PMID- 11252970 TI - [Organizational and operational work of death analysis committees: review of experiences]. AB - OBJECTIVES: Among deaths occurring in hospital, some are unexpected at the time of admission and unexplained by the course of the illness at the time of death. Health care givers agree that analysis of the circumstances of death is a powerful method for improving the quality of medical care but are nevertheless reluctant to set up mortality conferences. We present institution-wide experience with mortality conferences, detailing methods used and reviewing adverse care events and avoidable organizational and medical errors associated with unexpected deaths. METHODS: We reviewed literature cited in the Medline and Pascal Biomed databases and completed our findings with consultations with key-informants of hospital evaluation units. Institution-wide experiences alone were analyzed. RESULTS: The death analysis committees were composed of health care givers who review the deaths occurring in their units (internal committees). In rare cases, hospitals set up in addition an external committee for methodological assistance. Implicit criteria were almost always used for death case analysis. DISCUSSION: Death analysis by internal committees appears to contribute to physician's knowledge and provokes a global improvement in patient care. This analysis relies however on implicit criteria which leads to minimal reliability. The lack of reliability has two consequences: the impossibility of identifying causes of preventable deaths and related factors, and the impossibility of evaluating the impact of the internal committee. As there is no available epidemiological data, a sentinel system cannot be proposed: a systematic analysis of all deaths is therefore advisable. Other limitations on internal committees are the difficulty of obtaining autopsies and the problem of the confidentiality of death analysis, a serious handicap recognized by all physicians. A national guide on methodology should be developed. PMID- 11252971 TI - [The body packer syndrome]. AB - BACKGROUND: Ingestion of illicit drug packages is a well known method for transportation. These packages are prone to rupture causing overdose. The body packer syndrome may be overlooked in medical practice as illustrated by the following case report. CASE REPORT: A 19-year old male had convulsions followed by cardiac arrest during a flight. He was resuscitated in the plane, but he died a few hours after admission in intensive care unit. Chest and abdominal X-rays were considered normal. Cocaine metabolites were found in his urine. The death was considered suspicious. X-rays performed before medicolegal autopsy showed numerous packages in his digestive tract. Thirty-six packages were found in the stomach and intestine. Two were ruptured in the stomach. The cause of death was cocaine overdose caused by package rupture. DISCUSSION: The packages are usually visible on an standard abdomen X-ray. The drug is often wrapped in latex membranes or condoms. The air is trapped between the condoms by the nodes, forming two crescents visible on the X-ray. Surgery is preferred to laxatives when the packages are fragile with a high risk of rupture. PMID- 11252972 TI - [Concomitant active pulmonary tuberculosis and respiratory aspergillosis]. PMID- 11252973 TI - [Terminal care: 8 rules to follow]. PMID- 11252974 TI - [Acute pentobarbital poisoning]. PMID- 11252975 TI - [Elevated CA19.9 without a malignant tumor]. PMID- 11252976 TI - [Is it necessary to organize death analysis committees in our hospitals?]. PMID- 11252977 TI - [Regional hospitalization agencies and accreditation]. PMID- 11252978 TI - [Renal tolerance to amphotericin B]. PMID- 11252979 TI - [Drugs and drug abusers]. AB - DRUG ABUSERS: Drugs are widely used by toxicomaniacs to reproduce drug effects. Drug abusers generally start with psychotrops, but other abuse drug classes. Toxicomanic behavior leads to addictive practices that are difficult to control. BARBITURATES: Both the oral and intravenous routes are used. The expected result is a state of ecstasy with a feeling of comfort. Intoxication may cause respiratory depression. Barbiturates induce physical and psychic dependence. Abuse is not widespread with this class of drugs. BENZODIAZEPINES: Drug abuses widely use benzodiazepines orally or intravenously. They search for a flash effect, with sedation and a feeling of comfort. All benzodiazepines induce physical and psychic dependence. Death may result from combinations leading to respiratory depression. Flunitrazepam is the most widely abused benzodiazepine in France. It induces serious neuropsychic disorders. ANTIDEPRESSANTS: Few are used, mostly at high doses. OPIATES: Administration gives the same effect as heroine injection. Opiates induce physical and psychic dependence. The adverse effects are similar to those of morphine with a higher risk of respiratory depression. AMPHETAMINES: Few are used, either orally or intravenously. They induce a flash with excitation, euphoria, and a period of invincibility. This is followed by a period of depression with risk of suicide. Psychic dependence is high. ANTICHOLINERGIC ANTIPARKINSONIANS: These drugs are well known to abusers for their hallucinatory effect. They induce atropinic adverse effects and physical and psychic dependence. GAMMA-HYDROXYBUTYRATE: This anesthetic is used for its euphoria and sedation effects. It may induce falling sickness or coma, with a risk of respiratory depression. KETAMINE: Administered via the intranasal route, ketamine induces a state of indifference. Death has been observed. ANABOLIC AND ANDROGENIC STEROIDS: These drugs are used for their physical and psychic stimulating effect. They induce potentially dangerous adverse effects such as cardiovascular, hepatic, neurological and psychiatric disorders. Clinical signs of addiction and weaning are observed. OTHERS: Several other drug classes are used by abusers, including analgics, beta-adrenergic agents, nasal vasoconstrictors and corticosteroids. PMID- 11252980 TI - [Type 2 diabetes: what therapeutic strategy?]. AB - GOAL OF TREATMENT: Prevention of diabetic micro and macroangiopathy is the goal of treatment in type 2 diabetes mellitus. A well-controlled glucose level is the key to prevention of microangiopathy; there is no threshold level. Antihypertensive treatment, with the goal of blood pressure below 130/80 mmHg is also beneficial in preventing aggravation of microangiopathy. For macroangiopathy, prevention is based in priority on treatment of other risk factors for cardiovascular disease; the threshold level for drug treatment and the therapeutic objective are those defined for secondary prevention in non diabetic patients, i.e. blood pressure below 140/80 mmHg and LDL cholesterol under 1.30 g/l. The beneficial effect of lower glucose levels on preventing macrovascular risk was not formally demonstrated by the UKPDS, probably because the difference between the control and the treatment group HbA1c levels was minimal, 0.9 points. REVISITING STRATEGY: It is thus time to revisit the preventive strategy for type 2 diabetes mellitus, i.e. step-by-step increments, as currently proposed for worsening glucose levels. Metformine should be prescribed if the HbA1c is above normal in order to achieve the demonstrated benefit in prevention of microangiopathy and in the hope, motivated by pathophysiology data, of preventing insulin failure. Slow-release insulin at bedtime should be added to the oral hypoglycemiants if fasting glucose exceeds 1.60 or 1.80 g/l, even if the HbA1c remains below 8%. NEW HYPOGLYCEMIANTS: The role of these new agents in this more "aggressive" strategy remains to be defined. Glinides will have to demonstrate their superiority over sulfamides (fewer episodes of hypoglycemia with comparable efficacy) to justify their high cost. Glitazones will have to demonstrate a beneficial effect in second intention combination with metformine on cardiovascular morbidity mortality in type 2 diabetes patients with a metabolic insulin-resistance syndrome and visceral obesity. OBSERVANCE: Since patients with type 2 diabetes mellitus are often taking 3 to 6 tablets to control their glucose level, 3 to control blood pressure, plus another to lower the lipid level and finally one more for an antiplatelet effect reducing the number of tablets and patient education will most certainly help improve therapeutic observance. PMID- 11252981 TI - [Venous thrombosis from central venous catheterization in oncology]. AB - OBJECTIVES: The aim of this review article is to assess the epidemiology, diagnosis and treatment of deep venous thrombosis of the upper limbs which are becoming more common with the use of central catheterizations in oncology. RISK FACTORS: The incidence is differently estimated. The catheterization is the main risk factor which is added to the proper patient's risk factors in a highly thrombogenic context (cancer, chemotherapy, infection,...). The outcome can be limited to the catheter (thrombotic) dysfunction or can progress towards vein thrombosis, pulmonary embolism, most often asymptomatic or to a superior vena cava syndrome. Ultrasounds and CT scan have an important role although their efficacy have not been demonstrated yet. TREATMENT: Prophylactic anticoagulation with unfractionated heparin, low molecular weight heparin or oral anticoagulant seems to be effective. Low dose thrombolysis by bolus of urokinase or tissue plasminogen activator allows restoring the patency of the catheter in 70 to 90% cases of thrombotic dysfunction. The modalities of treatment of the vein thrombosis are much discussed. PMID- 11252982 TI - Mobilisation of Latin America to promote health. PMID- 11252983 TI - Health promotion in Chile: an evaluation of a national plan implementation. PMID- 11252984 TI - Mexico: health promotion initiatives. AB - Mexico is currently undergoing a Health Sector Reform to address the country's epidemiologic and demographic changes, deep socio-economic inequalities and their consequences on health. The Government and a diversified set of actors, mainly NGOs, are taking up health promotion initiatives. PMID- 11252985 TI - Impact of health promotion and education on the achievements of Cuban public health. PMID- 11252986 TI - Local integrated and sustainable development as a strategy for "radical health promotion" in Brazil. PMID- 11252987 TI - Public health policies and advocacy in Latin America: chances and environments to support these initiatives. PMID- 11252988 TI - Health promotion in the Americas: toward bridging the equity gap. PMID- 11252989 TI - Health promotion, health education and social communication on health: specificities, interfaces, intersections. PMID- 11252990 TI - Vertebrate neural cell-fate determination: lessons from the retina. AB - Postmitotic neurons are produced from a pool of cycling progenitors in an orderly fashion during development. Studies of cell-fate determination in the vertebrate retina have uncovered several fundamental principles by which this is achieved. Most notably, a model for vertebrate cell-fate determination has been proposed that combines findings on the relative roles of extrinsic and intrinsic regulators in controlling cell-fate choices. At the heart of the model is the proposal that progenitors pass through intrinsically determined competence states, during which they are capable of giving rise to a limited subset of cell types under the influence of extrinsic signals. PMID- 11252992 TI - The neurobiology of attachment. AB - It is difficult to think of any behavioural process that is more intrinsically important to us than attachment. Feeding, sleeping and locomotion are all necessary for survival, but humans are, as Baruch Spinoza famously noted, "a social animal" and it is our social attachments that we live for. Over the past decade, studies in a range of vertebrates, including humans, have begun to address the neural basis of attachment at a molecular, cellular and systems level. This review describes some of the important insights from this work. PMID- 11252991 TI - Molecular basis of long-term plasticity underlying addiction. AB - Studies of human addicts and behavioural studies in rodent models of addiction indicate that key behavioural abnormalities associated with addiction are extremely long lived. So, chronic drug exposure causes stable changes in the brain at the molecular and cellular levels that underlie these behavioural abnormalities. There has been considerable progress in identifying the mechanisms that contribute to long-lived neural and behavioural plasticity related to addiction, including drug-induced changes in gene transcription, in RNA and protein processing, and in synaptic structure. Although the specific changes identified so far are not sufficiently long lasting to account for the nearly permanent changes in behaviour associated with addiction, recent work has pointed to the types of mechanism that could be involved. PMID- 11252993 TI - The genetics of g in human and mouse. AB - The g factor refers to the substantial overlap that exists between individual differences in diverse cognitive processes in humans. In this article, I argue that a mouse model of g could provide a powerful analytic tool for exploring cognitive processes that are linked functionally by genes. PMID- 11252994 TI - The hands that hold the keys. PMID- 11252995 TI - Incentive memory. PMID- 11252996 TI - An X-orphan. PMID- 11252997 TI - Disappearing act with two barrels. PMID- 11252998 TI - The molecular dynamics of pain control. AB - Pain is necessary for survival, but persistent pain can result in anxiety, depression and a reduction in the quality of life. The discriminative and affective qualities of pain are both thought to be regulated in an activity dependent fashion. Recent studies have identified cells and molecules that regulate pain sensitivity and the parallel pathways that distribute nociceptive information to limbic or sensory areas of the forebrain. Here, we emphasize the cellular and neurobiological consequences of pain, especially those that are involved in the generation and maintenance of chronic pain. These new insights into pain processing will significantly alter our approach to pain control and the development of new analgesics. PMID- 11252999 TI - Initial patterning of the central nervous system: how many organizers? AB - For three-quarters of a century, developmental biologists have been asking how the nervous system is specified as distinct from the rest of the ectoderm during early development, and how it becomes subdivided initially into distinct regions such as forebrain, midbrain, hindbrain and spinal cord. The two events of 'neural induction' and 'early neural patterning' seem to be intertwined, and many models have been put forward to explain how these processes work at a molecular level. Here I consider early neural patterning and discuss the evidence for and against the two most popular models proposed for its explanation: the idea that multiple signalling centres (organizers) are responsible for inducing different regions of the nervous system, and a model first articulated by Nieuwkoop that invokes two steps (activation/transformation) necessary for neural patterning. As recent evidence from several systems challenges both models, I propose a modification of Nieuwkoop's model that most easily accommodates both classical and more recent data, and end by outlining some possible directions for future research. PMID- 11253000 TI - Neural plate patterning: upstream and downstream of the isthmic organizer. AB - Two organizing centres operate at long-range distances within the anterior neural plate to pattern the forebrain, midbrain and hindbrain. Important progress has been made in understanding the formation and function of one of these organizing centres, the isthmic organizer, which controls the development of the midbrain and anterior hindbrain. Here we review our current knowledge on the identity, localization and maintenance of the isthmic organizer, as well as on the molecular cascades that underlie the activity of this organizing centre. PMID- 11253001 TI - Monitoring the species of arsenic, chromium and nickel in milled coal, bottom ash and fly ash from a pulverized coal-fired power plant in western Canada. AB - The concentration of As, Cr and Ni and their speciation (As3+;5+, Cr3+;6+ and Ni0;2+) in milled coal, bottom ash and ash collected by electrostatic precipitator (ESP) from a coal fired-power plant in western Canada were determined using HGAAS, ICP-AES and XANES. The chemical fractionation of these elements was also determined by a sequential leaching procedure, using deionized water, NH4OAC and HCI as extracting agents. The leachate was analyzed by ICP-AES. Arsenic in the milled coal is mostly associated with organic matter, and 67% of this arsenic is removed by ammonium acetate. This element is totally removed from milled coal after extraction with HCI. Arsenic occurs in both the As3+ and the As5+ oxidation states in the milled coal, while virtually all (>90%) of the arsenic in bottom ash and fly ash appears to be in the less toxic arsenate (As5+) form. Both Ni and Cr in the milled coal are extracted by HCI, indicating that water can mobilize Ni and Cr in an acidic environment. The chromium is leached by water from fly ash as a result of the high pH of the water, which is induced during the leaching. Ammonium acetate removes Ni from bottom ash through an ion exchange process. Chromium in milled coal is present entirely as Cr3+, which is an essential human trace nutrient. The Cr speciation in bottom ash is a more accentuated version of the milled coal and consists mostly of the Cr3+ species. Chromium in fly ash is mostly Cr3+, with significant contamination by stainless steel from the installation itself. PMID- 11253002 TI - Total organic carbon analyzers as tools for measuring carbonaceous matter in natural waters. AB - For some utilities, new US drinking water regulations may require the removal of disinfection byproduct (DBP) precursor material as a means of minimizing DBP formation. The Environmental Protection Agency's Stage 1 DBP Rule relies on total organic carbon (TOC) concentrations as a measure of the effectiveness of treatment techniques for removing organic material that could act as DBP precursors. Accordingly, precise and accurate methods are needed for the determination of TOC and dissolved organic carbon (DOC) concentrations in raw and finished potable water supplies. This review describes the current analytical technologies and summarizes the key factors affecting measurement quality. It provides a look into the fundamental principles and workings of TOC analyzers. Current peroxydisulfuric acid wet ashing methods and combustion methods are discussed. Issues affecting quality control, such as non-zero blanks and preservation, are covered. Some of the difficulties in analyzing water for TOC and DOC that were identified up to 20 years ago still remain problematic today. Limitations in technology, reagent purity, operator skill and knowledge of natural organic matter (NOM) can preclude the level of precision and accuracy desirable for compliance monitoring. PMID- 11253003 TI - The application of ICP-SMS, GF-AAS and HG-AFS to the analysis of water and sediment samples from a temperate stratified estuary. AB - Three atomic spectrometry techniques, namely sector field inductively coupled plasma mass spectroscopy, graphite furnace atomic absorption spectrometry and hydride generation atomic fluorescence spectroscopy (ICP-SMS, GF-AAS and HG-AFS, respectively), housed at separate independent laboratories, were used to analyse water and sediment samples collected from the Huon River Estuary, SE Tasmania (Australia) in the Austral spring 1998. A dithiocarbamate-chelation/back extraction technique was used to separate and preconcentrate Co, Ni, Cu, Zn, Cd and Pb from eight collected water samples prior to analysis by ICP-SMS and GF AAS. A number of other elements in the waters were analysed directly (Mn, Fe and Zn by GF-AAS; As by HG-AFS), or following sample dilution (1 + 19: V, Mn, Fe, As, Mo, Ba and U by ICP-SMS). Where possible, previously corroborated GF-AAS and HG AFS techniques were used to verify obtained ICP-SMS results. From the analysis of four reference waters (SLEW-1 and -2, SLRS-3 and NASS-5), good agreement, to within +/- 10-20%, was typically found between certified (or information only values) and measured results (irrespective of analytical technique). Exceptions included Zn (and sometimes Fe) that could not be quantified by ICP-SMS due to elevated blank signals, and As which was found to lie below ICP-SMS detection limits. For Huon Estuary water samples, inter-method agreement was within +/- 10 20% (for those elements amenable to analysis by more than one technique). Nitric acid extracts of two certified reference materials (Buffalo River Sediment and BCSS-1) and six Huon Estuary sediments were analysed by ICP-SMS (for Al, Sc, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, Cd and Pb) and HG-AFS (for As). Results from the certified reference materials indicated extraction efficiencies of 60 70% (for most elements). A close correlation between ICP-SMS and HG-AFS was obtained for leachable As in the sediments. In terms of potential inorganic contaminants, the Huon Estuary was found to be a relatively 'clean' water system. The elemental concentrations measured in water and sediment samples from this region were found to lie within current Australian guidelines for estuaries. In general, no one analytical technique was able to accurately determine all elements in all samples from this relatively pristine estuarine environment. A combination of all three analytical techniques was necessary for the successful analysis of the elements considered in this study. PMID- 11253004 TI - Development of a harmonised phosphorus extraction procedure and certification of a sediment reference material. AB - A harmonised procedure for the determination of the forms of phosphorus in freshwater sediments, developed in the frame of the European Programme, Standards, Measurements and Testing (SMT) has been used for a certification campaign for a reference material. This operationally defined scheme is a good compromise between method performance and reproducibility. Furthermore, the method is rather simple to implement and could be used by water managers on a routine basis. A homogeneous and stable sediment reference material has been prepared and will be available before mid 2001. The so-called SMT protocol, together with the reference material, are useful tools in the field of water management, especially at a time when quality assurance is of paramount importance in laboratory analyses. Knowledge of the bioavailable forms of phosphorus is important not only for analysis of sediments but also for sludge and soils. Therefore, the SMT protocol could be extended to these materials. PMID- 11253005 TI - Fate and movement of atrazine, cyanazine, metolachlor and selected degradation products in water resources of the deep Loess Hills of Southwestern Iowa, USA. AB - The environmental fate and movement of herbicides widely used for weed control in corn are assessed for a deep loess soil in southwestern Iowa. Beginning in the early 1980s, the herbicide-based weed control program emphasized the application of atrazine (ATR) or cyanazine (CYN) and metolachlor (MET) for both broadleaf and grass control. Between 1992 and 1995, concentrations of ATR, desethylatrazine (DEA), desisopropylatrazine (DIA), CYN and MET were measured in rainwater, both shallow and deep vadose zone water, and well water. Results show that the frequency of herbicide detections and the range and distribution of occurrences are dependent upon both landscape position and temporal inputs of recharge water from rainfall. Generally, DIA was observed more frequently and in higher mean concentration in well water than DEA, while DEA was observed more frequently than DIA in vadose zone groundwater. A chromatographic analogy is suggested to explain the occurrence patterns observed for both parent herbicide and degradation products within the unsaturated zone water. Analysis of rainwater samples collected during this time also revealed low concentrations of ATR, CYN and MET, with the timing of the detections indicative of non-local transport. Results show that the deep loess soil conducts both water and agricultural chemicals relatively rapidly and as such represents a production system which is vulnerable to contamination of shallow groundwater by herbicide-derived chemicals. Results also illustrate the importance of including major herbicide degradation products in water resource impact assessment studies. PMID- 11253006 TI - Characterisation of mineralogical forms of barium and trace heavy metal impurities in commercial barytes by EPMA, XRD and ICP-MS. AB - This study was carried out to characterise the mineralogical forms of barium and the trace heavy metal impurities in commercial barytes of different origins using electron probe microanalysis (EPMA), X-ray diffraction (XRD) and inductively coupled plasma mass spectrometry (ICP-MS). Qualitative EPMA results show the presence of typically eight different minerals in commercial barytes including barite (BaSO4), barium feldspar, galena (PbS), pyrite (FeS2), sphalerite (ZnS), quartz (SiO2), and silicates, etc. Quantitative EPMA confirms that the barite crystals in the barytes contain some strontium and a little calcium, whereas trace heavy metals occur in the associated minerals. Analysis of aqua regia extracts of barytes samples by ICP-MS has shown the presence of a large number of elements in the associated minerals. Arsenic, copper and zinc concentrations correlate closely in all 10 samples. The findings suggest that barytes is not, as traditionally thought, an inert mineral, but is a potentially toxic substance due to its associated heavy metal impurities, which can be determined by an aqua regia digest without the need for complete dissolution of the barite itself. X ray powder diffraction was not informative as the complex barite pattern masks the very weak lines from the small amounts of associated minerals. PMID- 11253007 TI - Measurements of nitrogen dioxide in Greenland using Palmes diffusion tubes. AB - Measurements of nitrogen dioxide using the Palmes diffusion tubes in Uummannaq, Aasiaat, and Nuuk. all located along the west-coast of Greenland, have demonstrated that the levels of pollution at the most heavily impacted sites are comparable to levels in much larger towns in Denmark. The highest concentrations were, in general, observed near sites influenced by car traffic (peak concentrations of up to 16 ppbv), medium concentrations were observed in the residential areas (2 6 ppbv), and very low levels were found at the background locations in the town outskirts (1-2 ppbv). Observations of nitrogen dioxide concentrations less than 0.1 ppbv at a remote site, Akia, 25 km from Nuuk, indicate that, compared to local sources, long-range transport of nitrogen dioxide is not important in western Greenland. PMID- 11253008 TI - Levels of lead in atmospheric deposition in a large urban agglomeration in Poland. AB - Lead levels in wet and dry deposition were determined within this project. A network of 10 sampling stations was established. The stations were located in areas characterized by heavy traffic volumes, but away from industrial and/or municipal pollution sources. It was assumed, therefore, that lead in the samples collected was coming primarily from automobile emissions. Measurements were carried out over a period of one year. Both rain and snow samples were collected. Lead concentrations in the samples ranged from 0.6 to 141 microg dm(-3). They depended on street topography, traffic volume, average speed of the vehicles, frequency of traffic congestion and atmospheric conditions. The highest lead levels in deposition were observed during the cold season. PMID- 11253009 TI - Assessing dermal exposure to pesticides from non-agricultural uses: a UK health and safety executive (HSE) perspective. PMID- 11253010 TI - Trace element speciation at cell membranes: aqueous, solid and lipid phase effects. AB - Biological interest in trace element speciation has tended to be polarised in very different ways according to the concerns of the investigators. Much of the ecological interest has centred on the effects of metal ions in the external environment with the "free metal ion model" dominating discussions of potential toxicity. By way of contrast biochemists have been much more concerned with the stereochemical and kinetic aspects of specificity in metal protein interactions in the cytoplasm. Separating these two sets of interests are the membrane biophysicists whose studies have concentrated on the channel concept. All three groups have tended to ignore speciation onto the solid phase. In the overall biological context, trace element speciation in the cell is more concerned with kinetics and the evolution of specificity of interaction between diverse ligands than with the conditions for equilibrium. PMID- 11253011 TI - Influence of environmental parameters on the accuracy of nitrogen dioxide passive diffusion tubes for ambient measurement. AB - Two studies at three sites in the UK provided confirmation that systematic positive bias in NO2 diffusion tube measurement occurred because of changes to "within-tube" chemistry, rather than eddy diffusion at the mouth of the tube. In the first study in Cambridge, UK, sampler overestimation for 1 and 2 week exposures was compared to corresponding time-averaged monitor measurements (NO NO2-NOx, O3) and weather variables. Noninearity between sampler and monitor NO2 measurements was interpreted in terms of spatial and temporal variations in relative and absolute availability of NO, NO2 and O3 at the site. A maximum overestimation occurred for an exposure mean NO2/NOx approximately 0.5. The separate contributions of reduced NO2 photolysis and eddy diffusion were compared in Study II using samplers of two materials, acrylic and quartz, and of different lengths (40, 55, 71 and 120 mm) at three sites: Norwich background, Cambridge intermediate, London kerbside. For compared sites, NO2 measured by acrylic samplers was significantly higher than for equivalent quartz samplers. For quartz samplers [NO2]mean was only just above the monitor at Norwich and London; sampler/monitor NO2 = 1.04 (P = 0.59) and 1.01(P = 0.76), respectively. For acrylic samplers the order of [NO2]mean was 40 mm > 120 mm > 71 mm > or = 55 mm. Excepting 40 mm samplers, this accords with a chemical bias where co-diffusing NO and 03 molecules in longer tubes have more time to react to form excess NO2. Bias in 40 mm samplers is discussed. Eddy diffusion is negligible for standard samplers because [NO2]mean was equivalent for 55 mm and 71 mm acrylic samplers and close to monitor NO2 for 71 mm quartz tubes. Both studies showed that sampler accuracy was dependent on location. Significantly, overestimation was greatest (approximately 3-4 ppb) where the NO2 annual mean was approximately 20 ppb, close to the UK and EU air quality standard of 21 ppb. PMID- 11253012 TI - Evaluation of VOC measurments in the EXPOLIS study. Air Pollution Exposure Distributions within Adult Urban Urban Populations in Europe. AB - Personal exposures and microenvironment concentrations of 30 target VOCs were measured for 401 participants living in five European cities as a part of the EXPOLIS (Air Pollution Exposure Distributions within Adult Urban Populations in Europe) study. Measurements in Basel used an active charcoal (Carbotech) adsorbent as opposed to the Tenax TA used in the other study centres. In addition, within each centre, personal and microenvironment VOC sampling required different sampling pumps and, because of different sampling durations, different sampling flow rates. Thus, careful testing of the sampling and analysis procedures was required to ensure accuracy and comparability of collected data. Monitor comparison tests using Tenax TA showed a mean VOC concentration ratio of 0.95 between the personal and microenvironment monitors. The LODs for the target VOCs using Tenax TA ranged from 0.7 to 5.2 microg m(-3). The LODs for the 14 target compounds quantifiable using Carbotech ranged from 0.9 to 3.2 microg m( 3). Tenax TA field blanks showed no remarkable contamination with the target VOCs, except benzaldehyde, a known artefact with this adsorbent. Thus, the diffusion barrier system used prevented contamination of Tenax TA samples by passive diffusion during non-sampling periods. Duplicate and parallel evaluations of the Tenax TA and Carbotech showed an average difference of < 17% in VOC concentrations within the sampling methods, but a systematic difference between the methods (Tenax TA: Carbotech concentration ratio = 1.18-2.36). These field evaluations and quality assurance tests showed that interpretation and comparison of the results in any VOC monitoring exercise should be done on a compound by compound basis. It is also apparent that carefully planned and realised QA and QC (QA/QC) procedures are needed in multi-centre studies, where a common sampling method and laboratory analysis technique are not used, to strengthen and simplify the interpretation of observed VOC levels between participating centres. PMID- 11253013 TI - A comparison of sampling and analysis methods for low-ppbC levels of volatile organic compounds in ambient air. AB - A carefully designed study was conducted during the summer of 1998 to collect samples of ambient air by canisters and compare the analysis results to direct sorbent preconcentration results taken at the time of sample collection. Thirty two 1 h sample sets were taken, each composed of a "near-real-time" sample analyzed by an autoGC-MS XonTech 930/Varian Saturn 2000 system, and Summa and Silco canisters. Hourly total non-methane organic carbon (TNMOC), ozone, and meteorological measurements were also made. Each canister was analyzed on the autoGC-MS system for a target list of 108 volatile organic compounds (VOCs) and on a manual cryosampling GC-FID system. Comparisons were made between the collection and analysis methods. Because of the low sample loading (150-250 ppbC TNMOC), these comparisons were a stringent test of sample collection and analysis capabilities. The following specific conclusions may be drawn from this study. Reasonable precision (within 15% mean difference of duplicate analyses from the same canister) can be obtained for analyses of target VOCs at low-ppbC concentrations. Relative accuracy between the GC-MS and GC-FID analysis methods is excellent, as demonstrated by comparisons of analyses of the same canisters, if measurements are sufficiently above the detection limits. This is especially significant as the GC-MS and GC-FID were independently calibrated. While statistically significant differences may be observed between the results from canister and near-real-time samples, the differences were generally small and there were clear correlations between the canister results and the near-real-time results. Canister cleanliness limits detection below the EPA Method TO-14 acceptance standard of 0.2 ppbv (0.2-2 ppbC for target analytes). PMID- 11253014 TI - Application of natural zeolites for preconcentration of arsenic species in water samples. AB - Zeolites of the clinoptilolite type produced in Mexico and Hungary were investigated with respect to their sorption efficiency for various redox species of arsenic. Long-term experiments showed that arsenate remains stable for a long period in spiked deionised water and drinking water, as well as in the surface water of the Biela valley in Saxony, Germany. Both clinoptilolites are able to decrease the initial arsenic concentration of 200 microg l(-1) by more than 75% in deionised, drinking, ground and surface waters. In the case of the Mexican zeolite, both the arsenite and the arsenate concentrations (200 microg l(-1)) can be lowered from 200 microg l(-1) to 10 microg l(-1), which is the World Health Organisation's (WHO's) recommended maximum level. It was found that the presence of cations and anions in the natural waters of Biela, Germany, and Zimapan, Mexico, did not reduce the efficiency of the selected zeolites. The Hungarian zeolite removed 75% of the arsenate in the Zimapan water and only 50% when the sample was first acidified. This zeolite totally desorbed the fixed arsenic into a water volume that was half the volume in the adsorption experiment. PMID- 11253015 TI - Historical variation of elements with respect to different geochemical fractions in recent sediments from Pigeon Lake, Alberta, Canada. AB - Geochemical analysis of elements and organic matter were conducted on vertical profiles of the recent sediments from Pigeon Lake, Alberta, Canada, to determine historical variations in elemental content of the sediments as related to their geochemical fractions. The elements are grouped according to their affinity with different geochemical fractions, by using cluster analysis and sequential extraction experiments. As a result, four elemental fractions were identified: clastic mineral detritus; carbonate; organic; and elements that show less similarity to the previous groups perhaps due to anthropogenic input or the influence of other fractions, such as oxyhydroxides. Following the identification of geochemical fractions in the sediments, a three-step normalizing method was applied using parameters that represent each geochemical fraction. These normalizing techniques appear to be important in verifying whether the variation of elements is indeed the result of anthropogenic and/or natural activities. The normalized data are correlated with the recent history of human activity and natural events near Pigeon Lake. Given the age of the lake sediments, this correlation indicates that the depth profiles of elements after being normalized to the organic and carbonate fractions reflect the variation of detrital input into the lake. However, the former mainly corresponds to the coarse-grained clastic minerals originating from high-energy erosion as the result of natural events (e.g., flooding), whereas the latter corresponds to the low-energy erosion of the fine particles (enriched in lithophile elements) due to deforestation in the drainage basin. Normalizing to the clastic mineral detritus fraction results in the increase of heavy metals in the uppermost part of the sediment profiles, which coincides with industrial activities during the past two decades in central Alberta. However, the concentration of these elements is negligible, as compared to the quantities released by geogenic processes (erosion). PMID- 11253016 TI - The science of the unclean. PMID- 11253017 TI - Partitioning of mercury onto suspended sediments in estuaries. AB - Radiochemical partitioning experiments using 203Hg have been undertaken with mixtures of river, seawater and sediment samples taken from three geochemically contrasting UK estuaries: the Plym, Beaulieu and Mersey. Species of dissolved Hg were determined using reversed-phase C18 chelating columns and particulate species were determined by sequential leaching with 1 M NH4OAc and 1 M HCl. Mercury had a high particle reactivity with partition coefficients, KDs, ranging from 10(4) to 5 x 10(5) ml g(-1), depending on salinity, the chemical composition of the end-member waters, and on the physico-chemical characteristics of the sediment. Dissolved organic matter present in the waters (humic substances and/or anthropogenic compounds) was found to be the main factor governing the forms of dissolved Hg and their reactivity. From the spiked 203Hg, up to 95% of the dissolved metal was retained on the C18 columns for the Mersey waters, whereas this fraction was < 60% in the Plym and Beaulieu waters. Quasi-irreversible adsorption of Hg onto particles from each estuary was observed over a time-scale of a few hours and < 20% of total particulate Hg was released by the sequential leach. In this paper, physico-chemical processes are proposed to explain the estuarine behaviour of Hg and the results are discussed in terms of Hg availability in estuarine systems. PMID- 11253018 TI - Monitoring cyclical air/water elemental mercury exchange. AB - Previous experimental work has demonstrated that elemental mercury evasion from natural water displays a diel cycle; evasion rates during the day can be two to three times evasion rates observed at night. A study with polychlorinated biphenyls (PCBs) found that diurnal PCB air/water exchange rates exceeded nocturnal exchange rates by 32%. Given that the exchange rates of both PCBs and elemental mercury are dominated by the resistance in the aqueous thin film at the air/water interface and that water column elemental mercury concentrations in natural water bodies also display a diel cycle (and water column PCB concentrations do not) the findings here suggest that PCBs can serve as a tracer to assess the relative contribution of diel atmospheric temperature variations on elemental mercury air/water exchange rates. Using previously published data describing water column elemental mercury concentrations and the previously published diel mercury evasion model, four evasion scenarios are examined within the context of monitoring air/water toxicant exchange: constant atmospheric temperatures and constant water column elemental mercury concentrations; variable atmospheric temperatures and constant water column elemental mercury concentrations; constant atmospheric temperatures and variable water column elemental mercury concentrations; and variable atmospheric temperatures and variable water column elemental mercury concentrations. A scenario of monthly elemental mercury air/water exchange also is examined (at constant atmospheric and water column elemental mercury concentrations). Some of the findings include: (1) atmospheric temperature variations do have a significant effect on air/water toxicant exchange; (2) diel atmospheric temperature variations become more significant to overall diel toxicant exchange rates the closer the air/water system is to equilibrium conditions; (3) for refractory toxicants, average diel exchange rates are best estimated by averaging datasets obtained over a 24 h period or, at minimum, by measuring exchange rates at average atmospheric temperature values; (4) for elemental mercury, variable diel water column concentrations are likely to be the dominant contributor to variations in diel evasion rates; (5) diel atmospheric temperature variations amplify the magnitudes of both diel mercury evasion and absorption events and can shift maximum evasion rates to later in the day; (6) variations in monthly elemental mercury air/water exchange rates may exceed diel variations: and (7) 24 h and monthly monitoring efforts will likely be required to accurately describe diel and annual elemental mercury air/water exchange in a given system. PMID- 11253019 TI - The relevance of speciation in the remediation of soils and sediments contaminated by metallic elements--an overview and examples from Central Scotland, UK. AB - The environmental impact of metallic contaminants in soils and sediments is dependent both on the chemical speciation of the metal and the response of the matrix to biological and physicochemical conditions. These factors are responsible for the mobilisation of the metal from the solid into the aquatic phase and hence transport within the immediate vicinity, impacting on the rate of dispersal, dilution, uptake and transfer into living systems. The impact of changing environmental conditions on the contaminant inventory can be to enhance or moderate these phenomena, with subsequent consequences for the broader risk assessment of the contaminants. Remediation of metallic contaminants can only be brought about by their removal from the site or by establishing conditions which favour their retention in the solid phase. A wide range of in situ and ex situ approaches are available and a summary overview is presented. The examples show assessment at both the field and laboratory scale and demonstrate an equally wide range of success in achieving remediation targets. This can be attributed to limitations in ensuring that the desired conditions for the initial removal or immobilisation process are met and maintained over a suitable period of time. Three areas are reviewed which include: the transport and release of metallic contaminants in estuarine sediments and the assessment of their potential to impact on biota; terrestrial contamination systems involving the release of chromium from waste ore contaminating urban environments; the response of metal contaminated wastes to changing environmental conditions and the impact of natural bioremediation. The focus of the discussion is to highlight the generation of reliable speciation information and the problems associated with impact and risk assessment. Particular issues of concern are the laboratory to field scale evaluation of contaminant behaviour and the approach used to assess the reliability of remediation options. In conclusion, part of a recent initiative in risk assessment and the development of pilot scale experimental systems to study long-term behaviour are addressed as future goals to fill gaps in current research. PMID- 11253020 TI - Sulfur speciation monitored in situ with solid state gold amalgam voltammetric microelectrodes: polysulfides as a special case in sediments, microbial mats and hydrothermal vent waters. AB - Sulfur speciation was determined in real time in salt marsh microbial mats, subtidal sediments and hydrothermal vent diffuse flow waters using solid state gold-amalgam voltammetric microelectrodes. Chemical species were measured in situ without any sample manipulation or processing. The partially oxidized sulfur species detected were polysulfides, thiosulfate, elemental sulfur and tetrathionate. Fe(III) oxidation of hydrogen sulfide does not occur within the mats where microbially mediated processes are responsible for oxidation of H2S. In sediments and diffuse flow vent waters, Fe(III) phases are the direct oxidant of H2S. Sulfur speciation determined in this work is due to in situ biogeochemical processes and is not due to artefacts of sample manipulation. The voltammetric data show that polysulfides are the first detectable intermediate during sulfide oxidation which is consistent with previous laboratory studies. PMID- 11253021 TI - Determination of germanium in urine and its usefulness for biomonitoring of inhalation exposure to inorganic germanium in the occupational setting. AB - The present study aimed to assess whether urinary germanium concentration can be used as a biomarker of inhalation exposure to airborne dust from metallic germanium (Ge) or GeO2 in the occupational setting. A novel hydride generation based method coupled with fow-injection graphite furnace atomic absorption spectrometry (HG/FI-GFAAS) was developed for the determination of urinary germanium. It was found that urinary germanium concentration could be reliably determined by a standard additions method after thorough digestion of the urine and careful pH adjustment of the digest. The limit of detection (LOD) in urine for the HG/FI-GFAAS method was 0.25 microg Ge L(-1). In Belgian control male subjects, the urinary germanium concentration was below this LOD. In 75 workers currently exposed to inorganic germanium compounds, respirable and inhalable concentrations of germanium in the aerosols were measured on Monday and Friday at the job sites using personal air samplers. Spot-urine samples were collected on the same days before and after the work shift. The germanium concentrations of respirable dust correlated very well with those of inhalable dust and represented 20% of the inhalable fraction. Workers exposed to metallic Ge dust were on average ten times less exposed to germanium than those whose exposure involved GeO2 (3.4 versus 33.8 microg Ge m(-3)). This difference was reflected in the urinary germanium concentrations (3.4 versus 23.4 microg Ge g(-1) creatinine). Regression analysis showed that the concentration of germanium in the inhalable fraction explained 42% of the post-shift urinary germanium concentration either on Monday or on Friday, whereas in a subgroup of 52 workers mainly exposed to metallic germanium dust 57% (r = 0.76) of the Monday post-shift urinary germanium was explained. Urinary elimination kinetics were studied in seven workers exposed to airborne dust of either metallic Ge or GeO2. The urinary elimination rate of germanium was characterised by half-times ranging from 8.2 to 18.1 h (on average 12 h 46 min). The present study did not allow discrimination between the germanium species to which the workers were exposed, but it showed fast urinary elimination kinetics for inhalation exposure to dust of metallic Ge and GeO2. It pointed out that urine samples taken at the end of the work shift can be used for biological monitoring of inorganic germanium exposure in the occupational setting. PMID- 11253022 TI - Determination of total and EDTA extractable metal distributions in the colloidal fraction of contaminated soils using SdFFF-ICP-HRMS. AB - Newly developed methods involving an on-line combination of sedimentation field flow fractionation-inductively coupled plasma-high resolution mass spectrometry (SdFFF-ICP-HRMS) have been used to study the distributions of extractable heavy metals in a soil which had been treated with sewage sludge contaminated with Cu or Pb. The relationship of these metals with other elements in the colloidal fraction was also investigated. The colloidal fraction from the soil was obtained by repeated gravitational sedimentation and extracted with 0.11 M acetic acid, 0.1 M hydroxylamine hydrochloride, 0.05 M ethylenediaminetetraacetic acid disodium salt (EDTA) or aqua regia to assess the potential availability of the metals Cu and Pb. Large proportions of the Cu and Pb were extracted by EDTA, approaching that removed by aqua regia, whereas < 10% of the aqua regia extractable metals were removed by acetic acid and hydroxylamine chloride. The distributions of the heavy metals, the major mineral forming element (Al) and the elements forming sesquioxides (Fe and Mn) within different size classes (0.05-1 microm) of the colloidal fraction were measured using SdFFF-ICP-HRMS before and after extraction with EDTA. This information provides an insight into the composition of the colloids and the distributions of metal contaminants. In the contaminated soil colloids, the concentration of Fe, Mn and Pb is greatest in the smaller particles (<0.2 microm). In contrast, the Cu concentration is constant over the size range studied. Iron oxide surface coatings probably play a significant role in Pb adsorption on soil particles, but may be less important for Cu. The combination of selective chemical extraction, SdFFF and ICP-HRMS provides a means of determining the distribution of potentially available heavy metals within the colloidal fraction of contaminated soils. PMID- 11253023 TI - Monitoring of the natural environment by chemical speciation of elements in aerosol and sediment samples. AB - The development of a monitoring network for chemical speciation of elements of aerosol and sediment samples collected at Lake Balaton has been carried out. Sequential leaching procedures for the determination of the distribution of elements in aerosols (3 steps) and sediments (4 steps) were used. These methods were recently successfully applied to describe environmentally mobile and stable fractions of toxic metals. In aerosol matrices the partition of elements was accomplished by particle size and chemical bonding. In sediments the distribution was performed by chemical bonding. The processes are called fractionation of elements. Particular attention was paid to distinguishing between environmentally mobile and environmentally immobile fractions because these represent the two extreme modes by which the metals are bound to solid matrices. The monitoring objectives were to assess pollution effects on man and his environment and to identify any possible cause and effect relationship between pollutant concentrations and health effects. The results of dry and wet deposition rates showed that most of the toxic metals were dissolved in an aqueous phase and the wet deposition played an important role. It has been found that, while the concentration of Cd and Pb in aerosols is low (0.7 and 29 ng m(-3), respectively), environmentally mobile fractions are considerable. Based upon the data it can be concluded that the effect of the anthropogenic sources on the quality of the lake is minor. This has been the first attempt to correlate speciation results between aerosols and sediments. PMID- 11253024 TI - Speciation of arsenic using solid phase extraction cartridges. AB - Various solid phase extraction (SPE) cartridges were investigated for speciation of arsenite [As(III)], arsenate [As(v)], monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA). Cartridges containing different types of sorbent materials were tested for arsenic retention and elution characteristics. Alumina cartridges were found to completely retain all the four target arsenic species, and are suitable for removal and preconcentration purposes. For speciation analysis, different arsenic species were separated on the basis of their selective retention on and elution from specific cartridges. DMA was retained on a resin-based strong cation exchange cartridge and eluted with 1.0 M HCl. MMA and As(v) were both retained on a silica-based strong anion exchange cartridge and sequentially eluted with 60 mM acetic acid (for MMA) and 1.0 M HCl [for As(v)]. As(III) was not retained on either cartridge and remained in solution. Arsenic species in solution and those eluted from the cartridges were subsequently quantified by using flow injection with hydride generation atomic fluorescence spectrometry (FI-HGAFS) and hydride generation atomic absorption spectrometry (FI HGAAS). A detection limit of 0.05 microg L(-1) arsenic in water sample was achieved using HGAFS. An application of the method was demonstrated at a drinking water treatment facility. As(III) and As(v) species were determined in water at various stages of treatment. The method is suitable for routine determination of trace levels of arsenic in drinking water to comply with more stringent environmental regulations. PMID- 11253025 TI - In vivo distribution and speciation of [114mIn]InCl3 in the Wistar rat. AB - Five Wistar rats were given an intraperitoneal injection of [114mIn]InCl3 during four consecutive days. One hour after the last injection the rats were sacrificed. The in vivo distribution of 114mIn was studied in the blood and in different organs. Differential centrifugation was used to study the distribution in liver, kidney and spleen homogenate. Rat serum, packed cell lysate, urine and the cytosol of liver, kidney and spleen homogenate were examined by size exclusion fast protein liquid chromatography. The results showed that serum accounts for 90% of the indium activity in whole blood. Indium is preferentially accumulated within the liver, spleen and kidney, the highest amount of 114mIn being localised in the cytosolic fraction followed by the mitochondria. Size exclusion experiments showed that, in rat serum, indium is exclusively bound to transferrin. These results differed from earlier in vitro incubation experiments of human serum with 114mIn. It was not possible, from the experiments described herein, to conclude unequivocally whether indium is bound to haemoglobin of packed cell lysate or to another high molecular mass compound. Indium is associated with the high molecular mass fraction in liver, kidney and spleen cytosol; only in kidney are small amounts of 114mIn found in the low molecular mass fraction. The in vivo inhibitory effect of indium on the delta aminolaevulinic acid dehydratase (ALAD) enzymatic activity in red blood cells and kidney tissue, well documented by other researchers, could not be attributed to direct binding of indium with this enzyme. PMID- 11253026 TI - Soil solution extraction techniques for microbial ecotoxicity testing: a comparative evaluation. AB - The suitability of two different techniques (centrifugation and Rhizon sampler) for obtaining the interstitial pore water of soil (soil solution), integral to the ecotoxicity assessment of metal contaminated soil, were investigated by combining chemical analyses and a luminescence-based microbial biosensor. Two different techniques, centrifugation and Rhizon sampler, were used to extract the soil solution from Insch (a loamy sand) and Boyndie (a sandy loam) soils, which had been amended with different concentrations of Zn and Cd. The concentrations of dissolved organic carbon (DOC), major anions (F- , CI-, NO3, SO4(2-)) and major cations (K+, Mg2+, Ca2+) in the soil solutions varied depending on the extraction technique used. Overall, the concentrations of Zn and Cd were significantly higher in the soil solution extracted using the centrifugation technique compared with that extracted using the Rhizon sampler technique. Furthermore, the differences observed between the two extraction techniques depended on the type of soil from which the solution was being extracted. The luminescence-based biosensor Escherichia coli HB101 pUCD607 was shown to respond to the free metal concentrations in the soil solutions and showed that different toxicities were associated with each soil, depending on the technique used to extract the soil solution. This study highlights the need to characterise the type of extraction technique used to obtain the soil solution for ecotoxicity testing in order that a representative ecotoxicity assessment can be carried out. PMID- 11253027 TI - Summary paper of the EC Network on trace element speciation for analysts, industry and regulators--what we have and what we need. PMID- 11253028 TI - Measurement of styrene-7,8-oxide and other oxidation products of styrene in air. AB - Styrene-7,8-oxide (SO) is generated at low concentrations from the oxidation of styrene during the processing of reinforced plastics. Since exposure to SO has important health implications, we developed air sampling and analytical methods to measure low levels of airborne SO in the presence of styrene and its other oxidation products, namely phenylacetaldehyde (PAA) and acetophenone (AP). Both active and passive air monitors were used. The active sampling method, which employed adsorption on Tenax, was suitable for measuring SO, PAA and AP but had limited capacity for styrene due to breakthrough. The passive monitor employed a carbon adsorbent and was suitable for measurement of styrene and SO but not PAA and AP due to poor recovery. After sampling, the analytes were extracted from the adsorbents with ethyl acetate and measured by gas chromatography with flame ionization detection or mass spectrometry. By maintaining the injection port at 70 degrees C, the thermal rearrangement of SO to PAA was minimized. Recovery of styrene and SO from the passive monitor depended upon loading and was corrected by linearization of the Freundlich isotherm. The limits of detection for SO, PAA, and AP were 0.2 ppb using the active monitor, and for SO was 1 ppb using the passive monitor. The sampling precision for SO (RSD from personal measurements) was 5.0% for the passive monitor and was 13.4% for the active monitor over a range of exposures from 5-150 ppb. The corresponding precision for styrene was 5.3% for the passive monitor for levels ranging from 1.2 to 104 ppm. Measurements of 235 personal exposures with the active monitor in 12 facilities manufacturing fiberglass-reinforced plastics (FRP) showed that levels of AP and PAA were below 7.8 ppb and 5 ppb, respectively. In contrast, SO averaged 30.4 ppb (SE=2.4) in these FRP facilities, ranging from below 0.2 ppb to 190 ppb. The active monitor was also used to detect airborne SO at levels of approximately equals 1 ppb in one facility manufacturing styrene butadiene rubber, suggesting that SO is generally present during the polymerization of styrene. Personal passive monitoring in the 12 FRP facilities (n = 657) revealed mean concentrations of styrene ranging between 1.8 and 55.4 ppm, and for SO between 1.7 and 62.6 ppb. The ratio of the mean styrene level to the mean SO level varied between 449:1 and 1,635:1 among the 12 FRP facilities. PMID- 11253029 TI - Evaluation of airborne lead in the welding working environment. AB - Manual and automatic welding machines (which use leaded alloys) are considered to be important sources of the emission of lead fumes into the general air of the working environment. Three workplaces at a television factory were selected for the present study, to determine the control class of the working unit. The concentrations of conventional measurements ("A" sampling points) were lower than the administrative control level (statutory standard of lead, 150 microg m(-3)), whereas the maximum concentration of 264.1 microg m(-3) ("B" sampling point) was higher at one working unit than the administrative control level. However, the control classes varied between class III (bad) and class I (good). PMID- 11253030 TI - Indoor and outdoor formaldehyde concentrations in homes in residential areas in Greater Cairo. AB - Indoor and outdoor measurements of formaldehyde were conducted at seven flats located in residential areas in Greater Cairo, during spring and summer seasons 1999. The mean daytime formaldehyde concentrations in kitchens, bedrooms and living rooms were 89, 100 and 100 ppb, respectively, in the seven flats. Significant positive correlations were found between the concentrations of formaldehyde found in these three rooms. On the other hand, no significant differences were found between the mean formaldehyde concentrations in these three rooms. The maximum mean concentration of formaldehyde (147 ppb) was recorded in a new flat, while the minimum concentration (43 ppb) was observed in an old flat. The maximum hourly and daytime concentrations were 350 and 225 ppb, respectively. Air temperature, relative humidity and the age of the flat are factors affecting the emission and concentration of formaldehyde. The maximum indoor and outdoor formaldehyde concentrations were recorded during the summer season. During the spring, 38% of the samples indicated that the concentration of formaldehyde in the seven flats exceeded 0.1 ppm, the American Society of Heating, Refrigerating, and Air Conditioning Engineers' (ASHRAE) standard; in the summer, this figure increased to 53%. PMID- 11253031 TI - Microclimate monitoring of indoor environments using piezoelectric quartz crystal humidity sensors. AB - The aim of this paper is to describe indoor microclimate monitoring at two different locations, Sandham Memorial Chapel, in Hampshire, England, and the castle El Alcazar, in Segovia, Spain. Piezoelectric quartz crystal sensors with novel humidity sensitive poly(ethyleneimine) coatings and Pt resistance thermometers were used to measure the relative humidity (RH) and temperature gradients across one of the paintings of the British artist, Stanley Spencer, housed in Sandham Memorial Chapel. The measurement period extended from December 1997 to September 1998. Each coated crystal was set in its own housing from which temperature and RH measuring circuits were connected via a cable to the computer. The 9 month monitoring period incorporated the range in seasons from winter through to autumn. Between December and February the RH at the back of the painting was found to be lower than that at the front. In March and April the reverse was true. Additionally, there were large spikes in the data in some of the months for both RH and temperature, probably caused by direct sunlight falling on the sensors. At the second site monitoring was performed for a shorter period, from December 1997 to early January 1998. It served, however, to show that abrupt changes can occur in the microclimate surrounding the painting. These fluctuations can with time lead to alterations to the paint surface and result eventually in cracking and damage to the art work. PMID- 11253032 TI - Development of a calibration system to evaluate VOC losses in a branch enclosure. AB - Considerable uncertainties are associated with the experimental estimates of emission rates of different volatile organic compound (VOC) species from the biosphere to the atmosphere. Some of this uncertainty derives from the sampling and analytical procedures used in emission rate measurements. A calibration system was developed in order to evaluate possible errors in the measurements of biogenic emission rates using a branch enclosure system. Two types of calibration procedures were tested, a standard additions technique and an internal standard procedure. Both techniques were used to evaluate possible losses while sampling isoprene and monoterpenes, which are the most abundant VOCs of biogenic origin. The losses to Teflon lines and the empty sampling system were tested and losses to the branch enclosure system installed on two VOC emitting plant species were evaluated. A considerable loss of isoprene (approximately 18% of inflow concentration 65 ng l(-1)) to the empty enclosure system and to the system installed on the plant was measured, but no losses of monoterpenes were observed. PMID- 11253033 TI - Mercury and other elements in lichens near the INA Naftaplin gas treatment plant, Molve, Croatia. AB - This paper describes the use of epiphytic lichens as bioindicators for spatial monitoring of mercury and other elements in air near the natural gas treatment facilities at Molve, Croatia. It is well known that at this location the concentration of mercury in natural gas is very high and therefore it has to be removed from natural gas before further processing in order to prevent technological and environmental problems. In order to monitor the efficiency of an industrial facility for removal of mercury from natural gas, mercury measurements in air and lichens were performed during 9 months in 1997/1998. In situ lichens Parmelia sulcata, Xantoria parientina and Hypogymnia physodes and transplanted lichen species Hypogymnia physodes were used. A good correlation between mercury concentrations in air and lichens was found. The concentrations of barium and bromium were also significantly elevated in transplanted lichens, most probably related to technological process at the gas treatment plant and/or other geological factors. It was confirmed that lichens can successfully be used as bioindicators, provided a careful experiment is designed, particularly the amount of lichens transplanted, the duration of exposure and the initial levels and homogeneity of transplanted lichens. PMID- 11253034 TI - In vivo laser-induced fluorescence detection of pyrene in nematodes and determination of pyrene binding constants for humic substances by fluorescence quenching and bioconcentration experiments. AB - The bioconcentration of pyrene by bacterivorous thread worms (nematodes) of the species Caenorhabditis elegans was studied with laser-induced fluorescence (LIF) spectroscopy, fluorescence imaging and a radiotracer method. The vibronic band intensities of the LIF spectra indicated that the microenvironment of pyrene in the nematodes was similar to a low-polarity solvent, and thus provided direct evidence that pyrene was accumulated in lipid-rich areas inside the nematodes. The concentration of pyrene in the nematodes was estimated from the monomer/excimer fluorescence intensity ratio. Results from this method were in fair agreement with results using 14C labeled pyrene for measuring pyrene bioconcentration. Preliminary results indicated that LIF measurements of pyrene may be possible even in single nematodes. Fluorescence microscopic observations revealed that pyrene was not adsorbed on the outside of the organisms, but was strongly concentrated in restricted areas inside the worms. In the second part of the study, the effects of six different humic substances (HS) on the bioconcentration of pyrene were investigated and sorption coefficients (KDOC) calculated from reductions in bioconcentration (KDOC(biol)) were compared with sorption coefficients measured with a fluorescence quenching technique (KDOC(flu)). The results of these two different experimental methods agreed well (with KDOC(biol) being slightly lower than KDOC(flu), indicating that the fraction of pyrene that was determined as freely dissolved by the fluorescence quenching method was comparable to the bioavailable fraction. PMID- 11253035 TI - Spectrophotometric determination of ferbam (iron(III) dimethyldithiocarbamate) in commercial sample and wheat grains using 4,7-diphenyl-1,10-phenanthroline after extraction into mesityl oxide. AB - A procedure has been developed for the determination of iron(III) dimethyldithiocarbamate by converting it into iron(II)-bathophenanthroline complex, which is then extracted into mesityl oxide in the presence of potassium perchlorate. The absorbance is then measured at 534 nm against a reagent blank. Beer's law is obeyed over the concentration range 0.5-20 microg ml(-1) in the final solution. The method is sensitive, highly selective and can be used for the determination of ferbam in a commercial sample, and in mixtures with various dithiocarbamates (ziram, zineb, maneb, etc.) and in wheat grains. PMID- 11253036 TI - Quantification of protein adducts of hexahydrophthalic anhydride and methylhexahydrophthalic anhydride in human plasma. AB - Hexahydrophthalic anhydride (HHPA) and methylhexahydrophthalic anhydride (MHHPA) are two highly allergenic compounds used in the chemical industry. A method was developed for quantification of protein adducts of HHPA and MHHPA in human plasma. The plasma was dialysed and the anhydrides were hydrolysed from the proteins at mild acidic conditions. The released hexahydrophthalic acid (HHP acid) and methylhexahydrophthalic acid (MHHP acid) were purified by reversed solid phase extraction followed by derivatisation with pentafluorobenzyl bromide. The derivatives were analysed using GC-MS in negative ion chemical ionisation mode with ammonia as moderating gas. As internal standards, deuterium labelled HHP and MHHP acids were used. The detection limits were 0.06 pmol mL(-1) plasma for HHP acid and 0.03 pmol mL(-1) plasma for MHHP acid. The between-day precisions for HHP acid were 18% at 0.3 pmol mL(-1) and 8% at 4 pmol mL(-1). For MHHP acid, the precisions were 13% at 0.3 pmol mL(-1) and 9% at 4 pmol mL(-1). There were strong correlations (r=0.94 for HHPA and 0.99 for MHHPA) between total plasma protein adduct concentrations and serum albumin adduct levels. Workers exposed to time-weighted average air levels of HHPA between < 1 and 340 microg m( 3) and between 2 and 160 microg m(-3) for MHHPA had plasma adduct levels between the detection limits of the methods and 8.40 and 19.0 pmol mL(-1), respectively. PMID- 11253037 TI - Ascorbic acid reduction of active chlorine prior to determining Ames mutagenicity of chlorinated natural organic matter (NOM). AB - Many potable water disinfection byproducts (DBPs) that result from the reaction of natural organic matter (NOM) with oxidizing chlorine are known or suspected to be carcinogenic and mutagenic. The Ames assay is routinely used to assess an overall level of mutagenicity for all compounds in samples from potable water supplies or laboratory studies of DBP formation. Reduction of oxidizing disinfectants is required since these compounds can kill the bacteria or react with the agar, producing chlorinated byproducts. When mutagens are collected by passing potable water through adsorbing resins, active chlorine compounds react with the resin, producing undesirable mutagenic artifacts. The bioanalytical and chemoanalytical needs of drinking water DBP studies required a suitable reductant. Many of the candidate compounds failed to meet those needs, including 2,4-hexadienoic (sorbic) acid, 2,4-pentanedione (acetylacetone), 2-butenoic (crotonic) acid, 2-butenedioic (maleic and fumaric) acids and buten-2-ol (crotyl alcohol). Candidates were rejected if they (1) reacted too slowly with active chlorine, (2) formed mutagenic byproducts, or (3) interfered in the quantitation of known chlorination DBPs. L-Ascorbic acid reacts rapidly and stoichiometrically with active chlorine and has limited interactions with halogenated DBPs. In this work, we found no interference from L-ascorbic acid or its oxidation product (dehydroascorbic acid) in mutagenicity assays of chlorinated NOM using Salmonella typhimurium TA100, with or without metabolic activation (S9). This was demonstrated for both aqueous solutions of chlorinated NOM and concentrates derived from the involatile, ether-extractable chlorinated byproducts of those solutions. PMID- 11253038 TI - Comparison of degradation state and stability of different humic acids by means of chemolysis with tetramethylammonium hydroxide. AB - In the chemolysis products of extracted humic acids (HAs) and tetramethylammonium hydroxide, different compounds can be identified which allow a description of the sources of soil organic matter (SOM). It is possible to draw conclusions concerning stability, degree of intramolecular cross-linking and degradation of aliphatic and lignin components from the distinctive product ratios. The ratio of lignin derived acidic phenolic derivatives and their analogous aldehydes and the ratio of phenylpropenoic acids and analogous benzoic acids provide comparably good parameters for the characterization of the state of degradation of lignin compounds to relatively stable HA building blocks. Regarding the aliphatic chemolysis products, alpha,omega-dicarboxylic acid and methoxy fatty acid contents indicate an intramolecular cross-linking state within the HA molecules and the degree of stable aliphatic constituent parts. High contents of unsaturated fatty acids can be considered as an indicator of an easy degradable humic skeleton because with their double bonds they represent active sites for further transformations. The suitability of the carbon preference index as an indicator of a biogenic carbon source can be confirmed. It is remarkable how the results obtained from extracted humic acids are in accordance with the expectations for the different SOMs derived from land use. PMID- 11253039 TI - Comparison study of five analytical methods for the fractionation and subsequent determination of aluminium in natural water samples. AB - Five methods for aluminium fractionation used in different laboratories in Norway and Finland were compared using six control, 75 soil water and 10 lake water samples. Different fractionation principles [cation exchange, formation of the Pyrocatechol Violet (PCV) or quinolin-8-ol (oxine) complex], types of cation exchanger [Amberlite (Na/H) or Bond Elut (H)], reaction time (from 2.3 s), flow systems (flow injection analysis or segmented flow) and determination principles (molecular absorption spectrometry or ICP-AES) were tested. Determination of the 'labile' fraction was strongly dependent on the method used and the largest differences were observed between the ICP-AES method with cation exchange (Bond Elut H form) and the 'quickly reacting' method (oxine, 2.3 s). Different flow systems, both using cation exchange and determination of the PCV complex but with different reaction times and an extra acidification step, resulted in large differences in the 'reactive' and 'non-labile' fractions determined. However, the determination of the labile fraction gave similar results with both these methods. The two different types of cation exchanger used (with and without pH buffering and with different counter ions) in the ICP-AES methods resulted in differences, mainly because of a smaller 'non-labile' fraction in the non buffered system. The two flow injection systems (oxine and PCV complexation) showed common trends, which may be connected with the short reaction times used. Comparison with theoretical equilibrium calculations using the model ALCHEMI suggested that the best correlation for the determination of the 'labile' fraction were obtained with the ICP-AES method with an Amberlite column. PMID- 11253040 TI - A field test for the assessment of faecal contamination of potable water. AB - A simple, sensitive, rapid, inexpensive paper strip impregnated with Salmonellal E. coli medium (SEM) was formulated, and placed in a test tube. When potable water of 10 ml was added to the test tube it detected the faecal contamination of water samples within 16-48 h when incubated at room temperature from 20 to 35 degrees C. The positive results were indicated when the medium turned black (hydrogen sulfide production) for the presence of Salmonella sp. and/or the formation of a red ring (free indole from tryptophan) when a few drops of Kovac's reagent was added for the presence of coliform bacteria (E. coli). More than 600 water samples were tested with the new test (SEM) and results showed 99% agreement with that of the standard most probable number (MPN) coliform test and also proved highly successful in the field when it was employed to detect both Salmonella and E. coli. Different water testing laboratories involved in a water quality monitoring programme and governmental agencies evaluated the test media and reported that the test was user friendly, reliable and simple to perform in the field and will be especially useful for screening of both urban and rural water supplies for routine monitoring of bacteriological contamination. PMID- 11253041 TI - A study of the distribution of lead, cadmium and copper between water and kaolin, bentonite and a river sediment. AB - Experiments were carried out to monitor the equilibrium distribution of lead, cadmium and copper between an aqueous phase modelling natural water and a solid phase modelling natural sediment, under varying conditions. The aqueous phase was analysed using ETAAS and differential pulse anodic stripping voltammetry (DPASV), whereas XRD and FTIR were used to study the solid phase. Sorption isotherms at constant pH were measured. Conditional distribution constants were calculated as functions of the pH, the time of equilibration and the amount of solid material. The results obtained stress the need for standardization of the approaches to the study of water-sediment interactions in order to be able to evaluate and compare the extensive data from field measurements and to predict these interactions. PMID- 11253042 TI - A novel sensor for monitoring leakage of petroleum and other liquid hydrocarbons into soil environments. AB - This paper describes the use of a potentially implantable infrared reflectometer for the qualitative detection of petroleum and a number of other hydrocarbon solvents. A rugged, low-power, re-useable sensor was evaluated in the laboratory for its ability to detect petrol in soil. A hydrophobic fluoropolymer was used as the sensing surface due to its high selectivity for petroleum hydrocarbons. The photocurrent reflected by this surface from a near IR source was measured to test for petroleum saturation within the membrane, which in turn was an indicator of petroleum in the surrounding soil. The simplicity in the sensor design enabled a stable, low cost detection method for petroleum and other hydrocarbons, ideal for use in sub-surface applications. PMID- 11253043 TI - The invisible pollutant: health effects of EMFs. PMID- 11253044 TI - EU explains precaution principle. PMID- 11253045 TI - Liability proposals find few friends. PMID- 11253046 TI - Pesticides residues in food. PMID- 11253047 TI - Bioavailability: coming of age? PMID- 11253048 TI - Methodologies for determination of antimony in terrestrial environmental samples. AB - Methodologies for the environmental analysis of total antimony and aqueous chemical speciation are critically reviewed, including preparation techniques for aqueous and solid matrices and the determination of solid state partitioning and recommendations are given for future research directions. Concentrations of total antimony commonly present in aqueous and solid environmental samples are readily determined using present day analytical techniques. This has resulted primarily from technological advances in microwave digestion for solid matrices and the development of plasma based analyte detection systems. ICP-AES and ICP-MS techniques are both utilised for the environmental analysis of total antimony concentrations. However, ICP-MS is increasingly favoured as a result of reduced spectral interferences and the potential for analyte detection in the pg mL(-1) range. Determination of aqueous antimony speciation presents a number of complex analytical challenges and highly selective separation and identification techniques are required prior to detection. The majority of published techniques including common applications of hydride generation are insufficiently selective for the determination of intrinsic chemical speciation and often only oxidation state data are obtained. The recent in-line applications of HPLC-ICP-MS offer the potential for highly selective separations of aqueous antimony species and determination of detailed chemical speciation data. However, considerable development work is required to optimise chromatographic separations and identify uncharacterised species resident in environmental systems. Analytical techniques to aid the determination of antimony's associations with solid environmental matrices include the application of chemical extraction procedures and leaching experiments. To date, this area of analytical research has received little attention and further studies are required to elucidate this aspect of antimony's environmental chemistry. PMID- 11253049 TI - Genomic strategies to identify mammalian regulatory sequences. AB - With the continuing accomplishments of the human genome project, high-throughput strategies to identify DNA sequences that are important in mammalian gene regulation are becoming increasingly feasible. In contrast to the historic, labour-intensive, wet-laboratory methods for identifying regulatory sequences, many modern approaches are heavily focused on the computational analysis of large genomic data sets. Data from inter-species genomic sequence comparisons and genome-wide expression profiling, integrated with various computational tools, are poised to contribute to the decoding of genomic sequence and to the identification of those sequences that orchestrate gene regulation. In this review, we highlight several genomic approaches that are being used to identify regulatory sequences in mammalian genomes. PMID- 11253050 TI - Post-transcriptional gene silencing by double-stranded RNA. AB - Imagine being able to knock out your favourite gene with only a day's work. Not just in one model system, but in virtually any organism: plants, flies, mice or cultured cells. This sort of experimental dream might one day become reality as we learn to harness the power of RNA interference, the process by which double stranded RNA induces the silencing of homologous endogenous genes. How this phenomenon works is slowly becoming clear, and might help us to develop an effortless tool to probe gene function in cells and animals. PMID- 11253051 TI - Gliomagenesis: genetic alterations and mouse models. AB - Glioblastoma multiforme is the most malignant of the primary brain tumours and is almost always fatal. The treatment strategies for this disease have not changed appreciably for many years and most are based on a limited understanding of the biology of the disease. However, in the past decade, characteristic genetic alterations have been identified in gliomas that might underlie the initiation or progression of the disease. Recent modelling experiments in mice are helping to delineate the molecular aetiology of this disease and are providing systems to identify and test novel and rational therapeutic strategies. PMID- 11253052 TI - Genetic dissection of phenotypic diversity in farm animals. AB - Farm animal populations harbour rich collections of mutations with phenotypic effects that have been purposefully enriched by breeding. Most of these mutations do not have pathological phenotypic consequences, in contrast to the collections of deleterious mutations in model organisms or those causing inherited disorders in humans. Farm animals are of particular interest for identifying genes that control growth, energy metabolism, development, appetite, reproduction and behaviour, as well as other traits that have been manipulated by breeding. Genome research in farm animals will add to our basic understanding of the genetic control of these traits and the results will be applied in breeding programmes to reduce the incidence of disease and to improve product quality and production efficiency. PMID- 11253053 TI - Science or alchemy? AB - Hyperbole has become a common and accepted practice in science nowadays. We sell our results, we hide our ignorance and we use stock terms that gain spurious weight through repeated use. I illustrate from the field of developmental genetics. PMID- 11253054 TI - At the interfaces of epidemiology, genetics and genomics. AB - You come onto the court prepared for tennis but your partner seems to be ready for rugby. Neither of you is at all sure what it is that your opponent wants to play. The only recourse is to teach each other the rules of your own game and then decide whether you can collectively invent a new sport. Welcome to the dialogue at the intersections of epidemiology with genetics and genomics. PMID- 11253055 TI - Beyond consent: ethical and social issues in genetic testing. AB - Informed consent is a vital ethical doctrine in clinical medicine and, through genetic counselling, is being applied to genetic testing. But genetic testing raises issues that transcend the traditional concept of informed consent. Genetic tests are adopted without demonstrable clinical benefit, and the consequences of testing can reach beyond the individual to their families and communities. Understanding the social and cultural context of genetic testing will lead to more informed discussion and debate on these issues. PMID- 11253056 TI - Ageing. The fly that won't die. PMID- 11253057 TI - Developmental biology. Shaping gradients. PMID- 11253058 TI - Technology. ChIP on chips. PMID- 11253059 TI - The secret is in the head. PMID- 11253060 TI - Cancer biology. Manipulating the system. PMID- 11253061 TI - Alzheimer hotspot on 10. PMID- 11253062 TI - Association study designs for complex diseases. AB - Assessing the association between DNA variants and disease has been used widely to identify regions of the genome and candidate genes that contribute to disease. However, there are numerous examples of associations that cannot be replicated, which has led to skepticism about the utility of the approach for common conditions. With the discovery of massive numbers of genetic markers and the development of better tools for genotyping, association studies will inevitably proliferate. Now is the time to consider critically the design of such studies, to avoid the mistakes of the past and to maximize their potential to identify new components of disease. PMID- 11253063 TI - Quantitative trait loci in Drosophila. AB - Phenotypic variation for quantitative traits results from the simultaneous segregation of alleles at multiple quantitative trait loci. Understanding the genetic architecture of quantitative traits begins with mapping quantitative trait loci to broad genomic regions and ends with the molecular definition of quantitative trait loci alleles. This has been accomplished for some quantitative trait loci in Drosophila. Drosophila quantitative trait loci have sex-, environment- and genotype-specific effects, and are often associated with molecular polymorphisms in non-coding regions of candidate genes. These observations offer valuable lessons to those seeking to understand quantitative traits in other organisms, including humans. PMID- 11253064 TI - Genomic imprinting: parental influence on the genome. AB - Genomic imprinting affects several dozen mammalian genes and results in the expression of those genes from only one of the two parental chromosomes. This is brought about by epigenetic instructions--imprints--that are laid down in the parental germ cells. Imprinting is a particularly important genetic mechanism in mammals, and is thought to influence the transfer of nutrients to the fetus and the newborn from the mother. Consistent with this view is the fact that imprinted genes tend to affect growth in the womb and behaviour after birth. Aberrant imprinting disturbs development and is the cause of various disease syndromes. The study of imprinting also provides new insights into epigenetic gene modification during development. PMID- 11253065 TI - A bigger needle in a small haystack. PMID- 11253066 TI - Ancient origin of the Hox gene cluster. AB - The Hox gene cluster has a crucial function in body patterning during animal development. How and when this gene cluster originated is being clarified by recent data from Cnidaria, a basal animal phylum. The characterization of Hox like genes from Hydra, sea anemones and jellyfish has revealed that a Hox gene cluster is extremely ancient, having originated even before the divergence of these basal animals. PMID- 11253067 TI - Zebrafish genetics and vertebrate heart formation. AB - Forward-genetic analyses in Drosophila and Caenorhabditis elegans have given us unprecedented insights into many developmental mechanisms. To study the formation of organs that contain cell types and structures not present in invertebrates, a vertebrate model system amenable to forward genetics would be very useful. Recent work shows that a newly initiated genetic approach in zebrafish is already making significant contributions to understanding the development of the vertebrate heart, an organ that contains several vertebrate-specific features. These and other studies point to the utility of the zebrafish system for studying a wide range of vertebrate-specific processes. PMID- 11253068 TI - Epigenetics. Satellite tools. PMID- 11253069 TI - Regulating cholesterol by A, B, C. PMID- 11253070 TI - Rewiring the keyboard: evolvability of the genetic code. AB - The genetic code evolved in two distinct phases. First, the 'canonical' code emerged before the last universal ancestor; subsequently, this code diverged in numerous nuclear and organelle lineages. Here, we examine the distribution and causes of these secondary deviations from the canonical genetic code. The majority of non-standard codes arise from alterations in the tRNA, with most occurring by post-transcriptional modifications, such as base modification or RNA editing, rather than by substitutions within tRNA anticodons. PMID- 11253071 TI - X-chromosome inactivation: counting, choice and initiation. AB - In many sexually dimorphic species, a mechanism is required to ensure equivalent levels of gene expression from the sex chromosomes. In mammals, such dosage compensation is achieved by X-chromosome inactivation, a process that presents a unique medley of biological puzzles: how to silence one but not the other X chromosome in the same nucleus; how to count the number of X's and keep only one active; how to choose which X chromosome is inactivated; and how to establish this silent state rapidly and efficiently during early development. The key to most of these puzzles lies in a unique locus, the X-inactivation centre and a remarkable RNA--Xist--that it encodes. PMID- 11253072 TI - Genomics. Weed it and reap. PMID- 11253073 TI - Functional genomics. Comprehensive interference.. PMID- 11253074 TI - Biotechnology in the 1930s: the development of hybrid maize. AB - Hybrid maize was one of the first examples of genetic theory successfully applied to food production. When first introduced, it seemed almost miraculous; study hybrids convinced skeptical farmers that 'the professors' and their arcane science could do them some good. Strangely, the genetic basis of heterosis (hybrid vigour) was and still is unknown. But to this day, newer hybrids continue to outyield their predecessors; they do so because they are tougher and healthier. PMID- 11253075 TI - Cracking conditional transgenesis. PMID- 11253076 TI - Human embryonic stem cell research: ethical and legal issues. AB - The use of human embryonic stem cells to replace damaged cells and tissues promises future hope for the treatment of many diseases. However, many countries now face complex ethical and legal questions as a result of the research needed to develop these cell-replacement therapies. The challenge that must be met is how to permit research on human embryonic tissue to occur while maintaining respect for human life generally. PMID- 11253077 TI - Multifactorial genetics. A map of fear. PMID- 11253078 TI - Gene therapy. Precision and control. PMID- 11253079 TI - Gene expression. Escaping silence. PMID- 11253080 TI - [Use of VR (virtual reality) software for preoperative implantation fitting with an implantable hearing aid as an example]. AB - PURPOSE: To prove the feasibility of a preoperative fitting test for an implantable hearing aid using a VR environment. METHODS: A high-resolution spiral CT was performed after mastoidectomy in 10 temporal bone specimens. The bony structures were segmented and merged with the Computer-Aided Design (CAD) data of the hearing aid in a VR environment. For each specimen a three-dimensional fitting test was carried out by three examiners determining the implantability of the hearing aid. The implantation simulation was compared with the real implantation procedure performed by an experienced ENT surgeon. RESULTS: The used VR system enabled real-time 3D-visualisation and manipulation of CT- and CAD data. All objects could be independently moved in all three dimensions. The VR fitting test corresponded closely with the real implantation. The implantability of the hearing aid was properly predicted by all three examiners. CONCLUSION: Merging CT and CAD data in a virtual reality environment bears high potential for the pre-surgical determination of the fit and mountability of medical implants in complex anatomical regions. PMID- 11253081 TI - [Diagnosis of urethral diverticula and periurethral masses]. AB - AIM: To give recommendations for the standard diagnostic assessment of urethral diverticula and periurethral masses based on an evaluation of our results and a survey of the recent literature. METHODS: Group I (1981-1993) included 47 women in whom urethral diverticula (n = 34), periurethral cysts (n = 11), and periurethral leiomyomas (n = 2) were diagnosed and the results compared with the intraoperative findings. Diagnostic work-up comprised history taking, vaginal palpation, introitus ultrasound, double-balloon urethrography (DBU), voiding cystourethroscopy (VCU), excretion urography (EU), and urethrocystoscopy. Group II (1994-1996) included 12 women with urethral diverticula who were examined by DBU, 2D and 3D introitus ultrasound. The diagnostic accuracy of the different methods was assessed. RESULTS: Group I: The diagnostic accuracy in identifying urethral diverticula was 85.3% for palpation, 61.5% for introitus ultrasound, 93.8% for DBU, 37.5% for EU, and 30.0% for urethrocystoscopy. Of all imaging modalities used, only introitus ultrasound depicted periurethral cysts and leiomyomas. Group II: The DBU, 2D and 3D introitus ultrasound had a diagnostic accuracy of 100% in identifying urethral diverticula. CONCLUSIONS: Introitus ultrasound should be used as the basic diagnostic tool in clinically suspected urethral diverticula or periurethral masses and additional DBU should be restricted to cases with inconclusive findings. PMID- 11253082 TI - [Gd-BOPTA enhanced excretory MR urography without administration of diuretics]. AB - PURPOSE: To evaluate the feasibility and clinical utility of Gd-BOPTA enhanced excretory magnetic resonance urography without additional administration of diuretics in correlation with conventional urography. METHOD: 15 preoperative patients with pelvic tumors were examined at 1.5 T using a breath-hold high resolution 3D-FLASH sequence during first-pass as well as 5, 10, 15 minutes after i.v. injection of 0.05 mmol/kg BW Gd-BOPTA (MultiHance) without administration of diuretics. Post-processed coronal and multiplanar MIP reconstructions were compared to conventional excretory urography with regard to morphologic accuracy, anatomic variability, filling defects, cause and level of obstruction or compression, tumor visibility, and time-effectiveness by two independent radiologists. RESULTS: Visualization of the urinary tract by MRU was comparable to conventional excretory urography in 14 of 15 cases. Caliceal fornices were better delineated on conventional urographies, whereas MRU was considered superior in the assessment of the inferior ureter sections, the urinary bladder and obstructive tumors, whose extents could be clearly marked out. Examination times of both techniques were comparable. CONCLUSION: These first results show that non-diuretic Gd-BOPTA enhanced MRU is comparable to conventional excretory urography for the preoperative diagnosis of pelvic tumors. Further improvements of this technique seem possible by optimization of examination intervals and injection doses. PMID- 11253083 TI - [Signal changes of the bone marrow in MRI under long-term treatment with granulocyte colony-stimulating factors]. AB - PURPOSE: Recurrent infections in patients with glycogen storage disease (GSD) type Ib resulting from an associated neutropenia are frequently treated with granulocyte colony-stimulating factors (G-CSF). The aim of this study was to evaluate the changes occurring in bone marrow by magnetic resonance imaging (MRI) in these patients. MATERIAL AND METHODS: The distal femoral and tibial bones of six patients with GSD Ib were evaluated by MRI. Four of these patients were treated with G-CSF for at least 3.9 to a maximum of 8.2 years (mean 5.8 years). The imaging sequences encompassed spin-echo as well as short-time inversion recovery sequences. 4 of the 6 patients had bone marrow aspirations. RESULTS: The patients who had undergone therapy with G-CSF showed a marked increase in signal strength in STIR sequences which encompassed the entire medullar cavity. In T1 weighted images these areas were hypointense. Biopsies obtained from these patients showed a bone marrow hypercellularity. The patients without G-CSF therapy showed the same signal intensity changes but with a more discrete and localized pattern in the metaphyseal cavities. CONCLUSION: In subjects with GSD Ib, an increased myelopoetic activity of the bone marrow which is intensified under long-term treatment with G-CSF can be demonstrated by MRI. PMID- 11253084 TI - [Value of kinematic MRI in the evaluation of patients with exacerbated pain in cervical spine motion compared with static MRI]. AB - OBJECTIVE: To assess the clinical value of kinematic MR imaging in patients with cervical radiculopathy and increasing symptoms after provocative maneuvers at flexion, extension, axial rotation and coupled motion of the cervical spine. METHODS: Thirty-five patients with cervical disc herniation or cervical spondylosis in whom symptoms were elicited at flexion, extension, axial rotation and coupled motion were studied inside a positioning device using T2-weighted TSE, 2D-FLASH, and reformatted 3D DESS and 3D-FISP sequences. The images were evaluated for the size of disc herniations, the foraminal size and cervical cord displacement at provocative position compared with neutral position (0 degree). In addition, the value of kinematic MR images were interpretated with regard to changes in the therapeutic procedure and intraoperative patient positioning. RESULTS: Compared with the neutral position (0 degree) a change in disc herniations was not found in any (0%) of the provocative positions. In five patients (14%) cervical cord displacement was noted at axial rotation. The foraminal size varied depending on the provocative position, increasing at flexion and decreasing at extension. CONCLUSION: Kinematic MR imaging in patients with cervical radiculopathy and increasing symptoms at provocative maneuvers provides no additional information for the therapeutic decision-making process. PMID- 11253085 TI - [Comparison of 3D power doppler ultrasound, color doppler ultrasound and digital subtraction angiography in carotid stenosis]. AB - PURPOSE: To compare easy-to-perform three-dimensional power Doppler ultrasound (3D PDUS) to color Doppler ultrasound (CDUS) and digital subtraction angiography (DSA) in the assessment of internal carotid artery (ICA) stenoses in patients with severe atherosclerosis. METHODS: 26 ICA's (7 without stenosis, 4 low-, 4 middle-, and 11 high-grade stenoses) in 13 patients were examined with DSA, CDUS, and 3D PDUS. CDUS and 3D PDUS were performed with a 7.5 MHz standard transducer and a Sonoline Elegra ultrasound machine. The three methods were performed and interpreted by different persons who were not aware of the diagnoses. RESULTS: Regarding the degree of stenosis correlations between DSA and 3D PDUS were r = 0.98 (p < 0.001), between DSA and CDUS r = 0.97 (p < 0.001), and between CDUS and 3D PDUS r = 0.95 (p < 0.001). Sensitivity and specificity regarding the detection of a high-degree stenosis was 90% and 100% for 3D PDUS, and 100% and 93.3% for CDUS. CONCLUSION: Even in cases with severe atherosclerosis, both sonographic methods reveal similar results comparable to DSA. 3D PDUS does not result in a diagnostic improvement on CDUS, however, it does give the new opportunity for complete data storage, reconstruction, and survey presentations. PMID- 11253087 TI - [Quality assessment using AGIR: results and experience]. AB - PURPOSE: To evaluate whether a new software from the working group for interventional radiology (AGIR) is an appropriate tool for quality assurance in interventional radiology, and presentation of results acquired within the quality improvement process in 1999. PATIENTS AND METHODS: AGIR-defined parameters such as patient data, risk profile, given interventions as well as complications were registered by a recently developed software. Based on monthly data analyses, possible complications were identified and discussed in morbidity and mortality conferences. RESULTS: 1014 interventions were performed in our institution in 1999. According to criteria established by AGIR, the complication rate was 2.7%. In addition and according to SCVIR criteria, complications were distinguished quantitatively in five classes and semiquantitatively in minor and major groups. The result was a minor complication rate of 1.8%, and a major rate of 0.9%. There were no cases of death associated with the intervention. Further strategies were developed in order to reduce the complication rate. CONCLUSION: Extensive quality assurance methods can be integrated in daily routine work. These methods lead to an intensive transparency of treatment results, and allow the implementation of continuous quality improvements. The development of the software is a first step in establishing a nation-wide quality assurance system. Nevertheless, modification and additional definition of the AGIR predefined parameters are required, for example, to avoid unnecessary procedures. PMID- 11253086 TI - [Cerebral protection with balloon occlusion during carotid artery stent implantation--first experiences]. AB - PURPOSE: To assess the technical feasibility and the results of cerebral protection with the GuardWire Plus Temporary Occlusion & Aspiration System during carotid artery stenting for high-grade stenosis. PATIENTS AND METHODS: In 20 patients 20 carotid artery stenoses were treated with stent placement under cerebral protection. A contralateral carotid occlusion was an exclusion criteria for the use of the protection device. In all cases only aspiration, but no flushing was used before deflation of the occlusion balloon. In 17 of 20 patients diffusion-weighted (DW-)MRT imaging of the brain was performed before and 24 hours after the procedure. RESULTS: The stent implantation was successfully performed in all patients. In 3 patients neurologic symptoms occurred during the occlusion time. In these 3 patients the symptoms immediately disappeared after deflation of the balloon. In one case there was dilatation of the internal carotid artery at the site of the balloon inflation. In 3 of the 17 DW-MR images new ipsilateral cerebral lesions, in one case a new contralateral lesion occurred after the procedure. CONCLUSIONS: The cerebral protection procedure is technically feasible. The occlusion of the internal carotid artery was not tolerated by all patients. The DW-MR imaging demonstrated cerebral lesions indicating the occurrence of cerebral microemboli during the procedure. Further investigations are necessary to determine if the use of the cerebral protection device will improve the results of the carotid artery stenting for high-grade stenoses. PMID- 11253088 TI - Late distal perigraft endoleak after endovascular repair of an abdominal aortic aneurysm due to cranial migration of the iliac branch of a modular stent-graft. PMID- 11253089 TI - [Intramural duodenal hematoma after endoscopic biopsy]. PMID- 11253090 TI - [Unusual "contrast media extravasation" in separation urography]. PMID- 11253091 TI - [On: Indirect MR arthrography in the diagnosis rotator cuff injuries. By J. Rudolph, Berlin et. al. In: Fortsch Rontgenstr 2000; 172: 686-691]. PMID- 11253092 TI - [Radiation exposure in x-ray mammography]. AB - When discussing the radiation risk of X-ray mammography, the magnitude of the dose applied has decisive importance. The radiation exposure of the breast is the predominant factor in risk considerations, since it contributes more than 98% to the effective dose of this examination. At present, it is generally assumed that, with regard to cancer induction by ionizing radiation, the glandular tissue is the most vulnerable part in the breast. Therefore, the average glandular dose, i.e., the mean value of the absorbed dose in the glandular tissue, is used for a description of the radiation risk. The average glandular dose cannot be measured directly, but is calculated under certain assumptions from the experimentally determined entrance surface air kerma or entrance surface dose by the use of a so called conversion factors. During the seventies, i.e., in the era of the industrial type X-ray film, the mean value of the average glandular dose per exposure for a larger sample of patients (n > 100) was about 20 mGy. Due to the progress in radiographic technique such as, for example, the use of sensitive film-screen systems, optimized radiation qualities and modern automatic exposure control units this value has now decreased to about 1 mGy. Further dose reductions seem possible by the introduction of digital image receptors. PMID- 11253093 TI - [Percutaneous transpapillary extraction of biliary calculi for symptomatic choledocholithiasisafter unsuccessful endoscopic treatment]. AB - PURPOSE: Evaluation of a percutaneous transhepatic treatment of symptomatic choledocholithiasis in bile ducts that cannot be reached with the endoscope. METHODS: From January 1996 to August 2000 a transhepatic extraction of biliary calculus was performed in four patients. Endoscopic retrograde cholangiography (ERC) was not successful in any of the cases. Clinical symptoms were icterus in four cases, additional cholangitis or colics in two cases. First, a balloon dilation of the papilla was performed by a percutaneous transhepatic approach. For removal of bile duct stones, occlusion catheters and Dormia baskets were used. Technical success was defined as complete removal of bile duct stones. Clinical success was defined as normalization of cholestasis and inflammation parameters. In the follow-up an ultrasound examination was performed and blood samples were taken for control of cholestasis parameters. RESULTS: In all four cases treatment was technically and clinically successful. For complete removal of biliary calculus a second intervention was necessary in two cases. In each case an internal to external drainage was left over a mean of 7 days (3-13 days). In the mean follow-up of 30.5 months (6-50 months) all patients had persistent relief of symptoms. No further interventions were necessary. No complications were present. CONCLUSION: Percutaneous transpapillary extraction of biliary calculus is an effective alternative to surgery in patients with bile ducts, that cannot be reached with the endoscope. PMID- 11253095 TI - [Experimental radiology in the USA]. PMID- 11253094 TI - [Treatment of arterial traumas by the Wallgraft endoprosthesis]. AB - PURPOSE: Percutaneous peripheral interventional procedures as well as coronary interventions can be complicated by dissections and traumatic lesions of peripheral arteries. The aim of this study was to evaluate the efficacy of treatment for traumatic peripheral arterial lesions. MATERIAL AND METHODS: In this study we used the Wallgraft-Endoprothesis (Boston Scientific, USA), which is a self-expanding covered stent. In 17 patients a total number of 24 endoprostheses (mean length 6.4 cm) were implanted in iliac arteries. Indications for stenting were large dissections (n = 9), arterial perforations (n = 4), aneurysms (n = 3), and stent in stent implantation (n = 2). RESULTS: An immediate exclusion of the lesion could be achieved in all cases. There were no major procedural complications. The primary patency after a mean follow-up of 18 months was 82.4% (14/17). Early reocclusion was seen in two cases, one stent in stent reocclusion and one reocclusion after acute stent thrombosis. In one other case the angiography revealed relevant restenosis (> 75%). The patency could be restored in one of these three cases leading to a secondary patency rate of 94.1%. CONCLUSIONS: The Wallgraft-Endoprotheses seems to be safe and effective to seal large dissections and traumatic lesions of peripheral arteries, showing a high long-term patency rate. PMID- 11253096 TI - [Molecular imaging in magnetic resonance tomography and nuclear medicine]. AB - The identification of genetic and biochemical changes allows a more conclusive characterization and classification of disease. Up to now this information is mostly obtained through in vitro analysis after resection or biopsy by immunohistopathology and molecular biology. There is a definite need for non invasive detection and repeated monitoring of such changes in experimental research as well as in clinical trials. Therefore, it is necessary to develop radiological imaging techniques that not only visualize morphologic and physiologic alterations, but track genetic and biochemical processes. This short review reports some of the various ongoing research projects that address this problem and provide some very promising approaches. PMID- 11253097 TI - [Experimental and clinical approaches to lymph node imaging]. AB - Exact assessment of lymph nodes is crucial to tumor staging, choice of therapy and in predicting the outcome. Although imaging plays a central role in the evaluation of lymph nodes, current imaging methods have low sensitivity and specificity primarily because they rely on insensitive morphological criteria or because they have low special resolution. Because of this diagnostic dilemma invasive, expansive and uncomfortable diagnostic techniques and/or unnecessary aggressive therapies are still in use. This brief overview is intended to summarize current imaging strategies and to give an outlook on experimental and clinical strategies in lymph node imaging in cancer. PMID- 11253098 TI - [Progress in optical imaging]. AB - Different optical imaging technologies have significantly progressed over the last years. Besides advances in imaging techniques and image reconstruction, new "smart" optical contrast agents have been developed which can be used to detect molecular targets (such as endogenous enzymes) in vivo. The combination of novel imaging technologies coupled with smart agents bears great diagnostic potential both clinically and experimentally. This overview outlines the basic principles of optical imaging and summarizes the current state of the art. PMID- 11253099 TI - [Imaging of angiogenesis]. AB - Angiogenesis of tumors has gained tremendous attention in the research community over the last years. Novel anti-angiogenic treatment protocols are currently in various phases of clinical testing. Thus, the major challenge for Radiological diagnostics is to develop imaging methods which reliably measure the angiogenic tumor burden as well as tumor response to anti-angiogenic treatment in vivo. This article intends to provide a brief overview of imaging strategies for angiogenesis and anti-angiogenic treatment. PMID- 11253100 TI - [Experimental flow and perfusion measurements in the animal model with magnetic resonance tomography]. AB - AIM: Validation of non-invasive methods for morphologic and functional imaging of the kidney under physiologic and pathophysiologic conditions. MATERIAL AND METHODS: In chronically instrumented animals (foxhounds) comparative measurements of renal flow and perfusion were performed. Magnetic resonance imaging techniques were compared to data obtained from implanted flow probes and total kidney weight post mortem. In the MR system, different degrees of renal artery stenosis could be induced by means of an implanted inflatable cuff. The degree of stenosis was verified with high-resolution 3D contrast-enhanced MR angiography (3D-CE-MRA) using an intravascular contrast agent. RESULTS: The MR-data agreed well with the invasively obtained results. Artifacts resulting from the implanted flow probes and other devices could be kept to a minimum due to appropriate selection of the probe materials and measurement strategies. Stenoses could be reproduced reliably and quantified from the induced morphologic and functional changes. CONCLUSION: Morphologic and functional MR techniques are well suited for non-invasive in vivo assessment of renal blood flow physiology. PMID- 11253101 TI - [Static-dynamic MR urography. Comparison with excretory urography and scintigraphy in experimentally-induced urinary tract obstruction]. AB - PURPOSE: To assess the diagnostic value of combined static-dynamic MR urography (MRU) for the functional-morphological evaluation of experimentally induced urinary tract obstruction. METHODS: Static-dynamic MRU--combination study with a respiratory-triggered 3D-IR-TSE sequence and a dynamic 2D-FFE sequence after Gd DTPA and furosemide--was obtained in comparison with 99mTc-MAG3 diuretic renal scintigraphy (DRS), excretory urography (EU) and ultrasound (US) in 29 healthy piglets and in 20 piglets with surgically induced ureteric stenosis (total of 50 postoperative examination blocks). RESULTS: MRU allowed complete depiction of the urinary tract in all controls, in operated piglets the stenosis was always correctly identified. Quality of MRU was superior to EU in 36 of 43 comparative studies. Calculation of single kidney function from parenchymal renograms, and assessment of urinary excretion from whole-kidney renograms resulted in a highly significant agreement of MRU with DRS. CONCLUSION: Static-dynamic MR urography allows excellent depiction of experimentally induced urinary tract obstruction, and reliable assessment of individual renal function and urinary excretion. Two advantages of the method stand out, it does not require radiation and it permits a functional-morphological correlation. PMID- 11253102 TI - [Imaging the smallest tumor vessels using color Doppler ultrasound in an experiment]. AB - PURPOSE: We present an experimental, statistical approach to estimate the size required for small vessels to become detectable with color Doppler sonography. MATERIALS AND METHODS: A murine experimental tumor was examined with color Doppler sonography after injection of 1.5 ml of the contrast medium Levovist. Histologically, we measured vessel diameters inside the tumor as well as other, clearly identifiable locations. RESULTS: With color Doppler at a transmit frequency of 7 MHz, vessels were only detected in the tumor's environment, but not inside. From the 95% quantiles of the vessel diameter distribution found histologically, we estimate that vessels 80-140 microns in diameter or above may be detectable with color Doppler sonography, while vessels 40 microns in diameter or smaller are indetectable. CONCLUSIONS: Although a direct sonographic- histologic correlation is impossible for small vessels, a systematic assessment of the size distribution in clearly identifiable regions permits to estimate the sensitivity of color Doppler to detect blood flow in small vessels. According to our results, capillary blood flow is indetectable, and precapillary vessels may be detected only under optimal conditions. PMID- 11253103 TI - [Magnetic resonance-guided therapy with focused ultrasound. Non-invasive surgery of breast carcinoma?]. AB - Magnetic resonance guided focused ultrasound surgery (MRgFUS) has the potential to become an important therapy modality in the adjuvant, neoadjuvant or palliative cancer treatment. All ultrasound accessible regions are possible target areas, especially breast tumors. Ultrasound propagation is well predictable. The ultrasound energy can be focused to a defined spot through the intact skin, and temperatures of 60 degrees C to 85 degrees C can be induced locally for a few seconds that instantaneously necrose biological tissues, while sparing surrounding healthy tissue. In addition, MRI is sensitive to temperature allowing for online monitoring of the temperature focus. In this work we demonstrate our Heidelberg experiments from basic research and animal studies towards the clinical realization of MRgFUS in breast cancer patients. The most important of these experiments involved sheep as an appropriate model for the human breast. A new therapy setup is designated to treat human breast patients in a clinical 1.5 T MRI scanner. While the therapies have been successful so far without any side effects, the future clinical role of noninvasive MRgFUS has to be defined by clinical studies. PMID- 11253104 TI - [Pathomorphology of laser-induced interstitial tumor thermotherapy of the liver]. AB - The aim of this study was to analyse pathomorphological findings after treatment with laser induced tumor thermotherapy (LITT) on liver tissue and to correlate the results with magnetic resonance imaging. LITT was performed ex vivo and in vivo using a Neodym-YAG-Laser. Lesions were monitored by MR-thermometry ex vivo and by contrast-enhanced MRI in vivo. After LITT the lesions were examined macroscopically, histologically, and electronmicroscopically. LITT-induced tissue damage was qualitatively evaluated, classified, and quantified by means of digital image analysis. Four different zones of tissue damage were identified within the lesions. Adjacent to the applicator the tissue was completely ablated while more peripheral lesions exhibited only sublethal cell damages seen by EM. In vivo the pattern of tissue injury followed the lobular architecture of the liver tissue. Ultrastructural examination revealed only in areas of minor tissue injury intact sinusoidal patterns. MRI overestimated the diameter of the core zone of complete tissue ablation both ex vivo and to a lesser extent in vivo. PMID- 11253105 TI - [Diagnosis of acute lung embolism with spiral CT and 3D reconstruction. Development of an animal model and technical probe in an ex-vivo experiment]. AB - PURPOSE: To develop a model for simulation the CT morphologic situation of acute pulmonary embolism, to evaluate the accuracy of spiral CT and 3D reconstruction in the detection of artificial emboli and to investigate the influence of the orientation of emboli depending on z-axis orientation. MATERIALS AND METHODS: Standardized artificial emboli made of wax and of defined size and shape were positioned into the pulmonary arteries of porcine lungs. Castings of the embolized pulmonary arterial trees were made by injection of a special opaque resin. After performance of spiral CT the data sets of the emboli and the pulmonary arteries were post-processed. The 3D segmentations were compared with the anatomic preparation to evaluate the accuracy of spiral CT/3D reconstruction technique. Technical specimens simulating CT-morphology of acute embolized vessels underwent spiral CT in six different positions with respect to the z axis. The CT data were reconstructed using a standardized and a contrastadapted method with interactive correction. The 3D emboli were analysed under qualitative aspects, and measurements of their extent were done. RESULTS: In nearly 91%, there was complete agreement between CT and the corresponding findings at the anatomical preparation. Measurements of the 3D reconstructed technical specimens showed discrepancies of shape and size in dependence of the size of the original preparation, orientation and reconstruction technique. Overestimation up to 4 mm and underestimation to 2.2 mm were observed. Measurements of preparations with heights from 14 to 26 mm showed variances of +/- 1.5 mm (approximately 6-11%). CONCLUSION: The presented models are suitable to simulate CT morphology of acute pulmonary embolism under ex-vivo conditions. Accuracy in the detection of artificial emboli using spiral CT/3D reconstruction is affected by localization, size and orientation of the emboli and the reconstruction technique. PMID- 11253106 TI - [Multi-rotation CT and acute respiratory distress syndrome. Animal experiment studies]. AB - PURPOSE: Aim of the study was to investigate alveolar inspiration and expiration using multiscan CT. Results of a visual assessment using a scoring system were compared with density ranges known to represent alveolar ventilation best. METHOD: Pigs were examined before and after lavage-induced ARDS. All animals were examined using dynamic multiscan CT. The visual assessment was done by a scoring system proposed by Gattinoni. The results were compared with planimetric determination of defined density ranges. RESULTS: In the healthy lung, the visual analysis showed higher scores at lower airway pressures with a marked gradient, whereas at higher pressures neither opacities nor gradients were observed. In ARDS-lungs, the scores were double as high as in healthy lungs at low pressures. At the same time the differences between inspiration and expiration were minor. There was good correlation between lung density measurements and lung opacities under different airway pressures. In healthy lungs, the greatest area increase is found between -910 and -700 HU. The biggest area growth in the ARDS-model is observed between -910 and -300 HU. CONCLUSION: Dynamic multiscan CT allows for determining different ventilation-relevant lung compartments and lung density ranges. PMID- 11253107 TI - [CT-guided marking of lung lesions before minimally invasive operation]. AB - Video-assisted thoracoscopic surgery (VATS) is an established method for resection of suspicious pulmonary lesions. However, there are problems to detect small subpleural lesions. A procedure for localization of such lesions will be demonstrated. Since may 2000 our experience includes 5 patients (4m, 1f) suffering from solitary pulmonary lesions. In preparation of VATS a CT-guided marking was carried out using both a lasermarker system as well as a special marker system for lung lesions. All 5 procedures were successful. With the laser system the pulmonary nodule was exactly marked and the special wire was placed without any complications. Consequently, the pulmonary nodule was fixed. During video-assisted wedge resection the nodule can be tracted outside. Operating time was reduced in comparison to time consuming search of unmarked lesions. The combined application of CT-guided marking, transthoracic fixation of pulmonary nodule and VATS is recommended preoperatively. It should apply in lesions, which are located subpleural and thoracoscopically not visible. PMID- 11253108 TI - [Prevalence and clinical significance of computerized tomography verified idiopathic calcinosis of the basal ganglia]. AB - With increasing CT examinations of the cerebrum, the discovery of basal ganglia calcification becomes more frequent. In order to correlate these calcifications to the symptoms believed to be accompanied with Fahr's disease 2318 cranial CT scans were examined. There was an overall incidence of basal ganglia calcification of 12.5%. The most frequent location was the globus pallidus (96.4%). In the examined population there was no correlation found between the calcifications and symptoms having been described with striopallidentate calcifications. PMID- 11253109 TI - [Hematemesis of uncertain origin. Cavernous stomach hemangioma]. PMID- 11253110 TI - 21st Annual Conference on Dialysis. New Orleans, Louisiana, USA. February 19-21, 2001. Abstracts. PMID- 11253111 TI - Liver histopathology and biological correlates in five cases of fatal dengue fever in Vietnamese children. AB - We studied five fatal cases of dengue haemorrhagic fever (DHF), confirmed using the reverse transcriptase-polymerase chain reaction (RT-PCR) method, in Vietnamese children. The liver seems to be a target for dengue virus, so postmortem examinations were performed to investigate elementary lesions, local recruitment of inflammatory cells and whether the virus was present in target cells of the liver. We detected severe, diffuse hepatitis with midzonal necrosis and steatosis in two patients, focal areas of necrosis in two patients, and normal histology in one patient. Dengue virus antigen was detected using immunohistochemistry in hepatocytes from necrotic areas in four cases. There was no recruitment of polymorphonuclear cells, and no lymphocytes were detected in the liver lesions of patients who died from DHF. Lymphocytic infiltration occurred in only one hepatitis B virus-positive patient, with no signs of chronic hepatitis. Kupffer cells had mostly been destroyed in cases with focal or severe necrosis. TUNEL tests were positive in necrotic areas, with positive cells forming clusters, suggesting that an apoptotic mechanism was involved. Thus, we suggest that the hepatocyte and Kupffer cells may be target cells supporting virus replication and that the councilman body is an apoptotic cell, as in the pathogenesis of yellow fever. PMID- 11253112 TI - Expression of thrombospondin-1 in pancreatic carcinoma: correlation with microvessel density. AB - Thrombospondin-1 (TSP-1) is a multifunctional platelet and extracellular matrix protein that is involved in angiogenesis. Under certain pathological conditions, e.g., malignant tumors, high concentrations of TSP-1 work as an angiogenic agonist. Here we examined 98 pancreatic carcinomas with respect to TSP-1 immunoreactivity and its correlation to intratumoral microvessel density (MVD), a representation of the overall degree of angiogenesis in carcinomas. Northern blot analysis for TSP-1 mRNA was performed in seven additional cases. Eighty-seven tumors showed strong TSP-1 immunoreactivity, nine carcinomas were only weakly positive, and two lesions were negative for TSP-1. TSP-1 immunoreactivity was detected in the extracellular matrix, mostly at the invasion front of the tumor. Using Northern blot analysis, we observed high levels of TSP-1 mRNA in three out of seven pancreatic carcinomas. The mean MVD in pancreatic carcinoma was 38.8 vessels per mm2. Tumors with a high expression of TSP-1 showed a higher MVD and the correlation between TSP-1 immunoreactivity and microvessel density was highly significant (P=0.003). As a modulator of angiogenesis, TSP-1 is strongly expressed in most pancreatic adenocarcinomas and is likely to contribute to the extensive neovascularization and spread of this highly aggressive tumor. PMID- 11253113 TI - Protein expression of matrix metalloproteinases 2 and 9 and tissue inhibitors of metalloproteinase 1 and 2 in papillary thyroid carcinomas. AB - Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) play an important role in tumor invasion and metastasis. There have been only a few studies on the protein expression of MMPs and TIMPs in thyroid carcinomas. Therefore, we investigated the protein expression of MMP-2, MMP-9, TIMP-1 and TIMP-2 in 86 papillary thyroid carcinomas using immunohistochemistry, semiquantitative scoring morphometry of immunohistochemistry, gelatin zymography, and western blotting. We also examined the correlations between the immunohistochemical scores and several clinicopathological parameters. The immunoreactivities of MMP-2, MMP-9, TIMP-1, and TIMP-2 were largely located in the tumor cells or non-tumor follicular cells and to a much lesser extent in the fibroblasts and endothelial cells in the tumor and non-tumor regions. Compared with non-tumor regions, these four proteins tended to be overexpressed in the tumor cells; the overexpression was found in 64 of 86 (74%), 80 of 86 (93%), 79 of 86 (92%), and 64 of 86 (74%) cases for MMP-2, MMP-9, TIMP-1, and TIMP-2, respectively. Gelatin zymography showed distinct bands of MMP-2 and MMP-9 in tumor extracts but vague bands in non-tumor extracts. Western blotting revealed the specific bands of MMP-2 and MMP-9 in both tumor and non-tumor extracts. Morphometric scoring revealed that high expression of these proteins significantly correlated with large tumor size, presence of lymph node metastasis, high clinical stage, high intrathyroidal invasion, and high vascular invasion. These data suggest that MMP-2, MMP-9, TIMP-1, and TIMP-2 proteins and activities are increased in tumors cells of papillary thyroid carcinomas and that they play an important role in the invasion and metastasis of papillary thyroid carcinomas. PMID- 11253114 TI - On glomerular structural alterations in type-1 diabetes. Companions of early diabetic glomerulopathy. AB - Glomerular structural modifications were measured in kidney biopsies from two follow-up studies in type-1 diabetic patients with microalbuminuria and in kidney donors. Stereologic methods were used to obtain data on glomerular composition and absolute quantities per glomerulus to supplement data on diabetic glomerulopathy previously published. Diabetic patients at baseline (n=37) showed significant changes compared with controls (n=11). The volume fraction of tuft/glomerulus was increased, the proportion of capillary surface facing peripheral basement membrane was decreased (0.72+/-0.04 vs 0.77+/-0.03, P=0.0008), the ratio of mesangial surfaces, urinary/capillary, was decreased (0.67+/-0.17 vs 1.11+/-0.28, P<10(-4)), and the average capillary diameter was increased (8.9+/-0.9 microm vs 7.5+/-1.0 microm, P=0.0002). The total volume of mesangial extracellular material per glomerulus was increased (P=0.01), whereas glomerular volume was not significantly different from controls. Follow-up biopsies after antihypertensive treatment with ACE-inhibitor (n=7) or beta blocker (n=6; 36-48 months) and after intensive insulin treatment (n=7; 24-33 months) showed no change. In a conventionally treated group (n=9), the glomerular volume, the volume of extracellular material/glomerulus, and the capillary length increased. The mean capillary diameter did not correlate with the glomerular volume. In conclusion, the development of diabetic glomerulopathy entails structural modifications of the glomerular tuft. Antihypertensive and intensified insulin treatment seem to slow the progression of ultrastructural changes. PMID- 11253115 TI - Sequence of events in the glomerular capillary wall at the onset of proteinuria in passive Heymann nephritis. AB - Proteinuria in passive Heymann nephritis (PHN) results from complement-mediated glomerular injury, since complement depletion with cobra venom factor (CVF) prevents proteinuria. However, there are no comprehensive morphological studies identifying the sites of injury leading to onset of proteinuria. To address this issue, we attempted to locate sites of injury involved in the onset of proteinuria in PHN. PHN was induced in intact Munich-Wistar rats (PHN-rats, examined at days 3, 5, and 7) and in complement-depleted rats (CVF treated, PHN CVF-rats, examined at days 3 and 5). The distribution of endogenous albumin in the glomerular basement membrane (GBM) was studied in in situ drip-fixed glomeruli using immunogold immunocytochemistry, and glomerular anionic sites were visualized by polyethyleneimine staining. In addition, the ultrastructural localization of an epitope recognized by a proteinuria-inducing monoclonal antibody (called 5-1-6) directed against the slit diaphragm was examined. Significant proteinuria was seen in intact PHN-rats, starting at day 5. The intensity of gold labeling for endogenous albumin was significantly increased at the outermost site of the GBM (GBM interfacing foot process and the filtration slit, designated area O) at day 3 in both PHN-rats and PHN-CVF-rats in comparison to untreated controls. At day 5, labeling for albumin in area O was decreased in PHN-rats, but not in PHN-CVF-rats, where it was then higher; in PHN-rats, some areas between epithelial cells and subepithelial deposits were almost free of albumin labeling at day 7. There was no evidence of epithelial cell detachment in any group at day 5, but on day 7 limited focal detachment was seen exclusively in PHN-rats. In proteinuric rats, amorphous material that stained for albumin could be seen in the urinary space, without any exocytosis of labeling by glomerular epithelial cells. A significant reduction of intensity of staining for anionic sites was seen in parallel in both groups, but only in the regions of the lamina rara externa adjacent to subepithelial deposits. This local loss of charge might contribute to enhanced permeability to albumin in both PHN- and PHN-CVF-rats. Changes in the appearance of the filtration slits and in the density and distribution of antigen recognised by monoclonal antibody 5-1-6 were similar in PHN- and PHN-CVF-rats at day 5. Complement depletion prevented neither the reduction in anionic sites of the GBM nor the changes in the slit diaphragm observed. These data suggest that albumin leakage between the epithelial cell and the GBM (area O) could occur in PHN-rats, perhaps as a result of epithelial foot process changes. This may be the final link in the chain of events responsible for the onset of proteinuria in PHN. PMID- 11253116 TI - Radiation sclerosing proliferative atypical nephropathy of peritumoral tissue of renal-cell carcinomas after the Chernobyl accident in Ukraine. AB - After the Chernobyl accident, the morbidity of renal-cell carcinomas in Ukraine increased gradually from 4.7 to 7.5 per 100,000 of the total population. Cesium 137 (137Cs) is responsible for 80-90% of the internal radioactivity in people living in radiocontaminated areas of Ukraine, and 90% of 137Cs is eliminated through the kidneys. Histological and immunohistochemical study of proliferating cell nuclear antigen (PCNA) and K-ras protein was performed in peritumoral kidney tissues of 167 Ukrainian patients (groups I-III, according to varying degrees of internal exposure to radiation), and of 85 analog Spanish patients, as a control group. Our data showed in the majority of Ukrainian patients a radiation sclerosing proliferative atypical nephropathy (RSPAN) in association with an increase in the incidences of tubular epithelial nuclear atypia and carcinoma in situ (CIS). Areas of epithelial nuclear atypia and CIS of the cortex and medulla showed significant PCNA expression with means of extent as 12, 14, and 15% of stained nuclei in groups I, II, and III respectively. K-ras expression of the same areas occurred in 67, 87, and 85% of cases in groups I, II, and III respectively. The present study points to a strong relationship between the long term of low-dose radiation exposure of the Ukrainian population and the development of RSPAN as a possible precursor of malignancy. In addition, it confirms the possible initiator, promoter, or progressor role of chronic low level radiation of renal human carcinogenesis in Ukraine. PMID- 11253117 TI - Successful xenografting of cryopreserved primary pancreatic cancers. AB - In order to assess the suitability of cryopreserved neoplastic tissues for xenografting into nude (nu/nu) mice, we compared the take rate in 28 samples of pancreatic ductal carcinoma. Eleven fresh samples were implanted in nu/nu mice, and 17 were frozen in cryopreserving solution and implanted at a later time. All samples were examined for the presence of neoplastic tissue in cryostat sections. A total of 15 tumors grew in the animals; five from the freshly implanted samples and ten from those cryopreserved. Ten xenografted tumors were characterized for alterations in p53, K-ras, and p16 genes, which were found in six, eight, and nine cases, respectively. Our results demonstrate that the take rate for xenografting is comparable between cryopreserved and fresh tissue samples. The procedure allows for the exchange of tumor material between institutions and permits the establishment of centralized facilities for the storage of an array of different primary tumor samples suitable for the production of in vivo models of cancers. PMID- 11253118 TI - Primary malignant melanoma of the gallbladder in dysplastic naevus syndrome. AB - A case of gallbladder involvement by malignant melanoma in a 57-year-old woman is reported. The gallbladder, resected for cholelithiasis, harboured a pedunculated polypoid dark mass, which histologically revealed sheets and nests of epithelioid cells with hyperchromatic nuclei in the lamina propria and at the junctional level. These cells were pigmented (with positive reaction with Schmorl's stain and bleaching with peroxide) and showed immunohistochemical positivity for S-100, gp 100 antigen (HMB-45 antibody) and vimentin. The patient, affected by dysplastic naevus syndrome, had a melanoma in situ excised from the scalp 8 years earlier. The features of the investigated lesion address towards a diagnosis of primary gallbladder melanoma. Furthermore, this is the first time that the existence of such a controversial entity is sustained by the ultrastructural investigation of melanosomes, demonstrating the presence of two melanocitary populations, a typical one exclusively junctional and an atypical one both at the junctional level and in the lamina propria. PMID- 11253119 TI - A distinctive melanocytic lesion associated with melanoma-prone dysplastic naevus syndrome: the hybrid naevus. AB - Clinically and histologically, the concept of dysplastic nevi remains controversial. To elaborate more precise criteria for the nevi of patients with dysplastic naevus syndrome (DNS), we examined 58 nevi from seven DNS patients who developed one or several malignant melanomas. Clinical presentation and histomorphology were evaluated, and immunohistochemistry was performed using proliferation marker Ki-S5 and antibody DO-7 to the p53 protein. Sixty nevi from individuals without history of melanoma served as controls. Of the DNS nevi, 21 (36.2%) exhibited no morphological particularities. The remaining 37 nevi presented distinctive histological features consisting of a slight epidermal acanthosis, spitzoid vertically oriented nests of dyscohesive nevus cells, and single-standing atypical melanocytes in the basal cell layer of the epidermis. Immunohistochemical analysis revealed an average proliferation index of 2.5%, which significantly surpassed the mean growth fraction of conventional dysplastic nevi (<1%). No increase in p53 expression was observed. Characteristically, active proliferation was found in junctional single-standing melanocytes with or without nuclear atypia rather than in nest-shaped compounds. In conclusion, certain moles of patients with DNS possess distinctive features. The newly characterized criteria may provide a basis for the diagnosis of DNS and might help to identify patients at increased risk for malignant melanoma by examination of a single biopsy. PMID- 11253120 TI - Cytogenetic abnormalities of alveolar soft-part sarcomas using interphase fluorescent in situ hybridization: trisomy for chromosome 7 and monosomy for chromosomes 8 and 18 seem to be characteristic of the tumor. AB - Four alveolar soft-part sarcomas were investigated by means of standard immunohistochemistry and interphase cytogenetics to further characterize the immunophenotype and proliferative activity of this tumor. The main goal of this study was to explore the chromosomal changes of this rare soft-tissue sarcoma. One epithelial (KLI), three neurogenic [neuron specific enolase (NSE), PGP 9.5, and S100], and five myogenic (desmin, myoglobin, alpha-smooth mnuscle actin, alpha-sarcomeric actin, and MyoD1) markers were used for the immunophenotypical analysis. Proliferative activity was assessed using the Ki67 index. Twelve (peri)centromeric (1, 3, 4, 6, 7, 8, 10, 12, 15, 17, 18, and X) and one telomeric (17q25-qtel.) chromosomal probes were used for interphase cytogenetic analysis. Three of the cases showed cytoplasmic desmin and/or myoglobin, and one showed smooth muscle actin positivity. All of the four tumors had granular, cytoplasmic, possibly nonspecific MyoD1 and sarcomeric actin positivity. Two of the tumors were positive for vimentin, four gave focal and weak staining with neurogenic markers (four of four NSE, one of four S100, and four of four PGP 9.5), but none of them was positive with KLI. Alveolar soft-part sarcomas may show myogenic immunophenotype in a number of cases, which supports myogenic differentiation. Fluorescent in situ hybridization using alpha satellite chromosomal probes revealed significant alterations in all of the cases. Most frequent and repeated numerical changes, which seem to be characteristic of the neoplasm and may play an important part in its pathogenesis and/or progression, were trisomy 7, monosomy 8 and monosomy 18. PMID- 11253121 TI - Brenner tumors but not transitional cell carcinomas of the ovary show urothelial differentiation: immunohistochemical staining of urothelial markers, including cytokeratins and uroplakins. AB - To determine whether Brenner tumors and transitional cell carcinomas (TCCs) of the ovary are urothelial in type, the immunoprofiles of 14 Brenner tumors, including three malignant examples, and eight ovarian TCCs were compared with those of Walthard nests, urothelium, 12 urinary bladder TCCs and 17 ovarian adenocarcinomas (serous, endometrioid, mucinous, and undifferentiated type). The immunohistochemical stains used included those for cytokeratins CKs 5/6, CK7, CK8, CK13, and CK20, vimentin, CA125, and the specific urothelial differentiation marker uroplakin III. CK7 and CK8 were broadly expressed in most tumors of ovary and bladder examined, while vimentin was focally present in some ovarian TCCs and adenocarcinomas. As in normal and neoplastic bladder urothelium, urothelial markers, including uroplakin III, CK13, and CK20, were detected in the epithelial nests of Brenner tumors. Brenner tumor cells also expressed uroplakins Ia and II. CA125 was observed focally in some Brenner tumors. In contrast, TCCs of the ovary and Walthard nests lacked uroplakins and were essentially negative for CK20 and CK13 but quite strongly expressed CA125. This immunophenotype closely resembled that found in ovarian adenocarcinomas. Thus, it appears that the only true urothelial-type ovarian neoplasm is the Brenner tumor, whereas ovarian TCC most likely represents a poorly differentiated adenocarcinoma with a morphologic transitional cell pattern. These results may explain the controversies as expressed in the recent literature concerning TCC of the ovary and establish its place among the ovarian adenocarcinomas of mullerian type. PMID- 11253122 TI - Endobronchial juvenile hemangioma--a case report of a neonate including immunohistochemical monitoring and nuclear, cellular, and vascular morphometry. AB - A 3-month-old female child suffered from tachypnea and dyspnea with abnormal blood gas values. Chest X-rays revealed an increased transparency of the left lung and a mediastinal shift to the right side. High resolution computed tomography (CT) documented a narrowing of the left upper stem bronchus. Ensuing endoscopy detected an occlusive endobronchial tumor mass that did not infiltrate the bronchial cartilage as confirmed with endobronchial ultrasonic monitoring. Based on gross histological examination of the surgical specimen obtained using sleeve resection, the highly vascularized tumor exhibited an adenomatoid growth pattern with a rather homogeneous population of nuclei. The light microscopical presentation was consistent with a juvenile (infantile) hemangioma, which was confirmed using immunohistochemical examinations despite the display of neuroendocrine features. Although endobronchial juvenile hemangiomas are an extremely rare event in early childhood, this case underscores the necessity to not neglect its occurrence in differential diagnosis. PMID- 11253123 TI - Meningeal nodules in teratoma of the testis. AB - A case of a 52-year-old man with a mature adult teratoma is reported. Beside histologically mature tissues, this teratoma contained large areas of a meningiomatous proliferation in close proximity of a peripheral nerve and glial tissue. These meningiomatous proliferations were mostly seen in the peripheral parts of the teratoma surrounding the rest of teratomatous elements and were immunohistochemically EMA-positive and S-100 protein- and cytokeratin-negative. Identical meningothelial proliferations are well known in the skin and adjacent soft tissues of the scalp, where they have variously been called sequestrated meningoceles, meningeal hamartoma, cutaneous meningiomas, rudimentary meningocele, hamartoma of the scalp with ectopic meningothelial elements, or cutaneous heterotopic meningeal nodules. PMID- 11253124 TI - Sentinel node biopsy in melanoma. AB - There has, for a long time, been an ongoing discussion on whether the prophylactic removal of lymph nodes ("elective lymph node dissection") is of benefit for melanoma patients. More recently, "selective" lymphadenectomy ("sentinel node biopsy", SNB) has been proposed to evaluate the status of the first draining lymph node ("sentinel node") of the regional basin. Several studies now demonstrate that the sentinel node evaluation for underlying metastatic disease reflects the status of the entire lymph node region and is therefore a useful prognostic factor. A multi-institutional study highlighted SNB status as the most significant prognostic factor, superior to measurement of tumor thickness in primary melanoma. Different techniques to detect micrometastasis within the lymph node are under current evaluation. Histology and immunohistology using antibodies against melanoma-associated antigens are routinely performed in SNs. The clinical value of reverse-transcriptase polymerase chain reaction (RT-PCR)based search for minimal melanoma disease in lymph nodes remains unclear. PMID- 11253125 TI - Application of quantitative techniques for the assessment of gastric atrophy. PMID- 11253126 TI - Papillomavirus research update: highlights of the Barcelona HPV 2000 international papillomavirus conference. AB - The 18th international papillomavirus conference took place in Barcelona, Spain in July 2000. The HPV clinical workshop was jointly organised with the annual meeting of the Spanish Association of Cervical Pathology and Colposcopy. The conference included 615 abstracts describing ongoing research in epidemiology, diagnosis/screening, treatment/prognosis, immunology/human immunodeficiency virus, vaccine development/trials, transformation/progression, replication, transcription/translation, viral protein functions, and viral and host interactions. This leader summarises the highlights presented at the conference (the full text of the abstracts and lectures can be found at www.hpv2000.com). Relevant material in Spanish can be found at www.aepcc.org. PMID- 11253128 TI - Antibodies to extractable nuclear antigens. Has technological drift affected clinical interpretation? AB - Precipitating antibodies to extractable nuclear antigens are important in the diagnosis of connective tissue diseases. Disease associations are defined using gel based techniques. Alternative technologies have been introduced, including passive haemagglutination, enzyme linked immunosorbent assay, and western blotting. This leader contains a review of the literature on the clinical usefulness of these assays, together with knowledge gained from personal experience. Using the example of systemic lupus erythematosus, the sensitivity, specificity, and positive predictive value of the assays for disease is discussed, as is their differences in performance. The conclusion drawn is that disease specificity is method dependent. Validation and audit of performance of the method selected by the investigation laboratory is essential. PMID- 11253129 TI - Diagnosis of severe combined immunodeficiency. AB - Early diagnosis of severe combined immunodeficiency (SCID) is important to enable prompt referral to a supraregional centre for bone marrow transplantation before the occurrence of end organ damage secondary to infective complications. This review outlines clinical, microbiological, and immunopathological clues that aid the diagnosis of SCID and emphasises the multidisciplinary approach needed to diagnose and treat these infants. PMID- 11253130 TI - The usefulness of tyrosinase in the immunohistochemical assessment of melanocytic lesions: a comparison of the novel T311 antibody (anti-tyrosinase) with S-100, HMB45, and A103 (anti-melan-A). AB - AIM: To compare the sensitivity and staining pattern of the new immunohistochemical antibody to tyrosinase (T311) with S-100, HMB45, and the recently evaluated antibody to melan-A (A103) in a range of melanocytic lesions. METHOD: Archival, formalin fixed, paraffin wax embedded sections from 50 benign and malignant melanocytic lesions were stained immunohistochemically with anti tyrosinase, A103, S-100, and HMB45. They were scored semiquantitatively for the distribution and intensity of staining. RESULTS: All melanomas, with the exception of desmoplastic melanoma, showed some staining with all four antibodies. Overall, T311 and A103 showed an intermediate sensitivity compared with that of S-100 and HMB45. T311 stained most benign and malignant lesions strongly and diffusely with minimal background staining. Immunoreactivity was found to be patchy in some naevi, with weak or absent staining of the mature melanocytes. A103 showed strong and diffuse staining of all benign lesions and most melanomas with minimal background staining. S-100 was the most sensitive, with diffuse staining of most lesions, including desmoplastic and metastatic melanoma, but lacked specificity. HMB45 was the least sensitive antibody, frequently demonstrating patchy staining with absent staining in some benign naevi. CONCLUSIONS: S-100 remains the most sensitive marker of melanocytes. However, because of its lack of specificity, it should be used with at least one other more specific antibody. HMB45 is more specific, but lacks sensitivity; T311 is a reliable marker of melanocytes in paraffin wax embedded sections and is worth consideration for use in a staining panel, although it shows no additional benefit over A103. PMID- 11253127 TI - Antioxidants in health and disease. AB - Free radical production occurs continuously in all cells as part of normal cellular function. However, excess free radical production originating from endogenous or exogenous sources might play a role in many diseases. Antioxidants prevent free radical induced tissue damage by preventing the formation of radicals, scavenging them, or by promoting their decomposition. This article reviews the basic chemistry of free radical formation in the body, the consequences of free radical induced tissue damage, and the function of antioxidant defence systems, with particular reference to the development of atherosclerosis. PMID- 11253132 TI - Acute erythremic myelosis (true erythroleukaemia): a variant of AML FAB-M6. AB - AIMS: Classic erythroleukaemia (acute myeloid leukaemia M6, or M6 AML) is defined as an excess of myeloblasts in an erythroid predominant background. Leukaemia variants in which the primitive blast cells are demonstrably erythroid are extremely rare and poorly characterised. Variably referred to as "true erythroleukaemia" or "acute erythremic myelosis", they are often included within the M6 AML category even though they do not meet strict criteria for this type of AML. METHODS: Two cases of acute erythroid neoplasia are presented with clinical, morphological, immunophenotypic, and cytogenetic analysis. RESULTS: Both patients presented with profound anaemia, one in a setting of long standing myelodysplasia. Bone marrow examination revealed a predominant population of highly dysplastic erythroid cells in both cases. In one case, the liver was infiltrated by neoplastic erythroid cells. Both patients died within four months of diagnosis. CONCLUSIONS: This report illustrates that cases of acute leukaemia occur in which the dominant neoplastic cell is a primitive erythroid cell without an accompanying increase in myeloblasts. This does not preclude the neoplastic clone originating in a multipotent haemopoietic stem cell, as suggested by cases arising in patients with myelodysplasia. Acute erythremic myelosis should be recognised as a distinct variant of M6 AML. PMID- 11253131 TI - Downregulation of transforming growth factor beta type II receptor in laryngeal carcinogenesis. AB - AIMS: To investigate whether anomalies of transforming growth factor beta type II receptor (TGF-beta RII) expression occur in the early stages of laryngeal carcinogenesis and to assess their importance in the development of laryngeal squamous cell carcinoma. TGF-beta RII status was examined in laryngeal premalignant lesions coupled with malignant evolution and compared with a control group of similar lesions without progression to cancer. METHODS: Immunohistochemical staining for TGF-beta RII was performed on 15 paraffin wax embedded biopsies from patients with precancerous laryngeal lesions who subsequently developed invasive squamous cell carcinoma of the larynx, and on 30 control biopsies from patients who did not develop cancer in a comparable follow up period. In addition, DNA extracted from 18 preneoplastic lesions and eight squamous cell carcinomas was amplified by the polymerase chain reaction at the poly A and the poly GT regions of the TGF-beta RII gene. RESULTS: In the group of lesions with progression to carcinoma, 11 of 15 cases showed loss (< 20% of epithelial cells) of TGF-beta RII immunoreactivity, whereas among non-evolved lesions only five of 30 had similar altered expression of the receptor (p < 0.001, two tailed Fisher's exact test). All squamous cell carcinomas showed a degree of receptor expression comparable with that of the corresponding preneoplastic lesion, with the exception of one case, in which loss of the receptor was evident only in invasive cancer. Mutation of the poly A sequence of the TGF-beta RII gene was identified in only one precancerous lesion and in the subsequent squamous cell carcinoma. CONCLUSIONS: These findings indicate that the downregulation of TGF-beta RII is an early event in laryngeal carcinogenesis, which may result in the loss of TGF-beta mediated growth inhibition, thereby facilitating the progression of laryngeal precancerous lesions to squamous cell carcinoma. PMID- 11253133 TI - Immunohistochemical study of the expression of MUC5AC and MUC6 in breast carcinomas and adjacent breast tissues. AB - AIM: To study the protein expression patterns of MUC5AC and MUC6 in normal and diseased breast tissues and to compare their expression with that of a mucin (MUC1) normally expressed in mammary tissues. METHODS: Formalin fixed, paraffin wax embedded tissue from 69 cases of invasive breast carcinoma and surrounding breast tissue was studied immunohistochemically with monoclonal antibodies against MUC1 (SM3), MUCSAC (CLH2), and MUC6 (CLH6), using the avidin-biotin peroxidase method. RESULTS: MUC5AC was detected in five of 68 cases of invasive carcinoma including one of three cases of pure colloid carcinoma. MUCSAC expression in the adjacent normal breast epithelium was present in one of 29 cases and in one of two cases of ductal carcinoma in situ. None of 15 cases of ductal hyperplasia without atypia was positive for MUCSAC. MUC6 was present in 15 of 65 cases of invasive carcinoma, in four of 29 cases of normal adjacent epithelium, two of 15 cases of ductal hyperplasia without atypia, and one of two cases of ductal carcinoma in situ. MUC1 immunoreactivity detected by the SM3 antibody was present in 50 of the 67 cases of invasive carcinoma, but expression was also detected in benign epithelium. All invasive carcinomas expressing MUCSAC were positive for MUC1 and four were positive for MUC6. No significant association was found between the expression of these mucins and tumour size, histological grade, node status, oestrogen receptor status, p53 positivity, or c ErbB-2 overexpression. CONCLUSIONS: This study documents the expression of two different mucins (MUCSAC and MUC6) not described as being expressed by normal breast tissues in a minority of breast carcinomas, as well as in normal and hyperplastic epithelium. Although the role of mucins in malignant transformation and the progression of breast cancer is not well understood, in some cases, there is probably an upregulation of several genes that encode distinct mucin proteins. PMID- 11253134 TI - Overwhelming infection in asplenic patients: current best practice preventive measures are not being followed. AB - AIMS: Patients without spleens are at increased risk of overwhelming infection. Recently, greater efforts, including the publication of national guidelines, have been made to improve the management of asplenic individuals. In theory, risks of serious sepsis can be reduced by good advice, immunisation, and antibiotic prophylaxis. In practice, such preventive measures might not be followed or may fail. A study of recent cases of overwhelming postsplenectomy infection (OPSI) was undertaken to examine specific associated factors and to determine whether currently recommended preventive measures are being followed. METHODS: Cases of OPSI were identified and reported mainly by microbiologists across the country using a specifically designed proforma. Data including the nature of the infection and vaccination/ antibiotic prophylaxis history since splenectomy were obtained. RESULTS: Seventy seven cases were reported. The age range varied from 3 months (congenital asplenia) to 87 years. In those who had undergone surgical splenectomy, the time interval between surgery and OPSI varied from 24 days to 65 years. Overall mortality reached 50%, with underlying haematological malignancy associated with the highest death rate. Streptococcus pneumoniae caused approximately 90% episodes. Only 31% individuals had received pneumococcal vaccination before OPSI. Seven of 17 pneumococcal infections in immunised cases could be considered vaccine failures. Few patients had been adequately advised on antibiotic prophylaxis or other measures. CONCLUSIONS: Currently accepted best practice for managing asplenic patients is not being followed. Some OPSI cases may still be preventable but many asplenic individuals remain unrecognised. The compilation of asplenic patient registers might help to implement agreed policies with audit necessary to evaluate compliance. More is needed to ensure optimal management for this cohort of the population. PMID- 11253135 TI - Genetic diversity in the Helicobacter pylori cag pathogenicity island and effect on expression of anti-CagA serum antibody in UK patients with dyspepsia. AB - AIMS: To investigate variation within the cag pathogenicity island (PAI) of Helicobacter pylori isolated from patients with dyspepsia in mid-Essex, and to evaluate the effect on expression of anti-CagA antibody. METHODS: Sixty two isolates of H pylori cultured from gastric biopsies were screened by specific PCR assays for the presence of cagA and other gene markers (cagD and cagE, and virD4) in the cag PAI. An enzyme linked immunosorbent assay (ELISA) kit (Viva Diagnostica helicobacter p120) was used to test for anti-CagA IgG antibody in matching sera. Isolates were also genotyped by vacuolating cytotoxin polymerase chain reaction (PCR) analysis, and tested for absence of the complete cag PAI (empty site PCR assay). RESULTS: Forty one of the H pylori isolates had a cag PAI containing cagA. One strain had no cagA but other cag PAI loci were present, whereas the remaining 20 strains had no detectable cag PAI markers. Anti-CagA IgG antibody was detected in 34 sera by the ELISA assay, and when compared with the cag PAI genotype of the infecting strain, accuracy, sensitivity, and specificity were 92%, 87%, and 100%, respectively. The seven discrepant or borderline strains in the ELISA were all vacA s1 but differed in other genotypic markers. CONCLUSIONS: The cag PAI was widely distributed in H pylori from patients with dyspepsia in mid-Essex who had different gastric pathologies. Infection with a strain having an uninterrupted cag PAI was associated with the presence of anti CagA antibody in most patients. Discrepant ELISA results, mostly for elderly patients with duodenal ulcers, were attributed to cagA associated variation, particularly to the presence of mixed cagA+/cagA- cell variants in the infecting strain population. Tests for anti-CagA serum antibody were unreliable for predicting severity of clinical disease associated with H pylori infection in this series of patients. PMID- 11253138 TI - Macrocryosectioning of the prostate: a simple technique. AB - Whole mount sections of the prostate are widely used in many laboratories. Macrocryosections of the gland; that is, whole mount frozen sections of the prostate from radical prostatectomies represent a useful new research protocol. The technique is very simple and does not require expensive equipment. PMID- 11253137 TI - Expression of cyclins and cyclin dependent kinases in human benign and malignant melanocytic lesions. AB - AIMS: The regulation of cell proliferation is a key event in normal development, pathophysiological responses to injury, and tumorigenesis. The orderly progression of cells through the cell cycle depends on a finely tuned balance between the concentrations of activated cyclins and cyclin dependent kinases. This study was undertaken to compare the expression of cell cycle regulators in benign and malignant melanocytic lesions during tumour progression. METHODS: Immunohistochemistry was used to analyse 49 primary cutaneous malignant melanomas, 18 metastatic melanomas, and 12 histologically confirmed naevus cell naevi for their expression of cyclins (A, B1, D1, D2, D3, and E) and cyclin dependent kinases (CDK1, CDK2, and CDK4). RESULTS: Cyclin E and CDK2 had the highest expression patterns in human cutaneous melanomas and metastases and correlated positively with histological type and tumour stage. Cyclins B1, D2, and D3 had significantly increased expression in metastases, but normal or even decreased expression in primary melanomas. However, cyclins A and D1, and CDK1 and CDK4 were expressed very weakly in situ with no significant differences between naevi, melanomas, or metastases, and there was no correlation with histopathological staging. The specificity of recognition by the antibodies used was confirmed by western blotting on a panel of seven human melanoma cell lines. Cyclins A, B, and E were expressed by all seven, whereas cyclin D1 was detectable in six of seven and CDK2 and cdc2 were present in five of seven lines analysed. CONCLUSIONS: Taken together, this study demonstrated a significant increase of cyclin E and CDK2 expression during tumour progression in malignant melanomas. PMID- 11253136 TI - Expression of killer cell inhibitory receptors is restricted to true NK cell lymphomas and a subset of intestinal enteropathy-type T cell lymphomas with a cytotoxic phenotype. AB - BACKGROUND/AIMS: Killer inhibitory receptors (KIR) have a modulating effect on the cytotoxic functions of natural killer (NK) cells and T cells. Because lymphoma cells often have the same receptors as their non-neoplastic counterparts, this study investigated the expression of KIR on well defined groups of NK and T cell lymphomas, with and without a cytotoxic phenotype, from different sites of origin. METHODS: Nine CD56+/CD3- NK cell lymphomas, 29 CD3+/CD56- T cell lymphomas with a cytotoxic phenotype, and 19 T cell lymphomas without a cytotoxic phenotype were stained for KIR using monoclonal antibodies specific for CD94, CD158a, and CD158b. In addition, the expression of KIR was studied on normal lymphoid tissues. RESULTS: KIR expression was seen in five of nine true NK cell lymphomas including three of four nasal, one of four cutaneous, and one of one intestinal lymphoma nasal type. Double staining for CD56 and CD94 in normal lymphoid tissues revealed that KIR was predominantly expressed by CD56+ NK cells and sporadically on CD8+ T cells. Moreover, enteropathy-type T cell lymphomas with a cytotoxic phenotype showed KIR expression (three cases expressing CD94 and one case expressing CD158a). All nodal and extranodal nonintestinal T cell lymphomas with or without a cytotoxic phenotype lacked expression of KIR. CONCLUSIONS: These results show that KIR expression is restricted to CD56+/CD3- true NK cell lymphomas originating from the nose, gut, and skin, as well as in a subset of extranodal T cell lymphomas originating from the small intestine, which possessed a cytotoxic phenotype. Thus, the presence of KIR on NK/T cell lymphomas seems to mimic the distribution of KIR found on NK and T cells in normal lymphoid tissue. PMID- 11253139 TI - Detection of fastidious bacteria in cardiac valves in cases of blood culture negative endocarditis. AB - The diagnosis of blood culture negative endocarditis is still a problem. Fastidious bacteria such as bartonella and coxiella are responsible for cases of blood culture negative endocarditis, the identification of which is mainly based on serological and DNA studies only available in specialised centres. Therefore, a routine technique is needed in surgical pathology laboratories to detect these bacteria in cardiac valve tissue sections. This report describes a staining technique, the Gimenez stain, feasible and sensitive in detecting bartonella and coxiella in two cases of blood culture negative endocarditis. PMID- 11253140 TI - Solid and papillary epithelial neoplasm arising in heterotopic pancreatic tissue of the mesocolon. AB - AIM: Solid and papillary epithelial neoplasm (SPEN) is an uncommon pancreatic tumour. Very rarely it has also been described outside the pancreas, usually arising from heterotopic pancreatic tissue. This report summarises all the published extrapancreatic SPENs and documents the sixth such case arising from heterotopic pancreatic tissue of the transverse mesocolon in a 15 year old girl. METHODS/RESULTS: Histological and immunohistochemical examination revealed typical papillary and solid areas composed of columnar, cuboidal, and round cells, which were focally positive for vimentin, cytokeratin, neurone specific enolase, carcinoembryonic antigen, alpha1-antitrypsin, alpha1-antichymotrypsin, and negative for neuroendocrine markers (neurofilament, PGP 9.5, chromogranin A, synaptophysin, and S100), p53, and oestrogen and progesterone receptors. Electron microscopy showed scant zymogen but no neurosecretory granules. In agreement with the flow cytometric result s of diploidy, comparative genomic hybridisation (CGH) did not reveal loss or gain of genetic material, and the in situ hybridisation analysis of the RB1 and p53 genes revealed no abnormality in the 13q and 17p arms. CONCLUSIONS: Immunohistochemical and electron microscopic data support exocrine differentiation. The CGH and the flow cytometric results suggest a subtle, yet unknown genetic change, rather than a large genetic alteration. RB1 and p53 in situ hybridisation ruled out the role of deletion at these sites in the pathogenesis of SPEN. Interestingly, review of the published and the present heterotopic pancreatic SPENs identified the mesocolon as the most common anatomical site (four of six), despite the very rare occurrence of ectopic pancreatic tissue at this site. PMID- 11253141 TI - What constitutes a histological confirmation of cancer? A survey of terminology interpretation in two English regions. AB - AIMS: To compare interpretation by cancer registries and histopathologists of phrases that might confirm a diagnosis of cancer. METHODS: One hundred and thirty one consultant pathologists were sent a questionnaire containing 37 phrases used in pathology reports, including those indicating cancer and those not. Pathologists were asked to indicate whether each phrase confirmed the disease, ruled it out, or was uncertain, together with a subjective estimate of how frequently they used the phrase. RESULTS: There was a 58% response rate with similar interpretation between regions. There were some differences in frequency of use. At least 50% of respondents considered 12 terms as confirmatory (for nine the lower 95% confidence limit was greater than 66%). CONCLUSIONS: The registry should consider ignoring four of the 13 terms currently regarded as confirmatory. Terminology used in pathology reports should be standardised across registries. Registries and coding departments should use empirical evidence to assess which phrases confirm a diagnosis. PMID- 11253142 TI - Evidence for loss of heterozygosity of 5q in sporadic haemangiomas: are somatic mutations involved in haemangioma formation? AB - BACKGROUND/AIMS: Haemangiomas are common benign tumours of infancy that consist of rapidly proliferating endothelial cells. A locus for an autosomal dominant predisposition to haemangioma has been identified recently on chromosome 5q. This study aimed to investigate loss of heterozygosity on chromosomes 5 and 9 in haemangiomas. METHODS: Sporadic proliferative phase haemangiomas were microdissected. Polymerase chain reaction amplification and analysis of microsatellite markers on chromosomes 5 and 9 was carried out. RESULTS: There was a significant loss of heterozygosity for markers on chromosome 5q in haemangioma tissue, when compared with either markers from chromosome 5p (p < 0.05) or markers from chromosome 9 (p < 0.05). CONCLUSIONS: These results suggest that haemangioma formation might be associated with somatic mutational events, and provides evidence that a locus on 5q is involved in the formation of sporadic haemangiomas. PMID- 11253143 TI - Mature renal teratoma and a synchronous malignant neuroepithelial tumour of the ipsilateral adrenal gland. PMID- 11253144 TI - Random, chance, or hazard? PMID- 11253145 TI - Thrombophilia testing: science or medicine? PMID- 11253146 TI - 2001 AAHPERD (American Alliance for Health, Physical Education, Recreation and Dance) National Convention and Exposition. Cincinnati, Ohio, USA. March 2001. Abstracts. PMID- 11253147 TI - Effects of food restriction on systolic mechanical behavior of the ventricular pump in middle-aged and senescent rats. AB - Previous work from our laboratory has revealed that the intrinsic contractility of the left ventricle is depressed in rats at 24 months, and the ventricular internal resistance shows declines with age. The aim of this study was to determine whether food restriction (FR) delays the development of age-related changes in left ventricular (LV) contractility and internal resistance. Male Fischer 344 rats that began FR at the ages of 12 and 18 months were fed on alternate days for 6 months and compared with age-matched ad libitum (AL)-fed rats. Rats studied at the ages of 18 and 24 months were referred to as middle aged and senescent rats, respectively, and were anesthetized and thoracotomized. We measured LV pressure and ascending aortic flow waves by using a high-fidelity pressure sensor and an electromagnetic flow probe, respectively. The elastance resistance model was used to generate Emax and Qmax to describe the physical properties of the left ventricle; Emax is the maximal systolic elastance to represent the myocardial contractility; Qmax is the theoretical maximal flow to be inversely related to the LV internal resistance. Neither age nor diet affected basal heart rate, LV end-systolic pressure, or cardiac output. Emax normalized to LV weight (Emaxn) exhibited a decline from 941.9+/-62.7 mmHg/ml-g to 690.2+/-57.5 mmHg/ml-g with age in AL-fed rats but not FR rats. Qmax showed an increase with age from 36.55+/-2.78 ml/s to 44.22+/-2.62 ml/s in AL-fed rats or from 36.01+/ 2.09 ml/s to 43.52+/-2.74 ml/s in FR rats. There was no effect of diet on Qmax. In conclusion, FR prevents or delays the reduction in myocardial contractility that occurred between 18 and 24 months of age in AL rats. However, FR does not affect the age-related changes in ventricular internal resistance. PMID- 11253148 TI - Effects of caloric restriction on skeletal muscle mitochondrial proton leak in aging rats. AB - Long-term caloric restriction (CR) retards aging processes and increases maximum life span. We investigated the influence of CR on mitochondrial proton leaks in rat skeletal muscle. Because CR lowers oxidative damage to mitochondrial membrane lipids and proteins, we hypothesized that leak would be lower in mitochondria from old CR rats than in age-matched controls. Three groups (n = 12) were studied: 4-month-old "young" control rats (body weight: 404 g +/- 7 SEM), 33 month-old CR rats (body weight: 262 g +/- 3), and 33-month-old control rats (body weight: 446 g +/- 5). CR rats received 67% of the energy intake of old control rats, with adequate intakes of all essential nutrients. Maximum leak-dependent O2 consumption (State 4) was 23% lower in CR rats than in age-matched controls, whereas protonmotive force values were similar, supporting our hypothesis. The overall kinetics of leak were similar between the two groups of old rats; in the young, kinetics indicated higher protonmotive force values. The latter indication is consistent with aging-induced alterations in proton leak kinetics that are independent of dietary intervention. There was no influence of age or diet on serum T4 level, whereas T3 was lower in young than in old control rats. These results support and extend the oxidative stress hypothesis of aging. PMID- 11253149 TI - Dietary restriction and aging: comparative tests of evolutionary hypotheses. AB - Dietary restriction (DR) increases life span in many types of animals. The response to chronic DR may be an adaptation to environments with variable food levels. This study uses the comparative method to test evolutionary predictions about the origin of the response to DR, using data from 10 species of rotifers. Most species, but not all, responded to DR by increasing mean life span, maximum life span, reproductive life span, mortality rate doubling time, and initial mortality rate. Interspecific comparisons did not show the predicted correlations between the strength of the response to DR and either reproductive life span, age of first reproduction, or total reproduction. There was support for the idea that the response to chronic DR is associated with changes in reproductive allocation during short-term periods of starvation: species that reduced reproduction when starved increased their life spans under DR, whereas species that continued to reproduce when starved decreased their life spans under DR. PMID- 11253150 TI - Familial excess longevity in Utah genealogies. AB - We evaluated the influence of family history on longevity by examining longevity in a cohort of 78,994 individuals drawn from the Utah Population Database (UPDB) who were born between 1870 and 1907, and lived to at least age 65. We examined Mendelian genetic and social modes of transmission of excess longevity (the difference between observed and expected longevity) by varying weighted kinship contributions over different classes of relatives. The genetic component of the variation in excess longevity measured as heritability, h2, was approximately 0.15 (95% confidence interval [CI] 0.12-0.18). Among siblings of probands who reached the 97th percentile of excess longevity (+ 14.8 years, currently age 95 for men and 97 for women), the relative risk of recurrence (lambdas) was 2.30 (95% CI 2.08-2.56). In sibships whose relatives were in the top 15% of the distribution for familial excess longevity, the value of lambdas increased substantially, indicating that considering the longevity of distant relatives may be helpful in the selection of families in which to identify genes influencing aging and longevity. PMID- 11253151 TI - Protective efficacy of L-carnitine on acetylcholinesterase activity in aged rat brain. AB - The purpose of this research was to study the activity of acetylcholinesterase in various regions of young and aged rat brain before and after L-carnitine supplementation. Two groups of male albino rats were used for this study (4 and 24 months of age). L-carnitine was administered intraperitoneally 300 mg/kg/d using physiological saline as a vehicle for 7, 14, and 21 days. The activity of acetylcholinesterase was measured in the cerebral cortex, the hippocampus, the hypothalamus, the striatum and the cerebellum. Highly significant variation was observed in a duration dependent manner in the hippocampus, the striatum, and the cortex of aged rats after L-carnitine supplementation when compared with young controls. Our results indicate that treatment of aged rats with L-carnitine restored the level of acetylcholinesterase. PMID- 11253152 TI - Calorie restriction and skeletal mass in rhesus monkeys (Macaca mulatta): evidence for an effect mediated through changes in body size. AB - Little is known regarding the effects of prolonged calorie restriction (CR) on skeletal health. We investigated long-term (11 years) and short-term (12 months) effects of moderate CR on bone mass and biochemical indices of bone metabolism in male rhesus monkeys across a range of ages. A lower bone mass in long-term CR monkeys was accounted for by adjusting for age and body weight differences. A further analysis indicated that lean mass, but not fat mass, was a strong predictor of bone mass in both CR and control monkeys. No effect of short-term CR on bone mass was observed in older monkeys (mean age, 19 years), although young monkeys (4 years) subjected to short-term CR exhibited slower gains in total body bone density and content than age-matched controls. Neither biochemical markers of bone turnover nor hormonal regulators of bone metabolism were affected by long term CR. Although osteocalcin concentrations were significantly lower in young restricted males after 1 month on 30% CR in the short-term study, they were no longer different from control values by 6 months on 30% CR. PMID- 11253153 TI - Pinning down frailty. PMID- 11253154 TI - The use of COX-2-specific inhibitors: is it all hype or is it evidence based? PMID- 11253155 TI - Treatment of older persons with hypercholesterolemia with and without cardiovascular disease. AB - Hypercholesterolemia is a risk factor for new coronary events in older men and women. Secondary prevention trials have demonstrated in persons with coronary artery disease (CAD) and hypercholesterolemia that statin drugs reduced in older persons all-cause mortality, cardiovascular mortality, coronary events, coronary revascularization, stroke, and intermittent claudication. Statins have also been shown to slow progression of coronary atherosclerotic plaques in persons with CAD, to reduce restenosis after coronary stent implantation, and to decrease myocardial ischemia in persons with CAD. Older men and women with CAD, prior atherothrombotic brain infarction, peripheral arterial disease, or extracranial carotid arterial disease and a serum low-density lipoprotein (LDL) cholesterol level higher than 125 mg/dl despite diet should be treated with statin drug therapy to lower the serum LDL cholesterol level below 100 mg/dl. Primary prevention trials have shown that statins were also effective in reducing cardiovascular events in older persons with hypercholesterolemia. On the basis of data from the Air Force/Texas Coronary Atherosclerosis Prevention Study, the physician should consider using statins in persons aged 65-80 years without cardiovascular disease with a serum LDL cholesterol level above 130 mg/dl and serum high-density lipoprotein cholesterol level below 50 mg/dl. PMID- 11253156 TI - Frailty in older adults: evidence for a phenotype. AB - BACKGROUND: Frailty is considered highly prevalent in old age and to confer high risk for falls, disability, hospitalization, and mortality. Frailty has been considered synonymous with disability, comorbidity, and other characteristics, but it is recognized that it may have a biologic basis and be a distinct clinical syndrome. A standardized definition has not yet been established. METHODS: To develop and operationalize a phenotype of frailty in older adults and assess concurrent and predictive validity, the study used data from the Cardiovascular Health Study. Participants were 5,317 men and women 65 years and older (4,735 from an original cohort recruited in 1989-90 and 582 from an African American cohort recruited in 1992-93). Both cohorts received almost identical baseline evaluations and 7 and 4 years of follow-up, respectively, with annual examinations and surveillance for outcomes including incident disease, hospitalization, falls, disability, and mortality. RESULTS: Frailty was defined as a clinical syndrome in which three or more of the following criteria were present: unintentional weight loss (10 lbs in past year), self-reported exhaustion, weakness (grip strength), slow walking speed, and low physical activity. The overall prevalence of frailty in this community-dwelling population was 6.9%; it increased with age and was greater in women than men. Four-year incidence was 7.2%. Frailty was associated with being African American, having lower education and income, poorer health, and having higher rates of comorbid chronic diseases and disability. There was overlap, but not concordance, in the cooccurrence of frailty, comorbidity, and disability. This frailty phenotype was independently predictive (over 3 years) of incident falls, worsening mobility or ADL disability, hospitalization, and death, with hazard ratios ranging from 1.82 to 4.46, unadjusted, and 1.29-2.24, adjusted for a number of health, disease, and social characteristics predictive of 5-year mortality. Intermediate frailty status, as indicated by the presence of one or two criteria, showed intermediate risk of these outcomes as well as increased risk of becoming frail over 3-4 years of follow-up (odds ratios for incident frailty = 4.51 unadjusted and 2.63 adjusted for covariates, compared to those with no frailty criteria at baseline). CONCLUSIONS: This study provides a potential standardized definition for frailty in community-dwelling older adults and offers concurrent and predictive validity for the definition. It also finds that there is an intermediate stage identifying those at high risk of frailty. Finally, it provides evidence that frailty is not synonymous with either comorbidity or disability, but comorbidity is an etiologic risk factor for, and disability is an outcome of, frailty. This provides a potential basis for clinical assessment for those who are frail or at risk, and for future research to develop interventions for frailty based on a standardized ascertainment of frailty. PMID- 11253157 TI - Associations of subclinical cardiovascular disease with frailty. AB - BACKGROUND: Frail health in old age has been conceptualized as a loss of physiologic reserve associated with loss of lean mass, neuroendocrine dysregulation, and immune dysfunction. Little work has been done to define frailty and describe the underlying pathophysiology. METHODS: Frailty status was defined in participants of the Cardiovascular Health Study (CHS), a cohort of 5,201 community-dwelling older adults, based on the presence of three out of five clinical criteria. The five criteria included self-reported weight loss, low grip strength, low energy, slow gait speed, and low physical activity. We examined the spectrum of clinical and subclinical cardiovascular disease in those who were frail (3/5 criteria) or of intermediate frailty status (1 or 2/5 criteria), compared to those who were not frail (0/5). We hypothesized that the severity of frailty would be related to a higher prevalence of reported cardiovascular disease (CVD), as well as to a greater extent of CVD, measured by noninvasive testing. RESULTS: Of 4,735 eligible participants, 2,289 (48%) were not frail, 299 (6%) were frail, and 2.147 (45%) were of intermediate frailty status. Those who were frail were older (77.2 yrs) compared to those who were not frail (71.5 yrs) or intermediate (73.4 yrs) (p < .001). Frailty status was associated with clinical CVD and most strongly with congestive heart failure (odds ratio [OR] = 7.51 (95% confidence interval [CI] = 4.66-12.12). In those without a history of a CVD event (n = 1.259), frailty was associated with many noninvasive measures of CVD. Those with carotid stenosis >75% (adjusted OR = 3.41), ankle-arm index <0.8 (adjusted OR = 3.17) or 0.8-0.9 (adjusted OR = 2.01), major electrocardiography (ECG) abnormalities (adjusted OR = 1.58), greater left ventricular (LV) mass by echocardiography (adjusted OR = 1.16), and higher degree of infarct-like lesions in the brain (adjusted OR = 1.71), were more likely to be frail compared to those who were not frail. The overall associations of each of these noninvasive measures of CVD with frailty level were significant (all p < .05). CONCLUSIONS: Cardiovascular disease was associated with an increased likelihood of frail health. In those with no history of CVD, the extent of underlying cardiovascular disease measured by carotid ultrasound and ankle-arm index, LV hypertrophy by ECG and echocardiography, was related to frailty. Infarct-like lesions in the brain on magnet resonance imaging were related to frailty as well. PMID- 11253159 TI - Informants' knowledge of reproductive history and estrogen replacement. AB - BACKGROUND: There has been much interest in assessing estrogen use in healthy older women and those with Alzheimer's disease. However, data for the women with Alzheimer's disease must be obtained from an informant. The aim of this study was to better understand what informants are likely to know about reproductive history and estrogen use. METHODS: Reproductive history data from informants of Alzheimer's patients were modeled by comparing responses from 40 cognitively healthy older women with that of a designated informant. The designated informants were similar in demographics to informants for patients with Alzheimer's disease. RESULTS: Informant data regarding reproductive history was likely to be accurate, when known. However, 30% of the subjects did not identify an informant who had personal knowledge of them. Of those informants who had personal knowledge of the subject, accuracy for those who reported that they knew the information varied depending on the aspect of reproductive history assessed (age of menarche, 29%; age of menopause, 20%; pregnancies, 63%; live births, 92%; hysterectomy, 92%; and postmenopausal estrogen use, 82%). Daughters served as the most likely and most accurate informants in this study. CONCLUSION: This study demonstrates that information obtained from informants for patients with Alzheimer's disease is likely to be accurate for some but not all aspects of reproductive history. Of concern for such studies will be the 30% of patients who do not have an informant with personal knowledge about them. PMID- 11253158 TI - Functional status and health-related quality of life of elderly osteoarthritic patients treated with celecoxib. AB - BACKGROUND: This study evaluates the impact of celecoxib on functional status, health-related quality of life (HRQOL), and safety of elderly patients (> or =70 years) with osteoarthritis (OA) of the knee and/or hip. METHODS: Data were pooled from three prospective, randomized, multicenter, double-blind, parallel group trials, each having a 12-week treatment period. Multicenter studies were conducted in the United States and Canada. Data for patients diagnosed with active OA of the knee and/or hip in a flare state who were 70 years of age and older were included in the comparison of therapeutic doses of celecoxib or naproxen versus placebo (N = 768). Elderly patients from each of the three trials who were randomly assigned to groups treated with a placebo. 200 mg/day of celecoxib, 400 mg/ day of celecoxib, or 1000 mg/day of naproxen were included in this analysis. The Western Ontario and McMaster Universities Osteoarthritis Index was used to measure functional status. The Short Form-36 was used as a general measure of HRQOL. Safety was assessed according to the incidence and type of adverse reactions as reported by the patients and the rate of withdrawal due to adverse events. RESULTS: At the end of the treatment period, patients in the celecoxib groups had significant improvement in both functional status and HRQOL in comparison with the placebo group. The effects of total daily doses of 200 mg of celecoxib, 400 mg of celecoxib, and 1000 mg of naproxen on functioning and HRQOL were not found to be significantly different from each other. The incidence of serious adverse events and withdrawal from the studies due to adverse events were similar in the celecoxib groups as they were in the placebo group. Overall, the naproxen group reported a significantly higher incidence of gastrointestinal adverse events than did the placebo and the 200-mg-daily celecoxib groups. CONCLUSIONS: This study showed that celecoxib and naproxen significantly improved functional status and HRQOL in elderly patients compared with those treated with a placebo. Celecoxib-treated patients were also found to experience safety and tolerability similar to that of the placebo-treated patients. PMID- 11253160 TI - Age-associated differences in responses to noxious stimuli. AB - BACKGROUND: Although population-based studies typically report age-associated increases in clinical pain, laboratory-based pain assessment procedures generally indicate diminished pain sensitivity with age. The majority of these studies have utilized noxious thermal stimuli as the method of pain induction. However, other pain assessment methodologies, including ischemic pain induction, may have a more meaningful relationship to clinical pain. The present study examined the effects of age on responses to a variety of experimental noxious stimuli. In addition, relationships between cardiovascular measures and pain responses were investigated in both older and younger subjects. METHODS: Responses to thermal, mechanical, and ischemic pain were assessed in 34 younger (mean age, 22.4 years) and 34 older adults (mean age, 62.2 years). In addition, relationships between resting blood pressure and pain responses were assessed separately for older and younger participants. RESULTS: Although group differences in thermal and mechanical pain responses did not achieve statistical significance, older individuals demonstrated substantially lower ischemic pain thresholds and tolerances assessed via the modified submaximal effort tourniquet procedure (ps < .01). Overall, higher resting arterial blood pressures were associated with increased pain thresholds and tolerances, although relationships between blood pressure and ischemic pain variables were evident only for the younger group. CONCLUSIONS: These findings indicate that age-related differences in responses to experimental noxious stimuli vary as a function of the pain induction task, with older individuals showing greater sensitivity to clinically relevant stimuli. In addition, the absence of a relationship between blood pressure and ischemic pain responses in older adults may suggest potential functional decrements in at least one endogenous pain-modulatory system. PMID- 11253161 TI - Effects of policosanol in older patients with type II hypercholesterolemia and high coronary risk. AB - BACKGROUND: The present study was undertaken to investigate the effects of policosanol in older patients with type II hypercholesterolemia and more than one concomitant atherosclerotic risk factor. METHODS: After 6 weeks on a lipid lowering diet, 179 patients randomly received a placebo or policosanol at doses of 5 followed by 10 mg per day for successive 12-week periods of each dose. Policosanol (5 and 10 mg/d) significantly (p < .001) reduced low-density lipoprotein cholesterol (LDL-C; 16.9% and 24.4%, respectively) and total cholesterol (TC; 12.8% and 16.2%, respectively), while significantly (p < .01) increasing (p < .001) high-density lipoprotein cholesterol (HDL-C) by 14.6% and 29.1%, respectively. RESULTS: Policosanol significantly decreased (p < .01) the ratios of LDL-C to HDL-C (29.1%) and TC to HDL-C (28%) at study completion, although triglycerides remained unchanged. Policosanol, but not the placebo, significantly improved (p .01) cardiovascular capacity, which was assessed using the Specific Activity Scale. No serious adverse experiences occurred in policosanol patients (p < .01), compared with seven adverse experiences (7.9%) reported by placebo patients. CONCLUSIONS: This study shows that policosanol is effective, safe, and well tolerated in older hypercholesterolemic patients. PMID- 11253163 TI - Diagnostic ultrasound induces change within numbers of cryptal mitotic and apoptotic cells in small intestine. AB - Recent work on gas filled organs, including the lung and small intestine, has concentrated on the hemorrhaging effects of ultrasound, with little attention paid to cell cycle perturbations and apoptosis--two very sensitive indicators of environmental insult. This study addresses this by exploring the effects of ultrasound on these two features. The anterior abdominal surface of anaesthetised male, adult CD1 mice was shaved and exposed to ultrasound. An 8 MHz linear array transducer was manually swept from the midline to the left mouse flank on a continuous basis. Each mouse was scanned for 15 minutes with B mode and color flow modes selected. The Thermal Index registered 1.0. Groups of mice were killed at various times after treatment, the small intestine was excised and histologically examined. Analysis of the data demonstrates a statistically significant 22% reduction in numbers of mitotic figures at 4.5 hours after the ultrasonic insult (p = 0.011). Numbers of apoptotic bodies increased by 153% (p=0.003), 166% (p=0.014) and 160% (0.001) at 1, 3 and 4.5 hours post-treatment respectively. These preliminary results suggest that bioeffects of ultrasound maybe more diverse than previously described. Further work will establish thresholds and explore mechanisms for these deterministic effects. PMID- 11253162 TI - Characterization of RGS5 in regulation of G protein-coupled receptor signaling. AB - RGS proteins (regulators of G protein signaling) serve as GTPase-activating proteins (GAPs) for G alpha subunits and negatively regulate G protein-coupled receptor signaling. In this study, we characterized biochemical properties of RGS5 and its N terminal (1-33)-deleted mutant (deltaN-RGS5). RGS5 bound to G alpha(i1), G alpha(i2), G alpha(i3), G alpha(o) and G alpha(q) but not to G alpha(s) and G alpha13 in the presence of GDP/AIF4-, and accelerated the catalytic rate of GTP hydrolysis of G alpha(i3) subunit. When expressed in 293T cells stably expressing angiotensin (Ang) AT1a receptors (AT1a-293T cells), RGS5 suppressed Ang II- and endothelin (ET)-1-induced intracellular Ca2+ transients. The effect of RGS5 was concentration-dependent, and the slope of the concentration-response relationship showed that a 10-fold increase in amounts of RGS5 induced about 20-25% reduction of the Ca2+ signaling. Furthermore, a comparison study of three sets of 293T cells with different expression levels of AT1a receptors showed that RGS5 inhibited Ang II-induced responses more effectively in 293T cells with the lower density of AT1a receptors, suggesting that the degree of inhibition by RGS proteins reflects the ratio of amounts of RGS proteins to those of activated G alpha subunits after receptor stimulation by agonists. When expressed in AT1a-293T cells, deltaN-RGS5 was localized almost exclusively in the cytosolic fraction, and exerted the inhibitory effects as potently as RGS5 which was present in both membrane and cytosolic fractions. Studies on relationship between subcellular localization and inhibitory effects of RGS5 and deltaN-RGS5 revealed that the N terminal (1-33) of RGS5 plays a role in targeting this protein to membranes, and that the N terminal region of RGS5 is not essential for exerting activities. PMID- 11253164 TI - Peroxynitrite inhibits the activity of ornithine decarboxylase. AB - Polyamines are required during cell proliferation, whereas NO has anti proliferative properties. Ornithine decarboxylase (ODC) is a critical enzyme for the synthesis of polyamines. We tested the hypothesis that the modification of ODC by peroxynitrite (OONO-), a short-lived free radical formed from NO and superoxide produces a fall in ODC activity, and therefore polyamine synthesis and cell proliferation. The treatment of a rat recombinant ODC (rODC) with OONO- resulted in a dose-dependent inhibition of rODC activity with an IC50 of approximately 100 microM. A Western blot employing a specific antibody to nitrotyrosine revealed a dose-dependent nitration of rODC tyrosine residues. When intact IEC-6 cells were treated with ONOO-, ODC activity decreased by 49%. These data suggest a correlation between ODC activity and nitration, and a possible mechanism by which NO synthesis may modulate polyamine synthesis. PMID- 11253165 TI - Arginine vasopressin inhibits apoptosis of rat glomerular mesangial cells via V1a receptors. AB - Arginine vasopressin (AVP) promotes proliferation of glomerular mesangial cells. We examined whether AVP modulates an apoptosis of cultured rat glomerular mesangial cells at 3-17th passages. The agarose gel electrophoresis demonstrated that AVP attenuated a ladder formation stimulated by the serum deprivation. The quantitation of oligonucleosomes by ELISA also showed that AVP suppressed the serum deprivation-induced apoptosis. Such an antiapoptotic effect of AVP was dose dependent. An AVP V1a receptor antagonist, d(CH2)5Tyr(Me)AVP, abolished the antiapoptotic effect of AVP. The inhibitory effect of AVP on the apoptosis was reduced by staurosporine and mimicked by phorbol-12-myristate-13-acetate. These results suggest that AVP inhibits serum deprivation-induced apoptosis of glomerular mesangial cells via V1a receptor-protein kinase C pathway. PMID- 11253166 TI - High cholesterol diet down regulates the activity of activator protein-1 but not nuclear factor-kappa B in rabbit brain. AB - Cardiovascular risk factors and alterations in cholesterol metabolism are implicated in the pathogenesis of Alzheimer's dementia (AD). The hypercholesterolemic rabbit model of atheroslerosis and AD was utilized in this study to examine oxidative stress related changes in the brain. The high cholesterol diet induced dramatic increases in plasma and liver cholesterol concentrations, but brain cholesterol levels remained constant. Similar effects have been found regarding lipid oxidation products. The amounts of conjugated dienes, trienes and thiobarbituric acid reactive substances (TBARS) significantly increased in the plasma of cholesterol treated animals while the brain cortex showed no signs of increased lipid peroxidation. The oxidative damage sensitive nuclear transcription factor kappa B (NF-kappaB) and activator protein-1 (AP-1) diverged in their responses. Accordingly, the AP-1 DNA binding activity decreased by more than 50% in brain nuclear protein extracts while the NF-kappaB binding activity remained unaltered by the hypercholesterol diet. These results indicate that despite the relative resistance of the central nervous system to dietary manipulation of its lipid composition and lipid peroxidation products, chronic dietary intake of cholesterol can alter the function of certain proteins involved in regulation of gene expression in the brain. PMID- 11253167 TI - Exercise and ovarian steroid hormones: their effects on mitochondrial respiration. AB - The effect of exercise on mitochondria respiration was studied in gastrocnemius muscle of ovariectomized rats, pseudopregnant rats, and estrous rats. The estrous cycles were followed by vaginal smears. Rats were made pseudopregnant (PSP) by 45 s cervical stimulation with a glass rod on the day of estrous. The treadmill protocol (21 m/min, 10 grade uphill) induced a significant decrease in state 3 oxygen consumption (oxidative phosphorylation) in estrous (0.26 +/- 0.02 vs. 0.49 +/- 0.05 microatoms O min(-1) mg protein(-1)) and ovariectomized rats (0.18 +/- 0.03 vs. 0.40 +/- 0.03 microatoms O min(-1) mg protein(-1)). In contrast, pseudopregnant and progesterone-treated ovariectomized rats did not decrease state 3 nor state 4 respiratory rates. These results show that the effect of exercise on mitochondria respiration does vary according to the hormonal status. PMID- 11253168 TI - Involvement of 1,2-diacylglycerol in improvement of heart function by etomoxir in diabetic rats. AB - Abnormal lipid metabolism has been proposed to be involved in the pathogenesis of diabetic cardiomyopathy. In this study, we measured myocardial lipid levels, including 1,2-diacylglycerol (1,2-DAG) and ceramide (CM), and myocardial function in diabetic rats. We also evaluated the effects of etomoxir (ETM), a carnitine palmitoyl transferase I inhibitor, on diabetic rat hearts from the viewpoints of alterations in lipid second messengers and myocardial function. Rats were injected with streptozotocin (60 mg/kg) to induce diabetes and were treated 5 weeks later with ETM (18 mg/kg) for 8 days. In diabetic rats, heart rate, systolic blood pressure, and fractional shortening were significantly reduced compared with those in controls. Treatment of diabetic rats with ETM ameliorated myocardial dysfunction other than heart rate. Myocardial 1,2-DAG levels in diabetic rats were significantly elevated compared with those in controls, while myocardial CM levels were not. ETM treatment caused an additional increase in myocardial 1,2-DAG levels in diabetic rats, but the CM levels did not change. There was a marked difference in fatty acid pattern of 1,2-DAG between diabetic and ETM-treated diabetic rat hearts. The fatty acids 18:1 and 18:2 were significantly increased and the fatty acids 16:0, 18:0, 20:4, and 22:6 were significantly reduced in ETM-treated diabetic rat hearts. These data suggest 1,2 DAG is involved in ameliorating myocardial dysfunction in diabetic rats and that its source is different between diabetic and ETM-treated diabetic rats. CM is unlikely to be involved in the pathogenesis of diabetic cardiomyopathy or the improvement of cardiac contractility in diabetic rats by ETM. PMID- 11253169 TI - Mechanisms involved in the cardiovascular alterations immediately after spinal cord injury. AB - The early cardiovascular effects resulting from an acute spinal cord injury (SCI) produced by a contusion procedure at T5-T6 were evaluated in anaesthetized rats. The mean arterial pressure (MAP) and heart rate (HR) were measured during one hour after the injury. A marked decrease in MAP and HR was observed immediately after injury, followed by an abrupt increase in MAP. These changes were observed between 3 and 9 min and the basal values were recovered after 20 min. Fall in the MAP and HR and increase in MAP induced by SCI were abolished by atropine. The interruption of the parasympathetic outflow by vagotomy also significantly diminished the fall and increase in MAP and the fall in HR. Likewise, pre treatment with nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) completely abolished the effects produced by SCI. These data suggest that after SCI the decrement in MAP and HR was probably due to acetylcholine release from parasympathetic fibers and NO from endothelial source probably by a cholinergic stimulation. Additionally, the MAP increase observed was probably due to a reflex compensatory vasoconstriction. PMID- 11253171 TI - Profile of acetylcholinesterase in brain areas of male and female rats of adult and old age. AB - Inhibition of acetylcholinesterase (AChE)-metabolizing enzyme of acetylcholine, is presently the most important therapeutic target for development of cognitive enhancers. However, AChE activity in brain has not been properly evaluated on the basis of age and sex. In the present study, AChE activity was investigated in different brain areas in male and female Sprague-Dawley rats of adult (3 months) and old (18-22 months) age. AChE was assayed spectrophotometrically by modified Ellman's method. Specific activity (micromoles/min/mg of protein) of AChE was assayed in salt soluble (SS) and detergent soluble (DS) fractions of various brain areas, which consists of predominantly G1 and G4 molecular isoforms of AChE respectively. The old male rats showed a decrease (40-55%) in AChE activity in frontal cortex, striatum, hypothalamus and pons in DS fraction and there was no change in SS fraction in comparison to adult rats. In the old female rats the activity was decreased (25-40%) in frontal cortex, cerebral cortex, striatum, thalamus, cerebellum and medulla in DS fraction whereas in SS fraction the activity was decreased only in hypothalamus as compared to adult. On comparing with old male rats, old female rats showed increase in AChE activity in cerebral cortex, hippocampus and hypothalamus of DS fraction and decrease in hypothalamus of SS fraction. There was a significant increase in AChE activity in DS fraction of cerebral cortex, hippocampus, hypothalamus, thalamus and cerebellum in female as compared to male adult rats. However, no significant change in AChE activity was found in the SS fraction, except hypothalamus between these groups. Thus it appears that age alters AChE activity in different brain regions predominantly in DS fraction (G4 isoform) that may vary in male and female. These observations have significant relevance to age related cognitive deficits and its pharmacotherapy. PMID- 11253170 TI - The influence of NO-containing ruthenium complexes on mouse hippocampal evoked potentials in vitro. AB - The influence of different, nitric oxide-containing ruthenium complexes on the evoked potentials recorded from the CA1 region of the mouse hippocampus in vitro has been investigated. Of the compounds tested, only trans [(NO)(P(OEt)3)(NH3)4Ru](PF6)3 (1-2.5 mM) exerted a strong facilitatory action on the population spike, the EPSP, and the spontaneous activity. Its activity probably depends upon its ability to release NO following reduction. The phosphito ligand is important both in terms of adjusting the reduction potential of the complex to be biologically accessible and in labilizing the coordinated NO. The effects of this compound could not be reversed by perfusion. Scavenging NO in slices preincubated with oxyhemoglobin prior to the addition of this compound eliminated its neurophysiological effects. The control molecules trans [(P(OEt)3)2(NH3)4Ru](PF6)2, trans-[(H2O)(P(OEt)3) (NH3)4Ru](PF6)3, and [(NO)(NH3)5Ru]Cl3, which are structurally similar, but unable to generate NO, were ineffective. NaNO2 suppressed neuronal firing. Attempts to induce Long-Term Potentiation (LTP) at the time of maximal effect of trans [(NO)(P(OEt)3)(NH3)4Ru](PF6)3 were unsuccessful, suggesting that the mechanism of amplification induced by trans-[(NO)(P(OEt)3)(NH3)4Ru](PF6)3 and LTP may share common pathways. PMID- 11253172 TI - Pregnancy and labor increase the capacity of human myometrial cells to secrete parathyroid hormone-related protein. AB - Parathyroid hormone-related protein (PTHrP), a oncofetal gene product possessing smooth muscle relaxant properties, has been found in rat and human uterine smooth muscle cells (USMC) where it is postulated to regulate myometrial tone and/or blood flow. Studies investigating the gestational regulation of PTHrP in human USMC have not been performed. This study was conducted to determine if pregnancy alters the capacity of USMC to secrete or respond to PTHrP. USMC cultures were established from 8 hysterectomy specimens (H) and 7 non-laboring (NP) and 5 laboring term pregnant uterine biopsies (LP). PTHrP secretion was measured at baseline and in response to TGF-beta1 using a immunoradiometric assay. The USMC response to PTHrP was assessed by incubating cultures with human (1-34)PTHrP and measuring cellular cAMP by radioimmunoassay. We found that cultures from the groups did not differ with respect to basal PTHrP secretion. TGF-beta1, on the other hand, produced dose-dependent increases in secreted PTHrP in each group such that LP>NP>H at 12 hrs and LP>NP and H 24 hrs. Maximal responses were found at 24 hrs in cells treated with 10 ng/ml TGF-beta1 (LP: 2034+/-366 vs NP: 1485+/ 427; H: 1250+/-202 fmol/mg). Incubation of cultures with PTHrP produced dose dependent increases in cAMP production, with 10(-7) M increasing levels by 64%. Neither pregnancy nor labor significantly affected the cAMP response. These findings indicate that the human myometrium has the capacity to increase PTHrP secretion during pregnancy and labor through a TGF-beta-dependent pathway. Such findings are consistent with a role of PTHrP in enhancing uterine blood flow. PMID- 11253174 TI - European science. Flagship E.U. research program aims for pan-European panacea. PMID- 11253175 TI - Foot-and-mouth disease. U.K. outbreak is latest in global epidemic. PMID- 11253173 TI - NMDA recepter-mediated neuroprotection by essential oils from the rhizomes of Acorus gramineus. AB - Acori graminei Rhizoma (AGR) is shown to exhibit a number of pharmacological actions including sedation and anticonvulsive action. To further characterize its actions in the CNS, the present study evaluated the effects of essential oils (EO) from AGR on the excitotoxic neuronal cell death induced in primary rat cortical cell cultures. EO inhibited the glutamate-induced excitotoxicity in a concentration-dependent manner, with the IC50 of 0.241 mg/ml. EO exerted more potent neuroprotection against the toxicity induced by NMDA (IC50 = 0.139 mg/ml). In contrast, the AMPA-induced toxicity was not inhibited by EO. Receptor-ligand binding studies were performed to investigate the neuroprotective action mechanism. EO dramatically inhibited the specific bindings of a use-dependent NMDA receptorion channel blocker [3H]MK-801, indicating an NMDA receptor antagonist-like action. However, the bindings of [3H]MDL 105,519, a ligand selective for the glycine binding site of NMDA receptor, were not considerably inhibited. These results demonstrated that EO extracted from AGR exhibited neuroprotective effects on cultured cortical neurons through the blockade of NMDA receptor activity, and that the glycine binding site appeared not to be the major site of action. PMID- 11253176 TI - 2002 budget. NIH gets big boost; lobbyists want more. PMID- 11253177 TI - Research ethics. Query by Congress halts new policy. PMID- 11253178 TI - Planetary science. Cosmic misfits elude star-formation theories. PMID- 11253179 TI - China. Two honored, other prizes go unclaimed. PMID- 11253180 TI - India. Work starts on first science satellite. PMID- 11253181 TI - Microbiology. Possible new route to polyketide synthesis. PMID- 11253182 TI - Cell biology. Nobel laureates lobby for stem cells. PMID- 11253184 TI - Brazil. New industry taxes boost science budget. PMID- 11253183 TI - Neurobiology. Working memory helps the mind focus. PMID- 11253185 TI - Virology. Sublimating egos for a common goal. PMID- 11253186 TI - AAAS meeting. Lake Vostok: stirred, not shaken. PMID- 11253187 TI - AAAS meeting. Stem cells make brain cells. PMID- 11253188 TI - AAAS meeting. The melting snows of Kilimanjaro. PMID- 11253190 TI - AAAS meeting. Fighting diplomatic technophobia. PMID- 11253189 TI - AAAS meeting. The flip side of obesity research. PMID- 11253191 TI - Hemophilia. After a setback, gene therapy progresses...gingerly. PMID- 11253192 TI - Essays on science and society. Monkeys in the back garden. PMID- 11253193 TI - Phylogenetics. Which mammalian supertree to bark up? PMID- 11253194 TI - Solid state organic chemistry. Stepping it up. PMID- 11253195 TI - Development. The art of making a joint. PMID- 11253196 TI - Geophysics. When the compass stopped reversing its poles. PMID- 11253197 TI - Cosmology. Probing the early universe with the SZ effect. PMID- 11253198 TI - Cell cycle. Some importin news about spindle assembly. PMID- 11253200 TI - U.S. census 2000: an update. PMID- 11253199 TI - Evolution. Nota bene. Wolbachia and wasp evolution. PMID- 11253201 TI - U.S. science policy. New faces please and puzzle researchers. PMID- 11253202 TI - Solar, planetary studies. Two fields prepare to take the long view. PMID- 11253203 TI - Human genome. NIH considers paying to use private database. PMID- 11253204 TI - Paleontology. New fossil fills gap in bird evolution. PMID- 11253205 TI - Transgenic animals. Infant monkey carries jellyfish gene. PMID- 11253206 TI - Circadian rhythms. Mutant gene speeds up the human clock. PMID- 11253207 TI - Evolution of sockeye salmon ecotypes. PMID- 11253208 TI - Selfish DNA and the origin of genes. PMID- 11253209 TI - Scientific whaling. PMID- 11253210 TI - Tenured faculty on soft money. PMID- 11253211 TI - No error in vote counts--in principle. PMID- 11253212 TI - No error in vote counts--in principle. PMID- 11253213 TI - Disasters. Volcano fatalities--lessons from the historical record. PMID- 11253214 TI - Particle physics. The search for the Higgs boson. PMID- 11253215 TI - Neuroscience. Seeking categories in the brain. PMID- 11253216 TI - Molecular metals. Staying neutral for a change. PMID- 11253217 TI - Ecology. Bird navigation--computing orthodromes. PMID- 11253218 TI - Electron spin resonance in studies of membranes and proteins. AB - We provide a review of current electron spin resonance (ESR) techniques for studying basic molecular mechanisms in membranes and proteins by using nitroxide spin labels. In particular, nitroxide spin label studies with high-field/high frequency ESR and two-dimensional Fourier transform ESR enable one to accurately determine distances in biomolecules, unravel the details of the complex dynamics in proteins, characterize the dynamic structure of membrane domains, and discriminate between bulk lipids and boundary lipids that coat transmembrane peptides or proteins; these studies can also provide time resolution to studies of functional dynamics of proteins. We illustrate these capabilities with recent examples. PMID- 11253219 TI - [The 42nd Congress of the Japanese Society of Clinical Hematology. Kurashiki City, Japan. November 8-10, 2000. Abstracts]. PMID- 11253220 TI - [The 47th Meeting of the Japanese Society of Clinical Pathology. Fukushima, Japan. November 2-4, 2000. Abstracts]. PMID- 11253221 TI - [The 50th General Meeting of the Japanses Society of Allergology. Yokohama, Japan. November 30-December 2, 2000. Abstracts]. PMID- 11253222 TI - [The 46th Annual Meeting of the Japanese Association for Laboratory Animal Science. May 20-22, 1999. Ichikawa City, Japan. Abstracts]. PMID- 11253223 TI - [Regional meeting of the Japanese Society of Otolaryngology. 2000. Abstracts]. PMID- 11253224 TI - Deciding right from wrong: more complicated than it might appear. PMID- 11253225 TI - Emergency contraception and conscience clauses. PMID- 11253226 TI - Nutraceuticals: sorting out fact, fiction, and uncertainty. AB - Scientific evaluation of herbal products has been limited, yet herbal products are the most commonly consumed health care products. Because of known pharmacological effects and potential interactions of many of these compounds with therapeutic drugs, a history of herbal and dietary supplement intake should be considered part of the routine medical history and should be evaluated before any changes in prescription drugs and before medical procedures. PMID- 11253227 TI - Case report: 1884. Carcinoma of the stomach. PMID- 11253228 TI - Case report: 2000. Carcinoma of the stomach. PMID- 11253229 TI - Pharmacists' right to conscientious objection: the debate over Preven. PMID- 11253231 TI - Cancer of the male breast. AB - Cancer of the breast in men is an infrequent but serious problem. The epidemiology and clinical features of the disease generally parallel those of breast cancer in women, but affected men tend to be older, have subareolar tumors, and present in more advanced stages. Treatment of men follows the trends for treatment of women, with less radical surgery and more frequent use of systemic adjuvant therapy. Stage and axillary nodal status are the most important prognostic indicators for men. Although overall survival rates of males remain inferior to those of women, the indications are that breast cancer in men is equally curable in comparable stages. Men need to be sensitive to the signs of breast cancer and seek early consultation. PMID- 11253230 TI - Gender differences in the cross-sectional and cohort age dependence of cause specific mortality: the United States, 1962 to 1995. AB - OBJECTIVE: Gender- and age-specific mortality rates were calculated to determine the role of behavioral and biologically based differences in health. DESIGN: Changes in age-specific mortality rates were calculated for males and females across select time periods, in addition to gender- and cause-specific mortality age trajectories for birth cohorts for heart disease, stroke, and lung and breast cancer. Differences between cohort mortality age trajectories and the age pattern of mortality rates were related to socioeconomic differences between genders and to biological risk factors. SUBJECTS: U.S. males and females aged 30 to 95+ over the 34-year period 1962 to 1995. METHODS: Gender- and age-specific mortality rates were computed and displayed using the logarithmic transform of the rates for five-year age categories. RESULTS: Major gender cross-sectional mortality differences were found for heart disease, stroke, and lung cancer, with males being disadvantaged except for stroke. Less variation was found for lung cancer in recent male cohorts compared to large rates of change for recent female cohorts. Female breast cancer mortality showed modest variation, with most of the cohort differential in risk in postmenopausal breast cancer risk. Relative risk of lung cancer with age was greater for females. CONCLUSION: Differences in male and female cohort mortality rates underlie many of the period changes for heart disease, stroke, and lung cancer. Cohort differences were largest for lung cancer, although the cohort-specific smoking patterns these rates imply could also be responsible for some of the heart disease and mortality differences. The breast cancer mortality rates showed consistency with the concept of two distinct etiologies for breast cancer: premenopausal and postmenopausal. Large cohort increases in female lung cancer mortality may be alterable by behavioral measures and education. PMID- 11253233 TI - Gender-specific therapy: medication-related concerns and issues regarding HRT in women. PMID- 11253232 TI - Androgen physiology and the cutaneous pilosebaceous unit. AB - Hormones play an important role in cutaneous physiology. The androgen hormones are of particular relevance in modulating hair growth and sebum production in the pilosebaceous unit, which is comprised of the hair follicle and sebaceous gland. Testosterone arising from the circulation is converted peripherally to its more potent, reduced form, dihydrotestosterone, by the enzyme 5 alpha-reductase. Dihydrotestosterone is primarily responsible for androgen receptor binding and exerting end-organ effects. The clinical sequelae of enhanced local androgen production include androgenetic alopecia, hirsutism, and acne. These disorders can be fraught with significant psychosocial ramifications because of their highly visible nature and impact on perceptions of masculinity and femininity. PMID- 11253234 TI - Reconceptualizing medical necessity. PMID- 11253235 TI - Androgens and female sexuality. AB - An accumulating body of data indicates that many women experience a cluster of symptoms that are responsive to testosterone treatment and may be due to androgen deficiency. Characteristically, affected women complain of low libido, persistent fatigue, and diminished well-being and are found to have low circulating bioavailable testosterone. Whether the apparent therapeutic effects of testosterone are mediated via the androgen receptor or as a consequence of metabolism to estrogen is not known. Despite the lack of understanding of the mechanism(s) by which testosterone may enhance libido, the prescription of testosterone to women in a variety of formulations is becoming increasingly popular. This article provides an overview of the rationale for testosterone therapy in women, offers a broad definition of androgen deficiency in women based on the clinical experience of the author, and outlines the currently available options and potential risks of testosterone replacement in women. PMID- 11253236 TI - Gender differences in patients undergoing coronary artery bypass surgery, from a mandatory statewide database. AB - OBJECTIVE: To determine why women have a higher mortality rate than men when undergoing coronary artery bypass grafting. DESIGN: Retrospective analysis of patients entered in a mandatory state database. PARTICIPANTS: 19,224 patients who underwent coronary artery bypass grafting in New York State in 1995. METHOD: The authors evaluated data pertaining to 27 variables. They conducted univariate analysis using the Student t test for continuous variables and the chi-square test or the Fisher exact test for discrete data. They conducted multivariate analysis using a logistic regression model. RESULTS: Analysis of body surface area revealed that smaller size was a risk factor for both women and men. Analysis of age demonstrated increased risk for mortality in women in both older and younger subpopulations. Other significant variables included a lesser degree of revascularization and less frequent use of the internal mammary artery in women. CONCLUSION: Smaller size and advanced age alone do not explain why female gender is an independent risk factor for mortality from coronary artery bypass grafting. Increased mortality is probably due to the fact that women have more comorbid conditions than men at the time of referral, perhaps because they are not being evaluated aggressively enough. PMID- 11253237 TI - Gender-specific health care in diabetes mellitus. AB - This article reviews information on health care issues of specific clinical relevance for women with diabetes mellitus (type 1, type 2, and gestational diabetes mellitus), high-lighting several topics that require careful attention by the physician and the diabetes management team. Type 2 diabetes is more prevalent among women than men, making prevention and early detection particularly important in the treatment of women. Major areas of health care concern for women with diabetes include cardiovascular disease, mental health, infections, and contraception and fertility. Knowledge of lifespan issues from adolescence through menopause is crucial to the management of women with diabetes. PMID- 11253238 TI - The 1st Annual Conference on Gender-Specific Medicine. PMID- 11253239 TI - Medicine then and now: how far have we come? How much farther will we go? PMID- 11253240 TI - Protecting personal medical information. PMID- 11253241 TI - Case report: 1884. Mitral and aortic regurgitation. PMID- 11253243 TI - Gender and dietary influences on drug clearance. PMID- 11253244 TI - Sexual function and aging in men and women: community and population-based studies. AB - Research has consistently shown a decline in sexual activity with age for both men and women, with the reasons differing by gender. For men, the decline is primarily due to age, poor health, and medications, all of which have been related to erectile dysfunction. For women, health is less of a factor; having a functioning partner is a more important variable. Menopause has some negative impact on women's sexual interest and desire, but psychosocial factors appear to be more important. This article reviews research on age, gender, and human sexuality in the general (nonpatient) population. Additional longitudinal research is recommended to evaluate the long-term effects of declining ovarian function on postmenopausal women; the ways in which aging influences partners over time; and the impact of new medications and generational changes in sexual mores on sexual function in both genders. PMID- 11253245 TI - Gender differences in gastroesophageal reflux disease. AB - Gastroesophageal reflux disease (GERD) is a common chronic condition in the United States, affecting as many as 40% of adults. Although questionnaire-based studies have found the prevalence of the disease to be equal in men and women, the relative prevalence of GERD in males and females has yet to be established by quantitative, clinical evaluation. Moreover, preliminary research suggests that there are gender differences in the pathology and symptomatology of GERD, and the increased prevalence of GERD in pregnancy may indicate that sex hormones play a role in the disease. Additional research is necessary to confirm these findings. PMID- 11253246 TI - Clinical implications of prostate-specific antigen in men and women. AB - Prostate-specific antigen (PSA) is a valuable tumor marker for prostate cancer. Although it is indeed produced at an extremely high level by the prostate, PSA is also expressed in many female tissues, especially those regulated by sex steroid hormones. PSA is detected in both normal and abnormal breast tissue, as well as in various breast fluids, including milk, nipple aspirate, and cyst fluid. Clinical studies suggest that the presence of PSA in breast tissue may indicate a favorable prognosis for breast cancer patients. Levels of PSA in nipple aspirate fluid, however, may be indicative of breast cancer risk. Concentrations of PSA in serum are elevated in pregnant women as well as in women who have excess androgens. More studies are necessary to determine the clinical implications of the presence of PSA in amniotic fluid and female serum. PMID- 11253247 TI - Cross-national epidemiology of depression and gender. AB - Depressive disorders are 1.5 to 3 times more frequent in women than in men. This article reviews the evidence of gender differences in depression across countries in terms of definition, frequency, pattern, and burden. It attempts to confirm that these differences are real and not the result of any methodological bias; it examines what factors account for these differences; and it considers the implications of these differences for dealing with the increasing public health burden of this common mental disorder. PMID- 11253248 TI - Gender and ethnic differences in health behaviors and risk factors for coronary disease among urban teenagers: the PATH program. AB - OBJECTIVE: To assess gender and ethnic differences among teenagers in heart health behaviors, risk factors for coronary heart disease (CHD), and cardiovascular fitness. DESIGN: Observations consist of cross-sectional data collected prior to a school-based health promotion intervention program. PARTICIPANTS: Teenage girls (N = 865) and boys (N = 497) from three New York City high schools. The ethnic composition of this sample was 20% Asian-American, 40% African-American, 25% Hispanic, and 15% white. METHOD: Subjects were compared on the following: height, weight, body mass index, percentage body fat, total cholesterol, blood pressure, heart health knowledge, family history, socioeconomic status, dietary habits, smoking, physical activity, and estimated aerobic capacity. Differences were assessed with independent t tests, analysis of variance, and chi-square statistical techniques. RESULTS: Compared with girls, boys were more active and had higher estimated aerobic capacity, higher systolic blood pressure, and better self-perception of health. Compared with boys, girls had higher cholesterol, percentage body fat, and heart health knowledge scores and ate fewer foods high in saturated fat, cholesterol, salt, and simple sugars. Among girls, African-Americans had the highest blood pressure, cholesterol, body mass index, and intake of foods high in saturated fat, cholesterol, and sugar. Among boys, Hispanics had the highest body mass index and percentage body fat and the lowest heart health knowledge scores. White girls and white boys were the most frequent smokers. CONCLUSIONS: Poor health behaviors and risk factors for CHD occurred frequently among urban teenagers. In general, teenage girls had poorer health behaviors and a greater prevalence of risk factors than teenage boys, even though they scored better in heart health knowledge testing. Ethnic comparisons revealed poorer health behaviors and higher prevalence of risk factors in African-American and Hispanic teens compared with white and Asian American teens. Results support the need for health promotion intervention among urban teenagers. PMID- 11253249 TI - Multiple sclerosis may require gender-specific treatment. PMID- 11253250 TI - The NIH and women's health: praising--and criticizing--the right things. PMID- 11253251 TI - Domestic violence is a medical issue. PMID- 11253252 TI - The electrocardiographic QT interval and its prolongation in response to medications: differences between men and women. AB - Ventricular repolarization corrected for heart rate as measured by the electrocardiographic QTc interval differs in women compared to men. Although the mechanism remains unelucidated, the data are convincing that the effects of drugs that prolong ventricular repolarization are more likely to produce torsades de pointes arrhythmias in both healthy women and women with cardiac disease compared to men of similar ages and health status. Avoidance of such drugs and careful monitoring during necessary therapeutic administration of drugs that prolong ventricular repolarization is especially necessary in women. Electrolyte abnormalities such as hypokalemia and hypomagnesemia must be carefully avoided in patients receiving drugs that can produce QT prolongation. Because the recognition of this risk for drugs other than cardiac and psychotropic drugs has only recently emerged, it can be anticipated that the list of drugs known to produce such effects will grow. PMID- 11253253 TI - Systemic lupus erythematosus in males: a retrospective study with a review of the literature. AB - OBJECTIVE: To delineate the clinical pattern of a cohort of male patients with systemic lupus erythematosus (SLE) and compare it with previously reported data. DESIGN: Retrospective review of hospital records. SUBJECTS: Male patients (n = 11) who were diagnosed with SLE and admitted to a 500-bed university hospital between 1990 and 1998. Eight of the men were African-American and three were Latino. The mean age was 36 years (range, 29-46). METHODS: Clinical and laboratory data were collected according to a well-established protocol. Imaging and invasive studies (including aspirations and biopsies) were also recorded. RESULTS: Nine of the patients (82%) had renal involvement, with five needing dialysis within a year of presentation. Five patients had neurologic involvement: two presented with psychosis and three with seizures. Eight patients had hematologic involvement, and seven had serosal and articular involvement. Cutaneous lesions (discoid lupus) were noted in only one patient. A majority of the patients were noncompliant and were lost to follow-up; therefore, ultimate outcome could not be clearly delineated. CONCLUSION: Renal, serosal, neurologic, articular, and hematologic involvement occurred frequently in our sample of male patients with SLE. The most striking finding was the high frequency of renal involvement with poor prognosis. A high index of suspicion for SLE in males may permit earlier diagnosis and may dictate the need for more aggressive therapy. PMID- 11253254 TI - Psychoneuroimmunology and the faith factor. AB - A recent systematic review of research on religion and health has found a consistent relationship between religion and better mental health as well as greater social support. There also appears to be a relationship between religious involvement and better physical health, although the mechanism for this effect is poorly understood. One way that religion could impact physical health is through neuroendocrine and immune mechanisms, or psychoneuroimmunology (PNI). This article describes the physiological mechanisms by which the mind affects the body, reviews research substantiating the link between psychosocial processes and immune functioning, and examines the limited research that has addressed the religion-PNI relationship. Gender differences are emphasized. PMID- 11253255 TI - Sexual pharmacology in the 21st century. AB - Sexual dysfunction is highly prevalent in both sexes. Considerable progress has been made in the development of new pharmacologic treatments since the approval of sildenafil in 1998. A variety of oral erectogenic agents are available or are in late-phase development, including centrally active dopamine agonists (e.g., sublingual apomorphine), peripheral nonselective alpha-blockers (e.g., oral phentolamine), and other phosphodiesterase type-5 inhibitors (e.g., vardenafil). These drugs have recently been evaluated for the treatment of female sexual arousal disorder, although results to date have been inconclusive. Pharmacologic therapies have also been proposed for the treatment of premature ejaculation and hypoactive sexual desire disorder. Strong evidence exists for the value of serotonergic drugs (e.g., selective serotonin reuptake inhibitors) in the treatment of premature ejaculation. Further research is needed, particularly on the effects of these drugs on female sexual dysfunction. PMID- 11253256 TI - Mood disorders and the reproductive cycle. AB - Women have a significantly higher risk for developing mood disorders than men. Although reasons for this gender difference are not fully understood, it is clear that changing levels of reproductive hormones throughout women's life cycles can have direct or indirect effects on mood. Fluctuations in reproductive hormones may interactively affect neuroendocrine, neurotransmitter, and circadian systems. Reproductive hormones also may affect response to some antidepressant drugs and alter the course of rapid-cycling mood disorders. Nonpharmacologic interventions, such as light therapy and sleep deprivation, may be beneficial for mood disorders linked to the reproductive cycle. These interventions may have fewer side effects and a greater potential for patient compliance than some antidepressant drugs. PMID- 11253258 TI - Upholding life at all stages. PMID- 11253257 TI - Gender differences in the relationship between insulin-mediated glucose utilization and sex hormones in young African-Americans. AB - OBJECTIVE: To determine whether there are gender differences in insulin-mediated glucose utilization and if sex hormones correlate with measures of insulin sensitivity in young adult African-Americans. DESIGN: Cross-sectional case (women)-control (men) study. PARTICIPANTS: African-American men and women aged 27 to 35 years. Excluded were known diabetics, individuals on antihypertensive therapy, and women taking exogenous estrogen preparations. METHODS: Procedures included anthropometric and blood pressure measurement, oral glucose tolerance test, sex hormone assay, and euglycemic hyperinsulinemic clamp. Procedures for data analysis included two-way analysis of variance and Pearson's correlation coefficients. RESULTS: Data were analyzed on 104 men and 142 women with a mean age of 31.5 years. Insulin sensitivity was lower in women than in men. When insulin-mediated glucose utilization was corrected for body fat, there was no gender difference in insulin sensitivity. There was a significant correlation of androgen status with insulin sensitivity, but this relationship was divergent between men and women. For men, the correlation between insulin sensitivity and free testosterone was positive (r = .36, P < .001). For women, this correlation was negative (r = -.28, P = .001). CONCLUSION: These data on young African Americans demonstrate no gender differences in insulin sensitivity when glucose utilization is corrected for adipose mass. Androgen status is significantly linked with insulin sensitivity, but the relationship is divergent in men and women. Insulin resistance in young women is strongly associated with relative androgen excess, which may augment the risk for cardiovascular disease. PMID- 11253259 TI - Hormone therapy and smoking. PMID- 11253260 TI - Medical necessity. PMID- 11253261 TI - Gender differences in recovery from coronary artery disease. PMID- 11253262 TI - HRT, HERS, and the medical community: controversies and confusion or, "what is truth?" said Pilate. PMID- 11253264 TI - Authenticity and identity in contemporary adoptive families. PMID- 11253263 TI - Congestive heart failure medications: is there a rationale for sex-specific therapy? AB - The etiology and physiology of symptomatic heart failure may differ by sex. Prognostic markers for outcomes of CHF may also have sex-specific sensitivity. Insufficient numbers of women have been enrolled in most large interventional survival studies of pharmacologic therapy for CHF to test for sex-specific efficacy or responses. There are some data that demonstrate sex-specific lack of efficacy and increase in side effects of therapy. Given that the number of women admitted to hospitals with CHF each year exceeds the number of men, there is an obvious need to specifically and definitively evaluate the pharmacologic therapy of CHF in women. PMID- 11253265 TI - Urinary tract infection risk factors and gender. AB - Urinary tract infections (UTIs) are more common among women than men, although the prevalence in elderly men and women is similar. Most of the research on UTI has focused on young, sexually active women who are at high risk for developing an infection. The predominant UTI risk factors in young women are sexual intercourse and the use of spermicidal contraceptives. Other important UTI risk determinants in selected age groups include anatomic and physiologic factors, such as obstructing lesions and estrogen deficiency; genetic factors, such as blood group secretor status; antibiotic exposure; functional status; and possibly receptive anal intercourse and HIV infection. PMID- 11253266 TI - Influence of gender on prevention of myocardial infarction by antihypertensives and acetylsalicylic acid: the HOT study. AB - OBJECTIVE: The aims of the Hypertension Optimal Treatment (HOT) Study were to investigate the relationship between three levels of target office diastolic blood pressure (BP; < or = 90, < or = 85, and < or = 80 mm Hg) and cardiovascular death, myocardial infarction (MI), and stroke in hypertensive patients, and to examine the effects of 75 mg of acetylsalicylic acid (ASA) daily versus placebo. DESIGN: Randomized, double-blind study. This substudy assessed the influence of gender on the incidence of MI. SUBJECTS: A total of 18,790 patients (mean age, 61.5 years; range, 50-80 years). METHODS: Patients were randomized and followed for an average of 3.8 years until 71,051 patient-years had elapsed and 683 events, including 215 MIs, had occurred. RESULTS: There were significantly fewer MIs in the lowest diastolic BP target group (P = .034) in women (n = 8883); a similar but smaller trend was not statistically significant in men. The effect of ASA on preventing MI was also influenced by gender (P = .38 in women; P = .001 in men [lowered by 42%]). CONCLUSION: Lowering diastolic BP to about 80 mm Hg in hypertensive women and administering 75 mg of ASA daily to well-treated hypertensive men reduces MI in patients with essential hypertension. PMID- 11253267 TI - Gender differences in osteoporosis treatment: a review of clinical research. AB - Morbidity from osteoporosis in the aging population is substantial in both men and women; however, therapeutic options approved by the Food and Drug Administration are open only for use in women. A literature search was performed to define gender differences in pharmacologic osteoporosis treatments in published research. Only two studies on the efficacy of pharmacologic osteoporosis interventions in both genders were found, both of which involved glucocorticoid-treated patients. There is clearly a lack of information on osteoporosis treatment approaches in men compared to women, despite the increasingly recognized risk of osteoporosis in men. Although there are no approved therapies for osteoporosis in men in the United States, the scant but promising evidence suggests potential benefit from alendronate and risedronate in men as well as women. Further gender-comparative research regarding nonestrogenic approaches to osteoporosis is warranted. PMID- 11253269 TI - Thrombotic stroke in an otherwise healthy middle-aged female related to the use of continuous-combined conjugated equine estrogens and medroxyprogesterone acetate. PMID- 11253268 TI - Gender differences in asthma in childhood and adolescence. AB - Asthma is a common chronic disease that can have a significant impact on individuals' daily lives. It is characterized by wheeze, shortness of breath, chest tightness, and cough secondary to airway inflammation and hyperresponsiveness to a variety of stimuli. Asthma is far more common in boys than girls during early childhood. The prevalence equalizes between the genders during adolescence and then switches to a female predominance in adulthood. This article reviews the epidemiology and possible pathophysiologic mechanisms for the observed differences in asthma between the genders. In practical terms, the impact of asthma may be different according to gender in terms of daily activities for children and adolescents. The implications of gender differences in asthma for the health professional will also be discussed. PMID- 11253270 TI - Use of female standard leads to underdiagnosis of osteoporosis in men. PMID- 11253271 TI - Blood pressure predicts heart disease differently in males and females. PMID- 11253272 TI - Health and the quality of life. PMID- 11253273 TI - Social environment, behavior, and schistosomiasis in an urban population in the northeast of Brazil. AB - The objective of our study was to gain greater insight into the pattern of water contact in relation to schistosomiasis among residents of Sao Lourenco da Mata, a town in Pernambuco, a state in the Northeast of Brazil. We had two primary objectives: 1) to identify water contact activities that were more likely to produce infection and 2) to examine the socioeconomic factors behind water contact activities. Using a sample of persons 10-25 years old, we carried out a population-based case-control study to investigate the first objective, and a cross-sectional study for the second objective. We found that leisure water contact with flowing water (stream or river) was significantly associated with schistosomiasis. Variables showing a statistically significant association with leisure water contact were economic sector, income, and level of education of the head of the household; type of housing; possessions inside the house; type of domestic water supply; and method of excreta collection. We introduced these variables into a multivariate model to select the ones that were most closely associated with leisure water contact. We used a stepdown procedure, and two variables were retained in the final model: type of domestic water supply and type of housing. We concluded that a decrease in leisure water contact was associated with better socioeconomic conditions. Our results suggest that with the subjects we studied in Sao Lourenco da Mata there was a socioeconomic determination for leisure water contact. Our data highlight the importance of a broad and integrated approach in studying water contact activities and in implementing behavioral interventions for schistosomiasis prevention and control. PMID- 11253274 TI - [Resistance of Plasmodium falciparum to 3 antimalarials in Turbo (Antioquia, Colombia), 1998]. AB - In 1998 we determined in vivo and in vitro the frequency and the degree of resistance of Plasmodium falciparum to the three antimalarials (chloroquine, amodiaquine, and sulfadoxine/pyrimethamine) most utilized in the municipality of Turbo (in the area of Uraba, Antioquia, Colombia), in a sample representative of the population with malaria. We carried out clinical and parasitological analyses over a 14-day period using the standard test recommended by the World Health Organization. In vivo, P. falciparum showed resistance to chloroquine, amodiaquine, and sulfadoxine/pyrimethamine, with a frequency of 97%, 7%, and 13%, respectively. In vitro, the corresponding figures were 21%, 23%, and 9%, respectively. For chloroquine the level of agreement between the in vivo and in vitro results was 23%. PMID- 11253275 TI - [Effect of sodium fluoride mouth rinses containing xylitol and sorbitol on the number of Streptococcus mutans from human saliva]. AB - The objective of this study was to assess the effect of 0.05% sodium fluoride solutions containing 2.5% or 12.5% xylitol on the number of Streptococcus mutans in the human mouth. Fifty boys between 8 and 16 years of age participated in this double-blind crossover study. Of the original 50 boys, 33 finished the study. Participants were randomly divided into four groups. The following solutions were employed: placebo solution; 0.05% sodium fluoride solution; 0.05% sodium fluoride + 2.5% xylitol + 2% sorbitol; 0.05% sodium fluoride + 12.5% xylitol + 2% sorbitol. Each solution was used for a 28-day period (20 mL/day, twice a day), with a 10-day washout period between solutions. There were no significant differences (P = 0.32) between the two xylitol-containing solutions (2.5% vs. 12.5%) concerning the number of Streptococcus mutans. However, there was a significant difference between these two xylitol-containing solutions and the sodium fluoride and placebo solutions (P < 0.001). Our results suggest that the 0.05% sodium fluoride solutions containing either 2.5% or 12.5% xylitol caused a significant reduction in the number of Streptococcus mutans. PMID- 11253276 TI - [Feasibility of a vaccine against rotavirus for developing countries]. PMID- 11253277 TI - [Phage typing of Salmonella enteritidis isolated from clinical, food, and poultry samples in Chile]. AB - Since 1994 an extensive epidemic of infections with Salmonella enteritidis (S. enteritidis) has affected Chile. In order to understand the diversity of infective sources, the possible origin of the epidemic, and the epidemiological relationships between clinical, food, and poultry isolates, we carried out phage typing of three groups of samples: 1) 310 S. enteritidis clinical samples collected between 1975 and 1996, 2) 47 food isolates obtained during S. enteritidis outbreaks, and 3) 27 strains isolated in surveillance studies of poultry-raising establishments. With the clinical samples, a total of 13 phage types were identified, 2 isolates could not be typed, and 1 was considered atypical. The phage types that were identified most frequently were 1 (56.8%) and 4 (31.3%), trailed by type 8 (4.8%) and type 28 (1.9%). Over time and in different regions of the country there were major changes in the distribution of the phage types. In the first years of collection the only phage types registered were 8 and 28, which disappeared around 1980 and then began reappearing sporadically in 1996. With the gradual S. enteritidis expansion that started in 1988, in the central and southern areas of the country phage type 4 began to appear; that type had not been found before in Chile. In 1991 in the northern area of the country phage type 1 began to predominate; it was another type that had not been reported before in Chile. In the food isolates the only phage types identified were 1 and 4, which were also the most common in the poultry isolates. Phage typing of S. enteritidis has proved to be useful in guiding the epidemiological analysis of the infections caused by this pathogen. PMID- 11253278 TI - New directions for the Revista/Journal. PMID- 11253279 TI - [Heparin induced thrombocytopenia, left ventricular thrombus and cerebral embolism during an acute myocardial infarction]. AB - HISTORY AND CLINICAL FINDINGS: A 58-year-old male patient was admitted to the emergency room after reanimation in hypotensive shock, under sedation and respiration. INVESTIGATIONS: The cardiac laboratory markers were within normal limits. The ECG demonstrated a sinustachycardia with typical infarction signs. The echocardiogram showed a hypokinesia of the cardiac apex. DIAGNOSTICS, TREATMENT AND COURSE: Acute myocardial infarction was treated with a systemic rt PA lysis. The course was complicated by: confirmed heparin-induced thrombocytopenia, left ventricular thrombus and a cerebral embolism. After multiple adaptations of the anticoagulation regimen, the patient was discharged in a good condition without major neurologic deficits. CONCLUSION: With a careful anticoagulation regimen it is possible to achieve a successful outcome in major complications such as heparin-induced thrombocytopenia, left ventricular thrombus and cerebral embolism. PMID- 11253280 TI - [Sweet syndrome and erythema nodosum in ulcerative colitis, refractory to steroids: successful treatment with tacrolimus]. AB - BACKGROUND: Inflammatory bowel disease is accompanied by cutaneous manifestations in about 10% of cases. Erythema nodosum and pyoderma gangraenosum are most frequently observed, which often subside on treatment of the underlying disease. CASE REPORT: A 30-year-old male with a history of long-standing ulcerative colitis experienced an acute attack despite treatment with azathioprine. Further he noticed dull red, elevated and tender maculae on the forelegs. A disseminated and papulosquamous exanthema arose on the back of the trunk and the upper extremities without pruritus. Well-being was compromised and blood sampling revealed an inflammatory response. High-dose steroids with antibiotics were without benefit until they were combined with tacrolimus, an immunosuppressive agent acting similar to ciclosporin. Remission occurred rapidly and the skin lesions resolved. Six months later the patient is currently still in remission and developed no signs of recurrent exanthema. CONCLUSION: The cutaneous lesions are thought to be related to ulcerative colitis and were classified as erythema nodosum and Sweet syndrome. This is the first report on the successful use of tacrolimus in steroid-refractory ulcerative colitis with extraintestinal cutaneous involvement. PMID- 11253281 TI - [Gastroenteritis due to Plesiomonas shigelloides--rare cases in the Western world]. AB - BACKGROUND: Plesiomonas shigelloides is a common pathogen in tropical regions, whereas it is rarely isolated in temperate climates. It is most often found in surface water and fish. During the last 10 years it was found to cause gastroenteritis 6 times in Ludwigshafen. Not all of these patients reported a trip to foreign countries. CASE REPORT: A 54-year-old male patient was hospitalized after a trip to Malaysia with strong greenish watery diarrhea and chills. On physical examination we saw a dehydrated patient in severely reduced general condition. The stool frequency was 30/d. The laboratory examinations only showed elevated parameters of inflammation. Plesiomonas shigelloides was cultivated in the stool cultures. With appropriate substitution of fluid and electrolytes, and antidiarrheal therapy the patient resumed a normal diet without any complications. Three days later his bowel movements were normal and his general condition was greatly improved. We withheld antibiotic therapy because of the noncomplicated course of illness. CONCLUSION: In Germany infections with Plesiomonas shigelloides are rare, an increase is observed because of increasing tourism to tropical regions. The course of infection is sometimes asymptomatic, but usually patients develop an acute gastroenteritis. Especially immunocompromised patients can show serious courses of infection. Plesiomonas shigelloides should be included in the differential diagnosis of acute gastroenteritis after journeys to tropical regions. Some of our patients, however, denied traveling to tropical regions. They also denied consuming seafood, which indicates a risk of infection in Germany. Still an infection with Plesiomonas shigelloides seems to be rare in northern European countries. PMID- 11253282 TI - [Beau's lines and chemotherapy]. PMID- 11253283 TI - [Interpretation of efficacy measures derived from 2 X 2 tables for the evaluation of diagnostic tests and treatment]. AB - BACKGROUND: To describe the efficacy of diagnostic tests and the effect of treatment a number of measures are used, which can be derived from 2 x 2 tables of frequencies. For the comprehension of these measures the knowledge of their properties in the framework of probability theory is necessary. MATERIAL AND METHOD: After an introduction of basic terms such as probability, odds, joint and conditional probability the usual measures sensitivity, specificity, likelihood ratio, positive and negative predictive value, relative risk, odds ratio, relative risk reduction, absolute risk reduction, and number needed to treat are presented and explained. In particular, the importance of disease prevalence and baseline risk for the interpretation of these measures is pointed out by means of examples. CONCLUSION: If the disease prevalence or the baseline risk is not appropriately taken into account, the efficacy of a diagnostic test and the effect of a treatment are overestimated, especially in screening and prevention trials. PMID- 11253284 TI - [Public health costs of influenza in Germany 1996- a cost-of-illness analysis]. AB - Influenza is often seen as an unproblematic and self-limiting disease despite putting a high burden on patients as well as being of high socio-economic relevance to societies. This analysis aims to visualize that influenza deserves a rise in attention especially because of its socio-economic relevance, based on a transparent methodology. This analysis investigates the costs caused by influenza in Germany in 1996. The analysis is based on a costs model that takes in official aggregated statistical and publicly available data. A top-down approach based on the ICD-9 Code 487 is used. The costs of influenza in 1996 came to nearly DM 5 billion. Only a small proportion of this was due to direct treatment costs (DM 0.6 billion). The direct costs are made up of 304 million DM (52.8%) for ambulatory care, 214 million DM (37.1%) for prescription-only and OTC medication, and 58 million DM (10.1%) for inpatient treatment including rehabilitation measures. The bulk of the indirect costs, some DM 4.4 billion (99.5%), was due to unfitness for work, whereas only about 21 million DM (0.5%) was accounted for by occupational disability and deaths. On the basis of an estimated 4 million cases of influenza in 1996 (no epidemics), the costs work out at about 1,237 DM per patient per year. In the event of an epidemic, the German economy would be faced with correspondingly higher costs of over DM 10.5 billion. Effective prevention and treatment strategies such as better pre-season vaccination rates, early diagnosis and effective antiviral therapy can help to reduce this financial burden. PMID- 11253285 TI - [Assessment of diabetic alterations of microcirculation by means of capillaroscopy and laser-Doppler anemometry]. AB - BACKGROUND: Only minor changes of skin capillary morphology have been described in diabetic patients by means of capillaroscopy, whereas cutaneous microvascular dysfunction is well known. We examined correlations between functional and morphological abnormalities of the capillaries in Type 1 and 2 diabetic patients and the influence of diabetes duration on capillary morphology. PATIENTS AND METHODS: Density, diameters and morphology of nailfold capillaries were investigated in diabetic patients, 16 Type 1 and 19 Type 2, and compared to age- and sex-matched control groups. Capillary blood cell velocities (CBV) during rest and after 3-minute arterial occlusion were measured in the dorsal middle phalangeal area of the left ring finger by means of laser Doppler anemometry. RESULTS: Capillary density, width and arterial limb diameter were similar in Type 1 and 2 diabetic patients compared to their controls. Capillary diameters of the apical part and the venous limb were enlarged in the combined analysis of Type 1 and 2 diabetic patients compared to the control group (apex: 19.2 +/- 0.6 microns vs 17.4 +/- 0.6 microns, p = 0.0243; venous limb: 17.3 +/- 0.5 microns vs 15.9 +/ 0.4 microns, p = 0.0238). Tortuous capillaries were more often observed in Type 1 (n = 13 vs n = 7, p = 0.028) and 2 diabetic patients (n = 16 vs n = 9, p = 0.019) than in controls. In Type 1 diabetic patients an inverse correlation (r = 0.52; p = 0.019) was found between capillary density and resting CBV. In Type 2 diabetic patients capillary apex diameter correlated positively with peak CBV (r = 0.49; p = 0.017). Disease duration correlated inversely with arterial limb diameter (r = -0.48; p = 0.020) and width of the capillaries (r = -0.48; p = 0.018) in Type 2 diabetic patients. CONCLUSION: Tortuous and dilated capillaries, indicating microangiopathy, were found in the skin of diabetic patients by means of capillaroscopy. Using laser Doppler anemometry it is possible to assess impairment of postocclusive reactive hyperemia, due to diabetic microvascular dysfunction, in single capillaries. Correlations between morphological and functional microcirculatory alterations in diabetes may be explained by hemodynamic changes, depending on diabetes duration. PMID- 11253286 TI - [Hypertensive disorders in pregnancy]. AB - BACKGROUND: Hypertensive complications contribute to maternal and fetal morbidity. Hypertensive diseases in pregnancy comprise various disorder from transient hypertension to the dangerous preeclampsia/eclampsia. Diagnosis of these diseases requires an understanding of the normal physiological adaptations during pregnancy. PATHOGENESIS: The primary cause of preeclampsia/eclampsia is a disturbed growth of throphoblast cells, probably induced by an altered maternal immunotolerance. The consequence is a dysfunction of endothelial cells with a decrease in perfusion of the uterus and placenta. The normal balance between vasoconstrictors and vasodilators is changed in favor of vasoconstrictors. Complex changes in the renin-angiotensin system have been detected resulting in an increased angiotensin II-mediated vasoconstriction. The reduction in perfusion of the uterus and placenta eventually leads to preeclampsia/eclampsia and growth retardation of the fetus. Manifest preeclampsia/eclampsia is characterized by disturbed microcirculation of target organs such as brain, liver and kidney. An involvement of the liver causes the HELLP syndrome. THERAPY: Various pharmacological approaches to prevent preeclampsia/eclampsia showed disappointing results, but patients with a risk for the eventual development of preeclampsia/eclampsia should be identified, closely monitored, and hypertension should be treated. A systolic blood pressure > 170 mm Hg and diastolic blood pressure > 100 mm Hg should be treated. Drugs such as alpha-methyldopa and dihydralazine that are well-characterized in their fetal effects are the primary choice for the treatment of hypertension in pregnancy. ACE-inhibitors and angiotensin II receptor antagonists are absolutely, diuretics are relatively contraindicated. The causal therapy for preeclampsia/eclampsia is delivery. Gravida before the 33th week of pregnancy should be admitted, hypertension should be treated, and the fetus should be monitored by duplex ultrasound and cardiotocography. New data suggest that early treatment with glucocorticoids may prevent the manifestation of HELLP syndrome. Hypertensive pregnant patients should be treated in tertiary centers with an interdisciplinary approach involving obstetricians, neonatologists, and nephrologists. PMID- 11253287 TI - [Increasing incidence and mortality of non-Hodgkin lymphomas. An epidemiological review of recent studies on risk factors for non-Hodgkin lymphoma]. AB - BACKGROUND: Non-Hodgkin's lymphoma (NHL) are among the small number of malignant tumors with markedly increasing incidence and mortality rates in the recent past. This trend is particularly obvious in industrialized countries. The causes of the observed increase remain unclear. METHODS: We conducted a Medline search to identify case control and cohort studies on medical, biological and selected environmental risk factors of non-Hodgkin's lymphoma published between 1992 and 1998. Methodological aspects and results of identified studies are presented in tabular form. We furthermore discuss the role of various risk factors for the observed trends in non-Hodgkin's lymphoma. RESULTS: We identified a total of 64 studies. Iatrogenic immunosuppression as well as numerous diseases associated with an impaired immune system have clearly been recognized as etiological factors for non-Hodgkin's lymphoma. However, they can explain only a small percentage of the rate increases of non-Hodgkin's lymphoma. Similarly, the HIV/AIDS epidemic in Germany is responsible only for a small proportion of the rising figures of non-Hodgkin's lymphoma; other viral agents are currently being associated with a few distinct subtypes of non-Hodgkin's lymphoma. Smoking and nutritional factors are weakly or not at all associated with non-Hodgkin's lymphoma and can not account for the disease trends. CONCLUSION: The observed secular rise in incidence and mortality of non-Hodgkin's lymphoma can only partly be explained. New epidemiologic studies should focus on risk factors associated with the function of the immune system and on possible interactions between different etiological factors. PMID- 11253289 TI - The quiet revolution in dentistry. PMID- 11253288 TI - Management of cancer pain. PMID- 11253290 TI - Let's face it. PMID- 11253291 TI - Insurance companies and third-party payments. PMID- 11253292 TI - Carbon fibre posts. PMID- 11253293 TI - Why the success of dentistry's charitable foundation should matter to you. PMID- 11253294 TI - Occlusion: the standard of care. PMID- 11253295 TI - Occlusion: the "science-based" approach. PMID- 11253296 TI - Is there a sound basis for deciding how many dentists should be trained to meet the dental needs of the Canadian population? Systematic review of literature (1968-1999). AB - A systematic review was conducted of the literature on human resources planning (HRP) in dentistry in Canada, critically assessing the scientific strength of 1968-1999 publications. Inclusion and exclusion criteria were applied to 176 peer reviewed publications and "grey literature" reports. Thirty papers were subsequently assessed for strength of design and relevance of evidence to objectively address HRP. Twelve papers were position statements or experts' reports not amenable for inclusion in the system. Of the remaining 18 papers, 4 were classified as projections from manpower-to-population ratios, 4 as dental practitioner opinion surveys, 8 as estimates of requisite demand to absorb current capacity and 2 as need-based, demand-weighted studies. Within the 30.5 years reviewed, 53.4% of papers were published between 1982 and 1987. Overall, many papers called for a reduction in human resources, a message that dominated HRP during the 1980s, or noted an increase in the demand for services. HRP publications often had questionable strength or analytic frameworks. The paradigm of busyness-scarcity evolved from a belief around an economic model for the profession into a fundamental tenet of HRP. A formal analysis to establish its existence beyond arbitrary dentist:population ratios has usually been lacking. PMID- 11253297 TI - Autogenous tooth transplantation: an alternative to dental implant placement? AB - Autogenous tooth transplantation, or autotransplantation, is the surgical movement of a tooth from one location in the mouth to another in the same individual. Once thought to be experimental, autotransplantation has achieved high success rates and is an excellent option for tooth replacement. Although the indications for autotransplantation are narrow, careful patient selection coupled with an appropriate technique can lead to exceptional esthetic and functional results. One advantage of this procedure is that placement of an implant supported prosthesis or other form of prosthetic tooth replacement is not needed. This article highlights the indications for autogenous tooth transplantation using 3 case reports as examples. A review of the recommended surgical technique as well as success rates are also discussed. PMID- 11253298 TI - The use of resin cements in restorative dentistry to overcome retention problems. AB - The use of resin cements in combination with dentin bonding agents can result in superior attachment of prostheses to tooth structure. This paper describes four clinical cases in which dentin-bonded resin cements were used to overcome retention problems. In the first case, a detached fixed partial denture, which was in good condition when separated, was recemented to abutment teeth prepared with less-than-ideal angle of convergence. In the second case, a detached all porcelain crown was recemented with a dentin-bonded resin cement after appropriate surface treatment. In the third case, a porcelain-fused-to-metal crown made for a molar tooth was cemented to a short clinical crown, avoiding crown-lengthening surgery. In the fourth case, a 3-unit fixed partial denture was recemented to abutments with less-than-ideal supporting features. Dentin-bonded resin cements can help to extend the life of detached prostheses until the patient is financially prepared for replacement or it can help to avoid crown lengthening surgery. PMID- 11253299 TI - Bacteriophage therapy for bacterial infections. Rekindling a memory from the pre antibiotics era. PMID- 11253302 TI - Dirt, disgust, and disease. Is hygiene in our genes? PMID- 11253303 TI - Human language and our reptilian brain. The subcortical bases of speech, syntax, and thought. PMID- 11253304 TI - The significance of wheat in the Dakota Territory, human evolution, civilization, and degenerative diseases. PMID- 11253305 TI - Personal reflections on the "animal-rights" phenomenon. PMID- 11253306 TI - Sir John Eccles, 1903-1997. Part 1. Onto the demonstration of the chemical nature of transmission in the CNS. PMID- 11253307 TI - The problems of seeing and saying in medicine and poetry. PMID- 11253308 TI - Neurology, technology, and the diagnostic imperative. AB - Advancements in diagnostic technologies have revolutionized the field of neurology. The use of these tools in the course of neurological evaluations is driven by a strong version of the diagnostic imperative, with the goal of precisely identifying the locus and extent of disease processes. Because of the discrepancy between the sophistication of these technologies and the availability of therapeutic interventions, there is active debate regarding the appropriate use of these tools when the diagnosis is clear, or when no change is made to the therapeutic management. A narrow view of management that is bounded by the availability of pharmacological or surgical interventions results in a more rigid dichotomy between the needs of doctors and patients. A broader view that relaxes the constraint between diagnostic procedures and interventions is more in keeping with the observation that many acts are performed for the benefit of doctors and patients alike. An historical and ethical analysis of the diagnostic imperative, with attention to the rise of innovative medical technologies and current concepts of therapeutic intervention, can help clarify the principles of medical paternalism and beneficence that guide current models of decision making in the neurological sciences. PMID- 11253309 TI - [The post-perfusion syndrome after operations performed with extracorporeal circulation]. AB - Postperfusion syndrome (PPS) is a dreaded complication of cardiac surgery operation in extracorporeal circulation (ECC). Four factors play a key role in its pathophysiology: 1. contact of blood with the material of ECC, 2. release of activated leucocytes from pulmonary bed after the release of aortic cross-clamp, 3. translocation of endotoxin due to gut ischemia and its consequent reperfusion, 4. activation of coagulation, fibrinolytic, kallikrein-kinin and complement systems. The occurrence of PPS can be limited by reducing the ECC time and/or cross-clamp time, by using membrane oxygenator in the system of ECC, by using polyester or polypropylene in the set of ECC, by using heparin-coated set or leucocyte filter in the system of ECC, by application of pharmacological dosis of corticosteroids prior to the ECC, by early enteral nutrition. In eligible patients it is possible to eliminate the risk of PPS completely by using the operation without ECC. The experience of the authors with above mentioned problems is given and confronted with literature. PMID- 11253310 TI - [Effect of polycyclic aromatic hydrocarbons on the immune system]. AB - Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous compounds in our environment. They are formed during incomplete burning of coal, oil, gas, wood, garbage or other organic substances such as tobacco and charbroiled meat. PAHs enter the environment mostly as releases to air from volcanoes, forest fires, residential wood and coal burning and exhaust from automobiles and trucks. There are more than 100 different PAHs. Some of them have danger toxic properties including mutagenic and carcinogenic potential. Since 1993 PAHs are classified as the compounds which can cause immunosuppression. They reduce the body resistance against infection and cancer diseases. Immunosuppressive, carcinogenic and hypersensitivity effects of some PAHs representatives were described well in experimental studies. On the other hand, only sporadic information about immunological changes after long-term occupational exposure to PAHs were found from existing human epidemiological database. In addition, these studies usually did not cover the complex immunological profile represented by cellular and humoral activity. PMID- 11253311 TI - [Prion diseases. Subacute spongiform encephalopathies in humans and animals (neural "anthropo- and zooprionoses")]. AB - General review concerning current problems of subacute spongiform encephalopathies (Prion-diseases) in humans and in animals. The work is based on the aetiological conception of "Prion" as an allosteric conformational variant of the Prion-protein with autocatalytic "infectious" properties. The therm "Anthropoprionosis" and "Zooprionosis" are recommended by the author for human and animal Prion-diseases. Actual medical problems of various Prion-diseases are discussed in more detail, particularly those of aetiopathogenesis, genetics and molecular-biological principles in Prion-replication as well as the consensual neuropathological and clinical diagnostical criteria. Neuropathological findings are evaluated based on personal experience of diagnosed and/or consulted CJD cases. On the ultrastructural level the morphological proof of abnormal secondary phagosomes in neurons of the CNS with accumulation of protein material ("prionosomes") is documented. Finally, the risks of CJD and the infectious potential of various organs and contaminated materials are also mentioned. In the methodical part, safety precautions in handling histological material from stereotactic biopsies and necropsies of brain are described, including safety techniques of autopsy in cases of CJD. PMID- 11253312 TI - [Phenotyping of cytochrome P-450 1A2 and N-acetyltransferase (NAT2) using the in vivo caffeine test as a tool for determining individual susceptibility to selected xenobiotics]. AB - An in vivo phenotyping method of CYP1A2 (cytochrome P-450 1A2) and NAT2 (N acetyltransferase, isoform 2) based on caffeine test (Butler et al. Pharmacogenetics 1992;2:116-127) with modifications has been established. Molar concentrations of caffeine and its metabolites in urine were determined by HPLC with gradient elution and spectral detection (200-350 nm). The results obtained with this method were sufficiently accurate and precise and allowed to calculate the respective parameters which were subsequently used to evaluate the CYP1A2 and NAT2 activities. PMID- 11253313 TI - [Changes in late potentials in the time period after myocardial infarct]. AB - Late potentials are thought to be a non-invasive marker of discrete morphologic and electrophysiologic changes of the ventricular myocardium caused by fibrosis due to myocardial infarction. To find out whether there are significant changes in the signal-averaged ECG (SA ECG) over one year we examined 30 patients following myocardial infarction. When compared to the initial findings, significantly decreased incidence of late potentials was found after one year. Most of the changes in the filtered QRS were located in the terminal portion of the fQRS--in the area of low amplitude signals. Significant differences were noted in the changes found in patients with anterior myocardial infarction (MI) compared with the changes found in patients with posterior MI. PMID- 11253314 TI - Methodological considerations for researchers and practitioners using pedometers to measure physical (ambulatory) activity. AB - Researchers and practitioners require guidelines for using electronic pedometers to objectively quantify physical activity (specifically ambulatory activity) for research and surveillance as well as clinical and program applications. Methodological considerations include choice of metric and length of monitoring frame as well as different data recording and collection procedures. A systematic review of 32 empirical studies suggests we can expect 12,000-16,000 steps/day for 8-10-year-old children (lower for girls than boys); 7,000-13,000 steps/day for relatively healthy, younger adults (lower for women than men); 6,000-8,500 steps/day for healthy older adults; and 3,500-5,500 steps/day for individuals living with disabilities and chronic illnesses. These preliminary recommendations should be modified and refined, as evidence and experience using pedometers accumulates. PMID- 11253315 TI - Individual differences and similarities in the stability, timing consistency, and natural frequency of rhythmic coordinated actions. AB - Under preferred speed conditions, 15 adults undertook bimanual in-phase and antiphase tapping, clapping, galloping, galloping while clapping, and crawling on their hands and feet. We measured stability of interlimb coordination (standard deviation of mean interlimb relative phasing), single limb timing consistency (coefficient of variation of mean single limb cycle durations), and natural limb frequency. Pearson product-moment correlations among tasks established that only the natural limb frequencies were significantly correlated (specifically among gross motor actions in which larger contributions of inertial loads contribute to natural frequencies). Intraclass correlations were high for tasks, meaning that within each task, all participants performed similarly. Thus, only frequency has a tendency to show a common time-based process within a participant, but common time-based processes exist between participants. PMID- 11253316 TI - Perceptual decision making for baseball pitch recognition: using P300 latency and amplitude to index attentional processing. AB - This study was designed to examine the perceptual and attentional processes associated with the effects of administering a cost-benefit precuing paradigm to intermediate and advance-level baseball batters. Psychophysiological and performance data obtained from 10 advanced and 10 intermediate-level players were compared. A total of 400 pitches (200 fastballs, 200 curveballs) was randomly presented via a large projection screen, and participants pressed one of two buttons to indicate the type of pitch thrown. Verbal precues were given for 300 of the pitches. Of those, 75% were valid, and 25% were invalid. Electroencephalographic data collected from the P location was used to assess the latency and amplitude of P300. Analysis of variance (Skill Level x Precue x Pitch) for P300 and reaction time (RT) indicated that intermediate batters produced shorter P300 latencies, larger P300 amplitudes, longer RTs, and less correct responses than the advanced batters; the effects were more pronounced for the curveballs. These results suggest that intermediate batters are less efficient in their perceptual decision-making processes due to greater limitations in attentional capacity when compared with advanced batters. PMID- 11253317 TI - Manipulating generalized motor program difficulty during blocked and random practice does not affect parameter learning. AB - Blocked practice engenders more trial-to-trial response stability, which is thought to be crucial for developing the generalized motor program (GMP) but not parameter learning (Lai, Shea, Wulf, & Wright, 2000). It was hypothesized that reducing the difficulty of the GMP might permit additional cognitive resources to be allocated to learning the parameter requirements. However, GMP theory maintains the independence of the memories governing the GMP and parameters. This notion suggests that manipulating the difficulty of the GMP should have no effect on the blocked practice participant's ability to successfully specify the appropriate parameters. Participants learned a simple or complex relative timing pattern under either blocked or random practice conditions. Smaller GMP errors were exhibited for the simple relative timing patterns, but this was not associated with improvements in parameter specification following blocked practice. A clear advantage for parameter specification was evident in transfer following random practice. Taken together, these data support the theoretical separation of the GMP and parameter processes. PMID- 11253318 TI - Training patterns of athletes during pregnancy and postpartum. AB - The purpose of the present investigation was to examine exercise patterns and psychological variables mediating a return to training and competition after pregnancy. Competitive female athletes who had given birth within the last 10 years completed surveys concerning (a) training patterns before, during, and after childbirth, (b) childbirth complications and training advice, (c) perceptions of success in their postpartum comebacks to training, and (d) self efficacy, social support, and perceived barriers to training during pregnancy and after childbirth. Results indicated that women decreased both cardiovascular and resistance training during pregnancy. Additionally, training efforts during pregnancy were independent of those during the pre- and postpartum periods. This finding suggests that athletes may be able to alter their training patterns during pregnancy without a significant impact on their postpartum training program. PMID- 11253319 TI - Children's self-efficacy, motivational intentions, and attributions in physical education and sport. AB - The purpose of this study was to examine how differences in children's self efficacy, age, and gender impact motivational intentions, future self-efficacy, and attributions following perceptions of failure. Children, ages 8-14 years (N = 289), were assigned to either high or low self-efficacy groups, and measures of intended effort, persistence, choice, future self-efficacy, and attributions for failure were collected following a failure scenario. Results indicated that children with higher self-efficacy chose to participate and had higher future self-efficacy scores than those with lower self-efficacy. Higher efficacy children attributed failure to lack of effort, whereas, those with lower efficacy attributed failure to lack of ability. Age-related differences were also found with choice to participate, effort, and future self-efficacy. PMID- 11253320 TI - Physical self and physical activity relationships in college women: does social physique anxiety moderate effects? AB - This research assessed whether social physique anxiety moderated the relationship between physical self-perceptions and the level of physical activity involvement in young women. Participants were 354 female students who completed the Social Physique Anxiety Scale (SPAS), Physical Self-Perception Profile, Self Administered 7-Day Physical Activity Recall (PAR), and Leisure Time Exercise Questionnaire (LTEQ). Both physical activity measures were significantly related to the SPAS and all physical self-perceptions. Multiple regressions showed that only self-perceptions of conditioning significantly predicted PAR (R2 = .24) and LTEQ (R2 = .30). SPA did not add any unique variance in predicting activity, and no moderator effects were found for either PAR or LTEQ. Findings suggest that perception of physical conditioning is the dominant predictor of physical activity levels in young women, and social physique anxiety does not moderate this relationship. PMID- 11253321 TI - Using principal components analysis to identify individual differences in vertical jump performance. PMID- 11253322 TI - Inhibition of maximal voluntary isometric torque production by acute stretching is joint-angle specific. PMID- 11253323 TI - Effects of aging, grip span, and grip style on hand strength. PMID- 11253324 TI - Examining the structure of physical self-description using an American university sample. PMID- 11253325 TI - Role modeling attitudes and physical activity and fitness promoting behaviors of HPERD professionals and preprofessionals. PMID- 11253326 TI - [Chlamydia pneumoniae and myocardial infarction (CLINF). Preliminary randomized controlled study with clarithromycin. CLAINF protocol]. PMID- 11253327 TI - Ventricular arrhythmias and left ventricular hypertrophy in essential hypertension. AB - BACKGROUND: Patients with essential hypertension and/or left ventricular hypertrophy and ventricular arrhythmias suffer from an increased mortality rate. In all previous studies on hypertension, the criterion for inclusion was diastolic blood pressure > 95 mmHg. This is a low selective threshold. Our study attempted to evaluate the incidence of ventricular arrhythmia in hypertensive patients not receiving pharmacological treatment and diagnosed by 24-h ambulatory blood pressure monitoring (ABPM), therefore using a more selective criterion than WHO guidelines. METHODS: Hundred-twenty-height consecutive patients with hypertension diagnosed on the basis of WHO guidelines were screened for 24-h ambulatory blood pressure measurement. Eighty-five (66.4%) presented a 24-h mean blood pressure > 135/85 mmHg. All 85 patients were screened for M-mode, B-mode echocardiography, PW Doppler and 24-h ECG Holter recordings. RESULTS: Sixty patients (70.6%) were affected by left ventricular hypertrophy and 25 were free (29.4%). Thirty-six patients (42.4%) had left ventricular diastolic dysfunction, 49 were free (57.6%). According to Lown and Wolf's classification of ventricular arrhythmia, 20 patients (23.5%) presented Grade I arrhythmia, 5 (5.9%) presented Grade II, 4 (4.7%) Grade III, 9 (10.6%) Grade IVA, 20 (23.5%) Grade IVB, 12 (14.1%) Grade V and 15 patients (17.6%) were free from premature ventricular complexes, namely Grade 0 arrhythmia. Left ventricular hypertrophy was found to correlate significantly with the arrhythmia score, r = 0.552 for p < 0.0001. Moreover, left ventricular diastolic dysfunction correlated significantly with the arrhythmia score, r = 0.495 for p < 0.0001. There was also a good correlation between left ventricular hypertrophy and left ventricular diastolic dysfunction, r = 0.616 for p < 0.0001. Among patients affected by left ventricular diastolic dysfunction and left ventricular hypertrophy, the correlation with the arrhythmia score was even closer, r = 0.586 for p < 0.0007. CONCLUSIONS: We conclude that by using a more selective criterion for the diagnosis of hypertension, we can identify patients with a highly significant statistical correlation between left ventricular hypertrophy and ventricular arrhythmia score, and also between diastolic dysfunction and the ventricular arrhythmia score, due to a more severe stage of disease. It is useful to detect those patients affected by ventricular arrhythmias for the primary prevention of major cardiovascular events. PMID- 11253328 TI - Normothermic versus hypothermic perfusion during cardiopulmonary bypass. A randomized study on 132 patients. AB - BACKGROUND: A prospective randomized trial to compare normothermic CPB with hypothermic CPB has been performed. METHODS: 132 patients undergoing CPB were randomized into two groups: group 1 underwent normothermic CPB and group 2 hypothermic CPB (between 26 and 30 degrees C). RESULTS: Any significant difference was observed between the groups with regard to hospital mortality, blood transfusions, incidence of neurologic deficits and hematocrit, blood hemoglobin levels, platelet counts, plasma concentrations of glutamic-pyruvic transminase, glutamic-oxaloacetic transaminase, creatine kinase, valued at the 12th and 24th postoperative hour and at the 2nd, 3rd, and 4th postoperative day. A significant difference was observed between the groups with regard to tracheal extubation time, discharge time from the intensive care unit and inotropic drug infusion. The normothermic CPB patients group needed shorter time for tracheal extubation and discharge from the intensive care unit: this difference may be ascribed to a shorter inotropic drug infusion. Any increased surgical risks have been observed. CONCLUSIONS: In conclusion, we think that normothermic CPB is favourable because it can reduce costs, it can improve the management of a cardiac surgery unit and it is more comfortable for patients. PMID- 11253329 TI - Surgery of abdominal aorta in octogenarians. Can indications be extended? AB - BACKGROUND: This paper describes the authors' experience with the management of the abdominal aorta in patients aged over 80 years. METHODS: Ten urgent procedures were performed on patients older than 80 years during a 2 year period. In 4 cases surgery was performed because of a ruptured aneurysm of the subrenal abdominal aorta, in 2 cases for active symptomatic aneurysms, in 3 cases for severe lower limb ischemia (occlusion of the iliac and femoral arteries) and in 1 case for a secondary aortoenteric fistula. RESULTS: The operative mortality rate was 20% (2 cases with a ruptured aneurysm). Five patients are still alive in good health conditions (one of them had been operated twice for two different diseases). Even if our findings refer to a small number of patients, although similar series on emergency operations found in the literature are not substantially larger, the results do not advise against operative treatment of the abdominal aorta in cases requiring a direct approach, even in patients over 80 years of age. CONCLUSIONS: If this treatment strategy is obviously adopted in emergency conditions, as with the patients we are reporting on, since the alternative to operation is usually death, it should also be carefully considered in elective circumstances, where alternative treatments such as endovascular stents did not to date obtain better results. In the elective scenario all the necessary biological and physical parameters as well as the patient's age should be taken into proper account in deciding whether to operate. This is specially true now that the average life spans of an individual is longer so that patients, who may incur serious problems if left untreated, may be offered a better quality of life. PMID- 11253330 TI - Isolated femoral profundoplasty using endarterectomised superficial femoral artery for limb salvage in the elderly. AB - BACKGROUND: The deep femoral artery provides the primary blood supply to the thigh, and in addition serves as the major collateral channel for bypassing the obstructed superficial femoral artery. The purpose of isolated profundoplasty is to relieve a significant stenosis and improve perfusion of the ischaemic leg. METHODS: Twenty-seven patients with critical limb ischaemia underwent isolated profundoplasty in the Vascular Unit of Meir General Hospital, using endarterectomised superficial femoral artery (ESFA) as an arterial patch. Nineteen patients were men. The average age was 72 (65-79). The presenting symptoms: rest pain: 18 (67%), ischaemic foot ulcer: 7 (28%), pedal gangrene: 2 (7%). Selection criteria for isolated profundoplasty: 1) > 50%: stenosis of arteria profunda femoris lumen. 2) Adequate profunda: popliteal collateral system. 3) Adequate arterial inflow: common femoral artery. RESULTS: There was no operative mortality or immediate operative failure. All 27 limbs were improved: relief of rest pain, healing of ischaemic ulcers and good healing after minor amputations (transphalangeal, transmetatarsal). Follow-up period ranged from 12 to 45 months (mean 30 months) and was based on clinical investigation + ankle/brachial Doppler measurements. All patients remained asymptomatic with improvement of limb function--either to the present or until their death. CONCLUSIONS: In view of our favourable experience, we feel that isolated profundoplasty still has a place in vascular surgery practice--when limb revascularization in elderly patients considered at high risk is dangerous and when there is impossible below knee vascular reconstruction. We recommend the use of ESFA as a patch for long segment profundoplasty--with all advantages of an autogenous material. PMID- 11253331 TI - [A new questionnaire for assessing the quality of life of patients with intermittent claudication]. AB - BACKGROUND: Quality of life assessment is becoming increasingly relevant for evaluating the impact of disease and treatments and for deciding priorities when allocating resources. This is especially true in intermittent claudication where the goal of therapy is not the cure of the disease but rather to alleviate its symptoms and improve the patient's functional capabilities. At present, however, no generic scale fits all criteria for the ideal quality of life measuring in intermittent claudication. METHODS: We developed a questionnaire aimed at evaluating the specific limitations encountered by claudicants in the physical activity and in the social and emotional functioning. The present study evaluated the questionnaire for validity, reliability, and sensitivity to change, attributes considered to be essential for a questionnaire to be useful. RESULTS: In 30 patients with intermittent claudication, the scores of the four sections of the questionnaire significantly correlated with the scores of the corresponding sections of the Nottingham Health Profile. This indicates that the questionnaire is valid. For each of the four subscales, the intraclass correlation coefficient was > 0.75, thus showing a high test re-test reliability. Also the internal consistency is strong with alpha coefficient ranging from 0.79 to 0.89. Finally, the questionnaire was administered to 9 patients before and 4 weeks after percutaneous transluminal angioplasty for claudication. After the intervention, the improvement in walking performance paralleled the improvement in quality of life. This indicates that the questionnaire is sensitive to change. CONCLUSIONS: Our questionnaire appears to be a valid and reliable quality of life measure in intermittent claudication. PMID- 11253332 TI - [Ischemic cardiovascular diseases. Correlation with Helicobacter pylori infection]. AB - Coronary heart disease is the primary cause of mortality in western countries. The well-established ("classical") risk factors cannot fully explain epidemiological variations of this disease. From several years infections have been linked to ischemic vascular events and recent studies pointed to the role of Helicobacter pylori (H. pylori), a spiral Gram negative bacterium, that chronically infects human stomach and is involved in the pathogenesis of gastritis and peptic ulceration. Systematic reviews of studies have suggested the existence of a possible weakly positive association between this bacterium and coronary heart disease, but this could be due to confounding bias and influenced by the degree of investigations heterogeneity. Experiments from animal studies demonstrated that H. pylori infection in mice induces the formation of platelet aggregates and in contrast to Chlamydia pneumoniae it has not been found in the plaque: therefore, the role of H. pylori, could be even more important in the acute phase of myocardial infarction. There is the need for extensive prospective studies to evaluate the incidence of these diseases in relation to the presence of H. pylori infection. Appropriately randomized studies employing an antibiotic treatment for patients affected by ischemic vascular disease will answer the question of whether H. pylori has a causal role in the pathogenesis of acute myocardial infarction and ischemic stroke. PMID- 11253333 TI - [TNF alpha and heart failure]. AB - Tumor necrosis factor alpha (TNF alpha) is a cytokine with proinflammatory properties which produces negative inotropic effects on the heart. It is produced in a variety of conditions such as septic shock, acute myocarditis, reperfusion injury, and congestive hear failure (CHF). This production is probably due to activation of immune elements localized in the heart or periphery, or both. TNF alpha acts by binding to two specific receptors: TNF-R1 and TNF-R2. These two proteins have different effects. TNF-R1 has cytotoxic and antiviral activity, induces fibroblast proliferation, and mediates apoptosis. TNF-R2 is involved in septic shock and in lymphocyte proliferation. They both have negative inotropic effect on the heart. It has been showed that these receptors are down-regulated in congestive heart failure, while their soluble forms (sTNF-R1 and sTNF-R2) increase with the severity of symptoms. However the significance of this increase is still unclear. The role of Fas, a receptor protein that induces apoptosis, is also examined. Fas and its ligand have homologies respectively with TNF alpha and TNF-R. Also the soluble form of Fas (sFas) increases in relation to heart failure and is related to soluble forms of the similar receptor family, therefore it is possible that the same stimuli lead the three receptors to act together. SFas, as well as sTNF receptors, may play an important role in CHF. PMID- 11253334 TI - [Venous thromboembolism. Introduction]. AB - The main aim of the treatment of deep venous thrombosis (DVT) is to prevent the onset of the main complications: embolism (acute) and post-phlebothrombotic syndrome-PPS--(late complication). If not treated, during the acute phase DVT presents extension and/or embolism in 60% of cases and pulmonary embolism is potentially fatal in 5-10%. PPS is the most frequent complication (up to 70% of cases). The treatment of acute DVT has been based for over thirty years on heparin and oral anticoagulants: thrombolytic agents and low molecular weight (LMWH) heparins have been introduced more recently. Anticoagulants treatment is continued for 3-6 months (or longer in the event of recidivation or thrombophilia). LMWH have proved more effective and easier to manage than non fractioned heparin. The association of thrombolytic and heparin presents no advantages compared to the use of heparin alone. It is currently reserved for cases of venous gangrene and acute massive pulmonary embolism. The possibility of surgical embolectomy or the use of catheters should only be considered in treatment is ineffective or contraindicated. The aim of this paper is to analyse the treatment of DVT and the diagnostic, clinical, laboratory and instrumental procedures used, and to describe the most up-to-date indications for its diagnosis and treatment. PMID- 11253335 TI - [Cardiologic diagnosis of pulmonary embolism: echocardiography]. AB - Pulmonary embolism (PE) represents the third more frequent cardiovascular disease following the acute coronary artery disease and stroke. The most important predisposing clinical condition for PE is represented by the deep-vein thrombosis. The clinical diagnosis of PE has a very low accuracy; so the clinical suspect has to be necessarily directed towards the performance of diagnostic procedures. Among the most used procedures, the echocardiography has a diagnostic role but also a prognostic one. Moreover, it offers precious informations useful to perform the most suitable treatment. The echocardiography features which suggest the presence of pulmonary embolism are: right ventricle and atrium dilatation, right ventricular hypokinesia, systolic flattening of the interventricular septum, tricuspid regurgitation, pulmonary artery dilatation, disappearance or reduction of the inspiratory collapse of the inferior vena cava and presence of eventual embolic sources. According to the involvement degree of right ventricular function, it is generally possible to identify a different survival. The subgroup of patients with moderate or severe right ventricular dysfunction shows a high in hospital and within 1 year death rate. For this reason the right ventricular dysfunction degree together with the hemodynamic stability, are the most important parameters in the therapeutic choice. If there is no right ventricular dysfunction a treatment with heparin is indicated. In presence of right ventricular dysfunction and hemodynamic instability, the thrombolytic treatment is necessary. If the patient is hemodynamically stable, a transesophageal echocardiography is recommended; in case of central thrombosis the thrombolytic therapy or surgery are needed, while if no embolic material is shown the heparin treatment is advisable. PMID- 11253336 TI - [Ultrasonographic diagnosis of venous thrombosis: color Doppler and power Doppler]. AB - Eco color-Doppler (ECD) and Power Doppler (PD) currently allow an accurate and early diagnosis of deep venous thrombosis (DVT). "Ecocontrasts" boost sensitivity. The simple test of ultrasound probe compression shows excellent sensitivity in the proximal venous segments, comparable to a full ECD: the latter is preferable for distal segments. Ultrasound probe compression is not sufficient as the sole test when studying suspected leg DVT, but may form an integral part of a complete ECD study. The accuracy of ECD used in segments that are difficult to evaluate (iliac axes, inferior vena cava) has improved considerably thanks to PD. At these levels, it is certainly not possible to use ultrasound probe compression alone. The same is true for the diagnosis of recidivation, where a complete ECD is required to evaluate thrombotic overlap indicating true rethrombosis of partial or fully recanalised blood vessels. Phlebography no longer represents the gold standard for diagnosis compared to ultrasound techniques. Its use is limited to discriminating between genuinely doubtful and discordant cases of DVT and the search for occult embolic sources in patients with severe pulmonary embolism. The usefulness of an extensive study of venous vessels is gaining increasing confirmation, using the full potential of eco Duplex methods in the systematic search for thrombotic lesions. The possibility of obtaining as much information as possible during the first complete scan reduces the costs of the procedure and is more accurate for the diagnosis of suspected DVT recidivation. PMID- 11253337 TI - [Evidence-based guidelines for prevention and therapy of venous thromboembolism]. AB - Recently, evidence-based guidelines for the prevention and therapy of venous thromboembolism have been published. Prophylaxis: in General Surgery patients with moderate to severe risk need to be treated with unfractioned (UFH) or low molecular weight (LMWH) heparin. Non pharmacological methods must be reserved to patients with high hemorrhagic risk and in association to heparin to patients with particularly high thromboembolic risk. In high risk Ortopedic Surgery prophylaxis with high doses LMWH or oral anticoagulants (OA) is indicated. Il Neurosurgical Surgery and in politraumatized patients prophylaxis must be individualized taking account of hemorrhagic risk; patients with acute medullary lesion with paraplegia must be treated with LMWH. In Internal Medicine conditions which determine prolonged bed rest need prophylaxis with UFH or LMWH. In pregnancy, pharmacological prophylaxis is indicated only in cases of preceding thrombotic events or documented congenital risk factors. THERAPY: deep venous thrombosis or sub-massive pulmonary embolism must be treated with anticoagulant doses of UFH or LMWH (100 U antiXa/Kg twice daily). OA must be continued for a time identifiable on the basis of underlying disease. In massive or sub-massive pulmonary embolism with hemodynamic instability thrombolysis is indicated. In heparin induced thrombocytopenia alternative antithrombotic treatments must be employed. PMID- 11253338 TI - [Current role of the surgeon in the treatment of venous thromboembolism]. AB - The most frequent complications of deep venous thrombosis (DVT) are post thrombotic syndrome (PTS) and pulmonary embolism (PE); the main purpose of DVT therapy is to prevent their onset. A range of treatment is now available, including physical, medical and surgical forms. Physical: elastic compression mobilization postural therapy. Medical: anticoagulants (heparin, LMW heparins, oral anticoagulants), thrombolytic agents. Surgical: if DVT is diagnosed at an early stage, anticoagulant treatment may be accompanied by attempted surgical deobstruction, above all if DVT is localised at the popliteal and femoral confluents (greater risk of evolution towards severe "ischemic" forms potential cause of venous gangrene). These treatments include locoregional endogenous thrombolysis followed after phlebography, by surgical thrombectomy in the event of thrombotic residue, enabling the possible embolization of the pulmonary district using a caval filter. The combined medical and surgical approach reduces the long-term incidence of PTS. Temporary caval filters are also available. In short, a modern approach to the clinical problem of DVT now takes the form of early diagnosis and combined thrombolysis-surgery, which appears to be the most appropriate choice, ensuring the best form of venous functional recovery. However, this requires hospitalization in specialist units and multidisciplinary skills (hematological, medical, radiological and surgical) to ensure the best results. PMID- 11253340 TI - [Diagnosis of deep venous thrombosis]. AB - Deep venous thrombosis (DVT) is often difficult to diagnose based on objective data alone (sensitivity 60-66%, specificity 20-72%, according to a number of studies) and this leads to the possible misuse of instrumental tests. It is important to establish standard clinical protocols to select those cases that call for further diagnostic tests. The classic gold standard, phlebography, is invasive, expensive, not always available, and occasionally inadequate or risky. Non-invasive diagnostic methods (Doppler, plethysmography, eco-Doppler) is sufficiently sensitive for proximal venous occlusions, but not for distal occlusion. D-dimer assay is an accurate means of excluding the presence of DVT, but should be associated with instrumental diagnosis to increase its accuracy in asymptomatic patients. In order to optimize the use of clinical and anamnestic data, the authors propose measuring standard parameters using a scoring system. In cases with a score of > 3 (high risk) the frequency of DVT confirmed by phlebography is 75% in medium-risk patients the frequency is 17% low risk patients reveal a frequency of 3%. Patients with a medium- to high risk (overall score > 1) and positive Doppler for DVT, the diagnosis is reasonably certain and treatment may be started. In low-risk patients (score < or = 0) and negative Doppler, the diagnosis of DVT is highly improbable and no therapy is indicated. Discordant results between the clinical and Doppler tests underline the need for further instrumental tests. PMID- 11253339 TI - [Evolution in the pharmacological treatment of venous thrombosis according to evidence-based medicine]. AB - Today therapeutic protocols must be in accordance with Recommendations derived by Randomized Controlled Trials (RCT) Evidences. Deep Venous Thrombosis (DVT), post thrombotic syndrome and pulmonary embolism (PE) are different forms of the thromboembolic venous disease. The Authors, according with Evidence-Based Medicine, review the most significant RCT about Low-Molecular-Weight Heparin (LMWH). It has been proved that LMWH is more efficacious, easier to administrate and with less significant side effects than Unfractioned Heparin (UH) in DVT treatment. Its higher anti-Xa than anti-IIa activity provides higher anti thrombotic properties and lower haemorrhagic risk. LMWH does not require anticoagulant monitoring and allows outpatient--ambulatory care. RCT also showed lower PE ratio and lower haemorrhagic risk with LMWH outpatient care than with UH in-hospital care for DVT. RCT showed also a long-term lower DVT relapse and PE incidence with LMWH than with oral anticoagulants. The Authors report their own experience with LMWH and early ambulation for the treatment of DVT versus standard UH therapy. Their retrospective analysis confirms lower incidence of complications: growth of the thrombus, severe haemorrhages, PE. PMID- 11253341 TI - [Physical treatment of deep venous thrombosis: bed rest or mobilization?]. AB - The need of prolonged bed-rest for the treatment of Deep Venous Thrombosis (DVT), which was considered essential to control the thrombotic phenomenon and to prevent Pulmonary Embolism (PE) until ten years ago, has now been critically reviewed in the light of the great success of the Low Molecular Weight Heparin (LMWH) in medical therapy of DVT. There is a great evidence for bed-rest and immobility to play a pivotal role in the growth and in the progression of a venous thrombosis. The Authors emphasize, both on the international reports and their own experience, that, in most cases, medical treatment of DVT consists of an outpatient--ambulatory care based on immediate mobilization and ambulation, on external compression therapy, on early LMWH administration and late oral anticoagulation. This regimen provides great benefits in order to prevent PE, to improve the quality of life, to reduce the hospital and the anticoagulant monitoring charges. PMID- 11253342 TI - [Compression therapy in deep venous thrombosis]. AB - External compression, both intermittent by pneumatic pumps and continuous by anelastic or elastic bandages and by graduate compression stockings, play a pivotal role in prophylaxis of Deep Venous Thrombosis (DVT). The use of external compression in DVT therapy and in prophylaxis of pulmonary embolism (PE) and of post-thrombotic syndrome has not been validated as well as in DVT prophylaxis. The pathophysiologic properties of the external compression and the most recent evidences about the early mobilization of the patients with DVT and about Low Molecular Weight Heparin (LMWH) therapy suggest the advantages of the external compression. The authors review the most important clinical investigations about early use of external compression in DVT joined with pharmacological therapy: the results have been the reduction of the growth of the thrombus, the reduction of PE ratio, the prevention of the post-thrombotic syndrome, the indirect improvement of the quality of life. Finally the authors confirm the recommendations about the use of physical therapy with early mobilization and external compression joined with LMWH anticoagulation in DVT. PMID- 11253344 TI - [Superficial thrombophlebitis]. AB - Thrombophlebitis of the superficial veins (SVT) of the leg is usually regarded as a mild and uncomplicated disease. Although this is generally true for acute thrombosis of the branches of the saphenous vein, the natural history of SVT involving the main trunk may not be as benign. The association of SVT with deep venous thrombosis (DVT) has been reported to range from 17 to 40%; the progression of the thrombotic process from the greater saphenous vein into the deep venous system has been reported in 8.6% of the cases. For this reason, even if symptoms of DVT are lacking, it is necessary to use duplex ultrasonography to be certain that DVT does not exist concurrently with SVT. In a recent study we found that saphenous-vein thrombi embolize even when no femoral-vein involvement is evident. Of 21 patients included in the study, findings compatible with a high probability of pulmonary embolism were detected in 7 (33.3%, 95% CI, 14.6 to 57.0), although clinical symptoms were present only in 1. The risk of pulmonary embolism is similarly high in patients with and without thrombosis at the sapheno femoral junction. These patients presumably would benefit from anticoagulation, but such a benefit remains to be proven. Superficial thrombophlebitis, in the absence of DVT proven by duplex ultrasonography, is generally treated with nonsteroidal anti-inflammatory agents. A prospective randomized study is being carried out at our Institution evaluating therapeutic doses of anticoagulant drugs in SVT. Interim report suggests that, in thrombophlebitis of the thigh, high fixed doses of unfractioned heparin are more effective than low doses for the prevention of early and late venous thromboembolic complications and are not associated with an appreciable bleeding risk. PMID- 11253343 TI - [State of the art: low-molecular-weight heparin and beyond]. AB - Approximately 20 years ago a new family of antithrombotic compounds started to be investigated: the low-molecular weight heparins (LMWH). The rationale for their use was based on the evidence that the inhibition of the Factor Xa of blood coagulation was less marked than that of Factor IIa when using the LMWHs as compared to unfractioned heparin (HF). This particular mechanism of action was considered to be of advantage regarding the safety profile (the pro-haemorrhagic effect) compared to HF. Today we know that the real advantage of LMWHs is due to their high bioavailability which makes safe and reliable their subcutaneous administration without laboratory monitoring. The LMWHs are equally effective than HF for the treatment of acute Deep Vein Thrombosis. For the prophylaxis of Venous Thromboembolism, LMWHs are indicated as first choice in high-risk patients such as those undergoing major orthopaedic surgery. The future development of this family of drugs encompasses the launch of the pentasaccharide which is a pure anti-/Xa inhibitor. PMID- 11253345 TI - Caveat emptor. PMID- 11253346 TI - Keeping your patients informed. PMID- 11253347 TI - Professional integrity in publishing. PMID- 11253348 TI - Oral infections and CVD. PMID- 11253349 TI - Prosthodontics and esthetics. PMID- 11253350 TI - The ethics of water fluoridation. PMID- 11253351 TI - The periodontal disease classification system of the American Academy of Periodontology--an update. AB - Until recently, the accepted standard for the classification of periodontal diseases was the one agreed upon at the 1989 World Workshop in Clinical Periodontics. This classification system, however, had its weaknesses. In particular, some criteria for diagnosis were unclear, disease categories overlapped, and patients did not always fit into any one category. Also, too much emphasis was placed on the age of disease onset and rate of progression, which are often difficult to determine. Finally, no classification for diseases limited to the gingiva existed. In 1999, an International Workshop for a Classification of Periodontal Diseases and Conditions was organized by the American Academy of Periodontology to address these concerns and to revise the classification system. The workshop proceedings have been published in the Annals of Periodontology. The major changes to the 1989 proceedings and the rationale for these changes are summarized here. In addition, the potential impact of these changes is discussed. PMID- 11253352 TI - Oral and maxillofacial manifestations of multiple sclerosis. AB - Multiple sclerosis is a chronic demyelinating disease of the central nervous system which mostly affects young adults living in the northern hemisphere. It is a disease primarily found in temperate climates, being rare in the tropics and increasing in frequency with distance from the equator. Canada has one of the highest prevalence rates in the world. Dentists should be familiar with the clinical manifestations that affect the oral and maxillofacial areas as well as patients' general health. Three of the most frequent oro-facial symptoms include trigeminal neuralgia, trigeminal sensory neuropathy and facial palsy. Dentists should also be aware of the importance of this disease in the diagnosis, treatment and prognosis of certain oro-facial lesions or conditions. This paper reviews 2 cases of multiple sclerosis, highlights its oro-facial manifestations and discusses the dental implications of the disease. PMID- 11253353 TI - Unconventional dentistry: Part V. Professional issues, concerns and uses. AB - This is the last in a series of 5 articles providing a contemporary overview and introduction to unconventional dentistry (UD) and its correlation to unconventional medicine (UM). UD and UM both present important concerns for health care professionals and for the general public. Professional concerns include risks to the practitioner and the patient. UD is of special concern because of the potential harm of invasive dental procedures. Nonetheless, because some UD practices may be of benefit to the patient, decision-making issues and guidelines for UD practice are suggested. PMID- 11253354 TI - Restoration of endodontically treated teeth with carbon fibre posts--a prospective study. AB - BACKGROUND: A prospective study was started in 1995 to evaluate the success of carbon fibre reinforced epoxy resin (CFRR) posts used to restore endodontically treated teeth. All the teeth in the study had lost more than 50% of their coronal structure. METHODS: Fifty-nine carbon fibre Composiposts cemented with Metabond and built up with Core Paste cores were placed into the teeth of 47 patients. Each tooth received a full-coverage restoration (porcelain fused to metal crown) and was followed for 6.7-45.4 months (average = 28.0 months, standard deviation = 10.7). RESULTS: Results for 52 teeth in 42 patients were analyzed. There were no fractures. The overall failure rate was 7.7% and the cumulative survival rate was 89.6% at the end of the follow-up period. The only statistically significant finding (p = 0.04) was that posts in lower premolars were at higher risk of failure. CONCLUSION: CFRR posts are among the most predictable systems available today. CFRR posts in the upper anterior teeth are associated with a higher success rate and longer life than those placed in premolars, especially lower premolars. This study contributes to the growing body of evidence that supports the use of CFRR posts in the restoration of endodontically treated teeth. PMID- 11253355 TI - Interneurons unbound. AB - Local-circuit, gamma-aminobutyric acid-releasing inhibitory interneurons of the hippocampus and cortex have traditionally been considered as the regulators of principal neuron activity--the yin to the excitatory yang. Recent evidence indicates that, in addition to that role, their network connectivity and the properties of their intrinsic voltage-gated currents are finely tuned to permit inhibitory interneurons to generate and control the rhythmic output of large populations of both principal cells and other populations of inhibitory interneurons. This review brings together recently described properties and emerging principles of interneuron function that indicate a much more complex role for these cells than just providers of inhibition. PMID- 11253356 TI - Neurotrophins as synaptic modulators. AB - The role of neurotrophins as regulatory factors that mediate the differentiation and survival of neurons has been well described. More recent evidence indicates that neurotrophins may also act as synaptic modulators. Here, I review the evidence that synaptic activity regulates the synthesis, secretion and action of neurotrophins, which can in turn induce immediate changes in synaptic efficacy and morphology. By this account, neurotrophins may participate in activity dependent synaptic plasticity, linking synaptic activity with long-term functional and structural modification of synaptic connections. PMID- 11253357 TI - Neural basis of deciding, choosing and acting. AB - The ability and opportunity to make decisions and carry out effective actions in pursuit of goals is central to intelligent life. Recent research has provided significant new insights into how the brain arrives at decisions, makes choices, and produces and evaluates the consequences of actions. In fact, by monitoring or manipulating specific neurons, certain choices can now be predicted or manipulated. PMID- 11253358 TI - The splice of life: alternative splicing and neurological disease. AB - Splicing of pre-messenger RNA is regulated differently in the brain compared with other tissues. Recognition of aberrations in splicing events that are associated with neurological disease has contributed to our understanding of disease pathogenesis in some cases. Neuron-specific proteins involved in RNA splicing and metabolism are also affected in several neurological disorders. These findings have begun to bridge what we know about the mechanisms regulating neuron-specific splicing and our understanding of neural function and disease. PMID- 11253359 TI - Recognition memory: what are the roles of the perirhinal cortex and hippocampus? AB - The hallmark of medial temporal lobe amnesia is a loss of episodic memory such that patients fail to remember new events that are set in an autobiographical context (an episode). A further symptom is a loss of recognition memory. The relationship between these two features has recently become contentious. Here, we focus on the central issue in this dispute--the relative contributions of the hippocampus and the perirhinal cortex to recognition memory. A resolution is vital not only for uncovering the neural substrates of these key aspects of memory, but also for understanding the processes disrupted in medial temporal lobe amnesia and the validity of animal models of this syndrome. PMID- 11253360 TI - Expressing what's on your mind: DNA arrays and the brain. AB - Questions about brain function and disease are being addressed with parallel genomic approaches. High-density DNA arrays make it possible to monitor the expression levels of thousands of genes at a time, and are being used to address old questions in new ways and to generate new hypotheses about the workings of the brain. PMID- 11253361 TI - Interpretations of retrograde amnesia: old problems redux. AB - Recent evidence indicates that an old memory reactivated by cueing becomes labile and vulnerable to an amnesic treatment. Although the 'reconsolidation' concept derived from these findings challenges the traditional consolidation theory, here we argue that the new concept suffers from some of the same limitations as the earlier model. We propose an alternative retrieval-based theory that accommodates the recent data, as well as other puzzling related observations. PMID- 11253362 TI - The expanding polymerase universe. AB - Over the past year, the number of known prokaryotic and eukaryotic DNA polymerases has exploded. Many of these newly discovered enzymes copy aberrant bases in the DNA template over which 'respectable' polymerases fear to tread. The next step is to unravel their functions, which are thought to range from error prone copying of DNA lesions, somatic hypermutation and avoidance of skin cancer, to restarting stalled replication forks and repairing double-stranded DNA breaks. PMID- 11253363 TI - Secrets of actin-based motility revealed by a bacterial pathogen. AB - Actin-based cell motility is a complex process involving a dynamic, self organizing cellular system. Experimental problems initially limited our understanding of this type of motility, but the use of a model system derived from a bacterial pathogen has led to a breakthrough. Now, all the molecular components necessary for dynamic actin self-organization and motility have been identified, setting the stage for future mechanistic studies. PMID- 11253364 TI - Apoptosis in neurodegenerative disorders. AB - Neuronal death underlies the symptoms of many human neurological disorders, including Alzheimer's, Parkinson's and Huntington's diseases, stroke, and amyotrophic lateral sclerosis. The identification of specific genetic and environmental factors responsible for these diseases has bolstered evidence for a shared pathway of neuronal death--apoptosis--involving oxidative stress, perturbed calcium homeostasis, mitochondrial dysfunction and activation of cysteine proteases called caspases. These death cascades are counteracted by survival signals, which suppress oxyradicals and stabilize calcium homeostasis and mitochondrial function. With the identification of mechanisms that either promote or prevent neuronal apoptosis come new approaches for preventing and treating neurodegenerative disorders. PMID- 11253365 TI - Grabbing the cat by the tail: manipulating molecules one by one. AB - Methods for manipulating single molecules are yielding new information about both the forces that hold biomolecules together and the mechanics of molecular motors. We describe here the physical principles behind these methods, and discuss their capabilities and current limitations. PMID- 11253366 TI - Nuclear compartmentalization and gene activity. AB - The regulated expression of genes during development and differentiation is influenced by the availability of regulatory proteins and accessibility of the DNA to the transcriptional apparatus. There is growing evidence that the transcriptional activity of genes is influenced by nuclear organization, which itself changes during differentiation. How do these changes in nuclear organization help to establish specific patterns of gene expression? PMID- 11253367 TI - The meteoric rise of regulated intracellular proteolysis. AB - It is often the case in biology that research into breaking things down lags behind research into synthesizing them, and this is certainly true for intracellular proteolysis. Now that we recognize that intracellular proteolysis, triggered by attaching multiple copies of a small protein called ubiquitin to target proteins, is fundamental to life, it is hard to believe that 20 years ago this field was little more than a backwater of biochemistry studied by a handful of laboratories. Among the few were Avram Hershko, Aaron Ciechanover and Alexander Varshavsky, who were recently awarded the Albert Lasker award for basic medical research for discovering the importance of protein degradation in cellular physiology. This Timeline traces how they and their collaborators triggered the rapid movement of ubiquitin-mediated proteolysis to centre stage. PMID- 11253368 TI - Biological machines: from mills to molecules. AB - Although scientific progress is usually represented as being linear, it may, in fact, have a cyclical character--some discoveries may be forgotten or lost (at least temporarily), and themes may reappear through the centuries. Consider, for example, the concept of 'molecular machines', from the exciting phase of research that flourished in the seventeenth century, to the idea of machines that is at centre stage in modern cell biology. PMID- 11253369 TI - Slow axonal transport: stop and go traffic in the axon. AB - Efforts to observe the slow axonal transport of cytoskeletal polymers during the past decade have yielded conflicting results, and this has generated considerable controversy. The movement of neurofilaments has now been seen, and it is rapid, infrequent and highly asynchronous. This motile behaviour could explain why slow axonal transport has eluded observation for so long. PMID- 11253370 TI - Cadherins in embryonic and neural morphogenesis. AB - Cadherins not only maintain the structural integrity of cells and tissues but also control a wide array of cellular behaviours. They are instrumental for cell and tissue polarization, and they regulate cell movements such as cell sorting, cell migration and cell rearrangements. Cadherins may also contribute to neurite outgrowth and pathfinding, and to synaptic specificity and modulation in the central nervous system. PMID- 11253371 TI - National survey identifies gender differences in rosacea. PMID- 11253372 TI - Mental health problems may go undetected in men, nonwhites. PMID- 11253373 TI - Gender difference demonstrated in HIV viral load: researchers call for new treatment guidelines. PMID- 11253374 TI - African and Asian women may be more susceptible to HIV-1. PMID- 11253375 TI - The changing demographics of AIDS. PMID- 11253376 TI - The real Fourth of July story. PMID- 11253377 TI - The road ahead in women's health research. PMID- 11253378 TI - Estrogens for heart disease: a report from the annual session of the American College of Cardiology. PMID- 11253379 TI - Q & A with the expert on cardiovascular disease. PMID- 11253380 TI - Lateralized cortical perfusion in women with Alzheimer's disease. AB - OBJECTIVE: To define the pathophysiologic substrate of gender differences in cognition and behavior in Alzheimer's disease (AD). DESIGN: Hemispheric regional cerebral blood flow (rCBF) was studied in a consecutive series of dementia patients using single photon emission computed tomography (SPECT). PARTICIPANTS: Subjects included 300 outpatients who were studied with SPECT as part of a diagnostic evaluation for degenerative dementia or memory disorder. METHOD: Based on qualitative descriptions by a radiologist, subjects were classified as having unilateral left, unilateral right, bilateral, or no perfusion defects. Semiquantitative analysis of SPECT images was also performed using region of interest radionuclide counts normalized to the cerebellum. RESULTS: Among 174 females and 126 males, unilateral left hemisphere defects were found more commonly in women than men (24% vs 10%; chi 2 = 9.4; P = .009). This observation was most significant for the 103 females and 62 males regarded as having probable AD (26% vs 8%; chi 2 = 9.3; P = .01). In a multiple regression model of clinical variables, shorter duration of disease and female gender were significant independent predictors of the unilateral left hemisphere pattern among those with probable AD. Age, family history, education, handedness, and severity of cognitive impairment were not significant contributors. CONCLUSIONS: Women with AD exhibit greater heterogeneity in rCBF than men. Asymmetry in rCBF occurs more often in women. In some cases, this is related to relative preservation of right hemisphere function in women. PMID- 11253381 TI - Gender differences in neurotoxicity of the nigrostriatal dopaminergic system: implications for Parkinson's disease. AB - This article describes the progression of steps followed to demonstrate a gender difference associated with Parkinson's disease (PD) and to gain an understanding of the basis, mechanisms, and implications of this gender specificity. First, a review of the literature on PD shows a greater incidence in men. Next, data are presented from a series of laboratory studies in animal models of PD that suggest a basis for this gender difference: estrogen appears to act as a neuroprotectant of the striatal dopaminergic system. One mechanism for this effect may be that estrogen inhibits the uptake of neurotoxins capable of producing degeneration within dopaminergic neurons. Finally, some of the potential neurologic implications of manipulating estrogen in premenopausal and postmenopausal women are considered. PMID- 11253382 TI - Atrial fibrillation: are there gender differences? AB - The incidence of atrial fibrillation is greater in men than in women, but this gap closes with advancing age. More women with atrial fibrillation have underlying valvular disease, and more men with this condition have underlying coronary artery disease. Atrial fibrillation increases mortality and the incidence of stroke in both sexes. However, women in particular (especially those over 75 years old) may be at increased risk for embolism and long-term mortality. Gender is also an important feature affecting the selection of antiarrhythmic drugs for atrial fibrillation, because women are more likely to develop drug induced arrhythmias. Stroke prevention with anticoagulation in chronic atrial fibrillation is a priority in both men and women; however, women derive the most benefit from it. PMID- 11253383 TI - Gender differences in chronic headache in a treatment-seeking population. AB - OBJECTIVE: To evaluate gender differences in headache description, frequency, disability, and psychological distress in a treatment-seeking population. DESIGN: Consecutive treatment-seeking headache patients were questioned about headache characteristics and comorbid psychological distress. SUBJECTS: 37 males and 90 females seeking treatment at a university headache clinic. METHOD: Subjects were evaluated by a neurologist and assigned headache diagnoses according to International Headache Society criteria. Subjects also completed several psychological self-report measures. Gender comparisons were made for both headache characteristics and psychological measures using the Student t test and chi-square test. RESULTS: Headache characteristics and comorbid psychological symptoms were similar in men and women. Men were more likely than women to report headache-related disability. Headache triggers were similar in men and women, although exercise was reported as a more frequent trigger in men, and women were more likely to identify foods as usual headache triggers. Men and women reported similar responsivity to treatment. CONCLUSIONS: Treatment recommendations for chronic headache should be similar for men and women. However, evaluation of disability and exercise as headache triggers should be actively sought in male patients, while women should be educated about dietary adjustments and avoidance of eating irregularities. PMID- 11253384 TI - The effects of sexual abuse on body image, self-image, and sexual activity of women. AB - OBJECTIVES: To evaluate the differences between females who had been sexually abused and those who had not been sexually abused in body image, self-image, self consciousness, and relationships; to determine the effect of sexual abuse on sexual intimacy and behavior; and to identify consequences of sexual abuse. METHODS: A sample of 1664 females (832 sexually abused and 832 not sexually abused) who responded to a survey in Shape magazine were strictly matched on age and body mass index. RESULTS: Females who had been sexually abused reported more body dissatisfaction and self-consciousness, less satisfaction with themselves and in relationships, and less comfort with having sex with the lights on and undressing in front of their sexual partner than females who had not been sexually abused. Additional consequences of sexual abuse included lack of control over the body, eating disorders, and sexual identity confusion. Females who had been sexually abused were less likely to use contraception regularly than those without a history of sexual abuse. DISCUSSION: Results are presented with respect to limitations of the study and suggestions for treatment interventions. CONCLUSIONS: The impact of sexual abuse permeates all areas of survivors' lives. Clinicians have the opportunity to help their clients respond to the trauma of abuse. PMID- 11253386 TI - [Reaction time to visual stimuli in child development]. AB - The simple and differential reaction time and time of cognitive processes were studied in 3-7-year-old children using age-adapted computer technique. The reaction time significantly decreased with age in parallel with improvement of cognitive processes. An experimental method is proposed, which makes it possible to determine what kind of cognitive process is responsible for age-related decrease in the reaction time. PMID- 11253387 TI - [Saccade latency during probability presentation of the visual targets in man]. AB - The latent periods of saccadic eye movements in response to peripheral visual stimuli were measured in 8 right-handed healthy subjects using Posner's paradigm "COST-BENEFIT". In 6 subjects, the saccade latency in response to visual target presented in expected location in valid condition was shorter than that in neutral condition ("benefit"). Increase in saccade latency in response to the visual target presented in unexpected location in valid condition versus neutral condition took place only in 4 subjects ("cost"). A decrease in left-directed saccade latency in response to expected target presented in the left hemifield and increase in saccade latency in response to unexpected left target in comparison with analogous right-directed saccades were observed in valid condition. This phenomenon can be explained by the dominance of the right hemisphere in the processes of spatial orientation and "disengage" of attention. PMID- 11253388 TI - [Increasing selectivity of defense behavior during development of pied flycatcher nestlings]. AB - The development of defense reaction was studied in the wildlife and experimentally in 7 broods of altricial pied flycatcher (Ficedula hypoleuca) nestlings. Field studies demonstrated that passive-defense response first appeared on the 4th day of the nest life. It developed from the cessation of begging in young relatively satiated nestlings to characteristic freezing response independent of the level of feeding motivation in older nestlings. Older nestlings also acquire the defense reaction in response to novel visual stimuli. The efficiency of the natural stimulus for defense behavior (species-specific alarm call) nongradually changes during the nest life attaining the 100% level only on the 11th posthatching day. During the initial phase of defense behavior development, the reaction can be induced by different rhythmically organized stimuli. Later it becomes considerably more selective and other rhythmic and acoustic signals become much less effective than the alarm call. PMID- 11253389 TI - [Motor activity and emotional response in the open field test in rats after pharmacologic stimulation or blockade of neuropeptides in terminals of primary sensory neurons]. AB - Effects of high and low doses of capsaicin on the open-field behavioral patterns were examined in Wistar rats. The treated animals exhibited a significant increase in locomotion, grooming, and exploratory activity. PMID- 11253390 TI - [Dose-response dependence of the excitatory effect of acetylcholine on Helix lucorum neurons after orthodromic tetanization]. AB - Hill numbers before and after tetanic stimulation were calculated from dose response dependence between the amplitude of the acetylcholine-induced inward current and the amplitude of the iontophoretic current through a micropipette filled with acetylcholine. Semi-intact Helix lucorum preparation was used. Acetylcholine-induced inward currents were recorded using two-electrode voltage clamp technique. Tetanic stimulation evoked changes in dose-response dependence but did not modify the slopes of dose-response plots (Hill numbers were 1.42 + 0.15 before and 1.41 + 0.15 after tetanization). It was concluded that increase in cholinosensitivity in LPa3 and RPa3 neurons after the orthodromic tetanic stimulation of nervus intestinalis is not accompanied by changes in the number of ligand-binding sites per acetylcholine receptor molecule or proportion of nicotinic and muscarinic cholinoreceptors. PMID- 11253391 TI - [Research in psychophysiology at the Saint Retersburg State University]. PMID- 11253392 TI - [Effect of substance P on ethanol consumption in subchronically alcoholized rats in the limited scheduled access paradigm]. AB - The effect of substance P on alcohol intake was studied in rats using a limited scheduled access paradigm. Ascending doses of substance P (100 and 200 micrograms/kg) were administered intraperitoneally to rats approximately 30 minute prior to their 1-hour-per-day access to alcohol. Each dose was administered for 3 successive days, and the effect of substance P was compared to that of saline solution control. Substance P at the dose of 100 micrograms/kg had no effect on alcohol consumption but significantly decreased the alcohol intake at the dose of 200 micrograms/kg. Thus, substance P reduces the alcohol motivation of rats in a limited scheduled access paradigm. PMID- 11253393 TI - [Motor skill recovery in rats with various forelimb preference subjected to lesions in the caudate nucleus]. AB - Rats were trained for retrieving a sunflower seed from a tube using a forelimb. In the process of training, all the animals were divided in two groups: with the right- and left-limb preference. The period of learning was divided in two emotional (after the period of anxiety the animal takes a sunflower seed from the flow of a chamber) and three motor stages (different degrees of dexterity and precision in reaching a seed in the tube and grasping it). After a rat learned the skilled movement, the caudate nucleus was lesioned contralaterally to the preferred limb. After surgery, the animals were tested for recovery of the motor skill. The stages of recovery were the same as during learning. "Right-" and "left-handed" rats were compared in the percentage of animals performing each stage, duration of each stage, the depth of the tube reached by the animal's forelimb, and time required for obtaining the best results. The "right-handed" rats dominated in the percentage of animals performing the emotional stages and later began to recover the skilled movement (the crucial motor stage) but faster acquired their individual maximal depth of reaching a seed in the tube than the "left-handed" animals. The recovery of the skilled movement in the "left-handed" animals was a more gradual process than that in the "right-handed" ones. PMID- 11253395 TI - [Resonance phenomena to rhythmic light stimulation with various intensity and frequency in human EEG]. AB - The photoinduced resonance EEG response in the occipital area (O1 and O2) of right-handed men during 12-s intermittent photic stimulation was studied as a function of flash frequency (6, 10, or 16 Hz) and intensity (5 levels from 0.05 to 0.7 J). The EEG power in the narrow band coinciding with stimulation frequency was FFT-extracted in 3-s intervals before, during, and after each stimulation. It was found that increase in flash intensity was accompanied by an enhancement of the resonance EEG response and decrease in time of reaching its maximal value. These changes were to a greater extent characteristic for the right hemisphere. The low-intensity stimulation induced more pronounced resonance effects in the left hemisphere, whereas the high-intensity flashes to a greater extent involved the right hemisphere. The asymmetry of the EEG response to stimulation of the middle intensity was slight, and the time of reaching the maximal level of the resonance activation was about 6-8 s. A relatively high level of the resonance EEG response was observed during stimulation with the frequency of 10 Hz, even in case of its minimal intensity. The most pronounced resonance EEG response was induced in the right occipital area by the high-intensity 16-Hz stimulation. The enhanced sensitivity of the right hemisphere to intensity of flashes is interpreted as an indication of interhemispheric differences in nonspecific adaptive mechanisms of the brain. PMID- 11253396 TI - [Hemispheric organization of verbal memory functions in seasonal winter depression: electrophysiological analysis]. AB - Spatial organization of EEG power and coherence during memorization of dichotically presented lists of words were studied in patients with winter depression (N = 17) and control subjects (N = 22). In contrast to the control subjects, the depressed patients were characterized by the higher theta power in the right parietal and posterior temporal regions and the dominance of the alpha 2 in the left midfrontal area. The patients also differed in the lower theta 2 coherence in the left hemisphere and lower alpha 1 coherence in the right hemisphere. These effects showed different intrahemispheric distribution. The interhemispheric EEG coherence in the theta 2 range between the frontal areas and alpha 1 coherence between the left frontal and right posterior areas was lower in the patients than in the control subjects. Verbal-emotional interaction in depressions are discussed. PMID- 11253398 TI - [Natural coordination of the head and limb movements and its transformation by learning in dogs]. AB - Kinematic analysis of the head and the forelimb movements in dogs has been done during elaboration the instrumental tonic forelimb flexion when the head was bent down to foodwell. It was found, that in naive dogs the forelimb flexion was accompanied by anticipatory lifting of the head, the head lowering evoked extension of the flexed forelimb. Therefore simultaneous holding of the lifted limb and the lowered head was impossible and could be achieved only by learning. Studying of the dynamics of transformation of the innate (natural) head-forelimb coordination during learning has shown that innate relationship between phasic head-forelimb movements, which was lost at the early stage could spontaneously restore for a short time in trained dogs. It was found between low-amplitude head forelimb oscillations which did not disturb the learnt tonic forelimb flexion, when the head was bent down. The innate coordination is supposed to be an inborn and in the given conditions the only possible way of the forelimb lifting, in which the anticipatory lifting of the head might facilitate the limb flexion. That's why lowering of the head provoked extension of the flexed limb. Contrary to the known hypothesis [4] that the mechanism of elaboration of the novel coordination is connected with suppression of the interfering innate coordination, it is proposed to consider the elaborated coordination as the novel way of the forelimb lifting in the forced posture of the lowered head. The novel flexion of the forelimb, as supposed, became possible by changing its innate organization (muscular pattern). PMID- 11253399 TI - [Physiology of the higher nervous activity at the frontier of centuries. Report at the conference "Actual problems of biology" dedicated to the 70th anniversary of the Biology Department of the M. V. Lomonosov Moscow State University]. PMID- 11253397 TI - [Various analogues of the second-signal behavior in monkeys during their symbolic "description" of properties of planimetric figures]. AB - An attempt was made of experimental simulation of some components of the "second signal" (verbal) behavior in rhesus monkeys (Macaca mulatta) under conditions of their "dialog" with a computer. Three monkeys were trained to perform manipulative actions on keyboard simulator to "describe" the properties of planimetric figures. For the period of research, the monkeys have learned to reproduce combinations of two symbols ("bigrams") that were used by them for a "description" up to 12 geometrical figures of various sizes. The data obtained indicate that rhesus monkeys are able not only to generalize the properties of a certain class of visual images at the perceptual level but also to use symbols for the description of these generalized properties. PMID- 11253400 TI - [Effect of isatin on the electrical activity in the rat hippocampus cells]. AB - In vitro superfusion of rat hippocampal slices with isatin changed the population spikes. Isatin perfusion produced two clear effects. 50 microM isatin it increased the amplitude of the population spike in the CA1 evoked by stimulation of stratum radiatum. This effect was readily reversible. 100 microM isatin decreased the population spike amplitude with minimal effect on its latency. High initial response were more suppressed. This effect on the population spike amplitude was not eliminated even after 1 h of washing with saline. The data obtained suggest that isatin-induced electrophysiological changes are involved into the anticonvulsant effect of isatin. PMID- 11253401 TI - [Effects of sex factors and hemispheric localization on the involvement of serotonin from the frontal cortex, striatum, and nucleus accumbens into the processing of novel and repeatedly presented information in rats]. AB - The effects of novel or relevant (a single exposure to experimental chamber) and irrelevant (20 exposures to experimental chamber) stimuli on the levels of serotonin (5-HT) and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) in the frontal cortex, striatum, and nucleus accumbens in the left and right hemispheres were studied in male and female rats. It was found that 5-HT and 5-HIAA contents in the frontal cortex changed in response to neither relevant nor irrelevant stimuli. However, there were hemispheric difference in 5-HT and 5-HIAA in the frontal cortex of intact animals. The level of 5-HT in males and the level of 5 HIAA in females were higher in the left frontal cortex. In females, the level of 5-HIAA in the left striatum decreased in response to the novel stimulus. Sex differences in: a) 5-HT metabolism (increase in the level of 5-HIAA in males and increase in 5-HT in females) and b) lateralization (the striatal 5-HT metabolism in males changed bilaterally and only in the left hemisphere in females) were observed in reactions to irrelevant stimuli. Both in male and female rats, serotonin content in the nucleus accumbens changed only in response to the irrelevant stimuli. The 5-HT level increased in the left and right hemispheres independently of sex, but hemispheric difference was revealed only in females, in which the serotonin level was higher in the left nucleus accumbens. It is concluded that serotonergic neurotransmitter mechanisms are involved in hemispheric and sex differences in selective attention. PMID- 11253402 TI - [Post-tetanic modification of the efficacy of excitatory transmission in neuronal networks with interhemispheric connections]. AB - The modifiable reciprocal transcallosal monosynaptic excitatory connections were for the first time detected in vivo experiments in rat motor cortex using multiunit recording and crosscorrelation analysis, It was shown that high frequency microstimulation (MCS) of a small group of cortical cells of one hemisphere produces long-term changes in the efficacy of transcallosal excitatory connections, and also ipsilateral connections in both hemispheres. The posttetanic changes appear as long-term potentiation (LTP) and long-term depression (LTD). The bursting neurons were found to have more favorable conditions for the induction of LTP of most converging inputs (in contrast to cells with other discharge patterns). Both LTP and LTD could be simultaneously induced in synapses formed by axon collaterals of a callosal cell on several neurons. LTP and LTD could be simultaneously obtained at diverse synapses of the same cell. The number of spontaneously active callosal neurons as well as the number and efficacy of transcallosal connections increased after the MCS, whereas the number and efficacy of ipsilateral connections decreased. Basing on these data we assume that the ipsilateral inhibition is more effective than the transcallosal inhibition. MCS results in the modification of the pattern of initially existing connections between numerous neurons of an ensemble including cells of both hemispheres. PMID- 11253403 TI - [Effect of antibodies against S100 proteins on neural plasticity in sensitized and naive snails]. AB - The influence of antibodies against total S100 protein fraction (AB-S100) and S100b protein (AB-S100b) on the activity of LP11 and RP11 neurons were studied in naive snails and during the nociceptive sensitization. Application of AB-S100 or AB-S100b (0.1 mg/ml) initiated membrane depolarization, increase in its excitability, and depression of neural responses to sensory stimulation in nonsensitized snails. The sensitization produced facilitation of neural transmission and increase in membrane excitability. Exposure to AB-S100 or AB S100b (0.1 mg/ml) during sensitization substantially reduced its effects on neural transmission and membrane excitability. The difference between the extent of synaptic facilitation in neurons of sensitized snails and neurons of snails sensitized under conditions of AB-S100 or AB-S100b application was comparable with synaptic depression in neurons of naive snails produced by the isolated application of AB-S100 or AB-S100b. Application of AB-S100 of AB-S100b in the dose of 0.01 mg/ml did not change the parameters of neural activity. The obtained evidence suggest that S100 proteins (in particular, S100b) in L-RP11 neurons are involved in the mechanisms of membrane excitability, regulation of membrane potential and synaptic transmission in naive snails and in the mechanisms of membrane plasticity in the neurons during development of nociceptive sensitization. PMID- 11253404 TI - [Formation of C/EBP transcription factors and possible ways of regulation of their activity during learning in Helix]. AB - Defensive conditioning in Helix is associated with activation of C/EBP family of transcription factors. The incubation of the CNS with serotonin that imitates the effects of conditioning or stimulation of adenylate cyclase with forscolin also increase the DNA-binding activity of C/EBP. The induction of protein kinase C by phorbol ester does not stimulate the formation of these transcription complexes. However, a slight increase in the cAMP-induced influence on the activation of C/EBP transcription factors was observed during a combined action of phorbol ester and serotonin. The more substantial cAMP-induced rise in DNA-binding activity of C/EBP family transcription factors was observed during increase in the intracellular Ca2+ concentration (incubation of the CNS with calcium ionophor A23187 and forscolin). Thus, the convergence of Ca2+ and cAMP-dependent regulating signals that reflect the interaction of different modal stimuli during learning can produce the intensification of formation of the C/EBP transcription complexes. PMID- 11253405 TI - [Sensorimotor reactivity (acoustic startle response) in rats with experimental depressive syndrome]. AB - Alterations of the sensorimotor responses in Wistar rats with experimental dopamine deficit-dependent depressive syndrome induced by neurotoxin 1-methyl-4 phenyl-1,2,3,6-tetrahydropyridine were measured by acoustic startle. In rats with innate high level of anxiety the development of behavioral depression was accompanied by the decrease in startle amplitude. In rats with innate low level of anxiety the decrease in startle amplitude did not reach the statistical significance. Correlation between the anxiety-phobic level and the expression of behavioral depression was not revealed. Independently of the initial anxiety phobic level, in rats with depressive syndrome at the stage of behavioral rehabilitation after the neurotoxin withdrawal the prepulse inhibition of the acoustic startle was decreased as compared to control animals. It is suggested that the decrease in startle amplitude and, to a greater extent, the decrease in prepulse inhibition may characterize the development of dopamine deficit dependent states. PMID- 11253406 TI - [Effect of fractions of the cerebrospinal fluid from patients with drug dependence on basic electrophysiological characteristics of the rat olfactory cortex slices]. AB - Heroin addicts at the initial stage of abstinence syndrome were subjected to detoxication by liquorosorption technique. The fractions of their cerebrospinal fluid obtained by the thin layer chromatography technique were analyzed. The substances extracted from the cerebrospinal fluid of drug addicts, presumably peptides, negatively affected the conductive function and synaptic transmission in surviving slices of the olfactory cortex of rats. The conclusion was drawn about a possibility of application of surviving rat brain slices as test object for estimation of the extent of purification of the cerebrospinal fluid from toxic endogenous substances after the liquorosorption. PMID- 11253407 TI - Surfing the Odornet: exploring the role of smell in life and healing. PMID- 11253409 TI - The ill effects of nagging. PMID- 11253408 TI - Usefulness of guggul (Commiphora mukul) for osteoarthritis of the knee: An experimental case study. PMID- 11253410 TI - Distinguishing science from scientists, round two. PMID- 11253411 TI - Measuring the statistical probability of dreams? PMID- 11253412 TI - The illusion of nonlocality. PMID- 11253413 TI - Straus gives state of the center, CAM on PUBMED unveiled. PMID- 11253414 TI - Research in Ayurveda: where do we go from here? PMID- 11253415 TI - Ayurveda: a historical perspective and principles of the traditional healthcare system in India. AB - Ayurveda, the science of life, is a comprehensive medical system that has been the traditional system of healthcare in India for more than 5000 years. This medical system was well established around 2500 to 600 BC, when it evolved into 2 schools: the School of Physicians and the School of Surgeons, similar to allopathy. Charak Samhita, Susrut Samhita, and Ashtang Hridaya Samhita are the Senior Triad texts, and Madhav Nidan Samhita, Sarangdhar Samhita, and Bhavprakash Samhita are the Junior Triad texts. Around 600 BC. Ayurveda was branched into internal medicine; pediatrics; psychiatry; surgery; eye, ear, nose, and throat; toxicology; geriatrics; and eugenics/aphrodisiacs. The body is composed of 3 body doshas, 3 mental doshas, 7 dhatus, and malas. The harmony among the body doshas of vata (nervous system), pitta (enzymes), and kapha (mucus) and the gunas, or mental doshas (which are human attributes: satogun [godly], rajas [kingly], and tamas [evil]), constitutes health, and their disharmony constitutes disease. The management of illness requires balancing the doshas back into a harmonious state through lifestyle interventions, spiritual nurturing, and treatment with herbo mineral formulas based on one's mental and bodily constitution. PMID- 11253416 TI - Healthcare and disease management in Ayurveda. AB - Because the disharmony of mental doshas (satogun, rajogun, and tamogun) and body doshas (vata, pitta, and kapha) are the major cause of illness, the goal of illness management in Ayurveda is to bring back harmony among the doshas. The management includes clinical examination, diagnosis, and dietary and lifestyle interventions and treatment. The clinical examination consists of Astha Sthana Pariksha (8-point diagnosis: pulse-diagnosis, urine, stool, tongue, voice and body sound, eye, skin, and total body appearance examinations) and examination of the digestive system and the patient's physical strength. The treatment consists of cleansing (Panchkarma), palliation (improve digestion, remove toxic waste, fasting, observe thirst, exercise, sunbathing, and meditation), mental nurturing, and spiritual healing depending on the disturbed doshas and the patient's constitution. The preferred use of bhasms and herbal formulas over the respective metallic salts or the single herbs is discussed. This review suggests a great potential for integration of Ayurvedic therapies into the healthcare system in the United States. PMID- 11253417 TI - Complementary and alternative medicine: applications and implications for cognitive functioning in elderly populations. AB - OBJECTIVE: Aged populations in the United States are growing in numbers, and stand to be affected most by the changing shape of healthcare delivery. Within these elderly populations, the problem of decreased cognitive functioning due to dementing disorders is rising. Recent compelling research on complementary and alternative medicine interventions targeted at cognitive deficits in the elderly is reviewed in this survey. DATA SOURCES: A literature review was undertaken to identify original clinical research studies, review articles, chapters, and books on treating cognitive deficits in the elderly. Contact with complementary and alternative medicine researchers provided additional information concerning developments in this field. STUDY SELECTION: Research studies that were methodologically sound were selected for review. More purely clinical studies also were included to provide a thorough overview of the limited amount of accumulated knowledge in this field. DATA SYNTHESIS: A qualitative synthesis of the above data was used to comprehensively present all information accumulated to date in this field. CONCLUSIONS: Although still in the preliminary stages of development, clinical research exploring the benefits of complementary and alternative therapies for cognitive deficits among the elderly shows a significant level of promise that warrants a further investment of resources. PMID- 11253419 TI - Scott Gerson, MD. Ayurvedic medicine. Interview by Bonnie Horrigan. PMID- 11253418 TI - Modulation of neuroimmune parameters during the eustress of humor-associated mirthful laughter. AB - CONTEXT: Humor therapy and the related mirthful laughter are suggested to have preventive and healing effects. Although these effects may be mediated by neuroendocrine/neuroimmune modulation, specific neuroimmune parameters have not been fully investigated. OBJECTIVE: To determine the efficacy of mirthful laughter to modulate neuroimmune parameters in normal subjects. DESIGN: A series of 5 separate studies based on a multivariate repeated measures design, with post hoc simple contrast analysis. SETTING: The schools of medicine and public health at Loma Linda University, Loma Linda, Calif. SUBJECTS: 52 healthy men. INTERVENTION: Viewing of a humor video for 1 hour. Blood samples were taken 10 minutes before, 30 minutes into, and 30 minutes and 12 hours after the intervention. MAIN OUTCOME MEASURES: Natural killer cell activity; plasma immunoglobulins; functional phenotypic markers for leukocytes including activated T cells, nonactivated T cells, B cells, natural killer cells, T cells with helper and suppressor markers, and assessment of plasma volume and compartmental shifts; plasma cytokine--interferon-gamma; and total leukocytes with subpopulations of lymphocytes, granulocytes, and monocytes. RESULTS: Increases were found in natural killer cell activity (P < .01); immunoglobulins G (P < .02), A (P < .01), and M (P < .09), with several immunoglobulin effects lasting 12 hours into recovery from initiation of the humor intervention; functional phenotypic markers for leukocyte subsets such as activated T cells (P < .01), active cytotoxic T cells (P < .01), natural killer cells (P = .09), B cells (P < .01), helper T cells (P < .02), uncommitted T cells with helper and suppressor markers (P < .02), helper/suppressor ratio (P = .10) with several leukocyte subset increase effects lasting 12 hours after the humor experience; the cytokine interferon gamma (P = .02), with increases lasting 12 hours; total leukocytes (P < .05), with specific subpopulation lymphocytes during the intervention (P < .01) and 90 minutes into recovery (P < .05); and granulocytes during the intervention (P < .05) and 90 minutes following the intervention (P < .01). CONCLUSION: Modulation of neuroimmune parameters during and following the humor-associated eustress of laughter may provide beneficial health effects for wellness and a complementary adjunct to whole-person integrative medicine therapies. PMID- 11253420 TI - Meditation: myths and misconceptions. PMID- 11253421 TI - Dolphinswim: finding your 'first self' in the deep blue sea. PMID- 11253422 TI - [Genetic explanation for vertical evolution]. AB - The genetic theory of natural selection proposed by Fisher takes into account differential reproduction success of organisms, which may be estimated by using the Malthusian parameter as fitness. However, the minimum possible value of this parameter depends on ecological stability of an organism, which determines the probability of the survival and participation in reproduction for each viable offspring. In the course of vertical evolution, leading to an increase in the level of biological organization, ecological stability of organisms increases, and this might be accompanied by a decrease in their fitness. In the macroevolutionary process, alterations in ecological stability of organisms, including those responsible for an increase in the level of biological organization, are basic and primary changes whereas alterations in fitness are additional and secondary. PMID- 11253423 TI - [Interlineage distribution and characteristics of the structure of two subfamilies of Drosophila melanogaster MDG4 (gypsy) retrotransposon]. AB - The distribution of two variants of MDG4 (gypsy) was analyzed in several Drosophila melanogaster strains. Southern blot hybridization revealed the inactive variant of MDG4 in all strains examined and active MDG4 only in some of them. Most of the strains harboring the active MDG4 variant were recently isolated from natural populations. It is of interest that the active MDG4 prevailed over the inactive one only in strains carrying the mutant flamenco gene. Several lines were analyzed in more detail. The number of MDG4 sites on salivary-gland polytene chromosomes was established via in situ hybridization, and MDG4 was tested for transposition using the ovoD test. PMID- 11253424 TI - [Effect of integrating the pJFF350 vector into the 85-MDa plasmid of Azospirillum brasilense Sp245 on bacterial flagellation and mobility]. AB - Results of genetic analysis of three derivatives of Azospirillum brasilense Sp245 (strains BK570, SK051, and SK248) carrying cointegrates of plasmids 85-MDa and pJFF350 (the vector for omegon mutagenesis), which manifest abnormalities in flagellation and motility, are presented. It was shown for the first time that the integration of the suicide vector into one of Azospirillum resident plasmids is accompanied by the formation of various fusion products and changes in flagellation and motility of these bacteria, such as the loss of the polar (Fla) and lateral (Laf) flagella in SK051; inactivation of Fla and Laf in SK248; and Fla-dependent acceleration of expansion in semiliquid media in BK570. PMID- 11253425 TI - [Complementation analysis of mutants of the associative bacteria Azospirillum brasilense Sp245 and S27, defective in mobility and flagellation]. AB - Three mutants of Azospirillum brasilense Sp245 incapable of both formation of the polar flagellum (Fla-phenotype) and swarming in semisolid media (Swa-phenotype) were characterized. These mutants were shown to have lost the 85-MDa plasmid and to carry the Tn5-Mob transposon and pSUP5011 vector in different regions of their genomes. With the use of A. brasilense Sp245 gene bank, the capacity for both polar flagellum formation and swarming was restored in the above mutants and in the previously generated transposon mutants A. brasilense Sp245 and S27. The transconjugants obtained were only slightly motile in the liquid culture. In the gene bank of Sp245, the recombinant plasmids carrying wild-type fla/swa loci were identified. PMID- 11253426 TI - [Damaging effect of antibiotics on the structure of synaptonemal complexes of meiotic chromosome of mice]. AB - The structure of synaptonemal complexes (SCs) of chromosomes of mouse primary spermatocytes were studied using electron microscopy on days 1, 10, and 36 after the completion of per os administration of drugs belonging to three groups of antibiotics: tetracyclins, macrolides, and fluoroquinolones. The antibiotics were administered to mice during ten days. At the substages of early and middle pachytene, heteromorphic SC bivalents and fragments of chromosome-core elements were detected in spermatocytes at all times studied after the administration of the antibiotics of three groups. As cells passed through the period from early to middle pachytene, the number of cells containing heteromorphic SC bivalents and the fragments of chromosome cores gradually decreased, which could be an indication of selection of cells with chromosomal aberrations. A high level of associations between the X chromosome and autosome bivalents (including heteromorphic ones) also favors this suggestion. A gradual decrease in the number of chromosomal aberrations was detected, as time elapsed from the completion of antibiotics administration. The study of sperm obtained from epididymises of males did not reveal significant differences in both morphology and motility of sperm between males of the control and experimental groups. PMID- 11253427 TI - [Dependence of the combining ability of mutant sugar beet plants on their phenotype]. AB - Repetitive gametic selection for a higher frequency of the Adh1-S semilethal mutant allele of the alcohol dehydrogenase (ADH) gene yielded viable homozygotes Adh1-SS. The plants varied in phenotype from weak mutant to nearly normal (restored). Phenotypically different plants were individually tested for combining ability. This parameter was high in plants with the mutant phenotype and tended to decrease, rather than further increase, in plants with a restored normal phenotype. The results are discussed in terms of viability restoration mechanisms in homozygotes for semilethal mutant alleles. PMID- 11253428 TI - [Hydrocarbon-binding peptides from bean plant lectins in connection with various host specific macrosymbionts during development of symbiosis with rhizobium bacteria]. AB - The nucleotide sequences of 280-360-bp domains of lectin genes from 20 legume species belonging to 17 genera have been determined. A computer analysis of the sequences has been performed with the LASERGENE package. Based on this analysis, we constructed the phylogenetic tree of the lectins, which reflects their phylogenetic and evolutionary relationships, and predicted the amino-acid sequences of the corresponding protein domains. Features of the structure of the hydrocarbon-binding lectin domains were elucidated in some species of legume genera from the temperate climatic zone. The domains were highly variable and contained the consensus sequence AspTrePheXxxAsxXxxXxxTrpAspProXxxXxxIns/DelArgHis bearing the bulk of amino acid replacements, insertions, and deletions. An association between legume groups (including species from different genera and tribes) symbiotic with the same rhizobium species and the similarity between the hydrocarbon-binding domains of lectins from these plants was found. PMID- 11253429 TI - [Genetic diversity of natural populations of Arabidopsis thaliana (L.) Heynh. in Karelia]. AB - The genetic structure of ten natural populations of Arabidopsis thaliana (L.) Heynh. at eight isozyme loci was studied. The populations were located in the northern part of the species range, 200 km from the north to the south along the Onega Lake coast in Karelia. Considerable genetic diversity (P99% = 43.7, Hobs = 0.003) was revealed that is not typical of populations of self-pollinating plant species. A direct correlation between the proportion of polymorphic loci and geographical latitude was shown (r = 0.68; P < 0.05). It is suggested that a high polymorphism level in Karelian Arabidopsis thaliana (L.) populations increasing from the south to the north is due to extreme environmental conditions in the northern part of the species range. The distribution of genetic diversity within and between populations is typical of self-pollinating species: the larger part of the total diversity resides among populations (GST = 0.583). PMID- 11253430 TI - [Radiation mapping of the short arm of swine chromosome 2]. AB - In recent years, maps of mammalian genomes have been acquiring increasingly higher resolution. Integration of maps of different types has become possible. As a tool in integrating maps of mammalian genomes of different types, high resolution mapping with radiation-induced hybrids (RH) is used. Here, we present an RH6000 map of the short arm of porcine chromosome 2. The map contains 15 microsatellites and five genes (for parathyroid hormone, lactate dehydrogenase A, myogenic factor, follicle-stimulating hormone beta, and calpain I). The RH panel was obtained on the basis of a hybrid cell line bearing the single porcine chromosome 2 against the background of mink chromosomes. The mean frequency of preserving markers examined in the panel was 18.3%. Integration of four genes in the panel and a comparison of gene order in homologous regions of human and porcine chromosomes are presented. PMID- 11253431 TI - [Effect of selection for behavior of wild gray rats (Rattus norvegicus) on the morphology of the adrenal cortex]. AB - A comparative morphological and functional study of adrenal cortex was conducted in adult male Norway rats selected for domestic and aggressive behavior. Morphological changes were shown to occur in the structure of zona fasciculata of adrenal cortex during selection for domestic behavior. PMID- 11253432 TI - [Karyotype of Chukotka char from Estikhed Lake (Eastern Chukotka)]. AB - The karyotype of chars from the Estikhed Lake (Eastern Chukotka) was examined. This karyotype comprises 78 chromosomes, NF = 98. Marker chromosomes include one pair of submetacentrics, one pair of large acrocentrics, and one pair of large subtelocentrics with very short second arms. Nucleolus organizer regions are located in telomeric regions of short arms of marker submetacentric chromosomes. Small heterochromatin blocks are observed in centromeric regions of most chromosomes. The Chukotka char karyotype is very similar to that of Taranetz char Salvelinus taranetzi from the Achchen Lake: these karyotypes differ only in stability of the chromosome number. PMID- 11253433 TI - [Genetic variability for number of CAG-repeats in the X-linked androgen receptor gene and embryonal cell death in humans]. AB - Distribution of the X-linked androgen receptor gene (AR) alleles with different number of CAG repeat units in the first exon was analyzed in spontaneous abortuses having the 46,XX karyotype. The increased frequency of alleles with low number of trinucleotide repeat units in these specimens compared to adult healthy women was revealed. For more detailed analysis, all AR alleles were grouped into three classes: A, with a low CAG repeat number (21 to 23); B, with a moderate repeat number (24 to 28); and C, with a high maximum repeat number (29 to 32). Statistically significant among-sample differences in the frequencies of AA genotypes, which prevailed in spontaneous abortuses, were revealed. The samples tested were also different by the mean squared distance between the AR allele lengths, which constituted 81.8 +/- 0.4 and 53.2 +/- 0.5 in healthy women and spontaneous abortuses, respectively. Based on the data on inverse correlation between the length of the CAG repeat sequence and transcriptional activity of the androgen receptor gene, a hypothesis on selection against female individuals with the small number of CAG repeats in the AR gene and high transcriptional activity of this locus in prenatal period of human ontogeny is advanced. PMID- 11253434 TI - [Haplotype of Y-chromosomes in the Central Asia population]. AB - The distribution of alleles and haplotypes of three diallellic Y-specific loci (YAP, DYF155S2, and Tat) in the populations of Kyrgyz, Uzbeks and Tajiks was analyzed. In Kyrgyzes and Uzbeks, a relatively high frequency of the DYF155S2 deletion (20 and 12.5%, respectively) and the C allele at the Tat locus (11.2 and 8.3%, respectively) were revealed. In the populations of southern Kyrgyzes and Uzbeks, two chromosomes carrying the YAP+ allele were detected. In both cases the YAP+ allele was found within the YAP+/DYF155S2+/TatT haplotype. The Tajik population was monomorphic in respect to the polymorphisms studied. The Tajiks demonstrated the presence of only the YAP-/DYF155S2+/TatT haplotype. This haplotype appeared to be most frequent in Kyrgyz (78.8%) and Uzbeks (83.3%). The question on the origin and the distribution of Y-chromosome variants in Eurasia are discussed. PMID- 11253435 TI - [A method of evaluating the stationary rate of gene migration (a cartographic approach)]. AB - A method of geographic mapping of the stationary (limiting) gene migration rate has been developed. The method is based on approximation of the empirical distribution of gene frequencies by a theoretical steady-state distribution. The maximum likelihood method and the chi 2 minimization method are used to obtain consistent estimations of the gene migration rate as a parameter of the steady state distribution. The new method makes it possible to determine the geographical distribution of the ratio between the properties of the population migration structure described by the stepping-stone and island models and to construct a geographical map of chi 2 values. This map approximately reflects the distribution of natural selection pressure on the gene pool if genetic processes are quasisteady. PMID- 11253436 TI - [Analysis of polymorphism of the glutathione S-transferase gene in populations of the Volga-Ural region]. AB - The frequency of the GSTM1 gene deletion homozygotes in eight populations of the Volga-Ural region belonging according to linguistic classification to Turkic (Bashkirs, Tatars, and Chuvashs), Finno-Ugric (Maris, Komis, Mordovians, and Udmurts), and Eastern-Slavic (Russians) ethnic groups, was examined by means of PCR technique. The frequency of the deletion homozygotes varied from 41.4% in Bashkirs to 61.3% in Mordovians. The mean deletion frequency comprised 50.1%, which was consistent with the data for European populations (chi 2 = 0.009). PMID- 11253437 TI - Computer use and physical inactivity in young adults: public health perils and potentials of new information technologies. AB - Physical inactivity contributes to premature mortality and morbidity and increasing prevalences of overweight and obesity in industrialized countries. Computer use is an increasingly common sedentary behaviour, potentially displacing physical activity. Physical activity and computer use were examined in 697 young adults (18-30 years). Energy expenditure estimates were derived from self-reported walking, moderate, and vigorous activity; participants were classified as sedentary, low, moderate, or high in their level of activity. For multivariate analyses, two categories of physical activity were used: inactive (sedentary/low activity; < 800 kcal.week-1) or active (moderate/high activity; > or = 800 kcal.week-1). Time spent in computer-related activities was summed, and computer use tertiles calculated (< 3 hours.week-1; 3-8 hours.week-1; > 8 hours.week-1). Those in the highest tertile of computer use were most likely to be inactive (p = 0.003) and most likely to report computer use as a barrier to physical activity (p < 0.001). The majority of those in the top two tertiles of computer use, and of the inactive, preferred obtaining information from computers than from conventional print media. These findings suggest that computer use plays a significant role in the discretionary time of young adults and is negatively associated with physical activity. Computer-mediated communication has potential in disseminating interventions to increase physical activity in young adults. PMID- 11253438 TI - Neuroendocrine and leukocyte responses and pulmonary function to acute stressors. AB - Although stress is linked to asthma exacerbation, underlying mechanisms are unclear. Given the shared relevance to stress and asthma, select neuroendocrine and immune responses to acute stressors and their impact on pulmonary function were examined, comparing responses between students with (n = 20) and without childhood asthma (n = 16). Students were challenged with speech and math tasks. Blood samples were collected five times: before tasks, immediately after first and second tasks, and 15 and 60 minutes posttasks. Pulmonary function was measured four times, excluding midtask point. Stress reactivity patterns did not differ between two groups. However, all measures showed significant changes across the challenge. Plasma epinephrine and norepinephrine rose during tasks and declined after tasks, p < .001. Cortisol mainly declined after tasks, p = .03. Leukocyte count increased during tasks with increased lymphocyte percentage that declined after tasks, while neutrophil percentage changed opposite to lymphocytes, p < .001 each. Changes in pulmonary function were significant, p < .05, but were not predicted by the magnitude of neuroendocrine and immune changes. Instead, neuroendocrine and immune levels explained 33%-51% of variance on concurrent pulmonary function. Findings indicate that acute stress induces significant neuroendocrine and immune changes that can affect pulmonary function. However, stress reactivity needs further investigation with larger samples and people with a more severe form of asthma. PMID- 11253440 TI - Behavioral epidemiology: a systematic framework to classify phases of research on health promotion and disease prevention. AB - Although the term "behavioral epidemiology" has been used in the literature since the late 1970s, it has not been clearly defined. A behavioral epidemiology framework is proposed to specify a systematic sequence of studies on health related behaviors, leading to evidence-based interventions directed at populations. The phase are: 1--establish links between behaviors and health; 2- develop measures of the behavior; 3--identify influences on the behavior; 4- evaluate interventions to change the behavior; 5--translate research into practice. Mature research areas are expected to have more studies in the latter phases. Recent volumes of four journals (Annals of Behavioral Medicine, Health Psychology, Journal of Nutrition Education, Tobacco Control) were audited, and empirical studies were classified into these phases. Phase 3 studies were common (identifying influences on behaviors; 27% to 50%), and Phase 2 studies were least common (measurement; 0% to 15%). Annals of Behavioral Medicine and Health Psychology were low on Phase 4 (intervention studies; 9% and 11%, respectively). The Journal of Nutrition Education was the only journal reviewed that had a substantial number (20%) of Phase 5 studies (translating research into practice). The behavioral epidemiology framework can be used to evaluate the status of research on health behaviors and to guide research policies. PMID- 11253439 TI - Sun protection behaviors and stages of change for the primary prevention of skin cancers among beachgoers in southeastern New England. AB - Sun exposure is the most important avoidable cause of skin cancers. We report characteristics of a representative sample (N = 2,324) of beachgoers in Southeastern New England during the summer of 1995. This sample was not employing adequate sun protection behaviors (83% did not often avoid the sun during midday and only 45% often used sunscreen). Important demographic and skin cancer risk factor differences in sun protective behaviors and stages of change for sun protection were found, especially differences based on age, gender, and degree of sun sensitivity. Consistent with previous research, increased age, female gender, and greater sun sensitivity were each independently associated with more sun protective behaviors. These findings underscore the need for interventions targeting high-risk populations, such as those receiving high-intensity sun exposures at the beach. PMID- 11253441 TI - Health beliefs and pediatric emergency department after-care adherence. AB - This study's purpose was to apply a multivariate adaptation of the Health Belief Model (HBM) to examine parental adherence to pediatric emergency department (ED) after-care instructions. Parents/legal guardians (n = 162) of children ages 0-17 years with minor (noncritical) conditions (e.g. abrasion/contusion, laceration) completed health beliefs and demographics questionnaires while waiting for their child to be seen. Postdischarge, children's medical records were reviewed for after-care instructions and insurance status, and parents were phoned to assess adherence to specific after-care instructions. In logistic regressions, health beliefs (barriers, severity, susceptibility) and child age significantly predicted several postdischarge adherence behaviors, including home care procedures and prescription medication purchase. Results are discussed as they relate to the effects of specific health belief variables and the need for further refinement of the HBM in accordance with the changing health care system; clinical applications are proposed. PMID- 11253442 TI - Psychosocial factors and intervention-associated changes in those factors as correlates of change in fruit and vegetable consumption in the Maryland WIC 5 A Day Promotion Program. AB - This study sought to examine: (a) the relative effects of baseline demographic and psychosocial factors and an intensive nutritional intervention on changes in fruit and vegetable consumption in low-income, ethnically diverse women served by the Special Supplemental Nutrition Program for Women, Infants, and Children (WIC) program in Maryland; (b) whether this intervention, designed to modify psychosocial factors associated with fruit and vegetable consumption, was successful in changing these factors; and (c) whether changes in these factors were associated with increased consumption. The same women from 15 WIC program sites were surveyed at baseline and postintervention 8 months later. Women randomized to the intervention group showed significantly greater mean change in self-efficacy, attitudes, social support, and knowledge of national consumption recommendations than control group women. Changes in all psychosocial factors were significantly associated with nutrition session attendance and increased consumption. In a hierarchical model, changes in these factors accounted for most of the intervention effect on increased consumption. PMID- 11253443 TI - Cortisol secretion throughout the day, perceptions of the work environment, and negative affect. AB - The effects of explanatory variables derived from a work stress model (the effort reward imbalance model) on salivary cortisol were assessed. A multilevel analysis was used to distinguish the effects of single occasion and multiple occasion measurements of work stress and effect on cortisol. The single (or cross sectional) factors include Effort-Reward Imbalance (ERI), need for control, negative affect, and other enduring factors (type of occupation, gender, and smoking). The multiple occasion measurements include momentary negative mood, Momentary Demand-Satisfaction Ratio (MD-SR), sleep quality, work load (workday versus day off), at work (versus not being at the workplace), and lunch. The effect of time of day on cortisol was controlled for before the effects of these variables were determined. Momentary negative mood but not trait negative affect was positively associated with ambulatory measured cortisol. The variables from the work stress model--effort, reward, need for control, and the multiple occasion measurements of demand and satisfaction--did not affect cortisol. As could be expected, time of day had an effect on cortisol, but a hypothesised interaction with momentary negative mood was not found. Additionally, the results show that the time course of cortisol differs between individuals and that the effect of sleep quality on cortisol can vary from person to person. This points to the necessity of continued efforts to single out sources of individual variability. The finding that variables derived from the effort-reward imbalance model are not related with cortisol does not support the hypothesis that ERI leads to short-term changes in cortisol, indicating no relation with hypothalamic pituitary-adrenal (HPA) axis activity. On the other hand, the present results invite further qualification of negative affect as a potential determinant of HPA activity, at least, as far as can be deduced from cortisol measurements. PMID- 11253444 TI - The use of nicotine replacement therapy during hospitalization. AB - Recent findings suggest that smokers who are hospitalized experience significant craving for cigarettes. Thus, nicotine replacement therapy (NRT) may be a particularly important tool for use during hospitalization. The goal of this study is to evaluate the utilization of the transdermal nicotine patch and/or nicotine gum by hospitalized smokers. The data represented in this article are from 580 smokers who participated in a study of a motivational intervention for smoking cessation that was delivered during hospitalization. The primary outcome for this analysis was use of NRT during hospitalization. The results revealed that, among the entire sample, only 7.1% of the overall sample used NRT during hospitalization; 6% of the hospitalized smokers used the transdermal nicotine patch, and 1.1% used nicotine gum. Use of NRT was significantly greater among patients who reported that they were doing anything to help themselves quit smoking at the time of admission (OR = 4.1), those who were seriously planning to quit smoking within the next 30 days (OR = 2.36), those who were nicotine dependent (OR = 2.81), and those for whom a physician had ever offered to prescribe NRT (OR = 1.9). The finding that there is a very low rate of NRT use during hospitalization provides important information to hospital-based care providers and smoking cessation intervention planners. Barriers to NRT use among hospitalized patients should be identified, and strategies designed to maximize use when appropriate. The AHCPR Guideline on Smoking Cessation recommends routine use of NRT in health care settings. Further research is needed to determine why NRT use was so low. In addition, these data suggest that efforts to increase NRT use during hospitalization are needed. PMID- 11253445 TI - The use of refusal postcards in recruiting older adults. AB - This article examines whether a refusal postcard makes recruitment more efficient or instead reduces response rates to a telephone survey of older adults. Medicare health maintenance organization (HMO) members were randomly sampled in sequential phases. All samples received an initial contact letter from a HMO geriatrician. A refusal postcard was included in the first sample (N = 178); however, the remaining six samples did not receive this postcard (N = 1,003). An overall refusal rate of 32% was observed when postcards were included versus a 14% rate of refusal when postcards were excluded (p < .001). When potential respondents were reached by telephone, refusal rates were similar (9% versus 10%). Despite the higher refusal rate among the sample receiving the refusal postcard, no significant differences in demographics, health, and health behaviors were observed between the two final sample groups completing the survey. We conclude that refusal postcards greatly increase the refusal rates without offering any prescreening advantage in the recruitment process of older adults and could increase the costs of recruitment for a telephone survey. Furthermore, use of a refusal postcard precludes individuals from making fully informed decisions about participating in research. PMID- 11253446 TI - Medicare: intentions, effects, and politics. PMID- 11253447 TI - How not to think about Medicare reform. PMID- 11253448 TI - A tale of two Editions: Marmor's The Politics of Medicare and the study of health politics after thirty years. Essay Review. PMID- 11253449 TI - Political analysis and medical care: The Politics of Medicare reconsidered. Essay Review. PMID- 11253450 TI - Lasting Impact ... The Politics of Medicare. Essay Review. PMID- 11253451 TI - Medicare and the politics of incrementalism. Essay review. PMID- 11253452 TI - The Politics of Medicare north and south. Essay review. PMID- 11253453 TI - Comments on JHPPL Review Symposium. Essay review. PMID- 11253454 TI - The social roles of Medicare: assessing Medicare's collateral benefits. AB - The Medicare program incorporates a number of functions that go beyond providing health insurance to its beneficiaries. These activities, which we refer to as "collateral" functions, may have important health consequences but are also an increasing source of controversy. In this essay we develop a conceptual framework for categorizing these involvements, introduce some additional options that might complement Medicare's current collateral functions, assess the reaction of policy elites and Medicare's current beneficiaries to these alternatives, and evaluate the role that collateral activities play for Medicare's core mission. A case can be made for expanding some collateral involvements, but only if the Health Care Financing Administration has the strategic direction and administrative capacity to effectively implement these activities. PMID- 11253455 TI - Paying for Medicare: benefits, budgets, and Wilbur Mills's policy legacy. AB - Medicare features an unusually complex financing design. The Hospital Insurance Trust Fund pays for Part A of Medicare (hospital stays), while the Supplementary Medical Insurance Trust Fund finances Part B (doctor visits, outpatient care, and certain home health services). At a time when Medicare policy is generating debate, this article takes a new analytical look at the origins and consequences of the program's peculiar bifurcated structure. Addressing historians of the U.S. welfare state as well as contemporary health policy reformers, the article focuses on the crucial role of legendary Ways and Means Committee chair Wilbur Mills in Medicare's enactment in 1965. The central theme of the article is that fiscal conservatism and a commitment to budgetary restraint constitute important elements of Medicare's original political understanding. Contrary to analysts who argue that Medicare's financing design has produced "perverse" effects, we argue that it has served a valuable social function by encouraging policy makers to confront periodically the costs of one of the largest and fastest-growing federal programs. An argument can be made that Medicare's original division requires modification in order to integrate health care delivery changes of the past few decades. It is crucial, however, for reformers not to lose sight of the policy goals, including fiscal rectitude, that motivated the adoption of Medicare's bifurcated structure in the first place. PMID- 11253456 TI - The Medicare Cost Report and the limits of hospital accountability: improving financial accounting data. AB - Health policy makers, legislators, providers, payers, and a broad range of other players in the health care market routinely seek information on hospital financial performance. Yet the data at their disposal are limited, especially since hospitals' audited financial statements--the "gold standard" in hospital financial reporting--are not publicly available in many states. As a result, the Medicare Cost Report (MCR), filed annually by most U.S. hospitals in order to receive payment for treating Medicare patients, has become the primary public source of hospital financial information. However, financial accounting elements in the MCR are unreliable, poorly defined, and lacking in critical detail. Comparative analyses of MCRs and matched, audited financial statements reveal long-standing problems with the MCR's data, including major differences in reported profits; variations in the reporting of both revenues and expenses; an absence of relevant details, such as charity care, bad debt, operating versus nonoperating income, and affiliate transactions; an inconsistent classification of changes in net assets; and a failure to provide cash flow statements. Because of these problems, MCR financial data give only a limited and often inaccurate picture of the financial position of hospitals. Audited financial statements provide a more complete perspective, enabling analysts to address important questions left unanswered by the MCR data. Regulatory action is needed to create a national database of financial information based upon audited statements. PMID- 11253457 TI - Headache as the sole presentation of acute myocardial infarction in two elderly patients. AB - Myocardial infarction presenting solely as an acute, severe headache is underdiagnosed in elderly patients. In patients over 80 years of age, myocardial infarction presents more commonly with atypical symptoms than with chest pain. The authors describe two patients who presented with headache as the only symptom of an acute myocardial infarction. The authors recommend that acute myocardial infarction be considered in the differential diagnosis of acute, severe headache in elderly patients. PMID- 11253458 TI - Post-tussive sinus node slowing. PMID- 11253460 TI - 2:1 atrioventricular block partly concealed by P-on-T: misleading lead II. PMID- 11253459 TI - Small thrombus on sinotubular ridge of ascending aorta. PMID- 11253461 TI - Cardiac amyloidosis in nonagenarians. PMID- 11253462 TI - Ethical issues in the management of geriatric cardiac patients. PMID- 11253463 TI - Preventive cardiology--its applicability at elderly age. PMID- 11253464 TI - Elderly patients at risk for coronary heart disease or stroke: selecting an ideal product for lipid lowering. AB - Coronary heart disease is an affliction of the elderly: 84% of those who die from the disease are over 65 years of age. In patients over 55 years, the incidence of stroke more than doubles with each decade of life. The 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors, or statins, have been shown to lower cholesterol and lipids in both middle-aged and elderly patients in large clinical trials. Some statins have been shown to improve endothelial function and vasodilation and to normalize thrombin formation, which may be among the mechanisms involved in both coronary event and stroke prevention. PMID- 11253465 TI - Smoking and obesity are associated with the progression of aortic stenosis. AB - OBJECTIVE: The purpose of the study was to identify clinical predictors of progression of aortic stenosis. BACKGROUND: The natural history of valvular aortic stenosis includes a latency period followed by an unpredictable progression. Recent investigations have shown an association between risk factors for atherosclerosis and the presence of aortic stenosis. The authors hypothesized that atherosclerosis risk factors are also associated with the progression of aortic stenosis. METHODS: In a retrospective study, patients with a diagnosis of aortic stenosis were identified by continuous wave Doppler and a follow-up study of at least 6 months. Clinical data at the time of the index echocardiogram were obtained from review of patients' medical records. Independent risk factors for the progression of aortic stenosis were identified by stepwise logistic regression analysis. RESULTS: One hundred twenty-three patients were identified, and complete data were obtained for 87 patients (mean age, 70.7 +/- 10 years; men, 81%; mean follow-up, 2.54 +/- 1.6 years). The initial gradient was mild in 61% of patients and moderate in 31%. The mean rate of progression was 6.3 +/- 13 mm Hg/year. Mild aortic stenosis in 36% of patients at the time of the index echocardiogram progressed to moderate or severe over an average of 2.9 +/- 2.0 years. Independent clinical factors associated with a progression of 5 mm Hg/year or greater included a history of smoking (relative risk [RR] = 3.06; 95% confidence interval [CI] = 1.09-8.61; p = 0.034) and body mass index (RR = 1.16; 95% CI = 1.03-1.30; p = 0.013). Hypertension, diabetes, cholesterol, age, gender, and coronary artery disease were not independently associated with progression. CONCLUSIONS: Body mass index and a history of smoking are independent predictors of significant progression of aortic stenosis, defined as > 5 mm Hg/year. The rate of progression of aortic stenosis is variable. However, a substantial number of patients have progression of even initially mild aortic stenosis within a relatively short period of time. The effect of controlling atherosclerosis risk factors on the rate of progression of aortic stenosis remains to be determined. PMID- 11253466 TI - Syncope in the elderly: new trends in diagnostic approach and nonpharmacologic management. AB - Syncope in the elderly is an important health care issue because of the large patient population, challenging diagnostic and therapeutic approaches, and potentially devastating consequences. Significant comorbidity and atypical clinical presentations render a precise determination of the cause of syncope difficult. Recent studies suggest that noninvasive tests, such as carotid sinus massage or tilt-table testing, can be helpful in the diagnostic workup. It has been shown that permanent pacemaker therapy benefits elderly patients with carotid sinus hypersensitivity, and younger patients with recurrent vasovagal syncope. The implantable wireless loop recorder can be effective in documenting transient arrhythmias as causes of syncope in selected patients. Elderly patients with syncope and a low ejection fraction are at increased risk of sudden death due to malignant ventricular arrhythmia. Electrophysiologic study and electrophysiology-guided therapy should be considered in this segment of the population. PMID- 11253467 TI - Carotid sinus syndrome and falls in older adults. AB - Retrospective and circumstantial evidence supports an overlap between symptoms of falls and syncope in older adults. Because of this overlap, we undertook a prospective, explanatory, single-center study of cardiac pacing for falls in patients with carotid sinus syndrome in a consecutive series of over 56,000 adult visitors to an emergency department. One third attended because of a fall; one in five fallers had unexplained falls, and one third of these had carotid sinus hypersensitivity, of whom one half had a cardioinhibitory or mixed response that may be expected to respond to cardiac pacing. In a randomized, controlled trial of a subset of these patients, cardiac pacing was shown to significantly reduce subsequent fall rates by two thirds and syncopal rates during 1-year follow-up. The current pacing rate for carotid sinus syndrome is much higher in our practice than in other series because our facility is dedicated to falls and syncope in older subjects who have direct access to referring physicians and family practitioners. Of the implants in our region, 24% are for carotid sinus syndrome, compared with 43% for atrioventricular block, 20% for sick sinus syndrome, and 12% for atrial fibrillation. These rates do not include pacing in patients who fall, but rather reflect pacing rates consistent with American College of Cardiology guidelines for carotid sinus syndrome, such as recurrent syncope. These preliminary results from a local explanatory study are now being tested in a multicenter study entitled Syncope and Falls in the Elderly: Pacing and Carotid Sinus Evaluation (SAFE-PACE II). PMID- 11253469 TI - [Granular cell tumor in the region of the hypophysis. Case report]. AB - We present a case of a 60-year old woman with a history of stroke with suspected aneurysm of anterior communicating artery. Angiography of cerebral arteries excluded the presence of an aneurysm. Magnetic resonance imaging showed the presence of a tumour in the area of the sella turcica. The tumour was resected, totally, using fronto-orbital approach. Histopathological examination showed granular cell type. The patient was released from the hospital in good health without any neurological deficits and hormonal disorders. On the basis of our case and reports of other authors we can conclude that benign character, usually with well defined boundaries from surrounding tissue and poorly vascularised, makes it easy to remove and gives good surgical results for this type of tumours. PMID- 11253468 TI - [Transcatheter closure of patent foramen ovale in patients after cryptogenic stroke]. AB - Paradoxical embolism through a patent foramen (PFO) is a possible mechanism of ischaemic stroke in patients with cryptogenic stroke. Occlusion of PFO in such patients is considered by some authors as most effective in stroke prevention. We present our initial experience with transcatheter closure of PFO with the new self-expanding device--the Amplatzer PFO occluder in three young patients (age < 50 years). Each of them experienced at least one ischaemic stroke episode, without a left heart or carotid source and each had an interatrial communication with right-to-left shunting during Valsalva manoeuvre on echocardiography. The PFO's were closed completely without complications, under transoesophageal echo guidance in general anaesthesia. Complete closure was confirmed at one-month follow-up echocardiogram in each patient. No repeat cerebral accidents occurred at that time. The procedures were relatively easy and the clear presentation of the implant on TEE and fluoroscopy, made implantation fully controlled. The unique feature of the device is, that until release it can easily be retrieved, repositioned or removed. Transcatheter closure of PFO with the Amplatzer PFO occluder may become the new therapeutic option for patients with cryptogenic stroke and presumed paradoxical embolism. PMID- 11253470 TI - [Dysembryoplastic neuroepithelial tumor (DNT)--case report and literature review]. AB - Neuroepithelial dysembryoplastic tumour was first described by Daumas-Duport in 1988 and in WHO classification was included into the group of neuronal and mixed neuroglial tumours. This is a benign and very rare tumor with a good prognosis occurring in children and young adults. The tumour caused characteristic clinical symptoms: epileptic fits, supratentorial, intracortical localisation, most often in temporal lobe and specific nodular architecture with heterogenic cell composition. Oligodendrocyte-like cells, glial and neuronal elements are usually found. The authors present a case of a 24-years old female with partial epileptic sensorial symptomatology. CT examination revealed a tumour in the left parietal lobe. Histological findings showed a typical texture of DNT. The tumour has no tendency for recurrence even in case of incomplete removal and does not require chemotherapy nor radiotherapy which is significantly important for accurate diagnosis, in order to avoid an aggressive therapy in young patients. PMID- 11253471 TI - [A letter concerning the article by J. Krzyzanowski et. al.: "A case of schizophrenia-like psychosis...", N.Nch.Pol. 1998, 32, 433]. PMID- 11253472 TI - [A letter to the editor concerning opinions about the usefulness of radiculography for diagnosis of lumbar disc diseases]. PMID- 11253473 TI - [Confidentiality and medicine]. PMID- 11253474 TI - [Incidence of atherosclerotic lesions in internal carotid arteries and leg arteries of patients with ischemic stroke]. AB - Atherosclerosis is a chronic process developing in all arteries. The aim of the study was the evaluation of the frequency of coexistence of atherosclerotic lesions in internal carotid arteries (ICA) and in the arteries in the lower extremities (LEA) in patients with cerebrovascular disease. Doppler examination of ICA and LEA was performed in 168 patients. The extension and character of pathological changes in ICA was estimated by of colour duplex ultrasonography, the grade of disorders of blood flow in LEA was according to the values of ankle:arm index (a:a ind.). Changes in LEA correlated proportionally to the size and extent of the pathology in ICA. In 46 patients with normal blood flow in ICA, 93.4% had normal blood flow in LEA. In 61 patients with small atherosclerotic changes in ICA, in 47.5% a decrease of blood flow in LEA (a:a ind. < 0.90) was found. In the group of 42 patients with severe stenosis of ICA, abnormal blood flow (a:a ind. < 0.70) in LEA in 40.5% was observed. CONCLUSION: 1. Atherosclerotic changes in the arteries of the patients with ischaemic stroke are generalised. 2. Normal blood flow in ICA is usually associated with normal blood flow in LEA and atherosclerotic changes in ICA are usually associated with disturbances of blood flow in LEA. PMID- 11253475 TI - [Blood platelet activation markers in patients with acute cerebral infarction during the earliest stage of the disease--evaluation using flow cytometry methods]. AB - Platelet activation seems to play a critical role in a number of vascular diseases, including stroke. The aim of our study was whole-blood flow cytometry evaluation of platelet activation markers: P-selectin (CD62), glycoprotein-53 (CD63) and platelet-derived microparticle in vivo in patients with acute cerebral infarction. We investigated 50 patients (29 men and 21 women, mean age 67.8) with acute cerebral infarction. Parameters of platelet activation were measured on the 1st, 3rd and 7th day after stroke onset. Comparisons were made with 20 control patients matched age. Compared to controls (1.6 +/- 0.7%) the stroke patients showed higher expression of CD62 on the 1st (2.6 +/- 1.4%), 3rd (3.5 +/- 2.3%) and 7th (3.0 +/- 2.0%) day after stroke onset. The differences were statistically significant on all days (p < 0.05). Compared to controls (1.5 +/- 0.6%) the stroke patients had also higher expression of CD63 on the 1st (1.9 +/- 0.6%), 3rd (2.0 +/- 0.6%) day and they showed higher level of platelet-derived microparticles on the 3rd (13.0 +/- 3.0%) day after stroke onset. The differences were statistically significant (p < 0.05). Elevated expression of CD62, CD63 and platelet-derived microparticles level indicate platelet activation during the acute phase of ischaemic stroke. Flow cytometry is a useful tool for in vivo assessment of platelet activation. PMID- 11253476 TI - [Evaluation of dysarthria with the assistance of acoustic speech analysis in patients with amyotrophic lateral sclerosis]. AB - The aim of the study was to assess dysarthria in ALS subjects using acoustic speech analysis. The study was performed in 47 definite or probable ALS patients aged 29-76 years (mean age 53.7 yr.) and in 30 age and sex matched healthy control subjects. Neurological examination showed 15 dysarthric ALS subjects. Acoustic speech analysis is a quantitative, computer-acoustic method estimating dysarthria and based on assessing of sound distance from speech sound tests. In both group the mean sound distance between chosen sounds was compared to a basic pattern and was measured on time-frequency computer acoustic analyses (delta f = 125 Hz, delta T = 9 ms, delta s = 0.5 dB). Our results demonstrated that all sounds were incorrect in all ALS subjects. These abnormalities were significantly increased in the dysarthric ALS subjects. The mean sound distances which separated ALS from control subjects is 0.2 (by Euclidian principle) in 4 out of 5 measured sounds. We suggest that it is possible to detect and measure dysarthria in ALS patients based on the acoustic speech analysis, also in the limb onset ALS subjects. PMID- 11253478 TI - [Assessment of blood flow velocity during cognitive stimulation in persons with suspected hydrocephalus]. AB - The aim of this work is to evaluate middle cerebral arteries (MCA) blood flow velocity changes during performance of linguistic and visuospatial cognitive tasks. Two groups were investigated: eight patients with suspected hydrocephalus without perceptible cognitive disturbances, and eight healthy persons. Blood flow velocity in left and right MCA was recorded with transcranial Doppler sonography while the examined patients were performing three different tasks. The analysis of the results showed differences between the groups concerning both increase of blood flow velocity values and performance patterns. Compared to healthy individuals less increase in blood flow velocity during performance of all tasks and no difference in haemodynamic changes between both hemispheres during task performance were observed in patients with hydrocephalus. The results obtained suggest, that the pattern of functional lateralization in brain in patients with suspected hydrocephalus is probably changed. PMID- 11253477 TI - [Subacute sclerosing panencephalitis (SSPE) in Poland in 1996-1999. Phase VII of epidemiologic studies]. AB - In the seventh phase of epidemiological studies of SSPE data were gathered on patients who developed the disease in 1996-1999. In this time period the diagnosis was confirmed in only 10 cases (4 in 1996, 4 in 1997, 0 in 1998 and 2 in 1999). This is a significant reduction of incidence in relation to the preceding stages, in particular to the years 1993-1995 in which 49 new cases were still reported. This is not ruling out the possibility that at the beginning of the years 2,000 cases would be diagnosed in which the first symptoms developed several or more months earlier. PMID- 11253479 TI - [Surgical treatment of pineal and tectal tumors from the subtentorial and supracerebellar approach]. AB - The results of operative treatment of pineal and tectal tumours by infratentorial, supracerebellar approach were presented. There were 15 patients with pineal and 2 with tectal tumours. Clinical symptoms of the disease were as follows: signs of intracranial hypertension, tectal lesion, impairment of gait and hypoacusis. Diagnostic procedure included CT, MRI, and panangiography investigation. Application of infratentorial, supracerebellar approach enabled total resection of tumours in 14 patients (82.3%) and subtotal in 2 patients (11.8%) In 1 patient we managed to do only diagnostic biopsy (5.9%). This approach makes possible removal of pineal and tectal tumours growing in the midline toward the anterior part of III ventricle and also posterior fossa. The advantage of this approach is fact that the operation takes place below the vein of Galen. Drooping by gravity cerebellum creates enough places for microsurgical operation. During control investigation 13 patients (76%) had no announce any complaints and were without neurological disorders. PMID- 11253480 TI - [Risk of hemorrhage and size of hematoma in patients with cerebral arteriovenous malformations]. AB - The occurrence of intracranial haemorrhage, epileptic seizures and focal neurological signs was estimated in the group of 40 patients with AVMs. Size of AVM, its localization was established by angiography. Maximum diameter of a haematoma was measured on preoperative CT. We found that small AVMs (diameter < 3 cm) are more often the source of intracranial haemorrhage than larger ones. Intracerebral haematoma was the larger the lesser was AVM. Intracerebral haematoma has no influence on immediate and long-term results of operative treatment of AVMs. PMID- 11253481 TI - [The role of the bcl-2 gene in programmed death of neurons]. AB - Alterations in counterbalance between proliferation and cell death contribute to the pathogenesis of many neurological disorders. The main form of cell death that occurs in the developing nervous system and in many pathological states is apoptosis. Many factors participate in the regulation of programmed cell death. Bcl-2 gene family members modulate the sensitivity of cells to death. Some of them promote cell death and the others, such as bcl-2, prevent apoptosis. Bcl-2 shows a wide expression in the nervous system and protects neurons from various toxic insults. This review focuses on the role of bcl-2 in the apoptosis occurring in some neurological diseases. Understanding of apoptotic threshold may lead to new strategies for the treatment in neurology. PMID- 11253482 TI - [Venous infarcts]. AB - Venous infarcts are uncommon and frequently misdiagnosed as arterial infarcts, intracerebral haemorrhages or tumours on CT. Cerebral venous thrombosis is a condition with large variety of causes. However, in 20 to 35% of causes, no cause is found. The clinical features depend on the location of venous thrombosis. Clinical signs are mainly headache, hemiparesis, cranial nerves paresis, epilepsy, TIA. The question about the proper way of treatment and duration of treatment remains open. PMID- 11253483 TI - [The role of molecular genetics in diagnosis of hereditary motor-sensory neuropathy]. AB - Hereditary motor-sensory neuropathies (HMSN) are a heterogeneous group of disorders of peripheral nervous system. Four genes in HMSN have been characterized so far i.e.: PMP22, MPZ, Cx32 and EGR-2. The advent of molecular genetic techniques over the past few years has provided identification of molecular defects in a few forms of HMSN. The present study describes the application of modern molecular genetic methods, which are used in the studies of HMSN. Southern blot hybridisation, Fluorescence in situ hybridisation (FISH), Short Tandem Repeat analysis (STR), Semiquantitative PCR analysis (SQ-PCR), Single Strand Conformation Polymorphism method (SSCP), Heteroduplex analysis (HD) and finally DNA automated sequencing are described in the present paper. In the conclusions the advantages and limits of mentioned methods of DNA analysis in HMSN have been described. PMID- 11253484 TI - [Modelling cerebrovascular circulation from the viewpoint of autoregulation mechanisms]. AB - The rationale for cerebral blood flow modelling is to extrapolate the possible effects of the pharmacological agents and significance of different pathologic states. Most authors present linear, statistical models. Such modelling does not take into consideration many important factors. Autoregulation of cerebral circulation is the most important one. It maintains cerebral blood flow on a constant level despite significant changes of systemic pressure. Influence of autoregulation is necessary for evaluation of the different anatomical configurations of the main supplying arteries and the cerebral arterial circle of Willis. This paper presents original, non-linear and dynamic model of the cerebral circulation with factor of autoregulation. Comparison of the data from the modelling and from the clinical observations shows high correlation. PMID- 11253485 TI - [Use of stereotactic methods in neurosurgery practice]. AB - Beginnings and historical evolution of stereotaxis and its role in a contemporary neurosurgery are performed. Application of this technique in clinical practice is reviewed. Brief description of stereotactic methods in neurooncology, movements disorders surgery, chronic pain therapy, radiosurgery, modern methods of neuronavigation as well as photodynamic and gene therapy is presented. PMID- 11253486 TI - [Peridural fibrosis in lumbar disc surgery--pathogenesis, clinical problems and prophylactic attempts]. AB - Postoperative peridural fibrosis is unavoidable adverse effect of lumbar disc surgery. This process is disadvantageous both to the patient and to the surgeon. It is assumed that peridural fibrosis is responsible for as much as 25% of all Failed Back Surgery Syndrome. In case of reherniated discs requiring reoperation epidural scar may cause technical difficulties. Thus the prevention or inhibition of postoperative peridural fibrosis and adhesions is an essential goal for successful lower back surgery. The authors review new opinions on pathophysiology of peridural fibrosis, clinical aspects of the process, results of experimental approaches for limiting peridural fibrosis and perspective of anti-adhesion gel Adcon-L. PMID- 11253487 TI - [Anaplastic oligoastrocytoma of the spinal cord: diagnostic difficulties. Case report]. AB - The authors report a rare case of 49-years old woman with rapidly progressing anaplastic oligoastrocytoma primarily localized in the spinal cord. Increasing spastic paresis of the right lower limb was concomitant with decrease in superficial sensibility in this limb and the right side of the trunk below Th10 level, suggested a lesion within the spinal cord. However, it was the difficult confirming the diagnosis by spinal MR imaging, and the negative result of the first MR examination (performed 5 weeks after manifestation of first clinical symptoms of the disease) delayed surgical treatment. During the next 3 weeks the neurological syndrome increased to spastic paraparesis with sphincters dysfunction and decrease in superficial and vibratory sensibility within the lower limbs and the trunk below the Th10 level. The second MR examination of the spine revealed an intraspinal tumour at Th8-Th10 levels. Surgical (partial excision of the tumour) and radiation treatment was given. Histopathological examination of tumour tissue showed the presence of anaplastic oligoastrocytoma. During the follow-up of our patient we found cerebral foci, probably of metastatic origin ascending with cerebrospinal fluid. More than 5 months after the diagnosis was established the patient died of primary disease. The imaging parameters of both spinal MR examinations were similar, however, on second examination narrower field of vision was used. In both cases Magnevist was administered. Discussing factors which might be responsible for the false negative result of spinal MR examination--the examination of choice for detection of proliferative transformation--the authors take artefacts connected with respiratory and circulatory function, peristaltic movements, and with field of vision into consideration. PMID- 11253488 TI - [Long-term safety of using tiagabine in epilepsy]. AB - Tiagabine (TGB) is a novel antiepileptic drug efficacious for the treatment of partial epilepsies. The aim of that work is short presentation of current data concerning long-term safety of TGB. Tolerance to TGB does not develop with long term therapy. Idiosyncratic reaction and changes in haematology and chemistry values have not been associated with TGB therapy. The most common adverse effects are dizziness, asthenia, nervousness, tremor, diarrhoea and depression. The current data do not show any evidence of relationship between visual field constriction and TGB treatment. No adverse effects on cognitive abilities have been found. There are contradictory data concerning tiagabine-induced nonconvulsive status epilepticus. Because of high safety and efficacy TBG is an important new antiepileptic drug for the treatment of intractable partial epilepsies. PMID- 11253489 TI - [Gabitril as an additive drug in therapy of intractable epileptic seizures in children]. AB - Tiagabine (Gabitril, Sanofi Synlhelabo) new antiepileptic drug was used in add-on therapy in 25 children with resistant partial complex and secondary generalized seizures. Treatment was carried out in children aged 4-17 years with low dose escalation from 5 to 45 mg/day, in three doses until good clinical effects were obtained. In 3 patients aged 4 years, in 11 children aged 5-12 years and in 11 children aged above 17 years Gabitril was used. Follow up period was 8-10 months. Frequency of epileptic seizures before implementation of Gabitril treatment, even during polytherapy with 2 or more antiepileptic drugs was several to hundred per day (status epilepticus was observed in 2 children with Rasmussen syndrome). During the observation 5 children became seizure free, in 11 patients reduction in seizures frequency above 50% was observed and in 9 children effects of treatment were not good enough. Gabitril was well tolerated, and any adverse events were observed in add-on therapy. Preliminary observation and good results of add-on therapy with Gabitril are positive. Drug is safe and generally well tolerated with good effects at add-on therapy in 64% children with resistant partial complex and secondary generalized seizures. PMID- 11253490 TI - [Open multicenter study of the effectiveness and safety of gabitril in epileptic patients with partial seizures]. AB - The paper presents the results obtained by 53 investigators implementing the first Polish multicentre study of the effectiveness and safety of tiagabine (Gabitril). The study included 81 patients with refractory epilepsy with partial seizures. The duration of the study was 16 weeks. For the initial 6 weeks, Gabitril was gradually introduced till a dose of 30 mg/day was achieved. Within the subsequent 10 weeks the treatment effectiveness was observed and monitored, with the provision that the dose could be increased. The final analysis included 62 patients, while in 12 subjects the treatment was discontinued in less than 16 weeks. The results indicate a very beneficial effect of Gabitril on the frequency of seizures in patients with drug-resistant epilepsy. Almost 1 of the analyzed patients were seizure free. The most beneficial effects with respect to seizure number and intensity reduction were noted in subjects with partial complex and partial seizures with secondary generalization. The dynamic character of the decrease in seizure frequency was best observed between the first and third month of therapy. In 2/3 of patients the recommended dose was achieved and maintained. Less than 15% of subjects were excluded from the study, mainly due to lack of therapeutic effects. The number and character of adverse effects observed in the course of the present study did not differ from these noted in long-term Gabitril trials. The drug was demonstrated to exert no effect on vital functions and laboratory parameters. The results confirm the high effectiveness of Gabitril in treatment of patients with partial seizures and a good tolerance of this agent. PMID- 11253491 TI - [Evaluation of tiagabine efficacy in different groups of patients with epilepsy]. AB - The aim of study is to discuss from a clinicians viewpoint the use of tiagabine in the clinical setting, in particular how we can relate our current knowledge to individuals in our clinics. The choice for individual and the clinician is multi layered dependent on many factors. Namely is based on patient characteristics, their beliefs and wishes, age, seizure type or syndrome and comorbidity. The next factor influencing choice is drug characteristics; pharmacology, mode of action, efficacy, tolerability, practical use and cost. Very important is fear of unknown. Particularly important are new drugs, which can be safety use in groups of special concern like children, women of reproductive age, the elderly, patients with psychiatric illness and mental handicaps. In these cases the risk of fetal malformations, interactions with other drugs, safety profile should be taken into consideration. Tiagabine represents an important new therapeutic option for patients with drug resistant partial seizures. The role of tiagabine in the management of patients with intellectual disability is especially emphasised since tiagabine has a low side-effects profile in the cognitive area. Comparison of data for new antiepileptic drugs provide information for selection among treatments when a second drugs is needed to improve control of seizures. However, there are numerous caveats in use of these summary data. First, the data from clinical trials cannot be compared directly because of variability in placebo response and potentially in other population characteristics. Second, the needs of individual patients differ. CONCLUSION: There is important to stress that no controlled clinical trials have provided algorithm for the best add-on drug or combination of drugs. So there is need to describe the homogenic groups of patients and final choice should be based on the need of the individual patient (sex, age, pregnancy, mental status) for superior seizure control versus minimal adverse effects. PMID- 11253492 TI - [Individualized obstetrical care for women with a history of sexual abuse]. AB - The consequences of a history of sexual abuse for pregnancy and delivery are illustrated in case histories of three women, aged 28, 27 and 31 years respectively. The first woman (who had a history of sexual abuse, like the other women) reacted to labour with dissociative behaviour. This was anticipated by giving her directions for the proceedings during labour, which helped restoring contact. The second patient expressed insecurity during her prenatal check-ups, the reaction to which was inadequate. When labour started her fear had grown and led to an early plea for a caesarean section. An epidural made caesarean section redundant. The third patient had re-experiences during her prenatal check-ups. External examination or an ultrasound were a burden to her. Together with the patient a detailed treatment plan was made which gave her control over the situation. It is advised to question all women during prenatal care about a possible history of sexual abuse and if found, to discuss the consequences of these experiences in her everyday life, especially during her pregnancy. The obstetrical care-giver has to be aware of the circumstances that might provoke traumatic memories and anxious feelings. By knowing and anticipating these events -also called triggers--, the professional can give 'tailor-made' care during pregnancy and delivery. PMID- 11253493 TI - [Williams syndrome: new insights into genetic etiology, pathogenesis and clinical aspects]. AB - Williams syndrome (WS) is a developmental disorder characterized by distinct facial features, congenital heart disease, mental retardation and a gregarious personality. The majority of people with this disorder have a submicroscopic deletion of genes in chromosome band 7q11.23. This deletion can be detected using fluorescence in situ hybridization (FISH). Although the condition is usually sporadic a few familial cases with autosomal dominant inheritance have been described. A clinical scoring system has been developed by Selicorni with which a diagnosis of 'Williams syndrome' can be made; in all patients in whom the diagnosis was made in this way FISH results were positive. PMID- 11253494 TI - [Development and developmental disorders of the human brain. II. Development of the cerebral cortex and major tract systems]. AB - In the development of the cerebral cortex, two phases can be distinguished: (a) the formation of the preplate, a superficial layer essential for a normal lamination of the cerebral cortex; (b) the formation of the cortical plate. The cortical plate divides the preplate into a superficial marginal zone (the future layer I) and the subplate. The transient subplate is important for the formation of thalamocortical projections. Most cortical neurons arise in the ventricular zone of the pallium and migrate along radial glial cells (radial migration) to the cortical plate. The gamma-aminobutyric acid (GABA)ergic cortical interneurons, however, originate from the ganglionic eminences and reach the cerebral cortex through tangential migration. PMID- 11253495 TI - [Diagnostic imaging of brain maturation in premature infants]. AB - In 3 studies brain maturation in 42 preterm infants with no or minor cerebral deviations was examined with MRI. With a scoring system 5 stadia in gyral development were recognized, which corresponded to the gestational age (duration of amenorrhoea of the pregnant woman). At a gestational age between 30 and 34 weeks, myelin was present in many brain stem, diencephalic and cerebellar structures, but only in a few hemispheric structures. Little progress in myelination was noticed up to the gestational age of 46 weeks. Remnants of the germinal matrix formed a sharp contrast with the immature unmyelinated periventricular white matter and had a characteristic appearance on MR images before term age. Broad periventricular zones with subtle change of signal intensity were also physiological in this age group. Zones with more outspoken signal intensity change compared with the rest of the cerebral white matter were, however, not physiological and represent mild ischaemic change of the periventricular white matter. PMID- 11253496 TI - [Diagnostic image (27). Normal multiparous cervix, around ovulation, with early transformation zone]. AB - Colposcopic image of early transformation zone of portio cervicis without abnormalities in a 31-year-old multiparous woman. PMID- 11253497 TI - [Lung cancer in the Netherlands in the period 1989-1997: the epidemic is not over yet]. AB - OBJECTIVE: To describe and interpret changes in incidence, mortality and survival of lung cancer in the Netherlands in the period 1989-1997. DESIGN: Secondary data analysis. METHODS: Data on the incidence of lung cancer were collected from the Dutch Cancer Registration (1989-1997), on mortality from Statistics Netherlands (CBS; 1989-1994), on the incidence of lung cancer in other European countries from EUROCIM (1990-1994), on survival of Dutch lung cancer patients from the Comprehensive Cancer Centre Amsterdam (1988-1997) and the Comprehensive Cancer Centre South (1988-1992) and on survival of other European lung cancer patients from EUROCARE (1985-1989). Incidence rates were calculated per 100,000 person years and standardized by age according to the European population structure. Survival was calculated as the ratio of observed survival among the lung cancer patients and the expected survival of the general population. RESULTS: The incidence of lung cancer among men decreased from 109 to 93, whereas that among women increased from 18 to 23. The incidence of lung cancer among Dutch men was high in comparison to other European countries, whereas that among women was average. The trends in lung cancer incidence were probably related to the trends in past smoking behaviour. Mortality decreased among men from 106 to 91 and increased among women from 15 to 20. Survival was better for younger patients, a localised tumour, and better for squamous cell carcinoma or adenocarcinoma than for large-cell undifferentiated or small-cell carcinoma. The relative 5-year survival was 12%, the relative 1-year survival 39%; these were good in comparison with other European countries. CONCLUSION: The incidence and mortality of lung cancer among Dutch men decreased, but still in 1997 almost 20 men in the Netherlands died each day of lung cancer. Among women the end of the increase is not in sight and in 1997 over 5 women died each day of lung cancer. PMID- 11253498 TI - [Pain in the symphyseal region after parturition: possibly osteomyelitis]. AB - A 29-year-old woman 3 weeks after her first childbirth suffered from atypical and progressive pain in the pelvis, which turned out to be a symptom of osteomyelitis of the pubic bone. She recovered after treatment with antibiotics and 6 weeks' stabilization of the pelvis. Symptoms of osteomyelitis resemble those of pubic osteitis, symphyseal rupture and symphysiolysis. Radiologically, osteomyelitis is characterized by development of infiltrates, cortical involvement and local osteopenia. Isolation of micro-organisms in a bone culture after puncture is regarded as proof of the diagnosis. The treatment is primarily with antibiotics, if abscesses or sequestra develop these should be relieved and/or removed. PMID- 11253499 TI - [Delirium due to increase in clozapine level during an inflammatory reaction]. AB - Between January 1999 and May 2000, the Netherlands Pharmacovigilance Foundation LAREB received five reports of patients with clozapine intoxication attributed to inflammation. The reports all concerned men with schizophrenia, aged 63, 54, 41, 45 en 42 years. The occurrence of increased clozapine levels during inflammation, and normalisation after recovery, strongly suggest a causal relationship. No other possible explanations were found. A three to five-fold increase occurred in most instances, but one patient experienced a ten-fold increase compared with the basal levels of clozapine. Three of the patients developed a delirium as an intoxication symptom, probably due to anticholinergic effects on the central nervous system. In case of an inflammatory reaction in patients on clozapine treatment, the physician should be aware of the possibility of clozapine intoxication and delirium. Measuring clozapine levels during infection and dosing based on these levels can minimise the adverse effects of clozapine intoxication. PMID- 11253500 TI - [Value of a good example: a search for role models in practical clinical teaching]. AB - The importance of the role model function of the 'teacher' (m/f) was always realized to some extent in practical clinical teaching, but appears to be gaining renewed interest at present. There exists, however, a discrepancy between the importance attached by many experienced clinicians and their students to the role model on the one hand, and the amount of systematical study that has been dedicated to it on the other hand. The available date show an impressionistic image of the qualities these students and interns consider important in role models. However, the 'why' of the importance of role models is seldom considered. In an era in which it is considered important that 'learning targets', 'learning moments' and 'transfer mechanisms' are defined in detail, the function of role models in practical clinical teaching should also be given more attention. Insights from other fields of knowledge than the medical field might be of value in designing research leading to a clearer image of the function and the value of role models in practical clinical education. PMID- 11253501 TI - [Meticulous handling of electronic patient records in family practice]. PMID- 11253502 TI - [Embolization as treatment for postpartum hemorrhages]. PMID- 11253503 TI - [Embolization as treatment for postpartum hemorrhages]. PMID- 11253504 TI - Cutaneous manifestations in inflammatory bowel disease. AB - Numerous extraintestinal manifestations in various organ systems have been reported to be associated with inflammatory bowel disease (IBD). Aim of the present paper was to evaluate the frequency of cutaneous manifestations in Crohn's disease (CD) and ulcerative colitis (UC) with respect to their location, the activity and location of the underlying disease, the treatment options and the time to remission. METHODS: The medical records of 1043 inpatients with CD and UC were screened retrospectively for extraintestinal symptoms with special regard to cutaneous manifestations. RESULTS: The prevalence of cutaneous manifestations in IBD was 22/1043 (2.1%; 18 women, 4 men; age: 31.41 +/- 9.9 [21 51] yrs.). In 15/22 patients (68.2%) the cutaneous manifestations were associated with CD, in 7/22 patients (31.8%) UC was confirmed. In 6/22 patients (27.3%) pyoderma gangrenosum (PG) was diagnosed, in 16/22 patients (72.7%) erythema nodosum (EN). EN and PG predominately occurred at the lower legs: in 68.1% the tibia was the main affection site. Other locations like breast or anus were rare. In 16/22 patients (72.7%) an acute phase of the underlying disease was evident, in 6/22 patients (27.3%) CD or UC were in remission. In patients with CD a colonic involvement was found in 86.7%. Arthritis was the most frequent coexisting extraintestinal manifestation in CD (53.3%) and UC (28.8%). Drug treatment was performed with high doses of glucocorticoids and salicylates. The time to remission in patients with EN was significantly shorter as compared to PG (5.3 +/- 1.8 vs. 19.6 +/- 14.2 weeks; p < 0.001). In 5/22 patients (22.7%) cutaneous manifestations reoccurred after a symptom-free interval. All efflorescenses reoccurred during an active phase of the underlying disease at the same manifestation site as the initial presentation. CONCLUSION: In this series the prevalence of cutaneous manifestations in IBD was 22/1043 (2.1%). EN and PG were more frequent in women with IBD, in CD, and during the acute phases of the underlying disease. EN and PG predominately affect the lower legs. Cutaneous manifestations respond well to an acute phase therapy of the underlying disease. The time to remission was significantly shorter in EN as compared to PG. However, relapses have to be considered in a relevant subgroup of patients. PMID- 11253505 TI - Pilot study of interferon-alpha high-dose induction therapy in combination with ribavirin plus amantadine for nonresponder patients with chronic hepatitis C. AB - Ribavirin plus interferon-alpha (IFN alpha) combination has led to a marked advance in the treatment of IFN alpha-naive or relapser patients with chronic hepatitis C but was shown to be only marginally effective in IFN alpha nonresponders. We therefore conducted a pilot study to see whether an intensified treatment protocol might be more effective in inducing a virological response in patients who had not responded virologically to previous IFN alpha monotherapy. 14 nonresponder patients with histologically proven chronic hepatitis C were included in the study. Patients received 9 MU IFN alpha-2a daily for one week followed by 9 MU IFN alpha every second day for further 5 weeks. With the beginning of the seventh week, patients were treated with 6 MU IFN alpha thrice in week (tiw) for a period of 6 weeks (until week 12). IFN alpha was continued up to 48 weeks at a dose of 3 MU IFN alpha tiw. Ribavirin (1000-1200 mg/day) and amantadine sulphate (200 mg/day) was given orally for 48 weeks. One patient discontinued therapy after first IFN alpha injection and one other patient after 12 weeks of therapy because of side effects. The remaining 12 patients completed treatment according to the protocol. An initial virological response at week 24 was achieved in 2 of the 14 patients (14%) and both patients remained HCV RNA negative at the end of treatment. However, both patients relapsed 4 weeks after completion of therapy, and therefore none of the patients achieved a virological sustained response. Viral dynamics studies showed a marked decline in hepatitis C viremia during the first 6 weeks of high-dose IFN alpha. After IFN alpha dose reduction, however, viremia stabilized or increased in most patients. These data indicate, that even triple therapy with high-dose IFN alpha plus ribavirin and amantadine fails to improve significantly the response rates in IFN alpha nonresponders. PMID- 11253506 TI - Weekly 24-h infusion of high-dose 5-fluorouracil (5-FU) with folinic acid (FA) in adjuvant therapy of colon cancer. AB - After curative resection (R-0) of primary colon cancer or curative metastases resection, the efficacy, toxicity and compliance of a weekly 24-h infusion of high-dose 5-FU with folinic acid was examined in a prospective feasibility trial. From June 1995 to June 1997, 19 patients were included, 11 patients with UICC stage III and 8 patients with UICC stage IV colon cancer. The patients received weekly 500 mg/m2 of calcium folinic acid (Rescuvolin) as a 1-2 h-infusion on an outpatient basis following a 24-h infusion of 2,000 mg/m2 5-FU via a pump system (intermate LV5 Baxter). The adjuvant therapy was administered for 6 months. 90% of the patients received the planned 18 chemotherapy applications. The total 5-FU dose given to each patient amounted to 34.4 g/m2 in 6 months, thus corresponding to 95% of the planned 5-FU total dose. The main toxicity was diarrhea CTC toxicity grade 3 in the case of 16% of the patients. After a median follow-up of 51 months (range: 37-59 months), 82% of the patients (9 out of 11) with stage III remained free of recurrence. The 2 cases of recurrence belonged to the pN2 subgroup. In stage IV only 12% of the patients (1 out of 8) remained free of recurrence. On an adjuvant basis, a weekly 24-h infusion of high-dose 5-FU with folinic acid is accompanied by a good complicance and a high-dose intensity of 5 FU. Now it is tested within randomized phase III trials of the "Arbeitsgemeinschaft Gastroenterologische Onkologie (AGO)" of the "Deutsche Gesellschaft fur Verdauungs- und Stoffwechselerkrankungen" for UICC stage III colon cancer. Concerning stage IV, adjuvant therapy was not effective, a fact that seems to justify new drugs and new therapeutic strategies. PMID- 11253507 TI - [Study Group of Gastroenterological Pathology of the German Society of Pathology. Recommendations for celiac disease/sprue diagnosis]. AB - The diagnosis of celiac disease (CD) is based upon histological findings in duodenal or jejunal biopsies. In the past few years it has turned out that the development of CD lesion in the small bowel is a dynamic process which may present in various histological forms. At one end of the spectrum is a mucosa with normal architecture and an increase in intraepithelial lymphocytes (IEL), at the other end is the classical flat mucosa. Histological features supporting the diagnosis of CD are architectural changes of the villi and/or crypts, an increase in lamina propria cell density and an increase in IEL counts. For diagnostic purposes and for monitoring of CD patients an exact histological classification of the histological findings has to be given. This has become possible by using a modified Marsh classification. In the present paper the histological presentation of CD is presented as well as the modified Marsh classification and the most important differential diagnoses. PMID- 11253508 TI - [Ulcerative esophagitis and colitis as rare manifestations of Adamantiades-Behcet disease]. AB - We report the case of a 39-year-old German women who suffered from chronic inflammatory bowel disease which had not been exactly classified for 6 years. In the course of the disease she developed recurrent iritis, nodular skleritis, oral and genital ulcer, erythema nodosum and axillary folliculitis. For the first time retrosternal pain occurred. Endoscopy of the upper gastrointestinal tract showed mid esophageal ulcer formation. Colonoscopy demonstrated ulcerative colitis with no involvement of rectal mucosa. Histologically a cutaneous vasculitis in the lower limb was seen and diagnosis of Adamantiades-Behcet disease with marked gastrointestinal involvement and rare manifestation in the esophagus was established. A therapy with prednisolone, azathioprine and mesalazine was started. Under this regimen stomatitis, esophagitis and cutaneous vasculitis resolved, while the colitis showed partial remission now for up to one year. PMID- 11253509 TI - [Opisthorchiasis simulating a malignancy]. AB - We report on 2 patients from Siberia suffering from an infection with the parasite Opisthorchis felineus. The unusual course of their disease pretended in case 1 an eosinophilic leukemia and in case 2 a malignoma of the gallbladder. The Opisthorchiasis is endemic in large areas of Asia and Russia. Humans acquire the infection by eating raw fresh-water fish. Symptoms are nonspecific, but detection of eosinophilia in travellers or residents of endemic areas should induce analysis for specific antibodies against Opisthorchis species. Opisthorchiasis is known to be a precursor of cholangiocarcinoma. Malignoma which was initially suspected could be excluded in both cases and the patients were cured by oral administration of Praziquantel, 40-75 mg/kg body weight for 1 day in 3 divided doses. PMID- 11253510 TI - [Anisakiasis of the stomach--a case report from Germany]. AB - Anisakiasis or "herring worm disease" is one of the most important parasitic diseases of the gastrointestinal tract in Japan. In 1988 Lorenz and Warzok published 8 cases of intestinal anisakiasis in Eastern Germany. In 1988 Spehn et al. reported a case of gastric anisakiasis in an AIDS patient. Here, we describe a case of gastric anisakiasis in Germany with an impressive serious clinical course. The symptoms--acute abdominal cramps, severe chest pain, diarrhoea, sub febrile temperatures and leucocytosis--followed 4 h after consumption of raw herring, which was homemade pickled in vinegar. The conventional and the endoscopic ultrasonography showed a thickened gastric wall made of mainly thickened submucosa. The larvae of Anisakis in the gastric mucosa were found and extracted endoscopically. Acute and severe abdominal pain after eating raw fish is an indication for early gastroscopy. The endoscopical extraction of possible larvae is the only effective therapy, as anthelmintics against nematodes (mebendazole, albendazole, thiabendazole) are ineffective. PMID- 11253511 TI - [Palliative therapy of pancreatic adenocarcinoma]. AB - The palliative therapy of pancreatic carcinoma consists of best supportive care (BSC) including nutritional and pain therapy, and antineoplastic remedies. This is complemented by interventional endoscopy aiming to treat obstructive jaundice and/or gastric or duodenal obstruction by implantation of endoprostheses. Pain therapy is standardized for the most part according to the WHO guidelines; nutrition of the cancer patient, however, is sometimes disregarded. For palliative chemotherapy, 5-fluorouracil and gemcitabine can be recommended. Combinations thereof with other cytostatic drugs or radiation are subject to ongoing studies. A variety of novel therapeutic concepts, e.g. immunomodulation or suicide gene therapy, have demonstrated good effects in animal studies. Unfortunately, very few of these have entered clinical studies (phase I and phase II). This will be the focus of future research in this field. PMID- 11253512 TI - [Does a diet high in fiber prevent development of colorectal adenomas?]. PMID- 11253513 TI - [Thalidomide in treatment of Crohn disease: a new approach for managing chronic inflammation?]. PMID- 11253514 TI - Long-term follow-up of patients with iron deficiency anemia. PMID- 11253515 TI - [The Journal of Orthopedics online]. PMID- 11253516 TI - [Sensomotor function while wearing lumbar support ortheses]. AB - AIM: Flexible lumbar corsets should contribute to an improved active and passive stabilisation of the lumbar spine. In the present study, the effects of flexible corsets on sensory-motor abilities have been investigated. METHODS: 24 healthy subjects (m = 10, f = 14, 26.7 years) performed angle- and force-reproduction tasks with and without wearing a flexible corset. The subjects tried to reproduce trunk flexion angles of 20 degrees und 40 degrees as well as forces of 33% and 66% of the maximum strength of the back extensor muscles. Additionally, muscular reactions were measured during destabilizing stimuli. RESULTS: No significant differences were found for the reproduction tasks. When wearing the corset, the left erector spinae muscles showed a delayed onset (+15 ms, p < 0.05), and the obliquus abdominis reached maximal activity more rapidly (-18 ms, p < 0.05). In the preinnervation phase the obliquus abdominis (-33%, p < 0.01), in the eccentric phase the erector spinae (-27%, p < 0.05) and the right obliquus abdominis (-46%, p < 0.05) and in the concentric phase the right obliquus abdominis (-35%, p < 0.01) and the rectus abdominis (-32%, p < 0.05) showed lower activities. CONCLUSION: The reduced muscle activities with destabilizing stimuli can be interpreted as the result of an improved passive stabilisation of the lumbar region by wearing a flexible corset. PMID- 11253517 TI - [Efficacy of MRI--whole spine image in diagnosis of vertebral metastases--results of a prospective study]. AB - AIM: To prospectively investigate the outcome efficacy of whole spine MRI for diagnosis and treatment in patients with suspected metastases of the spine. METHODS: All patients older than 50 years with newly diagnosed back pain and/or newly diagnosed spine-related neurological symptoms without a diagnosis by other imaging modalities were accepted in this study. A whole spine MRI and a detailed MRI per spine region with suspicious lesions were performed using a Siemens Magnetom Expert 1.0 Tesla machine. Outcome efficacy was determined by assessing further therapy and result for the patient. RESULTS: In all 15 patients of the year 1999 whole spine MRI allowed us to determine the definite diagnosis and treatment. Plain X-ray and 99mTc bone scanning gave a diagnostic suspicion but no definite diagnosis or therapeutic consequence. CONCLUSIONS: MRI of the spine including whole spine images allows clear cut decision making in diagnosis and treatment of cases suspicious for metastatic disease of the spine. Careful history taking and clinical examination provide enough information to opt for whole spine MRI as the first choice investigation. This will provide maximum benefit to the patient and avoid examination cascades. PMID- 11253518 TI - [Radiologic and computerized tomography evaluation of pedicle screw placement in lumbar spondylodesis]. AB - PURPOSE: To determine the accuracy of a standard roentgenogram for the placement control of pedicle screws following spinal fusion. METHOD: From 1995 to 1997 we performed computed tomography (CT) after material removal following lumbar and lumbosacral spinal fusion in 16 patients. We compared the placement of the 76 pedicle screws in plain X-rays after spinal fusion with their appearance on CT. A correct placement was defined as no penetration of the pedicle cortex, no contact of the lateral or ventral cortex of the vertebral body or joint, and a sufficient screw length. RESULTS: We found correct placement of 58 screws (76.3%) on the standard roentgenogram, whereas by CT only 46 (60.5%) were placed correctly. The results of both radiological examinations correlated (correct/incorrect placement) for only 54 screws (71.1%). A penetration of the ventral cortex of the vertebral body in 21 cases (27.6%) was identified in only 11 screws (14.5%) on the postoperative X-rays. The two radiological methods in this instance correlated for 62 screws (81.6%). CONCLUSION: The value of postoperative standard roentgenogram for the placement control of pedicle screws following spinal fusion is low. By this method possible contact with the aorta, vena cava, dura or vertebral joint cannot sufficiently be excluded. PMID- 11253519 TI - [Joint biomechanics and design of modern knee prostheses--time for revised thinking!]. AB - AIM AND METHOD: This review article summarises new knowledge about knee kinematics and induces a new discussion about the design of total knee arthroplasty (TKA) components. RESULTS: According to these new observations, knee flexion is not linked to femoral rollback but to a rotational movement between tibia and femur. The axis of this rotation is situated in the medial compartment of the knee when an intact anterior craciate ligament is present and not centrally through the tibial spines. In case of ACL insufficiency, such as that following TKA, the center of rotation shifts into the lateral compartment. Furthermore, the form of the posterior femoral condyle is not elliptical but round. CONCLUSION: Rotational movements between femoral component and tibial baseplate with the polyethylene-inlay have to be possible. One needs an asymmetric surface of the polyethylene-inlay, because different movements occur in the medial compartment than in the lateral compartment. The option to construct the posterior femoral condyle with a single radius allows a high congruency with the articulating inlay. The surgeon should let the new findings influence his choice of a TKA system. A closer analysis of modern prosthetic designs with either fixed or mobile bearings reveals that a few systems have already incorporated some of the new kinematic aspects of the knee. PMID- 11253520 TI - [Surgical correction and stabilization of neuromuscular scoliosis--2-4-year results of dorsal and one-stage ventro-dorsal operated patients]. AB - QUESTION: In order to evaluate the outcome of the operative treatment of neuromuscular scoliosis 45 patients were studied prospectively. METHODS: 27 Patients were operated by posterior correction and fusion using the Munster Posterior Doublerod-System (MPDS) (GI). 18 Patients were treated with one-stage ventro-dorsal procedure in combination of VDS with MPDS (GII). For all patients, medical and radiographic records were available, with a minimum follow-up of 2 years. Postoperative management, bloodloss and complications will be discussed. RESULTS: In group I the main curve (76.3 degrees), by an average flexibility of 36.1%, were corrected by 53.5% and 52.3% at follow up. The mean pelvic obliquity (7.7 degrees) averaged 53.8%. The mean major scoliosis of group II (107.1 degrees), by an average flexibility of 25%, gets improved by 61%. At most recent follow-up, the mean correction was 61.5%. The mean pelvic obliquity (23.1 degrees) averaged 73.2% and 70.6% respectively. The mean bloodloss in group I was 1840 ml and in group II 2180 ml. CONCLUSION: The data in the current study support the benefit of the operative treatment of patients with severe neuromuscular scoliosis. The quality of life gets improved by stability in seating and standing by correction of pelvic obliquity and trunk instability. PMID- 11253521 TI - [A validated finite element model of the human spine--description of the model and initial application]. AB - AIM: To generate a finite-element model of the human cervical spine and evaluate the first application of the model to the analysis of new c-spine implants. METHODS: CT-data were used to generate a three-dimensional, anisotrophic, linear model of the human C4-C7 motion segments using the software ANSYS 5.4. As a next step, anterior cervical fusion and plate fixation using mono- and bicortical screws was simulated in the model. Loading of the finite-element models was simulated using pure moments of +/- 2.5 Nm in flexion/extension, axial left/right rotation, and left/right lateral bending. The range of motion was calculated. The results were compared to the results of an in vitro study using human cadaveric c spine segments C4-C7, with the same implants and moments on both the intact and surgically treated specimens. RESULTS: The results obtained by the finite-element model were always within one standard deviation of the results of the in vitro study. CONCLUSION: Keeping in mind the simplifications of such a mathematical model, it may be used for a first analysis of the shape of new c-spine implants or to predict the initial stability of a new device. PMID- 11253522 TI - [Water jet discotomy with microinvasive approach--in vitro testing and initial clinical aspects of a new procedure]. AB - AIM: The difference in consistence of the nucleus pulposus and the annulus fibrosus allows the water jet to selectively remove the nucleus in a closed vertebral disc at a certain pressure range. The aim of the study was to investigate the use of water jet cutting in microinvasive spinal surgery. METHODS: A comparison in terms of efficiency between the water jet and those of the laser and APLD (automatic percutaneous lumbar discotomy) was achieved by plastic reconstruction of the resected spaces using the in-vitro-model of the spinal column of young pigs. The in-vitro-study was followed by a prospective clinical study with 21 patients. RESULTS: The in-vitro-employment of the three different methods showed that there were no significant differences in volume of the removed nucleus material. During the use of the hydro jet at 50 bar and simultaneous suction the intradiscal pressure measured in vitro remained below 1 bar. Clinical tests on the 21 patients showed good to very good results in 71% of the patients tested (mean follow-up 5.8 months). No complications were found. As working mechanism the pure mechanical effect and the influence on chemical processes within the nucleus remain points for discussion. CONCLUSION: The current studies results demonstrate that hydrojet spinal surgery might be a safe new method for surgery of disc protrusion and contained prolapse. PMID- 11253523 TI - [Significance of jet lavage for in vitro and in vivo cement penetration]. AB - AIM: The purpose of this study was to determine the efficacy of pulsatile jet lavage and manual syringe lavage with regard to their cleansing capabilities as measured by cement penetration into cancellous bone both in vivo and in vitro. METHODS: Three separate experiments were performed. Study A: In a cadaver study 36 left human cadaver femora were used for implantation of cemented femoral components. Conventional broaches were used for femoral preparation. Bone lavage was carried out either using jet lavage or manual syringe lavage of equal volume. The allocation to two different lavage groups was randomised. In both groups high pressurising cementing techniques were implemented with the use of a proximal seal and additional finger packing. Study B: To guarantee standardised cement pressurisation and equal bone quality, the influence of jet lavage (1000 ml) versus syringe lavage (1000 ml) was studied in 11 paired human cadaver femora in an additional study without prosthesis implantation. The specimens were imbedded in specially designed pots. Bone cement was applied in a retrograde manner and subjected to a standard pressure protocol with a constant force of 3000 N. Study C: To directly compare the effectiveness of both pulsatile jet and syringe lavage with regard to cement penetration in vivo, a new sheep model allowing for standardised bilateral, simultaneous cement pressurisation was used. After femoral neck osteotomies both femoral cavities of 10 sheep were prepared for retrograde cement application. After randomisation one side was lavaged with 250 ml irrigation using a bladder syringe, the contralateral femur with the identical volume but using a pulsatile lavage. A specially designed apparatus was used to allow for bilateral simultaneous cement pressurisation. ANALYSIS: In all studies horizontal sections were obtained from the femoral specimens at predefined levels using a diamond saw. Microradiographs were taken and analysed using image analysis to assess cement penetration into cancellous bone. RESULTS: Study A: Compared with syringe lavage the use of jet lavage significantly improved the penetration of cement into cancellous bone (p = 0.027). In the presence of strong, dense cancellous bone the findings were more pronounced. Study B: Our results show that in equal quality bone, the use of jet lavage yields significantly (p < 0.001) improved cement penetration compared to syringe lavage specimens. Study C: The results of the in vivo study confirmed the superiority of jet lavage bone surface preparation (p = 0.002). CONCLUSIONS: The use of jet lavage yields significantly improved interdigitation between cancellous bone and cement both in vitro and in vivo and should be regarded as mandatory in cemented total hip arthroplasty. High pressurising techniques are effective means to improve cement penetration, but should only be administered with jet lavage to reduce the risk of fat embolism. PMID- 11253524 TI - [Cryosurgical ablation of bone tissue with a newly developed miniature cryoprobe- adaptation of the method for use in bones in vitro and in vivo]. AB - AIM: Up to now, modern miniature cryoprobes have been used successfully for local destruction of soft tissue tumors without damaging adjacent healthy tissue. In this study, the methodology of cryoablation was applied to bone and the freezing effect as well as the cooling capacity of the probe was examined in vivo and in vitro. METHODS: Freezing was performed by cooling one or two probes, with a diameter of 3.2 mm to -180 degrees C with liquid nitrogen. The cooling capacity of the probes was determined under thermic control by an in vitro measurement on human bone, followed by an in vivo measurement on femoral and tibia bones of a sheep. RESULTS: The in vitro freezings achieved a sufficient tissue cooling using one or two cryoprobes. The simultaneous use of 2 probes resulted in a synergistic effect between the probes. According to the body heat, the registered temperature curves, during the in vivo freezings, showed a more flat trend. Nevertheless, temperatures below -50 degrees C were achieved at a distance of 1 cm from the probe due to the synergistic effect. Local or systematical intraoperative complications have not been observed. CONCLUSION: An adequate tissue cooling of bone matrix can be achieved within in vivo freezings through the use of one or more miniature cryoprobes so that the use of this probe could possibly become an alternative or supplement to the surgical resection of pathologic bone processes. PMID- 11253526 TI - [Histomorphology versus three-dimensional ultrasound morphology of the rotator cuff]. AB - AIM: Sonographic evaluation is a reliable method for the detection of rotator cuff tears. However, the diagnostic value of the different echogenic patterns of the rotator cuff, especially in elderly patients, has led to a controversial discussion. The present study elucidates whether three-dimensional ultrasound can increase the diagnostic significance of different sonographic echogenicity patterns of tendons compared to histopathological tissue degeneration. METHOD: To evaluate the sonographic appearance of tendons in elderly patients, the sonographic echogenicity of 33 rotator cuff specimens with macroscopic absence of rotator cuff lesions was classified in three groups and compared with the histopathological morphology. RESULTS: The sonographic appearance of the degenerated rotator cuff in elder patients correlates with a reduced sonographic echogenicity. The two-dimensional ultrasound evaluation of the rotator cuff led to a sensitivity of 84.0%, a specificity of 42.9%, a positive predictive value of 63.6%, a negative predictive value of 69.2%, and an accuracy of 65.2%. The three dimensional ultrasound evaluation of the rotator cuff led to a sensitivity of 91.6%, a specificity of 50.0%, a positive predictive value of 66.6%, a negative predictive value of 84.6% and an accuracy of 71.%, respectively. CONCLUSION: As a high incidence of false positive results has to be taken into consideration, neither the two-dimensional, nor the three-dimensional ultrasound evaluation seems to lead to a reliable correlation. PMID- 11253525 TI - [Inadequate detectability of early stages of coxarthrosis with conventional roentgen images]. AB - AIM: The study was undertaken to determine the value of standard radiographs in the early stages of osteoarthritis. METHODS: Standard radiographs and arthro MRI's from thirty hips operated on for early arthrosis (age 25-57 years) were independently analyzed by two orthopaedic surgeons and one radiologist blinded from the intra-operative findings. The radiographs were read on two occasions two months apart. The radiographic findings were then compared to the intra-operative findings. RESULTS: Intra-operatively, all cases had a labral lesion and, in all but three of the cases, there was a major acetabular cartilage lesion. Each investigator diagnosed all of the labral and/or cartilage lesions on the arthro MRI. However, on average, the investigators judged 20% (10-35%) of the hips to be normal on the standard radiographs. The probability of detecting an abnormal hip joint was statistically significantly better with arthro-MRI in four of six readings (p < 0.05) and there was a trend in favor of the arthro-MRI in the other two readings (p < 0.1). Intra-observer agreement when using the Tonnis classification of arthrosis on standard radiographs was 0.26 (-0.1-0.62), 0.69 (0.42-0.96) and 0.83 (0.53-1) [kappa-statistic, (95% confidence interval)]. The interobserver agreement was 0.24 (-0.07-0.55). CONCLUSION: Plain radiographs in the early stages of osteoarthrosis of the hip are neither reliable nor valid to diagnose the onset of disease. Therefore, in the case of a normal radiograph and clinical suspicion of arthrosis, a "normal" radiograph does not exclude the diagnosis and an artho-MRI should be obtained for further evaluation. PMID- 11253527 TI - [Non-cerebrovascular complication in chirotherapy manipulation of the cervical vertebrae]. AB - PURPOSE: Chirotherapy is a popular and successful management option for reversible functional disorders of the cervical spine. Though rarely observed, complications do occur, mainly involving the cerebrovascular system. By means of the here described case and a literature survey, we aim to highlight non cerebrovascular complications of chirotherapeutic cervical spine manipulation. RESULTS: A 43-year-old male initially consulted an ENT specialist, suffering from tinnitus aurium and loss of hearing ability. His hearing significantly increased after intravenous drug therapy, but the tinnitus remained. During chiropractic manipulation of the cervical spine by an orthopaedic surgeon for the tinnitus, the patient described severe neck pain following a clearly audible clicking sound. Scans of the cervical spine prior to and after manipulation showed an intracapsular/intraosseus oedema of the facet joints C2/C3 with lesion of the nerve root C3, most probably induced by chirotherapy. CONCLUSION: Although complications after chiropractic manipulation are extremely rare, treatment of the spine, especially the cervical spine, is not wholly harmless. An adequate history taking followed by clinical and radiographic patient evaluation is necessary to keep the risk of iatrogenic trauma at a minimum. Above all, the chiropractic manipulation of the cervical spine belongs in the hands of a qualified and experienced medical practitioner. PMID- 11253528 TI - [Rope pulley isokinetic system in shoulder rehabilitation--initial results]. AB - AIM: Of this study was to evaluate the results of a shoulder rehabilitation program of different shoulder diseases, based on an isokinetie pulley system ("Moflex", Recotec/Bernina, Swiss). METHOD: In this prospective study 70 patients participated in a standardized rehabilitation program (instability: n = 19; rotator cuff disorders: n = 23; impingement syndrome without lesion: n = 16; others: n = 12; operative therapy: n = 47). The major aspect of the program was an isokinetic pulley system. RESULTS: Isokinetic training with the used device affords strict monitor-feedback to avoid critical torque values. Strength which was attained without relevant pain was almost linearly increased by a mean of 31% until the 20th day of rehabilitation, workload by 79%. At the end of the rehabilitation program the strength of the affected (mostly dominant) shoulder was 15% higher than in the unaffected shoulder; the respective workload values were almost equal. CONCLUSION: These first results demonstrate the value of the isokinetic pulley system in the rehabilitation of the investigated shoulder diseases. The equipment may be used already in an early postoperative state. First results of strength increases using an isokinetic pulley system in shoulder rehabilitation are presented. PMID- 11253529 TI - [The course of uncertain mono-arthritis--a follow-up over 30 years]. AB - INTRODUCTION: Despite intensive investigations the original of a monarthritis may remain unclear. In this present study, relevant parameters for prognosis are sought for such cases. METHODS: In a retrospective analysis 501 records, kept over a period of thirty years, of patients with a first diagnosis of arthritis have been evaluated. RESULTS: The percentage of monarticular arthritis which remained unclear is 48% (n = 246). Of these patients, 63% (n = 318) were male, 37% female. A diagnosis was possible in 26% by puncture, in 47% by biopsy of the synovia. White blood cell count and sedimentation rate were of no help for diagnosis. The great joints were mostly affected, especially the knee (148 cases) and the hip (115 cases). Diagnosis of monarthritis was seldom in younger patients (7%). 119 of the 246 cases of arthritis of unknown origin became pain free (48.4%). Influence of age or sex could not be shown in this group of patients. However, younger patients tended more often to become pain free (73% in the group under 10 years of age). DISCUSSION: Exact history, precise primary examination, laboratory tests and radiological investigations are most important for diagnostic reliability. Whenever possible a puncture of the joint and in persisting cases also a biopsy should be performed. With careful examination more cases should be diagnosed. PMID- 11253530 TI - [Compensation of the role of vitamin D in the skeletal system by calcium]. PMID- 11253531 TI - [World peak of traumatologists in Davos]. PMID- 11253532 TI - [Case-mix adjusted reimbursement system based on the Australian Refined Diagnostic Related Groups (AR-DRG) for orthopedic and trauma surgery clinics]]. PMID- 11253533 TI - [Results of endovascular exclusion of abdominal aortic aneurysms]. AB - According to the principles of minimal invasive surgery a reduced operative trauma and complication rate are the main goals of endovascular aneurysm exclusion. From July 1996 to August 1999 we found 75 patients eligible for endovascular aneurysm therapy. The procedure was primarily successful in 95% of the patients. In the first time we had to switch to open surgery in 4 patients. There was 1 procedure related death. Most frequent later complications were graft occlusion (9.7%) and type I, II and III leakages (14.1%), which were partially repaired using endovascular techniques and partially by use of open surgery. In a selected group of patients the middle term results of infrarenal aneurysm therapy by endovascular stent graft implantation are promising. PMID- 11253534 TI - [Results and complications of endovascular therapy of aortic aneurysms]. AB - INTRODUCTION: Endovascular repair (ER) has been established as an alternative treatment option for aortic aneurysm (AA) in case of a suitable morphology. However, there are specific problems related to diagnostic and therapeutic management including potential complications of the new procedure. PATIENTS AND METHODS: Between 8/1996 and 11/1999, 41 patients (6 female, mean age 67.9 (range 55-84) years) underwent an operation with the intention of ER. Modular, self expanding stent-grafts were used for aorto-biiliacal (36), aorto-monoiliacal (1), and aorto-aortal (infrarenal-1, thoracic-1) aortic aneurysm (AA) exclusion. Postoperatively and during the follow-up period, diagnostic measures included clinical investigation, native X-ray, and color-coded Doppler sonography, and spiral computed tomography, and digital subtraction angiography. Results were analysed with special reference to complications and resulting therapeutic consequences. RESULTS: Technical success was achieved in 36/41 patients (87.8%). There were 2 primary distal endoleaks and 3 conversions because of lacking vascular access. Of 4 primary endoleaks, a proximal one was treated successfully by overstenting, a distal one was sealed off by iliac extension, and 2 distal ones were treated conservatively. Three secondary endoleaks, a proximal and 2 distal ones, required conversion each by retro- and transperitoneal approach. Presently, there are 4 endoleaks, with the maximal aortic diameter remaining constant except one case. Five secondary occlusions of an iliac limb (4) or artery (1) were treated by thrombectomy (1), PTA (1), PTA with overstenting (1), and cross-over (1) or ilicofemoral bypass (1). Three patients died of unrelated disease during the follow-up period. DISCUSSION: On condition of a critical indication, improved diagnostic management and further refinement of stent-graft systems ER constitutes an alternative, minimally invasive treatment option for AA. Long-term results must be obtained by means of continued prospective and comparative studies to definitely evaluate ER. PMID- 11253535 TI - [Intraoperative and early postoperative results after laparoscopic implantation of aortofemoral bifurcation prostheses]. AB - PURPOSE: The evaluation of multiple intra- and early postoperative parameters in patients undergoing laparoscopic aortobifemoral bypass grafting. METHODS: The charts of 22 patients who underwent laparoscopic aortobifemoral grafting between February 2nd, 1996 and April 30th, 1999 were retrospectively reviewed. RESULTS: All patients were men. The mean age was 54 +/- 7.2 years with the mean body weight being 76.3 +/- 10.9 kg and the Body-Mass-Index (BMI) 23 +/- 2.8. Claudication was present in 20 patients; one patient had rest pain and one patient suffered from tissue loss. The mean Ankle-Brachial-Index (ABI) was 0.57 +/- 0.1. In four cases conversion to open technique became necessary. The mean operation time was 316 +/- 73 min and the mean aortic cross-clamp time was 73 +/- 20 min. The mean intraoperative blood-loss reached 689 +/- 461 ml. The mean duration of postoperative ventilator support was 6.0 +/- 5.8 hours and the patients left the ICU after 2.2 +/- 3.2 days. Oral intake was allowed after a mean of 2.6 +/- 2.9 days and the central venous lines were removed after 3.8 +/- 3.9 days. The administration of analgetic drugs was required for 2.9 +/- 3.9 days. The mean length of stay in the hospital was 9.6 +/- 5.5 days. CONCLUSIONS: Laparoscopic aortobifemoral bypass grafting is feasible in a selected group of patients. Despite relatively long operation times and the use of a pneumoperitoneum, we did not encounter significant cardiopulmonary adverse effects. PMID- 11253536 TI - [Laparoscopic aortic surgery in a swine model using a newly designed intestine retraction system]. AB - BACKGROUND: Recently, the feasibility of laparoscopic surgery for aortic occlusive and aneurysmal disease has been demonstrated in clinical studies. However, aortic access is compromised by poor exposure of the operative field from uncontrolled bowel. Currently available retractors are inadequate. The development of new retracting instruments therefore may facilitate laparoscopic aortic surgery. METHODS: Two paddles inserted in a polyester bilayer (mobile device, group A) or a mesh net fixed to the abdominal wall (fixed device, group B) were used to retain the bowel. Six female piglets (28-30 kg) in each group underwent laparoscopic aortic surgery. RESULTS: Mean time to deploy the device was 22 +/- 12 min in group A and 36 +/- 34 min in group B (n.s.). Vascular surgery time averaged 60 +/- 24 min in group A and 68 +/- 16 min in group B (n.s.), the time to withdraw the nets was respectively 3.6 +/- 1.2 min and 13.5 +/- 8.2 min (p < 0.05). Total surgery time was 155 +/- 41 min vs. 174 +/- 49 min (n.s.). Two retraction failures have been registered (1 in group A and 1 in group B). No major complications were documented. CONCLUSIONS: Both nets provided adequate exposure of the infrarenal aorta. Vascular surgery time and blood loss were similar in both groups. Nevertheless, the handling of the mobile device (group A) seemed to be more comfortable in direct comparison. The newly developed retraction devices might facilitate the performance of laparoscopic aortic surgery. PMID- 11253537 TI - [Endoscopic aortofemoral bypass reconstruction: experimental and clinical results]. AB - According to the present experiences, the new videoendoscopic vascular instrument systems have fulfilled the basic requirements for performing videoendoscopic aortoiliac surgery. The animal studies (n = 25) showed clearly, that the piglet is a reproducible and favourable model to acquire the basic surgical techniques for videoendoscopic aortoiliac surgery, since its retroperitoneal anatomy is similar to that of humans. The transabdominal-retroperitoneal approach on human cadavers (n = 6) was soon abandoned, since particularly in obese cases it caused burdensome sliding of the intestine into the operative field adjacent to the renal vessels. The intensive investigations on human cadavers (n = 20) has meanwhile allowed us to constantly expose the aorta above the inferior mesenteric artery up to the left renal vessels without the need to sacrifice the inferior mesenteric artery (total operating time: 3-4.5 hours, aortic reconstruction averagely 42 minutes). The prolonged operating time during the extraperitoneal approach, especially in case of accidental tearing of the retroperitoneum has led us to evaluate the transabdominal paracolic approach (n = 6). This procedure with right-sided positioning of the patient offers a broad operating area and retains the intestine in the right side of the abdomen throughout, thus reducing the operating time in our initial 6 cases to a mean of 2.9 hours (range: 2.5-3.5 hours). As to our limited clinical experience and in accordance to other surgical groups the video assisted aortoiliac surgery (n = 12) by performing a small abdominal incision for the aortic anastomosis under direct vision seems to be an alternative procedure to the total videoendoscopic aortoiliac surgery (n = 7). Further experimental research, consistent development and refinement of videoendoscopic vascular instruments and prospective evaluation of the different videoendoscopic approaches to the aortoiliac vessels are necessary to evaluate the use of the videoendoscopic surgical technique, to clarify its advantages and to probably generalize the procedure in the near future. PMID- 11253538 TI - [Clinical experiences with 43 cases of laparoscopic reconstruction in aortoiliac occlusive diseases: analysis of morbidity, effectiveness and treatment results]. AB - Between October 1995 and April 2000 we performed 30 aortic laparoscopic reconstructions and 13 procedures on the iliac vessels. Severe aortic calcifications, poor cardiac and pulmonary status, and extensive intraabdominal adhesions were considered as contraindications for the laparoscopic procedure. Operative time, conversion rate, surgical and cardiopulmonary complications were recorded for an intention- to treat-analysis. Clinical criteria and Duplex ultrasound scans were used to document the patency of the grafts. The duration of analgetic therapy, nursing care, and hospital stay characterized the postoperative recovery. Mean surgical time was 302 minutes (min) in the aortic group (Ao) and 251 min in the iliac group (Ia). Seven conversions (5 in the aortic and 2 in the iliac group) occurred for clamping problems (n = 3), aortic lesion (n = 1), iliac vein lesion (n = 1), time consuming dissection (n = 1), and suturing failure (n = 1). Cardiac morbidity rate was 6.6% (Ao) and 7.7% (Ia). Two patients suffered pulmonary complications after aortic surgery (6.6%). Mean duration of analgetic therapy was 2.4 d (Ao) and 2.5 d (Ia). Nursing care was necessary for 4.4 d (Ao) and 3.1 d (Ia). At discharge the reconstructions were patent and ischemia symptoms subsided in all cases. Mean hospital stay was 9.1 d (Ao) and 6.7 d (Ia). At follow-up after a mean interval of 5.2 months (Ao) and 8.3 mo (Ia) patency was documented in all cases. Disease progression in the adjacent native vessels (n = 3) and beyond the graft (n = 1) was revealed by ultrasound scan, requiring a further vascular procedure. In our laparoscopic experience cardiac and pulmonary morbidity proved similar to traditional vascular surgery. The advantage of the minimally invasive approach seemed to be a more comfortable recovery. However, operative time and conversion rate emphasize the technical challenge of the procedure, which should be performed only in highly selected cases. PMID- 11253539 TI - [Hemobahn endoprosthesis: experience with percutaneous placing in the arteria femoralis superficialis]. AB - The self-expanding Polytetrafluoroethylene-lined nitinolstent Hemobahn offers from its design a prophylaxis for re-occlusion after angioplasty of long femoral artery occlusions. During the last two years we placed in 33 patients at 35 extremities a total of 53 Hemobahns. Fontaine's clinical stages were: 30 x IIb, 1 x III and 4 x IV. The treated lesions were femoral stenoses (3x) and occlusions (32x with mean distance 12 cm). Follow-up-visits were after 3, 6 and 12 months. The mean ankle-brachial-index changed from 0.49 to 0.85, post-interventional Fontaine's stages were: 24 x I, 5 x IIa, 2 x IIb and 4 x IV. The primary inhospital patency rate was 91%. After 3 months all primary/secondary patency rates were: 77%/87%, after 6 months 62%/85% and after 1 year 40%/67%. Causes for re-occlusions were a poor "run off", calcifications and multiple vessel lesions. The results for "ideal indications" were much better. Such "ideal indications" applied to: > or = 1 cm "healthy" proximal and distal vascular segments, missing lesions in the popliteal artery, at least one open lower leg vessel and no severe calcifications. The primary/secondary patency rate was inhospital 100% (n = 23), after 3 months 95/100%, after 6 months 85/100% and after 1 year 80/100%. Side effects were local haematoma, pain in the prosthesis and temporarily temperature reactions. Sonographically no re-stenoses at the limb ends were observed. Hemobahn-endoprostheses are suited for therapy of long femoral artery occlusions. The medium patency rate of "ideal indications" is higher than that of angioplasty with or without uncovered stents. PMID- 11253540 TI - [Femoro-distal ePTFE bypass grafting using femoro-crural patch prosthesis (FCPP). Results of a prospective clinical study]. AB - Femorodistal bypass using exclusively PTFE is known to have a poor prognosis, mostly because of the development of myointimal hyperplasia (MIH). Several vein patch techniques are established but the role of hemodynamics within the anastomotic site has only been explained insufficiently and is hardly considered clinically. In a prospective study, between 6/1992 and 7/1998 129 patients (89 m/40 f, mean age 65.2 +/- 10.0 years) with critical limb ischemia and no usable saphenous vein were included to undergo femorodistal ePTFE bypass grafting with a new, hemodynamically optimized distal end-to-side anastomosis. Patients were followed at 6-month intervals with clinical investigation and color-coded Doppler sonography. Primary and secondary graft patency (PPR, SPR), limb salvage, and patient survival were calculated according to Kaplan-Meier. With a median follow up of 45 (range 6 to 72) months, PPR and SPR at 1, 3 and 5 years were 63.0, 35.7 and 27.6% and 74.5, 44.8%, and 37.6%, respectively. Limb salvage at 1, 3 and 5 years was 86.4%, 78.7% und 73.2%. There was no perioperative mortality. Graft infection occurred in 7 patients (5.2%). ePTFE bypass grafting represents a valuable option for infragenicular and crural reconstruction in the absence of autologous vein. The new anastomotic design was feasible and represents another adjunct to possibly improve patency of femorodistal bypass allografts. PMID- 11253541 TI - [Biodegradation of a PTFE prosthesis]. AB - Processes of biodegradation of PTFE--as they were not seen before for this type of graft--are demonstrated by clinical examples. As it is possible to subsume these results to regularly described processes of incorporation of alloplastic material, it is to be expected that there will be more cases than those observed by us up to now. PMID- 11253542 TI - [Revascularization of the profunda femoris artery in ischemia of the stump after above knee amputation]. AB - We describe a severe complication after above knee amputation: persisting ischemia of the stump, and its treatment by vascular surgery. Only 4 (2.3%) of 168 above knee amputations performed in our department between June 1985 and September 1999 were followed by ischemia of the stump non responding to conservative treatment. We decided on subsequent revascularization of the ipsilateral deep femoral artery. A femorofemoral bypass graft was successfully implanted. A reamputation or an exarticulation of the hip joint could be avoided. We call special attention to the importance of the deep femoral artery. PMID- 11253543 TI - [Our surgical heritage: the surgical school of Halle]. AB - The knowledge of the historical development of surgery including its increasing specialization is important for our efforts to preserve the unity of surgery. In the past distinguished surgeons essentially influenced the progress of surgery. The list of the directors of the surgical department of the Halle university hospital in the last 150 years with the outstanding Richard von Volkmann is an excellent example for this influence and will be presented here. PMID- 11253544 TI - [Prevention of postoperative wound infections. Evidence-based recommendations]. AB - Among all hospitalized patients, surgical site infections (SSI) are the third most frequently hospital-acquired-infection. SSIs remain a substantial cause of morbidity and mortality among surgical patients. This may be partially explained by the emergence of antimicrobial-resistant pathogens and the increased numbers of patients who are elderly and/or have a wide variety of chronic, debilitating, or immunocompromising underlying diseases. This is why it is essential to implement SSI prevention measures. In April 1999 the Centers for Disease Control and Prevention (CDC) presented the "Guideline for Prevention of Surgical Site Infection". The recommendations represent the consensus of the Hospital Infection Control Practices Advisory Committee (HICPAC) regarding strategies for the prevention of SSIs. Whenever possible, the recommendations are based on data from well-designed scientific studies. This guideline is a major step forward and is also essential to optimize the management of surgical patients in Germany. PMID- 11253545 TI - [Location of the aortic bifurcation in man and its practical significance in vascular surgery]. AB - Based on analysis of arteriography in 300 subjects of different age and sex, as well as 4 human fetuses, we demonstrated that in the course of an individual's life aortal bifurcation moves caudally. In a fetus, bifurcation is located at the level of L3, while in an adult it moves to the level of L4. After the age of 50, the position of bifurcation moves gradually caudally and can reach--in the advanced age--as far as the level of L5/S1 disc. The changes of position during a lifetime are statistically significant. A shift in the position of the bifurcation of abdominal aorta can be of significance for surgical interventions on the abdominal aorta. PMID- 11253546 TI - [Results of surgical therapy and classification of non-ruptured abdominal aortic aneurysms]. AB - INTRODUCTION: Surgery for symptomatic aortic abdominal aneurysms (sAAA) is associated with an increased mortality and morbidity compared to asymptomatic aortic aneurysms (aAAA). With the advent of endovascular therapy, an alternative therapeutic modality has become available. Endovascular therapy, however, depends on certain morphologic criteria, whereas open surgery can be performed on any type of AAA. The purpose of this study was to analyse our data of surgical treatment of non ruptured AAA and to identify the amount of patients in whom endovascular therapy would have been possible. METHODS: Retrospective analysis of the medical data of all patients operated upon non ruptured AAA in our department by 3 responsible vascular surgeons from 1995-1999. RESULTS: 225 consecutive patients with a median age of 65 (42-95) years were included in the study. There were 184 (82%) male and 41 (18%) female patients with 143 (63.5%) aAAA and 82 (36.5%) sAAA. Patients with sAAA underwent emergency aneurysm repair and had a significantly increased aneurysm diameter compared to the aAAA, who underwent elective surgical aneurysm repair (6.9 +/- 1.6 cm vs. 6 +/- 1.2 cm; p = 0.002). A total of 11 (4.9%) patients had an inflammatory AAA. Smoking was found to be the only significant increased preoperative risk factor in the group of sAAA compared to aAAA (91 vs. 35 patients; p = 0.008). Morbidity was significantly increased in the patients with sAAA compared to the aAAA (55% vs. 31.5%; p = 0.041) The mortality however did not differ significantly in the two groups (2 vs. 3 patients; p = 0.691). Considering morphological criteria of the AAA, endovascular therapy would have been possible in 59 (26%) patients. However, in 24 (11%) of the 59 patients, endovascular therapy was not feasible because of aortic kinking, heavy calcification of the aneurysm neck, a patent inferior mesenteric artery or atherosclerotic diseased iliac arteries. Consequently, only 35 (15%) patients would have qualified for an endovascular therapy. DISCUSSION: Surgical therapy can be performed in patients with asymptomatic and symptomatic AAA with an equal low mortality. This finding underlines the fact, that surgical therapy still remains the standard therapy for AAA. In addition, in our study only a relative small amount of patients would have qualified for an endovascular therapy. PMID- 11253547 TI - [Transplantation surgery. I]. PMID- 11253548 TI - [DEGUM standards for sonographic examinations in gynecology and obstetrics]. PMID- 11253549 TI - [3D-sonography as "2D-M-mode sonography in fetal echocardiography]. AB - AIM: Establishing a technique employing 3D-ultrasound for the echocardiographic examination of foetuses. METHOD: M-mode sonography has reached considerable importance in the detection of arrhythmia as well as pathological changes of the heart valves and contraction disorders of the foetal heart. The variability of the position of the foetus leads to problems typically arising with this method. The ventro-anterior position along the long axis of the foetal heart, being the optimal position for prenatal echocardiography, allows no examination in M-mode because the plane of the valve lies vertically in relation to the direction of the ultrasound. A special appliance of 3D-sonography solves the problem by providing a 2-dimensional examination technique of the heart in M-mode independent of the position of the foetal heart. The method allows simultaneous measurement of the movement of different parts of the myocardium in several different planes. By fixing the position of the applicator (in B-mode) and choosing the 3D-sequence in the "free-hand-mode" one gets a 2-dimensional time tissue-block which contains all movements in the chosen axis. These "time planes" can be deliberately selected for the examination of different planes in the fixed B-mode image which shows their variations along the time axis. RESULTS: Our study describes the application and evaluation of this examination during pregnancy between the 28th and 40th week. Reliable images of the AV-valves or the aortic valve could be produced in 73% of examinations. PMID- 11253550 TI - [Single umbilical artery. Effect on fetal growth and doppler flow velocity wave form]. AB - OBJECTIVE: Fetuses with single umbilical arteries (SUA) are often small for gestational age (SGA). We tried to clarify the following questions: 1. In which range of the normal reference chart for 2 umbilical arteries are the S/D-ratios of SUA located? 2. Is it possible to correctly predict the fetuses at risk for asphyxia or fetal death using these reference charts in SNA fetuses? 3. How do placental weight and histological findings in the fetoplacental vessel influence flow patterns in SUAs? 4. How does the vessel diameter of SUAs influence fetal growth? PATIENTS AND METHODS: 25 fetuses with SUA were examined by means of ultrasound and Doppler ultrasound 1 to 9 times with measurements of arterial diameter and S/D-ratio. These data were compared with results of normal umbilical cords (data from the literature) and correlated with fetal outcome and histological placental findings. RESULTS: Out of 94 individual measurements the S/D-ratio was found to be below the 10th percentile in 28 cases (30%), between the 10th and 50th percentile in 31 cases (33%), between 50 and 90th 20 cases (21%), and above the 90th percentile of the reference chart in 15 cases (16%). 14 fetuses were SGA, 2 presented an intrauterine death, 2 were born with chromosomal aberrations and 3 with malformations. 5/6 fetuses with caesarean section for asphyxia or fetal demise were correctly detected to show an elevated S/D-ratio, 1 case of intrauterine death was not. 16/19 placentas were found to be of low weight, in 14 we found villous malmaturation. 18/22 examined SNAs had an elevated diameter above the +2s range of the reference chart for normal arteries. The ratio of the diameters of the umbilical vein and artery was above 2 in 12/14 SGA fetuses, indicating that the SNA's diameter was less than 50% of that of the veins in these cases. CONCLUSION: Widening of the SNA lumen leads to a reduction of the S/D-ratio. An SNA diameter of less than 50% of that of the vein results in intrauterine growth retardation. Elevated S/D-ratio correctly identifies SNA fetuses at risk. PMID- 11253551 TI - [Breast cancer recurrence versus scar. Ultrasonographic differentiation using Levovist as the contrast medium]. AB - AIM: To determine the scope of improving the distinction between a postoperative scar and the recurrence of a breast carcinoma through the use of the ultrasound echo enhancer Levovist? METHOD: In 23 patients with 26 lesions a colour-coded duplex sonography examination before and after administration of Levovist was performed. The parameters investigated were: degree of enhancement, number of tumour vessels and the pattern of vascular morphology and anatomy. RESULTS: Recurrences (n = 15) demonstrated a greater number of vessels and a stronger enhancement after administration of Levovist. Individual vessels were also visible in scars (n = 11). Further evaluations with respect to the pattern of the tumour vascularization are therefore necessary with the exception of one false positive and one negative result a clear distinction was possible. CONCLUSION: The administration of the ultrasound echo enhancer clearly improved the otherwise difficult distinction between a scar and a tumour recurrence through sonography and mammography. Further studies with larger number of patients are necessary to establish the value of the method. PMID- 11253552 TI - Anatomical variations in the internal jugular veins of cancer patients affecting central venous access. Anatomical variation of the internal jugular vein. AB - PURPOSE: Establishing a reliable central venous access is an important procedure in clinical haematology and oncology. The purpose of this study was to determine how anatomical variations in the internal jugular vein (IJV) and its position in relation to the common carotid artery (CCA) in cancer patients affects external landmark puncture. PATIENTS AND METHODS: In 113 patients with haematological or oncological diseases we examined sonographically potential target regions for placement of a central catheter via the IJV. RESULTS: 36% of our patients showed anatomical variations in the IJV and surrounding tissue. CONCLUSIONS: External landmark puncture may be difficult in a considerable number of patients since the IJV might not be situated in the presumed location anteriorly or laterally to the CCA, or a normal lumen may not be present in approximately 1/3 of cancer patients. This study supports the use of ultrasound-guided techniques for central venous catheters particularly in haematological and oncological patients. PMID- 11253553 TI - [Sonographic findings in interstitial lung diseases]. AB - Up to now, sonography of the thorax has not been taken into consideration in cases of interstitial lung disease because of its technical limitations. This study was aimed at examining pleural and subpleural alterations in cases of interstitial lung disease and comparing the sonographic method with the usual imaging procedures such as x-ray and computer tomography. PATIENTS AND METHODS: 24 patients, aged 25 to 70 who were diagnosed as suffering from an interstitial lung disease and underwent sonography of the thorax, were analysed with regard to the following criteria: 1 pleural effusions, 2 pleural fragmentations, 3 subpleural infiltrations > 2 mm, 4 pleural nodules. RESULTS: Six patients were diagnosed to have small pleural effusions which had not been visible on x-ray scans. 14 patients showed pleural fragmentations, 10 patients had subpleural infiltrations, and in one patient pleural nodules could be detected. Out of the 24 patients suffering interstitial lung disease, sonographic alterations were diagnosed in 17 cases (= 71%). Among the 9 patients definitively suffering from sarcoidosis, five had had a completely inconspicuous ultrasonic result, four of them with sarcoidosis stage I. CONCLUSION: Sonography of the thorax has proved to be an excellent complementing examination method in cases of interstitial lung disease. The advantages lie in the possible detection of small pleural effusions and small subpleural infiltrations, where this method can also be used to monitor therapy. PMID- 11253554 TI - [Simultaneous transthoracic echocardiography and transcranial doppler ultrasonography with ultrasound contrast agents for the detection of a patent foramen ovale in diving medicine]. AB - OBJECTIVE: Presentation of a non-invasive method for the identification of a patent foramen ovale (PFO) in screening of diving-candidates or for the investigation of diving accidents. METHOD: Simultaneous transthoracic echocardiography and transcranial Doppler sonography of the middle cerebral artery using ultrasound contrast media. RESULTS: In an unselected collective of amateur and professional divers a PFO was identified according to statistical prediction. CONCLUSION: This method can be used in diving medicine for PFO detection as an alternative procedure to transesophageal echocardiography. PMID- 11253555 TI - [Optimal age for hip sonography screening]. AB - In the German Federal Republic the guidelines from 1996 for screening by hip sonography recommend a practice in two parts: at birth (3rd to 10th day) examination of newborn babies presenting risk factors. Lateron (4th to 5th week) screening of all babies regardless of previous findings. The effectiveness of this splitting was not proven at that time, when these guidelines were introduced. In our experience over 15 years in hip sonography of nearly 15,000 babies, the screening of all babies immediately after birth--regardless of risk factors--shows to be effective as well as economic. PMID- 11253556 TI - [Technical errors in the application of Graf's hip sonography method]. AB - AIM: The aim of the study is to point out possible examination errors because of a titled transducer, and to analyse their sources. METHODS: The results obtained by experienced examiners using the correct examination technique were compared to results obtained under defined deviations from the standard. With the use of a computer model the course taken by the ultrasound waves in the neonatal hip was simulated and the sonographic image reconstructed. RESULTS: Twisted as well as tilted positions of the transducer lead to significant errors in alpha, the acetabular roof angle. This is in accordance with the computer model predictions. Caudo-cranially tilted probes lead to false-positive, therapeutically relevant incorrect diagnoses. CONCLUSION: The computer simulation predicts a high degree of accuracy of hip sonography in Graf's technique under standard conditions. The effectiveness of sonographic infant hip screening largely depends on the ability to adhere to the standardized approach, minimizing possible errors. A mechanical transducer guide prevents tilting errors by reducing the degrees of variability in positioning the transducer. PMID- 11253557 TI - [Color-coded doppler sonographic demonstration of intrahepatic venous collaterals in Budd-Chiari syndrome]. AB - By means of colour Doppler ultrasound, veno-venous shunts were found in 3 cases, leading to the diagnosis of Budd-Chiari Syndrome. Pulsed and colour Doppler ultrasound showed a reduction in venous blood flow and the reversal of blood flow in the venous collaterals. Venous spectral Doppler wave forms in the veno-venous shunts were flattened and aphasic in all cases. Sonographic findings were confirmed by cavography. Our cases show that pulsed and colour Doppler ultrasound are a valuable tool in the initial diagnosis of veno-venous shunts associated with Budd-Chiari Syndrome. PMID- 11253558 TI - The role of Doppler velocity histograms of hepatic veins in early detection of cirrhotic changes of the liver parenchyma. AB - AIM: In a prospective study we measured the velocity ranges of the Doppler Velocity Histogram (DVH) of the hepatic venous system at two levels (-3 dB and -6 dB) below the maximum power level of the Doppler wave in order to investigate if that allows a higher selectivity between the Doppler tracings in hepatic veins of patients with liver cirrhosis and healthy subjects. METHODS: The DVH was measured in 23 healthy subjects and 31 patients with liver cirrhosis of different etiologies at power levels of -3 and -6 dB in the right and middle hepatic vein after an overnight fast. The DVH measurements were performed at the maximum of phase I of the hepatic venous flow in which we assessed the Peak-, Mode-, Mean Value, and the velocity range (bandwidth). RESULTS: At both power levels the bandwidth (BW) of the DVH in the right and middle hepatic vein was significantly higher in cirrhotics than in healthy subjects (-3 dB: RHV: p = 0.048, MHV: p = 0.006; -6 dB: RHV and MHV: p < 0.0005). The selectivity between healthy subjects and cirrhotics is higher at the -6 dB level than at the -3 dB level. CONCLUSION: The DVH-measurement is a useful additional device in early sonographic detection of cirrhotic liver parenchyma changes. DVH-velocity range measurements at a level of -6 dB below the maximum power level reveal a better selectivity between healthy subjects and cirrhotic patients than measurements at the -3 dB level. It is recommendable to perform velocity range measurements at different power levels within a single frozen image. PMID- 11253559 TI - [Evolution and function of sleep]. AB - This paper presents a brief historical review of the beginning of scientific studies on sleep research and evaluation of the electrophysiological basics. Subsequently, hypotheses are presented and discussed on phylogenesis and function of NONREM- and REM-sleep as well as aspects of the regulation of energy conservation and temperature. Sleep may have first evolved out of rest/activity cycles. Later on, with the evolution of complex brains, sleep may also assume importance in development and maintenance of these structures. Hence, special emphasis is placed on the influence of NONREM and REM sleep on the reorganisation of neuronal networks. PMID- 11253560 TI - [Somatization--somatoform disorders--etiopathological models]. AB - A multifactorial model is obligatory to comprise the etiopathogenesis of somatization. Thereby, different aspects of genetics, neurology, neurophysiology, psychophysiology, endocrinology, psychology of personality, perception and cognition, social learning, knowledge of illness, public conceptualisation of illness, negative life events, chronic psychosocial stressors, reduced coping skills, lacking supporting systems, systems of social reinforcement, coexistence/comorbidity of psychiatric disorders, traumapsychology and psychodynamics can be outlined. PMID- 11253561 TI - [Treatment courses and therapeutic effectiveness of a psychogeriatric day hospital]. AB - Since 1976, 37 psychogeriatric day hospitals have been set up in Germany. As only few empirical studies on the effectiveness of this day hospital system are available even 25 years on, the treatment course of 58 patients attending the psychogeriatric day hospital run by the Department of Psychiatry, University of Tubingen was evaluated. The course analysis was based on a series of tests comprising the following questionnaires: Social Situation Scale, Activities of Daily Living (ADL), Instrumental Activities of Daily Living (IADL), Munich Quality of Life Dimension List (MLDL), Mini Mental Status (MMS), Hamilton Depression Scale, and Cumulative Illness Rating Scale (CIRS). Women accounted for three-quarters of the sample, and men for one-quarter. The average age was 72 years. The fact that only 16% of the patients had been transferred from psychiatric hospitals and that more than half had been referred directly by their local doctors underlines the acute-psychiatric nature of the institution. More than 80% of the patients were discharged to their previous place of residence. A highly significant improvement in affective state was verified by the Hamilton Depression Scale. Significant improvements were also registered in the following items: general satisfaction with life, state of health, physical and mental performance, and scope for personal development. The quality of life, measured with the Munich Quality of Life Dimension List (MLDL), improved in nine of the nineteen items covered, with correction of the significance level alpha taken into account. The treatment outcomes confirmed the therapeutic effectiveness of this type of psychogeriatric institution from aspects of acute psychiatry too and should encourage the continued expansion of partial hospitalization facilities for the elderly. PMID- 11253562 TI - [Monotherapy of Parkinson's disease with budipine. A double blind comparison with amantadine]. AB - In a randomised double blind parallel-group study in three centers budipine, a diphenylpiperidin derivate, was compared to amantadine with respect to efficacy and safety in the monotherapy of mild to moderate Parkinson's disease (PD). From 53 patients of either sex 27 patients were randomised to 3 x 20 mg/d budipine and 26 patients to 3 x 100 mg/d amantadine. The duration of treatment was 4 weeks in 1 center (21 patients) and 12 weeks in the other 2 centers (32 patients). Safety was measured by vital signs, ECG, adverse event recording and clinical laboratory. Both drugs caused a clinically relevant and statistically significant (p < 0.001) improvement of Parkinsonian symptoms according to the Webster-Rating Scale (WRS) as compared to pretreatment values. With respect to the total WRS score sum there was no difference between the groups (p > 0.05; n.s.), while budipine showed a significantly (p < 0.05) better effect on the main symptom tremor after 12 weeks. During amantadine treatment more adverse events were observed than after budipine intake. Two patients left the study prematurely, one in the amantadine group due to psychiatric adverse events and one in the budipine group because of insufficient efficacy. PMID- 11253563 TI - [Deprivation versus importation: a model explaining the increase of suicide rates in custody]. AB - High suicide rates in jail, lock-up or prison settings have given rise to a debate about whether suicides result chiefly from the type of people confined, or from the types of places they are confined in, the types of confinement. This is summarily framed by the terms of an associated debate in criminology, between importation and deprivation theory. This paper describes the importation versus deprivation theory, concerning the circumstances in Austrian prisons and jails. The article reports on all completed suicides over the period from 1947 to 1999 (n = 410). The increase of suicide rates in Austrian jails and prisons is significant over the last fifty years. While the rate was stable between 1947 and 1975, we have a significantly increasing rate since 1975. In 1975 there was an important legislational reform of the criminal law in Austria. The implications of this reform are discussed in the light of the importation/deprivation theory. PMID- 11253564 TI - [Measurement of the outcome quality as beginning of total quality management in otorhinolaryngology]. AB - BACKGROUND: The last years are characterized by a change of paradigms in the evaluation of medical therapeutical strategies. The term quality of life (QoL) has been established in oncology and could be demonstrated as a subject of highly qualified investigations which were published during the last years. An extendend quality management is correlated with the possibility of cost-effectiveness measurement and arrangement in a time characterized by decreasing financial resources of the hospitals and university departments. The experiences in the field of quality analysis could offer precious tools to optimize structures, processes and results of treatments. This aim is strongly supported by the expected law "Gesundheitsreform 2000". METHOD AND RESULTS: Therefore reliable, valid tools are needed to calculate the effects of therapies. The combination of global and specific questionnaires include the possibility to compare oncologic data of different entities of malignomas. A couple of questionnaires in otorhinolaryngology were developed in the last decade involving the fields of malignomas of the upper aerodigestive tract, rhinosinusitis etc. CONCLUSIONS: Ishikawa, the founder of "Total Quality Management" and of the "Quality Circ", pointed out the high importance of customer satisfaction. Whereas the patient is in the mid point of medical treatment there are hardly any investigations about customer satisfaction published. Therefore it is necessary to develop and establish validated tools for the measurement of quality dimensions. The short, middle and long-lasting effects of a treatment could be calculated by quality of life measurements. The QoL is here of very strong importance, especially since economic aspects are included. PMID- 11253565 TI - [The Interesting Case No. 42. Differential diagnosis of epistaxis]. AB - Epistaxis is a symptom and one of the most frequent medical emergencies. In most cases haemorrhages concern the anterior parts of the septum, in particular the Locus Kiesselbachi. Thus they are harmless and therapy is easy to handle. We report a case of a 55-year-old lady with relapsing epistaxis due to a pseudoaneurysm after surgery of a meningioma of the sphenoid bone. This type of epistaxis is rare and may culminate into a life-threatening event. The case demonstrates the importance of an exact differential diagnostic evaluation by use of modern imaging techniques for severe and life-threatening symptomatic nose bleeding. PMID- 11253566 TI - [Recommendations for the Provincial Chambers of Physicians of the Federal Republic of Germany on the handling of applications for authorization to conduct continuing education in ENT specialty]. PMID- 11253567 TI - [Visualization of cochlear implant electrode movements in the cochlea by using x ray microscopy]. AB - BACKGROUND: The movements of the electrode cable of a cochlear implant inside the cochlea during the insertion procedure generally are invisible, even in temporal bone experiments. Yet, the development of new designs of electrodes, their positioning near to the modiolus etc. requires an exact knowledge on the dynamic behaviour of the electrodes inside the cochlea. The exclusive method for looking through the undamaged cochlea walls is by x-ray technique. METHODS: A specific x ray tube with a focal spot size of barely 5 microns allows useful direct magnifications of more than 20-30 times. We performed temporal bone experiments with this tube and studied the behaviour of conventional and perimodiolar electrode cables of a MedEl-Cl during the insertion procedure under fluoroscopic viewing. RESULTS: The high resolution imaging revealed the influence of stiffness of the electrode cable on the insertion depth of a conventional electrode. If the angle of the direction of insertion was too steep, the resulting kinking of the cable at the medial wall of the basal turn was instantly visible. The direct visualisation of the movements of the cable was useful in optimizing the design and stiffness of various prototypes of perimodiolar electrodes for a deep insertion. CONCLUSIONS: The dynamic examination technique disclosed that the stiffness of actual electrode cables is not optimally adapted to the form of the cochlear cavity for deep insertion. The non-destructive visualisation technique will facilitate the development of advanced electrode design, especially for various concepts of perimodiolar electrodes. PMID- 11253568 TI - [Intraoperative measurement of stapes mobility using a hand-guided electromagnetic probe]. AB - BACKGROUND: The hearing results of otosurgery are still unsatisfactory. Even after successful implantation of middle ear prostheses there often remains an air bone gap of 30 dB or more. As possible reasons dislocation of the prostheses due to scar growth, changes in prostheses' attachment or ventilation disorders are being discussed. Decreased stapes mobility, which has been judged only manually up to now, is supposed to be a further reason. METHOD: We are introducing a new electromagnetic probe. The output signal of this device is proportional to the impedance of the stapes-annular ligament cochlear fluid system at the sensor's resonance frequency (2.4 kHz). The advantage of this system is characterised by its hand-guidance. Injury of the sensitive stapes-annular ligament due to tremor movements of the surgeon can be excluded using a special construction of the sensor head. The maximum force of the sensor's tip onto the stapes during measurement is limited to below 5 mN. RESULTS: Preliminary measurement results of 20 patients are presented with normal and abnormal stapes mobility. These results are compared to the subjective impression of the surgeon, who usually tested the stapes mobility by hand. As a result of our investigations probe measurements can detect more exactly decreased mobility of the stapes than the surgeon. CONCLUSIONS: Our device may help to detect latent stapes fixation caused by chronic inflammation of the middle ear. The intraoperative measurement of stapes mobility may influence the strategy of the surgeon. Furthermore it would be helpful in patient consulting prior to a revision-tympanoplasty with predicting the potential hearing improvement. PMID- 11253570 TI - [Normal sense of smell in Kallmann syndrome. A case report]. AB - BACKGROUND: Kallmann's syndrome (KS) was first mentioned in 1944 as an association of anosmia and hypogonadotropic hypogonadism. Causes are multiple genetic defects the most common of which is the x-linked KS appearing mostly in men. However, autosomal dominant and autosomal recessive forms have also been described. PATIENT: We present a case of KS with normosmia (male, 39 years of age). All symptoms of hypogonadotropic hypogonadism were present. RESULTS: Psychophysical olfactory testing revealed left-sided anosmia with right-sided normosmia which was confirmed by electrophysiological measures of olfactory function. Magnetic resonance imaging indicated aplasia of the left olfactory tract and bulb, whereas the right-sided structures appeared to be normal. CONCLUSIONS: As indicated in this case with lateralized anosmia and contralateral normosmia, overall olfactory function strongly depends on the "best" nostril. Therefore, in many clinical situations, lateralized olfactory testing appears to be extremely important. PMID- 11253569 TI - [Extended indications for decompression of the optic nerve: a differentiating analysis of restriction of the visual function, also in comatose patients]. AB - BACKGROUND: The care of indirect traumatic optic neuropathy is still treated differently. Special diagnostic and therapeutic difficulties exist in comatose patients without definable visual acuity and in patients with complex failure of the visual field with central visual acuity still receiving sufficiently. METHOD: The total collective of optic nerve decompressions within the period between January 1992 and August 2000 was comprised of 66 patients. 18 of these 66 patients (average age: 40.5 years) were comatose and required critical care. 3 of the 48 consciousness-clear patients showed post-traumatically different defects of the visual field with a visual acuity of 0.3, 0.4 and 1.0. The indication for surgical decompression in both groups was based on the ophthalmological findings and the CT-finding of traumatization of the optic nerve, or the orbit apex respectively. RESULTS: During the subsequent postoperative examinations (on average 12.3 months postoperatively) five patients within the group of the unconscious patients showed a normal visual acuity (0.9-1.0), 3 patients a visual acuity of 0.7, 0.4 and 0.3 and one a visual acuity of 0.1. Six patients remained amaurotic. Three patients died from the general consequences of injury. Improvement of visual field and correction of stereoscopic function occurred in all patients, operated on primarily because of the described visual defects. CONCLUSIONS: Due to these results the indication for the decompression of the optic nerve should find special consideration in comatose patients and in cases of severe restriction of the visual field. PMID- 11253571 TI - [Open surgical treatment of laryngo-tracheal stenoses in children]. AB - BACKGROUND: Treatment strategy in laryngo-tracheal stenoses in children has for a long time been conservative treatment with tracheostomy or bougination in hope of a more or less spontaneous resolution of the stenosis during growth of the child. The results of this option as well as the endoscopic treatment with different laser systems has proved to be rather disappointing. A child with a tracheostomy means a heavy load for the parents to look after this child as well as a constant threat from complications by displacement or plugging of the cannula. METHOD: A retrospective chart review of 22 children, aged between two months and 15 years at the time of surgery with laryngo-tracheal stenoses treated by different open surgical procedures. RESULTS: The aetiology of the stenoses was prolonged endotracheal intubation in 12 children, congenital stenoses in 3 children, unsuccessful laser treatment for acquired stenoses in 3 children, subglottic hemangioma in 3 children and a transglottic cyst in 1 child. 17 cases were treated by laryngo-tracheal reconstruction with rib cartilage graft, 3 crico tracheal resections, and 2 laryngofissures. Five children could be treated without tracheostomy. From the remaining 17 cases 13 could be decannulated, 1 child died one week after surgery from his congenital heart disease. 3 children are still with a tracheostomy, two of them had had endoscopic laser therapy alio loco before. CONCLUSIONS: Open laryngo-tracheal surgery for paediatric airway stenoses is a successful treatment option besides endoscopic management for selected cases. Resection surgery seems to be indicated for severe stenoses with proliferative scar tissue formation. Depending on personal experience and post operative facilities procedures without tracheostomy but prolonged post-operative intubation are possible single-stage-solutions. PMID- 11253572 TI - [The enjoyment of red wine is influenced by the shape of the wine glass]. AB - BACKGROUND: It is frequently claimed that the glass shape has a direct impact on wine aroma. METHODS, PATIENTS: Healthy volunteers (43 m, 46 f, age 19-60 years) tried 3 glasses. Three glasses of different shape but of the same height and of comparable opening diameter were used. Glasses were of "tulip"-like, "beaker" like, and "egg"-like shapes. Intensity, hedonic tone, and quality of a red wine were rated before and after drinking. RESULTS: Both intensity and hedonic ratings of wines from different glasses were influenced by glass shape. Glass shape also influenced the complexity of wine odors. This appeared not to be related to the esthetic impression the glasses made. CONCLUSIONS: The present data indicate that egg-shaped glasses, compared to "tulip" or "beaker" glasses, appear to produce higher intensity and higher complexity of wine odors. This may relate to the trapping of odors in egg-shaped glasses. PMID- 11253573 TI - [Surgery of the mediastinum. I. Mediastinal dissection]. PMID- 11253574 TI - Community workers as extension of nursing personnel. PMID- 11253575 TI - Issues in cross-cultural quality-of-life research. AB - PURPOSE: To examine cross-cultural measurement of quality of life (QOL) and issues to consider in adapting quality-of-life instruments. ORGANIZING CONSTRUCT: Health-related quality of life. METHODS: Review of the literature on cross cultural QOL using the Cumulative Index to Nursing & Allied Health Literature (CINAHL), 1982 to February 2000, and Medline, 1966 to May 2000, databases. FINDINGS: Use of research instruments beyond the samples with which they were initially tested, particularly if the new samples are cross-cultural, presents considerable challenges. The findings indicate consideration of these problems: phenomenon of interest, cross-cultural versus cross-national, salience, conceptual equivalence, cultural hegemony versus cultural validity, cultural equivalence versus verbal equivalence, fidelity versus appropriateness, privacy versus disclosure, appropriateness of format, and resource utilization for translation. Although the literature indicates increased discussion of theoretical, conceptual, and operational approaches to measuring cross-cultural QOL, problems continue in adapting instruments from one culture to another. CONCLUSIONS: Many issues about cross-cultural QOL were identified. By addressing these issues, researchers can develop appropriately translated and validated quality-of-life instruments to advance knowledge about cross-cultural QOL. PMID- 11253576 TI - The shifting perspectives model of chronic illness. AB - PURPOSE: To present the Shifting Perspectives Model of Chronic Illness, which was derived from a metasynthesis of 292 qualitative research studies. DESIGN: The model was derived from a metasynthesis of qualitative research about the reported experiences of adults with a chronic illness. The 292 primary research studies included a variety of interpretive research methods and were conducted by researchers from numerous countries and disciplines. METHODS: Metastudy, a metasynthesis method developed by the author in collaboration with six other researchers consisted of three analytic components (meta-data-analysis, metamethod, and metatheory), followed by a synthesis component in which new knowledge about the phenomenon was generated from the findings. FINDINGS: Many of the assumptions that underlie previous models, such as a single, linear trajectory of living with a chronic disease, were challenged. The Shifting Perspectives Model indicated that living with chronic illness was an ongoing and continually shifting process in which an illness-in-the-foreground or wellness-in the-foreground perspective has specific functions in the person's world. CONCLUSIONS: The Shifting Perspectives Model helps users provide an explanation of chronically ill persons' variations in their attention to symptoms over time, sometimes in ways that seem ill-advised or even harmful to their health. The model also indicates direction to health professionals about supporting people with chronic illness. PMID- 11253577 TI - Accuracy of children's self-reported adherence to treatment. AB - PURPOSE: To examine the relationship between self-reported and electronically monitored adherence to a recommended asthma treatment and to assess the accuracy of the diary data reported by school-age children. DESIGN: A randomized, controlled clinical trial of the effectiveness of an asthma self-management program. The relationship between self-reported and electronically recorded daily peak expiratory flow rate (PEFR) adherence was assessed in a sample of 42 children, ages 7 through 11 years with moderate to severe asthma in one community in West Virginia, USA. Cognitive social learning theory served as the framework for the intervention. METHODS: At-home adherence to PEFR monitoring during the 5 week study was evaluated using the self-report Asthma Diary and an electronic PEFR meter. Recommended twice daily (morning and evening) PEFR monitoring was measured. RESULTS: Self-reported and electronically recorded PEFR adherence were modestly correlated. Self-reported adherence was significantly higher than electronically monitored adherence during Week 5. Accuracy of the self-reported PEFR declined over time, and over half of the children fabricated at least one PEFR value during the final week of the study. CONCLUSIONS: Clinicians often evaluate the efficacy of prescribed treatment for children with chronic conditions based on the children's self-reported diary data. The findings indicate that these children's self-reported adherence behaviors contained errors. Parent education regarding supervision of children's adherence, including validating the accuracy of diary data, is critical for successful self-management in children with chronic conditions. PMID- 11253578 TI - Correlates of resilience in homeless adolescents. AB - PURPOSES: To (a) describe reasons adolescents give for their homelessness, (b) explore relationships among resilience and selected risk and protective factors, (c) identify differences in risk and protective factors by gender and sexual orientation, and (d) determine best predictors of resilience. DESIGN: A descriptive and exploratory correlational design was used to collect and analyze data from a convenience sample of 59 homeless adolescents who sought health and social services from a community street-outreach project in central Texas in 1998. METHODS: A paper and pencil survey consisting of valid measures (Resilience Scale, UCLA-Revised Loneliness Scale, Beck Hopelessness Scale, Social Connectedness Scale, and Death-Related Attitude Schedule) was administered in a street-outreach setting. FINDINGS: Nearly half the sample (47%) reported a history of sexual abuse and 36% self-identified as gay, lesbian, or bisexual in orientation. Over half (51%) were thrown out of their homes by their parents, 37% left home because their parents disapproved of their alcohol or drug use, and nearly one-third left home because parents sexually abused them. Lack of resilience was significantly related to hopelessness, loneliness, life threatening behaviors, and connectedness, but not to gender or sexual orientation. Hopelessness and connectedness explained 50% of the variance in resilience. CONCLUSIONS: Participants who perceived themselves as resilient, although disconnected from other people, were less lonely, less hopeless, and engaged in fewer life-threatening behaviors than were those who perceived themselves as not being resilient. They survived by adapting to street life and by becoming overly self-reliant. Findings may be useful in planning interventions to promote health and well-being in this vulnerable population. PMID- 11253579 TI - Unresolved feelings of guilt and shame in the maternal role with substance dependent African American women. AB - PURPOSE: To identify care needs of African American women residing in an inner city transitional home for substance abuse, with the goal of facilitating their ability to move successfully through treatment and recovery. DESIGN: Ethnography. The convenience sample consisted of 12 key participants and 18 general participants. Data were collected 1996-1998. METHOD: Participant observation and focused interviews were used to collect data. Participants were asked open-ended questions designed to determine their care needs as they experienced treatment and recovery for substance abuse. FINDINGS: Unresolved feelings of guilt and shame associated with perceptions of failure in the maternal role during their active addiction were discovered to be critical issue and possible barrier to successful treatment for African American women in a residential program for treatment of substance abuse. CONCLUSIONS: Unresolved feelings of guilt and shame associated with consequences of the use and abuse of substances, particularly with perceptions of failure in the maternal role, could serve as barriers to successful treatment and recovery for substance-dependent African American women. Nursing actions designed to facilitate healing with these women could offer the potential for improved maternal and child health and well being. PMID- 11253580 TI - Application of Leventhal's self-regulation model to Chinese immigrants with type 2 diabetes. AB - PURPOSE AND BACKGROUND: To demonstrate the application of Leventhal's Self Regulation Model with a group of Chinese immigrants with type 2 diabetes. DESIGN AND METHODS: Using qualitative methods and a convenience sample of 30 Chinese immigrants, interviews were analyzed by categorizing data according to the components of the Leventhal model. Participants were recruited from a U.S. West Coast Chinatown health center and were interviewed to identify beliefs about health and illness that are shaped by cultural factors. FINDINGS: Application of the self-regulation model indicated that participants were unclear about the etiology and chronicity of diabetes and interpreted the illness as stigmatizing. Coping strategies included wishful thinking, belief in powerful others, keeping diabetes a secret, and avoiding social situations. Participants lacked the ability to appraise the effects of their coping strategies. CONCLUSIONS: Health care providers can help people with type 2 diabetes develop critical-thinking strategies instead of relying on sets of rules to gain control of blood glucose levels. The self-regulation model was useful in profiling a vulnerable group whose diabetes management, social environment, and self-image could be improved through thoughtful patient education strategies. PMID- 11253582 TI - Critique of review of therapeutic touch. PMID- 11253581 TI - Nurse practitioners and outcomes. PMID- 11253583 TI - Critique of review of therapeutic touch. PMID- 11253584 TI - Nurses' experiences of restructuring in three Ontario hospitals. AB - PURPOSE: To describe the effects of restructuring, particularly redeployment, on nurses' personal and work lives, and to compare the utility of "survivor syndrome" and empowerment as alternative concepts for understanding these effects and planning change. METHODS: Twenty-six focus groups or interviews were held with 59 nurses working in three hospitals in Ontario, Canada. FINDINGS: Participants described how restructuring strategies had affected them as individuals, as members of nursing teams, and as employees. In each of these aspects of their work lives, relationships became less integrated, their work activities became less controllable, and the changes compromised their ability to deliver effective care. CONCLUSIONS: Restructuring intensifies structural weaknesses in professions, such as nursing, whose members are primarily employed by bureaucracies. Nurses may not find survivor syndrome a useful model to explain their low morale following restructuring because it identifies nurses as "patients" in need of therapy. An empowerment model that takes into account nurses' concerns about uncertainty and integration may be more fruitful for devising strategies to enhance their ability to practice effectively in hospital settings. PMID- 11253585 TI - A gender perspective on conflict management strategies of nurses. AB - PURPOSE: To apply a gender perspective to synthesis of research findings on conflict management. ORGANIZING CONSTRUCT: The Thomas-Kilmann Mode Instrument (TKI), for measuring five conflict-handling strategies: avoiding, compromising, collaborating, accommodating, and competing. METHOD: Nursing research studies with the TKI and other studies are synthesized from perspectives in three gender theories. CONCLUSIONS: Findings were that two conflict management strategies, avoiding and compromising, were used predominantly by all categories of nurses. Possible reasons for over- and underuse of the remaining three strategies (collaborating, accommodating, competing) are described. Implications of these findings for nurses and nursing organizations are discussed. PMID- 11253586 TI - Advanced education and health policy. PMID- 11253587 TI - Need for DNR policies. PMID- 11253588 TI - History and trends in clinical information systems in the United States. AB - PURPOSE: To provide a synopsis of issues about clinical information systems for nurses not schooled in nursing informatics. ORGANIZING CONSTRUCT: The past, present, and future of clinical computing, including major factors resulting in the early hospital information systems (HIS) and decision support systems (DSS) in the United States, current advances and issues in managing clinical information, and future trends and issues. METHODS: Literature review and analysis. FINDINGS AND CONCLUSIONS: The first HIS and DSS were used in the late 1960s and were focused on applications for acute care. The change from fee-for service to managed care required a change in the design of clinical information systems toward more patient-centered systems that span the care continuum, such as the computer-based patient record (CPR). Current difficulties with CPR systems include lack of systems integration, data standardization, and implementation. Increased advances in information and technology integration and increased use of the Internet for health information will shape the future of clinical information systems. PMID- 11253589 TI - An exploratory study of clinical decision-making in five countries. AB - PURPOSE: To identify the cognitive processes nurses use in their decision-making in long- and short-term care settings in five countries, and the demographic variables associated with their decision-making. METHOD AND SAMPLES: The instrument used was a 56-item questionnaire that has been shown to be reliable in earlier studies. The sample consisted of five convenience samples of registered nurses working in either geriatric wards (n = 236) or acute medical-surgical wards (n = 223) in hospitals or nursing homes in Canada, Finland, Sweden, Switzerland, and the United States. FINDINGS: Five models of decision-making were identified on the basis of factor analysis. They represent both analytical and intuitive cognitive processes. Analytical cognitive processes were emphasized in information collection, problem definition, and planning of care, and intuitive cognitive processes were emphasized in planning, implementing, and evaluating care. Professional education, practical experience, field of practice, and type of knowledge were significantly associated with decision-making models as well as with country of residence of the participants. The highest proportion of analytically oriented decision-makers was found among nurses in long-term care, the decision-making of nurses in short-term care was more intuitively oriented. CONCLUSIONS: The results indicate that decision-making of participants varied from country to country and in different nursing situations. Future research should be focused on reasons for these differences, the relationship between the task and the nurses' type of knowledge, and how nurses use their knowledge to make decisions in different nursing situations. PMID- 11253590 TI - Globalization, nursing, and health for all. PMID- 11253591 TI - Ethics in qualitative research. AB - PURPOSE: To critically examine ethical issues in qualitative research. ORGANIZING CONSTRUCT: The ethical principles of autonomy, beneficence, and justice are guides for researchers to address initial and ongoing tensions between the needs and goals of the research and the rights of participants. METHODS: Research literature, ethics literature, and researcher experiences. CONCLUSIONS: Ethical principles can be used to guide the research in addressing the initial and ongoing issues arising from qualitative research in order to meet the goals of the research as well as to maintain the rights of the research participants. PMID- 11253592 TI - Surgical oncologic principles. AB - In this article, there is a return to the beginning of the last century, retracing the evolution of the lung cancer epidemic. As lung cancer increased in frequency, the steps developed to investigate and treat the disease are recalled. At the beginning of the new millennium, the position of surgery in the management of lung cancer is summarized. The role of the surgeon in the investigation and treatment of lung cancer, whether with curative or palliative intent, is evaluated. The principles of surgical management are enunciated, how these principles are presently understood is discussed, and how the disease, its prevention, and treatment may develop in the new millennium is addressed. PMID- 11253593 TI - Stage IIIB non-small-cell lung cancer. AB - Highly selected patients with locally advanced non-small-cell lung cancer achieving cure do not contribute meaningfully to the overall prognosis of stage III. This finding is true particularly if clinicians rely on currently available therapeutic modalities (chemotherapy, radiation therapy, surgery) or refinements thereof. Understanding of the molecular biology continues to improve; it is more likely that in the new millenium, the real breakthroughs in staging and therapy for this high-risk, poor-prognosis group will come from the integration of molecular modalities in the clinical application. PMID- 11253594 TI - The management of non-small-cell lung cancer with oligometastases. AB - The standard care for patients with non-small-cell lung cancer is chemotherapy of supportive care, with surgery being reserved for palliation of symptoms; however, there is a small group of patients with a finite number of extrathoracic metastases (oligometastases) who may experience improved survival by resection of their metastases and the primary site, with or without systemic treatment. This article summarizes the theoretic basis for resection of metastatic lung cancer, reviews the available data addressing management of disease metastatic to the lung, brain, and adrenal glands, and outlines a model for a future clinical trial to investigate the area further. PMID- 11253595 TI - Treatment of metastatic non-small-cell lung cancer. AB - The management of advanced NSCLC remains a daunting challenge; however, new tools are available for treating this malignancy, and continued progress is likely. The future is bright, with a myriad of opportunities to exploit our ever-expanding knowledge of tumor biology. What is perhaps most needed, however, is development of new methods to prevent children and young adults from ever taking up the use of tobacco. In addition, clinicians need new techniques to assist those who are already addicted to escape from tobacco's death grip. Sadly, most users of tobacco still fail to recognize the dangers of their habit. This needs to change. PMID- 11253597 TI - Treatment of stage I lung cancer. AB - This article reviews changes in the roles of surgeons, medical oncologists, and radiation oncologists that have occurred over the past 20 years. A gradual evolution will certainly continue over the next several decades in the multidisciplinary quest to improve the dismal survival results. PMID- 11253596 TI - Small-cell lung cancer. AB - The management of small-cell lung cancer (SCLC) evolved rapidly through the 1980s, but has stalled since then. The relative roles of surgery and radiotherapy in this former systemic disease have been worked out well. The chemotherapy of SCLC has progressed less rapidly. This article discusses the latest developments in each modality of treatment of SCLC and summarizes current treatment strategies. PMID- 11253598 TI - Novel molecular and immunotherapeutic strategies for lung cancer. AB - Lung cancer therapy in the future will be guided by specific characteristics of the individual tumor specimens. The molecular cancer phenotyping will allow for targeted approaches based on the amplification of oncogenes, lack of tumor suppressor genes, dysregulation of growth factors, and angiogenesis or matrix metalloproteinases. Specific immunotherapeutic approaches based on. PMID- 11253599 TI - Postoperative surveillance following lung cancer resection. AB - The purpose of this article is to review routine clinical surveillance testing regimens used in the past and those currently being practiced. In addition, the authors predict follow-up strategies that will be routinely practiced within the next decade. PMID- 11253600 TI - Stage II (N1) lung cancer. AB - As therapies evolve and mature, the authors predict that the next 20 years will see significant advances in the understanding of non-small-cell lung cancer (NSCLC), in molecular stratification of NSCLC, and significant improvement in survival and cure rates. This survival will be achieved through early detection and combined treatments using effective surgical interventions, improved radiotherapeutics, and especially significantly enhanced, rationally designed systemic therapies. PMID- 11253601 TI - Stage II (T3) lung cancer. AB - The tumors classified under T3 are diverse and usually present a difficult problem; however, with the improvements in surgery, radiation, and systemic therapies described above, a significant improvement in survival and quality of life can be anticipated for the future. PMID- 11253602 TI - Evaluation and management of patients with stage IIIA (N2) non-small-cell lung cancer. AB - Management of the stage IIIA (N2) patient remains one of the most controversial areas in thoracic oncology. The potential curability of at least some of these patients has encouraged the development of more complex, aggressive, and toxic multimodality treatment regimens. PMID- 11253603 TI - Induction therapy of adult acute lymphocytic leukemia without the use of vincristine or prednisone. AB - In the last 30 years, a multitude of treatment regimens for adult acute lymphocytic leukemia (ALL) has been developed. Essentially, all of these regimens use an induction therapy vincristine, prednisone, and an anthracycline intensified with L-asparaginase or cyclophosphamide. Though such regimens induce most patients to enter a remission, relapse is frequent, and most adult patients ultimately die of their disease. The author postulated that further refinements in this approach to induction therapy were unlikely to markedly improve treatment results in this disease. Therefore, the author is studying a new intensive strategy using cytarabine with a single very high dose of mitoxantrone (without vincristine or prednisone) as induction therapy for adult patients with ALL. PMID- 11253604 TI - Autologous stem cell transplantation for acute lymphocytic leukemia in adults. AB - Autologous bone marrow transplantation remains an investigational treatment for adult ALL. Despite many anecdotal studies showing efficacy, the rarity of ALL has prevented the large randomized trials necessary to confirm effectiveness. Candidates for autoBMT include adult patients in first CR with adverse risk factors and all patients who have experienced disease relapse. It remains debatable which preparative regimen is optimal, whether purging is necessary, or if chemotherapy or immunotherapy administered after transplantation can decrease disease relapse. Overall, every effort should be made to enter ALL patients on well-designed randomized multi-institutional trials. These trials should compare autologous transplantation to newer more intensive chemotherapy regimens and should take into account the heterogeneity of ALL. A quality of life analysis should be performed as one high-dose treatment may be less toxic and better tolerated than multiple cycles of consolidation chemotherapy. Strategies aimed at enhancing an autologous graft-versus-leukemia effect after transplantation may enhance long-term survival. Many more studies are needed to further define the optimal role of autoBMT in adult ALL. PMID- 11253605 TI - Central nervous system involvement in adult acute lymphocytic leukemia. AB - With effective CNS prophylaxis, most adults with ALL may remain free of CNS leukemia. Several combinations of IT chemotherapy, high-dose systemic chemotherapy, and cranial irradiation have been used with varying results. Excellent prophylaxis can be achieved without cranial irradiation, and in view of the potential acute and long-term toxicity of radiation, these methods may be preferable. A prophylactic approach tailored to the risk of CNS leukemia was shown to be valuable in childhood ALL and in at least one adult study. Further studies should focus on defining risk groups for CNS leukemia and designing effective prophylaxis for each group. More research is needed to define the intensity and duration of treatment and the role of cranial irradiation in the treatment of isolated CNS relapses. PMID- 11253606 TI - Salvage therapy for refractory or relapsed acute lymphocytic leukemia. AB - The overall prognosis for patients with relapsed or refractory adult ALL remains poor. Further insight into the biology of ALL is required, and novel therapeutic agents are needed to counter mechanisms of resistance. A palliative approach to the management of multiply relapsed or refractory ALL should be supplanted by enrollment into clinic trials to promote drug discovery. Monitoring of minimal residual disease may allow an earlier intervention before overt clinical relapse and improve outcome; prospective studies are needed. Attainment of a second or later CR should be followed by allogeneic BMT when feasible owing to the paucity of long-term survivors with salvage chemotherapy alone. PMID- 11253607 TI - Adult acute lymphocytic leukemia. Future research directions. AB - This article reviews future research of potential benefit to adults with acute lymphocytic leukemia (ALL). This new research includes dose intensive anthracyclines, novel agents, STI571 for Philadelphia chromosome-positive ALL, and risk-oriented strategies. PMID- 11253608 TI - Burkitt's acute lymphocytic leukemia (L3ALL) in adults. AB - Burkitt's acute lymphocytic leukemia is a rare type of adult ALL, probably difficult to distinguish from disseminated Burkitt's lymphoma involving the bone marrow. This tumor is highly proliferative and tends to involve the CNS at diagnosis or early during the disease course. It shows rapid chemosensitivity, initially leading to the risk of severe acute tumor lysis syndrome. Principles of its treatment, by comparison with the other types of ALL, include: 1. A low-dose chemotherapy prephase to prevent acute tumor lysis syndrome. 2. Multiagent chemotherapy using high-dose cyclophosphamide, an anthracycline, high-dose MTX, high-dose ara-C, and probably VP16. A short and intensive treatment (6 to 8 months) without maintenance is indicated. 3. Early intensive CNS treatment, with multiple triple intrathecal injections, high-dose MTX, and high-dose ara-C, and possibly cranial irradiation. Autologous or allogeneic stem cell transplantation do not seem to be useful in first CR. Using such approaches, recent results suggest that about two thirds of L3ALL in adults can be cured, more than in any other type of adult ALL. PMID- 11253609 TI - Lymphoblastic lymphoma. AB - Recent advances in the unique clinicopathologic entity of lymphoblastic lymphoma (and its variants) are discussed in this article, which details the natural history, molecular biology, prognosis, and outcome with various chemotherapy regimens. Improved outcome with the newer intensive chemotherapy regimens and the role of modalities such as autologous intensification, allogeneic bone marrow transplant, and radiotherapy are discussed. PMID- 11253610 TI - High-dose anthracycline induction in adult acute lymphocytic leukemia. AB - Many anthracyclines seem to be essential components of chemotherapy in adult ALL patients. The results of different studies suggest that 1. anthracyclines increase the probability of CR 2. complete response is obtained earlier with the use of high-dose anthracyclines 3. the optimal timing for anthracycline administration is probably the very early phase of the disease, because an earlier CR is a favorable prognostic indicator in ALL 4. high-dose DNM may be beneficial: given at high dosage in a dose-intensive way it can reduce the occurrence of relapse. There is evidence from randomized and nonrandomized studies that anthracyclines increase the CR rates when added to other classic components of ALL therapy. The dose and dose intensity of DNM during induction may influence duration of CR and long-term prognosis. Although anthracyclines in induction are only part of a complex program of therapy for adult ALL, the dose and dose intensity of anthracyclines may become important components of the armamentarium against this devastating disease. PMID- 11253611 TI - Allogeneic stem cell transplantation for acute lymphocytic leukemia in adults. AB - Acute lymphocytic leukemia remains a difficult disease to treat in adults. Allogeneic bone marrow transplantation can cure some patients with ALL, but GVHD transplant-related mortality and disease relapse remain problematic. Immunotherapeutic approaches aimed at eliminating minimal residual disease and enhancing the GVL effect may decrease relapse rates and improve overall survival. Because of the rarity of this disease in adults, every new patient should be entered in well-designed, peer-reviewed, investigational trial. PMID- 11253612 TI - A whole-body physiologically based pharmacokinetic model incorporating dispersion concepts: short and long time characteristics. AB - In whole-body physiologically based pharmacokinetic (PBPK) models, each tissue or organ is frequently portrayed as a single well-mixed compartment with distribution, perfusion rate limited. However, single-pass profiles from isolated organ studies are more adequately described by models which display an intermediate degree of mixing. One such model is the dispersion model, which successfully describes the outflow profiles from the liver and the perfused hindlimb of many compounds, under a variety of conditions. A salient parameter of this model is the dispersion number, a dimensionless term, which characterizes the relative axial spreading of compound on transit through the organ. We have developed a whole-body PBPK model wherein the distribution of drug on transit through each organ is described by the dispersion model with closed boundary conditions incorporated. The model equations were numerically solved using finite differencing methods, in particular, the method of lines. An integrating routine suitable for solving stiff sets of equations was used. Physiological parameters, blood flows, and tissue volumes, were taken from the literature, as were the tissue dispersion numbers, which characterize the mixing properties of each tissue; where none could be found, the value was set as that for liver. On solution, tissue, venous and arterial blood concentration-time profiles are generated. The profiles exhibited both short and long time characteristics. Oscillations were observed in the venous and arterial profiles over the first 10 min of simulation for the rat. On scale-up to human, the effects were seen over a 30 min period. Longer time effects of tissue distribution involve buildup of drug in the large tissues of distribution: skeletal muscle, skin, and adipose. The extent of distribution in the large tissues was somewhat dependent on the magnitude of the dispersion number, the lower the dispersion number, the greater the extent of distribution after an intravenous bolus dose. The model has a distinct advantage over the well-stirred organ whole-body PBPK model in its ability to describe both short and long time characteristics. PMID- 11253613 TI - The rate and extent of oral bioavailability versus the rate and extent of oral absorption: clarification and recommendation of terminology. AB - In the literature, the meanings of the terms oral absorption and oral bioavailability of drugs vary greatly. Absorption has been considered to take place at the mucosal membrane of the gastrointestinal (GI) tract. It has also been defined as the process from the site of drug administration to the site of measurement. In the latter definition, the extent of oral absorption depends on the extent of first-pass elimination in the gut wall and liver even though a drug may be completely absorbed from the GI tract. Moreover, these two terms have also been used interchangeably. Inconsistency in the definition of these two terms has led to varying interpretations of these terms among students, researchers and laymen, and such an inconsistency seems undesirable. Apparently because of these inconsistencies, improper correlations between the Caco-2 permeability or intestinal permeability and the oral bioavailability of drugs subject to extensive first-pass effect may have occurred. It is suggested that absorption be defined as movement of drug across the outer mucosal membranes of the GI tract, while bioavailability be defined as availability of drug to the general circulation or site of pharmacological actions. Since transit times (this may range from about 1 min to several hours) across enterocytes, liver, lungs, and the peripheral venous sampling tissue are virtually unknown for all drugs, this factor alone would favor the use of "oral bioavailability rate" rather than "oral absorption rate" in all routine studies. PMID- 11253614 TI - Indirect pharmacodynamic models for responses with multicompartmental distribution or polyexponential disposition. AB - Basic indirect response models where drug alters the production (kin) of the response variable (R) based on the Hill function previously assumed one compartment distribution of the response variable and simple first-order loss (kout) of R. These models were extended using convolution theory to consideration of two-compartment distribution of R and/or polyexponential loss of R. Theoretical equations and methods of data analysis were developed and simulations are provided to demonstrate expected response behavior based on biexponential response dissipation. The inhibition model was applied to our previous data for inhibition of circadian cortisol secretion by prednisolone. The presence of multicompartment response variables and/or polyexponential loss complicates the response patterns and resolution of pharmacologic parameters of indirect response models and requires careful experimental and data analysis approaches in order to properly evaluate such pharmacodynamic responses. The occurrence of these alternative distribution or disposition components does not alter the area under the effect curve (AUCE) which remains identical to the basic models. Model misselection was addressed by testing fittings comparing the basic and new models. Use of the former for these more complex models does not severely perturb the calculated cardinal dynamic parameters. These models may provide improved insights into indirect responses with complexities in distribution or disposition. PMID- 11253615 TI - Peripheral link model as an alternative for pharmacokinetic-pharmacodynamic modeling of drugs having a very short elimination half-life. AB - Attempts to obtain estimates of pharmacokinetic-pharmacodynamic (PK-PD) parameters for mivacurium with traditional central link models were unsuccessful in many patients. We hypothesized that a link model with the peripheral compartment would be more appropriate for mivacurium in view of its extremely rapid plasma clearance and its potential elimination by tissue pseudocholinesterases. For validation purposes, the peripheral link model was applied to other neuromuscular blocking agents (NMBA), i.e., atracurium and doxacurium which have respectively an intermediate and a long elimination half life. Assuming peripheral elimination in PK-PD modeling was investigated but found to have no impact on the estimation of PK-PD parameters. Our results indicate that, for drugs having intermediate and long elimination half-lives, EC50 values are similar with either the central or peripheral link model. For mivacurium, a peripheral link model enables PK-PD modeling in all subjects, with more precision in the PK-PD parameter estimates and a better fitting of the effect data when compared to the central link model. For these reasons, a peripheral link model should be preferred for mivacurium. PMID- 11253616 TI - Population pharmacokinetic modeling of steady state carbamazepine clearance in children, adolescents, and adults. AB - Carbamazepine (CBZ) clearance decreases from childhood to adulthood and the factors determining this change could include age, size, autoinduction, or maturational changes. This study aims to describe the population pharmacokinetics of CBZ in children and young adults and test the hypothesis that CBZ clearance correlates with weight, surface area, and age. CBZ therapeutic drug monitoring data (sparse data) were collected from child and adult epileptics, and rich data were obtained from a bioequivalence study of CBZ in young adults. Population pharmaco-kinetic analysis was performed using NONMEM V. Forward stepwise, multiple regression was performed on the covariates. Bootstrap validation was performed. A total of 946 observations from 91 subjects, ages 0.7-37 years, were collected and analyzed. A one-compartment, first-order absorption and elimination model, with exponential interindividual error and additive residual error models was developed. The population model was: Clearance (Lhr-1) = ((2.24 x Surface area (m2)) + (0.047 x Dose (mg.kg-1)); Volume of distribution (L) = 0.37 x weight (kg); Absorption rate constant = 0.013 (hr-1). CBZ clearance increased with surface area and dose. PMID- 11253617 TI - An identifiability analysis of a parent-metabolite pharmacokinetic model for ivabradine. AB - The paper considers the structural identifiability of a parent-metabolite pharmacokinetic model for ivabradine and one of its metabolites. The model, which is linear, is considered initially for intravenous administration of ivabradine, and then for a combined intravenous and oral administration. In both cases, the model is shown to be unidentifiable. Simplification of the model (for both forms of administration) to that proposed by Duffull et al. (1) results in a globally structurally identifiable model. The analysis could be applied to the modeling of any drug undergoing first-pass metabolism, with plasma concentrations available from drug and metabolite. PMID- 11253618 TI - Patient confidentiality takes on a new meaning. PMID- 11253619 TI - Sequential bilateral total knee arthroplasty. AB - As the senior citizen population has grown, the incidence of osteoarthritis and joint replacement has increased. Bilateral total knee arthroplasty (BTKA) can be performed sequentially during one anesthetic. Studies have shown the complication rates differ only slightly for total knee arthroplasty procedures performed sequentially during one anesthetic or separately requiring two hospitalizations. With the use of staggered tourniquet deflation times, efficient OR time and motion techniques, and careful postoperative management, patients can achieve successful outcomes after BTKA during one anesthetic. PMID- 11253620 TI - An integrative review of pressure relief in surgical patients. AB - Effective patient positioning has been an important issue throughout the history of the nursing profession. Pressure ulcers result from prolonged pressure, which causes skin, tissue, or muscle damage. Surgical patients present a unique challenge in preventing pressure ulcers because they are immobile and unable to perceive the discomfort of prolonged pressure. The purpose of this integrative review is to examine risk factors associated with pressure ulcer development in surgical patients and to examine pressure-relieving support surfaces to determine if they significantly reduce intraoperative tissue pressure and result in a lower incidence of postoperative pressure ulcers. Most of the research focuses on long term care units, with little attention given to the acute care setting. Although the pathophysiology and etiology of pressure ulcers are well documented by years of research, the OR as an etiologic factor is largely undefined. PMID- 11253622 TI - Nurses and their right to whistle blow. PMID- 11253621 TI - Nurses as working women. AB - Women today make up nearly half of the nation's workforce. A number of these women have children at home, and women also often are responsible for providing care to older adult family members or friends. The different roles assigned to women in today's society are burdensome, particularly for nurses who deal with the stress of managed care, downsizing, the nursing shortage, caring for increasingly ill patients, long and irregular hours, and daily crises. There are ways that nurses can modify their work situation, however, to help them cope with the rigors of the workplace and the home. PMID- 11253623 TI - Evaluating the findings of qualitative research. AB - This article provides an overview of strategies used to support and evaluate qualitative research. Research consumers should critique qualitative research to see how the concepts of credibility, transferability, and confirmability are met in a study. Further, as qualitative research evolves and transforms the understanding of phenomena, the strategies for evaluating credibility may evolve and change, as well. Qualitative research just as personal knowing, is a dynamic process. PMID- 11253624 TI - [Treatment of coronary risk factors by general practitioners in Austria]. AB - Aim of the study was to investigate attitudes and practice of coronary prevention in offices of general practitioners and internists. 67 Austrian physicians took part in a mail survey focussing on hyperlipidemia. 96.6% of participating physicians recommend dietary intervention at least at total cholesterol levels of 250 mg/dl, 98.0% prescribe lipid-lowering drugs at least at total cholesterol levels of 300 mg/dl. At corresponding levels of total/HDL cholesterol ratios and especially at corresponding levels of LDL-cholesterol the proportion was lower. On average 37.9% of physicians spend up to 5 minutes for patients with hyperlipidemia, 10.3% spend more than 15 minutes. The time frame is similar in overweight patients, and bigger in patients with hypertension and diabetes. Dietary therapy is estimated similarly successful in patients with hyperlipidemia and overweight, but estimated more successful in patients with hypertension and diabetes. Drug therapy of hyperlipidemia is estimated more successful than in overweight and diabetes, and worse compared with hypertension. Main measures for improving prevention are more time and specific postgraduate education. The majority of physicians feel that within the last five years quality has improved both in the outpatient and the inpatient care. PMID- 11253625 TI - Serum creatine kinase elevation in a medical department. AB - Serum creatine kinase (CK) levels are diagnostic markers for acute myocardial infarction. Many other causes however, including neuromuscular disorders, may induce serum CK elevation as well. Aim of the study was to investigate the prevalence of potential causes for serum CK elevation in a medical department. In particular we were interested in the recognition of patients in whom serum CK elevation was due to a neuromuscular disorder. Included in this prospective study were 100 consecutive patients in whom the CK level, determined at admission, was increased (> 70 IU/l in female, > 80 IU/l in male patients). After admission we looked for the presence of causes known to induce CK elevation. Patients with no potential cause for CK elevation were invited for follow-up investigations three months later. If no potential cause could be found and if CK was elevated again on this occasion, the patient was referred for a comprehensive neurological investigation. The prevalence of patients with CK elevation was 11.2%. The 100 patients (44 female, 56 male) were aged from 23 to 94 (mean 67) years. In 95% CK elevation was only up to 500 IU/l. The most frequent cause for serum CK elevation was acute myocardial infarction in 32%. Further frequent causes were drug intake (32%), fall (24%), haematoma (17%), intramuscular injection (16%) and malignancy (11%). In 61% of the cases at least two potential causes for serum CK elevation could be detected. Neuromuscular disorders were found in only 2%. This study shows that serum CK elevation occurs in 11% of patients admitted to a medical department and can be explained by acute myocardial infarction in only 32%. In almost two thirds of the patients, more than one potential cause for serum CK elevation can be found thus making CK elevation a rather unspecific finding. Neuromuscular disorders are rarely found as a cause of serum CK elevation in a medical department. PMID- 11253626 TI - Radiofrequency catheter ablation of the bundle branch reentrant ventricular tachycardia. AB - Bundle branch reentrant ventricular tachycardia (BBRVT) has a suitable anatomic substrate for radiofrequency catheter ablation. However, the experience with this treatment is still small. In the current study, we examined the safety and the long-term efficacy of radiofrequency ablation in the cure of patients with BBRVT. Four patients with BBRVT, identified during electrophysiological study, underwent temperature-controlled radiofrequency ablation of the right bundle branch (RBB). All of them had syncope and structural heart disease with reduced left ventricular ejection fraction. The baseline examination revealed an intraventricular block, prolonged HV interval and inducible sustained VT because of bundle branch reentry in all patients. RBB was successfully abolished in all patients after the delivery of 3 +/- 1 radiofrequency pulses. After ablation, a permanent pacemaker was implanted in one patient with significantly prolonged HV interval. All patients were free of BBRVT during a mean follow-up of 20 months. One patient received implantable cardioverter-defibrillator for myocardial VT five months after ablation. Two patients developed congestive heart failure. Radiofrequency catheter ablation of the RBB is a safe and highly effective therapeutic procedure for definitive cure of BBRVT. Long-term prognosis of these patients depends mainly on the underlying heart disease and the treatment of other VT. PMID- 11253627 TI - Nocturnal sleep-disturbing nicotine craving: a newly described symptom of extreme nicotine dependence. AB - In our research on smoking and nicotine dependence we have noticed a sleep disturbance, which is a further symptom of extreme nicotine dependence. We call this symptom "nocturnal sleep-disturbing nicotine craving" (NSDNC). NSDNC is characterised by craving for cigarettes during the individual sleep times. The smoker awakes (one or several times per week) during his regular sleep time, and has to smoke a cigarette before he/she continues sleeping. This symptom can be explained by the decreasing nicotine levels during the sleep time, which results in nicotine craving. However, NSDNC should be carefully separated from other sleep disturbances, or sleep disturbing events (nycturia, medication side effects), when nicotine craving is not the main reason for awakening. PMID- 11253628 TI - Expanding our interventional skills: placement of totally implantable injection ports by internists/intensivists. AB - Totally implantable injection ports are usually placed by surgeons or radiologists using fluoroscopic guidance. In a prospective study we evaluated the efficacy of percutaneous insertion of these devices without the use of fluoroscopic control by internists/intensivists experienced in the placement of permanent cuffed catheters. The supraclavicular approach to the subclavian vein was chosen for first line puncture site because of its low rate of malpositions and complications. 101 ports were inserted in 101 consecutive patients, 96 from the supraclavicular approach. Difficulties in introducing the catheter through the peel-away sheath, misplacement into adjacent vessels, secondary migration, or fragmentation of a line were not observed. Function was excellent in all ports. Three pneumothoraces (3%) and three arterial punctures (3%), none of which required intervention, were recorded. Two ports (2%) had to be revised, one due to local hematoma and another because of inadequate catheter length. Catheter survival was 94% in a 30-month observation period. Placement of totally implantable port systems by internists/intensivists experienced in placing central venous lines is safe and efficient, thus the implantation can easily be performed with minimal technical expenditure in the setting of an intensive care unit. The supraclavicular approach is suitable for insertion of permanent infusion port systems without fluoroscopic control. PMID- 11253629 TI - [Identification of high risk patients with severe, but asymptomatic aortic stenosis]. AB - Whether asymptomatic patients with severe aortic stenosis benefit from surgery remains unclear. We report our data recently published in the New England Journal of Medicine on the natural history of this disease and predictors of outcome. PMID- 11253631 TI - A silent change. PMID- 11253630 TI - [Coronary artery bypass graft dysfunction--clinical presentation, laboratory and electrocardiographic parameters]. AB - The recurrence of symptoms after coronary artery bypass surgery is often caused by bypass-dysfunction. In this study we tried to determine factors related to the long-term patency of arterial and venous bypass grafts. We evaluated all patients with bypass grafts undergoing coronary angiography in the year 1998 at our hospital (163 patients, mean age 67 years, mean interval since the operation 79 months, a total of 341 venous bypasses (VBP), 386 peripheral venous anastomoses and 85 arterial (LIMA = left internal mammarial artery) bypasses. The data were collected by a retrospective analysis of the hospital records. Statistics were performed using the Wilcoxon-Mann-Whitney-U test. After an interval of 53 months LIMA-bypasses were patent without stenosis in 92%. Symptoms were caused in only 2% by a dysfunction of the LIMA-graft. The patency of venous bypass grafts decreased with time (5 years after the operation 74% were patent without stenosis, 5-10 years 56%, more than 10 years 35%, p < 0.01). We found clear relations between the function of the venous grafts and the clinical presentation (patent grafts without stenosis in 43% with acute coronary syndromes, in 57% with stable angina [p = 0.08] and in 86% with atypical angina [p < 0.0001 for the difference between each of the first two and the last syndrome]), the resting-ECG (65% patent VBP without stenosis with normal ST-segments and 49% with abnormal ST segments, p < 0.01), the body-mass-index (70% patent VBP without stenosis with a BMI < 25 and 56% with a BMI > 30, p = 0.05) and the erythrocyte sedimentation rate after 2 hours (79% patent VBP with an ESR < or = 20 mm vs. 64% with an ESR > 49 mm, p = 0.02). The function of VBP after coronary artery bypass graft (CABG) procedure depends primarily upon the interval since the operation. In addition, we found correlations with clinical presentation, resting-ECG, body-mass-index and erythrocyte-sedimentation rate as a possible marker of inflammation in bypass atherosclerosis. Therefore, inflammatory processes seem to play an important role in the development of venous graft dysfunction. PMID- 11253632 TI - The effect of glycosylation on emulsifying and structural properties of bovine beta-casein. AB - A number of attempts have been made to improve the functional properties of milk proteins by chemical modifications. One of such modifications is glycosylation which was carried out to determine the effect of covalent binding of glucose molecules to beta-casein (beta CN) on its emulsifying properties. It was found that up to six molecules of glucose were bound to one molecule of beta CN. Glycosylated beta CN produced smaller emulsion droplets than the intact beta CN. Increases in emulsion-forming and -stabilizing properties were observed. A prerequisite of proteins to form emulsions is their adsorption onto oil/water interface. Therefore the secondary structure of intact and glycosylated beta CN, both in solution and adsorbed onto a hydrophobic teflon/water interface also were studied by far-ultra violet circular dichroism (CD). It appeared, that after glycosylation the degree of helicity of intact beta CN decreased and the incorporation of glucose moieties most likely resulted in a type beta-turn formation after adsorption onto the interface. PMID- 11253633 TI - Cholesterol content in European bovine milk fats. AB - Data about the cholesterol content in edible fats like bovine milk fat are important for balancing the cholesterol intake with food. A comparison of 3 different cholesterol determination methods showed that with the direct analysis by a 25 m long TAP steel capillary column the same results could be obtained as with a time-consuming saponification standard method including thin-layer chromatographic cleaning and subsequent silylation. On the other hand with a rapid direct method using a short packed column 21% unsaponifiables as e.g. minor sterols or hydrocarbons could be found in the "cholesterol peak". The analysis of 1142 German milk fats led to a mean cholesterol content of 265.6 +/- 20.0 mg/100 g fat (range: 204.4 to 382.5). For 165 milk fats from other 12 EU-countries, a similar mean cholesterol content of 258.5 +/- 19.9 mg/100 g fat (range: 215.0 to 331.6) was detected. Compared with sufficiently fed cows, underfed cows demonstrated an approx. 10.1% lower mean cholesterol content (238.7 +/- 9.7 mg/100 g fat). On the other hand, during the first 7 days post partum, the colostrum showed a significantly higher mean cholesterol content of 327.2 +/- 99.0 mg/100 g fat (n = 15; range: 213.1 to 583.9). Further, with special conditions as feeding of rape-seed the cholesterol content can be significantly lowered by 8-13%. An extraordinary lowering up to 50% can be reached by dry fractionation of milk fat (stearin "hard" fraction). PMID- 11253634 TI - Biochemical and technological studies on the production of isolated guar protein. AB - Guar seeds contain 32% crude protein. Therefore, attempts were made to prepare protein isolates from guar seed flour (GSF) by extraction in different media (distilled water, salt solution, alkali solution alone or in combination) followed by a precipitation at acid pH. From the four technologies adopted, mixed salt-alkali solution was found to be the most satisfactory for extraction of protein from GSF. The highest amount of product was obtained in the mixed technology along with the highest amount of protein (87.5%). Protein isolates were also nutritionally evaluated following well-established rat bioassay procedures in a comparative study with casein as standard. The protein isolates are rich in lysine but poor in sulphur-containing amino acids such as methionine and cysteine. Protein isolates obtained by mixed salt-alkali solution showed high water and oil absorption as well as good emulsifying and foaming stability. The results indicate that protein isolates can be used as a supplementary source of protein in different food industries. PMID- 11253635 TI - In vitro protein and starch digestibility of pearl millet (Pennisetum gluacum L.) as affected by processing techniques. AB - The effect of malting and blanching on the in vitro protein and starch digestibility of pearl millet (Pennisetum gluacum L.) were investigated. Pearl millet seeds were subjected to malting [steeping (16 h), germination (48 and 72 h) and kilning (24 h at 50 degrees C)] and blanching (30 s at 98 degrees C) treatments, before grinding to flour. The results indicated that both the treatments improved the in vitro digestibility significantly. Malting appreciably improved the in vitro protein (14-26%) and starch (86-112%) digestibility and improvement by malting was significantly higher than blanching. The effect of malting with 72 h of germination was most remarkable in improving the in vitro protein and starch digestibility. PMID- 11253636 TI - Fusariotoxins in kernels of winter wheat cultivars field samples collected during 1993 in Poland. AB - In South-Eastern region of Poland (near Lublin), where frequency of scab (fusariosis) is much higher than in other parts of the country, during harvest of 1993 kernels of 25 winter wheat cultivars were collected. On the basis of morphological studies Fusarium graminearum was found in 42% of investigated samples while other fungi appeared less frequently: F. nivale and F. poae (35%), F. avenaceum (31%) and F. culmorum (12%). Chemical analysis (by HPLC) revealed that the tested cultivars were contaminated with deoxynivalenol (96% of investigated samples), its acetyl derivatives (48%), nivalenol (76%) and moniliformin (28%). The average levels of the metabolite concentrations were as follows: 104; 16; 97; and 63 micrograms/kg, respectively. Co-occurrence of 2 toxic metabolites was found in the following percentage of the positive samples: deoxynivalenol and nivalenol (72%), deoxynivalenol and moniliformin, as well as nivalenol and moniliformin (24%). Usually (71-83% of contaminated samples) mycotoxins were accumulated in the concentration range > or = 10, < 100 micrograms/kg. PMID- 11253637 TI - Enrichment of Egyptian 'Balady' bread. Part 2. Nutritional values and biological evaluation of enrichment with decorticated cracked broadbeans flour (Vicia faba L.). AB - Decorticated cracked broadbeans flour (DCBF) was used to replace 5%, 10%, 15% and 20% of the wheat flour (WF) in the Egyptian 'Balady' bread. The nutrient composition of DCBF, WF, wheat bread, and DCBF-fortified bread was studied. When DCBF fortification was increased from 0 to 20%, there was an increase of 36% in protein, 18% in fat, 123% in calcium, 52% in phosphorus and 40% in iron contents. DCBF contained greater amounts [g/16 g N] of lysine and histidine compared to WF. All essential amino acids increased when DCBF was substituted for WF from 0 to 20%, except methionine, which decreased. The biological quality of the breads was investigated. The protein efficiency ratio for 10%. DCBF bread (1.60) was found to be significantly greater (P < 0.05) than that of 5%-DCBF bread (1.48) and wheat bread (1.17). PMID- 11253638 TI - Chemical composition and nutritional characteristics of some hull less and hulled barley cultivars grown in India. AB - Three hull less and nine hulled barley cultivars were grown during 1995-96 at Research Farm, CCS Haryana Agricultural University, Hisar, India and were used for chemical analysis such as nutritional and antinutritional parameters. Dolma, a hull less barley cultivar had higher contents of protein, fat, starch, in vitro digestibility of protein and starch, in vitro availability of Ca, Fe and Zn. Hulled cultivar BH-331 showed higher values of phytic acid (925 mg/100 g), polyphenols (625 mg/100 g) and amylase inhibitor activity (169 AIU) which might have contributed towards poor in vitro digestibility of Ca, Fe and Zn. Phytic acid and polyphenols manifested significantly negative correlation with in vitro availability of Ca, Fe and Zn and protein digestibility. Whereas amylase inhibitor activity showed significant and negative correlation (-0.992) only with starch digestibility. PMID- 11253639 TI - A survey on organochlorine pesticide residues in butter and cracked wheat available in Turkish markets. AB - This study has been conducted with a view to examine the level of organochlorine pesticide residues which are Quintozene (PCNB), Dichlorodiphenyl trichloroethane (p,p'-DDT), Dichlorodiphenyl dichloroethylene (p,p'-DDE) and Dichlorodiphenyl dichloroethane (p,p'-DDD), exiting in butter and cracked wheat sold in Ankara local markets. Thus, the use of some pesticides is restricted due to the factors such as the environmental pollution and for saving the health of consumers. Therefore, the aim of this study is to analyse the results achieved from such a restriction. 100 samples of butter and 50 samples of cracked wheat were obtained from the different markets in order to be analysed. After extracting the organochlorine pesticides from the samples, the amounts have been determined by gas liquid chromatography-ECD method. The results of this study revealed that the butter sold in Ankara local markets did not contain organochlorine pesticide residues while the cracked wheat contained PCNB and lindane residues although the amounts of residues had been found in the Food and Agriculture Organization (FAO) tolerance limits. PMID- 11253640 TI - Derivative spectrophotometric determination of 5-(hydroxymethyl)-2-furaldehyde (HMF) and furfural in Locust bean extract. AB - Furfural and 5-(hydroxymethyl)-2-furaldehyde (HMF) are useful indicators of accurate storage temperature of food samples. A first derivative spectrophotometric method was developed for the determination of the furfural and HMF level in Locust bean pekmez. In addition, the levels of the furfural and HMF were investigated during Locust bean pekmez preparation. The recoveries were generally nearly 100%. This method is also rapid and accurate and results in economical analyses of furaldehydes. PMID- 11253641 TI - The effect of air-drying on glycosidically bound volatiles from seasonally collected origano (Origanum vulgare ssp. hirtum) from Croatia. AB - The glycosidically bound volatiles were isolated from fresh and airdried origano by exhaustive percolation with ethyl acetate. After the purification of glycosidic fraction, the enzymatic hydrolysis with beta-glucosidase was performed. The obtained aglycones were analysed by GC-MS. Eighteen compounds were identified. The seasonal variations of main aglycone contents from the fresh plant material were: thymoquinone (3.10-6.18 mg/kg), benzyl alcohol (1.33-3.62 mg/kg), 3,5,5-trimethyl-4-(3-hydroxy-1-buthenyl)-2-cyclohexen-1-on (0.51-3.35 mg/kg), 2-phenyl ethanol (0.42-2.98 mg/kg), eugenol (0.93-2.55 mg/kg), thymol (0.70-1.40 mg/kg) and carvacrol (0.88-1.40 mg/kg). The season of collecting influenced the content and composition of the glycosidically bound volatiles of origano. Air-drying effected mostly the yield of these compounds. PMID- 11253643 TI - Survival of Listeria monocytogenes during the manufacture and ripening of Turkish white cheese. AB - Listeria monocytogenes was enumerated during the manufacture and ripening of Turkish White cheese with particular reference to a) pasteurized milk, b) cheese milk after inoculation with L. monocytogenes (0 h), c) after curd formation (2 h), d) curd after pressing (6 h), e) curd after pH was reduced (17 h), f) curd after salting (32 h), and g) cheeses during ripening. Cheeses were also examined periodically for total solids, moisture and salt contents, pH values and aerobic plate count. An increase in the number of L. monocytogenes was observed during manufacture. Following salting and throughout the storage period, numbers of L. monocytogenes decreased at a rate depending on the salt concentration, starter activity and storage time. The initial microbial number had a significant (P < 0.01) effect on the survival of L. monocytogenes during the storage period. PMID- 11253642 TI - Investigation on the nutritive value and microbiological quality of wild quail carcasses. AB - Quail meats have many advantages and superiority one the other species of poultry. This study was planned to throw plenty of light on gross chemical composition, lipid fractions, fatty acids composition, amino acids composition, of thigh and breast of male and female wild quail meat as well as the microbiological quality. The mean values of moisture, protein, fat, ash and energy contents ranged from 60.1 to 69.2%, 55.0 to 68.8%, 28.8 to 42.1%, 2.40 to 3.63% and 696 to 1000 kJ, respectively. Seven fractions of lipids (phospholipids, monoglycerides, cholesterol, diglycerides, free fatty acids, triglycerides and hydrocarbons) were estimated. The individual fatty acids were determined. The mean total unsaturated fatty acids represented 73.9, 66.8, 60.2 and 67.5% of the total fatty acids in thigh male, breast male, thigh female and breast female quail, while that of saturated fatty acids were 25.1, 30.1, 32.0 and 30.4%, respectively. The essential fatty acids in thigh and breast males were 34.8 and 29.0% against 25.7 and 28.1% in females. Amino acids composition were varied from 82.6 to 95.2 g/100 g protein in thigh, breast of male and female wild quails. The essential amino acids were illustrated. The mean values of psychotrophic, Pseudomonas, Enterobacteriaceae, coliforms, Streptococci and Staph. aureus were 4 x 10(4), 1 x 10(2), 4 x 10(3), 3 x 10(3), 6 x 10(2) and 1 x 10(3) cfu/g, respectively. E. coli, Enterobacter agglumerans, E. cloacae, Morganella morgani, Proteus mirabilis, and P. vulgaris could be isolated in varying percentages. Neither Salmonellae nor Clostridium perfringens could be isolated from the examined quails. The public health aspects for the estimated and isolated criteria were outlined. PMID- 11253644 TI - Investigation of kinetics of zinc biosorption by Saccharomyces cerevisiae cells. AB - The highest amount of zinc (approximately 90%) is bound after 3 h of contact at low initial (total) concentrations of zinc in yeast broth, 2.0-16.0 g/l at 10-30 degrees C. The equilibrium between bound and free zinc ions is established after 6 h of contact time, independently on total zinc concentration in yeast milk. No bigger changes of content of zinc bound to yeast cells was determined at 10 degrees C and 30 degrees C. 40% of bound zinc in the equilibrium state is bound during the first 15 min of contact of zinc ions and yeast cells at all initial (total) zinc concentrations in yeast milk both at 10 degrees C and 30 degrees C. The KEKAM (Kolmogorov-Erofeev-Kozeeva-Avrami-Mampel) equation can be used for the description of kinetics of zinc biosorption by yeast cells, for the ranges of zinc concentration of 2.0-16.0 g/l at 30 degrees C (mean correlation coefficient 0.96) and 10.0-16.0 g/l at 10 degrees C (mean correlation coefficient 0.95). PMID- 11253645 TI - Application of capillary electrophoresis for separating 1-O-glycopyranosyl sinapate from rapeseed. PMID- 11253646 TI - Antioxidant activity of essential oils from various spices. PMID- 11253647 TI - Nutrient distribution and zinc bioavailability. Estimation in some tropical edible mushrooms. PMID- 11253648 TI - Influence of added menadione (vitamin K3) on dissolution and dimerization of tocopherols and autoxidation of triacylglycerols during storage of plant oils. AB - The effect of added menadione (vit. K3) to stored rapeseed and soybean oil on the dissolution and dimerization of natural tocopherols and autoxidation of triacylglycerols was studied. Commercial rapeseed and soybean oils with added menadione and pure oil were stored at +20 degrees C in brown and transparent glass bottles. During storage their peroxide value, changes of the content of fatty acids and dissolution of tocopherols were measured. Destruction of individual tocopherols in tested oils with added menadione stored in both dark and transparent glass bottles were greater than in the oil samples without menadione. However, the degradation of individual tocopherols was different in each oil. Addition of menadione to the rapeseed and soybean oils resulted in the accelerated process of autoxidation of these oils. The oxidative state of the oils was lower in the oils stored in transparent bottles than in the oils stored in brown bottles. The prooxidant activity of menadione was additionally activated by light. Menadione added to the vegetable oils influenced the dissolution of natural tocopherols as well as their dimerization. It is conduced that addition of menadione to vegetable oils decreases their nutritive value. PMID- 11253649 TI - Sticking together and sorting things out: adhesion as a force in development. AB - During development it is not sufficient for cells to differentiate properly--they must also become physically grouped into appropriate structures, to form skin on the outside, and blood and muscle on the inside. How does this three-dimensional patterning occur? One classic explanation for this resolution of cells and tissues into distinct three-dimensional structures has been that as cells differentiate, they develop differential adhesive properties, and that these affinity differences allow cells to sort out from one another. This classic hypothesis is being investigated with increasing intensity, as recent work on the Drosophila wing and the vertebrate brain has shown that signalling between tissues is essential for the establishment of differential affinities. PMID- 11253650 TI - Capitalizing on large-scale mouse mutagenesis screens. AB - Variation is the crux of genetics. Mutagenesis screens in organisms from bacteria to fish have provided a battery of mutants that define protein functions within complex pathways. Large-scale mutation isolation has been carried out in Caenorhabditis elegans, Drosophila melanogaster and zebrafish, and has been recently reported in the mouse in two screens that have generated many new, clinically relevant mutations to reveal the power of phenotype-driven screens in a mammal. PMID- 11253652 TI - Natives or immigrants: modern human origin in east Asia. AB - East Asia is one of the few regions in the world where a relatively large number of human fossils have been unearthed--a discovery that has been taken as evidence for an independent local origin of modern humans outside of Africa. However, genetic studies conducted in the past ten years, especially using Y chromosomes, have provided unequivocal evidence for an African origin of East Asian populations. The genetic signatures present in diverse East Asian populations mark the footsteps of prehistoric migrations that occurred tens of thousands of years ago. PMID- 11253651 TI - Chemical genetics: ligand-based discovery of gene function. AB - Chemical genetics is the study of gene-product function in a cellular or organismal context using exogenous ligands. In this approach, small molecules that bind directly to proteins are used to alter protein function, enabling a kinetic analysis of the in vivo consequences of these changes. Recent advances have strongly enhanced the power of exogenous ligands such that they can resemble genetic mutations in terms of their general applicability and target specificity. The growing sophistication of this approach raises the possibility of its application to any biological process. PMID- 11253653 TI - Mobile elements and the human genome. AB - Genomic DNA is often thought of as the stable template of heredity, largely dormant and unchanging, apart from perhaps the occasional point mutation. But it has become increasingly clear that DNA is dynamic rather than static, being subjected to rearrangements, insertions and deletions. Much of this plasticity can be attributed to transposable elements and their genomic relatives. PMID- 11253654 TI - Molecular melodies in high and low C. AB - For 50 years now, one of the enigmas of molecular evolution has been the C-value paradox, which refers to the often massive, counterintuitive and seemingly arbitrary differences in genome size observed among eukaryotic organisms. For example, the genome of the fruitfly Drosophila melanogaster is 180 megabases (Mb), whereas that of the European brown grasshopper Podisma pedestris is 18,000 Mb. The difference in genome size of a factor of 100 is difficult to explain in view of the apparently similar levels of evolutionary, developmental and behavioural complexity of these organisms. PMID- 11253655 TI - Environmental health and genomics: visions and implications. AB - The relationship between genes and the environment can be compared to a loaded gun and its trigger. A loaded gun by itself causes no harm; it is only when the trigger is pulled that the potential for harm is released. Genetic susceptibility creates an analogous situation, where the loaded gun is one or a combination of susceptibility genes (alleles) and the trigger is an environmental exposure. The key objective of the Environmental Genome Project is to identify alleles that confer susceptibility to the adverse effects of environmental agents. Here we discuss the goals of the Environmental Genome Project, its implications and, in particular, its potential effect on our ability to assess human disease risk in the future. PMID- 11253656 TI - Engineering American society: the lesson of eugenics. AB - We stand at the threshold of a new century, with the whole human genome stretched out before us. Messages from science, the popular media, and the stock market suggest a world of seemingly limitless opportunities to improve human health and productivity. But at the turn of the last century, science and society faced a similar rush to exploit human genetics. The story of eugenics--humankind's first venture into a 'gene age'--holds a cautionary lesson for our current preoccupation with genes. PMID- 11253657 TI - Linkage disequilibrium. Highs and lows. PMID- 11253658 TI - Mitochondrial Genetics. A clever way to model defects.... PMID- 11253659 TI - Mitochondrial Genetics...and a new way to rescue them. PMID- 11253660 TI - That damned elusive polygene. PMID- 11253661 TI - Viral evolution. Not such a variable clock? PMID- 11253662 TI - Development biology. Nodal signalling gets foxy. PMID- 11253663 TI - Counting the calories to immortality. PMID- 11253664 TI - Spermatogenesis. Give me a break. PMID- 11253665 TI - A leg-up for crickets. PMID- 11253666 TI - Gene therapy: trials and tribulations. AB - The art and science of gene therapy has received much attention of late. The tragic death of 18-year-old Jesse Gelsinger, a volunteer in a Phase I clinical trial, has overshadowed the successful treatment of three children suffering from a rare but fatal immunological disease. In the light of the success and tragedy, it is timely to consider the challenges faced by gene therapy--a novel form of molecular medicine that may be poised to have an important impact on human health in the new millennium. PMID- 11253667 TI - [Liver surgery: technical and conceptional changeover]. PMID- 11253668 TI - [Dissection techniques in liver surgery]. AB - The first liver resection was performed in 1888. Since then a wide variety of dissection techniques have been introduced. The blunt dissection was replaced by novel methods, i.e. the CUSA technique and the Jet Cutter for major liver resections. These methods represent selective dissection techniques; whereas non selective methods include the scalpel, scissors, linear stapling cutter, high frequency coagulation, and the laser technique. The aim of this review article is the comparison of the different resection techniques in liver surgery, focussing on blood loss and resection time. PMID- 11253669 TI - [Anatomical and atypical liver resections]. AB - Liver resection has evolved to an established treatment for various malignant primary and secondary hepatic tumours, some benign tumours, and other conditions. The anatomical approach, the preferred concept of the author, rests on knowledge of the intrahepatic segmentation according to the portal structure branching and the course of major hepatic veins. As most of the malignant tumours respect the corresponding intrahepatic boundaries this resectional approach offers superior tumour clearance and, probably, better long-term outcome. Besides the four standard resections along the main fissure and left intersectorial plane, respectively, there are less common sector-orientated procedures including central hepatectomies and operations along the right intersectorial plane. Segment-orientated resections are defined by additional use of the transverse boundary according to the cranially and caudally directed third-order ramification of the portal trunks. Despite the advantage of anatomical resections there are rational indications for non-anatomical procedures such as removal of small benign tumours, excision of HCC in liver cirrhosis, re-resection following major hepatectomies, an excision biopsy in a non-resectable situation, and liver trauma care. Irrespective of the resectional approach, routine use of intraoperative ultrasound, maintenance of a low central venous pressure during parenchyma transsection, intermittent hilar clamping, and ischemic preconditioning all contribute to a safe and oncologically effective operation. In the future, augmentation of the liver remnant by preoperative portal vein embolisation, and multicentre trials on multidisciplinary strategies, may help to enhance resectability and to improve both safety and long-term outcome. PMID- 11253670 TI - [The anterior approach to hemihepatectomy]. AB - When performing right hemihepatectomy, the anterior approach can be employed as an alternative to the conventional procedure in which, before dividing parenchymal tissue, the right lobe of the liver is completely mobilized, the vena cava exposed from the lateral aspect, and the veins entering it from the right half of the liver divided outside the organ. In particular, in the case of large or capsule-broaching tumors of the right liver lobe, exposure of the retrohepatic vena cava from the anterior aspect facilitates work on disection of the hepatic veins. Since with this approach the right lobe of the liver does not need to be removed out of its bed, perfusion of the remnant liver and the venous return via the vena cava remain unimpaired during the entire operation. Since manipulation of the tumor is minimal (no-touch technique), the risk of iatrogenic tumor perforation or hematogenous dissemination of malignant cells during surgery is reduced, and a surgical radical en bloc resection is facilitated. These technical and oncological benefits of the anterior approach must, however, be weighed against the increased risk of bleeding from hepatic veins during divisioning of the parenchyma, since, owing to the fact that the right lobe of the liver is not mobilised, such bleeding is frequently difficult to deal with. PMID- 11253671 TI - [Ex situ resection and resection of the in situ perfused liver: are there still indications?]. AB - Most liver tumors can be removed with conventional resection techniques employing partial or total vascular occlusion when needed. Duration of tolerable warm ischemia has not yet been defined, but it seems to be well tolerated up to 60 min. In a few cases with extended vascular resection and reconstruction liver protection by hypothermic perfusion is advantageous. This can be achieved by in situ perfusion, ante situm resection or ex situ resection. Major reconstruction of hepatic vessels with good technical access should be performed under in situ hypothermic protection using veno-venous bypass. Tumors involving the hepatic venous confluence and/or retrohepatic vena cava should be approached by either the in situ, or preferentially, the ante situm resection technique. The indication for an ex situ liver resection resulting in autotransplantation of the remnant liver exists only in rare cases for oncological reasons. PMID- 11253673 TI - [Laparoscopic gastric banding using a modified gastric band with loops and a microport system. A report on 25 cases]. AB - INTRODUCTION: The laparoscopic application of an adjustable silicone gastric band is an established procedure in the surgical treatment of pathologic adiposity. PATIENTS AND METHODS: Twenty-five patients with morbid obesity underwent the laparoscopic application of a gastric band between November 1998 and June 2000. A modified band (GastroBelt II) was used. RESULTS: Early and late complications were rare in comparison with previous procedures. Complications, which often require surgical intervention, such as slipping of the stomach (8-12%) or pouch dilatation (2-4%), were not observed. The total morbidity rate and mortality rate were both 0%. CONCLUSION: Critical selection of the patients before the operation and appropriate compliance produced an average loss of weight of 7% of the overweight at 6 weeks, 20% at 3 months and 28% at 6 months after the operation. Preconditions for this operation are a coordinated operation team and a clearly defined standardized postoperative care concept. PMID- 11253672 TI - [Therapeutic strategies in malignant soft tissue tumors. Results of the soft tissue tumor register study of the Surgical Oncology Working Group]. AB - INTRODUCTION: This study, carried out by the Surgical Oncology Working Group (CAO) of the German Society for Surgery, was performed to analyse the strategies in the treatment of soft tissue sarcomas in adults. METHODS: In a period of 19 months the data on 292 patients suffering from soft tissue sarcomas, treated in 99 surgical departments in Germany, were analysed prospectively. A special questionnaire was developed including pretherapeutic biopsy, previous treatment, definitive surgical treatment, combined modality approach and histopathological results. RESULTS: Thirty-nine per cent of the tumours were treated in university hospitals, 36% in medical centres, 24% in regional hospitals. During the observation period two patients were treated on average (median) by each hospital. Limb-sparing treatment was performed in 96% of the extremity tumours. There was no significant difference in the frequency of R0 resections between the different hospitals. At the university hospitals local extended operations and additive measures were used more often. The indication for adjuvant radiotherapy differed: after compartmental resection, adjuvant radiotherapy was performed in 39% of cases (19/49); after wide-excision of high-grade tumours, in 45% of cases (20/44) no adjuvant radiotherapy was necessary. In spite of less radical treatment in tumours of the trunk, additional radiotherapy was not more frequently performed. CONCLUSION: To improve the quality in the treatment of soft tissue sarcomas it seems to be of great importance to avoid inadequate initial treatment (18%), to respect the rules of oncological surgery (tumour rupture in 7% of cases), to improve the histopathological examination (no R classification in 5-12%) and to develop guidelines for multimodality treatment. PMID- 11253674 TI - [Contribution to the nomenclature of variations of the cystic artery]. AB - INTRODUCTION: Safe procedures for laparoscopic cholecystectomy demand good knowledge of the anatomy of the terminal part of the cystic artery and its variations, and also precise dissection in and around the hepatobiliary triangle. METHOD: Good laparoscopic visualisation enables recognition of the variation of the cystic artery. Our observations are based on 1000 cholecystectomies. RESULTS: We have described and named variations of the terminal part of the cystic artery. Group I comprises the five variations of the cystic artery within the hepatobiliary triangle: (a) "normal" position; (b) frontal cystic artery; (c) backside; (d) multiple; (e) short cystic artery that arises from an aberrant right hepatic artery. Group II consists of variations of the cystic artery that approach--the gallbladder beyond the hepatobiliary triangle: (a) "low-lying"; (b) transhepatic; (c) "recurrent" cystic artery. CONCLUSION: Our classification is simple and easy to memorize and will considerably facilitate safe laparoscopic cholecystectomy. PMID- 11253675 TI - [Spiral CT cholangiography is not suitable for routine diagnosis before laparoscopic cholecystectomy]. AB - INTRODUCTION: The purpose of this prospective controlled study was to evaluate the diagnostic potential of spiral computed tomographic (CT) cholangiography in patients undergoing laparoscopic cholecystectomy. METHODS: 60 patients (17 men, 43 women, mean age 54.5 years, range 15-84 years) with symptomatic cholecystolithiasis were included in this study. After infusion of meglumine jodoxamate, all patients underwent upper abdominal spiral CT. The results of the spiral CT scan were then compared with endoscopic retrograde cholangiography (ERC) or intraoperative cholangiography. RESULTS: In 53 patients (88%) CT cholangiography was considered to be technically adequate for interpretation, but was suboptimal in 4 patients (7%) and nondiagnostic in 3 patients (5%), respectively. CT cholangiography showed a stone free common bile duct in 51 patients which was correct in all cases. CT cholangiography predicted a common bile duct stone in 6 patients which proved to be correct in 4 patients but was found to be incorrect in 2 patients. CONCLUSION: Spiral CT cholangiography is useful for the diagnosis of common bile duct stones. Because of the low positive predicting value routine use before laparoscopic cholecystectomy is not justified. PMID- 11253676 TI - [Hypothermia in patients with burn injuries: influence of prehospital treatment]. AB - Hypothermia following pre-hospital treatment of burn patients is a common risk with increasing lethality. Soon after admission to our burn unit, the body temperature of 212 adult patients with more than 5% total body surface area burned was documented. We found no influence of the time of pre-hospital care and cold-water treatment alone on the body temperature. If the patients were not anesthetized, the initial temperature was normal. Only the anesthetized and artificial ventilated patients were hypothermic. We conclude that hypothermia is not a problem of the non-anesthetized and cold-water-treated patient. However, all anesthetized patients must be carefully treated to avoid hypothermia as an important complication in the pre-hospital management. PMID- 11253677 TI - [Surgical concepts and results in necrotizing fasciitis]. AB - INTRODUCTION: Necrotizing fasciitis (NF) is a rapidly progressive soft tissue infection involving primarily the superficial fascia and subcutaneous tissue. The disease is caused by Streptococcus pyogenes or synergistic infection of anaerobic and facultative anaerobic bacteria. Further characteristics are severe, intolerable pain and a mortality of 30-50%. PATIENTS AND METHODS: From January 1996 to January 2000 12 patients underwent treatment for NF. The patients' charts were investigated retrospectively. RESULTS: In 7 patients the bacterial cultures showed a mixed, polymicrobial infection and in 5 cases only Streptococcus pyogenes. The NF was localized at the upper extremity (2), abdomen (3), back (1), hip (2) and lower extremity (4). The area involved was 8 (4-11)% of the total body surface. The surgical procedures in 12 patients were debridement (60x), local transposition flap (2), free muscle flaps (3), lower leg amputation (1) and split thickness skin graft (3x). Four patients developed streptococcal toxic shock syndrome and two died. In total there were four deaths with a mortality of 33%. In the "survivor group" the time to diagnosis was 2.8 (1-7) days, the time to radical surgery 3.3 (1-9) days. In the "mortality group" it was 6.8 (3-10) days or 9.3 (6-12) days. CONCLUSION: The prognosis of NF seems to be influenced by the site of the infection, because 4 out of 6 patients with NF of the abdomen, back or hip died, but all patients with NF of the extremities survived. The age of the patient is not a key parameter, because also young and previously healthy people also die from the streptococcal toxic shock syndrome. The interval between diagnosis and radical debridement appears to be the crucial factor in terms of prognosis, since early diagnosis and prompt, radical surgery improves the survival rate. PMID- 11253678 TI - [Pedicled flap plasty in closure of defects after recurrence of tumors of the back]. AB - Resection of recurrent tumors of the back often results in large transmural defects. Sometimes, radiation therapy is applied after the first operation, leading to indurated, badly vascularized skin and surrounding tissue, often complicated by infection. For this reason, it is necessary to remove not only the tumor but also the surrounding tissue. To reconstruct such wide defects, we use large axial pattern flaps with extension through a fasciocutaneous flap. This leads to safe reconstruction of a complex defect with well-vascularized tissue. We report on two patients with recurrent tumors of the back, reconstructed with combined, extended axial pattern flaps. PMID- 11253680 TI - [Superior mesenteric vein syndrome: a case of duodenal stenosis caused by atypical malposition of the superior mesenteric vein]. AB - Cases in which mesenteric vessels lead to stenosis of the duodenum are very rare. Several cases have been reported of patients suffering from stenosis of the last third of the duodenum due to a malpositioning of the superior mesenteric artery or the left renal vein. We report a 78-year-old patient who was suffering from dyspepsia, pain in the upper abdominal region, nausea, and vomiting. The medical history revealed that the patient had undergone a subtotal gastrectomy according to Billroth II at the age of 19 because of similar complaints. In the last 20 years the patient had to be laparotomized several times for ileus of the small intestine. Now the patient presented abdominal complaints with nausea and pressure in the upper abdominal region. Assuming an efferent-loop syndrome and adhesions, the patient was laparotomized. We discovered malpositioning of the superior mesenteric vein, leading to stenosis of the superior part of the duodenum. In fact, 60 years ago surgeons performed a duodenojejunostomy, circumvening the stenosis of the duodenum. With a "delay of 60 years", we then performed a subtotal gastrectomy according to Billroth II. The postoperative course was uneventful; the patient had no complaints and increased in body weight. To our knowledge, this is the first time that a stenosis of the duodenum due to malpositioning of the superior mesenteric vein has been observed. PMID- 11253679 TI - [The Longo and Milligan-Morgan hemorrhoidectomy. A prospective comparative study of 300 patients]. AB - INTRODUCTION: Stapled haemorrhoidectomy according to Longo for treating reducible anal and mucosal prolapse appears to be very simple technically. In order to investigate the feasibility and to delineate the value of this procedure it was compared to standard open haemorrhoidectomy according to Milligan-Morgan in a prospective study. METHOD: In 1998 and 1999, 300 patients with third-degree haemorrhoids were operated on either with a Milligan-Morgan or a Longo technique. Intraoperative and postoperative performance--complications, length of surgery, consumption of analgetics, hospital stay, return to work--were evaluated. Follow up was 1, 3 and 6 months. RESULTS: Length of surgery for both types of operation was the same. In comparison with the conventional open excision, patients with a stapled haemorrhoidectomy required considerably less analgetics. Hospital stay was shorter and return to work quicker and there were fewer morphological residues, e.g. skin tags. However, the costs of the procedure were considerably higher because of the disposable instrument. CONCLUSION: This new surgical procedure of supra-anodermal resection according to Longo offers advantages in the repair of prolapsing haemorrhoidal disease. Despite its primary simplicity, in our opinion the surgeon has to have colorectal, especially proctologic, experience. Although costs are significantly higher, this procedure might replace conventional techniques in many cases with prolapsing haemorrhoids. PMID- 11253681 TI - [Sweat gland carcinoma in the axillary area. A case report and review of the literature]. AB - The sweat gland carcinoma is a rare tumor, representing in approximately 1% of primary skin lesions. In the literature 220 cases of sweat gland carcinomas have been presented over the last three decades. The medium age is 57 years, with an equal male-to-female distribution. Topographically, the tumor is located at the lower limbs (32.9%), the upper extremities (28%), and the head (26%). Involvement of the trunk is rare. Diagnosis can be complicated as the carcinoma is a rare entity, with no correlation of its histologic classification and biologic presentation. This can only implicate the difficulties in treatment. PMID- 11253682 TI - [Soft tissue chondromatosis of the foot: diagnosis and therapy]. AB - Chondromas of the soft tissue are uncommon, benign tumors. They are most frequently found adjacent to periarticular tissues or tenosynovium with a predilection for the hands and feet. We report a case of a patient with a large, increasing tumor of the left foot. Because of subjective complaints and mechanical irritation the patient was scheduled for surgery. The preoperative radiological work-up showed popcorn like clusters of calcifications around the dorsum of the foot. The tumor mass was reduced using several incisions. Histology showed soft tissue chondromatosis of the foot without malignancy. Following the operative procedure, the patient was free of pain and able to wear normal shoes. This case illustrates the clinical and radiological characteristics as well as the surgical treatment of progressive soft tissue chondromatosis. PMID- 11253684 TI - [Comments on the subject of the thyroid gland: "Monitoring of the recurrent laryngeal nerve" and "Evidence-based medicine" [7,10,14]]. PMID- 11253683 TI - [An update of the German version of AEP (Appropriateness Evaluation Protocol): metric properties and practical experiences]. AB - The development of a German version of the Appropriateness Evaluation Protocol (AEP) allows for the first time the evaluation of hospital admissions and bed days in Germany. The instrument is based on international experience and has been adopted in cooperation with acknowledged members of German surgical and medical societies. The AEP showed excellent reliability in general internal medicine as well as in surgery. The validity is comparable to international studies, although further research is necessary. Approximately 90% of surgical cases could be evaluated according to the criteria of the AEP; the remaining patients were evaluated using the "override option". The majority of inappropriate care is due to poor documentation in medical records and management deficiencies during inpatient care. PMID- 11253685 TI - [Artificial pacemaker implantation]. PMID- 11253686 TI - [Effects of the DRG-based payment system. What action requirement exists for the surgeon?]. PMID- 11253687 TI - [Surgery 2010. Study on the development of surgeon demand until the year 2010]. PMID- 11253688 TI - [The troublesome expert]. PMID- 11253690 TI - [On the problem of prophylaxis of late effects induced by internal irradiation using immunomodulators]. AB - Using the experimental modeling of late effects induced by intake of radionuclides (131I, 238,239Pu, 90Sr) the attempt to investigate the possibility of protection using immune modulators was made. Co-polymer of 2-methil-5 vinilprilidin and N-vinilpirrholidon (sovidon), G immunoglobulin and interferon, Fc fragment of immunoglobulin and S. parathyphi B. Breslau vaccine were used as immune modulators. The positive prophyletie influence of immune modulators was found for average life span, radiation action, immune status, of rates of occurrence and developments of breast tumors induced by 131I, 239Pu and 90Sr. At the same time the increase in osteosarcoma cumulative rate induced by immune modulators in case of body intake of 238Pu and 90Sr was registered. PMID- 11253689 TI - [Effect of prolonged gamma-irradiation on the functional state of heart and its adrenergic regulation at hypothyroidism]. AB - The study of heart isolated by Langendorf's method has shown that the prolongated gamma-irradiation of euthyroid rats in 1.0 Gy dose (2.8 x 10(-7) Gy/sec) causes the decrease in contraction ability, myocardium relaxation and functional response of heart to the stimulation of beta-adrenergic receptors, and the increase in myocardium reaction to the stimulation of alpha-adrenergic receptors. The irradiation of hypothyroid animals leads to more significant and long changes in contraction function of heart and its adrenergic regulation. PMID- 11253691 TI - [Study of 137Cs transfer to the rat offspring with breast milk and a modifying effect of ferrocene on this process]. AB - 137Cs transfer to the lactating rats' offspring has been investigated following daily administration of the radionuclide to the females. Certain patterns have been established for 137Cs accumulation dynamics in the female rats and their progeny during 30 days of suckling. Radiocaesium accumulation multiplicity in both female rats and offspring, as well as absorbed dose formation and a modifying effect of ferrocinum on these processes have been studied. PMID- 11253692 TI - [Experimental study of the intensity of phosphorus radionuclide uptake by samples of algobacterial community of the Yenisei river]. AB - The experiments in which phosphorus radionuclide was added to samples of algobacterial community of the Yenisei River taken near the production area of the Mining-and-Chemical Combine (Zheleznogorsk) made possible determining the uptake rates and coefficients of radionuclide accumulation by microorganisms. Ratios between processes of adsorption and accumulation of phosphorus radionuclide by components of water seston (suspended matter) have been determined. The portion of the specific radioactivity of phosphorus adsorbed by unit mass of seston (and algae) has been found to be not more than 7% of the activity accumulated by algal cells. PMID- 11253693 TI - [APUD- and RAA- regulatory systems in acute radiation injury]. AB - An experimental morpho-functional assessment of rat APUD and RAA systems status in acute radiation injury (cerebral form) was performed. The named regulatory systems were found to display an actual momentary post-exposure reaction, followed by subsequent functional changes of a distinct phase character. The latter were shown to determine in many respects the clinical picture and in some cases the outcome of the disease. PMID- 11253694 TI - [Effect of lysophosphatidylcholine on the radiation-induced lipid peroxidation in liposomes]. AB - The effect of lysophosphatidylcholine (LPC) on the lipid peroxidation process (LPO) in liposomes of rat liver phosphatidylcholine initiated by irradiation of 137Cs source was studied. The formation of diene conjugates (DC) is shown to increase dramatically on incorporation of more than 10% LPC into liposomes. The dependence of DC proportion on the irradiation dose is practically linear in the range of 0 to 5 kGy. The DC concentration in the liposomes without LPC grows at least to dose of 3.3 kGy and is unchanged on further irradiation. The malonic dialdehide accumulation follows the similar dependence. The LPC effect is neutralized by the incorporation of cholesterol into liposomes. The product of free radical fragmentation of LPC, palmitoxyacetone, practically has no influence on the DC concentration. The reasons of LPC effect on the irradiation initiated LPO in liposomes is discussed. PMID- 11253695 TI - [The role of labile complexes with oxygen in antioxidant activity of carotenoids]. AB - The present research work was done with the main purpose to study early stages of interaction of carotenoids (Car) with molecular oxygen and clarify their role in the mechanism responsible for Car radiochemical stability and carotenoid ability to decrease concentration of the most active oxygen transients like superoxide anion radicals (O2.-). Alcoholic and phosphate buffer (pH 7.5) solutions of carotenoid fucoxanthin (Fx) were used for investigation of the oxygen effect on the absorption spectra in the UV-Visible range. Special analysis of time dependent reversible shifts of absorption bands of evacuated Fx solution after contact with O2 indicated existence of equilibrium between two distinct forms of Car: Fx and the labile charge transfer complex (Fx+delta...O2-delta). The velocity of the achievement of equilibrium state and a degree of reversibility depend on chemical structure of the carotenoid, oxygen content and the solvent nature. Radiation-chemical methods were used to confirm the important role of primary Car oxocomplexes in different redox processes. It appeared that the yield of radiation-chemical bleaching of Fx, G-Fx, is 0.02-0.05 molecule/100 eV in the presence of oxygen, which in hundred times less the yield achieved in anaerobic conditions. The obtained results provide the evidence of Fx high level of stability under radiation, and demonstrate the supreme importance of reversible oxocomplex (Fx+delta...O2-delta) in stabilizing carotenoids in aerobic medium. The pulse radiolysis method with spectrophotometric registration of transients was used for generation and studying of mechanism O2.- interaction with different carotenoids. Introduction of any carotenoids containing oxygen (10(-5) M) in phosphate buffer solutions (pH 7.5) caused a red-shift of absorption maximum (from 5 to 15 nm) and difference in kinetics of O2.- decay. These results prove that radiation generated esolv- are directly accepted by (Car...O2) with consequent formation of superoxocomplexes (Car...O2.-) instead of O2.-. On the base of detecting the following transformation of superoxocomplexes the peroxocomplex (Car+...O2(2-)) was identified. In case of Fx a peroxocomplex (Fx+...O2(2-)) had absorption band with lambda max at approximately 360 nm. It is very important to mention that beta-carotene does not cause the similar effect and gets easily oxydized when exposed to the air. PMID- 11253696 TI - [Cytosol reception of estradiol and nuclear acception of estrogen-receptor complexes in target organs of female rats after gamma-irradiation]. AB - Condition of cytosol reception of estradiol and nuclear acception of estrogen receptor complexes in target organs of female rats subjected to gamma-radiation was studied. It was established that the action of ionizing radiation of 0.5 and 1.0 Gy dose (dose rate 6.2 sGy per minute) was combined with a decrease (liver, uterus) of cytosol estrogen-receptor complexes and their modification and that those effects were realized through the lessening of the nuclear acception capability in liver cells of radiation-exposed animals. PMID- 11253698 TI - [Analysis of stable chromosome aberrations using G-banding stain in lymphocyte of patients with high dose radiation injury]. AB - The analysis was performed on 514 blood lymphocytes from a person accidentally exposed to 137Cs. Blood samples were collected 1 year after exposure three times at intervals of one month. Terminal deletions and simple translocations were found to predominate in all cases. No differences between these cases were observed on analysing total frequency of stable chromosome aberrations. However, the frequency of terminal deletions decreased and frequencies of exchange-type aberration increased with time after exposure. Chromosome #4 was more involved in stable aberrations than it would be expected from the relative chromosome lengths. Clonal aberrations del-ter (5)(q31 or 32) were found. PMID- 11253697 TI - [Effect of microbial derived agents on the level of blood cytokines, hematological status and survival of mice following combined radiation injury]. AB - Mice models of acute radiation and combined radiation injuries (whole body irradiation + thermal burn) were used for the experiments. Blood serum response of IL-1 beta, TNF-alpha, IL-6, IL-3, and GM-CSF was evaluated after single injection of bacterially derived products (BDPs). As it was established, BDPs revealed different ability for cytokine increasing activity: Imuvert (extract from Serratia marcescens) > synthetic trehalose dicorynomycolate > heat-killed Lactobacillus acidophilus. The capacity of BDPs to enhance survival of animals did not depend on its cytokine-inducing and hematopoietic activities. PMID- 11253699 TI - [Mechanisms of cerebral radiation syndrome]. AB - Postradiation DNA breaks activate the repair enzyme, Adenosine diphosphoribosil transferase, which consumes NAD as a substrate and causes neuronal NAD and then ATP pool depletion; here this is considered to be the crucial links of the cerebral radiation syndrome (CRS) mechanism. Two ways of its metabolic correction were examined: (a) prevention of postradiation NAD depletion by administration of the enzyme inhibitor and (b) shunting NAD-dependent oxidative phosphorilation path of ATP resynthesis by administration of substrate of NAD-independent oxidation. PMID- 11253700 TI - [State of hemopoiesis under exposure to low doses of ionizing radiation and emotional stress during treatment with anxiolytic aphobazole]. AB - In experiments on rats it was found that aphobazole administered before emotional stress is capable to stop violations of adaptive reactions and compensator capabilities of a hemopoietic system in conditions of development of an emotional stress in early terms after gamma-irradiation with a dose of 0.9 Gy. PMID- 11253701 TI - [Effect of natural radioprotector carnosine on histamine-diamine oxidase system of rat's myocardium after action of various extreme factors]. AB - The unbalance in system histamine-diaminooxydase in myocardium of rats subjected to various stress actions (intensive cold influence, hyperthermia and hypoxia) was found. The preventive application of a natural radioprotector carnosine prevented apparent changes, that gives the basis to assume about carnosine ability to enlarge a nonspecific resistance of an organism. PMID- 11253702 TI - [Effect of low-intensity pulse-modulated microwave on human blood aspartate aminotransferase activity]. AB - Pulse-modulated microwaves (frequency 2375 MHz, intensity: 2 microW/cm2 and 8 microW/cm2, pulse modulation from 50 to 390 Hz with step of 20 Hz; exposure time 5 min) changed the activity of aspartataminotranspherase of the donor blood. Aspartataminotranspherase activity was strongly dependent both on modulation frequency and microwave intensity. Maximum activity was found at 390 Hz and 8 microW/cm2. Maximum observed activity was about six times greater than control level of activity. PMID- 11253703 TI - [Effect of low intensity pulse-modulated electromagnetic radiation on activity of alkaline phosphatase in blood serum]. AB - The change in alkaline phosphotase activity in vitro with frequencies modulation at low intensity of pulse-modulated electromagnetic radiation was experimentally shown (EMR, 2375 MHz, intensity: 0.8, 8.0; 40.0 microW/cm2; range modulation: 30 310 Hz; time of interaction: 1-3 min). Revealed effects could be regarded as an evidence of informative character of interaction of modulated EMR. PMID- 11253704 TI - [The study of some mechanisms of injurious effects of low-frequency noises]. AB - Low-frequency noise has been shown to cause certain functional changes in the organism of laboratory animals that manifest in changes of the state of the regulatory systems and metabolic disturbances at cellular and subcellular levels. The obtained data support the hypothesis of the mechanism of injurious effect of this physical factor including two basically correlated ways, i.e. the central mechanism associated with overexcitation of the hypotalamohypophysis-adrenal system and mediating the homeostatic parameters of the body, and the local one which is determined by a direct effect of low-frequency noise on highly organized structure of membraneous and genetic apparatus of cells. PMID- 11253705 TI - [Modifying effect of low-intensive electromagnetic radiation on the irradiated cells]. AB - The normalising effect of low-intensive electromagnetic radiation on the viability of cell culture of L929 line, irradiated with gamma-quantums of 137Cs in a wide dose range was demonstrated. PMID- 11253706 TI - [Oligomeric proteins: modulation of UV-sensitivity and characteristics of photochemical transformations]. AB - The results of our research devoted to the study of photophysical and photochemical transformations of some compound proteins of oxidative, antioxidant, transport and immune systems of an organism under conditions of different microenvironment (presence of chemical modificators, the change of pH, temperature, doses and spectral composition of ultraviolet irradiation) have been analyzed. Methods and the degree of modulation of UV-sensitivity of proteins of the mentioned systems have been discussed. Conditions, on which the used chemical agents show photoprotective and photosensitizing effect on protein molecules, have been found. PMID- 11253707 TI - Labelled leukocytes for diagnosis of infectious diseases. Our experience in labelling and clinical usefulness. AB - BACKGROUND: To review our experience in infectious diseases diagnosis, using a simple labelling technique. METHODS: We made 101 scans in 91 patients with suspected infectious diseases confirmed by clinical, histologic or cultural specimens; a clinical or instrumental follow-up was available for every patient. Patients were divided into four subgroups: A: Inflammatory Bowel Diseases; B: Septic Syndromes; C: Bone diseases; D: Others. We used 99mTc-HMPAO in 80 scans and 111In in 21. In 20/80 frozen and stored HMPAO was used. RESULTS: 99mTc-HMPAO frozen and stored labelling yield was: 60%(SD15%), 99mTcHMPAO: 62(12)(p n.s.), 111In 75%(10%), (p < 0.05). Frozen-stored HMPAO was sterile and apyrogen. 27 had positive scan without leukocytosis or neutrophilia. No correlation between leukocytosis or neutrophilia and yield was observed. Transit lung times ranged between 14-16 min without differences among three radiopharmaceuticals. In each Group and in the sample as a whole True Positive, False Positive, True Negative, False Negative, Sensitivity, Specificity and Accuracy, were calculated. In Group B and in the sample as a whole Predictive Positive Value and Negative Predictive Value were also evaluated. CONCLUSIONS: Labelling yields and "viability" were good using the three radiopharmaceuticals; labelling procedure was simple and safe. Accuracy PPV, NPV, cost-effectiveness were good; 111In is the choice for diagnosis of bone and joint infections. Frozen-stored HMPAO should be introduced as a cost-saving labelling procedure in practice. PMID- 11253708 TI - [The T index in the elderly with altered values of free thyroxine and/or thyrotropin. A new thyroid diagnostic-therapeutic index]. AB - BACKGROUND: Routinary evaluation of FT4 and TSH, in elderly in-patients without thyroid disease shows frequent, isolated and, often unclear, changes from normal values of plasma TSH and FT4 concentrations. A new parameter called "T index" derived from the product of FT4 and TSH values has been used by the authors. In a previous work they studied "T index" variations in 1257 elderly subjects with normal FT4 and TSH levels. They have determined the "T index" theoretic model of distribution and identified a normality range (values from 5.78 to 50.76), a suspect range (values from 2.92 to 5.78 and from 50.76 to 68.6) and a pathologic range (values less than 2.92 and more than 68.6). Aim of this study was to investigate "T index" variations in a group of 357 elderly subjects with altered FT4 and/or TSH levels. METHODS: Patients were divided into eight groups according to different concentrations of FT4 and/or TSH levels. "T index" results were expressed as mean, standard deviation, maximum and minimum values. Patients were therefore divided into three groups (normality range, suspect range, pathologic range) according to "T index" distribution as previously described. RESULTS: In 20% of elderly people hospitalized, we found alterations of thyroid hormones levels represented mostly by: a) FT4 normal; TSH low. b) FT4 normal; TSH high, c) FT4 low; TSH normal. In case of hypothalamus-hypophysis hypothyroidism and in case of hyperthyroidism, "T index" score is, usually, less than 2.92, whereas in case of hypothalamus-hypophysis hypothyroidism and hypothyroidism "T index" score is, nearly always, more than 68.6. When TSH levels are into the range of normality "T index" score has always normality levels. The "T index" helps us to divide patients with subclinical hyperthyroidism into two groups: one with values from 5.78 to 2.92 (suspect range), and the other, with values less than 2.92 pathologic range). In patients with subclinical hypothyroidism, some patients have values from 50.76 to 68.6 (suspect range), other patients have values more than 68.6 (pathologic range). CONCLUSIONS: In case of subclinical hypothyroidism or subclinical hyperthyroidism the use of "T index" seems to be a good way to select the cases in which it is better to start pharmacological treatment (hormonal replacement or thyroid inhibition) from those which is better to follow up. PMID- 11253709 TI - [Sex hormones and male osteoporosis. A physiologic prospective for prevention and therapy]. AB - BACKGROUND: Male osteoporosis is often an underestimate and non-acknowledged pathology because it is less frequent then post-menopausal women osteoporosis. The causes of osteoporosis in males were reviewed, considering the importance of the assessment of sexual hormones, even without symptoms of hypogonadism. METHODS: Sixty-two patients ranging in age from 45 to 75 years, males, were studied. None of them had assumed steroids nor other drugs causing osteoporosis. These patients have been subjected to bone mineral analysis (BMA) together with the following sexual hormones: LH, FSH, total testosterone, free testosterone, SHBG, estrone, estradiol. Two methods of BMA have been employed (Norland 2780 and UBA 575 Walker and Sonix). RESULTS: When osteoporosis or osteopenia were present, always there were modifications of sex hormones values. Statistic evaluation (linear regression, Pearson coefficient, Multiple regression, Backward Stepwise Multiple Regression), showed that there was a significant association between total testosterone and osteoporosis. CONCLUSIONS: Total testosterone resulted the most predictive sex hormone for the loss of bone mass; therefore, it is important to evaluate sex hormones in males with osteoporosis, for a correct and "physiological" therapy. PMID- 11253710 TI - [Evaluation of the efficacy and tolerance of radioactive fangotherapy in gonarthrosis. Comparative study versus short wave therapy]. AB - BACKGROUND: Spa therapy is frequently used in daily rheumatological practice, but its benefit remains to be evaluated. The purpose of this paper is to evaluate the effectiveness and tolerability of mud-packs and mineral baths with fluorurate radioactivity water from the thermal resort of Merano (Bolzano) versus short wave therapy in patients with osteoarthritis of the knees. METHODS: Forty-eight patients were treated for a period of two weeks with mineral water baths and mineral mud-packs and twenty-four patients were treated with short-wave therapy for the same period. Assessment criteria were spontaneous pain ratings on a visual analogue scale (VAS), functional impairment (Lequesne's index), quality of life index (AIMS1), analgesic and/or non-steroidal anti-inflammatory drugs consumption and patient and physician assessment of effectiveness and tolerability of treatments. These criteria were recorded at the beginning and at the end of the treatments and after 3 months. RESULTS: A significant improvement (p < 0.0001) in the Lequesne's index, in the VAS and a significant decrease in analgesic consumption was achieved in both groups up to 15 days. The improvement remains to the end of the follow-up period only in the group treated with spa therapy. CONCLUSIONS: This study suggests that spa therapy has a prolonged, beneficial, symptomatic effect in osteoarthritis of the knees. PMID- 11253711 TI - [The role of telomere-binding proteins in carcinogenesis]. AB - Normal somatic cells have a defined number of divisions, a limited capacity to proliferative. The telomeres, sequences of TTAGGG repeats at the ends of chromosomes, are considered the direct responsible of the control of the cellular cycle. In fact, the progressive shortening of telomere length at each cellular division, causes the entrance of the cells in a phase of senescence and than apoptosis. The maintenance of the length of telomeres is carried out through: the telomerase, a DNA polymerase reverse transcriptase that extends sequence TTAGGG repeats, or the alternative lengthening of telomeres (ALT), between which the adaptive mechanisms, inactivation of TRF1, a protein bound to the telomeres with the functions of inhibiting the telomerase activity and Tankirase-PARP, an enzymatic complex that ADP-ribosylate TRF1 and reduce its binding to DNA. The alteration of the mechanism of maintenance of the telomeres length (Telomerase, TRF1, Tankirase-PARP) may represent a first step toward the cell immortalization and cancerogenesis. Together with the alteration of the control mechanisms of the telomere length, also the cell genic contest should be considered. In fact, the oncogene activation and/or oncosuppressor gene inactivation (p53, Rb, ras) may allow or reduce the cancerogenesis. From this point of view, the telomerase, the TRF1, Tanchirase-PARP and other proteins involved in telomere length could be, in a near future, used as new indicators of prognosis and as markers for new anti cancer therapies. PMID- 11253712 TI - [Rehabilitative intervention after a myocardial infarct]. AB - The purpose of the present review was to determine whether exercise training improves cardiac function in patients with prior myocardial infarction. Home exercise programs for patients with myocardial infarction effectively improve their ability to exercise as well as quality of life. A computer-based, automated, telemetry system comprising central and peripheral computers and telephone line was developed. Myocardial infarction patients were evaluated for peak oxygen uptake and anaerobic threshold. Some studies have in fact suggested that using echocardiography, exercise training in patients with reduced left ventricular function after a myocardial infarction leads to further myocardial damage, including wall thinning, infarct and expansion. A more recent analysis by these investigators suggested that training actually has a beneficial effect on the remodeling process. Many factors appear to influence the extent of the remodeling process, including attenuation by Ace inhibition therapy, extent and site of the infarction, hypertension and continued ischemia. Some results suggest early physical training may be safe and improve the autonomic nerve balance and exercise tolerance in patients with acute myocardial infarction. PMID- 11253714 TI - [Surgical aspects as a solution to carotid occlusive pathology. Clinical case]. AB - A significant clinical case presenting carotid occlusion pathology; successfully treated with the most recent surgical techniques is reported. The etiopathogenetic problem of this pathology which is of remarkable scientific interest in the medical literature is underlined. PMID- 11253713 TI - [Clinical approach in hypertensive elderly patients]. AB - Hypertension in the elderly represents a cardiovascular risk factor which increases due to ageing and to the raise of blood pressure (BP) values. The occurrence of hypertension depends on an interaction between genes and environment. An available antihypertensive therapy causes a reduction in the incidence of cardiovascular events. An antihypertensive therapy in the elderly must take into account: in these subjects BP might be spontaneously lower over 30 mmHg in 24 hours; people normally have a postprandial BP reduction; sudden raises or falls of pressure cause cerebral hypoperfusion; some adverse vents of hypertensive drugs worsen their quality of life, not reducing myocardial hypertrophy; possible electrolytic troubles might worsen a congestive heart failure; drastic diets cause a raise in the incidence of colorectal tumours; a high heart rate increases the risk of sudden death; a chronic NSAID intake might cause or aggravate a hypertensive state; a reduction of natrium chlorure and lipides in the diet might cause a BP fall. In short, the BP reduction should be gradual in the hypertensive elderly in order to avoid the occurrence of cardiovascular events, diets should be balanced, rich in fibres and vitamins to avoid colorectal tumours. Besides, NSAID must be used by these patients for a short time and all therapeutic interventions should improve their quality of life. PMID- 11253715 TI - Hyperpyrexia secondary to acute hypocalcemic status. Is there a role for calcitonin therapy in bone metabolism derangements due to anticonvulsant drugs? AB - Hypocalcemia, hyperparathyroidism, hypovitaminosis D, hypocalcitoninemia and decreased bone mass are side effects of several anticonvulsant drugs. Since calcitonin inhibits the mineral mobilization of bone and augments minerals bone content, combined therapy with calcitonin, calcium, vitamin-C and vitamin-D was administrated to a patient with severe anticonvulsant disturbances of bone metabolism. Calcitonin hypersensitivity was evident. The symptomatology, characterized by the rare hypocalcemic hyperpyrexia, regressed after calcium infusion. PMID- 11253716 TI - [Dynamics of the non-specific reactivity of the organism in time]. AB - Experiments with 10 mongrel rats and dogs showed that individual reactivity of animals to hypobaric hypoxia was same during repeated exposure in two weeks. Investigations of 20 dogs demonstrated relatively permanent types of group reactivity, i.e. hyper-, normal, and hyporeactivity to adrenaline injections every 2 months over a year period. Each type of group reactivity was marked by phase variation in the count of peripheral blood leukocytes with the relation of group mean values unchanged. Analysis of mean values of adaptive levels in 340 subjects with varying individual reactivity over 10 years also pointed to undulation of these values in each group. At the same time, the difference in mean values of this parameter among the groups was kept constant. Hence, the group specific reactivity has a long-range character as determined by testing. PMID- 11253718 TI - [The imitation of the effects of otolith-ocular asymmetry by eccentric rotation]. AB - Today, investigation of the vestibuloocular reactions is the mainstream method of studying the vestibular asymmetry. Analysis of experimental data requires a model of otolith-ocular interaction. The proposed model is based on the literary data concerning measurements of ocular counter-rotation (OCR) and luminous line rotation (LLR) in experiments with eccentric rotation. The method utilizes a number of simplifications and suppositions, the basic of which are linearity of all phases of transformation of mechanic stimulus with the exception of the afferents' transmission function (proportionality of the nervous response to acceleration; the otolith-ocular response is proportional to the nervous response). It was demonstrated that the model qualitatively imitates the behavior of OCR and LLR in response to axifugal acceleration of the utricular otoliths and permits analysis of the role of various parameters of the otolith-ocular interaction. Comparison of calculated and experimental dependence of OCR and LLR on acceleration can help understanding of the otolith asymmetry. PMID- 11253719 TI - [Sub-circadian rhythms as an instrument for the determination of and prediction for the state of the organism]. AB - The paper deals with the analysis of literature concerning the role and hierarchy of circadian rhythms in the structure of rhythmic processes in organism. Substantiation is given to the concept according to which the diagnostic and prognostic value of subcircadian rhythms is predetermined by their wide representation at different levels of the living system organization, high sensitivity to external impacts, and responsiveness to all sorts of shifts in the state of organism. The above considerations are used in the physiology of labor, experimental and clinical medicine. Illustration can be determination of and prediction for the state of patients and subjects exposed to head-down bedrest, g loads and other debilitating factors based on investigations of subcircadian rhythms. The author presents his own data on determination of and prediction for individual tolerance for passive standing test with account of subcircadian rhythms. Special emphasis is placed on the data about subcircadian rhythms of animals during spaceflight. PMID- 11253717 TI - [The role of individual reaction s of thermal and water-electrolyte metabolism to suited immersion]. AB - Role of individual reactions of thermal and water-electrolyte metabolism and their relation to the mechanisms of hormonal control were studied during suited immersion (SI). The investigation consisted of 177 man/exposures from 3 hrs up to 3 days of 46 male volunteers. Parameters of mass, heat and water-electrolyte metabolism, and hormonal control were determined on a background of monitoring consumption of special ratios. Based on the complex evaluation of tolerance of weight-free SI for 1 to 3 days, parameters of thermal and hydroionic homeostasis were found to be the predictor variables. Extreme types of reactions to SI were in the ratio of 1:4 (tolerant/intolerant subjects). An extreme reaction appears to be an individual constitutional characteristic of the body the dimensions of which are equal during repeated immersion. As for the water-electrolyte parameters, extreme levels of SI tolerance were significantly different in the amount and velocity of renal excretion of liquids and liquid losses due to perspiration (897 +/- 253 ml and 2010 +/- 218 ml in SI tolerant and intolerant subjects, serotonin variability in everyday life and serotonin dynamics during SI (significant increase in tolerant subjects and no dynamics in intolerant subjects after 24 hrs in SI). The complex evaluation of the reaction showed nonstationary thermal state and reduction of oral temperature by 1 degree C in SI intolerant subjects after initial 1-1.5 days of exposure. In the course of 3 days, the negative water balance showed an increase; body mass losses were in parallel with the development of motion sickness and hyperventilation, and disturbance in the rhythm of the sympathoadrenal activity. The renin-aldosterone system reacted most actively out of all the hormonal systems and did not adapt to SI within a day and a half. Thermal state of SI tolerant subjects was stationary during 1-1.5-d immersion; oral temperature dropped by 0.2-0.4 degree C, liquid losses and AP deviations were relatively small and blood osmolality remained unchanged on a background of stable adaptive reactions of various components of the neuroendocrine system. Adaptation to SI occurred in the period from one day to a day and a half. PMID- 11253720 TI - [Characteristics of super dwarf wheat metabolism in microgravity]. AB - Metabolism of sLt during Russian-US experiment GREENHOUSE-2 (July 9, 1996-January 17, 1997) within the MIR/NASA space research program and in laboratory Svet experiments in 1995-1996 was studied. Chemical, biochemical and pigment analyses of the flight and laboratory plants were made after the first (dry biomass) and second vegetation (photosynthetically active 41-d old plants). Data on the composition of leaves and stems of ground and flight wheat do not attest any biologically significant shifts in plant metabolism. There were slight changes in accumulation and migration of several macro- and microelements, protein nitrogen and phosphororganic compounds in microgravity. Lowered content of lignin, a critical supportive element for cellular walls was observed only during early stages of vegetation. In the Mir experiment, concentrations of photosynthetically active pigments also decreased a little but the chlorophyills-carotenoids balance was not upset. PMID- 11253721 TI - [Sensitivity in vitro of test-cultures and collection s of microoganisms isolated from surfaces of a pressurized complex to detergents]. AB - Human subjects are a prime source of microbial contamination of the diving gear and the interior of deep-water diving complexes (DWDC). Disinfecting measures cannot ensure infectious safety of deep-water dives. Investigations in vitro were to compare effectiveness of a standard and a novel detergent against the microorganisms isolated from DWDC-250 at SRC-IBMP. It was discovered that 21% of microbial strains were resistant to the standard 3% solution of hydrogen peroxide. Diluted in ratio 1:10, polycept Demos (alkyl dimethyl benzyl ammonium chloride AB 0.5%) demonstrated 100% effectiveness in usual conditions and in hyperbaric experiments with various gaseous mixtures. Biocide Demos should be further tested during simulation of dives in DWDC-250. PMID- 11253722 TI - [Comparative study of ion homeostasis and lipid peroxidation in human serum during prolonged exposure to ozone]. AB - Concentrations of the ions of calcium, sodium, potassium, chloride-anions and end products of lipid peroxidation in blood serum of patients with bronchopulmonary pathologies were investigated on a background of prolonged exposure in ozone. Ion homeostasis was found to alter as products of lipid peroxidation accumulated. Suppositions regarding possible mechanisms of impairment of ion homeostasis were made. PMID- 11253723 TI - [Regulation of vertical posture in patients with children's cerebral paralysis treated with the method of proprioceptive correction]. AB - Maintenance of vertical posture is a complex coordination act dependent on a multi-level system of regulation. Functional state of the visual system may contribute significantly to the implementation of the mechanisms of vertical posture maintenance. Stabilographic examination of healthy teenagers and patients afflicted with the main CCP forms with the method of dynamic proprioceptive correction revealed dependence of posture stability and involvement of the visual analyzer on sex. PMID- 11253724 TI - [Method of integral evaluation of alterations in the body systems due to various factors on the basis of generalized parameters]. AB - The paper presents generalized parameters for quantitative evaluation of shifts in human health and separate body systems due to external factors based on available circumstantial data of prophylactic medical examination of different cohorts of population and professional groups. Test of the generalized logarithmic parameter proved its applicability to the quantitative evaluation of radiation damage to and recovery processes in the blood-forming system following repeated acute exposures. A significant fall in the rate of recovery of the system and increases in the number of fractions and the total absorbed dose were demonstrated. PMID- 11253725 TI - [The effect of the microwave energy on the water, contaminated by vegetative forms of microorganisms]. AB - The paper presents a technology and a device for super-high sterilization, and results of investigation on the effect of microwave energy on water contaminated by vegetative forms of microorganisms. Temperatures bringing death to microorganisms in water flow were determined. In addition to the thermal effect of microwave energy, a non-thermal "specific" effect of super-high frequencies on various microorganisms (staphylococci, E. coli and Pseudomonas aeruginosa) was also studied. PMID- 11253727 TI - [Russian conference on Organism and Environment: Life Support and Protection of Humans in Extreme Conditions with international participation]. PMID- 11253726 TI - [Investigation of the methods of extension of useful life of nanofiltration membranes in washing water regenerating system]. AB - Transport and selective characteristics of Russian nanofiltration membranes OPMN K were investigated in order to find approaches to extend useful life of the membranes and, therefore, of the nanofiltration assembly and the washing water regenerating system at large. Hygienic cleansing agents as washing water ingredients pollute but not irreversibly impair the membranes. Flushing of membranes after regeneration of various model detergents was shown to recover the membrane permeability virtually up to original level. PMID- 11253728 TI - [Methodological approaches to utilization of telemedicine capabilities in teaching medical ecology and space medicine]. PMID- 11253729 TI - [Initial experiences with primiparous women using a new kind of Epi-no labor trainer]. AB - PURPOSE: The effectiveness of a vaginal dilatator (Epi-no) in avoiding episiotomies and improving the fetal outcome was examined. DATA SOURCES AND METHODS: Fifty pregnant women were included in our prospective study and took part in the prepartal birth training program with Epi-no. Matched-pairs were compared for the rate of episiotomy and perineal tears, neonatal APGAR score, average time of training, duration of labour and analgesia during delivery. RESULTS: We found a significant reduction in the rate of episiotomies in the group of women who participated in the birth training program with Epi-no (EG: 49%) compared to women who did not take part in our training program (NEG: 82%). Also the rate of perineal tears was twice as high in the latter (4% vs. 2%). Moreover, children of women of the EG showed better one-minute-APGAR-scores. In addition to this we found a significant reduction in the average duration of the second stage of labour in the EG (29 min) if compared with the NEG (54 min). Women in the EG had a lower rate of PDA (16% vs. 36%) and needed less analgesics than those in the NEG. Women of the EG who delivered without episiotomy had trained on average two days longer than women who had had an episiotomy. CONCLUSION: Birth training with Epi-no decreases the rate of episiotomies in primiparous significantly. PMID- 11253730 TI - [Quality criteria and risks for malpractice in ultrasound prenatal diagnosis. What is allowed--what may be missed?]. AB - BACKGROUND: As with all new diagnostic options in medicine, great hope was placed in the introduction of high-resolution prenatal sonography. Progress tends to carry with it a demanding attitude of higher expectations. Patients and doctors alike may initially overestimate the possibilities of medical advances. QUESTIONS: The question at hand is whether objective criteria can validate a positive influence of prenatal ultrasound on fetal outcome. How should a sonographic routine screening be structured, and what legal aspects need to be taken into consideration? MATERIAL AND METHODS: This is an attempt to survey the heterogeneous pool of internationally published data with regard to these critical questions. Only a rationally devised analysis of possibilities and restrictions of routine prenatal sonography can answer the question of "what--if anything--may be missed?" RESULTS AND CONCLUSIONS: Screening ultrasound matches sonography on indication. Despite controversial data a discussion of different studies leads to a positive conclusion on the benefits of ultrasound monitoring in pregnant women. Ultrasound-screening has an explicit effect on medical and economic issues as well as on litigation. PMID- 11253731 TI - [Emergency cervix cerclage in amniotic sac prolapse--a realistic option for prolonging pregnancy]. AB - BACKGROUND: We tried to evaluate, whether emergency operative closure of the cervix (EOCC) is a realistic option for prolongation of pregnancy in cases with early opening of the cervix and prolapse of the amniotic sac (PAS) into the vagina. PATIENTS AND METHODS: We report on 16 patients with PAS between 15 + 3 and 28 + 1 weeks of gestation and cervix dilatation between 2 and 8 cm. After antibiotic and tocolytic treatment we performed EOCC in 7 cases and EOCC + Cerclage in 9 cases. Pregnancy follow up and fetal outcome were analysed retrospectively. RESULTS: Mean gestational age at delivery was 33 + 1 weeks (9 cases > 32 + 0 weeks, 2 cases between 28 + 0 and 31 + 6 weeks, 3 cases between 25 + 0 and 27 + 6 weeks, 1 case with rupture of membranes during operation and immediate cesarean section at 28 + 1 weeks, 1 miscarriage at 23 + 3 weeks). Time between EOCC and delivery was between 0 and 146 days (mean 56.3 days), 14 fetuses survived healthy. The best results were obtained after EOCC + cerclage. CONCLUSION: If antibiotic and tocolytic treatment was successful in stopping local infection and contractions, EOCC is an acceptable and mostly successful procedure to prolong pregnancy. PMID- 11253732 TI - [Treatment of posthemorrhagic hydrocephalus with intraventricular administration of recombinant plasminogen activator (rt-PA) and dexamethasone--possible prevention of permanent shunt implantation?]. AB - BACKGROUND: In spite of substantial advances in intensive care for extremely preterm babies, including continuous monitoring and minimal handling, prevention of intracerebral hemorrhage is not successful in some cases. Unless treated adequately, a progressive posthemorrhagical hydrocephalus leads to considerable impairment of neurological outcome. Several conservative treatment strategies have been published, but often fail to avoid a permanent ventriculoperitoneal shunt implantation with its possible complications. PATIENTS AND METHODS: Our therapeutical regimen of posthemorrhagical hydrocephalus focussed on resolving the mismatch between production and resorption of intracerebral fluid (ICF) by direct lysis of intracranial hematoma and fast decrease of ICF protein concentration. To achieve this goal, we instilled repeated doses of recombinant human tissue plasminogen activator (rt-PA) and dexamethasone intraventricularly, generally via external ventricle drainage systems used also to relieve the elevated intracranial pressure. RESULTS: Relevant complications were not observed. Conservative management without shunt operation was only possible in one out of the treated patients. CONCLUSIONS: Controlled multicenter studies are needed for statistical assessment of treatment efficiency. PMID- 11253734 TI - Pediatric visceral leishmaniasis in Austria: diagnostic difficulties in a non endemic region. AB - Visceral leishmaniasis is usually fatal if left untreated. In Europe it is mainly caused by Leishmania infantum which is endemic in the whole Mediterranean region. While visceral leishmaniasis classically affects children, adults increasingly suffer infections in regions which are known to be endemic for HIV. Nowadays up to 70% of the patients with visceral leishmaniasis in southern Europe are HIV infected adults. The diagnosis is known to be especially difficult to establish in this group of patients because of a frequently atypical clinical presentation, but even in non-HIV-infected patients visceral leishmaniasis often represents a diagnostic challenge particularly when the patient is living in a non-endemic region. We report on four children with visceral leishmaniasis diagnosed at St. Anna Children's Hospital, Vienna, in the last decade. Diagnostic difficulties arose (1) from inexperience with this rare disease, (2) from a long incubation period (6 to 8 months) and (3) from a travel history apparently unsuspicious for the contraction of what is considered a 'tropical' disease. In one case, specific problems resulted (4) from clinical appearance and laboratory data mimicking hemophagocytic lymphohistiocytosis. Consequently even in regions where leishmaniasis is not endemic, diagnostic efforts should be undertaken to rule out this disease especially in patients with the presumptive diagnosis of hemophagocytic lymphohistiocytosis. PMID- 11253733 TI - [Blood volume changes and oxygenation during labor--a laser spectroscopic analysis]. AB - BACKGROUND: Hemodynamic patterns and the distribution of blood volume are influenced by the onset of labour. This study evaluates patterns of oxygenation and changes of blood volume after the induction of contractions. METHODS: A prototype NIR-laser spectrometer was applied in 28 cases. RESULTS: We found a rise of blood volume and oxygen saturation (HbO2) during the peak of contractions. Cerebral tissue oxygenation was monitored by registration of relative changes of cytochrome-aa3. No significant change of tissue oxygenation was detected during variable volume load. DISCUSSION: This implicates compliance of the redox state during labour. PMID- 11253735 TI - Sepsis-associated purpura fulminans in adults. AB - Sepsis-associated purpura fulminans is defined as septicemia, shock, disseminated intravascular coagulation and circulatory failure leading to multiple organ dysfunction. 40-70% of patients with sepsis-associated purpura fulminans die. Early prognostic factors in adults have not been well delineated yet. Aim of our study was 1) to evaluate currently used scoring systems for meningococcal septicemia in the setting of sepsis-associated purpura fulminans and 2) to assess if other parameters are feasible as early prognostic factors. From 1.1 1994 31.12.1998 twelve patients (female: 7; mean age: 31 (21; 43) years) were studied. Six patients (50%) died within 2 hours and 7 days after admission despite standard intensive treatment. On admission non-survivors had a more pronounced degree of disseminated intravascular coagulation compared to survivors (platelet count 18000 (15000; 45000) G/l vs. 119.000 (111000; 152000) G/l, (p = 0.03); fibrinogen 67 (50; 108) mg/dl vs. 356 (234; 483) mg/dl, (p = 0.02); PTZ 28% (20%; 30%) vs. 44% (35%; 51%), (p = 0.05); aPTT 120 (120; 128) sec vs. 46 (44; 69) sec, (p = 0.001). Severity of lactic acidosis was significantly higher in non survivors than in survivors (pH 7.08 (6.92; 7.21) vs. pH 7.4 (7.25; 7.4), (p = 0.02); lactate 13.5 (11; 15) mval/l vs. 6.0 (4.4; 6) mval/l, (p = 0.02); data presented as median (25-75% interquartile range). In our patients the Glasgow Meningococcal Septicemia Prognostic Score (GMSPS) and the Niklasson-Score failed to distinguish between survivors and non-survivors (GMSPS 7 (6; 11) vs 7.5 (7; 9) out of 15; predicted mortality according to Niklasson-Score 73% vs 88%). There was no difference in the APACHE II Score (22 (18.5, 24) vs 22 (20.25, 26)). The severity of disseminated intravascular coagulation assessed by routine laboratory parameters and the degree of lactic acidosis on admission were the strongest predictors of outcome in patients with sepsis-associated purpura fulminans. Scoring systems developed for patients with meningococcal septicemia are of limited value in the setting of sepsis-associated purpura fulminans. PMID- 11253737 TI - Post-traumatic changes in insulin-like growth factor type 1 and growth hormone in patients with bone fractures and traumatic brain injury. AB - The aim of the study was to determine whether changes in serum levels of growth hormone (GH) and insulin-like growth factor type 1 (IGF-1) are related to the phenomenon of enhanced osteogenesis in patients with bone fracture combined with traumatic brain injury (TBI), which would also suggest their involvement in post traumatic stress and their applicability in the promotion of bone fracture healing. GH values were increased during the initial post-traumatic period in all patients (those with bone fractures or TBI alone or combined injury associated with enhanced osteogenesis), declining to normal values afterwards. However, a further increase in GH was only observed in patients with combined injury overlapping with the time of clinically manifested enhanced osteogenesis. Serum levels of IGF-1 were above normal throughout the study period (14 weeks) in patients with TBI only, but not if TBI was combined with bone fractures followed by enhanced osteogenesis. In these patients IGF-1 values increased gradually during fracture healing, as was also the case in patients with bone fractures alone. Thus, different patterns of post-traumatic changes in both GH and IGF-1 were seen in patients with TBI or bone fractures in comparison to those with combined injury, indicating the involvement of these substances in the post traumatic stress response and in the phenomenon of enhanced osteogenesis in patients with bone fractures and TBI. PMID- 11253736 TI - Fibrinolytic parameters and insulin resistance in young survivors of myocardial infarction with heterozygous familial hypercholesterolemia. AB - A characteristic feature of patients with heterozygous familial hypercholesterolemia (FH) is the premature occurrence of coronary artery disease because of elevated LDL cholesterol levels. Hyperinsulinemia and insulin resistance, important characteristics of the cardiovascular dysmetabolic syndrome (CDS), were found to be associated with coronary artery disease in FH subjects, as in the general population. We investigated whether hypofibrinolysis, as part of CDS, is independently associated with symptomatic coronary artery disease in these high-risk patients. Clinical examination (body mass index, waist circumference, blood pressure) and blood analysis (plasma tissue plasminogen activator (t-PA) antigen, plasminogen activator inhibitor (PAI-1) antigen and activity, fibrinogen, serum lipids and lipoproteins, fasting glucose and insulin) were carried out in 39 male patients with heterozygous FH (aged 46.6 +/- 8.8 years). Insulin resistance was calculated using the homeostasis model assessment (HOMA) mathematical model. Thirteen of the patients had suffered a myocardial infarction (MI) 5 to 8 years ago (aged 47.8 +/- 6.1 years) and 26 were free of coronary artery disease (aged 45.9 +/- 9.9 years). There was no difference in total and LDL cholesterol between the two groups. Patients with previous myocardial infarction had significantly higher levels of insulin, insulin resistance, triglycerides, t-PA antigen, PAI-1 antigen and activity, and significantly lower values of HDL cholesterol. Other widely recognised risk factors for coronary artery disease, such as smoking, systolic and diastolic blood pressure, obesity and age, did not differ significantly between the groups. In the logistic regression model, PAI-1 antigen, as a marker of hypofibrinolysis, emerged as an independent risk factor for the occurrence of myocardial infarction (odds ratio 1.55; p = 0.02). In summary our results suggest that the impairment of fibrinolytic activity resulting from elevated levels of PAI-1 antigen and activity and t-PA antigen is an independent variable in CDS associated with the premature occurrence of myocardial infarction in male patients with FH. PMID- 11253738 TI - Acute effects of nicorandil on glucose tolerance in subjects with borderline fasting blood glucose levels. AB - The acute effect of the anti-ischemic potassium channel opener nicorandil on glucose tolerance and post-challenge insulin levels was investigated in 11 subjects (6 males and 5 females, age 59 +/- 2 years) with borderline fasting blood glucose in a single blinded randomised study. All participants were submitted to two oral glucose tolerance tests in randomised order, once without any premedication and once 30 minutes after oral administration of 20 mg nicorandil. This single dose of nicorandil significantly increased blood glucose levels at 120 minutes (173 +/- 16 vs. 150 +/- 11 mg/dl, p < 0.05 by ANOVA) and 180 minutes (106 +/- 11 vs. 88 +/- 7 mg/dl, p < 0.05 by ANOVA) after ingestion of 75 mg of glucose. Serum insulin levels were not significantly altered. In conclusion we suggest that controlled studies in patients with coronary artery disease should be performed to investigate whether long term treatment with nicorandil increases progression rates from impaired glucose tolerance to type-II diabetes and/or from normal to impaired glucose tolerance with a possibly negative impact on the course of cardiovascular disease in comparison to conventional anti-anginal drugs. PMID- 11253739 TI - Unilateral high-altitude pulmonary edema (HAPE): a case report and discussion of pathophysiology. AB - High-altitude pulmonary edema (HAPE), a potentially life-threatening altitude adaptation disorder, is considered to be caused by an exaggerated increase in pulmonary blood pressure and a non-cardiogenic rise in pulmonary vascular permeability subsequent to alveolar hypoxia. A 40-year-old male mountaineer was affected by an advanced stage of HAPE at high altitude (Monte Rosa plateau, 4000 m). The symptoms abated immediately after the patient descended from the altitude. However, six hours after the symptoms had resolved, radiographic signs of pulmonary edema, confined to the right lung, were seen. This rarely described unilateral radiological pattern of HAPE resolved completely within two days. We suggest that aspiration events of nasal secretion, the right sleeping position at night and an elevated right diaphragm reduced the patient's compensatory hyperventilation capacity of the right lung. The resulting increased alveolar hypoxia in the right lung was responsible for unilateral edema. The pathophysiological mechanism underlying unilateral HAPE is discussed. PMID- 11253740 TI - [Fracture and embolization of a central venous port catheter without prior compression between the clavicle and the 1st rib]. AB - A central venous port catheter inserted infraclavicularily via the subclavian route may be compressed by the clavicle and the adjacent first rib. It's appearance on chest x-ray has been previously described as the 'pinch-off phenomenon' and requires the removal of the catheter due to a significant risk of fracture. We report the case of a catheter fracture without prior evidence of pinch-off. The free catheter fragment was embolized into the right atrium and caused pericardial effusion. Percutaneous removal of the fragment was attempted but failed and thus open heart surgery with a cardiopulmonary bypass was required. The myocardial lesion caused by the catheter was sutured. PMID- 11253741 TI - Emergency angioplasty and stent deployment for acute occlusion of an anomalous single coronary artery (all three coronary arteries from one ostium in the right sinus of Valsalva). AB - A single coronary ostium is traditionally considered to be of little clinical significance. We report a case of a single ostium in the right sinus of Valsalva, giving rise to the right coronary artery, from which the left main coronary artery originated. Sudden death occurred seven days after acute gastrointestinal bleeding and subsequent interruption of aspirin therapy. Acute coronary angiography following successful resuscitation revealed an ascending thrombus in the right coronary artery. The patient underwent a complex percutaneous coronary angioplasty with stent deployment. We conclude that coronary artery disease may lead to severe ischemia with a large area at risk and major complications in patients with coronary anomalies. Patients with acute stent implantation might benefit from platelet aggregation even in cases of recent intestinal bleeding. PMID- 11253742 TI - The Vienna Stroke Registry--objectives and methodology. The Vienna Stroke Study Group. AB - The Vienna Stroke Registry (VSR) comprises nine neurological departments of Vienna and was established in 1998. This article describes the objectives and methodology of the VSR. The goals of the VSR are as follows: (1) to document the quality (structure, process, outcome) of medical services and to construct a database which can be used for future adjustment of medical services in Vienna (early stroke intervention, concept of stroke units, rehabilitation services); (2) to guide educational programs; (3) to adjust or establish algorithms for clinical decision making and to analyze predictors of outcome; (4) to document changes in diagnostic and therapeutic strategies over time; (5) to formulate hypotheses about the etiology, pathophysiology, clinical course, and outcome of stroke. Since October 1998 all patients with the presumed diagnosis of a transitory ischemic attack (TIA) or stroke who had been admitted to one of the participating centers within 72 hours of the onset of symptoms were included in the VSR. All patients are prospectively documented according to standardized detailed protocols. The collected data refer to demographic parameters; medical, particularly, vascular history; vascular risk factors; laboratory and technical investigations (including cerebrovascular, cardiological, and neuroradiological findings); details of pharmacological and non-pharmacological treatment; factors influencing the time from the onset of symptoms to hospital admission and the start of therapy; clinical and etiological classification according to pre specified criteria; neurological and functional impairment at specified time points; structured follow-up investigations at 3, 12, and 24 months after the event. Until September 2000, 2300 patients had been included in the VSR. The target number of 3500 patients will be achieved in the second half of 2001. A baseline description of the VSR population will be given separately. PMID- 11253743 TI - Is endotoxemia a risk factor for atherosclerosis? PMID- 11253744 TI - [Stroke control]. PMID- 11253745 TI - Anemia in critical illness. AB - Anemia is a frequent finding in patients treated in ICUs and results in a high number of red blood cell transfusions. Many patients are already admitted to ICUs with subnormal hemoglobin values. Surgery, frequent phlebotomies and overt bleeding episodes are obvious reasons for continuous blood loss during the ICU stay. However, these causes are usually not sufficient to explain the total blood consumption of critically ill patients, which may amount to several liters. Reduced red cell life span and occult gastrointestinal bleeding are possibly important contributory factors. Irrespective of the cause the erythropoietic response to anemia is severely blunted, as a consequence of an inappropriate increase in erythropoietin production, diminished iron availability and direct inhibitory effects of inflammatory cytokines. The importance of anemia for the course and outcome of critically ill patients and its optimal therapy remain to be defined. Considering red blood cell transfusions recent evidence indicates that a target range of 7-9 g/dl hemoglobin is at least as safe and may even be superior compared to a more liberal transfusion strategy. However, the optimal transfusion trigger in relation to patient comorbidity requires further investigation. Rigorous strategies of blood conservation may help to avoid transfusions. Red blood cell substitutes and recombinant erythropoietin are promising treatment options that are currently under investigation. PMID- 11253746 TI - Recombinant granulocyte colony-stimulating factor (G-CSF) in infectious diseases: still a debate. AB - Granulocyte colony-stimulating factor (G-CSF), a central mediator of the endogenous response to infection and inflammation, is approved for use in the prevention of infection-related complications in patients with nonmyeloid malignancies during antineoplastic therapy associated with high risk of severe neutropenia. Administration of granulocyte colony-stimulating factor results in improvement of host defence paired with anti-inflammatory effects. There is evidence from animal and clinical studies that administration of granulocyte colony-stimulating factor may also be beneficial in non-neutropenic infections. This review focuses mainly on the results of different animal and clinical studies of granulocyte colony stimulating factor used in the treatment of severe infections and sepsis. PMID- 11253747 TI - [Paintings of an intensive care: "Lived or not lived, survived nevertheless at the end"]. PMID- 11253748 TI - Demographic features, clinical characteristics and laboratory findings in children with multiple erythema migrans in Slovenia. AB - OBJECTIVE: To identify demographic, clinical and laboratory characteristics of children with multiple erythema migrans (EM) in Slovenia. METHODS: We prospectively studied patients aged 15 years and less, examined at our department for multiple EM in 1996 and 1997. Demographic and clinical data were collected by means of a questionnaire. In addition, basic haematological and biochemical investigations, serologic testing, and Borrelia cerebrospinal fluid and blood cultures were performed. RESULTS: Ninety-five children (44 girls, 55 boys) aged 1 to 13.5 (median, 4.5) years fulfilled the inclusion criteria. A tick bite was recalled by 23%. The incubation period was 10.5 (range, 1 to 150) days, the duration of skin lesions before the initial examination 4 (range, 1 to 54) days, and the median number of skin lesions, 4.5 (range, 2 to 35). The initial disease was mild in 81% of patients. Local and systemic symptoms were reported by 11% and 30% of children, respectively. Clinical signs accompanying EM lesions were found in 42%. Cerebrospinal pleocytosis (predominantly lymphocytic) was seen in 18% of patients; none of them had frank clinical evidence of central nervous system involvement. Serum IgM and IgG antibodies were detected in 28% and 22% of children, respectively. In 3/79 (4%) patients, Borreliae were isolated from the blood and in 2/83 (2%) from the cerebrospinal fluid. In 2/81 (2%) children, borrelial IgG intrathecal antibody production was demonstrated. CONCLUSIONS: Multiple EM in Slovene children is a mild disease. However, some patients had an associated, usually asymptomatic, infection of the central nervous system. PMID- 11253749 TI - [Cost evaluation by a patient questionnaire: pilot study of a weekly cost diary]. AB - Weekly cost diaries are instruments to measure direct and indirect costs prospectively by using patient data. First we searched MEDLINE for information concerning the use of diaries in health care and their methodological evaluation. Based on a Dutch weekly cost diary we developed an instrument for patients with acute or chronic back pain to be completed following participation in an inpatient rehabilitation measure. Its use was tested in an explorative pilot study. We asked for all costs and resource use due to back pain (all direct medical and non medical costs as well as indirect costs) occurring in a 4-week follow-up period, irrespective of the cost carrier. The total response rate was 58%. Patients spent an average 13 minutes a week for completing the questionnaire, without reporting any major methodological difficulties. Some 30% percent of overall costs were direct costs, the majority being non-recurring costs for assistive devices such as mattresses and mattress frames. Excluding these, monthly direct costs per patient were 270 DM on average. Indirect costs, mainly due to absence from salaried work, amounted to an average 1634 DM per patient, with marked variation. Our study results show that this instrument is basically useful and feasible in this indication. Further studies with larger and representative samples are needed to evaluate data quality. It is suggested that weekly cost diaries can be useful tools in particular in decentralized health care systems to measure costs from the societal perspective. PMID- 11253750 TI - [Community-based rehabilitation for severely ill psychiatric patients?]. AB - A central aim of reformatory efforts, as a consequence of the "Psychiatrieenquete" 1975 (a fundamental report of the situation of psychiatry in Germany), had been dehospitalisation of patients with chronic mental illness and their reintegration into the community. Despite a meanwhile well-developed range of community-based services, patients with severe mental illness only rarely get adequate care by these services. This holds especially true for patients with an unfavourable course of disease such as schizophrenia, severe personality disorder, skid-row alcoholism with multiple problems or for patients with double diagnosis. The reasons are barriers set up by the various services and their underlying concepts as well as structural problems in the health care system. Adapted to the special needs for help of these patients, we present a model for the community-based care of this group, combining elements of community psychiatry, addiction treatment and help for the homeless. PMID- 11253751 TI - [Relationship between illness, rehabilitation and work (ZERA)--an training program for medical-vocational rehabilitation of the mentally ill]. AB - A training programme for rehabilitation of people with mental illness is presented, designed particularly for schizophrenic patients who need occupational rehabilitation. The training goal is to connect aspects of the schizophrenic disorder with vocational issues. It is aimed at supporting the participants in developing a realistic and appropriate vocational perspective in line with their illness related restrictions and current vocational possibilities, seeking to find out the individual's optimal ability to take stress so as to avoid over- or understimulation in vocational respects. The ZERA training has been developed for implementation in different medical and vocational rehabilitation settings for persons with mental illness. An initial control-group study was carried out to evaluate the effectiveness of this group training approach, and preliminary results have revealed changes in the experimental group in accordance with the objectives of the training and encourage further research. PMID- 11253752 TI - [Development of a patient questionnaire for assessment of motivation for rehabilitation(PAREMO)]. AB - Following the assumption that the motivation to participate in a rehabilitation program is a multidimensional construct we surveyed experts to develop a first version of the patient questionnaire of rehabilitation motivation (PAREMO). In this article we describe the results of the pilot study with regard to the test theoretical analysis of the questionnaire. Patients of cardiologic, orthopaedic and psychosomatic rehabilitation clinics were participants of this study. After several subsequent steps of analysis the PAREMO was reduced from an initial 150 items to 46 items. The questionnaire now contains a structure of six factors: 1. need for assistance and psychological burden of suffering, 2. restrictions in everyday life because of physical burden of suffering, 3. reactions of significant others to the illness, 4. readiness to change in terms of preventive behaviour, 5. hopelessness and scepticism, and 6. initiative and knowledge. These factors explain almost 50% of the total variance. Cronbach's Alphas range between 0.71 and 0.91 for the subscales, the corrected item total correlation means range between 0.45 and 0.65. The statistical results as well as the naming of the scales are preliminary to this date, they are being reanalysed in another study. PMID- 11253753 TI - [Psychomotor development of children with psychiatric disorders assessed by Movement-ABC-Checklist and Movement-ABC-Test]. AB - Research was done about the agreement between test results and checklist information concerning psychomotor development. The Movement ABC checklist and the Movement-ABC-test were administered in 34 children from the child and adolescent psychiatry unit of the UZ Gasthuisberg, of Leuven, Belgium. The children were 25 boys and 9 girls aged 4 to 12 years. There was a moderate association between the test results of the children and the judgements of the carepersons, suggesting that use of the checklist instead of testing cannot be deemed a good alternative. PMID- 11253755 TI - [Effects of rehabilitation after hip replacement surgery on postoperative complaints regarding the disease and limitation of function]. AB - BACKGROUND: The study was conducted in co-operation with a German health insurance fund (Gmunder Ersatzkasse, GEK) to identify determinants of outcomes of hip surgery from the patient's perspective. METHODS: In September 1997 all beneficiaries (age 40-75 yrs.) who had been treated in hospital for osteoarthrosis of the hip (ICD-715/820) (n = 1352) were sent a questionnaire on average 5.2 months (T1) postoperatively. The standardized questionnaire contained, among others, items about pre- and postoperative subjective assessment of disease specific symptoms (Lequesne Index), complications, comorbidity (Katz Index), health related quality of life (SF-36) and discharge (home or for inpatient rehabilitation). The response rate at T1 was 67.8%. Patients with hip surgery (n = 390) were sent a second questionnaire 17.2 months (T2) postoperatively. After the two mailings, data from 293 patients were available for analysis. Descriptive and multivariate analyses (GSK model) were performed to reveal determinants of disease specific symptom alleviation. RESULTS: Patients (57.6% male) were 61 years of age on average, and 61.2% reported no comorbidity. 88.4% had undergone total hip replacement. A third of the patients reported at least one complication. 70.6% were discharged for inpatient rehabilitation. Univariately, a substantial (and statistically highly significant) decrease was observed in the Lequesne Index over time: (recalled) preoperative: 14.2 points; postoperative T1: 5.6 pts.; postoperative T2: 4.4 pts. This result is confirmed by multivariate analyses (estimated values: pre = 13.8; T1 = 6.9; T2 = 5.7) although it is modified by an interaction effect between the variables "Lequesne Index" and "Discharge". In patients discharged home, the preoperative Lequesne Index is an estimated 13.3, at T1 = 6.9 and T2 = 6.2. The respective estimated values for patients discharged for inpatient rehabilitation are: preoperative 14.3; T1 = 6.9; T2 = 5.2. CONCLUSIONS: Patients receiving inpatient rehabilitation scored higher on the Lequesne Index (higher burden of disease) before hip surgery. In the short term, their improvements are higher than those of the patient group discharged home (-7.4 pts. versus -6.4 pts.) and continue to be higher in the medium term (-9.1 pts. versus -7.1 pts.). Inpatient rehabilitation after hip surgery leads to better disease specific health outcomes than direct discharge home. PMID- 11253754 TI - [Comparison of Hannover Functional Ability Questionnaire (FFbH) and the SF-36 subscale "Physical Functioning"]. AB - Pretesting of a questionnaire for evaluating day-patient rehabilitation, which included comparison of the "Hannover Functional Ability Questionnaire" (FFbH) and the SF-36 subscale "physical functioning" (SF-36 PF), gave rise to more extensive validity testing of the two instruments. In the framework of an expanded pretest, a questionnaire including FFbH and SF-36 PF was sent out to 520 adult former patients who had undergone inpatient orthopaedic rehabilitation following total hip or knee replacement an average 16 months ago. Return rate was 76%. Checks on plausibility showed no complaints about the Hannover Functional Ability Questionnaire while, on the SF-36 subscale, it was found that 16 of 374 patients (4.3%) had confused positive and negative ratings. Moreover, problems were found concerning the wording of several items (understanding, multidimensionality). In contrast to the FFbH, the SF-36 subscale shows weaknesses in the formulation of its items and particularly its ratings. PMID- 11253756 TI - [Self-help groups as a component of ambulatory community-based rehabilitation after acquired brain damage]. AB - Self-help groups are communities of people with chronic illness or disabilities and their families who join in an effort to cope with their condition and/or psychosocial live problems through mutual help. Their areas of work range from therapeutically focussed support to legal and social counselling and support to the personal help an individual member may need. Self-help groups for the various health conditions or disabilities are considered an important component of ambulatory rehabilitation at community level. Along with social, cultural and leisure activities, health education plays an important role in terms of prevention. For survivors of stroke or brain injury, however, the availability of self-help offers continues to be highly inadequate despite a huge need for assistance, counselling and follow-along services after discharge from rehabilitation in this very patient group. PMID- 11253757 TI - Injury in America: the role of alcohol and other drugs--an EAST position paper prepared by the Injury Control and Violence Prevention Committee. PMID- 11253758 TI - A tracheostomy complication resulting from acquired tracheomalacia: case report. PMID- 11253759 TI - Blunt injury of the intrapericardial great vessels. PMID- 11253760 TI - Renal vein thrombosis after martial arts trauma. PMID- 11253761 TI - Evaluation and management of traumatic posterior urethral disruption with flexible cystourethroscopy. AB - BACKGROUND: We sought to consolidate evaluation and management of traumatic urethral disruption using cystourethroscopic evaluation without retrograde urethrogram or suprapubic cystostomy placement. METHODS: We review our experience with initial flexible cystourethroscopic evaluation of suspected urethral injury from blunt trauma with placement of a Council urethral catheter to provide primary endoscopic realignment of the urethra. RESULTS: Access into the bladder was achieved in 8 of 10 patients. After a mean follow-up of 18 months (range, 9 27 months) in the six living patients, only three have required treatment for urethral stricture--direct vision internal urethrotomy in two, and open perineal urethroplasty in one. Urinary continence has been achieved in five of six patients. CONCLUSION: Primary flexible cystourethroscopy with placement of a urethral catheter streamlines evaluation of traumatic posterior urethral injury. In the presence of partial disruption it provided stricture-free outcomes in three of three surviving patients. PMID- 11253762 TI - Ciliary corona and lentricular halo. PMID- 11253763 TI - Iatrogenic keratectasia. PMID- 11253764 TI - Iatrogenic keratectasia. PMID- 11253765 TI - Iatrogenic keratectasia. PMID- 11253766 TI - Opposite clear corneal incisions. PMID- 11253767 TI - LASIK in pediatric eyes. PMID- 11253768 TI - Potential source of pseudoaccomodation in young pseudophakic patients. PMID- 11253770 TI - [Influence of masticatory muscle function on dental occlusion and maxillofacial development]. PMID- 11253771 TI - [Day surgery and the anesthetic management]. PMID- 11253769 TI - Effects of troglitazone in patients with type 2 diabetes mellitus not adequately controlled by sulfonylureas. PMID- 11253772 TI - Plasmodium falciparum gametocyte periodicity. PMID- 11253773 TI - Serological responses to Cryptosporidium infection. PMID- 11253774 TI - Gill morphology and acid-base regulation in freshwater fishes. AB - This review examines the recent advances in our understanding of the mechanisms of ion transport and acid-base regulation in the freshwater fish gill. The application of a combination of morphological, immunocytochemical and biochemical techniques has yielded considerable insight into the field. An important mechanism for regulation of Cl- uptake/base excretion is by morphological modification of the gill epithelium. During acidosis, the chloride cell associated Cl-/HCO3- exchanger is effectively removed from the apical epithelium because of a covering by adjacent pavement cells; this mechanism reduces base excretion and contributes to the compensation of the acidosis. In addition, acidosis induces changes in both the surface structure and ultrastructure of pavement cells. Evidence is accumulating to support the hypothesis that Na+ uptake/H+ excretion is accomplished by the pavement cell. Further, specific localization of a V-type H+-ATPase on the pavement cell epithelium and an increased expression during acidosis provides support for the model originally proposed, that this exchange is accomplished by an electrochemically coupled H+ ATPase/Na+ channel mechanism. PMID- 11253775 TI - Urea and water permeability in dogfish (Squalus acanthias) gills. AB - We used a perfused gill preparation from dogfish to investigate the origin of low branchial permeability to urea. Urea permeability (14C-urea) was measured simultaneously with diffusional water permeability (3H2O). Permeability coefficients for urea and ammonia in the perfused preparation were almost identical to in vivo values. The permeability coefficient of urea was 0.032 x 10( 6) cm/sec and of 3H2O 6.55 x 10(-6) cm/sec. Adrenalin (1 x 10(-6) M) increased water and ammonia effluxes by a factor of 1.5 and urea efflux by a factor of 3.1. Urea efflux was almost independent of the urea concentration in the perfusion medium. The urea analogue thiourea in the perfusate had no effect on urea efflux, whereas the non-competitive inhibitor of urea transport, phloretin, increased efflux markedly. The basolateral membrane is approximately 14 times more permeable to urea than the apical membrane. We conclude that the dogfish apical membrane is extremely tight to urea, but the low apparent branchial permeability may also relate to the presence of an active urea transporter on the basolateral membrane that returns urea to the blood and hence reduces the apical urea gradient. PMID- 11253776 TI - Branchial gas transfer models. AB - Gas transfer in fish gills is simulated by a simple counter-current model, with ventilation, water-blood transfer and blood flow characterized by conductances. The ventilation and perfusion conductances are products of flow rate and effective solubility. The diffusion conductance of water-blood transfer (diffusing capacity) is considered to depend on diffusion properties of both the water-blood tissue barrier and of interlamellar water. In the gills of the dogfish Scyliorhinus stellaris, more than half of the total resistance to O2 diffusion was located into interlamellar water. Complicating factors like water shunt, blood shunt, ventilation-perfusion maldistribution, pulsatile flow, diffusion in blood and reaction of O2 with hemoglobin may reduce the O2 transfer efficacy predicted by the simple model. In S. stellaris, the effect of such complicating factors appeared to be minor in most conditions, but in other species and/or conditions, more complex models might be required. PMID- 11253777 TI - Effect of water alkalinity on gill CO2 exchange and internal PCO2 in aquatic animals. AB - In addition to metabolic CO2 production and gill ventilatory flow rate, expired water PCO2 is very dependent on water acid-base balance in a complex way. This is particularly true in carbonated waters at low ambient PCO2 and high pH, where CO2 excreted in the gill water may be buffered by carbonate ions, leading to an increased CO2 capacitance coefficient. The higher the carbonate alkalinity (CA) and the lower the inspired PCO2 (i.e., the higher the inspired water pH), the stronger the carbonate buffering and the smaller the increase of PCO2 in the gill water during respiratory CO2 exchanges. As a consequence, as shown by a number of reported data, increasing the CA leads to blood hypocapnia and respiratory alkalosis at constant low, but not at high, inspired PCO2. In the low range of inspired PCO2, internal PCO2 becomes very sensitive to even small changes of water PCO2, which may explain at least in part the large variability of reported blood PCO2 values in gill breathers. Water CA also influences the amplitude of respiratory acid-base disturbances caused by changes of the gill ventilatory flow rate. Carbonate buffering of excreted CO2 and thus dependence of blood PCO2 on water alkalinity requires catalysis of CO2 hydration by carbonic anhydrase, that must be available from the water side of the gill epithelium. PMID- 11253778 TI - Gill blood flow control. AB - The arrangement of the fish gill vasculature is quite complex, and varies between the different fish groups. The use of vascular casting techniques has greatly enhanced our knowledge of the anatomy of the branchial microcirculation, not least through the contributions of Pierre Laurent and co-workers at Strasbourg. At different physiological situations, the contact surface between water and blood (functional surface area) varies to balance oxygen uptake against osmotic water flow ("respiratory-osmoregulatory compromise"). This is controlled by nerves and by blood-borne or locally released substances that affect blood flow patterns in the gill. Histochemical techniques have been used to demonstrate neurotransmitter substances in the branchial innervation. In combination with physioly-osmoregulatory compromise" at different physiological situations. PMID- 11253780 TI - Invited editorial/introduction to nitric oxide and the respiratory musculature: A short history of nitric oxide in skeletal muscle function. PMID- 11253779 TI - Gills of antarctic fish. AB - We review the literature on the way the structure of icefish gills relates the physiology of these haemoglobin-less fishes. Vascular casting confirmed earlier reports that the only special feature of the gills is the large size of the blood vessels, especially the prominent and continuous marginal channels Isolated perfused gill arches were used to study the effects of changes in afferent and efferent pressure on gill resistance and tritiated water influx in Chionobathyscus dewitti. Increasing perfusion rate did not change gill resistance, but there were moderate proportional increases in water influx. Reducing efferent pressure increased gill resistance but did not affect water influx. In both C. dewitti and Cryodraco antarcticus gills perfused at constant flow rate, noradrenaline produced concentration-dependent decreases in gill resistance and, with high concentrations, increases in water influx. Fixation while perfusion continued was used to compare blood space dimensions in control, noradrenaline-treated and unperfused gills. Noradrenaline caused large increases in the thickness of the lamellar blood space and increased lamellar height, despite a greatly reduced afferent pressure. This suggests that modulation of pillar cell active tension might be involved in control of lamellar perfusion. The possible relationship between gill water fluxes and lamellar recruitment is discussed. PMID- 11253782 TI - Structure and function of the axillary organ of the gulf toadfish, Opsanus beta (Goode and Bean). AB - The structure of the axillary organ of a batrachoidid species, the gulf toadfish (Opsanus beta Goode and Bean 1879), has been examined and several simple experiments designed to elucidate its function performed. Electron microscopy (EM) studies revealed cells and structures suggesting secretory and iono regulatory roles (e.g., abundant intracytoplasmic secretory particles, rough endoplasmic reticulum, sparse Golgi bodies, indented epithelial cells with microvilli, numerous endocytotic vesicles, etc.). Our physiological experiments allowed us to reach several conclusions: the organs do not excrete significant quantities of urea relative to other areas of the fish (head and gills), the organs do not secrete a substance that is toxic to a teleost test fish (Gambusia affinis), the secretions do not induce short-term modifications in locomotory activity of other gulf toadfish (e.g., by pheromonal means) and the secretions do not inhibit the growth of several species of microorganisms in culture. The function of the organ and its secretions remains unknown, representing a fertile area for research on structure and function in comparative physiology. PMID- 11253781 TI - Role of nitric oxide on diaphragmatic contractile failure in Escherichia coli endotoxemic rats. AB - Contractile dysfunction of the respiratory muscles plays an important role in the genesis of respiratory failure during sepsis. Nitric oxide (NO), a free radical that is cytotoxic and negatively inotropic in the heart and skeletal muscle, is produced in large amounts during sepsis by a NO synthase inducible (iNOS) by LPS and/or cytokines. The aim of this study was to investigate whether iNOS was induced in the diaphragm of Escherichia coli endotoxemic rats and whether inhibition of iNOS induction or of NOS synthesis attenuated diaphragmatic contractile dysfunction. Rats were inoculated intravenously (IV) with 10 mg/kg of E. coli endotoxin (LPS animals) or saline (C animals). Six hours after LPS inoculation animals showed a significant increase in diaphragmatic NOS activity (L-citrulline production, P < 0.005). Inducible NOS protein was detected by Western-Blot in the diaphragms of LPS animals, while it was absent in C animals. LPS animals had a significant decrease in diaphragmatic force (P < 0.0001) measured in vitro. In LPS animals, inhibition of iNOS induction with dexamethasone (4 mg/kg IV 45 min before LPS) or inhibition of NOS activity with N(G)-methyl-L-arginine (8 mg/kg IV 90 min after LPS) prevented LPS-induced diaphragmatic contractile dysfunction. We conclude that increased NOS activity due to iNOS was involved in the genesis of diaphragmatic dysfunction observed in E. coli endotoxemic rats. PMID- 11253783 TI - Cyclic GMP is a second messenger by which nitric oxide inhibits diaphragm contraction. AB - We have shown that endogenous nitrogen oxides (NOx) modulate excitation contraction coupling in diaphragm. Because cyclic GMP (cGMP) is a second messenger for nitric oxide (NO) inhibition of smooth muscle contraction, we rested the hypothesis that NO acts via cGMP in diaphragm. Fiber bundles from rat diaphragm were studied in vitro. Immunohistochemical analysis using a cGMP specific monoclonal antibody confirmed the presence of cGMP in the subsarcolemmal region, near nitric oxide synthase (NOS). cGMP measured by ELISA in control muscle (0.27 pmol/mg +/- 0.01 SE) was significantly increased by the NO donor S nitroso-N-acetylcysteine 1 mM (0.55+/-0.05; N = 6; P < 0.001). Contractile studies showed that the nitric oxide synthase inhibitor N-nitro-L-arginine (L NNA) 10 mM increased submaximal (40 Hz) tetanic force (P < 0.0001). L-NNA effects were exaggerated by the guanylate cyclase inhibitor LY83583 5-10 microM; force at 40 Hz was increased (P < 0.001). L-NNA effects were partially reversed by 8-bromo cGMP 1 mM (8-Br-GMP; a cell-permeable cGMP analogue; P < 0.0001) or dipyridamole 10 microM (DPM; a phosphodiesterase inhibitor; P < 0.0001). 8-Br-GMP and DPM produced more-complete L-NNA reversal in combination (P < 0.0001). We conclude that cGMP functions as a second messenger by which NO inhibits diaphragm contraction. PMID- 11253784 TI - Effect of nitric oxide synthase inhibitor on diaphragmatic function after resistive loading. AB - We studied the effect of a nitric oxide synthase inhibitor, Nomega-Nitro-L arginine-methyl-ester (L-NAME), on in vitro diphragmatic function both at rest (control) or after inspiratory resistive loading (IRL). Sprague-Dawley rats were anesthetized, instrumented, and then the following experimental groups: (1) controls; (2) L-NAME (100 mg/kg/body weight intravenously alone); (3) IRL alone; and (4) L-NAME + IRL. The IRL protocol consisted of applying a variable resistor to the inspiratory limb of a two-way valve at 70% of maximal airway pressure until apnea. After the experiment, the animals were sacrificed and diaphragmatic strips were obtained for activity of constitutive nitric oxide synthase (cNOS) and measurements of in vitro contractile properties: tetanic (Po) and twitch tensions (Pt). cNOS activity was significantly decreased in the L-NAME and L-NAME + IRL groups (P < or = 0.05) as compared with control and IRL groups. L-NAME alone did not affect Po or Pt. However, in both IRL groups, with and without was a significant decrease in Po and Pt. This reduction was comparable in both groups. In summary, our data showed that L-NAME resulted in a significant decrease cNOS activity, but in vitro contractility was impaired. PMID- 11253785 TI - Activity of nitric oxide synthase in the ventilatory muscle vasculature. AB - We evaluated in the in situ vascularly isolated canine diaphragm the role of nitric oxide (NO) in the regulation of basal vascular resistance and vascular responses to increased muscle activity (active hyperemia), brief occlusions of the phrenic artery (reactive hyperemia), and changes in arterial pressure. The vasculature of the left hemidiaphragm was either pump-perfused at a fixed flow rate or autoperfused with arterial blood from the femoral artery. Endothelial nitric oxide synthase (NOS) activity was inhibited by intraphrenic infusion of L arginine analogues such as N(G)-nitro-L-arginine, N(G)-nitro-L-arginine methyl ester and argininosuccinic acid. Active hyperemia was produced by low (2 Hz) frequency stimulation of the left phrenic nerve. Reactive hyperemia was measured in response to 10, 20, 30, 60, and 120 sec duration occlusions of the left phrenic artery and was quantified in terms of postocclusive blood flow, vascular resistance, hyperemic duration, and hyperemic volume. Infusion of NOS inhibitors into the vasculature of the resting diaphragm increased phrenic vascular resistance significantly and to a similar extent. Reactive hyperemic volume and reactive hyperemic duration were also significantly attenuated after NOS inhibition, however, peak reactive hyperemic dilation was not influenced by NOS inhibition. It was also found that enhanced NO release contribute by about 41% to active dilation elicited by continuous 2 Hz stimulation. In addition, NOS inhibition had no effect on O2 consumption of the resting diaphragm, but significantly attenuated the rise in diaphragmatic O2 consumption during during 2 Hz stimulation. The decline in diaphragmatic O2 consumption was due to reduction in blood flow. These results indicate that NO release plays a significant role in the regulation of diaphragmatic vascular tone and O2 consumption. PMID- 11253786 TI - Nitric oxide effects on force-velocity characteristics of the rat diaphragm. AB - The present experiments tested nitric oxide (NO) effects on shortening velocity and power production in maximally activated rat diaphragm at 37 degrees C. Diaphragm fiber bundles (n = 10/group) were incubated at 37 degrees C in Krebs Ringer solution containing no added drug (control), the NO synthase inhibitor Nomega-nitro-L-arginine (L-NNA; 10 mM), the NO donor sodium nitroprusside (SNP; 1 mM), or a combination (L-NNA + SNP). Loaded shortening velocity was measured via the load-clamp technique over a range of afterloads. Unloaded shortening velocity (Vo) was measured in control and L-NNA-treated bundles (n = 12/group) by using the slack test. Force-velocity data fitted to the Hill equation determined a Vmax of 13.7+/-0.4 lengths/s, contradicting the notion that rat diaphragm Vmax declines at temperatures > 35 degrees C. In contrast, L-NNA decreased Vmax (P < 0.05), loaded shortening velocity (P < 0.001), and power production (P < 0.001), but did not change Vo or maximal isometric force. All L-NNA effects were prevented by coincubating fiber bundles with L-NNA + SNP. SNP alone had no effect on any variable. These data indicate that endogenous NO is essential for optimal myofilament function during active shortening. PMID- 11253787 TI - Nitric oxide modulates excitation-contraction coupling in the diaphragm. AB - We investigated the enzymatic source, cellular production, and functional importance of nitric oxide (NO) in rat diaphragm. Neuronal and endothelial isoforms of constituitive nitric oxide synthase (nc-NOS, ec-NOS) were identified by immunostaining. NOS activity measured in diaphragm homogenates averaged 5.1 pmol/min/mg. Passive diaphragm fiber bundles produced NO derivatives (NOx) at the rate of 0.9 pmol/min/mg as measured by the cytochrome c reduction assay; NO production was confirmed by photolysis/ chemiluminescence measurements. Endogenous NO depressed diaphragm contractile function. The force of submaximal contraction was increased by NOS inhibitors, an effect that was stable for up to 60 min and was reversed by NO donors. We conclude that diaphragm muscle fibers express nc-NOS, ec-NOS, or both; passive myocytes produce NOx; and NO or NO derivatives inhibit force production by modulating excitation-contraction coupling. PMID- 11253788 TI - Effects of L-NAME and L-arginine on diaphragm contraction in a septic animal model. AB - The effects of nitric oxide on diaphragm contraction after endotoxin administration were studied in Wistar rats. The animals were divided into seven treatments: a saline-injected group as control, three groups injected with L-NAME (0.01, 0.1, 1 mg/kg) and three groups injected with L-arginine (1, 10, 100 mg/kg). Escherichia coli endotoxin was injected into the peritoneal cavity 15 min later. Twitch kinetics and force-frequency curves were measured 0, 2, and 4 hr after endotoxin injection. In the control group, the force-frequency curves significantly decreased from 0 hr to 4 hr. In the L-NAME group, the force frequency curves at 4 hr showed significant increases in a dose-dependent manner. In the L-arginine group, the force-frequency curve with 100 mg/kg at 4 hr showed a significant increase. There was no consistent change in the contraction time, half relaxation time, or fatiguability. NADPH diaphorase histochemistry performed on diaphragm muscle samples 4 hr after endotoxin injection showed positive in the control and L-arginine group, but was only weakly observed in L-NAME group. These data suggest that nitric oxide contributes to the endotoxin induced diaphragm contractile deterioration. PMID- 11253789 TI - Electrophorus electricus as a model system for the study of membrane excitability. AB - The stunning sensations produced by electric fish, particularly the electric eel, Electrophorus electricus, have fascinated scientists for centuries. Within the last 50 years, however, electric cells of Electrophorus have provided a unique model system that is both specialized and appropriate for the study of excitable cell membrane electrophysiology and biochemistry. Electric tissue generates whole animal electrical discharges by means of membrane potentials that are remarkably similar to those of mammalian neurons, myocytes and secretory cells. Electrocytes express ion channels, ATPases and signal transduction proteins common to these other excitable cells. Action potentials of electrocytes represent the specialized end function of electric tissue whereas other excitable cells use membrane potential changes to trigger sophisticated cellular processes, such as myofilament cross-bridging for contraction, or exocytosis for secretion. Because electric tissue lacks these functions and the proteins associated with them, it provides a highly specialized membrane model system. This review examines the basic mechanisms involved in the generation of the electrical discharge of the electric eel and the membrane proteins involved. The valuable contributions that electric tissue continues to make toward the understanding of excitable cell physiology and biochemistry are summarized, particularly those studies using electrocytes as a model system for the study of the regulation of membrane excitability by second messengers and signal transduction pathways. PMID- 11253790 TI - The disequilibrium pH: a tool for the localization of carbonic anhydrase. AB - The disequilibrium pH is defined as any discrepancy between the measured pH and the pH which would exist if CO2-HCO3-H+ reactions were at equilibrium. Measurement of the disequilibrium pH can be used to assess the status of CO2-HCO3 -H+ reactions and, in combination with carbonic anhydrase (CA) or CA inhibitor treatments, may also be used to localize CA. Renal physiologists have used disequilibrium experiments to determine that HCO3- reabsorption in the kidney tubule occurs via proton secretion, and that CA activity is available to ultrafiltrate CO2-HCO3-H+ reactions in the proximal convoluted tubule, but not the distal tubule. Disequilibrium experiments were also used in investigating the availability of CA to CO2-HCO3--H+ reactions in water at the fish gill; the opposing results obtained in two studies have not yet been resolved. Respiratory physiologists have used the disequilibrium technique in vivo and with saline perfused preparations to assess the availability of CA to plasma CO2-HCO3--H+ reactions following gas exchange. Saline-perfused preparations enable direct localization of CA activity, while in vivo measurements encompass the numerous factors affecting CO2-HCO3--H+ equilibration in a multi-phase solution. Given the many organs in which membrane-bound CA activity has now been identified, the usefulness of the disequilibrium pH technique has increased beyond its original applications in renal and pulmonary physiology. PMID- 11253791 TI - Novelty response in the weakly electric fish Gymnotus carapo: seasonal differences and the participation of the telencephalon in its modulation. AB - The arousal or novelty response (increased frequency of electric organ discharge in the presence of an electric stimulus delivered by a pair of electrodes placed in the water of the experimental chamber) and its habituation were studied in Gymnotus carapo, a weak field electric fish, in different seasons (September and March) and in animals with extensive or restricted telencephalic lesions. The novelty response was reduced but not abolished by repeated stimuli at fixed intervals in normal animals both in September (pre-mating season) and in March (beginning of the dry season). The magnitude of the novelty response did not differ between sham-operated animals and animals with extensive (detelencephalization) or restricted telencephalic lesions (dorsocentral area) within their respective groups (September or March). The novelty response to the first applied stimulus was significantly greater in normal, sham-operated, and derelencephalated animals in September compared to March, although baseline firing rate did not differ. The maximum extent of response reduction occurred after the 18th stimulus in September for all three experimental groups, whereas in the 4 groups tested in March the maximum reduction occurred between the 16th and 72nd stimulus, indicating that in March there is a greater fluctuation in the arousal level. PMID- 11253792 TI - Effect of variation in photoperiod and light intensity on oxygen consumption, lactate concentration and behavior in crayfish Procambarus clarkii and Procambarus digueti. AB - The effects of light intensity and duration on metabolic and behavioral parameters of two species of crayfish, Procambarus clarkii and Procambarus digueti, were studied. Sixty animals of each species were submitted to high irradiance conditions of two different photoperiod lengths, one normal light/dark (LD) 12:12 and one extreme LD 20:4 for 2 weeks. Hemolymph, lactate and oxygen consumption were determined throughout the experimental period. Simultaneously in 18 additional animals of each species, motor activity was individually recorded under the same control and experimental conditions. Both species showed a decrease in oxygen uptake and an increase in hemolymph lactate concentration. The statistical significance of this finding was higher for LD 20:4. This extreme condition evoked a significant decrease of motor activity in P. clarkii and a high mortality rate in P. digueti. P. digueti did not survive after the experiment, whereas P. clarkii survived and adapted to the laboratory conditions. Changes in metabolic and behavioral parameters could indicate different adaptation abilities in these species. PMID- 11253793 TI - Gill protein turnover: costs of adaptation. AB - Measurements of gill protein synthesis, and hence turnover, were greatly facilitated over the last decade by the application of "flooding dose" methodology to non-mammalian species. Numerous studies show that in fish and aquatic invertebrates, gills are among the most active tissues with respect to protein turnover, this being true under a variety of environmental and nutritional conditions. The main components being turned over in fish gills are probably collagen, primarily in the gill arches, and epithelial cell proteins in the filaments, both arches and filaments having similar protein synthesis rates. Intriguingly, differences are apparent between protein synthesis rates of adjacent holobranchs, the first (most anterior) being significantly more active than the second or third, perhaps hinting at functional differences between holobranchs. Experimental estimates of energetic costs for protein synthesis, derived from cycloheximide treatment of isolated perfused gills, give a maximum value of 14 mmol O2/g protein synthesized, which is about double theoretical costs. Environmental stressors, such as heavy metals or acid/aluminum, have variable effects on branchial protein turnover. Limited data suggest that zinc or acid exposure depresses protein synthesis, whereas acid/aluminum increases it quite markedly. Calculations indicate that whereas effects within the gills may be substantial, in terms of whole animal energetics, the costs of branchial adaptation are likely to be small. PMID- 11253794 TI - Relative distribution of blood flow in rats during surface and submerged swimming. AB - The relative distribution of blood flow was investigated in conscious rats with a radiological imaging technique that utilizes technetium-99m ethyl cysteinate dimer (99mTc-ECD). The objective of the study was to determine the effects of locomotory activity on the distribution of blood flow during a dive response. We compared the relative distribution of systemic flow in rats at rest, surface swimming and during periods of voluntarily initiated underwater swimming. The pattern of blood flow differed considerably between the three groups of rats. In resting controls, blood flow was widely distributed throughout the whole body with the thoraco-abdominal region receiving the largest fraction of cardiac output. During surface swimming blood shifted towards the exercising limbs, while during underwater swimming systemic blood flow was largely restricted to the head and thorax. However, the active front and hind limbs were not rendered totally ischemic. This suggests that the demands of exercising skeletal muscle partially over-ride the peripheral vasoconstriction during asphyxic diving in conscious rats. Furthermore, relative blood flow to the head increased during underwater swimming, which supports the view that there is a preferential maintenance of blood flow to the brain. PMID- 11253795 TI - Evaporative water loss, corporal temperature and the distribution of sympatric fiddler crabs (Uca) from south Texas. AB - Desiccation and thermal stress are among the primary factors limiting terrestriality in crustaceans. Water loss was estimated as weight change in five sympatric species of Uca from south Texas for periods up to 7 hr in dry air. Simultaneously, corporal temperature was measured with a thermocouple placed under the carapace. To estimate integumental permeability to water, 115 mm2 portions of dorsal carapace were glued to U-shaped tubes containing a crab Ringer's solution. These were exposed to dry air and water permeability was estimated from weight change. In whole-animal studies, most rapid weight loss occurred in the first 5 min of exposure to dry air as the body temperature fell below ambient (25 degrees C) in all species. The three most terrestrial species exhibited significant survival over more aquatic congeners after prolonged desiccation. The greatest rate of water loss was observed in Uca subcylindrica which lost 22.9+/-3.0% body weight. Uca panacea and Uca spinicarpa lost 14.1+/ 1.6% and 18.5+/-1.8%, respectively. Based on blood osmolarity changes, Uca longisignalis and Uca rapax were more resistant to water loss than Uca subcylindrica under these conditions. Water loss from sections of the dorsal carapace were highest in Uca spinicarpa (10.4 mg/hr/cm2) and Uca longisignalis (8.9 mg/ hr/cm2). Uca subcylindrica and Uca panacea were intermediate (4.5 and 4.2 mg/hr/cm2) while Uca rapax expressed the lowest value (2.9 mg/hr/cm2). These observations support the notion that water loss can effectively lower body temperature in fiddler crabs. However, an inverse relationship between terrestriality and integumental permeability was not evident in these sympatric congeners. Ultimately a balance between physiological and behavioral mechanisms must be achieved for adaptation to the semi-arid habitats in south Texas. PMID- 11253796 TI - Oxygen consumption and carbon dioxide production in male prairie deermice (Peromyscus maniculatus bairdii) in different reproductive conditions and group densities. AB - Natural and laboratory populations of Peromyscus exhibit a profound but reversible reproductive inhibition related to population density. Our earlier studies described the endocrine physiology of inhibited animals which resembles a condition of delayed puberty, but they did not reveal a primary mechanism for the induction and maintenance of the inhibition. These studies indicated that reproductive inhibition could be associated with an overall change in general metabolism. To test this hypothesis, oxygen consumption (VO2) and carbon dioxide production (VCO2) were measured in three groups of Peromyscus maniculatus males that were: 1) reproductively-proven, 2) reproductively-inhibited, or 3) recovered from inhibition. We found that the mean of the 2-hr period with the lowest VO2 (the Resting Metabolic Rate, or RMR) was significantly lower, and the mean Respiratory Exchange Ratio (RER) was significantly higher in reproductively inhibited males compared with reproductively-proven males. In addition, previously inhibited males allowed to recover reproductive function had a significantly higher mean VO2, while the mean RER was not different from reproductively-proven males. Moreover, and contrary to some studies with other species, increasing the ambient carbon dioxide concentration or the caging densities to as high as six animals did not significantly affect oxygen consumption when compared with the corresponding values for individuals. Taken together, these findings indicate that the reproductive inhibition observed in P. maniculatus laboratory populations is causally associated with a significant reduction in general metabolism and that this metabolic reduction which is associated with reproductive-inhibition is not induced by a CO2 signal or induced by absolute density, per se. PMID- 11253797 TI - Homeostatic and circadian control of body temperature in the fat-tailed gerbil. AB - The interplay of homeostasis and circadian rhythmicity in the control of body temperature was studied in the fat-tailed gerbil (Pachyuromys duprasi). In a first study, the body temperature rhythm of 8 gerbils maintained at 24 degrees C under a 14L:10D light-dark cycle was studied by telemetry. Data from 9 other species of small mammals were also obtained for comparison. The gerbils were found to exhibit a robust rhythm of body temperature (the most robust of the 10 species) with a high plateau during the dark phase of the light-dark cycle and a low plateau during the light phase. In a second experiment, 5 gerbils were allowed to select the temperature of their environment by moving along a thermal gradient. The animals consistently selected higher ambient temperatures during the light phase of the light-dark cycle (when their body temperature was at the low plateau). In a third experiment, the metabolic response of 8 gerbils to an acute cold exposure was determined by indirect calorimetry. Greater cold-induced thermogenesis was observed during the light phase. The fact that the animals selected higher ambient temperatures and displayed greater cold-induced thermogenesis when their body temperature was lower contradicts the hypothesis that the body temperature rhythm is caused by a rhythmic oscillation of the thermoregulatory set point. PMID- 11253798 TI - Interactions between ion and gas transfer in freshwater teleost fish. AB - Carbonic anhydrase and proton ATPase are co-distributed, being restricted to the apical regions of the gill epithelium of freshwater teleosts. Carbonic anhydrase supplies protons to the apical proton ATPase. Carbonic anhydrase is absent from the basal regions of the gill epithelium. Plasma flowing through the gills has no available carbonic anhydrase activity and plasma CO2/bicarbonate reactions are uncatalyzed. Thus, bicarbonate dehydration in plasma is negligible, and catalyzed bicarbonate dehydration occurs in erythrocytes in blood flowing through the gills. This results in tight coupling of carbon dioxide excretion to oxygen uptake and the evolution of hemoglobins with large Haldane effects but low buffering capacities, typical of many freshwater teleosts. Tight coupling of carbon dioxide and oxygen transfer in these fish also ensures that the Root shift does not impair oxygen uptake at the gills. Under these conditions, there is a selective advantage for hemoglobins with a Root shift. The presence of a Root shift augments oxygen transfer to the tissues in general and the eye and swimbladder in particular. PMID- 11253799 TI - Effects of exhausting exercise and catecholamines on K+ balance, acid-base status and blood respiratory properties in carp. AB - The potential role of adrenergic mechanisms in the recovery of potassium balance and acid-base status following 5 min of exhausting exercise was studied in carp. The extracellular metabolic H+ load after exercise matched the lactate load, suggesting similar release rates of H+ and lactate from white muscle. Blockage of alpha-adrenoceptors by phentolamine or beta-adrenoceptors by propranolol neither influenced absolute magnitudes nor recovery kinetics of extracellular H+ and lactate loads. The arterial oxygen tension increased following exercise, but blood oxygen transport was not improved via a red cell beta-adrenergic response or modulation of the red cell nucleoside triphosphate content. Exercise induced an increase in extracellular [K+] which was corrected within 30-60 min of recovery. The recovery of K+ balance was not influenced by blockage of adrenergic receptors. Red cell [K+] changed only insignificantly following exercise, whereby a possible function of the red cells as a temporary depository for K+ during the extracellular hyperkalaemia could not be established. The minimal influence of catecholamines on the measured parameters during recovery from exercise was supported by an absence of change in these parameters upon adrenaline injection in resting carp. PMID- 11253800 TI - Adenosine deaminase activity in rat intestine: assay with a microdialysis technique. AB - Using microdialysis, we measured adenosine deaminase activity in rat intestine by detecting inosine, a breakdown product of adenosine. The dialysis probe consisted of a 3 x 0.22 mm dialysis fiber with a 50,000 mol wt cut off. When the probe was perfused at 1 microl/min in vitro, the average relative recovery rate of inosine was 22.1+/-0.9%). The dialysis probe was implanted in the intestinal mucosa and perfused with Tyrode solution containing adenosine at 1 microl/min. The dialysate samples were analyzed for inosine by high-performance liquid chromatography with ultraviolet (HPLC-UV) detection at 260 nm. When adenosine (100-1000 microM) was perfused, the level of inosine increased dose-dependently and was saturatable at about 1 mM adenosine. The ED50 of adenosine was 192.6 microM, with a maximum attainable inosine concentration of 59.7 microM. In the presence of aminoguanidine, a adenosine deaminase inhibitor (10 mM or 10 n mol/microl/min), the elevation of inosine was not observed. The dialysis technique makes it possible to measure adenosine deaminase activity in intestinal mucosa. PMID- 11253801 TI - Experimental inoculations with Ostertagia ostertagi or exposure to artificial illumination alter peripheral cortisol in dairy calves (Bos taurus). AB - A series of experiments were conducted on dairy calves (Bos taurus) to assess, by way of circulating cortisol, the impact of a parasitic infection as a systemic stressor. The first study was designed to assess the effects of chronic stress on dairy calves resulting from a large bolus inoculation of the nematode parasite, Ostertagia ostertagi. Peripheral cortisol concentrations and adrenal cortical competency to adrenocorticotropic hormone (ACTH) challenge were utilized as indicators of chronic stress for 5 weeks. Calves were cleared of nematodes by anthelminthic treatment after the third week of infection. Calves were challenged with ACTH on weeks 0 and 2, and blood samples were obtained at a 12 x 10-min bleeding schedule. Cortisol concentrations were significantly higher (P < 0.05) in the infected calves than in the uninfected calves. The maximal response level to the ACTH challenge was also higher while the calves were infected. Two additional experiments were conducted to investigate the effects of experimental procedures that became evident during Experiment 1. Firstly, calves that had previously been fitted with jugular cannulae were sampled from 3 hr predawn until 5 hr after dawn under red- or white-light incandescent illumination. Calves under red lights had lower initial cortisol concentrations but increased to the concentrations in calves under white lights, indicating a compounding effect of lighting with the procedures of blood-sample acquisition. Secondly, 12 calves were inoculated with 10,000, 100,000, or 200,000 third-stage, infective larvae of O. ostertagi. Blood samples were obtained similarly to the regimen in Experiment 1. Cortisol concentrations were elevated only in the 200,000-dose group during week 3, correlating with the period immediately after emergence of the young adult parasites from the gastric glands. Continuous emergence of these parasites might induce chronic hyperadrenocorticism and the concomitant negative consequences on metabolic and immunological processes. PMID- 11253802 TI - Light-induced extracellular changes of calcium and sodium concentrations in the Planarian ocellus. AB - Ion-selective double-barreled microelectrodes inserted into a planarian ocellus were used to monitor the ocellus potential and the changes in extracellular concentrations of Ca2+ (Ca(o)) and Na+ (Na(o)) caused by a 0.5-sec light flash or sustained (120s) illumination. Ca(o) and Na(o) were slightly decreased following a flash. Sustained illumination caused a biphasic change in Ca(o) (a rapid decrease followed by a slow increase) and a tonic decrease in Na(o). When Na+ in the planarian saline was replaced by Li+ or choline+, the increase in Ca(o) was prevented: sustained illumination induced only a decrease in Ca(o). These results suggest that illumination induces influxes of both Ca+ and Na+ into planarian photoreceptors, and that the Ca2+ influx is rapidly followed by a Na-dependent Ca2+ efflux due to Na-Ca exchange. PMID- 11253803 TI - Protein secretion from rat submandibular acini and granular ducts: effects of exogenous VIP and substance P during parasympathetic nerve stimulation. AB - The influences of exogenous vasoactive intestinal peptide (VIP) and substance P on the release of peroxidase from acini and true tissue kallikrein (rK1) from granular ducts of the rat submandibular gland were studied during continuous parasympathetic stimulation. Parasympathetic nerve impulses caused a moderate flow of saliva (mean +/- SD, 108+/-26 microl/g tissue/min) that had a low protein concentration (174+/-88 microg/ml). The outputs of peroxidase and rK1 were minimal (14.3+/-11.8 pmol DCF/g tissue/min and 6.5+/-3.4 nmol AFC/g tissue/min, respectively). When administered intravenously, VIP had no apparent effect on the overall flow rate, but caused a significant increase in the output of peroxidase; 450% at 1 microg/kg and a further 10-fold increase at 10 microg/kg. In contrast, substance P (1 microg/kg) evoked a marked increase in flow rate (68%), and peroxidase secretion increased only 3-fold. The output of rK1 was unaffected by either VIP or substance P. Our results support the hypothesis that acinar, but not granular duct, protein secretion is evoked by non-adrenergic, non-cholinergic peptides released from parasympathetic nerve terminals. PMID- 11253805 TI - The origins, fate, and ecological significance of free amino compounds released by freshwater pulmonate snails. AB - The mass-specific accumulation rates (MSAR) of both total (TFAC) and individual free amino compounds (FAC) in conditioned media were measured by HPLC, using the orthophthaldialdehyde (OPA) methods, in the following cases: (a) laboratory reared freshwater snails (B. glabrata) with chemosterilized shells; (b) Biomphalaria glabrata with non-chemosterilized shells; (c) B. glabrata faeces; (d) isolated shells of B. glabrata; and (e) 10 other species of freshwater gastropods from the Lewes Brooks, East Sussex, U.K. The MSAR values for B. glabrata show that 95% of the TFAC's (predominantly ethanolamine, phosphoserine, and the amino acids leucine, isoleucine, valine, aspartic acid, and glycine/threonine) originated from the snails themselves as the faeces and shells contributed only 5.0 and 0%, respectively. In contrast, epizootic organisms on the shells of all 10 snail species from the Lewes Brooks released significant amounts of FAC with the two smallest species (Planorbis vortex and Planorbis contortus) having the highest MSAR values. The MSAR for isolated B. glabrata mucus was 42.45 micromol x g(-1)h(-1). As 500 mg snails can release 16.67 mg of mucus daily, this could potentially result in the daily loss of 707.5 micromol of FAC. The cost/benefits of mucus secretion and the various anatomical, physiological, biochemical, and ecological mechanisms which allow freshwater snails to recover FAC's lost as a result of a high rate of urine production in their hypotonic environment, are discussed. PMID- 11253804 TI - In situ potentiation of the glutathione-binding protein for the tentacle ball formation by a protease and efficient ingestion of prey in hydra. AB - Within minutes, brief treatment with trypsin potentiated tentacle ball formation in Hydra japonica, a new behavioral response to reduced glutathione. With the potentiation of this behavioral response, new glutathione-binding proteins were immediately detected after the trypsin treatment of live Hydra, indicating that trypsin activated the glutathione-binding protein in situ. Fixed brine shrimp (Artemia francisca) were more efficiently ingested in the presence of trypsin and S-methylglutathione (GSM) than in the presence of GSM alone, suggesting a biological role of this behavioral potentiation by trypsin in the feeding chain of Hydra. Ingestion of live A. francisca was significantly reduced in the presence of soybean trypsin inhibitor, suggesting that a protease, possibly released from the wounded prey, plays a role in the feeding in vivo. As for Hydra swallowing its captured prey, a small hydra head piece was isolated and measured as it crept along a thin nylon line; advancement of the head was the same in the presence of both GSM alone, and in that of GSM and trypsin together. Together, these results indicate that the chemoreceptor potentiated in situ by a trypsin like protease specifically evokes tentacle ball formation resulting in an efficient transfer of prey on the tentacle to the mouth. PMID- 11253806 TI - Active proton and urea transport by amphibian skin. AB - Proton secretion in frog skin is mediated by an electrogenic H+ pump. Pharmacological and immunocytological approaches have identified this pump as belonging to the V-ATPase class. The key role of this V-ATPase in proton secretion (acid-base balance) and as a membrane energizer of other solute transport from very dilute solutions is outlined. It is shown that the frog skin constitutes a model of a V-ATPase-dependent Na+ transport mechanism applicable to other freshwater animals. On the other hand, attempts to implicate the V-ATPase in the active urea transport that develops through the skin of salt-adapted frogs have failed; the nature of the different urea transporters located on apical and basal epithelial cell membranes and those responsible for active urea reabsorption remain to be identified. PMID- 11253808 TI - Desiccation resistance of two semiterrestrial isopods, Ligia exotica and Ligia taiwanensis (Crustacea) in Taiwan. AB - The ability to resist desiccation stress was examined in two semiterrestrial Ligia species, Ligia exotica Roux and L. taiwanensis Lee, in Taiwan, under a certain desiccation condition. L. exotica exhibited the longer survival time, lower weight-specific rates of water loss, and the slightly higher ability of tolerance to water loss, compared to L. taiwanensis. In each species, the animal size displays a positive correlation to the survival time and total ability to resist desiccation, yet this size effects on the weight-specific water loss rate is negative. Neither water content nor maximum tolerance to water loss shows the association with the animal size in both species. The path ways and magnitudes of the interactions between these traits of desiccation resistance are analyzed and diagrammed using a stepwise regression model. In this model, the body sizes of animal can explain the most part of the variations in the survival time. The body size has a direct effect and an indirect effect, through the effect on water loss rate, on the time that the experimental animals can survival under this desiccated condition. These results suggest that L. exotica attains larger size than does L. taiwanensis, a lower transpiration rate and, consequently, a greater ability in desiccation resistance. The performances of these interactions in the desiccated resistance are more advantageous for L. exotica to migrate and colonize in variable land habitats within a certain limit, and as a result that L. exotica shows a broader distribution pattern than did L. taiwanensis in Taiwan. PMID- 11253807 TI - Accumulation of manganese in the haemolymph, nerve and muscle tissue of Nephrops norvegicus (L.) and its effect on neuromuscular performance. AB - Exposure of Norway lobsters, Nephrops norvegicus (L.) for 3 weeks to manganese concentrations, (5 & 10 mg Mn l(-1) (90-180 microM)), led to its accumulation in various body tissues. The highest concentration was in nerve tissue (brain and abdominal ganglia) which had up to 6 times (on wet wt. basis) the manganese concentration of the exposure concentration, whereas the haemolymph accumulated 3 times and the muscle tissue only 0.5 times the exposure concentration. In the haemolymph the manganese was bound mainly to protein, predominantly (80-90%) to the respiratory protein haemocyanin, as the concentration was 14 times higher in the protein fraction than in the supernatant. Manganese did not substitute for copper in the haemocyanin, as the copper concentration remained constant despite the manganese exposure. The possibility that manganese exposure induced neurotoxic effects sufficient to reduce neuromuscular performance was assessed from the kinematics of free tail-flip swimming, and from measures of the forces produced by abdominal movements in tethered animals. No significant reduction in tail flip velocity or flexion force, but a significant reduction in the maximum post-flip extension force was found. No correlation was found between the manganese concentration in a single tissue or different fractions of the haemolymph and the post-flip extension, except for a weak negative correlation with the manganese concentration in the abdominal ganglion. The ecophysiological implications of these results are discussed. PMID- 11253809 TI - Effect of microalgal diets and commercial wheatgerm flours on the lipid profile of Ruditapes decussatus spat. AB - The influence of both the lipid composition of microalgal diets and commercial flours on the lipid classes and fatty acids of Ruditapes decussatus spat was studied. These aspects of the nutritional value of the diets were discussed in relation to the growth of the spat. Four diets were tested; Diet A, composed of 100% of the daily food ration of microalgae; Diet B, composed of 100% of wheatgerm; Diet C, composed of 50% of microalgae and 50% of wheatgerm; and Diet D, composed of 25% of microalgae and 75% of wheatgerm. The microalgal cells present a higher lipid content than that for wheatgerm. Tahitian Isochrysis cells have phospholipids and triacylglycerols as majority lipids, whereas in the wheatgerm particles, the lipids more abundant are triacylglycerols. Fatty acid content was higher in the microalgal cells than in the wheatgerm particles. The n 3 fatty acids were the most abundant acids in the microalgae, whereas the n-6 fatty acids were in the wheatgerm. The n-3 PUFA were not detected in wheatgerm. Phospholipids were the main lipids present in the clam spat, followed by triacylglycerols. Other lipid classes, detected in significantly lower amounts, included free fatty acids, sterols, and sterol ester + waxes. The composition of fatty acids in the spat was influenced by the fatty acid composition of the diet. Highest spat growth rates were observed with those diets that present a higher phospholipid/triacylglycerol relation. A negative correlation in the relation n 6/n-3 vs. growth has also been observed, with better growth rates in diets with a lower ratio. If the fatty acid 20:5n-3 and 22:6n-3 considered "essential" for marine animals were not present in the diet, they were not present in the spat either. Desaturation and elongation capabilities of R. decussatus spat were also discussed. PMID- 11253810 TI - Comparative lectin-histochemistry on the pre-epithelial mucus layer in the distal colon of conventional and germ-free rats. AB - The mucin composition of the rat distal colonic pre-epithelial mucus layer (PML) was studied by lectin histochemistry in conventional (CV), and germ-free (GF) rats to define effects exerted by the gut flora. No peanut agglutinin (PNA) binding was observed in the PML of GF rats, while the PML of their CV counterparts showed a considerable PNA linkage, indicating terminal Gal-beta1,3 GalNAc residues. Soybean agglutinin (SBA) and Helix pomatia agglutinin (HPA) stained the PML mucins in CV and in GF rats, indicating terminal GalNAc moieties. A quantitative difference in the Limax flavus agglutinin (LFA) binding capacity was found between CV and GF rats, indicating terminal sialic acid moieties: the staining intensity of bound LFA/ FiTC was higher in CV rats than in GF rats. No linkage of Datura stramonium agglutinin (DSA) and of wheat germ agglutinin (WGA) was found in the PML of GF rats, indicating the absence of terminal GlcNAc, while in CV rats, a clearly marked border was visible next to the luminal content as a "nipple edge" when stained with DSA or WGA. Canavalia ensiformis agglutinin (ConA), indicative for branched mannose, stained PML mucins and goblet cell mucins of GF rat distal colon. In CV rats, both locations were free of ConA binding sites. These results suggest degrading effects, exerted by the gut flora on the rat colonic pre-epithelial mucus layer. PMID- 11253811 TI - Responses of class R3 retinal ganglion cells of the frog to moving configurational bars: effect of the stimulus velocity. AB - Discrimination of 'prey' (bars elongated in the direction of movement; W- or H orientation) and 'non-prey' (bars perpendicular to the direction of movement; A- or V-orientation) stimuli in freely moving amphibians is velocity-invariant. Whether or not this phenomenon is present in cells belonging to a general decision making neuronal process remains questionable. Present investigations report the effect of the angular velocity of the stimulus on the discrimination function of class R3 (transient ON-OFF) retinal ganglion cells. The main conclusions of this work are the following: (1) irrespective of the angular velocity, class R3 neurons always prefer vertically (A-) to horizontally (W-) oriented stripes as long as the stimulus length remains inferior to the receptive field size; (2) this preference for small A-stimuli is best expressed when stimuli are moved at V = 7.6 degrees/s; (3) a preference reversal is induced by stripes longer than the receptive field via a dual process involving both spatial and temporal mechanisms; (4) this preference reversal is velocity-dependent: the longer the bar, the faster the velocity should be. PMID- 11253812 TI - ATP-sensitive K+ channels in cardiac muscle from cold-acclimated goldfish: characterization and altered response to ATP. AB - ATP-sensitive potassium channels (K(ATP)) play an important, if incompletely defined, role in myocardial function in mammals. With the discovery that K(ATP) channels are also present at high densities in the hearts of vertebrate ectotherms, speculation arises as to their function during periods of cold acclimation and depressed ATP synthesis. We used single-channel and intracellular recording techniques to examine the possibility that channel activity would be altered in cardiac muscle from goldfish (Carassius auratus) acclimated at 7+/-1 degrees C relative to control (21+/-1 degrees C). As previously observed in mammals, K(ATP) channels in isolated ventricular myocytes were inwardly rectified with slope conductances of 63 pS. However, channel mean open-time and overall open-state probability (Po) were significantly increased in cells from the cold acclimated animals. In addition, K(ATP) channels in cells from fish acclimated at 7 degrees were nearly insensitive to the inhibitory effects of 2 mM ATP, whether studied at 7 or at 21 degrees C. Transmembrane action potential duration (APD) in hearts of cold-acclimated fish studied at 21 degrees was significantly shorter than that observed in hearts of warm-acclimated fish at the same temperature; this difference was eliminated by the K(ATP) channel antagonist glibenclamide (5 microM). These data suggest that K(ATP) channels in the hearts of cold-acclimated animals are more active and less sensitive to ATP-inhibition than those in warm acclimated fish, possibly reflecting a functional adaptation to promote tolerance of low temperatures in this species. PMID- 11253813 TI - Anatomical position of heart in snakes with vertical orientation: a new hypothesis. AB - It has been observed that climbing arboreal snakes have hearts closer to the head than nonclimbing terrestrial or aquatic snakes. The closeness to the head is said to minimize the work of the heart in pumping blood to the head. However, there is ample evidence that the gravitational pressure in the arteries going to the head is counterbalanced (neutralized) by the gravitational pressure of the blood in the veins going down to the heart. Hence, the heart does not do extra work so, another explanation must be sought. It is proposed that the position of the heart may be related to the filling pressure of the heart which is influenced by the compliance of the vessels above and below the heart. Some observations suggest that the caudal vessels in climbing snakes are less compliant than that of aquatic snakes. This tends to move the hydrostatic indifferent point closer to the head and provides an adequate filling pressure in climbing snakes in the vertical position. PMID- 11253814 TI - The development of the ventilatory response to cold in very young rats. AB - To assess the range of functional responses of the ventilatory apparatus of developing rats and the degree to which ventilatory function is developed in advance of other functional characteristics, rat pups at five ages (between 4 and 20 days old) were exposed to temperatures of 28, 32 and 36 degrees C while in a flow through metabolic chamber modified to serve as a whole body plethysmograph. Ventilatory frequency, tidal volume and oxygen extraction 'efficiency' (EO2 = VO2/FEO2 x VI) were measured at each age and temperature. Mean breathing frequency at 4 days old was 2.56 breaths per second, decreasing to 1.99 at 20 days old. There was insignificant modification of breathing frequency with temperature. Four day old rat pups at 28 degrees C had mass specific tidal volumes of 0.017 ml/g, 142% of the value at 36 degrees C (0.012 ml/g). Twenty day old pups at 28 degrees C had mass specific tidal volumes of 0.027 ml/g, also 142% of the thermoneutral value (0.019 ml/g at 32 degrees C). At all ages, increases in tidal volumes were similar and increases in tidal volume were the only response to increased metabolic demand. Oxygen extraction 'efficiency' was about half that previously observed in adult rodents. These observations of ventilation during a cold challenge suggest that although structural development is not complete until much later, functional development is sufficient, either at birth or shortly thereafter. PMID- 11253815 TI - Intestinal uptake of lipovitellin from brine shrimp (Artemia franciscana) by larval inland silversides (Menidia beryllina) and striped bass (Morone saxatilis). AB - Intestinal uptake of lipovitellin (LV) from brine shrimp (Artemia franciscana) in larval inland silversides (Menidia beryllina) and striped bass (Morone saxatilis) was described using immunocytochemistry. Polyclonal antisera were raised against two subunits of LV (LV68 and LV190). When tested by immunocytochemistry, anti LV68 showed cross-reactivity with some of the pancreatic cells especially in inland silversides. Therefore anti-LV190 was used to localize immunoreactive LV. Inland silversides at 14 days after hatching were fed Artemia nauplii and then sampled 4, 8, 12 hr after feeding. Similar experiments were carried out by using striped bass at 5 days and 15 days of age. They were sampled at 2, 4, 8, and 12 hr after feeding. Anterior enterocytes showed no evidence of uptake; however, the brush border of the cells of inland silversides reacted with the antiserum. Posterior enterocytes took up the LV and/or, possibly, their immunoreactive breakdown products. The pattern of uptake included accumulation in supranuclear vacuoles and digestion in supranuclear vacuoles, as suggested by the decay of the immunoreactivity over time. Thus, the posterior intestine of these larval fishes is the site of uptake and digestion of LV, an important nutritive component in the food of many larval fishes; this supports earlier findings using non nutritive marker proteins. PMID- 11253816 TI - 5-(N,N-Dimethyl)-amiloride to discriminate the Unidirectional electrolyte transports in rat small intestine and proximal colon in vivo. AB - The effect of dimethyl-amiloride (DMA), a selective Na+/H+ exchange blocker, was studied on electrolyte net fluxes and unidirectional fluxes of Na and Cl at four levels of rat intestine in vivo in basal conditions. DMA was applied intraluminally at concentrations of 10(-4) and 10(-3) M in the model of ligated loops prepared from duodenum, proximal jejunum, distal ileum and ascending colon in fasted Sprague Dawley rats. Two iso-osmotic test solutions were used: (1) hypo ionic: Na+ 80 mM and (2) iso-ionic: Na+ 148 mM, pH 8.2. 22Na was placed in the loop and 36Cl was given by intravenous route at the beginning of the experiment. Na+/H+ was calculated by two different means, one was based on pH variation following amiloride inhibition of Na influx, the other on the calculation of the passive Na transport. The quantitative evaluation shows that Na/H exchange largely contributes to the electroneutral absorption and luminal pH regulation. The exchanger activity decreases from duodenum, jejunum, ileum and colon where it is completed by K/H exchange to assure low colon luminal pH. PMID- 11253817 TI - Sialic acid concentration of brain gangliosides: variation among eight mammalian species. AB - Sialic acid is a vital component of brain gangliosides which play an essential role in the transmission and storage of information in the brain. The concentration of bound sialic acid in gangliosides and free sialic acid in the brain cortex of eight different mammals [human, chimpanzee (Pan troglodytes), rat (Rattus norvegicus), mouse (Mus musculus), rabbit (Oryctolagus cuniculus), sheep (Ovis aries), cow (Bos indicus) and pig (Sus scrofa)] were compared. Total sialic acid concentration (890+/-103 microg/g wet weight tissue, mean+/-SE, n = 6) was 2 4 times higher in the human brain compared with the other species studied (0.001 < p < 0.05). There was no significant difference between human males and females. The rank order of adult brain sialic acid after humans (in microg/g) was rat (493+/-23, n = 12), mouse (445+/-29, n = 16), rabbit (380+/-18, n = 6), sheep (323+/-43, n = 6), cow (304+/-14, n = 6) and pig (252+/-14, n = 6). Apart from the cow vs the sheep, the differences between species were statistically significant (p < 0.05). In the mouse, cow and sheep, total sialic acid concentration increased during maturation by 18-32% (p < 0.05). In a 2-year-old chimpanzee, the sialic acid concentration in the left lobe of the brain cortex was 25% higher than that of right lobe at 6 weeks of age (p < 0.05). Free sialic acid was higher in the human brain cortex (41+/-3 microg/g) than that of the rat and mouse (32+/-3 and 25+/-5 microg/g respectively) and absent from other species. Variation in brain sialic acid concentration among different animals has implications for the evolution of the brain and may affect learning ability in animals. PMID- 11253818 TI - Urea production and transport in teleost fishes. AB - Teleosts appear to have retained the genes for the urea cycle enzymes. A few species express the full complement of enzymes and are ureotelic (e.g., Lake Magadi tilapia) or ammoniotelic (e.g., largemouth bass), whereas most species have low or non-detectable enzyme activities in liver tissue and excrete little urea (e.g., adult rainbow trout). It was surprising, therefore, to find the expression of four urea cycle enzymes during early life stages of rainbow trout. The urea cycle may play a role in ammonia detoxification during a critical time of development. Exposure to alkaline water (pH 9.0-9.5) or NH4Cl (0.2 mmol/l) increased urea excretion by several-fold in trout embryos, free embryos and alevin. Urea transport is either by passive simple diffusion or via carried mediated transport proteins. Molecular studies have revealed that a specialised urea transport protein is present in kidney tissue of elasmobranchs, similar to the facilitated urea transporter found in the mammalian inner medulla of the kidney. PMID- 11253819 TI - Hormone metabolism by the fish gill. AB - The fish gill, like the mammalian lung, is ideally situated to process circulating biomolecules because: 1) the gill is the only organ perfused by the entire cardiac output, 2) the in-series positioning of branchial and systemic circulations permits "conditioning" of blood immediately before systemic perfusion and 3) gill microcirculation is extensive, providing substantial endothelial/pillar cell surface in contact with plasma. In addition, two or three distinct circulatory pathways within the gill may differentially affect plasma substrates, raising the possibility of vasoactive control of hormone titers. Hormones may be activated or inactivated by the gill, the latter involving extraction (uptake) from the plasma, metabolism by enzymes on the endothelial surface without uptake or uptake plus intracellular metabolism. Over 60% of angiotensin I (ANG I) is activated to angiotensin II (ANG II) in a single transit through the gill lamellae by pillar cell angiotensin-converting enzyme, whereas both ANG I and II are inactivated by the non-lamellar filamental vasculature. Gills may accumulate and store (uptake 1) or degrade (uptake 2) catecholamines via intracellular monoamine oxidase and catechol-O-methyl transferase enzymes, and they show substrate preference for norepinephrine over epinephrine. Similar processes may exist for serotonin. Atrial natriuretic peptides are efficiently (60-90%) extracted from plasma in vivo by C-type clearance receptors. Fifty percent of an endothelin-1 bolus is removed in a single transit through the gill circulation, arginine vasotocin extraction is modest and bradykinin is virtually unaffected. Arachidonic acid is completely extracted by the gill, whereas extraction of prostaglandins I2 and E2 is only 13 and 5%, respectively. Intense cytochrome P450 immunofluorescence in the pillar cells suggests that the gill vasculature may be an important site of detoxification and production of biologically active epoxides. Thus, gills appear to be potent and selective effectors of hormonal signals. PMID- 11253820 TI - Physiology and biochemistry of the pseudobranch: an unanswered question? AB - The structure and function of the pseudobranch has long interested scientists, but its overall role has remained a mystery. Previous studies have attributed respiratory, endocrine, osmoregulatory and sensory roles to the pseudobranch, and the present review concentrates on new findings. Perfusion experiments on the pseudobranch of the rainbow trout (Oncorhynchus mykiss) using both erythrocyte suspensions and Ringer solution have shown that this organ is able to generate values for the respiratory quotient (RQ) greater than 1.0. The release of carbon dioxide into the perfusate was found to be largely independent of flow between perfusion rates of 120-190 microl/min and could be inhibited by acetazolamide (10(-5) M), indicating a role for carbonic anhydrase. Noradrenaline (10(-5) M) had no effect on oxygen consumption or carbon dioxide release of the pseudobranch. The rate of carbon dioxide release was also dependent on the pH of the pre-pseudobranch perfusate, carbon dioxide release being reduced at lower perfusate pH values. Based on the glucose balance of the isolated saline-perfused rainbow trout pseudobranch and on the enzyme profiles for the rainbow trout, cod, swordfish and deep-water grenadier pseudobranch, it is suggested that the pentose phosphate shunt might be a source of carbon dioxide, yielding the high RQ values found for this organ. Most evidence now available indicates that the pseudobranch is integrally linked with the choroid rete and the supply of oxygen to the retina of the fish eye. PMID- 11253821 TI - Fish red blood cells: characteristics and physiological role of the membrane ion transporters. AB - Several membrane ion transporters playing a role in gas transport and exchanges, cell volume regulation and intracellular acid-base regulation have been identified in fish red blood cells (RBCs). This short review focuses on Na+/K+ATPase and its role in establishing the ionic gradients across the membrane, on the Cl-/HCO3- exchanger and its key role in respiration and possibly in inducing a chloride conductance, on the Na+/H+ exchanger and the recent advances on its molecular mechanisms of activation and regulation, on the different types of K-Cl cotransports, the different hypotheses and suggested models and their role in cell volume regulation. There is no evidence in the literature for ionic channels in fish RBCs. We present original data obtained with the patch-clamp technique that shows for the first time the existence of a DIDS-sensitive chloride anionic conductance measured in whole cell configuration and the presence of a stretch-activated nonselective cationic channel recorded in cell-attached and excised inside-out configuration. The part played by these ionic conductances is discussed in relation with their possible involvement in volume regulation. PMID- 11253822 TI - Passive and active transport properties of a gill model, the cultured branchial epithelium of the freshwater rainbow trout (Oncorhynchus mykiss). AB - Branchial epithelia of freshwater rainbow trout were cultured on permeable supports, polyethylene terephthalate membranes ("filter inserts"), starting from dispersed gill epithelial cells in primary culture. Leibowitz L-15 media plus foetal bovine serum and glutamine, with an ionic composition similar to trout extracellular fluid, was used. After 6 days of growth on the filter insert with L 15 present on both apical and basolateral surfaces, the cultured preparations exhibited stable transepithelial resistances (generally 1000-5000 ohms cm2) typical of an electrically tight epithelium. Under these symmetrical conditions, transepithelial potential was zero, and unidirectional fluxes of Na+ and Cl- across the epithelium and permeability to the paracellular marker polyethylene glycol-4000 (PEG) were equal in both directions. Na+ and Cl- fluxes were similar to one another and linearly related to conductance (inversely related to resistance) in a manner indicative of fully conductive passive transport. Upon exposure to apical fresh water, transepithelial resistance increased greatly and a basolateral-negative transepithelial potential developed. At the same time, however, PEG permeability and unidirectional effluxes of Na+ and Cl- increased. Thus, total conductance fell, and ionic fluxes and paracellular permeability per unit conductance all increased greatly, consistent with a scenario whereby transcellular conductance decreases but paracellular permeability increases upon dilution of the apical medium. In apical fresh water, there was a net loss of ions from the basolateral to apical surfaces as effluxes greatly exceeded influxes. However, application of the Ussing flux ratio criterion, in two separate series involving different methods for measuring unidirectional fluxes, revealed active influx of Cl- against the electrochemical gradient but passive movement of Na+. The finding is surprising because the cultured epithelium appears to consist entirely of pavement-type cells. PMID- 11253823 TI - Relationships between branchial chloride cells and gas transfer in freshwater fish. AB - The gill lamellar epithelium is composed of two predominant cell types, pavement cells and mitochondria-rich chloride cells. The chloride cells play a vital role in ionic regulation because they are the sites of Ca2+ and Cl- uptake from water. Consequently, lamellar chloride cell proliferation occurs in response to ionoregulatory challenges so as to increase the ion-transporting capacity of the gill. It has been argued that such chloride cell proliferation might increase the thickness of the blood-to-water diffusion barrier and thereby impede gas diffusion. This review focuses on the potential negative consequences of chloride cell proliferation on gas transfer and possible compensatory mechanisms that might minimise the extent of respiratory impairment. Two approaches were used to evoke chloride cell proliferation in rainbow trout, hormone treatment (growth hormone/cortisol) and exposure to soft water. In all cases, chloride cell proliferation was associated with a pronounced thickening of the lamellar diffusion barrier. The thickening of the diffusion barrier was associated with a significant impairment of gas transfer. Subsequent studies revealed that several compensatory physiological responses occurred concurrently with the chloride cell proliferation to alleviate or reduce the detrimental consequences of the thickened diffusion barrier. These included hyperventilation, an increased affinity of haemoglobin-oxygen binding and earlier onset of catecholamine release during acute hypoxia. PMID- 11253824 TI - Transport mechanisms of seawater teleost chloride cells: an inclusive model of a multifunctional cell. AB - This review assembles recent information on seawater-type chloride cells of marine teleost fish and evaluates the secretion of Na+, Cl-, K+, H+ and NH4+ and the absorption of Ca2+. The evidence for the distribution (apical vs basolateral) and the abundance of the various ion pumps, cotransporters, channels and exchangers is assessed and an inclusive model is constructed. Relationships among the transport systems are presented to suggest that many, if not all, of these systems may be operating simultaneously in individual, multifunctional chloride cells. PMID- 11253826 TI - AIDS research. HIV inhibitor blocks virus from cell. PMID- 11253825 TI - Fisheries science. Ear bones reveal homing tendencies. PMID- 11253827 TI - Paleontology. Mammoth hunters put hopes on ice. PMID- 11253828 TI - Paleoanthropology. Oldest human DNA reveals Aussie oddity. PMID- 11253829 TI - Paleoanthropology. Skull study targets Africa-only origins. PMID- 11253830 TI - Exquisite Chinese fossils add new pages to book of life. PMID- 11253831 TI - A peek at China's paleontological bounty. PMID- 11253833 TI - Internal fights, looting hinder work in the field. PMID- 11253832 TI - Research kicks into high gear after a long, uphill struggle. PMID- 11253834 TI - Fruitful collaborations follow a two-way street. PMID- 11253835 TI - Quantitative modeling of category learning in amnesic patients. AB - Category rule learning was examined in two amnesic patients using the perceptual categorization task (e.g., Ashby & Gott, 1988; Filoteo & Maddox, 1999). Traditional accuracy-based analyses as well as quantitative model-based analyses were performed. Unlike accuracy-based analyses, the model-based approach allowed us to examine both categorization rule learning and variability in the trial-by trial application of the participant's categorization rule. The results indicated that the amnesic patients were as accurate as the controls in learning a complex, nonlinear rule over a large number of trials. The model-based analysis indicated that, in general, the amnesic patients learned the categorization rule as well as controls and applied their rule as consistently as controls. Categorization performance on a second day of testing revealed that amnesic patients can retain the categorization rule over a 24-h period. These results suggest that the brain regions damaged in amnesia are not involved in category learning or memory for the category structures. PMID- 11253836 TI - Executive function in fluency and recall measures among children with Tourette syndrome or ADHD. AB - This study assessed two relevant aspects of executive dysfunction in children with either Tourette syndrome (TS) or ADHD. Process variables derived from existing neuropsychological measures were used to clarify the executive function construct. Clustering of responses on measures of verbal fluency, figural fluency, and verbal learning was examined to assess strategic response organization. Rule breaks, intrusions, and repetition errors were recorded to assess inhibition errors. No significant differences were found among the three groups (TS, ADHD, and controls) on tasks of response organization (clustering). In our sample, both the ADHD and the TS groups were largely free from executive function impairment, and their performance on the fluency and list learning tasks was in the average range. There was a significant group difference on one of the disinhibition variables, with both TS and ADHD groups showing significantly more intrusions on verbal list learning trials than controls. When more traditional total score variables were analyzed among the three groups, there were no significant differences; however, analysis of effect size revealed medium-to large effect sizes for Letter Word Fluency total score differences (ADHD vs. controls), and for Semantic Word Fluency total score differences (ADHD vs. TS), with the ADHD group having weaker performance in both comparisons. Results provide some support for the use and analysis of process variables-particularly those related to inhibition and intrusion errors, in addition to the total score variables when assessing executive function deficits in children with ADHD and TS. While group differences may be found, children with uncomplicated TS should not routinely be considered to have significant executive function impairments, and when deficits are found, they may be attributable to other comorbid disorders. PMID- 11253837 TI - Impaired skill learning in children with heavy prenatal alcohol exposure. PMID- 11253838 TI - Auditory working memory in HIV-1 infection. AB - We evaluated auditory working memory in 41 HIV-seropositive (HIV+) and 37 HIV seronegative (HIV-) male drug users, employing a modified version of the Letter Number Span Task developed by Gold and colleagues. We added a control condition to the standard task in order to evaluate more directly the contribution of the processing component to the working memory deficits with the effects of storage demands minimized. HIV+ subjects performed significantly more poorly compared to controls on an index of working memory processing derived from raw scores obtained under the two testing conditions. These findings are consistent with our previous reports that HIV-related working memory deficits are evident across multiple informational domains; further, the deficit appears to involve multiple component functions of working memory. Converging findings from recent working memory studies and from primate and neuroimaging investigations suggest that functional abnormalities of prefrontal cortex should receive greater emphasis in models of neurocognitive aspects of HIV-1 infection, which have typically emphasized "subcortical" deficits. PMID- 11253839 TI - Long-term effects of high-dose zidovudine treatment on neuropsychological performance in mildly symptomatic HIV-positive patients: results of a randomized, double-blind, placebo-controlled investigation. AB - This study examined the treatment outcome of high-dose (1500 mg/day) zidovudine (AZT) on neuropsychological (NP) functioning (Trailmaking Test A & B, WAIS-R Digit Symbol, and Rey Auditory Verbal Learning Test) across a 12-month period in mildly symptomatic HIV-1 seropositive men (n = 46 at entry) enrolled in a randomized, double-blind, placebo-controlled trial (VA Cooperative Studies Program #298). Neither short-term (0-6 months) nor long-term (0-12 months) AZT administration revealed enhancement in NP performance. The results suggest that, although AZT may afford patients prophylactic benefits, protracted high-dose AZT treatment does not improve NP functioning in mildly symptomatic HIV-positive individuals. PMID- 11253840 TI - Longitudinal evaluation of cognitive disorder in Huntington's disease. AB - The study investigated longitudinal change in cognitive function in 87 patients with Huntington's disease (HD), using a range of neuropsychological tests, which tap mental manipulative abilities, memory, and frontal executive skills. Over a 1 year period the largest changes were noted in letter fluency, object recall, and Stroop Test performance, whereas no changes were noted over more than 3 years on the modified Wisconsin Card Sorting Test. Contrary to expectation, greater change was evident over 1 year for tasks with low compared to high cognitive demands. The differential sensitivity of tasks was attributed in part to inherent characteristics of the tests themselves: their capacity to detect minor gradations of change and their vulnerability to practice effects. However, the greater change for relatively automatic, speed-based tasks with low cognitive demands was interpreted as reflecting the evolution of HD, with a greater magnitude of change occurring in basal ganglia than cortical function. One purpose of the study was to identify tasks sensitive to the progression of HD and hence most suitable for the evaluation of therapies. Despite reaching statistical significance by virtue of the large group size, numerical differences in test scores over 1 year were very small, suggesting that the use of such tests to evaluate change in individuals or small groups of subjects would be problematic. The data highlight the slow progression of HD, the limitations of standard cognitive tests in detecting change over short periods, and the need for therapeutic studies that encompass a relatively prolonged time frame. PMID- 11253842 TI - Brain activation on fMRI and verbal memory ability: functional neuroanatomic correlates of CVLT performance. AB - We have recently reported (Saykin et al., 1999b) selective activation of left medial temporal lobe structures during processing of novel compared to familiar words using functional magnetic resonance imaging (fMRI). The current study describes the relationship between a widely used clinical test of verbal learning, the California Verbal Learning Test (CVLT), and the previously reported fMRI activations. Thirteen right-handed healthy adult participants were studied with whole brain echo-planar fMRI while listening to novel and recently learned (familiar) words intermixed pseudorandomly in an event-related design. These participants were also tested with the CVLT. Scores for CVLT Trial 1 (immediate encoding of novel words) and recognition discriminability (recognition of familiar vs. novel words) were correlated with fMRI signal change during processing of novel versus familiar words using a covariance model implemented in SPM96. For the novel words analysis, voxels in the right anterior hippocampus correlated significantly with Trial 1 (r = .76 at the maxima). For the recognition analysis, a significant cluster of voxels was found in the right dorsolateral prefrontal cortex (r = .88 at the maxima). Our prior results of separable left medial temporal activation to novel and familiar words, together with results of the covariance analyses reported here, suggest that in addition to the left medial temporal lobe (MTL) regions that are engaged during novel and familiar word processing, the right hippocampus and right frontal lobe are also involved, particularly in those participants with better memory ability. This positive relationship between fMRI activation and CVLT performance suggests a role for these right hemisphere regions in successful memory processing of verbal material, perhaps reflecting more efficient encoding and retrieval strategies that facilitate memory. PMID- 11253844 TI - Rates of forgetting on three measures of verbal learning using retention intervals ranging from 20 min to 62 days. AB - Previous research has examined age effects in rates of forgetting at short delay intervals of 20-30 min. The present study examined age effects in three verbal memory tasks at longer delay intervals of up to 62 days. Study participants consisted of 371 community-dwelling men and women comprising three age groups 20 39, 40-59, and 60-79 years. Age differences in acquisition and 20-min delayed recall were found on each of the memory tasks (paragraph, word list, and word pairs). However, all age groups showed equivalent rates of forgetting after this short delay interval. When participants were required to retain information for longer delay intervals (i.e., 1-62 days), increasing age was associated with faster rates of forgetting for day 1, but not over longer delay intervals. Age differences in rates of forgetting for longer delay intervals and the facilitating effects of prompted recall are discussed in terms of encoding and storage versus retrieval processes. PMID- 11253843 TI - Mesial temporal, diencephalic, and striatal contributions to deficits in single word reading, word priming, and recognition memory. AB - Fifty-three volunteer participants were studied with the fade-in task (Ostergaard, 1998) to measure naming latency, word priming, and recognition memory performance. and with morphometric magnetic resonance imaging (MRI) techniques to measure volumes of mesial temporal lobe, diencephalic, striatal, and neocortical structures. The relationship between measures of cerebral volume loss and performance deficits was modeled using simultaneous regression analyses in which the behavioral measures were dependent variables. The results suggested that damage in both hippocampal and amygdala/entorhinal areas as well as damage in the diencephalon and the nucleus accumbens all contributed independently to the severity of recognition-memory deficits. Both caudate nucleus damage and hippocampal damage contributed independently to increased naming latency (slowed single-word reading). Finally, only damage in the hippocampus appeared to result in decreased word priming. These results provide further evidence against the assertion that word priming represents a form of memory unaffected by damage to the mesial temporal lobes. PMID- 11253841 TI - Selective deficits in verbal working memory associated with a known genetic etiology: the neuropsychological profile of duchenne muscular dystrophy. AB - Forty-one boys diagnosed with Duchenne muscular dystrophy (DMD) were each compared to an unaffected sibling on a battery of neuropsychological tests. Verbal. visuospatial, attention/memory, abstract thinking, and academic achievement skills were tested. Results indicated the boys with DMD performed similarly to their siblings on the majority of measures, indicating intact verbal, visuospatial, long-term memory, and abstract skills. However, the DMD group did significantly more poorly than their siblings on specific measures of story recall, digit span, and auditory comprehension, as well as in all areas of academic achievement (reading, writing, and math). This profile indicates that verbal working memory skills are selectively impaired in DMD, and that that likely contributes to limited academic achievement. The association between the known impact of the genetic mutation on the development of the central nervous system and boys' cognitive profile is discussed. PMID- 11253845 TI - Risky decision making in Huntington's disease. AB - In the clinical setting, Huntington's disease is associated with problems in judgment and decision making, however, the extent of these problems and their association with clinical characteristics have not been assessed. Recently, a laboratory-based simulated gambling task has been used to quantify similar decision-making deficits in ventromedial frontal lobe damaged participants. We hypothesized that participants with Huntington's disease (HD) would show deficits on this gambling task. For this study, 14 HD participants were asked to make 100 selections from four decks of cards with varied payoffs in order to maximize winnings of play money. They were compared to 22 participants with Parkinson's disease (PD) and 33 healthy controls. After an initial period in which participants had to learn contingencies of the decks, the HD group made fewer advantageous selections than the PD and control groups. In HD, the number of advantageous selections in the gambling task was correlated with measures of memory and conceptualization but not disinhibition. Thus, people with HD may have had difficulties learning or remembering win/loss contingencies of the decks, or they may have failed to consistently take these into account in their card selections. These findings are consistent with current models of frontal subcortical brain circuits and behavior. PMID- 11253846 TI - Circannual changes in the secondary sexual adornments of semifree-ranging male and female mandrills (Mandrillus sphinx). AB - Male mandrills (Mandrillus sphinx) have spectacular secondary sexual adornments. These include red and blue sexual skin on the face, rump, and genitalia; a sternal scent-marking gland; and a "fatted" rump. Mandrills are seasonal breeders, and in other seasonally-breeding primate species members of both sexes may show increased expression of secondary sexual characteristics during the mating season. We examined changes in male secondary sexual adornments and testosterone levels, in relation to seasonal changes in the female reproductive cycle and sexual skin morphology, in two semifree-ranging mandrill groups. Females showed circannual changes in sexual skin tumescence, and periods of tumescence peaked from May-July in a long-established group. However, formation of a second, smaller group, two years previous to commencement of the study, disrupted the seasonal pattern of sexual skin tumescence and births. As the groups occupied adjacent enclosures, it appears that social factors, as well as physical environment, affected the seasonal patterning of reproduction in females. Male mandrills, by contrast, did not exhibit marked circannual changes in secondary sexual traits. Although adult male testicular volume and circulating testosterone levels increased significantly during the mating season, sexual skin coloration and rump "fattedness" showed no consistent changes with season. There was some evidence to suggest that maturing males (ages 5-8 yr) showed increased development of red sexual skin during mating periods, but once males had fully developed secondary sexual adornments, they remained stable throughout the year. The possible reasons for this are discussed in relation to intermale competition and social organization in mandrills. PMID- 11253847 TI - Corticotropin-releasing hormone-binding protein in primates. AB - In humans, placental corticotropin-releasing hormone (CRH) production has been linked to the determination of gestational length, and a late gestational fall in CRH-binding protein (CRH-BP) has been linked to the onset of parturition. Expression of placental CRH mRNA is limited to primates, and only in man has a circulating CRH-BP been described. As the fall in CRH-BP in late gestation has been associated with parturition in humans, we sought to determine whether a CRH BP circulated in the plasma of other primates. It is unclear whether maternal plasma CRH concentrations are elevated in New World monkeys and prosimians. We have therefore performed CRH plasma measurements in the blood of pregnant marmosets, in several species of lemur, and in pregnant and fetal rhesus monkeys as a positive control. Using gel chromatography, CRH-BP was detected in the human, gorilla, chimpanzee, orangutan, gibbon, macaque, squirrel monkey, and marmoset, but was absent in the mandrill, spider monkey, and lemur. CRH was detected in the plasma of pregnant marmosets and rhesus monkeys. CRH was also detected in the fetal rhesus monkey, but at lower concentrations than in maternal plasma. CRH immunoreactivity was not detectable in the plasma of pregnant lemurs or in extracts of lemur placenta. In conclusion, a circulating binding protein for CRH exists in all species of apes but occurs variably among New World and Old World monkeys and is absent in lemurs. The variable occurrence of the CRH-BP does not support a role for this protein in the mechanism of parturition in primates. Maternal CRH is elevated in the pregnant marmoset and rhesus, and may play a role in the pregnancy of New and Old World monkeys. PMID- 11253848 TI - Cotton-top tamarins (Saguinus oedipus) fail to show mirror-guided self exploration. AB - To investigate the problem of inter- and intraspecific differences on the mirror test, we conducted two experiments on cotton-top tamarins. Experiment 1 employed a technique similar to one used recently on chimpanzees, and provided no evidence of mirror-mediated touching of the marked area. In a control condition, involving colored dye applied to one arm, two subjects also failed to show self-directed touching, even though they clearly looked at their newly dyed arm. Under these test conditions, cotton-top tamarins fail to show mirror-guided self-exploration. Experiment 2 examined whether this failure was due to insufficient mirror exposure, as well as other details of the testing conditions. In particular, we replicated the design of a previously successful experiment on mirror-mediated recognition in tamarins [Hauser et al., 1995], providing four new animals with a protracted period (three weeks) of mirror exposure prior to dying their hair. In parallel with results from Experiment 1, we observed no evidence of mirror mediated behavior (recognition) in Experiment 2. PMID- 11253849 TI - Density and population structure of owl monkeys (Aotus azarai) in the Argentinean Chaco. AB - Owl monkeys are small monogamous primates ranging over a wide area extending from Panama to the Chaco region of northern Argentina. The Chaco, an alluvial plain covering over one million km2 of Argentina, Bolivia, Brazil, and Paraguay, consists of a mosaic of grasslands, savannas, xeric thorn forests, and gallery forests. The region shows significant seasonal variation in climate, rainfall, and food availability. The goal of this study was to determine the density, size, and structure of a population of Aotus azarai in the seasonal gallery forests of the eastern Argentinean Chaco. Reported population density, as well as group size and composition are based on data collected from 11 groups contacted on approximately 900 occasions, and observed for over 2,000 hours during a three year period. Group and individual densities were 16 groups/km2 and 64 individuals/km2, respectively. Approximately half of the groups (n = 5) were small groups which had three individuals most of the time and never more than four, whereas the remaining groups were large groups composed of four or five individuals, and sometimes even six or seven individuals. This is the first study of A. azarai based on monitoring of a relatively large number of distinct groups. Our data suggest that owl monkeys in the seasonal subtropical forests of Formosa live at a density as high as those reported for owl monkey populations observed in tropical forests. The data also show that the social groups in the owl monkey population are of comparable size and composition to those characteristic of populations in the tropics. PMID- 11253850 TI - Evaluating the risk of cardiac toxicity. PMID- 11253852 TI - Phenylacetate pharmacokinetics based on iterative two-stage population analysis. AB - STUDY OBJECTIVE: To determine the population pharmacokinetics of phenylacetate using iterative two-stage analysis implemented with ADAPT 11 software. SETTING: United States government research hospital. DESIGN: Retrospective pharmacokinetic analysis. SUBJECTS: Sixty-seven patients with refractory solid tumors. INTERVENTION: Subjects received from 1-10 courses/individual (total 141 courses) of therapy with either twice-daily administration or continuous infusions of phenylacetate. MEASUREMENTS AND MAIN RESULTS: Extensive plasma concentration measurements were performed after the initial dose or start of infusion, with sparse sampling during subsequent courses of therapy. Phenylacetate plasma concentration-time profiles were described by a one-compartment, capacity-limited clearance model with incorporation of parameters to describe extent of induction of clearance and the rate of induction. Median estimates for volume of distribution, maximum rate of drug elimination, Michaelis-Menten constant, and induction factor, and rate of onset of induction of drug clearance were 0.33 (0.26, 0.48) L/kg, 21.8 (16.3, 28.0) mg/kg/hour, 94.6 (48.8, 153.0) mg/L, 1.28 (1.06, 1.66), and 0.0038 (0.0019, 0.0058) hour(-1), respectively. CONCLUSION: The results of this study are similar to previous pharmacokinetic evaluations using the Abbottbase PKS system but suggest that earlier analyses were suboptimal. PMID- 11253851 TI - Effects of adjunctive treatment with combined cytokines in a neutropenic mouse model of multidrug-resistant Enterococcus faecalis septicemia. AB - STUDY OBJECTIVE: To examine whether the antienterococcal efficacy of a regimen of gentamicin plus vancomycin combined with granulocyte colony-stimulating factor (G CSF) is enhanced by concurrent therapy with interferon-gamma (IFN-gamma). SETTING: Hospital laboratory. INTERVENTION: Mice rendered neutropenic by cyclophosphamide were intraperitoneally inoculated with a gentamicin- and vancomycin-resistant Enterococcus faecalis isolate. MEASUREMENTS AND MAIN RESULTS: Infected animals were randomized into treatment groups that received G CSF alone or in combination with various dosages of IFN-gamma. Additional groups of animals received vancomycin; G-CSF, G-CSF plus vancomycin, IFN-gamma, and G CSF; or vancomycin with both cytokines. Addition of IFN-gamma to G-CSF regimen resulted in no significant change (p>0.05) in survival, compared with treatment with G-CSF alone. Also, the antienterococcal efficacy of antibiotic plus G-CSF was not modified by coadministration of IFN-gamma. CONCLUSION: This study suggests that adjunctive application of combined cytokines may not be more beneficial than only G-CSF in combination with an antibiotic to treat multidrug resistant enterococcal infection. PMID- 11253853 TI - The safety and antiviral effect of protease inhibitors in children. AB - STUDY OBJECTIVE: To determine the safety and antiviral effect of protease inhibitors (PIs) over 36 months in pediatric patients infected with the human immunodeficiency virus (HIV). DESIGN: Observational study SETTING: Pediatric immunodeficiency clinic. PATIENTS: Twenty-one children. INTERVENTION: Demographics, dosage regimens, genotype data, viral RNA and CD4+ lymphocyte counts, adverse drug events (ADEs), laboratory tests, and compliance were evaluated over 3 years. Data were analyzed by chi2, repeated measures analysis of variance, and paired t tests. MEASUREMENTS AND MAIN RESULTS: Twenty-one pediatric patients (aged 3 mo-15 yrs) received PIs over the study period. Average daily doses were ritonavir 26 mg/kg in 12 patients, nelfinavir 94 mg/kg in 16, indinavir 49 mg/kg in 5, and saquinavir 43 mg/kg in 4. Five patients developed resistance to an existing PI. Overall compliance was 70%. Baseline HIV-1 RNA plasma concentrations were significantly higher than average follow-up concentrations during 3-36 months in patients taking ritonavir (p<0.001) and nelfinavir (p<0.001). Sample size was insufficient for indinavir or saquinavir. Sixty ADEs occurred, diarrhea being most common. Of patients with ADEs, 55% required increased monitoring and 43% treatment. Ritonavir was associated with the most ADEs (28), followed by nelfinavir (16), indinavir (11), and saquinavir (5). Significant increases between baseline and follow-up cholesterol levels were found with ritonavir (p=0.02) and nelfinavir (p=0.001), and for serum creatinine (p=0.02) and triglycerides (p=0.02) with ritonavir. Follow-up triglycerides were significantly higher than baseline for indinavir (p=0.003). CONCLUSION: Nelfinavir and ritonavir were effective in decreasing HIV-1 viral loads and improving CD4+ lymphocyte counts. Ritonavir was associated with more ADEs than other PIs. Changes in cholesterol, serum creatinine, and triglycerides were noted with some PIs. PMID- 11253854 TI - Evaluation of very low-dose subcutaneous vitamin K during postoperative warfarin therapy. AB - STUDY OBJECTIVE: To determine the effect of very low-dose subcutaneous vitamin K (SCVK) compared with withholding warfarin for above-target international normalized ratio (INR) values after joint surgery. DESIGN: Historical controlled study. SETTING: University hospital. SUBJECTS: One hundred thirty-nine patients beginning warfarin after total joint surgery. INTERVENTION: For a high INR, warfarin was either withheld or SCVK 100, 300, or 400 microg was administered, depending on INR value. MEASUREMENTS AND MAIN RESULTS: The primary outcome was change in INR from the day of intervention (day 1) to the next day (day 2). Adjusting for day 1 INR, the mean day 2 INR was 2.10 (95% confidence interval [CI] 1.86-2.33) after SCVK, compared with 2.73 (95% CI 2.50-2.96) in controls. This corresponded to declines of -0.72 and -0.08, respectively (p=0.001). CONCLUSION: In orthopedic patients starting warfarin therapy, very low-dose SCVK was more effective than withholding warfarin in reducing high INRs. Investigations in other populations and assessment of the effect of low-dose SCVK on postoperative bleeding are indicated. PMID- 11253855 TI - Risk assessment for antimicrobial agent-induced QTc interval prolongation and torsades de pointes. AB - Over the past several years a multitude of new pharmaceutical agents have been released to the market. Several of them were withdrawn altogether or their use severely restricted to certain indications due to unexpected adverse events, including fatalities. Progress in developing new compounds clearly has surpassed our technology, in some cases, to measure and predict certain toxicities. Prolongation of the QT interval, which may lead to potentially life-threatening ventricular arrhythmias such as torsades de pointes, is one example. Regulatory agencies such as the Food and Drug Administration are increasing standards by which drugs are evaluated for cardiac toxicity related to QT interval prolongation. It is imperative that clinicians be knowledgeable of the risk factors for QT prolongation and avoid the use of culpable agents in patients at risk for QT prolongation. PMID- 11253856 TI - Clinical use of new antithrombotic therapies for medical management of acute coronary syndromes. AB - Prevention and management of acute coronary syndromes (ACS) are focal points of interest among health care providers. Acute coronary syndromes is an all encompassing term that refers to unstable angina, non-Q wave myocardial infarction, and Q wave myocardial infarction. These syndromes are usually the result of atherosclerotic plaque rupture leading to thrombus formation in a coronary artery. Heparin and aspirin are traditional antithrombotic treatments. They typically are administered with antiischemic therapies and often with fibrinolytic agents for patients with ST segment elevation associated with acute myocardial infarction. Although aspirin and heparin are important, they have significant limitations that have prompted development of newer antithrombotic approaches. Adenosine diphosphate inhibitors have been evaluated as either alternatives or adjunctive treatment to aspirin. Glycoprotein IIb-IIIa receptor inhibitors, low-molecular-weight heparins, and direct thrombin inhibitors have been studied as concurrent therapy with, or as alternatives to, heparin. PMID- 11253857 TI - Does prophylaxis against atrial fibrillation after cardiac surgery reduce length of stay or hospital costs? AB - Atrial fibrillation (AF) is common after cardiac surgery and may result in stroke, need for permanent pacemaker, hemodynamic instability, and sustained AF. Studies determining the effect of prophylaxis against AF in cardiac surgery on length of stay (LOS) and hospital cost were reviewed. A MEDLINE search from January 1966-November 2000 was conducted using the medical subject heading (MeSH) "atrial fibrillation." The search was limited to clinical trials in the English language. In five of the seven studies reviewed, the frequency of postoperative AF was significantly reduced with prophylactic amiodarone or sotalol. In only one study were LOS and costs significantly reduced. Whereas there is strong evidence that prophylactic drug therapy reduces the frequency of postoperative AF, there is little evidence of an economic advantage. Future studies are warranted that examine costs beyond initial hospitalization; compare prophylaxis administered selectively to high-risk patients with prophylaxis administered universally; and compare prophylaxis with the combination of rate control and anticoagulation with cardioversion if AF persists. PMID- 11253858 TI - Comparison of preoperative skin preparation products. AB - We compared application, drying, and removal times as well as user satisfaction of four preoperative skin preparation products. All products were applied to 25 subjects, allowed to dry, and removed. Operating room personnel who applied the products were asked to complete a user-satisfaction survey. Application and drying times were longest with the povidone iodine paint and scrub product (p<0.05). That product had the highest rating for overall user satisfaction. Cost minimization analysis revealed that although alcohol-containing products had lower overall costs, savings may not outweigh associated safety risks. PMID- 11253859 TI - Periorbital cellulitis secondary to Conidiobolus incongruus. AB - A previously healthy, 18-month-old girl developed edema and erythema around her left eye 1 week after getting sand in that eye. The patient did not respond to oral or intravenous antibiotics. A mass developed around the eye, and biopsy revealed Conidiobolus incongruus. The patient failed to respond to amphotericin B 1 mg/kg, and susceptibility tests indicated multiantifungal resistance. A combination of antifungal therapy, hyperbaric oxygen, and surgery was required for successful treatment. Three months after treatment the child was disease free. There is no definitive therapy for Conidiobolus incongruus infections, although various drugs have been administered with some success. When susceptibility tests determine multidrug resistance, radical resection with antifungal chemotherapy and hyperbaric oxygen may be necessary as well as lifesaving. PMID- 11253861 TI - New milestones achieved in fluoroquinolone safety. PMID- 11253860 TI - A case report of warfarin resistance due to azathioprine and review of the literature. AB - A 32-year-old woman with a history of deep vein thrombosis (DVT), steroid dependent ulcerative colitis, and osteoporosis was prescribed azathioprine for a steroid-sparing effect. Stable, therapeutic international normalized ratios (INRs) were obtained with warfarin 35 mg/week during a 6-week postpartum course to treat an initial DVT. Ten days after completing warfarin therapy, azathioprine was begun. A recurrent DVT occurred 9 days later, and an increase in warfarin dosage to 120 mg/week was necessary to achieve an INR of 2.0-3.0. The patient denied changes in warfarin adherence, dietary vitamin K intake, or medical conditions. The addition of azathioprine was the only change in her regimen. Review of the literature found only limited reports of an interaction between warfarin and azathioprine, with increases in warfarin requirements of 3-4 times. Careful monitoring and caution are recommended when administering these two drugs concomitantly. PMID- 11253862 TI - Gatifloxacin-induced QTc prolongation and ventricular tachycardia. PMID- 11253863 TI - Activated partial thromboplastin time evaluation. PMID- 11253864 TI - Single-isomer beta-agonists. AB - A new generation of bronchodilators is being developed for acute asthma management-single-isomer beta-agonists. These drugs consist only of the active bronchodilatory isomer (eutomer); they do not have the inactive and potentially harmful isomer (distomer) that is present in marketed racemic beta-agonists. Clinical studies comparing the effectiveness of (R)-albuterol (levalbuterol) with racemic albuterol established a strong rationale for using single-isomer beta agonists in place of the racemic mixture: reduced dosages provide equivalent bronchodilatory effects with fewer beta-mediated side effects. Higher dosages achieve superior bronchodilation in episodes of severe asthma and may reduce costs of emergency department treatment. PMID- 11253865 TI - Do antihistamines have a role in asthma therapy? AB - The coexistence of allergic rhinitis with asthma is widely recognized by clinicians. Histamine, a common mediator for both diseases, has a substantial role in the pathophysiology of asthma through its ability to produce smooth muscle contraction and promote vascular permeability. Because antihistamines often are administered to manage allergic rhinitis symptoms, the effects of antihistamines in asthma should be evaluated. The usefulness of first-generation antihistamines is limited by their side-effect profile, namely sedation and cognitive impairment. Second-generation antihistamines have only a modest effect in attenuating bronchospasm induced by histamine, cold air, exercise, and allergen bronchoprovocation, suggesting that second-generation antihistamines do not have a direct role as a single agent for treating asthma. Studies have shown that controlling allergic rhinitis with antihistamines has a small, indirect effect in improving asthma symptoms. Future work should be directed at improving the potency of antihistamines and defining their antiinflammatory activity. PMID- 11253866 TI - Leukotriene receptor antagonists in the treatment of asthma. AB - The role of leukotriene receptor antagonists in the treatment of persistent asthma is in evolution. Pivotal 8-12-week, randomized, controlled trials in both adults and children have shown efficacy, as defined by standard asthma outcomes. Tolerance to therapy did not develop, nor did rebound worsening of asthma symptoms once therapy was withdrawn. In a comparator trial of montelukast versus beclomethasone, the average percentage change from baseline in forced expiratory volume in 1 second was greater with the inhaled corticosteroid preparation; however, improvements in other asthma outcomes were similar. There was considerable heterogeneity of pulmonary response with both treatments, with good and poor responders in both groups. In an open-label, crossover comparison of montelukast versus cromolyn, both parents and children preferred montelukast, thus regimen adherence was greater with montelukast. Additional long-term, randomized, controlled trials will define the effectiveness of leukotriene receptor antagonists compared with established controllers, thus determining the leukotriene receptor antagonists' place in asthma management. PMID- 11253867 TI - Clinical considerations in the use of inhaled corticosteroids for asthma. AB - Inhaled corticosteroids are the most potent and effective therapy for treating asthma. They exert their pharmacologic action through activation of the glucocorticoid receptor, which helps regulate gene transcription. Corticosteroids also directly inhibit several inflammatory mediators involved in the pathophysiology of asthma. In randomized, controlled clinical trials, inhaled corticosteroids, as monotherapy, are superior to other therapies in improving lung function and clinical outcomes in patients with asthma. However, the use of inhaled corticosteroids is limited by concerns of dose-related adverse effects, including growth suppression and decreased bone density. Combination regimens with these agents and other long-term therapies are beneficial in maintaining asthma control while minimizing dose-related toxicities. Several inhaled corticosteroid products are available in the United States. They differ in potency; however, clinical efficacy is similar when equipotent doses are administered. A variety of factors influence product selection and patient response, including the therapeutic ratio, pharmacokinetic properties, and the inhalation delivery device. In addition, adherence to therapy and the patient's skill in administering the inhaled drug contribute to the therapeutic outcome. PMID- 11253868 TI - Drug treatment of airway inflammation in asthma. AB - Several methods are available for assessing drug effects on airway inflammation and the antiinflammatory effects of drugs for asthma. Cromolyn and theophylline are well-established drugs for the treatment of asthma, and each has antiinflammatory properties. Drugs in development include those aimed at inhibiting inflammatory mediators and immunoglobulin E function; clinical studies, however, have been conducted largely in patients with moderate to severe asthma. PMID- 11253869 TI - Phylogenetic analysis of infectious salmon anaemia virus isolates from Norway, Canada and Scotland. AB - The sequences of gene segments 2 and 8 from 10 different isolates of infectious salmon anaemia virus (ISAV) sampled in Norway, Canada and Scotland between 1987 and 1999 were determined and compared. Pairwise comparisons revealed a high degree of homology between the European isolates, with identities of 98 to 100% for both genes examined. The Canadian isolate showed identities of 84 and 87 to 88% with the European isolates for the nucleotide sequence of segments 2 and 8, respectively. Phylogenetic analyses were performed to establish the interrelationship between the European virus isolates. The evolutionary rate based on 4 Norwegian isolates clustered together in the analysis of segment 2 was calculated to be 0.96 x 10(-3) nucleotides site(-1) yr(-1). On the basis of this mutation rate it was estimated that the Norwegian Glesvaer 90 and Canadian Bay of Fundy 97 isolates diverged around 1900, which coincides with transportation of salmonids between Europe and North America starting in the late nineteenth century. PMID- 11253870 TI - Recombinant vaccines against infectious hematopoietic necrosis virus: production by the Caulobacter crescentus S-layer protein secretion system and evaluation in laboratory trials. AB - We report the development of an IHNV vaccine produced by a new protein production system based on the bacterium Caulobacter crescentus. The subunit vaccines that were tested contain a 184 amino acid segment of the IHNV glycoprotein in different fusion arrangements with the C. crescentus S-layer protein. Relative percent survival of 26 to 34% was demonstrated in rainbow trout fry for a vaccine that contained the 184 amino acid segment of the IHNV glycoprotein fused to the C terminal one-quarter of the S-layer protein. Inclusion of the universal mammalian T-cell epitopes developed from the measles fusion protein or the tetanus toxin protein did not increase the effectiveness of the IHNV-G/S-layer recombinant protein. PMID- 11253871 TI - Detection of Yersinia ruckeri in rainbow trout blood by use of the polymerase chain reaction. AB - We evaluated a polymerase chain reaction (PCR) method for detecting Yersinia ruckeri, the bacterial pathogen causing enteric redmouth disease (ERM), in blood of rainbow trout Oncorhynchus mykiss. Identification of the PCR product was confirmed by Southern blot hybridization with a 32P-labeled oligonucleotide probe matching a sequence within the small subunit ribosomal RNA gene of Y. ruckeri. Following a 1 h immersion of rainbow trout in water with 4.5 x 10(6) colony forming units of Y. ruckeri l(-1), the PCR was positive for all blood samples from 1 h (first sample) to 5 d and was negative from 9 to 30 d (last sample). Fish in this experiment did not show signs of disease, probably because they had been vaccinated against Y. ruckeri. To test this method with naturally infected fish, 42 rainbow trout from hatcheries were examined. Four of these fish had clinical signs of ERM and were infected with Y. ruckeri based on bacteriological culture. The PCR method detected Y. ruckeri in blood, intestine, liver, and trunk kidney from the 4 fish with ERM and from 5 additional rainbow trout that were bacteriologically negative for Y. ruckeri. Three of 5 rainbow trout from streams receiving effluent from hatcheries were positive for Y. ruckeri when tested with PCR, although there was no growth of Y. ruckeri on culture plates inoculated with the same samples. Samples were successfully stored for 1 wk in lysis buffer at 25 degrees C. This study demonstrated that a non-lethal blood sample can be used with PCR to detect Y. ruckeri. PMID- 11253872 TI - Identification of fish-parasitic Myxobolus (Myxosporea) species using a combined PCR-RFLP method. AB - Polymerase chain reaction (PCR) with primers specific for the family Myxobolidae was used to amplify a part of the 18S ribosomal RNA gene of Myxobolus species. The length of the amplified fragments was approximately 1600 base pairs. Six Myxobolus species identified on the basis of morphological features were compared using a combined PCR-RFLP method. The cleavage patterns generated by 2 frequent cutter restriction enzymes (HinfI and MspI) were suitable for the differentiation of the examined Myxobolus species. PMID- 11253873 TI - Occurrence of Ichthyophthirius multifiliis within the peritoneal cavities of infected channel catfish Ictalurus punctatus. AB - Ichthyophthirius multifiliis is a ciliated protozoan parasite that infects the skin and gills of freshwater fish. This report describes the unusual finding of I. multifiliis within the peritoneal cavities of experimentally infected channel catfish Ictalurus punctatus. Twenty catfish fingerlings were exposed to I. multifiliis theronts using a standardized protocol. Five infected fish and 2 control fish were killed at various time points after infection and their tissues examined. Formalin-fixed, paraffin embedded sections were processed for light microscopy and immunohistochemical detection of I. multifiliis immobilization antigen. Trophonts were observed in skin and gill sections of all exposed fish. Parasites were associated with epithelial hyperplasia, focal areas of cellular disruption and necrosis. In addition to these usual sites of infection, individual trophonts were unexpectedly found within the peritoneal cavities of 4 fish. Staining for parasite antigen facilitated their detection within abdominal adipose tissue or adjacent to intestines. This discovery is interesting as it suggests I. multifiliis may be found in tissues other than the skin and gills during the course of a normal infection. PMID- 11253874 TI - Histopathology of cultured sea bream Sparus aurata infected with sanguinicolid trematodes. AB - The present study is the first report of a sanguinicolid infection affecting sea bream Sparus aurata cultured in net cages in the NE of Spain. The disease was associated with trickling mortalities during the cold season (1999 and 2000). Examination of gill wet mounts of the affected population revealed that sanguinicolid infection was present in 82.6 and 100% of the fish sampled in 1999 and 2000, respectively. Adult flukes, which were located in the kidney, were tentatively identified as members of the family Sanguinicolidae, subfamily Cardicolinae. Eggs and miracidia were found in the gill vascular structures. The inflammatory response triggered by the parasites was moderate and the lesions caused by either eggs and miracidia in the gills or adult flukes in the kidney were not extremely severe, possibly because of the moderate intensity of the parasitosis. Histological observations of sanguinicolid infected sea bream presented here are compared with those reported in other fish species. The role played on sea bream morbility and mortality by other factors (occurrence of a simultaneous moderate monogenean infection, immunological impairement related to low water temperatures) is discussed. PMID- 11253875 TI - First report of the invasive eel pest Pseudodactylogyrus bini in North America and in wild American eels. AB - We detected 2 species of monogenean gill worms, Pseudodactylogyrus bini (Kikuchi, 1929) Gusev, 1965 and P. anguillae (Yin & Sproston, 1948) Gusev, 1965 (Monopisthocotylea: Pseudodactylogyridae), on American eel Anguilla rostrata in 2 rivers in South Carolina, USA. One of these, P. anguillae, was reported 5 yr ago from Nova Scotia; as well as in South Carolina, we also discovered it in 2 localities in Chesapeake Bay. Differences in the morphologies of specimens of either species of worm from North America and northeastern Asia were negligible. Similarly, the level of variation in sequences in the ITS2 (internal transcribed spacers) region of ribosomal RNA was minor, and not consistent with geographical origin. These data indicate that these monogeneans invaded North America only recently, possibly in parallel with the nematode Anguillicola crassus (which is known to have been introduced with commercial imports of foreign eels). We map the current global occurrence of these monogeneans, and conclude that their dispersal from northeastern Asia was largely as a result of the eel trade, and has probably been secondarily augmented by longshore migration of infected eels, and possibly also by transport in ballast waters. With present technology, all eel stocks must still be collected from the wild; unless shipments are disinfected at quarantine, these and other eel pathogens (such as A. crassus) are likely to continue to colonise other regions of the world. PMID- 11253877 TI - Complete nucleotide sequence of the S10 genome segment of grass carp reovirus (GCRV). AB - Hemorrhagic disease, caused by the grass carp reovirus (GCRV), is one of the major diseases of grass carp in China. Little is known about the structure and function of the gene segments of this reovirus. The S10 genome segment of GCRV was cloned and the complete nucleotide sequence is reported here. The S10 is 909 nucleotides long and contains a large open reading frame (ORF) encoding a protein of 276 amino acids with a deduced molecular weight of approximately 29.7 kDa. Comparisons of the deduced amino acid sequence of GCRV S10 with those of other reoviruses revealed no significant homologies. However, GCRV S10 shared a putative zinc-finger sequence and a similar distribution of hydrophilic motifs with the outer capsid proteins encoded by Coho salmon aquareovirus (SCSV) S10, striped bass reovirus (SBRV) S10, and mammalian reovirus (MRV) S4. It was predicted that this segment gene encodes an outer capsid protein. PMID- 11253876 TI - Mass mortality of the Japanese pearl oyster Pinctada fucata martensii in relation to water temperature, chlorophyll a and phytoplankton composition. AB - Mass mortalities of the Japanese pearl oyster Pinctada fucata martensii have widely occurred in western Japan since 1994. The causes of these mass mortalities are at present not thoroughly understood. In this study, we investigated oyster survival in relation to some environmental factors such as water temperature, concentration of chlorophyll a and density or composition of phytoplankton. The examined mass mortality occurred from September to December 1998, and the color on the adductor muscle of the oysters was red-brown, suggesting an infectious disease. Oysters that became moribund during the experiment lost weight, while the weight of unaffected oysters increased. The cell density of Nitzschia spp., an inedible algae for the oyster, in Uchiumi Bay increased before and during the mass mortality event. From the results of our study, we hypothesize that P. fucata martensii was weakened by starvation because of the dominance of inedible food and then contracted an infectious disease that resulted in mortality. PMID- 11253878 TI - Pathology associated with an aquareovirus in captive juvenile Atlantic halibut Hippoglossus hippoglossus and an experimental treatment strategy for a concurrent bacterial infection. AB - A large-scale mortality of larval and juvenile halibut Hippoglossus hippoglossus occurred at a semi-commercial halibut farm in Atlantic Canada. Investigation of the cause revealed aquareovirus particles in necrotic liver tissue of affected fish. Cytopathic effect on CHSE-214 cell lines occurred from all fish cultured for viruses, and the viral morphology of the particles in culture was consistent with that observed in necrotic host tissue. The virus was placed in the family of Reoviridae, genus Aquareovirus based on morphology and RT-PCR results. Multifocal hepatocellular necrosis was a consistent finding in all fish as well as acute necrosis of proximal renal tubules. Concurrent bacterial infections were present in some specimens. Fish experimentally treated with oxytetracycline or a combination of oxytetracycline and chloramine-T had a significantly lower mortality rate than untreated fish. Fish treated with chloramine-T alone had a significantly elevated mortality rate compared to controls. Despite supportive medical therapy, mortality levels in treated and untreated groups remained elevated, supporting the hypothesis that the primary pathogen was of viral origin. This is the first report of elevated mortalities in Atlantic halibut associated with an aquareovirus. PMID- 11253880 TI - Urethral diverticulum in females: 25 years experience at Ramathibodi Hospital. AB - We retrospectively reviewed the urethral diverticulum in females from 1972 to 1997. Sixty seven patients were found in this study. Nine per cent were nulliparous and the rest were multiparous with the mean of 2.2 births (range 1 6). Voiding cystourethrography and intravenous pyelography were the main diagnostic investigations (92.4%). Stones in the diverticulum were found in 4.4 per cent. The treatment included marsupialization for the diverticulum at distal urethra in 14 per cent and diverticulectomy for the diverticulum at middle and proximal urethra in 86 per cent. The complications included 1.4 per cent of stress incontinence and 4.4 per cent of recurrent infection. PMID- 11253879 TI - Survival of the North American strain of viral hemorrhagic septicemia virus (VHSV) in filtered seawater and seawater containing ovarian fluid, crude oil and serum-enriched culture medium. AB - The North American strain of viral hemorrhagic septicemia virus (NA-VHSV) could be recovered for up to 40 h in natural filtered seawater (27 ppt) with a 50% loss of infectivity after approximately 10 h at 15 degrees C. Addition of 10 ppb North Slope crude oil to the seawater had no effect on virus survival. However, when various concentrations of teleost ovarian fluid were added to seawater, virus could be recovered after 72 h at 0.01% ovarian fluid and after 96 h at 1.0%. When cell culture medium supplemented with 10% fetal bovine serum was added to the seawater, 100% of the virus could be recovered for the first 15 d and 60% of the virus remained after 36 d. These findings quantify NA-VHSV infectivity in natural seawater and demonstrate that ovarian fluid, which occurs naturally during spawning events, significantly prolongs the survival and infectivity of the virus. The extended stabilization of virus in culture medium supplemented with serum allows for low titer field samples to be collected and transported in an unfrozen state without significant loss of virus titer. PMID- 11253882 TI - Consequences of hip fracture among Thai women aged 50 years and over: a prospective study. AB - OBJECTIVES: To compare post-discharge outcomes of hip-fractured Thai women aged 50 and over with age and sex-matched controls. SUBJECTS AND METHOD: From 1995 to 1997, 60 Thai women aged 50 years and over with hip fracture who had been admitted to the King Chulalongkorn Memorial Hospital and their age and sex matched controls (n = 60) were recruited in a case-control study. These 120 patients were followed for at least 1 year after discharge from the hospital by telephone and/or mailed questionnaire to obtain information about outcomes including death, dependency status and new fracture. Relatives of missing subjects were contacted and interviewed about the outcome status of the patients. RESULTS: The mean age (SD) of those with and without hip fracture was 71.7 (7.6) and 71.2 (8) years, respectively. Of these 120 subjects, 3 cases and 3 controls could not be contacted. The longest follow-up period was 32 months. Means periods (SD) of follow-up among cases and controls were 18.8 (6.7) and 18.1 (6.6) months, respectively. Eleven cases and 5 controls died during the follow-up period. Seven cases and 3 controls died within 1 year after hospitalisation. The survival rate of the cases clearly separated from that of the controls after 1 year. There was a statistical significance of survival between the cases and controls (p < 0.05). The mean (SD) BAI and CAI scores one year after discharge of hip fractured subjects (n = 50) were 17.3 (3.4) and 5.5 (2.3), respectively. The mean (SD) BAI and CAI scores one year after discharge of the control subjects (n = 54) were 16.9 (5) and 5.3 (2.5), respectively. There was no statistically significant difference between dependency status among the two groups. Three (5.2%) cases and one (1.8%) control had new fractures during the follow-up period (no statistical significance). CONCLUSION: This study showed that appropriate management of hip fracture could maintain the dependency status of hip-fractured women for one year. However, Thai women aged 50 years and over with hip fracture had a higher mortality rate than those without hip fracture which suggests that hip fracture might be a sign of poor health status among these elderly women. PMID- 11253881 TI - Multiple intracranial aneurysms: incidence and management outcome in King Chulalongkorn Memorial Hospital. AB - We retrospectively reviewed the 380 patients on whom surgery was performed for intracranial aneurysms between January 1987 and December 1997. The incidence of multiple intracranial aneurysms (MIA) in our hospital was 8.7 per cent (33/380 cases). The management outcome of 33 patients with MIA was assessed 6 months after SAH. The outcome was poorer for patients with MIA than for those with a single intracranialaneurysm (SIA). The mortality and morbidity in all grades were 24.2 per cent in patients with MIA and 16.7 per cent and 19.6 per cent respectively in those with SIA. Delayed neurological deficit and treatment outcome of poor grade patients had significant contribution to outcome in patients with MIA, more than in patients with SIA. PMID- 11253883 TI - Intracameral pilocarpine in topical phacoemulsification. AB - PURPOSE: To evaluate the efficacy and corneal toxicity of intracameral pilocarpine. METHOD: A randomized, control trial using contralateral eye as control was designed to evaluate the effect of intracameral pilocarpine during phacoemulsification in 30 patients. 0.13 mg/ml pilocarpine in BSS was used as an irrigating solution to remove viscoelastic agents at the end of the operation while BSS was used in the control group. The outcome measurements composed of intraoperative pre and post irrigation pupil diameter, pre and post operative endothelial cell count and corneal thickness. SETTING: Priests Hospital. RESULTS: The pre-irrigation pupil size in the pilocarpine group and the control group was 7.62 +/- 0.75 mm and 7.60 +/- 0.77 mm respectively. The post-irrigation pupil size in the pilocarpine group and the control group was 5.40 +/- 0.79 mm and 7.18 +/- 0.79 mm. There were no statistically differences in pre and post-operative endothelial cell density, central corneal thickness, and the average corneal thickness between the pilocarpine group and the control group during six months follow-up. CONCLUSION: Intracameral pilocarpine in a low concentration (0.13 mg/ml) effectively constricts the pupil without significant changes of corneal endothelium compared to the control group. PMID- 11253884 TI - Dog-bite injuries at the Animal Bite Clinic of the Thai Red Cross Society in Bangkok. AB - Canine rabies remains a public health problem in Thailand and other developing countries. This study of animal bites at the Animal Bite Clinic at the Queen Saovabha Memorial Institute revealed that: (1) The majority of patients were bitten by dogs and the time of the attack was mostly during the day. (2) School aged children are at the highest risk for animal bites. (3) The most common site of injury are the legs and foot (64.2%), with the second most common site being the hands and fingers (21.2%). (4) Only 48 per cent of patients received rabies vaccine 1-2 days after being exposed. There was considerable delay before the rest received treatment. Solving Thailand' s rabies problem depends on control of canine rabies and educational campaigns. Public education must be an integral part of efforts to decrease the incidence of animal bites and assurance that they are managed properly. PMID- 11253885 TI - Clinical and laboratory findings in patients with pulmonary embolism in Phramongkutklao Hospital. AB - Pulmonary embolism (PE) was believed to be a rare disease and often misdiagnosed in Thailand. Only a few cases of PE in Thai patients have been reported. The purpose of this study was to describe the characteristics of history, physical examination and laboratory investigations in Thai patients with PE. Forty-nine patients diagnosed as PE in Phramongkutklao Hospital between 1994 and 1998 were included in the study. All patients underwent complete history, physical examination and appropriate laboratory studies. The mean age of this patient group was 53 years. Thirty-four per cent of these patients were first suspected of lung embolism while the others were misdiagnosed as congestive heart failure, myocardial infarction, pneumonia or septic shock. The most common syndrome was isolated dyspnea. Interestingly, chronic thromboembolic pulmonary hypertension which is uncommonly found in western countries was diagnosed in 12 per cent of our patients. Dyspnea, pleuritic pain, leg swelling, cough, tachypnea, tachycardia and increased pulmonary component of second heart sound were common symptoms and signs. A high-probability ventilation/perfusion lung scan and deep vein thrombosis were demonstrated in 93 per cent and 55 per cent of our patients, respectively. The mortality rate was 10 per cent. PMID- 11253887 TI - Treatment of benign paroxysmal positional vertigo by canalith repositioning procedure: experience from Srinagarind Hospital. AB - INTRODUCTION: Benign paroxysmal positional vertigo (BPPV) is one of the most common causes of vertigo. The diagnosis is confirmed by observing a classical response during the Dix-Hallpike maneuver. The cause of BPPV is usually idiopathic. There are two popular hypotheses described regarding the pathogenesis of BPPV. The first one is the "cupulolithiasis" hypothesis, and the second hypothesis, the so-called "canalithiasis" hypothesis. The clinical course of BPPV is spontaneous recovery in weeks or months. Treatments for BPPV have ranged from no intervention to surgical treatment. The new treatment, "Canalith-repositioning procedure (CRP)" which was introduced by Epley in 1992 produces a very high rate of success. This treatment has caused interest and has been modified and studied worldwide in recent years. OBJECTIVE: To study the efficacy of the canalith repositioning procedure that we modified from Epley's maneuver in the treatment of BPPV patients. DESIGN: A descriptive study. The BPPV patients, who came to the neurotologic clinic at Srinagarind Hospital from January 1997 to December 1998, were treated with our technique that was modified from Epley's maneuver. We neither used pre-medication, a mastoid oscillator, nor post-treatment instruction. RESULTS: The total number of patients included in this study was 19. The efficacy of this procedure for curing nystagmus and vertigo was 89.5 per cent. One patient did not follow-up and one patient did not respond to the CRP. Complication such as vago-vagal reflex, lateral canalithiasis, occurred in 5.3 per cent of the patients. The recurrence of BPPV in our study was 26.3 per cent. However, CRP was also effective in treatment of both patients with recurrence as well as those without recurrence. CONCLUSION: The canalith-repositioning procedure that is modified from Epley is effective in the treatment of BPPV. PMID- 11253886 TI - Safety and acceptable initial outcomes of reused balloon catheters for percutaneous transluminal coronary angioplasty. AB - OBJECTIVES: This study was conducted to compare the safety and initial outcomes applying reused balloon (RB) catheters with those of attained new balloon (NB) catheters when performing percutaneous transluminal coronary angioplasty. BACKGROUND: Recently, PTCA procedures have been used increasingly for the treatment of patients with coronary heart disease. In the era of national economic constraint, reused balloon catheters will reduce the cost of expensive, imported coronary angioplasty devices. Hence, data concerning the safety and success rate of RB catheters compared with NB catheters are urgently required. METHODS: Prospective comparative study between reused and new balloon catheters for coronary angioplasty. Data forms were completed after each procedure and before the patient was discharged after an 18-month period. RESULTS: From July 1996 to December 1997, 221 cases (121-RB, 100-NB) were enrolled. Mean age, ejection fraction, diseased vessel and lesion characteristics were similar in both groups. The number of lesions was much higher performed in the RB than in the NB group (1.7 +/- 0.9 vs 1.4 +/- 0.8, p = 0.02). The RB group had more cases of acute myocardial infarction than the NB group (7.4% vs 1%, p = 0.003), however, the angiographic and case success rate were the same (99.5% vs 97.9% and 98.3% vs 97% respectively). Major adverse cardiac events in RB amounted to 1.7 per cent and for NB to 1.0 per cent (p = ns). The total amount of balloons used in RB was much higher than in the NB group (1.5 +/- 0.6 vs 1.1 +/- 0.3, p = <0.0001). There were neither infection nor positive blood cultures in either group. CONCLUSIONS: Reused balloon catheters can be safely used for percutaneous transluminal coronary angioplasty with a high success rate. The total cost of angioplasty can be reduced without a decline in efficacy. PMID- 11253888 TI - Percutaneous balloon mitral valvulotomy with transesophageal echocardiographic monitoring: experience in Khon Kaen University. AB - To study the results and complications of Percutaneous Balloon Mitral Valvulotomy with Transesophageal Echocardiographic monitoring in patients with symptomatic mitral stenosis. From November 1996 to November 1998, PBMV with TEE monitoring was performed in 107 patients with symptomatic mitral stenosis. There were 72 females and 35 males, aged 19 to 65 years (mean 37.63). The mitral valve was successfully dilated in 104 patients. Immediately after PBMV, there was significant reduction of mean mitral valve gradient (17.89 +/- 6.7 mm Hg to 6.21 +/- 3.02 mm Hg), mean left atrial pressure (26.67 +/- 6.61 mm Hg to 13.97 +/- 4.7 mm Hg), mean pulmonary artery pressure (35.21 +/- 13.03 mm Hg to 27.71 +/- 10.31 mm Hg). Mitral valve area was increased from 0.80 +/- 0.24 cm2 to 1.75 +/- 0.42 cm2 and cardiac output was increased from 3.84 +/- 0.97 L/min to 4.74 +/- 1.09 L/min. Mitral regurgitation was detected in 20 patients, severe mitral regurgitaion appeared in one patient. None of these patients required emergency surgery. Cardiac tamponade was detected in one case and resolved by pericardiocentesis. TEE was well tolerated and no complications of TEE were detected. PBMV aided by TEE is safe and well tolerated. PMID- 11253889 TI - Situational analysis of the health insurance market and related educational needs in the era of health care reform in Thailand. AB - The purposes of this study were to explore the situation of health insurance in Thailand, to compare public and private perspectives and to identify related educational needs. Between March and April of 1998, the study employed in-depth interviews of 12 public and private major stakeholders of the health insurance systems, including policy makers, providers and insurers. Additional inputs were gathered in a brainstorming session with 41 participants from organizations with important roles in regulating, monitoring, paying, or providing health care services, as well as research and education. The findings indicated the health insurance market was expanding. But there was no national policy on health insurance. Insurance-related law was outdated. Public and private schemes overlapped, and were generally characterized by inadequate risk diversification, overutilization of services, lack of effective cost containment, inconsistent service quality, and poor understanding of health insurance principles. There were needs for more education and training in various aspects of health services management and health-insurance related functions. Consequently, continuing education and training related to health insurance services for policy makers, system administrators, managers, providers and insurers are strongly recommended during the health-care reform process. PMID- 11253890 TI - Prenatal diagnosis: 10-year experience. AB - To evaluate the indications and results of prenatal diagnosis of the high risk pregnant women attending the antenatal care clinic at Maharaj Nakorn Chiang Mai Hospital, Chiang Mai University during 1988-1998, we analysed 2,315 amniocenteses, 1,000 cordocenteses, and 11,895 obstetric ultrasound examinations. Among the amniocentesis group, 2,017 cases (87%) were done with the indication of advanced maternal age. The prevalence of major abnormal fetal chromosomes among high risk pregnancies was 1:58. Of 1,000 cases with cordocentesis, the most common indication was fetal risk of severe thalassemia (658 cases; 65.8%) and followed by fetal risk of chromosome abnormalities (272 cases; 27.2%). In the group of cordocentesis for diagnosis of thalassemia, 99 and 49 pregnancies were affected with Hb Bart's disease and homozygous beta-thalassemia, respectively. Thirty three cases with indication of chromosome analysis had fetuses with abnormal chromosomes. The major indications of ultrasonography included suspicion of intrauterine growth restriction (IUGR), determination of gestational age and screening anomalies, respectively. In conclusion, our extensive experience has enabled us to prenatally detect most fetuses with severe thalassemia, and fetuses with abnormal chromosomes as well as anomalies in a significant number, contributing a great deal to our population. Therefore, we recommend that systematic prenatal diagnosis, either amniocentesis, cordocentesis or ultrasound should be provided to every high risk pregnant woman for a healthy newborn. PMID- 11253891 TI - Prevalence of video games among Thai children: impact evaluation. AB - The present study was performed in order to determine prevalence and favored types of video games among altogether 679 primary and secondary school children in Thailand. To that end, the authors distributed questionnaires comprising detailed questions as to demographic data, playing frequency, available location and preferred type of video games among the parents of the children and adolescents to be investigated. Consistent with the literature, our results showed an early onset of video game playing (7.6 years), a higher prevalence among boys compared with girls, and a predilection for games invoking some aggressive behavior. In conclusion, although health hazards created by video game playing have remained beyond proof we still recommend parents and teachers to play a more active part as to the choice of games and the time spent playing. PMID- 11253892 TI - Esophageal foreign bodies. AB - A retrospective study of 310 patients with foreign bodies in the esophagus was analyzed. The most common age of the esophageal foreign body patients was between 0 to 9 years old (32.6%), and a coin was the most common foreign object in children. Bone, fish bone and bolus of meat were found commonly in adults, while dentures were encountered in the old age group. History of foreign body ingestion, dysphagia and odynophagia were usually presented by the patients. Roentgenographic study was useful in diagnosis and plan of management. Rigid esophagoscopy under general anesthesia is recommended in all patients with suspected history of foreign body, even though plain films of the chest and neck failed to demonstrate any significant findings. Complications can be reduced if the treatment is conducted within 24 hours of foreign body impaction. PMID- 11253893 TI - The risks of prostate cancer detection by transrectal ultrasound guide biopsy in Thai men with abnormal prostatic-specific antigen or abnormal digital rectal examination. AB - OBJECTIVE: To determine the risks of prostate cancer detection in Thai men with abnormal prostatic-specific antigen (PSA) or abnormal digital rectal examination (DRE). MATERIAL AND METHOD: One hundred and forty four Thai men with abnormal PSA or abnormal DRE or both were biopsied at the prostate gland with the use of transrectal ultrasound guide biopsy (TRUSBX). The risks of prostate cancer detection were evaluated. RESULTS: Mean age was 65.7 years old (S.D. = 9.88). The risks of positive biopsy according to the PSA levels of 0-4 ng/ml, 4.1-10 ng/ml, 10.1-20 ng/ml, 20.1-50 ng/ml, 50.1-100 ng/ml and more than 100 ng/ml were 6.25 per cent, 6.67 per cent, 10.8 per cent, 33.3 per cent, 60 per cent and 100 per cent, respectively. The risks of positive biopsy according to DRE appearances of total hard consistency, nodule, induration and benign prostatic hyperplasia were 57.1 per cent, 23.5 per cent, 34.6 per cent and 10 per cent, respectively. Of 144 men, 32 had adenocarcinoma of prostate. Radical prostatectomy was performed on 15 patients with clinically localized disease. Ten patients (66.6%) had free margin on their pathological specimens and 6 (40%) had organ confined disease. CONCLUSION: PSA testing alone or DRE alone is not a perfect test to diagnose prostate cancer since prostate cancer may present in men with normal PSA or men with no suspicious cancer DRE. For early detection of prostate cancer, both PSA testing and DRE need to be performed. When either PSA testing or DRE or both is abnormal, TRUSBX should be carried out. PMID- 11253894 TI - Successful combination chemotherapy (vincristine, procarbazine, etoposide, and prednisolone) in the treatment of inoperable, radioresistant low grade astrocytoma: a case report. AB - A case of successful combination chemotherapy using vincristine, procarbazine, VP 16 and prednisolone to treat an inoperable low grade astrocytoma is presented. This patient, whose tumor was also resistant to radiotherapy, had well controlled symptoms after the initiation of chemotherapy. A brain CT scan demonstrated disappearance of the tumor mass after eight courses of a combination chemotherapy regimen. She is at present symptom-free 80 months after diagnosis. This result suggests that combination chemotherapy may offer treatment modalities for low grade astrocytoma. PMID- 11253895 TI - Downbeating nystagmus and postural hypotension due to basilar invagination. AB - Downbeating nystagmus is an involuntary vertical rhythmic eye movement with the fast component in the downward direction. The sign indicates a craniocervical disorder. The most common cause is the Arnold-Chiari malformation, followed by cerebellar degeneration. Basilar invagination is a rare cause of downbeating nystagmus. However, with appropriate treatment its prognosis is good. Here, we report a case of basilar invagination which presented with downbeating nystagmus and postural hypotension. A 31 year-old Thai male patient had a 20 year history of postural hypotension. He had recurrent pneumonia and cough-induced syncope a year before admission. He complained of symptoms of an acute febrile illness and a productive cough. The physical examination showed high grade fever, postural hypotension and medium crepitation in the right upper lobe. The neurological examination showed downbeating nystagmus, atrophy and fasciculation of the right side of the tongue, atrophy of the right sternocleidomastoid muscle, mild weakness of the extremities and generalized hyperreflexia. The cervical spine X ray revealed upward displacement of the vertebral bodies of C1 and C2, with a mild narrowing of the space between C1 and the occiput. The CT-myelogram and MRI showed upward displacement of C1 with overriding of the dens over the anterior lip of the foramen magnum; this also compressed the medulla. Syringomyelia was seen at the C1-C5 level. We report a patient who presented with postural hypotension, recurrent pneumonia and downbeating nystagmus due to basilar invagination. The symptoms were aggravated by cough which caused an increase in intracranial pressure. This resulted from medulla compression in the foramen magnum by the first cervical spine. The treatment of choice was surgical decompression. PMID- 11253896 TI - Hallervorden-Spatz syndrome in two siblings diagnosed by clinical features and magnetic resonance imaging. AB - The Hallervorden-Spatz syndrome (HSS) is a rare condition characterized by extrapyramidal and pyramidal signs, dystonia, dysarthria, retinal degeneration, dementia and a progressive course. The development of magnetic resonance imaging (MRI) has increased the number of clinical and pathological reports of HSS. MRI pallidal abnormalities are called "eye of the tiger" signs. The combination of clinical features and MRI findings can be considered as highly suggestive of a diagnosis of HSS. Patient 1 was a 28 year old man who had been well until the age of 25 years. He developed dysarthria, difficulty with his gait and dystonia in his arms at the age of 28 years. Patient 2 was a 33 year old man who was the older brother of the first patient. He developed gait difficulty, tongue dystonia and dystonia of both arms at the age of 25 years. Each patient had spastic gait, dysarthria, dystonic posturing of both arms and generalized hyperreflexia, but no Kayser-Fleischer rings or retinitis pigmentosa. Blood chemistry, urine copper, serum copper and serum ceruloplasmin were all normal. MRI of the brain showed the "eye of the tiger" sign in the globus pallidus on T2 weighted images. These siblings had clinical features and MRI findings consistent with HSS. They are the first to be reported in Thailand. PMID- 11253898 TI - Anesthetic management of cerebral aneurysm clipping using the deep hypothermic circulatory arrest technique: a case report. AB - Deep hypothermic circulatory arrest may prove advantageous during surgery of some technically difficult brain lesions. This technique was first applied in one patient with a large intracavernous aneurysm which had failed standard neurosurgical techniques. For this technique to be successful the cooperation of neurosurgeons, cardiovascular surgeons, anesthesiologists, perfusionists and nurses is essential. Techniques aimed at improving the outcome include a short period of circulatory arrest, the depth of hypothermia, barbiturate administration, coagulation management and well-controlled blood glucose levels. The total time of circulatory arrest and the thiopentone dosage were 61 minutes and 1,700 mg respectively. The lowest core temperature was 13.9 degrees C. The positive outcome supports the use of this technique in selected patients with complex intracranial vascular lesions who may not be operable by standard techniques. PMID- 11253897 TI - Persistent Mullerian duct syndrome in adult: a case report. AB - A twenty-eight year old phenotypically and genotypically normal male adult was admitted with a right inguinal hernia, a right retractile testis and left cryptorchidism. During surgery, he was found to have a uterus in the right spermatic cord and a left undescended testis (intra abdominal type). The uterus was excised, left orchidectomy, right orchidoplexy and right herniorrhaphy were performed. This rare case is reported as persistent Mullerian Duct Syndrome (PMDS) which is caused by impaired action of Mullerian Inhibiting Substance (MIS) in regressing the Mullerian duct. PMID- 11253899 TI - Cutaneous tuberculosis in three HIV-infected patients. AB - Although tuberculosis is a common disease in patients infected with HIV, cutaneous presentation is not commonly found. The authors report three HIV infected patients with cutaneous tuberculosis and lung involvement. Patient 1 presented with a nodular skin lesion on the right forearm and the diagnosis was confirmed by histopathology and PCR study. Patients 2 and 3 presented with generalized erythematous papules and vesicopustules on the trunk and extremities. Culture grew M. tuberculosis in patient 2 and M. tuberculosis DNA was detected in the skin lesion of patient 3 by the PCR method. PMID- 11253900 TI - About the death penalty in Thailand. PMID- 11253901 TI - Commensurate ratings of health care. AB - A new descriptive item statistic, termed the mean cumulative logit (MCL), is advocated for scoring ratings of health care at the population level. The advantages of the MCL are demonstrated on data from the Consumer Assessment of Health Plans Study (CAHPS). The CAHPS data require (1) the comparison of binary and ordinal ratings in a common metric, (2) a treatment of unit and item nonresponse, and (3) the control of ordinal item correlations. These requirements are handled by a cumulative logit model that is applicable to unweighted (incomplete) and weighted (complete) data. The former case gives item satisfactions from a patient perspective. The latter case generates these satisfactions as social utilities from a provider perspective. From both of these viewpoints the perceived quality of health care is greater for fee-for-service plans than managed care plans in the field test population studied here. PMID- 11253903 TI - Measuring disability: application of the Rasch model to activities of daily living (ADL/IADL). AB - This paper describes a comparative analysis of (ADL) and (IADL) items administered to two samples, 4,430 persons representative of older Americans, and 605 persons representative of patients with rheumatoid arthrisit (RA). Responses are scored separately using both Likert and Rasch measurement models. While Likert scoring seems to provide information similar to Rasch, the descriptive statistics are often contrary if not contradictory, and estimates of reliability from Likert are inflated. The test characteristic curves derived from Rasch are similar despite differences between the levels of disability with the two samples. Correlations of Rasch item calibrations across three samples were .71, .76, and .80. The fit between the items and the samples, indicating the compatibility between the test and subjects, is seen much more clearly with Rasch with more than half of the general population measuring the extremes. Since research on disability depends on measures with known properties, the superiority of Rasch over Likert is evident. PMID- 11253902 TI - Maintaining instrument quality while reducing items: application of Rasch analysis to a self-report of visual function. AB - While efficiency has been of concern in the measurement of health care outcomes, little attention has been devoted to methods that achieve efficient, shortened instruments that have good psychometric properties. The purpose of this study was to show how Rasch analysis could be used to reduce the number of items in an instrument while maintaining credible psychometric properties. This approach was applied to the Visual Function-14 (VF-14), a self-report of 14 vision-dependent activities, designed to measure the need for and outcomes of cataract surgery. An instrument which contained the VF-14 plus an additional 10 items that were developed for the study (VF-24) was administered to sixty-one patients (73.7+/ 9.5 years) about to undergo extracapsular cataract removal at one of two surgical centers. Rasch analysis (BIGSTEPS) of the VF-14 showed a number of limitations to the original instrument, including: 1) unequal use of the five rating categories, 2) ceiling effect, 3) several other gaps where patient abilities did not match with item difficulties, and 4) sets of items that appeared redundant, (i.e., having the same calibration level). To resolve the first three of these problems, the rating scale was converted to a three-point scale and BIGSTEPS was run with all 24 items. (10 additional items added to the VF-14 designed to "fill in" the gaps). The conversion to a three-point scale and the increase in items resulted in some improvement in the matching of item difficulty to patient ability, as evidenced by a slight decrease in gaps. The addition of items resulted in improvements in person separation (2.55 to 2.99) and Cronbach's alpha (.83 to .91) but did not substantially reduce the ceiling effect and furthermore resulted in an increase in item redundancy. The final practical improvement undertaken was to reduce the number of items while attempting to maintain the psychometric qualities of the instrument as a whole. Three criteria were used in deciding to remove items: 1) high mean square, 2) low mean square and 3) items having similar calibrations. In addition, if an analysis showed that the removal of an item substantially decrease person separation, that item was retained for further analyses. Relative to the original VF-14, the resulting VF-10 showed less redundancy of items while person separation (2.20) and Cronbach's alpha (.89) remained relatively intact. The study demonstrates that Rasch analysis, while effective in elucidating the metrics of an original instrument, can also be useful in designing modifications of instruments that are both efficient and psychometrically sound. PMID- 11253904 TI - Payment and provider profiling of episodes of illness of clinical illnesses involving rehabilitation. AB - The prevailing trend in American health care finance for the last two decades and likely for the forseeable future, is the movement from a system based on fee for service payments to one dominated by capitation arragements. At the core of this change is a shifting of risk from payers to providers. Such a fundamental transition is not without its difficulties. This is exemplified by some of the problems experienced by the Health Care Financing Administration (HCFA) as it has begun to encourage beneficiaries to move from traditional fee for service Medicare to capitated HMOs. The most recently published regulations involving risk adjustment of rates of payment for managed care organizations (MCOs) did not find much favor as the statistical power was poor and the clinical meaningfulness of the risk adjustment was highly problematic. Specifically, the risk adjustment explained less than 10% of the variation in costs (compared to approximately 30% when DRGs were first implemented). From a clinical perspective, the methodology only used hospitalization data for adjustment of capitation rates; that is, anyone who was not hospitalized was assumed to be healthy! It is known that payment for rehabilitation services is among the most difficult to understand and predict of all medical care. This article will summarize the development of a new risk adjustment methodology which should be particularly useful for payment and monitoring of episodes of clinical conditions that involve rehabilitation. The article will conclude with directions for future research. PMID- 11253905 TI - Descriptive epidemiology of male breast cancer in Osaka, Japan. AB - Male breast cancer is rare. The total number of incidence in Osaka for the period of 1966-95 was 182. Male-to-female ratio for breast cancer incidence was 1:164 in Osaka during this period. Mean age of the male breast cancer incidence was 63.3. The numbers of incidence and the crude incidence rates for male breast cancer have increased during the last 3 decades, while the age-standardized rates have remained constant. The age-specific incidence rates for males showed a gradual increase with age, while those for females showed a steep increase beginning at twenty years of age and a peak around 45-49 or 50-54 years old. The age standardized incidence rates of male breast cancer were lower in Japan than in European countries and North America, as were those of female breast cancer. Distributions of the histological type and the extent of disease were not significantly different between males and females. Relative 5-year survival for the male breast cancer was, however, lower than that for the female, especially in the "regional" stage and "distant" stage. Further studies on the sex difference in survival will be mandatory based on high-quality hospital cancer registries' data, which provide detailed information on the clinical stage and treatment. PMID- 11253906 TI - Characteristics of cataracts in the Chinese Singaporean. AB - PURPOSE: To conduct an epidemiological survey of cataracts and examine the characteristics of lens opacities in Chinese Singaporeans. The results are then compared with those from two similar surveys previously done in Japan in Noto Area, Honshu, and Amami, Kyushu, respectively. SUBJECTS AND METHODS: 468 subjects of > or = 50 years of age, who responded to the invitation to participate, were examined. Examination principally included photo-documentation of the anterior and posterior segments of both eyes. Evaluation and grading of lens opacities were done using graphical analysis of Scheimpflug and retro-illumination images. Inter-group comparisons were based on statistical analysis of cataract prevalence and distribution. RESULTS: The prevalence of clear lenses decreased with aging with no significant difference between males and females--a finding common to Singapore and the two Japanese study groups. The prevalence of cataracts (or lens opacities of Grade II and above) in 60-79 year-old Singapore subjects was significantly higher than Noto and Amami subjects in the same age group. Further, cortical opacity was the main type in Singapore subjects in their 50s and which was significantly higher than Noto subjects of the same age group. In all age groups, the distribution and prevalence of both nuclear and subcapsular types in the Singapore group were higher than the two Japanese study groups. CONCLUSIONS: Cataracts in Chinese Singaporeans are characterized by a high prevalence of nuclear opacities which was generally seen in tropics and sub-tropics. Our study also suggested the involvement of solar-UV in cortical cataracts as well as that of additional risk factors, such as environmental temperature and race, in nuclear and subcapsular cataract formation. PMID- 11253907 TI - A necessary and sufficient condition of comparability for using standardized mortality ratio (SMR). AB - A necessary and sufficient condition of comparability for using SMR was studied mathematically by considering the equivalence between SMR and CMF, as CMF was a perfectly comparable index. This condition was expressed by either proportionality of mortality vectors or proportionality of projected person-years to the plane spanned by mortality vectors of reference and index groups. We could obtain another expression of the condition, in which affect of distortions were easily understood, which consist of three factors: distortion of proportionality of mortality, distortion of person-years and similarity of distortions. Our results were applied to study the mortality of biliary tract cancer in Ibaraki Prefecture. Places where absolute difference between CMF and SMR exceeds some criterion (say, 0.15) were Satomi, Ushiboiri, Nihari in males and Gozenyama, Suifu and Asahi in females. All three distortion indices exceeded their upper 95% percentiles in Satomi in males. PMID- 11253908 TI - The defibrillation system of basic emergency medical technicians in Japan: a comparison with other systems from a 14-year review of out-of-hospital cardiac arrest reports. AB - Although seven years have passed after basic emergency medical technician (EMT) defibrillation system was introduced in Japan, the overall survival for out-of hospital cardiac arrest (OHCA) remains poor. We investigated factors leading to such an unanticipated result in Japan by comparing the data of OHCA in Japan with those in other countries. We obtained population-based OHCA data from three communities in Japan. We also performed a comprehensive literature and manual search to identify reports that included rates for incidence and survival or provided sufficient data for the calculation of these rates of OHCA. Statistical analysis was performed to compare survival and incidence rates between the communities. We identified 36 articles from 16 countries by a comprehensive literature search. There was no significant difference in incidence and survival rates among communities in Japan. Although the incidence rate of collapse witnessed OHCA with ventricular fibrillation (VF) was much lower in Japan than western countries, the proportions of survival from it were comparable to those. Basic-EMT defibrillation system in Japan has yielded excellent result in terms of the survival of VF cases. However, much lower proportion of VF to all cases is responsible for lower overall rates of survival from OHCA in Japan. PMID- 11253909 TI - An international comparison of the involvement of epidemiology in the most frequently cited publications in the field of clinical medicine. AB - The objectivity, validity and credibility of research in clinical medicine can be enhanced by the appropriate involvement of epidemiology. However, the overall contribution of epidemiology to clinical research, either as a methodology or as a resource for research, has been poorly quantified. We therefore assessed the involvement of epidemiology in influential publications in the field of clinical medicine, and made an international comparison on a quantitative basis. The 500 most frequently cited papers published during 1981-96 in the field of clinical medicine in the US, the UK, and Japan were compared in terms of epidemiological involvement using predetermined criteria. The three criteria were based on the indexing of relevant MeSH keywords, publication types, or the departmental affiliations of the authors. For all three criteria, the proportion of clinical papers with epidemiological involvement was the highest in the US, followed by the UK, whereas it was the lowest in Japan. The difference was almost four-fold between the US and Japan. There was also an increasing trend of epidemiological involvement in publications of clinical medicine over the years, which was more apparent in the US than in either the UK or Japan. These findings may reflect inter-country differences in resources as well as in the stance towards evidence based health sciences. PMID- 11253910 TI - Randomized controlled trials conducted in Japan as a comparison with top-ranking countries. PMID- 11253911 TI - Long-term prognosis after stroke: a community-based study in Japan. AB - Stroke is the leading cause of severe disability in the elderly. Under the national insurance for care and assistance for the elderly starting in 2000, data must be obtained on the prognostic status of stroke patients in communities. We identified 322 incident strokes in six communities (total census population=71,610) during the two- or three-years survey period between 1987 and 1990, and we completed a follow-up of the respective prognoses of most of these patients at one, three, and five years after the onset (n=315 stroke patients) (98%). One year after stroke, 33% of the 315 strokes were dead, 13% were dependent, and 54% were independent. After three years, 44% were dead, 13% were dependent, and 43% were independent. After five years, 52% were dead, 9% were dependent, and 39% were independent. The long-term prognosis was poorer with increased age, and poorer for women than for men except in the case of men ages less than 55 years old at onset. Among patients who were dependents, the proportion of taken care at home was approximately 30% one year after onset, and 50% three to five years after onset. It is estimated that approximately 17 dependents from 127 incident strokes in a population of around 70,000 every year. Because the average survival time of dependents was about 4 years, the prevalence of dependents is estimated to be 68, indicating that the prevalence is about 10 persons per 10,000. Over the period of this study, and as compared with the reported proportions in community-based studies in the 1970's, the proportion of deaths declined and that of independents increased, probably due to reduced severity of stroke. However, the proportion of dependents did not change significantly over time. Thus, under the terms of the new national insurance, it is essential for family and communities to cooperate in taking care of dependent stroke patients. PMID- 11253912 TI - Refinement of the NMR structures of alpha-conotoxin MI using molecular dynamics simulation with explicit solvent water and a full molecular force field. AB - Three NMR structures of alpha-conotoxin MI, a potent antagonist of the nicotinic acetylcholine receptor, have been refined using molecular dynamics (MD) simulation with explicit water. Although the convergence of the NMR structures of alpha-conotoxin MI was not sufficient to provide detailed structural features, the average structures obtained from MD simulations converged to one conformation, providing structural characteristics. The resulting structure was also found to be consistent with the results of amide proton-exchange experiments. These results demonstrate that MD simulation with explicit solvent water is very useful in refining NMR structures. PMID- 11253913 TI - Interaction of diphenyltin(IV) dichloride with some selected bioligands. AB - The interaction of diphenyltin(IV) with selected bioligands having a variety of model functional groups were investigated using the potentiometric technique. The hydrolysis constants of diphenyltin(IV) cation and the step-wise formation constants of the complexes formed in solution were calculated using the non linear least-squares program MINIQUAD-75. The participation of different ligand functional groups in binding to organotin is discussed. The concentration distribution of the various complex species was evaluated as a function of pH. PMID- 11253914 TI - Novel non-peptide GPIIb/IIIa antagonists: synthesis and biological activities of 2-[4-[2-(4-amidinobenzoylamino)-2-(substituted)acetyl]-3-(2-methoxy-2-oxoethyl)-2 oxopiperazinyl] acetic acids. AB - To improve the in vitro and in vivo potency of our first low molecular weight GPIIb/IIIa antagonist 1 (TAK-029), a series of 2-[4-[2-(4-amidinobenzoylamino)-2 (substituted)acetyl]-3-(2-methoxy-2-oxoethyl)-2-oxopiper-azinyllacetic acids were synthesized through modification of the glycine moiety of 1 and evaluated for their ability to inhibit in vitro adenosine 5'-diphosphate (ADP)-induced platelet aggregation of guinea pig platelet rich plasma (PRP). Among the compounds examined, the (3S,2S)-4-methoxyphenylalanine derivative 4h showed the most potent antagonistic activity with an IC50 value of 13 nM. Dose-dependent inhibition of ex vivo platelet aggregation was achieved with oral administration of 4h (0.3-1.0 mg/kg) to guinea pigs. Complete inhibition was observed for up to 8 h, and 43% inhibition could still be observed 24 h after oral administration of 1.0 mg/kg. The long-lasting antiplatelet effect of 4h suggests that 4h would be suitable for once-a-day dosing. Structure-activity relationships (SAR) were examined in the series of the phenylalanine derivatives. An increase in the electron density around the 4-position of the phenyl ring of the phenylalanine moiety led to an increase in the antiplatelet activity, suggesting the existence of a hydrophobic and electrostatic interaction site in addition to the ionic binding sites in the GPIIb/IIIa. PMID- 11253915 TI - Orally active GPIIb/IIIa antagonists: synthesis and biological activities of masked amidines as prodrugs of 2-[(3S)-4. AB - To improve the in vivo potency of the potent GPIIb/IIIa antagonist 2-[(3S)-4 [(2S)-2-(4-amidinobenzoylamino)-3-(4-methoxyphenyl)propanoyl]-3-(2-methoxy-2 oxoethyl)-2-oxopiperazinyljacetic acid (4), the amidino group was converted to an oxadiazole ring, thiadiazole ring or substituted amidoxime group. These groups were expected to be metabolized to an amidino group in vivo. The compounds synthesized were evaluated for their potency to inhibit the ex vivo adenosine 5' diphosphate (ADP)-induced aggregation of guinea pig platelets. Among the compounds examined, the methoxycarbonyloxyamidine 8a exhibited the most potent ex vivo inhibitory activity with a fast onset and prolonged duration of action after oral administration. PMID- 11253916 TI - Indirect potentiometric titration of sulphamethoxazole in the presence of trimethoprim in co-trimazole tablets using copper based mercury film electrode. AB - A simple and rapid indirect potentiometric titration of sulphamethoxazole in the presence of trimethoprim contained in co-trimazole tablets is described. The method is based on the formation of a complex of sulphamethoxazole with a known excess of silver ions and the titration of unreacted silver ion potentiometrically using an inexpensive lab-made copper based mercury film electrode (CBMFE). The titration conditions have been optimized for the determination of 1.0-10.0 mg of sulphamethoxazole in pure and dosage forms. The precision and accuracy of the method have been assessed by the application of lack of fit test and other statistical methods. Overall mean recovery and relative standard deviations obtained were 99.88% and 1.32% (n=7) respectively. No interference was caused by other excipients present in pharmaceutical dosage forms. The application of this method for sulphamethoxazole assay in the presence of trimethoprim in tablets was validated by the comparison of results obtained by the proposed method with that of the British Pharmacopoeia (BP) method using F- and t-statistical tests of significance. PMID- 11253917 TI - Bufadienolides and a new lignan from the bulbs of Urginea maritima. AB - Thirty three compounds were obtained from the bulbs of Urginea maritima (Liliaceae). The compounds were identified by means of fast atom bombardment mass spectrometry (FAB-MS), nuclear magnetic resonance (1H-NMR), (13C-NMR), nuclear overhauser effects (NOE) and two dimensional (2D) NMR. Ten of them were new natural compounds. Nine were bufadienolides and only one was lignan in these compounds. PMID- 11253918 TI - Efficient photocleavage of DNA by cationic porphyrin-acridine hybrids with the effective length of diamino alkyl linkage. AB - Positively charged porphyrins bearing an acridine with various lengths of diamino alkyl linkage, 5-[4-[(6-chloro-2-methoxy-9 acridyl)aminoalkylaminocarbonyl]phenyl]-10,15,20-tris(4-N methylpyridiniumyl)porphine triiodide, alkyl=ethyl, butyl, hexyl, or octyl, were synthesized. They exhibited more enhanced photocleavage activity of pUC18 plasmid DNA than TMPyP, meso-tetrakis(4-N-methylpyridiniumyl)porphine, which is well known to bind to DNA tightly and to cleave DNA effectively; the hybrid linked with the hexamethylene chain showed particularly high activity. An equilibrium dialysis experiment demonstrated that the binding ability of the hybrids to calf thymus (CT) DNA correlated quantitatively with the photocleavage activity. The lack of the substantial red-shift of the Soret maxima of the hybrids through the titration with CTDNA denied the intercalative binding of the porphyrin part. In their circular dichroism (CD) spectral change on binding to CTDNA, two negative peaks appeared at 275 nm and at 285-290 nm in the UV range. The latter negative peak was observed for hybrids, but not for TMPyP, and thus we assigned it to induced CD (ICD) derived from intercalation of acridine chromophore. In the visible range, the hybrids showed only a positive peak around their Soret maxima, and this feature suggested the porphyrin moiety lay in the DNA groove. In addition, the length of the linker markedly influenced the ellipticity of their visible ICD, suggesting that the proximity of the porphyrin moiety to DNA was greatly affected by the linker. PMID- 11253919 TI - Hydrogen peroxide-induced deacetylation of acetyl resorufin as a novel indicator reaction for fluorometric detection of glucose using only glucose oxidase. AB - Hydrogen peroxide (H2O2)-induced deacetylation of non-fluorescent acetyl resorufin (1) to fluorescent resorufin (2) as a novel indicator reaction for fluorometric detection of glucose using only glucose oxidase (GOD) is described. When a 1:1:1 mixture of 1 (in CH3CN), glucose, and GOD (each in pH 7.4 phosphate buffer) was incubated at 25 degrees C under aerobic conditions, the resulting solution turned yellow to fluorescent pink due to 2. The formation of 2 was markedly retarded on incubation under anaerobic conditions. When a mixture of 1 and H2O2 was incubated under aerobic conditions, the formation of 2 was noted as in the case of the enzymatic reaction of 1. These results demonstrated that the observed color change is brought about through deacetylation of 1 to 2 induced by H2O2 generated in GOD-catalyzed oxidation of glucose. With regard to the fluorometric traces of the enzymatic reaction with 1 (0.2 mM), GOD (0.5 mg/ml), and glucose at 25 degrees C, fluorescence intensity exhibited a linear relationship against glucose concentration between 0.2 and 2.0 mm, with a correlation coefficient of 0.997. Neither ascorbic acid, uric acid, nor bilirubin significantly interfered with the transformation of 1 to 2 through GOD-catalyzed oxidation of glucose. PMID- 11253921 TI - Dimer and superstructure of the active form of a vitamin D3; 1 alpha,24(R)dihydroxy-vitamin D3 monohydrate, C27O3H44.H2O. AB - The crystals of 1alpha24(R)dihydroxy-vitamin D3 monohydrate, C27O3H44.H2O are orthorhombic in the space group P2(1)2(1)2(1) with cell dimensions a=25.719, b=42.572, c=9.851A and Z=16. The asymmetric unit consists of two subunits with b/8, and each subunit contains a dimer in which two molecules are hydrogen-bonded through water molecules into non-crystallographical symmetry of C2. The two-fold axes are the straight lines, x=1/2, z=0.256 and x=1/2, z=0.623. The two dimers are the same in the rigid ring part, but differ in the conformation of the flexible chains. The dimers further make C2 symmetry between the rigid ring parts to form a superstructure, and the two-fold axis of the straight line, y=1/8, z=0.435 goes through a point that is a little apart from the hypercenter (1/2, 1/8, 1/2). The structure was solved by integrated Patterson and direct methods and refined on Fo2 under restraints. The final R1 is 0.228 on Fo for 1623 reflections with Fo>3sigma, resolutions 1.0-3.0 A, 313 restraints, 490 parameters and average Ueq=0.120. Not all the atoms of the chains appeared nor the hydrogen atoms. The missing atoms of the dimer were modeled from another pair molecule by C2 symmetry and hydrogen atoms were added. The structure of the dimer was optimized by ab initio molecular orbital of HF/6-31G. PMID- 11253920 TI - Relationship between effects of phenolic compounds on the generation of free radicals from lactoperoxidase-catalyzed oxidation of NAD(P)H or GSH and their DPPH scavenging ability. AB - The influence of various phenolic compounds on the lactoperoxidase (LPO)/hydrogen peroxide (H2O2)-catalyzed oxidation of biochemical reductants such as reduced beta-nicotinamide adenine dinucleotide (NADH), reduced beta-nicotinamide adenine dinucleotide phosphate (NADPH) or reduced glutathione (GSH) was investigated by electron spin resonance (ESR) spectroscopy. Micromolar quantities of phenolic compounds such as 17beta-estradiol, phenol, and p-chlorophenol enhanced the LPO/H2O2-catalyzed oxidation of NAD(P)H or GSH to generate a large amount of superoxide radical (O2*-) or glutathione thiyl radical (GS*), while, phenolic compounds such as quercetin and Trolox C greatly suppressed the generation of O2* and GS*. In order to elucidate the effects of phenolic compounds on the generation of O2*- and GS*, their quenching activities for a stable radical, 1,1 diphenyl-2-picrylhydrazyl (DPPH), were investigated by ESR spectroscopy. 17beta Estradiol, phenol, and p-chlorophenol showed very weak scavenging activities for DPPH, but quercetin and Trolox C showed strong activities. This suggests that the ability of phenolic compounds to enhance LPO/H2O2-catalyzed oxidation of NAD(P)H or GSH relates inversely to their ability to quench DPPH. That is, phenolic compounds having weak quenching activity against DPPH may enhance the LPO/H2O2 catalyzed oxidation of NAD(P)H or GSH to generate a large amount of O2*- or GS*. PMID- 11253923 TI - Steroidal saponins from Hemerocallis fulva var. kwanso. AB - Two steroidal saponins, hemeroside A and B, were isolated from the aerial part of Hemerocallis fulva var. kwanso for the first time. The structures of these compounds were established as 24S-hydroxy-neotokorogenin 1-O-alpha-L arabinopyranosyl 24-O-beta-D-glucopyranoside (1) and isorhodeasapogenin 3-O-beta D-glucopyranosyl-(1-->3)-[beta-D-xylopyranosyl-(1-->2)]-beta-D-glucopyranosyl-(1- >4)-beta-D-galactopyranoside (2) through NMR experiments. PMID- 11253922 TI - (10Z)- and (10E)-19-fluoro-1alpha,25-dihydroxyvitamin D3: an improved synthesis via 19-nor-10-oxo-vitamin D. AB - An efficient synthetic route to (10Z)- and (10E)-19-fluoro-1alpha,25 dihydroxyvitamin D3 was developed. The key feature of this pathway is the introduction of a 19-fluoromethylene group to a (5E)-19-nor-10-oxo-vitamin D derivative. The 10-oxo-compound was obtained via a 1,3-dipolar cycloaddition reaction of (5E)-1alpha,25-dihydroxyvitamin D with in situ generated nitrile oxide followed by ring cleavage of the formed isoxazoline moiety with molybdenum hexacarbonyl. Conversion of the keto group of (5E)-19-nor-10-oxo-vitamin D to the E and Z fluoromethylene group was achieved through a two-step sequence involving a reaction of lithiofluoromethyl phenyl sulfone followed by the reductive desulfonylation of the alpha-fluoro-beta-hydroxy sulfone. The dye-sensitized photoisomerization of the (5E)-19-fluorovitamin D afforded the desired (5Z)-19 fluorovitamin D derivatives, (10Z)- and (10E)-19-fluoro-1alpha,25 dihydroxyvitamin D3. PMID- 11253924 TI - Cyclooxygenase-2 inhibitory cerebrosides from phytolaccae radix. AB - A mixture of cerebrosides, called poke-weed cerebrosides, was purified from Phytolaccae Radix (Phytolaccaceae) and characterized as 1-O-beta-D glucopyranosides of phytosphingosine type ceramides comprised of a common long chain base (2S,3S,4R,8Z)-2-amino-8-octadecene-1,3,4-triol and fatty acids. The fatty acyl chain of ceramide moieties was determined as (2R)-2 hydroxypentacosanoic acid, (2R)-2-hydroxylignoceric acid, (2R)-2 hydroxytricosanoic acid, (2R)-2-hydroxybehenic acid, (2R)-2-hydroxypalmitic acid, and palmitic acid. The pokeweed cerebroside inhibited the cyclooxygenase-2 dependent phase of prostaglandin D2 generation in bone marrow-derived mast cells in a concentration dependent manner with an IC50 of 6.2 microg/ml. PMID- 11253925 TI - Selective electrocatalytic oxidation of N-alkyl-N-methylanilines to N alkylformanilides using nitroxyl radical. AB - Electrocatalytic oxidation of N-alkyl-N-methylanilines was studied using 4 benzoyloxy-2,2,6,6-tetramethyl-piperidinyl-N-oxyl as a nitroxyl radical. The reaction with N-alkyl-N-methylanilines led to direct formation of N alkylformanilides in the presence of H2O in reaction media in adequate conversion (>75.8%), high current efficiency (>89.2%) and high selectivity (>93.8%). PMID- 11253926 TI - A benzofuran glycoside and an acetylenic acid from the fungus Laetiporus sulphureus var. miniatus. AB - A new benzofuran glycoside, masutakeside I (1) and a new C10 acetylenic acid, masutakic acid A (2) were isolated from the fruiting bodies of the fungus Laetiporus sulphureus var. miniatus. Their structures were established by spectroscopic and chemical methods. The known compounds 3-6 were also obtained and identified as egonol, demethoxyegonol, egonol glucoside and egonol gentiobioside. Some of these compounds exhibited cytotoxicity against Kato III cells. PMID- 11253927 TI - Stereoselective reactions. XXXIIII. Design and synthesis of chiral bidentate amines having a bulky group on the chiral carbon. AB - Based on the solution structures of chiral bidentate lithium amides ((R)-3a,b) having a phenyl group on the chiral carbon, chiral bidentate amines ((R)-5a,b 8a,b and (S)-9a,b) having a bulkier group instead of a phenyl group on the chiral carbon were designed and synthesized. PMID- 11253929 TI - The practical synthesis of (2S)-7-methoxy-1,2,3,4-tetrahydro-2-naphthylamine via optical resolution of 2-(3-methoxybenzyl)succinic acid. AB - We describe the practical synthetic route for (2S)-7-methoxy-1,2,3,4-tetrahydro-2 naphthylamine 1(2S)-2-amino-7-methoxytetraline; (S)-AMT]. (2R)-2-(3 Methoxybenzyl)succinic acid [(R)-1] was obtained by the optical resolution of 2 (3-methoxybenzyl)succinic acid (1) as the salt of (1R,2S)-2 (benzylamino)cyclohexylmethanol (7), and (R)-1 was converted to the optically active (2S)-7-methoxy-1,2,3,4-tetrahydro-2-naphthoic acid [(S)-2] by the intramolecular Friedel-Crafts reaction followed by catalytic hydrogenation. (S) AMT was obtained from the acid (S)-2 by Hofmann rearrangement without racemization. PMID- 11253928 TI - Preparation and root growth-modulatory activity of N-substituted 2-acetylamino-2 ethoxycarbonyl-3-(2-furyl)propanamides. AB - N-Substituted 2-acetylamino-2-ethoxycarbonyl-3-(2-furyl)propanamides (8) were synthesized through the reaction of amines (13) with 2-acetylamino-2 ethoxycarbonyl-3-(2-furyl)propanoic acid (3b), which was prepared via condensation of 2-(bromomethyl)furan (10b) with diethyl acetamidomalonate, followed by partial hydrolysis of the resultant diethyl ester (3a) in the presence of barium hydroxide. However, bulky amines such as tert-butylamine or 2 trifluoromethylaniline did not afford the corresponding diamides (8). The biological activity of the prepared diamides (8) as root growth modulators was examined by germination assay using rape and leek seeds. N-(5-Bromo-2-thiazolyl)- and N-(4-chloro-2-benzothiazolyl)-2-acetylamino-2-ethoxycarbonyl-3-(2 furyl)propanamides (8h, i) both potently inhibited the root growth of rape seedlings, but were less effective in the case of leek seeds. The herbicide 2,4 dichlorophenoxyacetic acid completely inhibited root growth in both cases. PMID- 11253930 TI - Functional analysis of the iron(II) etiocorrphycene incorporated in the myoglobin heme pocket. AB - The iron(III) complex of 2,7,12,17-tetraethyl-3,6,11,18-tetra-methylcorrphycene, an isomeric heme, was complexed with apomyoglobin to examine the ligand binding ability of the novel macrocycle under physiological conditions. The reconstituted holoprotein was found to be functionally active at pH 7.4 and 20 degrees C and to bind oxygen and carbon monoxide reversibly with a half-saturation pressure at 6.7 and 3.5mmHg, respectively. Equilibrium affinities for these ligands are one to two orders of magnitude lower than those reported for native myoglobin. The functional anomaly was ascribed to the geometric and electronic strain on the iron(II) atom in the trapezoidal coordination core of corrphycene. PMID- 11253931 TI - Microencapsulation of pesticides and their improved handling safety. AB - The purpose of pesticide formulations and recent requirements of pesticide industry are discussed, and the importance of research work on pesticide delivery systems, is pointed out. For this purpose, the microcapsules, as one of the typical controlled release formulations, play an important role. Handling safety of the pesticides is improved considerably by microencapsulation due to hazard and exposure reduction. PMID- 11253932 TI - Encapsulation of retinoids in solid lipid nanoparticles (SLN). AB - SLN have been suggested for a broad range of applications, such as intravenous injection, peroral, or dermal administration. The incorporation of the drug in the core of the SLN has to be ensured for these applications, but the inclusion of drugs in SLN is poorly understood. This study is a contribution to further describe the inclusion properties of colloidal lipids and to propose incorporation mechanisms. Besides the well known methods to investigate entrapment of actives in nanoparticles such as DSC or microscopy, the present study focussed on yet a different approach. Based on the different chemical stability of retinoids in water and in a lipid phase, a method to derive information on the distribution of the drug between SLN-lipid and the water phase was established. Comparing different lipids, glyceryl behenate gave superior entrapment compared to tripalmitate, cetyl palmitate and solid paraffin. Comparing three different drugs, entrapment increased with decreasing polarity of the molecule (tretinoin < retinol < retinyl palmitate). The encapsulation efficacy was successfully enhanced by formulating SLN from mixtures of liquid and solid lipids. These particles were solid and provided better protection of the sensitive drugs than an emulsion. X-ray investigations revealed that good encapsulation correlated with a low degree of crystallinity and lattice defects. With highly ordered crystals, as in the case of cetyl palmitate, drug expulsion from the carrier was more pronounced. PMID- 11253934 TI - In vivo evaluation of submicron emulsions with pilocarpine: the effect of pH and chemical form of the drug. AB - Submicron emulsions containing 2.0% w/v pilocarpine as pilocarpine HCl, soybean oil (10% w/v) and egg lecithin (1.2% w/v) were formulated. Emulsions at pH 5.0, 6.5 and 8.5 were applied to the rabbit's eye, and the reduction in pupil diameter was measured for 6 h. The miotic effect was compared with that obtained with aqueous solutions at the same pH. A prolonged miotic effect was observed when the submicron emulsion was used as a vehicle. After application of emulsions at pH 5.0, 6.5 or 8.5, the time when 20% reduction of pupil diameter was still observed was 3.9 +/- 1.1 h, 4.3 +/- 1.3 h and 5.3 +/- 0.8 h, respectively, while, after application of a solution, this parameter was shorter by 30-40%. AUC(0-6h) values were larger after application of the submicron emulsions in comparison to aqueous solutions; however, statistically significant differences were only observed for emulsions at pH 6.5. Although the bioavailability of the drug is pH dependent, emulsions at higher pH cannot be considered for clinical use because of pilocarpine degradation which occurs with a similar rate as in aqueous solutions. Introduction of pilocarpine into the oily phase in the form of pilocarpine base or its oleate did not improve either the physicochemical or the pharmacological properties of the formulations. Irrespective of the pH and chemical form of pilocarpine used for emulsion preparation, practically all drug was found in the aqueous phase of the emulsion; thus, partitioning to the oily phase was negligible. PMID- 11253933 TI - Air-filled polymeric microcapsules from emulsions containing different organic phases. AB - Air-filled polymeric microcapsules for use as a contrast agent in ultrasonography have been prepared by the freeze-drying of different oil-in-water emulsions. The water phases consisted of a block copolymer in water. The organic phases consisted of a biodegradable polyester dissolved in (-)-camphene, cyclooctane, cyclohexane or tricyclene, which were relatively poor solvents for the polyester. A polymeric wall was, therefore, precipitated at the droplet surface early in the process, i.e. during freezing. Removing the solvent during freeze-drying, resulted in air-filled microcapsules. The microcapsules were suspended in saline after freeze-drying. All the suspensions contained echogenic microcapsules with a volume mean diameter of approximately 5-7 microm. Microscopic investigations showed that the microcapsules were spherical and hollow. Tricyclene and, to some degree, (-)-camphene were found unsuitable for industrial production due to melting points above 30 degrees C. Cyclooctane and cyclohexane were investigated as replacements for the initially chosen (-)-camphene, since they are liquids over a wider temperature range. These solvents gave improved yields, measured both as particle volume concentration per amount of polymer in suspension and acoustic attenuation at 3.5 MHz per amount of polymer in suspension, although the freeze-drying cycle was not optimized for these systems. PMID- 11253935 TI - Characterization of protein-loaded poly(epsilon-caprolactone) microparticles based on a factorial design. AB - This study was designed to systematically investigate the characteristics of bovine serum albumin (BSA) loaded poly(epsilon-caprolactine) (PCL) microparticles based on a 2(4) factorial experiment. The influences of polyvinyl pyrrolidone (PVP) concentration, BSA/PCL ratio, w/o/o/o ratio, and PEG/PCL ratio on the surface morphology, particle size, as well as the yield of microparticles, encapsulation efficiency of BSA, and in vitro release properties were evaluated. The microparticles were prepared by the w/o/o/o solvent evaporation method. The structure of BSA retained its integrity using this technique. The mean particle sizes of BSA-loded microparticles were in the range of 20-50 microm, and a highly porous morphology existed in these microparticles, irrespective of the formulations. The production yields of microparticles were in the range of 52.1 89.0%, and the encapsulation efficiencies were in the range of 13.8-68.3%. The burst release of BSA was in the range of 6.9-69.0%. The volume ration of the multi-phases significantly affected the encapsulation efficiency of BSA in PCL microparticles, and the initial amount of BSA encapsulated by PCL in terms of BSA/PCL ratio significantly affected the amount of BSA released at the end of 14 days (p < 0.05). PMID- 11253936 TI - Influence of pluronics on protein-loaded poly(epsilon-caprolactone) microparticles. AB - The objective of this study was designed to elucidate the importance of adding pluronics to PCL microparticles for the delivery of proteins. The influences of the type and the amount of pluronic on the surface morphology and release properties of protein-loaded PCL microparticles were evaluated. Microparticles were prepared by the w/o/o/o solvent evaporation technique with an ultrasonicator. All of the microparticles prepared were spherical in shape, with a rough surface due to crystallization of PCL in the microparticles. The pluronics efficiently prevented microparticles from aggregation, and the size of microparticles prepared was significantly reduced. The significant increase in the encapsulation efficiency and decrease in the burst release of protein from PCL microparticles were achieved by using pluronic F127. Incorporation of pluronic F127 increased the hydrophilicity of the matrix, which further retained protein in blended microparticles. Both pluronics PE10100 and L61 significantly reduced the crystallinity of PCL microparticles. Nevertheless, this result did not further influence the release property of BSA from these microparticles. PMID- 11253937 TI - Microencapsulation of ascorbic acid: effect of process variables on product characteristics. AB - This study deals with a comparative investigation of the characteristics of ascorbic acid microcapsules prepared by different methods, such as thermal phase separation, melt dispersion, solvent evaporation and spray drying. Scanning electron microscopy (SEM), release tests and size distribution were used for the evaluation of product characteristics. The results show that microencapsulated ascorbic acid could prevent the ascorbic acid colour change, retard its core release rate, and generally mask its acid taste. In the thermal phase separation, molecular weight (Mw) of ethyl cellulose (EC) and the addition of polyisobutylene (PIB) significantly influenced the aggregation and release rate of microcapsules. In the melt dispersion method, spherical particles were prepared by using carnauba. The ascorbic acid release rate was found to be slower in the case of carnauba-encapsulated ascorbic acid than that made by EC using other methods. In the solvent evaporation method, a higher Mw of EC and the addition of plastizer were also found to be important for good encapsulation. In the spray drying method, loss of ascorbic acid was found to be minimum during microencapsulation. Starch and beta-cyclodextrin encapsulated ascorbic acid delayed the degradation of ascorbic acid during storage at 38 degrees C and relative humidity 84.0%. PMID- 11253938 TI - Characterization of calcitonin-containing liposome formulations for intranasal delivery. AB - Calcitonin-containing liposome formulations were characterized to obtain information for evaluation of their feasibility in intranasal delivery. The parameters of liposomal charge characteristics, charge inducing agent concentration, calcitonin concentration and pH of the medium on the loading efficiency and leakage behaviour, and the chemical stability of calcitonin in liposomes were investigated. Results showed that the loading efficiency of calcitonin increased with increasing the added concentration of calcitonin. The magnitude of the loading efficiency due to the liposomal charge of negative, positive and neutral characteristics was in the order of negatively charged liposome > neutral liposome > positively charge liposome. The increase of molar ratio of phosphatidylserine in liposomes showed an increase of loading efficiency; while, the increase of molar ratios of stearylamine showed a decrease of loading efficiency. The loading efficiency at pH 7.4 was greater than that at pH 4.3. The leakage of positively charged liposomes was greater than that of neutral and negatively charged liposomes. The leakage at pH 4.3 was faster than that at pH 7.4. The leakage of positively charged liposomes increased as temperature increased. The chemical stability of calcitonin in both solution and liposomes demonstrated a pseudo-first-order kinetic degradation. Less degradation was observed at pH 3.4 and 4 degrees C. The degradation rate of calcitonin in solution, or in positively charged, negatively charged, and neutral liposomes, exhibited no significant difference. The particle size of the calcitonin containing liposomes after storage for 1 month at pH 4.3 and 4 degrees C showed little change. PMID- 11253939 TI - Encapsulating aspirin into a surfactant-free ethyl cellulose microsphere using non-toxic solvents by emulsion solvent-evaporation technique. AB - Microencapsulation of aspirin in ethylcellulose was studied in a surfactant-free, water-in-oil type of emulsification/solvent evaporation process using non-toxic solvents. Ethanol was used as the dispersed phase and soybean oil as the continuous phase. The recovered weight, particle size distribution, aspirin loading efficiency, and the aspirin release rate of microspheres were analysed. The addition of a small amount of non-solvent (water) prior to the emulsification was found to have a significant impact on the microencapsulation process. Adding non-solvent increases the recovered weight and the size of the microspheres. The addition of non-solvent also markedly changes the microsphere characteristics, resulting in a coarser surface and an increased release rate. Increasing the emulsification evaporation temperature increases the size of the microsphere, but reduces the recovered weight and loading efficiency. The release rate follows a first-order kinetics and Higuchi matrix kinetics at low concentrations of non solvent, suggesting a monolithic system with aspirin uniformly distributed in the microsphere. PMID- 11253940 TI - Chitosan/gelatin microspheres prepared by modified emulsification and ionotropic gelation. AB - Chitosan microspheres were prepared by an emulsion-phase separation technique without the use of chemical cross-linking agents; alternatively, ionotropic gelation was employed in a w/o emulsion. The possibility of three kinds of anions (tripolyphosphate, citrate and sulphate) to interact with chitosan was investigated by turbidimetric titration. The results indicate that there are electrostatic interactions between the above anions and chitosan in a certain region of solution pH (1.0-7.5 for sulphate/chitosan, 4.5-7.5 for citrate/chitosan and 1.9-7.5 for tripolyphosphate/chitosan), that is related to the natural characteristics of the anions. Out of the pH region where anions interact with chitosan, no microspheres were formed. However, even in the pH region where anions can interact with chitosan, only irregular microparticles were obtained in the case of the conventional emulsification and ionotropic gelation method, while spherical microspheres with diameters in the range of tens of microns were obtained when a modified process was employed. The key point of the modified process is the introduction of gelatin and allowing the ionic cross linking process of chitosan/gelatin w/o emulsions to take place under coagulation conditions at a low temperature. The surface of sodium sulphate cross-linked chitosan/gelatin and sodium citrate cross-linked chitosan/gelatin microspheres was very smooth, but large gaps were observed on the surface of tripolyphosphate/chitosan microspheres. The increase of stirring speed led to a decrease in diameter and a narrowing in size distribution. PMID- 11253942 TI - Literature alerts. PMID- 11253943 TI - Principles and applications of the polymerase chain reaction. AB - The polymerase chain reaction (PCR), a primer-directed in vitro enzymatic reaction for the production of a specific DNA fragment, has dramatically facilitated molecular biology approaches to fundamental research questions, as well introduced DNA testing into a variety of clinical diagnostic areas. Some of the principles of PCR, several recent advances in PCR technology and selected applications are reviewed. A variety of methods for analyzing allelic sequence diversity in amplified DNA have been developed. Genetic testing of the HLA loci, and other genes encoding cytokines and cytokine receptors, has had a significant impact on disease susceptibility studies and in clinical transplantation. PMID- 11253944 TI - History of DNA typing for the human MHC. AB - The last twenty years have seen an exponential growth in the application of DNA technology to the field of Histocompatibility and Immunogenetics (H&I). Initially, this was confined to a few research laboratories. However, the development and application of several different DNA methods by many laboratories has led to the situation whereby nearly every H&I laboratory performs some DNA typing for the detection of HLA alleles. It would not be unfair to say that H&I has shown diagnostic laboratories in other disciplines how useful the DNA techniques can be. This review attempts to summarise the history and application of DNA methods in the field of Histocompatibility (Table 1). PMID- 11253941 TI - Preparation and characterization of hCG-loaded polylactide or poly(lactide-co glycolide) microspheres using a modified water-in-oil-in-water (w/o/w) emulsion solvent evaporation technique. AB - A modified w/o/w emulsion solvent evaporation technique was adopted to prepare human Chorionic Gonadotropin (hCG)-loaded polylactide (PLA) or poly(lactide-co glycolide) (PLGA) microspheres. The effects of preparative parameters, such as stirring rate, polymer MW and concentration, and the composition of both the inner aqueous phase and oil phase etc., on hCG entrapment efficiency and microsphere characteristics were investigated. It was found that by adding 20% glycerol into the inner aqueous phase and 40% acetone into the oil phase, smooth microspheres approximately 1 microm in diameter could be produced with high hCG entrapment efficiency (>90%). In vitro release test showed a burst release of hCG from PLGA (75:25) microspheres, followed by sustained release of 55% hCG over 2 months. The initial hCG burst from PLGA microspheres increased with the glycerol concentration in the inner aqueous phase, but decreased to a low value (ca. 20%) with the addition of acetone into the oil phase, which could be attributed to the associated changes in surface morphology of the microspheres. In vivo experiments demonstrated that a single shot of hCG-loaded PLGA microspheres could produce a comparable antibody response with the inoculation of free hCG four times. PMID- 11253945 TI - Molecular typing for the MHC with PCR-SSP. AB - Sequence-specific amplification (SSP) is simply a form of polymerase chain reaction (PCR) which involves designing one or both primers so that they will or will not allow amplification (the 3'-mismatch principle). Its origins are probably legion, i.e. many people probably thought of it at the same time. For example, in 1988 a group from Guy's Hospital, London, described a form of SSP for HLA-DR4 detection and in the same year a group from Upjohn described its use at the American Society of Histocompatibility and Immunogenetics (ASHI). Both are published in abstract form (British Society of Rheumatology and ASHI). The 3' mismatch principle can be used to identify virtually any single nucleotide point mutation (SNP) within one or two PCR-SSP reactions and the first peer-reviewed statements of this came in 1989 (1, 2). Thus, although the use of SSP probably began around 1990, it was 5 years before its popularity erupted, mainly due to the work of Olerup & Zetterquist (3, 4), who defined its potential for solid organ transplantation. It is now the method of choice for high resolution HLA typing in many laboratories. In addition, over a thousand applications for genes outside the MHC are in the literature. PMID- 11253946 TI - High and intermediate resolution DNA typing systems for class I HLA-A, B, C genes by hybridization with sequence-specific oligonucleotide probes (SSOP). AB - DNA typing systems for alleles of the HLA class I loci A, B, C at intermediate (IR) and high resolution (HR) levels were developed. The approaches combine locus specific amplification of genomic DNA by the polymerase chain reaction (PCR) and hybridization with sequence-specific oligonucleotide probes (SSOP). The SSOP were designed to match nucleotide sequences at all polymorphic sites of exons 2 and 3 at these loci. Alleles and genotypes for these loci are assigned by their unique hybridization patterns. Some genotypes with particular allele combinations resulted in the same hybridization pattern. These genotype ambiguities were resolved by performing additional group-specific amplifications with appropriate GSA primers and hybridization with informative SSOP. At intermediate resolution level, many groups of alleles of HLA-A and B with the same serologic equivalence resulted in the same hybridization pattern. Both HR and IR typing approaches required the design, validation and testing of locus- and group-specific primers and sequence-specific oligonucleotide probes (SSOP). Single locus-specific amplification and hybridization with sets of 67 SSOP for HLA-A, 99 for HLA-B and 57 for HLA-C allowed us to identify unequivocally the majority of A, B, C alleles at HR level. To resolve ambiguous genotypes at HLA-B, we performed 4 GSA with 5' primers at codon 45-46 and hybridization with selected sets of SSOP. About 22,415 high resolution typing results were obtained (4,953A, 6,621B, 10,841C). In these samples, 63 HLA-A, 170 HLA-B and 40 HLA-C alleles were observed. In the course of these studies, more than 30 new alleles have been identified. In IR testing, sets of 39 SSOP for HLA-A typing and 59 SSOP for HLA-B typing allowed us to obtain maximal resolution of the majority of common genotypes. To achieve IR level, the majority of SSOP selected were those that span codons encoding amino acid residues located in the alpha helical segments of the class I molecules. A total of 50,522 samples were typed for HLA-A and B at IR level. Approximately 2.0% of them carried ambiguous genotypes associated with alternative switches of Bw4/w6 related sequences. All these ambiguities were resolved by Bw4/Bw6 GSA by 2 primer pairs (77N-IALR-83/3B.1; 77S-DLRG-83/3B.1) and hybridization with 9 selected SSOP Testing was performed by individual hybridization of replicate dot blot membranes with the SSOP of the corresponding set. The approach was robust and cost effective in large-scale HLA class I molecular typing. The resolution provided by HLA-A, B IR reached serologic split-level or higher. The description of primers and probes for HLA class I typing may be utilized as starting elements for development of second generation methods with a more rapid turn around time. PMID- 11253947 TI - HLA typing by Reference Strand Mediated Conformation Analysis (RSCA). AB - Reference Strand Mediated Conformation Analysis (RSCA) is a novel conformational method that offers high resolution and high sample throughput for typing the HLA class I and class II genes. This technique differs from conventional sequence based typing methodologies in that the HLA type is assigned on the basis of accurate measurement of conformation-dependent DNA mobility in polyacrylamide gel electrophoresis (AGE). RSCA utilises a fluorescent-labelled locus-specific reference DNA to selectively modify the molecular conformation of the tested DNA. The use of laser-based instrumentation and computer software, in addition to internal DNA markers for correction of gel variability, allows the discrimination of HLA alleles which differ by one nucleotide in a DNA fragment nearly as large as a kilobase in length. RSCA has been successfully applied in blind studies of HLA typing, demonstrating that is reproducible, able to identify new alleles and able to resolve ambiguous heterozygous combinations. PMID- 11253948 TI - Oligonucleotide arrays for high resolution HLA typing. AB - The methodology and applications of oligonucleotide array technology are reviewed. An oligonucleotide array system for typing HLA-B alleles is described. Oligonucleotide probes representing all known polymorphisms in exons 2 and 3 of HLA-B are immobilized on glass slides. Exons 2 and 3 are amplified from human genomic DNA using fluorescent-labeled primers and subsequently allowed to hybridize to an oligonucleotide array on glass supports. The presence of positive hybridization is detected by fluorescence scanning. HLA-B alleles are assigned by quantitative analysis of hybridization patterns. Proof of principle has been established in blinded testing of heterozygous DNA samples. PMID- 11253949 TI - Mapping HLA for single nucleotide polymorphisms. AB - Knowledge of DNA sequence variation may help us understand how genetic variability gives rise to functional variability and, in so doing, revolutionize the development of strategies to combat and prevent disease. Single nucleotide polymorphisms (SNPs) are stable, inherited, biallelic, single base pair differences which are present in the human genome at a density of 1 to 10 per 1,000 nucleotides. It is anticipated that SNPs will account for much of the functional heterogeneity in gene expression and protein activity exhibited in the human population. Susceptibility to or protection from a number of diseases, particularly those of autoimmune etiology, has been associated with specific alleles of the human leukocyte antigen (HLA) complex. Interestingly, the precise molecular defects in the HLA genes are unknown and the notion that non-HLA genes, within the same chromosomal region, are involved remains a formal possibility. We have determined the nucleotide sequence of a contiguous 2.2 Mbp segment of chromosome six that includes all of the HLA class I region, and have identified over 10,000 SNPs therein. Because of the derivative knowledge of gene and SNP content and position, the scientific community is now uniquely poised to identify disease-contributory SNPs that lie within the MHC. PMID- 11253950 TI - Typing for human platelet alloantigens. AB - Antibodies to platelet alloantigens, and sometimes to isoantigens, induce severe clinical problems such as neonatal alloimmune thrombocytopenia (NAIT), post transfusion purpura (PTP) and refractoriness to platelet transfusions (PTR). For example, NAIT affects approximately 1 in 5,000 live births. It is essential, therefore, to screen pregnant women for platelet antibodies in order to save babies' lives. Almost 40 years ago, two platelet alloantigen systems were discovered using relatively simple methods, namely the platelet agglutination test and the complement fixation test. However, these methods were not sensitive enough to identify all antibodies in mothers and patients, even in those with severe clinical problems. Tremendous effort has been devoted to establish more sensitive and reliable methods. In recent years, excellent new serological and immunochemical methods have been established and several new platelet antigen systems have been discovered. Simultaneously, newly developed molecular genetic techniques have been introduced for the typing and analysis of human platelet alloantigen systems. These methods allow DNA typing for cases in which serological typing is not available. In this article, the history of studies on human platelet alloantigen systems and isoantigens, the nomenclature of platelet alloantigen systems and their alleles, the present status of antibody detection and typing techniques and, finally, ethnic variations in platelet antigen profiles are reviewed. PMID- 11253951 TI - Analysis of amplified variable number tandem repeat loci for evaluation of engraftment after hematopoietic stem cell transplantation. AB - Many methods have been used to evaluate engraftment of donor cells after hematopoietic stem cell transplantation in allogeneic recipients. The identification of variable number tandem repeat loci in the human genome and the development of methods for testing amplified DNA segments containing these polymorphic loci have provided a novel and highly convenient approach for assessing the success of hematopoietic stem cell transplantation. We discuss the advantages and limitations of this approach as it has been implemented in our laboratory. We summarize the reasons for testing genetic markers after hematopoietic stem cell transplantation, using illustrations from our own experience. This information is designed to help disseminate similar methods in laboratories that do not yet provide such services, to assist laboratory personnel in understanding the clinical and research needs of physicians who request such services and to assist clinicians who interpret the results of genetic marker studies. PMID- 11253952 TI - The use of PCR technology for detecting minimal residual disease in patients with leukemia. AB - The study of minimal residual disease (MRD) aims to understand the biology and clinical significance of leukemia that persists in patients who are in complete pathologic remission. The detection of MRD most consistently has been associated with subsequent relapse in childhood acute lymphoblastic leukemia (ALL), t(15;17) acute myeloid leukemia (AML) and chronic myeloid leukemia (CML) following marrow transplantation. However, in several settings, MRD has been detected in patients enjoying long-term remission. The study of MRD is thus evolving from identifying patients at high risk of relapse to explaining how leukemia can persist for years in an otherwise "cured" patient. PMID- 11253953 TI - Cognitive impairment in schizophrenia is the core of the disorder. AB - Patients with schizophrenia exhibit an exceedingly wide range of symptoms from a variety of domains. The cardinal features are abnormal ideas (such as delusions); abnormal perceptions (such as hallucinations); formal thought disorder (as evidenced by disorganized speech); motor, volitional, and behavioral disorders; and emotional disorders (such as affective flattening or inappropriateness). In addition to these diverse, and sometimes bizarre symptoms, it has become increasingly apparent that the disorder is, to variable degrees, accompanied by a broad spectrum of cognitive impairments. This review addresses the question of whether the cognitive deficits seen in schizophrenic patients are the core features of the disorder. In other words, we explore whether schizophrenia is best characterized by symptoms or cognitive deficits (we suggest the latter) and moreover, whether there is a specific cognitive deficit profile that may assist in diagnosis. First, we discuss what the cognitive deficits are. Then we address in turn the reality, frequency, predictive validity, specificity, course and susceptibility to neuroleptic effects of these cognitive impairments. In brief, we argue that various cognitive deficits are enduring features of the schizophrenia illness, that they are not state-related and are not specific to subtypes of the illness, and, more specifically, that working memory and attention are characteristically impaired in patients with schizophrenia, irrespective of their level of intelligence. Last, we conclude that problems in these cognitive domains are at the very core of the dysfunction in this disease. PMID- 11253954 TI - Impairment of GABAergic transmission in depression: new insights from neuroimaging studies. AB - Several lines of evidence suggest that abnormalities in GABAergic neurotransmission are associated with the neurobiology of depression. Animal studies demonstrate that GABA agonists and antagonists can modulate commonly used behavioral models of depression and that chronic administration of antidepressant drugs induce marked changes in GABAergic function. In humans, depressed patients have lower plasma and CSF GABA concentrations than nondepressed comparison subjects. The recent discovery that several anticonvulsant and GABA-mimetic agents possess mood stabilizing and antidepressant properties has further increased interest in these findings. Novel imaging techniques now allow investigation of the GABAergic contribution to affective disorder pathophysiology. Through the techniques of PET, SPECT, and MRS, GABAergic function can be evaluated in vivo. Preliminary studies employing these techniques are finding new evidence suggesting that GABAergic abnormalities are associated with stress, anxiety, and depression. This article reviews the existing literature investigating the possible involvement of GABA in the neurobiology of depression and briefly highlights how these novel neuroimaging techniques can be used to further assess this hypothesis. PMID- 11253955 TI - Herpes simplex virus-mediated gene transfer as a tool for neuropsychiatric research. AB - There is an enormous initiative to establish causal relationships between brain biology (including patterns of gene expression) and behavior. Unfortunately, genetic intervention is not accomplished easily in the brain. One strategy is to engineer and deliver to the brain specialized viral vectors that carry a gene (or genes) of interest, thereby exploiting the natural ability of viruses to insert genetic information into cells. When delivered to the brain, these vectors cause infected cells to increase expression of the genes of interest. Viral vectors are particularly useful when the goal is to manipulate expression of a single gene in a specific brain region, at a specific time, and in animals that developed normally. There are several types of virus that can be adapted for use as viral vectors, including those based on herpes simplex virus (HSV-1), adenovirus (AV), adeno-associated virus (AAV), and lentivirus. Although each vector has its own unique advantages and disadvantages, this rapidly evolving technology has the potential to revolutionize neuropsychiatric research by offering the opportunity to establish, with anatomical and temporal specificity, causal relations between altered expression of individual gene products and alterations in complex behavior. PMID- 11253957 TI - Sidestepping the Ottawa Mine Ban Treaty. PMID- 11253956 TI - Ethanol and NMDA receptor signaling. AB - NMDA receptors are a multi-subunit family of ionotropic receptors activated by the neurotransmitter glutamate. Localized primarily postsynaptically in neurons, they play an important role in mediating excitatory synaptic neurotransmission and are implicated in a wide variety of important calcium-dependent neuronal processes. Experimental animals expressing mutant forms of NMDA subunits display abnormal behaviors and locomotor and cognitive impairments. Over the last 10 years, a wealth of studies has indicated that NMDA receptors are an important site of action for ethanol in the brain. The effects of acute ethanol on NMDA receptor function is discussed herein, with particular focus on efforts to define a molecular site of action of ethanol on the receptor. While it is clear that the ethanol sensitivity of NMDA receptors is influenced by subunit composition, it is also apparent that posttranslational factors such as phosphorylation and protein protein interactions are important in modulating this sensitivity. PMID- 11253958 TI - Complexity of issue of dietary trans fatty acids. PMID- 11253959 TI - Examination of context of medicine. PMID- 11253960 TI - Transcription factors and muscle cachexia: is there a therapeutic target? PMID- 11253962 TI - Starting the day the right way. PMID- 11253961 TI - Is there a role for follicle-stimulating-hormone receptor in familial dizygotic twinning? PMID- 11253963 TI - Rape as a war crime: putting policy into practice. PMID- 11253964 TI - Foot-and-mouth epidemic: the choices. PMID- 11253965 TI - Help by and for medical editors. PMID- 11253967 TI - Association between trans fatty acid intake and 10-year risk of coronary heart disease in the Zutphen Elderly Study: a prospective population-based study. AB - BACKGROUND: Evidence on the relation between trans fatty acid intake and coronary heart disease is limited. We investigated this relation in a Dutch population with a fairly high trans fatty acid intake, including trans fatty acids from partly hydrogenated fish oils. METHODS: We prospectively studied 667 men of the Zutphen Elderly Study aged 64-84 years and free of coronary heart disease at baseline. We used dietary surveys to establish the participants' food consumption patterns. Information on risk factors and diet was obtained in 1985, 1990, and 1995. After 10 years of follow-up from 1985-95, there were 98 cases of fatal or non-fatal coronary heart disease. FINDINGS: Between 1985 and 1995, average trans fatty acid intake decreased from 4.3% to 1.9% of energy. After adjustment for age, body mass index, smoking, and dietary covariates, trans fatty acid intake at baseline was positively associated with the 10-year risk of coronary heart disease. The relative risk for a difference of 2% of energy in trans fatty acid intake at baseline was 1.28 (95% CI 1.01-1.61). INTERPRETATION: A high intake of trans fatty acids (all types of isomers) contributes to the risk of coronary heart disease. The substantial decrease in trans fatty acid intake, mainly due to industrial lowering of trans contents in Dutch edible fats, could therefore have had a large public-health impact. PMID- 11253966 TI - Comparison of two standard chemotherapy regimens for good-prognosis germ-cell tumours: a randomised trial. Australian and New Zealand Germ Cell Trial Group. AB - BACKGROUND: Most patients with metastatic germ-cell tumours are cured with chemotherapy. However, the optimum chemotherapy regimen is uncertain, and there is variation in international practice. We did a multicentre randomised trial to compare two standard chemotherapy regimens for men with good-prognosis germ-cell tumours. METHODS: Good prognosis was defined by modified Memorial Sloan-Kettering criteria. The first regimen (regimen A) was based on treatment recommendations from Indiana University and comprised three cycles of 20 mg/m2 cisplatin on days 1-5, 100 mg/m2 etoposide on days 1-5, and 30 kU bleomycin on days 1, 8, and 15, repeated every 21 days. The second regimen (regimen B) was based on the control regimen of a published randomised clinical trial and comprised four cycles of 100 mg/m2 cisplatin on day 1, 120 mg/m2 etoposide on days 1-3, and 30 kU bleomycin on day 1, repeated every 21 days. The primary outcome measure was overall survival. Analysis was by intention to treat. FINDINGS: 166 patients were randomised, 83 to each regimen. The trial was stopped when the second planned interim analysis met predefined stopping rules. The median follow-up was 33 months. Overall survival was substantially better with regimen A (three vs 13 deaths, hazard ratio 0.22 [95% CI 0.06-0.77], p=0.008). This difference was due to deaths from cancer (one vs nine), and not deaths from treatment (two vs two) and remained significant after adjustment for other prognostic factors (0.25 [0.07-0.88], p=0.03). INTERPRETATION: In men with good-prognosis germ-cell tumours, the regimen developed at Indiana University is superior to the alternative regimen studied in this trial. The lower total dose and dose-intensity of bleomycin and the lower dose-intensity of etoposide in regimen B could be responsible for the worse outcome. PMID- 11253968 TI - Clinical picture: bronchiolitis obliterans with organising pneumonia during interferon beta-1a treatment. PMID- 11253970 TI - Influence of context effects on health outcomes: a systematic review. AB - BACKGROUND: Throughout history, doctor-patient relationships have been acknowledged as having an important therapeutic effect, irrespective of any prescribed drug or treatment. We did a systematic review to determine whether there was any empirical evidence to support this theory. METHODS: A comprehensive search strategy was developed to include 11 medical, psychological, and sociological electronic databases. The quality of eligible trials was objectively assessed by two reviewers, and the type of non-treatment care given in each trial was categorised as cognitive or emotional. Cognitive care aims to influence patients' expectations about the illness or the treatment, whereas emotional care refers to the style of the consultation (eg, warm, empathic), and aims to reduce negative feelings such as anxiety and fear. FINDINGS: We identified 25 eligible randomised controlled trials. 19 examined the effects of influencing patients' expectations about treatment, half of which found significant effects. None of the studies examined the effects of emotional care alone, but four trials assessed a combination of both cognitive and emotional care. Three of these studies showed that enhancing patients' expectations through positive information about the treatment or the illness, while providing support or reassurance, significantly influenced health outcomes. INTERPRETATION: There is much inconsistency regarding emotional and cognitive care, although one relatively consistent finding is that physicians who adopt a warm, friendly, and reassuring manner are more effective than those who keep consultations formal and do not offer reassurance. PMID- 11253969 TI - Sensitisation, asthma, and a modified Th2 response in children exposed to cat allergen: a population-based cross-sectional study. AB - BACKGROUND: Although asthma is strongly associated with immediate hypersensitivity to indoor allergens, several studies have suggested that a cat in the house can decrease the risk of asthma. We investigated the immune response to cat and mite allergens, and asthma among children with a wide range of allergen exposure. METHODS: We did a population-based cross-sectional study of children (aged 12-14 years), some of whom had symptoms of asthma and bronchial hyper-reactivity. Antibodies to mite (Der f 1) and cat (Fel d 1) allergens measured by isotype (IgG and IgG4) specific radioimmunoprecipitation assays were compared with sensitisation and allergen concentrations in house dust. FINDINGS: 226 children were recruited, 47 of whom had symptoms of asthma and bronchial hyper-reactivity. Increasing exposure to mite was associated with increased prevalence of sensitisation and IgG antibody to Der f 1. By contrast, the highest exposure to cat was associated with decreased sensitisation, but a higher prevalence of IgG antibody to Fel d 1. Thus, among children with high exposure, the odds of sensitisation to mite rather than cat was 4.0 (99% CI 1.49-10.00). Furthermore, 31 of 76 children with 23 microg Fel d 1 at home, who were not sensitised to cat allergen had >125 units of IgG antibody to Fel d 1. Antibodies to Fel d 1 of the IgG4 isotype were strongly correlated with IgG antibody in both allergic and non-allergic children (r=0.84 and r=0.66, respectively). Sensitisation to mite or cat allergens was the strongest independent risk factor for asthma (p<0.001). INTERPRETATION: Exposure to cat allergen can produce an IgG and IgG4 antibody response without sensitisation or risk of asthma. This modified T-helper-2 cell response should be regarded as a form of tolerance and may be the correct objective of immunotherapy. The results may also explain the observation that animals in the house can decrease the risk of asthma. PMID- 11253971 TI - Effect of alcohol consumption on systemic markers of inflammation. AB - BACKGROUND: Epidemiological studies suggest that light to moderate alcohol intake is associated with lower all-cause mortality than abstention or heavy alcohol intake, primarily through reduced risk of coronary heart disease. The underlying mechanisms are incompletely understood. METHODS: We investigated the association between alcohol consumption (assessed by a 7-day food record) and concentrations of C-reactive protein (CRP), alpha1-globulins, alpha2-globulins, albumin, and transferrin, and leucocyte count in a sample of 2006 men and women aged 18-88 years participating in a national health survey carried out in former West Germany in 1987-88. Analyses were based on 781 men and 995 women with complete data. FINDINGS: Among men, alcohol consumption showed a U-shaped association with mean values of CRP (p for linear term 0.65, for quadratic term 0.048), alpha1 globulins (p=0.20, 0.0006), alpha2-globulins (p=0.82, 0.31), and leucocyte count (p=0.51, 0.26) even after adjustment for age, smoking, body-mass index, HDL and LDL cholesterol, history of hypertension, education, and income. There were inverted U-shaped associations between the negative acute-phase reactants albumin (p=0.41, 0.006) and transferrin (p=0.14, 0.28) and alcohol intake. In women, the associations were less strong for CRP (p=0.35, 0.31), leucocyte count (p=0.28, 0.15), and transferrin (p=0.86, 0.83). Concentrations of alpha1-globulins and alpha2-globulins were inversely related to alcohol consumption, and albumin showed a positive association with increasing alcohol intake in women. INTERPRETATION: Non-drinkers and heavy drinkers had higher CRP concentrations than moderate drinkers. In view of the robust association between markers of inflammation, especially CRP, and risk of coronary heart disease, an anti inflammatory action of alcohol could contribute to the link between moderate consumption and lower cardiovascular mortality. PMID- 11253972 TI - Ascending cellulitis after an insect bite. PMID- 11253973 TI - Diagnosis of rotator cuff tears. AB - Rotator cuff tears account for almost 50% of major shoulder injuries but are sometimes difficult to diagnose. To aid diagnosis, we did a prospective study, comparing results of 23 clinical tests from 400 patients with and without rotator cuff tears. Three simple tests were predictive for rotator cuff tear: supraspinatus weakness, weakness in external rotation, and impingement. When all three were positive, or if two tests were positive and the patient was aged 60 or older, the individual had a 98% chance of having a rotator cuff tear; combined absence of these features excluded this diagnosis. PMID- 11253974 TI - Towards the eradication of hookworm in an isolated Australian community. AB - Hookworm (Ancylostoma duodenale) and other enteric parasites such as Giardia and Hymenolepis are common in Aboriginal communities in northem Australia, and their presence is associated with iron deficiency, anaemia, and failure to thrive. We report the outcome of a sustained, community-based control programme that used regular albendazole in one isolated community. Whereas hookworm has been effectively controlled by the programme, no sustained effect on the prevalence of Giardia and Hymenolepis was seen; the control of these parasites will depend on improvements in health education. This programme might serve as a model for community-based or population-based drug treatment programmes elsewhere. PMID- 11253975 TI - Mild head injury, anticoagulants, and risk of intracranial injury. AB - We studied intracranial damage in patients with mild head injuries who were taking warfarin. Of the 215,785 individuals who visited the Mount Auburn and Beth Israel accident and emergency departments during our study, we identified records for 144 patients by anticoagulation status and computed tomography (CT) imaging. We retrospectively reviewed these patients and ten (7%, 95% CI 3-11) with clinically important injuries on cranial CT. Our findings suggest that patients with head injuries who receive anticoagulants have a similar or greater risk of intracranlal injury to those falling into a previously defined moderate-risk category, invalidating a previous conclusion that CT scanning is unnecessary in such patients. PMID- 11253976 TI - Mutations in the follicle-stimulating hormone receptor and familial dizygotic twinning. AB - Concentrations of follicle-stimulating hormone (FSH) have an important role in multiple ovulation. An association has been reported between mutations in the FSH receptor (FSHR) in a family with increased twinning frequency. We sequenced the transmembrane region of FSHR (located on chromosome 2) in 21 unrelated mothers of dizygotic twins and found no differences to the published sequence. A linkage study of 183 sister pairs and trios, in which all sisters had given birth to spontaneous dizygotic twins, excluded linkage to this region of chromosome 2. We conclude that mutations in FSHR are not a common cause of familial dizygotic twinning. PMID- 11253977 TI - Velopharyngeal incompetence and chromosome 22q11 deletion. AB - Chromosome 22q11 deletion gives rise to various phenotypes, including cardiac malformations, velopharyngeal abnormalities, absent thymus, and neurological defects. We assessed, in a prospective study, chromosome 22q11 deletion in 50 of 144 patients with velopharyngeal incompetence in the absence of overt clefting. 18 (12.5% of the whole cohort and 36% of patients tested for the deletion) had the 22q11 deletion. This frequency differs from an estimated population prevalence of 0.025% and suggests a need for screening for the 22q11 deletion in these patients. PMID- 11253978 TI - Drug firms take South Africa's government to court. PMID- 11253979 TI - Macrodoctor, come meet the nanodoctors. PMID- 11253980 TI - Bush boosts research funds despite general austerity. PMID- 11253981 TI - UK consultant censured for failure to act on junior's research fraud. PMID- 11253982 TI - Fragmented US health-care system needs major reform. PMID- 11253983 TI - UK government focuses resources to reduce health inequalities. PMID- 11253984 TI - Cushing's syndrome. AB - During the past 30 years, there have been advances in understanding of the pathogenesis of Cushing's syndrome and in differential diagnosis of its various forms. Improved diagnostic tests and procedures have increased the ability to recognise even mild hypercortisolism and have provided the means to obtain an accurate diagnosis. Despite these advances, the occurrence of unusual clinical presentations and laboratory shortcomings may produce diagnostic problems and challenge clinical intuition. This article reviews recent pathogenic views, new tests, and new diagnostic problems in the evaluation of Cushing's syndrome. Atypical clinical presentations of hypercortisolism and some laboratory shortcomings that may confuse the diagnosis of Cushing's syndrome are also reported. PMID- 11253985 TI - The uses of error: iatrogenic hepatitis. PMID- 11253986 TI - Communication and information technology in medical education. AB - The past few years have seen rapid advances in communication and information technology (C&IT), and the pervasion of the worldwide web into everyday life has important implications for education. Most medical schools provide extensive computer networks for their students, and these are increasingly becoming a central component of the learning and teaching environment. Such advances bring new opportunities and challenges to medical education, and are having an impact on the way that we teach and on the way that students learn, and on the very design and delivery of the curriculum. The plethora of information available on the web is overwhelming, and both students and staff need to be taught how to manage it effectively. Medical schools must develop clear strategies to address the issues raised by these technologies. We describe how medical schools are rising to this challenge, look at some of the ways in which communication and information technology can be used to enhance the learning and teaching environment, and discuss the potential impact of future developments on medical education. PMID- 11253987 TI - Chagas' disease. PMID- 11253988 TI - Drug use during pregnancy. PMID- 11253989 TI - Drug use during pregnancy. PMID- 11253990 TI - Levonorgestrel-releasing intrauterine devices. PMID- 11253991 TI - Levonorgestrel-releasing intrauterine devices. PMID- 11253992 TI - Liposomal amphotericin B. PMID- 11253993 TI - CCR5-delta32 polymorphism in asthma. PMID- 11253994 TI - Complementary and alternative medicine. PMID- 11253995 TI - Complementary and alternative medicine. PMID- 11253996 TI - Complementary and alternative medicine. PMID- 11253997 TI - Ciprofloxacin resistance in gonococci. PMID- 11253998 TI - Onset of coeliac disease and interferon treatment. PMID- 11253999 TI - Fluticasone and asthma. PMID- 11254001 TI - Discomfort with fine-needle aspiration cytology of the breast. PMID- 11254000 TI - Discomfort with fine-needle aspiration cytology of the breast. PMID- 11254002 TI - Holmes-Adie syndrome and Lyme disease. PMID- 11254003 TI - Eyelid movement disorders and electromyography. PMID- 11254004 TI - Yoga and chemoreflex sensitivity. PMID- 11254005 TI - Relapse of systemic lupus erythematosus. PMID- 11254006 TI - Troponins in prediction of cardiotoxic effects. PMID- 11254007 TI - Prescribing in the UK. PMID- 11254009 TI - Has the world forgotten Bhopal? PMID- 11254008 TI - Emergency contraception in Chile. PMID- 11254010 TI - Has the world forgotten Bhopal? PMID- 11254011 TI - Has the world forgotten Bhopal? PMID- 11254013 TI - Big bang--black hole? PMID- 11254012 TI - Intellectual property. PMID- 11254014 TI - Diabetes on the rise worldwide and webwide. PMID- 11254015 TI - The diagnostic boundaries of bipolar disorder. PMID- 11254016 TI - Abnormalities of cAMP signaling in affective disorders: implication for pathophysiology and treatment. AB - OBJECTIVE: During the last decade, much attention has been given to the role of signal transduction pathways in affective disorders. This review describes the possible role of the cAMP signaling in such disorders. METHODS: Among the components of cAMP signaling, this review focuses on the cAMP-dependent phosphorylation system. We analyzed the basic components of the cAMP-dependent phosphorylation system and the preclinical evidence supporting their involvement in the biochemical action of antidepressants and mood stabilizers. The clinical data available until now, concerning the possible link between the cAMP-dependent phosphorylation system and the pathophysiology of affective disorders, are also reviewed. RESULTS: The studies herein presented demonstrated that the levels and the activity of cAMP-dependent protein kinase are altered by antidepressants and mood stabilizers. Furthermore. these medications are able to modify the phosphorylation state, as well as the levels of some of the cAMP-dependent protein kinase substrates. More recently, clinical studies have reported abnormalities in the cAMP-dependent phosphorylation system in both peripheral cells and the postmortem brain of patients with affective disorders. CONCLUSIONS: Overall, these studies support an involvement of cAMP signaling in affective disorders. The precise knowledge of the findings has the potential to improve the understanding of pharmacotherapy and to provide directions for the development of novel biochemical and genetic research strategies on the pathogenesis of affective disorders. PMID- 11254017 TI - A plea for integrity of the bipolar disorder concept. PMID- 11254018 TI - Valproate treatment and the risk of hyperandrogenism and polycystic ovaries. AB - Long-term administration of valproate to women with epilepsy has been suggested to result in increased risk of hyperandrogenism and polycystic ovaries. In preliminary reports involving patients treated for several years, the reported rates were as high as 43% for polycystic ovaries and 17% for hyperandrogenism. In particular, when therapy started before the age of 20 years, the rates of either one of these complications were as high as 80%. Surprisingly, these reports have been relatively ignored in the psychiatric literature to date. As increasing numbers of bipolar patients are in long-term treatment with valproate, there is an important need for further research that clarifies the relationship between long-term administration of valproate and other mood stabilizers and the potential development of reproductive endocrinologic abnormalities, and for increased awareness among clinicians and patients of the unknown potential for these worrisome side-effects. PMID- 11254019 TI - Pharmacologic loading in the treatment of acute mania. AB - The rapid and safe reduction of manic symptoms is an important initial goal of the pharmacologic treatment of acute mania. The pharmacokinetics and studies of pharmacologic loading of lithium, valproate, and carbamazepine were reviewed. In addition, the feasibility of administering other agents with potential efficacy in mania, e.g., atypical antipsychotics and new anticonvulsants, was discussed. Further double-blind, controlled studies with adequate sample sizes comparing loading strategies with more gradual titration schedules of candidate antimanic agents are needed. PMID- 11254020 TI - Inositol as an add-on treatment for bipolar depression. AB - OBJECTIVE: Inositol is a constituent of the intracellular phosphatidyl inositol (PI) second messenger system, which is linked to various neurotransmitter receptors. Inositol crosses the blood-brain barrier in pharmacological doses, and has shown efficacy in a small double-blind study of unipolar depression. This pilot study evaluated its potential efficacy and safety in bipolar depression. METHODS: Twenty-four consenting adult men and women with DSM-IV bipolar depression (bipolar I = 21; bipolar II = 3) were randomly assigned to receive either 12 g of inositol or D-glucose as placebo for 6 weeks. Efficacy and safety ratings were done weekly. Thymoleptic medications (lithium, valproate, carbamazepine) in stable doses and at therapeutic levels at study entry were continued unchanged. RESULTS: Two subjects receiving placebo dropped out early due to worsening or non-adherence to the protocol. Among the 22 subjects who completed the trial, six (50%) of the inositol-treated subjects responded with a 50% or greater decrease in the baseline Hamilton Depression Rating Scale (HAM-D) score and a Clinical Global Improvement (CGI) scale score change of 'much' or 'very much' improved, as compared to three (30%) subjects assigned to placebo, a statistically nonsignificant difference. On the Montgomery-Asberg Depression Rating Scale (MADRS), eight (67%) of twelve inositol-treated subjects had a 50% or greater decrease in the baseline MADRS scores compared to four (33%) of twelve subjects assigned to placebo (p = 0.10). Inositol was well tolerated with minimal side effects, and thymoleptic blood levels were unaltered. CONCLUSIONS: These pilot data suggest a controlled study with an adequate sample size, and the appropriate rating scale may demonstrate efficacy for inositol in bipolar depression. The tolerability and the 'natural substance' aspect of inositol may be particularly appealing to subjects with bipolar depression. PMID- 11254022 TI - Cholesterol levels in mood disorders: high or low? AB - OBJECTIVES: To assess cholesterol levels in patients with mood disorders. METHODS: All consecutively admitted patients meeting inclusion criteria (n = 50) who were hospitalized in an affective disorders unit received assessments of cholesterol levels. Correlations were made with diagnosis using DSM-IV criteria, current mood states, and other clinical and demographic features of illness. Exclusion criteria included current alcohol abuse, medical illnesses that could influence cholesterol levels, eating disorders, and age greater than 70 years. RESULTS: Cholesterol levels did not differ based on diagnostic status of unipolar depression or bipolar disorder. In the total sample, cholesterol levels were lower in patients with current manic (170.2 +/- 38.9, p = 0.05) and depressive (182.0 +/- 42.0) than in mixed (226.4 +/- 43.3) episodes (p = 0.05). In subgroups of patients with bipolar disorder, manic episodes (169.9 +/- 38.8, n = 9) were associated with lower cholesterol levels than depressive (201.0 +/- 49.4) or mixed (226.4 +/- 44.4) episodes (p = 0.02 for comparison of manic and mixed episodes). Body mass index (BMI), age, alcohol use, and gender did not account for these findings. CONCLUSIONS: Cholesterol levels were lower in manic and depressive than in mixed episodes. No differences were found between diagnoses of unipolar or bipolar mood disorders. Cholesterol may be a state rather than a trait function, and may be influenced by the acute mood state. PMID- 11254023 TI - Amantadine in depressive patients with Borna disease virus (BDV) infection: an open trial. AB - OBJECTIVE: Originally introduced into pharmacotherapy as an antiviral compound, amantadine was shown to also have multiple pharmacological eftfects on the central nervous system. In addition. only a few studies reported on certain antidepressive properties of amantadine. This effect was highlighted by the discovery of its antiviral effect on Borna disease virus (BDV), which is hypothesized to be an etiopathogenetic factor to subtypes of affective disorders. Therefore, the therapeutical use of amantadine in BDV-infected depressive patients was investigated. METHODS: In this open trial, amantadine was added to antidepressive and or mood-stabilizing compounds treating BDV-infected depressed patients (n = 25) with bipolar or major depressive disorders. Amantadine was given twice a day (100-300 mg/day) for a mean of 11 weeks. Antidepressive treatment response was measured on the Hamilton rating scale for depression (HAM D) and/or with an operationalized diagnostic criteria system (OPCRIT: version 3.31). Virological response was measured by expression of BDV infection parameters in blood samples. RESULTS: The overall response rate of the amantadine augmentation in the BDV-infected patients with regard to depressive symptoms was 68% after a mean of 2.9 weeks of treatment. Bipolar I patients improved faster and did not show any following hypomania. In addition, the decrease of depression tended to correspond with the decrease in viral activity. CONCLUSION: Amantadine appears to show a remarkable antidepressive efficacy in BDV-infected depressive patients. The antidepressive effect in this open trial appeared to be comparable to standard antidepressives, possibly being a result of its antiviral effect against BDV as a potentially relevant etiopathogenetic factor in these disorders. PMID- 11254021 TI - An open study of methylphenidate in bipolar depression. AB - BACKGROUND: The treatment of bipolar depression is problematic. Mood stabilizing agents are often inadequate, while antidepressants may induce mania or mood destabilization. Methylphenidate has been advocated as an effective antidepressant agent in unipolar depression, and depression secondary to medical illness. Amphetamine administration has been shown to reduce manic behavior. These independent observations suggest that methylphenidate may be a safe and effective agent in bipolar depression. METHODS: Fourteen depressed subjects with DSM-IV bipolar illness and a Hamilton-depression (HAM-D) scale score of at least 15 had methylphenidate added to a stable mood stabilizer regiment. Patients were followed weekly for 4 weeks and then biweekly for an additional 8 weeks. RESULTS: HAM-D scores dropped from 16.9 +/- 1.79 SD at baseline to 9.4 +/- 9.73 on week 12 (p = 0.12, t = 1.84, df= 6) and 9.8 +/- 7.56 on last observation carried forward (LOCF) (p = 0.019, t = 2.8, df = 10). Psychiatric symptom assessment scale (PSAS) scores dropped from 17.9 +/- 5.63 at baseline to 4.8 +/- 7.47 at week 12 (p = 0.016, t = 4.02, df= 4) and 6.3 +/- 6.75 on LOCF (p = 0.007, t = 3.74, df = 7). Three individuals stopped secondary to anxiety, agitation, and hypomania, respectively. CONCLUSION: In this brief, open study, methylphenidate was effective and relatively safe in depressed bipolar subjects. PMID- 11254024 TI - Comorbidity of obsessive-compulsive disorder in recovered inpatients with bipolar disorder. AB - OBJECTIVE: To determine the frequency of obsessive-compulsive disorder (OCD) in inpatient subjects with bipolar disorder (BD) and to examine the clinical characteristics of BD subjects with OCD. METHOD: The sample consisted of 143 inpatient subjects with DSM-III-R BD-I and BD-NOS (BD-II), recovered from a current episode of either depression or mania. Demographic and clinical variables were obtained on the day of admission. Current comorbid conditions including OCD were determined by the Structured Clinical Interview for DSM-III-R Ifollowing recovery from the acute affective episode. RESULTS: The frequency of current OCD was 7% (N = 10). All BD subjects with OCD were BD-II, were male, and had a diagnosis of current dysthymia. They had fewer episodes and a higher incidence of prior suicide attempts than bipolar subjects without OCD. None of the bipolar subjects with OCD fulfilled criteria for cyclothymia. CONCLUSIONS: Our findings suggest that BD-II, OCD, dysthymia, and suicidality cluster together in some subjects with BD. We discuss the clinical implications of our findings. PMID- 11254025 TI - Searching high and low: a review of the genetics of bipolar disorder. AB - OBJECTIVES: To review the methodologies and findings in the genetics of bipolar disorder (BPD), and to suggest future directions for research. METHODS: Reports of family, twin, adoption, linkage, association, cytogenetic, and animal model studies, and segregation analyses in English, were identified from multiple MEDLINE searches. Hand searches were carried out in bibliographies from review articles. RESULTS: Family, twin, and adoption studies have provided strong evidence for a genetic etiology in BPD. Early reports of linkage of BPD to DNA markers at several chromosomal sites have not proven robust, perhaps because of the complex nature of BPD inheritance. However, linkage findings in the 1990s, on chromosomes 18, 21q, 12q, and 4p, have provided leads that are being pursued through both genetic and physical mapping. No gene has yet been definitively implicated in BPD. CONCLUSIONS: Strategies for increasing the power to detect BPD genes include: (1) dividing the phenotype into genetically meaningful subtypes to decrease heterogeneity: and (2) ascertaining a very large family sample--a multicenter study now in progress will collect 700 bipolar I sibling pairs. BPD may result from several genes acting in concert so that new multilocus statistical methods could enhance the capacity to detect loci involved. Family based association studies using a very large number of newly identified single nucleotide polymorphisms (SNPs) may allow for more efficient screening of the genome. As the Human Genome Project approaches its goal of isolating all genes by 2003, the data generated is likely to speed identification of candidate BPD genes. PMID- 11254026 TI - Effect of 17-beta-estradiol and ginsenoside Rg1 on reactive microglia induced by beta-amyloid peptides. AB - The reactive microglias induced by 25 micromol of beta-amyloid peptides (Abeta25 35) and/or IFN-gamma can initiate the microglial respiratory burst and release NO, respectively. Oxidative stress and inflammatory function have been implicated in Alzheimer's disease (AD). We showed that 10 micromol 17-beta-estradiol (E2) and 1-10 micromol ginsenoside Rg1 (Rg1) could prevent the toxicity of Abeta25-35 and/or IFN-gamma to microglias, inhibit the microglial respiratory burst activity and decrease the accumulation of NO. These results demonstrated the protectional effect of E2 or Rg1 on neuron from damaging by reactive microglias in AD. PMID- 11254027 TI - Two new diterpenoids from Mallotus apelta Muell.Arg. AB - Two new diterpenoids, malloapeltene (6,8-dihydroxy-cembrene-5-one) and malloapeltin (4alpha,15,16-trihydroxy-dolabradane) were isolated and characterized from the petroleum ether fraction of the alcoholic extract of Mallotus apelta Muell.Arg. Their structures were determined by spectral methods. PMID- 11254028 TI - New guaianolides from Artemisia selengensis. AB - A new guaianolide artselenin (1) and a new dimeric guaianolide artselenoid (2), along with 10 known compounds, were isolated from the aerial parts of Artemisia selengensis. Their structures were elucidated by spectroscopic methods. Two dimensional NMR techniques were used to make complete assignments for the 1H- and 13C-NMR chemical shifts of the two new guaianolides. PMID- 11254029 TI - Neopallavicinin from the Taiwanese liverwort Pallavicinia subciliata. AB - Neopallavicinin (2), a diastereomer of pallavicinin (1), was identified from the Taiwanese liverwort Pallavicinia subciliata. The structure of neopallavicinin was deduced by spectroscopic analysis. Three chemotypes may be classified for the species Pallavicinia subciliata. PMID- 11254030 TI - Total synthesis of (-)-incrustoporin. AB - (-)-Incrustoporin (1) has been synthesized using aldol condensation of ethyl p tolyl-acetate (2) and (2R)-benzoyloxy-butanal (3), followed by acid-catalyzed deprotection of the benzoyl group, lactone ring-closure, and elimination of the beta-OH in a one-pot manner. The aldehyde 3 was prepared from the commercially available D-mannitol by a two-directional strategy. PMID- 11254031 TI - Styryllactones from the rhizomes of Goniothalamus griffithii. AB - Three new styryl-lactones 8-acetylgoniofufurone(1), 7-acetylgonio-pypyrone(3), and 5-acetylgoniopypyrone(4), along with ten known compounds, goniofufurone(2), goniopypyrone(5), goniothalamin, goniothalenol, (+)-isoaltholactone, goniodiol, 7 acetylgoniodiol, goniotriol, 8-acetylgoniotriol, 9-deoxygoniopypyrone were isolated from the rhizomes of Goniothalamus griffithii Hook f. et. Thoms. Their structures were elucidated by IR, MS, NMR spectra and chemical evidence. All compounds showed cytotoxic activities against human cancer cell lines. PMID- 11254032 TI - Triterpenoid saponins from Vaccaria segetalis. AB - A new triterpenoid saponin, named segetoside C (1), and two known saponins, vaccaroid A (vaccaroside A) (2) and dianoside G (3), have been isolated from the seeds of Vaccaria segetalis. On the basis of chemical reaction and spectral data, the structure of segetoside C (1) has been established as: gypsogenic acid-28-O [beta-D-glucopyranosyl-(1-->3)]-[6-O-acetyl-beta-D-glucopyra-nosyl-(1-->2)-beta-D glucopyranosyl-(1-->6)]-beta-D-glucopyranoside. PMID- 11254033 TI - Structures and stereochemistry of pseudolarolides K and L, novel triterpene dilactones from pseudolarix kaempferi. AB - Pseudolarolides K(1) and L(2), two novel triterpenedilactones, were isolated from the seeds of Pseudolarix kaempferi, and their structures characterized from spectral data. PMID- 11254034 TI - Chemical pattern recognition of three Chinese herbal medicines from the genus Stephania lour. AB - Chemical pattern recognition was applied to three Chinese herbal medicines from the genus Stephania Lour., viz. S. kwangsiensis Lo, S. viridiflavens Lo and M. Yang and S. mashanica Lo and B.N. Chang. Based on the chemical features obtained from HPLC, SIMCA program was carried out and the results showed that the classification accuracy was 100%. In addition, the obtained features showed three major classes by NLM. The results of both methods were consistent with those of plant taxonomical identification. It suggested that chemical pattern recognition could be a helpful method to classify and identify Chinese herbal medicines. PMID- 11254035 TI - A new furofuran mono-lactone from Forsythia suspensa. AB - A new furofuran mono-lactone, named forsythenin, was isolated from the fruits of Forsythia suspensa, together with the known compounds, ocotillone, ocotillol monoacetate, (6'-O-palmitoyl)-sitosterol-3-O-beta-D-glucoside and palmitic acid. The structure of the new compound was elucidated on the basis of spectroscopic means and X-ray crystallography. PMID- 11254036 TI - New polyoxygenated cyclohexenes from Uvaria calamistrata. AB - Five new polyoxygenated cyclohexenes, named uvacalol A (1), B (2), C (3), D (4) and E (5) were isolated from the roots of Uvaria calamistrata. On the basis of spectral analysis and chemical derivatization, including the preparation of Mosher esters, the structures of compound 1-5 were established as (2R,3S,4R,5S)-2 acetoxyl-5-ethoxyl-1-benzoyloxymethylcyclohex-1(6)-ene3,4-diol-3-benzoate, (2R,3S,4R,5S)-2-acetoxyl-5-ethoxyl-1-benzoyloxymethylcyclohex-1(6)-ene-3,4-diol-4 benzoate, (2R,3S,4R,5S)-5-ethoxyl-1-benzoyloxymethylcyclohex-1(6)-ene-2,3,4-triol 3-benzoate, (2R,3S,4R,5S)-3-methoxyl-1-benzoyloxymethylcyclohex-1(6)-ene-2,3,5 triol and (2R,3S,4R,5S)-2-acetoxyl-1-benzoyloxymethylcyclohex-1(6)-ene-3,4,5 triol-5-benzoate, respectively. PMID- 11254038 TI - Trace level determination of perchlorate in water matrices and human urine using ESI-FAIMS-MS. AB - High-field asymmetric waveform ion mobility spectrometry (FAIMS) separates perchlorate from interfering isobaric ions of bisulfate and dihydrogenphosphate in the gas-phase. The use of a new FAIMS prototype and waveform generator, along with the use of a mixed carrier gas, in this electrospray ionisation (ESI)-FAIMS mass spectrometry (MS) study gave a detection limit for perchlorate in a relatively "clean" matrix of tap water of 0.050 ppb. Flow injection analysis (FIA) of dilutions of fortified waste water, a fortified river water certified reference material (CRM; SLRS-4, National Research Council of Canada), and a fortified human urine Standard Reference Material (SRM; 2381, National Institute of Standards and Technology) gave detection limits of 0.37 ppb, 0.50 ppb, and 4.8 ppb, respectively, in the undiluted matrices. PMID- 11254037 TI - Microsystems technology for remote monitoring and control in sustainable agricultural practices. AB - The future development of advanced sensor and microsystems technologies for use in agriculture is briefly discussed, within a general appraisal of the industrial requirements and the potential economic and ecological benefits to be derived from the employment of novel sensing systems in sustainable agricultural practices, such as precision farming. It is proposed that these technologies are potentially important monitoring tools for farmers and growers, and that a concerted programme of research and development is required to satisfy the future requirements of the agricultural industry. PMID- 11254039 TI - The development of a chemical 'fingerprint' to characterise dissolved organic matter in natural waters. AB - A suite of twelve assays has been used to 'fingerprint' dissolved organic matter (DOM). The assays were applied directly to filtered natural water samples. Temperature, pH and conductivity accounted for the environmental conditions on site. Bulk carbon characteristics were assayed by measuring UV absorbance at 200 and 240 nm, colour in grade Hazen, DOC (dissolved organic carbon), fluorescence (excitation 370 nm, emission 450 nm) and the complexation of phenol itself. Measuring hydroxybenzenes ('monophenolics'), polyhydroxybenzenes ('polyphenolics') and total phenolics with the Gibbs, Prussian Blue and Folin Ciocalteau assays, respectively, determined the phenolics pool. The methodology was tested on six freshwater sites in North Wales chosen to provide differences in vegetation, land-use and water chemistry and sampled once during each season. A novel approach for the presentation of the data has been developed that combines all range normalised assay results for each site and each season within one polar plot, hence the term 'fingerprint'. The data was also analysed using principal component factor analysis. Assays characterised as determining the chemical properties of DOM contributed to Factor 1 and explained 59% of the variation in the data. Assays apparently determined by the water matrix, contributed to Factor 2 and explained 20% of the variation within the data. The factor scores obtained for each site showed more variation for assays relating to the chemical properties of DOM than to the surrounding water matrix. The methodology was found to detect chemical changes within DOM for each site throughout the year and different responses for different sites. PMID- 11254040 TI - A field evaluation of international monitoring guidelines for the biological effects of tributyltin. AB - The Oslo and Paris Commission (OSPAR) Guidelines for monitoring the biological effects of tributyltin compounds (TBT) were evaluated using data collected for preparation of a Celtic Seas Quality Status Report. Two types of survey were undertaken: broad scale, to determine wide impacts in coastal waters; and localised, around representative harbours to establish ranges of effect from recognised input points. This evaluation indicates that results from the broad scale surveys can be used to compare different areas of coastline. Nucella is widespread away from point sources although many individuals show some degree of imposex. Populations are generally not at risk. The localised surveys indicate that, in certain situations, the monitoring objectives can be met in the OSPAR Guidelines (subject to minor amendment). Criteria are identified for the selection of point sources suitable for monitoring under OSPAR Guidelines. PMID- 11254041 TI - Assessing metal sorption on the marine alga Pilayella littoralis. AB - Increasing interest in the development of biological materials for metal sorption led us to investigate the brown marine alga, Pilayella littoralis, as a biological sorbent. This work focuses on the harvest, preparation and evaluation of P. littoralis from Nahant beaches for use as a metal biosorbent. This biomass was used in batch tests with synthetic solutions. Its metal uptake properties, including metal binding capacity, the pH dependence of metal uptake and the kinetics of metal sorption, were investigated. Most metal sorption occurred within the first 5 min of exposure and the metals were optimally bound to the algae at pH 5.5. The algal binding capacities for Al(III), Cd(II), Co(II), Cr(VI), Cu(II), Fe(III), Ni(II) and Zn(II), were 2,000, 430, 560, 90, 850, 700, 390 and 450 micromol g(-1) of dried biomass, respectively. Metals were desorbed with 0.12 mol l(-1) HCl and determined by inductively coupled plasma atomic emission spectrometry (ICP-AES). PMID- 11254043 TI - Removal of atrazine and four organophosphorus pesticides from environmental waters by diatomaceous earth-remediation method. AB - A new viable remediation technique based on the use of diatomaceous earth is proposed to improve the ecological system. Its ability to remove atrazine and the four organophosphorus pesticides parathion-methyl, chlorpyriphos, fenamiphos and methidathion from river and waste waters has been proven. A series of experiments including variable conditions, such as temperature, pH, contact time, pesticide concentration and adsorbent quantity, were performed to demonstrate the efficiency of pesticide removal from three different water samples. The batch experiments showed that diatomaceous earth was able to remove 95% of chlorpyriphos, 75% of methidathion and parathion-methyl and 55% of atrazine and fenamiphos from all types of waters tested. The individual adsorption of each pesticide on diatomaceous earth could be described by the Freundlich isotherm and a tentative adsorption mechanism was proposed. The Freundlich coefficient (Kf) and Freundlich constant (1/n) appeared to be closely related to the physicochemical properties (Kow, solubility) of the compounds. The actual results support the conclusion that diatomaceous earth has the potential to serve as an extractant in remediation techniques. PMID- 11254042 TI - Interlaboratory trial to determine the analytical state-of-the-art of bromate determination in drinking water. AB - The new European Directive for water intended for human consumption has established a regulatory level for bromate at 10 microg L(-1). This Maximum Admissible Concentration requires analytical methods with detection limits of a least 2.5 microg L(-1). A project funded by the Standards, Measurements and Testing Programme of the European Commission has enabled the improvement and/or development of methods for the determination of bromate at such concentration levels. This collaborative work was concluded by the organisation of an interlaboratory trial involving 26 European laboratories, which enabled the testing of both a draft ISO Standard method and alternative methods. This paper presents the results of this interlaboratory trial, along with results of a bromate stability study. The progress made with respect to the analytical state of-the-art for bromate will greatly benefit the quality of measurements carried out in water quality monitoring. PMID- 11254044 TI - Trend, seasonal and multivariate modelling study of wet precipitation data from the Austrian Monitoring Network (1990-1997). AB - The aim of the present study was to analyse the data structure of a large data set from rainwater samples collected during a long-term interval (1990-1997) by the Austrian Precipitation Monitoring Network. Eleven sampling sites from the network were chosen as data sources (chemical concentrations of major ions only) covering various location characteristics (height above sea level, rural and urban sampling positions, Alpine rim and Alpine valley disposition, etc.). The analytical results were treated by the application of already classical environmetric approaches, such as linear regression analysis, time-series analysis and principal components analysis (PCA). For most of the sampling sites, a distinct trend of acidity decrease of the wet precipitation was observed. An overall decrease in sulfate concentration for the whole period and all sites of 3.9% year(-1) (2.0 muequiv. L(-1) year(-1)) was found. The free acidity decrease for most of the sites was between 3.5 and 10.9% year(-1). No significant linear trends were found for nitrate. Base cations either decreased (mean percentage decrease for calcium was 5.4% year(-1) and for magnesium 4.4% year(-1)) or did not show any significant change (sodium, potassium). The overall decrease in ammonium concentration was 2.3% year(-1). Further, some typical "rural" (summer minima and winter maxima) and "urban" (winter minima and spring maxima) seasonal behaviour for the majority of the sites in consideration could be defined, indicating the influence of local emission sources. Several latent factors, named "anthropogenic", "crustal" and "mixed salt", were revealed by the multivariate modelling procedure (PCA) possessing a similar structure for most of the sites. The unavoidable exceptions observed were indications of the influence of sporadic local events (construction and agricultural activities, secondary emission sources, etc.), and an effort was made to explain these exceptions. PMID- 11254045 TI - Development of perfluorocarbon tracer technology for underground leak location. AB - A method has been developed for the atmospheric sampling and analysis of four perfluorocarbon tracer (PFT) compounds simultaneously at the parts per trillion (ppt) level. PFTs were pre-concentrated using adsorbent tube air sampling. Analysis was achieved by thermal desorption (TD) and gas chromatography (GC) with electron capture detection (ECD). Efficient separation of the PFTs from the other sample constituents was achieved by use of a capillary porous layer open tubular (PLOT) GC column without the need to cool the GC oven to sub-ambient temperatures using liquid coolants (M. de Bortoli and E. Pecchio, J. High Resolut. Chromatogr., 1985, 8, 422) or for a catalytic destruction step to remove interferents (T. W. D'Ottavio, R. W. Goodrich and R. N. Dietz, Environ. Sci. Technol., 1986, 20, 100). Results from test field trials with two volatile PFTs that were buried to simulate an underground leaking cable were successful. The PFTs were detected above ground level to pinpoint the leak position. The highest tracer concentrations were detected within 1 m of the simulated leak positions 2 days after tracer burial. The developed technology was applied to an oil leaking high voltage electricity cable. One PFT was added to the cable oil which enabled detection of the oil leak to within 3 m. The reported method has many advantages over currently used leak detection methods and could, in the future, be applied to the detection of underground leaks in a variety of cables and pipes. PMID- 11254046 TI - Characterization of landfill leachates and studies on heavy metal removal. AB - This study covers a thorough characterisation of landfill leachates emerging from a sanitary landfill area. The landfill leachates were obtained in the acidic stage of landfill stabilisation. Their organic content was high as reflected by the high BOD5 (5 day biological oxygen demand) and COD (chemical oxygen demand) values. They were also highly polluted in terms of the parameters TKN (total Kjeldahl nitrogen), NH4-N, alkalinity, hardness and heavy metals. Nickel was present in these wastewaters at a significant concentration. With regard to the high heavy metal content of these wastewaters, several physicochemical removal alternatives for the heavy metals Cu, Pb, Zn, Ni, Cd, Cr, Mn and Fe were tested using coagulation, flocculation, precipitation, base addition and aeration. Additionally, COD removal and ammonia stripping were examined. Co-precipitation with either alum or iron salts did not usually lead to significantly higher heavy metal removal than lime alone. The major methods leading to an effective heavy metal removal were aeration and lime addition. Nickel and cadmium seemed to be strongly complexed and were not removed by any method. Also lead removal proved to be difficult. The results are also discussed in terms of compliance with standards. PMID- 11254047 TI - Collaborative evaluation of the analytical state-of-the-art of platinum, palladium and rhodium determinations in road dust. AB - In order to control the quality of platinum, palladium and rhodium determinations in road dust, the Standards, Measurements and Testing Programme (formerly BCR) of the European Commission has started a project, the final aim of which is to certify a road dust reference material for its contents of platinum group elements. The first part of this project consisted of an interlaboratory study, which aimed to test the feasibility of the preparation of a candidate road dust reference material and to detect and remove most of the pitfalls observed in platinum, palladium and rhodium determinations. This paper presents the main results of this interlaboratory study carried out prior to the certification campaign. The concordance of the data obtained by the participating laboratories for the three elements was considered to reflect the state-of-the-art and was encouraging enough to decide on the organization of a certification campaign to be conducted during the year 2000. The progress made with respect to the analytical state-of-the-art for these elements will be of great benefit to the quality of measurements carried out in environmental monitoring in this particular field. PMID- 11254048 TI - A new method for the determination of trichloroacetic acid in spruce foliage and other environmental media. AB - Trichloroacetic acid (TCAA) is a phytotoxic chemical, present throughout the environment. The majority of methods for analysis of TCAA require chemical derivatisation and multiple extraction steps prior to analysis by gas chromatography. Here, a new analytical method for TCAA determination in environmental matrices is reported. The method is based on a modified Nielsen Kryger steam distillation that combines into one 1 h reflux the thermal decarboxylation of TCAA to CHCl3 and the partitioning and concentration of the CHCl3 into 5 ml of hexane, which is analysed by GC. Sample preparation is minimal and no matrix standard additions are required. The background CHCl3 in the sample is removed prior to extraction by degassing the solution for 1 h with nitrogen. Optimisation of the method gave recoveries from three separate solutions of 0.31 ppb aqueous TCAA standards of 93 +/- 15% (n = 9), 110 +/- 9% (n = 9) and 105 +/- 11% (n = 6). The extraction method has been compared with a decarboxylation and headspace GC method for determination of TCAA in Sitka spruce needles. No significant difference in TCAA concentration or replicate precision between the two methods was observed. PMID- 11254049 TI - Direct on-filter assay of subtilisin-type proteolytic enzymes for rapid analysis of analyte captured from the workplace atmosphere. AB - A simple assay for some proteolytic enzymes has been developed which can be performed directly on the surface of a cellulose nitrate filter used to capture the analyte during workplace monitoring for health and safety purposes. Following air sampling the analysis is performed on the filter which is retained within the air sampler. This involves two steps: first, a 15 min incubation in which the captured enzyme is dissolved and then digests an alkaline-phosphatase-labelled antibody immobilised as a small dot on the surface of the filter; and second, is a 10 min incubation with substrate solution, which follows an in situ wash under a vacuum. During the incubation colour develops on the spot at the location of the immobilised enzyme antibody conjugate. The intensity of the spot can be assessed visually within the sampler to ascertain the presence or absence of captured enzyme, or alternatively quantitative results can be obtained using an optical scanner. The limit of detection is 5 ng per filter for subtilisin (20 ng for visual discrimination between this standard and the zero). The assay is stable to the effects of ambient air sampling at 31 min(-1) for 18 h. PMID- 11254050 TI - Intercomparison of five PM10 monitoring devices and the implications for exposure measurement in epidemiological research. AB - Five different instruments for the determination of the mass concentration of PM10 in air were compared side-by-side for up to 33 days in an undisturbed indoor environment: a tripod mounted BGI Inc. PQ100 gravimetric sampler with a US EPA certified Graseby Andersen PM10 inlet; an Airmetrics Minivol static gravimetric sampler; a Casella cyclone gravimetric personal sampler; an Institute of Occupational Medicine gravimetric PM10 personal sampler; and two TSI Inc. Dustrak real-time optical scattering personal samplers. For 24 h sampling of ambient PM10 concentrations around 10 microg m(-3), the estimated measurement uncertainty for the two gravimetric personal samplers was larger (approximately +/- 20%) compared with estimated measurement uncertainty for the PQ100/Graseby Andersen sampler (< +/- 5%). Measurement uncertainty for the Dustraks was lower (approximately +/- 15% on average) but calibration of the optical response against a reference PM10 method is essential since the Dustraks systematically over-read PM10 determined gravimetrically by a factor approximately 2.2. However, once calibrated, the Dustrak devices demonstrated excellent functionality in terms of ease of portability and real-time data acquisition. Estimated measurement uncertainty for PM10 concentrations determined with the Minivol were +/- 5%. The Minivol data correlated well with PQ100/Graseby Andersen data (r= 0.97, n = 18) but were, on average, 23% greater. The reason for the systematic discrepancy could not be traced. Intercomparison experiments such as these are essential for assessing measurement error and revealing systematic bias. Application of two Dustraks demonstrated the spatial and temporal variability of exposure to PM10 in different walking and transport microenvironments in the city of Edinburgh, UK. For example, very large exposures to PM10 were identified for the lower deck of a double-decker tour bus compared with the open upper deck of the same vehicle. The variability observed emphasises the need to determine truly personal exposure profiles of PM10 for quantifying exposure response relationships for epidemiological studies. PMID- 11254051 TI - Airborne thermal degradation products of polyurethene coatings in car repair shops. AB - A methodology for workplace air monitoring of aromatic and aliphatic, mono- and polyisocyanates by derivatisation with di-n-butylamine (DBA) is presented. Air sampling was performed using midget impinger flasks containing 10 ml of 0.01 mol l(-1) DBA in toluene and a glass-fibre filter in series after the impinger flask, thereby providing the possibility of collecting and derivatising isocyanates in both the gas and particle phases. Quantification was made by LC-MS, monitoring the molecular ions [MH]+. Air samples taken with this method in car repair shops showed that many different isocyanates are formed during thermal decomposition of polyurethane (PUR) coatings. In addition to isocyanates such as hexamethylene (HDI), isophorone (IPDI), toluene (TDI) and methylenediphenyl diisocyanate (MDI), monoisocyanates such as methyl (MIC), ethyl (EIC), propyl (PIC), butyl (BIC) and phenyl isocyanate (PhI) were found. In many air samples the aliphatic monoisocyanates dominated. During cutting and welding operations, the highest levels of isocyanates were observed. In a single air sample from a welding operation in a car repair shop, the highest concentrations found were: MIC, 290; EIC, 60; PIC, 20; BIC, 9; PhI, 27; HDI, 105; IPDI, 39; MDI, 4; and 2,4-TDI and 2,6-TDI 140 microg m(-3). Monitoring the particle size distribution and concentration during grinding, welding and cutting operations showed that ultrafine particles (< 0.1 microm) were formed at high concentrations. Isocyanates with low volatility were mainly found in the particle phase, but isocyanates with a relatively high volatility such as TDI, were found in both the particle and gas phases. PMID- 11254052 TI - The determination of carbonyl compounds in air using a robotic sampling preparation system integrated to a gas chromatograph with a nitrogen-phosphorus detector. AB - A totally automated solid phase extraction gas chromatography procedure was developed for the sampling and analysis of carbonyl compounds in air. In this system, two PrepStation modules were used, one for the preparation and elution of 2,4-dinitrophenylhydrazine silica cartridges, and the other for air sampling. The sample collected by the sampling module was eluted to an autosampler vial in the PrepStation module and then transferred to the gas chromatograph for analysis via a robotic arm. The sampling module was modified to enable air sampling via an external pump. A typical run by this technique required 142 min, 100 min for air sampling and 42 min for the other operations, including a GC analysis time of 25 min. Recoveries of at least 85% were obtained for all compounds studied. The detection limits for formaldehyde, acetaldehyde and acetone were 2.2, 2.7 and 2.2 ppbv, respectively. All operations, including the conditioning of the cartridges, were performed without any intervention from the analyst. PMID- 11254053 TI - Phosphine sampling and analysis using silver nitrate impregnated filters. AB - In the field of industrial hygiene, besides the necessity of monitoring phosphine with direct reading apparatus to prevent accidents, there is a need for a method of sampling and analysing phosphine to control workers' exposure. The use of filters impregnated with silver nitrate to collect arsine, phosphine and stibine in workplace air has been described in the literature. Having previously chosen this type of filter to collect arsine, we studied its characteristics for phosphine capture. A filter impregnated with sodium carbonate was used both as a prefilter to collect the particles and to trap arsenic trioxide. After dissolving the silver compounds in nitric acid, ICP emission spectrometry was used to carry out the analysis. This article describes the comparative sampling we performed in a microelectronic laboratory and in a fumigation chamber (130 samples) to determine the concentration of AgNO3 impregnation solution to be used, the detection limit of the method and the retention capacity of the impregnated filters. Interference with other gases reacting with silver nitrate was studied and the storage time for sampled filters and analysis solutions was checked. The detection limit of the adopted method is better than 1 microg per filter, and the retention capacity exceeds 300 microg per filter. The problem of how to sample phosphine when H2S, NH3, or HCl is present has been solved, but the problem of sampling phosphine in atmospheres where acetylene evolves remains. Sampled filters and filter solutions are stable for more than three months at ambient temperature. PMID- 11254054 TI - BTX measurements with diffusive samplers in the vicinity of a cokery: comparison between ORSA-type samplers and pumped sampling. AB - In the framework of new European directives in the field of ambient air quality assessment specific requirements for measurement methods for benzene will be defined and have to be met in the future. In a residential area in the vicinity of a cokery a comprehensive monitoring programme for BTX compounds (benzene, toluene, xylenes) was carried out with diffusive samplers. Measurement results from 20 sites show a distinct spatial distribution of benzene concentrations which is caused by the emissions of the plant. Comparisons with two pumped measurement methods reveal excellent agreement in the case of benzene. An assessment of measurement uncertainty for the diffusive method is presented in terms of standard deviations from parallel measurements (precision) and by comparison with a reference method according to national and international standards. Data quality objectives required by an upcoming European directive for benzene can be met by diffusive sampling. PMID- 11254056 TI - Shipboard techniques based on flow injection analysis for measuring dissolved Fe, Mn and Al in seawater. AB - An overview is presented of sampling techniques and flow injection analysis (FIA) methods for low concentrations of Fe, Mn and Al in filtered seawater. On the basis of sampling procedures, filtration techniques, accuracy, blanks, detection limits, intercalibration results and oceanographic consistency, the feasibility of these FIA methods was evaluated. It was found that these metals could be measured on board with a minimum risk of contamination and with good accuracy even at low subnanomolar levels (<0.5 nM). Results for reference seawater were in the case of Fe-FIA and Mn-FIA in excellent agreement with the certified values. Data from samples analyzed by Fe-FIA and by cathodic stripping voltametry (CSV) compared well, as did Mn-FIA and GFAAS. All three methods gave results that were mostly in good agreement with data from the same ocean regions published by other research groups. Two different types of surface water sampling were also tested and compared, namely conventional hand filling of a sample bottle from a rubber dinghy away from the ship, and underway pumping of seawater using a 'tow fish'. The latter method gave the best results. Also, conventional membrane filtration and cartridge filtration for large volume filtration were compared using Fe and Al data from water column samples. Good agreement was found for both filter types, although for defining dissolved metal species the latter filter type was preferred. PMID- 11254055 TI - Development of a novel passive sampling system for the time-averaged measurement of a range of organic pollutants in aquatic environments. AB - A new sampling system has been developed for the measurement of time-averaged concentrations of organic micropollutants in aquatic environments. The system is based on the diffusion of targeted organic compounds through a rate-limiting membrane and the subsequent accumulation of these species in a bound, hydrophobic, solid-phase material. It provides a novel and robust solution to the problem of monitoring in situations where large temporal fluctuations in pollutant levels may occur. Accumulation rates are regulated by choice of diffusion-limiting membrane and bound solid-phase material and have been found to be dependent on the physico-chemical properties of individual target analytes. Two separate prototype systems are described: one suitable for the sampling of non-polar organic species with log octanol/water partition coefficient (log P) values greater than 4, the other for more polar species with log P values between 2 and 4. Both systems use the same solid-phase material (47 mm C18 Empore disk) as a receiving phase but are fitted with different rate-limiting membrane materials (polysulfone for the polar and polyethylene for the non-polar analytes). The two systems complement each other and together can be used for sampling a wider range of organic analytes than generally possible using current passive sampling techniques. Calibration data are presented for both devices. In each case, linear uptake kinetics were sustained, under constant conditions, for deployment periods of between 1 and 9 days. The effects of water temperature and turbulence on sampling rates have been quantitatively assessed. The performance of the system was further investigated by means of field exposures for one and two weeks in marine environments where calibrated samplers were used to determine the time-averaged concentrations of the polar biocides diuron and irgarol 1051. The quantitative results obtained using the passive sampler were compared with those obtained using spot sampling. PMID- 11254057 TI - Toxaphene: a challenging analytical problem. AB - The analysis of toxaphene, a highly complex mixture of chlorinated bornanes, bornenes and camphenes, is a challenging problem, especially as individual congeners are present at trace levels in biota and other relevant samples. The complicated nomenclature of the compounds of interest is briefly discussed. Gas chromatographic techniques are the most important tools in toxaphene analysis because of their high resolution. The main focus of attention is devoted to important steps in the GC-based analytical procedure, such as sample preparation and injection, separation by single-column and multidimensional GC, both heart cut and comprehensive, and detection by electron capture and, increasingly, MS based detection/identification. The gradual shift from total toxaphene to individual compound analysis is discussed, and the growing interest in enantiomer selective separations is highlighted. The problems encountered when selecting appropriate indicator compounds for rapid toxaphene screening are also addressed. PMID- 11254058 TI - Monitoring the quality of the marine environment. AB - As part of the monitoring program from Instituto Hidrografico, since 1981 sediment and water samples have been collected from four different estuarine areas located along the continental coast of Portugal. The concentrations of different parameters were measured in the water and sediment samples. After normalization, the concentrations of chromium, mercury, lead and zinc in the sediments from the different areas were compared. PMID- 11254059 TI - The QUASIMEME laboratory performance study of polycyclic aromatic hydrocarbons (PAHs): assessment for the period 1996-1999. Quality Assurance of Information in Marine Environmental Monitoring in Europe. AB - The results of the laboratory performance studies held during the period 1996-99 and involving the determination of PAHs in sediments and biota have been assessed. Overall 67% of the data submitted met the QUASIMEME criterion for satisfactory performance, although differences in performance were apparent between the two main analytical techniques employed: gas chromatography and high performance liquid chromatography. Further development of PAH methodology utilising coupled gas chromatography mass spectrometry is encouraged as the most appropriate means of meeting the future requirements for the determination of a wider range of PAH analytes. More reference materials are also urgently required for use in marine monitoring programmes, and they need to be certified for a wider range of PAHs than hitherto, including both parent PAHs and their alkylated derivatives. PMID- 11254060 TI - Determination of organochlorinated compounds and petroleum hydrocarbons in sediment sample IAEA-408. Results from a world-wide intercalibration exercise. AB - A sediment sample from the intertidal mudflats of the Tagus estuary was prepared, homogenised and distributed globally to laboratories as the IAEA-408 intercomparison material for the analyses of organochlorinated pesticides, PCBs and petroleum hydrocarbons (PHs). A total of 48 laboratories from 36 countries reported their results. The data from participants show that there still remain some difficulties with the accurate determination of organic contaminants such as pesticides and polycyclic aromatic hydrocarbons (PAHs). More consistent interlaboratory results were obtained for PCBs congeners. The final results of this intercomparison exercise enable individual participants to assess their performance and, where necessary, to introduce appropriate modifications in their analytical procedures. Furthermore, as a series of statistical criteria was fulfilled for a number of compounds, the sample IAEA-408 can now be used as a reference material for quality control in the determination of some persistant organic pollutants (POPs) in marine sediment samples. PMID- 11254061 TI - Sediment composition and normalisation procedures: an example from a QUASH project sediment exercise. AB - The QUASH UE-Project was designed to assess the reliability of normalisation approaches to compensate the influence of natural process affecting the distribution and concentration of contaminants in sediment. The focus of this paper was to test the influence on normalisation procedures of an inorganic matrix using a sample collected in the Venice Lagoon, Italy. PMID- 11254062 TI - The determination and distribution of Zn in surface water samples collected in the northeast Atlantic Ocean. AB - Dissolved Zn concentrations were determined in surface water samples collected on line along transects in the eastern North Atlantic in spring (March 1998). Two frontal zones could be identified in the research area by a change in salinity, temperature and nutrient concentrations. One zone was identified at 42 degrees N, separating the North Atlantic central water (NACW) and the Atlantic surface water (ASW) from each other, and another one crossing the continental slope at 12 degrees and 8 degrees E, respectively. Variability in Zn concentrations was observed near these zones, not only as a result of a change of water mass, but also due to external Zn sources. Surface Zn concentrations were 0.5-1 nM and 2 nM in the NACW and ASW, respectively, increasing to 4 nM over the continental shelf and finally 5-6 nM in the English Channel. Contributions of Zn derived from shelf sediments appear to be the major source for the enriched surface values in the continental zone. PMID- 11254063 TI - QUASIMEME-QUASH. Quality Assurance of Information for Marine Environmental Monitoring in Europe-Quality Assurance of Sampling and Sample Handling. PMID- 11254064 TI - Unhealthy particles. PMID- 11254065 TI - Monitoring of pesticides in the UK environment. PMID- 11254066 TI - Refining the costs analyses of the Fort Bragg evaluation: the impact of cost offset and cost shifting. AB - A key aim of the evaluation of the Fort Bragg Demonstration was to determine whether delivering services through a continuum of care lowered expenditures on mental health services. The evaluation clearly showed that expenditures were actually higher in the Demonstration. Critics of the evaluation claimed that the evaluation's perspective on costs was too narrow-in particular, that the Demonstration produced cost shifting and cost offset that were not captured by the evaluation. New data allow us to include a broader array of costs: mental health services received outside the catchment areas, general medical services for the children themselves, and mental health services used by family members. Results showed that reductions in other costs do partially offset higher expenditures on mental health services for children at the Fort Bragg Demonstration. However, even when broader costs are included, total family expenditures are still substantially higher at the Demonstration. PMID- 11254067 TI - Predictors of barriers to treatment and therapeutic change in outpatient therapy for antisocial children and their families. AB - This study examined the role of parent psychopathology and quality of life in predicting barriers to participation in outpatient treatment and therapeutic change among clinically referred children. Children (N = 169) referred for oppositional, aggressive, and antisocial behavior and their families participated. The major findings were that (1) higher levels of parent psychopathology and lower levels of quality of life predicted the subsequent emergence of perceived barriers to treatment in the parents and therapeutic changes among the children, (2) these effects were not explained by socioeconomic disadvantage or severity of child dysfunction, (3) perceived barriers and therapeutic changes were related and this relation was not explained by other family and child predictors, (4) as the level of perceived barriers increased among families, the amount of therapeutic change and the proportion of children who made a marked change decreased, and (5) parent perception of the relevance and demandingness of treatment were salient dimensions contributing to the relation between perceived barriers and therapeutic change. The conceptual and applied implications of relating barriers to treatment and therapeutic change are discussed. PMID- 11254068 TI - Multisystemic therapy versus hospitalization for crisis stabilization of youth: placement outcomes 4 months postreferral. AB - Hospitalization and out-of-home placement data for 113 youth participating in a randomized trial comparing home-based multisystemic therapy (MST; n = 57) with hospitalization (n = 56) for psychiatric crisis stabilization were analyzed following the completion of MST treatment--approximately 4 months post approval for emergency psychiatric hospitalization. Analyses showed that MST prevented any hospitalization for 57% of the participants in the MST condition and reduced the overall number of days hospitalized by 72%. Importantly, the reduction in use and length of hospitalization was not offset by increased use of other placement options, as MST reduced days in other out-of-home placements by 49%. The cost implications for the viability of MST as an alternative to hospitalization for youth presenting psychiatric emergencies are discussed. PMID- 11254069 TI - From case management to court clinic: examining forensic system involvement of persons with severe mental illness. AB - The study examined the flow of a state mental health agency's case-managed clients into its forensic mental health court clinic systems for evaluation of competency to stand trial (CST) for a criminal offense. An analysis of merged encounter data from the case management and court clinic systems revealed that roughly 2% of the case-managed population were referred to court clinics for evaluation of CST during a 1-year period, but that these 2% represented roughly one eighth of that year's court clinic evaluees. The likelihood of this involvement was higher for males, African-Americans, and Latinos, and for persons with a history of substance abuse, and also was associated with higher levels of previous hospitalization. In addition, CST evaluees were more likely to be non White, male, and uninsured than were case-managed evaluees. These data indicate that demographic characteristics, substance abuse, and lack of insurance are potential risk factors for forensic and, by inference, criminal justice system involvement among persons with mental illness. PMID- 11254070 TI - Integrating vocational services on case management teams: outcomes from a research demonstration project. AB - Recent innovations to improve employment rates among persons with psychiatric disabilities include "hybrid case management/employment services." Project WINS was a research/demonstration project which integrated specialized vocational services into case management teams. In this report, client outcomes of WINS involvement are evaluated, using a quasiexperimental, longitudinal design. On almost all the work-related variables, participants in the immediate and delayed treatment conditions displayed better outcomes than those in the control condition, as did individuals receiving moderate or substantial service versus no/ minimal services. To address possible selection bias due to the quasiexperimental nature of the design, further analyses used baseline differences across conditions and participation levels as covariates. Results of multivariate analyses showed some anomalous findings regarding significant positive effects for the delayed, but not the immediate treatment condition versus the no-treatment control group. However, in similar analyses involving participation level as the independent variable, a moderate or substantial amount of service increased the odds of working by almost five times and also positively affected three other work-related variables. While limitations of this quasiexperimental design are noted, the results appear promising enough to support replications of WINS. PMID- 11254072 TI - Transmission of Theileria parva in the traditional farming sector in the Southern Province of Zambia during 1997-1998. AB - The incidence of first contact with the protozoan Theileria parva was determined in two traditional cattle herds in the Southern Province of Zambia during a period of average rainfall in 1997 and 1998, following a drought in the previous two years. Compared to that period, there was a marked increase in the number of rainy season first contacts attributable to transmission by Rhipicephalus appendiceulatus adults. However, there were still more dry season contacts that resulted from nymphal transmission. These results highlight the important role that climate plays in the transmission of theileriosis in the Southern Province of Zambia. PMID- 11254071 TI - Immunoprotective efficacy of a purified 39 kDa nymphal antigen of Hyalomma anatolicum anatolicum. AB - Soluble nymphal antigens (HNAg) were purified by immunoaffinity chromatography using CNBr-activated Sepharose 4B coupled with immunoglobulin ligands from animals immunized with HNAg and 69-71% protected against challenge infestations, and 8% recovery of the purified protein (Aff-HNAg) was obtained. Following immunization of crossbred calves (Bos indicus x Bos taurus) with 1600 sg of Aff HNAg in three divided doses, significant rejections of larvae (p<0.001, 84.2%), nymphs (p<0.05, 61.4%) and adults (p<0.05, 58.7%) were recorded. No significant changes were recorded in the engorgement weights of the larvae and nymphs, but there was a significant (p<0.05) reduction in the weight of the engorged adults. Immunization conferred a significant decrease in the numbers of resultant nymphs (p<0.001) and adults (p<0.001) that had fed on the immunized animals. SDS-PAGE analysis identified a 39 kDa protein, previously isolated from larvae of Hyalomma anatolicum anatolicum, as the antigen responsible for the induction of resistance against all the stages of the tick. PMID- 11254073 TI - Evaluating sugarcane diets for dairy cows using a digestion model. AB - To eliminate unnecessary feeding trials, a mechanistic model of sugarcane digestion was used in the search for suitable supplements to improve milk production. Milk production simulated by the model was compared with data observed in four feeding trials published in the tropical literature where crossbred dairy cows were fed sugarcane/urea diets with different types of supplements. The predicted effects of the supplements on the ruminal microbial population, concentrations of ammonia and volatile fatty acids were also compared with the published results in one experiment. The model indicated the nutrient most limiting milk production for the different feeding situations. The addtion of Leucaena to the basal sugarcane/urea improved the availability of amino acids and long-chain fatty acids, with energy becoming the limiting factor. Supplementation with rice bran increased the availability of energy and long chain fatty acids, but amino acids then became the limiting factor. Supplementation with both Leucaena and rice bran further improved the milk yield, but availability of energy now limited milk production. Supplementation with Leucaena increased milk production more than supplementation with king grass. The main reason for this increase was increased amino acid absorption due to increased microbial outflow. In all feeding situations, the average difference between the predicted milk production and that observed experimentally was 0.57 kg/d (ranging from 0.08 to 1 kg/d). PMID- 11254074 TI - Farmers' perceptions of the impacts of tsetse and trypanosomosis control on livestock production: evidence from southern Burkina Faso. AB - Interviews with all the households in the Agropastoral Zone (ZAP) of Yale, southern Burkina Faso, were conducted in 1994 and again in 1997 to assess the impacts of a tsetse control programme implemented from 1994, using insecticide impregnated targets and pour-on treatments of all cattle with deltamethrin 1%. In the absence of health and productivity monitoring, data were collected in single visit surveys in order to generate quantitative estimates of relevant reproductive performance variables for cattle and to assess changes in the inputs used and outputs produced. The results indicate a 25% increase in herd size and an increase in the number of oxen from 0.1 to 1.1 per household; a reduction in mortality from 63.1% to 7.1% and reductions in the rates of abortions and stillbirth of 55.9% and 51.3%, respectively; and an increase in the rate of live births of 57.6% and in the milk yield from 0.2 to 2.2 litres/cow per day in the dry season. These results show the dramatic impacts that trypanosomosis control can have on Zebu cattle exposed to high tsetse challenge. Well-designed surveys can be a cost-effective way to obtain estimates of productivity impacts that can be used to simulate projections of herd growth and meat and milk production in herd models. While there are many confounding factors associated with farmers' perception of a gain in productivity, these estimates form a useful alternative to subjective assessments in modelling the economic benefits of tsetse and trypanosomosis control. PMID- 11254075 TI - The handling and short-haul road transportation of spent buffaloes in relation to bruising and animal welfare. AB - The handling of 100 spent buffaloes during transportation by truck from an animal market to a slaughterhouse was observed so as to assess the influence of sex, body condition (weak, normal or heavy), body size based on the height at the hump (small, medium or large) and handling method (dragging, dragging + hitting or lifting + hitting) on the manpower requirement and the time spent in both loading and unloading. A buffalo could be loaded onto a truck with the help of 2.8 labourers in 66 s, whereas the average manpower and time needed for unloading a buffalo were 1.9 labourers and 26 s, respectively. Sex and size had no significant effect on the manpower requirement or the time spent in loading and unloading. Animals in normal body condition needed more (p<0.05) manpower (3.2 labourers) for loading compared to weak animals (2.5 labourers) or heavy animals (2.6 labourers). The manpower and time required for loading and unloading were least (p<0.05) when a stick was used while dragging. Excessive steepness of the loading ramp caused some of the animals to fall down during loading, whereas a slippery truck floor, due to increased eliminative activities by nervous animals, might result in buffaloes going down during unloading. The mean number of bruises was 2.44 per head. As muscle tissues were involved in about 90% of the bruises, considerable economic losses occurred through bruising, which necessitated removal of damaged tissues post mortem. Most of the bruises were found on the hind limbs (43.4%), followed by the abdomen and udder region (21.3%), shoulder, neck and back (16.0%) and perianal region (11.1%). PMID- 11254076 TI - Evaluation of the feeding value of palm press fibre using in vitro digestibility techniques. AB - Palm press fibre (PPF) was obtained from two sources, a small-scale oil palm processing unit and a large-scale factory processing unit, and its chemical composition was determined. In vitro digestibility techniques were used to assess the feeding value of untreated, defatted and sodium hydroxide-treated PPF. For the NaOH treatment, 0.5 g oven-dried PPF was treated for 24 h with 5% NaOH in three ways: treated and not washed (NaNW); treated and washed (NaW); and treated after milling (NAD). The results indicate that, on a dry matter basis, PPF is low in nitrogen (12-13 g/kg), moisture (37-90 g/kg) and ash (53-62 g/kg), but high in ether extract (269-355 g/kg), neutral detergent fibre (532-768 g/kg), acid detergent fibre (375-548 g/kg) and lignin (219 g/kg). The in vitro dry matter digestibility values were low for the samples from both sources, but the large scale factory-processed PPF had higher in vitro dry matter digestibility (0.215 vs 0.166) and in vitro organic matter digestibility (0.196 vs 0.145). Defatting the PPF and treating it with 5% NaOH solution significantly (p < 0.01) improved both the dry matter and organic matter digestibility. Washing the NaOH-treated PPF resulted in a higher digestibility of dry matter as against NaNW or NAD. These results suggest that defatting and treatment with 5% NaOH would improve the feeding value of PPF. PMID- 11254077 TI - Clinically manifested major health problems of crossbred dairy herds in urban and periurban production systems in the central highlands of Ethiopia. AB - The major clinical diseases of crossbred dairy herds were investigated for two years in a milk shed in Addis Ababa. Animals in 38 herds were randomly selected and visited weekly. Diagnosis of diseases and causes of death were based on clinical observation. Disease conditions were categorized into 8 groups, the mean annual incidence for all diseases being 44.7%. Reproductive diseases and clinical mastitis were most frequently observed, whereas gastrointestinal disorders, respiratory tract diseases, locomotor disorders and metabolic diseases each occurred in less than 5% of the cattle. Specific infectious and miscellaneous disease conditions each had annual incidence of about 6%. Cows and young stock were most affected and diseases were more frequent in urban situations than in periurban situations. Herd size and season significantly influenced the incidence of disease but study year did not. The crude mortality rate was 4.2%. PMID- 11254078 TI - An outbreak of Salmonella enteritidis infection in pygmy hogs (Sus salvanius). AB - An outbreak of salmonellosis was recorded in captive pygmy hogs (Sus salvanius), a critically endangered species of mammal. Of 42 captive animals maintained for conservation breeding by the Pygmy Hog Conservation Programme, Guwahati, Assam, India, 7 (16.67%) died within 3 days. The organism associated with this outbreak was identified as Salmonella enteritidis. The organisms were highly susceptible to chloramphenicol, gentamicin, norfloxacin and cefotaxim but were resistant to ampicillin, oxytetracycline, mezlocillin and sulfamerazin. The strain belonged to phage type 13a/7 and harboured two plasmids (38 and 44 megadaltons). The organisms were enterotoxigenic in CHO cell assay and were found to carry stn, sef and pef genes. PMID- 11254079 TI - Oscillatory model of novelty detection. AB - A model of novelty detection is developed which is based on an oscillatory mechanism of memory formation and information processing. The frequency encoding of the input information and adaptation of natural frequencies of network oscillators to the frequency of the input signal are used as the mechanism of information storage. The resonance amplification of network activity is used as a recognition principle for familiar stimuli. Application of the model to novelty detection in the hippocampus is discussed. PMID- 11254081 TI - Global dynamics of a network of stochastic neurons maximizes local mutual information. AB - We define a stochastic neuron as an element that increases its internal state with probability p until a threshold value is reached; after that its internal state is set back to the initial value. We study the local information of a stochastic neuron between the message arriving from the input neurons and the response of the neuron. We study the dependence of the local information on the firing probability alpha of the synaptic inputs in a network of such stochastic neurons. The values of alpha obtained in the simulations are the same as those obtained theoretically by maximization of local mutual information. We conclude that the global dynamics maximizes the local mutual information of single units, which means that the self-selected parameter value of the population dynamics is such that each neuron behaves as an optimal encoder. PMID- 11254080 TI - Independent neurons representing a finite set of stimuli: dependence of the mutual information on the number of units sampled. AB - We study the capacity with which a system of independent neuron-like units represents a given set of stimuli. We assume that each neuron provides a fixed amount of information and that the information provided by different neurons has a random overlap. We derive analytically the dependence of the mutual information between the set of stimuli and the neural responses on the number of units sampled. For a large set of stimuli, the mutual information rises linearly with the number of neurons and later saturates exponentially at its maximum value. PMID- 11254082 TI - Properties of synaptic transmission and the global stability of delayed activity states. AB - The influence of synaptic channel properties on the stability of delayed activity maintained by recurrent neural networks is studied. The duration of excitatory post-synaptic current (EPSC) is shown to be essential for the global stability of the delayed response. The NMDA receptor channel is a much more reliable mediator of the reverberating activity than the AMPA receptor, due to a longer EPSC. This allows one to interpret the deterioration of the working memory observed in NMDA channel blockade experiments. The key mechanism leading to the decay of the delayed activity originates in the unreliability of synaptic transmission. The optimum fluctuation of the synaptic currents leading to the decay is identified. The decay time is calculated analytically and the result is confirmed computationally. PMID- 11254083 TI - Efficient temporal processing with biologically realistic dynamic synapses. AB - Synapses play a central role in neural computation: the strengths of synaptic connections determine the function of a neural circuit. In conventional models of computation, synaptic strength is assumed to be a static quantity that changes only on the slow timescale of learning. In biological systems, however, synaptic strength undergoes dynamic modulation on rapid timescales through mechanisms such as short term facilitation and depression. Here we describe a general model of computation that exploits dynamic synapses, and use a backpropagation-like algorithm to adjust the synaptic parameters. We show that such gradient descent suffices to approximate a given quadratic filter by a rather small neural system with dynamic synapses. We also compare our network model to artificial neural networks designed for time series processing. Our numerical results are complemented by theoretical analyses which show that even with just a single hidden layer such networks can approximate a surprisingly large class of nonlinear filters: all filters that can be characterized by Volterra series. This result is robust with regard to various changes in the model for synaptic dynamics. PMID- 11254084 TI - Competition in the development of nerve connections: a review of models. AB - The establishment and refinement of neural circuits involve both the formation of new connections and the elimination of already existing connections. Elimination of connections occurs, for example, in the development of mononeural innervation of muscle fibres and in the formation of ocular dominance columns in the visual cortex. The process that leads to the elimination of connections is often referred to as axonal or synaptic competition. Although the notion of competition is commonly used, the process is not well understood-with respect to, for example, the type of competition, what axons and synapses are competing for, and the role of electrical activity. This article reviews the types of competition that have been distinguished and the models of competition that have been proposed. Models of both the neuromuscular system and the visual system are described. For each of these models, the assumptions on which it is based, its mathematical structure, and the extent to which it is supported by the experimental data are evaluated. Special attention is given to the different modelling approaches and the role of electrical activity in competition. PMID- 11254085 TI - Detection of ICAM-1 in experimentally induced colitis of ICAM-1-deficient and wild-type mice: an immunohistochemical study. AB - Adhesion molecules (e.g. ICAM-1, CD 54) are known to be upregulated on activated vascular endothelial cells during inflammatory reactions. To study the role of ICAM-1 in intestinal inflammation in vivo, we induced acute experimental colitis in wild-type (C57BL/6) mice and ICAM-1-deficient mice, by feeding the animals with 3% dextran sodium sulphate (DSS) in drinking water for 7 days. In the control strain the immunohistochemical staining showed a very pronounced endothelial upregulation of ICAM-1 after the DSS treatment observed in areas of inflammatory infiltrate, especially in venules or arterioles of the propria and submucosa, and partly in the mesocolon. DSS-fed ICAM-1-deficient mice showed no endothelial enhancement and only faint staining of venules or capillaries approaching that encountered in the control ICAM-1-deficient animals. Our data indicate that ICAM-1 may play a crucial role in the development of acute intestinal inflammation, consistent with our finding that ICAM-1 deficiency can obviate severe forms of experimentally induced colitis in mice. PMID- 11254086 TI - Characterization of monoclonal antibodies to glycodelin and recombinant glycodelin. AB - Glycodelin-A belongs to the lipocalin superfamily. Although it is associated with normal endometrial growth during the menstrual cycle, fertilization and normal pregnancy in humans, the molecular mechanism of its biological action has not been elucidated. To undertake studies to understand the functional relevance of any molecule, obtaining large quantities of the protein becomes essential. With the ultimate aim of purifying glycodelin either from its natural sources (human amniotic fluid) or the recombinant glycodelin from bacterial recombinant lysates, we raised monoclonal antibodies to this protein. As immunogens, recombinant glycodelin expressed in E. coli and Pichia pastoris as well as glycodelin from amniotic fluid were used. The monoclonal antibodies generated were characterized with respect to binding to both the native as well as the recombinant proteins using ELISA, immunoblotting, and immunohistochemistry. PMID- 11254087 TI - Lectin histochemical identification of the carbohydrate moieties on N- and O linked oligosaccharides in the duct cells of the testis of an amphibian urodele, the spanish newt (Pleurodeles waltl). AB - The aim of the present work was to study the carbohydrate moieties present on N- and O-linked oligosaccharides of duct cells of a urodele amphibian testis, by means of lectin histochemistry. It was found that duct cells have a carbohydrate composition that includes alpha(1,3)-, alpha(1,4)- or alpha(1,6)-linked Fuc and Man on N-linked oligosaccharides, Gal and GlcNAc on O-linked oligosaccharides, and DBA-positive GalNAc, alpha(1,2)-linked Fuc and Neu5Ac alpha(2,3)Gal beta(1,4)GlcNAc on both N- and O-linked oligosaccharides. All the duct cells showed the same lectin labelling pattern, the only exception being some sparse duct cells that showed the sequence Neu5Ac alpha(2,6)Gal/GalNAc. The possible roles of duct cells in sperm maturation and the hypothesis for a common origin of duct and follicle (Sertoli) cells in the urodele testis are discussed. PMID- 11254088 TI - Atrial natriuretic peptide and guanylin-activated guanylate cyclase isoforms in human sweat glands. AB - The ultracytochemical localization of membrane-bound guanylate cyclases A and C, stimulated by atrial natriuretic peptide and guanylin respectively, has been studied in human sweat glands. The results showed that the peptides stimulated guanylate cyclases A and C in both eccrine and apocrine glands. In the secretory cells, enzymatic activity was present on the plasma membranes and on intracellular membranes involved in the secretory mechanism. In eccrine glands, the cells of the excretory duct also presented enzymatic activity on the plasma membranes. In both glands, myoepithelial cells, surrounding the secretory cells, exhibited only guanylate cyclase A activity. These localizations of enzymatic activity suggest a role for both atrial natriuretic peptide and guanylin in regulating glandular secretion. PMID- 11254089 TI - An immunocytochemical study of the pituitary gland of the white seabream (Diplodus sargus). AB - The adenohypophysis of the white seabream (Diplodus sargus) was studied using histochemical and immunocytochemical techniques. The adenohypophysis was composed of rostral pars distalis, proximal pars distalis and pars intermedia. Prolactin (anti-chum salmon prolactin positive) and adrenocorticotropic (anti-human ACTH positive) cells were found in the rostral pars distalis. Prolactin cells were organized into follicles, while ACTH cells were arranged in cords around neurohypophyseal tissue branches that penetrated the rostral pars distalis. In the proximal pars distalis, somatotropic (anti-chum salmon and anti-gilthead seabream growth hormone positive), gonadotropic (anti-chum salmon beta gonadotrophin II and anti-carp beta-gonadotrophin II positive, but anti-chum salmon beta-gonadotrophin I negative) and thyrotropic (anti-human beta thyrotropin positive) cells were observed. Growth hormone cells were restricted to the dorsal and ventral part of the proximal pars distalis. They were clustered or surrounded the neurohypophyseal branches. Only one type of gonadotrophin cell was identified and they were clustered or isolated in the proximal pars distalis. Scattered groups of thyrotropin cells were located throughout the proximal pars distalis. In the pars intermedia somatolactin (anti-chum salmon and anti-gilthead seabream somatolactin positive) and melanotropic (anti-alpha-melanotropic hormone positive) cells were localized. In addition, gonadotrophin cells surrounded the pars intermedia or distributed evenly between somatolactin and melanotropic hormone cells. Somatolactin cells were periodic acid-Schiff negative and surrounded the neurohypophyseal branches intermingled with melanotropic cells. These cells were also immunoreactive to anti-human ACTH antiserum. PMID- 11254090 TI - Irregular costameres represent nitric oxide synthase-1-positive sarcolemma invaginations enriched in contracted skeletal muscle fibres. AB - NADPH diaphorase histochemistry and NOS-1 immunohistochemistry on 60 microm thick frozen sections of rat extensor digitorum longus muscles led to the detection of prominent rings clearly encompassing the surface of the muscle fibres. These so far unknown costameres were usually found as doublets flanking a space of about 2 microm width. Because these costameric doublets did not appear in regular periods, we designate them irregular costameres to discriminate them from regular ones with a 1 microm periodicity overlying Z-discs and M-lines. Irregular costameres were thicker than the regular ones and free of intercostameres. Immunohistochemistry demonstrated that NOS-1 was co-localized with integral (beta dystroglycan, alpha-sarcoglycan) and peripheral (caveolin-3, dystrophin) members of the enlarged dystrophin complex in the irregular costameres but not with non sarcolemmal organized proteins (myosin heavy chain, alpha-actinin, desmin and sarcoplasmic reticulum-located Ca2+-dependent ATPase-1). Invaginations of the sarcolemma to form irregular costameres were observed. In teased myofibres the sarcolemma between two following irregular costameres was ballooned, while the irregular costameres themselves clamped the fibres together. Finally, the number of detectable irregular costameres was significantly increased in maximally contracted extensor digitorum longus muscles generated by electric stimulation but decreased in mechanically stretched ones. Combining these observations, we hypothesize that irregular costameres belong to a reserve zone for the sarcolemma necessary for the contraction/relaxation cycle in myofibres. PMID- 11254091 TI - The distribution of carbonic anhydrase II in human, pig and rat oesophageal epithelium. AB - Carbonic anhydrase II (CA II) is present in human oesophageal epithelial cells and probably involved in protecting the mucosa against acidic gastric refluxate. If this is the case, then it is likely that the enzyme will be more concentrated at or near the gastro-oesophageal junction. To answer this question, and determine whether CA II is present and similarly distributed in other species, we also examined the oesophageal epithelium of the rat and pig. In the rat, CA II was largely absent from the oesophageal epithelium, but present in the stratified squamous epithelium of the gastric forestomach as an approximately 2 mm-long collar around the entrance to the corpus, a site that roughly corresponds to the gastro-oesophageal junction in other animals. The enzyme was present mainly in basal and prickle cells. In upper and middle pig oesophagus, CA II was largely confined to basal cells and isolated groups of stratified superficial prickle cells. CA II-containing epithelial cells were highly concentrated in the thickened epithelium at the gastro-oesophageal junction (about four-times thicker than upper or middle). Reactive cells were present throughout the depth of the epithelium, but noticeably more concentrated in the basal and superficial prickle cell layers. CA II was also prominent in the most superficial cell layers in islands of the oesophageal mucosa within the gastric cardia. In man, CA II was confined largely to the basal half of the epithelium in the upper and middle regions of oesophagus. The distribution of CA II at the gastro-oesophageal junction took different forms. In general, there were more CA II-reactive cells at or closer to the lumen. The superficial prickle cell layers tended to exhibit more CA II than the deeper layers, with basal and epibasal cells containing little or no enzyme. In other regions of the same specimens, CA II-containing cells were present from the basal to the most luminal layers. If CA II in oesophageal epithelial cells in the region of the gastro-oesophageal junction (or in the case of the rat the forestomach/corpus junction) is important in the defence against refluxate, then it is in a vulnerable site, since bile salts are potent inhibitors of the enzyme. The action of bile salts on CA II may be an important factor in the initiation of oesophageal disease. PMID- 11254093 TI - ELISA and direct immunofluorescence test to detect equine arteritis virus (EAV) using a monoclonal antibody directed to the EAV-N protein. AB - A monoclonal antibody (mAb) directed against the equine arteritis virus (EAV) nucleocapsid (N) protein was used for indirect enzyme-linked immunosorbent assays (ELISAs) using viral antigen from different sources. The same mAb was labelled with fluorescein isothiocyanate for direct immunofluorescence tests (DIFTs). The N-specific mAb appeared to be suitable for the detection in both ELISA and DIFT of different EAV strains and field isolates from semen and tissue samples after passage in lines of RK-13, Vero and fetal equine kidney cells. The ELISA described is an easy and fast method which can be used in most cases to replace the microneutralization test to prove the EAV specificity of the cytopathic effect of cell cultures. The DIFT, however, is more sensitive than both the ELISA and the microneutralization test because EAV antigen can be detected even in cell cultures without or with very weak cytopathic effect. PMID- 11254092 TI - Appropriate cytochemical controls for differentiating calcium-specific ATPase from ecto-ATPase. PMID- 11254094 TI - A new product with formic acid for Varroa jacobsoni Oud. control in Argentina. I. Efficacy. AB - An organic product based on formic acid in a gel matrix was evaluated for use in Varroa control under autumnal climatic conditions in Argentina. Twenty colonies each received two gel packets with formic acid in two applications and numbers of falling mites were registered. After this treatment colonies received two other acaricides in order to compare efficacy. Average final efficacy in colonies treated with the organic product was 92% with a low variability. The gel matrix kept an adequate formic acid concentration inside the colonies with only two applications. This product is, therefore, a good alternative for Varroa control because it is organic, easy to use and presents a low variability in final efficacy between colonies. No queen, brood, or adult honeybee mortality was registered. PMID- 11254095 TI - Effect of endobronchial challenge with Actinobacillus pleuropneumoniae serotype 9 of pigs vaccinated with a vaccine containing Apx toxins and transferrin-binding proteins. AB - The efficacy of a subunit vaccine containing the Apx toxins of Actinobacillus pleuropneumoniae and transferrin-binding proteins was determined. Ten pigs were vaccinated twice with the vaccine. Eight control animals were injected twice with a saline solution. Three weeks after the second vaccination, all pigs were endobronchially inoculated with 10(6.5) colony-forming units (CFU) of an A. pleuropneumoniae serotype 9 strain. In the vaccine group, none of the pigs died after inoculation. Only one pig of the control group survived challenge. Surviving pigs were killed at 7 days after challenge. The mean percentage of affected lung tissue was 64% in the control group and 17% in the vaccine group. Actinobacillus pleuropneumoniae was isolated from the lungs of all animals. The mean bacterial titres of the caudal lung lobes were 5.0 x 10(8) CFU/g in the control group and 3.0 x 10(6) CFU/g in the vaccine group. It was concluded that the vaccine induced partial protection against severe challenge. PMID- 11254096 TI - Comparison of methods for the determination of antimicrobial resistance in Staphylococcus aureus from bovine mastitis. AB - The results of three standard methods (broth dilution, agar dilution, disk diffusion) and an experimental modification of the microdilution method for determination of resistance to ampicillin, cephalotin, cloxacillin, neomycin, novobiocin, penicillin and streptomycin were compared using 151 Staphylococcus aureus isolates obtained from cases of mastitis. The accuracy of the dilution methods was compared by determination of minimum inhibition concentrations (MIC, MIC50, MIC90 and modal MIC) and by assessment of the agreement within the tolerance of +/-1 dilution step in 2-fold dilution series. The results of the dilution methods were further compared with those of the reference disk diffusion method and the strains were classified as sensitive or resistant using the interpretation criteria for human strains. The comparisons indicated that MIC characteristics and the final classification as sensitive or resistant were method-dependent. Resistance to aminoglycoside antibiotics was observed more often when using broth dilution methods, especially when the broth was supplemented with lactose. PMID- 11254098 TI - Arthritis after experimental infection with Yersinia enterocolitica 0:3 in rabbits. AB - Arthritis in rabbits was caused after experimental oral infection with Yersinia enterocolitica (serotype 0:3, biotype 4, pYV+). Clinical and laboratory signs, bacterial dissemination to the viscera, immune response and morphological findings were studied from day 1 to day 40 post-infection (p.i.). Augmentation of body temperature and erythrocyte sedimentation rate occurred on day 1, and on day 8 p.i. was accompanied by leucopenia. The number of alveolar macrophages was increased up to the 15th day p.i., in contrast to peritoneal macrophage numbers. Extensive bacterial colonization of the internal organs was detected at necropsy until the end of the experiment. Analysis of the cell immune response revealed activation of B cells in peripheral blood, spleen and thymus as well as augmentation of T-cell number in the lymphoid organs examined on days 15, 28 and 40 p.i. Histological changes typical of a generalized infection, such as purulent meningoencephalitis, catarrhal pneumonia and lymphadenitis, were observed. Clinical and morphological manifestations of arthritis were also established. The results obtained show that Y. enterocolitica (serotype 0:3, pYV+) induces a generalized, non-lethal infection in Chinchilla rabbits, complicated by arthritis. PMID- 11254097 TI - Variations among unbred heifers in the activities of polymorphonuclear leucocytes from the mammary gland and blood. AB - The phenotypic characteristics are described for the activity of polymorphonuclear leucocytes NMN) obtained by either lavage of the cavity system of juvenile mammary glands stimulated with a synthetic muramyl dipeptide analogue or isolation from the peripheral blood. Attention was paid to the variability of characteristics and its sources, and to correlations among them. The following characteristics were investigated in 27 clinically healthy, unbred Bohemian Red Pied x Holstein heifers: migration activity in situ, number of phagocytosing PMN, phagocytotic index, bactericidal activity of PMN and unstimulated and zymosan stimulated luminol-dependent chemiluminescence. Considerable individual variation was found in the characteristics. Significant differences between blood PMN and PMN from lavages after influx induction were found for bactericidal activity (P < 0.05) and chemiluminescence (P < 0.01). A significant correlation between blood PMN and mammary gland PMN was found only for the number of phagocytosing cells (r = 0.329; P < 0.01). Highly significant positive correlations (P < 0.01) were demonstrated between the number of phagocytosing PMN [a], phagocytotic index [b], and bactericidal activity [c] in both blood PMN (r(ab) = 0.602; r(ac) = 0.565; r(bc) = 0.529) and mammary gland PMN (r(ab) = 0.730, r(ac) = 0.618, r(bc) = 0.589). No significant correlation was demonstrated for non-stimulated (NS), zymosan-stimulated (ZS), or opsonized zymosan-stimulated (OZS) chemiluminescence with any of the other characteristics of phagocytotic activity, in either blood PMN or mammary gland PMN (P > 0.05). The animal was a highly significant source of variability for all the phagocytotic activity characteristics (P < 0.01). Udder quarter was a non-significant source of variability for all the characteristics of phagocytotic activity except for NS chemiluminescence (P < 0.05) and ZS or OZS chemiluminescence (P < 0.01). However, udder quarter was a non-significant source of variability of chemiluminescence indices ZS/NS and OZS/NS (P > 0.05). It has been demonstrated that in situ migration activity, the number of phagocytosing PMN, phagocytotic index, bactericidal activity of PMN and chemiluminescence indices of PMN collected from juvenile mammary glands of unbred heifers after influx induction can be regarded as candidate early markers of resistance to mammary infections. PMID- 11254100 TI - Prevalence of eae and shiga toxin genes among Escherichia coli strains isolated from healthy calves. AB - Strains of Escherichia coli (n = 390) isolated from 132 healthy, 4-8-week old calves, were tested by polymerase chain reaction (PCR) for the eae (intimin) gene and shiga toxin genes (stx1 and stx2). All strains were also analysed for F5, F17 and F41 fimbriae and for the heat-labile (LT) and heat-stable (STI and STII) genetic markers. Overall, the eae gene was detected in 84 (21.5%) of the strains tested. Only 21 (5.4%) isolates were positive for stx1 (18 strains) or stx2 (three strains); nine of the stx1-positive isolates also possessed the eae gene. A high percentage (29.2%) of the isolates tested expressed F17 but no enterotoxin genes were detected. None of the eae- or stx-positive strains belonged to the O157 serogroup. PMID- 11254099 TI - Aspects of the transmission of protection against Mycoplasma hyopneumoniae from sow to offspring. AB - The aims of this study were to describe the variation in concentration of antibodies to Mycoplasma hyopneumoniae in the serum and colostrum of sows, and to compare the amount of antibodies in colostrum with that obtained in the serum of the smallest piglets in a litter. In addition, the efficacy of the passive immunity in natural conditions was studied. The study was performed in a sow pool herd (600 sows) that was endemically infected with M. hyopneumoniae. Blood samples were collected from sows 19 days (n = 25) before and 3 days (n = 15) after farrowing, and a colostrum sample (n = 25) was collected on the day of farrowing. All samples were analysed for antibodies to M. hyopneumoniae with a monoclonal blocking enzyme-linked immunosorbent assay (ELISA). Twelve sows (48%) were high-responders with respect to antibody concentration in colostrum. The amount of blocking decreased in serum during the last weeks of pregnancy and 3 days post-farrowing it was only 53% of the level found in colostrum. At the age of 14 days, 30 of the smallest piglets were weaned. They were divided into three experimental groups, being the offspring of high-responding sows, low-responding sows, or a mix of high- and low-responding sows. The groups were transported to three separated isolation units and were followed until slaughter. At slaughter, lung lesions were not found. Nor could M. hyopneumoniae be demonstrated either by cultivation or by polymerase chain reaction. However, a significant increase in absorbance values, assessed by an indirect-ELISA, was demonstrated in groups established from low-responding sows. It was concluded that a high antibody level in colostrum appeared to protect piglets from M. hyopneumoniae. PMID- 11254101 TI - Computerized comparison of the protein compositions of Erysipelothrix rhusiopathiae and Erysipelothrix tonsillarum strains. AB - Protein profiles of six Erysipelothrix rhusiopathiae strains, five Erysipelothrix tonsillarum strains and three Erysipelothrix strains of uncertain taxonomic position were studied by sodium dodecyl sulphate-polyactylamide gel electrophoresis (SDS-PAGE). In a computerized comparison of the protein patterns of the strains, the level of similarity between the strains was determined. The SDS-PAGE protein bands were divided into 14 groups based on molecular weight. The relative distribution of proteins within these groups was used to characterize the strains. These distribution patterns were analysed by computing Pearson's correlation coefficient between strains, and by cluster analysis based on Euclidean distances and the unweighted pair-group method of arithmetic averages (UPGMA). The geometric mean of the similarities calculated by Pearson's correlation coefficient was 0.980 +/- 0.018 between the E. rhusiopathiae strains and 0.979 +/- 0.013 for E. tonsillarum strains. The value was 0.932 +/- 0.036 between the strains belonging to different species. However, a threshold value applicable for identification of a given strain to a species could not be established. Of the three strains of uncertain taxonomic position, the strains designated Rotzunge and Iszap 4 had a protein composition more similar to that of E. tonsillarum than to that of the E. rhusiopathiae type strain. The strain designated Pecs 56, which may be a member of a new species according to literature data, gave inconsistent results by the two methods used. The computerized evaluation method developed here is suitable for the comparison of the protein composition of the strains and for the construction of the protein similarity tree by cluster analysis. PMID- 11254102 TI - Cloning and analysis of the mouse follistatin promoter. AB - Follistatin is a secreted protein, which functions as an antagonist of different members of the TGF-beta superfamily, including activin and bone morphogenetic proteins. Expression of follistatin is tightly regulated during mouse development both spatially and temporally. In order to study the regulation of follistatin expression in the mouse embryo we have cloned and analyzed part of the 5' flanking region of the murine follistatin gene. Primer extension and RNase protection assays demonstrate that the murine follistatin promoter region has at least three distinct transcription initiation sites, which are each preceded by a TATA box. All of the transcription initiation sites are located within the first 500 bp upstream of the translational start site. Sequence analysis of this 500 bp region revealed several consensus binding sites for transcription factors including AP-1, Brachyury-T, CREB, Sp1, AP-2 and Tcf. To test whether the 5' region displays promoter activity, we transfected various 5' flanking region deletion constructs into F9 embryonal carcinoma (EC) cells and into P19 EC cells. In these two cell lines a region of only 262 bp upstream of the translation start site could drivereporter expression in a manner that reflects endogenous mRNA expression. PMID- 11254103 TI - The evolution of glutamate synthase. AB - DNA coding for the ferredoxin-dependent glutamate synthase (EC 1.4.7.1) of spinach chloroplasts has been cloned and sequenced. It consists of 5015 bp and starts with the codon for the N-terminal cysteine of the mature protein. Ferredoxin-dependent glutamate synthase is one of the key enzymes in the early stages of ammonia assimilation in plants, algae and cyanobacteria. In addition to the ferredoxin-dependent enzyme, there are two other forms of glutamate synthase, one of which uses NADH as the electron donor and a second that uses NADPH. Although all three forms catalyze the reductive transamidation of the amido nitrogen from glutamine to 2-oxoglutarate to form two molecules of glutamate, ferredoxin-dependent glutamate synthases differ from the NADH and NADPH-dependent forms in subunit composition and amino acid sequence. The recent availability of sequence data for glutamate synthases from spinach and from two archael species has produced a clearer and more detailed picture of the evolution of this key enzyme in nitrogen metabolism and the origins of the two subunit/domain structure of the enzyme. PMID- 11254104 TI - Identification of a approximately 30S size non-ribosomal Saccharomyces cerevisiae RNA that is rapidly labeled on its 3' end by ATP or UTP. AB - Cell-free extracts prepared from S. cerevisiae cells were incubated in the presence of [alpha-32P]-labeled ATP, CTP, GTP or UTP. An RNA larger than ribosomal 25S RNA with an apparent size of approximately 30S was prominently labeled on its 3' end in the presence of ATP or UTP but not with CTP or GTP. This labeled RNA was not hybrid-selected by cloned yeast ribosomal DNA; in addition, this approximately 30S RNA was not cleaved by RNase H in the presence of complementary deoxyribooligonucleotides to rRNA. These two lines of evidence show that this approximately 30S RNA is not structurally related to ribosomal RNA gene repeat. The cell-free extracts prepared from yeast cells containing temperature sensitive poly(A) polymerase adenylated this novel yeast RNA at restrictive temperature with efficiency similar to extracts prepared from wild-type yeast cells. These data show that the enzyme responsible for adenylation of this approximately 30S RNA is distinct from mRNA poly(A) polymerase. While the human SRP RNA 3' adenylating enzyme in the HeLa cell extract adenylated human SRP or Alu RNAs, the yeast adenylating enzyme did not adenylate the human SRP or Alu RNAs in vitro; these data indicate species specificity for this adenylating enzyme. PMID- 11254105 TI - Characterization of human and mouse H19 regulatory sequences. AB - H19 is expressed in a large percentage of bladder tumors, but not expressed in healthy bladder tissue. The aim of this study is to define H19 optimal transcriptional regulatory sequences in tumor cells, which can potentially be used to control expression of a toxin gene in constructs to be used in bladder cancer gene therapy trials in mice and human. Transient expression assays revealed that elements responsible for promoter activity are contained within the 85 bp upstream region. The transcriptional activity of this region was strongly inhibited by the methylation of the Hpa II sites. A modest cell specificity is conferred by the upstream sequences. The human and murine promoter activities were significantly increased by the human H19 4.1 kb enhancer sequence. The 85 bp H19 upstream region contains all the elements to interact with the enhancer. We showed that the human H19 promoter is highly active in a murine bladder carcinoma cell line, justifying its use to drive the expression of a cytotoxin gene in gene therapy trials in mice. PMID- 11254106 TI - Characterization of 5'-flanking region of human aggrecanase-1 (ADAMTS4) gene. AB - Aggrecanase-1, also known as ADAMTS4 (a disintegrin and metalloproteinase with thrombospondin motifs 4), cleaves at the Glu373-Ala374 site of aggrecan, thereby indicating aggrecan degradation. It is thought that ADAMTS4 plays a pivotal role in inflammatory joint diseases and cartilage degradation. To elucidate the mechanisms of regulation of ADAMTS4 gene expression, we cloned the 5'-flanking region of the human ADAMTS4 gene and characterized its promoter activity by means of reporter assay using porcine chondrocytes and NIH3T3 cells. Reporter gene analysis using deletion variants suggested that the region between -383 and +10 relative to the tentative transcription start site is necessary for full promoter activity; this region contains one Sp1 and three AP2 sites. In addition, the segment between -726 and -384 appears to contain silencer element(s). A complete deletion mutant of the nuclear factor I (NFI) binding site at -441 to -429 resulted in recovery of the promoter activity in chondrocytes, but not in NIH3T3 cells. Thus, the NFI site is involved in negative regulation of the human ADAMTS4 promoter activity in chondrocytes. PMID- 11254108 TI - DNA-protein cross-linking in nuclei of immature and mature chicken erythrocytes. AB - DNA-protein cross-linkages were formed in isolated nuclei from immature and mature chicken erythrocytes by reaction with cis-diammine dichloroplatinum. On the basis of electrophoretic behaviour, the most abundant proteins involved in the cross-linking appeared to be present also in preparations of nuclear matrix. The maturation of the erythrocyte, which is accompanied by transcriptional inactivation, leads to a decrease in the amount of DNA-interacting proteins, to a loss of proteins capable of a specific recognition of DNA sequences and, unexpectedly, to the appearence of some new DNA-protein interactions. At least three cross-linked proteins were found predominantly or exclusively in nuclei of immature cells, and three others in those of mature ones. The three DNA-bound proteins, typical of mature erythrocytes, were not found among the components of a high-salt preparation of nuclear matrix. The results obtained suggest that, in addition to the well-known histone H5 and MENT protein, these newly identified DNA-bound proteins contribute to the formation of the condensed, inactive chromatin characteristic of mature erythrocyte. PMID- 11254107 TI - Differential scanning calorimetry of chromatin at different levels of condensation. AB - The thermal denaturation of calf thymus total chromatin and of fractions enriched in heterochromatin or euchromatin, has been investigated by differential scanning calorimetry and compared to that of calf thymus DNA and DNA-histone complexes. In our experimental conditions, chromatin melts in three thermal transitions: the main one, assigned to separation of the DNA double helix, occurs at 83 degrees C, while the other two occur at 63 degrees C and 74 degrees C. The data show that: (a) the transition enthalpy for denaturation of DNA in the total chromatin and in DNA-histone complexes is nearly the same as that of DNA in solution; (b) the transition at 63 degrees C is present in the thermogram of the heterocromatin enriched fraction, while it is completely absent in that of the euchromatin enriched one. The results suggest that this transition can be attributed to the higher order structures of heterochromatin. PMID- 11254109 TI - Human SP1 but not human AP1 binding to the TGF-beta element in the 5' flanking region of the rat PROalpha1(I) collagen gene. AB - The consensus TGF-beta element (TGCCCACGGCCAG) located at approximately -161Obp from the start site of transcription of the rat pro alpha1(I) collagen gene has recently been shown to be required for the basal promoter activity of this gene (Meisler et al., J. Cell Biochem. 75: 196, 1999). Site directed mutation of this TGF-beta element resulted in almost complete abolishment of the basal promoter activity of the fibroblasts transfected with the 3.6 ColCat plasmid which contains a 3.6 kb portion of the 5' flanking region of the rat pro alpha1(I) collagen gene linked to the reporter gene, chloramphenicol acetyltransferase (CAT). Southwestern analysis of the nuclear protein binding to the TGF-beta element revealed a 34,000 Da complex while after UV-crosslinking, studies revealed a TGF-beta element nuclear protein complex of 82,000 Da (Ritzenthaler et al., J. Biol. Chem. 268: 13625, 1993). Thus, a multiple protein TGF-beta DNA element complex may exist which may promote the transcription of the rat pro alpha1(I) collagen gene. Since literature findings indicate that a nuclear factor interacts with an SP1-like binding site of the human pro alpha1(I) collagen promoter and an AP-1 binding sequence has been shown to be involved in the regulation of the human pro alpha2(I) collagen gene and both these binding sequences are TGF-beta1 responsive, we determined whether the TGF-beta element located in the 5' flanking region of the rat pro alpha1(I) collagen gene formed complexes with either of these nuclear factors or both. PMID- 11254110 TI - Responses to ultraviolet-B in cell lines from hereditary melanoma kindreds. AB - Ultraviolet-B (UV-B) triggers a cascade of events involving cell cycle control genes leading ultimately to DNA repair or apoptosis. The hypothesis examined here is that the genetic abnormality predisposing to melanoma affects the ability of the cell to respond appropriately to UV-B, so favouring mutagenesis. Epstein-Barr virus-transformed lymphoblastoid cell lines from hereditary melanoma kindreds were irradiated with UV-B, and changes in p53, p21 and Bcl-2 expression and cell cycle phase distribution were analysed. Twenty-two cell lines were tested: 12 carriers of melanoma susceptibility and 10 non-carriers (unaffected first degree relatives). At 24 h after irradiation with 50 J/m2, 15 of the 22 cell lines showed a rise in G2/M. After 400 J/m2, all the cell lines showed a reduction or loss of G2/M and 17 of the 22 showed an S phase delay. More carriers than noncarriers of melanoma susceptibility showed significant S phase delay after 50 J/m2 (seven out of 12 carriers versus two out of 10 non-carriers). Six of the 10 pairs (carrier versus non-carrier) tested showed discordant cell cycle responses; however the nature of the difference was not universal. Bcl-2 reduction was seen 4 h post-irradiation in all the carriers and non-carriers. The p53 and p21 responses, although showing some individual variations, were not related to carrier status. These results show individual variations in response to UV-B irradiation among cell lines from the members of hereditary melanoma kindreds, but no consistent differences between carriers and non-carriers of melanoma susceptibility. PMID- 11254111 TI - The MEK1 inhibitor PD98059 sensitizes C8161 melanoma cells to cisplatin-induced apoptosis. AB - The regulation of apoptosis is believed to be dependent on the balance of the activities of different intracellular signalling systems. Activation of the SAPK/JNK pathway is implied in pro-apoptotic signalling, while activation of the MEK1/ERK pathway may have a viability-promoting effect. We show here that treatment with the MEK1 inhibitor PD98059 sensitizes the human melanoma cell line C8161 to cisplatin-induced apoptosis. In these cells, cisplatin at 40 microM did not elicit significant cell death, whereas massive cell death was seen when cells were pretreated for 20 h with 40 microM PD98059 before the addition of cisplatin. Concomitant addition of PD98059 and cisplatin did not have any sensitizing effect, and PD98059 on its own did not induce apoptosis. However, in three other human melanoma cell lines PD98059 did not potentiate cisplatin-induced apoptosis. Instead, in one of these cell lines (AA), PD98059 protected against cisplatin induced cytotoxicity. We conclude that blocking of the MEK1/ERK pathway may, in some instances, potentiate the cytotoxic effect of cisplatin on human melanoma cell lines, whereas in other instances it may have a protective effect. Thus it cannot be regarded as a general approach to sensitizing melanoma cells to drug induced apoptosis. PMID- 11254113 TI - Automated skin lesion screening--a new approach. AB - Automated melanoma diagnosis is a popular focus of research, with numerous papers describing techniques and results. In our study, we identified two possible problems with the current method of automated diagnosis, where systems are intended to reproduce histopathology results. We propose a new method of identifying problematic skin lesions, namely attempting to reproduce algorithmically the perceptions of dermatologists as to whether the lesion should be excised. In the best case, our initial model reproduced the decision of dermatologists in over 80% of cases. These results suggest that reproducing the decision to excise may be a valuable adjunct to current methodology. PMID- 11254112 TI - Comparative analysis of immunocritical melanoma markers in the mouse melanoma cell lines B16, K1735 and S91-M3. AB - The mouse melanoma cell lines B16, K1735 and Cloudman S91-M3 (and various sublines) are frequently used as melanoma models. Extensive comparative data of their immunological features are not available. In order to define the immunological profiles of these cell lines, relevant tumour markers were studied. S91-M3 melanoma cells constitutively expressed high levels of major histocompatibility complex (MHC) I, in contrast to K1735-M2 and B16-F1 cells. MHC II expression was restricted to B16-F1 cells following interferon-gamma treatment. Tyrosinase, tyrosinase-related protein-2 and gp100 were detected in B16-F1 and S91-M3 cells, but not in K1735-M2 cells. Constitutive surface expression and secretion of intercellular adhesion molecule-1 was found on S91-M3 cells. No substantial secretion of interleukin-10 could be detected. In contrast, low levels of latent transforming growth factor-beta were found in the cell supernatants of B16-F1 and K1735-M2 cells. The expression pattern of Fas, FasL and FLICE inhibitory protein was comparable in all three cell lines. Thus our findings indicate that each cell line presents a characteristic immunological profile, confirming that B16-F1 is an appropriate murine tumour model for tumours with low levels of MHC I but expressing melanoma-associated antigens. S91-M3 represents a complementary, more immunogenic model. In contrast, K1735-M2 does not seem to be an appropriate model for melanoma. PMID- 11254114 TI - Differentiation between pigmented Spitz naevus and melanoma by digital dermoscopy and stepwise logistic discriminant analysis. AB - Epiluminescence light microscopy (ELM) has proven useful in the diagnosis of pigmented skin lesions (PSLs). However, in some cases this technique does not sufficiently increase the diagnostic accuracy in distinguishing pigmented Spitz naevi (PSNs) from melanoma. With the aim of obviating these problems of qualitative interpretation, methods based on the mathematical analysis of PSLs, such as digital dermoscopy analysis (DDA), have recently been developed. In the present study we used a digital dermoscope (DBDermo-MIPS, Dell'Eva-Burroni) to analyse PSNs and melanomas with similar clinical and dermoscopic features for any correlation between variables and to determine its discriminating power with respect to histological diagnosis. The 100 lesions underwent histological examination by three experienced dermatopathologists and were identified as PSNs (43) or melanomas (57). Thirty-six parameters were identified as possible discriminating variables and were grouped in four categories: geometry, colour, texture, and islands of colour. Statistical analysis was used to identify the variables with the highest discriminating power. Stepwise discriminant analysis selected only four variables: entropy, minimum diameter, red lesion value and peripheral dark (the means of these variables were higher in melanomas than in PSNs). Thus the combined use of digital dermoscopy and stepwise logistic discriminant analysis made it possible to single out the best objective variables for distinguishing PSN and melanoma. PMID- 11254116 TI - Does intensive histopathological workup by serial sectioning increase the detection of lymph node micrometastasis in patients with primary cutaneous melanoma? AB - Various histopathological techniques have been developed in order to improve the detection of micrometastasis in the regional lymph nodes of patients with malignant melanoma. Our standard histopathological examination of lymph nodes included haematoxylin and eosin (H & E) staining and immunohistochemistry (IH) using antibodies to HMB-45 and S-100 proteins of three paraffin sections at one level. In addition, lymph nodes were examined by molecular biological methods using tyrosinase reverse transcription-polymerase chain reaction (RT-PCR). In this study, we investigated the use of step sections and IH in lymph nodes regarded as negative by standard histopathology but positive by tyrosinase RT PCR, suggesting the presence of tumour cells. In a series of 76 consecutive patients with stage I and II cutaneous melanoma, a total of 156 regional lymph nodes were examined by H & E staining, IH and tyrosinase RT-PCR. All lymph nodes were bisected along their long axis for separate evaluation. In 21 patients, at least one lymph node in the regional nodal basin reported as tumour-negative by standard histopathology was demonstrated to express tyrosinase (total number of nodes = 33). These 33 lymph nodes were re-examined by H & E and IH at 10 additional levels of the paraffin block. Only one lymph node from one patient had occult melanoma cells in deeper levels detected exclusively by IH. Six out of 20 patients with positive findings exclusively on tyrosinase RT-PCR developed tumour recurrences during a median follow-up of 34 months. We therefore conclude that additional step sectioning with IH does not significantly increase the detection of tumour-positive lymph nodes. Patients with melanoma cells detected exclusively by RT-PCR, however, were shown to be at increased risk for tumour recurrence. PMID- 11254115 TI - Gamma probe guided biopsy of the sentinel node in malignant melanoma: a multicentre study. AB - Sentinel lymph node biopsy was attempted in 336 patients with clinically node negative cutaneous melanoma. All patients were injected with technetium-99m labelled radiocolloid, with 108 patients simultaneously receiving vital blue dye for sentinel node identification. Sentinel lymph nodes were identified in 329 patients, giving a technical success rate of 97.9%. Metastatic disease was identified in 39 (11.9%) of the patients in whom sentinel nodes were found. Patients with negative sentinel nodes were observed and patients with positive sentinel nodes underwent comprehensive lymph node dissection. The presence of metastatic disease in the sentinel nodes and primary tumour depth by Breslow or Clark levels were joint predictors of survival based on Cox proportional hazards modelling. Disease recurrences occurred in 26 (8.8%) patients with negative sentinel lymph nodes, with isolated regional recurrences as the first site in 10 (3.4%). No patients with Clark level II primary tumours were found to have positive sentinel nodes or disease recurrences. One patient with a thin (<0.75 mm) Clark level III primary had metastatic disease in a sentinel node. Patients with metastases confined to the sentinel nodes had similar survival rates regardless of the number of nodes involved. PMID- 11254117 TI - Detection of circulating melanoma cells in peripheral blood by a two-marker RT PCR assay. AB - The aim of this study was to develop a highly sensitive two-marker assay for the detection of circulating melanoma cells in patients' blood using a reverse transcriptase-polymerase chain reaction (RT-PCR). We analysed the usefulness of two different sets of markers: tyrosinase and MUC-18 (TYR/MUC-18), and tyrosinase and MART 1 (TYR/MART 1). Total cellular RNA was isolated from 337 blood samples from 80 melanoma patients at different stages of the disease. All patients had undergone primary surgery. Assay sensitivity and specificity were confirmed using three different melanoma cell lines and two different fibroblast lines. In addition, blood from 47 healthy subjects and 10 patients with non-melanoma cancer was used as a negative control. We found that two-marker analysis is more accurate than the single tyrosinase assay. The frequency of melanoma cell detection in patients' blood was about 10% higher when the TYR/MART 1 two-marker assay was used. Using this assay we did not find any statistical correlation between the molecular markers and the UICC stage of disease or the Breslow thickness or Clark level of the primary melanoma. The frequency of melanoma cell detection with the TYR/MUC-18 two-marker assay was even higher than the TYR/MART 1 assay, but unfortunately the MUC-18 transcript was also present in about 20% of healthy subjects. Therefore we do not recommend the use of MUC-18 as a standard value marker. PMID- 11254118 TI - Single-agent DTIC versus combination chemotherapy with or without immunotherapy in metastatic melanoma: a meta-analysis of 3273 patients from 20 randomized trials. AB - It is currently unclear whether any combination therapy for the treatment of metastatic melanoma is superior to standard single-agent dacarbazine (DTIC) in terms of tumour response and overall survival. The available randomized clinical trial data were combined in a meta-analysis to address this question. Initially a thorough MEDLARS search was conducted covering the time period from January 1970 to January 1999. This literature search was supplemented by manual searches of study bibliographies (including review articles) and review of relevant textbooks. The meta-analysis was performed according to a prospective protocol using strict study eligibility criteria. Data derived from randomized controlled trials comparing single-agent DTIC with combination chemo/immunotherapy were combined using a fixed effects model. Data were stratified into three combination therapy groups: DTIC-containing regimens, non-DTIC-containing therapy, and chemotherapy plus immunotherapy. The primary outcome of interest was the proportion of patients demonstrating a complete or partial response to treatment. A total of 20 randomized trials comprising 3273 patients were initially combined in a meta-analysis. This yielded an odds ratio (OR) of 1.23 (95% confidence interval [CI] 1.02-1.48), demonstrating that combination drug therapies are associated with a 23% increase in response rate compared with single-agent DTIC. The combination of DTIC plus interferon-alpha produced a tumour response rate 53% greater (95% CI 1.10-2.13) than that seen with DTIC alone. This increase was greater than that seen with DTIC-containing multi-drug regimens, which had an OR of 1.33 (95% CI 0.99-1.78). No difference in overall survival was demonstrated. Non-DTIC-containing treatment programmes showed no advantage over DTIC in terms of tumour response rate (OR = 0.77, 95% CI 0.45-1.32). The combination of DTIC and interferon-alpha appears more active than standard single-agent DTIC in metastatic melanoma. Further randomized clinical trials employing a DTIC plus interferon arm are necessary to confirm these results. PMID- 11254119 TI - Mobile hospital rooms to fight melanoma. PMID- 11254120 TI - Evidence for orthologous S-locus-related I genes in several genera of Brassicaceae. AB - In the Brassica genus, self-incompatibility (SI) is considered to be controlled by the combined action of several highly polymorphic genes located at the S locus. These genes, including the S-Locus Gene (SLG), and the S-Receptor Kinase (SRK) are all members of the complex multigenic S-family. The S-Locus Related I gene (SLR1) is a member of the S-family, but is not involved in SI control since it is not linked to the S-locus and is essentially monomorphic. Here we confirm or demonstrate the occurrence of SLR1 as highly diverged but not very polymorphic genes in several genera of the Brassicaceae family (Arahidopsis, Brassica, Hirschfeldia, Raphanus, Sinapis). They show similar expression patterns with respect to location (stigmatic papillae), developmental stage (before and during anthesis) and transcript size (1.6 kb). In addition, they are assumed to be involved in the same biological function (late pollen adhesion). These features suggest that the pollen adhesion function might have evolved towards self-pollen recognition through duplication of SLR1 and recruitment of a protein kinase gene. PMID- 11254121 TI - Characterization of the reverse transsulfuration gene mecB of Acremonium chrysogenum, which encodes a functional cystathionine-gamma-lyase. AB - In Acremonium chrysogenum, biosynthesis of cysteine for the formation of cephalosporin has been proposed to occur through the reverse transsulfuration pathway. A gene, named mecB, has been cloned from an A. chrysogenum C10 genomic library in lambdaEMBL3-ble. The cloned DNA fragment encodes a protein of 423 amino acids with a deduced molecular mass of 45 kDa that shows great similarity to cystathionine-gamma-lyases from Saccharomyces cerevisiae and other eukaryotic organisms. The protein was shown to be functional because it restores growth on methionine to A. nidulans C47 (mecB10), a mutant that is known to be defective in cystathionine-gamma-lyase. The cloned gene did not complement A. nidulans mecA or metG mutants. Enzyme activity assays confirmed that the cloned mecB gene encodes a cystathionine-gamma-lyase activity. The mecB gene is present in a single copy in the wild-type A. chrysogenum (Brotzu's strain) and also in the A. chrysogenum strain C10, a high cephalosporin producer. The gene is localized on chromosome VIII (5.3 Mb), as shown by hybridization to A. chrysogenum chromosomes resolved by pulsed-field gel electrophoresis. Transcription of the mecB gene gives rise to a major transcript of 1.5 kb and a minor one of 1.7 kb. The transcript levels were not significantly affected by addition of DL-methionine to the culture, indicating that expression of this gene is not regulated by methionine. The availability of this gene provides a very useful tool for understanding the proposed role of cystathionine-gamma-lyase in splitting cystathionine to supply cysteine for cephalosporin biosynthesis. PMID- 11254122 TI - Characterization of the lys2 gene of Acremonium chrysogenum encoding a functional alpha-aminoadipate activating and reducing enzyme. AB - A 5.2-kb NotI DNA fragment isolated from a genomic library of Acremonium chrysogenum by hybridization with a probe internal to the Penicillium chrysogenum lys2 gene, was able to complement an alpha-aminoadipate reductase-deficient mutant of P. chrysogenum (lysine auxotroph L-G-). Enzyme assays showed that the alpha-aminoadipate reductase activity was restored in all the transformants tested. The lys2-encoded enzyme catalyzed both the activation and reduction of alpha-aminoadipic acid to its semialdehyde, as shown by reaction of the product with p-dimethylaminobenzaldehyde. The reaction required NADPH, and was not observed in the presence of NADH. Sequence analysis revealed that the gene encodes a protein with relatively high similarity to members of the superfamily of acyladenylate-forming enzymes. The Lys2 protein contained all nine motifs that are conserved in the adenylating domain of this enzyme family, a peptidyl carrier domain, and a reduction domain. In addition, a new NADP-binding motif located at the N-terminus of the reduction domain that may form a Rossmann-like betaalphabeta-fold has been identified and found to be shared by all known Lys2 proteins. The lys2 gene was mapped to chromosome I (2.2 Mb, the smallest chromosome) of A. chrysogenum C10 (the chromosome that contains the "late" cephalosporin cluster) and is transcribed as a monocistronic 4.5-kb mRNA although at relatively low levels compared with the beta-actin gene. PMID- 11254123 TI - Multiple cellular processes affected by the absence of the Rpb4 subunit of RNA polymerase II contribute to the deficiency in the stress response of the yeast rpb4(delta) mutant. AB - We previously described the isolation of yeast mutants (sex mutants) that secrete reduced amounts of mature alpha-factor when it is synthesized as part of a fusion with prosomatostatin. In the present study we show that the sex3-1 mutant displays pleiotropic phenotypes. These include an abnormal morphology, an osmoremediable caffeine sensitivity, reduced secretion of mature alpha-factor, a weakened cell wall and a marked deficiency in halotolerance. Cloning of the SEX3 gene revealed that it is identical to the RPB4 gene. This gene encodes the fourth largest subunit of yeast RNA polymerase II, which has been postulated to play a major role in the response to stress. We show that transcriptional activation in response to either a cell wall stress or to growth in the presence of elevated salt concentrations is minimally affected by the loss of RPB4 function. However, whereas the levels of several mRNAs are similarly reduced (by about 30%) in rpb4 mutants grown in rich medium at moderate temperature, some transcripts, in particular ZDS1, are more abundant. An increase dosage of ZDS1, or of genes involved in cell wall assembly and in secretion (RHO1 and SR077, respectively), partially suppresses the sensitivity of rpb4delta cells to high temperature, heat shock and stationary phase. Collectively, our results indicate that the loss of Rpb4p perturbs several cellular functions that contribute to the inappropriate stress response of rpb4delta yeast. We therefore conclude that this RNA poiymerase II subunit is not specifically involved in the stress response. PMID- 11254124 TI - The mitochondrial inner membrane protein Lpe10p, a homologue of Mrs2p, is essential for magnesium homeostasis and group II intron splicing in yeast. AB - The yeast ORF YPL060w/LPE10 encodes a homologue of the mitochondrial protein Mrs2p. These two proteins are 32% identical, and have two transmembrane domains in their C-terminal regions and a putative magnesium transporter signature, Y/F-G M-N, at the end of one of these domains. Data presented here indicate that Lpe10p is inserted into the inner mitochondrial membrane with both termini oriented towards the matrix space. Disruption of the LPE10 gene results in a growth defect on non-fermentable substrates (petite phenotype) and a marked defect in group II intron splicing. The fact that in intron-less strains lpe10 disruptants also exhibit a petite phenotype indicates that functions other than RNA splicing are affected by the absence of Lpe10p. In the mitochondria, concentrations of magnesium, but not of several other divalent metal ions, are increased when Lpe10p is overexpressed and reduced when it is absent. Magnesium concentrations are raised to normal levels and growth on non-fermentable substrates is partially restored by the expression of CorA, the bacterial magnesium transporter, in the lpe10 disruptant. These features are similar to those previously reported for Mrs2p, suggesting that Lpe10p and Mrs2p are functional homologues. However, they cannot easily substitute for each other. Their roles in magnesium homeostasis and, possibly as a secondary effect, in RNA splicing are discussed. PMID- 11254125 TI - Essential genes in proximal 3L heterochromatin of Drosophila melanogaster. AB - We have further characterized essential loci within the centric heterochromatin of the left arm of chromosome 3 (3L) of Drosophila melanogaster, using EMS, radiation and P element mutagenesis. We failed to find any new essential genes, a result that suggests a lower-than-average gene density in this region. Mutations affecting expression of the most proximal gene [lethal 1, l1 or l(3)80Fj] act as dominant suppressors of Polycomb (Pc), behavior which is consistent with a putative trithorax group (trx-G) gene. The third gene to the left of the centromere [lethal 3, l3 or l(3)80Fh] is likely to correspond to verthandi (vtd), a known trx-G gene that plays a role in the regulation of hedgehog (hh) expression and signalling. The intervening gene [lethal 2, l2 or l(3)80Fi] is required throughout development, and mutant alleles have interesting phenotypes; in various allelic combinations that survive, we observe fertility, bristle, wing, eye and cuticle defects. PMID- 11254126 TI - Cycloheximide resistance conferred by novel mutations in ribosomal protein L41 of Chlamydomonas reinhardtii. AB - Although most eukaryotic cells are sensitive to the 80S ribosome inhibitor cycloheximide (CYH), naturally occurring CYH resistance is widespread amongst yeast species. The primary determinant of resistance appears to be a single residue within ribosomal protein L41; resistance is acquired by the substitution of a conserved proline (P56) by a glutamate residue. We have isolated the L41 gene (RPL41) from the green alga Chlamydomonas reinhardtii, and investigated the molecular basis of CYH resistance in various mutant strains. In both the wild type strain and the mutant act-1, a proline is found at the key position in L41. However, analysis of six independently isolated act-2 mutants reveals that all have point mutations that replace the proline with either leucine or serine. Of the two changes, the leucine mutation confers significantly higher levels of CYH resistance. This work identifies the ACT-2 locus as RPL41 and provides a possible dominant marker for nuclear transformation of C. reinhardtii. PMID- 11254127 TI - The chaperonin GroEL and other heat-shock proteins, besides DnaK, participate in ribosome biogenesis in Escherichia coli. AB - It has been shown that in Escherichia coli the chaperone DnaK is necessary for the late stages of 50S and 30S ribosomal subunit assembly in vivo. Here we focus on the roles of other HSPs (heat-shock proteins), including the chaperonin GroEL, in addition to DnaK, in ribosome biogenesis at high temperature. GroEL is shown to be required for the very late 45S-->50S step in the biogenesis of the large ribosome subunit, but not for 30S assembly. Interestingly, overproduction of GroES/GroEL can partially compensate for a lack of DnaK/DnaJ at 44 degrees C. PMID- 11254128 TI - Functional analysis of the Bacillus subtilis y shD gene, a mutS paralogue. AB - In the course of the Bacillus subtilis genome sequencing project, an ORF called yshD was identified, and its product was classified as a mismatch repair protein. Further analysis of the YshD primary sequence showed that the protein belongs to the MutS2 protein family, sharing a high degree of identity with the Thermootoga Inaritima protein TM1278 (34%) and with the so-called MutS2 protein sl11772 of Synechocystis (32%). The COG1193 family of MutS-like proteins is made up of polypeptides that have been predicted from genomic sequencing data from various prokaryotes, but their biological role has not yet been analysed. The functional study of yshD revealed that the gene is constitutively transcribed during the life cycle of B. subtilis, and in minimal medium expression remains at appreciable levels until very late in stationary phase. Fluctuation tests with yshD knock-out mutants did not indicate any role for the protein in preventing the accumulation of spontaneous forward mutations to RifR, nor was any functional interaction with MutS or MutL suggested in fluctuation experiments with mutants lacking combinations of the three genes. Nevertheless, the mutation spectrum observed in the rpoB gene in the deltayshD strain has some characteristic features. The gene does not seem to be involved in the prevention of interspecific recombination in transformation-competent cells. PMID- 11254129 TI - Cytoplasmic kinase and phosphatase activities can induce PsaF gene expression in the absence of functional plastids: evidence that phosphorylation/dephosphorylation events are involved in interorganellar crosstalk. AB - PsaF is a nuclear gene for subunit III of the reaction center of photosystem I, and its expression is stimulated by cytokinins and light, when monitored at the mRNA level or at the level of GUS activity directed by chimeric promoter::uidA gene fusions in transgenic tobacco. These inductive effects can be mimicked by pertussis toxin, serotonin, phorbol acetate myristate or Ca2+, suggesting the involvement of heterotrimeric G proteins, phospholipids and Ca2+-dependent processes. Both breakdown products of the phosphatidylinositol cycle, inositol triphosphate (IP3) and diacylglycerol (or its homolog phorbol myristate acetate, PMA) appear to be involved. The IP3-dependent pathway requires kinase activity, and the signal operates via a 42-bp Ca2+-responsive element located between positions -220 and -178, while the PMA-dependent pathway requires phosphatase activity and a binding element that lies further upstream in the promoter. The effects of G proteins, phospholipids and Ca2+ on GUS gene expression are restricted to tissues with functional plastids, while modulation of phosphatase and kinase activities activates the responsive PsaF promoter regions even in photobleached material. Thus, activation of kinases and phosphatases can bypass the plastid-mediated inhibition of PsaF gene expression in tobacco seedlings. One cytoplasmic target which reflects the functional state of the plastids is protein kinase C. The enzyme can be efficiently phosphorylated in protein extracts from seedlings in which plastid function is impaired, but not in extracts from green tissue. PMID- 11254130 TI - The mtmVUC genes of the mithramycin gene cluster in Streptomyces argillaceus are involved in the biosynthesis of the sugar moieties. AB - Mithramycin is a glycosylated aromatic polyketide produced by Streptomyces argillaceus, and is used as an antitumor drug. Three genes (mtmV, mtmU and mtmC) from the mithramycin gene cluster have been cloned, and characterized by DNA sequencing and by analysis of the products that accumulate in nonproducing mutants, which were generated by insertional inactivation of these genes. The mtm V gene codes for a 2,3-dehydratase that catalyzes early and common steps in the biosynthesis of the three sugars found in mithramycin (D-olivose, D-oliose and D mycarose); its inactivation caused the accumulation of the nonglycosylated intermediate premithramycinone. The mtmU gene codes for a 4-ketoreductase involved in D-oliose biosynthesis, and its inactivation resulted in the accumulation of premithramycinone and premithramycin A , the first glycosylated intermediate which contains a D-olivose unit. The third gene, mtmC, is involved in D-mycarose biosynthesis and codes for a C-methyltransferase. Two mutants with lesions in the mtmC gene accumulated mithramycin intermediates lacking the D mycarose moiety but containing D-olivose units attached to C-12a in which the 4 keto group is unreduced. This suggests that mtmC could code for a second enzyme activity, probably a D-olivose 4-ketoreductase, and that the glycosyltransferase responsible for the incorporation of D-olivose (MtmGIV) shows some degree of flexibility with respect to its sugar co-substrate, since the 4-ketoanalog is also transferred. A pathway is proposed for the biosynthesis of the three sugar moieties in mithramycin. PMID- 11254131 TI - Miscoding and misincorporation of 8-oxo-guanine during leading and lagging strand synthesis in Escherichia coli. AB - We examined whether strand identity with respect to DNA replication influences strand bias for 8-oxo-7,8-dihydroguanine (8-oxoG) mutagenesis. The specificity of 8-oxoG mutagenesis was determined in a mutM mutY or a mutT strain carrying the supF gene on one of two vectors that differed only in the orientation of supF with respect to a unique origin of replication. Most of the supF mutations in the mutM mutY strain were base substitutions (67%), predominantly G:C-->T:A transversions (> 64%), while the majority in the mutT strain were base substitutions (> 92%), predominantly A:T-->C:G transversions (> 91%). The distributions of frequently mutated sites of G:C-->T:A and A:T-->C:G transversions in the supF gene in the mutM mutY and mutT strains, respectively, did not differ markedly between the two vectors. These results suggest that gene orientation is not an important determinant of the strand bias of 8-oxoG mutagenesis. PMID- 11254133 TI - Isolation and characterization of the fission yeast gene Sprpa12+ reveals that the conserved C-terminal zinc-finger region is dispensable for the function of its product. AB - RNA polymerase I of Saccharomyces cerevisiae contains a small subunit, A12.2, encoded by RPA12, that was previously shown to be involved in the assembly and/or stabilization of the largest subunit, A190, of RNA polymerase I. To examine whether an equivalent subunit is present in another eukaryotic RNA polymerase I, we have cloned a Schizosaccahromyces pombe cDNA that is able to complement the rpa12 mutation in S. cerevisiae. The gene, named Sprpa12+, encodes a polypeptide of 119 amino acids that shows 55% identity to S. cerevisiae A12. 2 over its entire length, including two zinc-finger motifs. Disruption of the chromosomal Sprpa12+ gene shows that it is required for growth at higher temperatures but not at lower temperatures. Expression of Sprpa190+/nuc1+, which encodes the largest subunit of the S. pombe RNA polymerase I, from a multicopy plasmid can partially suppress the growth defect of the Sprpa12 disruptant at higher temperatures. These findings suggest that A12.2 subunit is functionally and structurally conserved between S. cerevisiae and S. pombe. Finally, the analysis of mutants suggests that SpRPA12 requires the zinc-finger domain in the N-terminal region but not the one in the C-terminal region for its function. PMID- 11254132 TI - A compromised yeast RNA polymerase II enhances UV sensitivity in the absence of global genome nucleotide excision repair. AB - Nucleotide excision repair is the major pathway responsible for removing UV induced DNA damage, and is therefore essential for cell survival following exposure to UV radiation. In this report, we have assessed the contributions of some components of the RNA polymerase II (Pol II) transcription machinery to UV resistance in Saccharomyces cerevisiae. Deletion of the gene encoding the Pol II elongation factor TFIIS (SII) resulted in enhanced UV sensitivity, but only in the absence of global genome repair dependent on the RAD7 and RAD16 genes, a result seen previously with deletions of RAD26 and RAD28, yeast homologs of the human Cockayne syndrome genes CSB and CSA, respectively. A RAD7/16-dependent reduction in survival after UV irradiation was also seen in the presence of mutations in RNA Pol II that confer a defect in its response to SII, as well as with other mutations which reside in regions of the largest subunit of Pol II not involved in SII interactions. Indeed, an increase in UV sensitivity was achieved by simply decreasing the steadystate level of RNA Pol II. Truncation of the C terminal domain and other RNA Pol II mutations conferred sensitivity to the ribonucleotide reductase inhibitor hydroxyurea and induction of RNR1 and RNR2 mRNAs after UV irradiation was attenuated in these mutant cells. That UV sensitivity can be a consequence of mutations in the RNA Pol II machinery in yeast cells suggests that alterations in transcriptional programs could underlie some of the pathophysiological defects seen in the human disease Cockayne syndrome. PMID- 11254134 TI - Characterisation of novel target promoters for the dexamethasone inducible/tetracycline-repressible regulator TGV using luciferase and isopentenyl transferase as sensitive reporter genes. AB - The chimeric transcriptional activator TGV mediates dexamethasone (dx)-inducible and tetracycline (tc)-repressible transgene expression in tobacco (dx-on/ tc-off system). The expression profiles of four different synthetic target promoters, comprising multiple TGV binding sites upstream of a core promoter, were characterised using the sensitive luciferase assay. Induction factors of over 1,000 were measured in roots and leaves of over 30% of the transgenic plants, irrespective of the promoter used. Promoters PTF and PTax, which carry the -48 to +1 region of the Cauliflower Mosaic Virus 35S promoter, showed higher expression levels in both the uninduced and induced states than PTop10 and PTFM, which harbour several point mutations in this region. Moreover, PTax expressed higher background activities than PTF, indicating that the sequence of the synthetic regulatory region can influence background levels. The usefulness of the dx-on/tc off system for experiments addressing gene function was demonstrated by using it to control the expression of isopentenyl transferase. This enzyme catalyses the rate-limiting step in cytokinin biosynthesis and causes phenotypic effects even at low expression levels. Only dx-induced transgenic plants displayed phenotypic alterations indicative for increased cytokinin synthesis (e.g. outgrowth of lateral buds). Simultaneous treatment of selected buds with the antiinducer tc suppressed bud growth. This result suggests that cytokinins cannot serve as mobile signals to elicit the release of apical dominance in tissues compromised for enhanced cytokinin synthesis. PMID- 11254135 TI - Isolation, characterization, inheritance and linkage of microsatellite DNA markers in white spruce (Picea glauca) and their usefulness in other spruce species. AB - Microsatellite DNA/simple-sequence-repeat (SSR) loci were identified, isolated and characterized in white spruce (Picea glauca) by screening both a non-enriched partial genomic library and a partial genomic library enriched for (AG/TC)n containing clones. Inheritance and linkage of polymorphic SSR loci were determined in F1 progeny of four controlled crosses. We also assessed the compatibility and usefulness of the P. glauca microsatellite DNA markers in five other Picea species. Twenty-four microsatellites were identified by sequencing 32 clones selected from screens of 5,400 clones from the two libraries. The (AG/TC)n microsatellites were the most abundant in the non-enriched library. Eight microsatellite DNA loci were of the single-copy type, and six of these were polymorphic. A total of 87 alleles were detected at the six polymorphic SSR loci in 32 P. glauca individuals drawn from several populations. The number of alleles found at these six SSR loci ranged from 2 to 22, with an average of 14.5 alleles per locus, and the observed heterozygosity ranged from 0.48 to 0.91, with a mean of 0.66 per locus. Parents of the controlled crosses were polymorphic for five of the six polymorphic SSR loci. Microsatellite DNA variants at each of these five SSR loci followed a single-locus, codominant, Mendelian inheritance pattern. Joint two-locus segregation tests indicated complete linkage between PGL13 and PGL14, and no linkage between any of the remaining SSR loci. Each of the 32 P. glauca individuals examined had unique single or two-locus genotypes. With the exception of non-amplification of PGL12 in P. sitchensis, P. mariana, and P. abies and the monomorphic nature of PGL7 in P. mariana, primer pairs for all six polymorphic SSR loci successfully amplified specific fragments from genomic DNA and resolved polymorphic microsatellites of comparable sizes in P. engelmanni, P. sitchensis, P. mariana, P. rubens, and P. abies. The closely related species P. mariana and P. rubens, and P. glauca and P. sitchensiss could be distinguished by the PGL12 SSR marker. The microsatellite DNA markers developed and reported here could be used for assisting various genetics, breeding, biotechnology, tree forensics, genome mapping, conservation, restoration, and sustainable forest management programs in spruce species. PMID- 11254137 TI - The Neurospora crassa colonial temperature-sensitive 3 (cot-3) gene encodes protein elongation factor 2. AB - At elevated temperatures, the Neurospora crassa mutant colonial, temperature sensitive 3 (cot-3) forms compact, highly branched colonies. Growth of the cot-3 strain under these conditions also results in the loss of the lower molecular weight (LMW) isoform of the Ser/Thr protein kinase encoded by the unlinked cot-1 gene, whose function is also involved in hyphal elongation. The unique cot-3 gene has been cloned by complementation and shown to encode translation elongation factor 2 (EF-2). As expected for a gene with a general role in protein synthesis, cot-3 mRNA is abundantly expressed throughout all asexual phases of the N. crassa life cycle. The molecular basis of the cot-3 mutation was determined to be an ATT to AAT transversion, which causes an Ile to Asn substitution at residue 278. Treatment with fusidic acid (a specific inhibitor of EF-2) inhibits hyphal elongation and induces hyperbranching in a manner which mimics the cot-3 phenotype, and also leads to a decrease in the abundance of the LMW isoform of COT1. This supports our conclusion that the mutation in cot-3 which results in abnormal hyphal elongation/branching impairs EF-2 function and confirms that the abundance of a LMW isoform of COT1 kinase is dependent on the function of this general translation factor. PMID- 11254136 TI - Functional asymmetry of the two nucleotide binding domains in the ABC transporter Ste6. AB - The yeast a-factor transporter Ste6 is a member of the ABC transporter family and is closely related to human MDR1. We constructed a set of 26 Ste6 mutants using a random mutagenesis approach. Cell fractionation experiments demonstrated that most of the mutants, with the notable exception of those with alterations in TM1, are transported to the plasma membrane, the presumptive site of action of Ste6. Trafficking, therefore, does not seem to be affected in most of the mutants. To identify regions in Ste6 that interact with the ABC transporter "signature motif" (LSGGQ) we screened for intragenic revertants of the LSGGQ mutant M68 (S507N). Suppressor mutations were identified in TM12 and upstream of TM6. Surprisingly, these mutations also suppressed the Walker A mutation G397D, which should be defective in ATP-binding and hydrolysis at NBD1. Photoaffinity labeling experiments with 8-azido-[alpha-32P]ATP showed that ATP binding at NBD2 is reduced by the suppressor mutation in TM12. The experiments further suggest that the two NBDs of Ste6 are not equivalent and affect each other's ability to bind and hydrolyze ATP. PMID- 11254138 TI - Two RpoH homologs responsible for the expression of heat shock protein genes in Sinorhizobium meliloti. AB - We identified two rpoH-related genes encoding sigma32-like proteins from Sinorhizobium meliloti, a nitrogen-fixing root-nodule symbiont of alfalfa. The genes, rpoH1 and rpoH2, are functionally similar to rpoH of Escherichia coli because they partially complemented an E. coli rpoH null mutant. We obtained evidence indicating that these genes are involved in the heat shock response in S. meliloti. Following an increase in temperature, synthesis of several putative heat shock proteins (Hsps) was induced in cultures of wild-type cells: the most prominent were 66- and 60-kDa proteins, both of which are suggested to represent GroEL species. The other Hsps could divided into two groups based on differences in synthesis kinetics: synthesis of the first group peaked 5-10 min, and expression of the other group 30 min, after temperature upshift. In the rpoH1 mutant, inducible synthesis of the former group was markedly reduced, whereas that of the latter group was not affected. Synthesis of both the 66- and 60-kDa proteins was partially reduced. While no appreciable effect was observed in the rpoH2 single mutant, the rpoH2 mutation had a synergistic effect on the 60-kDa protein in the rpoH1- background. The results indicate that two distinct mechanisms are involved in the heat shock response of S. meliloti: one requires the rpoH1 function, while rpoH2 can substitute in part for the rpoH1 function. Moreover, the rpoH1 mutant and rpoH1 rpoH2 double mutant exhibited Nod+ Fix- and Nod- phenotypes, respectively, on alfalfa. PMID- 11254139 TI - The stringent response, sigmaH-dependent gene expression and sporulation in Bacillus subtilis. AB - Recently, we found that transcription of the yvyD gene of Bacillus sibtilis from a sigmaH-dependent promoter is induced in response to amino acid starvation. This induction occurred in the wild type, but not in the relA mutant, suggesting that ppGpp might be involved in the activa tion of sigmaH. In order to substantiate this finding we looked for other sigmaH-dependent genes which also required an active relA gene product for activation in amino acid-starved cells. Three additional transcription units, namely spo0A, spoVG and the ytxGHI operon (also known as the csb40 operon), were found to be under the dual control of RelA and sigmaH in amino acid-starved cells, but many other sigmaH-dependent genes were not activated under these conditions. Of particular interest is the observation that spo0A is one of the genes that need a functional relA gene product for sigmaH-dependent gene activation. Furthermore, in sporulation medium, both Spo0A accumulation and phosphorylation are delayed in the relA deletion strain, leading to a delay in sporulation. A comparison between a relA point mutant and a relA deletion mutant revealed that the relA deletion mutant exhibits a more pronounced delay in the appearance of spores. These results suggest that the stringent response is not required for sporulation per se, but rather enhances the efficiency of sporulation. PMID- 11254140 TI - DNA photolyase homologs are the major UV resistance factors in the cyanobacterium Synechocystis sp. PCC 6803. AB - In this study, the unicellular photosynthetic cyanobacterium Synechocystis sp. PCC 6803 was used as a model phototroph to study the contribution of enzymatic photoreactivation to the overall protection against UV irradiation. We have isolated genes encoding two DNA photolyase homologs, phrA and phrB, from Synechocystis 6803. phrA encodes an 8-hydroxy-5-deazariboflavin (HDF) type, Class I DNA photolyase. By complementing a photolyase-deficient mutant strain of Escherichia coli, we demonstrated that PhrA is a DNA photolyase. Analysis of a phrA knockout mutant strain suggested that this gene is responsible for the majority of the observed UV resistance in Synechocystis 6803. Similar studies on phrB demonstrated that it also contributes to photoreactivation, but to a much lesser degree. Based on these findings, we conclude that enzymatic photoreactivation is the primary process used for repairing UV-induced damage in Synechocystis 6803. PMID- 11254141 TI - Expression of the seqA gene is negatively modulated by the HU protein in Escherichia coli. AB - The SeqA protein acts as a regulator of chromosomal replication initiation in Escherichia coli by sequestering hemi-methylated oriC, effectively blocking methylation and therefore preventing rapid re-initiation. The level of SeqA protein is maximal at mid-log phase and decreases when cells enter late-log phase. In hup mutants that lack the HU protein, the maximal seqA expression is also seen at mid-log phase, but seqA expression, as well as SeqA levels and activity, is increased by up to four fold relative to that in the wild type. These results suggest that the HU protein functions as a negative modulator of seqA expression. PMID- 11254142 TI - Overview: extreme environments. PMID- 11254143 TI - Do parasites live in extreme environments? Constructing hostile niches and living in them. AB - We develop the hypothesis that parasites do not invade extreme environments, i.e. hostile hosts, but rather 'create' them. We argue that parasites may have driven the evolution of the constitutive and adaptive immune system. This leads to several implications. First, parasites respond to 'genes to kill' by 'genes to survive' and this triggers an indefinite selection of measures and counter measures. Second, these coevolutionary arms races may lead to local adaptation, in which parasite populations perform better on local hosts. Third, the evolution of the immune system, whose responses are predictable, may allow parasites to specialize, to evade and even to manipulate. Finally we show that the correlations between the increase in the antibody repertoire, the expansion of MHC loci and parasite pressures support our hypothesis that both host complexity and parasite pressures can be invoked to explain the diversity of antibodies, T receptors and MHC molecules. PMID- 11254144 TI - Analysis of parasite host-switching: limitations on the use of phylogenies. AB - Even the most generalist parasites usually occur in only a subset of potential host species, a tendency which reflects overriding environmental constraints on their distributions in nature. The periodic shifting of these limitations represented by host-switches may have been an important process in the evolution of many host-parasite assemblages. To study such events, however, it must first be established where and when they have occurred. Past host-switches within a group of parasites are usually inferred from a comparison of the parasite phylogeny with that of the hosts. Congruence between the phylogenies is often attributed to a history of association by descent with cospeciation, and incongruence to host-switching or extinction in 'duplicated' parasite lineages (which diverged without a corresponding branching of the host tree). The inference of host-switching from incongruent patterns is discussed. Difficulties arise because incongruence can frequently be explained by different combinations of biologically distinct events whose relative probabilities are uncertain. Also, the models of host-parasite relationships implicit in historical reconstructions may often not allow for plausible sources of incongruence other than host switching or duplication/extinction, or for the possibility that colonization could, in some circumstances, be disguised by 'false' congruence. PMID- 11254146 TI - Desiccation survival of parasitic nematodes. AB - The ability of certain species of parasitic nematodes to survive desiccation for considerable periods is a fascinating example of adaptation to the demands of fluctuating environments that occasionally can become extreme and life threatening. Behavioural and morphological adaptations associated with desiccation survival serve primarily to reduce the rate of drying, either to prolong the time taken for the nematode's water content to reach lethal low levels or, in true anhydrobiotes, to enable the structural and biochemical changes required for long-term survival to take place. Examples of these adaptations are reviewed, together with information on the factors involved in rehydration that ensure successful exit from the dormant state. Information on desiccation survival is central to effective management and control options for parasitic nematodes. It is also required to assess the feasibility of enhancing the longevity of commercial formulations of entomopathogenic nematodes, both before and after application; current research and future prospects for enhancing survival of these bio-insecticides are discussed. PMID- 11254145 TI - Digenean parasites of deep-sea teleosts: a review and case studies of intrageneric phylogenies. AB - Studies on the digenean parasites of deep-sea (> 200 m depth) teleosts are reviewed and two case study generic phylogenies are presented based on LSU rDNA and ND1 mtDNA sequences. The phylogeny of the lepocreadiid genus Lepidapedon, the most common deep-sea digenean genus, is not clearly resolved as the two gene trees are not compatible. It can be inferred, however, that the genus has radiated in the deeper waters off the continental shelf, mainly in fishes of the gadiform family Macrouridae. Steringophorus, a fellodistomid genus, is better resolved. In this case a deep-sea radiation is also indicated, but the pattern of host-specificity is not clear, with evidence of much host-switching. Results of studies of the parasites of the macrourid fish Coryphaenoides (Nematonurus) armatus from various depths have reinforced recent views on the lack of zoned depth-related communities in the deep-sea. The diversity of deep-sea digeneans is relatively low with only 18 families (of about 60) reported. Little, or nothing, is known from most deep-sea areas and nothing from trenches and mid-ocean ridge systems. PMID- 11254147 TI - Parasite adaptation to extreme conditions in a desert environment. AB - Deserts represent universally recognized extreme environments for animal life. This paper documents the highly specialized adaptations of Pseudodiplorchis americanus, a monogenean parasite of the desert toad, Scaphiopus couchii. Building on a long-term record of parasite population ecology (continuing since the early 1980s), field studies focus on the effects of severe drought in the Sonoran Desert, Arizona, in the mid 1990s. This provides a test of the ability of the host-parasite system to tolerate exceptional perturbation. The analysis provides new insight into parasite infection dynamics in a natural wildlife system through integration of host and parasite population age structure. The environmental check interrupted host recruitment in 1993-95 and parasite recruitment in 1995-97. This produced an imprint in age structure and infection levels recognizable over several years: parasite recruitment failure reduced transmission 2-3 years later. The host (maximum life span 17 years) tolerated the disruption but the impact was more serious for the parasite (life span 3 years) leading to extinction of some previously stable populations. Despite this demonstration of a rare event exacerbating external environmental constraints, experimental studies suggest that the internal (host) environment normally creates the most severe conditions affecting P. americanus. Only about 3% of parasites survive from invasion until first reproduction. Post-invasion factors including host immunity, characteristic of most parasite life cycles, constitute a greater constraint upon survival than external conditions, even in a desert environment. PMID- 11254149 TI - Parasites and low temperatures. AB - Low temperatures affect the rate of growth, development and metabolism of parasites and when temperatures fall below 0 degrees C may expose the parasite to the potentially lethal risk of freezing. Some parasites have mechanisms, such as diapause, which synchronise their life cycle with favourable seasons and the availability of hosts. Parasites of endothermic hosts are protected from low temperatures by the thermoregulatory abilities of their host. Free-living and off host stages, however, may be exposed to subzero temperatures and both freezing tolerant and freeze-avoiding strategies of cold hardiness are found. Parasites of ectothermic hosts may be exposed to subzero temperatures within their hosts. They can rely on the cold tolerance adaptations of their host or they may develop their own mechanisms. Exposure to low temperatures may occur within the carcass of the host and this may be of epidemiological significance if the parasite can be transmitted via the consumption of the carcass. PMID- 11254148 TI - The survival of monogenean (platyhelminth) parasites on fish skin. AB - This review deals with the problems faced by those monogenean (platyhelminth) parasites that attach themselves to fish skin. The structure of the skin and the ways in which the posterior hook-bearing haptor achieves virtually permanent attachment to the skin are considered. Small marginal hooklets are specialized for attachment to superficial host epidermal cells, finding anchorage in the terminal web of keratinous tonofilaments, while large hooks (hamuli) may penetrate into and lodge in the collagenous dermis. The complementary roles of suction and sticky secretions in haptor attachment and the role of the pharynx in temporary attachment during feeding are also considered. During leech-like locomotion the haptor is briefly detached and, at this critical time, the anterior end is strongly fixed to the wet, current-swept and possibly slimy skin by a sticky secretion. This secretion is deployed on paired pads or discs, the latter sometimes backed up by suction. After attachment by the haptor is re established, the special tegument covering the anterior adhesive areas may be instrumental in their instant release. The role of fish skin in the phenomenon of host specificity and in the generation of a defensive response against monogeneans is considered and site-specificity of parasites on the host's body is discussed. Possible selection pressures exerted by predatory 'cleaner' organisms are briefly evaluated. PMID- 11254151 TI - A guest editorial from abroad: medicolegal opinion--time for peer review. PMID- 11254150 TI - Pentastomids and the tetrapod lung. AB - Pentastomids comprise a highly specialized taxon of arthropod-like parasites that probably became adapted to the lungs of amphibians and reptiles early in their long evolutionary history. Few other macroparasites exploit this particular niche. Pentastomids are often large, long-lived and yet they cause little observable pathology in lungs, despite being haematophagous. The lungs of all tetrapods are lined with pulmonary surfactant, a remarkable biological material consisting of a complex mixture of phospholipids, neutral lipids and proteins that has the unique ability to disperse over the air-liquid lining of the lung. In the lower tetrapods it acts as an anti-glue preventing adhesion of respiratory surfaces when lungs collapse during swallowing prey or upon expiration. In mammals, pulmonary surfactant also plays a critical role regulating the activity of alveolar macrophages, the predominant phagocytes of the lower airways and alveoli. This review outlines the evidence suggesting that lung-dwelling pentastomids, and also nymphs encysted in the tissues of mammalian intermediate hosts, evade immune surveillance and reduce inflammation by coating the chitinous cuticle with a their own stage-specific surfactant. The lipid composition of surfactant derived from lung instars of the pentastomid Porocephalus crotali cultured in vitro is very similar to that recovered from the lung of its snake host. Pentastomid surfactant, visualised as lamellate droplets within sub parietal cells, is delivered to the cuticle via chitin-lined efferent ducts that erupt at a surface density of < 400 mm(-2). The fidelity of the system, which ensures that every part of the cuticle surface is membrane-coated, testifies to its strategic importance. Two other extensive glands discharge membrane associated (hydrophobic?) proteins onto the hooks and head; some have been purified and partly characterized but their role in minimising inflammatory responses is, as yet, undetermined. PMID- 11254152 TI - Fetal biometry: clinical, pathological, and technical considerations. AB - Sonographic measurements of fetal ultrasound parameters are the basis for accurate determination of gestational age and detection of fetal growth abnormalities. Selection of the most useful single biometric parameter depends on the timing and purpose of measurement and is influenced by specific limitations. CRL (crown-rump length) is the best parameter for early dating of pregnancy. Biparietal diameter (BPD) maintains the closest correlation with gestational age in the second trimester. In cases of variation in the shape of the skull, head circumference is an effective alternative. Abdominal circumference is the most useful dimension to evaluate fetal growth, and femur length is the best parameter in the evaluation of skeletal dysplasia. Use of multiple predictors improves the accuracy of estimates. An individual approach to each pregnancy is recommended for fetal growth assessment. The various epidemiological factors involved in fetal growth should be considered and specific charts for different communities should be used when possible. The methods of fetal weight estimation with their limitations and potential errors are presented. Clinical application of fetal biometry in abnormal growth is discussed in cases of small- and large-for gestational-age fetuses, chromosomal aberrations, and skeletal dysplasias. PMID- 11254153 TI - Deeply infiltrating endometriosis: implications, diagnosis, and management. AB - Deeply infiltrating endometriosis was described in the early part of the last century. Only recently, has there become a greater awareness and understanding of this form of endometriosis aided in part by advances in laparoscopic surgical technology in techniques. The clinical implications of the disease as well as diagnosis and current management are reviewed. PMID- 11254154 TI - Obstetrical delivery of the HIV-positive woman: legal and ethical considerations. AB - Every year, thousands of perinatally HIV-infected children are born, resulting in debate about appropriate HIV treatment and interventions for pregnant women. Recent medical studies endorse the use of the cesarean delivery to reduce vertical (mother to infant) transmission of HIV. In addition to medical questions, this practice raises legal and ethical considerations for the attending physician. In the context of AIDS prevention, the potential exists for reasoned and well-informed decision making to give way to encouragement, and even duress, in cases where a woman refuses recommended surgical delivery. However, in such cases, the role of the physician should remain as that of an informed educator and counselor, enabling the patient to exercise her autonomy and personal choice within her social and cultural context. PMID- 11254155 TI - The ductus venosus. AB - Until recently, our information on the ductus venosus was based on postmortem and experimental studies. The present review relates to the modern concept of this vein predominantly founded on clinical studies. Recent publications show that the blood distribution through the ductus venosus is particularly sensitive to changes in umbilical venous pressure, blood viscosity, and an active regulation of diameter of the entire ductus venosus. The mean fraction of umbilical blood shunted through the ductus is reduced from 30% to 20% during the second half of the human pregnancy, indicating that, during this period, the fetal liver has a higher priority than the shunting through the ductus venosus, apart from the compensatory redistribution needed during extreme challenges of placental compromize and hypoxemia. Additionally, the ductus venosus acts as a transmission line to the umbilical vein for pulse waves generated in the heart. These waves, reflecting cardiac function, are substantially influenced by the local variation of impedance and compliance. PMID- 11254156 TI - Doppler assessment of the fetal venous system. AB - This article describes the achievements in Doppler measurements of the fetal venous circulation with emphasis on the clinical impact of these techniques. In rhesus isoimmunization, fetal venous flow assessment gives useful information on the fetal haematologic condition and on the impact of blood transfusion. In first trimester fetuses, Doppler evaluation of the ductus venosus and umbilical vein could attribute to the detection of cardiac defects and/or chromosomal abnormalities. The inferior vena cava flow velocity waveform could be studied in cases of fetal arrhythmias to be able to diagnose the type of fetal heart rhythm disturbances. One of the more important applications of venous Doppler assessment is its use in the evaluation of the intrauterine growth retarded fetus, who is suffering from placental insufficiency. Decrease of the late diastolic flow component in the ductus venosus waveform and the presence of umbilical venous pulsations are distinct alterations, which have been detected before cardiotocogram deterioration occurs. The clinical possibilities of venous Doppler measurements are limited, and the use of the techniques requires intensive training. PMID- 11254157 TI - Three-dimensional ultrasonography in early pregnancy. AB - Three-dimensional ultrasound was introduced into clinical use over a decade ago. Early attempts on three-dimensional ultrasound were primitive and limited to experimental use only. Recently, it was used as a powerful tool to display three dimensional fetal appearance and assess normal and pathological conditions. Because of the improvement in image resolution and reconstruction speed, three dimensional ultrasound is now more and more popular in many centers, and used commonly as an adjunct in prenatal diagnosis. Indeed, a skillful sonographer may usually make the correct diagnosis of fetal anomaly with simply two-dimensional ultrasound. Nonetheless, the situation is gradually changing, because some anomalies have been diagnosed only based on the findings of three-dimensional ultrasound. In spite of that, the application of three-dimensional ultrasound is still less discussed in early pregnancy. In this article, the authors clarify the capability of three-dimensional ultrasound in different anatomic areas, and catalog its clinical merits in the early pregnancy to date. We also present the safety guidelines for its use in the first trimester. PMID- 11254158 TI - Ductus venosus blood flow in chromosomally abnormal fetuses at 11 to 14 weeks of gestation. AB - This article reviews the role of ductus venosus (DV) Doppler evaluation in the screening for aneuploidies at 11 to 14 weeks of gestation. Ductus venosus flow velocity waveforms were obtained immediately before fetal karyotyping in 515 consecutive singleton pregnancies at 11 to 14 weeks. We found 446 normal and 69 abnormal karyotypes. Abnormal flow in the DV was the only significant difference between both groups. Sensitivity of the test was 80% and false positive rate < 1%. Normal karyotype but abnormal flow in the DV was recorded in 17 of 446 cases, 7 presenting a cardiac defect. Increased nuchal translucency seems to be related, in most cases, to early cardiac dysfunction. Chromosomal abnormalities and/or cardiac defects were often found in cases with increased nuchal translucency and abnormal flow in the DV. We suggest that the evaluation of ductal flow between 11 to 14 weeks of gestation should be adopted as a second level screening test to reduce invasive test rate derived from the exclusive measurement of nuchal translucency. PMID- 11254159 TI - Transvaginal 3D and Doppler ultrasonography of the fetal brain. AB - Transvaginal sonographic approach to the fetal brain, which provides detailed information about the fetal intracranial morphology, opened a new field in medicine, "neurosonography." The clinical significance of 3D ultrasound for prenatal diagnosis has been discussed since three-dimensional ultrasound was introduced in obstetrics. Three-dimensional ultrasound has several functions: surface reconstruction, multiplanar image analysis, three-dimensional sono angiography, and volume calculation. In this article, we introduce transvaginal three-dimensional ultrasound for the assessment of fetal head and brain. Surface mode shows not only fetal head abnormality such as acrania but also normal cranial bones and sutures in the first trimester. Rotation of the brain volume image and multiplanar analysis enable tomographic visualization as magnetic resonance imaging. Sono-angiography shows the brain circulation three dimensionally and extracted volume images of target organ provide information on detailed intracranial conditions. The technology is easy, noninvasive, and reproducible methods, and produces comprehensible and objective information. PMID- 11254160 TI - Umbilical artery Doppler velocimetry--an update. AB - Doppler ultrasonography was introduced into clinical obstetric practice over 20 years ago. It is also accepted that a variety of common obstetrical complications such as preeclampsia and intrauterine growth restriction have their origin in abnormal development of the placental vasculature and this could be reflected in abnormal Doppler velocimetry. Doppler velocimetry of the umbilical artery has been the subject of multiple clinical studies but results have often been disputed due to differences in study populations and methodologies. In recent years, meta-analysis of randomized clinical trials have shown that incorporation of Doppler velocimetry into clinical practice will reduce perinatal mortality in high-risk patients. This article reviews the data of the meta-analyses as it pertains to the management of high-risk pregnant patients. PMID- 11254161 TI - Cadmium-mediated oxidative stress in alveolar epithelial cells induces the expression of gamma-glutamylcysteine synthetase catalytic subunit and glutathione S-transferase alpha and pi isoforms: potential role of activator protein-1. AB - Exposure of rat alveolar epithelial cells to 10 micromol/L CdCl2 causes time dependent increases in steady-state mRNA levels of the gamma-glutamylcysteine synthetase catalytic (heavy) subunit (gamma-GCS) and of glutathione S-transferase isoforms (GST-alpha and GST-pi). The expression of gamma-GCS was significantly increased as early as 2 h after addition of cadmium. Maximal induction of gamma GCS mRNA (approximately 4-fold), at 8 h, was subsequently followed by increases in gamma-GCS activity/protein and glutathione (GSH) levels. Maximal elevations in GST-pi (approximately 2-fold) and GST-alpha (approximately 10-fold) transcripts, at 8 and 24 h, respectively, were also accompanied by enhanced GST activity. Cadmium-induced oxidative stress, assessed by alterations in GSH homeostasis and an accelerated rate of intracellular oxidant production, could constitute early events in the signal transduction pathway mediating these responses. The dimeric transcription factor, activator protein-1 (AP-1), may also play a regulatory role in this process. This association is suggested by transcriptional activation of the immediate-early response genes, c-fos and c-jun, within 15 min after exposure to cadmium and by the enhancement of AP-1 DNA binding activity, involving a c-Jun protein complex, which is maximally induced (approximately 4-fold) by 2 h. These molecular changes likely function together to protect alveolar epithelial cells against cadmium toxicity. PMID- 11254162 TI - Effect of cisplatin on mitochondrial protein, glutathione, and succinate dehydrogenase in Dalton lymphoma-bearing mice. AB - Cisplatin treatment of tumor-bearing mice resulted a significant decrease of protein in the tissues studied (liver, kidney, and Dalton lymphoma) and also in their mitochondrial fractions. As compared to respective tissues, the protein decrease was noted to be more conspicuous in their mitochondrial fractions. Similarly, mitochondrial glutathione also decreased significantly in the tissues. However, succinate dehydrogenase activity was selectively decreased in the kidney and Dalton lymphoma cells, whereas in liver it remained almost unchanged. An increase in serum urea concentration and kidney mitochondrial lipid peroxidation was also observed after cisplatin treatment. It is suggested that the cisplatin induced biochemical changes in mitochondria involving mitochondrial protein, glutathione, and succinate dehydrogenase could be the important potent cellular sites contributing to toxicity/cytotoxicity after cisplatin treatment. PMID- 11254163 TI - Glucocorticoid receptor-Hsp90 interaction in the liver cytosol of cadmium intoxicated rats. AB - The influence of cadmium on the rat liver glucocorticoid receptor (GR) binding capacity, on the cytosolic level of 90 kDa heat shock protein (Hsp90), and on the association of the two proteins was investigated. The results showed that the mode of metal application led to diverse alterations in hormone binding to the GR. Reduction of the GR binding capacity observed after in vitro treatment was proportional to the applied metal concentrations. In animals administered different doses of cadmium, GR binding capacity was not reduced, except in those that received the highest dose. A concomitant elevation of Hsp90 level was detected both in the cytosol and within the GR untransformed heterocomplexes. The results suggest that cadmium-induced reduction of the GR binding capacity seen in vitro was prevented in intact animals by the elevated level of Hsp90 within the GR heterocomplexes. PMID- 11254164 TI - The effect of medroxyprogesterone acetate on bone marrow and testis during cytotoxic chemotherapy. AB - The effect of medroxyprogesterone acetate (MPA) on the mitotic activity of bone marrow and testis during chemotherapy was investigated experimentally in an animal study. A total of 120 male Swiss albino mice were included in this study. Six groups were formed, each consisting of 20 mice. Low-dose MPA (LD-MPA) (15 mg/kg), high-dose MPA (HD-MPA) (100 mg/kg), LD-MPA plus cyclophosphamide (CP) (65 mg/kg), HD-MPA plus CP (65 mg/kg), and CP (65 mg/kg) were administered to the test groups and no drug was administered to the control group. Bone marrow samples and testis were examined for mitotic activity rate (MAR) on days 0, 18, 22, 26, and 30. In groups with regimens containing CP, MAR of hematopoietic cells in bone marrow was suppressed significantly (p<0.05). There was no difference in MAR of hematopoietic cells in bone marrow between the groups given MPA or not (p>0.05). Mitotic activity rate of the testis cells was significantly suppressed in groups with regimens containing MPA (p<0.05). In conclusion, MPA inhibited mitotic activity of testis, but there was no effect on the mitotic activity of bone marrow. These data do not seem to confirm the hypothesis of a myeloprotective effect of MPA. PMID- 11254166 TI - Radiation dosimetry using Fricke-infused gels and magnetic resonance imaging. AB - We have witnessed the advancements of MRI-Fricke-infused gel dosimetry since its commencing in 1984. Over the years, many efforts have been spent to improve the method's efficacy, i.e., to improve its dose-response sensitivity, reproducibility and measurement accuracy. In this article, we give a review of the development of this relatively new dosimetric method. An example of applying this method to gamma knife stereotactic radiosurgery dose distribution mapping is also given. PMID- 11254167 TI - Development of lipase in nursing piglets. AB - This experiment was designed to investigate the development of gastric lipase and pancreatic lipase in nursing piglets. During the nursing period, the lipase activity was measured at one, 7, 14, 21 and 28 days of age. The results showed that the gastric mucosa weight, pancreas weight, body weight, the specific activity of gastric lipase and pancreatic lipase, and the total activity of gastric lipase and pancreatic lipase increased with the age of the piglets. The development of the specific activity of gastric lipase slowed before the nursing piglets reached 3 weeks of age, but the total activity of gastric lipase at day 28 was significantly higher than that at day 21. The specific activity and total activity of pancreatic lipase were at low levels during the first two weeks of life and then developed quickly from days 21 to 28. It was observed that the specific activity and total activity of gastric lipase were lower than those of pancreatic lipase. PMID- 11254168 TI - Selection of valid and reliable EEG features for predicting auditory and visual alertness levels. AB - A selection procedure with three rules, high efficiency, low individual variability, and low redundancy, was developed to screen electroencephalogram (EEG) features for predicting behavioral alertness levels. A total of 24 EEG features were derived from temporal, frequency spectral, and statistical analyses. Behavioral alertness levels were quantified by correct rates of performance on an auditory and a visual vigilance task, separately. In the auditory task study, a subset of three EEG features, the relative spectral amplitudes in the alpha (alpha%, 8-13 Hz) and theta (theta%, 4-8 Hz) bands, and the mean frequency of the EEG spectrum (MF), was found to be the best combination for predicting the auditory alertness level. In the visual task study, the mean frequency of the beta band (Fbeta, 13-32 Hz) was the only EEG feature selected. The application of an averaging subwindow procedure within a moving time window to EEG analysis increased the predictive power of EEG features and decreased the disturbing effect of movement artifacts on the EEG data. PMID- 11254165 TI - Development of a highly sensitive in vitro phototoxicity assay using the SkinEthic reconstructed human epidermis. AB - The reconstituted human epidermis model SkinEthic was used to evaluate the phototoxicity of topically applied chemicals. For comparison with published data, we first tested a library of 13 nonphototoxic (NPT) and phototoxic (PT) compounds, applied onto SkinEthic reconstituted human epidermal tissues, in a protocol as close as possible to the one described by Liebsch using another skin tissue model. The results showed that, under these nonoptimized conditions, the SkinEthic model was already able to fully discriminate between known NPT and PT compounds. Furthermore, these epidermal tissues being highly resistant to UVA irradiation, it was possible to increase irradiation by (at least) 3-fold without decrease in tissue viability. In such conditions, the phototoxicity assay is much more sensitive, so that the model is expected to be of great interest for the detection not only of strong but also of weak phototoxic compounds. PMID- 11254170 TI - Prediction of soil depth using a soil-landscape regression model: a case study on forest soils in southern Taiwan. AB - Techniques for conventional forest soil surveys in Taiwan need to be further developed in order to save time and money. Although some soil-landscape regression models have been developed to describe and predict soil properties and depths, they have seldom been studied in Taiwan. This study establishes linear soil-landscape regression models related to soil depths and landscape factors found in the forest soils of southern Taiwan. These models were evaluated by validating the models according to their mean errors and root mean square errors. The study was carried out at the 60,000 ha Chishan Forest Working Circle. About 310 soil pedons were collected. The landscape factors included elevation, slope, aspect, and surface stone contents. Sixty percent of the total field samples were used to establish the soil-landscape regression models, and forty % were used for validation. The sampling strategy indicated that each representative pedon covers an area of about 147 ha. The number of samples was appropriate considering the available time and budget. The single variate and/or multivariate linear regression soil-landscape models were successfully established. Those models revealed significant inter-relations among the soil depths of the B and B+BC horizons, solum thickness, and landscape factors, including slope and surface stone contents (p < 0.003). The mean errors in the validation of the soil landscape model were low and acceptable for this case study. In addition, the slope data derived from the DEM (digital elevation model) database in this case study were used to predict the soil depths of the B, B+BC horizons and the solum thickness without carrying out a field survey. Surface stone should be collected in a field soil survey to increase the precision of soil depth prediction of the B and B+BC horizons, and the solum thickness. PMID- 11254169 TI - Inhibition of lignin peroxidase-mediated oxidation activity by ethylenediamine tetraacetic acid and N-N-N'-N'-tetramethylenediamine. AB - The mineralization rate of LC-[1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane] (DDT) was reduced by 90% on the 18th day in fungal cultures of Phanerochaete chrysosporium in the presence of 8 mM ethylenediamine tetraacetic acid (EDTA). In the presence of 8 mM N-N-N'-N'-tetramethylenediamine (TEMED), the mineralization rate of 14C-DDT was reduced by 80%. In the presence of 2 mM or 10 mM EDTA, 95% inhibition of lignin peroxidase (LiP) mediated veratryl alcohol oxidase activity and 97% inhibition of LiP mediated iodide oxidase activity occurred. TEMED caused 79% inhibition of veratryl alcohol oxidase activity and 92% inhibition of iodide oxidase activity when the amount used was 2 mM and 10 mM, respectively. In the presence of Zn(II) with slight molar excess of the EDTA concentration, reversed the EDTA mediated non-competitive inhibition of LiP catalyzed veratryl alcohol or iodide oxidation, Zn(II) also reversed the inhibition of LiP catalyzed veratryl alcohol oxidase activity caused by chelators other than EDTA and TEMED. In addition to Zn(II), several other metal ions also relieved EDTA mediated inhibition of veratryl alcohol and iodide oxidase activity catalyzed by LiP. The ability of veratryl alcohol to inhibit iodide oxidation catalyzed by LiP showed that veratryl alcohol could inhibit LiP mediated iodide oxidase activity. Increasing the concentration of iodide was also shown to inhibit veratryl alcohol oxidation. Kinetic analysis showed that the reaction was competitive inhibition. PMID- 11254171 TI - Delineation of CTG repeats and clinical features in a Taiwanese myotonic dystrophy family. AB - Myotonic dystrophy (DM) is an inherited, autosomal dominant muscular disease which is primarily caused by a CTG trinucleotide expansion mutation on chromosome 19q13.3. The size of this trinucleotide repeat is related both to the age of onset and to the severity of the clinical manifestation. This disease is very rare in Taiwan, and clinical and genetic study on DM has not yet been documented in this area. Here, we present both clinical features and degrees of CTG expansion for a Taiwanese DM family. All of the DM patients examined in this family showed obvious clinical manifestations by age 30, which included facial and limb muscle weakness with atrophy, myotonia, and ptosis. In addition, individual DM members also exhibited variable phenotypes, which may reflect the complexity of the pathogenic mechanism. Because the collection of blood specimens was considered to be an invasive procedure, a genetic study on this DM family was performed using buccal cells. Our results confirmed that four members showing classic symptoms of DM had CTG repeat expansion in the DMI locus, and that one member with ptosis and minor muscle weakness in the right foot was a normal homozygote for CTG repeat. These data demonstrate that buccal cells can provide clear and reliable results, and thus, are suitable for a family study of DM. PMID- 11254172 TI - Measurement of the size and orientation of human masseter and medial pterygoid muscles. AB - To gain a better understanding of biting and chewing performance, the size and orientation of the masseter and medial pterygoid muscles in living humans were studied. Twenty-seven young males having complete dentition, class I dental occlusion and normal muscle and jaw function were examined using magnetic resonance images of the head between the zygomatic arch and hyoid bone. The sections were parallel to the palatal plane, and the thickness was 3 mm without a gap. A computer software program (Medical Dental Image, MDI) was developed to identify and calculate the area of each cross section of the muscle, and the volume of the muscle was then estimated. The axis of the muscle was determined by connecting the centroids of the sections in the lower and upper 1/3 of the whole muscle. The effective muscle cross section area was then calculated by resectioning the muscle perpendicularly to the muscle axis. It was found that the mean masseter muscle volume was around 31 cm3, and that the mean medial pterygoid muscle volume was 11 cm3. Their mean effective cross section areas were around 6.2 cm2 and 3.5 cm2, respectively. The axis of the masseter muscle was more perpendicular to the palatal plane and parallel to the sagittal plane than was the medial pterygoid muscle. The results suggest that the use of magnetic resonance images (MRI) is an effective noninvasive measurement technique for determining the size and orientation of masseter and medial pterygoid muscles. This technique can be employed in future studies on human bite force evaluation and masticatory function. PMID- 11254173 TI - Effects of mutant Vpr/Vpx on HIV-2 assembly demonstrated by immunoelectron microscopy. AB - The virion-associated accessory proteins Vpr and Vpx of human immunodeficiency virus type 2 (HIV-2) are required for efficient viral replication. Vpr could be important for Vpx assembly. To investigate the interaction of Vpr and Vpx with respect to the effects of reverse transcriptase (RT) activity, viral particle information and Vpx expression site directed mutagenesis was carried out to construct Vpr, Vpx and double Vpr/Vpx HIV-2 mutants. These mutants were used for infection of peripheral blood mononuclear cells (PBM), human acute lymphoblastomic leukaemia cells (CEM-CM3) and HeLa CD4+ cells. Visualization of Vpx expression was carried out using FITC and gold labelling by means of laser scanning confocal microscopy and semi quantitative immunoelectron microscopy. Intracellular and extracellular localizations of Vpx were determined by means of fine structural analysis. Up to 80-90% reduction in the RT activity, total number of viral particles, and average Vpx expression was observed after infection of target cells with the Vpr mutant strains. In addition, intracellular Vpx expression was reduced to 51.2% with the Vpr mutant. Only 0.02% Vpx expression was detected after mutation at amino acid 62. These results provide evidence that Vpr or Vpr/Vpx mutants reduce RT activity and interfere with the expression of Vpx in HIV-2 particles during viral assembly. Vpr is efficient for Vpx corporation during viral assembly. PMID- 11254174 TI - Foot-and-mouth disease virus: a long known virus, but a current threat. AB - Foot-and-mouth disease virus (FMDV) was the first animal virus identified. Since then, FMDV has become a model system in animal virology and a considerable amount of information on its structure, biology and vaccinology has been obtained. However, the disease that this virus produces (FMD) still constitutes one of the main animal health concerns. In this review, we have attempted to summarise the state of the knowledge in different basic and applied areas of FMDV research, with emphasis on those aspects relevant to the control of the disease. PMID- 11254175 TI - Identification of optimal regions for phylogenetic studies on VP1 gene of foot and-mouth disease virus: analysis of types A and O Argentinean viruses. AB - An analysis of the informative content of sequence stretches on the foot-and mouth disease virus (FMDV) VPI gene was applied to two important viral serotypes: A and O. Several sequence regions were identified to allow the reconstruction of phylogenetic trees equivalent to those derived from the whole VPI gene. The optimal informative regions for sequence windows of 150 to 250 nt were predicted between positions 250 and 550 of the gene. The sequences spanning the 250 nt of the 3' end (positions 400 to 650), extensively used for FMDV phylogenetic analyses, showed a lower informative content. In spite of this, the use of sequences from this region allowed the derivation of phylogenetic trees for type A and type O FMDVs which showed topologies similar to those previously reported for the whole VP1 gene. When the sequences determined for viruses isolated in Argentina, between 1990 and 1993, were included in these analyses, the results obtained revealed features of the circulation of type A and type O viruses in the field, in the months that preceded the eradication of the disease in this country. Type A viruses were closely related to an Argentinean vaccine strain, and defined an independent cluster within this serotype. Among the type O viruses analysed, two groups were distinguished; one was closely related to the South American vaccine strains, while the other was grouped with viruses of the O3 subtype. In addition, a detailed phylogeny for type A FMDV is presented. PMID- 11254176 TI - Deletion of the UL21 gene in Pseudorabies virus results in the formation of DNA deprived capsids: an electron microscopy study. AB - We studied the morphogenesis of three pseudorabies virus mutants lacking parts of the gene homologous to the UL21 gene of the herpes simplex virus type 1. The mutants were examined in an SK-6 cell-line, in an SK-6 cell-line expressing the UL21 gene product, in porcine lung alveolar macrophages (PLAM) and in porcine nasal mucosa explants. Although on SK-6 cells and PLAM, the virus-assembly and egress of mutant virus M155, lacking almost the entire UL21 gene, was similar to that of the rescued PRV mutant, M155 producing virions containing little or no DNA (A-type particles). Virus mutants M133 and M134 (lacking 23 and 232 amino acids respectively) produced more C-type particles. In SK-6 cells stably expressing the UL21-encoded protein, all mutants produced C-type particles. All mutants produced C-type particles in nasal mucosa explants, indicating that the UL21-gene product is not essential for virus production in porcine tissue. These results support and extend previous work that indicated a role for the UL21 encoded protein in the packaging of newly replicated viral DNA. PMID- 11254177 TI - Fructosamine and glycated hemoglobin in the assessment of glycaemic control in dogs. AB - Fructosamine and glycated hemoglobin (HbA1c) concentrations were measured simultaneously in 222 dogs (96 healthy and 126 sick dogs). The dogs were divided into 3 groups according to the glucose concentration: hypo, hyper and euglycaemic dogs. Serum fructosamine concentrations were measured by the reduction test with nitroblue tetrazolium. A turbidimetric inhibition immunoassay and specific polyclonal antibodies were used to evaluate glycated hemoglobin concentrations. A significant correlation was found between glucose concentration and either fructosamine (r = 0.63, p < 0.0001) or glycated hemoglobin (r = 0.82, p < 0.0001). The correlation was higher in hyperglycaemic dogs for fructosamine (r = 0.80, p < 0.0001) and in hypoglycaemic dogs for glycated hemoglobin (r = 0.91, p < 0.005). We found a significant correlation between serum fructosamine and glycated hemoglobin (r = 0.65, p < 0.0001 ) when all the dogs were studied. A significant correlation was observed between serum fructosamine and glycated hemoglobin only in hyperglycaemic dogs (r = 0.82, p < 0.0003). Thus, fructosamine and HbA1c may be considered for use in screening tests for diabetes mellitus in dogs and clinical tests for monitoring control and evaluation of the diabetic animal's response to treatment. The choice of the analytical assay depends on the characteristic and analytical opportunities of the laboratory, as well as the number of serum samples to be analysed. PMID- 11254178 TI - Tissue and intracellular distribution of rhodanese and mercaptopyruvate sulphurtransferase in ruminants and birds. AB - Cyanide detoxification is catalysed by two enzymes: rhodanese [thiosulphate: cyanide sulphurtransferase, E.C. 2.8.1.1], and 3-mercaptopyruvate sulphurtransferase [3-MST, EC. 2.8.1.2]. In the present work, the activity of the two enzymes in the crude extracts of different tissues and in the mitochondrial and cytosolic fractions of tissues from some ruminants (camels, cattle and sheep) and birds (chickens and pigeons) have been compared. Rhodanese activity was almost exclusively present in the mitochondrial fraction. In ruminants and chickens the highest activity of rhodanese was found in the liver, followed by the kidney. In pigeons, however, the enzyme activity was the highest in the kidneys. In camels' tissues, the rhodanese activity was significantly (P < 0.05) lower than in cattle or sheep, and the enzyme activities in the two latter species were similar. The activity of 3-MST in the crude extract of tissues from camels was similar to that in sheep, but higher than that in cattle. The enzyme activity was equally distributed between the mitochondrial and cytosolic fractions in the liver and kidneys of camels, cattle and sheep. PMID- 11254179 TI - Interferon-alpha-producing cells are localized in gut-associated lymphoid tissues in transmissible gastroenteritis virus (TGEV) infected piglets. AB - Transmissible gastroenteritis virus (TGEV) infection of piglets results in a very rapid and massive release of IFN-alpha in serum and secretions. The objective of this work was to characterize the IFN-alpha-producing cells (IPC) in tissues of TGEV-infected piglets. Caesarean-derived colostrum-deprived piglets were infected orally with the TGEV virulent Miller strain and IPC were characterized in situ by immunohistochemistry, using a rabbit anti-pig IFN-alpha antiserum. IPC were almost exclusively detected in intestinal tissues and mesenteric lymph nodes (MLN), as early as 6 h post inoculation (p.i.), with a peak at 12-18 h. They disappeared by 24 h. IPC were localized between enterocytes in the small intestine epithelial layer, in the lamina propria, around the Peyer's patches and, at highest frequency, in MLN. Very few IPC were present in the spleen and popliteal lymph nodes of infected piglets. Double immunohistochemical staining for IFN-alpha and leukocyte markers on MLN cryosections showed that IPC were mainly Swine Leukocyte Antigen (SLA) class II positive, and were not stained by an anti-macrophage (SWC3a) MAb. In addition, double staining with anti-TGEV and anti-IFN-alpha MAbs showed that viral antigens were present in MLN, close to IPC. These results show for the first time the presence of IPC in gut mucosa and gut associated lymphoid tissues in response to an enteropathogenic virus. Moreover, this work shows that IFN-alpha released in serum is likely to originate almost exclusively from gut IPC triggered locally by viral antigens to produce IFN alpha, since there were very few IPC in spleen or peripheral lymph nodes. MHC class II molecule expression by gut-associated IPC suggests that these cells may be the in vivo mucosal counterparts of the dendritic cells recently shown to produce IFN-alpha after in vitro viral induction. PMID- 11254180 TI - Residual foot-and-mouth disease virus antibodies in French cattle and sheep six years after the vaccination ban. AB - A serological survey was carried out on French cattle to establish a reference pattern of residual vaccine antibodies and non-specific reactions against the foot-and-mouth disease virus 6 years after the ban on vaccination and in the absence of any foot-and-mouth disease outbreak. Most of the multi-vaccinated cattle still displayed high titres of antibodies and up to 50% of those which had received a single injection still had antibodies. Non-specific reactors were also recorded among animals born during and after 1991. Most of them displayed low titres close to the threshold. Sheep were also tested and, as for cattle, 4.6% of non-specific reactors were recorded, with titres close to the threshold for two thirds of them. As part of these animals have been resampled and retested, sera revealed negative confirming that these animals are true non-specific reactors. Serological testing as a mean of FMD control should take these facts into account. PMID- 11254181 TI - Responses of Fasciola hepatica infected sheep to various infection levels. AB - The response to Fasciola hepatica was studied in sheep infected with 5, 30, 150 metacercariae. The animals were necropsied 12 weeks post-infection (p-i) for counting and measuring flukes. Cellular and humoral responses were detected by peripheral eosinophil count, peripheral blood lymphocyte proliferation with excretory-secretory products (FhESP) and ELISA. All sheep were infected at necropsy except one sheep which was infected with 5 metacercariae. Mean parasitic intensities were 40%, 44% and 27% of the infection dose in sheep infected with 5, 30, 150 metacercariae respectively. FhESP-specific lymphocyte responses of the 3 infected groups were significantly enhanced in weeks 3 and 4 p-i (p < 0.05). The kinetics of the specific humoral response were similar for the 3 infected groups but the antibody level was significantly lower in animals infected with 5 metacercariae than in the 2 other infected groups from week 5 p-i to week 12 p-i (p < 0.05). Peripheral eosinophil count was significantly enhanced (p < 0.05) in infected groups. The numbers of peripheral eosinophils were significantly different between the 3 infected groups in week 3, 4 and 6 p-i and were related to infection level. These results confirm that sheep are highly susceptible to F. hepatica infection, even when infection pressure is very low. Peripheral eosinophilia was dependent of the infection level. The immune response was similar in sheep infected with various numbers of flukes. PMID- 11254182 TI - IgG response against infective larvae of Dirofilaria immitis in experimentally infected cats. AB - Somatic antigens from third stage larvae of Dirofilaria immitis (SL3) were used to detect IgG response against heartworm infection in 8 experimentally infected cats. A moderate specific anti-SL3 IgG response was found one month post infection. Afterwards, antibodies decreased reaching a basal level 4 months post infection and remained at this level until the end of the study. 6 months post infection. Western blot analysis showed specific recognition of polypeptides of 79, 73, 60, 52, 40 and 39 kDa by sera from infected cats 1 month post-infection, but not by sera taken prior to the infection. The low antigenicity of the SL3 antigen in the cat should allow the parasite to escape the host's immune response. PMID- 11254184 TI - Determination of free-form of cocaine in rat brain by liquid chromatography electrospray mass spectrometry with in vivo microdialysis. AB - A rapid liquid chromatography-electrospray mass spectrometry (LC-ES-MS) method with in vivo microdialysis for the determination of free-form of cocaine (COC) in rat brain has been developed. A C18 column and a gradient elution were employed for the separation. The [M+H]+ (m/z=304) and a fragmented ion (m/z=182) were detected using positive ion mode detection. Selective ion monitoring was utilized for quantitative measurement. The linearity of this assay was good ranging from 0.01 to 1.0 microM (r2=0.999). The inter- and intra-day precisions showed relative standard deviations ranging from 1.0% to 3.3% and 1.0% to 3.6%, respectively. In addition, the detection of one COC metabolite, benzoylecgonine (BE), by this assay was also investigated. The linearity, precision, and detection limit associated with this method for BE were determined. The application of this newly developed method was demonstrated by examining the pharmacokinetics of COC in rat brain. PMID- 11254183 TI - Capillary electrophoresis-based immunoassay for insulin antibodies with near infrared laser induced fluorescence detection. AB - A noncompetitive capillary electrophoresis (CE)-based immunoassay with near infrared laser induced fluorescence detection (NIR-LIF) for insulin antibodies has been developed. In the assay, insulin was derivatized with a NIR fluorescent dye (NN382; LI-COR). Insulin antibodies were detected via the formation of an immunocomplex. Parameters affecting the separation such as pH, voltage and ionic strength were investigated. Furthermore, it was found that increasing the ramp time of the applied voltage improved the detection limit of the assay by an order of magnitude. The detection limit of the assay was 1.1 nM. PMID- 11254185 TI - High-performance liquid chromatography with electrochemical detection for the determination of 7-monohydroxyethylrutoside in plasma. AB - MonoHER (7-monohydroxyethyl rutoside) is a semisynthetic flavonoid, which can be used as a modulator for doxorubicin-induced cardiotoxicity. To study the pharmacokinetics of monoHER in mice and human an HPLC procedure was developed to measure the level of monoHER in plasma. After extraction of monoHER with methanol, the supernatant was equally diluted (v/v) with 25 mM phosphate buffer (pH 3.33). This solution was analysed by HPLC, using a reversed-phase ODS column, with a mobile phase consisting of 49% methanol and 51% of an aqueous solution containing 10 mM sodium dihydrogen phosphate (pH 3.4), 10 mM acetic acid and 36 microM EDTA. The retention time of monoHER was about 5.2 min. The lower limit of quantification of monoHER was set at 0.3 microM and the calibration line was linear up to 75 microM. The within-day accuracy and precision of the quality control samples (0.45, 1.0, 10 and 40 microM) were better than 15 and 13%, respectively. The between-day accuracy and precision were less than 3, 20%, respectively. The recovery of monoHER (using quality control concentrations) was concentration independent and ranged from 90.5 to 95.3% except for the lowest quality control, 0.45 microM, of which the recovery was 85%. The concentration of monoHER in plasma decreased with 10% when stored at -80 degrees C for one month and with 20% when stored at -20 degrees C for 3 weeks. The repeated injection of monoHER in aliquots of 10 microM, stored in the autosampler tray (4 degrees C), showed a consistent decrease during a run: 15% over 24 h. To compensate for this decrease, sample duplicates were analysed in a mirror image sequence. PMID- 11254186 TI - Quantitation of reduced and total glutathione at the femtomole level by high performance liquid chromatography with fluorescence detection: application to red blood cells and cultured fibroblasts. AB - A new rapid and highly sensitive HPLC method with ortho-phthalaldehyde (OPA) pre column derivatization has been developed for determination of reduced glutathione (GSH) and total glutathione (GSHt) in human red blood cells and cultured fibroblasts. OPA derivatives are separated on a reversed-phase HPLC column with an acetonitrile-sodium acetate gradient system and detected fluorimetrically. An internal standard (glutathione ethyl ester) is added to facilitate quantitation. Total glutathione is determined after reduction of disulfide groups with dithiothreitol; the oxidized glutathione (GSSG) concentration is calculated by subtraction of the GSH level from the GSHt level. The assay shows high sensitivity (50 fmol per injection, the lowest reported), good precision (C.V. <5.0%), an analytical recovery of GSH and GSSG close to 100%, and linearity (r > 0.999). This HPLC technique is very simple and rapid. Its wide applicability and high sensitivity make it a convenient and reliable method for glutathione determination in various biological samples. PMID- 11254187 TI - Development and validation of a column-switching high-performance liquid chromatographic method for the determination of sanfetrinem in rat and dog plasma by direct injection. AB - A direct injection column-switching HPLC method was developed and validated for quantification of sanfetrinem in rat and dog plasma. Following dilution with buffer, samples were directly injected onto the system. The analyte was retained in an enrichment column while endogenous plasma components were eluted to waste. Sanfetrinem was then back-flushed to the analytical column for separation and quantification with an ultraviolet detector. Sample batch size was increased by adding a washing phase of the enrichment column and by alternating the injections between two enrichment columns. The method is very simple and sample preparation is minimal. The method has been fully validated and shown to be specific, accurate and reproducible. PMID- 11254188 TI - Narrowbore high-performance liquid chromatography for the simultaneous determination of sildenafil and its metabolite UK-103,320 in human plasma using column switching. AB - A fully automated narrowbore high-performance liquid chromatography method with column switching was developed for the simultaneous determination of sildenafil and its active metabolite UK-103,320 in human plasma samples without pre purification. Diluted plasma sample (100 microl) was directly introduced onto a Capcell Pak MF Ph-1 column (20x4 mm I.D.) where primary separation occurred to remove proteins and concentrate target substances using 15% acetonitrile in 20 mM phosphate solution (pH 7). The drug molecules eluted from the MF Ph-1 column were focused in an intermediate column (35x2 mm I.D.) by a valve switching step. The substances enriched in the intermediate column were eluted and separated on a phenyl-hexyl column (100x2 mm I.D.) using 36% acetonitrile in 10 mM phosphate solution (pH 4.5) when the valve status was switched back. The method showed excellent sensitivity (detection limit of 10 ng/ml), good precision (RSD < or = 2.3%) and accuracy (bias: +/-2.0%) and speed (total analysis time 17 min). The response was linear (r2 > or = 0.999) over the concentration range 10-1000 ng/ml. PMID- 11254189 TI - Application of a new analytical method using gas chromatography and gas chromatography-mass spectrometry for the azide ion to human blood and urine samples of an actual case. AB - We have established a practical and reliable method to identify and quantify the azide ion in human whole blood and human urine by transforming the ion into pentafluorobenzyl azide (PFBN3). PFBN3 was simply derived from a reaction of the ion with an excess amount of pentafluorobenzyl bromide (PFBBr). The excess amount of PFBBr was removed from the products by its reaction with sodium thiosulfate. PFBN3 in the sample was detected in high sensitivity by gas chromatography with nitrogen-phosphorus detector (GC-NPD) and gas chromatography-mass spectrometry (GC-MS). The lower detection limits of the ion by GC-NPD were 5 ng/ml for human whole blood sample and 0.5 ng/ml for human urine sample at S/N=3. On the other hand, they were 100 ng/ml for human whole blood sample and 10 ng/ml for human urine sample by the full-scan mode of GC-MS. The analytical method was applied to identification and quantification of the ion in the actual whole blood and urine samples of the victims in an actual criminal case. PMID- 11254190 TI - Optimized determination of trace jet fuel volatile organic compounds in human blood using in-field liquid-liquid extraction with subsequent laboratory gas chromatographic-mass spectrometric analysis and on-column large-volume injection. AB - A practical and sensitive method to assess volatile organic compounds (VOCs) from JP-8 jet fuel in human whole blood was developed by modifying previously established liquid-liquid extraction procedures, optimizing extraction times, solvent volume, specific sample processing techniques, and a new on-column large volume injection method for GC-MS analysis. With the optimized methods, the extraction efficiency was improved by 4.3 to 20.1 times and the detection sensitivity increased up to 660 times over the standard method. Typical detection limits in the parts-per-trillion (ppt) level range were achieved for all monitored JP-8 constituents; this is sufficient for assessing human fuels exposures at trace environmental levels as well as occupational exposure levels. The sample extractions are performed in the field and only solvent extracts need to be shipped to the laboratory. The method is implemented with standard biological laboratory equipment and a modest bench-top GC-MS system. PMID- 11254191 TI - Determination of salicylate, gentisic acid and salicyluric acid in human urine by capillary electrophoresis with laser-induced fluorescence detection. AB - Acetylsalicylic acid (Aspirin) is rapidly metabolized to salicylic acid (salicylate) and other compounds, including gentisic acid and salicyluric acid. Monitoring of salicylate and its metabolites is of toxicological, pharmacological and biomedical interest. Three capillary electrophoresis (CE) methods featuring alkaline aqueous buffers, laser-induced fluorescence (LIF) detection and no solute extraction or derivatization have been explored. A competitive binding, electrokinetic capillary-based immunoassay is developed that recognizes the presence of salicylate and gentisic acid in urine. Differentiation of the two compounds, however, is problematic. With appropriate ultraviolet excitation, many salicylate-related compounds are fluorescent so that CE with direct urine injection and LIF detection permits the determination of salicylate, gentisic acid and salicyluric acid. Using a HeCd laser with 325 nm produces interference free monitoring of all three compounds. Using 257 nm excitation from a frequency doubled Ar ion laser, native fluorescence of an endogenous urinary compound that co-migrates with gentisic acid is observed. With wavelength-resolved fluorescence detection, however, the two substances are distinguished. Furthermore, this technique, with comparison to literature data, permits the putative assignment of several peaks to other salicylate metabolites, namely glucuronide conjugates of salicylate and salicyluric acid. All three CE-LIF techniques have been applied to toxicological patient urines and urines collected after ingestion of 500 mg acetylsalicylic acid. CE results compare favorably with those obtained by a commercial fluorescence polarization immunoassay and by a conventional photometric assay. PMID- 11254192 TI - Development of liquid chromatographic method for the analysis of kanamycin residues in varicella vaccine using phenylisocyanate as a derivatization reagent. AB - A liquid chromatographic method for the determination of the aminoglycoside kanamycin in varicella vaccine is described. Kanamycin sulfate was derived with phenylisocyanate (PIC) and triethylamine for 10 min at 70 degrees C and chromatographed on a alkylamide-bonded column, Suplex pKb-100. A derivative of kanamycin sulfate was attached to four phenylisocynato groups and that molecular mass was confirmed with liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS). The kanamycin-PIC derivative was found to have a retention time of 11.7 min using an eluent composed of 40% acetonitrile in water at 1.2 ml/min column flow-rate. Detection was at a wavelength of 240 nm. Recoveries ranging from 97.5 to 99.8% were found. The correlation coefficient was greater than 0.9998 over the range between 10 and 100 microg/ml. The method precision of within-day assay showed a 0.5 to 4.0% coefficient of variation (n = 5) ranging from 10 to 70 microg/ml of kanamycin concentration levels. Kanamycin PIC derivative in reaction solution was stable for 24 h at room temperature. A simple and efficient method for the analysis of the kanamycin in varicella vaccine was developed and validated. PMID- 11254193 TI - Evaluation of urinary dihydrocodeine excretion in human by gas chromatography mass spectrometry. AB - Urinary metabolic pattern after the therapeutic peroral dose of dihydrocodeine tartrate to six human volunteers has been explored. Using the GC-MS analytical method, we have found that the major part of the dose administered is eliminated via urine within the first 24 h. However, the analytical monitoring of dihydrocodeine and its metabolites in urine was still possible 72 h after the dose was administered. The dihydrocodeine equivalent amounts excreted in urine in 72 h ranged between 32 and 108% of the dose, on average 62% in all individuals. The major metabolite excreted into urine was a 6-conjugate of dihydrocodeine, then in a lesser amount a 6-conjugate of nordihydrocodeine (both conjugated to approximately 65%). The O-demethylated metabolite dihydromorphine was of a minor amount and was 3,6-conjugated in 85%. Traces of nordihydromorphine and hydrocodone were confirmed as other metabolites of dihydrocodeine in our study. This information can be useful in interpretation of toxicological findings in forensic practice. PMID- 11254195 TI - Improved assay method for the determination of pyronaridine in plasma and whole blood by high-performance liquid chromatography for application to clinical pharmacokinetic studies. AB - An improved high-performance liquid chromatography method using a diisopropyl-C14 reversed-phase column (Zorbax Bonus-RP column) and a liquid-liquid extraction technique with UV detection is presented for the analysis of pyronaridine in human whole blood and plasma. Tribasic phosphate buffer (50 mM, pH 10.3) and diethyl ether were used for liquid-liquid extraction. The mobile phase consists of acetonitrile-0.08 M potassium dihydrogen phosphate buffer (13:87, v/v) with the pH 2.8 adjusted by orthophosphoric acid. Amodiaquine was found to be a suitable internal standard for the method. The quantification limit with UV detection at 275 nm was 3 ng on-column for both plasma and blood samples. The method was applied to plasma and blood specimens from a rabbit after a single intramuscular dose of pyronaridine tetraphosphate (20 mg/kg as base). From this in vivo study, evidence was found that pyronaridine is concentrated in blood cells, with a blood:plasma ratio ranging from 4.9 to 17.8. We conclude that blood is the preferred matrix for clinical pharmacokinetic studies. PMID- 11254194 TI - On-line, continuous and automatic monitoring of extracellular malondialdehyde concentration in anesthetized rat brain cortex. AB - An assay for in vivo, continuous and automatic monitoring of extracellular malondialdehyde concentrations in anesthetized rat brain cortex was developed. This method involved the use of microdialysis perfusion, on-line derivatization and on-line high-performance liquid chromatographic analysis. Microdialysate from an implanted microdialysis probe was on-line reacted with thiobarbituric acid at 80 degrees C for 10 min prior to on-line collection and automatic injection into a HPLC system equipped with a fluorescence detector. This method gave a linear response between the concentrations of the malondialdehyde in the microdialysates and the TEP solution where the microdialysis probe was placed. This method was used to observe the increased extracellular malondialdehyde production following elevated extracellular glutamate levels, which were achieved by perfusion of L trans-pyrrolidine-2,4-dicarboxylate, a competitive inhibitor of glutamate uptake transporter. PMID- 11254196 TI - Separation of olanzapine, carbamazepine and their main metabolites by capillary electrophoresis with pseudo-stationary phases. AB - Conditions were worked out for the separation of carbamazepine, olanzapine, and their main metabolites carbamazepine 10,11-epoxide, 10-hydroxycarbamazepine, and desmethylolanzapine. The separation was based on electrokinetically driven methods in the capillary format. The main difficulty in separating these compounds is related to their different chemical classes. Whereas the carbamazepine members are amides, and are electrically neutral, the olanzapine members have aliphatic amino groups and are thus cationic under most experimental conditions. Different additives were applied as pseudo-stationary phases to implement selectivity. Poly(diallyldimethylammonium), PDADMA, is a polycationic replaceable and soluble polymer, that interacts mainly according to the polarisability of the analyte molecules. The MEKC principle was applied with the common SDS as micelle former. In both systems, only partial resolution of the analytes was obtained. The most favorable system consisted of a charged, oligomeric additive: full separation of all analytes within 4 min was achieved with heptakis-6-sulfato-beta-cyclodextrin (7 mM) in 30 mM borate buffer, pH 8.5. PMID- 11254197 TI - Assay of human platelet guanylate cyclase activity by high-performance liquid chromatography with fluorescence derivatization. AB - A selective and sensitive high-performance liquid chromatography (HPLC) method with fluorescence derivatization for the assay of guanylate cyclase (GC) activity is described. GTP and cGMP, which are the substrate and the product of GC, respectively, and other guanine-containing compounds are selectively converted by the reaction with (3,4-dimethoxyphenyl)glyoxal to the fluorescent derivatives. The derivatives were separated by reversed-phase HPLC. The limit of detection at a signal-to-noise ratio of 3 for cGMP was 10 fmol on the column. The sensitivity of this method was less than that of the conventional radioisotopic method, but this method is simple and convenient. Human platelet GC activity was measured, and the effects of some compounds were investigated. PMID- 11254198 TI - Quantitative determination of olanzapine in rat brain tissue by high-performance liquid chromatography with electrochemical detection. AB - A sensitive assay was developed for the measurement of olanzapine in rat brain tissue using HPLC with electrochemical detection. The assay has a lower limit of quantitation of 0.5 ng/ml in tissue homogenate and utilizes a liquid-liquid extraction followed by reversed-phase HPLC for the quantitative analysis of olanzapine. The method provided a linear response for olanzapine over a concentration range of 0.5-100 ng/ml with a coefficient of determination (r2) greater than 0.9995. The extraction efficiencies of olanzapine and internal standard (LY170158) were greater than 82% in brain tissue. The intra-assay and inter-assay relative errors ranged from -5.38 to 17.60% and -3.25 to 10.53%, respectively. The intra-assay and inter-assay RSD values were in the range of 1.12 to 6.96% and 3.78 to 6.68%. Long-term stability studies showed that brain tissue homogenate samples spiked with olanzapine and internal standard are stable at -70 degrees C for at least 110 days. However, a room temperature stability study showed that olanazapine was not stable in brain homogenate if the sample was exposed at 25 degrees C longer than 2 h. This method has been used for the study of the disposition and pharmacokinetics of olanzapine in male Sprague Dawley rats. PMID- 11254199 TI - Characterization of immunoreactive acetyl-Ser-Asp-Lys-Pro in human plasma and urine by liquid chromatography-electrospray mass spectrometry. AB - The tetrapeptide AcSDKP, a natural and specific substrate of angiotensin I converting enzyme (ACE), is a negative regulator of hematopoiesis. AcSDKP has been measured in various biological media using an enzyme immunoassay (EIA), but its presence in human plasma and urine has not been formally established. By using immunoaffinity extraction and liquid chromatography-electrospray mass spectrometry, we demonstrate that AcSDKP-like immunoreactivity measured with EIA in plasma and urine samples from untreated, captopril- (an ACE inhibitor) and AcSDKP-treated subjects corresponds to AcSDKP. The present study confirms that AcSDKP is naturally present in human plasma and urine and that EIA is reliable for its measurement in such media. PMID- 11254201 TI - Development and validation of a high-performance liquid chromatographic method for the determination of gamma-hydroxybutyric acid in rat plasma. AB - A method for the determination of gamma-hydroxybutyric acid (GHB) in rat plasma was developed using solid-phase extraction (SPE) and high-performance liquid chromatography (HPLC) with UV detection. GHB was isolated from plasma using strong anion-exchange SPE columns. The chromatographic separation was performed on a C18 Aqua column. The lower limit of quantification was 10 microg/ml using 60 microl of plasma. The linearity of the calibration curves was satisfactory as indicated by correlation coefficients of >0.990. The within-day and between-day precision were <10% (n=24), the accuracy was nearly 101%. Plasma concentrations in rats after GHB infusion determined by HPLC-UV were compared with the corresponding concentrations determined with a validated gas chromatographic-mass spectrometric method by orthogonal distance regression. A good correlation was observed and a t-test indicated no significant differences from 0 and 1 for the intercept and slope, respectively. PMID- 11254200 TI - Automated high-performance liquid chromatography method for the determination of rosiglitazone in human plasma. AB - A robust, fully automated assay procedure for the determination of rosiglitazone (I, BRL-49653) in human plasma has been developed. Plasma concentrations of I were determined using automated sequential trace enrichment of dialysates (ASTED) coupled to reversed-phase high-performance liquid chromatography. Sequential automated dialysis of human plasma samples was followed by concentration of the dialysate by trace enrichment on a C18 cartridge. Drug and internal standard, SB 204882 (II) were eluted from the trace enrichment cartridge by mobile phase (0.01 M ammonium acetate, pH 8-acetonitrile, 65:35, v/v) onto the HPLC column (a Novapak C18, 4 microm, 100x5 mm radial compression cartridge) protected by a Guard-Pak C18 cartridge. The compounds were detected by fluorescence detection, using an excitation wavelength of 247 nm, and emission wavelength of 367 nm. The lower limit of quantitation of the method was 3 ng/ml (200 microl aliquot) with linearity demonstrated up to 100 ng/ml. Within- and between-run precision and accuracy of determination were better than 10% across the calibration range. There was no evidence of instability of I in human plasma following three complete freeze-thaw cycles and samples can be safely stored for at least 7 months at -20 degrees C. This method has been successfully utilised to provide pharmacokinetic data throughout the clinical development of rosiglitazone. PMID- 11254202 TI - Quantitation of itraconazole in rat heparinized plasma by liquid chromatography mass spectrometry. AB - A liquid chromatographic-mass spectrometric (LC-MS) assay was developed and validated for the determination of itraconazole (ITZ) in rat heparinized plasma using reversed-phase HPLC combined with positive atmospheric pressure ionization (API) mass spectrometry. After protein precipitation of plasma samples (0.1 ml) with acetonitrile containing nefazodone as an internal standard (I.S.), a 50 microl aliquot of the supernatant was mixed with 100 microl of 10 mM ammonium formate (pH 4.0). An aliquot of 25 microl of the mixture was injected onto a BDS Hypersil C18 column (50x2 mm; 3 microm) at a flow-rate of 0.3 ml/min. The mobile phase comprising of 10 mM ammonium formate (pH 4) and acetonitrile (60:40, v/v) was used in an isocratic condition, and ITZ was detected in single ion monitoring (SIM) mode. Standard curves were linear (r2 > or = 0.994) over the concentration range of 4-1000 ng/ml. The mean predicted concentrations of the quality control (QC) samples deviated by less than 10% from the corresponding nominal values; the intra-assay and inter-assay precision of the assay were within 8% relative standard deviation. Both ITZ and I.S. were stable in the injection solvent at room temperature for at least 24 h. The extraction recovery of ITZ was 96%. The validated assay was applied to a pharmacokinetic study of ITZ in rats following administration of a single dose of itraconazole (15 mg/kg). PMID- 11254203 TI - Determination of aloenin, barbaloin and isobarbaloin in aloe species by micellar electrokinetic chromatography. AB - Aloenin, barbaloin and isobarbaloin in JP Aloe, Aloe barbadensis (Aloe vera) and Aloe arborescens Miller var. natalensis Berger (Aloe arborescens Miller) were determined by micellar electrokinetic chromatography (MEKC) with 50 mM sodium dodecyl sulfate. Aloenin, barbaloin and isobarbaloin were well separated by MEKC and as little as 5.5 pg/11 nl of the three compounds could be detected. The determination took around 14 min. PMID- 11254204 TI - Resolution of isomers of sorbitolparaben esters by chromatographic and electrophoretic techniques. AB - Pharmaceutical preparations usually contain preservatives and sweeteners. When parabens are used as preservatives and a polyol as a sweetener, a transesterification reaction may happen, yielding the transester polyol-paraben. The products formed in the transesterification reaction of methylparaben and sorbitol were analyzed by micellar electrokinetic chromatography and by HPLC. Up to six positional isomers of sorbitolparaben (SPB) can be produced. However, only three peaks were found by HPLC. The higher efficiency and resolution power of MEKC allowed one to resolve five peaks. Results were compared with those obtained by capillary zone electrophoresis in borate buffer, where the separation of isomers occurred in a different way, because of a complexation between SPB and borate. PMID- 11254205 TI - Short-strong hydrogen bonds and a low barrier transition state for the proton transfer reaction in RNase A catalysis: a quantum chemical study. AB - There is growing evidence that some enzymes catalyze reactions through the formation of short-strong hydrogen bonds as first suggested by Gerlt and Gassman. Support comes from several experimental and quantum chemical studies that include correlation energies on model systems. In the present study, the process of proton transfer between hydroxyl and imidazole groups, a model of the crucial step in the hydrolysis of RNA by the enzymes of the RNase A family, is investigated at the quantum mechanical level of density functional theory and perturbation theory at the MP2 level. The model focuses on the nature of the formation of a complex between the important residues of the protein and the hydroxyl group of the substrate. We have also investigated different configurations of the ground state that are important in the proton transfer reaction. The nature of bonding between the catalytic unit of the enzyme and the substrate in the model is investigated by Bader's atoms in molecule theory. The contributions of solvation and vibrational energies corresponding to the reactant, the transition state and the product configurations are also evaluated. Furthermore, the effect of protein environment is investigated by considering the catalytic unit surrounded by complete proteins--RNase A and Angiogenin. The results, in general, indicate the formation of a short-strong hydrogen bond and the formation of a low barrier transition state for the proton transfer model of the enzyme. PMID- 11254206 TI - Characterization of the molecular motions of constitutively active G protein coupled receptors for parathyroid hormone. AB - The molecular mechanism of constitutive activity of the G protein-coupled receptor for human parathyroid hormone (PTH1) has been examined by molecular dynamics (MD) simulations. The single point mutations H223R, T410P, and I458R, of the PTH1 receptor result in ligand-independent receptor activation. Extensive MD simulations indicate that each of the mutations, through different mechanisms, lead to very similar conformational changes of the third intracellular loop. The structural changes, centered on K405 in the C-terminus of the third intracellular loop, can be traced back to the single-point mutations by calculation of the forces and torques responsible for the collective motions of the receptor. This analysis indicates a direct correlation between the conformational preferences of the cytoplasmic loop and the mutations in different locations of the receptor: TM2 (H223R), TM6 (T410P), and TM7 (1458R). Given the pivotal role of the third intracellular loop of PTH1 in coupling to the G proteins, the structural changes induced by these single-point mutations may be responsible for the ligand-free activation of the receptor. These results coupled with the high-resolution structure of the third cytoplasmic loop of PTH1, previously determined in our laboratory, provide unique insight into the mechanism of ligand free activation of the PTH1 receptor. PMID- 11254207 TI - Molecular characterization of a laminin-derived oligopeptide with implications in biomimetic applications. AB - The molecular properties of the laminin-derived oligopeptide, H-CDPGYIGSR-NH2, have been investigated with the aid of a tandem computer simulation/experimental approach. The simulation studies placed a particular emphasis on studying the oligopeptide in aqueous media, as well as in a grafted or immobilized state. The simulations revealed the presence of a stable double hydrogen bond between arginine (R) and aspartic acid (D) residues. The mutation of the terminal arginine with lysine, another hydrophilic and positively charged amino acid, resulted in a drastic structural change, thus suggesting a major role of the terminal arginine residue in the overall oligopeptide's conformation and, hence, its bioactivity. In addition, the involvement of the aspartic acid residue in overall peptide structural stabilization also illustrates a previously undetermined role for this region (i.e. CDPG) of the oligopeptide. A subsequent in vitro experiment demonstrated a significant loss of bioactivity upon mutating the terminal residue from arginine to lysine, thereby corroborating the overall findings of the computational model. PMID- 11254208 TI - Molecular dynamics simulations of urea and thermal-induced denaturation of S peptide analogue. AB - Molecular dynamics simulations of the S-peptide analogue AETAAAKFLREHMDS in water at 278 and 358 K, and in 8 M urea at 278 K were performed. The results show agreement with experiments. The helix is stable at low temperature (278 K), while at 358 K, unfolding is observed. The effects of urea on protein stability have been studied. The data support a model in which urea denatures proteins by: (1) diminishing the hydrophobic effect by displacing water molecules from the solvent shell around nonpolar groups; and (2) binding directly to amide units (NH and CO groups) via hydrogen bonds. The results of cluster analysis and essential dynamics analysis suggest that the mechanism of urea and thermal-induced denaturation may not be the same. PMID- 11254210 TI - Temperature-induced conformational changes in amyloid beta(1-40) peptide investigated by simultaneous FT-IR microspectroscopy with thermal system. AB - Temperature-dependent secondary structures of the amyloid beta(1-40) peptide in the solid state were studied by simultaneous Fourier transform infrared/differential scanning calorimetry (FT-IR/DSC) microspectroscopic system with the heating-cooling-reheating cycle. The result indicates that a thermal transition temperature at 45 degrees C was easily obtained from the three dimensional plot of the transmission FT-IR spectra as a function of temperature. Furthermore, the thermal-dependent conformational transformations, due to denaturation and aggregation, of solid amyloid beta(1-40) were mainly evidenced by reducing the compositions from 37 to 20-24% for alpha-helical and random coil structures but increasing the components from 27 to 45% for intermolecular beta sheet structures. Thermal-irreversible behavior and a poor thermal stability of solid amyloid beta(1-40) were also observed from the poor restoration of the secondary conformational changes in the heated sample. PMID- 11254209 TI - Cholesterol favors phase separation of sphingomyelin. AB - The phase behavior of mixed lipid dispersions representing the inner leaflet of the cell membrane has been characterized by X-ray diffraction. Aqueous dispersions of phosphatidylethanolamine:phosphatidylserine (4:1 mole/mole) have a heterogeneous structure comprising an inverted hexagonal phase H(II) and a lamellar phase. Both phases coexist in the temperature range 20-45 degrees C. The fluid-to-gel mid-transition temperature of the lamellar phase assigned to phosphatidylserine is decreased from 27 to 24 degrees C in the presence of calcium. Addition of sphingomyelin to phosphatidylethanolamine/phosphatidylserine prevents phase separation of the hexagonal H(II) phase of phosphatidylethanolamine but the ternary mixture phase separates into two lamellar phases of periodcity 6.2 and 5.6 nm, respectively. The 6.2-nm periodicity is assigned to the gel phase enriched in sphingomyelin of molecular species comprising predominantly long saturated hydrocarbon chains because it undergoes a gel-to-fluid phase transition above 40 degrees C. The coexisting fluid phase we assign to phosphatidylethanolamine and phosphatidylserine and low melting point molecular species of sphingomyelin which suppresses the tendency of phosphatidylethanolamine to phase-separate into hexagonal H(II) structure. There is evidence for considerable hysteresis in the separation of lamellar fluid and gel phases during cooling. The addition of cholesterol prevents phase separation of the gel phase of high melting point sphingomyelin in mixtures with phosphatidylserine and phosphatidylethanolamine. In the quaternary mixture the lamellar fluid phase, however, is phase separated into two lamellar phases of periodicities of 6.3 and 5.6 nm (20 degrees C), respectively. The lamellar phase of periodicity 5.6 nm is assigned to a phase enriched in aminoglycerophospholipids and the periodicity 6.3 nm to a liquid-ordered phase formed from cholesterol and high melting point molecular species of sphingomyelin characterized previously by ESR. Substituting 7-dehydrocholesterol for cholesterol did not result in evidence for lamellar phase separation in the mixture within the temperature range 20-40 degrees C. The specificity of cholesterol in creation of liquid-ordered lamellar phase is inferred. PMID- 11254211 TI - Dissecting the free energy of formation of the 1:1 actomyosin complex. AB - The behaviour of solutions of pure myosin, of pure F-actin and of the equimolar mixture of myosin and of F-actin is studied. It is found that the chemical potential of the two proteins, in separate solutions, increases monotonically with the increase of protein osmotic pressure. A method is presented to determine the chemical potential of the 1:1 actin-myosin complex formed from equimolar solutions of myosin and of F-actin (as monomer). This is the first evaluation of the chemical potential of actomyosin under conditions similar to those of skeletal muscle. It is found that the filament suspensions of myosin and of the 1:1 actin-myosin complex display a high non-ideal behavior as well as distinctly different energy profiles as a function of protein osmotic pressure. This supports the hypothesis that, in muscle: (a) detached cross-bridge change significantly their free energy when sarcomere is shifting from the relaxed to the active or to the rigor state; and (b) the cross-bridge attachment-detachment process is accompanied by changes of muscle protein osmotic pressure. PMID- 11254212 TI - A kinetic study on the interaction of deprotonated purine radical cations with amino acids and model peptides. AB - By use of pulse radiolysis techniques, the radical cations of purine nucleotides have been successfully produced by the SO4- ion oxidation. Time-resolved spectroscopic evidence is provided that the one-electron-oxidized radicals of dAMP and dGMP can be efficiently repaired by aromatic amino acids (including tyrosine and tryptophan) via electron transfer reaction. As a model peptide, Arg Tyr-AcOH was also investigated with regard to its interaction with deprotonated purine radical cations. The rate constants of the electron transfer reactions were determined to be (1 approximately 5) x 10(8) dm(3) mol(-1) s(-1). These results suggest that the aromatic amino acids in DNA-associated proteins may play some role in electron transfer reactions through DNA. PMID- 11254213 TI - Thermodynamics of interaction of water vapour with 20 different poly-L-amino acids. AB - The uptake of water vapour by 20 different polyaminoacids have been evaluated by an isopiestic vapour pressure technique in absence of solute at three different temperatures (22 degrees C, 30 degrees C and 37 degrees C). The water vapour adsorption isotherm for different polyaminoacids in the range of water activity varying between zero and unity apparently agreed with that expected from a type III BET isotherm. From the linear BET plots, obeyed in the lower range of water activity, the BET constants n(m) and Qm for different polyamines have been evaluated. The amount of water vapour adsorbed (n1) was calculated in moles/kg of polyaminoacids as well as in moles/mole of amino acid residue. Its value at unit water activity (deltan(o)1) has been evaluated by an extrapolation method and the results support that the multilayer adsorption of water vapour at the interface of polyaminoacids takes place. Values of deltan(o)1 are strictly comparable in terms of moles per kg rather than mole per mole unit. In case of beta lactoglobulin (betalg), lysozyme and BSA, theoretically obtained deltan(o)1 values were observed to be considerably larger than experimental values of deltan(o)1. This indicated that amino acid residues in the polypeptide release a large amount of water due to the formation of a globular structure. Using the Bull equation in the integrated form, standard free energies, deltaGo(w) for water-polyamino acid binding interaction at two different temperatures have been evaluated. Based on the Clausius-Clapeyron equation in an integrated form, the integral enthalpy for water-polyamino acid interaction has also been evaluated. PMID- 11254214 TI - Comparison between the thermodynamic features of alpha1-antitrypsin and human albumin partitioning in aqueous two-phase systems of polyethyleneglycol-dextran. AB - The partitioning features of human serum albumin and alpha1-antitrypsin in aqueous two-phase systems of dextran and polyethyleneglycol were studied. The effect of factors that affect the electrostatic term of Albertsson equation such as pH, ionic strength, presence of neutral salts as well as those which affect the non-electrostatic term such as polyethyleneglycol mol. wt. and temperature were assayed. At room temperature, the positive entropy and enthalpy changes associated to the partition may be due to a release of part of the structured water in the domain of proteins caused by H-bonds rupture when the proteins are transferred to the upper phase. This behaviour may be explained on the basis of a preferential hydration of the proteins in presence of dextran (bottom phase) and a preferential interaction of polyethyleneglycols with the protein domain (top phase). The electrostatic interactions were similar for both proteins due to the proximity of their isoelectric point and similar dissociation profiles of their prototropic groups. PMID- 11254215 TI - Kinetic phase behavior of distearoylphosphatidylethanolamine dispersed in glycerol. AB - Phase behavior of distearoylphosphatidylethanolamine dispersed in excess glycerol has been examined by differential scanning calorimetry. Transformation from lamellar-gel to lamellar crystalline phase was found to take place at temperatures near 74.9 degrees C upon cooling and near 76.3 degrees C during heating scans. The transition can also be observed under isothermal conditions at temperature in this range. The kinetics of the transformation from lamellar-gel to lamellar-crystal phase was analyzed by the well-known Avrami equation. The apparent Avrami exponents were found to be approximately 1.6. The effective dimensionality of the growth pattern can then be set as 1, after taking into account the contribution of nucleation at the examination temperatures. The activation energy of the phase transition was estimated as approximately 255 kJ mol(-1). The data are discussed in terms of development of successful cryoprotective strategies using glycerol. PMID- 11254217 TI - The role of the gel <=> liquid-crystalline phase transition in the lung surfactant cycle. AB - Lipid polymorphism plays an important role in the lung surfactant cycle. A better understanding of the influence of phase transitions on the formation of a lipid film from dispersions of vesicles will help to describe the mechanism of action of lung surfactant. The surface pressure (or tension) of dispersions of DPPC, DMPC, and DPPE unilamellar vesicles was studied as a function of temperature. These aggregates rapidly fuse with a clean air-water interface when the system is at their phase transition temperature (Tm), showing a direct correlation between phase transition and film formation. Based on these results, an explanation on how fluid aggregates in the alveolar subphase can form a rigid monolayer at the alveolar interface is proposed. PMID- 11254216 TI - X-ray diffraction study on ordered, disordered and reconstituted intercellular lipid lamellar structure in stratum corneum. AB - From small angle X-ray diffraction for the stratum corneum of hairless mouse, it was obtained that in the normal stratum corneum, the 1st, 2nd and 3rd order diffraction peaks for the intercellular lipid lamellar structure appear at 13.8, 6.87 and 4.59 nm, respectively and also a broad hump for the 4th order reflection appears as observed by the previous researchers. In the damaged stratum corneum prepared by the treatment of sodium dodecyl sulfate, these small-angle diffraction peaks disappear and only the broad maxima remain around the 1st, 2nd and 3rd order diffraction peaks. These facts indicate that in the normal stratum the lamellar structure is ordered and in the damaged stratum corneum the lamellar structure is disordered. Furthermore, in the reconstituted lamellar structure obtained by immersing into the dilute suspension of the mixture of ceramide 3, cholesterol and stearic acid, the 1st, 2nd and 3rd order diffraction peaks reappear at 13.3, 6.67 and 4.44 nm, respectively. This fact indicates that the reorganization of the ordered lamellar structure takes place by adding the mixture to the damaged stratum corneum. PMID- 11254218 TI - Group additivity calculations of the thermodynamic properties of unfolded proteins in aqueous solution: a critical comparison of peptide-based and HKF models. AB - A recent paper in this journal [Amend and Helgeson, Biophys. Chem. 84 (2000) 105] presented a new group additivity model to calculate various thermodynamic properties of unfolded proteins in aqueous solution. The parameters given for the revised Helgeson-Kirkham-Flowers (HKF) equations of state for all the constituent groups of unfolded proteins can be used, in principle, to calculate the partial molar heat capacity, C(o)p.2, and volume, V2(0), at infinite dilution of any polypeptide. Calculations of the values of C(o)p.2 and V2(0) for several polypeptides have been carried out to test the predictive utility of the HKF group additivity model. The results obtained are in very poor agreement with experimental data, and also with results calculated using a peptide-based group additivity model. A critical assessment of these two additivity models is presented. PMID- 11254219 TI - Response to 'Group additivity calculations of the thermodynamic properties of unfolded proteins in aqueous solution: a critical comparison of peptide-based and HKF models'. PMID- 11254220 TI - Polypeptide expression in prostate hyperplasia and prostate adenocarcinoma. AB - Cells were collected from prostate hyperplasias (n = 6) and prostate carcinomas (n = 6) and subjected to two-dimensional gel electrophoresis (2-DE). The resulting polypeptide patterns were analysed with the PDQUEST computer software. Malignant tumors showed significant increases in the level of expression of proliferating cell nuclear antigen (PCNA), calreticulin, HSP 90 and pHSP 60, oncoprotein 18(v), elongation factor 2, glutathione-S-transferase pi (GST-pi), superoxide dismutase and triose phosphate isomerase. In addition, decreases in the levels of tropomyosin-1 and 2 and cytokeratin 18 were observed in prostate carcinomas compared to prostate hyperplasias. This pattern of alterations is similar to that observed in other carcinomas in our previous studies. All malignant tumors showed simultaneous alterations in 5 or more of 9 markers studied, whereas only one case of benign hyperplasia showed alterations in 5 markers. The EST-data base for prostate tumors available from NCI (CGAP) was searched for the expression of the mRNAs corresponding to proteins identified in our gels. Large differences in the relative expression of mRNAs and proteins were observed. Our data show alterations in the pattem of polypeptide expression in prostate carcinomas which are similar to those observed in other carcinomas. PMID- 11254221 TI - Spontaneous apoptosis, oxidative status and immunophenotype markers in blood lymphocytes of AIDS patients. AB - Peripheral blood mononuclear cells (PBMC) from 251 HIV-positive drug abusers of known clinical stage and from 40 healthy donors were tested for conventional immunologic markers (CD3, CD4, CD8, CD19, CD14, CD16/CD56, CD45 and HLA-DR). Additional cell parameters and the occurrence of spontaneous apoptosis (programmed cell death) were investigated on freshly isolated PBMC by flow cytometric measurement of either annexin-V bound to plasma membrane phosphatidylserine or propidium iodide uptake. The activity of gamma glutamyltransferase (gamma-GT), an ectoenzyme contributing to the synthesis of the intracellular antioxidant glutathione (GSH) and involved in early apoptosis, was also determined in these cells. Immunocompetent T-cell counts were lower in HIV+ patients, with the exception of CD8+ and HLA-DR+ lymphocytes. The external binding of annexin-V was significantly higher in HIV+ PBMC and occurred in both CD8+ and CD4+ T-lymphocyte subsets. The activity of gamma-GT, was significantly lower in the PBMC from HIV+ patients, indicating that the redox status of PBMC may be affected in HIV+ individuals. Finally, the most dominant features characterising patients receiving antiretroviral therapy were greater long-term stability in the distribution of various cell parameters excepted the level of apoptosis. PMID- 11254222 TI - Biological characterisation of superficial bladder cancer by bivariate cytokeratin 7/DNA analysis, flow cytometric assessment of MIB- 1, and an immunohistochemical study. AB - A total of 238 cases of bladder carcinoma stages Ta, Tis, T1 were submitted prospectively to multiparameter flow cytometry and immunohistochemical study in order to determine the biological aggressiveness of the tumour. DNA index (DI), S phase fraction (SPF) obtained by bivariate cytokeratin 7/DNA analyses, and the immunohistochemical evaluation of p53 and MIB-1 were studied in relation to the traditional prognostic factors in bladder cancer (stage and grade). the variance analysis results showed that DNA aneuploidy was significantly associated with high stage (p = 0.0001), high grade (p = 0.0001), high SPF value > or = 5.5% (p = 0.0001), MIB-1 positivity > or = 31% (p = 0.0001) and high expression of p53 (staining involving > 50% of cells, p = 0.0001). Even if there was no statistical significance the hypotetraploid class (1.70 < DI < 1.89) showed poor prognostic biomarkers more frequently than the other aneuploid classes. Out of 238 cases, 101 were also submitted to flow cytometric measurement of MIB-1 (fMIB-1) to study the correlation between cell proliferation and DNA content. Data obtained from fresh, 3:1 methanol/acetone fixed samples were compared with values obtained from both cell cycle analysis methods and routine application of the MIB-1 immunostaining in histological sections. fMIB-1 values were positively correlated with SPF values (r = 0.801, p < 0.01) and S+G2M fraction (percentage of cells in S and in G2M phases) (r = 0.763, p < 0.01) but no correlation with paraffin sections was found. A fMIB-1 value > 7% was strongly associated with aneuploidy (p = 0.0001). The determination of DNA content coupled with the study of the epithelial (cytokeratin 7) and proliferative (MIB-1) markers could be useful in providing important information on the biological behaviour of superficial bladder tumours. PMID- 11254223 TI - Analysis of erythrocyte glycophorin-A variants by flow cytometry in lung disease patients detects the effect of tobacco smoke. AB - The glycophoryn A (GPA) assay evaluates somatic in vivo mutations. It is considered a cumulative biodosimeter for genotoxic exposures and is under evaluation in cancer risk assessment. GPA, a polymorphic membrane protein of the erythrocytes, determines the MN blood groups. The NO and NN variant frequencies (VF) may be detected in MN subjects (about 50% of the population) by flow cytometry using two differently labelled antibodies. We explored if GPA NO and NN VF might be relevant to the assessment of individual lung cancer risk and susceptibility, in a small population with a high prevalence of heavy tobacco smokers: 8 lung cancer patients and 16 subjects with non-malignant lung diseases associated with increased risk of lung cancer. There was a wide interindividual variability and complete overlap between non-neoplastic and neoplastic patients. A significant positive correlation was seen with smoking duration in NO VF (p = 0.04, age-adjusted). Current smokers (n = 12) displayed higher NO values than never (n = 1) or ex-smokers (n = 11), 36.3 +/- 18.2 and 21.0 +/- 13.2, respectively (p < 0.01). No association was shown with occupational exposure. The present exploratory study suggests that assessment of individual lung cancer risk and susceptibility by the GPA assay does not seem to be feasible. The assay appears to provide a biomarker of longterm exposure to tobacco smoke. PMID- 11254224 TI - Digital imaging analysis for the study of endotoxin-induced mitochondrial ultrastructure injury. AB - Primary defects in mitochondrial function have been implicated in over 100 diverse diseases. In situ, mitochondria possess unique and well-defined morphology in normal healthy cells, but diseases linked to defective mitochondrial function are characterized by the presence of morphologically abnormal and swollen mitochondria with distorted cristae. In situ study of mitochondrial morphology is established as an indicator of mitochondrial health but thus far assessments have been via subjective evaluations by trained observers using discontinuous scoring systems. Here we investigated the value of digital imaging analysis to provide for unbiased, reproducible, and convenient evaluations of mitochondrial ultrastructure. Electron photomicrographs of ileal mucosal mitochondria were investigated using a scoring system previously described by us, and also analyzed digitally by using six digital parameters which define size, shape, and electron density characteristics of over 700 individual mitochondria. Statistically significant changes in mitochondrial morphology were detected in LPS treated animals relative to vehicle control using both the subjective scoring system and digital imaging parameters (p < 0.05). However, the imaging approach provided convenient and high throughput capabilities and was easily automated to remove investigator influences. These results illustrate significant changes in ileal mucosal mitochondrial ultrastructure during sepsis and demonstrate the value of digital imaging technology for routine assessments in this setting. PMID- 11254225 TI - High altitude medicine and biology in North Chile. PMID- 11254226 TI - Acute medical problems in the Himalayas outside the setting of altitude sickness. AB - Well-recognized medical threats at high altitude (>2,500 m) include acute mountain sickness (AMS), high altitude pulmonary edema (HAPE), and high altitude cerebral edema (HACE). Thousands of travelers in the Himalayas are exposed annually to these often life-threatening syndromes. Their recognition and treatment has advanced considerably in recent years. In the Himalayas, we frequently see acute medical problems outside the setting of AMS and the two types of altitude edemas. Many of these other conditions are also hypoxia related and sometimes may mimic the classic high altitude illnesses of AMS, HAPE, and HACE. Although the vast majority of these medical problems are neurological, pulmonary and other organ system dysfunction also occur. These "non-high altitude sickness" disease entities in persons who sojourn to remote mountainous environments are reviewed in this paper to enhance their recognition, diagnosis, and treatment. PMID- 11254227 TI - Cardiovascular responses to arginine vasopressin blockade during acute hypoxemia in the llama fetus. AB - The fetal llama has a marked increase in the peripheral vascular resistance and no augmentation of brain blood flow during hypoxemia. In spite of the substantial plasma arginine-vasopressin (AVP) increase during hypoxemia, up to 8 times greater than in fetal sheep, there are no changes of carotid and femoral blood flows during hypoxemia with a V1 receptor blockade, as is seen in the fetal sheep. The aim of this study was to assess the role of AVP function in mediating the combined ventricular output and organ blood flow in the hypoxemic llama fetus. Six fetal llamas at 0.65 of gestation were instrumented under general anesthesia, and cardiorespiratory responses and blood flows determined under normoxemic and hypoxemic conditions. The AVP effect was determined using a V1 antagonist during normoxemic and hypoxemic conditions. Organ blood flows were measured with the radioactive microsphere technique. No significant differences in organ blood flow or in their vascular resistances were seen between the control and treated fetuses during hypoxemia. We conclude that V1 blockade did not have any important role in the cardiovascular response to acute hypoxemia in the llama fetus, in contrast with lowland fetuses. AVP may be playing a role in other regions, possibly in kidney or lung, during hypoxemia. PMID- 11254228 TI - Serial changes in spirometry during an ascent to 5,300 m in the Nepalese Himalayas. AB - The aims of the present study were to determine the changes in forced vital capacity (FVC), forced expiratory volume in 1 sec (FEV1) and peak expiratory flow (PEF), during an ascent to 5,300 m in the Nepalese Himalayas, and to correlate the changes with arterial oxygen saturation measured by pulse oximetry (SpO2) and symptoms of acute mountain sickness (AMS). Forty-six subjects were studied twice daily during an ascent from 2,800 m (mean barometric pressure 550.6 mmHg) to 5,300 m (mean barometric pressure 404.3 mmHg) during a period of between 10 and 16 days. Measurements of FVC, FEV1, PEF, SpO2, and AMS were recorded. AMS was assessed using a standardized scoring system. FVC fell with altitude, by a mean of 4% from sea level values [95% confidence intervals (CI) 0.9% to 7.4%] at 2,800 m, and 8.6% (95% CI 5.8 to 11.4%) at 5,300 m. FEV1 did not change with increasing altitude. PEF increased with altitude by a mean of 8.9% (95% CI 2.7 to 15.1%) at 2,800 m, and 16% (95% CI 9 to 23%) at 5,300 m. These changes were not significantly related to SpO2 or AMS scores. These results confirm a progressive fall in FVC and increase in PEF with increasing hypobaric hypoxia while FEV1 remains unchanged. The increase in PEF is less than would be predicted from the change in gas density. The fall in FVC may be due to reduced inspiratory force producing a reduction in total lung capacity; subclinical pulmonary edema; an increase in pulmonary blood volume, or changes in airway closure. The absence of a correlation between the spirometric changes and SpO2 or AMS may simply reflect that these measurements of pulmonary function are not sufficiently sensitive indicators of altitude-related disease. Further studies are required to clarify the effects of hypobaric hypoxia on lung volumes and flows in an attempt to obtain a unifying explanation for these changes. PMID- 11254229 TI - Nocturnal O2 enrichment of room air at high altitude increases daytime O2 saturation without changing control of ventilation. AB - In a randomized, double-blind study, 24 sea-level residents drove to 3,800-m altitude in 1 day, and then slept the first night in either ambient air or 24% oxygen, and the second night in the treatment that they did not receive on the first night. Oxygen enrichment, compared with ambient air, resulted in significantly fewer apneas, and significantly less time spent in periodic breathing during the night. The increase in SaO2 between evening and morning was significantly higher after sleeping in the oxygen-enriched atmosphere, compared with ambient air. However, this significant improvement in SaO2 did not persist into mid-day. The overnight treatment did not alter the ventilatory response to hypoxia or to carbon dioxide as measured the following morning. The results suggest that the elevation in SaO2 following overnight oxygen enrichment is probably not due to a change in the control of ventilation, but possibly to differences in subclinical lung pathology. PMID- 11254230 TI - Transient high altitude expressive aphasia. AB - Transient focal neurological deficits have been described in sojourners to high altitude. We present two cases of transient expressive aphasia in well acclimatized high altitude climbers. We speculate that this type of transient focal neurological impairment may represent migraine aura, and we discuss other reports of transient focal neurological deficit at high altitude. PMID- 11254232 TI - My self-portrait. PMID- 11254231 TI - The mysterious death of Dr. Jacottet on Mont Blanc. PMID- 11254233 TI - A comparison between septic bursitis caused by Staphylococcus aureus and those caused by other organisms. AB - Septic bursitis is an infection that usually involves olecranon and prepatellar bursae. Staphylococcus aureus is responsible for around 80% of cases. However, information regarding bursitis caused by non-Staphylococcus aureus microorganisms (NSAB) is scant. In this paper we describe the characteristics of NSAB and emphasise differences between these and Staphylococcus aureus bursitis (SAB). A retrospective study of all cases with septic bursitis seen between January 1991 and June 1998 at one university hospital was conducted. Only cases in which bursal fluid culture yielded growth of a microorganism were analysed. A literature review was conducted for completeness. Fifty-seven episodes of septic bursitis in 56 patients were studied: 47 of these were caused by Staphylococcus aureus and 11 by non-Staphylococcus aureus microorganisms. Forty-three SAB patients were male (91%). Mean age at diagnosis was 50 years (range 20-85 years). The presentation of bursitis had a seasonal trend, with a peak in the summer. Twenty-three patients (51%) had occupations involving frequent or sustained pressure on the bursae. Other risk factors were recent trauma in 11 (23%), alcoholism in six (13%), pre-existing bursal disease in five (11%), and chronic obstructive pulmonary disease in four (9%). There were 20 cases of olecranon bursitis (43%), 25 of prepatellar bursitis (53%) and two of first metatarsophalangeal bursitis. Characteristics of patients from the literature review were similar. Eight NSAB patients (73%) were male. Mean age at diagnosis was 46.9 (range 29-83 years). Two patients were plumbers and one a stonemason. Five (45%) had neither putative systemic nor local risk factors. There were five olecranon (45%), five prepatellar (45%), and one external malleolus bursitis. Infection by a mixed flora was common. Unlike SAB, the presentation of cases did not have a seasonal trend. The clinical spectrum of non-Staphylococcus aureus bursitis (NSAB) differs from that of Staphylococcus Aureus bursitis (SAB), and this should be considered in the initial diagnosis of septic bursitis. PMID- 11254234 TI - The effect of balneotherapy at the Dead Sea on the quality of life of patients with fibromyalgia syndrome. AB - Fibromyalgia (FS) is an idiopathic chronic pain syndrome defined by widespread non-articular musculoskeletal pain and generalised tender points. As there is no effective treatment, patients with this condition have impaired quality of life (QoL). The aim of this study was to assess the possible effect of balneotherapy at the Dead Sea area on the QoL of patients with FS. Forty-eight subjects participated in the study; half of them received balneotherapy, and half did not. Their QoL (using SF-36), psychological well-being and FS-related symptoms were assessed prior to arrival at the spa hotel in the Dead Sea area, at the end of the 10-day stay, and 1 and 3 months later. A significant improvement was reported on most subscales of the SF-36 and on most symptoms. The improvement in physical aspects of QoL lasted usually 3 months, but on psychological measures the improvement was shorter. Subjects in the balneotherapy group reported higher and longer-lasting improvement than subjects in the control group. In conclusion, staying at the Dead Sea spa, in addition to balneotherapy, can transiently improve the QoL of patients with FS. Other studies with longer follow-up are needed to support our findings. PMID- 11254235 TI - Osteoarthritis of the scaphoidtrapezium joint: an early sign of calcium pyrophosphate dihydrate disease. AB - The aim of this study was to determine the value of scaphoidtrapezium osteoarthritis (ST osteoarthritis) as an early sign of calcium pyrophosphate dihydrate disease (CPDD) in a cohort of patients undergoing surgery for osteoarthritis of the first carpometacarpal joint. We examined whether patients with cartilage calcification of the wrist at the time of operation had ST osteoarthritis, indicating CPDD at an earlier time (retrospective study), and whether patients with ST osteoarthritis but without cartilage calcification at the time of surgery develop radiological or clinical signs of CPDD at a later time (prospective study). From 1 January 1989 to 31 December 1995 a total of 169 patients (from an orthopaedic clinic) with a diagnosis of osteoarthritis of the first carpometacarpal joint were included in the study; 167 underwent surgery and two were treated without. Of the 16 patients showing calcification on surgery and therefore included in the retrospective study, 12 had prior radiographs, of which eight showed ST osteoarthritis. Among these, four had no concomitant cartilage calcification in the prior radiographs. Of the 32 patients in the prospective group having ST osteoarthritis but no calcifications at the time of surgery, 27 could be clinically examined. Of these, two showed cartilage calcifications on the follow-up radiographs of the hands. The presence of ST osteoarthritis is a helpful diagnostic finding for the diagnosis of CPDD, especially in cases without radiographic cartilage or fibrocartilage calcification of the wrist. ST osteoarthritis may then point to the correct diagnosis. PMID- 11254236 TI - Suppressed levels of serum cortisol following high-dose oral dexamethasone administration differ between healthy postmenopausal females and patients with established primary vertebral osteoporosis. AB - Hypercortisolism and glucocorticoid treatment, even in a low dose or administered topically, may influence bone metabolism. It was the aim of this study to investigate whether there might be differences in the regulation of endogenous cortisol secretion between patients with established primary vertebral osteoporosis and healthy controls. Suppressed morning serum cortisol concentrations in a 3 mg dexamethasone overnight suppression test were compared in well-defined healthy postmenopausal women (n = 149) and osteoporotic patients classified as having established primary vertebral osteoporosis with no clinical features of hypercortisolism (n = 78). Suppressed cortisol in the healthy controls was 1.08 +/- 0.44 microg/dl and in the primary osteoporotics 1.58 +/- 1.42 microg/dl (p < 0.0001). Of the investigated primary osteoporotics 15.4% (n = 12) had suppressed cortisol levels above the 97.5th percentile (1.96 microg/dl) of the healthy controls. Subgroup analysis regarding the influence of gonadal steroid hormone replacement in both groups and gender in the osteoporotic group did not change the results. Four of the 12 patients with incomplete suppressive cortisol underwent adrenal endosonography, unilateral adrenal nodular hyperplasia being detected in three cases. In two patients the diagnosis was confirmed by histology and normalisation of a dexamethasone suppression test following endoscopic adrenalectomy. These data yield evidence for a difference in the regulation of cortisol secretion following high-dose dexamethasone administration between healthy subjects and a subgroup of patients with primary osteoporosis. This might be due to a relevant amount of autonomous cortisol secretion in some of these patients; however, even cortisol resistance has to be taken into account. PMID- 11254237 TI - Osteoprotegerin and osteoprotegerin-ligand balance: a new paradigm in bone metabolism providing new therapeutic targets. AB - Osteoprotegerin-ligand, also called Osteoclast Differentiation Factor or RANK ligand, its receptor RANK and its decoy receptor Osteoprotegerin are key molecules regulating osteoclast differentiation and activation. In this view we discuss structure and expression of these molecules, the genetic models addressing their function and their role in in vivo models of osteoclast differentiation and activation. The new paradigm that has evolved from these studies, is not only important in normal bone homeostasis but also appears to play a role in different diseases that affect the skeleton, such as osteoporosis, inflammatory joint disease and cancer. It has opened a new era in bone research by increasing our molecular knowledge and providing new therapeutic targets in bone disease. PMID- 11254238 TI - Cytokine levels in serum of patients with juvenile rheumatoid arthritis. AB - We investigated serum levels of interleukin (IL)-1beta, IL-6, IL-8, IL-12 and tumour necrosis factor (TNF)-alpha in JRA patients during both active and inactive phases of the disease. The systemic JRA patients had the highest IL 1beta and IL-6 levels during both active and inactive periods. In the systemic group IL-1beta, IL-6 and IL-12 levels during the active period were elevated compared to the inactive period (p = 0.0173, p = 0.0359 and p = 0.0117, respectively). Levels of these cytokines during the inactive stage were still greater than those of controls. IL-8 and TNF-alpha levels during both active and inactive periods were comparable to controls. IL-1beta correlated strongly with CRP and ESR (p = 0.008 and p = 0.031, respectively). IL-6 correlated significantly with CRP (p = 0.002). IL-12 levels were found to be correlated with ESR and CRP (p = 0.03 and p = 0.04, respectively). In active polyarticular JRA patients, IL-6 levels were elevated compared to the inactive phase, and the control (p = 0.001) IL-12 levels decreased significantly with clinical remission (p = 0.018). There was a strong correlation between 11-12 levels and number of joint with limited motion (p = 0). In oligoarticular JRA patients, IL-12 levels during active period were greater than in the controls and there was a marked decrease in IL-12 levels when the patients entered the inactive phase (p = 0.001) In conclusion, IL-1beta, IL-6 and IL-12 may play an important role in JRA and may be used as a marker of disease activity. PMID- 11254239 TI - Subsets in psoriatic arthritis formed by cluster analysis. AB - The aim of the study was to create subgroups among psoriatic arthritis patients on the basis of dermatological features, clinical pattern of arthritis, and laboratory, immunological and radiological findings. Data on 100 patients were expressed in a standardised form and entered into hierarchical cluster analysis according to Ward's method. Seven subgroups were created. Fifty-six patients with mild psoriasis were sorted into a 'polyarticular group'. Two 'RA-like groups' were formed, differing from each other serologically and in axial involvement. In an 'oligoarticular group' (18 patients) serious skin disease and female gender predominancy were found to be characteristic. Eight patients with polyarticular arthritis were assigned to an 'erythrodermal group', in which polyarticular arthritis, mutilating, severe arthritis and a history of erythroderma were characteristic. Close to this group on the dendrogram eight women were sorted into a 'distal form'. Sausage fingers were frequent, and nail dystrophy was present in every case. In a 'pustular group' (three patients) the different type of skin involvement was considered and nail dystrophy was common. In the newly created subgroups not only the arthritic status, but also the type of the skin disease, played a determining role. PMID- 11254240 TI - Hypothalamus-hypophysis-thyroid axis, triiodothyronine and antithyroid antibodies in patients with primary and secondary Sjogren's syndrome. AB - It has been well established that, anti-thyroglobulin antibodies (ATG) and anti microsomal antibodies (AMC) may be present in various thyroid disorders and other systemic autoimmune diseases, including Sjogren's syndrome (SS). However, presence of circulating autoantibodies to thyroid hormones, i.e. both to triiodothyronine (T3) and tetraiodothyronine (T4), has not been studied extensively in SS. Autoantibodies to T3 and T4 are very important, because serum T3 and T4 levels may be detected spuriously higher or lower, due to the presence of these autoantibodies. Their presence should be suspected when measured serum thyroid hormone levels are not consistent with clinical status of the patient. SS is a slowly progressive, inflammatory autoimmune disease, affecting primarily the exocrine glands. Thyroid gland, being a target in some autoimmune diseases, is well known to be affected in SS as well. Keeping this possibility in mind, we investigated T3 autoantibody levels and thyroid gland involvement in patients with SS. Twenty-six SS patients (F/M:22/4) with a mean age of 46.6 years, were recruited in this study. Twelve of them were accepted as primary SS (pSS), while others had secondary SS (sSS) (7 with rheumatoid arthritis (RA), 3 with systemic lupus erythematosus (SLE), 3 with progressive systemic sclerosis (PSS) and 1 with sarcoidosis). Thyroid function tests, including T3, T4, fT3, fT4, TSH, ATG, AMC, T3 antibody measurements, thyroid scintigraphy, thyroid ultrasonography and TRH stimulation tests were performed in all patients. We compared our results with those of the twenty healthy normal controls. Serum ATG and/or AMC were detected in three patients with pSS (25%) and no patients with sSS. No significant difference could be shown in the other parameters, including T3 autoantibodies and thyroid function tests. TRH stimulation test was also normal, showing that the hypothalamus-hypophysis-thyroid axis was not affected in patients both with pSS and sSS. In conclusion, we found that T3 autoantibody levels in pSS, were not significantly higher than sSS and normal controls. PMID- 11254241 TI - Pneumatosis cystoides intestinalis in a patient with polymyositis. AB - A 29-year-old Japanese woman with polymyositis (PM) developed pneumatosis cystoides intestinalis (PCI). The patient experienced a gradual onset of mild epigastric abdominal pain and abdominal distension. Radiographs revealed pneumatosis intestinalis involving the small bowel, and colonoscopy revealed submucosal cysts. Treatment with antibiotics and high-flow oxygen resulted in remission. This is an extremely rare case of PCI in a patient with PM. PMID- 11254243 TI - Glycogenosis type V (McArdle's disease) mimicking atypical myositis. AB - A 13-year-old girl was referred to our clinic because of a positive rheumatoid factor test, muscle pain and weakness. Laboratory evaluation revealed an increased ESR, hypergammaglobulinaemia, antinuclear antibodies, circulating immune complexes, complement consumption and elevated serum creatine kinase (CK) activity. A needle biopsy of the dolent muscle showed normal routine histology. Immunohistochemistry disclosed single lymphocytes and a weak myocytic HLA class I expression. The diagnosis of myositis was considered and corticosteroids were initiated, leading to an increase of complement levels and a decrease of CK activity and ESR. She subjectively felt stronger but still reported exercise intolerance and metabolic myopathy was considered. Myophosphorylase activity was completely lacking, establishing the diagnosis of McArdle's disease. CK level was found to be elevated in an obese 4-year-old brother too, who refused extensive walking but reported no muscle pain. Myophosphorylase deficiency was demonstrated by histochemistry and by biochemical analysis of his muscle. The female case illustrates that in children with the clinical picture of inflammatory myopathy and serological but not clinical response to therapy underlying metabolic muscle disorders should be excluded. Since the pathogenesis of polymyositis remains unclear, we speculate that inflammatory changes observed in the muscles may have been initiated by muscular damage resulting from the underlying metabolic disease. The serological changes remained unexplained and may contribute to a so far undeterminable connective tissue disease. PMID- 11254242 TI - Bilateral femoral osteomyelitis with knee arthritis due to Salmonella enteritidis in a patient with systemic lupus erythematosus. AB - A bilateral knee septic arthritis due to Salmonella enteritidis developed in a female patient affected by long-standing systemic lupus erythematosus (SLE) with cardiac and renal involvement treated with immunosuppressants and corticosteroids. Because avascular necrosis and multiple osteomyelitic areas were detected at the same time in both right and left femoral condyles, an early localisation of Salmonella into the bone was assumed. Involvement of the joints was regarded as consequence of local dissemination of infection. Ampicilline (0.2 g/kg body weight daily for 2 months) plus ciprofloxacin (1.5 g daily for 12 months) and withdrawal of immunosuppressants appeared to be effective in preventing complications of infection. PMID- 11254244 TI - An unusual presentation of Behcet's disease: intestinal perforation. AB - Behcet's disease (BD), when first described in 1937, consisted of three symptoms: recurrent oral and genital ulcerations and iridocyclitis. Today, it is known that BD is a multisystemic chronic vasculitic disorder which may involve both arteries and veins of all sizes, as well as the central nervous and gastrointestinal systems. The rate of gastrointestinal involvement of BD varies in different populations, being more common in Japan (50%-60%) and less common in the Mediterranean basin, including Turkey (0%-5%). We present a 34-year-old Turkish woman with BD who had ileal and colonic ulcerations complicated by perforation and gastrointestinal bleeding. Special emphasis was placed on the differential diagnosis between Crohn's disease (CD) and BD with gastrointestinal involvement. PMID- 11254246 TI - Minocycline-associated lupus-like syndrome with ulnar neuropathy and antiphospholipid antibody. AB - Peripheral neuropathy in association with minocycline-induced lupus-like reaction has not previously been reported. We present a case of probable minocycline induced lupus associated with antiphospholipid antibodies and an ulnar neuropathy which has slowly improved since the discontinuation of minocycline. PMID- 11254245 TI - Induction of remission with intravenous immunoglobulin and cyclophosphamide in steroid-resistant Evans' syndrome associated with dermatomyositis. AB - Evans' syndrome is characterised by the simultaneous or sequential occurrence of Coombs'-positive haemolytic anaemia (AIHA) and immune thrombocytopenia without underlying aetiology. It has been found to be associated with collagen vascular diseases, especially systemic lupus erythematosus (SLE) and scleroderma. However, Evans' syndrome with dermatomyositis is very rare. A 59-year-old woman, who had been taking high-dose prednisolone for a month and cyclosporin for 10 days for dermatomyositis, developed purpura on the left popliteal fossa. The platelet and haemoglobin levels decreased to 77,000/mm3 and 9.8 g/dl, respectively. Antiplatelet antibody was positive. Thrombocytopenia responded to intravenous immunoglobulin (IVIG) for a short time, but further decreased in a week. Her blood film showed features of haemolytic anaemia. Laboratory findings showed reticulocytosis and a positive direct Coombs' test. Bone marrow examination showed a mild hyperplasia of erythroid precursors and megakaryocytes. The patient was successfully treated with cyclophosphamide in addition to oral prednisolone. AIHA in connective tissue disease may develop gradually and show a benign clinical course in most patients. Therefore, we suggest that patients with dermatomyositis and anaemia should always be checked for haemolysis if there is no other explanation. PMID- 11254247 TI - Clinical manifestation of POEMS syndrome with features of connective tissue disease. AB - The POEMS (polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, skin changes) syndrome is a rare plasma cell disease with multiorgan involvement and varying clinical manifestations. We report a 38-year-old man who presented with scleroderma-like skin changes of the hands and feet, sicca and Raynaud's syndrome, pleural effusions, glomerulopathy, polyneuropathy, hepatosplenomegaly and lymphadenopathy. Steroid treatment was started on the assumption of a connective tissue disease and led to a temporary improvement. During the further course of the disease, hypothyreosis, monoclonal gammopathy and osteosclerotic bone lesions were detected, leading to the diagnosis of POEMS syndrome. This case emphasises the need to consider POEMS syndrome as a differential diagnosis in patients with signs of connective tissue disease and polyneuropathy. PMID- 11254248 TI - Pancytopenia and colitis with Clostridium difficile in a rheumatoid arthritis patient taking methotrexate, antibiotics and non-steroidal anti-inflammatory drugs. AB - Methotrexate (MTX) is widely used despite its side-effects. We describe a rheumatoid arthritis (RA) patient taking low-dose MTX who developed severe pancytopenia and colitis with Clostridium difficile after the administration of antibiotics for acute pyelonephritis. Our case suggests that low-dose MTX may seriously interact with antibiotics and that these side-effects should always be considered when RA patients are treated with MTX and antibiotics. PMID- 11254250 TI - Methotrexate-treated arthritis and lymphoproliferative disease--coincidence only? AB - This paper describes four patients with inflammatory joint diseases treated with methotrexate who developed lymphoma either of unusual type or with bizarre clinical features. The pathogenesis of this unusual condition is discussed and the difficulty in differentiating it from the features of the initial disease is emphasised. PMID- 11254249 TI - Distal extremity swelling with pitting oedema in rheumatoid arthritis. AB - Distal extremity swelling with pitting oedema due to altered lymphatic drainage has been reported in some patients with rheumatoid arthritis (RA). The resistant to-therapy oedema usually affected the upper limbs in an asymmetrical pattern. Until now, extensor tenosynovial involvement has not been described in RA patients suffering from distal extremity swelling with pitting oedema. Three patients are described: two of them had predominant extensor tenosynovial involvement in their hands, with impaired lymphatic drainage demonstrated by (MRI) and lymphoscintigraphy, respectively. In both cases the oedema was chronic and not responsive to treatment. One patient had extensor tenosynovial involvement without impaired lymphatic drainage. In this case, the oedema remitted completely after a few days of corticosteroid therapy. None of them showed differences in serum levels of vascular endothelial growth factor (VEGF), whether they were RA patients with no pitting oedema or healthy volunteers. PMID- 11254251 TI - Dyslexia and visual-spatial talents: compensation vs deficit model. AB - There are both theoretical and empirical reasons to support the hypothesis that dyslexia is associated with enhancement of right-hemisphere, visual-spatial skills. However, the neurological evidence is neutral with respect to whether dyslexic visual-spatial abilities should be superior (a compensation model) or inferior (a deficit model). In three studies we tested the hypothesis that dyslexia is associated with superior visual-spatial skills. Individuals with dyslexia not only failed to show superiority on a range of visual-spatial tasks, even when tasks were presented without time constraints, but also demonstrated a deficit on many tasks. Whereas we found attentional problems associated with dyslexia, these did not explain our findings. Results are discussed in terms of the apparent conflict between the failure to find any visual-spatial talent associated with dyslexia and the fact that dyslexia is overrepresented in certain visual-spatial professions. PMID- 11254252 TI - Performance in picture naming and word comprehension: evidence for common neuronal substrates from intraoperative language mapping. AB - Correlations of naming ability and performance in the Token Test are known from aphasia; however, the mechanisms underlying these correlations are unclear. Naming tasks are commonly used in intraoperative mapping for identification of cortical areas involved in language processing. In the present study, we measured performance in an elementary Token Test task, i.e., single word comprehension, during electrical stimulation of cortical sites at which this stimulation previously had disturbed the naming process. It was found that at about half of the sites at which naming could be disturbed electrical stimulation also led to disturbances in Token Test performance, indicating that there are common neural structures critical for performance in both tasks. These findings are discussed in terms of a multilayered semantic network in which the level of the simultaneous binding of features into concepts and the level of units that are organized with respect to semantic relations may be disturbed separately. PMID- 11254253 TI - Disorder in sequential speech perception: a case study on pure word deafness. AB - We described disorders of a patient which were uniquely restricted to speech perception of syllable sequences after brain damage. The results of series of experiments using syllable sequences showed "negative recency effect," in which the subject's repetition performance at the latter syllable position was remarkably poor. Experimental analyses suggested that the "negative recency effect" could be due to dual factors: the lower rate of processing of speech sounds and the memory load of holding processes of preceding syllables imposed on the succeeding phonological processing. The results also suggested that the holding processes which imposed the memory load on the succeeding auditory phonological coding processing were modality nonspecific. PMID- 11254254 TI - Semantic priming in Alzheimer's dementia: evidence for dissociation of automatic and attentional processes. AB - The nature of the semantic memory deficit in dementia of the Alzheimer's type (DAT) was investigated in a semantic priming task which was designed to assess both automatic and attention-induced priming effects. Ten DAT patients and 10 age matched control subjects completed a word naming semantic priming task in which both relatedness proportion (RP) and stimulus-onset asynchrony (SOA) were varied. A clear dissociation between automatic and attentional priming effects in both groups was demonstrated; however, the DAT subjects' pattern of priming deviated significantly from that of the normal controls. The DAT patients failed to produce any priming under conditions which encouraged automatic semantic processing and produced facilitation only when the RP was high. In addition, the DAT group produced hyperpriming, with significantly larger facilitation effects than the control group. These results suggest an impairment of automatic spreading activation in DAT and have implications for theories of semantic memory impairment in DAT as well as models of normal priming. PMID- 11254255 TI - Anomalous cerebral language organization: acquired crossed aphasia in a dextral child. AB - Following a dramatic change of its reported incidence, it was only recently recognized that acquired crossed aphasia in dextral children represents a highly exceptional phenomenon. We describe in a three epoch time-frame model the aphasic and neurocognitive manifestations of an additional case and focus briefly on its anatomoclinical configurations. In our patient, a right parietal cortico subcortical hemorrhagic lesion caused an initially severe aphasia. After remission of the global aphasic symptoms in the acute phase, an adynamic output disorder with relatively severe auditory-verbal comprehension disturbances developed. In addition to the adynamia of self-generated speech, formal language investigations performed 3 weeks postonset, revealed agrammatism, hypertonic dysarthria, and dysprosodia. A substantial improvement of the aphasic disorder was objectified 83 days postonset. Neuropsychological investigations disclosed both dominant and nondominant hemisphere dysfunctions. Reassessment of neurocognitive functions after a 10-year period evidenced discrete residual anomia, confined to visual confrontational naming and a discrete visuo-perceptual syndrome. Given the posterior localization of the lesion, the syndrome shift from global to predominantly adynamic aphasia represents a finding beyond the plausible anatomoclinical expectations holding in general for the uncrossed, classic types of childhood and adult aphasia. As the first representative of crossed aphasia in dextral children with an anomalous lesion-aphasia profile, our case provides evidence to enrich the discussion on lateralization and intrahemispherical organization of language functions in both childhood and adult aphasia. PMID- 11254256 TI - Agrammatic Broca's aphasia is not associated with a single pattern of comprehension performance. AB - One influential hypothesis posits that the brain regions implicated in Broca's aphasia are responsible for specific syntactic operations that are necessary for the comprehension and production of sentences (Grodzinsky, 1986, 1990, in press). The empirical basis of this hypothesis is the claim that Broca's aphasics have no difficulty understanding sentences in the active voice (and other "canonical" sentence types, such as subject relatives and clefts with negative predicates), but perform at chance level with passive voice constructions (and other "noncanonical" sentences such as object-gap relatives and object clefts). In the face of well-established results indicating that Broca's aphasics can exhibit several different performance patterns on these sentence types, Grodzinsky, Pinango, Zurif, and Drai (1999) argued that these conflicting results do not challenge the theory when the data are analyzed appropriately. They carried out a creative statistical analysis of the comprehension performance of published cases of Broca's aphasia and concluded that all of these cases are in agreement with the predicted pattern: chance on passives and 100% correct on actives. Here we show that the statistical reasoning adopted by Grodzinsky et al. (1999) is flawed. We also show that the comprehension performance of a substantial number of the Broca's aphasics in their own sample does not conform to the pattern required. Rather, contrary to these authors' claim, Broca's aphasia is not associated with a consistent pattern of sentence comprehension performance, but allows for a number of distinct patterns in different patients. PMID- 11254257 TI - Broca's aphasia is associated with a single pattern of comprehension performance: a reply. PMID- 11254258 TI - The measurement of chance performance in aphasia, with specific reference to the comprehension of semantically reversible passive sentences: a note on issues raised by Caramazza, Capitani, Rey, and Berndt (2001) and Drai, Grodzinsky, and Zurif (2001). PMID- 11254259 TI - Sometimes a noun is just a noun: comments on Bird, Howard, and Franklin (2000). AB - Bird, Howard, and Franklin (2000) have proposed a semantic-conceptual explanation of grammatical category-specific deficits that attributes impairments in noun and verb processing to two distinct mechanisms. According to their account, apparent deficits in verb production are not category specific, but rather result from the lower imageability of verbs compared to concrete nouns. Noun deficits are said to result from differences in the distribution of semantic feature types such that damage to sensory features results in disproportionate impairments in naming nouns, especially animate nouns, compared to verbs. However, this hypothesis, which we call the "extended sensory/functional theory" (ESFT), fails on several counts. First, the assumption that representations of living things are more heavily freighted with sensory features than are those of nonliving objects does not have any reliable empirical basis. Second, the ESFT incorrectly predicts associations between deficits in processing sensory features and living things or functional features and nonliving things. Finally, there are numerous cases of patients with grammatical category-specific deficits that do not seem to be consistent with damage at the semantic level. All of this suggests that the ESFT is not a useful model for considering grammatical (or semantic) category-specific deficits. PMID- 11254260 TI - Noun-verb differences? a question of semantics: a response to shapiro and caramazza. AB - We presented a new model using the sensory/functional theory of semantic category deficits to explain noun/verb deficits in aphasia. The predictions arising from this model were tested on a small number of patients exhibiting grammatical and/or semantic category specific deficits in picture naming. The results lent support for the theory presented. Shapiro and Caramazza (this issue) raised several objections to this theory (which they call the "extended sensory functional theory," or ESFT). In this article we address their concerns about the validity of the ESFT and conclude that it is indeed a useful model that provides a parsimonious explanation for many diverse patterns of deficits. PMID- 11254262 TI - Effects of Alkylamines on the Percolation Phenomena in Water/AOT/Isooctane Microemulsions. AB - We carried out an investigation on the influence of several alkylamines, frequently present in reactions carried out in microemulsions, on the properties of the water/AOT/isooctane system. The presence of alkylamines has an important effect on the electrical percolation phenomena. This effect of amines on the electrical percolation of microemulsions of AOT/isooctane/water can be explained by taking into account the ability of these substrates to associate with the AOT film in the microemulsion, the basicity of the amine, and the different solubility of the amine in the three pseudophases of the system. Copyright 2000 Academic Press. PMID- 11254263 TI - Potentiometric and Spectroscopic Characterization of Copolypeptide-Surfactant Complexes Formed by a Cooperative Binding System. AB - The binding behavior of an anionic surfactant, sodium dodecyl sulfate (SDS), to a series of L-lysine-containing copolypeptides in aqueous solutions was investigated in relation to the conformational change of copolypeptide-surfactant complexes with the use of potentiometric and spectroscopic techniques. The present results of CD spectra and the binding isotherm of SDS by copolypeptides of opposite charges can lead us to conclude that SDS binds cooperatively to the positively charged side groups of a series of copolypeptides used in this work, resulting in the formation of a micelle-like cluster due to an additional hydrophobic interaction among bound SDS ions. Solid-state properties of the stoichiometric copolypeptide-SDS complexes were also examined by using CD and FT IR spectroscopies; (Lys, Tyr) (1:1) and (4:1) systems adopt a beta-pleated sheet conformation, while (Lys, Trp) (4:1) and (Lys, Phe) (1:1) systems adopt an alpha helical conformation. Based on the results of FT-IR spectra, in all cases surfactant alkyl chains of SDS in the solid complexes were in an extended conformation. Copyright 2000 Academic Press. PMID- 11254264 TI - Flocs in Shear and Strain Flows. AB - Preflocculated ferric hydroxide flocs were subjected to either a simple shear flow or a two-dimensional straining flow, and their motion was optically observed. Digital image analysis was applied to extract information on orientation and deformation from the digitized frames. It was found that the simple shear flow led to a rotation of the flocs whose motion can be understood from the behavior of a solid ellipsoid. In the extensional flow, no continuous rotation occurred and flocs were broken apart along the axis of straining. The rupture forces estimated from an ellipsoid model were found to be in the range of 0.1 N/m(2). Copyright 2000 Academic Press. PMID- 11254265 TI - Structures of Fibrous Supramolecular Assemblies Constructed by Amino Acid Surfactants: Investigation by AFM, SANS, and SAXS. AB - Aqueous gel-like solutions of N-acyl-L-aspartic acids (C(n)Asp, n=14, 16, 18) and N-dodecanoyl-beta-alanine (C(12)Ala) were prepared at pH 5-6 at room temperature. Structures of supramolecular assemblies in the solutions were investigated by atomic force microscopy (AFM), small-angle neutron scattering (SANS), and small angle X-ray scattering (SAXS). The cross-sectional radii, 22-30 A, of helical, fibrous assemblies were obtained from analysis of SANS for 1% gel-like C(n)Asp solutions. Three Bragg spacings were observed in a SANS spectrum for a 6% C(16)Asp solution. C(n)Asp molecules are associated into the unit chain of a helical bilayer strand with a diameter of 50-60 A. Unit chains where linear bilayers twist form a double strand with helical sense of approximately 650-A pitch. It was confirmed from AFM images that cylindrical fibers in a gel-like C(12)Ala solution had a circular cross-section. The SAXS spectrum showed characteristic Bragg spacings. Cylindrical C(12)Ala fibers consist of multilamellar layers of period approximately 34-A. The fibers are laterally organized with period 365-380 A. Copyright 2000 Academic Press. PMID- 11254266 TI - Nitrogen Sorption Studies of Silica Particles Obtained in Emulsion and Microemulsion Media. AB - The internal surface structures of silica aerogel particles synthesized using different catalysts in emulsion and microemulsion media have been investigated by means of N(2) adsorption and desorption isotherms. Surface fractal dimensions have been computed using different methods: Frankel-Halsey-Hill plots of the adsorption isotherms, the thermodynamic fractal isotherm equation, and a modification of the thermodynamic fractal isotherm equation. Silica aerogels synthesized in emulsion media with an acidic catalyst have a high specific surface area without micropores and show two separate ranges of scales where the surface fractal dimensions are different and constant. Silica aerogels synthesized in emulsion media with a basic catalyst have a moderate specific surface area with a high percentage of micropores and show constant surface fractal dimensions over a larger range. Silica aerogels synthesized in microemulsion media with a basic catalyst have a low specific surface area with a moderate percentage of micropores and show a moderate range of scales over which the surface fractal dimension is constant. Analyses by both the thermodynamic and modified thermodynamic methods give similar ranges of the surface fractal dimensions of the silica particles. Copyright 2000 Academic Press. PMID- 11254267 TI - Sorptive Characteristics of Acridine-Surfactant-Phyllosilicate Systems. AB - The effects of the presence of a cationic surfactant on the sorption of two typical nitrogen heterocyclic compounds (NHCs) on swelling and nonswelling phyllosilicates were investigated. The addition of a cationic surfactant to a NHC phyllosilicate system can either solubilize the sorbed NHC molecules or immobilize the suspended NHC molecules, depending upon the concentration and sequence of the surfactant added. The presence of surfactant molecules on the clay mineral surfaces promoted sorption of more neutral NHC molecules due to hydrophobic effect, and this resulted in a considerable increase in sorption at high pH conditions. Both surfactant and NHC molecules were sorbed not only on the external surfaces of the phyllosilicates but also on the interlayer spaces of swelling-type clay minerals, and the resulting change in basal spacings indicates the competition between the two groups of molecules in the interlayer space. Copyright 2000 Academic Press. PMID- 11254268 TI - Heteroflocculation of Amidine Polystyrene Latex and Anticarsia gemmatalis Nucleopolyhedrovirus as a Model System for Studying Sunlight Protection. AB - Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) is a baculovirus specific for the control of an important soybean defoliator. The baculovirus is comprised of double-stranded DNA, occluded in a proteinaceous structure called a polyhedron. Ultraviolet sunlight is the most destructive factor that affects the persistence of the virus in the field. In the present study, we use a model system wherein the pathogen is covered by another particle of opposite charge in order to test the effectiveness of a physical barrier as a protection against sunlight. Heteroflocculation experiments were carried out using two different age batches of AgMNPV and amidine polystyrene latex particles. The assessment of heteroflocculation was achieved by zeta potential and adsorption isotherm measurements, and by scanning electron microscopy. Despite the great difference in potentials between latex particles and the baculovirus, low-affinity isotherms were obtained in both pure water and 0.1 mM KCl. Adsorbed latex particles were easily washed out from the polyhedron surface. This low affinity could be attributed to the presence of a strongly repulsive hydration force of short range operating on the system. The results suggest that the failure to obtain a good physical barrier against sunlight might be attributed to the difficulty in keeping the polyhedron surface covered. Copyright 2000 Academic Press. PMID- 11254269 TI - Experimental Study on Contact Angle Patterns: Liquid Surface Tensions Less Than Solid Surface Tensions. AB - The interpretation of contact angles in terms of solid surface tensions is not trivial. In the past, we and others have postulated that contact angles should be measured with liquid of surface tension larger than the anticipated solid surface tension, i.e., gamma(lv)>gamma(sv). This has recently been disputed. It is also not entirely obvious how to proceed experimentally since gamma(sv) is not known initially. Typically, one starts with a liquid of high gamma(lv) (such as water) and goes lower. We have stopped in the past when the contact angles became small. A question arises as to what would happen if we would go on. Contact angles of liquids with gamma(lv) less than or near gamma(sv) were measured on eight polymer coated solid surfaces. The experimental contact angle patterns for gamma(lv)gamma(sv) were compared. Results suggest that contact angle interpretation in terms of solid surface tensions requires contact angles to be measured for gamma(lv)>gamma(sv) because the Young equation is not applicable for gamma(lv)/=350-nm radius for an ionic strength of 0.01 M). Under these conditions, our calculations predict a transient period of fast aggregation into the secondary minimum followed by slow primary aggregation. The aggregation in this second regime is found to take place at a lower rate than what would be expected in the absence of the secondary minimum or from an earlier linearized model for secondary aggregation. The crossover time between the two regimes strongly depends on the particle size but not on the particle concentration, which however determines the degree of aggregation reached within the fast regime. We also conclude that a previously observed severe discrepancy between measured and predicted aggregation rate constants for submicron particles is not due to the neglect of secondary aggregation in the theoretical treatment. Copyright 2000 Academic Press. PMID- 11254287 TI - Inorganic Ligand Effects on Pb(II) Sorption to Goethite (alpha-FeOOH). AB - Macroscopic measurements show that Pb(II) uptake on iron-(hydr)oxides can be altered significantly by dissolved carbonate (enhanced up to 18% at pH 5 and decreased above pH approximately 6.5 in analyses at 1 atm CO(2)). This study elucidates the molecular-scale processes giving rise to these macroscopic effects by characterizing the structures of Pb(II) sorption complexes formed on goethite (alpha-FeOOH) in the presence of carbonate using in situ Pb L(III)-EXAFS and ATR FTIR spectroscopies. Bond valence and structural constraints are applied to develop mineral surface site-specific models for Pb sorption. Under all conditions studied (pH 5-7, Gamma(Pb)=0.4-4umol/m(2), and P(CO(2))=0-1 atm), Pb(II) forms predominantly inner-sphere edge-sharing (bidentate and/or tridentate) complexes with Fe(O,OH)(6) octahedra (R(Pb-Fe) approximately 3.3 A). Corner-sharing complexes (R(Pb-Fe) approximately 3.9 A) are observed only in low pH (5) samples (P(CO(2)) 0-1 atm). Consistent with this pH sensitivity, site specific analyses suggest that the relative abundance of corner-sharing sites reflects changes in the proton affinity of triply coordinated sites on the goethite (110) surface as suggested previously. FTIR results suggest the existence of ternary surface complexes in which carbonate groups bond to Pb as monodentate ligands. EXAFS data indicate that these ternary complexes are bound to the surface through Pb, forming metal-bridged (Type A) complexes. Findings are summarized as structural models and corresponding mineral surface site-specific chemical reactions. Copyright 2000 Academic Press. PMID- 11254288 TI - Inorganic Ligand Effects on Pb(II) Sorption to Goethite (alpha-FeOOH). AB - The effects of sulfate anions on the uptake of Pb(II) onto goethite were investigated at the molecular level using in situ Pb L(III)-EXAFS and ATR-FTIR spectroscopies. Macroscopic uptake data show that Pb uptake can be enhanced by at least 30% at pH 5 in the presence of 3.16 mM sulfate and that sulfate uptake at pH 7 can be enhanced by more than a factor of 3 in the presence of 1.0 mM Pb. Consistent with behavior in sulfate-free systems, Pb(II) forms inner-sphere complexes sharing either corners or edges with Fe(O,OH)(6) octahedra under all conditions studied. The relative fraction of corner-sharing complexes is, however, significantly enhanced in the presence of sulfate at pH 5, 6, and 7 (all conditions studied) and additional sulfate species with C(3v) or lower point symmetry were noted in the presence of Pb by ATR-FTIR. Drawing on bond valence and structural constraints developed in J. D. Ostergren et al. (2000, J. Colloid Interface Science 224, 000-000), these results indicate formation of Type A ternary complexes bonded to the surface through Pb that is bound as a bridging bidenate complex to two adjacent A-type (singly coordinated) surface oxygens (( identical withFe-O)(2)-Pb-OSO(3)). Copyright 2000 Academic Press. PMID- 11254289 TI - Quantitative Determination of Asphaltenes and Resins in Solution by Means of Near Infrared Spectroscopy. Correlations to Emulsion Stability. AB - Near-infrared (NIR) spectroscopy in the range 1100-2250 nm together with a latent variable regression technique is used to analyze the content of asphaltene and resins in solution. It is shown that this technique is capable of determining the amount of these components individually. w/o emulsions were prepared from the separated components of asphaltenes and resins from crude oils. The stability was directly determined with the critical voltage in a dielectric instrumentation. The emulsion stability decreased linearly with an increase in the resin/asphaltene ratio. A final linear model correlating the critical voltage and the analytical concentrations (from the NIR spectra) could be established for this model system. Copyright 2000 Academic Press. PMID- 11254290 TI - Hydrophobically Associating Alginate Derivatives: Surface Tension Properties of Their Mixed Aqueous Solutions with Oppositely Charged Surfactants. AB - The comparative study of the interfacial properties of an anionic polysaccharide, sodium alginate (Alg), and its hydrophobically modified derivative (Alg-C(12)), covalently substituted by dodecyl chains (12% mol/mol saccharide unit), was carried out in the absence or in the presence of an oppositely charged surfactant, dodecyltrimethylammonium bromide (DTAB). The drastically different behaviors which were observed are interpreted in terms of the arrangement and mobility of the hydrophobic long alkyl chains, depending on the nature of their fixation, covalent or ionic, on the polysaccharide backbone. Copyright 2000 Academic Press. PMID- 11254291 TI - Cation Exchange and Sorption Properties of TIN(IV) Phosphate. AB - The ion exchange and sorption properties of crystalline Tin bis(monohydrogen orthophosphate) monohydrate of composition Sn(HPO(4))(2).H(2)O were studied in an aqueous solution of KCl over the temperature range 300-320 K, varying the pH and metal ion concentration in the solution. The data were explained on the basis of the law of chemical equilibrium. The metal ion sorption data were fitted to the Langmuir adsorption equation to evaluate the Langmuir parameters. The extent of adsorption was found to increase with an increase in temperature and metal ion concentration in the selectivity order Cu(2+)>Co(2+)>Ni(2+). The Langmuir parameters were used to calculate the thermodynamic functions like standard entropy, enthalpy, and free energy changes during the process of sorption. Copyright 2000 Academic Press. PMID- 11254292 TI - Effect of Temperature on Micelle Formation in Aqueous NaBr Solutions of Octyltrimethylammonium Bromide. AB - Association processes in aqueous solutions of octyltrimethylammonium bromide, C(8)TAB, have been studied in aqueous NaBr solutions at temperatures from 20 to 55 degrees C. The values of the critical micelle concentration, CMC, were determined from the intersections of two straight line portions of the plots of the relationship between adiabatic compressibility of the solutions and surfactant concentration. The value of the CMC thus determined exhibits minima at a certain temperature, T(min). The value of T(min) shifts toward a lower temperature with increasing NaBr concentration. Based on the most probable micelle size model, the values of thermodynamic functions of micelle formation have been estimated at various temperatures. Copyright 2001 Academic Press. PMID- 11254293 TI - Hyperbolic Spirals as Surface Structures in Thin Layers. AB - When thin layers of 4-chloro-3-methylphenol and a copolymer of methyl(methacrylate) and maleic acid dissolved in acetone are dried by solvent evaporation, various surface structures appear. Besides linear surface deformations that can ramify like fractals, spirals of the hyperbolic type have been found. The surface structures are due to crystallization processes and flows caused by surface tension differences. The spirals are surface elevations with grooves on both sides as shown by surface profile measurements by means of a microscope interferometer. The addition of surfactants reduces the structure formation. A large surfactant concentration prevents the structure formation. Copyright 2001 Academic Press. PMID- 11254294 TI - Dispersed Molecular Aggregates. AB - Colloidal dispersions of tungstic acid (H(2)WO(4)) have been prepared in water/(TX-100+alkanol)/n-heptane water-in-oil microemulsion media by reacting Na(2)WO(4) with HCl. The effects of alkanol chain length, TX-100/alkanol mass ratio, temperature, and dilution at different [water]/[TX-100] mole ratios (omega) have been studied by the dynamic light scattering technique. The formation of H(2)WO(4) in the microwater pool has been established by FT-IR measurements. The particle sizes and shapes in microemulsion media and in isolated states have been measured by TEM and SEM techniques. The enthalpy of formation of H(2)WO(4) in the water pool of the microemulsions has also been determined microcalorimetrically. Copyright 2001 Academic Press. PMID- 11254295 TI - Physicochemical Investigations on the Interaction of Surfactants and Salts with Polyvinylpyrrolidone in Aqueous Medium. AB - Microcalorimetric investigations have been carried out onthe interaction of the surfactants sodium cholate, sodium deoxycholate, tetradecyltrimethylammonium bromide, cetyl(hexadecyl)trimethylammonium bromide, and p-tert-octylphenoxy polyoxy-ethylene ether (Triton X-100) and the salts potassium iodide, sodium benzoate, sodium bromide, and sodium salicylate with the neutral polymer polyvinylpyrrolidone (PVP). The enthalpy of dilution of the surfactants has been measured in the absence and presence of the polymer and the results are compared to determine the effect of PVP on the micellization of the surfactants and the energetics of the process. As well, the micellization activity of the surfactants in the presence of the polymer has been studied by conductometric and fluorimetric methods. The enthalpy of dilution of the salts has been measured to provide an understanding of the nature and magnitude of their interaction with PVP. Copyright 2001 Academic Press. PMID- 11254296 TI - In Situ Immobilization of Ultrafine Particles Synthesized in a Water/Oil Microemulsion. AB - We investigated the in situ immobilization of ultrafine particles synthesized in a water/oil (w/o) microemulsion to silica for its possible application to supported metal catalysts. ZnS particles immobilized to silica by the ME method were consistent with those synthesized in a w/o microemulsion. Therefore, ZnS particles in a w/o microemulsion could be immobilized to silica without aggregation by this method. The relationship between the method of synthesizing Rh ultrafine particles in a w/o microemulsion and the diameter and diameter distribution of Rh particles immobilized to silica was studied. Rh-SiO(2) catalysts with a sharp diameter distribution could be prepared by immobilizing Rh hydrazine complex particles because these complex particles would be very stable in a w/o microemulsion. The Rh particle diameters of Rh-SiO(2) catalysts prepared by changing the amount of silica produced were almost identical. Accordingly, the Rh particle diameter of Rh-SiO(2) catalysts could be controlled independently of Rh content by the ME method. Copyright 2001 Academic Press. PMID- 11254297 TI - Mixed Monolayers of Cyclosporin-A and Phospholipids at the Air-Water Interface. AB - Cyclosporin-A (CsA) is a pharmaceutical product which has a polypeptide structure and immunosuppressive activity. It may be administered in the form of liposomes, and this has a series of advantages. Therefore, in this paper, the possible interaction between CsA and phospholipids is studied. The superficial monolayer technique has been used as a liposome membrane model. From the isotherms of mixed monolayers of CsA and different phospholipids it is possible to prove that these systems obey the molecular area additivity rule when spread as mixed monolayers. Applying the Defay-Crisp phase rule, it is reasonable to conclude that these systems are immiscible given that they orientate themselves differently on thesurface. Copyright 2001 Academic Press. PMID- 11254298 TI - Shape Transition in Micelles of Undecylammonium Chloride in the Presence of Sodium Chloride and n-Butanol: A Study by Dynamic Light Scattering. AB - Dynamic light scattering measurements have been performed on micellar solutions of undecylammonium chloride in the presence of 0 to 0.5 mol dm(-3) n-butanol and 0 to 0.2 mol dm(-3) sodium chloride. The results support opposing effects of these additives on micellar growth, with the n-butanol causing a reduction of micelle size. The dependence of the aggregation number on the hydrodynamic radius suggests a transition from sphere to prolate ellipsoid when the aggregation number approximates that predicted for the maximum aggregation number of a spherical micelle. Micelle dimensions calculated from the aggregation number, conformational considerations, and packing criteria of the monomers in the micelle are compared with those from dynamic light scattering data. Copyright 2001 Academic Press. PMID- 11254299 TI - Scaling Behavior of the Rheological Properties of Montmorillonite Suspensions: Correlation between Interparticle Interaction and Degree of Flocculation. AB - In this work we investigate some aspects of the rheological behavior of sodium montmorillonite (NaMt) suspensions in the pH range 3 to 9, of NaCl concentrations between 10(-3) and 10(-1) M, and of solid concentrations between 5 and 11% w/v. Three kinds of experiments were performed: steady-state viscometry, oscillatory test, and creep recovery. The physical quantities of interest were the yield stress sigma(y) of the suspensions, the elastic rigidity modulus G', and the instantaneous elastic compliance. Furthermore, G' was obtained from oscillatory tests in three different experiments: determination of the viscoelastic linear region, oscillograms, and the gelation process. All quantities were found to scale with the concentration of solids, C, according to a power law of the form Y=k(y)C(n). The exponents n were found to change from approximately 3 to approximately 6 when the pH was increased from 3 to 9 (at constant ionic strength 10(-2) M), although values as high as 10 were estimated when the NaCl concentration was reduced to 1 mM. Such values of n correlate well with the characteristics of the edge-to-face (E-F), edge-to-edge (E-E), and face-to-face (F-F) interparticle interactions. The minimum values of n correspond to maximum E F attractions, whereas the largest n are associated with strong F-F repulsions. Copyright 2001 Academic Press. PMID- 11254300 TI - Study of the Reaction 2-(p-Nitrophenyl)Ethyl Bromide + OH(-) in Sulfobetaine Aqueous Micellar Solutions in the Presence and Absence of Added Salts. AB - The reaction of dehydrobromination of 2-(p-nitrophenyl)ethyl bromide with hydroxide ions has been studied in aqueous micellar solutions of N-tetradecyl-N,N dimethyl-3-ammonio-1-propanesulfonate, SB3-14. The kinetic effects of added salts (NaF, NaCl, NaBr, and NaNO(3)) on the reaction rate in SB3-14 aqueous micellar solutions have also been studied. They were rationalized by considering the binding of the anions, which come from the salt, to the sulfobetaine micelles and their competition with the reactive hydroxide ions for the micellar surface. The equilibrium binding constant of the 2-(p-nitrophenyl)ethyl bromide to the sulfobetaine micelles was estimated by recording the changes in the spectra of the organic substrate when the SB3-14 concentration in the micellar medium changed. This value was in agreement with that obtained from fitting of kinetic data. The second-order rate constant in the micellar pseudophase revealed that the reaction is faster in SB3-14 micelles than in water. This acceleration seems independent of the presence of added salts and can be explained by considering that SB3-14 micelles favor reactions in which charge is delocalized in the transition state. Copyright 2001 Academic Press. PMID- 11254301 TI - A Study of the Measurement of Surface and Interfacial Tension by the Maximum Liquid Drop Volume Method. AB - The maximum liquid drop volume (nu(max)) is measured using a back-suction technique with a micrometer syringe piston. The residual amount of liquid on the tip was determined and the tension data calculated from the theoretical correction factors are well in accordance with those from the falling drop volume (nu(f)). Copyright 2001 Academic Press. PMID- 11254302 TI - A Study of the Measurement of Surface and Interfacial Tension by the Maximum Liquid Drop Volume Method. AB - The maximum liquid drop volume (v(max)) is measured by using a back-suction micrometer syringe piston technique. Different very viscous liquids are measured by (v(max)) and (v(f)) methods to observe the effect of viscosity on tension measurement. No apparent viscosity effect was observed in surface tension data obtained by using Harkins-Brown factors and the theoretical correction factors in the viscosity range 5.9-100,900 mP. Copyright 2001 Academic Press. PMID- 11254303 TI - Microstructured Polyacrylamide Hydrogels Prepared Via Inverse Microemulsion Polymerization. AB - The synthesis by a two-stage polymerization process of microstructured polyacrylamide hydrogels with large swelling capacity and improved mechanical properties is reported. First, crosslinked polyacrylamide particles of nanosize scale are made by inverse microemulsion polymerization. These particles are then dried and redispersed in an aqueous solution of acrylamide and polymerized in the presence of a crosslinking agent. The microstructured hydrogels, in contrast to transparent conventional polyacrylamide hydrogels, are translucid due to the presence of the dispersed particles. The swelling capacity of these hydrogels increases as the particle content increases and their Young and elastic moduli (at equilibrium swelling) diminish only slightly. Mechanical tests disclose that the microstructured hydrogels have larger Young moduli than conventional hydrogels with an identical degree of swelling. Copyright 2001 Academic Press. PMID- 11254305 TI - Surface Chemical Studies on Sphalerite and Galena Using Bacillus polymyxa. AB - The interaction of sphalerite and galena with cells of Bacillus polymyxa was investigated through adsorption, electrokinetic, flotation, and flocculation studies. Adsorption experiments indicated that a higher amount of the cells was adsorbed onto galena compared to sphalerite. The adsorption density of the cells onto galena was almost independent of pH while that onto sphalerite was found to continuously decrease with increasing pH. The adsorption isotherms of the bacterial cells on galena and sphalerite exhibited Langmuirian behavior. Electrokinetic measurements showed that the negative electrophoretic mobilities of the cells were reduced in magnitude in proportion to the time of interaction with either sphalerite or galena. Similar trends were observed in the cases of sphalerite and galena after interaction with the cells. However, the magnitude of the reduction in the electrophoretic mobilities was found to be greater for galena than for sphalerite. Flotation tests revealed that galena was almost completely depressed after interaction with the cells both in the absence and in the presence of the collector. In contrast, the addition of collector and activator to sphalerite, which was initially interacted with the cells, restored the floatability at and beyond pH 8.5. Selective flotation tests on a synthetic mixture of galena and sphalerite confirmed that sphalerite could be preferentially floated from galena, which was depressed by the bacterial cells. Selective flocculation tests further demonstrated that galena could be flocculated from sphalerite, which was dispersed in the presence of cells of B. polymyxa at pH 9-9.5. Copyright 2001 Academic Press. PMID- 11254304 TI - Study of the Leacril Dyeing Process by a Cationic Dye from an Emulsion System. AB - Adsorption studies of a cationic dye, Rhodamine B, from an emulsion phase on Leacril fabric at different temperatures were conducted. The emulsion phase consisted of n-hexadecane emulsified by isopropyl alcohol (1 M) and stabilized by tannic acid. In the alcohol solution Rhodamine B was dissolved. The kinetics of its adsorption and desorption is discussed. The changes in Leacril surface free energy components in the dyeing process were also determined. The adsorption data show that the presence of an emulsion increases the dye adsorption at room temperature (293 K) and at 313 K, while at 333 K it is smaller than that from Rhodamine solution alone. However, Rhodamine desorbs more when adsorbed from the solution. Surface free energy components differ for the Leacril samples dyed at different temperatures, and the most hydrophobic surface was obtained for the samples dyed at 333 K, where the electron-donor component is the lowest one. In general, the work of water spreading is close to zero, except for the above sample for which it is relatively highly negative. Possible mechanisms of the dye adsorption are discussed. Copyright 2001 Academic Press. PMID- 11254306 TI - Surface Chemical Studies on Sphalerite and Galena Using Bacillus polymyxa. AB - Biodissolution tests reveal the release of lead/zinc species from galena/sphalerite, respectively, while biosorption experiments confirm interaction of cells of Bacillus polymyxa (B. polymyxa) with the metal ions of interest. The amount of exo-polysaccharides is found to be the highest in the case of galena-interacted cells, followed by the Bromfield medium-grown cells while the sphalerite-interacted cells have the least, based on ruthenium red adsorption studies. In contrast, the sphalerite-interacted cells assay the highest amount of protein while the galena-interacted cells have the lowest amount, on a comparative basis. The adsorption of xanthate onto galena is found to be diminished in the presence of the cells whereas the xanthate adsorption density for activated sphalerite is unaffected in the pH range 9-11. Additionally, the cell surface hydrophobicity tests confirm that the sphalerite interacted cells are more hydrophobic relative to the galena-interacted cells. FTIR spectroscopic data lend support to the higher adsorption density of the cells onto galena vis-a-vis sphalerite. The higher exo-polysaccharide and lower protein contents together with the hydrophilic nature of the galena-interacted cells could be the contributing factors to the selective flocculation and depression of galena. In a similar manner, the higher protein and lower exo polysaccharide contents as well as the greater hydrophobicity of the sphalerite interacted cells favor its floatability and dispersion. Copyright 2001 Academic Press. PMID- 11254307 TI - Thermodynamic Studies of Aqueous m-s-m Gemini Surfactant Systems. AB - The specific conductance, surface tension, and apparent molar volume properties of aqueous solutions of two series of m-s-m gemini surfactants-one having a constant spacer s(=3) with m=8, 10, 12, and 16 and the other having a constant alkyl chain length m(=12) with variable spacer length 2/=Li(+) approximately Na(+)>K(+)>NH(+)(4)>Rb(+)Tl(+) approximately Ag(+) approximately Cs(+). The parameters of Frumkin and the virial isotherms have been calculated. There is a rather strong repulsion among the adsorbed anions. To define the energy of the specific interaction between the anions and the adsorbed monolayer, the technique of E. Goddard et al. (J. Colloid Interface Sci. 24, 297 (1967)) was applied. Its application for two-phase systems permits us to define the parameters of the ion exchange reaction on the basis of measurements of interfacial tension. The ion exchange constants calculated by various methods have been compared. Copyright 2001 Academic Press. PMID- 11254324 TI - Effect of Salts on the Formation of C(8)-Lecithin Micelles in Aqueous Solution. AB - The effect of divalent and trivalent salts (CaCl(2), CaBr(2), MgCl(2), MgBr(2), LaCl(3), CeCl(3), La(NO(3))(3), and Ce(NO(3))(3)) on the micelle formation in C(8)-lecithin solutions was investigated using the techniques of static and dynamic light scattering. The critical micelle concentration (cmc), mean hydrodynamic radius (R(h)), gyration radius (R(g)), and weight-average molecular weight of the micelles were measured as functions of salt identity and concentration, amphiphile concentration, and temperature. It was found that the micelles in solutions of magnesium are less likely to form and less stable; their standard enthalpy is less exothermic as the ionic strength increases. On the contrary, the micelles in solutions of calcium and trivalent salts form easily, and are more stable; their standard enthalpy is also more exothermic as the ionic strength increases. Based on our model of the Gibb's free energy for the salt added solutions, we obtained the following formula for the effect of salts on cmc: ln(cmc)'=ln(cmc)+k(1) I(1/2)+k(2)I, where (cmc)' and (cmc) are the critical micelle concentrations in salt-added and salt-free solutions, respectively, I is the ionic strength, and k(1) and k(2) are the salt effect parameters. The agreement between the formula and the experimental data for all the systems under study shows that the formula is more satisfactory than those suggested previously by other authors in describing the effect of salts on the cmc in the micellar solutions of not only zwitterionic but also nonionic surfactants. Copyright 2001 Academic Press. PMID- 11254325 TI - The Elasticity of Adsorption Layers of Reorientable Surfactants. AB - A theoretical model is discussed which describes the relaxation of an adsorption layer under a harmonic surface area perturbation (oscillating bubble method). The model assumes sufficiently high oscillation frequencies that a diffusional exchange of matter can be neglected. As relaxation process the change in the molar area of an adsorbed molecule is considered, caused for example by changing the orientation at the interface. The model calculations show that the use of a reorientation isotherm first leads to a different dilational elasticity isotherm as compared to that obtained for a Langmuir adsorption model. Moreover, it is shown that the surface relaxation due to molecular orientation changes can influence the viscoelasticity significantly. Qualitative comparisons with literature data show good agreement. Copyright 2001 Academic Press. PMID- 11254326 TI - Synthesis of Strontium Ferrite Ultrafine Particles Using Microemulsion Processing. AB - The strontium ferrite ultrafine particles have been prepared using the microemulsion processing. The mixed hydroxide precursor was obtained via the coprecipitation of Sr(2+) and Fe(3+) in a water-in-oil microemulsion of water/CTAB/n-butanol/isooctane. According to the investigation on the thermochemical properties by TGA/DTA and the phase analysis by XRD, it was shown that the precursor could yield pure strontium ferrite after calcination at 700 degrees C for 5 h while using an appropriate molar ratio of Sr/Fe in microemulsions. From TEM measurement, the diameters of the precursor and calcined particles were 3.8+/-0.7 and 50-100 nm, respectively. The magnetic properties characterized by a SQUID magnetometer showed that the saturation magnetization, remanent magnetization, coercivity, and squareness ratio were 55 emu/g, 28 emu/g, 492 Oe, and 0.51, respectively. The magnetization was also observed to increase with the decrease of temperature at 5-400 K. Compared with those reported earlier, the quite low coercivity implies the potential application of final product in the high-density perpendicular recording media. Copyright 2001 Academic Press. PMID- 11254327 TI - Synthesis of Ferrierite-Type Zeolite in the Presence of a Catalytic Amount of Pyrrolidine and Sodium Bis(2-ethyhlhexyl) Sulfosuccinate. AB - Synthesis of ferrierite (FER) type zeolite with varying Si to Al ratios in the presence of a catalytic amount of pyrrolidine and sodium bis(2-ethylhexyl) sulfosuccinate (AOT) in aqueous media is reported. The surfactant moieties direct pyrrolidine molecules in a particular fashion, resulting in a sixfold decrease in the required amount of template as compared to conventional procedure, for FER crystallization. The product obtained was highly crystalline and pure. In the presence of AOT alone, ferrierite co-crystallized with the ZSM-5 phase, indicating AOT is not acting as a structure-directing agent and a small concentration of pyrrolidine ( approximately 2 wt%) template is essential for the ferrierite crystallization. A scanning electron micrograph showed uniformity in crystals (average 2-3 um) consisting of broad plate type morphology. The crystal structure of FER (AOT/Py) maintains structural integrity until about 1000 degrees C. FER (AOT/Py) has been further characterized by employing XRD, XRF, IR, and TG/DTA techniques. Copyright 2001 Academic Press. PMID- 11254328 TI - The Hydrotrope Action of Sodium Xylenesulfonate on the Solubility of Lecithin. AB - The activity of sodium xylenesulfonate in aqueous solution was determined by vapor pressure measurements, the results of which were transformed to mean activities. In addition, surface tension, conductivity, and partial molar volume were determined to obtain complementary information about potential association structures. No sudden change was observed, indicating the association to take place in the concentration range where the increased solubility of lecithin was observed, contrary to the case of surfactant micellization. The solubilization of lecithin in the aqueous solutions of sodium xylenesulfonate showed the association to be cooperative between the two amphiphiles. A model is proposed for the interpretation of the catastrophic incorporation threshold concentration observed in the solubility curve of lecithin. Copyright 2001 Academic Press. PMID- 11254329 TI - Dispersion Destabilization in Magnetic Water Treatment. AB - The destabilization of fine nonmagnetic particles as one of the possible mechanisms for magnetic water treatment (MWT), an alternative method for scale control in industrial water processing and amelioration of dispersion separations, is discussed. Numerical results (based on an electrical double-layer theory) for the theoretical model of surface neutralization due to ion shifts from the bulk of the solution toward the particle surfaces, are presented to show the theoretical possibility of accelerated coagulation of scale-forming particles during and after MWT. Copyright 2001 Academic Press. PMID- 11254330 TI - Evidence of Shear Rate Dependence on Restructuring and Breakup of Latex Aggregates. AB - Small-angle static light scattering has been used to probe the evolution of aggregate size and structure in the shear-induced aggregation of latex particles. The size of aggregates obtained from the particle-sizing instrument (Coulter LS230) was compared with the size of those obtained with another approach utilizing the Guinier equation on the scattering data. Comparison of the two methods for studying the effects of mixing on the evolution of the aggregate size with time revealed similar trends. The aggregate structures were quantified in terms of their fractal dimensions on the grounds of the validity of Rayleigh-Gans Debye scattering theory for the fractal aggregates. Analysis of the scattering patterns of aggregates verified that restructuring of the aggregates occurred as the aggregates were exposed to certain shear environments, resulting in a scale dependent structure that could not be quantified by a fractal dimension. The effect of restructuring on aggregate size was particularly noticeable when the aggregates were exposed to average shear rates of 40 to 80 s(-1), whereas no significant restructuring occurred at lower shear rates. At 100 s(-1), the fragmentation of aggregates appeared to be more significant than aggregate compac tion. Copyright 2001 Academic Press. PMID- 11254331 TI - Solubilization of Polar Oils in Surfactant Self-Organized Structures. AB - The cloud temperature of 2 wt% C(12)EO(8) aqueous solutions decreases upon addition of sarcosinate-lauroyl isopropyl (SLIP), 1-dodecanol, and m-xylene, whereas it increases in glycerol tris(2-ethylhexanoic) ester (TEH), isopropyl myristate (IPM), and saturated hydrocarbon systems. A three-phase microemulsion is formed at equal weights of water and oil in the IPM system, but a lamellar liquid crystal (L(alpha)) is present in the SLIP system at the balanced temperature. The effect of added oil on the phase transition of the hexagonal (H(1)) phase was also investigated by means of SAXS study. The H(1)-L(alpha) transition occurs upon addition of SLIP or 1-dodecanol whereas the H(1)-I(1) (discontinuous micellar cubic) phase transition takes place in TEH or IPM systems. These differences in phase behavior are attributed to the placement of solubilized oil in micelles: In the former systems, oil tends to penetrate in the surfactant palisade layer and induces the surfactant layer curvature in micelles to be less positive, while the penetration tendency is small and the opposite effect on the curvature is induced upon addition of the latter oils. Copyright 2001 Academic Press. PMID- 11254332 TI - Micellar Effects on S(N)2 Reactions of Alkyl Naphthalene-2-sulfonates:The Role of Hydrophobic Substituents. AB - Reactivities of methyl naphthalene-2-sulfonate, MeONs, and its 6-alkyl derivatives, alkyl=Me, n-C(6)H(13) and n-C(12)H(25), 6-Me-MeONs, 6-Hex-MeONs, and 6-Do-MeONs, respectively, are compared for reactions in cetyl trialkylammonium bromide micelles, n-C(16)H(33)NR(3)Br, R=Me, Et, n-Pr, n-Bu, CTABr, CTEABr, CTPABr, CTBABr, respectively. Similar experiments were made on reactions of Br(-) with n-butyl and n-decyl naphthalene-2-sulfonates, BuONs, and DeONs, respectively. Reactions with OH(-) were followed in cetyl trialkylammonium hydroxide, alkyl=Me, Et, n-Pr, CTAOH, CTEAOH, and CTPAOH, solubility permitting. Some reactions with OH(-) or Br(-) were also followed in mixed-ion systems of CTAOMs or CTPAOMs (OMs=MeSO(3)). Micellar rate effects were analyzed by using pseudophase treatments including interionic competition in mixed-ion systems. Second-order rate constants in the micellar pseudophase increase systematically with increases in substrate hydrophobicity and for reactions with Br(-), but not OH(-), they also increase with bulk of the cationic head group. Copyright 2001 Academic Press. PMID- 11254333 TI - Rheological Behavior of Titanium Dioxide Suspensions. AB - The rheological properties of titanium dioxide dispersed in water are measured over a wide range of powder concentrations, temperatures, and pH values. The value of intrinsic viscosity of titanium dioxide measured with an Ubbelohde capillary viscometer is 3.55, which is useful for determining the shape and aggregation property of the particles. The yield stress and steady shear viscosity of titanium dioxide with broad and narrow particle size distributions were measured over a wide range of solid volume fractions on a Brabender rheometer. It is observed that the rheological properties of the suspensions are quite different due to the difference in particle size distributions. Quemada, Casson, and Zhou's models were used to fit the experimental data and useful parameters were obtained. Calculated data are also in good agreement with the experimental data. As expected, the shear viscosity and yield stress decrease with increasing temperature. But when the temperature is around 50 degrees C, yield stress increases with increasing temperature while shear viscosity exhibits a complex behavior. The phenomena are very interesting and special. The Peclet number was used to analyze the shear thickening behavior. Models were also used to describe the shear viscosity under different temperatures and the master plots of the reduced variables eta/eta(infinity) vs t(c)gamma; at different temperatures are superimposed, which means the agreement is fair and the models are suitable to describe the rheological properties of titanium dioxide suspensions. pH effects were investigated on a Rheometrics RFS-II rheometer and it was found that pH can change the surface charge of the particles, which also affects the rheological behavior. The pH at which maximum shear viscosity and yield stress occur is in concordance with the isoelectric point. Copyright 2001 Academic Press. PMID- 11254334 TI - Voltammetric Characterization of trans-Dioxo Ethylenediamine Complexes of Re(V) in Aqueous Solutions. AB - The electrochemical behavior of trans-[Re((V))O(2)(en)(2)]I and trans [Re((V))O(2) (en)(2)]ClO(4) (en=ethylenediamine) complexes was studied by cyclic voltammetry on Au electrodes. Experiments were performed in aqueous solutions at pH 7.0 and at room temperature. The complex voltammogram was characterized by Re containing species, assigned to the [Re((V))O(2)(en)(2)](+)/[Re((IV))O(2)(en)(2)] couple, and I-containing species. To overcome I interference, the electrochemical response of Re complexes was segregated by performing a reductive desorption of adsorbed I from Au. Copyright 2001 Academic Press. PMID- 11254335 TI - Sugar Amphiphiles as Revealing Dopants for Induced Chiral Nematic Lyotropic Liquid Crystals. AB - The existence of phase chirality in lyotropic liquid crystals still raises questions. The mechanisms behind the transfer of chirality throughout the long range orientational order are not yet obvious. Guest/host systems with chiral dopants in achiral host phases offer the capability of systematic investigations. We demonstrate that the large amount of accessible sugar amphiphiles exhibits remarkable structure/property relations. Their helical twisting power HTP increases strongly with the number of sugar units of a dopant molecule. The spatial range of the chirality information reaching from a chirally doped micelle to adjacent aggregates is essential for the development of phase chirality. The induced twist of the lyotropic nematic host phase is highly sensitive to small changes of the sugar type (e.g., galacto- to glucopyranose). Depending on the nature of the host phase, either the alpha- or the beta-linkage of the sugar to the hydrophobic moiety of the sugar dopant results in larger HTP values. We propose that our amphiphilic sugar derivatives act like antennae to transfer chirality information. Their effectiveness as chiral dopants is due to a hydrophobic anchoring within the micelles and an extension of their chiral moiety far into the intermicellar region. The chirality transfer works especially well if the hydrophilic and chiral sugar moieties are oriented toward a neighboring micelle in the direction of the helix axis. Copyright 2001 Academic Press. PMID- 11254336 TI - Adsorption of Poly(ethylene oxide)-Poly(lactide) Copolymers. Effects of Composition and Degradation. AB - The effect of chemical degradation of two diblock copolymers of poly(ethylene oxide) (E) and poly(lactide) (L), E(39)L(5) and E(39)L(20), on their adsorption at silica and methylated silica was investigated with in situ ellipsometry. Steric stablization of polystyrene dispersions was investigated in relation to degradation. Hydrolysis of the poly(lactide) block of the copolymers was followed at different temperatures and pH by using HPLC to measure the occurrence of lactic acid in solution. The block copolymers were quite stable in pH-unadjusted solution at low temperature, whereas degradation was facilitated by increasing temperature or lowering of the pH. Lower degradation rates of E(39)L(20) where observed at low temperature in comparison with those of E(39)L(5), whereas the degradation rates of the copolymers were quantitatively similar at high temperature. The adsorption of the copolymers at methylated silica substrates decreased with increasing degree of degradation due to the reduction in the ability of hydrophobic block to anchor the copolymer layer at the surface. At silica the adsorption initially increased with increasing degradation, particularly for E(39)L(20) due to deposition of aggregates onto the surface. After extensive degradation the adsorption of the copolymers at both silica and methylated silica resembled that of the corresponding poly(ethylene oxide) homopolymer. Overall, it was found that the eventual reduction in adsorption occurred at a lower degree of degradation for E(39)L(5) than for E(39)L(20). Mean field calculations showed a reduced anchoring for the block copolymers with decreasing poly(lactide) block length at hydrophobic surfaces. In accordance with this finding, it was observed that polystyrene dispersions were stabilized by E(39)L(20) or E(39)L(5) in a way that depended on both the lactide block length and the degree of degradation. Upon degradation of the hydrophobic block, stabilization of the polystyrene dispersions was maintained initially, but eventually degradation resulted in destabilization. The average residual copolymer concentration required for stabilization of the polystyrene dispersions was much higher than the corresponding concentration of intact copolymer required for stabilization. Copyright 2001 Academic Press. PMID- 11254337 TI - Structure Analysis of Montmorillonite Intercalated with Cetylpyridinium and Cetyltrimethylammonium: Molecular Simulations and XRD Analysis. AB - Molecular mechanics and molecular dynamics simulations combined with X-ray powder diffraction were used in structure investigation of montmorillonite intercalated with cetylpyridinium (CP) and cethyltrimethylammonium (CTA) cations. Molecular modeling revealed the interlayer structure and differences in intercalation behavior of CP and CTA cations in montmorillonite. The experimental and calculated values of basal spacing were in good agreement for both intercalates: in the case of CP-montmorillonite d(exp)=20.59 A, d(calc)=20.60 A; for CTA montmorillonite d(exp)=18.00 A and d(calc)=18.10 A. CTA-montmorillonite exhibits significantly higher total sublimation energy and higher host-guest interaction energy than the CP-montmorillonite. The main difference between both intercalates is in charge distribution on the host layers and guest species. The charge transfer from the guest species to the host layer is higher in CTA montmorillonite than in CP-montmorillonite, and consequently the charge polarization between the host and guest layers is much higher in CTA montmorillonite. This leads to much stronger host-guest electrostatic interaction in the case of CTA-montmorillonite. Copyright 2001 Academic Press. PMID- 11254339 TI - Forces between a Rigid Probe Particle and a Liquid Interface. AB - The effect of disjoining pressure between a rigid spherical probe particle (attached to an AFM cantilever) and a liquid interface (e.g., oil/water or air/water) is treated in an analytic manner to describe the total force F exerted on the probe as a function of the distance X of the probe from the rigid substrate (AFM stage) on which the liquid interface resides. Two cases (i) a flat interface under gravity and (ii) a drop whose size is sufficiently small that gravity can be neglected have been examined. A simple numerical algorithm is given for computing F(X) (the AFM observable) from a given form for the disjoining pressure. Numerical results are displayed for electrostatic probe/interface interactions which reveal the linear compliance regime experimentally observed in AFM experiments on these systems. The slope of the linear compliance regime is shown to be a function of the properties of the interface (capillary length, particle radius, drop size, contact angle of drop on rigid substrate etc.). Copyright 2001 Academic Press. PMID- 11254338 TI - Thermally Induced Transient Activity Changes of Plasmin Adsorbed onto Bare and Fibrinogen-Modified Graphite and Glassy Carbon Surfaces. AB - The occurrence of a thermally induced first-order transition affecting the amidolytic activity of plasmin adsorbed onto bare and protein-modified graphite and glassy carbon was demonstrated in the 10-45 degrees C temperature range in the presence of a chromogenic substrate. Modification of the surfaces was achieved upon spontaneous adsorption of plasmin to surfaces bearing a coating of fibrinogen, whether electrochemically oxidized or not. The amount of fibrinogen adsorbed at graphite was determined by ELISA. The kinetics of the transition was characterized by its starting temperature (T(c)), which was between 14 and 19 degrees C, the first-order rate constant, and the activation energy E(a) deduced from Arrhenius plots. Results showed the absence of a correlation between T(c), E(a), and contact angle variations. It is therefore likely that these variables address separate steps in a complex pathway of reactions undergone by plasmin under mild thermal constraints. Copyright 2001 Academic Press. PMID- 11254340 TI - Effect of Corona Discharge Treatment on the Dyeability of Low-Density Polyethylene Film. AB - The effect of corona discharge on low-density polyethylene (LDPE) film was studied in terms of surface functionality and surface energetics of the film surfaces, improving the dyeability. The introduction of a polar group (O=C-O, C=O, and C-O) to a corona-treated LDPE film with acrylic acid could be confirmed by ESCA. The Owens-Wendt and Wu models using geometric means were studied to analyze the surface free energy of corona-treated film. It was found that the corona-treated LDPE film did lead to an increase in surface free energy, mainly due to the increase of its specific (or polar) component as the corona discharge power increased. Also, the K/S values were increased as the concentrations of dye increased. From the acid-base interaction point of view, it was found that the graft polymerization of acrylic acid onto the corona-treated LDPE film plays an important role in growing the acidic character which is one of the specific components of surface free energy, resulting in improving the dyeability with basic dyeing agent. A direct linear relationship is shown between the O(1s)/C(1s) ratio and the resulting K/S value or the specific component for this work. Copyright 2001 Academic Press. PMID- 11254341 TI - The Pesticide 3-(3,4-Dichlorophenyl)-1,1-dimethylurea (Diuron) Immobilized on Silica Gel Surface. AB - A route for 3-(3,4-dichlorophenyl)-1,1-dimethylurea (diuron) immobilization on silica gel was established after reacting at the first stage the precursor silylant agent 3-trimethoxysilylpropylamine to the support. The pesticide was covalently bonded to available amine groups of the precursor, giving 1.03 mmol of amine per gram of silica. Infrared, (13)C, and (29)Si NMR spectra are in agreement with the proposed reaction between nitrogen of the amine group of the previously anchored silica to carbon on the para-position of the aromatic ring of the pesticide. The immobilization is clearly affected by the presence or absence of disprotonating agent, to give 12.50 and 68.40% reaction yield, respectively; these results were confirmed through elemental analysis. Copyright 2001 Academic Press. PMID- 11254342 TI - Electrochemistry and Electrocatalysis with Myoglobin in Biomembrane-Like DHP-PDDA Polyelectrolyte-Surfactant Complex Films. AB - The polyelectrolyte-surfactant complex DHP-PDDA was prepared by reacting the anionic surfactant dihexadecylphosphate (DHP) with polycationic poly(diallyldimethylammonium) (PDDA). Thin films made from DHP-PDDA on solid substrates demonstrated an ordered multibilayer structure by XRD and DSC. Incorporated myoglobin (Mb) in DHP-PDDA films on pyrolytic graphite (PG) electrodes showed a pair of well-defined and nearly reversible cyclic voltammetric peaks for the Mb Fe(III)/Fe(II) couple at about -0.3 V vs SCE in pH 7.0 buffers. Electron transfer between Mb and PG electrodes was greatly facilitated in the film microenvironment. The positions of the Soret absorption band suggest that Mb maintains its secondary structure similar to its native state in DHP-PDDA films in the medium pH range. Mb could act as an enzyme-like catalyst in DHP-PDDA films as demonstrated by catalytic reduction of trichloroacetic acid, nitrite, and oxygen with a decrease in the electrode potentials required. Mb-DHP-PDDA films may thus have potential application as biosensors. Copyright 2001 Academic Press. PMID- 11254343 TI - Iterative Solution Method for the Linearized Poisson-Boltzmann Equation: Indirect Boundary Integral Equation Approach. AB - An iterative solution scheme is proposed for solving the electrical double-layer interactions governed by the linearized Poisson-Boltzmann equation. The method is based on the indirect integral equation formulation with the double-layer potential kernel of the linearized Poisson-Boltzmann equation. In contrast to the conventional direct integral equation approach that yields Fredholm integral equations of the first kind, the indirect integral equation approach yields well posed Fredholm integral equations of the second kind. The eigenvalue analysis reveals that the spectral radius of the double-layer integral operator is always less than one. Thus, iterative solution schemes can be successfully implemented for solving the electrical double-layer interactions for very large and complex systems. The utility of the iterative indirect method is demonstrated for several examples which include spherical and spheroidal particles. Copyright 2001 Academic Press. PMID- 11254345 TI - Comparison of Gaseous Molecular Adsorption Properties of Microporous Manganese Oxides and Crystalline Aluminosilicates. AB - The N(2), H(2)O, and NH(3) adsorption properties on hollandite-type (H-Hol) and birnessite-type (H-Bir) hydrous manganese oxides were compared with those on zeolitic crystals. It was found that the effective pore openings of H-Bir and H Hol are below 0.3 nm. Although the geometric spaces on H-Bir and H-Hol available for physical adsorption from a gaseous phase are much smaller than those on zeolites, they can provide a strong chemisorption field for H(2)O and NH(3) intercalation. Copyright 2001 Academic Press. PMID- 11254344 TI - The Electrophoretic Mobility and Electric Conductivity of a Concentrated Suspension of Colloidal Spheres with Arbitrary Double-Layer Thickness. AB - The electrophoresis in a monodisperse suspension of dielectric spheres with an arbitrary thickness of the electric double layers is analytically studied. The effects of particle interactions are taken into account by employing a unit cell model, and the overlap of the double layers of adjacent particles is allowed. The electrokinetic equations, which govern the ionic concentration distributions, the electric potential profile, and the fluid flow field in the electrolyte solution surrounding the charged sphere in a unit cell, are linearized assuming that the system is only slightly distorted from equilibrium. Using a perturbation method, these linearized equations are solved with the surface charge density (or zeta potential) of the particle as the small perturbation parameter. Analytical expressions for the electrophoretic mobility of the colloidal sphere in closed form correct to O(zeta) are obtained. Based on the solution of the electrokinetic equations in a cell, a closed-form formula for the electric conductivity of the suspension up to O(zeta(2)) is derived from the average electric current density. Comparisons of the results of the cell model with different conditions at the outer boundary of the cell are made for both the electrophoretic mobility and the electric conductivity. Copyright 2001 Academic Press. PMID- 11254346 TI - Electron-Transfer-Induced Molecular Reorientations: The Benzoquinone/Hydroquinone Reaction at Pd(111)-(square3xsquare3)R30 degrees -I Studied by EC-STM. AB - The benzoquinone/hydroquinone (Q/H(2)Q) redox reaction has been studied by electrochemical-scanning tunneling microscopy (EC-STM) at a Pd(111) (square3xsquare3)R30 degrees -I electrode surface in a solution that contained 10(-4) M H(2)Q in 0.05 M H(2)SO(4); iodine-pretreatment of the Pd(111) surface was to prevent chemisorption of organic-derived species. The molecule-resolved EC STM images indicated that: (i) at a potential where only H(2)Q is present in solution, a self-assembled (square21xsquare21)R10.9 degrees -eta(6)-H(2)Q monolayer is produced in which the H(2)Q molecules are oriented parallel to the surface; (ii) at a potential where partial oxidation (to Q) occurs, a self assembled (square21xsquare21)R10.9 degrees -eta(6)-QH adlayer is generated, where QH represents quinhydrone, an equimolar mixture of Q and H(2)Q; in this structure, the Q and H(2)Q molecules are oriented vertically, face-to-face, and arranged alternately along a given row, reminiscent of the crystal structure of quinhydrone. The partial oxidation-induced molecular reorientation, which is reversible, most likely arises from favorable Q-H(2)Q face-to-face interactions; that is, complete oxidation would yield only flat-oriented Q species. Unfortunately, at potentials where only Q would be present in solution, I catalyzed corrosion of the Pd starts to occur, which leads to noise-laden EC-STM images. Copyright 2001 Academic Press. PMID- 11254347 TI - Vertebrate cranial placodes I. Embryonic induction. AB - Cranial placodes are focal regions of thickened ectoderm in the head of vertebrate embryos that give rise to a wide variety of cell types, including elements of the paired sense organs and neurons in cranial sensory ganglia. They are essential for the formation of much of the cranial sensory nervous system. Although relatively neglected today, interest in placodes has recently been reawakened with the isolation of molecular markers for different stages in their development. This has enabled a more finely tuned approach to the understanding of placode induction and development and in some cases has resulted in the isolation of inducing molecules for particular placodes. Both morphological and molecular data support the existence of a preplacodal domain within the cranial neural plate border region. Nonetheless, multiple tissues and molecules (where known) are involved in placode induction, and each individual placode is induced at different times by a different combination of these tissues, consistent with their diverse fates. Spatiotemporal changes in competence are also important in placode induction. Here, we have tried to provide a comprehensive review that synthesises the highlights of a century of classical experimental research, together with more modern evidence for the tissues and molecules involved in the induction of each placode. PMID- 11254348 TI - Putative stem cells and the lineage of rod photoreceptors in the mature retina of the goldfish. AB - The retinas of teleost fish grow continuously, in part, by neuronal hyperplasia and when lesioned will regenerate. Within the differentiated retina, the growth associated hyperplasia results in the generation of new rod photoreceptors only, whereas injury-induced neurogenesis results in the regeneration of all retinal cell types. It is believed, however, that both new rod photoreceptors and regenerated neurons originate from the same populations of intrinsic progenitors. Experiments are described here that attempt to identify in the normal retina of goldfish neuronal progenitors intrinsic to the retina, particularly those which have remained cryptic because they divide infrequently. Long-term, systemic exposure to bromodeoxyuridine (BrdU) was used to label these cells. Five populations of proliferative cells were labeled: microglia, which are briefly described but not studied further; retinal progenitors in the circumferential germinal zone (CGZ); and rod precursors in the outer nuclear layer (ONL), both of which have been well characterized previously; and two populations of slowly dividing cells in the inner nuclear layer (INL). The majority of these cells have a fusiform morphology, whereas the remaining ones are spherical. Longitudinal BrdU labeling suggests that the fusiform cells migrate to the ONL to replenish the pool of rod precursors. A subset of the spherical cells express pax6, although none are stained with markers of differentiated amacrine or bipolar cells. It is hypothesized that these rare, pax6-expressing cells are retinal stem cells, which give rise to the pax6-negative fusiform cells. Based on these data, two models are proposed: the first describes the lineage of rod photoreceptors in goldfish; the second is a consensus model of neurogenesis in the retinas of all teleosts. PMID- 11254349 TI - Myf-5 is transiently expressed in nonmuscle mesoderm and exhibits dynamic regional changes within the presegmented mesoderm and somites I-IV. AB - Myf-5 is one of four myogenic regulatory factors that play important roles in skeletal muscle development. This study provides detailed analysis of Myf-5 expression during early chick development using an in situ hybridization technique that has been optimized to detect low level Myf-5 transcripts. This facilitated detection of heretofore unrecognized dynamic changes in Myf-5 expression patterns. Myf-5 expression is first detected at stage 3 in the primitive streak and exhibits transient low-level expression in nonmyogenic mesoderm. Myf-5 is later expressed in the presegmented mesoderm (psm) in a reiterating pattern that is coordinated with somitogenesis and also colocalizes with the Notch ligand C-Delta-1. In somites (S) I-IV, Myf-5 expression exhibits dynamic regional changes, and in somites rostral to S IV, Myf-5 is expressed at higher levels in muscle precursors in the dorsomedial somite. Semiquantitative comparison of Myf-5 mRNA levels in the psm and in myotome-containing somites indicates about a 10-fold difference. The expression pattern of Myf-5 differs from that of MyoD, which we find is expressed only in the dorsomedial somite. These data reveal that Myf-5 is expressed at low levels several stages before muscle differentiation occurs and suggest that only a subset of cells that initially express Myf-5 will upregulate its expression and differentiate as muscle. PMID- 11254350 TI - Bicaudal-D is essential for egg chamber formation and cytoskeletal organization in drosophila oogenesis. AB - Bicaudal-D (Bic-D) is required for the transport of determinant mRNAs and proteins to the presumptive oocyte, an essential step in the differentiation of the oocyte. Bic-D protein contains four well-defined heptad repeat domains characteristic of intermediate filament proteins. We characterized the ovarian phenotypes of females expressing mutant Bic-D proteins (Bic-D(H)) deleted for each of the heptad repeat domains. The altered migration of follicle cells we observe in mutant ovaries suggests that Bic-D functions in the germline and directs the inward migration of somatic follicle cells. In the germarium Bic-D is required for the organization of the egg chamber and the structural integrity of the oocyte and nurse cells. Examination of the polarized microtubule network in Bic-D(H) ovaries shows that Bic-D function is required for both the establishment of the polarized microtubule network and its maintenance throughout oogenesis. To explain the multiple functions suggested by the pleiotropic Bic-D phenotype, we propose that Bic-D protein could form itself a filamentous structure and represent an integral, essential part of the cytoskeleton. PMID- 11254351 TI - Analyses of reproductive interactions that occur after heterospecific matings within the genus Caenorhabditis. AB - Formation of zygotes in internally fertilizing organisms requires a number of successful interactions between oocytes and sperm within a receptive female reproductive tract. These interactions are usually assumed to be species specific. For most species, it is either not possible to inseminate females with sperm from a different species or not possible to observe the consequences of such an insemination because the female is opaque. Nematodes of the genus Caenorhabditis are optically transparent and prior work indicates copulation between individuals of two different species is possible. We have used a series of vital stains and other cytological methods to analyze sperm after cross species mating. We present here a series of analyses of the postcopulatory, prefertilization interactions among three Caenorhabditis species and find that reproductive biology is conserved, to varying degrees, among all three species. This approach allows investigation into which in vivo interactions between sperm and both oocytes and the somatic gonad have been maintained during the reproductive isolation that accompanies speciation. PMID- 11254352 TI - Altered primary myogenesis in NFATC3(-/-) mice leads to decreased muscle size in the adult. AB - Signal transduction pathways involving calcineurin and its downstream effector NFAT have been implicated in regulating myogenesis. Several isoforms of NFAT exist that may differentially contribute to regulating skeletal muscle physiology. The purpose of this study was to determine the role of the NFATC3 isoform in skeletal muscle development. Adult mice lacking NFATC3 have reduced muscle mass compared to control mice. The smaller size of the muscles is not due to atrophy or blunted myofiber growth, but rather to a reduced number of myofibers. This reduction in myofiber number is not limited to a specific fiber type nor are the proportions of fiber types altered. The lower fiber number found in the adult NFATC3(-/-) mice is a consequence of impaired muscle development during embryogenesis. Immunohistochemical studies of E15 EDL muscles indicate that the total number of primary myofibers is decreased in NFATC3(-/-) embryos. At E17.5 no further decrease in primary myofiber number occurs; the size and organization of the myofibers are unaltered, and secondary myogenesis proceeds normally, suggesting a role for NFATC3 during early events in primary myogenesis. These results suggest a heretofore unknown role for the transcription factor NFAT in early skeletal muscle development. PMID- 11254353 TI - Sperm factor initiates capacitance and conductance changes in mouse eggs that are more similar to fertilization than IP(3)- or Ca(2+)-induced changes. AB - We used patch clamp electrophysiology and concurrent imaging with the Ca(2+) sensitive dye, fura-2, to study the temporal relationship between membrane capacitance and conductance and intracellular free Ca(2+) concentration ([Ca(2+)](i)) during mouse egg fertilization. We found an approximately 2 pF step increase in egg membrane capacitance and a minor increase in conductance with no change in [Ca(2+)](i) at sperm fusion. This was followed approximately 1 min later by a rise in [Ca(2+)](i) that led to larger changes in capacitance and conductance. The most common pattern for these later capacitance changes was an initial capacitance decrease, followed by a larger increase and eventual return to the approximate starting value. There was some variation in this pattern, and sub-microM peak [Ca(2+)](i) favored capacitance decrease, while higher [Ca(2+)](i) favored capacitance increase. The magnitude of accompanying conductance increases was variable and did not correlate well with peak [Ca(2+)](i). The intracellular introduction of porcine sperm factor reproduced the postfusion capacitance and conductance changes with a similar [Ca(2+)](i) dependence. Raising [Ca(2+)](i) by the intracellular introduction of IP(3) initiated fertilization-like capacitance changes, but the conductance changes were slower to activate. Capacitance decrease could be induced when [Ca(2+)](i) was increased modestly by activation of an endogenous Ca(2+) current, with little effect on resting conductance. These results suggest that net turnover of the mouse egg surface membrane is sensitive to [Ca(2+)](i) and that sperm and the active component of sperm factor may be doing more than initiating the IP(3) mediated release of intracellular Ca(2+). PMID- 11254354 TI - Agenesis of the scapula in Emx2 homozygous mutants. AB - The shoulder and pelvic girdles represent the proximal bones of the appendicular skeleton that connect the anterior and posterior limbs to the body trunk. Although the limb is a well-known model in developmental biology, the genetic mechanisms controlling the development of the more proximal elements of the appendicular skeleton are still unknown. The knock-out of Pax1 has shown that this gene is involved in patterning the acromion, while the expression pattern candidates Hoxc6 as a gene involved in scapula development. Surprisingly, we have found that scapula and ilium do not develop in Emx2 knock-out mice. In the homozygous mutants, developmental abnormalities of the brain cortex, the most anterior structure of the primary axis of the body, are associated with important defects of the girdles, the more proximal elements of the secondary axis. These abnormalities suggest that the molecular mechanisms patterning the more proximal elements of the limb axis are different from those patterning the rest of appendicular skeleton. While Hox genes specify the different segments of the more distal part of the appendicular skeleton forming the limb, Emx2 is concerned with the more proximal elements constituting the girdles. PMID- 11254355 TI - Drosophila single-minded represses gene transcription by activating the expression of repressive factors. AB - The Drosophila single-minded gene controls CNS midline cell development by both activating midline gene expression and repressing lateral CNS gene expression in the midline cells. The mechanism by which Single-minded represses transcription was examined using the ventral nervous system defective gene as a target gene. Transgenic-lacZ analysis of constructs containing fragments of the ventral nervous system defective regulatory region identified sequences required for lateral CNS transcription and midline repression. Elimination of Single minded:Tango binding sites within the ventral nervous system defective gene did not affect midline repression. Mutants of Single-minded that removed the DNA binding and transcriptional activation regions abolished ventral nervous system defective repression, as well as transcriptional activation of other genes. The replacement of the Single-minded transcriptional activation region with a heterologous VP16 transcriptional activation region restored the ability of Single-minded to both activate and repress transcription. These results indicate that Single-minded indirectly represses transcription by activating the expression of repressive factors. Single-minded provides a model system for how regulatory proteins that act only as transcriptional activators can control lineage-specific transcription in both positive and negative modes. PMID- 11254356 TI - Regulatory analysis of the mouse Hoxb3 gene: multiple elements work in concert to direct temporal and spatial patterns of expression. AB - The expression pattern of the mouse Hoxb3 gene is exceptionally complex and dynamic compared with that of other members of the Hoxb cluster. There are multiple types of transcripts for Hoxb3 gene, and the anterior boundaries of its expression vary at different stages of development. Two enhancers flanking Hoxb3 on the 3' and 5' sides regulate Hoxb2 and Hoxb4, respectively, and these control regions define the two ends of a 28-kb interval in and around the Hoxb3 locus. To assay the regulatory potential of DNA fragments in this interval we have used transgenic analysis with a lacZ reporter gene to locate cis-elements for directing the dynamic patterns of Hoxb3 expression. Our detailed analysis has identified four new and widely spaced cis-acting regulatory regions that can together account for major aspects of the Hoxb3 expression pattern. Elements Ib, IIIa, and IVb control gene expression in neural and mesodermal tissues; element Va controls mesoderm-specific gene expression. The most anterior neural expression domain of Hoxb3 is controlled by an r5 enhancer (element IVa); element IIIa directs reporter expression in the anterior spinal cord and hindbrain up to r6, and the region A enhancer (in element I) mediates posterior neural expression. Hence, the regulation of segmental expression of Hoxb3 in the hindbrain is different from that of Hoxa3, as two separate enhancer elements contribute to expression in r5 and r6. The mesoderm-specific element (Va) directs reporter expression to prevertebra C1 at 12.5 dpc, which is the anterior limit of paraxial mesoderm expression for Hoxb3. When tested in combinations, these cis elements appear to work as modules in an additive manner to recapitulate the major endogenous expression patterns of Hoxb3 during embryogenesis. Together our study shows that multiple control elements direct reporter gene expression in diverse tissue-, temporal-, and spatially restricted subset of the endogenous Hoxb3 expression domains and work in concert to control the neural and mesodermal patterns of expression. PMID- 11254357 TI - A role for BMP signalling in heart looping morphogenesis in Xenopus. AB - The heart develops from a linear tubular precursor, which loops to the right and undergoes terminal differentiation to form the multichambered heart. Heart looping is the earliest manifestation of left-right asymmetry and determines the eventual heart situs. The signalling processes that impart laterality to the unlooped heart tube and thus allow the developing organ to interpret the left right axis of the embryo are poorly understood. Recent experiments in zebrafish led to the suggestion that bone morphogenetic protein 4 (BMP4) may impart laterality to the developing heart tube. Here we show that in Xenopus, as in zebrafish, BMP4 is expressed predominantly on the left of the linear heart tube. Furthermore we demonstrate that ectopic expression of Xenopus nodal-related protein 1 (Xnr1) RNA affects BMP4 expression in the heart, linking asymmetric BMP4 expression to the left-right axis. We show that transgenic embryos overexpressing BMP4 bilaterally in the heart tube tend towards a randomisation of heart situs in an otherwise intact left-right axis. Additionally, inhibition of BMP signalling by expressing noggin or a truncated, dominant negative BMP receptor prevents heart looping but allows the initial events of chamber specification and anteroposterior morphogenesis to occur. Thus in Xenopus asymmetric BMP4 expression links heart development to the left-right axis, by being both controlled by Xnr1 expression and necessary for heart looping morphogenesis. PMID- 11254358 TI - Pulmonary hypoplasia in mice lacking tumor necrosis factor-alpha converting enzyme indicates an indispensable role for cell surface protein shedding during embryonic lung branching morphogenesis. AB - Many membrane-bound protein precursors, including cytokines and growth factors, are proteolytically shed to yield soluble intercellular regulatory ligands. The responsible protease, tumor necrosis factor-alpha converting enzyme (TACE/ADAM 17), is a transmembrane metalloprotease-disintegrin that cleaves multiple cell surface proteins, although it was initially identified for the enzymatic release of tumor necrosis factor-alpha (TNF-alpha). Mammalian lung growth and development are tightly controlled by cytokines and peptide growth factors. However, the biological function of the cell shedding mechanism during lung organogenesis is not understood. We therefore evaluated the role of TACE as a "sheddase" during lung morphogenesis by analyzing the developmental phenotypes of lungs in mice with an inactive TACE gene in both in vivo and ex vivo organ explant culture. Neonatal TACE-deficient mice had visible respiratory distress and their lungs failed to form normal saccular structures. These newborn mutant lungs had fewer peripheral epithelial sacs with deficient septation and thick-walled mesenchyme, resulting in reduced surface for gas exchange. At the canalicular stage of E16.5, the lungs of TACE mutant mice were impaired in branching morphogenesis, inhibited in epithelial cell proliferation and differentiation, and delayed in vasculogenesis. Embryonic TACE knockout mouse lungs (E12) branched poorly compared to wild-type lungs, when placed into serumless organ culture. Gene expression of both surfactant protein-C and aquaporin-5 were inhibited in cultured TACE-mutant embryonic lungs, indicating defects in both branching and peripheral epithelial cytodifferentiation in the absence of TACE protein. Furthermore, both the hypoplastic phenotype and the delayed cytodifferentiation in TACE-deficient lungs were rescued by exogenous addition of soluble stimulatory factors including either TNF-alpha or epidermal growth factor in embryonic lung culture. Thus, the impaired lung branching and maturation without TACE suggest a broad role for TACE in the processing of multiple membrane-anchored proteins, one or more of which is essential for normal lung morphogenesis. Taken together, our data indicate that the TACE-mediated proteolytic mechanism which enzymatically releases membrane-tethered proteins plays an indispensable role in lung morphogenesis, and its inactivation leads to abnormal lung development. PMID- 11254359 TI - Pecam-1 is a modulator of stat family member phosphorylation and localization: lessons from a transgenic mouse. AB - PECAM-1 (CD31) is a member of the immunoglobin (Ig) superfamily of cell adhesion molecules whose expression is restricted to hematopoietic and vascular cells. PECAM-1 can recruit adapter and signaling molecules via its immunoreceptor tyrosine activation motif (ITAM), suggesting that PECAM-1 plays a role in signal transduction pathways. To study the involvement of PECAM-1 in signaling cascades in vivo, we used the major histocompatibility (MHC) I gene promoter to target ectopic PECAM-1 expression in transgenic mice. We noted an attenuation of mammary gland development at early stages of virgin ductal branching morphogenesis. STAT5a, a modulator of milk protein gene expression during lactation, was localized to the nuclei of ductal epithelial cells of 6-week-old virgin PECAM-1 transgenics, but not in control mice. This correlated with decreases in ductal epithelial cell proliferation and induction of p21, an inhibitor of cell cycle progression. Using in vitro model systems we demonstrated PECAM-1/STAT5a association and found that residue Y701 in PECAM-1's cytoplasmic tail is important for PECAM-1/STAT5 association and that PECAM-1 modulates increases in STAT5a tyrosine phosphorylation levels. We suggest that by serving as a scaffolding, PECAM-1 can bring substrates (STAT5a) and enzymes (a kinase) into close proximity, thereby modulating phosphorylation levels of selected proteins, as previously noted for beta-catenin. PMID- 11254360 TI - Protein tyrosine phosphatase PTP1 negatively regulates Dictyostelium STATa and is required for proper cell-type proportioning. AB - The protein tyrosine phosphatase PTP1, which mediates reversible phosphorylation on tyrosine, has been shown to play an important regulatory role during Dictyostelium development. Mutants lacking PTP1 develop more rapidly than normal, while strains that overexpress PTP1 display aberrant morphology. However, the signalling pathways involved have not been characterised. In reexamining these strains, we have found that there is an inverse correlation between levels of PTP1 activity, the extent of tyrosine phosphorylation on Dictyostelium STATa after treatment with cAMP, and the proportion of the slug population exhibiting STATa nuclear enrichment in vivo. This suggests that PTP1 acts to attenuate the tyrosine phosphorylation of STATa and downstream STATa-mediated pathways. Consistent with this, we show that when PTP1 is overexpressed, there is increased expression of a prestalk cell marker at the slug posterior, a phenocopy of STATa null slugs. In ptp1 null strains, STATa tyrosine phosphorylation and nuclear enrichment in the slug anterior is increased. There is also a change in the prestalk to prespore cell ratio. Synergy experiments suggest that this is due to a cell-autonomous defect in forming the subset of prespore cells that are located in the anterior prespore region. PMID- 11254361 TI - Contribution of BDNF-mediated inhibition in patterning avian skin innervation. AB - Multiple factors are involved in the development and regulation of sensory innervation in skin. The findings we report here suggest that brain-derived neurotrophic factor (BDNF)-mediated inhibition may play an important role in determining the pattern of sensory innervation in avian skin. In birds, cutaneous innervation is restricted to dermis, where axons form a ring of innervation around the base of each feather. Here we show that both BDNF message and protein are more abundant in avian epidermis than dermis when innervation is being established; the BDNF in dermis is localized to feather buds. In vitro, BDNF caused growth cones of NGF-dependent dorsal root ganglion neurons to collapse. Similarly, outgrowth of neurites toward BDNF-secreting fibroblasts was inhibited. The inhibitory effects of BDNF appear to be mediated by the low-affinity p75 neurotrophin receptor, rather than a trk receptor. Thus, the distribution of BDNF in embryonic avian skin and the inhibitory effects of BDNF on cutaneous neurites in vitro suggest that BDNF may be important in restricting axons from entering the epidermis and the core of feather buds during development in vivo. PMID- 11254362 TI - During multicellular migration, myosin ii serves a structural role independent of its motor function. AB - We have shown previously that cells lacking myosin II are impaired in multicellular motility. We now extend these results by determining whether myosin contractile function is necessary for normal multicellular motility and shape control. Myosin from mutants lacking the essential (mlcE(-)) myosin light chain retains the ability to form bipolar filaments that bind actin, but shows no measurable in vitro or in vivo contractile function. The contractile function is necessary for cell shape control since mlcE(-) cells, like myosin heavy-chain null mutants (mhcA(-)), were defective in their ability to control their three dimensional shape. When mixed with wild-type cells in chimeric aggregation streams, the mlcE(-) cells were able to move normally, unlike mhcA(-) cells which accumulated at the edges of the stream and became distorted by their interactions with wild-type cells. When mhcA(-) cells were mixed with mlcE(-) streams, the mhcA(-) cells were excluded. The normal behavior of the mlcE(-) cells in this assay suggests that myosin II, in the absence of motor function, is sufficient to allow movement in this constrained, multicellular environment. We hypothesize that myosin II is a major contributor to cortical integrity even in the absence of contractile function. PMID- 11254363 TI - Regulation of transcription factors MHR4 and betaFTZ-F1 by 20-hydroxyecdysone during a larval molt in the tobacco hornworm, Manduca sexta. AB - During the last larval molt in Manduca sexta, a number of transcription factors are sequentially expressed. Unlike E75A and MHR3, whose mRNAs are induced when the ecdysteroid titer increases, the expression of MHR4 mRNA occurs transiently at the onset of the decline of ecdysteroid titer followed by betaFTZ-F1 mRNA expression when the ecdysteroid titer becomes low. When day 2 fourth epidermis was exposed to 20-hydroxyecdysone (20E) in vitro, MHR4 mRNA appeared between 12 and 21 h, peaked at 24 h, and then declined. Using the protein synthesis inhibitors cycloheximide and anisomycin both in vivo and in vitro, we found that the MHR4 transcript was directly induced by 20E and required the presence of 20E for its expression. The accumulation of MHR4 mRNA, however, did not occur until a 20E-induced inhibitory protein(s) disappeared. This control of MHR4 expression is unique among the ecdysone-induced transcription factors. When the epidermis was cultured with 20E, betaFTZ-F1 mRNA was not induced until after the removal of 20E as previously found for Drosophila and the silkworm Bombyx mori. The presence of juvenile hormone had no effect on accumulation of either transcript. PMID- 11254364 TI - Characterization of [3H]CGP 12177 binding to beta-adrenergic receptors in intact eel hepatocytes. AB - The aim of this study was to characterize [3H]CGP 12177 (CGP) binding to beta adrenergic receptors in isolated hepatocytes of the European eel (Anguilla anguilla), in which the involvement of cAMP in epinephrine-induced glucose release has been previously observed. Specific binding of CGP was saturable, reversible, and linear as a function of cell number. Analysis of binding data suggested a single class of binding sites, with a Kd of 1.31 nM and a number of approximately 7000 beta-adrenergic receptors per cell. The potency order of specific inhibition of [3H]CGP binding was CGP > propranolol > or = alprenolol >> butoxamine > or = atenolol, while phentolamine and prazosin failed to significantly displace the tracer at concentrations up to 100 microM. The binding kinetics of CGP were closely related to its biological effect. In fact, the drug dose-dependently counteracted the enhancement of intracellular cAMP levels induced by epinephrine in isolated hepatocytes with a Kd of 1.06 nM. Moreover, it antagonized the hormone-induced stimulation of adenylyl cyclase activity in hepatic membranes as well as of glucose release from cells. These data clearly show that beta-adrenergic receptors are coupled to the adenylyl cyclase/cAMP transduction pathway in eel liver. PMID- 11254366 TI - Effects of implantation of hypertrophied androgenic glands on sexual characters and physiology of the reproductive system in the female red claw crayfish, Cherax quadricarinatus. AB - The role of the androgenic gland (AG), an organ unique to male Crustacea, in the development of sex characters and physiology of the reproductive system has not been fully documented in the red claw crayfish, Cherax quadricarinatus. To investigate the role of the AG in this species, the effect of implanting hypertrophied AGs into immature female animals was followed. Of the female animals with AG implants, 91.6% developed male-like propodi, including the red patch characteristic of males of this species. The development of female secondary sex characteristics such as a wider abdomen, a wider endopod, and simple setation was inhibited. At the end of the experiment, the ovaries of the AG-implanted females contained mostly lipid-stage oocytes, with a small number of oocytes at the early yolk stage. The gonadosomatic index of the AG-implanted females was significantly lower than that of the control (sperm duct-implanted or sham-operated) females, which had mature oocytes with a well-defined perinuclear zone and yolk globules. An immunohistochemical test using an antibody developed against a 106-kDa secondary vitellogenic polypeptide showed only slight immunoreactivity in the oocytes of AG-implanted females compared with abundant immunoreactivity in control ovaries. In the polypeptide profile of the high density lipoprotein (HDL) from the hemolymph of AG-implanted females, the 206- and 79-kDa secondary vitellogenesis-specific polypeptides were not found, whereas they were present in the profile of control females. In contrast, the female specific 177-kDa polypeptide was present in the polypeptide profile of hemolymph HDL of both AG-implanted females and control females. It seems therefore that while secondary sex characters were masculinized under the influence of the implanted AG, the process of vitellogenesis was suppressed but not fully eliminated in the AG-implanted females. PMID- 11254365 TI - The distribution of melanin-concentrating hormone in the lamprey brain. AB - In addition to its novel, colour-regulating hormonal role in teleosts, the melanin-concentrating hormone (MCH) serves as a neuromodulatory peptide in all vertebrate brains. In gnathostome vertebrates, it is produced in several neuronal cell groups in the hypothalamus. The present work examines the organisation of the MCH system in the brain of lampreys, which separated from gnathostome vertebrates at an early stage in evolution. In all three lamprey genera examined Petromyzon, Lampetra, and Geotria spp.-MCH perikarya were found in one major anatomical site, the periventricular dorsal hypothalamic nucleus of the posterior hypothalamus. Axons from these cell bodies projected medially into the ventricular cavity, and laterally to the neuropile of the lateral hypothalamus. From here, they extended anteriorly and posteriorly to the fore- and hindbrain. Other fibres extended dorsomedially to the habenular nucleus. In Lampetra, but not in Petromyzon, MCH fibres were seen in the pituitary neurohypophysis, most prominantly above the proximal pars distalis. The hypothalamic region in which the MCH perikarya are found forms part of the paraventricular organ (PVO), which is rich in monoamines and other neuropeptides. The association of MCH neurones with the PVO, which occurs also in many other nonmammalian vertebrates, may reflect the primary location of the MCH system. These MCH neurones were present in ammocoetes, postmetamorphic juveniles, and adults. They were more heavily granulated in adults than in young lampreys but showed no marked change in secretory appearance associated with metamorphosis or experimental osmotic challenge to indicate a role in feeding or osmoregulation. In sexually maturing Lampetra fluviatilis, however, a second group of small MCH neurones became detectable in the telencephalon, suggesting a potential role in reproduction and/or behaviour. PMID- 11254367 TI - Induction of gene expression in sheepshead minnows (Cyprinodon variegatus) treated with 17beta-estradiol, diethylstilbestrol, or ethinylestradiol: the use of mRNA fingerprints as an indicator of gene regulation. AB - The recent interest in hormonally active environmental contaminants has sparked a drive to find sensitive methods to measure their effects on wildlife. A molecular based assay has been developed to measure the induction of gene expression in sheepshead minnows (Cyprinodon variegatus) exposed in vivo to the natural and pharmaceutical estrogens 17beta-estradiol, ethinylestradiol, and diethylstilbestrol. This method used differential display reverse transcriptase polymerase chain reaction assays to compare the expression of individual mRNAs from control and estrogen-exposed fish. Forty-eight differentially expressed cDNAs were isolated by this method, including cDNAs for vitelline envelope proteins and vitellogenin. The mRNA expression patterns for fish injected with a pharmacological dose of estradiol (5 mg/kg) were identical to those obtained in fish receiving constant aqueous exposure to 212 ng estradiol/liter. Further, the cDNA "fingerprint" pattern observed in the estradiol-treated fish also matched that obtained in fish receiving continuous-flow aqueous exposures to 192 ng ethinyl estradiol/liter and a nominal concentration of 200 ng diethylstilbestrol/liter. The results demonstrate a characteristic expression pattern for genes upregulated by exposure to a variety of natural and anthropogenic estrogens and suggest this approach may be valuable to examine the potential effects of environmental contaminants on other endocrine-mediated pathways of reproduction, growth, and development. PMID- 11254368 TI - Electron microscopic evidence for multiple types of secretory vesicles in bovine chromaffin cells. AB - It has been previously shown that the neuron-like chromaffin cells from the bovine adrenal medulla are heterogeneous. Among other differences, the cells also differed in secretory vesicles represented in their cytoplasm. The present study investigates the types of secretory vesicles in bovine chromaffin cells by electron microscopy. Morphometric analysis revealed five types of electron-dense secretory vesicles in chromaffin cells. These were as follows: elementary large catecholamine-storing chromaffin granules of rounded shape, large dense core vesicles of ovoid and rod-like shapes, small dense core vesicles as well as ribosome-coated vesicles of intermediate density. Among the electron-lucent vesicles there were small synaptic-like microvesicles, endocytotic clathrin coated vesicles, growth cone vesicles, and emptied large light core vesicles. The structural and functional backgrounds of different types of secretory vesicles are described, focusing on their formation and potential role. PMID- 11254369 TI - Endocrine correlates of male polymorphism and alternative reproductive tactics in the Azorean rock-pool blenny, Parablennius sanguinolentus parvicornis. AB - In the Azorean rock-pool blenny male sexual polymorphism occurs. Larger and older males (M+ males) fully express male secondary sex characters (SSC), particularly an anal gland that produces a sex pheromone, whereas smaller and younger sexually active males do not express SSC (M- males). Two mating tactic types can be identified among M+ males: nest-holders that establish nests and court females and floaters that move around in the breeding area and try to achieve parasitic fertilizations and/or to take over nests. Two behavioral tactic types can also be identified within M- males: satellites that are associated with particular nests and actively participate in territorial defense (when females go inside the nest to spawn they try to enter to fertilize some of the eggs) and sneakers that do not help nest holders (when spawning occurs they also try to enter the nest to fertilize eggs). It was found that M+ males have significantly higher levels of 11-ketotestosterone (KT), but not testosterone (T), than M- males [M+ male androgen levels (mean +/- SE): total T = 11.6 +/- 3.0 ng ml(-1), total KT = 4.5 +/- 1.1 ng ml(-1); M- male androgen levels (mean +/- SE): total T = 9.6 +/- 1.0 ng ml(-1), total KT = 2.5 +/- 1.1 ng ml(-1)]. There were no differences in plasma T or KT among individuals using different mating tactics within the same male morph; that is, among M+ males, nest-holders did not differ in androgen levels from floaters [nest-holder androgen levels (mean +/- SE): total T = 12.3 +/- 4.4 ng ml(-1), total KT = 4.3 +/- 1.4 ng ml(-1); floater androgen levels (mean +/- SE): total T = 5.9 +/- 0.8 ng ml(-1), total KT = 3.4 +/- 0.3 ng ml(-1)], and among M- males, satellites did not differ in androgen levels from sneakers [satellite androgen levels (mean +/- SE): total T = 7.7 +/- 1.5 ng ml(-1), total KT = 1.3 +/- 0.3 ng ml(-1); sneaker androgen levels (mean +/- SE): total T = 8.3 +/- 1.6 ng ml(-1), total KT = 1.4 +/- 0.3 ng ml(-1)]. Thus, the observed differences appear to be correlated with the expression of different male morphotypes and not with the expression of different behavioral tactics within the morphotype. Androgen levels were not correlated with the behavior activity of nest-holders, except for a negative correlation between KT levels and parental behavior. Furthermore, nest-holder males that succeeded in having females spawn in their nests during the observation period had significantly lower KT levels than unsuccessful males. Since behavioral observations preceded blood sampling in time, it is suggested that these results indicate a negative relationship between KT and parental care, since successful males were parenting when blood samples were collected. Male SSC were better correlated with KT than with T and the use of total blood levels (i.e., free + conjugates) yielded higher correlation coefficients than when only the free fraction of each steroid was considered. Since conjugates are nonactive metabolites of the free androgen they should reflect active free steroids in a previous time. Thus, their incorporation into the hormonal measurements increases the time frame captured, and because steroids are released in a pulsatile way, this time-integrated measure can be more meaningful than the free steroids, which represent a snapshot of the hormone levels at a given point in time. PMID- 11254370 TI - Effects of glucocorticoids on cartilage growth and response to IGF-I in the tilapia (Oreochromis mossambicus). AB - To study the effects of glucocorticoids and IGF-I on the modulation of growth in the tilapia Oreochromis mossambicus, we employed an epiceratobranchial cartilage radioisotope incorporation assay, wherein radiolabeled sulfate and thymidine uptakes are measured in vitro to indicate proteoglycan synthesis and cell proliferation, respectively. Cartilage explants were cultured with cortisol or dexamethasone with or without recombinant bovine insulin-like growth factor-I. Cortisol directly inhibited sulfate uptake at 100 and 1000 ng/mL concentrations in a concentration-dependent manner but inhibited thymidine uptake significantly only at the 1000 ng/mL concentration. Dexamethasone inhibited sulfate and thymidine uptake at concentrations similar to the effective concentrations of cortisol. Cortisol did not inhibit IGF-I stimulation of sulfate uptake at any of the concentrations tested. Furthermore, cortisol did not inhibit thymidine uptake when IGF-I was present in the medium. Cortisol appears to act directly on cartilage and not by interacting with the IGF-I system. However, the physiologically significant role of cortisol is mainly an inhibitory one on cartilage metabolism. The data generally indicate an inhibitory role for glucocorticoids on cartilage growth but an inability to counter the stimulation of sulfate uptake by IGF-I. PMID- 11254371 TI - Effects of estrogens in vitro and in vivo on cartilage growth in the tilapia (Oreochromis mossambicus). AB - To study the effects of estrogens on cartilage growth in the tilapia Oreochromis mossambicus, an epiceratobranchial cartilage radioisotope incorporation assay was employed to measure proteoglycan synthesis and prechondrocyte proliferation by incorporation of radiolabeled sulfate and thymidine, respectively. Cartilage explants were cultured with estrogens with or without recombinant bovine insulin like growth factor-I (IGF-I). In vitro experiments using the natural teleost estrogen, 17beta-estradiol (E2), showed a trend toward inhibition of sulfate incorporation and an inhibition of thymidine incorporation at higher doses (10 micrograms/ml), but not at physiological levels. E2 also showed a trend toward inhibition of sulfate and thymidine incorporation in the presence of IGF-I. Similar results were found with other estrogenic compounds in vitro: ethinylestradiol, diethylstilbestrol (DES), genistein, and nonylphenol. Ethinylestradiol inhibited sulfate and thymidine incorporation at 1000 ng/ml in the presence of IGF-I. DES inhibited thymidine incorporation at 1000 ng/ml in untreated or IGF-I-exposed cartilage. Genistein inhibited sulfate incorporation at 100 micrograms/ml in IGF-I-exposed cartilage and inhibited thymidine uptake at 1, 10, and 100 micrograms/ml in untreated and IGF-I-exposed cartilage. Nonylphenol inhibited sulfate uptake at 100 microM in untreated and IGF-I-exposed cartilage. Nonylphenol alone at 10 and 100 microM inhibited thymidine uptake. In IGF-I-exposed cartilage nonylphenol inhibited thymidine uptake at 100 microM. Fish receiving estrogen injections (E2 or DES) in vivo at a concentration of 2 micrograms/g body weight showed increased sulfate incorporation by cartilage in vitro. Stimulation in vivo by estrogens, in contrast to the inhibition by high doses in vitro, may be a result of the influence of estrogen on pituitary growth hormone release. PMID- 11254372 TI - Follicle-stimulating hormone regulates steroidogenic enzymes in cultured cells of the chick embryo ovary. AB - This investigation addresses the potential regulation of enzymes involved in the biosynthesis of steroid hormones during early stages of gonadal development by follicle-stimulating hormone (FSH). Gonadal cells of 10-day-old chick embryo and cells of the left ovary of 18-day-old chick embryo were cultured for 60 h in a defined medium with or without the addition of FSH (2.0 IU/ml). At the end of the culture, cells were recovered and evaluated by biotransformation of tritiated steroid precursors and mRNA levels were evaluated by RT-PCR. The production of estrone from androstenedione was increased in the FSH-treated cells, both human FSH (hFSH) and recombinant human FSH (rhFSH), indicating a stimulatory effect on aromatase (P450arom). Similarly, the intensity of the band corresponding to P450arom mRNA was higher in hFSH and rhFSH than in control and chorionic gonadotropin (hCG) groups. The P450arom stimulation was observed in the ovary of 10- and 18-day-old chick embryo. The transformation of dehydroepiandrosterone to androstenedione was taken as evidence of 3beta-hydroxysteroid dehydrogenase function. This enzyme was stimulated in the cultured ovarian cells of 18-day-old chick embryos treated with hFSH and rhFSH compared with controls. The production of pregnenolone in the mitocondrial fraction of 18-day-old chick embryo ovary was increased when cultured with hFSH and rhFSH. This observation together with the increase in the band intensity corresponding to mRNA of P450 cholesterol side chain cleavage indicates stimulation by FSH treatment; hCG produced a similar effect. Somatic cells of the medullary cords are proposed to be FSH target cells in the ovary of the chick embryo. PMID- 11254373 TI - The effects of a single long photoperiod on induction and dissipation of reproductive photorefractoriness in European starlings. AB - In many birds, long photoperiods stimulate gonadal maturation but also cause photorefractoriness, leading to gonadal regression. While much is known about the neuroendocrinology of photostimulation, little is known about photorefractoriness, partly due to lack of an experimental model. This study aimed to develop a model to test whether a single long photoperiod (LP) initiates the mechanism leading to photorefractoriness. It made use of the fact that in castrated European starlings, luteinizing hormone (LH) is low in photorefractory birds and high in photosensitive birds. In the first experiment, groups of castrated photorefractory birds were transferred from a long to a short photoperiod and then exposed to one LP every 5, 10, 20, 30, 40, or 60 days. In birds exposed to one LP every 5 days, LH stayed low. In birds exposed to one LP every 10 days, each LP caused a pulse in LH, but mean LH remained low. One LP every 20 days increased LH 6 days later, followed by a decrease at 11 days and then a further increase, so that overall, LH increased slowly. In birds exposed to one LP every 30, 40, or 60 days, LH increased at the same rate as in short photoperiod controls. Each LP did not cause a significant increase in LH, but did cause a decrease. A second experiment examined the changes in LH following a LP in more detail. Castrated starlings had been exposed to one LP every 14 days for 16 weeks. On the day of the final LP, LH values were midway between photorefractory and photosensitive values. The LP caused an increase in LH from the second day to a peak after 7 days. Thereafter, LH declined to initial values after 14 days, followed, in the absence of further LP, by a second increase to photosensitive values. These results suggest that the mechanisms causing photostimulation and photorefractoriness are both initiated during the first long photoperiod. That gonadal maturation precedes regression must reflect the relative rates at which the two processes reach completion. PMID- 11254374 TI - Steroid profiles in cultured female jundia, the Siluridae Rhamdia quelen (Quoy and Gaimard, Pisces Teleostei), during the first reproductive cycle. AB - The jundia Rhamdia quelen (Quoy and Gaimard) is a teleost species from the Siluridae family and is an important species for aquaculture in temperate and subtropical climates. Gonad and blood tissue samples were taken from cultured jundia females between 1998 and 1999. Plasma concentrations of 17beta-estradiol (E(2)), testosterone (T), 11-ketotestosterone (11-KT), 17-hydroxy-4-pregnene-3,20 dione (17-P), 17,20beta-dihydroxy-4-pregnen-3-one (17,20beta-P), and 17,20beta,21 trihydroxy-4-pregnen-3-one (20beta-S) were measured by radioimmunoassay and potential correlations with the stage of oogenesis and sexual maturation examined. During the experimental period two spawning episodes were observed. Plasma concentrations of E(2) increased progressively during oocyte development, simultaneously with the appearance of yolk vesicles and increasing amounts of deposited yolk. In female jundia, the T peak occurred in October and was coincident with the peak in gonadosomatic index. Two distinct peaks of progestogens were detected, corresponding to the two spawning episodes, suggesting that one or more of these steroids might act as the "maturational inducing steroid" in jundia. Unusually large amounts of 11-KT were also measured in the plasma of mature jundia females. The identity of 11-KT was confirmed by thin-layer chromatography. Although the profiles of the other steroids are compatible with the roles proposed for the action of these hormones in other teleosts, the role of 11-KT, normally found only in males, is unknown. PMID- 11254375 TI - Pituitary and interrenal function in gilthead sea bream (Sparus aurata L., Teleostei) after handling and confinement stress. AB - Dynamics of adrenocorticotropin (ACTH), alpha melanocyte-stimulating hormone (alpha-MSH), N-acetylated-beta-endorphin (N-ac-beta-END), cortisol, and growth hormone (GH) were investigated in gilthead sea bream (Sparus aurata) stressed by handling plus confinement. As indices of the secondary stress response, plasma levels of glucose, lactate, and plasma ions were monitored. Within 1 h, plasma cortisol and ACTH levels increased above the control values but GH levels decreased. Subsequently, at 24 h cortisol and ACTH levels had declined, but were still higher than in controls, whereas GH levels had recovered after 4 h. Regarding the melanotrope peptides, there were no differences in plasma levels of alpha-MSH and N-ac-beta-END, but pituitary stores of these peptides were severely depleted already after 1 h, as were ACTH stores. Pituitary contents of proopiomelanocortin (POMC)-derived hormones did not show significant differences from 72 h onward. Therefore, the results indicate that both handling and confinement affected the corticotropes of the pars distalis and the melanotropes of the neurointermediate lobe but at different magnitudes. The possible involvement of corticotropin-releasing hormone (CRH) in the regulation of pituitary POMC-producing cell types under these conditions was indicated by the in vitro dose-dependent effect of the peptide on release of ACTH, alpha-MSH, and N-ac-beta-END. The corticocotropes appeared more responsive, and approximately 10 fold more sensitive, to CRH compared with the melanotropes. The ACTH-releasing potency of 1 nM CRH was inhibited 75% following pretreatment of the whole pituitary gland with 400 nM of the CRH antagonist alpha-helical CRH(9-41). PMID- 11254376 TI - Non-clonability correlates with genomic instability: a case study of a unique DNA region. AB - Instability of eukaryotic DNA in constructs propagated in prokaryotic hosts is a frequently observed phenomenon. With the exception of a very high A+T-content and the presence of multiple repetitions, no general rule at the basis of this phenomenon is actually known. The intergenic spacer located between the pi and alpha(D) chicken alpha-type globin genes is frequently deleted from recombinant phages and plasmids. Here we have cloned this DNA fragment using a specially designed bacterial strain (SURE competent cells, Stratogene). Comparative analysis of DNA of recombinant clones bearing deletions and clones containing the intact genomic DNA fragment has revealed two important DNA sequence motifs that contribute to the unclonability of eukaryotic DNA in prokaryotic cells. First, the similarity to bacterial transposons (i.e. the presence of repeats flanking a several kilobase DNA fragment) may cause the loss of the fragment during propagation of the recombinant DNA in E. coli. Second, a high content of rotationally correlated kinkable elements (TG*CA steps) may result in non clonability of the DNA sequence. Interestingly, the latter type of "unclonable" DNA sequence motifs identified in the globin gene domain is unstable (frequently rearranged) also in the eukaryotic chromosome resulting in a local polymorphism. In the chicken domain of alpha globin genes this unstable DNA sequence seems to be partially protected by interaction with nuclear matrix proteins. PMID- 11254378 TI - Ab initio phasing of high-symmetry macromolecular complexes: successful phasing of authentic poliovirus data to 3.0 A resolution. AB - A genetic algorithm-based computational method for the ab initio phasing of diffraction data from crystals of symmetric macromolecular structures, such as icosahedral viruses, has been implemented and applied to authentic data from the P1/Mahoney strain of poliovirus. Using only single-wavelength native diffraction data, the method is shown to be able to generate correct phases, and thus electron density, to 3.0 A resolution. Beginning with no advance knowledge of the shape of the virus and only approximate knowledge of its size, the method uses a genetic algorithm to determine coarse, low-resolution (here, 20.5 A) models of the virus that obey the known non-crystallographic symmetry (NCS) constraints. The best scoring of these models are subjected to refinement and NCS-averaging, with subsequent phase extension to high resolution (3.0 A). Initial difficulties in phase extension were overcome by measuring and including all low-resolution terms in the transform. With the low-resolution data included, the method was successful in generating essentially correct phases and electron density to 6.0 A in every one of ten trials from different models identified by the genetic algorithm. Retrospective analysis revealed that these correct high-resolution solutions converged from a range of significantly different low-resolution phase sets (average differences of 59.7 degrees below 24 A). This method represents an efficient way to determine phases for icosahedral viruses, and has the advantage of producing phases free from model bias. It is expected that the method can be extended to other protein systems with high NCS. PMID- 11254377 TI - A Mutation in the C-terminal domain of the RNA polymerase alpha subunit that destabilizes the open complexes formed at the phage phi 29 late A3 promoter. AB - Regulatory protein p4 from Bacillus subtilis phage phi29 activates the viral late A3 promoter mainly by stabilizing the binding of RNA polymerase (RNAP) to it as a closed complex. This requires an interaction between protein p4 residue Arg120 and the C-terminal domain (CTD) of the RNAP alpha subunit. Several acidic residues of the alpha-CTD, considered as plausible targets for p4 residue Arg120, were individually changed into alanine. In addition, a truncated alpha subunit lacking the last four residues, two of which are acidic, was obtained. The modified alpha subunits were purified and reconstituted into RNAP holoenzyme in vitro. Protein p4 was found to be unable to activate the late A3 promoter when residue Glu297 of the alpha subunit was changed to Ala, a modification that did not impair transcription from several other promoters. Interestingly, protein p4 could stabilize the modified RNAP at the A3 promoter as a closed complex, although the open complexes formed were unstable and did not proceed to elongation complexes. Our results indicate that the change of the alpha residue Glu297 into Ala destabilizes the open complexes formed at this promoter, but not at other promoters. Considered in the context of earlier findings indicating that the RNAP alpha-CTD may participate in the transition from closed to intermediate complexes at some other promoters, the new results expand and clarify our view of its role in transcription initiation. PMID- 11254379 TI - A-like guanine-guanine stacking in the aqueous DNA duplex of d(GGGGCCCC). AB - We have used CD spectroscopy, NMR spectroscopy and unrestrained molecular dynamics to study conformational properties of a DNA duplex formed by the self complementary octamer d(GGGGCCCC). Its unusual CD spectrum contains features indicating A-like stacking of half of the bases, whereas the other half stack in a B-like fashion. Unrestrained molecular dynamics simulations converged to a stable B-like double-helix of d(GGGGCCCC). However, the double-helix contained a central hole whose size was half of that occurring in structure A. In the canonical structure B, the hole does not exist at all because the base-pairs cross the double-helix centre. The cytosine bases were stacked in the duplex of d(GGGGCCCC) as in structure B, while stacking of the guanine bases displayed features characteristic for structure A. NMR spectroscopy revealed that the A like guanine-guanine stacking was accompanied by an increased tendency of the deoxyribose rings attached to the guanine bases to be puckered in an A-like fashion. Otherwise, the duplex of d(GGGGCCCC) showed no clash, no bend and no other significant deviation from structure B. The present analysis demonstrates a remarkable propensity of the guanine runs to stack in an A-like fashion even within the B-DNA framework. This property explains why the oligo(dG). oligo(dC) tracts switch into structure A so easily. Secondly, this property may influence replication, because structure A is replicated more faithfully than structure B. Thirdly, the oligo(dG) runs might have played an important role in early evolution, when DNA took on functions that originally evolved on RNA. Fourthly, the present study extends the vocabulary of DNA secondary structures by the heteronomous duplex of d(GGGGCCCC) in which the B-like strand of oligo(dC) is bound to the A-like strand of oligo(dG). PMID- 11254380 TI - Relationship of DNA structure to internal dynamics: correlation of helical parameters from NOE-based NMR solution structures of d(GCGTACGC)(2) and d(CGCTAGCG)(2) with (13)C order parameters implies conformational coupling in dinucleotide units. AB - The coupling between the conformational properties of double-stranded DNA and its internal dynamics has been examined. The solution structures of the isomeric DNA oligomers d(GCGTACGC)(2) (UM) and d(CGCTAGCG)(2) (CTSYM) were determined with (1)H NMR spectroscopy by utilizing distance restraints from total relaxation matrix analysis of NOESY cross-peak intensities in restrained molecular dynamics calculations. The root-mean-square deviation of the coordinates for the ensemble of structures was 0.13 A for UM and 0.49 A for CTSYM, with crystallographic equivalent R(c)=0.41 and 0.39 and sixth-root residual R(x)=0.11 and 0.10 for UM and CTSYM, respectively. Both UM and CTSYM are B-form with straight helical axes and show sequence-dependent variations in conformation. The internal dynamics of UM and CTSYM were previously determined by analysis of (13)C relaxation parameters in the context of the Lipari & Szabo model-free formalism. Helical parameters for the two DNA oligomers were examined for linear correlations with the order parameters (S(2)) of groups of (13)C spins in base-pairs and dinucleotide units of UM and CTSYM. Correlations were found for six interstrand base-pair parameters tip, y-displacement, inclination, buckle and stretch with various combinations of S(2) for atoms in Watson-Crick base-pairs and for two inter-base-pair parameters, rise and roll with various combinations of S(2) for atoms in dinucleotides. The correlations for the interstrand base-pair helical parameters indicate that the conformations of the deoxyribose residues of each strand are dynamically coupled. Also, the inter-base-pair separation has a profound effect on the local internal motions available to the DNA, supporting the idea that rise is a principal degree of freedom for DNA conformational variability. The correlations indicate collective atomic motions of spins that may represent specific motional modes in DNA, and that base sequence has a predictable effect on the relative order of groups of spins both in the bases and in the deoxyribose ring of the DNA backbone. These observations suggest that an important functional outcome of DNA base sequence is the modulation of both the conformation and dynamic behavior of the DNA backbone. PMID- 11254381 TI - Structural biochemistry of a type 2 RNase H: RNA primer recognition and removal during DNA replication. AB - DNA replication and cellular survival requires efficient removal of RNA primers during lagging strand DNA synthesis. In eukaryotes, RNA primer removal is initiated by type 2 RNase H, which specifically cleaves the RNA portion of an RNA DNA/DNA hybrid duplex. This conserved type 2 RNase H family of replicative enzymes shares little sequence similarity with the well-characterized prokaryotic type 1 RNase H enzymes, yet both possess similar enzymatic properties. Crystal structures and structure-based mutational analysis of RNase HII from Archaeoglobus fulgidus, both with and without a bound metal ion, identify the active site for type 2 RNase H enzymes that provides the general nuclease activity necessary for catalysis. The two-domain architecture of type 2 RNase H creates a positively charged binding groove and links the unique C-terminal helix loop-helix cap domain to the active site catalytic domain. This architectural arrangement apparently couples directional A-form duplex binding, by a hydrogen bonding Arg-Lys phosphate ruler motif, to substrate-discrimination, by a tyrosine finger motif, thereby providing substrate-specific catalytic activity. Combined kinetic and mutational analyses of structurally implicated substrate binding residues validate this binding mode. These structural and mutational results together suggest a molecular mechanism for type 2 RNase H enzymes for the specific recognition and cleavage of RNA in the RNA-DNA junction within hybrid duplexes, which reconciles the broad substrate binding affinity with the catalytic specificity observed in biochemical assays. In combination with a recent independent structural analysis, these results furthermore identify testable molecular hypotheses for the activity and function of the type 2 RNase H family of enzymes, including structural complementarity, substrate-mediated conformational changes and coordination with subsequent FEN-1 activity. PMID- 11254382 TI - Effects of ligand binding on the association properties and conformation in solution of retinoic acid receptors RXR and RAR. AB - In higher eukaryotes, vitamin A derived metabolites such as 9-cis and all-trans retinoic acid (RA), are involved in the regulation of several essential physiological processes. Their pleiotropic physiological effects are mediated through direct binding to cognate nuclear receptors RXRs and RARs that act as regulated transcription factors belonging to the superfamily of nuclear hormone receptors. Hormone binding to the structurally conserved ligand-binding domain (LBD) of these receptors triggers a conformational change that principally affects the conserved C-terminal transactivation helix H12 involved in transcriptional activation. We report an extensive biophysical solution study of RAR alpha, RXR alpha LBDs and their corresponding RXR alpha/RAR alpha LBD heterodimers combining analytical ultracentrifugation (AUC), small-angle X-ray and neutron scattering (SAXS and SANS) and ab initio three-dimensional shape reconstruction at low resolution. We show that the crystal structures of RXRs and RARs LBDs correlate well with the average conformations observed in solution. Furthermore we demonstrate the effects of 9-cisRA and all-transRA binding on the association properties and conformations of RXR alpha and RAR alpha LBDs in solution. The present study shows that in solution RAR alpha LBD behaves as a monomer in both unliganded and liganded forms. It confirms the existence in solution of a ligand-induced conformational change towards a more compact form of the LBD. It also confirms the stability of the predicted RXR alpha/RAR alpha LBD heterodimers in solution. SAS measurements performed on three different types of RXR alpha/RAR alpha LBD heterodimers (apo/apo, apo/holo and holo/holo) with respect to their ligand-binding site occupancy show the existence of three conformational states depending on the progressive binding of RA stereoisomers on RAR alpha and RXR alpha LBD subunits in the heterodimeric context. These results suggest that the subunits are structurally independent within the heterodimers. Our study also underlines the particular behaviour of RXR alpha LBD. In solution unliganded RXR alpha LBD is observed as two species that are unambiguously identified as homotetramers and homodimers. Molecular modelling combined with SAS data analysis allows us to propose a structural model for this autorepressed apo tetramer. In contrast to the monomeric state observed in the crystal structure, our data show that in solution active holo-RXR alpha LBD bound to 9-cisRA is a homodimer regardless of the protein concentration. This study demonstrates the crucial role of ligands in the regulation of homodimeric versus heterodimeric association state of RXR in the NR signalling pathways. PMID- 11254383 TI - Crystal structure of the stromelysin-3 (MMP-11) catalytic domain complexed with a phosphinic inhibitor mimicking the transition-state. AB - Stromelysin-3 (ST3) is a matrix metalloproteinase (MMP-11) whose proteolytic activity plays an important role in tumorigenicity enhancement. In breast cancer, ST3 is a bad prognosis marker: its expression is associated with a poor clinical outcome. This enzyme therefore represents an attractive therapeutic target. The topology of matrix metalloproteinases (MMPs) is remarkably well conserved, making the design of highly specific inhibitors difficult. The major difference between MMPs lies in the S(1)' subsite, a well-defined hydrophobic pocket of variable depth. The present crystal structure, the first 3D-structure of the ST3 catalytic domain in interaction with a phosphinic inhibitor mimicking a (d, l) peptide, clearly demonstrates that its S(1)' pocket corresponds to a tunnel running through the enzyme. This open channel is filled by the inhibitor P(1)' group which adopts a constrained conformation to fit this pocket, together with two water molecules interacting with the ST3-specific residue Gln215. These observations provide clues for the design of more specific inhibitors and show how ST3 can accommodate a phosphinic inhibitor mimicking a (d, l) peptide. The presence of a water molecule interacting with one oxygen atom of the inhibitor phosphinyl group and the proline residue of the Met-turn suggests how the intermediate formed during proteolysis may be stabilized. Furthermore, the hydrogen bond distance observed between the methyl of the phosphinic group and the carbonyl group of Ala182 mimics the interaction between this carbonyl group and the amide group of the cleaved peptidic bond. Our crystal structure provides a good model to study the MMPs mechanism of proteolysis. PMID- 11254384 TI - The high-resolution X-ray crystallographic structure of the ferritin (EcFtnA) of Escherichia coli; comparison with human H ferritin (HuHF) and the structures of the Fe(3+) and Zn(2+) derivatives. AB - The high-resolution structure of the non-haem ferritin from Escherichia coli (EcFtnA) is presented together with those of its Fe(3+) and Zn(2+) derivatives, this being the first high-resolution X-ray analysis of the iron centres in any ferritin. The binding of both metals is accompanied by small changes in the amino acid ligand positions. Mean Fe(A)(3+)-Fe(B)(3+) and Zn(A)(2+)-Zn(B)(2+) distances are 3.24 A and 3.43 A, respectively. In both derivatives, metal ions at sites A and B are bridged by a glutamate side-chain (Glu50) in a syn-syn conformation. The Fe(3+) derivative alone shows a third metal site (Fe( C)( 3+)) joined to Fe(B)(3+) by a long anti-anti bidentate bridge through Glu130 (mean Fe(B)(3+) Fe(C)(3+) distance 5.79 A). The third metal site is unique to the non-haem bacterial ferritins. The dinuclear site lies at the inner end of a hydrophobic channel connecting it to the outside surface of the protein shell, which may provide access for dioxygen and possibly for metal ions shielded by water. Models representing the possible binding mode of dioxygen to the dinuclear Fe(3+) pair suggest that a gauche micro-1,2 mode may be preferred stereochemically. Like those of other ferritins, the 24 subunits of EcFtnA are folded as four-helix bundles that assemble into hollow shells and both metals bind at dinuclear centres in the middle of the bundles. The structural similarity of EcFtnA to the human H chain ferritin (HuHF) is remarkable (r.m.s. deviation of main-chain atoms 0.66 A) given the low amino acid sequence identity (22 %). Many of the conserved residues are clustered at the dinuclear centre but there is very little conservation of residues making inter-subunit interactions. PMID- 11254385 TI - Structure and functionality of a designed p53 dimer. AB - P53 is a homotetrameric tumor suppressor protein involved in transcriptional control of genes that regulate cell proliferation and death. In order to probe the role that oligomerization plays in this capacity, we have previously designed and characterized a series of p53 proteins with altered oligomeric states through hydrophilc substitution of residues Met340 or Leu344 in the normally tetrameric oligomerization domain. Although such mutations have little effect on the overall secondary structural content of the oligomerization domain, both solubility and the resistance to thermal denaturation are substantially reduced relative to that of the wild-type domain. Here, we report the design and characterization of a double-mutant p53 with alterations of residues at positions Met340 and Leu344. The double-mutations Met340Glu/Leu344Lys and Met340Gln/Leu344Arg resulted in distinct dimeric forms of the protein. Furthermore, we have verified by NMR structure determination that the double-mutant Met340Gln/Leu344Arg is essentially a "half-tetramer". Analysis of the in vivo activities of full-length p53 oligomeric mutants reveals that while cell-cycle arrest requires tetrameric p53, transcriptional transactivation activity of monomers and dimers retain roughly background and half of the wild-type activity, respectively. PMID- 11254386 TI - The energetics of the interaction of BamHI endonuclease with its recognition site GGATCC. AB - The interaction of BamHI endonuclease with DNA has been studied crystallographically, but has not been characterized rigorously in solution. The enzyme binds in solution as a homodimer to its recognition site GGATCC. Only six base-pairs are directly recognized, but binding affinity (in the absence of the catalytic cofactor Mg(2+)) increases 5400-fold as oligonucleotide length increases from 10 to 14 bp. Binding is modulated by sequence context outside the recognition site, varying about 30-fold from the bes t (GTG or TAT) to the worst (CGG) flanking triplets. BamHI, EcoRI and EcoRV endonucleases all have different context preferences, suggesting that context affects binding by influencing the free energy levels of the complexes rather than that of the free DNA. Ethylation interference footprinting in the absence of divalent metal shows a localized and symmetrical pattern of phosphate contacts, with strong contacts at NpNpNpGGApTCC. In the presence of Mg(2+), first-order cleavage rate constants are identical in the two GGA half-sites, are the same for the two nicked intermediates and are unaffected by substrate length in the range 10-24 bp. DNA binding is strongly enhanced by mutations D94N, E111A or E113K, by binding of Ca(2+) at the active site, or by deletion of the scissile phosphate GpGATCC, indicating that a cluster of negative charges at the catalytic site contributes at least 3-4 kcal/mol of unfavorable binding free energy. This electrostatic repulsion destabilizes the enzyme-DNA complex and favors metal ion binding and progression to the transition state for cleavage. PMID- 11254387 TI - Thermodynamics of trimer-of-hairpins formation by the SIV gp41 envelope protein. AB - The gp41 envelope protein mediates the entry of primate immunodeficiency viruses into target cells by promoting the fusion of viral and cellular membranes. The structure of the gp41 ectodomain core represents a trimer of identical helical hairpins in which a central trimeric coiled-coil made up of three amino-terminal helices is wrapped in an outer layer of three antiparallel carboxyl-terminal helices. Triggering formation of this fusion-active gp41 conformation appears to cause close membrane apposition and thus overcome the activation energy barrier for lipid bilayer fusion. We present a detailed description of the folding thermodynamics of the simian immunodeficiency virus (SIV) gp41 core by using a recombinant trimeric N34(L6)C28 model. Differential scanning calorimetry and spectroscopic experiments on denaturant-induced and thermal unfolding indicate that the free energy of association of three 68 residue N34(L6)C28 peptides to a trimer-of-hairpins is 76 kJ mol(-1) at pH 7.0 and 25 degrees C in physiological buffer. The associated enthalpy change, Delta H(unf), is 177 kJ mol(-1), while the entropy of unfolding, Delta S(unf), is 0.32 kJ K(-1) mol(-1). The temperature of maximal stability is close to 20 degrees C. The unfolding heat capacity increment is approximately 9 kJ K(-1) mol(-1) (approximately 45 J K(-1) mol residue(-1)), which is lower than expected for unfolding of the trimer to an extended and fully hydrated polypeptide chain. Replacement by isoleucine of the polar residues Thr582 or Thr586 buried in the N-terminal trimeric coiled-coil interface leads to very strong stabilization of the trimer-of-hairpins, 30-35 kJ mol(-1). Single-point mutations in the central coiled-coil thus strongly stabilize the gp41 core structure. These thermodynamic characteristics may be important for the refolding of the gp41 envelope protein into its fusion-active conformation during membrane fusion. PMID- 11254388 TI - Core and surface mutations affect folding kinetics, stability and cooperativity in IL-1 beta: does alteration in buried water play a role? AB - Interleukin-1 beta (IL-1 beta) is a cytokine and a member of the beta-trefoil superfamily of protein structures. An interesting feature in the folding of IL-1 beta, shared with some other members of the same topological family, is the existence of a slow step in folding to the native conformation from a discrete intermediate. Wanting to probe the nature of this slow step in the folding of WT IL-1 beta (tau(1)=45 seconds), we made ten sequence variants of IL-1 beta (L10A, T9Q, T9G, C8S, C8A, N7G, N7D, L6A, R4P, and R4Q), where all mutations are located along strand 1. This strand is not protected from hydrogen exchange until late in folding. Most of the mutations showed little effect on the kinetics of folding for IL-1 beta. However, C8 is clearly involved in both the late and the early steps in folding, while sequence variants at L10 and L6 affect only late events in folding. The value of the slowest relaxation time, tau(1), which is associated with the rate of native protein formation, increased for the refolding of C8S, while C8A, L6A, and L10A showed smaller but systematic increases in the value of tau(1.)For both C8S and C8A, the value of the step associated with formation of the intermediate, tau(2), was independent of denaturant concentration. In addition, mutations in the hydrophobic core (L10A, C8A, C8S, and L6A) and, surprisingly, along the surface (T9G, T9Q, and N7G) alter the stability. The most destabilizing mutations show changes in equilibrium unfolding cooperativity, which is atypical for destabilizing mutations in IL-1 beta. Crystallographic studies indicate that mutations along strand 1 may alter the number of ordered water molecules within the core. Thus, side-chain replacement in this region can disrupt essential main-chain interactions mediated by ordered water contacts in a highly cooperative network of hydrogen bonding. PMID- 11254389 TI - An interfacial mechanism and a class of inhibitors inferred from two crystal structures of the Mycobacterium tuberculosis 30 kDa major secretory protein (Antigen 85B), a mycolyl transferase. AB - The Mycobacterium tuberculosis 30 kDa major secretory protein (antigen 85B) is the most abundant protein exported by M. tuberculosis, as well as a potent immunoprotective antigen and a leading drug target. A mycolyl transferase of 285 residues, it is closely related to two other mycolyl transferases, each of molecular mass 32 kDa: antigen 85A and antigen 85C. All three catalyze transfer of the fatty acid mycolate from one trehalose monomycolate to another, resulting in trehalose dimycolate and free trehalose, thus helping to build the bacterial cell wall. We have determined two crystal structures of M. tuberculosis antigen 85B (ag85B), initially by molecular replacement using antigen 85C as a probe. The apo ag85B model is refined against 1.8 A data, to an R-factor of 0.196 (R(free) is 0.276), and includes all residues except the N-terminal Phe. The active site immobilizes a molecule of the cryoprotectant 2-methyl-2,4-pentanediol. Crystal growth with addition of trehalose resulted in a second ag85B crystal structure (1.9 A resolution; R-factor is 0.195; R(free) is 0.285). Trehalose binds in two sites at opposite ends of the active-site cleft. In our proposed mechanism model, the trehalose at the active site Ser126 represents the trehalose liberated by temporary esterification of Ser126, while the other trehalose represents the incoming trehalose monomycolate just prior to swinging over to the first trehalose site to displace the mycolate from its serine ester. Our proposed interfacial mechanism minimizes aqueous exposure of the apolar mycolates. Based on the trehalose-bound structure, we suggest a new class of antituberculous drugs, made by connecting two trehalose molecules by an amphipathic linker. PMID- 11254390 TI - Predicting the functional consequences of non-synonymous single nucleotide polymorphisms: structure-based assessment of amino acid variation. AB - We have developed a formalism and a computational method for analyzing the potential functional consequences of non-synonymous single nucleotide polymorphisms. Our approach uses a structural model and phylogenetic information to derive a selection of structure and sequence-based features serving as indicators of an amino acid polymorphim's effect on function. The feature values can be integrated into a probabilistic assessment of whether an amino acid polymorphism will affect the function or stability of a target protein. The method has been validated with data sets of unbiased mutations in the lac repressor and lysoyzyme. Applying our methodology to recent surveys of genetic variation in the coding regions of clinically important genes, we estimate that approximately 26-32 % of the natural non-synonymous single nucleotide polymorphisms have effects on function. This estimate suggests that a typical person will have about 6240-12,800 heterozygous loci that encode proteins with functional variation due to natural amino acid polymorphism. PMID- 11254391 TI - Structures of bovine glutamate dehydrogenase complexes elucidate the mechanism of purine regulation. AB - Glutamate dehydrogenase is found in all organisms and catalyses the oxidative deamination of l-glutamate to 2-oxoglutarate. However, only animal GDH utilizes both NAD(H) or NADP(H) with comparable efficacy and exhibits a complex pattern of allosteric inhibition by a wide variety of small molecules. The major allosteric inhibitors are GTP and NADH and the two main allosteric activators are ADP and NAD(+). The structures presented here have refined and modified the previous structural model of allosteric regulation inferred from the original boGDH.NADH.GLU.GTP complex. The boGDH.NAD(+).alpha-KG complex structure clearly demonstrates that the second coenzyme-binding site lies directly under the "pivot helix" of the NAD(+) binding domain. In this complex, phosphates are observed to occupy the inhibitory GTP site and may be responsible for the previously observed structural stabilization by polyanions. The boGDH.NADPH.GLU.GTP complex shows the location of the additional phosphate on the active site coenzyme molecule and the GTP molecule bound to the GTP inhibitory site. As expected, since NADPH does not bind well to the second coenzyme site, no evidence of a bound molecule is observed at the second coenzyme site under the pivot helix. Therefore, these results suggest that the inhibitory GTP site is as previously identified. However, ADP, NAD(+), and NADH all bind under the pivot helix, but a second GTP molecule does not. Kinetic analysis of a hyperinsulinism/hyperammonemia mutant strongly suggests that ATP can inhibit the reaction by binding to the GTP site. Finally, the fact that NADH, NAD(+), and ADP all bind to the same site requires a re-analysis of the previous models for NADH inhibition. PMID- 11254392 TI - Pairwise sequence alignment below the twilight zone. AB - Improved sequence alignment at low pairwise identity is important for identifying potential remote homologues in database searches and for obtaining accurate alignments as a prelude to modeling structures by homology. Our work is motivated by two observations: structural data provide superior training examples for developing techniques to improve the alignment of remote homologues; and general substitution patterns for remote homologues differ from those of closely related proteins. We introduce a new set of amino acid residue interchange matrices built from structural superposition data. These matrices exploit known structural homology as a means of characterizing the effect evolution has on residue substitution profiles. Given their origin, it is not surprising that the individual residue-residue interchange frequencies are chemically sensible. The structural interchange matrices show a significant increase both in pairwise alignment accuracy and in functional annotation/fold recognition accuracy across distantly related sequences. We demonstrate improved pairwise alignment by using superpositions of homologous domains extracted from a structural database as a gold standard and go on to show an increase in fold recognition accuracy using a database of homologous fold families. This was applied to the unassigned open reading frames from the genome of Helicobacter pylori to identify five matches, two of which are not represented by new annotations in the sequence databases. In addition, we describe a new cyclic permutation strategy to identify distant homologues that experienced gene duplication and subsequent deletions. Using this method, we have identified a potential homologue to one additional previously unassigned open reading frame from the H. pylori genome. PMID- 11254393 TI - In memoriam. Edward H. Bloch (1914-2000). Pioneer in the microscopic study of and the microcirulation of living tissues and organs. PMID- 11254394 TI - Hyperthermic pretreatment decreases microvascular protein leakage and attenuates hypotension in anaphylactic shock in rats. AB - Systemic anaphylaxis is a life-threatening allergic reaction and its pathologic conditions, such as edema, bronchospasm, and hypotension, have been attributed to release of vasoactive mediators. Heat shock protein (HSP) is known to play a protective role in living cells under various stresses. In these studies, we investigated the protective role of heat shock response in anaphylactic shock, focusing on changes of blood pressure (BP) and vascular permeability. Adult sensitized rats were injected intravenously with Evans blue (EB) and challenged with bovine serum albumin (BSA). The rats were treated with whole-body hyperthermia at 41.5 +/- 0.5 degrees for 15 min 24 h before BSA challenge. Vascular protein leakage in tissues was analyzed with the EB technique. The results showed that BSA challenge induced EB extravasation in all sensitized rats. EB values (EB/tissue; microg/g) in heart and lung (112.3 +/- 41 and 244.4 +/- 90.6; mean +/- SD; n = 6) in the nonheated rats were significantly higher than those (33.4 +/- 23.3 and 103.4 +/- 63.9; n = 9) in the heated rats (P < 0.05). The results showed that BSA challenge caused BP to fall drastically in the sensitized rats. BP in the heated rats was significantly higher than BP in the nonheated rats from 4 to 15 min during anaphylactic shock (P < 0.001). Inducible HSP72 appeared overexpressed in heart, lung, and liver tissue in the heated rats tested by Western immunoblotting. The results indicate that reduction of increased protein leakage and attenuation of hypotension may result from induction of HSP by whole-body hyperthermia. PMID- 11254395 TI - Isolation and in vitro characterization of human dermal microvascular pericytes. AB - Pericytes cover the abluminal surface of capillaries and venules and are thought to play an important role in microvascular regulation and pathology. The purpose of this study was to isolate and characterize human dermal microvascular pericytes (HDMPC), a minor cell type in the skin but a relatively easily obtainable human source of tissue. We developed and compared two procedures that differed in the preselection method. Isolation of dermal microvessel fragments from neonatal foreskins by trypsin digestion was followed by mechanical release of subepidermal tissue, collagenase treatment, and sieving through 100- and 30 microm meshes. After subcultivation, pericytes were preselected either by isolation of outgrowing capillary fragments or by 3G5-coupled magnetic beads. Pericytes were selected finally by cultivation of single cells in endothelial cell-conditioned media. Cultured HDMPC were seen to be large and well spread with irregular edges and prominent stress fibers. They lack contact inhibition, are positive for 3G5 antigen, alpha-smooth muscle actin, and vimentin, and are negative for the endothelial cell marker CD31, diI-acetylated low-density lipoprotein uptake, cytokeratin 5, 6, and 18, and S100 protein. Using both preselection methods, we could establish purified cell cultures of HDMPC. The results of these studies represent the first report of HDMPC isolation. PMID- 11254396 TI - Protective effect of aminoguanidine upon capillary and submesothelial anionic sites. AB - This study evaluates albuminuria and peritoneal permeability to albumin in control and diabetic rats, as well as in diabetic animals treated with subcutaneously injected aminoguanidine hydrochloride (Ag) (5 mg/100 g/day), during a follow-up period of 6 months. Aminoguanidine effectively prevented albuminuria and albumin extravasation in the mesenteric interstitial tissue (control, 0.43 +/- 0.11 microg EB/100 g of dry tissue, Ag, 0.60 +/- 0.44; untreated diabetic animals, 1.22 +/- 0.73; control and Ag group vs untreated diabetic rats, P < 0.001). Albumin D/P ratio of the aminoguanidine-exposed rats (0.017 +/- 0.011) was higher than that of controls (0.008 +/- 0.002), but significantly lower (P < 0.001) than values observed in the untreated group of animals (0.046 +/- 0.003). Administration of aminoguanidine preserved both submesothelial and subendothelial electronegative charges. For capillary basement membrane (BM), control at zero time, 32 +/- 4 AS/microm BM; control at 6 months, 33.4; aminoguanidine-treated rats, 35 +/- 2. For submesothelial BM, control at zero time, 33 +/- 3; control at 6 months, 32 +/- 3; aminoguanidine-treated rats, 35 +/- 3. Splitting and thickening of both basement membranes were not prevented by the therapeutic intervention. We conclude that the shielding effect of aminoguanidine upon the permselectivity capabilities of the endothelial and mesothelial monolayers appears to be connected, basically to the preservation of anionic fixed charges. PMID- 11254397 TI - Endothelial K(ca) channels mediate flow-dependent dilation of arterioles of skeletal muscle and mesentery. AB - The role of Ca(2+)-activated potassium channels (K(Ca)) in flow-initiated intracellular events in microvessels is not known. We hypothesized that K(Ca) channels in the arteriolar endothelium are responsible for the mechanotransduction of flow/shear stress-induced arteriolar dilation in skeletal muscle and mesentery of rats. The active diameter of arterioles isolated from gracilis (80 mm Hg) and cremaster (60 mm Hg) muscles and mesentery (80 mm Hg) at a constant intraluminal pressure was 53 +/- 3, 77 +/- 5, and 72 +/- 6 microm, respectively. Their passive diameter (in Ca(2+)-free solution) was 113 +/- 3, 152 +/- 12, and 121 +/- 7 microm, respectively. At a constant intraluminal pressure stepwise increases in perfusate flow (25, 40, and 14 microL/min in 5, 10, and 2 microL/min steps) elicited a gradual increase in diameter of all three groups of arterioles up to 93 +/- 5, 137 +/- 11, and 102 +/- 7 microm, respectively. Flow induced dilations of arterioles were eliminated by intraluminal administration of iberiotoxin (ibTX 10(-9) M), an inhibitor of high conductance K(Ca) channels (BK(Ca)). In contrast, arteriolar dilations to acetylcholine and sodium nitroprusside were not altered by this agent, indicating that BK(Ca) channels are not involved in the receptor-mediated endothelial synthesis of nitric oxide (NO) and that the inhibitor did not affect the action of NO on smooth muscle. Abluminal application of ibTX (10(-8) M) did not affect flow-dependent dilation. We conclude that in arterioles of several tissues activation of endothelial BK(Ca) channels is an obligatory step in the transduction of the signal initiated by changes in intraluminal flow/shear stress, leading to the release of endothelial factors evoking dilation. PMID- 11254399 TI - Microvessel oxyhemoglobin saturation does not reflect tissue oxygen tension in human melanoma xenografts. PMID- 11254398 TI - Microvascular permeability of human melanoma xenografts to macromolecules: relationships to tumor volumetric growth rate, tumor angiogenesis, and VEGF expression. AB - The effective microvascular permeability of human melanoma xenografts to albumin Evans blue was measured and related to tumor volumetric growth rate, rate of tumor angiogenesis, and expression of vascular endothelial growth factor (VEGF) in an attempt to identify mechanisms regulating the microvascular permeability of tumors to macromolecules. Three melanoma lines (A-07, R-18, and U-25) were included in the study. Effective microvascular permeability was assessed by using the indicator diffusion method. Intradermal and intratumor angiogenesis assays were used to measure the rate of tumor angiogenesis. VEGF expression was studied by ELISA, immunohistochemistry, Western blotting, and measurement of tumor induced formation of ascitic fluid. The effective microvascular permeabilities of albumin-Evans blue were determined to be (1.5 +/- 0.2) x 10(-6) cm/s (A-07), (1.1 +/- 0.4) x 10(-6) cm/s (R-18), and (0.9 +/- 0.3) x 10(-6) cm/s (U-25). These values are high compared with those measured for other tumor lines and are not significantly different. Correlations between the effective microvascular permeability of albumin-Evans blue and tumor volumetric growth rate, rate of tumor angiogenesis, or VEGF expression were not found. The three last-mentioned parameters differed significantly among the melanoma lines and covered a broad range of values relative to those of other experimental tumors. Our study suggests that the effective microvascular permeability of macromolecules can be high even in slowly growing tumors, poorly angiogenic tumors, and tumors showing low VEGF expression. PMID- 11254400 TI - Kinetics of placenta growth factor/vascular endothelial growth factor synergy in endothelial hydraulic conductivity and proliferation. AB - Vascular endothelial growth factor (VEGF) was originally discovered as vascular permeability factor because of its ability to increase microvascular permeability to plasma proteins. Since then, it has been shown to induce proliferation and migration in endothelial cells. Placenta growth factor (PlGF) is a member of the VEGF family of growth factors, but has little or undetectable mitogenic activity on endothelial cells. Intriguingly, however, PlGF was able to potentiate the action of low concentrations of VEGF on endothelial cell growth and macromolecule permeability in vitro. Here we show that PlGF can potentiate the effects of VEGF on the hydraulic conductivity of certain endothelial cells and that the duration of pretreatment with PlGF determines the resulting response. Hydraulic conductivity (Lp) was calculated from the water flux across the monolayer of human umbilical vein endothelial cells (HUVECs) or bovine aortic endothelial cells (BAECs). After 2 h of exposure to VEGF(165), the Lp of BAEC monolayers increased threefold, but the Lp of HUVEC monolayers did not increase. PlGF alone induced a small (63%) increase in Lp in BAECs, but not in HUVECs. BAEC, but not HUVEC, monolayers exposed first to PlGF and then to VEGF exhibited a seven- to eightfold increase in Lp. This enhancement in BAEC Lp could be observed for 4 h after the administration of PlGF. PlGF also potentiated the effect of VEGF on BAEC proliferation. Thus, augmentation of VEGF action by PlGF depends on the duration of PlGF exposure and on the origin of endothelial cells. PMID- 11254401 TI - FGF-2 promotes disassembly of actin cytoskeleton and shape changes in murine vascular cells. PMID- 11254402 TI - An in vitro model to evaluate regulatory mechanisms of antigen expression by normal pulmonary vessel endothelial cells. PMID- 11254403 TI - Isolation and purification of canine adipose microvascular endothelial cells. PMID- 11254404 TI - Coronary vascular resistance and ST-segment changes during coronary microvascular spasm. PMID- 11254406 TI - Patient satisfaction with cleft lip and palate services in a regional centre. AB - We sent a questionnaire to the parents of 478 children aged between 3 and 14 years who are under the care of the cleft team at the Queen Victoria Hospital, East Grinstead, and received 341 replies. A wide variety of questions were asked about aspects of patient satisfaction, and the results are reported and discussed. There was a high level of satisfaction with the service provided but 30% of parents would like to be more involved in treatment-planning decisions; 33% thought they had either not enough or no knowledge about cleft lip and palate and its treatment. Only 8% of parents would rather have seen the specialists separately than together in the joint clinic. PMID- 11254407 TI - Superiorly based flap pharyngoplasty: the degree of postoperative "tubing" and its effect on speech. AB - It is recognised that superiorly based pharyngeal flaps tend to contract resulting in narrowing and lowering of the flaps. If lateral pharyngeal-wall motion is unable to close against the "tubed" flap or if the flap migrates below the level of medial displacement of the lateral pharyngeal walls, velopharyngeal insufficiency will result. The extent of this phenomenon of flap contracture or shrinkage has not been previously quantified. A consecutive series of 120 superior flap pharyngoplasty operations were assessed critically and carefully. The mean width of the harvested flap measured 89% of the width of the pharyngeal posterior wall and shrank over 6 months to 45% of the lateral pharyngeal diameter. The relations between speech results, complication rate and remaining flap width are analysed. All flaps shrink but to a varying degree. PMID- 11254408 TI - A prospective study of the effect of botulinum toxin A on masseteric muscle hypertrophy with ultrasonographic and electromyographic measurement. AB - We evaluated the effect of botulinum toxin A on masseteric muscle hypertrophy by using ultrasound and electromyography. Five patients (four with bilateral and one with unilateral masseteric muscle hypertrophy) were studied prospectively. In each patient, ultrasound-guided percutaneous intramuscular injection of botulinum toxin A was carried out. The change in muscle bulk was evaluated using serial ultrasonography and the electrical activity was assessed with electromyography. All five patients (nine hypertrophic muscles) showed a good response, with the maximal effect of a 31% reduction in muscle bulk seen 3 months after treatment. The effect remained stable one year after injection for six of the hypertrophic muscles, whereas three muscles needed a second injection to maintain the atrophy. This preliminary prospective study suggests that botulinum toxin A is a safe alternative method of treating masseteric muscle hypertrophy. However, the effect may be temporary and further intramuscular injection may be required to maintain atrophy. PMID- 11254409 TI - Reconstruction of the retroauricular fold: a personal technique. AB - In auricular reconstruction emphasis is placed on carving the rib-cartilage framework. The three-dimensional frame is very important in obtaining a good anatomical shape but often a good shape is not complemented by a good projection of the auricle. In order to avoid obliteration of the retroauricular fold we use a cartilage wedge covered by a local fascial flap. We have treated 17 ears in 16 patients with this technique and have obtained satisfactory results in all cases, achieving a mean projection of 1.7cm between the mastoid plane and the helical rim. PMID- 11254410 TI - Platysma muscle turnover flap correction of tracheostomy scarring deformity. AB - There are numerous techniques for the reconstruction of the tracheostomy scarring deformity that give acceptable results. In this paper, we present a new method for improving this defect, using two rectangular turnover platysma muscle flaps to fill the concavity. The skin is then sutured over them. We have used this technique in10 patients over the last 2 years and achieved good aesthetic results. Our follow-up has shown that we have achieved a stable reconstruction with no functional impairment. PMID- 11254412 TI - The salute splint for axillary contractures. AB - A new splint for use in the treatment of burn contractures of the axilla is presented. The splint is easy to deploy, comfortable for patients and achieves the same results as other splints. PMID- 11254411 TI - The use of a bilaminate artificial skin substitute (Integra) in acute resurfacing of burns: an early experience. AB - Integra artificial skin provides immediate full-thickness reconstruction for cutaneous burns. The clinical outcome appears to be superior in terms of final function and cosmesis. Consequently the use of such a skin substitute is being heralded as the future treatment of choice, particularly for massive burns where autologous donor skin is limited. The three cases reported here describe the senior author's early experience with Integra and highlight some of the difficulties and successes encountered. A high rate of dermal graft loss and slow epidermal engraftment have tempered the original enthusiasm, but with growing experience the final outcome justifies the continued use of Integra in our unit. PMID- 11254413 TI - The Derriford Appearance Scale (DAS59): a new psychometric scale for the evaluation of patients with disfigurements and aesthetic problems of appearance. AB - The DAS59 has been designed and developed to meet the need for an objective measure of the spectrum of psychological distress and dysfunction that is characteristic of disfigurements, deformities and aesthetic problems of appearance. Content validity has been assured by basing the scale's items on a detailed autobiographical study of representative patients. Internal consistency is high (0.98) and test-retest reliabilities are good (general population: 0.75; clinical population: 0.86). Correlations with other appropriate standardised tests show good criterion validity and good construct validity. Factor analysis of 2741 data sets (general population and clinical population) identified three factors that are not feature specific and two that are (bodily and sexual features, facial features). The DAS59 thus generates a full-scale score and five factorial sub-scale scores. The DAS59 has been standardised on the clinical population across a range of patient groups and on the general population subdivided into those concerned and those not concerned about appearance. The DAS59 is highly sensitive as a measure of change following treatment with large and significant preoperative-postoperative reductions in full-scale and factorial scores of patients treated for facial features or bodily/sexual features. The DAS59 offers benefits for patient selection in both cosmetic and reconstructive plastic surgery and in the evaluation of outcome. It provides valid and reliable data for clinical audit and governance and for evaluating the merits of one treatment protocol against another. PMID- 11254414 TI - Prevalence of concern about physical appearance in the general population. AB - Using information gathered in the introductory sections of the Derriford Appearance Scales (DAS24 and DAS59), the prevalence and epidemiological characteristics of concern about physical appearance have been determined for a carefully constructed sample of the general population of southwest Devon (rural and urban). In all, 2108 usable replies were received from a postal survey of a targeted population of 5400 men and women, aged 18 and over and randomly selected with constraints for age, sex and socio-economic status. The prevalence of concern about physical appearance was highest among women through to age 60 and younger men. There was no association with socio-economic status or living status. Concerns about the nose, weight and skin disorders were reported most frequently by both men and women and additionally concerns about breasts and abdomen were reported by women and premature balding by men. The mean DAS24 and DAS59 full-scale scores of 19% of male and 25% of female responders who were concerned about appearance exceeded the mean scores of preoperative patients undergoing reconstructive and cosmetic plastic surgery. Concern about appearance is widespread in the general population. More often than not, concern is about one feature only, which runs counter to the hypothesis that concern about appearance reflects a neurotic trait. The high levels of measured psychological distress and dysfunction found in a substantial minority of those in the general population who are concerned about appearance highlight the need for appropriate services. PMID- 11254416 TI - Management of paraffinoma of the breast: 10 years' experience. AB - Paraffin injection was regarded as a simple and effective method of improving body contour. It was widely used in breast augmentation until the long-term complication of paraffinoma was recognised. Paraffinoma of the breast can present as a spectrum of disease ranging from a painless mass to a destructive ulcer simulating breast cancer. This makes it difficult to make the correct diagnosis and provide suitable treatment. Eight patients with paraffinoma of the breast have been managed at a teaching hospital over a 10 year period. All were females, with a mean age of 65.6 years (range: 57-73 years). The average time between paraffin injection and the onset of symptoms was 24 years (range: 11-30 years). These patients have been followed up for between 3 and 10 years (mean: 6.1 years). One patient died of congestive heart failure 4 years after bilateral mastectomy for painful paraffinomas of the breasts. Another patient had a coexisting infiltrative ductal carcinoma of the breast. The clinical presentations, radiological appearances, histopathological features and the treatment are discussed. PMID- 11254415 TI - How many procedures to make a breast? AB - The construction of a new breast after mastectomy involves fashioning the breast mound and creating a projecting nipple and a coloured areola. This should involve three episodes for a patient, but is this the experience of patients embarking on breast reconstruction? We identified 177 patients who had undergone breast reconstruction between 1 September 1997 and 31 March 1999. The clinical records for 164 of these patients were found and the data summarised. The techniques, complications and other ancillary procedures experienced by this group of patients are presented. Multiple procedures are likely to be required to complete breast reconstruction, and the patient should be so counselled from the outset. Particular problems may be encountered with each technique and this should be borne in mind when selecting a procedure for each patient, especially in the context of immediate reconstruction where avoiding any delay to adjuvant treatment is a consideration. PMID- 11254417 TI - Ambulant vacuum-assisted closure of skin-graft dressing in the lower limbs using a portable mini-VAC device. AB - A skin graft may fail to adhere to the recipient site because of fluid collecting between the graft and the area being treated. We have devised a simple procedure, consisting of a vacuum-sealed dressing, to fix skin grafts on the lower limbs. A fully portable, battery-operated aspirator continuously draws secretions through a vacuum-sealed dressing, preventing accumulation of fluid underneath the graft. Patients are not confined to bed, thus reducing nursing time. The procedure was applied successfully in seven out of nine patients treated for ulcers of the lower limbs. PMID- 11254418 TI - Topical negative pressure for treating chronic wounds: a systematic review. AB - Some wounds take a long time to heal, fail to heal or recur, causing significant pain and discomfort to the patient and cost to the National Health Service. This review assesses the effectiveness of topical negative pressure (TNP) in treating chronic wounds. The Cochrane Wounds Group Specialised Trials Register was searched for randomised controlled trials (RCTs) that evaluated the effectiveness of TNP on chronic-wound healing. Eligibility for inclusion, data extraction and details of trial quality were conducted by two reviewers independently. A narrative synthesis of results was undertaken as only two small trials, with different outcome measures, fulfilled the selection criteria. Trial 1 considered any type of chronic wound, trial 2 considered diabetic foot ulcers. Both trials compared TNP with saline-gauze dressings. Trial 1 reported a statistically significant difference in the percentage change in wound volume after 6 weeks, in favour of TNP. Trial 2 reported a difference in the number of days to healing and a difference in the percentage change in wound surface area after 2 weeks, in favour of TNP. These two small trials provide weak evidence to suggest that TNP may be superior to saline-gauze dressings in terms of wound healing. However, due to the small sample sizes and the methodological limitations of the studies, these findings must be interpreted with extreme caution. The effects of TNP on cost, quality of life, pain and comfort were not reported. It was not possible to determine the optimum TNP regimen. Further high-quality RCTs that address these issues are required. PMID- 11254419 TI - Contact dermatitis complicating pinnaplasty. AB - Proflavine allergy is uncommon, occurring in approximately 6% of patients attending contact dermatitis clinics. Proflavine wool is used by many surgeons in the UK as a dressing that can be moulded to conform to the contours of a corrected prominent ear. It may have bacteriostatic properties. We present a case where contact dermatitis in response to proflavine developed after pinnaplasty. This caused diagnostic confusion, a lengthened hospital stay and an unsightly hypertrophic scar. PMID- 11254420 TI - Recurrent pleomorphic adenoma of the palate in a child. AB - A rare case of recurrent pleomorphic adenoma of the palate in a 9-year-old boy is presented. Pleomorphic adenoma is relatively rare in children compared with its incidence in adults. However, it is the most common benign epithelial tumour of the salivary glands. The majority of pleomorphic adenomata in children occur in the major salivary glands, mainly the parotid gland. Pleomorphic adenomata of the minor salivary glands are rare in children and mainly occur in the palatal glands. Of the few cases of pleomorphic adenoma of the palate reported in children, only one case showed recurrence of the tumour after primary excision. We present the second case of recurrent pleomorphic adenoma of the palate in a child. PMID- 11254421 TI - Reconstruction of the inner canthus region with a forehead muscle flap: a report on three cases. AB - We report our experience of using a forehead flap to repair the defect left by the excision of skin tumours in the medial canthal region involving both eyelids in three patients. Both eyelids and the inner canthus were reconstructed using a myofascial flap taken from the forehead, combined with septal chondro-mucosal grafts, oral mucosa and skin grafts. After a careful anatomical study of the vascularisation of the frontal region, we used only the frontal myofascial portion, a part of the forehead muscle vascularised by the deep branch of the supraorbital artery and by the supratroclear artery; the skin left behind is adequately nourished by the fine mesh of anastomoses in the area between the two supratroclear arteries, the supraorbital artery and the terminal vessels of the superficial temporal artery. The particularly thin, elastic and resistant features of this flap enabled us to repair a loss of substance in a difficult area with a successful outcome in terms of morphology, function and cosmetic appearance. PMID- 11254422 TI - Argyria caused by an earring. AB - The staining of skin by silver is termed argyria and is grey-blue in colour. This may be caused by a number of mechanisms such as ingestion and direct implantation. We report an unusual case, caused by an impacted earring, where the skin discoloration was not entirely typical of argyria. This may have been due to copper impurities present in the earring. The literature on the subject is also reviewed. PMID- 11254423 TI - Spread of a recurrent lentigo maligna into a graft: a case for conservative treatment. AB - Lentigo maligna is an in situ malignant melanoma for which the treatment of choice is surgical excision. The recurrence rate in lentigo maligna is high and hence other treatments, such as cryotherapy, laser therapy, radiotherapy and Mohs' chemosurgery, have been described. Despite the high recurrence rate, spread of a lentigo maligna into a skin graft is rare. We describe a case of a recurrent lentigo maligna spreading into a skin graft, which, along with the cases described in the literature, highlights the presence of a group of low-grade malignant lentiginous lesions that may be managed by careful follow-up and observation. PMID- 11254424 TI - Expanding silicone granuloma. AB - Complications of breast augmentation using silicone implants have been the subject of much discussion. We report a single case of a silicone granuloma, which has exhibited unusual behaviour in that it has grown rapidly and significantly. Whilst silicone granulomata have been reported on many occasions in the past, to our knowledge this is the first report of a rapidly growing example. PMID- 11254425 TI - Congenital sternal cleft. AB - A cleft of the sternum is a rare congenital anomaly. We present a case of a sternal cleft in a 7-year-old boy. A split iliac bone graft covered with the sternocostal portion of a pectoralis major flap was used to reconstruct the defect. PMID- 11254426 TI - Modified dorsalis pedis flap for coverage of a pretibial pressure sore after hip rotationplasty. AB - Rotationplasty is a well-established procedure after total femur resection, especially in children. Rehabilitation is superior to disarticulation of the hip or hemipelvectomy because patients regain hip and knee function.(1) A tight fit of the prosthetic shaft is essential. The pretibial area has a low physiological resistance to pressure and shear forces, and is thus at increased risk of developing pressure-related complications. Skin defects with exposure of skeletal elements require flap coverage. The dorsalis pedis flap is one of the surgical options available for skin coverage of the proximal anterior leg. It can be rotated to cover almost any site on the anterior aspect of the leg if the pedicle is mobilised up to the anterior tibial artery.(2)Since donor site complications are common, this flap has few indications.(3) PMID- 11254427 TI - Acute exertional compartment syndrome in an athlete. AB - Acute exertional compartment syndrome is a rare condition, associated with strenuous, unaccustomed exercise. This report describes its onset in a professional footballer during a regular training session. It is often diagnosed late due to lack of awareness and patient stoicism. We illustrate the consequences of delay and reinforce the need for prompt and decisive fasciotomy if complications are to be avoided. PMID- 11254428 TI - Extra-abdominal desmoid tumour of the breast: review of the primary management and the implications for breast reconstruction. AB - This case report illustrates the presentation and management of an extra abdominal desmoid tumour of the breast. A review of the literature describing the aetiology, pathology and risk of recurrence was undertaken to determine how current understanding of this rare tumour may affect the management of patients, should they require breast reconstruction after radical excision of the primary tumour. The natural progression of the disease is variable and there are no markers predictive of recurrence or regression. Primary lesions should be assessed with respect to their anatomical site of origin (i.e. whether they arise within the breast or invade the breast from the underlying musculo-aponeurotic tissue) and the extent of local invasion. Radical excision of the tumour with clear histological margins decreases the likelihood of recurrence. Tumours arising from the musculo-aponeurotic system have increased risks of recurrence and of developing multifocal primary tumours in specific anatomical territories. Local recurrences should be assessed for extent and anatomical distribution, and radical excision performed as for a primary tumour. Radiotherapy can be used as an alternative treatment if radical excision of a primary or recurrent tumour would cause severe functional loss or mutilation. Radiotherapy can be used for positive histological margins following tumour excision. There is a higher risk of recurrence in the first 3 years after primary excision, and breast reconstruction may be best delayed for this period. Surgical trauma has been implicated in the aetiology of recurrence and the patient should be informed of this prior to breast reconstruction. PMID- 11254429 TI - TRAM and DIEP flap zones. PMID- 11254430 TI - Three-dimensional skin-graft contraction in full-thickness grafts. PMID- 11254431 TI - The use of conchal-cartilage grafts in the closure of recurrent palatal fistulae. PMID- 11254432 TI - Sentinel lymph node biopsy in malignant melanoma. PMID- 11254433 TI - Vascularised temporoparietal fascial flap for closure of an orocutaneous fistula. PMID- 11254435 TI - Fine bone rasps in unexpected places. PMID- 11254434 TI - Large scalp and skull defects in aplasia cutis congenita. PMID- 11254436 TI - Multiple pilomatrixomata without myotonic dystrophy. PMID- 11254437 TI - Johann Friedrich Dieffenbach: successful use of leeches in plastic surgery in the 1820s. PMID- 11254438 TI - MRSA - again. PMID- 11254439 TI - On discovery, genomes, the Society, and society. PMID- 11254440 TI - 2000 ASHG Award for Excellence in Education: introductory speech for F. Clarke Fraser. PMID- 11254441 TI - 2000 ASHG Award for Excellence in Education. Resetting our educational sights: unconstructing the public's dreams and nightmares of the genetic revolution. PMID- 11254442 TI - Mutations of MLC1 (KIAA0027), encoding a putative membrane protein, cause megalencephalic leukoencephalopathy with subcortical cysts. AB - Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is an autosomal recessive disorder characterized by macrocephaly, deterioration of motor functions with ataxia, and spasticity, eventuating in mental decline. The brain appears swollen on magnetic resonance imaging, with diffuse white-matter abnormalities and the invariable presence of subcortical cysts. MLC was recently localized on chromosome 22q(tel). We have narrowed down the critical region by linkage analysis of 11 informative families with MLC to a region of approximately 250 kb, containing four known genes. One family with two patients who were siblings did not display linkage between the MLC phenotype and any of the analyzed microsatellite markers on chromosome 22q(tel), suggesting genetic heterogeneity and the existence of at least a second MLC locus. The maximum two point LOD score for the 11 families was 6.6 at recombination fraction .02. Twelve different mutations in seven informative and six uninformative families were found in one of the candidate genes, KIAA0027, which we renamed "MLC1." The gene encodes a putative membrane protein with eight predicted transmembrane domains. The patients of one family were compound heterozygotes for mutations that both introduced stop codons. The mutations further included frameshifts, splice acceptor mutations, a putative splice-donor mutation, and amino acid substitutions of residues in predicted transmembrane domains. These data provide strong evidence that mutations of MLC1 cause the disease. PMID- 11254443 TI - Disruption of a novel gene (IMMP2L) by a breakpoint in 7q31 associated with Tourette syndrome. AB - Gilles de la Tourette syndrome (GTS) is a complex neuropsychiatric disorder characterized by multiple motor and phonic tics. We identified a male patient with GTS and other anomalies. It was determined that he carried a de novo duplication of the long arm of chromosome 7 [46,XY,dup(7)(q22.1-q31.1)]. Further molecular analysis revealed that the duplication was inverted. The distal chromosomal breakpoint occurred between the two genetic markers D7S515 and D7S522, which define a region previously shown to be disrupted in a familiar case of GTS. Yeast and bacterial artificial chromosome clones spanning the breakpoints were identified by means of FISH analysis. To further characterize the distal breakpoint for a role in GTS, we performed Southern blot hybridization analysis and identified a 6.5-kb SacI junction fragment in the patient's genomic DNA. The DNA sequence of this fragment revealed two different breaks in 7q31 within a region of approximately 500 kb. IMMP2L, a novel gene coding for the apparent human homologue of the yeast mitochondrial inner membrane peptidase subunit 2, was found to be disrupted by both the breakpoint in the duplicated fragment and the insertion site in 7q31. The cDNA of the human IMMP2L gene was cloned, and analysis of the complete 1,522-bp transcript revealed that it encompassed six exons spanning 860 kb. The possible role of IMMP2L and several other candidate genes within the region of chromosomal rearrangement, including NRCAM, Leu-Rch Rep, and Reelin, is discussed. The 7q31 breakpoint interval has also been implicated in other neuropsychiatric diseases that demonstrate some clinical overlap with GTS, including autism and speech-language disorder. PMID- 11254445 TI - A novel SCN1A mutation associated with generalized epilepsy with febrile seizures plus--and prevalence of variants in patients with epilepsy. AB - We recently described mutations of the neuronal sodium-channel alpha-subunit gene, SCN1A, on chromosome 2q24 in two families with generalized epilepsy with febrile seizures plus (GEFS+) type 2. To assess the contribution that SCN1A makes to other types of epilepsy, 226 patients with either juvenile myoclonic epilepsy, absence epilepsy, or febrile convulsions were screened by conformation-sensitive gel electrophoresis and manual sequencing of variants; the sample included 165 probands from multiplex families and 61 sporadic cases. The novel mutation W1204R was identified in a family with GEFS+. Seven other coding changes were observed; three of these are potential disease-causing mutations. Two common haplotypes, with frequencies of .67 and .33, were defined by five single-nucleotide polymorphisms (SNPs) spanning a 14-kb region of linkage disequilibrium. An SNP located 18 bp upstream of the splice-acceptor site for exon 3 was observed in 7 of the 226 patients but was not present in 185 controls, suggesting possible association with a disease mutation. This work has confirmed the role of SCN1A in GEFS+, by identification of a novel mutation in a previously undescribed family. Although a few candidate disease alleles were identified, the patient survey suggests that SCN1A is not a major contributor to idiopathic generalized epilepsy. The SCN1A haplotypes and SNPs identified here will be useful in future association and linkage studies. PMID- 11254444 TI - Neuronal sodium-channel alpha1-subunit mutations in generalized epilepsy with febrile seizures plus. AB - Generalized epilepsy with febrile seizures plus (GEFS+) is a familial epilepsy syndrome characterized by the presence of febrile and afebrile seizures. The first gene, GEFS1, was mapped to chromosome 19q and was identified as the sodium channel beta1-subunit, SCN1B. A second locus on chromosome 2q, GEFS2, was recently identified as the sodium-channel alpha1-subunit, SCN1A. Single-stranded conformation analysis (SSCA) of SCN1A was performed in 53 unrelated index cases to estimate the frequency of mutations in patients with GEFS+. No mutations were found in 17 isolated cases of GEFS+. Three novel SCN1A mutations-D188V, V1353L, and I1656M-were found in 36 familial cases; of the remaining 33 families, 3 had mutations in SCN1B. On the basis of SSCA, the combined frequency of SCN1A and SCN1B mutations in familial cases of GEFS+ was found to be 17%. PMID- 11254446 TI - Melanocortin-1 receptor gene variants determine the risk of nonmelanoma skin cancer independently of fair skin and red hair. AB - Melanocortin-1 receptor (MC1R) gene variants are associated with fair skin and red hair and, independently of these, with cutaneous malignant melanoma. The association of MC1R gene variants with nonmelanoma skin cancer is largely unknown. A total of 838 subjects were included in the present study: 453 patients with nonmelanoma skin cancer and 385 subjects with no skin cancer. The coding sequence of the human MC1R gene was tested using single-stranded conformation polymorphism analysis followed by sequencing of unknown variants. Risk of skin cancer dependent on the various MC1R gene variants was estimated using the exposure odds ratio. We investigated whether subjects with MC1R variant alleles were at increased risk of developing nonmelanoma skin cancer and, if so, whether this increased risk was mediated by fair skin and red hair. A total of 27 MC1R gene variants were found. The number of carriers of one, two, or three MC1R gene variants was 379 (45.2%), 208 (24.8%), and 7 (0.9%), respectively. A strong association between MC1R gene variants and fair skin and red hair was established, especially the variants Arg151Cys and Arg160Trp (P < .0001). Carriers of two variant alleles were at increased risk for developing cutaneous squamous cell carcinoma (odds ratio 3.77; 95% confidence interval [CI] 2.11 6.78), nodular basal cell carcinoma (odds ratio 2.26; 95% CI 1.45-3.52), and superficial multifocal basal cell carcinoma (odds ratio 3.43; 95% CI 1.92-6.15), compared with carriers of two wild-type alleles. Carriers of one variant allele had half the risk. The highest relative risks of nonmelanoma skin cancer were found in carriers of the Asp84Glu, His260Pro, and Asp294His variant alleles, and the risk was only slightly lower for carriers of the Val60Leu, Val92Met, Arg142His, Arg151Cys, and Arg160Trp variant alleles. When subjects were stratified by skin type and hair color, analysis showed that these factors did not materially change the relative risks. These findings indicate that MC1R gene variants are important independent risk factors for nonmelanoma skin cancer. PMID- 11254447 TI - Localization of a novel locus for autosomal recessive early-onset parkinsonism, PARK6, on human chromosome 1p35-p36. AB - The cause of Parkinson disease (PD) is still unknown, but genetic factors have recently been implicated in the etiology of the disease. So far, four loci responsible for autosomal dominant PD have been identified. Autosomal recessive juvenile parkinsonism (ARJP) is a clinically and genetically distinct entity; typical PD features are associated with early onset, sustained response to levodopa, and early occurrence of levodopa-induced dyskinesias, which are often severe. To date, only one ARJP gene, Parkin, has been identified, and multiple mutations have been detected both in families with autosomal recessive parkinsonism and in sporadic cases. The Parkin-associated phenotype is broad, and some cases are indistinguishable from idiopathic PD. In > or = 50% of families with ARJP that have been analyzed, no mutations could be detected in the Parkin gene. We identified a large Sicilian family with four definitely affected members (the Marsala kindred). The phenotype was characterized by early-onset (range 32 48 years) parkinsonism, with slow progression and sustained response to levodopa. Linkage of the disease to the Parkin gene was excluded. A genomewide homozygosity screen was performed in the family. Linkage analysis and haplotype construction allowed identification of a single region of homozygosity shared by all the affected members, spanning 12.5 cM on the short arm of chromosome 1. This region contains a novel locus for autosomal recessive early-onset parkinsonism, PARK6. A maximum LOD score 4.01 at recombination fraction .00 was obtained for marker D1S199. PMID- 11254448 TI - Evaluation of linkage and association of HPC2/ELAC2 in patients with familial or sporadic prostate cancer. AB - To investigate the relationship between HPC2/ELAC2 and prostate cancer risk, we performed the following analyses: (1) a linkage study of six markers in and around the HPC2/ELAC2 gene at 17p11 in 159 pedigrees with hereditary prostate cancer (HPC); (2) a mutation-screening analysis of all coding exons of the gene in 93 probands with HPC; (3) family-based and population-based association study of common HPC2/ELAC2 missense variants in 159 probands with HPC, 249 patients with sporadic prostate cancer, and 222 unaffected male control subjects. No evidence for linkage was found in the total sample, nor in any subset of pedigrees based on characteristics that included age at onset, number of affected members, male-to-male disease transmission, or race. Furthermore, only the two previously reported missense changes (Ser217Leu and Ala541Thr) were identified by mutational analysis of all HPC2/ELAC exons in 93 probands with HPC. In association analyses, family-based tests did not reveal excess transmission of the Leu217 and/or Thr541 alleles to affected offspring, and population-based tests failed to reveal any statistically significant difference in the allele frequencies of the two polymorphisms between patients with prostate cancer and control subjects. The results of this study lead us to reject the three alternative hypotheses of (1) a highly penetrant, major prostate cancer susceptibility gene at 17p11, (2) the allelic variants Leu217 or Thr541 of HPC2/ELAC2 as high-penetrance mutations, and (3) the variants Leu217 or Thr541 as low-penetrance, risk-modifying alleles. However, we did observe a trend of higher Leu217 homozygous carrier rates in patients than in control subjects. Considering the impact of genetic heterogeneity, phenocopies, and incomplete penetrance on the linkage and association studies of prostate cancer and on the power to detect linkage and association in our study sample, our results cannot rule out the possibility of a highly penetrant prostate cancer gene at this locus that only segregates in a small number of pedigrees. Nor can we rule out a prostate cancer modifier gene that confers a lower-than-reported risk. Additional larger studies are needed to more fully evaluate the role of this gene in prostate cancer risk. PMID- 11254449 TI - HPC2 variants and screen-detected prostate cancer. AB - Two studies have reported significant associations between susceptibility to prostate cancer and two common missense variants of the HPC2/ELAC2 gene, with estimated relative risks in the range of two- to threefold. We investigated whether these polymorphisms could be informative in the prediction of the presence of prostate cancer in men undergoing prostatic biopsy for the evaluation of an elevated serum-PSA level (> or = 4.0 ng/ml). We genotyped 944 men who underwent a prostate biopsy at our institution, as well as a control population of 922 healthy, unselected women from the same population. The prevalence of the HPC2 Ala541Thr allele was similar in men with prostate cancer (6.3%), men with other prostatic conditions (6.8%), and healthy women (6.3%) (P = .83). We conclude that HPC2 genotyping is unlikely to be a useful adjunct to PSA in the prediction of the presence of biopsy-detected prostate cancer in asymptomatic men. PMID- 11254452 TI - Assessment of parent-of-origin effects in linkage analysis of quantitative traits. AB - Methods are presented for incorporation of parent-of-origin effects into linkage analysis of quantitative traits. The estimated proportion of marker alleles shared identical by descent is first partitioned into a component derived from the mother and a component derived from the father. These parent-specific estimates of allele sharing are used in variance-components or Haseman-Elston methods of linkage analysis so that the effect of the quantitative-trait locus carried on the maternally derived chromosome is potentially different from the effect of the locus on the paternally derived chromosome. Statistics for linkage between trait and marker loci derived from either or both parents are then calculated, as are statistics for testing whether the effect of the maternally derived locus is equal to that of the paternally derived locus. Analyses of data simulated for 956 siblings from 263 nuclear families who had participated in a linkage study revealed that type I error rates for these statistics were generally similar to nominal values. Power to detect an imprinted locus was substantially increased when analyzed with a model allowing for parent-of-origin effects, compared with analyses that assumed equal effects; for example, for an imprinted locus accounting for 30% of the phenotypic variance, the expected LOD score was 4.5 when parent-of-origin effects were incorporated into the analysis, compared with 3.1 when these effects were ignored. The ability to include parent of-origin effects within linkage analysis of quantitative traits will facilitate genetic dissection of complex traits. PMID- 11254450 TI - A genomewide screen in multiplex rheumatoid arthritis families suggests genetic overlap with other autoimmune diseases. AB - Rheumatoid arthritis (RA) is an autoimmune/inflammatory disorder with a complex genetic component. We report the first major genomewide screen of multiplex families with RA gathered in the United States. The North American Rheumatoid Arthritis Consortium, using well-defined clinical criteria, has collected 257 families containing 301 affected sibling pairs with RA. A genome screen for allele sharing was performed, using 379 microsatellite markers. A nonparametric analysis using SIBPAL confirmed linkage of the HLA locus to RA (P < .00005), with lambdaHLA = 1.79. However, the analysis also revealed a number of non-HLA loci on chromosomes 1 (D1S235), 4 (D4S1647), 12 (D12S373), 16 (D16S403), and 17 (D17S1301), with evidence for linkage at a significance level of P<.005. Analysis of X-linked markers using the MLOD method from ASPEX also suggests linkage to the telomeric marker DXS6807. Stratifying the families into white or seropositive subgroups revealed some additional markers that showed improvement in significance over the full data set. Several of the regions that showed evidence for nominal significance (P < .05) in our data set had previously been implicated in RA (D16S516 and D17S1301) or in other diseases of an autoimmune nature, including systemic lupus erythematosus (D1S235), inflammatory bowel disease (D4S1647, D5S1462, and D16S516), multiple sclerosis (D12S1052), and ankylosing spondylitis (D16S516). Therefore, genes in the HLA complex play a major role in RA susceptibility, but several other regions also contribute significantly to overall genetic risk. PMID- 11254451 TI - Multipoint linkage-disequilibrium-mapping approach based on the case-parent trio design. AB - In the present study we propose a multipoint approach, for the mapping of genes, that is based on the case-parent trio design. We first derive an expression for the expected preferential-allele-transmission statistics for transmission, from either parent to an affected child, for an arbitrary location within a chromosomal region demarcated by several genetic markers. No assumption about genetic mechanism is needed in this derivation, beyond the assumption that no more than one disease gene lies in the region framed by the markers. When one builds on this representation, the way in which one may maximize the genetic information from multiple markers becomes obvious. This proposed method differs from the popular transmission/disequilibrium test (TDT) approach for fine mapping, in the following ways: First, in contrast with the TDT approach, all markers contribute information, regardless of whether the parents are heterozygous at any one marker, and incomplete trio data can be utilized in our approach. Second, rather than performing the TDT at each marker separately, we propose a single test statistic that follows a chi(2) distribution with 1 df, under the null hypothesis of no linkage or linkage disequilibrium to the region. Third, in the presence of linkage evidence, we offer a means to estimate the location of the disease locus along with its sampling uncertainty. We illustrate the proposed method with data from a family study of asthma, conducted in Barbados. PMID- 11254453 TI - Efficient multipoint linkage analysis through reduction of inheritance space. AB - Computational constraints currently limit exact multipoint linkage analysis to pedigrees of moderate size. We introduce new algorithms that allow analysis of larger pedigrees by reducing the time and memory requirements of the computation. We use the observed pedigree genotypes to reduce the number of inheritance patterns that need to be considered. The algorithms are implemented in a new version (version 2.1) of the software package GENEHUNTER. Performance gains depend on marker heterozygosity and on the number of pedigree members available for genotyping, but typically are 10-1,000-fold, compared with the performance of the previous release (version 2.0). As a result, families with up to 30 bits of inheritance information have been analyzed, and further increases in family size are feasible. In addition to computation of linkage statistics and haplotype determination, GENEHUNTER can also perform single-locus and multilocus transmission/disequilibrium tests. We describe and implement a set of permutation tests that allow determination of empirical significance levels in the presence of linkage disequilibrium among marker loci. PMID- 11254454 TI - A new statistical method for haplotype reconstruction from population data. AB - Current routine genotyping methods typically do not provide haplotype information, which is essential for many analyses of fine-scale molecular genetics data. Haplotypes can be obtained, at considerable cost, experimentally or (partially) through genotyping of additional family members. Alternatively, a statistical method can be used to infer phase and to reconstruct haplotypes. We present a new statistical method, applicable to genotype data at linked loci from a population sample, that improves substantially on current algorithms; often, error rates are reduced by > 50%, relative to its nearest competitor. Furthermore, our algorithm performs well in absolute terms, suggesting that reconstructing haplotypes experimentally or by genotyping additional family members may be an inefficient use of resources. PMID- 11254455 TI - An extensive analysis of Y-chromosomal microsatellite haplotypes in globally dispersed human populations. AB - The genetic variance at seven Y-chromosomal microsatellite loci (or short tandem repeats [STRs]) was studied among 986 male individuals from 20 globally dispersed human populations. A total of 598 different haplotypes were observed, of which 437 (73.1%) were each found in a single male only. Population-specific haplotype diversity values were.86-.99. Analyses of haplotype diversity and population specific haplotypes revealed marked population-structure differences between more isolated indigenous populations (e.g., Central African Pygmies or Greenland Inuit) and more-admixed populations (e.g., Europeans or Surinamese). Furthermore, male individuals from isolated indigenous populations shared haplotypes mainly with male individuals from their own population. By analysis of molecular variance, we found that 76.8% of the total genetic variance present among these male individuals could be attributed to genetic differences between male individuals who were members of the same population. Haplotype sharing between populations, phi(ST) statistics, and phylogenetic analysis identified close genetic affinities among European populations and among New Guinean populations. Our data illustrate that Y-chromosomal STR haplotypes are an ideal tool for the study of the genetic affinities between groups of male subjects and for detection of population structure. PMID- 11254456 TI - High-resolution analysis of human Y-chromosome variation shows a sharp discontinuity and limited gene flow between northwestern Africa and the Iberian Peninsula. AB - In the present study we have analyzed 44 Y-chromosome biallelic polymorphisms in population samples from northwestern (NW) Africa and the Iberian Peninsula, which allowed us to place each chromosome unequivocally in a phylogenetic tree based on >150 polymorphisms. The most striking results are that contemporary NW African and Iberian populations were found to have originated from distinctly different patrilineages and that the Strait of Gibraltar seems to have acted as a strong (although not complete) barrier to gene flow. In NW African populations, an Upper Paleolithic colonization that probably had its origin in eastern Africa contributed 75% of the current gene pool. In comparison, approximately 78% of contemporary Iberian Y chromosomes originated in an Upper Paleolithic expansion from western Asia, along the northern rim of the Mediterranean basin. Smaller contributions to these gene pools (constituting 13% of Y chromosomes in NW Africa and 10% of Y chromosomes in Iberia) came from the Middle East during the Neolithic and, during subsequent gene flow, from Sub-Saharan to NW Africa. Finally, bidirectional gene flow across the Strait of Gibraltar has been detected: the genetic contribution of European Y chromosomes to the NW African gene pool is estimated at 4%, and NW African populations may have contributed 7% of Iberian Y chromosomes. The Islamic rule of Spain, which began in a.d. 711 and lasted almost 8 centuries, left only a minor contribution to the current Iberian Y-chromosome pool. The high-resolution analysis of the Y chromosome allows us to separate successive migratory components and to precisely quantify each historical layer. PMID- 11254457 TI - Shwachman-Diamond syndrome with exocrine pancreatic dysfunction and bone marrow failure maps to the centromeric region of chromosome 7. AB - Shwachman-Diamond syndrome (SDS) is an autosomal recessive disorder characterized by exocrine pancreatic insufficiency and hematologic and skeletal abnormalities. A genomewide scan of families with SDS was terminated at approximately 50% completion, with the identification of chromosome 7 markers that showed linkage with the disease. Finer mapping revealed significant linkage across a broad interval that included the centromere. The maximum two-point LOD score was 8.7, with D7S473, at a recombination fraction of 0. The maximum multipoint LOD score was 10, in the interval between D7S499 and D7S482 (5.4 cM on the female map and 0 cM on the male map), a region delimited by recombinant events detected in affected children. Evidence from all 15 of the multiplex families analyzed provided support for the linkage, consistent with a single locus for SDS. However, the presence of several different mutations is suggested by the heterogeneity of disease-associated haplotypes in the candidate region. PMID- 11254460 TI - Adverse metabolic consequences of HIV protease inhibitor therapy: the search for a central mechanism. AB - Although the clinical introduction of human immunodeficiency virus (HIV) protease inhibitors (PIs) has resulted in a dramatic decline in HIV-related morbidity and mortality, it is now recognized that PI therapy is associated with serious adverse metabolic effects, including peripheral lipoatrophy, increased visceral fat, hyperlipidemia, and insulin resistance. Despite increasing awareness of this metabolic syndrome, the etiology of these side effects remains obscure. This review critically examines current mechanistic hypotheses in the context of the available experimental data. To date, a single unifying explanation for this syndrome has not been confirmed. As data accumulate, it is becoming clear that PIs lack precision in their cellular targets and it is likely that many of the side effects of these drugs are due to inhibition of a number of unrelated molecules. PMID- 11254458 TI - Homozygosity mapping places the acrodermatitis enteropathica gene on chromosomal region 8q24.3. AB - Acrodermatitis enteropathica (AE) is a rare autosomal recessive pediatric disease characterized by dermatitis, diarrhea, alopecia, and growth failure. The disease results from insufficient uptake of zinc by the intestine and can be fatal unless the diet is supplemented with zinc. To map the gene responsible for AE, a genomewide screen was performed on 17 individuals, including 4 affected individuals, in a consanguineous Jordanian family. Three markers-D8S373, D10S212, and D6S1021-had a pattern consistent with tight linkage to a recessive disease: one allele in the affected sibs and multiple alleles in unaffected sibs and parents. Two-point parametric linkage analysis using FASTLINK identified one region, D8S373, with a maximum LOD score >1.5 (1.94 at D8S373: recombination fraction.001). Twelve additional markers flanking D8S373 were used to genotype the extended family, to fine-map the AE gene. All five affected individuals including one who was not genotyped in the genomewide screen-were found to be homozygous for a common haplotype, spanning approximately 3.5 cM, defined by markers D8S1713 and D8S2334 on chromosomal region 8q24.3. To support these mapping data, seven consanguineous Egyptian families with eight patients with AE were genotyped using these markers, and six patients from five families were found to be homozygous in this region. Multipoint analysis with all consanguineous families, by Mapmaker/Homoz, resulted in a maximum LOD score of 3.89 between D8S1713 and D8S373. Sliding three-point analysis resulted in a maximum LOD score of 5.16 between markers D8S1727 and D8S1744. PMID- 11254459 TI - Correcting for a potential bias in the pedigree disequilibrium test. PMID- 11254461 TI - Effects of clenbuterol on insulin resistance in conscious obese Zucker rats. AB - The present study was conducted to determine the effect of chronic administration of the long-acting beta(2)-adrenergic agonist clenbuterol on rats that are genetically prone to insulin resistance and impaired glucose tolerance. Obese Zucker rats (fa/fa) were given 1 mg/kg of clenbuterol by oral intubation daily for 5 wk. Controls received an equivalent volume of water according to the same schedule. At the end of the treatment, rats were catheterized for euglycemic hyperinsulinemic (15 mU insulin. kg(-1). min(-1)) clamping. Clenbuterol did not change body weight compared with the control group but caused a redistribution of body weight: leg muscle weights increased, and abdominal fat weight decreased. The glucose infusion rate needed to maintain euglycemia and the rate of glucose disappearance were greater in the clenbuterol-treated rats. Furthermore, plasma insulin levels were decreased, and the rate of glucose uptake into hindlimb muscles and abdominal fat was increased in the clenbuterol-treated rats. This increased rate of glucose uptake was accompanied by a parallel increase in the rate of glycogen synthesis. The increase in muscle glucose uptake could not be ascribed to an increase in the glucose transport protein GLUT-4 in clenbuterol treated rats. We conclude that chronic clenbuterol treatment reduces the insulin resistance of the obese Zucker rat by increasing insulin-stimulated muscle and adipose tissue glucose uptake. The improvements noted may be related to the repartitioning of body weight between tissues. PMID- 11254462 TI - Evidence that amylin stimulates lipolysis in vivo: a possible mediator of induced insulin resistance. AB - The present study investigated the role of amylin in lipid metabolism and its possible implications for insulin resistance. In 5- to 7-h-fasted conscious rats, infusion of rat amylin (5 nmol/h for 4 h) elevated plasma glucose, lactate, and insulin (P <0.05 vs. control, repeated-measures ANOVA) with peak values occurring within 60 min. Despite the insulin rise, plasma nonesterified fatty acids (NEFA) and glycerol were also elevated (P < 0.001 vs. control), and these elevations (80% above basal) were sustained over the 4-h infusion period. Although unaltered in plasma, triglyceride content in liver was increased by 28% (P < 0.001) with a similar tendency in muscle (18%, P = 0.1). Infusion of the rat amylin antagonist amylin-(8-37) (125 nmol/h) induced opposite basal plasma changes to amylin, i.e., lowered plasma NEFA, glycerol, glucose, and insulin levels (all P < 0.05 vs. control); additionally, amylin-(8-37) blocked amylin-induced elevations of these parameters (P < 0.01). Treatment with acipimox (10 mg/kg), an anti-lipolytic agent, before or after amylin infusion blocked amylin's effects on plasma NEFA, glycerol, and insulin but not on glucose and lactate. We conclude that amylin could exert a lipolytic-like action in vivo that is blocked by and is opposite to effects of its antagonist amylin-(8-37). Further studies are warranted to examine the physiological implications of lipid mobilization for amylin-induced insulin resistance. PMID- 11254463 TI - Dexamethasone inhibits the stimulation of muscle protein synthesis and PHAS-I and p70 S6-kinase phosphorylation. AB - Glucocorticoids inhibit protein synthesis in muscle. In contrast, insulin and amino acids exert anabolic actions that arise in part from their ability to phosphorylate ribosomal p70 S6-kinase (p70(S6k)) and eukaryotic initiation factor (eIF)4E binding protein (BP)1 (PHAS-I), proteins that regulate translation initiation. Whether glucocorticoids interfere with this action was examined by giving rats either dexamethasone (DEX, 300 microg. kg(-1). day(-1), n = 10) or saline (n = 10) for 5 days. We then measured the phosphorylation of PHAS-I and p70(S6k) in rectus muscle biopsies taken before and at the end of a 180-min infusion of either insulin (10 mU. min(-1). kg(-1) euglycemic insulin clamp, n = 5 for both DEX- and saline-treated groups) or a balanced amino acid mixture (n = 5 for each group also). Protein synthesis was also measured during the infusion period. The results were that DEX-treated rats had higher fasting insulin, slower glucose disposal, less lean body mass, and decreased protein synthetic rates during insulin or amino acid infusion (P < 0.05 each). DEX did not affect basal PHAS-I or p70(S6k) phosphorylation but blocked insulin-stimulated phosphorylation of PHAS-I- and amino acid-stimulated phosphorylation of both PHAS-I and p70(S6k) (P < 0.01, for each). DEX also increased muscle PHAS-I concentration. These effects can, in part, explain glucocorticoid-induced muscle wasting. PMID- 11254464 TI - Preoperative oral carbohydrate treatment attenuates immediate postoperative insulin resistance. AB - Postoperative insulin resistance is a well-characterized metabolic state that has been shown to correlate with the length of postoperative stay in hospital. Preoperative intravenous or oral carbohydrate treatment has been shown to attenuate the development of postoperative insulin resistance measured 1 day after surgery. To study the effects of preoperative oral carbohydrate treatment on postoperative changes in insulin resistance and substrate utilization, in the absence of postoperative confounding factors, 15 patients were double-blindly treated with either a carbohydrate-rich beverage (12.5%) (n = 8) or placebo (n = 7) before undergoing total hip replacement surgery. Insulin sensitivity, endogenous glucose release, and substrate oxidation rates were measured before and immediately after surgery. Whole body insulin sensitivity decreased by 18% in the treatment group vs. 43% in the placebo group (P < 0.05, Student's t-test for unpaired data). In both groups, the major mechanism of insulin resistance was an inhibition of insulin-induced nonoxidative glucose disposal after surgery. The better preservation of insulin sensitivity in the treatment group was attributable to a less reduced glucose disposal in peripheral tissues and increased glucose oxidation rates. PMID- 11254465 TI - Local inhibition of nitric oxide temporarily stimulates aldosterone secretion in conscious sheep in vivo. AB - The effect of localized blockage of endogenous nitric oxide (NO) on basal aldosterone secretion was studied in conscious sheep with autotransplanted adrenal glands. We have shown that infusion of the NO synthase (NOS) inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME; 130 microg/l blood flow) significantly stimulated basal aldosterone secretion rate (ASR). This stimulatory effect was seen up to 4 h of infusion. Beyond this time point, however, the elevated ASR level was not sustained, and it was seen to drop markedly to lower than control values at 5 h. L-NAME had no effect on cortisol secretion rate (FSR) during the first 4 h of infusion, but a significant reduction in FSR was seen by the 8-h time point. Adrenal blood flow was consistently decreased in association with long L-NAME infusion. Additionally, L-NAME was shown to have no effect on aldosterone secretion when infused systemically. We conclude that the relationship between NO and aldosterone secretion is an inhibitory one, in which NO seems to have a negative effect on basal aldosterone secretion. PMID- 11254466 TI - Plasma protein synthesis in patients with low-grade nephrotic proteinuria. AB - Overt nephrotic syndrome is characterized by albumin and fibrinogen hyperproduction and reduced very low density lipoprotein apolipoprotein B-100 (VLDL apoB-100) clearance. Whether similar changes also occur in low-grade proteinuria is not known. Thus we measured albumin, fibrinogen, and VLDL apoB-100 kinetics in six patients with modest proteinuria and normal creatinine clearance (P) and in ten control subjects (C) by leucine tracer infusion and precursor product relationships. In P, plasma albumin concentration was decreased (P < 0.003), whereas concentrations of fibrinogen and VLDL apoB-100 were increased (P < 0.001). In P, albumin fractional secretion rate (FSR) was increased (P < 0.01), fibrinogen FSR was normal, and VLDL apoB-100 FSR was decreased (P < 0.03). As a result, in P, absolute secretion rates (ASR) of albumin and fibrinogen were increased (P < 0.03), whereas VLDL apoB-100 ASR was normal. Albumin FSR was inversely correlated to oncotic pressure in P but not in C. These findings suggest that low-grade nephrotic proteinuria is characterized by simultaneous multiple alterations in turnover rates of albumin, fibrinogen, and VLDL apoB-100. Their pathogenesis, however, appears to be multifactorial. PMID- 11254467 TI - Flux control in the rat gastrocnemius glycogen synthesis pathway by in vivo 13C/31P NMR spectroscopy. AB - To determine the relative contributions of glucose transport/hexokinase, glycogen synthase (GSase), and glycolysis to the control of insulin-stimulated muscle glycogen synthesis, we combined 13C and 31P NMR to quantitate the glycogen synthesis rate and glucose 6-phosphate (G-6-P) levels in rat (Sprague-Dawley) gastrocnemius muscle during hyperinsulinemia at euglycemic (E) and hyperglycemic (H) glucose concentrations under thiopental anesthesia. Flux control was calculated using metabolic control analysis. The combined control coefficient of glucose transport/hexokinase (GT/Hk) for glycogen synthesis was 1.1 +/- 0.03 (direct measure) and 1.14-1.16 (calculated for a range of glycolytic fluxes), whereas the control coefficient for GSase was much lower (0.011-0.448). We also observed that the increase in in vivo [G-6-P] from E to H (0.22 +/- 0.03 to 0.40 +/- 0.03 mM) effects a supralinear increase in the in vitro velocity of GSase, from 14.6 to 26.1 mU. kg(-1). min(-1) (1.8-fold). All measurements suggest that the majority of the flux control of muscle glycogen synthesis is at the GT/Hk step. PMID- 11254468 TI - Acetylcholine increases intracellular Ca2+ in the rat pituitary folliculostellate cells in primary culture. AB - Pituitary folliculostellate cells (FSCs) are thought to partially inhibit pituitary hormone secretion through a paracrine mechanism. In this process, one of the important questions is what factors regulate the function of FSCs. Because ACh is synthesized in and possibly released from the corticotrophs and lactotrophs, we examined whether FSCs respond to ACh by the method of Ca2+ imaging in primary cultured FSCs from male Wistar rats. ACh (30 nM-3 microM) increased intracellular calcium concentration ([Ca2+](i)) of FSCs in a concentration-dependent manner, with an initial rapid rise followed by a relatively sustained increase. The complete block of the response by atropine and pirenzepine suggests involvement of muscarinic receptors. Depletion of the stored Ca2+ by thapsigargin blocked the response completely. Blockers of phospholipase C, U-73122 and neomycin, suppressed significantly the rise of [Ca2+](i). These results suggest that ACh increases [Ca2+](i) in FSCs by activating phospholipase C, presumably through activation of M(1) receptors. The rise in [Ca2+](i) could trigger a variety of Ca2+-dependent cellular processes, including the synthesis and release of bioactive substances, which in turn act on endocrine cells. PMID- 11254469 TI - Cortisol and GH secretory dynamics, and their interrelationships, in healthy aged women and men. AB - We studied 130 healthy aged women (n = 57) and men (n = 73), age 65-88 yr, with age-related reductions in insulin-like growth factor I and gonadal steroid levels to assess the interrelationships between cortisol and growth hormone (GH) secretion and whether these relationships differ by sex. Blood was sampled every 20 min from 8:00 PM to 8:00 AM; cortisol was measured by RIA and GH by immunoradiometric assay, followed by deconvolution analyses of hormone secretory parameters and assessment of approximate entropy (ApEn) and cross-ApEn. Cortisol mass/burst, cortisol production rate, and mean and integrated serum cortisol concentrations (P < 0.0005), and overnight basal GH secretion (P < 0.05), were elevated in women vs. men. Integrated cortisol concentrations were directly related to most measures of GH secretion in women (P < 0.01) and with mean and integrated GH concentrations in men (P < 0.05). Integrated GH concentrations were directly related to mean and integrated cortisol levels in women (P < 0.005) and men (P < 0.05), with no sex differences. There were no sex differences in cortisol or GH ApEn values; however, the cross-ApEn score was greater in women (P < 0.05), indicating reduced GH-cortisol pattern synchrony in aged women vs. men. There were no significant relationships of integrated cortisol secretion with GH ApEn, or vice versa, in either sex. Thus postmenopausal women appear to maintain elevated cortisol production in patterns that are relatively uncoupled from those of GH, whereas mean hormone outputs remain correlated. PMID- 11254470 TI - Differential GH-releasing hormone regulation of GHRH receptor mRNA expression in the rat pituitary. AB - To understand the capacity of growth hormone-releasing hormone (GHRH) to regulate expression of the GHRH receptor, we studied the effects of GHRH on GHRH receptor mRNA expression in immature and adult rats by use of pituitary cell culture and immunoneutralization approaches. Pituitary cell cultures from neonatal (2-day old) and adult (70-day-old) rats were treated with GHRH for 4, 24, or 72 h. The effect of GHRH on GHRH receptor mRNA expression depended on the duration of GHRH exposure in both age groups; short-term (4 h) GHRH treatment significantly reduced GHRH receptor mRNA expression (P < 0.05), whereas intermediate treatment (24 h) restored GHRH receptor mRNA to basal levels, and long-term treatment (72 h) stimulated GHRH receptor mRNA expression (P < 0.02). The long-term stimulatory effect of GHRH on GHRH receptor mRNA expression required the presence of serum in the culture medium, and, in the absence of serum, the stimulatory effect was completely abolished. Moreover, the capacity of the pituitary to increase GHRH receptor mRNA expression in response to 72-h GHRH treatment was age dependent, with neonatal pituitaries exhibiting a much greater stimulatory effect than adult pituitaries (P < 0.025). Immunoneutralization of endogenous GHRH significantly reduced GHRH receptor mRNA expression in neonatal (P < 0.004), juvenile (P < 0.003), and mature (P < 0.004) pituitaries compared with age-matched controls. Taken together, these results indicate that GHRH is a potent regulator of GHRH receptor gene expression in immature and mature pituitaries; however, the nature and direction of GHRH regulation of its receptor depend significantly on several variables, including the duration of GHRH exposure, the presence of permissive components in serum, and the developmental stage of the pituitary. PMID- 11254471 TI - Elevated intramyocellular lipid concentration in obese subjects is not reduced after diet and exercise training. AB - To determine the effects of weight loss on intramyocellular energy substrates, vastus lateralis muscle biopsies were taken from six obese subjects (body mass index 34 +/- 5 kg/m(2)) before, after 15 wk of energy restriction (ER; -700 kcal/day), and after a further average 20.7 +/- 1.6 wk of endurance training plus low-fat diet (ET-LFD). Body weight fell from 100 +/- 6 to 89 +/- 6 kg during ER and to 84 +/- 4 kg after ET-LFD. Lipids and glycogen were histochemically measured in type I, IIA, and IIB fibers. Total muscle glycogen content (MGC; per 100 fibers) decreased after ER [from 72 +/- 13 to 55 +/- 8 arbitrary units (AU)]. A similar but not significant decrease was seen in total muscle lipid content (MLC; 14 +/- 5 to 9 +/- 1 AU). After ET-LFD, MGC returned to initial values (74 +/- 8 AU), and MLC approached near-initial values (12 +/- 3 AU). Individual fiber lipid concentration did not change throughout the protocol in all fiber types, whereas glycogen concentration increased after ET-LFD. The training effects of ET LFD were measured as increasing activities of key mitochondrial enzymes. Although total muscle energy reserves can be reduced after weight loss, their concentration within individual myofibers remains elevated. Weight loss does not appear sufficient to correct the potential detrimental effects of high intracellular lipid concentrations. PMID- 11254472 TI - Mineralocorticoid and glucocorticoid receptors inhibit UCP expression and function in brown adipocytes. AB - Uncoupling proteins (UCP), specific mitochondrial proton transporters that function by uncoupling oxidative metabolism from ATP synthesis, are involved in thermoregulation and control of energy expenditure. The hibernoma-derived T37i cells, which possess functional endogenous mineralocorticoid receptors (MR), can undergo differentiation into brown adipocytes. In differentiated T37i cells, UCP1 mRNA levels increased 10- to 20-fold after retinoic acid or beta-adrenergic treatment. Interestingly, UCP2 and UCP3 mRNA was also detected. Aldosterone treatment induced a drastic decrease in isoproterenol- and retinoic acid stimulated UCP1 mRNA levels in a time- and dose-dependent manner (IC(50) approximately 1 nM aldosterone). This inhibition was unaffected by cycloheximide and did not modify UCP1 mRNA stability (half-life time = 5 h), indicating that it occurs at the transcriptional level. It involves both the MR and/or the glucocorticoid receptor (GR), depending on the retinoic or catecholamine induction pathway. Basal UCP3 expression was also significantly reduced by aldosterone, whereas UCP2 mRNA levels were not modified. Finally, as demonstrated by JC1 aggregate formation in living cells, aldosterone restored mitochondrial membrane potential abolished by isoproterenol or retinoic acid. Our results demonstrate that MR and GR inhibit expression of UCP1 and UCP3, thus participating in the control of energy expenditure. PMID- 11254473 TI - Time course changes in IGFBP-1 after treadmill exercise and postexercise food intake in rats. AB - Prolonged exercise increases circulating insulin-like growth factor binding protein-1 (IGFBP-1) in humans and animals, but its physiological significance is unknown. This study examined 1) time-course changes in plasma IGFBP-1 and hepatic IGFBP-1 mRNA expression after exercise, 2) changes in IGFBP-1 in relation to plasma glucose, insulin, and IGF-I, and 3) the impact of feeding a postexercise meal on the IGFBP-1 response. Food-deprived male rats were vigorously run on a treadmill and compared with nonexercised controls at 15 min and 1, 4, 8, and 12 h after exercise. Circulating insulin concentrations in exercised rats were lower than in controls at 15 min and 1 h, whereas plasma glucose and IGF-I remained unaffected. Circulating and hepatic expression of IGFBP-1 was markedly increased above that of controls at 15 min, 1 h, and 12 h. In a separate experiment, one half of the exercised animals received a nutritionally complete meal immediately after the experimental run. The meal elevated plasma insulin and glucose concentrations at 15 min and 1 h. Despite this change in nutritional status, serum IGFBP-1 concentrations and hepatic IGFBP-1 abundance remained elevated at 15 min and 1 h. These results demonstrate that the IGFBP-1 response to a single bout of treadmill exercise is short in duration and independent of insulin, glucose, and amino acid availability. PMID- 11254474 TI - Autoregulation of glucose production in men with a glycerol load during rest and exercise. AB - Related to hepatic autoregulation we evaluated hypotheses that 1) glucose production would be altered as a result of a glycerol load, 2) decreased glucose recycling rate (Rr) would result from increased glycerol uptake, and 3) the absolute rate of gluconeogenesis (GNG) from glycerol would be positively correlated to glycerol rate of disappearance (R(d)) during a glycerol load. For these purposes, glucose and glycerol kinetics were determined in eight men during rest and during 90 min of leg cycle ergometry at 45 and 65% of peak O2 consumption (.VO2 (peak)). Trials were conducted after an overnight fast, with exercise commencing 12 h after the last meal. Subjects received a continuous infusion of [6,6-(2)H(2)]glucose, [1-(13)C]glucose, and [1,1,2,3,3 (2)H(5)]glycerol without (CON) or with an additional 1,000 mg (rest: 20 mg/min; exercise: 40 mg/min) of [2-(13)C]- or unlabeled glycerol added to the infusate (GLY). Infusion of glycerol dampened glucose Rr, calculated as the difference between [6,6-(2)H(2)]- and [1-(13)C]glucose rates of appearance (R(a)), at rest [0.35 +/- 0.12 (CON) vs. 0.12 +/- 0.10 mg. kg(-1). min(-1) (GLY), P < 0.05] and during exercise at both intensities [45%: 0.63 +/- 0.14 (CON) vs. 0.04 +/- 0.12 (GLY); 65%: 0.73 +/- 0.14 (CON) vs. 0.04 +/- 0.17 mg. kg(-1). min(-1) (GLY), P < 0.05]. Glucose R(a) and oxidation were not affected by glycerol infusion at rest or during exercise. Throughout rest and both exercise intensities, glycerol R(d) was greater in GLY vs. CON conditions (rest: 0.30 +/- 0.04 vs. 0.58 +/- 0.04; 45%: 0.57 +/- 0.07 vs. 1.19 +/- 0.04; 65%: 0.73 +/- 0.06 vs. 1.27 +/- 0.05 mg. kg(-1). min(-1), CON vs. GLY, respectively). Differences in glycerol R(d) (DeltaR(d)) between protocols equaled the unlabeled glycerol infusion rate and correlated with plasma glycerol concentration (r = 0.97). We conclude that infusion of a glycerol load during rest and exercise at 45 and 65% of .VO2(peak) 1) does not affect glucose R(a) or R(d), 2) blocks glucose Rr, 3) increases whole body glycerol R(d) in a dose-dependent manner, and 4) results in gluconeogenic rates from glycerol equivalent to CON glucose recycling rates. PMID- 11254475 TI - Glutamine supplementation promotes anaplerosis but not oxidative energy delivery in human skeletal muscle. AB - The aims of the present study were twofold: first to investigate whether TCA cycle intermediate (TCAI) pool expansion at the onset of moderate-intensity exercise in human skeletal muscle could be enhanced independently of pyruvate availability by ingestion of glutamine or ornithine alpha-ketoglutarate, and second, if it was, whether this modification of TCAI pool expansion had any effect on oxidative energy status during subsequent exercise. Seven males cycled for 10 min at approximately 70% maximal O2) uptake 1 h after consuming either an artificially sweetened placebo (5 ml/kg body wt solution, CON), 0.125 g/kg body wt L-(+)-ornithine alpha-ketoglutarate dissolved in 5 ml/kg body wt solution (OKG), or 0.125 g/kg body wt L-glutamine dissolved in 5 ml/kg body wt solution (GLN). Vastus lateralis muscle was biopsied 1 h postsupplement and after 10 min of exercise. The sum of four measured TCAI (SigmaTCAI; citrate, malate, fumarate, and succinate, approximately 85% of total TCAI pool) was not different between conditions 1 h postsupplement. However, after 10 min of exercise, SigmaTCAI (mmol/kg dry muscle) was greater in the GLN condition (4.90 +/- 0.61) than in the CON condition (3.74 +/- 0.38, P < 0.05) and the OKG condition (3.85 +/- 0.28). After 10 min of exercise, muscle phosphocreatine (PCr) content was significantly reduced (P < 0.05) in all conditions, but there was no significant difference between conditions. We conclude that the ingestion of glutamine increased TCAI pool size after 10 min of exercise most probably because of the entry of glutamine carbon at the level of alpha-ketoglutarate. However, this increased expansion in the TCAI pool did not appear to increase oxidative energy production, because there was no sparing of PCr during exercise. PMID- 11254476 TI - V. Stress and irritable bowel syndrome. AB - Different types of stress play important roles in the onset and modulation of irritable bowel syndrome (IBS) symptoms. The physiological effects of psychological and physical stressors on gut function and brain-gut interactions are mediated by outputs of the emotional motor system in terms of autonomic, neuroendocrine, attentional, and pain modulatory responses. IBS patients show an enhanced responsiveness of this system manifesting in altered modulation of gastrointestinal motility and secretion and in alterations in the perception of visceral events. Functional brain imaging techniques are beginning to identify brain circuits involved in the perceptual alterations. Animal models have recently been proposed that mimic key features of the human syndrome. PMID- 11254478 TI - Swallow-related cerebral cortical activity maps are not specific to deglutition. AB - Cortical representation of swallow-related motor tasks has not been systematically investigated. In this study, we elucidated and compared these cortical representations to those of volitional swallow using block-trial and single-trial methods. Fourteen volunteers were studied by functional magnetic resonance imaging. Cortical activation during both swallowing and swallow-related motor tasks that can be performed independent of swallowing, such as jaw clenching, lip pursing, and tongue rolling, was found in four general areas: the anterior cingulate, motor/premotor cortex, insula, and occipital/parietal region corresponding to Brodmann's areas 7, 19, and 31. Regions of activity, volume of activated voxels, and increases in signal intensity were found to be similar between volitional swallow and swallow-related motor tasks. These findings, using both block-trial and single-trial techniques, suggest that cerebral cortical regions activated during swallowing may not be specific to deglutitive function. PMID- 11254477 TI - IV. Helicobacter pylori strain-specific activation of signal transduction cascades related to gastric inflammation. AB - Helicobacter pylori strains that possess the cag pathogenicity island induce more severe gastritis and augment the risk of developing peptic ulcer disease and distal gastric cancer. A specific mechanism by which cag(+) strains may enhance gastritis is strain-selective regulation of interleukin (IL)-8 production. On contact with gastric epithelial cells, H. pylori activates multiple signal transduction cascades that regulate IL-8 secretion, including nuclear factor kappaB and mitogen-activated protein kinases, and these events are dependent on genes within the cag island. An independent effect of cag-mediated cellular contact is translocation and phosphorylation of H. pylori proteins within the host epithelial cell. The redundancy of intracellular signaling cascades activated by H. pylori and the divergent epithelial cell responses induced by components of the cag island may contribute to the ability of this organism to persist for decades within the gastric niche. PMID- 11254479 TI - Endogenous corticosteroids modulate Clostridium difficile toxin A-induced enteritis in rats. AB - We examined the role of glucocorticoids in acute inflammatory diarrhea mediated by Clostridium difficile toxin A. Toxin A (5 microg) or buffer was injected in rat ileal loops, and intestinal responses were measured after 30 min to 4 h. Ileal toxin A administration increased plasma glucocorticoids after 1 h, at which time the toxin-stimulated secretion was not significant. Administration of the glucocorticoid analog dexamethasone inhibited toxin A-induced intestinal secretion and inflammation and downregulated toxin A-mediated increase of macrophage inflammatory protein-2. Adrenalectomy followed by replacement with glucocorticoids at various doses suggested that intestinal responses to toxin A were related to circulating levels of glucocorticoids. Administration of the glucocorticoid receptor antagonist RU-486 enhanced toxin A-mediated intestinal secretion and inflammation. We conclude that C. difficile toxin A causes increased secretion of endogenous glucocorticoids, which diminish the intestinal secretory and inflammatory effects of toxin A. PMID- 11254480 TI - Different types of contractions in rat colon and their modulation by oxidative stress. AB - The aim of this study was to investigate the modulation of in vitro rat colonic circular muscle contractions by dextran sodium sulfate (DSS)-induced inflammation and in spontaneous inflammation in HLA-B27 rats. We also examined the potential role of hydrogen peroxide (H(2)O(2)) in modulating excitation-contraction coupling. The muscle strips from the middle colon generated spontaneous phasic contractions and giant contractions (GCs), the proximal colon strips generated primarily phasic contractions, and the distal colon strips were mostly quiescent. The spontaneous phasic contractions and GCs were not affected by inflammation, but the response to ACh was suppressed in DSS-treated rats and in HLA-B27 rats. H(2)O(2) production was increased in the muscularis of the inflamed colon. Incubation of colonic muscle strips with H(2)O(2) suppressed the spontaneous phasic contractions and concentration and time dependently reduced the response to ACh; in the middle colon, it also increased the frequency of GCs. We conclude that H(2)O(2) mimics the suppression of the contractile response to ACh in inflammation. H(2)O(2) also selectively suppresses phasic contractions and increases the frequency of GCs, as found previously in inflamed dog and human colons. PMID- 11254481 TI - Endothelin ET(A) and ET(B) receptors in postnatal intestine. AB - We aimed to characterize endothelin (ET) receptors in the swine intestinal vasculature and to determine ischemia-reperfusion (I/R) effects on these receptors. Saturation and competitive binding assays were performed on mesenteric artery protein membranes from 1- and 40-day-old animals, both control and those subjected to 1 h of partial ischemia followed by 6 h of reperfusion in vivo. Scatchard analysis of saturation binding with (125)I-labeled ET-1 in membranes from endothelium-denuded (E(-)) vessels revealed that the maximum number of binding sites was greater in younger animals. Competitive (125)I-ET-1 binding was significant for a one-site model with ET-1, ET-3, and sarafotoxin S6c (S6c) in membranes from endothelium-intact (E(+)) and E(-) vessels in both age groups. The maximum number of ET-1 binding sites was significantly greater in younger animals. In the presence of the ET(A) receptor antagonist BQ-123, competitive (125)I-ET-1 binding was significant for a one-site model with ET-1 and S6c in membranes from E(+) vessels in both age groups. The maximum number of ET-1 binding sites was significantly greater in younger animals. After I/R, the maximum number of ET-1 binding sites was unchanged. In the presence of BQ-123, specific binding by ET-1 and S6c was eliminated in both age groups after I/R. These results suggest that both ET receptor populations are expressed to a greater degree in younger animals and I/R significantly affects the ET(B) receptor. PMID- 11254482 TI - Immunolocalization, ontogeny, and regulation of microsomal triglyceride transfer protein in human fetal intestine. AB - To examine the multiple stages of lipoprotein packaging during development, we studied localization, ontogeny, and regulation of microsomal transfer protein (MTP), a crucial protein for lipid transport. With the use of immunofluorescence, MTP was identified in villus and crypt epithelial cells in different regions of human fetal intestine, including colon. Staining was detected as early as the 13th wk of gestation in all gut segments and was almost entirely confined to the columnar epithelial cells of the jejunum and colon. Unlike immunofluorescence, which provides qualitative but not quantitative information on MTP signal, enzymatic assays revealed a decreasing gradient from proximal small intestine to distal, as confirmed by immunoblot. Activity of MTP in small intestinal explants cultured for different incubation periods (0, 4, 8, and 24 h) peaked at 4 h but remained insensitive to different concentrations of oleic acid. Also, a trend toward increasing MTP activity was observed at 20-22 wk of gestation. Finally, in strong contrast to jejunal efficiency, colonic explants displayed impaired lipid production, apolipoprotein biogenesis, and lipoprotein assembly, in association with poor expression of MTP. These findings provide the first evidence that human fetal gut is able to express MTP and emphasize the distinct regional distribution, regulation by oleic acid, and ontogeny of MTP. PMID- 11254483 TI - The role of protein kinase C isozymes in TNF-alpha-induced cytotoxicity to a rat intestinal epithelial cell line. AB - Tumor necrosis factor (TNF)-alpha can induce cytotoxicity and apoptosis in a number of cell types and has been implicated in the regulation of many inflammatory processes. It has been suggested that protein kinase C (PKC) is one of the intracellular mediators of the actions of TNF-alpha. In the present study, the role of PKC isoforms in TNF-alpha-mediated cytotoxicity and apoptosis in intestinal cells was investigated using the rat epithelial cell line, IEC-18. Cells were incubated with TNF-alpha in the presence or absence of the transcription inhibitor actinomycin D (AMD). The extent of cell damage was enhanced when AMD was added to incubation medium, suggesting that new protein synthesis plays a role in the cytotoxic action of TNF. TNF-alpha also induced the translocation of PKC-alpha, -delta, and -epsilon from cytosol to the membrane fraction of the intestinal cells. Furthermore, the cytotoxic and apoptotic effects of TNF were reduced by pretreating the cells with the PKC-epsilon translocation inhibitor, PKC-epsilonV1-2. In contrast, although cells incubated with the phorbol ester phorbol 12-myristate 13-acetate (PMA) also displayed an increase in cell injury, the extent of cytotoxicity and apoptosis was not enhanced by AMD. Furthermore, PMA-induced cell damage was reduced by rottlerin, a PKC-delta inhibitor. Caspase-3, an enzyme implicated in the mediation of apoptosis, was activated in cells in response to either TNF-alpha or PMA stimulation, and its effects on this activity were reduced by selective inhibition of PKC-epsilon and -delta, respectively. Furthermore, inhibition of caspase-3 activity reduced apoptosis. These data suggest that activation of selective PKC isoforms mediate the effects of TNF-alpha on intestinal epithelial cell injury. PMID- 11254484 TI - Arterial compliance in patients with cirrhosis: stroke volume-pulse pressure ratio as simplified index. AB - Arterial function may be altered in patients with cirrhosis. We determined compliance of the arterial tree (C(1)) in relation to systemic and splanchnic hemodynamic derangement and clinical variables. C(1) and the stroke volume-pulse pressure index (SV/PP) were significantly higher (+62% and +40%, respectively; P < 0.001) in cirrhotic patients (n = 49) than in control subjects (n = 19), and a close correlation between C(1) and SV/PP was found in both cirrhotic patients (r = 0.86, P < 0.001) and control subjects (r = 0.96, P < 0.001). Univariate analysis showed significant relations between C(1) and SV/PP on one side and age, sex, body weight, portal pressure, systemic hemodynamics, biochemical variables, and severity of disease on the other. In the multiple-regression analysis, sex, age, mean arterial blood pressure, systemic vascular resistance, and biochemical variables were significant independent predictors of SV/PP (P < 0.005-0.00001). In conclusion, arterial compliance is elevated in cirrhosis. A simplified SV/PP index seems to reflect abnormalities in the arterial compliance of these patients. PMID- 11254485 TI - Roles of 5-HT receptors in the release and action of secretin on pancreatic secretion in rats. AB - 5-Hydroxytryptamine (serotonin, 5-HT) is a hormone and neurotransmitter regulating gastrointestinal functions. 5-HT receptors are widely distributed in gastrointestinal mucosa and the enteric nervous system. Duodenal acidification stimulates not only the release of both 5-HT and secretin but also pancreatic exocrine secretion. We investigated the effect of 5-HT receptor antagonists on the release of secretin and pancreatic secretion of water and bicarbonate induced by duodenal acidification in anesthetized rats. Both the 5-HT(2) receptor antagonist ketanserin and the 5-HT(3) receptor antagonist ondansetron at 1-100 microg/kg dose-dependently inhibited acid-induced increases in plasma secretin concentration and pancreatic exocrine secretion. Neither the 5-HT(1) receptor antagonists pindolol and 5-HTP-DP nor the 5-HT(4) receptor antagonist SDZ-205,557 affected acid-evoked release of secretin or pancreatic secretion. None of the 5 HT receptor antagonists affected basal pancreatic secretion or plasma secretin concentration. Ketanserin or ondansetron at 10 microg/kg or a combination of both suppressed the pancreatic secretion in response to intravenous secretin at 2.5 and 5 pmol x kg(-1) x h(-1) by 55-75%, but not at 10 pmol x kg(-1) x h(-1). Atropine (50 microg/kg) significantly attenuated the inhibitory effect of ketanserin on pancreatic secretion but not on the release of secretin. These observations suggest that 5-HT(2) and 5-HT(3) receptors mediate duodenal acidification-induced release of secretin and pancreatic secretion of fluid and bicarbonate. Also, regulation of pancreatic exocrine secretion through 5-HT(2) receptors may involve a cholinergic pathway in the rat. PMID- 11254486 TI - Sulfate and chloride transport in Caco-2 cells: differential regulation by thyroxine and the possible role of DRA gene. AB - The current studies were undertaken to establish an in vitro cellular model to study the transport of SO and Cl(-) and hormonal regulation and to define the possible function of the downregulated in adenoma (DRA) gene. Utilizing a postconfluent Caco-2 cell line, we studied the OH(-) gradient-driven (35)SO and (36)Cl(-) uptake. Our findings consistent with the presence of an apical carrier mediated (35)SO/OH(-) exchange process in Caco-2 cells include: 1) demonstration of saturation kinetics [Michaelis-Menten constant (K(m)) of 0.2 +/- 0.08 mM for SO and maximum velocity of 1.1 +/- 0.2 pmol x mg protein(-1) x 2 min(-1)]; 2) sensitivity to inhibition by DIDS (K(i) = 0.9 +/- 0.3 microM); and 3) competitive inhibition by oxalate and Cl(-) but not by nitrate and short chain fatty acids, with a higher K(i) (5.95 +/- 1 mM) for Cl(-) compared with oxalate (K(i) = 0.2 +/ 0.03 mM). Our results also suggested that the SO/OH(-) and Cl(-)/OH(-) exchange processes in Caco-2 cells are distinct based on the following: 1) the SO/OH(-) exchange was highly sensitive to inhibition by DIDS compared with Cl(-)/OH(-) exchange activity (K(i) for DIDS of 0.3 +/- 0.1 mM); 2) Cl(-) competitively inhibited the SO/OH(-) exchange activity with a high K(i) compared with the K(m) for SO, indicating a lower affinity for Cl(-); 3) DIDS competitively inhibited the Cl(-)/OH(-) exchange process, whereas it inhibited the SO/OH(-) exchange activity in a mixed-type manner; and 4) utilizing the RNase protection assay, our results showed that 24-h incubation with 100 nM of thyroxine significantly decreased the relative abundance of DRA mRNA along with the SO/OH(-) exchange activity but without any change in Cl(-)/OH(-) exchange process. In summary, these studies demonstrated the feasibility of utilizing Caco-2 cell line as a model to study the apical SO/OH(-) and Cl(-)/OH(-) exchange processes in the human intestine and indicated that the two transporters are distinct and that DRA may be predominantly a SO transporter with a capacity to transport Cl(-) as well. PMID- 11254487 TI - Role of cholecystokinin in the intestinal phase of pancreatic circulation in dogs. AB - The regulatory mechanisms of postprandial pancreatic hyperemia are not well characterized. The aim of this study is to clarify the role of cholecystokinin (CCK) in the intestinal phase of pancreatic circulation. Pancreatic, gastric, and intestinal blood flows were measured by ultrasound transit-time blood flowmeters in five conscious dogs. Pancreatic and gastric secretion and blood pressure were also monitored. Synthetic CCK octapeptide (CCK-8) or gastrin heptadecapeptide (gastrin-17) was infused intravenously, and milk was infused into the duodenum with or without loxiglumide, a specific CCK-A receptor antagonist. CCK-8 induced dose-related increases of pancreatic, but not gastric or intestinal, blood flow and protein secretion without affecting systemic blood pressure. Gastrin-17 did not affect pancreatic blood flow. An intraduodenal infusion of milk increased pancreatic and intestinal blood flows and pancreatic protein secretion. Loxiglumide completely inhibited pancreatic blood flow and protein responses to CCK-8 and milk but not the intestinal blood flow response. CCK is a potent and specific pancreatic vasodilator, with its effect mediated by CCK-A receptors. CCK plays an important role in the regulation of the intestinal phase of the pancreatic circulation in dogs. PMID- 11254488 TI - Lactase synthesis is pretranslationally regulated in protein-deficient pigs fed a protein-sufficient diet. AB - The in vivo effects of protein malnutrition and protein rehabilitation on lactase phlorizin hydrolase (LPH) synthesis were examined. Five-day-old pigs were fed isocaloric diets containing 10% (deficient, n = 12) or 24% (sufficient, n = 12) protein. After 4 wk, one-half of the animals in each dietary group were infused intravenously with [(13)C(1)]leucine for 6 h, and the jejunum was analyzed for enzyme activity, mRNA abundance, and LPH polypeptide isotopic enrichment. The remaining animals were fed the protein-sufficient diet for 1 wk, and the jejunum was analyzed. Jejunal mass and lactase enzyme activity per jejunum were significantly lower in protein-deficient vs. control animals but returned to normal with rehabilitation. Protein malnutrition did not affect LPH mRNA abundance relative to elongation factor-1alpha, but rehabilitation resulted in a significant increase in LPH mRNA relative abundance. Protein malnutrition significantly lowered the LPH fractional synthesis rate (FSR; %/day), whereas the FSR of LPH in rehabilitated and control animals was similar. These results suggest that protein malnutrition decreases LPH synthesis by altering posttranslational events, whereas the jejunum responds to rehabilitation by increasing LPH mRNA relative abundance, suggesting pretranslational regulation. PMID- 11254489 TI - Ambulatory 24-h colonic manometry in healthy humans. AB - Our aim was to investigate motor activity of the healthy, relatively unprepared colon in the ambulatory state. Twenty-five age- and gender-matched adults had a six-sensor solid-state probe inserted into the proximal transverse colon without sedation. Subjects ambulated freely and ate standard meals. In 528 h of recording, we found a lower (P < 0.05) area under the curve during the night. Waking induced a threefold increase in motility, whereas meals induced a twofold increase. Women showed less activity (P < 0.05) in the transverse/descending colon than men. The transverse/descending colon showed more (P < 0.05) activity than the rectosigmoid colon. Seven patterns were recognized; predominantly, they were simultaneous, propagated, or periodic bursts of 3-cycles/min (cpm) waves. A specialized propagating pressure wave with a high amplitude (>105 mmHg) and a prolonged duration (>14 s) occurred in all subjects (mean 10/day), mostly after waking, after meals, or with defecation. A 3-cpm motor activity was seen in the rectosigmoid region predominantly at night. The colon exhibits a wide spectrum of pressure activity around the clock, with gender and regional differences and circadian rhythm. This comprehensive study provides qualitative and quantitative normative data for colonic manometry. PMID- 11254490 TI - Mast cell-independent impairment of host defense and muscle contraction in T. spiralis-infected W/W(V) mice. AB - In response to nematode infection, the host presumably attempts to create an unfavorable environment to prevent larval penetration of the host and to expedite parasite expulsion from the gut. In this study, we have used W/W(V) mice with or without mast cells after bone marrow reconstitution (BMR-W/W(V)) to examine the role of mast cells in the host response. W/W(V), BMR-W/W(V), and wild-type (+/+) mice were infected with Trichinella spiralis. Infected W/W(V) mice exhibited less tissue damage and experienced a delay in worm expulsion and a greater degree of larval penetration of the gut leading to encystment in skeletal muscle. Tissue injury was greater and worm expulsion was normalized in BMR-W/W(V) mice, but larval penetration remained unchanged. Spontaneous contractile activity of jejunal muscle was disrupted in W/W(V) mice, as was the contractile response to carbachol. These abnormalities were also present in BMR-W/W(V) mice. These results indicate that mast cells mediate tissue damage and contribute to the timely expulsion of nematodes from the gut during primary infection. PMID- 11254491 TI - Inflammation enhances reflex and spinal neuron responses to noxious visceral stimulation in rats. AB - To improve understanding of sensory processes related to visceral inflammation, the effect of turpentine-induced inflammation on reflex (cardiovascular/visceromotor) and extracellularly recorded lumbosacral dorsal horn neuron responses to colorectal distension (CRD) was investigated. A 25% solution of turpentine, applied to the colorectal mucosa, produced inflammation, decreased compliance of the colonic wall, and enhanced reflex responses in unanesthetized rats within 2-6 h. At 24 h posttreatment, pressor responses to CRD (80 mmHg, 20 s) were 20% greater, and intraluminal pressures needed to evoke visceromotor reflexes were 30% lower than controls. Parallel electrophysiological experiments in spinal cord-transected, decerebrate rats demonstrated that two neuronal subgroups excited by CRD were differentially affected by turpentine administered 24 h before testing. During CRD, abrupt neurons were 70% less active and sustained neurons were 25% more active than similar neurons in controls. In summary, reflex and neuronal subgroup (sustained neurons) responses to CRD were both potentiated by chemical inflammation. This suggests that the neurophysiological basis for inflammation-induced increases in reflex responses to CRD is increased activity of this neuronal subgroup. PMID- 11254492 TI - GABA(B) receptors on vagal afferent pathways: peripheral and central inhibition. AB - To investigate GABA(B) receptors along vagal afferent pathways, we recorded from vagal afferents, medullary neurons, and vagal efferents in ferrets. Baclofen (7 14 micromol/kg i.v.) reduced gastric tension receptor and nucleus tractus solitarii neuronal responses to gastric distension but not gastroduodenal mucosal receptor responses to cholecystokinin (CCK). GABA(B) antagonists CGP-35348 or CGP 62349 reversed effects of baclofen. Vagal efferents showed excitatory and inhibitory responses to distension and CCK. Baclofen (3 nmol i.c.v. or 7-14 micromol/kg i.v.) reduced both distension response types but reduced only inhibitory responses to CCK. CGP-35348 (100 nmol i.c.v. or 100 micromol/kg i.v.) reversed baclofen's effect on distension responses, but inhibitory responses to CCK remained attenuated. They were, however, reversed by CGP-62349 (0.4 nmol i.c.v.). In conclusion, GABA(B) receptors inhibit mechanosensitivity, not chemosensitivity, of vagal afferents peripherally. Mechanosensory input to brain stem neurons is also reduced centrally by GABA(B) receptors, but excitatory chemosensory input is unaffected. Inhibitory mechano- and chemosensory inputs to brain stem neurons (via inhibitory interneurons) are both reduced, but the pathway taken by chemosensory input involves GABA(B) receptors that are insensitive to CGP-35348. PMID- 11254493 TI - Activation of NF-kappaB by hepatitis B virus X protein through an IkappaB kinase independent mechanism. AB - pX, the hepatitis B virus-encoded transcription coactivator, is involved in viral infection in vivo. pX stimulates the activity of several transcription factors including nuclear factor-kappaB (NF-kappaB), but the mechanism of activation is poorly understood. The IkappaB kinase complex (IKK) mediates activation of NF kappaB in response to various extracellular stimuli, including inflammatory cytokines like tumor necrosis factor and interleukin 1, human T cell lymphoma virus 1 Tax protein, and tumor promoters like phorbol esters. It is not known whether IKK also mediates activation of NF-kappaB by pX. Here we report that IKK was not essential for activation of NF-kappaB by pX. Expression of pX resulted in the degradation of IkappaBalpha in the absence of its phosphorylation at Ser(32) and Ser(36) residues. Although pX stimulated the activity of cotransfected IKK beta when it was overexpressed, it failed to activate endogenous IKK. Furthermore, expression of pX stimulated NF-kappaB nuclear translocation and transcriptional activity in IKK-gamma-null fibroblast 5R cells. Our data indicate that pX stimulates NF-kappaB activity through a mechanism that is dependent on IkappaBalpha degradation but not on IKK activation. PMID- 11254494 TI - Determinants of terminal mesenteric artery resistance during the first postnatal month. AB - Experiments were conducted to delineate the vascular effector systems that contribute to setting mesenteric vascular tone in swine during the first postnatal month. Terminal mesenteric arteries (TMA), which function as resistance vessels, were studied in vitro with a microvascular perfusion system allowing independent pressure and flow manipulation. When pressure was varied 0-100 mmHg in the absence of flow, TMA from 1-day-old animals demonstrated myogenic vasoconstriction, whereas TMA from 40-day-old animals did not. In 1- but not 40 day-old TMA, the endothelin A (ET(A)) receptor antagonist BQ-610 shifted the pressure-diameter curve upward, whereas the ET(B) receptor antagonist BQ-788 and the L-arginine analog N(G)-monomethyl-L-arginine (L-NMMA) shifted the curve downward; in all instances, myogenic vasoconstriction was preserved. Flow eliminated myogenic vasoconstriction in 1-day-old TMA, i.e., diameter increased as a function of pressure. The effect of BQ-610 was lost under flow conditions; however, BQ-788 and N-acyl-L-Trp-3,5-bis-(trifluoromethyl) benzyl ester, an antagonist specific to the substance P neurokinin-1 (NK(1)) receptor, shifted the pressure-diameter curve downward in the presence of flow, whereas L-NMMA restored myogenic vasoconstriction. Adding flow had no effect on the pressure-diameter relationship in 40-day-old TMA. Other blocking agents, including prazosin, losartan, indomethacin, and charybdotoxin, had no effect on the pressure-diameter relationship in either age group under flow or no-flow conditions. Constitutive production of nitric oxide (NO) and endothelin-1 participates in setting resistance in 1-day-old TMA, and important stimulants to NO production include flow and activation of ET(B) and NK(1) receptors. In contrast, 40-day-old TMA act as passive conduits in which the elastic properties of the vessel are the primary determinant of diameter. PMID- 11254496 TI - Targeted disruption of the Nhe1 gene fails to inhibit beta(1)-adrenergic receptor induced parotid gland hypertrophy. AB - Chronic beta(1)-adrenergic receptor activation results in hypertrophy and hyperplasia of rodent salivary gland acinar cells. Na(+)/H(+) exchanger isoform 1 (NHE1) regulates cell volume and the induction of cell proliferation in many tissues. To investigate the relationship between NHE1 and the response of parotid glands to beta(1)-adrenergic agonists, we examined by Northern blot analysis NHE1 expression in saline-treated mice and mice 30 min and 2, 6, and 24 h after isoproterenol injection. NHE1 transcripts increased approximately 50% by 2 h, and a more than twofold increase was noted at 24 h. Isoproterenol did not acutely increase Na(+)/H(+) exchanger activity; however, exchanger activity was significantly elevated by 24 h. To test whether NHE1 activity is essential for inducing salivary gland hypertrophy in vivo, mice with targeted disruption of Nhe1 were treated with isoproterenol. Na(+)/H(+) exchanger activity was absent in acinar cells from Nhe1(-/-) mice, nevertheless, the lack of NHE1 failed to inhibit isoproterenol-induced hypertrophy. These data directly demonstrate that acinar cell hypertrophy induced by chronic beta(1)-adrenergic receptor stimulation occurs independently of NHE1 activity. PMID- 11254495 TI - SCFA increase intestinal Na absorption by induction of NHE3 in rat colon and human intestinal C2/bbe cells. AB - Short-chain fatty acids (SCFA), produced by colonic bacterial flora fermentation of dietary carbohydrates, promote colonic Na absorption through mechanisms not well understood. We hypothesized that SCFA promote increased expression of apical membrane Na/H exchange (NHE), serving as luminal physiological cues for regulating colonic Na absorptive capacity. Studies were performed in human colonic C2/bbe (C2) monolayers and in vivo. In C2 cells exposed to butyrate, acetate, proprionate, or the poorly metabolized SCFA isobutyrate, apical membrane NHE3 activity and protein expression increased in a time- and concentration dependent manner, whereas no changes were observed for NHE2. In contrast, no significant changes in brush-border hydrolase or villin expression were noted. Analogous to the in vitro findings, rats fed the soluble fiber pectin exhibited a time-dependent increase in colonic NHE3, but not NHE2, protein, mRNA, and brush border activity. These changes were region-specific, as no changes were observed in the ileum. We conclude that luminal SCFA are important physiological cues for regulating colonic Na absorptive function, allowing the colon to adapt to chronic changes in dietary carbohydrate and Na loads. PMID- 11254497 TI - Expression and immunolocalization of aquaporin water channels in rat exocrine pancreas. AB - Both the acinar and ductal cells of the pancreas secrete a near-isotonic fluid and may thus be sites of aquaporin (AQP) water channel expression. Northern blot analysis of mRNA from whole rat pancreas revealed high levels of AQP1 and AQP8 expression, whereas lower levels of AQP4 and AQP5 expression were just detectable by RT-PCR Southern blot analysis. Immunohistochemistry showed that AQP1 is localized in the microvasculature, whereas AQP8 is confined to the apical pole of the acinar cells. No labeling of acinar, ductal, or vascular tissue was detected with antibodies to AQP2-7. With immunoelectron microscopy, AQP8 labeling was observed not only at the apical membrane of the acinar cells but also among small intracellular vesicles in the subapical cytoplasm, suggesting that there may be regulated trafficking of AQP8 to the apical plasma membrane. To evaluate the contribution of AQPs to the membrane water permeability, video microscopy was used to measure the swelling of acinar cells in response to hypotonic stress. Osmotic water permeability was reduced by 90% following exposure to Hg(2+). Since AQP8 is confined to the apical membrane, the marked effect of Hg(2+) suggests that other water channels may be expressed in the basolateral membrane. PMID- 11254498 TI - Regulated MIP-3alpha/CCL20 production by human intestinal epithelium: mechanism for modulating mucosal immunity. AB - Human intestinal epithelial cells secrete an array of chemokines known to signal the trafficking of neutrophils and monocytes important in innate mucosal immunity. We hypothesized that intestinal epithelium may also have the capacity to play a role in signaling host adaptive immunity. The CC chemokine macrophage inflammatory protein (MIP)-3alpha/CCL20 is chemotactic for immature dendritic cells and CD45RO(+) T cells that are important components of the host adaptive immune system. In these studies, we demonstrate the widespread production and regulated expression of MIP-3alpha by human intestinal epithelium. Several intestinal epithelial cell lines were shown to constitutively express MIP-3alpha mRNA. Moreover, MIP-3alpha mRNA expression and protein production were upregulated by stimulation of intestinal epithelial cells with the proinflammatory cytokines tumor necrosis factor-alpha or interleukin-1alpha or in response to infection with the enteric bacterial pathogens Salmonella or enteroinvasive Escherichia coli. In addition, MIP-3alpha was shown to function as a nuclear factor-kappaB target gene. In vitro findings were paralleled in vivo by increased expression of MIP-3alpha in the epithelium of cytokine-stimulated or bacteria-infected human intestinal xenografts and in the epithelium of inflamed human colon. Mucosal T cells, other mucosal mononuclear cells, and intestinal epithelial cells expressed CCR6, the cognate receptor for MIP-3alpha. The constitutive and regulated expression of MIP-3alpha by human intestinal epithelium is consistent with a role for epithelial cell-produced MIP-3alpha in modulating mucosal adaptive immune responses. PMID- 11254499 TI - Direct cell-to-cell contact between Kupffer cells and hepatocytes augments endotoxin-induced hepatic injury. AB - This study focuses on the importance of direct contact between Kupffer cells (KCs) and hepatocytes (HCs) during the hepatic inflammatory response using an in vitro approach. The lipopolysaccharide (LPS)-induced inflammatory response in monocultures of porcine HCs and KCs were compared with cocultures prepared either with direct contact between KCs and HCs (DC cocultures) or without direct contact using cell culture membrane inserts. Our data show that DC cocultures exhibited the highest production of tumor necrosis factor (TNF)-alpha, interleukin-6, and nitric oxide (NO) compared with the other cultures. Immunohistochemical studies revealed that TNF-alpha was exclusively produced by KCs, whereas HCs were responsible for NO production after LPS stimulation. Biotransformation capacity, as determined by cytochrome P-450 and UDP glucuronosyl transferase enzyme activities, was most significantly decreased in DC cocultures. These results provide evidence that direct contact between KCs and HCs favors the extensive TNF alpha production by KCs but in turn affects HC functionality and viability. These findings suggest that direct contact between KCs and HCs plays a key role in the development of a fulminating hepatic inflammatory response. PMID- 11254500 TI - Ca(2+) signaling in porcine duodenal glands by muscarinic receptor activation. AB - The duodenal glands have been thought to play an important role in defense against proximal duodenal ulcer; however, the secretory mechanisms of these glands remain to be determined. In isolated duodenal acinar cells of the pig, we investigated the effects of ACh on intracellular Ca(2+) concentration ([Ca(2+)](i)) and on membrane currents with fura 2 fluorometry and the patch clamp technique. ACh caused a transient increase in [Ca(2+)](i), and the increase was markedly inhibited by atropine or 4-diphenylacetoxy-N-methylpiperidine methiodide but not by hexamethonium, pirenzepine, or methoctramine. The expression of mRNA for the M(3) subtype far exceeded that for either M(1) or M(2) as revealed by real-time quantitative PCR and in situ hybridization. The rise in [Ca(2+)](i) evoked by ACh was largely inhibited by thapsigargin but slightly affected by extracellular Ca(2+) deprivation. Caffeine had no effect on [Ca(2+)](i). ACh elicited Ca(2+)-dependent K(+) currents, a finding similar to the response to inositol 1,4,5,-trisphosphate applied intracellularly. These results suggest the presence of M(3) receptors linked to Ca(2+) release in porcine duodenal glands. PMID- 11254501 TI - Interleukin-5 inhibition of biliary cell chloride currents and bile flow. AB - Recent studies have detected significant elevations of interleukin (IL)-5 mRNA in the liver parenchyma of patients with both primary biliary cirrhosis and acute rejection after liver transplantation. In both of these disorders, intrahepatic biliary epithelial cells (BECs) are the targets of injury. We hypothesized that BECs may themselves express IL-5 receptors that may modulate key biliary functions. RNAs coding for IL-5alpha and -beta receptors were amplified by RT/PCR from a biliary cell line derived from a human cholangiocarcinoma (Mz-ChA-1) and verified by DNA sequencing. IL-5 receptor distribution was detected immunocytochemically on Mz-ChA-1 cells, immortalized murine BEC, bile duct ligated rat liver, and isolated cholangiocytes. Patch-clamp studies on Mz-ChA-1 cells showed that IL-5 inhibits 5'-N-ethylcarboxamidoadenosine-stimulated chloride currents. Additional functional studies showed that IL-5 inhibits secretin-induced bile flow. We conclude that BECs express IL-5 receptors and that IL-5 modulates BEC chloride currents and fluid secretion. Since IL-5 has previously been associated with cholestatic liver disease, we speculate that IL-5 may contribute to liver injury through its effects on biliary secretion. PMID- 11254502 TI - Neutrophil transepithelial migration: regulation at the apical epithelial surface by Fc-mediated events. AB - Neutrophil (PMN) transepithelial migration is a major effector of epithelial defense in inflammatory diseases involving mucosal surfaces. However, major receptor-ligand interactions between epithelial cells and PMN remain incompletely characterized. To better define the molecular events involved in PMN interactions with epithelial cells, we produced a monoclonal antibody called g82 that inhibited PMN transepithelial migration in the physiological basolateral-to apical direction. The g82 antigen localized to the apical surface of human colonic epithelium and was significantly upregulated under inflammatory conditions. Immunoprecipitation revealed two polypeptides of M(r) 207 and 32 kDa. F(ab')(2) fragments from g82 IgG had no effect on transmigration, suggesting Fc dependence. Further experiments confirmed dependence on the PMN Fc receptor CD32A and that the observed effects were secondary to a failure of PMN to detach from the apical epithelial surface. These Fc-mediated events were epitope specific since binding, isotype-matched antibodies did not affect detachment. These results identify a new mechanism for retention of PMN at the apical epithelial surface following transepithelial migration. This pathway may be important in pathogen clearance and mucosal pathophysiology associated with autoimmunity. PMID- 11254503 TI - Colonic production and expression of IL-4, IL-6, and IL-10 in neonatal suckling rats after LPS challenge. AB - It has been demonstrated that the neonatal suckling rat is more susceptible to endotoxin [lipopolysaccharide (LPS)]-induced colonic damage compared with weaned littermates. There is evidence to suggest that differences in the production of certain cytokines, including interleukin (IL)-4, IL-6, and IL-10, are associated with intestinal inflammation in children. We have examined the production, localization, and mRNA detection of these cytokines in suckling and weaned rat colons after bacterial LPS challenge. Suckling (10 day old) and weaned (25 day old) rats were injected with LPS (3 mg/kg ip). Colon samples were taken up to 4 h after treatment, and cytokines were measured by ELISA. LPS-induced cytokine levels were significantly different in suckling rats compared with weaned rats. Cytokine localization to the colonic mucosa was evident in suckling rats up to 4 h after LPS administration but was not consistently seen in weaned rats. The mRNA for cytokines examined were detected by RT-PCR in suckling but not in weaned rat colons after LPS treatment. Induction of neutropenia via anti-neutrophil serum (ANS) administration did not affect cytokine mRNA detection in neonates after LPS treatment. Weaned animals displayed positive detection of all cytokines examined after ANS. Therefore, we have shown that the suckling rat displays a different production and expression of colonic IL-4, IL-6, and IL-10 compared with weaned littermates after LPS challenge. Furthermore, neutrophils may be implicated in colonic cytokine expression after LPS challenge in rats. PMID- 11254504 TI - The human Na(+)/H(+) exchanger NHE2 gene: genomic organization and promoter characterization. AB - The Na(+)/H(+) exchanger (NHE) 2 belongs to a family of plasma membrane transporters involved in intracellular pH and cell volume regulation. We recently reported cloning of human NHE2 (hNHE2) from a colonic cDNA library. Northern blot analysis has identified NHE2 mRNA only in small intestine, prostate, kidney, colon, and skeletal muscle. In this study, we describe the structure and 5' regulatory region of the hNHE2 gene. The hNHE2 gene spans >90 kb and is organized in 12 exons intervened by 11 introns. All introns contain the conserved GT and AG dinucleotides at the donor and acceptor sites, respectively. The hNHE2 gene was mapped to chromosome 2q11.2. Primer extension analysis revealed a single transcription initiation site in human colonic adenocarcinoma cell lines. Analysis of the DNA nucleotide sequences of a 1.4-kb fragment of the 5'-flanking region shows no canonical TATA or CAAT boxes. However, the promoter region contains several potential cis-regulatory elements such as Sp1, early growth response-1, activator protein-2, MyoD, p300, nuclear factor-kappaB, myeloid zinc finger protein-1, caudal-related homeobox (Cdx) gene A, and Cdx protein-2 binding sites. In transient transfection studies, a reporter construct containing the 1.4 kb promoter region exhibited low luciferase activity levels. However, after deletion upstream of -664, its activity increased approximately threefold. Thus our data suggest that an inhibitory element may exist in the NHE2 promoter 5' upstream region. PMID- 11254505 TI - Epidermal growth factor increases surface hydrophobicity and resistance to acid in the rat duodenum. AB - Epidermal growth factor (EGF) is produced in Brunner's glands and plays a role in healing and repair of duodenal ulcers. We examined the participation of zwitterionic phospholipids of mucus in the effects of EGF. Under anesthesia, groups of rats received an intraduodenal bolus of either saline or EGF. Some rats received subcutaneous indomethacin followed by EGF or EGF followed by a detergent (5% Brij 35, a nonionic detergent that solubilizes luminal phospholipids). Thirty minutes after treatment, mucosal surface hydrophobicity and phospholipid concentration in the mucus layer were measured. Matched groups of rats were challenged with 0.5 M HCl, instilled intraduodenally 30 min after treatment, and mucosal damage was assessed 1 h after acid challenge. Exogenous EGF significantly increased surface hydrophobicity and phosphatidylcholine concentration in the mucus layer. EGF treatment also reduced mucosal damage induced by acid. However, indomethacin pretreatment or detergent administration after EGF abolished both protection against acid and changes in the mucus layer. These data suggest that EGF increases duodenal resistance to luminal acid via stimulation of mucosal zwitterionic phospholipids. PMID- 11254506 TI - Menopause, hormone replacement therapy, and sleep-disordered breathing: are we ready for the heat? PMID- 11254507 TI - Detailed occupational history: the cornerstone in diagnosis of asbestos-related lung disease. PMID- 11254508 TI - Basic science in ventilator-induced lung injury: implications for the bedside. PMID- 11254509 TI - Reinfection tuberculosis: how important is it? PMID- 11254510 TI - In the cards was ARDS: how we discovered the acute respiratory distress syndrome. PMID- 11254511 TI - Trauma critical care. AB - The surgical approach to the most injured patients has changed in recent years. Many patients arrive in the intensive care unit with problems that in the past would have been definitively addressed in the operating room, or led to the patient's demise due to continued attempts to complete all surgical procedures, despite deteriorating physiology. As a result, the triad of hypothermia, acidosis, and coagulopathy, along with the frequent complication of abdominal compartment syndrome, are critical factors that require correction in the intensive care unit. Prompt correction is necessary not only to allow expeditious completion of required surgical procedures, but because this triad, unless interrupted, invariably leads to death during resuscitation. PMID- 11254512 TI - Prevalence of sleep-disordered breathing in women: effects of gender. AB - The prevalence of sleep-disordered breathing has not been well studied in women, especially in terms of the effects of age, body mass index (BMI), and menopause. We evaluated this question using a two-phase random sample from the general population. In Phase I, 12,219 women and 4,364 men ranging in age from 20 to 100 yr were interviewed; and in Phase II, 1,000 women and 741 men of the Phase I subjects were selected for one night of sleep laboratory evaluation. The results of our study indicated that, for clinically defined sleep apnea (apnea/hypopnea index > or = 10 and daytime symptoms), men had a prevalence of 3.9% and women 1.2%, resulting in an overall ratio of sleep apnea for men to women of 3.3:1 (p = 0.0006). The prevalence of sleep apnea was quite low in premenopausal women (0.6%) as well as postmenopausal women with hormone replacement therapy (HRT) (0.5%). Further, in these women the presence of sleep apnea appeared to be associated exclusively with obesity (BMI > or = 32.3 kg/m2). Postmenopausal women without HRT had a prevalence of sleep apnea that was significantly higher than the prevalence in premenopausal women with HRT (2.7 versus 0.6%, p = 0.02) and was more similar to the prevalence in men (3.9%), although it remained significantly less when controlling for age and BMI (p = 0.001). These data combined indicate that menopause is a significant risk factor for sleep apnea in women and that hormone replacement appears to be associated with reduced risk. PMID- 11254513 TI - Clearance of filtered fluid from the lung during exercise: role of hyperpnea. AB - During strenuous exercise in sheep, lung lymph flow increases within seconds and rises to levels 7- to 10-fold over baseline. Concomitant with the flow increase, the lymph protein content rapidly decreases to levels consistent with severe capillary hypertension. This pattern of clearance of filtered fluid is quite different than is seen with the passive capillary hypertension that results from mechanical obstruction of the mitral valve. In passive capillary hypertension, the increase in lymph flow and reduction in lymph protein content develop over several hours. The purpose of this study was to discover if these observed differences in edema clearance are related to the hyperpnea that accompanies exercise. Sheep were instrumented for continuous measurement of pulmonary arterial, left atrial, and systemic pressures, cardiac output by ultrasound, lung lymph flow, and ventilation. First, hemodynamics, ventilatory, and lymph clearance variables were measured during moderate exercise at 2.8 mph on a treadmill. Second, on a separate occasion, sheep were induced to hyperventilate to the same minute ventilation as during exercise, using modest CO2 stimulation. Lymph flow and hemodynamics were unaffected by this hyperpnea. The third arm of the experiment was to raise pulmonary microvascular pressure at rest to the level seen with exercise by means of a balloon catheter placed in the mitral valve. Lymph flow rose and protein content decreased slowly and to a lower degree than seen with exercise despite a comparable microvascular pressure. Finally, left atrial hypertension and induced hyperpnea were combined in sheep at rest, and the resulting lymph flow and protein content were the same as seen with exercise at similar pressures and ventilation. We conclude that hyperpnea is a major mechanism of interstitial liquid clearance during exercise, and may be largely responsible for preventing pulmonary edema that might occur at the high microvascular pressures of strenuous exercise. PMID- 11254514 TI - Spirometric pulmonary function in healthy preschool children. AB - The purpose of this study was to evaluate spirometric lung function in normal children ages 3 to 6 yr. Spirometric measurements were obtained at nursery and daycare centers by experienced pediatric pulmonary function technicians. Of 307 children recruited, 259 fulfilled our criteria as normal. Of these, 82.6% (214) were able to perform technically acceptable and reproducible maneuvers during a testing session limited to 15 min. The regression model with log-transformed parameters of pulmonary function and height had the best correlations. After accounting for height in the model, other physical traits and health questionnaire items did not contribute significantly. PEFR, FVC, FEV1, and FEF25 75 all increased with increasing height; correlation coefficients were 0.73, 0.93, 0.92, and 0.67, respectively. The group mean coefficients of variation for replicate measurements of PEFR, FVC, FEV1, and FEF25-75 were 7.8%, 2.5%, 2.7%, and 8.3%, respectively. There was a significant decrease in the ratio FEV1/FVC with increasing height; the mean predicted FEV1/FVC was 0.97 at 90 cm height and 0.89 at 125 cm height. In conclusion, reproducible spirometry can be obtained in the majority of preschool children and has the potential to improve our assessment and management of pulmonary disease. PMID- 11254515 TI - The impact of insulin-dependent diabetes on ventilatory control in the mouse. AB - Insulin-dependent diabetes mellitus (IDDM) can lead to ventilatory depression and decreased sensitivity to hypercapnia. We examined relationships between ventilation, plasma insulin, leptin, ketones, and blood glucose levels in two mouse models of IDDM: (1) streptozotocin-induced diabetes in C57BL/6J mice on a regular diet or with induced obesity from a high fat diet; and (2) spontaneous diabetes mellitus in NOD-Ltj mice. In both mouse models, IDDM resulted in depression of the hypercapnic ventilatory response (HCVR). This ventilatory depression was not associated with decreases in plasma insulin or leptin levels. There was, however, a strong association between the duration of hyperglycemia, the decline in HCVR, and increased glycosylation of the diaphragm. Hyperventilation was observed in only six of 14 C57BL/6J obese wild-type mice, despite a significant degree of diabetic ketoacidosis (DKA) in all 14 animals. In mice with DKA, there was a significant correlation between the increase in baseline minute ventilation (V E) and hyperleptinemia (r = 0.77, p < 0.01). In leptin-deficient C57BL/6J-Lep(ob) mice, low levels of both V E and ketones were observed. These results suggest that: (1) depression of the HCVR in IDDM is associated with hyperglycemia and glycosylation of the diaphragm; and (2) the hyperventilation of DKA is leptin dependent. PMID- 11254516 TI - Rapid declines in FEV1 and subsequent respiratory symptoms, illnesses, and mortality in coal miners in the United States. AB - Coal mine dust exposure is associated with accelerated loss of lung function. We assessed long-term health outcomes in two groups of underground coal miners who during previous mine surveys had shown either high rates of FEV1 decline (cases, n = 310) or relatively stable lung function (referents, n = 324). Cases and referents were matched initially for age, height, smoking status, and FEV1. We determined vital status for 561 miners, and obtained a follow-up questionnaire for 121 cases and 143 referents. Responses on the follow-up questionnaire were compared with those on the last previous mine health survey questionnaire. Cases showed a greater incidence of symptoms than did referents for cough, phlegm production, Grades II and III dyspnea, and wheezing, and greater incidences than referents of chronic bronchitis and self- reported asthma and emphysema. More cases than referents (15% versus 4%) left mining before retirement because of chest illnesses. After controls were applied for age and smoking, cases had twice the risk of dying of cardiovascular and nonmalignant respiratory diseases and a 3.2-fold greater risk of dying of chronic obstructive pulmonary disease than did referents. Rapid declines in FEV1 experienced by some coal miners are associated with subsequent increases in respiratory symptoms, illnesses, and mortality from cardiovascular and nonmalignant respiratory diseases. PMID- 11254517 TI - Transforming growth factor-beta2 induces pleurodesis significantly faster than talc. AB - Transforming growth factor-beta2 (TGF-beta2) has recently been shown to produce effective pleurodesis in rabbits. Conventional pleurodesing agents such as talc act by inducing pleural injury, which results in acute inflammation and fibrosis. TGF-beta2 is a profibrotic cytokine capable of producing fibrosis without inducing significant pleural inflammation. We hypothesize that intrapleural administration of TGF-beta2 would (1) produce an effective pleurodesis faster; (2) stimulate more collagen deposition, and (3) induce less inflammation when compared with intrapleural injection of talc. Thirty rabbits were divided into two groups and given either TGF-beta(2) (1.7 microg) or talc slurry (400 mg/kg) via a chest tube. Five rabbits from each group were killed at Days 1, 4, and 7. Gross pleurodesis was graded from 1 (none) to 8 (complete symphysis). The microscopic pleural inflammation and fibrosis were graded from 0 to 4. Pleural thickening and the total area of collagen deposition were compared. Intrapleural injection of TGF-beta2 produced effective pleurodesis within 7 d (median pleurodesis score = 7 at Day 7). At Day 7, TGF-beta2 induced significantly more collagen deposition (19.4 +/- 19.6% versus 4.6 +/- 2.9% of total area of pleura at Day 7), higher pleural fibrosis score (3.0 +/- 1.0 versus 1.8 +/- 0.5), and pleural thickness (286 +/- 191 versus 85 +/- 37 microm) than did talc. There was no difference in the degree of pleural inflammation between the two groups at Day 7 (2.6 +/- 0.9 for TGF-beta2 versus 2.4 +/- 0.6 for talc) or at any other time points. In conclusion, the intrapleural administration of TGF-beta2 produced excellent pleurodesis in rabbits at a rate faster than talc slurry and all other pleurodesing agents investigated before. TGF-beta2 stimulated more collagen deposition without inducing excess inflammation when compared with talc slurry. TGF-beta2 may have advantages over talc slurry in the management of recurrent pleural effusion and pneumothorax. PMID- 11254518 TI - Etiology of severe pneumonia in the very elderly. AB - The etiology of severe pneumonia requiring mechanical ventilation in the very elderly has been imprecise because of lack of comprehensive studies and low yield of diagnostic approach. Overall, 104 patients 75 yr of age and older with severe pneumonia were studied prospectively at two university-affiliated hospitals. Microbial investigation included blood culture, serology, pleural fluid, and bronchoalveolar secretions. Streptococcus pneumoniae (14%), gram-negative enteric bacilli (14%), Legionella sp. (9%), Hemophilus influenzae (7%), and Staphylococcus aureus (7%) were the predominant pathogens in community-acquired pneumonia (CAP). Staphylococcus aureus (29%), gram-negative enteric bacilli (15%), Streptococcus pneumoniae (9%), and Pseudomonas aeruginosa (4%) accounted for most isolates of nursing home-acquired pneumonia (NHAP). The case fatality rate was 55% (53% for CAP and 57% for NHAP; p > 0.5). Activity of Daily Living (ADL) Index, pulmonary, endocrine and central nervous system (CNS) comorbidities were associated with distinct microbial etiology. By multivariate analysis, hospital mortality was associated independently with 24-h urine output (odds ratio [OR], 5.6; 95% confidence interval [CI], 2.5 to 7.9; p < 0.001), septic shock (OR, 4.3; 95% CI, 1.9 to 8.9; p = 0.0059), radiographic multilobar involvement (OR, 3.7; 95% CI, 1.8 to 15.6; p = 0.02), and inadequate antimicrobial therapy (OR, 2.6; 95% CI, 1.4 to 23.9; p = 0.034). Further studies should focus on identifying effective antimicrobial regimens in randomized trials. PMID- 11254519 TI - Autoadjusting CPAP therapy based on impedance efficacy, compliance and acceptance. AB - Constant continuous positive airway pressure (CPAP) is the treatment of choice for the obstructive sleep apnea syndrome (OSAS). To enable the pressure to be matched more accurately to actual requirements, and thus increase patient acceptance, an autoadjusting device based on the measurement of upper airway impedance was developed (APAP(FOT)). We investigated the efficacy and compliance in continuous use at home. Fifty-two patients were treated (randomized crossover) with CPAP and APAP(FOT) for 6 wk each. Respiratory disturbances, sleep profile, and arousals improved significantly with both modes (AHI: baseline, 35.1 +/- 26/h; APAP(FOT), 5.0 +/- 5.2; CPAP, 4.3 +/- 6.3; p < 0.001 baseline versus each mode). The mean pressure with APAP(FOT) was significantly reduced as compared with CPAP (CPAP, 7.8 +/- 1.5 cm H2O; APAP(FOT), 5.7 +/- 1.8 cm H2O; p < 0.001). Under APAP(FOT) the pressure was lower than that under CPAP for 81.5 +/- 21% of the time. Although overall use did not differ, 75% of the patients preferred APAP(FOT) for home treatment. We conclude that APAP(FOT) is as efficacious as constant CPAP in the treatment of OSAS. The treatment pressure can be reduced significantly, and sleep microstructure improved with APAP(FOT). These might be the reasons for patient preference of automatic therapy. PMID- 11254520 TI - Predictors of successful extubation in neurosurgical patients. AB - A respiratory therapist-driven weaning protocol incorporating daily screens, spontaneous breathing trials (SBT), and prompts to caregivers has been associated with superior outcomes in mechanically ventilated medical patients. To determine the effectiveness of this approach in neurosurgical (NSY) patients, we conducted a randomized controlled trial involving 100 patients over a 14-mo period. All had daily screens of weaning parameters. If these were passed, a 2-h SBT was performed in the Intervention group. Study physicians communicated positive SBT results, and the decision to extubate was made by the primary NSY team. Patients in the Intervention (n = 49) and Control (n = 51) groups had similar demographic characteristics, illness severity, and neurologic injuries. Among all patients, 87 (45 in the Control and 42 in the Intervention group) passed at least one daily screen. Forty (82%) patients in the Intervention group passed SBT, but a median of 2 d passed before attempted extubation, primarily because of concerns about the patient's sensorium (84%). Of 167 successful SBT, 126 (75%) did not lead to attempted extubation on the same day. The median time of mechanical ventilation was 6 d in both study groups, and there were no differences in outcomes. Overall complications included death (36%), reintubation (16%), and pneumonia (9%). Tracheostomies were created in 29% of patients. Multivariate analysis showed that Glasgow Coma Scale (GCS) score (p < 0.0001) and partial pressure of arterial oxygen/fraction of inspired oxygen ratio (p < 0.0001) were associated with extubation success. The odds of successful extubation increased by 39% with each GCS score increment. A GCS score > or = 8 at extubation was associated with success in 75% of cases, versus 33% for a GCS score < 8 (p < 0.0001). Implementation of a weaning protocol based on traditional respiratory physiologic parameters had practical limitations in NSY patients, owing to concerns about neurologic impairment. Whether protocols combining respiratory parameters with neurologic measures lead to superior outcomes in this population requires further investigation. PMID- 11254521 TI - Antibody to intercellular adhesion molecule 1 (CD54) decreases survival and not lung injury in baboons with sepsis. AB - Neutrophil influx into the lung is an important event in the pathogenesis of acute lung injury in gram-negative sepsis. We hypothesized that administration of a monoclonal antibody to intercellular adhesion molecule 1 (ICAM-1, CD54), a molecule mediating neutrophil adhesion to endothelial cells, would decrease neutrophil sequestration and transmigration in the lung and attenuate lung injury in Escherichia coli sepsis. Sepsis was induced in 12 baboons primed with heat killed E. coli (1 x 10(9) CFU/kg) 12 h before infusion of live bacteria (1 x 10(10) CFU/kg). Six animals received monoclonal antibody to CD54 (1 mg/kg) intravenously at the time of live E. coli infusion. After 48 h or when blood pressure could not be maintained, tissues were harvested and bronchoalveolar lavage (BAL) samples were obtained. Median survival time was decreased in anti CD54-treated animals. This group also had decreased mean arterial pressure, increased metabolic acidosis, and decreased urine output. Measures of lung injury including gas exchange, lung lavage protein and lactate dehydrogenase (LDH), lung thiobarbituric acid-reactive species, and lung histology, including alveolar neutrophil volumes, were unaffected by treatment. The effect of anti-CD54 on neutrophil influx into tissues as measured by myeloperoxidase was organ specific. These data show that monoclonal antibody to CD54 does not ameliorate acute lung injury in E. coli sepsis, and septic primates given anti-CD54 have worsened metabolic parameters and decreased survival. PMID- 11254522 TI - Fluticasone inhibits but does not reverse allergen-induced structural airway changes. AB - Ethical and technical reasons limit the possibility of evaluating the effects of inhaled corticosteroids on structural changes in airways of humans with asthma. We therefore evaluated whether fluticasone propionate (FP) modifies airway remodeling, induced by repeated allergen exposure in rats. Sensitized BN rats were exposed to aerosolized ovalbumin (OA) for 2 wk. To assess the effect of FP on the development of or on established airway remodeling, animals were treated with aerosolized FP or placebo during allergen exposure or for 2 wk afterward. Compared with animals exposed to phosphate-buffered saline (PBS), OA-challenged animals developed an increase in total airway wall area, enhanced fibronectin deposition, epithelial cell proliferation, goblet cell hyperplasia, and airway hyperresponsiveness. Concomitant treatment with FP decreased all allergen-induced structural changes without being able to reverse them to normal. Initiating FP treatment after the allergen exposure had no effect on any of the OA-induced structural airway changes. The increase in total airway wall area, enhanced fibronectin deposition, and epithelial cell proliferation persisted. The goblet cell hyperplasia disappeared spontaneously. In conclusion, concomitant treatment with FP partly inhibits structural airway changes as well as hyperresponsiveness induced by OA exposure. Post hoc treatment fails to reverse established airway remodeling. PMID- 11254523 TI - Antipneumolysin antibody titers in HIV-seropositive injection drug users before and after pneumococcal bacteremia. AB - Lower baseline antipneumolysin antibody (alpha-PLY) levels have been found in populations with a higher incidence of pneumococcal infections. To determine whether predisease alpha-PLY titer is associated with invasive pneumococcal disease in HIV-seropositive injection drug users (IDU), we utilized a prospective cohort of IDU in Baltimore to compare alpha-PLY titers before bacteremia in 28 HIV- seropositive IDU cases with alpha-PLY titers in 56 matched (CD4 and seroconversion date) HIV-seropositive IDU control subjects and 28 matched (calendar time) HIV-seronegative IDU control subjects remaining free of pneumococcal disease. We also compared the postinfection fold-rise of alpha-PLY titers in cases relative to the change in alpha-PLY titers in control subjects during the same interval; alpha-PLY titers were measured using quantitative ELISA, and functional activity was assessed using antihemolysin assays. Predisease alpha-PLY titer did not differ between cases (66 units) and HIV seropositive control subjects (70 units, p = 0.56) or HIV-seronegative control subjects (80 units, p = 0.10). There was a significant difference in fold-rise of alpha-PLY titers postdisease between cases (1.18) and HIV-seronegative control subjects (0.76), p = 0.03. Baseline alpha-PLY titers do not differ significantly between HIV-seropositive IDU who develop pneumococcal bacteremia from HIV seropositive and HIV-seronegative IDU control subjects remaining free of severe pneumococcal disease. PMID- 11254524 TI - Obstructive sleep apnea-hypopnea and related clinical features in a population based sample of subjects aged 30 to 70 yr. AB - The prevalence and related clinical features of obstructive sleep apnea-hypopnea (OSAH) in the general population were estimated in a two-phase cross-sectional study. The first phase, completed by 2,148 subjects (76.9%), included a home survey, blood pressure, and a portable respiratory recording, whereas in the second, subjects with suspected OSAH (n = 442) and a subgroup of those with normal results (n = 305) were invited to undergo polysomnography (555 accepted). Habitual snoring was found in 35% of the population and breathing pauses in 6%. Both features occurred more frequently in men, showed a trend to increase with age, and were significantly associated with OSAH. Daytime hypersomnolence occurred in 18% of the subjects and was not associated with OSAH. An apnea hypopnea index (AHI) > or = 10 was found in 19% of men and 15% of women. The prevalence of OSAH (AHI > or = 5) increased with age in both sexes, with an odds ratio (OR) of 2.2 for each 10-yr increase. AHI was associated with hypertension after adjusting for age, sex, body mass index, neck circumference, alcohol use, and smoking habit. This study adds evidence for a link between OSAH and hypertension. PMID- 11254525 TI - Peak flow as predictor of overall mortality in asthma and chronic obstructive pulmonary disease. AB - Lung function is a strong predictor of overall mortality in asthma and chronic obstructive pulmonary disease (COPD). FEV1 is considered to be the "gold standard," whereas peak expiratory flow (PEF) is mostly used in absence of FEV1 measurements. We compared the predictive power of PEF and FEV1, measured after maximal bronchodilation, which included a short course of oral corticosteroids. The study population comprised 491 asthmatics and 1,095 subjects with COPD. Pulmonary function tests were performed between 1983 and 1988, and survival data were obtained by September 1997, when 127 asthmatics and 723 subjects with COPD had died. Predictors of survival were examined by Cox proportional hazards analyses. After controlling for age, smoking, sex, and body mass index, we found best PEF to be at least equal to best FEV1 as predictor of overall mortality in subjects with COPD. The predictive power of best PEF was in part maintained after controlling for best FEV1. In asthma, best FEV1 seemed to be a better predictor of mortality than best PEF. Despite close correlation to FEV1, PEF apparently provides independent prognostic information in patients with COPD. This may be due to PEF and FEV1 reflecting different components of COPD, i.e., chronic bronchitis, small airways disease, and emphysema. Furthermore, extrapulmonary components such as muscle mass and general "vigour" probably affect PEF to a greater extent than they affect FEV1. PMID- 11254526 TI - Worsening of asthma in children allergic to cats, after indirect exposure to cat at school. AB - Exposure to cat allergen at school might exacerbate symptoms in asthmatic children with cat allergy. To study this, we identified 410 children, 6-12 yr of age, who were being treated for asthma (inhaled steroids and beta-agonists), were allergic to cats, and had no cat at home. Peak expiratory flow (PEF), asthma symptoms, medication, fever and/or sore throat, and contact with furred pets were recorded twice daily during the last week of summer holidays and the second and third weeks of school. The number of cat owners in each class was recorded. Ninety-two children with asthma reported no contact with furred pets. Among these, children who attended classes with > 18% (median value) cat owners reported significantly decreased PEF, more days with asthma symptoms, and increased use of medication after school started. Those in classes with < or = 18% cat owners reported no change. Children in classes with many cat owners ran a 9-fold increased risk of exacerbated asthma after school start compared with children in classes with few cat owners, after adjusting for age, sex, and fever and/or sore throat. Thus, asthma symptoms, PEF, and the use of asthma medication in children with cat allergy may be affected by indirect cat exposure at school. PMID- 11254527 TI - FeNO measured at fixed exhalation flow rate during controlled tidal breathing in children from the age of 2 yr. AB - We have outlined a new method to measure exhaled nitric oxide on-line at fixed flow rate during controlled tidal breathing (FeNO [controlled]) in young children aged 2 yr and older. FeNO(controlled) measures NO on-line during operator controlled tidal breathing. The operator targets the exhaled flow of the child within preset limits of 0.4-0.6 L/s by continuously adjusting an expiratory resistance. FeNO(controlled) is estimated during end exhalation. We have validated this method against the reference method of the single breath on-line (SBOL) maneuvre (FeNO[SBOL]) and compared it with NO in mixed exhaled air collected in a bag (FeNO [mixed]). Sixty-seven children were studied: 16 school children and 51 children aged 2-5 yr; 14 of the young children were healthy, 22 had asthma treated with regular inhaled budesonide, and 15 had mild episodic wheeze treated with inhaled terbutaline as necessary. FeNO (controlled) showed good agreement with FeNO(SBOL) (factor difference 0.7-1.4), whereas FeNO(mixed) showed poor agreement with FeNO(SBOL) (factor difference 0.51-5.37). FeNO(controlled) (mean [95% confidence interval]) was 6 ppb (4-8 ppb) in young children with asthma, 5 ppb (3-7 ppb) in young children with mild episodic wheeze, and 3 ppb (2-4 ppb) in healthy control subjects (asthma versus control subjects: p = 0.006; episodic wheeze versus control subjects: p = 0.057). FeNO(controlled) increased from 4 ppb (2-7 ppb) to 13 ppb (10-18 ppb) (p < 0.0001) when the mean daily maintenance dose of budesonide was tapered in nine young children with asthma. FeNO(controlled) is feasible in young children from age 2 and shows better agreement with FeNO(SBOL) than FeNO(mixed). FeNO(controlled) covaries with asthma disease severity and steroid dose. FeNO(controlled) is therefore suggested as a noninvasive diagnostic tool for monitoring asthma disease activity in young children with asthma from the age of 2 yr. PMID- 11254528 TI - Surface of localized pleural plaques quantitated by computed tomography scanning: no relation with cumulative asbestos exposure and no effect on lung function. AB - To evaluate if there is a relation between the size of asbestos plaques and the level of past exposure and pulmonary function, we measured the surface of localized pleural plaques found on high-resolution (HR) CT scan, using a computerized video display unit-imaging system, in 73 workers (mean age, 43.5 yr) who had worked from 23 to 27 yr in an asbestos-cement factory. Their estimated cumulative exposure to asbestos ranged from 16.4 to 98.7 fiber-years/ ml (mean, 26.3 fiber-years/ml). Lung function measurements included lung volumes, maximal expiratory flows, and diffusing capacity. A control group of 21 workers was examined by the same procedures. Plaques were detected by CT in 51 (70%) asbestos exposed subjects and in none of the control subjects. The average calculated plaque surface was 47.9 +/- 61.7 cm2 (median, 22.1 cm2; range, 0 to 278.4 cm2). There was no relation between plaque surface and cumulative asbestos exposure (p = 0.24). In the 51 subjects with pleural plaques, the surface of the pleural lesions was not related to cumulative asbestos exposure, or to smoking history or time since first exposure. Neither the presence nor the extent of the plaques was correlated with lung function parameters. PMID- 11254529 TI - Ventilation-induced chemokine and cytokine release is associated with activation of nuclear factor-kappaB and is blocked by steroids. AB - Recent clinical trials have shown that the survival of patients with acute respiratory distress syndrome (ARDS) is improved by ventilation with reduced volumes. These studies suggested that overinflation of the lungs causes overactivation of the immune system. The present study investigated the hypothesis that ventilation with increased tidal volumes results in early responses similar to those caused by stimulation with one of the major risk factors for ARDS: bacterial lipopolysaccharide (LPS). We therefore compared the effects of ventilation (-10 cm H2O or -25 cm H2O end-inspiratory pressure) and LPS (50 microg/ml) on nuclear factor (NF)-kappaB activation, chemokine release, and cytokine release in isolated perfused lungs obtained from BALB/C mice. We found that both LPS and ventilation with -25 cm H2O (overventilation; OV) caused translocation of NF-kappaB, which was abolished by pretreatment with the steroid dexamethasone. Furthermore, both treatments resulted in similar increases in perfusate levels of alpha-chemokines (macrophage inflammatory protein; [MIP]-2; KC), beta-chemokines (macrophage chemotactic protein-1; MIP-1alpha), and cytokines (tumor necrosis factor-alpha, interleukin-6), which were largely prevented by dexamethasone pretreatment. In LPS-resistant C3H/HeJ mice, only OV, and not LPS, caused translocation of NF-kappaB and release of MIP-2. We conclude that OV evokes early inflammatory responses similar to those evoked by LPS (i.e., NF-kappaB translocation and release of proinflammatory mediators). The NF-kappaB translocation elicited by OV appears to be independent of Toll-like receptor 4 and not due to LPS contamination introduced by the ventilator. Our data further suggest that steroids might be considered as a subsidiary treatment during artificial mechanical ventilation. PMID- 11254530 TI - Exogenous reinfection with tuberculosis on a European island with a moderate incidence of disease. AB - The frequency and determinants of exogenous reinfection and of endogenous reactivation of tuberculosis in patients previously treated are poorly understood. In Gran Canaria Island, Spain, between 1991 and 1996, 962 tuberculosis cases were confirmed by culture. Drug susceptibility testing was performed on available bacterial isolates and IS6110-based RFLP genotyping was carried out. Twenty-three patients (2.4%) had two positive cultures separated by at least 12 mo, 18 of whom had bacterial DNA available for genotypic analysis. The initial and final isolates from eight (44%) were different genotypes, indicating exogenous reinfection. Six of them were retreated after cure and two retreated after default. Six were HIV seronegative and two were HIV seropositive. Endogenous reactivation was seen in the remaining 10 patients of whom eight were retreated after default and two after cure. Three of the eight (38%) being retreated after default developed multidrug resistance. One genotype was responsible for a second episode of tuberculosis in five cases, three exogenous reinfections and two endogenous reactivations. In the context of a moderate incidence of tuberculosis, exogenous reinfection is an important cause of TB recurrence, even in HIV-seronegative patients. PMID- 11254531 TI - Temporal association between airway hyperresponsiveness and airway eosinophilia in ovalbumin-sensitized mice. AB - The temporal association between airway inflammation and airway hyperresponsiveness (AHR) has been analyzed in BALB/c mice sensitized to, and subsequently exposed to, a single intranasal challenge of ovalbumin (OVA). In OVA sensitized/challenged animals only a small increase in responsiveness to methacholine (MCh) was seen at 8 h, peaked at 24 to 48 h, and resolved by 96 h. An early bronchoalveolar lavage fluid (BALF) neutrophil infiltrate (peaking at 8 h postchallenge; approximately 72% total cells was observed) that returned to baseline by 48 h. BALF eosinophil numbers did not increase until 48 h (approximately 32% of total cells), peaked at 96 h (approximately 38% total cells), and remained elevated at 8 d (approximately 27% total cells). Airway tissue eosinophilia preceded changes in BALF. Eosinophil peroxidase (EPO) levels in BALF were elevated in OVA-sensitized/challenged mice at 48 h only. BALF TNF alpha levels peaked at 8 h, whereas IL-5 and IL-4 levels peaked at 24 h. IL-13 levels were increased at both 24 and 48 h. Mucus-positive cells were not observed in the airway epithelium until 48 h. Administration of IL-5 or VLA-4 antibody prior to OVA challenge prevented the development of AHR in sensitized mice as well as BALF and tissue eosinophilia. These data identify a temporal association between Th2 cytokine production, tissue eosinophil infiltration and activation, and, importantly, both the development and resolution kinetics of AHR. Moreover, the antibody studies further support the association of eosinophilia with the pathogenesis of AHR. PMID- 11254532 TI - Magnetic resonance imaging of the upper airway in children with Down syndrome. AB - As compared with control subjects, children with Down syndrome have different size and shape relationships among tissues composing the upper airway, which may predispose them to obstructive sleep apnea (OSA). We hypothesized that Down syndrome children without OSA have similar subclinical differences. We used magnetic resonance imaging to study the upper airway in 11 Down syndrome children without OSA (age, 3.2 +/- 1.4 yr) and in 14 control subjects (age, 3.3 +/- 1.1 yr). Sequential T1- and T2-weighted spin-echo axial and sagittal images were obtained. We found a smaller airway volume in subjects with Down syndrome (1.4 +/ 0.4 versus 2.3 +/- 0.8 cm(3) in controls, p < 0.005). Subjects with Down syndrome had a smaller mid- and lower face skeleton. They had a shorter mental spine-clivus distance (5.7 +/- 0.6 versus 6.2 +/- 0.4 cm, p < 0.05), hard palate length (3.2 +/- 0.4 versus 3.7 +/- 0.2 cm, p < 0.005), and mandible volume (11.5 +/- 3.7 versus 16.9 +/- 2.9 cm3, p < 0.0005). Adenoid and tonsil volume was significantly smaller in the subjects with Down syndrome. However, the tongue, soft-palate, pterygoid, and parapharyngeal fat pads were similar to those of control subjects. This study shows that Down syndrome children without OSA do not have increased adenoid or tonsillar volume; reduced upper airway size is caused by soft tissue crowding within a smaller mid- and lower face skeleton. PMID- 11254533 TI - Endothelial cell death and decreased expression of vascular endothelial growth factor and vascular endothelial growth factor receptor 2 in emphysema. AB - Emphysema due to cigarette smoking is characterized by a loss of alveolar structures. We hypothesize that the disappearance of alveoli involves apoptosis of septal endothelial cells and a decreased expression of lung vascular endothelial growth factor (VEGF) and its receptor 2 (VEGF R2). By terminal transferase dUTP nick end labeling (TUNEL) in combination with immunohistochemistry, we found that the number of TUNEL+ septal epithelial and endothelial cells/lung tissue nucleic acid (microg) was increased in the alveolar septa of emphysema lungs (14.2 +/- 2.0/microg, n = 6) when compared with normal lungs (6.8 +/- 1.3/microg, n = 7) (p < 0.01) and with primary pulmonary hypertensive lungs (2.3 +/- 0.8/microg, n = 5) (p < 0.001). The cell death events were not significantly different between healthy nonsmoker (7.4 +/- 1.9/microg) and smoker (5.7 +/- 0.7/microg) control subjects. The TUNEL results were confirmed by single-stranded DNA and active caspase-3 immunohistochemistry, and by DNA ligation assay. Emphysema lungs (n = 12) had increased levels of oligonucleosomal-length DNA fragmentation when compared with normal lungs (n = 11). VEGF, VEGF R2 protein, and mRNA expression were significantly reduced in emphysema. We propose that epithelial and endothelial alveolar septal death due to a decrease of endothelial cell maintenance factors may be part of the pathogenesis of emphysema. PMID- 11254534 TI - Role of nitric oxide in vascular permeability after combined burns and smoke inhalation injury. AB - Patients with severe burn and/or smoke inhalation injury suffer both systemic and pulmonary vascular hyperpermeability. We hypothesized that nitric oxide (NO) produced by inducible nitric oxide synthase (iNOS) plays a role in the changes in microvascular permeability seen with this injury. To test the hypothesis, we administered mercaptoethylguanidine (MEG), a selective iNOS inhibitor, to conscious sheep subjected to a combined smoke inhalation and third-degree burn injury to 40% of total body surface area. The sheep were surgically prepared for chronic study with lung and prefemoral lymph fistulas in order to estimate microvascular permeability. Both the groups and a control group of animals showed an increase in iNOS protein and message in their lungs. The control animals showed significant increases in either plasma or lymph NO2-/NO3- (NOx) concentration at 24 h after injury, with associated cardiac depression and hemoconcentration. The airway epithelium stained for nitrotyrosine. In the treatment group, NOx did not increase significantly in plasma or lymph throughout the experiment, there was no nitrotyrosine staining, hemodynamic depression was not observed, and the fluid requirement was significantly less than in the control group. Changes in pulmonary microvascular permeability were significantly suppressed by inhibition of iNOS. However, there was no significant difference between the two study groups in the microvascular permeability of burned tissue. These data suggest that NO produced by iNOS plays an important role in the changes in systemic and pulmonary microvascular permeability in combined smoke inhalation/third-degree burn injury, but does not affect the vascular permeability of third-degree-burned tissue in this type of injury. PMID- 11254535 TI - Protective role of heme oxygenases against endotoxin-induced diaphragmatic dysfunction in rats. AB - Reactive oxygen species are strongly implicated in diaphragmatic dysfunction during sepsis. We investigated whether the heme oxygenase (HO) pathway, which is a powerful protective cellular system, protects the diaphragm against oxidative stress and contractile failure during sepsis. A basal expression of both the inducible and constitutive HO protein isoforms (HO-1 and HO-2, respectively) was found in the diaphragm. Enhanced HO-1 expression in diaphragmatic myocytes was observed 24 h after Escherichia coli endotoxin (lipopolysaccharide, LPS) inoculation and remained elevated for at least 96 h. Enhanced HO-1 expression was also observed in the rectus abdominis and soleus muscles and in the left ventricular myocardium of endotoxemic animals. Diaphragmatic HO-2 expression was not modified by endotoxin. Diaphragmatic HO activity exhibited a biphasic time course characterized by a transient decrease during the first 12 h followed by a significant increase at 24 h, corresponding to HO-1 induction. Diaphragmatic force was significantly reduced 24 h after LPS, concomitantly with muscular oxidative stress. Administation of an inhibitor of heme oxygenase activity, zinc protoporphyrin IX (ZnPP-IX), further impaired muscular oxidative stress and contractile failure. By contrast, increased levels of HO-1 expression obtained by pretreatment of rats with hemin, a powerful inducer of HO-1, completely prevented LPS-mediated diaphragmatic oxidative stress and contractile failure. This protective effect was reversed by ZnPP-IX. These results show an important protective role for the HO pathway against sepsis-induced diaphragmatic dysfunction, which could be related to its antioxidant properties. PMID- 11254536 TI - Fas/FasL-dependent apoptosis of alveolar cells after lipopolysaccharide-induced lung injury in mice. AB - To determine the possible contribution of apoptosis in the pathogenesis of acute lung injury (ALI), we investigated Fas antigen (Fas), Fas ligand (FasL), perforin, granzyme A, and granzyme B expressions in a murine model of ALI after intratracheal instillation of Escherichia coli lipopolysaccharide (LPS: 0.3-30 microg) into the left lung. Lung injury, examined by water-to-dry weight ratio and albumin leakage, demonstrated maximal epithelial injury 1 d after 30 microg LPS instillation. Expressions of the proapoptosis molecules' mRNA were dose dependently up-regulated, with maximal expression in the early phase in the instilled lung and most apparent 1 d after LPS instillation. Negligible mRNA expression of proapoptosis molecules was observed in noninstilled lungs. The terminal deoxynucleotidyl transferase-mediated dUTP biotin nick end labeling (TUNEL) demonstrated positive signals in neutrophils and macrophages as well as in alveolar wall cells of the instilled lung 1 d after LPS instillation. Immunohistochemistry demonstrated that Fas was up-regulated in alveolar and inflammatory cells and FasL-positive inflammatory cells migrated into the air spaces in the LPS-instilled lung. Intratracheal administration of P2 antibody, which is an anti-Fas blocking antibody, attenuated the lung injury after 30 microg LPS instillation without attenuating mRNA expressions of proapoptosis molecules and neutrophil accumulation in the lung. In contrast, concanamycin A, which inhibits the function of perforin, did not alter the outcome after LPS instillation. These results indicate that the Fas/FasL system could be important in the pathogenesis of LPS-induced ALI, and proper regulation of the FasL/Fas system might be important for potential treatment of ARDS. PMID- 11254537 TI - Transient transgene expression of decorin in the lung reduces the fibrotic response to bleomycin. AB - Pulmonary fibrosis is a chronic progressive disease with no effective therapy. Transforming growth factor beta (TGF-beta) is thought to be a key profibrotic mediator and blocking its activity is therefore one of the targets of new treatment strategies for fibrosis. Decorin is an endogenous proteoglycan and one of the known inhibitors of TGF-beta. The short half-life of peptide-based therapeutics makes gene transfer a promising approach to achieve prolonged protein levels in the lung. Replication-deficient adenovirus was used to deliver decorin transgene (AdDec) to the airways by a single intranasal injection in a murine bleomycin model of lung fibrosis. The ability of vector-derived decorin to inhibit TGF-beta was examined in a bioassay and its effect on bleomycin-induced pulmonary fibrosis was determined by histomorphology and lung hydroxyproline. In vitro, supernatant from cells infected with AdDec abrogated the bioactivity of TGF-beta in a dose-dependent manner whereas control virus (AdDL70) had no effect. In vivo, treatment of bleomycin-injected mice with AdDec substantially reduced the fibrogenic response compared with control virus (hydroxyproline: bleomycin/AdDec, 1.96 microg/mg; bleomycin/AdDL70, 3.05 microg/mg; p = 0.0005). These results suggest that a single administration of AdDec was able to generate a local pulmonary environment that effectively blocked the fibrogenic response to bleomycin by inhibition of TGF-beta. PMID- 11254538 TI - The transcription factor early growth-response factor 1 modulates tumor necrosis factor-alpha, immunoglobulin E, and airway responsiveness in mice. AB - Early growth-response factor 1 (Egr-1) is a sequence-specific transcription factor that plays a regulatory role in the expression of many genes important in inflammation, cell growth, apoptosis, and the pathogenesis of disease. In vitro studies suggest that Egr-1 is capable of regulating the expression of tumor necrosis factor-alpha (TNF-alpha) and other genes involved in airway inflammation and reactivity following allergen stimulation. On the basis of these data, we hypothesized that in the absence of Egr-1, the TNF-alpha response and subsequent downstream inflammatory events that usually follow allergen challenge would be diminished. To test our hypothesis Egr-1 knock-out (KO) mice were examined in an ovalbumin (OVA)-induced model of airway inflammation and reactivity, and compared with identically treated wild-type (WT) control mice. In response to OVA sensitization and airway challenge, KO mice had diminished TNF-alpha mRNA and protein in the lungs and mast cells compared with WT mice. Interestingly, the KO mice had elevated IgE levels at baseline and after allergen challenge compared with WT mice. Furthermore, the airways of KO mice were hyporesponsive to methacholine challenge at baseline and after allergen challenge. These data indicate that Egr-1 modulates TNF-alpha, IgE, and airway responsiveness in mice. PMID- 11254539 TI - Human collagenase (matrix metalloproteinase-1) expression in the lungs of patients with emphysema. AB - Pulmonary emphysema is believed to result from an imbalance between proteolytic enzymes and their inhibitors. Multiple studies have examined the presence of various proteases within the bronchoalveolar lavage fluid from patients with chronic obstructive pulmonary disease (COPD). However, to date extensive examination of the lung parenchyma for the expression of destructive enzymes has not yet been determined. The following study examines the lung parenchyma of 23 patients with emphysema and 8 normal control samples for the expression of matrix matalloproteinase-1 (MMP-1), MMP-12, and MMP-9. We report here that interstitial collagenase (MMP-1) RNA, protein, and activity are present in the lung parenchyma of patients with emphysema and not in the lung of normal control subjects. In contrast, metalloelastase (MMP-12) expression is absent in these samples. Immunohistochemistry studies localized MMP-1 to the Type II pneumocyte in patients with emphysema and not normal control subjects or smokers without emphysema. This observation demonstrates that the lung is altered in emphysema such that the Type II pneumocyte secretes MMP-1 and suggests that MMP-1 may be an important enzyme involved in the destruction of the lung in the human disease. In addition, the induction of a proteolytic enzyme within the Type II pneumocyte suggests that the cells within the lung itself are capable of producing degradative enzymes in this disease process. PMID- 11254540 TI - Acute renal failure and erratum. PMID- 11254541 TI - Response to oxygen breathing in ALI/ARDS patients. PMID- 11254542 TI - Respiratory syncytial virus and reactive airway disease. New developments prompt a new review. PMID- 11254543 TI - Epidemiologic and clinical evidence of a respiratory syncytial virus-reactive airway disease link. PMID- 11254544 TI - Respiratory syncytial virus and asthma. The role of monocytes. PMID- 11254545 TI - Potential mechanisms causing delayed effects of respiratory syncytial virus infection. PMID- 11254546 TI - Treatment and prevention of respiratory syncytial virus lower respiratory tract infection. Long-term effects on respiratory outcomes. PMID- 11254547 TI - Neural mechanisms of respiratory syncytial virus-induced inflammation and prevention of respiratory syncytial virus sequelae. PMID- 11254548 TI - Plasminogen binding and activation by Mycoplasma fermentans. AB - The binding of plasminogen to Mycoplasma fermentans was studied by an immunoblot analysis and by a binding assay using iodine-labeled plasminogen. The binding of 125I-labeled plasminogen was inhibited by unlabeled plasminogen, lysine, and lysine analog epsilon-aminocaproic acid. Partial inhibition was obtained by a plasminogen fragment containing kringles 1 to 3 whereas almost no inhibition was observed with a fragment containing kringle 4. Scatchard analysis revealed a dual phase interaction, one with a dissociation constant (kd) of 0.5 microM and the second with a kd of 7.5 microM. The estimated numbers of plasminogen molecules bound were calculated to be 110 and 790 per cell, respectively. Autoradiograms of ligand blots containing M. fermentans membrane proteins incubated with 125I labeled plasminogen identified two plasminogen-binding proteins of about 32 and 55 kDa. The binding of plasminogen to M. fermentans enhances the activation of plasminogen to plasmin by the urokinase-type plasminogen activator (uPA), as monitored by measuring the breakdown of chromogenic substrate S-2251. Enhancement was more pronounced with the low-molecular-weight and the single-chain uPA variants, known to have low plasminogen activator activities. The binding of plasminogen also promotes the invasion of HeLa cells by M. fermentans. Invasion was more pronounced in the presence of uPA, suggesting that the ability of the organism to invade host cells stems not only from its potential to bind plasminogen but also from the activation of plasminogen to plasmin. PMID- 11254549 TI - Differential regulation of Bvg-activated virulence factors plays a role in Bordetella pertussis pathogenicity. AB - Bordetella pertussis, the causative agent of whooping cough, regulates expression of many virulence factors via a two-component signal transduction system encoded by the bvgAS regulatory locus. It has been shown by transcription activation kinetics that several of the virulence factors are differentially regulated. fha is transcribed within 10 min following a bvgAS-inducing signal, while prn is transcribed after 1 h and ptx is not transcribed until 2 to 4 h after induction. These genes therefore represent early, intermediate, and late classes of bvg activated promoters, respectively. Although there have been many insightful studies into the mechanisms of BvgAS-mediated regulation, the role that differential regulation of virulence genes plays in B. pertussis pathogenicity has not been characterized. We provide evidence that alterations to the promoter regions of bvg-activated genes can alter the kinetic pattern of expression of these genes without changing steady-state transcription levels. In addition, B. pertussis strains containing these promoter alterations that express either ptx at an early time or fha at a late time demonstrate a significant reduction in their ability to colonize respiratory tracts in an intranasal mouse model of infection. These data suggest a role for differential regulation of bvg-activated genes, and therefore for the BvgAS regulatory system, in the pathogenicity of B. pertussis. PMID- 11254550 TI - Streptococcus iniae virulence is associated with a distinct genetic profile. AB - Streptococcus iniae causes meningoencephalitis and death in commercial fish species and has recently been identified as an emerging human pathogen producing fulminant soft tissue infection. As identified by pulsed-field gel electrophoresis (PFGE), strains causing disease in either fish or humans belong to a single clone, whereas isolates from nondiseased fish are genetically diverse. In this study, we used in vivo and in vitro models to examine the pathogenicity of disease-associated isolates. Strains with the clonal (disease associated) PFGE profile were found to cause significant weight loss and bacteremia in a mouse model of subcutaneous infection. As little as 10(2) CFU of a disease-associated strain was sufficient to establish bacteremia, with higher inocula (10(7)) resulting in increased mortality. In contrast, non-disease associated (commensal) strains failed to cause bacteremia and weight loss, even at inocula of 10(8) CFU. In addition, disease-associated strains were more resistant to phagocytic clearance in a human whole blood killing assay compared to commensal strains, which were almost entirely eradicated. Disease-associated strains were also cytotoxic to human endothelial cells as measured by lactate dehydrogenase release from host cells. However, both disease-associated and commensal strains adhered to and invaded cultured human epithelial and endothelial cells equally well. While cellular invasion may still contribute to the pathogenesis of invasive S. iniae disease, resistance to phagocytic clearance and direct cytotoxicity appear to be discriminating virulence attributes of the disease-associated clone. PMID- 11254551 TI - Induction of inducible nitric oxide synthase-NO* by lipoarabinomannan of Mycobacterium tuberculosis is mediated by MEK1-ERK, MKK7-JNK, and NF-kappaB signaling pathways. AB - Nitric oxide (NO*) expression by inducible nitric oxide synthase (iNOS) is an important host defense mechanism against Mycobacterium tuberculosis in mononuclear phagocytes. The objective of this investigation was to examine the role of mitogen-activated protein (MAP) kinase (MAPK) and nuclear factor kappaB (NF-kappaB) signaling pathways in the regulation of iNOS and NO* by a mycobacterial cell wall lipoglycan known as mannose-capped lipoarabinomannan (ManLAM). Specific pharmacologic inhibition of the extracellular-signal-regulated kinase (ERK) or NF-kappaB pathway revealed that both these signaling cascades were required in gamma interferon (IFN-gamma)-ManLAM-induced iNOS protein and NO2 expression in mouse macrophages. Transient cotransfection of dominant-negative protein mutants of the c-Jun NH2-terminal kinase (JNK) pathway revealed that the MAP kinase kinase 7 (MKK7)-JNK cascade also mediated IFN-gamma-ManLAM induction of iNOS promoter activity whereas MKK4 did not. Overexpression of null mutant IkappaBalpha, a potent inhibitor of NF-kappaB activation, confirmed that the IkappaBalpha kinase (IKK)-NF-kappaB signaling pathway enhanced IFN-gamma-ManLAM induced iNOS promoter activity. By contrast, activated p38mapk inhibited iNOS induction. These results indicate that combined IFN-gamma and ManLAM stimulation induced iNOS and NO. expression and that MEK1-ERK, MKK7-JNK, IKK-NF-kappaB, and p38mapk signaling pathways play important regulatory roles. PMID- 11254552 TI - Leukotriene B4 augments neutrophil phagocytosis of Klebsiella pneumoniae. AB - Neutrophils play a critical role in the clearance of bacteria from the lung and other organs by their capacity for phagocytosis and killing. Previously, we identified an important role for the leukotrienes in rat alveolar macrophage phagocytosis of Klebsiella pneumoniae. In this report, we explored the possibility that the leukotrienes play an important role in phagocytosis by neutrophils as well. Inhibition of endogenous leukotriene synthesis by 5 lipoxygenase knockout in mice or by pharmacologic means in human peripheral blood neutrophils attenuated phagocytosis of opsonized K. pneumoniae. Reduced phagocytosis was also observed in human neutrophils pretreated with a leukotriene B4 receptor but not a cysteinyl-leukotriene receptor antagonist. While leukotriene B4 reconstituted defective phagocytosis in leukotriene-deficient neutrophils and enhanced phagocytosis in neutrophils capable of leukotriene synthesis, leukotriene C4, leukotriene D4, 5-hydroperoxyeicosatetraenoic acid, and 5-oxo-eicosatetraenoic acid were ineffective. To determine the opsonin dependence of the leukotriene B4 augmentation of phagocytosis, we assessed the ability of leukotriene B4 to modulate neutrophil phagocytosis and the adherence of sheep erythrocytes opsonized with immunoglobulin G or the complement fragment C3bi. While leukotriene B4 augmented both Fc receptor- and complement receptor mediated phagocytosis, increased adherence to leukotriene B4-treated neutrophils was limited to complement opsonized targets. In conclusion, we have identified a novel role for leukotriene B4 in the augmentation of neutrophil phagocytosis mediated by either the Fc or complement receptor. PMID- 11254553 TI - Chemokine-dependent neutrophil recruitment in a murine model of Legionella pneumonia: potential role of neutrophils as immunoregulatory cells. AB - The roles of CXC chemokine-mediated host responses were examined with an A/J mouse model of Legionella pneumophila pneumonia. After intratracheal inoculation of 10(6) CFU of L. pneumophila, the bacterial numbers in the lungs increased 10 fold by day 2; this increase was accompanied by the massive accumulation of neutrophils. Reverse transcription-PCR data demonstrated the up-regulation of CXC chemokines, such as keratinocyte-derived chemokine, macrophage inflammatory protein 2 (MIP-2), and lipopolysaccharide-induced CXC chemokine (LIX). Consistent with these data, increased levels of KC, MIP-2, and LIX proteins were observed in the lungs and peaked at days 1, 2, and 2, respectively. Although the administration of anti-KC or anti-MIP-2 antibody resulted in an approximately 20% decrease in neutrophil recruitment on day 2, no increase in mortality was observed. In contrast, the blockade of CXC chemokine receptor 2 (CXCR2), a receptor for CXC chemokines, including KC and MIP-2, strikingly enhanced mortality; this effect coincided with a 67% decrease in neutrophil recruitment. Interestingly, anti-CXCR2 antibody did not affect bacterial burden by day 2, even in the presence of a lethal challenge of bacteria. Moreover, a significant decrease in interleukin-12 (IL-12) levels, in contrast to the increases in KC, MIP-2, and LIX levels, was demonstrated for CXCR2-blocked mice. These data indicated that CXCR2-mediated neutrophil accumulation may play a crucial role in host defense against L. pneumophila pneumonia in mice. The increase in lethality without a change in early bacterial clearance suggested that neutrophils may exert their protective effect not through direct killing but through more immunomodulatory actions in L. pneumophila pneumonia. We speculate that a decrease in the levels of the protective cytokine IL-12 may explain, at least in part, the high mortality in the setting of reduced neutrophil recruitment. PMID- 11254555 TI - Mucosal vaccination with recombinantly attenuated staphylococcal enterotoxin B and protection in a murine model. AB - Previous work in our laboratory revealed that mice parenterally vaccinated with recombinantly attenuated staphylococcal enterotoxin (SE) or toxic shock syndrome toxin 1 develop protective antibodies against a lethal intraperitoneal (i.p.) toxin challenge. This study investigated the efficacy of nasal and oral immunizations with an SEB vaccine (SEBv) toward an i.p. or mucosal (via an aerosol) toxin challenge. Both vaccination routes, with the immunoadjuvant cholera toxin (CT), elicited comparable SEB-specific immunoglobulin A (IgA) and IgG levels in saliva. Nasal or oral inoculations also generated SEB-specific IgA, IgG, and IgM in the serum, but the nasal route yielded higher specific IgG titers. SEBv alone, when given nasally or orally, did not induce any detectable SEB-specific antibody. Mice vaccinated mucosally were protected against a 50% lethal dose of wild-type SEB given i.p. or mucosally, thus demonstrating that nasal or oral administration of this SEBv, with CT, elicits systemic and mucosal antibodies to SEB that protect against SEB-induced lethal shock. PMID- 11254554 TI - Micrococcus luteus teichuronic acids activate human and murine monocytic cells in a CD14- and toll-like receptor 4-dependent manner. AB - Teichuronic acid (TUA), a component of the cell walls of the gram-positive organism Micrococcus luteus (formerly Micrococcus lysodeikticus), induced inflammatory cytokines in C3H/HeN mice but not in lipopolysaccharide (LPS) resistant C3H/HeJ mice that have a defect in the Toll-like receptor 4 (TLR4) gene, both in vivo and in vitro, similarly to LPS (T. Monodane, Y. Kawabata, S. Yang, S. Hase, and H. Takada, J. Med. Microbiol. 50:4-12, 2001). In this study, we found that purified TUA (p-TUA) induced tumor necrosis factor alpha (TNF alpha) in murine monocytic J774.1 cells but not in mutant LR-9 cells expressing membrane CD14 at a lower level than the parent J774.1 cells. The TNF-alpha inducing activity of p-TUA in J774.1 cells was completely inhibited by anti-mouse CD14 monoclonal antibody (MAb). p-TUA also induced interleukin-8 (IL-8) in human monocytic THP-1 cells differentiated to macrophage-like cells expressing CD14. Anti-human CD14 MAb, anti-human TLR4 MAb, and synthetic lipid A precursor IV(A), an LPS antagonist, almost completely inhibited the IL-8-inducing ability of p TUA, as well as LPS, in the differentiated THP-1 cells. Reduced p-TUA did not exhibit any activities in J774.1 or THP-1 cells. These findings strongly suggested that M. luteus TUA activates murine and human monocytic cells in a CD14 and TLR4-dependent manner, similar to LPS. PMID- 11254556 TI - Interactions of surfactant proteins A and D with Saccharomyces cerevisiae and Aspergillus fumigatus. AB - Surfactant proteins A (SP-A) and D (SP-D) are members of the collectin family of calcium-dependent lectins and are important pulmonary host defense molecules. Human SP-A and SP-D and rat SP-D bind to Aspergillus fumigatus conidia, but the ligand remains unidentified. To identify a fungal ligand for SP-A and/or SP-D, we examined the interactions of the proteins with Saccharomyces cerevisiae. SP-D but not SP-A bound yeast cells, and EDTA inhibited the binding. SP-D also aggregated yeast cells and isolated yeast cell walls. Treating yeast cells to remove cell wall mannoprotein did not reduce SP-D binding, and SP-D failed to aggregate chitin. However, SP-D aggregated yeast glucan before and after treatment with a beta(1-->3)-glucanase, suggesting a specific interaction between the collectin and beta(1-->6)-glucan. In support of this idea, SP-D-induced yeast aggregation was strongly inhibited by pustulan [a beta(1-->6)-linked glucose homopolymer] but was not inhibited by laminarin [a beta(1-->3)-linked glucose homopolymer]. Additionally, pustulan but not laminarin strongly inhibited SP-D binding to A. fumigatus. The pustulan concentration for 50% inhibition of SP-D binding to A. fumigatus is 1.0 +/- 0.3 microM glucose equivalents. Finally, SP-D showed reduced binding to the beta(1-->6)-glucan-deficient kre6 yeast mutant. Taken together, these observations demonstrate that beta(1-->6)-glucan is an important fungal ligand for SP-D and that glycosidic bond patterns alone can determine if an extended carbohydrate polymer is recognized by SP-D. PMID- 11254557 TI - Synergistic effect of muramyldipeptide with lipopolysaccharide or lipoteichoic acid to induce inflammatory cytokines in human monocytic cells in culture. AB - An analog of 1alpha,25-dihydroxyvitamin D3, 22-oxyacalcitriol (OCT), differentiated human monocytic THP-1 and U937 cells to express membrane CD14 and rendered the cells responsive to bacterial cell surface components. Both THP-1 and U937 cells expressed Toll-like receptor 4 (TLR4) on the cell surface and TLR4 mRNA in the cells, irrespective of OCT treatment. In contrast, OCT-treated U937 cells scarcely expressed TLR2 mRNA, while OCT-treated THP-1 cells expressed this transcript. Muramyldipeptide (MDP) by itself exhibited only a weak ability to induce secretion of inflammatory cytokines such as interleukin-8 (IL-8) in the OCT-differentiated THP-1 cells but showed marked synergistic effects with Salmonella lipopolysaccharide (LPS) or lipoteichoic acid (LTA) from Staphylococcus aureus, both of which exhibited strong activities. Combinatory stimulation with LPS plus LTA did not show a synergistic effect on OCT differentiated THP-1 cells. Similar results were observed in OCT-differentiated U937 cells, although combination experiments were carried out only with MDP plus LPS. Anti-CD14 monoclonal antibody (MAb) MY4, anti-TLR4 MAb HTA125, and the synthetic lipid A precursor LA-14-PP almost completely inhibited the IL-8 inducing activities of LTA as well as LPS on OCT-treated THP-1 cells, but these treatments increased MDP activity. OCT-treated THP-1 cells primed with MDP exhibited enhanced production of IL-8 upon stimulation with LPS, while the cells primed with LPS showed no change in production upon stimulation with MDP. MDP up regulated mRNA expression of an adapter molecule to TLRs, MyD88, to an extent similar to that for LPS in OCT-treated THP-1 cells. These findings suggested that LTA as well as LPS activated human monocytic cells in a CD14- and TLR4-dependent manner, whereas MDP exhibited activity in a CD14-, TLR4-, and probably TLR2 independent manner and exhibited synergistic and priming effects on the cells for cytokine production in response to various bacterial components. PMID- 11254559 TI - Epitope mapping of monoclonal antibodies capable of neutralizing cytotoxic necrotizing factor type 1 of uropathogenic Escherichia coli. AB - Cytotoxic necrotizing factor type 1 (CNF1) of uropathogenic Escherichia coli belongs to a family of bacterial toxins that target the small GTP-binding Rho proteins that regulate the actin cytoskeleton. Members of this toxin family typically inactivate Rho; however, CNF1 and the highly related CNF2 activate Rho by deamidation. Other investigators have reported that the first 190 amino acids of CNF1 constitute the cellular binding domain and that the CNF1 enzymatic domain lies within a 300-amino-acid stretch in the C terminus of the toxin. Amino acids 53 to 75 appear to be critical for cell receptor recognition, while amino acids Cys866 and His881 are considered essential for deamidation activity. To delineate further the functional domains of CNF1, we generated 16 monoclonal antibodies (MAbs) against the toxin and used them for epitope mapping studies. Based on Western blot immunoreactivity patterns obtained from a series of truncated CNF1 proteins, this panel of MAbs mapped to epitopes located throughout the toxin, including the binding and enzymatic domains. All MAbs showed reactivity to CNF1 by Western and dot blot analyses. However, only 7 of the 16 MAbs exhibited cross reactivity with CNF2. Furthermore, only three MAbs demonstrated the capacity to neutralize toxin in either HEp-2 cell assays (inhibition of multinucleation) or 5637 bladder cell assays (inhibition of cytotoxicity). Since CNF1 epitopes recognized by neutralizing MAbs are likely to represent domains or regions necessary for the biological activities of the toxin, the epitopes recognized by these three MAbs, designated JC4 (immunoglobulin G2a [IgG2a]), BF8 (IgA), and NG8 (IgG2a), were more precisely defined. MAbs JC4 and BF8 reacted with epitopes that were common to CNF1 and CNF2 and located within the putative CNF1 binding domain. MAb JC4 recognized an epitope spanning amino acids 169 to 191, whereas MAb BF8 mapped to an epitope between amino acids 135 and 164. Despite the capacity of both MAbs to recognize CNF2 in Western blot analyses, only MAb BF8 neutralized CNF2. MAb NG8 showed reactivity to a CNF1-specific epitope located between amino acids 683 and 730, a region that includes a very small portion of the putative enzymatic domain. Taken together, these findings identify three new regions of the toxin that appear to be critical for the biological activity of CNF1. PMID- 11254558 TI - Identification of new genes involved in the virulence of Listeria monocytogenes by signature-tagged transposon mutagenesis. AB - Listeria monocytogenes is a gram-positive, facultative intracellular pathogen that can cause severe food-born infections in humans and animals. We have adapted signature-tagged transposon mutagenesis to L. monocytogenes to identify new genes involved in virulence in the murine model of infection. We used transposon Tn1545 carried on the integrative vector pAT113. Forty-eight tagged transposons were constructed and used to generate banks of L. monocytogenes mutants. Pools of 48 mutants were assembled, taking one mutant from each bank, injected into mice, and screened for those affected in their multiplication in the brains of infected animals. From 2,000 mutants tested, 18 were attenuated in vivo. The insertions harbored by these mutants led to the identification of 10 distinct loci, 7 of which corresponded to previously unknown genes. The properties of four loci involving putative cell wall components were further studied in vitro and in vivo. The data suggested that these components are involved in bacterial invasion and multiplication in the brain. PMID- 11254561 TI - Analysis of transcriptionally active gene clusters of major outer membrane protein multigene family in Ehrlichia canis and E. chaffeensis. AB - Ehrlichia canis and E. chaffeensis are tick-borne obligatory intramonocytic ehrlichiae that cause febrile systemic illness in humans and dogs, respectively. The current study analyzed the pleomorphic multigene family encoding approximately 30-kDa major outer membrane proteins (OMPs) of E. canis and E. chaffeensis. Upstream from secA and downstream of hypothetical transcriptional regulator, 22 paralogs of the omp gene family were found to be tandemly arranged except for one or two genes with opposite orientations in a 28- and a 27-kb locus in the E. canis and E. chaffeensis genomes, respectively. Each locus consisted of three highly repetitive regions with four nonrepetitive intervening regions. E. canis, in addition, had a 6.9-kb locus which contained a repeat of three tandem paralogs in the 28-kb locus. These total 47 paralogous and orthologous genes encoded OMPs of approximately 30 to 35 kDa consisting of several hypervariable regions alternating with conserved regions. In the 5'-end half of the 27-kb locus or the 28-kb locus of each Ehrlichia species, 14 paralogs were linked by short intergenic spaces ranging from -8 bp (overlapped) to 27 bp, and 8 remaining paralogs in the 3'-end half were connected by longer intergenic spaces ranging from 213 to 632 bp. All 22 paralogs, five unknown genes, and secA in the omp cluster in E. canis were transcriptionally active in the monocyte culture, and the paralogs with short intergenic spaces were cotranscribed with their adjacent genes, including the respective intergenic spaces at both the 5' and the 3' sides. Although omp genes are diverse, our results suggest that the gene organization of the clusters and the gene locus are conserved between two species of Ehrlichia to maintain a unique transcriptional mechanism for adaptation to environmental changes common to them. PMID- 11254560 TI - Fibronectin attachment protein homologue mediates fibronectin binding by Mycobacterium avium subsp. paratuberculosis. AB - Attachment of Mycobacterium avium subsp. paratuberculosis to host tissue and penetration of mucosal surfaces are pivotal events in the pathogenesis of Johne's disease. Fibronectin (FN) binding is required for attachment and internalization of several mycobacteria by epithelial cells in vitro. The objective of this study was to further characterize the FN binding activity of M. avium subsp. paratuberculosis. Although the bacteria bound FN poorly at pH above 7, brief acid pretreatment greatly enhanced FN binding within the pH range (3 to 10) studied. A 4.6-kbp fragment from an M. avium subsp. paratuberculosis genomic library was found to contain a 1,107-bp open reading frame that shows very high nucleotide sequence identity with that of the FN attachment protein (FAP) gene of M. avium subsp. avium. Pretreatment of FN with an FN-binding peptide from M. avium subsp. avium FAP abolished FN binding, indicating that M. avium subsp. paratuberculosis binds FN in a FAP-dependent manner. Pretreatment of M. avium subsp. paratuberculosis with anti-FAP immunoglobulin G did not abrogate FN binding; blocking occurred only when anti-FAP was added together with FN. FAP was detected by immunofluorescence only in lipid-extracted M. avium subsp. paratuberculosis. Western blotting and immunoelectron microscopy revealed that FAP is located near the interior of the cell envelope of M. avium subsp. paratuberculosis. The results indicate that a FAP homologue mediates the attachment of FN to M. avium subsp. paratuberculosis. Further, given the subcellular location of FAP, it is considered that this protein operates at the terminus of a coordinated FN binding system in the cell envelope of M. avium subsp. paratuberculosis. PMID- 11254562 TI - Type II protein secretion is a subset of the PilD-dependent processes that facilitate intracellular infection by Legionella pneumophila. AB - Previously, we had demonstrated that a Legionella pneumophila prepilin peptidase (pilD) mutant does not produce type IV pili and shows reduced secretion of enzymatic activities. Moreover, it displays a distinct colony morphology and a dramatic reduction in intracellular growth within amoebae and macrophages, two phenotypes that are not exhibited by a pilin (pilE(L)) mutant. To determine whether these pilD-dependent defects were linked to type II secretion, we have constructed two new mutants of L. pneumophila strain 130b. Mutations were introduced into either lspDE, which encodes the type II outer membrane secretin and ATPase, or lspFGHIJK, which encodes the pseudopilins. Unlike the wild-type and pilE(L) strains, both lspDE and lspG mutants showed reduced secretion of six pilD-dependent enzymatic activities; i.e., protease, acid phosphatase, p nitrophenol phosphorylcholine hydrolase, lipase, phospholipase A, and lysophospholipase A. However, they exhibited a colony morphology different from that of the pilD mutant, suggesting that their surfaces are distinct. The pilD, lspDE, and lspG mutants were similarly and greatly impaired for growth within Hartmannella vermiformis, indicating that the intracellular defect of the peptidase mutant in amoebae is explained by the loss of type II secretion. When assessed for infection of U937 macrophages, both lsp mutants exhibited a 10-fold reduction in intracellular multiplication and a diminished cytopathic effect. Interestingly, the pilD mutant was clearly 100-fold more defective than the type II secretion mutants in U937 cells. These results suggest the existence of a novel pilD-dependent mechanism for promoting L. pneumophila intracellular infection of human cells. PMID- 11254563 TI - Critical role of CD14 for production of proinflammatory cytokines and cytokine inhibitors during sepsis with failure to alter morbidity or mortality. AB - We investigated the immunopathophysiologic responses during sepsis induced by cecal ligation and puncture (CLP) in CD4-deficient (CD14 knockout [CD14KO]) mice. Our studies were designed to specifically test the role of CD14 in the inflammatory response to sepsis and to ascertain if alterations would improve morbidity or mortality. Sepsis was induced using the CLP model with appropriate antibiotic treatment. The severity of sepsis increased in the CD14KO mice with increasing puncture size (18 gauge [18G], 21G, and 25G). Following CLP, body temperature (at 12 h) and gross motor activity levels of the sham and 25G CLP groups recovered to normal, while the 21G and 18G CLP groups exhibited severe hypothermia coupled with decreased gross motor activity and body weight. There were no significant differences in survival, temperature, body weight, or activity levels between CD14KO and control mice after 21G CLP. However, CD14KO mice expressed two- to fourfold less pro-inflammatory (interleukin-1beta [IL 1beta], tumor necrosis factor [TNF], and IL-6) and anti-inflammatory (IL-10, IL-1 receptor antagonist, and TNF receptors I and II) cytokines in the blood after 21G CLP. Plasma levels of the chemokines macrophage inflammatory protein 2alpha and KC were similarly reduced in CD14KO mice. A similar trend of decreased cytokine and cytokine inhibitor levels was observed in the peritoneal cavity of CD14KO mice. Our results indicate that the CD14 pathway of activation plays a critical role in the production of both pro-inflammatory cytokines and cytokine inhibitors but has minimal impact on the morbidity or mortality induced by the CLP model of sepsis. PMID- 11254565 TI - Flagellum of Legionella pneumophila positively affects the early phase of infection of eukaryotic host cells. AB - Legionella pneumophila, the etiologic agent of Legionnaires' disease, contains a single, monopolar flagellum which is composed of one major subunit, the FlaA protein. To evaluate the role of the flagellum in the pathogenesis and ecology of Legionella, the flaA gene of L. pneumophila Corby was mutagenized by introduction of a kanamycin resistance cassette. Immunoblots with antiflagellin-specific polyclonal antiserum, electron microscopy, and motility assays confirmed that the specific flagellar mutant L. pneumophila Corby KH3 was nonflagellated. The redelivery of the intact flaA gene into the chromosome (L. pneumophila Corby CD10) completely restored flagellation and motility. Coculture studies showed that the invasion efficiency of the flaA mutant was moderately reduced in amoebae and severely reduced in HL-60 cells. In contrast, adhesion and the intracellular rate of replication remained unaffected. Taking these results together, we have demonstrated that the flagellum of L. pneumophila positively affects the establishment of infection by facilitating the encounter of the host cell as well as by enhancing the invasion capacity. PMID- 11254564 TI - Complete nucleotide sequence and analysis of the locus of enterocyte Effacement from rabbit diarrheagenic Escherichia coli RDEC-1. AB - The pathogenicity island termed the locus of enterocyte effacement (LEE) is found in diverse attaching and effacing pathogens associated with diarrhea in humans and other animal species. To explore the relation of variation in LEE sequences to host specificity and genetic lineage, we determined the nucleotide sequence of the LEE region from a rabbit diarrheagenic Escherichia coli strain RDEC-1 (O15:H ) and compared it with those from human enteropathogenic E. coli (EPEC, O127:H6) and enterohemorrhagic E. coli (EHEC, O157:H7) strains. Differing from EPEC and EHEC LEEs, the RDEC-1 LEE is not inserted at selC and is flanked by an IS2 element and the lifA toxin gene. The RDEC-1 LEE contains a core region of 40 open reading frames, all of which are shared with the LEE of EPEC and EHEC. orf3 and the ERIC (enteric repetitive intergenic consensus) sequence present in the LEEs of EHEC and EPEC are absent from the RDEC-1 LEE. The predicted promoters of LEE1, LEE2, LEE3, tir, and LEE4 operons are highly conserved among the LEEs, although the upstream regions varied considerably for tir and the crucial LEE1 promoter, suggesting differences in regulation. Among the shared genes, high homology (>95% identity) between the RDEC-1 and the EPEC and EHEC LEEs at the predicted amino acid level was observed for the components of the type III secretion apparatus, the Ces chaperones, and the Ler regulator. In contrast, more divergence (66 to 88% identity) was observed in genes encoding proteins involved in host interaction, such as intimin (Eae) and the secreted proteins (Tir and Esps). A comparison of the highly variable genes from RDEC-1 with those from a number of attaching and effacing pathogens infecting different species and of different evolutionary lineages was performed. Although RDEC-1 diverges from some human infecting EPEC and EHEC, most of the variation observed appeared to be due to evolutionary lineage rather than host specificity. Therefore, much of the observed hypervariability in genes involved in pathogenesis may not represent specific adaptation to different host species. PMID- 11254566 TI - Preexposure of murine macrophages to CpG oligonucleotide results in a biphasic tumor necrosis factor alpha response to subsequent lipopolysaccharide challenge. AB - Bacterial DNA and synthetic oligonucleotides containing CpG sequences (CpG-DNA and CpG-ODN) provoke a proinflammatory cytokine response (tumor necrosis factor alpha [TNF-alpha], interleukin-12 [IL-12], and IL-6) and increased mortality in lipopolysaccharide (LPS)-challenged mice via a TNF-alpha-mediated mechanism. It was hypothesized that preexposure of macrophages to CpG-ODN would result in an increased TNF-alpha response to subsequent LPS challenge in vitro. Using the murine macrophage cell line RAW 264.7, we demonstrated both a rapid proinflammatory cytokine response (TNF-alpha) and a delayed inhibitory cytokine response (IL-10) with CpG-ODN. Preexposure of macrophages to CpG-ODN for brief periods (1 to 3 h) augmented TNF-alpha secretion and mRNA accumulation following subsequent LPS challenge (1 microg/ml). However, prolonged preexposure to CpG-ODN (6 to 9 h) resulted in suppression of the TNF-alpha protein and mRNA response to LPS. The addition of anti-IL-10 antibody to CpG-ODN during preexposure resulted in an increase in the LPS-induced TNF-alpha response over that induced by CpG-ODN preexposure alone. Thus, while brief preexposure of macrophages to CpG-ODN augments the proinflammatory cytokine response to subsequent LPS challenge, prolonged preexposure elicits IL-10 production, which inhibits the TNF-alpha response. Although the initial proinflammatory effects of CpG-DNA are well established, the immune response to CpG-DNA may also include autocrine or paracrine feedback mechanisms, leading to a complex interaction of proinflammatory and inhibitory cytokines. PMID- 11254567 TI - DNA vaccines expressing a fusion product of outer surface proteins A and C from Borrelia burgdorferi induce protective antibodies suitable for prophylaxis but Not for resolution of Lyme disease. AB - DNA vaccines encoding the outer surface protein A (OspA) of Borrelia burgdorferi have been shown to induce protective humoral responses capable of preventing but not curing infection in mice. Subsequent studies showed that an established infection or disease could be resolved by passive transfer of antibodies to OspC. In the present study, DNA vaccines encoding either the OspC antigen alone or fused to OspA and under the transcriptional control of the human elongation factor 1alpha promoter were evaluated for their protective and/or curative potential. In contrast to ospA-containing plasmids, none of the six constructs with ospC alone were immunogenic in vivo, independent of whether they contained promoter or leader sequences from ospA and/or ospC, or alternatively, the signal sequence of the human tissue plasminogen activator. Solely, a DNA vaccine encoding an OspA-OspC fusion product led to expression of the respective polypeptide chain in transfected cells in vitro and to the induction of OspA- and OspC-specific antibodies in vivo. Immune sera raised against the OspA-OspC fusion product conveyed full protection against subsequent infection, most probably via OspA-specific antibodies, but were unable to resolve infection. PMID- 11254568 TI - Reduced virulence of a Bordetella bronchiseptica siderophore mutant in neonatal swine. AB - One means by which Bordetella bronchiseptica scavenges iron is through production of the siderophore alcaligin. A nonrevertible alcaligin mutant derived from the virulent strain 4609, designated DBB25, was constructed by insertion of a kanamycin resistance gene into alcA, one of the genes essential for alcaligin biosynthesis. The virulence of the alcA mutant in colostrum-deprived, caesarean delivered piglets was compared with that of the parent strain in two experiments. At 1 week of age, piglets were inoculated with phosphate-buffered saline, 4609, or DBB25. Two piglets in each group were euthanatized on day 10 postinfection. The remainder were euthanatized at 21 days postinfection. Clinical signs, including fever, coughing, and sneezing, were present in both groups. Nasal washes performed 7, 14, and 21 days postinoculation demonstrated that strain DBB25 colonized the nasal cavity but did so at levels that were significantly less than those achieved by strain 4609. Analysis of colonization based on the number of CFU per gram of tissue recovered from the turbinate, trachea, and lung also demonstrated significant differences between DBB25 and 4609, at both day 10 and day 21 postinfection. Mild to moderate turbinate atrophy was apparent in pigs inoculated with strain 4609, while turbinates of those infected with strain DBB25 developed no or mild atrophy. We conclude from these results that siderophore production by B. bronchiseptica is not essential for colonization of swine but is required for maximal virulence. B. bronchiseptica mutants with nonrevertible defects in genes required for alcaligin synthesis may be candidates for evaluation as attenuated, live vaccine strains in conventionally reared pigs. PMID- 11254569 TI - Characterization of a cell surface protein of Clostridium difficile with adhesive properties. AB - Our laboratory has previously shown that Clostridium difficile adherence to cultured cells is enhanced after heat shock at 60 degrees C and that it is mediated by a proteinaceous surface component. The present study was undertaken to identify the surface molecules of this bacterium that could play a role in its adherence to the intestine. The cwp66 gene, encoding a cell surface-associated protein of C. difficile 79-685, was isolated by immunoscreening of a C. difficile gene library with polyclonal antibodies against C. difficile heated at 60 degrees C. The Cwp66 protein (66 kDa) contains two domains, each carrying three imperfect repeats and one presenting homologies to the autolysin CwlB of Bacillus subtilis. A survey of 36 strains of C. difficile representing 11 serogroups showed that the 3' portion of the cwp66 gene is variable; this was confirmed by sequencing of cwp66 from another strain, C-253. Two recombinant protein fragments corresponding to the two domains of Cwp66 were expressed in fusion with glutathione S transferase in Escherichia coli and purified by affinity chromatography using gluthatione-Sepharose 4B. Antibodies raised against the two domains recognized Cwp66 in bacterial surface extracts. By immunoelectron microscopy, the C-terminal domain was found to be cell surface exposed. When used as inhibitors in cell binding studies, the antibodies and protein fragments partially inhibited adherence of C. difficile to cultured cells, confirming that Cwp66 is an adhesin, the first to be identified in clostridia. PMID- 11254570 TI - Induction of protective immunity against Streptococcus mutans colonization after mucosal immunization with attenuated Salmonella enterica serovar typhimurium expressing an S. mutans adhesin under the control of in vivo-inducible nirB promoter. AB - The purpose of the present study was to evaluate the effectiveness of an attenuated Salmonella enterica serovar Typhimurium vaccine strain expressing the saliva-binding region (SBR) of the Streptococcus mutans antigen I/II adhesin, either alone or linked with the mucosal adjuvant cholera toxin A2 and B subunits (CTA2/B) and under the control of the anaerobically inducible nirB promoter, in inducing a protective immune response against S. mutans infection. BALB/c mice were immunized by either the intranasal or the intragastric route with a single dose of 10(9) or 10(10) Salmonella CFU, respectively. The Salmonella vaccine strain expressing an unrelated antigen (fragment C of tetanus toxin [TetC]) was also used for immunization as a control. Samples of serum and secretion (saliva and vaginal washes) were collected prior to and following immunization and assessed for antibody activity by enzyme-linked immunosorbent assay. Anti-SBR antibodies were detected in the serum and saliva of experimental animals by week 3 after immunization. A booster immunization at week 17 after the initial immunization resulted in enhanced immune responses to the SBR. The serum immunoglobulin G subclass profiles were indicative of T helper type 1 responses against both the vector and the SBR antigen. To determine the effectiveness of these responses on the protection against S. mutans infection, mice were challenged after the second immunization with a virulent strain of S. mutans which was resistant to tetracycline and erythromycin. Prior to the challenge, mice were treated for 5 days with tetracycline, erythromycin, and penicillin. S. mutans was initially recovered from all of the challenged mice. This bacterium persisted at high levels for at least 5 weeks in control TetC-immunized or nonimmunized mice despite the reappearance of indigenous oral organisms. However, mice immunized with Salmonella clones expressing SBR or SBR-CTA2/B demonstrated a significant reduction in the number of S. mutans present in plaque compared to the control groups. These results provide evidence for the effectiveness of the Salmonella vector in delivering the SBR antigen for the induction of mucosal and systemic immune responses to SBR. Furthermore, the induction of a salivary anti SBR response corresponded with protection against S. mutans colonization of tooth surfaces. PMID- 11254571 TI - DNA from protozoan parasites Babesia bovis, Trypanosoma cruzi, and T. brucei is mitogenic for B lymphocytes and stimulates macrophage expression of interleukin 12, tumor necrosis factor alpha, and nitric oxide. AB - The activation of innate immune responses by genomic DNA from bacteria and several nonvertebrate organisms represents a novel mechanism of pathogen recognition. We recently demonstrated the CpG-dependent mitogenic activity of DNA from the protozoan parasite Babesia bovis for bovine B lymphocytes (W. C. Brown, D. M. Estes, S. E. Chantler, K. A. Kegerreis, and C. E. Suarez, Infect. Immun. 66:5423-5432, 1998). However, activation of macrophages by DNA from protozoan parasites has not been demonstrated. The present study was therefore conducted to determine whether DNA from the protozan parasites B. bovis, Trypanosoma cruzi, and T. brucei activates macrophages to secrete inflammatory mediators associated with protective immunity. DNA from Escherichia coli and all three parasites stimulated B-lymphocyte proliferation and increased macrophage production of interleukin-12 (IL-12), tumor necrosis factor alpha (TNF-alpha), and nitric oxide (NO). Regulation of IL-12 and NO production occurred at the level of transcription. The amounts of IL-12, TNF-alpha, and NO induced by E. coli and protozoal DNA were strongly correlated (r2 > 0.9) with the frequency of CG dinucleotides in the genome, and immunostimulation by DNA occurred in the order E. coli > or = T. cruzi > T. brucei > B. bovis. Induction of inflammatory mediators by E. coli, T. brucei, and B. bovis DNA was dependent on the presence of unmethylated CpG dinucleotides. However, at high concentrations, E. coli and T. cruzi DNA-mediated macrophage activation was not inhibited following methylation. The recognition of protozoal DNA by B lymphocytes and macrophages may provide an important innate defense mechanism to control parasite replication and promote persistent infection. PMID- 11254572 TI - Effect of mycobacterial phospholipids on interaction of Mycobacterium tuberculosis with macrophages. AB - This study demonstrates that pretreatment of macrophages with phosphatidylinositol, of either soya bean or mycobacterial origin, results in a down-regulation of the binding and uptake of Mycobacterium tuberculosis by the phagocytes. We also describe the novel observation that cardiolipin induces an increase in the binding and uptake of M. tuberculosis by macrophages. Neither phospholipid interacts with macrophages via the 2F8 epitope of scavenger receptor A, and treatment of macrophages with either phospholipid results in a down regulation of CR3 function and tumor necrosis factor alpha production by the phagocyte. We have also shown that the ability of macrophages to interact with mycobacteria is greatly affected by an as yet unidentified product from the interaction of chloroform and polypropylene tubes. PMID- 11254573 TI - Spa33, a cell surface-associated subunit of the Mxi-Spa type III secretory pathway of Shigella flexneri, regulates Ipa protein traffic. AB - The Mxi-Spa type III secretion system of Shigella flexneri directs the host cell contact-induced secretion of a set of invasins, referred to as Ipas. In this study, we examined the role of Spa33 in Ipa secretion. A spa33-null mutant was both noninvasive and unable to translocate the Ipas from inner membrane to outer membrane (OM) positions of the Mxi-Spa transmembrane channel. Spa33 was found to be a Mxi-Spa substrate that is translocated to the bacterial cell surface upon the induction of Ipa secretion. This mobility may serve to drive Ipa translocation within Mxi-Spa toward OM positions. Consistent with a second distinct role in regulating Ipa traffic, the overexpression of Spa33 also blocked Ipa secretion and resulted in Ipa accumulation at the OM. Co-overexpression of Spa33 and another OM-associated element, Spa32, did not disrupt Ipa secretion, suggesting an interaction between the two proteins and an effect on the mechanism which serves to regulate Ipa release from the OM. These findings indicate that Spa33 is a mobile element within Mxi-Spa, which is required to control Ipa translocation into and out of OM positions of the secretory structure. PMID- 11254574 TI - Activation of endothelium by Borrelia burgdorferi in vitro enhances transmigration of specific subsets of T lymphocytes. AB - Lyme disease, caused by Borrelia burgdorferi, is characterized by the accumulation of lymphocytes and monocytes in the affected tissue. Endothelial cells line the blood vessel walls and control the trafficking of inflammatory leukocytes from the blood into the surrounding tissues. A model of the blood vessel wall, consisting of human umbilical vein endothelial cells (HUVEC) grown on amniotic connective tissue, was utilized to examine the effects of B. burgdorferi on the transendothelial migration of T lymphocytes. Maximal migration occurred when the HUVEC-amnion cultures were preincubated with B. burgdorferi for 24 h and T lymphocytes were added for an additional 4 h, yielding a two- to fourfold increase compared to migration across unstimulated cultures. The number of T lymphocytes that migrated was proportional to the number added. The anti inflammatory cytokine interleukin 10 (IL-10), added during activation of the HUVEC, significantly diminished (by an average of 70% +/- 21%) the migration of T lymphocytes across endothelium stimulated for 8 or 24 h with B. burgdorferi, but not IL-1. Compared to the initially added population of T lymphocytes, the population that migrated across untreated endothelium or HUVEC activated with B. burgdorferi or IL-1 contained a significantly smaller percentage of CD45RA+RO- (naive) cells and a greater proportion of CD45RA+RO+ cells. The migratory population was also enriched for CD8+ T lymphocytes when the endothelium was incubated with either control medium or B. burgdorferi, but not IL-1. B. burgdorferi thus activates endothelium in a manner that promotes the transmigration of T lymphocytes, and IL-10 inhibits this activation. These data further suggest that endothelium plays an active role in promoting the recruitment of specific subpopulations of T lymphocytes. PMID- 11254575 TI - Comparison of the exoS gene and protein expression in soil and clinical isolates of Pseudomonas aeruginosa. AB - Exoenzyme S (ExoS) is translocated into eukaryotic cells by the type III secretory process and has been hypothesized to function in conjunction with other virulence factors in the pathogenesis of Pseudomonas aeruginosa. To gain further understanding of how ExoS might contribute to P. aeruginosa survival and virulence, ExoS expression and the structural gene sequence were determined in P. aeruginosa soil isolates and compared with ExoS of clinical isolates. Significantly higher levels of ExoS ADP-ribosyltransferase (ADPRT) activity were detected in culture supernatants of soil isolates compared to those of clinical isolates. The higher levels of ADPRT activity of soil isolates reflected both the increased production of ExoS and the production of ExoS having a higher specific activity. ExoS structural gene sequence comparisons found the gene to be highly conserved among soil and clinical isolates, with the greatest number of nonsynonymous substitutions occurring within the region of ExoS encoding GAP function. The lack of amino acid changes in the ADPRT region in association with a higher specific activity implies that other factors produced by P. aeruginosa or residues outside the ADPRT region are affecting ExoS ADPRT activity. The data are consistent with ExoS being integral to P. aeruginosa survival in the soil and suggest that, in the transition of P. aeruginosa from the soil to certain clinical settings, the loss of ExoS expression is favored. PMID- 11254576 TI - Cloning and expression of the gene which encodes a tube precipitin antigen and wall-associated beta-glucosidase of Coccidioides immitis. AB - We report the structure and expression of the Coccidioides immitis BGL2 gene which encodes a previously characterized 120-kDa glycoprotein of this fungal respiratory pathogen. The glycoprotein is recognized by immunoglobulin M tube precipitin (TP) antibody present in sera of patients with coccidioidomycosis, a reaction which has been used for serodiagnosis of early coccidioidal infection. The deduced amino acid sequence of BGL2 shows 12 potential N glycosylation sites and numerous serine-threonine-rich regions which could function as sites for O glycosylation. In addition, the protein sequence includes a domain which is characteristic of family 3 glycosyl hydrolases. Earlier biochemical studies of the purified 120-kDa TP antigen revealed that it functions as a beta-glucosidase (EC 3.2.1.21). Its amino acid sequence shows high homology to several other reported fungal beta-glucosidases which are members of the family 3 glycosyl hydrolases. Results of previous studies have also suggested that the 120-kDa beta glucosidase participates in wall modification during differentiation of the parasitic cells (spherules) of C. immitis. In this study we showed that expression of the BGL2 gene is elevated during isotropic growth of spherules and the peak of wall-associated BGL2 enzyme activity correlates with this same phase of parasitic cell differentiation. These data support our hypothesis that the 120 kDa beta-glucosidase plays a morphogenetic role in the parasitic cycle of C. immitis. PMID- 11254577 TI - Human monoclonal antibodies against Pseudomonas aeruginosa lipopolysaccharide derived from transgenic mice containing megabase human immunoglobulin loci are opsonic and protective against fatal pseudomonas sepsis. AB - Pseudomonas aeruginosa is a significant human pathogen, and no vaccine is commercially available. Passive antibody prophylaxis using monoclonal antibodies (MAb) against protective P. aeruginosa epitopes is an alternative strategy for preventing P. aeruginosa infection, but mouse MAb are not suitable for use in humans. Polyclonal human antibodies from multiple donors have variable antibody titers, and human MAb are difficult to make. We used immunoglobulin-inactivated transgenic mice reconstituted with megabase-size human immunoglobulin loci to generate a human MAb against the polysaccharide (PS) portion of the lipopolysaccharide O side chain of a common pathogenic serogroup of P. aeruginosa, 06ad. The anti-PS human immunoglobulin G2 MAb made from mice immunized with heat-killed P. aeruginosa was specific for serogroup 06ad pseudomonas. The MAb was highly opsonic for the uptake and killing of P. aeruginosa by human polymorphonuclear leukocytes in the presence of human complement. In addition, 25 microg of the MAb protected 100% of neutropenic mice from fatal P. aeruginosa sepsis. DNA sequence analysis of the genes encoding the MAb revealed V(H)3 and Vkappa2/A2 variable-region genes, similar to variable region genes in humans immunized with bacterial PS and associated with high avidity anti-PS antibodies. We conclude that human MAb to P. aeruginosa made in these transgenic mice are highly protective and that these mice mimic the antibody response seen in humans immunized with T-cell-independent antigens such as bacterial PS. PMID- 11254578 TI - Membrane-associated proteins of a lipopolysaccharide-deficient mutant of Neisseria meningitidis activate the inflammatory response through toll-like receptor 2. AB - The recent isolation of a lipopolysaccharide (LPS)-deficient mutant of Neisseria meningitidis has allowed us to explore the roles of other gram-negative cell wall components in the host response to infection. The experiments in this study were designed to examine the ability of this mutant strain to activate cells. Although it was clearly less potent than the parental strain, we found the LPS-deficient mutant to be a capable inducer of the inflammatory response in monocytic cells, inducing a response similar to that seen with Staphylococcus aureus. Cellular activation by the LPS mutant was related to expression of CD14, a high-affinity receptor for LPS and other microbial products, as well as Toll-like receptor 2, a member of the Toll family of receptors recently implicated in host responses to gram-positive bacteria. In contrast to the parental strain, the synthetic LPS antagonist E5564 did not inhibit the LPS-deficient mutant. We conclude that even in the absence of LPS, the gram-negative cell wall remains a potent inflammatory stimulant, utilizing signaling pathways independent of those involved in LPS signaling. PMID- 11254579 TI - Evidence that the Campylobacter fetus sap locus is an ancient genomic constituent with origins before mammals and reptiles diverged. AB - Campylobacter fetus bacteria, isolated from both mammals and reptiles, may be either subsp. fetus or subsp. venerealis and either serotype A or serotype B. Surface layer proteins, expressed and secreted by genes in the sap locus, play an important role in C. fetus virulence. To assess whether the sap locus represents a pathogenicity island and to gain further insights into C. fetus evolution, we examined several C. fetus genes in 18 isolates. All of the isolates had 5 to 9 sapA or sapB homologs. One strain (85-387) possessed both sapA and sapB homologs, suggesting a recombinational event in the sap locus between sapA and sapB strains. When we amplified and analyzed nucleotide sequences from portions of housekeeping gene recA (501 bp) and sapD (450 bp), a part of the 6-kb sap invertible element, the phylogenies of the genes were highly parallel. Among the 15 isolates from mammals, serotype A and serotype B strains generally had consistent positions. The fact that the serotype A C. fetus subsp. fetus and subsp. venerealis strains were on the same branch suggests that their differentiation occurred after the type A-type B split. Isolates from mammals and reptiles formed two distinct tight phylogenetic clusters that were well separated. Sequence analysis of 16S rRNA showed that the reptile strains form a distinct phylotype between mammalian C. fetus and Campylobacter hyointestinalis. The phylogenies and sequence results showing that sapD and recA have similar G + C contents and substitution rates suggest that the sap locus is not a pathogenicity island but rather is an ancient constituent of the C. fetus genome, integral to its biology. PMID- 11254580 TI - Different regions of the malaria merozoite surface protein 1 of Plasmodium chabaudi elicit distinct T-cell and antibody isotype responses. AB - In this study we have investigated the antibody and CD4 T-cell responses to the well-characterized malaria vaccine candidate MSP-1 during the course of a primary Plasmodium chabaudi chabaudi (AS) infection. Specific antibody responses can be detected within the first week of infection, and CD4 T cells can be detected after 3 weeks of infection. The magnitude of the CD4 T-cell response elicited during a primary infection depended upon the region of MSP-1. In general, the highest precursor frequencies were obtained when a recombinant MSP-1 fragment corresponding to amino acids 900 to 1507 was used as the antigen in vitro. By contrast, proliferative and cytokine responses against amino acids 1508 to 1766 containing the C-terminal 21-kDa region of the molecule were low. The characteristic interleukin 4 (IL-4) switch that occurs in the CD4 T-cell population after an acute blood stage P. c. chabaudi infection was only consistently observed in the response to the amino acid 900 to 1507 MSP1 fragment. A lower frequency of IL-4-producing cells was seen in response to other regions. Although the magnitudes of the immunoglobulin G antibody responses to the different regions of MSP-1 were similar, the isotype composition of each response was distinct, and there was no obvious relationship with the type of T helper cells generated. Interestingly, a relatively high antibody response to the C-terminal region of MSP-1 was observed, suggesting that T-cell epitopes outside of this region may provide the necessary cognate help for specific antibody production. PMID- 11254581 TI - Stage-dependent role of nitric oxide in control of Trypanosoma cruzi infection. AB - Trypanosoma cruzi, the causative agent of Chagas' disease, is known to be susceptible to nitric oxide (NO)-dependent killing by gamma interferon-activated macrophages. Mice deficient for inducible nitric oxide synthase (iNOS) are highly susceptible to T. cruzi, and inhibition of iNOS from the beginning of infection was reported to lead to an increase in trypomastigotes in the blood and to high mortality. In the present study, we investigated whether NO production is essential for the control of T. cruzi in all phases of the infection. BALB/c mice were treated at different time intervals after T. cruzi infection with an iNOS inhibitor, aminoguanidine or L-N6-(1-iminoethyl)-lysine (L-NIL). Treatment initiated with the beginning of the infection resulted in 100% mortality by day 16 postinfection (p.i.). If treatment was started later during the acute phase at the peak of parasitemia (day 20 p.i.), all the mice survived. Parasitemia was cleared and tissue amastigotes became undetectable in these mice even in the presence of the iNOS inhibitor L-NIL. Inhibition of iNOS in the chronic phase of the infection, i.e., from day 60 to day 120 p.i., with L-NIL did not result in a reappearance of parasitemia. These data suggest that while NO is essential for T. cruzi control in the early phase of acute infection, it is dispensable in the late acute and chronic phase, revealing a fundamental difference in control mechanisms compared to those in infections by other members of the order Kinetoplastida, e.g., Leishmania major. PMID- 11254582 TI - Plesiomonas shigelloides enters polarized human intestinal Caco-2 cells in an in vitro model system. AB - This study provides the first definitive evidence that the gram-negative bacterium Plesiomonas shigelloides adheres to and enters eukaryotic intestinal host cells in vitro. P. shigelloides is increasingly regarded as an emerging enteric pathogen and has been implicated in intestinal and extraintestinal infections in humans. However, the establishment of its true role in enteric disease has been hindered by inadequacies in experimental design, deficiencies in clinical diagnosis, and the lack of an appropriate animal model. In this investigation, an in vitro system was used to evaluate plesiomonad pathogenesis. Differentiated epithelium-derived Caco-2 cell monolayers inoculated apically with 12 isolates of P. shigelloides from clinical (intestinal) origins were examined at high resolution using transmission electron microscopy. Bacterial cells were observed adhering to intact microvilli and to the plasma membrane on both the apical and the basal surfaces of the monolayer. The bacteria entered the Caco-2 cells and were observed enclosed in single and multiple membrane-bound vacuoles within the host cell cytoplasm. This observation suggests that initial uptake may occur through a phagocytic-like process, as has been documented for many other enteropathogens. P. shigelloides also was noted free in the cytosol of Caco-2 cells, suggesting escape from cytoplasmic vacuoles. Differences in invasion phenotypes were revealed, suggesting the possibility that, like Escherichia coli, P. shigelloides comprises different pathogenic phenotypes. PMID- 11254583 TI - Micrococci and peptidoglycan activate TLR2-->MyD88-->IRAK-->TRAF-->NIK-->IKK-->NF kappaB signal transduction pathway that induces transcription of interleukin-8. AB - This study was done to elucidate the signal transduction pathway of interleukin-8 (IL-8) induction by gram-positive bacteria. Bacteria (micrococci) and peptidoglycan (PGN) induced transcription of IL-8 in HEK293 cells expressing Toll like receptor 2 (TLR2) and CD14 but not in those expressing TLR1 or TLR4. A mutation within the NF-kappaB site in the IL-8 promoter abrogated transcriptional induction of IL-8 by the two stimulants. Dominant negative myeloid differentiation protein (MyD88), IL-1 receptor-associated kinase (IRAK), NFkappaB inducing kinase (NIK), and IkappaB kinase (IKK) mutant forms completely inhibited micrococcus- and PGN-induced activation of NF-kappaB and expression of the gene for IL-8. Induction of NF-kappaB was partially inhibited by dominant negative tumor necrosis factor receptor-associated kinase 6 (TRAF6) but not TRAF2, whereas induction of IL-8 gene was partially inhibited by both TRAF6 and TRAF2. These data indicate that micrococci and PGN induce TLR2-dependent activation of the gene for IL-8 and that this activation requires MyD88, IRAK, NIK, IKK, and NF kappaB and may also utilize TRAF6 and, to a lesser extent, TRAF2. PMID- 11254584 TI - Different subsets of enteric bacteria induce and perpetuate experimental colitis in rats and mice. AB - Resident bacteria are incriminated in the pathogenesis of experimental colitis and inflammatory bowel diseases. We investigated the relative roles of various enteric bacteria populations in the induction and perpetuation of experimental colitis. HLA-B27 transgenic rats received antibiotics (ciprofloxacin, metronidazole, or vancomycin-imipenem) in drinking water or water alone in either prevention or treatment protocols. Mice were treated similarly with metronidazole or vancomycin-imipenem before or after receiving 5% dextran sodium sulfate (DSS). Germfree transgenic rats were colonized with specific-pathogen-free enteric bacteria grown overnight either in anaerobic or aerobic atmospheres. Nontransgenic rats colonized with anaerobic bacteria served as negative controls. Although preventive metronidazole significantly attenuated colitis in transgenic rats and DSS-treated mice, it had no therapeutic benefit once colitis was established. Ciprofloxacin also partially prevented but did not treat colitis in B27 transgenic rats. In both animal models vancomycin-imipenem most effectively prevented and treated colitis. Germfree transgenic rats reconstituted with enteric bacteria grown under anaerobic conditions had more aggressive colitis than those associated with aerobic bacteria. These results suggest that a subset of resident luminal bacteria induces colitis, but that a complex interaction of commensal aerobic and anaerobic bacteria provides the constant antigenic drive for chronic immune-mediated colonic inflammation. PMID- 11254585 TI - Lack of expansion of major histocompatibility complex class Ib-restricted effector cells following recovery from secondary infection with the intracellular pathogen Listeria monocytogenes. AB - Sublethal infection of BALB/c mice with the intracellular bacterial pathogen Listeria monocytogenes leads to the development of antilisterial immunity with concurrent stimulation of major histocompatibility complex (MHC) class Ia- and Ib restricted CD8+ effector T cells. Secondary L. monocytogenes infection is followed by an accelerated increase in the number of Listeria-specific CD8+ cells and rapid clearance of the bacterium from the murine host. Recovery from secondary infection is associated with increased levels of effector cell function, as measured by gamma interferon secretion following coculture of immune cells with L. monocytogenes infected APCs in vitro, as well as antilisterial cytotoxicity, as measured by effector cell recognition of L. monocytogenes infected target cells. We assessed the frequency of L. monocytogenes-specific MHC class I-restricted cells following secondary infection by ELISPOT assays utilizing coculture of immune cells with L. monocytogenes-infected antigen presenting cells that express MHC class Ia and/or Ib molecules. We found that the antilisterial Qa-1b (MHC class Ib)-restricted effector subset is not detected as an expanded population following secondary infection compared to the frequency of this effector population as measured following recovery from primary infection. This is in contrast to the frequency of antilisterial H2-Kd (MHC class Ia) restricted effector cells, which following recovery from secondary infection are detected as an expanded population, and appears to undergo a substantial expansion event 3 to 4 days post-secondary infection. These results are consistent with the conclusion that although Listeria-specific MHC class Ib restricted effector cells are present following recovery from secondary infection, this subset does not appear to undergo the expansion phase that is detected for the MHC class Ia-restricted effector cell response. PMID- 11254586 TI - Salmonella enterica serovar typhimurium induces cell death in bovine monocyte derived macrophages by early sipB-dependent and delayed sipB-independent mechanisms. AB - It was previously demonstrated that Salmonella enterica serovar Typhimurium induces cell death with features of apoptosis in murine macrophages. Mice infected with Salmonella serovar Typhimurium develop systemic disease without diarrhea, whereas the infection in cattle and in humans is localized and characterized by diarrhea. Considering these clinical disease expression differences between mice and cattle, we investigated whether serovar Typhimurium is cytotoxic for bovine macrophages. Macrophages infected with serovar Typhimurium grown in the logarithmic phase quickly underwent cell death. Macrophages infected with stationary-phase cultures or with a mutant lacking sipB underwent no immediate cell death but did develop delayed cytotoxicity, undergoing cell death between 12 and 18 h postinfection. Both pathways were temporarily blocked by the general caspase inhibitor Z-VAD-Fmk and by the caspase 1 inhibitor Z-YVAD-Fmk. Comparisons of macrophages from cattle naturally resistant or susceptible to intracellular pathogens indicated no differences between these two genetic backgrounds in terms of susceptibility to serovar Typhimurium-induced cell death. We conclude that Salmonella serovar Typhimurium induces cell death in bovine macrophages by two distinct mechanisms, early sipB mediated and delayed sipB-independent mechanisms. PMID- 11254587 TI - Antibody against surface-bound C5a peptidase is opsonic and initiates macrophage killing of group B streptococci. AB - The capsular polysaccharides of group B streptococci (GBS) are a primary focus of vaccine development. Immunogenicity and long-lasting protection are best achieved by conjugating polysaccharides to a T-cell-dependent protein antigen. Streptococcal C5a peptidase (SCPB) is a conserved surface protein that is expressed by all streptococcal serotypes tested to date, and it is a possible carrier protein that could itself induce a protective immune response. Clearance of GBS from lungs, mucosal surfaces, or blood probably depends on the opsonophagocytic response of tissue-specific macrophages and polymorphonuclear leukocytes (PMNs). In this study, we examined the potential of antibody directed against SCPB from a serotype II strain to enhance the capacity of mouse bone marrow macrophages (from primary cultures) and human PMNs in whole blood to kill GBS in vitro. Our experiments demonstrated that Streptococcus serotypes Ia, Ib, II, III, and V, preopsonized with anti-SCPB antibody, were killed more rapidly by cultured macrophages and PMNs in whole blood than were nonopsonized GBS. The increased rate of killing was accompanied by an increased macrophage oxidative burst. Furthermore, opsonization was serotype transparent. Immunization with SCPB conjugated to capsular polysaccharide type III produced polysaccharide-specific antibodies. It is interesting that this antiserum promoted serotype-independent killing of streptococci. These data support the use of SCPB in a GBS polysaccharide conjugate vaccine. SCPB not only enhanced the immunogenicity of polysaccharide components of the vaccine, but it might also induce additional serotype-independent protective antibodies. PMID- 11254588 TI - Essential role for cellular phosphoglucomutase in virulence of type 3 Streptococcus pneumoniae. AB - Synthesis of the Streptococcus pneumoniae type 3 capsule requires the pathway glucose-6-phosphate (Glc-6-P) --> Glc-1-P --> UDP-Glc --> UDP-glucuronic acid (UDP-GlcUA) --> (GlcUA-Glc)(n). The UDP-Glc dehydrogenase and synthase necessary for the latter two steps, and essential for capsule production, are encoded by genes (cps3D and cps3S, respectively) located in the type 3 capsule locus. The phosphoglucomutase (PGM) and Glc-1-P uridylyltransferase activities necessary for the first two steps are derived largely through the actions of cellular enzymes. Homologues of these enzymes, encoded by cps3M and cps3U in the type 3 locus, are not required for capsule production. Here, we show that cps3M and cps3U also are not required for mouse virulence. In contrast, nonencapsulated isolates containing defined mutations in cps3D and cps3S were avirulent, as were reduced capsule isolates containing mutations in pgm. Insertion mutants that lacked PGM activity were avirulent in both immunologically normal (BALB/cByJ) and immunodeficient (CBA/N) mice. In contrast, a mutant (JY1060) with reduced PGM activity was avirulent in the former but had only modestly reduced virulence in the latter. The high virulence in CBA/N mice was not due to the lack of antibodies to phosphocholine but reflected a growth environment distinct from that found in BALB/cByJ mice. The reduced PGM activity of JY1060 resulted in enhanced binding of complement and antibodies to surface antigens. However, decomplementation of BALB/cByJ mice did not enhance the virulence of this mutant. Suppressor mutations, only some of which resulted in increased capsule production, increased the virulence of JY1060 in BALB/cByJ mice. The results suggest that PGM plays a critical role in pneumococcal virulence by affecting multiple cellular pathways. PMID- 11254589 TI - Novel molecular variants of allele I of the Escherichia coli P fimbrial adhesin gene papG. AB - P fimbriae of extraintestinal pathogenic Escherichia coli mediate digalactoside specific adherence via the tip adhesin molecule PapG, which occurs in three known variants (I to III), which are encoded by the corresponding three alleles of papG. In the present study, newly discovered variants of papG allele I and the respective wild-type source strains were characterized. One of the new papG allele I variants conferred a unique agglutination phenotype that combined the phenotypes associated with papG alleles I, II, and III. Comparative hydrophilicity analysis of predicted PapG peptides revealed regions that might explain the observed phenotypic similarities and differences between the PapG variants. The new papG allele I variants occurred either as the sole papG allele or together with both papG alleles II and III, rather than with only papG allele III, as in archetypal strains J96 and CP9. They also occurred in the absence of the usual F13 papA allele. One of the new papG allele I variants occurred in a serogroup O6 strain that, according to random amplified polymorphic DNA analysis, was phylogenetically distant from the "J96-like" clonal group of E. coli O4:H5, which includes all previously identified examples of papG allele I. Cluster analysis of nucleotide and predicted peptide sequences suggested that papG allele I represents the earliest evolutionary branch from a common papG ancestor. These results demonstrate unexpected diversity within papG allele I and, together with previous findings, suggest that the J96-like clonal group of E. coli O4:H5 may represent the original source of papG within the species. PMID- 11254590 TI - Immunoglobulin A (IgA) responses and IgE-associated inflammation along the respiratory tract after mucosal but not systemic immunization. AB - The purpose of the present study was to determine the extent of immunologic responses, particularly immunopathologic responses, within the upper and lower respiratory tracts after intranasal immunization using the mucosal adjuvant cholera toxin (CT). BALB/c mice were nasally immunized with influenza virus vaccine combined with CT. The inclusion of the mucosal adjuvant CT clearly enhanced generation of antibody responses in both the nasal passages and lungs. After nasal immunization, antigen-specific immunoglobulin A (IgA) antibody forming cells dominated antibody responses throughout the respiratory tract. However, IgG responses were significant in lungs but not in nasal passages. Furthermore, parenteral immunization did not enhance humoral immunity in the upper respiratory tract even after a nasal challenge, whereas extrapulmonary lymphoid responses enhanced responses in the lung. After nasal immunization, inflammatory reactions, characterized by mononuclear cell infiltration, developed within the lungs of mice but not in nasal passages. Lowering dosages of CT reduced, but did not eliminate, these adverse reactions without compromising adjuvancy. Serum IgE responses were also enhanced in a dose-dependent manner by inclusion of CT. In summary, there are differences in the generation of humoral immunity between the upper respiratory tract and the lung. As the upper respiratory tract is in a separate compartment of the immune system from that stimulated by parenteral immunization, nasal immunization is an optimal approach to generate immunity throughout the respiratory tract. Despite the promise of nasal immunization, there is also the potential to develop adverse immunopathologic reactions characterized by pulmonary airway inflammation and IgE production. PMID- 11254591 TI - Immunochemical and biological characterization of three capsular polysaccharides from a single Bacteroides fragilis strain. AB - Although Bacteroides fragilis accounts for only 0.5% of the normal human colonic flora, it is the anaerobic species most frequently isolated from intra-abdominal and other infections with an intestinal source. The capsular polysaccharides of B. fragilis are part of a complex of surface polysaccharides and are the organism's most important virulence factors in the formation of intra-abdominal abscesses. Two capsular polysaccharides from strain NCTC 9343, PS A1 and PS B1, have been characterized structurally. Their most striking feature is a zwitterionic charge motif consisting of both positively and negatively charged substituent groups on each repeating unit. This zwitterionic motif is essential for abscess formation. In this study, we sought to elucidate structural features of the capsular polysaccharide complex of a commonly studied B. fragilis strain, 638R, that is distinct from strain 9343. We sought a more general picture of the species to establish basic structure-activity and structure-biosynthesis relationships among abscess-inducing polysaccharides. Strain 638R was found to have a capsular polysaccharide complex from which three distinct carbohydrates could be isolated by a complex purification procedure. Compositional and immunochemical studies demonstrated a zwitterionic charge motif common to all of the capsular polysaccharides that correlated with their ability to induce experimental intra-abdominal abscesses. Of interest is the range of net charges of the isolated polysaccharides-from positive (PS C2) to balanced (PS A2) to negative (PS 3). Relationships among structural components of the zwitterionic polysaccharides and their molecular biosynthesis loci were identified. PMID- 11254592 TI - Interleukin-10 stimulates Coxiella burnetii replication in human monocytes through tumor necrosis factor down-modulation: role in microbicidal defect of Q fever. AB - Coxiella burnetii, an obligate intracellular bacterium, is the agent of Q fever. The chronic form of the disease is associated with the overproduction of interleukin-10 and deficient C. burnetii killing by monocytes. We hypothesized that the replication of C. burnetii inside monocytes requires a macrophage deactivating cytokine such as interleukin-10. In the absence of interleukin-10, C. burnetii survived but did not replicate in monocytes. C. burnetii replication (measured 15 days) was induced in interleukin-10-treated monocytes. This effect of interleukin-10 is specific since transforming growth factor beta1 had no effect on bacterial replication. C. burnetii replication involves the down modulation of tumor necrosis factor (TNF) release. First, interleukin-10 suppressed C. burnetii-stimulated production of TNF. Second, the addition of recombinant TNF to interleukin-10-treated monocytes inhibited bacterial replication. Third, the incubation of infected monocytes with neutralizing anti TNF antibodies favored C. burnetii replication. On the other hand, deficient C. burnetii killing by monocytes from patients with chronic Q fever involves interleukin-10. Indeed, C. burnetii replication was observed in monocytes from patients with Q fever endocarditis, but not in those from patients with acute Q fever. Bacterial replication was inhibited by neutralizing anti-interleukin-10 antibodies. As monocytes from patients with endocarditis overproduced interleukin 10, the defective bacterial killing is likely related to endogenous interleukin 10. These results suggest that interleukin-10 enables monocytes to support C. burnetii replication and to favor the development of chronic Q fever. PMID- 11254594 TI - Cellular basis of early cytokine response to Plasmodium falciparum. AB - Uncertainty remains about the cellular origins of the earliest phase of the proinflammatory cytokine response to malaria. Here we show by fluorescence activated cell sorter analysis that gammadelta T cells and CD14+ cells from nonimmune donors produce tumor necrosis factor and that gammadelta T cells also produce gamma interferon within 18 h of contact with mycoplasma-free Plasmodium falciparum-infected erythrocytes in vitro. This early cytokine response is more effectively induced by intact than by lysed parasitized erythrocytes. However, the IFN-gamma response to lysed parasites is considerably enhanced several days after peripheral blood mononuclear cells are primed with low numbers of intact parasitized erythrocytes, and in this case it derives from both alphabeta and gammadelta T cells. These data show that naive gammadelta T cells can respond very rapidly to malaria infection but that malaria fever may involve a multistage process in which the priming of both gammadelta and alphabeta T-cell populations boosts the cytokine response to lysed parasite products released at schizont rupture. PMID- 11254593 TI - Involvement of HxuC outer membrane protein in utilization of hemoglobin by Haemophilus influenzae. AB - Haemophilus influenzae can utilize different protein-bound forms of heme for growth in vitro. A previous study from this laboratory indicated that nontypeable Haemophilus influenzae (NTHI) strain N182 expressed three outer membrane proteins, designated HgbA, HgbB, and HgbC, that bound hemoglobin or hemoglobin haptoglobin and were encoded by open reading frames (ORFs) that contained a CCAA nucleotide repeat. Testing of mutants expressing the HgbA, HgbB, and HgbC proteins individually revealed that expression of any one of these proteins was sufficient to allow wild-type growth with hemoglobin. In contrast, mutants that expressed only HgbA or HgbC grew significantly better with hemoglobin-haptoglobin than did a mutant expressing only HgbB. Construction of an isogenic hgbA hgbB hgbC mutant revealed that the absence of these three gene products did not affect the ability of NTHI N182 to utilize hemoglobin as a source of heme, although this mutant was severely impaired in its ability to utilize hemoglobin-haptoglobin. The introduction of a tonB mutation into this triple mutant eliminated its ability to utilize hemoglobin, indicating that the pathway for hemoglobin utilization in the absence of HgbA, HgbB, and HgbC involved a TonB-dependent process. Inactivation in this triple mutant of the hxuC gene, which encodes a predicted TonB-dependent outer membrane protein previously shown to be involved in the utilization of free heme, resulted in loss of the ability to utilize hemoglobin. The results of this study reinforce the redundant nature of the heme acquisition systems expressed by H. influenzae. PMID- 11254595 TI - Influence of major histocompatibility complex on bacterial composition of fecal flora. AB - Very little is known about how the host genome influences the composition of the gastrointestinal flora, largely due to the great number and diversity of bacteria present in the flora and the difficulties of using traditional methods of bacterial isolation and identification. We have approached the problem by studying bacterium-derived cellular fatty acids in the stool samples of six mouse strains congenic for the major histocompatibility complex (MHC). The results obtained indicate that the composition of the fecal flora is genetically regulated. In addition to undefined gene loci, MHC alone has a pronounced effect, since mice with different MHC in the same background have significantly different fecal floras. Demonstration of the genetic influence on the gastrointestinal flora opens a new approach to studying the pathogenesis of bacterially induced diseases. PMID- 11254596 TI - Evaluation of De-O-acetylated meningococcal C polysaccharide-tetanus toxoid conjugate vaccine in infancy: reactogenicity, immunogenicity, immunologic priming, and bactericidal activity against O-acetylated and De-O-acetylated serogroup C strains. AB - The polysaccharide capsule of serogroup C Neisseria meningitidis (MenC) has been integral to vaccine development. Licensed MenC vaccines contain the O-acetylated (OAc+) form of polysaccharide. Some MenC strains have de-O-acetylated (OAc-) polysaccharide, which may affect antibody specificity and functional activity when used in a vaccine. We evaluated an OAc-MenC conjugate-tetanus toxoid conjugate (MCC-TT) vaccine given concomitantly with whole-cell diphtheria-tetanus pertussis, Haemophilus influenzae type b, and oral polio immunization in 83 infants at 2, 3, and 4 months of age. Serum bactericidal activities (SBA) against OAc+ and OAc- MenC strains and OAc+ and OAc- polysaccharide-specific immunoglobulin G (IgG) levels were evaluated. MCC-TT vaccine was well tolerated. All infants produced SBA titers of > or = 8 after a single dose at 2 months of age. The SBA geometric mean titer for OAc+ strain C11 increased from 2.7 (95% confidence interval [CI] 2.2 to 3.2) to 320 (95% CI, 237 to 432), 773 (95% CI, 609 to 982), and 1,063 (95% CI, 856 to 1319) after one, two, and three doses of MCC-TT, respectively. OAc- IgG levels were twice as high as OAc+ IgG levels after the primary series of MCC-TT vaccine, and the SBA was significantly higher against the OAc- MenC strain. Antibody responses to booster vaccination with either OAc+ MenC polysaccharide vaccine (MACP) or a fourth dose of MCC-TT at 14 months of age provided evidence of immunologic memory. The acetylation status of the booster vaccine influenced the specificity of the response, with significantly higher OAc- IgG levels and SBA after MCC-TT vaccine compared to MACP vaccine but similar OAc+ antibody levels. MCC-TT vaccine is highly immunogenic and primes for immunologic memory against OAc+ and OAc- MenC strains in infancy. PMID- 11254597 TI - Expression of Chlamydia pneumoniae polymorphic membrane protein family genes. AB - The genome of the obligate intracellular bacterium Chlamydia pneumoniae CWL029 encodes a family of 21 proteins with predicted outer membrane localization. These polymorphic membrane proteins (Pmps) are heterogeneous in both amino acid sequence and predicted size but are unified by the conserved amino acid motifs GGAI and FXXN repeated in the N-terminal half of each protein. Reverse transcriptase PCR analysis showed that all pmp genes are transcribed. To determine whether all proteins are expressed, specific antisera were generated by immunization with mutually exclusive synthetic peptides representing each of the 21 predicted Pmps. Each antiserum reacted with, and was typically immunospecific for, the corresponding peptide immunogen by enzyme-linked immunosorbent assay. Western blot analyses of purified elementary bodies showed that 11 of the 21 Pmps were detectable. Attempts to demonstrate by Sarykosyl fractionation that the Pmps were localized to the outer membrane revealed that several of the Pmps were unstable and readily degraded. Analyses of additional C. pneumoniae strains showed that although some Pmps are conserved, others vary between strains, in both molecular weight and level of expression. PMID- 11254598 TI - Flow cytometric determination of Panton-Valentine leucocidin S component binding. AB - The binding of the S component (LukS-PV) from the bicomponent staphylococcal Panton-Valentine leucocidin to human polymorphonuclear neutrophils (PMNs) and monocytes was determined using flow cytometry and a single-cysteine substitution mutant of LukS-PV. The mutant was engineered by replacing a glycine at position 10 with a cysteine and was labeled with a fluorescein moiety. The biological activity of the mutant was identical to that of the native protein. It has been shown that LukS-PV has a high affinity for PMNs (Kd = 0.07 +/- 0.02 nM, n = 5) and monocytes (Kd = 0.020 +/- 0.003 nM, n = 3) with maximal binding capacities of 197,000 and 80,000 LukS-PV molecules per cell, respectively. The nonspecifically bound molecules of LukS-PV do not form pores in the presence of the F component (LukF-PV) of leucocidin. LukS-PV and HlgC share the same receptor on PMNs, but the S components of other staphylococcal leukotoxins, HlgA, LukE, and LukM, do not compete with LukS-PV for its receptor. Extracellular Ca2+ at physiological concentrations (1 to 2 nM) has only a slight influence on the LukS-PV binding, in contrast to its complete inhibition by Zn2+. The down-regulation by phorbol 12 myristate 13-acetate (PMA) of the binding of LukS-PV was blocked by staurosporine, suggesting that the regulatory effect of PMA depends on protein kinase C activation. The labeled mutant form of LukS-PV has proved very useful for detailed binding studies of circulating white cells by flow cytometry. LukS PV possesses a high specific affinity for a unique receptor on PMNs and monocytes. PMID- 11254599 TI - Profiles of immunoglobulin M (IgM) and IgG antibodies against defined carbohydrate epitopes in sera of Schistosoma-infected individuals determined by surface plasmon resonance. AB - We report here that sera of children and adults infected with Schistosoma mansoni, S. haematobium, or S. japonicum contain antibodies against GalNAcbeta1 4(Fucalpha1-2Fucalpha1-3)GlcNAc (LDN-DF) and to a lesser extent to Galbeta1 4(Fucalpha1-3)GlcNAc (Lewis(x)) and GalNAcbeta1-4GlcNAc (LDN). Surface plasmon resonance (SPR) spectroscopy was used to monitor the presence of serum antibodies to neoglycoconjugates containing these carbohydrate epitopes and to define the immunoglobulin M (IgM) and IgG subclass distribution of the antibodies. The serum levels of antibodies to LDN-DF are high related to LDN and Lewis(x) for all examined groups of Schistosoma-infected individuals. A higher antibody response to the LDN-DF epitope was found in sera of infected children than in sera of infected adults regardless of the schistosome species. With respect to the subclasses, we found surprisingly that individuals infected with S. japonicum have predominantly IgG antibodies, while individuals infected with S. mansoni mainly show an IgM response; high levels of both isotypes were measured in sera of individuals infected with S. haematobium. These data provide new insights in the human humoral immune response to schistosome-derived glycans. PMID- 11254600 TI - Involvement of CD14 and toll-like receptors in activation of human monocytes by Aspergillus fumigatus hyphae. AB - Invasive fungal infections represent an increasing problem associated with high mortality. The present study was undertaken to identify leukocyte subsets that are activated by hyphal fragments in a whole-human-blood model, as well as to examine the involvement of CD14 and Toll-like receptors (TLRs) in activation of monocytes by hyphae. Incubation of whole human blood with hyphal fragments from Aspergillus fumigatus and Scedosporium prolificans for 6 h caused induction of mRNAs for tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-6 in T cells, B cells, and monocytes, but not in granulocytes, as analyzed by reverse transcription-PCR with mRNA isolated from very pure populations of these leukocyte subsets. In primary adherent human monocytes, induction of TNF alpha by hyphal fragments was dependent on plasma. Heat treatment of plasma at 56 degrees C for 30 min strongly reduced the ability of plasma to prime for activation. Pretreatment of human monocytes with different concentrations (1, 3, and 10 microg/ml) of monoclonal antibody (MAb) HTA125 (anti-TLR4) or MAb 18D11 (anti-CD14) for 30 min inhibited the release of TNF-alpha induced by hyphal fragments in a dose-dependent manner. Maximal inhibitions of 35 and 70% were obtained with 10 microg of HTA125 and 18D11 per ml, respectively. In contrast, pretreatment with MAb TL2.1 (anti-TLR2) did not affect signaling induced by hyphae. Pretreatment with the lipid A antagonist B975 blocked lipopolysaccharide signaling but did not inhibit TNF-alpha production induced by hyphal fragments. Our results suggest that T cells, B cells, and monocytes are involved in the innate immune response to invasive fungal pathogens and that serum components are relevant for activation of monocytes by hyphae. CD14 and TLR4 may be involved in signaling of Aspergillus hyphae in monocytes, but further studies to elucidate this issue are warranted. PMID- 11254601 TI - Major histocompatibility complex class II-independent generation of neutralizing antibodies against T-cell-dependent Borrelia burgdorferi antigens presented by dendritic cells: regulation by NK and gammadelta T cells. AB - We previously showed that adoptive transfer of Borrelia burgdorferi-pulsed dendritic cells (DCs) into syngeneic mice protects animals from challenge with tick-transmitted spirochetes. Here, we demonstrate that the protective immune response is antibody (Ab) dependent and does not require the presence of major histocompatibility complex (MHC) class II molecules on DCs. Mice sensitized with B. burgdorferi-pulsed MHC class II-deficient (MHC class II(-/-)) DCs mounted a humoral response against protective antigens, including B. burgdorferi outer surface protein A (OspA) and OspC. B-cell help for the generation of neutralizing anti-OspC immunoglobulin G Abs could be provided by gammadelta T cells. In contrast, anti-OspA Ab production required the presence of alphabeta T cells, although this pathway could be independent of MHC class II molecules on antigen presenting cells. Moreover, depletion of NK cells prior to transfer of antigen pulsed MHC class II(-/-) DCs resulted in significant increases in the levels of neutralizing Abs induced by DCs. Altogether, these data suggest that the initial interactions between DCs and innate immune cells, such as gammadelta and NK cells, can influence the generation of a protective humoral response against B. burgdorferi antigens. PMID- 11254602 TI - Multilocus sequence typing of Streptococcus pyogenes and the relationships between emm type and clone. AB - Multilocus sequence typing (MLST) is a tool that can be used to study the molecular epidemiology and population genetic structure of microorganisms. A MLST scheme was developed for Streptococcus pyogenes and the nucleotide sequences of internal fragments of seven selected housekeeping loci were obtained for 212 isolates. A total of 100 unique combinations of housekeeping alleles (allelic profiles) were identified. The MLST scheme was highly concordant with several other typing methods. The emm type, corresponding to a locus that is subject to host immune selection, was determined for each isolate; of the >150 distinct emm types identified to date, 78 are represented in this report. For a given emm type, the majority of isolates shared five or more of the seven housekeeping alleles. Stable associations between emm type and MLST were documented by comparing isolates obtained decades apart and/or from different continents. For the 33 emm types for which more than one isolate was examined, only five emm types were present on widely divergent backgrounds, differing at four or more of the housekeeping loci. The findings indicate that the majority of emm types examined define clones or clonal complexes. In addition, an MLST database is made accessible to investigators who seek to characterize other isolates of this species via the internet (http://www.mlst.net). PMID- 11254603 TI - Identification of polymorphic outer membrane proteins of Chlamydia psittaci 6BC. AB - The genomes of Chlamydia spp. encode a family of putative outer membrane proteins, referred to as polymorphic outer membrane proteins (POMPs), which may play a role in the avoidance of host immune defenses. We analyzed avian strain 6BC of Chlamydia psittaci by polyacrylamide gel electrophoresis for the expression of POMPs. At least six putative POMPs were identified on the basis of their size (90 to 110 kDa) and labeling with an outer membrane-specific probe, 3 (trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine. Three of the putative POMPs reacted with antiserum raised against a recombinant ovine C. psittaci strain POMP, and two possessed surface-exposed, trypsin-sensitive sites. The POMPs were dependent on disulfide bonds for their maintenance in sodium lauryl sarcosine- and sodium dodecyl sulfate-insoluble complexes but did not appear to be interpeptide disulfide bond cross-linked. The putative POMPs were found to be synthesized during the late phase of the chlamydial developmental cycle, cotemporally with the cysteine-rich doublet periplasmic proteins. PMID- 11254604 TI - Clostridium perfringens iota-toxin: mapping of receptor binding and Ia docking domains on Ib. AB - Clostridium perfringens iota-toxin is a binary toxin consisting of iota a (Ia), an ADP-ribosyltransferase that modifies actin, and iota b (Ib), which binds to a cell surface protein and translocates Ia into a target cell. Fusion proteins of recombinant Ib and truncated variants were tested for binding to Vero cells and docking with Ia via fluorescence-activated cytometry and cytotoxicity experiments. C-terminal residues (656 to 665) of Ib were critical for cell surface binding, and truncated Ib variants containing > or = 200 amino acids of the C terminus were effective Ib competitors and prevented iota cytotoxicity. The N-terminal domain (residues 1 to 106) of Ib was important for Ia docking, yet this region was not an effective competitor of iota cytotoxicity. Further studies showed that Ib lacking just the N-terminal 27 residues did not facilitate Ia entry into a target cell and subsequent cytotoxicity. Five monoclonal antibodies against Ib were also tested with each truncated Ib variant for epitope and structural mapping by surface plasmon resonance and an enzyme-linked immunosorbent assay. Each antibody bound to a linear epitope within the N terminus (residues 28 to 66) or the C terminus (residues 632 to 655). Antibodies that target the C terminus neutralized in vitro cytotoxicity and delayed the lethal effects of iota-toxin in mice. PMID- 11254605 TI - Persistent Chlamydia trachomatis infections resist apoptotic stimuli. AB - Microbial modulation of apoptosis has added a new dimension of understanding to the dynamic interaction between the human host and its microbial invaders. Persistent infection can be a by-product of inhibition of apoptosis and may significantly impact the pathogenesis of diseases caused by organisms such as Chlamydia trachomatis. We compared apoptotic responses among HeLa 229 cells acutely and persistently infected and mock infected with serovar A/HAR-13. Persistence was induced by gamma interferon at 0.2 and 2.0 ng/ml. Cells were treated with etoposide or staurosporine at 24-h intervals and assayed for apoptosis by cell count, DNA ladder formation, and cytochrome c translocation. From the 24- to 120-h time points, infected cultures were 87 and 90% viable for etoposide and staurosporine treatment, respectively, and produced no DNA ladder, and cytochrome c remained in the mitochondria. In contrast, mock-infected cells were 22 and 37% viable for etoposide (P = 0.0001) and staurosporine (P = 0.01), respectively, and displayed characteristic DNA ladders, and cytochrome c was translocated. We found that resistance to apoptotic stimuli was identical in acute and persistent infections. Since cytochrome c was not translocated from the mitochondrion, caspase-9 activity was likely not involved. The expression of chlamydial hsp60, a known stimulator of inflammation in vivo, was measured in both active and persistent infections by Western blot, with increased production in the latter with or without staurosporine treatment. Chlamydial disregulation of apoptosis and the ensuing persistence of organisms offer an alternative pathogenic mechanism for chlamydial scarring observed in trachoma and infertility populations via sustained inflammation induced by immunoreactive molecules such as hsp60. PMID- 11254606 TI - SarS, a SarA homolog repressible by agr, is an activator of protein A synthesis in Staphylococcus aureus. AB - The expression of protein A (spa) is repressed by global regulatory loci sarA and agr. Although SarA may directly bind to the spa promoter to downregulate spa expression, the mechanism by which agr represses spa expression is not clearly understood. In searching for SarA homologs in the partially released genome, we found a SarA homolog, encoding a 250-amino-acid protein designated SarS, upstream of the spa gene. The expression of sarS was almost undetectable in parental strain RN6390 but was highly expressed in agr and sarA mutants, strains normally expressing high level of protein A. Interestingly, protein A expression was decreased in a sarS mutant as detected in an immunoblot but returned to near parental levels in a complemented sarS mutant. Transcriptional fusion studies with a 158- and a 491-bp spa promoter fragment linked to the xylE reporter gene disclosed that the transcription of the spa promoter was also downregulated in the sarS mutant compared with the parental strain. Interestingly, the enhancement in spa expression in an agr mutant returned to a near-parental level in the agr sarS double mutant but not in the sarA sarS double mutant. Correlating with this divergent finding is the observation that enhanced sarS expression in an agr mutant was repressed by the sarA locus supplied in trans but not in a sarA mutant expressing RNAIII from a plasmid. Gel shift studies also revealed the specific binding of SarS to the 158-bp spa promoter. Taken together, these data indicated that the agr locus probably mediates spa repression by suppressing the transcription of sarS, an activator of spa expression. However, the pathway by which the sarA locus downregulates spa expression is sarS independent. PMID- 11254607 TI - Stimulation of dendritic cells via CD40 enhances immune responses to Mycobacterium tuberculosis infection. AB - The resolution of pulmonary tuberculosis (TB) critically depends on the development of the Th1 type of immune responses, as exemplified by the exacerbation of TB in IL-12-deficient mice. Therefore, vaccination strategies optimizing IL-12 production by antigen-presenting cells (APC) in response to mycobacteria may have enhanced protective efficacy. Since dendritic cells (DC) are the critical APC for activation of CD4+ and CD8+ T cells, we examined whether stimulation of Mycobacterium bovis bacillus Calmette Guerin (BCG)-infected DC via CD40 increased their ability to generate Th1-oriented cellular immune responses. Incubation of DC with an agonistic anti-CD40 antibody activated CD40 signaling in DC, as shown by increased expression of major histocompatibility complex class II and costimulatory molecules, mRNA production for proinflammatory cytokines and interleukin 12 (IL-12) p40. This activation pattern was maintained when DC were stimulated with anti-CD40 antibody and infected with BCG. Importantly, CD40 stimulated BCG-infected DC displayed increased capacity to release bioactive IL 12 and to activate gamma interferon (IFN-gamma) producing T cells in vitro. Moreover, when C57BL/6 mice were immunized with these DC and challenged with aerosol Mycobacterium tuberculosis, increased levels of mRNA for IL-12 p40, IL 18, and IFN-gamma were present in the draining mediastinal lymph nodes. However, the mycobacterial burden in the lungs was not reduced compared to that in mice immunized with BCG-infected non-CD40-stimulated DC. Therefore, although the manipulation of DC via CD40 is effective for enhancing immune responses to mycobacteria in vivo, additional strategies are required to increase protection against virulent M. tuberculosis infection. PMID- 11254608 TI - Guillain-Barre syndrome- and Miller Fisher syndrome-associated Campylobacter jejuni lipopolysaccharides induce anti-GM1 and anti-GQ1b Antibodies in rabbits. AB - Campylobacter jejuni infections are thought to induce antiganglioside antibodies in patients with Guillain-Barre syndrome (GBS) and Miller Fisher syndrome (MFS) by molecular mimicry between C. jejuni lipopolysaccharides (LPS) and gangliosides. We used purified LPS fractions from five Campylobacter strains to induce antiganglioside responses in rabbits. The animals that received injections with LPS from GBS-associated strains developed anti-GM1 and anti-GA1 antibodies. Animals injected with LPS from one MFS-related C. jejuni strain produced anti GQ1b antibodies. Rabbits that were injected with Penner O:3 LPS had a strong anti LPS response, but no antiganglioside reactivity was observed. The antiganglioside specificity in the rabbits reflected the specificity in the patients from whom the strains were isolated. In conclusion, our results indicate that an immune response against GBS- and MFS-associated C. jejuni LPS results in antiganglioside antibodies. These results provide strong support for molecular mimicry as a mechanism in the induction of antiganglioside antibodies following infections. PMID- 11254610 TI - Mosaic genes and mosaic chromosomes: intra- and interspecies genomic variation of Streptococcus pneumoniae. AB - Streptococcus pneumoniae remains a major causative agent of serious human diseases. The worldwide increase of antibiotic resistant strains revealed the importance of horizontal gene transfer in this pathogen, a scenario that results in the modulation of the species-specific gene pool. We investigated genomic variation in 20 S. pneumoniae isolates representing major antibiotic-resistant clones and 10 different capsular serotypes. Variation was scored as decreased hybridization signals visualized on a high-density oligonucleotide array representing 1,968 genes of the type 4 reference strain KNR.7/87. Up to 10% of the genes appeared altered between individual isolates and the reference strain; variability within clones was below 2.1%. Ten gene clusters covering 160 kb account for half of the variable genes. Most of them are associated with transposases and are assumed to be part of a flexible gene pool within the bacterial population; other variable loci include mosaic genes encoding antibiotic resistance determinants and gene clusters related to bacteriocin production. Genomic comparison between S. pneumoniae and commensal Streptococcus mitis and Streptococcus oralis strains indicates distinct antigenic profiles and suggests a smooth transition between these species, supporting the validity of the microarray system as an epidemiological and diagnostic tool. PMID- 11254609 TI - Down-regulation of interleukin-8 secretion from Mycobacterium tuberculosis infected monocytes by interleukin-4 and -10 but not by interleukin-13. AB - Interleukin-8 (IL-8), a CXC chemokine, has a central role in leukocyte recruitment to areas of granuloma formation in tuberculosis. In the present studies, we investigated the effect of the T(H)2-derived cytokines IL-4, IL-10, and IL-13 on Mycobacterium tuberculosis-induced IL-8 secretion from purified human monocytes. Our results demonstrate that IL-4 and IL-10 have a down regulatory effect on IL-8 secretion and that this effect is dose dependent. IL-10 has a greater effect than IL-4 on secretion, and autologous IL-10 secreted from M. tuberculosis-infected monocytes also down-regulates IL-8 secretion. The down regulatory effect is partly a result of reduced IL-8 mRNA accumulation analyzed by reverse transcription-PCR. When combined, 1 microM IL-4 and IL-10 had an additive effect in decreasing IL-8 secretion and transcription; there was no synergy of action. IL-13 did not have any significant effect on IL-8 gene expression or secretion. The inhibitory effect of IL-10 but not of IL-4 is associated with decreased nuclear binding of the key activating transcription factor NF-kappaB. We show for the first time that M. tuberculosis causes up regulation of nuclear binding of Oct-1 detected by electromobility gel shift assay. However, neither AP-1 nor Oct-1 nuclear binding was altered by IL-4 or IL 10. In summary, this study demonstrates that type 2 responses have an important role in the regulation of M. tuberculosis-induced IL-8 expression but that the mechanisms by which the different cytokines act are distinct. PMID- 11254611 TI - Role of capsule in the pathogenesis of fowl cholera caused by Pasteurella multocida serogroup A. AB - We have constructed a defined acapsular mutant in Pasteurella multocida X-73 (serogroup A:1) by disrupting the hexA gene through the insertion of a tetracycline resistance cassette. The genotype of the hexA::tet(M) strain was confirmed by PCR and Southern hybridization, and the acapsular phenotype of this strain was confirmed by electron microscopy. The hexA::tet(M) strain was attenuated in both mice and chickens. Complementation of the mutant with an intact hexAB fragment restored lethality in mice but not in chickens. In contrast to the results described previously for P. multocida serogroup B (J. D. Boyce and B. Adler, Infect. Immun. 68:3463-3468, 2000), the hexA::tet(M) strain was sensitive to the bactericidal action of chicken serum, whereas the wild-type and complemented strains were both resistant. Following inoculation into chicken muscle, the bacterial count of the hexA::tet(M) strain decreased significantly, while the wild-type and complemented strains both grew rapidly over 4 h. The capsule is thus an essential virulence determinant in the pathogenesis of fowl cholera. PMID- 11254612 TI - Identification of stress-responsive genes in Streptococcus mutans by differential display reverse transcription-PCR. AB - Streptococcus mutans, which causes dental caries in the human oral cavity and occasionally causes infective endocarditis in the heart, withstands adverse environmental stress through diverse alterations in protein synthesis. Differential gene expression in response to environmental stress was analyzed by RNA fingerprinting using arbitrarily primed PCR with a panel of 11mer primers designed for differential display in Enterobacteriaceae. Dot and Northern blot hybridization confirmed that the transcription of several genes was up- or down regulated following exposure to acid shock from pH 7.5 to 5.5. RNA of a gene designated AP-185 (acid-stress protein) was induced specifically by acid treatment, while RNA of GSP-781 (general-stress protein) was up-regulated significantly when bacteria were exposed to high osmolarity and temperature, as well as low pH. The deduced amino acid sequence of AP-185 shares homology (78% identity) with branched-chain amino acid aminotransferase. Cloning and sequence analysis of GSP-781 revealed a potential secreted protein of a molecular mass of about 43 kDa and with a pI predicted to be 5.5. Transcriptional levels of another gene, designated AR-186 (acid-repressed protein), which encodes putative aconitase, were repressed by acid treatment but were enhanced by plasma or serum components. Analogous results were identified in icd and citZ genes, and repression of these genes, along with AR-186, was also observed when they were exposed to high osmolarity and temperature. These results indicate that differential regulation of specific genes at the transcriptional level is triggered by different stress and that genes responsible for glutamate biosynthesis in the citrate pathway are coordinately regulated during the stress response of S. mutans. PMID- 11254613 TI - Transcriptional regulation of divergent capsule biosynthesis and transport operon promoters in serogroup B Neisseria meningitidis. AB - The clinically important serogroups B, C, Y, and W-135 of Neisseria meningitidis produce sialic acid capsules that are critical in pathogenesis. In each of these serogroups, the capsule transport (ctrABCD) and capsule biosynthesis (synABCD) operons are divergently transcribed from putative promoters located in a 134-bp intergenic region (J. S. Swartley, J. H. Ahn, L. J. Liu, C. M. Kahler, and D. S. Stephens, J. Bacteriol. 178:4052-4059, 1996). In this study we further assessed the role of the intergenic sequence in the transcriptional regulation of the sialic acid capsules of N. meningitidis. Insertional mutagenesis or deletions of the 134-bp sequence in the serogroup B meningococcal strain NMB resulted in a marked reduction or elimination of ctrABCD and synABCD transcription, with a concomitant loss of encapsulation. Chromosomal transcriptional lacZ-ermC reporter fusions of syn and ctr promoters were constructed through allelic exchange. Using these constructs, both operons were found to be constitutively transcribed in meningococci, the biosynthesis operon about fourfold higher than the transport operon. Both promoters showed increased activity during stationary-phase growth. In addition to the promoters, a 70-bp 5' untranslated region (UTR) upstream of synA was found to have a direct repeat and an inverted repeat that overlapped three putative integration host factor binding sites. Mutation of this 70-bp UTR and of the direct repeat upregulated both syn and ctr transcription. Regulation through the synA UTR was absent in a K1 Escherichia coli strain that produces identical capsular polysaccharide, implicating species-specific regulation. Meningococcal sialic acid capsule expression is initiated by divergent promoters in a 134-bp intergenic region, is repressed at the transcriptional level by the 5' UTR of synA, is increased during stationary-phase growth, and shows species specific regulation. Transcriptional regulation is another important control point for sialic capsule expression in N. meningitidis. PMID- 11254614 TI - Streptococcus parasanguis fimbria-associated adhesin fap1 is required for biofilm formation. AB - The sanguis streptococci are primary colonizers of the tooth surface and thus form the foundation for the complex multiple species biofilm known as dental plaque. In addition, these bacteria can colonize native and prosthetic heart valves and are a common cause of endocarditis. Little is known about the molecular mechanisms governing multiple or single species biofilm development within this group of organisms. Using an in vitro assay for biofilm formation, we determined that (i) Streptococcus parasanguis FW213 can form biofilms on inert surfaces such as polystyrene and (ii) environmental and nutritional factors, such as glucose, affect S. parasanguis biofilm formation. Several isogenic mutants of FW213 were tested in the biofilm assay. Strains containing mutations in fap1, a gene encoding a protein required for assembly of fimbriae, were deficient in biofilm formation. Mutants defective in recA, PepO endopeptidase activity, or the production of a fimbriae-associated protein, FimA, were still capable of biofilm formation. Phase-contrast microscopy was used to follow biofilm development by wild-type and fap1 mutant strains on plastic coverslips over time. Wild-type FW213 attached to the surface, formed aggregates of cells, and eventually formed a dense layer of cells that included microcolonies. In contrast, few fap1 mutant cells were observed attached to the surface, and no cell aggregates or microcolonies were formed. These results suggest that the long peritrichous fimbriae of FW213 are critical for the formation of biofilms on solid surfaces. PMID- 11254615 TI - Activation of protein tyrosine kinases by Coxiella burnetii: role in actin cytoskeleton reorganization and bacterial phagocytosis. AB - Coxiella burnetii, the agent of Q fever, is an obligate intracellular microorganism that grows in monocytes/macrophages. The internalization of virulent organisms by monocytes is lower than that of avirulent variants and is associated with actin cytoskeleton reorganization. We studied the activation of protein tyrosine kinases (PTKs) by C. burnetii in THP-1 monocytes. Virulent organisms induced early PTK activation and the tyrosine phosphorylation of several endogenous substrates, including Hck and Lyn, two Src-related kinases. PTK activation reflects C. burnetii virulence since avirulent variants were unable to stimulate PTK. We also investigated the role of PTK activation in C. burnetii-stimulated F-actin reorganization. Tyrosine-phosphorylated proteins were colocalized with F-actin inside cell protrusions induced by C. burnetii, and PTK activity was increased in Triton X-100-insoluble fractions. In addition, lavendustin A, a PTK inhibitor, and PP1, a Src kinase inhibitor, prevented C. burnetii-induced cell protrusions and F-actin reorganization. We finally assessed the role of PTK activation in bacterial phagocytosis. Pretreatment of THP-1 cells with lavendustin A and PP1 upregulated the uptake of virulent C. burnetii but had no effect on the phagocytosis of avirulent organisms. Thus, it is likely that PTK activation by C. burnetii negatively regulates bacterial uptake by interfering with cytoskeleton organization. PMID- 11254616 TI - Analysis of chicken mucosal immune response to Eimeria tenella and Eimeria maxima infection by quantitative reverse transcription-PCR. AB - The recent cloning of chicken genes coding for interleukins, chemokines, and other proteins involved in immune regulation and inflammation allowed us to analyze their expression during infection with Eimeria. The expression levels of different genes in jejunal and cecal RNA extracts isolated from uninfected chickens and chickens infected with Eimeria maxima or E. tenella were measured using a precise quantitative reverse transcription-PCR technique. Seven days after E. tenella infection, expression of the proinflammatory cytokine interleukin-1beta (IL-1beta) mRNA was increased 80-fold. Among the chemokines analyzed, the CC chemokines K203 (200-fold) and macrophage inflammatory factor 1beta (MIP-1beta) (80-fold) were strongly upregulated in the infected ceca, but the CXC chemokines IL-8 and K60 were not. However, the CXC chemokines were expressed at very high levels in uninfected cecal extracts. The levels of gamma interferon (IFN-gamma) (300-fold), inducible nitric oxide synthase (iNOS) (200 fold), and myelomonocytic growth factor (MGF) (50-fold) were also highly upregulated during infection with E. tenella, whereas cyclooxygenase 2 showed a more modest (13-fold) increase. The genes upregulated during E. tenella infection were generally also upregulated during E. maxima infection but at a lower magnitude except for those encoding MIP-1beta and MGF. For these two cytokines, no significant change in expression levels was observed after E. maxima infection. CD3+ intraepithelial lymphocytes may participate in the IFN-gamma upregulation observed after infection, since both recruitment and upregulation of the IFN-gamma mRNA level were observed in the infected jejunal mucosa. Moreover, in the chicken macrophage cell line HD-11, CC chemokines, MGF, IL-1beta, and iNOS were inducible by IFN-gamma, suggesting that macrophages may be one of the cell populations involved in the upregulation of these cytokines observed in vivo during infection with Eimeria. PMID- 11254617 TI - Antibody recognition of rodent malaria parasite antigens exposed at the infected erythrocyte surface: specificity of immunity generated in hyperimmune mice. AB - In regions where malaria is endemic, inhabitants remain susceptible to repeated reinfection as they develop and maintain clinical immunity. This immunity includes responses to surface-exposed antigens on Plasmodium sp.-infected erythrocytes. Some of these parasite-encoded antigens may be diverse and phenotypically variable, and the ability to respond to this diversity and variability is an important component of acquired immunity. Characterizing the relative specificities of antibody responses during the acquisition of immunity and in hyperimmune individuals is thus an important adjunct to vaccine research. This is logistically difficult to do in the field but is relatively easily carried out in animal models. Infections in inbred mice with rodent malaria parasite Plasmodium chabaudi chabaudi AS represent a good model for Plasmodium falciparum in humans. This model has been used in the present study in a comparative analysis of cross-reactive and specific immune responses in rodent malaria. CBA/Ca mice were rendered hyperimmune to P. chabaudi chabaudi (AS or CB lines) or Plasmodium berghei (KSP-11 line) by repeated infection with homologous parasites. Serum from P. chabaudi chabaudi AS hyperimmune mice reacted with antigens released from disrupted P. chabaudi chabaudi AS-infected erythrocytes, but P. chabaudi chabaudi CB and P. berghei KSP-11 hyperimmune serum also contained cross-reactive antibodies to these antigens. However, antibody activity directed against antigens exposed at the surfaces of intact P. chabaudi chabaudi infected erythrocytes was mainly parasite species specific and, to a lesser extent, parasite line specific. Importantly, this response included opsonizing antibodies, which bound to infected erythrocytes, leading to their phagocytosis and destruction by macrophages. The results are discussed in the context of the role that antibodies to both variable and invariant antigens may play in protective immunity in the face of continuous susceptibility to reinfection. PMID- 11254618 TI - Virulence of a phosphoribosylaminoimidazole carboxylase-deficient Candida albicans strain in an immunosuppressed murine model of systemic candidiasis. AB - The relative pathogenicities of three Candida albicans strains differing in the function of ADE2 (the gene encoding phosphoribosylaminoimidazole carboxylase) were evaluated in a murine candidiasis model. C. albicans strain CAI7 (ade2/ade2), previously constructed by site-specific recombination, was avirulent in immunosuppressed mice compared to the parent strain, CAF2-1, and a heterozygous ADE2/ade2 strain obtained by transforming CAI7 with a wild-type allele. The reduced virulence of CAI7 was correlated with the inability to proliferate in either synthetic medium or serum without the exogenous addition of >10 microg of adenine/ml. The loss of virulence upon site-specific disruption of the ade2 locus, and the restoration of wild-type virulence with the repair of just one ade2 allele, confirmed that the ADE2 gene and de novo purine biosynthesis were required for Candida pathogenicity. The potential of the phosphoribosylaminoimidazole carboxylase enzyme as a novel target for antifungal drug discovery is discussed. PMID- 11254619 TI - Haemophilus ducreyi associates with phagocytes, collagen, and fibrin and remains extracellular throughout infection of human volunteers. AB - In a previous study, Haemophilus ducreyi was found in the pustule and dermis of samples obtained at the clinical end point in the human model of infection. To understand the kinetics of localization, we examined infected sites at 0, 24, and 48 h after inoculation and at the clinical end point. Immediately after inoculation, bacteria were found predominantly in the dermis but also in the epidermis. Few bacteria were detectable at 24 h; however, by 48 h, bacteria were readily seen in the pustule and dermis. H. ducreyi was associated with polymorphonuclear leukocytes and macrophages in the pustule and at its base, but was not associated with T cells, Langerhans' cells, or fibroblasts. H. ducreyi colocalized with collagen and fibrin but not laminin or fibronectin. Association with phagocytes, collagen, and fibrin was seen as early as 48 h and persisted at the pustular stage of disease. Optical sectioning by confocal microscopy and transmission electron microscopy both failed to demonstrate intracellular H. ducreyi. These data identify collagen and fibrin as potentially important targets of adherence in vivo and strongly suggest that H. ducreyi remains extracellular throughout infection and survives by resisting phagocytic killing in vivo. PMID- 11254621 TI - HtrA homologue of Legionella pneumophila: an indispensable element for intracellular infection of mammalian but not protozoan cells. AB - Legionella pneumophila replicates within alveolar macrophages, and possibly, alveolar epithelial cells and also within protozoa in the aquatic environment. Here we characterize an L. pneumophila mutant defective in the HtrA/DegP stress induced protease/chaperone homologue and show that HtrA is indispensable for intracellular replication within mammalian macrophages and alveolar epithelial cells and for intrapulmonary replication in A/J mice. Importantly, amino acid substitutions of two conserved residues in the catalytic domain of (H103mapstoR and S212mapstoA) and in-frame deletions of either or both of the two conserved PDZ domains of HtrA abolish its function. Interestingly, the htrA mutant exhibits a parental-type phenotype in intracellular replication within the protozoan host Acanthamoeba polyphaga. We used a promoterless lacZ fusion to the htrA promoter to probe the phagosomal microenvironment harboring L. pneumophila within macrophages and within A. polyphaga for the exposure to stress stimuli. The data show that expression through the htrA promoter is induced by 12,000- to 20,000 fold throughout the intracellular infection of macrophages but its induction is by 120- to 500-fold within protozoa compared to in vitro expression. Data derived from confocal laser scanning microscopy reveal that in contrast to the parental strain, phagosomes harboring the htrA mutant within U937 macrophages colocalize with the late endosomal-lysosomal marker LAMP-2, similar to killed L. pneumophila. Coinfection experiments examined by confocal laser scanning microscopy show that in communal phagosomes harboring both the parental strain and the htrA mutant, replication of the mutant is not rescued, while replication of a dotA mutant control, which is normally trafficked into a phagolysosome, is rescued by the parental strain. Our data show, for the first time, that the stress response by L. pneumophila (mediated, at least in part, by HtrA) is indispensable for intracellular replication within mammalian but not protozoan cells. PMID- 11254620 TI - Rhoptry-associated protein 1-binding monoclonal antibody raised against a heterologous peptide sequence inhibits Plasmodium falciparum growth in vitro. AB - Monoclonal antibodies (MAbs) specific for Plasmodium falciparum rhoptry associated protein 1 (RAP-1) were generated and tested for inhibition of parasite growth in vitro. The majority of indirect immunofluorescence assay (IFA)-positive MAbs raised against recombinant RAP-1 positions 23 to 711 (rRAP-1(23-711)) recognized epitopes located in the immunodominant N-terminal third of RAP-1. MAbs specific for the building block 35.1 of the synthetic peptide malaria vaccine SPf66 also yielded an IFA staining pattern characteristic for rhoptry-associated proteins and reacted specifically with rRAP-1 and parasite-derived RAP-1 molecules p67 and p82. Cross-reactivity with RAP-1 was blocked by the 35.1 peptide. Epitope mapping with truncated rRAP-1 molecules and overlapping peptides identified the linear RAP-1 sequence Y218KYSL222 as a target of the anti-35.1 MAbs. This sequence lacks primary sequence similarity with the 35.1 peptide (YGGPANKKNAG). Cross-reactivity of the anti-35.1 MAbs thus appears to be associated with conformational rather than sequence homology. While the anti-35.1 MAb SP8.18 exhibited parasite growth-inhibitory activity, none of the tested anti rRAP-1(23-711) MAbs inhibited parasite growth, independently of their fine specificity for the RAP-1 sequences at positions 33 to 42, 213 to 222, 243 to 247, 280 to 287, or 405 to 446. The growth-inhibitory activity of MAb SP8.18 was, however, accelerated by noninhibitory anti-RAP-1 MAbs. Results demonstrate that in addition to fine specificity, other binding parameters are also crucial for the inhibitory potential of an antibody. PMID- 11254622 TI - Mu-like Prophage in serogroup B Neisseria meningitidis coding for surface-exposed antigens. AB - Sequence analysis of the genome of Neisseria meningititdis serogroup B revealed the presence of an approximately 35-kb region inserted within a putative gene coding for an ABC-type transporter. The region contains 46 open reading frames, 29 of which are colinear and homologous to the genes of Escherichia coli Mu phage. Two prophages with similar organizations were also found in serogroup A meningococcus, and one was found in Haemophilus influenzae. Early and late phage functions are well preserved in this family of Mu-like prophages. Several regions of atypical nucleotide content were identified. These likely represent genes acquired by horizontal transfer. Three of the acquired genes are shown to code for surface-associated antigens, and the encoded proteins are able to induce bactericidal antibodies. PMID- 11254623 TI - Immunological studies of chronic ocular toxoplasmosis: up-regulation of major histocompatibility complex class I and transforming growth factor beta and a protective role for interleukin-6. AB - A murine model was used to characterize the local immune and inflammatory response during ocular toxoplasmosis. Major histocompatibility complex (MHC) class I, normally expressed at low levels in immune-privileged sites such as the eye, was up-regulated during infection as determined by competitive reverse transcriptase (RT)-PCR and immunocytochemistry for both beta2-microglobulin and the MHC class I heavy chain. However, the eyes of chronically infected mice also had increased levels of mRNA transcripts for transforming growth factor beta, a cytokine associated with immune privilege and constitutively expressed in normal eyes. Transcripts for a number of inflammatory mediators, including interleukin-6 (IL-6), were increased during chronic infection. The role of IL-6 was further investigated by comparing disease progression and the development of the local immune response in wild-type (WT) and IL-6-deficient mice (IL-6(-/-) mice). Following infection, IL-6(-/-) mice developed more severe inflammation in the retina and vitreous humor compared with WT mice. This increased severity of disease was associated with reduced ocular IL-1alpha and increased tumor necrosis factor alpha mRNA production compared with WT mice. Moreover, the increased severity of disease in IL-6(-/-) mice correlated with increased eye parasite burden as determined by RT-PCR for the Toxoplasma gondii bradyzoite-specific LDH2 gene. These results demonstrate alterations to components of immune privilege as a result of ocular toxoplasmosis and a role for IL-6 in controlling parasite numbers and inflammation in the eye. PMID- 11254624 TI - Mycobacterium bovis BCG-induced granuloma formation depends on gamma interferon and CD40 ligand but does not require CD28. AB - Progressive granuloma formation is a hallmark of chronic mycobacterial infection. Granulomas are localized, protective inflammatory reactions initiated by CD4+ T cells, which contribute to control of bacterial growth and blockade of bacterial dissemination. In order to understand the costimulatory requirements that allow CD4+ T cells to directly or indirectly induce granulomas, we studied granuloma formation after 6 weeks in Mycobacterium bovis BCG-infected CD28- and CD40 ligand (CD40L)-deficient mice and compared it to granuloma formation in infected wild type inbred mice and infected cytokine-deficient mice. We characterized granulomas morphologically in liver sections, analyzed granuloma infiltrating cells by flow cytometry, and measured cytokine production by cultured granuloma cells. CD28-deficient mice have no defect at the local inflammatory site, inasmuch as they form protective granulomas and control bacterial growth. However, there are fewer activated T cells in the spleen compared to infected wild-type animals, and quantitative differences in the cellular composition of the granuloma are observed by flow cytometry. In CD40L-deficient mice, the granuloma phenotype is very similar to the phenotype in gamma interferon (IFN gamma)-deficient mice. Both IFN-gamma-deficient and CD40L-deficient mice form granulomas which prevent bacterial dissemination, but control of bacterial growth is significantly impaired. The relative proportion of CD4+ T cells in granulomas from both CD28(-/-) and CD40L(-/-) mice is significantly decreased compared with wild-type animals. Both models demonstrate that the phenotype and activation stage of systemic T cells do not always correlate with the phenotype and activation stage of the localized granulomatous response. PMID- 11254625 TI - Helicobacter pylori resists phagocytosis by macrophages: quantitative assessment by confocal microscopy and fluorescence-activated cell sorting. AB - Helicobacter pylori infection of the stomach epithelium is characterized by an infiltration of polymorphonuclear and mononuclear cells. These immune cells contribute to mucosal damage which may eventually lead to gastritis, peptic ulcer, gastric cancer, and/or MALT-associated gastric lymphoma. Here we show that H. pylori inhibits its own uptake, as well as in trans the phagocytosis of Neisseria gonorrhoeae, by human and murine macrophages. This antiphagocytic activity is dependent on the presence of the cag pathogenicity island in the H. pylori genome. We demonstrate that H. pylori also expresses its antiphagocytic activity towards the myelomonocytic cell line JOSKM, thus providing a potent model for the study of the interaction between H. pylori and phagocytes. Our data were obtained using laser confocal microscopy and flow cytometry after quenching the fluorescence of labeled extracellular bacteria. The antiphagocytic activity of H. pylori may explain the persistence of H. pylori and its pathological consequences. The use of cell lines and flow cytometry will hopefully facilitate progress in our understanding of the immune escape of these persistent bacteria. PMID- 11254627 TI - Gamma interferon prevents the inhibitory effects of oxidative stress on host responses to Escherichia coli infection. AB - Oxidative stress occurs in animals challenged with bacterial endotoxin and can affect the expression of important host inflammatory genes. However, much less is known about the effects of oxidative stress on responses to gram-negative bacteria. The current study compared the effects of redox imbalance on hepatic responses of mice to Escherichia coli bacteria versus purified endotoxic lipopolysaccharide (LPS). Oxidative stress induced by glutathione depletion virtually eliminated hepatic tumor necrosis factor alpha responses to both E. coli and LPS. Inducible NO synthase (iNOS) and intercellular adhesion molecule-1 (ICAM-1) expression was also markedly inhibited by glutathione depletion in LPS challenged mice, but was unaffected in E. coli-infected animals. Three findings suggested that gamma interferon (IFN-gamma) production explained the differences between LPS and bacterial challenge. Glutathione depletion completely inhibited the IFN-gamma response to LPS, but only partially inhibited IFN-gamma production in infected mice. Exogenous IFN-gamma restored iNOS and ICAM-1 responses to LPS in stressed mice. Conversely, IFN-gamma-deficient, glutathione-depleted mice showed a marked decrease in iNOS and ICAM-1 expression when challenged with E. coli. These findings indicate that both the nature of the microbial challenge and the production of IFN-gamma can be important in determining the effects of redox imbalance during gram-negative bacterial infections. PMID- 11254626 TI - Complete DNA sequence and comparative analysis of the 50-kilobase virulence plasmid of Salmonella enterica serovar Choleraesuis. AB - The complete nucleotide sequence of pKDSC50, a large virulence plasmid from Salmonella enterica serovar Choleraesuis strain RF-1, has been determined. We identified 48 of the open reading frames (ORFs) encoded by the 49,503-bp molecule. pKDSC50 encodes a known virulence-associated operon, the spv operon, which is composed of genes essential for systemic infection by nontyphoidal Salmonella. Analysis of the genetic organization of pKDSC50 suggests that the plasmid is composed of several virulence-associated genes, which include the spvRABCD genes, plasmid replication and maintenance genes, and one insertion sequence element. A second virulence-associated region including the pef (plasmid encoded fimbria) operon and rck (resistance to complement killing) gene, which has been identified on the virulence plasmid of S. enterica serovar Typhimurium, was absent. Two different replicon regions, similar to the RepFIIA and RepFIB replicons, were found. Both showed high similarity to those of the pO157 plasmid of enterohemorrhagic Escherichia coli O157:H7 and the enteropathogenic E. coli (EPEC) adherence factor plasmid harbored by EPEC strain B171 (O111:NM), as well as the virulence plasmids of Salmonella serovars Typhimurium and Enteritidis. Comparative analysis of the nucleotide sequences of the 50-kb virulence plasmid of serovar Choleraesuis and the 94-kb virulence plasmid of serovar Typhimurium revealed that 47 out of 48 ORFs of the virulence plasmid of serovar Choleraesuis are highly homologous to the corresponding ORFs of the virulence plasmid of serovar Typhimurium, suggesting a common ancestry. PMID- 11254628 TI - Subcytocidal attack by staphylococcal alpha-toxin activates NF-kappaB and induces interleukin-8 production. AB - Formation of transmembrane pores by staphylococcal alpha-toxin can provoke a spectrum of events depending on target cell species and toxin dose, and in certain cases, repair of the lesions has been observed. Here, we report that transcriptional processes are activated as a response of cells to low toxin doses. Exposure of monocytic (THP-1) or epithelial (ECV304) cells to 40 to 160 ng/ml alpha-toxin provoked a drop in cellular ATP level that was followed by secretion of substantial amounts of interleukin-8 (IL-8). Cells transfected with constructs comprising the proximal IL-8 promoter fused to luciferase or to green fluorescent protein cDNA exhibited enhanced reporter gene expression following toxin treatment. Electrophoretic mobility shift and immunofluorescence assays demonstrated that IL-8 secretion was preceded by activation of NF-kappaB. Transfection experiments conducted with p65/p50 double-deficient cells showed that activation of the IL-8 promoter/reporter by toxin was absolutely dependent on NF-kappaB. In contrast, this transcription factor was not required for lesion repair. Attack of cells by low doses of a pore-forming toxin can lead to transcriptional gene activation, which is followed by production of mediators that may contribute to the initiation and propagation of inflammatory lesions. PMID- 11254629 TI - CD8+ T cells have an essential role in pulmonary clearance of nontypeable Haemophilus influenzae following mucosal immunization. AB - A rodent respiratory experimental model has proved useful for investigating the immune mechanisms responsible for clearance of bacteria from the lungs. Immunohistochemical studies in immune and nonimmune rats have identified the cellular kinetics of response to bacterial pulmonary infection for CD8+, CD4+, and gammadelta+ T cells; B cells; and the expression of major histocompatibility complex class II (MHC-II). During the course of bacterial clearance, there was no apparent proliferation or extravasation of lymphocytes, nor was there increased expression of MHC-II in nonimmune animals despite an influx of polymorphonuclear leukocytes, whereas in immunized animals there was an early influx of CD8+ and gammadelta+ T cells, followed by enhanced expression of the MHC-II marker, cellular infiltration by polymorphonuclear leukocytes, and finally an increased number of CD4+ T cells. Depletion of CD8+ T cells confirmed their vital contribution in the preprimed immune response to pulmonary infection by significantly decreasing the animals' ability to clear bacteria following challenge. PMID- 11254630 TI - Resolution of secondary Chlamydia trachomatis genital tract infection in immune mice with depletion of both CD4+ and CD8+ T cells. AB - The essential role of T cells in the resolution of primary murine Chlamydia trachomatis genital tract infection is inarguable; however, much less is known about the mechanisms that confer resistance to reinfection. We previously established that CD4+ T cells and B cells contribute importantly to resistance to reinfection. In our current studies, we demonstrate that immune mice concurrently depleted of both CD4+ T cells and CD8+ T cells resisted reinfection as well as immunocompetent wild-type mice. The in vivo depletion of CD4+ and CD8+ T cells resulted in diminished chlamydia-specific delayed-type hypersensitivity responses, but antichlamydial antibody responses were unaffected. Our data indicate that immunity to chlamydial genital tract reinfection does not rely solely upon immune CD4+ or CD8+ T cells and further substantiate a predominant role for additional effector immune responses, such as B cells, in resistance to chlamydial genital tract reinfection. PMID- 11254631 TI - Proinflammatory and proapoptotic activities associated with Bordetella pertussis filamentous hemagglutinin. AB - Filamentous hemagglutinin (FHA) is a dominant cell surface-associated Bordetella pertussis adhesin. Recognition that this protein is secreted in significant amounts and that bacterial adhesins may have other activities, prompted an assessment of FHA effects on human macrophages. Incubation of human macrophage like U937 cells with preparations of FHA resulted in dose-dependent cytotoxicity, with death of 95% of treated cells after 24 h. Based on the use of four independent methods, death of these cells could be largely attributed to apoptosis. FHA-associated apoptosis was also observed in THP-1 macrophage-like cells, fresh human peripheral blood monocyte-derived macrophages (MDM), and BEAS 2B human bronchial epithelial cells. Infection of MDM with wild-type B. pertussis resulted in apoptosis within 6 h, while infection with an FHA-deficient derivative strain was only 50% as effective. FHA-associated cytotoxicity was preceded by host cell secretion of tumor necrosis factor alpha (TNF-alpha), a potential proapoptotic factor. However, pretreatment of cells with a neutralizing anti-TNF-alpha monoclonal antibody inhibited only 16% of the FHA-associated apoptosis. On the other hand, a blocking monoclonal antibody directed against TNF alpha receptor 1 inhibited FHA-associated apoptosis by 47.7% (P = 0.0001), suggesting that this receptor may play a role in the death pathway activated by FHA. Our in vitro data indicate that secreted and cell-associated FHA elicits proinflammatory and proapoptotic responses in human monocyte-like cells, MDM, and bronchial epithelial cells and suggest a previously unrecognized role for this prominent virulence factor in the B. pertussis-host interaction. PMID- 11254633 TI - Gamma interferon-producing CD4+ T lymphocytes in the lung correlate with resistance to infection with Mycobacterium tuberculosis. AB - The human immune system efficiently limits the replication of Mycobacterium tuberculosis in most infected individuals. Only 5 to 10% of infected people develop clinical tuberculosis, a sign of the inability of the immune system to control the infection. We have studied the C3H/HeJ (C3H) and C57BL/6 (B6) inbred mouse strains, which differ in their susceptibility to tuberculosis, in order to ascertain the immunological determinants of a successful immune response against M. tuberculosis and to establish a system to identify genes that influence susceptibility to tuberculosis. We found that the resistant B6 mice were able to control infection in both the lung and spleen, while susceptible C3H mice were incapable of limiting bacteria growth, especially in the lung, and succumbed to infection within 4 weeks. We determined that the susceptibility of C3H mice was independent of the Toll-like receptor 4 (tlr4) genetic locus and allelic major histocompatibility complex differences. Although the splenic immune responses were similar in the two mouse strains, the local immune responses in the lungs of the infected mice differed greatly. The pulmonary immune response in resistant B6 mice was characterized by an early influx of both CD4+ and CD8+ lymphocytes that produced gamma interferon (IFN-gamma). In contrast, the immune response of C3H mice in the lung was characterized by a delayed and decreased influx of lymphocytes, which produced little IFN-gamma. These results suggest an important role for the early appearance of IFN-gamma-producing lymphocytes in the lung in resistance to infection with M. tuberculosis. PMID- 11254632 TI - Curli fibers mediate internalization of Escherichia coli by eukaryotic cells. AB - Curli fibers are adhesive surface fibers expressed by Escherichia coli and Salmonella enterica that bind several host extracellular matrix and contact phase proteins and were assumed to have a role in pathogenesis. The results presented here suggest that one such role is internalization into host cells. An E. coli K 12 strain transformed with a low-copy vector containing the gene cluster encoding curli fibers (csg operon) was internalized by several lines of eukaryotic cells. The internalization could be correlated with a high level of curli fiber expression and was abolished by disruption of the csg operon. The ability to be internalized by eukaryotic cells could be conferred even by the curli fiber gene cluster of a noninvasive K-12 strain, but the homologous csg cluster from a virulent septicemic E. coli isolate mediated a higher level of internalization. The finding that curli fibers promote bacterial internalization indicates a new role for curli fibers in pathogenesis. PMID- 11254634 TI - Pseudomonas aeruginosa-induced apoptosis involves mitochondria and stress activated protein kinases. AB - Pseudomonas aeruginosa, a gram-negative facultative pathogen, causes severe infections in immunocompromised and cystic fibrosis patients. However, the molecular details of the interaction between P. aeruginosa and mammalian cells are still largely unknown. Here we demonstrate that infection of human conjunctiva epithelial Chang cells with the well-characterized P. aeruginosa strain PAO-I results in rapid induction of apoptosis. Apoptosis was mediated by mitochondrial alterations, in particular mitochondrial depolarization, synthesis of reactive oxygen intermediates, and release of cytochrome c, as well as an activation of Jun N-terminal kinases (JNK). Stimulation of these events was dependent on upregulation of CD95 on infected cells, and a deficiency of CD95 or the CD95 ligand prevented mitochondrial changes, JNK activation, and apoptosis upon infection. Further, efficient apoptosis of Chang epithelial cells required infection with live P. aeruginosa, adhesion but not invasion of the bacteria, and expression of the type III secretion system in PAO-I. The data indicate a type III secretion system-dependent, sequential activation of several signaling pathways by P. aeruginosa PAO-I, resulting in apoptosis of the infected cell. PMID- 11254635 TI - Gallinacin-3, an inducible epithelial beta-defensin in the chicken. AB - Gallinacin-3 and gallopavin-1 (GPV-1) are newly characterized, epithelial beta defensins of the chicken (Gallus gallus) and turkey (Meleagris gallopavo), respectively. In normal chickens, the expression of gallinacin-3 was especially prominent in the tongue, bursa of Fabricius, and trachea. It also occurred in other organs, including the skin, esophagus, air sacs, large intestine, and kidney. Tracheal expression of gallinacin-3 increased significantly after experimental infection of chickens with Haemophilus paragallinarum, whereas its expression in the tongue, esophagus, and bursa of Fabricius was unaffected. The precursor of gallinacin-3 contained a long C-terminal extension not present in the prepropeptide. By comparing the cDNA sequences of gallinacin-3 and GPV-1, we concluded that a 2-nucleotide insertion into the gallinacin-3 gene had induced a frameshift that read through the original stop codon and allowed the chicken propeptide to lengthen. The striking structural resemblance of the precursors of beta-defensins to those of crotamines (highly toxic peptides found in rattlesnake venom) supports their homology, even though defensins are specialized to kill microorganisms and crotamines are specialized to kill much larger prey. PMID- 11254636 TI - Antimycobacterial agent based on mRNA encoding human beta-defensin 2 enables primary macrophages to restrict growth of Mycobacterium tuberculosis. AB - Human macrophages are hosts for Mycobacterium tuberculosis, the causative agent of tuberculosis, which killed approximately 1.87 million people in 1997. Human alveolar macrophages do not express alpha- or beta-defensins, broad-spectrum antimicrobial peptides which are expressed in macrophages from other species more resistant to infection with M. tuberculosis. It has been previously reported that M. tuberculosis is susceptible to killing by defensins, which may explain the difference in resistance. Defensin peptides have been suggested as a possible therapeutic strategy for a variety of infectious diseases, but development has been hampered by difficulties in their large-scale production. Here we report the cellular synthesis of human beta-defensin 2 via highly efficient mRNA transfection of human macrophages. This enabled mycobactericidal and mycobacteristatic activity by the macrophages. Although human macrophages are difficult to transfect with plasmid vectors, these studies illustrate that primary macrophages are permissive for mRNA transfection, which enabled expression of a potentially therapeutic protein. PMID- 11254637 TI - Intracellular Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in buccal epithelial cells collected from human subjects. AB - The mouth may provide an accessible model for studying bacterial interactions with human cells in vivo. Using fluorescent in situ hybridization and laser scanning confocal microscopy, we found that human buccal epithelial cells from 23 of 24 subjects were infected with intracellular bacteria, including the periodontal pathogens Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis, as well as other species which have yet to be identified. Buccal cell invasion may allow fastidious anaerobes to establish themselves in aerobic sites that otherwise present an unfavorable environment. Exfoliated buccal epithelial cells might provide a protected route for bacterial transmission between different oral sites within and between hosts. PMID- 11254638 TI - Antigastric autoantibodies in ferrets naturally infected with Helicobacter mustelae. AB - Infection with Helicobacter pylori has been associated with induction of autoantibodies that cross-react with the gastric mucosa. There have been discordant reports as to whether or not these autoantibodies arise due to molecular mimicry between H. pylori and host cell antigens on parietal cells. In this study, we investigated whether molecular mimicry by H. mustelae causes autoantibodies in infected ferrets. Serum from H. mustelae-infected ferrets reacted with parietal cells in the ferret gastric mucosa but not with duodenal or colonic mucosa. These sera did not react with the blood group A epitope on erythrocytes or H. mustelae lipopolysaccharide, and absorption with H. mustelae whole cells or red blood cells did not remove autoantibodies. In conclusion, ferrets naturally infected with H. mustelae generate antibodies that react with parietal cells, but these autoantibodies are not due to molecular mimicry. PMID- 11254639 TI - Boosting vaccine for tuberculosis. AB - An effective new vaccine for the control of tuberculosis is badly needed. While current research attempts to improve vaccination are concentrating on new prophylactic or immunotherapeutic vaccines, virtually no emphasis has been placed on boosting individuals already inoculated with Mycobacterium bovis BCG. It is shown here that mice vaccinated with BCG gradually lose their capacity to resist an aerosol challenge infection as they age. However, if these mice are inoculated with the 30-kDa mycolyl transferase A protein in midlife, after challenge when aged they express levels of protection in the lungs comparable to those of young mice, associated with minimal pathological damage. PMID- 11254640 TI - Differential gene expression during meningeal-meningococcal interaction: evidence for self-defense and early release of cytokines and chemokines. AB - Using microarray technology, we studied the early differential expression of 3,528 genes in human meningothelial cells in response to meningococcal challenge. Thirty-two genes were up-regulated, and four were down-regulated. Those up regulated included the tumor necrosis factor alpha, interleukin-6 (IL-6), and IL 8 (but not IL-1beta) genes, suggesting that meningeal cells may be a local and early source of these cytokines. Also, a trend in up-regulation of anti-apoptotic genes and down-regulation of pro-apoptotic genes was observed. This is the first evidence that meningothelial cells may mount cytoprotective responses to pathogenic bacteria. PMID- 11254641 TI - Intracellular crystal formation as a mechanism of cytotoxicity in murine pulmonary Cryptococcus neoformans infection. AB - Rod-like crystalline structures formed during eosinophilic Cryptococcus neoformans pneumonia in C57BL/6 mice. Crystals were found associated with yeast cells and free in host cell cytoplasm. The crystals apparently formed because of the interaction of a host protein with the cryptococcal polysaccharide. Crystal formation likely contributes to pathogenesis by causing cellular damage. PMID- 11254642 TI - Protective role of lung surfactant protein D in a murine model of invasive pulmonary aspergillosis. AB - The protective effects of intranasal administration of amphotericin B (AmB), human SP-A, SP-D and a 60-kDa fragment of SP-D (rSP-D) were examined in a murine model of invasive pulmonary aspergillosis (IPA). The untreated group of IPA mice showed no survival at 7 days postinfection. Treatment with AmB, SP-D, and rSP-D increased the survival rate to 80, 60, and 80%, respectively, suggesting that SP D (and rSP-D) can protect immunosuppressed mice from an otherwise fatal challenge with Aspergillus fumigatus conidia. PMID- 11254643 TI - Generation and characterization of a defined mutant of Streptococcus suis lacking suilysin. AB - A defined allelic-replacement mutant of the sly gene, encoding a thiol-activated cytolysin, from a European isolate of Streptococcus suis serotype 2 was generated and characterized. Unlike the parental strain, it is nonhemolytic, noncytotoxic for cultured macrophage-like cells, avirulent in a mouse infection model, yet only slightly attenuated in a porcine model of systemic infection. PMID- 11254644 TI - Induction of cell-associated chemokines after endotoxin administration to healthy humans. AB - Erythrocytes express the Duffy antigen receptor for chemokines. Endotoxin injection into humans induced high levels of interleukin-8 (IL-8), growth-related oncogene alpha, and monocyte chemoattractant protein 1 in circulating erythrocytes. IL-8 was also recovered from mononuclear and polymorphonuclear cells. Cell-associated chemokines may more accurately reflect their production than plasma concentrations. PMID- 11254645 TI - Borrelia burgdorferi proteins whose expression is similarly affected by culture temperature and pH. AB - Previously, we had demonstrated the upregulation in the expression of several proteins, including the lipoproteins OspC and P35, of Borrelia burgdorferi in the stationary growth phase. Since the expression of OspC is also known to be affected by culture temperature and pH, we examined the effects of both variables on the expression of the remaining stationary-phase-upregulated proteins. Our study revealed that the expression of each of the remaining stationary-phase upregulated proteins, P35 included, was also influenced by culture temperature; these proteins were selectively expressed at 34 degrees C but not at 24 degrees C. Significantly, the expression of a majority of these proteins was also affected by culture pH, since they were abundantly expressed at pH 7.0 (resembling the tick midgut pH of 6.8 during feeding) but only sparsely at pH 8.0 (a condition closer to that of the unfed tick midgut pH of 7.4). We propose that this group of B. burgdorferi proteins, which in culture is selectively expressed under conditions of 34 degrees C and pH 7.0, may be induced in the tick midgut during the feeding event. Furthermore, the differential and coordinate expression of these proteins under different environmental conditions suggests that the encoding genes may be coregulated. PMID- 11254646 TI - Evidence of involvement of the mannose receptor in adhesion of Borrelia burgdorferi to monocyte/macrophages. AB - The mannose receptor (MR) plays an important role in the recognition of some pathogens in nonopsonic phagocytosis and in antigen presentation to T cells. We found that Borrelia burgdorferi, the agent of Lyme borreliosis, adheres to monocyte-derived macrophages and to rat MR-transfected cells but not to untransfected cells. Antibodies to MR and sugars such as mannose, mannan, fucose, and some lectins significantly lowered the adhesion, confirming participation of the MR in the binding. PMID- 11254647 TI - Identification of the exported proteins of the oral opportunistic pathogen Actinobacillus actinomycetemcomitans by using alkaline phosphatase fusions. AB - A phoA fusion library of Actinobacillus actinomycetemcomitans genomic DNA has been screened to identify genes encoding exported and secreted proteins. A total of 8,000 colonies were screened, and 80 positive colonies were detected. From these, 48 genes were identified with (i) more than half having homology to known or hypothetical Haemophilus influenzae genes, (ii) 14 having no ascribed function, and (iii) 4 having very limited or no homology to known genes. The proteins encoded by these genes may, by virtue of their presence on the cell surface, be novel virulence determinants. PMID- 11254648 TI - Secretion of listeriolysin by Brucella suis inhibits its intramacrophagic replication. AB - The introduction into Brucella suis 1330 of a plasmid allowing the heterologous expression of a hybrid cytolysin containing listeriolysin from Listeria monocytogenes, and its export via the Escherichia coli hemolysin secretion pathway, resulted in secretion of active listeriolysin monitored by erythrocyte lysis. In contrast to observations with the nonhemolytic control strain, the phagosomes of infected human monocytes containing the hemolytic B. suis were partially disrupted, and this strain failed to multiply in human macrophage-like cells. These results added strong evidence supporting the proposal that the phagosome of the macrophage was the predominant niche of brucellae in their mammalian hosts. PMID- 11254649 TI - Trypanosome-derived oligopeptidase B is released into the plasma of infected rodents, where it persists and retains full catalytic activity. AB - A trypsin-like serine peptidase activity, levels of which correlate with blood parasitemia levels, is present in the plasma of rats acutely infected with Trypanosoma brucei brucei. Antibodies to a trypanosome peptidase with a trypsin like substrate specificity (oligopeptidase B [OP-Tb]) cross-reacted with a protein in the plasma of trypanosome-infected rats on a Western blot. These antibodies also abolished 80% of the activity in the plasma of trypanosome infected rats, suggesting that the activity may be attributable to a parasite derived peptidase. We purified the enzyme responsible for the bulk of this activity from parasite-free T. b. brucei-infected rat plasma and confirmed its identity by protein sequencing. We show that live trypanosomes do not release OP Tb in vitro and propose that disrupted parasites release it into the host circulation, where it is unregulated and retains full catalytic activity and may thus play a role in the pathogenesis of African trypanosomiasis. PMID- 11254650 TI - Saccharomyces boulardii stimulates intestinal immunoglobulin A immune response to Clostridium difficile toxin A in mice. AB - Saccharomyces boulardii is a nonpathogenic yeast that protects against antibiotic associated diarrhea and recurrent Clostridium difficile colitis. The administration of C. difficile toxoid A by gavage to S. boulardii-fed BALB/c mice caused a 1.8-fold increase in total small intestinal immunoglobulin A levels (P = 0.003) and a 4.4-fold increase in specific intestinal anti-toxin A levels (P < 0.001). Enhancing host intestinal immune responses may be an important mechanism for S. boulardii-mediated protection against diarrheal illnesses. PMID- 11254651 TI - HLA-A*01-restricted cytotoxic T-lymphocyte epitope from the Plasmodium falciparum circumsporozoite protein. AB - Here, we report the identification of a novel CD8+ cytotoxic T-lymphocyte epitope on the Plasmodium falciparum circumsporozoite protein (3D7; amino acids 310 to 319 [EPSDKHIKEY]) that is restricted by HLA-A*01 and is recognized by human volunteers immunized with irradiated P. falciparum sporozoites. HLA-A*01 is the second most common HLA allele among Caucasians. PMID- 11254652 TI - Oligosaccharide-based information in endoplasmic reticulum quality control and other biological systems. PMID- 11254653 TI - Phospholipase C-gamma1 is required for the induction of immediate early genes by platelet-derived growth factor. AB - To explore the functional role of phospholipase C-gamma1 (PLC-gamma1) in the induction of immediate early genes (IEGs), we have examined the influence of Plcg1 gene disruption on the expression of 14 IEG mRNAs induced by platelet derived growth factor (PDGF). Plcg1-null embryos were used to produce immortalized fibroblasts genetically deficient in PLC-gamma1 (Null cells), and retroviral infection of those cells was used to derive PLC-gamma1 re-expressing cells (Null+ cells). In terms of PDGF activation of PDGF receptor tyrosine phosphorylation as well as the mitogen-activated protein kinases Erk1 and Erk2, Null and Null+ cells responded equivalently. However, the PDGF-dependent expression of all IEG mRNAs was diminished in cells lacking PLC-gamma1. The expression of FIC, COX-2, KC, JE, and c-fos mRNAs were most strongly compromised, as the stimulation of these genes was reduced by more than 90% in cells lacking PLC-gamma1. The combination of PMA and ionomycin, downstream analogs of PLC activation, did provoke expression of mRNAs for these IEGs in the Null cells. We conclude that PLC-gamma1 is necessary for the maximal expression of many PDGF induced IEGs and is essential for significant induction of at least five IEGs. PMID- 11254654 TI - BMK1 mediates growth factor-induced cell proliferation through direct cellular activation of serum and glucocorticoid-inducible kinase. AB - Activation of the mammalian mitogen-activated protein kinase known as BMK1 is required for growth factor-induced cell proliferation. To understand the mechanism by which BMK1 mediates this cellular response, this kinase was used as bait in a yeast two-hybrid-based library screening. Here, we report the identification of serum and glucocorticoid-inducible kinase (SGK) as a cellular protein that physically interacts with BMK1. During growth factor-induced cell stimulation, BMK1 activates SGK by phosphorylation at serine 78. This BMK1 mediated phosphorylation event is necessary for the activation of SGK and, more importantly, for cell proliferation induced by growth factors. PMID- 11254655 TI - Mnl1p, an alpha -mannosidase-like protein in yeast Saccharomyces cerevisiae, is required for endoplasmic reticulum-associated degradation of glycoproteins. AB - The endoplasmic reticulum (ER) has a mechanism to block the exit of misfolded or unassembled proteins from the ER for the downstream organelles in the secretory pathway. Misfolded proteins retained in the ER are subjected to proteasome dependent degradation in the cytosol when they cannot achieve correct folding and/or assembly within an appropriate time window. Although specific mannose trimming of the protein-bound oligosaccharide is essential for the degradation of misfolded glycoproteins, the precise mechanism for this recognition remains obscure. Here we report a new alpha-mannosidase-like protein, Mnl1p (mannosidase like protein), in the yeast ER. Mnl1p is unlikely to exhibit alpha1,2-mannosidase activity, because it lacks cysteine residues that are essential for alpha1,2 mannosidase. However deletion of the MNL1 gene causes retardation of the degradation of misfolded carboxypeptidase Y, but not of the unglycosylated mutant form of the yeast alpha-mating pheromone. Possible roles of Mnl1p in the degradation and in the ER-retention of misfolded glycoproteins are discussed. PMID- 11254656 TI - Multiple N-CoR complexes contain distinct histone deacetylases. AB - N-CoR (nuclear receptor corepressor) is a corepressor for multiple transcription factors including unliganded thyroid hormone receptors (TRs). In vitro, N-CoR can interact with the Sin3 corepressor, which in turn binds to the histone deacetylase Rpd3 (HDAC1), predicting the existence of a corepressor complex containing N-CoR, Sin3, and histone deacetylase. However, previous biochemical studies of endogenous Sin3 complexes have failed to find an N-CoR association. Xenopus laevis eggs and oocytes contain all of the necessary components for transcriptional repression by unliganded TRs. In this study, we report the biochemical fractionation of three novel macromolecular complexes containing N CoR, two of which possess histone deacetylase activity, from Xenopus egg extract. One complex contains Sin3, Rpd3, and RbAp48; the second complex contains a Sin3 independent histone deacetylase; and the third complex lacks histone deacetylase activity. This study describes the first biochemical isolation of endogenous N CoR-containing HDAC complexes and illustrates that N-CoR associates with distinct histone deacetylases that are both dependent and independent of Sin3. Immunoprecipitation studies show that N-CoR binds to unliganded TR expressed in the frog oocyte, confirming that N-CoR complexes are involved in repression by unliganded TR. These results suggest that N-CoR targets transcriptional repression of specific promoters through at least two distinct histone deacetylase pathways. PMID- 11254657 TI - Control of retinoic acid receptor heterodimerization by ligand-induced structural transitions. A novel mechanism of action for retinoid antagonists. AB - Heterodimerization of retinoic acid receptors (RARs) with 9-cis-retinoic receptors (RXRs) is a prerequisite for binding of RXR.RAR dimers to DNA and for retinoic acid-induced gene regulation. Whether retinoids control RXR/RAR solution interaction remains a debated question, and we have used in vitro and in vivo protein interaction assays to investigate the role of ligand in modulating RXR/RAR interaction in the absence of DNA. Two-hybrid assay in mammalian cells demonstrated that only RAR agonists were able to increase significantly RAR interaction with RXR, whereas RAR antagonists inhibited RXR binding to RAR. Quantitative glutathione S-transferase pull-down assays established that there was a strict correlation between agonist binding affinity for the RAR monomer and the affinity of RXR for liganded RAR, but RAR antagonists were inactive in inducing RXR recruitment to RAR in vitro. Alteration of coactivator- or corepressor-binding interfaces of RXR or RAR did not alter ligand-enhanced dimerization. In contrast, preventing the formation of a stable holoreceptor structure upon agonist binding strongly altered RXR.RAR dimerization. Finally, we observed that RAR interaction with RXR silenced RXR ligand-dependent activation function. We propose that ligand-controlled dimerization of RAR with RXR is an important step in the RXR.RAR activation process. This interaction is dependent upon adequate remodeling of the AF-2 structure and amenable to pharmacological inhibition by structurally modified retinoids. PMID- 11254661 TI - Borrelia burgdorferi and Treponema pallidum: a comparison of functional genomics, environmental adaptations, and pathogenic mechanisms. PMID- 11254663 TI - Homocysteine: a sulph'rous fire. PMID- 11254662 TI - The simple sequence contingency loci of Haemophilus influenzae and Neisseria meningitidis. PMID- 11254664 TI - Pathogenesis of hemangioma. PMID- 11254665 TI - The SAMP1/Yit mouse: another step closer to modeling human inflammatory bowel disease. PMID- 11254666 TI - Conserved responses to oxygen deprivation. PMID- 11254667 TI - Hyperhomocysteinemia enhances vascular inflammation and accelerates atherosclerosis in a murine model. AB - Although hyperhomocysteinemia (HHcy) is a well-known risk factor for the development of cardiovascular disease, the underlying molecular mechanisms are not fully elucidated. Here we show that induction of HHcy in apoE-null mice by a diet enriched in methionine but depleted in folate and vitamins B6 and B12 increased atherosclerotic lesion area and complexity, and enhanced expression of receptor for advanced glycation end products (RAGE), VCAM-1, tissue factor, and MMP-9 in the vasculature. These homocysteine-mediated (HC-mediated) effects were significantly suppressed, in parallel with decreased levels of plasma HC, upon dietary supplementation with folate and vitamins B6/B12. These findings implicate HHcy in atherosclerotic plaque progression and stability, and they suggest that dietary enrichment in vitamins essential for the metabolism of HC may impart protective effects in the vasculature. PMID- 11254668 TI - Mutation in pre-mRNA adenosine deaminase markedly attenuates neuronal tolerance to O2 deprivation in Drosophila melanogaster. AB - O2 deprivation can produce many devastating clinical conditions such as myocardial infarct and stroke. The molecular mechanisms underlying the inherent tissue susceptibility or tolerance to O2 lack are, however, not well defined. Since the fruit fly, Drosophila melanogaster, is extraordinarily tolerant to O2 deprivation, we have performed a genetic screen in the Drosophila to search for loss-of-function mutants that are sensitive to low O2. Here we report on the genetic and molecular characterization of one of the genes identified from this screen, named hypnos-2. This gene encodes a Drosophila pre-mRNA adenosine deaminase (dADAR) and is expressed almost exclusively in the adult central nervous system. Disruption of the dADAR gene results in totally unedited sodium (Para), calcium (Dmca1A), and chloride (DrosGluCl-alpha) channels, a very prolonged recovery from anoxic stupor, a vulnerability to heat shock and increased O2 demands, and neuronal degeneration in aged flies. These data clearly demonstrate that, through the editing of ion channels as targets, dADAR, for which there are mammalian homologues, is essential for adaptation to altered environmental stresses such as O2 deprivation and for the prevention of premature neuronal degeneration. PMID- 11254669 TI - Th1-type responses mediate spontaneous ileitis in a novel murine model of Crohn's disease. AB - We describe here the immunologic characterization of a new mouse strain, SAMP1/Yit, which spontaneously develops a chronic intestinal inflammation localized to the terminal ileum. The resulting ileitis bears a remarkable resemblance to human Crohn's disease. This strain of mice develops discontinuous, transmural inflammatory lesions in the terminal ileum with 100% penetrance by 30 weeks of age. The intestinal inflammation is characterized by massive infiltration of activated CD4+ and CD8alpha(+)TCRalphabeta(+) T cells into the lamina propria and is accompanied by a dramatic decrease in the intraepithelial lymphocyte CD8alpha(+)TCRgammadelta(+)/CD8alpha(+)TCRalphabeta(+) ratio. The results of adoptive transfer experiments strongly suggest that CD4+ T cells that produce a Th1-like profile of cytokines, e.g., IFN-gamma and TNF, mediate the intestinal inflammation found in SAMP1/Yit mice. In addition, pretreatment of adoptive transfer recipients with a neutralizing anti-TNF antibody prevents the development of intestinal inflammation, suggesting that TNF plays an important role in the pathogenesis of intestinal inflammation in this model. To our knowledge, these data provide the first direct evidence that Th1-producing T cells mediate intestinal inflammation in a spontaneous animal model of human Crohn's disease. PMID- 11254671 TI - Inducible nitric oxide synthase mediates the change from retinal to vitreal neovascularization in ischemic retinopathy. AB - Intravitreal neovascular diseases are a major cause of blindness worldwide. It remains unclear why neovessels in many retinal diseases spread into the physiologically nonvascularized vitreous rather than into the ischemic retinal areas, where the angiogenic factors are released. Here we show that inducible nitric oxide synthase (iNOS) is expressed in the ischemic retina. Using iNOS knockout mice and the iNOS inhibitor 1400W, we demonstrate that iNOS expression inhibits angiogenesis locally in the avascular retina, mediated at least in part by a downregulation of VEGF receptor 2 (VEGFR2) in cells adjacent to iNOS expressing cells. At the same time, pathological intravitreal neovascularization is considerably stronger in iNOS-expressing animals. These findings demonstrate that iNOS plays a crucial role in retinal neovascular disease and show that it offers an ideal target for the control of vitreal neovascularization through improvement of the vascularization of the hypoxic retina. PMID- 11254670 TI - A novel vascular smooth muscle chymase is upregulated in hypertensive rats. AB - While greater than 80% of angiotensin II (Ang II) formation in the human heart and greater than 60% in arteries appears to result from chymase activity, no cardiovascular cell-expressed chymase has been previously reported. We now describe the cloning of a full-length cDNA encoding a novel chymase from rat vascular smooth muscle cells. The cDNA encompasses 953 nucleotides, encodes 247 amino acids, and exhibits 74% and 80% homology in amino acid sequence to rat mast cell chymase I and II, respectively. Southern blot analysis indicates that the rat vascular chymase is encoded by a separate gene. This chymase was induced in hypertrophied rat pulmonary arteries, with 11-fold and 8-fold higher chymase mRNA levels in aortic and pulmonary artery smooth muscle cells from spontaneously hypertensive than in corresponding tissues from normotensive rats. We assayed the activity of the endogenous enzyme and of a recombinant, epitope-tagged chymase in transfected smooth muscle cells and showed that Ang II production from Ang I can be inhibited with chymostatin, but not EDTA or captopril. Spontaneously hypertensive rats show elevated chymase expression and increased chymostatin inhibitable angiotensin-converting activity, suggesting a possible role for this novel enzyme in the pathophysiology of hypertension. PMID- 11254672 TI - The discoidin domain receptor tyrosine kinase DDR1 in arterial wound repair. AB - Collagens act as important signaling molecules regulating vascular smooth muscle cell responses during arterial wound repair. Discoidin domain receptors (DDRs) are a novel class of receptor tyrosine kinases that bind to several collagens and stimulate matrix metalloproteinase (MMP) production, but little is known about their expression and function in the vasculature. We posited a critical role for the DDRs controlling smooth muscle cell migration and proliferation and thus repair following arterial injury. Smooth muscle cells were isolated from the aortas of mice with a targeted deletion of the DDR1 gene (DDR1-null) and studied in culture using models that mimic critical steps in neointimal thickening. Our studies suggest that DDR1 plays an important role in regulating attachment to collagen, chemotaxis, proliferation, and MMP production in smooth muscle cells. Following mechanical injury to the carotid arteries, cross-sectional area of the neointima was significantly lower in DDR1-null mice than in wild-type mice. There was also a significant decrease in collagen deposition in the injured arteries of the DDR1-null mice. Our results support the hypothesis that DDR1 plays an important role as a collagen receptor, mediating intimal thickening after vascular injury. PMID- 11254673 TI - LP-BM5 virus-infected mice produce activating autoantibodies to the AMPA receptor. AB - Autoantibodies to alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors may contribute to chronic hyperexcitability syndromes and neurodegeneration, but their origin is unclear. We examined LP-BM5 murine leukemia virus-infected mice, which manifest excitotoxic brain lesions and hypergammaglobulinemia, for the presence of AMPA-receptor Ab's. Endogenous IgG accumulated upon neurons in the neocortex and caudate/putamen of infected mice and interacted with native and recombinant AMPA-receptor subunits with the following relative abundance: GluR3 > or = GluR1 > GluR2 = GluR4, as determined by immunoprecipitation. In a radioligand assay, IgG preparations from infected mice specifically inhibited [(3)H]AMPA binding to receptors in brain homogenates, an activity that was lost after preadsorbing the IgG preparation to immobilized LP-BM5 virus. These IgGs also evoked currents when applied to hippocampal pyramidal neurons or to damaged cerebellar granule neurons. These currents could be blocked using any of several AMPA receptor antagonists. Thus, anti-AMPA receptor Ab's can be produced as the result of a virus infection, in part through molecular mimicry. These Ab's may alter neuronal signaling and contribute to the neurodegeneration observed in these mice, actions that may be curtailed by the use of AMPA-receptor antagonists. PMID- 11254674 TI - Clonality and altered behavior of endothelial cells from hemangiomas. AB - Hemangioma, the most common tumor of infancy, is a benign vascular neoplasm of unknown etiology. We show, for the first time to our knowledge, that endothelial cells from proliferating hemangioma are clonal, and we demonstrate that these hemangioma-derived cells differ from normal endothelial cells in their rates of proliferation and migration in vitro. Furthermore, migration of hemangioma endothelial cells is stimulated by the angiogenesis inhibitor endostatin, unlike the inhibition seen with normal endothelial cells. We conclude that hemangiomas constitute clonal expansions of endothelial cells. This is consistent with the possibility that these tumors are caused by somatic mutations in one or more genes regulating endothelial cell proliferation. PMID- 11254676 TI - Imprinting of the G(s)alpha gene GNAS1 in the pathogenesis of acromegaly. AB - Approximately 40% of growth hormone-secreting pituitary adenomas have somatic mutations in the GNAS1 gene (the so-called gsp oncogene). These mutations at codon 201 or codon 227 constitutively activate the alpha subunit of the adenylate cyclase-stimulating G protein G(s). GNAS1 is subject to a complex pattern of genomic imprinting, its various promoters directing the production of maternally, paternally, and biallelically derived gene products. Transcripts encoding G(s)alpha are biallelically derived in most human tissues. Despite this, we show here that in 21 out of 22 gsp-positive somatotroph adenomas, the mutation had occurred on the maternal allele. To investigate the reason for this allelic bias, we also analyzed GNAS1 imprinting in the normal adult pituitary and found that G(s)alpha is monoallelically expressed from the maternal allele in this tissue. We further show that this monoallelic expression of G(s)alpha is frequently relaxed in somatotroph tumors, both in those that have gsp mutations and in those that do not. These findings imply a possible role for loss of G(s)alpha imprinting during pituitary somatotroph tumorigenesis and also suggest that G(s)alpha imprinting is regulated separately from that of the other GNAS1 products, NESP55 and XLalphas, imprinting of which is retained in these tumors. PMID- 11254675 TI - Uroporphyrinogen III synthase erythroid promoter mutations in adjacent GATA1 and CP2 elements cause congenital erythropoietic porphyria. AB - Congenital erythropoietic porphyria, an autosomal recessive inborn error of heme biosynthesis, results from the markedly deficient activity of uroporphyrinogen III synthase. Extensive mutation analyses of 40 unrelated patients only identified approximately 90% of mutant alleles. Sequencing the recently discovered erythroid-specific promoter in six patients with a single undefined allele identified four novel mutations clustered in a 20-bp region: (a) a -70T to C transition in a putative GATA-1 consensus binding element, (b) a -76G to A transition, (c) a -86C to A transversion in three unrelated patients, and (d) a 90C to A transversion in a putative CP2 binding motif. Also, a -224T to C polymorphism was present in approximately 4% of 200 unrelated Caucasian alleles. We inserted these mutant sequences into luciferase reporter constructs. When transfected into K562 erythroid cells, these constructs yielded 3 +/- 1, 54 +/- 3, 43 +/- 6, and 8 +/- 1%, respectively, of the reporter activity conferred by the wild-type promoter. Electrophoretic mobility shift assays indicated that the 70C mutation altered GATA1 binding, whereas the adjacent -76A mutation did not. Similarly, the -90C mutation altered CP2 binding, whereas the -86A mutation did not. Thus, these four pathogenic erythroid promoter mutations impaired erythroid specific transcription, caused CEP, and identified functionally important GATA1 and CP2 transcriptional binding elements for erythroid-specific heme biosynthesis. PMID- 11254677 TI - CCR6-deficient mice have impaired leukocyte homeostasis and altered contact hypersensitivity and delayed-type hypersensitivity responses. AB - CCR6 expression in dendritic, T, and B cells suggests that this beta-chemokine receptor may regulate the migration and recruitment of antigen-presenting and immunocompetent cells during inflammatory and immunological responses. Here we demonstrate that CCR6-/- mice have underdeveloped Peyer's patches, in which the myeloid CD11b+ CD11c+ dendritic-cell subset is not present in the subepithelial dome. CCR6-/- mice also have increased numbers in T-cell subpopulations within the intestinal mucosa. In 2,4-dinitrofluorobenzene-induced contact hypersensitivity (CHS) studies, CCR6-/- mice developed more severe and more persistent inflammation than wild-type (WT) animals. Conversely, in a delayed type hypersensitivity (DTH) model induced with allogeneic splenocytes, CCR6-/- mice developed no inflammatory response. The altered responses seen in the CHS and DTH assays suggest the existence of a defect in the activation and/or migration of the CD4(+) T-cell subsets that downregulate or elicit the inflammation response, respectively. These findings underscore the role of CCR6 in cutaneous and intestinal immunity and the utility of CCR6-/- mice as a model to study pathologies in these tissues. This article was published online in advance of the print edition. The date of publication is available from the JCI website, http://www.jci.org. PMID- 11254678 TI - Cutting edge: differential signaling requirements for activation of assembled cyclin D3-cdk4 complexes in B-1 and B-2 lymphocyte subsets. AB - B-1 lymphocytes represent a distinct B cell subset with unusual mitogenic responses. PMA alone promotes proliferation in B-1 cells, but not in splenic B-2 cells. Although cyclin D2-cyclin-dependent kinase 4 (cdk4) complexes mediate early retinoblastoma gene product (pRb) phosphorylation in B-1 cells, the transient nature of their accumulation cannot account for the continued increase in pRb phosphorylation, which is maximal at 24 h. We show herein that PMA promotes the accumulation of functional cyclin D3-cdk4 complexes in B-1 cells following loss of cyclin D2. PMA also induces accumulation of cyclin D3-cdk4 complexes in B-2 cells; however, these complexes do not phosphorylate pRb. Thus, PMA is sufficient to induce synthesis and assembly of cyclin D3-cdk4 complexes in B-1 and B-2 cells; however, PMA triggers cyclin D3-cdk4 activation only in B-1 cells. These results reveal a novel regulatory step that controls activation of cyclin D3-cdk4 complexes whose function segregates differentially in B cell subsets. PMID- 11254679 TI - Cutting edge: in vivo identification of TCR redistribution and polarized IL-2 production by naive CD4 T cells. AB - TCR aggregation at the point of contact with an APC is thought to play an important role in T cell signal transduction. However, this potentially important phenomenon has never been documented during an immune response in vivo. Here we used immunohistology to show that the TCR on naive Ag-specific CD4 T cells in the lymph nodes of mice injected with Ag redistributed to one side of the cell. In cases where the APC could be identified, the TCR was concentrated on the side of the T cell facing the APC. In those T cells that produced IL-2, the TCR and IL-2 localized to the same side of the cell. In vivo IL-2 production depended on costimulatory signaling through CD28, whereas TCR redistribution did not. These results show that Ag-stimulated CD4 T cells produce IL-2 in a polarized fashion and undergo CD28-independent TCR redistribution in vivo. PMID- 11254680 TI - Antigen-independent Th2 cell differentiation by stimulation of CD28: regulation via IL-4 gene expression and mitogen-activated protein kinase activation. AB - To delineate the molecular mechanisms regulating Th2 cell differentiation, CD28 mediated generation of Th2 effectors was analyzed. In the absence of TCR ligation CD28 stimulation induced Th2 differentiation of memory but not of naive CD4(+) T cells, whereas costimulation via CD28 and the TCR enhanced Th2 differentiation from naive T cells but suppressed it from memory T cells. Stimulation of T cells via the CD28 pathway, therefore, provided critical signals facilitating Th2 cell differentiation. By comparing the responses to CD28 stimulation in memory and naive T cells and by using specific inhibitors, signaling pathways were defined that contributed to Th2 differentiation. CD28-induced Th2 differentiation required IL-4 stimulation and the activation of the mitogen-activated protein kinases p38 and extracellular signal-regulated kinases 1/2. CD28 engagement directly initiated IL-4 gene transcription in memory T cells and induced activation of phosphatidylinositol 3-kinase, p38, and c-Jun NH(2)-terminal kinase/stress-activated protein kinase pathways. Extracellular signal-regulated kinase phosphorylation that was necessary for Th2 differentiation, however, required stimulation by IL-2. These results indicate that optimal TCR-independent generation of Th2 effectors requires coordinate signaling via the CD28 and IL-2 pathways. TCR-independent generation of Th2 effectors might provide a mechanism to control Th1-dominated cellular inflammation. PMID- 11254681 TI - Stromal cell-derived factor-1-induced LFA-1 activation during in vivo migration of T cell hybridoma cells requires Gq/11, RhoA, and myosin, as well as Gi and Cdc42. AB - Dissemination of T cell hybridomas in mice, a model for in vivo migration of memory T cells and for T lymphoma metastasis, depends on the chemokine stromal cell-derived factor-1 (SDF-1) and the integrin LFA-1 and correlates well with invasion into fibroblast cultures. In addition to the known role of the pertussis toxin-sensitive heterotrimeric GTPase G(i), we show that also the pertussis toxin insensitive GTPase G(q/11) is required for dissemination and invasion. Furthermore, we show that the small GTPases, Cdc42 and RhoA, are involved, and that invasion is blocked by inhibitors of actinomyosin contraction. G(q/11), RhoA, and contraction are specifically required for LFA-1 activation, since 1) they are essential for LFA-1-dependent migration toward low SDF-1 concentrations through ICAM-1-coated filters, but not for migration toward high SDF-1 levels, which is LFA-1 independent; 2) G protein (AlF(4)(-))-induced adhesion to ICAM-1 requires RhoA and contraction; 3) constitutively active G(q) induces aggregation, mediated by LFA-1. We previously reported that binding of this activated LFA-1 to ICAM-1 triggers a signal, transduced by the zeta-associated protein 70 tyrosine kinase, that activates additional LFA-1 molecules. This amplification of LFA-1 activation is essential for invasion. We show here that zeta-associated protein 70-induced LFA-1 activation requires neither Cdc42 and RhoA nor contraction and is thus quite different from that induced by SDF-1. We conclude that two modes of LFA-1 activation, with distinct underlying mechanisms, are required for the in vivo migration of T cell hybridomas. PMID- 11254682 TI - Expression of Ly49E and CD94/NKG2 on fetal and adult NK cells. AB - Murine NK cells express inhibitory receptors belonging to the Ly49 and CD94/NKG2 family. Ly49E and CD94 are the only NK cell receptor transcripts detectable in fetal NK cells. Still unproved is the surface expression of Ly49E on NK cells. Here we generated two novel mAbs, a mAb recognizing Ly49E with cross-reactivity to Ly49C, and a mAb against NKG2A/C/E. Ly49E was immunoprecipitated as a disulfide-linked homodimer with 46-kDa subunits. Removal of N-linked carbohydrates revealed a 31-kDa protein backbone. NKG2A was immunoprecipitated as a 38-kDa protein. Although the frequency of fetal NK cells expressing Ly49E was higher than 25%, it decreased drastically from 2 wk after birth. Phenotypic analysis showed that approximately 90% of fetal NK cells and approximately 50% of adult NK cells express high levels of CD94/NKG2. The remaining 50% of adult NK cells expressed low surface levels of CD94/NKG2. Expression of Ly49E and CD94/NKG2 was not restricted to NK cells, but was also observed on NK T and memory T cells. Functional analysis showed that sorted Ly49E(+) and CD94/NKG2(+) fetal NK cells could discriminate between MHC class I-positive and MHC class I negative tumor cells. We also demonstrated that Ly49E becomes phosphorylated following pervanadate stimulation of fetal NK cells. The expression levels of Ly49E and CD94/NKG2 were similar in wild-type compared with beta(2) microglobulin(-/-) mice. In conclusion, generation of mAbs against Ly49E and NKG2 extended the phenotypic and functional characterization of NK cells. PMID- 11254683 TI - Regulatory activity of autocrine IL-10 on dendritic cell functions. AB - IL-10 is a critical cytokine that blocks the maturation of dendritic cells (DCs), but the relevance of autocrine IL-10 on DC functions has not been investigated. In this study, we found that immature monocyte-derived DCs released low but sizeable amounts of IL-10. After stimulation with bacteria, LPS, lipoteichoic acid, or soluble CD40 ligand, DCs secreted high levels of IL-10. Addition of an anti-IL-10-neutralizing Ab to immature DCs as well as to soluble CD40 ligand- or LPS-maturing DCs led to enhanced expression of surface CD83, CD80, CD86, and MHC molecules and markedly augmented release of TNF-alpha and IL-12, but diminished IL-10 mRNA expression. Moreover, DCs treated with anti-IL-10 Ab showed an increased capacity to activate allogeneic T cells and primed naive T cells to a more prominent Th1 polarization. DC maturation and IL-10 neutralization were associated with enhanced accumulation of the IL-10 receptor binding chain (IL 10R1) mRNA and intracellular IL-10R1 protein. In contrast, surface IL-10R1 and IL 10 binding activity diminished in mature DCs. These results indicate that autocrine IL-10 prevents spontaneous maturation of DCs in vitro, limits LPS- and CD40-mediated maturation, and increases IL-10 production by DCs. Moreover, IL-10R expression appears to be regulated by both transcriptional and posttranscriptional mechanisms. Endogenous IL-10 and IL-10R can be relevant targets for the manipulation of DC functions. PMID- 11254684 TI - Identification of an IL-2 binding protein in the saliva of the Lyme disease vector tick, Ixodes scapularis. AB - A potent inhibitor of mitogen-stimulated T cell proliferation exists in the saliva of several species of hard ticks, including the Lyme disease vector tick, Ixodes scapularis. Our characterization of this phenomenon has led to the identification of a possible mechanism for the T cell inhibitory activity of I. scapularis saliva. The T cell inhibitor can overcome stimulation of mouse spleen cells with anti-CD3 mAb; however, a direct and avid interaction with T cells does not appear to be necessary. Tick saliva inhibits a mouse IL-2 capture ELISA, suggesting that a soluble IL-2 binding factor is present in the saliva. This hypothesis was verified by using a direct binding assay in which plate immobilized tick saliva was shown to bind both mouse and human IL-2. Elimination of the IL-2 binding capacity of saliva in the in vitro assays by trypsin digestion demonstrated that the IL-2 binding factor is a protein. These experiments comprise the first demonstration of the existence of such a secreted IL-2 binding protein from any parasite or pathogen. This arthropod salivary IL-2 binding capacity provides a simple mechanism for the suppression of T cell proliferation as well as for the activity of other immune effector cells that are responsive to IL-2 stimulation. Relevance of the tick T cell inhibitory activity to the human immune system is demonstrated by the ability of tick saliva to inhibit proliferation of human T cells and CTLL-2 cells grown in the presence of human IL-2. PMID- 11254685 TI - Gamma delta T cells are needed for ocular immune privilege and corneal graft survival. AB - It has been recognized for over a century that the anterior chamber of the eye is endowed with a remarkable immune privilege. One contributing component is the Ag specific down-regulation of systemic delayed-type hypersensitivity (DTH) that is induced when Ags are introduced into the anterior chamber. This phenomenon, termed anterior chamber-associated immune deviation (ACAID), culminates in the generation of regulatory cells that inhibit the induction (afferent suppression) and expression (efferent suppression) of DTH. Since gamma delta T cells play a major role in other forms of immune regulation, we suspected they might contribute to the induction and expression of ACAID. Mice treated with anti-gamma delta Ab failed to develop ACAID following anterior chamber injection of either soluble Ag (OVA) or alloantigens (spleen cells). Additional experiments with knockout mice confirmed that mice lacking functional gamma delta T cells also fail to develop ACAID. Using a local adoptive transfer of DTH assay, we found that gamma delta T cells were required for the generation of regulatory T cells, but did not function as the efferent regulatory cells of ACAID. The importance of gamma delta T cells in corneal allograft survival was confirmed by blocking gamma delta T cells with GL3 Ab before corneal transplantation. While in vivo treatment with normal hamster serum had no effect on corneal graft survival, infusion of anti-gamma delta Ab resulted in a profound increase in corneal allograft rejection. Thus, gamma delta T cells are needed for sustaining at least one aspect of ocular immune privilege and for promoting corneal allograft survival. PMID- 11254686 TI - In vivo neutralization of human IL-6 (hIL-6) achieved by immunization of hIL-6 transgenic mice with a hIL-6 receptor antagonist. AB - Neutralization of IL-6 represents an attractive therapeutic option in several diseases, including B cell neoplasia, osteoporosis, and autoimmunity. Therapeutic attempts in humans have shown that administration of injectable doses of a mAb to IL-6 does not provide efficient neutralization of the cytokine in vivo. Therefore, alternative approaches are needed. In this study, we evaluated whether the Ab response to human IL-6 (hIL-6) elicited by vaccination with Sant1 (a hIL-6 variant with seven amino acid substitutions) was able to fully correct in vivo the clinical and biological effects of a chronic endogenous overproduction of hIL 6 in the hIL-6-transgenic NSE/hIL-6 mice. Because of the overexpression of hIL-6, occurring since birth, with circulating levels in the nanogram per milliliter range, NSE/hIL-6 mice have a marked decrease in growth rate, associated with decrease in insulin-like growth factor I levels, and represent an animal model of the growth impairment associated with human chronic inflammatory diseases. Following immunization with Sant1, but not with hIL-6, NSE/hIL-6 mice developed high titers of polyclonal Abs to hIL-6. The Abs, acquired by transplacental transfer, effectively neutralized IL-6 activities in vivo as shown by the complete correction of the growth defect and normalization of insulin-like growth factor levels in the hIL-6-transgenic offspring. Immunization with Sant1 could therefore represent a novel and simple therapeutic approach for the specific neutralization of IL-6 in humans. PMID- 11254687 TI - Biological activity of soluble CD100. I. The extracellular region of CD100 is released from the surface of T lymphocytes by regulated proteolysis. AB - CD100 is the first semaphorin described in lymphoid tissues, where it has been shown to be associated with a serine kinase activity. Semaphorins are molecules involved in axon pathfinding during nerve development and act as repellent guidance cues. In the nervous system semaphorins exist as either membrane-bound or secreted forms. We report here a spontaneous processing of membrane CD100, suggesting that it is also produced as a diffusable semaphorin from lymphoid cells. Monomeric and homodimeric forms of CD100 are expressed by T lymphocytes and CD100-transfected fibroblasts. We demonstrate that CD100 is released through a proteolytic process blocked by metalloprotease inhibitors. In T cells, only soluble CD100 dimers are produced, suggesting that CD100 dimerization is required for proteolysis. In agreement, we observe that increasing membrane dimers strongly favors shedding of the molecule. By expressing a CD100 molecule mutated at cysteine 674 into a COS cell system, we additionally demonstrate that this particular residue in the extracellular domain of the molecule is required for dimerization. Finally, we show that staurosporine, a serine kinase inhibitor, enhances the membrane cleavage of CD100. Together these results demonstrate that membrane CD100 is cleaved by a metalloprotease-dependent process, which is probably regulated by phosphorylation. Mainly, these findings shed light on a possible function for the semaphorin region of CD100 as a long range guidance cue in the immune system. PMID- 11254688 TI - Biological activity of soluble CD100. II. Soluble CD100, similarly to H-SemaIII, inhibits immune cell migration. AB - CD100 is a human 150-kDa homodimer expressed at the surface of most hemopoietic cells, and its gene belongs to the Ig and semaphorin gene families. Semaphorin genes encode soluble and membrane-bound proteins, most of which have been shown to act as chemorepellents on growth cone guidance. CD100 is discrete, as it is a transmembrane leukocyte surface molecule that can also exist in a soluble form. While our previous studies using mAbs suggested that the transmembrane form of CD100 plays a role in lymphocyte activation, no function was shown for its soluble form. Here, we investigated the effect of soluble CD100 in a cell migration assay; both CD100 spontaneously shed from a stable transfectant and soluble recombinant CD100 inhibited spontaneous and chemokine-induced migration of human monocytes. Interestingly, only the dimeric form of CD100 exerted an effect. Moreover, soluble CD100 inhibited migration of cells from monocytic and B cell lineages. A similar inhibitory effect on migration was observed with H SemaIII, but not H-SemaIV, semaphorins. In addition, both CD100 and H-SemaIII were recognized by two CD100 mAbs in an ELISA, and one of these mAb abolished the inhibitory effect of each of these semaphorins. We also provide evidence that CD100 and H-SemaIII act through the same receptor on immune cells, which is not neuropilin-1. Furthermore, we describe a function on immune cells for H-SemaIII, a semaphorin to date only studied in the nervous system. PMID- 11254689 TI - Multiple antigen-specific processing pathways for activating naive CD8+ T cells in vivo. AB - Current knowledge of the processing of viral Ags into MHC class I-associated ligands is based almost completely on in vitro studies using nonprofessional APCs (pAPCs). This is two steps removed from real immune responses to pathogens and vaccines, in which pAPCs activate naive CD8(+) T cells in vivo. Rational vaccine design requires answers to numerous questions surrounding the function of pAPCs in vivo, including their abilities to process and present peptides derived from endogenous and exogenous viral Ags. In the present study, we characterize the in vivo dependence of Ag presentation on the expression of TAP by testing the immunogenicity of model Ags synthesized by recombinant vaccinia viruses in TAP1( /-) mice. We show that the efficiency of TAP-independent presentation in vitro correlates with TAP-independent activation of naive T cells in vivo and provide the first in vivo evidence for proteolytic processing of antigenic peptides in the secretory pathway. There was, however, a clear exception to this correlation; although the presentation of the minimal SIINFEKL determinant from chicken egg OVA in vitro was strictly TAP dependent, it was presented in a TAP-independent manner in vivo. In vivo presentation of the same peptide from a fusion protein retained its TAP dependence. These results show that determinant-specific processing pathways exist in vivo for the generation of antiviral T cell responses. We present additional findings that point to cross-priming as the likely mechanism for these protein-specific differences. PMID- 11254690 TI - Endogenous oocyte antigens are required for rapid induction and progression of autoimmune ovarian disease following day-3 thymectomy. AB - Female (C57BL/6xA/J)F(1) mice undergoing thymectomy on day 3 after birth (d3tx) developed autoimmune ovarian disease (AOD) and autoimmune disease of the lacrimal gland. As both were prevented by normal adult CD25(+) T cells, regulatory T cell depletion is responsible for d3tx diseases. AOD began as oophoritis at 3 wk. By 4 wk, AOD progressed to ovarian atrophy with autoantibody response against multiple oocyte Ag of early ontogeny. The requirement for immunogenic endogenous ovarian Ag was investigated in d3tx female mice, d3tx male mice, and d3tx neonatally ovariectomized (OX) females. At 8 wk, all mice had comparable lacrimalitis but only those with endogenous ovaries developed AOD in ovarian grafts. The duration of Ag exposure required to initiate AOD was evaluated in d3tx mice OX at 2, 3, or 4 wk and engrafted with an ovary at 4, 5, or 6 wk, respectively. The mice OX at 2 wk did not have oophoritis whereas approximately 80% of mice OX at 3 or 4 wk had maximal AOD, thus Ag stimulus for 2.5 wk following d3tx is sufficient. AOD progression requires additional endogenous Ag stimulation from the ovarian graft. In mice OX at 3 wk, ovaries engrafted at 5 wk had more severe oophoritis than ovaries engrafted at 6 or 12 wk; moreover, only mice engrafted at 5 wk developed ovarian atrophy and oocyte autoantibodies. Similar results were obtained in mice OX at 4 wk. Thus endogenous tissue Ag are critical in autoimmune disease induction and progression that occur spontaneously upon regulatory T cell depletion. PMID- 11254691 TI - Quantitative analysis of the immune response to mouse non-MHC transplantation antigens in vivo: the H60 histocompatibility antigen dominates over all others. AB - Minor histocompatibility Ags (minor H Ags) are substantial impediments to MHC matched solid tissue and bone marrow transplantation. From an antigenic standpoint, transplantation between MHC-matched individuals has the potential to be remarkably complex. To determine the extent to which the immune response is simplified by the phenomenon of immunodominance, we used peptide/MHC tetramers based on recently discovered minor H Ags (H60, H13, and HY) and monitored in vivo CD8 T cell responses of female C57BL/6 mice primed with MHC-matched, but background-disparate, male BALB.B cells. CD8 T cells against H60 overwhelmed responses to the H13 and HY throughout primary and secondary challenge. H60 immunodominance was an inherent quality, overcoming a lower memory precursor frequency compared with that of H13 and evoking a T cell response with diverse TCRV beta usage. IFN-gamma staining examining congenically defined minor H Ags extended H60 dominance over additional minor H Ags, H28, H4, and H7. These four minor H Ags accounted for up to 85% of the CD8 T cell response, but H60 stood out as the major contributor. These findings show that immunodominance applies to antigenically complex transplantation settings in vivo and that the responses to the H60 minor H Ag dominates in this model. We suggest that immunodominant minor H Ags are those that result from the absence of a self analog. PMID- 11254692 TI - Diversity of the killer cell Ig-like receptors of rhesus monkeys. AB - Because the killer cell Ig-like receptors (KIRs) have only been characterized in humans and chimpanzees, we do not have a full understanding of their evolutionary history. Therefore, cDNAs encoding the KIR molecules of five rhesus monkeys were characterized, and were found to differ from the KIR molecules identified in humans and chimpanzees. Whereas only one KIR2DL4 molecule is detected in humans and chimpanzees, two distinct KIR2DL4 homologues were identified in the monkeys. Although the two human KIR3DL molecules are limited in their polymorphism, the KIR3DL homologues in the monkeys were highly polymorphic. Up to five KIR3DL homologues were identified in each monkey that was studied, and eleven distinct KIR3DL molecules were detected in the five rhesus monkeys. Two novel families of KIR molecules were identified in the rhesus monkeys, KIR3DH and KIR1D. The KIR3DH molecules have three Ig domains, transmembrane domains homologous to KIR2DL4 molecules that contain an arginine, and short cytoplasmic domains. With these features, the KIR3DH molecules resemble the activating forms of the human KIR molecules. The KIR1D molecule encodes only one complete Ig domain before a frame shift in the second Ig domain occurs, leading to early termination of the molecule. Multiple splice variants of KIR1D exist that encode at least one Ig domain, as well as transmembrane and cytoplasmic domains. The extensive diversity of the rhesus monkey KIR3DL homologues and the novel KIR3DH and KIR1D molecules suggests that the KIR family of molecules has evolved rapidly during the evolution of primates. PMID- 11254693 TI - IL-9 and IL-13 production by activated mast cells is strongly enhanced in the presence of lipopolysaccharide: NF-kappa B is decisively involved in the expression of IL-9. AB - Mast cells, due to their ability to produce a large panel of mediators and cytokines, participate in a variety of processes in adaptive and innate immunity. Herein we report that in primary murine bone marrow-derived mast cells activated with ionomycin or IgE-Ag the bacterial endotoxin LPS strongly enhances the expression of IL-9 and IL-13, but not IL-4. This costimulatory effect of LPS is absent in activated mast cells derived from the LPS-hyporesponsive mouse strain BALB/c-LPS(d), although in these cells the proinflammatory cytokine IL-1 can still substitute for LPS. The enhanced production of mast cell-derived IL-13 in the presence of IL-1 is a novel observation. Coactivation of mast cells with LPS leads to a synergistic activation of NF-kappa B, which is shown by an NF-kappa B driven reporter gene construct. In the presence of an inhibitor of NF-kappa B activation, the production of IL-9 is strongly decreased, whereas the expression of IL-13 is hardly reduced, and that of IL-4 is not affected at all. NF-kappa B drives the expression of IL-9 via three NF-kappa B binding sites within the IL-9 promoter, which we characterize using gel shift analyses and reporter gene assays. In the light of recent reports that strongly support critical roles for IL-9 and IL-13 in allergic lung inflammation, our results emphasize the potential clinical importance of LPS as an enhancer of mast cell-derived IL-9 and IL-13 production in the course of inflammatory reactions and allergic diseases. PMID- 11254694 TI - Regulation of activator protein-1 and NF-kappa B in CD8+ T cells exposed to peripheral self-antigens. AB - The transcriptional events that control T cell tolerance to peripheral self Ags are still unknown. In this study, we analyzed the regulation of AP-1- and NF kappa B-mediated transcription during in vivo induction of tolerance to a self Ag expressed exclusively on hepatocytes. Naive CD8(+)Desire (Des)(+) T cells isolated from the Des TCR-transgenic mice that are specific for the H-2K(b) class I Ag were transferred into Alb-K(b)-transgenic mice that express the H-2K(b) Ag on hepatocytes only. Tolerance develops in these mice. We found that the self reactive CD8(+)Des(+) T cells were transiently activated, then became unresponsive and were further deleted. In contrast to CD8(+)Des(+) T cells activated in vivo with APCs, which express high AP-1 and high NF-kappa B transcriptional activity, the unresponsive CD8(+)Des(+) T cells expressed no AP-1 and only weak NF-kappa B transcriptional activity. The differences in NF-kappa B transcriptional activity correlated with the generation of distinct NF-kappa B complexes. Indeed, in vivo primed T cells predominantly express p50/p50 and p65/p50 dimers, whereas these p50-containing complexes are barely detectable in tolerant T cells that express p65- and c-Rel-containing complexes. These observations suggest that fine regulation of NF-kappa B complex formation may determine T cell fate. PMID- 11254695 TI - p62dok negatively regulates CD2 signaling in Jurkat cells. AB - p62(dok) belongs to a newly identified family of adaptor proteins. In T cells, the two members that are predominantly expressed, p56(dok) and p62(dok), are tyrosine phosphorylated upon CD2 or CD28 stimulation, but not upon CD3 ligation. Little is known about the biological role of Dok proteins in T cells. In this study, to evaluate the importance of p62(dok) in T cell function, we generated Jurkat clones overexpressing p62(dok). Our results demonstrate that overexpression of p62(dok) in Jurkat cells has a dramatic negative effect on CD2 mediated signaling. The p62(dok)-mediated inhibition affects several biochemical events initiated by CD2 ligation, such as the increase of intracellular Ca(2+), phospholipase C gamma 1 activation, and extracellular signal-regulated kinase 1/2 activation. Importantly, these cellular events are not affected in the signaling cascade induced by engagement of the CD3/TCR complex. However, both CD3- and CD2 induced NF-AT activation and IL-2 secretion are impaired in p62(dok) overexpressing cells. In addition, we show that CD2 but not CD3 stimulation induces p62(dok) and Ras GTPase-activating protein recruitment to the plasma membrane. These results suggest that p62(dok) plays a negative role at multiple steps in the CD2 signaling pathway. We propose that p62(dok) may represent an important negative regulator in the modulation of the response mediated by the TCR. PMID- 11254696 TI - Nuclear shuttling of mitogen-activated protein (MAP) kinase (extracellular signal regulated kinase (ERK) 2) was dynamically controlled by MAP/ERK kinase after antigen stimulation in RBL-2H3 cells. AB - The mitogen-activated protein kinase (MAPK) cascade consists of the MAPK (extracellular signal-regulated kinase 2; ERK2) and its activator, MAPK kinase (MAP/ERK kinase; MEK). However, the mechanisms for activation of ERK2 have not been defined yet in cells. Here, we used fluorescent protein-tagged ERK2 and MEK to examine the localization of ERK2 and MEK in living rat basophilic leukemia (RBL-2H3) cells. ERK2 was mainly in the cytoplasm in resting cells but translocated into the nucleus after the ligation of IgE receptors. The import of ERK2 reached the maximum at 6--7 min, and then the imported ERK2 was exported from the nucleus. MEK mainly resided in the cytoplasm, and no significant MEK translocation was detected statically after ligation of IgE receptors. However, analysis of the dynamics of ERK2 and MEK suggested that both of them rapidly shuttle between the cytoplasm and the nucleus and that MEK regulates the nuclear shuttling of ERK2, whereas MEK remains mainly in the cytoplasm. In addition, the data suggested that the sustained calcium increase was required for the optimal translocation of ERK2 into the nucleus in RBL-2H3 cells. These results gave a new insight of the dynamics of ERK2 and MEK in the nuclear shuttling of RBL-2H3 cells after the ligation of IgE receptors. PMID- 11254697 TI - The NK cell MHC class I receptor Ly49A detects mutations on H-2Dd inside and outside of the peptide binding groove. AB - The NK cell inhibitory receptor Ly49A recognizes the mouse MHC class I molecule H 2D(d) and participates in the recognition of missing self. Previous studies indicated that the determinant recognized by Ly49A exists in alpha1/alpha2 domain of H-2D(d). Here we have substituted polymorphic as well as conserved residues of H-2D(d) alpha1/alpha2 domain (when compared with H-2K(d), which does not interact with Ly49A). We then tested the ability of the H-2D(d) mutants to interact with Ly49A by soluble Ly49A tetramer binding and NK cell cytotoxicity inhibition assays. Individual introduction of mutations converting the H-2D(d) residue into the corresponding H-2K(d) residue (N30D, D77S, or A99F) in H-2D(d) partially abrogated the interaction between Ly49A and H-2D(d). Introduction of the three mutations into H-2D(d) completely abolished Ly49A recognition. Individual introduction of D29N or R35A mutation into the residues of H-2D(d) that are conserved among murine MHC class I severely impaired the interaction. The crystal structure of H-2D(d) reveals that D77 and A99 are located in the peptide binding groove and that N30, D29, and R35 are in the interface of the three structural domains of MHC class I: alpha1/alpha2, alpha3, and beta(2)-microglobulin. These data suggest that Ly49A can monitor mutations in MHC class I inside and outside of the peptide binding groove and imply that inhibitory MHC class I-specific receptors are sensitive to mutations in MHC class I as well as global loss of MHC class I. Our results also provide insight into the molecular basis of Ly49A to distinguish MHC class I polymorphism. PMID- 11254698 TI - A role for accessibility to self-peptide-self-MHC complexes in intrathymic negative selection. AB - Whether intrathymic-positive and -negative selection of conventional alpha beta T cells occur in anatomically distinct sites is a matter of debate. By using a system composed of two distinct immune receptors, the Y-Ae mAb and the 1H3.1 (V alpha 1/V beta 6) TCR, both directed against the 52--68 fragment of the I-E alpha chain (E alpha 52--68) bound to I-A(b), we examined the occurrence of negative selection imposed in vivo by a self-peptide-self-MHC class II complex with differential tissue expression. 1H3.1 TCR-transgenic (Tg) mice were bred to mice having an I-E alpha transgene with expression directed to all MHC class II positive cells, restricted to thymic epithelial cells, or restricted to B cells, dendritic cells, and medullary thymic epithelial cells. All 1H3.1 TCR/I-E alpha double-Tg mice revealed a severely diminished thymic cellularity. Their lymph node cells were depleted of V beta 6(+)CD4(+) cells and were unresponsive to E alpha 52--68 in vitro. The absolute number of CD4(+)CD8(+) thymocytes was drastically reduced in all combinations, indicating that negative selection caused by an endogenously expressed self-determinant can effectively occur in the thymic cortex in vivo. Moreover, both cortical epithelial cells and, interestingly, the few cortical dendritic cells were able to support negative selection of CD4(+)CD8(+) thymocytes, albeit with a distinct efficiency. Collectively, these observations support a model where, in addition to the avidity of the thymocyte/stromal cell interaction, in vivo negative selection of autoreactive TCR-Tg T cells is determined by accessibility to self-peptide-self MHC complexes regardless of the anatomical site. PMID- 11254699 TI - Distinguishing self from nonself: immunogenicity of the murine H47 locus is determined by a single amino acid substitution in an unusual peptide. AB - Histocompatibility (H) Ags are responsible for chronic graft rejection and graft vs host disease in solid tissue and bone marrow transplantation among MHC-matched individuals. Here we defined the molecular basis of self-nonself discrimination for the murine chromosome 7 encoded H47 histocompatibility locus, known by its trait of graft-rejection for over 40 years. H47 encodes a novel, highly conserved cell surface protein containing the SCILLYIVI (SII9) nonapeptide in its transmembrane region. The p7 isoleucine-to-phenylalanine substitution in SII9 defined the antigenic polymorphism and T cell specificity. Despite absence of the canonical consensus motif and weak binding to D(b) MHC I, both H47 peptides were presented to CTLs. However, unlike all the other known H loci, the relative immunogenicity of both H47 alleles varied dramatically and was profoundly influenced by neighboring H loci. The results provide insights into the peptide universe that defines nonself and the basis of histoincompatibility. PMID- 11254700 TI - Inducible expression of Stat4 in dendritic cells and macrophages and its critical role in innate and adaptive immune responses. AB - Autocrine activation of APC by IL-12 has recently been revealed; we demonstrate here that inducible expression of Stat4 in APC is central to this process. Stat4 is induced in dendritic cells (DC) in a maturation-dependent manner and in macrophages in an activation-dependent manner. Stat4 levels directly correlate with IL-12-dependent IFN-gamma production by APC as well as IFN-gamma production by DC during Ag presentation. The Th2 cytokines IL-4 and IL-10 suppress Stat4 induction in DC and macrophages when present during maturation and activation, respectively, diminishing IFN-gamma production. In contrast, IL-4 has no effect on Stat4 levels in mature DC and actually augments IFN-gamma production by DC during Ag presentation, indicating that IL-4 acts differently in a spatiotemporal manner. The functional importance of Stat4 is evident in Stat4(-/-) DC and macrophages, which fail to produce IFN-gamma. Furthermore, Stat4(-/-) macrophages are defective in NO production in response to IL-12 and are susceptible to TOXOPLASMA: Autocrine IL-12 signaling is required for high-level IFN-gamma production by APC at critical stages in both innate and adaptive immunity, and the control of Stat4 expression is likely an important regulator of this process. PMID- 11254702 TI - Spontaneous and continuous cyclooxygenase-2-dependent prostaglandin E2 production by stromal cells in the murine small intestine lamina propria: directing the tone of the intestinal immune response. AB - The mechanisms allowing the gastrointestinal immune system to avoid an inappropriate inflammatory response to nonpathogenic luminal Ags are poorly understood. We have previously described a role for cyclooxygenase (COX)-2 dependent arachidonic acid metabolites produced by the murine small intestine lamina propria in controlling the immune response to a dietary Ag. To better understand the role of COX-2-dependent arachidonic acid metabolites produced by the lamina propria, we examined the pattern of expression and the cellular source of COX-2 and COX-2-dependent PGE(2). We now demonstrate that non-bone marrow derived lamina propria stromal cells have basal COX-2 expression and that COX-2 dependent PGE(2) production by these cells is spontaneous and continuous. The other mucosal and nonmucosal lymphoid compartments examined do not share this phenotype. In contrast to the majority of descriptions of COX-2 expression, COX-2 expression by lamina propria stromal cells is not dependent upon exogenous stimuli, including adhesion, LPS signaling via Toll-like receptor 4, or the proinflammatory cytokines TNF-alpha, IFN-gamma, and IL-1 beta. These findings, in conjunction with the known immunomodulatory capacities of PGs, suggest that COX-2 expression by the small intestine lamina propria is a basal state contributing to the hyporesponsiveness of the intestinal immune response. PMID- 11254701 TI - B cell-deficient (mu MT) mice have alterations in the cytokine microenvironment of the gut-associated lymphoid tissue (GALT) and a defect in the low dose mechanism of oral tolerance. AB - Peripheral immune tolerance following i.v. administration of Ag has been shown to occur in the absence of B cells. Because different mechanisms have been identified for i.v. vs low dose oral tolerance and B cells are a predominant component of the gut-associated lymphoid tissue (GALT) they may play a role in tolerance induction following oral Ag. To examine the role of B cells in oral tolerance we fed low doses of OVA or myelin oligodendrocyte glycoprotein to B cell-deficient ( microMT) and wild-type C57BL/6 mice. Results showed that the GALT of naive wild-type and microMT mice was characterized by major differences in the cytokine microenvironment. Feeding low doses of 0.5 mg OVA or 250 microg myelin oligodendrocyte glycoprotein resulted in up-regulation of IL-4, IL-10, and TGF-beta in the GALT of wild-type but not microMT mice. Upon stimulation of popliteal node cells, in vitro induction of regulatory cytokines TGF-beta and IL 10 was observed in wild-type but not microMT mice. Greater protection against experimental autoimmune encephalomyelitis was found in wild-type mice. Oral tolerance in microMT and wild-type mice was found to proceed by different mechanisms. Anergy was observed from 0.5 mg to 250 ng in microMT mice but not in wild-type mice. Increased Ag was detected in the lymph of microMT mice. No cytokine-mediated suppression was found following lower doses from 100 ng to 500 pg in either group. These results demonstrate the importance of the B cell for the induction of cytokine-mediated suppression associated with low doses of Ag. PMID- 11254703 TI - GIF inhibits Th effector generation by acting on antigen-presenting B cells. AB - Glycosylation-inhibiting factor (GIF) is a 13-kDa cytokine secreted from T cells. Administration of bioactive recombinant GIF inhibits IgG1 and IgE Ab responses in vivo. Treatment of B cells with the cytokine reduces the secretion of IgG1 and IgE induced by LPS and IL-4. To examine the effect on cognate T-B interaction, GIF was added to low-density B cells from MD4 transgenic (Tg) mice, which express B cell receptor specific for hen egg lysozyme (HEL). The B cells were subsequently pulsed with HEL-OVA conjugate and cultured with OVA-specific naive CD4 T cells from DO11.10 Tg mice. Treatment of Ag-presenting B cells with GIF reduced expansion and IL-2 secretion of naive T cells and rendered them hyporesponsive to antigenic restimulation, resulting in 50--95% reduction of IL-4 and IFN-gamma secretion upon restimulation with Ag. GIF dramatically inhibited Th effector generation when it was added to B cells before pulsing with HEL-OVA, whereas it showed little to no effect when added after B cells were pulsed with Ag. GIF was more effective when B cells from MD4 Tg mice were pulsed with HEL-OVA than when they were pulsed with OVA. This cytokine did not affect Th effector generation when B cells or irradiated splenocytes pulsed with OVA(323--339) peptide stimulated naive DO11.10 T cells. Confocal microscopy revealed that GIF inhibited internalization of HEL by B cells from MD4 Tg mice. Therefore, the cytokine may regulate early steps of Ag presentation involving B cell receptors to diminish Th effector generation from naive CD4 T cells. PMID- 11254704 TI - IFN-gamma/TNF-alpha synergism as the final effector in autoimmune diabetes: a key role for STAT1/IFN regulatory factor-1 pathway in pancreatic beta cell death. AB - Fas ligand (FasL), perforin, TNF-alpha, IL-1, and NO have been considered as effector molecule(s) leading to beta cell death in autoimmune diabetes. However, the real culprit(s) in beta cell destruction have long been elusive, despite intense investigation. We and others have demonstrated that FasL is not a major effector molecule in autoimmune diabetes, and previous inability to transfer diabetes to Fas-deficient nonobese diabetic (NOD)-lpr mice was due to constitutive FasL expression on lymphocytes from these mice. Here, we identified IFN-gamma/TNF-alpha synergism as the final effector molecules in autoimmune diabetes of NOD mice. A combination of IFN-gamma and TNF-alpha, but neither cytokine alone, induced classical caspase-dependent apoptosis in insulinoma and pancreatic islet cells. IFN-gamma treatment conferred susceptibility to TNF-alpha induced apoptosis on otherwise resistant insulinoma cells by STAT1 activation followed by IFN regulatory factor (IRF)-1 induction. IRF-1 played a central role in IFN-gamma/TNF-alpha-induced cytotoxicity because inhibition of IRF-1 induction by antisense oligonucleotides blocked IFN-gamma/TNF-alpha-induced cytotoxicity, and transfection of IRF-1 rendered insulinoma cells susceptible to TNF-alpha induced cytotoxicity. STAT1 and IRF-1 were expressed in pancreatic islets of diabetic NOD mice and colocalized with apoptotic cells. Moreover, anti-TNF-alpha Ab inhibited the development of diabetes after adoptive transfer. Taken together, our results indicate that IFN-gamma/TNF-alpha synergism is responsible for autoimmune diabetes in vivo as well as beta cell apoptosis in vitro and suggest a novel signal transduction in IFN-gamma/TNF-alpha synergism that may have relevance in other autoimmune diseases and synergistic anti-tumor effects of the two cytokines. PMID- 11254705 TI - Nonobese diabetic mice display elevated levels of class II-associated invariant chain peptide associated with I-Ag7 on the cell surface. AB - Peptide presentation by MHC class II molecules plays a pivotal role in determining the peripheral T cell repertoire as a result of both positive and negative selection in the thymus. Homozygous I-A(g7) expression imparts susceptibility to autoimmune diabetes in the nonobese diabetic mouse, and recently, it has been proposed that this arises from ineffectual peptide binding. Following biosynthesis, class II molecules are complexed with class II-associated invariant chain peptides (CLIP), which remain associated until displaced by Ag derived peptides. If I-A(g7) is a poor peptide binder, then this may result in continued occupation by CLIP to the point of translocation to the cell surface. To test this hypothesis we generated affinity-purified polyclonal antisera that recognized murine CLIP bound to class II molecules in an allele-independent fashion. We have found abnormally high natural levels of cell surface class II occupancy by CLIP on nonobese diabetic splenic B cells. Experiments using I-A transfected M12.C3 cells showed that I-A(g7) alone was associated with elevated levels of CLIP, suggesting that this was determined solely by the amino acid sequence of the class II molecule. These results indicated that an intrinsic property of I-A(g7) would affect both the quantity and the repertoire of self peptides presented during thymic selection. PMID- 11254706 TI - Suppression of Il-12 transcription in macrophages following Fc gamma receptor ligation. AB - Ligating Fc gamma R on macrophages results in suppression of IL-12 production. We show that Fc gamma R ligation selectively down-regulates IL-12 p40 and p35 gene expression at the level of transcription. The region responsive to this inhibition maps to the Ets site of the p40 promoter. PU.1, IFN consensus sequence binding protein, and c-REL: form a complex on this element upon macrophage activation. Receptor ligation abolishes the binding of this PU.1-containing activation complex, and abrogates p40 transcription. A dominant-negative construct of PU.1 diminishes IL-12 p40 promoter activity and endogenous IL-12 p40 protein secretion. Thus, the specificity of IL-12 down-regulation following receptor ligation lies in the inhibition of binding of a PU.1-containing complex to the Ets site of the IL-12 promoter. These findings provide evidence demonstrating for the first time the importance of PU.1 in the transcriptional regulation of IL-12 gene expression. PMID- 11254707 TI - STAT6 mediates eotaxin-1 expression in IL-4 or TNF-alpha-induced fibroblasts. AB - Eosinophils are attracted to sites of allergic inflammation by a number of chemoattractants including eotaxin-1. This chemokine can be secreted from epithelial cells and fibroblasts after IL-4 and TNF-alpha stimulation in a synergistic fashion. TNF-alpha activated gene expression at the transcriptional level in a STAT6-dependent manner, because: 1) eotaxin-1 promoter luciferase constructs were TNF-alpha inducible in STAT6-defective HEK293 cells only on cotransfection of STAT6 expression vector, an effect that was partially mediated by activation-induced binding of NF-kappa B proteins to a composite STAT6/NF kappa B element; 2) reporter constructs defective in STAT6 DNA binding did not respond to TNF-alpha stimulation; 3) eotaxin-1 protein secretion was detected only in STAT6-transfected HEK293 cell supernatants on TNF-alpha treatment; and 4) a trans-dominant negative STAT6 protein inhibited TNF-alpha-induced eotaxin-1 secretion in primary fibroblasts. TNF-alpha inducibility of the IL-8 and monocyte chemoattractant protein-1 genes was not dependent on STAT6 expression in the same experimental systems. The inducing effect of IL-4 and IL-13 was also mediated by STAT6. The synergistic effect of IL-4 and TNF-alpha observed at the RNA and the protein level was not seen at the promoter level. The data demonstrate that both IL-4 and TNF-alpha induce eotaxin-1 expression at the level of transcription via a STAT6-mediated pathway. PMID- 11254708 TI - NF-kappa B and STAT5 play important roles in the regulation of mouse Toll-like receptor 2 gene expression. AB - Toll-like receptor 2 (TLR2) is involved in the innate immunity by recognizing various bacterial components. We have previously reported that TLR2 gene expression is rapidly induced by LPS or inflammatory cytokines in macrophages, and by TCR engagement or IL-2/IL-15 stimulation in T cells. Here, to investigate the mechanisms governing TLR2 transcription, we cloned the 5' upstream region of the mouse TLR2 (mTLR2) gene and mapped its transcriptional start site. The 5' upstream region of the mTLR2 gene contains two NF-kappa B, two CCAAT/enhancer binding protein, one cAMP response element-binding protein, and one STAT consensus sequences. In mouse macrophage cell lines, deletion of both NF-kappa B sites caused the complete loss of mTLR2 promoter responsiveness to TNF-alpha. NF kappa B sites were also important but not absolutely necessary for LPS-mediated mTLR2 promoter activation. In T cell lines, mTLR2 responsiveness to IL-15 was abrogated by the 3' NF-kappa B mutation, whereas 5' NF-kappa B showed no functional significance. The STAT binding site also seemed to contribute, as the deletion of this sequence significantly reduced the IL-15-mediated mTLR2 promoter activation. EMSAs confirmed nuclear protein binding to both NF-kappa B sites in macrophages following LPS and TNF-alpha stimulation and to the 3' NF-kappa B site in T cells after IL-15 treatment. Thus, NF-kappa B activation is important but differently involved in the regulation of mTLR2 gene expression in macrophages and T cells following LPS or cytokine stimulation. PMID- 11254709 TI - The TCR repertoire of an immunodominant CD8+ T lymphocyte population. AB - The TCR repertoire of an epitope-specific CD8(+) T cell population remains poorly characterized. To determine the breadth of the TCR repertoire of a CD8(+) T cell population that recognizes a dominant epitope of the AIDS virus, the CD8(+) T cells recognizing the tetrameric Mamu-A*01/p11C(,CM) complex were isolated from simian immunodeficiency virus (SIV)-infected Mamu-A*01(+) rhesus monkeys. This CD8(+) T cell population exhibited selected usage of TCR V beta families and complementarity-determining region 3 (CDR3) segments. Although the epitope specific CD8(+) T cell response was clearly polyclonal, a dominance of selected V beta(+) cell subpopulations and clones was seen in the TCR repertoire. Interestingly, some of the selected V beta(+) cell subpopulations and clones maintained their dominance in the TCR repertoire over time after infection with SIV of macaques. Other V beta(+) cell subpopulations declined over time in their relative representation and were replaced by newly evolving clones that became dominant. The present study provides molecular evidence indicating that the TCR repertoire shaped by a single viral epitope is dominated at any point in time by selected V beta(+) cell subpopulations and clones and suggests that dominant V beta(+) cell subpopulations and clones can either be stable or evolve during a chronic infection. PMID- 11254710 TI - Differential expression of Fas ligand in Th1 and Th2 cells is regulated by early growth response gene and NF-AT family members. AB - Inducible expression of Fas ligand (CD95 ligand) by activated T cells and the resulting apoptosis of CD95-bearing cells is a critical component of peripheral T cell homeostasis and cytotoxic effector mechanisms. Transcriptional control of the expression of Fas ligand has been attributed to a number of factors, including early growth response gene 2 (Egr2), Egr3, Sp1, and NF-AT, although a direct contribution of NF-AT is controversial. The present study confirms a role for Egr factors and indicates that NF-AT is essential for optimal expression of murine Fas ligand through a direct interaction with an NF-AT consensus element. The role of these factors was further defined by studying the differential expression of Fas ligand in Th1 and Th2 lines derived from DO11.10 TCR transgenic mice. EMSA analyses of a composite Egr/NF-AT site showed recruitment of Sp1 to this site in Th2 cells, but not in Th1 cells. Furthermore, gel-shift analyses demonstrated the binding of Egr1, 2, and 3 in Th2 cells and Egr1 and 2, but not Egr3 in Th1 cells at a known Egr site. Northern analysis corroborated the lack of Egr3 in Th1 cells. Differential usage of these transcription factors by Th1 and Th2 cells suggests a potential mechanism underlying the differential expression of Fas ligand by distinct T cell lineages. PMID- 11254711 TI - Cysteinylation of MHC class II ligands: peptide endocytosis and reduction within APC influences T cell recognition. AB - Peptides bind cell surface MHC class II proteins to yield complexes capable of activating CD4(+) T cells. By contrast, protein Ags require internalization and processing by APC before functional presentation. Here, T cell recognition of a short peptide in the context of class II proteins occurred only after delivery of this ligand to mature endosomal/lysosomal compartments within APC. Functional and biochemical studies revealed that a central cysteine within the peptide was cysteinylated, perturbing T cell recognition of this epitope. Internalization and processing of the modified epitope by APC, was required to restore T cell recognition. Peptide cysteinylation and reduction could occur rapidly and reversibly before MHC binding. Cysteinylation did not disrupt peptide binding to class II molecules, rather the modified peptide displayed an enhanced affinity for MHC at neutral pH. However, once the peptide was bound to class II proteins, oxidation or reduction of cysteine residues was severely limited. Cysteinylation has been shown to radically influence T cell responses to MHC class I ligands. The ability of professional APC to reductively cleave this peptide modification presumably evolved to circumvent a similar problem in MHC class II ligand recognition. PMID- 11254713 TI - HIV-1 Tat inhibits IL-2 gene transcription through qualitative and quantitative alterations of the cooperative Rel/AP1 complex bound to the CD28RE/AP1 composite element of the IL-2 promoter. AB - Dysregulation of cytokine secretion plays an important role in AIDS pathogenesis. Here, we demonstrate that expression of HIV-1 Tat protein in Jurkat cells induces a severe impairment of IL-2 but not TNF gene transcription. Interestingly, this inhibition correlates with the effect of the viral protein on the transactivation of the CD28RE/AP1 composite element (-164/-154), but not with that observed on the NFAT/AP1 site of the IL-2 gene promoter, neither with the effect on NF-kappa B- nor AP1-independent binding sites. Endogenous expression of Tat induced a decrease in the amount of the specific protein complex bound to the CD28RE/AP1 probe after PMA plus calcium ionophore stimulation. This effect was accompanied by qualitative alterations of the AP1 complex. Thus, in wild-type Jurkat cells, c jun was absent from the complex, whereas in Tat-expressing cells, c-jun was increasingly recruited overtime. By contrast, similar amounts of c-rel and a small amount of NFAT1 were detected both in wild type and in Jurkat Tat(+) cells. Furthermore, Tat not only induced the participation of c-jun in the cooperative complex but also a decrease in its transactivation activity alone or in combination with c-rel. Thus, the interaction of Tat with the components of this rel/AP1 cooperative complex seems to induce quantitative and qualitative alterations of this complex as activation progresses, resulting in a decrease of IL-2 gene transcription. Altogether our results suggest the existence of tuned mechanisms that allow the viral protein to specifically affect cooperative interactions between transcription factors. PMID- 11254712 TI - NF-kappa B p50-dependent in vivo footprints at Ig S gamma 3 DNA are correlated with mu-->gamma 3 switch recombination. AB - NF-kappa B has been demonstrated to play critical roles in multiple aspects of immune responses including Ig H chain isotype switching. To better define the specific roles the p50 subunit of NF-kappa B plays in mu-->gamma 3 switch recombination (SR), we systematically evaluated p50-deficient B cells for activities that are strongly correlated with SR. B cell activation with LPS plus anti-IgD-dextran plus IL-5 plus IL-4 plus TGF-beta produced normal levels of proliferation and gamma3 germline transcripts in p50-deficient B cells, but mu- >gamma 3 SR was impaired. In vitro binding studies previously showed that NF kappa B p50 homodimer binds the switch nuclear B-site protein (SNIP) of the S gamma 3 tandem repeat. Ligation-mediated PCR in vivo footprint analysis demonstrates that the region spanning the SNIP and switch nuclear A-site protein (SNAP) binding sites of the S gamma 3 region are contacted by protein in normal resting splenic B cells. B cells that are homozygous for the targeted disruption of the gene encoding p50 (-/-) show strong aberrant footprints, whereas heterozygous cells (+/-) reveal a partial effect in S gamma 3 DNA. These studies provide evidence of nucleoprotein interactions at switch DNA in vivo and suggest a direct interaction of p50 with S gamma 3 DNA that is strongly correlated with SR competence. PMID- 11254714 TI - Novel androgen-dependent promoters direct expression of the C4b-binding protein alpha-chain gene in epididymis. AB - C4b-binding protein (C4BP) is a large plasma protein composed of seven alpha chains and one beta-chain and is involved in the fluid phase regulation of the classical pathway of the complement system. Complement inhibitory activity is located in the alpha-chain, and its mRNA has been detected only in liver to date. Here, we have isolated cDNA clones encoding the alpha-chain of guinea pig C4BP (C4BP alpha) and have demonstrated significant C4BP alpha mRNA expression in epididymis as well as liver. The level of C4BP alpha transcripts increased in the epididymis after birth, while it remained constant in the liver. C4BP alpha mRNA was also detected in the normal murine epididymis at a significant level, but it decreased drastically after castration, suggesting that epididymal expression of the C4BP alpha gene is regulated by androgen. Gene analysis of guinea pig C4BP alpha indicated that liver and epididymis C4BP alpha mRNA share the coding region and 3'-untranslated region, but are transcribed from independent promoters on a single-copy gene. Two novel epididymis-specific promoters were identified in the region corresponding to the first intron of liver transcripts. The binding motif for hepatocyte NF-1 occurs in the promoter used for transcription of liver C4BP alpha, whereas androgen-responsive elements occur in both promoters used in the epididymis. These findings present a novel link between complement regulators and reproduction. Furthermore, variation in the 5'-untranslated regions, arising from alternative splicing of the newly identified exons, is demonstrable in the guinea pig C4BP alpha transcripts. PMID- 11254715 TI - Regulation of transcriptional activity of the murine CD40 ligand promoter in response to signals through TCR and the costimulatory molecules CD28 and CD2. AB - We have analyzed the murine CD40 ligand promoter with regard to stimulation of transcriptional activity in Jurkat T cells after signaling via the TCR and the costimulatory molecules CD28 and CD2. TCR engagement was necessary for the induction of transcriptional activity from the CD40 ligand promoter, and costimulation through either CD28 or CD2 further increased the activity. Analysis of promoter deletants showed that the DNA elements needed for transcriptional activity induced by costimulatory molecules were located within two regions containing previously identified transcription factor NFAT sites. Further studies of the proximal NFAT site showed that it was not dependent on AP-1 binding for transcriptional activity induced by costimulation through CD28. Instead, a region between the TATA box and the proximal NFAT site was shown to bind proteins of the early growth response family and to contribute to NFAT-mediated transcriptional activation. PMID- 11254716 TI - Identification of a novel lipopolysaccharide-inducible gene with key features of both A kinase anchor proteins and chs1/beige proteins. AB - Mutations in chs1/beige result in a deficiency in intracellular transport of vesicles that leads to a generalized immunodeficiency in mice and humans. The function of NK cells, CTL, and granulocytes is impaired by these mutations, indicating that polarized trafficking of vesicles is controlled by CHS1/beige proteins. However, a molecular explanation for this defect has not been identified. Here we describe a novel gene with orthologues in mice, humans, and flies that contains key features of both chs1/beige and A kinase anchor genes. We designate this novel gene lba for LPS-responsive, beige-like anchor gene. Expression of lba is induced after LPS stimulation of B cells and macrophages. In addition, lba is expressed in many other tissues in the body and has three distinct mRNA isoforms that are differentially expressed in various tissues. Strikingly, LBA-green-fluorescent protein (GFP) fusion proteins are localized to vesicles after LPS stimulation. Confocal microscopy indicates this protein is colocalized with the trans-Golgi complex and some lysosomes. Further analysis by immunoelectron microscopy demonstrates that LBA-GFP fusion protein can localize to endoplasmic reticulum, plasma membrane, and endocytosis vesicles in addition to the trans-Golgi complex and lysosomes. We hypothesize that LBA/CHS1/BG proteins function in polarized vesicle trafficking by guiding intracellular vesicles to activated receptor complexes and thus facilitate polarized secretion and/or membrane deposition of immune effector molecules. PMID- 11254717 TI - Antigen-specific Th1 but not Th2 cells provide protection from lethal Trypanosoma cruzi infection in mice. AB - Infection with Trypanosoma cruzi results in the development of both type 1 and type 2 patterns of cytokine responses during acute and chronic stages of infection. To investigate the role of Th1 and Th2 subsets of CD4(+) T cells in determining the outcome of T. cruzi infection in mice, we have developed T. cruzi clones that express OVA and have used OVA-specific TCR-transgenic T cells to generate OVA-specific Th1 and Th2 cells. BALB/c mice receiving 10(7) OVA-specific Th1 cells and then challenged with OVA-expressing T. cruzi G-OVA.GPI showed significantly lower parasitemia and increased survival in comparison to mice that received no cells. In contrast, recipients of OVA-specific Th2 cells developed higher parasitemias, exhibited higher tissue parasitism and inflammation, and had higher mortality than recipients of Th1 cells after infection with T. cruzi G OVA.GPI. Mice receiving a mixture of both Th1 and Th2 OVA-specific cells also were not protected from lethal challenge. The protective effect of the OVA specific Th1 cells was OVA dependent as shown by the fact that transfer of OVA specific Th1 or Th2 cells failed to alter the course of infection or disease in mice challenged with wild-type T. cruzi. Immunohistochemical analysis of OVA specific Th1 and Th2 cells at 4, 15, and 30 days postinfection revealed the persistence and expansion of these cells in mice challenged with T. cruzi G OVA.GPI but not in mice infected with wild-type T. cruzi. We conclude that transfer of Ag-specific Th1 cells but not Th2 cells protect mice from a lethal infection with T. cruzi. PMID- 11254718 TI - Depletion of alveolar macrophages exerts protective effects in pulmonary tuberculosis in mice. AB - Mycobacterium tuberculosis bacilli are intracellular organisms that reside in phagosomes of alveolar macrophages (AMs). To determine the in vivo role of AM depletion in host defense against M. tuberculosis infection, mice with pulmonary tuberculosis induced by intranasal administration of virulent M. tuberculosis were treated intranasally with either liposome-encapsulated dichloromethylene diphosphonate (AM(-) mice), liposomes, or saline (AM(+) mice). AM(-) mice were completely protected against lethality, which was associated with a reduced outgrowth of mycobacteria in lungs and liver, and a polarized production of type 1 cytokines in lung tissue, and by splenocytes stimulated ex vivo. AM(-) mice displayed deficient granuloma formation, but were more capable of attraction and activation of T cells into the lung and had increased numbers of pulmonary polymorphonuclear cells. These data demonstrate that depletion of AMs is protective during pulmonary tuberculosis. PMID- 11254719 TI - Cancer vaccine design: a novel bacterial adjuvant for peptide-specific CTL induction. AB - The recent identification of tumor Ags as potential vaccines has prompted the search for efficient adjuvants and delivery systems, especially in the case of peptide-based vaccination protocols. Here, we investigated the adjuvant potential of the recombinant 40-kDa outer membrane protein of Klebsiella pneumoniae (P40) for specific CTL induction. We studied the CTL response induced in HLA A*0201/K(b) transgenic mice immunized with peptides derived from two melanoma associated differentiation Ags, the HLA-A*0201-restricted decapeptide Melan-A(26- 35) substituted at position 2 and the K(b)-restricted tyrosinase-related protein 2(181--188) T cell epitope. We found that both peptides are able to generate a specific CTL response when mixed with the protein in the absence of conventional adjuvant. This CTL response is a function of the amount of P40 used for immunization. Moreover, the CTL response generated against the tyrosinase-related protein 2(181-188) peptide in presence of P40 is associated with tumor protection in two different experimental models and is independent of the presence of CD4(+) T lymphocytes. Thus, the recombinant bacterial protein P40 functions as a potent immunological adjuvant for specific CTL induction. PMID- 11254720 TI - Toll-like receptor 4 mediates intracellular signaling without TNF-alpha release in response to Cryptococcus neoformans polysaccharide capsule. AB - Toll-like receptors (TLR) 2 and 4 are cell surface receptors that in association with CD14 enable phagocytic inflammatory responses to a variety of microbial products. Activation via these receptors triggers signaling cascades, resulting in nuclear translocation of NF-kappa B and a proinflammatory response including TNF-alpha production. We investigated whether TLRs participate in the host response to Cryptococcus neoformans glucuronoxylomannan (GXM), the major capsular polysaccharide of this fungus. Chinese hamster ovary fibroblasts transfected with human TLR2, TLR4, and/or CD14 bound fluorescently labeled GXM. The transfected Chinese hamster ovary cells were challenged with GXM, and activation of an NF kappa B-dependent reporter construct was evaluated. Activation was observed in cells transfected with both CD14 and TLR4. GXM also stimulated nuclear NF-kappa B translocation in PBMC and RAW 264.7 cells. However, stimulation of these cells with GXM resulted in neither TNF-alpha secretion nor activation of the extracellular signal-regulated kinase 1/2, p38, and stress-activated protein kinase/c-Jun N-terminal kinase mitogen-activated protein kinase pathways. These findings suggest that TLRs, in conjunction with CD14, function as pattern recognition receptors for GXM. Furthermore, whereas GXM stimulates cells to translocate NF-kappa B to the nucleus, it does not induce activation of mitogen activated protein kinase pathways or release of TNF-alpha. Taken together, these observations suggest a novel scenario whereby GXM stimulates cells via CD14 and TLR4, resulting in an incomplete activation of pathways necessary for TNF-alpha production. PMID- 11254721 TI - Diversity of epitope and cytokine profiles for primary and secondary influenza a virus-specific CD8+ T cell responses. AB - Screening with the flow cytometric IFN-gamma assay has led to the identification of a new immunogenic peptide (SSYRRPVGI) [corrected] from the influenza PB1 polymerase (PB1(703--711)) and a mimotope (ISPLMVAYM) from the PB2 polymerase (PB2(198--206)). CD8(+) T cells specific for K(b)PB1(703) make both IFN-gamma and TNF-alpha following stimulation with both peptides. The CD8(+) K(b)PB1(703)(+) population kills PB2(198)-pulsed targets, but cell lines stimulated with PB2(198) neither bind the K(b)PB1(703) tetramer nor become CTL. This CD8(+)K(b)PB1(703)(+) population is prominent in the primary response to an H3N2 virus, although it is much less obvious following secondary challenge of H1N1-primed mice. Even so, we can now account for >40% of the CD8(+) T cells in a primary influenza pneumonia and >85% of those present after H3N2 --> H1N1 challenge. Profiles of IFN-gamma and TNF-alpha staining following in vitro stimulation have been traced for the four most prominent influenza peptides through primary and secondary responses into long-term memory. The D(b)NP(366) epitope that is immunodominant after the H3N2 --> H1N1 challenge shows the lowest frequencies of CD8(+) IFN-gamma(+)TNF alpha(+) cells for >6 wk, and the intensity of IFN-gamma staining is also low for the first 3 wk. By 11 wk, however, the IFN-gamma/TNF-alpha profiles look to be similar for all four epitopes. At least by the criterion of cytokine production, there is considerable epitope-related functional diversity in the influenza virus specific CD8(+) T cell response. The results for the K(b)PB1(703) epitope and the PB2(198) mimotope also provide a cautionary tale for those using the cytokine staining approach to identity antigenic peptides. PMID- 11254722 TI - CXC chemokine receptor-2 ligands are required for neutrophil-mediated host defense in experimental brain abscesses. AB - We have developed a mouse brain abscess model by using Staphylococcus aureus, one of the main etiologic agents of brain abscesses in humans. Direct damage to the blood-brain barrier was observed from 24 h to 7 days after S. aureus exposure as demonstrated by the accumulation of serum IgG in the brain parenchyma. Evaluation of brain abscesses by immunohistochemistry and flow cytometry revealed a prominent neutrophil infiltrate. To address the importance of neutrophils in the early containment of S. aureus infection in the brain, mice were transiently depleted of neutrophils before implantation of bacteria-laden beads. Neutrophil depleted animals consistently demonstrated more severe brain abscesses and higher CNS bacterial burdens compared with control animals. S. aureus led to the induction of numerous chemokines in the brain, including macrophage-inflammatory protein (MIP)-1alpha/CCL3, MIP-1beta/CCL4, MIP-2/CXCL1, monocyte chemoattractant protein-1/CCL2, and TCA-3/CCL1, within 6 h after bacterial exposure. These chemokines also were expressed by both primary cultures of neonatal mouse microglia and astrocytes exposed to heat-inactivated S. aureus in vitro. Because neutrophils constitute the majority of the cellular infiltrate in early brain abscess development, subsequent analysis focused on MIP-2 and KC/CXCL1, two neutrophil-attracting CXC chemokines. Both MIP-2 and KC protein levels were significantly elevated in the brain after S. aureus exposure. Neutrophil extravasation into the brain parenchyma was impaired in CXCR2 knockout mice and was associated with increased bacterial burdens. These studies demonstrate the importance of the CXCR2 ligands MIP-2 and KC and neutrophils in the acute host response to S. aureus in the brain. PMID- 11254723 TI - The alpha 4 beta 1 (very late antigen (VLA)-4, CD49d/CD29) and alpha 5 beta 1 (VLA-5, CD49e/CD29) integrins mediate beta 2 (CD11/CD18) integrin-independent neutrophil recruitment to endotoxin-induced lung inflammation. AB - The beta(2) integrin cell adhesion molecules (CAM) mediate polymorphonuclear leukocyte (PMNL) emigration in most inflamed tissues, but, in the lung, other yet to be identified CAMs appear to be involved. In Lewis rats, the intratracheal injection of Escherichia coli-LPS induced acute (6-h) PMNL accumulation in the lung parenchyma (280 x 10(6) by myeloperoxidase assay; PBS control = 35 x 10(6)) and bronchoalveolar lavage fluid (BALF = 27 x 10(6); PBS = 0.1 x 10(6)). Parenchymal accumulation was not inhibited by a blocking Ab to beta(2) integrins and only minimally inhibited (20.5%; p < 0.05) in BALF. We examined the role of alpha(4)beta(1) and alpha(5)beta(1) integrins and of selectins in this PMNL recruitment. Treatment with mAbs to alpha(4)beta(1) or alpha(5)beta(1), even in combination, had no effect on PMNL accumulation induced by intratracheal LPS. However, anti-alpha(4) combined with anti-beta(2) mAbs inhibited PMNL recruitment to the parenchyma by 56% (p < 0.001) and to BALF by 58% (p < 0.01). The addition of anti-alpha(5) mAb to beta(2) plus alpha(4) blockade inhibited PMNL accumulation further (by 79%; p < 0.05). In contrast, blockade of L-, P-, and E selectins in combination or together with beta(2), alpha(4), and alpha(5) integrins had no effect. LPS-induced BALF protein accumulation was not inhibited by treatment with anti-beta(2) plus alpha(4) mAbs, but was prevented when alpha(5)beta(1) was also blocked. Thus, while selectins appear to play no role, alpha(4)beta(1) and alpha(5)beta(1) function as major alternate CAMs to the beta(2) integrins in mediating PMNL migration to lung and to pulmonary vascular and epithelial permeability. PMID- 11254724 TI - Human leukocyte subpopulations from inflamed gut bind to joint vasculature using distinct sets of adhesion molecules. AB - Reactive arthritis can be triggered by inflammatory bowel diseases. We hypothesized that migration of mucosal immune cells from inflamed gut to joints could contribute to the development of reactive arthritis. Here we isolated gut derived leukocytes from patients with Crohn's disease and ulcerative colitis. Using function-blocking mAbs and in vitro frozen section adhesion assays we studied whether these cells bind to synovial vessels and which molecules mediate the interaction. The results showed that mucosal leukocytes from inflammatory bowel diseased gut bind well to venules in synovial membrane. Small intestinal lymphocytes adhered to synovial vessels using multiple homing receptors and their corresponding endothelial ligands (CD18-ICAM-1, alpha(4)beta(7)/alpha(4)beta(1) integrin-VCAM-1, L-selectin-peripheral lymph node addressins, and CD44). Of these, only ICAM-1 significantly supported binding of immunoblasts. In contrast, P-selectin glycoprotein ligand-1-P-selectin interaction accounted for practically all synovial adherence of mucosal macrophages. In addition, blocking of vascular adhesion protein-1 significantly inhibited binding of all these leukocyte subsets to joint vessels. We conclude that different leukocyte populations derived from inflamed gut bind avidly to synovial vessels using distinct repertoire of adhesion molecules, suggesting that their recirculation may contribute to the development of reactive arthritis in inflammatory bowel diseases. PMID- 11254725 TI - Granulocyte macrophage-colony-stimulating factor mRNA is stabilized in airway eosinophils and peripheral blood eosinophils activated by TNF-alpha plus fibronectin. AB - Airway eosinophils show prolonged in vitro survival compared with peripheral blood eosinophils (PBEos). Recent studies have shown that autocrine production and release of GM-CSF is responsible for enhanced survival, but the mechanisms controlling cytokine production remain obscure. We compared GM-CSF mRNA decay in eosinophils from bronchoalveolar lavage (BALEos) after allergen challenge or from PBEos. BALEos showed prolonged survival in vitro (60% at 4 days) and expressed GM CSF mRNA. The enhanced survival of BALEos was 75% inhibited at 6 days by neutralizing anti-GM-CSF Ab. Based on transfection studies, GM-CSF mRNA was 2.5 times more stable in BALEos than in control PBEos. Treatment of PBEos with fibronectin and TNF-alpha increased their in vitro survival, GM-CSF mRNA expression, and GM-CSF mRNA stability to a comparable level as seen in BALEos. These data suggest that TNF-alpha plus fibronectin may increase eosinophil survival in vivo by controlling GM-CSF production at a posttranscriptional level. PMID- 11254726 TI - Evidence for a role for SAM68 in the responses of human neutrophils to ligation of CD32 and to monosodium urate crystals. AB - SAM68 (Src-associated in mitosis 68 kDa) is a member of the signal transduction of activator RNA novel gene family coding for proteins postulated to be involved in signal transduction and activation of RNA. It has been implicated through its phosphorylation status in the control of the transition from the G(1) to the S phases during mitosis. However, the implication and role of SAM68 in nonproliferative cells are unknown. The present study was initiated to examine the role of SAM68 in the phagocytic responses of the terminally differentiated human neutrophils. The results obtained show that SAM68 is present in human neutrophils and that it is tyrosine phosphorylated in response to stimulation by monosodium urate crystals or by ligation of CD32. Stimulation of neutrophils by these agonists decreases the association of SAM68 with Sepharose-conjugated poly U beads. Additionally, the amount of immunoprecipitable SAM68 was modulated differentially after stimulation by monosodium urate crystals or by CD32 engagement indicating that the posttranslational modifications and/or protein associations of SAM68 induced by these two agonists differed. The results of this study provide evidence for an involvement of SAM68 in signal transduction by phagocytic agonists in human neutrophils and indicate that SAM68 may play a role in linking the early events of signal transduction to the posttranscriptional modulation of RNA. PMID- 11254728 TI - Two proteins modulating transendothelial migration of leukocytes recognize novel carboxylated glycans on endothelial cells. AB - We recently showed that a class of novel carboxylated N:-glycans was constitutively expressed on endothelial cells. Activated, but not resting, neutrophils expressed binding sites for the novel glycans. We also showed that a mAb against these novel glycans (mAbGB3.1) inhibited leukocyte extravasation in a murine model of peritoneal inflammation. To identify molecules that mediated these interactions, we isolated binding proteins from bovine lung by their differential affinity for carboxylated or neutralized glycans. Two leukocyte calcium-binding proteins that bound in a carboxylate-dependent manner were identified as S100A8 and annexin I. An intact N terminus of annexin I and heteromeric assembly of S100A8 with S100A9 (another member of the S100 family) appeared necessary for this interaction. A mAb to S100A9 blocked neutrophil binding to immobilized carboxylated glycans. Purified human S100A8/A9 complex and recombinant human annexin I showed carboxylate-dependent binding to immobilized bovine lung carboxylated glycans and recognized a subset of mannose-labeled endothelial glycoproteins immunoprecipitated by mAbGB3.1. Saturable binding of S100A8/A9 complex to endothelial cells was also blocked by mAbGB3.1. These results suggest that the carboxylated glycans play important roles in leukocyte trafficking by interacting with proteins known to modulate extravasation. PMID- 11254727 TI - An increase in circulating mast cell colony-forming cells in asthma. AB - We compared a potential to generate mast cells among various sources of CD34(+) peripheral blood (PB) cells in the presence of stem cell factor (SCF) with or without thrombopoietin (TPO), using a serum-deprived liquid culture system. From the time course of relative numbers of tryptase-positive and chymase-positive cells in the cultured cells grown by CD34(+) PB cells of nonasthmatic healthy individuals treated with G-CSF, TPO appears to potentiate the SCF-dependent growth of mast cells without influencing the differentiation into mast cell lineage. CD34(+) PB cells from asthmatic patients in a stable condition generated significantly more mast cells under stimulation with SCF alone or SCF+TPO at 6 wk of culture than did steady-state CD34(+) PB cells of normal controls. Single-cell culture studies showed a substantial difference in the number of SCF-responsive or SCF+TPO-responsive mast cell progenitors in CD34(+) PB cells between the two groups. In the presence of TPO, CD34(+) PB cells from asthmatic children could respond to a suboptimal concentration of SCF to a greater extent, compared with the values obtained by those of normal controls. Six-week cultured mast cells of asthmatic subjects had maturation properties (intracellular histamine content and tryptase/chymase enzymatic activities) similar to those derived from mobilized CD34(+) PB cells of nonasthmatic subjects. An increase in a potential of circulating hemopoietic progenitors to differentiate into mast cell lineage may contribute to the recruitment of mast cells toward sites of asthmatic mucosal inflammation. PMID- 11254729 TI - The expression of prostaglandin E receptors EP2 and EP4 and their different regulation by lipopolysaccharide in C3H/HeN peritoneal macrophages. AB - The expression and regulation of the PGE receptors, EP(2) and EP(4), both of which are coupled to the stimulation of adenylate cyclase, were examined in peritoneal resident macrophages from C3H/HeN mice. mRNA expression of EP(4) but not EP(2) was found in nonstimulated cells, but the latter was induced by medium change alone, and this induction was augmented by LPS. mRNA expression of EP(4) was down-regulated by LPS but not by medium change. PGE(2) increased the cAMP content of both LPS-treated and nontreated cells. ONO-604, an EP(4) agonist, also increased cAMP content in nonstimulated cells and in cells treated with LPS for 3 h, but not for 6 h. Butaprost, an EP(2) agonist, was effective only in the cells treated with LPS for 6 h. The inhibitory effects of ONO-604 on TNF-alpha and IL 12 production were equipotent with PGE(2) at any time point, but the inhibitory effects of butaprost were only seen from 14 h after stimulation. PGE(2) or dibutyryl cAMP alone, but not butaprost, reduced EP(4) expression, and indomethacin reversed the LPS-induced down-regulation of EP(4), indicating that the down-regulation of EP(4) is mediated by LPS-induced PG synthesis and EP(4) activation. Indeed, when we used C3H/HeJ (LPS-hyporesponsive) macrophages, such reduction in EP(4) expression was found in the cells treated with PGE(2) alone, but not in LPS-treated cells. In contrast, up-regulation of EP(2) expression was again observed in LPS-treated C3H/HeJ macrophages. These results suggest that EP(4) is involved mainly in the inhibition of cytokine release, and that the gene expression of EP(2) and EP(4) is differentially regulated during macrophage activation. PMID- 11254730 TI - Expression and characterization of the chemokine receptors CCR2 and CCR5 in mice. AB - The chemokine receptors CCR2 and CCR5 play important roles in the recruitment of monocytes/macrophages and T cells. To better understand the role of both receptors in murine models of inflammatory diseases and to recognize potential problems when correlating these data to humans, we have generated mAbs against murine CCR2 and CCR5. In mice CCR2 is homogeneously expressed on monocytes and on 2--15% of T cells, closely resembling the expression pattern in humans. In contrast to humans, murine NK cells are highly CCR5 positive. In addition, CCR5 is expressed on 3--10% of CD4 and 10--40% of CD8-positive T cells and is weakly detectable on monocytes. Using a model of immune complex nephritis, we examined the effects of inflammation on chemokine receptor expression and found a 10-fold enrichment of CCR5(+) and CCR2(+) T cells in the inflamed kidneys. The activity of various chemokines and the antagonistic properties of the mAbs were measured by ligand-induced internalization of CCR2 and CCR5 on primary leukocytes. The Ab MC-21 (anti-CCR2) reduced the activity of murine monocyte chemotactic protein 1 by 95%, whereas the Ab MC-68 (anti-CCR5) blocked over 99% of the macrophage inflammatory protein 1alpha and RANTES activity. MC-21 and MC-68 efficiently blocked the ligand binding to CCR2 and CCR5 with an IC(50) of 0.09 and 0.6--1.0 microg/ml, respectively. In good correlation to these in vitro data, MC-21 almost completely prevented the influx of monocytes in thioglycollate-induced peritonitis. Therefore, both Abs appear as useful reagents to further study the role of CCR2 and CCR5 in murine disease models. PMID- 11254731 TI - A comparison of mediators released or generated by IFN-gamma-treated human mast cells following aggregation of Fc gamma RI or Fc epsilon RI. AB - The high affinity receptor for IgG (Fc gamma RI, CD64) is expressed on human mast cells, where it is up-regulated by IFN-gamma and, thus, may allow mast cells to be recruited through IgG-dependent mechanisms in IFN-gamma-rich tissue inflammation. However, the mediators produced by human mast cells after aggregation of Fc gamma RI are incompletely described, and it is unknown whether these mediators are distinct from those produced after activation of human mast cells via Fc epsilon RI. Thus, we investigated the release of histamine and arachidonic acid metabolites and examined the chemokine and cytokine mRNA profiles of IFN-gamma-treated cultured human mast cells after Fc gamma RI or Fc epsilon RI aggregation. Aggregation of Fc gamma RI resulted in histamine release and PGD(2) and LTC(4) generation. These responses were qualitatively indistinguishable from responses stimulated via Fc epsilon RI. Aggregation of Fc epsilon RI or Fc gamma RI led to an induction or accumulation of 22 cytokine and chemokine mRNAs. Among them, seven cytokines (TNF-alpha, IL-1beta, IL-5, IL-6, IL 13, IL-1R antagonist, and GM-CSF) were significantly up-regulated via aggregation of Fc gamma RI compared with Fc epsilon RI. TNF-alpha mRNA data were confirmed by quantitative RT-PCR and ELISA. Furthermore, we confirmed histamine and TNF-alpha data using IFN-gamma-treated purified human lung mast cells. Thus, aggregation of Fc gamma RI on mast cells led to up-regulation and/or release of three important classes of mediators: biogenic amines, lipid mediators, and cytokines. Some cytokines, such as TNF-alpha, were released and generated to a greater degree after Fc gamma RI aggregation, suggesting that selected biologic responses of mast cells may be preferentially generated through Fc gamma RI in an IFN-gamma rich environment. PMID- 11254732 TI - Lipopolysaccharide activates Akt in human alveolar macrophages resulting in nuclear accumulation and transcriptional activity of beta-catenin. AB - Exposure of human alveolar macrophages to bacterial LPS results in activation of a number of signal transduction pathways. An early event after the alveolar macrophage comes in contact with LPS is activation of the phosphatidylinositol 3 kinase (PI 3-kinase). This study evaluates the downstream effects of that activation. We observed that LPS exposure results in phosphorylation of Akt (serine 473). We found this using both phosphorylation-specific Abs and also by in vivo phosphorylation with (32)P-loaded cells. AKT activation resulted in the phosphorylation-dependent inactivation of glycogen synthase kinase (GSK-3) (serine 21/9). We found that both of these events were linked to PI 3-kinase because the PI 3-kinase inhibitors, wortmannin and LY294002, inhibited LPS induced phosphorylation of both AKT and GSK-3. Inactivation of GSK-3 has been shown to reduce the ubiquitination of beta-catenin, resulting in nuclear accumulation and transcriptional activity of beta-catenin. Consistent with this, we found that LPS caused an increase in the amounts of PI 3-kinase-dependent nuclear beta-catenin in human alveolar macrophages and expression of genes that require nuclear beta-catenin for their activation. This is the first demonstration that LPS exposure activates AKT, inactivates GSK-3, and causes accumulation and transcriptional activity of beta-catenin in the nucleus of any cell, including alveolar macrophages. PMID- 11254733 TI - Requirement of A1-a for bacillus Calmette-Guerin-mediated protection of macrophages against nitric oxide-induced apoptosis. AB - The role of apoptosis in regulating the course of intracellular microbial infection is not well understood. We studied the relationship between apoptotic regulation and bacillus Calmette-Guerin (BCG) treatment in murine peritoneal exudate macrophages (PEM) and the J774 macrophage cell line. In both PEM and J774 cells, mRNA expression of the anti-apoptotic gene, A1, was selectively induced by BCG treatment as compared with other bcl2 family members (bcl-w, bcl-2, bcl-xl, bcl-xs, bax, bak, bad). In PEM, A1 expression was maximal by 8 h postinfection and was abrogated by the proteasomal inhibitor MG-132. The induction was independent of protein synthesis as well as the p38 mitogen-activated protein kinase and phosphatidylinositol 3-kinase pathways and did not require live organism. Three genes encoding closely related isoforms of A1 were all expressed; however, the A1-a isoform displayed the greatest fold induction in PEM. BCG induced A1 expression was associated with protection of host macrophages from NO mediated apoptosis in both PEM and J774 cells. BCG-mediated protection was abrogated in PEM derived from A1-a(-/-) mice, indicating a requirement of A1-a for survival of inflammatory macrophages. PMID- 11254734 TI - Macrophages transfected with adenovirus to express IL-4 reduce inflammation in experimental glomerulonephritis. AB - Nephrotoxic nephritis (NTN) is characterized by acute macrophage-dependent inflammation and serves as a model of human glomerulonephritis. In this study we have transfected rat macrophages with recombinant adenovirus expressing IL-4 (Ad IL4) and demonstrated that these transfected macrophages develop fixed properties as a result of transfection, as shown by reduced NO production in response to IFN gamma and TNF. Ad-IL4-transfected macrophages localized with enhanced efficiency to inflamed glomeruli after renal artery injection in rats with NTN compared with adenovirus expressing beta-galactosidase (Ad-beta gal)-transfected macrophages and produced elevated levels of the cytokine in glomeruli in vivo for up to 4 days. The delivery of IL-4-expressing macrophages produced a marked reduction in the severity of albuminuria (day 2 albuminuria, 61 +/- 15 mg/24 h) compared with unmodified NTN (day 2 albuminuria, 286 +/- 40 mg/24 h; p < 0.01), and this was matched by a reduction in the number of ED1-positive macrophages infiltrating the glomeruli. Interestingly, the injection of IL-4-expressing macrophages into single kidney produced a marked reduction in the numbers of ED1-positive macrophages in the contralateral noninjected kidney, an effect that could not be mimicked by systemic delivery of IL-4-expressing macrophages. This implies that the presence of IL-4-expressing macrophages in a single kidney can alter the systemic development of the inflammatory response. Macrophage transfection and delivery provide a valuable system to study and modulate inflammatory disease and highlight the feasibility of macrophage-based gene therapy. PMID- 11254735 TI - Reduced early alcohol-induced liver injury in CD14-deficient mice. AB - Activation of Kupffer cells by gut-derived endotoxin is associated with alcohol induced liver injury. Recently, it was shown that CD14-deficient mice are more resistant to endotoxin-induced shock than wild-type controls. Therefore, this study was designed to investigate the role of CD14 receptors in early alcohol induced liver injury using CD14 knockout and wild-type BALB/c mice in a model of enteral ethanol delivery. Animals were given a high-fat liquid diet continuously with ethanol or isocaloric maltose-dextrin as control for 4 wk. The liver to body weight ratio in wild-type mice (5.8 +/- 0.3%) was increased significantly by ethanol (7.3 +/- 0.2%) but was not altered by ethanol in CD14-deficient mice. Ethanol elevated serum alanine aminotransferase levels nearly 3-fold in wild-type mice, but not in CD14-deficient mice. Wild-type and knockout mice given the control high-fat diet had normal liver histology, whereas ethanol caused severe liver injury (steatosis, inflammation, and necrosis; pathology score = 3.8 +/- 0.4). In contrast, CD14-deficient mice given ethanol showed minimal hepatic changes (score = 1.6 +/- 0.3, p < 0.05). Additionally, NF-kappa B, TGF-beta, and TNF-alpha were increased significantly in wild-type mice fed ethanol but not in the CD14 knockout. Thus, chronic ethanol feeding caused more severe liver injury in wild-type than CD14 knockouts, supporting the hypothesis that endotoxin acting via CD14 plays a major role in the development of early alcohol-induced liver injury. PMID- 11254736 TI - Nonphlogistic clearance of late apoptotic neutrophils by macrophages: efficient phagocytosis independent of beta 2 integrins. AB - Neutrophils undergo constitutive death by apoptosis, leading to safe nonphlogistic phagocytosis and clearance by macrophages. Recent work has shown that before secondary necrosis, neutrophils exhibiting classical features of apoptosis can progress to a morphologically defined late apoptotic state. However, whether such neutrophils could be safely cleared was unknown. We now report that human late apoptotic neutrophils could be purified from cultured neutrophil populations undergoing constitutive death and were subsequently ingested by human monocyte-derived macrophages by serum-independent mechanisms that did not trigger the release of IL-8 or TNF-alpha. Such ingestion was specifically inhibited by Abs to thrombospondin-1 and the alpha(v)beta(3) vitronectin receptor. Murine bone marrow-derived macrophage phagocytosis of late and early apoptotic neutrophils occurred by similar mechanisms, proceeding with the same efficiency as that observed for wild-type controls when macrophages from [alpha(m)](-/-) or [beta(2)](-/-) mice were used. We conclude that specific nonphlogistic, beta(2) integrin-independent mechanisms involving thrombospondin-1 and alpha(v)beta(3) allow macrophages to ingest late apoptotic neutrophils without eliciting inflammatory cytokine secretion. PMID- 11254737 TI - T cells of multiple sclerosis patients target a common environmental peptide that causes encephalitis in mice. AB - Multiple sclerosis (MS) is a chronic autoimmune disease triggered by unknown environmental factors in genetically susceptible hosts. MS risk was linked to high rates of cow milk protein (CMP) consumption, reminiscent of a similar association in autoimmune diabetes. A recent rodent study showed that immune responses to the CMP, butyrophilin, can lead to encephalitis through antigenic mimicry with myelin oligodendrocyte glycoprotein. In this study, we show abnormal T cell immunity to several other CMPs in MS patients comparable to that in diabetics. Limited epitope mapping with the milk protein BSA identified one specific epitope, BSA(193), which was targeted by most MS but not diabetes patients. BSA(193) was encephalitogenic in SJL/J mice subjected to a standard protocol for the induction of experimental autoimmune encephalitis. These data extend the possible, immunological basis for the association of MS risk, CMP, and CNS autoimmunity. To pinpoint the same peptide, BSA(193), in encephalitis-prone humans and rodents may imply a common endogenous ligand, targeted through antigenic mimicry. PMID- 11254738 TI - Frequencies of neuroantigen-specific T cells in the central nervous system versus the immune periphery during the course of experimental allergic encephalomyelitis. AB - Direct measurements of the frequency and the cytokine signature of the neuroantigen-specific effector cells in experimental allergic encephalomyelitis (EAE) are a continuing challenge. This is true for lymphoid tissues, and more importantly, for the CNS itself. Using enzyme-linked immunospot analysis (ELISPOT) assays, we followed proteolipid protein (PLP) 139--151-specific T cells engaged by active immunization of SJL mice. The total numbers of PLP(139--151) specific CD4 cells were highest before disease onset. At this time, these cells resided in lymphoid and nonlymphoid tissues, but were not detected in the CNS. While the PLP(139--151)-specific cells reached high frequencies in the CNS during clinical EAE, in absolute numbers, less than 20% of them were present in the target organ, with the majority residing in the periphery throughout all stages of the disease. The numbers of PLP(139--151)-specific cells gradually declined in both compartments with time. While eventually this first wave of effector cells completely disappeared from the CNS, PLP(178--191)-specific cells became engaged, being detected first in the CNS. These data suggest that throughout all stages of EAE, the effector cells in the CNS are recruited from a vast peripheral reservoir, and that the second wave of effector cells is engaged while the first wave undergoes exhaustion. PMID- 11254739 TI - Persistence of pathogenic CD4+ Th1-like cells in vivo in the absence of IL-12 but in the presence of autoantigen. AB - Despite recent successful treatment of murine autoimmune disease with anti-IL-12 mAb, it has not yet been addressed whether anti-IL-12 mAb can also be effective in late stages of disease and whether it can provide lasting protection against recurrence, especially during continued presence of autoantigen. We used a newly developed psoriasis model in scid/scid mice, which allows easy tracking of pathogenic T cells, to show that when anti-IL-12 mAb is given for 2 wk (1 mg/wk) in the late stage of severe disease, inflammation is greatly reduced, as measured by ear thickness and histology (scores, 1.1 +/- 0.1 vs 2.0 +/- 0.4). Moreover, prolonged treatment (4 wk) of chronic psoriatic mice with high doses of mAb (1 mg/wk; prolonged active anti-inflammatory treatment (PAAIT)) results in the almost complete resolution of lesions (scores, 0.3 +/- 0.1 vs 2.7 +/- 0.2). Surprisingly, however, despite these significant treatment results, the psoriasis like lesions return soon after the anti-IL-12 mAb treatment is discontinued. This rapid relapse of disease may be attributed to large populations of activated CD4(+) T cells present in the lymph nodes of PAAIT animals still expressing an effector/memory phenotype (CD45RB(low), L-selectin(low)). Upon stimulation in vitro such PAAIT lymph node cells secrete high amounts of IFN-gamma (129 ng/ml); when transferred into naive scid/scid animals they are able to rapidly induce disease without costimulation. Our data indicates an alternative IL-12 independent pathway for pathogenic Th-1-like cells in vivo during the chronic phase of disease that allows these cells to persist and maintain their pathogenicity in the draining lymph tissue of the autoimmune site. PMID- 11254740 TI - Specific immunotherapy by genetically engineered APCs: the "guided missile" strategy. AB - We tested the hypothesis that APCs genetically engineered to present an Ag and to express Fas ligand (FasL) simultaneously can target and eliminate Ag-specific T cells. Transgenic T cells specific for influenza hemagglutinin (HA) were used as targets. We prepared recombinant vaccinia virus vectors (VVV) to transfer the gene constructs individually or simultaneously into APCs. We prevented unwanted viral replication by attenuating the VVVs with psoralen-UV light treatment. For presentation of the HA Ag, APCs were transduced with cDNA for HA flanked by sequences of the lysosome-associated membrane protein that direct efficient processing and presentation of the Ag by APCs. As a "warhead" for the APCs, we transduced them with the gene for FasL, which induces apoptosis of Fas-expressing activated T cells. To protect the transduced APCs from self-destruction by FasL, we transferred cDNA for a truncated form of Fas-associated death domain, which inhibits Fas-mediated cell death. Our results show that the engineered APCs effectively expressed the genes of interest. APCs transduced with VVV carrying all three gene constructs specifically killed HA-transgenic T cells in culture. Coculture with T cells specific for an unrelated Ag (OVA) had no significant effect. Our in vitro findings show that APCs can be genetically engineered to target and kill Ag-specific T cells and represent a promising novel strategy for the specific treatment of autoimmune diseases. PMID- 11254741 TI - Reactivity of anti-proliferating cell nuclear antigen (PCNA) murine monoclonal antibodies and human autoantibodies to the PCNA multiprotein complexes involved in cell proliferation. AB - Proliferating cell nuclear Ag (PCNA) occurs as a component of multiprotein complexes during cell proliferation. We found the complexes to react with murine anti-PCNA mAbs, but not with anti-PCNA Abs in lupus sera. The complexes were purified from rabbit thymus extract by affinity chromatography using anti-PCNA mAbs (TOB7, TO17, and TO30) and analyzed by ELISA, immunoprecipitation, immunoblotting, and HPLC gel filtration. That PCNA was complexed with other proteins was demonstrated by its copurification with a group of proteins excluded by an HPLC G3000 SW column. Although immunoblot analysis showed the mAbs to react exclusively with the 34-kDa PCNA polypeptide, they nonetheless immunoprecipitated the same group of proteins, confirming the interaction of the isolated PCNA with other proteins. Anti-PCNA sera, including AK, which reacts with biologically functional sites on PCNA, did not react with complexed PCNA, but did react with it once it was dissociated from the complexes. PCNA complexes in turn reacted with murine anti-DNA mAbs, as well as with Abs against p21, replication protein A, DNA helicase II, cyclin-dependent kinases 4 and 5, and topoisomerase I. These findings suggest that the PCNA complexes purified using anti-PCNA mAbs comprise the "protein machinery" for DNA replication and cell cycle regulation. They also suggest that anti-PCNA mAbs are useful tools with which to characterize the protein-protein interactions within PCNA complexes, as well as the autoimmune responses to proteins interacting with PCNA, which may shed light on the mechanisms of autoantibody production in lupus patients. PMID- 11254743 TI - Disturbances in the normal regulation of SREBP-sensitive genes in PPAR alpha deficient mice. AB - Peroxisome proliferator-activated receptor alpha (PPAR alpha)-null mice were used to investigate the nature of the relationship between the normal circadian rhythm of hepatic PPAR alpha expression and the expression of the lipogenic and cholesterogenic sterol regulatory element-binding protein (SREBP)-regulated genes, acetyl-CoA carboxylase, fatty acid synthase (FAS), and 3-hydroxy-3 methylglutaryl-CoA reductase (HMG-CoAR). The expression of FAS and HMG-CoAR varied rhythmically over the diurnal cycle in the normal mice, with patterns that were the opposite of that of PPAR alpha. The diurnal variation of lipogenic and cholesterogenic gene expression was attenuated or abolished in the PPAR alpha null mice. This resulted in decreased expression compared with normal mice, but only during the dark phase of the cycle, when food intake was high. The diurnal variation in hepatic fatty acid and cholesterol synthesis was also abolished in the PPAR alpha-null animals and the variations in the concentration of plasma triacylglycerol, nonesterified fatty acids, and cholesterol were all attenuated. The failure of HMG-CoAR expression to increase during the feeding period in the PPAR alpha-null mice was associated with a decrease in hepatic nonesterified cholesterol content and an increase in cholesteryl ester compared with normal mice. There was no defect in the downregulation of hepatic HMG-CoAR mRNA in response to dietary cholesterol in the PPAR alpha-null mice. Under these conditions, hepatic PPAR gamma expression increased in both the control and PPAR alpha-deficient mice. The results suggest that PPAR alpha-deficiency disturbs the normal circadian regulation of certain SREBP-sensitive genes in the liver, but does not affect their response to dietary cholesterol. -- Patel, D. D., B. L. Knight, D. Wiggins, S. M. Humphreys, and G. F. Gibbons. Disturbances in the normal regulation of SREBP-sensitive genes in PPAR alpha-deficient mice. J. Lipid Res. 2001. 42: 328--337. PMID- 11254742 TI - Lysosomal cholesterol derived from mildly oxidized low density lipoprotein is resistant to efflux. AB - In atherosclerotic lesions, macrophages store lipid in cytoplasmic inclusions and lysosomes. Regression studies show that lysosomal lipid is not as easily cleared as cytoplasmic inclusion lipid. Macrophages enriched with mildly oxidized low density lipoprotein (oxLDL) accumulate cholesteryl ester (CE) and free cholesterol (FC) in lysosomes. We examined whether lysosomal stores of cholesterol from oxLDL are cleared from THP-1 and mouse macrophages. As in previous studies, oxLDL-enriched THP-1 macrophages accumulated substantial lysosomal cholesterol. Surprisingly, less than 12% of oxLDL-derived lysosomal CE was cleared to efficient FC acceptors (e.g., cyclodextrins, apolipoprotein/phosphatidylcholine vesicles, and fetal bovine serum). Filipin staining showed that lysosomes of oxLDL-treated THP-1 cells contained FC, and despite removal of most of the cell FC (70--80%) by incubation with cyclodextrins, filipin staining of FC in lysosomes did not diminish. Also, when THP-1 macrophages were incubated with [(3)H]CE oxLDL, 73--76% of the [(3)H]CE was retained in a lysosomal hydrolysis resistant pool. In contrast, greater than 90% of acetylated low density lipoprotein (acLDL) [(3)H]CE was hydrolyzed. Furthermore, [(3)H]FC liberated from oxLDL [(3)H]CE was released at a slower rate to cyclodextrins than was [(3)H]FC from acLDL [(3)H]CE. In contrast, only 27% of oxLDL [(3)H]CE was resistant to hydrolysis in mouse macrophages, and the [(3)H]FC generated from oxLDL and acLDL [(3)H]CE was released to cyclodextrins at similar rates. We conclude that lack of hydrolysis and efflux of oxLDL cholesterol is not exclusively inherent in oxLDL, but also requires specific cell factors present in one cell type but not the other.--Yancey, P. G., and W. G. Jerome. Lysosomal cholesterol derived from mildly oxidized low density lipoprotein is resistant to efflux. J. Lipid Res. 2001. 42: 317--327. PMID- 11254744 TI - Atomic resolution structure analysis of beta' polymorph crystal of a triacylglycerol: 1,2-dipalmitoyl-3-myristoyl-sn-glycerol. AB - The crystal structure of the beta'-2 form of a mixed chain triacylglycerol (TAG), 1,2-dipalmitoyl-3-myristoyl-sn-glycerol (PPM), was determined to a final reliability factor of 0.074. This work is the first to resolve the atomic-level structure of the beta' polymorph, which is of the highest functionality among multiple polymorphs in asymmetric TAG. In particular, fat crystals present in food emulsions are in beta', whose transformation into beta causes deterioration in their physical properties. beta'-2, one of the two beta' forms of PPM, forms a monoclinic unit cell with a space group of C2; Z = 8, a = 16.534(5) A, b = 7.537(2) A, c = 81.626(9) A; beta = 90.28(2) degrees, V = 10171(3) A(3), density = 1.018 g/cm(3), and mu = 4.96 cm(-1). The following characteristics were obtained: 1) two asymmetric units, named A and B, form a hybrid-type orthorhombic perpendicular subcell; 2) the two asymmetric units reveal different glycerol conformations: trans for sn-1 palmitic acid and sn-2 palmitic acid, but gauche for sn-3 myristic acid in A; and trans for sn-2 palmitic acid and sn-3 myristic acid, but gauche for sn-1 palmitic acid in B; 3) a unit lamellae reveals a four chain-length structure consisting of two double-layer leaflets; 4) the two double layer leaflets are combined end-by-end in a unit lamellae; and 5) the chain axes are alternatively inclined against the lamellar interface. -- Sato, K., M. Goto, J. Yano, K. Honda, D. R. Kodali, and D. M. Small. Atomic resolution structure analysis of beta' polymorph crystal of a triacylglycerol: 1,2-dipalmitoyl-3 myristoyl-sn-glycerol. J. Lipid Res. 2001. 42: 338--345. PMID- 11254746 TI - Ergosterol biosynthesis in novel melanized fungi from hypersaline environments. AB - Halotolerant and halophilic melanized fungi were recently described in hypersaline waters. A close study of the sterol composition of such fungi, namely Hortaea werneckii, Alternaria alternata, Cladosporium sphaerospermum, Cladosporium sp., and Aureobasidium pullulans revealed the dominance of ergosterol and the presence of 29 intermediates of its biosynthesis pathway. The presence or absence of intermediates from distinct synthesis routes gave insight into the operative synthetic pathways from 4,4,14-trimethylcholesta-8,24-dien-3 beta-ol (lanosterol) to ergosterol in melanized fungi and in Saccharomyces cerevisiae, a reference yeast cultured in parallel. In all studied melanized fungi, initial methylation at C-24 took place before C-14 and C-4 demethylation, involving a different reaction sequence from that observed in S. cerevisiae. Further transformation was observed to occur through various routes. In A. alternata, isomerization at C-7 takes place prior to desaturation at C-5 and C 22, and methylene reduction at C-24. In addition to these pathways in Cladosporium spp., H. werneckii, and A. pullulans, ergosterol may also be synthesized through reduction of the C-24 methylene group before desaturation at C-5 and C-22 or vice versa. Moreover, in all studied melanized fungi except A. alternata, ergosterol biosynthesis may also proceed through C-24 methylene reduction prior to C-4 demethylation. -- Mejanelle, L., J. F. Lopez, N. Gunde Cimerman, and J. O. Grimalt. Ergosterol biosynthesis in novel melanized fungi from hypersaline environments. J. Lipid Res. 2001. 42: 352--358. PMID- 11254745 TI - Partitioning of polyunsaturated fatty acids, which prevent cardiac arrhythmias, into phospholipid cell membranes. AB - It has been demonstrated in animal studies that polyunsaturated fatty acids (PUFA) prevent ischemia-induced malignant ventricular arrhythmias, a major cause of sudden cardiac death in humans. To learn how these PUFA, at low micromolar concentrations, exert their antiarrhythmic activity, we studied their effects in vitro on the contractions of isolated cardiac myocytes and the conductances of their sarcolemmal ion channels. These fatty acids directly stabilize electrically every cardiac myocyte by modulating the conductances of specific ion channels in their sarcolemma. In this study, we determined the molar ratio of PUFA to the moles of phospholipid (PL) in cell membranes to learn if the ratio is so low as to preclude the possibility that the primary site of action of PUFA is on the packing of the membrane PL. [(3)H]-arachidonic acid (AA) was used to measure the incorporation of PUFA, and the inorganic phosphorous of the PL was determined as a measure of the moles of PL in the cell membrane. Our results indicate that the mole percent of AA to moles of phospolipid is very low (< or =1.0) at the concentrations that affect myocyte contraction and the conductance of voltage dependent Na(+) and L-type Ca(2)+ channels in rat cardiomyocytes and in alpha subunits of human myocardial Na(+) channels. In conclusion, it seems highly unlikely that these fatty acids are affecting the packing of PL within cell membranes as their way of modulating changes in cell membrane ion currents and in preventing arrhythmias in our contractility studies. -- Pound, E. M., J. X. Kang, and A. Leaf. Partitioning of polyunsaturated fatty acids, which prevent cardiac arrhythmias, into phospholipid cell membranes. J. Lipid Res. 2001. 42: 346--351. PMID- 11254747 TI - Maternal essential fatty acid deficiency depresses serum leptin levels in suckling rat pups. AB - Dietary lipid quantity and quality have recently been shown to affect serum leptin levels in adult rats. Moreover, suckling pups from dams fed a high fat diet had increased serum leptin levels. The aim of the present study was to analyze the influence of essential fatty acid (EFA) deficiency on serum leptin levels in dams and their pups during the suckling period. For the last 10 days of gestation and throughout lactation, pregnant rats were fed a control or an EFA deficient (EFAD) diet. The levels of leptin and EFA in the serum of the dams and pups were analyzed 1, 2, and 3 weeks after delivery. In parallel, serum levels of glucose and corticosterone were analyzed in the pups. Low serum leptin levels were found in the control lactating dams during the entire lactation period compared with the age-matched nonlactating animals. The leptin concentrations in the lactating dams fed the EFAD diet were lower compared with those fed the control diet. The serum leptin levels of suckling pups from dams on the EFAD diet were markedly decreased compared with controls (P < 0.05). The reduced serum leptin levels could not be explained by nutritional restriction as evaluated by serum levels of glucose and corticosterone. These results indicate the importance of the EFA composition of the maternal diet for serum leptin levels in both dams and pups. EFA deficiency in lactating dams may cause long-term effects on the pups through dysregulation of leptin and leptin-dependent functions. -- Korotkova, M., B. Gabrielsson, L. A. Hanson, and B. Strandvik. Maternal essential fatty acid deficiency depresses serum leptin levels in suckling rat pups. J. Lipid Res. 2001. 42: 359--365. PMID- 11254748 TI - Oxysterols in the circulation of patients with the Smith-Lemli-Opitz syndrome: abnormal levels of 24S- and 27-hydroxycholesterol. AB - Infants with the cholesterol synthesis defect Smith- Lemli-Opitz syndrome (SLO) have reduced activity of the enzyme 7-dehydrocholesterol-7-reductase and accumulate 7-dehydrocholesterol, with the highest concentration in the brain. As a result of the generally reduced content of cholesterol, plasma levels of oxysterols would be expected to be reduced. 24S-hydroxycholesterol is almost exclusively formed in the brain, whereas 27-hydroxycholesterol is mainly formed from extrahepatic and extracerebral cholesterol. In accordance with the expectations, sterol-correlated plasma levels of 24S-hydroxycholesterol were reduced in infants with SLO (by about 50%). In contrast, the sterol-correlated levels of 27-hydroxycholesterol in the circulation were markedly increased. No side-chain oxidized metabolites of 7-dehydrocholesterol were detected in the circulation. Recombinant human CYP27 had markedly lower 27-hydroxylase activity toward 7-dehydrocholesterol than towards cholesterol. HEK293 cells expressing 24S hydroxylase active toward cholesterol had no significant activity towards 7 dehydrocholesterol. The plasma levels of 3 beta,7 alpha-dihydroxy-5-cholestenoic in the patients acid were reduced, suggesting a generally reduced metabolism of 27-oxygenated steroids. It is concluded that the accumulation of 7 dehydrocholesterol in the brains of patients with SLO reduces formation of 24S hydroxycholesterol. The condition is associated with markedly increased circulating levels of 27-hydroxycholesterol, most probably due to reduced metabolism of this oxysterol. We discuss the possibility that the circulating levels of 24S-hydroxycholesterol may be used as a marker for the severity of the disease.--Bjorkhem, I., L. Starck, U. Andersson, D. Lutjohann, S. von Bahr, I. Pikuleva, A. Babiker, and U. Diczfaulsy. Oxysterols in the circulation of patients with the Smith-Lemli-Opitz syndrome: abnormal levels of 24S- and 27 hydroxycholesterol. J. Lipid Res. 2001. 42: 366--371. PMID- 11254749 TI - Dietary diacylglycerol suppresses high fat and high sucrose diet-induced body fat accumulation in C57BL/6J mice. AB - Diacylglycerol (DG) comprises up to approximately 10% of various edible oils. In the present study, we examined the effects of dietary DG consisting mainly of 1,3 species on body weight, body fat accumulation, and mRNA levels of various genes involved in energy homeostasis in obesity-prone C57BL/6J mice. Five-month feeding with the high triacylglycerol (TG) diet (30% TG + 13% sucrose) resulted in significant increases in body weight, visceral fat accumulation, and circulating insulin and leptin levels compared with mice fed the control diet (5% TG). Compared with mice fed the high TG diet, body weight gain and visceral fat weight were reduced by 70% and 79%, respectively, in those fed the high DG diet (30% DG + 13% sucrose). In addition, circulating leptin and insulin levels were reduced to the respective control levels. Compared with high TG feeding, high DG feeding suppressed the elevation of leptin mRNA expression in adipose tissue, and up regulated acyl-coenzyme (Co)A oxidase and acyl-CoA synthase mRNA expression in the liver. These results indicate that dietary DG is beneficial for suppression of high fat diet-induced body fat accumulation. Furthermore, it is suggested that structural differences in DG and TG, but not the composition of fatty acid, markedly affect nutritional behavior of lipids. -- Murase, T., T. Mizuno, T. Omachi, K. Onizawa, Y. Komine, H. Kondo, T. Hase, and I. Tokimitsu. Dietary diacylglycerol suppresses high fat and high sucrose diet-induced body fat accumulation in C57BL/6J mice. J. Lipid Res. 2001. 42: 372--378. PMID- 11254750 TI - Role of individual amino acids of apolipoprotein A-I in the activation of lecithin:cholesterol acyltransferase and in HDL rearrangements. AB - The central region of apolipoprotein A-I (apoA-I), spanning residues 143--165, has been implicated in lecithin:cholesterol acyltransferase (LCAT) activation and also in high density lipoprotein (HDL) structural rearrangements. To examine the role of individual amino acids in these functions, we constructed, overexpressed, and purified two additional point mutants of apoA-I (P143R and R160L) and compared them with the previously studied V156E mutant. These mutants have been reported to occur naturally and to affect HDL cholesterol levels and cholesterol esterification in plasma. The P143R and R160L mutants were effectively expressed in Escherichia coli as fusion proteins and were isolated in at least 95% purity. In the lipid-free state, the mutants self-associated similarly to wild-type protein. All the mutants, including V156E, were able to lyse dimyristoylphosphatidylcholine liposomes. In the lipid-bound state, the major reconstituted HDL (rHDL) of the mutants had diameters similar to wild type (96- 98 A). Circular dichroism and fluorescence methods revealed no major differences among the structures of the lipid-free or lipid-bound mutants and wild type. In contrast, the V156E mutant had exhibited significant structural, stability, and self-association differences compared with wild-type apoA-I in the lipid-free state, and formed rHDL particles with larger diameters. In this study, limited proteolytic digestion with chymotrypsin showed that the V156E mutant, in lipid free form, has a distinct digestion pattern and surface exposure of the central region, compared with wild type and the other mutants. Reactivity of rHDL with LCAT was highest for wild type (100%), followed by P143R (39%) and R160L (0.6%). Tested for their ability to rearrange into 78-A particles, the rHDL of the two mutants (P143R and R160L) behaved normally, compared with the rHDL of V156E, which showed no rearrangement after the 24-h incubation with low density lipoprotein (LDL). Similarly, the rHDL of V156E was resistant to rearrangement in the presence of apoA-I or apoA-II. These results indicate that structural changes are absent or modest for the P143R and R160L mutants, especially in rHDL form; that these mutants have normal conformational adaptability; and that LCAT activation is obliterated for R160L.Thus, individual amino acid changes may have markedly different structural and functional consequences in the 143--165 region of apoA-I. The R160L mutation appears to have a direct effect in LCAT activation, while the P143R mutation results in only minor structural and functional effects. Also, the processes for LCAT activation and hinge mobility appear to be distinct even if the same region of apoA-I is involved. -- Cho, K-H., D. M. Durbin, and A. Jonas. Role of individual amino acids of apolipoprotein A-I in the activation of lecithin:cholesterol acyltransferase and in HDL rearrangements. J. Lipid Res. 2001. 42: 379--389. PMID- 11254751 TI - Cholesterol sulfate stimulates involucrin transcription in keratinocytes by increasing Fra-1, Fra-2, and Jun D. AB - Lipids that are synthesized de novo in the epidermis, including fatty acids, oxysterols, 1,25-dihydroxyvitamin D(3), and farnesol, can regulate the differentiation of normal human keratinocytes (NHK). Cholesterol sulfate (CS), an epidermal lipid that is produced in the upper nucleated layers of the epidermis coincident with terminal differentiation, has been shown to play a role in the regulation of the late stages of keratinocyte differentiation, including formation of the cornified envelope. In the present study, we determined i) whether CS regulates involucrin (INV), an early keratinocyte differentiation marker, and ii) the mechanism by which CS regulates differentiation. mRNA and protein levels of INV, a precursor protein of the cornified envelope, increased 2 to 3-fold in NHK incubated in the presence of CS. In contrast, cholesterol had no effect on INV protein or mRNA levels. Transcriptional regulation was assessed in NHK transfected with INV promoter-luciferase constructs. CS increased luciferase reporter activity approximately 2- to 3-fold in NHK transfected with a 3.7-kb INV promoter construct. Deletional analysis revealed a CS-responsive region of the INV promoter located between bp --2452 and --1880. A 5-base pair (bp) mutation of the AP-1 site (bp --2117 to --2111) within this responsive region abolished CS responsiveness, suggesting a role for the AP-1 complex in the regulation of INV transcription by CS. Electrophoretic mobility shift analysis demonstrated increased binding of nuclear extracts isolated from CS-treated NHK to AP-1 DNA as compared with vehicle-treated controls. Incubation of the nuclear extract with the appropriate antibodies showed that the AP-1 DNA-binding complex contained Fra-1, Fra-2, and Jun D. Western blots demonstrated that CS treatment increased the levels of Fra-1, Fra-2, and Jun D, and Northern analyses revealed that CS increased mRNA levels for these same AP-1 factors. These data indicate that CS, an endogenous lipid synthesized by keratinocytes, regulates the early stages of keratinocyte differentiation, and may do so through its ability to modulate levels of AP-1 proteins. -- Hanley, K., L. Wood, D. C. Ng, S. S. He, P. Lau, A. Moser, P. M. Elias, D. D. Bikle, M. L. Williams, and K. R. Feingold. Cholesterol sulfate stimulates involucrin transcription in keratinocytes by increasing Fra-1, Fra-2, and Jun D. J. Lipid Res. 2001. 42: 390--398. PMID- 11254752 TI - Vesicle-binding properties of wild-type and cysteine mutant forms of alpha(1) domain of apolipoprotein B. AB - Previous studies demonstrated that structural perturbation of the alpha(1) domain of apolipoprotein B (apoB) blocked the initiation of lipoprotein assembly. We explored the hypothesis that this domain may interact with the inner leaflet of the endoplasmic reticulum membrane in a manner that may nucleate microsomal triglyceride transfer protein-dependent lipid sequestration. ApoB-17 (amino terminal 17% of apoB), which contains most of the alpha(1) domain, was expressed stably in rat hepatoma cells and recovered from medium in lipid-poor form. On incubation with phospholipid vesicles composed of 1-myristol-2-myristoyl-sn glycero-3-phosphocholine or 1-palmitoyl-2-oleoyl-sn-gylycero-3-phosphocholine, apoB-17 underwent vesicle binding and was recovered in the d < 1.25 g/ml gradient fraction. To determine whether vesicle binding is disrupted by the same structural perturbations that block lipoprotein assembly in vivo, apoB-17 was subjected to partial and complete chemical reduction. Although normally a soluble peptide, mild reduction of apoB-17 caused its precipitation, suggesting that hydrophobic, solvent-inaccessible domains within the alpha(1) domain of apoB are stabilized by intramolecular disulfide bonds. In contrast to apoB-17 chemically reduced in vitro, forms of apoB-17 bearing pairwise cysteine-to-serine substitutions were recovered in soluble form from transiently transfected COS-1 cell extracts. Although individual disruption of disulfide bond 2 or 4 in apoB-28 and apoB-50 was previously shown to block lipoprotein assembly in vivo, these alterations had no impact on the ability of apoB-17 to bind to phospholipid vesicles in vitro or on its capacity to form recombinant lipoprotein particles. These results suggest that while the vesicle/lipid-binding property of the alpha(1) domain may reflect an essential role required for the initiation of lipoprotein formation, some other aspect of alpha(1) domain function is perturbed by disruption of native disulfide bonds. -- DeLozier, J. A., J. S. Parks, and G. S. Shelness. Vesicle-binding properties of wild-type and cysteine mutant forms of alpha(1) domain of apolipoprotein B. J. Lipid Res. 2001. 42: 399--406. PMID- 11254753 TI - Supplementation of postmenopausal women with fish oil does not increase overall oxidation of LDL ex vivo compared to dietary oils rich in oleate and linoleate. AB - Although replacement of dietary saturated fat with monounsaturated and polyunsaturated fatty acids (MUFA and PUFA) has been advocated for the reduction of cardiovascular disease risk, diets high in PUFA could increase low density lipoprotein (LDL) susceptibility to oxidation, potentially contributing to the pathology of atherosclerosis. To investigate this possibility, 15 postmenopausal women in a blinded crossover trial consumed 15 g of sunflower oil (SU) providing 12.3 g/day of oleate, safflower oil (SA) providing 10.5 g/day of linoleate, and fish oil (FO) providing 2.0 g/day of eicosapentaenoate (EPA) and 1.4 g/day of docosahexaenoate (DHA). During CuSO(4)-mediated oxidation, LDL was depleted of alpha-tocopherol more rapidly after FO supplementation than after supplementation with SU (P = 0.0001) and SA (P = 0.05). In LDL phospholipid and cholesteryl ester fractions, loss of n-3 PUFA was greater and loss of n-6 PUFA less after FO supplementation than after SU and SA supplementation (P < 0.05 for all), but loss of total PUFA did not differ. The lag phase for phosphatidylcholine hydroperoxide (PCOOH) formation was shorter after FO supplementation than after supplementation with SU (P = 0.0001) and SA (P = 0.006), whereas the lag phase for cholesteryl linoleate hydroperoxide (CE18:2OOH) formation was shorter after FO supplementation than after SU (P = 0.03) but not SA. In contrast, maximal rates of PCOOH and CE18:2OOH formation were lower after FO supplementation than after SA (P = 0.02 and 0.0001, respectively) and maximal concentrations of PCOOH and CE18:2OOH were lower after FO supplementation than after SA (P = 0.03 and 0.0006, respectively). Taken together, our results suggest that FO supplementation does not increase the overall oxidation of LDL ex vivo, especially when compared with SA supplementation. Consequently, health benefits related to increased fish consumption may not be offset by increased LDL oxidative susceptibility.-- Higdon, J. V., S. H. Du, Y. S. Lee, T. Wu, and R. C. Wander. Supplementation of postmenopausal women with fish oil does not increase overall oxidation of LDL ex vivo compared to dietary oils rich in oleate and linoleate. J. Lipid Res. 2001. 42: 407--418. PMID- 11254754 TI - Reversal of docosahexaenoic acid deficiency in the rat brain, retina, liver, and serum. AB - The loss of docosahexaenoic acid (DHA) from the retina or brain has been associated with a loss in nervous-system function in experimental animals, as well as in human infants fed vegetable oil-based formulas. The reversibility of the loss of DHA and the compensation by an increase in the n-6 docosapentaenoic acid (DPAn-6) was studied in young adult rats. Long-Evans rats were subjected to a very low level of n-3 fatty acids through two generations. The F2 generation, n 3-deficient animals at 7 weeks of age were provided a repletion diet containing both alpha-linolenate and DHA. A separate group of F2 generation rats had been maintained on an n-3-adequate diet of the same composition. Tissues from the brain, retina, liver, and serum were collected on weeks 0, 1, 2, 4, and 8 from both groups of animals. The concentrations of DHA, DPAn-6, and other fatty acids were determined and the rate of recovery and length of time needed to complete DHA recovery were determined for each tissue. The DHA level in the brain at 1 and 2 weeks after diet reversal was only partially recovered, rising to approximately 20% and 35%, respectively, of the n-3-adequate group level. Full recovery was not obtained until 8 weeks after initiation of the repletion diet. Although the initial rate of retinal DHA accretion was greater than that of brain DHA, the half-time for DHA recovery was only marginally greater. On the other hand, the levels of DHA in the serum and liver were approximately 90% and 100% replaced, respectively, within 2 weeks of diet reversal. A consideration of the total amounts and time courses of DHA repleted in the nervous system compared with the liver and circulation suggests that transport-related processes may limit the rate of DHA repletion in the retina and brain.-- Moriguchi, T., J. Loewke, M. Garrison, J. N. Catalan, N. Salem, Jr. Reversal of docosahexaenoic acid deficiency in the rat brain, retina, liver, and serum. J. Lipid Res. 2001. 42: 419--427. PMID- 11254755 TI - Nutritional lipid emulsions modulate cellular signaling and activation of human neutrophils. AB - Although numerous studies suggest that nutritional lipids modulate human immune responses, the mechanism behind this observation remains unclear. On the basis of the hypothesis that lipids might affect cellular signaling we evaluated the effects of various lipid emulsions on two major pathways involved in neutrophil activation: second messenger (Ca(2)+) mobilization and protein kinase C (PKC) activation. Activation by opsonized yeast particles (serum-treated zymosan; STZ) increased cytosolic [Ca(2)+] ([Ca(2)+](i)) in neutrophils, with an initial slow rise that turned into a fast phase until a plateau was reached. The PKC activator 4-alpha-phorbol 12-myristate 13-acetate (PMA) markedly increased the initial STZ induced [Ca(2)+](i) rise. This PMA effect was mimicked by emulsions containing medium-chain triglycerides (MT), but not by long-chain triglycerides (LT) or structured lipids (SL). However, like PMA, all emulsions decreased the STZ induced [Ca(2)+](i) plateau and all activated purified PKC, suggesting that only MT emulsions activate PKC in the context of the intact cell. MT, like PMA, evoked a leftward shift of the dose-response curve for the STZ-induced [Ca(2)+](i) rise, indicating PKC-dependent sensitization of neutrophils for stimulation by STZ. This study is the first to show that nutritional lipids distinctively modulate cellular signaling and stimulation of neutrophils through effects on calcium mobilization and PKC activation: i) MT emulsions sensitize neutrophils for STZ in a PKC-dependent manner, and ii) MT, LT, and SL emulsions all reduce the stimulatory effect of STZ in a nonspecific manner. -- Wanten, G., S. van Emst-de Vries, T. Naber, and P. Willems. Nutritional lipid emulsions modulate cellular signaling and activation of human neutrophils. J. Lipid Res. 2001. 42: 428--436. PMID- 11254756 TI - Cholesterol metabolism in primary biliary cirrhosis during simvastatin and UDCA administration. AB - Little is known about the effects of cholesterol-lowering agents in hypercholesterolemic patients with primary biliary cirrhosis (PBC). The aim of this study was to compare the changes induced by simvastatin and ursodeoxycholic acid (UDCA) on cholesterol metabolism in patients with PBC and preserved liver function. Six patients with PBC were administered simvastatin (40 mg/day) for 30 days and, after a washout period of 30 days, ursodeoxycholic acid (600 mg/day) for 30 days. Serum levels of lathosterol, campesterol, 7 alpha hydroxycholesterol, and 27-hydroxycholesterol were measured by gas chromatography mass spectrometry. During simvastatin administration, reduction of cholesterol levels (34% in 30 days) was paralleled by the decrease of lathosterol (55%), whereas concentrations of campesterol and of the two hydroxysterols were not substantially modified. During ursodeoxycholic acid administration, a trend toward a decrease of serum cholesterol concentrations was observed after only one year of treatment, and these changes were paralleled by the decrease of campesterol serum levels. Both simvastatin and UDCA were well tolerated, and a reduction of serum liver enzyme levels occurred with the latter. Simvastatin proved to be safe and effective in reducing serum cholesterol levels in patients with PBC by an inhibitory effect on cholesterol synthesis occurring within 24 h. -Del Puppo, M., M. Galli Kienle, A. Crosignani, M. L. Petroni, B. Amati, M. Zuin, and M. Podda. Cholesterol metabolism in primary biliary cirrhosis during simvastatin and UDCA administration. J. Lipid Res. 2001. 42: 437--441. PMID- 11254757 TI - GPI-specific phospholipase D associates with an apoA-I- and apoA-IV-containing complex. AB - Glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) is abundant in serum and associates with high density lipoproteins (HDL). We have characterized the distribution of GPI-PLD among lipoproteins in human plasma. Apolipoprotein (apo)-specific lipoproteins containing apoB (Lp[B]), apoA-I and A-II (Lp[A-I, A II]), or apoA-I only (Lp[A-I]) were isolated using dextran sulfate and immunoaffinity chromatography. In six human plasma samples with HDL cholesterol ranging from 39 to 129 mg/dl, 79 +/- 14% (mean +/- SD) of the total plasma GPI PLD activity was associated with Lp[A-I], 9 +/- 12% with Lp[A-I, A-II], and 1 +/- 1% with Lp[B]; and 11 +/- 10% was present in plasma devoid of these lipoproteins. Further characterization of the GPI-PLD-containing lipoproteins by gel-filtration chromatography and nondenaturing polyacrylamide and agarose gel electrophoresis revealed that these apoA-I-containing particles/complexes were small (8 nm) and migrated with pre-beta particles on agarose electrophoresis. Immunoprecipitation of GPI-PLD with a monoclonal antibody to GPI-PLD co-precipitated apoA-I and apoA IV but little or no apoA-II, apoC-II, apoC-III, apoD, or apoE. In vitro, apoA-I but not apoA-IV or bovine serum albumin interacted directly with GPI-PLD, but did not stimulate GPI-PLD-mediated cleavage of a cell surface GPI-anchored protein. Thus, the majority of plasma GPI-PLD appears to be specifically associated with a small, discrete, and minor fraction of lipoproteins containing apoA-I and apoA IV. -- Deeg, M. A., E. L. Bierman, and M. C. Cheung. GPI-specific phospholipase D associates with an apoA-I- and apoA-IV-containing complex. J. Lipid Res. 2001. 42: 442--451. PMID- 11254758 TI - Regulation of sphingolipid and glycosphingolipid metabolism in extrahepatic tissues by endotoxin. AB - The host response to infection and inflammation is associated with multiple alterations in lipid metabolism. We have shown that endotoxin [lipopolysaccharide (LPS)] stimulates hepatic sphingolipid synthesis and increases ceramide and glucosylceramide (GlcCer) content in circulating lipoproteins in Syrian hamsters. LPS also increases the activity and mRNA levels of serine palmitoyltransferase (SPT) and GlcCer synthase, the committed enzymes in sphingolipid and glycosphingolipid (GSL) synthesis, respectively, in the liver. To determine whether sphingolipid and GSL metabolism are regulated in other tissues during the host response to infection, we examined the effect of LPS on the regulation of SPT and GlcCer synthase in extrahepatic tissues in Syrian hamsters. LPS significantly increased SPT activity in spleen and kidney after 16 h of treatment, but had no effect on SPT activity in lung and brain, suggesting that the effect of LPS on sphingolipid metabolism is tissue specific. LPS also increased SPT mRNA levels in spleen and kidney by approximately 3-fold, suggesting that the increase in SPT activity is due to an increase in SPT mRNA expression. LPS significantly increased GlcCer synthase activity in spleen and kidney, and produced 4- and 15-fold increases in GlcCer synthase mRNA levels in spleen and kidney, respectively. LPS treatment increased GlcCer content by 1.3 fold in spleen and by 6.2-fold in kidney. LPS also increased the content of ceramide trihexoside by 1.7-fold in spleen. These results suggest that LPS regulates sphingolipid and GSL metabolism in spleen and kidney. An increase in GSL metabolites in spleen and kidney during the host response to infection and inflammation may be required for modulation of immune responses and regulation of cell growth. -- Memon, R. A., W. M. Holleran, Y. Uchida, A. H. Moser, C. Grunfeld, and K. R. Feingold. Regulation of sphingolipid and glycosphingolipid metabolism in extrahepatic tissues by endotoxin. J. Lipid Res. 2001. 42: 452- 459. PMID- 11254759 TI - Adenovirus transduction of 3T3-L1 cells. AB - 3T3-L1 cells offer an excellent model system for studies of differentiation and biochemistry of fat cells. However, these cells are limited in their utility by the low efficiency with which DNA can be introduced by transfection. Gene delivery by viral vectors, particularly adenovirus, has proven a powerful means for introduction of genes into certain cell types. Furthermore, adenovirus transduction has been used to study mechanisms involved in the differentiation of 3T3-L1 cells into mature fat cells. We show in this study that 3T3-L1 cells are inefficiently transduced by adenovirus. The potential advantages offered by adenovirus transduction led us to examine methods designed to enhance transduction of 3T3-L1 cells by adenovirus. Of these methods, polylysine-mediated enhancement demonstrates considerable promise because it permits up to 100% of cells to be transduced and because it does not inhibit differentiation of 3T3-L1 cells. -- Orlicky D. J., and J. Schaack. Adenovirus transduction of 3T3-L1 cells. J. Lipid Res. 2001. 42: 460--466. PMID- 11254760 TI - Neurology in practice: another supplement: why? PMID- 11254761 TI - The clinical impact of epilepsy genetics. AB - An overemphasis on molecular genetic advances in epilepsy is in danger of missing the major contribution that clinical genetic studies make in predicting the likely benefit of molecular research efforts. Genetic epidemiology, twin, and family studies suggest that some individual epilepsy genes raise the risk for developing many different types of epilepsy but that specific combinations of these genes determine specific epilepsy phenotypes. Experimental data show how differences in drug response can result from inherited differences in drug sensitivity and there is emerging data on the genetic basis of harmful adverse drug reactions and teratogenesis. These developments predict a future in which epilepsy is ultimately classified on the basis of its oligogenic architecture, effective treatments are tailored to the appropriate patient and harmful adverse drug reactions avoided in those who are sensitive. PMID- 11254762 TI - Magnetic resonance, magnetisation transfer, and diffusion weighted imaging correlates of optic nerve, brain, and cervical cord damage in Leber's hereditary optic neuropathy. PMID- 11254763 TI - The outcome of epilepsy surgery. PMID- 11254765 TI - Magnetic resonance imaging, magnetisation transfer imaging, and diffusion weighted imaging correlates of optic nerve, brain, and cervical cord damage in Leber's hereditary optic neuropathy. AB - OBJECTIVES: Leber's hereditary optic neuropathy (LHON) is a mitochondrial disease leading to bilateral loss of central vision and severe optic nerve atrophy. A subtype of LHON presents additional clinical and MRI aspects indistinguishable from those of multiple sclerosis (MS) (LHON-MS). In patients with LHON or LHON MS, an assessment was made of (a) the severity of optic nerve damage, using MRI and magnetisation transfer imaging (MTI), and (b) the presence and extent of macroscopic and microscopic pathology in the brain and cervical cord, using MRI and MT ratio (MTR) and mean diffusivity (D) histogram analysis. METHODS: Ten patients with LHON, four with LHON-MS, and 20 age and sex matched healthy controls were studied. For the optic nerve and the brain, dual-echo turbo spin echo (TSE), T1 weighted spin echo, and MT images were obtained. For the brain, fast fluid attenuated inversion recovery (fast FLAIR) and diffusion weighted images were also obtained. For the cervical cord, fast short tau inversion recovery (STIR) and MT images were obtained. The volume and the average MTR value of both the optic nerves were measured. MTR and histograms of the normal appearing brain tissue (NABT) and MTR histograms of the whole cervical cord tissue were created. RESULTS: The mean values of optic nerve volumes and MTR were significantly lower in patients with LHON than in healthy controls. Mean NABT-MTR histogram peak height was significantly lower in patients with LHON than in controls, whereas no significant difference was found for any of the cervical cord MTR histogram derived measures. Average diffusivity (D) was higher in patients with LHON than in controls. Optic nerve volume and MTR value and mean NABT-MTR were lower in patients with LHON-MS than in those with LHON. CONCLUSIONS: The severity of optic nerve pathology in LHON is measurable in vivo using MRI and MTI. MTR and histogram analysis suggests that microscopic brain damage occurs in LHON and that it is more severe in the MS-like form of the disease. PMID- 11254764 TI - Intraspinal steroids: history, efficacy, accidentality, and controversy with review of United States Food and Drug Administration reports. PMID- 11254766 TI - Predictors of outcome and pathological considerations in the surgical treatment of intractable epilepsy associated with temporal lobe lesions. AB - OBJECTIVES: To evaluate the influence of clinical, investigative, and pathological factors on seizure remission after temporal lobectomy for medically intractable epilepsy associated with focal lesions other than hippocampal sclerosis. METHODS: From a series of 234 consecutive "en bloc" temporal resections for medically intractable epilepsy performed between 1976 and 1995, neuropathological examination disclosed a focal lesion in 80. The preoperative clinical, neuropsychological, interictal EEG, and neuroimaging characteristics of these patients were assembled in a computerised database. The original neuropathological material was re-examined for lesion classification and completeness of removal. The presence of additional cortical dysplasia and mesial temporal sclerosis was also noted. Survival analysis was performed using Kaplan Meier curves and actuarial statistics. Logistic regression analysis was used to establish the independent significance of the clinical variables. RESULTS: The probability of achieving a 1 year seizure remission was 71% by 5 years of follow up. Factors predicting a poor outcome on multivariate analysis included the need for special schooling and a long duration of epilepsy. Generalised tonic-clonic seizures, interictal EEG discharges confined to the resected lobe, demonstration of the lesion preoperatively on CT, and complete histological resection of the lesion were not predictive of outcome. Neuropsychological tests correctly predicted outcome in left sided cases but apparently congruent findings in right sided resections were associated with a poor outcome. Pathological reclassification established the dysembryoplastic neuroepithelial tumour as the commonest neoplasm (87%) in this series, with a significantly better seizure outcome than for developmental lesions, such as focal cortical dysplasia. CONCLUSIONS: The findings highlight the importance of dysembryoplastic neuroepithelial tumour in the pathogenesis of medically refractory lesional temporal lobe epilepsy and the prognostic significance of preoperative duration of epilepsy emphasises the need for early recognition and surgical treatment. Cognitive and behavioural dysfunction, however, is associated with a lower seizure remission rate, independent of duration of epilepsy. PMID- 11254767 TI - Transient lesion in the splenium of the corpus callosum: three further cases in epileptic patients and a pathophysiological hypothesis. AB - OBJECTIVE: Focal lesions limited to the splenium of the corpus callosum (SCC) are rare and little is known about their aetiology. Three patients were examined for presurgical evaluation in epilepsy with a transient lesion in the SCC and a pathophysiological hypothesis is presented. METHODS: Three patients were identified with a circumscribed lesion in the centre of the corpus callosum. Follow up MRI was performed, the medical records examined retrospectively, and the literature reviewed. RESULTS: The patients showed identical lesions in the SCC with reduced T1 and increased T2 signal intensity and an unaffected marginal hemline of a few mm. Patients were asymptomatic and control MRIs showed complete normalisation within 2 months. Patients had been treated with antiepileptic drugs (AEDs) without signs of toxicity. In all patients AEDs were rapidly reduced for diagnostic purposes, but only one had psychomotor seizures, 5 days before imaging. CONCLUSIONS: A transient lesion in the SCC has so far only been described in 13 patients with epilepsy and has been interpreted either as reversible demyelination due to AED toxicity or transient oedema after secondary generalised seizures. The data confirm neither of these hypotheses. A transient lesion in the SCC seems to be a non-specific end point of different disease processes leading to a vasogenic oedema. This suggests, in these patients, a multifactorial pathology triggered by transient effects of AEDs on arginine vasopressin and its function in fluid balance systems in a condition of vitamin deficiency. The complete and rapid reversibility in all cases without specific intervention is emphasised and any invasive diagnostic or therapeutic approach is discouraged. PMID- 11254768 TI - Deep brain stimulation for the treatment of Parkinson's disease: subthalamic nucleus versus globus pallidus internus. AB - OBJECTIVES: Deep brain stimulation of the basal ganglia has become a promising treatment option for patients with Parkinson's disease who have side effects from drugs. Which is the best target-globus pallidus internus (GPi) or subthalamic nucleus (STN)-is still a matter of discussion. The aim of this prospective study is to compare the long term effects of GPi and STN stimulation in patients with severe Parkinson's disease. PATIENTS AND METHODS: Bilateral deep brain stimulators were implanted in the GPi in six patients and in the STN in 12 patients with severe Parkinson's disease. Presurgery and 3, 6, and 12 months postsurgery patients were scored according to the CAPIT protocol. RESULTS: Stimulation of the STN increased best Schwab and England scale score significantly from 62 before surgery to 81 at 12 months after surgery; GPi stimulation did not have an effect on the Schwab and England scale. Stimulation of the GPi reduced dyskinesias directly whereas STN stimulation seemed to reduce dyskinesias by a reduction of medication. Whereas STN stimulation increased the unified Parkinson's disease rating scale (UPDRS) motor score, GPi stimulation did not have a significant effect. Fluctuations were reduced only by STN stimulation and STN stimulation suppressed tremor very effectively. CONCLUSION: Stimulation of the GPi reduces medication side effects, which leads to a better drug tolerance. There was no direct improvement of bradykinesia or tremor by GPi stimulation. Stimulation of the STN ameliorated all parkinsonian symptoms. Daily drug intake was reduced by STN stimulation. The STN is the target of choice for treating patients with severe Parkinson's disease who have side effects from drugs. PMID- 11254769 TI - Levodopa reversible loss of the Piper frequency oscillation component in Parkinson's disease. AB - OBJECTIVES: Although Parkinson's disease is typically characterised by bradykinesia, rigidity, and rest tremor, the possibility that two additional motor deficits are manifest during small hand muscle activity was explored namely, weakness and abnormal physiological tremor. METHODS: A paradigm previously used in normal subjects reliably records the strength, tremor and surface EMG of index finger abducting contractions against a compliant (elastic) resistance. In addition to the well known physiological tremor at around 10 Hz, there are other co existing peak tremor frequencies at around 20 and 40 Hz; the last of these frequencies corresponds to the range of EMG Piper rhythm. The same technique was used to study parkinsonian patients while on and off dopaminergic medication. RESULTS: The maximum strength of finger abduction produced by first dorsal interosseous contraction was considerably lower when patients were off medication (mean (SD) 6.27 (1.49) N when off v 12.33 (3.64) N when on). There was also a marked reduction in the power of Piper frequency finger tremor (p<0.0005) and EMG (p<0.0005) oscillations that did not simply result from weaker contraction. CONCLUSION: As the components of physiological tremor at higher frequencies are thought to derive from CNS oscillations important in motor control, their loss in parkinsonism in association with severe off symptoms may represent an important pathophysiological link between dopaminergic depletion and parkinsonian motor deficits. PMID- 11254770 TI - Reducing everyday memory and planning problems by means of a paging system: a randomised control crossover study. AB - OBJECTIVES: To evaluate a paging system designed to improve independence in people with memory problems and executive deficits. METHODS: After a successful pilot study, a randomised control trial was conducted involving a crossover design with 143 people aged between 8 and 83 years. All had one or more of the following: memory, planning, attention, or organisation problems. Most had sustained a traumatic head injury or a stroke although a few had developmental learning difficulties or other conditions. The crossover design ensured that some people received a pager after a 2 week baseline whereas others were required to wait for 7 weeks after the baseline before receiving the pager. Participants were assessed at three time periods-namely, at baseline, 7 weeks, and at 14 weeks postbaseline. RESULTS: More than 80% of those who completed the 16 week trial were significantly more successful in carrying out everyday activities (such as self care, self medication, and keeping appointments) when using the pager in comparison with the baseline period. For most of these, significant improvement was maintained when they were monitored 7 weeks after returning the pager. CONCLUSIONS: This particular paging system significantly reduces everyday failures of memory and planning in people with brain injury. PMID- 11254771 TI - Pentagon copying is more impaired in dementia with Lewy bodies than in Alzheimer's disease. AB - OBJECTIVES: In many cases the clinical differentiation of patients with dementia with Lewy bodies (DLB) from those with Alzheimer's disease (AD) has been difficult. Because many neuropsychological studies have reported greater visuospatial/constructional impairment in DLB than in AD, it was determined whether accuracy in copying the interlocking pentagons item on the mini mental state examination (MMSE) may be helpful in distinguishing patients with DLB from those with AD relatively early in the course of the dementia. METHODS: All cases of neuropathologically proved DLB and AD in the Center for Alzheimer Disease and Related Disorders brain bank were retrospectively reviewed, and the first available MMSE for each was retrieved. Only patients with MMSE scores > or = 13 were included, indicating mild to moderate dementia. The patients' copies of the interlocking pentagons were analyzed and graded as acceptable or unacceptable according to the original instructions for grading the MMSE. RESULTS: Seventeen patients with DLB and 27 patients with AD were identified for whom MMSE with copies of the interlocking pentagons were available. Two patients with DLB (MMSEs 22 and 27) drew the pentagons acceptably, by contrast with 16 of the patients with AD (MMSEs 13-28). An unacceptable copy was associated with DLB with a sensitivity of 88% and a specificity of 59% (p = 0.002). CONCLUSIONS: For patients with MMSE scores > or = 13, an inability to accurately copy the pentagons suggests that the diagnosis is more likely DLB than AD. The results confirm the work of others on visuospatial/constructional impairment in DLB and indicate that this feature may be helpful in its diagnosis. PMID- 11254772 TI - Gamma knife radiosurgery of recurrent central neurocytomas: a preliminary report. AB - OBJECTIVES: A series of three recurrent central neurocytomas treated by gamma knife radiosurgery (GKRS), which were initially totally resected, are described. Up to now, no reports exist on this treatment modality for this rare tumour entity. METHODS: Three male patients, aged between 20 and 25 years, presented with large intraventricular tumours. Total tumour removal was achieved by a single surgical procedure (one patient) or two operations (two patients). Neuropathological investigation showed a central neurocytoma, immunohistochemically all three tumours expressed a neuronal antigenic profile typical for neurocytomas, and the MIB-1 proliferation index ranged from 2.4% to 8.7%. Each patient experienced a tumour recurrence after 5 to 6 years. The recurrence was multifocal in two and a singular tumour mass in one patient. Gamma knife radiosurgery was performed. The tumours were enclosed within the 30% to 60% isodoseline, and delivered a tumour marginal dose of 9.6 to 16 Gy. During the follow up period, the patients were tested clinically and the volume of the tumours was measured on MRI. RESULTS: Within follow up periods of 1 to 5 years, control MRI showed a significant decrease of the tumour mass in all cases. None of the patients developed new neurological symptoms after GKRS. Two patients returned to work in their previous employment, whereas one patient remained permanently disabled due to a pre-existing visual impairment and abducens palsy. CONCLUSION: GKRS proved to be a useful tool in the treatment of recurrent central neurocytomas. Tumour control and even tumour shrinkage can be achieved with a single procedure and a low risk of morbidity. PMID- 11254773 TI - Clinical trials of multiple sclerosis monitored with enhanced MRI: new sample size calculations based on large data sets. AB - OBJECTIVE: A new parametric simulation procedure based on the negative binomial (NB) model was used to evaluate the sample sizes needed to achieve optimal statistical powers for parallel groups (with (PGB) and without (PG) a baseline correction scan). It was also used for baseline versus treatment (BVT) design clinical trials in relapsing-remitting (RR) and secondary progressive (SP) multiple sclerosis (MS), when using the number of new enhancing lesions seen on monthly MRI of the brain as the measure of outcome. METHODS: MRI data obtained from 120 untreated patients with RRMS selected for the presence of MRI activity at baseline, 66 untreated and unselected patients with RRMS, and 81 untreated and unselected patients with SPMS were fitted using an NB distribution. All these patients were scanned monthly for at least 6 months and were all from the placebo arms of three large scale clinical trials and one natural history study. The statistical powers were calculated for durations of follow up of 3 and 6 months. RESULTS: The frequency of new enhancing lesions in patients with SPMS was lower, but not significantly different, from that seen in unselected patients with RRMS. As expected, enhancement was more frequent in patients with RRMS selected for MRI activity at baseline than in the other two patient groups. As a consequence, the estimated sample sizes needed to detect treatment efficacy in selected patients with RRMS were smaller than those of unselected patients with RRMS and those with SPMS. Baseline correction was also seen to reduce the sample sizes of PG design trials. An increased number of scans reduced the sample sizes needed to perform BVT trials, whereas the gain in power was less evident in PG and PGB trials. CONCLUSION: This study provides reliable estimates of the sample sizes needed to perform MRI monitored clinical trials in the major MS clinical phenotypes, which should be useful for planning future studies. PMID- 11254774 TI - Demyelinating disorder of the central nervous system occurring in black South Africans. AB - OBJECTIVES: To investigate the nature and cause in eight black South African patients of a recurrent (multiphasic), remitting, and relapsing demyelinating disease of the CNS. METHODS: The clinical and laboratory investigations and radiological manifestations of these patients were documented. RESULTS: Each patient had two or more acute attacks of demyelinating disease affecting the CNS. The clinical presentations of the patients were predominantly those of multiphasic neuromyelitis optica (NMO). Brain MRI in these patients showed features consistent with those described for acute disseminated encephalomyelitis (ADEM), as well as lesions that are described in multiple sclerosis. There was involvement of the corpus callosum in addition to typical ADEM lesions. Laboratory investigations excluded all other known causes of multiphasic CNS demyelination. Oligoclonal antibodies were not detected in these patients at any time. The patients were all from a population with a low risk for MS (black South Africans). CONCLUSION: The patients described here represent a new phenotypic expression of a recurrent (multiphasic), steroid sensitive, inflammatory demyelinating disorder of the CNS occurring in black South Africans. The disorder is either a distinct inflammatory demyelinating disorder of the CNS of as yet unknown aetiology, or a varied form of MS (ADEM/NMO) occurring in a population with a low risk (where the genetic trait and environmental risk factors for MS do not exist) for MS. PMID- 11254776 TI - Wilson's disease presenting in a family with an apparent dominant history of tremor. AB - A patient with Wilson's disease is described who presented with dystonic tremor in a family with an apparent dominant history of tremor. Subsequent investigation showed that the patient's mother had essential tremor, with molecular analysis of the ATP7B gene excluding the possibility of pseudodominant inheritance. This case highlights the importance of considering the possibility of Wilson's disease in every young patient with a movement disorder, even where the clinical picture does not suggest a recessively inherited disorder. PMID- 11254775 TI - Acute functional reorganisation of the human motor cortex during resection of central lesions: a study using intraoperative brain mapping. AB - OBJECTIVES: Brain plasticity is supposed to allow the compensation of motor function in cases of rolandic lesion. The aim was to analyse the mechanisms of functional reorganisation during surgery in the central area. METHODS: A motor brain mapping was performed in three right handed patients without any neurological deficit, operated on for a slow growing lesion near the rolandic region (two precentral resected under general anaesthesia and one retrocentral removed under local anaesthesia to allow also sensory mapping) using intraoperative direct electrical stimulations (5 mm space tips bipolar stimulator probe, biphasic square wave pulse current: 1 ms/phase, 60 Hz, 4 to 18 mA). RESULTS: For each patient, the motor areas of the hand and forearm in the primary motor cortex (M1) were identified before and after lesion removal with the same stimulation parameters: the same eloquent sites were found, plus the appearance after resection of additional sites in M1 inducing the same movement during stimulations as the previous areas. CONCLUSIONS: Multiple cortical representations for hand and forearm movements in M1 seem to exist. In addition, the results demonstrate the short term capacity of the brain to make changes in local motor maps, by sudden unmasking after tumour resection of a second redundant site participating in the same movement. Finally, it seems not necessary for the whole of the redundant sites to be functional to provide normal movement, a concept with potential implications for surgery within the central region. PMID- 11254777 TI - "The other" Babinski's sign: paradoxical raising of the eyebrow in hemifacial spasm. PMID- 11254778 TI - Reduction of striatal glucose metabolism in McLeod choreoacanthocytosis. AB - McLeod syndrome is a distinct form of neuroacanthocytosis. Its defining feature is the depression of erythrocyte Kell antigens. The underlying X chromosomal mutations cause a dysfunction of an erythrocyte membrane protein Kx. A choreatic movement disorder with caudate atrophy in CT and MRI has been reported in McLeod syndrome later in the course of the disease. Positron emission tomography with 18F-deoxyglucose (FDG) was performed in two unrelated affected men. In the older patient, progressive chorea was seen from the 5th decade. In the second patient there were no signs of a movement disorder at the age of 28. Positron emission tomography disclosed a reduction of the striatal FDG uptake in both patients, with accentuation in patient 1. Frontal lobe metabolism was not affected. Basal ganglia dysfunction with early impairment of striatal glucose metabolism thus seems obligatory for McLeod syndrome, as found in other forms of chorea with or without acanthocytosis. PMID- 11254779 TI - Sir David Bruce (1855-1931) and Thermistocles Zammit (1864-1935). PMID- 11254780 TI - Combined functional magnetic resonance imaging and diffusion tensor imaging demonstrate widespread modified organisation in malformation of cortical development. AB - A patient with a mild left hemiparesis and a malformation of cortical development in the right hemisphere was investigated with fMRI (functional magnetic resonance imaging) and DTI (diffusion tensor imaging). The motor cortex was studied using a finger tapping fMRI experiment. The fibre orientation was studied by displaying the principal eigenvector of the diffusion tensor in the spatially normalised brain of the patient and of control subjects. In addition, the anisotropy (directionality) of water diffusion of the patient was statistically compared with control subjects. The malformation was located in the right central region in the expected position of the motor cortex. fMRI showed activation anterior and posterior to the malformation. DTI disclosed that fibres with rostrocaudal orientation, presumably representing the pyramidal tract, were deviating from their normal orientation and passing around the malformation. There were widespread regions of reduced anisotropy affecting both hemispheres. In conclusion, fMRI and DTI provided concordant information showing widespread modified functional and structural organisation including regions which appeared normal on standard imaging. PMID- 11254781 TI - Neurology in ancient faces. AB - BACKGROUND: Clinical paleoneurology is almost non-existent, but recognition of neurological diseases in ancient people might be possible by scrutinising portraits apparently representing people as they appeared in life. METHODS: About 200 mummy portraits painted in colour at the beginning of the first millennium were examined. Thirty two skulls excavated at Hawara in the Fayum (northern Egypt), where most of the portraits were found were measured, and nine caliper measures on each side of the skulls were taken. The right/left ratios were statistically analyzed by analysis of variance (ANOVA). One skull was subjected to 3D CT scanning and transilluminated. RESULTS: Two patients were found with progressive facial hemiatrophy (Parry-Romberg syndrome), three with deviations of the visual axes (tropia) and one with oval pupils (corectopia). CONCLUSIONS: Clinical paleoneurology is possible in the absence of a living nervous system. The patients probably had focal epilepsy, hemiplegic migraine, and autonomic nervous system dysfunction. PMID- 11254782 TI - Diffusion imaging shows abnormalities after blunt head trauma when conventional magnetic resonance imaging is normal. AB - The investigation and management of patients after head injury must include the accurate and complete identification of cerebral damage. Using diffusion tensor imaging, abnormalities of diffusion in patients with head injuries and unremarkable MRI have been shown for the first time. PMID- 11254783 TI - Inflammatory cytokines in subarachnoid haemorrhage: association with abnormal blood flow velocities in basal cerebral arteries. AB - Subarachnoidal release of inflammatory cytokines (interleukin (IL)-1beta, IL-6, and tumour necrosis factor (TNF)-alpha) was characterised in 35 patients with subarachnoid haemorrhage (SAH) and control subjects and compared with development of complicating haemodynamic abnormalities in basal cerebral arteries and clinical outcome. Serial analysis allowed the observation of a subacute response profile of these key mediators of inflammation in the subarachnoidal space. This compartmentalised inflammatory host response was closely associated in time and extent with development of increased blood flow velocities in the basal cerebral vessels as recorded by transcranial Doppler sonography. Moreover, intrathecal secretion of inflammatory cytokines was significantly increased in patients with poor clinical outcome. Together, these findings suggest a role of excessive compartmentalised inflammatory host response in pathogenesis of cerebrovascular complications after SAH. PMID- 11254784 TI - Treatment of sialorrhoea with ultrasound guided botulinum toxin type A injection in patients with neurological disorders. AB - OBJECTIVES: To investigate the safety and efficacy of ultrasound guided botulinum toxin type A (BTX-A) injections into salivary glands for the treatment of sialorrhoea in patients with neurological disorders. METHODS: The parotid and submandibular glands of 10 patients were injected with BTX-A using ultrasound guidance. Before injection, the baseline rate of salivation was assessed using a visual analogue scale. Postinjection, assessments were repeated at regular intervals for up to 1 year. RESULTS: Of the 10 patients treated, nine (90%) reported a subjective reduction in salivation post-treatment and one patient (10%) found no improvement. Visual analogue scale scores showed a reduction of 55% in the mean rate of salivation for all patients and a reduction of 60.8% for the group of responders. No serious adverse events occurred and no procedure related complications were reported. CONCLUSIONS: This is the first study to report (1) the injection of BTX-A (BOTOX) into both parotid and submandibular glands, and (2) the use of ultrasound guidance during the administration of BTX-A into salivary glands. The results suggest that the technique is safe and that BTX A injections are effective for the treatment of sialorrhoea in patients with neurological disorders. PMID- 11254785 TI - Rub epilepsy: a somatosensory evoked reflex epilepsy induced by prolonged cutaneous stimulation. AB - To delineate rub epilepsy--a type of reflex epilepsy induced by prolonged or repetitive cutaneous stimulation in a circumscribed area of the body--three cases are presented, as well as one of tooth brushing epilepsy for comparison. In all three cases of rub epilepsy, cutaneous stimuli in a particular body area on the left side initially induced a sensory jacksonian march in the middle of, or in close vicinity to, the trigger zone, which led to subsequent unilateral tonic contractions with intact consciousness. By contrast, a motor jacksonian seizure without sensory aura was induced in the patient with tooth brushing epilepsy. A review of cases with rub epilepsy, including those in this paper, disclosed a striking consistency in clinical manifestations. The symptomatology of the induced seizures indicates a propagation of epileptic discharges from the postcentral gyrus to the supplementary motor area. Rub epilepsy is proposed as a separate clinical entity, clearly demarcated from other somatosensory evoked reflex epilepsies such as startle and tooth brushing epilepsy. PMID- 11254786 TI - Measurement of spinal cord area in clinically isolated syndromes suggestive of multiple sclerosis. AB - Atrophy of the spinal cord is known to occur in multiple sclerosis but the cause and the timing of its onset are not clear. Recent evidence suggests that atrophy may start to occur early in the disease. The aim was to determine whether atrophy of the spinal cord could be detected in vivo using MRI techniques, in patients presenting with a clinically isolated syndrome, which in many cases is the earliest clinical stage of multiple sclerosis. The cross sectional area of the spinal cord was measured in 43 patients presenting with a clinically isolated syndrome and 15 matched controls. T2 weighted imaging of the brain was also performed to determine the number and volume of high signal lesions consistent with disseminated demyelination. Both patients and controls were restudied after 1 year. The spinal cord area was significantly smaller in the 74% of patients with an abnormal brain MRI at presentation than in controls (mean areas 73.9 mm(2) and 78.1 mm(2) respectively, p=0.03). No significant difference was found in the spinal cord area between controls and patients with normal baseline brain imaging. The annual rate of change in patients did not differ significantly from controls. In conclusion, the finding of a smaller cord area in the subgroup of patients with clinically isolated syndrome with the highest risk of developing multiple sclerosis-that is, with an abnormal brain MRI, suggests that atrophy has developed in some patients with multiple sclerosis even before their first clinical symptoms. However, the lack of a detectable change in cord area over 1 year of follow up contrasts strikingly with the results of an earlier study of patients with relapsing-remitting multiple sclerosis, suggesting that the rate of atrophy increases as the disease becomes more established. PMID- 11254787 TI - Disability and quality of life in Charcot-Marie-Tooth disease type 1. AB - OBJECTIVES: Charcot-Marie-Tooth disease type I (CMT1) is a hereditary sensorimotor neuropathy causing variable degrees of handicap. The risk for relevant disability in respect to genetic counselling is unknown. An attempt was made to define it. METHODS: Disability and ambulation of 50 patients with CMT1 were scored by the Hauser ambulation index score and the Rankin scale. Rankin score 2 was subdivided into 2a (independent without relevant slowness) and 2b (independent, though at the cost of excessive time consumption). The sickness impact profile was assessed and compared with patients 6 months after stroke who were without mental deficit. To define at which degree sickness and disability become relevant for genetic counselling, the patients were asked whether they would refrain from childbearing if the children were at risk of inheriting a disease that caused as much disability as they experienced themselves. RESULTS: Subdivision of Rankin score 2 was reliable and improved validity. High disability significantly predicted an attitude against childbearing (stepwise logistic regression) only with this subdivision. Thirty six per cent of the patients voted against childbearing. The cut off for relevant disability in respect to childbearing was a Rankin score higher than 2a, which was present in 44% of the patients. Psychosocial impact was comparable with patients with stroke and similar disability. Depression was present in 18% of the patients. CONCLUSION: Subdivision of Rankin score 2 is recommended for the assessment of longstanding disability in neuromuscular disorders. Disability becomes relevant for the attitude towards childbearing as soon as everyday activities become markedly slow (Rankin score 2b). Relevant disability occurred in 44% of the patients. Emotional stress in CMT is similar to that of patients with stroke and comparable disability. PMID- 11254788 TI - Iron overload without the C282Y mutation in patients with epilepsy. AB - To test the hypothesis that iron overload predisposes to epilepsy, transferrin saturation in 130 patients with epilepsy and sex and age matched 128 control subjects without epilepsy were studied. Mean transferrin saturation was significantly higher in the epilepsy group (39.9 (SD 19.6)%) than in the control group (29.1 (SD 14.9)%). Abnormally high transferrin saturations (men>60%, women>48%) were found in 10 patients with epilepsy but in only one subject without epilepsy. Antiepileptic drugs did not affect the transferrin saturation. Of the 11 with abnormally high transferrin saturation, two with epilepsy were heterozygotic for H63D in the haemochromatosis gene but no patient had the C282Y mutation. These results indicate that iron overload other than the C282Y mutation underlies epilepsy. PMID- 11254789 TI - High dose naltrexone for dyskinesias induced by levodopa. AB - Ten patients with Parkinson's disease and levodopa induced dyskinesias (LIDs) took part in this randomised, placebo controlled, double blind, crossover trial to assess the efficacy and tolerability of high dose oral naltrexone for LIDs in Parkinson's disease. Patients received naltrexone (5 mg/kg/day) or placebo for 2.5 weeks with 1 week wash out in between. Dyskinesias and motor function were assessed with a levodopa challenge, unified Parkinson's disease rating scale (UPDRS), the unified dyskinesia rating scale (UDRS), and patient diaries. Eight patients completed the trial. There was a small reduction in LIDs measured by patient diaries with naltrexone (20.5 (SD 24.9)%) compared with placebo (-4.1 (SD 22.6)%), p<0.05, although no difference was found by other subjective or objective measures. Naltrexone was well tolerated and caused no significant differences in UPDRS motor scores or off time. This study suggests that short term therapy with high dose naltrexone (250-350 mg/day) has no or minimal effect on reducing LIDs in Parkinson's disease. PMID- 11254790 TI - Pallidal and thalamic neurostimulation in severe tardive dystonia. AB - A 70 year old woman presented with a 6 year history of medically refractory severe tardive dystonia. After informed consent, a bilateral stereotactic electrode placement targeting the ventral intermediate thalamic nucleus (VIM) and the globus pallidus internus (GPi) was performed. After bilateral stimulation of the GPi, the patient showed a clear and stable improvement of the painful dystonic syndrome within hours. Stimulation of the VIM did not improve the hyperkinetic movements and simultaneous stimulation of both the GPi and the VIM did not result in any additional benefit. The possible pathophysiological mechanisms are discussed. PMID- 11254791 TI - Indicators of rapid clinical recovery in Guillain-Barre syndrome. AB - To elucidate the features of patients with Guillain-Barre syndrome who show markedly rapid clinical recovery, clinical, serological, and electrophysiological data of 80 consecutive patients were reviewed. Antigangliosides, and Campylobacter jejuni and Haemophilus influenzae antibodies were measured by enzyme linked immunosorbent assays. Nine (11%) patients showed rapid recovery (improvement by two or more Hughes grades within 2 weeks). They often had electrodiagnosis of acute motor axonal neuropathy (AMAN; 67%), preserved tendon reflexes (44%), anti-GM1 antibodies (89%), preceding H influenzae infection (44%), and received immunoglobulin treatment (44%). On the other hand six patients with poor prognosis often had AMAN (100%) and anti-GM1 antibody (83%), but a higher incidence of preceding C jejuni infection (83%). It is concluded that patients with Guillain-Barre syndrome with AMAN and anti-GM1 antibodies have either faster or slower recoveries. Among the axonal subgroup of patients with Guillain-Barre syndrome, preserved tendon reflexes, H influenzae infection, and the patient having received immunoglobulin treatment may be indicators of rapid recovery. PMID- 11254792 TI - Neurology in practice: cerebrovascular disease. PMID- 11254793 TI - Assessment and investigation of stroke and transient ischaemic attack. PMID- 11254794 TI - Radiology of stroke. PMID- 11254795 TI - Medical management of stroke. PMID- 11254796 TI - Identification and management of difficult stroke and TIA syndromes. PMID- 11254797 TI - 1999 Optident prize and William Houston Medal of the Royal College of Surgeons of Edinburgh. AB - This paper describes the clinical orthodontic treatment of three cases which were awarded the 1999 Optident prize and the William Houston Medal. PMID- 11254798 TI - The integration of functional and fixed appliance treatment. AB - This article describes a functional appliance system to correct Class II problems, which is clipped on to bands, cemented to the teeth. This appliance has several advantages, as the patient cannot remove it. It acts on the teeth and jaws for 24 hours each day, patient co-operation is not a problem, and as a result the treatment time is short. Any fixed appliance system can be added while the functional phase is being completed so allowing full integration of the two treatment systems. PMID- 11254799 TI - A case of anterior open bite developing during adolescence. AB - Imaging studies have reported on the relationship between temporomandibular joint (TMJ) degeneration and facial deformity. These studies have suggested that mandibular growth is affected by TMJ degeneration, resulting in altered skeletal structure as mandibular retrusion. However, there are very few longitudinal case reports on TMJ osteoarthrosis (OA). Progressive open bite occurred in an adolescent patient with TMJ OA. Cephalometric analysis showed a downward and backward rotated mandible, and a labial inclination of the upper incisor. Magnetic resonance imaging showed internal derangement without reduction and erosion in the right and the left condyles. Although the cause of open bite is unclear in this case, tongue thrusting, and internal derangements in the temporomandibular joint were suspected as causes of the open bite. PMID- 11254800 TI - A modified approach for obtaining cephalograms in the natural head position. AB - Cephalograms taken in the Natural Head Position (NHP), and related to the true vertical and the true horizontal reference planes should logically replace the planes used in conventional cephalometry. This has not happened because of the difficulties in radiographically recording the NHP. This paper presents a modified approach to capture the true vertical reference line on the patients' face itself in NHP, which is then transferred to the conventional lateral cephalogram. PMID- 11254802 TI - Orthodontic treatment for disabled children--a survey of patient and appliance management. AB - The objective of this article was to investigate the management problems encountered during the orthodontic treatment of children with disability, and took the form of a retrospective analysis. The investigation took place at the Center for the Treatment of Cranio-facial Disorders, Department of Orthodontics, Hebrew University Hadassah School of Dental Medicine, Jerusalem, Israel, between years 1989 and 1997. The subjects were the 37 children with mental and/or physical disability whose orthodontic treatment was either completed or nearly completed, whose parents were given a questionnaire. Thirty-five patients responded with a mean age of 13 years (range 7-21 years), representing 94.6 per cent of the sample. Most of the patients (94.3 per cent) were able to conclude the orthodontic treatment and 91.4 per cent of the parents reported that the added responsibilities were either negligible or bearable. In 63 per cent of the children, compliance increased during the treatment as anxiety decreased. The problems encountered with fixed appliances were generally more severe than with removable appliances. The two major obstacles were attendance at frequent and regular intervals (37.1 per cent) and maintaining an appropriate level of oral hygiene (37.1 per cent). Children with a disability are able and willing to undergo orthodontic treatment. Recommendations intended to facilitate management are presented. PMID- 11254801 TI - A validated finite element method study of orthodontic tooth movement in the human subject. AB - The aim of the study was to develop a 3D computer model of the movement of a maxillary incisor tooth when subjected to an orthodontic load. A novel method was to be developed to directly and accurately measure orthodontic tooth movement in a group of human volunteers. This was to be used to validate the finite element based computer model. The design took the form of a prospective experiment at a laboratory at the University of Wales in 1996/7. A laser apparatus was used to sample tooth movement every 0.01 seconds over a 1-minute cycle for 10 healthy volunteers, whilst a constant 0.39 N load was applied. This process was repeated on eight separate occasions and the most consistent five readings taken for each subject. Data were used to calculate the physical properties of the periodontal ligament (PDL). The data gleaned by this method were used to validate the 3D FEM model. This was formed of 15,000 four-noded tetrahedral elements. Tooth displacements ranged from 0.012 to 0.133 mm. An appropriate elastic modulus of 1 N/mm(2) and Poisson's Ratio of 0.45 was derived for the PDL. Strain analysis, using the model, suggested that a maximum PDL strain of 4.77 x 10(-3) was recorded at the alveolar crest, while the largest apical strain recorded was 1.55 x 10(-3). The maximum strains recorded in the surrounding alveolar bone were 35 times less than for the PDL. A novel method for direct measurement of PDL physical properties in the human subject has been developed. The validated FEM model lends further evidence that the PDL is the main mediator of orthodontic tooth movement. PMID- 11254803 TI - The use of the Index of Orthodontic Treatment need (IOTN) in a school population and referred population. AB - The aim of this study is to assess the need for orthodontic treatment in a Turkish school population and a group of population referred for orthodontic treatment. The study groups were 250 school children, 11-14 years of age, and 250 patients, 11-14 years of age, referred to the department of orthodontics. The Index of Orthodontic Treatment Need (IOTN) was used by two examiner in order to estimate the treatment need. The differences between the IOTN values for the boys and girls were also not statistically significant in both groups. When the dental health component of IOTN is considered, 38.8 per cent of Turkish school population showed great need treatment, 24.0 per cent moderate need treatment and slight or no need was 37.2 per cent. On the other hand, the referred population represented an 83.2 per cent great need treatment, 12.0 per cent moderate need treatment, 4.8 per cent no need treatment according to the DHC. The AC of IOTN in school population resulted in 4.8 per cent great need, 4.8 per cent moderate need, 90.4 per cent no need. These percentage were 36.8 per cent great need, 17.6 per cent moderate need, 45.2 per cent no need in referred population. Grade 8 was 28.8 per cent out of the 36.8 per cent great need percentage in referred population. Therefore, it can be concluded that the ectopic canines were the driving factor for the referred population. PMID- 11254804 TI - Dentoskeletal morphology in children with juvenile idiopathic arthritis compared with healthy children. AB - The aim of this study was to evaluate the dentoskeletal relationships in children with juvenile idiopathic arthritis (JIA) compared to healthy children without significant differences in relation to age and sex, by means of lateral cephalometric radiographs. Cephalometric, as well as dental panoramic radiographs were taken of 66 JIA children (27 males and 39 females; age range: 6-19 years; mean age: 11.9 years). The control group consisted of 37 healthy children unaffected by JIA seeking orthodontic treatment, with Class I occlusion (17 males and 20 females; age range: 7.5-17 years; mean age: 11.9 years). All cephalometric landmarks were identified and digitized, and calculations were performed by means of a computerized cephalometric system. The cephalometric findings indicated a tendency towards retrognathism with a short mandible. The lower facial height was increased and the growth pattern of the face was biased towards the vertical direction (clockwise, i.e. with a tendency to open bite) and the interincisal angle was less in the JIA children compared to the healthy children. These findings were in general more pronounced by the JIA children with polyarticular type of the disease as well as with affected condyles. Our study indicated that the dentoskeletal morphology in children with JIA presented some special characteristics when compared to healthy children, which could be attributed to the effects of the disease. PMID- 11254805 TI - An ex vivo study to investigate bond strengths of different tooth types. AB - This study aimed to identify the presence and pattern of differences in ex vivo shear bond strength between tooth types when bonding orthodontic brackets using Right-On, and took the form of a prospective laboratory study of bond strength on different tooth types, at the Newcastle University Dental School Materials Science Laboratory, 1997-1999. Ex vivo bond strength testing was undertaken using the technique described by Fox et al. (BJO 18, 125-130, 1991) on a total of 120 extracted incisor, canine, and premolar teeth of each dental arch. Analysis was by one-way ANOVA with Tukey's pairwise comparisons, and by Weibull Analysis. Shear stress to failure (measured in MPa) was recorded on Instron 5567 universal testing machine. Significant differences in mean bond strength existed between different tooth-type series. Canine (upper 12.3, lower 12.1) and premolar (upper 11.9, lower 10.9) teeth exhibited higher strengths than incisors (upper 6.9, lower 9.0). The results of this study confirm that ex vivo bond strength is not uniform across all teeth. PMID- 11254807 TI - An orthodontic patient administration system (OPAS) for complete departmental management. AB - There is a requirement for effective management and audit in today's hospital environment. This paper discusses some of the principal requirements of a computer program for comprehensive orthodontic department management and describes in detail one system. PMID- 11254808 TI - Functional occlusion: I. A review. AB - The features that constitute an "ideal" functional occlusion have not been conclusively established. Orthodontic treatment has the capacity to change static and functional occlusal relationships fundamentally. In this article, we present the evidence on which features of the occlusion are reported to be detrimental to the teeth and masticatory system Deficiencies in this research area are highlighted, together with the need for prospective longitudinal trials to clarify the requirements of an ideal functional occlusion Based on the existing evidence this paper suggests which occlusal features may be significant in producing an "ideal" functional occlusion As no long-term studies exist to measure the impact of non-ideal occlusal relationships on the dentition, it is debatable whether orthodontic treatment should be prolonged in order to ensure that "ideal" occlusal contacts are achieved As the occlusion tends to "settle" in the period following appliance removal, we propose that it may be more appropriate to examine the functional occlusal relationships after retention has ceased rather than prolong active orthodontic treatment to achieve "ideal" functional occlusal goals. PMID- 11254810 TI - Orthodontics in Nigeria: journey so far and the challenges ahead. AB - The practice of orthodontics in Nigeria has witnessed a gradual, but steady development since its introduction about three decades ago. The undergraduate orthodontic training that was fashioned after the British model has evolved from a concentrated course of lectures to a more clinical/practical orientated programme. The local postgraduate training, however, needs to be restructured and strengthened in order to face the challenges ahead. The importance of constant upgrading of knowledge and skills in all areas of orthodontics is also emphasized. PMID- 11254811 TI - An update on orthodontic manpower in ireland. AB - There has been considerable debate in Europe over the past few years on manpower requirements in orthodontics. In some countries today the need for orthodontic care cannot be accommodated due to lack of professional manpower whereas in others a surplus of orthodontic treatment facilities exists. The aim of the present study was to establish a baseline for orthodontic demographics in the Republic of Ireland. The number of orthodontists currently practising in Ireland was identified together with the number of Irish graduates currently on training programmes. Population figures were obtained from the Central Statistics Office. The orthodontic manpower situation has altered dramatically in the Republic of Ireland over the past 20 years. The number of 12-year-olds per orthodontist has reduced over the past 18 years from 2773 in 1980 to 890 in 1998. The age profile of the orthodontists presently practising in Ireland is low with an expected retirement over the next 20 years of only 28 of the 69 orthodontists identified. This study provides baseline information on orthodontic manpower in Ireland, and will facilitate Ireland's participation in similar or comparative studies in the future. PMID- 11254812 TI - Undergraduate orthodontic & paediatric dentistry education in Europe--the DentEd project. AB - As a result of a European Union funded project (DentEd), a programme of visits to dental schools throughout Europe has been underway since 1998. This report describes the philosophy behind DentEd, gives a brief description of the features of a visitation, and covers the orthodontic and paediatric dentistry teaching as reported in 26 different dental schools in 16 different countries. It is based on a report submitted to DentEd from a small working group that looked at various aspects of educational provision within the two disciplines across Europe. The value of this information to teachers within the two disciplines and to the wider dental community is briefly discussed. The report recommends the adoption of an integrated course for orthodontics and paediatric dentistry. The main objectives are that the student should be able to understand orofacial and psychosocial growth and development of the child, recognize aberrant growth and development, and manage the behaviour of the child, their straightforward preventive, restorative and occlusal needs, and to make appropriate and timely referral. PMID- 11254813 TI - Tobacco control: from concern for the lung to global political action. PMID- 11254814 TI - Nottingham University and British American Tobacco. PMID- 11254815 TI - Statement on malignant mesothelioma in the United Kingdom. PMID- 11254816 TI - Relationship between anxiety, depression, and morbidity in adult asthma patients. AB - BACKGROUND: Symptoms of disease reported by patients reflect the effects of the disease process within the individual and the person's physical and mental ability to tolerate or otherwise cope with the limitations on their functioning. This study examines the relationship between asthma symptoms, disease severity, and psychological status in patients being managed in routine primary healthcare settings. METHODS: One hundred and fourteen subjects from four GP practices, two inner city and two suburban, were studied. Symptoms were assessed by means of the Asthma Quality of Life questionnaire (AQLQ) and a locally devised Q score, and psychological status with the Hospital Anxiety and Depression (HAD) scale. Spirometric values and details of current asthma treatment (BTS asthma guidelines treatment step) were recorded as markers of asthma severity. RESULTS: Symptoms as measured by AQLQ correlated with peak expiratory flow (r(S) = 0.40) and with BTS guidelines treatment step (r(S) = 0.25). Similarly, the Q score correlated with peak expiratory flow (r(S) = 0.44) and with BTS guidelines treatment step (r(S) = 0.42). Similar levels of correlation of forced expiratory volume in one second (FEV(1)) with symptoms were reported. HAD anxiety and depression scores also correlated to a similar extent with these two symptom scores, but there was hardly any correlation with lung function. Logistic regression analysis showed that HAD scores help to explain symptom scores over and above the effects of lung function and BTS guidelines treatment step. Symptoms, depression, and anxiety were higher for inner city patients while little difference was observed in objective measures of asthma. CONCLUSIONS: Asthma guidelines suggest that changing levels of symptoms should be used to monitor the effectiveness of treatment. These data suggest that reported symptoms may be misleading and unreliable because they may reflect non-asthma factors that cannot be expected to respond to changes in asthma treatment. PMID- 11254817 TI - Bone mineral density in subjects with mild asthma randomised to treatment with inhaled corticosteroids or non-corticosteroid treatment for two years. AB - BACKGROUND: Inhaled corticosteroids are clearly beneficial for patients with asthma of moderate severity, but the risks and benefits of using them in patients with milder asthma are less clear. We have compared the change in bone mineral density over 2 years in adults with mild asthma randomised to receive an inhaled corticosteroid or non-corticosteroid treatment. METHODS: Subjects with mild asthma (mean forced expiratory volume in one second (FEV(1)) 86% predicted, mean age 35 years, taking beta agonists only) were randomised to receive inhaled budesonide, inhaled beclomethasone dipropionate, or non-corticosteroid treatment for 2 years in a prospective randomised open study in 19 centres in France, New Zealand, Spain, and the UK. The corticosteroid dose was adjusted according to a written self-management plan. The main outcome measure-change in bone mineral density after 6, 12, and 24 months-was measured "blind". Secondary outcomes included lung function, the relation between change in bone density and inhaled steroid dose and change in biochemical markers of bone metabolism. RESULTS: Of 374 subjects randomised, 239 (64%) completed the study and were included in the analysis. The median daily doses of inhaled budesonide (n=87) and beclomethasone (n=74) were 389 microg and 499 microg, respectively. Subjects treated with an inhaled corticosteroid had better asthma control than those in the reference group (n=78). Change in bone mineral density did not differ between the three groups over the 2 years, nor did it correlate with changes in markers of bone metabolism. The mean change in bone mineral density over 2 years in the budesonide, beclomethasone dipropionate, and reference groups was 0.1%, -0.4%, and 0.4% for the lumbar spine and -0.9%, -0.9%, and -0.4% for neck of the femur. Mean daily dose of inhaled steroid was related to reduction in bone mineral density at the lumbar spine but not at the femoral neck. CONCLUSION: In subjects with mild asthma an inhaled corticosteroid provided better asthma control than alternative non-corticosteroid treatment with no difference in change in bone mineral density over 2 years. The relation between dose of inhaled corticosteroid and change in bone density at the lumbar spine may be due to a direct effect of inhaled corticosteroids on bone. Since inhaled steroid dose is also related inversely to lung function, an effect of asthma severity on bone density was also possible. PMID- 11254818 TI - Adverse effects of oral corticosteroids in relation to dose in patients with lung disease. AB - BACKGROUND: The adverse effects of oral corticosteroids are widely recognised but there are few quantitative data on which to base advice to patients. In a two part cross sectional study we compared adverse effects in patients with lung disease taking oral corticosteroids and control subjects and related the adverse effects to corticosteroid dose in the patient group. METHODS: Data on oral corticosteroid use, lifestyle, fractures, and other possible adverse effects were collected by questionnaire and compared between a community based cohort of patients taking continuous or frequent intermittent oral corticosteroids for asthma, chronic obstructive pulmonary disease, or alveolitis and age and sex matched control subjects. Dose related effects were explored in the corticosteroid group using cumulative dose quartiles and multiple logistic regression. RESULTS: A total of 367 patients (> or = 50 years, 48% female) and 734 control subjects completed the questionnaire. The cumulative incidence of fractures since the time of diagnosis was 23% for patients taking oral corticosteroids and 15% in the control group (odds ratio (OR) 1.8; 95% confidence interval (CI) 1.3 to 2.6). Patients were more likely to have had a fracture of the vertebrae (OR 10; 95% CI 2.9 to 34), hip (OR 6; 95% CI 1.2 to 30), and ribs or sternum (OR 3.2, 95% CI 1.6 to 6.6) than control subjects. They also reported a significant increase in cataracts, use of antacids, muscle weakness, back pain, bruising, oral candidiasis, and having fewer teeth. The effects of oral corticosteroids were dose related: the odds ratio for patients in the highest compared with the lowest cumulative dose quartile (median prednisolone dose 61 g versus 5 g) ranged from 2 for all fractures to 9 for vertebral fractures and bruising. CONCLUSIONS: By quantifying the morbidity associated with the use of oral corticosteroids, this study should help to rationalise their long term use. PMID- 11254819 TI - Controlled low flow off line sampling of exhaled nitric oxide in children. AB - BACKGROUND: The aim of this study was to validate exhaled nitric oxide (eNO) values obtained with an alternative off line, single breath, low flow balloon sampling method against on line sampling according to ERS and ATS guidelines in children who could perform both methods. METHODS: One hundred and twenty seven white children of median age 14.1 years, all pupils of a secondary school, participated in the study. They performed the two different sampling techniques at three different flows of 50, 100, 150 ml/s. Additional measurements were done in random subgroups to determine the influence of the dead space air on eNO values obtained off line by excluding the first 220 ml of exhaled air. All children completed a questionnaire on respiratory and allergic disorders and underwent spirometric tests. RESULTS: The off line eNO values were significantly higher than the on line values at all flows. At 50 ml/s the geometric mean (SE) off line eNO was 18.7 (1.1) ppb and the on line eNO was 15.1 (1.1) ppb (p<0.0001). However, when dead space air was discarded, off line and on line values were similar: at 50 ml/s off line eNO was 17.7 (1.0) ppb and on line eNO 16.0 (1.2) ppb. There was a good agreement between off line eNO values without dead space air and on line eNO: for 50 ml/s the mean on/off line ratio was 0.95 (95% agreement limits 0.63 to 1.27). The off line eNO level at 50 ml/s in 80 children with negative questionnaires for asthma, rhinitis, and eczema was 13.6 (1.0) ppb compared with 33.3 (1.1) ppb in the remaining children with positive questionnaires on asthma and allergy and/or recent symptoms of cold (p<0.0001). CONCLUSIONS: In children, off line assessment of eNO using constant low flow sampling and excluding dead space air is feasible and produces similar results as on line assessment with the same exhalation flow rate. Both sampling methods are sufficiently sensitive to differentiate between groups of otherwise healthy school children with and without self-reported asthma, allergy, and/or colds. We propose that, for off line sampling, similar low flow rates should be used as are recommended for on line measurements. PMID- 11254820 TI - Prenatal risk factors of wheezing at the age of four years in Tanzania. AB - BACKGROUND: A study was undertaken to assess the interactions between prenatal exposures, early life infections, atopic predisposition, and allergen exposures in the development of wheezing up to the age of 4 years in a tropical region of Africa. METHODS: The study subjects comprised children born at the district hospital in Ifakara, Tanzania during a 1 year period who were participating in a trial of iron supplementation and malaria chemoprophylaxis during the first year of life and followed for up to 4 years. From this group of subjects, 658 (79%) participated in the interview at 18 months and 528 (64%) in a second interview at 4 years. Wheezing was measured with the ISAAC questionnaire. A hospital based inpatient and outpatient surveillance system was set up to document all attendance by study children for any cause, including episodes of clinical malaria and lower respiratory tract infections. Total IgE levels and malaria parasites were measured in maternal and cord blood. Total IgE was also measured at 18 months of age. Indoor environmental levels of Der p I and Fel d I were determined using an enzyme linked immunosorbent assay at the same time as the interview at the age of 18 months. RESULTS: The prevalence of wheezing at 4 years is common in Ifakara (14%, range 13-15%). The presence of malaria parasites in cord blood (odds ratio, OR = 6.84, 95% CI 1.84 to 24.0) and maternal asthma (OR = 8.47, 95% CI 2.72 to 26.2) were positively associated with wheezing at the age of 4 years, and cord blood total IgE was negatively associated (OR = 0.24, 95% CI 0.07 to 0.85) (all p<0.05). Parasitaemia at birth was not related to total IgE levels in cord blood (p=0.6). Clinical episodes of malaria during infancy were not associated with wheezing, and nor were levels of indoor aeroallergens. CONCLUSION: These findings suggest that events occurring during pregnancy may play a role in the future appearance of wheezing, although the results must be interpreted with caution because of the small numbers studied. PMID- 11254821 TI - Study of community acquired pneumonia aetiology (SCAPA) in adults admitted to hospital: implications for management guidelines. AB - BACKGROUND: Since the last British study of the microbial aetiology of community acquired pneumonia (CAP) about 20 years ago, new organisms have been identified (for example, Chlamydia pneumoniae), new antibiotics introduced, and fresh advances made in microbiological techniques. Pathogens implicated in CAP in adults admitted to hospital in the UK using modern and traditional microbiological investigations are described. METHODS: Adults aged 16 years and over admitted to a teaching hospital with CAP over a 12 month period from 4 October 1998 were prospectively studied. Samples of blood, sputum, and urine were collected for microbiological testing by standard culture techniques and new serological and urine antigen detection methods. RESULTS: Of 309 patients admitted with CAP, 267 fulfilled the study criteria; 135 (50.6%) were men and the mean (SD) age was 65.4 (19.6) years. Aetiological agents were identified from 199 (75%) patients (one pathogen in 124 (46%), two in 53 (20%), and three or more in 22 (8%)): Streptococcus pneumoniae 129 (48%), influenza A virus 50 (19%), Chlamydia pneumoniae 35 (13%), Haemophilus influenzae 20 (7%), Mycoplasma pneumoniae 9 (3%), Legionella pneumophilia 9 (3%), other Chlamydia spp 7 (2%), Moraxella catarrhalis 5 (2%), Coxiella burnetii 2 (0.7%), others 8 (3%). Atypical pathogens were less common in patients aged 75 years and over than in younger patients (16% v 27%; OR 0.5, 95% CI 0.3 to 0.9). The 30 day mortality was 14.9%. Mortality risk could be stratified by the presence of four "core" adverse features. Three of 60 patients (5%) infected with an atypical pathogen died. CONCLUSION: S pneumoniae remains the most important pathogen to cover by initial antibiotic therapy in adults of all ages admitted to hospital with CAP. Atypical pathogens are more common in younger patients. They should also be covered in all patients with severe pneumonia and younger patients with non-severe infection. PMID- 11254822 TI - Atypical adenomatous hyperplasia of the lung: a clinicopathological study of 118 cases including cases with multiple atypical adenomatous hyperplasia. AB - BACKGROUND: Atypical adenomatous hyperplasia (AAH) of the lung is a putative precursor lesion of adenocarcinoma, according to many immunohistochemical and genetical studies, but few clinicopathological studies on a large number of cases have been reported. The aim of this study was to clarify the clinicopathological characteristics of lung cancer patients with AAH lesions. METHODS: A retrospective study was carried out on 508 consecutive primary lung cancer patients operated on at National Cancer Center Hospital East. The relationship between the number and location of AAH lesions and the clinicopathological features of the lung cancer patients was analysed statistically. RESULTS: A total of 311 AAH lesions were found in 118 (23.2%) of the 508 cases. AAH lesions were detected in 121 of 572 lobes examined, usually in both upper lobes, and occurred most frequently in patients with adenocarcinoma (OR 2.97; 95% CI 1.82 to 4.85). AAH lesions were more frequently detected in patients with multiple primary carcinomas than in those with a single carcinoma (OR 3.06; 95% CI 1.56 to 6.00). The presence of AAH lesions was not significantly correlated with sex, age, smoking status, familial history of malignancy, or preceding malignancy. Patients with multiple AAH lesions were found to have a significantly higher frequency of preceding malignancies. CONCLUSIONS: The present study highlights the clinicopathological characteristics of AAH lesions, showing them to be significantly associated with both adenocarcinoma and multiple primary carcinoma of the lung and suggesting common factors in the histogenesis of multiple AAH lesions and preceding malignancy. PMID- 11254823 TI - Sputum induction as a research tool for sampling the airways of subjects with cystic fibrosis. AB - BACKGROUND: Sputum induction (SI) has proved to be a reliable non-invasive tool for sampling inflammatory airway contents in asthma, with distinct advantages over collection of expectorated sputum (ES) and bronchoalveolar lavage (BAL). A study was undertaken to evaluate the safety of SI and to assess if it might be an equally valuable outcome tool in patients with cystic fibrosis (CF). METHODS: The safety of the procedure was examined and sample volume, cell counts, cytokine concentrations, and bacterial culture results obtained by SI, spontaneous ES, and fibreoptic bronchoscopy were compared in 10 adults with CF. RESULTS: SI was well tolerated and was preferred to BAL by all subjects. The mean (SE) sample volume obtained by SI was significantly greater than ES (6.74 (1.46) ml v 1.85 (0.33) ml, p = 0.005). There was no significant difference in the number of cells per ml of sample collected. There was a difference in the mean (SD) percentage of non epithelial, non-squamous cells collected (67 (28)%, 86 (21)%, and 99 (1)% for ES, SI, and BAL, respectively). These percentage counts were different between ES and both SI and BAL (p=0.03 and p=0.006, respectively). Cell differential counts (excluding squamous cells) from all collection methods were similar (mean (SD) 84 (9)%, 87 (7)%, and 88 (11)% polymorphonuclear cells for ES, SI, and BAL, respectively). The concentrations of interleukin (IL)-8 and tumour necrosis factor (TNF)-alpha were the same in all three samples when corrected for dilution using urea concentration. The test specific detection rate for recovery of bacteriological pathogens was 79% for SI, 76% for ES, and 73% for BAL. CONCLUSION: SI offers safety advantages over BAL and may be a more representative airway outcome measurement in patients with CF. PMID- 11254824 TI - Increasing prevalence of asthma diagnosis and symptoms in children is confined to mild symptoms. AB - BACKGROUND: The prevalence of childhood asthma is increasing but few studies have investigated trends in asthma severity. We investigated trends in asthma diagnosis and symptom morbidity between an eight year time period in a paired prevalence study. METHODS: All children in one single school year aged 8-9 years in the city of Sheffield were given a parent respondent questionnaire in 1991 and 1999 based on questions from the International Survey of Asthma and Allergy in Children (ISAAC). Data were obtained regarding the prevalence of asthma and wheeze and current (12 month) prevalences of wheeze attacks, speech limiting wheeze, nocturnal cough and wheeze, and exertional symptoms. RESULTS: The response rates in 1991 and 1999 were 4580/5321 (85.3%) and 5011/6021 (83.2%), respectively. There were significant increases between the two surveys in the prevalence of asthma ever (19.9% v 29.7%, mean difference 11.9%, 95% confidence interval (CI) 10.16 to 13.57, p<0.001), current asthma (10.3% v 13.0%, mean difference 2.7%, 95% CI 1.44 to 4.03, p<0.001), wheeze ever (30.3% v 35.8%, mean difference 5.7%, 95% CI 3.76 to 7.56, p<0.001), wheeze in the previous 12 months (17.0% v 19.4%, mean difference 2.5, 95% CI 0.95 to 4.07, p<0.01), and reporting of medication use (16.9% v 20%, mean difference 3.0%, 95% CI 1.46 to 4.62, p<0.001). There were also significant increases in reported hayfever and eczema diagnoses. CONCLUSIONS: Diagnostic labelling of asthma and lifetime prevalence of wheeze has increased. The current 12 month point prevalence of wheeze has increased but this is confined to occasional symptoms. The increased medication rate may be responsible for the static prevalence of severe asthma symptoms. The significant proportion of children receiving medication but reporting no asthma symptoms identified from our 1999 survey suggests that some children are being inappropriately treated or overtreated. PMID- 11254827 TI - Respiratory diseases in pregnancy. Introduction. PMID- 11254825 TI - Effect of intravenous pamidronate on bone mineral density in adults with cystic fibrosis. AB - BACKGROUND: Low bone mineral density (BMD) is prevalent in adults with cystic fibrosis. The aim of this study was to assess the effect of intravenous pamidronate on BMD in these subjects. METHODS: Patients were invited to participate if they had a BMD Z score of -2 or less in the lumbar spine, proximal femur, or distal forearm. Patients were randomised to receive either 30 mg intravenous pamidronate every 3 months + 1 g calcium daily (pamidronate group) or 1 g calcium daily (control group). All pancreatic insufficient patients were prescribed oral vitamin D supplements. RESULTS: After 6 months of treatment the pamidronate group (n=13) showed a significant increase in absolute BMD compared with the control group (n=15) in the lumbar spine (mean difference 5.8% (CI 2.7% to 8.9%)) and total hip (mean difference 3.0% (CI 0.3% to 5.6%)). However, the pamidronate group showed a reduction in BMD compared with the control group in the distal forearm (mean difference -1.7% (CI -3.7% to 0.3%)). The use of pamidronate was associated with a high incidence of bone pain in non corticosteroid treated individuals. CONCLUSION: Intravenous pamidronate increases axial BMD in adults with cystic fibrosis, but the high incidence of bone pain associated with this treatment might limit its use. PMID- 11254829 TI - Diaphragm strength in acute systemic lupus erythematosus in a patient with paradoxical abdominal motion and reduced lung volumes. AB - Diaphragmatic weakness is reported as a common feature of the shrinking lung syndrome of systemic lupus erythematosus (SLE). However, in chronic stable SLE it has been shown that, despite poor performance of voluntary tests of diaphragm strength, twitch pressures obtained by stimulating the phrenic nerves are normal. We present a patient with acute SLE and pulmonary involvement who, despite having paradoxical abdominal motion and low maximal inspiratory pressures during voluntary manoeuvres, had normal diaphragm strength when assessed with magnetic stimulation of the phrenic nerves. Following immunosuppressive therapy symptoms and lung function improved, yet diaphragm contractility remained normal and unchanged. We suggest that this case supports the view that reduced diaphragm muscle contractility per se does not explain the small volume lungs and respiratory symptoms in patients with acute SLE. PMID- 11254828 TI - Asthma in pregnancy. PMID- 11254830 TI - Perioperative neuropsychologic testing. PMID- 11254826 TI - Long term sequelae of bronchopulmonary dysplasia (chronic lung disease of infancy). PMID- 11254831 TI - The use of neurocognitive tests in evaluating the outcome of cardiac surgery: some methodologic considerations. PMID- 11254832 TI - A comparison of neuropsychologic deficits after extracardiac and intracaradiac surgery. AB - OBJECTIVE: To compare the incidence of neuropsychologic deficits 1 week and 6 months after coronary artery bypass graft (CABG) surgery (extracardiac) and valve surgery with or without CABG surgery (intracardiac) using reliable change indices to define the incidence of neuropsychologic deficits. DESIGN: Prospective study. SETTING: Cardiac surgical unit in a university teaching hospital. PARTICIPANTS: Patients scheduled for elective multiple-graft (> or =3 grafts) CABG surgery (n = 59), or elective valve surgery (with or without concomitant CABG surgery) (n = 50) and a matched sample of nonsurgical controls (n = 53). INTERVENTIONS: Neuropsychologic assessments were performed 1 day before surgery, 7 days and 6 months after surgery. MEASUREMENTS AND MAIN RESULTS: The 7-day assessment showed no significant differences between valve surgery patients and CABG surgery patients in the incidence of neuropsychologic deficits. When reassessed 6 months postoperatively, the valve group displayed a significantly higher incidence of deficits on the digit symbol test compared with the CABG group (valve 26.7% v CABG 6.8%). In the CABG group, there was a significant change in the incidence of deficits per patient from 7 days to 6 months (p = 0.03) that was not evident in the valve group. CONCLUSION: There are some differences in the neuropsychologic outcome of extracardiac and intracardiac surgery. Patients undergoing isolated CABG surgery showed a greater reduction in the incidence of persisting deficits at 6 months than patients undergoing valve surgery with or without CABG surgery. This finding warrants further investigation, with particular attention to patients undergoing combined valve and coronary artery procedures. PMID- 11254834 TI - Neuropsychologic testing within 18 hours after cardiac surgery. AB - OBJECTIVE: To undertake neuropsychologic testing within 18 hours of cardiac surgery after fast-track anesthesia. DESIGN: Prospective study. SETTING: University hospital, single center. PARTICIPANTS: Fifty patients undergoing first time elective coronary artery surgery. INTERVENTIONS: A neuropsychologic test battery was administered preoperatively and 18 hours and 5 days after surgery. MAIN RESULTS: Seven patients were withdrawn, and 9 patients did not attempt the postoperative tests (on both occasions) because of medical complications. Thirty patients completed > or =4 tests at both postoperative occasions. Of these, 9 patients (30%) showed a deficit in > or =2 tests at 18 hours postoperatively, and 3 (10%) showed a deficit at 5 days postoperatively. CONCLUSION: In the absence of medical complications and despite the difficulties, early postoperative neuropsychologic testing is possible after fast-track anesthesia. Such testing has the potential to more clearly define the course of cognitive decline after cardiac surgery. PMID- 11254833 TI - Early neurobehavioral disorders after cardiac surgery: a comparative analysis of coronary artery bypass graft surgery and valve replacement. AB - OBJECTIVE: To analyze neurobehavioral disorders in the early postoperative period after valve replacement and coronary artery bypass graft (CABG) surgery. DESIGN: Prospective study. SETTING: University hospital. PARTICIPANTS: Patients undergoing elective cardiac surgery with cardiopulmonary bypass; 42 patients in the valve replacement surgery group and 42 patients in the CABG surgery group, with both groups matched post hoc for age, sex, and preoperative cognitive status. MEASUREMENTS AND MAIN RESULTS: All subjects were investigated preoperatively as well as 2 and 7 days postoperatively with a comprehensive neuropsychologic and neuropsychiatric assessment. The groups did not significantly differ with respect to the incidence of postoperative neuropsychiatric disorders. Valve replacement surgery patients exhibited more severe neuropsychologic deficits and showed a slower recovery than patients who underwent CABG surgery. In both groups, postoperative neuropsychologic alterations were most marked in fluency, arithmetic, and memory performance. CONCLUSION: These results indicate that patients after valve replacement surgery have a higher risk of postoperative neuropsychologic alterations mainly attributable to temporal lobe dysfunction. This finding corresponds to a specific vulnerability of hippocampal structures to transient hypoxia. PMID- 11254835 TI - S-100beta protein levels do not correlate with stroke in patients undergoing carotid endarterectomy under general anesthesia. AB - OBJECTIVE: To establish the S-100beta protein profile during carotid artery surgery to show a possible correlation between postoperative stroke and this biochemical marker. DESIGN: Prospective, nonrandomized study. SETTING: Departments of anesthesiology, biochemistry, and vascular surgery in a single university hospital. PARTICIPANTS: One hundred patients consecutively scheduled for carotid endarterectomy. MEASUREMENTS AND MAIN RESULTS: Postoperative neurologic complications were defined as major, occurrence of a postoperative permanent stroke, or minor, occurrence of a new postoperative transient ischemic attack lasting < 2 hours. Serum samples were obtained before induction, before carotid artery cross-clamping, after declamping, at the end of surgery, during recovery, and on the first postoperative day. Concentrations of S-100beta were analyzed using a commercially available kit (LIA-mat S300 analyzer, Byk-Sangtec Medical, Bromma, Sweden). Ninety-five patients awoke without a neurologic defect. Three patients experienced a permanent stroke, and 2 patients had a transient ischemic attack. S-100 basal values were unrelated to preoperative status, including hypertension, neurologic status, renal function, and degree of the carotid lesion. S-100 concentration increased slightly but significantly at the end of surgery and remained stable until the first postoperative day. S-100 profile during the procedure was independent of the duration of carotid artery cross-clamping and the need for a shunt. S-100 serum level was not significantly different in the patients with a postoperative ischemic event in comparison with the entire group. The S-100 profile was not increased in 2 of 3 patients with a permanent stroke and in 1 of 2 patients with a transient ischemic attack in comparison with the 95 patients with uneventful recovery. CONCLUSION: S-100 concentration slightly increased at the end of surgery and remained high until the first postoperative day in all patients. S-100 was not significantly different in the patients with postoperative stroke. S-100 did not serve as a marker for postoperative stroke after carotid artery surgery. This fact must be taken into account during further investigations of S-100. PMID- 11254836 TI - Blood S-100 protein concentration in children undergoing cardiac surgery. AB - OBJECTIVES: To investigate plasma levels of the betabeta isomer of S-100 protein and to assess the relationship between post-cardiopulmonary bypass (CPB) levels of this marker and a variety of perioperative and patient factors in children undergoing cardiac surgery. DESIGN: Prospective study. SETTING: University hospital. PARTICIPANTS: Twenty-five children. INTERVENTIONS: Blood samples (2 mL) for S-100 determinations were collected after the induction of anesthesia, 30 minutes after aortic cross-clamping, 1 hour after the termination of CPB, and 5 and 24 hours after the operation. Electroencephalogram activity was recorded, and neurologic examination was performed on all children 1 day before and 10 days after the operation. Lowest values of nasopharyngeal temperature, mean arterial pressure, arterial carbon dioxide tension (PaCO2), pH, and hematocrit during CPB were recorded. MEASUREMENTS AND MAIN RESULTS: The overall change in S-100 during the study period was found to be statistically significant (p < 0.0001). Correlation between deltaS-100 and age (r = -0.45; p = 0.04), body surface area (r = -0.63; p = 0.002), nasopharyngeal temperature (r = -0.55; p = 0.01), and PaCO2 (r = -0.55; p = 0.009) was statistically significant in infants and children. Multivariate regression analysis indicated significant effects of PaCO2 and body surface area on deltaS-100 levels and area under the curve values. CONCLUSION: In contrast to newborns, infants and older children showed prominent increases in S-100 protein concentration. Lack of pathologic electroencephalogram findings and neurologic signs in the postoperative period precludes the clinical use of S-100 protein concentration as a sensitive marker of cerebral injury. PMID- 11254837 TI - Comparison of two sites of inflow pressure measurement during retrograde cerebral perfusion. AB - OBJECTIVE: To determine whether internal jugular venous valves influence inflow pressure during retrograde cerebral perfusion. DESIGN: Prospective study. SETTING: Community hospital, university setting, single institution. PARTICIPANTS: Ten patients undergoing reconstructive aortic arch surgery with profound hypothermic circulatory arrest. INTERVENTIONS: During retrograde cerebral perfusion, inflow pressure was continuously measured at 2 separate sites relative to the left internal jugular venous valve (ie, superior vena cava inflow catheter [infravalvular pressure] and rostral left internal jugular vein [supravalvular pressure]). MEASUREMENTS AND MAIN RESULTS: Infravalvular pressure of 29.8 +/- 3.5 mmHg and supravalvular pressure of 22.7 +/- 0.8 mmHg were significantly different (mean difference, 7.1 +/- 3.6 mmHg; p = 0.041). In 8 patients, the pressure difference was <6 mmHg; whereas in 2 patients, the pressure difference was >20 mmHg. Bland and Altman analysis revealed 95% limits of agreement on mean bias of -12.9 to 27.8 mmHg. CONCLUSION: Internal jugular venous valves can obstruct retrograde cerebral perfusion inflow, manifest by an inflow pressure difference between the superior vena cava and internal jugular vein. In the presence of competent internal jugular venous valves, measurement of inflow pressure in the superior vena cava may be an inaccurate estimate of actual cerebral perfusion pressure. Internal jugular vein pressure should be monitored to avoid inadvertent cerebral hypoperfusion. PMID- 11254838 TI - Correlation of peripheral venous pressure and central venous pressure in surgical patients. AB - OBJECTIVE: To determine the degree of agreement between central venous pressure (CVP) and peripheral venous pressure (PVP) in surgical patients. DESIGN: Prospective study. SETTING: University hospital. PARTICIPANTS: Patients without cardiac dysfunction undergoing major elective noncardiac surgery (n = 150). MEASUREMENTS AND MAIN RESULTS: Simultaneous CVP and PVP measurements were obtained at random points in mechanically ventilated patients during surgery (n = 100) and in spontaneously ventilating patients in the postanesthesia care unit (n = 50). In a subset of 10 intraoperative patients, measurements were made before and after a 2-L fluid challenge. During surgery, PVP correlated highly to CVP (r = 0.86), and the bias (mean difference between CVP and PVP) was -1.6 +/- 1.7 mmHg (mean +/- SD). In the postanesthesia care unit, PVP also correlated highly to CVP (r = 0.88), and the bias was -2.2 +/- 1.9 (mean +/- SD). When adjusted by the average bias of -2, PVP predicted the observed CVP to within +/-3 mmHg in both populations of patients with 95% probability. In patients receiving a fluid challenge, PVP and CVP increased similarly from 6 +/- 2 to 11 +/- 2 mmHg and 4 +/ 2 to 9 +/- 2 mmHg. CONCLUSION: Under the conditions of this study, PVP showed a consistent and high degree of agreement with CVP in the perioperative period in patients without significant cardiac dysfunction. PVP -2 was useful in predicting CVP over common clinical ranges of CVP. PVP is a rapid noninvasive tool to estimate volume status in surgical patients. PMID- 11254839 TI - Electric impedance for evaluation of body fluid balance in cardiac surgical patients. AB - OBJECTIVE: To evaluate whether electric impedance can be used to monitor body fluid balance and fluid distribution in cardiac surgical patients. DESIGN: Prospective clinical study. SETTING: Heart Center, Rigshospital, Copenhagen. PARTICIPANTS: Sixteen consecutive patients scheduled for cardiac surgery. MEASUREMENTS AND MAIN RESULTS: Body weight, fluid balance, central hemodynamics, and total and segmental body impedance were examined perioperatively. During semisupine rest before surgery, changes in impedance indicated relocation of fluid from the legs to the thorax, mostly in the extracellular space. After surgery, weight and fluid balance increased by 3.87 +/- 0.35 kg and 1.86 +/- 0.16 L (mean +/- SE, p < 0.01) and remained elevated through the next 2 days. Impedance decreased by 30% over the thorax, by 24% over the abdomen, by 2% over the leg, and by 4% over the entire body. Changes in total and thoracoabdominal impedances had the highest correlation to the fluid balance (r = -0.86 and r = 0.87). After correction of impedance values by the constant from the regression model, the mean difference in estimation of fluid changes obtained by electric impedance and by fluid balance was 0 +/- 0.1 L at the range of changes of 4.6 L. CONCLUSION: Alterations in electric impedance closely follow changes in fluid balance during the perioperative period. This method can be used in clinical practice to control postoperative body fluid balance in cardiac surgical patients. PMID- 11254840 TI - Platelet glass bead retention predicts bleeding after cardiac surgery. AB - OBJECTIVE: To determine if the platelet glass bead retention assay can predict bleeding after cardiac surgery. DESIGN: Prospective, observational study. SETTING: Large, tertiary care, academic medical center. PARTICIPANTS: Forty-three adult patients scheduled to undergo elective cardiac surgery employing cardiopulmonary bypass (CPB). MEASUREMENTS AND MAIN RESULTS: Whole blood samples were observed for platelet count, prothrombin time, and platelet (glass bead) retention assay. The platelet retention and prothrombin times were independent univariant and multivariant predictors of bleeding after CPB (r = 0.554, p = 0.0002 and r = 0.655, p = 0.00001). CONCLUSION: The platelet glass bead retention assay measures dynamic platelet function and is sensitive to the CPB-induced adhesion and aggregation defect and correlates with postoperative blood loss. Modification of this platelet function assay used with the prothrombin time may provide a simple and effective diagnostic approach to bleeding after CPB. PMID- 11254841 TI - The effects of recent aspirin ingestion on platelet function in cardiac surgical patients. AB - OBJECTIVE: To examine the effects of the preoperative aspirin-free interval on platelet function in cardiac surgical patients. DESIGN: Prospective clinical investigation. SETTING: University-affiliated teaching hospital. PARTICIPANTS: Patients undergoing elective coronary artery bypass graft surgery (n = 100). INTERVENTIONS: The patients were divided into 3 groups based on the number of days since they last ingested aspirin: < or =2 days, 3 to 7 days, and >7 days. Preoperative platelet function was assessed in all patients using platelet aggregation responses to arachidonic acid, 5 microg/mL, and Platelet Function Analyser (PFA100) collagen/epinephrine closure times. MEASUREMENTS AND MAIN RESULTS: Patients who ceased aspirin < or =2 days preoperatively had weaker platelet aggregation responses (18.5% +/- 7% maximum aggregation, mean +/- SD, n = 36) than patients who ceased aspirin 3 to 7 days preoperatively (68.8% +/- 29%, n = 48, p < 0.001) or >7 days preoperatively (68.3% +/- 28%, n = 16, p < 0.001). Similarly, patients who ceased aspirin < or =2 days preoperatively had longer PFA100 closure times (168 +/- 52 sec) than patients who ceased aspirin 3 to 7 days preoperatively (122 +/- 43 sec, p < 0.001) or >7 days preoperatively (128 +/ 42 sec, p < 0.01). The percentage of abnormal responses was also greatest in the aspirin < or =2 days group. CONCLUSION: Cardiac surgical patients who ingest aspirin < or =2 days preoperatively have greater impairment of platelet function than patients who have a longer preoperative aspirin-free interval. PMID- 11254842 TI - Another point of view on the mechanism of thrombin generation during cardiopulmonary bypass: role of tissue factor pathway inhibitor. AB - OBJECTIVE: To determine the role of tissue factor and tissue factor pathway inhibitor (TFPI) in coagulation activation during cardiopulmonary bypass (CPB). DESIGN: Prospective, observational study. SETTING: Operating room in a city hospital. PARTICIPANTS: Thirty-one patients undergoing cardiac surgery. MEASUREMENTS AND MAIN RESULTS: The plasma levels of tissue factor antigen (tissue factor), total and free TFPI, several markers of thrombin generation (prothrombin fragment F1+2, thrombin antithrombin complex, and fibrinopeptide A), and heparin concentration were measured. Blood samples were obtained after induction of anesthesia (baseline level), before and after CPB, and at the end of the surgery. Despite an average heparin concentration of 2.9 +/- 0.2 IU/ mL, markers of thrombin generation, fibrin formation and its degradation (D-dimer) were observed during CPB. Significant increases of total and free TFPI levels (p < 0.0001) were found during CPB associated with lower tissue factor concentration (p < 0.0001) compared with the baseline values. Heparin concentration correlated with levels of total TFPI (r2 = 0.613, p < 0.0001) and free TFPI (r2 = 0.689, p < 0.0001). Tissue factor concentration showed significant negative correlations with levels of total TFPI (r2 = 0.128, p = 0.0003) and free TFPI (r2 = 0.070, p = 0.0078). CONCLUSION: These data indicate that TFPI release by heparin probably has an important role in the suppression of the tissue factor-dependent coagulation pathway during CPB. These changes occur along with ongoing thrombin generation and its activation. Either insufficient prevention of thrombin generation by TFPI or indirect activation of the intrinsic coagulation pathway occurs during CPB. PMID- 11254845 TI - Hemodynamic instability after parachute-jumping trauma: role of transesophageal echocardiography. PMID- 11254843 TI - Intraoperative transesophageal ultrasonography can measure renal blood flow. AB - OBJECTIVE: To determine the feasibility of acquiring Doppler-derived indices of renal blood flow by transesophageal ultrasonography in the perioperative period. DESIGN: Prospective, sequential, institutional review board-approved study. SETTING: University teaching hospital. PARTICIPANTS: Nine patients with normal renal function, scheduled for elective primary coronary artery bypass graft surgery. INTERVENTIONS: Two-dimensional images of renal parenchyma and Doppler measurement of intrarenal arterial blood flow during internal mammary dissection were acquired. To effect renal blood flow changes, the renal vasodilator dopamine, 2 microg/kg/min, was infused for 20 minutes after baseline measurements were made. Renal Doppler measurements were repeated to determine whether transesophageal ultrasonography can follow these changes. MEASUREMENTS AND MAIN RESULTS: Hemodynamic measurements (heart rate, mean arterial blood pressure, cardiac output, and cardiac index) and Doppler velocity measurements of intrarenal arterial blood flow (peak systolic, end-diastolic, and mean velocity) were made at time 1 (T1 = baseline) and at time 2 (T2 = after 20 minutes of dopamine infusion). The derived Doppler indices, pulsatility index and resistive index, were calculated according to standard formulae. Measurements were compared by paired Student's t-test (two-tailed, p < 0.05, significant). There were no statistical differences between cardiac index (2.10 +/- 0.93 L/min/m2 v 2.21 +/- 0.92 L/min/m2, p = 0.254) and mean arterial pressure (82.3 +/- 11.2 mmHg v 83.3 +/- 14.5 mmHg, p = 0.872) between T1 and T2. Systolic renal velocity increased from 44.7 +/- 13.0 cm/s to 63.0 +/- 20.4 cm/s (p = 0.005), diastolic velocity increased from 12.7 +/- 4.0 cm/s to 22.4 +/- 7.8 cm/s (p = 0.0003), and mean velocity increased from 22.5 +/- 6.6 cm/s to 34.1 +/- 11.7 cm/s (p = 0.003) after infusion of dopamine. These results indicate an increase in renal blood flow from baseline values. The pulsatility index decreased from 1.44 +/- 0.29 to 1.21 +/- 0.24 (p = 0.0005), whereas the resistive index decreased from 0.71 +/- 0.06 to 0.64 +/- 0.06 (p = 0.0004) after dopamine. Reductions in pulsatility and resistive indices indicate a reduction in renal vascular resistance. CONCLUSION: This study demonstrates the ability to acquire two-dimensional images of kidney and renal arterial Doppler velocities using transesophageal ultrasonography during cardiac surgery. Transesophageal renal arterial Doppler waveform analysis can follow changes in renal blood flow patterns secondary to interventional therapy. PMID- 11254844 TI - The effects of fenoldopam on coronary conduit blood flow after coronary artery bypass graft surgery. AB - OBJECTIVE: To quantify the effects of fenoldopam, 0.1 microg/kg/min, on left internal mammary artery (LIMA) and saphenous vein blood flow after coronary anastomosis. DESIGN: Prospective, randomized, double-blind, placebo-controlled trial. SETTING: University teaching hospital, single institution. PARTICIPANTS: Thirty-one American Society of Anesthesiologists III patients undergoing elective coronary revascularization. INTERVENTIONS: A perivascular ultrasonic flow probe (Linton Instrumentation, Norfolk, UK) was placed around the LIMA and saphenous vein graft after coronary anastomosis. MEASUREMENTS AND MAIN RESULTS: Immediately before and at 5-minute intervals for 15 minutes after starting the infusion, blood flow was measured in the LIMA and one saphenous vein graft using a transit time ultrasonic flow probe. Heart rate, blood pressure, and central venous pressure were documented at these time points. Administration of fenoldopam, 0.1 microg/kg/min, did not alter heart rate or blood pressure. A small, nonsignificant increase in LIMA blood flow occurred during the 15-minute study period (30 +/- 12 to 35 +/- 10 mL/min) in patients who received fenoldopam. No significant changes occurred in the placebo group. CONCLUSIONS: The findings indicate that fenoldopam, 0.1 microg/kg/min, did not influence coronary conduit blood flow to a clinically significant extent. The small increase in LIMA blood flow may be of greater importance in high-risk patients or in the prevention of coronary arterial spasm. PMID- 11254846 TI - An unusual cause of systolic anterior motion of the mitral valve. PMID- 11254847 TI - Shone's anomaly complicated by ascending aortic aneurysm in a pregnant woman. PMID- 11254848 TI - Endovascular thoracic aortic aneurysm repair using a single catheter for spinal anesthesia and cerebrospinal fluid drainage. PMID- 11254849 TI - Management of perioperative ventricular dysfunction. AB - With the recognition of the clinical importance of the right ventricle; the development of new techniques for the perioperative evaluation of RV function, particularly transesophageal echocardiography; and new treatment modalities (pharmacologic and mechanical), clinicians will be able to more accurately diagnose and precisely manage patients who have sustained RV injury. PMID- 11254850 TI - Case 1--2001: intraoperative embolization of a right atrial thrombus. PMID- 11254851 TI - Case 2--2001: detection of subepicardial hematoma after percutaneous transluminal coronary angioplasty. PMID- 11254852 TI - Case 3--2001: multiplane transesophageal echocardiography in minimally invasive surgery for coronary artery fistula. PMID- 11254853 TI - Case 4--2001: perioperative transesophageal echocardiography as a diagnostic and monitoring tool in pediatric oncologic surgery. PMID- 11254854 TI - Pro: aprotinin should be used in patients undergoing hypothermic circulatory arrest. PMID- 11254855 TI - Con: aprotinin should not be used in patients undergoing hypothermic circulatory arrest. PMID- 11254856 TI - An interesting transesophageal echocardiographic finding during a beating heart procedure. PMID- 11254857 TI - Unexpected, transesophageal echocardiography-detected left ventricular microbubbles during off-pump coronary artery bypass graft surgery. PMID- 11254858 TI - Left atrial thrombi associated with left ventricular vent catheter. PMID- 11254859 TI - Hematoma compression over the right internal jugular vein resulting in a pulseless ipsilateral arm. PMID- 11254860 TI - Carinal hook wrapped in curvature maneuver: an easy insertion technique for Carlens endobronchial catheter intubation. PMID- 11254861 TI - Reversal of hypoxemia using insufflation of oxygen during one-lung ventilation with a wire-guided endobronchial blocker. PMID- 11254862 TI - Introduction: The evolution of early systemic therapy. PMID- 11254863 TI - Adjuvant and neoadjuvant therapy in prostate cancer. AB - While surgery and radiation therapy remain the only definitive treatments for prostate cancer, single modality therapy has been associated with high failure rates in patients with aggressive disease. Although hormonal therapy has been effective in cases of metastatic disease, the timing of treatment with respect to definitive therapy remains controversial. This review will explore the efficacy of hormonal and chemotherapy in both the adjuvant and neoadjuvant settings. A MEDLINE search was performed to identify pertinent articles regarding both adjuvant and neoadjuvant therapy in prostate cancer. Articles of historical relevance in addition to those using large patient numbers with a randomized design were reviewed preferentially. Since hormonal therapy has been considered standard treatment at the time of cancer progression after definitive therapy, many of the randomized trials essentially compared adjuvant therapy to delayed therapy. Historical trials using adjuvant hormonal therapy have been limited due to difficulties in clinical staging, as well as toxicities attributed to the formulations used. More recently, hormonal therapy has been found to delay disease progression, increase disease-free survival, and decrease mortality when given immediately after prostatectomy or radiation therapy in selected patients. Neoadjuvant hormonal therapy can improve disease-free survival and local control when given before radiation therapy; it has only decreased positive surgical margins when given prior to radical prostatectomy. Although hormonal therapy given immediately after either radical prostatectomy or radiation therapy is highly effective, the side effects of persistent long-term use must be weighed for each patient. While the use of chemotherapy has been limited by the lack of active agents, newer combinations have shown effectiveness in patients with hormone refractory disease, raising the possibility of their use in the adjuvant setting. PMID- 11254864 TI - Adjuvant and neoadjuvant treatment of breast cancer. AB - Treatment of early breast cancer has been revolutionized during the past 30 years and new data continue to refine our knowledge of systemic treatments for this stage of disease. The updated worldwide overview has confirmed that, in terms of recurrence and survival, the balance of the known long-term benefits and risk favors some months of adjuvant polychemotherapy and/or a few years of tamoxifen for a wide range of patients. Both the overview and individual trials have shown that anthracycline-containing regimens can achieve additional reduction of the risk of disease relapse and death over cyclophosphamide, methotrexate, and fluorouracil (CMF)-like regimens. Paclitaxel-containing regimens appear promising, but require additional confirmation with longer follow-up. By contrast, controversy still exists on the role of high-dose chemotherapy in high risk patients. Primary (neoadjuvant) chemotherapy is a new modality to treat large operable breast cancers and offers the possibility of breast conservation with treatment results at least similar to those achieved with classical adjuvant regimens. In the near future, newer agents and information gained on the role of prognostic and predictive factors will probably increase the effectiveness of adjuvant and neoadjuvant treatments. PMID- 11254865 TI - Adjuvant therapy of colon cancer. AB - The primary curative therapy of colorectal cancer is surgical resection. However, within the last 15 years, prospectively randomized appropriately powered clinical trials have convincingly demonstrated that adjunctive postoperative adjuvant chemotherapy is of benefit to all patients with node-positive disease (stage III) and arguably to high-risk node-negative (stage II) cases. In the United States, the clinical trials encompassing greater than 5,000 cases have demonstrated that fluorouracil (5-FU)/leucovorin used in a variety of doses and schedules improves disease-free and overall survival in resected node-positive (stage III) colorectal cancer. The postoperative use of 5-FU/leucovorin for approximately 6 months represents standard of care for such patients. Current clinical trials are evaluating the role of nonfluorinated pyrimidine chemotherapeutic agents in adjuvant chemotherapy for resected large bowel cancer. 5-FU/leucovorin combined with irinotecan (CPT-11) versus 5-FU/leucovorin are being tested in a national intergroup clinical trial. Another trial is evaluating 5-FU/leucovorin plus oxaliplatin versus 5-FU/leucovorin alone. These clinical trials will be important in defining the appropriate standard of care for patients with resected colorectal cancer, since recent studies in advanced colorectal cancer in the United States and in Western Europe have demonstrated that combinations of 5 FU/leucovorin and CPT-11 or 5-FU/ leucovorin and oxaliplatin are superior to 5 FU/leucovorin alone. PMID- 11254866 TI - The use of sentinel lymphadenectomy to identify candidates for postoperative adjuvant therapy of melanoma and breast cancer. AB - Sentinel lymphadenectomy (SLND) is fast becoming the procedure of choice for staging primary breast carcinoma and melanoma. This simpler and less morbid alternative to standard lymph node dissection can increase the rate of detecting nodal disease. Because the tumor status of the regional lymph nodes remains a significant prognostic tool in both diseases, clinicians may use SLND to facilitate selection of patients for adjuvant chemotherapy. Although SLND has been validated by institutions worldwide, it continues to evolve. The following review will examine current data and controversies surrounding this emerging technology. PMID- 11254867 TI - New prognostic factors for breast cancer recurrence. AB - Decisions regarding the use of adjuvant therapy for breast cancer are strongly influenced by the risk of disease recurrence and death. These risks are now determined by examining the currently recognized breast cancer prognostic factors including clinical stage, axillary nodal status, tumor size and grade, hormone receptor status, and presence of lymphovascular involvement. Newer factors are being evaluated in an attempt to more precisely define disease-related prognosis. This article provides an overview of issues that need to be considered when analyzing studies of prognostic factors, as well as a review of the currently recognized and the newer candidate prognostic factors. PMID- 11254868 TI - Adjuvant immunotherapy for melanoma and colorectal cancers. AB - Immune approaches to the therapy of colorectal cancer and melanoma have substantially evolved over the past years, from treating patients with nonspecific immune stimulants to a focus on the use of tumor-associated antigens (TAAs) either by passive immune therapy with antibodies targeted directly to tumor cells or by active immune therapy via vaccination with tumor cells, tumor cell lysates, peptides, carbohydrates, gene constructs encoding proteins, or anti idiotype antibodies that mimic TAAs. We review the different immunotherapeutic approaches to the treatment of melanoma and colorectal cancers, and summarize relevant clinical trials. PMID- 11254869 TI - The potential role of antivascular therapy in the adjuvant and neoadjuvant treatment of cancer. AB - Antivascular therapy may be considered as one of the most promising approaches in the treatment of cancer. Antivascular treatment may be divided in antiangiogenesis and vascular targeting. Antiangiogenic therapy prevents neovascularization by inhibiting proliferation, migration, and/or differentiation of endothelial cells. Vascular targeting is directed at the existing tumor vasculature. Several inhibitors of angiogenesis are currently being tested in clinical cancer trials. The challenge for the next decade is to incorporate antivascular approaches into conventional treatment strategies. This review will summarize the conceptual basis of antivascular therapy and discuss the potential role of these agents in the adjuvant, neoadjuvant, and perioperative treatment of cancer. PMID- 11254870 TI - Secondary chemoprevention of upper aerodigestive tract tumors. AB - Patients with successfully treated upper aerodigestive tract (UADT) tumors commonly develop second primary tumors (SPTs). These tumors occur more often than chance would predict, arise in both the upper or lower aerodigestive tracts, are frequently preceded by leukoplakia, and are a major cause of treatment-related failure. Measures to control SPTs include primary prevention with tobacco and alcohol abstinence, surveillance endoscopy, and secondary chemoprevention. Chemoprevention is the administration of natural or synthetic substances to suppress or reverse the malignant process. Secondary chemoprevention of the UADT is the suppression or reversal of leukoplakia and/or SPTs. Classic antioxidant micronutrients such as retinoids, carotenoids, and certain other agents have been effective in nonrandomized and randomized clinical trials, but treatment is uncertain and recurrences common. These facts, coupled with recent harmful effects of beta-carotene in two clinical trials, stress the need for additional basic science, translational, and clinical research. Chemoprevention is a promising new technology, but is not currently standard therapy for the secondary prevention of UADT tumors. PMID- 11254871 TI - Complementary and alternative medicine in early-stage breast cancer. AB - Complementary and alternative medicine (CAM) are becoming increasingly popular in many medical situations, particularly among patients with cancer. CAM encompasses a range of modalities including dietary and vitamin supplements, mind-body approaches, acupuncture, and herbal medicines. In contrast to standard chemotherapeutic and hormonal regimens used for the adjuvant treatment of early stage breast cancer, controlled clinical trials have generated few data on the relationship between CAM and the outcomes of recurrence or survival, or even overall quality of life and safety. The objectives of CAM treatments are manifold: the reduction of toxicities of therapy, improvement in cancer-related symptoms, enhancement of the immune system, and even a direct anticancer effect. The primary basis of CAM rests on empirical evidence and case studies, as well as theoretic physiologic effects. In some cases, laboratory or clinical data lend support to these modalities. Some types of CAM are based on ancient Oriental forms of medicine founded on centuries of experience documented through oral and written text. Nevertheless, the paucity of evidence in the clinical setting limits firm conclusions about the effectiveness or safety of most CAM approaches in breast cancer. This review will summarize the basis for the application of certain CAM modalities in the therapy of early-stage breast cancer and will highlight some of the directions of investigative work that could lead to a rational integration of CAM into conventional adjuvant therapy. PMID- 11254872 TI - The Bcr N-terminal oligomerization domain contributes to the full oncogenicity of P190 Bcr/Abl in transgenic mice. AB - The Bcr/Abl P190 oncoprotein is responsible for the development of Philadelphia chromosome positive acute lymphoblastic leukemia (ALL). The Bcr moiety in Bcr/Abl activates the Abl tyrosine kinase, an ingredient essential for the transforming capability of Bcr/Abl. Residues 1-63 of Bcr form an N-terminal oligomerization domain and are key to Abl activation in vitro. Mice transgenic for P190 BCR/ABL reproducibly develop an aggressive B-lineage lymphoblastic leukemia/lymphoma. Here we test the hypothesis that residues 1-63 of Bcr have a major in vivo contribution to the oncogenicity of Bcr/Abl P190 by the generation of mice transgenic for an N-terminal deleted form of P190. We find that although the transgene is expressed in the bone marrow of mice at an early age, the incidence of leukemogenesis is greatly diminished as compared to mice transgenic for non mutated P190 Bcr/Abl. Sporadic hematological malignancies which did develop showed decreased levels of phosphotyrosine as compared to those of wild-type P190 transgenics, although Ras was activated. These results demonstrate that the Bcr oligomerization domain contributes to the oncogenicity of Bcr/Abl in vivo. PMID- 11254873 TI - D-mannoheptulose phosphorylation by hexokinase isoenzymes. AB - D-mannoheptulose is a specific inhibitor of D-glucose phosphorylation by hexokinase isoenzymes. In the present study, the phosphorylation of this heptose was investigated by either a spectrophotometric or radioisotopic procedure. Using yeast hexokinase, the phosphorylation of 25 mM D-mannoheptulose only represented 0.02% of that of 5 mM D-glucose. Such a percentage was increased to 3.93% in the case of bovine heart hexokinase. In the latter case, the Km for D-mannoheptulose was close to 0.2 mM and both D-glucose (0.1-1.0 mM) and D-glucose 6-phosphate (also 0.1-1.0 mM) inhibited the phosphorylation of the heptose (0.03-0.60 mM). Human B-cell glucokinase also catalyzed the phosphorylation of D-mannoheptulose (0.1 mM), which was now increased in a bell-shaped manner by D-glucose (1.0-20 mM). Likewise, rat parotid gland, liver and pancreatic islet homogenates catalyzed the phosphorylation of D-[3H]mannoheptulose. The results obtained in these three tissues differed from one another by their absolute values (per mg wet wt.), relative values (by reference to the phosphorylation rate of 10 mM D glucose), and sensitivity to inhibition by D-glucose (10 mM). PMID- 11254874 TI - Coordinate immunoreactivity to vascular endothelial growth factor receptor-2 and its ligand suggests a paracrine regulation during the development of the vascular system in the chick embryo bursa of Fabricius. AB - The bursa of Fabricius is a lymphoid organ of the chick which plays an important role in the development of the immune system. The role of angiogenic factors in the development of the vascular system of this organ has been poorly investigated. Vascular endothelial growth factor (VEGF) is a major regulator of endothelial cell proliferation, angiogenesis and vascular permeability, and its activities are mediated by two receptors, VEGFR-1 and VEGFR-2. In this study we have investigated by immunohistochemistry the VEGF and VEGFR-2 immunoreactivity in developing bursa of Fabricius. Starting from day 10 of incubation, the endodermal epithelium reacts with VEGF and gives rise to the lymphoid follicles, while the vascular endothelium reacts with VEGFR-2. These data support the view that VEGF acts as a paracrine stimulator of angiogenesis in the avian embryo and confirm the requirement of the endodermal layer for the normal formation of blood vessels by mesodermal cells. PMID- 11254875 TI - Sesamolin from sesame seed inhibits proliferation by inducing apoptosis in human lymphoid leukemia Molt 4B cells. AB - The exposure of human lymphoid leukemia Molt 4B cells to sesamolin, a component of sesame seed led to both growth inhibition and the induction of apoptosis. Morphological change showing apoptotic bodies was observed in the cells treated with sesamolin. The fragmentation of DNA by sesamolin to oligonucleosomal-sized fragments that are characteristics of apoptosis was observed to be concentration- and time-dependent. These findings suggest that growth inhibition of Molt 4B cells by sesamolin results from the induction of apoptosis in the cells. PMID- 11254876 TI - Regulated expression of a dominant negative protein kinase C epsilon mutant inhibits the proliferation of U-373MG human astrocytoma cells. AB - The tight regulation of protein kinase C (PKC) activity is crucial for maintaining normal cell proliferation. Excessive PKC activity leads to uncontrolled growth and malignant transformation. It has been reported that the activity of PKC is higher in astroglial cell lines than in normal astrocytes. Previously, we demonstrated that PKC epsilon is overexpressed in astroglial cell lines and in samples from primary high-grade astroglial brain tumors. Because there are no PKC epsilon isozyme-specific inhibitors, we chose a genetic approach to confirm that PKC epsilon is a potential target for inhibiting astroglial cell proliferation. We regulated the expression of a dominant negative PKC epsilon mutant (PKC epsilon 1-401 encoding amino acid 1-401) in U-373MG human astrocytoma cells using a tetracycline-regulated expression vector and established stable clones. Induction of expression of the dominant negative PKC epsilon mutant by the addition of doxycycline, a tetracycline derivative, completely blocked proliferation of U-373MG cells in proliferation and clonogenic assays. Although the induction of the dominant negative PKC epsilon mutant did not markedly affect mitogen-induced tyrosine phosphorylation of the mitogen-activated protein (MAP) kinases, it inhibited the induction of c-Fos protein expression by substance P (SP) and fetal bovine serum (FBS). These results clearly show that the expression of dominant negative PKC epsilon leads to the inhibition of U-373MG cellular proliferation demonstrating that this isozyme may be a potential therapeutic target for astroglial brain tumors. PMID- 11254877 TI - Non-invasive approach for diagnosing atrophic gastritis using the 13C-bicarbonate breath test. AB - The experiments presented here were done to evaluate whether the levels of CO2 in respiratory air during the 13C-bicarbonate breath test (13C-BBT) may be used as a marker of non-invasive diagnosis of the levels of atrophic gastritis. Twenty eight patients with chronic gastritis and five healthy volunteers were enrolled in the study. Moreover, experimental gastritis was induced in rats by N-methy-N nitro-N-nitrosoguanidine. In human, the levels of atrophic gastritis were evaluated from the vascular pattern of the gastric fornix. Total delta 13CO2 calculated from the 13C-BBT and the mucosal thickness ratio (MTR) were measured in rats with experimental gastritis. The levels of 13CO2 were significantly higher from patients with a vascular pattern at the fornix than in those without a vascular pattern (p<0.01). There was a good correlation between MTR and the levels of 13CO2, in rats with experimental gastritis (p<0.01). These findings indicate that the levels of 13CO2 during 13C-BBT reflect the levels of atrophic gastritis and show its clinical significance for non-invasive evaluation of atrophic gastritis. This has important clinical implications in selecting Helicobacter pylori-positive cases for therapy and follow-up. PMID- 11254878 TI - HGF/SF modifies the interaction between its receptor c-Met, and the E cadherin/catenin complex in prostate cancer cells. AB - The effect of HGF/SF on the association between the E-cadherin/catenin complex and the tyrosine kinase receptor c-Met, was examined in prostate cancer cells LNCap FGC. Stimulation by HGF/SF showed E-cadherin and beta-catenin to be co precipitated and located at areas of cell-cell contact with the HGF/SF receptor c Met, as detected by immunoprecipitation and immunofluorescence respectively. Furthermore, continued exposure to this motogen increased the level of co precipitations between the E-cadherin/catenin complex with c-Met, and also increased tyrosine phosphorylation of c-Met. In contrast, continued stimulation by HGF/SF decreased the level of co-localised peripheral staining and increased the level of cytoplasmic staining. In conclusion, the association between the E cadherin/catenin complex with the HGF/SF receptor c-Met, may influence or regulate intercellular adhesion in prostate cancer following stimulation by HGF/SF. PMID- 11254879 TI - Base sequence-specific attack of stilbene estrogen metabolite(s) on the mitochondrial DNA: implications in the induction of instability in the mitochondrial genome in the kidney of Syrian hamsters. AB - We have demonstrated previously that diethylstilbestrol is metabolized to diethylstilbestrol reactive metabolites by mitochondrial enzymes in vitro. In vitro, these reactive intermediates bind to mitochondrial DNA. Here we have investigated the in vivo formation of diethylstilbestrol adducts with mitochondrial DNA, the nature of mitochondrial DNA-diethylstilbestrol adducts, and the influence of diethylstilbestrol adduction on in vitro replication of a mitochondrial gene. Diethylstilbestrol administration to male hamsters produced several adducts in mitochondrial DNA of both kidney and liver. The total relative adduct levels were 5- to 6-fold higher in mitochondrial DNA than in nuclear DNA. The chromatographic mobility of mitochondrial DNA adducts formed in vivo were similar to that of dGMP-DES quinone adducts formed in vitro. The identity of mitochondrial DNA adducts formed in vivo was further confirmed as dGMP diethylstilbestrol quinone adducts by rechromatography and cochromatography. Using a DNA polymerase arrest assay we found that the DES quinone attack on a mitochondrial respiratory gene, i.e., the gene for subunit III of cytochrome c oxidase (COIII), was specific for guanine residues that were adjacent to cytosine residues. Long-term treatment with diethylstilbestrol produced tumors in the kidney, and the level of COIII transcripts was 5- to 10-fold higher in tumor samples than age-matched control kidneys. These findings suggest that i) mitochondrial DNA appears more susceptible to formation of diethylstilbestrol adducts than nuclear DNA, ii) the DNA adducts formed by DES were predominantly with guanines, iii) the adducted bases stopped DNA polymerase-mediated in vitro replication of the COIII gene, and iv) long-term exposure of hamsters to diethylstilbestrol elevated the expression of COIII mRNA. These results suggest that obstruction of replication of the mitochondrial genes by covalent modifications of the mitochondrial DNA by diethylstilbestrol may produce mitochondrial genomic instability in vivo and may provide an explanation for the DES-induced mitochondrial structural abnormality. PMID- 11254880 TI - Expression of multidrug resistance associated transporters (MDR1, MRP1, LRP and BCRP) in porcine oocyte. AB - Transporters such as P-glycoprotein (MDR1), multidrug resistance protein 1 (MRP1), lung resistance-related protein (LRP) and breast cancer resistance protein (BCRP) are associated with multidrug resistance in various carcinoma cell lines. The expression of these molecules has been also characterized in human normal tissues. However, the expression of these molecules in oocyte is still unclear. In order to obtain more insight into the physiological role of these transporters, their expression in porcine oocyte were examined by reverse transcriptase-polymerase chain reaction. MDR1, MRP1 and LRP genes, but not BCRP gene were found to be expressed in porcine oocyte. After the subcloning and sequence analysis of MDR1, MRP1 and LRP genes, the high homology of these transporters were observed between porcine and human gene. These findings suggest that MDR1, MRP1 and LRP play an important physiological role(s) in an oocyte. PMID- 11254881 TI - Prolonged orexin administration stimulates steroid-hormone secretion, acting directly on the rat adrenal gland. AB - Orexins A and B are two hypothalamic peptides, that play a role in the central control of food intake. Orexins act via two subtypes of receptors: OX1R which is selective for orexin A, and OX2R which binds both orexins. Reverse transcription polymerase chain reaction demonstrated the expression of both OX1R and OX2R gene in the adrenal cortex of adult female rats. The prolonged systemic administration of orexins A and B (20 ng/kg x day, for 7 days) affected neither adrenal weight and the morphology of adrenocortical zones (as evaluated by morphometric techniques) nor ACTH plasma concentration in rats. In contrast, the treatment with both orexins increased plasma concentration of both aldosterone and corticosterone. Taken together, these findings indicate that orexins exert a marked direct chronic secretagogue action on adrenocortical cells, acting through both OX1R and OX2R. PMID- 11254882 TI - Assessment of B-cell mass in isolated islets exposed to D-[3H]mannoheptulose. AB - D-mannoheptulose was recently proposed to be transported into cells mainly at the intervention of GLUT2. In the present study, it was investigated whether advantage could be taken from such a situation to assess the contribution of insulin-producing B-cells to the total mass of isolated rat pancreatic islets. After 60 min incubation at 37 degrees C in the presence of 8.3 mM D-glucose, the intracellular distribution space of D-[3H]mannoheptulose (0.1 mM) averaged, in islets from control and streptozotocin-induced diabetic rats, respectively 49.0+/ 2.3 and 6.2+/-1.5% of the corresponding intracellular 3HOH space, all values being corrected for extracellular contamination as judged from the distribution space of [U-14C]sucrose (1.0 mM). These findings indicate that the present approach indeed allows to assess the relative contribution of B-cells to total islet mass for purpose of comparison between animals with different metabolic and/or hormonal status. PMID- 11254883 TI - Decrease in Ca2+-ATPase activity in the brain plasma membrane of rats with increasing age: involvement of brain calcium accumulation. AB - The alteration in Ca(2+)-ATPase activity in the brain plasma membrane of rats with increasing age was investigated. Calcium content in the brain tissues was significantly raised in aged rats (50 weeks old) as compared with that of young rats (5 weeks old). Increasing age caused a significant decrease in Ca(2+)-ATPase activity in the brain plasma membranes. The presence of N-ethylmaleimide (2.5 or 5 mM), a modifying reagent of thiol (SH)-groups, in the reaction mixture caused a significant decrease in the brain plasma membrane Ca(2+)-ATPase activity of young and aged rats, while dithiothreitol (2.5 or 5 mM), a protecting reagent of SH groups, produced a significant increase in the enzyme activity, indicating that the SH-group is an active site of Ca(2+)-ATPase. The active site of Ca(2+)-ATPase may not be impaired by ageing. The brain plasma membrane Ca(2+)-ATPase activity of young rats was significantly reduced in the presence of dibutyryl cyclic AMP (10(-7)-10(-5) M) or inositol 1, 4, 5-trisphosphate (10(-7)-10(-5) M) in the reaction mixture. Such an decrease was not seen in aged rats. The responsibility for signaling factors seemed to be weakened by ageing. Calmodulin (2.5 and 5 microg/ml) or regucalcin (10(-8) and 10(-7) M), a Ca(2+)-regulating protein, did not have an effect on Ca(2+)-ATPase activity. This study demonstrates that ageing induces a decrease in Ca(2+)-ATPase activity in the brain plasma membranes. This finding suggests a cellular mechanism by which ageing causes calcium accumulation in brain. PMID- 11254884 TI - Bax expression as a prognostic marker of postoperative chemoradiotherapy for patients with esophageal cancer. AB - Postoperative chemoradiotherapy was introduced to improve the survival of patients with esophageal squamous cell carcinoma (ESCC). However, considerable number of patients still die of cancer recurrence despite curative operation plus chemoradiotherapy. This indicates that some ESCCs are chemoradio-resistant. To prevent unnecessary treatment and to improve the effect of post-operative adjuvant therapy, it seems to be important to investigate biological markers of chemoradio-sensitivity in ESCC. Loss of Bax expression has been reported to be associated with poor response to chemotherapy in breast cancer, and Bax promotes apoptosis in cells. Abnormal expression of Bax may play an important role in chemoradio-sensitivity in malignant tumors. In this study, we retrospectively investigated the prognostic significance of the expressions of Bax and p53 in patients with ESCC. Immunoreactivities of Bax and p53 were evaluated in 141 surgically resected ESCC by using monoclonal antibodies. Prognoses of 141 patients with or without postoperative chemoradiotherapy were compared among groups with high and low expressions of Bax or p53. High immunoreactivities of Bax and p53 were detected in 49 cases (33.1%) and in 70 cases (47.3%), respectively. Loss of Bax expression was detected more frequently in p53-positive tumors. Bax expression correlated with favorable prognosis (P=0.016) in 57 patients with postoperative chemoradiotherapy. However, in 84 patients without adjuvant therapy, the prognostic significance of Bax was minimal. Moreover, in patients with or without postoperative chemoradiotherapy, p53 expression did not correlate with the prognosis. Bax expression may be a good marker for chemoradio sensitivity in patients with ESCC. PMID- 11254885 TI - Presence of endogenous morphine and morphine 6 glucuronide in human heart tissue. AB - Human atria contain the opiate alkaloids morphine and morphine 6 glucuronide as determined by high performance liquid chromatography coupled to electrochemical detection. This method found endogenous morphine and morphine 6 glucuronide at 106.28+/-61.58 and 48.32+/-24.63 (+/- SD) ng/gm wet weight, respectively. Identification of these opiates was confirmed by nano electrospray ionization double quadrupole orthogonal acceleration time of flight mass spectrometry. Furthermore, human saphenous vein fragments did not contain morphine as determined by these methods. Fragmentation from a selected precursor ion by collision-induced dissociation of endogenous morphine 6 glucuronide (462.14 da) yields morphine (286.14 Da) obtained from the heart tissues and with the authentic material, further demonstrating the presence of endogenous morphine. Thus, vascular tissues appear to contain endogenous opiate alkaloids. PMID- 11254886 TI - Transforming growth factor-beta 1 and insulin-like growth factor-1 expression in ovarian endometriotic cysts: a preliminary study. AB - Increased concentrations of TGF-beta 1 in endometriotic tissue are considered important in the pathophysiology of endometrial diseases since TGF-beta 1 may inhibit natural killer activity and induce angiogenesis and proliferation of endometrial stromal cells. In the present study we report on TGF-beta 1, IGF-1 and their receptor localization, as detected by Northern hybridization and immunohistochemistry, in ovarian endometriotic tissues removed during surgical procedures. We detected comparable expression of IGF-1 and IGF-1 receptor in the stromal and epithelial compartments, thus confirming disregulated expression of the IGF system in ovarian endometriosis. On the contrary, strongly increased TGF beta 1 steady state level mRNA expression was detected in all endometriotic samples. In addition, we demonstrated weak TGF-beta 1 immunohistochemical expression in the epithelial lining and intense expression in the cellular stroma of ovarian endometriomas, thus suggesting that TGF-beta 1 could have an important role in the maintenance and propagation of the disease. On the basis of these preliminary results we can assume that TGF-beta 1, IGF-1 and their receptors may play an important role in the pathogenesis of endometriosis. PMID- 11254887 TI - Induction of apoptosis in murine lymphoma cells by cyclosporin A. AB - The aim of this study was to investigate if CsA could induce apoptosis in the murine T-lymphoma cell line LBC, whose growth is inhibited by this immunosuppressive drug. CsA induced programmed cell death in LBC cells with typical features of apoptosis demonstrated by exposure of phosphatidyl serine residues on the cell membrane, the decrease of cell DNA content, chromatin condensation, and nuclear fragmentation. Apoptosis was evident within 12 h after CsA incubation, with a maximal effect at 48 h, in a time and dose-dependent fashion. In addition, the role of apoptosis inhibitors (Bcl-2 and Bcl-x) and the apoptosis inducer (Bax) in CsA induced-apoptosis was evaluated. The expression of Bcl-2 and Bax proteins were high in LBC cells and following CsA treatment the expression of these proteins as well as Bcl-XL decreased. In this work we demonstrated that cell growth inhibition following CsA treatment in LBC was paralleled by the induction of apoptosis thus providing an interesting animal model to identify the mechanism participating in the regulation of apoptotic genes by CsA in T-cell neoplasms and to assess preclinical in vivo trials of T cell lymphoma-related disorders. PMID- 11254888 TI - The nuclear receptors FXR and LXRalpha: potential targets for the development of drugs affecting lipid metabolism and neoplastic diseases. AB - The orphan nuclear receptors FXR and LXRalpha have become challenging targets for the discovery of new therapeutic agents. Bile acids and hydroxysterol intermediates are the respective natural ligands of these two structurally and functionally closely related receptors. Both FXR and LXRalpha; are thought to play a major role in the control of cholesterol catabolism by regulating the expression of cholesterol 7alpha-hydroxylase, the rate limiting enzyme of bile acid synthesis. Reverse cholesterol transport might also be affected by FXR and LXR since they control the expression of PLTP and CETP, two proteins involved in the transfer of phospholipid, cholesterol and cholesteryl esters among plasma lipoproteins. A new class of potent synthetic activators of FXR, the 1,1 bisphosphonate esters, has been discovered which up regulate the Intestinal Bile Acid Binding Protein gene (I-BABP) as demonstrated for chenodeoxycholic acid, however there are no known synthetic activators yet identified for LXRalpha. The evaluation of FXR as a potential target for the development of drugs affecting plasma cholesterol can take advantage of the fact that the activators of FXR (farnesol, bile acids and the 1,1-bisphosphonate esters) have been studied in various in vitro and in vivo models. Administration of chenodeoxycholic acid to animals and man did not result in the increase in plasma cholesterol expected from a decrease in cholesterol 7alpha-hydroxylase expression. Like farnesol, the 1,1-bisphosphonate esters increase the rate of degradation of HMGCoA reductase and have the unexpected property of inducing hypocholesterolemia in normal animals. The natural and synthetic FXR agonists trigger differentiation, inhibit cell proliferation and are potent inducers of apoptosis. The 1,1-bisphosphonate ester SR-45023A (Apomine) is presently being developed as an antineoplastic drug. PMID- 11254889 TI - Endurance exercise training and reproductive endocrine dysfunction in men: alterations in the hypothalamic-pituitary-testicular axis. AB - Research indicates that endurance exercise training has significant effects upon the reproductive endocrine system of humans. Until recently, this effect was thought to be limited primarily to women. However, a growing body of evidence demonstrates that the male reproductive endocrine system is also effected. Specifically, the circulating hormonal levels of testosterone are found to be at low concentrations; and, the hypothalamic-pituitary-testicular axis that regulates testosterone production is altered in endurance trained men. The physiological mechanism inducing the lower testosterone is currently unclear; but in many respects, these men display hypogonadotropic hypogonadism characteristics. Currently, the time course of the changes in the reproductive endocrine system is unresolved and in need of much furthers scientific investigation. The evidence available, however, suggests that a slowly developing process requiring years of exercise training results in these changes. Potentially, the lowered testosterone levels of the endurance-trained male could disrupt some of their anabolic or androgenic dependent processes. To date, there are only a limited number of findings suggesting that a consistent disruption of testosterone dependent processes occur due to endurance exercise training (e.g., oligo-spermatogenesis). Conversely, the alterations in testosterone concentration brought about by endurance training could have cardiovascular protective effects and thus be beneficial to the health of these men. PMID- 11254890 TI - Pharmacological interference with transcriptional control of osteoblasts: a possible role for leptin and fatty acids in maintaining bone strength and body lean mass. AB - Osteoblasts pass through a sequence of events controlled by hormones and transcriptional factors ensuring proper development of phenotype and functional properties until the osteoblast enter the osteocyte phenotype and/or undergo apoptosis. During its life cycle, the osteoblasts proliferate, deposit matrix proteins and mineralize it until they turn into osteocytes believed to constitute a mechanosensor mesh giving feed-back to the osteoblast to initiate bone modeling or remodeling necessary for the making or remaking of proper bone architecture and strength. It appears that several factors common to osteoblast and adipocyte differentiation determine their entry into different functional stages. Such factors are insulin, growth hormone (GH), insulin-like growth factor type I (IGF I), transforming growth factor beta (TGFbeta), platelet derived growth factor (PDGF), fibroblast growth factor (FGF), cytokines (e.g. interleukins, interferon and tumor necrosis factor alpha (TNF alpha), bone morphogenetic proteins (BMPs), glucocorticoids, retinoic acid (RA), prostaglandins and cAMP-elevating hormones. The focus of this article is to review the effects of leptin on bone cells and bone turnover, the peroxisome proliferator-activated receptors (PPARs) in the regulation of bone and fat cell differentiation, hormones and fatty acids on the orchestration of osteoblast and adipocyte derived regulatory signals, and mechanostimulation of bone on the mechanisms by which the above mentioned factors modulate osteoblast and adipocyte function. The hypothesis or concept is that prescription of a certain treatment regimen to correct bone turnover, without attempting to assess how hormonal homeostasis, nutritional factors and physical exercise may interact locally, will remain far from optimal, and may even prove detrimental to the patient's health condition. PMID- 11254891 TI - Proteomics as a tool in the pharmaceutical drug design process. AB - Proteomics is a technology platform that is gaining widespread use in drug discovery and drug development programs. Defined as the protein complement of the genome, the proteome is a varied and dynamic repertoire of molecules that in many ways dictates the functional form that is taken by the genome. The importance of proteomics is a direct consequence of the central role that proteins play in establishing the biological phenotype of organisms in healthy and diseased states. Moreover, proteins constitute the vast majority of drug targets against which pharmaceutical drug design processes are initiated. By studying interrelationships between proteins that occur in health and disease and following drug treatment, proteomics contributes important insight that can be used to determine the pathophysiological basis for disease and to study the mechanistic basis for drug action and toxicity. Proteomics is also an effective means to identify biomarkers that have the potential to improve decision making surrounding drug efficacy and safety issues based on data derived from the study of key tissues and the discovery and appropriate utilization of biomarkers. PMID- 11254892 TI - The chemical and biological aspects of fluoroquinolones: reality and dreams. AB - A vast array of fluoroquinolones having excellent broad-spectrum activity form an invaluable part of the present anti-infective armory of the clinicians. A number of these compounds are today's blockbusters of the antibacterial market due to their therapeutic efficacy having tolerable side effects and thus challenging the predominance of well established beta-lactam antibiotics which are becoming more prone to the resistant pathogenic bacteria. Since the discovery of nalidixic acid the development of fluoroquinolones has experienced an exponential growth and is being continued with more vigor to obtain better drugs having multifunctional action. This article attempts to review the current developments of the chemical and biological aspects of fluoroquinolones in a chronological manner touching upon their antibacterial properties based on the structure activity relationship while pointing out to their mode of action. It also provides an insight into a variety of approaches resulting in elegant manipulations of their basic skeleton and some breakthroughs in their synthetic strategies of a few widely used drugs, which had helped in accelerating their market growth as well as continuing research for newer fluoroquinolones. Since the mode of action of fluoroquinolones being different from beta-lactams and their transportation to the target site is slow several dual action quinolonyl-beta-lactams (Penicillins, Cephalosporins, Penems, Cephems, Carbapenams etc.) have come as a major breakthrough among the hybrid antibiotics. While focusing on the multifunctional activities of such compounds, this review briefly points out to the current trends in various techniques for de novo drug design and development of newer therapeutic molecules, which hold future promises in combating the fight against drug resistant bacteria as it still remains to be won. PMID- 11254893 TI - Type II topoisomerases as targets for quinolone antibacterials: turning Dr. Jekyll into Mr. Hyde. AB - Quinolones are a very important family of antibacterial agents that are widely prescribed for the treatment of infections in humans. Although the founding members of this drug class had little clinical impact, successive generations include the most active and broad spectrum oral antibacterials currently in use. In contrast to most other anti-infective drugs, quinolones do not kill bacteria by inhibiting a critical cellular process. Rather, they corrupt the activities of two essential enzymes, DNA gyrase and topoisomerase IV, and induce them to kill cells by generating high levels of double-stranded DNA breaks. A second unique aspect of quinolones is their differential ability to target these two enzymes in different bacteria. Depending upon the bacterial species and quinolone employed, either DNA gyrase or topoisomerase IV serves as the primary cytotoxic target of drug action. While this unusual feature initially stymied development of quinolones with high activity against Gram-positive bacteria, it ultimately opened new vistas for the clinical use of this drug class. In addition to the antibacterial quinolones, specific members of this drug family display high activity against eukaryotic type II topoisomerases, as well as cultured mammalian cells and in vivo tumor models. These antineoplastic quinolones represent a potentially important source of new anticancer agents and provide an opportunity to examine drug mechanism across divergent species. Because of the clinical importance of quinolones, this review will discuss the mechanistic basis for drug efficacy and interactions between these compounds and their topoisomerase targets. PMID- 11254894 TI - Helicobacter pylori: current chemotherapy and new targets for drug design. AB - Helicobacter pylori infection is a major cause of many diseases of the gastrointestinal tract, including gastritis, non-ulcer dyspepsia, peptic ulcer disease, and gastric cancers. It is estimated that more than half of the human race is affected by this organism. Although effective treatments are available which will eliminate the organism in about 90 percent; of cases in developed countries, the pandemic occurrence of Helicobacter pylori infection coupled with its ability to develop resistance to our current arsenal of antimicrobial regimens and subsequently reinfect patients makes the pathogenic potential of this microorganism a major global health concern. Provided is a review of the current and evolving therapeutic regimens used in the eradication of Helicobacter pylori, the difficulties associated with in vitro drug screening, as well as potentially new therapeutic targets. In addition, the discovery, the unique physiology, biochemistry, and pathogenicity of this remarkable microorganism is examined. PMID- 11254895 TI - Lipid-based antifungal agents: current status. AB - Immunocompromised patients are well known to be predisposed to developing invasive fungal infections. These infections are usually difficult to diagnose and more importantly, the resulting mortality rate is high. The limited number of antifungal agents available and their high rate of toxicity are the major factors complicating the issue. However, the development of lipid-based formulations of existing antifungal agents has opened a new era in antifungal therapy. The best examples are the lipid-based amphotericin B preparations, amphotericin B lipid complex (ABLC; Abelcet), amphotericin B colloidal dispersion (ABCD; Amphotec or Amphocil), and liposomal amphotericin B (AmBisome). These formulations have shown that antifungal activity is maintained while toxicity is reduced. This progress is followed by the incorporation of nystatin into liposomes. Liposomal nystatin formulation is under development and studies of it have provided encouraging data. Finally, lipid-based formulations of hamycin, miconazole, and ketoconazole have been developed but remain experimental. Advances in technology of liposomes and other lipid formulations have provided promising new tools for management of fungal infections. PMID- 11254896 TI - The impact of phospholipid transfer protein (PLTP) on HDL metabolism. AB - High-density lipoproteins (HDL) play a major protective role against the development of coronary artery disease. Phospholipid transfer protein (PLTP) is a main factor regulating the size and composition of HDL in the circulation and plays an important role in controlling plasma HDL levels. This is achieved via both the phospholipid transfer activity of PLTP and its capability to cause HDL conversion. The present review focuses on the impact of PLTP on HDL metabolism. The basic characteristics and structure of the PLTP protein are described. The two main functions of PLTP, PLTP-mediated phospholipid transfer and HDL conversion are reviewed, and the mechanisms and control, as well as the physiological significance of these processes are discussed. The relationship between PLTP and the related cholesteryl ester transfer protein (CETP) is reviewed. Thereafter other functions of PLTP are recapitulated: the ability of PLTP to transfer cholesterol, alpha-tocopherol and lipopolysaccharide (LPS), and the suggested involvement of PLTP in cellular cholesterol traffic. The discussion on PLTP activity and mass in (patho)physiological settings includes new data on the presence of two forms of PLTP in the circulation, one catalytically active and the other inactive. Finally, future directions for PLTP research are outlined. PMID- 11254897 TI - The potential role of adenosine in the pathophysiology of the insulin resistance syndrome. AB - An increased intracellular availability of the co-enzyme A esters of long-chain fatty acids is thought to underlie many aspects of the insulin resistance syndrome. However, the cause of clustering of a hyperdynamic circulation, sympathetic activation, hypertension, hyperuricaemia, and a raised haematocrit in the insulin resistance syndrome remains to be elucidated. We propose a mechanism that expands the etiological role of long-chain fatty acids. By inhibiting adenine nucleotide translocators, elevated intracellular concentrations of the co enzyme A esters of long-chain fatty acids impair mitochondrial oxidative phosphorylation. This is expected to result in a chronic systemic increase in extracellular adenosine concentrations. As adenosine stimulates the sympathetic nervous system, induces systemic vasodilatation, stimulates erythropoiesis, and induces renal vasoconstriction with renal sodium retention, increased extracellular ADO concentrations may be the common denominator explaining the above-mentioned and still unexplained phenomena associated with the insulin resistance syndrome. Along the same lines, hyperuricaemia can be explained by the fact that adenosine is broken down to urate and because of increased renal urate retention. PMID- 11254899 TI - Acute-phase HDL in phospholipid transfer protein (PLTP)-mediated HDL conversion. AB - In reverse cholesterol transport, plasma phospholipid transfer protein (PLTP) converts high density lipoprotein(3) (HDL(3)) into two new subpopulations, HDL(2) like particles and prebeta-HDL. During the acute-phase reaction (APR), serum amyloid A (SAA) becomes the predominant apolipoprotein on HDL. Displacement of apo A-I by SAA and subsequent remodeling of HDL during the APR impairs cholesterol efflux from peripheral tissues, and might thereby change substrate properties of HDL for lipid transfer proteins. Therefore, the aim of this work was to study the properties of SAA-containing HDL in PLTP-mediated conversion. Enrichment of HDL by SAA was performed in vitro and in vivo and the SAA content in HDL varied between 32 and 58 mass%. These HDLs were incubated with PLTP, and the conversion products were analyzed for their size, composition, mobility in agarose gels, and apo A-I degradation. Despite decreased apo A-I concentrations, PLTP facilitated the conversion of acute-phase HDL (AP-HDL) more effectively than the conversion of native HDL(3), and large fusion particles with diameters of 10.5, 12.0, and 13.8 nm were generated. The ability of PLTP to release prebeta from AP-HDL was more profound than from native HDL(3). Prebeta-HDL formed contained fragmented apo A-I with a molecular mass of about 23 kDa. The present findings suggest that PLTP-mediated conversion of AP-HDL is not impaired, indicating that the production of prebeta-HDL is functional during the ARP. However, PLTP-mediated in vitro degradation of apo A-I in AP-HDL was more effective than that of native HDL, which may be associated with a faster catabolism of inflammatory HDL. PMID- 11254898 TI - Omapatrilat, a dual angiotensin-converting enzyme and neutral endopeptidase inhibitor, prevents fatty streak deposit in apolipoprotein E-deficient mice. AB - Angiotensin-converting enzyme (ACE) is mainly responsible for converting angiotensin I (AI) to angiotensin II (AII), and ACE inhibitors prevent atherosclerosis in animal models. Neutral endopeptidase 24.11 (NEP) degrades substance P, kinins and atrial natriuretic peptide (ANP), and aortic wall NEP activity was found to be increased in atherosclerosis. In the present study, we have evaluated the effect of candoxatril, a NEP inhibitor, and of omapatrilat, a dual ACE and NEP inhibitor, on the development of fatty streak in apolipoprotein E (apoE)-deficient mice. Groups of ten male apoE-deficient mice were given either placebo, candoxatril 50 mg/kg per day, or omapatrilat 10, or 100 mg/kg per day for 4 months. None of the treatments influenced body weight, serum total or HDL cholesterol. Compared with the placebo, candoxatril did not protect the mice from fatty streak deposit. In contrast, omapatrilat dose dependently inhibited the constitution of fatty streak in apoE-deficient mice. The precise advantages of the dual ACE and NEP inhibition versus the inhibition of only ACE should now be considered in the prevention of atherosclerosis as well as in the occurrence of its complications. PMID- 11254900 TI - Effects of high-cholesterol diet on the interendothelial clefts and the associated junctional complexes in rat aorta. AB - The arterial endothelial intercellular cleft (AEC) and its associated junctional complex (JC) are the determinants of permeability to macromolecules. This study analyzed frequencies of AEC and JC profile types in the rat thoracic aorta at 1 and 12 months after feeding the animals with a normal or a high-cholesterol diet. Rats on either a normal diet or high-cholesterol diet for 12 months showed more of the simple 'end to end' or 'overlap' types (P < 0.01) but fewer complex 'interdigitating' type (P < 0.01) of AEC compared to the 1 month group. With regard to JC, the frequencies of gap junctions were decreased (P < 0.01) while the tight junctions and the normal junctionless complex were increased (P < 0.01) after 12 months of normal diet as compared with 1 month on the normal diet. These changes in frequencies for gap junction and tight junction were even greater for the high-cholesterol diet than for the normal diet treatment. Moreover, the incidence of open junctions was also noticeably increased after 12 months of high cholesterol diet. These findings suggest that the proportions of the AEC and JC were highly responsive to aging whereas those of JC were more susceptible to the high-cholesterol diet treatment. PMID- 11254901 TI - Effect of chronic insulin treatment on NO production and endothelium-dependent relaxation in aortae from established STZ-induced diabetic rats. AB - The hypothesis that the impaired endothelial function seen in streptozotocin (STZ)-induced diabetic rats may result from an increased nitric oxide (NO) metabolism was tested. Acetylcholine (ACh) increased the nitrite NO(2-) and nitrate (NO(3-)) levels in the perfusates from both control and diabetic aortic strips, although the level of NO(2-) was significantly lower in diabetic rats while the NO(3-) level was significantly higher. Both effects (decrease in NO(2-) and increase in NO(3-)) were ameliorated by chronic administration of insulin to diabetic rats but NOx (NO(2-) plus NO(3-)) was increased. The expression of endothelial nitric oxide synthase (eNOS) was significantly increased by chronic administration of insulin to diabetic rats. A decrease in NO(2-) and an increase in NO(3-) occurred following treatment of control aortae with hypoxanthine/xanthine oxidase. Incubating diabetic aortic strips with superoxide dismutase (SOD) normalized the production of both NO(2-) and NO(3-). Both the basal and the ACh-stimulated production of O(2)(-) were significantly higher in diabetic rats than in controls. These results demonstrate that the ACh-induced relaxation of aortic strips was significantly impaired in diabetic rats and that this impairment may be due to an abnormal oxidative metabolism of NO, rather than to a decrease in NOS mRNA and NO production. PMID- 11254902 TI - Rho-Rho kinase is involved in smooth muscle cell migration through myosin light chain phosphorylation-dependent and independent pathways. AB - Although Rho, a small GTPase, has been demonstrated to play an important role in the smooth muscle contraction and relaxation, little is known about the involvement of Rho protein in smooth muscle cell (SMC) migration. In this study the role of Rho-Rho kinase pathway was examined in SMC migration induced by platelet-derived growth factor (PDGF) and lysophosphatidic acid (LPA). C3 transferase, a specific inhibitor of Rho, blocked SMC migration induced by PDGF and LPA. Y-27632, a specific inhibitor of Rho kinase, a direct target molecule of Rho, inhibited PDGF and LPA-induced SMC migration in a concentration dependent manner. Although rapid increase in myosin light chain (MLC) phosphorylation in SMC treated with LPA was observed, no enhanced MLC phosphorylation was detected in response to PDGF. Y-27632 suppressed LPA-induced as well as basal level of MLC phosphorylation. ML-9, a specific inhibitor of myosin light chain kinase (MLCK), inhibited PDGF and LPA-induced SMC migration without the suppression of MLC phosphorylation at 5 min incubation, suggesting that MLCK may contribute to SMC migration via mechanism other than MLC phosphorylation. These results suggest that Rho-Rho kinase pathway is implicated in SMC migration and that different signaling pathways downstream of Rho-Rho kinase may be involved in LPA and PDGF induced SMC migration. MLC phosphorylation via Rho-Rho kinase pathway appears to be implicated in LPA-dependent SMC migration. Whereas PDGF-mediated SMC migration is independent of increased MLC phosphorylation and other target molecules downstream of Rho-Rho kinase seem to be involved. PMID- 11254903 TI - Macro-porosity is necessary for the reduction of neointimal and medial thickening by external stenting of porcine saphenous vein bypass grafts. AB - BACKGROUND: placing external non-restrictive macro-porous stents around porcine vein grafts prevents neointima formation and medial thickening in both the short and long term. Whether the porosity of the stent material influences this effect, however, has not been determined. Therefore, the effect on neointimal and medial thickening of external macro-porous (polyester) and micro-porous (polytetrafluorethylene) stents of equal diameter were compared. The effect on expression of platelet-derived growth factor (PDGF), a potent mediator of vascular smooth muscle cell migration and proliferation and its receptors was also investigated. METHODS AND RESULTS: saphenous vein-carotid artery interposition grafting was performed in Landrace pigs with external placement of 8 mm diameter macro- and micro-porous stents contralaterally. One month after surgery, graft wall dimensions, PDGF and PDGF receptor expression and cell proliferation using proliferating cell nuclear antigen (PCNA) were measured on histological sections. Macro-porous stents significantly reduced neointimal and medial thickening compared with micro-porous stents (0.1+/-0.02 vs. 0.25+/-0.03 mm, P<0.002, and 0.10+/-0.02 vs. 0.17+/-0.02 mm, P<0.014, respectively). Macro porous stents significantly reduced the percentage of cells expressing PDGF and PCNA, compared with micro-porous stents (36+/-9 vs. 80+/-7, P < 0.002, and 11+/-3 vs. 21+/-2, P < 0.02, respectively). The percentage of cells expressing PDGF receptors was similar with both the stent types. Adventitial microvessel formation occurred across macro-porous stents but was markedly suppressed by micro-porous stents. CONCLUSIONS: porosity is crucial to the efficacy of external stents in reducing neointima formation in porcine vein grafts. Decreases in PDGF expression and cell proliferation accompany the reduction in neointima formation. In addition, macro-porous stents allow adventitial microvessels to connect with the vasculature outside the stent, thereby potentially improving oxygenation. Although external stenting is highly effective in reducing neointima formation after vein grafting, the properties of the stent material necessary for this effect have not been defined. This study establishes that macro-porosity is one essential feature required to reduce PDGF expression cell proliferation and neointima formation. PMID- 11254904 TI - Medial thickenings of coronary artery and the aging risk factor for atherosclerosis. AB - The objective here is to inquire what kind of coronary artery is it that tends to acquire atheroma: When an atheroma is found somewhere in the specimen (YesA specimen), what do we see in the specimen far away from the atheroma? Previous studies found thicker intima in YesA specimens than in NoA specimens, but with equal numbers of smooth muscle cells (SMC's). Thickness per SMC strongly predicted atheroma, so much so that the risk factor age was fully explained statistically. This study now finds that the medial layer is also thicker in YesA specimens, and with medial SMC numbers equal to those in NoA specimens. Hence, the aging risk factor appears to induce excessive thickness per SMC as a generalized property throughout the whole specimen in the medial as well as intimal layers, with excessive production of collagenous matrix acting as an initial, rate limiting step in plaque formation. In the intima, atheroma tends to occur when average thickness per SMC exceeds the threshold value of 8.6 microm/SMC. The extreme high value found in the most severely affected medial sample was 4.2 microm/SMC, and this failure to approach the threshold could explain the medial resistance to fatty degenerations. PMID- 11254905 TI - A HMG-CoA reductase inhibitor possesses a potent anti-atherosclerotic effect other than serum lipid lowering effects--the relevance of endothelial nitric oxide synthase and superoxide anion scavenging action. AB - We have determined whether the anti-atherosclerotic effect of a 3-hydroxy-3 methyl-glutaryl-CoA (HMG-CoA) reductase inhibitor (fluvastatin) is mediated through nitric oxide (NO) as well as affecting plasma lipids. NO related vascular responses, endothelial nitric oxide synthase (eNOS) mRNA and superoxide anion (O(2)(-)) release were examined in vascular walls of oophorectomized female rabbits fed 0.5% cholesterol chow for 12 weeks with or without fluvastatin (2 mg/kg per day). Serum lipid profile was not different between two groups. NO dependent responses stimulated by acetylcholine and calcium ionophore A23187 and tone related basal NO response induced by N(G)-monomethyl-L-arginine acetate (L NMA); nitric oxide synthase inhibitor were all improved by fluvastatin treatment. Endothelium independent vasorelaxation induced by nitroglycerin was not different between the two groups of rabbits' arteries. Fluvastatin treatment increased cyclic GMP concentration in aorta of rabbits. eNOS mRNA expression and O(2)(-) release were measured in aorta using competitive reverse transcription-polymerase chain reaction (RT-PCR) and with lucigenin analogue, 2-methyl-3,7-dihydroimidazol [1,2-a]pyrazine-3-one (MCLA) chemiluminescence methods. eNOS mRNA in the endothelial cells of aorta was significantly up-regulated and O(2)(-) production was significantly reduced in fluvastatin treated rabbit aorta. Anti-macrophage staining area, but not anti-smooth muscle cell derived actin stained area in the aorta was also reduced by fluvastatin treatment. Conclusion, fluvastatin, a HMG CoA reductase inhibitor, retards the initiation of atherosclerosis formation through the improvement of NO bioavailability by both up-regulation of eNOS mRNA and decrease of O(2)(-) production in vascular endothelial cells, and this means that part of the anti-atherosclerotic effect of fluvastatin may be due to nonlipid factors. PMID- 11254906 TI - Preferential pharmacological inhibition of macrophage ACAT increases plaque formation in mouse and rabbit models of atherogenesis. AB - The cholesteryl ester, foam cell-enriched vulnerable plaque is a principle pharmacological target for reducing athero-thrombosis. Acyl CoA:cholesterol Acyl Transferase (ACAT) catalyzes the esterification of free cholesterol in intestine, liver, adrenal and macrophages, leading in the latter cells to intracellular cholesteryl ester accumulation and foam cell formation in the arterial intima. Previous studies suggested the existence of several isoforms of ACAT with different tissue distribution and this has largely been confirmed by molecular cloning of ACAT-1 and ACAT-2. We developed a series of ACAT inhibitors that preferentially inhibited macrophage ACAT relative to hepatic or intestinal ACAT based on in vitro assays and ex vivo bioavailability studies. Four of these compounds were tested in three models of atherosclerosis at oral doses shown to give sufficient bioavailable monocyte/macrophage ACAT inhibitory activity. In fat fed C57BL/6 mice, chow fed apo E-/- mice and KHC rabbits, the various ACAT inhibitors had either no effect or increased indices of atherosclerotic foam cell formation. Direct and indirect measurements suggest that the increase in plaque formation may have been related to inhibition of macrophage ACAT possibly leading to cytotoxic effects due to augmented free cholesterol. These results suggest that pharmacological inhibition of macrophage ACAT may not reduce, but actually aggravate, foam cell formation and progression. PMID- 11254907 TI - Dexamethasone inhibits macrophage accumulation after balloon arterial injury in cholesterol fed rabbits. AB - Macrophages play a critical role in the development and progression of atherosclerosis. This study was designed to examine the effect of the glucocorticoid, dexamethasone, (Dex), on macrophage accumulation after acute arterial injury. Twenty New Zealand white rabbits were fed a 2% cholesterol, 6% peanut oil, rabbit chow diet for one month prior to bilateral balloon dilatation of the femoral arteries. Ten rabbits received Dex (1 mg/kg, im.) the day before and then daily for 7 days after arterial injury; control rabbits received vehicle only. Seven days after injury, Dex treatment resulted in a 96% and 77% reduction (P < 0.002) in the mean number of macrophages accumulating in the intima and media, respectively. This effect was apparently not due to a reduction in the number of circulating monocytes or to the ability of monocytes from Dex treated animals to adhere to endothelium or migrate in response to a chemotactic signal, determined in vitro under static conditions. It was associated with a 61% reduction in monocyte chemoattractant protein-1 (MCP-1) antigen (P < 0.004) in the injured arterial wall (media+intima). Glucocorticoids may be useful in attenuating the inflammatory response and subsequent foam-cell accumulation after arterial injury. PMID- 11254908 TI - Interaction between monocytes and vascular endothelial cells induces adrenomedullin production. AB - Adrenomedullin (AM), a potent vasodilator peptide, has natriuretic effects, and its plasma concentration is elevated in cardiovascular diseases. In the present study, we investigated the induction of AM expression due to interactions between THP-1 cells (human monocytic cell line) and human umbilical cord vein endothelial cells (HUVECs). AM levels in the culture medium were measured by radioimmunoassay. The luciferase vector containing the 5'-flanking region of the human AM gene was transfected into either HUVECs or THP-1 cells. Addition of THP 1 cells to HUVECs for 48 h induced marked increases in AM levels, which were 16 fold higher than those of HUVECs alone. Luciferase vectors containing the 5' flanking region of human AM gene (pLCF-1534) were transferred into THP-1 cells or HUVECs. Addition of THP-1 cells to pLCF-1534-transfected HUVECs induced an increase in luciferase activity in cell lysates, which was 5-fold higher than that of the transfected HUVECs alone. In contrast, the luciferase activity of lysates from pLCF-1534-transfected THP-1 cells was not affected by coculture with HUVECs. A separate coculture experiment revealed that direct contact of THP-1 cells and HUVECs contributed to enhanced AM production in the cocoulture. Co incubation of the cell membrane fraction from THP-1 cells augmented AM production by HUVECs. Both anti-interleukin (IL)-1alpha antibody and IL-1 receptor antagonist significantly inhibited AM production in the cocultures. The cell-to cell interaction between monocytes and HUVECs induces AM production by HUVECs, which may play an important role in the pathogenesis of vascular disorders. PMID- 11254909 TI - Effects of heparin and aspirin on circulating P-selectin, E-selectin and von Willebrand Factor levels in healthy men. AB - As thrombin stimulates P-selectin expression on platelets and its release into plasma, we hypothesized that enhancing antithrombin activity by unfractionated heparin (UFH) could decrease plasma levels of circulating (c)P-selectin, (c)E selectin, and von Willebrand Factor (vWF). Hence the effect of UFH and aspirin were examined on these activation markers in healthy volunteers. UFH decreased cP selectin levels by -10% (CI: -16 - (-4%); P = 0.005) at 24 h, but did not change levels of vWF-Ag. In contrast, aspirin did not affect cP-selectin levels but decreased vWF-Ag levels by -12% (CI: -18 - (-7%); P = 0.005) at 24 h. Neither drug affected cE-selectin levels. Thus, UFH decreases cP-selectin levels, which may reflect decreased platelet activation in vivo. An increase in cP-selectin under UFH therapy should alert the clinician to look for platelet destruction. PMID- 11254910 TI - Fluvastatin therapy improves microcirculation in patients with hyperlipidaemia. AB - The purpose of this study was to investigate the effect of fluvastatin on the microcirculation of patients with hyperlipidaemia (low-density lipoprotein cholesterol > 160 mg/dL, triglycerides < 350 mg/dl) inadequately controlled by diet. After a dietary run-in of 4 weeks, patients were randomised in a double blind study to receive fluvastatin 40 mg twice daily (n = 24) or placebo (n = 24) for 12 weeks. The effect on microcirculation was assessed using capillary microscopy and laser Doppler fluxmetry at the nailfold at baseline and at 6 and 12 weeks after initiation of therapy. Capillaroscopy showed that fluvastatin improved microcirculation, i.e. time to peak flow during postocclusive reactive hyperaemia dropped from 19.7 +/- 7.2 s at baseline to 12.3 +/- 9.5 s at week 6 (P < 0.01) and 10.6 +/- 6.5 s at week 12 (P < 0.0001). These results were confirmed using laser Doppler fluxmetry to study microcirculation in thermoregulatory capillaries at the same site. A significant decrease in total and LDL-cholesterol was achieved during fluvastatin therapy. In conclusion, fluvastatin therapy improves microcirculation in nutritive as well as thermoregulatory capillaries in hypercholesterolaemic patients within 6 weeks. PMID- 11254911 TI - Oxidized LDL and thickness of carotid intima-media are associated with coronary atherosclerosis in middle-aged men: lower levels of oxidized LDL with statin therapy. AB - We investigated the relation between serum lipids including oxidized LDL and the severity of coronary atherosclerosis. Serum lipids and oxidized LDL was measured in 62 men (33-66 years), who underwent diagnostic coronary angiography and sonography to measure the carotid intima-media thickness. LDL oxidation was found in chemical analyses to be due to conjugated fatty acids in cholesteryl esters and triglycerides. Regression analysis indicated that the carotid intima-media thickness and the ratio of LDL diene conjugation to LDL cholesterol (the ox LDL:LDL ratio) were the only factors associated independently with the severity of coronary atherosclerosis. The patients with multi-vessel disease who did not use lipid lowering therapy had a 50% thicker carotid intima media (P = 0.030) and a 41% higher ox-LDL:LDL ratio (P = 0.020) than patients with normal vessels. Further, patients with multi-vessel disease on statin therapy had a 24% lower ox LDL:LDL ratio than the subjects with multi-vessel disease who did not use lipid lowering drugs (P = 0.027), although the concentration of LDL cholesterol did not differ between the groups. This study supports the hypothesis that lipid oxidation plays a role in the development of atherosclerosis. PMID- 11254912 TI - Immunohistochemical localization of Betacellulin, a member of epidermal growth factor family, in atherosclerotic plaques of human aorta. AB - Betacellulin (BTC), a new member of the EGF family, has been reported to be a potent mitogen for rat vascular smooth muscle cells (SMCs). BTC mRNA is known to be expressed in several human organs. However, the localization of BTC in human vascular tissues has not yet been clarified. We investigated whether or not BTC protein is involved in the pathogenesis of human atherosclerosis. Recombinant human BTC showed a mitogenic activity on cultured human aortic SMCs by measuring [3H]thymidine incorporation. The immunohistochemical localization of BTC, SMCs, macrophages, EGF receptors and ErbB4 was examined in autopsied human aortas. BTC was detected in both intimal and medial SMCs of the aortic wall. The percentage of BTC-positive medial SMCs in early types of atherosclerotic lesions decreased with age, but in adult, it was significantly higher in advanced types than in early types of atherosclerotic lesions. BTC-positive SMCs were predominantly localized in the medial side of the intima. Furthermore, numerous BTC-positive SMCs and macrophages were observed around the core lesion of atherosclerotic plaques. Receptors for BTC, EGF receptor and ErbB4, were expressed on SMCs, suggesting that BTC is associated with EGF receptor family-mediated signaling. BTC is produced in human aortic tissue and might play important roles in atherogenesis. PMID- 11254913 TI - Oral 17beta-estradiol and medroxyprogesterone acetate therapy in postmenopausal women increases HDL particle size. AB - Menopause is accompanied by changes in lipoprotein particles that include an increase in density of low density lipoproteins (LDL) and high density lipoproteins (HDL) particles. The effect of 3 months of oral hormone replacement therapy (HRT) on lipoprotein particle size in postmenopausal women who were randomized to (1) estrogen replacement therapy (ERT) alone (either 17beta estradiol (1 mg) or conjugated equine estrogens (CEE) (0.625 mg); (2) combination therapy (17beta-estradiol plus medroxyprogesterone acetate (MPA) or CEE plus MPA); and (3) placebo were examined. Lipoprotein subclass concentrations and particle size were quantified by nuclear magnetic resonance spectroscopy (NMR). Combination HRT resulted in significant (P=0.002) increases in HDL particle size as compared with those on placebo formulations or ERT alone. Women assigned to combined HRT had lower concentrations of smaller HDL particles after 3 months (P=0.005) and higher concentrations of larger HDL particles (P=0.02), whereas women assigned to ERT or placebo experienced non-significant changes. In summary, combined HRT increases HDL particle size by altering concentrations of the smallest and largest HDL subspecies. PMID- 11254914 TI - Left ventricular mass and correlated atherosclerotic risk factors in young adolescents: report from Chin-Shan community cardiovascular study in Taiwan. AB - Various subclinical disease indicators can be used as an early stage marker of atherosclerosis. Left ventricular (LV) mass has been related to cardiovascular morbidity and mortality. The distribution of LV mass in Chinese is rarely studied and nothing is known about its relationships with various atherosclerotic risk factors in young teenagers, in particular, aspects of lipid profiles. We performed a community-based survey of 523 males and 555 females, aged 12-15, in Chin-Shan, a suburb area near Taipei, Taiwan. LV mass was calculated from the Penn convention. Normalized LV mass by height with power of 2.7 was defined. LV mass and normalized LV mass were significantly greater in males than in females. There were significant positive correlation coefficients between LV mass and age, blood pressure, body mass index, low density lipoprotein cholesterol (LDL-C), apolipoprotein (Apo) B, fasting insulin levels and significant negative correlation coefficients between LV mass and high density lipoprotein cholesterol (HDL-C) and Apo A1 level in both genders. Multiple linear regression models showed gender and body mass index (BMI) were important factors associated with LV mass or normalized values for adolescents. Age and systolic blood pressure were also significant predictors of LV mass, but not of normalized LV mass values. LV mass values were found to be negatively associated with HDL-C values at marginal statistically significant level. Age and BMI are the most significant factors of echocardiographic LV mass distributions in young adolescent in Taiwan. LV mass may also be associated with atherosclerotic risk factors. PMID- 11254915 TI - Expression of very low density lipoprotein receptor mRNA in circulating human monocytes: its up-regulation by hypoxia. AB - Although very low density lipoprotein (VLDL) receptor expression by macrophages has been shown in the vascular wall, it is not clear whether or not circulating monocytes express the VLDL receptor. We investigated the expression of VLDL receptor mRNA in human peripheral blood monocytes and monocyte-derived macrophages by reverse transcriptase polymerase chain reaction (RT-PCR) and nucleotide sequencing after subcloning of PCR product. VLDL receptor mRNA was detected both in peripheral blood monocytes and monocyte-derived macrophages. Expression of VLDL receptor mRNA was upregulated by hypoxia in monocytes, whereas treatment with oxidized LDL, interleukin-1beta or monocyte chemoattractant protein-1 did not affect the levels of VLDL receptor mRNA in monocytes and macrophages. The present study shows a novel response of VLDL receptor mRNA to hypoxia, suggesting a role for VLDL receptor in the metabolism of lipoproteins in the vascular wall and the development of atherosclerosis. PMID- 11254916 TI - Coronary atherosclerosis and oxidative stress as reflected by autoantibodies against oxidized low-density lipoprotein and oxysterols. AB - Clinical studies and animal experiments have demonstrated that oxidized low density lipoprotein (oxLDL) and oxysterols play important roles in atherogenesis. OxLDL is immunogenic, and autoantibodies (Ab) against oxLDL are detectable in serum. We investigated the relevance of oxysterols and Ab against-oxLDL to coronary artery disease (CAD) in 183 patients undergoing coronary angiography. Patient groups included angiographically normal subjects (< 75% stenosis), others with spasm (> 75% narrowing in response to acetylcholine), and some others with fixed stenosis (> 75%). The group with stenosis was subdivided into patients with stable and unstable angina. Serum concentrations of autoantibodies and 25-, 27-, and 7-beta-hydroxycholesterols were significantly higher in the stenotic group than in the normal group (P < 0.01, P < 0.05, P < 0.05, and P < 0.05, respectively). Antibodies, but not oxysterol concentrations, were significantly greater in subjects with unstable than with stable angina (P < 0.01). We conclude that anti-oxLDL antibody and oxysterol concentrations are associated with coronary artery stenosis, and that oxidative stress may be greatly increased in unstable angina. PMID- 11254917 TI - Fenofibrate raises plasma homocysteine levels in the fasted and fed states. AB - The effect of fenofibrate (FEN), compared with placebo (PL), on total plasma homocysteine (tHcy) levels in the fasted and fed states has been examined. Twenty men with established coronary artery disease (CAD) or with at least two cardiovascular risk factors, who had elevated plasma triglyceride levels (> 2.3 mmol/l) and reduced HDL-C levels (< 0.91 mmol/l), and in whom a fibric acid derivative was clinically indicated were studied. The study was a randomized, PL controlled, double-blind study designed to test the effect of micronized FEN on postprandial lipemia. Plasma tHcy levels were investigated as a post-hoc analysis. After a 4-week dietary stabilization period, patients were randomized to PL or FEN (200 mg/day) for 8 weeks, followed by an 8-h postprandial study, consisting of 1 g fat/kg body weight (35% cream). The methionine content of cream was approximately 0.53 mg/ml. A 5-week washout period was then followed by a second 8-week treatment period (FEN or PL), at the end of which a second postprandial study was undertaken. Blood was sampled in the fasted state (0 h) and postprandially at 2, 4, 6 and 8 h. Plasma was stored at -80 degrees C for homocysteine, vitamins B(6), B(12) and folate measurements. FEN caused a marked decrease in all triglyceride-rich lipoprotein parameters, no change in LDL-C, and an increase in HDL-C levels. Fen treatment was associated with an increase in fasting tHcy (PL: 10.3+/-3.3 micromol/l to FEN: 14.1+/-3.8 micromol/l, 40.4+/ 20.5%, P < 0.001) and fed tHcy levels 6 h post-fat load (PL: 11.6+/-3.3 micromol/l vs. FEN: 17.1+/-5.4 micromol/l, P < 0.001). Homocysteine levels were increased by the fat load; PL: 14% (P < 0.001) and FEN: 21%, P < 0.001 at the 2, 4, 6 and 8 h time points. Change in tHcy level on FEN was not associated with changes in plasma levels of folate, vitamins B(6) or B(12) or creatinine. Amino acid analysis revealed that methionine and cysteine were significantly increased on FEN (P < 0.005). The incidence of hyperhomocysteinemia (defined as tHcy level >14 micromol/l) was PL: 2/20 (10%) and FEN: 9/20 (45%) (chi(2) = 4.51, P = 0.034). There was no correlation between changes in plasma triglyceride levels and tHcy levels. Since tHcy is considered an emerging cardiovascular risk factor, the ability of FEN to increase plasma tHcy levels could potentially mitigate the potential of this drug to protect against cardiovascular disease. PMID- 11254918 TI - Comparative study of HMG-CoA reductase inhibitors on fibrinogen. AB - Statins have a variable response on fibrinogen, and these changes may have implications on cardiovascular events. In this randomized placebo-controlled crossover study, we evaluated whether changes in fibrinogen levels were different between atorvastatin and other statin-treated patients. Adult coronary heart disease (CHD) patients aged 39-83 years with LDL cholesterol levels > or = 130 mg/dl were randomized to atorvastatin 80 mg (n = 84) and one of the following statins: fluvastatin 80 mg (n = 23), lovastatin 80 mg (n = 20), pravastatin 40 mg (n = 22) or simvastatin 40 mg (n = 20) each for 12 weeks in either order. Fibrinogen was analyzed by an automated method of Clauss. Three independently acquired samples were obtained within 1 min of tourniquet application, and each specimen was measured in duplicate. Statistical analyses were performed using a mixed model repeated measures analysis of variance procedure with SAS version 6.12. There were no significant changes in fibrinogen between treatment groups. This study evaluated changes in fibrinogen with established pre-analytical and analytical procedures known to minimize variability in fibrinogen measurement, and we did not observe any differences in fibrinogen levels in the treatment groups. PMID- 11254919 TI - Partially hydrogenated soybean oil reduces postprandial t-PA activity compared with palm oil. AB - The effects of dietary trans fatty acids on fasting and diurnal variation in hemostatic variables are not known. This study compares the effects of three diets with three different margarines, one based on palm oil (PALM-diet), one based on partially hydrogenated soybean oil (PHSO, TRANS-diet) and one with a high content of polyunsaturated fatty acids (PUFA-diet) on diurnal postprandial hemostatic variables. A strictly controlled dietary Latin square study was performed and nine young female participants consumed each of the diets for 17 days in a random order. The sum of the cholesterol-increasing fatty acids (C12:0, C14:0, C16:0) was 36.3% of total fatty acids in the PALM-diet, the same as the sum of saturated-(C12:0, C14:0, C16:0) (12.5%) and trans fatty acids (23.1%) in the TRANS-diet. The sum of C12:0, C14:0 and C16:0 was 20.7% in the PUFA-diet. The amount of fat made up 30-31% of energy in all diets. Nine participants completed the study. The diurnal postprandial state level of tissue plasminogen activator (t-PA) activity was significantly decreased on the TRANS-diet compared with the PALM-diet. t-PA activity was also decreased on the PUFA-diet compared with PALM diet but the difference was below statistical significance (P=0.07, Bonferonni adjusted). There were no significant differences in either fasting levels or in circadian variation of t-PA antigen, PAI-1 activity, PAI-antigen, factor VII coagulant activity or fibrinogen between the three diets. Our results indicate that dietary trans fatty acids from PHSO has an unfavourable effect on postprandial t-PA activity and thus possibly on the fibrinolytic system compared with palm oil. PMID- 11254920 TI - Human triglyceride-rich lipoprotein apo E kinetics and its relationship to LDL apo B-100 metabolism. AB - Apolipoprotein (apo) E is a multifunctional protein that can act as a ligand for lipoprotein receptors. The receptor-mediated clearance of the triglyceride-rich lipoproteins (TRL) chylomicrons and VLDL from plasma is, in part, dependent on apo E. Enrichment of VLDL with apo E is thought to enhance receptor-mediated clearance of VLDL resulting in a low rate of conversion of VLDL to LDL. However, the kinetic mechanism controlling the concentration of apo E in VLDL is not known. We conducted kinetic studies on apo E in the TRL fraction (d < 1.006 g/ml) and apo B-100 in the TRL and LDL (d = 1.019-1.063 g/ml) fractions to assess the kinetic determinants of apo E concentration in TRL and to determine the effects that TRL apo E production and clearance rates have on the production rate of LDL apo B-100. Nineteen males between the ages of 24 and 73 underwent a primed constant infusion with deuterated leucine tracer in the constantly-fed state. Apo B-100 from TRL and LDL, and apo E from TRL were isolated and their tracer incorporation measured by gas chromatography/mass spectrometry. The residence time and production rates of each protein were determined from the kinetic data using the SAAM II modeling program. The residence time and production rate of TRL apo E were about one-half that of TRL apo B-100 (1.8 +/- 1.0 vs. 2.9 +/- 2.1 h and 14.5 +/- 11.0 vs. 27.6 +/- 17.3 mg/kg per day, respectively). The production rate of TRL apo E was weakly correlated with the production rate of TRL apo B-100 (r = 0.424, P = 0.07). Multiple regression analysis showed that the residence time of TRL apo B-100 and the relative TRL apo E production rate (relative to the TRL apo B100 production rate) were negatively associated with LDL apo B-100 production rate, accounting for 68% of its variability. We conclude that (1) the concentration of apo E in TRL is highly correlated to its production rate, suggesting that production rate regulates the TRL apo E concentration, and (2) individuals with a relatively short TRL apo B-100 residence time and those producing TRL with a relatively low apo E content have the highest LDL apo B-100 production rates. PMID- 11254921 TI - Second nation-wide study of atherosclerosis in infants, children and young adults in Japan. AB - This paper reports the results of the second nation-wide cooperative study of atherosclerosis in young Japanese, aged from 1 month to 39 years, who were autopsied between 1991 and 1995. Atherosclerotic lesions in 1066 aortas and 974 coronary arteries were classified into fatty streaks, fibrous plaques and complicated lesions and quantificated with the point-counting method. The results of this study were compared with those of the former study, which was conducted 13 years earlier in almost the same fashion as this study. Atherosclerosis of aorta, which was determined by surface involvement (SI) of atherosclerotic lesions and atherosclerotic index (AI), increased with age in both sexes of the former and the present studies and their tendency for the progression of the extent of atherosclerotic lesions appeared to be similar. In the coronary arteries, the mean values of SI and AI in the males of the present study were greater significantly than those in the male of the former studies and in the female of the both studies in the third and fourth decades. This difference suggests that atherosclerotic lesions are increasing in young Japanese males. It also suggests that these subjects may be increasingly susceptible to atherosclerotic cardiovascular disease with increasing age. PMID- 11254922 TI - Coronary atherosclerosis in unheralded sudden coronary death under age 50: histo pathologic comparison with 'healthy' subjects dying out of hospital. AB - AIM: sudden coronary death (SCD) in older individuals is generally associated with extensive coronary atherosclerosis, although it may be the first manifestation of ischaemic heart disease. In younger age-groups, SCD may occur in the presence of less severe disease. We sought to (1) examine the extent of coronary atherosclerosis in young victims of SCD compared with age- and sex matched controls, (2) analyse the composition of atherosclerotic plaques in these patients, (3) identify the predominant mechanism of SCD, and (4) evaluate the possibility of detecting this mechanism on the basis of morphologic plaque features, in particular presence and amount of lipid accumulation and calcific deposits. METHODS AND RESULTS: coronary arteries were obtained at autopsy from 28 victims of SCD under age 50 with no prior clinical manifestation of ischaemic heart disease (IHD) and no myocardial scar formation and from 16 age- and sex matched subjects dying of noncardiac causes out of hospital. Sections of all available major coronary arteries were cut in 5-mm intervals to yield a total of 1357 histologic sections, which were analysed using digitised planimetry. Victims of SCD had significantly more major coronary arteries per subject with luminal area narrowing > or = 75% than controls (on average, 2.1 vs. 0.2). Plaque area per histologic section was 5.1 +/- 2.1 mm(2) in SCD cases and 2.0 +/- 0.9 mm(2) in controls (P < 0.001). The major constituent of all plaques was fibrous tissue. Lipid core area per section was 0.49 +/- 0.59 mm(2) in SCD cases and 0.004 +/- 0.01 mm(2) in controls (P < 0.001), and calcified plaque area was 0.18 +/- 0.19 mm(2) in SCD cases and 0.02 +/- 0.05 mm(2) in controls (P < 0.001), both defining significant differences between SCD cases and controls. Arterial thrombosis, most often with underlying plaque rupture was the mechanism of SCD in > 80% of the cases. Considering histologic sections with > or = 50 and with > or = 75% area stenosis, plaque rupture was independently predicted by lipid core area. Calcific deposits were a frequent feature of plaque rupture but were only associated with it in univariate analysis. CONCLUSIONS: the extent and severity of coronary atherosclerosis in young victims of SCD as the first manifestation of IHD was substantially greater than in age-and sex-matched controls and comparable with that previously reported in SCD cases with a broader age range. Lipid core and calcified plaque areas provided for excellent separation between the two groups, which may have implications for identifying persons at increased risk for SCD by non invasive visualisation and assessment of the coronary arteries. PMID- 11254923 TI - Apolipoproteins E and C-III in apo B- and non-apo B-containing lipoproteins in middle-aged women from the Stanislas cohort: effect of oral contraceptive use and common apolipoprotein E polymorphism. AB - Oral contraceptive (OC) use and common apo E polymorphism are well known to modify serum lipid and lipoprotein concentrations. The combined effect of OC use and apo E genotype on the concentration of apo E or apo C-III in apo B- (apo E LpB or apo C-III-LpB) or in non-apo B-containing lipoparticles (apo E-Lp-non-B or apo C-III-Lp-non-B) are unknown. Our study comprised 613 women, aged 30-45 years, genotyped for common apo E polymorphism and who differed in their combined low dose OC consumption. The concentrations of apo C-III, apo C-III-LpB and apo C-III Lp-non-B were significantly higher in OC users than in non-users by 13, 23 and 8% respectively, without significant interaction with the apo E genotype. The concentrations of apo E and apo E-Lp-non-B were significantly lower (differences being -14% and -31% respectively) in OC users than in controls whereas the apo E LpB concentration was significantly higher (+19%), resulting in a redistribution of apo E from Lp-non-B towards LpB. Total apo E and apo E-Lp-non-B concentrations were higher in subjects carrying the epsilon2 allele and lower in those with the epsilon4 allele when compared to epsilon3/epsilon3 subjects (P < 0.001). The opposite held for the apo E- LpB concentration (P < 0.05). The main finding is the significant interaction between apo E genotype and OC use (P < 0.01) on apo E Lp-non-B concentration, the epsilon4 carriers showing the smallest differences between OC users and non-users in comparison with the epsilon2 or epsilon3/epsilon3 carriers. These results suggest that the common apo E polymorphism can modulate the OC use effect. PMID- 11254926 TI - The effects of left- versus right-hemisphere lesions on the sensitivity to intra- and interconceptual semantic relationships. AB - Studies on split-brain, normal and brain-damaged subjects suggest differences in the processing of semantic relationships by the two hemispheres. Various authors have conceived of this distinction in terms of the paradigmatic/syntagmatic dissociation, and the connotative and denotative meanings of words, and as reflecting different types of links between words. Drews has suggested that a left-hemisphere lesion would affect the processing of intraconceptual relationships, while a right-hemisphere lesion would impair the processing of interconceptual relationships. The goal of this study was to test this hypothesis, using a number of intra- and interconceptual semantic relationships. Pairs of common words were submitted to left-hemisphere brain-damaged and right hemisphere brain-damaged subjects. The task required subjects to indicate whether or not there was any relationship between the words. The results only partially support the hypothesis. The right/left opposition applied to only one of three types of intraconceptual relationships (whole-part relation) and to one of two types of interconceptual relationship (same location relation). This partially unexpected result is discussed in reference to other studies. PMID- 11254924 TI - Postprandial hypertriglyceridemia impairs endothelial function by enhanced oxidant stress. AB - AIMS: it appears that hypertriglyceridemia (HTG) is a risk factor of atherosclerosis as demonstrated by recent studies. In this study, we analyzed the effects of acute HTG on endothelial function and oxidative stress, which are important mechanisms in the pathogenesis of atherosclerosis. METHODS AND RESULTS: in a high fat meal group (n = 11), serum triglycerides and PMA-activated leukocyte O(2)(-)* production were significantly (P < 0.005) increased from 146 +/- 69 mg/dl and 4.09 +/- 0.93 nmol/10(6) cells/min preprandially to 198 +/- 88 mg/dl and 5.49 +/- 1.19 nmol/10(6) cells/min, respectively, 2 h after eating a high-fat meal. The flow-mediated endothelium-dependent brachial artery dilation (FMD; high-resolution ultrasound) was decreased from 13.7 +/- 3.3% preprandially to 8.2 +/- 3.7%, 2 h after eating a high-fat meal (P < 0.005). However, following a low-fat meal (n = 9), there were no significant changes in triglycerides, leukocyte O(2)(-)* production and FMD. Changes of serum triglycerides were correlated negatively (r = -0.650, P < 0.005) with changes of FMD, but were correlated positively (r = 0.798, P < 0.001) with changes of leukocyte O(2)(-)* production, which - in turn - were correlated negatively (r = -0.784, P < 0.001) with changes of FMD in all study subjects (mean age: 56 years, n = 20). CONCLUSIONS: this study suggests that acute HTG causes endothelial dysfunction via enhanced oxidant stress and this may pave the way for the development of atherosclerosis under chronic conditions. PMID- 11254928 TI - Handedness, criminality, and sexual offending. AB - A very large database was used to investigate whether men with a history of criminality and/or sexual offending have a higher incidence of nonright handedness (NRH) relative to a control sample of nonoffender men. The sample (N>8000) comprised interviews by investigators at the Kinsey Institute for Sex and Reproduction in Indiana. The general offender group and a subsample of sex offenders (e.g. pedophiles) had a significantly higher rate of NRH relative to the control (nonoffender) men. In addition, evidence was found that the general criminality/NRH relationship might result from increased educational difficulties that some nonright-handers experience. In contrast, education was unrelated to the handedness/pedophilia relationship, suggesting that there may be a different mechanism underlying the handedness/pedophile relationship than the handedness/(general) criminality relationship. Finally, as a cautionary note, it is stressed that the effects are small and that NRH should not be used as a marker of criminality. PMID- 11254927 TI - Path integration following temporal lobectomy in humans. AB - Path integration, a component of spatial navigation, is the process used to determine position information on the basis of information about distance and direction travelled derived from self-motion cues. Following on from studies in the animal literature that seem to support the role of the hippocampal formation in path integration, this facility was investigated in humans with focal brain lesions. Thirty-three neurosurgical patients (17 left temporal lobectomy, LTL; 16 right temporal lobectomy, RTL) and 16 controls were tested on a number of blindfolded tasks designed to investigate path integration and on a number of additional control tasks (assessing mental rotation and left-right orientation). In a test of the ability to compute a homing vector, the subjects had to return to the start after being led along a route consisting of two distances and one turn. Patients with RTL only were impaired at estimating the turn required to return to the start. On a second task, route reproduction was tested by requiring the subjects to reproduce a route consisting of two distances and one turn; the RTL group only were also impaired at reproducing the turn, but this impairment did not correlate with the homing vector deficit. There were no group differences on tasks where subjects were required to reproduce a single distance or a single turn. The results indicate that path integration is impaired in RTL patients only and suggest that the right temporal lobe plays a role in idiothetic spatial memory. PMID- 11254929 TI - Asymmetric influences of pointing on saccade latency in hemi-Parkinson's disease. AB - The objective of this study was to investigate whether eye-hand coupling was preserved or not in PD. We studied predictive saccade performance during hand pointing in six Parkinson's disease (PD) patients with asymmetrical motor signs compared to nine age-matched healthy subjects. The motor responses (saccades and hand pointing) were elicited under open loop conditions (without vision of the hand), by a visual target stepping at a predictable location (10 degrees right and left from the centre) and time. The subjects had to simultaneously move the eyes and point with the finger to the visual target alternating at one of three fixed frequencies (0.25, 0.5 and 1 Hz), for 30 cycles. This task was performed in two sessions balanced over the subjects: one session of ocular saccades only and another session of combined ocular saccades and manual pointing. In the PD group, motor performance was perturbed particularly in terms of increased latencies of hand movements. Interestingly, during pointing, associated predictive saccade disorders were tightly related to the defects of the pointing hand. Indeed, with respect to the latency of predictive saccades alone, the predictive saccade latency during hand pointing significantly decreased in the control group and in the PD group when using the non-affected hand. In contrast, for the PD group when using the affected hand, the saccade latency was increased from the latency values of predictive saccades induced without pointing. Moreover, in the control and in the PD groups, the correlation between eye and hand latencies was highly significant, suggesting an intact eye-hand coupling. No saccadic amplitude disorders were found in either condition. These results demonstrate that eye-hand coupling is preserved in PD, as revealed by the possible beneficial or adverse effects on the ocular saccades, respectively, of the less- or more-affected hand motor responses. This eye-hand coupling mechanism likely involves regions other than the nigro-striatal pathways affected in PD. PMID- 11254930 TI - Parametric control of fingertip forces during precision grip lifts in children with DCD (developmental coordination disorder) and DAMP (deficits in attention motor control and perception). AB - Twenty boys with developmental coordination disorder (DCD), 11 of whom had associated attention deficit disorder (ADD), were compared with an age-matched control group of 12 boys to examine mechanisms that adapt the grip force at the digit-object interface in a precision grip task. An experimental grip object equipped with pressure transducers registered the grip forces (normal to the surface) and the load force (tangential to the surface) generated by the fingertips. The surface of the object was changed to vary the frictional properties. Both study groups exhibited disturbances of the basic coordination of forces in the initial phase of the movement, manifested by longer time latencies and higher force levels than the control group. All subjects were able to adapt the force output in response to the friction at the digit-object interface. Higher grip forces and safety margins were documented for the DCD group in comparison to the controls. Furthermore, there was greater variation in the parametric control of the grip force in the DCD group. The results suggest that the control of the grip force is similar in children with DCD, regardless of whether they have associated ADD or not, but it is impaired in comparison to that of controls. PMID- 11254931 TI - Event-related potential correlates of verbal and pictorial feature comparison in aphasics and controls. AB - Event-related potentials (ERPs) were recorded from 19 aphasic patients and 18 controls in four versions of a feature comparison task, in which the verbal or pictorial representation of a first stimulus (S1) had to be compared with the verbal or pictorial representation of a second stimulus (S2) presented 2 s later. These tasks were designed to cover some of the discriminatory variance of the token test (TT) including the analytical isolation, encoding and short-term storage of individual features of objects, independent of auditory verbal comprehension. Aphasics made more errors and had longer response latencies than controls in all four tasks, performance being poorest when verbal stimuli had to be processed. ERP analyses - restricted to subjects performing well above chance and to trials with correct responses - were confined to the slow wave (SW) (250 750 ms post-S1-onset) and the contingent negative variation (CNV) preceding the S2. There was no overall group difference that would have suggested that the patients activated different cortical areas than controls on correct performance. A left-hemispheric predominance of the negative SW was found in all four tasks and in both groups, although it was more pronounced in aphasics, and more pronounced in non-fluent than in fluent aphasics. The CNV was characterized by a left-hemispheric accentuation which was more pronounced in controls than in aphasics, particularly in tasks with a verbal S2. Results indicate that successful feature comparisons in the present tasks activate primarily left anterior cortical areas. During encoding and short-term storage this activation is more pronounced in aphasics than in controls. PMID- 11254932 TI - New questions on the hemispheric encoding/retrieval asymmetry (HERA) model assessed by divided visual-field tachistoscopy in normal subjects. AB - According to the hemispheric encoding/retrieval asymmetry (HERA) model, based on data obtained through functional neuroimaging, the left and right prefrontal cortices are preferentially, and, respectively, involved in long-term episodic memory encoding and retrieval. In this study, the HERA model was tested from a behavioral perspective using divided visual-field tachistoscopy. A recognition paradigm with both verbal and visuospatial materials was devised to differentiate memory-related effects (encoding vs. retrieval) from effects linked to the materials. The paradigm used lists of 12 and four items to assess long-term episodic memory and short-term memory, respectively. The aim of the latter condition was to test whether the HERA model is applicable in short-term memory. For long-term episodic memory, the data obtained validated the HERA model; the direction of the hemispheric asymmetry was found to depend on the type of materials used, whereas its magnitude was determined by the type of memory process. For verbal short-term memory, the HERA model seems to be confirmed. The pre-existing representations of the material could take into account the similarity of the hemispheric asymmetry pattern between short-term memory and long-term memory. In contrast, for visuospatial short-term memory, Baddeley's working memory model seems to better explain our results insofar as the asymmetries were essentially linked to the material in encoding but not in retrieval. This latter difference between short-term memory and long-term indicates that processes involved in LTM depend on episodic processes per se, hence, lending more support for the HERA model. Accordingly, these two memory systems seem to bring into play two different modes of hemisphere specialization. PMID- 11254933 TI - Dynamic position sense during a cyclical drawing movement: effects of application and withdrawal of tendon vibration. AB - The present study addressed the stability of dynamic position sense across time as well as the resetting of position sense following its disturbance by means of tendon vibration. Blindfolded subjects drew circles within a specific location of the workspace for a duration of 28 s and at a repetition rate of 1 s(-1). To study the stability of dynamic position sense, the changes in circle drawing performance across time were studied (control condition). Resetting of dynamic position sense was studied by application and subsequent withdrawal of biceps tendon vibration during movement (vibration condition). The results showed that the spatial characteristics of circle drawing in the control condition did not change significantly over the course of the 28 s trial, suggesting that dynamic position sense does not drift systematically across time. Whereas vibration leads to a decrease in diameter, a deterioration of the circularity of the pattern, and a drift of the hand movement toward the body, a restoration of these features was obtained during withdrawal of vibration. This suggests that subjects are able to reset dynamic position sense to reasonable values without the help of vision during active cyclical movement. PMID- 11254934 TI - Decision making in patients with spinal cord damage: afferent feedback and the somatic marker hypothesis. AB - Damasio has proposed an influential model of human decision making - the somatic marker hypothesis (Damasio AR. Descates' Error. London: Papermac/Macmillan, 1994), where he argues that somatic feedback to the brain influences decision making in man. It is proposed that when choosing between options that differ in relative risk, a somatic marker (e.g. a 'gut feeling') feeds back to the brain and influences cognitive appraisal. In the present study patients who had suffered a complete tetraplegia at the level of the sixth cervical vertebra were compared with matched healthy control subjects. As the spinal injury group have reduced somatic/peripheral feedback via the spinal cord, it was predicted, based on the somatic marker hypothesis, that they may demonstrate riskier behaviour than controls. All subjects completed the Iowa Gambling Task, a computerised card playing game where the player is instructed to try and win as much money as possible over 100 selections from one of four decks. The rules are not disclosed in advance, and the player gradually 'learns' that two of the decks are 'high risk' and lead to significant financial losses. Healthy individuals have previously been shown to learn to avoid the risky decks, whereas patients with medial frontal lobe damage (Bechara A, Damasio AR, Damasio H, Anderson SW. Insensitivity to future consequences following damage to human prefrontal cortex. Cognition 1994;50:7-15) and those with peripheral neuropathy (Bechara A, Tranel D, Wilson J, Heberlein AS, Ross M, Damasio AR, 1998. Impaired decision-making in peripheral neuropathy. Society for Neuroscience Abstracts 24:1176) select an excessive number from the risky decks, and consequently lose money. In the present study there were no significant differences between the spinal sectioned and healthy control groups in either card selection strategy or net financial outcome. This result suggests that in terms of the somatic marker hypothesis, feedback to the brain from the periphery via the cranial vagus and other nerves and the hormonal route may be equally or more influential than afferent feedback transmitted via the spinal cord. PMID- 11254935 TI - Processing of rapid auditory information in epileptic patients with left temporal lobe damage. AB - The role of the temporal lobes in processing time-related (temporal) information was tested in a task of anisochrony (or irregularity) discrimination assessing the temporal processing of sequential auditory information according to different presentation rates or tempos (between 80 and 1000 ms inter-onset intervals, IOI). Epileptic patients with either left (LHA, n=8) or right (RHA, n=10) hippocampal atrophy participated in this study as well as six normal control subjects (NC). For all the subjects, an effect of tempo was obtained, thresholds deteriorating for the 80 ms IOI as compared to longer IOI (the latter conforming to Weber's law). Furthermore, there was a specific impairment of rapid anisochrony discrimination (80 ms IOI) for patients with LHA as compared to patients with a RHA and NC, whereas no significant deficit was observed in these patients for longer IOI. These findings are in agreement with studies carried out in patients with left-hemisphere damage associated with massive language disorders and suggest the specialisation of left medial temporal lobe structures in processing rapid sequential auditory information. However, these results might also be explained by an auditory memory deficit in the presence of left temporal lobe dysfunction, as opposed to a specific impairment of fast sequential temporal processing. PMID- 11254938 TI - Interleukin-11 (IL-11) in human endometrium: expression throughout the menstrual cycle and the effects of cytokines on endometrial IL-11 production in vitro. AB - Interleukin-11 is a member of the IL-6 family of cytokines. Its presence in mouse decidua has been shown and experiments in genetically modified mice have suggested the importance of its receptor in stromal cell decidualization. In this study we used immunocytochemistry to determine expression of IL-11 in human endometrium. The effects of TNFalpha, IL-1alpha and TGFbeta on IL-11 production by epithelial and stromal cells was also investigated. Immunocytochemical staining in sections cut from 19 endometrial biopsies obtained throughout the menstrual cycle showed that IL-11 was expressed in both human endometrial epithelial and stromal cells, with epithelial staining being more intense than that seen in the stromal cells, at all times except the late secretory phase when the intensity was similar. Basal IL-11 production by cultured epithelial cells was greater than basal production by stromal cells. IL-1alpha, TNFalpha and TGFbeta (0.1-10 ng/ml) all caused a concentration-dependent increase in IL-11 production by both epithelial and stromal cells, but stimulated: basal values were greater for stromal than epithelial cells for all three cytokines. This work shows, for the first time, the presence of IL-11 within the human endometrium and that its production is controlled by other cytokines, which are postulated to play a role in implantation. Thus IL-11 may also play an important role in human endometrial function and embryo implantation. PMID- 11254936 TI - Identifying and separating the effects of practice and of cognitive ageing during a large longitudinal study of elderly community residents. AB - In protracted longitudinal studies of cognitive changes in old age volunteers must be repeatedly tested. Even with intervals of several years between assessment, this raises the possibility that improvements due to practice mask other changes. This problem is much more acute in brief studies of cognitive changes associated with progressive pathologies such as Alzheimer's disease or the effects of clinical interventions. Both types of study also encounter problems of selective dropout of frail and less able individuals leaving relatively 'elite' survivors. An analysis of data from repeated testing at 2-3 years intervals on the AH4 (1) intelligence test is presented to illustrate how a random effects model can be used to identify and disassociate age-related changes and practice effects at the population level, after effects of selective dropout and of background demographical variables have been taken into consideration. This analysis also provides some new, substantive empirical findings. Age-related changes are relatively slight between 49 and 70 years but much more marked between 70 and 80 years. Even with assessment points, several years apart the population average effect of practice is large relative to that of age-related change. Variation between individuals increases as samples age, providing the first clear evidence from a longitudinal study for marked individual differences in trajectories of cognitive ageing. PMID- 11254939 TI - Inhibition of antigen transport by expression of infected cell peptide 47 (ICP47) prevents cell surface expression of HLA in choriocarcinoma cell lines. AB - Cell surface expression of HLA class I (including non-classical HLA-G) in JEG3 (choriocarcinoma cell line) was blocked by stable transfection with the sequence encoding the Herpes simplex virus protein, infected cell peptide 47 (ICP47) inserted into a vector pCEP4. Intracellular expression of ICP47 protein in ICP47 transfected cells was demonstrated. The lack of HLA cell surface expression was likely to be due to blockage of peptide transport from the cytoplasm into the endoplasmic reticulum by ICP47. ICP47 is known to block the heterodimeric transporter associated with antigen processing (formed from TAP1 and TAP2). Western blotting with a polyclonal antibody to the C-terminus of TAP1 showed high expression of TAP1 in BeWo and JEG3, but not JAR cells, expression that was strongly upregulated by gamma-interferon. Gamma-interferon also upregulated the cell surface expression of HLA class I. TAP1 was strongly expressed in MC2 and MC3 extravillous cytotrophoblast cell lines immortalised with the SV40 large T antigen. The results suggest a role for non-classical HLA in the presentation of antigenic peptides to the immune system. PMID- 11254940 TI - The impact of chronic estrogen deprivation on immunologic parameters in the ovariectomized rhesus monkey (Macaca mulatta) model of menopause. AB - A large clinical literature suggests that estradiol (E(2)) plays a critical role in immune function. To further explore the relationship between E(2) and immune function, we examined a variety of immunological parameters in a rhesus monkey model of menopause and hormone replacement therapy. Rhesus monkeys (Age, 13.7+/ 2.6 years) were ovariectomized and received either sham (n=10) or estradiol (n=10) replacement implants. Nine months post-ovariectomy, a variety of immunologic parameters were measured. E(2)-deprivation reduced natural killer cell activity and increased serum soluble gp130 levels. There was a trend for an increased proportion of CD8(+) (P=0.12) and HLA-DR(+)CD3(+) cells (P=0.15) and decreased proportion of eosinophils (P=0.11) in the E(2)-deprived monkeys. There was no difference in leukocyte distribution, CD28, CD56, CD4, CD8/CD45, colony forming units-granulocyte/monocytes formation, peripheral blood mononuclear cell apoptotic rate, or serum TNF, TNF-R1, TNF-R2, IL-6, soluble IL-6R, and IL-1 between the groups. These data demonstrate that E(2)-deprivation affects several aspects of immune function. These findings may have implications for menopause associated changes of immune function that occur in women. PMID- 11254942 TI - Evaluation of the toxicity of chemical compounds using digestive acini of the bivalve mollusc Pecten maximus L. maintained alive in vitro. AB - The digestive gland of bivalve molluscs is a model of choice for experiments in ecotoxicology because of its implication in detoxification processes moreover of its classical functions in digestive phenomena. All physiological deteriorations of this organ, related or not to pollution, can lead to animal death. The recent development of a method allowing digestive acini of Pecten maximus to be maintained alive in vitro for 96 h opens up new research prospects in ecotoxicology. The action of contaminants considered to be cytotoxic or genotoxic in the literature were tested on this model. The results show the high cytotoxicity of ethylmethane sulphonate 80 and 5 mM after 2 h of contact with acini. Other compounds such as 4-nitroquinoline-N-oxide 0.1 mM, cadmium chloride 10(-5) M and atrazine 10(-4) M, which were weakly toxic after 2 h, became highly toxic after 48 h of contact. Compounds such as 4-nitroquinoline-N-oxide 1 mM, which were not cytotoxic after 2 h, proved to be the most genotoxic of all those tested. Others, such as MMS 1 mM, cadmium chloride 10(-4) and 10(-5) M, and atrazine 10(-5) M, showed an unconfirmed tendency to be genotoxic. The results obtained with this 'acinus' model seem more readily transposable to the whole organism than those obtained with 'isolated cell' models, in that acini can be considered as digestive glands 'in miniature'. PMID- 11254941 TI - Phenotypic and functional characterization of rhesus monkey decidual lymphocytes: rhesus decidual large granular lymphocytes express CD56 and have cytolytic activity. AB - In this study, we carried out a phenotypic and functional characterization of lymphocytes isolated from the uterine endometrium of the pregnant rhesus monkey. A majority (80%) of these cells were CD56(bright+), CD3- had typical large granular lymphocyte/uterine natural killer (NK) cell morphology and contained numerous cytoplasmic granules. Flow cytometric evaluation showed that rhesus decidual CD56(bright+) cells shared other phenotypic features of human uterine NK cells, including low levels of CD45RA and CD62L expression. A majority of the rhesus uterine CD56(bright+) cells expressed low levels of CD 16 but were CD2-. In contrast, most rhesus CD16+ peripheral blood cells were CD56-. In addition to the primary population of CD56(bright+) cells, a minor subset of smaller and less granular CD56(intermediate+) decidual lymphocytes was identified, the majority of which were CD16-, CD2(+). Decidual CD56+ cells did not express monocyte/macrophage markers, including CD14, CD64 and CD68. Decidual lymphocytes effectively lysed K562, Raji and particularly 721.221 targets in cytotoxicity assays. Together, these results suggest that as in human pregnancy, rhesus decidual CD56(bright+) cells represent a distinct lymphocyte subset that belongs to the NK cell lineage. PMID- 11254943 TI - Determination of acute mortality in adults and sublethal embryo responses of Palaemonetes pugio to endosulfan and methoprene exposure. AB - Adult grass shrimp (Palaemonetes pugio) were exposed to endosulfan or methoprene for 96 h and LC(50) values were calculated. Male and female P. pugio cohorts were also exposed to endosulfan for 96 h in an attempt to determine potential differences in sensitivity between the sexes. Results from the methoprene exposure indicated that this pesticide was not acutely toxic to adult grass shrimp at 1 mg l(-1). Due to the lack of sensitivity, sex specific tests with methoprene were not performed. The calculated LC(50) for a population of grass shrimp, including both males and females exposed to endosulfan, was 0.62 microg l(-1). The LC(50) determinations for the sex specific tests were 0.92 microg l( 1) for males and 1.99 microg l(-1) for females. Following these acute exposures, reproductively active grass shrimp were chronically exposed to 200 ng l(-1) endosulfan or 1 mg l(-1) methoprene and were allowed to produce embryos. The resulting embryos were assessed for potential sublethal toxicity. There were no observed differences in the percent successfully hatching or larval mortality 3 days post hatch among the treatments. However, endosulfan treated embryos had a significantly increased hatching time (9.76 days compared to 8.72 days in controls). Methoprene treated embryos also took longer to hatch (9.55 days), but this delay was not significantly different from controls. These findings suggest that endosulfan may preferentially affect male grass shrimp and exposed female grass shrimp may produce embryos with delayed hatching times. PMID- 11254944 TI - DNA integrity and total oxyradical scavenging capacity in the Mediterranean mussel, Mytilus galloprovincialis: a field study in a highly eutrophicated coastal lagoon. AB - In Mediterranean coastal lagoons, the combination of human impact and wide variability of natural environmental factors can lead to upsets in ecosystem homeostasis resulting in biodiversity decline. Oxidative damage has been causally linked to various kinds of environmental stress, both natural and artificial, the result being impairment of cellular functions. DNA damage and the efficiency of antioxidant defences in Mytilus galloprovincialis from the highly eutrophicated Orbetello Lagoon (Tuscany, Italy) were investigated, respectively by the single cell gel electrophoresis (or Comet test) and the total oxyradical scavenging capacity assay. Results showed significantly higher levels of DNA damage in mussels collected from the inner parts of the lagoon compared to specimens from more external sites. Specimens with the lower genetic integrity also exhibited a reduced efficiency in neutralizing three potent cellular oxidizing species, namely peroxyl radicals (ROO*), hydroxyl radicals (*OH) and peroxynitrite (HOONO), suggesting the involvement of reactive oxygen species in mediating the genetic damage. The analyzed biological parameters also showed a seasonal variability with a minimum of both DNA integrity and antioxidant scavenging efficiency during the warm months and an opposite trend in winter. The potential of analyzed techniques is discussed for the assessment of both anthropogenic and natural disturbance. PMID- 11254945 TI - Reproducibility of toxicity across mode of toxic action in the Tetrahymena population growth impairment assay. AB - Toxicity data collected in a laboratory setting are the primary source of potency information used for regulatory, modeling, or risk assessment purposes. However, the relative reproducibility of such toxicity data is rarely discussed. This study investigated the reproducibility of growth impairment data for the freshwater ciliate Tetrahymena pyriformis exposed to a structurally diverse group of chemicals of varying hydrophobicity within different modes of toxic action, either non-covalent narcosis or covalent electro(nucleo)philicity. The proportions of chemicals representing each mode of toxic action, or mechanism of action within each mode, were not chosen to emulate the occurrence of manufactured chemicals or chemicals within the TETRATOX database. Chemicals for which prior toxicity data existed were re-tested and reproducibility was evaluated. The toxic potency values of the selected chemicals were largely reproducible after re-testing of the toxic potency, as 98% of the chemicals had re-test toxicity values within one log unit of the original potency value. To further scrutinize the reproducibility of toxicity values, differences between values were investigated by mode of toxic action. A stringent criterion for reproducibility was enforced, which dictated that the re-tested toxicity value must be encompassed by the fiducial interval (FI) of the original toxicity value and vice versa for the chemical to be considered reproducible. Toxicity values of 28 of the 50 re-tested chemicals conformed to the criterion set for reproducible values. Of the nonreproducible chemicals, seven were narcotics: four nonpolar or neutral narcotics and three other narcotics (e.g. polar narcotics). However, four of these seven narcotics did have toxicity values encompassed by one FI, but not the other FI. The remaining chemicals that did not have reproducible potency measurements were electro(nucleo)philic in nature. Certain toxicophores were highly represented among these chemicals. These included quinone derivatives, electron releasing amino and hydroxyl moieties, and electron withdrawing nitro substituents, often in tandem with strong leaving groups (i.e. halogens), and unsaturated alcohols. Lack of reproducibility was common among the chemicals that elicited toxicity after either abiotic or biotic transformation. There was no clear trend between hydrophobicity and lack of reproducibility. While data are limited, these results suggest that toxic potency values of chemicals acting via the electro(nucleo)philic mode of toxic action could be more susceptible to non reproducibility. Ramifications of such lack of reproducibility could manifest in predictive toxicology models and their use in regulatory and risk assessment endeavors. PMID- 11254946 TI - Experimental exposure of juvenile chinook (Oncorhynchus tshawytscha) to bleached kraft mill effluent: hepatic CYP1A induction is correlated with DNA adducts but not with organochlorine residues. AB - Juvenile chinook salmon (Oncorhynchus tshawytscha) were exposed for 28 days to 0 16% treated effluent from an elemental chlorine free (ECF) bleached kraft pulp mill. Fresh effluent was diluted with river water drawn from upstream of the effluent diffuser. Fish were tested for biochemical responses to identify if the effluent would cause significant effects at concentrations spanning those present in the Fraser River, BC, Canada during winter conditions. Hepatic ethoxyresorufin O-deethylase (EROD) activity was increased significantly at all effluent concentrations and hepatic cytochrome P450 1A (CYP1A) protein was increased in all but 2% effluent. Hepatic DNA adduct concentrations were increased significantly at 8 and 16% effluent. These data indicate that there is a significant increase in all three responses at concentrations similar to those found in the receiving waters (4%) and that a dose-response relationship exists between BKME concentration and the responses measured. Carcasses contained low (< 0.2 pg g(-1)) concentrations of polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs). Polychlorinated biphenyls (PCBs) were at higher concentrations, accounting for 77% of the total contaminant burden, expressed as TEQ, but contaminants do not appear to have bioaccumulated in this study. It is likely that the PCB concentrations were due to contaminated commercial fish feed. Bile did not contain detectable levels of polycyclic aromatic hydrocarbons (PAHs) as measured by synchronous fluorescence spectrometry. PMID- 11254947 TI - Cadmium induces apoptosis and genotoxicity in rainbow trout hepatocytes through generation of reactive oxygene species. AB - Cadmium poses a serious environmental threat in aquatic ecosystems but the mechanisms of its toxicity remain unclear. The purpose of this work was first to determine whether cadmium induced apoptosis in trout hepatocytes, second to determine whether or not reactive oxygen species (ROS) were involved in cadmium induced apoptosis and genotoxicity. Hepatocytes exposed to increasing cadmium concentrations (in the range of 1-10 microM) showed a molecular hallmark of apoptosis which is the fragmentation of the nuclear DNA into oligonucleosomal length fragments, resulting from an activation of endogenous endonucleases and recognized as a 'DNA ladder' on conventional agarose gel electrophoresis. Exposure of hepatocytes to cadmium led clearly to the DEVD-dependent protease activation, acting upstream from the endonucleases and considered as central mediators of apoptosis. DNA strand breaks in cadmium-treated trout hepatocytes was assessed using the comet assay, a rapid and sensitive single-cell gel electrophoresis technique used to detect DNA primary damage in individual cells. Simultaneous treatment of trout hepatocytes with cadmium and the nitroxide radical TEMPO used as a ROS scavenger, reduced significantly DNA fragmentation, DEVD-related protease activity and DNA strand breaks formation. These results lead to a working hypothesis that cadmium-induced apoptosis and DNA strand breaks in trout hepatocytes are partially triggered by the generation of ROS. Additional studies are required for proposing a mechanistic model of cadmium-induced apoptosis and genotoxicity in trout liver cells, in underlying the balance between DNA damage and cellular defence systems in fish. PMID- 11254949 TI - Haemoproteus and Schistosoma synthesize heme polymers similar to Plasmodium hemozoin and beta-hematin. AB - Many parasites digest hemoglobin as an amino acid source, but only a few produce heme polymer pigment instead of catabolizing heme via heme oxygenase. This work compares purified heme polymers produced by Haemoproteus columbae and Schistosoma mansoni to that of Plasmodium falciparum hemozoin and synthetic beta-hematin. Fourier-transform infrared spectroscopy identifies the signature peaks of the common iron-carboxylate bond characteristic in all four heme polymers. However, all pigments could be distinguished by quite different three-dimensional structure visualized by Field Emission Inlens Scanning Electron Microscopy. Both P. falciparum and H. columbae heme polymers had a symmetrical shape unlike the amorphous S. mansoni heme polymer and beta-hematin. All four heme pigments serve as templates for heme polymer extension, which was inhibitable by chloroquine and other quinoline antimalarials. The polymers showed different levels of resistance to hydrogen peroxide degradation. This work identifies another genus, Haemoproteus, capable of intracellular heme polymer formation. The different three-dimensional structures of each pigment implicate genus specific formation of heme polymer, variation of inhibition of polymer extension by the quinolines and degradation by hydrogen peroxide. PMID- 11254951 TI - Serial analysis of gene expression (SAGE) in Plasmodium falciparum: application of the technique to A-T rich genomes. AB - The advent of high-throughput methods for the analysis of global gene expression, together with the Malaria Genome Project open up new opportunities for furthering our understanding of the fundamental biology and virulence of the malaria parasite. Serial analysis of gene expression (SAGE) is particularly well suited for malarial systems, as the genomes of Plasmodium species remain to be fully annotated. By simultaneously and quantitatively analyzing mRNA transcript profiles from a given cell population, SAGE allows for the discovery of new genes. In this study, one reports the successful application of SAGE in Plasmodium falciparum, 3D7 strain parasites, from which a preliminary library of 6880 tags corresponding to 4146 different genes was generated. It was demonstrated that P. falciparum is amenable to this technique, despite the remarkably high A-T content of its genome. SAGE tags as short as 10 nucleotides were sufficient to uniquely identify parasite transcripts from both nuclear and mitochondrial genomes. Moreover, the skewed A-T content of parasite sequence did not preclude the use of enzymes that are crucial for generating representative SAGE libraries. Finally, a few modifications to DNA extraction and cloning steps of the SAGE protocol proved useful for circumventing specific problems presented by A-T rich genomes. PMID- 11254950 TI - Sialic acid-dependent binding of baculovirus-expressed recombinant antigens from Plasmodium falciparum EBA-175 to Glycophorin A. AB - The Plasmodium falciparum Erythrocyte Binding Antigen-175, EBA-175, is a soluble merozoite stage parasite protein which binds to glycophorin A surface receptors on human erythrocytes. We have expressed two conserved cysteine-rich regions, region II and region VI, of this protein as soluble His-tagged polypeptides in insect cell culture, and have tested their function in erythrocyte and glycophorin A binding assays. Recombinant region II polypeptides comprised of the F2 sub-domain or the entire region II (F1 and F2 sub-domains together) bound to erythrocytes and to purified glycophorin A in a manner similar to the binding of native P. falciparum EBA-175 to human red cells. Removal of sialic acid residues from the red cell surface totally abolished recombinant region II binding, while trypsin treatment of the erythrocyte surface reduced but did not eliminate recombinant region II binding. Synthetic peptides from three discontinuous regions of the F2 sub-domain of region II inhibited human erythrocyte cell binding and glycophorin A receptor recognition. Immune sera raised against EBA 175 recombinant proteins recognized native P. falciparum-derived EBA-175, and sera from malaria-immune adults recognized recombinant antigens attesting to both the antigenicity and immunogenicity of proteins. These results suggest that the functionally-active recombinant region II domain of EBA-175 may be an attractive candidate for inclusion in multi-component asexual blood stage vaccines. PMID- 11254952 TI - Identification of Leishmania major cysteine proteinases as targets of the immune response in humans. AB - In this study, we report the identification of two parasite polypeptides recognized by human sera of patients infected with Leishmania major. Isolation and sequencing of the two genes encoding these polypeptides revealed that one of the genes is similar to the L. major cathepsin L-like gene family CPB, whereas the other gene codes for the L. major homologue of the cysteine proteinase a (CPA) of L. mexicana. By restriction enzyme digestion of genomic DNA, we show that the CPB gene is present in multiple copies in contrast to the cysteine proteinase CPA gene which could be unique. Specific antibodies directed against the mature regions of both types expressed in Escherichia coli were used to analyze the expression of these polypeptides in different stages of the parasite's life cycle. Polypeptides of 27 and 40 kDa in size, corresponding to CPA and CPB respectively, were detected at higher level in amastigotes than in stationary phase promastigotes. Purified recombinant CPs were also used to examine the presence of specific antibodies in sera from either recovered or active cases of cutaneous leishmaniasis patients. Unlike sera from healthy uninfected controls, all the sera reacted with recombinant CPA and CPB. This finding indicates that individuals having recovered from cutaneous leishmaniasis or with clinically apparent disease have humoral responses to cysteine proteinases demonstrating the importance of these proteinases as targets of the immune response and also their potential use for serodiagnosis. PMID- 11254953 TI - Targeting of soluble proteins to the rhoptries and micronemes in Toxoplasma gondii. AB - Rhoptry and microneme organelles of the protozoan parasite Toxoplasma gondii are closely associated with host cell adhesion/invasion and establishment of the intracellular parasitophorous vacuole. In order to study the targeting of proteins to these specialized secretory organelles, we have engineered green fluorescent protein (GFP) fusions to the rhoptry protein ROP1 and the microneme protein MIC3. Both chimeras are correctly targeted to the appropriate organelles, permitting deletion analysis to map protein subdomains critical for targeting. The propeptide and a central 146 amino acid region of ROP1 are sufficient to target GFP to the rhoptries. More extensive deletions result in a loss of rhoptry targeting; the GFP reporter is diverted into the parasitophorous vacuole via dense granules. Certain MIC3 deletion mutants were also secreted into the parasitophorous vacuole via dense granules, supporting the view that this route constitutes the default pathway in T. gondii, and that specific signals are required for sorting to rhoptries and micronemes. Deletions within the cysteine rich central region of MIC3 cause this protein to be arrested at various locations within the secretory pathway, presumably due to improper folding. Although correctly targeted to the appropriate organelles in living parasites, ROP1-GFP and MIC3-GFP fusion proteins were not secreted during invasion. GFP fusion proteins were readily secreted from dense granules, however, suggesting that protein secretion from rhoptries and micronemes might involve more than a simple release of organellar contents. PMID- 11254954 TI - In-vitro competition analysis of procyclin gene and variant surface glycoprotein gene expression site transcription in Trypanosoma brucei. AB - In Trypanosoma brucei, alpha-amanitin-resistant transcription characteristic of RNA polymerase I is initiated at ribosomal RNA gene (RRNA), procyclin gene (GPEET or EP1), and variant surface glycoprotein gene expression site (VSG ES) promoters. The three promoter types do not share obvious sequence homologies, but contain a proximal domain I and a distal domain II within 80 bp upstream of the transcription initiation site. RRNA, GPEET and EP1, but not the VSG ES promoter, require additional upstream sequences for full activity. In the present study, we competed in-vitro transcription of circular template DNA with linear DNA fragments to identify promoter domains responsible for binding and sequestering essential trans-acting transcription factors. For the GPEET promoter, we found that domain III, located between positions -141 and -92, was most important for the DNA fragment to exert a transcription competition effect, whereas domain I, the only element absolutely required for transcription, was not. Moreover, insertions between promoter domains II and III reduced both transcription from the GPEET promoter and competition with the GPEET promoter fragment, suggesting that these two domains cooperate in the formation of a stable DNA-protein complex. Taken together, these results indicate a promoter structure very similar to that of the Saccharomyces cerevisiae RRNA promoter. In contrast, VSG ES promoter analysis showed that domains I and II are both necessary and sufficient to compete transcription. Despite this structural difference, our analysis provide evidence that GPEET and VSG ES promoters interact with a common factor that is also important for RRNA promoter transcription. PMID- 11254955 TI - Conservation of metacyclic variant surface glycoprotein expression sites among different trypanosome isolates. AB - We identified in a Trypanosoma brucei brucei strain (AnTat 1) an expression site for a metacyclic variant surface glycoprotein (MVSG) gene (MVSG) that was previously characterized in a T. b. rhodesiense strain (WRATat 1.1). The 3.4 kb sequences of the two expression sites are 99.6% identical, with no differences in the sequence of the 1.5 kb MVSG. Two other MVSGs in the WRATat 1.1 genome are not present in the AnTat 1 genome. In addition, five other T. b. brucei and T. b. rhodesiense strains, isolated in the same geographic region as the two former strains, do not contain any of these three MVSGs. Two of these five strains, however, appear to possess a very similar MVSG expression site, but with different MVSGs in it. Thus, the presence of the same MVSG in the same expression site in two different isolates is unusual and may be the result of genetic exchange in the field between T. b. brucei and T. b. rhodesiense isolates. Analysis of other African trypanosome strains for the presence of the three WRATat 1.1 MVSG expression sites demonstrated that the expression sites' promoter sequences are much more likely to be present than are specific MVSGs, suggesting that loss of MVSGs is the result of replacement by other VSGs. The promoter region of the MVSG expression site active in the WRATat 1.1 MVAT7 variant was found to be highly conserved among T. b. brucei, T. b. rhodesiense and T. b. gambiense group 2 isolates, whereas it does not occur in the T. b. gambiense group 1 isolates tested. A phylogenetic analysis of this promoter region sequence shows that the T. b. gambiense group 2 isolates form a monophyletic clade well separated from the T. b. brucei/T. b. rhodesiense isolates. Thus, whilst the T. b. brucei, T. b. rhodesiense and T. b. gambiense group 2 isolates are closely related but heterogenous, molecular tools may be developed to distinguish T. b. gambiense group 2 isolates from the others. PMID- 11254956 TI - Diversity and dynamics of the minichromosomal karyotype in Trypanosoma brucei. AB - The genome of African trypanosomes contains a large number of minichromosomes. Their only proposed role is in the expansion of the parasites' repertoire of telomeric variant surface glycoprotein (VSG) genes as minichromosomes carry silent VSG gene copies in telomeric locations. Despite their importance as VSG gene donors, little is known about the actual composition of the minichromosomal karyotype and the stability of its inheritance. In this study we show, by using high-resolution pulsed-field electrophoresis, that a non-clonal trypanosome population contains an extremely diverse pattern of minichromosomes, which can be resolved into less complex clone-specific karyotypes by non-selective cloning. We show that the minichromosome patterns of such clones are stable over at least 360 generations. Furthermore, using DNA markers for specific minichromosomes, we demonstrate the mitotic stability of these minichromosomes within the population over a period of more than 5 years. Length variation is observed for an individual minichromosome and is most likely caused by a continuous telomeric growth of approximately 6 bp per telomere per cell division. This steady telomeric growth, counteracted by stochastic large losses of telomeric sequences is the most likely cause of minichromosome karyotype heterogeneity within a population. PMID- 11254957 TI - Bacteria expressing single-chain immunotoxin inhibit malaria parasite development in mosquitoes. AB - Single-chain immunotoxins are ideal tools to selectively kill infectious agents. In applying this technology to block transmission of malaria parasites in the mosquito vector, we have constructed a single-chain immunotoxin composed of a single-chain antibody fragment (scFv) directed to Pbs2l on the surface of Plasmodium berghei ookinetes linked to a lytic peptide, Shiva-1. The single-chain immunotoxin was expressed in Escherichia coli, and the protein was purified by a Ni-NTA column. The single-chain immunotoxin was initially shown to exhibit greater killing properties for P. berghei ookinetes in vitro compared with the scFv or synthetic Shiva-1 peptide alone. In an attempt to block malaria transmission by genetically engineered bacteria, recombinant E. coli harboring the single-chain immunotoxin gene were introduced into the mosquito midgut by membrane feeding. The number of infected mosquitoes and their oocyst densities were significantly reduced when the mosquitoes were subsequently allowed to feed on P. berghei-infected mice. These results indicate not only that a single-chain immunotoxin with enhanced parasiticidal activity could form a basis for the development of more effective malaria therapeutic agents, but also that introduction of genetically engineered bacteria into anopheline mosquitoes may offer a practical approach to the regulation of malaria transmission. PMID- 11254958 TI - Characterization of the Trypanosoma cruzi Cdc2p-related protein kinase 1 and identification of three novel associating cyclins. AB - Several Cdc2p-related protein kinases (CRKs) have been described in trypanosomatids but their role in the control of the cell cycle nor their biological functions have been addressed. In Trypanosoma cruzi two CRKs have been identified, TzCRK1 and TzCRK3. In this work we further characterize T. cruzi CRK1 and report the identification of three novel associating cyclins. We demonstrate that CRK1 levels and localization do not vary during the cell cycle, and show that it is localized in the cytoplasm, discrete regions of the nucleus, and is highly concentrated in the mitochondrion DNA (kinetoplast), suggesting a putative control function in this organelle. Using purified anti-CRK1 IgGs, we immunoprecipitated from the soluble fraction of T. cruzi epimastigote forms a protein kinase activity which is not inhibited by CDK inhibitors. In addition, we co-precipitated with p13Suc1p beads a kinase activity that was inhibited by the CDK inhibitor flavopiridol and olomoucine. Lastly, using the yeast two-hybrid system we identified three novel cyclin-like proteins able to associate with TzCRK1, and demonstrate that two of these cyclins also bind the T. cruzi CRK3 protein, indicating that these two CRKs are cyclin-dependent kinases. PMID- 11254959 TI - Characterization of a candidate Trypanosoma brucei U1 small nuclear RNA gene. AB - We have previously shown that the poly(A) polymerase (PAP) gene of Trypanosoma brucei is interrupted by an intervening sequence. It was postulated that removing this intron by cis-splicing requires a yet unidentified U1 small nuclear RNA (snRNA), which in other organisms engages in base-pair interactions across the 5' splice site during early spliceosome assembly. Here we present a characterization of a 75 nucleotide long candidate T. brucei U1 snRNA. Immunoprecipitation studies indicate that a trimethylguanosine cap structure is present at the 5' end and that the RNA is bound to core proteins common to spliceosomal ribonucleoprotein particles. The U1 snRNA has the potential for extensive intermolecular base pairing with the PAP 5' splice site. We used block replacement mutagenesis to identify sequences necessary for in vivo expression of U1 snRNA. We found that at least two cis-acting elements, tRNA-like A and B boxes, located in the 5' flanking region are necessary for U1 snRNA synthesis; no internal sequences close to the transcription start site are essential, suggesting a promoter architecture distinct from other trypanosome U-snRNA genes. PMID- 11254960 TI - The immunodominant 17-kDa antigen from Cryptosporidium parvum is glycosylphosphatidylinositol-anchored. AB - Cryptosporidium parvum is a protozoan parasite of the intestinal epithelium that has caused numerous outbreaks of diarrheal illness in humans. During our studies of the host immune response to C. parvum infection, we noted that two of the immunodominant surface antigens of the sporozoite stage of the parasite readily extract into Triton X-114. We recently cloned the immunodominant 17-kDa surface antigen and suggested that the carboxy-terminal peptide sequence may satisfy the requirements for GPI anchor addition. In the work presented here, we were able to show that the 17-kDa antigen could be metabolically labeled in vitro with tritiated ethanolamine and that the antigen contained myo-inositol. The antigen was cleaved by GPI-PLD but not by PI-PLC and it could be converted to a water soluble form by chemical deglycosylation. We suggest that the 17-kDa antigen is indeed GPI anchored and that the anchor contains an acylated inositol and either a lyso-acyl- or a diacyl-glycerol. We are currently working to determine what role the anchor may play in the human immune response to this antigen. PMID- 11254961 TI - A receptor-like flagellar pocket glycoprotein specific to Trypanosoma brucei gambiense. AB - Trypanosoma brucei gambiense and T. b. rhodesiense are protozoan parasites causing sleeping sickness in humans due to their resistance to lysis by normal human serum (NHS). Based on the observation that the resistance gene of T. b. rhodesiense encodes a truncated form of the variant specific glycoprotein (VSG), we cloned a similar gene in T. b. gambiense using reverse transcription-linked polymerase chain reaction with VSG-specific primers. This gene, termed TgsGP for T. gambiense-specific glycoprotein, was found to be specific to T. b. gambiense. It is located close to a telomere and is transcribed by a pol II RNA polymerase, only at the bloodstream stage of the parasite development. TgsGP encodes a 47-kDa protein consisting of a N-terminal VSG domain presumably provided with a glycosylphosphatidylinositol (GPI) anchor sequence, similar to the pESAG6 subunit of the trypanosomal transferrin receptor. TgsGP is located in the flagellar pocket, and contains the linear N-linked polyacetyllactosamine characteristic of the endocytotic machinery of T. brucei. These observations strongly suggest that TgsGP is a T. b. gambiense specific receptor. Since stable expression of this protein in T. b. brucei did not confer resistance to NHS, TgsGP may either need another factor to achieve this purpose or fulfils another function linked to adaptation of the parasite to man. PMID- 11254962 TI - Mutagenesis of dihydrofolate reductase from Plasmodium falciparum: analysis in Saccharomyces cerevisiae of triple mutant alleles resistant to pyrimethamine or WR99210. AB - Inhibitors of dihydrofolate reductase (DHFR) have been a mainstay of chemotherapy of falciparum malaria for >50 years. Unfortunately, point mutations in DHFR are the major cause of resistance to drugs of this class and mutations have rapidly diminished the clinical effectiveness of these drugs. We designed a simple yeast based system to produce and analyze point mutations in the Plasmodium falciparum DHFR domain of the DHFR-thymidylate synthase gene that confers resistance to pyrimethamine (PM), the major antifolate currently used in malaria treatment, or to WR99210, an experimental antifolate. We used PCR mutagenesis, screened >1000 DHFR alleles that encoded functional enzymes and studied approximately 100 that were more resistant than a naturally occurring resistant allele (N51I and S108N). The IC(50) values for both drugs were determined for a subset of 44 alleles that carried only a single new mutation. Mutations that increased resistance to PM 10 100 fold (to >10(-4) M) were identified in three regions of the DHFR domain - around amino acids 50, 188 and 213. In contrast, mutations that caused WR resistance were far less common and only conferred approximately 10-fold resistance (to approximately 10(-7) M). Even more interesting, only the mutations at 188 increased resistance to WR and mutations in the 213 and other regions either had no effect or actually increased sensitivity to WR. This collateral hypersensitivity raises the possibility that opposing selection for resistance/sensitivity to PM and WR might be used to slow selection of populations of P. falciparum resistant to antifolate treatment. PMID- 11254964 TI - Initiator and upstream elements in the alpha2-tubulin promoter of Giardia lamblia. AB - Giardia lamblia, one of the earliest diverging eukaryotes and a major cause of diarrhea world-wide, has unusually short intergenic regions, raising questions concerning its regulation of gene expression. We have approached this issue through examination of the alpha2-tubulin promoter and in particular investigated the function of an AT-rich element surrounding the transcription start site. Its placement and the ability of this sequence to direct transcription initiation in the absence of any other promoter elements is similar to the initiator element in higher eukaryotes. However, the sequence diversity of extremely short (8-10 bp) initiator elements is surprising, as is their ability to independently direct substantial levels of transcription. We also identified a large AT-rich element located between -64 and -29 bp upstream of the transcriptional start site and show using both deletions and site-specific mutations of this region that sequences between -60 and the start of transcription are important for promoter strength; interestingly this AT-rich sequence is not highly conserved among different Giardia promoters. These data suggest that while the overall structure of the core promoter has been conserved throughout eukaryotic evolution, significant variation and flexibility is allowed in element consensus sequences and roles in transcription. In particular, the short and diverse sequences that function in transcription initiation in Giardia suggest the potential for relaxed transcriptional regulation. PMID- 11254963 TI - Complementation of Plasmodium berghei TRAP knockout parasites using human dihydrofolate reductase gene as a selectable marker. AB - Previously we have used the Plasmodium dihydrofolate reductase thymidylate synthase (DHFR-TS) selectable marker to generate Plasmodium berghei TRAP null mutant parasites. These TRAP null mutants do not glide and they showed a great reduction in their ability to infect mosquito salivary glands and the hepatocytes of the vertebrate host. Thus far, complementation of these knockout parasites was not possible due to the lack of additional selectable markers. Recently, a new selectable marker, based on the human dihydrofolate reductase (hDHFR) gene, has been developed which confers resistance to the antifolate drug WR99210. This drug has been found to be highly active against pyrimethamine-sensitive and -resistant strains of P. berghei. In this study, we have used the hDHFR gene as a second selectable marker for the complementation of P. berghei TRAP null mutant parasites. Restoration of the TRAP null mutant parasites to the wild-type phenotype was achieved in this study via autonomously replicating episomes bearing a wild-type copy of the TRAP gene. This is the first report of complementation of a mutant phenotype in malaria parasites. PMID- 11254965 TI - In vivo epitope tagging of Trypanosoma brucei genes using a one step PCR-based strategy. PMID- 11254966 TI - Uridine insertion/deletion RNA editing in Leishmania tarentolae mitochondria shows cell cycle dependence. PMID- 11254967 TI - Deletion analysis of the 5' flanking sequence of the Plasmodium gallinaceum sexual stage specific gene pgs28 suggests a bipartite arrangement of cis-control elements. PMID- 11254968 TI - Bacterial replication initiator DnaA. Rules for DnaA binding and roles of DnaA in origin unwinding and helicase loading. AB - We review the processes leading to the structural modifications required for the initiation of replication in Escherichia coli, the conversion of the initial complex to the open complex, loading of helicase, and the assembly of two replication forks. Rules for the binding of DnaA to its binding sites are derived, and the properties of ATP-DnaA are described. We provide new data on cooperative interaction and dimerization of DnaA proteins of E. coli, Streptomyces and Thermus thermophilus, and on the stoichiometry of DnaA-oriC complexes of E. coli. PMID- 11254969 TI - Multiple pathways regulating DnaA function in Escherichia coli: distinct roles for DnaA titration by the datA locus and the regulatory inactivation of DnaA. AB - Escherichia coli DnaA protein forms a multimeric complex at the chromosomal origin of replication (oriC), where a series of initiation reactions occurs and DNA polymerase III holoenzyme is loaded. The ATP-bound form of DnaA, which is active for initiation, is converted to the inactive ADP-bound form through interaction with the sliding clamp, the beta subunit of DNA polymerase III holoenzyme loaded on DNA. This negative regulation, termed RIDA, is required for preventing untimely initiations. Here, we asked if RIDA is functionally related to another negative regulation, DnaA titration by the datA site. The datA site can harbor hundreds of DnaA molecules, and is also required for preventing untimely initiations. We reveal here that, in growing cells of the datA(+) and datA-deleted strains, the ATP-DnaA levels were both maintained in a limited range of about 20-30% of the total ATP- plus ADP-DnaA molecules. This indicates that RIDA functions in the absence of datA. In synchronized datA-deleted cells, the ATP-DnaA level fluctuated in a manner similar to that observed in datA(+) cells. This suggests that RIDA operates independent from DnaA titration to datA. We suggest that these two mechanisms may play complementary roles during the cell cycle to prevent untimely initiations and thus ensure the scheduled initiation. PMID- 11254970 TI - Escherichia coli DnaA protein--phospholipid interactions: in vitro and in vivo. AB - DNA replication in Escherichia coli is controlled at the initiation stage, possibly by regulation of the essential activity of DnaA protein. The cellular membrane has long been hypothesized to be involved in chromosomal replication. Accumulating evidence, both in vitro and in vivo, that supports the importance of membrane phospholipids influencing the initiation activity of DnaA is reviewed. PMID- 11254971 TI - Defective initiation in an Escherichia coli dnaA(Cs,Sx) mutant. AB - Mutations in the Escherichia coli gene for initiation of DNA replication, dnaA, which suppress the polymerization defect and growth inhibition caused by temperature-sensitive (Ts) mutations in the polymerization gene, dnaX, are called Cs,Sx. We show here that these mutations, on their own, also cause defects in initiation, including inhibition of initiation at both low (20 degrees C) and high (44 degrees C) temperatures and asynchronous initiation at both the permissive (34 degrees C) and suppression (39 degrees C) temperatures. These findings suggests a relationship between partially defective initiation and suppression of the polymerization defect, both of which occur at 39 degrees C. PMID- 11254972 TI - DnaA protein dependent denaturation of negative supercoiled oriC DNA minicircles. AB - The DnaA protein binds specifically and tightly to oriC supercoiled 641 bp minicircle DNA. The binding of the initiator promoted a partial unwinding of the superhelical oriC minicircle (Mc-oriC). Open complexes are detected either by a change in the linking number or by the sensitivity to the attack of a single strand specific Bal 31 nuclease. The open complex is found only in the presence of the DnaA protein. PMID- 11254973 TI - Partitioning of the Escherichia coli chromosome: superhelicity and condensation. AB - Segregation in Escherichia coli, the process of separating the replicated chromosomes into daughter progeny cells, seems to start long before the duplication of the genome reaches completion. Soon after initiation in mid-cell region, the daughter oriCs rapidly move apart to fixed positions inside the cell (quarter length positions from each pole) and are anchored there by yet unknown mechanism(s). As replication proceeds, the rest of the chromosome is sequentially unwound and then refolded. At termination, the two sister chromosomes are unlinked by decatenation and separated by supercoiling and/or condensation. Muk and Seq proteins are involved in different stages of this replication-cum partition process and thus can be categorized as important partition proteins along with topoisomerases. E. coli strains, lacking mukB or seqA functions, are defective in segregation and cell division. The nucleoids in these mutant strains exhibit altered condensation and superhelicity as can be demonstrated by sedimentation analysis and by fluorescence microscopy. As the supercoiling of an extrachromosomal element (a plasmid DNA) was also influenced by the mukB and seqA mutations we concluded that the MukB and SeqA proteins are possibly involved in maintaining the general supercoiling activity in the cell. The segregation of E. coli chromosome might therefore be predominantly driven by factors that operate by affecting the superhelicity and condensation of the nucleoid (MukB, SeqA, topoisomerases and additional unknown proteins). A picture thus emerges in which replication and partition are no longer compartmentalized into separable stages with clear gaps (S and M phases in eukaryotes) but are parallel processes that proceed concomitantly through a cell cycle continuum. PMID- 11254975 TI - SMC proteins in bacteria: condensation motors for chromosome segregation? AB - SMC proteins are a ubiquitous protein family, present in almost all organisms so far analysed except for a few bacteria. They function in chromosome condensation, segregation, cohesion, and DNA recombination repair in eukaryotes, and can introduce positive writhe into DNA in vitro. SMC proteins and the structurally homologous MukB protein are unusual ATPases that form antiparallel dimers, with long coiled coil segments separating globular ends capable of binding DNA. Recently, SMC proteins have been shown to be essential for chromosome condensation, segregation and cell cycle progression in bacteria. Identification of a suppressor mutation for MukB in topoisomerase I in Escherichia coli suggests that SMC proteins are involved in negative DNA supercoiling in vivo, and by this means organize and compact chromosomes. A model is discussed in which bacterial SMC proteins act after an initial separation of replicated chromosome origins into the future daughter cell, separating sister chromatids by condensing replicated DNA strands within both cell halves. This would be analogous to a pulling of DNA strands into opposite cell halves by a condensation mechanism exerted at two specialised subregions in the cell. PMID- 11254974 TI - The Escherichia coli SeqA protein binds specifically to two sites in fully and hemimethylated oriC and has the capacity to inhibit DNA replication and affect chromosome topology. AB - The SeqA protein was identified as a factor that prevents reinitiation of newly replicated, hemimethylated origins. SeqA also seems to inhibit initiation of fully methylated origins, thus contributing to the regulation of chromosomal replication. The SeqA protein was found to bind to two sites in the left part of the origin, near the AT-rich region where strand separation takes place during initiation of replication. The same binding sites seemed to be preferred irrespective of whether the origin was in the newly replicated (hemimethylated) state or not. In addition to binding specifically to groups of GATC sites, the SeqA protein was capable of interacting non-specifically with negatively supercoiled DNA, restraining the supercoils in a fashion similar to that seen with histone-like protein HU. The restraint of supercoils by SeqA was, in contrast to that of HU, cooperative. PMID- 11254976 TI - Does the parallel evolution pattern between the replication-segregation proteins and HU have a biological significance? AB - The bacterial chromosome is a highly compacted nucleoproteic structure. Its apparent disordered morphology is difficult to conciliate with newly discovered mechanisms governing the propagation of genetic information between mother and daughter cells. Recent experiments in bacterial genetics, biochemistry and cytology from a number of laboratories are beginning to unravel how at each cell division, DNA replication and segregation proteins interact spatially with specific DNA motifs to orchestrate replication and movement of replication forks and chromosomes. We propose here a method to confirm and perhaps extend these experiments by in silico protein sequence comparisons and phylogeny. This analysis showed a parallel evolution between the histone-like protein HU and key protein factors involved in DNA replication and chromosome segregation. PMID- 11254977 TI - Experiments on movement of DNA regions in Escherichia coli evaluated by computer simulation. AB - During the cell cycle of Escherichia coli DNA is replicated and segregated over two prospective daughter cells. Nucleoids as a whole separate gradually in line with cell elongation, but sub-nucleoid DNA regions may behave differently, separating non-gradually. We tested the ability of three models to predict the outcome of a fluorescent in situ hybridisation (FISH) experiment. We did this by comparing computer-simulated data with experimental data. The first model predicts gradual separation in line with cell elongation. The second model predicts that origins stick together for some time after duplication before one copy jumps to the other side of the cell (non-gradual separation). The simulated data of these models are very similar, indicating that FISH is not a suitable method to distinguish between these two models. The third model predicts that origins may be anywhere within the nucleoid(s). We found that simulated data using the third model resemble the experimental data most. However, DNA regions are not randomly localised in the cell, although their localisation is fuzzy. We propose that movement of DNA regions is the result of a combination of factors. Nucleoid segregation (or the forces behind it) dictates the overall direction of movement. Other factors, of which we show that diffusion could be an important one, move DNA in other directions giving rise to non-gradual movement in individual cells and contributing to variation in intracellular position per cell length in a population of cells. PMID- 11254978 TI - Coordinating DNA replication with cell division: lessons from outgrowing spores. AB - Progress in solving the long-standing puzzle of how a cell coordinates chromosome replication with cell division is significantly aided by the use of synchronous cell populations. Currently three systems are employed for obtaining such populations: the Escherichia coli 'baby machine', the developmentally-controlled cell cycle of Caulobacter crescentus, and Bacillus subtilis germinated and outgrowing spores. This review examines our current understanding of the relationship between replication and division and how the use of B. subtilis outgrowing spores and, more recently its combination with immunofluorescence microscopy, has contributed significantly to this important area of biology. About 20 years ago, and also more recently, this system was used to show convincingly that termination of DNA replication is not essential for a central septum to form, raising the possibility that the early stages of division occur well before termination. It has also been demonstrated that there is no major synthesis of the division initiation proteins, FtsZ and DivIB, linked to initiation, progression or completion of the first round of chromosome replication accompanying spore outgrowth. This has led to the suggestion that the primary link between chromosome replication and cell division at midcell is not likely to occur through a control over the levels of these proteins. Very recent work has employed a combination of the use of B. subtilis outgrowing spores with immunofluorescence microscopy to investigate the relationship between midcell Z ring assembly and the round of chromosome replication linked to it. The results of this work suggest a role for initiation and progression into the round of replication in blocking midcell Z ring formation until the round is complete or almost complete, thereby ensuring that cell division occurs between two equally partitioned chromosomes. PMID- 11254979 TI - Cell-cycle research with synchronous cultures: an evaluation. AB - The baby-machine system, which produces new-born Escherichia coli cells from cultures immobilized on a membrane, was developed many years ago in an attempt to attain optimal synchrony with minimal disturbance of steady-state growth. In the present article, we put forward a model to describe the behaviour of cells produced by this method, and provide quantitative evaluation of the parameters involved, at each of four different growth rates. Considering the high level of selection achievable with this technique and the natural dispersion in interdivision times, we believe that the output of the baby machine is probably close to optimal in terms of both quality and persistence of synchrony. We show that considerable information on events in the cell cycle can be obtained from populations with age distributions very much broader than those achieved with the baby machine and differing only modestly from steady state. The data presented here, together with the long and fruitful history of findings employing the baby machine technique, suggest that minimisation of stress on cells is the single most important factor for successful cell-cycle analysis. PMID- 11254980 TI - Hypothesis: membrane domains and hyperstructures control bacterial division. AB - The mechanism responsible for creating the division site in the right place at the right time in bacteria is unknown. It has been attributed to the formation of proteolipid domains in the cytoplasmic membrane surrounding the nucleoids. We interpret the growing evidence for this hypothesis by invoking hyperstructures, which exist at a level of organization intermediate between macromolecules and genes. Non-equilibrium hyperstructures comprise the genes, mRNA proteins and lipids required for a particular function such as cell division, and assemble and disassemble according to the needs of the cell. PMID- 11254981 TI - Approaching the physiological functions of penicillin-binding proteins in Escherichia coli. AB - A rigid shell of peptidoglycan encases and shapes bacteria and is constructed and maintained by a diverse set of enzymes, among which are the penicillin-binding proteins (PBPs). Although a great deal has been learned about how these proteins synthesize and modify peptidoglycan, the physiological functions of the multitude of bacterial PBPs remain enigmatic. We approached this problem by combining PBP mutations in a comprehensive manner and screening for effects on biochemical processes involving the passage of proteins or nucleic acids across the cell wall. The results indicate that the PBPs or their peptidoglycan product do have significant biological functions, including roles in determination of cell shape, in phage resistance, in induction of capsule synthesis, and in regulation of autolysis. PMID- 11254982 TI - Enzymology of elongation and constriction of the murein sacculus of Escherichia coli. AB - Multiple deletions in murein hydrolases revealed that predominantly amidases are responsible for cleavage of the septum during cell division. Endopeptidases and lytic transglycosylases seem also be involved. In the absence of these enzymes E. coli grows normally but forms chains of adhering cells. Surprisingly, mutants lacking up to eight different murein hydrolases still grow with almost unaffected growth rate. Therefore it is speculated that general enlargement of the murein sacculus may differ from cell division by using transferases rather than the two sets of hydrolytic and synthetic enzymes as seems to be the case for the constriction process. A model is presented that describes growth of the murein of both Gram-positive and -negative bacteria by the activity of murein transferases. It is speculated that enzymes exist that catalyze a transpeptidation of the pre existing murein onto murein precursors or nascent murein by using the chemical energy present in peptide cross-bridges. Such enzymes would at the same time cleave bonds in the murein net and insert new material into the growing sacculus. PMID- 11254983 TI - Transcription of the Escherichia coli dcw cluster: evidence for distal upstream transcripts being involved in the expression of the downstream ftsZ gene. AB - Escherichia coli strains VIP596 and VIP597 have been constructed to compare the amount of transcription of the ftsZ gene derived from proximal promoters in the ddlB-ftsZ region with that originating in the upstream regions of the dcw cluster. Both strains have in common a beta-galactosidase reporter fusion located at the ddlB locus, but differ in that VIP597 has a transcription terminator Omega interposon located downstream from lacZ. In addition, these strains have the ddlB, ftsQ, ftsA and ftsZ genes under the control of the IPTG-inducible promoter P(tac), allowing to control artificially ftsZ expression for normal cell division to take place. When beta-galactosidase activity was measured in VIP596 and VIP597 and compared to the levels measured in strain VIP407, in which the lacZ reporter fusion is located in the ftsZ gene, they were found to account for nearly 66% of the total transcription entering into ftsZ. This result indicates that the reduction in ftsZ transcription observed when the promoters in the ddlB-ftsA region are disconnected from the upstream sequences of the dcw cluster (as observed by Flardh et al., Mol. Microbiol. 30 (1998) 305-316) in strain VIP490) is the direct consequence of the interruption in the transcription originated upstream and not due to the effect of such sequences on the promoters proximal to ftsZ. PMID- 11254984 TI - Conserved sequence motif at the C-terminus of the bacterial cell-division protein FtsA. AB - FtsA is an essential part of the septal ring structure in bacterial cell division. Two peptide-protein interactions are known in this process: FtsA and ZipA bind the C-terminus of FtsZ, the bacterial tubulin homologue, which is the first septal component to appear at the septum. Our recent crystal structure of FtsA revealed a possible peptide binding site on FtsA and a long disordered C terminal region. Here we show that all FtsA proteins contain a conserved 10-13 residue motif at the C-terminal end that may facilitate targeting of downstream septal components. PMID- 11254985 TI - Perpendicular planes of FtsZ arcs in spheroidal Escherichia coli cells. AB - Division planes in Escherichia coli, usually restricted to one dimension of the rod-shaped cell, were induced at all possible planes by transforming the cells to spheroids with mecillinam (inactivating PbpA). Such cells displayed many nucleoids and arcs of FtsZ, genetically tagged to green fluorescent protein, that developed to rings at constriction sites all around their surface. These observations are consistent with the view (Woldringh et al., J. Bacteriol. 176 (1994) 6030-6038) that nucleoids, forced during replication to segregate in the length axis of the cell by the rigid bacillary envelope, induce assembly of FtsZ to division rings in between them. PMID- 11254987 TI - Discrimination of cartoons and photographs in pigeons: effects of scrambling of elements. AB - Four groups of pigeons were trained on four visual discrimination tasks using people photographs, pigeon photographs, cartoons of people, and cartoons of pigeons. After completion of the discrimination, the subjects were tested with new stimuli in the same stimulus categories but never used during the training, and scrambled stimuli in which elements of the original stimulus were spatially randomized. The subjects showed generalization to the new stimuli except for the people cartoons. When scrambled stimuli were presented, the subjects showed suppression of responding to the photographs but not to the people cartoon. These results suggest that the pigeons recognized both the elements and spatial arrangement of the photographs but ignored the spatial arrangement of the human cartoons. PMID- 11254986 TI - FtsZ rings in mukB mutants with or without the Min system. AB - The site of cell division in Escherichia coli is defined by formation of the Z ring between the two segregated daughter nucleoids. Positioning of the Z ring, composed of the highly conserved and tubulin-like FtsZ protein, appears to be negatively regulated by both the nucleoid and the oscillating MinCD inhibitor proteins. MukB protein is probably involved in nucleoid condensation, and in the absence of MukB, the negative effect of the nucleoid on Z rings appears to be partially suppressed. In this study, we examined the localization of Z rings in cells lacking both the Min system and MukB. In the Deltamin DeltamukB double null mutant, essentially all nucleoid-free zones, either at the cell poles or at non polar sites between nucleoids, contained Z rings. However, a significant proportion of Z rings also formed on top of nucleoids. Interestingly, Z ring clusters often formed at gaps between nucleoids, and some of the rings within the clusters were clearly positioned on top of nucleoids. These results provide further evidence that the negative topological effect of nucleoids in cells lacking MukB is partially but not totally suppressed, and that the absence of the Min system allows more promiscuous Z ring formation. PMID- 11254989 TI - The mechanisms of kin discrimination and the evolution of kin recognition in vertebrates: a critical re-evaluation. AB - I re-examine the four most widely proposed mechanisms of kin discrimination among vertebrates and conclude that the current categorization of kin discrimination mechanisms has been counterproductive because it has a hindered a clear understanding of the basic mechanisms by which animals discriminate kin. I suggest that there likely is only one authentic mechanism of kin discrimination and that this mechanism is learning, particularly associative learning and habituation. Observed differences in the way animals discriminate between kin and non-kin are due only to the cues (e.g., individually-distinctive, family distinctive, or self) that are used, and not to different mechanisms per se. I also consider whether kin discrimination is mediated by specially evolved kin recognition systems, defined as neural mechanisms that allow animals to directly classify conspecifics as either kin or non-kin. A preliminary analysis of vertebrate recognition systems suggests that specialized neural, endocrine, and developmental mechanisms specifically for recognizing kin have not evolved. Rather, kin discrimination results from an extension of other, non-specialized sensory and cognitive abilities of animals, and may be derived from other forms of social recognition, such as individual, group, or species recognition. PMID- 11254988 TI - Validation of a new system for the automatic registration of behaviour in mice and rats. AB - A newly developed behaviour registration system, Laboratory Animal Behaviour Observation, Registration and Analysis System (LABORAS) for the automatic registration of different behavioural elements of mice and rats was validated. The LABORAS sensor platform records vibrations evoked by animal movements and the LABORAS software translates these into the corresponding behaviours. Data obtained by using LABORAS were compared with data from conventional observation methods (observations of videotapes by human observers). The results indicate that LABORAS is a reliable system for the automated registration of eating, drinking, grooming, climbing, resting and locomotion of mice during a prolonged period of time. In rats, grooming, locomotion and resting also met the pre defined validation criteria. The system can reduce observation labour and time considerably. PMID- 11254990 TI - Development of play behavior between potential guide dogs for the blind and human raisers. AB - This study shows longitudinal development of human-dog interactions in a guide dog raising program. Social play interactions between potential guide dogs for the blind (puppies) and their adult female raisers (PWs) were videotaped at home during the period from two months to 11 or 12 months of the puppies' age. The puppies and PWs established close proximity relationships by two to three months of age. The older the puppies became, the shorter time the puppies and PWs spent in intensive fighting and chasing play. The older the puppies became, the longer the puppies and PWs spent in waiting, seeking and possessing together play which needed the puppies' self-control, concentration and ability to cooperate with the PWs. Whether or not these behavioral tendencies which the puppies showed were appropriate for guide dog candidates is also discussed. PMID- 11254991 TI - Effort- and time-cost effects on demand curves for food by pigeons under short session closed economies. AB - Demand curves for food were compared under the effort- and time-cost conditions using response-initiated fixed-ratio (FR) and fixed-interval (FI) schedules. For the effort-cost conditions, two pigeons were exposed to FR 3, 30, 90, and 150 for six sessions each. The time equivalent of each ratio was a FI schedule, each FI value equal to an average time from the first peck on the ratio to reinforcement (3-s access to mixed grain). The experiment was repeated in 1.5-, 3.0-, and 4.5-h closed economies in a different order for each pigeon. Food consumption as a function of time-based unit price (time equivalent per reinforcement duration) for each session length showed moderate convergence on a single demand function for the two cost conditions. When the demand function was separately fitted to each cost condition, however, the ranges of inelastic demand were generally larger in time-cost than effort-cost conditions. These curve-fitting analyses suggest that although time is a critical cost factor decreasing consumption at moderate prices, food intake under the effort-cost condition decreases more rapidly than under the time-cost condition as unit price increases. The analyses provide useful descriptions for the functional difference of costs. PMID- 11254992 TI - Transitive predatory relationships of spider species (Arachnida, Araneae) in laboratory tests. AB - Interspecific predation of spiders was studied in the laboratory in view of possible competition in the wild. Certain species killed other species even if handicapped by smaller size. Thirty eight spider species were involved in such a relationship and their predatory relationships were significantly reliable and transitive ('linear' or 'non-triangular'). A theridiid species (Theridion tinctum) showed the highest rank in terms of killing seven 'beta species', i.e. species capable of killing at least one alien species of larger size than themselves. Another theridiid (Steatoda grossa) obtained the second rank by killing five beta species. Experiments in both the wild and laboratory may, further, investigate other factors than body size that may be relevant to competition, such as behaviour-related characteristics (e.g. web structure and biting speed) and ecological factors (e.g. different susceptibilities of the species to parasite or predator attack). PMID- 11254993 TI - Songs of Holcostethus strictus (Fabricius): a different repertoire among landbugs (Heteroptera: Pentatomidae). AB - Songs emitted during mating by male and female Holcostethus strictus were recorded as substrate vibrations. Spectra of the vibrational signals have a dominant frequency peak between 100 and 260 Hz and in this respect reflect the general characteristic of the family Pentatomidae. Songs of H. strictus differ from the song repertoire of the southern green stink bug Nezara viridula (L.) (Pentatomidae) in many respects. The female calling and courtship songs differ in echeme and phrase duration. The male calling song is composed of spectrally different subunits. The male courtship song is characterised by three types of spectrally and temporally different echemes. The male copulatory song is composed of echemes of two types, which constitute a phrase of less regular temporal structure. In H. strictus, males start to sing first and female songs are less complex than in N. viridula. The female calling song is evoked by male calling and does not trigger male response. The female and male courtship song phrases are superimposed on one another and we have not observed any obvious regularity in their exchange. The possible role of different songs in H. strictus is discussed and compared with that in other pentatomide landbug species. PMID- 11254994 TI - Effects of ageing on allocentric and egocentric spatial strategies in the Wistar rat. AB - This study was designed to assess the effect of ageing on spatial (allocentric and egocentric) strategies in rats. Two different tasks were designed for this purpose: one involving Morris' circular pool (distal extramaze cues) and another using the T water maze (egocentric cues). In the first task, the aged rats showed some difficulty in acquiring allocentric spatial learning skills. After increasing the number of trials in this task, there was no significant improvement in the performance of the aged group of rats compared to the adult group. However, in the second spatial task (using egocentric cues), both age groups gave a similar performance. Therefore, the effect of ageing on spatial learning depends on the strategy required to acquire this learning. PMID- 11254995 TI - Trade-off between floor level and floor material in farmed silver foxes. AB - Farmed silver foxes (Vulpes vulpes) were allowed to balance their known preference for an elevated floor against their presumed preference for a sand floor. In Experiment 1, foxes had to choose between two identical cages, connected with an opening. One cage had a wire floor and the other had a sand floor, but the cages either were on the same (low or elevated) or on different levels (one cage 40 cm higher than the other). In Experiment 2, the cage pairs were connected with a 1.2 m long wire-mesh tunnel, one cage was always on a higher level (50 cm) than the other. In Experiment 1, foxes always preferred the sand floor during their active time. They also preferred the sand floor for resting, if it was on the same level as wire floor, but did not show any genuine preference if the floors were on different levels. In Experiment 2, foxes never preferred the lower floor. They preferred the elevated sand floor for activity and the elevated wire floor for lying. When two floors were identical they preferred the elevated one. Their rest consisted of 11-22 bouts, a major part of these being spent in the preferred cage. They also preferred a previous lying site to a new one, often exclusively and independently of floor material. In Experiment 1 foxes preferred the sand floor whereas in Experiment 2 they preferred the elevated floor indicating that the ability of a trade-off situation to rank resources depends on the method it is inflicted. PMID- 11254996 TI - Dissociation of food-finding and tentacle-lowering, following food-attraction conditioning in the snail, Helix aspersa. AB - The assumption that tentacle-lowering and food-finding may be used interchangeably as measures of food-attraction conditioning was examined in the snail, Helix aspersa. A brief pairing of an odor with the opportunity to feed (food-attraction conditioning) resulted in increased tendency to orient to that odor (food-finding), when tested the following day. In addition, conditioned snails exhibited increased levels of tentacle-lowering. A more detailed analysis revealed that a subset of conditioned snails exhibited successful food-finding in the absence of tentacle-lowering, and that another subset of conditioned snails exhibited increased levels of tentacle-lowering in the absence of successful food finding. These results suggest that caution should be observed when comparing results across these two response systems. PMID- 11254997 TI - Pigeons' timing of an arbitrary and a naturalistic auditory stimulus: tone versus cooing. AB - Previous animal research has traditionally used arbitrary stimuli to investigate timing in a temporal bisection procedure. The current study compared the timing of the duration of an arbitrary, auditory stimulus (a 500-Hz tone) to the timing of the duration of a naturalistic, auditory stimulus (a pigeon cooing). In the first phase of this study, temporal perception was assessed by comparing psychophysical functions for the duration of tone and cooing signals. In the first set of tests, the point of subjective equality (PSE) was significantly lower for the tone than for the cooing stimulus, indicating that tones were judged longer than equivalent durations of cooing. In the second set of tests, gaps were introduced in the tone signal to match those present in the cooing signal, and no significant difference in the PSE for the tone or the cooing signal was found. A repetition of the testing conducted with gaps removed from the tone signal, failed to replicate the difference in the PSEs for the tone and cooing signals originally obtained. In the second phase of the study, memory for the duration of tone and cooing was examined, and a choose-long bias was found for both signals. Based on these results, it appears that, for pigeons, there may be no significant differences in either temporal perception or temporal memory for arbitrary, auditory signals and more complex, naturalistic, auditory signals. PMID- 11254998 TI - Urine marking and social dominance in male house mice (Mus musculus domesticus). AB - House mice use urine marking for a variety of forms of social communication. Urine marking varies with dominance status; socially dominant male house mice urine mark more than those that are socially subordinate. Experiment I was designed to confirm this previous finding. Experiment II was designed to test whether urine marking, measured prior to testing males for aggression, could be used to predict social dominance. Mice were tested for urine marking in 20 cmx40 cm rectangular cages with filter paper below the wire mesh bottom of the cage. In Experiment I, groups of four males were tested in a round robin design to assess social dominance and were then placed individually in urine marking cages. Social dominance was a significant predictor of the number of 1 cm squares that contained urine marks, both with regard to interior squares and for perimeter squares in the test cage. In Experiment II, groups of four males were first tested individually in urine marking cages and then used for round robin aggressive encounters to assess social dominance. The number of interior squares with urine marks, and, to a lesser extent, the number of perimeter squares with urine marks, were both significant predictors of aggression scores and social dominance status. Being able to judge social dominance without having the mice encounter each other could be a valuable tool for future work; confounding effects on such parameters as hormone levels could be avoided while obtaining an estimate of male social dominance status. PMID- 11254999 TI - Effect of melatonin supplementation on the sexual development in European quail (Coturnix coturnix). AB - At the end of their wintering phase, male European quails were exposed to a stimulation photoperiod of light/dark 12:12 h for 10 days to induce sexual development. A daily oral melatonin supplementation was then given to one group of treated males (N=11) and the alcohol solvent was given to a control group of males (N=10). These solutions were provided during the final 3 h of the photophase for 28 days, then during the final 4 h for 18 days. There were no significant differences between the two groups with respect to fat levels. However, 3 weeks after the beginning of melatonin supplementation, the sexual development of the treated birds slowed down. The importance of this decline varied to a greater or lesser degree between individual birds. When melatonin supplementation stopped, sexual development resumed. Activity recordings revealed a decrease in feeding activity when melatonin supplementation was provided. However, this response showed important interindividual variability. The birds that produced the most marked responses to melatonin during the first 3 weeks of supplementation were those that also showed the most obvious decline in sexual development. It seems that, in European quail, a wild migratory species that always shows a natural biological annual rhythm, a melatonin signal could play a role in regulating reproduction. PMID- 11255000 TI - Are there multiple mating strategies in blue sheep? PMID- 11255001 TI - Male and female agonistic and affiliative relationships in a social group of farm cats (Felis catus L.). AB - A dominance hierarchy based on the outcome of agonistic encounters was found among male and female domestic cats. A female dominated over some males. The dominance concept is also discussed in terms of social bonding. The relationships among adult females were amicable, whereas adult males showed reciprocal tolerance. The flow of affiliative behaviour was directed mainly from females to one male of the group. The analysis of marking behaviour showed that this male sprayed urine and rubbed the perioral and cheek regions of the face on the objects of the environment at a higher rate than the other members of the group. Nevertheless, rubbing the perioral and cheek regions of the face on objects was not correlated to dominance rank, possibly because it has some function in social communication other than territorial defence against strangers. No relationships have been found between claw scratching, rolling on the ground and social rank, or between the former and other marking behaviour. It is concluded that claw scratching and rolling were not utilised to mark territory. PMID- 11255002 TI - Gene duplication and mobile genetic elements in the morning glories. AB - We review gene duplication and subsequent structural and functional divergence in the anthocyanin biosynthesis genes in the Japanese and common morning glories and discuss their evolutionary implications. These plants appear to contain at least six copies of the CHS gene and three tandem copies of the DFR gene. Of these, the CHS-D and DFR-B genes are mainly responsible for flower pigmentation and mutations in these genes confer white flowers. We compared the genomic sequences of these duplicated genes between the two morning glories and found small mobile element-like sequences (MELSs) and direct repeats (DRs) in introns and intergenic regions. The results indicate that the MELS elements and DRs play significant roles in divergence after gene duplication. We also discuss DNA rearrangements occurring before and after speciation of these morning glories. DNA transposable elements belonging to the Ac/Ds or En/Spm families have acted as major spontaneous mutagens in these morning glories. We also describe the structural features of the first Mu-related element found in the morning glories and polymorphisms found in the same species. PMID- 11255003 TI - Regulation of mRNA stability in mammalian cells. AB - The regulation of mRNA decay is a major control point in gene expression. The stability of a particular mRNA is controlled by specific interactions between its structural elements and RNA-binding proteins that can be general or mRNA specific. Regulated mRNA stability is achieved through fluctuations in half-lives in response to developmental or environmental stimuli like nutrient levels, cytokines, hormones and temperature shifts as well as environmental stresses like hypoxia, hypocalcemia, viral infection, and tissue injury. Furthermore, in specific disorders like some forms of neoplasia, thalassemia and Alzheimer's disease, deregulated mRNA stability can lead to the aberrant accumulation of mRNAs and the proteins they encode. This review presents a discussion of some recently identified examples of regulated and deregulated mRNA stability in order to illustrate the diversity of genes regulated by alterations in the degradation rates of their mRNAs. PMID- 11255004 TI - Genetic analysis of the T4 holin: timing and topology. AB - The t protein of bacteriophage T4 shares with other holins the ability to cause the formation of a lethal membrane lesion which allows the phage endolysin to attack the peptidoglycan. Moreover, T, like other holins, acts in a saltatory manner at a precisely programmed time in the vegetative cycle. Unlike other holins, however, T has the unique ability to postpone its lethal function in response to a secondary infection by a T-even phage during the vegetative cycle. A signal transduction system that responds to the secondary infection is thought to be encoded by some of the numerous r genes, defined by mutations that abolish this lysis-inhibition (LIN) response. The primary structure of T differs from two main structural patterns found in more than 30 orthologous groups of holins. Genetic approaches were taken to probe the t sequence for features involved in membrane localization, functional timing and LIN regulation. Gene fusion analysis indicates that T has a single TMD near the N-terminus, with the bulk of the protein residing in the periplasm. Mapping and phenotypic analysis of deletion and point mutations in t indicates that the periplasmic domain of T is the major determinant of the timing mechanism and is involved in the LIN response. PMID- 11255005 TI - Nucleotide sequence and structure of the mouse carbonic anhydrase III gene. AB - At least 14 distinct isozymes of carbonic anhydrase have been identified in mammals. These enzymes catalyze the hydration of carbon dioxide and are essential for regulation of cellular pH and carbon dioxide transport. Carbonic anhydrase III is highly expressed in certain tissues, including muscle and fat where it constitutes up to 25% of the soluble protein. We cloned a cDNA encoding mouse carbonic anhydrase III. This cDNA contains 1653 bp, consisting of 79 bp in the 5' UTR, a 780 bp open reading frame, and 794 bp of the 3' UTR, including two potential polyadenylation signals. Fluorescent in situ hybridization confirmed the existence of a single copy of the gene on chromosome 3. We then isolated the genomic DNA for mouse carbonic anhydrase III and analyzed its structure. The gene consists of seven exons and six introns which span 10.5 kb. The 5' flanking region of the genomic DNA is notable for a pyrimidine rich region consisting of two dinucleotide repeats containing 23 and 20 TC pairs separated by the same 15 bp spacer. PMID- 11255006 TI - Variants of the 5'-untranslated region of the bovine growth hormone receptor mRNA: isolation, expression and effects on translational efficiency. AB - The growth hormone receptor (GHR) gene in cattle is expressed as multiple GHR mRNA variants that differ in the 5'-untranslated region (5'-UTR). Three GHR mRNA 5'-UTR variants (named 1A, 1B, and 1C) were isolated in previous studies. Six additional GHR mRNA 5'-UTR variants (named 1D, 1E, 1F, 1G, 1H, and 1I) were discovered in the present study by using rapid amplification of cDNA ends (RACE). Ribonuclease protection assays (RPA) indicated that variant 1A was exclusively expressed in liver, variant 1F was expressed in liver and muscle, and most of the remaining GHR 5'-UTR variants were expressed in a variety of tissues including liver and muscle. The RPA also indicated that in liver and skeletal muscle, two important GH target tissues, the most abundant GHR mRNA 5'-UTR variants were 1A and 1B (liver) and 1B and 1C (muscle), respectively. In vitro translation assays indicated that the GHR 5'-UTR variants also differed in their effects on the translation of a luciferase reporter mRNA. Notably, the GHR 5'-UTR 1B, which is a highly expressed GHR 5'-UTR variant in vivo, strongly inhibited translation of the luciferase reporter mRNA. The observations in the present study suggest that GHR expression in the bovine may be controlled by translational mechanisms in addition to complex transcriptional mechanisms. PMID- 11255007 TI - ERGL, a novel gene related to ERGIC-53 that is highly expressed in normal and neoplastic prostate and several other tissues. AB - We have identified a new gene, that is highly expressed in normal and neoplastic prostate, and is also expressed in cardiac atrium, salivary gland, spleen and selective cells in the CNS. Database analyses of ESTs indicated prostate specificity but experimental results showed the expression in other tissues. The full length transcript is 1800 bp with an open reading frame of 526 aa. The amino terminal 230 residues of the expressed protein has high homology to a family of lectins, especially to the sugar binding domain of ERGIC-53. We therefore designate the new gene ERGL (ERGIC-53-like). There is a transmembrane domain at amino acid positions 468-482 suggesting that the product of ERGL is a type-I membrane protein. In prostate there are two fully processed transcripts one of which is a splice variant with a deletion in the region of the transmembrane domain of the protein. PMID- 11255008 TI - LHX3 transcription factor mutations associated with combined pituitary hormone deficiency impair the activation of pituitary target genes. AB - The Lhx3 LIM homeodomain transcription factor is critical for pituitary gland formation and specification of the anterior pituitary hormone-secreting cell types. Two mutations in LHX3, a missense mutation changing a tyrosine to a cysteine and an intragenic deletion that results in a truncated protein lacking the DNA-binding homeodomain, have been identified in humans. These mutations were identified in patients with retarded growth and combined pituitary hormone deficiency and also abnormal neck and cervical spine development. For both the LHX3a and LHX3b isoforms, we compared the ability of wild type and mutant LHX3 proteins to trans-activate pituitary genes, bind DNA recognition elements, and interact with partner proteins. The tyrosine missense mutation inhibits the ability of LHX3 to induce transcription from selected target genes but does not prevent DNA binding and interaction with partner proteins such as NLI and Pit-1. Mutant LHX3 proteins lacking a homeodomain do not bind DNA and do not induce transcription from pituitary genes. These studies demonstrate that mutations in the LHX3 isoforms impair their gene regulatory functions and support the hypothesis that defects in the LHX3 gene cause complex pituitary disease in humans. PMID- 11255009 TI - Polyadenylation of ribosomal RNA by Candida albicans. AB - Candida albicans is the leading fungal pathogen in immunocompromised patients such as those with AIDS and malignancies. It is a polymorphic organism existing as a unicellular yeast or as filamentous forms that include pseudohyphae and true hyphae. While studying the early period of hyphal transformation, comparing cDNAs from yeast to those in early transition, we were surprised to find 25S rRNA represented frequently in our differential display assays, suggesting that our reverse transcription with poly-T primers was copying rRNA with extended poly-A 3' ends. We now report that both the yeast forms and germinating organisms polyadenylate some of their 25S rRNA transcripts. We also found a rapid and transient enhancement of this process upon stimulation with serum. These data indicate that 25S rRNA polyadenylation is part of the biological repertoire of C. albicans and its transient upregulation just prior to hyphal development raises the possibility of a regulatory role in this transition. PMID- 11255010 TI - Cloning of maltase gene from a methylotrophic yeast, Hansenula polymorpha. AB - The Hansenula polymorpha maltase structural gene (HPMAL1) was isolated from a genomic library by hybridization of the library clones with maltase-specific gene probe. An open reading frame of 1695 nt encoding a 564 amino-acid protein with calculated molecular weight of 65.3 kD was characterized in the genomic DNA insert of the plasmid p51. The protein sequence deduced from the HPMAL1 exhibited 58 and 47% identity with maltases from Candida albicans and Saccharomyces carlsbergesis encoded by CAMAL2 and MAL62, respectively, and 44% identity with oligo-alpha-1,6-glucosidase from Bacillus cereus. The recombinant Hansenula polymorpha maltase produced in Escherichia coli hydrolyzed p-nitrophenyl-alpha-D glucopyranoside (PNPG), sucrose, maltose and alpha-methylglucoside and did not act on melibiose, cellobiose, trehalose and o-nitrophenyl-beta-D galactopyranoside (ONPG). The affinity of the recombinant enzyme for its substrates increased in the order maltose .05). After LASIK, corneal sensation was significantly decreased at 3 days, 1 week, and 1 month; it recovered slightly at 3 months, although it remained significantly less than preoperatively. CONCLUSIONS: Laser in situ keratomileusis was associated with a negative effect on corneal sensation, which was markedly greater than the effect with PRK and was evident for at least 3 months after surgery. PMID- 11255048 TI - Laser in situ keratomileusis after penetrating keratoplasty. AB - PURPOSE: To assess the outcomes of laser in situ keratomileusis (LASIK) after penetrating keratoplasty (PKP). SETTING: Hospital de Clinicas de Porto Alegre, Department of Ophthalmology, Federal University of Rio Grande do Sul, Porto Alegre, RS, Brazil. METHODS: Fourteen eyes of 13 patients who had LASIK after PKP were retrospectively reviewed. The interval between LASIK and PKP was at least 1 year, and the follow-up after LASIK was also at least 1 year. All patients had a stable refractive error for a minimum of 6 months after all sutures were removed, regular and symmetric topographic astigmatism, and a minimal ultrasonic central corneal pachymetry of 500 microm. The Chiron Automatic Corneal Shaper and the Meditec Aesculap MEL 60 excimer laser were used. RESULTS: At 12 months, mean myopia decreased from -5.33 diopters (D) +/- 4.22 (SD) to 0.19 +/- 1.71 D, mean hyperopia decreased from +5.04 +/- 3.32 D to + 0.42 +/- 0.46 D, and mean astigmatism decreased from 5.37 +/- 2.12 D to 2.82 +/- 2.42 D (47.5% of mean percentage reduction). Retreatment was necessary in 42.9% of eyes because of cylindrical undercorrection. Uncorrected visual acuity improved in 11 eyes (78.6%). Best spectacle-corrected visual acuity improved in 6 eyes (42.8%) and was maintained in 4 eyes (28.6%); 5 eyes (35.7%) lost 1 Snellen line. Intraoperative complications included 1 buttonhole flap. Postoperative complications included interface epithelial ingrowth at the periphery (2 eyes) and pseudophakic retinal detachment 2 years after LASIK (1 eye). CONCLUSION: Laser in situ keratomileusis after PKP safely and predictably corrected the spherical component of the refraction. However, the predictability of LASIK in correcting post-PKP astigmatism was poor. PMID- 11255049 TI - Implantable contact lens for moderate to high myopia: short-term follow-up of 2 models. AB - PURPOSE: To confirm the safety, efficacy, and predictability of the surgical correction of moderate to high myopia by the ICM V3 and ICM V4 implantable contact lenses (ICLs), with emphasis on vaulting, intraocular pressure (IOP), and pigment dispersion. SETTING: University Eye Hospital, Lausanne, Switzerland. METHODS: Thirty-two eyes had implantation of an ICL. In 22 eyes with a mean spherical equivalent (SE) of -11.5 diopters (D), the target was emmetropia; in 10 eyes with a mean SE of -22.3 D, the goal was a reduction in the myopia. Nineteen eyes received the ICM V3 ICL and 13, the ICM V4 ICL. The mean follow-up was 7.4 months. RESULTS: The mean postoperative SE in the 32 eyes was -2.16 D. Best spectacle-corrected visual acuity was maintained or improved in all eyes. In the 22 eyes targeted to achieve emmetropia, 10 (45%) were within +/-1.00 D; 15 (68%) had an uncorrected visual acuity of 20/40 or better and 4 (18%), of 20/20 or better. Vaulting of the ICL over the crystalline lens was more pronounced with the V4 than with the V3, and the difference was statistically significant. Subtle, localized anterior subcapsular opacification was encountered in 4 eyes. In 3 of them, the ICL (model V3) vaulting was minimal and 1 ICL (model V4) did not show any vaulting. Eighteen eyes had an IOP higher than the preoperative level, and the difference was statistically significant. No correlation was seen between final IOP and vaulting. Pigment dispersion on the ICL did not appear to be related to vaulting or ICL thickness. CONCLUSION: Implantation of an ICL was effective in correcting moderate to high myopia of up to -17.50 D. Although the procedure appears to be safe, the predictability of the refractive outcome must be improved. The new generation of ICLs for myopia (ICM V4) offers a better vault over the crystalline lens than the older models (ICM V3), which should decrease the risk of cataract. No explanation was found for the IOP increase in several eyes 3 months or more after surgery. PMID- 11255050 TI - Hyperopic laser in situ keratomileusis to treat overcorrected myopic LASIK. AB - PURPOSE: To evaluate the safety and efficacy of hyperopic laser in situ keratomileusis (LASIK) in treating hyperopia caused by overcorrected myopic LASIK and to evaluate a new technique to place the hyperopic treatment after lifting the initial myopic flap. SETTING: Open-access outpatient excimer laser surgical facility. METHODS: A retrospective analysis was performed of 54 eyes in 47 patients who had spherical hyperopic LASIK by 21 surgeons for the treatment of significant hyperopia after overcorrected LASIK for myopia. In 42 eyes, the initial LASIK flaps were lifted and in 12 eyes, new flaps were cut. The mean age of the 25 men (53%) and 22 women (47%) was 48.2 years +/- 8.4 (SD). Outcome measures included refractive error, uncorrected visual acuity (UCVA), best spectacle-corrected visual acuity (BSCVA), and complications. The mean follow-up was 2.97 months. RESULTS: In eyes in which postoperative emmetropia was attempted (n = 45), the mean spherical equivalent improved from +1.21 +/- 0.49 diopters (D) preoperatively to -0.38 +/- 0.50 D postoperatively (P <.001). The mean UCVA improved from 20/38.6 +/- 16.3 to 20/27.4 +/- 9.4 (P <.001). At the last follow up, 69% of eyes were within +/-0.5 D and 96% were within +/-1.0 D of emmetropia; 42% had a UCVA of 20/20 and 96% had a UCVA of 20/40 or better. No eyes lost 2 or more lines of BSCVA. No vision-threatening complications occurred. Results in patients who had initial flaps lifted and those who had new flaps cut were statistically indistinguishable. On average, achieved hyperopic corrections were 18% greater than intended. CONCLUSION: Hyperopic LASIK was safe, predictable, and effective in the treatment of hyperopia caused by overcorrected myopic LASIK. Results were similar whether the original flap was lifted or a new one was cut. PMID- 11255051 TI - Surgically induced astigmatism after hyperopic and myopic photorefractive keratectomy. AB - PURPOSE: To compare the axis and magnitude of surgically induced refractive astigmatism (SIA) after hyperopic and myopic photorefractive keratectomy (PRK). SETTING: Department of Ophthalmology, University of Texas Southwestern Medical Center, Dallas, Texas, USA. METHODS: In this single-center retrospective study, the VISX Star S2 excimer laser was used to create a peripheral annular ablation profile to correct spherical hyperopia in 23 eyes of 16 consecutive patients. Attempted corrections ranged from +0.50 diopter (D) to +4.25 D with 0 to 1.00 D of astigmatism. The same laser was used to create a central ablation profile to correct spherical myopia in 25 eyes of 17 consecutive patients. Attempted corrections ranged from -2.25 to -6.50 D with 0 to 1.00 D of astigmatism. The absolute change in refractive astigmatism was calculated by taking the difference in magnitudes of astigmatism before and after laser treatment without regard to axis. Axis and magnitude of SIA were analyzed by vector differences. Magnitudes were compared using the Student t test, and axial shifts were compared using the chi-square test. All patients were followed for a minimum of 6 months. RESULTS: The mean changes in absolute astigmatism were 0.29 +/- 0.28 D at 3 months and 0.34 +/- 0.29 D at 6 months after hyperopic PRK and 0.40 +/- 0.35 D at 3 months and 0.39 +/- 0.36 D at 6 months after myopic PRK. The mean vectoral magnitudes were 0.49 +/- 0.29 at 3 months and 0.52 +/- 0.25 at 6 months after hyperopic PRK and 0.48 +/- 0.39 at 3 months and 0.44 +/- 0.38 at 6 months after myopic PRK. The mean values for SIA (the centroid) were 0.10 +/- 0.57 D x 113 degrees at 3 months and 0.15 +/- 0.57 D x 131 degrees at 6 months after hyperopic PRK and 0.04 +/- 0.63 D x 160 degrees at 3 months and 0.08 +/- 0.58 D x 171 degrees at 6 months after myopic PRK. There was no statistically significant difference between the 2 groups in vectoral axis or magnitude of SIA. CONCLUSION: Surgically induced astigmatism after hyperopic PRK was comparable to astigmatism induced by myopic PRK. A peripheral annular ablation for hyperopic correction, similar to a central ablation in myopic PRK, did not appear to result in uneven corneal healing causing astigmatism. PMID- 11255052 TI - Correlation between ultraviolet radiation level and the incidence of late-onset corneal haze after photorefractive keratectomy. AB - PURPOSE: To investigate the correlation between environmental changes in ultraviolet (UV) radiation levels and the incidence of late-onset cornea haze (LOCH) after photorefractive keratectomy (PRK). SETTING: SynsLaser Clinic, Tromso, Norway. METHODS: The study comprised 404 eyes that had myopic PRK and photoastigmatic refractive keratectomy from February 1996 through July 1998. The high latitude (70 degrees N) of the observation site provided "natural laboratory" conditions to look at the occurrence of LOCH with high and low UV radiation levels, which occurred during summers and winters, respectively. The diagnostic criterion for LOCH was acute haze of grade > or =2 occurring between 4 and 12 months postoperatively. RESULTS: The follow-up ranged from 12 to 41 months. Of the 314 eyes that met the inclusion criteria, 11 developed LOCH when the environmental UV-radiation level was high. No eye developed LOCH when the level was low. The correlation between a high level of environmental UV radiation and the occurrence of LOCH was statistically significant (P =.001). CONCLUSION: Environments with high UV-radiation levels may increase the risk of LOCH after PRK in eyes with moderate to high myopia. Use of UV-protective eyewear should be encouraged during the first year after PRK. PMID- 11255053 TI - Surgically induced astigmatism after implantation of intacs intrastromal corneal ring segments. AB - PURPOSE: To analyze surgically induced astigmatism (SIA) after implantation of Intacs intrastromal corneal ring segments. SETTING: Multicenter clinical trials in the United States. METHODS: Data from 11 investigational sites involved in the Phase II and III trials of Intacs for the United States Food and Drug Administration were retrospectively analyzed. The distribution of Intacs thicknesses implanted in 449 eyes was 0.25 mm in 148 eyes, 0.30 mm in 151 eyes, and 0.35 mm in 150 eyes. Refractive astigmatism was measured by subjective manifest refraction preoperatively and 1 week and 1, 2, 3, 6, 9, and 12 months postoperatively. The mean simple change in astigmatism and the surgically induced refractive change were determined by vector analysis. RESULTS: Mean induced astigmatism at 12 months was 0.13 diopter (D) +/- 0.52 (SD). Induced astigmatism was more frequently with the rule (44%) than against the rule (26%) or oblique (30%). Maximal mean astigmatism was 0.50 +/- 1.09 D and occurred at 7 days. Mean induced astigmatism increased with segment thickness (0.01 D, 0.17 D, and 0.21 D for the 0.25 mm, 0.30 mm, and 0.35 mm segments, respectively). Mean surgically induced refractive change in cylinder power in all eyes at 12 months by vector analysis was 0.17 D x 92. CONCLUSION: Mean SIA was not clinically meaningful 12 months after Intacs implantation. PMID- 11255054 TI - Clinical results of phacoemulsification with the use of Healon5 or Viscoat. AB - PURPOSE: To compare the ophthalmic viscosurgical devices Healon5 (viscoadaptive) and Viscoat (dispersive) regarding their overall clinical performance during phacoemulsification and posterior chamber intraocular lens (IOL) implantation as well as their influence on intraocular pressure (IOP). SETTING: Department of Ophthalmology, Ruprecht-Karls-University Heidelberg, Heidelberg, Germany. METHODS: In this prospective randomized patient- and observer-masked clinical study, the performance of Healon5 (sodium hyaluronate 2.3%) and Viscoat (sodium hyaluronate 3.0%-chondroitin sulfate 4.0%) was assessed by 3 surgeons during cataract surgery in 90 patients. Surgeons used a 5-point scale for the subjective assessment of the ease of injection, maintenance capacity during continuous curvilinear capsulorhexis, remaining capacity during phacoemulsification, facilitation of IOL implantation, removal from the eye, transparency, and overall performance throughout surgery. Intraocular pressure was measured preoperatively and 24 hours and 7 days postoperatively. Best corrected visual acuity was assessed preoperatively and 7 days postoperatively. RESULTS: Overall intraoperative product performance was assessed as good or very good in 34 of 44 patients (77%) in the Healon5 group and in 16 of 46 patients (35%) in the Viscoat group (P <.001). Retention in the anterior chamber was graded good or very good in 36 patients (82%) in the Healon5 group and in 23 (50%) in the Viscoat group (P =.001). There were no statistically significant between-group differences in mean IOP preoperatively and 24 hours postoperatively. CONCLUSIONS: Surgeons graded Healon5 better than Viscoat in overall surgical performance and retention in the anterior chamber during phacoemulsification. These data support that Healon5 adapts to each step during surgery. PMID- 11255055 TI - Intraocular pressure after small incision cataract surgery: temporal sclerocorneal versus clear corneal incision. AB - PURPOSE: To compare intraocular pressure (IOP) after phacoemulsification and foldable intraocular lens (IOL) implantation using a temporal sclerocorneal or clear corneal incision. SETTING: Department of Ophthalmology, Johannes Gutenberg University, Mainz, Germany. METHODS: One hundred patients (100 eyes) with cataract having phacoemulsification with posterior chamber IOL implantation were randomly assigned to receive a temporal sclerocorneal or clear corneal tunnel incision. Intraocular pressure was measured preoperatively and 6 hours, 1, 2, and 3 days, and 5 months postoperatively. Statistical significance was determined by nonparametric group comparisons using 2-sample random Wilcoxon tests. RESULTS: Six hours postoperatively, the median IOP increase was significantly higher in the sclerocorneal tunnel group (57%) than in the clear corneal incision group (18%) (P <.001). No significant between-group difference in IOP was found at 1, 2, or 3 days or 5 months. At 5 months, IOP was 0.6 mm Hg lower than preoperatively in the sclerocorneal tunnel group and 1.5 mm Hg lower in the clear corneal group. CONCLUSIONS: After phacoemulsification and foldable IOL implantation, the immediate postoperative IOP increase was higher in eyes having a sclerocorneal incision than in those having a clear corneal tunnel incision. These results could be important in eyes with decreased outflow facility or preexisting optic nerve damage. PMID- 11255056 TI - Intraocular pressure after phacoemulsification and intraocular lens implantation in nonglaucomatous eyes with and without exfoliation. AB - PURPOSE: To evaluate intraocular pressure (IOP) after phacoemulsification and intraocular lens (IOL) implantation in nonglaucomatous eyes with and without exfoliation. SETTING: Department of Ophthalmology, Helsinki University Central Hospital, Helsinki, Finland. METHODS: One hundred ninety-six eyes that had phacoemulsification with IOL implantation were examined. Eyes with a history of intraocular disease or surgery that could affect IOP were excluded. The study comprised the remaining 160 eyes: 23 with exfoliation (EXF group) and 137 without exfoliation (non-EXF group). Follow-up data were available for 136 eyes. The same surgeon performed all surgeries. Patients were examined on the first postoperative day and after 1 week, 4 months, and 1.0 to 2.7 years. RESULTS: One day postoperatively, IOP rose in the EXF group from a mean preoperative level of 16.3 mm Hg +/- 2.7 (SD) to 21.0 +/- 8.5 mm Hg, a 28.4% increase (P =.0061). In the non-EXF group, mean IOP rose from 16.2 +/- 3.4 mm Hg to 20.5 +/- 5.7 mm Hg, a 29.9% increase (P =.001). In 4 eyes (17.4%) in the EXF group and 8 eyes (5.8%) in the non-EXF group, IOP increased to 30 mm Hg or higher at 1 day. After this, significant IOP decreases occurred in both the EXF and non-EXF group, respectively, as follows: 14.2 +/- 3.0 mm Hg (12.0% decrease from preoperative value; P =.001) and 15.0 +/- 2.9 mm Hg (5.9%; P =.001) 1 week postoperatively; 12.8 +/- 2.7 mm Hg (20.2%; P =.0002) and 13.8 +/- 2.7 mm Hg (13.2%; P =.001) after 4 months; and 12.3 +/- 2.6 mm Hg (23.2%; P =.0001) and 12.7 +/- 2.7 mm Hg (21.2%; P =.001) after 1.0 to 2.7 years. There was no significant difference between the 2 groups. CONCLUSION: After phacoemulsification with IOL implantation, IOP decreased significantly and remained lower than preoperatively in eyes with and without exfoliation. One day postoperatively, transient pressure peaks were more common in eyes with exfoliation. One eye without exfoliation developed glaucoma. PMID- 11255057 TI - Cystoid macular edema after pediatric intraocular lens implantation: fluorescein angioscopy results and literature review. AB - PURPOSE: To evaluate the occurrence of cystoid macular edema (CME) after lens extraction, anterior vitrectomy, and intraocular lens implantation (IOL) in children using angioscopy after administration of oral fluorescein. SETTING: Centers in Tamil Nadu, India, and Hong Kong, China. METHODS: This study comprised 18 children (28 eyes) who had cataract extraction, posterior capsulorhexis, anterior vitrectomy, and in-the-bag IOL implantation. The presence of CME was evaluated 1 week and 4 to 6 weeks after surgery using fluorescein angioscopy. RESULTS: Anterior chamber fibrin occurred in 4 eyes (14.3%). Fluorescein angioscopy was performed 1 week after surgery in all eyes and after 1 month in 25 eyes (89.3%). No eye demonstrated the presence of CME on fluorescein angioscopy. CONCLUSIONS: Cystoid macular edema did not occur in the early period after pediatric cataract surgery using current surgical techniques. Longer follow-up is required to ascertain the incidence of CME in the late postoperative period. PMID- 11255058 TI - Posterior subcapsular and nuclear cataract after vitrectomy. AB - PURPOSE: To examine the morphological changes in the postvitrectomy lens and to monitor the development of these changes over time. SETTING: Oxford Eye Hospital, Oxford, United Kingdom. METHODS: In this prospective study, 33 consecutive phakic patients having pars plana vitrectomy were recruited. Cataract development was quantified by clinical grading and digital Scheimpflug image analysis. Slitlamp biomicroscopy and photography were used to document the morphological appearance. The main outcome measures were the incidence, morphology, and development of posterior subcapsular and nuclear cataract. RESULTS: A characteristic, transient posterior subcapsular cataract (PSC) was present in 89% (17 of 19) of tamponade patients within 24 hours of surgery. Of the patients who had vitrectomy without tamponade, 9% (1 of 11) developed similar changes. Nuclear opacity developed in 61% (11 of 18) of tamponade patients and in 50% (3 of 6) of nontamponade patients. A longer term retrospective review of the same patients' case notes revealed nuclear cataract in 67% (12 of 18) of tamponade cases and 30% (3 of 10) of nontamponade cases. Eighteen percent (2 of 11) of nontamponade cases and 67% (14 of 21) of tamponade cases had cataract surgery after a 10.7 month and a 12.4 month follow-up, respectively. CONCLUSIONS: Vitrectomy and tamponade produced a characteristic transient PSC in the immediate postoperative period. Disruption of fluid balance in the region of the posterior lens was suggested by the morphological appearance. The acute changes resolved but were followed by accelerated nuclear opacification. PMID- 11255060 TI - Effect of a capsular tension ring on the shape of the capsular bag and opening and the intraocular lens. AB - PURPOSE: To evaluate the effect of a capsular tension ring (CTR) on the shape of the capsular bag, the extent of the capsular opening, and the shape of intraocular lenses (IOLs). SETTING: Department of Ophthalmology, College of Medicine, The Catholic University of Korea, Seoul, Korea. METHODS: The corneal button was removed from porcine eyes in vitro. After phacoemulsification was performed, an IOL alone or an IOL and CTR were inserted in the capsular bag in 6 groups of 5 eyes each. The eyes were examined from the posterior aspect using a Miyake technique to assess capsular bag shape and the capsular opening. To evaluate effects of the CTR on IOL shape, rabbit eyes had phacoemulsification and IOL implantation with and without placement of a CTR in vivo. The IOLs were removed from enucleated eyes 3 months postoperatively and compared with unused control IOLs. RESULTS: The differences between the maximum and minimum diameters of the capsular bags and capsular openings were significantly less in groups with a CTR. Intraocular lens size (difference from haptic to haptic) decreased significantly in eyes with only an IOL compared with normal controls or eyes with both an IOL and CTR. CONCLUSIONS: The CTR preserved the integrity of the capsular bag diameter, capsular opening, and IOL shape. It is likely that CTR implantation can avert contracture of the capsular bag and capsular opening, preventing IOL decentration. PMID- 11255059 TI - Topical anesthesia for cataract surgery: a population-based perspective. AB - PURPOSE: To analyze the anesthetic regimen and sedation in a population-based cohort of unselected cataract surgery cases operated on with the goal of maximizing the percentage of patients with topical anesthesia and no sedation. SETTING: Department of Ophthalmology, Norrlands University Hospital, Umea, Sweden. METHODS: This prospective observational population-based study comprised all patients having cataract surgery during a 1 year period at 1 institution. Data were collected from the patients' records, which were standardized. Outcome measures were use of preoperative sedation, type of anesthesia, intraoperative complications, and adverse events. RESULTS: The study comprised 890 cases performed by 4 surgeons. Seventy-two percent of patients had no sedation and topical anesthesia only. All patients except 1 who had previous cataract surgery with topical anesthesia chose the same method for their second-eye surgery. The rate of posterior capsule rupture was in the expected range for a population with high incidences of pseudoexfoliation and mature cataract. CONCLUSIONS: It is possible to achieve a high percentage of effective topical anesthesia for cataract surgery in a population-based setting. The findings have implications for cost-containment in health services. PMID- 11255061 TI - Experimental studies of capsular equator rings of soft latex. AB - PURPOSE: To determine the efficacy of wide equatorial rings of various materials in reducing posterior capsule opacification (PCO) and maintaining the circular contour of the capsular equator in a rabbit model. SETTING: Laboratory for Intraocular Microsurgery and Implants, Goldschleger Eye Research Institute, Tel Aviv University, Sheba Medical Center, Tel-Hashomer, Israel. METHODS: Closed rings made of poly(methyl methacrylate), Teflon(R), polyurethane, and latex were introduced into the capsular bags of rabbit eyes after removal of the lens material. The soft latex rings were deemed the only practical rings for atraumatic implantation through a small incision. Latex rings of 2 dimensions with rectangular edges were implanted in 8 rabbits. One eye served as a control after lens removal and no ring implantation. The eyes were followed for up to 5 months and then evaluated by light microscopy. RESULTS: Posterior capsule opacification was minimal or absent in all animals implanted with the latex rings. Mild to moderate inflammatory reaction was noted in most eyes. It was caused by the ring material or secondary to long-standing pressure on the ciliary tissue. The rings also maintained well the circular contour of the capsular bags. CONCLUSIONS: Equatorial rings of soft latex with sharp rectangular edges were effective in minimizing PCO and maintaining the shape of the capsule after lens removal in rabbits. Soft rings made of a biocompatible material may be effective in humans. PMID- 11255062 TI - Identification of free radicals produced during phacoemulsification. AB - PURPOSE: To detect, identify, and quantitate free radicals produced during conditions similar to phacoemulsification cataract surgery. SETTING: Research laboratory at the Biotechnology Center, Utah State University, Logan, Utah, USA. METHODS: All experiments were performed using a Series Ten Thousand phacoemulsifier (Alcon Laboratories) modified to make a 10 mL continuous circulation loop (to increase sensitivity). The irrigating solution was passed through a 3 mL chamber in line with the circulation loop, and electron spin resonance spin trapping methods were used to detect, identify, and quantitate free radical production during phacoemulsification. As an additional indication of hydroxyl radical production, the hydroxylation of salicylate and thiocyanate was detected by high-performance liquid chromatography and spectrophotometry, respectively. RESULTS: The hydroxyl radical was formed when phacoemulsification was performed in the presence of solutions containing spin trap in double deionized water or balanced salt solution (BSS). Hydroxyl radical production was linear with respect to phacoemulsification time. Production of the hydroxyl radical was not observed when phacoemulsification was performed with anaerobic solutions, indicating a requirement for oxygen in radical production. The concentration of trapped hydroxyl radical was reduced in the presence of balanced salt solution with bicarbonate, dextrose, and glutathione (BSS Plus). Upon phacoemulsification, both salicylate and thiocyanate underwent hydroxylation when included in the irrigating solution, confirming the generation of the hydroxyl radical. Additional tests discounted the formation of superoxide or hydrogen peroxide during phacoemulsification. CONCLUSIONS: Hydroxyl radical was produced by phacoemulsification in the presence of aerobic solutions. Hydroxyl radical production was dependent on the presence of molecular oxygen and was not generated as a result of the homolytic cleavage of water. The amount of hydroxyl radical detected was directly proportional to phacoemulsification time and was reduced in the presence of BSS Plus. Other reactive oxygen species such as superoxide, hydrogen peroxide, and ozone were not detected during phacoemulsification under these conditions. PMID- 11255063 TI - Methicillin-resistant Staphylococcus aureus keratitis after laser in situ keratomileusis. AB - A 50-year-old man had uneventful bilateral laser in situ keratomileusis (LASIK) for moderate myopia (-4.50 diopters sphere, both eyes). Twelve days postoperatively, he developed unilateral bacterial keratitis. Cultures revealed methicillin-resistant Staphylococcus aureus. The antibiotic regimen was adjusted, and he regained an uncorrected visual acuity of 20/40 and a best spectacle corrected visual acuity (BSCVA) of 20/15. Bacterial keratitis after LASIK is a rare occurrence. Aggressive use of cultures and fortified antibiotics can prevent significant loss of BSCVA, even when a resistant organism is the cause. PMID- 11255064 TI - Vitrectomy to remove a posteriorly dislocated endocapsular tension ring. AB - We treated a patient who had a posteriorly dislocated endocapsular ring associated with decreased vision and intravitreal cortical remnants. The ring was removed by uneventful pars plana vitrectomy. By the last examination, best corrected visual acuity had improved to 6/12 and intraocular pressure had stabilized to within normal limits. A posteriorly dislocated endocapsular ring is a rare complication of cataract surgery. Its removal by pars plana vitrectomy under direct observation is effective and safe. PMID- 11255066 TI - Management of atypical epicapsular star. AB - An 18-year-old woman had brown pigment deposits on the central anterior capsule of the lens associated with poor visual acuity and significant anisometropia. Surgical removal of the deposits, phacoemulsification, and intraocular lens implantation resulted in visual improvement. Epicapsular stars are formed by the confluence of multiple fine pigmentary deposits. Although they cause amblyopia, this does not appear to be dense. Surgical removal of these central deposits may improve vision, even if the patient presents as an adult. PMID- 11255065 TI - Intraocular lens implantation in a child with monocular cataract and anterior persistent hyperplastic primary vitreous. AB - A 3-year-old girl had phacoemulsification during which the presence of anterior persistent hyperplastic primary vitreous (PHPV) was discovered. Visual rehabilitation comprised contact lens use for 1 year. However, visual acuity deteriorated gradually because of secondary cataract formation. In a second surgery 1 year after the first, the posterior capsule was incised, followed by an anterior vitrectomy and intraocular lens implantation. At the last follow-up 6 months after the second surgery, there was no evidence of ocular complications and best corrected visual acuity was 0.6. PMID- 11255067 TI - Monotherapy trials in antiepileptic drugs: are modified "presurgical studies" a way out of the dilemma? AB - Monotherapy trials in epilepsy are confronted with a dilemma: either they are in conflict with ethical requirements, or they are scientifically not meaningful. Monotherapy trials, which are performed as controlled studies randomizing patients to ineffective (pseudo)placebo treatment, are incompatible with the Helsinki Declaration. On the other hand, equivalence or noninferiority studies using an active-control design do not permit valid conclusions on the efficacy of the test drug. Therefore, they do not fulfill scientific requirements for trials on new drugs. As an alternative approach, a monotherapy trial design for epilepsy patients undergoing presurgical evaluation was outlined. During presurgical evaluation, antiepileptic drugs are routinely tapered down for seizure recording. This situation is used for a placebo-controlled short-term monotherapy trial. Four trials according to this design have been completed so far. Recently, several points of concern have been raised against this design, especially for matters of ethics and external validity. In the present article, these objections are outlined and discussed. In the proposed modification the randomization and the titration of the test drug or control begins prior to the presurgical investigations. The advantages are: the test drug does not have to be titrated quickly, pure monotherapy conditions are achieved, and the subjects do not have to experience more seizures than are required for the presurgical evaluation. PMID- 11255069 TI - Idiopathic generalized epilepsies with pure grand mal: clinical data and genetics. AB - OBJECTIVE: To analyze the clinical features and family history of patients with idiopathic generalized epilepsy (IGE), with pure grand mal (GM), divided into epilepsies with GM occurring exclusively on awakening (GMA) and random GM (RGM). METHODS: We studied retrospectively 98 patients from a large epilepsy outpatient clinic. All patients had a full clinical examination and computed cerebral tomography scans (CCT) or magnetic resonance imaging (MRI) when feasible. We analyzed seizure type, seizure frequency, provocative factors, prognosis, electroencephalography (EEG) findings and family history. RESULTS: Sixty-eight patients had GMA and 30 had RGM. The mean age at seizure onset was 16.6 years (+/ 6.3 S.D., range: 5-41) and 16.7 years in those with RGM (+/-7.5 S.D., range: 4 42, NSD). Patients with GMA had a longer course of active epilepsy (median 8.5 years) compared to RGM (median 2 years). Seizure-provoking factors, especially sleep deprivation, were significantly (P=0.001) more common in patients with GMA (52/68, 77%) than in the group with RGM (13/30, 43%). Of all patients, 23% (23/98) reported first degree relatives with seizures or epilepsy. Pure GM was found in 41% (12/29) of affected first degree relatives, other idiopathic generalized epilepsy syndromes were less frequently observed (4/29, 14%). The concordance rate was high within the syndrome - none of the patients with RGM had an affected relative with GMA and vice versa only two of affected relatives of GMA patients had RGM. CONCLUSION: GMA seems to be associated with a longer duration of active epilepsy, a higher relapse rate and a stronger tendency to be precipitated by seizure provoking factors. The different concordance rates between the syndromes suggest a genetically different background. PMID- 11255068 TI - Combined influence of cyclic arousability and EEG synchrony on generalized interictal discharges within the sleep cycle. AB - PURPOSE: to analyze the activating role of cyclic alternating pattern (CAP) and EEG synchrony on generalized interictal paroxysms in the first part of the night, when all sleep patterns are represented. METHODS: nocturnal polysomnographic investigation was accomplished on a randomized series of 18 subjects with an active form of primary generalized epilepsy (PGE), but only six patients showed a complete and regular profile of the first two sleep cycles (SCs). Completeness and regularity of the selected SCs consisted in the absence of intervening wakefulness, in the presence of all sleep stages, and in the identification of three main units, (a) a descending branch, dominated by the build-up of EEG synchrony in the transition from light to deep non-rapid eye movement (NREM) sleep; (b) a trough, where the magnitude of EEG synchrony is greatest and gives rise to stages 3 and 4; (c) an ascending branch characterized by a decrease of EEG synchrony preceding the onset of rapid eye movement (REM) sleep. Generalized paroxysms were evaluated in terms of discharge rates (number of interictal bursts per minute of sleep) and distribution within the investigated sleep parameters. RESULTS: the discharge rates decreased from SC1 to SC2, with higher values quantified during NREM sleep (mean, 2.8) compared with REM sleep (mean, 0.8). Both SCs showed a progressive decrease of activation across the three units, from the highest discharge rates reached during the descending branches (mean, 3.6) to the more attenuated discharge rates during the troughs (mean, 2.4) down to the lowest rates during the ascending limbs (mean, 1.1). The magnitude of activation during the descending branches was closely related to the CAP condition (mean, 5.2) and to the powerful effect of phase A (mean, 13.9). The great majority (82%) of EEG discharges occurring in phase A were distributed within the A1 subtypes (identified by sequences of k-complexes or delta bursts). CONCLUSIONS: within the first two SCs, the features of NREM sleep endowed with the major activating power on generalized bursts are represented by the rise of EEG synchrony (descending branch) and by the A phases of CAP involved in the regulation of its build-up. PMID- 11255070 TI - No effect of 50 Hz magnetic field observed in a pilot study on pentylenetetrazol induced seizures and mortality in mice. AB - This study was planned so as to evaluate whether magnetic field exposure has any significant effect on the pentylenetetrazol (PTZ)-induced seizures. Mice were exposed to 50 Hz, 2 G (0.2 mT) magnetic field in glass cages for 1 h. Sham exposure was produced by turning off the current while the animals were in the same exposure volume. Then, PTZ was administered intraperitoneally (i.p.) at a dose of 60 mg/kg and the animals were observed for 30 min. Subsequently, the latency to seizure onset, total seizure duration, the number of seizure episodes and mortality were recorded for each subject. There was no evidence for a significant effect of the 50 Hz magnetic field on the mean number of PTZ induced seizures, seizure latency, total seizure duration and mortality (P>0.05). As a conclusion the present study failed to provide any further support for a therapeutic potential of magnetic field. PMID- 11255071 TI - Temporal lobe seizure interhemispheric propagation time depends on nonepileptic cortical cerebral blood flow. AB - In some patients with epilepsy, activation of eloquent cortex using various forms of environmental stimulation and mental activity may induce seizures. The increased neuronal activity resulting from cortical stimulation may be associated with increased regional cerebral blood flow. The vascular steal theory of temporal lobe epilepsy suggests that as nonepileptogenic cortical cerebral blood flow (CBFn) increases, temporal lobe epileptogenicity increases as a result, in part, of decreasing interhemispheric propagation time (IHPT). Recently, IHPT has been shown to be a quantitative electrocorticographic measure of temporal lobe epileptogenicity. In the current study, long-term combined subdural-EEG and surface cortical cerebral blood flow (CBF) monitoring was performed to test the hypothesis that IHPT depends upon CBFn. The results show that IHPT is a nonlinear (negative exponential) function of nonepileptic cortical CBF (r=0.507, df=32, t= 2.204, P<0.05). In temporal lobe epilepsy, nonepileptic cortical hypoperfusion may represent a protective mechanism for delaying interhemispheric seizure propagation. The fact that IHPT decreases exponentially with increasing CBFn suggests that small increases in CBFn should substantially decrease IHPT and increase epileptogenicity. This study confirms that inter-hemispheric propagation time depends upon perfusion of nonepileptogenic cortex. PMID- 11255072 TI - Alteration of GLUR2 expression in the rat brain following absence seizures induced by gamma-hydroxybutyric acid. AB - We explored the involvement of the glutamate receptor subunit B (GluR2) in the mechanism of absence seizures induced by gamma-hydroxybutyric acid (GHB). The expression and distribution of GluR2 protein in rat brain were examined during and after GHB-induced absence seizures. The data indicate that GluR2 protein expression significantly decreases following the onset of absence seizures. The suppression of GluR2 expression was prolonged and it outlasted the duration of the continuous absence seizure activity. The alteration of GluR2 protein levels was accompanied by a re-distribution of GluR2 expression from laminae V to IV in cerebral cortex. We also analyzed the duration and latency of absence seizures induced by GHB 72 h following an initial GHB-induced absence seizure, a time when suppression of GluR2 protein was maximal. The second absence seizure was significantly more prolonged than the first. These data may indicate that the putative down-regulation of GluR2 following GHB-induced absence seizure could have contributed to the potentiation of subsequent seizures in animals. A related hypothesis posed by the data is that down-regulation of GluR2 is involved in the mechanisms of the maintenance of recurrent absence seizure activity once it is initiated and therefore, may contribute to the chronicity of seizures in absence epilepsy. PMID- 11255073 TI - Reliability, validity and responsiveness of a revised scoring system for the Liverpool Seizure Severity Scale. AB - This report examines the reliability, validity and responsiveness of a revised scoring system for the Liverpool Seizure Severity Scale (LSSS). The revised scoring system was validated using archival data from an observational study and a randomized controlled study. Factor analyses confirmed that a single dimension captured how patients evaluate the severity of their most severe seizures occurring during a recall period. The revised scoring system repositions the severity score to range from 0 (no seizures) to 100 (most severe possible). Scores based on the new system were reliable, had construct validity (known groups validity), and were responsive to changes in the patients' epilepsy as noted by their physicians. Results suggest that future epilepsy studies assessing seizure severity should incorporate the revised LSSS scoring system and a modified version of the questionnaire that simplifies self-assessment and analyses. The modified version of the LSSS and its scoring system are appended to this report. PMID- 11255074 TI - Comparison the cognitive effect of anti-epileptic drugs in seizure-free children with epilepsy before and after drug withdrawal. AB - We studied the cognitive effects of antiepileptic drugs (AED), by investigating epileptic children who were seizure-free for at least 2 years and who had undergone fixed monotherapy. Seventy consecutive epileptic children (25 with carbamazepine (CBZ), 22 with phenobarbital (PB), and 23 with valproate (VPA)) were examined by Wechsler Intelligence Scale for Children-Revised (WISC-R) and auditory event-related potentials (P(300)) at three sessions: before AED reduction, then 1 and 7 months after complete withdrawal of treatment. There were no significant differences in IQ and subtests scores of WISC-R in any group at any of the three sessions. P(300) latencies were significantly increased in the children receiving PB but not in children receiving CBZ or VPA. P(300) amplitudes were increased but not significantly different among the three groups. These findings suggest that PB may affect cognitive function on children, but the changes of P(300) latencies may improve after discontinuation. PMID- 11255076 TI - The promise of monoclonal antibodies for the therapy of cancer. PMID- 11255075 TI - Inhibition of phenytoin hydroxylation in human liver microsomes by several selective serotonin re-uptake inhibitors. AB - Several case reports have indicated that the selective serotonin re-uptake inhibitor (SSRI) fluoxetine increases phenytoin blood levels when given concurrently. The mechanism of this drug-drug interaction has been attributed to inhibition of CYP2C9-catalyzed hydroxylation of phenytoin to its major oxidative metabolite in humans, para-hydroxyphenyl phenyl hydantoin (HPPH). With a bank of human liver microsomes (HLM), four SSRIs (fluoxetine, norfluoxetine, sertraline, and paroxetine) were tested for inhibition of HPPH formation. Initially, the K(m) and V(max) values of phenytoin hydroxylation to HPPH were determined in the individual HLM samples. The average K(m) (n=8) was 9.7+/-2.9 microM. The V(max) varied fivefold, with an average value of 113+/-53 pmol HPPH/min/nmol CYP450. All of the SSRIs inhibited HPPH formation; resulting Ki values were 31.1+/-10.1 microM (fluoxetine) (n=5), 51.1+/-9.4 microM (norfluoxetine) (n=3), 52.2+/-21.5 microM (sertraline) (n=3), and 80.0+/-7.2 microM (paroxetine) (n=3). Sulfaphenazole (10 microM), utilized as a positive control for inhibition of HPPH formation, inhibited phenytoin hydroxylation (>95%) in all HLM samples. Diclofenac hydroxylation to 4'-OH diclofenac, a specific marker for CYP2C9 activity, was determined in HLM1-HLM6 and was highly correlated with HPPH formation in HLM1-HLM6, indicating that phenytoin hydroxylation in human liver microsomes is largely due to CYP2C9. This work presents direct evidence that the effect of fluoxetine on phenytoin blood levels may be explained by inhibition of CYP2C9-catalyzed phenytoin hydroxylation. In light of typical SSRI blood levels observed in patients, this study also suggests that the risk of a SSRI-phenytoin interaction is highest with fluoxetine and norfluoxetine, and less likely with sertraline and paroxetine. PMID- 11255077 TI - Development of a minimally immunogenic variant of humanized anti-carcinoma monoclonal antibody CC49. AB - Monoclonal antibody (MAb) CC49 reacts with a pancarcinoma antigen, tumor associated glycoprotein (TAG)-72. To circumvent human anti-murine antibody (HAMA) responses in patients, we earlier developed a humanized CC49 (HuCC49) by grafting the complementarity-determining regions (CDRs) of MAb CC49 onto variable light (VL) and variable heavy (VH) frameworks of the human MAbs LEN and 21/28'CL, respectively. With the aim of minimizing its immunogenicity further, we have now generated a variant HuCC49 MAb by grafting the specificity-determining residues (SDRs) of MAb CC49 onto the frameworks of the human MAbs. Based on the evaluation of its binding affinity for TAG-72 and its reactivity with anti-idiotypic antibodies present in sera from patients who have been treated with murine CC49, this variant retains its antigen-binding activity and shows minimal reactivity with anti-idiotypic antibodies in patients' sera. Development of this variant, which is a potentially useful clinical reagent for diagnosis and therapy of human carcinomas, demonstrates that for humanization of a xenogeneic antibody grafting of the potential SDRs should be sufficient to retain its antigen-binding properties. PMID- 11255078 TI - Development of ABX-EGF, a fully human anti-EGF receptor monoclonal antibody, for cancer therapy. AB - Overexpression of epidermal growth factor receptor (EGFr) has been demonstrated on many human tumors, and the increase in receptor expression levels has been linked with a poor clinical prognosis. Blocking the interaction of EGFr and the growth factors could lead to the arrest of tumor growth and possibly result in tumor cell death. To this end, using XenoMouse technology, ABX-EGF, a human IgG2 monoclonal antibody (mAb) specific to human EGFr, has been generated. ABX-EGF binds EGFr with high affinity (5x10(-11) M), blocks the binding of both EGF and transforming growth factor-alpha (TGF-alpha) to various EGFr-expressing human carcinoma cell lines, and inhibits EGF-dependent tumor cell activation, including EGFr tyrosine phosphorylation, increased extracellular acidification rate, and cell proliferation. In vivo ABX-EGF prevents completely the formation of human epidermoid carcinoma A431 xenografts in athymic mice. More importantly, administration of ABX-EGF without concomitant chemotherapy results in complete eradication of established tumors. No tumor recurrence was observed for more than 8 months following the last antibody injection, further indicating complete tumor cell elimination by the antibody. Inhibition of human pancreatic, renal, breast and prostate tumor xenografts which express different levels of EGFr by ABX-EGF was also achieved. Tumor expressing more than 17000 EGFr molecules per cell showed significant growth inhibition when treated with ABX-EGF. ABX-EGF had no effect on EGFr-negative tumors. The potency of ABX-EGF in eradicating well established tumors without concomitant chemotherapy indicates its potential as a monotherapeutic agent for treatment of multiple EGFr-expressing human solid tumors, including those where no effective chemotherapy is available. Utilization of mAbs directed to growth factor receptors as cancer therapeutics has been validated recently by the tumor responses obtained from clinical trials with Herceptin, the humanized anti-HER2 antibody, in patients with HER2 overexpressing metastatic breast cancer. Being a fully human antibody, ABX-EGF is anticipated to exhibit a long serum half-life and minimal immunogenicity with repeated administration, even in immunocompetent patients. These results demonstrate the potent anti-tumor activity of ABX-EGF and its therapeutic potential for the treatment of multiple human solid tumors that overexpress EGFr. PMID- 11255079 TI - Systemic radiotherapy in metastatic breast cancer using 90Y-linked monoclonal MUC 1 antibodies. AB - Radioimmunotherapy (RIT) is a promising approach for treating metastatic breast cancer. Initial clinical trials using 131I radioimmunoconjugates, and more recent studies employing 90Y, have demonstrated objective, although transient, antitumor effects in heavily pretreated patients with minimal toxicity. Antibodies targeting unique epitopes of epithelial glycoprotein mucin (MUC-1) on breast cancer cell surfaces that have been studied in patients include BrE-3 (murine and humanized) and m170 (murine). Both antibodies react with at least 90% of breast cancers. In these and other RIT trials, myelosuppression has been the dose limiting toxicity. However, this toxicity has been successfully circumvented with the use of autologous peripheral blood stem cell transplantation, and recent clinical trials have escalated 90Y doses up to 50 mCi/m(2). The therapeutic index indicates that using these agents with stem cell support should deliver 9000 to 18000 rads to metastatic tumors. Development of improved chelates that are readily metabolized in the liver may reduce doses to this organ, projected to be next in line as dose-limiting. Combination therapy will be required to produce durable benefits in metastatic breast cancer. Low dose taxanes are synergistic with RIT in preclinical studies and when administered in the optimal sequence could sensitize tumor cells to the continuous low dose radiation delivered by RIT, without increasing toxicity. The addition of systemically administered tumor targeting radiation therapy using RIT as part of combined modality therapy may enhance the rate of complete response and, in patients with minimal metastatic disease, could lead to curative therapy. PMID- 11255080 TI - Antibody therapy for residual disease in acute myelogenous leukemia. AB - The elimination of minimal residual disease remains one of the most promising applications of monoclonal antibody (mAb)-based therapies. An early study showed that treatment with iodine-131 (131I)-labeled anti-CD33 mAb M195 had antileukemic effects when given as postremission therapy to patients with acute promyelocytic leukemia (APL) in second remission. This treatment, however, was limited by significant myelosuppression and by the development of neutralizing human antimouse antibodies. Treatment with native HuM195, a humanized version of murine M195, eliminated minimal residual disease detectable by reverse transcription polymerase chain reaction (RT-PCR) in 50% of patients. Patients with newly diagnosed APL treated with all-trans retinoic acid followed by HuM195 and consolidation chemotherapy had an 87% 3-year disease-free survival. Radioimmunotherapy with short-ranged, high-energy alpha particle-emitting isotopes may increase the potency of native mAbs while avoiding the nonspecific cytotoxicity of beta-emitting constructs. Targeted alpha particle therapy with bismuth-213-HuM195 showed significant antileukemic activity in patients with relapsed or refractory acute myelogenous leukemia. Antibody-drug conjugates, such as gemtuzumab ozogamicin, composed of a humanized anti-CD33 mAb and calicheamicin, have produced complete remissions in patients with relapsed AML and are likely to be active in the postremission setting. PMID- 11255081 TI - Preclinical and clinical data with bispecific antibodies recruiting myeloid effector cells for tumor therapy. AB - Bispecific antibodies constitute a novel approach to improve antibody efficacy. In vitro, constructs to recruit myeloid effector cells have been extensively investigated, and first animal data in human Fc receptor transgenic mice confirmed their promising therapeutic potential. Clinical experience with these constructs demonstrated acceptable toxicity, and support therapeutic efficacy in subgroups of patients. However, limited availability, unacceptable immunogenicity, and unfavorable pharmacokinetics of bispecific compared to conventional antibodies often hampered clinical studies. As solutions to these problems are available today, bispecific antibodies hold promise to improve therapeutic efficacy of antibody-based approaches in the near future. PMID- 11255082 TI - Internet pharmaco-economic studies in metastatic colorectal cancer. AB - An interactive Web site has been developed: http://smbh7.smbh.univ-paris13.fr, which uses a Markov model to calculate the management costs for metastatic colorectal cancer. This site allows drug usage costs, daily tariff costs per site, local ISA point values and the cost to the society of the chemotherapies prescribed to be recorded by cycle in a de-centralised manner. The overall cost of treatment may be calculated by one of these four units from the time when the first chemotherapy was administered until the patient has escaped from first or second line treatment. The median time to progression and the median survival time are key parameters used to calculate costs as they determine the number of patients who remain on treatment, course by course. Effectiveness results have been measured in terms of progression free survival or of global survival. Eight treatment strategies have been examined. It is possible to add new treatment regimens or new compounds into the existing pre-formatted tables. This software enables budgets to be planned depending on the regimen used and the number of patients treated. It also allows the different treatment options to be classified with respect to their incremental cost effectiveness ratio, which is defined by the additional cost of one treatment option compared to another divided by the corresponding increase in effectiveness. PMID- 11255083 TI - Differentiated thyroid carcinoma in the elderly. AB - The overall prognosis of patients with differentiated thyroid cancer is excellent, but the prognosis is rapidly worsening, when the disease is diagnosed in elderly patients. Old patients more often present with poor prognostic features, such as large tumors, follicular or Hurthle cell subtypes, extrathyroidal growth and distant metastases. Therefore, an optimal therapeutic approach is recommended. Current therapy includes a total thyroidectomy, if necessary combined with a lymph node dissection and followed by high dose radioiodine ablation. Radioiodine therapy in elderly patients meets specific problems, concerning thyroid hormone withdrawal, side effects of 131I and nursing problems. Additional treatment of residual, recurrent or metastatic disease must be tailored, according to the stage of the disease, and should not be denied on the basis of chronological age. Lifelong treatment with suppressive thyroid hormone therapy does not lead to important long-term side effects at old age. PMID- 11255086 TI - Feasibility of MRI in the diagnosis of acute diverticulitis: initial results. AB - PURPOSE: The purpose of this study was to evaluate MRI as a diagnostic tool in patients with suspected acute sigmoid diverticulitis. Furthermore, we sought to develop an optimal imaging protocol in these patients. PATIENTS AND METHODS: Eleven patients with suspected acute diverticulitis were included in the study. All patients were imaged in a 1.0 T clinical scanner using a body-array coil. Imaging sequences were single-shot TSE, HASTE-, STIR- and TrueFisp- sequence. All were obtained in the frontal plane. The diagnosis was verified by a single experienced investigator, using ultrasound, and overall clinicopathological outcome. RESULTS: MRI enabled visualization of signs of an acute diverticulitis in all patients. However, the diagnosis of acute diverticulitis was obtained in 10 patients only. The mean imaging time was 17.5+/-5.5 min. STIR- and TrueFisp sequences alone displayed all findings, e.g pericolonic exsudation, edema and segmental narrowing, whereas SSTSE and HASTE-sequences showed no additional information. Therefore, it appeared that the imaging protocol could be restricted to STIR- and TrueFisp-sequences. CONCLUSION: MRI is feasible as a fast, accurate and investigator-independent diagnostic tool in patients with suspected acute diverticulitis. To prove its value in comparison to computed tomography or ultrasound, further studies are needed. PMID- 11255087 TI - Chronic liver disease: relaxometry in the brain after liver transplantation. AB - Relaxometry revealed changes in the basal ganglia in T(1) and T(2) relaxation times due to liver disease. Manganese is probably responsible for T(1) and T(2) shortening (as the concentration is known to be higher in both the liver and blood due to hepatic cirrhosis). The aim of this study was to follow possible recovery after liver transplantation by MR relaxometry. Together with a group of 20 healthy volunteers we scanned 53 patients before and after liver transplantation (some of them repeatedly). Both T(1) and T(2) values were evaluated in the basal ganglia, thalamus, and frontal white matter. T(1) relaxation time was shortened by approx. 20-25% compared to the control group, probably the result of manganese deposition in the brain caused by hepatic cirrhosis. After liver transplantation the relaxation time recovered gradually with almost normal values reached approx. 2 years after surgery. T(1) recovery was observed in all evaluated structures. Similar results were observed with T(2) relaxation in the basal ganglia and thalamus. In the white matter T(2) remained low even 2 years after surgery. PMID- 11255088 TI - Vascular and perfusion imaging using encapsulated laser-polarized helium. AB - In this work, the use of hyperpolarized (HP) 3He for in vivo intravascular imaging on animal is reported. To overcome the problem of the low solubility of helium in blood, we propose an approach based on helium encapsulation in lipid based carrier agents. The mean diameter of the 3He microbubbles, measured equal to 3.0+/-0.2 microm, makes it possible to conduct in vivo studies. In vitro spectroscopy yielded a longitudinal relaxation time T(1) equal to 90 s and an apparent transverse relaxation time T(2)(*) of 4.5 ms. Angiographic imaging (venous and cardiac cavity visualization), as well as lung perfusion imaging, were demonstrated in rats using intravenous injections of microbubble suspensions. Suitable signal and spatial resolution were achieved. The potential of this technique for lung perfusion assessment was assessed using an experimental animal embolism model. Lung perfusion defects and recovery towards a normal perfusion state were visualized. This study was completed with the demonstration of a new ventilation-perfusion lung exploration method based entirely on HP 3He. PMID- 11255089 TI - Diffusion-weighted imaging of the spine using radial k-space trajectories. AB - INTRODUCTION: Diffusion-weighted MR imaging (DWI) of the spine requires robust imaging methods, that are insensitive to susceptibility effects caused by the transition from bone to soft tissue and motion artifacts due to breathing, swallowing, and cardiac motion. The purpose of this study was to develop a robust imaging method suitable for DWI of the spine. METHODS AND SUBJECTS: A radial k space spin echo sequence has been implemented, which is self-navigating because each acquisition line passes through the origin of k-space. Influence of cardiac motion and associated flow of cerebrospinal fluid is minimized by cardiac gating with a finger photoplethysmograph. The sequence has been tested on a 1.5T system. Diffusion-weighted images of six normal volunteers were acquired in the sagittal plane with 4 b values between 50 and 500 s mm(-2). Because of the symmetries of the cord, diffusion measurements in the head-foot (HF) or left-right (LR) directions were sufficient to measure the dominant effects of anisotropy. RESULTS: The apparent diffusion coefficients (ADCs) measured, respectively, in the LR and HF directions were (0.699+/-0.050)x10(-3) and (1.805+/-0.086)x10(-3) mm(2) s(-1) in the spinal cord, (1.588+/-0.082)x10(-3) and (1.528+/-0.052)x10(-3) mm(2) s(-1) in the intervertebral disks, and (0.346+/-0.047)x10(-3) and (0.306+/ 0.035)x10(-3) mm(2) s(-1) in the vertebrae of the cervicothoracic spine. CONCLUSION: Diffusion-weighted spin echo sequences with radial trajectories in k space provide a means of achieving robust, high quality diffusion-weighted imaging and measuring ADCs in the spine. The application of the diffusion weighting gradients in different directions allows diffusion anisotropy to be measured. PMID- 11255092 TI - Overrepresentation of point mutations in the Sp1 site of the non-coding control region of BK virus in bone marrow transplanted patients with haemorrhagic cystitis. AB - BACKGROUND: Haemorrhagic cystitis (HC) in allogeneic bone marrow transplanted (BMT) patients is associated with reactivation of BK virus (BKV) manifested as BK viruria. However, it has been suggested that BKV reactivation alone is not responsible for HC, since BKV can be detected in the urine of 50-90% of all adult BMT patients. OBJECTIVES: In the present study, we analysed if BK viruses with specific mutations in the non-coding control region (NCCR) or in the region encoding the major capsid protein (VP1) were more frequently associated to the appearance of HC in BMT patients. STUDY DESIGN: The NCCR and the region encoding VP1 of BKV excreted in the urine from 25 BMT patients, 16 with and nine without HC, were sequenced by an ABI Prism Big Dye terminator cycle sequencing ready reaction kit. RESULTS AND CONCLUSIONS: A statistically significant (P=0.019) overrepresentation of C to G mutations within the NCCR Sp1 binding site was observed in 7/16 (43%) patients with HC (six cases at position 249 (P=0.035) and one case at position 251), as compared with 0/9 (0%) of the patients without HC. Major differences were not observed in the VP1 sequences of patients with and without HC. BKV WW and WWT-variants as well as BKV subtype I were most commonly encountered in both groups of patients. In conclusion, C to G point mutations, within the BKV NCCR Sp1 binding site, were significantly more common in patients with HC, suggesting that these mutations may be indicative for the clinical diagnosis of HC and could influence the virulence of the virus. PMID- 11255093 TI - Clinical features of patients with acute respiratory illness and rhinovirus in their bronchoalveolar lavages. AB - BACKGROUND: Several reports in selected populations suggest that human rhinovirus (HRV) may be responsible for lower respiratory tract infections or pneumonia. We describe clinical features of all patients with rhinovirus cultured from their bronchoalveolar lavage (BAL) during a 10-yr period in a tertiary care center. METHODS: Results for viral culture of all lower respiratory specimens performed during a 10-year period at the University of Virginia Health Sciences Center were reviewed. A case was defined as any patient with a positive culture for HRV in a BAL specimen. A comprehensive review of the patients' medical records was performed. In one case, in situ hybridization (ISH) was performed in order to identify whether rhinoviral RNA was present in bronchial biopsy specimens. RESULTS: During the 10-year study period viruses were identified in 431 lower respiratory tract specimens, and were most frequently cytomegalovirus or herpes simplex virus. Twenty patients (ages, 2.5-86 year) had a bronchoalveolar specimen culture positive for HRV. All had an abnormal chest radiograph, 60% were admitted to the intensive care unit, and 25% expired during their hospitalization. In 18 patients (90%) various severe underlying conditions were identified including solid organ transplants in seven, malignancies in four and AIDS in two. An immunosuppressive disease or condition requiring immunosuppressive therapy was present in all cases. In addition to HRV, one or more potential pathogens were identified in respiratory specimens from 14 patients (70%). Histopathological abnormalities, ranging from fibropurulent debris in alveoli to diffuse alveolar damage, were present in 6 of 13 bronchial biopsies. In two cases without any other significant pathogens than HRV, acute inflammations with fibropurulent debris in alveoli were observed. One lung transplant patient showed intermittent recovery of HRV in her respiratory specimens during a 15-week time period, but ISH did not show HRV RNA in bronchial epithelial cells. CONCLUSION: Our observations suggest that HRV recovery from BALs or lower respiratory tract samples in highly immunocompromised patients is associated with severe lower respiratory tract illness. Whether HRV directly causes viral pneumonia or predispose to pulmonary injury and/or superinfection remains uncertain. PMID- 11255090 TI - Use of spin echo T(2) BOLD in assessment of cerebral misery perfusion at 1.5 T. AB - Inadequate blood supply relative to metabolic demand, a haemodynamic condition termed as misery perfusion, often occurs in conjunction with acute ischaemic stroke. Misery perfusion results in adaptive changes in cerebral physiology including increased cerebral blood volume (CBV) and oxygen extraction ratio (OER) to secure substrate supply for the brain. It has been suggested that the presence of misery perfusion may be an indication of reversible ischaemia, thus detection of this condition may have clinical impact in acute stroke imaging. The ability of single spin echo T(2) to detect misery perfusion in the rat brain at 1.5 T owing to its sensitivity to blood oxygenation level dependent (BOLD) contrast was studied both theoretically and experimentally. Based on the known physiology of misery perfusion, tissue morphometry and blood relaxation data, T(2) behaviour in misery perfusion was simulated. The interpretation of these computations was experimentally assessed by quantifying T(2) in a rat model for cerebral misery perfusion. CBF was quantified with the H(2) clearance method. A drop of CBF from 58+/-8 to 17+/-3 ml/100 g/min in the parieto-frontal cortex caused shortening of T(2) from 66.9+/-0.4 to 64.6+/-0.5 ms. Under these conditions, no change in diffusion MRI was detected. In contrast, the cortex with CBF of 42+/-7 ml/100 g/min showed no change in T(2). Computer simulations accurately predicted these T(2) responses. The present study shows that the acute drop of CBF by 70% causes a negative BOLD that is readily detectable by T(2) MRI at 1.5 T. Thus BOLD may serve as an index of misery perfusion thus revealing viable tissue with increased OER. PMID- 11255094 TI - Evaluation of two current generation enzyme immunoassays and an improved isolation-based assay for the rapid detection and isolation of rotavirus from stool. AB - BACKGROUND: Rapid and accurate rotavirus testing is important in decisions involving patient care and management. Quality assurance testing needs to be periodically performed, especially among widely used assays having a direct impact on patient care. OBJECTIVES: To evaluate the current generation Kallestad Pathfinder Direct antigen Detection system (PTH), and the widely used Rotaclone(R) Rotavirus EIA Diagnostic Kit (RTC), in comparison with an improved cell culture amplification-antigen detection (CCA-Ag) isolation-based assay. STUDY DESIGN: Two hundred stool specimens (specimen stored at > or =-75 degrees C), which had been previously tested by PTH, were tested by RTC and CCA-Ag. Discordant specimens were retested by PTH, blocking assay, polyacrylamide gel electrophoresis (PAGE), and/or electron microscopy (EM). RESULTS: Among 200 stool specimens, 197 were in accord by PTH, RTC and CCA-Ag. The sensitivity, specificity, positive and negative predictive values for RTC, PTH and CCA-Ag were, 100, 99, 99, 100, 100, 99, 99, 100; and 98, 100, 100, 98%, respectively. Among five initially discordant specimens, two required a period of 10 days to affect isolation. A non-cultivatable (CCA-Ag negative) but true positive specimen, was identified as rotavirus group A serotype G2 by RT-PCR. Four true positive but discordant specimens were blocking assay negative using one or both EIA kits. CONCLUSIONS: PTH and RTC are excellent rotavirus detection system. However, PTH is more expensive (ca. $3.50 vs. $2.00 per test), mandates a slightly longer turn-around time (ca. 1 vs. 1.5 h), and necessitates slightly more hands-on manipulative/preparative steps. Blocking assay was not a reliable confirmatory test for the resolution of specimen discordancy. A combination of CCA-Ag, PAGE, EM, and/or perhaps RT-PCR, is recommended as an appropriate test panel for the resolution of discordant results during assay evaluation. The newly modified and simplified 48-h rotavirus isolation-based assay may serve as a base line methodology in laboratory evalaution studies, as a laboratory support methodology during drug/vaccine efficacy trials, or for the testing of sources (e.g., biopsy/autopsy tissues) not approved for assay by commercial rotavirus kits. PMID- 11255095 TI - Prospective analysis of 61 cases of enteroviral meningitis: interest of systematic genome detection in cerebrospinal fluid irrespective of cytologic examination results. AB - BACKGROUND: Enteroviruses are the most commonly identified cause of viral meningitis. Detection of the enterovirus genome in cerebrospinal fluid (CSF) using reverse-transcription polymerase chain reaction (PCR) has proved to be useful in diagnosis and is more rapid and sensitive than viral cultures. In routine practice, cytologic examination results of CSF are obtained swiftly and PCR indication is performed as a second step. OBJECTIVES: The aim of this study was to determine, by analysis of complete data from CSF results for 61 cases of proven enteroviral meningitis, whether cytologic CSF findings can be used to establish viral etiology and to indicate if PCR assay should be performed. STUDY DESIGN: From a prospective study of children admitted during 1997 for suspected enterovirus meningitis in which PCR and viral cultures of CSF were systematically performed, we selected 61 patients with proven enterovirus meningitis. We compared global white cell count (WCC), relative percentage of lymphocytes/neutrophils, PCR and culture for enterovirus, patient age, and clinical data. RESULTS: 92% of patients (56/61) had positive PCR in CSF and in 48% (29/61) enterovirus was isolated in CSF. Nine patients (14.75%) had WCC<10/mm(3); eight of them had positive PCR and two had positive culture. There were comparable numbers of CSF with a predominance of lymphocytes (n=25) and CSF with a predominance of neutrophils (n=22), and of positive PCR and positive cultures of CSF in the two groups. Results were not influenced by the age of the patients. CONCLUSION: Irrespective of other CSF parameters, it seems difficult to dispense with PCR assay for enterovirus genome detection. It should be introduced as a true rapid routine test. Early reporting of a positive PCR result could result in a considerable saving in health resources. PMID- 11255096 TI - Application of HIV-1 genotypic-resistance testing prevents the evolution of further resistance mutations in heavily pretreated patients. AB - BACKGROUND: Resistance-associated mutations in HIV-1 evolve even under highly active antiretroviral therapy. OBJECTIVE: To evaluate the clinical efficacy of genotypic-resistance testing (GRT), to estimate the potential of a given antiretroviral therapy for prevention of further resistance mutations. STUDY DESIGN: Ten patients were treated prospectively with drugs, according to the results of a GRT. Five patients were allocated to group I in which antiretroviral therapy could be switched to an effective regimen (consisting of at least three sensitive drugs, from at least two different classes of antiretroviral substances). Five patients (group II) had no option for effective therapy, and continued to be treated non-effectively (at least one applicated substance class only intermediately sensitive, or resistant). GRT and quantitative viral cultures were performed longitudinally for 8 months. Also, plasma HIV-1 RNA, total CD4+ cells, and rates of productively infected CD4+ cells were determined. RESULTS: All the patients in group I showed a significant decrease of HIV-RNA of >1 log/ml (mean, -1.35 log/ml, P=0.025). The mean increase of CD4+ cells was 46 (not significant). The rate of productively infected CD4+ cells decreased significantly (mean, -16 productively infected CD4+ cells per 10(6) total CD4+ cells, P=0.04). In this group no further resistance mutations were detected after 8 months. In group II, none of the patients showed a significant decrease of HIV 1 RNA (mean, +0.05 log/ml), total CD4+ cells decreased (mean, -35, not significant), the rate of productively infected CD4+ cells increased significantly (mean, +124 productively infected CD4+ cells per 10(6) total CD4+ cells, P=0.04), and 4 of 5 patients had additional mutations in the RT gene conferring multi-drug resistance within 8 months (P=0.048). CONCLUSIONS: GRT is predictive of the efficacy of a therapeutic regimen, in particular regarding evolution of further resistance mutations. PMID- 11255097 TI - Congenital HCMV infection: a collaborative and comparative study of virus detection in amniotic fluid by culture and by PCR. AB - Cytomegalovirus (HCMV) infection is the leading cause of congenital virus infection in developed countries, affecting an estimated 1% of births. This antenatal infection can cause serious sequelae. Strategies for prevention and treatment must, therefore, be agreed upon, entailing a preliminary performance assessment of antenatal virus diagnosis techniques. Between 1992 and 1999, HCMV serology status was established for 19456 pregnant women in four French hospitals. Seronegative patients (55.4%) were given serology screening, and antenatal diagnosis was given to 152 women who had shown seroconversion during their pregnancies (1.4%). The detection of HCMV transmission from mother to fetus was finally established in 95 cases, using polymerase chain reaction (PCR) and viral culture methods for detecting HCMV in the amniotic fluid. These results were compared with viral culture of children's urine after birth, enabling us to distinguish between children really infected in utero (30%) and non-infected children (70%). The results of the virus culture and those of PCR were identical in 94 of the 95 cases, with one discrepancy (culture-/PCR+). The two diagnosis techniques had identical sensitivity (72%), with culture proving slightly more specific than PCR (98.4% as opposed to 96.9%). Positive prediction values for culture and for PCR were, respectively, 95.6 and 91.3%. Antenatal virus diagnosis on amniotic fluid was negative with both techniques in 8 out of 29 cases of children born with HCMV infection (VPN=89%). Over half of these wrongly negative results can be explained by amniocentesis carried out too early in the pregnancy or too early with respect to the mother's primary infection. PMID- 11255098 TI - Epstein-Barr virus (EBV) infection in infancy. AB - BACKGROUND: Epstein-Barr virus (EBV) has been shown to be the cause of infectious mononucleosis (IM) and has more complicated associations with several malignant diseases. These EBV associated diseases provide a strong incentive for the development of an EBV vaccine. Most primary EBV infection during infancy and early childhood is mild or subclinical. Little is known about its infection in infancy. The pattern of EBV serological response during infancy may be important for vaccine management. OBJECTIVES: this study has served to clarify the epidemiology and serology of primary EBV infection during early infancy. STUDY DESIGN: longitudinal serum samples from 66 Hong Kong infants were tested for EBV antibodies by immunofluorescence. Cord blood and sequential serum samples from these infants were taken at birth and then at 4-month intervals up to 2 years of age. RESULTS: maternal antibodies were present at different levels in all cord blood specimens and in serum samples of 8 infants at 4-month of age. Evidenced by VCA-IgG seroconversion, 60.6% (40/66) infants were infected during the first 2 years of life. One episode occurred before 8 months of age but, thereafter and for the remaining 16 months of follow-up until the infants were 2 years of age, the infection occurred at essentially a constant rate affecting about 20% of the remaining seronegative infants every 4 months. CONCLUSIONS: the abrupt onset of the infection after a delay of 8 months is a remarkable feature of primary EBV infection during infancy, which implicates a protective role for maternal antibodies. Persisting maternal antibodies may additionally serve to contain the infection once it occurred. This may partly explain why, unlike during adolescence, primary EBV infection early in life is usually asymptomatic. PMID- 11255099 TI - Evaluation of a pan-reactive hantavirus enzyme immunoassay and of a hantavirus immunoblot for the diagnosis of nephropathia epidemica. AB - BACKGROUND: Nephropathia epidemica (NE) caused by the hantavirus serotype Puumala (PUUV) is endemic in large parts of Europe. The prognosis of this disease is usually good. However, a rapid serological diagnosis is important to differentiate NE from potentially more severe renal conditions. OBJECTIVE: To evaluate the diagnostic usefulness of a novel pan-reactive hantavirus enzyme immunoassay (EIA) and of a novel hantavirus immunoblot (IB). STUDY DESIGN: Three groups of serum samples were tested with both assays: 79 samples from 43 patients with acute NE, 27 samples from healthy adults, and 29 tricky samples from patients with autoantibodies, with acute Epstein-Barr virus (EBV) or cytomegalovirus (CMV) infections, and from pregnant women. RESULTS: With the EIA, all but two of the early samples of the NE patients and all of the follow-up samples were positive for hantavirus IgG. All control samples were negative. The IgM EIA was positive in 42 of the 43 primary NE samples. Weak IgM EIA reactions were observed for some of the serum samples from patients with acute EBV and CMV infections. Optimal sensitivity and specificity values for the EIA were achieved when both the IgG and the IgM results were considered for the diagnosis of acute NE. All of the early NE samples reacted with the hantavirus nucleocapsid proteins in the IgG IB and all but one of these samples in the IgM IB. Cross reactions between the PUUV and the Hantaan antigens were very common. Several of the control samples did show borderline or positive bands, but these were mostly bands against only one hantavirus antigen in either the IgG or the IgM IB. The presence of at least three hantavirus bands (PUUV or HTNV) in the IgG and IgM assays was highly predictive of acute NE. CONCLUSION: Both assays were highly sensitive for the diagnosis of acute NE. However, the specificity of the IB IgM was only 76%. The specificity of both the IB and the EIA can be increased by modifications of the result interpretation. PMID- 11255100 TI - CMV gB genotypes and outcome of vertical transmission: study on dried blood spots of congenitally infected babies. AB - BACKGROUND: The role of the virulence of the infecting cytomegalovirus (CMV) strain in the transmission of the virus from mother to fetus and the outcome of the fetal infection has not received much attention yet. Molecular analysis of the gene coding for the surface glycoprotein B (gB) has been used to investigate the relationship between genotype and virulence in groups of immunosuppressed patients. OBJECTIVES: (1) to assess the prevalence of different gB genotypes in babies with congenital CMV infection; (2) to investigate the possible relationship between genotype and severity of congenital CMV disease; (3) to evaluate the possibility of using dried blood on Guthrie cards (DBS) for genotyping. STUDY DESIGN: CMV DNA was extracted from DBS and from urine/saliva samples collected in the first two weeks of life of 98 congenitally infected babies, half of which were symptomatic at birth. Genotyping was performed through RFLP analysis of the region corresponding to the cleavage site of the gB protein. RESULTS: The most prevalent genotype was gB1 (42%) followed by gB3 (26%), gB2 (19%) and gB4 (13%). Rates of disease and CNS damages were higher among children infected by gB1 (35%, 17%) and gB3 (31%, 28%) than in those infected by gB2 and gB4 (20%, 17% and 13%, 15%, respectively). These differences however did not reach the statistical significance. The parallel typing of DBS and urine/saliva strains gave a full concordance of results. CONCLUSIONS: All four major CMV gB genotypes (gB1-4) can cause a congenital infection but none seems to be associated to the development and the severity of disease. The possibility of using the neonatal DBS for genotyping opens a way to the examination of large numbers of cases of congenital CMV infection. PMID- 11255102 TI - Nucleotide divergences in the core promoter and precore region of genotype D hepatitis B virus in patients with persistently elevated or normal ALT levels. AB - BACKGROUND: Mutation in the hepatitis B virus precore codon 28, creating a translational stop codon and double 1762-1764 T/A mutations in the core promoter region, controlling the transcription of the precore RNA and the core RNA have been suggested to correlate with the HBeAg status of patients with HBV infection. OBJECTIVES: The aim of the study was to further investigate the association of nucleotide divergences in both core promoter and precore regions with liver cell injury (reflected by ALT levels) in patients with chronic HBV infection. STUDY DESIGN: The sequences of the core promoter and the precore region of HBV isolated from 67 patients, all having genotype D and subtype ayw were analyzed. The patients were divided into two groups and four subgroups according to their HBeAg and Anti-HBe status, and ALT profile. RESULTS: It was found that the nucleotide divergences in the core promoter but not in the precore region were higher in patients having persistently elevated serum ALT than in serum ALT normal patients in both HBeAg positive and Anti-HBe positive groups (P<0.05). The number of T/A and A1896 stop codon mutations did not yield a statistically significant difference between ALT normal and elevated groups. It was also found that 1762 1764 T/A and precore A 1896 mutation existed in five and six out of 29 HBeAg positive patients, respectively. In 38 anti-HBe positive patients, 1762-1764 T/A and precore A1896 mutation were detected in three and 16 patients respectively, and coexisted in 10 patients. CONCLUSIONS: Precore A 1896 stop codon mutation seems to play an essential role in the loss of HBeAg in Turkish patients. Serum viremia levels of HBV in patients having precore stop codon and/or T/A mutation were not significantly different from the other patients carrying wild type strains. Nucleotide variability in the core promoter region may be one of the factors linked to hepatitis B disease activity. PMID- 11255101 TI - Antibodies against full-length Tat protein and some low-molecular-weight Tat peptides correlate with low or undetectable viral load in HIV-1 seropositive patients. AB - BACKGROUND: The efficacy of a specific humoral response to transactivating Tat protein was studied in a group of HIV-1 seropositive drug addicts, who had previously received a similar course of anti-retroviral treatment with two reverse transcriptase inhibitors. OBJECTIVES: The aim of the study was to evaluate the meaning of an immune response to Tat protein in HIV-1 seropositive patients with different levels of HIV-1 RNA viremia. STUDY DESIGN: The study analyzed the presence of anti-Tat antibody reacting either with full-length Tat or with individual overlapping Tat-peptides (Tat(6-14), Tat(11-24), Tat(36-50), Tat(46-60), Tat(56-70) and Tat(65-80)), in a group of HIV-1 seropositive subjects with different peripheral blood viral loads. Plasma samples were examined by immunoenzymatic assay for the presence of anti-Tat IgG antibody and for the quantification of peripheral blood (plasma) viral load by branched DNA assay. RESULTS: The large majority of HIV-1 patients showed detectable levels of serum IgG to full-length-Tat, and the anti-Tat antibody level presented an inverse correlation with viral load magnitude. The analysis of antibody levels against individual overlapping Tat-peptides clearly showed that an undetectable viral load was significantly associated with the presence of a high antibody concentration against Tat(6-14), Tat(36-50) and Tat(46-60) (P=0.002, P=0.027 and P<0.001, respectively). CONCLUSION: In HIV-1-infected patients, a strong humoral immune response against HIV-1 Tat protein is inversely correlated to peripheral blood viral load and, in particular, a high level of antibody against Tat peptides containing amino acid residues 6-14 (Tat(6-14)), 36-50 (Tat(36-50)) and 46-60 (Tat(46-60)) is associated with an undetectable plasma viral load. These findings may help to tailor anti-HIV-1 Tat-containing vaccines. PMID- 11255103 TI - [3H]Ro 15-1788 binding sites to brain membrane of the saltwater Mugil cephalus. AB - The equilibrium binding parameters of the benzodiazepine antagonist [3H]Ro 15 1788 (8-fluoro-3-carboethoxy-5,6-dihydro-5-methyl-6-oxo-4H-imidazol-[1,5-a]-1,4 benzodiazepine) were evaluated in brain membranes of the saltwater teleost fish, Mugil cephalus. To test receptor subtype specificity, displacement studies were carried out by competitive binding of [3H]Ro 15-1788 against six benzodiazepine receptor ligands, flunitrazepam [5-(2-fluoro-phenyl)-1,3-dihydro-1-methyl-7-nitro 2H-1,4-benzodiazepin-2-one], alpidem [N,N-dipropyl-6-chloro-2-(4 chlorophenyl)imidazo[1,2-a]pyridine-3-acetamide], zolpidem [N,N-6 trimethyl-2-(4 methyl-phenyl)imidazo[1,2-a]pyridine-3-acetamide hemitartrate], and beta-CCM (methyl beta-carboline-3-carboxylate). Saturation studies showed that [3H]Ro 15 1788 bound saturatably, reversibly and with a high affinity to a single class of binding sites (Kd value of 1.18-1.5 nM and Bmax values of 124-1671 fmol/mg of protein, depending on brain regions). The highest concentration of benzodiazepine recognition sites labeled with [3H]Ro 15-1788 was present in the optic lobe and the olfactory bulb and the lowest concentration was found in the medulla oblongata, cerebellum and spinal cord. The rank order of displacement efficacy of unlabelled ligands observed suggested that central-type benzodiazepine receptors are present in one class of binding sites (Type I-like) in brain membranes of Mugil cephalus. Moreover, the uptake of 36Cl- into M. cephalus brain membrane vesicles was only marginally stimulated by concentrations of GABA that significantly enhanced the 36Cl- uptake into mammalian brain membrane vesicles. The results may indicate a different functional activity of the GABA-coupled chloride ionophore in the fish brain as compared with the mammalian brain. PMID- 11255104 TI - Tissue metallothionein, apoptosis and cell proliferation responses in Atlantic salmon (Salmo salar L.) parr fed elevated dietary cadmium. AB - Atlantic salmon parr were reared for 4 months on experimental diets supplemented with 0 (control), 0.5, 5, 25, 125, or 250 mg Cd x kg(-1) feed to establish a threshold concentration for dietary cadmium exposure by assessing early adaptive cellular responses. At the end of the experiment, the lowest dietary Cd concentration that caused significant accumulation in the gut, kidney and muscle was 5 mg Cd x kg(-1) compared to the control group. Over time, dietary Cd accumulated first in the gut (after 1 month), followed by the kidney (2 months), and later by muscle (4 months). Highest Cd accumulation (100-fold) was found in the gut. A significant increase in regulated cell death and proliferation in salmon fed 125 mg Cd x kg(-1) compared to control fish appeared efficient in preventing gross histopathological damage in the intestine. The highest increase in metallothionein levels was found in the kidney, and metallothionein (MT) levels increased disproportionally to Cd accumulation at increased exposure concentrations. It was concluded that MT was not directly associated with long term Cd accumulation. Atlantic salmon showed increased metallothionein levels in the kidney at a median effective concentration (concentration of dietary Cd giving 50% of the maximum increase in metallothionein, EC50) of 7 mg Cd x kg(-1), indicating toxic exposure at this concentration. PMID- 11255105 TI - Energetics in Atlantic salmon (Salmo salar L.) parr fed elevated dietary cadmium. AB - Atlantic salmon (Salmo salar L.) parr were reared for 4 months on experimental diets supplemented with Cd (0.5, 5, 25, 125, or 250 mg Cd x kg(-1)) to assess the long-term energetic changes based on the digestibility and biochemical deposition of the major dietary nutrients and to evaluate a maximum tolerable dietary toxicant concentration. Growth did not differ significantly (P>0.05) from the control groups. The biochemical composition of the carcass, but not the viscera, was negatively affected by dietary Cd exposure. The significant decreases in protein, lipid, and glycogen concentrations in the carcass (P<0.05, 25 mg x kg( 1) compared to control groups) caused a reduction in calculated whole-body energy content in fish fed 125 mg x kg(-1)compared to control groups. This reduction in calculated whole-body energy content was explained by a concurrent significant disturbance to the gastrointestinal function (measured as reduced digestibility). Only at the highest dietary Cd exposure (250 mg x kg(-1)), increased metabolic costs to cope with Cd toxicity was thought to contribute significantly to the reduction in carcass energy content. The most important factor effecting calculated total energetics was nutrient digestibility. Based on the logarithmic effective median concentration for reduced calculated energy digestibility (dietary Cd concentration corresponding to 50% reduction, EC50), the maximum tolerable dietary Cd concentration is 11 mg x kg(-1) diet. PMID- 11255106 TI - Factors involved in the (near) anoxic survival time of Cerastoderma edule: associated bacteria vs. endogenous fuel. AB - The effect of several antibiotics, molybdate and hydrogen sulfide was tested on anoxic tolerance of the cockle Cerastoderma edule, as well as utilisation of glycogen. The aim was to evaluate the role of fuel depletion and growth of bacteria as a cause of mortality. The exponential increase of sulfide and ammonium occurred in anoxic natural seawater incubations and to a lesser extend in artificial, sulfate free, seawater. This could be strongly decreased by antibacterial agents, which led to improved survival time by approximately two fold. Molybdate suppressed sulfide formation also, but did not affect survival time. Exogenous sulfide showed a negative effect on survival time at pH 6.8 and induced stronger accumulation of free glucose, D-lactate and L-alanine. This was not the case at pH 8.2. Fifty percent (LT50) of cockles in anoxic seawater died after 3.5 days still with half the initial glycogen concentration present. However, in the presence of chloramphenicol (LT50 7.9 days), the cockles utilised their endogenous fuel almost completely. In both incubations there was initially a strong increase of D-lactate and L-alanine. The D-lactate levels subsequently decreased again, probably due to bacterial consumption. This study strongly indicates that in anoxic closed systems, infection by pathogenic bacteria is the first cause of death and not exhaustion of endogenous fuel depots. PMID- 11255107 TI - Expression of metallothionein in the liver and kidney of rats is influenced by excess dietary histidine. AB - It is well known that excess dietary histidine induces the metabolic changes in copper and zinc. Therefore, this study was carried out to clarify whether excess dietary histidine alters the gene expressions of metallothionein-1 and metallothionein-2 in the liver and kidney. Male rats were fed the control (ad libitum and pair-fed) or histidine-excess (50 g of L-histidine per kg of diet) diet for 0, 1 and 3 days. The levels of liver metallothionein-1 and metallothionein-2 mRNA were markedly lower in the rats fed the histidine-excess diet as compared to those of the control (ad libitum and pair-fed) diet, when fed for 1 or 3 days. The levels of renal metallothionein-1 and metallothionein-2 mRNA in the rats fed the histidine-excess diet were higher or tended to be higher as compared with the rats fed the control (ad libitum and pair-fed) diet when fed for 1 or 3 days, respectively. At the same time, hepatic copper content was decreased and renal zinc content was increased by dietary histidine. It thus appears, that such a response on the level of liver metallothionein mRNA might be related to the contents of liver copper, but of kidney metallothionein mRNA might be due to the content of zinc. PMID- 11255108 TI - No detectable DNA excision repair in UV-exposed hepatocytes from two catfish species. AB - DNA repair is a critical process in protecting cellular genetic information from mutation. Nucleotide excision repair (NER) is a mechanism by which cells correct DNA damage caused by agents that form bulky covalent adducts and UV photoproducts such as thymine dimers and 6-4 photoproduct. NER, sometimes called dark repair, is generally accepted as being low in fish compared to mammals. This study was designed to quantitate NER in two related catfish species that have known differential sensitivities to liver carcinomas. The original hypothesis was that the more cancer resistant species, channel catfish (Ictalurus punctatus), would have more efficient DNA repair compared to the more sensitive brown bullhead (Ameriurus nebulosus). In order to measure NER, primary cultured hepatocytes of both species were exposed to UV light (10-40 J/m2) and collected at 0, 24, 48 and 72 h after exposure. Total DNA was extracted from the cells and incubated with T4 endonuclease V. Using alkaline gel electrophoresis, endonuclease sensitive sites (ESS) were quantified. Results from the ESS assay indicated there was a UV dose response increase in thymine dimers from 0 to 40 J/m2. However, no repair (decrease in number of ESS) occurred in either fish species over a 72-h time period. When cells were exposed to photoreactivating fluorescent light, repair was detected. These studies highlight the difficulty of measuring NER in fish and are consistent with the low levels of NER reported by other researchers in fish. PMID- 11255109 TI - Studies on the anti-mitogenic, anti-phage and hypotensive effects of several ribosome inactivating proteins. AB - An investigation was conducted to compare the anti-mitogenic, anti-phage and hypotensive activities of several ribosome inactivating proteins (RIPs) in order to ascertain whether the RIPs differed in their potencies in the various bioassays. Agrostin, luffin and saporin elicited a dose-dependent suppression of the mitogenic response of murine splenocytes to concanavalin A. The three RIPs were approximately equipotent in this regard, with near maximal inhibition attained at a dose of 83 nM and approximately 50% inhibition at 830 pM. Trichosanthin was slightly more potent than the three aforementioned RIPs. All of these RIPs were capable of inhibiting the replication of phage M13 in the bacterium Escherichia coli, the ranking of potencies being luffin>trichosanthin>agrostin when tested at a concentration of 3.5 microM. The RIPs gelonin and saporin did not exert a conspicuous antiviral effect at the same dose. After intravenous administration into normotensive rats via the external jugular vein, the RIPs saporin, trichosanthin, gelonin and momordin evoked a mild hypotensive response while luffin and agrostin were inactive. The hypotensive response, however, lacked dose dependence. The RIPs trichosanthin, momordin and gelonin did not affect the blood pressure response to angiotensin I. Chemical modification of the arginine residues of the RIPs brought about a reduction in their ability to inhibit cell-free translation. It appears that the ranking of potency of RIPs in one bioassay was different from the rankings in other assays. PMID- 11255110 TI - Xenobiotic metabolism in Australian marsupials. AB - The Australian marsupials are significant and unique Australian fauna. Xenobiotic metabolism is the process of enzymatic modification of xenobiotics, which include the chemicals, such as agricultural chemicals and natural dietary toxins, that these animals may be exposed to. Very little is known about the enzymes involved in xenobiotic metabolism in this unique group of animals. Folivore marsupials such as the koala (Phascolarctos cinereus and the brushtail possum (Trichosurus vulpecula) represent unique adaptation which has only been relatively superficially examined to date. We provide an overview of our current knowledge of marsupial xenobiotic metabolism. PMID- 11255111 TI - Monoamine pharmacology of the lobster cardiac ganglion. AB - Monoamine agonists and antagonists were applied to the lobster cardiac ganglion in an attempt to clarify the different actions of 5-hydroxytryptamine (5HT) and dopamine (DA) on this rhythmic pattern generator. Experiments were designed to determine whether the similar responses to 5HT and DA applied to the anterior region of the ganglion could be separated by pharmacological approaches, and whether the different responses to 5HT applied to the anterior and posterior regions of the ganglion could be attributed to mediation by different receptors. A small number of the 5HT agonists which were tested mimic the effects of 5HT, in that they increase the frequency of bursting and decrease burst duration when applied to the whole ganglion, but decrease burst frequency and increase burst duration when applied only to the posterior half. Other 5HT agonists decrease frequency and prolong bursts when applied to the whole ganglion. Of the DA agonists tested, none acts as DA itself does. Rather, they mimic the effects of 5HT applied to the posterior ganglion, by slowing bursting and prolonging bursts. The actions of agonists do not correspond in any clear way to the receptor specificities as defined in vertebrates. Most antagonists tested do not show similar specificities to their effects in vertebrates. In particular, most of the DA antagonists tested are more effective in blocking exogenous 5HT than DA. One monoamine agonist directly alters the properties of endogenous burst-organizing potentials (driver potentials) in the motorneurons of the ganglion. PMID- 11255112 TI - The effect of hydrogen peroxide on isolated body wall of the lugworm Arenicola marina (L.) at different extracellular pH levels. AB - The effect of hydrogen peroxide on the rate of tissue oxygen consumption, on intracellular pH (pH(i)) and on malondialdehyde (MDA) accumulation was studied in isolated body wall tissue of the lugworm Arenicola marina (L.). H2O2 effects were investigated at various levels of pH(i) by changing medium pH (pH(e)). The largest decrease of tissue oxygen consumption (by 17% below controls), as well as the highest degree of MDA accumulation (four-fold compared to control values) after H2O2 exposure were found at acidic pH(e) of 6.4. This was attributed to the higher redox potential of H2O2 in acidic solutions. Oxygen consumption at alkaline pH(e) (8.5) was not affected by H2O2. MDA accumulation in the tissue was considerably lower than at pH(e) 7.4 or 6.4. Despite pH dependent alterations of H2O2 redox potential, we observed more or less constant pH(e) independent acidification of the tissue upon exposure to H2O2. We attributed the acidification to an inhibition of ATP consuming proton equivalent ion transport across the cellular membrane. Inactivation of carrier proteins is discussed to be responsible for the decrease in tissue oxygen consumption. However, with a larger effect on oxygen consumption at acidic pH(e) values, the latter may not be the only explanation, but additional impairment of other energy demanding processes may be involved. PMID- 11255113 TI - Toxicity of coelomic fluid of the earthworm Eisenia foetida to vertebrates but not invertebrates: probable role of sphingomyelin. AB - The coelomic fluid (CF) of the earthworm Eisenia foetida exhibits a wide variety of biological activities. We found that the CF was not toxic to 42 species, belonging to seven invertebrate phyla, almost all in aquatic adults and larvae exposed to CF. Eleven teleostean species tested died in 0.2-1% CF mostly between 10 and 120 min and the effects were dose-dependent. Tadpoles of the toad Bufo japonicus formosus died in 0.4-2% CF between 80 and 225 min depending upon size, with larger tadpoles surviving longer. Before dying, all experimental tadpoles developed curled and shrunken tails. The Okinawa tree lizard, soft-shelled turtle, Japanese quail, mouse and rat all died after i.v. injection of CF (above 20 microl/kg). Thus, CF was not toxic to invertebrates, but toxic to vertebrates. After heating, CF lost its toxicity to fish, tadpoles and mice. Both CF and lysenin incubated with sphingomyelin-liposomes (SM-liposomes) were no longer toxic, suggesting the involvement of SM in the toxicity. Lysenin, which is a constituent of CF and known to bind specifically to sphingomyelin, exhibited toxicity similar to that of CF. Thus, lysenin in CF is probably responsible for the toxic effects of CF by binding to SM in vertebrate tissues. The bodies of invertebrates might contain little or no SM, while those of vertebrates do contain SM. The coelomic fluid of the earthworm Pheretima communissima has no toxicity to mouse. PMID- 11255114 TI - Identification of P1 and P2 purinoceptors in the aorta of the lizard (Agama sp.). AB - In the isolated Agama lizard aorta, acetylcholine (ACh; 3 nM-100 microM), noradrenaline (NA; 30 nM-0.3 mM), adrenaline (Adr; 30 nM-300 microM), adenosine 5'-triphosphate (ATP; 30 nM-1 mM), alpha,beta-methylene ATP (alpha,beta-meATP; 10 nM-10 microM), beta,gamma-methylene ATP (beta,gamma-meATP; 0.1-300 microM), 2 methylthio ATP (2-meSATP; 30 nM-30 microM) and high concentrations of uridine triphosphate (UTP; 1 microM-1 mM), all produced constriction. The P2 receptor antagonists pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS; 30 microM), suramin (0.1 mM) and Reactive blue 2 (30 microM) all raised vascular tone and could not be utilized and the antagonist 2'-O-(trinitrophenyl) ATP (TNP ATP; 0.1 microM) had no effect on responses to the ATP analogues. alpha,beta MeATP (3 microMx3) desensitised responses to alpha,beta-meATP (10 microM) and beta,gamma-meATP (0.3 mM), but not to ATP (0.3 mM) or 2-meSATP (30 microM). On pre-constricted aorta (EC50 concentration of either ACh or Adr), adenosine (1 microM-1 mM), the A1-selective agonist N6-cyclopentyl adenosine (CPA; 1-300 microM) [but not the A2- and A3-selective agonists CGS 21680 and IB-MECA respectively (both up to 30 microM)] and sodium nitroprusside (10 nM-100 microM) produced vasodilatation. Adenosine vasodilatation was antagonised by 8-p sulfophenyl-theophylline (8-pSPT; 30 microM) but not by N(omega)-nitro-L-arginine methyl ester (L-NAME; 0.1 mM). ATP (up to 0.3 mM), 2-meSATP (up to 10 microM) and UTP (up to 1 mM) were not vasodilators. In summary, A1 receptors mediating relaxation and excitatory P2X1 receptors were identified in the smooth muscle of the lizard aorta. However, in contrast to mammalian aorta, P2Y receptors on endothelial cells mediating vasodilatation via nitric oxide do not appear to be present. PMID- 11255115 TI - Prey specificity, comparative lethality and compositional differences of coral snake venoms. AB - Toxicities of crude venoms from 49 coral snake (Micrurus sp.) populations, representing 15 nominal taxa, were examined in both laboratory mice and in native prey animals and compared with data gathered from two non-micrurine elapids and a crotalid, which served as outgroups. These venoms were further compared on the basis of 23 enzymatic activities. Both toxicities and enzymatic activities were analyzed with respect to natural prey preferences, as determined from stomach content analyses and literature reports. Venoms of nearly all Micrurus for which prey preferences are known, are more toxic to natural prey than to non-prey species. Except for amphisbaenians, prey are more susceptible to venoms of Micrurus that feed upon them, than to venoms of those that eat other organisms. All venoms were more toxic i.v.>i.p.>i.m. Route-specific differences in toxicity are generally greatest for preferred prey species. Cluster analyses of venom enzymatic activities resulted in five clusters, with the fish-eating M. surinamensis more distant from other Micrurus than even the crotalid, Bothrops moojeni. Ophiophagous and amphisbaenian-eating Micrurus formed two close subclusters, one allied to the outgroup species Naja naja and the other to the fossorial, ophiophagous Bungarus multicinctus. Prey preference is shown to be the most important determinant of venom composition in Micrurus. PMID- 11255116 TI - Tissue-specific expression of two metallothionein genes in common carp during cadmium exposure and temperature shock. AB - Two metallothionein cDNA isoforms (MT-1 and MT-2) were isolated from carp (Cyprinus carpio) by RT-PCR. Sequence analysis of the cDNAs revealed two amino acid differences between the coding regions and markedly different 3' untranslated ends. Gene-specific primers were selected and used in RT-PCR reactions to measure the basal MT-1 and MT-2 mRNA levels and to follow the inducer-specific expression of MT genes in different tissues during in vivo studies. In the brain and muscle, the uninduced levels of the two MT mRNAs were similar. In the kidney and liver, the MT-1 gene product predominated, while in the heart the relative expression levels of the two genes were opposite. Both the MT-1 and MT-2 mRNA levels increased with Cd concentration in a time- and dose dependent manner. The expression of MT-2, however, was more responsive to a high Cd concentration. In parallel with the induction of the MTs by Cd, we followed the accumulation of this metal in the kidney and liver. Although the Cd level was always higher in the kidney during treatment, the rate of accumulation was higher in the liver. Cold stress resulted in a significantly higher induction of MT-1 than of MT-2, while heat shock had no effect on the expression of either gene. PMID- 11255117 TI - Biochemical and morphological changes in carp (Cyprinus carpio L.) liver following exposure to copper sulfate and tannic acid. AB - As a consequence of human activity various toxicants reach the aquatic ecosystems; humics may interact with them and may change their toxicity. Many fish are exposed to a considerable concentration of humics and pollutants. Because of paucity of data on the biochemical action of tannins in the presence of the fungicide CuSO4 a comparative study was undertaken. The alterations of redox-parameters in carp liver were monitored and tissue necrosis was followed by measuring the plasma transaminase activities and by electron microscopy. Tannic acid, a representative phenolic/humic compound, exerted prooxidant effects in carp, which may be partially due to formation of prooxidant intermediates/end products via its biotransformation. Alternatively, tannic acid may partially inhibit the antioxidant enzymes of fish. The response to CuSO4 was more severe. Although tannic acid alone acted as a prooxidant in fish, electron micrographs demonstrated that it reduced the necrotizing effect of copper, which may be due to the complexing activity of tannic acid with the biomolecules of the hepatocytes and to the H2O2-degrading activity of tannin-CuSO4 combination. Our results indicate that the heavy metal-detoxifying capacity of tannin may be significant; however, tannin-exposure alone or combined with metals may be toxic for fish due to enzyme inhibition and oxidative stress induction. PMID- 11255118 TI - The physiological role of the creatine kinase system: evolution of views. AB - The development of ideas concerning the buffer and transport functions of the creatine kinase system is described. The concept of ATP compartmentation at sites of its production and utilization is critically analyzed. Kinetic, thermodynamic, and structural data used as a basis for a hypothesis on the structural and functional coupling of mitochondrial creatine kinase and adenine nucleotide translocase are comprehensively analyzed, and experimental evidence inconsistent with this hypothesis is presented. It seems that the mitochondrial creatine kinase may serve to equilibrate ADP concentration in the intermembrane space with fluctuating ADP concentrations in the cytoplasm. It is suggested that creatine kinase molecules bound to other intracellular structures (e.g., to myofibrils) may equilibrate local ADP concentrations with those present in the cytoplasm. PMID- 11255119 TI - Matrix metalloproteinases of normal human tissues. AB - This review considers biochemical properties of the family of matrix metalloproteinases (MMPs) of normal human tissues and the involvement of these enzymes in morphogenesis. Four main MMP subfamilies are characterized, and a group of other MMPs is described. Data on mechanisms of activation and inhibition of MMPs in certain tissues during various physiological processes (embryogenesis, angiogenesis, tissue growth and involution) are considered. Information about tissue inhibitors of MMP is presented, and the ability of these inhibitors to regulate the activity of MMPs is analyzed. PMID- 11255120 TI - Expression and properties of bacteriophage T4 gene product 11. AB - A plasmid vector for expression of bacteriophage T4 gene product 11 (gp11) in E. coli cells has been constructed. Gp11 is a baseplate protein that connects short tail fibers providing irreversible adsorption of the virus on a cell. A method based on chromatography on hydroxyapatite has been developed for purification of recombinant gp11. The protein is active in an in vitro complementation assay and transforms defective phage particles lacking gp11 into infective ones. Gel filtration data suggest that the biologically active protein is a trimer. According to CD spectroscopy and sequence analysis data, the polypeptide chain of gp11 contains not less than 20% alpha-helical segments, about 30% beta-structure, and belongs to the class of alpha/beta structural proteins. PMID- 11255121 TI - Two exopolyphosphatases of the cytosol of the yeast S. cerevisiae: comparative characteristics. AB - In cell-free extracts of the yeast Saccharomyces cerevisiae that had been transferred from phosphate-deficient (-P) medium to complete (+P) medium ("hypercompensation" conditions), the specific and the total polyphosphatase activities increased (by 50 and 60%, respectively) compared to the control that was transferred from (+P) medium to (+P) medium. Specific and total polyphosphatase activities under "hypercompensation" conditions increased by 25 and 43% in cytosol, by 33 and 100% in vacuoles, and by 50 and 50% in the total membrane fraction, respectively. In contrast, the polyphosphatase activity in the cell envelope somewhat decreased compared to the control. Under the growth conditions indicated above, a novel high molecular weight polyphosphatase was revealed in the cytosol fraction along with the previously studied 40-kD polyphosphatase. Unlike the 40-kD polyphosphatase, which is most active with tripolyphosphate, this novel enzyme has a molecular mass of more than 440 kD and is most active with high molecular weight polyphosphates. This polyphosphatase is insensitive to antibodies that suppress the activity of the 40-kD polyphosphatase of the cytosol. In a number of properties, the high molecular weight polyphosphatase of the cytosol resembles the polyphosphatase of vacuoles, but it differs from the polyphosphatases of nuclei and mitochondria of S. cerevisiae. The ratio of the low and high molecular weight polyphosphatases depends on the culture growth conditions. Under "hypercompensation" conditions, the total activity of the high molecular weight polyphosphatase in the cytosol is five times higher than that of the 40-kD polyphosphatase. During growth without re inoculation, the 40-kD polyphosphatase is predominant in the cytosol; its total activity in dependence on the growth stage is 3.5-12.5 times higher than the activity of the high molecular weight form. PMID- 11255122 TI - Macroscopic aggregation of tobacco mosaic virus coat protein. AB - The relationship between processes of thermal denaturation and heat-induced aggregation of tobacco mosaic virus (TMV) coat protein (CP) was studied. Judging from differential scanning calorimetry "melting" curves, TMV CP in the form of a trimer-pentamer mixture ("4S-protein") has very low thermal stability, with a transition temperature at about 40 degrees C. Thermally denatured TMV CP displayed high propensity for large (macroscopic) aggregate formation. TMV CP macroscopic aggregation was strongly dependent on the protein concentration and solution ionic strength. By varying phosphate buffer molarity, it was possible to merge or to separate the denaturation and aggregation processes. Using far-UV CD spectroscopy, it was found that on thermal denaturation TMV CP subunits are converted into an intermediate that retains about half of its initial alpha-helix content and possesses high heat stability. We suppose that this stable thermal denaturation intermediate is directly responsible for the formation of TMV CP macroscopic aggregates. PMID- 11255124 TI - Influence of ethanol and phosphatidylethanol on the activity of pancreatic phospholipase A2. AB - Hydrolysis of dioleoylphosphatidylethanol (DOPEt) and dioleoylphosphatidylcholine (DOPC) catalyzed by phospholipase A2 (PLA2) from porcine pancreas has been studied in single-component and binary liposomes in the absence and in the presence of ethanol. DOPEt (an anionic phospholipid) was found to increase the rate of hydrolysis of zwitterionic DOPC in liposomes under the action of PLA2. PMID- 11255123 TI - Natural hidden autoantibodies react with negatively charged carbohydrates and xenoantigen Bdi. AB - Immunoglobulin preparations from sera of healthy donors contain polyspecific autoantibodies interacting with DNA and other charged antigens. These antibodies belong to the IgG class and can exist in the free or hidden state. The hidden antibody activity can be revealed after ion-exchange chromatography on QAE Sephadex A-50. Immunoenzyme assay was used to assess the interactions of both free and hidden antibodies with different carbohydrates. The hidden antibodies were only able to interact with different polyanionic carbohydrates and neutral xenoantigen Bdi. PMID- 11255125 TI - Participation of free radicals in photoreduction of protochlorophyllide to chlorophyllide in an artificial pigment-protein complex. AB - The primary stages of protochlorophyllide phototransformation in an artificially formed complex containing heterologously expressed photoenzyme protochlorophyllide-oxidoreductase (POR), protochlorophyllide, and NADPH were investigated by optical and ESR spectroscopy. An ESR signal (g = 2.002; H = 1 mT) appeared after illumination of the complex with intense white light at 77 K. The ESR signal appeared with simultaneous quenching of the initial protochlorophyllide fluorescence, this being due to the formation of a primary non-fluorescent intermediate. The ESR signal disappeared on raising the temperature to 253 K, and a new fluorescence maximum at 695 nm belonging to chlorophyllide simultaneously appeared. The data show that the mechanism of protochlorophyllide photoreduction in the complex is practically identical to the in vivo mechanism: this includes the formation of a short-lived non-fluorescent free radical that is transformed into chlorophyllide in a dark reaction. PMID- 11255126 TI - Swelling of root cell walls as an indicator of their functional state. AB - The swelling capacity of cell walls isolated from different parts of lupine root was investigated. The water content in fragments of intact roots (Q) and swelling coefficient of standardized samples of cell walls (Kcw) were determined, and the dependences of Q and Kcw on the distance from the root tip (L) were plotted. It was shown that the change in Q value along the stretch of the lupine root reaches its maximum at distances of 1.5-6 cm or 7-12 cm from the root tip in 7-day-old and 14-day-old seedlings, respectively, whereas the Kcw value distribution over the root length is virtually invariable. In the radial direction, both the Q and Kcw values in cortex tissues are about twice higher than in the central cylinder. In our opinion, the changes of both Q and Kcw in the radial direction are associated with different degrees of cross-linking between polymer chains in cell wall structures of root cortex and central cylinder. The results of measurement of the Kcw value are consistent with the widely accepted mechanisms of water transport in roots in the radial direction. These data show that water transport through apoplast to the border between the cortex and central cylinder is accompanied by an increase in the resistance to water flow. Among other factors, this increase is due to a greater degree of cross-linking between cell wall polymers in the central cylinder. The results of measurement of the swelling coefficient of standardized cell wall samples in water and in 10 mM KCl at different pH values show that the swelling capacity of root cell walls varies according to the physicochemical properties of synthetic ion exchangers. Cell walls shrink (cell wall volume decreases) as ion concentration in solution increases and pH decreases. This causes an increase in the hydraulic resistance (or a decrease in the hydraulic conductivity) of apoplast. It was concluded that swelling is determined by the physicochemical properties of the cell wall, whereas the change in the swelling capacity induced by variation of external or internal conditions is an element of the mechanism of regulation of volume water flow in roots. PMID- 11255128 TI - Biosynthesis of chlorophyll of photosystem II reaction centers in plant leaves in early stages of etiolation. AB - Spectral methods were used to study the sequences of chlorophyll biosynthesis reactions in etiolated pea, bean, and maize plants in early stages (3-4 days) of growth. For these juvenile plants, along with the reaction chain known for mature (7-9 day-old) plants, a new reaction chain was found which started with phototransformation of the long-wavelength form PChld 686/676 into PChld 653/648. (PChld 653/648 differs from the main known precursor form PChld 655/650). The subsequent photoreduction of PChld 653/648 leads to the formation of Chld 684/676, which is transformed into Chl 688/680 in the course of a dark reaction. After completion of this reaction, fast (20-30 sec) quenching of the fluorescence of the reaction product is observed with the formation of non-fluorescent Chl 680. The reaction accompanied by pigment fluorescence quenching is absent in pea mutants with depressed function of Photosystem II reaction centers. This suggests that the newly found reaction chain leads to the formation of chlorophyll of the Photosystem II reaction center. PMID- 11255127 TI - Proteins of brown seaweeds as inhibitors of endo-1-->3-beta-D-glucanases of marine invertebrates. AB - It has been found that aqueous-ethanol extracts of brown seaweeds contain substances inhibiting endo-1-->3-beta-D-glucanases, the digestive enzymes of marine mollusks. The inhibitors were detected in 14 of 21 brown seaweeds investigated. An irreversible protein inhibitor possessing high specificity toward endo-1-->3-beta-D-glucanases of marine mollusks was isolated from the brown seaweed Laminaria cichorioides. As determined by gel filtration, the molecular mass of the inhibitor is 46 kD. The value of [I]50 (10(-8) M) for the inhibitor is comparable with the corresponding value for natural inhibitors of amylases from terrestrial plants. The results of chemical modification indicated that tryptophan, glutamic acid, aspartic acid, histidine, and probably tyrosine residues are important for the interaction of the inhibitor with the enzyme. PMID- 11255129 TI - Phosphorylation of the protein encoded by the first open reading frame of the MDG4 transposable element (gypsy) by homologous and heterologous casein kinases type 2. AB - Homogeneous casein kinase type 2 (CK2) was obtained from oocytes of Rana temporaria and cells of Drosophila melanogaster by chromatography on heparin Sepharose, phosphocellulose, and Mono Q columns using a Pharmacia FPLC system. The procedure was first successfully used for the purification of CK2 from the Drosophila melanogaster cell culture. It has been shown that the protein encoded by the first open reading frame (ORF) of the gypsy transposable element (MDG4) is an effective protein substrate both for homologous and heterologous CK2 from the oocytes of Rana temporaria in vitro. Both enzymes catalyze the incorporation of two moles of phosphate per mole of protein. The Km and Vmax values for the reaction catalyzed by CK2 from the Drosophila cell culture were 32.5 +/- 2.1 nM and 70.97 +/- 1.89 nmol/min per microg, respectively, and for CK2 from oocytes, these values were 37.6 +/- 2.8 nM and 66.02 +/- 2.15 nmol/min per microg, respectively. PMID- 11255130 TI - Charge separation in photosynthetic reaction centers under femtosecond excitation. AB - The process of electron transfer from the primary electron donor P* to the primary electron acceptor BA in the reaction center of Rhodobacter sphaeroides R 26 under 30 fsec pulse excitation was studied in this work with the aim of establishing a relationship between the nuclear subsystem motion and charge transfer. For this purpose the fsec and psec oscillations in the bands of stimulated emission of P* and in the band of reaction product BA- at 1020 nm were investigated. It was established that the reversible formation of the P+BA- state is characterized by two vibration modes (130 and 320 cm(-1)) and connected with an arrival of the wavepacket induced by fsec excitation to the intersection of potential surfaces P*BA and P+BA-. The irreversible formation of the P+BA- state with the time constant of 3 psec is followed by oscillations with frequencies of 9 and 33 cm(-1). These results show that the irreversibility of electron transfer is determined by two factors: 1) by a difference between the energy width of the wavepacket and the gap between the named surfaces; 2) by a difference between the duration of wavepacket residence near the intersection of the surfaces and the relaxation time of the P+BA- state. PMID- 11255131 TI - Role of glutathione-dependent peroxidase in regulation of lipoperoxide utilization in malignant tumors. AB - Glutathione content, the activity of glutathione-dependent enzymes (glutathione reductase, glutathione peroxidase, and glutathione S-transferase), and also SOD (superoxide dismutase) and catalase were studied in human malignant tumors (uterus, breast, and ovaries) and normal tissues. Glutathione level and the activity of glutathione-dependent enzymes were 2-3 times higher in the malignant tumors than in normal tissues. A negative correlation between the level of glutathione and glutathione-dependent enzymes (glutathione peroxidase and glutathione S-transferase) in tumors and the efficacy of postoperative chemotherapy may characterize the degree of tumor resistance to chemotherapy and therefore may have prognostic value. Low SOD and catalase activity and high activity of glutathione-dependent enzymes in tumors suggest that glutathione peroxidase and glutathione S-transferase play a major role in peroxide utilization in malignant tumors. PMID- 11255132 TI - Identification of DNA polymerase delta in eggs of a teleost fish (loach). AB - DNA polymerase found in an extract from eggs of the teleost fish Misgurnus fossilis (loach) has been identified as an enzyme of the delta type. The enzyme was purified 4000- to 5000-fold from the extract by liquid chromatography. The DNA polymerase activity was sensitive to the inhibiting action of aphidicolin but resistant to N2-(p-n-butylphenyl)-2'-deoxyguanosine 5'-triphosphate (BuPdGTP). The enzyme activity correlates with the presence of a polypeptide with molecular mass of 120-130 kD that interacts specifically with polyclonal antibodies against calf thymus DNA polymerase delta as revealed by Western blotting and is presumably the catalytic subunit of the enzyme. The loach DNA polymerase possesses the 3'-->5'-exonuclease activity specific to single-stranded DNA and catalyzes distributive elongation of primers in primer-template complexes. PMID- 11255133 TI - The relationship of mode and intensity of training on resting metabolic rate in women. AB - The present cross-sectional study was designed to investigate the relationship between exercise training and resting metabolic rate (RMR). The focus of this investigation was to compare RMR in aerobically trained (AT), resistance trained (RT), and untrained (UNT) women. Subjects were also classified as highly trained (HT), moderately trained (MT), or untrained (UNT) in order to examine the relationship between RMR and level of training. Sixty-one women between the ages of 18 and 46 years volunteered to serve as subjects in this study. Each subject completed measurements of body composition, maximal oxygen uptake (VO2max), and two consecutive measurements of RMR. The data presented show that there was no significant difference in resting metabolic rate between resistance-trained, aerobically trained, and control subjects. However, when grouped by intensity of training, there was a trend for an increased resting metabolic rate (kcal/day) in the highly trained subjects, regardless of mode of training. PMID- 11255134 TI - The effects of a 20-week exercise training program on resting metabolic rate in previously sedentary, moderately obese women. AB - The present study was designed to investigate the effects of exercise training on resting metabolic rate (RMR) in moderately obese women. It was hypothesized that exercise training would increase resting metabolic rate. Nineteen previously sedentary, moderately obese women (age = 38.0 +/- 0.9 years, percent body fat = 37.5 +/- 0.8) trained for 20 weeks using either resistance training (RT) or a combination of resistance training and walking (RT/W). The high intensity resistance-training program was designed to increase strength and fat-free mass and the walking program to increase aerobic capacity. There was also a non exercising control group (C) of 9 subjects in this study. Fat-free mass was significantly increased in both the RT (+1.90 kg) and RT/W (+1.90 kg) groups as a result of the training program. No group showed significant changes in fat mass or relative body fat from pre- to post-training. Aerobic capacity was slightly, though significantly, increased in the RT/W group only. The RT group showed a significant increase (+44 kcal x day(-1)), while the RT/W group showed a significant decrease (-53 kcal x day(-1)) in resting metabolic rate post training. RT can potentiate an increase in RMR through an increase in fat-free mass, and the decrease in RMR in the RT/W group may have been a result of heat acclimation from the walk training. PMID- 11255135 TI - The assessment of frequency of iron deficiency in athletes from the transferrin receptor-ferritin index. AB - The transferrin receptor-ferritin index (sTfR/logFerr) was determined in 131 male and 121 female athletes in order to assess the frequency of iron deficiency (threshold value of that index taken as 1.8). Blood was drawn for determining morphological indices as well as sTfR, ferritin, iron, total iron binding capacity (TIBC), and haptoglobin. A significantly (p <.01) higher incidence of iron deficiency was observed in women (26%) than in men (11%). The iron deficiency was latent, since no subject was found to be anemic. The plasma iron was significantly lower and TIBC higher (p <.001) in both iron-deficient subgroups than in the non-deficient ones. This confirmed the latent character of iron deficiency. Some hematological indices (Hb, MCH, MCHC, MCV) were significantly lower in iron-deficient female athletes than in male athletes, which suggested a more profound iron deficiency in the former. The sTfR/logFerr index might thus be useful in detecting iron deficiency in athletes, especially in those with erythropoiesis disorders, since physical loads may affect the widely used ferritin levels. PMID- 11255136 TI - Effect of potassium phosphate supplementation on perceptual and physiological responses to maximal graded exercise. AB - This investigation evaluated the effect of oral potassium phosphate supplementation on ratings of perceived exertion (RPE) and physiological responses during maximal graded exercise tests (GXT). Eight highly trained endurance runners completed a GXT to anchor the Borg 15-point RPE scale and two double-blind counterbalanced GXTs. Subjects ingested either 4,000 mg x day(-1) of phosphate (PHOS) or a placebo (PLA) for 2 days. Two weeks separated GXTs. Phosphate levels obtained immediately prior to the GXTs were greater in PHOS than PLA. No differences between PHOS and PLA were noted for the submaximal and maximal physiological responses. RPE for the overall body were lower during PHOS than PLA at intensities corresponding to 70-80% of VO2max. This suggests that oral potassium phosphate supplementation mediates perceived exertion during moderately intense exercise. PMID- 11255137 TI - Effectiveness of glycerol as a rehydrating agent. AB - On two occasions, 8 male subjects completed a dehydration protocol, immediately followed by a 180-min rehydration protocol, then a subsequent exercise bout. During each dehydration session, subjects lost 3.1 +/- 0.4% body weight (BW) following discontinuous exercise in the heat (40 degreesC, 33% rh). During the first 30 min of rehydration, subjects ingested either 1.0-g glycerol x kg body weight(-1) + 30% of the total rehydration water volume (GLY), or 30% of the total rehydration water volume without glycerol (CON). The five remaining ingestions (every 30 min) were equal to 14% of the remaining fluid volume and were identical in nature. Fluid volume ingested equaled fluid volume lost during dehydration. Following the 180 min rehydration period, subjects cycled (appoximately 50% VO2 peak) in the heat (40 degrees C, 33% rh) until volitional exhaustion. Three observations were made: (a) Following glycerol-induced rehydration, time to volitional exhaustion was greater during the subsequent exercise bout in the heat (CON: 38.0 +/- 2.0, GLY 42.8 +/- 1.0 min, p <.05); (b) glycerol-induced rehydration significantly increased plasma volume restoration within 60 min and at the end of the 180-min rehydration period; and (c) total urine volume was lower and percent rehydration was greater following GLY, but neither was significantly different. PMID- 11255138 TI - Blood glucose responses to carbohydrate feeding prior to exercise in the heat: effects of hypohydration and rehydration. AB - This study assessed the plasma glucose (PG) and hormonal responses to carbohydrate ingestion, prior to exercise in the heat, in a hypohydrated state versus partial rehydration with intravenous solutions. On separate days, 8 subjects (21.0 +/- 1.8 years; 57.3 +/- 3.7 ml x kg(-1) x min(-1)) exercised at 50% VO2max in a 33 degree C environment until a 4% body weight loss was achieved. Following this, subjects were rehydrated (25 ml x kg(-1)) with either: 0.45% IV saline (45IV), 0.9% IV saline (9IV), or no fluid (NF). Subjects then ingested 1 g x kg(-1) of carbohydrate and underwent an exercise test (treadmill walking, 50% VO2max, 36 degrees C) for up to 90 min. Compared to pre-exercise level (294 mg x dl(-1)), PG increased significantly (>124 mg x dl(-1)) at 15 min of the exercise test in all trials and remained significantly elevated for 75 min in NF, 30 min more than in the 2 rehydration trials. Although serum Insulin increased significantly at 15 min of exercise in the 45IV trial (7.2 +/- 1.2 vs. 23.7 +/- 4.7 mIU x ml(-1)), no significant differences between trials were observed. Peak plasma norepinephrine was significantly higher in NF (640 +/- 66 pg x ml(-1)) compared to the 45IV and 9IV trials (472 +/- 55 and 474 +/- 52 pg x ml(-1), respectively). In conclusion, ingestion of a small solid carbohydrate load prior to exercise in the 4% hypohydration level resulted in prolonged high PG concentration compared to partial IV rehydration. PMID- 11255139 TI - Regulation of skeletal muscle amino acid metabolism during exercise. AB - The contribution of amino acid oxidation to total energy expenditure is negligible during short-term intense exercise and accounts for 3-6% of the total adenosine triphosphate supplied during prolonged exercise in humans. While not quantitatively important in terms of energy supply, the intermediary metabolism of several amino acids-notably glutamate, alanine, and the branched-chain amino acids-affects other metabolites, including the intermediates within the tricarboxylic acid (TCA) cycle. Glutamate appears to be a key substrate for the rapid increase in muscle TCA cycle intermediates (TCAI) that occurs at the onset of moderate to intense exercise, due to a rightward shift of the reaction catalyzed by alanine aminotransferase (glutamate + pyruvate <==> alanine + 2 oxoglutarate). The pool of muscle TCAI declines during prolonged exercise, and this has been attributed to an increase in leucine oxidation that relies on one of the TCAI. However, this mechanism does not appear to be quantitatively important due of the relatively low maximal activity of branched-chain oxoacid dehydrogenase, the key enzyme involved. It has been suggested that an increase in TCAI is necessary to attain high rates of aerobic energy production and that a decline in TCAI may be a causative factor in local muscle fatigue. These topics remain controversial, but recent evidence suggests that changes in TCAI during exercise are unrelated to oxidative energy provision in skeletal muscle. PMID- 11255140 TI - Exercise, protein metabolism, and muscle growth. AB - Exercise has a profound effect on muscle growth, which can occur only if muscle protein synthesis exceeds muscle protein breakdown; there must be a positive muscle protein balance. Resistance exercise improves muscle protein balance, but, in the absence of food intake, the balance remains negative (i.e., catabolic). The response of muscle protein metabolism to a resistance exercise bout lasts for 24-48 hours; thus, the interaction between protein metabolism and any meals consumed in this period will determine the impact of the diet on muscle hypertrophy. Amino acid availability is an important regulator of muscle protein metabolism. The interaction of postexercise metabolic processes and increased amino acid availability maximizes the stimulation of muscle protein synthesis and results in even greater muscle anabolism than when dietary amino acids are not present. Hormones, especially insulin and testosterone, have important roles as regulators of muscle protein synthesis and muscle hypertrophy. Following exercise, insulin has only a permissive role on muscle protein synthesis, but it appears to inhibit the increase in muscle protein breakdown. Ingestion of only small amounts of amino acids, combined with carbohydrates, can transiently increase muscle protein anabolism, but it has yet to be determined if these transient responses translate into an appreciable increase in muscle mass over a prolonged training period. PMID- 11255142 TI - Message from the Program Committee Chair. PMID- 11255141 TI - Amino acids and endurance exercise. AB - Although skeletal muscle is capable of oxidizing selected amino acids, exercise in the fed and carbohydrate-replete condition results in only a small increase in amino acid utilization. Nevertheless, it may be important to increase the dietary protein requirements of active individuals. There is ongoing debate as to whether the amino acids for oxidation are derived from the free amino acid pool, from net protein breakdown, or a combination of both. There has been interest in the potential ergogenic benefits of amino acid ingestion; however, BCAA ingestion does not appear to affect fatigue during prolonged exercise, there is little support from controlled studies to recommend glutamine ingestion for enhanced immune function, and although glutamine stimulates muscle glycogen synthesis, its addition to carbohydrate supplements provides no additional benefit over ingestion of carbohydrate alone. PMID- 11255143 TI - The conference at a glance. PMID- 11255145 TI - Conference Schedule. PMID- 11255144 TI - Westin Crown Center Hotel meeting rooms. PMID- 11255146 TI - AAPA poster and presentation schedule. PMID- 11255148 TI - Evaluation of the interaction between biodegradation and sorption of phenanthrene in soil-slurry systems. AB - This work develops and utilizes a non-steady-state model for evaluating the interactions between sorption and biodegradation of hydrophobic organic compounds in soil-slurry systems. The model includes sorption/desorption of a target compound, its utilization by microorganisms as a primary substrate existing in the dissolved phase, and/or the sorbed phase in biomass and soil, oxygen transfer, and oxygen utilization as an electron acceptor. Biodegradation tests with phenanthrene were conducted in liquid and soil-slurry systems. The soil slurry tests were performed with very different mass transfer rates: fast mass transfer in a flask test at 150 rpm, and slow mass transfer in a roller-bottle test at 2 rpm. The results of liquid tests indicate that biodegradation of the soil-soluble organic fraction did not significantly enhance the biodegradation rate. In the slurry tests, phenanthrene was degraded more rapidly than in liquid tests, but at a similar rate in both slurry systems. Modeling analyses with several hypotheses indicate that a model without biodegradation of compound sorbed to the soil was not able to account for the rapid degradation of phenanthrene, particularly in the roller-bottle slurry test. The model with sorbed-phase biodegradation and the same biokinetic parameters, but unique mass transfer coefficients, simulated the experimental data in both slurry tests most successfully. Reduced mass transfer resistance to bacteria attached to the soil is the most likely phenomenon accounting for rapid sorbed-phase biodegradation. PMID- 11255149 TI - High-rate continuous production of lactic acid by Lactobacillus rhamnosus in a two-stage membrane cell-recycle bioreactor. AB - It is important to produce L(+)-lactic acid at the lowest cost possible for lactic acid to become a candidate monomer material for promising biodegradable polylactic acid. In an effort to develop a high-rate bioreactor that provides high productivity along with a high concentration of lactic acid, the performance of membrane cell-recycle bioreactor (MCRB) was investigated via experimental studies and simulation optimization. Due to greatly increased cell density, high lactic acid productivity, 21.6 g L(-1) h(-1), was obtained in the reactor. The lactic acid concentration, however, could not be increased higher than 83 g/L. When an additional continuous stirred tank reactor (CSTR) was attached next to the MCRB a higher lactic acid concentration of 87 g/L was produced at significant productivity expense. When the two MCRBs were connected in series, 92 g/L lactic acid could be produced with a productivity of 57 g L(-1) h(-1), the highest productivity among the reports of L(+)-lactic acid that obtained lactic acid concentration higher than 85 g/L using glucose substrate. Additionally, the investigation of lactic acid fermentation kinetics resulted in a successful model that represents the characteristics of lactic acid fermentation by Lactobacillus rhamnosus. The model was found to be applicable to most of the existing data with MCRBs and was in good agreement with Levenspiel's product-inhibition model, and the Luedeking-Piret equation for product-formation kinetics appeared to be effective in representing the fermentation kinetics. There was a distinctive difference in the production potential of cells (cell-density-related parameter in Luedeking-Piret equation) as lactic acid concentration increases over 55 g/L, and this finding led to a more precise estimation of bioreactor performance. PMID- 11255150 TI - Utilization of a palladium-metal oxide semiconductor (Pd-MOS) sensor for on-line monitoring of dissolved hydrogen in anaerobic digestion. AB - The use of a hydrogen-sensitive palladium-metal oxide semiconductor (Pd-MOS) sensor in combination with a membrane for liquid-to-gas transfer for the detection of dissolved hydrogen was investigated. The system was evaluated with known concentrations of dissolved hydrogen in water. The lowest concentration detected with this set-up was 160 nM. The method was applied to monitoring of a laboratory-scale anaerobic digestion process employing mixed sludge containing mainly food/industrial waste. Pulse loads of glucose were added to the system at different levels of microbial activity, and the microbial status of the culture was reflected in the dissolved hydrogen response. Simultaneous headspace hydrogen measurements were performed, and at the lower levels of dissolved hydrogen no corresponding headspace hydrogen could be detected. When glucose was added to a resting culture the dissolved hydrogen response was rapid and the first response could be detected 9 min after addition of glucose, whereas headspace hydrogen concentrations increased only after 80 to 110 min. This indicates limitations in the liquid-to-gas hydrogen transfer and illustrates the importance of hydrogen monitoring in the liquid. The sensor system developed is flexible, the membrane is easily replaceable, and the probe for liquid-to-gas hydrogen transfer can be adjusted easily to large-scale applications. PMID- 11255147 TI - Fractionation of soybean proteins with pressurized carbon dioxide as a volatile electrolyte. AB - Fractionation of specific proteins from plant material is a complex and involved science, yet pure protein extracts are in high demand by a wide range of food and pharmaceutical industries. In this study carbon dioxide has been used as a volatile electrolyte to isoelectrically precipitate two major protein constituents of soybean. Carbon dioxide was shown to be effective in purifying glycinin and beta-conglycinin in a three-step process as 95% and 80% concentrated fractions with precipitation yields of 28% and 21%, respectively. Recycling of the mixed precipitate of the intermediary step enables complete separation into the concentrated fractions. Fractionation acidity was precisely controlled by a simple modification of pressure. In addition, the occurrence of a pH overshoot was prevented at any point in the fractionation vessel, as the pH minimum was defined by its equilibrium relationship with carbon dioxide operating pressure. The removal of the glycinin precipitate was an important factor in the purification procedure. The yield of the individual concentrated glycinin and beta-conglycinin precipitate fractions was a function of carbon dioxide pressure, extract concentration and, to a much lesser extent, temperature. PMID- 11255151 TI - Effect of mass transfer limitations on the enzymatic kinetic resolution of epoxides in a two-liquid-phase system. AB - Optically active epoxides can be obtained by kinetic resolution of racemic mixtures using enantioselective epoxide hydrolases. To increase the productivity of the conversion of sparingly aqueous soluble epoxides, we investigated the use of a two-phase aqueous/organic system. A kinetic model which takes into account interphase mass transfer, enzymatic reaction, and enzyme inactivation was developed to describe epoxide conversion in the system by the epoxide hydrolase from Agrobacterium radiobacter. A Lewis cell was used to determine model parameters and results from resolutions carried out in the Lewis cell were compared to model predictions to validate the model. It was found that n-octane is a biocompatible immiscible solvent suitable for use as the organic phase. Good agreement between the model predictions and experimental data was found when the enzyme inactivation rate was fitted. Simulations showed that mass transfer limitations have to be avoided in order to maximize the yield of enantiomerically pure epoxide. Resolution of a 39 g/L solution of racemic styrene oxide in octane was successfully carried out in an emulsion batch reactor to obtain (S)-styrene oxide in high enantiomeric excess (>95% e.e.) with a yield of 30%. PMID- 11255152 TI - Using historical data for bioprocess optimization: modeling wine characteristics using artificial neural networks and archived process information. AB - Optimization of fermentation processes is a difficult task that relies on an understanding of the complex effects of processing inputs on productivity and quality outputs. Because of the complexity of these biological systems, traditional optimization methods utilizing mathematical models and statistically designed experiments are less effective, especially on a production scale. At the same time, information is being collected on a regular basis during the course of normal manufacturing and process development that is rarely fully utilized. We are developing an optimization method in which historical process data is used to train an artificial neural network for correlation of processing inputs and outputs. Subsequently, an optimization routine is used in conjunction with the trained neural network to find optimal processing conditions given the desired product characteristics and any constraints on inputs. Wine processing is being used as a case study for this work. Using data from wine produced in our pilot winery over the past 3 years, we have demonstrated that trained neural networks can be used successfully to predict the yeast-fermentation kinetics, as well as chemical and sensory properties of the finished wine, based solely on the properties of the grapes and the intended processing. To accomplish this, a hybrid neural network training method, Stop Training with Validation (STV), has been developed to find the most desirable neural network architecture and training level. As industrial historical data will not be evenly spaced over the entire possible search space, we have also investigated the ability of the trained neural networks to interpolate and extrapolate with data not used during training. Because a company will utilize its own existing process data for this method, the result of this work will be a general fermentation optimization method that can be applied to fermentation processes to improve quality and productivity. PMID- 11255153 TI - Determination of the toxicity of several aromatic carbonylic compounds and their reduced derivatives on Phanerochaete chrysosporium using a Pseudomonas putida test system. AB - We tested four aromatic carbonylic compounds and their corresponding reduced derivatives, possible substrates, and products of a biotransformation for toxicity against the white-rot fungus Phanerochaete chrysosporium. The bacterium Pseudomonas putida, which has been proven to be a good test organism for investigating toxic effects, was used as a primary screen. For both P. chrysosporium and P. putida, all ketones showed a higher toxicity than their corresponding alcohol derivatives. Within one chemical group a direct correlation between the hydrophobicity (logP values) of the compounds and their toxicity could be observed. Furthermore, all tested compounds also caused an isomerization of cis to trans unsaturated fatty acids in P. putida, a mechanism of this bacterium to adapt its membrane to toxic environmental influences. Toxicity of aromatic carbonylic compounds in an established biotransformation system with P. chrysosporium can be estimated by calculating the corresponding logP values of the substrates and potential products. P. putida can be used to test the toxicity of aromatic ketones to the basic diomycete P. chrysosporium. PMID- 11255155 TI - Chemo-enzymatic D-enantiomerization of DL-lactate. AB - We investigated the total conversion of racemic lactate, L-lactate, and pyruvate into D-lactate, which is very useful as a starting material for the synthesis of chiral compounds and much more valuable than the L-enantiomer by means of coupling of L-specific oxidation of the racemate with L-lactate oxidase and non enantiospecific reduction of pyruvate to DL-lactate with sodium borohydride. In this one-pot system, L-lactate was enantiospecifically oxidized to an achiral product, pyruvate, which was chemically reduced to DL-lactate leading to a turnover. Consequently, either DL-lactate, L-lactate, or pyruvate was fully converted to the D-enantiomer. We optimized the reaction conditions: DL-lactate was converted to D-lactate in 99% of the theoretical yield and with more than 99% enantiomeric excess. DL-alpha-Hydroxybutyrate and alpha-ketobutyrate were converted also to D-alpha-hydroxybutyrate in the same way, though slowly. PMID- 11255154 TI - Human insulin from a precursor overexpressed in the methylotrophic yeast Pichia pastoris and a simple procedure for purifying the expression product. AB - The methylotrophic yeast Pichia pastoris, which proved successful in producing many heterologous proteins, was used to express an insulin precursor. A transformant with a high copy number of the gene integrated into the chromosome was obtained by the dot-blotting method. In high-density fermentation using a simple culture medium composed mainly of salt and methanol, the expression level reached 1.5 g/L. A simple two-step method was established to purify the expression product from the culture medium with an overall recovery of about 80%. After tryptic transpeptidation, human insulin with full receptor binding capacity and biological activity was obtained. In the presence of zinc, the recombinant human insulin could be crystallized in the rhombohedral form. PMID- 11255156 TI - Glycogen metabolism in aerobic mixed cultures. AB - In this study, the metabolism of glycogen storage and consumption in mixed cultures under aerobic conditions is described. The experimental results are used to calibrate a metabolic model, which as sole stoichiometric variables has the efficiency of oxidative phosphorylation (delta) and maintenance requirement in units of adenosine triphosphate (m(ATP)). Using the experimental data and values from the literature we show that delta and m(ATP) are strongly coupled and that the values determined for glycogen and poly-beta-hydroxybutyrate (PHB) metabolism are similar. We also demonstrate that storage of glycogen and subsequent growth occur without significant loss of energy, as compared with direct growth on glucose. For kinetic modeling, Monod kinetics is used most commonly in activated sludge models to describe the rate of microbial transformation. Monod kinetics, however, does not provide a good description of the data obtained. Second-order kinetics gives a better description of the rate of glycogen degradation. Formation and consumption of glycogen appears to be much faster than for PHB. PMID- 11255157 TI - High-cell-density fermentation for production of L-N-carbamoylase using an expression system based on the Escherichia coli rhaBAD promoter. AB - A high-cell-density fed-batch fermentation for the production of heterologous proteins in Escherichia coli was developed using the positively regulated Escherichia coli rhaBAD promoter. The expression system was improved by reducing of the amount of expensive L-rhamnose necessary for induction of the rhamnose promoter and by increasing the vector stability. Consumption of the inducer L rhamnose was inhibited by inactivation of L-rhamnulose kinase encoding gene rhaB of Escherichia coli W3110, responsible for the first irreversible step in rhamnose catabolism. Plasmid instability caused by multimerization of the expression vector in the recombination-proficient W3110 was prevented by insertion of the multimer resolution site cer from the ColE1 plasmid into the vector. Fermentation experiments with the optimized system resulted in the production of 100 g x L(-1) cell dry weight and 3.8 g x L(-1) of recombinant L-N carbamoylase, an enzyme, which is needed for the production of enantiomeric pure amino acids in a two-step reaction from hydantoins. PMID- 11255158 TI - Kinetics of the lipase-catalyzed synthesis of glucose esters in acetone. AB - A simple kinetic model derived from a ping-pong bi-bi mechanism is proposed to describe the lipase-catalyzed esterification of glucose with fatty acids. The mathematical expressions derived from this model have been tested using several sets of data obtained from reactions carried out under different reaction conditions. The predicted values provide very good fits of the experimental data for temperatures from 30 to 60 degrees C, enzyme loadings from 90 to 180 mg, and fatty acid concentrations from 0.33M to 1M. Experiments conducted at different temperatures permit one to estimate an activation energy of approximately 12 kcal/mol for the rate-limiting step of the reaction (formation of the acyl-enzyme complex). The model also considers the kinetics of inactivation of the biocatalyst during the reaction. PMID- 11255159 TI - Cell separation mediated by differential rolling adhesion. AB - Recently, we showed a correlation between the maturity of hematopoietic stem and progenitor cells during development and rolling efficiency on selectins. These findings motivated us to explore a novel separation that exploits differences in selectin-mediated rolling adhesion between populations of cells. We extend the use of a previously developed cell-free system to study the separation of populations of sialyl Lewis x (sLe(x))-coated microspheres designed to roll with different average velocities on L-selectin chimeric substrates under well-defined flow. Results show that a separation that exploits differences in average rolling velocities between cell or microsphere populations is attainable. Excellent recovery and purity values for the slower rolling, or more desirable, populations are obtained and can be estimated from rolling velocity measurements. We also assess the feasibility of a selectin-mediated separation of adult bone marrow cell populations using previously obtained rolling velocity and rolling flux data for CD34+ and CD34- adult bone marrow cells on L-selectin substrates. We believe that a cell separation mediated by differential rolling adhesion can be used to enrich populations of hematopoietic stem and progenitor cells from an adult bone marrow cell preparation and that this method possesses several major advantages over existing antibody-mediated cell-affinity chromatography technologies. PMID- 11255160 TI - Kinetics and mass-transfer phenomena in anaerobic granular sludge. AB - The kinetic properties of acetate-degrading methanogenic granular sludge of different mean diameters were assessed at different up-flow velocities (V(up)). Using this approach, the influence of internal and external mass transfer could be estimated. First, the apparent Monod constant (K(S)) for each data set was calculated by means of a curve-fitting procedure. The experimental results revealed that variations in the V(up) did not affect the apparent K(S)-value, indicating that external mass-transport resistance normally can be neglected. With regard to the granule size, a clear increase in K(S) was found at increasing granule diameters. The experimental data were further used to validate a dynamic mathematical biofilm model. The biofilm model was able to describe reaction diffusion kinetics in anaerobic granules, using a single value for the effective diffusion coefficient in the granules. This suggests that biogas formation did not influence the diffusion-rates in the granular biomass. PMID- 11255161 TI - Preparation, stability, and in vitro performance of vesicles made with diblock copolymers. AB - Vesicles made completely from diblock copolymers-polymersomes-can be stably prepared by a wide range of techniques common to liposomes. Processes such as film rehydration, sonication, and extrusion can generate many-micron giants as well as monodisperse, approximately 100 nm vesicles of PEO-PEE (polyethyleneoxide polyethylethylene) or PEO-PBD (polyethyleneoxide-polybutadiene). These thick walled vesicles of polymer can encapsulate macromolecules just as liposomes can but, unlike many pure liposome systems, these polymersomes exhibit no in-surface thermal transitions and a subpopulation even survive autoclaving. Suspension in blood plasma has no immediate ill-effect on vesicle stability, and neither adhesion nor stimulation of phagocytes are apparent when giant polymersomes are held in direct, protracted contact. Proliferating cells, in addition, are unaffected when cultured for an extended time with an excess of polymersomes. The effects are consistent with the steric stabilization that PEG-lipid can impart to liposomes, but the present single-component polymersomes are far more stable mechanically and are not limited by PEG-driven micellization. The results potentiate a broad new class of technologically useful, polymer-based vesicles. PMID- 11255162 TI - Evolution of a recombinant (gucoamylase-producing) strain of Fusarium venenatum A3/5 in chemostat culture. AB - Fusarium venenatum JeRS 325 is a transformant of strain A3/5 which produces Aspergillus niger glucoamylase (GAM) under the control of a Fusarium oxysporum trypsin-like protease promoter. The evolution of JeRS 325 was studied in glucose limited chemostat cultures grown on NaNO3 or (NH4)2SO4 as the nitrogen source. Thirteen mutants which were more highly branched and four mutants which were more sparsely branched than the parental strain were isolated from the NaNO3 chemostat. The highly branched mutants detected in this chemostat did not displace the sparsely branched population. The mutants isolated from the NaNO3 chemostat complemented representative strains previously isolated from glucose limited chemostat cultures of F. venenatum A3/5 grown on (NH4)2SO4, but showed little complementation between themselves. By contrast, a highly branched mutant isolated from the (NH4)2SO4 chemostat culture displaced the sparsely branched mycelial population. None of the mutants isolated from the NaNO3 or (NH4)2SO4 chemostats produced as much GAM as JeRS 325. Southern blot analysis showed that all except one mutant had lost copies of both the glucoamylase and the acetamidase (the selectable marker) genes. However, specific GAM production was not necessarily correlated with the extent of glaA gene loss observed. Further, 10 of the mutants had lost the ability to grow on acetamide as the sole nitrogen source, although they retained copies of the amdS gene. In competition studies, mutants which could not utilize acetamide displaced mutants which could. The presence of foreign DNA in JeRS 325 resulted in a reduced specific growth rate (compared to A3/5), but the presence of the foreign DNA did not prevent the evolution of the strain or the isolation of mutants which had improved growth rates. PMID- 11255163 TI - Activity of different Candida antarctica lipase B formulations in organic solvents. AB - The activity of different formulations of Candida antarctica lipase B (CALB), such as crude CALB, purified CALB, purified CALB lyophilized with PEG (CALB + PEG) or oleic acid (CALB + OA), and the commercial formulation Novozym 435, was determined in toluene, carbon tetrachloride, and 1,4-dioxane at various water activities (a(w)). The reaction between vinylacetate and 1-octanol was used as the model reaction and both transesterification (formation of 1-octylacetate) and hydrolytic (formation of acetic acid from vinylacetate) activities were determined. For equal amounts of lipase protein, CALB + PEG (and to a lesser extent CALB + OA) displayed higher activity than that of the other formulations; for instance, in toluene (a(w) < 0.1), it was 260-, 13-, and 1.8-fold more active than crude CALB, purified CALB, and Novozym 435, respectively. Moreover, the transesterification activity of CALB + PEG was of the same order of magnitude (51%) of the activity shown by the enzyme in the hydrolysis of vinylacetate in aqueous buffer. These results suggest that PEG and oleic acid could act as lyoprotectants, preventing the formation of intermolecular interactions during the lyophilization process that might be responsible for protein denaturation. No diffusional limitation was observed for CALB + PEG-catalyzed reactions. Purified CALB, in contrast to the other formulations, showed a marked activity increase (2.1 to 7.8-fold) as a function of a(w) and, in 1,4-dioxane, it was 3.5-fold more active when it was added to the solvent after previous dissolution of the lyophilized powder in water. PMID- 11255164 TI - Gas phase biotransformation reaction catalyzed by baker's yeast. AB - The gas phase continuous production of acetaldehyde from ethanol and hexanol from hexanal using dried baker's yeast was studied as an alternative approach to conventional processes. The effects of water activity, activity of substrates, and amount of yeast on the performance of the continuous bioreactor were investigated. The extent of yeast hydration and ethanol activity are the most important factors affecting yeast activity and stability. PMID- 11255165 TI - Biodegradable polymeric scaffolds for musculoskeletal tissue engineering. AB - Biodegradable scaffolds have played an important role in a number of tissue engineering attempts over the past decade. The goal of this review article is to provide a brief overview of some of the important issues related to scaffolds fabricated from synthetic biodegradable polymers. Various types of such materials are available; some are commercialized and others are still in the laboratories. The properties of the most common of these polymers are discussed here. A variety of fabrication techniques were developed to fashion polymeric materials into porous scaffolds, and a selection of these is presented. The very important issue of scaffold architecture, including the topic of porosity and permeability, is discussed. Other areas such as cell growth on scaffolds, surface modification, scaffold mechanics, and the release of growths factors are also reviewed. A summary outlining the common themes in scaffold-related science that are found in the literature is presented. PMID- 11255166 TI - Gene-expression profiling of human osteoblasts following treatment with the ionic products of Bioglass 45S5 dissolution. AB - The effect of the ionic products of Bioglass 45S5 dissolution on the gene expression profile of human osteoblasts was investigated by cDNA microarray analysis of 1,176 genes. Treatment with the ionic products of Bioglass 45S5 dissolution increased the levels of 60 transcripts twofold or more and reduced the levels of five transcripts to one-half or less than in control. Markedly up regulated genes included RCL, a c-myc responsive growth related gene, cell cycle regulators such as G1/S specific cyclin D1, and apoptosis regulators including calpain and defender against cell death (DAD1). Other significantly up-regulated genes included the cell surface receptors CD44 and integrin beta1, and various extracellular matrix regulators including metalloproteinases-2 and -4 and their inhibitors TIMP-1 and TIMP-2. The identification of differentially expressed genes by cDNA microarray analysis has offered new insights into the mode of action of bioactive glasses and has proven to be an effective tool in evaluating their osteoproductive properties. PMID- 11255167 TI - In-depth morphological changes and embrittlement near the wear surface of UHMWPE inserts from uncemented hip systems. AB - Polyethylene hip cups were examined with optical and electron microscopy after permanganic etching, a technique that allows in-depth examination from the articulating surface downward. In addition to wear features present on the surface, novel defects were revealed in implants after retrieval from the body but not in as-manufactured controls. They were incipient cracks that indicated the existence of an embrittled layer extending 10 microm or more into the implant from the wear surface after exposure to the body environment. The lengths of the cracks, which were perpendicular to the tensile stresses responsible for their formation, were mostly more or less parallel to the wear surface. The embrittlement and cracking revealed are probably major contributors to the wear of polyethylene implants in the body. Poor particle consolidation may be a contributory factor, but it was not observed to be the primary cause of implant wear within the body. PMID- 11255168 TI - Effects of apatite and wollastonite containing glass-ceramic powder and two types of alumina powder in composites on osteoblastic differentiation of bone marrow cells. AB - Previously we developed a composite consisting of apatite and wollastonite containing glass-ceramic (AW-GC) powder and bisphenol-a-glycidyldimethacrylate (Bis-GMA)-based resin (designated AWC), and demonstrated that AWC showed direct contact with living bone. Another new composite consisting of mainly the delta crystal phase of alumina bead powder and Bis-GMA-based resin (designated ABC) was developed. Although alumina ceramics are bioinert and a composite filled with the pure alpha-crystal phase of alumina powder (designated alphaALC) did not allow direct bone formation in vivo, ABC was shown to have excellent osteoconductivity. One purpose of this study was to investigate whether AW-GC powder in a composite promotes osteoblastic differentiation of rat bone marrow cells as AW-GC bulk did. Another purpose was to evaluate the effects of the delta-crystal phase of alumina powder in a composite on osteoblastic differentiation. In a cell culture with dexamethasone, alkaline phosphatase (AP) activity at both days 7 and 14, and the levels of osteocalcin mRNA and alpha1(I) collagen mRNA at day 14 and osteopontin mRNA at day 7, were highest on AWC, followed by ABC, and finally alphaALC. Scanning electron microscopy showed more abundant mineralized globules and a fibrous collagen matrix on AWC at day 14, followed by ABC. In a cell culture without dexamethasone, AP activity at both days 7 and 14, and the level of osteopontin mRNA at day 7, were higher on ABC than on any other composite, whereas osteocalcin mRNA could not be detected. These results indicate that AW-GC powder in a composite promotes osteoblastic differentiation of bone marrow cells intensively when supplemented with dexamethasone. The delta-crystal phase of alumina powder in a composite promotes greater osteoblastic differentiation than the alpha-crystal phase of alumina powder. PMID- 11255169 TI - Effect of polymethylmethacrylate particles on mature bone in the optical bone chamber. AB - Reaction of mature bone and its vasculature to 3.33 +/- 0.19 microm polymethylmethacrylate (PMMA) particles at a concentration of 2.5 x 10(8)/cc was measured using optical bone chamber implant intravital microscopy. Twelve adult female New Zealand White rabbits were divided into six receiving Healon alone (controls) and six receiving Healon plus PMMA. The particles were introduced to the bone chamber compartment after removing its overlying optical element, which was immediately reinstalled. Reaction was monitored weekly over a 6-week period using video and photographic imaging. Bone was labeled before treatment with oxytetracycline and after treatment with alizarin complexone. Perfusing blood was labeled with fluorescein isothiocyanate dextran-70 kDa (FITC-D70). Parameters measured were net bone resorption, from black and white images, bone turnover, from color images, vascularity, and average vessel caliber. Neither bone turnover nor vessel caliber were significantly affected at the p < or = 0.050 level over time. In contrast, bone resorption was significantly greater and vascularity significantly less in the presence of PMMA. It was inferred that any differences in bone turnover were masked by resorption of new bone. It was concluded that the lack of a PMMA effect on average vessel caliber meant that the vascularity effects were not due to angiogenesis, but to vessel recruitment (or its opposite), an effect more consistent with inflammation than repair. The lack of vascularity increase in PMMA-treated compartments also suggested that increased resorption was a local phenomenon, because blood supply had not increased to provide the extra osteoclasts required for observed net bone loss. PMID- 11255170 TI - An X-ray photoelectron spectroscopy study of the process of apatite formation on bioactive titanium metal. AB - Bioactive titanium metal, prepared by treatment with NaOH followed by an annealing stage to form a sodium titanate layer with a graded structure on its surface, forms a biologically active bone-like apatite layer on its surface in the body, and bonds to bone through this apatite layer. In this study, process of apatite formation on the bioactive titanium metal in a simulated body fluid was investigated using X-ray photoelectron spectroscopy. The bioactive titanium metal formed Ti-OH groups soon after soaking in the simulated body fluid, via the exchange of the Na(+) ions in the sodium titanate on its surface with H(3)O(+) ions in the fluid. The Ti-OH groups on the metal combined with the calcium ions in the fluid immediately to form a calcium titanate. After a long period, the calcium titanate on the metal took the phosphate ions as well as the calcium ions in the fluid to form the apatite nuclei. The apatite nuclei then proceeded to grow by consuming the calcium and phosphate ions in the fluid. These results indicate that the Ti-OH groups formed on the metal induce the apatite nucleation indirectly, by forming a calcium titanate. The initial formation mechanism of the calcium titanate may be attributable to the electrostatic interaction of the negatively charged Ti-OH groups with the positively charged calcium ions. PMID- 11255171 TI - Three-dimensional printing and porous metallic surfaces: a new orthopedic application. AB - As-cast, porous surfaced CoCr implants were tested for bone interfacial shear strength in a canine transcortical model. Three-dimensional printing (3DP) was used to create complex molds with a dimensional resolution of 175 microm. 3DP is a solid freeform fabrication technique that can generate ceramic pieces by printing binder onto a bed of ceramic powder. A printhead is rastered across the powder, building a monolithic mold, layer by layer. Using these 3DP molds, surfaces can be textured "as-cast," eliminating the need for additional processing as with commercially available sintered beads or wire mesh surfaces. Three experimental textures were fabricated, each consisting of a surface layer and deep layer with distinct individual porosities. The surface layer ranged from a porosity of 38% (Surface Y) to 67% (Surface Z), whereas the deep layer ranged from 39% (Surface Z) to 63% (Surface Y). An intermediate texture was fabricated that consisted of 43% porosity in both surface and deep layers (Surface X). Control surfaces were commercial sintered beaded coatings with a nominal porosity of 37%. A well-documented canine transcortical implant model was utilized to evaluate these experimental surfaces. In this model, five cylindrical implants were placed in transverse bicortical defects in each femur of purpose bred coonhounds. A Latin Square technique was used to randomize the experimental implants left to right and proximal to distal within a given animal and among animals. Each experimental site was paired with a porous coated control site located at the same level in the contralateral limb. Thus, for each of the three time periods (6, 12, and 26 weeks) five dogs were utilized, yielding a total of 24 experimental sites and 24 matched pair control sites. At each time period, mechanical push-out tests were used to evaluate interfacial shear strength. Other specimens were subjected to histomorphometric analysis. Macrotexture Z, with the highest surface porosity, failed at a significantly higher shear stress (p = 0.05) than the porous coated controls at 26 weeks. It is postulated that an increased volume of ingrown bone, resulting from a combination of high surface porosity and a high percentage of ingrowth, was responsible for the observed improvement in strength. Macrotextures X and Y also had significantly greater bone ingrowth than the controls (p = 0.05 at 26 weeks), and displayed, on average, greater interfacial shear strengths than controls, although they were not statistically significant. PMID- 11255172 TI - Mechanical properties and cell cultural response of polycaprolactone scaffolds designed and fabricated via fused deposition modeling. AB - A number of different processing techniques have been developed to design and fabricate three-dimensional (3D) scaffolds for tissue-engineering applications. The imperfection of the current techniques has encouraged the use of a rapid prototyping technology known as fused deposition modeling (FDM). Our results show that FDM allows the design and fabrication of highly reproducible bioresorbable 3D scaffolds with a fully interconnected pore network. The mechanical properties and in vitro biocompatibility of polycaprolactone scaffolds with a porosity of 61 +/- 1% and two matrix architectures were studied. The honeycomb-like pores had a size falling within the range of 360 x 430 x 620 microm. The scaffolds with a 0/60/120 degrees lay-down pattern had a compressive stiffness and a 1% offset yield strength in air of 41.9 +/- 3.5 and 3.1 +/- 0.1 MPa, respectively, and a compressive stiffness and a 1% offset yield strength in simulated physiological conditions (a saline solution at 37 degrees C) of 29.4 +/- 4.0 and 2.3 +/- 0.2 MPa, respectively. In comparison, the scaffolds with a 0/72/144/36/108 degrees lay-down pattern had a compressive stiffness and a 1% offset yield strength in air of 20.2 +/- 1.7 and 2.4 +/- 0.1 MPa, respectively, and a compressive stiffness and a 1% offset yield strength in simulated physiological conditions (a saline solution at 37 degrees C) of 21.5 +/- 2.9 and 2.0 +/- 0.2 MPa, respectively. Statistical analysis confirmed that the five-angle scaffolds had significantly lower stiffness and 1% offset yield strengths under compression loading than those with a three-angle pattern under both testing conditions (p < or = 0.05). The obtained stress-strain curves for both scaffold architectures demonstrate the typical behavior of a honeycomb structure undergoing deformation. In vitro studies were conducted with primary human fibroblasts and periosteal cells. Light, environmental scanning electron, and confocal laser microscopy as well as immunohistochemistry showed cell proliferation and extracellular matrix production on the polycaprolactone surface in the 1st culturing week. Over a period of 3-4 weeks in a culture, the fully interconnected scaffold architecture was completely 3D-filled by cellular tissue. Our cell culture study shows that fibroblasts and osteoblast-like cells can proliferate, differentiate, and produce a cellular tissue in an entirely interconnected 3D polycaprolactone matrix. PMID- 11255173 TI - In vivo response to biodegradable controlled antibiotic release systems. AB - In this study, the major goal was to evaluate in vitro and in vivo findings by macroscopy, radiology, and histology to determine the effectiveness of therapy of experimental implant-related osteomyelitis with antibiotic carrier rods constructed of microbial polyesters. The polymers used were poly(3 hydroxybutyrate-co-4-hydroxyvalerate) [P(3-HB-co-4-HB)] and poly(3 hydroxybutyrate-co-3-hydroxy- valerate) [P(3-HB-co-3-HV)]. Both the Sulperazone and the Duocid-P(3-HB-co-4-HB) rods with a drug to polymer ratio of 1:1 (w/w) were effective in treating the bone infection that was experimentally initiated by inoculation of a hemolytic strain of Staphylococcus aureus (coagulase positive; phage type 52/52b) together with metal implants into the medullary area of rabbit tibia. Macroscopical data revealed that the effectiveness of therapy was apparent at week 6 for all categories tested. Radiological findings with Duocid- and Sulperazone-loaded P(3-HB-co-4-HB) rods improved significantly when judged by changes in periosteal elevation, widening of bone shaft, new bone formation, and soft-tissue deformation after 6 weeks of implantation. Histologically the signs of infection were found to subside by weeks 3 and 6. Inflammatory cells were replaced with bone-forming cells upon treatment with Sulperazone-P(3-HB-co-4-HB) and Duocid-P(3-HB-co-4-HB). Osteoblastic activity was prominent. Intramedullary inflammation, although still present, started to be replaced by fibrous or bony tissue. Histological findings presented the subsidence of infection. In summary, the antibiotic-loaded biopolymeric rods appeared to have potential as a new controlled-release system for the treatment of implant related osteomyelitis and chronic osteomyelitis. PMID- 11255174 TI - Selective differentiation of mammalian bone marrow stromal cells cultured on three-dimensional polymer foams. AB - Bone marrow stromal cells (BMSC) are pluripotent progenitor cells that can regenerate different skeletal tissues in response to environmental signals. In this study, we used highly porous, structurally stable three-dimensional polymer foams in conjunction with specific regulatory molecules to selectively differentiate mammalian BMSC into either cartilaginous or bone-like tissues. Bovine BMSC were expanded in monolayers and cultured on 5-mm-diameter, 2-mm-thick foams made of poly(lactic-co-glycolic acid) and poly(ethylene glycol). Constructs maintained their original size and shape for up to 4 weeks of culture and supported BMSC growth and production of extracellular matrix (ECM). By proper use of chondrogenic (dexamethasone, insulin, transforming growth factor-beta1) or osteogenic (dexamethasone, beta-glycerophosphate) medium supplements, we could control whether the generated ECM was cartilaginous (containing collagen type II and sulfated glycosaminoglycans) or bone-like (containing osteocalcin, osteonectin, and mineralized foci). After 4 weeks of cultivation, cartilaginous and bone-like ECM were uniformly distributed throughout the construct volume and respectively represented 34.2 +/- 9.3% and 12.6 +/- 3.2% of the total available area. BMSC culture on poly(lactic-co-glycolic acid)/poly(ethylene glycol) foams provides a three-dimensional model system to study the development of mesenchymal tissues in vitro and has potential applications in engineering autologous grafts for skeletal tissue repair. PMID- 11255175 TI - Compressive stress-relaxation of human atherosclerotic plaque. AB - Knowledge of the mechanical properties of human atherosclerotic plaque is fundamental to understanding atherosclerosis and its treatment. Data are scant, however, particularly with respect to the time-dependent nature of plaque behavior. Previous experiments in our lab showed that human plaques do not exhibit the traditional preconditioning behavior common to most soft tissues. In particular, the behaviors of three classes of plaques differed fundamentally in response to multiple, successive, cyclic compression protocols. In this report, we demonstrate that plaques exhibit different responses to successive relaxation tests in uniaxial compression. Not only is there significant relaxation, but there are composition-dependent differences in the general character of the relaxation responses. Such information on the time-dependent behavior is important for the design of clinical protocols such as stenting or angioplasty wherein the atherosclerotic vessel is subjected to persistent or multiple short duration loadings. This study presents a step toward a better understanding of the biomechanical behavior of atherosclerotic plaques; however, the need for much more data remains. PMID- 11255178 TI - Effect of cellulose acetate materials on the oxidative burst of human neutrophils. AB - Following adverse clinical events involving seven patients undergoing renal dialysis using 12-year-old cellulose acetate hemodialyzers, this in vitro study was proposed in an effort to characterize the inflammatory response to the constituent cellulose acetate (CA) fiber materials. Chemiluminescence (CL) and apoptosis assays were used to determine whether human neutrophils were activated by CA fiber materials and/or are sensitive to degradation/alteration of these fibers over time. Furthermore, the study examined in vitro assays with human neutrophils using a CA film, the solvents used in the film preparation and CA resin. The film could be cut to identical sized pieces in an effort to compare hemodialysis material effects in standardized amounts. For the CL assays, 60-min exposure was followed by secondary stimulation with n-formyl-met-leu-phe (fMLP) or phorbol-12-myristate-13-acetate (PMA). Short-term exposure (60-min postintroduction to CA materials) increased the inflammatory response as measured by the respiratory burst of neutrophils (p < or =.05), with CA fiber exposure significantly compared with cells alone. There was a trend toward an increased response with exposure to older fibers with secondary PMA stimulation. Apoptosis was increased 12% with exposure to the more aged fibers versus 2% with the new fibers. The fiber storage component, glycerol, significantly inhibited the oxidative response (p < or =.001; > or =80% suppression with concentrations of 5 20%). The solvents used in film preparation, N,N-dimethylacetamide and tetrahydrofuran, produced greater than a 70% and 60% suppression, respectively, of CL activity for all concentrations > or =1%. More work is needed to determine the specific nature of the interaction of inflammatory cells with CA materials, but early evidence suggests that neutrophils are activated by CA and display an altered response to more aged fibers. PMID- 11255179 TI - Small-diameter compliant arterial graft prosthesis: Design concept of coaxial double tubular graft and its fabrication. AB - To minimize compliance mismatch between native artery and arterial graft prosthesis over the entire pressure regions, we proposed a coaxial double tubular artificial graft which consists of an enhanced compliant inner tube and a less compliant outer tube, both of which were fabricated using well-controlled multiply micropored segmented polyurethane (SPU) films. Double tubular grafts were coaxially assembled by inserting the inner tube into the outer tube. First, the pressure-diameter (P-D) relationship of canine common carotid arteries, which exhibited a "J" curve, was determined as a targeted artery. Two determinant variables, the pressure-induced distensibility of each tube and the intertubular space distance, were defined and formulated in several models of coaxial double tubular SPU grafts, which had various intertubular space distances, micropore densities, and wall thicknesses. The distensibility of the inner tube determined the distensibility in the low-pressure regions, which was adjusted using wall thickness and microporosity. Thinner films with higher porosities resulted in a high pressure-induced distensibility. On the other hand, a low pressure-induced distensibility in the high-pressure regions was realized using an outer tube with a thicker wall and lower microporosity. The transition point from low- to high pressure regions was determined by the intertubular distance using the theoretical values. On the basis of these results, we presented a prototype model of a coaxial double tubular graft that exhibited well-matched compliance with canine carotid artery. PMID- 11255180 TI - Bioactive bone cement: Effect of silane treatment on mechanical properties and osteoconductivity. AB - A novel bioactive bone cement (GBC) was developed with newly designed bioactive MgO-CaO-SiO(2)-P(2)O(5)-CaF(2) glass beads as the inorganic filler and high molecular weight poly(methyl methacrylate) as the organic matrix. The purpose of this study was to examine the relationship between the amount of the silane coupling agent (gamma-methacryloxy propyl trimethoxy silane) used to treat the glass beads and the mechanical and biological properties of the resultant bone cement. Serial changes in the cement over time were also investigated. Five different kinds of cement, in which the glass beads were treated with different amounts of the coupling agent, were prepared. The quantities of the coupling agent were 0 (control), 0.1, 0.2, 0.5, and 1.0% (w/w) of the glass beads, and the cements were designated GBCs0, GBCs0.1, GBCs0.2, GBCs0.5, and GBCs1.0, respectively. After soaking in water at 75 degrees C for 5 days, GBCs0.1 and GBCs0.2 had significantly higher bending strengths than the other cements. Each GBC was packed into intramedullar canals of rat tibiae to evaluate osteoconductivity, as determined by affinity indices. Rats were killed 4 and 8 weeks after the operation. The affinity index was calculated for each GBC and equaled the length of bone in direct contact with the cement and was expressed as a percentage of the total length of the cement surface. Histologically, new bone had formed along all of the GBC surfaces within 4 weeks. At each time interval, a decreasing trend in the affinity index of GBC was found as the amount of the coupling agent increased. At 8 weeks, no significant change in the affinity index occurred when the amount of the coupling agent increased from 0 to 0.2%, whereas a significant decrease in the affinity index was observed when the amount of the coupling agent increased from 0 to 0.5 or 1.0%. The affinity indices for all the GBCs increased significantly up to 8 weeks. When both the mechanical properties and osteoconductivity were taken into consideration, GBCs0.1 and GBCs0.2 were the best cements, and they showed excellent osteoconductivity and strong enough mechanical properties for clinical use. PMID- 11255176 TI - Bone tissue engineering in a rotating bioreactor using a microcarrier matrix system. AB - A novel approach was utilized to grow in vitro mineralized bone tissue using lighter-than-water, polymeric scaffolds in a high aspect ratio rotating bioreactor. We have adapted polymer microencapsulation methods for the formation of hollow, lighter-than-water microcarriers of degradable poly(lactic-co-glycolic acid). Scaffolds were fabricated by sintering together lighter-than-water microcarriers from 500 to 860 microm in diameter to create a fully interconnected, three-dimensional network with an average pore size of 187 microm and aggregate density of 0.65 g/mL. Motion in the rotating bioreactor was characterized by numerical simulation and by direct measurement using an in situ particle tracking system. Scaffold constructs established a near circular trajectory in the fluid medium with a terminal velocity of 98 mm/s while avoiding collision with the bioreactor wall. Preliminary cell culture studies on these scaffolds show that osteoblast-like cells readily attached to microcarrier scaffolds using controlled seeding conditions with an average cell density of 6.5 x 10(4) cells/cm(2). The maximum shear stress imparted to attached cells was estimated to be 3.9 dynes/cm(2). In addition, cells cultured in vitro on these lighter-than-water scaffolds retained their osteoblastic phenotype and showed significant increases in alkaline phosphatase expression and alizarin red staining by day 7 as compared with statically cultured controls. PMID- 11255181 TI - In vitro biocompatibility of resorbable experimental glass ceramics for bone substitutes. AB - Tricalcium phosphate ceramics (TCPs) are increasingly used as bone substitutes. They demonstrate good biocompatibility and degrade relatively slowly. New glass ceramics based on calcium alkali orthophosphates (Ca(2)KNa(PO(4))(2)) were developed that degrade faster than TCP but could have reduced biocompatibility due to their high solubility. Therefore, they were modified by a neutralizing surface treatment. The aim of this study was to evaluate the biocompatibility of some of these ceramics, GB1a, GB9, and GB14, which differ in the amount of added Na, K, Mg, or Si ions, with standard and modified surfaces. The in vitro cytotoxicity of the ceramics GB1a, GB9, and GB14 was determined by the agar diffusion and filter test and the microculture tetrazolium (MTT) assay. In order to investigate the influence of surface modification, these three ceramics were compared to their surface-treated counterparts, GB1aN, GB9N, and GB14N. GB1a, the ceramic with the highest in vitro solubility, showed the strongest toxic influence in all cell culture tests. GB9 and GB14 produced better results. In contrast, the counterparts with modified surfaces exhibited no (GB9N, GB14N) or weak (GB1aN) signs of cytotoxicity. It is concluded that the toxicity of the ceramics GB1a, GB9, and GB14 depends on their solubility. A positive influence of the surface treatment on in vitro biocompatibility was demonstrated. Therefore, the surface-treated glass ceramics could be promising materials for bone replacement. PMID- 11255182 TI - Molded porous poly (L-lactide) membranes for guided bone regeneration with enhanced effects by controlled growth factor release. AB - The aim of this study was to develop platelet-derived growth factor (PDGF-BB) loaded moldable porous poly (L-lactide) (PLLA)-tricalcium phosphate (TCP) membranes for guided bone regeneration (GBR) therapy. The membranes were designed to fit various types of bone defect sites. PDGF-BB-dissolved PLLA-TCP in methylene chloride-ethyl acetate solution was cast on a dome shaped metallic mold to fabricate a model membrane. The release rate of PDGF-BB, the osteoblast attachment test, and guided bone regeneration potential were evaluated with PDGF BB-loaded PLLA-TCP membranes. Regular pores were generated throughout the membrane mainly due to phase inversion of PLLA-methylene chloride-ethyl acetate solution. A therapeutic amount of PDGF-BB was released from the membrane. The release rate could be controlled by varying the initial loading content of PDGF BB. A significant amount of cells attached onto the PDGF-BB-loaded membrane rather than onto the unloaded membrane. Dome shaped bone formation was achieved in rabbit calvaria at 4 weeks. This indicated that restoration of bone defects to the bone's original shape can be made possible by using molded membranes, which guide bone regeneration along with providing sufficient spaces. Bone forming efficiency was increased remarkably due to PDGF-BB release from PLLA-TCP membranes. These results suggested that the PDGF-BB releasing molded PLLA-TCP membrane may potentially improve GBR efficiency in various types of bone defects. PMID- 11255183 TI - Synthesis and characterization of macroporous chitosan/calcium phosphate composite scaffolds for tissue engineering. AB - Chitosan scaffolds reinforced by beta-tricalcium phosphate (beta-TCP) and calcium phosphate invert glass were fabricated with a low-cost, bioclean freeze-drying technique via thermally induced phase separation. The microstructure, mechanical performance, biodegradation, and bioactivity of the scaffolds were studied. The composite scaffolds were macroporous, and the pore structures of the scaffolds with beta-TCP and the glass appeared very different. Both the compressive modulus and yield strength of the scaffolds were greatly improved, and reinforced microstructures were achieved. The bioactivity tests showed a continuous decrease in both Ca and P concentrations of a simulated body fluid (SBF) after the scaffolds with beta-TCP were immersed in the SBF for more than 20 h, which suggests that an apatite layer might be formed on the scaffolds. However, the same was not observed for the pure chitosan scaffolds or the scaffolds incorporated with the glass. This was further confirmed by micrographs from scanning electron microscopy. This study suggests that the desirable pore structure, biodegradation rate, and bioactivity of the composite scaffolds might be achieved through controlling the ratio of chitosan and calcium phosphates or beta-TCP and the glass. PMID- 11255184 TI - Preferential degradation of osteoclasts by titanium tetrachloride. AB - Although titanium alloys are known to be biocompatible with bone tissue after implantation in human beings, the effect of titanium on osteoclasts remains to be studied. We examined the effect of titanium salt on the formation and survival of osteoclasts in cell culture. The addition of 10 microM titanium tetrachloride caused a decrease in the cell number of osteoclast-like cells induced in bone marrow cell cultures taken from mice. The addition of 10 microM titanium tetrachloride caused degradation of the disaggregated osteoclasts taken from neonatal rats and a decrease in bone resorption. Along with the increase in the degradation of osteoclasts, the number of apoptotic cells increased. Titanium tetrachloride dose-dependently decreased the cell number and alkaline phosphatase activity of osteoblastic cell cultures taken from rat calvaria. However, these concentrations were 30-40 times higher than those in the case of osteoclast-like cell formation. These results showed that titanium ions caused a preferential degradation of osteoclasts rather than osteoblasts, most likely by apoptosis. PMID- 11255185 TI - Effects of digestion protocols on the isolation and characterization of metal metal wear particles. I. Analysis of particle size and shape. AB - Isolation of metal wear particles from hip simulator lubricants or tissues surrounding implants is a challenging problem because of small particle size, their tendency to agglomerate, and their potential for chemical degradation by digestion reagents. To provide realistic measurements of size, shape, and composition of metal wear particles, it is important to optimize particle isolation and minimize particle changes due to the effects of the reagents. In this study (Part I of II), transmission electron microscopy (TEM) was used to examine and compare the effects of different isolation protocols, using enzymes or alkaline solutions, on the size and shape of three different types of cobalt based alloy particles produced from metal-metal bearings. The effect on particle composition was examined in a subsequent study (Part II). Large particles (<1200 nm) were generated by dry abrasion of CoCrMo alloy against itself and small particles (<300 nm) were generated by hip simulator testing of a metal-metal implant pair in the presence of either distilled-deionized water or a 95% bovine serum solution. The reagents changed particle size and to a lesser extent particle shape. For both large particles and small particles generated in water, the changes in size were more extensive after alkaline than after enzymatic protocols and increased with alkaline concentration and time in solution, up to twofold at 2 h and threefold at 48 h. However, when isolating particles from 95% serum, an initial protective effect of serum proteins and/or lipids was observed. Because of this protective effect, there was no significant difference in particle size and shape for both oval and needle-shaped particles after 2 h in 2N KOH and after enzymatic treatments. However, round particles were significantly smaller after 2 h in 2N KOH than after enzymatic treatments. Particle composition may also have been affected by the 2N KOH treatment, as suggested by a difference in particle contrast under TEM, an issue examined in detail in Part II. PMID- 11255186 TI - Effects of digestion protocols on the isolation and characterization of metal metal wear particles. II. Analysis of ion release and particle composition. AB - The isolation of metal wear particles from hip simulator lubricants is important for understanding wear mechanisms and the tissue response to particulate material. Part I of this study demonstrated that isolation protocols involving digestion reagents can chemically attack metal-metal wear particles, reducing their size and changing their shape. In part II of this study, Co and Cr ion concentrations in solution after each digestion protocol were measured by flame atomic absorption spectrometry, and wear particle composition was determined by X ray analysis spectra. The exposure of wear particles in water to alkaline solutions caused an increasing release of Cr ions in solution with alkaline concentration and time, and a corresponding decrease in particle Cr peak intensity on X-ray spectra. As a result, particles exposed to 12N KOH for 48 h displayed Co peaks and no Cr. In contrast, enzymatic protocols caused a release of Co ions in solution and a corresponding decrease in particle Co peak intensity on X-ray spectra, especially with sodium phosphate as a buffer. However, when isolating particles from 95% serum, there was an initial protective effect of serum proteins, presumably because of their binding to Co and Cr. As a result, the extent of Cr ion release from metal wear particles in 95% serum after alkaline treatments was diminished, although still present, whereas both enzymatic protocols resulted in a negligible release of Co and Cr ions into solution. Particle composition analysis after enzymatic treatments revealed the presence of chromium oxide particles and CoCrMo particles with variable Co/Cr ratios. After alkaline treatments, the chromium oxide particles increasingly disappeared with time and alkaline concentration, demonstrating a change in particle composition after these treatments. This study demonstrated that digestion reagents can induce chemical changes that affect particle composition. Of all the protocols tested, the enzymatic protocols were the least damaging to the particles and appeared to be the best compromise for isolation and characterization of metal particles, especially in 95% serum. Special care on the choice of buffers should be taken when isolating particles from a lower concentration of serum. PMID- 11255187 TI - Modulation of osteosarcoma cell growth and differentiation by silane-modified surfaces. AB - The effects of growing the Saos-2 human osteosarcoma cell line onto surfaces containing -CH(3), -OH, -COOH, -NH(2), and C6H5 groups obtained by silane modification were examined. These cells were used because of the great importance of bone cells in many aspects of biomaterials research. Silane-modified surfaces were characterized by contact angle measurements and, subsequently, surface energies were calculated. Cells grown on clean glass, as well as those grown on glass surfaces containing the functional groups cited above, were examined by light and scanning electron microscopy and assessed for their growth characteristics (i.e., determination of cell number and Ki67 antigen expression). The data presented seemed to indicate that if Saos-2 cells are grown on silane modifed surfaces containing the methyl (CH(3)), hydroxyl (OH), and phenyl (C6H5) functional groups, their proliferation is slowed down while growth of these cells on glass surfaces modified with amino (NH(2)) and carboxyl (COOH) groups did not significantly affect growth. Once it was demonstrated that these three functional groups induce significant variations in proliferation, cells grown on these surfaces were also tested for apoptosis and expression of important markers of bone cell differentiation (i.e., osteonectin and osteopontin) by flow cytometry and eventual rearrangement of these markers by fluorescence microscopy. The data suggested that growth of Saos-2 cells on CH(3) induces the most evident morphological changes while growth of these cells on OH and C6H5 brings about the greater variations in osteonectin and osteopontin. We hypothesized that these changes are indicative of an increase in differentiation of Saos-2 cells when grown on the OH and C6H5 groups. PMID- 11255189 TI - Comparison of plasma-sprayed hydroxyapatite coatings and hydroxyapatite/tricalcium phosphate composite coatings: in vivo study. AB - This study aimed to compare biological properties, including osteoconduction, osseointegration, and shear strength, between plasma-sprayed hydroxyapatite (HA) and HA/tricalcium phosphate (TCP) coatings, using a transcortical implant model in the femora of canines. After 3 and 12 weeks of implantation, the implants with surrounding bone were assessed histologically in undecalcified sections in backscattered electron images (BEIs) under a scanning electron microscope (SEM). After short-term (3 week) follow-up, both coatings conducted new bone formation and revealed direct bone-to-coating contact. The HA/TCP coating could not enhance early host-to-coating responses. At 12 weeks, serious dissolution of the HA/TCP coatings evidently occurred. By the new bone healing index (NBHI) and apposition index (AI), we found no significant difference between HA/TCP-coated implants and HA-coated implants throughout all implant periods. At 12 weeks of implantation, some particles dissociated from the HA/TCP coating were found within the remodeling canal. After push-out measurements, the shear strength and failure mode of HA/TCP-coated implants were similar to those of HA-coated implants, and no statistical differences were found between either coating. Consequently, this study indicates that HA/TCP coatings have excellent biological response and may be considered suitable bioactive ceramic coatings for short-term clinical use. PMID- 11255188 TI - Both cyclooxygenase-1 and cyclooxygenase-2 mediate osteoblast response to titanium surface roughness. AB - Previous studies suggest that the enhanced expression of the osteoblastic phenotype exhibited by MG63 osteoblast-like cells on rough Ti surfaces (R(a) 4-5 microm) involves increased production of prostaglandin. Inhibition of prostaglandin synthesis by indomethacin blocks surface-roughness-dependent decreases in cell proliferation and increases in alkaline phosphatase activity and the production of osteocalcin and TGF-beta1. This study examined the hypothesis that the increase in expression of the osteoblastic phenotype noted in MG63 cells cultured on rough Ti surfaces is mediated by inducible cyclooxygenase 2 (Cox-2) whereas Cox-1 modulates prostaglandin production and phenotypic expression of the cells under standard conditions and on smooth Ti surfaces. MG63 cells were cultured on tissue culture plastic, smooth Ti (PT, R(a) = 0.60 microm), and two rough Ti surfaces with differing morphologies (SLA, R(a) = 3.97 microm and TPS, R(a) = 5.21 microm). At 24 h after plating, media were replaced with media containing the general Cox inhibitor indomethacin (10(-7)M), the Cox-1 inhibitor resveratrol (1 or 10 microM), or the Cox-2 inhibitor NS-398 (1 or 10 microM). Media were changed again after 48 h. Five days after plating, osteocalcin, PGE(2), and TGF-beta1 content of the conditioned media were determined. Cell numbers were assessed in the same cultures used for determination of osteocalcin production. Cell layer protein and alkaline phosphatase specific activity were assessed in cultures used to measure PGE(2) and TGF-beta1. Indomethacin, resveratrol, and NS-398 had no effect on cell number. Indomethacin blocked the surface-roughness-dependent increase in PGE(2) production by up to 80%. Similarly, resveratrol inhibited up to 50% of the PGE(2) production on smooth surfaces and up to 80% on rough surfaces. In contrast, NS 398 had no effect on PGE(2) production by cells on smooth surfaces but caused a 60% reduction in cultures on rough surfaces. Indomethacin reduced alkaline phosphatase on all surfaces below basal levels. However, neither resveratrol nor NS-398 had an effect. Indomethacin blocked the stimulatory effect of surface roughness on osteocalcin production while resveratrol only partially reduced osteocalcin production, and NS398 completely blocked the surface-dependent increase. TGF-beta1 production on rough surfaces was blocked by indomethacin. The effects of resveratrol and NS-398 were dose dependent, but neither agent caused total inhibition of the increase noted on SLA, and only resveratrol blocked the increase on TPS. These results indicate that both Cox-1 and Cox-2 are involved in the response of osteoblasts to surface roughness with respect to production of PGE(2), TGF-beta1, and osteocalcin. While prostaglandin mediates the effects of surface roughness on alkaline phosphatase, neither Cox-1 nor Cox-2 appears to be involved, at least with respect to the two inhibitors used. PMID- 11255191 TI - Novel cell immobilization method utilizing centrifugal force to achieve high density hepatocyte culture in porous scaffold. AB - Cell seeding is one of the key procedures in the construction of tissue engineered organs. In our previous efforts to create a bioartificial liver, high density cultures of hepatocytes (>1 x 10(7) cells/1 cm(3)-substrate) and long term maintenance of metabolic function were achieved with a packed-bed reactor utilizing porous poly(vinyl formal) (PVF) resin as a scaffold. However, a low seeding efficiency of about 30% remains a major obstacle to the scaleup of the reactor. In the present study, a new cell seeding method, centrifugal cell immobilization (CCI), which is based on alternating centrifugation and resuspension, was used to achieve high-density seeding and improve the seeding efficiency. Using the CCI method, the maximum density of the immobilized hepatocytes reached 3.8 x 10(7) cells/1 cm(3)-PVF, and the seeding efficiency was improved to about 43% after a relatively short immobilization process (about 15 min). Moreover, further improvement of the seeding efficiency was obtained by serial immobilization procedures. Thus, we concluded that this method is useful and effective for seeding cells into 3-dimensional scaffolds. PMID- 11255190 TI - In vivo biocompatibility of carbodiimide-crosslinked collagen matrices: Effects of crosslink density, heparin immobilization, and bFGF loading. AB - Collagen matrices, crosslinked using N-(3-dimethylaminopropyl)-N' ethylcarbodiimide (E) and N-hydroxysuccinimide (N), were previously developed as a substrate for endothelial cell seeding of small-diameter vascular grafts. In the present study, the biocompatibility of various EN-crosslinked collagen matrices was evaluated following subcutaneous implantation in rats for periods up to 10 weeks. The effects of the crosslink density, referred to as the number of free primary amino groups per 1,000 amino acid residues (EN10, EN14, EN18, or EN22), the amount of heparin immobilized to EN14, and the effect of preloading heparinized EN14 with basic fibroblast growth factor (bFGF) on the induced tissue reaction were studied. EN-crosslinked collagen was biocompatible at both early and late time intervals, and matrices with high crosslink densities (i.e., EN14, EN10) especially demonstrated a significantly decreased antigenic response when compared to noncrosslinked collagen. Furthermore, increased crosslinking resulted in a decreased degradation rate. Immobilization of heparin onto EN14 resulted in a similar to EN14 (thus without heparin) or somewhat reduced tissue reaction, but fibrin formation and vascularization were increased with increasing quantities of immobilized heparin. Matrices preloaded with bFGF also demonstrated good biocompatibility, especially in combination with higher amounts of immobilized heparin. The latter matrices [EN14 with high heparin and bFGF, thus EN14-H (0.4)F and EN14-H(1.0)F] demonstrated significantly increased vascularization for periods up to 3 weeks. Neither heparin immobilization nor bFGF preloading induced an increased antigenic response. It is concluded that the results of this study justify further evaluation of bFGF preloaded, heparin immobilized EN14 collagen, as a matrix for endothelial cell seeding in experimental animals. PMID- 11255192 TI - Effect of thermal treatment on bioactive glass microstructure, corrosion behavior, zeta potential, and protein adsorption. AB - Bioactive glass ceramic is characterized by high mechanical strength and a slow rate of bone bonding. To understand the factors contributing to a decrease in the rate of bone bonding to bioactive glass ceramic, we evaluated the effect of different percentages of bioactive glass crystallization on corrosion behavior, zeta potential, and serum protein adsorption. X-ray diffraction analysis showed that heat treatment of bioactive glass in the temperature range 550 degrees -700 degrees C resulted in the precipitation of Na(2)Ca(2)Si(3)O(9) crystals in the glass matrix. The percentage of crystallization increased in the order: 5%, 8%, 45%, and 83% after thermal treatment at 550 degrees, 600 degrees, 650 degrees, and 700 degrees C/1 h, respectively. Scanning electron microscopic analyses of bioactive glass treated at 550 degrees C showed major glass in glass-phase separation. Moreover, energy-dispersive X-ray analyses indicated that during crystallization P is concentrated in the glassy phase. Induced-coupled plasma analyses showed that after 24 h immersion in simulated body fluid, the concentration of the released P ion increased as the crystallization percentage of bioactive glass increased. zeta potential of bioactive glass samples containing 5% crystallization had a statistically significant higher negative value than control untreated bioactive glass (p <.02). Control untreated bioactive glass adsorbed a statistically significant higher amount of serum protein than bioactive glass samples containing 5% crystallization (p <.02). Results of our study suggest that inhibition of protein adsorption might be responsible for the slow rate of bone bonding to bioactive glass ceramic. It is also possible that conformation changes inhibit the activity of the protein adsorbed onto thermally treated bioactive glass. PMID- 11255193 TI - Evaluation of carboxymethylcellulose, hydroxypropylmethylcellulose, and aluminum hydroxide as potential carriers for rhBMP-2. AB - Conventional iliac crest nonvascularized corticocancelous bone grafts and bone flaps have been used to treat bony defects. However, these treatments have some limitations, namely, the availability of donor tissue, donor site morbidity, difficulty to shape the bone flap to the defect, and complexity of the surgery. The bone morphogenetic protein (rhBMP-2) is osteoinductive. However, its implantation requires a matrix (carrier) in order to define the shape of the resulting bone and to retain the protein at the site for the time required for induction to occur. When the ideal carrier is found, an unlimited supply of material would be available for all applications where bone is needed. In this in vitro study, we evaluated the suitability of some potential carriers for rhBMP-2 by measuring the alkaline phosphatase (ALP) activity of fibroblast cultures. Either rhBMP-2 or sodium carboxymethylcellulose significantly increased the ALP activity, when used alone. When sodium carboxymethylcellulose was combined with rhBMP-2, there was an increase in the ALP activity, but lower than those obtained when the products were used alone. Hydroxypropylmethylcellulose alone did not affect ALP activity. However, the combination of rhBMP-2 with hydroxypropylmethylcellulose did not increase the ALP activity, despite the presence of rhBMP-2. Aluminium hydroxide proved to be an unsuitable rhBMP-2 adsorbent. PMID- 11255194 TI - Degradation behaviors of biodegradable macroporous scaffolds prepared by gas foaming of effervescent salts. AB - Biodegradable polymeric scaffolds for tissue engineering were fabricated by a gas foaming/salt-leaching method using a combination of two effervescent salts, ammonium bicarbonate and citric acid. Poly(D,L-lactic-co-glycolic acid) (PLGA) in a state of gel-like paste was first produced by precipitation of PLGA dissolved in chloroform into ethanol. The polymer slurry was mixed with sieved particles of ammonium bicarbonate, molded, and then immersed in an aqueous solution of citric acid to generate macroporous scaffolds. The scaffolds had relatively homogeneous pore structures throughout the matrix and showed an average pore size of 200 microm and over 90% porosity. By adjusting the concentration of citric acid in the aqueous medium, it was possible to control porosity as well as mechanical strength of the scaffolds. In vitro degradation studies of three different scaffolds having lactic/glycolic acid molar ratios of 75/25, 65/35, and 50/50 exhibited marked swelling behaviors at different critical time points. The swollen matrices had a hydrogel-like internal structure. It was found that massive water uptake into the degrading scaffolds induced matrix swelling, which facilitated the hydrolytic scission of PLGA chains with concomitant disintegration of the matrices. PMID- 11255195 TI - Adsorption characteristics of proteins on calcium phosphates using liquid chromatography. AB - The adsorption characteristics of various proteins on beta-Ca(3)(PO(4))(2) (beta TCP), CaHPO(4). 2H(2)O (DCPD) and CaHPO(4) (DCPA) have been investigated using liquid chromatography. Acidic proteins were eluted from these calcium phosphate surfaces by potassium phosphate buffer (KP) but not by KCl and MgCl(2), whereas basic proteins were eluted by all three eluents. Such elution behavior can be interpreted by considering two kinds of adsorbing sites on the calcium phosphates, namely positively charged sites (Ca sites) which adsorb acidic proteins, and negatively charged sites (P sites) which adsorb basic proteins. A similar behavior has also been observed on Ca(10)(PO(4))(6)(OH)(2) (HAp) and Ca(8)H(2)(PO(4))(6). 5H(2)O (OCP). The order of the ratios of Ca-to-P sites was estimated to be DCPD > OCP > HAp >> DCPA >> beta-TCP, which is in agreement with the order of the surface zeta potentials. In addition, the (0k0) surface of DCPD is suggested to consist mainly of Ca sites. The total number of Ca sites on beta TCP is thought to be significantly smaller than the other calcium phosphates. PMID- 11255196 TI - Tissue reactions to epoxy-crosslinked porcine heart valves post-treated with detergents or a dicarboxylic acid. AB - Calcification limits the long-term durability of xenograft glutaraldehyde (GA) crosslinked heart valves. Previously, a study in rats showed that epoxy crosslinked heart valves reduced lymphocyte reactions to the same extent as the GA-crosslinked control and induced a similar foreign-body response and calcification reaction. The present study was aimed at reducing the occurrence of calcification of epoxy-crosslinked tissue. Two modifications were carried out and their influence on cellular reactions and the extent of calcification after 8 weeks' implantation in weanling rats was evaluated. First, epoxy-crosslinked valves were post-treated with two detergents to remove cellular elements, phospholipids and small soluble proteins, known to act as nucleation sites for calcification. The second approach was to study the effect of the impaired balance between negatively and positively charged amino acids by an additional crosslinking step with a dicarboxylic acid. The detergent treatment resulted in a washed-out appearance of especially the cusp tissue. With the dicarboxylic acid, both the cusps and the walls had a limited washed-out appearance. The wall also demonstrated some detachment of the subendothelium. After implantation, both detergent and dicarboxylic acid post-treatment histologically resulted in reduced calcification at the edges of cusps and walls. However, total amounts of calcification, measured by atomic emission spectroscopy, were not significantly reduced. Data concerning the presence of lymphocytes varied slightly, but were in the same range as the GA-crosslinked control, i.e., clearly reduced compared with a noncrosslinked control. It is concluded that both the double detergent and the dicarboxylic acid post-treatment of epoxy-crosslinked heart valve tissue do not represent a sound alternative in the fabrication of heart valve bioprostheses. PMID- 11255197 TI - Chemical and biological evaluation of endotoxin contamination on natural rubber latex products. AB - Relationship between pyrogenicity and bacterial endotoxin contamination on latex products was demonstrated by chemical analysis and biological assays. In commercially available latex products' surveillance, water extracts prepared from one surgical glove and two silicone elastomer-coated Foley catheters sterilized by gamma-irradiation were obviously pyrogenic in rabbits. The induced fever was monophasic at low dose of the pyrogenic extracts and biphasic at high dose. These extracts exhibited limulus amebocyte lysate gelation activity, and induced inflammatory cytokine (interleukin-1, interleukin-6, and tumor necrosis factor alpha) production from MM6-CA8 human monocytoid cells. These biological properties, including pyrogenicity, completely disappeared by treating the pyrogenic extracts with endotoxin-adsorbent affinity column. Limulus amebocyte lysate activity and cytokine production from MM6-CA8 cells induced by the extracts were significantly decreased by endotoxin inhibitors, an active fragment peptide of an 18-kDa cationic antimicrobial protein and a synthetic lipid A B464 analogue. Furthermore, very small amounts of 2-keto-3-deoxyoctonate and 3-hydroxy fatty acid, which are common constituents of bacterial endotoxins, were detected by gas chromatography-mass spectrometry analysis of the pyrogenic extracts. These findings clearly showed that the pyrogenicity found in these latex products originated from endotoxins contaminating the products. PMID- 11255198 TI - Chronic antigen-specific immune-system activation may potentially be involved in the loosening of cemented acetabular components. AB - Previous studies have attempted to determine whether aseptic loosening and osteolysis are caused by a T cell-mediated type IV hypersensitivity reaction or a nonspecific foreign body reaction involving phagocytic macrophages. The purpose of this study was to examine the role of the B7-CD28 costimulatory pathway (which is indicative of an activated immune response) in loosening and osteolysis of total joint replacements (TJRs). We harvested periprosthetic tissues from 24 loose, cemented, all polyethylene, acetabular components in patients undergoing revision total hip replacement surgery for aseptic loosening. Prostheses were classified radiographically as to whether ballooning, scalloping osteolysis was present or not. Monoclonal antibodies were used to identify macrophages, antigen presenting cells (APCs) expressing B7-1 or B7-2, total T lymphocytes, and T cells expressing CD28 or CTLA-4. The large numbers of positive cells, including macrophages, T cells, and APCs in both groups are substantially higher than previously reported. Macrophages constituted the predominant cell type, the majority of which were APCs. B7-1 was expressed by 18.3% of all cells, and B7-2 was expressed by 61.0% of cells. Despite the fact that there were no statistically significant differences in expression of proteins in the B7-CD28 pathway between the osteolytic and nonosteolytic groups, the magnitude of positive staining suggests that the process of aseptic loosening (not osteolysis) may involve proteins of the B7-CD28 pathway, particularly B7-2. One possible antigenic stimulus is protein-coated particulate wear debris from prosthetic materials. PMID- 11255199 TI - Surface modification of titanium through amino group implantation. AB - We modified Ti surfaces by implantation of amino (NH(2+)) groups at 10(16) and 10(17) cm(-2). The implanted surfaces were characterized by X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), scanning Auger electron spectroscopy (AES), and second ion mass spectroscopy (SIMS). The experimental results showed that the implanted Ti specimens were covered by a dominant hydrocarbon overlayer due to contamination and the surface oxide layer of implanted specimens became thicker. XPS, AES, and SIMS depth profiles showed that implanted elements had a typical ion implantation distribution and that titanium nitride (TiN) was formed. PMID- 11255201 TI - Chronic fatigue syndrome and fibromyalgia: clinical assessment and treatment. AB - Chronic fatigue syndrome (CFS) and fibromyalgia (FM) are closely related illnesses of uncertain etiology. This article reviews the research literature on these biobehavioral conditions, with an emphasis on explanatory models, clinical evaluation of comorbid psychiatric disorders, assessment of stress factors, pharmacologic and alternative therapies, and cognitive-behavioral treatment studies. Furthermore, clinical protocols suitable for professional practice are presented based on an integration of the authors' clinical observations with published data. The article concludes with the recognition that mental health professionals can offer substantial help to these patients. PMID- 11255202 TI - Psychological issues and treatments for people with diabetes. AB - This article examines psychological issues and their treatment among people with diabetes. The paper contains two main sections, one dealing with diagnosable clinical disorders, and the other with more mundane but nevertheless important subclinical problems in living with diabetes. We review the published literature on prevalence, manifestation, consequences, and treatment of psychological disorders in persons with diabetes, primarily depression, anxiety, and eating disorders. In describing everyday problems in living with diabetes we expand our sources beyond the published literature to include our own clinical and consulting experiences as well as our unpublished qualitative research. These problems include dietary restrictions, self-monitoring of blood glucose, taking insulin injections, and lack of support from family and health care professionals. We describe methods for dealing with such problems and discuss the tension between focusing on emotional distress versus practical issues of disease management. Finally, we briefly present some potentially positive consequences of living with diabetes so that readers can be aware of the inspirational aspects of personal experience with this disease. PMID- 11255203 TI - Multiple sclerosis: empirical literature for the clinical health psychologist. AB - This article reviews the empirical literature related to clinical health psychology in multiple sclerosis (MS). MS is a disease in which the immune system attacks the central nervous system. As such, the interactions between medical and psychological variables are complex, and potentially of considerable importance to patients. Common neuropsychological and psychological problems associated with MS and their etiologies are reviewed. The effects of stress and depression on MS exacerbation are discussed, including clinical, immune, endocrine, and neuroimaging findings. The types of coping common in MS and their effects on adjustment are discussed. The empirical literature on psychological and neuropsychological intervention is reviewed. The small literature on caregiving in MS is also summarized. PMID- 11255204 TI - Quality of life and breast cancer: relationship to psychosocial variables. AB - The aim of this article is to shed more light on the relationship between quality of life and aspects of the psychosocial experience for women with breast cancer. The literature is briefly reviewed, including highlights of the psychosocial consequences of cancer, an exploration of the relationship of psychosocial variables to cancer, and a brief review of psychosocial interventions for cancer. Further, preliminary findings of an on-going NCI study are introduced. Finally, clinical implications are discussed. The purpose of this article is to provide a context and foundation on which future researchers and clinicians can build. Ultimately, we suggest that the biomedical model of disease, though crucial, does not take into account all of the complex factors involved in cancer. The current literature lends support to the argument that a broader, more integrative framework, which includes psychosocial factors, is needed. PMID- 11255205 TI - Psychosocial aspects of the organ transplant experience: what has been established and what we need for the future. AB - This article briefly describes the current status and limitations of the organ transplant process that has now become a routine medical procedure. The article discusses how transplantation is not a cure for end-stage organ disease but an alternative form of treatment with both potential medical and psychosocial problems. Both transplant candidates and recipients encounter psychosocial problems. The article examines how these psychosocial problems affect transplant patients prior to transplant, immediately following surgery, and posttransplant. The psychosocial problems include psychiatric diagnoses, individual and family adjustment and relationship problems, sexual dysfunction, return-to-work (RTW) difficulties, and compliance problems and variables related to noncompliance. The article also reviews the special problems of pediatric and adolescent transplant recipients. The need for empirically supported interventions is noted in each of the problem areas. The author outlines problems with previous research studies that hamper solid interpretations of the data, and discusses literature suggesting that the psychosocial problems of transplant candidates and recipients are likely to be underreported. The article concludes with recommendations about the need to switch research efforts toward intervention studies in the problem areas already solidly identified by the literature. PMID- 11255206 TI - Health education groups for caregivers in an HMO. AB - The short-term effectiveness of a Health Education Group (HEP) intervention program for spouses of frail older adults was compared to the usual care (UC) offered to the spouses of frail older persons in a staff model health maintenance organization. HEP is a multicomponent group program offered in eight weekly, two hour group sessions, and ten monthly, two-hour follow-up group sessions. It includes emotion-focused and problem-focused coping strategies, education, and support. One-hundred and five spouses were recruited and randomly assigned to HEP (n = 58) or UC (n = 47). Spouse caregivers and care recipients were assessed within two weeks of intervention and within two weeks after the completion of the eight weekly group meetings. The results indicate that, for caregivers, HEP was more effective than UC in reducing depression, maintaining social integration, increasing effectiveness in solving pressing problems, increasing knowledge of community services and how to access them, changing caregivers' feelings of competence, and the way they respond to the care giving situation. No significant differences, however, were found between care recipients in the two arms of the study on any of the outcome measures. PMID- 11255207 TI - Quality of life measurement in children and adolescents: issues, instruments, and applications. AB - There is increasing interest in measuring quality of life (QL) in children and adolescents, but this interest has developed without careful attention given numerous important issues. Consequently, there is much diversity and confusion in this measurement area. We discuss at a conceptual level herein how to construe and define QL, approach its measurement, and the implications of for whom this is done. Methodological issues pertaining to validation, proxy report, and child development are also discussed. Guidelines for selecting QL measures are provided and a set of generic QL measures is recommended for further consideration. Finally, applications of the QL concept in the policy, service and care, and science areas are delineated. PMID- 11255208 TI - Pain and emotion: new research directions. AB - Recently, there has been growing interest in the relation between pain and emotion. Numerous recent studies have been conducted in this area. This article provides an introduction to this interesting area by highlighting selected research topics including studies on: stress and pain, negative emotional states and pain, catastrophizing and pain, the fear of pain, emotional regulation processes and pain, the effects of enhancing emotional regulation on pain, and the relation of emotional distress to treatment seeking in persons having pain. The article concludes with a discussion of important directions for future research in this area. PMID- 11255209 TI - Cytological effects of 60 Hz magnetic fields on human lymphocytes in vitro: sister-chromatid exchanges, cell kinetics and mitotic rate. AB - Incubation for 72 h of human peripheral blood cultures in the presence of 60 Hz sinusoidal magnetic fields (MF) at magnetic flux densities of 1.0, 1.5, and 2.0 mT led to stimulation of lymphocyte proliferation but had no influence on the frequency of sister-chromatid exchanges (SCE). The cytotoxic potential of MF combined with the mutagen Mitomycin-C also was analyzed. An opposite effect between MF exposure and Mitomycin-C treatment in terms of cell kinetics and mitotic rate was found, whereas no variation in SCE frequency was observed for this coexposure condition. PMID- 11255210 TI - Chromosomal aberrations in lymphocytes of employees in transformer and generator production exposed to electromagnetic fields and mineral oil. AB - The objective was to study the risk of cytogenetic damage among high voltage laboratory workers exposed to electromagnetic fields and mineral oil. This is a cross sectional study of 24 exposed and 24 matched controls in a Norwegian transformer factory. The exposure group included employees in the high voltage laboratory and in the generator soldering department. Electric and magnetic fields and oil mist and vapor were measured. Blood samples were analyzed for chromosomal aberrations in cultured lymphocytes. In addition to conventional cultures, the lymphocytes were also treated with hydroxyurea and caffeine. This procedure inhibits DNA synthesis and repair in vitro, revealing in vivo genotoxic lesions that are repaired during conventional culturing. In conventional cultures, the exposure group and the controls showed similar values for all cytogenetic parameters. In the DNA synthesis- and repair-inhibited cultures, generator welders showed no differences compared to controls. Among high voltage laboratory testers, compared to the controls, the median number of chromatid breaks was doubled (5 vs. 2.5 per 50 cells; P<0.05) the median number of chromosome breaks was 2 vs. 0.5 (P>0.05) and the median number of aberrant cells was 5 vs. 3.5 (P<0.05). Further analysis of the inhibited culture data from this and a previous study indicated that years of exposure and smoking increase the risk of aberrations. We conclude that there was no increase in cytogenetic damage among exposed workers compared to controls in the conventional lymphocyte assay. In inhibited cultures, however, there were indications that electromagnetic fields in combination with mineral oil exposure may produce chromosomal aberrations. PMID- 11255211 TI - The determinants of Canadian children's personal exposures to magnetic fields. AB - Study of the health effects of magnetic fields often depends on identifying determinants and hence indicators of personal exposure. This study identified determinants of children's exposure to magnetic fields and constructed a prediction model for them. For 632 children participating in a case-control study of childhood leukemia, we made direct measures of exposure over 48 h using a portable device, together with observations on candidate determinants. A child's age and sex, the proportion of time spent in the home, and their parents' education or income were very weak predictors of (logged) mean 48 h magnetic field (R(2) < 1%). More important were province (R(2) = 8.0%) and type of residence (R(2) = 11.3%). Low temperatures at the time of measurement were associated with high fields (about 20% increase for each 10 degrees C below 14, R(2) = 4.9%). Several visible attributes of wiring around residences predicted exposure, mostly captured in the Wertheimer-Leeper wire code (R(2) = 13.5%). Stationary 24 h measurement in the bedroom (R(2) = 63.3%) and spot measurements outside the house (R(2) = 40.7%) predicted personal exposures best. Adding other minor predictors increased only slightly variance explained by 24 h stationary (R(2) = 66.2%) and spot (R(2) = 46.8%) measurements. Without spot or stationary measurements, the best model was much less powerful (R(2) = 29.0%). We conclude that spot measurements outside the residence provide a moderately effective basis for estimating exposure for children living there, but do not perform as well as 24 h stationary measurements in the child's bedroom. Although several other easily-observed variables were associated with personal exposure, they were weak determinants, either individually or in combination. PMID- 11255212 TI - Long-term effects of 60-Hz electric vs. magnetic fields on IL-1 and IL-2 activity in sheep. AB - This study was designed to assess the effect of exposure to long-term extremely low-frequency electric and magnetic fields (ELF-EMF) from a 500 kV transmission line on IL-1 and IL-2 activity in sheep. The primary hypothesis was that the reduction in IL-1 activity observed in our two previous short-term studies (10 months) was due to EMF exposure from this transmission line. To repeat and expand these studies and to characterize the components of EMF responsible for the previously observed reduction in IL-1 activity, the current experiment examined not only the effect of exposure to electric and magnetic fields, but also the magnetic field component alone. In the current study, IL-2 was examined to characterize the effects of EMF exposure on an indicator of T cell responses. 45 Suffolk ewe lambs were randomized into three groups of 15 animals each. One group of animals was placed in the EMF pen, located directly beneath the transmission line. A second group was placed in the shielded MF (magnetic field only) pen, also directly beneath the transmission line. The third group of animals was placed in the control pen located several hundred meters away from the transmission line. During the 27 month exposure period, blood samples were taken from all animals monthly. When the data were analyzed collectively over time, no significant differences between the groups were found for IL-1 or IL-2 activity. In previous studies ewe lambs of 8-10 weeks of age were used as the study animals and significant differences in IL-1 activity were observed after exposure of these animals to EMF at mean magnetic fields of 3.5-3.8 microT (35-38 mG) and mean electric fields of 5.2-5.8 kV/m. At the start of the current study EMF levels were reduced as compared to previous studies. One interpretation of the current data is that magnetic field strength and age of the animals may be important variables in determining whether EMF exposure will affect IL-1 activity. PMID- 11255213 TI - Studies of the interactions between melatonin and 2 Hz, 0.3 mT PEMF on the proliferation and invasion of human breast cancer cells. AB - Interactions between the hormone melatonin at pharmacological concentrations (10( 3) M) and 2 Hz, 0.3 mT pulsed electromagnetic fields (PEMF) on the proliferation and invasion of human breast cancer cells were studied in vitro. Three types of human breast cancer cells were used in this study: MDA-MB-435, MDA-MB-231, and MCF-7. Results showed that cellular growth of MDA-MB-231 cells, which were reported to be lowly metastatic, and MCF-7 cells, which were reported to be nonmetastatic, were both significantly reduced by melatonin regardless of the presence of the field. Results also showed that MDA-MB-435 and MDA-MB-231 cells were invasive, with MDA-MB-231 cells being more invasive than the MDA-MB-435 cells for both unexposed and experimental-PEMF groups. In addition, invasion studies showed that MCF-7 cells were not invasive and that melatonin did not have any effects on the invasion of these cells, with or without the PEMF. It is also suggested that since metastasis requires growth and invasion into tissue, anti invasion agents can be used in conjunction with melatonin to prevent formation of secondary metastases. The overall studies suggest that PEMF at 2 Hz, 0.3 mT does not influence cancer metastasis; while having clinical merit in the healing of soft tissue injury, this field has shown no influence on cancer cells as 60 Hz power line fields have. PMID- 11255214 TI - Large granular lymphocytic (LGL) leukemia in rats exposed to intermittent 60 Hz magnetic fields. AB - An animal model for large granular lymphocytic (LGL) leukemia in male Fischer 344 rats was utilized to determine whether magnetic field exposure can be shown to influence the progression of leukemia. We previously reported that exposure to continuous 60 Hz, 1 mT magnetic fields did not significantly alter the clinical progression of LGL leukemia in young male rats following injection of spleen cells from donor leukemic rats. Results presented here extend those studies with the following objectives: (a) to replicate the previous study of continuous 60 Hz magnetic field exposures, but using fewer LGL cells in the inoculum, and (b) to determine if intermittent 60 Hz magnetic fields can alter the clinical progression of leukemia. Rats were randomly assigned to four treatment groups (18/group) as follows: (1) 1 mT (10 G) continuous field, (2) 1 mT intermittent field (off/on at 3 min intervals), (3) ambient controls ( < 0.1 microT), and (4) positive control (5 Gy whole body irradiation from cobalt-60 four days prior to initiation of exposure). All rats were injected intraperitoneally with 2.2 x 10(6) fresh, viable LGL leukemic spleen cells at the beginning of the study. The fields were activated for 20 h per day, 7 days per week, and all exposure conditions were superimposed over the natural ambient magnetic field. The rats were weighed and palpated for splenomegaly weekly. Splenomegaly developed 9-11 weeks after transplantation of the leukemia cells. Hematological evaluations were performed at 6, 8, 10, 12, 14, and 16 weeks of exposure. Peripheral blood hemoglobin concentration, red blood cells, and packed cell volume declined, and total white blood cells and LGL cells increased dramatically in all treatment groups after onset of leukemia. Although the positive control group showed different body weight curves and developed signs of leukemia earlier than other groups, differences were not detected between exposure groups and ambient controls. Furthermore, there were no overall effects of magnetic fields on splenomegaly or survival in exposed animals. In addition, no significant and/or consistent differences were detected in hematological parameters between the magnetic field exposed and the ambient control groups. PMID- 11255215 TI - Induction of primary root curvature in radish seedlings in a static magnetic field. AB - Primary roots of radish (Raphanus sativus L.) seedlings were exposed to an inhomogeneous static magnetic field generated by a permanent magnet, during continuous rotation on a 0.06 rpm clinostat, thereby reducing the unilateral influence of gravity. The roots responded tropically to the static magnetic field with the tropism appearing to be negative. These roots responded significantly (P < 0.05) to the south pole of the magnet. The significant tropic response was found for a magnetic flux density of 13-68 mT, for a field gradient of 1.8-14.7 T/m, and for the product of magnetic field and field gradient of 0.023-1.0 T(2)/m. A small, but insignificant, response of the roots to the north pole has also been found. PMID- 11255216 TI - Why arguments based on photon energy may be highly misleading for power line frequency electromagnetic fields. AB - When evaluating possible mechanisms by which low frequency electromagnetic fields may have a biological effect, arguments based on photon energy have often been used in a misleading way. For visible light the concept of photons has proved to be very useful in explaining experimental findings. However, the concept of photons cannot be used without major modifications in describing phenomena related to near field problems at power frequency (50 or 60 Hz) electric and magnetic fields. For this regime, the photon description is very complex. A very high number of highly coherent photons must be used in a quantum electrodynamic description of low frequency electromagnetic field phenomena. Thus, one-photon interaction descriptions must be replaced by multiple-photon interaction formalism. However, at low frequencies, a classical electromagnetic field description is far more useful than quantum electrodynamics. There is in principle no difference in how much energy an electron can pick up from a low frequency electric field as compared to from a high frequency photon. Thus, the total gain in energy is not limited to the energy carried by a single photon, which is E = hv, where h is Planck's constant and (v) is the frequency of the radiation. However, the time scale of the primary event in a mechanism of action is very different for ionizing radiation compared to power line frequency fields. The advice is to consider the time scale given by the inverse of the frequency of the fields, rather than photon energy, when one use physics as a guidance in evaluating possible mechanisms for biological effects from low frequency electromagnetic fields. PMID- 11255217 TI - Variations of dose and electrode spacing for rat breast cancer electrochemical treatment. AB - Electrochemical treatment (EChT) with direct current delivered through implanted electrodes has been used for local control of solid tumors in humans. This study tested the hypothesis that rat breast cancer responses to EChT are dependent on electrode spacing and dose, and explored suitable parameters for treating breast cancers with EChT. Rat breast cancers were initiated by injecting 1 x 10(6) MTF-7 cells to the right mammary gland fat pad of Fisher 344 female rats. The rats were randomly divided into designated experimental groups when the tumors grew to approximately 2 x 2 x 2 cm. One hundred and thirty rats were used for a survival study and 129 for a pathology study. A 4-channel EChT machine was used to administer coulometric doses. The survival study indicated that local tumor control rate is less than 40% in the 40 coulomb (C) and 60 C groups and more than 70% in the 80 and 100 C groups. Sixty six rats died of primary tumors, including all 10 rats in the control group. Once a rat's primary tumor was controlled, no recurrence was found. The main reason for terminating the primary tumor-free rats (51) was lymph node metastasis. Thirteen tumor-free rats survived for more than 6 months. The pathology study showed a significant dose effect on EChT induced tumor necrosis. At 10, 20, 40, and 80 C, the fraction showing necrosis were 39.7, 52.3, 62, and 77.7%, respectively (P UMR 106 cells>ROS 17/2.8. Although all three cell lines had similar numbers of GR (50,000/cell), glucocorticoid modulation of alkaline phosphatase activity and cell proliferation was only detectable in ROS 17/2.8 cells. Further studies showed that 11 beta-HSD2 activity in each of the cells was potently stimulated by both A and B, but not by synthetic dexamethasone. This effect was blocked by the 11 beta-HSD inhibitor, 18 beta-glycyrrhetinic acid (but not by GR or MR antagonists) suggesting direct, allosteric regulation of 11 beta HSD2 activity. These data indicate that in osteosarcoma cells 11 beta-HSD2 plays a key role in controlling GR-mediated responses; cells with relatively high levels of 11 beta-HSD2 activity were insensitive to glucocorticoids, whilst cells with low levels showed functional responses to both dexamethasone and B. In addition to the established effects of 11 beta-HSD2 in protecting MR in the kidney and colon, our data suggest that 11 beta-HSD2 in bone represents an important pre-receptor mechanism in determining ligand availability to GR. PMID- 11255229 TI - Expression of SPARC/osteonectin/BM4O in the human gut: predominance in the stroma of the remodeling distal intestine. AB - SPARC is a glycoprotein of the extracellular matrix that exhibits a number of biological functions such as disruption of cell adhesion and modulation of matrix metalloprotease expression. These properties, in concert with the expression of the molecule during development, repair, and neoplastic progression, suggest that SPARC has an important role in remodeling in a variety of tissues. However, the role of SPARC in the intestine is unclear since the development expression and tissular origin of SPARC in this organ appears to be species-dependent. As a first step to investigate the function of SPARC in the tissues of the intestine, we have analyzed its expression at the protein and mRNA levels in the human fetal and adult small intestinal and colonic mucosa as well as in intestinal cell models. Our results show that SPARC expression is differentially regulated during development and along the length of the human intestine. In the colon, SPARC was predominantly found at the epithelial-mesenchymal interface at the fetal stage, below detection levels in the normal adult, but re-expressed in the stroma of colonic tumors. In the small intestine, low levels of SPARC expression were observed at an early stage of morphogenesis (between 9 and 11 weeks) but expression was not detected at subsequent developmental stages nor was it induced in the mucosa of Crohn's disease. While SPARC appeared to be produced mainly by mesenchymal and stromal cells in the intact intestine it was not detected in colon cancer cells. Taken together, these results indicate that SPARC is subject to an onco-fetal pattern of expression in the stroma of the colonic mucosa while its expression is much more restricted in the small intestine, suggesting a differential involvement of this molecule in the extracellular matrix remodeling occurring along the length of the developing and diseased human intestinal mucosa. PMID- 11255230 TI - Mechanisms regulating c-met overexpression in liver-metastatic B16-LS9 melanoma cells. AB - Liver selected B16-LS9 melanoma cells show a dramatic overexpression of the proto oncogene c-met, the cellular receptor for hepatocyte growth factor/scatter factor. As a consequence, c-met becomes constitutively active, and the cells become more responsive to hepatocyte growth factor stimulation. We have investigated the molecular mechanisms regulating c-met expression in both the parental line B16-F1, which has low expression levels, and the liver-specific B16 LS9, overexpressing c-met. Overexpression is observed at the protein and mRNA levels, however without further evidence of gene amplification or rearrangement. c-met promoter activity was higher in B16-LS9 than B16-F1 cells, and also a nuclear run-off showed higher transcription levels in B16-LS9 cells. Moreover, we found that c-met mRNA had a longer half-life in B16-LS9 cells, thus indicating also the involvement of post-transcriptional regulation mechanisms. Finally, we found evidence that autonomous activation of the melanocortin receptor-1 (MCR-1) is at least partially responsible for c-met upregulation in B16-LS9 cells, since treatment of the cells with a potent MSH antagonist (the agouti peptide) has strong down-regulatory effects. PMID- 11255232 TI - Effect of the neuropeptide substance P on the rat bone marrow-derived osteogenic cells in vitro. AB - Substance P containing, thin, sensory nerve fibres have been demonstrated in bone and bone marrow. However the role of substance P in bone tissue is not fully understood. We investigated the effects of substance P on the growth and development of rat bone marrow-derived osteogenic cells in vitro. To examine this, the marrow-derived osteogenic cells were treated from 3rd to 6th day of subculture with substance P at concentrations 10(-10), 10(-9) and 10(-8)M. [(3)H] thymidine, L-2,3-[(3)H]-proline incorporation, protein accumulation, alkaline phosphatase activity, and calcium deposition were measured in cultures. Substance P slightly stimulated [(3)H]-thymidine incorporation at 10(-10) M. Protein accumulation and L-2,3-[(3)H]-proline incorporation were enhanced in a dose dependent manner. Simultaneous application of spantide, a substance P receptor antagonist, could not block substance P-induced L-2,3-[(3)H]-proline incorporation probably because of statistically significant effect of spantide itself. Calcium deposits were significantly lower (about 30%) in cultures treated with SP. This effect was probably due in part by the fall in alkaline phosphatase activity which in substance P treated cultures was decreased about 17%. Our results indicate that substance P could be one of the factors modulating bone metabolism. PMID- 11255231 TI - Functional effect of point mutations in the alpha-folate receptor gene of CABA I ovarian carcinoma cells. AB - The alpha-folate receptor (alpha FR) is overexpressed in 90% of nonmucinous ovarian carcinomas. In addition to the known role of alpha FR binding and mediating the internalization of folates, functional interaction of alpha FR with signaling molecules was recently shown. To identify a model to study the role of alpha FR in ovarian carcinoma, we characterized the alpha FR gene in the ovarian carcinoma cell line CABA I in comparison to a reference line, IGROV1. In CABA I cells, Northern blot analysis revealed an alpha FR transcript of the expected length and FACS analysis indicated receptor expression on the cell membrane; however, RNase protection assay revealed no specific signals. Southern blot and genomic PCR analysis suggested the presence of a rearrangement(s) involving the 5' region of the gene in CABA I cells as compared to IGROV1 cells. Cloning and sequencing of CABA I alpha FR cDNA revealed several point mutations. The partitioning of alpha FR in membrane microdomains from CABA I cells and its association with regulatory molecules was comparable to that of IGROV1 cells. By contrast, the alpha FR expressed on the CABA I cell membrane bound folic acid with lower affinity, and ectopic expression of the corresponding cDNA in CHO cells confirmed impaired folic acid binding. Thus, CABA I cells may provide a tool to delineate functional domains of the alpha FR. PMID- 11255233 TI - Indole-3-propionic acid, a melatonin-related molecule, protects hepatic microsomal membranes from iron-induced oxidative damage: relevance to cancer reduction. AB - Excessive free iron and the associated oxidative damage are commonly related to carcinogenesis. Among the antioxidants known to protect against iron-induced oxidative abuse and carcinogenesis, melatonin and other indole compounds recently have received considerable attention. Indole-3-propionic acid (IPA), a deamination product of tryptophan, with a structure similar to that of melatonin, is present in biological fluids and is an effective free radical scavenger. The aim of the study was to examine the effect of IPA on experimentally induced oxidative changes in rat hepatic microsomal membranes. Microsomes were preincubated in presence of IPA (10, 3, 2, 1, 0.3, 0.1, 0.01 or 0.001 mM) and, then, incubated with FeCl(3) (0.2 mM), ADP (1.7 mM) and NADPH (0.2 mM) to induce oxidative damage. Alterations in membrane fluidity (the inverse of membrane rigidity) were estimated by fluorescence spectroscopy and lipid peroxidation by measuring concentrations of malondialdehyde+4-hydroxyalkenals (MDA+4-HDA). IPA, when used in concentrations of 10, 3 or 2 mM, increased membrane fluidity, although at these concentrations it did not influence lipid peroxidation significantly. The decrease in membrane fluidity due to Fe(3+) was completely prevented by preincubation in the presence of IPA at concentrations of 10, 3, 2 or 1 mM. The enhanced lipid peroxidation due to Fe(3+) was prevented by IPA only at the highest concentration (10 mM). It is concluded that Fe(3+)-induced rigidity and, to a lesser extent, lipid peroxidation in microsomal membranes may be reduced by IPA. However, IPA in high concentrations increase membrane fluidity. Besides melatonin, IPA may be used as a pharmacological agent to protect against iron-induced oxidative damage to membranes and, potentially, against carcinogenesis. PMID- 11255234 TI - Promoter activity of the rat connexin 43 gene in NRK cells. AB - Cellular communication mediated by gap junctions plays a major role in organ function. Gap junction channels are formed by the organization of polypeptide subunits, termed connexins (Cx), on the cell surface of adjacent cells. One mechanism to regulate gap-junctional communication is by change in Cx expression. In the present study, the promoter region of the rat Cx43 gene was characterized. Nested deletions of the 5' flanking region of the first Cx43 exon were coupled to the human growth hormone gene and transfected into normal rat kidney (NRK) cells, that express this gene constitutively. The minimal region of the Cx43 gene that showed maximal promoter activity was localized within 110 bp upstream of the transcriptional initiation site. One particular subregion that contains a Sp-1 binding site (located within 98--93 bp from the transcriptional initiation site) was found to sustain Cx43 promoter activity to the same extent as that of the 110 bp promoter region. Mutations of this Sp-1 binding site abolished transcriptional activity and DNA-protein interactions. These observations suggest that the Sp-1 binding site plays a major role in the basal transcriptional activity of Cx43 gene in NRK cells. PMID- 11255235 TI - Early growth response factor-1 mediates insulin-inducible vascular endothelial cell proliferation and regrowth after injury. AB - Hyperinsulinemia in diabetes mellitus is a significant risk factor in the development of atherosclerosis and early restenosis after balloon angioplasty. These manifestations could be mediated by the ability of insulin to potentiate the cellular proliferative and reparative response of vascular cell types to local stimuli. Here we demonstrate that insulin stimulates DNA synthesis in aortic endothelial cells. Reverse transcription-polymerase chain reaction and Northern blotting revealed that insulin induces the expression and transcriptional activity of the immediate early gene and zinc finger transcription protein, early growth response factor-1 (Egr-1). Western immunoblot analysis revealed that insulin-inducible Egr-1 expression was inhibited using phosphorothioate-specific antisense oligonucleotides targeting Egr-1 mRNA. These agents blocked endothelial cell DNA synthesis stimulated by insulin in a dose dependent manner and inhibited the capacity of insulin to potentiate the reparative response of endothelial cells to mechanical injury in vitro. These oligonucleotides also attenuated wound repair in smooth muscle cells. DNA synthesis induced by insulin was suppressed by inhibitors of two upstream activators of Egr-1, extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3-phosphate (PI 3-K), whereas p38 kinase inhibitors had no effect. These present findings demonstrate that insulin-inducible DNA synthesis and repair after injury are processes critically dependent upon the activation of Egr-1. Additionally, they implicate this transcription factor as a potential target for the inhibition of restenosis in diabetics. PMID- 11255236 TI - Baculovirus-expressed vitamin D-binding protein-macrophage activating factor (DBP maf) activates osteoclasts and binding of 25-hydroxyvitamin D(3) does not influence this activity. AB - Vitamin D-binding protein (DBP) is a multi-functional serum protein that is converted to vitamin D-binding protein-macrophage activating factor (DBP-maf) by post-translational modification. DBP-maf is a new cytokine that mediates bone resorption by activating osteoclasts, which are responsible for resorption of bone. Defective osteoclast activation leads to disorders like osteopetrosis, characterized by excessive accumulation of bone mass. Previous studies demonstrated that two nonallelic mutations in the rat with osteopetrosis have independent defects in the cascade involved in the conversion of DBP to DBP-maf. The skeletal defects associated with osteopetrosis are corrected in these mutants with in vivo DBP-maf treatment. This study evaluates the effects of various forms of DBP-maf (native, recombinant, and 25-hydroxyvitamin D(3) bound) on osteoclast function in vitro in order to determine some of the structural requirements of this protein that relate to bone resorbing activities. Osteoclast activity was determined by evaluating pit formation using osteoclasts, isolated from the long bones of newborn rats, incubated on calcium phosphate coated, thin film, Ostologic MultiTest Slides. Incubation of osteoclasts with ex vivo generated native DBP-maf resulted in a dose dependent, statistically significant, activation of the osteoclasts. The activation was similar whether or not the vitamin D binding site of the DBP-maf was occupied. The level of activity in response to DBP-maf was greater than that elicited by optimal doses of other known stimulators (PTH and 1,25(OH(2)D(3)) of osteoclast function. Furthermore, another potent macrophage activating factor, interferon--gamma, had no effect on osteoclast activity. The activated form of a full length recombinant DBP, expressed in E. coli showed no activity in the in vitro assay. Contrary to this finding, baculovirus-expressed recombinant DBP-maf demonstrated significant osteoclast activating activity. The normal conversion of DBP to DBP-maf requires the selective removal of galactose and sialic acid from the third domain of the protein. Hence, the differential effects of the two recombinant forms of DBP-maf is most likely related to glycosylation; E. coli expressed recombinant DBP is non glycosylated, whereas the baculovirus expressed form is glycosylated. These data support the essential role of glycosylation for the osteoclast activating property of DBP-maf. PMID- 11255237 TI - Age-dependent decline in bone nodule formation stimulating activity in rat serum is mainly due to the change in the corticosterone level. AB - The replacement of fetal bovine serum with rat serum in a culture medium brought about a marked increase in the formation of mineralized bone nodules (BN) in primary cultures of rat calvarial cells. These effects of rat serum were most prominent when added during the early phase of the culture, indicating that the serum factor mainly acts on the cells during the growing phase. A significant increase in BN formation was observable at final rat serum concentration as low as 1%, and the effect was dependent on serum concentration, at least up to 10%. The addition of rat serum also increased alkaline phosphatase (ALP) activity, collagen synthesis, and DNA synthesis in calvarial cells. BN formation stimulating activity was extractable with ethyl acetate. The ethyl acetate extract was purified by TSK-GEL OH-120 column chromatography by monitoring the stimulation of ALP activity in ROS 17/2.8 cells. The chromatographic behavior of the ALP activity was found to be identical to that of corticosterone, the major glucocorticoid in rodents and the preincubation of the purified fraction with anticorticosterone antibody abolished the ALP stimulating activity. These results suggest that BN formation stimulating activity in rat serum is mainly attributable to corticosterone. The concentration of serum corticosterone decreased with age in parallel with BN formation stimulating activity, which suggests that the physiological level of corticosterone may have a regulatory role in the maintenance of osteoblast function. PMID- 11255238 TI - Binding and uptake of differently oxidized low density lipoprotein in mouse peritoneal macrophages and THP-1 macrophages: involvement of negative charges as well as oxidation-specific epitopes. AB - Oxidatively modified low-density lipoprotein (LDL) has been found in vivo, and oxidized LDL (oxLDL) could bind to scavenger receptors, leading to foam cell formation. Macrophages bear a number of different scavenger receptors for oxLDL, and macrophages of different origins may have a different scavenger receptor repertoire. In addition, LDL oxidized to different degrees may differ in the ability to bind macrophage scavenger receptors. In this study, we characterized the patterns of the binding and uptake of differently oxidized LDL in mouse peritoneal macrophages (MPM) and human THP-1 macrophages, and the influence of negative charge and oxidation-specific epitopes in oxLDL on these processes. Thresholds of increased binding and uptake in MPM were found when LDL was oxidized to the degrees with a relative electrophoretic mobility (REM) of 2.6 (minor threshold) and 3.0 (major threshold), corresponding to 49 and 57%, respectively, of the loss of free amino groups in these oxLDL. There was no threshold for the binding of oxLDL to THP-1 macrophages, while for uptake, a major threshold with REM of 3.0 (57% free amino groups lost) was found. The presence of the F(ab')(2) fragments of the monoclonal antibody OB/04, which was raised against copper-oxidized LDL, led to the reduction of the binding and uptake, respectively, of Eu(3+)-oxLDL (REM:3.6) in MPM by 31 and 29%, and by 19 and 22% in THP-1 macrophages. It is concluded that LDL oxidized to different degrees binds differently to macrophages, and the patterns of binding and uptake are different for MPM and human THP-1 macrophages. Both, the negative charge and the oxidation-specific epitopes of oxLDL are involved in these processes. PMID- 11255240 TI - Estimation of allele frequencies with data on sibships. AB - Allele frequencies are generally estimated with data on a set of unrelated individuals. In genetic studies of late-onset diseases, the founding individuals in pedigrees are often not available, and so one is confronted with the problem of estimating allele frequencies with data on related individuals. We focus on sibpairs and sibships, and compare the efficiency of four methods for estimating allele frequencies in this situation: (1) use the data for one individual from each sibship; (2) use the data for all individuals, ignoring their relationships; (3) use the data for all individuals, taking proper account of their relationships, considering a single marker at a time; and (4) use the data for all individuals, taking proper account of their relationships, considering a set of linked markers simultaneously. We derived the variance of estimator 2, and showed that the estimator is unbiased and provides substantial improvement over method 1. We used computer simulation to study the performance of methods 3 and 4, and showed that method 3 provides some improvement over method 2, while method 4 improves little on method 3. PMID- 11255239 TI - Tree-based recursive partitioning methods for subdividing sibpairs into relatively more homogeneous subgroups. AB - We propose a new splitting rule for recursively partitioning sibpair data into relatively more homogeneous subgroups. This strategy is designed to identify subgroups of sibpairs such that within-subgroup analyses result in increased power to detect linkage using Haseman-Elston regression. We assume that the subgroups can be defined by patterns of non-genetic binary covariates measured on each sibpair. The data we consider consists of the squared difference of a quantitative trait measurement on each sibpair, estimates of identity-by-descent (IBD) values at each genetic marker, and binary covariate data describing characteristics of the sibpair (e.g., race, sex, family history of disease). To test the efficacy of this method in linkage analysis, we performed two simulation experiments. In the first, we simulated a mixture consisting of 66.6% of the sibpairs with no linkage and 33.3% of the sibpairs with genetic linkage to one marker. The two groups were distinguished by the value of a single binary covariate. We also simulated one unlinked marker and one random covariate to include as noise in the data. In the second experiment, we simulated a mixture consisting of 55% of the sibpairs with no genetic linkage, 22.5% of the sibpairs with genetic linkage to one marker, and 22.5% of the sibpairs with linkage to a different marker. Each subgroup was defined by a distinct pattern of two binary covariates. We also simulated one unlinked marker and two random covariates to include as noise in the data. Our simulation studies found that we can significantly increase the overall power to detect linkage by fitting Haseman Elston regression models to homogeneous subgroups with only a small increase in the false-positive rate. Second, the splitting rule can correctly identify important covariates and linked markers. Third, recursive partitioning of sibpair data using this splitting rule can correctly identify sibpair subgroups. These results indicate that partitioning sibpairs into homogeneous subgroups is feasible and significantly increases the power to detect linkage, thus demonstrating the practical utility and potential this new methodology holds. PMID- 11255241 TI - Stratified case sampling and the use of family controls. AB - We compare the asymptotic relative efficiency (ARE) of different study designs for estimating gene and gene-environment interaction effects using matched case control data. In the sampling schemes considered, cases are selected differentially based on their family history of disease. Controls are selected either from unrelated subjects or from among the case's unaffected siblings and cousins. Parameters are estimated using weighted conditional logistic regression, where the likelihood contributions for each subject are weighted by the fraction of cases sampled sharing the same family history. Results showed that compared to random sampling, over-sampling cases with a positive family history increased the efficiency for estimating the main effect of a gene for sib-control designs (103 254% ARE) and decreased efficiency for cousin-control and population-control designs (68-94% ARE and 67-84% ARE, respectively). Population controls and random sampling of cases were most efficient for a recessive gene or a dominant gene with an relative risk less than 9. For estimating gene-environment interactions, over-sampling positive-family-history cases again led to increased efficiency using sib controls (111-180% ARE) and decreased efficiency using population controls (68-87% ARE). Using case-cousin pairs, the results differed based on the genetic model and the size of the interaction effect; biased sampling was only slightly more efficient than random sampling for large interaction effects under a dominant gene model (relative risk ratio = 8, 106% ARE). Overall, the most efficient study design for studying gene-environment interaction was the case-sib control design with over-sampling of positive-family-history-cases. PMID- 11255242 TI - Stoppage: an issue for segregation analysis. AB - Segregation analysis assumes that the observed family-size distribution (FSD), i.e., distribution of number of offspring among nuclear families, is independent of the segregation ratio p. However, for certain serious diseases with early onset and diagnosis (e.g., autism), parents may change their original desired family size, based on having one or more affected children, thus violating that assumption. Here we investigate "stoppage," the situation in which such parents have fewer children than originally planned. Following Brookfield et al. [J Med Genet 25:181-185, 1988], we define a stoppage probability d that after the birth of an affected child, parents will stop having children and thus not reach their original desired family size. We first derive the full correct likelihood for a simple segregation analysis as a function of p, d, and the ascertainment probability pi. We show that p can be estimated from this likelihood if the FSD is known. Then, we show that under "random" ascertainment, the presence of stoppage does not bias estimates of p. However, for other ascertainment schemes, we show that is not the case. We use a simulation study to assess the magnitude of bias, and we demonstrate that ignoring the effect of stoppage can seriously bias the estimates of p when the FSD is ignored. In conclusion, stoppage, a realistic scenario for some complex diseases, can represent a serious and potentially intractable problem for segregation analysis. PMID- 11255243 TI - Genome-wide linkage analyses of total serum IgE using variance components analysis in asthmatic families. AB - Variance components models were used to analyze total IgE levels in families ascertained though the Collaborative Study of the Genetics of Asthma (CSGA) using a genome-wide array of polymorphic markers. While IgE levels are known to be associated with clinical asthma and recognized to be under strong genetic control (here the heritability was estimated at 44-60% in the three racial groups), specific genes influencing this trait are still largely unknown. Multipoint analysis of 323 markers yielded little indication of specific regions containing a trait locus controlling total serum IgE levels (adjusted for age and gender). Although a number of regions showed LOD statistics above 1.5 in Caucasian families (chromosome 4) and in African-American families (chromosomes 2 and 4), none yielded consistent evidence in all three racial groups. Analysis of total IgE adjusted for gender, age and Allergy Index (a quantitative score of skin test sensitivity to 14 common aeroallergens) was conducted on these data. In this analysis, a much stronger signal for a trait locus controlling adjusted log[total IgE] was seen on the telomeric end of chromosome 18, but only in Caucasian families. This region accounted for most of the genetic variation in log[total IgE], and may represent a quantitative trait locus for IgE levels independent of atopic response. Oligogenic analysis accounting simultaneously for the contribution of this locus on chromosome 18 and other chromosomal regions showing some evidence of linkage in these Caucasian families (on chromosomes 2, 4 and 20) failed to yield significant evidence for interaction. PMID- 11255244 TI - Gibbs sampling-based segregation analysis of asthma-associated quantitative traits in a population-based sample of nuclear families. AB - Asthma is a common, complex human disease. Elevated serum immunoglobulin E (IgE) levels, elevated blood eosinophil counts, and increased airway responsiveness are physiological traits that are characteristic of asthma. Few studies have investigated major gene effects for these traits in a population-based sample. Further, it is not known if any putative major genes may be common to two or more of these traits. We investigated the existence and nature of major genes modulating asthma-associated quantitative traits in an Australian population based sample of 210 Caucasian nuclear families. The sharing of these major genes was also investigated. Segregation analysis was based upon a Markov Chain Monte Carlo (Gibbs sampling) approach as implemented in the program BUGS v0.6. All models included adjustment for age, height, tobacco smoke exposure, and gender. The segregation of total IgE levels, blood eosinophil counts, and dose-response slope (DRS) of methacholine challenge were all consistent with major loci at which a recessive allele acted to increase or decrease the phenotype. The respective estimated frequencies of the recessive alleles were 68% (total IgE), 10% (blood eosinophil count), and 27% (DRS). Extensive modelling suggested that the major loci controlling total serum IgE levels, blood eosinophil counts, and airway responsiveness represent different genes. These data provide evidence, for the first time, of the existence of at least 3 distinct genetic pathways involving major gene effects on physiological traits closely associated with asthma. These results have implications for gene discovery programs. PMID- 11255245 TI - Genome-wide linkage analysis of blood pressure in Mexican Americans. AB - The genetic mechanisms that control variation in blood pressure level are largely unknown. One of the first steps in understanding those mechanisms is the localization of the genes that have a significant effect on blood pressure. We performed genome scans of systolic (SBP) and diastolic blood pressure (DBP) on a population-based sample of families in the San Antonio Family Heart Study. A likelihood-based Mendelian model incorporating genotype-specific effects of sex, age, age(2), BMI, and blood pressure (SBP or DBP, as appropriate) as covariates was used to perform two-point lodscore (Z) linkage on 399 polymorphic markers. Results showed that the genotype-specific covariate effects were highly significant for both SBP and DBP. Linkage results showed that a quantitative trait locus (QTL) influencing DBP was significantly linked to D2S1790 (Z = 3.92, theta = 0.00) and showed suggestive linkage to D8S373 (Z = 1.92, theta = 0.00). A QTL influencing SBP showed suggestive linkage to D21S1440 (Z = 2.82, theta = 0.00) and D18S844 (Z = 2.09, theta = 0.11). Without the genotype-specific effects in the model, the linkage to D2S1790 was not even suggestive (Z = 1.33, theta = 0.09); thus genotype-specific modeling was crucial in detecting this linkage. A comparison with linkage studies based in other populations showed that the significant linkage to D2S1790 has been replicated at the same marker in the Quebec Family Study. The replicated significant linkage at D2S1790 may begin to establish the locations of the genes that significantly affect blood pressure across several human ethnic groups. PMID- 11255247 TI - On the total expected study cost in two-stage genome-wide search designs for linkage analysis using the mean test for affected sib pairs. PMID- 11255246 TI - Genetic epidemiology of visceral leishmaniasis in northeastern Brazil. AB - Familial clustering of disease, racial differences in asymptomatic:disease ratios, and studies of mice all point to a genetic component for disease susceptibility in visceral leishmaniasis. Analysis of 87 multi-case pedigrees (824 individuals; 138 nuclear families) from a region of northeastern Brazil endemic for Leishmania chagasi demonstrates a high relative risk ratio (lambda(2S) = 34) to further siblings of affected sibling pairs. Complex segregation analysis using POINTER and COMDS show that all single locus models, as well as polygenic and multifactorial models, provide a significantly (P < 0.001) better fit to the data than a sporadic model. Of the genetic models, the general single locus model was not significantly different from additive or dominant single locus models, all of which gave a gene frequency for the putative disease susceptibility allele of approximately 0.002. The general single locus model was strongly favored (P < 0.001) over a recessive single gene model. Using POINTER, polygenic and multifactorial models were clearly rejected (P < 0.001 in all cases) in favor of the general single locus model. Using COMDS, the analysis was extended to consider two locus models. Results under a general two-locus model did not differ significantly from the dominant, additive, or general single locus models. Under this model, one locus was estimated at a gene frequency of 0.0017, i.e., in the same range as the disease susceptibility locus for the most favored single gene models, with the second locus at a much lower frequency of 0.0002. Hence, the data support the hypothesis that a single major gene may be important in determining disease susceptibility in this population. To identify the gene(s) involved, a genome scan with replication using two subsets of these larger pedigrees with power to detect linkage is in progress. PMID- 11255248 TI - Mutation of the homologue of GDP-mannose pyrophosphorylase alters cell wall structure, protein glycosylation and secretion in Hansenula polymorpha. AB - A Hansenula polymorpha mutant with enhanced ability to secrete a heterologous protein has been isolated. The mutation defines a gene, designated OPU24, which encodes a protein highly homologous to GDP-mannose pyrophosphorylase Psa1p/Srb1p/Vig9p of Saccharomyces cerevisiae and CaSrb1p of Candida albicans. The opu24 mutant manifests phenotypes similar to those of S. cerevisiae mutants depleted for GDP-mannose, such as cell wall fragility and defects in N- and O glycosylation of secreted proteins. The influence of the opu24 mutation on endoplasmic reticulum-associated protein degradation is discussed. The GenBank Accession No. for the OPU24 sequence is AF234177. PMID- 11255249 TI - Yeast 2 microm plasmid copy number is elevated by a mutation in the nuclear gene UBC4. AB - The copy number of the Saccharomyces cerevisiae endogenous 2 microm plasmid is under strict control to ensure efficient propagation to the daughter cell without significantly reducing the growth rate of the mother or the daughter cell. A recessive mutation has been identified that resulted in an elevated but stable 2 microm plasmid copy number, which could be complemented by a genomic DNA clone containing the UBC4 gene, encoding an E2 ubiquitin-conjugating enzyme. A ubc4::URA3 deletion resulted in the same elevated 2 microm plasmid copy number. An analysis of the endogenous 2 microm transcripts revealed that the steady-state abundance of REP1, REP2, FLP and RAF were all increased 4-5-fold in the mutant. Analysis of the mutant ubc4 allele identified a single base pair mutation within the UBC4 coding region, which would generate a glutamic acid to lysine amino acid substitution within a region of conserved tertiary structure located within the first alpha-helix of Ubc4p. These investigations represent the first molecular characterization of a mutation within a Saccharomyces cerevisiae nuclear gene shown to affect 2 microm steady-state plasmid copy number and implicate the ubiquitin-dependent proteolytic pathway in host control of 2 microm plasmid copy number. PMID- 11255250 TI - Regulation of the expression of endopolygalacturonase gene PGU1 in Saccharomyces. AB - Previous work in our laboratory has shown that Saccharomyces bayanus strain SCPP is the only reported yeast expressing the three types of pectolytic enzymes: pectin esterases, pectin lyases and polygalacturonases. One of these enzymes, the endopolygalacturonase (endoPG), hydrolyses plant-specific polysaccharide pectin. The endoPG encoding gene (PGU1) is also present in Saccharomyces cerevisiae. It has been shown that this endoPG is required for the development of pseudohyphae. Using genomic DNA, the PGU1-1 and PGU1-2 promoters of these strains have been amplified and used to construct gene fusions with the beta-galactosidase gene. On the basis of beta-galactosidase measurements, we compared the expression of both promoters in different environmental conditions in order to identify their modulation. We have shown that the PGU1 gene is upregulated by the presence of the pectin and the product resulting from endopolygalacturonase activity. Moreover, expression of the PGU1 is also enhanced under respiratory and filament formation conditions. PMID- 11255251 TI - In situ localization of beta-glucans in the cell wall of Schizosaccharomyces pombe. AB - The chemical composition of the cell wall of Sz. pombe is known as beta-1,3 glucan, beta-1,6-glucan, alpha-1,3-glucan and alpha-galactomannan; however, the three-dimensional interactions of those macromolecules have not yet been clarified. Transmission electron microscopy reveals a three-layered structure: the outer layer is electron-dense, the adjacent layer is less dense, and the third layer bordering the cell membrane is dense. In intact cells of Sz. pombe, the high-resolution scanning electron microscope reveals a surface completely filled with alpha-galactomannan particles. To better understand the organization of the cell wall and to complement our previous studies, we set out to locate the three different types of beta-glucan by immuno-electron microscopy. Our results suggest that the less dense layer of the cell wall contains mainly beta-1,6 branched beta-1,3-glucan. Occasionally a line of gold particles can be seen, labelling fine filaments radiating from the cell membrane to the alpha galactomannan layer, suggesting that some of the radial filaments contain beta 1,6-branched beta-1,3-glucan. beta-1,6-glucan is preferentially located underneath the alpha-galactomannan layer. Linear beta-1,3-glucan is exclusively located in the primary septum of dividing cells. beta-1,6-glucan only labels the secondary septum and does not co-localize with linear beta-1,3-glucan, while beta 1,6-branched beta-1,3-glucan is present in both septa. Linear beta-1,3-glucan is present from early stages of septum formation and persists until the septum is completely formed; then just before cell division the label disappears. From these results we suggest that linear beta-1,3-glucan is involved in septum formation and perhaps the separation of the two daughter cells. In addition, we frequently found beta-1,6-glucan label on the Golgi apparatus, on small vesicles and underneath the cell membrane. These results give fresh evidence for the hypothesis that beta-1,6-glucan is synthesized in the endoplasmic reticulum-Golgi system and exported to the cell membrane. PMID- 11255252 TI - Use of Saccharomyces cerevisiae in the identification of novel transcription factor DNA binding specificities. AB - Members of the steroid/hormone nuclear receptor superfamily regulate target gene transcription via recognition and association with specific cis-acting sequences of DNA, called hormone response elements (HREs). The identification of novel HREs is fundamental to understanding the physiological function of nuclear receptor mediated signalling pathways. A number of these receptors are transcriptionally active, or can be induced to an active state, when expressed in the yeast strain Saccharomyces cerevisiae. This aspect of nuclear receptor activity was used to screen random rat genomic DNA fragments for their ability to function as a HRE for the farnesoid X-activated receptor (FXR). An isolated genomic fragment mediated FXR transcriptional activation without the co-expression of the retinoid X receptor (RXR), a receptor previously thought to be an obligate heterodimer partner for FXR function. This genomic sequence of DNA contained a pair of highly conserved HRE half-sites arranged in an everted orientation and separated by 3 bp (ER3). Furthermore, it was located 240 bp from a highly conserved TATA box motif. A minimal ER3 sequence of DNA was further demonstrated to function as a FXR HRE and was bound in vitro by FXR-expressing yeast extracts. Using RT-PCR, an expressed mRNA fragment was identified within an 8 kb region downstream of the putative TATA box motif. This sequence of DNA was observed to bear homology to a cDNA found in mouse blastocyst. These findings define a novel FXR DNA binding specificity but, more importantly, these data suggest that this strategy might be universally applied to any transcription system that can be reconstituted in yeast. PMID- 11255254 TI - Construction of FLAG and histidine tagging vectors for Schizosaccharomyces pombe. AB - Schizosaccharomyces pombe is becoming an increasingly popular model system for investigating important cellular processes. To facilitate detection, purification and functional studies of Sz. pombe gene products, we constructed two tagging expression vectors for use in Sz. pombe. These vectors allow proteins to be expressed ectopically as fusion proteins with a FLAG epitope and six histidine residue tags attached to their N-terminus or C-terminus. The function and applicability of these vectors were examined and the results are shown using the N-terminal tagging vector encoding Sfc6p, a subunit of the Sz. pombe RNA polymerase III general transcription factor, TFIIIC. PMID- 11255253 TI - Cloning and characterization of the Hansenula polymorpha homologue of the Saccharomyces cerevisiae MNN9 gene. AB - A gene homologous to Saccharomyces cerevisiae MNN9 has been cloned and characterized in the methylotrophic yeast Hansenula polymorpha. This gene was cloned from a H. polymorpha genomic DNA library using the S. cerevisiae MNN9 gene as a probe. The H. polymorpha MNN9 homologue (HpMNN9) contained a 1062 bp open reading frame encoding a predicted protein of 354 amino acids. The deduced amino acid sequence showed 58% and 51% identity, respectively, with the S. cerevisiae and Candida albicans Mnn9 proteins. Disruption of HpMNN9 leads to phenotypic effects suggestive of cell wall defects, including detergent sensitivity and hygromycin B sensitivity. The hygromycin B sensitivity of S. cerevisiae mnn9 null mutant was complemented in the presence of the HpMNN9 gene. The DNA sequence of the H. polymorpha homologue has been submitted to GenBank with the Accession No. AF264786. PMID- 11255256 TI - Disruption and basic phenotypic analysis of six novel genes from the right arm of chromosome XII of Saccharomyces cerevisiae. AB - Six open reading frames (ORFs) of unknown function from the right arm of Saccharomyces cerevisiae chromosome XII were deleted in two genetic backgrounds by disruption cassettes with regions of short flanking homology. This work was carried out within the framework of the EUROFAN consortium. The SFH disruption cassettes, obtained by PCR, were made by amplification of the kanMX marker module with oligonucleotides containing approximately 40 bp of homology to either the promoter or translation terminator regions of the relevant ORF. Transformants resistant to geneticin (G418) were selected. The SFH disruption cassettes were cloned into a bacterial vector. Each cognate gene was also cloned into a yeast centromeric plasmid. Sporulation and tetrad analysis of the disrupted heterozygous strains revealed that ORF YLR153c (now known as ACS2) is essential. Basic phenotypic analysis was performed on haploid deletants of both mating types of the five non-essential ORFs, YLR082c (now known as SRL2), YLR149c, YLR151c, YLR152c and YLR154c. Plate growth tests on different media at 15 degrees C, 30 degrees C and 37 degrees C did not reveal any significant differences between parental and mutant cells. Mating and sporulation efficiencies were not affected in any of the viable disruptants as compared to wild-type cells. PMID- 11255255 TI - Plasmids with the Cre-recombinase and the dominant nat marker, suitable for use in prototrophic strains of Saccharomyces cerevisiae and Kluyveromyces lactis. AB - Two plasmids are described which can be used to remove the "loxP-markerMX-loxP" cassettes in strains lacking the ura3 mutation. Both contain the Cre-recombinase under control of the GAL1 promoter and the natMX cassette with the dominant marker nat, which gives yeasts resistance to the antibiotic ClonNat. pNatCre contains ARSH and CEN6 for maintenance in Saccharomyces cerevisiae. pKlNatCre has a Kluyveromyces lactis replication origin and centromere in addition. PMID- 11255257 TI - Current awareness on yeast. AB - In order to keep subscribers up-to-date with the latest developments in their field, this current awareness service is provided by John Wiley & Sons and contains newly-published material on yeasts. Each bibliography is divided into 10 sections. 1 Books, Reviews & Symposia; 2 General; 3 Biochemistry; 4 Biotechnology; 5 Cell Biology; 6 Gene Expression; 7 Genetics; 8 Physiology; 9 Medical Mycology; 10 Recombinant DNA Technology. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted. (4 weeks journals - search completed 3rd Jan. 2001) PMID- 11255258 TI - Atobe J. H., Hirata M. H., Hoshino-Shimizu S., Schmal M. R., Mamizuka E. M. One step heminested PCR for amplification of Neisseria meningitidis DNA in cerebrospinal fluid. J Clin Lab Anal 14:193-199, 2000. AB - On page 195, in Fig. 1, the labels of boxes RW01 and DG74 are switched. The label RW01 in the right box should read DG74 (1540-1521 position), and the label DG74 in the left box should read RW01 (1189-1170 position). Also in Fig. 1, the primer DG74 listed in the "Primers" box above the legend should read 5' AGGAGGTGATCCAACCGCA-3', and not 5'-AGGAGGTGATCCAACGCGA-3' as printed. The publisher regrets the error. PMID- 11255259 TI - Expression of RFG/ELE1alpha/ARA70 in normal and malignant prostatic epithelial cell cultures and lines: regulation by methylation and sex steroids. AB - RET fused gene (RFG)/ELE1alpha/androgen receptor-associated protein 70(ARA70) was first found to be involved in the activation of the RET proto-oncogene in thyroid neoplasm and has recently been shown to be a ligand-dependent transcriptional coregulator for androgen receptor (AR). The functionality of RFG/ELE1alpha/ARA70 remains controversial, and little is known about factors regulating its expression in the prostate. Of significant interest is whether this molecule is involved in prostate carcinogenesis. Using reverse transcriptase-polymerase chain reaction semiquantitation, we compared RFG/ELE1alpha/ARA70 mRNA levels in four prostate cancer cell lines (LNCaP, TSU-Pr1, DU-145, and PC-3) with those found in primary cultures of normal prostatic epithelial cells (PrECs). In addition, we examined the effects of androgen and antiandrogen, estrogen and antiestrogen, and a demethylating agent on RFG/ELE1alpha/ARA70 mRNA expression levels in AR- and AR+ PC-3 cells. Reduced levels of RFG/ELE1alpha/ARA70 message were observed in all four prostate cancer cell lines when compared with normal PrECs in primary cultures. RFG/ELE1alpha/ARA70 mRNA levels in PC-3 cells, which express both estrogen receptor subtypes, were upregulated by 17beta-estradiol and inhibited by the antiestrogen ICI-182780. In PC-3(AR+) cells, which were genetically engineered to express AR, exposure to androgen upregulated RFG/ELE1alpha/ARA70 mRNA expression, whereas treatment with 4-hydroxyflutamide lowered expression of this transcript. Furthermore, treatment of DU-145 cells, which did not express RFG/ELE1alpha/ARA70 transcripts, with a demethylating agent reactivated transcription of this gene. Polymerase chain reaction analyses of monochromosomal human-rodent hybrid panels localized a putative RFG/ELE1alpha/ARA70 isoform on human chromosome 5q31.1-31.2. In summary, we identified sex hormones and DNA hypermethylation as regulators of RFG/ELE1alpha/ARA70 expression in prostate cancer cells. In addition, we found reduced levels of RFG/ELE1alpha/ARA70 expression in prostate cancer cell lines when compared with expression levels in normal PrECs in culture. These findings suggest that RFG/ELE1alpha/ARA70 may be involved prostate carcinogenesis and that it may serve as a key mediator of estrogen-androgen synergism. PMID- 11255260 TI - Fifteenth Aspen Cancer Conference: mechanisms of toxicity, carcinogenesis, and cancer prevention. PMID- 11255262 TI - Promotion of S-phase entry and cell growth under serum starvation by SAG/ROC2/Rbx2/Hrt2, an E3 ubiquitin ligase component: association with inhibition of p27 accumulation. AB - The sensitive-to-apoptosis gene (SAG) was initially identified as a redox inducible, apoptosis-protective protein and subsequently found to be the second family member of regulator of cullins (ROC)/RING box protein (Rbx)/Hrt, which acts as a component of E3 ubiquitin ligase. We report here that SAG promoted cell growth under serum starvation. Microinjection of SAG mRNA into quiescent NIH/3T3 cells induced S-phase entry as determined by [(3)H]-thymidine incorporation. Likewise, overexpression of SAG by either adenovirus infection of immortalized human epidermal keratinocytes (Rhek-1) or DNA transfection of SY5Y human neuroblastoma cells induced cell proliferation under serum starvation. Because cyclin-dependent kinase inhibitors (CKIs), including p21, p27, and p57, are degraded through the ubiquitin pathway, we tested whether SAG-induced cell growth is associated with CKI degradation. Although there was no significant difference in the levels of p21 and p57 between the vector controls and SAG-overexpressing cells, serum starvation induced 10- to 18-fold accumulation of p27 in control Rhek-1 cells. Accumulation of p27 was remarkably inhibited (only 2 to 5-fold) in SAG-infected cells. Inhibition of p27 accumulation was also observed in stably SAG-overexpressing SY5Y cells. Significantly, SAG-associated inhibition of p27 accumulation was largely abolished by the treatment with a proteasome inhibitor. In vivo binding of SAG and Skp2, an F-box protein that promotes p27 ubiquitination, was detected, and the binding was enhanced in SAG-overexpressing cells grown under serum starvation. Thus, SAG-induced growth with serum withdrawal appears to be associated with SAG-mediated p27 degradation. Mol. Carcinog. 30:37-46, 2001. PMID- 11255263 TI - Involvement of Fas receptor and not tumor necrosis factor-alpha receptor in ultraviolet-induced activation of acid sphingomyelinase. AB - Fas receptor and tumor necrosis factor receptor-1 (TNFR1) mediate the activation of acid sphingomyelinase (ASMase), which catalyzes the hydrolysis of sphingomyelin to ceramide. Ceramide acts as a second messenger in mediating cell growth, differentiation, stress response, and apoptosis. Ultraviolet (UV) irradiation induces Fas receptor and TNFR1 aggregation. However, the roles of Fas receptor and TNFR1 in mediating UV-induced ASMase activation have not been explored. In this report, we demonstrate that Fas receptor, not TNFR1, mediated UV-induced activation of ASMase. Our data indicate that ASMase activity was not induced with UV irradiation but by TNFalpha in MCF-7 cells that expressed low levels of Fas receptor. In contrast, ASMase was activated by UV irradiation or TNFalpha treatment in Fas stably transfected MCF-7 cells. Immunofluorescence staining of TNFR1 on MCF-7 cells showed that TNFR1 was aggregated after treatment with UV irradiation or TNFalpha. However, UV-induced aggregation of TNFR1 did not lead to induction of ASMase activity. These results suggest that Fas receptor aggregation is solely responsible for UV-induced activation of ASMase. Further, with the use of BJAB and dominant-negative Fas-associated death domain-containing protein (FADD) stably transfected BJAB cells, we demonstrated that dominant negative FADD partly inhibited UV-induced ASMase activation. Our results suggest that FADD is involved in UV-induced and Fas-mediated signaling pathways for activation of ASMase. Mol. Carcinog. 30:47-55, 2001. PMID- 11255261 TI - Somatic INK4a-ARF locus mutations: a significant mechanism of gene inactivation in squamous cell carcinomas of the head and neck. AB - The INK4a-ARF locus is located on human chromosome 9p21 and is known to encode two functionally distinct tumor-suppressor genes. The p16(INK4a) (p16) tumor suppressor gene product is a negative regulator of cyclin-dependent kinases 4 and 6, which in turn positively regulate progression of mammalian cells through the cell cycle. The p14(ARF) tumor-suppressor gene product specifically interacts with human double minute 2, leading to the subsequent stabilization of p53 and G(1) arrest. Previous investigations analyzing the p16 gene in squamous cell carcinomas of the head and neck (SCCHNs) have suggested the predominate inactivating events to be homozygous gene deletions and hypermethylation of the p16 promoter. Somatic mutational inactivation of p16 has been reported to be low (0-10%, with a combined incidence of 25 of 279, or 9%) and to play only a minor role in the development of SCCHN. The present study examined whether this particular mechanism of INK4a/ARF inactivation, specifically somatic mutation, has been underestimated in SCCHN by determining the mutational status of the p16 and p14(ARF) genes in 100 primary SCCHNs with the use of polymerase chain reaction technology and a highly sensitive, nonradioactive modification of single stranded conformational polymorphism (SSCP) analysis termed "cold" SSCP. Exons 1alpha, 1beta, and 2 of INK4a/ARF were amplified using intron-based primers or a combination of intron- and exon-based primers. A total of 27 SCCHNs (27%) exhibited sequence alterations in this locus, 22 (22%) of which were somatic sequence alterations and five (5%) of which were a single polymorphism in codon 148. Of the 22 somatic alterations, 20 (91%) directly or indirectly involved exon 2, and two (9%) were located within exon 1alpha. No mutations were found in exon 1beta. All 22 somatic mutations would be expected to yield altered p16 proteins, but only 15 of them should affect p14(ARF) proteins. Specific somatic alterations included microdeletions or insertions (nine of 22, 41%), a microrearrangement (one of 22, 5%), and single nucleotide substitutions (12 of 22, 56%). In addition, we analyzed the functional characteristics of seven unique mutant p16 proteins identified in this study by assessing their ability to inhibit cyclin dependent kinase 4 activity. Six of the seven mutant proteins tested exhibited reduced function compared with wild-type p16, ranging from minor decreases of function (twofold to eightfold) in four samples to total loss of function (29- to 38-fold decrease) in two other samples. Overall, somatic mutation of the INK4a/ARF tumor suppressor locus, resulting in functionally deficient p16 and possibly p14(ARF) proteins, seems to be a prevalent event in the development of SCCHN. Mol. Carcinog. 30:26-36, 2001. PMID- 11255264 TI - Cutaneous squamous cell carcinoma and p53 codon 72 polymorphism: a need for screening? AB - The association between human papillomavirus (HPV)-associated cervical cancer and cutaneous squamous cell carcinoma and codon 72 polymorphism in the p53 gene is not unequivocal. Especially, it is not known whether carriers of the arginine form have an increased risk of cancer that necessitates screening. The alternative is that the polymorphism is a tumor marker instead of a risk factor. We set out a case-control study to determine the risk of squamous cell carcinoma of the skin in individuals with the p53 codon 72 arginine genotype in order to establish the possible need for screening. The distribution of the different p53 codon 72 genotypes was examined in 86 subjects with a history of cutaneous squamous cell carcinoma and in 168 controls. Additionally, 121 subjects who had had histologically proven basal cell carcinoma and 108 subjects who had had non familial malignant melanoma were tested. p53 polymorphism was evaluated by polymerase chain reaction (PCR) using DNA samples from peripheral blood lymphocytes. In a subgroup of patients with squamous cell carcinoma and controls, the presence of epidermodyplasia verruciformis human papillomavirus (EV-HPV) DNA was determined in plucked eyebrow hair. Differences in the distributions of the genotypes among cases and controls were calculated, and univariate and multivariate analyses were performed to assess the risk to develop cutaneous squamous cell carcinoma in the presence of the p53 codon 72 arginine genotype. Frequency distributions of the three different genotypes (homozygous for the arginine allele, heterozygous for the two alleles, and homozygous for the proline allele) were similar among the squamous cell carcinoma group and the control group: 47.1%, 46.0% and 6.9% versus 47.8%, 45.8% and 6.4%, respectively. Statistical analysis showed no significant differences between these groups. In patients with squamous cell carcinoma and controls who harbored EV-HPV DNA in their plucked eyebrow hair, similar results were obtained. The distributions of the p53 codon 72 genotypes in the basal cell carcinoma and malignant melanoma group were also not significantly different from the control group. p53 codon 72 arginine homozygosity does not appear to represent a significant risk factor for cutaneous squamous cell carcinoma and screening seems not to be indicated. Mol. Carcinog. 30:56-61, 2001. PMID- 11255265 TI - Elevated expression of SAG/ROC2/Rbx2/Hrt2 in human colon carcinomas: SAG does not induce neoplastic transformation, but antisense SAG transfection inhibits tumor cell growth. AB - Sensitive-to-apoptosis gene (SAG)/regulator of cullins (ROC)2/Rbx2/Hrt2 is a newly identified component of SCF E3 ubiquitin ligase that controls cell-cycle progression by promoting ubiquitination and degradation of cell-cycle inhibitors. We recently found that SAG protects cells from apoptosis induced by redox agents, promotes S-phase entry and cell growth under serum starvation, and is required for yeast growth. In the present study, we report that the SAG protein level was elevated in six of 10 human colon carcinoma tissues (60%) as compared with adjacent normal tissues from the same patient. SAG overexpression in preneoplastic cells in a JB6 tumor promotion-and-progression model did not induce neoplastic transformation, and SAG overexpression in NIH/3T3 cells did not induce transforming foci formation, suggesting that SAG is not a dominant oncogene. However, when DLD-1 human colon carcinoma cells were transfected with antisense SAG, monolayer growth was significantly inhibited, as shown by a decreased number of stable colonies in the plate after normalization with transfection efficiency. Stable clones that expressed antisense SAG showed a 50% decrease in their ability to form colonies when grown in soft agar versus clones that did not express antisense SAG. We found an inverse correlation in four of 10 tumors between the levels of SAG and p27, a cyclin-dependent kinase inhibitor. We concluded that SAG is not causally related to cellular transformation, but its overexpression may be important for the maintenance of tumor cell phenotype. Therefore, targeting SAG expression may have therapeutic value in cancer treatment. Mol. Carcinog. 30:62 70, 2001. PMID- 11255266 TI - The microsomal epoxide hydrolase Tyr113His polymorphism: association with risk of ovarian cancer. AB - Functional significance has been demonstrated in vitro for the exon 3 T-->C Tyr113His amino acid substitution polymorphism of the microsomal epoxide hydrolase (EPHX) gene. The higher activity or fast TT genotype was previously reported to be associated with an increased risk of ovarian cancer, and this association may reflect enhanced activation of endogenous or exogenous substrates to more reactive and mutagenic derivatives. Components of cigarette smoke are examples of exogenous substrates subject to such bioactivation, and smoking exposure may thus modify the risk associated with the EPHX polymorphism. We examined 545 cases of epithelial ovarian cancer and 287 unaffected controls for this EPHX T-C genetic variant to investigate whether, in the Australian population, the TT genotype was associated with (i) specific ovarian tumor characteristics; (ii) risk of ovarian cancer, overall or for specific subgroups; and (iii) risk of ovarian cancer in smokers specifically. Genotyping was carried out using the Perkin-Elmer ABI Prism 7700 Sequence Detection System for fluorogenic polymerase chain reaction allelic discrimination. Stratification of the ovarian cancer cases according to tumor behavior (low malignant potential or invasive), grade, stage, and p53 immunohistochemical status failed to show any heterogeneity with respect to the genotype defined by the EPHX polymorphism. There was a suggestion of heterogeneity with respect to histologic subtype (P=0.03), largely due to a decreased frequency of the TT genotype in endometrioid tumors. EPHX genotype distribution did not differ significantly between unaffected controls and ovarian cancer cases (overall, low malignant potential, or invasive) either overall or after stratification by smoking status. However, the TT genotype was associated with a decreased risk of invasive ovarian cancer of the endometrioid subtype specifically (age-adjusted odds ratio=0.38, 95% confidence interval=0.17-0.87). The results suggest that the proposed EPHX mediated bioactivation of components of cigarette smoke to mutagenic forms is unlikely to be involved in the etiology of ovarian cancer in general but that a greater rate of EPHX-mediated detoxification may decrease the risk of endometrioid ovarian cancer. Mol. Carcinog. 30:71-78, 2001. PMID- 11255267 TI - Immune tolerance in hemophilia and treatment of hemophiliacs with an inhibitor. PMID- 11255268 TI - Mouse plasmacytoma: an experimental model of human multiple myeloma. AB - BACKGROUND AND OBJECTIVES: There is no ideal animal model for human multiple myeloma (MM). All the models resemble the human disease in some respect, but none of them fulfils all the criteria of a perfect animal model. EVIDENCE AND INFORMATION SOURCES: The pristane oil (2,6,10,12-tetramethylpentadecane)-induced mouse plasmacytoma (MPC) model is the most widely used and accepted model and has provided the most data on plasmacytomagenesis so far. This model gives the opportunity to study the role of c-myc dysregulations, the mechanisms leading to cytogenetic changes involving Ig genes, the role of chronic inflammatory factors, the role of interleukin-6 (IL-6), insulin-like growth factor-I, prostaglandins, as well as signal transduction pathways in the neoplastic process. Therapeutic agents have been successfully tested. Although MPC growth is usually restricted to the peritoneal environment, intraperitoneal injection of MPC cell suspensions can reproduce the disseminated characteristics of the human disease in recipients. The IL-6 transgene and knockout models are important tools for clarifying the role of IL-6 in the pathogenesis of MM. Transgenic mice and retroviral gene transfer facilitate the study of oncogenes in neoplastic transformation. Spontaneous development of plasmacytomas in C57BL/ KaLwRij aging mice has several advantages, mainly because the disseminated growth, the typical bone lesions and renal involvement resemble, in part, the human disease. Furthermore, this model has already proved useful in studies on the effect of bisphosphonate in the treatment of bone disease in MM. The severe combined immunodeficiency (SCID) mouse model is also very attractive. A disseminated-like disease can be reproduced in this model. Multiple osteolytic bone lesions and bone marrow involvement are generated, and conventional drugs applied in the treatment of human multiple myeloma have proven to be effective. Nevertheless, the immune system of SCID mice basically differs from that of a MM patient. PERSPECTIVES: Taken together, all these models have contributed to our understanding of MM, but demonstrate the opportuness of developing a more appropriate model of the human disease. PMID- 11255269 TI - Quantitative analysis of Fas and bcl-2 expression in hematopoietic precursors. AB - BACKGROUND AND OBJECTIVES: We investigated the expression of bcl-2 and CD95 (Apo1 /Fas) on CD34+ cells obtained from bone marrow (BM), mobilized peripheral blood (MPB), and umbilical cord blood (UCB) samples. The expression of bcl-2 and Fas was then compared with that of other markers usually associated with immaturity; functional tests using the agonistic antibody anti- Fas CH11 were also carried out. DESIGN AND METHODS: The analysis was performed by flow cytometry on purified CD34+ cells in a three (CD95 PE, CD34 APC and CD71 FITC) and in a four (CD38 PE, HLA-DR PerCP, CD34 APC and bcl-2 FITC) fluorescence assay. RESULTS: The results were expressed as mean fluorescence index (MFI); bcl-2 expression was significantly higher (p < 0.001) in BM (3.73 +/- 0.63) than in MPB (2.47 +/- 0.39) and UCB (2.38 +/- 0.58); Fas was significantly less expressed (p < 0.001) in UCB (1.27 +/- 0.78) than in MBP (3.63 +/- 2.19) and BM (4.56 +/- 1.69). CD34 expression was significantly (p < 0.001) brighter in UCB compared to in MBP and BM, while CD38 and CD71 were significantly (p = 0.005 and p < 0.001, respectively) more expressed in BM than in MPB and UCB. Fas values were directly correlated to CD38; both Fas and bcl-2 were directly related to CD71 and inversely to CD34. Culture assays showed that hematopoietic precursor cells from BM, MPB and UCB had a low susceptibility to undergo Fas-mediated apoptosis. INTERPRETATION AND CONCLUSIONS: In conclusion, bcl-2 and Fas are less expressed in UCB than in MPB and BM; early hematopoietic precursor cells are relatively resistant to CD95-triggered apoptosis; the observed correlation between Fas/bcl-2 and markers of immaturity suggests that they may be determinants of commitment in early hematopoietic precursors. PMID- 11255270 TI - Reticulocyte transferrin receptor (TfR) expression and contribution to soluble TfR levels. AB - BACKGROUND AND OBJECTIVES: Transferrin receptor (TfR) expression in erythroid cells is regulated by a number of factors, including iron status and erythropoietin (Epo) stimulation. However, the impact of these factors on reticulocyte TfR expression in vivo has never been studied. A soluble form of TfR (sTfR) is present in serum in proportion to the mass of cellular TfR. Although sTfR shedding by reticulocytes and erythroblasts has been demonstrated in vitro, the contribution of reticulocyte TfR to serum sTfR has never been evaluated in vivo. DESIGN AND METHODS: We measured directly the total number of reticulocyte TfR in normal rats of different age and iron status, as well as in animals experiencing various conditions and treatments aimed at altering erythropoietic activity and iron status, including rHuEpo therapy, hemolytic anemia, phlebotomies, hypertransfusions, thiamphenicol-induced red cell aplasia or inflammation. In addition, we examined the impact of repeated hypertransfusions with normal, reticulocyte-poor and reticulocyte-rich blood on serum sTfR levels. RESULTS: The number of TfR molecules per reticulocyte was around 50,000 in young rats but was around 100,000 in older animals. These values remained constant in most conditions and in particular were not influenced by iron supplementation or iron overload. However, functional iron deficiency as well as rHuEpo therapy resulted in increased reticulocyte TfR expression. In addition, TfR numbers in reticulocytes were elevated in the early phase of recovery after acute hemolysis or red cell aplasia but normalized soon after. Hypertransfusion experiments clearly demonstrated that reticulocytes can contribute substantially to sTfR levels in vivo. INTERPRETATION AND CONCLUSIONS: TfR numbers are regulated in vivo by the same factors as in vitro, in particular iron deficiency and erythropoietin stimulation. Circulating reticulocytes contribute significantly to serum sTfR levels. PMID- 11255271 TI - Real-time quantification of different types of bcr-abl transcript in chronic myeloid leukemia. AB - BACKGROUND AND OBJECTIVES: The most common translocation in chronic myeloid leukemia (CML) t(9;22) (q34;q22) produces the BCR/ABL fusion gene. We set up and evaluated a rapid and reliable real-time reverse-transcription-polymerase chain reaction (RT-PCR) approach using TaqMan technology for detection and quantification of bcr-abl transcripts in CML patients at diagnosis and during therapy. DESIGN AND METHODS: A pair of primers and probe complementary to ABL exon 2 were designed, enabling detection of the most frequent bcr-abl transcripts, and also of the normal ABL-Ia transcript as an internal control. Conditions were established to amplify less than 1(-10) target molecules/reaction and detect one CML cell in 10(6) cells from healthy donors. To determine the utility of the assay, we quantified the bcr-abl/ABL-Ia ratio in 59 bone marrow samples (45 samples with evidence of different Ph+ chromosome percentages and 14 samples in complete cytogenetic remission) from 48 CML patients, 34 of them at diagnosis and 14 in clinical remission (CR). In 14 cases, this ratio was compared with results obtained by a competitive-quantitative RT-PCR/capillary electrophoresis method from contemporary specimens. RESULTS: By real-time RT-PCR, the median value of bcr-abl/ABL-Ia ratio at diagnosis was 15.334 (range 3.3 28.81) and fell to 0.9 (range 0.003-26.1) in CR. The median value of bcr-abl/ABL Ia ratio at cytogenetic remission was 0.7 (range 0.003-2.83). The real-time bcr abl/ABL-Ia ratios correlated with those obtained by competitive RT-PCR (p < 0.0001) and the percentage of Ph+ metaphases (p < 0.0001). The high sensitivity and specificity of the real-time RT-PCR procedure was confirmed in all 14 patients with minimal residual disease. INTERPRETATION AND CONCLUSIONS. We conclude that this real-time RT-PCR procedure is a reliable and sensitive method of monitoring CML patients after therapy, and that the bcr-abl/ABL-Ia ratio correlates strongly with cytogenetic analysis. PMID- 11255272 TI - ALK positive lymphohistiocytic variant of anaplastic large cell lymphoma in an adult. AB - BACKGROUND AND OBJECTIVES: The lymphohistiocytic (LH) variant of anaplastic large cell lymphoma (ALCL) has, for a long time, been considered typical of children and adolescents. The aim of this study is a detailed characterization of a case of this peculiar ALCL subtype affecting an adult patient. DESIGN AND METHODS: A 36-year old male presented with diffuse adenopathy and systemic symptoms (high fever, anorexia, asthenia); a diagnosis of CD30+/ALK+ ALCL, LH variant, was morphologically suspected and corroborated by immunohistochemistry that was crucial for the definitive diagnosis and subtyping. RESULTS: The neoplastic population consisted of cells highly variable in size and shape but more often isolated and largely obscured by a predominant reactive cellular infiltrate of histiocytes and plasma cells. The lymphoma cells exhibited a null non-B non-T antigenic profile, but reacted strongly for the Ber-H2/CD30, EMA, ALKc anti-TIA-1 monoclonal antibodies. The patient underwent chemotherapy plus bone marrow transplantation and, one year after diagnosis, he is well and in complete remission. INTERPRETATION AND CONCLUSIONS: Our findings provide additional evidence that: a) ALK+ lymphoma represents a single disease with a broad spectrum of morphology; b) clinicians and pathologists should be aware of the possible occurrence of LH variant of ALK+ ALCL also in adults in whom a favorable response to therapy may be expected despite systemic disease and an aggressive clinical presentation. PMID- 11255273 TI - Whole-body positron emission tomography using 18F-fluorodeoxyglucose compared to standard procedures for staging patients with Hodgkin's disease. AB - BACKGROUND AND OBJECTIVES: Accurate staging is essential in order to determine appropriate treatment in Hodgkin's disease (HD). (18)F-fluorodeoxyglucose positron emission tomography ((18)F-FDG PET) offers the advantage of metabolic imaging that is largely independent of morphologic criteria. In the present study we evaluated the role of (18)F-FDG PET compared to routine procedures for the staging of patients with HD. DESIGN AND METHODS: Thirty-three patients with HD underwent standard staging procedures (clinical examination, laboratory screening, chest X-ray, computed tomography (CT) of the chest and abdomen and bilateral bone marrow biopsies) and a whole-body (18)F-FDG PET study. In clinical examination, an isolated lymph node > 1 cm or multiple lymph nodes > or = 1 cm in size were considered abnormal. Positive findings at both clinical examination or CT and (18)F-FDG PET were regarded as actual locations of disease. Negative findings with both methods were regarded as true negative (no involvement by HD). In cases of discrepancy, response to treatment and follow-up data were used to assess the overall accuracy of the patient's original evaluation. RESULTS: Completely concordant results in lymph node staging were observed in 20 patients. The two staging procedures indicated complementary information in 1 patient. Conventional staging indicated more pathologic lymph node areas in 6 patients (at least 1 false positive). (18)F-FDG PET showed more sites in 6 patients. The sensitivity of (18)F-FDG PET in detecting all known pathologic lymph nodes was 83% for peripheral lymph nodes, 91% for thoracic lymph nodes and 75% for abdominal and pelvic lymph nodes. Conventional staging procedures and (18)F-FDG PET indicated the same tumor stage in 26 patients. Based on (18)F-FDG PET, downstaging was suggested in 4 patients, including a biopsy-proven case. However in 1 of these cases this was incorrect. (18)F-FDG PET suggested upstaging in 3 patients. Based on conventional staging or (18)F-FDG PET the same treatment strategy was defined in 32 patients. In one patient (18)F-FDG PET downstaged disease extension (stage IIIA-->IIA) that would have suggested radiotherapy as a possible treatment option. INTERPRETATION AND CONCLUSIONS: (18)F-FDG PET provides an easy and efficient whole-body method for the evaluation of patients with HD. (18)F-FDG PET never missed tumor masses >1 cm. (18)F-FDG PET detected additional sites of disease not seen by conventional procedures and identified absence of disease in some sites suspected to be involved. However, in our patients this did not translate into changes in treatment strategy. PMID- 11255274 TI - Serum interleukin-10 levels are an independent prognostic factor for patients with Hodgkin's lymphoma. AB - BACKGROUND AND OBJECTIVES: Interleukin-10 (IL-10) is a pleiotropic cytokine which increases bcl-2 levels and protects cells from steroid or doxorubicin-induced apoptosis. Hodgkin and Reed-Sternberg (HRS) cells bear functional IL-10 receptors. Thus serum IL-10 (sIL-10) might inhibit apoptosis in HRS cells, which could occur as a result of either chemotherapy or the crippled immunoglobulin genes. DESIGN AND METHODS: We determined sIL-10 levels in 122 patients with Hodgkin's lymphoma (HL), treated with ABVD or equivalent regimens with or without radiotherapy, and correlated them with presenting clinical and laboratory features, as well as failure-free survival (FFS) and overall survival. RESULTS: Elevated sIL-10 levels ( > or = 10 pg/mL) were detected in 55 patients (45%), and were correlated with advanced stage and elevated serum b2-microglobulin levels. At 7 years FFS was 85% vs. 63% for patients with normal vs. elevated sIL-10 levels, respectively (p=0.01); overall survival was 97% vs. 73% (p=0.005). Multivariate analysis with Cox's proportional hazards model demonstrated that elevated sIL-10 levels were the strongest independent predictor of FFS, and were also associated with inferior overall survival. INTERPRETATION AND CONCLUSIONS: We conclude that sIL-10 levels are elevated in 45% of patients with HL, and are associated with inferior FFS and overall survival, independently of other established prognostic factors. PMID- 11255275 TI - Fludarabine in combination with cyclophosphamide or with cyclophosphamide plus mitoxantrone for relapsed or refractory low-grade non-Hodgkin's lymphoma. AB - BACKGROUND AND OBJECTIVES: We report the activity of two combinations of fludarabine (FLU), one with cyclophosphamide (FLU/CY) and the second with CY plus mitoxantrone (FLU/CY/MITO). The aim of the study was to evaluate the activity and toxicity of these two schedules in patients with non-Hodgkin's lymphoma (NHL). DESIGN AND METHODS: Twenty-two patients with recurrent low grade non-Hodgkin's lymphoma (LGL) received FLU/CY (FLU 25 mg/m(2) days 1 to 3, CY 300 mg/m(2) days 1 to 3), and 31 patients received FLU/CY/MITO (FLU 25 mg/m(2) days 1 to 3, CY 300 mg/m(2) days 1 to 3, mitoxantrone 10 mg/m(2) day 1). Patients received antibiotic oral prophylaxis during all treatments and growth factors (G-CSF) when grade III granulocytopenia (WHO scale) occurred. RESULTS: Of the 53 patients, 31 achieved complete remission (CR) (58%) and 16 partial remission (PR) (30%). Response was similar in both arms of the study. After 3 courses, 77% of patients who achieved CR showed a complete disappearance of disease. Seventy-nine per cent of patients experienced granulocytopenia. Few patients had fever, all without infection. One patient died with fever of unknown origin three months after completion of six courses of treatment. INTERPRETATION AND CONCLUSIONS: Both treatments were seen to be effective in recurrent low-grade NHL. Antibiotic prophylaxis with G-CSF support seems to reduce treatment-related infection. PMID- 11255277 TI - Low molecular weight heparins prevent thrombin-induced thrombo-embolism in mice despite low anti-thrombin activity. Evidence that the inhibition of feed-back activation of thrombin generation confers safety advantages over direct thrombin inhibition. AB - BACKGROUND AND OBJECTIVES: Thrombin-induced thromboembolism in mice is a model in which the feed-back clotting activation produced by the injected enzyme greatly contributes to fibrin accumulation in lungs and to mortality. Using this model we have previously shown that activated human protein C (aPC), by interrupting endogenous clotting activation at a high level (factors Va and VIIIa), prevents mortality inducing only a minor hemostatic impairment. With the same model we have now compared the antithrombotic and prohemorrhagic effects of two low molecular weight heparins (LMWHs), reviparin and tinzaparin, which are expected to inhibit preferentially the positive feed-back triggered by thrombin (anti Xa activity), with those of unfractionated heparin (UFH) and PEG-hirudin, which inhibit mainly or exclusively thrombin activity (anti IIa activity). DESIGN AND METHODS: Pulmonary thromboembolism was induced in mice by i.v. injection of bovine thrombin (1,000U/kg). Drugs (from 0.12 to 1.2 mg/kg) were given as bolus injection 2 min prior to thrombin challenge and mortality was assessed within 15 min. The bleeding time was assessed by a tail tip transection model. Activated partial thromboplastin time (aPTT), thrombin clotting time (TcT), fibrinogen assay and anti Xa activity determination were performed in citrated plasma from saline- or drug-treated animals. RESULTS: All drugs protected mice from thrombin induced mortality in a dose-dependent way. At comparable antithrombotic dosages, the anti IIa activity generated in plasma (assessed by TcT) was highest with UFH, intermediate with tinzaparin and very low with reviparin. Accordingly, the fibrinogen drop, which is caused mainly by the injected thrombin, was prevented by the heparins to an extent that was fairly well related to their anti IIa activity. aPTT and bleeding time, used as measures of hemorrhagic risk, were markedly more prolonged by UFH than by reviparin. Tinzaparin, instead, had an intermediate effect. Interestingly, PEG-hirudin, at equipotent antithrombotic dosages, caused a prolongation of bleeding time comparable to that observed with UFH. INTERPRETATIONS AND CONCLUSIONS: Our data show that, in our model, drugs acting at a high level of the blood clotting cascade, like LMWHs with a high anti Xa/anti IIa ratio, display a better antithrombotic/prohemorrhagic profile than drugs acting prevalently on thrombin. PMID- 11255276 TI - Efficacy of vinorelbine, epirubicin and prednisone combination regimen in pretreated elderly patients with aggressive non-Hodgkin's lymphoma. AB - BACKGROUND AND OBJECTIVES: To assess the efficacy and toxic profile of the NAEPP protocol, a regimen including vinorelbine, epirubicin and prednisone, in a particularly troublesome subset of patients: pretreated elderly patients with aggressive non-Hodgkin's lymphoma (NHL). DESIGN AND METHODS: From November 1998 to January 2000, 20 pretreated patients who had all relapsed after first-line VNCOP-B chemotherapy were enrolled in a phase II trial and treated with the NAEPP regimen: vinorelbine (25 mg/m(2) i.v. on days 1 and 8), epirubicin (40 mg/m(2) i.v. on days 1 and 8), and prednisone (40 mg/m(2) on days 1 and 8) with granulocyte colony-stimulating factor administered at 5 mg/kg/day on days 2-5 and days 9-12. Chemotherapy was repeated every 4 weeks for a total of 6 cycles. RESULTS: Six (30%) patients achieved complete remission (CR) and 7 (35%) had partial responses (PR), giving an overall response rate of 65%. The response rate was not affected either by type of relapse presentation (nodal versus nodal plus extranodal), presence of bulky disease, or time of relapse. No major toxic effects were recorded. INTERPRETATION AND CONCLUSIONS: These preliminary data suggest that the NAEPP regimen is an effective combination with a low toxicity profile in elderly pretreated patients with aggressive NHL. Further trials using NAEPP as a consolidation phase following first-line treatment are needed to establish the advantage in terms of CR rate and relapse-free survival in these patients. PMID- 11255278 TI - Hematopoietic stem cell transplantation for high-risk adult patients with severe aplastic anemia; reduction of graft failure by enhancing stem cell dose. AB - BACKGROUND AND OBJECTIVES: The main causes of failure after allogeneic hematopoietic stem cell transplantation (HSCT) in patients with severe aplastic anemia (SAA) are graft-versus-host disease (GVHD), infection and graft failure, often exacerbated by large numbers of transfusions and prolonged disease duration before transplant. This study retrospectively analyzes the outcome and factors related to survival or graft failure in high-risk patients with SAA receiving HSCT in our institution. DESIGN AND METHODS: Between January 1995 and December 1999, 40 consecutive adult patients who were multi-transfused (more than 40 units of red blood cells +/- platelets) and/or had a 3 years or longer period prior to transplant were enrolled. Their median age was 27.5 years (range, 16 to 43) and 21 (52.5%) were women. All donors were human leukocyte antigen (HLA)-matched siblings. Before transplant, 29 patients (72.5%) received a course of antithymocyte globulin (ATG) and cyclosporin A (CsA). The median interval from diagnosis to transplant was 59 months (range, 2 to 216). The median number of transfusions was 115 units (range, 10 to 480). All patients received a conditioning regimen of cyclophosphamide, ATG, and procarbazine. Our patients received either bone marrow (BM) alone (n=20) or BM+peripheral blood stem cells (PBSC) (n=20) as a stem cell source. T-cells of PBSC were depleted using the CD34 enrichment method. GVHD prophylaxis consisted of CsA and short-term methotrexate. RESULTS: In the BM+PBSC group, neutrophil recovery to 0.5 x 10(9)/L and platelet recovery to 20 x 10(9)/L were achieved more rapidly than in the BM group (p=0.005 and 0.039, respectively). The incidences of graft failure, grade II to IV acute GVHD, and chronic GVHD were 22.5%, 12.8% and 23.1%, respectively. Graft failure occurred in 2 of 20 patients (10%) receiving BM+PBSC and in 7 of 20 (35%) receiving BM alone (p=0.069). Seven of 9 patients who had graft failure received a booster treatment and recovered normal marrow function. GVHD incidence was comparable between the BM+PBSC and BM groups. Six patients (15%) died from graft failure (n=2), interstitial pneumonia (n=2), cyclophosphamide-induced heart failure (n=1), and chronic GVHD followed by pneumonia (n=1). The Kaplan-Meier estimate of survival was 83.7% with a median follow-up duration of 40.5 months (range 8-67). In multivariate analysis only chronic GVHD adversely influenced survival (p=0.042). INTERPRETATION AND CONCLUSIONS: These results suggest that HSCT is an effective treatment for multi-transfused SAA patients with prolonged disease duration. It is highly possible that the infusion of a large number of stem cells leads to a reduction of graft failure and a faster speed of engraftment. Booster treatment is successful in achieving engraftment in patients with graft failure. PMID- 11255279 TI - Pharmacokinetic study of the new cyclosporine-A formulation (Neoral) in adult allogeneic bone marrow transplant recipients. AB - BACKGROUND AND OBJECTIVES: A major problem encountered during oral cyclosporin-A (CsA) administration to prevent acute graft-versus-host-disease (GVHD) after allogeneic bone marrow transplantation (allo-BMT) is its irregular pharmacokinetics. The aim of this study was to evaluate the pharmacokinetics of Neoral, a new water-free microemulsion formulation of CsA. DESIGN AND METHODS: Eighteen patients aged over 18 were enrolled into the study. When able to eat normally after allo-BMT, patients received CsA orally and after 4 days a 12-hour CsA pharmacokinetic profile was constructed. Three patients received Sandimmune 10 mg/kg/day, 5 patients received Neoral 7.5 mg/kg/day and 10 patients Neoral 5 mg/kg/day. CsA concentration was analyzed on whole blood by high-performance liquid chromatography (HPLC). RESULTS: Neoral showed concentration-time profiles characterized by a smooth and faster rise to the Cmax value compared to that produced by Sandimmune. The comparison between pharmacokinetic parameters obtained in patients receiving Neoral 5 mg/kg/day or 7.5 mg/kg/day showed a proportional increase of the AUC (4776+/-1084 vs. 7746+/-2006 ng/mL h) and C(max) (1027+/-203 vs. 1514+/-231 ng/mL). In all patients to whom 7.5 mg/kg/day of Neoral were given, C(trough) levels were always above the threshold of 200 ng/mL. INTERPRETATION AND CONCLUSIONS: Our data suggest that oral administration of Neoral 7.5 mg/kg/day early after allo-BMT may represent an appropriate dose resulting in adequate CsA C(trough) levels without significant renal toxicity. PMID- 11255280 TI - S65c frequency in Italian patients with hemochromatosis, porphyria cutanea tarda and chronic viral hepatitis with iron overload. PMID- 11255281 TI - Quantification of bcr/abl mRNA expression by a rapid real-time reverse transcription-polymerase chain reaction assay in patients with chronic myeloid leukemia. PMID- 11255283 TI - Flow-cytometric detection of minimal residual disease in adult acute lymphoblastic leukemia. PMID- 11255282 TI - p230 does not always predict a mild clinical course in myeloid malignancies: e19a2 bcr/abl fusion transcript with additional chromosome abnormalities in a patient with acute monoblastic leukemia (M5a). PMID- 11255284 TI - Complex karyotype in one patient with small cell variant of T-prolymphocytic leukemia. Analysis by G-banding and comparative genomic hybridization. PMID- 11255285 TI - Anti-heparin-platelet factor 4 antibodies after cardiopulmonary bypass: role of HLA expression. PMID- 11255286 TI - Preoperative screening: the rationale of measuring APTT in risk assessment. PMID- 11255287 TI - The role of human herpesvirus-6 in delayed engraftment in stem cell transplant patients in China. PMID- 11255288 TI - Indwelling catheter-related central venous thrombosis during bone marrow transplantation. PMID- 11255289 TI - The irreplaceable image: Nail transverse white bands induced by antileukemic chemotherapy. PMID- 11255290 TI - The irreplaceable image: Nails changes in onco-hematologic patients. PMID- 11255291 TI - Management of blood donors with a low level of exposure to vCJD. PMID- 11255292 TI - Severe thrombocytopenia after an infusion of abciximab. PMID- 11255293 TI - Demystyfication versus trivialization of stem cell transplantation. PMID- 11255295 TI - What is the true risk of thrombosis in patients carrying thrombogenic mutations? PMID- 11255296 TI - Thrombocytopenic Purpura after Recombinant Hepatitis B Vaccine. A rare association. PMID- 11255298 TI - Presidential address. XVIII National Conference of the Indian Academy of Pediatrics, February 8, 2001. PMID- 11255297 TI - Current practices in transfusion medicine. PMID- 11255299 TI - Clinical profile of HIV infection. AB - OBJECTIVE: To study the clinical profile of human immunodeficiency virus (HIV) infection in children. DESIGN: Prospective. SETTING: HIV clinic at a pediatric tertiary care center in an urban metropolis. METHODS: From August 1994 onwards, 285 HIV positive children were referred to the HIV clinic. These included those intramural deliveries born to HIV positive mothers, those referred from other centers with a positive HIV ELISA (enzyme-linked immunosorbent assay) test and those screened routinely at our center in view of transfusion dependence and found to be HIV positive. After informed consent from either parent, the HIV status of all referred patients was retested by ELISA. RESULTS: Two hundred and thirteen (74.73%) patients were below the age of five years. Vertical transmission as the route of infection was documented in 247 (86.66%), 33 (11.57%) were infected through blood and in 5 (1.75%), the mode of transmission could not be ascertained. The clinical features noted were protein energy malnutrition in 127 (44.56%), pulmonary and extrapulmonary tuberculosis in 84 (29.47%), hepatosplenomegaly in 82 (28.77%), persistent generalized lymphadenopathy in 67 (23.50%), skin lesions in 63 (22.10%), chronic diarrhea in 43 (15.08%), oral thrush in 42 (14.73%), pyrexia of unknown origin in 36 (12.63%), chronic lung disease in 32 (11.22%), chronic hypertrophic parotitis in 27 (9.47%), chronic ottorrhea in 26 (9.12%), recurrent lower respiratory tract infection in 24 (8.42%), neurological manifestations of non-tuberculous origin in 13 (4.56%) and Pneumocystis carinii pneumonia in 11(3.88%). Forty-eight (16.84%) were asymptomatic, 30 (10.52%) died of AIDS during the study period and 39 (13.68%) have been lost to follow up. CONCLUSION: Vertical transmission was the commonest mode of infection. Perinatally infected children become symptomatic by five years of age. Protein energy malnutrition, hepatosplenomegaly and persistent generalized lymphadenopathy were common presenting features. Tuberculosis was the major co-infection. Chronic hypertrophic parotitis and chronic lung disease were distinguishing features of this study. Encephalopathy was associated with poor outcome. PMID- 11255301 TI - Enteral nutrition for critically ill patients. PMID- 11255300 TI - Biochemical assessment of iodine deficiency disorders in Baroda and Dang districts of Gujarat State. AB - OBJECTIVE: (i) To assess the severity of Iodine Deficiency Disorders (DD) in Baroda and Dang Districts of Gujarat, using biochemical prevalence indicators of IDD; and (ii) To establish a biochemical baseline, in a sub-sample of the large population of Gujarat, that could be used to monitor the effectiveness of iodine replacement program. METHODS: 1,363 children (<1-15 years) were studied and data was collected on dietary habits, anthropometric and biochemical parameters such as height, weight and urinary iodine (UI) and blood TSH respectively. BSA and BMI were calculated. Drinking water and salt were analyzed for iodine content. RESULTS: Median true urinary iodine was 65 microg/I (interquartile-range 38-108). Mean TSH was 2.08 mU/1 (SD +/- 2.06) and 6% of the studied population had whole blood TSH values > 5 mU/1. Females from both districts were affected more by iodine deficiency as evidenced by lower true urinary iodine and higher mean TSH levels. The interfering substances were significantly higher in Baroda boys and Dang girls as compared to their counterparts (< 0.001). Boys were more malnourished than girls as evidenced by lower BMI. Dang district was more severely affected by IDD as compared to Baroda. Drinking water in Dang district was lacking in iodine content. Iodine in salt varied at around 7 to 2000 PPM. CONCLUSIONS: IDD is a public health problem in Gujarat. Baroda district is a new pocket of IDD. Dang district is the worse affected. The expression of IDD in these two districts of Gujarat revealed interplay of multiple factors. PMID- 11255302 TI - The National Family Health Survey (1998-99): childhood mortality. PMID- 11255303 TI - Seropositivity rate for HIV infection in hospitalized children on selective screening. PMID- 11255304 TI - Incidence of low birth weight in rural Ballabgarh, Haryana. PMID- 11255305 TI - Calf circumference as an alternative to birth weight for identification of low birth weight babies. PMID- 11255306 TI - Correlation of plasma color index with serum bilirubin in neonatal jaundice. PMID- 11255307 TI - Prevalence of rubella antibody in school going girls. PMID- 11255308 TI - Burkitts lymphoma developing in a child with hyper immunoglobulin E syndrome. PMID- 11255309 TI - India's first successful pediatric liver transplant. PMID- 11255310 TI - Autosomal recessive polycystic kidney disease with congenital hepatic fibrosis and encephalocele. PMID- 11255311 TI - Mucormycosis of the neonatal gastrointestinal tract. PMID- 11255312 TI - Pleomorphic xanthoastrocytoma. PMID- 11255313 TI - Hepatitis B vaccine and pregnancy. PMID- 11255315 TI - Protection provided by hepatitis B vaccine. PMID- 11255317 TI - Congenital lymphedema. PMID- 11255318 TI - Homozygous familial hypercholesterolemia. PMID- 11255319 TI - Physical signs in children with pneumonia. PMID- 11255320 TI - Recurrent abdominal pain--a reappraisal? PMID- 11255322 TI - Juvenile recurrent parotitis. PMID- 11255323 TI - Anti-CD20 monoclonal antibody (rituximab) for refractory erosive stomatitis secondary to CD20(+) follicular lymphoma-associated paraneoplastic pemphigus. PMID- 11255324 TI - A case of trichloroethylene hypersensitivity syndrome. PMID- 11255325 TI - The impact of psoriasis on quality of life: results of a 1998 National Psoriasis Foundation patient-membership survey. AB - BACKGROUND: Psoriasis can have a profound impact on a patient's quality of life. OBJECTIVES: To assess patients' perspectives on the impact of psoriasis on their lifestyle and emotional well-being and the social ramifications of living with the disease; to determine the range of therapies available; and to ascertain patients' satisfaction with the management of their disease. DESIGN: A 4-page, self-administered questionnaire was mailed on July 13 and 14, 1998, to the entire membership of the National Psoriasis Foundation (N = 40 350), and followed by a telephone survey of responders with severe psoriasis. MAIN OUTCOME MEASURES: Patients' perspectives on the psychosocial impact of psoriasis and the effectiveness of the management of their disease. RESULTS: Of the 40 350 questionnaires mailed out, a response rate of 43% was realized. The most frequent symptoms experienced by the mail-survey respondents were scaling (94%), itching (79%), and skin redness (71%); 39% reported that psoriasis covered 10% or more of their bodies. A total of 6194 patients with severe psoriasis were entered into the database for the telephone survey. Of these, 79% reported that psoriasis had a negative impact on their lives, 40% felt frustrated with the ineffectiveness of their current therapies, and 32% reported that treatment was not aggressive enough. CONCLUSIONS: The unprecedented response to the survey provides compelling evidence that individuals with psoriasis believe that the disease has a profound emotional and social as well as physical impact on their quality of life. Many patients with psoriasis, particularly those with severe disease, are frustrated with the management of their disease and by the perceived ineffectiveness of their therapies. Physicians may need to improve communication with their patients and should reevaluate their management of psoriasis. PMID- 11255326 TI - Comparative efficacy of treatments for pediculosis capitis infestations: update 2000. AB - OBJECTIVE: To evaluate the pediculicidal and ovicidal activity of 5 head lice products. DESIGN: In vitro pediculicidal and ovicidal product comparison. SETTING: Tropical field station in Panama. PARTICIPANTS: Head lice and eggs were harvested from healthy children infested with Pediculus capitis. INTERVENTION: Within 2 hours of capture, lice were placed in continuous, direct contact with the pediculicide products and observed at regular intervals. Fresh, viable eggs were immersed in the pediculicides for 10 minutes, rinsed, air-dried, and incubated for 2 weeks. MAIN OUTCOME MEASURES: Percentage of lice dead at regular observation intervals between 5 minutes and 3 hours of continuous exposure to the pediculicide and percentage of eggs not hatched after 2 weeks. RESULTS: All lice treated with Ovide lotion (0.5% malathion) were dead within 10 minutes and none of the eggs hatched. There was no significant change in the effectiveness of 0.5% malathion lotion or A-200 shampoo compared with the results of an earlier study (1986). There were significant declines in the pediculicidal activity of RID and the ovicidal activity of lindane. Nix (1% permethrin), which was not on the market at the time of the original study, killed lice in less than 30 minutes, and ovicidal activity ranged from 73% to 90% (diluted and undiluted, respectively). CONCLUSIONS: Ovide lotion (0.5% malathion) was the fastest-killing pediculicide and the most effective ovicide. One percent lindane shampoo was the slowest-acting pediculicide and least effective ovicide. Nix was highly effective in both undiluted and diluted forms. PMID- 11255327 TI - The frequency of common skin conditions in preschool-age children in Australia: atopic dermatitis. AB - OBJECTIVE: To determine the prevalence and severity of atopic dermatitis in a stratified cross-section of preschool-age children examined throughout Victoria, Australia. DESIGN: A cross-sectional skin survey using a selected cluster sample of the various centers throughout Victoria. SETTING: The study population included Victorian children attending child-care centers, preschools, and Maternal and Child Health Centres, with the reference population being Australian children aged 5 years and younger. PARTICIPANTS: Of 1634 potential participants, 1116 children (68.3%) were examined. INTERVENTION: A dermatologist performed a total skin examination, including head and neck, limbs, and trunk, on all children. The diaper area was examined in children younger than 12 months. MAIN OUTCOME MEASURE: All parents were administered a questionnaire to elicit demographic information, history of skin conditions, and family history of skin problems or related diseases. The examiner recorded the presence, site, and severity of atopic dermatitis for calculation of age- and sex-specific prevalence rates. RESULTS: The age- and sex-adjusted point prevalence was 30.8% (95% confidence interval [CI], 28.0%-33.5%). Most children (63.7%) were classified as having minimal or mild disease. Only 5.8% of children with atopic dermatitis did not have face or flexural involvement. Of the 237 children with atopic dermatitis and information available, 209 used 1 or more products to treat their condition. CONCLUSIONS: Atopic dermatitis is common, decreasing in prevalence after the first 3 years of life. Most children have mild disease requiring little if any treatment, and much could be prevented with simple measures. Educational programs directed at those caring for preschool-age children that provide information on simple preventive measures, where practical, and sources of advice for treatment, if necessary, could substantially reduce the morbidity of this condition in predisposed children. PMID- 11255328 TI - Association of human herpesvirus 6 infection with drug reaction with eosinophilia and systemic symptoms. AB - BACKGROUND: There is a current debate regarding the association of human herpesvirus 6 (HHV-6) infection and drug reaction with eosinophilia and systemic symptoms (DRESS). METHODS: Seven consecutive patients hospitalized with DRESS were enrolled in a prospective study to evaluate evidence of active HHV-6 infection. OBSERVATIONS: The imputable drugs were carbamazepine (5 patients), ibuprofen (1 patient), and sulfasalazine (1 patient). All patients were seropositive for anti-HHV-6 IgG antibodies. Anti-HHV-6 IgM antibodies were detected in 4 of the 7 patients with a seroconversion in 2 patients. Neither anti cytomegalovirus nor anti-Epstein-Barr virus early antigen IgM antibody was detected. Human herpesvirus 6 genome was not detected by polymerase chain reaction in the first serum sample of all patients. It was weakly detected in skin lesions in the last patient tested by polymerase chain reaction but was not found in uninvolved skin. CONCLUSIONS: The results suggest an association between HHV-6 active infection (primo-infection or reactivation) and severe DRESS. Absence of anti-cytomegalovirus or anti-Epstein-Barr virus early antigen IgM antibodies argues against a nonspecific viral reactivation. Human herpesvirus 6 infection may play a role in the development of DRESS in susceptible patients. Some drugs with reactive metabolites could favor reactivation and propagation of HHV-6. PMID- 11255330 TI - Icodextrin cutaneous hypersensitivity: report of 3 psoriasiform cases. AB - BACKGROUND: Icodextrin is proposed as a new osmotic agent for use in peritoneal dialysis. Because of its recent use, adverse reactions are not well known. Cutaneous adverse effects have been described. We report 3 cases of cutaneous hypersensitivity to icodextrin and discuss the pathogenesis of this reaction. OBSERVATIONS: The cutaneous adverse reaction was psoriasiform in our 3 cases. The eruption was generalized with acute generalized exanthematous pustulosis in 1 case, and limited to the palms and soles in 1 case. It occurred 10 to 15 days after icodextrin therapy was initiated. In patient 1, the results of a rechallenge with icodextrin were positive. Icodextrin therapy was discontinued in all patients. CONCLUSIONS: Some cases of cutaneous reactions to icodextrin have been reported in the literature, but they are rare. As in our cases, most eruptions are psoriasiform, limited to the palms and soles, or extensive. Although the etiology is unclear, a hypersensitivity reaction, with the formation of immunocomplexes, is probable. PMID- 11255329 TI - Differentiation and clonality of lesional lymphocytes in pityriasis lichenoides chronica. AB - BACKGROUND: Pityriasis lichenoides chronica (PLC) and pityriasis lichenoides et varioliformis acuta (PLEVA) are benign T-cell diseases that share several overlapping clinicopathologic features, leading many to believe that they exist as a spectrum rather than as single entities. Previous molecular studies have shown that PLEVA is a clonal lymphoproliferative disorder. To further characterize the immunohistologic features of PLC and to determine whether PLC demonstrates clonality, we studied 6 cases of PLC using a frozen section immunoperoxidase technique and polymerase chain reaction/denaturing gradient gel electrophoresis. OBSERVATIONS: All 6 cases showed a mild to moderate superficial and deep perivascular infiltrate composed predominantly of CD4(+) T cells, admixed with Langerhans cells and macrophages; most were associated with an HLA DR(+) epidermis. Three of 6 cases involved monoclonal T-cell receptor gamma (TCR gamma) gene rearrangements detected by V gamma 1-8/J gamma 1-2 and V gamma 9/J gamma 1-2 primers. CONCLUSIONS: Our findings enhance existing data showing that PLC shares many immunohistologic features with PLEVA and indicating that PLC is frequently a clonal T-cell disease. This provides further evidence that PLC and PLEVA are interrelated processes within the larger group of T-cell lymphoproliferative disorders. PMID- 11255331 TI - Antibiotic prophylaxis for full-face laser resurfacing: is it necessary? AB - OBJECTIVE: To evaluate the need for antibiotic prophylaxis when performing full face laser resurfacing. METHOD: Prospective study of 31 patients undergoing full face laser resurfacing, 17 with and 14 without antibiotic prophylaxis. OBSERVATION: Four of 14 patients without antibiotic prophylaxis had microbiologic and clinical evidence of infection. None of the 17 patients with antibiotic prophylaxis had clinical infection. Early treatment prevented adverse sequelae in the 4 patients who developed infection. CONCLUSION: Antibiotic prophylaxis against Staphylococcus aureus is useful but not essential, because meticulous wound care and close clinical monitoring of patients daily with routine bacterial swabs can detect infection early. PMID- 11255332 TI - Photodynamic therapy for large or multiple patches of Bowen disease and basal cell carcinoma. AB - BACKGROUND: Photodynamic therapy (PDT) using topical delta-aminolevulinic acid (delta-ALA) is an effective treatment for Bowen disease and certain basal cell carcinomas (BCCs), but its place in clinical practice remains to be established. Patients with large and/or multiple lesions of Bowen disease or BCC can represent a considerable therapeutic challenge. We suggest that delta-ALA PDT may be of particular benefit in such patients. OBSERVATION: In an open study, 35 (88%) of 40 large patches of Bowen disease, all with a maximum diameter greater than 20 mm, cleared following 1 to 3 treatments of delta-ALA PDT, although 4 patches recurred within 12 months. delta-Aminolevulinic acid PDT was also used to treat 40 large BCCs, with an identical 88% initial clearance (after 1-3 treatments), with 4 recurrences within 34 months (range, 12-60 months). In 10 further patients with multiple (> or =3) patches of Bowen disease, 44 (98%) of 45 patches cleared following delta-ALA PDT, although 4 lesions recurred over 12 months. In 3 patients with multiple BCCs, PDT cleared 52 (90%) of 58 lesions, with 2 recurrences during 41 months (range, 12-52 months). Treatments were well tolerated, with only 5 patients with solitary large lesions requiring local anesthesia. CONCLUSIONS: delta-Aminolevulinic acid PDT is an effective tissue sparing modality achieving good cosmesis. We propose that delta-ALA PDT be considered as a first-line therapy for large and/or multiple areas of Bowen disease and superficial BCCs. PMID- 11255333 TI - A meta-analysis of reverse transcriptase-polymerase chain reaction for tyrosinase mRNA as a marker for circulating tumor cells in cutaneous melanoma. AB - OBJECTIVE: To systematically review the use of reverse transcriptase-polymerase chain reaction (RT-PCR) for tyrosinase messenger RNA as a molecular serum marker for metastatic melanoma. DATA SOURCES: Computerized searches (1966-1999) of the PubMed and MDConsult databases and a manual search of retrieved article references. STUDY SELECTION: Cohort studies containing test subjects and negative controls were reviewed. DATA EXTRACTION: Three investigators independently screened abstracts for relevant studies and 2 investigators independently reviewed all eligible studies. DATA SYNTHESIS: Of 127 identified studies, 50 were reviewed in detail and 23 met all inclusion criteria. From these 23 studies, the PCR methods, the total number of patients, the number of control subjects, and the number of RT-PCR-positive patients per stage were analyzed. Results of RT-PCR for tyrosinase messenger RNA were positive in 18% (95% confidence interval [CI], 3%-22%) patients for stage I disease, 28% (95% CI, 23%-34%) for stage II disease, 19% (95% CI, 16%-21%) for stage I/II localized disease, 30% (95% CI, 26%-34%) for stage III disease, and 45% (95% CI, 41%-50%) for stage IV disease. Specificities were 100% in all but 1 study. Results of RT-PCR were positive in only 0.4% of healthy controls and patients with nonmelanoma cancer. CONCLUSIONS: The lack of data on the outcome of stage I, II, and III patients who were RT-PCR positive and the low prevalence of RT-PCR positivity in patients with known stage IV disease limit the applicability of this test at this time. Ongoing and future studies on a quantitative RT-PCR, amplification of multiple melanoma-associated antigens, and use of the test as a prognostic indicator might improve the utility of this molecular serologic tool. PMID- 11255334 TI - How can hand searching the dermatological literature benefit people with skin problems? PMID- 11255335 TI - Direct medical costs for surgical and medical treatment of condylomata acuminata. AB - OBJECTIVE: To determine which treatment modalities for condylomata acuminata are associated with the lowest direct medical costs. DESIGN: Cost-effectiveness analysis. SETTING: Ambulatory private practice, primary or specialty care. PATIENTS OR OTHER PARTICIPANTS: Adults with no presenting complaints other than condylomata acuminata. INTERVENTIONS: Construction of a cost-effectiveness model. From a literature review, extraction of commonly accepted guidelines regarding duration and frequency as well as reports of efficacies of typical treatment regimens; from Medicare physician fee schedules, costs of physician visits and physician-administered treatments; from published data, average wholesale prices of medications. MAIN OUTCOME MEASURE: Estimated direct medical costs per complete clearance associated with different treatment options for condylomata acuminata. RESULTS: Mean direct medical costs per complete clearance are lowest for surgical excision ($285). Other low-cost modalities are loop electrosurgical excision procedure ($316), electrodesiccation ($347), carbon dioxide laser ($416), podofilox ($424), and pulsed-dye laser ($479). Higher-cost modalities are cryotherapy ($951), trichloroacetic acid ($986), imiquimod ($1255), podophyllum resin ($1632), and interferon alfa-2b ($6665). CONCLUSION: Surgical modalities, including excision, electrodesiccation, loop electrosurgical excision procedure, and laser, as well as podofilox are low-cost options for the treatment of condylomata acuminata. PMID- 11255336 TI - Challenges to the hierarchy of evidence: does the emperor have no clothes? PMID- 11255337 TI - Vitiligo: the evolution of cultured epidermal autografts and other surgical treatment modalities. PMID- 11255339 TI - Psoriasis from the patient's point of view. PMID- 11255340 TI - The drug hypersensitivity syndrome: what is the pathogenesis? PMID- 11255341 TI - Recurrent papules on the left upper extremity of a 50-year-old man. PMID- 11255342 TI - Asymptomatic, firm, yellow-brown plaques of the lower eyelids and chest. PMID- 11255343 TI - A peculiar pattern of alopecia. PMID- 11255344 TI - Nonhealing perianal ulcer. PMID- 11255345 TI - Use of minocycline and soft tissue pigmentation: close association. PMID- 11255346 TI - Solar pruritus: a symptom, not a diagnosis. PMID- 11255348 TI - Folliculitis decalvans and tufted folliculitis are specific infective diseases that may lead to scarring, but are not a subset of central centrifugal scarring alopecia. PMID- 11255350 TI - Scarring alopecia and ethnicity. PMID- 11255352 TI - Histopathologist: to step section or not? PMID- 11255354 TI - Melanoma diagnosis by computerized analysis of clinical images. PMID- 11255355 TI - Epiluminescence microscopy: a reevaluation of its purpose. PMID- 11255358 TI - The role of UV light in the pathogenesis of digital papular calcific elastosis. PMID- 11255359 TI - Experimental contact sensitivity in patients with previous exposure to high-dose puva photochemotherapy. PMID- 11255360 TI - Topical psoralen photochemotherapy with lethal outcome. PMID- 11255361 TI - Do specific human papillomavirus types cause psoriasis? PMID- 11255362 TI - Butcher's warts: dermatological heritage or testable misinformation? PMID- 11255369 TI - A piece of my mind: the condition we don't discuss. PMID- 11255371 TI - 20 years after AIDS emerges, HIV's complexities still loom large. PMID- 11255370 TI - New guidelines for cardiopulmonary resuscitation and emergency cardiac care: changes in the management of cardiac arrest. PMID- 11255372 TI - New HIV therapy guidelines. PMID- 11255373 TI - Playing it cool in stroke research. PMID- 11255378 TI - Marital stress and coronary heart disease. PMID- 11255382 TI - Linezolid and reversible myelosuppression. PMID- 11255383 TI - Methylmercury and neurodevelopment: reanalysis of the Seychelles Child Development Study outcomes at 66 months of age. PMID- 11255384 TI - Continuation pharmacotherapy in the prevention of relapse following electroconvulsive therapy: a randomized controlled trial. AB - CONTEXT: Electroconvulsive therapy (ECT) is highly effective for treatment of major depression, but naturalistic studies show a high rate of relapse after discontinuation of ECT. OBJECTIVE: To determine the efficacy of continuation pharmacotherapy with nortriptyline hydrochloride or combination nortriptyline and lithium carbonate in preventing post-ECT relapse. DESIGN: Randomized, double blind, placebo-controlled trial conducted from 1993 to 1998, stratified by medication resistance or presence of psychotic depression in the index episode. SETTING: Two university-based hospitals and 1 private psychiatric hospital. PATIENTS: Of 290 patients with unipolar major depression recruited through clinical referral who completed an open ECT treatment phase, 159 patients met remitter criteria; 84 remitting patients were eligible and agreed to participate in the continuation study. INTERVENTIONS: Patients were randomly assigned to receive continuation treatment for 24 weeks with placebo (n = 29), nortriptyline (target steady-state level, 75-125 ng/mL) (n = 27), or combination nortriptyline and lithium (target steady-state level, 0.5-0.9 mEq/L) (n = 28). MAIN OUTCOME MEASURE: Relapse of major depressive episode, compared among the 3 continuation groups. RESULTS: Nortriptyline-lithium combination therapy had a marked advantage in time to relapse, superior to both placebo and nortriptyline alone. Over the 24 week trial, the relapse rate for placebo was 84% (95% confidence interval [CI], 70%-99%); for nortriptyline, 60% (95% CI, 41%-79%); and for nortriptyline lithium, 39% (95% CI, 19%-59%). All but 1 instance of relapse with nortriptyline lithium occurred within 5 weeks of ECT termination, while relapse continued throughout treatment with placebo or nortriptyline alone. Medication-resistant patients, female patients, and those with more severe depressive symptoms following ECT had more rapid relapse. CONCLUSIONS: Our study indicates that without active treatment, virtually all remitted patients relapse within 6 months of stopping ECT. Monotherapy with nortriptyline has limited efficacy. The combination of nortriptyline and lithium is more effective, but the relapse rate is still high, particularly during the first month of continuation therapy. PMID- 11255385 TI - Survival after AIDS diagnosis in adolescents and adults during the treatment era, United States, 1984-1997. AB - CONTEXT: Declines in the number of acquired immunodeficiency syndrome (AIDS) deaths were first observed in 1996, attributed to improvements in antiretroviral therapy and an increase in the proportion of persons receiving therapy. OBJECTIVE: To examine national trends in survival time among persons diagnosed as having AIDS in 1984-1997. DESIGN, SETTING, AND SUBJECTS: Retrospective cohort study using data from a population-based registry of AIDS cases and deaths reported in the United States. MAIN OUTCOME MEASURE: Months of survival after AIDS diagnosis through December 31, 1998, compared by year of diagnosis. RESULTS: Among 394 705 persons with an AIDS-defining opportunistic illness (OI) diagnosed in 1984-1997, median survival time improved from 11 months for 1984 diagnoses to 46 months for 1995 diagnoses. Among persons with an OI diagnosed in 1996 and 1997, 67% were alive at least 36 months after diagnosis and 77% were alive at least 24 months after diagnosis, respectively. Among 296 621 AIDS cases diagnosed during 1993-1997, 65% were based on immunologic criteria and 35% on OI criteria; 80% were among men; and 42% were among non-Hispanic blacks, 40% among non Hispanic whites, 17% among Hispanics, 1% among Asians/Pacific islanders, and less than 1% among American Indians/Alaska natives. The probability of surviving at least 24 months increased from 67% for those with immunologic diagnoses in 1993 to 90% in 1997 and from 49% for those with OI diagnoses in 1993 to 80% in 1997. Survival time increased with each year of diagnosis from 1984 to 1997 for blacks, whites, and Hispanics. The greatest annual survival gains occurred among persons receiving an AIDS diagnosis in 1995 and 1996. CONCLUSIONS: Survival time after AIDS diagnosis improved from 1984 to 1997. While AIDS incidence is declining, improved survival times present a growing public health challenge as the number of persons living with chronic human immunodeficiency virus disease/AIDS increases. PMID- 11255387 TI - Drug-induced QT prolongation in women during the menstrual cycle. AB - CONTEXT: Women have a higher incidence of torsades de pointes than men, but it is not known if the risk of drug-induced torsades de pointes varies during the menstrual cycle. OBJECTIVES: To determine if the degree of QT prolongation in response to ibutilide varies with the menstrual cycle phase and to compare QT prolongation between women and men. DESIGN AND SETTING: Cohort study of men and women who received the same intervention conducted between November 1998 and November 2000 at a general clinical research center of a university hospital. PARTICIPANTS: A volunteer sample of 58 healthy adults (38 men and 20 women) aged 21 to 40 years. INTERVENTION: A low dose of ibutilide (0.003 mg/kg), infused intravenously for 10 minutes. Subjects were monitored for 120 minutes. Women received the intervention on 3 separate occasions to correspond with menstrual cycle phases, which were verified by using hormonal assays. MAIN OUTCOME MEASURE: QT interval, recorded from electrocardiogram at timed intervals during and after ibutilide infusion and standardized for variations in heart rate (QTc). RESULTS: Maximum (mean [SD]) millisecond increase in QTc after ibutilide infusion was greater for women during menses (63 [13]) and the ovulatory phase (59 [17]) compared with women during the luteal phase (53 [14]) and compared with men (46 [16]; P =.002 vs menses and P =.007 vs ovulation). Progesterone (r = -0.40) and progesterone-to-estradiol ratio (r = -0.41), but not estradiol (r = 0.14) or testosterone (r = 0.09), were inversely correlated with ibutilide-induced QT prolongation. CONCLUSIONS: Menstrual cycle and sex differences exist in QTc responses to ibutilide, with the greatest increase in QTc corresponding to the first half of the menstrual cycle. PMID- 11255386 TI - Acute respiratory tract infections and mannose-binding lectin insufficiency during early childhood. AB - CONTEXT: Hospital-based studies have found that increased susceptibility to certain infections is associated with low serum levels of mannose-binding lectin (MBL) due to MBL variant alleles. However, the contribution of MBL insufficiency to incidence of common childhood infections at a population level is unknown. OBJECTIVE: To investigate the effect of MBL insufficiency on risk for acute respiratory tract infection (ARI) in unselected children younger than 2 years. DESIGN AND SETTING: Population-based, prospective, cohort study conducted in Sisimiut, Greenland. PARTICIPANTS: Two hundred fifty-two children younger than 2 years who were followed up weekly between August 1996 and August 1998 for morbidity surveillance. MAIN OUTCOME MEASURE: Risk of ARI, based on medical history and clinical examination, compared by MBL genotype, determined from blood samples based on presence of structural and promoter alleles. RESULTS: A 2.08 fold (95% confidence interval [CI], 1.41-3.06) increased relative risk (RR) of ARI was found in MBL-insufficient children (n = 13) compared with MBL-sufficient children (n = 239; P<.001). The risk association was largely restricted to children aged 6 to 17 months (RR, 2.92; 95% CI, 1.78-4.79) while less effect (RR, 1.47; 95% CI, 0.45-4.82) and no effect (RR, 1.00; 95% CI, 0.42-2.37) was shown among children aged 0 to 5 months and 18 to 23 months, respectively. CONCLUSION: These data suggest that genetic factors such as MBL insufficiency play an important role in host defense, particularly during the vulnerable period of childhood from age 6 through 17 months, when the adaptive immune system is immature. PMID- 11255388 TI - Histamine poisoning associated with eating tuna burgers. AB - CONTEXT: Histamine poisoning occurs when persons ingest fish in which bacteria have converted histidine to histamine, a process that usually can be controlled by storage at low temperatures. From 1994 to 1997, North Carolina averaged 2 cases annually; however, from July 1998 to February 1999, a total of 22 cases of histamine fish poisoning were reported. OBJECTIVES: To examine the increase in histamine case reports, identify risk factors for poisoning, and develop recommendations for prevention. DESIGN AND SETTING: Case series evaluated in North Carolina from July 1998 to February 1999. SUBJECTS: Reported case-patients with 2 of the following symptoms within 2 hours of eating tuna: rash, facial flushing, vomiting, diarrhea, dyspnea, a tight feeling in the throat, headache, or a metallic or peppery taste in the mouth. RESULTS: Twenty cases occurred during 5 outbreaks, and there were 2 single occurrences. Of the 22 persons affected, 19 (86%) sought emergency medical care. All case-patients ate tuna: 18 ate tuna burgers, 2 ate salad containing tuna, and 2 ate filets. Tuna samples (available from 3 outbreaks) had histamine levels above the Food and Drug Administration regulatory level of 50 ppm (levels were between 213 and 3245 ppm). In 19 cases, the tuna used to prepare burgers or salads was frozen and thawed more than once before serving. Violations of recommended temperature controls were identified in 2 of the 5 restaurants, accounting for 14 (64%) cases. CONCLUSIONS: Tuna burgers, a relatively new menu item in restaurants, were associated with an increase in histamine poisoning cases in North Carolina. Tuna ground for burgers can be susceptible to both temperature fluctuations and bacterial contamination. PMID- 11255389 TI - Perspectives on care at the close of life. Management of dyspnea in patients with far-advanced lung disease: "once I lose it, it's kind of hard to catch it... ". AB - Dyspnea is a common problem among patients with interstitial fibrosis, lung cancer, cystic fibrosis, and chronic obstructive pulmonary disease. The slow but steady progression of such diseases, often punctuated by acute exacerbations or secondary illnesses, can lead to decision-making dilemmas among patients and their caregivers, such as when to accept mechanical ventilation, when to forgoe aggressive therapies, and when to make formal end-of-life care plans. Two cases, a 74-year-old woman with dyspnea secondary to emphysema and a 65-year-old woman with recurrent lung cancer and severe exertional fatigue and dyspnea, illustrate how dyspneic patients approaching the end of life can be evaluated and treated. Four management strategies for dyspnea are discussed: reducing ventilatory impedance, reducing ventilatory demand, improving respiratory muscle function, and altering central perception. Physicians should encourage end-stage lung disease patients and their families to discuss issues such as hospitalization and mechanical ventilation, to prepare advance directives, and to participate in a plan to manage their dyspnea. PMID- 11255391 TI - Electroconvulsive therapy: time to bring it out of the shadows. PMID- 11255390 TI - Magnetic resonance angiography for the evaluation of lower extremity arterial disease: a meta-analysis. AB - CONTEXT: Magnetic resonance angiography (MRA) is a rapidly evolving technique that has been reported to be accurate for assessment of lower extremity arterial disease. OBJECTIVE: To obtain the best available estimates of the diagnostic performance of MRA in patients with lower extremity arterial disease. DATA SOURCES: Studies published from January 1985 through May 2000 in English, German, or French, identified from the MEDLINE, EMBASE, and Current Contents databases. STUDY SELECTION: Studies were included that allowed construction of 2 x 2 contingency tables for detection of stenosis greater than 50% or occlusion with MRA or arteriography in patients with claudication or critical ischemia. DATA EXTRACTION: Two observers graded the following elements of study quality: consecutively enrolled patients, prospective study design, clear cut-off levels, blinded assessment, and clear description of MRA technique. Summary receiver operating characteristic analysis was performed to examine the influence of year of publication, all methodological criteria, arterial tract, number of subdivisions within arterial tracts, and MRA technique on diagnostic performance. DATA SYNTHESIS: Of 3583 studies initially identified, 34 were included that evaluated MRA in 1090 patients (72% men; median age, 65 years). Magnetic resonance angiography was highly accurate for assessment of all lower extremity arteries. Three-dimensional gadolinium-enhanced (3-D Gd) MRA improved diagnostic performance compared with 2-D MRA (relative diagnostic odds ratio, 2.8 [95% confidence interval, 1.2-6.4]), adjusted for number of subdivisions within arterial tracts. The estimated points of equal sensitivity and specificity were 94% and 90% for 3-D Gd MRA and 2-D MRA, respectively. CONCLUSIONS: Magnetic resonance angiography is highly accurate for assessment of the entire lower extremity for arterial disease. Three-dimensional Gd-enhanced MRA improves diagnostic performance compared with 2-D MRA. PMID- 11255392 TI - Why do some individuals have more infections than others? PMID- 11255399 TI - A piece of my mind: searching for Margaret. PMID- 11255393 TI - The patient-physician relationship. Patient-physician communication during outpatient palliative treatment visits: an observational study. AB - CONTEXT: Improving health-related quality of life (HRQL) is an important goal of palliative treatment, but little is known about actual patient-physician communication regarding HRQL topics during palliative treatment. OBJECTIVES: To investigate the content of routine communication regarding 4 specific HRQL issues between oncologists and their patients and to identify patient-, physician-, and visit-specific factors significantly associated with discussion of such issues. DESIGN: Observational study conducted between June 1996 and January 1998. SETTING: Outpatient palliative chemotherapy clinic of a cancer hospital in the Netherlands. PARTICIPANTS: Ten oncologists and 240 of their patients (72% female; mean age, 55 years) who had incurable cancer and were receiving outpatient palliative chemotherapy. MAIN OUTCOME MEASURES: Patient and physician questionnaires and audiotape analysis of communication regarding daily activities, emotional functioning, pain, and fatigue during an outpatient consultation using the Roter Interaction Analysis System. RESULTS: Physicians devoted 64% of their conversation to medical/technical issues and 23% to HRQL issues. Patients' communication behavior was divided more equally between medical/technical issues (41%) and HRQL topics (48%). Of the independent variables investigated, patients' self-reported HRQL was the most powerful predictor of discussing HRQL issues. Nevertheless, in 20% to 54% of the consultations in which patients were experiencing serious HRQL problems, no time was devoted to discussion of those problems. In particular, these patients' emotional functioning and fatigue were unaddressed 54% and 48% of the time, respectively. Discussion of HRQL issues was not more frequent in consultations in which tumor response was evaluated. CONCLUSION: Despite increasing recognition of the importance of maintaining patients' HRQL as a goal of palliative treatment, the amount of patient-physician communication devoted to such issues remains limited and appears to make only a modest contribution, at least in an explicit sense, to the evaluation of treatment efficacy in daily clinical practice. PMID- 11255400 TI - Treatment of postmenopausal osteoporosis. PMID- 11255401 TI - Women, exercise, and aging: strong message for the "weaker" sex. PMID- 11255402 TI - Communication gaps hinder full recovery from depression. PMID- 11255403 TI - Are autoimmunologists in many women's future? PMID- 11255408 TI - Ethical issues in embryonic stem cell research. PMID- 11255409 TI - Ethical issues in embryonic stem cell research. PMID- 11255412 TI - Options for increasing the supply of living organ donors. PMID- 11255414 TI - Orthostatic hypotension and chronic fatigue syndrome. PMID- 11255415 TI - Orthostatic hypotension and chronic fatigue syndrome. PMID- 11255416 TI - Orthostatic hypotension and chronic fatigue syndrome. PMID- 11255418 TI - Industry-sponsored grand rounds and prescribing behavior. PMID- 11255419 TI - Hepatic gamma-cystathionase deficiency in patients with AIDS. PMID- 11255420 TI - Physical activity and coronary heart disease in women: is "no pain, no gain" passe? AB - CONTEXT: Physically active women have lower coronary heart disease (CHD) rates than inactive women. However, whether the association differs by intensity of activity or in women at high risk for CHD is unclear. OBJECTIVE: To examine the relation between physical activity, specifically investigating walking (a light to-moderate activity depending on pace), and CHD among women, including those at high risk for CHD. DESIGN, SETTING, AND PARTICIPANTS: Cohort study of 39 372 healthy female health professionals aged 45 years or older, enrolled throughout the United States between September 1992 and May 1995, with follow-up to March 1999. Recreational activities, including walking and stair climbing, were reported at study entry. MAIN OUTCOME MEASURE: Correlation of CHD with energy expended on all activities, vigorous activities, and walking. RESULTS: A total of 244 cases of CHD occurred. Adjusting for potential confounders, the relative risks (RRs) of CHD for less than 200, 200-599, 600-1499, and 1500 or more kcal/wk expended on all activities were 1.00 (referent), 0.79 (95% confidence interval [CI], 0.56-1.12), 0.55 (95% CI, 0.37-0.82), and 0.75 (95% CI, 0.50-1.12), respectively (P for linear trend =.03). Vigorous activities were associated with lower risk (RR, 0.63; 95% CI, 0.38-1.04 comparing highest and lowest categories). Walking also predicted lower risk among women without vigorous activities. Among these women, the multivariate RRs for walking 1 to 59 min/wk, 1.0 to 1.5 h/wk, and 2 or more h/wk, compared with no regular walking, were 0.86 (95% CI, 0.57 1.29), 0.49 (95% CI, 0.28-0.86), and 0.48 (95% CI, 0.29-0.78), respectively. For walking paces of less than 3.2 km/h (2.0 mph), 3.2 to 4.7 km/h (2.0-2.9 mph), and 4.8 km/h (3.0 mph) or more, compared with no regular walking, RRs were 0.56 (95% CI, 0.32-0.97), 0.71 (95% CI, 0.47-1.05), and 0.52 (95% CI, 0.30-0.90), respectively. When analyzed simultaneously, time spent walking (P for linear trend =.01) but not walking pace (P for linear trend =.55) predicted lower risk. The inverse association between physical activity and CHD risk did not differ by weight or cholesterol levels (P for interaction =.95 and.71, respectively), but there were significant interactions by smoking and hypertension status. Physical activity was inversely related to risk in current smokers but not hypertensive women (P for interaction =.01 and.001, respectively). CONCLUSIONS: These data indicate that even light-to-moderate activity is associated with lower CHD rates in women. At least 1 hour of walking per week predicted lower risk. The inverse association with physical activity was also present in women at high risk for CHD, including those who were overweight, had increased cholesterol levels, or were smokers. PMID- 11255421 TI - Enhanced surveillance for pregnancy-associated mortality--Maryland, 1993-1998. AB - CONTEXT: Deaths occurring among women who are pregnant or who have had a recent pregnancy have a devastating impact on the family and community. It is important to understand the magnitude and causes of pregnancy-associated mortality so that comprehensive strategies can be formulated to prevent such deaths. OBJECTIVE: To ascertain the number and causes of pregnancy-associated deaths using enhanced surveillance techniques. DESIGN, SETTING, AND SUBJECTS: Retrospective, cross sectional analysis of death certificate data of reproductive-age women, live birth and fetal death records, and medical examiner records in Maryland during 1993-1998. MAIN OUTCOME MEASURE: Number of pregnancy-associated deaths, defined as death from any cause during pregnancy or within 1 year of delivery or pregnancy termination, by source of data and cause of death. RESULTS: A total of 247 pregnancy-associated deaths were ascertained. Twenty-seven percent (n = 67) were identified through cause-of-death information obtained from death certificates, 70% (n = 174) through linkage of death records with birth and fetal death records, and 47% (n = 116) through review of medical examiner records. Homicide was the leading cause of pregnancy-associated death (n = 50; 20%), and cardiovascular disorders were the second-leading cause (n = 48; 19%). CONCLUSIONS: In this Maryland sample, comprehensive identification of pregnancy associated deaths was accomplished only after collecting information from multiple sources and including all deaths occurring up to 1 year after delivery or pregnancy termination. This enhanced pregnancy mortality surveillance led to the disturbing finding that a pregnant or recently pregnant woman is more likely to be a victim of homicide than to die of any other cause. By broadening pregnancy mortality to include all possible causes, previously neglected factors may assume increased importance in prenatal and postpartum care. PMID- 11255422 TI - Estrogen replacement therapy and ovarian cancer mortality in a large prospective study of US women. AB - CONTEXT: Postmenopausal estrogen use is associated with increased risk of endometrial and breast cancer, 2 hormone-related cancers. The effect of postmenopausal estrogen use on ovarian cancer is not established. OBJECTIVES: To examine the association between postmenopausal estrogen use and ovarian cancer mortality and to determine whether the association differs according to duration and recency of use. DESIGN AND SETTING: The American Cancer Society's Cancer Prevention Study II, a prospective US cohort study with mortality follow-up from 1982 to 1996. PARTICIPANTS: A total of 211 581 postmenopausal women who completed a baseline questionnaire in 1982 and had no history of cancer, hysterectomy, or ovarian surgery at enrollment. MAIN OUTCOME MEASURE: Ovarian cancer mortality, compared among never users, users at baseline, and former users as well as by total years of use of estrogen replacement therapy (ERT). RESULTS: A total of 944 ovarian cancer deaths were recorded in 14 years of follow-up. Women who were using ERT at baseline had higher death rates from ovarian cancer than never users (rate ratio [RR], 1.51; 95% confidence interval [CI], 1.16-1.96). Risk was slightly but not significantly increased among former estrogen users (RR, 1.16; 95% CI, 0.99-1.37). Duration of use was associated with increased risk in both baseline and former users. Baseline users with 10 or more years of use had an RR of 2.20 (95% CI, 1.53-3.17), while former users with 10 or more years of use had an RR of 1.59 (95% CI, 1.13-2.25). Annual age-adjusted ovarian cancer death rates per 100 000 women were 64.4 for baseline users with 10 or more years of use, 38.3 for former users with 10 or more years of use, and 26.4 for never users. Among former users with 10 or more years of use, risk decreased with time since last use reported at study entry (RR for last use <15 years ago, 2.05; 95% CI, 1.29 3.25; RR for last use >/=15 years ago, 1.31; 95% CI, 0.79-2.17). CONCLUSIONS: In this population, postmenopausal estrogen use for 10 or more years was associated with increased risk of ovarian cancer mortality that persisted up to 29 years after cessation of use. PMID- 11255423 TI - Mortality, CD4 cell count decline, and depressive symptoms among HIV-seropositive women: longitudinal analysis from the HIV Epidemiology Research Study. AB - CONTEXT: The impact of depression on morbidity and mortality among women with human immunodeficiency virus (HIV) has not been examined despite the fact that women with HIV have substantially higher rates of depression than their male counterparts. OBJECTIVE: To determine the association of depressive symptoms with HIV-related mortality and decline in CD4 lymphocyte counts among women with HIV. DESIGN: The HIV Epidemiologic Research Study, a prospective, longitudinal cohort study conducted from April 1993 through January 1995, with follow-up through March 2000. SETTING: Four academic medical centers in Baltimore, Md; Bronx, NY; Providence, RI; and Detroit, Mich. PARTICIPANTS: A total of 765 HIV-seropositive women aged 16 to 55 years. MAIN OUTCOME MEASURES: HIV-related mortality and CD4 cell count slope decline over a maximum of 7 years, compared among women with limited or no depressive symptoms, intermittent depressive symptoms, or chronic depressive symptoms, as measured using the self-report Center for Epidemiologic Studies Depression Scale. RESULTS: In multivariate analyses controlling for clinical, treatment, and other factors, women with chronic depressive symptoms were 2 times more likely to die than women with limited or no depressive symptoms (relative risk [RR], 2.0; 95% confidence interval [CI], 1.0-3.8). Among women with CD4 cell counts of less than 200 x 10(6)/L, HIV-related mortality rates were 54% for those with chronic depressive symptoms (RR, 4.3; 95% CI, 1.6-11.6) and 48% for those with intermittent depressive symptoms (RR, 3.5; 95% CI, 1.1-10.5) compared with 21% for those with limited or no depressive symptoms. Chronic depressive symptoms were also associated with significantly greater decline in CD4 cell counts after controlling for other variables in the model, especially among women with baseline CD4 cell counts of less than 500 x 10(6)/L and baseline viral load greater than 10 000 copies/microL. CONCLUSIONS: Our results indicate that depressive symptoms among women with HIV are associated with HIV disease progression, controlling for clinical, substance use, and sociodemographic characteristics. These results highlight the importance of adequate diagnosis and treatment of depression among women with HIV. Further research is needed to determine if treatment of depression can not only enhance the mental health of women with HIV but also impede disease progression and mortality. PMID- 11255424 TI - Reproductive period and risk of dementia in postmenopausal women. AB - CONTEXT: Exogenous estrogen use may lower risk of dementia in postmenopausal women. A relationship between long-term exposure to endogenous estrogens and incident dementia has been hypothesized but not studied. OBJECTIVE: To determine whether a longer reproductive period, as an indicator of longer exposure to endogenous estrogens, is associated with lower risk of dementia and Alzheimer disease (AD) in women who have natural menopause. DESIGN AND SETTING: The Rotterdam Study, a population-based prospective cohort study conducted in the Netherlands. PARTICIPANTS: A total of 3601 women aged 55 years or older who did not have dementia at baseline (1990-1993) and had information on age at menarche, age at menopause, and type of menopause. Participants were reexamined in 1993 1994 and 1997-1999 and were continuously monitored for development of dementia. MAIN OUTCOME MEASURES: Incidence of dementia, based on Diagnostic and Statistical Manual of Mental Disorders, Revised Third Edition criteria, and AD, based on National Institute of Neurological Disorders and Stroke/Alzheimer's Disease and Related Disorders Association criteria, compared by quartiles of reproductive period among women with natural menopause. RESULTS: During 21 046 person-years of follow-up (median follow-up, 6.3 years), 199 women developed dementia, including 159 who developed AD. After adjusting for age, dementia was not clearly associated with length of reproductive period. However, after adjusting for multiple covariates, women with natural menopause and more reproductive years had an increased risk of dementia (adjusted rate ratio [RR] for women with >39 reproductive years [highest quartile] compared with <34 reproductive years [lowest quartile], 1.78; 95% confidence interval [CI], 1.12-2.84). The adjusted RR per year of increase was 1.04 (95% CI, 1.01-1.08). For risk of AD, the adjusted RRs were 1.51 (95% CI, 0.91-2.50) and 1.03 (95% CI, 1.00-1.07), respectively. Risk of dementia associated with a longer reproductive period was most pronounced in APOE epsilon4 carriers (adjusted RR for >39 reproductive years compared with <34 reproductive years, 4.20 [95% CI, 1.97-8.92] for dementia and 3.42 [95% CI, 1.51-7.75] for AD), whereas in noncarriers, no clear association with dementia or AD was observed. CONCLUSION: Our findings do not support the hypothesis that a longer reproductive period reduces risk of dementia in women who have natural menopause. PMID- 11255425 TI - Ipriflavone in the treatment of postmenopausal osteoporosis: a randomized controlled trial. AB - CONTEXT: Data on the efficacy and safety of ipriflavone for prevention of postmenopausal bone loss are conflicting. OBJECTIVES: To investigate the effect of oral ipriflavone on prevention of postmenopausal bone loss and to assess the safety profile of long-term treatment with ipriflavone in postmenopausal osteoporotic women. DESIGN AND SETTING: Prospective, randomized, double-blind, placebo-controlled, 4-year study conducted in 4 centers in Belgium, Denmark, and Italy from August 1994 to July 1998. PARTICIPANTS: Four hundred seventy-four postmenopausal white women, aged 45 to 75 years, with bone mineral densities (BMDs) of less than 0.86 g/cm(2). INTERVENTIONS: Patients were randomly assigned to receive ipriflavone, 200 mg 3 times per day (n = 234), or placebo (n = 240); all received 500 mg/d of calcium. MAIN OUTCOME MEASURES: Efficacy measures included spine, hip, and forearm BMD and biochemical markers of bone resorption (urinary hydroxyproline corrected for creatinine and urinary CrossLaps [Osteometer Biotech, Herlev, Denmark] corrected for creatinine), assessed every 6 months. Laboratory safety measures and adverse events were recorded every 3 months. RESULTS: Based on intent-to-treat analysis, after 36 months of treatment, the annual percentage change from baseline in BMD of the lumbar spine for ipriflavone vs placebo (0.1% [95% confidence interval (CI), -7.9% to 8.1%] vs 0.8% [95% CI, -9.1% to 10.7%]; P =.14), or in any of the other sites measured, did not differ significantly between groups. The response in biochemical markers was also similar between groups (eg, for hydroxyproline corrected for creatinine, 20.13 mg/g [95% CI, 18.85-21.41 mg/g] vs 20.67 mg/g [95% CI, 19.41-21.92 mg/g]; P =.96); urinary CrossLaps corrected for creatinine, 268 mg/mol (95% CI, 249-288 mg/mol) vs 268 mg/mol (95% CI, 254-282 mg/mol); P =.81. The number of women with new vertebral fracture was identical or nearly so in the 2 groups at all time points. Lymphocyte concentrations decreased significantly (500/microL (0.5 x 10(9)/L]) in women treated with ipriflavone. Thirty-one women (13.2%) in the ipriflavone group developed subclinical lymphocytopenia, of whom 29 developed it during ipriflavone treatment. Of these, 15 (52%) of 29 had recovered spontaneously by 1 year and 22 (81%) of 29 by 2 years. CONCLUSIONS: Our data indicate that ipriflavone does not prevent bone loss or affect biochemical markers of bone metabolism. Additionally, ipriflavone induces lymphocytopenia in a significant number of women. PMID- 11255426 TI - Hormone replacement therapy and cognition: systematic review and meta-analysis. AB - CONTEXT: Some observational data suggest that hormone replacement therapy (HRT) may reduce the risk of cognitive decline and dementia but results have been conflicting. OBJECTIVE: To review and evaluate studies of HRT for preventing cognitive decline and dementia in healthy postmenopausal women. DATA SOURCES: Studies with English-language abstracts identified in MEDLINE (1966-August 2000), HealthSTAR (1975-August 2000, PsychINFO (1984-August 2000); Cochrane Library databases; and articles listed in reference lists of key articles. STUDY SELECTION: Randomized controlled trials and cohort studies were reviewed for the effects of HRT on cognitive decline; cohort and case-control studies were reviewed for dementia risk. No randomized controlled trials regarding dementia risk were identified. DATA EXTRACTION: Twenty-nine studies met inclusion criteria and were rated. Two reviewers rated study quality independently and 100% agreement was reached on Jadad scores and 80% agreement was reached on US Preventive Services Task Force quality scores. A final score was reached through consensus if reviewers disagreed. DATA SYNTHESIS: Studies of cognition were not combined quantitatively because of heterogeneous study design. Women symptomatic from menopause had improvements in verbal memory, vigilance, reasoning, and motor speed, but no enhancement of other cognitive functions. Generally, no benefits were observed in asymptomatic women. A meta-analysis of observational studies suggested that HRT was associated with a decreased risk of dementia (summary odds ratio, 0.66; 95% confidence interval, 0.53-0.82). However, possible biases and lack of control for potential confounders limit interpretation of these studies. Studies did not contain enough information to assess adequately the effects of progestin use, various estrogen preparations or doses, or duration of therapy. CONCLUSIONS: In women with menopausal symptoms, HRT may have specific cognitive effects, and future studies should target these effects. The meta-analysis found a decreased risk of dementia in HRT users but most studies had important methodological limitations. PMID- 11255428 TI - Implications of low diagnostic reproducibility of cervical cytologic and histologic diagnoses. PMID- 11255429 TI - Women's health-filling the gaps. PMID- 11255427 TI - Interobserver reproducibility of cervical cytologic and histologic interpretations: realistic estimates from the ASCUS-LSIL Triage Study. AB - CONTEXT: Despite a critical presumption of reliability, standards of interpathologist agreement have not been well defined for interpretation of cervical pathology specimens. OBJECTIVE: To determine the reproducibility of cytologic, colposcopic histologic, and loop electrosurgical excision procedure (LEEP) histologic cervical specimen interpretations among multiple well-trained observers. DESIGN AND SETTING: The Atypical Squamous Cells of Undetermined Significance-Low-grade Squamous Intraepithelial Lesion (ASCUS-LSIL) Triage Study (ALTS), an ongoing US multicenter clinical trial. SUBJECTS: From women enrolled in ALTS during 1996-1998, 4948 monolayer cytologic slides, 2237 colposcopic biopsies, and 535 LEEP specimens were interpreted by 7 clinical center and 4 Pathology Quality Control Group (QC) pathologists. MAIN OUTCOME MEASURES: kappa Values calculated for comparison of the original clinical center interpretation and the first QC reviewer's masked interpretation of specimens. RESULTS: For all 3 specimen types, the clinical center pathologists rendered significantly more severe interpretations than did reviewing QC pathologists. The reproducibility of monolayer cytologic interpretations was moderate (kappa = 0.46; 95% confidence interval [CI], 0.44-0.48) and equivalent to the reproducibility of punch biopsy histopathologic interpretations (kappa = 0.46; 95% CI, 0.43-0.49) and LEEP histopathologic interpretations (kappa = 0.49; 95% CI, 0.44-0.55). The lack of reproducibility of histopathology was most evident for less severe interpretations. CONCLUSIONS: Interpretive variability is substantial for all types of cervical specimens. Histopathology of cervical biopsies is not more reproducible than monolayer cytology, and even the interpretation of LEEP results is variable. Given the degree of irreproducibility that exists among well-trained pathologists, realistic performance expectations should guide use of their interpretations. PMID- 11255430 TI - Examining homicide's contribution to pregnancy-associated deaths. PMID- 11255435 TI - Cerebrospinal fluid beta-amyloid and tau proteins for the diagnosis of Alzheimer disease. PMID- 11255436 TI - Tau mutations--center tent or sideshow? PMID- 11255437 TI - New biochemical markers in Alzheimer disease. PMID- 11255438 TI - Age-related memory decline: current concepts and future directions. AB - The effect of age on memory and the brain has been the focus of many studies. Results have identified critical questions that need to be addressed to further our understanding of age-related memory decline: Is cognitive decline diffuse or selective? Where does memory decline localize to anatomically? Does decline represent an abnormal state? What are the causes of memory decline? What level of analysis is needed to investigate age-related cortical changes? These questions are reviewed herein, and attempts at early answers are discussed. PMID- 11255439 TI - Glutamate transporters in neurologic disease. PMID- 11255440 TI - Evaluation of CSF-tau and CSF-Abeta42 as diagnostic markers for Alzheimer disease in clinical practice. AB - OBJECTIVE: To evaluate the diagnostic potential of cerebrospinal fluid (CSF) levels of tau and beta-amyloid protein ending at amino acid 42 (Abeta42) as biomarkers for Alzheimer disease (AD) in clinical practice. DESIGN: A 1-year prospective study. SETTING: Community population-based sample of all consecutive patients admitted for investigation of cognitive symptoms to the Pitea River Valley Hospital, Pitea, Sweden. PATIENTS: A total of 241 patients with probable AD (n = 105), possible AD (n = 58), vascular dementia (n = 23), mild cognitive impairment (n = 20), Lewy body dementia (n = 9), other neurological disorders (n = 3), and psychiatric disorders (n = 5) and nondemented individuals (n = 18). MAIN OUTCOME MEASURES: Cerebrospinal fluid tau and CSF-Abeta42 were assayed each week as routine clinical neurochemical analyses. Sensitivity and specificity were defined using the regression line from 100 control subjects from a multicenter study. Positive and negative predictive values were calculated for different prevalence rates of AD. RESULTS: We found increased CSF-tau and decreased CSF Abeta42 levels in probable and possible AD. Sensitivity was 94% for probable AD, 88% for possible AD, and 75% for mild cognitive impairment, whereas specificity was 100% for psychiatric disorders and 89% for nondemented. Specificity was lower in Lewy body dementia (67%) mainly because of low CSF-Abeta42 levels and in vascular dementia (48%) mainly because of high CSF-tau levels. Sensitivity for CSF-tau and CSF-Abeta42 increased in patients with AD possessing the ApoE epsilon4 allele, approaching 100%. At a prevalence of AD of 45%, the positive predictive value was 90% and the negative predictive value was 95%. CONCLUSIONS: Cerebrospinal fluid tau and CSF-Abeta42 have so far been studied in research settings, under conditions providing data on the optimal performance. We examined a prospective patient sample, with assays run in clinical routine, giving figures closer to the true performance of CSF-tau and CSF-Abeta42. The predictive value for AD was greater than 90%. Therefore, these biomarkers may have a role in the clinical workup of patients with cognitive impairment, especially to differentiate early AD from normal aging and psychiatric disorders. PMID- 11255441 TI - Frequency of tau gene mutations in familial and sporadic cases of non-Alzheimer dementia. AB - BACKGROUND: Mutations in the tau gene have been reported in families with frontotemporal dementia (FTD) linked to chromosome 17. It remains uncertain how commonly such mutations are found in patients with FTD or non-Alzheimer dementia with or without a positive family history. OBJECTIVE: To determine the frequency of tau mutations in patients with non-Alzheimer dementia. PATIENTS AND METHODS: One hundred one patients with non-Alzheimer, nonvascular dementia, most thought to have FTD. Of these, 57 had a positive family history of dementia. Neuropathologic findings were available in 32. The tau gene was sequenced for all exons including flanking intronic DNA, portions of the 3' and 5' untranslated regions, and at least 146 base pairs in the intron following exon 10. RESULTS: Overall, the frequency of the tau mutations was low, being 5.9% (6/101) in the entire group. No mutations were found in the 44 sporadic cases. However, 6 (10.5%) of the 57 familial cases and 4 (33%) of the 12 familial cases with tau pathologic findings had mutations in the tau gene. The most common mutation was P301L. CONCLUSIONS: We conclude that tau mutations are uncommon in a neurology referral population with non-Alzheimer dementia, even in those with a clinical diagnosis of FTD. However, a positive family history and/or tau pathologic findings increase the likelihood of a tau mutation. There must be other genetic and nongenetic causes of FTD and non-Alzheimer dementia, similar to the etiologic heterogeneity present in Alzheimer disease. PMID- 11255442 TI - Ratio of 8-hydroxyguanine in intact DNA to free 8-hydroxyguanine is increased in Alzheimer disease ventricular cerebrospinal fluid. AB - BACKGROUND: Markers of oxidative stress are increased in cerebrospinal fluid (CSF) of patients with Alzheimer disease (AD), although none of those reported are appropriate diagnostic markers because of the overlap between patients with AD and control subjects. OBJECTIVE: To determine the ratio of 8-hydroxyguanine (8 OHG) levels in intact DNA to free 8-OHG in the ventricular CSF of patients with AD and age-matched control subjects. The most prominent marker of DNA oxidation is 8-OHG. METHODS: Free 8-hydroxy-2'-deoxyguanosine (8-OHdG) was isolated from ventricular CSF taken at autopsy from 18 subjects with AD and 7 control subjects using solid-phase extraction columns. Levels were measured as the hydrolysis product, 8-OHG, using gas chromatography/mass spectrometry with selective ion monitoring. Intact DNA was isolated from the same CSF and the levels of 8-OHG were determined in the intact structures. Stable-labeled 8-OHG was used for quantification. RESULTS: A statistically significant (P<.05) 108-fold increase in the ratio of 8-OHG in intact DNA to free 8-OHG was observed in patients with AD. Analysis of the data distribution indicated that the lowest AD ratio was 3.5 times higher than the highest control ratio; there was no overlap of the 2 populations. CONCLUSION: Although the data for each individual measurement demonstrates overlap between patients with AD and control subjects, the ratio of 8-OHG intact in DNA to free 8-OHG demonstrates a delineation between patients with AD and control 8-OHG subjects and may be useful as a marker of disease progression or the efficacy of therapeutic antioxidant intervention. PMID- 11255443 TI - Mild cognitive impairment represents early-stage Alzheimer disease. AB - BACKGROUND: Mild cognitive impairment (MCI) is considered to be a transitional stage between aging and Alzheimer disease (AD). OBJECTIVE: To determine whether MCI represents early-stage AD by examining its natural history and neuropathologic basis. DESIGN: A prospective clinical and psychometric study of community-living elderly volunteers, both nondemented and minimally cognitively impaired, followed up for up to 9.5 years. Neuropathologic examinations were performed on participants who had undergone autopsy. SETTING: An AD research center. PARTICIPANTS: All participants enrolled between July 1990 and June 1997 with Clinical Dementia Rating (CDR) scores of 0 (cognitively healthy; n = 177; mean age, 78.9 years) or 0.5 (equivalent to MCI; n = 277; mean age, 76.9 years). Based on the degree of clinical confidence that MCI represented dementia of the Alzheimer type (DAT), 3 subgroups of individuals with CDR scores of 0.5 were identified: CDR 0.5/DAT, CDR 0.5/incipient DAT, and CDR 0.5/uncertain dementia. MAIN OUTCOME MEASURE: Progression to the stage of CDR 1, which characterizes mild definite DAT. RESULTS: Survival analysis showed that 100% of CDR 0.5/DAT participants progressed to greater dementia severity over a 9.5-year period. At 5 years, rates of progression to a score of CDR 1 (or greater) for DAT were 60.5% (95% confidence interval [CI], 50.2%-70.8%) for the CDR 0.5/DAT group, 35.7% (95% CI, 21.0%-50.3%) for the CDR 0.5/incipient DAT group, 19.9% (95% CI, 8.0%-31.8%) for the CDR 0.5/uncertain dementia group, and 6.8% (95% CI, 2.2%-11.3%) for CDR 0/controls. Progression to greater dementia severity correlated with degree of cognitive impairment at baseline. Twenty-four of the 25 participants with scores of CDR 0.5 had a neuropathologic dementing disorder, which was AD in 21 (84%). CONCLUSIONS: Individuals currently characterized as having MCI progress steadily to greater stages of dementia severity at rates dependent on the level of cognitive impairment at entry and they almost always have the neuropathologic features of AD. We conclude that MCI generally represents early-stage AD. PMID- 11255444 TI - Mild cognitive impairments predict dementia in nondemented elderly patients with memory loss. AB - BACKGROUND: Some elderly individuals exhibit significant memory deficits but do not have dementia because their general intellect is preserved and they have no impairments in everyday activities. These symptoms are often a precursor to Alzheimer disease (AD), but sometimes dementia does not occur, even after many years of observation. There is currently no reliable way to distinguish between these 2 possible outcomes in an individual patient. We hypothesized that clear impairments in at least 1 cognitive domain in addition to memory would help identify those who will progress to AD. OBJECTIVE: To determine whether nondemented patients with impairments in memory and other domains are more likely than those with memory impairment alone to develop AD. DESIGN AND METHODS: In a retrospective study, we evaluated 48 nondemented, nondepressed patients with clinical and psychometric evidence of memory impairment who were followed up for 2 or more years. Age-adjusted normative criteria were used to identify whether additional impairments were present in language, attention, motor visuospatial function, and verbal fluency at this initial evaluation. The presence or absence of dementia after 2 years and at the most recent neurological evaluation was compared in subjects with normal scores in all 4 of these cognitive areas apart from memory (M-) and those with impairment in 1 or more of these areas (M+). Outcomes were adjusted for age, intelligence at initial evaluation, and years of education. RESULTS: Of the 48 nondemented patients with memory loss, 17 met M- criteria, leaving 31 in the M+ group. Deficits in block design were the most frequent abnormality other than memory loss. At the 2-year follow-up, 1 M- subject (6%) had progressed to AD, whereas 15 (48%) of the M+ group had progressed to AD (P =.003). At the most recent follow-up (mean +/- SD, 4.0 +/- 2.0 years), 4 (24%) of the M- patients progressed to AD compared with 24 (77%) of the M+ patients (P<.001). CONCLUSIONS: Among nondemented elderly patients, memory loss alone rarely progresses to dementia in the subsequent 2 years. However, the risk of dementia is significantly increased among patients with clear cognitive impairments beyond memory loss. Further study is needed to determine whether patients with impairments limited to memory loss have a distinctive clinical course or pathophysiology. PMID- 11255445 TI - Response of patients with Alzheimer disease to rivastigmine treatment is predicted by the rate of disease progression. AB - BACKGROUND: Evidence suggests that disease severity predicts the response of patients with Alzheimer disease (AD) to cholinesterase inhibitor treatment, raising the question of whether disease progression also predicts response to this treatment. OBJECTIVE: To evaluate retrospectively whether rate of disease progression during placebo treatment affects response to subsequent rivastigmine tartrate therapy for patients with mild to moderately severe AD. DESIGN: A 26 week, open-label extension study following a 26-week, double-blind, randomized, placebo-controlled trial. SETTING: Outpatient research centers at 22 sites in the United States. PATIENTS: We studied 187 of 235 patients originally randomized to receive placebo treatment in the double-blind phase of the trial who continued with open-label (rivastigmine) extension therapy. INTERVENTION: Placebo treatment for 26 weeks followed by rivastigmine treatment, 2 to 12 mg/d, for 26 weeks. MAIN OUTCOME MEASURES: Alzheimer's Disease Assessment Scale-cognitive subscale (ADAS Cog), Progressive Deterioration Scale, Mini-Mental State Examination, and Global Deterioration Scale scores. RESULTS: Rivastigmine administration during open label extension therapy benefited patients who had progressed slowly and those who had progressed rapidly during initial double-blind placebo treatment. Slowly progressive patients responded with a mean 1.03-point improvement in the week 26 (ie, start of open-label rivastigmine treatment) ADAS-Cog score at 12 weeks of rivastigmine treatment (week 38 of treatment; P =.02 vs week 26). However, more rapidly progressive patients had a significantly larger mean 4.97-point improvement from the week 26 ADAS-Cog score at 12 weeks (with respect to week 26 of treatment and slowly progressive patient scores, P<.001 for both). Thus, a more rapid disease progression rate while receiving placebo treatment was predictive of a significantly stronger patient response to rivastigmine therapy. This relation also was observed with the other 3 outcome measures and was still apparent when accounting for disease severity. CONCLUSIONS: Rate of disease progression for patients with mild to moderate AD seems to predict response to rivastigmine treatment. Patients with more rapidly progressive disease might be particularly likely to benefit from rivastigmine therapy. PMID- 11255446 TI - Open-label, multicenter, phase 3 extension study of the safety and efficacy of donepezil in patients with Alzheimer disease. AB - BACKGROUND: Donepezil hydrochloride is a selective acetylcholinesterase inhibitor approved for the symptomatic treatment of mild to moderately severe Alzheimer disease (AD). Controlled clinical trials of up to 24 weeks have demonstrated that donepezil treatment (5 and 10 mg/d) significantly improves cognition and global function. OBJECTIVE: To investigate the long-term benefits of donepezil treatment in patients with AD. DESIGN: Multicenter, open-label, 144-week extension of 2 US phase 3, double-blind, placebo-controlled clinical trials: a 15-week study (12 weeks of treatment followed by a 3-week placebo washout) and a 30-week study (24 weeks of treatment followed by a 6-week placebo washout). INTERVENTIONS: All patients (N = 763) initially received donepezil, 5 mg/d, for 6 weeks, after which an increase to 10 mg/d was encouraged. MEASURES: Primary efficacy measures were the Alzheimer's Disease Assessment Scale-cognitive subscale and the Clinical Dementia Rating-Sum of the Boxes. RESULTS: After the shorter 3-week placebo washout, donepezil-associated benefits remained above original baseline values for an additional 24 weeks of open-label treatment. Benefits on Alzheimer's Disease Assessment Scale-cognitive subscale scores for patients who received 10 mg/d in the double-blind study were evident compared with the other groups for 108 weeks of open-label treatment. In contrast, donepezil-associated benefits were lost after the 6-week placebo washout, and scores decreased below original baseline values for all patient groups. Although scores improved relative to the new open-label study baseline scores after drug use was restarted, patients remained below original baseline values. The most common adverse events were associated with the nervous and digestive systems and were generally mild and transient; 17% of patient discontinuations were associated with adverse events. CONCLUSIONS: Donepezil is an effective and safe drug for the long-term symptomatic treatment of mild to moderately severe AD for up to 144 weeks (2.8 years), and sustained treatment may confer some advantages. PMID- 11255447 TI - Postmenopausal estrogen replacement therapy and the risk of Alzheimer disease. AB - BACKGROUND: Previous studies have examined the relation between postmenopausal estrogen replacement therapy (ERT) and the risk of Alzheimer disease (AD). The findings have been inconsistent, since some studies have been interpreted as showing a protective effect while others have reported no effect. OBJECTIVE: To determine whether exposure to ERT is associated with a reduced risk of AD. DESIGN: Population-based nested case-control study. SETTING: The United Kingdom based General Practice Research Database. PATIENTS: The base cohort consisted of women who were recipients of ERT (n = 112 481) and a similar cohort of women who did not use estrogens (n = 108 925). The 2 cohorts were restricted to women born on or before January 1, 1950. From the 2 cohorts, we identified and verified 59 newly diagnosed cases of AD and 221 matched control subjects. MAIN OUTCOME MEASURE: Prior and current use of ERT in cases compared with controls. RESULTS: Among the 59 newly diagnosed cases of AD, 15 (25%) were current estrogen users, while among the controls, 53 (24%) were current users. The adjusted odds ratio comparing all current estrogen recipients with nonrecipients was 1.18 (95% confidence interval, 0.59-2.37). In estrogen users who took the drug for 5 years or longer compared with nonusers, the odds ratio was 1.05 (95% confidence interval, 0.32-3.44). Odds ratios were similar for estrogen recipients who received estrogens alone and recipients who received combined estrogen-progestin treatment. CONCLUSION: The use of ERT in women after the onset of menopause was not associated with a reduced risk of developing AD. PMID- 11255448 TI - Amyloid precursor protein in platelets of patients with Alzheimer disease: effect of acetylcholinesterase inhibitor treatment. AB - BACKGROUND: Amyloid precursor protein (APP) forms with apparent molecular weights of 130, 110, and 106 kd are present in human platelets. It has been demonstrated that Alzheimer disease (AD) is specifically associated with a decreased APP forms ratio in platelets. OBJECTIVE: To investigate whether acetylcholinesterase (AChE) inhibitor treatment modifies the ratio of platelet APP forms in patients with AD. PATIENTS AND METHODS: From a large sample of patients with probable AD, 30 with mild to moderate AD were selected. Each patient underwent a clinical evaluation including the Mini-Mental State Examination (MMSE) and platelet APP forms analysis at baseline and after 30 days. During this interval, 20 of 30 patients with AD were treated with donepezil hydrochloride (5 mg/d), a piperidine phosphate-based cholinesterase inhibitor. Platelets were subjected to Western blot analysis using monoclonal antibody (22C11). The ratio between the immunoreactivity of the higher-molecular-weight APP form (130 kd) and the lower forms (106 and 110 kd) was measured. RESULTS: All patients taking donepezil completed the 30 days of treatment without adverse effects. The platelet APP forms ratio at baseline did not differ between the 2 AD groups (mean +/- SD optical density ratio: untreated AD, 0.47 +/- 0.12; treated AD, 0.38 +/- 0.18), whereas a significant difference was found at follow-up (mean +/- SD optical density ratio: untreated AD, 0.45 +/- 0.17; treated AD, 0.77 +/- 0.29; P<.001). A significant improvement in MMSE scores in treated AD patients was observed from baseline (16.9 +/- 3.8) to 30 days (18.9 +/- 4.42) (P<.009, 30 days vs baseline), but no significant correlation was found in treated AD patients between MMSE score improvement and APP forms/ratio increase (P =.09). CONCLUSIONS: Administration of AChE inhibitors increases the ratio of APP forms in platelets of patients with AD, suggesting a potential effect of AChE inhibitors on APP trafficking or processing in a peripheral cell. PMID- 11255449 TI - A method for estimating progression rates in Alzheimer disease. AB - BACKGROUND: The ability to predict progression of disease in patients with Alzheimer disease (AD) would aid clinicians, improve the validation of biomarkers, and contribute to alternative study designs for AD therapies. OBJECTIVE: To test a calculated rate of initial decline prior to the first physician visit (preprogression rate) for its ability to predict progression during subsequent follow-up. METHODS: We calculated preprogression rates for 298 patients with probable or possible AD (using the criteria of the National Institute of Neurological and Communicative Disorders and Stroke and the Alzheimer's Disease and Related Disorders Associations (NINCDS-ADRDA) with a formula using expected Mini-Mental State Examination (MMSE) scores, scores at presentation, and a standardized estimate of duration. The patients are being followed up longitudinally in our Alzheimer Disease Research Center. The time to clinically meaningful deterioration, defined as an MMSE score drop of 5 or more points, was compared for patients stratified as slow, intermediate, and rapid progressors based on the preprogression rate. Cox regression analysis was used to examine the contribution of demographic variables (age, sex, ethnicity, and level of education), initial MMSE scores, estimated symptom duration, and the calculated preprogression rate to the time it took to reach the end point across the groups. RESULTS: Both initial MMSE (hazard ratio, 0.95 (0.002); z = 4.19; P<.001) and the calculated preprogression rate (hazard ratio, 1.06 (0.019); z = 3.16; P =.002) were significant in determining time to clinically meaningful decline during longitudinal follow-up (Cox regression analysis). Slow, intermediate, and rapid progressors (based on preprogression rates) experienced significantly different time intervals to clinically meaningful deterioration, with the slow progressors taking the longest time, the intermediate progressors in the middle, and the rapid progressors reaching the end point first (log rank chi(2)(1) = 9.81, P =.002). CONCLUSION: An easily calculable rate of early disease progression can classify patients as rapid, intermediate, or slow progressors with good predictive value, even at initial presentation. PMID- 11255450 TI - An investigation of clinical correlates of Lewy bodies in autopsy-proven Alzheimer disease. AB - BACKGROUND: Studies of patients meeting clinical and pathologic criteria for Alzheimer disease (AD) have not consistently found associations between the presence of Lewy bodies (LBs) at postmortem examination and a higher frequency during life of the clinical features of dementia with LBs. OBJECTIVE: To evaluate the clinical correlates of LBs in patients with AD. DESIGN AND METHODS: Fifty-one patients were diagnosed as having probable AD during life and met pathologic criteria for AD. Semiquantitative ratings for LBs were obtained in 4 brain regions: substantia nigra, cingulate, insular cortex, and hippocampus. The patients had been followed up semiannually for up to 9.9 years before death, and clinical features associated with dementia with LBs, including extrapyramidal signs and visual hallucinations, were assessed at each study visit. Logistic regression analyses determined whether patients who had LBs were more likely than those without LBs to express specific clinical signs during follow-up. Cox analyses determined whether patients with LBs developed clinical signs or died earlier. Generalized estimating equations were used to compare rates of cognitive or functional change. RESULTS: Nineteen of the 51 patients had at least 1 LB in one of the studied regions. In no case was a significant relation noted between LBs and the presence of a measured clinical sign. No LB measure was associated with an increased risk of developing any of the evaluated clinical signs earlier in the disease. There was no association between the presence of LBs and more rapid mortality or more rapid disease progression. CONCLUSIONS: In patients diagnosed as having AD during life, we did not observe a relation of LBs noted during postmortem examination with the presence of any clinical feature that we assessed or with the rapidity of disease progression. The relation between LBs and specific clinical manifestations may be tenuous in these patients. PMID- 11255451 TI - Alterations of striatal dopamine receptor binding in Alzheimer disease are associated with Lewy body pathology and antemortem psychosis. AB - BACKGROUND: Lewy bodies (LB) are present in at least 20% to 30% of persons with Alzheimer disease (AD) and contribute to the risk of psychosis and to excess cognitive burden. OBJECTIVE: To determine whether altered striatal dopamine receptor binding is associated with LB and psychosis in AD. DESIGN: Postmortem case control. SETTING: Alzheimer's Disease Research Center at the University of Pittsburgh (Pa). PARTICIPANTS: Consecutive cases from the Alzheimer's Disease Research Center brain bank, neuroleptic free for at least 1 month prior to death, with neuropathologic diagnoses of AD with LB (AD + LB, n = 14), AD without LB (AD, n = 13), or normal brains (n = 8). MAIN OUTCOME MEASURES: Dopamine D1, D2, and D3 receptor densities, and affinities as determined by selective saturation binding studies in striatal tissue. RESULTS: Subjects with AD + LB, compared with those with AD, demonstrated increased D1 receptor density and decreased D2 and D3 receptor density. D3 receptor density was selectively increased, however, in AD subjects with a history of psychosis, independent of the presence or absence of LB. The effect of neuroleptic treatment on D3 binding was further examined in an additional group of subjects who had received neuroleptics near the time of death. Neuroleptic treatment reduced D3 affinity with no effect on D3 density. CONCLUSIONS: Alzheimer disease with LB is associated with selective alterations in dopamine receptor density, which may contribute to the distinct clinical profile of this group. The D3 receptor may be an important target of neuroleptic treatment of psychosis in AD. PMID- 11255452 TI - Factors associated with incident human immunodeficiency virus-dementia. AB - BACKGROUND: Antecedents to human immunodeficiency virus-dementia (HIV-D) are poorly understood. OBJECTIVE: To identify risk factors for HIV-D. METHODS: Subjects who are positive for HIV who have CD4+ counts either below 200/microL or below 300/microL with evidence of cognitive impairment were enrolled in this study. Neurologic, cognitive, functional, and laboratory assessments were done semiannually for up to 30 months. Human immunodeficiency virus-dementia was diagnosed using American Academy of Neurology criteria for probable HIV-1 associated dementia complex. RESULTS: One hundred forty-six nondemented patients were enrolled, 45 of whom subsequently met criteria for incident HIV-D. In univariate analyses using the Cox proportional hazards regression model, the following variables were significantly associated with time to develop dementia: cognitive: abnormal scores on Timed Gait, Verbal Fluency, Grooved Pegboard, and Digit Symbol tests; attention-memory, psychomotor, and executive function domain scores; and the diagnosis of minor cognitive/motor disorder; neurologic and medical: increased abnormalities on the neurologic examination, extrapyramidal signs, history of HIV-related medical symptoms; functional: higher reported role or physical function difficulties. Depression was also a strong risk factor, along with sex, hematocrit, hemoglobin, and beta2-microglobulin levels. In a multivariate model that used cognitive domain scores, covariates with significant hazard ratios included depression, executive dysfunction, and the presence of minor cognitive/motor disorder. CONCLUSION: Cognitive deficits, minor cognitive/motor disorder, and depression may be early manifestations of HIV-D. PMID- 11255453 TI - The effect of brain atrophy on cerebral hypometabolism in the visual variant of Alzheimer disease. AB - BACKGROUND: Brain glucose metabolic rates measured by positron emission tomography can be more affected by partial volume effects in Alzheimer disease (AD) than in healthy aging because of disease-associated brain atrophy. OBJECTIVE: To determine whether the distinct distribution of cerebral metabolic lesions in patients with the visual variant of AD (AD + VS) represents a true index of neuronal/synaptic dysfunction or is the consequence of brain atrophy. SETTING: Government research hospital. DESIGN: Resting cerebral metabolic rate for glucose was measured with positron emission tomography in a cross-sectional study of AD and AD + VS groups and in healthy control subjects. Segmented magnetic resonance images were used to correct for brain atrophy. PATIENTS: Patients with AD + VS had prominent visual and visuospatial symptoms. There were 15 patients with AD, 10 with AD + VS, and 37 age-matched control subjects. MAIN OUTCOME MEASURE: Measurement of the rate of cerebral glucose metabolism. RESULTS: Before atrophy correction, the AD + VS group, compared with the control subjects, showed hypometabolism in primary and extrastriate visual areas and in parietal and superior temporal cortical areas. Compared with the AD group, the AD + VS group showed hypometabolism in visual association areas. After atrophy correction, hypometabolism remained significantly different between patients and controls and between the 2 AD groups. CONCLUSIONS: The reductions in cerebral hypometabolism represent a true loss of functional activity and are not simply an artifact caused by brain atrophy. The different patterns of hypometabolism indicate the differential development of the lesions between the AD and AD + VS groups. PMID- 11255454 TI - Neuronal cyclooxygenase 2 expression in the hippocampal formation as a function of the clinical progression of Alzheimer disease. AB - BACKGROUND: Prior studies have shown that cyclooxygenase 2 (COX-2), an enzyme involved in inflammatory mechanisms and neuronal activities, is up-regulated in the brain with Alzheimer disease (AD) and may represent a therapeutic target for anti-inflammatory treatments. OBJECTIVE: To explore COX-2 expression in the brain as a function of clinical progression of early AD. DESIGN AND MAIN OUTCOME MEASURES: Using semiquantitative immunocytochemistry, we analyzed COX-2 protein content in the hippocampal formation in 54 postmortem brain specimens from patients with normal or impaired cognitive status. SETTING AND PATIENTS: Postmortem study of nursing home residents. RESULTS: The immunointensity of COX-2 signal in the CA3 and CA2 but not CA1 subdivisions of the pyramidal layers of the hippocampal formation of the AD brain increased as the disease progressed from questionable to mild clinical dementia as assessed by Clinical Dementia Rating. COX-2 signal was increased in all 3 regions examined among cases characterized by severe dementia. CONCLUSION: Neuronal COX-2 content in subsets of hippocampal pyramidal neurons may be an indicator of progression of dementia in early AD. PMID- 11255455 TI - Frontal lobe hypometabolism predicts cognitive decline in patients with lacunar infarcts. AB - BACKGROUND: A proportion of patients with subcortical lacunes will suffer progressive cognitive dysfunction, but the basis for this decline is controversial and little is known about predicting cognitive decline in these patients. Studies of Alzheimer disease have shown that imaging measures of temporal and parietal metabolism and blood flow predict disease course. OBJECTIVE: To determine whether regional cerebral glucose metabolism predicts cognitive decline by testing 2 opposing hypotheses: (1) temporoparietal activity predicts decline (based on the idea that concomitant Alzheimer disease causes decline) vs (2) frontal hypometabolism predicts decline (based on evidence that subcortical frontal circuits are especially vulnerable to small vessel ischemia). DESIGN: Prospective cohort study. SETTING: University outpatient dementia center. PATIENTS: A convenience sample of 26 patients with radiologically defined lacunes and baseline cognitive function ranging from normal to moderately demented. MAIN OUTCOME MEASURES: Regional cerebral metabolism was quantitated in the form of atrophy-corrected positron emission tomographic activity ratios in cortical regions that were defined a priori. Patients were followed up at a mean of 1.8 years, and the dependent variable was rate of change in the Mini-Mental State Examination score. RESULTS: Bilateral and right hemisphere dorsolateral frontal metabolism significantly predicted cognitive decline, with right dorsolateral frontal metabolism explaining 19% of the variance. No other positron emission tomographic region was a significant predictor, nor were demographic variables or baseline Mini-Mental State Examination scores significant predictors. CONCLUSION: Cognitive decline in patients with lacunes may result in part from progressive vascular compromise in subcortical frontal circuits. PMID- 11255456 TI - Physical activity and risk of cognitive impairment and dementia in elderly persons. AB - CONTEXT: Dementia is common, costly, and highly age related. Little attention has been paid to the identification of modifiable lifestyle habits for its prevention. OBJECTIVE: To explore the association between physical activity and the risk of cognitive impairment and dementia. DESIGN, SETTING, AND SUBJECTS: Data come from a community sample of 9008 randomly selected men and women 65 years or older, who were evaluated in the 1991-1992 Canadian Study of Health and Aging, a prospective cohort study of dementia. Of the 6434 eligible subjects who were cognitively normal at baseline, 4615 completed a 5-year follow-up. Screening and clinical evaluations were done at both waves of the study. In 1996-1997, 3894 remained without cognitive impairment, 436 were diagnosed as having cognitive impairment-no dementia, and 285 were diagnosed as having dementia. MAIN OUTCOME MEASURE: Incident cognitive impairment and dementia by levels of physical activity at baseline. RESULTS: Compared with no exercise, physical activity was associated with lower risks of cognitive impairment, Alzheimer disease, and dementia of any type. Significant trends for increased protection with greater physical activity were observed. High levels of physical activity were associated with reduced risks of cognitive impairment (age-, sex-, and education-adjusted odds ratio, 0.58; 95% confidence interval, 0.41-0.83), Alzheimer disease (odds ratio, 0.50; 95% confidence interval, 0.28-0.90), and dementia of any type (odds ratio, 0.63; 95% confidence interval, 0.40-0.98). CONCLUSION: Regular physical activity could represent an important and potent protective factor for cognitive decline and dementia in elderly persons. PMID- 11255457 TI - Dementia with Lewy bodies studied with positron emission tomography. AB - OBJECTIVE: To report a case initially fulfilling the clinical criteria for probable Alzheimer disease, although later clinical features suggested dementia with Lewy bodies. Oxygen 15-labeled positron emission tomograms revealed a pattern of hypometabolism characteristic of Alzheimer disease. At post mortem, there was no evidence of the pathological features of Alzheimer disease, but diffuse cortical Lewy bodies were seen in the pigmented brainstem nuclei and cerebral cortex. DESIGN: A case report. SETTING: Tertiary referral center. PATIENT: A 65-year-old white man presented with a 3-year history of memory loss and language difficulties. RESULTS: Oxygen 15-labeled positron emission tomograms revealed hypometabolism in the frontal, temporal, and parietal lobes, more severe on the left than right. Metabolism in the left caudate was just outside the 95% reference range. Occipital metabolism was normal. CONCLUSIONS: Positron emission tomographic studies have been reported to show occipital hypometabolism in dementia with Lewy bodies, in addition to the characteristic posterior bitemporal biparietal pattern of Alzheimer disease. We suggest that although this finding may favor a diagnosis of dementia with Lewy bodies, it is not necessary for diagnosis. PMID- 11255458 TI - Vasovagal syncope: the contributions of Sir William R. Gowers and Sir Thomas Lewis. PMID- 11255459 TI - How amyotrophic lateral sclerosis got its name: the clinical-pathologic genius of Jean-Martin Charcot. PMID- 11255460 TI - Lower incidence of Alzheimer disease in an Indian community compared with an American community. PMID- 11255462 TI - Effect of anti-inflammatory medications on neuropathological findings in Alzheimer disease. PMID- 11255464 TI - Cortical inflammation in dementia with Lewy bodies. PMID- 11255466 TI - Small concomitant cerebrovascular lesions are not important for cognitive decline in severe Alzheimer disease. PMID- 11255468 TI - Sudden sensorineural hearing loss: does application of glucocorticoids make sense? AB - BACKGROUND: Treatment of sudden sensorineural hearing loss (SSNHL) consists of administration of blood flow-promoting drugs with or without the addition of glucocorticoids. General guidelines based on scientific data do not currently exist. OBJECTIVE: To investigate the effect of glucocorticoids on the treatment of SSNHL. SETTING: Academic medical center. PATIENTS AND METHODS: We retrospectively analyzed the audiograms of 603 patients with SSNHL: 301 patients (cared for between January 1, 1986, and December 31, 1991) received intravenous blood flow-promoting drugs without glucocorticoids and 302 patients (cared for between January 1, 1992, and December 31, 1998) received intravenous blood flow promoting drugs with glucocorticoids (intravenous +/- oral application). The age distribution of patients with SSNHL in lower, middle, and higher frequencies was similar in both groups. RESULTS: Patients with SSNHL in lower and middle frequencies (250-2000 Hz) who received glucocorticoids (prednisolone-21-hydrogen succinate) showed significantly better recovery of hearing levels compared with those who did not receive glucocorticoids (P<.05). There was no significant difference at higher frequencies between the 2 groups. Patients with SSNHL throughout all frequencies (pancochlear hearing loss) who received glucocorticoids also had significantly better recovery of hearing levels compared with those who received blood flow-promoting drugs alone (P<.05). Also, patients with elevated blood sedimentation rates had better improvement of their hearing levels after receiving glucocorticoids. CONCLUSIONS: Administration of glucocorticoids should be recommended for treatment of patients with SSNHL. In particular, patients with SSNHL in the lower and middle frequency range and pancochlear hearing loss have significantly better recovery of hearing levels. PMID- 11255469 TI - Laser myringotomy in different age groups. AB - OBJECTIVE: To study the qualities of laser myringotomy (LM) as a treatment for middle ear ventilation problems. DESIGN: Prospective study and follow-up of consecutive cases of adults, children, and infants. Patients were observed for up to 2 years. SETTING: Children underwent LM, with or without adenoidectomy, under general anesthesia in the operating room. Adults and infants underwent LM under topical anesthesia, as an outpatient procedure. PATIENTS: All consecutive patients with either secretory otitis media (SOM) (adults and children) or acute otitis media (AOM) (infants) who agreed to participate were included without selection. INTERVENTION: Myringotomy was performed using new laser equipment, enabling a 0.1-second ablation with changeable diameter. OUTCOME MEASURES: Close follow-up, with microscopic examination of all ears. Findings were noted on the medical charts. RESULTS: Among all age groups, 136 ears were followed up. Perforation lasted a mean 22 days in adults, 17 days in children, and 11 days in infants. Patient age was found to be a significant determining factor for duration of perforation (P =.002). Laser myringotomy in the anterior and inferior areas lasted longer than posterior LM (P<.001). In patients with SOM, during the time the LM was patent, all ears were ventilated. In children, 38% of SOM cases resolved after a single LM treatment. All infants with AOM recovered promptly without antibiotic treatment. CONCLUSIONS: Laser myringotomy is a convenient, quick procedure that can be performed in the medical office with the use of topical anesthesia and is suitable for patients with AOM or for those who need short-term ventilation for SOM. It was found to be a safe alternative to ventilation tubes in these patients. In AOM, it was used instead of antibiotics and gave prompt relief from symptoms and cure of the AOM. PMID- 11255470 TI - Craniofacial, temporal bone, and audiologic abnormalities in the spectrum of hemifacial microsomia. AB - OBJECTIVES: To evaluate the clinical, audiologic, and temporal bone computed tomograpic findings in patients with hemifacial microsomia and to use the OMENS (each letter of the acronym indicates 1 of the following 5 dysmorphic manifestations: O, orbital asymmetry; M, mandibular hypoplasia; E, auricular deformity; N, nerve involvement; and S, soft tissue deficiency) grading system to assess possible correlations between the severity of dysmorphic features with the type of abnormalities in the temporal bone and with degree of hearing deficit. DESIGN: Retrospective study. SETTING: Tertiary care children's hospital. PATIENT: Forty patients with hemifacial microsomia. RESULT: Mandibular hypoplasia and auricular abnormalities were the most common clinical manifestations, present in 39 patients (97%) and 38 patients (95%), respectively. Conductive hearing loss was noted in 35 patients (86%) and sensorineural hearing loss in 4 patients (10%). Facial nerve weakness was present in 20 patients (50%). Twenty patients had unilateral aural atresia, 12 patients had unilateral aural stenosis, and 7 patients had bilateral anomalies. Moderate hypoplasia or atresia of the middle ear was noted in 36 patients (90%) and ossicles were malformed in 30 patients (75%). Hypoplasia of the oval window was the most common inner ear abnormality. CONCLUSIONS: Severity of craniofacial features (total OMENS score) significantly correlated with the degree of temporal bone abnormality, but no correlation was noted with the degree or type of hearing loss. We recommend the following: (1) use of the OMENS classification system for documentation and analysis of dysmorphic finding in hemifacial microsomia; (2) complete audiologic evaluation in all patients with hemifacial microsomia regardless of the type of craniofacial abnormalities; and (3) temporal bone computed tomography for further evaluation of hearing deficit. PMID- 11255471 TI - Narrow internal auditory meatus: an idiopathic case confirming the origin and pathway of vestibular evoked myogenic potentials in humans. AB - OBJECTIVE: To confirm the origin and pathway of vestibular evoked myogenic potentials (VEMPs) in humans. DESIGN: Case study. SETTING: University hospital. PATIENT: A patient with a narrow internal auditory meatus (IAM). MAIN OUTCOME MEASURES: Imaging studies and functional studies concerning the seventh and eighth cranial nerves. RESULTS: Of the 4 nerves in the IAM, all but the cochlear nerve had normal function and normal courses, despite the pronounced narrowing of the IAM. The facial nerve had a normal diameter, but the vestibular nerves were thinner. Imaging revealed that the cochlear nerve was absent or extremely thinned. Both the cochlea and the cochlear nerve showed no function in the affected ear, although the VEMPs were evoked normally. CONCLUSION: Our results definitively support the vestibular origin of VEMPs in humans. PMID- 11255472 TI - Management of orbital subperiosteal abscess in children. AB - OBJECTIVES: To present guidelines for the management of an orbital subperiosteal abscess (SPA) in children and to assess the efficacy and safety of transnasal endoscopic drainage of an orbital SPA. SETTING: Tertiary care children's hospital. PATIENTS: Nineteen patients treated for an SPA between July 1997 and December 1999. The age of the patients ranged from 17 months to 14 years (mean, 6 years). The male-female ratio was 10:9. Treatment modalities included transnasal endoscopic drainage (n = 11), external drainage (n = 3), and intravenous antibiotics alone (n = 5). RESULTS: Bilateral pansinusitis was the most common cause. All patients received an initial trial of intravenous antibiotics. Based on the Fisher exact test, no statistically significant differences were detected for age, sex, presence of gaze restriction, and radiographic findings. Based on multiple logistic regression, degree of proptosis was the only significant multivariate predictor of surgery (P =.003). The estimated probability of surgery was 6% when there was no proptosis, and 92% for 2 mm of proptosis. The location of the SPA determined the route of surgical drainage. Eleven patients with a medially based SPA underwent drainage via the transnasal endoscopic approach, and 3 with a superior SPA underwent drainage externally. The external approach was associated with a longer hospital stay (median, 7 days) than either the endoscopic or the intravenous antibiotic approach (median, 5 days). PMID- 11255473 TI - Cricotracheal resection in children. AB - OBJECTIVE: To review our experience with cricotracheal resection in a pediatric population. DESIGN: Prospective case review of a cohort of patients undergoing cricotracheal resection. SETTING: Tertiary care pediatric hospital. PATIENTS: Forty-four consecutive patients undergoing cricotracheal resection between January 1, 1993, and December 31, 1998. MAIN OUTCOME MEASURES: Decannulation rates. RESULTS: Thirty-eight (86%) of the 44 children are decannulated. The ultimate decannulation rate was independent of the presenting grade of subglottic stenosis. Fourteen children (100%) had a primary cricotracheal resection; all are decannulated. Twenty-one children had a salvage cricotracheal resection, and 19 (90%) are decannulated. Nine children had an extended cricotracheal resection, of whom 5 (56%) are decannulated. A primary cricotracheal resection was performed on a child on whom no previous open airway procedure had been performed. A salvage cricotracheal resection was performed on a child on whom previous open airway reconstruction had not resulted in an adequate airway. An extended cricotracheal resection was performed on a child on whom the cricotracheal resection was combined with a second procedure, either additional expansion cartilage grafting or an open arytenoid procedure. Most of these children had complex airway pathologic conditions. CONCLUSION: Cricotracheal resection complements standard laryngotracheal reconstruction techniques in a pediatric population. PMID- 11255474 TI - Brain-derived neurotrophic factor-enriched collagen tubule as a substitute for autologous nerve grafts. AB - BACKGROUND: Autologous nerve interposition grafts are frequently harvested by head and neck surgeons. The sacrifice of these donor nerves guarantees some degree of morbidity, including sensory loss, additional incision sites with associated potential complications, and prolonged operative time. An alternative to autologous nerve grafting is, therefore, desirable. OBJECTIVE: To determine if a collagen tubule (CT) filled with either a plain collagen gel or a brain-derived neurotrophic factor (BDNF)-enriched collagen gel could be used to achieve functional and histologic outcomes equivalent to an autologous nerve graft in bridging a 15-mm nerve gap in the rabbit facial nerve. DESIGN: A prospective, randomized, blinded animal study with a control group. METHODS: Thirty rabbit facial nerves were resected (15-mm segments) to create nerve gaps. The gaps were bridged using 1 of 3 methods, assigned randomly: a reversed facial nerve (control), a collagen gel-filled CT, or a BDNF-enriched collagen gel-filled CT. The animals were evaluated after 6 weeks in a blinded fashion for functional nerve recovery, axon count, and axonal diameter. RESULTS: There were no significant differences between the autologous nerve graft group, the collagen gel-filled CT group, or the BDNF-enriched collagen gel-filled CT group (n = 10 for each group) for functional nerve recovery (P =.94). The mean axon count and the mean axonal diameter were highest in the BDNF-enriched collagen gel-filled CT group, but these differences failed to reach statistical significance (P =.18 and.96, respectively). CONCLUSIONS: Collagen tubules filled with BDNF-enriched collagen gel appear to be at least as good as autologous nerve grafts for bridging short facial nerve gaps. Larger experimental studies are warranted to determine if clinical trials are justified. PMID- 11255475 TI - Prospective evaluation of eyelid function with gold weight implant and lower eyelid shortening for facial paralysis. AB - OBJECTIVES: To assess which signs and symptoms were relieved by gold weight implantation and which signs and symptoms persisted. DESIGN: Prospective observational cohort. SETTING: Tertiary care neurotology and oncology center. PATIENTS: Sixteen (4 males and 12 females) consecutive patients whose average age was 56 years (age range, 31-76 years). Inclusion criteria were gold weight implant, lagophthalmos of 2 mm or more, and a House-Brackmann score of 3 or less at the completion of follow-up. Mean follow-up was 13 months. INTERVENTIONS: Each patient received a gold weight implant. Six of these patients underwent a lower eyelid procedure. MAIN OUTCOME MEASURES: Surgical complications, static and dynamic lagophthalmos, static and dynamic corneal coverage, visual acuity, keratitis, topical treatment, and patient satisfaction. RESULTS: There were no extrusions. The preoperative mean lagophthalmos was 7.5 mm and the postoperative mean was 0.5 mm, (P<.001). Corneal coverage with eye closure before implantation was 73% and after implantation was 100%, (P<.001). Corneal coverage with normal (reflex) blink was less than 50% in 9 of 14 patients. When wearing correction, no patients had 20/20 visual acuity. The mean patient satisfaction score before the procedure was 3.5 and after was 7.1, (P<.001). Patient satisfaction was most closely related to visual acuity. The relationship was linear and statistically significant (P<.04). CONCLUSIONS: Gold weight implantation provides significant reduction in lagophthalmos and significant improvement in corneal coverage. But owing to delayed closure time and disrupted tear film, irritation may persist. As a result, some patients require ongoing topical treatment of the eye, which can compromise visual acuity. PMID- 11255476 TI - Incidental parathyroidectomy during thyroid surgery does not cause transient symptomatic hypocalcemia. AB - OBJECTIVES: To identify any risk factors for incidental parathyroidectomy and to define its association with symptomatic postoperative hypocalcemia. DESIGN: Retrospective study. SETTING: Tertiary referral cancer center. PATIENTS: Consecutive patients who underwent thyroid surgery between 1991 and 1999. Patients who underwent procedures for locally advanced thyroid cancer requiring laryngectomy, tracheal resection, or esophagectomy were excluded. INTERVENTIONS: All pathology reports were reviewed for the presence of any parathyroid tissue in the resected specimen. Slides were reviewed, and information regarding patient demographics, diagnosis, operative details, and postoperative complications was collected. MAIN OUTCOME MEASURE: Identification of parathyroid tissue in resected specimens and postoperative symptomatic hypocalcemia. RESULTS: A total of 141 thyroid procedures were performed: 69 total thyroidectomies (49%) and 72 total thyroid lobectomies (51%). The findings were benign in 68 cases (48%) and malignant in 73 cases (52%). In the entire series, incidental parathyroidectomy was found in 21 cases (15%). Parathyroid tissue was found in intrathyroidal (50%), extracapsular (31%), and central node compartment (19%) sites. The performance of a concomitant modified radical neck dissection was associated with an increased risk of unplanned parathyroidectomy (P =.05). There was no association of incidental parathyroidectomy with postoperative hypocalcemia (P =.99). Multivariate analysis identified total thyroidectomy as a risk factor for postoperative hypocalcemia (P =.008). In the entire study group, transient symptomatic hypocalcemia occurred in 9 patients (6%), and permanent hypocalcemia occurred in 1 patient who underwent a total thyroidectomy and concomitant neck dissection. CONCLUSIONS: Unintended parathyroidectomy, although not uncommon, is not associated with symptomatic postoperative hypocalcemia. Modified radical neck dissection may increase the risk of incidental parathyroidectomy. Most of the glands removed were intrathyroidal, so changes in surgical technique are unlikely to markedly reduce this risk. PMID- 11255477 TI - Dynamic tissue expansion of the larynx in a canine model. AB - OBJECTIVES: To test whether staged, progressive, monitored, dynamic tissue expansion is possible in the larynx and to evaluate its effectiveness in dilating and augmenting constricting cicatricial lesions. DESIGN: Animal study. SETTING: Research facility, tertiary care medical center. SUBJECTS: Thirteen dogs, 3 with laryngotracheal stenosis. INTERVENTIONS: Dogs underwent laryngeal splits, tracheostomy, and insertion of inflatable stents. In 7 normal dogs, stents were progressively inflated by air in predetermined increments during 11 days. In 3 normal dogs and 3 with laryngotracheal stenosis, stents were gradually expanded by water. Stents were kept in place for 21 days. After removal, dogs were observed for 25 days. Five died of complications of tracheostomy. MAIN OUTCOME MEASURES: Airway diameter measured by endoscopy before the induction of stenosis, before the laryngeal splitting procedure, after stent removal, and before euthanasia. RESULTS: The lumen increased, then shrank somewhat after stent removal. In 2 surviving dogs with laryngotracheal stenosis and water-expanded stents, the lumen was 82.5% larger than baseline at stent removal and 71.0% larger at euthanasia. In 2 surviving normal dogs with water-expanded stents, lumen size increased by 50.0% at stent removal, and in 1 dog surviving to day 46, it was 17.0% larger. In 5 surviving dogs with air-inflated stents, lumen size was 39.0% larger at stent removal and 8.0% larger at day 46. Histologically, fibrous tissue developed in the gaps between the splayed margins of the laryngeal cartilages. CONCLUSIONS: The larynx may be dynamically expanded. Although the maximal diameter is not maintained, final cross-sectional areas are larger. PMID- 11255478 TI - Individual monitoring of aspirin desensitization. AB - BACKGROUND: Patients with aspirin-sensitive rhinosinusitis, which is frequently associated with intrinsic bronchial asthma, can be desensitized by long-term treatment with oral aspirin. The exact mechanisms of this desensitization remain obscure, but modulations of the eicosanoid pathway occur and can be monitored with the help of a practicable in vitro assay on mixed leukocyte cultures. OBJECTIVE: To monitor the effect of low-dose aspirin desensitization therapy, 100 mg/d, objectively by an in vitro assay. DESIGN: In a prospective study, 30 patients with aspirin intolerance, who were treated following a desensitization protocol with a dose of oral aspirin of only 100 mg/d were followed up for 1 year and reassessed every 3 months clinically and in vitro. RESULTS: Twenty-five patients showed a normalization of in vitro eicosanoid levels during this period, 4 showed some improvement, and 1 showed no therapeutic effect on eicosanoid release. Clinical follow-up revealed a low recurrence rate of nasal polyposis, with recurrent disease only in 4 individuals who also showed no normalization of eicosanoid release levels. Furthermore, a reduction of the average incidence of purulent episodes of sinusitis was seen after 1 year. Of 12 patients with asthma, 9 experienced marked improvement in pulmonary function. Of 16 individuals with a marked impairment of nasal breathing, 14 felt an increase of nasal patency, and 7 of 11 patients with pretreatment hyposmia had an improved sense of smell after 1 year. CONCLUSIONS: Desensitization therapy in patients with aspirin-sensitive rhinosinusitis can be successfully performed with low oral doses of aspirin, and the individual course throughout the desensitization can be monitored with the help of an in vitro analysis of eicosanoid release from mixed leukocyte cultures. PMID- 11255479 TI - Familial bilateral vocal cord paralysis and Charcot-Marie-tooth disease type II C. PMID- 11255480 TI - Neurofibroma of the larynx in neurofibromatosis: preoperative computed tomography and magnetic resonance imaging. AB - Neurofibromas of the larynx are extremely rare. They occur in association with neurofibromatosis less frequently than solitary neurofibromas. However, most laryngeal tumors in neurofibromatosis are neurofibromas. This disorder has 2 histological subtypes, which require different surgical approaches because of their biological differences. Few cross-sectional imaging studies have been performed in neurofibroma of the larynx. We describe a 44-year-old man with neurofibromatosis and nonplexiform neurofibroma of the larynx and discuss the role of preoperative computed tomography and magnetic resonance imaging in this case. The tumor was removed completely using an endolaryngeal approach without an external incision. It was possible to distinguish subtypes preoperatively on cross-sectional imaging. Magnetic resonance imaging provided more sensitive information in the diagnosis of this tumor than computed tomography. Preoperative cross-sectional imaging should be performed to help the surgeon diagnose and choose an appropriate surgical approach for this disorder. PMID- 11255481 TI - Pathology forum: quiz case. Diagnosis: intranasal glomus tumor. PMID- 11255482 TI - Radiology forum: quiz case. Diagnosis: rhabdomyosarcoma with perineural intracranial invasion. PMID- 11255483 TI - The regular practice of telemedicine: telemedicine in otolaryngology. PMID- 11255484 TI - The challenges and potential of otolaryngological telemedicine. PMID- 11255485 TI - Like it or not, telemedicine is here: making it work for us. PMID- 11255486 TI - Magnetic resonance imaging and computed tomographic findings after temporal lobe injury. PMID- 11255487 TI - Treatment of Frey syndrome with botulinum toxin type F. PMID- 11255488 TI - Gadolinium-enhanced MRI and sudden hearing loss. PMID- 11255490 TI - High cassava production and low dietary cyanide exposure in mid-west Nigeria. AB - OBJECTIVE: To investigate if high cassava production levels indicate high consumption and high dietary cyanide exposure in three villages situated within the area of Nigeria with higher cassava production than predicted by a geographic model for cassava production in Africa. DESIGN: Exploratory assessment of: cassava production and processing by qualitative research methods and quantification of residual cyanogens in products; cassava consumption by food frequency and weighed food records and dietary cyanide exposure by urinary thiocyanate and linamarin. SETTING: Rural communities of Afuze, Ebue and Ofabo in mid-west Nigeria. SUBJECTS: 110 subjects from 42 households in three villages for food frequency interviews; 118 subjects in nine Ofabo households for weighed food records. RESULTS: Cassava cultivation was reported to have increased in the preceding 20 years. It was consumed daily by 37 (88%) households, but its mean contribution to daily energy intake was only 13% The range of residual cyanogens in cassava foods was 0 to 62 mg HCN equivalent/kg dry weight (dw). Ten samples (19%) had levels above the 10 mg HCN equivalent/kg dw FAO/WHO safety limit. Mean urinary thiocyanate and linamarin were 51 and 20 micromol/L, indicating low cyanogen intake and dietary cyanide exposure. CONCLUSION: High cassava production levels did not result in high consumption and high dietary cyanide exposure levels, therefore cassava production levels cannot be used to predict consumption or cyanide exposure levels in the study area. A large part of the production is explained by intensive sales. PMID- 11255491 TI - Nutrition and handgrip strength of older adults in rural Malawi. AB - OBJECTIVE: To examine the relationship between the nutritional status and handgrip strength of older people in rural Malawi. DESIGN: Cross-sectional study. SETTING: : Lilongwe rural, Malawi, situated approximately 35-50 km from the city. SUBJECTS: Ninety seven males and 199 females participated in this study. METHODS: Selected anthropometric measurements were taken and nutrition indices were computed using standard equations. Handgrip strength was measured using an electronic grip strength dynamometer. RESULTS: The mean handgrip strength (in kg) for men was significantly higher than for women vs. In addition, there was a significant decline in handgrip strength with age in both sexes. Furthermore, handgrip strength was positively correlated to the following nutritional status indicators: BMI in males and in females), mid-upper arm circumference (MUAC) in males and in females) and arm-muscle area (AMA) in males and in females). After controlling for potential confounders, namely sex, height and age, the correlations between handgrip strength and the nutrition indices were still significant. CONCLUSION: The results of this study support the hypothesis that poor nutritional status is associated with poor handgrip strength. Malawian males had both lower handgrip strength and lower arm muscle area than their counterparts from industrialised countries. However, Malawian females had similar handgrip strength despite lower arm muscle area, in comparison with women from industrialised countries, reflecting perhaps their higher level of physical activity. Further studies are required to determine whether by alleviating nutritional problems a concomitant improvement in handgrip strength can be obtained. PMID- 11255492 TI - The accuracy of self-reported weight by overweight and obese women in an outpatient setting. AB - OBJECTIVE: To investigate the accuracy of self-reported weight of overweight and obese women and characterisation of under-, correct- and over-reporters based on a number of related variables. DESIGN: Weight was self-reported before entering the study. At baseline, actual weight was recorded, and demographic, health, nutritional, psychological and physical activity questionnaires were completed. SETTING: A hospital outpatient department. SUBJECTS: Participants were 131 women aged 18-64 years with a body mass index attending a Comprehensive Weight Management Programme. OUTCOME MEASURES: The accuracy of self-reported weight was investigated for the total group, and the subjects were then categorised into three groups according to accuracy of self-reported weight (under-, correct- and over-reporters). The relationship between these accuracy groups and demographic, health, nutritional, and psychological variables and physical activity was examined, to characterise the under-, correct- and over-reporters. RESULTS: Although not statistically significantly different, the total group of women tended to underestimate their weight by 0.8 (+/-3.6) kg. Categorisation according to the accuracy groups revealed that 29% underestimated their weight by 2 kg or more, 19% overestimated their weight by 2 kg or more, and only 52% correctly estimated their weight within 2 kg. Some trends and statistically significant differences between the accuracy groups concerning certain variables, e.g. height, age, income, education, contraceptive pill usage, smoking and food choices were evident. CONCLUSIONS: Self-reported weight of a group of overweight/obese individuals may be a valid and reliable indicator of actual weight, but self-reported weight of an overweight/obese individual can not be interpreted similarly. Further research is necessary to ensure reliable characterisation of under-, over- and correct reporters. PMID- 11255493 TI - Association between nutrition knowledge and nutritional intake in middle-aged men from Northern France. AB - OBJECTIVE: The way in which nutrition knowledge transforms into dietary behaviour and nutrient intake may vary among populations. Therefore, the goal of the study was to examine whether nutrition knowledge is associated with nutritional intake in middle-aged men who are at major risk of cardiovascular disease. DESIGN: Cross sectional population study aimed at comparing the response to a nutrition quiz with food habits and nutrient intake determined by a 3-day food record. SETTING: Men of the Urban Community of Lille (France) examined at home. SUBJECTS: 361 men aged 45-64 y, randomly selected from the electoral rolls. RESULTS: Subjects were separated in a high-score and a low-score group according to their responses to the nutrition quiz. Subjects in the high-score group had better educational and higher income levels than those from the low-score group. Multivariate analysis, adjusting on educational and socio-economic levels and other confounding variables - such as age, body mass index, cigarette smoking, physical activity and energy intake underreporting - showed that subjects in the high-score group were more often consumers of olive oil (36 vs. 12%; ), cheese (85 vs. 76%; or cereals (27 vs. 15%; and less often consumers of sunflower oil (51 vs. 68%; or dry vegetables (12 vs. 22%; than those in the low-score group. Subjects in the high-score group had lower intakes of fat vs. and especially of monounsaturated fat of animal origin vs. than individuals in the low-score group. CONCLUSION: Nutrition quiz score is associated with specific patterns of food choices and nutrient intake suggesting that nutrition knowledge influences dietary behaviour in middle-aged men from Northern France. PMID- 11255494 TI - Disparities in vegetable and fruit consumption: European cases from the north to the south. AB - OBJECTIVE: To present disparities in consumption of vegetables and fruits in Europe and to discuss how educational level, region and level of consumption influence the variation. DESIGN: A review of selected studies from 1985 to 1997. SETTING/SUBJECTS: 33 studies (13 dietary surveys, nine household budget surveys and 11 health behaviour surveys) representing 15 European countries were selected based on criteria developed as part of the study. Association between educational level and consumption of vegetables and fruits was registered for each study and common conclusions were identified. RESULTS: In the majority of the studies, with the exception of a few in southern and eastern Europe, consumption of vegetables and fruits was more common among those with higher education. The results suggest that in regions where consumption of vegetables and fruits is more common, the lower social classes tend to consume more of these than the higher social classes. CONCLUSIONS: The differences in the patterns of disparities in vegetable and fruit consumption between regions, as well as within populations, need to be considered when efforts to improve nutrition and health are planned. PMID- 11255495 TI - Anaemia among non-pregnant women in rural Bangladesh. AB - OBJECTIVE: To estimate the prevalence and severity of anaemia among non-pregnant women in rural Bangladesh and describe its social distribution. DESIGN: A cross sectional study conducted in February-March 1996. Haemoglobin concentration was measured on a capillary blood sample by cyanmethaemoglobin method. The World Health Organization (WHO) classification was used to define anaemia. SETTING: Twelve randomly selected villages in Fulbaria thana of Mymensingh district, about 110 km northwest of Dhaka city in Bangladesh. SUBJECTS: A systematically selected sample of 179 non-pregnant apparently healthy women aged 15-45 years. RESULTS: Anaemia was highly prevalent (73%; 95%CI 67-79%). Most of the women had mild (52%) or moderate (20%) anaemia, but a few of them suffered from severe anaemia (1%). Ascaris was common (39%) while hookworm was not (1%). The anaemia prevalence had no statistically significant association with age, parity or Ascaris infestation Women with less than 1 year of schooling, who were landless or who reported having an economic deficit in the household had significantly higher prevalence of anaemia There was a significantly increasing trend in anaemia prevalence with decreasing socioeconomic situation (SES). However anaemia was common in all social strata. CONCLUSIONS: Although the overall anaemia prevalence among non-pregnant rural women is high, only a few women suffer from severe anaemia. Women of all SES groups irrespective of their age and parity are affected by anaemia. PMID- 11255496 TI - Towards healthier food policies - a new action plan for Europe. PMID- 11255498 TI - Nutrition and the health of Europe's children. PMID- 11255499 TI - Vitamin and mineral intakes in European children. Is food fortification needed? AB - OBJECTIVE: To provide an overview of vitamin and mineral intakes among children and adolescents in European countries and to present results from studies showing the impact of food fortification. DESIGN: Comparative analysis of a number of nutritional studies among children and adolescents performed during the last decade in certain European countries. SETTING: Spain, France, UK, North Ireland, Portugal, Germany. SUBJECTS: Europeans aged 6 to 18. RESULTS: Dietary surveys across Europe showed that varying levels of nutrient adequacy existed from one country to another, and that even within the same country, there were important nutritional gaps between different regions. In general, studies are difficult to compare, and information for many countries was missing. The results suggest that children and adolescents are the population group most likely to have higher risk of nutritional deficiencies, particularly for iron, vitamins C, E, B(6) and folates. In France, Ireland, UK and Spain, food fortification, and particularly of breakfast cereals, has positively contributed to increasing vitamin and mineral intakes in childhood and adolescence. CONCLUSIONS: Information on vitamin and mineral intakes in European children is less available than in adults. Fortified foods may contribute to reducing nutrient inadequacy in European children and adolescents, but should not replace nutrition education. PMID- 11255500 TI - Childhood obesity in Europe: a growing concern. AB - Estimation of the prevalence and secular trends in paediatric obesity in Europe is impeded by methodological problems in the definition of obesity and the paucity of data sets that mirror the demographic, cultural and socioeconomic composition of the European population. The available prevalence data show that paediatric obesity is increasing throughout Europe but the patterns vary with time, age, sex and geographical region. The highest rates of obesity are observed in eastern and southern European countries. Even within countries there may be marked variability in the rates of obesity. It is unclear whether these trends are a simple consequence of an overall increase in fatness in Europe or whether there may be sub-groups of children who, at certain ages, are either particularly susceptible to environmental challenges or are selectively exposed to such challenges. In addition to the general increase in adiposity in European youth, there is also evidence of an increasing degree of obesity, particularly in older children and adolescents. No definite conclusions can be made about the respective contribution of energy intake and physical activity to the increasing prevalence of obesity. Changing demographic and social circumstances are linked to childhood obesity but it is highly unlikely that these interact in similar ways in the genesis of obesity in different individuals and population groups. In conclusion, the limited understanding of the variability in susceptibility to obesity in European youth provides powerful justification for the development of preventive strategies which are population based rather than selectively targeted at high-risk children. PMID- 11255501 TI - Calcium, physical activity and bone health--building bones for a stronger future. AB - Adequate provision of nutrients composing the bone matrix and regulating bone metabolism should be provided from birth in order to achieve maximal bone mass, compatible with individual genetic background, and to prevent osteoporosis later in life. Low calcium intake (<250 mg day(-1)) in children is associated with both a reduced bone mineral content and hyperparathyroidism. Optimal calcium intake is, however, still a matter of controversy. The minimisation of fracture risk would be the ideal functional outcome on which to evaluate lifetime calcium intakes, but proxy indicators, such as bone mass measurements or maximal calcium retention, are used instead. Calcium recommendations in Europe and the United States are based on different concepts as to requirements, leading to somewhat different interpretations of dietary adequacy. Minerals and trace elements other than calcium are involved in skeletal growth, some of them as matrix constituents, such as magnesium and fluoride, others as components of enzymatic systems involved in matrix turnover, such as zinc, copper and manganese. Vitamins also play a role in calcium metabolism (e.g. vitamin D) or as co-factors of key enzymes for skeletal metabolism (e.g. vitamins C and K). Physical activity has different effects on bone depending on its intensity, frequency, duration and the age at which it is started. The anabolic effect on bone is greater in adolescence and as a result of weight-bearing exercise. Adequate intakes of calcium appear necessary for exercise to have its bone stimulating action. PMID- 11255502 TI - Establishing good dietary habits -- capturing the minds of children. AB - OBJECTIVE: To review the psycho-social research with respect to relevance for the development of nutritional education strategies. RESULTS: The eating behaviour of the newborn baby is controlled by innate preferences and dislikes, and by biological self-regulation. These innate control-systems are modified by learning processes, most importantly by the mere exposure to unknown food, by social influences, and by associating the physiological consequences of food intake with taste cues. The last decades have witnessed a change of the social meaning of food and eating, and the social context of eating is subject to dramatic changes. While on the one hand, prevalence of overweight and obesity is increasing, even young children deliberately practise weight control measures ranging from selective food choice to self-induced vomiting thus including behaviours which are clearly symptomatic of eating disorders. Such behaviour is motivated by unrealistic conceptions of a healthy body weight and shape. Children are interested in a range of nutrition topics. However, these topics have to be related to direct perceivable benefits from nutrition. CONCLUSIONS: Educational strategies should: firstly, focus on providing a variety of foods, including a range of nutrient-dense 'healthy' food and encouraging children to taste it; secondly, provide a stable and predictive pattern of social eating occasions to promote the social meaning and importance of eating and to enable social learning of food preferences; and finally, encourage a positive body image by providing advice and reassurance regarding the range of healthy and acceptable body weights and shapes. PMID- 11255503 TI - School-based nutrition education: lessons learned and new perspectives. AB - Nutrition is a major environmental influence on physical and mental growth and development in early life. Food habits during infancy can influence preferences and practices in later life and some evidence suggests fair to moderate tracking of food habits from childhood to adolescence. Studies support that good nutrition contributes to improving the wellbeing of children and their potential learning ability, thus contributing to better school performance. Children and young people who learn healthy eating habits, are encouraged to be physically active, to avoid smoking and to learn to manage stress, have the potential for reduced impact of chronic diseases in adulthood. Nutrition education is a key element to promoting lifelong healthy eating and exercise behaviours and should start from the early stages of life; it should also address the specific nutritional needs associated with pregnancy, including reinforcing breastfeeding. Food habits are complex in nature and multiple conditioning factors interact in their development. Young children do not choose what they eat, but their parents decide and prepare the food for them. During infancy and early childhood the family is a key environment for children to learn and develop food preferences and eating habits. As they grow and start school, teachers, peers and other people at school, together with the media and social leaders, become more important. Progressively children become more independent and start making their own food choices. The peer group is very important for adolescents and has a major influence in developing both food habits and lifestyles. Community trials suggest that nutrition education is an accessible effective tool in health promotion programmes with a focus on the development of healthy eating practices. PMID- 11255504 TI - Eat to live or live to eat? Do parents and children agree? AB - The lifestyles and diets of children in developed countries is changing rapidly in response to the social and cultural climate, as well as the availability of an increasing range of foods. The aim of this on-going study is to assess the trends in food-related behaviours of children and their attitudes towards food and nutrition. The study also monitors markers for physical activity, which have been presented elsewhere (Bellisle et al, 2000). Three successive surveys (1993, 1995, and 1997) were carried out on samples of 1000 French children aged 9--11 years and their mothers. RESULTS: Attitudes towards food varied markedly between children and their mothers. Children viewed food primarily as a necessity of life, whereas mothers viewed food primarily as a pleasure for their child, with necessity and nutrition given a lower importance. Contrary to popular belief children's enjoyment of 'unhealthy' and 'healthy' foods was similar. French-fries were the favourite food for 92% of children, closely followed by pasta (89%). Fruit and candy received similar scores (82% and 81% respectively), suggesting that external factors such as convenience are the prime barrier to fruit consumption rather than the enjoyment factor. Over the three surveys a strong persistence of the traditional French meal pattern has been demonstrated, with breakfast, lunch and evening meals eaten by 97%, 96% and 99% of children respectively. One increasing occurrence is the viewing of television during meal times with 25% of breakfasts, 46% of afternoon snacks and 41% of evening meals consumed in front of the television. Children's attitudes towards food demonstrated an overwhelming trust of their mother and her ability to provide them with nourishing foods. Mothers often found it difficult to reconcile the demands of family life, employment, healthy eating and the maintenance of a pleasurable atmosphere at mealtimes. CONCLUSIONS: Overall, children and mothers appeared to have divergent attitudes towards food. An understanding of children's enjoyment of foods considered more 'healthy' or less 'healthy' enables effective nutrition messages to be developed for children. However, further work is needed to explore the long-term impact of mothers' attitudes towards food and mealtimes on a child's relationship to food and eating habits during adolescence through to adulthood. PMID- 11255506 TI - Retinitis pigmentosa: mutations in a receptor tyrosine kinase gene, MERTK. PMID- 11255505 TI - Triclosan and fatty acid synthesis in Plasmodium falciparum: new weapon for an old enemy. PMID- 11255507 TI - Getting the beat. PMID- 11255508 TI - Non-redundant tumour supressor functions of transforming growth factor beta in breast cancer. PMID- 11255509 TI - Glutamate synthase: an archaeal horizontal gene transfer? PMID- 11255510 TI - Information theory: a guide in the investigation of disease. PMID- 11255511 TI - Developmental regulation and complex organization of the promoter of the non coding hsr(omega) gene of Drosophila melanogaster. AB - The nucleus-limited large non-coding hsr(omega)-n RNA product of the 93D or the hsr(omega) gene of Drosophila melanogaster binds to a variety of RNA-binding proteins involved in nuclear RNA processing. We examined the developmental and heat shock induced expression of this gene by in situ hybridization of nonradioactively labelled riboprobe to cellular transcripts in intact embryos, larval and adult somatic tissues of wild type and an enhancer-trap line carrying the hsr(omega) 05241 allele due to insertion of a P-LacZ-rosy+ transposon at -130 bp position of the hsr(omega) promoter. We also examined LacZ expression in the enhancer-trap line and in two transgenic lines carrying different lengths of the hsr(omega) promoter upstream of the LacZ reporter. The hsr(omega) gene is expressed widely at all developmental stages; in later embryonic stages, its expression in the developing central nervous system was prominent. In spite of insertion of a big transposon in the promoter, expression of the hsr(omega) 05241 allele in the enhancer-trap line, as revealed by in situ hybridization to hsr(omega) transcripts in cells, was similar to that of the wild type allele in all the embryonic, larval and adult somatic tissues examined. Expression of the LacZ gene in this enhancer-trap line was similar to that of the hsr(omega) RNA in all diploid cell types in embryos and larvae but in the polytene cells, the LacZ gene did not express at all, neither during normal development nor after heat shock. Comparison of the expression patterns of hsr(omega) gene and those of the LacZ reporter gene under its various promoter regions in the enhancer-trap and transgenic lines revealed a complex pattern of regulation, which seems to be essential for its dynamically varying expression in diverse cell types. PMID- 11255512 TI - Characterization of chicken riboflavin carrier protein gene structure and promoter regulation by estrogen. AB - The chicken riboflavin carrier protein (RCP) is an estrogen induced egg yolk and white protein. Eggs from hens which have a splice mutation in RCP gene fail to hatch, indicating an absolute requirement of RCP for the transport of riboflavin to the oocyte. In order to understand the mechanism of regulation of this gene by estrogen, the chicken RCP gene including 1 kb of the 5' flanking region has been isolated. Characterization of the gene structure shows that it contains six exons and five introns, including an intron in the 5' untranslated region. Sequence analysis of the 5' flanking region does not show the presence of any classical, palindromic estrogen response element (ERE). However, there are six half site ERE consensus elements. Four deletion constructs of the 5' flanking region with varying number of ERE half sites were made in pGL3 basic vector upstream of the luciferase-coding region. Transient transfection of these RCP promoter deletion constructs into a chicken hepatoma cell line (LMH2A) showed 6-12-fold transcriptional induction by a stable estrogen analogue, moxesterol. This suggests that the RCP gene is induced by estrogen even in the absence of a classical ERE and the half sites of ERE in this promoter may be important for estrogen induction PMID- 11255513 TI - Early localization of NPA58, a rat nuclear pore-associated protein, to the reforming nuclear envelope during mitosis. AB - We have studied the mitotic reassembly of the nuclear envelope, using antibodies to nuclear marker proteins and NPA58 in F-111 rat fibroblast cells. In earlier studies we have proposed that NPA58, a 58 kDa rat nuclear protein, is involved in nuclear protein import. In this report, NPA58 is shown to be localized on the cytoplasmic face of the envelope in interphase cells, in close association with nuclear pores. In mitotic cells NPA58 is dispersed in the cytoplasm till anaphase. The targeting of NPA58 to the reforming nuclear envelope in early telophase coincides with the recruitment of a well-characterized class of nuclear pore proteins recognized by the antibody mAb 414, and occurs prior to the incorporation of lamin B1 into the envelope. Significant protein import activity is detectable only after localization of NPA58 in the newly-formed envelope. The early targeting of NPA58 is consistent with its proposed role in nuclear transport. PMID- 11255515 TI - Incipient sexual isolation in the nasuta-albomicans complex of Drosophila: no choice experiments. AB - Drosophila nasuta nasuta and Drosophila nasuta albomicans are cross-fertile races of Drosophila. Hybridization between these races in the laboratory has given rise to new races (Cytoraces), among which karyotypic composition differs from one another and also from those of the parental races. In this study, we search for the evidence of incipient reproductive isolation among the parental races and four Cytoraces by assessing the fraction of no-matings, mating latency and copulation duration in all possible types of homo- and heterogamic crosses (N = 4184). In no-choice conditions, the latency time (time to initiation of copulation) is lower in homogamic crosses than in heterogamic crosses for both parental races and Cytoraces. Latency time and copulation duration are negatively correlated, whereas fraction of no matings is positively correlated with latency time. Thus these six closely related races of the nasuta-albomicans complex show the initiation of the earliest stages of pre-zygotic isolation, manifested as a tendency for matings to be initiated earlier and more often, and for a longer duration, among homogamic rather than heterogamic individuals PMID- 11255514 TI - Pleiotropic morphological and abiotic stress resistance phenotypes of the hyper abscisic acid producing Abo- mutant in the periwinkle Catharanthus roseus. AB - The pleiotropic properties of a abo abo (Abo-) gamma-ray induced mutant of Catharanthus roseus cv. Nirmal, selected among the M2 generation seeds for ability to germinate at 45 degrees C, are described. The mutant produced seeds possessing tricotyledonous embryos, unlike the typically dicotyledonous embryos present in the wild type Abo+ seeds. In comparison to Abo+ adults, the mutant plants had short stature and lanceolate leaves. The vascular bundles in the leaves and stem were poorly developed. Leaf surfaces were highly trichomatous, epidermal, cortex and mesophyll cells were small sized and a large majority of stomata were closed. Besides high temperature, the mutant was salinity and water stress tolerant. The abscisic acid (ABA) content in the leaves was about 500-fold higher. The genetic lesion abo responsible for the above pleiotropy was recessive and inherited in Mendelian fashion. The seedlings and adult plants of the mutant accumulated higher proline than Abo+ plants. The phenotypes of abo abo mutants permitted the conclusions that (i) the mutant synthesizes ABA constitutively, (ii) both ABA-dependent and ABA independent pathways for proline and betaine accumulation are functional in the mutant, and (iii) cell division, elongation and differentiation processes in embryo and adult plant stages are affected in the mutant PMID- 11255516 TI - Biological factors underlying regularity and chaos in aquatic ecosystems: simple models of complex dynamics. AB - This work is focused on the processes underlying the dynamics of spatially inhomogeneous plankton communities. We demonstrate that reaction-diffusion mathematical models are an appropriate tool for searching and understanding basic mechanisms of complex spatio-temporal plankton dynamics and fractal properties ofplanktivorous fish school walks PMID- 11255517 TI - Chaos and regular dynamics in model multi-habitat plankton-fish communities. AB - This work is focused on the role of diffusive interaction between separate habitats in a patchy environment in plankton pattern formation. We demonstrate that conceptual reaction-diffusion mathematical models constitute an appropriate tool for searching and understanding basic mechanisms of plankton pattern formation and complex spatio-temporal plankton dynamics PMID- 11255518 TI - Risk factors for coronary heart disease in men 18 to 39 years of age. AB - BACKGROUND: Few studies have evaluated the long-term predictive capacity of risk factors for death from coronary heart disease in men younger than 40 years of age. OBJECTIVE: To assess the predictive capacity and discriminatory ability of major coronary risk factors in predicting death from coronary heart disease in young men. DESIGN: Prospective cohort study with 20 years of follow-up. SETTING: 84 companies in the Chicago area that participated in the Chicago Heart Association Detection Project in Industry (1967-1973). PARTICIPANTS: 11 016 men 18 to 39 years of age (mean age, 29.7 years) at baseline were the primary focus of this report; 8955 men 40 to 59 years of age at baseline served as a reference group. MEASUREMENTS: The main end point was death from coronary heart disease. RESULTS: All major risk factors-age, serum cholesterol level, systolic blood pressure, and cigarette smoking-were significantly associated with death from coronary heart disease over 20 years in young men. Relative risks for the major risk factors were of generally similar magnitude in young and middle-aged men. Receiver-operating characteristic curves for the best predictive model yielded an area under the curve of 0.82, indicating that standard risk factors were highly predictive of long-term outcome in young men. CONCLUSIONS: Major coronary disease risk factors, many of which are modifiable, are strong contributors to prediction of future risk, even in young men. These data may help in formulating appropriate strategies to identify young men at heightened risk for death from coronary heart disease in later adulthood. PMID- 11255519 TI - Use of genotypic resistance testing to guide hiv therapy: clinical impact and cost-effectiveness. AB - BACKGROUND: Genotypic sequencing for drug-resistant strains of HIV can guide the choice of antiretroviral therapy. OBJECTIVE: To assess the cost-effectiveness of genotypic resistance testing for patients acquiring drug resistance through failed treatment (secondary resistance) and those infected with resistant virus (primary resistance). DESIGN: Cost-effectiveness analysis with an HIV simulation model incorporating CD4 cell count and HIV RNA level as predictors of disease progression. DATA SOURCES: Published randomized trials and data from the Multicenter AIDS Cohort Study, the national AIDS Cost and Services Utilization Survey, the Red Book, and an institutional cost-accounting system. TARGET POPULATION: HIV-infected patients in the United States with baseline CD4 counts of 0.250 x 10(9) cells/L. TIME HORIZON: Lifetime. PERSPECTIVE: Societal. INTERVENTIONS: Genotypic resistance testing and clinical judgment, compared with clinical judgment alone, in two contexts: after initial treatment failure (secondary resistance testing) and before initiation of antiretroviral therapy (primary resistance testing). OUTCOME MEASURES: Life expectancy, quality-adjusted life expectancy, and cost-effectiveness in dollars per quality-adjusted life-year (QALY) gained. RESULTS OF BASE-CASE ANALYSIS: Secondary resistance testing increased life expectancy by 3 months, at a cost of $17 900 per QALY gained. The cost-effectiveness of primary resistance testing was $22 300 per QALY gained with a 20% prevalence of primary resistance but increased to $69 000 per QALY gained with 4% prevalence. RESULTS OF SENSITIVITY ANALYSIS: The cost-effectiveness ratio for secondary resistance testing remained under $25 000 per QALY gained, even when effectiveness and cost of testing and antiretroviral therapy, quality-of life weights, and discount rate were varied. CONCLUSIONS: Genotypic antiretroviral resistance testing following antiretroviral failure is cost effective. Primary resistance testing also seems to be reasonably cost-effective and will become more so as the prevalence of primary resistance increases. PMID- 11255521 TI - Endoscopic ultrasonography-guided fine-needle aspiration biopsy of suspected pancreatic cancer. AB - BACKGROUND: In many institutions, computed tomography (CT)-guided percutaneous fine-needle aspiration (FNA) has become the procedure of choice for biopsies of pancreatic mass lesions. This method of biopsy and others, such as endoscopic retrograde cholangiopancreatography (ERCP) cytology, are problematic because of a substantial false-negative rate. OBJECTIVE: To investigate the yield of endoscopic ultrasonography-guided FNA biopsies in patients who had negative results on CT-guided biopsy or negative cytologic findings on ERCP sampling. DESIGN: Prospective cohort study. SETTING: Tertiary care university medical center. PATIENTS: 102 patients (median age, 65 years; 58 men and 44 women) with suspected pancreatic cancer who fulfilled the above criteria were prospectively identified and underwent endoscopic ultrasonography-guided FNA biopsy. MEASUREMENTS: The operating characteristics of endoscopic ultrasonography-guided FNA for diagnosing pancreatic masses were determined. Surgical pathology or long term follow-up (median, 24 months) was used to identify false-positive or false negative results. RESULTS: Median mass size was 3.5 cm x 2.7 cm. A median of 3.4 passes were performed. Cytologic results on endoscopic ultrasonography-guided FNA biopsy were positive in 57 patients, negative in 37, and inconclusive or nondiagnostic in 8. No false-positive results were observed. A diagnosis of pancreatic cancer was subsequently confirmed in 3 patients who had tested negative (false-negative results) and 1 of the 8 patients with nondiagnostic results. Of these 4 patients, 3 had cytologic evidence of chronic pancreatitis on endoscopic ultrasonography-guided FNA biopsy. The 95% CI for the likelihood ratio for a positive test result contained all values greater than or equal to 9.7. The likelihood ratio for a negative test result was 0.05 (CI, 0.02 to 0.15). The posterior probability of pancreatic cancer after a definitely positive result was at least 93.5% by a conservative lower 95% confidence limit; after a definitely negative test result, it was 6.9%. The prevalence of pancreatic cancer was 59.8% (61 of 102 patients). Self-limited complications occurred in 3 of the 102 patients (2.9% [CI, 0.6% to 8.4%]). CONCLUSION: Endoscopic ultrasonography-guided FNA biopsy may play a valuable role in the evaluation of a pancreatic mass when results on other biopsy methods are negative but pancreatic cancer is suspected. PMID- 11255520 TI - Recovery of ventricular function after myocardial infarction in the reperfusion era: the healing and early afterload reducing therapy study. AB - BACKGROUND: Patients with reduced left ventricular function and ventricular enlargement after myocardial infarction are at significantly greater risk for congestive heart failure and death. Nevertheless, recovery of ventricular function occurs in a significant proportion of patients after myocardial infarction, and modern reperfusion strategies have been associated with increased recovery of function. OBJECTIVE: To determine the extent and predictors of recovery of ventricular function after anterior Q-wave myocardial infarction in the reperfusion era. DESIGN: Subgroup analysis of the Healing and Early Afterload Reducing Therapy study. SETTING: 35 medical centers in the United States and Canada. PATIENTS: 352 patients with Q-wave anterior myocardial infarction. INTERVENTION: Placebo for 14 days, followed by full-dose (10 mg) ramipril until day 90; low-dose (0.625 mg) ramipril for 90 days; or full-dose ramipril for 90 days. All patients underwent reperfusion therapy. MEASUREMENTS: Echocardiography was performed on day 1 (before randomization), day 14, and day 90 after myocardial infarction. Left ventricular volume and ejection fraction were measured and wall-motion analyses were performed at all three time points in 249 patients and at baseline in an additional 12 patients who died during follow-up. Echocardiographic and nonechocardiographic predictors of ventricular recovery were examined. RESULTS: By day 90, 55 of 252 (22%) patients who had abnormal ejection fraction and wall-motion abnormalities on day 1 demonstrated complete recovery of function (ejection fraction in the normal range and infarct segment length of 0%), and an additional 36% (91 of 252 patients) demonstrated partial recovery of function. At 90 days, 53% (132 of 249) of patients had greater than 5% improvement in ejection fraction, whereas only 16% (39 of 249) had a decrease in ejection fraction of more than 5%. The majority of functional improvement occurred by day 14 after infarction. Of various clinical and echocardiographic measures obtained on day 1, peak creatine kinase level was the strongest independent predictor of subsequent recovery of ventricular function in multivariate analysis. Each 100-unit increase in peak creatine kinase was associated with a 4.3% decreased odds of recovery (P < 0.001) after adjustment for ejection fraction on day 1, extent of akinesis or dyskinesis, treatment regimen, Killip class, age, and sex. CONCLUSION: Significant myocardial stunning with subsequent improvement of ventricular function occurred in the majority of patients after Q-wave anterior myocardial infarction. A lower peak level of creatine kinase, an estimate of the extent of necrosis, is independently predictive of recovery of function. Early functional assessment (day 1 after acute myocardial infarction) had limited ability to predict recovery of ventricular function. PMID- 11255522 TI - Warfarin therapy for an octogenarian who has atrial fibrillation. AB - In North America, atrial fibrillation is associated with at least 75 000 ischemic strokes each year. Most of these strokes occur in patients older than 75 years of age. The high incidence of stroke in very elderly persons reflects the increasing prevalence of atrial fibrillation that occurs with advanced age, the high incidence of stroke in elderly patients, and the failure of physicians to prescribe antithrombotic therapy in most of these patients. This failure is related to the increased risk for major hemorrhage with advanced age, obfuscating the decision to institute stroke prophylaxis with antithrombotic therapy. This case-based review describes the risk and benefits of prescribing antithrombotic therapy for a hypothetical 80-year-old man who has atrial fibrillation and hypertension, and it offers practical advice on managing warfarin therapy. After concluding that the benefits of warfarin outweigh its risks in this patient, we describe how to initiate warfarin therapy cautiously and how to monitor and dose the drug. We then review five recent randomized, controlled trials that document the increased risk for stroke when an international normalized ratio (INR) of less than 2.0 is targeted among patients with atrial fibrillation. Next, we make the case that cardioversion is not needed for this asymptomatic patient with chronic atrial fibrillation. Instead, we choose to leave the patient in atrial fibrillation and to control his ventricular rate with atenolol. Later, when the INR increases to 4.9, we advocate withholding one dose of warfarin and repeating the INR test. Finally, when the patient develops dental pain, we review the analgesic agents that are safe to take with warfarin and explain why warfarin therapy does not have to be interrupted during a subsequent dental extraction. PMID- 11255523 TI - HIV resistance testing in clinical practice: a Qaly-fied success. PMID- 11255524 TI - Principles of appropriate antibiotic use for treatment of acute respiratory tract infections in adults: background, specific aims, and methods. AB - The need to decrease excess antibiotic use in ambulatory practice has been fueled by the epidemic increase in antibiotic-resistant Streptococcus pneumoniae. The majority of antibiotics prescribed to adults in ambulatory practice in the United States are for acute sinusitis, acute pharyngitis, acute bronchitis, and nonspecific upper respiratory tract infections (including the common cold). For each of these conditions-especially colds, nonspecific upper respiratory tract infections, and acute bronchitis (for which routine antibiotic treatment is not recommended)-a large proportion of the antibiotics prescribed are unlikely to provide clinical benefit to patients. Because decreasing community use of antibiotics is an important strategy for combating the increase in community acquired antibiotic-resistant infections, the Centers for Disease Control and Prevention convened a panel of physicians representing the disciplines of internal medicine, family medicine, emergency medicine, and infectious diseases to develop a series of "Principles of Appropriate Antibiotic Use for Treatment of Acute Respiratory Tract Infections in Adults." These principles provide evidence based recommendations for evaluation and treatment of adults with acute respiratory illnesses.This paper describes the background and specific aims of and methods used to develop these principles. The goal of the principles is to provide clinicians with practical strategies for limiting antibiotic use to the patients who are most likely to benefit from it. These principles should be used in conjunction with effective patient educational campaigns and enhancements to the health care delivery system that facilitate nonantibiotic treatment of the conditions in question. PMID- 11255525 TI - Principles of appropriate antibiotic use for treatment of nonspecific upper respiratory tract infections in adults. PMID- 11255526 TI - Principles of appropriate antibiotic use for treatment of nonspecific upper respiratory tract infections in adults: background. AB - The following principles of appropriate antibiotic use for adults with nonspecific upper respiratory tract infections apply to immunocompetent adults without complicating comorbid conditions, such as chronic lung or heart disease.1. The diagnosis of nonspecific upper respiratory tract infection or acute rhinopharyngitis should be used to denote an acute infection that is typically viral in origin and in which sinus, pharyngeal, and lower airway symptoms, although frequently present, are not prominent. 2. Antibiotic treatment of adults with nonspecific upper respiratory tract infection does not enhance illness resolution and is not recommended. Studies specifically testing the impact of antibiotic treatment on complications of nonspecific upper respiratory tract infections have not been performed in adults. Life-threatening complications of upper respiratory tract infection are rare.3. Purulent secretions from the nares or throat (commonly observed in patients with uncomplicated upper respiratory tract infection) predict neither bacterial infection nor benefit from antibiotic treatment. PMID- 11255527 TI - Principles of appropriate antibiotic use for acute sinusitis in adults. PMID- 11255528 TI - Principles of appropriate antibiotic use for acute rhinosinusitis in adults: background. AB - The following principles of appropriate antibiotic use for adults with acute rhinosinusitis apply to the diagnosis and treatment of acute maxillary and ethmoid rhinosinusitis in adults who are not immunocompromised.1. Most cases of acute rhinosinusitis diagnosed in ambulatory care are caused by uncomplicated viral upper respiratory tract infections. 2. Bacterial and viral rhinosinusitis are difficult to differentiate on clinical grounds. The clinical diagnosis of acute bacterial rhinosinusitis should be reserved for patients with rhinosinusitis symptoms lasting 7 days or more who have maxillary pain or tenderness in the face or teeth (especially when unilateral) and purulent nasal secretions. Patients with rhinosinusitis symptoms that last less than 7 days are unlikely to have bacterial infection, although rarely some patients with acute bacterial rhinosinusitis present with dramatic symptoms of severe unilateral maxillary pain, swelling, and fever.3. Sinus radiography is not recommended for diagnosis in routine cases. 4. Acute rhinosinusitis resolves without antibiotic treatment in most cases. Symptomatic treatment and reassurance is the preferred initial management strategy for patients with mild symptoms. Antibiotic therapy should be reserved for patients with moderately severe symptoms who meet the criteria for the clinical diagnosis of acute bacterial rhinosinusitis and for those with severe rhinosinusitis symptoms-especially those with unilateral facial pain-regardless of duration of illness. For initial treatment, the most narrow spectrum agent active against the likely pathogens, Streptococcus pneumoniae and Haemophilus influenzae, should be used. PMID- 11255529 TI - Principles of appropriate antibiotic use for acute pharyngitis in adults. PMID- 11255531 TI - Principles of appropriate antibiotic use for treatment of acute bronchitis in adults. PMID- 11255530 TI - Principles of appropriate antibiotic use for acute pharyngitis in adults: background. AB - The following principles of appropriate antibiotic use for adults with acute pharyngitis apply to immunocompetent adults without complicated comorbid conditions, such as chronic lung or heart disease, and history of rheumatic fever. They do not apply during known outbreaks of group A streptococcus.1. Group A beta-hemolytic streptococcus (GABHS) is the causal agent in approximately 10% of adult cases of pharyngitis. The large majority of adults with acute pharyngitis have a self-limited illness, for which supportive care only is needed.2. Antibiotic treatment of adult pharyngitis benefits only those patients with GABHS infection. All patients with pharyngitis should be offered appropriate doses of analgesics and antipyretics, as well as other supportive care.3. Limit antibiotic prescriptions to patients who are most likely to have GABHS infection. Clinically screen all adult patients with pharyngitis for the presence of the four Centor criteria: history of fever, tonsillar exudates, no cough, and tender anterior cervical lymphadenopathy (lymphadenitis). Do not test or treat patients with none or only one of these criteria, since these patients are unlikely to have GABHS infection. For patients with two or more criteria the following strategies are appropriate: a) Test patients with two, three, or four criteria by using a rapid antigen test, and limit antibiotic therapy to patients with positive test results; b) test patients with two or three criteria by using a rapid antigen test, and limit antibiotic therapy to patients with positive test results or patients with four criteria; or c) do not use any diagnostic tests, and limit antibiotic therapy to patients with three or four criteria. 4. Throat cultures are not recommended for the routine primary evaluation of adults with pharyngitis or for confirmation of negative results on rapid antigen tests when the test sensitivity exceeds 80%. Throat cultures may be indicated as part of investigations of outbreaks of GABHS disease, for monitoring the development and spread of antibiotic resistance, or when such pathogens as gonococcus are being considered.5. The preferred antibiotic for treatment of acute GABHS pharyngitis is penicillin, or erythromycin in a penicillin-allergic patient. PMID- 11255532 TI - Principles of appropriate antibiotic use for treatment of uncomplicated acute bronchitis: background. AB - The following principles of appropriate antibiotic use for adults with acute bronchitis apply to immunocompetent adults without complicating comorbid conditions, such as chronic lung or heart disease.1. The evaluation of adults with an acute cough illness or a presumptive diagnosis of uncomplicated acute bronchitis should focus on ruling out serious illness, particularly pneumonia. In healthy, nonelderly adults, pneumonia is uncommon in the absence of vital sign abnormalities or asymmetrical lung sounds, and chest radiography is usually not indicated. In patients with cough lasting 3 weeks or longer, chest radiography may be warranted in the absence of other known causes.2. Routine antibiotic treatment of uncomplicated acute bronchitis is not recommended, regardless of duration of cough. If pertussis infection is suspected (an unusual circumstance), a diagnostic test should be performed and antimicrobial therapy initiated.3. Patient satisfaction with care for acute bronchitis depends most on physician patient communication rather than on antibiotic treatment. PMID- 11255534 TI - My first fee. PMID- 11255535 TI - Distant healing. PMID- 11255536 TI - Distant healing. PMID- 11255538 TI - The alcohol hangover. PMID- 11255540 TI - Self-study from web-based and printed guideline material. PMID- 11255545 TI - Physicians for human rights. PMID- 11255542 TI - Hypothyroidism in two patients after hepatic arterial chemoembolization. PMID- 11255546 TI - Infertility and the establishment of pregnancy--overview. PMID- 11255547 TI - In vitro maturation of oocytes. AB - Only about 400 of the one million oocytes present at birth will be ovulated, while the rest will die by atresia. The ability to rescue oocytes destined to die and mature them in vitro would provide invaluable information about folliculogenesis and oocyte maturation, and could provide oocytes for infertile women. In vitro maturation (IVM) is challenging in the human because folliculogenesis is a lengthy process encompassing many complex cellular changes in the oocyte and its surrounding follicle cells. A few live births have resulted from the maturation and fertilization of immature human oocytes aspirated from small antral follicles. Furthermore, it is possible to grow primordial follicles to pre-antral stages in slices of ovarian tissue, and support antrum formation in isolated pre-antral follicles. However, we are still a considerable way from growing and maturing pre-antral follicles to pre-ovulatory stages in vitro. The importance of the follicular environment for producing a healthy and developmentally competent oocyte is illustrated by the oocyte's susceptibility to errors during meiosis. This counsels considerable caution in the development of IVM for clinical application. PMID- 11255548 TI - Cryopreservation of gonadal tissue and cells. AB - With the advent of assisted reproductive technology and an improved understanding of cryobiology, strategies have been developed which allow the long-term storage of gametes and embryos. Furthermore, in the light of the growing numbers of young adults and children who have been sterilised by successful cancer treatment, the need to protect their fertile potential has become more urgent. While semen cryopreservation is available for men, the methods for preserving oocytes are unreliable and neither method is suitable for prepubertal children. Research attention is, therefore, focusing on the low temperature banking of immature germ cells with the aim to restore natural fertility if possible or, alternatively, to use culture technology to produce ripe gametes for assisted conception. While there is no universal method for fertility conservation, gonadal tissue banking, in theory, is a practical alternative to gamete storage which can be utilised by both adults and children. PMID- 11255549 TI - Male infertility and intracytoplasmic sperm injection (ICSI). AB - Micro-assisted fertilization in the form of intracytoplasmic sperm injection (ICSI) has truly revolutionised the treatment options for couples with impaired semen quality, and those with both obstructive and non-obstructive azoospermia. In general, the major issues which relate to the success of ICSI are those related to the success of conventional IVF, and the high multiple pregnancy rate remains a major cause for concern. There is growing evidence that the short-term health of ICSI offspring is relatively unremarkable, but our growing understanding of the genetic basis of much of male subfertility and of the impaired genomic integrity which characterises the oligozoospermic phenotype indicate a cautious approach to the longer term health of ICSI offspring. PMID- 11255550 TI - Lifestyle and environmental contribution to male infertility. AB - This chapter is an overview of recent developments in our understanding and thinking about the importance and nature of environmental effects on sperm counts and fertility in the human male. This area is plagued by imperfect studies, not necessarily because of imperfect design but because of other 'uncontrollable' constraints. The available data, therefore, need to be placed in context and account taken of the limitations of our understanding or, more correctly, our ignorance. As we enter the new millennium, one of the saddest scientific aspects of human reproduction and infertility is our persisting ignorance about the causes and treatment of male infertility. With one notable exception (Y chromosome microdeletions) there has been little advance in our understanding of the causes of male infertility and its direct treatment over the past 20 years. Although most infertile men can now be offered the chance of fertility via ICSI, it is largely ignored that this does not represent treatment of the patient's infertility (which will persist unchanged), but is a means of circumventing the problem and leaving it for the next generation to tackle. There are many reasons for our ignorance about the causes of infertility, and some of these are outlined below in order to emphasise how this limits our ability to establish whether or not specific lifestyle and environmental factors do, or do not, affect human male reproductive function. PMID- 11255551 TI - Premature ovarian failure. AB - On average, the menopause occurs at the age of 50 years, with 1% of women continuing to menstruate beyond the age of 60 years and 1% whose menopause occurs before 40 years. Arbitrarily, a menopause before the age of 40 years is defined as 'premature'. PMID- 11255552 TI - Genetic basis of male fertility. AB - We are in the age of genetic discovery. Now the human genome has been completely sequenced1, there will be increasing understanding and ability to manipulate biochemical pathways downstream of genes. At the same time, further development of in vitro fertilization (IVF) and intracytoplasmic sperm injection(ICSI) will enable procreation in situations that were formerly impossible and when there may be an increased possibility of genetic abnormality. Furthermore, preimplantation diagnosis will enable defects to be diagnosed and will give the opportunity for the couple to decide whether to continue with treatment towards a pregnancy or not. Thus, there is a need for clinicians to have a good knowledge of the gentics and hereditary aspects of male (and indeed female) infertility and for couples to have access to correct information and expert counselling. Also, there are ethical implications of these scientific and clinical advances for the future child, the individual, the couple and society. There is increasing public unease about this new science of reproduction and, in the UK, there is regulation by law; thus, there is a need for clinicians and scientists to give accurate information in everday language to the public. PMID- 11255553 TI - Pre-implantation genetic diagnosis. AB - Genetic disorders are a major cause of miscarriage and fetal death. Pre implantation genetic diagnosis (PGD) can be used to diagnose genetic defects before pregnancy has occurred by creating embryos by IVF, then removing single cells which are genetically analysed using FISH or PCR. Although successful, the techniques have many difficulties because they are highly specialised and at the extreme limit of sensitivity. Newer techniques, however, can rapidly diagnose multiple defects including chromosomal aneuploidy, sex and single gene defects. Embryonic cells can also be DNA fingerprinted to ensure that contamination has not occurred. As embryo screening can increase IVF success rates and decrease miscarriage rates, it will be increasing offered in routine IVF rather than just those patients at high genetic risk. These new, low cost techniques may ultimately allow PGD to be offered to all IVF patients regardless of risk. PMID- 11255554 TI - Advances in ultrasound assessment in the establishment and development of pregnancy. AB - Current data demonstrate that angiogenesis in the ovaries and uterus is an essential component of both follicular and luteal phases of menstrual cycle, tightly correlating with the levels of bioactive substances such as hormones, growth factors and interleukins. Ultrasound is used principally to demonstrate follicular growth, a receptive triple layer endometrium and to exclude pathologies such as fibroids and ovarian tumours. However, the development of new technologies such as CDI, CPA, 3D-US, 3D-CPA is now set to expand the role of ultrasound in the assessment of the processes in the ovaries, uterus and early pregnancy. There is growing evidence that studies of peri-follicular vascularity will predict the development of a healthy oocyte and subsequently an embryo. Endometrial blood flow studies with conventional CDI and the newer techniques of CPA and 3D-CPA will be important in predicting endometrial receptivity. Ovarian stromal vascularity appears to correlate with vascular endothelial growth factor (VEGF) levels and high vascularity is associated with PCO and a risk of ovarian hyperstimulation syndrome. 3D-CPA may improve our ability to assess ovarian and endometrial vascularization and blood circulation, to diagnose tubal patency. Increasingly, 3D ultrasound is being applied to diagnose the pathology of early singleton and multiple pregnancies. PMID- 11255555 TI - Overview of advances in contraception. AB - For most of man's existence, natural checks on fertility ensured that the numbers of the population more or less matched the resources available. It is only in the last few generations that man has so dominated the natural environment that the numbers of people in the world have increased exponentially, unchecked by natural disasters such as disease and starvation. Coincidental with extended life expectancy, the fertility rate of individual women has increased with the advent of bottle feeding and the decline in the contraceptive effects of breast feeding. Contraception has become increasingly important to individuals, allowing them to break free from the 'burden of excessive fertility'. PMID- 11255556 TI - Hormonal contraception in the male. AB - The hormonal approach to male contraception is based on the suppression of gonadotrophin secretion with secondary suppression of spermatogenesis. This can be achieved by administration of testosterone or other androgen alone, but combined administration with a progestogen or GnRH analogue allows the dose of testosterone to be reduced to physiological replacement doses. This approach has been investigated for many years but without identification of a regimen which results in sufficient suppression of spermatogenesis to provide ensured contraception in all men, safely and conveniently. The reasons for this are discussed, and recent developments towards a regimen that fulfills all these criteria are described. Crucial to development of any new product is that it will be used: surveys of both men and women indicate firmly positive attitudes towards a 'male pill'. There are, therefore, grounds for cautious optimism that the next decade may see the introduction of the first novel male contraceptive for several hundred years. PMID- 11255557 TI - Emergency contraception. AB - Knowledge and use of emergency contraception world-wide is extremely limited. Recent research has demonstrated that levonorgestrel alone is at least as effective as the Yuzpe regimen and is much better tolerated. Levonorgestrel is likely to become the method of choice in the early 21st century. Mifepristone is highly effective even at doses which are not abortifacient. Efficacy cannot be calculated precisely, but all presently available methods seem to prevent at least 74% of unwanted pregnancies. The Yuzpe regimen inhibits or delays ovulation, but there is no good evidence that it prevents implantation. There are no data on the mechanism of action of levonorgestrel alone and the mode of action of mifepristone depends on when in the reproductive cycle it is used. Accessibility to emergency contraception is limited by the requirement for it to be prescribed by a doctor. Advanced provision of emergency contraception may prevent a significant number of unwanted pregnancies. PMID- 11255558 TI - Novel delivery systems in contraception. AB - Contraception has mainly remained the responsibility of women. The sexually active time during the fertile period of life may last over 30 years and, increasingly, it is more than 10 years before the first baby. It is, therefore, natural that convenient long-acting contraceptive methods are becoming more and more appealing. The discovery of poly(dimethylsiloxane) as a carrier, and controlled release polymers for small molecule drugs allowed the development of contraceptive devices releasing steroids for several years. While contraceptive implants and intra-uterine systems are already marketed in many countries, contraceptive vaginal rings are in their late development phase. The key features of these long acting delivery systems are convenience, efficacy, reversibility and positive long-term health effects. Since these methods are based on new concepts, the provider needs to be prepared for extensive counselling. PMID- 11255559 TI - Health consequences of combined oral contraceptives. AB - During the past 40 years, combined oral contraceptives (COCs) have become a key component of modern fertility regulation programmes. Today, an estimated 100 million women throughout the world use this method of contraception. With such wide-spread usage, it is perhaps not surprising that COCs have been the subject of extensive medical research. PMID- 11255560 TI - Progress towards the development of non-peptide orally-active gonadotropin releasing hormone (GnRH) antagonists: therapeutic implications. AB - Gonadotropin-releasing hormone (GnRH) is a decapeptide (pGlu-His-Trp-Ser-Tyr-Gly Leu-Arg-Pro-Gly.NH2) which is produced from a precursor polypeptide in hypothalamic neurons and secreted in a pulsatile fashion to stimulate the secretion of LH and FSH via its interaction with a cognate receptor on gonadotropes. Low doses of the native peptide delivered in a pulsatile manner to mimic that found in the hypothalamic portal vessels restore fertility in hypogonadal patients, and are also effective in treating cryptorchidism and delayed puberty. Administration of high doses of GnRH, or agonist analogues, causes desensitization of the gonadotrope with consequent decline in gonadal gametogenesis and steroid and peptide hormone synthesis. This phenomenon finds extensive therapeutic application in clinical medicine in a wide spectrum of disease (Table 1). In addition, GnRH analogues have promise as new generation male and female contraceptives in conjunction with steroid hormone replacement. GnRH antagonists inhibit the reproductive system through competition with endogenous GnRH for the receptor and, in view of their rapid effects, are being increasingly used for the above mentioned applications. The peptide agonists and antagonists currently available require parenteral administration, typically in the form of long-acting depots. A new generation of non-peptide GnRH antagonists are beginning to emerge which should allow oral administration and, therefore, may provide greater flexibility of dosing, lower costs and increased patient acceptance. PMID- 11255561 TI - Novel agents to modulate oestrogen action. AB - As women enter the menopause, the majority suffers symptoms associated with a dramatic fall in circulating levels of 17 beta-oestradiol and oestrone. As a result, the oestrogen protective effect against coronary artery disease and osteoporosis is lost. To solve these problems, hormone replacement therapy is often used. However, there are a number of side-effects including increased risk from breast and uterine cancer that can limit compliance. New drugs, called selective oestrogen modulators (SERMs), have been developed to mimic oestrogen's effects on the liver, heart and bones but without its harmful effects on the breast and uterus. SERMs are structurally diverse compounds that bind to oestrogen receptors and elicit agonist or antagonist responses depending on the target tissue and hormonal milieu. The drugs are being used, or evaluated, for the prevention of hormone-responsive breast cancer, osteoporosis and cardiovascular disease in postmenopausal women. Tamoxifen is the endocrine treatment of choice for breast cancer, but it also has beneficial effects on bone density and serum lipids in postmenopausal women. Recently, tamoxifen was shown to decrease the risk of invasive breast cancer in women at high risk. However, tamoxifen has some stimulatory effects on the endometrium. Raloxifene is used to prevent osteoporosis and fractures. Raloxifene also lowers circulating cholesterol and the incidence of invasive breast cancer in postmenopausal women but does not stimulate the endometrium. The SERMs have evolved from mere laboratory curiosities into drugs that hold promise for preventing several major diseases associated with ageing in women. PMID- 11255562 TI - Angiogenesis and its control in the female reproductive system. AB - The rapid, controlled and cyclical nature of angiogenesis in the female reproductive tract suggests that interference with this process should provide a novel approach to manipulation of reproductive function. Many factors involved in the regulation of angiogenesis have been identified, and the possibility of stimulating or inhibiting these paracrine control mechanisms is being addressed using current advances in the development of angiogenic and anti-angiogenic compounds. Studies with animal models indicate that the normal processes of folliculogenesis, ovulation and corpus luteum function in the ovary, and the control of menstruation and implantation in the endometrium could be profoundly influenced by manipulation of angiogenesis. Novel therapeutic agents targeted to the angiogenic pathway may also have a wide range of applications in pathological processes in the reproductive tract such as cancer, endometriosis, fibroid growth, and ovarian hyperstimulation syndrome. PMID- 11255563 TI - Treatment of male sexual dysfunction. AB - Male sexual dysfunction is a prevalent condition in the population, is a major health problem and has previously been both under diagnosed and under treated. There are now a number of treatments available that are safe and easy to use which provide an effective solution for most presenting patients. Oral drugs have recently become the first-line option for many men with about 60-70% of new presentations achieving success. Those who fail a trial of oral treatments have a number of other options available, which are able to provide erections sufficient for intercourse in many of the oral drug failures. All these options, their indications, side-effects and complications are outlined in this chapter. PMID- 11255564 TI - Consequences of long-term hormone replacement therapy. AB - The use of oestrogens in the longer term is an area of considerable current scientific and clinical debate. The extra-reproductive range of oestrogen actions is broad, with these steroid hormones and their receptors (ERs) being intimately involved in the normal function of, inter alia, the adult female skeleton, the cardiovascular system and the brain. Desirable as the restoration of normal circulating oestrogen may be after menopause, HRT use is compromised by the engagement of the reproductive sites of breast and uterus. This may cause concern to patient and physician alike due to the consequent imposition of cyclical bleeding and risk of breast malignancy. In the individual patient, therefore, a balance of risk against benefit has to be struck so that the patient may be precisely advised of the type and duration of oestrogen replacement which may be indicated in her own case. The advent of selective oestrogen receptor modulation with its ability to delete adverse effects in breast and endometrium, is a substantial pharmacological and clinical advance. PMID- 11255565 TI - Male germ cell transplantation: an experimental approach with a clinical perspective. AB - Germ cell transplantation was developed in strains of mice. The infusion of germ cell preparations into the seminiferous tubules of infertile hosts led to repopulation of the testis with donor germ cells and restored fertility. Meanwhile, this technique has become a powerful tool to study the expansion of the testicular stem cell population and the kinetics of spermatogonial proliferation and re-induction of spermatogenesis. Further approaches, such as the transfer of rat/hamster/rabbit germ cells into mouse testes or cryopreservation or culture of spermatogonia, have widened the spectrum of applications associated with germ cell transplantation. Since the devastating effects of chemo- or radiotherapy on spermatogonia are the cause of infertility in oncological patients, germ cell transplantation might become an alternative approach for gonadal protection in prepubertal and postpubertal male tumour patients. This review focusses on recent developments with respect to germ cell transplantation and highlights the problems and perspectives of future clinical applications. PMID- 11255566 TI - Sampling properties of the spectrum and coherency of sequences of action potentials. AB - The spectrum and coherency are useful quantities for characterizing the temporal correlations and functional relations within and between point processes. This article begins with a review of these quantities, their interpretation, and how they may be estimated. A discussion of how to assess the statistical significance of features in these measures is included. In addition, new work is presented that builds on the framework established in the review section. This work investigates how the estimates and their error bars are modified by finite sample sizes. Finite sample corrections are derived based on a doubly stochastic inhomogeneous Poisson process model in which the rate functions are drawn from a low-variance gaussian process. It is found that in contrast to continuous processes, the variance of the estimators cannot be reduced by smoothing beyond a scale set by the number of point events in the interval. Alternatively, the degrees of freedom of the estimators can be thought of as bounded from above by the expected number of point events in the interval. Further new work describing and illustrating a method for detecting the presence of a line in a point process spectrum is also presented, corresponding to the detection of a periodic modulation of the underlying rate. This work demonstrates that a known statistical test, applicable to continuous processes, applies with little modification to point process spectra and is of utility in studying a point process driven by a continuous stimulus. Although the material discussed is of general applicability to point processes, attention will be confined to sequences of neuronal action potentials (spike trains), the motivation for this work. PMID- 11255567 TI - A novel spike distance. AB - The discrimination between two spike trains is a fundamental problem for both experimentalists and the nervous system itself. We introduce a measure for the distance between two spike trains. The distance has a time constant as a parameter. Depending on this parameter, the distance interpolates between a coincidence detector and a rate difference counter. The dependence of the distance on noise is studied with an integrate-and-fire model. For an intermediate range of the time constants, the distance depends linearly on the noise. This property can be used to determine the intrinsic noise of a neuron. PMID- 11255568 TI - On synchrony of weakly coupled neurons at low firing rate. AB - The dynamics of a pair of weakly interacting conductance-based neurons, firing at low frequency, nu, is investigated in the framework of the phase-reduction method. The stability of the antiphase and the in-phase locked state is studied. It is found that for a large class of conductance-based models, the antiphase state is stable (resp., unstable) for excitatory (resp., inhibitory) interactions if the synaptic time constant is above a critical value tau(c)(s), which scales as the absolute value of log nu when nu goes to zero. PMID- 11255570 TI - Metabolically efficient information processing. AB - Energy-efficient information transmission may be relevant to biological sensory signal processing as well as to low-power electronic devices. We explore its consequences in two different regimes. In an "immediate" regime, we argue that the information rate should be maximized subject to a power constraint, and in an "exploratory" regime, the transmission rate per power cost should be maximized. In the absence of noise, discrete inputs are optimally encoded into Boltzmann distributed output symbols. In the exploratory regime, the partition function of this distribution is numerically equal to 1. The structure of the optimal code is strongly affected by noise in the transmission channel. The Arimoto-Blahut algorithm, generalized for cost constraints, can be used to derive and interpret the distribution of symbols for optimal energy-efficient coding in the presence of noise. We outline the possibilities and problems in extending our results to information coding and transmission in neurobiological systems. PMID- 11255569 TI - Population coding with correlation and an unfaithful model. AB - This study investigates a population decoding paradigm in which the maximum likelihood inference is based on an unfaithful decoding model (UMLI). This is usually the case for neural population decoding because the encoding process of the brain is not exactly known or because a simplified decoding model is preferred for saving computational cost. We consider an unfaithful decoding model that neglects the pair-wise correlation between neuronal activities and prove that UMLI is asymptotically efficient when the neuronal correlation is uniform or of limited range. The performance of UMLI is compared with that of the maximum likelihood inference based on the faithful model and that of the center-of-mass decoding method. It turns out that UMLI has advantages of decreasing the computational complexity remarkably and maintaining high-level decoding accuracy. Moreover, it can be implemented by a biologically feasible recurrent network (Pouget, Zhang, Deneve, & Latham, 1998). The effect of correlation on the decoding accuracy is also discussed. PMID- 11255571 TI - Effective neuronal learning with ineffective Hebbian learning rules. AB - In this article we revisit the classical neuroscience paradigm of Hebbian learning. We find that it is difficult to achieve effective associative memory storage by Hebbian synaptic learning, since it requires network-level information at the synaptic level or sparse coding level. Effective learning can yet be achieved even with nonsparse patterns by a neuronal process that maintains a zero sum of the incoming synaptic efficacies. This weight correction improves the memory capacity of associative networks from an essentially bounded one to a memory capacity that scales linearly with network size. It also enables the effective storage of patterns with multiple levels of activity within a single network. Such neuronal weight correction can be successfully carried out by activity-dependent homeostasis of the neuron's synaptic efficacies, which was recently observed in cortical tissue. Thus, our findings suggest that associative learning by Hebbian synaptic learning should be accompanied by continuous remodeling of neuronally driven regulatory processes in the brain. PMID- 11255573 TI - Blind source separation by sparse decomposition in a signal dictionary. AB - The blind source separation problem is to extract the underlying source signals from a set of linear mixtures, where the mixing matrix is unknown. This situation is common in acoustics, radio, medical signal and image processing, hyperspectral imaging, and other areas. We suggest a two-stage separation process: a priori selection of a possibly overcomplete signal dictionary (for instance, a wavelet frame or a learned dictionary) in which the sources are assumed to be sparsely representable, followed by unmixing the sources by exploiting the their sparse representability. We consider the general case of more sources than mixtures, but also derive a more efficient algorithm in the case of a nonovercomplete dictionary and an equal numbers of sources and mixtures. Experiments with artificial signals and musical sounds demonstrate significantly better separation than other known techniques. PMID- 11255572 TI - Temporal difference model reproduces anticipatory neural activity. AB - Anticipatory neural activity preceding behaviorally important events has been reported in cortex, striatum, and midbrain dopamine neurons. Whereas dopamine neurons are phasically activated by reward-predictive stimuli, anticipatory activity of cortical and striatal neurons is increased during delay periods before important events. Characteristics of dopamine neuron activity resemble those of the prediction error signal of the temporal difference (TD) model of Pavlovian learning (Sutton & Barto, 1990). This study demonstrates that the prediction signal of the TD model reproduces characteristics of cortical and striatal anticipatory neural activity. This finding suggests that tonic anticipatory activities may reflect prediction signals that are involved in the processing of dopamine neuron activity. PMID- 11255574 TI - Complexity pursuit: separating interesting components from time series. AB - A generalization of projection pursuit for time series, that is, signals with time structure, is introduced. The goal is to find projections of time series that have interesting structure, defined using criteria related to Kolmogoroff complexity or coding length. Interesting signals are those that can be coded with a short code length. We derive a simple approximation of coding length that takes into account both the nongaussianity and the autocorrelations of the time series. Also, we derive a simple algorithm for its approximative optimization. The resulting method is closely related to blind separation of nongaussian, time dependent source signals. PMID- 11255575 TI - Algebraic analysis for nonidentifiable learning machines. AB - This article clarifies the relation between the learning curve and the algebraic geometrical structure of an unidentifiable learning machine such as a multilayer neural network whose true parameter set is an analytic set with singular points. By using a concept in algebraic analysis, we rigorously prove that the Bayesian stochastic complexity or the free energy is asymptotically equal to lambda(1) log n - (m(1) - 1) log log n + constant, where n is the number of training samples and lambda(1) and m(1) are the rational number and the natural number, which are determined as the birational invariant values of the singularities in the parameter space. Also we show an algorithm to calculate lambda(1) and m(1) based on the resolution of singularities in algebraic geometry. In regular statistical models, 2lambda(1) is equal to the number of parameters and m(1) = 1, whereas in nonregular models, such as multilayer networks, 2lambda(1) is not larger than the number of parameters and m(1) > or = 1. Since the increase of the stochastic complexity is equal to the learning curve or the generalization error, the nonidentifiable learning machines are better models than the regular ones if Bayesian ensemble learning is applied. PMID- 11255576 TI - A new on-line learning model. AB - We introduce a new supervised learning model that is a nonhomogeneous Markov process and investigate its properties. We are interested in conditions that ensure that the process converges to a "correct state," which means that the system agrees with the teacher on every "question." We prove a sufficient condition for almost sure convergence to a correct state and give several applications to the convergence theorem. In particular, we prove several convergence results for well-known learning rules in neural networks. PMID- 11255577 TI - Why does research have so little impact on American drug policy? PMID- 11255578 TI - Drug wars in the post-Gutenberg galaxy: mass media as the next battleground. PMID- 11255579 TI - Conversation with Gabriel Romanus. PMID- 11255581 TI - Risk factors for HIV-1 seropositivity in drug users under 30 years old in Haiphong, Vietnam. AB - AIMS: To assess the prevalence of HIV infection among young drug users in Haiphong and, secondarily, to document the current patterns of drug use and sexual behavior in them. Design, setting and participants. A cross-sectional survey of drug users 15-30 years old, identified from police lists and by the snowballing method between March 15 and May 30, 1999, was conducted in Haiphong City. MEASUREMENTS: Subjects (n = 520: 514 males and six females) were interviewed and donated blood specimens for HIV-1 antibody testing. FINDINGS: Mean age of the subjects was 25 years (range 15-30 years). The prevalence of HIV among injecting drug users (IDUs) was 74% and among drug users not reporting injecting was 46% (92/201). Sixty-one per cent (319) reported injecting drugs. Among injectors, 72% used heroin, and 68% had shared needles. Factors related to the presence of HIV antibody among IDUs were sharing needles (OR: 4.12) and injecting more than 31 times per month (OR: 2.37). Extramarital sex within the last 6 months was reported by 44% of single and 24% of married IDUs. CONCLUSION: The high HIV-1 prevalence in drug users and their frequent sexual mixing with the non-drug-using population suggests that preventive interventions for reduction of high-risk drug taking and sexual activities are urgently needed in these populations. Interventions through public sexually transmitted infection (STI) clinics are unlikely to have much impact, as only 16% of IDUs with an STI attended a public STI clinic. PMID- 11255580 TI - Sleep-disordered breathing in stable methadone programme patients: a pilot study. AB - AIMS: To explore the possibility that stable MMP patients have sleep-disordered breathing (SDB) and abnormal sleep architecture defined by nocturnal sleep stages and sleep efficiency. DESIGN: Observational. SETTING: Regional Methadone Service and sleep disorders laboratory in a university affiliated hospital. Participants and measurements. Ten stable MMP patients and nine normal subjects were assessed clinically and with overnight polysomnography. FINDINGS: There were no differences in age, sex and body mass index between the groups. The methadone dose ranged between 50 and 120 mg/day. Six patients had central apnoea index (CAI) greater than 5, four had a CAI greater than 10 and three of these exhibited periodic breathing. No normal subject had central sleep apnoea. The patients had lower sleep efficiency (p < 0.05), less slow wave sleep (p < 0.01), less rapid eye movement sleep (p < 0.05) and more Stage 2 sleep (p < 0.05) than controls. CONCLUSIONS: Stable MMP patients have more sleep architecture abnormalities than controls and a higher prevalence of central sleep apnoea. Further studies are needed to confirm these findings, to delineate the mechanisms for the abnormalities and to assess whether the SDB is related to sudden death in stable MMP patients. We recommend that MMP patients have awake and sleep respiration assessed to identify those potentially at risk. PMID- 11255582 TI - Conduct problems and early cannabis initiation: a longitudinal study of gender differences. AB - AIM: To investigate the relationship between early conduct problems and early onset of cannabis use, with special emphasis on possible gender differences. DESIGN: A prospective longitudinal study of a national sample of 2436 adolescents. The sample was followed up over a year and a half, when the adolescents were in their early teens. SETTING: Norway. MEASUREMENTS: On the basis of an earlier study, conduct problems (CP) closely related to the criteria for conduct disorder (CD) in DSM-III-R were decomposed into three dimensions, labelled serious, aggressive and covert. Further, information was collected about alcohol intoxication, daily smoking and use of cannabis. A number of questions were posed about sexual interactions and perceived puberty development. Parental socio-economic status was measured according to the ISCO-88. Separate information was collected as to whether the parents were on social welfare or unemployed. A parental bonding measure (PBI) was used to measure the emotional relationship between respondents and parents. Further, a measure of parental monitoring was used, and information was also collected on other aspects of the family milieu, and on the adolescents' peers. Statistical models. Logistic regression analysis was employed. As the sample consisted of pupils clustered within classes within schools, a three-level error structure for the logistic regression model was estimated. FINDINGS: There was a strong association between early conduct problems and subsequent cannabis initiation. Also conduct problems at a potential subclinical level seemed to have great impact. The effect was significantly stronger in girls than in boys. Serious CP was found to have a moderate effect upon cannabis initiation in boys, whereas aggressive and covert CP had strong effects in girls. Early onset of puberty and early sexual involvement had no impact, whereas early use of cigarettes proved an important precursor to cannabis use. CONCLUSIONS: Conduct problems are important precursors of early onset cannabis use, but probably represent gender-specific aetiologies. There might be an important potential for prevention of early onset drug use in the prevention of early conduct problems, in particular for girls. PMID- 11255583 TI - Back-projection estimates of the number of dependent heroin users in Australia. AB - AIMS: To plan an appropriate response to heroin use in Australia, good estimates are needed of the numbers of dependent heroin users, the group who are most in need of treatment, most at risk of fatal opioid overdose and most at risk of contracting and transmitting blood-borne viruses. METHODS: Back-projection methods were used to estimate the numbers of people starting dependent heroin injecting in Australia between 1960 and 1997. Separate analyses were based on national opioid overdose deaths and numbers of new entrants to methadone treatment in New South Wales (NSW). Estimates of the rates at which dependent heroin users cease heroin use, commence methadone treatment or die from opioid overdoses were estimated from external data sources. RESULTS: Back-projection estimates derived from opioid overdose deaths indicated that there were 104 000 (lower limit of 72 000 and upper limit of 157 000) people who were heroin dependent in Australia between 1960 and 1997. Of these it was estimated that 67 000 (39 000-120 000) were still heroin dependent at the end of 1997. Back projection estimates based on numbers of new entrants to methadone treatment in NSW indicated that there were 108 000 (82 000-141 000) heroin-dependent people in Australia between 1960 and 1997, of whom 71 000 (47 000-109 000) were estimated to be heroin dependent at the end of 1997. Both analyses indicated that the number of heroin-dependent people in Australia has increased substantially from the early 1970s onwards. CONCLUSIONS: Back-projection estimates based on analyses of treatment entries and opioid overdose deaths provide an additional method for estimating the numbers of heroin-dependent people in the population. The addition of these methods to existing methods, using different data sources and statistical methods, should improve consensus estimates of the numbers of heroin dependent people. PMID- 11255584 TI - Social dimensions of adolescent substance use. AB - OBJECTIVES: The aim of this study was to explore in detail the relationship between various social aspects of young people's lives and substance use and differences in the degree of influence exerted by the different social factors as a function of age. Design, setting, participants. The study was a survey of pupils aged 11-16 in a stratified sample of five English schools. Data from 4516 participants were obtained in relation to their cigarette, alcohol and illicit drug use and their contact with the police, perceived academic achievements and future expectations, religious beliefs, family structure, the importance of family versus peer opinions and suspension from school. MEASURES: Cumulative, age specific preferences of substance misuse were compared. Logistic regression was used to rank the various risk factors. RESULTS: Substantial differences were found between substance users and non-users and the various risk factors being examined. For example, of those who had only been in trouble with the police, 18.8% used illegal drugs compared with 1.6% of those who had not had a police contact and who had no other risk factors. Many of these relationships were age sensitive. For instance, the negative relationship between belief in God and illicit drug use became stronger as age increased (non-believers: y = 8.1886x - 9.16 R(2) = 0.9484; believers: y = 5.1514x - 8.08 R(2) = 0.9247). These results suggest that, within this sample of English adolescents, there was a strong relationship between substance use and the social factors examined. Although there were differences depending upon whether cigarette, alcohol or illicit drug use was being modelled, logistic regression indicated that the social factors could be ranked in the following order of importance: concurrent use of the second and third substances, having been in trouble with the police, perceived poor academic performance and low future academic expectations, a lack of religious belief, coming from a non-intact family, favouring peer over family opinion and having been suspended from school. Many of these relationships were age-sensitive with substance use peaking at age 15. CONCLUSION: The models and relationships presented in this paper show that a constellation of behaviours are related to adolescent substance use. Also demonstrated is that behaviours cannot be considered in isolation, but need to be examined from an holistic or biopsychosocial standpoint. These relationships are complex and future research should consider not only causality of adolescent substance use, but also of the aetiology of the satellite behaviours. PMID- 11255585 TI - Measuring alcohol consumption and alcohol-related problems: comparison of responses from self-administered questionnaires and telephone interviews. AB - AIMS: Compared with surveys using self-administered questionnaires, telephone interviews generally yield higher coverage rates, have a lower proportion of missing values and result in fewer inconsistencies. Meta-analyses, however, show that responses to sensitive questions by telephone tend to be biased by social expectations. The aim of the study is to examine whether responses on alcohol consumption and alcohol-related problems differ with respect to mode of administration (self-administered vs. telephone). Design and participants. Data were analysed from the 1995 self-administered survey among 6427 subjects and from telephone surveys conducted annually between 1994 and 1996 yielding a pooled sample of 6193 subjects. MEASUREMENTS: Alcohol consumption within the last 30 days was measured using a beverage-specific quantity-frequency index. For a summary measure responses were converted into pure alcohol (ethanol) per day and categorized into no alcohol consumption (0 g), non-hazardous consumption (< or = 20 g for female and < or = 40 g for males) and hazardous consumption (> 20 g for females and > 40 g for males). Alcohol-related problems were assessed using the CAGE questionnaire with a cut-off point of at least two positive responses. FINDINGS: Using (cumulative) logistic regression, a significant mode effect was found for both alcohol consumption and alcohol-related problems. Lower beverage specific prevalences in the telephone mode were found to be responsible for the difference in the distribution of the summary consumption measure. CONCLUSIONS: Results indicate that patterns of drinking and alcohol-related problems are more easily reported in self-administration questionnaires compared to telephone interviews. PMID- 11255586 TI - Substance use and driving: the coexistence of risky and safe behaviors. AB - AIMS: Two risky behaviors (driving after drinking/getting drunk, riding with drinking drivers) and two safe behaviors (deciding not to drive under the influence of alcohol (DUI), preventing someone else from DUI) were examined in relation to use frequency and friends' DUI to determine whether individuals tend to engage in both types of behaviors. DESIGN: Self-report questionnaires were administered to a random sample of 1233 young adults in New Jersey (USA) on two occasions (mean age 21 and mean age 28). Structural equation modeling was used to assess the goodness of fit of a hypothesized model of cross-sectional and longitudinal relationships. FINDINGS: Relationships between the four behaviors were strongly positive for men and women at both occasions and were substantially accounted for by use frequency and friends' DUI. At the later age, however, it was necessary to add non-recursive pathways to the model, which were different for men and women. CONCLUSIONS: Findings suggest that (1) riding with drinking drivers plays an important role in the maintenance of the other behaviors and (2) most individuals vacillate between risky and safe behaviors indicating that drinking contexts are best viewed as risky decision-making situations requiring individuals to choose between riskier and safer courses of action. PMID- 11255587 TI - Randomized controlled trial of a midwife-delivered brief smoking cessation intervention in pregnancy. AB - OBJECTIVE: To evaluate the efficacy of a brief smoking cessation intervention with pregnant women practicable routinely by midwives. DESIGN: Midwives were randomized to deliver the experimental intervention or usual care. The 10-15 minute intervention was based on brief counselling, written materials, arrangements for continuing self-help support and feedback on expired-air carbon monoxide levels. The intervention was tailored to the women's needs: those who did not want to stop smoking received a brief motivational intervention, those who wanted to stop received an intervention designed to assist them and those that had stopped recently (recent ex-smokers) received a relapse prevention intervention. SETTING: Booking interviews with pregnant women in nine hospital and community trusts. SUBJECTS: A total of 1120 pregnant women in the third month of pregnancy (249 recent ex-smokers and 871 current smokers). MAIN OUTCOME MEASURES: Three indicators of biochemically validated abstinence were collected. Continuous abstinence for at least 3 months prior to delivery, point prevalence abstinence immediately post-delivery, and continuous abstinence from 3 months pre delivery to 6 months post-delivery. RESULTS: Only a small proportion of the women who would have been eligible to take part in the trial were actually recruited by 178 recruiting midwives, with lack of time being cited as the main barrier. The intervention and usual care groups differed in post-delivery point prevalence abstinence rates for recent ex-smokers (65% vs. 53%, p < 0.05, one-tailed), but not in other outcome measures. Overall, 54% of "recent ex-smokers" at booking and 7% of "current smokers" at booking had been abstinent for at least 3 months at the time of delivery, and 23% and 3%, respectively, were still abstinent by the time the child was 6 months old (i.e. 12 months post-intervention). Smoking status at follow-up was predicted by dependence indexed by time to first cigarette in the morning. CONCLUSIONS: A brief "one-off" smoking cessation intervention by midwives does not seem to be a practicable or effective method of helping pregnant smokers to stop. Other options such as tailored self-help materials and telephone counselling and other specialist treatments should be examined. Current smoking cessation rates in pregnancy are very low. PMID- 11255589 TI - Comment on "Shakespeare and the meaning of authorship". PMID- 11255588 TI - Differential uptake of a smoking cessation programme disseminated to doctors and midwives in antenatal clinics. AB - AIMS: Two methods of dissemination (simple and intensive) were used to disseminate a smoking cessation programme to doctors and midwives working in antenatal clinics. This paper describes the differential uptake of the smoking cessation programme by doctors and midwives. It investigates whether the number of smoking cessation interventions used differ due to the type of dissemination. It also examines the frequency with which doctors and midwives provide smoking cessation interventions after dissemination. DESIGN: Clinics were randomized to the method of dissemination (simple or intensive). Pre-post test design was used to examine the relationship between dissemination method and professional status at baseline and follow-up. A baseline survey collected data on the use of smoking cessation intervention in the clinics prior to dissemination. A follow-up survey was conducted 18 months after the dissemination. SETTING: Twenty-three public hospital antenatal clinics in NSW. PARTICIPANTS: All clinical staff (midwives and doctors) working in the clinic during the 1-2-week survey period prior to dissemination and 18 months after the dissemination were asked to participate. The response rate was 63% (223) at baseline and 64% (182) at follow-up. Only 48% of midwives and doctors at follow-up were working in the original clinic. MEASURES: The proportion of clinicians who initially adopted the programme; the proportion of clinicians who had used one or more programme components in the last week); the number of types of smoking cessation intervention provided (maximum = 13), and the estimated proportion of clients offered smoking cessation intervention. FINDINGS: More midwives than doctors "ever used" the programme (76% vs. 25%) and continued to implement (58% vs. 22%) the programme 18 months after dissemination. Both midwives and doctors increased the number of types of smoking cessation intervention offered at follow-up compared to baseline (mean difference 2.8). Midwives provided more smoking cessation interventions than doctors at baseline (mean difference 0.9) and at follow-up (1.6), regardless of method used to disseminate the programme. Midwives' mean estimates of the proportion of clients offered interventions were greater than doctors' (midwives' 59% vs. doctors' 35%) at follow-up. CONCLUSION: The dissemination of a smoking cessation programme increased the level of smoking cessation interventions used by doctors and midwives. Doctors and midwives differ in their uptake of smoking cessation programmes. This information can be used to plan programme dissemination strategies in the future. PMID- 11255590 TI - Role of delta-tubulin and the C-tubule in assembly of Paramecium basal bodies. AB - BACKGROUND: A breakthrough in the understanding of centriole assembly was provided by the characterization of the UNI3 gene in Chlamydomonas. Deletion of this gene, found to encode a novel member of the tubulin superfamily, delta tubulin, results in the loss of the C-tubule, in the nine microtubule triplets which are the hallmark of centrioles and basal bodies. Delta-tubulin homologs have been identified in the genomes of mammals and protozoa, but their phylogenetic relationships are unclear and their function is not yet known. RESULTS: Using the method of gene-specific silencing, we have inactivated the Paramecium delta-tubulin gene, which was recently identified. This inactivation leads to loss of the C-tubule in all basal bodies, without any effect on ciliogenesis. This deficiency does not directly affect basal body duplication, but perturbs the cortical cytoskeleton, progressively leading to mislocalization and loss of basal bodies and to altered cell size and shape. Furthermore, additional loss of B- and even A-tubules at one or more triplet sites are observed: around these incomplete cylinders, the remaining doublets are nevertheless positioned according to the native ninefold symmetry. CONCLUSIONS: The fact that in two distinct phyla, delta-tubulin plays a similar role provides a new basis for interpreting phylogenetic relationships among delta-tubulins. The role of delta-tubulin in C-tubule assembly reveals that tubulins contribute subtle specificities at microtubule nucleation sites. Our observations also demonstrate the existence of a prepattern for the ninefold symmetry of the organelle which is maintained even if less than 9 triplets develop. PMID- 11255591 TI - Issues in contemporary fluid management. AB - Fluid management strategies need to be guided by an understanding of the pathophysiologic mechanisms underlying fluid imbalance. In the hypovolaemic patient, reduced circulating blood volume and venous return and, in severe cases, altered tissue perfusion may initiate a cascade of pathophysiologic processes culminating in multiple organ failure. The objectives of fluid management are to maintain adequate blood pressure, tissue oxygenation and intravascular fluid volume. Both crystalloids and colloids can be useful for these purposes. In the hypovolaemic patient with normal pulmonary function, the use of colloids to maintain colloid osmotic pressure can limit the development of peripheral as well as pulmonary oedema. However, choice of fluid is less important in states of increased lung capillary permeability. Further evidence is needed to broaden understanding of the optimal roles for particular fluid management strategies. Experimental models can make an important contribution in gathering such evidence. Rigorous pharmacoeconomic studies are also needed to define the benefits and costs of differing fluid regimens. PMID- 11255592 TI - Pathophysiology of fluid imbalance. AB - Fluid imbalance can arise due to hypovolemia, normovolemia with maldistribution of fluid, and hypervolemia. Trauma is among the most frequent causes of hypovolemia, with its often profuse attendant blood loss. Another common cause is dehydration, which primarily entails loss of plasma rather than whole blood. The consequences of hypovolemia include reduction in circulating blood volume, lower venous return and, in profound cases, arterial hypotension. Myocardial failure may result from increased myocardial oxygen demand in conjunction with reduced tissue perfusion. Finally, anaerobic metabolism due to reduced perfusion may produce acidosis and, together with myocardial dysfunction, precipitate multi organ failure. The splanchnic organs are particularly susceptible to the deleterious effects of hypotension and hypovolemic shock, and these effects, depending upon their duration and severity, may be irreversible despite restoration of normovolemia by fluid administration. Patient monitoring in the intensive care unit typically relies upon central venous pressure devices, whereas the primary focus in the operating theater is blood volume deficit estimated from suction devices. However, estimates of intraoperative blood loss can be inaccurate, potentially leading to inappropriate fluid management. Normovolemia with maldistribution of fluid can be encountered in shock-specific microcirculatory disorders secondary to hypovolemia, as well as pain and stress. Consequent vasoconstriction and reduced tissue driving pressure, as well as leukocyte and platelet adhesion, and liberation of humoral and cellular mediators, may impair or abolish blood flow in certain areas. The localized perfusion deficit may contribute to multi-organ failure. Choice of resuscitation fluid may be important in this context, since some evidence suggests that at least certain colloids might be helpful in diminishing post-ischemic microvascular leukocyte adherence. Excessive volume administration may lead to fluid overload and associated impairment of pulmonary function. However, entry of fluid into the lungs may also be facilitated by increased vascular permeability in certain pathologic conditions, especially sepsis and endotoxemia, even in the absence of substantially rising hydrostatic pressure. Another condition associated with elevated vascular permeability is systemic capillary leak syndrome. The chief goal of fluid management, based upon current understanding of the pathophysiology of fluid imbalance, should be to ensure adequate oxygen delivery by optimizing blood oxygenation, perfusion pressure, and circulating volume. PMID- 11255593 TI - An alternative pathway for preclinical research in fluid management. AB - Recent meta-analyses have created uncertainties regarding the appropriate clinical role of colloid resuscitation fluids in critically ill patients and prompted changes in fluid management practice. Such changes may not be justified in view of methodological limitations inherent in the meta-analyses. Further research is nevertheless needed to resolve the questions raised concerning the relationship between choice of resuscitation fluid and patient outcome. Animal studies can play an important part by reliably indicating whether particular fluids are likely to prove effective and safe in clinical trials. It is important to avoid costly large-scale clinical trials that fail to demonstrate the clinical utility of the tested therapy, as resources expended in failed trials raise overall development costs and thereby restrict the range of therapies meeting criteria of commercial feasibility. Promising therapies may thus not be pursued, even though an urgent clinical need may exist. An alternative pathway of preclinical research may be of value in avoiding some of the major clinical trial failures of recent years, particularly in the area of sepsis. This alternative pathway commences with the formulation of hypotheses by therapeutics developers. Independent preclinical investigators are challenged, by means of a competitive request for proposals, to test the hypotheses in rigorous randomized studies employing clinically relevant animal models. Promising proposals would then be selected for further development with the aid of peer review. The results of the randomized animal studies, along with other preclinical data, could also be evaluated using accepted principles of 'critical appraisal' commonly applied to clinical trial results. This critical appraisal might, where appropriate, include meta-analysis of animal study findings. This alternative preclinical pathway to new product evaluation should be completed before the commencement of large-scale clinical trials. PMID- 11255594 TI - Albumin and artificial colloids in fluid management: where does the clinical evidence of their utility stand? AB - Key questions remain unresolved regarding the advantages and limitations of colloids for fluid resuscitation despite extensive investigation. Elucidation of these questions has been slowed, in part, by uncertainty as to the optimal endpoints that should be monitored in assessing patient response to administered fluid. Colloids and crystalloids do not appear to differ notably in their effects on preload recruitable stroke volume or oxygen delivery. Limited evidence nevertheless suggests that colloids might promote greater oxygen consumption than crystalloids. It remains unclear, in any case, to what extent such physiological parameters might be related to clinically relevant outcomes such as morbidity and mortality. Recent randomized controlled trial results indicate that, at least in certain forms of fluid imbalance, albumin is effective in significantly reducing morbidity and mortality. Much further investigation is needed, however, to determine the effects of colloid administration on clinically relevant outcomes in a broad range of critically ill patients. The ability of administered colloids to increase colloid osmotic pressure (COP) constitutes one mechanism by which colloids might reduce interstitial oedema and promote favourable patient outcomes. However, the applicability of this mechanism may be limited, due to the operation of compensatory mechanisms such as increased lymphatic drainage. Attempts to increase COP might also be less useful in states of increased vascular permeability such as acute respiratory distress syndrome, although this issue has by no means been settled by empirical data. Colloids are clearly more efficient than crystalloids in attaining resuscitation endpoints as judged by the need for administration of far smaller fluid volumes. Among the colloids, albumin offers several advantages compared with artificial colloids, including less restrictive dose limitations, lower risk of impaired haemostasis, absence of tissue deposition leading to severe prolonged pruritus, reduced incidence of anaphylactoid reactions, and ease of monitoring to prevent fluid overload. The cost of albumin, nevertheless, limits its usage. Crystalloids currently serve as the first-line fluids in hypovolaemic patients. Colloids can be considered in patients with severe or acute shock or hypovolaemia resulting from sudden plasma loss. Colloids may be combined with crystalloids to obviate administration of large crystalloid volumes. Further clinical trials are needed to define the optimal role for colloids in critically ill patients. PMID- 11255595 TI - Fluid balance and colloid osmotic pressure in acute respiratory failure: emerging clinical evidence. AB - Available evidence suggests that both hydrostatic and osmotic forces are important in the development of acute respiratory distress syndrome (ARDS) or, more broadly, acute lung injury (ALI). More than 80% of ARDS patients in a large scale randomized controlled trial (RCT) exhibited, at least intermittently, pulmonary artery wedge pressures (PAWP) above 18 mmHg. Retrospective analyses have shown that PAWP elevation is associated with increased mortality. Reduction in serum total protein (STP) has been shown, in a recent retrospective analysis of data from a sepsis patient population with a high frequency of ARDS, to be highly predictive of positive fluid balance, weight gain, development of ARDS, prolonged mechanical ventilation, and mortality. These findings suggest that therapy with diuretics and colloids might be of benefit in the prevention or treatment of ALI. A prospective RCT was designed and conducted to evaluate combination therapy with furosemide and albumin over a 5-day period in 37 ALI patients. Both mean serum albumin and mean STP increased promptly and substantially in furosemide + albumin recipients. The furosemide + albumin group also achieved a mean weight loss of 10 kg by the end of the treatment phase, and their weight loss exceeded that of placebo patients throughout. Hemodynamics improved in the treatment group during the 5-day protocol. Oxygenation, as assessed by the ratio between the fraction of inspired oxygen and the partial pressure of oxygen in arterial blood (PaO2/FiO2), was significantly higher within 24 h after commencement of treatment in the furosemide + albumin than the placebo group. No clinically important adverse effects of furosemide + albumin therapy were encountered. These results provide evidence that combined therapy with furosemide and albumin is effective in augmenting serum albumin and STP levels, promoting weight loss, and improving oxygenation and longer-term hemodynamic stability. Although mortality did not differ between groups, the RCT showed a trend toward reduced duration of mechanical ventilation and length of stay in the intensive care unit in patients receiving furosemide + albumin. The findings of the RCT further highlight the importance of both hydrostatic and osmotic forces in hypoxemic respiratory failure, a subject that requires further investigation. PMID- 11255598 TI - EDITORIAL. PMID- 11255596 TI - The appropriate role of colloids in managing fluid imbalance: a critical review of recent meta-analytic findings. AB - Three meta-analyses have recently been reported on the relationship between choice of resuscitation fluid and risk of mortality in critically ill patients. The relative risk of death (1.16-1.19) in two of the meta-analyses was slightly higher in colloid than crystalloid recipients; however, this observation was not statistically significant. In the third meta-analysis, 6% (95% confidence interval [CI], 3-9%) pooled excess mortality was documented in patients receiving albumin for hypovolaemia, burns or hypoalbuminaemia. The mortality difference in hypovolaemia patients (4%; 95% CI, 0-8%) was not statistically significant. A variety of serious limitations apply to the three meta-analyses, suggesting that their findings be interpreted cautiously. More than one-half of the randomized controlled trials (RCTs) included in the meta-analyses were reported prior to 1990 and hence do not reflect current practice. Each meta-analysis included only a subset of relevant RCTs, and therefore the scope of inferences to be drawn from the meta-analytic results is limited. The meta-analyses combined RCTs that were notably heterogeneous with respect to patient characteristics, type of illness, administered fluids and physiologic endpoints. Differences in illness severity, concomitant therapies and fluid management approaches were not taken into account. Very few of the RCTs were blinded. The meta-analyses do not support the conclusion that choice of resuscitation fluid is a major determinant of mortality in critically ill patients, nor do they support changes to current fluid management practice. Changes such as exclusive reliance on crystalloids would necessitate a reassessment of the goals and methods of fluid therapy. Since the effect on mortality may be minimal or non-existent, choice of resuscitation fluid should rest on whether the particular fluid permits the intensive care unit to provide better patient care. PMID- 11255600 TI - Challenges facing continuing medical education and the Saudi Council for Health Specialities. AB - As we leave the 20th century, continuing medical education faces many challenges in relation to its effectiveness, efficiency and quality. The young and promising Saudi Council for Health Specialties produces a document on accreditation of continuing medical education, which indicates its interest in this vital subject. This paper aims to enrich the approach to continuing medical education in Saudi Arabia by reviewing the main relevant challenges reported in literature and suggesting that the Saudi Council for Health Specialities may consider developing and implementing a continuing medical education charter that addresses the needs of all stakeholders and emphasizes high quality and cost-effective provision. PMID- 11255597 TI - Fluid management: the pharmacoeconomic dimension. AB - Cost is a key concern in fluid management. Relatively few data are available that address the comparative total costs of care between different fluid management regimens in particular clinical indications. Relevant costs of fluid-associated morbidity and mortality, including those incurred after intensive care unit or hospital discharge, also need to be considered in evaluating the cost-benefit ratios of administered fluids. Rigorously designed pharmacoeconomic studies are needed to delineate the costs and benefits of various approaches to fluid management. PMID- 11255601 TI - Are Saudi Arabian hospitals prepared for the threat of biological weapons? AB - The use of biological weapons has been recorded repeatedly in history. Until recently, biological terrorism had been little discussed or written about. However, events over the past 12 to 18 months have made it clear that likely perpetrators already envisage every possible scenario. Nations and dissident groups exist that have both the motivation and access to utilize biological weapons. In April 1994, a Russian biological weapons expert presented the conclusions of the Russian experts as to the agents most likely to be used: smallpox, anthrax, and plague. Health care workers in the Kingdom of Saudi Arabia (physicians, nurses, and emergency medical technicians) need to be aware of the seriousness of the threat of biological weapons, and to have an approach for the early identification, triage, and management of biological weapons victims. Clues to the occurrence of a bioterrorism attack include the abrupt onset of a large number of cases of a similar disease or syndrome, the occurrence of diseases with unusual geographic or seasonal distribution, and epidemics of non-endemic diseases. Health care workers must maintain a high index of suspicion, involve the hospital epidemiologist or infectious diseases specialist, identify a clear administrative chain-of-command to minimize confusion, and rely on existing networks such as the hospital disaster-and-safety committee to ensure a multidisciplinary response. Maximum readiness can be achieved by periodic readiness drills. PMID- 11255602 TI - Geographical distribution of biomedical publications from the Gulf Corporation Council countries. AB - OBJECTIVE: It was our purpose to perform a geographical analysis for the number of biomedical and clinical research publications from the six countries of the Gulf Cooperation Council over the past decade (1990-1999). METHODS: Medline was searched with the aid of the Internet provider PubMed. By using the advanced search option, entries were based on the country name for each of the Gulf Cooperation Council countries and the time period considered. RESULTS: The number of Medline-listed biomedical research papers published in the Gulf Cooperation Council countries over the last 10 years totaled 6,960 and increased by 14% over the past decade. The Kingdom of Saudi Arabia followed by Kuwait was by far the most prolific and accounted for 67 and 16% of publications. The research output from the United Arab Emirates and Oman grew steadily over the past decade, while it appeared to plateau for both Bahrain and Qatar. CONCLUSION: Taking into account that Gulf Cooperation Council countries have a relatively short history of research, the data show that the Gulf Cooperation Council countries are very prolific in terms of Medline-indexed biomedical research publications. PMID- 11255603 TI - Seroepidemiological study of Toxoplasma gondii infection in the human population in the Eastern Region. AB - OBJECTIVE: Toxoplasma gondii is an an obligate intracellular protozoan parasite that causes toxoplasmosis. The infection is worldwide, particularly in warm and moist climates. Few studies have been conducted on the prevalence of subclinical or overt disease in Saudi Arabia. No population-based study was conducted or the seroprevalence of toxoplasmosis in humans in Saudi Arabia and this initiated the present study. The present study aimed at studying seroprevalence of Toxoplasma IgG and IgM antibodies in sera from 5 areas in the Eastern Region of Saudi Arabia. METHODS: A population based epidemiological approach, prevalence according to lifestyle (urban or rural), gender (male or female) occupation and age. RESULTS: Inactive toxoplasmosis (IgG levels) is of rather high prevalence in the human population in the Eastern Region of Saudi Arabia (25%). On the other hand, active toxoplasmosis (acquired during pregnancy) is of rather low prevalence in this study (5%). Active toxoplasmosis (IgM levels) is positively related to the level of exposure, high in farmers and employees in village rural areas and low in children and students in urban areas. CONCLUSION: Patients with active toxoplasmosis are to be treated and made aware of their situation. Hygienic conditions in areas of rather high prevalence of active toxoplasmosis are to be more strictly imposed to minimize transmission of the disease. PMID- 11255604 TI - Epilepsy in Saudi children with cerebral palsy. AB - OBJECTIVE: To study the clinical characteristics, electroencephalographic and computerized axial tomography profile in Saudi children with cerebral palsy who suffer epilepsy in a university referral center. METHODS: A total of 113 patients with cerebral palsy and epilepsy was seen (67 boys, 46 girls) with an over all mean age 5.3 years (range.2-12 years) during the study period (January 1998 - December 1999). They all had clinical evaluation and standardized electroencephalographic studies interpreted by the same examiner. RESULTS: The main clinical features were language delay (61%), hypotonia (45%), hypertonia (38%), and behavioral abnormalities (41%). Seizure types included generalized in 96 (85%), and partial and complex partial with or without secondary generalization in 17 (15%). None of the patients had simple partial seizures. The generalized seizures were non-convulsive in 4 patients (3.5%), tonic/clonic 73 (65%), atonic 3 (3%), myoclonic 16 (14%), and mixed 2 (2%). Inter-ictal electroencephalographic abnormalities were epileptiform activity, generalized in 65 (57.5%) and focal 18 (16%), slow-wave activity in 58 (51%) and hypsarrythmia pattern in 6 (5%). Only 9 patients had normal electroencephalogram. The cranial computerized tomography findings were normal in 11.5%. The main abnormalities were cerebral atrophy (65%), hydrocephalus (8%) and agenesis of the corpus callosum (8%). CONCLUSION: The pattern of seizure type in patients with cerebral palsy and types of electroencephalogram abnormalities electroencephalogram and cranial computerized tomography are comparable to the results from studies in other clinical settings and environments. PMID- 11255605 TI - Corpus callosum agenesis. AB - OBJECTIVE: The objectives are to analyse corpus callosum agenesis in children with various neurological problems in a hospital set-up, and to study the neurological and systemic abnormalities associated with this condition. METHODS: The children with various neurological problems who underwent computerized tomography brain from January 1993 to December 1997, and were found to have corpus callosum agenesis, formed the subjects of this study. These children were examined for any syndromic association, congenital infections or metabolic defects. RESULTS: Out of 2164 children who underwent computerized tomography brain, 22 had corpus callosum agenesis (1%). Most cases were not syndromic and 64% were males. Epileptic disorders were noted in about one third of cases. CONCLUSION: Corpus callosum agenesis is an important anomaly in children with neurodevelopment handicaps, usually detected by neuroradiology. PMID- 11255606 TI - Pitfalls in cerebrospinal fluid test for the diagnosis of neurosyphilis. AB - OBJECTIVE: To determine the usefulness of cerebrospinal fluid tests in the diagnosis of neurosyphilis. METHODS: Two hundred and seven cerebrospinal fluid Venereal Disease Research Laboratories tests were performed at King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia between 1992 and 1997. The records of 14 cases with progressive neurological disease and reactive serum fluorescent treponemal absorbent antibodies or treponemal pallidum hemagglutination test were reviewed for clinical presentation, cerebrospinal fluid analysis and Venereal Disease Research Laboratories, neuro-imaging abnormalities and compatibility with the diagnosis of neurosyphilis. The diagnosis of neurosyphilis was made if the patient had reactive serum fluorescent treponemal absorbent antibodies or treponemal pallidum hemagglutination, history of progressive neurological disease and increased cerebrospinal fluid cells or protein. RESULTS: None of the 207 cerebrospinal fluid-Venereal Disease Research Laboratories tests were reactive. The diagnosis of neurosyphilis was made in 10 out of 14 cases with progressive neurological disease and reactive serum rapid plasma reagin, fluorescent treponemal absorbent antibodies and treponemal pallidum hemagglutination. CONCLUSION: We conclude that if reactive cerebrospinal fluid-Venereal Disease Research Laboratories is required to confirm or diagnose neurosyphilis, most cases will be overlooked. PMID- 11255607 TI - Penile implants in the treatment of organic impotence. AB - OBJECTIVE: To evaluate the reliability and safety of penile implants in the treatment of organic impotence at the Saudi Aramco - Dhahran Health Center. METHODS: A series of 108 cases of organic impotence that underwent 125 penile implantation procedures between 1988 and 1997 was reviewed. The follow-up period ranged between 6 months and 10 years. The mean age was 57.9 years (range 26-76). The prostheses used were AMS (American Medical System) inflatable (92 cases) and malleable (16 cases). RESULTS: There were no complications in 86 patients, (80%) who had functioning prostheses all through the follow-up period. Revision of the implants was required in 13 patients (14%). The causes of revision were severe infection, intolerable pain from an oversized malleable prosthesis, and dysfunction of the inflatable prostheses. Removal of the implant was necessary in severe infection, intolerable pain, and extrusion of the prosthesis. All 9 patients (8%) had inflatable prostheses and refused a second implant. There was no single mortality among our series. The overall procedure complications involved 26 out of 125 procedures (21%). It was shown that malleable penile prostheses have significantly lower procedure complications than the inflatable ones (p<0.05). CONCLUSION: Penile implants are reliable and safe modality of treatment for organic impotence with acceptable morbidity. PMID- 11255608 TI - Pattern of erectile dysfunction in Jeddah city. AB - OBJECTIVE: The aim of this study was to determine the demographic features of erectile dysfunction patients attending different specialized clinics in Jeddah city, and to identify possible risk factors associated with erectile dysfunction problem. METHODS: All newly erectile dysfunction patients (n=388) who attended 6 andrology and urology clinics within a period of 3 months were subjected to a modified structural interview questionnaire to collect demographic data and risk factors for erectile dysfunction. RESULTS: The study revealed the following results among erectile dysfunction patients; Saudi patients constituted (81%). The age ranged from 20-86 years with mean age of 43.23+12.56 years, 73% were married with one wife, 23.5% married with two wives, and 8% were single. About one-half (43%) were less than secondary education level. Retired patients constituted (13%) of all patients. Lack of exercise was the most frequent risk factor among 82% of patients, followed by smoking (56%), use of regular medication (44%), diabetes (30%), hypertension (15%), history of pelvic surgery (14%) alcoholism (13%), and drug addict (8%). CONCLUSION: Erectile dysfunction is a problem of not only old age but also of middle and young age. This might be attributed to the high frequency of some risk factors such as diabetes mellitus, hypertension, smoking, alcohol consumption, and drug addiction. This finding may reflect the necessity for construction of prevention strategies. PMID- 11255609 TI - Effect of maternal education on the rate of childhood handicap. AB - OBJECTIVE: The objectives of this study were to determine the relation between maternal education and various maternal risk factors, identify the impact of maternal education on the risk of childhood handicap and estimate the proportion of childhood handicap that can be prevented by maternal education. METHODS: Data was collected from all married women attending the two major maternity and child hospitals in Jeddah during April 1999. Women with at least one living child were interviewed for sociodemographic factors and having at least one handicapped child. The risk of having a handicapped child and the population attributable risk percent were calculated. RESULTS: Some potential risk factors are dominant in our society as approximately 30% of women did not attend school and 84% did not work. Consanguineous marriages accounted for about 43%. Pre-marriage counseling was limited as only 10% of women counseled before marriage. The proportion of unemployment and consanguineous marriages decreased significantly by increase in maternal education level. Conversely, the proportion of women reporting pre-marriage counseling increased significantly by increase in maternal education level. Approximately, 7% of women reported having at least one handicapped child. The risk of having a handicapped child showed a significant sharp decline with increase in maternal education level. At least 25% of childhood handicap can be prevented by achieving female primary education and up to half of cases can be prevented if mothers finish their intermediate education. CONCLUSION: Female education plays a major role in child health. The results of this study suggest investment in female education, which would have substantial positive effects in reducing incidence of childhood handicap in Jeddah. PMID- 11255611 TI - Hypertension control in a community health centre at Riyadh, Saudi Arabia. AB - OBJECTIVE: This study was carried out to determine the degree of control of hypertension and the most commonly used drugs for hypertensive patients attending our community health center. METHODS: A cross sectional study carried out by randomly examining the case notes of patients attending our primary care clinics. RESULTS: Case notes of 3747 patients were examined, 2064 (55%) females (mean age 23.76 years) and 1683 (45%) males (mean age 24.63 years). Prevalence of hypertension was 3% (108 patients), 3% (63 patients) and 3% (45 patients) for females and males respectively. Majority of patients 16 (35%) males and 32 (51%) females had blood pressure of 141-160/90-100 mmHg. Seventeen (37%) males and 15 (24%) female patients had blood pressure < 140/90 mm Hg. Among 108 hypertensive patients, 29 (65%) males and 44 (69%) females were on single drug. The most commonly used drugs were ACE inhibitors (35%), calcium channel blockers (17.5%) and beta- blockers (14%). CONCLUSION: This study like some other studies shows that control of hypertension falls short of recommended goals. There is need to adopt a strategy that incorporates health education about life style and proper protocol as this has been found useful in other studies. PMID- 11255610 TI - Prevalence of hypertension in obese and non-obese Saudis. AB - OBJECTIVE: Obesity occurs at a high prevalence in the Saudi population. Studies in literature show that hypertension occurs more frequently in obese individuals. This study was designed to determine the prevalence of hypertension in obese Saudis in comparison with results obtained in non-obese individuals. METHODS: The screening involved a statistically designed household screening program. Only adults 14-70 years of age were included in the study. Blood pressure (systolic and diastolic) was measured when the individuals were in sitting position and height and weight were used to calculate Body Mass Index. All individuals with Body Mass Index > 30 were classified as obese and hypertension was measured as systolic blood pressure > 140 and diastolic blood pressure > 90 or both. The prevalence of hypertension was calculated in the obese and non-obese group. Chi square analysis was carried out to determine the significance of the difference in prevalence in different groups. RESULTS: In the non-obese males and females the prevalence of hypertension was 4.8% and 2.8%. While in the obese group the prevalence was almost 1.6 times higher in the males (8%) and 3.52 times higher (8%) in the female obese. The results were separated on the basis of the province to which the population belonged and hypertension prevalence was calculated in the obese and non-obese. In each region the prevalence of hypertension was higher in the obese group compared to the non-obese group. Non-obese females had significantly lower hypertension prevalence than the male in the same province but the hypertension prevalence was higher in the females compared to the male in the obese group. Male in the Eastern, Southern and Western provinces did not show an increased hypertension prevalence in the obese. CONCLUSION: Since the prevalence of obesity is high in Saudis and since obesity and hypertension occur together and cause serious complications, it is strongly suggested that measures are adopted to decrease prevalence of obesity and its underlying complications. Awareness programs are required at the level of the general public for successful implication of preventive programs. PMID- 11255612 TI - Antibiotic susceptibilities of Helicobacter pylori. AB - OBJECTIVE: The aim of this study was to evaluate the prevalence of resistance among 83 Helicobacter pylori isolates cultured from biopsies taken during routine endoscopies in 1998-1999. METHODS: Minimum Inhibitory Concentration of amoxicillin, tetracycline, clarithromycin and metronidazole were determined by Epsilometer test. RESULTS: Forty-seven strains (57%) were resistant to metronidazole, and 27 (32.5%) were resistant to clarithromycin. Twenty of the 27 strains resistant to clarithromycin were also resistant to metronidazole. None of the strains were resistant to amoxicillin or tetracycline. CONCLUSION: A high percentage of patients from Bahrain were infected with resistant strains of Helicobacter pylori. Antibiotic resistance monitoring is very important and unified national treatment policies are needed. PMID- 11255613 TI - Rate of wound infection after clean surgery. AB - OBJECTIVE: The aim of this study is to determine the rate of wound infection after clean surgical operations without using of prophylactic antibiotics and to investigate the relation between surgical wound infection with patient s age, sex, type of hospital, and the difference in surgeons. METHODS: This study carried out in Sana'a city on 238 patients who underwent clean operations in two governmental and two private hospitals by four surgeons between 1998-1999. Patients at high risk of infection were excluded. RESULTS: The rate of wound infection was 8%. The study revealed statistically significant difference (P=0.011) in the infection with elderly patients (25% infection in elderly patients compared with 6% in less than 60 years old patients). It was also found that wound infection rate differs with the difference in surgeons; the rate did not exceed 3% with one surgeon (the author) in comparison with 13% with other surgeons. This difference is statically significant (P= 0.003). Differences in rates of infection with sex of the patient and type of the hospital were statistically insignificant. CONCLUSION: The study was concluded that the rate of wound infection after clean surgery without prophylactic antibiotics in Yemen is higher than many other countries, surgeon and the age of the patient were the risk factors of importance. It is suggested to give antibiotics as prophylaxis to elderly patients and to rise the awareness of the surgeons and nurses in order to improve their practice. PMID- 11255614 TI - Epidemiology of bronchial asthma among school boys in Al-Khobar city, Saudi Arabia. AB - OBJECTIVE: The objective of this cross-sectional study was to determine the prevalence of bronchial asthma among Saudi School boys at Al-Khobar city. METHODS: This is a cross-sectional study. The methodology included the distribution of a self administered questionnaire which was filled by the parents of 1482 school boys who satisfied the selection criteria of the study. RESULTS: The prevalence rate of Questionnaire Diagnosed Asthma and Physician Diagnosed Asthma were 9.5% and 8%. Questionnaire Diagnosed Asthma school boys and their parents suffered significantly higher rates of allergenic diseases and environmental factors (pets at home, passive smoking) than non Questionnaire Diagnosed Asthma. CONCLUSION: The prevalence of Questionnaire Diagnosed Asthma among schoolboys in Al-Khobar city was more than that which was described earlier. However, this rate was less than those reported from other parts of the Kingdom but higher than the ones reported from Arab, developing and European countries. There is evidence that a combination of genetic and environmental factors play a major role in the etiology of this disease. Based on the results of this study, appropriate and practical measures need to be taken to identify causes and initiate control programs. PMID- 11255616 TI - Hairy cell leukemia-variant. AB - Hairy cell leukaemia variant is a very rare chronic lymphoproliferative disorder and is closely related to hairy cell leukemia. We hereby describe a case of hairy cell leukaemia variant for the first time in Saudi Arabia. An elderly Saudi man presented with pallor, massive splenomegaly, and moderate hepatomegaly. Hemoglobin was 7.7 g/dl, Platelets were 134 x109/l and white blood count was 140x10 9/l with 97% being abnormal lymphoid cells with cytoplasmic projections. The morphology, cytochemistry, and immunophenotype of the lymphoid cells were classical of hairy cell leukaemia variant. The bone marrow was easily aspirated and findings were consistent with hairy cell leukaemia variant. PMID- 11255615 TI - Analysis of false positive and false negative cytological diagnosis of breast lesions. AB - OBJECTIVE: To study the reasons for interpretive errors in false negative and false positive diagnosis of breast carcinoma on fine needle aspiration cytology material. METHODS: We reviewed only those cases in which cytohistological discrepancies were found, where the cytologic material was abnormal and to some extent misinterpreted or both. RESULTS: There was only one false negative case (false negative fraction 0.32%) proved histologically as ductal carcinoma and four false positive cases (false positive fraction 1.2%); 2 fibroadenoma; 1 fibrocystic disease; and 1 stromal fibrosis. Smears of the two false positive fibroadenoma cases showed very high cellularity, overcrowded clusters and frequent stripped nuclei. The fibrocystic case showed tight clusters of apocrine cells and sheets of loosely aggregated macrophages that were over interpreted. Smears of the false negative ductal carcinoma was hypocellular overall, and the cells showed minimal nuclear pleomorphism. CONCLUSION: Overcrowded clusters and hypercellular smears should be carefully assessed for uniformity of cells and detailed nuclear and cytomorphological features. If the full-blown malignant cytomorphological changes are not visible, a diagnosis of suspicious or inconclusive should be made and frozen section recommended before surgery. Hypocellularity and relatively nuclear monomorphism are the reasons for failure to diagnose malignant breast lesions. Careful attention should be paid to extreme nuclear monomorphism and absence of naked bipolar cells. A cytologically atypical or suspicious diagnosis together with positive radiological and clinical findings should suggest a diagnosis of malignancy. PMID- 11255617 TI - Pseudolesion in left lobe of the liver due to superior vena cava obstruction. AB - We report a case of lymphoma in which abnormal strong enhancement in the medial segment of left lobe of liver during arterial phase of triphasic helical computed tomography due to superior vena cava obstruction. PMID- 11255618 TI - Hypothyroidism. The need for high degree of suspicion for early diagnosis. PMID- 11255619 TI - Insulin drip can be dangerous. PMID- 11255620 TI - Neonatal salmonella meningitis. PMID- 11255621 TI - The characteristics of systemic lupus erythematosus. PMID- 11255622 TI - Integrated health care systems: major issues and lessons learned. PMID- 11255624 TI - A revolutionary advance in disease management. PMID- 11255623 TI - Special report: health care staffing shortages in the news. PMID- 11255626 TI - Health risk as a cause of action. PMID- 11255625 TI - Will you respect me in the morning? The effect of Bill 68 on competent consent. PMID- 11255627 TI - Ontario's Community Treatment Orders: how did we get there and where do we go now? An advocate's perspective. PMID- 11255628 TI - In vivo histological changes occurring in hydroxyapatite cranial reconstruction- case report. AB - Histological changes were observed in a hydroxyapatite plate and hydroxyapatite granules used to repair a craniotomy defect and removed after 2 years and 9 months of use. The hydroxyapatite plates and granules had completely fused to the cranium, with new bone formation on the dural side extending in a three dimensional matrix along the pores with the Haversian system in the center. New bone formation was less extensive under the artificial dura than under normal dura. This finding suggests that the dura has the ability to promote bone formation. A new vessel was found along the interconnecting pores. The interconnecting pores allow osteoconduction in the hydroxyapatite plate, so new bone formation can progress. Hydroxyapatite has osteoconduction properties and is biocompatible, so gains strength in vivo through new bone formation, and is the ideal material for artificial bones. Factors important to achieving good bone formation after cranial reconstruction surgery include presence of the dura, and pore size approximate to the Haversian system (100-500 microns) and interconnecting pores in the hydroxyapatite plate. PMID- 11255629 TI - Brain hypothermia relieves severe brain swelling following acute major cerebral artery occlusion. AB - Seven patients were treated with brain hypothermia following acute major cerebral artery occlusion to utilize the suppressive effect against brain swelling. Five patients had internal carotid and two had proximal middle cerebral artery occlusion. Except for the first two cases, hypothermia was introduced early and the temperature reached 35.0 degrees C within 6 hours after the onset. The core temperature finally stabilized between 32 degrees C to 34 degrees C. Hypothermia had a suppressive effect against brain swelling and the temperature showed a significant correlation to intracranial pressure. Recurrence of brain swelling was observed during the rewarming process, but two patients became independent and three patients were moderately disabled in wheelchairs. Only two patients died. Brain hypothermia is an effective treatment for acute major cerebral artery occlusion through the relief of brain swelling. The overall outcome may be improved by combining brain hypothermia with other conventional therapies such as osmotherapy and external decompression implemented with an extended period of rewarming. PMID- 11255630 TI - Time course of expression of three nitric oxide synthase isoforms after transient middle cerebral artery occlusion in rats. AB - The involvement of nitric oxide synthase (NOS) in ischemia was evaluated by detecting the expression of neuronal NOS (nNOS), endothelial NOS (eNOS), and inducible NOS (iNOS) by the immunohistochemical method in the rat model of middle cerebral artery (MCA) occlusion. Transient MCA occlusion (2 hours) was induced in 32 male Wistar rats by extracranial insertion of a 3-0 nylon thread through the internal carotid artery into the MCA. Animals were killed at 0, 6, 24, 72, and 168 hours after MCA occlusion (n = 6, 6, 8, 6, and 6, respectively). The brains were fixed with periodate-lysine-paraformaldehyde, frozen, and sectioned. Sections were stained with polyclonal antibody against nNOS, eNOS, and iNOS. Each section was evaluated by microscopic observation (x100). The number of nNOS positive neurons was 41.6 +/- 5.8 (mean +/- SD) in the control hemisphere. nNOS was upregulated in the ischemic hemisphere (88.3 +/- 18.9), especially in the border zone at 6 hours after MCA occlusion. However, the number decreased to 36.4 +/- 3.6 and 26.3 +/- 7.3 in the ischemic hemisphere after 72 and 168 hours, respectively. eNOS immunoreactivity was present in the endothelium of major vessels at each time point. eNOS was not detected in the microvessels before ischemia, but faint staining was found in the endothelium at 6 hours after MCA occlusion. Immunostaining became more intense thereafter. Faint iNOS immunoreactivity was seen in the microvessels at 6 hours after MCA occlusion. Macrophages in the ischemic core and astrocytes in the border zone showed immunoreactivity to iNOS at 72 and 168 hours after MCA occlusion. Three types of NOS must be related to different stages of ischemic brain damage. nNOS may be neurotoxic in ischemia in the early phase, like iNOS in the late phase. On the other hand, eNOS seemed to be neuroprotective in all stages. These observations suggest the necessity for tailored therapeutic intervention against NOS isoforms at each stage in patients with ischemic stroke. PMID- 11255631 TI - Improvement of phase-contrast flow measurements: opposite directional flow encoding technique to eliminate the influence of the Maxwell term phase errors. AB - A new method termed the opposite directional flow-encoding (ODFE) technique is proposed to increase the accuracy and the reproducibility of phase-contrast flow measurements by correcting the non-linear background of velocity images induced by concomitant magnetic fields (Maxwell terms). In this technique, the volume flow rate is calculated from the difference of two region of interest (ROI) values derived from two velocity images obtained by reversing the flow-encoding direction. To evaluate the technique, various phantom experiments were carried out and volume blood flow rates of internal carotid arteries (ICAs) were measured in four volunteers. The technique could measure the volume flow rates of the phantom with higher accuracy (mean absolute percentage error = 1.04%) and reproducibility (coefficient of variation = 1.18%) than conventional methods. Flow measurements with the technique was not significantly affected by ROI size variation, measuring position, and flow obliquity not exceeding 30 degrees. The volume flow rates in the ICAs of a volunteer were measured with high reproducibility (coefficient of variation = 2.89% on the right, 1.48% on the left), and the flow measurement was not significantly affected by ROI size variation. The ODFE technique can minimize the effect of the non-linear background due to Maxwell terms. The technique allows use of ROIs of approximate size including the flow signal and provides accurate and objective phase-contrast flow measurements. PMID- 11255632 TI - Growth of occult arteriovenous malformation after cerebral hemorrhage demonstrated by serial magnetic resonance imaging--case report. AB - A 19-year-old male presented with sudden onset of right hemiparesis caused by left cerebral hemorrhage. Cerebral angiography demonstrated no vascular abnormality, and the hematoma was removed surgically. At operation, no abnormal vascular lesion was found in the brain adjoining the hematoma. Two years later, magnetic resonance (MR) imaging demonstrated a few foci of flow voids adjacent to the hematoma cavity. Four years after the hemorrhage, MR imaging showed more extensive flow-void abnormalities that indicated growth of an occult arteriovenous malformation (AVM). Cerebral angiography indicated a definite AVM supplied mainly by branches of the middle cerebral artery. Total resection of this lesion was performed. The histological diagnosis was typical AVM. Immunohistochemistry with vascular endothelial growth factor showed staining in the walls of the abnormal vessels. Serial MR imaging is very useful for the diagnosis and management of occult AVMs. PMID- 11255633 TI - Hemifacial spasm due to cerebellopontine angle meningiomas--two case reports. AB - A 54-year-old female and a 49-year-old female presented with complaints of hemifacial spasm. Both patients underwent surgery to remove cerebellopontine angle meningiomas. In one case, no vascular compression was observed at the root exit zone. The tumor was removed subtotally leaving residual tumor adhered to the lower cranial nerves. The hemifacial spasm disappeared immediately after the operation. The residual tumor was treated using gamma knife radiosurgery. In the other case, the root exit zone of the facial nerve was compressed by both the tumor and anterior inferior cerebellar artery and the tumor was removed totally. Postoperatively, the hemifacial spasm disappeared, but the patient suffered facial nerve paresis and deafness that was probably due to intraoperative manipulation. However, the facial nerve paresis gradually improved. Cerebellopontine angle meningioma with hemifacial spasm must be treated by surgical resection limited to preserve cranial nerve function. Subtotal removal with subsequent radiosurgery to treat the remaining tumor tissue is one option for the treatment of cerebellopontine angle meningioma. PMID- 11255634 TI - Intracranial meningeal malignant fibrous histiocytoma mimicking parasagittal meningioma--case report. AB - A 69-year-old female presented with a rare intracranial meningeal malignant fibrous histiocytoma (MFH). The neuroimaging appearance of this tumor was very similar to parasagittal meningioma. The tumor was grossly totally removed, and local irradiation of 50 Gy was performed. The histological diagnosis was MFH. The patient recovered from the preoperative deficits, and no recurrence was observed by 27 months after surgery. PMID- 11255635 TI - Midline posterior fossa teratoma--case report. AB - A 20-day-old female neonate presented with an immature teratoma in the midline posterior fossa. The tumor was totally removed but the patient died of pneumonia. Teratoma is a rare tumor, but very difficult to treat as the patients tend to be young, and the outcome is very poor. PMID- 11255636 TI - Migration of a lumboperitoneal shunt catheter into the spinal canal--case report. AB - A 50-year-old female suffered upward migration of a lumboperitoneal (LP) shunt catheter into the spinal canal, manifesting as disturbance of short-term memory. Revision of the shunt confirmed that the tube had migrated into the spinal canal. The tube was pulled back into the peritoneal cavity and attached firmly to the fascia with a new anchoring device. LP shunts have the advantages of technical simplicity and extracranial procedure, but firm fixation is recommended since movements of the spine may cause proximal tube migration. PMID- 11255637 TI - [Induction of antimeningitis immunity by synthetic peptides. II. Immunoactive synthetic fragments of OpaB protein from Neisseria meningitidis]. AB - Mice of various lines were immunized by 11 synthetic peptides that correspond to the sequences of fragments of the OpaB protein from the outer membrane of Neisseria meningitidis involving the known human T-helper epitopes and all the potential mouse T-helper epitopes calculated for the protein. The mice were immunized with the free peptides without their conjugation with a protein carrier. Most of the peptides were found to induce in mice the production of antipeptide antibodies. The mice protection against the experimental infection by a virulent strain of N. meningitidis of the B serotype was studied, and two peptides were shown to exert the most pronounced protective effect. PMID- 11255638 TI - [The study of the metal-binding region in human growth hormone using immobilized metal ion affinity gel-electrophoresis]. AB - The zinc(II)-binding affinities of recombinant human growth hormone and two its mutants, 14-33 and 14-95, were studied using Immobilized Metal Ion Affinity Gel electrophoresis (IMAG). The mutant hormones, composed of polypeptide chain segments of the human and porcine growth hormones, lacked His18, which may be crucial for binding of the intact hormone to the transition metal ions. The mutations did not affect the affinity of human growth hormone to immobilized zinc ions; the structural analysis implied that the human growth hormone contains two IDA-Zn(II) potential sorption sites formed by amino acid residues His21, Asp171, and Glu174 and/or His18 and Glu174. PMID- 11255639 TI - [Inhibition of inorganic pyrophosphatase from Escherichia coli with inorganic phosphate]. AB - The interaction of inorganic pyrophosphatase from E. coli with inorganic phosphate (Pi) was studied in a wide concentration range of phosphate. The apoenzyme gives two inactive compounds with Pi, a product of phosphorylation of the carboxylic group of the active site and a stable complex, which can be detected in the presence of the substrate. The phosphorylation occurs when Pi is added on a millimole concentration scale, and micromole concentrations are sufficient for the formation of the complex. The formation of the phosphorylated enzyme was confirmed by its sensitivity to hydroxylamine and a change in the properties of the inactive enzyme upon its incubation in alkaline medium. The phosphorylation of pyrophosphatase and the formation of the inactive complex occur upon interaction of inorganic phosphate with different subsites of the enzyme active sites, which are connected by cooperative interactions. PMID- 11255640 TI - [Polysaccharides of calcareous algae and their effect on calcification process]. AB - The composition and structure of polysaccharides from several groups of calcareous algae (including calcareous cyanobacteria), which differ in the calcification mode (extracellular, cell wall, or intracellular), are reviewed. Two families of marine algae, Corallinaceae (Rhodophyta) and Cocolithophoraceae (Prymnesiophyta = Haptophyta), are considered in detail; they exhibit the cell wall and intracellular calcification modes, respectively, and synthesize unusual polysaccharides that seem to directly participate in the calcification process. PMID- 11255641 TI - [Methods of preparation of recombinant proteins-cytokines. IV. Renaturation of recombinant human interleukin-3]. AB - Renaturation of recombinant human interleukin-3 produced as inclusion bodies in the transformed cells of Escherichia coli was studied and optimized. Importance was shown of removing from the protein solution the hydrophobic cellular components causing irreversible aggregation of the protein under renaturation conditions. An effect of pH on the secondary structure of the denatured protein was revealed by CD spectroscopy. It was thereby found that at pH 8.5, which is the optimal value for denaturation, the protein has the secondary structure most close to the native one. The isolation according to the scheme proposed allows preparation of interleukin-3 in 50% yield with 99% purity and biological activity 2 x 10(7) U/mg. PMID- 11255642 TI - [Synthesis and characteristics of modified oligodeoxyribonucleotides containing 2'-O-(2,3-dihydroxypropyl)uridine and 2'-O-(2-exoethyl)uridine]. AB - A new uridine derivative, 2'-O-(2,3-dihydroxypropyl)uridine, and its 3' phosphoramidite were obtained for direct introduction into oligonucleotides during automated chemical synthesis. Oligonucleotides 10 to 15 nt long harboring 2'-O-(2,3-dihydroxypropyl)uridine residues were synthesized; periodate oxidation of these oligomers gave oligonucleotides containing 2'-O-(2-oxoethyl)uridine residues. The presence of a reactive aldehyde group in 2' position of the carbohydrate moiety was confirmed by the interaction with p-nitrophenylhydrazine and methionine methyl ester. The thermostability of DNA duplexes containing modified units is practically indistinguishable from that of the natural analogues. PMID- 11255643 TI - [Isolation and characterization of polysaccharides from Tansy]. AB - Using extraction with 0.75% aqueous ammonium oxalate, the following polysaccharide fractions were isolated: tanacetans TVF, TVS, and TVR from floscules, sprouts, and roots, respectively, of Tanacetum vulgare L., spread throughout the European North of Russia. The sugar chain of tanacetan TVF consists of D-galacturonic acid (61.4%), arabinose (14.7%), galactose (10.2%), and rhamnose (3.7%) as the main constituents as well as xylose, glucose, mannose, apiose, and 2-O-methylxylose in trace amounts. Tanacetans TVS and TVR were shown to differ in the sugar quantitative composition. They contain 67 and 28% galacturonic acid, respectively. A partial acid hydrolysis of the tanacetan TVF gave a polysaccharide fragment TVF1, alpha-1,4-D-galacturonan (GalA 98.2%). Digestion with pectinase (alpha-1,4-D-polygalacturonase) resulted in fragment TVF3, containing residues of arabinose (27.1%) and galactose (17.3%). NMR spectroscopy allowed detection of the terminal residues of alpha-Araf and beta Galp as well as of the residues of alpha-Araf substituted in 3,5- and 5 positions. Thus, tanacetan TVF was proved to be a pectic polysaccharide. PMID- 11255644 TI - [Synthesis of novel porphyrin-quinone compounds]. AB - New porphyrin-quinone dyad systems containing spacer groups of various lengths and structures and sterically hindered 5,10,15,20-tetrakis(3,5-di-tert butylphenyl)porphyrin as an electron donor were synthesized. These compounds seem to be promising models for studying the photoinduced electron transfer. PMID- 11255645 TI - [Comparative study of DNA-specific dyes of the indole and benzimidazole derivates]. AB - Various models of complex formation of low-molecular ligands with DNA are considered. Using the Scatchard model, parameters of binding of fluorescent monophenylindole, monophenylbenzimidazole, and bisbenzimidazole dyes with calf thymus DNA were evaluated. By means of graphic (nonparametric) and correlation analysis, various spectral and complexation properties of these dyes in the presence of DNA are compared. PMID- 11255646 TI - [Geriatric nephrology: new branch of study about internal diseases?]. AB - This review refers to the main geriatric health problems, especially dysfunction of the autonomic sympathetic system and urine incontinence in the elderly, pathophysiology of aging kidney, as well as distinct clinical course of many kidney diseases in the advanced age. PMID- 11255647 TI - [Bone mineral density in dialysis patients: the optimal region of interest depending on parathormone levels]. AB - Frequency of osteoporosis in patients on dialysis with respect to the optimal Region of Interest (ROI) has not been established. Frequency of this complication varies in different studies, especially because of no agreement according to the optimal ROI. We studied 71 patients: 32 on CAPD and 39 on HD (31F, 40m. age mean 56 +/- 29) on dialysis for 3 to 81 months. BMD was measured using dual energy X ray densitometry (DXA) in L1-L4 segment of the vertebral column, femoral neck and forearm. The results of the BMD were presented as T-score in the standard deviation of the mean peak bone mass according to WHO criteria. Males and females were evaluated separately. Depending on the ROI, osteoporosis was diagnosed in 20 to 68% of women and in 2 to 40% of men. In females the most sensitive region of interest appeared to be forearm, in males it was proximal part of the femur. In these ROI's the lowest values of bone mineral density in both genders were found in patients with the level of iPTH higher than 200 pg/ml. We did not find any significant difference in BMD between patients on CAPD and HD. Values of bone mineral density were not related to the duration of dialysis therapy. On the results of our study it has been concluded, that optimal region of interest for BMD evaluation in dialysed patients depends on gender of the patient and functional status of parathyroid glands but is not related to either the method or duration of dialysis therapy. PMID- 11255648 TI - [Acute kidney failure in the course of rhabdomyolysis with hemodialysis in personal material from 1995-1999]. AB - Eleven cases (5 F + 6 M; mean age 48.0 years) of acute noninflammatory renal failure (ANRF) in the course of rhabdomyolysis (RBM) were treated with hemodialysis in years 1995-1999. The causes of RBM were the following: ischemia of lower limbs after vascular operations (4 cases), exhausting exercise with rapid body cooling (3 cases), multiorgan failure after traffic accident, acute myositis (1 case), status epilepticus (1 case), rapid clinical course of viral infection (1 case). It was necessary to perform from 1 to 13 hemodialyses in every patient. In nine cases, complete normalization of renal function during 5 to 30 days of therapy was achieved. Two patients died due to multiorgan complications after vascular operations despite effective dialysis therapy. The following correlation were found: positive between initial values of creatine phosphokinase (CPK) activity and creatinine and uric acid concentrations in the blood and negative correlation between CPK and serum calcium concentrations. The higher initial values of CPK activity were observed the more hemodialysis procedures were necessary and the longer time was needed to normalize renal function. On the base of initial, limited up to now, own results it seems that hemodialysis in ANRF in the course of RBM should be started immediately in cases with high activity of CPK in the blood (above 10,000 U/L). PMID- 11255649 TI - [Evaluation of nephrologic "risk" in a newly diagnosed case of plasma cell dyscrasias from personal material (1994-2000)]. AB - Among 149 patients with recently recognized plasma cell dyscrasia (PCD) in years 1994-2000 72 persons with serologically and nephrologically documented diagnostic profile were selected. In this group of pts we assessed dependence between degree of reduced glomerular filtration rate (GFR), evaluated by serum creatinine concentration and calculated with Barasckay's formula and hypercalcemia, hyperuricemia as well as type of monoclonal protein in urine. RESULTS: We revealed statistically significant higher values of calcium (p = 0.005), uric acid (p = 0.000001) concentrations and higher occurrence of Bence-Jones proteinuria (mainly kappa) in 22 patients with serum creatinine > 1.5 mg/dl in comparison with 50 patients with serum creatinine < or = 1.5 mg/dl. Among 72 patients, GFR > 90 ml/min, calculated with Barasckay's formula, was stated only in 9 patients (12.5%). There was no difference in nephrotoxity between kappa and lambda light chains with reference to serum creatinine concentration and GFR. The group of 12 patients with light chain dyscrasia (LCD) had higher degree of nephrotoxicity in comparison with other forms of PCD. On the basis of our study we concluded that patients with clinical suspicion of PCD, especially those with LCD are referred to a special Protein Laboratory too late, it means at the time of significant nephrological risk in the form of low glomerular filtration rate, hypercalcemia and hyperuricemia. PMID- 11255651 TI - [Long-term evaluation results for adequacy of continuous ambulatory peritoneal dialysis in patients with diabetes]. AB - The aim of the study was to analyze CAPD adequacy using own scoring including three groups of parameters: "Adequest" parameters, clinical and biochemical indices. The studied population comprised 17 patients treated with CAPD: 12 DN patients, in this number 6 female and 6 male, aged 44 +/- 10.9 years, treated with CAPD for 47 +/- 10.5 months, observed for 38.5 +/- 12.8 months were included in group 1 (studied group). 105 studies were performed in this group. Group 2 (control) consisted of 5 non-diabetic patients, including 2 female and 3 male, aged 51 +/- 13.6 years, treated with CAPD for 51 +/- 20.2 months, observed for 42.6 +/- 15.2 months. 47 studies were performed in group 2. On the base of own scoring group 1 was divided into two subgroups: 1A including 8 persons dialyzed adequately and 1NA consisting of 4 persons dialyzed inadequately. All patients from group 2 were dialyzed adequately. No significant differences concerning "Adequest" scoring as well as clinical and biochemical scorings were found between subgroup 1NA and group 2. The three kinds of scoring were significantly lower in subgroup 1NA in comparison to other groups of patients. The high level of consistence expressed as percent coefficients between particular scorings was stated in groups dialyzed adequately. This consistence was lower in group 1NA. The results of Spearman correlation test regarding three kinds of scoring were not as clear as consistence assessment using percent coefficients. The results of the study let to conclude that DN patients can reach similar level of CAPD adequacy to non-diabetics. High level of consistence between three kinds of scoring confirms the usefulness of own method of adequacy assessment. However the analysis of CAPD adequacy should not be based on single group of parameters. PMID- 11255650 TI - [Enhanced neuroticism in relation to cardiovascular risk factors in men]. AB - Chronic stress is well known as a cardiovascular risk factor in men. There are few reports referred to the relationship between cardiovascular risk factors and level of neuroticism in the psychological examination. We analysed 77 healthy men in mean age of 33.3 +/- 7.4 years being at risk of chronic professional stress. Level of neuroticism was examined by Geras scale and expressed as: low (1-4 sten) -group 1, mean (5-6 sten)--group 2 and high (7-10 sten)--group 3 level of neuroticism. The common biochemical, clinical and environmental cardiovascular risk factors were examined. In the whole group of 77 probands we identified 24 (31%) (group 2) and 10 (13%) (group 3) persons with mean and high level of neuroticism. In the group 2 we find the highest cholesterol (222.7 +/- 39.4 mg/dl), LDL cholesterol (147 +/- 35.1 mg/dl, triglicerydes (144.9 +/- 93.8 mg/dl) and apolipoprotein B (1.11 +/- 0.31 g/l) levels statistically higher than in the other group of neuroticism. In group 3 we find the highest glucose (96.5 +/- 7.4 mg/dl) and fibrinogen (353.8 +/- 39.2 mg/dl) levels. The worst results of the environmental inquiry were obtained in the group 2 (obesity (BMI > 30 kg/m2) in 25%, hypertension in 12.5%, nicotinism in 54%, low physical activity in 79% and positive to cardiovascular disease family history in 91%). Because we found correlation mostly in the group of mean level of neuroticism these results must be confirmed in the larger group of probands using other psychological tests. PMID- 11255652 TI - [Continuous ambulatory peritoneal dialysis: is long-term therapy possible?]. AB - The aim of this study was an analysis of long-term results of CAPD. 12 patients treated with CAPD over 5 years were analysed retrospectively. The most common cause of end-stage renal disease, present in 8 patients, was diabetes nephropathy. In the 1st, 3rd and 5th year of CAPD the biochemical parameters, dialysis adequacy, peritoneal transport were evaluated. Despite the fact that adequacy of CAPD was achieved, the progression of cardiovascular complications occurred, especially in diabetic patients with type 1 diabetes. It is suggested that early kidney transplantation should be considering, before cardiovascular complications progressed. PMID- 11255653 TI - [Problems with therapy for renovascular hypertension in Takayasu's arteritis--on the basis of two cases]. AB - Takayasu's arteritis is non-specific vasculitis, affecting aorta and its main branches. Renal involvement is usually manifested by renovascular hypertension followed by ischemic nephropathy. Early start of immunosuppressive treatment can substantially improve prognosis and delay progress of dangerous organic complications. On the basis of two cases of Takayasu's arteritis with renal involvement, we paid the attention to progressive nature of disease with renovascular hypertension, requiring corrective operation of renal arteries, despite intensive immunosuppressive treatment. PMID- 11255654 TI - [Goodpasture's syndrome--continuing clinical problem]. AB - On the case of too late diagnosed Goodpasture's syndrome with further fatal clinical course necessity of early diagnosis and aggressive immunosuppressive therapy is stressed. PMID- 11255655 TI - [Urinary tract infections in the elderly]. AB - This review refers to the special aspects of diagnosis and therapy of urinary tract infections in the elderly. PMID- 11255657 TI - [The role of peritoneal dialysis in intensive medical care: benefits and limitations]. AB - This review analyses the advantages and limitations of actually existing technics of continuous peritoneal dialysis (CPD) which is proposed more often as the renal replacement therapy of acute renal failure in patients hospitalized in intensive care units (ICU's). Application of CPD for routine practice in polish ICU's needs the permanent disposable equipment for manual and automatic CPD as well as training the medical staff in the nearest nephrological reference center for PD. The last one is especially urgent, because growing population of pts treated with CAPD and APD in Poland. Some of these pts might be hospitalized in ICU's because of extrarenal complications. They need to be simultaneously treated with the technics of CPD used ambulatory before hospitalization. PMID- 11255656 TI - [Stress as a risk factor for cardiovascular disease]. AB - This review refers to the experimental and clinical investigations on stress as a important risk factor of cardiovascular diseases. PMID- 11255658 TI - [Low turnover renal osteodystrophy--current state and perspectives]. AB - Invasive and non invasive diagnostic procedures as well as actually recommended prophylactic and conservative therapy of low turnover renal osteodystrophy are presented. PMID- 11255659 TI - [Glycosaminoglycans in diabetic nephropathy]. AB - This review refers to the nephro-protective role of glycosaminoglycans, especially in diabetic nephropathy. PMID- 11255660 TI - [The significance of adequacy for success of an automated peritoneal dialysis program]. AB - The dynamic development of APD causes the increasing importance of issues regarding the adequacy of this renal replacement therapy method. Basic parameters of effective dialysis are still Kt/V and weekly creatinine clearance. The lack of steady state of substance concentration in different body compartments is a potential cause of mistakes in calculation of APD adequacy kinetic parameters. The solute removal index (SRI) may be used as an alternative method of dialysis efficacy assessment. However, there is no "ideal" marker of adequacy. Therefore, it should be assessed on the base of analysis including kinetic, clinical and biochemical data. Residual renal function (RRF) is an important factor of successful APD, but there are different opinions concerning the influence of automatic dialysis on RRF preservation presented in literature. The characteristics of peritoneal transport is the basic criterion for the qualification of patients to APD program. Automated techniques are recommended for patients being high-average transporters and particularly high transporters. The evolution of APD creates new perspectives for adequacy assessment and improvement of dialysis efficacy. The performance of multicenter studies and usage of new parameters of adequate dialysis will be very important for the development of APD. The employment of alternative fluids, introduction of modernized catheters and construction of "intelligent cyclers" will create new possibilities for programming and monitoring of dialysis and improvement of patient's life quality. PMID- 11255661 TI - [Report from the 1st korean-Polish seminar: "Theoretical, technical and clinical aspects of renal replacement therapy (5/18-21/2000)]. AB - This was the first Korean-Polish seminar on renal replacement therapies which provided a survey of current research being carried out in both countries, delivered by leading experts in this field. The topics included malnutrition in uremia, biocompatibility, risk factors and complications in renal replacement therapy, diabetic nephropathy, new solutions for peritoneal dialysis, and peritoneal transport. The organizers strove to select the topics which are currently of interest to researchers from both countries with the purpose to compare results and stimulate discussion concerning possible collaboration in the future. PMID- 11255662 TI - [Report from the seventh international course on peritoneal dialysis, Vicenza, Italy, May 23-26, 2000]. AB - This report is devoted to the Seventh International Course on Peritoneal Dialysis with special interest to the place of peritoneal dialysis in so called "healthy" start of renal replacement therapy, more biocompatible dialysis solutions as well as limitations in the estimation of peritoneal dialysis adequacy using Kt/V for urea and weekly creatinine clearance. PMID- 11255663 TI - [Report from the XXVIII European Symposium on Calcified Tissues, Tampere, Finland, May 6-10, 2000]. AB - This report refers to the actual noninvasive diagnosis and therapy of bone diseases with special interest to osteoporosis. PMID- 11255665 TI - Human mitochondrial genetics. PMID- 11255664 TI - High-density arrays and insights into genome function. PMID- 11255666 TI - The p53 tumour suppressor protein. PMID- 11255668 TI - Genetic susceptibility in infectious diseases. AB - The outcome of infectious disease varies tremendously between individuals due to a number of factors and may therefore be viewed by the geneticist as complex traits. The identification of genes which influence disease outcome is, at present, a resource-intensive project and therefore should not be undertaken without clear evidence, preferably from twin studies, that the genetic contribution is significant. Although three principal techniques are available for the identification of disease susceptibility alleles, they are not applicable to all infectious diseases for logistical reasons. Whether a candidate polymorphic gene is identified through allele sharing studies, from interspecific crosses or taken from the currently available candidate list, the final evaluation will require carefully conducted disease association studies. As we move into the post genomic era, the identification of candidate polymorphisms and the characterization of their functional significance will rapidly increase, which will make the analysis of disease susceptibility in infectious diseases steadily more tractable. PMID- 11255667 TI - Production of active mammalian and viral proteases in bacterial expression systems. PMID- 11255669 TI - Emerging strategies for the chemical treatment of microbial biofilms. PMID- 11255670 TI - The effect of aeration upon the secondary metabolism of microorganisms. PMID- 11255671 TI - On-line, real-time measurements of cellular biomass using dielectric spectroscopy. PMID- 11255672 TI - Genetically modified food crops: current concerns and solutions for next generation crops. PMID- 11255674 TI - The use of chromatography to manufacture purer and safer plasma products. PMID- 11255673 TI - Metabolic engineering of plant cells in a space environment. PMID- 11255676 TI - Use and applications of subtractive antibody screening. PMID- 11255677 TI - Modulation of intestinal permeability: a novel and innovative approach for the oral delivery of drugs, macromolecules and antigens. PMID- 11255675 TI - Possible molecular mechanisms involved in nickel, zinc and selenium hyperaccumulation in plants. PMID- 11255678 TI - Diabetes mellitus and closed-loop insulin delivery. PMID- 11255679 TI - Gene therapy: development of immunostimulatory treatments for cancer. PMID- 11255680 TI - The effects of physical forces on cartilage tissue engineering. PMID- 11255681 TI - Laser reshaping of cartilage. PMID- 11255682 TI - Tools for molecular genetic epidemiology: a comparison of MADGE methodology with other systems. PMID- 11255683 TI - Deriving meaning from genomic information. PMID- 11255684 TI - [Trends in mortality in Serbia, excluding the provinces, 1973-1994]. AB - The war and break up of former Yugoslavia began in 1991. In May 1992 the United Nations imposed economic sanctions on Serbia and Montenegro which were suspended only in November 1995. The purpose of this study was to assess the effects of the war and UN sanctions on health of the population of Serbia without the provinces of Vojvodina and Kosovo. The period 1973-1994 was studied. Mortality data were derived from unpublished and published materials of the Federal Institute of Statistics [1]. Refugees, who, because of civil war, came to Serbia and Montenegro from other parts of former Yugoslavia, were not counted as a part of the population when mortality rates were calculated. Mortality rates were standardized directly using the "European population" as the standard [2]. The least square method was used to fit mortality rates to different trend curves. Linear trend was used whenever it significantly (p < 0.05) demonstrated the existing mortality rates. To measure the possible effect of the war and sanctions (WAS) on mortality between 1991 and 1994, dummy variable (variable WAS) consisting of 0's and 1's was made to signify the passage from the period before and the period after the beginning of the war and sanctions [3]. Over the period 1991-1994, characterized by the war and UN sanctions, in women aged 25-34, 35-44 and 75-84, total mortality was significantly higher than expected on the basis of the trend for the preceding period (p = 0.006, p = 0.000 and p = 0.015 respectively). The opposite effect was found in the age group 85+ (p = 0.012)/Table 2. Of major causes of death, in age group 25-34, mortality from endocrine diseases increased more rapidly in both sexes (p = 0.000) and mortality from urogenital diseases in women decreased more slowly than expected (p = 0.006). On the other hand, in age group 85+ mortality was significantly lower for cardiovascular diseases in both sexes (p = 0.035 and p = 0.006), for respiratory diseases in men (p = 0.011) and for neoplasms in women (p = 0.006)/Table 4. In addition, in the years 1991 and 1992 the increase in mortality from injuries and poisoning was evident in men aged 15-24, 25-34 and 85+ years (Graph 5). Our results show that over the period 1991-1994 changes in mortality were present in some age groups and were caused by certain groups of diseases. In men, besides mortality of infectious disease which decreased more slowly during 1991-1993 than expected, [4], the main departures were found in the mortality from injuries and poisoning and in mortality from endocrine diseases. The excess of death due to injuries and poisoning in the age group 15-34 can be explained as a direct consequence of the war. There were no military operations on the territory of Serbia, but young men from Serbia were nevertheless engaged in the war in other republics of former Yugoslavia. The outstanding increase in mortality caused by injuries and poisoning in men aged 85+ has two explanations. The first is the fact that suicide rate which was on an average of 86 per 100,000 over the period 1984-1990 rose to 140 per 100,000 during the period 1991-1993. In the year 1994 it fell to 92 per 100,000. Since there were no great differences in percent distribution of suicides among all deaths caused by injuries and poisoning in the two periods (27% in 1984-1990 and 20% in 1991-1993), it is clear that the rise of suicidal rate cannot be the only explanation for increased mortality from injuries and poisoning. In a situation when medical services were badly overextended (lack of medical equipment and proper maintenance of the existing equipment, lack of drugs and other medical inputs, a large number of wounded coming from Bosnia as well as numerous refugees) [5, 6], priority had to be given to younger age groups. Higher mortality due to endocrine diseases in men and women aged 25-34 years and higher mortality due to urogenital diseases in women of the same ages can be most probably attributed to poor medical supplies. Although formally excluded from the international economic blockade medical supplies were in practice badly affected by the fact that the dinar was rendered almost worthless and the Ministry of Health could no longer pay the medical inputs. In addition, bureaucratic hurdles of getting clearance from the UN added months of delay and made foreign companies unwilling to trade [5, 7]. The supply and distribution of drugs within the country was also irregular because communication lines were cut and local companies were not prepared to risk distributing drugs that nobody could pay for [7]. Higher than expected mortality in women aged 25-44 over the period 1991-1994 could be probably explained by their higher vulnerability (period of fertility) and the fact that the main burden of family survival was on them, so they had no time to think and to take care of their health. (ABSTRACT TRUNCATED) PMID- 11255686 TI - [Radiotherapy of locally advanced breast carcinoma in elderly female patients]. AB - INTRODUCTION: Breast cancer is the most frequent cancer in elderly patients (over 65 years). The recent data indicate that in women aged over 72 years the incidence of breast cancer is twice greater than in women aged 45 years. As more and more women are getting older, the total incidence of breast cancer can be expected to increase. The treatment of these patients is complicated by many other diseases including cardiovascular and pulmonary disorders associated with aging, and because chemotherapy and radical surgery are often contraindicated. MATERIAL AND METHODS: In an one year period in the Institute of Oncology and Radiology of Serbia a group of 53 elderly (65 years and more) patients with locally advanced breast cancer were treated. Twenty four patients (group A) were treated with hypofractionated (concentrated) radiotherapy. The irradiation was delivered to the breast with TD24-26 Gy with two tangentional portals and 19 Gy to regional lymphatics with anterior fields owner 8 fractions, breast and lymphatics alternatively. The same treatment plan was repeated after 28 days (split course). Co60 was used. Twenty nine patients (group B) were treated with conventional fractionated radiotherapy. Irradiation was delivered to the breast with 51 Gy tumour dose in 16 fractions and to the lymphatics with 45 Gy in 15 fractions. Breast and lymphatics were irradiated alternatively, during 31 working days. After 51 Gy the whole breast was boosted with 20 Gy tumour dose and axilla with TD 12 Gy. The concentrated radiotherapy is, in fact, an alternative for radical--conventional or protracted radiotherapy according to the so-called hypofractionated split course technique. Both techniques have very similar TDF factors. The aim of such a plan is the achievement of adequate tumour dose adapted to the age of patients (the patients should be treated in a smaller number of fractions). All patients were aged 65 years or were older. The median age in group A was 72 years and in group B 68 years. Also in all patients breast cancer was locally advanced (stadium III). In group A median follow-up was 29.79 months and in group B 23.62 months. RESULTS: All patients had acute skin reactions. In group A (irradiated with concentrated technique) 91.7% of patients had erythema, 8.3% dry desquamation, but moist desquamation was not observed. In group B (irradiated with conventional technique) 27.6% of patients had erythema, 55.2% dry desquamation and 17.2% moist desquamation. Delayed radiation changes manifested as fibrosis of the breast and region of axilla were noted in 29.24% of patients in group A and 13.8% in group B. The relapse in group A was 41.7% with median relapse free interval of 13.9 months and in group B 48.2% with relapse free interval of 15.6 months. There was no significant statistical difference between the two groups according to standard statistical methods (chi 2 = 0.96; DF = 3; p > 0.05). After approximately 30 months of follow-up, 50% of patients in group A are alive without signs of disease; 16.7% are alive with disease, and 16.7% are dead due to primary disease. In group B 24.1% of patients are alive without signs of disease; 24.1% are alive with disease; and 20.7% are dead due to primary disease. There was no significant statistical difference between the two groups (chi 2 = 4.09; DF = 4; p > 0.05). The overall survival rate in group A was 67% after 4 years and 53% in group B. Relapse free survival was 53% in group A after 4 years and 36% in group B. In conclusion, according to our study there was no statistically significant difference in local control between conventional and hypofractionated radiotherapy in the treatment in elderly patients. The main advantage of concentrated schedule is shortening of duration of irradiation, but the main disadvantage is a high incidence of fibrosis which makes difficult the evaluation of local control. Consensus about treatment of breast cancer in elderly women has not yet been clearly established. Our data suggest that hypofractionated schedule is an effective, suitable and comfortable therapeutic approach in the management of breast cancer in elderly women. PMID- 11255685 TI - [Potentiation of metaphit-induced audiogenic epilepsy with N-methyl-D-aspartate in rats]. AB - INTRODUCTION: Audiogenic seizures (AGS) are induced by high intensity sound stimulation in genetically susceptible rats or in animals subjected to chemical or electrical manipulation. Epileptic seizure may result from an impaired balance between excitation and inhibition in the CNS. The effect of NMDA (N-methyl-D aspartic acid) on metaphit 1-(1(3-isothiocyanatophenyl-ciclohexyl)-piperidine) induced audiogenic seizures was evaluated in rats. METHODS: Male Wistar albino rats were divided into 4 groups: 1) saline; 2) metaphit (10 mg/kg); 3) metaphit + NMDA; 4) NMDA (70 mg/kg). Animals were injected with metaphit intraperitoneally (i.p.) and exposed to sound stimulation (100 +/- 3 dB, 60 s) at hourly intervals. The incidence and severity (running, clonus and tonus) of seizures were analyzed. NMDA alone was administered i.p. to 6 rats. In group metaphit + NMDA only animals which did not exhibit any seizure during 8 hours were injected with NMDA i.p. after the 8th audiogenic testing. For electroencephalograph (EEG) recordings three gold-plated screws were used. Convulsive behaviour was assessed by incidence of motor seizure and by seizure severity grade, determined by use of a descriptive rating scale with range of 0-3; 0-no response; 1-wild running only; 2 wild running followed by clonic seizures of all four limbs with body rollover; 3 wild running progressing to generalized clonic convulsions and then a tonic extension of the fore and hind limbs and tail. Sound onset, seizure events, and sound offset, along with the animals behaviour (convulsive or other) were recorded as the correlates to the respective EEG responses. RESULTS: In most animals the administration of metaphit (10 mg/kg) resulted in electrographic abnormalities, elicited epileptiform activity in the form of spikes, polyspikes and spikewave complexes (Fig. 1.). Maximum incidence and severity of metaphit convulsions occurred 8 h after the injection (9/12, 75%) (Fig. 2, 3.), then abated gradually and disappeared 30 h later. NMDA (70 mg/kg) alone induced no seizure response but isolated spiking activity, and sporadic slow-wave complexes were recorded (Fig. 4). NMDA induced stereotyped behaviour in the form of asymmetric posture, loss of righting reflex and tonic hindlimb extension, which lasted for 60-90 min. Subconvulsive dose of NMDA potentiated the metaphit-induced audiogenic seizures in rats. Two hours after NMDA administration 3 of 17 metaphit treated rats convulsed, which in 8 previous testings never displayed seizures. Maximum incidence was 8 of 17 (53%), 5-6 h after NMDA administration and seizures lasted for 9 hours. DISCUSSION: Several authors reported that metaphit dose of 10 mg/kg accompanied by some REM sleep deprivation (REM-D) procedures [4], or subconvulsive doses of NMDA [25] provoked seizures of higher intensity and incidence. Metaphit treatment (10 mg/kg) followed 24 h later by NMDA dose of 50 mg/kg provoked no spontaneous convulsions, while metaphit in combination with a higher NMDA dose of 70 mg/kg resulted in spontaneous and AGS-induced seizures only in one time point [25]. It was found that the incidence and severity of convulsive responses were highest 8-12 h after metaphit injection (10 mg/kg) [23, 24]. Although about 8 h after metaphit administration the power spectra increased and were more intense in the period of sound onset and seizure events. CONCLUSION: The results of the present study strongly suggest that treatment of adult rats with the combination of metaphit and NMDA in the doses employed here followed by AGS provides a suitable animal model for examinations of epileptic seizures. PMID- 11255687 TI - [Angioedema caused by angiotensin-converting enzyme inhibitors]. AB - Angioedema is a rare, but important effect of arterial hypertension treatment with drugs which inhibit angiotensin-converting enzyme. Usually, it develops in the first week of therapy, but some atypical cases of the development of angioedema after several months to few years after the onset of the therapy have been reported. Undesired reactions caused by these drugs are probably not allergic, but they are caused by pharmacological effect of these drugs in persons with risk of allergic reaction. In this paper we present some patients with angioedema. PMID- 11255688 TI - [Hamartoma of the spleen]. AB - Hamartoma is a rare benign lesion of the spleen. Between 140 and 150 cases seem to have been described so far. Hamartoma of the spleen may appear as a single or multiple lesions which may tend to converge. It appears in all ages, mainly in elderly persons. About 20% of patients were described in paediatric subjects. Half of the patients have no symptoms, so that hamartomas were discovered by chance at autopsy. Other 50% of patients had pain, splenomegaly, haematologic abnormalities (most frequently thrombocytopenia or pancytopenia) and spontaneous rupture with intra-abdominal bleeding. In children, hamartoma of the spleen with haematologic abnormalities may be followed by growth retardation, frequent infections, fever and night sweating. The bigger the hamartoma the greater probability to cause symptoms. The exact preoperative diagnosis is rarely established. Hamartoma has to be taken into account always when tumour of the spleen is diagnosed, particularly in children. Splenectomy is the most frequent treatment of symptomatic hamartoma of the spleen. Partial splenic resection is the preferred surgery whenever it may be carried out, particularly in children. We report a 58 year old woman with a five-year history of left subcostal and lumbar pain in whom in the lower pole of moderately enlarged spleen a tumorous mass, 107 x 75 mm in diameter, was discovered on ultrasonography. She was submitted to splenectomy as well as to cholecystectomy due to gall bladder stones. Histological findings of the spleen showed hamartoma. She had an uneventful recovery. The pain disappeared after surgery. She stayed symptom free so far. PMID- 11255689 TI - [The role of insulin and hyperinsulinemia in the pathogenesis of the polycystic ovary syndrome]. PMID- 11255690 TI - [Chemotherapy of carcinoma of the urinary bladder]. PMID- 11255691 TI - [Indications for implantable defibrillator cardioverters]. PMID- 11255692 TI - Spontaneous conversion to sinus rhythm of recent (within 24 hours) atrial fibrillation. AB - OBJECTIVES: The purpose of the study was to determine the likelihood of spontaneous conversion of recent onset (< 24 hr) paroxysmal atrial fibrillation (Af) to sinus rhythm and to define clinical and echocardiographic characteristics which may predict it. METHODS: One hundred fifty-three consecutive adult patients admitted to the hospital with recent onset Af (< 24 hr) were studied. In each patient history, complete physical examination, 12-lead electrocardiogram, chest X-ray, routine hematological studies, serum electrolytes, troponin, thyroid function studies and a complete echocardiographic evaluation were performed. Patients hemodynamically unstable, with recent myocardial infarction, unstable angina, average ventricular rate > 150 beats/min, hyperthyroidism, congestive heart failure, left ventricular hypertrophy, valvular heart disease, and on antiarrhythmic drugs at the time of admission, were excluded. Patients were monitored without antiarrhythmic therapy for at least 24 hr from the onset of Af. RESULTS: Spontaneous conversion to sinus rhythm occurred in 109 patients (71.2%); among patients with spontaneous conversion 73.4% converted in the first 12 hr. Age, gender, other clinical characteristics, left ventricular dimensions and performance did not separate patients with or without spontaneous conversion. Left atrial size was significantly greater in patients without compared to patients with spontaneous conversion (p < 0.03); likewise increased left atrial size (> 40 mm) was seen more often in patients without compared to patients with spontaneous conversion (45% vs 22%, p < 0.05). CONCLUSIONS: Spontaneous conversion to sinus rhythm occurred in 71% of patients with recent onset (< 24 hr) Af. Left atrial size was the only predictor of spontaneous conversion in this highly selected group of patients. PMID- 11255693 TI - [Plasma brain natriuretic peptide as a prognostic indicator in patients with primary pulmonary hypertension]. AB - BACKGROUND: Plasma brain natriuretic peptide (BNP) level increases in proportion to the degree of right ventricular dysfunction in pulmonary hypertension. We sought to assess the prognostic significance of plasma BNP in patients with primary pulmonary hypertension. METHODS AND RESULTS: Plasma BNP was measured in 60 patients with primary pulmonary hypertension at diagnostic catheterization, together with atrial natriuretic peptide, norepinephrine, and epinephrine. Measurements were repeated in 53 patients after a mean follow-up period of 3 months. Forty-nine of the patients received intravenous or oral prostacyclin. During a mean follow-up period of 24 months, 18 patients died of cardiopulmonary causes. According to multivariate analysis, baseline plasma BNP was an independent predictor of mortality. Patients with a supramedian level of baseline BNP (> or = 150 pg/ml) had a significantly lower survival rate than those with an inframedian level, according to Kaplan-Meier survival curves (p < 0.05). Plasma BNP in survivors decreased significantly during the follow-up (217 +/- 38 to 149 +/- 30 pg/ml, p < 0.05), whereas that in nonsurvivors increased (365 +/- 77 to 544 +/- 68 pg/ml, p < 0.05). Thus, survival was strikingly worse for patients with a supramedian value of follow-up BNP (> or = 180 pg/ml) than for those with an inframedian value (p < 0.0001). CONCLUSIONS: A high level of plasma BNP, and in particular, a further increase in plasma BNP during follow-up, may have a strong, independent association with increased mortality in patients with primary pulmonary hypertension. PMID- 11255694 TI - [Cardioprotective mechanism of ischemic preconditioning is impaired by postinfarct ventricular remodeling through angiotensin II type 1 receptor activation]. AB - BACKGROUND: Activation of protein kinase C-linked receptors and subsequent opening of the mitochondrial adenosine triphosphate-sensitive K+ (mitoKatp) channel is crucial in preconditioning. This study examined whether post-infarct ventricular remodeling interferes with the preconditioning mechanism. METHODS AND RESULTS: Two weeks before isolation of hearts, rabbits underwent a sham operation or coronary ligation (COL) to induce remodeling. Isolated buffer perfused hearts were subjected to 30-min global ischemia/2-hr reperfusion, and infarct size was expressed as a percentage of the left ventricle (%I/LV), from which the scarred infarct by COL was excluded. Although %I/LV was similar in sham-operated and remodeled hearts (52.9 +/- 6.5% vs 45.8 +/- 5.2%), preconditioning with 2 episodes of 5-min ischemia protected sham-operated but not remodeled hearts (%I/LV = 18.1 +/- 2.5% vs 54.8 + 2.9%, p < 0.05). Infusion of valsartan (10 mg/kg/day; Val), an angiotensin II type 1 receptor blocker, for 2 weeks after COL prevented the ventricular remodeling and preserved the response to preconditioning (%I/LV = 27.4 +/- 3.8%), though Val alone did not change %I/LV. Diazoxide, a mitoKatp channel opener, protected both sham-operated and remodeled hearts (%I/LV = 14.1 +/- 3.1% and 8.3 +/- 3.6%). CONCLUSIONS: The myocardium remodeled after infarction is refractory to preconditioning, which is probably due to interruption of cellular signaling by preconditioning upstream of mitoKatp channels. An angiotensin II type 1 receptor blocker is beneficial not only for suppression of ventricular remodeling but also for preservation of the preconditioning mechanism. PMID- 11255695 TI - [Calcineurin inhibitor attenuates the development and induces the regression of cardiac hypertrophy in rats with salt-sensitive hypertension]. AB - BACKGROUND: It remains unclear how hemodynamic overload induces cardiac hypertrophy. Recently, activation of calcium-dependent phosphatase, calcineurin, has been elucidated to induce cardiac hypertrophy. In the present study, we examined the role of calcineurin in load-induced cardiac hypertrophy by using Dahl salt-sensitive (DS) rats, which develop both pressure and volume overload when fed a high salt diet. METHODS AND RESULTS: In the DS rat heart, the activity of calcineurin was increased and cardiac hypertrophy was induced by high salt diet. Treatment of DS rats with the calcineurin inhibitor FK506 (0.1 or 0.01 mg/kg every second day) from the age of 6 weeks to 12 weeks inhibited the activation of calcineurin in the heart in a dose-dependent manner and attenuated the development of load-induced cardiac hypertrophy and fibrosis without change of hemodynamic parameters. Additionally, treatment with 0.1 mg/kg every second day but not with 0.01 mg/kg every second day of FK506 from the age of 12 weeks to 16 weeks induced regression of cardiac hypertrophy in DS rats. Load-induced reprogramming of gene expression was also suppressed by the FK506 treatment. CONCLUSIONS: These results suggest that calcineurin is involved in the development of cardiac hypertrophy in rats with salt-sensitive hypertension and that inhibition of calcineurin could induce regression of cardiac hypertrophy. PMID- 11255696 TI - [Nitric oxide spares myocardial oxygen consumption through attenuation of contractile response to beta-adrenergic stimulation in patients with idiopathic dilated cardiomyopathy]. AB - BACKGROUND: The results of recent studies suggest that nitric oxide (NO) synthase may increase in the failing myocardium and that NO modulates the myocardial contractile response to beta-adrenergic stimulation. However, there are few data regarding the physiological role of NO in patients with heart failure. The aim of the present study was to address the role of NO in left ventricular contractile response to beta-adrenergic stimulation and corresponding oxygen expenditure in human heart failure. METHODS AND RESULTS: We studied 15 patients with heart failure due to idiopathic dilated cardiomyopathy (mean ejection fraction 0.33). We examined left ventricular contractility (Emax, the slope of end-systolic pressure-volume relation), left ventricular external work (EW), myocardial oxygen consumption (MVO2), and mechanical efficiency (measured as EW/MVO2) with the use of conductance and coronary sinus thermodilution catheters before and during dobutamine (DOB) infusion via a peripheral vein (4.8 +/- 0.3 micrograms.kg-1.min 1 i.v.). Heart rate was kept constant with atrial pacing. We carried out a similar protocol during the intracoronary infusion of the NO synthase inhibitor NG-monomethyl-L-arginine (L-NMMA; 200 mumol). DOB increased Emax, EW, and MVO2 (by 77 +/- 17%, 39 +/- 5%, and 21 +/- 5%, respectively), leading to an increase in mechanical efficiency (25.4 +/- 3.1% to 29.6 +/- 4.1%). L-NMMA alone did not significantly change these variables. Although the concurrent infusion of DOB with L-NMMA increased Emax, EW, and MVO2 (by 140 +/- 21%, 64 +/- 9%, and 35 +/- 5%, respectively) more than DOB alone, mechanical efficiency did not increase further (24.3 +/- 3.3% to 29.5 +/- 4.5%) because EW and MVO2 increased in parallel. CONCLUSIONS: These data suggest that in patients with idiopathic dilated cardiomyopathy, endogenous NO spares MVO2 through attenuation of left ventricular contractile response to beta-adrenergic stimulation while maintaining left ventricular energy-converting efficiency. PMID- 11255697 TI - [A 65-year-old man with syncope]. PMID- 11255699 TI - [Cost analysis of procedural fee for percutaneous transluminal coronary angioplasty at six Japanese hospitals]. AB - BACKGROUND: Disease-specific cost analysis is rarely performed in Japan, but is essential for reform of the healthcare reimbursement system and assessment of procedural fees. METHODS: The actual cost associated with the procedural fee of the percutaneous transluminal coronary angioplasty (PTCA) procedure was estimated by dividing into four categories: labor, disposable materials, expenses, and equipment costs. The special cost associated with PTCA devices such as angioplasty balloon and stent was responsible for the majority of PTCA-related hospitalization costs, but was not included in our survey. The six institutions that participated in the survey belong to the national, Red Cross, and Saiseikai organizations. Time study for labor was omitted, and procedural time was predetermined at 3 hours. RESULTS: The labor cost amounted to yen 65,000 to yen 98,000/procedure. To calculate the cost of disposable materials across all six hospitals, a universal amount of yen 60,000 from a model hospital was used. The expenses ranged from yen 1,000 to yen 39,000, and the expenditure plus capital cost from yen 95,000 to yen 224,000, showing significant differences between the hospitals. The total mean cost was yen 294,000 +/- 55,000, which indicated that every hospital was in deficit. CONCLUSIONS: Compared to Medicare in the United States of America, the procedural fee for Japanese physicians is extremely low, in contrast to the bloated special cost for devices, which causes significant pricing gaps between Japan and USA. The differences in total cost among the hospitals were mainly derived from the cost for angiographic equipment. Our survey did not include any private hospitals, but the PTCA-related procedural fee is less than the actual cost under the current health insurance reimbursement scheme at all hospitals. PMID- 11255700 TI - [Broad old anteroseptal myocardial infarction with coronary aneurysm showing marked improvement of myocardial perfusion by intravenous myocardial contrast echocardiography after successful coronary angioplasty: a case report]. AB - A 56-year-old man with old anteroseptal myocardial infarction was admitted to treat a gradually expanding coronary aneurysm. Coronary angiography demonstrated the aneurysm and delayed flow in the left coronary descending artery. The aneurysm was successfully treated with stenting. Technetium-99m-sestamibi single photon emission computed tomography (SPECT) showed a persistent defect in the anterior infarcted area after stenting. Myocardial contrast echocardiography was performed using SystemFIVE and EchoPAC. Levovist was injected (1 ml, 1.5 ml/sec; bolus + 1 ml/sec; continuous infusion) by Pulsar. Myocardial reperfusion was observed by B-mode and anatomical M-mode before and after intervention. Time intensity curves from the region of interest positioned within the interventricular septum showed the mean value at plateau increased from 10.8 to 25.1 dB. The persistent defect area demonstrated by SPECT was enhanced by myocardial contrast echocardiography after intervention. Myocardial contrast echocardiography is useful for the assessment of outcome after intervention and evaluation of improved coronary blood flow. PMID- 11255701 TI - [Cholesterol embolism after percutaneous transluminal coronary angioplasty procedure: a case report]. AB - A 68-year-old man with chest pain was treated under a diagnosis of angina pectoris based on coronary angiography by percutaneous transluminal coronary angioplasty including stent implantation performed by the femoral approach. About 1 month after intervention, his renal function deteriorated and purpura appeared on both feet, especially at the toe tips. He was treated under a tentative diagnosis of cholesterol embolism conservatively at the out-patient clinic. However, he was admitted by ambulance due to worsening renal failure 2 months later and died from multiple organ failure 2 weeks after admission. Autopsy examination identified cholesterol embolism due to crystal emboli in several organs. Cholesterol embolism rarely occurs after angiographic or interventional procedures, but is difficult to diagnose clinically and there is no established therapy. The number of intervention and angiography procedures is increasing, so the occurrence of embolism as a complication of these procedures might also increase. PMID- 11255698 TI - [Efficacy of cibenzoline in the termination and long-term prevention of paroxysmal atrial fibrillation: analysis based on the time of onset]. AB - OBJECTIVES: The efficacy of cibenzoline was assessed in the termination and long term prevention of paroxysmal atrial fibrillation in relation to the time of onset in a series of patients with paroxysmal atrial fibrillation. METHODS: Study of the termination included 92 patients (63 males, 29 females, mean age 64 +/- 12 years) and study of long-term prevention included 106 patients (77 males, 29 females, mean age 64 +/- 11 years; mean follow-up 32.7 +/- 18.8 months). Paroxysmal atrial fibrillation was classified into 3 types based on the time of onset: day type (AM 7:00-PM 5:00), night type (PM 5:00-AM 7:00) and mixed type (anytime). Successful termination was defined as pharmacological cardioversion within 30 min of the intravenous administration of 70 mg cibenzoline and efficacy of long-term prevention was presented as the event-free ratio of patients after oral administration of 300 mg/day cibenzoline. RESULTS: Successful pharmacological termination was achieved in 66.7% of the day type (n = 24), 70.0% of the night type (n = 48), and 41.6% of the mixed type (n = 20). There was a significantly higher success in the day type (p < 0.05), and tendency to success in the night type (p = 0.079) compared to the mixed type. The event-free ratios at 1, 3, 6, 12, 24 months after oral administration were 84.6%, 76.9%, 73.1%, 65.4%, 61.5% in the day type (n = 26), 92.0%, 80.0%, 80.0%, 72.0%, 60.0% in the night type (n = 25), and 81.8%, 61.8%, 47.3%, 30.9%, 23.6% in the mixed type (n = 55), respectively. Significantly higher success was achieved at 24 months in the day type and the night type compared to the mixed type (p < 0.05). CONCLUSIONS: Termination and long-term prevention of paroxysmal atrial fibrillation by cibenzoline has a high degree of efficacy in patients with both day and night onset of paroxysmal atrial fibrillation. PMID- 11255702 TI - Use of computer technology to help students with special needs. AB - Millions of students across the United States cannot benefit fully from a traditional educational program because they have a disability that impairs their ability to participate in a typical classroom environment. For these students, computer-based technologies can play an especially important role. Not only can computer technology facilitate a broader range of educational activities to meet a variety of needs for students with mild learning disorders, but adaptive technology now exists than can enable even those students with severe disabilities to become active learners in the classroom alongside their peers who do not have disabilities. This article provides an overview of the role computer technology can play in promoting the education of children with special needs within the regular classroom. For example, use of computer technology for word processing, communication, research, and multimedia projects can help the three million students with specific learning and emotional disorders keep up with their nondisabled peers. Computer technology has also enhanced the development of sophisticated devices that can assist the two million students with more severe disabilities in overcoming a wide range of limitations that hinder classroom participation--from speech and hearing impairments to blindness and severe physical disabilities. However, many teachers are not adequately trained on how to use technology effectively in their classrooms, and the cost of the technology is a serious consideration for all schools. Thus, although computer technology has the potential to act as an equalizer by freeing many students from their disabilities, the barriers of inadequate training and cost must first be overcome before more widespread use can become a reality. PMID- 11255703 TI - The impact of home computer use on children's activities and development. AB - The increasing amount of time children are spending on computers at home and school has raised questions about how the use of computer technology may make a difference in their lives--from helping with homework to causing depression to encouraging violent behavior. This article provides an overview of the limited research on the effects of home computer use on children's physical, cognitive, and social development. Initial research suggests, for example, that access to computers increases the total amount of time children spend in front of a television or computer screen at the expense of other activities, thereby putting them at risk for obesity. At the same time, cognitive research suggests that playing computer games can be an important building block to computer literacy because it enhances children's ability to read and visualize images in three dimensional space and track multiple images simultaneously. The limited evidence available also indicates that home computer use is linked to slightly better academic performance. The research findings are more mixed, however, regarding the effects on children's social development. Although little evidence indicates that the moderate use of computers to play games has a negative impact on children's friendships and family relationships, recent survey data show that increased use of the Internet may be linked to increases in loneliness and depression. Of most concern are the findings that playing violent computer games may increase aggressiveness and desensitize a child to suffering, and that the use of computers may blur a child's ability to distinguish real life from simulation. The authors conclude that more systematic research is needed in these areas to help parents and policymakers maximize the positive effects and to minimize the negative effects of home computers in children's lives. PMID- 11255705 TI - Federal programs to increase children's access to educational technology. PMID- 11255704 TI - Children's media culture in the new millennium: mapping the digital landscape. AB - A new "children's digital media culture" is swiftly moving into place on the Internet. In this article, the author describes the technological, demographic, and market forces shaping this new digital media culture and the rich array of Web sites being created for children and teens. Many nonprofit organizations, museums, educational institutions, and government agencies are playing a significant role in developing online content for children, offering them opportunities to explore the world, form communities with other children, and create their own works of art and literature. For the most part, however, the heavily promoted commercial sites, sponsored mainly by media conglomerates and toy companies, are overshadowing the educational sites. Because of the unique interactive features of the Internet, companies are able to integrate advertising and Web site content to promote "brand awareness" and "brand loyalty" among children, encouraging them to become consumers beginning at a very early age. The possibility that a child's exploration on the Internet might lead to inappropriate content, aggressive advertising, or even dangerous contact with strangers has given rise to a number of efforts to create "safe zones" for children--that is, places in cyberspace where children can be protected from both marketers and predators. Federal legislation now requires parental permission before commercial Web sites can collect personal information from children under age 13. Several companies offer filtering, blocking, and monitoring software to safeguard children from harmful content or predators. Generally lacking in debates concerning children's use of the Internet, however, is a more proactive definition of quality--one that would help ensure the creation and maintenance of Web sites that enhance children's learning and development and not merely keep them from harm. In the concluding section of this article, the author recommends actions to promote development of a quality media culture that would help children become good citizens as well as responsible consumers. PMID- 11255708 TI - Children and computer technology: analysis and recommendations. PMID- 11255706 TI - What children think about computers. PMID- 11255707 TI - Children and computers: new technology--old concerns. AB - Computer technology has ushered in a new era of mass media, bringing with it great promise and great concerns about the effect on children's development and well-being. Although we tend to see these issues as being new, similar promises and concerns have accompanied each new wave of media technology throughout the past century: films in the early 1900s, radio in the 1920s, and television in the 1940s. With the introduction of each of these technologies, proponents touted the educational benefits for children, while opponents voiced fears about exposure to inappropriate commercial, sexual, and violent content. This article places current studies on children and computers in a historical context, noting the recurrent themes and patterns in media research during the twentieth century. Initial research concerning each innovation has tended to focus on issues of access and the amount of time children were spending with the new medium. As use of the technology became more prevalent, research shifted to issues related to content and its effects on children. Current research on children's use of computers is again following this pattern. But the increased level of interactivity now possible with computer games and with the communication features of the Internet has heightened both the promise of greatly enriched learning and the concerns related to increased risk of harm. As a result, research on the effects of exposure to various types of content has taken on a new sense of urgency. The authors conclude that to help inform and sustain the creation of more quality content for children, further research is needed on the effects of media on children, and new partnerships must be forged between industry, academia, and advocacy groups. PMID- 11255709 TI - Who's wired and who's not: children's access to and use of computer technology. AB - As computer technology becomes increasingly prevalent throughout society, concerns have been raised about an emerging "digital divide" between those children who are benefitting and those who are being left behind. This article presents results from new analyses of national survey data describing children's differential access to computers in school and at home, and the varying conditions that affect how children experience computers. For example, responses from a nationwide survey of teachers suggest that, as of 1998, more than 75% of students had access to computers at school. In fact, those teaching lower-income students reported weekly use of computers more often than those teaching higher income students. But the nature of children's experiences using computers in school varied greatly by subject and teacher objectives, and the data suggest that lower-income students use computers more often for repetitive practice, whereas higher-income students use computers more often for more sophisticated, intellectually complex applications. Differences between low-income and high income children's access to home computers were far less subtle. Survey data indicate that only about 22% of children in families with annual incomes of less than $20,000 had access to a home computer, compared to 91% of those in families with annual incomes of more than $75,000. And among children with access, those in low-income families were reported to use the computer less than those in high income families, perhaps because most low-income families with computers lacked a connection to the Internet. The two most predictive factors of children's use of home computers were the child's age and the computer's capabilities. The author concludes that home access to computers will be a continued area of inequality in American society, and that schools must play a critical role in ensuring equal opportunity for less-advantaged children to access the benefits of the more intellectually powerful uses of computer technology. PMID- 11255710 TI - Changing how and what children learn in school with computer-based technologies. AB - Schools today face ever-increasing demands in their attempts to ensure that students are well equipped to enter the workforce and navigate a complex world. Research indicates that computer technology can help support learning, and that it is especially useful in developing the higher-order skills of critical thinking, analysis, and scientific inquiry. But the mere presence of computers in the classroom does not ensure their effective use. Some computer applications have been shown to be more successful than others, and many factors influence how well even the most promising applications are implemented. This article explores the various ways computer technology can be used to improve how and what children learn in the classroom. Several examples of computer-based applications are highlighted to illustrate ways technology can enhance how children learn by supporting four fundamental characteristics of learning: (1) active engagement, (2) participation in groups, (3) frequent interaction and feedback, and (4) connections to real-world contexts. Additional examples illustrate ways technology can expand what children learn by helping them to understand core concepts in subjects like math, science, and literacy. Research indicates, however, that the use of technology as an effective learning tool is more likely to take place when embedded in a broader education reform movement that includes improvements in teacher training, curriculum, student assessment, and a school's capacity for change. To help inform decisions about the future role of computers in the classroom, the authors conclude that further research is needed to identify the uses that most effectively support learning and the conditions required for successful implementation. PMID- 11255711 TI - Children and computer technology. A selected bibliography. PMID- 11255712 TI - [Evidence-based medicine. Science, paradigm or methodology?]. PMID- 11255713 TI - [Absence of right and left atrioventricular connexion]. AB - Fifty seven hearts with absence of atrioventricular (A-V) connection were studied morphologically to specify their types of ventriculoarterial connection and their associated anomalies; the anatomic features of the hearts were correlated with their echocardiographic and cardioangiographic images in order to establish their mutual correspondence. Fifty six hearts had situs solitus; fifty specimens had right absent A-V connection and six had left absent A-V connection. One had situs inversus. All the specimens had: A deep A-V sulcus at the site of the absent A-V connection, a dimple in the muscular floor of the involved atrium connected with the dilated and hypertrophic left ventricle, incomplete right ventricle without inlet portion, ventricular septal defect of variable dimensions (it was obliterated in two), atrial septal defect, the ventricular septum deviated from the crux cordis. The left absent A-V connection had ventricular inversion and discordant ventriculoarterial connection. In the right absent A-V connection the ventriculoarterial connections were concordant in thirty eight hearts, from which thirty four had pulmonary stenosis both infundibular and valvular (five had the tetrad of Fallot), two had pulmonary valve atresia and two had a dilated pulmonary artery; discordant in nine hearts, one with aortic atresia; double outlet, from the right ventricle in two, (one with the tetrad of Fallot) and from the left ventricle in one. The heart in situs inversus had ventricular inversion, right absent A-V connection (left-sided), single (right) ventricle and atresia of the left ventricle. The correlations between cardiac morphology and imaging were precise. Developmentally, this cardiopathy is the result of an ectopic unequally lateralized septation of the common atrioventricular canal, which separates two canals, one stenotic leading to atresia and the other which develops too wide. PMID- 11255714 TI - [Effect of cardiac rehabilitation in ischemic patients not undergoing coronary revascularization]. AB - Cardiac rehabilitation program was implemented on 65 patients rejected for bypass surgery and/or coronary angioplasty. The group was formed by 63 males (96.9%), 2 females (3.0). Their ages ranged from 54.5 +/- 8.7 year (mean +/- SD). Their risk's factors, were (RF): cigarette smoking, 25 patients (38.6%), hypercholesterolemia 47 (72.3%), hypertriglyceridemia 48 (73.8%), hypertension 33 (50.7%), diabetes mellitus 33 (46.1%), myocardial infarction 56 (86.1%). Coronary arteriography showed one vessel disease (25.5%), two vessels (38.3%) and three (36.1%). Ejection fraction (EF) > 50% in 39 (60%). The features evaluated were: number of obstructed vessels, (EF) and age. RESULTS: One patient died, fifty two (80%) were asymptomatic ten showed stable angina (15.3%) and three (4.6%) unstable angina. Tobacco addiction decreased by 92%. Hypertension was controlled in 42.2%. Hypercholesterolemia was normalized in 82.9%. Hypertriglyceridemia improved in 53%. Oxygen uptake VO2 and Oxygen pulse PO2 improved in 42 patients (64.6%). Cardiac rehabilitation is a good alternative to improve quality of life in patients in whom a coronary revascularization is not feasible. PMID- 11255716 TI - [Computerized helicoidal tomography of the coronary arteries vs coronary angiography]. AB - INTRODUCTION: The calcium score (CS) of the coronary arteries by computed tomography (CT) is an useful procedure for the diagnosis of obstructive coronary disease (OCD), with an average sensitivity of 82 +/- 6%, specificity of 88 +/- 2%, positive predictive value (PPV) of 57 +/- 7% and negative predictive value (NPV) of 96 +/- 2%. The objective of this trial was to compare helicoidal CT Scan with the traditional method and define sensitivity, specificity, Positive predictive value and negative predictive value against the coronary angiography. METHODS: From June of 1998 to March of 1999, one hundred and sixty six patients with coronary arteries CT were studied. The CT was done with an ELSCINT-CT Twin equipment and a software for the quantification of the coronary arteries CS in Hounsfield units. In forty one, coronary angiography was performed. A significant obstructive lesion was defined as > or = 70% of luminal stenosis in at least one artery, or > or = 50% in the left main and > or = 50% if some other artery was involved. This group was divided in accordance to the CS in two subgroups: A with a CS < or = 150 and B those with a CS > or = 151. RESULTS: In group A, 45% had significative lesions vs 95% in group B (p = 0.001). The sensitivity was 65%, specificity 95%, PPV 64% and the NPV 92%. Relative risk 2.08 (CI 95% 1.38-3.54) and Odds ratio 21.6 (CI 95% 2.43-191.37). CONCLUSIONS: Even though the small sample, CT is an useful procedure for the diagnosis of the OCD. PMID- 11255717 TI - [Non-invasive estimation of dP/dt of the left ventricle: is the measurement taken during the isovolumetric contraction?]. AB - Left ventricular dP/dt is estimated from mitral regurgitation (MR) jet as the rate of pressure rise (RPR) from 1 to 3 m/sec. In order to establish if this measure is made during the isovolumetric contraction (IC), we with MR studied 38 patients (age average 51 +/- 8 years) of different etiology. IC was estimated as pre-ejection time minus Q-first sound (S1). Velocity of the MR was measured at the onset and at the end of IC to estimate RPR during IC and time from 1 m/s to S1 (T1-S1) to indicate the mismatch between the two methods. RESULTS: There was not difference between RPR 1 to 3 m/s and RPR (IC). T 1-S1 was 26 +/- 24 ms indicating that the measure of RPR 1 to 3 m/s was made prior to the onset of IC. CONCLUSION: Noninvasive assessment of left ventricular dP/dt from 1 to 3 m/s is made prior to the onset of IC. PMID- 11255715 TI - [Atrioventricular discordance. Clinico-surgical experience 1990-2000]. AB - OBJECTIVE: To know the incidence of ventriculoarterial connections combined with atrioventricular discordance, associated lesions and surgical results, including the first case with anatomical correction. METHOD: All patients with atrioventricular discordance by echocardiography from 1990 to March 2000 were analyzed. RESULTS: Thirty six patients with atrioventricular discordance were found. Ages ranged from 0.1 to 46 years, with a mean 9.2 years (SD 5.9 years). Atrial chambers were situs solitus in 88.9%, inversus in 11.2%. The ventriculoarterial connections were discordant in 28 (77.7%), double outlet right ventricle in 4 (11.1%) (one of them was a "criss cross" heart), single outlet (pulmonary atresia) in 4 (11.1%), and double outlet left ventricle in 1 (2.7%). Associated lesions: Ventricular septal defect with pulmonary stenosis or atresia was present in 21 (58%), ventricular septal defect with no pulmonary obstruction was observed in 10 (28%). Five had tricuspid regurgitation with right ventricular disfunction, (two adults). Surgical results: 22 (61%) required 28 surgical procedures: 8 (36%) were palliative and 19 (86%) were corrective, one of them was our first anatomical correction. Operative mortality in all was 40.1%, postoperative atrioventricular block was observed in 9 (40.1%). CONCLUSION: Right ventricular dysfunction is not uncommon. Surgical results revealed high mortality and high pacemaker requirement for atrioventricular block. PMID- 11255719 TI - Aortic origin of the right pulmonary artery associated with ductus arteriosus in an adult. A case report. AB - We describe the case of a 26-year-old female in functional class I (NYHA), with aortic origin of the right pulmonary artery associated with a persistent ductus arteriosus and severe pulmonary artery hypertension (101/40-70 mm Hg), which remained elevated (89/40-60 mm Hg) after the administration of 100% oxygen. Right pulmonary artery pressure (125/60-86 mm Hg) was higher than that of main pulmonary artery and similar to aorta pressure. The patient was successfully treated: surgical closure of the ductus arteriosus and end-to-end anastomosis between the pulmonary artery and right pulmonary artery were carried out. Systolic pulmonary arterial pressure, estimated by echocardiography Doppler, was 60 mm Hg six months after surgery. Cross-sectional echocardiogram showed the anastomosis of the right pulmonary artery with the main pulmonary artery. Pulmonary gammagraphy showed both lungs perfused through the main pulmonary artery; right lung perfusion was lesser than left lung perfusion, 30 vs. 70% respectively. Aortic origin of a right or left pulmonary artery is a heart disease seen in patients during the course of the first year of life. Its frequency is < 1% among all the congenital cardiopathies and the survival rate to adult life is very low. The originality of this paper is the presentation of a rare congenital cardiopathy treated surgically in an adult. PMID- 11255718 TI - [Comparative study between bisoprolol and metoprolol, combined with hydrochlorothiazide, as antihypertensive therapy]. AB - The main objective of this research was to compare the efficacy and security of bisoprolol (B), a new cardioselective beta-blocker, that does not have intrinsic sympathomimetic activity, and metoprolol associated to hydrochlorothiazide (HCTZ), in the treatment of patients with mild to moderate hypertension. Sixty two hypertensive patients (47 females and 15 males) aged 20 to 70 years (mean 52.5 +/- 10.4) were included in a double-blind, placebo controlled and randomized clinical trial. After a two-weeks wash out period and a similar placebo phase, patients were randomly assigned to receive either a once-daily dosing of B (10 mg) with 6.25 mg of HCTZ, or M (100 mg) plus 6.25 mg of HCTZ during four-weeks. If there was no reduction below 90 mmHg at the end of this period, the dosing of either beta-blocker was doubled. After eight weeks of treatment, the mean decreases in systolic/diastolic blood pressures from baseline were 31.8/21.2 and 28.0/20.6 mmHg for B/HCTZ and M/HCTZ, respectively (p < 0.0001). There were no clinically significant changes from baseline in laboratory parameters in either group. Reduction in blood pressure with B/HCTZ is associated with adverse events and metabolic changes similar to those observed with other antihypertensive drugs. PMID- 11255720 TI - [Ergotism caused by automedication]. AB - The authors report 7 cases of gangrenous ergotism (six women and one man) secondary to an overdose of ergotamine ingested in order to relieve migraine crisis. In all cases, patients presented symptoms and signs of severe arterial constriction confirmed by echography and angiography. Hallucinations were absent. Ergotamine ingestion was discontinued and treatment was based on vasodilators and sympathectomy. After treatment, all seven patients showed clinical improvement with disappearance of the vasospastic symptoms and signs, and an increase in the plethysmographic index of blood perfusion, measured by Doppler echography. These changes were observed even in a patient who lost two toes of the right foot. Although, none of the patients presented hallucinations, the authors made reference to the historical first use of the ergot in magic and religious rites that took place in Eleusis, at the time of classic Greece, as well as the more recent mystic use of ergot in Salem, New England, in 1692. Migraine is indeed a serious disease, frequently causing despair to the patient, who attempts to alleviate the migraneous crisis with an overdose of ergotamine. Accordingly, physicians must be aware of prophylactic vasodilating drugs, reducing the risk of ergotism. PMID- 11255721 TI - [Metabolic therapy with glucose-insulin-potassium. Historical outline]. PMID- 11255722 TI - [Nuclear cardiology and its application in the study of the patient undergoing cardiac transplant]. PMID- 11255724 TI - A new frontier: applying comprehensive DM strategies to healthy members. AB - Extend disease management services to 'healthy' members. Why? Because with all the emphasis on high-cost, chronically ill patients, a health plan's most valuable asset is being ignored. At least that's the case being made by Nashville, TN-based American Healthways, which has rolled out a new program designed to put health plans in touch with all their members, not just the chronically ill. Through the use of predictive modeling, the program is designed to target higher risk members so preventive strategies can be employed. PMID- 11255723 TI - DM programs bring relief to migraine sufferers. PMID- 11255725 TI - Physician communication skills essential to an effective preventive care strategy. AB - Preventive services: Low-tech communications strategies needed to motivate patients. New-age Internet technologies can help, but the key to getting patients to take advantage of preventive services and screenings is physician advice. And the mastery of certain skills and strategies can give that advice much more mileage. Check out these proven approaches to more effective doctor-patient communications, and find out how health systems can complement the process. PMID- 11255726 TI - Exercise can be a powerful weapon against osteoporosis. AB - Are your exercise recommendations for osteoporosis patients off-base? At least one expert believes that the vast majority of clinicians are giving bad advice with regards to exercise, noting that the typical recommendations, focusing on weight-bearing exercise, are simply unrealistic for most people at risk for the bone-thinning disease. Instead, why not consider a more tolerable and less time consuming program of site-specific resistive exercise? It may not have as much impact on bone formation, but research suggests a modest program can provide valuable fracture-prevention benefits. PMID- 11255727 TI - Mutations of retinoblastoma gene (Rb-1) as a prognostic factor in children with acute leukemia and neuroblastoma. AB - Rb-1 is a tumor suppressor gene encoding for a nuclear phosphoprotein acting as a cell cycle regulator, normally expressed in hematopoietic cells and more often inactivated by point mutations with predominance for exons 20-24. The aim of this study is to correlate the retinoblastoma-1 (Rb-1) gene mutations with the prognosis and progression of childhood acute leukemia and neuroblastoma. Bone marrow slides from 26 children with leukemia (18 acute lymphoblastic leukemia [ALL] and 8 acute myeloid leukemia [AML]) and 4 children with neuroblastoma were studied. Exons 20, 21, and 22 were amplified using the polymerase chain reaction technique. Single strand conformational polymorphism (SSCP) and heterodoublex analysis were performed to detect mutations. In ALL cases, two samples in exon 20 (11.11%), one in exon 21 (5.56%), and four in exon 22 (22.22%) had altered conformation. All but one of these cases were classified as high-risk leukemia patients who either relapsed or never achieved remission. Two of the AML cases who did not achieve remission and one of the neuroblastoma cases with concomitant bone marrow infiltration had altered conformation as well. The SSCP and heterodoublex analysis showed that all but one who did not belong to the high risk group had the same altered conformation. These data suggest that Rb-1 gene could possibly be used as an independent prognostic factor for the acute leukemia of childhood and result in the intensification of chemotherapy. In solid tumors with bone marrow involvement it could play a role as a marker of aggressive disease. PMID- 11255728 TI - Evaluation of osteoporosis in thalassemia by quantitative computed tomography: is it reliable? AB - This study was conducted to quantify the degree of osteoporosis in thalassemic patients by single energy quantitative computed tomography (SEQCT) and to test the reliability of this method. On 38 thalassemic patients with osteoporosis and 38 normal control subjects, bone mineral density (BMD) measurements were done by SEQCT. BMD and standard deviation (SD) of the x-ray attenuation numbers of pixels within region of interest (ROI) of the measurement areas were compared between two groups. Mean BMD values for thalassemic patients and control group were 173.4 and 158.2 mg/cm3, respectively. Mean BMD value for thalassemic patient group was significantly higher. Mean SD values of ROI for control group and thalassemic patients were 41.4 and 71.1, respectively. The difference between the SD values was also statistically significant. Positive correlation was noted between SD values and patients' ages in the thalassemic group. Results of SEQCT method may not reflect the clinical and conventional radiographic findings of osteoporosis seen in thalassemic patient group and should be used cautiously. Other methods of BMD measurement, such as photon absorbsiometry and x-ray absorbsiometry, should also be investigated for their accuracy in this patient group. PMID- 11255729 TI - Mutism after surgical removal of a cerebellar tumor: two case reports. AB - The authors report two pediatric cases of transient mutism that occurred after surgical removal of a medulloblastoma and a pilocytic astrocytoma of the vermis and discuss the pathophysiology of this syndrome. Transient mutism has been described for the first time quite recently, even in cases where these tumors were also surgically removed before. Perhaps improvement in imaging and in surgical techniques made neurosurgeons more daring and some interventions that were judged impossible are routinely performed today. If this is the case, postoperative transient cerebellar mutism might be considered the price that must be paid in order to cure more patients with cerebellar tumors. PMID- 11255731 TI - Simultaneous occurrence of advanced neuroblastoma and acute myeloid leukemia. AB - The authors report the case of a 4-year-old boy with a diagnosis of stage IV neuroblastoma (NB), who had been treated with 6 cycles of cyclophosphamide, doxorubicin, cisplatin, and etoposide for 12 months. The patient reached partial remission and presented a diagnosis of acute myelomonocytic leukemia (M4 AML), confirmed by immunophenotyping. After 2 months of therapy for leukemia, the child died with both malignancies in activity. A necropsy histologically confirmed the simultaneity of the two diseases. The authors review the possibilities of this association. The review leads to the conclusion that AML can occur as a secondary malignancy after the onset of the neuroblastoma, or be suggested by a misdiagnosis. The simultaneous occurrence of both as described here is not, however, found in the literature, to the best of the authors' knowledge. PMID- 11255732 TI - Itraconazole-enhanced vincristine neurotoxicity in a child with acute lymphoblastic leukemia. AB - A boy with acute lymphoblastic leukemia (ALL) experienced life-threatening vincristine neurotoxicity while simultaneously exposed to itraconazole. Five pediatric and six adult cases of itraconazole-enhanced vincristine toxicity have been reported, all with ALL. Upon cessation of the itraconazole, the patient's symptoms resolved, which is similar to the outcome of the previously reported cases: 10 of 11 patients had complete resolution of symptoms. PMID- 11255730 TI - Metastatic osteosarcoma to the liver after treatment for synovial sarcoma: a case report. AB - Metastatic osteosarcoma most commonly affects the lungs and other bones. Hepatic metastasis at the time of diagnosis is extremely rare. A 14-year-old boy with synovial sarcoma of the left popliteal fossa was treated with surgical resection, radiotherapy for microscopic residual disease, and 1 year of chemotherapy (vincristine, cyclophosphamide, dactinomycin, and doxorubicin). Approximately 10 years after the initial diagnosis, a secondary osteosarcoma developed in the left proximal tibia. Computed tomography at presentation showed bilateral pulmonary metastases and large ossified nodules in the liver that demonstrated abnormal avidity on 99mTc MDP bone scan indicating hepatic metastasis. Despite chemotherapy (cisplatin, ifosfamide, high-dose methotrexate, and dacarbazine), the patient died of progressive disease 4 months after the diagnosis of the second cancer. Hepatic metastasis was found at the time of diagnosis of a secondary osteosarcoma and manifested as ossified nodules. The risk of radiation induced osteosarcoma should always be considered in decisions about treatment for soft-tissue sarcoma. PMID- 11255733 TI - Ki-1+ anaplastic large cell lymphoma in a child with unpredictable clinical course. AB - Ki-1+ anaplastic large cell lymphoma (Ki-1+ ALCL) is a subtype of non-Hodgkin lymphoma (NHL) with defined histopathological characteristics but with highly variable clinical presentation and outcome. Although in most of the patients the disease behaves as an intermediate- or high-grade lymphoma, some patients present with an indolent clinical course. Factors that determine the clinical behavior of this lymphoma have not yet been identified. A case is reported of a 13-year-old girl who initially presented with Ki-1+ ALCL but later developed recurrent localized cutaneous disease and followed a clinical course similar to that of a low-grade lymphoma. PMID- 11255735 TI - Idiopathic CD4+ T lymphocytopenia in two siblings. PMID- 11255734 TI - A rare tumor of craniofacial bones in children: a pediatric chondroblastic osteosarcoma case with diagnostic and therapeutic problems. AB - Osteosarcoma of the cranial facial region is uncommon and only rarely involves the ethmoid or sphenoid bones. The authors report on an unusual case of a 17-year old male presenting with chondroblastic osteosarcoma of the maxillary, ethmoid, and sphenoid sinuses who remains well and disease-free at 46 months. He was treated with anterior craniofacial resection followed by postoperative radiotherapy to the sight of the primary tumor. He did not receive chemotherapy because of emerging hepatitis-B infection and vasculitis. The literature on extragnathic craniofacial osteosarcomas is reviewed with particular emphasis on treatment options of this rare tumor. PMID- 11255736 TI - Myelodysplastic features in polyarteritis nodosa. PMID- 11255737 TI - Oxidative stress disturbances in erythrocytes of beta-thalassemia. PMID- 11255738 TI - Follicle-derived thyroid cancer in young people: the Duke experience. AB - Follicle-derived thyroid cancer is rare in the young. The authors examined a population with a low rate of radiation exposure and who were treated at a single institution. The records of 56 patients diagnosed before the age of 25 years were analyzed. The majority of patients presented with an asymptomatic thyroid mass. All patients were treated surgically and half received postoperative ablation with 131I. Recurrent disease was detected in 29%. The presence of local metastases at initial surgery was a predictor of recurrence. No patient presented with distant metastases and no patient died of thyroid cancer. Although radiation exposure remains a risk factor for thyroid cancer in the young, only a minority of patients with thyroid cancer have a known history of exposure. Patients who are diagnosed at a young age have a high rate of long-term recurrence, and should be followed closely throughout their lives. PMID- 11255739 TI - A process for putting the PPS patient first. AB - The guiding principle of home care is to put the needs of patients before financial gain. However, Medicare's prospective payment system (PPS) offers incentives to do just the opposite--to put a dollar gain ahead of the patient. The challenge is--in spite of PPS's incentives--to put the patient first and yet remain solvent. PMID- 11255741 TI - The ongoing challenge of defining quality. AB - Home care professionals want to provide high-quality care, and their rate of financial reimbursement is one of the variables that substantially affects their ability to do so. How will the advent of prospective payment, and the accompanying incentive to limit care, influence the quality of care? Clearly, agencies find themselves in a difficult position. Unless home health agencies are willing to stipulate that they had been previously providing unnecessary care, they would be disingenuous to suggest that quality will not be affected. Furthermore, for an agency to believe that quality has not been compromised in the face of evidence to the contrary could be seen as an institutional form of cognitive dissonance. PMID- 11255742 TI - Who's who in home care & hospice policy. Key Members of the U.S. Senate & House of Representatives for the 107th Congress. PMID- 11255743 TI - Credentialing, accountability, & ethics. Keys to home care PPS. AB - Successful organizations with a reputation for excellence know the value of raising the bar for accountability, adherence to standards, and ethical practice at all levels of operations and service delivery. Numerous resources and tools can assist home care agencies in achieving success with their values and ethics intact. PMID- 11255745 TI - HCFA launches new hospice initiatives. HAA seeks hospice input. PMID- 11255746 TI - NAHC's 2001. Legislative priorities. PMID- 11255744 TI - Quality of care compliance plans under PPS. AB - With the onset of Medicare home health Prospective Payment System (PPS), home care agencies must retool their internal compliance efforts to address the new risk areas. PPS presents a reversal of the incentives that existed under previous Medicare reimbursement principles, significantly reducing the risk of non compliance and fraud in the financing of services while dramatically increasing non-compliance risks in areas related to quality of care and access to services. PMID- 11255747 TI - HCFA quality improvement initiatives. AB - With PPS, it becomes even more important that agencies ensure that quality of care be maintained and improved. This article describes some of HCFA's initiatives in quality of care. PMID- 11255748 TI - Keeping high standards, caring traditions. PMID- 11255749 TI - The new age of opportunity & risk. Medicare PPS. AB - The home care community must take appropriate measures to ensure that PPS does not undermine the values and principles of home care. The temptation of PPS to achieve profit at the expense of quality of care and access to services must be confronted by the home care community through reinforced values systems and concrete risk management measures. PMID- 11255750 TI - [Current status of organotin studied in China and abroad]. AB - The toxicity of organotin compounds and the current pollution status in China and abroad is discussed. Regulations and limitations on the usage of organotin compounds in western countries are introduced briefly in this paper. The examination results showed that the most Chinese harbors, rivers as well as larger lakes were contaminated by organotin, especially by tribuyltin compounds. Regulations for limiting the use of butyltin compounds as antifouling paints in Chinese marine industries are imperatively needed. Actions toward for the investigation of organotin contamination levels in food chains and in household commodities are also highly desired. PMID- 11255751 TI - [Role of protein kinase in the proliferation of human embryonic pulmonary fibrolasts stimulated by the supernatants of crocidolite-exposed alveolar macrophages]. AB - In order to study the role of protein kinase in the proliferation of lung fibroblasts induced by crocidolite. An in vitro model was established by rabbit alveolar macrophage (AM) and human embryonic pulmonary fibroblasts (HEPF). Using MTT color response method to measure HEPF proliferation, the influence of the inhibitor or activator of protein kinase (PKA, PKC and TPK) on the proliferation of crocidolite-induced HEPF were investigated. TiO2 was taken as negative control and SiO2 positive control. The results showed that the inhibitors of PKA, PKC and TPK could all inhibit the proliferation of HEPF induced by crocidolite, their activators could also promote the proliferation of HEPF. There all existed significant dose-effect relationships (P < 0.01), and the intensity in crocidolite group was inhibited or activated more than that in the controls. Through acting intensity analysis, the intensity was found as follows: TPK > PKC > PKA. It was suggested that TPK, PKC and PKA signal pathways were all involved in the process of the proliferation of crocidolite-induced HEPF, but TPK maybe played a key role in this process. This study provide leads for further research on identifying the bioactive factors of proliferation of crocidolite-induce HEPF. PMID- 11255753 TI - [Measurement of formaldehyde in disinfectant with spectrophotometry]. AB - Formaldehyde in disinfectant was measured by acetylacetone spectrophotometry. The regression equation was linear when the formaldehyde concentration was smaller than 10 micrograms/ml. The detection limit was 0.08 microgram/ml. The accuracy was less than 3.9% and the recovery varied between 100.2%-108.2% for formaldehyde disinfectant with different concentration, which was similar to titrimetry. It was demonstrated that the method was simple, reproducible, sensitive, precise and accurate, making it important for assay of formaldehyde in disinfectant when disturbance caused titrimetry unavailable. PMID- 11255754 TI - [Collecting of trichloroaniline in the air by micropore filter membrane and determination by gas chromatography]. AB - Trichloroaniline in the air was collected with micropore filter membrane and desorbed with cyclohexane, separated with a column OV-17, OV-210 and determined by GC-ECD. The materals for the filter of collection, solution of desorping and flow-rate of sampling were selected. The detectable limit is 0.01 mg/L. When the concentration of standard solutions is 1-10 mg/L, the relative standard deviation (RSD) is 4.5%-2.3%. There is a linear relation within the range of 0.005-30 mg/L. The sampling efficiency is 98.7%-100%. The samplers were stable for at least 15 days. This method is proved to be accurate, sensitive, fast and simple for sampling and carrying, and it is method is suitable for the determination of trichloroaniline in air, for personal detection and detection in the fields. PMID- 11255755 TI - [Toxicokinetics of terephthalic acid]. AB - In order to study the toxicokinetics of terephthalic acid(TPA) in rats, and provide scientific basis for its biological exposed index (BEI), the concentrations of urine TPA in rats after single oral administration in dose of 100 mg/kg BW were determined by high pressure liquid chromatography. The toxicokinetic parameters were computed by using 3P97 program. The results showed that the first-order kinetics and two-compartment model were noted on the elimination of TPA. The main toxicokinetic parameters were as follows: Ka = 0.51/h, T1/2ka = 0.488 h, T1/2 alpha = 2.446 h, Tpeak = 2.160 h, Ku = 0.143/h, T1/2 beta = 31.551 h, Xu(max) = 10.00 mg. The excretion rates of TPA in urine were about 50%, 52% and 53% in 0-24 h, 0-48 h and 0-72 h respectively after administration. TPA is well absorbed when given orally and rapidly eliminated via urine. Urine TPA at the end of work shift should be considered as a biomarker of exposure for the occupational workers. PMID- 11255752 TI - [Cloning of cNDAs involved in malignant transformation induced by glycidyl methacrylate]. AB - The malignant transformation of human embryonic lung fibroblasts (HELF) induced by glycidyl methacrylate (GMA) in vitro was used as a model for comparing gene expression between the transformed cells and controls. Using mRNA differential display PCR(DD-PCR) technique, the three cDNA fragments up-regulated in the transformed cells were identified and highly homologous to human breast cancer transcription factor(ZABC1), Tbx3 and mouse ubiquitin conjugating enzyme(E2) gene. These results suggest that activation or inactivation of genes including ZABC1, Tbx3 and E2 may be involved in GMA-induced transformation. PMID- 11255756 TI - [Effects of high temperature and cigarette smoke on the development of nervous system in early rat embryos]. AB - In order to explore the effect of high temperature or cigarette smoke and their combined effect on the development of nervous system in early rat embryos, pregnant rats at the 8-11th day of gestation were exposed to high temperature and water soluble substances of cigarette smoke. All indexed of embryonic nervous system development were determined with experimental treat logical methods. The results showed that following the increase of temperature or cigarette smoke exposure all indexes correlated with embryonic nervous system development and morphological differentiation were changed, with an apparent dose-effect relationship (P < 0.01). The changes were related to specific phase of embryonic development. The results showed also that the nervous system development and morphological differentiation induced by both high temperature and cigarette smoke were higher than that by any single factor. PMID- 11255757 TI - [Study on the effects of DNA damage induced by cigarette smoke in male mice testicular cells using comet assay]. AB - The mainstream whole smoke was collected using atmosphere collector-U shape glass plate absorption tube with poly-hole, and both dimethyl sulfoxide(DMSO) and phosphate buffer solution (PBS) were used as absorbents respectively. Comet assay was used to detect the effects of DNA damage of the two solutions of cigarette smoke in male mice testicular cells. The results showed that both the two solutions of cigarette smoke contained some DNA damage agents that could induce the DNA single strand breakage in male mice testicular cells. Besides, the solution of cigarette smoke absorbed by DMSO had stronger cytotoxicity and genotoxicity than the solution of cigarette smoke absorbed by PBS. The research also suggested that collecting cigarette smoke like this paper and detecting DNA damage effects using comet assay might be an ideal measure in studying male reproducible toxicity. PMID- 11255759 TI - [Effects of homocysteine on post-implantation rat embryo cultured in vitro]. AB - In order to investigate the effect of homocysteine(HCY) on the development of rat embryos, the post-implantation whole embryo culture(WEC) technique was used. Neural plate stage(GD9.5) rat embryos were explanted in rat serum medium(immediately centrifugal serum, ICS) with D,L-HCY(0,0.15, 1.5,2.0,4.0,6.0,8.0,10.0 mmol/L), and cultured for 48 hours. The results showed that HCY did cause damage to embryonic development and the damages of HCY on embryos were characterized in a significant dose-response pattern. The minimum teratogenic dose of D,L-HCY was 0.15 mmol/L, the incidence of dysmorphogenic embryos was 8.33%. Significant inhibiting effects of HCY on yolk sac(including reduced yolk sac diameter, shriveled surface, small or defective yolk sac vessels) and embryonic growth and morpholological differentiation were apparent with increased HCY, while HCY were 4 mmol/L or more (P < 0.05). The abnormalities included neural tube defects, delayed cardiac tube formation, pericardial effusion, incomplete flexion, small or missing forelimb buds and irregular somites, etc. These findings suggested that HCY might exert a direct effect on embryos and a "double effect" both on yolk sac and embryos. PMID- 11255758 TI - [Effect of vitamin E on morphological variation of retinal ganglion cells after microwave radiation]. AB - To determine the morphological variation in the primary cultured pig retinal ganglion cells induced by microwave and the protection of VE can supply some experiment foundation for study of effect of microwave and its protection. Retinal ganglion cells of pig were cultured in vitro and added VE of different concentration, Each group was taken after 30 mW/cm2 microwave intensity radiated for 1 h in shielded room by 2450 MHz continuous wave physiotherapy machine. Immediately after radiation, the morphological variation of cells was observed by optics microscope and transmission electronic microscope. The result showed that a portion of cells congregated, with their axon disappeared after radiation. Mitochondria and endoplasmic reticulum are detected swelling by transmission electronic microscope. The results showed that A poptosis cells can be observed. Cells of VE added groups had not obvious changes with optics microscope, but could be found that mitochondria swelling lightly and integrate mitochondria cristae by transmission electronic microscope. The results showed that microwave induced the morphological damage in primary cultured retinal ganglion cells, VE could reduced the damage of retina ganglion cells by microwave in some extent. PMID- 11255760 TI - [Effects of conjugated linoleic acid on the metastasis of mouse melanoma B16-MB]. AB - In order to explore the effects of conjugated linoleic acid on tumor metastasis, the proliferation, gap junction intercellular communication (GJIC), adhesion to extracellular matrix, and metastasis cascades of tumor cells were imitated in cell culture. The results showed that 100 and 200 mumol/L conjugated linoleic acid could inhibit the proliferation and the adhesion of tumor cells to extracellular matrix, and could recover its gap junction intercellulor communication. PMID- 11255761 TI - [Effect of lead exposure during pregnancy on hippocampal long-term potentiation and expression of NMDAR-2A mRNA in the off-spring rats]. AB - Chronic lead exposure during brain development is known to affect cognitive and behavioral functions in children and animals. The lead exposure on pregnant rats was used as a model to examine the effects of lead on the long-term potentiation(LTP) in hippocampal dentate gyrus(DG) region in vivo and the expression of NMDAR-2A mRNA by in situ hybridization in the off-springs. Female rats were exposed to 0.5 g/L or 2 g/L lead acetate in drinking water since 10 days before mating to weaning. The filial rats of 70-90 days old with blood lead levels below 100 micrograms/L were prepared for recording their DG-evoked population spike(PS) in the hippocampal DG area in vivo. The results showed that the range of LTP were (136 +/- 31)% in low lead group, (145 +/- 30)% in high lead group, and (319 +/- 114)% in control group. The reduction of NMDAR-2A mRNA expression 34%-44% in the granule and pyramidal cell layers caused by lead were seen in 21 days old rats. It indicated that the cognitive deficits induced by low level lead exposure in early stage of life may persist to adulthood, and the modified NMDAR gene expression may play a key role in the cognitive deficits associated with lead exposure during development. PMID- 11255762 TI - [Study of fungus polysaccharides compounds (FPC) in inducing the apoptosis of liver cancer cell Bel-7402]. AB - To observe the influence of fungus polysaccharides compounds (FPC) in inducing human liver cancer cell Bel-7402 apoptosis in cell cultivating in vitro, the authors analyzed tumor inhibitive gene P53 expression in Bel-7402 apoptosis by applying double immuno-marker. The result showed that the multilevel of FPC could all apparently induce Bel-7402 apoptosis. With the enhancement of FPC concentration, the authors observed chromatin condensation in some phases companying with the characteristic apoptosis. In the meantime, it could also greatly reduce the G1 and S, with obviously dose-response relationship. The percentage of cell apoptosis increased with the enhancing of concentration. In the high-level group the authors found typical DNA ladder eletrophoresis stripe. The result showed that the mechanism of the FPC antineoplastic effect had an intimate relation with its induction to apoptosis and that the result of FPC inducing tumor cell apoptosis had the character of P53 independence. PMID- 11255763 TI - [Study on the inhibition of tea pigments on the adhesion between monocyte and endothelial cells]. AB - In order to study the effects of tea pigments (TP) on the adhesion between monocyte and endothelial cells induced by oxidized low density lipoprotein(Ox LDL), the fluorescence activated cell sorter (FACS) and the cell ELISA were applied. The results showed that TP could inhibit the adhesion between monocyte and endothelial cells(P < 0.05), and the expression of inter cell adhesion molecule(ICAM-1) and vascular cell adhesion molecule (VCAM-1) proteins in a dose dependent manner(P < 0.05). The results suggested that the inhibition of TP on ICAM-1 and VCAM-1 expression was one of the mechanisms of its antiadhesion between monocyte and vascular endothelial cells, as well as that of its antiatherosclerosis. PMID- 11255764 TI - [Effect of zinc on the activities of ATPase of erythrocyte membrane]. AB - The effects of zinc on the activities of Na+, K(+)-ATPase and Ca2+, Mg(2+)-ATPase of erythrocyte membrane were studied both in vivo and in vitro. Male adult rats were divided into three groups and fed on diets with different concentration of zinc (2.2, 28 and 128 mg/kg diet). Eight rats of each group were sacrificed after 25 days and the activities of the two ATPase were measured. The rest rats were changed diet for an other seven days to detect the sensitivities of the ATPases. The activities of these ATPase were also determined in fresh human erythrocyte membrane treated with different concentration of zinc (0, 5, 10, 50, 100, 500 mumol/L). The results showed that the activities of both Na+, K(+)-ATPase and Ca2+, Mg(2+)-ATPase were in close relationship with zinc concentration, too low or too high zinc could decrease them. Compared with Na+, K(+)-ATPase, Ca2+, Mg(2+)-ATPase was more sensitive to zinc deficiency and in a slower reaction to zinc supplement, but it could stand higher zinc concentration. It was concluded that zinc played an important role on maintaining the normal activities of Na+, K(+)-ATPase and Ca2+, Mg(2+)-ATPase. PMID- 11255765 TI - [Effect of five kinds of vegetable seed oil on serum lipid and lipid peroxidation in rats]. AB - The effects of vegetable seed oil on hyperlipidemia induced by high lipid diet in rats. Male adult Wistar rats were fed on the test diet containing 94% high lipid diet and 6% lard pinon seed oil, perilla seed oil, blackcurrent seed oil, borage seed oil and evening primrose seed oil respectively for 3 weeks. The results showed that the vale of trilyceride(TG), total cholesterol(TC), low density lipoprotein cholesterol (LDL-C), LDL-C/HDL-C(high density lipoprotein cholesterol) ratio increased and the vale of HDL-C/TC ratio and lecithin cholesterol acyltransferase(LCAT) activity decreased in the groups with vegetable seed oil were less than that of the control group. The results suggested that all the five kinds of vegetable seed oil had the effect of regulating lipid metabolism of hyperlipidemia rats to some extent. Pinon seed oil and borage seed oil may be well suited for the prevention of atherosclerosis. PMID- 11255766 TI - [Effect of a complex tea on reducing blood lipid in rabbits]. AB - In order to observe the effect of a complex tea composed of tea, ginkgo leaf, red koji and fructus lycii on reducing blood lipids in experimental rabbits, 40 rabbits weighted from 1.5 to 2.0 kg were divided randomly into 5 group: (I) normal control group, (II) hyperlipidemia model(positive control), (III) green tea control group, (IV) low dose complex tea group, and (V) high dose complex tea group. Blood triglyceride(TG), cholesterol(TC), lower density lipoprotein(LDL) and high density lipoprotein(HDL) were measured before experiment and 6.12 weeks later. Then the pathological changes of heart, aorta and liver were observed. After 12 weeks of experiment, the results showed that, (1) hyperlipidemia characterized by high TC has been induced by high-fat feeds in group II, (2) the complex tea can reduce body weight and eliminate the lipids deposition in aorta and liver. (3) the complex tea can decrease serum TG, TC,LDL and increase serum HDL in the high complex tea group. The results indicate that the complex tea is more effective than green tea on reducing blood lipids in experimental rabbits. PMID- 11255768 TI - [Chemical properties and scavenging effects on active oxygen radicals of tea polysaccharides]. AB - The chemical composition and scavenging effects of a kind of glycoprotein from tea on active oxygen free radicals were studied. The tea polysaccharides(TPS) were composed of arabinose, ribose, xylose, glucose and galactose. TPS(8.5-170 mg/L) could scavenge the .OH produced in deoxyribose-Fe2+ reaction and the scavenging rate was between 5.5% to 74.7%. PMID- 11255767 TI - [Effect of methionine supplementation on the selenium bioavailability in rats fed on grains from Keshan disease endemic area]. AB - In order to study the effect of dietary methionine on tissue selenium and glutathione peroxidase activity(GPX) in rats fed on grains from Keshan Disease endemic area, a nonpurified selenium (Se) deficient diet (containing Se 0.007 mg/kg diet) taken from Keshan Disease endemic area was supplemented with selenomethionine (SeMet) to provide selenium content in diet for 0.007, 0.06 and 0.50 mg/kg and added or not added with DL-methionine (Met) 4 g/kg to make 6 kinds of diet. Fifty four male weanling Wistar rats were randomly divided into 6 groups, consuming each one of the 6 diets for 8 weeks. The Se content and GPX activity in tissues of animals with Met supplementation were compared with those without Met supplementation. The results showed that adding Met did not result in significant changes of tissue Se content and GPX activity in animals having dietary Se of 0.007 mg/kg, except for lower muscle Se content. However, in animals having dietary Se of 0.06 mg/kg and supplementing Met resulted in selenium redistribution in tissues-decrease of Se content in muscle, increase of Se content in liver and blood and significant elevation of GPX activity in all tissues. In animals having methionine-added and dietary Se of 0.50 mg/kg, tissue Se content declined to various extent, while GPX activity remained unchanged. The results suggest that SeMet (main chemical form of Se in cereals) is preferentially incorporated into body protein when dietary methionine is limited. Once Met is supplemented, dietary SeMet would provide more Se for the syntheses of GPX and other selenoproteins. The results further suggest that marginal deficiency in sulfur-containing amino acids in the diet from Keshan Disease endemic area might be an additional factor for the development of Keshan Disease under selenium deficiency. PMID- 11255769 TI - [Survey of 3-monochloropropane-1,2-diol in soy sauce and similar products]. AB - A survey of 3-monochloropropane-1,2-diol (3-MCPD) in soy sauce and other similar products available in China has been completed. Thirty samples of soy, oyster sauce and other sauces were purchased from six supermarkets in Beijing during March and April 2000 and analysed using a validated method of analysis by capillary gas chromatography with mass spectrometric detection. The UK Food Advisory Committee (FAC) has advised a level of 0.01 mg/kg for 3-MCPD. Following reports that high levels had been detected in some brands of soy sauce in China. 3-MCPD was undetectable with a detection limit of 0.01 mg/kg in 15 of the 30 samples analyzed in the survey, with a further 4 samples (13.3%) containing very low levels of between 0.01 and 0.02 mg/kg. However, 5 samples (16.7%) contained 3 MCPD levels above 1 mg/kg. PMID- 11255770 TI - [Effects of methyl mercury chloride on nuclear factor-kappa B DNA binding activities of nuclear protein extracts from developing rat cerebra and cerebella]. AB - The effects of methyl mercury chloride (MMC) on DNA binding activities of nuclear factor kappa B(NF-kB) in developing rat cerebella and cerebra were investigated with electrophoretic mobility shift assays(EMSAs). The bindings of NF-kB in nuclei of rat cerebra and cerebella to kB probes showed two bands on gel shift in both control and experiment groups. NF-kB I and NF-kB II DNA binding activities of nuclear protein extracts from rat cerebra exposed to MMC in uterus, was lower than control groups on postnatal day 3 and 7, while that from rat cerebella was higher than control groups. The results suggested that the reactive abilities of neural cell to MMC between cerebra and cerebella were different. In binding reaction mixture, the quantities of MMC increased with the increase of NF-kB DNA binding activities of nuclear protein extracts. PMID- 11255771 TI - [Treatment of intermediate hypermetropia using laser in situ keratomileusis- retrospective study (1995-1999)]. AB - In this study, the authors present their findings concerning the usage of the LASIK method in 225 cases of hyperopia. This method has been used in our ELAS refractive centre from 1995 in the treatment of hyperopia up to +7.0 D. The average preoperative spherical equivalent in this group was +4.91 D, and in cycloplegic refraction 20 D (+/- 1.94 D). The postoperative average spherical equivalent was 0.53 D (+/- 0.58 D). Uncorrected visual acuity is used as the main criterion in assessing the effectiveness of the refractive procedure. The average uncorrected visual acuity was improved from a preoperative value of 0.4 (+/- 0.27) to a postoperative one of 0.8 (+/- 0.24). The average value of the best correct visual acuity worsened from 0.98 (+/- 0.31) preoperatively to 0.88 (+/- 0.23) postoperatively. This was perhaps caused by the side-effects of laser ablation (glare), which occurred in using LASIK with cases of hyperopia of over 0 D. For this reason, the choice of the LASIK method for correction of middle hyperopia should be made with caution. PMID- 11255772 TI - [Importance of laser peripheral iridoplasty in the treatment of acute closed angle glaucoma]. AB - The aim of this prospective study has been to evaluate effectivity of laser peripheral iridoplasty (ALPI) as a first choice therapy for intraocular pressure (IOP) reduction in acute primary angle-closure glaucoma (PACG). The study included 12 patients with the first attack of acute glaucoma in whom IOP in spite of continuing medical therapy for 12 hours persisted at the level of 5.32 kPa (40.0 mmHg) and higher. We used Argon laser manufactured by HGM E-005. Laser parameters were: spot size 300-500 um, time 0.2-0.5 s, intensity 300-500 mW. Laser burns were placed to the whole circumference of the iridocorneal angle numbering 50-60 with the use of Goldmann contact lens. Applanation tonometry was used to measure IOP shortly before the laser therapy and 1 and 2 hours after ALPI. The mean IOP prior to treatment was 6.00 +/- 1.46 kPa (51.7 +/- 11.04 mmHg). The mean IOP 1 hour after ALPI was 2.77 +/- 0.44 kPa (20.83 +/- 3.31 mmHg) and after 2 hours 2.39 +/- 0.26 kPa (18.0 +/- 2.0 mmHg) which represents 45.8% reduction in IOP. Two hours after treatment all patients had transparent corneas. The results of this study confirm that ALPI effectively reduces IOP and enhances transparency of the cornea in acute PACG for the definitive treatment by laser iridotomy. PMID- 11255773 TI - [Visual function in myopia 2 years after photorefractive keratectomy]. AB - 1. 86 myopes undergoing photorefractive keratectomy (PRK) with refraction from 0.25 D to -12.00 D were divided into 4 groups: A: up to -2.75 D, B: -3.00 D to 5.75 D, C: -6.00 D to -8.75 D and D: -9.00 D to -12.00 D. 20 emmetropes of the same median age were evaluated as a control group. The patients were examined 2 years after PRK. 2. Best corrected visual acuity (BCVA) increased in groups A, C, D and decreased in group B compared to values one year after PRK. Both changes were only nonsignificant. 3. Contrast sensitivity (CS) was equal to values one year after PRK in group A, the same was true for groups C and D with exception of lower values in the highest spatial frequency. In group B, CS decreased significantly compared to values one year after PRK. 4. In groups A, B a C number of patients without rest correction increased, regression was seen only in group B. 5. BCVA changed only nonsignificantly under glare of 41.1 cd/m2, 342.6 cd/m2 and 1360.4 cd/m2 in 97% of patients. PMID- 11255775 TI - [Clinical features and therapy of giant cell temporal arteritis]. AB - Giant-cell temporal arteritis is an urgent condition in ophthalmology as successful treatment depends on early diagnosis and effective therapy before the development of ophthalmological symptoms. The authors investigated on a long-term basis five patients. In the first one complete regression of general and ocular symptoms occurred and vision was preserved. The second patient was admitted already with loss of vision of one eye and despite intensive treatment it did not prove possible to save vision of the second eye. The third patient developed, after clinical recovery, a relapse which was again brought under control. In another patient of relatively younger age it proved possible to arrest the progress of the disease without loss of vision. In the last patient visual acuity improved after treatment. PMID- 11255774 TI - [The scotopic electroretinogram and age]. AB - 1. Electroretinographic examination according to recommendations of the International Society for Clinical Electrophysiology of Vision (ISCEV) was implemented in 115 healthy subjects of different age groups. 2. No significant difference was recorded in the results of men and women. 3. The amplitudes of all investigated retinal responses decline in a linear fashion with age and their peak latencies are prolonged. The mentioned changes are statistically significant even if changes of 0.5 to 0.8% per year are involved. PMID- 11255776 TI - [Intralenticular metal foreign bodies]. AB - OBJECTIVE: The paper is concerned with the problem of extraction of foreign metal bodies, which remained after a perforating injury in the patient's lens. The authors discuss early and late extraction of the opacifying lens. MATERIAL AND METHODS: In three successive accidents of young men (22-34 years) the authors compare the fate of intralenticular metal bodies and the lens. RESULTS: All three metal bodies were extracted, at the same time also surgery of the opacifying lens was performed with implantation of a posterior chamber intraocular lens into the sac after continual curvilinear capsulorrhexy. The postoperative visual acuity was 6/6. CONCLUSION: With regard to contemporary possibilities of careful solution of lenticular surgery, using high standard viscoelastic materials, it is advisable to remove primarily the foreign body from the lens along with the damaged opacified lens and replacement of the latter by a posterior chamber intraocular lens as the quickest possible solution of the posttraumatic state. PMID- 11255777 TI - [Glaucoma and myopia. The optic nerve papilla]. AB - The author investigated a group of 80 eyes of 40 myopic patients, 20 of whom are treated on account of glaucoma (17 cases of primary open angle glaucoma and 3 cases of pigmentary glaucoma), another 5 patients were on the records with suspected glaucoma and in 15 glaucoma was not detected. The work is focused on possible differentiation of typically myopic papillary changes and the papilla of myopic eyes altered by glaucoma. PMID- 11255778 TI - [A huge foreign body in the orbit]. AB - The authors present an account on a two-year-old child which while playing pushed the broken end of a pencil into the orbit above the bulbus. With regard to the nature of the object it was extracted by two-stage surgery. The injury did not affect the child's ocular functions. PMID- 11255779 TI - [Bilateral megalopapilla]. AB - The authors present an account on the unusual finding of enlarged optic discs in a nine-year-old boy. The clinical finding is consistent with bilateral megalopapilla with preserved intact central and peripheral vision. The authors draw attention to possible mistakes and errors which may be associated with this uncommon clinical finding. PMID- 11255780 TI - [Atypical forms of retinopathy of prematurity]. PMID- 11255781 TI - [Drainage implants in the treatment of glaucoma (part I)]. PMID- 11255782 TI - [Dr. Jan Vanysek: founder of the ophthalmology clinic in Hradec Kralove. A distinguished personality in Czechoslovak ophthalmology]. AB - Professor Vanysek was the first Czechoslovak ophthalmologist who overstepped the boundaries of the Republic and became one of the foremost scientist in ophthalmology. He was head of a department, consistent more with ideas of the 21st than 20th century. He founded two important ophthalmological school--in Hradec Kralove and Brno. He laid the foundations for an experimental institute in Brno which would have become one of the foremost European institutions if Vanysek had not been condemned by the totalitarian regime in the seventies. PMID- 11255783 TI - [ Johann Sebastian Bach and his blindness on the 250th anniversary of the artist's death]. PMID- 11255784 TI - [Isolated and combined occlusion of the cilioretinal artery and the central retinal vein]. AB - Cilioretinal artery occlusion is rare pathological condition because of infrequent occurrence of the cilioretinal artery in the human retina. It presents as an isolated entity, or as a combined cilioretinal artery--central retinal vascular, mostly venous occlusion. Authors present the course, angiographic features, prognosis, risk factors and final outcome of the isolated cilioretinal artery occlusion and combined cilioretinal artery--central retinal vein occlusion. Occlusion of the cilioretinal artery is believed to result from obstruction of the central retinal vein, as a primary process, in the case of combined cilioretinal artery--central retinal vein occlusion. PMID- 11255785 TI - Changing unhealthy behavior by the numbers. PMID- 11255786 TI - Internet-based program cuts benchmarking tasks. AB - Practicing by the numbers: The Medical Group Management Association has taken on the task of profiling U.S. physicians, and it's thinking big. The strategy is to entice physician participation with cheap enrollment fees and the promise of aggregated data fast. PMID- 11255787 TI - Three-pronged process improvement gets great buy-in from physicians. AB - Pneumonia process improvement. St. Joseph's Hospital in Tampa wanted to improve the process for the more than 600 people it sees every year with community acquired pneumonia. One strategy was to keep the data-collection and analysis tasks simple and focused. PMID- 11255788 TI - Measuring PPO performance is an inexact science at best. AB - The American Accreditation HealthCare Commission has found some PPOs, that are making headway in what is a fragmented and difficult task. PMID- 11255789 TI - Stroke care--there's room for improvement. AB - The HCIA-Sachs Institute points out exactly where and how U.S. hospitals can rise to the level of their top-performing peers. PMID- 11255790 TI - Induction of unscheduled DNA synthesis in hairless mouse epidermis by 8 methoxypsoralen plus ultraviolet A (PUVA). AB - Induction of unscheduled DNA synthesis (UDS) by 8-methoxypsoralen (8-MOP) plus ultraviolet A (UV-A) (PUVA) was investigated in the epidermis of female hairless mice by means of an in vivo--in vitro assay using a liquid scintillation counting method. Groups of three to five 8-week-old female hairless mice had 8-MOP applied once onto two areas of the back after stripping of the stratum corneum with adhesive tape to enhance skin penetration, and were irradiated with UV-A. Skin samples were taken and cultured in a medium containing [3H]thymidine with or without hydroxyurea (HU) for 2 hr. DNA of the epidermis was extracted, and the incorporation of [3H]thymidine into DNA and the DNA content were determined with a liquid scintillation counter and a fluorescence spectrophotometer, respectively. Induction of UDS was judged in terms of the UDS index [(the ratio of DNA synthesis in the presence of HU to that in its absence) x 100]. In a time course study, the UDS index was increased at 1, 2 and 24 hr after 1 x 10(5) J/m2 UV-A irradiation with 0.001% 8-MOP, reaching the maximum level at 24 hr. In a dose-response study, it was significantly increased at the dose of 1 x 10(5) J/m2 of UV-A at 24 hr with 0.001% 8-MOP, but showed no significant change at the doses of 0.5 x 10(5), 2 x 10(5) and 4 x 10(5) J/m2. In a further study on the effect of varying the dose of 8-MOP, the UDS index was significantly increased at 0.001 and 0.002% 8-MOP at 24 hr after 1 x 10(5) J/m2 UV-A irradiation, reaching the maximum level with 0.002% 8-MOP. The increase of the UDS index in these studies was less than 3-fold. These results show that PUVA causes a small induction of UDS, which might be due to slow DNA excision repair over a long period. PMID- 11255791 TI - Restitutive response of Mini rat liver to injury induced by a single oral administration of thioacetamide. AB - Mini rats are a transgenic rat strain carrying antisense gene for rat growth hormone (GH), resulting in retarded growth and a lower blood GH level (136 +/- 42.0 ng/mL) compared with that of age-matched parental strain Wistar rats (329 +/ 337 ng/mL). Mini rats have been used by several investigators as a GH deficiency model. In this work, we gave a single oral administration of thioacetamide (TAA), a hepatotoxicant, to both Mini rats and Wistar rats to ascertain the influence of GH deficiency on liver response to chemically induced injury and subsequent regeneration. TAA administration caused liver injury in both strains, with a greater extent of injury in Mini rats. Proliferation of bile epithelial cells and so-called oval cells was prominent at Day 3 in Mini rats only, and this change correlated well with serum total bilirubin concentrations. Antibody against Ki-67 antigen revealed that cellular proliferation after TAA-induced liver injury was suppressed but prolonged in the Mini rat liver. Although hepatic stellate cells and Kupffer cells/macrophages were more abundant in the livers of TAA-treated Mini rats, the hepatic expression patterns of hepatocyte growth factor and transforming growth factor beta 1 were comparable to those of Wistar rats. Insulin-like growth factor-I gene expression was significantly reduced in the Mini rat liver. Our results imply that a lower GH level may exacerbate chemically induced liver injury, enhance infiltration/proliferation of non-parenchymal cells, suppress regeneration of hepatocytes, and induce proliferation of bile epithelial cells and oval cells when the liver is injured by TAA. PMID- 11255792 TI - A possible relationship between abortions and placental embolism in pregnant rabbits given human granulocyte colony-stimulating factor. AB - Developmental toxicity studies were conducted in rats and rabbits with a human G CSF derivative (NTG). As reported for G-CSF, increases in abortions and fetal mortality were observed in rabbits, but not in rats given NTG. Histopathological examination of the rabbit placenta revealed accumulation of neutrophils in vessels and necrosis of the tissues surrounding these vessels. To assess the mechanism of abortion and fetal death in rabbits given G-CSF, 125I-labeled NTG was given intravenously on Day 18 of pregnancy after repeated administration of cold NTG on Days 6 through 17 of pregnancy, and the feto-maternal distribution of radioactivity was examined. In a rabbit given 20 micrograms/kg, high radioactivity was observed in the endometrium, placenta, and some parts of the decidua at 6 hr when the concentration of radioactivity in maternal blood had already decreased. At 24 hr after administration of 200 micrograms/kg NTG, high radioactivity was still detected in parts of the maternal placenta. These patterns of distribution suggest that embolism occurred in parts of the uterus and placenta which might have caused congestion. Radioactivity in the TCA precipitates in the fetus was low, suggesting that NTG does not readily transfer to the fetus. These results strongly suggest that neutrophils accumulated in the vessels of placenta and induced embolism leading to abortions and fetal mortality in the rabbits given G-CSF. PMID- 11255793 TI - Sperm motion analysis in rats treated with adriamycin and its applicability to male reproductive toxicity studies. AB - Adriamycin (ADR), an anticancer drug which induces testicular toxicity, was administrated to Slc:SD male rats at doses of 0.5, 1.0, and 2.0 mg/kg intravenously once a week for 4 weeks. The males treated with ADR were mated with untreated females, and sperm analyses (motion, count, and morphology) were performed. Sperm motion was analyzed by Hamilton-Thorne Sperm analyzer (HTM-IVOS) to investigate the useful parameters. Copulated females were necropsied at Day 13 of gestation, and reproductive status was evaluated. In ADR-treated groups, the testicular weight was dose-dependently decreased. Associated with this decrease was a depletion of the number of spermatogonia noted histopathologically at all dosage levels. Sperm morphological abnormalities, which were classified as tailless sperm and/or no-hook head sperm, were increased in both the 1.0 and 2.0 mg/kg groups. The males treated with ADR at 1.0 and 2.0 mg/kg had a decreased number of sperms per cauda epididymis. In sperm motion analysis, decreases in the percentage of motile sperm, percentage of progressive sperm, and sperm velocity (straight line velocity and curvilinear velocity) were noted at 2.0 mg/kg. Impaired fertility was noted at 2.0 mg/kg in the form of decreased numbers of implantations and live embryos, and an increased number of pre-implantation losses. In conclusion, ADR induced deterioration of sperm motion and sperm content, which were responsible for the adverse effect on male fertility. The most sensitive indicators to detect male reproductive toxicity induced by ADR were testicular weight and histopathological findings in the testis. Among the parameters generated by HTM-IVOS, the percentage of motile sperm, the percentage of progressive sperm, and sperm velocity are useful for assessing male fertility. PMID- 11255794 TI - Collaborative study on rat sperm motion analysis using CellSoft Series 4000 semen analyzer. AB - A collaborative study was conducted to determine useful and sensitive rat sperm motion parameters in a CellSoft Series 4000 semen analyzer to detect the effects of compounds on sperm motion. The effects on the sperm motion parameters were investigated using alpha-chlorohydrin, boric acid, ethinylestradiol, ethyl methanesulfonate, nitrazepam, nitrobenzene, ornidazole, sulfasalazine or valproic acid which are well known to induce reproductive or testicular toxicities. All compounds used in this study decreased percentage of motile sperm (% motile). Curvilinear velocity (VCL), maximum and mean amplitude of lateral head displacement (ALH max and ALH mean) were decreased by treatment with all compounds except for valproic acid. Treatment with alpha-chlorohydrin, ornidazole or sulfasalazine under mid-dosage regimens decreased only these parameters. Beat cross frequency (BCF) was increased by treatment with sulfasalazine. There were some treatments which caused either decreased or increased changes irrespective of dosage regimen in linearity, average radius, percentage of circular-swimming sperm out of motile sperm (circular/motile) and percentage of circular-swimming sperm out of all sperm (circular/all). Based on these results, we concluded that % motile, VCL, ALH max and ALH mean are considered useful and sensitive parameters for evaluating the effects of compounds on sperm motion. A parameter of BCF can be useful to detect the effects of specific compounds on sperm motion. Linearity, average radius, circular/motile and circular/all are not considered useful or sensitive indicators to detect the effects of compounds on sperm motion. PMID- 11255795 TI - Physical energy: fuel metabolism. PMID- 11255796 TI - Fatigue of anemia. PMID- 11255797 TI - Consumer perceptions of energy. PMID- 11255798 TI - The impact of the supply of glucose to the brain on mood and memory. PMID- 11255799 TI - Diet, neurochemicals, and mental energy. PMID- 11255800 TI - Dieting and weight loss: the energy perspective. PMID- 11255801 TI - Sleep and circadian rhythms: basic and clinical findings. PMID- 11255802 TI - Stress, fatigue, and behavioral energy. PMID- 11255803 TI - Energy for a new millennium--regulatory perspectives. PMID- 11255804 TI - Energy for a new millennium. PMID- 11255805 TI - Energy claims in the 21st century. PMID- 11255806 TI - Psychology and cultural aspects of energy. PMID- 11255807 TI - Metachromasia of the endocervical epithelium for delineating the internal os. AB - OBJECTIVE: To study the metachromatic stainability of the endocervical epithelium. STUDY DESIGN: Surgical specimens that included endocervical epithelium were deparaffinized and stained with toluidine blue at various pH levels and after enzyme digestion. Glycosaminoglycans were separated from endocervical tissue and analyzed by electrophoresis. RESULTS: The endocervical epithelium stained metachromatically purple with toluidine blue in pH 4.1 buffer, but the staining disappeared after chondroitinase digestion. The electrophoretic mobility of the glycosaminoglycan fraction separated from the endocervical epithelium was compatible with nonsulfated chondroitin. CONCLUSION: Endocervical epithelium was identifiable from metachromatic staining below the internal os, and the electrophoretic results of the tissue extract suggested that nonsulfated chondroitin is the main component of glycosaminoglycans in endocervical epithelium. PMID- 11255808 TI - Relative risk of high grade squamous intraepithelial lesion associated with prior abnormal Pap smears. AB - OBJECTIVE: Pap smear frequency remains controversial, especially for women with consecutive negative smears. We undertook the current study to ascertain the association of high grade squamous intraepithelial lesions (HSIL) and prior abnormal Paps. STUDY DESIGN: Women with biopsy-proven HSIL (cervical intraepithelial neoplasia 2 and 3) diagnosed between September 1996 and December 1997 and age-matched controls with a negative Pap obtained during the same time period were selected. RESULTS: Sixty-three cases (mean age = 32 years) of HSIL and 69 controls (mean age = 33 years) constituted the study population. Any prior abnormal diagnosis conferred a 15-fold increased risk of HSIL on the current Pap (50/63 vs. 14/69, P < .0001). When limited to the 60 women with at least three prior Paps, the odds ratio for HSIL on the current Pap with any prior abnormal was 18 (28/31 vs. 10/29, P < .0001). Three cases had at least three consecutive negative Paps prior to the diagnosis of HSIL. CONCLUSION: Women with one or more prior negative Pap smears had a significantly decreased risk of HSIL on the current Pap. Consecutive negative Paps did not appear to further decrease the risk; 10% of HSIL patients had had three or more consecutive prior negative Paps. To detect HSIL at its earliest stage, women should be advised to continue annual Pap screening in spite of consecutive negative results. PMID- 11255809 TI - Prophylactic actinomycin D for high-risk complete hydatidiform mole. AB - OBJECTIVE: To evaluate the effectiveness of one course of prophylactic actinomycin D in reducing the malignant sequelae requiring chemotherapy in high risk complete hydatidiform mole (CHM). STUDY DESIGN: A double-blind, randomized, controlled clinical trial was carried out at King Chulalongkorn Memorial Hospital. Sixty cases of CHM classified as high risk were recruited and randomly allocated to a chemoprophylactic or control group. Within one week after evacuation of molar tissues, actinomycin D was administered in the chemoprophylactic group. Patients in the control group were given only intravenous fluid and analgesic drugs. The number of patients with malignant sequelae who required therapeutic chemotherapy after evacuation of hydatidiform mole in each group was recorded. RESULTS: The incidence of malignant sequelae was 13.8% (95% confidence interval [CI] = 3.9-31.7%) in the chemoprophylactic group and 50.0% (95% CI = 31.3-68.7%) in the control group. The risk reduction of malignant sequelae with one course of actinomycin D chemoprophylaxis in high-risk CHM was 72.4% (95% CI = 26.7-89.6%) (P = .005). The side effects of prophylactic chemotherapy were stomatitis, nausea/vomiting, sore throat with oral ulcer and hair loss. CONCLUSION: One course of actinomycin D given as chemoprophylaxis decreased by 72.4% malignant sequelae after evacuation of molar tissue in patients with high-risk CHM. This may be particularly beneficial in patients with high-risk CHM who cannot be followed closely, whose compliance is in question and for whom hormonal follow-up is not available or unreliable. PMID- 11255810 TI - Distribution of transforming growth factor-beta isoforms TGF-beta 1, TGF-beta 2 and TGF-beta 3 and vascular endothelial growth factor in vulvar lichen sclerosus. AB - OBJECTIVE: To study the distribution of transforming growth factor beta (TGF beta) isoforms TGF-beta 1, TGF-beta 2 and TGF-beta 3 and vascular endothelial growth factor (VEGF) in vulvar lichen sclerosus. STUDY DESIGN: Biopsies were obtained from 10 patients with vulvar lichen sclerosus, snap frozen and stained with polyclonal antibodies to TGF-beta 1, TGF-beta 2, TGF-beta 3 and VEGF. Control tissues used were uninvolved thigh tissue from two of the lichen sclerosus patients and normal vulvar tissue obtained from eight patients during gynecologic procedures. Two specimens of morphea were also examined. RESULTS: Weak TGF-beta 1 staining was demonstrated in the epidermis of all the lichen sclerosus, morphea, thigh and five of the eight normal vulvar specimens. Slight increase in TGF-beta 1 staining was seen in the upper and middermis in 6 of the 10 lichen sclerosus specimens and in the morphea specimens as compared to the control tissue, and this staining was localized within cells. TGF-beta 2 staining was present throughout the epidermis in all the normal thigh, normal vulva, lichen sclerosus and morphea specimens. TGF-beta 2 staining was increased within cells in the upper and middermis of the lichen sclerosus and morphea specimens. TGF-beta 3 staining occurred in the basal half of the epidermis of all the control, lichen sclerosus and morphea specimens, and only slight upper dermal staining of a few individual cells was seen in 3 of the 10 lichen sclerosus specimens. VEGF staining was similar in the normal tissues, lichen sclerosus and morphea. CONCLUSION: These results suggest that TGF-beta may. PMID- 11255811 TI - Misoprostol administration in medical abortion. A comparison of three regimens. AB - OBJECTIVE: To determine the best regimen for using misoprostol after methotrexate in medical abortion with respect to outcome and side effects. STUDY DESIGN: In a cohort study, we compared vaginal misoprostol in one cohort of 134 women who used 800 micrograms dry tablets with 99 women who used 600 micrograms wet and 197 women who used 800 micrograms wet. These cohorts were compared with respect to outcome and side effects. RESULTS: The "dry" group had fewer completed abortions by day 8 (55.2% as compared to 69.7% and 71.1%, P = .008) but similar surgery rates. The dry group also had fewer side effects, especially fever and chills (4.5% as compared to 25.3% and 40.6%, P = .0001) and vomiting (8.2% as compared to 16.2% and 20.3%, P = .01). CONCLUSION: Of the three methods, the one consisting of 600 micrograms of wet misoprostol is the most effective for early completion of abortion and has the fewest side effects. PMID- 11255812 TI - Insertion of a slim fit tampon into the urethra. A case report. AB - BACKGROUND: A 14-year-old accidentally inserted a slim fit tampon into her urethra. Such a case could occur again in young teenagers who otherwise have never inserted a tampon and especially have yet to see a gynecologist for their first gynecologic examination. CASE: Accidental insertion of a slim fit tampon into the urethra by a 14-year-old necessitated cystoscopic resection of the engorged tampon. CONCLUSION: Although this is the only case reported of urethral tampon placement, one must include it as part of the differential diagnosis in assessing acute onset of pain or hematuria after the placement of a slim fit tampon. PMID- 11255813 TI - Acute liver failure in pregnancy. A case report. AB - BACKGROUND: Liver disease in pregnancy can be grossly divided into those disorders coincidentally occurring during the pregnant state and hepatic diseases limited to pregnancy. Numerous infectious agents can result in acute hepatitis and include not only the hepatitis viruses--A, B, C and E--but herpesvirus and cytomegalovirus as well. Coxsackie B viruses can cause several clinical presentations, ranging from asymptomatic to mild febrile illness to myocarditis and meningitis. Rarely has coxsackievirus infection been associated with fulminant hepatic failure. CASE: A Coxsackie B virus infection resulted in acute liver failure in a gravid woman. The patient was managed expectantly, with resolution of the liver disease and delivery five weeks after discharge. CONCLUSION: The onset of hepatic disease is insidious, with only vague symptoms or minor complaints often heralding the progression to liver failure. A careful history, physical examination and appropriate diagnostic tests can help determine the etiology of hepatic disease and help decide whether expectant management of the gravid patient or immediate delivery is appropriate. PMID- 11255814 TI - Spontaneous remission of cyclic neutropenia during pregnancy. A case report. AB - BACKGROUND: Cyclic neutropenia is characterized by regularly recurring episodes of neutropenia. It has been reported that pregnancy often has a mitigating effect on the symptoms. However, there have been no detailed studies on changes in the neutrophil count before, during and after pregnancy. CASE: A 24-year-old woman with cyclic neutropenia had a successful pregnancy, during which her symptoms improved spontaneously, with decreased severity. The disease recurred soon after pregnancy, and cyclic neutropenia was inherited by the child. CONCLUSION: Cyclic neutropenia may follow a favorable course during pregnancy, with decreased severity, but postpartum maternal and neonatal complications can occur. PMID- 11255815 TI - Broad ligament twin pregnancy. A case report. AB - BACKGROUND: Broad ligament pregnancy is an uncommon form of ectopic pregnancy. CASE: A 34-year-old, 11-week-pregnant woman, gravida 2, para 0, presented with left lower abdominal pain. She had undergone a right salpingectomy due to tubal pregnancy six years previously. She had a left broad ligament twin pregnancy, and excision of the pregnancy and left tube were performed. She was well at discharge and the six-week follow-up. CONCLUSION: This is the first case report of a broad ligament twin pregnancy after spontaneous conception. PMID- 11255816 TI - Recurrent second-trimester pregnancy loss. PMID- 11255817 TI - Urethral diverticulum in women. PMID- 11255818 TI - Postablative risk of endometrial carcinoma after high-dose estrogen. PMID- 11255819 TI - What is shoulder dystocia? PMID- 11255820 TI - Betamethasone alteration of the one-hour glucose challenge test in pregnancy. AB - OBJECTIVE: To determine if betamethasone administration alters the one-hour (50 g) glucose challenge test (GCT) and, if so, to determine the duration of this effect. STUDY DESIGN: Pregnant women with singleton gestations eligible to receive antenatal betamethasone (24-34 weeks) with no evidence of infection, diabetes mellitus, hepatic or pancreatic disease, betamimetic therapy or prolonged steroid use were invited to participate. Betamethasone (12 mg) was administered intramuscularly upon admission and repeated in 24 hours. After an 8 hour fast, a 50-g GCT was administered 24 hours (day 1) and 72 hours (day 3) after the second betamethasone injection. Plasma glucose samples were drawn one hour after the glucose challenge. Statistical analysis utilized the McNemar exact test. RESULTS: Seven patients, all with normal GCTs 7-10 days outside the study period, were enrolled. Six had abnormal GCT results on day 1 (P = .03). Three had abnormal results on day 3. CONCLUSION: Betamethasone administration results in abnormal one-hour GCTs in pregnant women. PMID- 11255821 TI - Prophylactic oophorectomy and ovarian cancer surveillance. Patient perceptions and satisfaction. AB - OBJECTIVE: To evaluate decision making, information gathering, satisfaction and regret in women at increased risk of ovarian cancer who had undergone prophylactic oophorectomy or ovarian cancer surveillance. STUDY DESIGN: Thirty women undergoing prophylactic oophorectomy (median age, 47 years) and 30 women who had undergone ovarian cancer surveillance (median age, 43) completed an in depth telephone interview consisting of open-ended questions. RESULTS: Most commonly cited concerns before prophylactic oophorectomy included the physical discomfort of surgery and recovery (40%) and issues of immediate menopause and hormone replacement (37%). Fourteen women (47%) would have liked more information prior to surgery. Two women (7%) expressed regret about their decision. The remaining 28 women (93%) undergoing prophylactic oophorectomy expressed no regret about the decision. Nine women (37%) would have liked more information prior to considering ovarian cancer surveillance. Nearly half the women undergoing surveillance did not recall receiving any information about prophylactic oophorectomy as an option. Fifteen women (50%) expressed some regret about ovarian cancer surveillance, and three were frankly dissatisfied. CONCLUSION: Few women undergoing prophylactic oophorectomy had regret about their decision, though half these women would have liked more information prior to surgery. Many women undergoing ovarian cancer surveillance had some regret about or dissatisfaction with their decision. PMID- 11255822 TI - In vitro fertilization and embryo transfer during natural cycles. AB - OBJECTIVE: To report the results of in vitro fertilization and embryo transfer (IVF/ET) performed during natural cycles. STUDY DESIGN: A prospective clinical study. RESULTS: Thirty-two cycles were started in 19 patients who had regular ovulatory cycles and tubal factors or unexplained infertility only as the cause of infertility. Egg collection was performed in 12 cycles, and four pregnancies resulted from ET in eight cycles. The pregnancy rates were 12.5% per cycle initiated, 33.3% per retrieval cycle and 50% per transfer. CONCLUSION: Natural cycle IVF/ET offers a low-cost alternative to patients with infertility. PMID- 11255823 TI - Caring for women with childbearing potential taking teratogenic dermatologic drugs. Guidelines for practice. AB - OBJECTIVES: Upon completion of this article, the reader should be able to: 1. Identify the dermatologic drugs with the highest risk of teratogenicity. 2. List the essential information to be obtained from a woman with childbearing potential being considered for treatment with a teratogenic dermatologic agent. 3. Discuss management principles for administration of teratogenic agents to women with childbearing potential. Dermatologic agents can be highly teratogenic, and the gynecologist must be aware of the issues involved in prescribing them. Prescribers should be aware of the risks and ensure that patients taking teratogens do not become pregnant during the course of therapy (or until the drug has been cleared from the body). This can involve taking menstrual and sexual histories and counseling the patient about sexual behavior and contraception. PMID- 11255824 TI - Taking a sexual history from and counseling women on teratogenic drugs. AB - OBJECTIVES: After studying this article, the reader should be able to: 1. Describe the relationship of sexual history taking and counseling to the safe prescription of teratogenic drugs. 2. Define the scope of the problem of unintended pregnancy and the importance of sexual history taking and counseling. 3. Review the skills of sexual counseling and taking a sexual health history. Although the need to take a sexual history and counsel patients about conception and contraception appears in several clinical situations, the process can be uncomfortable, and some physicians may avoid it or do it incompletely. When a patient is receiving a teratogenic agent, the need for proper history taking and counseling is critical. This article reviews ways to make the process more comfortable and rewarding for both the patient and physician. PMID- 11255825 TI - Contraception. Myths, facts and methods. AB - OBJECTIVES: After reading this article, the reader should be able to: 1. Recognize the mechanism of action, side effects, contraindications, precautions and instructions for use of a variety of contraceptive methods. 2. Understand the advantages and disadvantages of the various contraceptive methods. 3. List the common myths and misconceptions about conception and contraception, and recognize how they can influence contraceptive decisions. Unintended pregnancy is a serious problem in the United States. Counseling a patient about conception and contraception involves more than simply imparting information and answering questions. Clinicians should actively detect and correct any myths and misapprehensions on the patient's part. These myths are quite common and can interfere with treatment if not attended to. This article summarizes common myths about pregnancy and contraception and reviews the key facts about both. PMID- 11255826 TI - "Be smart, be safe, be sure". The revised Pregnancy Prevention Program for women on isotretinoin. AB - OBJECTIVES: After studying the information in this article, the reader should be able to: 1. Describe the purpose of the pregnancy prevention program. 2. Discuss the five most common reasons for unintended pregnancy. 3. List the components of the expanded pregnancy prevention program. Preventing unintended pregnancy is currently an unsolved problem in the United States, especially among teens. However, successful programs to minimize unintended pregnancy do exist and can serve as a model for other efforts. One such program is the Pregnancy Prevention Program, for use when prescribing isotretinoin to women with childbearing potential. Isotretinoin is a known teratogen and is prescribed disproportionately to teens, who are at higher risk of unintended pregnancy. The program has shown impressive effectiveness despite these handicaps, but since exposure to isotretinoin is so harmful to the fetus and some women still become pregnant while taking the drug, the program has been revised to reduce the failure rate further. PMID- 11255827 TI - Divided Congress faces challenging future. PMID- 11255828 TI - Who's covered by HIPAA? PMID- 11255830 TI - A visit to the cutting edge. PMID- 11255829 TI - Using physician profiling to improve documentation. PMID- 11255832 TI - Who should have access to your information? Privacy through the ethics lens. PMID- 11255833 TI - Crossing the spectrum: steps for making ethical decisions. AB - Resolving ethical dilemmas is never an easy process, but you can take certain steps to simplify it. The author provides a framework for addressing ethical issues that is useful to both organizations and individuals responsible for dealing with ethical challenges. PMID- 11255834 TI - Long-term care by the book: AHIMA releases practice standards. PMID- 11255835 TI - A diversity challenge: understanding cultural differences and communication. AB - Managing a diverse work group offers many challenges--especially when it comes to communication. As the HIM work force becomes more diverse, managers need to address this issue. The authors offer some strategies for better communication. PMID- 11255836 TI - From DNA to data privacy. AB - Genetic research is revolutionizing healthcare by predicting diseases and revealing cures. At the same time, the information presents new challenges to HIM professionals charged with keeping this vital data from the wrong hands. Here's a look at the role genetic information is playing in healthcare and what you need to know about keeping it private. PMID- 11255837 TI - Extenuating circumstances occasionally make physician record completion impossible. What is the recommended practice for retiring incomplete medical records? PMID- 11255838 TI - New PPS proposed for inpatient rehabilitation facilities. PMID- 11255839 TI - How to build a winning team. PMID- 11255840 TI - Practice brief. Documentation requirements for the acute care inpatient record. American Health Information Management Association. PMID- 11255841 TI - Committee considers ICD-9-CM changes for 2002. PMID- 11255842 TI - Ethical coding in the physician office. PMID- 11255843 TI - On the right side of the law. Interview by Anne Zender. PMID- 11255844 TI - Intimate partner violence: a health system's response. PMID- 11255845 TI - Organ donation program manages competing demands. PMID- 11255847 TI - [Heard but unable to meet]. PMID- 11255846 TI - Change in social work service in the VA system: tracking managers' perceptions. PMID- 11255848 TI - [New cardiac markers improve the diagnosis of infarction]. PMID- 11255849 TI - [Breech presentation--now what?]. PMID- 11255850 TI - [Lung cancer]. PMID- 11255851 TI - [Lung cancer]. AB - INTRODUCTION: The aim of this study was to examine our lung cancer patients, particularly their smoking habits and survival rates. MATERIAL AND METHODS: Information was recorded prospectively on 219 patients seen in our hospital from 1987 to 1992. RESULTS: Average age was 65.5 years (39-85 years). 27% were female. 9% were 49 years or younger, 13 men and seven women. Only six patient (3%) were non-smokers, 77% were smokers and 20% ex-smokers. 52 patients (24%) had small cell lung cancer (SCLC) and 167 (76%) non-small cell lung cancer (NSCLC). All six non-smokers had adenocarcinoma (two bronchioloalveolar cell carcinoma). 63% had advanced disease (stage IIIB and IV or small cell cancer, extensive disease). Only 22 patients (10%) were alive at five years: 19 patients with NSCLC who had got surgical treatment, one patient with NSCLC (stage IIIB) who had recieved radiotherapy, and two patients with SCLC, limited disease, treated with chemo- and radiotherapy. The surgically treated patients (23%) had a five-year survival of 50% (17/33) at stage I, 33% (2/6) at stage II, and none (0/7) at stage IIIA. INTERPRETATION: This poor outlook for patients with lung cancer and the fact that there are few non-smokers among them, tell us that the major emphasis must be on prevention. PMID- 11255852 TI - [Application of tobacco smoking regulations in restaurants in Tromso 1998]. AB - BACKGROUND: Under the Norwegian Environmental Tobacco Smoke Act, a minimum of 50% of tables in restaurants have to be in smoke-free areas. The Ministry of Health and Social Affairs has defined "smoke-free restaurants" as a priority objective in its anti-tobacco strategy. MATERIAL AND METHODS: We have investigated smoking policies in restaurants in the City of Tromso in Northern Norway, as reported by restaurateurs in 1998. Representatives of all the 85 restaurants, bars and pubs in the city were interviewed and their smoking policies and habits reported. This study was part of the local health authority's evaluation of the degree of compliance with the legislation. RESULTS: In 71% of establishments, at least 50% of tables were smoke-free; in 88%, smoking areas were in compliance with the legislation. 86% of restaurateurs reported a positive or neutral attitude to the legislation, 80% thought that their guests were of the same opinion. 47% smoked every day; however, there was no association between smoking habits and smoking policies. INTERPRETATION: Though the prevalence of smoking was high among restaurateurs, this did not affect their attitudes towards the Environmental Tobacco Smoke Act or their policies on smoking. PMID- 11255853 TI - [New cardiac markers--clinical benefits in early diagnosis of acute heart disease]. AB - BACKGROUND: New cardiac markers that may be analysed around the clock in emergency can now be performed in our hospitals with commercially available reagents and equipment. Upon the introduction of a new clinical biochemical regime based on these new markers for the diagnosis of acute coronary syndromes, we evaluated the clinical benefit achieved by the new set-up, especially with respect to early diagnosis. MATERIAL AND METHODS: cTroponinT, CK-MBmass, myoglobin and total-CKactivity were analysed in blood sample taken on admission, after 2-3 hours, and further once or twice over the next 24 hours in 300 patients admitted on suspicion of acute coronary syndromes (ACS). The study was based on results of the cardiac markers and information given on questionnaires by the physicians in charge. RESULTS: With the decision limits applied, CK-MB and myoglobin showed slightly higher sensitivity than cTroponinT for detecting acute myocardial infarction within the first 2-3 hours. cTroponinT showed the highest sensitivity for detecting heart muscle damage in patients with unstable angina. cTroponinT was the most cardiospecific marker. If the patient was considered not having ACS after the first few hours, only 3% ended with a diagnose of unstable angina and none with acute myocardial infarction. Of those considered certain ACS cases after the first few hours, 92% ended up with the diagnosis acute myocardial infarction or unstable angina. Treatment and/or supervision were changed in 68 of 220 patients based on the results of the two first blood samples, 85% of them to a lower level of supervision. INTERPRETATION: cTroponinT and CK-MB are useful markers for early detection of acute myocardial injuries. A prerequisite is that they are determined in two samples with an interval of at least two hours. In this case, myoglobin did not give additional information. Based on the results from two early blood samples, about one quarter of the patients could immediately be transferred to a less expensive level of care. PMID- 11255854 TI - [Myocardial injury in acute stroke assessed by troponin I]. AB - BACKGROUND: Acute ischaemic or haemorrhagic cerebrovascular events may produce myocardial damage. Cardiac troponin I is an indicator of cardiac cell injury with very high sensitivity and specificity. MATERIAL AND METHODS: We measured troponin I in 149 acute stroke patients admitted to the stroke unit of Trondheim University Hospital, Norway, in January to June 1999. RESULTS: 40 patients (27%) had troponin I values at 0.4 microgram/l or higher, indicating myocardial injury. 10 patients (6.7%) had troponin I values above 2.0 micrograms/l. Similarly, the mean value of CK-MB vas higher in the patients with myocardial injury, and these patients had more often ECG findings suggesting myocardial ischaemia. Patients with myocardial injury had a higher rate of previous TIA and heart failure. ECG showed atrial fibrillation in 13 of 39 patients with myocardial damage. Patients with detectable levels of troponin I had more embolic brain infarctions than thrombotic brain infarctions. Patients with myocardial injury did more often have abnormal values of CRP. 9 of 10 patients with troponin I-values above 2.0 micrograms/l had abnormal CRP values. No differences in glycosylated haemoglobin, cholesterol, heart rate, blood pressure or body temperature were found. Patients with the highest troponin I values had lower systolic blood pressure, and a higher heart rate, but these differences were not statistically significant. Patients with troponin I values above 2.0 micrograms/l had lower functional and neurological scores at admittance. Patients with myocardial injury were more often discharged to nursing homes. INTERPRETATION: Many patients with an acute stroke have at the same time a myocardial injury, determined by elevated troponin I values. PMID- 11255855 TI - [Hereditary neuropathy with pressure palsies]. AB - BACKGROUND: Hereditary neuropathy with liability to pressure palsies (HNPP) is an autosomal dominant polyneuropathy usually caused by a deletion in the gene coding for the peripheral nerve myelin protein 22 (PMP22). The patients usually get relapsing and remitting focal nerve symptoms due to mechanical factors like pressure or minor trauma that normal nerves tolerate. MATERIAL AND METHODS: Two patients from different families have been examined clinically, neurophysiologically and genetically by Southern blot and PCR techniques. RESULTS: The clinical and neurophysiological findings were typical of this disorder, and the DNA tests showed deletions in the PMP22 gene. INTERPRETATION: We discuss clinical, neurophysiological and molecular diagnostics, pathomechanisms, treatment and secondary prevention. Early diagnosis may be important for optimal management of the patients. PMID- 11255856 TI - [Surgery in chronic pulmonary embolism]. AB - BACKGROUND: Chronic lung thromboembolism may develop after episodes of acute lung emboli. Instead of being resoluted, the thromboembolic material is incorporated into the walls of the lung arteries causing obstruction of the blood flow. Secondarily, pulmonary hypertension develops and patients experience increasing dyspnoea on exertion. In chronic pulmonary thromboembolism, drug therapy is of little benefit. Surgical thromboendarterectomy of lung arteries has emerged as an effective treatment for these very sick patients. MATERIALS AND METHODS: This paper discusses our experience with seven patients who underwent pulmonary thromboendarterectomy during the five-year period 1995-99 at our institution. The preoperative and postoperative haemodynamic evaluation of all patients were similar. RESULTS: For five patients the working capacity was significantly improved, one had limited symptomatic effect, and one died perioperatively due to massive surgical bleeding. INTERPRETATION: Pulmonary thromboendarterectomy may be indicated in selected patients with severe dyspnoea due to pulmonary hypertension secondary to chronic pulmonary thromboembolism. PMID- 11255857 TI - [Spinal cord injury following knife stab wound]. AB - BACKGROUND: Problems about penetrating injuries are well known, but spinal cord damage is rare. Stab wounds to the spinal cord may be a new type of injury in our society. MATERIAL AND METHODS: We describe two patients brought to our hospital with stab wounds to the cervicothoracic region and major neurologic injury. One was treated initially only with cleaning and primary closure of the skin. After two weeks the intraspinal damage was repaired in our neurosurgical unit. The second patient was immediately brought to neurosurgery. The intraspinal damage was explored and the dural tear was closed. RESULTS: The first patient got a superficial infection and spinal fluid leakage after initial treatment. This resolved when the dura was closed. There were no wound complications in the second patient. Both demonstrated Brown-Sequard's syndrome. Neurologic recovery was much better in the first than in the second patient. INTERPRETATION: Minor penetrating wounds in the neck region may represent damage to the spinal cord with major neurologic injury. Further investigation with MR is preferable and we recommend immediate surgical treatment with closure of the dural tear. PMID- 11255858 TI - [Fetal alcohol syndrome--unnecessary suffering which has not become rarer. Eyes are affected in up to 90 per cent of the cases]. AB - Fetal alcohol syndrome, FAS, is characterized by pre- and/or postnatal growth deficiency, CNS dysfunction, typical facial features and evidence of prenatal alcohol exposure. FAS is often an unrecognized diagnosis and is based on clinical symptoms alone. The cognitive and behavioral disturbances have a great influence on the children's ability to learn and on their adult life. FAS is to a great extent found in areas with less than satisfactory socio-economic conditions. Since ophthalmologic involvement is frequent, an eye examination may be helpful in making a diagnosis. Prevention is essential since the brain damage prevails, despite comprehensive medical and social supportive efforts being made during childhood and in education. PMID- 11255859 TI - [Three cardiac mysteries--stunning, hibernation and ischemic preconditioning]. AB - BACKGROUND: Cardiovascular research has led to the identification of three new and important phenomena: myocardial stunning, myocardial hibernation, and ischaemic preconditioning. Myocardial stunning is characterised by transient contractile dysfunction that persists after reperfusion despite the absence of irreversible damage and despite restoration of normal or near normal coronary blood flow. Myocardial hibernation is a condition of sustained reduction of contractile function in hypoperfused but viable myocardium, which recovers completely upon reperfusion. Ischaemic preconditioning refers to a phenomenon by which one or more brief periods of myocardial ischaemia increases the ischaemic tolerance against infarction by endogenous adaptive mechanisms. MATERIAL AND METHODS: Current relevant literature obtained through PubMed search is reviewed with emphasis on occurrence of the phenomena, the therapeutic potential, and the underlying mechanisms. RESULTS: Several observations indicate that myocardial stunning, myocardial hibernation, and ischaemic preconditioning may occur in patients with coronary heart disease. Actually, an increasing amount of evidence indicates that these phenomena are of major importance with regard to myocardial ischaemic tolerance. The mechanisms underlying these phenomena are, however, not yet clarified. INTERPRETATION: A better understanding of the mechanisms underlying myocardial stunning, myocardial hibernation, and ischaemic preconditioning may provide a rational basis for development of therapeutic interventions that increase myocardial ischaemic tolerance. PMID- 11255860 TI - [Extrahepatic manifestations in hepatitis C. Are they overlooked?]. AB - BACKGROUND: There are accumulating documentation of autoimmune mediated extrahepatic manifestations of hepatitis C virus infection. The virus is hepatotrophic and lymphotrophic. It mutates frequently with subsequent inadequate immune response and chronic stimulation of T and B cells. This may be one explanation for the increased frequency of the autoimmune diseases associated with hepatitis C virus infection. MATERIAL AND METHODS: In this review of the literature published in the period of 1990 to 2000, we present the most common extrahepatic manifestations of the hepatitis C virus infection. RESULTS: Mixed cryoglobulinaemia, membranoproliferative glomerulonephritis and membranous glomerulonephritis are highly associated with hepatitis C infection. Other autoimmune diseases may also be associated with hepatitis C infection, but further documentation is necessary. INTERPRETATION: Extrahepatic manifestations of hepatitis C virus infection are associated with several autoimmune diseases. When diagnosing an autoimmune disease, a test for a coinfection of hepatitis C is highly recommended. Antiviral therapy with interferon may in some cases reduce the activity of the autoimmune disease. PMID- 11255861 TI - [Auscultation of the lungs--still a useful examination?]. AB - Auscultation of the lungs has been a central element in clinical examination since the early part of the nineteenth century. However, the role of the stethoscope in our diagnostic work-up has more and more been challenged by newer diagnostic equipment. Research carried out over the last 30 years has given us new knowledge about the physical basis of lung sounds and the meaning of the sounds. Electronic stethoscopes and computer-based analysis of digital lung sounds are now available. Lungs auscultation findings should be interpreted with caution and be related to the case history and other clinical findings. PMID- 11255862 TI - [Can use of antibiotics in acute bronchitis be reduced?]. AB - BACKGROUND: Acute bronchitis is one of the most common illnesses treated in the primary care setting. Most patients are treated with antibiotics, despite the fact that acute bronchitis is often a viral infection. There is little evidence that antibiotics are of any value in the treatment of this illness. Inappropriate use of antibiotics in the treatment of this and other infectious diseases contributes to the development of resistant bacteria. The purpose of this study was to reduce the prescribing of antibiotics for acute bronchitis in patients without underlying lung disease in the acute care clinic in Arendal, Norway. MATERIAL AND METHODS: The study had three phases, the first of which was a pilot study showing that 87% of patients with acute bronchitis received a prescription for antibiotics. The next phase was an educational intervention in which the physicians were informed of the inappropriately high prescribing rate, the lack of evidence that antibiotics are useful in the treatment of acute bronchitis, and the potential of C reactive protein (CRP) in the diagnose of this illness. The third phase of the study examined the treatment given to patients after the intervention. RESULTS: The antibiotic prescribing rate was reduced from 87% to 71% after the intervention. Doxycycline was prescribed most often, followed by penicillin and erythromycin. The use of CRP increased, and the rate of antibiotic prescriptions for patients with CRP < or = 20 was reduced after the intervention. There were fewer bronchitis diagnoses and more pneumonia diagnoses made after the intervention. INTERPRETATION: An educational intervention designed for prescribing doctors may reduce the use of antibiotics in the treatment of acute bronchitis. PMID- 11255863 TI - [Should iron preparations be available only by prescription?]. AB - Many persons associate fatigue and lassitude with iron deficiency and take extra iron "to be on the safe side". This is an unfortunate practice, as the early symptoms of iron deficiency anaemia and of hereditary iron overload (homozygous primary haemochromatosis) are similar. Primary haemochromatosis is considerably more prevalent than earlier believed. As many as 5 per 1,000 of the Norwegian population may have two mutated genes for haemochromatosis, while up to 15% may be carriers of a single mutated gene, and for these an extra intake of iron may be hazardous. The condition is highly underdiagnosed. In Norway at present, iron preparations of 60-100 mg are sold over the counter in pharmacies without prescription and often by self-service. However, no one should use iron tablets until iron deficiency and its cause has been ascertained. To avoid uncritical use of iron, iron preparations should be available only by doctor's prescription. Prolonged abuse of iron tablets may result in secondary haemochromatosis. PMID- 11255864 TI - [Hjalmar Schiotz and his tonometer]. AB - Hjalmar August Schiotz (1850-1927) was the first Norwegian professor of ophthalmology. Drawing on his rare technical skills and deep insight into advanced mathematics and physiological optics, he designed several ophthalmological instruments, the two most important being the ophthalmometer he designed together with Professor Javal in Paris in 1881 and his own tonometer in 1905. Throughout more than 50 years, the Schiotz tonometer upheld its position as the instrument most widely used world-wide for measuring intraocular pressure. PMID- 11255866 TI - [Gene therapy in HIV infection and other viral infections]. AB - BACKGROUND: Considerable problems in the management of HIV infection remain, even in the Western world. Not all patients respond satisfactorily to expensive highly active antiretroviral therapy (HAART). Relapses are frequent; often, but not always related to drug resistance, and side effects may be serious. New therapeutic strategies are therefore needed. MATERIAL AND METHODS: A systematic survey was made of protocols for gene therapy available in major databases, as well as of publications on gene therapy of HIV infection and other virus infections. RESULTS: A number of strategies for gene therapy in HIV infection are theoretically possible, several of which are already in phase I trials, in a few cases also in phase II trials. Several strategies are targeting various aspects of the life cycle of HIV, while others are aiming at improving the patient's own immune response to the virus. So far no clinical results have been published. Gene therapy is also in development for certain other viruses, including various herpes viruses as well as hepatitis B and C viruses. Gene therapy for cytomegalovirus retinitis with synthetic oligonucleotides based on the antisense principle is already available in clinical medicine. INTERPRETATION: Valuable information has been gained from the studies, which will be a great value in the further development of gene therapy for HIV infection and for other viral infections. PMID- 11255865 TI - [New preparations against influenza]. PMID- 11255867 TI - [Gene transfer--ways of administration, advantages, possible unsuitability and hazards]. AB - BACKGROUND: Reports about successful gene therapy are now published after a period of more than ten years of trial and error. The key problem is to get DNA from genes or gene fragments into the target cells to be transcribed. MATERIAL AND METHODS: A brief review of transfer techniques is given, based upon the authors' own research and the literature in the field. RESULTS: In most cases, a vector (modified virus DNA or RNA, or plasmid DNA) is used as a vehicle. Retrovirus (RNA virus), adenovirus (DNA virus) and adeno-associated virus (DNA virus) are frequently used. Non-viral vectors such as plasmid DNA, liposome linked DNA, protein DNA conjugates and artificial chromosomes are also relevant. Retrovirus has been used in about 60% of all gene therapy protocols. One problem is how to produce enough modified retrovirus for clinical use, hence retrovirus has mainly been used in ex vivo gene therapy, in which the number of target cells to be infected with the vector is limited and much lower than in in vivo therapy. INTERPRETATION: Increased insight into the genome has taught us that genes can partially or totally replace each other with regard to function, but they will not be expressed at the same time in the patient's life or in the same organ. In the future it may not be necessary to transfer a new gene; instead we may interfere with the regulation of another gene with a similar function. PMID- 11255868 TI - [Gene therapy in cancer]. AB - BACKGROUND: There has been a substantial increase in our understanding of the pathogenesis of cancer over the past decades, and new treatment modalities are being developed on the basis of this knowledge. MATERIAL AND METHODS: The literature is reviewed, and the status of gene therapy protocols approved in Norway is presented. RESULTS: About 70% of the more than 400 clinical gene therapy studies started are targeted at cancer. Several different principles are used, including gene therapy targeted at tumour suppressor genes, oncogenes, or central signaling molecules, as well as "suicide gene" therapy. In addition, various gene therapy protocols aim at strengthening immune responses. Most studies have been early clinical trials primarily designed to study safety, applicability and toxicity. Several of these phase I and II studies have, however, shown partial remission of tumours and, in rare cases, complete remission, although curation has not yet been shown. In some trials, including TP53 gene therapy trials, effects on tumour size have been observed in up to 50% of the patients. Up until now, only two phase III and one phase II/III studies have been initiated, but results from these studies have not yet been published. The two first gene therapy protocols approved in Norway are also targeted at cancer. So far, six patients in Norway have undergone gene therapy. INTERPRETATION: As of today, the results should be seen as promising for some of the principles which are being tried out; their clinical importance must, however, be documented in larger controlled clinical trials. PMID- 11255869 TI - [Gene therapy in cardiovascular diseases]. AB - Cardiovascular disease is the most frequent cause of death in the western hemisphere. Although significant advances in pharmacotherapy have been achieved, the morbidity and mortality of cardiovascular disease will probably remain a scourge of affluent societies for many years to come. This situation gives the scientific community and the pharmaceutical industry an impetus to develop novel therapeutical strategies. Gene therapy is a principle which may provide new means of targeting the pathophysiological mechanisms of cardiovascular disease and has recently drawn a lot of scientific attention, as the endothelial cells of the vasculature are in direct contact with the blood. Thus, many of the obstacles which are thought to hamper gene delivery may not apply in this system. Although there are many ongoing gene therapy trials in this patient group, the results that have so far been reported are from early phase studies (phase I and II studies) only. The published reports have investigated to what extent the growth factors vascular endothelial growth factor and fibroblast growth factor may improve revascularization in ischaemic myocardial tissue. Although the preliminary results are promising, it should be pointed out that phase III trials have not yet been started in this disease group. PMID- 11255870 TI - [Comparison of "the two cultures" in medicine. Scientific theory as a pedagogic and therapeutic tool]. PMID- 11255871 TI - [Is the time of compassion over?]. PMID- 11255872 TI - [Braatoy and psychomotor physiotherapy]. PMID- 11255873 TI - [Treatment of cerebral paresis--rebuff for Norway]. PMID- 11255874 TI - [Better trauma therapy--BEST or ATLS?]. PMID- 11255875 TI - [Abortion committees in Rogaland better than the recently reported]. PMID- 11255876 TI - [Risk of cancer with snuff?]. PMID- 11255877 TI - [Zyban in smoking cessation]. PMID- 11255878 TI - [Evolution of the catheterization laboratory: new instruments and imaging techniques]. AB - The Editorial Board of the Italian Heart Journal Supplement has planned to publish a series of four consecutive papers focusing on new highlights from the cardiac catheterization laboratory, developments of new devices, coronary imaging and physiological measurement. The first paper on cineless digital angiography and new fluoroscopy systems presented in this issue of the Journal has been prepared by Danzi et al. (Brescia). The paper follows the publication of the Vergara's work (Rovereto-TN) on a similar topic presented last year in a Symposium of the Gruppo Italiano di Studi Emodinamici (GISE) at the XXXI ANMCO meeting. The recent development of cineless digital angiography and the increasing interest in new digital fluoroscopic low energy systems represent a significant change in the management of cardiac images and have significant implications in the organization of a new catheterization laboratory. The second paper will focus on the clinical applications of intravascular ultrasound imaging and will be prepared by Di Mario (Milan). Intravascular ultrasound imaging has significantly contributed to the understanding of vascular adaptation to coronary artery disease and mechanisms of percutaneous coronary interventions. This new diagnostic approach is particularly useful in assessing the early stages of disease, evaluating occult atherosclerosis and guiding the strategy of coronary interventions. Unfortunately, the application of intravascular ultrasound imaging in clinical practice remains limited. The third paper prepared by Verna (Varese) will discuss the new techniques for physiological evaluation of coronary artery disease now available in the catheterization laboratory. The use of pressure and Doppler flow-wire in the assessment of functional significance of individual coronary artery stenoses and in the evaluation of the vasodilatory capacity of coronary microcirculation may provide new insights for the clinical decision at the catheterization table. In addition, the application of myocardial contrast echocardiography with new sonicated agents for intracoronary use may represent a novel approach to the direct evaluation of regional myocardial perfusion and microvascular integrity. Intracoronary stenting has become a widely used and accepted technique in percutaneous coronary interventions. The fourth article of this series, prepared by Silva (Milan), will discuss the advantages and limitations of stenting in different clinical settings based on the results of large clinical trials. In spite of the growing number of procedures safely performed in many countries, long-term results of intracoronary stenting may be sometimes disappointing in selected patients. In-stent restenosis still represents a clinical problem limiting long-term outcome. In addition, the increasing use of new antiplatelet agents in combination with most stenting procedures rise some concern about overall cost of interventions. The objective of this editorial program is to give a view of the changing scenario of the catheterization laboratory with the new methods and devices presently available. There may be significant implications and controversial issues of interest also for all clinical cardiologists. PMID- 11255879 TI - [New coronary imaging modality. I. Digital angiography (cineless) and low-energy digital fluoroangioscopy systems]. AB - The advent of digital medical imaging offered unique new possibilities of analyzing, visualizing and communicating medical images. This article reviews the impact of the digital technology in the cardiac catheterization laboratory and covers a range of topics such as the standard DICOM, the transition to cineless angiography, the digital cardiac archive, the network system for imaging exchange and the role of the cardiac digital mobile imaging systems. PMID- 11255880 TI - [Cardiovascular risk factors and prevention in women: similarities and differences]. AB - Epidemiological evidence shows that among women, the incidence of all, including less severe, coronary events is still increasing. However, owing both to diminished lethality as well as the reduction in the rate of acute myocardial infarction, mortality has globally decreased. The strong association observed between mortality and major cardiovascular risk factors as well as between their temporal changes and the occurrence of coronary disease makes the undertaking of multifactorial prevention strategies, including the formulation of risk charts for asymptomatic women and men, necessary. The different "penetrance" of risk factors in women, together with their interaction with female hormones, plays an important role in the development of cardiovascular disease. The excess risk of cigarette smoking is 2-4 times higher in women than in men and the correlation with the number of cigarettes smoked daily is distinct. However, the risk starts to decrease immediately after stopping and after 3-5 years is similar to that of non-smokers. In women, the association between hypertension, coronary artery disease and early mortality is stronger than in men: there is no threshold below which the risk disappears. Diet and lifestyle strongly influence the development of hypertension. For this reason, the American Heart Association/American College of Cardiology guidelines recommend adherence to a set of dietary and lifestyle habits including body weight control and physical activity. In particular, diet may modify the "penetrance" of risk factors in women: hence excess intake of saturated fatty acids associated with decreased cereals, fruit and vegetables does not only alter the lipid profile but also increases the risk of coronary disease. An elevated total/HDL cholesterol ratio and the presence of lipoprotein(a) constitute significant risk factors for coronary events. On the other hand, high HDL cholesterol levels (> 45 mg/dl) are considered to be protective in women. However, data on the efficacy of strategies aimed at reducing blood LDL levels in hypercholesterolemic women are limited and controversial. Pharmacological therapy is recommended in women with primary familial hypercholesterolemia and during menopause when the patient presents with two or more risk factors. Besides, pharmacological therapy is also indicated for women with a history of coronary artery disease in whom benefits exceed those observed in male patients with a similar clinical picture. In diabetic women, the risk of coronary mortality is increased 3 to 7-fold compared to the 2 to 3-fold increase observed in diabetic men. Diabetes definitely increases the effects of the other risk factors and modifies the protective effect by estrogens. However, to date, there is no evidence that keeping glucose levels within normal limits reduces the risk of coronary artery disease nor has a glycemic threshold capable of predicting mortality risk in diabetic women been established. For this reason, guidelines for such patients are aimed at keeping the other risk factors under strict control in order to significantly reduce their effect. Obesity results in a series of metabolic alterations that increase the risk of cardiovascular disease in both sexes. Although most, if not all, data confirm that obesity alone is not of predictive value, central obesity constitutes a risk factor for cardiovascular disease. A body mass index < 24.9 kg/m2 and a waist circumference < 80 cm are recommended so as to decrease the likelihood of developing a menopausal insulin-resistance syndrome. It has been demonstrated that in men, a sedentary lifestyle is correlated with a higher cardiovascular and all-cause mortality; some recent observational studies suggest a 25-30% decrease in the mortality risk for women who perform physical exercise. Current guidelines recommend at least 30 min daily of dynamic moderately vigorous activity, including brisk walking. Rather than to the reduction in the serum levels of endogenous estrogens, the increase in the incidence of disease and of mortality following menopause should be attributed to the age-related modifications in risk factors which result in an increased risk of coronary artery disease. In spite of the proved detrimental effect of estrogen deficiency on LDL- and HDL-cholesterol, on arterial smooth muscle cell proliferation and on insulin secretion and in spite of the data of numerous observational studies and of the HERS trial (all, however, with methodological limitations), clinical evidence does not justify widespread estrogen prescription, not even for purposes of secondary prevention. Besides, the dosages and the route of administration are still subject of debate. (ABSTRACT TRUNCATED) PMID- 11255881 TI - [Digital signature: new prospects for the information of the cardiologic clinical card]. AB - In the last few years, remarkable improvements have been made in computerized database systems used in cardiology. However, they will not easily lead to further relevant improvements unless the weaknesses and the gaps deriving from the obligation of forming and storing case sheets, according to law, are faced and resolved in an original way. This article covers the topic of the digital signature and how it could form the basis for a new powerful impulse to the process of informatization of cardiology records. The proposal of elaborating a totally computerized case sheet involves the need of rationalizing the flow of clinical information and of implementing a management system integrated with the hospital information system. The elimination of paper support will probably lead to an advantageous cycle that will involve the entire hospital, both clinically as well as administratively. PMID- 11255883 TI - [Dynamic electrocardiogram and syncope: a couple in real crisis?]. AB - Holter monitoring is still considered as a class I diagnostic tool for the assessment of syncope. This is due to poor yield of the diagnostic techniques available until the middle 1980s. However, considering the impressive progress made in the differential diagnosis of this clinical condition, such widespread use is no longer justified. The limitations of Holter monitoring are highlighted by its costs per diagnosis and, in particular, by comparison with other maneuvers, such as the head-up tilt test or the carotid sinus massage, which have a higher positive predictive value for patients with syncope of unknown origin. The diagnostic yield of Holter monitoring has to be considered particularly low among patients free from structural heart disease in whom the risk of arrhythmic syncope is extremely low. PMID- 11255882 TI - [Limits of cardiac functional adaptation in "top level" resistance athletes]. AB - BACKGROUND: Sports activity, particularly when performed at high level, provokes cardiovascular adjustments depending on the type of sport and on the level of the load. METHODS: We evaluated 15 athletes from the Italian national team during a non-agonistic period of cross country skiing, with non-invasive tests including exercise test, color Doppler echocardiography, Holter monitoring, physical examination and standard rest electrocardiogram. RESULTS: Physical examination, rest electrocardiogram, exercise testing and echocardiography were all within the range of the expected values for this type of subjects. Holter monitoring recorded during the periods of agonistic activity revealed significant hypokinetic arrhythmias such as severe bradycardia, pauses, I and II degree atrioventricular blocks, and complete atrioventricular block in 2 cases; these features were not observed on Holter monitoring recorded during the non-agonistic period. CONCLUSIONS: The perfect health status of subjects and their racing results may bring about physiological functional adjustments, but these observations suggest the need for a follow-up to evaluate possible pathologic outcomes. PMID- 11255884 TI - [Posterior echocardiography windows: usefulness in clinical practice]. AB - Posterior cardiac structures cannot always be imaged by means of standard transthoracic echocardiography. Left pleural effusion leading to pulmonary atelectasis and/or to displacement of air-filled pulmonary tissue displacement, allows ultrasound transmission from a patient's back to his heart through a liquid interface. In this study we present the clinical usefulness of echocardiographic posterior windows for the diagnosis of constrictive pericarditis and aortic dissection in 2 patients in whom the standard transthoracic approach did not permit diagnostic imaging. We conclude that, in the case of left pleural effusion, the use of posterior windows should be encouraged. PMID- 11255885 TI - [Guidelines for the diagnosis and treatment of acute pulmonary embolism]. PMID- 11255886 TI - [Heart rate recovery and treadmill exercise score as predictors of mortality in patients referred for exercise ECG]. PMID- 11255887 TI - [Low-dose transdermal testosterone therapy improves angina threshold in men with chronic stable angina. A randomized, double-blind, placebo-controlled study]. PMID- 11255888 TI - [Acute hemodynamic and clinical effects of a new inotropic agent, levosimendan, in patients with severe heart failure]. PMID- 11255889 TI - [Trends in the incidence of coronary heart disease and changes in diet and lifestyle in women] [Effects of estrogen replacement on the progression of coronary atherosclerosis]. PMID- 11255891 TI - [Various reasons for not discharging early]. PMID- 11255892 TI - [Uncomplicated myocardial infarction: importance of protected early discharge]. PMID- 11255893 TI - [Changes in medical treatment of heart failure in the light of large clinical trials]. AB - In the last years, the treatment of heart failure has radically changed, as has knowledge of this complex and heterogeneous clinical syndrome. This is largely due to the results of several multicenter clinical trials, which have been undertaken since the late 80's. These trials have not only contributed to the elaboration of present-day treatment protocols, but also to a better understanding of the pathophysiologic mechanisms involved in heart failure. In the past, heart failure was generally interpreted on the basis of pathophysiologic models according to which hemodynamic abnormalities played a very important role in determining the clinical presentation and evolution of the disease. This led to the use of digitalis, diuretics, inotropic drugs and vasodilators for the treatment of heart failure. More recently, improved knowledge of the pathophysiologic mechanisms involved in the progression of this disease has highlighted the central role and the complexity of various neurohormonal mechanisms. Antagonism of these systems has proved to be the only strategy which favorably modifies the natural history of heart failure. The proved effectiveness of ACE-inhibitors and particularly of beta-blockers in patients with heart failure and left ventricular systolic dysfunction was the most convincing demonstration of the validity of this model. However, the evolution and updating of the guidelines on the treatment of heart failure should only be considered as the first step in the development of strategies aimed at extending these principles to daily clinical practice and in particular to the real patient who is different from patients typically enrolled in heart failure trials. Moreover, the development of new effective models for the management of the ever-growing number of patients with heart failure is of utmost urgency. PMID- 11255894 TI - Changing the culture? PMID- 11255895 TI - Patients with alcohol problems--simple questioning is the key to effective identification and management. PMID- 11255896 TI - Diagnosis and prognosis of lower respiratory tract infections: a cough is not enough. PMID- 11255897 TI - Symptoms, signs, and prescribing for acute lower respiratory tract illness. AB - BACKGROUND: Most patients who consult with acute lower respiratory symptoms receive antibiotics, usually without evidence of significant infection. The physical signs at presentation of acute lower respiratory tract illness and the rate at which symptoms resolve and normal activities recover is not well documented. AIM: To examine in patients with lower respiratory tract infection (LRTi), their physical signs at presentation, their relationship to antibiotic prescribing, and symptom resolution and resumption of normal activities. DESIGN OF STUDY: Analysis of data collected prospectively during presentation of acute LRTi in primary care and from patient symptom diary cards. SETTING: Forty GPs who were members of an informal Community Respiratory Infection Interest Group recruited 391 patients to the study. METHOD: Information was collected on pulse, oral temperature, respiratory rate, abnormalities on auscultation, and details of any antibiotic prescription. Patients completed symptom diary cards for the following 10 days. RESULTS: Of the 391 patients who consulted 71% received antibiotics. A minority had abnormal physical signs: 17% had a pulse greater than 90 bpm, 15% a respiratory rate greater than 20 breaths per minute, 4% had a temperature greater than 38 degrees C, and 25% had an abnormality on auscultation. Antibiotic prescribing was more common in the presence of abnormal chest signs (odds ratio = 8.71, 95% confidence interval = 3.69-20.61) or discoloured sputum (OR = 2.67, 95% CI = 1.57-4.56). Ten days after consultation, 58% of patients were still coughing and 29% had not returned to normal activities. CONCLUSION: Abnormal physical signs at presentation do not explain the high rates of antibiotic prescribing nor do they predict persisting cough and functional impairment at 10 days. Reconsultation for the same symptoms within a month is common and is strongly related to persisting cough, but not abnormalities at presentation. PMID- 11255898 TI - The Othmer and DeSouza test for screening of somatisation disorder: is it useful in general practice? AB - BACKGROUND: Somatisation disorder is an underdiagnosed and difficult problem for family physicians. Early diagnosis of somatisers is a very important factor in improving health outcomes. AIM: To assess the validity of the Othmer and DeSouza test (a seven-item questionnaire) used by general practitioners as a screening instrument for the diagnosis of somatisation disorder in primary care. DESIGN OF STUDY: A cross-sectional study of patients presenting with unexplained multiple chronic physical symptoms. SETTING: A total of 149 patients were selected for the study by 29 family physicians in the primary health care centres of the Basque Health Service in the metropolitan area of Bilbao, Bizkaia, Spain. METHODS: Participating patients completed the Othmer and DeSouza test, carried out by family physicians. Their answers were compared with the results of the Schedules for Clinical Assessment in Neuropsychiatry (SCAN). This psychiatric interview was administered blind to 144 patients by trained psychiatrists. RESULTS: A total of 19% of patients were diagnosed as having somatisation disorder by the SCAN psychiatric interview. The discriminating capacity of the Othmer and DeSouza test for all possible screening thresholds (> or = 1, > or = 2, ... > or = 6 symptoms) was very low and positive predictive values ranged between 19% and 33%. With respect to negative predictive values, even in the absence of affirmative responses to all seven questions, the pretest probability of being a non somatiser case remained unchanged. CONCLUSION: Our data indicate that the Othmer and DeSouza test does not present clinically useful predictive values in primary care patients with suggestive symptoms of somatisation disorder. PMID- 11255899 TI - Shared care: a qualitative study of GPs' and hospital doctors' views on prescribing specialist medicines. AB - BACKGROUND: Shared care schemes have mainly centred on chronic diseases, such as asthma and diabetes. However, with increasing government emphasis on primary and secondary care integration and the effects of budget restraints, general practitioners (GPs) have been asked to take on the prescribing of specialist medicines. AIM: To elicit the views and experiences of GPs and hospital doctors about existing arrangements for shared care applied to the prescribing of specialist medicines. To identify a set of quality indicators for prescribing specialist medicines at the interface between primary and secondary care. DESIGN OF STUDY: A qualitative study based on semi-structured interviews. SETTING: Forty eight GPs and 13 hospital doctors in the former South Thames region. METHOD: The interviews focused on how far experiences with shared care compare with the arrangements currently in place for prescribing specialist medicines and identified the barriers to facilitators of effective shared care. RESULTS: A number of key themes were identified and these formed the basis for eight quality indicators relating to the prescribing of specialist medicines where treatment is shared between primary and secondary care. The themes centred around issues of clinical responsibility, 'cost-shifting', availability of medicines, GP satisfaction, and the nature of the prescribing relationship. CONCLUSION: Overall, GPs appeared dissatisfied with arrangements for prescribing specialist medicines, while hospital doctors were generally satisfied. The quality indicators will form the basis of a more extensive quantitative survey of GPs' perceptions of the arrangements for prescribing specialist medicines. PMID- 11255900 TI - Risk and prevention of type II diabetes: offspring's views. AB - BACKGROUND: People with a parent with type II diabetes have an increased risk of the disease. There is increasing evidence for the possibility of prevention, particularly by attaining and maintaining normal weight and adequate levels of physical exercise. No prior studies have reported awareness of risk and prevention in this high-risk group. AIM: To explore beliefs about personal risk of diabetes and prevention in people with a parent with type II diabetes. DESIGN OF STUDY: A total of 254 adults with type II diabetes were identified from five randomly selected practices in south London. Self-report questionnaires were sent to 152 eligible offspring of these patients. A total of 105 of the offspring returned the self-report questionnaires and participated in the study. SETTING: Five randomly selected practices in south London. METHODS: Patients with type II diabetes in five randomly selected practices in south London were asked if we might contact their offspring. One randomly selected offspring (over 18 years of age) from each family completed a self-report questionnaire. RESULTS: Of 254 adults with type II diabetes 152 had eligible offspring. A total of 105 (69%) of the offspring participated in the study. A total of 69 (66%) of these offspring believed their personal risk of developing diabetes was 'low'. At least 28 (28%) and maybe as many as 73 (70%) underestimated the risk of diabetes in offspring. Compared with the number thinking their current risk was low significantly more (95 versus 69) thought that their risk would be low if neither of their parents had diabetes. Fifty-seven (54%) thought prevention was possible. Sixteen thought taking more exercise was important for prevention and only seven thought that weight control was important. Many had good general knowledge about diabetes and its complications but awareness of the relationship between diabetes and cardiovascular disease was poor. CONCLUSIONS: People with a parent with type II diabetes are usually aware that they have an increased risk of diabetes. However, they often underestimate that risk and know little about potentially useful preventive strategies. They need accurate information about these matters if they are to reduce their risk of diabetes and its complications. PMID- 11255901 TI - A randomised controlled trial of delayed antibiotic prescribing as a strategy for managing uncomplicated respiratory tract infection in primary care. AB - BACKGROUND: Despite evidence that uncomplicated lower respiratory tract infection (cough) does not respond appreciably to antibiotics and that bacterial resistance is increasing, general practitioners (GPs) still prescribe frequently. AIM: To assess delayed antibiotic prescribing as a strategy for reducing the unnecessary use of antibiotics for cough in primary care. DESIGN OF STUDY: Open randomised controlled trial of delayed versus immediate prescribing of antibiotics. SETTING: One hundred and ninety-one adult patients with uncomplicated cough in 22 Scottish practices who would have received antibiotics under the GP's usual practice were randomised to receive either an immediate prescription (92 patients) or a delayed prescription (99 patients). METHOD: Delayed subjects were asked to wait a week before deciding whether to collect their prescription. Outcome measures included symptom duration, prescription uptake, patient satisfaction, patient enablement, and subsequent consultation rates. The 48 GPs who recruited patients were surveyed six months after the trial to see whether they used delayed prescribing as a part of their normal practice. RESULTS: Study and control groups were similar at baseline. Of the subjects in the delayed arm, 55% did not pick up their prescription. Although most patients were satisfied, more patients in the immediate arm were very satisfied with the treatment (P = 0.001) and the consultation (P = 0.03). The patients in the immediate arm were also more enabled (3.3 versus 2.4; P = 0.04), although more of them intended to consult for similar complaints in the future (85% versus 69%, P = 0.02). We were unable to detect any difference in actual consulting behaviour in the follow-up period (mean = 15 months [SD = 5 months]). Subsequently, 68% of GPs used delayed prescribing at least monthly; all gave the prescription to the patient. CONCLUSION: Delayed prescribing is effective at reducing the use of antibiotics for self-limiting cough; however, patients are less satisfied and enabled as a result. Patients may be deterred from consulting rather than becoming enabled. PMID- 11255902 TI - Screening properties of questionnaires and laboratory tests for the detection of alcohol abuse or dependence in a general practice population. AB - BACKGROUND: Early identification of alcohol abuse or dependence is important in general practice because many diseases are influenced by alcohol. General practitioners, however, fail to recognise most patients with alcohol problems. AIM: To assess the diagnostic performance of the CAGE and AUDIT questionnaires, their derivatives, and laboratory tests in screening for alcohol abuse or dependence in a primary care population (male and female patients), attending their general practitioner (GP). DESIGN OF STUDY: A diagnostic cross-sectional study. SETTING: A random sample of patients who were over 18 years of age (n = 1992) attending 69 general practices situated in the same region in Belgium. METHOD: Alcohol questionnaires (CIDI 1.1, section I, CAGE, AUDIT, AUDIT-C, Five Shot, and AUDIT Piccinelli) were completed, demographic information was recorded, and patients underwent conventional blood tests, including mean corpuscular volume, liver function tests, the gamma-glutamyl transferase test, and carbohydrate-deficient transferrin (CDT, estimated using %CDT). Calculations of sensitivity, specificity, positive predictive value, negative predictive value, odds ratios with their 95% CIs, and receiver operating characteristic (ROC) curves for different scores of the questionnaires and laboratory tests, using DSM III-R as the reference standard. RESULTS: The past-year prevalence of alcohol abuse or dependence in this population was 8.9% (178/1992) of which there were 132 male and 45 female patients attending a general practice. The GPs identified 33.5% of patients with alcohol abuse or dependence. Among male patients, all questionnaires had reasonable sensitivities between 68% and 93% and hence at lower cut-points than recommended. Only the sensitivity of the CAGE, even at its lowest cut-point of > or = 1 was lower (62%). In female patients the sensitivities were lower; however, odds ratios were higher for different questionnaires. The receiver operating characteristic (ROC) curves did not differ between the questionnaires. The laboratory tests had low diagnostic accuracy with areas under the ROC curves (AUCs) between 0.60 and 0.67 for female patients and 0.57 and 0.65 for male patients. CONCLUSIONS: This is one of the largest known studies on alcohol abuse or dependence among family care practices. We confirm earlier results that the AUDIT questionnaire seems equally appropriate for males and females; however, screening properties among male patients are higher. Nevertheless, the Five-Shot questionnaire is shorter and easier to use in a general practice setting and has nearly the same diagnostic properties in male and female general practice patient populations. We confirm that conventional laboratory tests are of no use for detecting alcohol abuse or dependence in a primary care setting. Also, the %CDT cannot been used as a screening instrument in this general practice population. PMID- 11255903 TI - General practitioners' and practice nurses' knowledge of how much patients should and do drink. AB - Despite evidence linking high levels of alcohol consumption to ill health, the number of people drinking above the 'sensible' limits is increasing. Clinicians in primary care can influence this trend by appropriate screening and advice. To do this they need to know the recommended sensible limits and also be able to translate commonly reported drinking levels into units of alcohol. A postal survey of 499 general practitioners and 343 practice nurses in Cornwall and South West Devon asked responders to calculate the number of units of alcohol contained in six different drinks and also state what they thought were the current sensible levels of consumption. The response rate was 63%. Less than 40% of responders were able to assess the units of alcohol in five out of the six drinks to within 10%. Over 70% of responders were unable to determine the alcohol content of all six drinks to within 30%. Forty-four per cent of responders now recommend an increased safe level of consumption at 28 units per week for men and 21 units per week for women, against the advice of the Royal Colleges and the BMA but in line with the levels suggested by the Government. PMID- 11255904 TI - Barriers to the development of collaborative research in general practice: a qualitative study. AB - General practice-based research activity is increasing rapidly, particularly for large, collaborative, multi-centre studies. We conducted semi-structured interviews with general practitioners and other professionals at practices in the East London and the City Health Authority area, to investigate the difficulties presented by becoming involved in these studies. Interviewees' main concerns were: time constraints; team motivation; the perception that external researchers have unrealistic expectations; the need for good communications throughout and, specifically, for good feedback from these researchers. PMID- 11255905 TI - Do income questions and seeking consent to link medical records reduce survey response rates? A randomised controlled trial among older people. AB - Traditional measures of socioeconomic status may not be reliable for older people and income may be a useful measure for research into inequalities in health. At the same time, researchers increasingly wish to link survey findings to individual data taken from medical records. For this, consent must be sought. To examine whether questions on household income and seeking consent for medical record linkage affected response rates, a postal health survey of patients aged 65 to 74 was undertaken in an inner London practice. The overall response rate was 62.8%. In this study, the inclusion of an income question or seeking consent to access medical records did not reduce response rates to a health survey among older people. PMID- 11255906 TI - Do children who become autistic consult more often after MMR vaccination? AB - A close temporal association has been reported between the measles, mumps, and rubella (MMR) vaccination and dramatic behavioural decline in children subsequently diagnosed as autistic. We hypothesised that such a decline would be reflected in increased consultations with the child's general practitioner. The Doctor's Independent Network database was used to examine whether children subsequently diagnosed as autistic consulted more frequently than controls after MMR vaccination. No difference in consulting behaviour was seen in the six months post MMR. Any dramatic effect of MMR on behaviour seems unlikely. PMID- 11255907 TI - Breast feeding. PMID- 11255908 TI - Is it time to review the idea of compliance with guidelines? PMID- 11255909 TI - Do condoms actively prevent non-HIV STIs? PMID- 11255911 TI - Ageing Britain. PMID- 11255910 TI - Sexually transmitted infections in primary care: a need for education. PMID- 11255914 TI - Evidence-based medicine. PMID- 11255913 TI - Summative assessment. PMID- 11255912 TI - Summative assessment. PMID- 11255915 TI - Demands for urgent care. PMID- 11255916 TI - Medical errors and adverse effects. PMID- 11255917 TI - Abdominal pain--bedside tricks? PMID- 11255918 TI - Diagnosis and general practice. PMID- 11255919 TI - A labor-supply analysis of cocaine self-administration under progressive-ratio schedules: antecedents, methodologies, and perspectives. AB - RATIONALE: Under a progressive-ratio (PR) schedule, a subject must complete increasing fixed-ratio (FR) response requirements to obtain reinforcers. Response requirements are increased until responding stops; the final ratio completed being the "break point" and providing an index of the relative effectiveness, or value, of the reinforcer to maintain behavior. OBJECTIVES: This review examines the historical and conceptual framework underlying the PR procedure and examines the concept of relative reinforcer value. Pharmacological analysis (based on receptor theory), and behavior analysis (based on microeconomic theory) are reviewed. METHODS: Using a microeconomic adaptation of the reinforcement model referred to as conservation, a mathematical model of PR performance is proposed based on the curvilinear relationship between economic supply and labor. Drug consumption and instrumental responding were assumed to reflect deviations from a balance point, defined as the levels of consumption and responding under no scheduled restraint. Data sets were re-analyzed in which several response sequences were examined in rhesus monkeys maintained on PR schedules of intravenous cocaine delivery. RESULTS: The modified conservation equation fitted the PR data accurately, and results consistent with both linear and concave labor supply functions were obtained. These results suggest that cocaine self administration under PR schedules conforms to labor-supply relationships characterized as inelastic (consumption is resistant to increases in schedule requirements) and unit elastic (at high response costs, consumption declines with no corresponding increase or decrease in total responding). CONCLUSIONS: The labor-supply methodology allows for a definition of the relative value of a drug reinforcer in PR studies based on changes in consumption across response costs. Specifically, relative reinforcer value is defined in terms of changes in behavior from a balance point, rather than as a property that determines the strength of the instrumental response. PMID- 11255921 TI - The effects of excitotoxic lesions of the basolateral amygdala on the acquisition of heroin-seeking behaviour in rats. AB - RATIONALE: Second-order schedules of drug-self-administration provide a method of examining drug-seeking behaviour, which is maintained in part by the presentation of a discrete, drug-associated light CS. Previous results have found that lesions of the basolateral amygdala (BLA) impair the acquisition of i.v. cocaine self administration under this type of schedule. OBJECTIVES: The present experiments examined the effects of excitotoxic lesions of the BLA on the acquisition of i.v. heroin self-administration under both continuous reinforcement and second-order schedules, in order to investigate possible commonalties in the neural basis of heroin- and cocaine-seeking behaviour. METHODS: Rats received quinolinic acid or sham vehicle lesions of the BLA prior to i.v. self-administration training. Initially, heroin self-administration under a continuous reinforcement schedule was acquired. Each active lever-press resulted in a 0.04 mg i.v. heroin infusion, paired with presentation of a 20-s light conditioned stimulus. Following acquisition of responding under this schedule, the response requirement was gradually increased to a second-order schedule of FI15(FR5:S). RESULTS: There was no effect of lesions of the BLA on the acquisition of heroin self-administration under a continuous reinforcement schedule. The acquisition of heroin-seeking behaviour under a second-order schedule of self-administration was not affected by lesions of the BLA, but lesioned rats showed a significantly higher baseline level of responding. CONCLUSIONS: These results indicate that the rewarding effects of heroin do not depend on the integrity of the BLA. The BLA is also not critically involved in mediating heroin-seeking behaviour under a second-order schedule of reinforcement, and this stands in marked contrast to the effects of BLA lesions on the acquisition of cocaine-seeking behaviour. These findings suggest that discrete heroin cues were not critical in maintaining heroin-seeking behaviour under the second-order schedule used here and that other learning systems are engaged in the control of this behaviour. PMID- 11255920 TI - Reinforcing and discriminative stimulus effects of RTI 111, a 3-phenyltropane analog, in rhesus monkeys: interaction with methamphetamine. AB - RATIONALE: The neuronal actions of methamphetamine (MA) include an increase in extracellular levels of monoamines, presumably via reverse transport involving the monoamine transporters. This action is thought to play an important role in the effects of MA. Therefore, in the present experiment, it was hypothesized that a monoamine uptake blocker would block behavioral effects of MA related to its abuse. OBJECTIVE: RTI 111, a newly synthesized 3-phenyltropane analog with high affinity for the dopamine, norepinephrine, and serotonin transporters, was evaluated alone and in combination with MA for its ability to block the reinforcing and discriminative stimulus effects of MA in rhesus monkeys. METHODS: RTI 111 (0.0003-0.03 mg/kg, i.v.) was made available to four rhesus monkeys for self-administration under a fixed-ratio 25 (FR 25) schedule of reinforcement. RTI 111 (0.01-0.1 mg/kg, i.m.) was also administered as a pretreatment (15 min prior) to four monkeys self-administering MA (0.0-0.3 mg/kg per injection, i.v.) on a progressive-ratio schedule of reinforcement. MA (0.01-1.0 mg/kg, i.m.), RTI 111 (0.001-0.1 mg/kg, i.m.), or the combination of MA and RTI 111 were administered to four monkeys trained to discriminate (+)-amphetamine (AMPH; 1.0 or 1.7 mg/kg, intragastric) from saline. RESULTS: When RTI 111 was made available for self administration under an FR 25 schedule it functioned as a positive reinforcer in all four monkeys tested. When RTI 111 was given as a pretreatment to monkeys self administering MA under a progressive-ratio schedule, the MA dose-response function shifted to the left and down. When RTI 111 or MA were given to monkeys trained to discriminate AMPH from saline, full AMPH-like responding was observed for both drugs. Given in combination, RTI 111 shifted the MA dose-response function to the left. CONCLUSIONS: These data suggest that RTI 111 is behaviorally similar to traditional psychomotor stimulants that act at the DA transporter and that it increases, rather than blocks, the behavioral potency of MA. PMID- 11255923 TI - Reinforcing effects of contingently administered subcutaneous injections of etonitazene in rats. AB - RATIONALE: Response-contingent injections of opioids have been shown to control behavior in various species. OBJECTIVE: To determine whether s.c. injections of etonitazene (ETZ) could maintain behavior in rats when administered under a single fixed-interval schedule. METHODS: Rats were trained to lever press for eight 45-mg food pellets under a single fixed-interval (FI) 10-min schedule of reinforcement: following passage of the 10-min interval, each lever press resulted in a pellet delivery until eight pellets were obtained. Delivery of the reinforcer was signaled by a change in visual stimulus conditions. Once stable responding for the food pellets under the FI 10-min schedule was established, a s.c. injection of 3.2 micrograms/kg ETZ was administered to the rat by the investigator following schedule completion and delivery of the food pellets. After receiving the drug injection, rats were returned to the experimental chamber for 30 min and exposed to the same stimulus conditions that accompanied food reinforcement. Across sessions, the number of food pellets was decreased until rats were responding solely for the drug. RESULTS: Responding for the s.c. administered drug stabilized and persisted across sessions. When saline vehicle injections were substituted for the drug injections, responses diminished across sessions to levels below that of the drug baseline. Subsequent alternating blocks of ETZ and vehicle injections produced respective increases and decreases in responding. CONCLUSION: This study demonstrates that response-contingent s.c. injections of a drug can control behavior in rats, systematically replicating a previous experiment that used the i.p. route. Since all pertinent operant behavior is emitted prior to the administration of drug, this procedure can be used for testing the reinforcing effects of a drug without interference from any direct (rate-altering) drug effects. The present findings also extend the conditions under which drugs of abuse may reinforce behavior. PMID- 11255922 TI - Heroin self-administration under a second-order schedule of reinforcement: acquisition and maintenance of heroin-seeking behaviour in rats. AB - RATIONALE: Second-order schedules of heroin self-administration provide a method of measuring heroin-seeking behaviour independently of the effects of the drug on motor behaviour and of investigating the role of heroin-associated stimuli in such heroin-seeking behaviour. OBJECTIVES: These experiments aimed to establish a second-order schedule of heroin self-administration in rats, similar to that already established in this laboratory for cocaine self-administration and to investigate the role of discrete heroin-associated stimuli in the maintenance of heroin-seeking behaviour under a second-order schedule of reinforcement. METHODS: Heroin i.v. self-administration (0.04 mg/infusion) was initially contingent upon a lever press, and each infusion was paired with presentation of a 20-s light conditioned stimulus (CS). Following acquisition of heroin self-administration, the response requirement was progressively increased so that, ultimately, responding was maintained under a fixed interval (FI) 15 min [fixed ratio (FR)5:S] second-order schedule. The effects of varying the dose of heroin (0.01 mg and 0.08 mg/infusion) and pre-treatment with the mu-opiate receptor antagonist, naloxone, on responding under a FI15(FR5:S) schedule were investigated. In addition, the role of the heroin-associated CS on responding was assessed by measuring the effects of omitting the CS during heroin-seeking behaviour and during extinction of responding, as well as the effect of CS presentation on the reinstatement of heroin-seeking behaviour following extinction. RESULTS: A second-order schedule of heroin self-administration was established. There were no clear effects on heroin-seeking behaviour of increasing or decreasing the dose of heroin. Although no effect of naloxone pre treatment was seen on heroin-seeking behaviour during the first, drug-free interval of responding, an extinction-like pattern of responding was seen in that interval during subsequent sessions. Omission of the light CS resulted in a reduction in levels of responding for i.v. heroin, indicating its role in maintaining heroin-seeking behaviour. However, under extinction conditions, response-contingent CS presentations did not affect the rate of extinction, nor did non-contingent presentations of the CS following extinction reinstate heroin seeking behaviour. CONCLUSIONS: These experiments have established a method of measuring heroin-seeking behaviour in rats by adopting a second-order schedule of i.v. heroin self-administration. The results indicate a relatively weak impact of discrete, heroin-associated cues on heroin-seeking behaviour relative to cocaine seeking behaviour studied under similar conditions. PMID- 11255924 TI - On the relationship between the dopamine transporter and the reinforcing effects of local anesthetics in rhesus monkeys: practical and theoretical concerns. AB - RATIONALE: Drugs that are self-administered appear to vary in their potency and effectiveness as positive reinforcers. Understanding mechanisms that determine relative effectiveness of drugs as reinforcers will enhance our understanding of drug abuse. OBJECTIVES: The hypothesis of the present study was that differences among dopamine transporter (DAT) ligands in potency and effectiveness as a positive reinforcers were related to potency and effectiveness as DA uptake inhibitors. Accordingly, self-administration of a group of local anesthetics that are DAT ligands was compared to their effects as DA uptake blockers in vitro in brain tissue. METHODS: Rhesus monkeys were allowed to self-administer cocaine and other local anesthetics i.v. under a progressive-ratio schedule. The same compounds were compared in standard in vitro DA uptake assays using monkey caudate tissue. RESULTS: The rank order of both potency and effectiveness as reinforcers was cocaine > dimethocaine > procaine > chloroprocaine. Tetracaine did not maintain self-administration. For inhibiting DA uptake, the potency order was cocaine > dimethocaine > tetracaine > procaine > chloro-procaine. At maximum, these compounds were equally effective in blocking DA uptake. Lidocaine did not inhibit DA uptake. CONCLUSIONS: The potency of local anesthetics as positive reinforcers is likely related to their potency as DA uptake inhibitors. Variation in their effectiveness as positive reinforcers was not a function of differences in effectiveness as DA uptake blockers, but may be related to relative potency over the concentrations that are achieved in vivo. Effects at sodium channels may limit the reinforcing effects of local anesthetics. PMID- 11255925 TI - Abuse liability of the anesthetic propofol: self-administration of propofol in rats under fixed-ratio schedules of drug delivery. AB - RATIONALE: Previous reports suggest that propofol (PPF) may have abuse potential in humans. Hence, we hypothesized that PPF could reinforce self-administration behavior in other species. Positive reinforcing effects of PPF could be interpreted as an index of abuse liability. OBJECTIVE: Acquisition and maintenance of i.v. PPF self-administration were examined in 12 rats. METHODS: Six rats were initially given access to methohexital (MHX, 2.0 mg/kg per infusion) under a fixed ratio (FR) 1 schedule, while the other six were initially given access to PPF (1.7 mg/kg per infusion). Once stable responding was established, various doses of PPF (0.56, 1.0, and 1.7 mg/kg per infusion) and vehicle (Intralipid 20%) were made available. RESULTS: The number of PPF infusions per session was an inverse function of dose, with 0.56 mg/kg and 1.0 mg/kg per infusion maintaining significantly more infusions per session than vehicle in most rats under the FR 1 schedule. For some rats, the number of vehicle infusions per session was equal to or greater than the number of PPF infusions. Increasing the response requirement to FR 5 decreased the number of vehicle infusions per session in these rats, while PPF maintained a higher number of infusions than vehicle under this FR value in six of seven rats. CONCLUSION: PPF served as a reinforcer in rats under FR schedules of i.v. drug delivery, adding to the extant evidence that it has abuse potential. PMID- 11255926 TI - Second-order schedules of drug reinforcement in rats and monkeys: measurement of reinforcing efficacy and drug-seeking behaviour. AB - RATIONALE AND OBJECTIVES: To review the literature on the use of second-order schedules of drug reinforcement in the context of experimental investigations of the neural and pharmacological mechanisms underlying addictive behaviour in general and drug-seeking behaviour in particular. METHODS: Second-order schedules of drug reinforcement are described in which responding is maintained not only by the self-administered drug, but also by contingent presentation of drug-paired stimuli that serve as conditioned reinforcers of instrumental behaviour. RESULTS: The behaviour of rats and monkeys responding under second-order schedules is discussed in relation to self-administered drug dose and the importance of drug associated cues in maintaining responding for cocaine, morphine or heroin. Drug seeking behaviour during the period before drug is self-administered is described and compared with drug-seeking behaviour derived from other procedures. In addition, results are summarised that demonstrate the differential involvement of the amygdala and prefrontal cortex in the acquisition of cue-controlled cocaine- and heroin-seeking behaviour, as well as the effects of drugs interacting with D3 dopamine, NMDA and AMPA receptors on drug-seeking behaviour and dopaminergic correlates of drug-paired stimuli presented non-contingently and during responding for cocaine under a second-order schedule. CONCLUSIONS: We argue that the first, drug-free interval (or other period) of responding under a second order schedule of reinforcement has particular utility in that it provides a measure of drug-seeking behaviour and reinforcing efficacy that are not affected by the pharmacological effects of recently administered drug. It also provides a means of investigating the role of drug-paired stimuli in drug-seeking behaviour, including its behavioural, neural and neurochemical basis. PMID- 11255927 TI - Conditioned place preference: what does it add to our preclinical understanding of drug reward? AB - RATIONALE: Among the various experimental protocols that have been used to measure drug reward in laboratory animals, conditioned place preference (CPP) has been one of the most popular. However, a number of controversial issues have surrounded the use of this experimental protocol. OBJECTIVE: The present review provides a theoretical overview of some critical issues relevant to CPP. The advantages and limitations of CPP are also covered. RESULTS: Based on modern and traditional theoretical formulations of Pavlovian conditioning, CPP appears to reflect a preference for a context due to the contiguous association between the context and a drug stimulus. Within this theoretical framework, it seems clear that CPP measures a learning process that is fundamentally distinct from drug self-administration. The main advantages of CPP are that it: (1) tests animals in a drug-free state; (2) is sensitive to both reward and aversion; (3) allows for simultaneous determination of CPP and locomotor activity; (4) is adaptable to a variety of species; (5) typically yields dose-effect curves that are monophasic rather than biphasic; and (6) has utility in probing the neural circuits involved in drug reward. The main limitations of CPP are that it: (1) is subject to interpretation based on the notion of novelty seeking; (2) is cumbersome for providing the graded dose-effect curves needed for answering some pharmacological questions; (3) is difficult to interpret when animals prefer one context prior to drug conditioning; and (4) lacks face validity as an experimental protocol of drug reward in humans. CONCLUSION: Despite some limitations, CPP provides unique information about the rewarding effect of contextual cues associated with a drug stimulus. PMID- 11255928 TI - Deconstructing relative reinforcing efficacy and situating the measures of pharmacological reinforcement with behavioral economics: a theoretical proposal. AB - BACKGROUND: Relative reinforcing efficacy has been assumed to be a homogeneous phenomenon referring to the behavior-strengthening or behavior-maintaining effects of a drug reinforcer. However, a variety of studies suggest that relative reinforcing efficacy may be heterogeneous. OBJECTIVES: The purpose of this theoretical proposal is to examine the difficulties associated with this conception of reinforcing efficacy and to explore whether relative reinforcing efficacy is a homogenous concept or whether it is composed of several functionally related heterogeneous phenomena. In examining this issue, we explore whether behavioral economic theory may address some of the challenges to the current conception of relative reinforcing efficacy and use this theory to suggest how the differing measures of reinforcing efficacy may relate to one another. RESULTS: Results indicate that peak-response rate and breakpoint are related to the economic measure of maximal output and elasticity of demand, respectively. Preference is related to and predicted by the relative location of the demand curves obtained under single schedule conditions. This behavioral economic analysis may provide a theoretical understanding of reinforcement that can reconcile results of studies that both support and fail to support the notion of reinforcing efficacy as a homogenous phenomenon. CONCLUSIONS: If this theoretical proposal is validated by additional studies, then like other natural phenomena found to be heterogeneous, the study of drug reinforcers may require the adoption of several new scientific terms, such as those used in behavioral economics, each of which has analytical precision and refers to homogeneous phenomena. PMID- 11255929 TI - Toward a mathematical description of dose-effect functions for self-administered drugs in laboratory animal models. AB - RATIONALE: The interpretation of dose-effect functions for self-administered drugs remains elusive. Since, for self-administered drugs, the amount of drug in an animal depends on its behavior, a mathematical theory of drug self administration must include terms relevant to receptor theory, as well as a description of how an organism's behavior affects the amount of drug in the animal over time. OBJECTIVE: A theory was constructed in which the ability of a dose to maintain responding was described in terms of receptor theory and the function relating rate of responding to amount of drug self-administered. The main predictions of the theory were that: 1) there should be no ascending limb for drugs self-administered under ratio schedules, 2) running rate of response should not change as a function of dose and, 3) pause duration should be an exponential function of dose. RESULTS: Low doses of cocaine were either self administered at high rates, or not at all. Run rates, though somewhat variable, did not change as an orderly function of dose. Pause duration could be well described by an exponential function. CONCLUSIONS: The theory provides an acceptable, though no doubt preliminary, description of drug self-administration. PMID- 11255930 TI - Agonist efficacy, drug dependence, and medications development: preclinical evaluation of opioid, dopaminergic, and GABAA-ergic ligands. AB - BACKGROUND: The general premise that receptor theory provides a useful framework for understanding the behavioral effects of psychoactive drugs has been a central tenet of behavioral pharmacology. OBJECTIVES: The purpose of this review is to reiterate this basic theme and, in particular, the proposition that current concepts of pharmacological efficacy can be effectively used to examine behavioral effects of drugs with abuse or dependence potential in a way that contributes to the discovery of drugs to treat drug dependence. EXPERIMENTAL DATA: The review begins by briefly introducing the concept of efficacy and follows with several illustrations of how our current understanding of efficacy can be used to address important research questions in drug discovery. In the first, the likelihood of developing novel opioid analgesics with reduced abuse potential is addressed by considering the different efficacy requirements for the discriminative-stimulus and antinociceptive effects of mu-opioids. From a pharmacologically different perspective within drug abuse research, the review continues with an exposition of efficacy-related differences in the behavioral effects of dopamine D1 agonists and how such differences might be exploited in different medications strategies for treating cocaine dependence. The principles of pharmacological efficacy also have come to guide the development of novel GABAA-related antianxiety medications, and this is illustrated in a discussion of the utility of low-efficacy agonists in the treatment of benzodiazepine dependence. The second half of the paper provides counterpoint to the several examples of how principles of efficacy can be applied in drug discovery. The counterpoint includes, first, a critical evaluation of how the concept of efficacy has been applied in the development of monoamine transport inhibitors as anti-cocaine medications and, in particular, the difficulties this may pose for data analysis. The review ends with a discussion of efficacy-based analysis in drug discrimination research and illustrates some of the obstacles that may be encountered in pharmacologically classifying drugs on this basis. CONCLUSIONS: Ample evidence indicates that many receptor systems can be activated in a graded manner and that principles of efficacy can be judiciously applied to understand and exploit the behavioral effects of drugs that result from such graded activation. However, as cautioned in the last sections, the misapplication of pharmacological concepts in behavioral studies of drugs may obscure their behavioral pharmacology and potentially confound drug discovery. PMID- 11255931 TI - d-amphetamine increases choice of cigarette smoking over monetary reinforcement. AB - RATIONALE: Psychomotor stimulants previously have been found to increase the frequency of cigarette smoking, but it is unclear whether this is due to a non specific increase in general activity or a specific increase in the reinforcing effects of smoking. OBJECTIVES: To investigate whether d-amphetamine increases the relative reinforcing effects of cigarette smoking. METHODS: Ninety minutes after d-amphetamine (7.5, 15 mg/70 kg) or placebo administration, 13 male and female subjects participated in 3-h sessions during which they could make a maximum of 20 choices between cigarette smoking (two puffs per choice), earning money ($0.25 per choice), or neither. In separate sessions, using the same subjects, the effects of d-amphetamine on the frequency of ad libitum smoking was assessed. RESULTS: During choice sessions, d-amphetamine dose-dependently increased smoking choices from 4.2 +/- 0.6 to 5.7 +/- 0.6. During sessions in which subjects smoked ad libitum, d-amphetamine increased number of cigarettes smoked from 2.8 +/- 0.4 to 3.8 +/- 0.6. Breath carbon monoxide (CO) levels, a measure of smoke exposure, showed corresponding dose-related increases. CONCLUSIONS: These results are consistent with previous findings that d amphetamine increases smoking and provide evidence that this effect is due to a drug-produced increase in the relative reinforcing effects of cigarette smoking. PMID- 11255932 TI - Kappa-opioid receptors and relapse-like drinking in long-term ethanol-experienced rats. AB - RATIONALE: The role of the dynorphin/kappa-opioid receptor system in ethanol reinforcement is unclear. OBJECTIVE: Examination of the effects of the highly selective kappa-opioid receptor agonist CI-977 (enadoline) and of the long-acting selective kappa-opioid receptor antagonist nor-binaltorphimine (nor-BNI) on relapse-like drinking measured by the alcohol deprivation effect (ADE) in long term ethanol-experienced rats. METHODS: Rats were either implanted with mini osmotic pumps delivering 0 or 0.01 mg/kg per h CI-977 or received two injections (12 h apart) of nor-BNI (0 or 5 mg/kg i.p.) before representation of alcohol after 2 weeks of alcohol deprivation in a four-bottle home cage drinking paradigm. In a second experiment, long-term ethanol-experienced rats trained in an operant ethanol self-administration paradigm received either acute CI-977 treatment (0, 0.003-0.1 mg/kg i.p.) or two injections (12 h apart) of nor-BNI (0 or 5 mg/kg i.p.) before a 23-h session. RESULTS: Chronic CI-977 potentiated ethanol intake and preference during the ADE. Acute CI-977 dose-dependently reduced total lever pressing activity demonstrating an unspecific sedative effect, except for the lowest dose (0.003 mg/kg), which selectively increased lever pressing for ethanol during basal drinking. Nor-BNI did not affect relapse like drinking at all. CONCLUSIONS: Stimulation of kappa-opioid receptors can increase ethanol intake, at least in long-term ethanol-experienced rats. Since kappa-opioid receptor agonists have aversive motivational consequences, increased ethanol drinking might be an attempt to counteract the aversive effects of this treatment. On the other hand, the nor-BNI experiments indicate that endogenous kappa-opioid receptor stimulation does not seem to be involved in relapse-like drinking after protracted abstinence. PMID- 11255933 TI - Symptoms of depression and survival experience among three samples of smokers trying to quit. AB - Symptoms of depression have been associated with increased smoking prevalence and failure to quit smoking in several cross-sectional and population-based studies. Few studies, however, have prospectively examined the ability of current symptoms of depression to predict failure to quit smoking in treatment-motivated smokers. Pretreatment depressed mood was assessed by 3 different methods in 3 separate samples, 2 of which comprised smokers receiving combined pharmacological and behavioral treatments and a 3rd in which smokers received self-help materials only. In all studies, time in days from quit day until the 1st cigarette was ascertained to document survival. Survival analyses showed that in all 3 studies survival time was significantly and negatively related to measures of even very low levels of pretreatment depressed mood. Results were replicated across 3 independent samples and were robust and uniformly clear, indicating that low levels of depressive symptoms assessed at baseline predict time to 1st cigarette smoked after attempted quitting. PMID- 11255934 TI - Activation of alcohol expectancies in memory in relation to limb of the blood alcohol curve. AB - Attempts to understand the mechanism by which alcohol expectancies might influence drinking have related activation of expectancies in memory to alcohol use. Limb of the blood alcohol curve, however, has not been considered. In the present study, 527 undergraduates completed the Anticipated Biphasic Alcohol Effects Scale and a drinking measure. Multidimensional scaling was used to map expectancies into memory network format, and likely activation of expectancies was empirically modeled. Heavier drinkers were most likely to activate positive and arousing expectancies associated with the ascending limb, whereas lighter drinkers were most likely to activate negative and sedating expectancies associated with the descending limb. These findings add to the literature suggesting that activation of expectancies in memory may be an important determinant of drinking behavior and a promising target for intervention strategies. PMID- 11255936 TI - A risk factor index predicting adolescent cigarette smoking: a 7-year longitudinal study. AB - The authors used longitudinal data to develop a risk factor index (RFI) for the prediction of smoking behavior in youth. Students were followed yearly from 6th through 12th grades in a prospective longitudinal study. Ten risk factors were identified and combined into an RFI. Data were analyzed with a generalized estimating equations approach. The RFI predicted both concurrent smoking and use of cigarettes in the succeeding year. It further predicted whether a youth would smoke at any point during his or her school career. Prediction was better for boys than for girls. Furthermore, the RFI better predicted heavy smoking than any use of cigarettes. The RFI could be useful in selecting youth for intensive prevention and early intervention efforts. Results also suggest the importance of further examination of gender differences in smoking behavior. PMID- 11255935 TI - Perceived control in adolescent substance use: concurrent and longitudinal analyses. AB - A sample of 9th-grade students (1,293 individuals, 51% girls) attending compulsory schools in Reykjavik, Iceland, was surveyed and followed up 3 years later. The relationship between perceived control and substance use is examined concurrently at age 14 for experimentation with tobacco and alcohol and longitudinally (14-17 years of age) for daily smoking, heavy drinking, and illicit drug use. Taking into account sociodemographic characteristics (family structure, socioeconomic status, and gender) and parental and peer use, the results of concurrent analyses indicate that adolescents who expressed more personal control were less likely to have smoked and to have had a drink at age 14. Longitudinal analyses showed that perceived control at age 14 predicted both daily smoking and illicit drug use among girls at age 17 but not among boys. Conversely, perceived control did not predict heavy drinking among adolescents. PMID- 11255937 TI - Initial validation of a computer-administered Addiction Severity Index: the ASI MV. AB - The Addiction Severity Index--Multimedia Version (ASI-MV) is a CD-ROM-based simulation of the interviewer-administered Addiction Severity Index (ASI). Clients in treatment (N = 202) self-administered the ASI-MV to examine the test retest reliability, criterion validity, and convergent-discriminant validity of the ASI-MV. Excellent test-retest reliability was observed for composite scores and severity ratings. Criterion validity, tested against the interviewer administered ASI, was good for the composite scores. For severity ratings, variable agreement was observed between the ASI-MV and each interviewer, suggesting poor interrater reliability among interviewers. This conclusion was bolstered by a finding of superior convergent-discriminant validity for both composite scores and severity ratings compared to the standard ASI. The ASI-MV is a viable alternative to the expensive and potentially unreliable interviewer administered version. PMID- 11255938 TI - Short- and long-term effects of fraternity and sorority membership on heavy drinking: a social norms perspective. AB - This study sought to determine whether the well-established relation between fraternity/sorority (Greek) membership and heavy alcohol use persists beyond the college years and whether some common third variables might account for the relation between Greek status and heavy drinking. During each of 4 years of college and 1 additional year, young adults (N = 319) completed measures of alcohol use, personality, alcohol expectancies, and environmental influences on drinking. Throughout the college years, Greeks consistently drank more heavily than non-Greeks. Statistically controlling for previous alcohol use did not eliminate this effect. However, Greek status did not predict postcollege heavy drinking levels. Also, perceived peer norms for heavy drinking mediated the relation between Greek affiliation and heavy alcohol use. Results are discussed in terms of situational determinants of heavy alcohol involvement in young adults. PMID- 11255939 TI - Cocaine withdrawal symptoms and initial urine toxicology results predict treatment attrition in outpatient cocaine dependence treatment. AB - This study evaluated the ability of cocaine withdrawal symptoms, measured by the Cocaine Selective Severity Assessment (CSSA) and initial urine toxicology results, to predict treatment attrition among 128 cocaine dependent veterans participating in a 4-week day hospital treatment program. The CSSA was administered and a urine toxicology screen was obtained at intake and at the start of the day hospital (about 1 week later). The combination of a positive urine toxicology screen and a high CSSA score at intake predicted failure to complete treatment. Urine toxicology results at the start of the day hospital, but not at intake, predicted failure to complete treatment. Among participants without other psychiatric illness, high CSSA scores at intake predicted failure to complete treatment. The presence of cocaine withdrawal symptoms and a positive urine toxicology screen are clinically useful predictors of treatment attrition. PMID- 11255940 TI - Positive alcohol expectancies and drinking behavior: the influence of expectancy strength and memory accessibility. AB - College student drinkers (N = 314) participated in a health survey in which they (a) completed an alcohol-related memory association task (expectancy accessibility measure), (b) rated their positive expectancies about alcohol use (expectancy strength measure), and (c) reported their level of alcohol involvement. Hierarchical regression analyses showed that both expectancy accessibility and expectancy strength predicted frequency of alcohol use and alcohol-related problems. Moreover, moderational analyses showed that the association between expectancy strength and frequency of alcohol use was greater for those who generated more alcohol responses on the expectancy association task. These findings suggest that the outcome association measure and Likert scale ratings of expectancies may assess distinct properties of expectancy representations, which may have independent and interactive effects on different aspects of drinking behavior. PMID- 11255941 TI - Course of functioning in adolescents 1 year after alcohol and other drug treatment. AB - Clinical course was studied in 131 male and female adolescents with current alcohol use disorder (AUD) at baseline (BL). Participants were classified into 4 groups according to their diagnosis and drinking pattern 1 year later. The 4 groups were compared with each other and with 37 community control participants. Results showed that over half of the clinical sample no longer had a current AUD at 1 year; about 64% were and 36% were not still drinking. BL discriminators of 1 year status were alcohol dependence, other drug use, and coping. All of the clinical groups tended to show improvement at 1 year in the main dependent variables, and the abstainers' level of drug use and coping were comparable with that of the community participants. These findings suggest that many adolescents improve in functioning during the 1 year after alcohol and drug treatment and that a stress and coping model is useful for studying clinical course of AUDs in adolescents. PMID- 11255942 TI - Clinical features of pathological gambling in an addictions treatment cohort. AB - This study examined the prevalence and descriptive psychopathology of pathological gambling in a heterogeneous treatment sample of 372 substance users. About 14% of male participants and 10% of female participants were identified as presumptive pathological gamblers (PGs) on the South Oaks Gambling Screen (SOGS). The authors contrasted 49 PGs with 323 participants who were not pathological gamblers (NPGs) on a host of variables measuring premorbid risk, pathological patterns of substance use, consequences of use, and psychiatric comorbidity. PGs showed more disturbance than NPGs on some measures of premorbid risk, pathological substance use, social consequences of use, and psychiatric comorbidity. Gambling status may be an important comorbid condition in addictions treatment settings and a significant covariate in research. PMID- 11255943 TI - [Contribution of Communal Hygiene Department to creation and development of norm setting basis of water sanitary legislation]. AB - Legal regulation of problems related to drinking water supply to the population is a pressing social problem, particularly during the transitional period of Russian economic development. Communal Hygiene Department participated in the formation and development of norm-setting base of drinking water supply to the population since the thirties. During recent 5 years the Department contributed to development of the "Law on Drinking Water and Drinking Water Supply" (the first in Russia), basic normative legislative acts regulating hygienic requirements to drinking water quality, choice and exploitation of sources of drinking water supply, zones of their sanitary protection, and many methodological documents. The paper sums up the principal basis of new generation documents of sanitary legislation concerning water. PMID- 11255944 TI - [Validation, development and biomedical evaluation of new food products for common and special use]. AB - Low level of provision of various population groups with animal proteins, poor choice of foodstuffs for common and dietetic use, inefficiency of meat and milk processing, loss of foodstuff quality as a result of industrial production determine the range of priority problems to be solved by specialists in nutrition hygiene. One of the main and most rapid approaches to solution of these problems is more rational utilization of the foodstuff potential created in the country for nutrition, but the chemical, sensory, and organoleptic properties of the manufactured foodstuffs do not always ensure their high biological value. Department of Hygiene of Nutrition, I. M. Setchenov Moscow Medical Academy, for many years has been engaged in development of biomedical rationale for effective use of raw material and processing protocols for preparation of common and dietetic food for various population groups. Special attention was paid to the effects of food-stuffs prepared by new technologies and recipes on metabolism. The most important results are summed up in this review. PMID- 11255945 TI - [Children's health in zone of radioactive pollution caused by the Chernobyl accident]. AB - The Chernobyl accident led to contamination of vast territories of Russia by man made radionuclides. One of such regions is the Tula region. For realization of programs aimed at alleviation of the accident consequences, medical and sanitary measures were taken, including many-year monitoring of radiation pollution in the region and health status of various population groups, primarily children. Complex studies were carried out, making use of radiation, sociohygienic, epidemiological, psychophysiological, anthropometric, clinical laboratory, cytogenetic, immunological, and other methods. Dynamic examinations of children of different age demonstrated that radioactive precipitations at a density of 5 15 Ci/m2 did not directly affect children's health; however essential deviations in the health status, such as changes in morbidity, physical development, and incidence of immunodeficiencies have been revealed. The authors consider that realization of health protection programs should concern primarily children. PMID- 11255946 TI - [Development of Public Health Department with course of economy]. AB - Transition of Russia to marketing economy necessitated reformation of public health system, which, in turn, necessitated economic education of medical staff. I. M. Setchenov Moscow Medical Academy accumulated sufficient experience in the solution of this problem many-year research validated the need in economic education of medical staff under conditions of public health reforms. This paper sums up the results of organization and experimental studies of introduction of public health economy in the curriculum. PMID- 11255947 TI - [Human microecology studies at Department of Microbiology with Virology and Immunology, I.M. Sechenov Moscow Medical Academy]. AB - Studies of human microflora in health and disease and during exposure to professional and ecological factors is a traditional problem solved for many years by staff members of Department of Microbiology with Virology and Immunology, I. M. Setchenov Moscow Medical Academy. The purpose of research is to develop methods and means for diagnosis and prevention of human microbiocenosis disorders. Fundamental and applied research in cooperation with prophylactic and clinical institutions and departments yielded data contributing to solution of many pressing problems in prevention and diagnosis of infectious diseases. PMID- 11255948 TI - [Research priorities at medical prophylaxis faculty]. AB - Medical Prophylaxis Faculty of I. M. Setchenov Moscow Medical Academy, which is now 70 years old, has made a significant contribution to development of sanitary epidemiological service in the country and training of specialists and research and training staff. The main forms of activities are training, research, and improvement of training and methodological work on the basis of research results, as well as upbringing of students. Despite the difficulties arising in the course of science and economy reforms, the departments of the Faculty continue fundamental and applied research. This paper sums up the activities of Medical Prophylaxis Faculty and outlines the main trends of research carried out at its departments. PMID- 11255949 TI - [Systemic vascular purpura: clinical etiological variants]. AB - Systemic vascular purpura (SVP) represents angiites, heterogeneous by etiology and pathogenesis. Their pathogenetic classification does not fully reflect the clinical picture, course, and prognosis. The authors suggest a clinical etiological approach to differential diagnosis of SVP. With this aim in view they examined 82 patients with SVP hospitalized at Clinic of Therapy and Occupational Diseases of I. M. Setchenov Moscow Medical Academy in 1993-1996. Etiologically associated differences in clinical manifestations of SVP were detected and clinical etiological variants of disease were distinguished. Endotoxemia was found to be often associated with purpura of different etiology, which can be regarded as a stage in the pathogenesis. The data allow creation of a clinical etiological classification of SVP and development of new therapeutic approaches. PMID- 11255951 TI - [Prevention of breast cancer]. AB - Analysis of 60 Russian and foreign publications demonstrated that the incidence of breast cancer and socio-biological characteristics of this disease necessitate improvement of methods for its prevention and treatment. Factors significantly increasing the probability of breast cancer are analyzed. Risk groups in need of thorough prophylactic examinations and treatment can be formed on the basis of these data. Some dyshormonal hyperplasias are considered as precancer. Active prophylactic measures, including surgery, are recommended for patients with such forms of mastopathy. There is a group of patients with 100% risk of breast cancer. Cancer can be prevented in them by prophylactic interventions, from resection of the mammary gland to subcutaneous mastectomy with plasty. Such strategy will have a positive impact on the incidence of breast cancer and the results of its treatment. PMID- 11255950 TI - [Thrombophilia and fetal loss syndrome]. AB - Pathological states associated with congenital and acquired hemostasis defects remain a pressing problem of practical obstetrics. Thrombophilia is responsible for 30-50% of habitual miscarriages, severe gestosis, detachment of normally attached placenta, preterm delivery, etc. Therefore, today correct therapeutic strategy determines the prevention of reproductive loss caused by thrombophilic state. A complex of studies aimed at drug prevention of miscarriages has been performed at Department of Obstetrics and Gynecology, Medical Prophylaxis Faculty of I. M. Setchenov Moscow Medical Academy. Seventy-two pregnant women without extragenital diseases with verified congenital or acquired hemophilia and fetal loss syndrome were observed. Various therapeutic protocols with low-molecular weight heparin were used. Positive clinical results were attained: no relapses of thrombosis and fetal loss occurred in the majority of cases, though the incidence of preterm labor and placental insufficiency remained high. PMID- 11255952 TI - [Relationship between changes in demographic structure and incidence of anthroponotic infections]. AB - Recent demographic shifts in Russia tell on the incidence of infectious diseases. With the aim of prediction, the authors analyzed the relationship between demographic changes in the country and the mean many-year level of anthroponotic infections with strained immunity in Moscow. Mathematical simulation methods were used. The data coincided with decreases in the incidence of scarlet fever and viral hepatitis A, observed in the nineties. On the other hand, the incidence of infectious diseases can return to the level of the beginning of the nineties irrespective of the demographic status in Russia. PMID- 11255953 TI - [Prospects of using hard dispersions in development of dosage forms for therapeutic and prophylactic use]. AB - Thermoanalytical, chromatographic, and microscopic methods of analysis were used for identification of hard dispersions (HD) and their differences from physical mixtures (PM) by benzene PEG-4000 model systems. A complex of physiocochemical methods showed that benzonal and PEG-4000 are not thermally destroyed during simultaneous melting at 140 degrees C. Differences in thermoanalytical characteristics of PM and HD, expressed in suppression of phase transition temperatures, changes in the type of melting peaks and heats, and in the absence of drug melting peak and heats in HD vs. PM confirm the formation of new physiochemical systems differing from PM. The resultant quantitative relationships between temperature depression and melting heats for HD and PM of different composition correlate with chromatographic findings. PMID- 11255954 TI - [Stages and results of Epidemiology Department activities]. AB - The paper sums up the activities of Department of Epidemiology of Medical Prophylaxis Faculty, I. M. Setchenov Moscow Medical Academy, in the 70 years of its existence. The Department plays the leading role in training specialists for sanitary and epidemiological service of the country. The main research, training, and methodological works in this speciality were prepared by the Department staff members, who prepared curricula for pre- and postgraduate training. For many years the department has been the methodological center in epidemiology training. Trends of research at the Department are concentrated on the pressing problems of general and special epidemiology. Due to activities of the department staff, epidemiological aspects have been studied and fundamentals of prevention and liquidation of many infectious diseases in the country have been formulated: enteric and streptococcal infections, typhus, tularemia, diphtheria, etc. Problems of epidemiological geography, including nosogeography, territorial and republican epidemiology have been investigated. Functioning of parasitic systems and mechanisms of epidemic process in some infections have been characterized with due regard for modern theoretical, methodological, and organization base of epidemiology. The data were used for developing new basis and organization forms of epidemiological surveillance of infectious diseases. PMID- 11255955 TI - [Possible difficulties in molecular-genetic expertise in cases with insufficiently high individual significance of the results (as exemplified by a complex case with disputable maternity)]. AB - Molecular genetic technologies used in forensic medical expert evaluations help quantitatively evaluate the significance of coincidence or non-coincidence of signs in personality identification and in expert evaluation of kinship identification (disputable paternity or maternity). The level of validity of evidences, which can be considered necessary and sufficient, is the principal problem in such cases. Analyzing a complex case with disputable maternity, the authors discuss problems illustrating the necessity of attaining a high level of validity of the results for drawing a justified expert conclusion. Only high validity standard can rule out errors in interpretation of the results, otherwise the significance of the detected complex of signs can be insufficient for an unambiguous solution of an expert task. PMID- 11255956 TI - [Immunoglobulins, binding opioid peptides, biogenic amines, and opiates, in substance abusers]. AB - Immunoglobulins binding morphine, biogenic amines, and an opioid peptide dermorphine were measured by solid-phase enzyme immunoassay in patients abusing narcotics. The patients' ages varied from 20 to 40, with the duration of narcotic use 1-17 years. Narcotic dependence was found to involve increased production of immunoglobulins binding opioid and monoamine neuromediators. Enzyme immunoassay of antibodies to opiates detects latent forms of narcomania in cases when the narcotic is not present in the body. These results serve the basis for creating a new method for analysis of narcotic abuse and evaluation of the duration of their use, which can be used in practical medicine and in forensic medical expert evaluations. PMID- 11255957 TI - [Detection of 1,4-benzodiazepine derivatives in liver tissues by immunochemical methods]. AB - The possibility of using immunochemical methods in general forensic chemical analysis of cadaveric material for narcotics is demonstrated. Conditions of isolation of 1,4-benzodiasepines from tissues and analysis of the resultant tissue extracts by polarization fluoroimmunoassay (PFIA) and solid-phase enzyme immunoassay are described. Time course of concentrations in tissue extracts stored for a long time has been studied by PFIA. Characteristics of Benzodiazepines Serum kits (Abbott) for PFIA of liver extracts are determined. PMID- 11255958 TI - [Features of extracting phenol and 4-methylphenol from aqueous solutions]. AB - Phenol and 4-methylphenol were extracted from aqueous solutions by 5 organic solvents. The degree of extraction depended on such factors as type of extracting agent, pH of aqueous phase, and water saturation of extracting agent. The necessary number of extractions for obtaining the preset volumes of studied compounds is calculated. PMID- 11255959 TI - [Detection of chlorinated carbohydrates in putrefactive biological material]. AB - Carbochlorohydrates were separated from putrefaction products of organic substances and volatile solvents by gas chromatographic column packed with CaA 5 A. A 100 x 0.4 cm column, used for forensic chemical analysis of fresh and putrefactive cadaveric material, proved to be highly effective. The data were confirmed by results of chemical analysis of distillates from the viscera. PMID- 11255960 TI - [Morphometric approaches to diagnosing the time of death]. AB - A new method has been used to evaluate the time of death. The relationship between the degree of autolysis of cadaveric viscera and time of death has been studied by histomorphometrical methods. The study was carried out on histological preparations of the lungs, heart, liver, spleen, and kidneys from 20 corpses of humans dead from craniocerebral injuries. Based on the results, tables were created for evaluating the time of death even at remote periods after death, when the results of other methods are unreliable. PMID- 11255961 TI - [Epidemiologic analysis of ephedrone substance abuse in the Primorye territory]. AB - Biochemical effects of ephedrone and their clinical manifestations are described. Statistical data on the prevalence of ephedrone narcomania in the Primorye territory are compared with the prevalence of use of psychostimulating narcotics in other regions (analysis of the files of thanatological department of Bureau of Forensic Medical Expert Evaluations in Moscow). Morphological changes in the viscera are described detail. The authors come to a conclusion on high incidence and multifactorial toxic effects of ephedrone and multilevel somatic involvement in ephedrone narcomania. PMID- 11255962 TI - [The possibilities of committing voluntary actions with a foreign body in the cranial cavity]. PMID- 11255963 TI - [Features of the corpse and place of occurrence in sexual suicide]. PMID- 11255964 TI - [Pathomorphological study in craniocerebral injury]. AB - Time course of the posttraumatic process in craniocerebral injuries is analyzed. Pathomorphological picture of changes in soft tissues of the head, meninx, and brain matter are described. Several periods are distinguished. Factors affecting the course of the posttraumatic process in the above-mentioned structures are discussed. PMID- 11255965 TI - [Forensic medical aspects of injuries inflicted with self-defense capsaicin aerosols]. AB - Toxicological and forensic medical characteristics of capsaicinoids are presented. Capsaicinoids represent a mixture of three basic compounds with similar structure: capsaicine, dihydrocapsaicine, and nordihydrocapsaicine, all of which are the primary components of red pepper extract (Oleoresin Capsicum). The advantages of this group of irritants in comparison with synthetic irritants are discussed. Probable routes of administration of the extract with regard to its hazard for the organism are considered. Results of experimental studies on nonbiological phantoms and volunteers are presented. PMID- 11255966 TI - [Staging for locoregional extension of rectal adenocarcinoma]. AB - The treatment of rectal carcinoma is mainly determined by its local extension. Preoperative staging of rectal carcinoma was assessed by different methods: digital rectal examination, transrectal ultrasound, computed tomography, and magnetic resonance imaging. Digital rectal examination had a diagnostic accuracy between 68 and 83 per cent. The accuracy of transrectal ultrasound was between 67 and 93 per cent for tumor staging and between 62 and 88 per cent for lymph node staging. The accuracy of computed tomography was between 33 and 77 per cent for tumor staging and between 22 and 73 per cent for lymph node staging. The overall accuracy of magnetic resonance imaging with body coil was between 59 and 95 per cent, and between 39 and 95 per cent for lymph node staging. Use of an endorectal coil allows a slightly more consistent degree of accuracy, with tumor staging accuracy between 66 and 91 per cent, and lymph node staging accuracy between 72 and 79 percent. Preoperative radiation therapy makes transrectal ultrasound and computed tomography less effective as staging techniques. PMID- 11255967 TI - [Direct and reservoir colonic-anal anastomoses. Short and long term results]. AB - STUDY AIM: This retrospective study was designed to assess the operative, oncologic and functional results of total proctectomy with coloanal anastomosis (CAA). PATIENTS AND METHOD: Between 1990 and 1994, 81 patients (44 males/37 females: mean age: 59 years) were operated for a cancer (n = 67) or a benign lesion (n = 14) of the rectum. Sixty-four patients had a straight CAA and 17 patients had a colonic J-pouch. RESULTS: There was no operative mortality. Two patients were reoperated for colonic necrosis and underwent abdominoperineal resection. An anastomotic leak was observed in 11 patients and its severity was decreased by a diverting stoma. An anastomotic stricture was observed in 10 patients. Of the 67 patients with cancer, 19 (28%) developed metastases and 11 (16%) developed local recurrence. The 5-year survival rate was 69%. Twelve months after the operation, continence was similar with the two types of CAA, but the mean daily stool frequency was lower in patients with a reservoir. With a long follow-up (mean = 9 years), functional results were good with regard to continence and stool frequency, almost similar with the two types of CAA; functional disorders (noctumal stools, fragmentation, urgency) were reported by 25 to 40% of patients. CONCLUSION: Total proctectomy with coloanal anastomosis yields good oncologic results. With regard to functional results, the superiority of the colonic J-pouch, which is observed in the first postoperative year, was lost beyond this period; long-term results are good for continence and stool frequency, but some disorders persist in a significant proportion of patients. PMID- 11255968 TI - [Hepatic artery aneurysms]. AB - AIM OF THE STUDY: The aim of this retrospective study was to report a series of nine aneurysms of the hepatic arteries, including real aneurysms (n = 4), pseudoaneurysms (n = 3) and false aneurysms (n = 2) observed from 1987 to 1999. PATIENTS: There were 7 men and 2 women (mean age: 58 years). In 3 cases, the aneurysm was asymptomatic and detected by sonography; in 4 cases it was revealed by rupture with a severe hemorrhage and in 2 cases by cholestasis. The aneurysm was located on right (n = 3), proper and common (n = 3), proper (n = 2), and common (n = 1) hepatic arteries. The aneurysm was associated with hepatocellular carcinoma (n = 1), carcinoma of the head of the pancreas (n = 1) and liver metastases (n = 1). METHODS AND RESULTS: Eight patients were operated and one of them was operated three times. Hepatic arterial blood supply was restored in 6 patients with simple suture (n = 1), Goretex graft (n = 2), allograft (n = 2) and autologous vein (n = 1), with one failure which required liver retransplantation. Only one of the three attempts of embolization was successful. One patient with surgical contraindications died from hemobilia after embolization failure. During follow-up, there was one thrombosis of the common hepatic artery which had been excluded and two late deaths: one from rupture of a false aneurysm after bypass with an allograft and one by terminal progression of the cancer. The other 6 patients were alive at the time of this study. CONCLUSION: Clinical characteristics and therapeutic indications of hepatic arterial aneurysm are variable. Management is usually surgical, while embolization is reserved for special circumstances. Restoration of the hepatic arterial blood supply is necessary in aneurysms located on the proper hepatic artery. PMID- 11255970 TI - [Zenker's diverticulum: diverticulopexy versus diverticulectomy]. AB - STUDY AIM: The aim of this retrospective, nonrandomized study was to compare the results of diverticulectomy and diverticulopexy in the treatment of Zenker's diverticulum. Over the 10-year period between 1988 and 1998, surgery for Zenker's diverticulum was performed in 40 patients. PATIENTS AND METHOD: The study group consisted of 23 men and 17 women with a mean age of 72 years. Only 39 patients were evaluated. In 19 patients, treatment consisted of cricopharyngeal myotomy and diverticulum suspension; in the other 19 patients, treatment consisted of diverticulectomy in addition to myotomy. Only one patient had a diverticulectomy without myotomy. RESULTS: There was no mortality and the morbidity rate was low: one fistula, one pneumonia, three cases of transient dysphonia and one hematoma. The results were excellent in 36 patients, and good in 3 patients. CONCLUSION: Cricopharyngeal myotomy with diverticulopexy is particularly suitable for geriatric patients. Diverticulectomy is proposed in the case of a diverticulum larger than 6 cm and for young patients to prevent the risk of malignant transformation. PMID- 11255969 TI - [Evaluation of somatostatin or octreotide efficacy in the treatment of external pancreatic fistulas]. AB - AIM OF THE STUDY: To evaluate the prevalence of pancreatic pseudocyst after persistent fistula closure with somatostatin or octreotide. To compare the patient characteristics according to the subsequent presence or absence of pseudocyst. PATIENTS AND METHODS: This retrospective study from January 1994 to August 1999 included 15 patients with an external pancreatic fistula. Fistula closure was observed for all patients with somatostatin or octreotide. CT scan was performed 66 +/- 34 days after the end of this treatment. RESULTS: CT scan was normal in 9 patients (favorable group) and showed pancreatic pseudocyst (failure group) in 6 patients. Pancreatic fistula etiologies were different between the two groups. The 5 patients presenting pancreatic fistula after duodenopancreatectomy belonged to the favorable group. Six of the 10 patients presenting pancreatic fistula after pseudocyst drainage belonged to the failure group. There were no other differences between the two groups. CONCLUSION: Persistent pancreatic fistula can be cured with somatostatin or octreotide. However, fistulas occurring after duodenopancreatectomy are more easily cured with somatostatin or octreotide than fistulas occurring after external pseudocyst drainage. Somatostatin or octreotide cannot be considered to be an effective treatment for pancreatic fistula occurring after pseudocyst drainage, despite the fact that 40% of them were permanently cured. PMID- 11255971 TI - [Segmental resection of tumoral invasion of the inferior vena cava without reconstruction]. AB - STUDY AIM: Oncological complete surgery of retroperitoneal tumours may require segmental resection of part of the invaded inferior vena cava. The aim of this retrospective study was to assess whether reconstruction of the inferior vena cava is necessary and presents any advantage. PATIENTS AND METHODS: This study included four patients who underwent partial resection of the inferior vena cava invaded by a retroperitoneal tumour, without reconstruction. Tumours were one renal cancer, one malignant phaeochromocytoma, one malignant retroperitoneal histiofibroma and one undifferentiated retroperitoneal carcinoma. The resection was located at the level of the renal confluence, associated with right nephrectomy, in 3 patients, and above this confluence, at the level of the retrohepatic inferior vena cava in 1 patient. RESULTS: Only one case of transient acute renal failure was observed during the postoperative course. One patient developed right deep vein thrombosis after three months and another one after 30 months. One patient died from cancer recurrence after 19 months. The other 3 patients were alive with anticoagulant therapy and without sequelae after 3, 6 and 15 years. PMID- 11255972 TI - [Laparoscopic gastroplasty for morbid obesity: prospective study of 300 cases]. AB - STUDY AIM: Laparoscopic gastric banding for morbid obesity is noninvasive and reversible. The aim of this prospective study was to report the preliminary results of this procedure in the first 300 patients. PATIENTS AND METHODS: From April 1997 to January 2000, 300 patients were laparoscopically operated for severe obesity: 266 women, 34 men, with a mean age of 40.1 years (range: 16-66). The mean preoperative weight was 118 kg (range: 85-195) and the mean body mass index (BMI) was 43.6 kg/m2 (range: 35.1-65.8). This is a recent and complete series with a mean follow-up of 10 months (range: 3-31). The primary endpoint was excessive weight loss (EWL) and the secondary endpoints were tolerance and morbidity. RESULTS: There were no postoperative deaths. The mean operating time was 129 minutes (range: 50-380). A conversion to laparotomy was necessary in 11 patients. The mean hospital stay was 4.76 days (range: 3-42). There were 29 complications (9.6%), 16 among the first 50 procedures: 14 patients underwent an abdominal reoperation (2 perforations, 3 early slippages, 7 late slippages, 2 incisional hernias); 6 had respiratory complications with 2 ARDS and 9 developed a complication related to the port. At one year, BMI decreased from 43.6 to 33.7 kg/m2 and EWL reached 44.2%; 80% of the patients lost 60% of their excess weight. CONCLUSION: Our experience is encouraging with an acceptable complication rate (5%) after 50 procedures. Slippage remains the main reason for close surveillance. Half of the excess weight can be comfortably lost in one year when the whole medical and surgical staff provide close support for each patient. PMID- 11255973 TI - [Hysteroscopic resection of submucous myomas: long term results]. AB - STUDY AIM: To evaluate the follow-up after operative hysteroscopic resection of submucous leiomyomas. PATIENTS AND METHODS: Between January 1990 and December 1996, 200 patients underwent operative hysteroscopic resection of 289 uterine leiomyomas. Indications were: menometrorrhagia (n = 159), postmenopausal metrorrhagia (n = 22), infertility (n = 19) as sole etiology. Sixteen patients had infertility and menometrorrhagia. RESULTS: The mean follow-up was 33.4 +/- 19.2 months. Twenty-three patients were lost to follow-up. Due to the large size of the leiomyomas, 35 patients had 2 or 3 resections and a total of 241 hysteroscopic resections were performed. Twelve complications (5%) occurred without death or need for intensive care. An improvement of clinical symptoms was observed in 74% of patients. The predictive factors of failure were: size (> 5 cm), number of intracavitary leiomyomas (> 3), hysterometry (> 12 cm), intramural myoma class 2 and association of leiomyomas. Eight of the 35 infertile patients subsequently became pregnant, but with only two live births (5.8%). CONCLUSION: Hysteroscopic myomectomy appears to be safe, effective and reproducible for the treatment of menstrual disorders. Intramural class 2 and larger leiomyomas constitute the limits of the endoscopic technique. PMID- 11255974 TI - [Traumatic rupture of an accessory spleen]. AB - A cyclist knocked over by a car was admitted with pain in the left upper quadrant that progressively worsened with haemorrhagic shock. Ultrasound showed a heterogenous spleen and perisplenic haematoma. Emergency laparotomy was performed and revealed rupture of an accessory spleen located on the splenocolic ligament. PMID- 11255975 TI - [Unusual testicular tumor: an ectopic spleen]. AB - The authors report a case of discontinuous splenogonadal fusion diagnosed after left orchidectomy. Ectopic spleen in the scrotum is a rare congential anomaly frequently associated with other anomalies, especially limb defects. The diagnosis is difficult. Preoperative isotope scanning and intraoperative pathological examination can be performed to avoid unnecessary orchidectomy. PMID- 11255976 TI - [How to chose a technique? Inguinal hernias]. PMID- 11255977 TI - [Multiple jejunal strictures caused by gastric heterotopia]. AB - A case of gastric heterotopia was discovered incidentally on a jejunal resection specimen in a 42-year-old patient operated for Koenig's syndrome present for 10 years. This anomaly was responsible for seven chronic ulcers with strictures at multiple levels. Gastric heterotopia, especially in the jejunum, is a rare anomaly, except in intestinal duplications and Meckel's diverticulum. The various complications are a direct result of the activity of the gastric glands: hemorrhage, Helicobacter pylori enteritis, perforation, chronic ulcer and obstructive syndrome; malignant adenocarcinomatous degeneration has also been reported. PMID- 11255979 TI - [Nissen by laparotomy or laparoscopy for gastroesophageal reflux. Randomized study]. PMID- 11255978 TI - [Jean Vaysse (1917-1975)]. AB - Jean Vaysse (1917-1975) participated during the 1950s et '60s in the great discoveries of modern surgery: renal grafts, cardiopulmonary bypasses, and surgery for arterial hypertension. He died prematurely. To right this injustice, to reveal the man out of the spotlight of hospital and university success stories though all the while participating decisively in some of surgery's greats movements, to find, above the pioneer, the questioning of the meaning of life, the nature of being and the knowledge of its private side: these are our intentions as we consider the life and work of Jean Vaysse. PMID- 11255980 TI - [Litigation and surgical practice in the United Kingdom]. PMID- 11255981 TI - [Randomized trial comparing primary versus secondary sigmoid resection in peritonitis due to sigmoid diverticulitis]. PMID- 11255982 TI - [New thrombolytic agents]. PMID- 11255983 TI - [Eflornithine for hair removal. Topical application for hirsutism]. PMID- 11255984 TI - [Are drug interactions predictable?]. PMID- 11255985 TI - [Malodor from the nose. Causes, diagnosis and therapy]. PMID- 11255987 TI - [Vegetarian nutrition--ideology or "evidence-based health benefit"?]. PMID- 11255986 TI - [Revaccination against pertussis. Recommendations from STIKO (Standing Vaccination Commission of the Robert Koch Institute)]. PMID- 11255988 TI - [Magnesium and sports?]. PMID- 11255989 TI - Tuberculosis. PMID- 11255990 TI - Unsafe injections: a potential source of HCV spread in Pakistan. PMID- 11255991 TI - Health care seeking behavior of pulmonary tuberculosis patients visiting TB Center Rawalpindi. AB - OBJECTIVE: To determine the health care seeking behavior of TB patients in seeking care and health care providers for delivering care. DESIGN: Cross sectional descriptive survey. The interviewers administered a standardized open ended questionnaire after training and pre-testing. SETTING: Federal Government TB Center, Rawalpindi. (specialized TB clinic). SUBJECT: One hundred and sixty newly registered TB patients at TB Center, Rawalpindi between 20 November to 21 December 1998. RESULTS: Prior to their consulting TB Center, 96% patients had already reported to a health care provider, i.e., to first, second or third health care providers. OF 154 patients, 48 were diagnosed as TB and only 29 (19%) of them received antituberculosis treatment. Most of the patients 118 (77%) consulted the health care provider within three weeks time. CONCLUSION: Delay is more on part of health care providers than on patients. Proper implementation of guidelines of TB program with public-private sector collaboration and continuing education of health care providers is necessary. PMID- 11255992 TI - Female genital tuberculosis revisted. AB - OBJECTIVE: To assess the clinical presentation of genital tuberculosis and to study various modes of diagnosis and treatment. SETTING: The Aga Khan University Hospital (AKUH), Karachi. METHOD: A retrospective case review of all index female cases of genital tuberculosis, admitted to AKUH over twelve years of period. RESULT: A total of 40 cases of genital tuberculosis were reported during this time period. Majority of cases were between 25-45 years. The commonest presenting symptoms were infertility (42.5%) and abdominal pain (42%). Others included fever, ascites, irregular vaginal bleeding, oligomenorrhea, chest pain and pain in the flanks. Main mode of treatment was antituberculous drug therapy for duration of nine months. Only 3 patients had successful pregnancies. CONCLUSION: Genital tuberculosis should be excluded when managing infertility in females. PMID- 11255993 TI - Morphological spectrum of ophthalmic tumors in northern Pakistan. AB - OBJECTIVE: The objective of the study was to assess the frequency and pattern of ophthalmic tumours in Northern Pakistan. METHODS: This study included all ophthalmic tumours diagnosed during a one year period (January to December 1992). RESULTS: One hundred and fourteen ophthalmic tumours were diagnosed at the Armed Forces Institute of Pathology (AFIP) and Pathology Department of the Army Medical College (AMC), Rawalpindi. Of these tumours, 70 were malignant (61.5%) and 44 were benign (38.5%). The age distribution of malignant ophthalmic tumours had two peaks. The first was seen in the paediatric age group and was mainly due to retinoblastoma. The second peak was seen above 50 years of age and was mainly due to conjunctival squamous cell carcinoma and malignant eyelid tumours which constituted 85% of the malignant ophthalmic tumours in paediatric age group. The average age at presentation of retinoblastoma was 3.8 years. The average age at presentation for squamous cell carcinoma was 56 years. Basal cell carcinoma was the most common malignant eyelid tumour (55%). The most common extraocular malignant orbital tumour was non-Hodgkin's lymphoma. Malignant melanoma of the uvea formed 22% of all melanomas diagnosed during this period. The most common benign tumours were naevi (33%), epidermal inclusion cysts (18%), choristomata (16%) and haemangioma (8%). The malignant ophthalmic tumours constituted 3% of all the malignant tumours diagnosed in Northern Pakistan during 1992 at AFIP and AMC, Rawalpindi. CONCLUSION: The ophthalmic tumours, both benign and malignant are not infrequent in clinical practice in Northern Pakistan. PMID- 11255994 TI - Urbanisation and coronary heart disease risk factors in South Asian children. AB - BACKGROUND: Coronary Heart Disease (CHD) and other Non Communicable Diseases (NCDs) are increasing globally. Comparison of various sections of the South Asian populations living at different levels of urbanization can help in understanding the role of demographic transition in the increased prevalence of these diseases in urbanized populations. OBJECTIVE: To compare the prevalence of certain CHD risk factors in 10-12 year old school children living at different levels of urbanization. METHOD: Differences in height, Body Mass Index (BMI), Waist Hip Ratio (WHR), Fasting Blood Glucose (FBG) and Total Blood Cholesterol (TBC) were studied. SUBJECTS: Anthropometric and biochemical measurements of six groups of 10-12 year old children, representing various urbanization categories, were studied. Three groups of children were recruited from Punjab, Pakistan: rural, middle income urban and high income urban and they were assigned urbanization rank (UR) 1, 2 and 3. Another three groups of children were recruited from Slough, UK: British Pakistani, British Indian, and British Caucasian and they were assigned urbanization rank 4, 5 and 6 respectively. RESULTS: Proportion of children having high CHD risk increased with urbanization rank. Increase in BMI and TBC with urbanization status was steadier than the increase in FBG and WHR. Stunting which have been found to have a positive association with obesity and increased risk of CHD was higher among the less urbanized groups. BMI and TBC of the urbanized South Asian groups were lower, but FBG was higher than the British Caucasian, who served as controls. CONCLUSION: These findings support the hypothesis that high CHD death rate among South Asians in UK may have its origin in the genetic predisposition to diabetes but are not likely to be solely due to this factor. The environmental factors like under nourishment in early life, adoption of urbanized life style or a combination of both could be the major determinants of CHD morbidity and mortality. PMID- 11255995 TI - Nitric oxide mediated effect of cyclo-oxygenase inhibitors. AB - OBJECTIVES: Non-Steroidal anti-inflammatory drugs (NSAIDS) have long been used as anti-inflammatory agents, yet their mode of action is not entirely clear. The inhibitory effects of NSAIDS on prostaglandin production can only partly explain their anti-inflammatory actions. This study was aimed at defining the role of cycl-oxygenase (COX) inhibitors on nitric oxide (NO) production in murine macrophages in vitro. METHODS: Murine macrophages were obtained from the peritoneum and after exposure, in vitro to lipopolysaccharide (LPS) produced nitrite, measured after 24 hours by Griess reaction. The macrophages were pre incubated with aspirin or indomethacin before activation with LPS. RESULTS: Treatment with aspirin resulted in an increase in nitric oxide production. A similar response was obtained with indomethacin treatment. CONCLUSION: This study shows that COX inhibitors significantly increase NO production in murine macrophages in vitro and this may be one of the mechanisms by which they exert their anti-inflammatory effects. PMID- 11255996 TI - The dynamics of tuberculosis treatment adherence. AB - AIMS: To establish various factors that affect TB treatment adherence over time. DESIGN/SETTING: Semi-structured questionnaire. All newly diagnosed cases of TB at Bethamia Hospital, Sialkot were interviewed at the beginning of treatment, one month of therapy and at the end of intensive phase. RESULTS: Perception of TB as a stigmatising disease was found related to early defaulting and to a lesser degree to late defaulting. Knowledge of TB in itself did not have a clear impact on defaulting, but the attitude towards interruption of treatment did. The strongest risk factor is irregularity of drug intake and appointment keeping. CONCLUSION: Strategies to improve treatment adherence should concentrate on methods to increase patient's motivation for treatment. PMID- 11255997 TI - Serum leptin levels in pregnant Pakistani females: relationship with body mass index and placental weight. AB - OBJECTIVE: To determine the serum leptin levels in Pakistani pregnant subjects at the time of delivery and to ascertain the relationship between serum leptin levels and related variables (weight, body mass index, placental weight, gestational age, parity) at delivery. SETTING: Lady Dufferin Hospital, and Ziauddin Hospital Karachi. METHODS: Leptin concentration was measured in 110 subjects from venous samples, using Active Human ELISA Kit (DSL-10-23100). Samples were selected according to the availability. RESULTS: Mean maternal weight, body mass index and placental weight were 64.3 +/- 13.8 kg, 27.1 +/- 5.8 kg/m2 and 523.5 +/- 90 gm, respectively. Gestational age was 36-41 weeks and maternal age was 18-35 years. Mean serum leptin level was 27.9 +/- 18.1 ng/ml. Serum leptin levels were found to be positively correlated with body weight (r = 0.73, p < 0.01), body mass index (r = 0.80, p < 0.01) and placental weight (r = 0.34, p < 0.05). Increased leptin levels were found in multigravida mothers (mean 30.2 +/- 17.5) than in primigravida mothers (23.1 +/- 18.9). CONCLUSION: Our results suggest that leptin does play role in body weight and energy regulation during pregnancy. The significant positive correlation between leptin and placental weight suggests that placenta may be the site of synthesis and/or secretion of leptin during pregnancy. PMID- 11255998 TI - Prevalence of histological reflux oesophagitis in H. pylori positive patients: effect of density of H. pylori and activity of inflammation. AB - OBJECTIVE: Recently there has been a great interest in the role of Helicobacter pylori in gastroesophageal reflux disease. Many studies do not show any significant difference in the overall prevalence of H. pylori in patients with endoscopic oesophagitis and controls. In this prospective study we assessed the influence of H. pylori density and activity of inflammation in different parts of stomach on histological oesophagitis. METHODS: One Hundred and forty consecutive patients undergoing endoscopy for dyspepsia and heartburn were evaluated. Three biopsies were taken from antrum and two each from corpus, cardia and lower oesophagus. Urease test (CLO test) was performed. Density and activity of infection was assessed in a semi-quantitative way. RESULTS: One Hundred and Fourteen (81%) patients from the 140 endoscoped, were positive for H. pylori and had H. pylori positive antral gastritis. Of these 114 cases, H. pylori were detectable in 104 (91%) of biopsies taken from corpus and 96 (84%) of biopsies from cardia. There was a strong correlation of density of H. pylori (0-3) in antrum with body and of body with cardia by Spearman correlation tests (p = 0.000). But H. pylori were more dense in antrum as compared to corpus and in corpus as compared to cardia (p = 0.0000 and 0.0003 respectively by Wilcoxon's rank test). Neutrophil activity and degree of mononuclear infiltrate were also greater in antrum as compared to corpus (p = 0.000 and 0.059). The activity and degree of inflammation was not significantly different in corpus-cardia pair. Out of 114 H. pylori positive patients, 75 had histological oesophogitis (p = 0.855). After excluding cases of hiatal hernia (H.H) and gapping lower oesophageal sphincter (LOS), number of H. pylori positive patients decreased to 73, out of these 50 had histological oesophagitis (p = 0.103). In all H. pylori positive patients with histological oesophagitis, H. pylori density (1-3) in antrum correlated with severity of oesophagitis (P = 0.011). Neutrophil activity in antrum and corpus also correlated with the severity of histological oesophagitis (P = 0.024 and 0.035 respectively). Correlation further improved after excluding cases of HH and gapping LOS (P = 0.002 for H. pylori density and 0.026 and 0.004 for activity in antrum and corpus). No correlation could be found of density and activity of infection in cardia with histological oesophagitis. CONCLUSION: Our H. pylori positive patients had more dense and severe infection in antrum. Those who had histological oesophagitis in addition showed a positive correlation of the density of H. pylori in antrum and neutrophil activity in antrum and corpus with the severity of histological oesophagitis. PMID- 11255999 TI - Tuberculosis control in Pakistan: critical analysis of its implementation. PMID- 11256000 TI - Adenomyosis with tuberculosis of uterus. PMID- 11256001 TI - The proposition: an insight into research. AB - Propositions form the basis for scientific research. The validity of a research study is, to a large extent, evaluated on the criteria of its propositions. For internal validity, study propositions provide information regarding precision of definitions, measurements, associations, confounding factors etc. that are considered in research. While for external validity, propositions form the premise for the deduction of inferences. The aim of this article is to help readers understand the propositions that are made in research. This article discusses those propositions, which are relevant to medical research. PMID- 11256002 TI - Can snoring kill? PMID- 11256003 TI - [Neuroscience within the program of biomedical research at the University of Navarra]. PMID- 11256004 TI - Intraoperative high dose rate brachytherapy can be used to salvage patients with previously irradiated head and neck recurrences. AB - PURPOSE: To develop a novel technique, intraoperative high-dose rate brachytherapy (IOHDR), in the treatment of previously irradiated head and neck cancers located at anatomical sites inaccessible to intraoperative electron beam radiotherapy (IOERT). METHODS: Between October 1992 and June 1997, seven patients (median age = 65 yrs; range = 52 to 71 years) with previously irradiated head and neck recurrences at anatomical sites inaccessible to IOERT in the base of skull were treated with IOHDR after maximal resection for microscopic residual disease. Treatment volume ranged from 6 cc to 24 cc. Six patients received 15 Gy of IOHDR at 0.5 cm; one received 10 Gy using custom-made surface foam applicators. RESULTS: The median follow-up was 59 months (range 33 to 67 months). It was technically feasible to deliver IOHDR in all seven patients at sites that were inaccessible to IOERT. The morbidity (observed in two patients) was acceptable and generally surgically related. Four of seven patients (57%) were locally controlled at IOHDR site. Two failed regionally, outside the IOHDR treated sites. The disease-free survival ranged from 3 to 30 months (median 9 months) with two patients still alive, disease-free at 28 and 30 months. CONCLUSIONS: IOHDR can be used, with limited toxicity, to treat previously irradiated head and neck cancers at sites inaccessible to IOERT. We are currently evaluating the addition of limited EBRT dose to improve the local control of these poor prognosis recurrent tumors. PMID- 11256005 TI - [Silicone in autoimmune diseases and cancer]. AB - In 1992 the Food and Drug Administration (FDA) announced the restriction of silicone gel-filled breast implants until research protocol studies evaluate the relationship of silicone to connective tissue diseases, and the association of the silicone implants with breast carcinoma. Since them comprehensive epidemiologic studies have concluded that there is no connection between breast implants and the known connective tissue diseases or between the implants and breast carcinoma. During the same year, The American College of Rheumatology said that it have not been demonstrated the relationship between silicone gel breast implants and any systemic disease. Although this, the FDA restriction continues. PMID- 11256007 TI - [Cystic pyeloureteritis]. AB - The pyelitis or cystic pyeloureteritis is a rare disease of unknown etiology. The clinic is unspecific and the treatment, medical and expectant. The importance of this disease consists of a correct differential diagnosis with other repletion defect imaging in the excretory tract and its frequent association to other diseases. PMID- 11256006 TI - [Epidemiology of tumors of the renal parenchyma]. AB - Renal cell carcinoma is responsible for about 2% of all cancer deaths in developed countries and represents 80-85% of all tumors of the kidney. Its etiology is still largely undefined. Its incidence varies among countries, with the highest rates in North Americans and Scandinavians. Its incidence is steadily rising in the last ten years. The location of the tumor suppressor gene on chromosome 3p has contributed to the understanding of tumor pathogenesis. Renal cell carcinoma occurs nearly twice as often in men as in women. Patients are generally more than 40 years old at diagnosis, usually in the fifth to seventh decade of life. This tumor is more common among urban than rural residents, but it was not a consistent association with education or socio-economic status. Recently large epidemiologic studies showed an increased risk of renal-cell cancer in relation to tobacco smoking, with a relative risk of about 2 for current smokers. Other established risk factors are elevated body mass index (mainly in women) and a family history of the disease. Occupational exposure to chemicals appears to have little significance, although associations with specific products, such as asbestos fibres, have been reported. Some relationship has been observed between renal-cell cancer and hypertension, use of anti hypertensives and kidney diseases, although this issue remains open to discussion. Data are inconsistent on the role of nutrition, mainly for fats and proteins, while vegetable and fruit consumption seems to convey some protection on renal-cell cancer risk. The risk of renal-cell cancer was not materially elevated in relation to coffee, tea and alcohol intake and, in women, oral contraceptive use, hormone replacement therapy, and menstrual factors. PMID- 11256008 TI - [The mind-brain problem as seen by a neurobiologist]. PMID- 11256009 TI - [Severe hepatotoxicity caused by amiodarone: description of a case]. AB - Amiodarone is useful for the treatment of ventricular and supraventricular arrhythmias, but it has been associated with several adverse effects. Amiodarone induced hepatotoxicity is a frequent complication and often consist in a mild and asymptomatic elevation of liver function tests, although severe cases have been described. A case of severe amiodarone-induced hepatotoxicity slowly resolving after discontinuation of the drug and in which liver biopsy was crucial for diagnosis is reported. Although there are no pathognomonic histopathological findings, phospholipidosis is the morphologic hallmark of amiodarone hepatotoxicity. PMID- 11256010 TI - [Fexofenadine: a antihistaminic. Review of its practical characteristics]. PMID- 11256011 TI - Goodbye to all that. An old man dying and a little girl kissing his hand. PMID- 11256012 TI - Need a mammogram? It could take a while. PMID- 11256013 TI - Do it yourself? Those easy-to-use home medical tests can be anything but. Some are downright misleading. PMID- 11256014 TI - [Diagnosis and treatment of somatoform disorders (functional physical complaints)]. AB - In modern classification systems, unexplained physical symptoms without an organic origin are labelled "somatoform disorders". This syndrome is very frequent and causes a lot of treatment costs as well as indirect costs (such as workers' compensation and others). In the past, effective treatment strategies were lacking. However, consideration of modern scientific results has made it possible to develop treatment approaches which find the acceptance of the patients and which are highly effective. A hierarchical approach is presented suggesting the following steps: a) Primary care: The consideration of management rules in primary care can prevent the chronicity of somatoform symptoms. b) Brief psychological and psychopharmacological treatments: Modern cognitive-behavioural approaches can help to cope with the symptoms and to improve the subjective well being. First results for pharmacological treatments are encouraging. c) Integrative inpatient treatment including intense psychotherapeutic and psychosomatic ingredients. Experts' judgements of course and prognosis in somatoform disorders should consider the following features: Duration and multiplicity of the complaints; comorbidity with other psychiatric and physical disorders; disability in different areas of life such as at work, family, leisure time; individual coping strategies; treatment approaches in the past. PMID- 11256015 TI - [Depressive disorders in children and adolescents]. AB - Depression is a fairly common disorder in children and adolescents. Symptoms include depressed mood, reduced energy, cognitive dysfunction and additional physical complaints. Depression may require in-patient treatment. In spite of effective psychopharmacological and psychotherapeutic treatment methods, depressive disorders tend to persist. PMID- 11256016 TI - [Psychological sequelae of accidents. A problem in accident and liability insurance]. AB - Active and successful management predominantly depends on good and trustful cooperation between the claims manager, the field representatives of the administration and the injured or damaged person. Only the early assessment and prompt and purposeful claims handling can minimize the risk of an unexpected claims development. In the early stage, particularly probands with acute psychological maldevelopment can still be helped effectively. The introduction of an adequate therapy is beneficial to the afflicted person and at the same time lowers the final developing costs. As early as in 1918, Horn stated that the early occupational reintegration, apart from granting compensation, positively affects the process of the psychological disturbance. In general, no substantial restriction on the quality of life occurs if the injuries of a physical and mental type have been treated successfully and if occupational reintegration has taken place. However, if a continuous performance loss remains in occupational life due to a chronic psychological disturbance and if the damaged person does not achieve his/her ability to work again, even the contribution of high compensation payments does generally not improve the quality of life. PMID- 11256017 TI - [Interdisciplinary management of chronic tinnitus (I)]. PMID- 11256018 TI - [Pain--diagnosis and therapy by organ specialists?]. PMID- 11256019 TI - [Evidence-based medicine and private health insurance]. PMID- 11256020 TI - [Comment on R. B. Pelka: Smoking and mortality]. PMID- 11256021 TI - [Significance of medical statistics in insurance medicine]. AB - Knowledge of medical statistics is of great benefit to every insurance medical officer as they facilitate communication with actuaries, allow officers to make their own calculations and are the basis for correctly interpreting medical journals. Only about 20% of original work in medicine today is published without statistics or only with descriptive statistics--and this trend is falling. The reader of medical publications should be in a position to make a critical analysis of the methodology and content, since one cannot always rely on the conclusions drawn by the authors: statistical errors appear very frequently in medical publications. Due to the specific methodological features involved, the assessment of meta-analyses demands special attention. The number of published meta-analyses has risen 40-fold over the last ten years. Important examples for the practical use of statistical methods in insurance medicine include estimating extramortality from published survival analyses and evaluating diagnostic test results. The purpose of this article is to highlight statistical problems and issues of relevance to insurance medicine and to establish the bases for understanding them. PMID- 11256022 TI - [Intima-media thickness in healthy probands without risk factors for arteriosclerosis]. AB - BACKGROUND AND OBJECTIVE: Ultrasound examination of the intima-media thickness (IMT) is a well accepted, highly reproducible method for the evaluation of early atherosclerosis. However, for the exact interpretation of these measurements reference values for IMT should be determined. The present work provides for the first time normal values for IMT of the common carotid artery and the arteries of the lower limbs (common femoral, superficial and popliteal arteries) in a German population of healthy subjects without risk factors. SUBJECTS AND METHODS: A total of 112 subjects (50 men, 62 women), aged 40 to 70, were examined, according to the following inclusion criteria: no diabetes or marked elevation of postprandial plasma glucose; non-smokers; no hypertension, obesity, dys/hyperlipidaemia or albuminuria; no history of coronary heart disease, stroke and peripheral arterial occlusion. IMT was measured and standardized by the method of Pignoli. Total cholesterol, triglycerides, HDL-cholesterol, HbA1c, plasma glucose (fasting and postprandial) and albuminuria were examined by routine methods. RESULTS: The mean values of double measurements bilaterally in the distal part of the common carotid artery in men were 0.79 (0.73 ... 0.84) mm (40-54 years) and 0.87 (0.81 ... 0.93) mm (55-70 years). In women the values were: -0.70 (0.67 ... 0.75) mm and 0.82 (0.75 ... 0.90) mm, resp. In the group aged 40-54 years the men showed significantly higher IMT of all examined vessels in comparison to the women. In the group aged 55-70 years somewhat higher IMT of the common carotid artery and significantly higher IMT of the arteries of the lower extremity were observed in men. In multivariate analysis age was found to be a significant determinant of IMT of all examined vessels. CONCLUSION: Intima media thickening of the common carotid artery in men aged 40-70 years is to be accepted if a single measurement of IMT exceeds > or = 1 mm. For women, aged 40 54 years, IMT is defined as pathological if one IMT is above 0.85 mm, and for those aged 55-70 years if IMT exceeds 1 mm. PMID- 11256023 TI - [Bendamustine, vincristine, prednisolone (BOP) in therapy of advanced low-grade non-Hodgkin lymphoma]]. AB - BACKGROUND AND OBJECTIVE: Low grade non-Hodgkin lymphomas (l-NHL) are rarely showing complete or sustained remissions to conventional chemotherapy. Thus, many therapeutic strategies try to improve the remission rates and outcome in relapsed and refractory l-NHL. Bendamustine (B) is a non-cross resistant alkylating agent shown to be highly effective in lymphoproliferative and other malignant diseases. In an open phase-II study we evaluated the efficacy and toxicity of B in combination with vincristine (O) and prednisolone (P) in heavily pretreated relapsed or refractory l-NHL. PATIENTS AND METHODS: 22 patients (median age 61.5 years, range 39-77 years) with relapsed or refractory low grade NHL: immunocytoma (IC) n = 11, centroblastic-centrocytic (CB-CC) n = 6, centrocytic (CC) n = 2, others n = 3, were treated with BOP as follows: patients up to 75 years: 60 mg/m2 B for 5 days; patients over 75 years: 50 mg/m2 B for 5 days. All patients received 2 mg vincristine (O) on day 1, 100 mg/m2 prednisolone (P) on day 1-5; repetition day 29. Prior to BOP patients were pretreated with 1-4 chemotherapy protocols. An average of 5 courses of BOP were administered (range 2-8). In most patients BOP was followed by a maintenance therapy (IFN-alpha n = 11, chlorambucil n = 4, etoposide n = 2). RESULTS: Objective remission was achieved in 19/22 (86%) patients, complete remission (CR) in 10/22 (45%), partial remission (PR) in 9/22 (41%) and no change (NC) in 3/22 (14%) patients. The mean duration of remission was 16.1 months. Predominant features of side effects of the BOP protocol were myelotoxicity of WHO grade III/IV in 8 of 109 cycles leukopenia (8%), thrombocytopenia 3 cyles (3%) and anaemia in 4 cycles (4%). We observed one WHO grade IV infectious episode. Other side effects were mild and rare. There was a decline of the CD4/8 in more than 50% of patients. However, these changes were not accompanied by a higher rate of infectious episodes. CONCLUSION: Salvage therapy of refractory and relapsed l-NHL with BOP results in a high objective remission rate. Together with a maintenance therapy most patients achieved a long-term disease-free survival. Myelotoxicity and the inversion of the CD4/CD8 ratio were frequently observed side effects. PMID- 11256024 TI - [Paraneoplastic Bazex acrokeratosis in adenocarcinoma of the stomach]. AB - HISTORY AND EXAMINATION: A 58-year-old man presented with a sudden eruption of hyperkeratoses on hands and feet. Alcohol abuse had been present 10 years ago, together with a gastric ulcer. Physical examination was unremarkable, except for plantar bizarre hyperkeratoses especially at the sides of the feet as well as some rhagades with spread to the dorsum of the toes. Similar but less severe hyperkeratoses were found on the palms. INVESTIGATIONS: Laboratory investigation was essentially normal. X-rays of the thorax and sinuses as well as a sonography of the stomach revealed, except for a fatty liver, no pathological findings. Gastroscopy revealed a chronic active gastritis on the small curvature of the stomach. Only a second gastroscopy and biopsy revealed an adenocarcinoma of the stomach. DIAGNOSIS, THERAPY AND COURSE: According to the clinical presentation a paraneoplastic acrokeratosis (Bazex syndrome) was diagnosed. The patient was given high-dose chemotherapy with subsequent stem-cell transplantation and gastrectomy. The skin problems resolved after the first cycles of chemotherapy. CONCLUSION: If clinically a paraneoplastic acrokeratosis is suspected extensive search for a malignancy has to be initiated, including a gastroscopy, because rarely, but as in the present case, a carcinoma of the stomach can be the underlying cause. PMID- 11256025 TI - [Current diagnostic methods for detection of Helicobacter pylori infection]. PMID- 11256026 TI - [Use of radiosynoviorthesis in inflammatory rheumatic joint diseases]. PMID- 11256027 TI - [Antioxidants in nutrition and arteriosclerosis]. PMID- 11256028 TI - [Abortions in forced female labor in the Third Reich]. PMID- 11256029 TI - [Infusion therapy with pentoxifylline and/or hydroxyethyl starch]. PMID- 11256030 TI - [Release of hospital discharge records to health insurance and medical offices of health insurance]. PMID- 11256031 TI - [Bone tumor]. PMID- 11256033 TI - [12 years experience with the physician in the practicum]. PMID- 11256032 TI - [Indications and results of video thoracoscopic sympathectomy]. PMID- 11256034 TI - [DMW theme and focal point issue: endocrinology]. PMID- 11256035 TI - [Treatment of obesity--where are we today?]. PMID- 11256036 TI - [Comparison of the effectiveness of two different dosages of levothyroxine-iodide combinations for the therapy of euthyroid diffuse goiter]. AB - BACKGROUND AND OBJECTIVE: Administration of levothyroxine and/or iodide can effectively reduce the volume of endemic goitre. However, TSH suppression during levothyroxine treatment may increase the number of recurrences through the persistence of intrathyroidal iodine deficiency. With special attention paid to the level of levothyroxine, a comparison was made of two dosages of combined levothyroxine and iodide. PATIENTS AND METHODS: 44 patients with diffuse euthyroid goitre were randomized to two treatment groups. Group A received 100 micrograms levothyroxine + 100 micrograms iodide, group B 75 micrograms levothyroxine + 150 micrograms iodide, all of them for three months. This was followed by three months without the medication. Intrathyroidal iodine concentration was measured at the onset of the study, then three months and six months later. At these same times thyroid volume was measured by ultrasound, as well as urinary iodine and various parameters of thyroid function. RESULTS: Thyroid gland volume was reduced in both groups (group A: -17.3%; group B: 14.8%; p < 0.001). There was no significant difference of intrathyroidal iodine concentration and thyroid volume between both groups. After the treatment period, TSH suppression was more marked in group A, while TSH rise was greater in group B. CONCLUSIONS: Both drug combinations resulted in comparable reduction of thyroid volume, while the intrathyroid concentration of iodine remained unchanged. The smaller rise of TSH after the treatment suggests that the dosage of 75 micrograms levothyroxine + 150 micrograms iodine is to be preferred. PMID- 11256037 TI - [Transsphenoidal hypophysectomy of a patient with an ACTH-producing pituitary adenoma and an "empty sella" after pretreatment with etomidate]. AB - HISTORY AND ADMISSION FINDINGS: A seventy-year-old woman was admitted to our hospital under suspicion of an adrenal Cushings's syndrome. Initial laboratory values showed elevated cortisol (834 nmol/l; normal: 180-640) which could not be suppressed after administration of 2 mg dexamethasone (632 nmol/l). Computed tomography of the abdomen showed a 19 x 34 mm mass in the region of the left adrenal gland. INVESTIGATIONS: ACTH levels were normal (42 ng/ml; 17-52). Serum cortisol remained high at 1021 nmol/l after administration of 8 mg dexamethasone. Four more doses of 2 mg dexamethasone were applied on 3 consecutive days, leading to a mild suppression of serum cortisol to 705 nmol/l, with urine cortisol levels dropping from 1915 to 101 nmol/l/24 h. The CRH-test produced a rise of serum cortisol from 895 to 1475 nmol, with ACTH rising from 42 to 68 pg/ml, a laboratory constellation consistent with the diagnosis of centrally located Cushing's syndrome (Cushing's disease). MRI failed to show an adenoma of the pituitary gland so that sinus petrosus sampling was done to confirm the diagnosis (ACTH central/peripheral 7:1; normal range < 2:1). TREATMENT AND COURSE: Serum cortisol rose to 1070 nmol/l and the patient developed pneumonia and contracted tinea. Prior to surgery we lowered the excessive cortisol levels with etomidate and successfully treated the pneumonia with antibiotics. Postoperatively clinical symptoms of Cushing's syndrome disappeared. The patient now presented with total insufficiency of the anterior pituitary. CONCLUSION: Exact hormone testing that may involve sinus petrosus sampling is necessary in diagnosing Cushing's syndrome. Even if radiological procedures cannot show an adenoma of the pituitary, transsphenoidal resection should be considered. Etomidate can lower excessive cortisol levels in seriously ill patients. PMID- 11256038 TI - [Predictive examinations of phenotypically healthy offspring of patients with a metabolic syndrome]. PMID- 11256039 TI - [Some may feel hot: significance of thermogenesis for energy metabolism and the treatment of obesity]. PMID- 11256040 TI - [Androgen deficiency in older men--what happens with testosterone substitution?]. PMID- 11256041 TI - [Hormone replacement therapy after menopause]. PMID- 11256042 TI - [Hormone replacement therapy--treatment with risk factors]. PMID- 11256043 TI - [Rapid and reliable detection of multiresistent Staphylococcus aureus (MRSA) by multiplex PCR]. AB - BACKGROUND AND OBJECTIVE: Staphylococci are widespread pathogens and are frequently associated with nosocomial infections. Many hospitals struggle with increasing amounts of methicillin-resistant Staphylococcus aureus (MRSA) which are "multiresistant" against all betalactam antibiotics. Often, applicable antibiotics for treatment are only glycopeptides like vancomycin and teicoplanin. In addition, MRSA infected patients require expensive intensive isolation measures and strict hygiene. To efficiently prevent dissemination of these pathogens rapid and reliable identification and a close collaboration between clinicians and microbiologists are required. The purpose of our study was to set up a rapid and reliable identification procedure for MRSA by the amplification of specific gene determinants by PCR in order to to efficiently support therapy and eradication of the pathogen. METHODS: 153 strains of staphylococci isolated from in-patients of the hospital of the Justus-Liebig University of Giessen were examined. The femB gene was used to differentiate between Staphylococcus aureus (S. aureus) and coagulase-negative staphylococci (CNS), a gene which allows the species-specific identification of methicillin-resistant (MRSA) and -susceptible S. aureus (MSSA). Additionally, MRSA harbor the mecA gene encoding methicillin resistance, which is absent in MSSA strains. RESULTS: Using a multiplex PCR with femB and mecA gene-specific oligonucleotides MRSA strains were unequivocally detected within 3 hours. The femB gene was detected in all 102 strains of S. aureus but in none of the 51 CNS. The mecA determinant was detected in 12 S. aureus. Among these, 11 strains were phenotypically methicillin-resistant and one strain was susceptible. The methicillin-resistance of this particular mecA positive/methicillin-susceptible strain (cryptic MRSA) was inducible by cultivation on agar plates supplemented with flucloxacillin. CONCLUSIONS: The described method specifically detects S. aureus and identifies phenotypical and cryptic MRSA. These cryptic MRSA are of particular relevance since they are undetectable using common phenotypically based detection methods. It is conceivable that the methicillin resistance of these strains is induced under antibiotic therapy with flucloxacillin and that the mec-encoded feature of methicillin-resistance can be transferred to previously methicillin-susceptible strains. Using the reliable detection of these strains by PCR, failure of flucloxacillin therapy is avoidable. PMID- 11256044 TI - [Deep sedation in gastrointestinal endoscopic interventions: safety and reliability of a combination of midazolam and propofol]. AB - BACKGROUND AND OBJECTIVE: Midazolam (M) is well known and established in endoscopic procedures for so-called conscious sedation. Propofol (P) is given during endoscopy for deep sedation. The combination of both (M/P) in endoscopic procedures is new. In this prospective study the safety of the combination was tested, in a second prospective study the combination M/P with Propofol alone was compared. PATIENTS AND METHODS: In the first study 143 patients undergoing 150 endoscopic procedures (expected > 30 min) were included. Deep sedation was induced by an i.v. bolus of 2.5 mg midazolam, followed by small doses of propofol. The sedation was performed by a second physician experienced in intensive care. In the second prospective study 64 patients undergoing two necessary endoscopic procedures were included: one deep sedation with propofol alone, the second one with combination of midazolam bolus followed by propofol. RESULTS: It was demonstrated that induction of deep sedation by an i.v. bolus of 2.5 mg midazolam, followed by small doses of propofol is safe, without undesirable side effects, e.g. respiratory or circulation depression. Recovery time increased (4 to 8 minutes) with the combination M/P, costs decreased by saving 59% propofol per minute. This may be important for longer endoscopic procedures. CONCLUSION: The combination of M/P for deep sedation during endoscopy may be useful when long procedures are expected or patients are at risk with propofol alone. PMID- 11256045 TI - [Leptospirosis after a staff outing]. AB - HISTORY AND CLINICAL FINDINGS: Three male colleagues aged between 34 and 38 years were admitted at the same time to three different Rhein-Main area Hospitals. They presented with a variety of symptoms, including high fever (39.0 to 40.0 degrees C), chills, headache with meningismus or facial paralysis, mild hepatitis and renal involvement. About 18 days before they had been together on a boat rafting tour when the boat capsized when they had fallen into a river in high flood. INVESTIGATIONS: Laboratory tests showed elevated inflammatory parameters, signs of a mild hepatitis and renal involvement. All patients had leptospirosis antibodies, detected by immunofluorescence test. In two cases there was evidence of antibodies against Leptospira interrogans serovar bataviae in the microscopic agglutination test (MAT). TREATMENT AND COURSE: The history and clinical presentation indicated leptospirosis in all patients, in two cases confirmed by laboratory findings. Following therapy with doxycycline or ceftriaxone, symptoms resolved quickly and permanently. CONCLUSION: Leptospires of serogroup Bataviae is a known pathogen of anicteric non-Weil leptospirosis. The symptoms are non specific and, moreover, in some cases the laboratory tests are negative, so that clinical diagnosis remains crucial. Typically there is a history of contact with contaminated water or urine. In our cases striking neurotropism was observed, which may be characteristic for this serovar. PMID- 11256046 TI - [Spontaneous rupture and embolization: a rare complication of port catheter implantation]. AB - HISTORY AND CLINICAL FINDINGS: A 70-year-old male patient had a venous port catheter implanted into his right subclavian vein for neoadjuvant radio chemotherapy of a rectal carcinoma (T3N0N0). Due to the patient's difficult venous access the catheter was left in situ after treatment. 31 weeks later he was admitted to the hospital because of parasternal and subclavicular pain. INVESTIGATIONS: Physical examination and an electrocardiogram revealed no abnormalities. A chest x-ray was performed. DIAGNOSIS, TREATMENT AND COURSE: The chest x-ray showed a normal location of the port-system but the tip of the catheter had embolized into the right atrium. The embolized fragment was extracted with a loop-snare technique and the reservoir of the system was removed under local anaesthesia without any complications. CONCLUSIONS: Despite its frequent use intravascular embolization of catheter fragments from implantable venous port-catheter systems present a rare but potentially life-threatening complication. Any implanted catheters should therefore be removed after completion of treatment or the system's integrity should be monitored on a regular basis. PMID- 11256047 TI - [Entrapment syndrome of the popliteal artery. A rare, but dangerous etiology of intermittent claudication in young patients]. PMID- 11256048 TI - [Cardiac arrhythmias in hyperthyroidism]. PMID- 11256049 TI - [Prospective epidemiological study of epigastric complaints (PRESTO)]. PMID- 11256050 TI - [Prospective epidemiological study of epigastric complaints (PRESTO)]. PMID- 11256051 TI - [Persistence of Chlamydia pneumoniae in coronary plaque tissue]. PMID- 11256052 TI - [IZKF W"urzburg: interdisciplinary is news]. PMID- 11256053 TI - [Italian forensic psychiatry: a story to be written]. PMID- 11256054 TI - Epidemiology of violence and mental illness. PMID- 11256055 TI - From homicide inquiries to high secure hospital inquiries: a decade of social and political restrictive pressure on forensic psychiatry. PMID- 11256056 TI - The growing public health importance of common mental disorders. The forgotten reality of the less developed world. AB - Non-communicable diseases are growing rapidly and mental problems represent a large proportion of this group. Most mental health burden is to be found in the community and at primary health care. There are large disparities in prevalence rates between and within countries. Latin American countries have consistently shown higher prevalence rates than most other countries in the world. PMID- 11256057 TI - Instrument development in the Italy-USA Collaborative Spectrum Project. AB - SCOPE: The Collaborative Spectrum Project aims to define subthreshold and atyical conditions not sufficiently characterized in the current diagnostic nomenclature and for which adequate assessment instruments are not available. This paper reports on the development and validation of new instruments to assess the spectrum of five psychiatric disorders. DESIGN: Three multicenter studies and one single-site study were conducted in Italy to assess the validity and reliability of the five spectrum interviews. Another cross-sectional study to validate the panic-agoraphobia spectrum has been conducted in Pittsburgh. SETTING: Outpatients attending various university clinics, university students and, in one Italian study, gym attenders were recruited for the studies. MAIN OUTCOME MEASURES: Five structured clinical interview to assess the spectrum of panic-agoraphobia (SCI PAS), mood (SCI-MOODS), social phobia (SCI-SHY), and the obsessive-compulsive (SCI-OBS) and eating disorder spectra (SCI-ABS) were administered along with a diagnostic interview and a number of self-report and interviewer-rated instruments. RESULTS: All the domains of the interview showed high test-retest reliability (intraclass correlation coefficient > 0.61) and satisfactory internal consistency. Mean domain scores were significantly higher in cases than in controls and in patients with the disorder of interest than in patients with other disorders. Convergent validity was satisfactory for panic-agoraphobia, social phobia and obsessive-compulsive spectrum domains. Differences emerged between SCI-ABS and self-report instruments assessing eating disorders. A cut-off score for the panic-agoraphobia spectrum was defined and its clinical validity was tested. CONCLUSIONS: The psychometric properties of the five spectrum interviews are very satisfactory, and studies are currently ongoing to test the clinical validity of all the spectra. Subthreshold and atypical symptoms deserve attention in epidemiological investigation. PMID- 11256058 TI - [Psychosocial problem disclosure during primary care consultations]. AB - OBJECTIVE: To examine how primary care patients with psychosocial problems actually introduce and present these topics. To examine the influence of some personality traits (emotional dependency and health locus of control) on psychosocial problem disclosure. DESIGN: "Case control". Cases had a GHQ-12 score equal or higher than three. Controls were matched with cases according to GP, GPs' attribution of absence or presence of emotional distress sex, age and presence of chronic illness. SETTING: Six single handed primary care practices. MEASURES: Clinical and Socio-demographic data, Social Problems List, List of 12 Threatening Life Events, GHQ-12, Multidimensional Health Locus of Control, Interpersonal Dependency Inventory and a Social Support Index. RESULTS: The introduction of psychosocial topics by patients is related to the attribution of emotional distress by GP. Patients identified correctly as not distressed present less often psychosocial topics compared to patients recognised as distressed. Distressed patients not recognised as such more often than the others did not offer any psychosocial cue. The personality measures did not influence the presentation of psychosocial problems. CONCLUSIONS: The lack of psycho-social cues from patients and of patient-centred skills contributed to the non recognition of emotional distressed patients. GPs' active facilitation of the presentation of psychosocial topics in such patients would improve the recognition of emotional distress. PMID- 11256059 TI - [Pattern of intervention and patients' satisfaction with community mental health services in Bologna]. AB - OBJECTIVE: To measure satisfaction with Community Mental Health Service (CSM) in patients and to check if it will be associated with some selected demographic and service variables. DESIGN: This study compares data from satisfaction scale administration in patients who attended CSM at least once every two months during 1998 and services that them have benefited from. SETTING: The Community Mental Health Service of Saragozza-Porto District in Bologna. Main outcome measures- Demographical, clinical and service variables taken from CSM informative system and Verona Service Satisfaction Scale (VSSS-32), a multidimensional instrument which measures satisfaction with community-based psychiatric service. RESULTS: Main results (145 subjects) pointed out higher satisfaction for technical and interpersonal skills of staff, information, drugs, help to get economical benefits and domiciliary care. Four issues stood out by comparing patients' satisfaction and their demographic, clinical and services' benefit features: 1) aged patients complained about high drugs costs and of contacts with other seriously ill patients into the waiting room; 2) relative's involvement was judged as insufficiently effective; 3) in young patients with serious mental disease high frequency of visits seemed to correlate with low satisfaction; 4) patients with complex pattern of intervention (including economical benefits) were the less satisfied ones with service's economical support. CONCLUSIONS: Patients' satisfaction differs according to their demographic and clinical features, but the pattern of intervention seems to influence their judgements too, sometimes in a incoherent way. PMID- 11256060 TI - [Needs for care in psychiatric patients: a systematic review. II. Needs for care on individual]. AB - OBJECTIVE: The present paper represents the second part of a larger review on the studies assessing needs for care in mental health field. In the first part, studies assessing needs for services in the general population were reviewed. In this second part studies assessing needs on individual level were reviewed and the role played by the assessment of individual needs in planning mental health care was discussed. METHODS: Studies published in international literature from January 1980 to June 1999 were reviewed. The studies were located through a computerised search of the databases MEDLINE and PsycLit; in addition, the reference lists of the studies located through the computerised search and the content of main international psychiatric journals were manually scanned in order to avoid possible omissions. Studies assessing needs in the general population and in psychiatric patients were reviewed separately. The latter were reviewed on the basis of the diagnostic category (psychotic and non psychotic patients) and setting of care (outpatients and inpatients). Studies on particular categories, such as homeless, children and adolescents, elderly and patients with aggressive behaviour, have been also reviewed. RESULTS: Psychotic patients show a wide range of both clinical and social needs. Patients followed by community-based mental health services show similar number of clinical and social needs, while inpatients (with the exception of acute patients) show more social needs. For both community-based mental health patients and inpatients, social needs are most frequently unmet and psychopharmacological treatments are used more frequently than psychotherapeutic or rehabilitative interventions. Non psychotic patients show lower number of needs than psychotic patients; these needs are both clinical and social, with a higher total number of clinical needs, but a higher number of unmet social needs. CONCLUSIONS: Studies on needs for care provide an interesting perspective on people suffering from mental illness. In the routine clinical practice needs assessment could be a valuable tool for providing specific, individualised and effective mental health care. PMID- 11256062 TI - Monkeying about with transgenics. PMID- 11256061 TI - Two-step regulation of asymmetry. PMID- 11256063 TI - Patterning back to front. PMID- 11256064 TI - Tiled arrays for gene hunting. PMID- 11256065 TI - RNA splicing. Chance findings. PMID- 11256066 TI - Snapshots of metabolic phenotypes. PMID- 11256067 TI - Equus--how it all began. PMID- 11256068 TI - Modifier genes in mice and humans. AB - An emerging theme of studies with spontaneous, engineered and induced mutant mice is that phenotypes often depend on genetic background, implying that genetic modifiers have a role in guiding the functional consequences of genetic variation. Understanding the molecular and cellular basis by which modifier genes exert their influence will provide insights into developmental and physiological pathways that are critical to fundamental biological processes, as well as into novel targets for therapeutic interventions in human diseases. PMID- 11256069 TI - Establishing sexual dimorphism: conservation amidst diversity? AB - The molecular mechanisms that control sexual dimorphism are very different in distantly related animals. Did sex determination arise several times with different regulatory mechanisms, or is it an ancient process with little surviving evidence of ancestral genes? The recent identification of related sexual regulators in different phyla indicates that some aspects of sexual regulation might be ancient. Studies of sex-determining mechanisms are beginning to reveal how sexual dimorphism arises and evolves. PMID- 11256070 TI - Function and evolution of the plant MADS-box gene family. AB - The function of MADS-box genes in flower and fruit development has been uncovered at a rapid pace over the past decade. Evolutionary biologists can now analyse the expression pattern of MADS-box genes during the development of different plant species, and study the homology of body parts and the evolution of body plans. These studies have shown that floral development is conserved among divergent species, and indicate that the basic mechanism of floral patterning might have evolved in an ancient flowering plant. PMID- 11256071 TI - Chromosomal stability and the DNA double-stranded break connection. AB - Genome stability is of primary importance for the survival and proper functioning of all organisms. Double-stranded breaks in DNA are important threats to genome integrity because they can result in chromosomal aberrations that can affect, simultaneously, many genes, and lead to cell malfunctioning and cell death. These detrimental consequences are counteracted by two mechanistically distinct pathways of double-stranded break repair: homologous recombination and non homologous end-joining. Recently, unexpected links between these double-stranded break-repair systems, and several human genome instability and cancer predisposition syndromes, have emerged. Now, interactions between both double stranded break-repair pathways and other cellular processes, such as cell-cycle regulation and replication, are being unveiled. PMID- 11256072 TI - The human Y chromosome, in the light of evolution. AB - Most eukaryotic chromosomes, akin to messy toolboxes, store jumbles of genes with diverse biological uses. The linkage of a gene to a particular chromosome therefore rarely hints strongly at that gene's function. One striking exception to this pattern of gene distribution is the human Y chromosome. Far from being random and diverse, known human Y-chromosome genes show just a few distinct expression profiles. Their relative functional conformity reflects evolutionary factors inherent to sex-specific chromosomes. PMID- 11256073 TI - Scientific perspectives on regulating the safety of genetically modified foods. AB - Regulation is often seen as the dull end of science. The recent storm over the introduction of genetically modified foods and the calls to regulate their consumption have had a negative effect on development of the science. Assuring the safety of genetically modified foods might raise questions where existing scientific data is limited and underline the need for further research. PMID- 11256074 TI - Human genome diversity: what about the other human genome project? AB - Although the Human Genome Project has been successful, the Human Genome Diversity Project, proposed in 1991, has so far failed to thrive. One of the main values in studying the human genome, however, will come from examining its variations and their effects. To do that in a systematic way, an active Human Genome Diversity Project, or something very similar, will ultimately prove vital. Such an effort will confront difficult ethical and political issues; this article reviews those issues and tries to show how they might be overcome. PMID- 11256075 TI - Ethical and legal implications of pharmacogenomics. AB - Pharmacogenomics is the application of genomics technology to the discovery and development of drugs. A greater understanding of the way in which individuals with a particular genotype respond to a drug allows manufacturers to identify population subgroups that will benefit most from a particular drug. The increasing emphasis on pharmacogenomics is likely to raise ethical and legal questions regarding, among other things, the design of research studies, the construction of clinical trials and the pricing of drugs. PMID- 11256076 TI - Multiple roles of EPH receptors and ephrins in neural development. AB - The control of cell movement during development is essential for forming and stabilizing the spatial organization of tissues and cell types. During initial steps of tissue patterning, distinct regional domains or cell types arise at appropriate locations, and the movement of cells is constrained in order to maintain spatial relationships during growth. In other situations, the guidance of migrating cells or neuronal growth cones to specific destinations underlies the establishment or remodeling of a pattern. Eph receptor tyrosine kinases and their ephrin ligands are key players in controlling these cell movements in many tissues and at multiple stages of patterning. PMID- 11256077 TI - Molecular physiology of P2X receptors and ATP signalling at synapses. AB - ATP is found in every cell, where it is a major source of energy. But in the nervous system, ATP also has additional actions, which include its role in fast synaptic transmission and modulation. Here I discuss the 'fast' actions of ATP at synapses, the properties of the receptors that are activated by ATP and the physiology of ATP signalling, with emphasis on its role in pain processing. PMID- 11256078 TI - Potassium leak channels and the KCNK family of two-P-domain subunits. AB - With a bang, a new family of potassium channels has exploded into view. Although KCNK channels were discovered only five years ago, they already outnumber other channel types. KCNK channels are easy to identify because of their unique structure--they possess two preforming domains in each subunit. The new channels function in a most remarkable fashion: they are highly regulated, potassium selective leak channels. Although leak currents are fundamental to the function of nerves and muscles, the molecular basis for this type of conductance had been a mystery. Here we review the discovery of KCNK channels, what has been learned about them and what lies ahead. Even though two-P-domain channels are widespread and essential, they were hidden from sight in plain view--our most basic questions remain to be answered. PMID- 11256079 TI - GLIA: listening and talking to the synapse. AB - Glial cells are emerging from the background to become more prominent in our thinking about integration in the nervous system. Given that glial cells associated with synapses integrate neuronal inputs and can release transmitters that modulate synaptic activity, it is time to rethink our understanding of the wiring diagram of the nervous system. It is no longer appropriate to consider solely neuron-neuron connections; we also need to develop a view of the intricate web of active connections among glial cells, and between glia and neurons. Without such a view, it might be impossible to decode the language of the brain. PMID- 11256080 TI - Computational modelling of visual attention. AB - Five important trends have emerged from recent work on computational models of focal visual attention that emphasize the bottom-up, image-based control of attentional deployment. First, the perceptual saliency of stimuli critically depends on the surrounding context. Second, a unique 'saliency map' that topographically encodes for stimulus conspicuity over the visual scene has proved to be an efficient and plausible bottom-up control strategy. Third, inhibition of return, the process by which the currently attended location is prevented from being attended again, is a crucial element of attentional deployment. Fourth, attention and eye movements tightly interplay, posing computational challenges with respect to the coordinate system used to control attention. And last, scene understanding and object recognition strongly constrain the selection of attended locations. Insights from these five key areas provide a framework for a computational and neurobiological understanding of visual attention. PMID- 11256081 TI - Eph receptors and neural plasticity. AB - Eph receptor tyrosine kinases are largely known for their involvement in brain development but, as some of these receptor tyrosine kinases are also expressed in adults, their possible role in the mature nervous system has begun to be explored. Evidence for the involvement of Eph receptors in synaptic plasticity, learning and memory is only emerging and needs corroboration. However, it is likely that the actions of Eph kinases in the adult brain will attract significant attention and become a fertile research area, as occurred in the case of the neurotrophins. PMID- 11256082 TI - Cognitive neuropsychiatry: towards a scientific psychopathology. AB - Cognitive neuropsychiatry represents a systematic and theoretically driven approach to explain clinical psychopathologies in terms of deficits to normal cognitive mechanisms. A concern with the neural substrates of impaired cognitive mechanisms links cognitive neuropsychiatry to the basic neurosciences. The emergence of cognitive neuropsychiatry in the 1990s illustrates the growing rapprochement between cognitive neuropsychology, clinical medicine and the neurosciences in addressing common questions about disorders of the mind/brain. In reviewing recent applications, we highlight how this hybrid discipline will make a distinctive contribution to the science of psychopathology. PMID- 11256083 TI - [Support intervention in grieving patients]. AB - OBJECTIVES: Principal: to describe the characteristics of grievers in an urban area. SECONDARY: To describe the risk factors for pathological grief, and the distribution of those cases of grief which are resolved and those tending to become chronic. DESIGN: Descriptive study. SETTING: Txantrea Health Centre, Pamplona. PATIENTS: Consecutive sampling of all the family members of people who died between January and July 1999. In 4 basic care units, a grief-monitoring procedure was followed with 62 bereaved persons for 8 months. In the 8 other basic care units, the 21 bereaved people recruited formed a control group. INTERVENTION: Initial interview with risk factors of pathological grief described in the PAPPS. Periodical consultations for help. Final interview with test from the Texas Revised Inventory of Grief Manual to determine the evolution of grief. MEASUREMENTS AND MAIN RESULTS: Over 90% had some risk factor. Considerable numbers were self-aware of difficulty in overcoming earlier grief. Over 50% did not attend even half their appointments, which was related to low income (p < 0.005) and background of depression (p < 0.04). Low income was related to worse evolution of grief (p < 0.01). There were high proportions of prolonged and delayed grief, without any differences found between the intervention and control groups. CONCLUSIONS: There were large numbers of bereaved people with risk factors, frequent failure to adhere to the proposed programme, and a high number of cases of lengthy and delayed grief. PMID- 11256084 TI - [Management of viral dermatologic lesions primary care]. AB - OBJECTIVE: To describe the characteristics of various viral tumoural lesions and their treatment by family doctors. DESIGN: Descriptive, retrospective study. SETTING: Urban health centre. PATIENTS: 225 patients registered at the centre. INTERVENTIONS: The following variables were collected over two years: sex, age, type of lesion, anatomical location and treatments used. MEASUREMENTS AND MAIN RESULTS: 51% of the sample were men, and 49% women. Almost half were between 15 and 34. The most common pathologies were common verrucas (72%), plantar warts (19%) and Molluscum contagiosum (8%). All the plantar warts were located on the lower limbs; condylomas and bowenoid papulosis on the genitals; 35% of the common verrucas on the head and neck (the rest on upper limbs); and 26% of the Molluscum contagiosum on the head and neck too (37% on the thorax and/or abdomen). All the genital lesions were treated medically. Cryotherapy was the most common treatment, used for all plantar warts, 53% of verrucas and 21% of the Molluscum contagiosum. Curettage was used on 80% of the Molluscum and 30% of the verrucas. 11% of the verrucas were sliced off. CONCLUSIONS: There are no conclusive data in the literature about the choice of technique for treating these lesions. We recommend cryotherapy for multiple lesions and in areas where surgery could have functional repercussions. Curettage seems effective for Molluscum contagiosum and where there is resistance to medical treatment. PMID- 11256085 TI - [Validity of new diagnostic criteria for type 2 diabetes mellitus. Impact of its application in a health care area]. AB - OBJECTIVES: To estimate how the number of diabetics known as type 2 is modified by applying the American Diabetes Association (ADA) new diagnostic criteria. To calculate the sensitivity, specificity and predictive values of the ADA diagnostic criteria. DESIGN: Transversal descriptive study. SCOPE OF THE STUDY: 15,451 people belonging to two urban health care centers. 1292 individuals were studied by routine random sampling. SUBJECTS OF THE STUDY: General population between 40 and 75 years of age. METHOD: Review of clinical histories, selecting the basal glycemias performed over the last three years, considering the last one in the event that more than one existed, and performing the necessary analysis if no data existed. A new glycemia measurement was carried out as well as a glucose tolerance test for those values > or = 110-139. Age, sex, and prior diagnosis of diabetes were also recorded. MEASUREMENTS AND RESULTS: The mean age was 56 years, 56.1% were females. Normal glycemias (< 110)--830 individuals (86.2%). The prevalence of diabetes was 10.5% when the WHO criteria were applied and 8.7% when ADA criteria were applied. Sensitivity was 39.29%, specificity was 100%, the positive predictive value was 100% and the negative predictive value was 98.5%. 49 individuals presented an altered basal glycemia (5.5%), 17 of whom (34.7%) were diabetic according to the WHO. CONCLUSIONS: The prevalence of diabetes when the WHO criteria are applied is significantly higher than when ADA criteria are applied (p = 0.000). The basal glycemia value of > or = 126 is less sensitive than the glucose tolerance test. No normoglycemic patient according to the ADA would be diabetic according to the WHO; however the ADA and the WHO classify the non-normoglycemics in different groups. The WHO criteria (scrupulously applied) are the better diagnostic method for diabetes in principle and the glucose tolerance test is a test not to be done away with. PMID- 11256086 TI - [Drug information in the Internet]. PMID- 11256087 TI - [Consensus document of the Spanish Society of Palliative Care (SECPAL) and the Spanish Society of Family and Community Medicine (semFYC). Domiciliary care for the patient with cancer in terminal phase]. PMID- 11256088 TI - [The patient with spinal cord lesions outside the hospital]. PMID- 11256089 TI - [The collapse of the hospital emergency services during the winter]. PMID- 11256090 TI - [The sexuality of university students: its perception and care]. PMID- 11256091 TI - [Coronary interventions and risk of hemorrhage]. PMID- 11256092 TI - [Do patients with cardiopathy stop smoking?]. PMID- 11256093 TI - [Profile of the patient with resistance to antituberculosis treatment in a basic health area]. PMID- 11256094 TI - [Increase of arterial pressure caused by generic atenolol]. PMID- 11256095 TI - [Health and gender in primary care consultations]. PMID- 11256096 TI - [Delay in the diagnosis of cancer]. AB - OBJECTIVE: A reduction in the time used to be able to diagnose an illness influences not only the well being of the patient but also the quality of the medical care. In this work, we have studied the amount of time needed to diagnose cancer. MATERIAL AND METHOD: Descriptive methods have been used: average, median, mode and standard deviation in order to know how long it takes to diagnose and treat cancer. The data have been taken from a hospital register. RESULTS: The longest time elapsed between the periods appears to be the one between the first symptoms and the date the diagnosis is made, which has been calculated to be 96 days on average. The remaining time periods studied showed a time lapse of about 26 days on median. DISCUSSION: The median has been taken as the measurement chosen to assess these time periods, which have been notably reduced in the last decade. CONCLUSIONS: The performance of the primary care doctor is of great importance, as he is normally the one who is faced with the precocious symptoms, as well as with the risk factors that persist in the patients. This doctor is therefore the one who can initiate both the early diagnosis and the medical education of the patients. PMID- 11256098 TI - [Design and validation of a questionnaire in Spanish for measuring the quality of life in postmenopausal women: the MENCAV questionnaire]. AB - OBJECTIVE: To calculate the validity and reliability of a questionnaire to measure the quality of life in post-menopausal women. DESIGN: Cross-sectional observation study. SETTING: Three health districts in the province of Cuenca. PARTICIPANTS: 203 menopausal women between 44 and 60 years of age, chosen through simple randomised sampling. MEASUREMENTS AND MAIN RESULTS: On the basis of the analysis of various scales of health-related quality of life, some for measuring physical disorders, and others for psychological disorders, a questionnaire was designed. This had 85 items structured as a Likert-like additive scale and explored four dimensions (physical health, psychological health, family environment and social support). In the view of a multi-disciplinary group of health professionals, it met criteria of logical validity and validity of contents for the different spheres that are usually affected in the menopause. After the administration of the questionnaire to the women taking part, a process of "purging" items began, in which those items with a reply frequency to one of the five options of below 5% or above 95% were discarded. Also discarded were those items for which the inter-item correlation was below 0.20, with analysis too of the behaviour of the Cronbach's alpha coefficient as each item was eliminated from its scale. Finally, through exploratory factorial analysis techniques (main components), the distribution of the items in dimensions was evaluated. A final questionnaire made up of 37 items in 5 dimensions, which we named physical health, psychological health, family environment, sexual relationships and social support was reached. This questionnaire had an overall Cronbach's alpha value of 0.84. Finally, the correlations existing between the sum of scores of the items on each of the dimensions were analysed. The correlation coefficients showed values between 0.08 and 0.69. CONCLUSIONS: The MENCAV questionnaire is the first in Spanish which assesses quality of life in menopausal women. Experts analysed its logical validity and contents validity and found it had a high degree of internal consistency. PMID- 11256097 TI - [Inventory of experiences in grief (IEG): adaptation to Spanish, reliability and validity]. AB - OBJECTIVE: To adapt to Castilian the Inventory of Experiences of Grief (IEG) of Catherine Sanders et al. (1977) and study its reliability and validity. DESIGN: In two stages: cross-cultural adaptation of a questionnaire and cross-sectional study with control group. SETTING: Primary care teams in Vizcaya. PARTICIPANTS: 147 people bereaved in the period between 3 months and 3 years before the study, and 36 who had lost no direct family member in the previous 5 years. MEASUREMENTS AND MAIN RESULTS: The IEG in American English was translated, back-translated and finally reviewed by Sanders and her colleagues, whose valuation was that the Castilian version was the same as the original. Reliability: the internal consistency of each of the scales of grief on the IEG (Cronbach's alpha) ran from 0.43 to 0.85. Factor validity: the first IEG factor adapted was similar to the original one (despair, somatization, anger, blame, depersonalisation and social isolation). Discriminating validity: all the grief scales on the IEG, except anxiety in face of death, discriminated (p < 0.05) between grieving and not grieving. Validity by hypothesis: the IEG scales showed differences (p < 0.05) between the bereaved according to sex, age, place of death of the spouse and time elapsed since death. Convergent validity: all the IEG grief scales correlated positively (p < 0.05) with the scales in the Texas Revised Inventory of Grief. CONCLUSIONS: The IEG adapted to Castilian is equivalent to the original and has similar reliability and validity. PMID- 11256100 TI - [In search of the Golden Fleece]. PMID- 11256099 TI - [Reactive hypoglycemia and neurocardiogenic syncope. A common origin or just coincidence?]. PMID- 11256101 TI - [DNA microchips: a new tool for the molecular diagnosis of cancer]. PMID- 11256102 TI - Evidence of a linkage between neurocardiogenic dysfunction and reactive hypoglycemia. AB - OBJECTIVE: Reactive hypoglycemia is a common medical problem whose pathophysiology is not completely understood. The objective of this study was to investigate the prevalence of autonomic nervous system abnormalities in patients with reactive hypoglycemia compared with controls. METHODS: Six women, mean age 31 +/- 5 years, with reactive hypoglycemia, and 5 healthy controls women aged 24 +/- 4 years were studied. We investigated the heart rate variability and blood pressure changes after an upright tilt with and without an isoproterenol infusion. A positive result was defined as syncope or presyncope associated with bradycardia, hypotension or both. RESULTS: In response to the orthostatic stress alone or in conjunction with a 4 micrograms isoproterenol infusion, 5 of 6 patients had a positive test as did one of the five control subjects. Patients had a baseline hyperadrenergic tone, with a sympathetic to parasympathetic ratio of 2.3 +/- 0.8 under basal conditions and 10.1 +/- 4.1 during the isoproterenol infusion, compared to 0.7 +/- 0.3 (p = 0.06) and 0.5 +/- 0.1 (p < 0.01) respectively, in the control group. CONCLUSION: Patients with reactive hypoglycemia may be at the extreme end of a spectrum of normal biologic variability, they may have an hyperadrenergic tone and, after a provocative stimulus, sympathetic nerve firing and or synaptic release of NE may not be sufficient to maintain an adequate vascular tone. Alternatively, the vascular response to NE may be impaired. An excessive and paradoxic vasovagal or parasympathetic response was not observed. PMID- 11256103 TI - Quantification and characterization of protein loss in continuous ambulatory peritoneal dialysis. AB - BACKGROUND/AIMS: Dialysate protein loss is involved in the etiology of hypoalbuminemia and malnutrition on CAPD. There is no information regarding the peritoneal transport of proteins in neither Mexican nor LatinAmerican CAPD patients. Therefore, the aim of this study was to quantify and characterize the peritoneal membrane transport of albumin (Alb), IgG, IgA, and IgM in CAPD patients. In addition, factors associated to protein losses were investigated. METHODS: Thirty-seven CAPD patients were randomly selected and subjected to a standard peritoneal equilibration test (PET). Alb, IgG, IgA, and IgM were measured during the PET in both serum and dialysate. RESULTS: Dialysate IgM was not detected with the employed nephelometry method. A significant continuous and gradual increasing pattern in Alb, IgG and IgA losses was observed throughout the PET. During the PET, patients with the fastest transport type displayed the lowest serum concentration of Alb (but not Igs) and the greatest dialysate loss of all the assayed proteins. The strongest predictor for Alb, IgG, and IgA losses was peritoneal transport rate in both the univariate and multivariate analyses. CONCLUSIONS: Peritoneal Alb, IgG, and IgA losses in Mexican CAPD patients are mainly dependent on peritoneal transport rate and dialysate dwell-time. PMID- 11256104 TI - [Clinical characteristics and course of severe hypercalcemia caused by primary hyperparathyroidism in surgically treated patients]. AB - AIM: To describe presentation, diagnosis, management and outcome of severe hypercalcemia due to primary hyperparathyroidism in a series of patients. METHODS: Clinical characteristics, presentation, diagnosis, acute preoperative medical management, surgical findings and strategy, short outcome and complications of a cohort of 21 patients with primary hyperparathyroidism (HPT) and severe hypercalcemia (serum calcium > or = 14 mg/dL) were analyzed. This group was selected from a total of 118 patients who underwent surgery for HPT in the time period. RESULTS: Mean age was 47 +/- 17 years-old and the male:female ratio was 4:17. A total of 95% of patients presented one or more symptoms related to hypercalcemia while 62% had an abnormal EKG and 76% also had radiological abnormalities. All patients received intense hydration often associated to diuretics. Uniglandular disease was found in 13 patients, multiglandular involvement was identified in 4 and parathyroid carcinoma was documented in other 4. Normalization of the calcemia was achieved in all patients with benign disease. CONCLUSIONS: Severe hypercalcemia was relatively frequent in our patients with HPT. Most patients were symptomatic and presented radiological or cardiac abnormalities. Surgical normalization of the calcemia was achieved in all patients with benign disease. PMID- 11256105 TI - [Cryptosporidium parvum infection in malnourished and non malnourished children without diarrhea in a Mexican rural population]. AB - Cryptosporidium parvum is associated with diarrheic disease and mainly affects children and immunocompromised hosts. In most of the cases, cryptosporidiosis infection is asymptomatic in immunocompetent subjects. The objectives of the study were to determine the frequency of asymptomatic infection caused by the parasite in children with and without malnutrition and to determine the risk factors associated to infection. METHODS: Children from one to fifteen years old without diarrhea were included, somatometry were performed. The socioeconomic and sanitary conditions were investigated for each family and community. The Faust method and Kinyoun stain were employed identify parasites and Cryptosporidium parvum in feces. Odds ratio (OR), 95% confidence intervals (75% CI), chi 2 Mantel Haenszel, Fisher exact test and chi 2 trends were calculated. RESULTS: One hundred thirty two children were included. In 10/132 (7.5%) cysts of Cryptosporidium were found, 7/71 in children with malnutrition (9.8%) and 3/61 without malnutrition (4.9%) p = 0.23. 69.7% of the children had parasitosis. According to the presence of C. parvum in feces, the different factors calculated were: Diarrhea in family OR = 5.82 (95%IC 0.86-39.18), not hand washing OR = 5.08 (95%IC 0.62-110.49), age less than 5 years old OR = 4.90 (95%IC 0.60-106.9), drinking non-potable water OR = 3.34 (95%IC 0.40-73.01) and malnutrition 2.11 (95%IC 0.46-10.89). Association was found between the number of people in the same house and the risk of infection (p = 0.005). The presence of diarrhea in the family (OR = 4.15, 95%IC 0.47-36.91) and drinking non-potable water (OR = 4.19, 95%IC 0.48-36.32) were the significant factors in the regression logistic model. CONCLUSIONS: The frequency of Cryptosporidium infection were 7.5%. Diarrhea in the family, overcrowding and drinking non-potable water were associated with C. parvum infection, malnutrition was not a significant risk factor. PMID- 11256106 TI - [Fulminant sepsis caused by Vibrio vulnificus. A case series]. AB - BACKGROUND: Vibrio vulnificus is a marine bacteria associated with the ingestion of raw shellfish or contact with seawater. It can produce wound infection, diarrhea and sepsis. The main risk factor for infection is the presence of chronic liver disease. Prior studies have shown mortality from 40% to 63%. OBJECTIVE: Report of 8 cases of disseminated infection with V. vulnificus causing fulminant sepsis. DESIGN: Series of cases. METHODS: We reviewed the database of the laboratory of clinical microbiology from 1990 to 1999. A computer-based review of the worldwide medical literature was also accomplished. RESULTS: There were 8 cases of V. vulnificus infection. All patients had chronic liver disease, 3 also had diabetes mellitus and 1 received immunosuppressive agents. Five patients were known to have ingested raw shellfish. The mean duration of illness before death was 4 days. All patients presented with sepsis, seven had cutaneous lesions. Five patients received early antimicrobial treatment during the first 24 hours and all of them in the first 48 hours. Regardless of susceptibility to the antimicrobial agents used, the mortality was of 87.5%. Disk-diffusion test showed 100% susceptibility to imipenem, ceftazidime and tetracycline; 83% to cefepime, ticarcillin and cotrimoxazole and 50% to quinolones. CONCLUSION: The V. vulnificus infection appears in patients with chronic liver disease and it is associated with high mortality. This infection has to be suspected in high-risk patients who have eaten raw shellfish and therapy must be initiated as soon as possible. PMID- 11256107 TI - [Sodium caseinate induces differentiation of 32D pluripotential hematopoietic cells]. AB - AIM: To determine the role of sodium caseinate (CasNa) in the modulation of hemopoiesis. MATERIALS AND METHODS: 32D cells, a murine hemopoietic multipotential cell line dependent on interleukin-3 (IL-3) for proliferation and survival, were used. These cells were cultured with 0.5 ng/mL of IL-3, together with different concentrations of CasNa. We evaluated: proliferation (direct counting under the microscope and use of thymidine 3H), morphological differentiation (giemsa staining), cytochemistry (specific staining for monocytes and granulocytes), and function (presence of Fc receptors and reduction of nitro blue tetrazolium). In addition, we determined cell viability through trypan blue exclusion and apoptosis using the TUNEL assay in situ. RESULTS: We showed that CasNa induced a decrease in cell proliferation, which is dose dependent, and is neither a result of a diminished cell viability, nor due to an increase in cell death through apoptosis. In addition, CasNa induces cell differentiation towards the monocytic lineage. CONCLUSIONS: CasNa has the capacity to differentiate 32D cells towards the monocytic lineage, and, importantly, has a potent differentiating activity on 32D cells being able to promote differentiation in a shorter time than the well known factors G-CSF and GM-CSF. PMID- 11256108 TI - [Transfection of TF-1 cell line with human bcl-2 proto-oncogene provides short term survival in absence of GM-CSF without changing the phenotype]. AB - Apoptosis is a process genetically controlled. The produce of the bcl-2 gene, bcl 2, is an anti apoptotic protein that is linked to the external membrane of the mitochondria. OBJECTIVE: To explore the possibility that bcl-2 transfection could change phenotype, response to mitogenic factor, and cell morphology on the TF-1 parental cell line and the bcl-2 transfectant TB-1 or TF-1neo. METHODS: We look at the expression of CD13, CD34 and c-Kit surface markers by flow cytometry. We have measured cell proliferation in response to GM-CSF and cell survival after GM CSF withdrawal by the MTT assay on the same cell lines. Apoptosis was evaluated by the apoptotic membrane blebbing set up at different times after serum and survival factor removal or tolerance to cytotoxic compounds from Justicia spicigera. RESULTS: According with our results, ectopic expression of the bcl-2 gene prevented apoptosis without changes in morphology or phenotype in the absence of GM-CSF and serum or the presence of the extract from Justicia spicigera. Consisting with the Bcl-2 function, we found that Bcl-2 did not change response to GM-CSF. Serum deprivation or GM-CSF withdrawal induces cell death at 36 hours in TF-1 and TF-1neo cells, whereas TB-1 cells undergo apoptotic membrane blebbing after 96 hours under the same conditions. CONCLUSIONS: Taken together, our data indicate that Bcl-2 is a short term anti apoptotic protein in TB-1 cell line, that does not affect response to GM-CSF neither CD13, CD34 nor c-Kit antigen expression. PMID- 11256109 TI - [A model for evaluating internal accuracy]. AB - OBJECTIVE: To provide an evaluation model of internal accuracy illustrating its use in a quality control program of clinical chemistry. METHODS: The model uses data transformed to percentage of assigned value (%AV) that allows the pooling of different controls. Precision is evaluated by the mean of the intracontrol coefficients of variation, and accuracy by the mean of means of the %VAs and its standard deviation. The model was used in 17,280 measurements of 23 analytes assayed in seven pairs of controls (medium and high concentration) during four years in two automated clinical chemistry analyzers. RESULTS: The model established inaccuracy of 5% to 25% in seven analytes, five of them enzymes. It also detected interanalyzer differences in accuracy of five analytes (22%) and an erroneous assigned value in 19 of 304 controls (6%). CONCLUSIONS: The model showed its capacity to dissect the sources of variation of accuracy. There was inaccuracy in 30% of the analytes but further studies are needed to validate these inaccuracies with controls of other sources. The model can be useful for control programs of any quantitative assay system. A summary description of the model's operation is given. PMID- 11256111 TI - [Surgical treatment of primary hyperparathyroidism]. PMID- 11256110 TI - [Interferons in neurology]. AB - Interferon (INF) plays an important role in the immune response against viral infections. INF is part of the big cytokines family. It has also shown modulating activity on numerous components of the immune system; it inhibits cellular division, thus counteracting proliferation of cancerogenic cells. It has also the ability to reduce complications in multiple sclerosis by immunologic mechanism involving. Th2 responses. Recently, the utility of INF in neurological cases has been explored. Good results have been observed using recombinant INF beta-1a and glucosylated INF beta-1 in patients with active multiple sclerosis relapsing remitting type (MS RR), in whom decreases the incidence and complications and possibly halts the progression of the disease. This manuscript will reviews the biologic activity of INF, the use, side effects and indications for using different INF's in neurological diseases specially in multiple sclerosis, particularly INF beta which are broadly accepted for clinical use in this pathology. We mention the Expanded Disability Status Scale (EDSS) to evaluate clinical involvement and recuperation after management. PMID- 11256112 TI - [Progesterone receptor isoforms: function and regulation]. AB - Progesterone participates in the regulation of several physiological processes in mammals. The biological response to progesterone is mediated by two forms of the progesterone receptor (PR) denominated PR-A and PR-B. The difference between them is that 164 amino acids of N-terminal of PR-B are absent in PR-A. Both PR isoforms are derived from a single gene but are generated from either alternative transcriptional or translational start sites, and are regulated by different estrogen-induced promoters. PR-B acts as a transcriptional activator in different cellular contexts whereas PR-A functions as a strong inhibitor of transcriptional activity. PR isoforms expression and function vary among target tissues such as the uterus, the mammary gland and the brain. The knowledge of the molecular mechanisms involved in the regulation of expression and function of PR isoforms will contribute to the understanding of fundamental biological processes such as sexual behavior and reproduction, and it will open the possibility of alternative therapies in fertility control as well as in the treatment of breast, endometrial and cerebral tumors. PMID- 11256113 TI - [Placental vitamin D: synthesis, regulation, and clinical implications]. PMID- 11256114 TI - [Death in medical practice. Excluded subjectivity]. AB - Medical practice when facing death is reviewed in this article. It is considered the result of two circumstances. First, the development of medical technology which has been an important support to physicians, but has led them to give less importance on patient's subjectivity. Second, changes on society' attitudes toward death which over time have led to its present negation. Nowadays, doctors do not respond to the emotional needs of their patients at the end of life. Medical students should receive education to be prepared to stay close to their patients when a cure is no longer possible. PMID- 11256116 TI - Human resources. Getting even. AB - A trust where a third of the workforce is a member of ethnic minority groups has set up a programme to enhance career advancement for this group. The programme aims to celebrate the achievements of ethnic minority staff and give them greater visibility within the organisation. Mentoring programmes and learning sets have been established with the support of senior. PMID- 11256115 TI - Litigation authority. For our eyes only. AB - The NHS Litigation Authority has a great deal of information on individual doctors that should be made available to trusts in order to ensure patient safety. The authority is so focused on managing clinical negligence cases that it neglects its wider mission to serve the NHS. The authority should be reformed to make it more accountable. PMID- 11256117 TI - Public health. A tale of two counties. AB - The development of primary care trusts requires health authority public health departments to work in new ways. Reviews of the public health function in two counties found widely varying views. A common understanding of organisations' responsibilities is crucial when developing public health in primary care. Public health networks can play a key role. Significant investment in training is required. PMID- 11256118 TI - Data briefing. Coronary heart disease. PMID- 11256119 TI - Nurse returners. Many happy returns. PMID- 11256120 TI - NVQs (national vocational qualifications)/training accounts. The bigger picture. PMID- 11256121 TI - MBAs. Worth a second lucre. PMID- 11256122 TI - IT training. Fine delivery. PMID- 11256123 TI - Human resources strategy. A working arrangement. AB - Now that the NHS plan's agenda is being turned into action, this series of special reports aims to assess progress in key areas. This report looks at training, a crucial aspect of the government's resolve to ensure NHS staff are treated better. A leadership academy, individual learning accounts for staff and better courses for nurse returners are some of the initiative promised in the plan. The report examines progress in these areas, as well as courses and schemes to help managers make the best of their careers. PMID- 11256124 TI - Leadership Centre for Health. One thing leads to another. Interview by Lynn Eaton. PMID- 11256125 TI - Leadership initiatives. Coach outings. PMID- 11256126 TI - [Microbiological contamination of intraoperatively collected erythrocyte concentrate in mechanical autotransfusion in tumor surgery]. AB - Intraoperative autotransfusion is an effective method of the autologous haemotherapy. Gamma irradiation of the salvaged erythrocyte concentrates leads to inactivation of malignant cells and allows retransfusion in oncologic surgery. The risk of microbiological contamination of the autologous blood product has not been systematically studied in this field of surgery. During the evaluation of intraoperative autotransfusion in oncologic surgery at the University Hospital of Leipzig, the salvaged blood of 46 patients who underwent tumour surgery (urology, gynaecology and neurosurgery) was processed to a washed erythrocyte concentrate with a Cell Saver 5 or a Haemolite 2 (Haemonetics). Specimens for microbiological studies were taken from the area of surgery, the reservoir bottle of the autotransfusion system and the salvaged erythrocyte concentrates. The investigation with plate cultures and the identification of microorganisms were performed according to standard procedures of microbiological diagnosis. The ethics committee of the University Hospital of Leipzig approved the study protocol. We found a microbiological contamination in 10 of 46 salvaged erythrocyte concentrates (22%), above all the growth of saprophytes (Streptococcus, Staphylococcus and Neisseria species). We also identified bacteria which have their habitats in the intestinal tract (E. coli and Enterococcus). The highest rate of microbial contaminated erythrocyte concentrates was found in urologic oncosurgery (37%). In neurosurgery (laminectomy and craniotomy including tumour extirpation), there was no bacterial growth in the salvaged erythrocyte concentrate. Intraoperative autotransfusion should be avoided during surgery touching the urinary tract, e.g. prostatectomy and cystectomy. In neurosurgery, it seems to be safe as far as microbiological contamination is concerned. To assess the real risk of microbiological contamination of the salvaged erythrocytes, more investigations are necessary to determine not only the extent and nature of contamination but also the clinical effects. PMID- 11256127 TI - The cuffed oropharyngeal airway in resuscitation. AB - The cuffed oropharyngeal airway (COPA, Mallinckrodt) may prove a useful airway adjunct in resuscitation by non-anaesthetists. Many hospital workers are familiar with the standard oropharyngeal airway but find it difficult to use with a face mask to provide effective positive pressure ventilation of the lungs. However, most inexperienced hospital basic life support providers can use the COPA effectively with minimal instruction. Recommended COPA size and cuff volumes for manual ventilation in adult males and females are respectively size 11 and 10 with up to 60 ml and 50 ml in each cuff. Simple airway manoeuvres taught in basic life support (lifting the chin, tilting back the head) also apply to use of the COPA. A leak around the cuff is common but does not usually impair the ability to ventilate the lungs. Future developments may include more widespread evaluation of the COPA in resuscitation and direct comparisons with other devices. PMID- 11256128 TI - [Hiatal hernia and risk of aspiration in anesthesia induction]. AB - Papers recently published in the literature have questioned whether residual gastric fluid volume at the time of induction is the most important risk factor for pulmonary aspiration. To estimate the risk, more factors than the gastric fluid volume have to be considered. Concomitant diseases such as the hiatal hernia must be considered. Pulmonary aspiration during induction of anaesthesia seems to be caused by a multifactorial process, which consists of gastric fluid volume, anaesthetic technique and concomitant disease. A fifty-year-old man was scheduled for elective cholecystectom. During induction, the patient surprisingly regurgitated and aspirated gastric fluid. Postoperatively, an additional barium swallowing x-ray examination showed a hiatal hernia. This case report shows that patients who report heartburn in their case history should be prophylactically treated as endangered by aspiration, even when they are considered to have an empty stomach. PMID- 11256129 TI - [Release of pro- and anti-inflammatory cytokines during different anesthesia procedures]. AB - The modulation of immune functions, induced by trauma, surgical interventions and anaesthesia, is thought to play a crucial role in the development of post traumatical or postoperative disorders. The balance of pro- and anti-inflammatory cytokines was shown to affect the outcome of the patients. This work studied the effects of different anaesthesiological procedures--total intravenous anaesthesia using Propofol/Sufentanil (TIVA) versus balanced inhalational anaesthesia using Trapanal/Sevoflurane (BIA) in patients with elective lumbal discectomia--on the secretion of various cytokines and their correlation to endocrine stress response. The concentrations of the pro-inflammatory cytokines IL-2, IL-6, IL-12 and IFN-gamma and their soluble receptor molecules as well as the concentrations of the anti-inflammatory cytokines IL-10, IL-1RA and TGF-beta were determined in plasma samples obtained pre-, intra- and postoperatively. Additionally, the plasma concentrations of the stress-related hormones cortisol, epinephrine and norepinephrine were measured. Changes in the cytokine profile were observed immediately after induction of anaesthesia. Significant differences were found particularly in IL-6 production as well as in the release of the soluble IL-2R alpha and the IL-1 receptor antagonist (IL-1RA). Whereas under BIA, the concentrations of IL-6 were found to be significantly elevated during the course of the study, the release of the soluble IL-2R alpha and the production of IL-1RA were reduced in this patient group in comparison to the TIVA group. The increase of the postoperative concentrations of cortisol, epinephrine and norepinephrine under BIA indicated enhanced activation of the hypothalamo-pituitary-adrenal axis and the sympathetic system. Thus, with respect to limitation of surgery associated stress, total intravenous anaesthesia seems to have a favourable effect. Moreover, induction of the release of anti-inflammatory mediators under TIVA might contribute to the prevention of excessive postoperative inflammation. Taken together, these data suggest that the anaesthesiological management may have considerable influence on the postoperative inflammatory process. This might be of particular relevance for surgical interventions in patients after injuries, infections or malignant diseases which are known to be associated with immune dysfunction. PMID- 11256130 TI - [2001: already!]. PMID- 11256131 TI - [Neuroborreliosis]. AB - Lyme disease, or borreliosis, is an endemic affection in Belgium. It is transmitted by a spirochete, Borrelia burgdorferi. The particularity of the infecting genomic group, Borrelia garinii, implies that half of the reported cases of Lyme disease in our country have neurologic manifestations. Due to the marked clinical heterogeneity and the difficult serologic diagnosis, neuroborreliosis is often part of the differential diagnosis in neurology. The antibiotic treatment is necessary because it decreases the risk of more advanced stages of the disease. We hope that a vaccination will soon be available in Belgium. PMID- 11256132 TI - [Is it reasonable to prescribe growth hormones in the elderly?]. AB - During normal aging, there is a +/- 60% decrease of the endogenous GH secretion ("somatopause"). However it is safe to prescribe GH therapy only to those people who show a clear cut decrease of GH release as evidenced by low integrated 24 hr secretion and/or low plasma IGF-1. In our view, a complete clinical check up must also show, on the one hand, an abnormal decrease of the optimal quality of life and, on the other hand a willing to maintain a reasonable intellectual and physical activity in the absence of major clinical and biological abnormalities. The benefits are likely to be an increase of muscular strength and of exercise tolerance, a decrease of trabecular osteopenia, a decrease of abdominal obesity, an increase of immunocompetence and a general improvement of the "quality of life". A "pharmacological" prescription is contra-indicated whereas very low dose regimen could induce, through feed back mechanism, a putative decrease of endogenous GH-RH (and GHRPs?) function whose deleterious psychoneuroendocrine effects remain to be demonstrated. PMID- 11256133 TI - [Prevalence and treatment of Helicobacter pylori in gastro-duodenal ulcers. An experience in Liege]. AB - Between April 1998 and July 1999, we prospectively investigated 152 patients with gastric or duodenal ulcer and we observed concomitant H. pylori infection in 72.8% and 78.5% respectively. We proposed to the GPs of these patients the recommended triple therapy (omeprazole 20 mg, amoxicillin 1 g and clarithromycin 500 mg (OAC) twice daily for 7 or 10 days). H. pylori eradication was tested using the C13-urea breath test. Our results showed a modest overall eradication rate of about 70%. We have to persuade the patients and the GPs of the benefit of antibiotics and of the importance of the correct dosages. We have to continue to follow the resistance against antibiotics. PMID- 11256134 TI - [Image of the month. Cutaneous Old World leishmaniasis]. PMID- 11256135 TI - [A day care center for management of early stage Alzheimer's disease]. AB - Alzheimer's disease represents an important problem for our societies, both in social and economical dimensions. Built in July 1997, the day care center of Liege aims at practicing strategic cognitive interventions in early Alzheimer patients or when pathological memory impairments occur. By means of an interdisciplinary work (by neuropsychologists and occupational therapists), the ultimate goal of the interventions is to facilitate the patient's autonomy in activities of daily living. Because of the variability of clinical symptoms of the disease, the goal of these cognitive interventions is to optimize the performance of each patient by making the most of its preserved abilities. Moreover, another goal of the day care center is also to support the caregivers, by giving them advices in order to adequately face with difficulties presented by the patient. PMID- 11256136 TI - [Dirty nails and mycotoxins]. AB - Some mycotoxins are among the most toxic natural products. They are metabolites produced by food-borne and feed-borne moulds. Dirty and soiled nails, and contaminated or infected onyxis by these specific moulds can be the origin of oral contamination and mycotoxicosis. Hence, nail cleaning, correct hygiene and treatment of mould onychomycosis have a potentially great impact on health. PMID- 11256137 TI - [How I treat...persistent atrial fibrillation, by internal cardioversion, in a patient with exreme obesity]. AB - An ethylic hypertensive patient with a BMI of 51.4 developed persistent atrial fibrillation with high ventricular rates. External electrical cardioversion was attempted, but failed in spite of high energy shocks (350 joules). Sinus rhythm was restored by internal cardioversion (12 joules). The value and indications of the techniques are briefly discussed. PMID- 11256138 TI - [Medical responsibility and legal malpractice]. AB - The principle that obliges anyone to answer for his acts, is one of the foundations of our law and ethic systems; so the fault committed by a doctor is punished, penally speaking, by the articles 418 and next of the penal code. These articles specify that a doctor's charge of unintentional blows and wounds or of unintentional homicide rests on the conjunction of three elements: a fault which can be defined as any act which wouldn't have been carried out by any other doctor of the same speciality who is normally cautious, competent and diligent and placed in identical circumstances, a harm: either the patient's death (homicide) or blows and wounds which can be caused by simple omission, an obvious relationship of cause and effect between the fault and the harm. The absence of a practioner's recording of his free and conscious patient's consent is considered similar to a fault by the doctrine. The same doctrine asserts that going beyond the patient's refusal to receive the suggested cares is considered as the same harm as intentional blows and wounds? PMID- 11256139 TI - [How I investigate...a myopathy with a muscle biopsy]. AB - Muscle biopsy is a valuable tool in the diagnosis of many neuromuscular disorders. It is an invasive investigation that will be considered after careful clinical examination, supplemented by biological and electrophysiological studies. The biopsy procedure and the subsequent processing of specimen in the laboratory significantly influence the information provided by the microscopic examination of muscle tissue. They are reviewed in details in this article. Close collaboration between clinicians and neuropathologists in the setting of an optimized procedure will further increase the diagnostic yield of muscle biopsy. PMID- 11256140 TI - [Pharma-clinics. The drug of the month. Celecoxib (Celebrex)]. AB - Celecoxib (Celebrex, Pharmacia) is a potent and selective inhibitor of the COX-2 isoform of cyclooxygenase which is used as nonsteroidal anti-inflammatory drug (NSAID). Its current indications are osteoarthritis and rheumatoid arthritis. The usual recommended daily dosage of celecoxib is 200 mg (in one or two intakes per day), to be increased up to 400 mg (two intakes per day) if necessary. Its clinical efficacy seems to be similar to that of other NSAIDs. However, its safety profile, especially gastro-intestinal tolerance and perhaps renal safety, is much better because of the COX-2 selectivity. PMID- 11256142 TI - [Clinical case of the month. Acute methanol intoxication]. AB - We report a fatal methanol intoxication and discuss its physiopathology, diagnosis and treatment. PMID- 11256141 TI - [Clinical study of the month. After the storm over central anorectic agents, the "STORM" study of sibutramine]. AB - The results of the "Sibutramine Trial of Obesity Reduction and Maintenance" (STORM) published in the last issue of December 2000 of the Lancet are summarized. This clinical trial (open label for the first 6 months and double blind versus placebo for a further 18 months) demonstrates the positive effect of sibutramine, a new anorectic drug, on long-term weight maintenance after weight loss in obese subjects. The benefit of sibutramine on the cardiovascular risk profile of the obese patient is more particularly discussed. PMID- 11256143 TI - [Pathophysiology of restless-legs syndrome. Review of current research]. AB - The primary pathomechanism of restless legs syndrome is unclear. Neurophysiological, pharmacological, and imaging studies have demonstrated a complex interaction between central and peripheral structures. The degree of integration of central and peripheral mechanisms is not known. Altered excitability at a spinal level modulated by supraspinal rhythms of the brainstem possibly plays an important role. There is no anatomical structural lesion and it is more likely that circadian disturbances of dopaminergic and/or opioidergic neurotransmission are involved. PMID- 11256144 TI - [Treatment of relapsing-remitting multiple sclerosis with recombinant interferon beta preparations]. AB - Three recombinant interferon-beta preparations, Betaferon (IFN beta-1b) and Avonex and Rebif (INF beta-1a) have been approved for the therapy of relapsing remitting multiple sclerosis (MS). The results from pharmacokinetic and pharmacodynamic studies with different interferon-beta preparations reflect only in part the mechanisms of action of this substance, because standard assays primarily measure antiviral bioactivity. These studies are often overinterpreted in relation to the therapy of MS. In this article, we critically evaluate the results of pharmacologic studies with respect to the dose-dependent efficacy of different interferon-beta preparations. PMID- 11256145 TI - [New insights in pathogenesis and therapy of sporadic inclusion body myositis (s IBM)]. AB - Sporadic inclusion body myositis (s-IBM) is a chronic progressive inflammatory myopathy which occurs preferentially in older patients. Histologic hallmarks are rimmed vacuoles and eosinophilic cytoplasmatic inclusions. The etiology is still unknown, but different pathogenetic mechanisms such as slow virus infection, autoimmunopathogenesis, myonuclear alterations, and mitochondrial defects have been implicated. A relation to neurodegenerative disorders and prion diseases has also been suggested. There is a poor response if any to immunosuppressive therapy. Stabilization of disease progression was shown only by intravenous immunoglobulin (IVIG) therapy. Future findings in the field of s-IBM pathogenesis may result in better therapeutic options. PMID- 11256146 TI - [Familial mitochondrial chronic progressive external ophthalmoplegia. Five families with differing genetics]. AB - Chronic progressive external ophthalmoplegia (CPEO) is considered the most frequent form of mitochondrial encephalomyopathies. Most cases occur sporadically. We investigated 18 consecutive patients with CPEO. Thirteen cases were sporadic and five cases were familial. In one family with maternal inheritance the mitochondrial point mutation A3243G was identified. In index patients of three other families multiple deletions of mitochondrial DNA were found. One of these families showed autosomal recessive inheritance. In the two other pedigrees a definitive determination of the mode of inheritance was impossible. The fifth family revealed autosomal dominant or maternal inheritance. In their index patient no alteration of mitochondrial DNA could be identified (including sequencing of hot spots for mitochondrial mutations). CONCLUSIONS: CPEO was familial in 28% of our patients. There are three different modes of inheritance: (i) maternal transmission associated with mitochondrial point mutations as it is known for other mitochondrial disorders, (ii) autosomal recessive, and (iii) autosomal dominant inheritance. In contrast to sporadic cases with single mitochondrial deletions autosomal inheritance can be associated with multiple deletions of mitochondrial DNA. They are due to so far unknown nuclear mutations. PMID- 11256148 TI - [Primary central nervous system lymphoma as a neurological manifestation of AIDS stage]. AB - In patients infected with human immunodeficiency virus (HIV), the risk of developing non-Hodgkin's lymphoma is over 100 times greater than with noninfected persons. Primary central nervous system lymphoma as a complication of the acquired immunodeficiency syndrome (AIDS) occurs in up to 2.4% of all cases and is strongly associated with the Epstein-Barr virus. The prognosis is very poor, with a mean survival time of 21 to 27 days without therapy and up to 119 days with radiation therapy. We describe the course of seven AIDS patients with histologically proven primary central nervous system lymphoma and present a review of clinical symptoms, diagnosis, and therapy. The main criteria for differential diagnosis from other secondary neuromanifestations such as cerebral toxoplasmosis, progressive multifocal leukoencephalopathy, abscesses, and infarctions are described. PMID- 11256147 TI - [Diagnostic imaging in refractory temporal lobe epilepsy. A comparison of MR volumetry and multivoxel-MR-spectroscopy for assessment of postoperative prognosis]. AB - While the importance of magnetic resonance (MR) spectroscopy, volumetry, and T2 relaxometry for preoperative localization has already been verified, the question arises as to what extent the individual techniques are useful for assessing postoperative prognosis with respect to seizures and neuropsychological outcome. In a prospective comparative study, 26 patients were examined preoperatively with a 1.5 T whole-body scanner. The MR spectroscopy was taken by high resolution 1 H proton spectroscopy, the volumetry with MP rage technique. The postoperative outcome was laid down using Engel's classification. Our results show that the metabolic changes can be divided into three groups using MR spectroscopy: unilateral, slightly bilateral, and severely bilateral to contralateral. In case of bilateral changes, the severity of metabolic changes in the nonoperated, contralateral side was decisive for postoperative outcome. The results from volumetry did not correlate with postoperative outcome. PMID- 11256149 TI - [Polyradiculopathy and ataxia: clinical manifestation of late recurrence of pilocytic astrocytoma with cerebral and spinal dissemination]. AB - A 14-year-old girl underwent surgery for cerebral pilocytic astrocytoma. Eighteen years later she developed acute ataxia and a polyneuropathic syndrome. A recurrence with cerebral and spinal leptomeningeal dissemination of the tumor was discovered. Treatment of these cases is always difficult, and no standard therapy scheme has been established so far. PMID- 11256150 TI - [Autochthonous cases of tick-borne encephalitis in Rhineland-Palatinate]. AB - Tick-borne encephalitis (TBE) is a neurotrophic viral disease which is endemic to certain regions. Such areas in Germany include Bavaria, Baden-Wurttemberg, and the Odenwald region in Hessen. So far, it has not been endemic to Rhineland Palatinate. There, only two single cases of TBE occurred in the years 1992 and 1997, near the town of Idar-Oberstein. We report two new cases of TBE which appeared in 1999 and two current cases from the Idar-Oberstein region which have been verified clinically and serologically. At admission, the patients suffered from headache, muscle pains, and high fever, in one case meningitis was suspected. In all four patients, serology for borrelia was negative in serum and CSF. The described cases indicate that it is possible to acquire TBE in Rhineland Palatinate, although only two cases have been reported in this area over the previous 10 years. Particularly in regions with a low incidence of TBE, the disease should be taken into consideration as a differential diagnosis. Studies of tick populations in regions with a low incidence can help in evaluating the benefit of possible vaccine recommendations by local public health authorities. PMID- 11256152 TI - [Austrian Society for Neurology]. PMID- 11256151 TI - [Escalating immunomodulatory therapy of multiple sclerosis. 1st supplement: December 2000]. AB - New clinical studies in multiple sclerosis provided data on the treatment of clinical isolated syndromes and secondary progressive forms which may have important implications for the optimal care of MS patients. The MSTKG critically evaluated the available data again and provides evidence-based recommendations for the application of immunoprophylactic therapies. Initiation of treatment after the first relapse may be indicated if there is clear evidence from MRI for subclinical dissemination of disease. Recent trials indicate that efficacy of therapy with IFN--is more likely with superimposed bouts or other indicators of inflammatory disease activity than without them in secondary progressive MS. If immunoprophylactic treatment is initiated with a provisional diagnosis of MS, confirmation of MS is essential. In long-term treated patients secondary treatment failure should be identified by follow-up examinations and other treatment options discussed. PMID- 11256153 TI - ["Sleep attacks" in Parkinson patients]. PMID- 11256154 TI - [Overestimation of the number of multiple sclerosis patients in Germany]. PMID- 11256155 TI - [Alfred Hauptmann (1881-1948). Comments on cover portrait]. PMID- 11256156 TI - [Interferon therapy of multiple sclerosis. A question of the correct dose?]. PMID- 11256157 TI - [Gabapentin therapy for pain]. AB - Gabapentin, which has been approved for add-on therapy of focal seizures, is increasingly used for treatment of neuropathic pain. Its analgesic effect is supposed to be due to reduction of glutamatergic transmission, improvement of GABAergic transmission and to binding to voltage-dependent calcium channels. Experimental studies demonstrated an ameliorating effect of gabapentin on neuropathic pain. Placebo-controlled studies revealed an efficacy of gabapentin against pain in diabetic neuropathy and postherpetic neuralgia and in prophylaxis of migraine. Case reports show an analgesic effect of gabapentin in trigeminus neuralgia and in reflex sympathetic dystrophy. The main adverse events are dizziness, ataxia and somnolence. Controlled studies, which compare the efficacy of gabapentin with that of the respective reference drug, are needed to evaluate its importance in treatment of pain. PMID- 11256158 TI - [Tauopathies--a new class of neurodegenerative diseases]. AB - Recently it was shown by several research groups that mutations in the gene encoding for the tau protein associated with microtubuli on chromosome 17 caused a distinct form of dementia named frontotemporal dementia and parkinsonism (FTDP 17). This disease includes familial asymmetrical frontal and, in the further course, frontotemporal dementia, parkinsonism, which is often initially sensitive to levodopa, signs of upper motor neuron degeneration, and, less commonly, amyotrophy. Tau is an intracellular protein of the cytoskeleton, which is responsible for the arrangement and stabilization of microtubuli. The discovery of mutations in the tau gene causing a distinct neurodegenerative disease in humans has firmly established the importance of the tau gene for neurodegenerative processes, not only in tauopathies but also in other degenerative disorders with tau pathology, such as corticobasal degeneration, supranuclear progressive paralysis, amyotropic lateral sclerosis, parkinsonism dementia complex of Guam, and Alzheimer's disease. Our experience with patients suffering from PTDP-17 shows that its phenotype varies more than was described in the first consensus conferences. In the future, it will be important to designate the diagnostic gold standard not by clinical description, but etiologic classification. PMID- 11256159 TI - [Clinical symptoms, origin, and therapy of the "pusher syndrome"]. AB - Stroke patients may exhibit the peculiar behavior of actively pushing away from the nonhemiparetic side, leading to lateral postural imbalance and a tendency to fall towards the paralyzed side. These patients use the nonparetic extremities to stem actively against attempts of passive correction towards upright orientation. This phenomenon has been called the "pusher syndrome". Recent findings disclose that the deficit leading to contraversive pushing is an altered perception of the body's orientation in relation to gravity. Pusher patients experience their body as upright when they are actually tilted to the nonhemiparetic side. In contrast, processing of visual and vestibular inputs for the determination of visual vertical was undisturbed. The results argue for a separate pathway in humans for sensing gravity apart from that for perception of the visual world. This second graviceptive system decisively contributes to our control of upright body posture. The present article describes this still largely unknown neurological disease. The clinical examination of contraversive pushing, its underlying disturbance, lesion location, and approaches for therapy are considered. PMID- 11256160 TI - [Posttraumatic dystonia. Review and legal aspects]. AB - More attention should be paid to dystonia as a consequence of trauma, particularly with regard to legal aspects. The underlying pathophysiological mechanisms of dystonia following central or peripheral trauma are largely unknown. Hemidystonia after severe head trauma is regarded to be due to contralateral basal ganglia lesions, particularly of the putamen. Focal and segmental dystonias follow various kinds of peripheral trauma. Central synaptic reorganisation due to altered peripheral input may play a role in its genesis. Clinically, post-traumatic dystonia differs from the idiopathic disease by the presence of accompanying pain or causalgia, limitation of the range of movement up to fixed posture, and poor response to conventional pharmacotherapy. If an expert opinion is requested, it is important to ascertain the diagnosis clinically and by EMG. To establish the cause-and-effect relationship between trauma and movement disorder, the severity of the injury, time course, and anatomical relationship must be taken into consideration. PMID- 11256161 TI - Follicular development in vitro. AB - There has been tremendous interest in recent years in the culture of oocytes and follicles. Although much of the research using follicle culture aims to increase understanding of the regulation of follicle development, an important goal has been to develop a method that will eventually allow maturation of human oocytes from the primordial follicle to the mature Graafian stage. We are still some way from this at present, although it has now been achieved in the mouse. In this article, we consider various methods of follicle culture for primordial, preantral, and antral follicles. In vitro development of primordial follicles has used primarily whole ovaries or ovarian fragments as a source of follicles. Culture of later stages of follicle development uses mainly isolated follicular units, either whole (with an intact basement membrane and, in some cases, attached thecal cells) or nonintact (oocyte-somatic cell complexes, which may or may not have remnants of basement membranes and/or thecal cells attached). PMID- 11256162 TI - Oocyte maturation. AB - The oocyte is dependent on granulosa cells to provide nutrients and regulatory signals. Granulosa cells must be at the appropriate stage of differentiation to initiate these signals and transmit them to the oocyte. Studies have shown that in vitro-matured oocytes from follicles in early stages of atresia are more competent to support embryonic development than those from actively growing follicles. The acquisition of developmental competence appears to occur prior to in vitro maturation and can be induced by gonadotropin-free coasting in vivo or postmortem ovary incubation in vitro. The acquisition of developmental competence is probably a common signaling or differentiation pathway that occurs in the oocyte and/or associated granulosa regardless of whether the oocyte is destined to ovulate or degenerate. Early follicle atresia is the visually discernible characteristic in vitro that is associated with increased developmental potential. PMID- 11256163 TI - Assessment of sperm competence. AB - The predictive values of four categories of established sperm function assays- computer-assisted semen analysis (CASA), induced-acrosome reaction testing, sperm penetration assay, and sperm-zona pellucida binding assays--are still unsure. In this article we examine the evaluation of sperm competence. We propose that assessment of sperm competence should include investigations at the nuclear, organelle, and cytoskeletal levels. In light of this, we discuss the assessment of sperm nuclear integrity as an alternative new method of analysis. We also question the merit of having such tests, whereby it may be an easier choice to direct these patients straight to an assisted reproduction treatment. PMID- 11256165 TI - Cytoskeletal aspects of assisted fertilization. AB - During mammalian fertilization, the zygotic centrosome organizes a large sperm aster, critical for uniting the male and female pronuclei prior to first mitosis. Fluorescent imaging of inseminated human oocytes has shown that centrosomal defects may result in abnormal microtubule nucleation preventing genomic union, suggesting a novel cause of fertilization failure. Working with rhesus monkey gametes, we have developed a preclinical model for understanding the cell biological basis of intracytoplasmic sperm injection (ICSI). Typically, ICSI results in abnormal nuclear remodeling during sperm decondensation due to the presence of the sperm acrosome and perinuclear theca, structures normally removed at the oolemma during IVF; this is turn causes a delay in the onset of DNA synthesis. These unusual modifications raise concerns that the ICSI procedure itself may result in chromatin damage during DNA decondensation and further highlight the need for a more rigorous assessment of methods of assisted reproduction prior to their global application. PMID- 11256164 TI - Human sperm-oocyte recognition and infertility. AB - The incidence of infertility related to both male and female factors continues to rise despite many advances in reproductive technologies. Some abnormalities in human gamete interaction have been shown to be due to defects in the sperm, and others have been attributed to defects in the zona pellucida (ZP). Our lack of understanding of molecular mechanisms involved in the interaction of human sperm with the ZP in fertile as compared with infertile females and males has been limited because of the unavailability of human oocytes and ethical restraints on experimental studies. It is becoming increasingly apparent that improved clinical assays are necessary for evaluating sperm-ZP interaction in order to assess the optimal procedures for successful fertilization and pregnancy. With advances in molecular biology, the genes encoding the three major human ZP proteins have been identified and complementary DNAs are available to begin to better evaluate the molecular basis of sperm-ZP interaction. PMID- 11256166 TI - ICSI and its outcome. AB - Since its introduction in 1992, intracytoplasmic sperm injection (ICSI) has become a popular assisted fertilization technique proved very efficient in treating male factor infertility. Many healthy children have been born worldwide from this procedure, and their physical and mental development appears to be within the normal limits. However, because of the peculiarity of the technique and the poor characteristics of the spermatozoa used, concern about the safety of ICSI still exist. In this article, we analyze the in vivo development of embryos conceived after ICSI as well as the obstetric outcome, occurrence of chromosomal abnormalities, and rate of congenital malformations in neonates born as a result of this treatment. A total of 2435 couples were studied in whom the male partners were presumed to be the cause of repeated failed attempts at in vitro fertilization (IVF) or had semen parameters that were unacceptable for conventional IVF treatment. Pregnancies resulting from 3573 ICSI cycles were analyzed; pregnancy outcome data were obtained from the records of obstetrician gynecologists and/or pediatricians. The overall clinical pregnancy (fetal heartbeat) rate was 44.8% with a resultant delivery rate of 39.2% per ICSI cycle (n = 1388). In 37 of the 77 miscarriages for which cytogenetic data were available, an autosomal trisomy was found in each and 29 additional pregnancies were terminated because of a chromosomal abnormality revealed by prenatal diagnosis. There was an equal distribution of vaginal deliveries and cesarean sections (n = 682 and n = 658, respectively). Of the 2059 neonates resulting from ICSI treatment, 38 (1.8%) presented with congenital abnormalities (22 major and 16 minor). When the frequency of miscarriages and congenital malformations was analyzed in terms of semen origin, the outcome was no different between ICSI and IVF. The course of pregnancies and occurrence of congenital malformations following treatment by ICSI are within the ranges obtained following conventional IVF. PMID- 11256167 TI - Oocyte and embryo polarity. AB - The development of all low-order animals and noneutherian mammals follows an organized, polarized directional course from fertilization through fetal development. New evidence points to a fundamental polarization during all steps of mammalian development, from the early oocyte through fertilization and gastrulation. The generator of this polarization is primarily at the genetic level, with the results of gene expression and checkpoints being manifested in phenotype. Although cell-cell interactions reinforce the polarization of the embryo, they are not the underlying means of establishing axes in eutherian embryos. The ability of mammalian cells to remain totipotent is only partial, with little evidence that isolated blastomeres can result in full fetal development. The isolated blastomere can, however, contribute to development if reintroduced into a polarized environment. Polarization begins in the unovulated oocyte and is reinforced at fertilization. The axes and polarization established at fertilization endure through to the blastocyst stage and define the axes during gastrulation and fetal development. PMID- 11256169 TI - Regulation of ionic homeostasis by mammalian embryos. AB - Control and regulation of cellular homeostasis are essential for normal embryo development and maintenance of viability. By understanding the role of ionic homeostasis in normal cell development and homeostatic control by the developing embryo, it is possible to develop culture systems that minimize cellular stress and therefore maintain embryo viability. This article discusses the regulation of intracellular levels of protons (pHi), calcium, magnesium, and phosphate in mammalian embryos. PMID- 11256168 TI - Junctional complexes in the early mammalian embryo. AB - Preimplantation embryos generate intercellular junctions during differentiation of the trophectoderm epithelium and the formation of the blastocyst. These membrane complexes comprise gap junctions, adherens junctions, tight junctions, and desmosomes, each performing fundamental roles in cellular communication, adhesion, and differentiation. The mouse embryo has been used as a model for the biogenesis of cell junctions. Their construction is achieved by temporally regulated gene expression programs. Mechanisms of junction membrane assembly include the timing of transcription, translation, and post-translational modifications of specific junctional proteins. Human embryos exhibit similar expression programs, and defects in these programs may contribute to reduced embryo viability. PMID- 11256170 TI - Embryo nutrition and energy metabolism and its relationship to embryo growth, differentiation, and viability. AB - Over the past decade there has been a resurgence of interest in the culture media used in clinical in vitro fertilization. Unfortunately, during this time more confusion than consensus appears to have developed regarding the composition of these media. In order to facilitate a clearer understanding of this field, it is important to understand the role of specific medium components and how their use is regulated by the embryo. The roles of the key nutrients glucose, pyruvate, lactate, and amino acids during the preimplantation period have therefore been presented. Analysis of how the embryo regulates the utilization of such nutrients has led to a clearer understanding of the embryo's requirements during the dynamic period of preimplantation development. From such information, sequential culture media have been developed along with novel noninvasive tests of embryonic viability. It is proposed that continued studies on the human embryo will lead to further improvements in embryo culture conditions and the optimization of viability assays, culminating in the ability to transfer single embryos for the majority of, if not all patients. PMID- 11256171 TI - Human embryonic stem cell technology. AB - Undifferentiated human embryonic stem (ES) cells can be cultured indefinitely and yet maintain the potential to form almost every cell in the adult human body. Therefore ES cells provide a model for understanding the differentiation and function of human tissue, offer new strategies for drug discovery and testing, and have the potential to provide new transplantation therapies for the treatment of a wide variety of human diseases. In this article, we describe the origin and properties of human ES cells, distinguish ES cells from other pluripotent stem cell lines, and discuss their implications for basic research and human medicine. PMID- 11256172 TI - [Exploration of the uterine cavity in the gynecologic preoperative diagnosis]. AB - BACKGROUND: The diagnostic accuracy of dilatation and curettage (D & C) was studied comparing retrospectively the results of histologic findings of D & C with the correspondent specimen from hysterectomy. METHODS: During five years, at the Institute of Gynecology and Obstetrics, II University of Studies in Naples, 260 women underwent hysterectomy, 160 of which underwent D & C prior to hysterectomy. The histologic findings were classified in: a) physiological endometrium; b) hyperplasia; c) polyps; d) atrophia; e) adenomatous hyperplasia; f) adenocarcinoma. During the period January 1989-October 1993, 260 patients underwent hysterectomy. The age was between 32 and 65 years. The indications to the intervention were: menometrorrhagia, hypogastric pains, dysmenorrhea, metrorrhagia, genital prolapse, urinary incontinence, anemia. Two hundred-sixty patients underwent hysterectomy, 160 of which underwent D & C prior to hysterectomy. Curettage was performed using a right size curette after dilatation of the uterine cervix using Hegar's metallic dilatator. Patients were submitted to general anesthesia. Histologic tissues were fixed with formalin and were sent to the Institute of Anatomopathology for examinations. RESULTS: The histologic results obtained by cavitary exploration have been compared with those reached by the analysis of the surgical samples. The results obtained confirm the reliability of D & C for the identification of endometrial lesions. CONCLUSIONS: Therefore, the diagnostic utility of cavitary exploration before hysterectomy is confirmed, particularly in selected cases. PMID- 11256173 TI - [Assisted labor among non European community pregnant women at the Gynecology and Obstetrics Clinic of the Verona University]. AB - BACKGROUND: The aim of this paper is to evaluate the role and the prevalence of the non-European Community pregnant women in our Institute during the period 1992 1998. The peculiarity of the female immigration in the world and particularly in Italy is stressed from the point of view of the different cultural, ethnic and religious problems of these women. METHODS: During the observed period 8972 women delivered; 434 of them came from non-European Community countries and their individual (age, country) and obstetric (parity, physiological or pathological evaluation of pregnancy, mode of delivery) data were observed. On the basis of the different countries of provenance these women have been subdivided into five groups (East Europe, North Africa and Middle East, Central Africa, Far East and Latin America). RESULTS: The percentages of preterm births (24.2% vs 23.1%), of < or = 1500 g newborns (6.9% vs 5.3%) and of caesarean sections (34.3% vs 27.7%) are higher in the non-European Community women that delivered in our Institute. In 222 (51.1%) cases the women delivered without induction of labour; while in 14.5% of cases it was induced. The length of labour and the genital conditions (episiotomy, tearing) were considered in all ethnic groups of women. CONCLUSIONS: On the basis of the literature and of the analysis of our data, some suggestions about the management of labour and delivery of non-European Community women in Italy are proposed. In particular, the problems of linguistic communication and of the hospital staff preparation in the assistance to labour and delivery are stressed. PMID- 11256174 TI - [Tamoxifen and giant endometrial polyps]. AB - Tamoxifen is a synthetic non-steroid anti-estrogen that has been used effectively for several years in the adjuvant treatment of breast cancer. Although its therapeutic effect is due to its anti-estrogenic properties, the drug also shows modest type B estrogen-receptor agonist activity during the menopausal period in which estrogens are at a low level. Owing to the fall in estrogen levels in menopause, tamoxifen provokes an up-regulation of both estrogen and progesterone receptors at an endometrial tissue is a direct consequence of this. This proliferation, which is the result of an inappropriate response of the basal layer and the basis for the onset of hyperplasia and polyps in the tissue. At standard therapeutic dosages, tamoxifen in postmenopausal women is associated with the onset of alterations in the vaginal and endometrial epithelium. Cases of endometrial hyperplasia, endometrial polyps, adenomyosis, endometriosis and fibromyomas are described in the literature. Endometrial polyps represent the most common pathology associated with TAM in women with previous breast cancer in menopause. The estrogenic stimulus to polyps following TAM treatment may be considerable, resulting in their growth to sizeable proportions, causing metrorrhagia and suspected neoplastic pathology. Two cases of patients receiving adjuvant treatment with tamoxifen for previous breast cancer, who presented two giant endometrial polyps of uncommon dimension, are reported. PMID- 11256175 TI - Aggressive angiomyxoma of the pelvis. A case report. AB - Aggressive angiomyxoma is a locally mesenchymal, benign, and rare neoplasm. The vulva, perineum, and pelvis are the most common sites of involvement. The preoperative diagnosis is postulated by CT, sonography, MR image and angiography. The immunohistochemical study reveals the definitive diagnosis. The therapy is only surgical, and because of its tendency to recur locally, the excision has to be as complete as possible. PMID- 11256176 TI - [Rubella vaccination in the post-partum: which is the best approach?]. AB - BACKGROUND: Two different post partum Rubella vaccination's programs for seronegative women in puerperium are compared. METHODS: In the Borgosesia hospital (Piedmont Region) the vaccination has been proposed by gynecologists already during pregnancy to all the pregnants proved to be negative at the Rubella's test in the initial pregnancy. The little number of enlisted patients is the consequence of the availability of only one doctor to take part in the vaccination's program. In this hospital the cost of vaccination was paid by the patient that had to buy the vaccine before hospitalization. In the Vigevano's hospital (Lombardia Region) the rubella's vaccination has been instead proposed by the nursery pediatrist to all the women who has just given birth in the period between 1995-1997, seronegative for rubella's test in pregnancy or in puerperium. RESULTS: The results (88% of the seronegative women vaccinated in Borgosesia versus 72.6% in Vigevano) show that the best way is to make the pregnant women aware by the gynecologist during pregnancy, even though the vaccine wasn't free for the patients. CONCLUSIONS: The action of pediatrists combined with the availability from hospital administration to offer a free vaccine will make possible to reach an almost total success of the Rubella vaccination's program. PMID- 11256177 TI - [Quaternary ammonium salts in gynecology and obstetrics]. AB - We have summarized chemical, physical, and microbiological characteristics of Quaternary Ammonium Salts: particularly, benzalchromium chloride and didecyl dimethyl-ammonium chloride characteristics were analyzed. These compounds may act as antimicrobial agents in different way: 1) they are surface-active agents and will denature protein or cause dissociation of an enzyme from its prosthetic group; 2) they may alter the cell permeability of bacteria and yeasts; 3) they may stimulate the glycolysis reaction and 4) may inhibit oxidation of lactate. These latest activities may play a role in maintenance of physiologic microbial ecology of vagina or in the re-establishment of the vaginal ecosystem after vaginitis or vaginosis. We have also summarized the physiologic variation of vaginal ecosystem during the different phases of women's life and the microbiology of vagina during vaginitis and vaginosis. The results of more recent studies about the therapeutic role of quaternary ammonium compounds in vulvo vaginal infections an in vaginosis are synthetically reported. We concluded that quarternary ammonium compounds are efficacious, handy and safe, in obstetrics and gynecology. A very good compliance and low costs of these compounds suggest that they may be used alone as well in association with specific antimicrobial agents in the treatment of most of gynecological infections, and particularly in bacterial vaginosis. PMID- 11256178 TI - [Female genital mutilation and legislation]. AB - This article deals with the legal aspect concerning female genital mutilations (FGM). Such a practice (a partial excision of the external genitalia) is highly widespread in Central Africa, especially in Ethiopia and Somalia, and currently involves approximately 130,000,000 women worldwide and, in Italy, about 30,000 women amongst the immigrant population. Since 1982 the World Health Organization (WHO), which condemns such a practice as injurious to women's rights and health, proposed that laws and professional codes prohibit it in all countries. Legislation, although insufficient as a sole measure, is considered indispensable for the elimination of FMG. Since a long time some western countries (Sweden, Great Britain, Belgium and Norway), involved by immigration from countries with FGM tradition, legislated with regard to FGM. In Italy, a specific law does not exist; however, FGM are not allowed by the article 5 of the Civil Code. Nevertheless, recently, several cases of mutilations took place: this led some members of the Parliament to introduce a bill in order to specifically forbid FGM. The authors believe that legislation could effectively support the job of prevention and education, which physicians may carry out in order to save little girls from the risk of familial tradition of genital mutilations. PMID- 11256179 TI - Role of the locus coeruleus in the static and dynamic control of posture. PMID- 11256180 TI - Remembering Nathaniel Kleitman. PMID- 11256181 TI - Induction of wakefulness and inhibition of active (REM) sleep by GABAergic processes in the nucleus pontis oralis. AB - The present study was undertaken to explore the role of brainstem GABAergic processes in the control of the behavioral states of sleep and wakefulness, and to compare the effects of GABAA agonists and antagonists with those of GABAB agonists and antagonists on these behavioral states. Accordingly, the following drugs were microinjected into the nucleus pontis oralis (NPO) in chronic, unanesthetized cats: muscimol (GABAA agonist), bicuculline (GABAA antagonist), baclofen (GABAB agonist) and phaclofen (GABAB antagonist). The percentage, latency, frequency and duration of each behavioral state were measured in order to quantify the effects of these microinjections on wakefulness and sleep. Microinjections of either muscimol or baclofen immediately induced wakefulness. There was a significant increase in the duration and the percentage of time spent in wakefulness as well as an increase in the latency to active (REM) sleep. These changes were accompanied by a decrease in the percentage of time spent in active and quiet sleep. In contrast, injections of bicuculline or phaclofen produced active sleep. The percentage of time spent in active sleep and the frequency of active sleep increased while the percentage of time spent in wakefulness and the latency to active sleep was significantly reduced. The effects of GABAA receptor agonists and antagonists on wakefulness and active sleep were comparable, but stronger than those of GABAB receptor agonists and antagonists. These data indicate that pontine GABAergic processes acting on both GABAA and GABAB receptors play a critical role in generating and maintaining wakefulness and in controlling the occurrence of state of active sleep. PMID- 11256183 TI - Sleep in a nonagenarian: N. Kleitman. PMID- 11256182 TI - Carbachol models of REM sleep: recent developments and new directions. AB - Since the early '60s, injections of a broad-spectrum muscarinic cholinergic agonist, carbachol, into the medial pontine reticular formation (mPRF) of cats have been extensively used as a tool with which to study the neural mechanisms of rapid eye movement (REM) sleep. During the last decade, new carbachol models of REM sleep were introduced, including chronically instrumented/behaving rats and "reduced" preparations such as decerebrate or anesthetized cats and rats. The combined results from these distinct models show interspecies similarities and differences. The dual nature, both REM sleep-promoting and wakefulness (or arousal)-promoting, of the cholinergic effects exerted within the mPRF is more strongly expressed in rats than in cats. This strengthens the possibility suggested by earlier central neuronal recordings that active wakefulness and REM sleep have extensive common neuronal substrates, and may have evolved from a common behavioral state. Carbachol studies using different intact and reduced models also suggest that powerful REM sleep episode-terminating effects originate in suprapontine structures. In contrast, the timing of REM sleep-like episodes in decerebrate models is determined by a pontomedullary neuronal network responsible for the generation of an ultradian cycle similar to the basic rest-activity cycle of N. Kleitman. Other presumed species differences, such as the more widespread distribution of carbachol-sensitive sites or the relative failure of carbachol to increase the duration of REM sleep episodes in rats when compared to cats, may be of a quantitative or technical nature. While carbachol and many other neurotransmitters and peptides microinjected into the mPRF evoke, enhance or suppress REM sleep, the most sensitive site(s) of their actions have not been fully mapped, and the nature of the cellular and neurochemical interactions taking place at the sites where carbachol triggers the REM sleep-like state remain largely unknown. Similarly, little is known about the pathways between the mPRF and medial medullary reticular formation, but the existing evidence suggests that they are reciprocal and essential for the generation of both natural and carbachol-induced REM sleep. Studies of the mesopontine cholinergic neurons, which are hypothesized to be the main source of endogenous acetylcholine for the mPRF, need to be extended to neurons of the mPRF and cells located functionally downstream from this important site for REM sleep, or both REM sleep and active wakefulness. PMID- 11256184 TI - State-related discharge of neurons in the brainstem of freely moving box turtles, Terrapene carolina major. AB - We have performed the first study of neuronal activity in freely-moving reptiles. 23 brainstem units were recorded from areas throughout the reticular formation, during wakefulness and quiescence in the box turtle. These units responded to various sensory stimuli and increased firing rates in relation to motor activity during wakefulness. All but one unit showed significant decreases in discharge during quiescence. Group I cells (32%) fired mostly during active movements and exhibited silent periods of 5 min or longer during quiescence while group II cells (68%) maintained slow tonic activity in quiescence. Polygraphic data showed no consistent, cyclically occurring phasic events during quiescence. PMID- 11256185 TI - A tribute to Nathaniel Kleitman. PMID- 11256186 TI - Active neocortical processes during quiescent sleep. AB - The neocortex and the thalamus constitute a unified oscillatory machine during different states of vigilance. The cortically generated slow sleep oscillation has the virtue of grouping other sleep rhythms, including those arising in the thalamus, within complex wave-sequences. Despite the coherent oscillatory activity in corticothalamic circuits, on the functional side there is dissociation between thalamus and neocortex during sleep. While dorsal thalamic neurons undergo inhibitory processes induced by prolonged spikebursts of GABAergic thalamic reticular neurons, the cortex displays, periodically, a rich spontaneous activity and preserves the capacity to process internally generated signals. Simultaneous intracellular recordings from thalamic and cortical neurons show that short-term plasticity processes occur after prolonged and rhythmic spike-bursts fired by thalamic and cortical neurons during slow-wave sleep oscillations. This may serve to support resonant phenomena and reorganize corticothalamic circuitry. PMID- 11256187 TI - From slow waves to sleep homeostasis: new perspectives. AB - EEG slow waves are the epitome of deep nonREM sleep. The level of slow-wave activity (SWA; defined as spectral power in the 0.5-4.5 Hz band) in the initial part of sleep is determined by prior sleep and waking, and thereby represents a marker of a homeostatic sleep regulating process (Process S). Models based on SWA were successful in simulating sleep architecture in a variety of experimental protocols. SWA is an exceptional sleep variable in that it is little influenced by circadian phase and variations of the photoperiod. There is recent evidence that it is not waking per se but the absence of sleep, which engenders a rise in sleep propensity. Thus animals emerging from the hypometabolic states of hibernation or daily torpor exhibit an increase in SWA akin to sleep deprivation. Recent human studies showed SWA to be a marker of a local, use-dependent facet of sleep. Selective activation of specific cortical areas during waking enhanced SWA over the activated region during sleep. A frontal predominance of power in the 2 Hz band was documented in the initial part of a normal sleep episode. Sleep homeostasis may be a valuable concept for exploring the evolutionary origin of sleep. Thus 'rest homeostasis' has been demonstrated in invertebrate species, and the search for homologies of rest and sleep on a molecular genetic level has begun. Conceptualizing and characterizing sleep as a regulated process may eventually shed light on its function. PMID- 11256188 TI - Hypothalamic sleep-promoting mechanisms: coupling to thermoregulation. PMID- 11256189 TI - Preoptic area sleep-regulating mechanisms. AB - Evidence is summarized for the existence of a sleep-regulating mechanism within the preoptic area of the hypothalamus, including the results of lesion, stimulation, and neuronal recording studies. Recent findings employing the c-fos protein immunohistochemical method, have localized putative sleep-regulatory neurons to the ventrolateral preoptic area (vlPOA) and the median preoptic nucleus (MnPn). Electrophysiological studies have confirmed the presence of neurons with sleep-related discharge in the vlPOA. Neurons in the vlPOA that exhibit c-fos protein immunoreactivity during sleep contain the inhibitory neuromodulators galanin and gamma-aminobutyric acid (GABA). These neurons also project to monoaminergic arousal systems, particularly the histaminergic cell groups in the posterior hypothalamus. POA neurons can be hypothesized to provide sleep-related inhibitory control over multiple arousal systems in the forebrain and brainstem. PMID- 11256190 TI - Pontine structures and mechanisms involved in the generation of paradoxical (REM) sleep. PMID- 11256191 TI - Clinical features in 130 patients with the velo-cardio-facial syndrome. The Leuven experience. AB - The velo-cardio-facial syndrome (VCFS) is a leading cause of velopharyngeal dysfunction and cleft palate and caused by a submicroscopic deletion in the long arm of chromosome 22 (band 22q11). During the last 5 years, 130 patients with a 22q11 deletion were diagnosed in Leuven. Most patients presented a wide variety of the classical features of the velo-cardio-facial syndrome. Velopharyngeal dysfunction was almost always present whereas an isolated cleft lip/palate was observed in a minority of patients. The velopharyngeal function can be evaluated by the classic combination of indirect and direct techniques. Because of the frequent occurrence of the velo-cardio-facial syndrome, estimated at around 1/4000 live births, and given the extremely broad clinical spectrum which makes clinical diagnosis difficult, screening of patients with velopharyngeal dysfunction for a deletion 22q11 is indicated. PMID- 11256192 TI - Voice similarity in identical twins. AB - If people are asked to discriminate visually the two individuals of a monozygotic twin (MT), they mostly get into trouble. Does this problem also exist when listening to twin voices? Twenty female and 10 male MT voices were randomly assembled with one "strange" voice to get voice trios. The listeners (10 female students in Speech and Language Pathology) were asked to label the twins (voices 1-2, 1-3 or 2-3) in two conditions: two standard sentences read aloud and a 2.5 second midsection of a sustained /a/. The proportion correctly labelled twins was for female voices 82% and 63% and for male voices 74% and 52% for the sentences and the sustained /a/ respectively, both being significantly greater than chance (33%). The acoustic analysis revealed a high intra-twin correlation for the speaking fundamental frequency (SFF) of the sentences and the fundamental frequency (F0) of the sustained /a/. So the voice pitch could have been a useful characteristic in the perceptual identification of the twins. We conclude that there is a greater perceptual resemblance between the voices of identical twins than between voices without genetic relationship. The identification however is not perfect. The voice pitch possibly contributes to the correct twin identifications. PMID- 11256193 TI - Wegener's granulomatosis triggered by infection? AB - Wegener's granulomatosis is a systemic disease of unknown origin, although recent studies suggest that auto-immune mechanisms and infection play a role in the pathogenesis of this disease. Wegener is characterized by a necrotizing vasculitis involving the lungs (pulmonary infiltrates), the upper airways and the kidneys (rapidly progressive glomerulonephritis). We present a case of a male patient admitted because of progressive deterioration of the general condition with weight loss, a unilateral neck mass, unilateral purulent rhinorrea and fever. CT-scan evaluation demonstrated a unilateral expanding mass in the sing nasal cavity, obliterating the ethmoid complex. MRI revealed signs of intracranial inflammatory reaction and onset of absedation. A malignancy was suspected but a diagnosis of Wegener's granulomatosis was established based on histologic criteria (nasal biopsy) and a positive titer for anti-cytoplasmic antibodies (cANCA). During follow-up, nasal carriage of Staphyloccocus Aureus could be documented. An overview of Wegener's granulomatosis will be provided with emphasis on the potential role of acute infections as a trigger for Wegener's granulomatosis and the head and neck manifestations. PMID- 11256194 TI - Seromucous maxillary sinusitis (SMMS): a clinicophysiological approach. AB - Chronic sinusitis, especially maxillary sinusitis is a common disorder in humans. Seromucous sinusitis is rarely described in the literature. The present study deals with the clinical and laboratory characteristics of a group of patients suffering from the above disorder. During the last 10 years, 32 patients suffering from seromucous maxillary sinusitis were enrolled in the study. Patients' charts were reviewed and tabulated according to age, sex, history, clinical symptoms and laboratory findings. Treatment was based on punction and drainage of the seromucous effluent. Results were also statistically evaluated. Flight trips and atypical episodes of nasal infection were the predisposing factors for seromucous maxillary sinusitis. The only clinical manifestation was coughing, for at least 12 weeks before diagnosis. Sinus effluent was composed by serous and mucous constituents with glue like structure. There were no differences between sexes in predisposing factors, or x-ray findings. The treatment is paracentesis and drainage and in one case of recurrence, middle meatotomy and sinus endoscopy. PMID- 11256195 TI - Oncocytic laryngeal cysts: a case report and literature review. AB - Some 150 cases of oncocytic laryngeal cysts have been published. We report another case of laryngeal oncocytic cysts with atypical presentation of acute, progressive stridor and sore throat. Literature was reviewed with special regard to etiology, clinical presentation, imaging, incidence, localization, associated lesions and treatment options. Oncocytic laryngeal cysts are rare, but may be underreported. They represent a separate clinicopathological entity in the group of all laryngeal cysic lesions and occur in persons over 60 years. The symptomatology varies from asymptomatic to hoarseness and dyspnea. Diagnosis is made by histological examination. Treatment is surgical. Although it is a benign lesion, follow up is recommended, as recurrence is possible. PMID- 11256196 TI - Giant congenital intracranial epidermoid tumor: a case report. AB - A case of a large epidermoid tumor in a middle-aged woman with limited symptomatology is reported. Intracranial epidermoid tumors are slow growing and benign lesions. Although the treatment of choice consists in complete resection, partial removal may be preferred in some very extended cases presenting with minimal symptomatology because of the lower morbidity involved. This case illustrates a key-hole approach via a retrolabyrinthine route with preservation of all labyrinthine structures, including the endolymphatic sac and duct. The epidermoid was partially removed by extensive intracapsular debulking under endoscopic control with the aim of decreasing cerebral compression. All cranial nerve functions were preserved and the 6 month postoperative imaging has remained unchanged over a 2-year long follow-up period. The literature regarding this rare pathology is also reviewed. PMID- 11256197 TI - Turnover of Haemophilus influenzae isolates in otitis-prone children. AB - Arbitrarily primed PCR with primer ERIC2 and RAPD Ready-to-Go beads was applied to study the epidemiology of non-typable Haemophilus influenzae (NTHI) isolated from the nasopharynx of children with recurrent otitis media (otitis-prone children). Thirty-five otitis-prone children (OP-children) were included. Three pairs of siblings were identified in the study population. This study is part of a large prospective multicentric trial investigating the efficacy of a new conjugated pneumococcal vaccine in OP-children (OMAVAX trial). During a 2-year study period, NTHI strains were isolated from nasopharyngeal swabs and, when possible, middle ear aspirates were collected in OP-children between 1 and 6 years of age. In 20 out of the 35 children, 48 H. influenzae isolates were obtained simultaneously from different sites (left and/or right ear and/or nasopharynx) of the same child as well as from siblings or during initial and follow-up visits of the same child. Subsequent genotyping indicated substantial genetic diversity among the H. influenzae isolates studied, since for a total of 48 isolates in 20 OP-children, 29 different genotypes were observed. Simultaneous isolation for different sampling sites (ear or nasopharynx) as well as for siblings resulted mostly in identical fingerprints. Longitudinal follow-up of H. influenzae isolates in the nasopharynx almost always resulted in different genotypes. We can therefore conclude that both cross colonization (between sampling sites within the same patient and between siblings) and turnover of H. influenzae isolates are high in OP-children. PMID- 11256198 TI - [Clinical considerations and statistical analysis on 100 patients with oral lichen planus]. AB - BACKGROUND: Oral Lichen planus (OLP) is an inflammatory disease, characterized by the presence of polygonal papules which can confluence and affect any part of the oral mucosa, and occurring in many different forms. The clinical picture of the popular form may be characterized exclusively by asymptomatic whitish striae, while the erosive form is extremely painful and disabling. The aim of this research was to evaluate the behavior of the reticular and erosive form of OLP taking into consideration the following three pathologies: hepatitis B, C and diabetes. METHODS: The data of 100 patients affected by OLP (43 males and 57 females) where analyzed, the patients have been observed at the G. Eastman hospital in Rome, Italy in the period between November 1995 and May 1998. Group I presented papular lesions, while group II presented atrophic-erosive lesions with or without the presence of papular lesions. RESULTS: A comparative analysis between the two groups, performed with chi 2 corrected by the Yates formula and if necessary followed by the Fisher test, demonstrated a higher average age and a larger extension of the oral lesions for group II (p < 0.05). The presence of viral hepatitis B was 6% overall, its prevalence in group II, 5% (p > 0.05); the presence of viral hepatitis C was 13% overall, its prevalence in group II 11%; the presence of diabetes was 10% overall, its prevalence in group II 7%. The buccal mucosa was the most frequently affected area in both groups. CONCLUSIONS: The results obtained confirm the possibility of an association between hepatitis and OLP, but a statistically significant relationship between hepatitis and OLP erosive type, has not been observed. PMID- 11256199 TI - [Correlation between edentulism, sleep disorders and arterial hypertension. Preliminary research]. AB - BACKGROUND: Respiratory disturbances during sleep are considered risk factors for arterial hypertension and cardiovascular diseases. Edentulism, by decreasing retro-pharyngeal space, may favor upper airway occlusion during sleep. Aim of the study is to evaluate whether edentulism is associated with greater prevalence of sleep disturbances, arterial hypertension and other cardiovascular diseases. METHODS: Eighty edentulous subjects with dentures (removed during sleep) and 57 subjects with natural teeth, matched for age, underwent assessment of oral conditions and recording of questionnaires on diseases and respiratory sleep disturbances. RESULTS: Edentulous subjects had a significantly higher prevalence of arterial hypertension and cardiovascular diseases, than subjects with natural teeth. CONCLUSIONS: In edentulous subjects, removing dentures during sleep may favor respiratory disorders, and increase the risk for hypertension and cardiovascular disease. PMID- 11256200 TI - [The influence of edentulism on spirometric values]. AB - BACKGROUND: Aim of the paper is to study whether edentulism, by causing a decrease in size and tone of pharyngeal musculature, may affect spirometric measurements. METHODS: Spirometry was recorded with and without dentures in 58 edentulous subjects, 36 asymptomatic normal subjects (N) and 22 patients with chronic obstructive pulmonary disease (COPD). In 10 subjects retropharyngeal space with and without dentures was assessed by cephalometry. RESULTS: In the N group, removing dentures produced a significant decrease in lung volumes and airflow rates, while in COPD patients it was ineffective. In both groups, retropharyngeal space was significantly decreased by removing dentures. CONCLUSIONS: Edentulism, by decreasing extrathoracic airway caliber, influences significantly spirometric measurements in normal subjects but not in those with COPD. PMID- 11256202 TI - [Complicated tooth replantation]. AB - The materials and techniques for tooth replantation in presence of complications are discussed on the basis of recent advances in oral medicine and surgery and dentistry. The following complications are examined: mucous and cutaneous wounds, retention of teeth and foreign bodies, bony and radicular fractures with or without tissue loss, root drying, infection. Other problems are also discussed: replantation of deciduous teeth, endodontic therapy, stabilization of replanted teeth, application of orthodontic forces. Some technical principles are presented for a better management of various clinical situations. At last, interesting clinical cases are reported. PMID- 11256201 TI - [Mouth opening in patients with systemic sclerosis: base analysis and during follow-up]. AB - BACKGROUND: A large number of orofacial abnormalities have been described in patients with Systemic Sclerosis (SSc) but no data are reported on the correlation with different subgroups of patients on the efficacy of different therapies. METHODS: In the present study mouth opening was retrospectively evaluated in 40 patients with SSc in whom measurement of interlabial distance was taken at the first clinical control and during follow-up. The data confirmed that the mouth opening is significantly decreased in patients with SSc independently from sclerosis subgroup, age or disease duration. RESULTS: Follow-up (8 +/- 8.3 years) showed a different behaviour of the parameter: in 12 patients (group I) no variation in mouth opening was detected, in 18 patients (group II) a decrease and in 10 patients (group III) an increase was observed. CONCLUSIONS: The only difference between the three groups was the treatment received: 80% of the patients of group III (p < 0.01) have been treated with cyclophosphamide (CF). Our data further support the efficacy of treatment with CF in patients with SSc. PMID- 11256203 TI - [Necrotic lesions of the oral cavity in chronic leukemia. Treatment with CO2 laser]. AB - The main characteristics of oral necrotic lesions in leukemia and the treatment and consequence of these pathologies are described. These lesions are classified as noma-like lesions, necrotic gangrenous ulcers of the oral cavity mucosa appearing in severe organic diseases. A new therapeutical approach in the treatment of oral necrotic lesions in chronic leukemia by the use of CO2 laser is tried and a case with the results obtained is presented. PMID- 11256204 TI - [Intraoperative lesions of the maxillary artery and its immediate treatment. Presentation of a clinical case]. AB - The authors discuss the possible vascular lesions that may occur during mandibular sagittal split ramus osteotomies with particular regard to the maxillary artery. A case of surgical lesion of the maxillary artery is presented and its course and anastomoses are analyzed. The ligation of the external carotid artery and its principal branches is the treatment of choice in case of maxillary artery lesion which is a life threatening event and needs immediate intervention. PMID- 11256205 TI - Canal filling in primary teeth. Description of two particular cases. AB - Endopedodontics is that branch of child dentistry that regards pulp treatment in childhood. It involves endodontic treatment of all primary teeth and incisives, and of the first permanent molars with immature apex. The pedodontist has to exactly evaluate the techniques and the material to use in treating his patients. It is important to realize that we are facing a continuously evolving and absolutely unstable dentition. Therefore, the success of our work is the result of a careful evaluation of diagnostic parameters and of the techniques and the material. In the current clinical cases it is obvious how the lack of only one of the parameters listed, could represent the cause of severe damage to the patient's dental health and it could lead to psychological discomfort (if not damage), considering the age of the patient. PMID- 11256206 TI - [Surgical removal of an osteotribe fractured during the extraction of an embedded third molar. Clinical case]. AB - Although rarely reported, the fracture of an osteotribe during the extraction of embedded lower third molars may have severe consequences on the surrounding anatomic structures possibly resulting in permanent damage to the patient. Operators must take special care when choosing instruments, which must comply with particular quality requisites, and during their use in order to avoid intraoperative accidents and subsequent complications. The authors describe a case that was brought to their attention in which the fragment of an osteotribe that had fractured during a previous operation was lodged in the submandibular fossa, penetrating the upper part of the homonymous gland. After the necessary clinical tests, the authors proceeded to remove the fragment thus leading to the resolution of the symptoms reported by the patient. PMID- 11256207 TI - Intelligence is universal in life. AB - Behaviorists assume that living things memorize random atoms of information (engrams), "reinforced" by success, just as in the neo-Darwinian mutation selection process. On the contrary we have to recognize the existence of organized and systematic responses in the learning process (Krechevsky). The animals seek desperately to "understand the meaning" of the world around them, by widening its context. Intelligence is not an exclusive prerogative of human mind. The minds of insects operate in the same way as that of man. Even a cell has a sort of intelligence (Cuenot). Consciousness is a state of awareness associated with enhanced mental activity. It occurs also in other "higher" animals (Thorpe). However human themselves are non conscious of their basic underlying motivations. Unconscious or ineffable knowledge plays a great role in shaping our world-view and in determining our influence on the Gaian hierarchy. PMID- 11256208 TI - Physiological differentiation of the mitochondria during Bufo bufo development. AB - 1) The oxygen consumption increases during Bufo bufo development in accordance with the two steps which border at the "heart beat" stage. 2) Cytochrome c oxidase activity is not proportional to the oxygen consumption: it is notable and constant in the first step, and it only increases in the second. 3) In the mitochondria of preneural embryos, citrate synthase, NADP+ dependent isocitrate dehydrogenase, and succinate dehydrogenase activities are very low in respect to malate dehydrogenase and glutamate oxaloacetate transaminase activities. The Krebs cycle results lowered at the condensing reaction level with acetyl accumulation when pyruvate is available. The same behavior has been observed in the Xenopus laevis oocytes and differentiated tissues. 4) The presence of a phosphagen system which is different from creatine phosphate and arginine phosphate, supporting ATP level, has been demonstrated in B. bufo embryos. 5) Mitochondria of postneural embryos are able to accomplish a complete Krebs cycle by increasing citrate synthase, and succinate dehydrogenase activities. 6) In all B. bufo development, malate dehydrogenase and glutamate oxaloacetate transaminase constitute a multienzymatic system by which the mitochondria accomplish a decarboxylic amino acid shunt required for the transformation of deutoplasm into protoplasm. This shunt is also operative in the X. laevis oocytes. 7) Through pyruvate production, by oxidative decarboxylation of malate, the NAD(P)+ dependent malic enzyme could carry out a fundamental anaplerotic function in the mitochondria which is specialized in the production of biosynthetic blocks belonging to the embryo in which the carbohydrates metabolism rather than the glycolytic activity is designed for pentose phosphate and glycerol phosphate synthesis for protein and cytomembrane production. 8) Consistent metabolic differences have been highlighted between B. bufo embryos and X. laevis embryos. PMID- 11256209 TI - Tobacco mosaic virus RNA as genetic determinant: genesis of a discovery. AB - It is generally held that the American geneticists Alfred Hershey and Martha Chase were the first to elucidate, in 1952, the genetic functions of phage DNA. The discovery of the genetic functions of RNA in a plant virus (Tobacco mosaic virus, TMV) is commonly attributed to the American plant virologist Heinz Fraenkel-Conrat, and to the Germans Alfred Gierer and Gerhard Schramm, who came to the same conclusion independently in 1956. In reality, the first understandings dated back to about 1940, when several scientists discovered that TMV infectivity was closely related to the presence of undamaged RNA in the virus particles. A very important but underestimated contribution came from the English group of Roy Markham, Kenneth Smith and Richard Matthews in 1948. This group purified and characterized an isometric plant virus, Turnip yellow mosaic virus, and first showed that virus infectivity depended on the presence of the RNA, concluding that nucleic acid was essential for virus multiplication. This finding was confirmed by the same group one year later but it laid neglected. After a five year period, in which several groups attempted to solve the question of the function of TMV RNA, the American electron microscopist Roger Hart offered, in 1955, further direct evidence which correlated RNA to TMV infectivity. One year later, three research groups (Fraenkel-Conrat; Gierer and Schramm; Max Lauffer, David Trkule and Anne Buzzell) obtained evidence that put an end to the question, which was (and is) fundamental to molecular Genetics because it demonstrated that RNA can function independently of DNA. PMID- 11256210 TI - Multiple-state model of ionic channel in reproducing the time course of electrophysiological events. AB - BACKGROUND: The predictions of the Hodgkin-Huxley model do not accurately fit all the measurements of voltage-clamp currents, gating charge and single-channel currents. There are many quantitative differences between the predicted and measured characteristics of the sodium and potassium channels. For example, the two-state gate model has exponential onset kinetics, whereas the sodium and potassium conductances show S-shaped activation and the sodium conductance shows an exponential inactivation. In this paper we shall examine a more general channel model that can more faithfully represent the measured properties of ionic channels in the membrane of the excitable cell. METHODS: The model is based on the generalisation of the notion of a channel with a discrete set of states. Each state has state attributes such as the state conductance, state ionic current and state gating charge. These variables can have quite different waveforms in time, in contrast with a two-state gate channel model, in which all have the same waveforms. RESULTS: The kinetics of all variables are equivalent: gating and ionic currents give equivalent information about channel kinetics; both the equilibrium values of the current and the time constants are functions of membrane potential. The results are in almost perfect concordance with the experimental data regarding the characteristics of nerve impulse. CONCLUSIONS: The expected values of the gating charge and the ionic conductance are weighted sums of the state occupancy probabilities, but the weights differ: for the expected value of the gating charge the weights are the state gating charges and for the expected value of the ionic conductance the weights are the state conductances. Since these weights are different, the expected values of the gating charge and the ionic conductance will differ. PMID- 11256220 TI - Ontogeny, phylogeny and the origin of biological information. AB - Accepting the evidence that evolution is largely finished and that sexual reproduction is incapable of supporting macroevolution, indicates that macroevolutionary changes were produced presexually through the cytological events associated with the first meiotic division. This reproductive mode is ideally suited to the production of new structural rearrangements of preexisting genetic information in instantaneous homozygous form. These new arrangements (position effects) produce new and discrete species. Thus, speciation results not from new genetic information, but rather from information already present (preformed). The several parallels that exist between epigenesis and preformation in both ontogeny (development) and phylogeny (evolution) are discussed. I propose that both of these phenomena have proceeded through the selective activation (derepression) of an enormous potential supply of information already present at the onset of each of these biological phenomena. Acceptance of these possibilities can serve to liberate us in our quest for the ultimate truth concerning these two closely related phenomena. PMID- 11256221 TI - [Rites and archetypes: from the inorganic to consciousness for a symbolic epistemology]. PMID- 11256222 TI - [What is your diagnosis? Melorheostosis middle phalanx digit IV]. PMID- 11256223 TI - [Status of scalp hair and therapy of alopecia in men in Switzerland]. AB - A community-based interview and a questionnaire of men visiting the dermatologist for treatment of hair loss were conducted in Switzerland, to characterize the significance of scalp hair and self-perception of hair loss in Swiss men, and to evaluate current treatment of hair loss. 508 men, aged 15-74 years, regardless of the degree of hair loss, were interviewed by telephone, and 308 patient questionnaires were completed by 19 dermatologists. The questions addressed by the interview were: degree of self-rated hair loss, time invested for hair care, use or reasons for rejecting hair growing agents, relevant criteria for scalp hair, self-assessment with respect to different "hair communication types". The questionnaire analysed the causes of hair loss, prior and current treatment modalities, and follow-up at the dermatologist. Respondents rated their hair loss on a 5-point, textual scale that ranged from 'no hair loss' to 'bald areas'. 43% reported hair loss to some extent. For 42% a full head of hair was very important, especially for men under 29 years, who invested more time for hair care and had not lost hair. Of men with hair loss, 26% previously applied hair growing agents. Of men consulting the dermatologist for hair loss, 90% had androgenetic alopecia. 37% were previously treated: prior treatment was in 59% minoxidil, in 4% finasteride (Propecia), in 7% Aminexil, in 7% dietary supplements, and in 6% conducted by the hair dresser. In 79% treatment was switched to Propecia: of these, 73% adhered to the follow-up consultations at the dermatologist. PMID- 11256224 TI - [Ambulatory treatment of anxiety disorder. Behavior therapy and drug therapy approaches]. AB - Phobic disorders and other anxiety disorders belong to the most frequent psychiatric illnesses. Since they can be overlapped by other mental illnesses, they are often overlooked in practice. For example, comorbidity between anxiety and depression is very high. Also, personality disorders and a person's biography can lay the ground to anxiety. A typical complication of anxiety disorders is dependence of substances, predominantly of tranquilizers and alcohol. By means of a clinical case example, the topic of anxiety disorders is discussed practically and the treatment concept is detailed. Already during ambulatory care in a general practitioner's office behavioral therapeutic strategies can be implemented besides drug treatment with relatively moderate effort. PMID- 11256225 TI - [Treatment of acute stroke with systemic thrombolysis--benefit or risk?]. PMID- 11256226 TI - ["I recently felt so odd"]. PMID- 11256227 TI - [Decreased tumor size in hypophyseal macroadenoma--empty sella syndrome]. PMID- 11256228 TI - [Breastfed infants and those sleeping with a pacifier awake more easily- indication of a decreased risk for sudden infant death?]. PMID- 11256229 TI - [Nutrition and life style prevent myocardial infarct and stroke: 14 years follow up of the US-American Nurses Health Study]. PMID- 11256231 TI - Comparison of inhibitory activities of donepezil and other cholinesterase inhibitors on acetylcholinesterase and butyrylcholinesterase in vitro. AB - This study was designed to compare the in vitro inhibitory effects on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) of donepezil and some other cholinesterase (ChE) inhibitors which have been developed for the treatment of Alzheimer's disease. The carbamate derivatives physostigmine and rivastigmine needed preincubation to exhibit appropriate anti-ChE activity. The maximum ChE inhibition by physostigmine developed within 30-60 min, while the inhibitory effect of rivastigmine on AChE and BuChE activities reached its peak after 48 and 6 h, respectively. The order of inhibitory potency (IC50) towards AChE activity under optimal assay conditions for each ChE inhibitor was: physostigmine (0.67 nM) > rivastigmine (4.3 nM) > donepezil (6.7 nM) > TAK-147 (12 nM) > tacrine (77 nM) > ipidacrine (270 nM). The benzylpiperidine derivatives donepezil and TAK-147 showed high selectivity for AChE over BuChE. The carbamate derivatives showed moderate selectivity, while the 4-aminopyridine derivatives tacrine and ipidacrine showed no selectivity. The inhibitory potency of these ChE inhibitors towards AChE activity may illustrate their potential in vivo activity. PMID- 11256230 TI - Insulin-like growth factor I inhibits cardiomyocyte apoptosis and the underlying signal transduction pathways. AB - The effects of insulin-like growth factor I (IGF-I) on cardiomyocyte apoptosis induced by hypoxia in cultured neonatal rat cardiomyocyte were investigated. Primary neonatal rat cardiomyocytes were cultured in 95% N2-5% CO2 to imitate the in vivo hypoxic condition. Electron microscopic observation revealed a series of typical morphological changes characteristic of apoptosis in cardiomyocytes under the hypoxic condition. DNA gel electrophoresis showed DNA laddering in an ischemic duration-dependent manner. The hypoxia-induced cardiomyocyte apoptosis was also evidenced by flow cytometry and TUNEL assay. DNA gel electrophoresis showed that IGF-I in a dose range of 10(-9)-10(-7) mol/l could significantly prevent the hypoxia-induced cardiomyocyte apoptosis. The protective effects of IGF-I against hypoxia-induced apoptosis could also be verified by flow cytometry and TUNEL assay. A tyrosine kinase inhibitor (genistein), a MAPK inhibitor (PD 098059) and a P13 kinase inhibitor (wortmannin) could also suppress the antiapoptotic effects of IGF-I. These results suggest that IGF-I can directly alleviate the hypoxia-induced cardiomyocyte apoptosis and that the three kinase routes mentioned above may be involved in its signaling pathways. PMID- 11256233 TI - A new method for radioiodination of polysaccharides and its use in biodistribution studies of an immunomodulating glycoconjugate (Immunoferon). AB - A new method for the labeling of polysaccharides by using tyrosine derivatives after CNBr activation is presented, along with its use for pharmacokinetic studies of an orally active immunomodulating noncovalent glycoconjugate, which was administered labeled either in the polysaccharide or the protein moiety. The relationship between the obtained results and the mechanism of action are also discussed. PMID- 11256232 TI - Investigation into the rapid effects of 17 beta-estradiol and neuroactive steroids upon beta-amyloid(25-35)-induced activation of phosphoinositide-specific phospholipase C in human platelets. AB - In the present study, the rapid effects of five steroids (17 beta-estradiol, progesterone, allopregnanolone, 3 alpha-hydroxy-5 alpha-pregnan-20-one and 3 alpha-hydroxy-5 beta-pregnan-20-one) and the plant steroid trans-resveratrol upon the calcium response to beta-amyloid(25-35) peptide (A beta(25-35)) in human platelets was measured. A beta(25-35) produced a robust increase in intracellular calcium due to a direct activation of phosphoinositide-specific phospholipase C. None of the steroids significantly affected the response to A beta(25-35). In contrast, trans-resveratrol appeared to increase the response to A beta(25-35) at a concentration that decreased the response to thrombin, although the possibility that these changes are artifactual could not be ruled out. It is concluded that although steroids affect human platelet Ca2+ homeostasis, this is not a rapid event, unless very high concentrations are used. PMID- 11256234 TI - Changes in rat liver monooxygenase activities after administration of atenolol, nifedipine and diltiazem alone and in combination. AB - The effects of the Ca2+ antagonists nifedipine (NF) and diltiazem (DL) and of the cardioselective beta 1-adrenergic blocking agent atenolol (AT) on the hexobarbital (HB) sleeping time and on the activity of some liver drug metabolizing enzyme systems in male Wistar rats were studied. Two hours after single oral administration, atenolol (150 mg/kg) did not change hexobarbital sleeping time, while nifedipine (50 mg/kg) and diltiazem (30 mg/kg) prolonged it by 171.2 and 99.6%, respectively. Coadministration of atenolol with diltiazem or with nifedipine significantly prolonged hexobarbital sleep by 205 and 283%, respectively. Administered alone, atenolol decreased the ethylmorphine-N demethylase (EMND) activity, but the amidopyrine-N-demethylase (APND) activity was not changed in any of the treated groups. Atenolol and nifedipine significantly increased aniline-4-hydroxylase (AH) activity and this effect was also observed with the combinations AT + NF and AT + DL. The NADPH cytochrome P 450 reductase activity was significantly decreased by nifedipine and diltiazem. Only nifedipine increased the total content of cytochrome P-450 (by 23.8%). Atenolol and diltiazem tended to increase the content of cytochrome b5 which was increased by nifedipine by 97.6%. The same effect was observed with the combinations AT + NF and AT + DL. The results suggest that NF, AT + NF and AT + DL produced the manifested changes in hepatic oxidative metabolism. The decreased EMND activity by atenolol, however, and the prolongation of hexobarbital sleeping time by nifedipine, diltiazem and their coadministration with atenolol did not correlate with enhanced microsomal P-450 and b5 content. PMID- 11256235 TI - AT1 receptor antagonism enhances angiotensin-II-facilitated carrageenan-induced paw edema. AB - Recent studies have demonstrated that the renin-angiotensin system (RAS) participates in the processes of inflammation. An active component of this system, angiotensin II (Ang II), differentially regulates the production of oxyradicals, nitric oxide, prostaglandins, platelet-activating factors and bradykinins by acting through AT1 or AT2 receptor subtypes. Many of the physiological actions of Ang II mediated through AT1 and AT2 receptors are opposite and thereby show physiological antagonism to each other. In the present study, we evaluated the effect of locally administered Ang II, the ACE inhibitor captopril and the AT1 receptor antagonist losartan in the carrageenan model of acute inflammation. Local administration of losartan (10-50 micrograms/paw) or Ang II (0.2-1 microgram/paw) alone did not induce inflammation, but significantly enhanced the carrageenan-induced edema in a dose-dependent manner. Coadministration of subeffective doses of losartan (10 micrograms/paw) and Ang II (0.2 microgram/paw) significantly potentiated the carrageenan-induced inflammation. In conclusion, the present study predicts that Ang II might be formed locally during carrageenan-induced acute inflammation. Potentiation of the Ang II effect in carrageenan-induced inflammation by losartan may be mediated through over-stimulation of unblocked AT2 receptors or due to stimulation of inflammatory pathways by unknown mechanisms. PMID- 11256236 TI - Involvement of GABAergic neurotransmission in the neurobiology of the apomorphine induced aggressive behavior paradigm, a model of psychotic behavior in rats. AB - The effect of treatment with the acute GABAA receptor agonist THIP and the GABAB receptor agonist baclofen on apomorphine-induced aggressive behavior was studied in adult male Wistar rats. Both THIP (10 mg/kg i.p.) and baclofen (8 mg/kg i.p.) attenuated the aggressiveness, thereby indicating the involvement of GABAergic neurotransmission in the mediation of apomorphine-induced aggressiveness. On the basis of our data it can be proposed that both GABAA and GABAB receptor subtypes are involved in the neurobiology of apomorphine-induced aggressive behavior, as this phenomenon is evidently subject to the general inhibitory effect of GABAergic neurotransmission. PMID- 11256239 TI - [The Internet]. PMID- 11256238 TI - Isoniazid does not affect bioavailability of cyclosporine in renal transplant recipients. AB - Transplant recipients are predisposed to develop opportunistic infections such as tuberculosis, and isoniazid (INH) is used in most antitubercular therapeutic and prophylactic protocols. Cyclosporine (CyA) bioavailability increases with the concomitant use of drugs that inhibit hepatic cytochrome P-450 enzymes. There are conflicting reports on a possible interaction between the two drugs. Seven renal transplant recipients on CyA (Sandimmun Neoral) with slow acetylation status and also requiring concomitant INH prophylaxis (300 mg/day) against tuberculosis were studied. There were no significant changes in CyA pharmacokinetic parameters including CyA trough levels, total CyA exposure and CyA clearance before and 2 weeks after instituting INH prophylaxis. There was also no statistically significant correlation between INH levels and changes in CyA pharmacokinetic parameters before and after administration of INH. Even after all post-INH pharmacokinetic parameters were adjusted for INH levels, the differences in the above pre- and post-INH parameters did not reach statistical significance. Renal function during the study period remained constant and there were no episodes of CyA toxicity or acute rejection during and up to 4 weeks of INH treatment. We conclude that concomitant administration of INH and CyA is safe and is not associated with any appreciable alterations in the bioavailability of CyA. PMID- 11256237 TI - Analysis of antinociceptive effects of flurbiprofen enantiomers in a rat model of arthritic pain. AB - The potential antinociceptive effects of the S(+)- and R(-)-enantiomers of flurbiprofen (SFB and RFB, respectively) were investigated when given intravenously to rats using the pain-induced functional impairment model in the rat (PIFIR), an animal model of arthritic pain. Groups of 6 rats received either vehicle or the enantiomer in turn and antinociception was determined by evaluating the dose-response curves over time. Although SFB and RFB produced dose dependent effects with similar efficacy (SFB: 277.4 +/- 29.9 au and RFB: 293.5 +/ 20.1 au), the R(-)-enantiomer was unable to produce any antinociceptive action when assessed at the same dose ranges as SFB. It was necessary to increase the dose of RFB by 100 times to produce similar antinociception. Accordingly, S(+) flurbiprofen was 100-fold more potent (ED50 = 0.33 +/- 0.13 mg/kg) than its antipode R(-)-(ED50 = 30.0 +/- 1.7 mg/kg). SFB generated from metabolic inversion (> 1%) after i.v. dosage of RFB, as well as impurities of SFB present in RFB preparations, tend to confirm the hypothesis that the efficacy of RFB achieved at 100 mg/kg, similar to that observed with 1 mg/kg of SFB, is attributable to SFB. PMID- 11256240 TI - [Characterization and distribution of Citrobacter species in a university hospital]. AB - OBJECTIVE: [corrected] a) To identify Citrobacter strains following the conventional biochemical reaction of Brenner and col; b) to evaluate the sensitivity and specificity of the O'Hara's method compared with Brenner's method, and c) to determine the rate and distribution of the strains in the clinical isolates. MATERIAL AND METHODS: One hundred and twenty two clinical isolates, characterized as Citrobacter spp. were collected between May of 1994 and August of 1997. Clinical isolates included inpatients and outpatients from Hospital de Clinicas. Strains were identified following the methods of Brenner and O'Hara. RESULTS: Methods of Brenner identified 111 of 122 strains: C. freundii 59 of 111; C. koseri 18 of 111; C. werkmanii 15 of 111; C. braakii 9 of 111; C. youngae 6 of 111 and C. amalonaticus 4 of 111. O'Hara's methods identified 104 of 111 strains (94%). C. freundii was recovered most frequently from urine and feces (p Fisher < 0.026 and 0.039 respectively), while C. koseri was isolated from urine principally (p Fisher < 0.0372). CONCLUSIONS: The genus Citrobacter is an important opportunistic pathogen that can be identified in clinical microbiology laboratories using O'Hara's method. PMID- 11256241 TI - [Valaciclovir in the treatment of initial infection by genital herpes virus: comparative study]. AB - BASIS: To know the therapeutic efficiency in the genital herpes of two drugs: acyclovir and valaciclovir. METHODS: There were included in the study 142 patients with diagnostic of clinic first episode by genital herpes in two equal groups of 71 patients each one. The distribution in both groups was random to receive one of the following treatment standards: acyclovir 200 mg by verbal each 5 hours, during 7 days; valaciclovir: 500 mg by verbal each 12 hours during 7 days being valued objective and subjective response to the treatment. RESULTS: The prevailing symptom was the pain (45% and 46.4%), followed by the warmth or burning sensation. The most frequent lesions in both groups were blisters (39.4% and 46.4%). The analysis response to the treatment in relationship to the symptoms as well as in the lesions it could be appreciated that there are not significant differences in the patients treated in both groups (p = 0.3). The adverse effect communicated by the discussed patients were scarce and similar in both groups. CONCLUSIONS: Both drugs are suitable for the treatment of the genital herpes. The advantage observed with the valaciclovir is the dosing comfort and the facility of completing the treatment. PMID- 11256242 TI - [Administration of granulocyte colony-stimulating factor in patients infected with human immunodeficiency virus and prolonged neutropenia]. AB - BACKGROUND: Persistent neutropenia is frequent in HIV infected patients with severe immunodeficiency. G-CSF induces proliferation and differentiation of granulocyte precursors. Our objective has been to assess the response to G-CSF therapy on patients with advanced HIV disease and prolonged neutropenia. METHODS: A retrospective analysis of databases containing demographic information, analytic controls and hospitalizations related to neutropenia for patients attending our Infectious Diseases Unit from December 1, 1992 to January 30, 98. The episodes with absolute neutrophil counts lower than 1,000 x 10(6)/l at least during 7 days which descend below 500 x 10(6)/l at any moment were included. RESULTS: 36 episodes were included. 9 episodes started on treatment with G-CSF. The median duration was 9 (3-76) weeks. Hospitalization with fever related to neutropenia was significantly less frequent in episodes which received G-CSF (22.2%) than episodes without (66.7%). CONCLUSION: In this study, a significantly lower risk of hospitalization due to fever and neutropenia was associated with administration of G-CSF in patients with absolute neutrophil counts lower than 500 x 10(6)/l. PMID- 11256244 TI - [Abdominal discomfort and soft stools in a habitual consumer of rare beef]. PMID- 11256243 TI - [Air evacuation of patients with potentially transmissible infectious diseases]. PMID- 11256245 TI - [Eye involvement in AIDS-related Kaposi sarcoma]. AB - BACKGROUND: The study evaluates the incidence of ocular compromise in the Kaposi's sarcoma associated with AIDS. METHODS: We revised the clinical histories of HIV seropositives patients seen in the ophthalmology department from January, 1994 to December, 1998. All patients were examined by direct visually and dilated fundus examination with the use of either a direct or an indirect ophthalmoscope. RESULTS: In 6,552 patients, ocular Kaposi's sarcoma was diagnosed in 17 (0.25%), predominantly in male sex (88.23%). The lesions predominated in eyelids, and the inferior has been the most affected. In only one female, the ocular compromise was the first neoplastic manifestation. CONCLUSIONS: The ocular compromise in the Kaposi's sarcoma is an alternative to be considered in AIDS patients with previous or simultaneous cutaneous or visceral involvement. Due to the few clinical signs of these lesions, a thorough ocular study is recommended in these patients. PMID- 11256248 TI - [Bilateral hip necrosis, corticoids, and human immunodeficiency virus protease inhibitors]. PMID- 11256247 TI - [Bacteremia caused by Brucella sp. with negative conventional serology]. PMID- 11256246 TI - [Spinal cord lesion in a patient with human immunodeficiency virus infection]. PMID- 11256249 TI - [Failure to detect Brucella melitensis in 3 hemoculture systems]. PMID- 11256250 TI - [Urinary infection caused by Enterococcus faecalis with a mucoid morphotype]. PMID- 11256251 TI - [Multiresistant tuberculosis caused by Mycobacterium bovis and human immunodeficiency virus infection. Are there new therapeutic possibilities?]. PMID- 11256252 TI - [Pacemaker-cable endocarditis and spondylodiscitis caused by Citrobacter koseri. Conservative treatment]. PMID- 11256253 TI - [Kaposi sarcoma simulating splenic abscesses in a woman with AIDS]. PMID- 11256254 TI - [Pneumonia caused by Hafnia alvei in a patient infected with human immunodeficiency virus]. PMID- 11256255 TI - [What is the cost of a prosthesis infection?]. PMID- 11256257 TI - A national survey: transporting patients within Australian hospitals. AB - A telephone survey was conducted to describe current practices and policies of patient transport in Australian hospitals. The survey had a 94% response rate. Results showed considerable variability and ambiguity throughout the sample in both practice and policy. Findings also indicated that criteria used for transport practices were predominantly shaped by physiological and technological considerations. Factors related to human and financial resources, as well as psychological and emotional aspects of the patient's condition, received little attention. PMID- 11256256 TI - [Diarrhea caused by adenovirus and astrovirus in hospitalized immunodeficient patients]. AB - BACKGROUND: Acute or chronic diarrheal illness are common complications in immunosuppressed patients such as human immunodeficiency virus (HIV)-infected, bone marrow or solid organ transplanted patients and those with leukaemias or other immune deficiency disorders. Due to the importance of recognizing the feasible etiologies of diarrhea in order to give the proper antimicrobial chemotherapy or to avoid a misdiagnosis of rejection in the case of transplanted patients, we have investigated adenovirus and astrovirus antigen in faeces from different immunosuppressed patients. PATIENTS AND METHODS: Stool samples from 258 immunodeficient patients hospitalized at University Hospital Complex of Santiago of Compostela with acute or persistent diarrhea were collected between 1997-99 and assayed for astrovirus and adenovirus antigen. Viral antigen was detected by EIA. Other common enteric pathogens were also assayed. RESULTS: Adenovirus antigen was positive in 5 cases (2%) and astrovirus antigen in 12 cases (5%). The most commonly patients infected was those with haematologic disorders and premature infants. HIV-infected patients were positive for astrovirus antigen in 3 cases. The majority of the cases were related with intestinal bacterial diseases or other circumstances, such as Clostridium difficile infection, both associated with prolonged antimicrobial therapy. CONCLUSIONS: Astrovirus and adenovirus have to be considered as enteropathogens specially in immunocompromised hospitalized patients. An accurate diagnosis about diarrhea etiology is advisable in order to give a specific antimicrobial therapy, when it be necessary, or to avoid a misdiagnosis of rejection, in transplanted patients. PMID- 11256258 TI - Hospital outpatient and emergency services in rural Victoria. AB - Outpatient and emergency services in rural hospitals have rarely been studied. This paper analyses routinely collected data, together with data from a survey of hospitals, to provide a picture of these services in Victorian public hospitals. The larger rural hospitals provide the bulk of rural outpatients and emergency services, particularly so for medical outpatients. Cost per service varies with the size of the hospital, possibly reflecting differences in complexity. Funding policies for rural hospital outpatient and emergency services should be sufficiently flexible to take into account the differences between rural hospitals. PMID- 11256259 TI - Determinants of length of stay: implications on differential funding for rural and metropolitan hospitals. AB - This study analysed and compared the determinants of length of inpatient stay between the rural and metropolitan public hospitals. The investigation was based on the 1998/99 Western Australia patient discharge data. A Cox regression model was used due to the high proportion of patient transfers in the rural hospitals. It was found that several variables were associated with length of stay (LOS) variations within Diagnosis Related Groups (DRG). The method provides additional insights to hospital management and clinicians in assessing the risk of prolonged hospitalisation. From a state government perspective, a DRG payment adjustment strategy may be developed for different categories of admitted patient episodes. The analysis has implications on the formulation of differential funding rates between rural and metropolitan hospitals. PMID- 11256260 TI - Integration between GPs and hospitals: lessons from a division-hospital program. AB - The aim of the study reported here was to evaluate current initiatives in GP hospital integration and highlight areas where further research, development and evaluation are required. Seven pre-existing GP-hospital programs were selected and given supplementary funding to allow for more effective evaluation. These local evaluations were then incorporated into a national program on GP-hospital collaboration. We found that the seven projects made substantial progress towards their goals, and in the process highlighted important aspects of successful collaboration. The collective evaluation of DHIP identified expected benefits of collaboration for patients (improved access to services, reduced anxiety, and fewer post discharge complications), for GPs (increased involvement in acute care and in hospital decision making), and for service organisations (stronger working relationships, increased capacity, and greater efficiency). Barriers to service integration were also identified, including the different cultures of Divisions and hospitals, their lack of internal coherence and the Commonwealth-state divide. The evaluation showed that much has been achieved in building the relationships and the capacity needed for GP-hospital collaboration, and that effective models exist. The current challenge is to extend successful models across health areas and make effective collaboration part of the normal system of care. Substantial progress towards integrated care relies on a shift from a focus on systems within general practice or hospital environments to a patient centred approach. This will require general practice, hospitals, community services and consumer organisations to form long term partnerships and move beyond their currently disjointed view of acute and community care. The development of practical indicators for integrated care will support the process and facilitate shared learning across Commonwealth and state divides. PMID- 11256261 TI - Post-treatment support for patients with haematological malignancies: findings from regional, rural and remote Queensland. AB - Social support is a significant factor in the cancer patient's psychosocial wellbeing. This paper presents the findings of recent Queensland-based research that explored the experience of families returning home to the regional, rural and remote sector after a family member completed specialist treatment for a haematological malignancy and related blood disorder in a major metropolitan centre. Family and friends are the key resource persons providing support. To a large degree, this is because of the absence of alternative sources. Even support from health professionals can be problematic and for many the only source of support is the specialist centre many hundreds of kilometres away. The primary response to developing supportive services needs to be through person to person contact either via telephone or newsletter. If targeted appropriately, there is also evidence that educational programs, support groups and volunteers would be an effective medium for providing support. Educating the local health professionals (GPs and nurses) about haematological conditions is a logical extension of providing support to families. PMID- 11256262 TI - The Innovative Ward Project: promoting innovation in health service delivery. AB - An Innovative Ward Project was undertaken as part of the planning for redevelopment of the Princess Alexandra Hospital. Two inpatient units (one medical and one surgical) became pilot areas for developing, implementing and evaluating innovative approaches to service delivery. The project focused on the key areas related to structural environment, information technology and redesign of work practices. This paper provides an overview which includes the key elements utilised to foster innovation. The challenges of disseminating and adopting successful innovations beyond the Innovative Wards are discussed. PMID- 11256263 TI - Transforming Allied Health. AB - In 1995, the Division of Allied Health at the Women's and Children's Hospital Adelaide (WCH) began a process of critical review of its service delivery models and organisational structure in order better to meet the vision and values of the WCH and the needs of consumers. This paper describes the change management process. Barriers to and facilitators of change are highlighted. The outcomes of the change process are described, including the new multidisciplinary team and program-based organisational structure and culture. PMID- 11256265 TI - The future shape of primary health care in New Zealand. AB - The Minister of Health in New Zealand earlier this year released a discussion document titled "The Future Shape of Primary Health Care" which outlines some far reaching proposals for the provision of primary health care services within New Zealand. This article sets the discussion document in the context of primary health care within New Zealand by examining current arrangements for primary health care, previous arrangements and the proposals outlined in the discussion document. PMID- 11256264 TI - What impact has managerialism had on a New South Wales area health service? AB - In a perfect world, the health public sector would be completely efficient and effective. In reality, managers, policymakers, politicians, academics, public sector employees and business representatives are constantly searching for new ways to orientate the public sector towards being more cost-effective, accountable, results- and outcome-orientated, task-specific and better organised and structured. In New South Wales (NSW), this has been most apparent in endeavours to bring about a change towards the philosophy of 'new managerial thinking' or corporate management. This paper explores the hypothesis that managerialism has significantly influenced the culture of the New England Area Health Service (NEAHS) and its relationship with its staff. To test this hypothesis, between 1996-1997 a self-administered questionnaire survey form was sent to a sample of the NEAHS staff across all work sites and all levels. It is concluded that during this time, the organisation was struggling with change management issues and the successful implementation of managerialist philosophy and its elements as evidenced by staff confusion, doubt and 'cultural shock'. PMID- 11256266 TI - The delegate's dilemma: ACAT re-assessment in hostels. AB - The provision for 'ageing in place' in the Aged Care Act of 1997 has provided an opportunity for hostel facilities to broaden their scope of care for older people. Aged Care Assessment Teams (ACATs) are required to provide assessments to give approval for high or low level entry to these facilities, and to provide approval for reclassification from low to high care. However, guidelines for ACAT assessments are contradictory with respect to the Resident Classification Scale (RCS) which provides the facility funding formula, thus creating gatekeeping compared with advocacy difficulties for the ACAT. If the facility can support a claim of high care need for a resident via the RCS but the ACAT (using different and less in-depth criteria) does not agree with that claim, then the care of that resident might be compromised due to inadequate funding. Recommendations made to solve this dilemma include conferring the right of the hostel staff to reclassify residents when necessary, with the responsibility for confirmation of that classification to remain with the trained validation officers from the Commonwealth Department of Health and Family Services, not the ACAT. PMID- 11256267 TI - Faith in the 'cultural fix': limits to a planned cultural change program in a rural health service. AB - This paper, by way of a narrative on the author's participation, explains the limits to a planned cultural change program in a large rural health service. Cultural change was identified by the CEO as crucial to the success of a major restructuring of the service, and the attitudes and beliefs of the 'old guard' were considered to be constraining progress. Advocates of cultural integration contend that shared core values across an organisation can overcome such obstacles. This is a matter of faith. An application of Habermasian theory suggests that organisational leaders are drawing on traditional/religious beliefs and practices to bolster their visions and missions at a time of motivational crisis. Though a need for cultural change in some sectors of the health services is acknowledged, the particular challenges in attempting to manipulate the traditionally embedded culture and sub-cultures of the health services is highlighted. An analysis of some of the ideas and beliefs surrounding authority, deference and discipline is undertaken. It is argued that the ritualistic reinforcement of these beliefs and the reproduction of sub-cultures along material and ideal interests militate against the implementation of objectives delineated by the CEO. While cultural analysis has revealed the irrational face of organisations and can bring to conscious awareness the taken-for-granted beliefs which inform behaviour, the cultural integrationists have a further agenda. They aim to manipulate organisational culture to subtly control employees' beliefs and hence behaviour. Cultural control is a covert form of top down authority that can be just as directive and centralizing as bureaucratic control. The author also maintains that cultural change programs alone cannot fix a problem that arose in the macro-economic sphere: a chronic lack of resources ever since the state responded to the economic crisis by cutting funds to health and welfare services. PMID- 11256268 TI - The National Demonstration Hospitals Program. AB - Fifty-five public hospitals in all Australian States and Territories participated in the first two phases of National Demonstration Hospitals Program (NDHP). The program was established in 1994 as part of a commitment by the then Department of Health and Family Services to reduce waiting times and improve health outcomes for patients. The program uses a collaborative approach to assist public hospitals to improve service delivery and patient care outcomes. Key results from Phases 1 and 2 of the NDHP have confirmed that identification of industry best practice, collaboration, knowledge sharing and innovation are key elements required to achieve positive health care reforms. PMID- 11256269 TI - Toward gender balance in the Australian medical workforce: some planning implications. AB - Men and women doctors participate differently in the workforce. As the proportion of women in the workforce increases, gender-based differences in workforce participation are raising important planning questions. For example, how will differences in hours worked per week impact on the number of trainee doctors required to meet future demand, why do some disciplines attract more women, what will be the impact on the practice of medicine if these trends continue and how does the training environment impact on variation in career decision. In this paper we summarise and discuss the findings of recent research undertaken by the Australian Medical Workforce Advisory Committee and outline some public policy responses. PMID- 11256270 TI - Would employment-based health savings accounts help Medicare? PMID- 11256271 TI - Innovations in medical education to meet workforce challenges. AB - The winds of change world-wide have swept medical education in the last fifteen years. Today, Australia's medical students are older and drawn from more diverse socio-economic, ethnic and geographic backgrounds than twenty years ago, and there is now an equal mix of men and women in medical school. Admission policies have been rewritten to broaden access with a range of entry options now available including direct entry from high school and graduate entry following a first degree. Curricula have been revised and modes of learning transformed. This paper describes these changes and discusses the implications for medical schools and for planning the future workforce. PMID- 11256272 TI - The Australian health workforce: facts and futures. AB - The quality of care received by a patient or consumer critically depends on the knowledge, skills and attitudes of the health workforce; the structure and functioning of the health workforce is critical to the structure and functioning of the health system overall. To a very large extent, diagnosis and treatment decisions call on the training and experience of the health professional. The quality of the interaction between a patient or consumer depends on the interpersonal and technical skills of health professionals. In a sense, health workers are important to defining the very nature of health care services. The importance of the health workforce is further highlighted by the fact that, as is typical of most service industries, labour accounts for a large proportion of health costs (around 80%). This paper provides an overview of the size and composition of the health workforce in Australia. It then reviews three segments of the workforce in more detail (medical, nursing and other health professionals) and reviews contemporary policy issues affecting those groups. PMID- 11256273 TI - Contemporary issues in the workforce and education of Australian midwives. AB - This paper, which is based on the preliminary findings of the Australian Midwifery Action Project (AMAP), outlines the issues around the midwifery labour force and education in Australia. One of the most alarming features is the lack of comprehensive data on midwives. Where data is available it demonstrates the shortage of midwives and the lack of consistency in educational programs for midwives within states and nationally. It is difficult to form a national picture with published sources of data because there are differences in definition and a lack of relevant information. Strategies for educational reform are discussed in relation to improving the supply and preparation of midwives. PMID- 11256274 TI - Medical workforce planning in Australia. AB - The Australian Medical Workforce Advisory Committee (AMWAC) was established by the Australian Health Ministers' Advisory Council (AHMAC) in 1995 to provide information and analysis to AHMAC and the profession about the medical workforce to inform the policy process. This article provides a brief history of the events leading to the formation of AMWAC and of the work of this committee, particularly its approach to medical workforce planning and the outcomes of some of its research. The paper concludes that the forces leading to and maintaining workforce geographic and structural maldistribution are better understood as a result of AMWAC studies and the work of other stakeholders. Further research is required to improve understanding of the hospital medical workforce and the factors influencing the career decisions of young doctors and to monitor the impact of strategies to improve workforce distribution. PMID- 11256275 TI - Absenteeism and the impact of a 38-hour week, rostered day off option. AB - We undertook a comparative analysis of nurses working in two consecutive years: one in a 40-hour standard working week and the other in a 38-hour week with a rostered day off per month, in order to determine whether there was any effect on absenteeism. We found that total absenteeism between the two years fell significantly from 4.58% to 4.36% (chi 2 = 5.09, P = 0.024). Sick leave decreased but not to a significant degree. We conclude that the change to the 38-hour week and 19-day month (rostered day off) arrangements led to a significant reduction in overall absenteeism but not in sick leave. However, the cost in implementing a 19-day month is far in excess of any savings made through absenteeism reductions. PMID- 11256276 TI - Australian Aboriginal trainee health service management program: a new initiative. AB - This paper explores the development, implementation and evaluation of the Australian Aboriginal trainee health service management program in New South Wales. In 1997, the two-year pilot program commenced with ten trainees. The program consisted of a combination of work-based placements, formal university education and Australian College of Health Service Executives (ACHSE) professional development sessions. The program has allowed trainees to gain professional skills and knowledge and broader work experience, in order to increase their employment opportunities throughout the Australian health care system. PMID- 11256277 TI - Effect of short-term treatment with bumetanide, quinapril and low-sodium diet on dogs with moderate congestive heart failure. AB - OBJECTIVE: To evaluate the effect of bumetanide, quinapril and a low-sodium diet on clinical response tolerance and side-effects on dogs with moderate congestive heart failure. DESIGN: A prospective clinical study, using 32 client-owned dogs with naturally occurring disease. PROCEDURE: Thirty-two dogs diagnosed with congestive heart failure (International Small Animal Cardiac Health Council stage II) due to mitral valve disease were included. During 4 weeks, patients received 0.5 mg/kg quinapril (Ectren, Menarini), 0.1 mg/kg bumetanide (Fordiuran, Boehringer Ingelheim) and a low sodium diet (CNM-CV, Purina) was fed. All dogs were examined weekly and results were treated statistically. RESULTS: The treatment was safe, effective and well-tolerated and no major side-effects were observed. There were no significant changes in measured haematological and biochemical variables, including serum electrolyte concentrations and urinary fractional excretion of electrolytes. CONCLUSION: This study suggests that the combined treatment with bumetanide, quinapril and low-sodium diet for controlling moderate CHF due to mitral insufficiency in dogs is simple, easy-to-administer and effective in controlling clinical signs and prompting improvement even after short-term treatment. PMID- 11256278 TI - Progressive neurological signs associated with systemic mastocytosis in a dog. AB - A 9-year-old dog was presented with nonregenerative anaemia and severe thrombocytopenia, diarrhoea, spinal hyperalgesia and progressive hindlimb paresis. A moderately well differentiated cutaneous mast cell tumour (MCT) was removed from the skin of the right elbow along with the enlarged right prescapular lymph node. Due to deterioration of the dog's neurological condition, euthanasia was performed. On necropsy examination, haemorrhage and accumulations of poorly differentiated mast cells were found in the lumbosacral region and cauda equina. This article describes an unusual presentation of systemic mastocytosis and the previously unreported finding of metastasis of mast cells to the spinal cord. PMID- 11256279 TI - Evaluation of high-speed treadmill videoendoscopy for diagnosis of upper respiratory tract dysfunction in horses. AB - OBJECTIVE: To evaluate high-speed treadmill videoendoscopy as a diagnostic technique and document the abnormalities found in Australian horses referred for poor performance associated with abnormal upper respiratory tract noise but where a definitive diagnosis could not be made at rest. DESIGN: A retrospective clinical study using client-owned horses. PROCEDURE: The clinical records and videorecordings of all horses referred to the University of Sydney for poor performance associated with abnormal upper respiratory tract noise during a 13 month period were examined. Only horses with a normal physical examination including absence of lameness, and where a definitive diagnosis of the cause of the abnormal upper respiratory tract noise could not be made from the resting videoendoscopic examination were included in the study. The age, gender, breed of horse and the purpose for which the horse was used were ascertained from the record and videorecordings were reviewed by the authors and any abnormalities noted. RESULTS: There were 37 horses included in the study. An upper airway abnormality was identified during high-speed treadmill videoendoscopy in 73% of horses. One abnormality was identified in 22 horses, 2 abnormalities in 4 horses and 3 abnormalities in 1 horse. Abnormalities identified included laryngeal hemiparesis (n = 15), axial deviation of the aryepiglottic folds (n = 10), pharyngeal collapse (n = 3), dorsal displacement of the soft palate (n = 2), epiglottic collapse (n = 1), axial deviation of the vocal cord (n = 1) and laryngeal collapse (n = 1). CONCLUSIONS: The results of this study are similar to reports from overseas and suggest high-speed treadmill videoendoscopy appears to be a useful technique to diagnose the cause of upper airway dysfunction in Australian horses referred specifically for poor performance associated with abnormal upper respiratory noise. However, a diagnosis will not be made in all horses undergoing this procedure. There were five horses with two or three abnormalities none of which were apparent at rest. This would suggest that in all horses making abnormal upper respiratory noise associated with poor performance, even where an abnormality is identified at rest, high-speed treadmill videoendoscopy should be performed for a complete diagnosis. PMID- 11256280 TI - Evaluation of an insulin-zinc suspension for control of naturally occurring diabetes mellitus in dogs. PMID- 11256281 TI - Gold standards and fool's gold. PMID- 11256282 TI - Canine coronavirus in Australian dogs. AB - OBJECTIVE: To estimate the frequency of serum antibodies (IgG and IgM) to canine coronavirus (CCV) in the Australian dog population and evaluate the role of CCV as a causative agent of gastroenteritis. DESIGN: A serological survey of antibodies to CCV among different dog populations. PROCEDURE: The development and characterisation of an indirect ELISA for the detection of antibodies (IgG and IgM) to CCV was undertaken. Sera collected from both diarrhoeal and non diarrhoeal dogs from various populations throughout Australia were tested for these antibodies to CCV. RESULTS: Serum samples (1396) collected from 1984 to 1998 were tested for the presence of IgG antibodies to CCV. Samples were divided into two categories on the basis of the number of dogs housed together. The groups were either an open population containing dogs housed as groups of three or less, or kennel populations. Sera from 15.8% of the open population and 40.8% of kennelled dogs were positive for CCV antibodies. The prevalence of antibodies varied from zero to 76% in kennelled dogs. About 23% of 128 dogs positive for IgG antibodies to CCV were also positive for IgM antibodies to CCV, indicating recent CCV infection. Of those dogs that were presented with clinical signs of gastroenteritis such as diarrhoea and vomiting (n = 29), 85% were positive in the IgM ELISA and 85.7% in the IgG ELISA for antibodies to CCV. In comparison, for those dogs presented without any history of gastroenteritis only 15% were positive for IgM and 30% positive for IgG. CONCLUSION: Serological evidence indicates that infection with CCV in dogs is widespread throughout the Australian mainland. The prevalence of antibodies varies greatly among different populations, with an average of 40.8% positive in kennelled populations and 15.8% in the open population. PMID- 11256283 TI - Efficacy of macrocyclic lactones for the control of larvae of the Old World screw worm fly (Chrysomya bezziana). AB - OBJECTIVE: To assess the efficacy of four macrocyclic lactones for the control of larvae of the Old World Screw-worm Fly (OWS), Chrysomya bezziana, and to examine the effects of excreted residues on the dung fauna. ANIMALS: 100 heifers were divided into five groups of 20 animals. One group remained untreated, whereas the other groups were treated respectively with pour-on formulations of moxidectin, eprinomectin or doramectin, or a sustained-release bolus of ivermectin. PROCEDURES: At intervals of 1 to 15 weeks after treatment, five cattle from each group were challenged with newly-laid eggs of OWS. The efficacy of each treatment was determined 48 h later by comparing the number of myiases in the treated and untreated groups. Abundance of fly larvae in naturally-voided dung pads and the survival of a species of dung beetle, Onthophagus sagittarius, were used to assess the effects of drug residues on the dung fauna. RESULTS AND CONCLUSIONS: Moxidectin showed no activity against larvae of OWS during the first 14 days after treatment. Eprinomectin provided protection for 3 days after dosing, but failed at days 7 and 14, whereas doramectin was effective at day 7, but not at days 14 or 21. In contrast, no myiases were established on bolus-treated cattle from 14 to 102 days after treatment. Faecal residues of moxidectin had no effect on the survival of larvae of dung-feeding flies, whereas those of eprinomectin and doramectin reduced survival for 1 to 2 weeks. Dung voided by bolus-treated cattle inhibited fly breeding and had adverse effects on the development and survival of O sagittarius for up to 15 weeks after treatment. PMID- 11256284 TI - The development and survival of three species of coprophagous insect after feeding on the faeces of sheep treated with controlled-release formulations of ivermectin or albendazole. AB - OBJECTIVE: To assess the toxicity from residues of controlled-release formulations of ivermectin and albendazole to insects that feed on sheep faeces. ANIMALS: In two consecutive years, groups of sheep were treated with controlled release capsules of ivermectin or albendazole. Untreated sheep were used as controls. PROCEDURES: Larvae of the bush fly, Musca vetustissima, and adults and larvae of the dung beetles, Onthophagus taurus and Euoniticellus fulvus were fed on faeces collected at intervals after drug treatment. In assays using beetles, treatment effects were assessed by comparing numbers of eggs laid, survival of juveniles and survival of mature and immature adults. Survival at time of pupariation was used in assays on flies. RESULTS AND CONCLUSIONS: Faeces from sheep treated with albendazole had no detectable effects on breeding by either flies or beetles. In contrast, faeces voided by sheep treated with controlled release capsules of ivermectin (CRI) precluded successful breeding by each of the species tested. No fly larvae and almost no beetle larvae survived in faeces collected up to 39 days after capsule administration. Newly-emerged O taurus also suffered significant mortality whereas those that survived underwent delayed sexual maturation. Ivermectin residues had no effect on the survival of sexually mature beetles, but reduced the fecundity of O taurus. A model simulating the effects of drug residues on dung beetle populations indicates that CRIs have the potential to cause substantial declines in beetle numbers, particularly if treatment coincides with spring emergence. PMID- 11256285 TI - Gastroenteritis associated with Helicobacter-like organisms and rotavirus in a reindeer (Rangifer tarandus). PMID- 11256286 TI - Communication of risk. PMID- 11256287 TI - Heartworm treatment--NRA decision opposed. PMID- 11256289 TI - Exotic animal diseases bulletin. February 2001 No. 76. PMID- 11256288 TI - SA newspaper fails to allow truth to be heard. PMID- 11256290 TI - Caged hens debate: silence reigns! PMID- 11256291 TI - Ashamed of AVA's caged hens policy. PMID- 11256292 TI - AVA and ACT showing the way to UK kennel clubs. PMID- 11256293 TI - A breeder defends docking dogs' tails. PMID- 11256294 TI - 'Research' redefined by letter writer. PMID- 11256295 TI - Animal welfare officers: should they be vets? PMID- 11256296 TI - NSW division representative on Companion Animal Advisory Board cautions over legal situation. PMID- 11256297 TI - Advertiser article wrong on docking. PMID- 11256298 TI - Concurrent Cryptosporidium and parvovirus infections in a puppy. AB - Cryptosporidium parvum, an intestinal coccidian parasite, was isolated from faeces and intestinal biopsies of a 9-week-old puppy with acute parvoviral gastroenteritis. Gene sequence analysis identified a Cryptosporidium genotype not previously recorded in Australia. The puppy recovered after treatment with crystalloid fluids, synthetic and natural colloids and jejunostomy tube feeding. PMID- 11256299 TI - Cannibals to cows: the path of a deadly disease. PMID- 11256300 TI - The last word. Our tired, our poor, our kids. PMID- 11256301 TI - [Pre-reduced TGY-V medium]. AB - Anaerobic bacteria need, for its multiplication, a reduce middle devoid of oxygen, who contains growth factors. We have prepared the prereduce middle TGY-V according to the recommendations of M. SEBALD [1]. This middle has been tested for its ability to permit the growth of the most exigent anaerobic bacteria. It could be used as a middle of transport and culture for any monomicrobial sampling that can contain anaerobic bacteria. PMID- 11256302 TI - [Production of monoclonal antibodies specific for the ABO blood group and rhesus D antigens]. AB - We report, in this work, the techniques to obtain four monoclonal antibodies specific of erythrocytes antigens. Three of this antibodies, react with the ABO Blood Groupe System (A,B and AB), are produced by three mouse hybridomas (M18 2F11, M18-4F6 et M19-45D6), obtained by fusion of Sp2/0 mouse myeloma cell with spleen cell of balb/c mice immunized with human red blood cells and selected on selectif medium (Hypoxanthin, Aminopterin, Thymidin) (HAT) and cloned by four limiting dilutions. Where as the fourth, it is an human monoclonal antibodies against Rhesus (D) produced by heterohybridomas, realized by fusion of X63 mouse myeloma cell with human B lymphocytes, from actively immunized persons by D antigen, that are purified and transformed by Epstein-Barr virus (EBV), and selected on selectif medium (HAT), presence of ouabain and then cloned by four limiting dilutions. The specificity of the antibodies produced, has been determined by direct hemagglutinin for the mouse monoclonal antibodies and artificial for the human anti-D. The determination of isotype the heavy and light chains is released by the technique immunoenzymatique (ELISA) and immunofixation has shown that mouse monoclonal antibodies belong to class IgM kappa, and human monoclonal antibodies anti-D belong to class IgG lambda. PMID- 11256304 TI - [Contribution of immunofixation and laser nephelometry in the diagnosis of monoclonal immunoglobulinopathy]. AB - Immunological diagnosis of principal's monoclonal gammapathies [Waldenstrom's macroglobulinemia and multiple myeloma], used classically electrophoresis and immunoelectrophoresis analysis (AIE) for detection of monoclonal components. Although its specificity and sensibility, the last method stays a long analysis with critical interpreting from time to time. The present study reports results obtained with 100 sera from subjects with these diseases when using immunofixation and nephelometry-laser witch tests amounts of IgG, IgA and IgM and kappa/lambda--ratio both methods together, which are simpler and more faster than AIE. PMID- 11256303 TI - [Sensitivity to beta lactams and macrolides of Streptococcus pyogenes isolated at the services from 1994 to 1998]. AB - The identification and antibiotic sensibility of 188 strains of streptocoques pyogenes had been performed from 1994 to March 1998. We were interested on active antibiotics against streptocoques pyogenes: beta lactamins (penicillin ampicillin) and macrolids (erythromycin). The aim of this study is to reveal resistances and calculate percentages of resistant strains to these antibiotics. The WHO recommend to treat streptococcal sore throat with penicillin, and with erythromycin when there is a proved inhibility to the penicillin. No penicillin resistant streptocoques pyogenes has been found in the world, that concords with our study: all strains (188 sp.) are penicillin sensitive. Macrolids resistant strains of streptocoques pyogenes has been described by a lot of countries with different distributions. In our study global percentage of resistant strains to erythromycin is 24% and 29% for throat specimen. PMID- 11256305 TI - [Iso-enzyme study of Echinococcus granulosus protoscoleces in Algeria]. AB - The purpose of this study is to identity the strains of Echinococcus granulosus the causative agent of unilocular hydatid in Algeria--a high endemic area. For this, the authors established a simple and reproductible electrophoretic techniques for iso enzyme analysis of protoscoleces. The enzymatic extracts of protoscoleces from various hosts and localisations of hidatic cystic analysed by these electrophoretic techniques showed variable electrophoretic profils witness the existence of various strains. PMID- 11256306 TI - [Leishmaniasis diagnosis: role of a simple medium for the culture of Leishmania]. AB - Leishmaniasis, Zoonoses or Anthroponoses, according to the focus, know an extension through the world and Algeria counts unfortunately more among countries who where touched. Parallelement to this extension the eco-epidemiology of leishmaniasis knex some important modifications and that. We can't surround theme if we don't dispose of a simple culture medium permeting the shoot of all species to leishmania. PMID- 11256307 TI - [Malaria chemoprophylaxis]. AB - The prevalence and the severity of malaria has increased since pesticide resistance against the Anopheles vector and amino 4 quinolone resistance against the plasmodial parasite become widespread. Chemoprophylaxis is based on the use of five drugs alone or in combination according to local resistance patterns and the lasting of sojourn on endemic area: Chloroquine, Proguanil, Chloroquine and Proguanil, Mefloquine, Doxycycline. It must be combined with preventive recommendation such the use of impregnated nets. PMID- 11256308 TI - [Validation of DNA tools in the identification of Leishmania sp. parasites in Algeria]. AB - The present study aimed at homogenizing the use of DNA tools for Leishmania parasite characterization in two endemic countries, Algeria and Tunisia. Two genomic DNA probes, pDK10 and pDK20, previously developped in Tunisia, were here applied to a collection of 41 isolates obtained from Algerian patients having cutaneous or visceral leishmaniases. These DNA tools allowed to discriminate among and to identify causal agents of cutaneous leishmaniasis, L. infantum and L. major. Apart from the pDK20--hybridization pattern obtained usually for the species L. infantum, new hybridization patterns were identified for isolates obtained from both visceral and cutaneous leishmaniases patients. Use of DNA probes in complement to isoenzyme typing offers interesting propects for a better description of transmission cycles. PMID- 11256309 TI - [Cryptococcosis. Report of 4 cases diagnosed at the El-Hadi FLICI hospital]. AB - The cryptococcosis is an opportunist infection, frequent during Acquired Immunodeficiency Syndrom (A.I.D.S.). It increases in the same way as the increase of immunodepression factors. This infection is due essentially to Cryptococcus neoformans. This paper deals with the analysis of four (04) cases of cryptococcosis admitted in El-Hadi FLICI hospital in Algiers from November 1997 to February 1998. Among these cases we have taken note of two patients struck doxn by A.I.D.S., one of them have undergone a kidney transplant, whereas the other suffered from sarcoidose and pulmonary tuberculosis. The diagnostic of cryptococcosis have been based essentially on C.S.E. examination. PMID- 11256310 TI - [Pneumocystis carinii pneumopathy in patients with AIDS. The first 3 cases reported in Algeria and review of the literature]. AB - Pneumocystis carinii pneumonia (PCP) is a severe and dangerous infection which afflicts patients with immune deficiency, particularly those with AIDS. This pathology isn't well known in Algeria. The aim of the present study is a contribution to make this disease more known in the algerian AIDS patients. It has focused on the research and the identification of P. carinii in the expectorations and in the bronchalveolar lavage (BAL) fluids of 14 patients with respiratory troubles: 13 were AIDS cases and one with an iatrogenic immune deficiency. 3 of these patients were women and the rest were men. Their average age was 35. PMID- 11256311 TI - [Immunocapture-hemagglutination technique]. AB - Virus specific IgM antibodies are very useful for the diagnosis of primer infection by the rubella and parotidis viruses. ELISA is the method usually used to detect IgM antibodies. The reactives and in some laboratories the apparatus are not always available. We carried out a serological method based on the immunocapture inhibition of hemagglutination of theses viruses by positive sera. 39 sera and 80 sera collected from patients and healthy population have been respectively studied to detect antirubella and antiparotidis IgM. The test appeared as sensitive and specific as the immunocapture IgM ELISA. PMID- 11256312 TI - [Congenital rubella resulting from maternal reinfection]. AB - Almost all the rubeolic reinfections are not apparent and are without any risk for the fetus. The news born's case (2nd in children) which is described here, presents a bilateral cataract. We have found positive IgA, IgG and IgM and an antibodies avidity of 70%. Concerning the mother, there are no IgM. The IgM and IgA are positive. The avidity of the antibodies is upper than 90%. During her first pregancy, the mother was subjected to the search of the antibodies, the results are positive (title = 60 ul/ml) and during her second pregnancy, the mother had rubeollic contact with her eldest son. Our conclusion is that it is a congenital rubella resulting from a reinfection. PMID- 11256313 TI - [Introduction to a study of specific antibodies (IgG and IgM) detection with ELISA in the diagnosis of poliomyelitis]. AB - Samples of single sera collected from 38 patients with different clinical diagnosis were studied in order to perform ELISA techniques with the purpose of detecting poliomyelitis IgG and IgM antibodies. The results were compared through antibody titration by neutralization test. 21 pairs of sera from infants suffering from acute flaccid paralysis were studied by ELISA-IgM, ELISA-IgG and neutralization test. Stool samples were collected from 20 of the latter patient. Wild poliovirus type 1 was isolated in 8 cases. ELISA-IgM technique was positive in 14 cases. The true positive poliomyelitis diagnosis was based on the persistence of flaccid paralysis 60 days after the onset and on wild poliovirus isolation with significant increase in antibody level. 16 cases were classified as poliomyelitis, 2 cases as non poliomyelitic paralysis and 3 cases as undetermined. 16 out of the 18 well established diagnosis were in agreement (88.8%) with the detection or not of IgM antibodies by ELISA. The specificity of these IgM ELISA antibodies was examined by studying 11 cases of lymphocytic meningitis. Cross reaction in serological responses between polioviruses and coxsackieviruses was observed. These cross reactions should be evaluated by studying a greater number of cases. The poliovirus ELISA-IgM is a sensitive, economical and rapid method to be used in poliomyelitis diagnosis to complete the neutralizing test and virus isolation. PMID- 11256314 TI - Production of immuns-serums for serotyping of streptococcus pyogene: quality control and epidemiological application. PMID- 11256315 TI - [Use of toxic fraction isolated from Algerian Androctonus australis hector scorpion venom for the assessment of anti-venom serum]. AB - Today, serotherapy is the only specific treatment used against the scorpionique envenomation. This treatment was associated with other drugs as a symptomatic and supportive care. The anti-venom against scorpion stings is still prepared using the conventional manufacturing production. To evaluate the efficacy of the anti venom, different protocols are used: The immunization of animals (rabbit and horse) with a whole toxic fraction prepared by molecular filtration was the first step to improve the anti-venom. Some observations also show the need for the quality control of this preparation of anti-venom. In this study, we describe the use the toxic fraction isolated from Androctonus australis Hector in the new protocol of anti-venom production. The quality control of the product is evaluated by using a combination of experimental immunological assay systems (ELISA, Hemaglutination, in vivo assessment of the neutralization of the lethal effect of the venom). PMID- 11256316 TI - [Genetic study of the autosomal recessive form of Charcot-Marie-Tooth in an Algerian family]. AB - Charcot-Marie-Tooth disease (CMT) is a hereditary neuropathy characterized by muscular atrophy and progressive sensitive alterations that affect limbs. The CMT is one of the most heterogenous diseases, clinically as well as genetically. At least twelve loci are responsible for the CMT phenotype, four of them for the autosomal recessive form. The aim of our work was to determinate the implication/exclusion of these four loci in an Algerian family by linkage analysis using microsatellites markers. We have tested the four loci on 8q13-21.1 (CMT4A), 11q23 (CMT4B), 5q23-33 (CMT4C) 8q24 (CMTAR). The haplotype reconstruction allowed us to exclude all the loci in this family, suggesting that the locus (gene) responsible for this form of CMT is localized elsewhere in the genome, thus providing an other observation of the great heterogeneity of the CMT, particularly autosomal recessive. PMID- 11256317 TI - [Blood determination of benzathine-penicillin used in acute joint rheumatism prophylaxis]. AB - Our actual work studies the effectiveness in vivo of the Benzathin penicillin that is realized on 88 subjects suffering from a stable rheumatic fever. It has shown that: The first hours after an intramuscular injection, the benzathin penicillin is found at an efficient concentration superior to 0.02 ug/ml at the level of the blood. The highest dose in the blood is obtained the first 24 hours. The amount of antibiotic at the level of the blood is very efficient during 4 weeks. PMID- 11256318 TI - [Refinement and study of the performance of a "Mycoplasma UG" kit for the research on urogenital Mycoplasmas]. AB - The pathogenic role of genital Mycoplasmas is no more disputably. The diagnosis is based on the culture which may present some technical difficulties. The present study consist in a perfecting of a miniaturised system called "Mycoplasma UG" for the research, identification and titration of genital Mycoplasmas. The performance of the kit is studied with an important number of different types samples by comparison with the commercial kit "Mycoplasma Duo" and the classic reference method. The results obtained are satisfactory and constitute an important element in favour of a more advanced validation before the kit's trading. PMID- 11256319 TI - [In vitro activity of trovafloxacin (CP-99,219) against 434 bacterial strains: comparative study with other fluoroquinolones]. AB - We have evaluated the activity of a new fluoroquinolone, trovafloxacin (CP 99,219), against 434 strains; 24 of them are resistant to ciprofloxacin. Against gram-negative bacteria, trovafloxacin has the same activity as the others fluoroquinolones. Against gram-positive bacteria, this antibiotic agent is very active. (MIC = 0.016 to 0.5 microgram/ml); Streptococcus is the most sensitive species. The ciprofloxacin resistant strains are also resistant to trovafloxacin. PMID- 11256320 TI - [In vitro study of antagonistic activity of bifidobacteria against Campylobacter and Escherichia coli causing gastroenteritis in children]. AB - Bifidobacteria represents the major constituents of the human intestinal microflora. It's a non mobile Gram (+) bacteria, which interest almost of the research about the relationships between the intestinal flora and human health. Bifidobacteria has an inhibition activity on the growth of the Campylobacter and E. coli G.E.I., in case of children gastro-intestinal infections by mixed cultivation in reconstituted infant milk adapted for the first age. This activity is the result of a group of mechanisms acting at the same time, it's probably the following: Competition of the nutrients indispensable for bacterial growth, action of the produced acid do not favorite the growth of the active pathogens germs (bacteriocine) with acid pH. PMID- 11256321 TI - [Study of antigenic groups among the different types of Clostridium perfringens]. AB - A comparative study of somatic antigens of different types of Clostridium perfringens with indirect immunofluorescence method was undertaking with 20 locals and foreigner strains in order to choose the valences that are better adapted to compose the Enterotoxemia vaccine. This study has shown a complete somatic antigens homology with A, B types and partial between D type and A, B types, moreover a complete heterology between C and D types. PMID- 11256322 TI - [Antibiotic sensitivity of Bacteroides fragilis group in Algeria]. AB - The antimicrobial resistance evolution of the Bacteroides fragilis group is the subject of international survey. Eighty-six clinical isolates collected in anaerobic service of I.P. A were tested for susceptibility to twelve antimicrobial agents. Chloramphenicol, metronidazole and imipenem proved to be the most active agents. After these agents, amoxicillin-clavulanic acid, carbenicillin and piperacillin were the most effective agents tested with respectively 15%, 17% and 17% of resistant isolates. Clindamicin and cefotaxin were active from only 70% and 65% of clinical isolates, and 71% of them were found resistant to cefotaxin with minimal inhibitory concentration above 32 ug/ml. beta Lactamasic profile determination according to Rolfe and Finegold modified method allowed to show five different beta lactamase types. The isoelectric points (pI) vary between 4.3 and 7.5 according to the enzymatic extracts of the clinical isolates. No transfer and no plasmid were observed with respectively imipenem and metronidazole resistant isolates. But, transconjugants were obtained with TRCcR isolate. PMID- 11256323 TI - [The Animal Health and Welfare Law: the veterinary statement as evidence and the role of veterinarians in legal procedures]. AB - Every veterinarian can be confronted with the request from a law enforcement agency to conduct a veterinary examination based on the Animal Health and Welfare Act and to produce a written report of that examination. This Veterinary Statement can be used in court. The Veterinary Statement is an important piece of evidence, so it should be formulated with care. The veterinarian in charge must be aware of the requirements that the Veterinary Statement must meet and the legal procedures that must be followed. PMID- 11256324 TI - [Shar-Pei dogs with recurring lameness]. PMID- 11256325 TI - [Is the carrying out of educational fetotomy in the living animal by a student ethical?]. PMID- 11256326 TI - [Continuation of discussion on educational fetotomy]. PMID- 11256327 TI - [Immunosterilization and integrity]. PMID- 11256328 TI - [Keep the secret or speak out?]. PMID- 11256330 TI - [Malignant melanoma: epidemiological data from the Dusseldorf area]. AB - Malignant melanoma represents only 1.5-2.5% of malignant neoplasms in Europe. Since 1990 the clinical data of melanoma patients are registered at the clinic for dermatology university Dusseldorf. 925 patients were treated between 1990 1996 (not including in situ melanomas, extracutaneous melanomas and multiple melanomas). In 1996 the incidence was 15 cases/100,000 habitants. A worldwide doubling of the morbidity is expected every ten years especially in the caucasian population. PMID- 11256329 TI - [What is your diagnosis? Secondary chondrocalcinosis crisis in primary hyperparathyroidism]. PMID- 11256331 TI - [Photocarcinogenesis]. AB - Sunlight-induced skin cancer is the most frequent cancer. Ultraviolet-B (UVB) (290-315 mm) and UVA (320-400 mm) radiation can induce DNA damage with resulting epithelial squamous cell carcinoma and melanoma by causing mutations and immunosuppressive effects that presumably contribute to photocarcinogenesis. The efficacy of photo- and photochemotherapeutic modalities is thought to result, at least in part, from the induction of immunomodulatory effects. In particular, UV radiation has been shown to affect (i) the production of soluble mediators, (ii) the expression of cell-surface receptors and (iii) to induce apoptosis in pathogenetically relevant cells. UVB radiation-induced immunomodulatory effects are limited to the epidermis, whereas UVA radiation-affects both epidermal and dermal cell populations. UVB and UVA radiation can exert essentially identical immunomodulatory effects, which result, however, from different photobiological mechanisms. UVB radiation-induced cyclobutane pyrimidine dimers within the DNA of epidermal cells are detrimental to human health. Photolyase-induced dimer repair completely prevented these UVB radiation-induced immunosuppressive effects as well as erythema and sunburn-cell formation. The Xeroderma pigmentosum (XP) is a rare syndrome of sensitivity to UV due to an inherited defect in nucleotide excision repair or daughter strand repair. Ionising radiation sensitivity is not part of the recognised syndrome. Extreme caution is advised before treating XP patients with radiotherapy. Determining the complementation group and radiosensitivity prior to treatment is recommended. PMID- 11256332 TI - [Adjuvant therapy of malignant melanoma]. AB - Malignant melanoma is resected with curative intent in 80-85% of the patients. In case of tumor thickness according to Breslow of > 4 mm and metastatic regional lymph nodes a high relapse rate and mortality of 50-90% is observed. Single agent chemotherapy with DTIC is effective as combination regimen, however does not significantly improve the relapse-free interval respectively the overall survival or quality of life. Several attempts with other adjuvant treatment modalities (e.g. BCG immunotherapy) are not convincing as well. The best results of adjuvant treatment in high-risk resected melanoma were published in 1996 by Kirkwood, who used interferon-alfa 2b. He could demonstrate a prolonged relapse free and overall survival. It does represent the current standard adjuvant treatment. However, the costs and toxicities of IFN are barriers to its widespread use. PMID- 11256333 TI - [Surgical therapy of malignant melanoma of the skin]. AB - Operative treatment is the therapy of choice in malignant melanoma under curative as well as palliative considerations. Today more conservative surgical strategies are preferred, however. Thus, the safety margins for the excision of the primary tumor have been significantly reduced. With the introduction of the minimally invasive sentinel node biopsy the elective lymph node dissection, a high morbidity intervention, has become obsolete. Sentinel node biopsy should be performed in all melanomas thicker than 0.75 mm, as the percentage of lymph nodes harvesting micrometastases increases in correspondence to the clinical tumor stage up to 75%. PMID- 11256334 TI - [Clinical problem solving]. PMID- 11256335 TI - The foreperiod effect revisited: conditioning as a basis for nonspecific preparation. AB - The foreperiod (FP) is the interval between a warning stimulus and the imperative stimulus. It is a classical finding that both the duration and the intertrial variability of FP considerably affects response time. These effects are invariably attributed to the participant's state of nonspecific preparation at the moment the imperative stimulus is presented. In this article, we examined a proposal by Los, S. A. (1996) [On the origin of mixing costs: exploring information processing in pure and mixed blocks of trials. Acta Psychologica, 94, 145-188] that the real-time development of nonspecific preparation during FP relies on the same principle as trace conditioning. To this end, we adjusted the formal conditioning model developed by Machado, A. (1997) [Learning the temporal dynamics of behavior. Psychological Review, 104 (2), 241-265], and fitted this model to a representative data set we obtained from nine participants. Although the model accounted for only a moderate proportion of the variance, it accurately reproduced several key features of the data. We therefore concluded that the model is a promising first step toward a theory of nonspecific preparation. PMID- 11256336 TI - Separate modifiability, mental modules, and the use of pure and composite measures to reveal them. AB - How can we divide a complex mental process into meaningful parts? In this paper, I explore an approach in which processes are divided into parts that are modular in the sense of being separately modifiable. Evidence for separate modifiability is provided by an instance of selective influence: two factors F and G (usually experimental manipulations) such that part A is influenced by F but invariant with respect to G, while part B is influenced by G but invariant with respect to F. Such evidence also indicates that the modules are functionally distinct. If we have pure measures MA and MB, each of which reflects only one of the parts, we need to show that MA is influenced by F but not G, while MB is influenced by G but not F. If we have only a composite measure MAB of the entire process, we usually also need to confirm a combination rule for how the parts contribute to MAB. I present a taxonomy of separate-modifiability methods, discuss their inferential logic, and describe several examples in each category. The three categories involve measures that are derived pure (based on different transformations of the same data; example: separation of sensory and decision processes by signal detection theory), direct pure (based on different data; example: selective effects of adaptation on spatial-frequency thresholds), and composite (examples: the multiplicative-factor method for the analysis of response rate; the additive-factor method for the analysis of reaction time). Six of the examples concern behavioral measures and functional processes, while four concern brain measures and neural processes. They have been chosen for their interest and importance; their diversity of measures, species, and combination rules; their illustration of different ways of thinking about data; the questions they suggest about possibilities and limitations of the separate-modifiability approach; and the case they make for the fruitfulness of searching for mental modules. PMID- 11256337 TI - Eccentric head positions bias random generation of leftward and rightward handle bar rotations. AB - We explored the effects of eccentric head positions on rapid rotations of a handle-bar in a modified randomization task. Subjects had to generate a random sequence of leftward and rightward handle-bar rotations; each pre-selected rotation was produced in response to a visual signal as rapidly as possible. Eccentric head positions induced a bias in that handle-bar rotations in the direction of the eccentric head position were chosen more frequently than handle bar rotations in the opposite direction. This is consistent with previous evidence on a spatial coupling. In contrast to response selection, response initiation was not affected by eccentric head positions. The kinematic characteristics, however, differed: rotations in the direction of the eccentric head position were of larger amplitude and longer duration than rotations in the opposite direction. This difference may have been secondary to a difference in the start positions of the handle-bar, which were shifted in the direction opposite to the eccentric head position. PMID- 11256338 TI - Movement observation affects movement execution in a simple response task. AB - The present study was designed to examine the hypothesis that stimulus-response arrangements with high ideomotor compatibility lead to substantial compatibility effects even in simple response tasks. In Experiment 1, participants executed pre instructed finger movements in response to compatible and incompatible finger movements. A pronounced reaction time advantage was found for compatible as compared to incompatible trials. Experiment 2 revealed a much smaller compatibility effect for less ideomotor-compatible object movements compared to finger movements. Experiment 3 presented normal stimuli (hand upright) and flipped stimuli (hand upside-down). Two components were found to contribute to the compatibility effect, a dynamic spatial compatibility component (related to movement directions) and an ideomotor component (related to movement types). The implications of these results for theories about stimulus-response compatibility (SRC) as well as for theories about imitation are discussed. PMID- 11256339 TI - Development of the functional visual field. AB - Andries Sanders' dissertation examined selective mechanisms in the functional visual field, and much of his work since has been concerned with the stages that underlie visual information processing particularly while making saccades. We argue that the study of orienting in the functional visual field is timely because it deals with the relation of covert attention shifts, eye movements and head movements to their underlying neurology. In our paper we develop a method to study learning of sequences at all ages from infants to adults. Our studies focus on how learning influences anticipatory eye movements. We examined the learning of unambiguous and context dependent sequences by 4-, 10-, and 18-month-old infants and undergraduates. We found clear learning of unambiguous sequences at 4 months, but learning of context dependent associations was found only in 18-month olds and in adults. We hypothesize that the learning of unambiguous sequences by 4-month-olds reflects maturation of a basal ganglia-parietal circuit related to adult implicit learning, while the learning of context dependent sequences requires development of frontal structures underlying more general attentional abilities. PMID- 11256340 TI - Concatenating familiar movement sequences: the versatile cognitive processor. AB - Earlier studies demonstrated that practicing a series of key presses in a fixed order yields memory representations (i.e., motor chunks) that can be selected and used for sequence execution as if familiar key pressing sequences are single responses. In order to examine whether these motor chunks are robust in different situations and whether preparation for one sequence may overlap with execution of another one, two experiments were carried out in which participants executed two highly practiced keying sequences in rapid succession in response to two simultaneously presented stimuli. The results confirmed robustness of motor chunks, even when the sequences included only two elements, and showed that preparation (and in particular, selection) of a forthcoming sequence may occur during execution of the earlier sequence. Sequences including only two keys appeared to be slowed more by concurrent preparation than longer sequences. Together these results suggest that the execution of familiar keying sequences is predominantly carried out by a dedicated motor processor, and that the cognitive processor can be allocated to preparing a forthcoming sequence (e.g., during execution of an earlier sequence) or, some times, to selecting individual sequence elements in parallel to the motor processor. PMID- 11256341 TI - Feature localization and identification. AB - Theories on visual search differ substantially with respect to the relationship they assume between localization and identification processes. The aim of the present study was to rigorously compare the alternative theoretical notions on how localization and identification processes are related. In two experiments, participants searched for a target with a unique line orientation among distractors containing another orientation. Localization and identification performance were measured in combination, as function of display size and target eccentricity. To compare the alternative theories, formal binomial models were developed and compared with respect to their goodness of fit to the individual data. The formal analyses showed that the model assuming identification processes to be conditioned on localization processes provided the best fit to the individual data. Furthermore, maximum likelihood estimates of the parameter corresponding to identification processes were differently affected by display size than identification performance was. The results were discussed in terms of their implication for current theories on visual search. PMID- 11256342 TI - Future challenges in perinatal-neonatal medicine. PMID- 11256343 TI - Leptospirosis in Kuala Lumpur and the comparative evaluation of two rapid commercial diagnostic kits against the MAT test for the detection of antibodies to leptospira interrogans. AB - The aim of the study was to look into the epidemiology of serodiagnosed cases of leptospirosis at the University Hospital and compare two commercial ELISA Assays to the Microscopic Agglutination Test (MAT). Demographic data for all serodiagnosed cases for the years 1991-1997 were collected. From this data, 104 sera (n = 104) were selected as samples for comparative evaluation of the commercial ELISAs (INDX Dip-S-Ticks and PanBio ELISA) to the MAT test. Thirty two (n = 32) negative control sera were selected from serodiagnosed cases of other differential diagnosis of leptospira infection. The MAT test is a standard test that detects agglutination antibodies to leptospira biflexa, while the INDX Dip-S Ticks is an ELISA dot test assaying for total anti-leptospira antibodies. The PanBio ELISA is a colorometric assay in test well strips to detect anti leptospira IgM. The sensitivity, specificity, and efficiency of tests were calculated at a MAT cut-off value of 1:320. Demographic data showed that leptospirosis peaks during March-May and Aug-Nov coinciding with the inter monsoon period with more men being infected than women and more adults than children. The sensitivity, specificity, and efficiency of test for the INDX Dip-S Ticks were 83.3%, 93.8% and 87.5% while the values for the PanBio ELISA were 54.2%, 96.9% and 71.3%. The suboptimal PanBio result could be related to the blocking effect of high IgG titres or could be related to the diagnostic MAT cut off values used in this study. The data hence reflects a pattern of transmission that is related to "wet" occupational risk factors. The commercial assays evaluated, are easier to perform but interpretation of results should be based on level of endemicity. The INDX Dip-S-Ticks allows this flexibility and is a practical alternative to the MAT test. PMID- 11256344 TI - Bedside assessment of swallowing: a useful screening tool for dysphagia in an acute geriatric ward. AB - AIM: Dysphagia is common in the elderly and is associated with increased morbidity and mortality. We undertook a prospective study to determine the usefulness of a simple bedside swallowing test in terms of (1) detecting previously undiagnosed dysphagia, (2) agreement of the doctor's assessment with that of the speech therapist, (3) impact on subsequent feeding modality, (4) predicting risk of subsequent pneumonia. METHOD: Patients in an acute geriatric ward who had no contra-indications to oral feeding were subjected to a bedside swallowing assessment by a geriatrician within 24 hours of admission. All patients found to be dysphagic were subsequently re-assessed by a speech therapist within 48 hours. In addition, every fifth patient deemed to have normal swallowing by the doctor was assessed by the speech therapist. RESULTS: Sixty five patients were studied. The doctor's assessment was in very good agreement with the assessment of the speech therapist (kappa = 0.87). Patients found to have dysphagia using the doctor's assessment protocol had an increased risk of developing pneumonia during their hospitalization (relative risk R.R.: 9.9 confidence interval C.I.: 1.2-81.2). Cough on swallowing and delayed swallowing were both found to be associated with an increased risk of developing pneumonia during the period of hospitalization (R.R.: 4.2, C.I.: 1.2-14.4; R.R.: 5.3, C.I.: 1.1-26.3 respectively). CONCLUSION: A simple bedside swallowing test can be used as an effective screening tool in detecting hitherto undiagnosed dysphagia. The validity of this tool in the diagnosis of aspiration requires further investigation. PMID- 11256345 TI - Recurrence of Helicobacter pylori infection and duodenal ulcer relapse, following successful eradication in an urban east Asian population. AB - We aimed to determine the rate of Helicobacter pylori (HP) recurrence and duodenal ulcer relapse in patients of a hospital in Singapore over a period of at least one year from the time of eradication. Ninety-six consecutive duodenal ulcer patients with biopsy-proven HP eradication and healed ulcer were seen at 3 month intervals, and follow-up endoscopy was performed when dyspepsia recurred, at the end of one year after eradication, or at the time of recall if the patient had been lost to follow-up. HP status was determined by antral and corpus biopsies and by antral cultures. Sixty-five had been given triple therapy, and 31 received dual therapy with omeprazole + amoxycillin or clarythromycin. Median time to follow-up endoscopy was 12 months. Six patients (6.25%) were positive for HP infection after eradication. Recurrence of HP infection was detected at 9 and 10 months after confirmation of HP eradication in two patients, and at between 13 and 20 months in the remaining four. Two of these had recurrent duodenal ulcer; all but one had erosive duodenitis. Two other patients had recurrent duodenal ulcer despite absence of HP reinfection; they admitted to taking low-dose aspirin. It was concluded that the recurrence of HP infection is low at the end of one year after successful eradication therapy in this urban East Asian population. Ulcer relapse occurred in 4.17% (4/96) of patients, and was associated with recurrent HP infection or NSAID exposure. PMID- 11256346 TI - Ultrasonography and computed tomography in a clinical algorithm for the evaluation of suspected acute appendicitis in children. AB - AIM: To evaluate the roles and effectiveness of US and CT in a clinical algorithm for the evaluation of children with suspected appendicitis. METHODS: Patients with suspected appendicitis were prospectively evaluated with ultrasound (US), and in some cases with CT, after they were graded to have high, intermediate or low clinical likelihood for appendicitis. Imaging findings were made known to clinicians who then decided on a line of management. Patho-histological examination and clinical follow-up established the final diagnoses, which were correlated with the imaging findings. The effect of imaging on the management of patients was examined. RESULTS: Overall, the sensitivity of US was 92.9%, specificity 96.9%, accuracy 96.0%, positive predictive value 89.7% and negative predictive value 97.9%. Imaging did not affect the decision to operate in 13/14 (92.9%) patients in the high likelihood subgroup. Imaging guided the clinicians to the right management pathway in 26/30 (86.7%) patients in the intermediate group. 77/82 (93.9%) of US was truly negative in the low likelihood group. CT was performed in 12 patients because of unsatisfactory US scans or incompatibility between the US and the clinical findings. CT correctly diagnosed the presence or absence of appendicitis in all 12 patients. CONCLUSION: US and CT are accurate modalities in the diagnosis of acute appendicitis in children. US is most useful in patients with equivocal clinical findings. US should be the first modality used to evaluate children with suspected appendicitis. CT should be reserved for cases where US is sub-optimal or where the findings are inconsistent with the clinical findings. PMID- 11256347 TI - Refractive errors and strabismus in premature Asian infants with and without retinopathy of prematurity. AB - AIM: In Caucasian populations, premature infants with retinopathy of prematurity (ROP) have been reported to have higher risks of developing refractive errors and strabimus. The purpose of this study is to evaluate the rate of these complications in Asian premature infants with and without ROP. METHODS: A retrospective case review of all premature infants referred to the Singapore National Eye Centre for ophthalmology screening. These included all neonates born earlier than 34 weeks gestational age and less than 1500 grams in birth weight. Standardized ophthalmology examinations including cycloplegic refraction and fundus examination at regular intervals were performed to determine the presence of ROP, refractive errors, squints and other ocular abnormalities until the patients were 3 years old. RESULTS: During 1991 to 1993, a total of 113 neonates were reviewed. Of these, 16 (14.2%) developed ROP. The risk of ROP was higher with lower birth weights and earlier gestational ages. At 1-year follow-up, the rate of myopia was 33.3% in babies with ROP compared to 3.7% in babies with no ROP (p < 0.001). The higher rates of myopia in babies with ROP remained with longer follow-up (33.3% and 25% in ROP group versus 3.4% and 3.8% in no ROP group, at 2 and 3 years respectively). There was no difference in rates of astigmatism or hyperopia throughout the 3 years. At 1 year follow-up, the rate of strabismus was 20% in the ROP group compared to 4.9% in the no ROP group (p = 0.07). However, this difference in rates of strabismus was not significant at 2 and 3 years of follow-up. CONCLUSION: Premature babies with ROP had higher rates of myopia and strabismus than those without ROP. Long-term follow-up of these babies is important for early detection and treatment of these ocular problems. PMID- 11256348 TI - Paediatric one lung anaesthesia by selective bronchial intubation. AB - One lung anaesthesia in paediatric patients may not always be achievable by bronchial blockade or double lumen tube intubation due to inadequate experiences or facilities. We attempted to isolate right lung by selectively intubating the left bronchus with single lumen tube on a 10 kg child. Optimal surgical condition and satisfactory oxygenation achieved but complicated with severe respiratory acidosis. The possible causes for hypercapnea in this child were discussed. PMID- 11256349 TI - Branch retinal artery occlusion secondary to a Hollenhorst plaque. AB - Retinal arterial circulation obstruction has serious implications. It may result in acute visual loss, but more significantly, it implies that the patient's systemic health needs further review and investigations in order to prevent severe and life-threatening consequences such as myocardial infarction and cerebrovascular accidents. We report a case of a patient with branch retinal artery occlusion with the presence of a Hollenhorst plaque. PMID- 11256350 TI - A case of recurrent disseminated granuloma annulare. AB - We present an 11-year-old boy who developed multiple pruritic, skin-coloured papules on his forehead, which subsequently spread to his trunk and limbs over a period of 6 months. Histology revealed granuloma annulare. The lesions underwent spontaneous regression over the next five years. He presented again when he was 18 years old, with a spontaneous eruption of multiple lesions on the trunk and the limbs 2 years after complete clinical remission from the first attack. A punch biopsy performed at this presentation revealed granuloma annulare again. No treatment was given and he is still being followed up. The tendency for spontaneous resolution of granuloma annulare is well recognised. Recurrent localized lesions often disappeared more rapidly than the original ones. Recurrence of disseminated GA, to our knowledge, has not been reported before. PMID- 11256351 TI - Cementoplasty and the oncologic population. AB - The first and only description of percutaneous cementoplasty, to date, has been described in the French medical literature in 1994. In this series of 12 cases, radiologists successfully instilled a cement derivative into the acetabulum under fluoroscopic control. As in these cases, the major indication for cementoplasty is to provide pain control and stabilization of an osteolytic lesion. Potential complications include physical or thermal damage to the adjacent neurovascular structures, either during needle positioning or from cement leakage, respectively. Although no absolute contraindications exist, one should proceed cautiously in patients with coagulopathies. Results may be suboptimal as well in patients with pathologic fractures. PMID- 11256352 TI - Q methodology--a journey into the subjectivity of human mind. AB - AIMS: This paper introduces a relatively new research methodological tool, known as Q method, useful in exploring issues related to human subjectivity. DESCRIPTION: Q methodology is unique as it combines the strengths of both qualitative and quantitative research traditions. The sequential steps involve generation of ideas about the research topics, clarification and refinement of the ideas, and rank ordering these ideas by the respondents in a quasinormal distribution. The data are extracted with by-person factor analysis and useful in exploring arrays of attitude either cross-sectionally or longitudinally over a period of time. CONCLUSION: Q methodology can be used to analyse opinions, perceptions, and attitudes in both clinical and non-clinical settings. It is a preferred method of human subjectivity study as it provides more in-depth analysis of complex subjectivity issues. PMID- 11256353 TI - Clinics in diagnostic imaging (52). Spinal cord schistosomiasis. AB - A 2-year-old Brazilian boy presented with bilateral leg weakness and constipation, followed by development of progressive paraparesis and bladder dysfunction. Neurological examination revealed flaccid paraparesis. Blood tests and CSF analysis showed eosinophilia. The MR examination revealed a spinal cord mass extending from T9 to L1 levels, with a heterogeneously-enhancing solid component and a cystic component. Stool tests for Schistosoma mansoni eggs were positive. The patient underwent surgery, the intramedullary mass was partially resected, and the diagnosis of spinal cord infection by Schistosoma mansoni was confirmed. After surgery, the patient was treated with praziquantel and oxamniquine. He was discharged with partial improvement of the lower extremity weakness and bowel/bladder function. The clinical and imaging features of spinal cord schistosomiasis are reviewed. PMID- 11256354 TI - The case of plagiarism in Wiadomosci Lekarskie. PMID- 11256355 TI - Utilization of psychosocial treatments by patients diagnosed with bipolar disorder and substance dependence. AB - We investigated psychosocial treatment interventions, mood symptoms, and substance use among 24 patients with bipolar disorder and substance dependence. Patients were assessed for 6 months following hospital discharge. Psychotherapy and Alcoholics Anonymous (AA) attendance decreased over time. Moreover, the focus of patients' psychotherapy changed over time, with decreasing emphasis on the patients' specific disorders. Mood symptoms and substance use did not change significantly over time, although there was a trend toward more frequent drug use over time. These findings point to infrequent utilization over time of psychosocial treatments focusing specifically on bipolar and substance use disorder. PMID- 11256356 TI - Early life stress can programme our health. AB - It is essential that the environment for a developing foetus is optimal for normal growth and maturation. Small perturbations in this environment may put that child at risk for developing cardiovascular, metabolic and cognitive deficits later in life. Evidence is accumulating that chronic stress while pregnant may result in lower birthweight babies and a heightened risk of mood disorders and cognitive deficits. Elevated glucocorticoid hormones, induced in the mother in response to the stress, appear to be mediators of events 'programming' the developing central nervous system of the foetus and rendering it susceptible to dysfunction in later life. PMID- 11256357 TI - Influence of ophthalmic viscosurgical device on the effects of intracameral anesthesia and stains. PMID- 11256358 TI - Looking through a cataract. PMID- 11256359 TI - Caveat emptor: editors beware. PMID- 11256360 TI - Bimonthly update. Nutrition and metabolism. PMID- 11256361 TI - Web Alert. PMID- 11256362 TI - Is needle biopsy of the liver necessary to diagnose HCC? PMID- 11256363 TI - Management of gastric fundal varices associated with a gastrorenal shunt. PMID- 11256364 TI - The science, economics, and effectiveness of combination therapy for hepatitis C. PMID- 11256365 TI - Screening for genetic haemochromatosis in blood samples with raised alanine aminotransferase. PMID- 11256366 TI - [Complete congenital cartilaginous trachea in a girl with Crouzon's syndrome]. AB - A tracheostomy was performed in a 4-month-old girl with Crouzon's syndrome because of upper respiratory obstruction. During the procedure the absence of tracheal rings was observed. These findings were confirmed by postoperative bronchoscopy. Subsequent surgical correction of the patient's craniofacial anomalies enabled decannulation when the patient was 10 months old. Complete cartilaginous trachea is very rare and is always associated with craniosynostotic syndromes. Tracheobronchial anomalies should be investigated in patients whose respiratory symptoms are not due to upper airway obstruction. PMID- 11256367 TI - Comparison of St John's Wort and imipramine. Sensitivity of assay is questionable. PMID- 11256368 TI - Addison's disease: after 40 years much remains the same. PMID- 11256369 TI - Editorial on management of anal warts is misleading. PMID- 11256370 TI - Access to undergraduate medical education is being broadened. PMID- 11256371 TI - Practices should determine their own strategies for identifying patients. PMID- 11256373 TI - AHA and federal agencies agree to cooperate to attack heart disease and stroke. PMID- 11256372 TI - Contaminated feed heightens "mad cow" fears in the United States. PMID- 11256374 TI - President Bush sends Medicare drug plan to Congress. PMID- 11256375 TI - Amphioxus goosecoid and the evolution of the head organizer and prechordal plate. AB - The organizer is a central feature of vertebrate embryogenesis and is functionally subdivided into the head organizer that gives rise primarily to the prechordal plate and induces forebrain structures, and the trunk/tail organizer that gives rise primarily to the notochord and induces more posterior structures. Goosecoid(gsc) encodes a homeodomain-containing transcription factor that is expressed in the vertebrate head organizer and prechordal plate, and can induce a secondary axis when expressed ectopically. To investigate the evolution of the vertebrate head organizer and prechordal plate, we have cloned and characterized a gsc homolog from the cephalochordate amphioxus. Amphioxus, it is important to note, lacks a prechordal plate in that the notochord extends to the extreme anterior end of the animal, and lacks elaborate differentiation of its forebrain. Gsc expression in amphioxus is initially localized during gastrulation to the mesendodermal layer of the dorsal lip of the blastopore. However, gsc expression in amphioxus is not maintained in anterior axial mesoderm, as is the case with the vertebrate prechordal plate. Rather, gsc is expressed in the dorsal axial mesoderm of the blastopore lip throughout gastrulation, appearing transiently in the presumptive notochord that underlies all regions of the amphioxus brain. The similarities in gsc expression in amphioxus and vertebrates suggest that a primitive version of the head organizer evolved prior to the origin of the vertebrates. The differences in gsc expression can be interpreted either as the loss of the prechordal plate domain in the cephalochordate lineage, or the gain of a distinct gsc-expressing prechordal plate that plays a role in forebrain induction in the vertebrate lineage. PMID- 11256376 TI - Characterization of the Hox cluster from the mosquito Anopheles gambiae (Diptera: Culicidae). AB - The Hox genes have been found to encode transcription factors, which specify the morphological identity of structures along the anteroposterior axis of animals ranging from worms to mice. The canonical set of nine genes is organized in a cluster in the genome of several protostomes and deuterostomes. However, within insects, whereas the Hox genes are organized in a single cluster in the beetle Tribolium castaneum, they are split into two separate groups in the flies Drosophila melanogaster and Drosophila virilis. The significance of a split Hox cluster is unknown and has been observed in only one organism outside the Drosophila lineage: the nematode Caenorhabditis elegans. We have cloned a majority of the Hox genes from the mosquito Anopheles gambiae (Diptera: Culicidae) and compared their genomic organization with that of Tribolium and Drosophila to determine if a split Hox cluster is found in dipterans aside from the Drosophilidae. We find that the Hox genes in Anopheles, as in Tribolium, are organized in a single cluster that spans a genomic region of at least 700 kb. This finding suggests that, within the insect genome, the partition of the Hox cluster may have evolved exclusively within the Drosophila lineage. The genomic structures of the resident genes, however, appear to be largely conserved between A. gambiae and D. melanogaster. PMID- 11256377 TI - Characterization of the Hox gene cluster in the malaria vector mosquito, Anopheles gambiae. AB - The Hox genes play a central role in regulating development and are involved in the specification of cell fates along the anteroposterior axis. In insects and vertebrates, these genes are clustered and organized in an arrangement that is largely conserved across evolutionary lineages. By exploiting the sequence conservation of the homeobox, orthologues of the Hox genes Sex combs reduced (Scr), fushi tarazu (ftz), Antennapedia (Antp), Ultrabithorax (Ubx), and abdominal-A (abd-A) have been isolated from the malaria vector mosquito, Anopheles gambiae. These genes were first identified in Drosophila, where they achieve a high level of functional complexity, in part, by the use of alternative promoters, polyadenylation sites, and splicing to generate different protein isoforms. Preliminary analyses of the Anopheles Hox genes suggest that they do not achieve their functional complexity in the same manner. Using a combination of in situ hybridization to polytene chromosomes and chromosome walking, the Anopheles Hox genes have been localized to a single cluster in the region 19D-E on chromosome 2R, a situation distinct from that of Drosophila where the Hox complex is split into two clusters. This study, therefore, provides a framework for future comparative analyses of the structure, organization, and expression of developmental regulatory genes between the lower and higher Diptera. Moreover, the genes that have been isolated enhance the genetic and physical maps of chromosome 2R in this medically important mosquito species. PMID- 11256378 TI - Molluscan engrailed expression, serial organization, and shell evolution. AB - Whether the serial features found in some molluscs are ancestral or derived is considered controversial. Here, in situ hybridization and antibody studies show iterated engrailed-gene expression in transverse rows of ectodermal cells bounding plate field development and spicule formation in the chiton, Lepidochitona cavema, as well as in cells surrounding the valves and in the early development of the shell hinge in the clam, Transennella tantilla. Ectodermal expression of engrailed is associated with skeletogenesis across a range of bilaterian phyla, suggesting a single evolutionary origin of invertebrate skeletons. The shared ancestry of bilaterian-invertebrate skeletons may help explain the sudden appearance of shelly fossils in the Cambrian. Our interpretation departs from the consideration of canonical metameres or segments as units of evolutionary analysis. In this interpretation, the shared ancestry of engrailed-gene function in the terminal/posterior addition of serially repeated elements during development explains the iterative expression of engrailed genes in a range of metazoan body plans. PMID- 11256379 TI - Evolutionary relationships between the amphibian, avian, and mammalian stomachs. AB - Although the gut is homologous among different vertebrates, morphological differences exist between different species. The most obvious variation in the guts of extant vertebrates appears in the stomach. To investigate the evolution of this structure, we compared the histology of the stomach and gastrointestinal tract in amphibian (Xenopus laevis), avian (Gallus gallus), and mammalian (Mus musculus) organisms, and defined the expression patterns of several genes within the developing guts of these lineages. In all three groups, we find that the anterior portion of the stomach has a similar glandular histology as well as a common embryonic expression of the secreted factors Wnt5a and BMP-4. Likewise, within the amniote lineages, the posterior nonglandular stomach and pyloric sphincter regions are also comparable in both histological and molecular phenotypes. The posterior stomach expresses Six2, BMPR1B, and Barx1, whereas the pyloric sphincter expresses Nkx2.5. Although the adult Xenopus stomach exhibits both glandular and aglandular regions and a distinct pyloric sphincter similar to that of the amniotic vertebrates, the histology of the Xenopus tadpole gut shows less distinct variation in differentiation in this region, which is most likely a derived condition. The molecular signature of the embryonic Xenopus gut correlates with the more derived morphology of the larval phase. We conclude that the global patterning of the gut is remarkably similar among the different vertebrate lineages. The distinct compartments of gene expression that we find in the gut be necessary for the unique morphological specializations that distinguish the stomachs from terrestrial vertebrates. PMID- 11256380 TI - Turning Hox "signatures" into synapomorphies. AB - It has recently been shown that the three metazoan superphyla that are recognized on the basis of 18S rDNA phylogenies--ecdysozoans, lophotrochozoans, and deuterostomes--each have characteristic Hox genes. This observation has been taken further, and these "signature" Hox genes have been looked for in taxa of uncertain affinity such as the mesozoa, in order to link them to one of the three superphyla. Here I point out that, in the absence of an out-group, these so called signature Hox genes are unpolarized characters and, as such, should not be used in this cladistic sense to determine phylogeny. Taking the example of the mesozoans, which have the Lox5 gene in common with the lophotrochozoans, I show that it is possible to polarize this character using paralogous Hox genes as proxy out-groups; however, due to the impossibility of reliable alignment outside the homeobox, only two residues of the Lox5 peptide are susceptible to this method. With this in mind, I find slim evidence for an association between mesozoans and lophotrochozoans. I demonstrate that the lophotrochozoan genes Lox2 and Lox4 would provide many more reliable residues that are truly indicative of lophotrochozoan affinity. Finally, I point out the potential problems in using unpolarized signatures to address the question of the position of the acoel flatworms. PMID- 11256382 TI - PCR template preparation for capillary DNA sequencing. AB - Fluorescence-based capillary DNA sequencing has facilitated the early completion of several complex sequencing projects. While capillary systems offer great benefits in terms of ease of use and automation, we find that they are sufficiently different from slab gel separation methodologies, demanding re examination of the protocols used to generate and use DNA sequencing templates. We have recently initiated a large-scale Human Open Reading Frame EST project involving 30 laboratories feeding 11 MegaBace 1000 capillary sequencers. The group has already produced more than 300,000 valid sequences. The most successful template preparation protocol we have found is described here. We have found that a crucial step is the standardization of the quantity and quality of the templates, which have been achieved by overnight bacterial culture followed by PCR using limiting amounts of primers. Using this protocol, there is no need for post-PCR purification, and the final preparation cost is US $0.09/template. After sequencing 10,848 templates using this protocol, 78% of the reads were accepted (after discarding vectors without inserts and inserts smaller than 100 nucleotides), and 85% of the total number of bases had Phred scores of 15 or above. PMID- 11256381 TI - Comparison of length of hospital stay for patients with known or suspected methicillin-resistant Staphylococcus species infections treated with linezolid or vancomycin: a randomized, multicenter trial. AB - STUDY OBJECTIVE: To compare hospital length of stay (LOS), weekly discharges, and days of antibiotic treatment with linezolid (intravenous with oral follow-up) and vancomycin (intravenous only). DESIGN: Multinational, randomized, phase III trial. SETTINGS: Hospitals in North America, Latin America, and Europe. PATIENTS: Four hundred sixty hospitalized patients with infections of known or suspected methicillin-resistant Staphylococcus species. INTERVENTION: Administration of linezolid or vancomycin. MEASUREMENTS AND MAIN RESULTS: For linezolid recipients, median LOS was 5 and 8 days shorter (p=0.05 and 0.003) in the complicated skin and soft tissue infection intent-to-treat (230 patients) and clinically evaluable (144) samples, and slightly but not significantly shorter in the overall intent to-treat (460) and clinically evaluable (254) samples. In all samples, linezolid recipients had more discharges in the first week of treatment and fewer days of intravenous therapy than vancomycin recipients. CONCLUSION: Our results support linezolid's ability to reduce medical resource use. PMID- 11256383 TI - Selective laser photocoagulation of communicating vessels in severe twin-twin transfusion syndrome in women with an anterior placenta. AB - BACKGROUND: We describe two techniques for the laser treatment of twin-twin transfusion syndrome in women with anterior placentas. TECHNIQUE: In the first technique, anastomoses were photocoagulated using a flexible endoscope through a single port. The second technique used a side-firing laser fiber with a rigid angled-view endoscope (two ports). EXPERIENCE: Seventy-two women had surgery between July 1997 and December 1999, 35 (48.6%) of whom had anterior placentas. Survival was similar for fetuses with anterior (80%) and posterior (75.6%) placentas, but operating time was significantly longer for those with anterior placentas (81.1 compared with 64.4 minutes for the anterior and posterior placentas, respectively; P = .02, Student t test). At least one fetus survived in 76% (16 of 21) of women treated with flexible endoscopes and 86% (12 of 14) of those treated with the side-firing lasers. Six of 72 women (8.3%) had patent vascular anastomoses on placental examination, and five of them had anterior placentas (P = .08, Fisher exact test). CONCLUSION: Although anterior placentas are surgically more challenging than posterior placentas, both techniques allow an effective percutaneous approach to the laser treatment of twin-twin transfusion syndrome. PMID- 11256384 TI - Wet smear compared with Gram stain diagnosis in asymptomatic pregnant women. PMID- 11256385 TI - Rethinking maternal-fetal conflict: gender and equality in perinatal ethics. PMID- 11256386 TI - Conserving biodiversity and ecosystem services. PMID- 11256387 TI - Financial conflict. Universities puncture modest regulatory trial balloon. PMID- 11256388 TI - Parkinson's research. Fetal cell transplant trial draws fire. PMID- 11256390 TI - Gender equity. NSF program targets institutional change. PMID- 11256391 TI - Neuroscience. Dyslexia: same brains, different languages. PMID- 11256389 TI - Human cloning. Experts assail plan to help childless couples. PMID- 11256393 TI - Astronomy. Stars rise from ashes in globular cluster. PMID- 11256392 TI - Cell biology. How bacterial flagella flip their switch. PMID- 11256394 TI - Innate immunity. Ancient system gets new respect. PMID- 11256395 TI - Innate immunity. Engineering protection for plants. PMID- 11256396 TI - Biophysical Society meeting. Crossover research yields scents and sensitivity. PMID- 11256397 TI - Research collaborations. NASA lab offers land to lure research partners. PMID- 11256398 TI - Demography. Sardinia's mysterious male Methuselahs. PMID- 11256399 TI - Demography. An island of 'genetic parks'. PMID- 11256400 TI - Neurobiology. Does alcohol damage female brains more? PMID- 11256401 TI - Deterring bioweapons development. PMID- 11256402 TI - Moribund funding in agricultural research. PMID- 11256403 TI - Porous sediments at the top of Earth's core? PMID- 11256404 TI - Tale of two kings. PMID- 11256405 TI - Climate change. Whither after The Hague? PMID- 11256407 TI - Geology. The deadliest intraplate earthquake. PMID- 11256406 TI - Development. How to stimulate your partner. PMID- 11256408 TI - Transduction. When worlds collide--trafficking in JNK. PMID- 11256409 TI - Ecology. The enemy of my enemy is my ally. PMID- 11256410 TI - Chemistry. A light-driven linear motor at the molecular level. PMID- 11256411 TI - Chemistry. Water on the move. PMID- 11256412 TI - And so they never mentioned it again. PMID- 11256413 TI - Genetic variation for the positioning of wing veins in Drosophila melanogaster. AB - To define the components of variation for wing shape in Drosophila in relation to what is known about the developmental control of wing patterning, we have characterized shape variation in the wings of 12 randomly chosen highly inbred lines. Despite large differences in wing size between males and females, and between flies reared at 18 degrees C or 25 degrees C, wing shape is remarkably unaffected by these variables and is highly line specific. The shape of each intervein region of the wing appears to be independently regulated at the genetic level, consistent with the role of secreted growth factors in establishing the locations of wing veins. Sex and temperature were found to have different effects on cell number in two intervein regions, with the result that wing shape is to a large extent independent of cell density. Dietary cholesterol was also shown to affect the breadth of the central intervein region, consistent with an effect on the strength of Hedgehog signaling during wing development. We conclude that wing shape is under tighter genetic control than wing size, and hypothesize that this control is achieved in large part by gene activity at the level of wing vein determination and differentiation. PMID- 11256414 TI - Development and evolution of adult feeding structures in Caenogastropods: overcoming larval functional constraints. AB - Comparative study of the developing foregut in three species of caenogastropods, including an herbivorous grazer (Lacuna vincta) and two carnivores (Euspira [Polinices] lewisii and Nassarius mendicus), suggests how the specialized adult foregut of a carnivorous neogastropod evolved within a life cycle having a planktotrophic larva. Postmetamorphic feeding structures (buccal cavity and radular sac) in all three species achieve advanced differentiation in the larval stage, permitting juvenile feeding at 3 days postmetamorphosis. Recent phylogenetic hypotheses for the Gastropoda predict that foregut developmental patterns in E. lewisii and N. mendicus are derived, relative to that of L. vincta. In hatching larvae of these three, the anlage of postmetamorphic feeding structures is a small patch of nonciliated cells embedded in the ventral wall of the larval foregut and the patch soon forms an outpocketing. During subsequent morphogenesis, Euspira lewisii and N. mendicus share a developmental novelty that involves semi-isolation of the developing, postmetamorphic buccal cavity and radular sac from the larval foregut and formation of a new, definitive mouth at metamorphosis. Nassarius mendicus, a neogastropod, embellishes this novelty by adding the entire anterior esophagus and valve of Leiblein (de novo structures) to the semi-isolated buccal cavity. Therefore, a valve and long stretch of muscular anterior esophagus, which are necessary for feeding with a pleurembolic proboscis, are preformed in the larval stage of this neogastropod without interfering with larval feeding. The inferred evolutionary events leading to postmetamorphic feeding specialization in N. mendicus are invisible in adults; they require reconstruction from comparative developmental analysis. PMID- 11256415 TI - The neural crest as a fourth germ layer and vertebrates as quadroblastic not triploblastic. PMID- 11256416 TI - Holomeric vs. meromeric segmentation: a tale of centipedes, leeches, and rhombomeres. AB - Explaining the origin and evolution of segmentation is central to understanding the body plan of major animal groups such as arthropods, annelids, and vertebrates. One major shortcoming of current views on segmentation is the failure to recognize the existence of two layers of segmentation. I distinguish here holomeric segmentation, involving the whole body axis (or the whole axis of an appendage) and producing "true" segments (eosegments); and meromeric segmentation, producing merosegments within one or more eosegment(s). In terms of developmental mechanisms, meromeric segmentation is probably the same as compartmentalization. This process follows two rules: (1) merosegments are formed from a stereotyped pattern of subdivisions, where only the merosegments in contact to the anterior or posterior boundary of the eosegment are allowed to divide; (2) contiguous eosegments undergoing meromeric segmentation generate merosegments according to identical lineage patterns apart from possible lineage truncation in one or a few terminal eosegments. The segmentation model proposed in this paper is mainly supported by evidence from comparative morphology, but it is compatible with known cellular and developmental mechanisms. The development of vertebrate rhombomeres, the annulation of leeches, the subdivision of the distal part of insect antenna into flagellomeres and the segmentation of centipedes are interpreted here in terms of meromeric segmentation. Some of these phenomena, like centipede segmentation, have thus far defied all attempts at an explanation, both in mechanistic (developmental) and phylogenetic terms. The model presented in this paper suggests a rich research agenda at all levels, from molecular and genetic to morphological and phylogenetic. PMID- 11256417 TI - The concept of developmental reprogramming and the quest for an inclusive theory of evolutionary mechanisms. AB - Evolutionary developmental biology has already made a major contribution to our understanding of evolutionary patterns, notably homology. However, while it has the potential to make an equally important contribution to our understanding of evolutionary mechanisms, and indeed to the integration of mechanism and pattern, it has not yet done so. This paper explores how this potential may be realized. In particular, I focus on the limitations of present-day neo-Darwinian theory, and indicate how a combination of the neo-Darwinian and "evo-devo" approaches provides a more inclusive view of evolutionary mechanisms with greater explanatory power. There is a particular focus on developmental reprogramming, which lies logically between mutation and selection, yet has been neglected in mainstream evolutionary theory. The inclusion of developmental reprogramming in the list of evolutionary mechanisms leads to a view that the direction of evolutionary change is determined by a combination of internal and external factors, rather than being controlled entirely by the environment. PMID- 11256418 TI - A diverse approach to arthropod development. PMID- 11256419 TI - Reproductive isolation in Caenorhabditis: terminal phenotypes of hybrid embryos. AB - Several interspecific combinations of the "elegans" group of Caenorhabditis species are cross-fertile. Most F1 hybrids from these crosses arrest during embryogenesis. Developmental defects observed in hybrid embryos include defects in gastrulation initiation, defects in embryonic compaction, and defects in embryonic elongation. These reproductive barriers have arisen multiple times in the evolution of Caenorhabditis. PMID- 11256420 TI - The alloresponse. AB - The alloresponse can be divided into two components. The first of these is allorecognition, which refers to the recognition of antigens, expressed on the surface of cells of non-self origin, by the host's lymphocytes. The second part is the immune effector mechanisms generated by this recognition process. The molecules recognised have been termed histocompatibility antigens and fall into two categories. The strongest responses are provoked by allogeneic major histocompatibility complex (MHC) antigens. T cells recognise these antigens either directly or after being processed like conventional antigens by antigen presenting cells, in what has been termed indirect presentation. In the context of MHC identity, responses are observed against the second category of antigens, namely minor histocompatibility antigens (mHAgs). Although weaker, these responses are of clinical importance, particularly in bone marrow transplant recipients. CD4+ T cells play a central role in orchestrating the immune response to alloantigens. They secrete cytokines to attract effector cells, such as macrophages and CD8+ T cells, into the graft and are able to interact with B cells that will secrete highly specific alloreactive antibodies. In clinical terms, the result of the immune response to transplanted allografts can be classified as hyperacute rejection, acute and chronic rejection. The immunological effector mechanisms involved in each of these processes are discussed. PMID- 11256421 TI - Indirect allorecognition in solid organ transplantation. AB - Recipient T cell recognition of donor major histocompatibility complex (MHC) alloantigens plays a central role in both acute and chronic rejection of human organ allografts. Two different pathways of T cell recognition of donor MHC alloantigens have been described. The direct pathway involves T cell recognition of intact MHC molecules expressed by donor antigen-presenting cells (APCs). The second, or indirect pathway, operates via T helper cell recognition of peptides derived from the processing and presentation of allogeneic MHC molecules on self APCs. At the onset of primary acute rejection, recipient CD4+ T cell responses to donor HLA-DR alloantigens are limited to a single dominant determinant present on one of the disparate alloantigens and restricted by one of the responder's HLA-DR molecules. In allograft recipients with recurring episodes of rejection, and/or at the onset of chronic rejection, recipient T cell reactivity may spread to other epitopes within the allogeneic MHC molecule, as well as to other alloantigens expressed by graft tissue. Both quantitative and qualitative alterations in T cell allopeptide reactivity are associated with increased risk of cellular and/or humoral rejection. These studies provide a basis for the design of new therapeutic strategies and for immunologic monitoring of transplant recipients. PMID- 11256422 TI - Role of alloimmunity in clinical transplantation. AB - HLA-specific humoral immunity, as a result of recipient allosensitization, induces hyperacute rejection of allogeneic kidney grafts. Cross-match tests are performed to avoid this complication. However, present techniques do not allow determination of HLA specificity of donor-reactive antibodies in the acute necro donor situation. New methods are described and discussed in this communication as well as the alloantibody specificities which are of clinical importance. Alloantibodies not only mediate hyperacute rejection, but may also participate in the acute rejection of organ grafts. Clinical associations between early immunological complications, such as acute rejection, in heart, liver and kidney allografted patients and pre-transplantation humoral alloimmunity emphasize the need for proper determination of donor-specific humoral immunity prior to transplantation. Acute rejection episodes are associated with an increased risk of subsequent chronic rejection. Cytomegalovirus (CMV) infection is also an important risk factor for chronic organ graft rejection as well as for chronic graft-versus-host (GvH) disease in bone marrow transplanted patients. CMV infection triggers autoantibody formation against CD13 in transplanted patients which, in turn, has been shown to be associated with the development of chronic GvH disease. Recently, alloactivation of immune reactivity in vitro was found to induce reactivation of latent CMV infection. We present here a hypothesis to explain the series of clinically associated events: acute rejection--CMV infection--chronic rejection. PMID- 11256423 TI - Identification of high and low responders to allografts. AB - The immune response to an allograft varies from one individual to another. This individual variation is, at least in part, due to genetic variation in the regulation of cytokine gene expression. High and low cytokine responses in vitro for tumour necrosis factor-alpha (TNF-alpha), transforming growth factor-beta 1 (TGF-beta1) and other cytokines can be predicted from an individual's cytokine genotype. Using the same genetic markers we have been able to show associations, in particular between TNF-alpha genotype of the recipient and acute allograft rejection and, similarly, between TGF-beta1 genotype and chronic rejection. The ability to identify high and low responders to allografts by a simple genetic test, and to predict who will suffer acute and chronic rejection, has implications for donor selection and recipient treatment as well as for the design and interpretation of clinical trials of new immunosuppressive agents. PMID- 11256424 TI - HLA compatibility and organ transplant survival. Collaborative Transplant Study. AB - The influence of HLA compatibility on organ transplant survival was analyzed in more than 150,000 recipients transplanted from 1987 to 1997 at transplant centers participating in the Collaborative Transplant Study. A statistically highly significant effect of HLA matching on graft and patient survival rates was found in the analysis of kidney transplants (P < 0.0001). Ten years after transplantation, the graft survival rate of first cadaver kidney transplants with a complete mismatch (6 HLA-A+B+DR mismatches) was 17% lower than that of grafts with no mismatch. During the first post-transplant year, the class II HLA-DR locus had a stronger impact than the class I HLA-A and HLA-B loci. During subsequent years, however, the influence on graft survival of the three loci was found to be equivalent and additive. For optimal graft outcome, compatibility at all three HLA loci is, therefore, desirable. The excellent correlation of HLA matching observed in recipients of cadaver kidneys with very short ischemic preservation (0-6 hours) or recipients of kidneys from living unrelated donors contradicts reports that short ischemia can eliminate the influence of matching. Although HLA has a significant effect on graft outcome regardless of the state of presensitization, the matching effect is potentiated in patients with highly reactive preformed lymphocytotoxic antibodies. Among first cadaver transplant recipients with an antibody reactivity against > 50% of the test panel, the difference in graft survival at 5 years between patients with 0 or 6 mismatches reached 30%. A collaborative project, in which molecular DNA typing methods were employed, showed that the correction of serological HLA typing errors by more accurate DNA typing results in a significantly improved HLA matching effect. Moreover, matching for the class II locus HLA-DP, a locus that can be typed reliably only by DNA methods, showed a significant effect in cadaver kidney retransplants, especially in the presence of preformed lymphocytotoxic antibodies. The analysis of heart transplants showed a highly significant impact of HLA compatibility on graft outcome (P < 0.0001). This result is of particular interest because donor hearts are not allocated according to the HLA match. A biasing influence of donor organ allocation (i.e. a preferential allocation of good matches to good risk recipients) can, therefore, be excluded. In liver transplantation, neither matching for HLA class I nor HLA class II could be shown to influence transplant outcome. PMID- 11256425 TI - The role of HLA matching in renal transplantation: experience from one center. AB - The influence of serology-based HLA matching on the risk of acute rejection episodes and of graft loss was analyzed in a material of 678 living donor (LD) and 997 cadaveric donor (CD) renal transplantations performed in our center in the period 1989-97. In LD transplantation, recipients of HLA-identical sibling grafts had the lowest rejection risk and the best graft survival, with a half life estimate of 30 years. One-HLA-haplotype mismatched grafts did better than two-haplotype mismatched related or unrelated donor grafts. Matching for HLA-DR significantly reduced the rejection risk of one-haplotype mismatched grafts. In CD first transplants, HLA-DR matched grafts had a lower incidence of rejection and better survival than HLA-DR mismatched grafts. Expected half-life for HLA-DR matched grafts was 12 years compared to less than 7 years for HLA-DR mismatched grafts. The effects of matching for HLA-A and -B did not reach statistical significance. In CD regrafts, a two-antigen mismatch for HLA-A or -DR led to a significantly poorer graft survival, but the panel-reactive antibody (PRA) status of the recipient was the most influential factor. In CD renal transplantation, we conclude that organ allocation based on matching for HLA-DR 1-14 is effective and not too difficult to obtain even in centers with a short patient waiting list. PMID- 11256426 TI - Acceptable HLA mismatches for highly immunized patients. AB - Highly sensitized patients have developed antibodies against many different HLA antigens due to previous pregnancies, blood transfusions or failed transplants. These antibodies cause a positive crossmatch with almost all potential organ donors. As a positive crossmatch is a contra-indication for transplantation, highly sensitized patients have a low chance of transplantation unless special strategies are introduced. One such strategy is the acceptable mismatch program, which has led to transplantation of more than 300 of these highly sensitized patients within Eurotransplant. Centers are participating in the program on a voluntary basis. Before a patient can be included in this program, extensive antibody screening is necessary to define those HLA-A and -B antigens towards which the patient has never formed antibodies. Organ donor selection is based on complete compatibility with the patients own HLA antigens in combination with the acceptable mismatches. If such a combination is identified, mandatory exchange takes place. Despite the success of the acceptable mismatch program, about 25% of the patients will never receive a donor offer. These are patients with rare HLA antigens or rare combinations of HLA antigens. In the last few years, this group of patients has had the advantage of two additional programs running within Eurotransplant. In the HIT (highly immunized tray) program, sera of highly sensitized patients are sent to the different centers and crossmatched with all ABO compatible donors. In the case of a negative crossmatch, mandatory exchange takes place. Secondly, these patients can benefit from the extra points they receive for their waiting time, high antibody reactivity and rare HLA type in the standard Eurotransplant allocation system. We conclude that the application of these three strategies will lead to a significantly increased transplantation rate of highly sensitized patients. PMID- 11256427 TI - The role of HLA matching in hematopoietic cell transplantation. AB - Hematopoietic cell transplantation (HCT) can be a life-saving therapy for patients with genetic and acquired hematologic diseases. Despite major advances in supportive care during HCT, immunological complications of the alloimmune response, including graft rejection and graft-versus-host disease (GVHD), remain major impediments to successful clinical outcomes. Although graft rejection mediated by host immune cells and GVHD mediated by donor immune cells can be prevented or mediated by immune suppression therapy, genetic HLA matching remains essential for successful strategies designed to minimize the risks of transplantation. The most favorable HCT results are seen in patients with a genotypically HLA-identical sibling donor, but the limited availability of matched related donors has severely restricted the clinical application of this therapy. Fortunately, the establishment of large unrelated volunteer donor registries now provides the opportunity to identify HLA matches for many patients who lack a family donor. The criteria for unrelated donor matching, however, are poorly defined. Until recently, an analysis of matching beyond HLA-identical siblings has been limited by typing technology. The introduction within the past few years of new methods for high resolution typing and definition of HLA alleles has had a profound impact on our ability to identify and interpret the multiple nucleotide sequence polymorphisms that encode HLA antigens. Preliminary studies clearly demonstrate the importance of precise matching at the allele level for successful transplantation. There remain, however, important unanswered questions about the relative importance of different HLA loci in matching strategies, as well as incomplete information about permissible limits of mismatching in different patient populations. PMID- 11256428 TI - Induction of tolerance to alloantigen. AB - The study of transplantation tolerance has been a major area of immunological research since the pioneering work by Medawar and colleagues. It has been classically defined as the absence of an immune response to a specific antigen in the setting of an otherwise normal immune system. The induction of tolerance to alloantigen in the transplantation setting would not only allow better graft survival but would also obviate the need for immunosuppressive therapy. The induction of tolerance is considered by many to be the "holy grail" of organ transplantation but, despite many years of experimental investigation, reliable induction of allograft tolerance has not been achieved in humans. There exists four fundamental mechanisms of T cell tolerance that may be operational in the transplantation setting: deletion, functional anergy, ignorance and regulation/suppression. Several strategies to induce tolerance in transplantation have been employed, but most of these tolerogenic strategies have been investigated in rodents and it remains to be determined whether they can be transferred into large outbred animal models or clinical transplantation. PMID- 11256429 TI - Evolution and development at the National Science Foundation. PMID- 11256430 TI - Evolution and development of gastropod larval shell morphology: experimental evidence for mechanical defense and repair. AB - The structural diversity of gastropod veliger larvae offers an instructive counterpoint to the view of larval forms as conservative archetypes. Larval structure, function, and development are fine-tuned for survival in the plankton. Accordingly, the study of larval adaptation provides an important perspective for evolutionary-developmental biology as an integrated science. Patterns of breakage and repair in the field, as well as patterns of breakage in arranged encounters with zooplankton under laboratory conditions, are two powerful sources of data on the adaptive significance of morphological and microsculptural features of the gastropod larval shell. Shells of the planktonic veliger larvae of the caenogastropod Nassarius paupertus [GOULD] preserve multiple repaired breaks, attributed to unsuccessful zooplankton predators. In culture, larvae isolated from concentrated zooplankton samples rapidly repaired broken apertural margins and restored the "ideal" apertural form, in which an elaborate projection or "beak" covers the head of the swimming veliger. When individuals with repaired apertures were reintroduced to a concentrated mixture of potential zooplankton predators, the repaired margins were rapidly chipped and broken back. The projecting beak of the larval shell is the first line of mechanical defense, covering the larval head and mouth and potentially the most vulnerable part of the shell to breakage. Patterns of mechanical failure show that spiral ridges do reinforce the beak and retard breakage. The capacity for rapid shell repair and regeneration, and the evolution of features that resist or retard mechanical damage, may play a more prominent role than previously thought in enhancing the ability of larvae to survive in the plankton. PMID- 11256431 TI - Wnt gene expression in sea urchin development: heterochronies associated with the evolution of developmental mode. AB - The Wnt genes encode a large family of conserved secreted proteins that are widely involved in animal development. The variety and ubiquity of this ancient family suggest that Wnt genes may have been important in the evolution of animal development, including early development. To test this hypothesis, we have characterized the expression of several Wnt genes in closely related sea urchins that exhibit radically different modes of early development. Wnt-1, -4, and -5 genes exhibit several conserved molecular and developmental characteristics, both within sea urchins and with Wnt genes examined in other animals (Ferkowicz et al. 1998). Here, we demonstrate that sea urchin Wnt-5 transcripts are specifically detected by in situ hybridization in discrete embryonic, larval, and developing adult tissues and processes: (1) in a band of vegetal ectoderm in mesenchyme blastula stage embryos, (2) in the larval ciliary bands, (3) in tissues that form the early adult rudiment (left coelomic pouch and overlying vestibular ectoderm), and (4) in the developing adult radial nervous system. We find that the sites of Wnt-5 transcript accumulation are conserved in species exhibiting either indirect or direct-developmental modes, suggesting that Wnt-5 function(s) have been conserved in sea urchin development. However, dramatic heterochronic changes in Wnt-5 gene expression have occurred in the direct-developing species that parallel the accelerated morphological changes that occur during direct development. These results suggest that heterochronic changes in the expression of conserved developmental regulatory genes, such as the Wnt family members, are agents of evolutionary change in animal development. PMID- 11256432 TI - Comparative development of fiber in wild and cultivated cotton. AB - One of the most striking examples of plant hairs is the single-celled epidermal seed trichome of cultivated cotton. The developmental morphology of these commercial "fibers" has been well-characterized in Gossypium hirsutum, but little is known about the pattern and tempo of fiber development in wild Gossypium species, all of which have short, agronomically inferior fiber. To identify developmental differences that account for variation in fiber length, and to place these differences in a phylogenetic context, we conducted SEM studies of ovules at and near the time of flowering, and generated growth curves for cultivated and wild diploid and tetraploid species. Trichome initiation was found to be similar in all taxa, with few notable differences in trichome density or early growth. Developmental profiles of the fibers of most wild species are similar, with fiber elongation terminating at about two weeks post-anthesis. In contrast, growth is extended to three weeks in the A- and F-genome diploids. This prolonged elongation period is diagnosed as a key evolutionary event in the origin of long fiber. A second evolutionary innovation is that absolute growth rate is higher in species with long fibers. Domestication of species is associated with a further prolongation of elongation at both the diploid and allopolyploid levels, suggesting the effects of parallel artificial selection. Comparative analysis of fiber growth curves lends developmental support to previous quantitative genetic suggestions that genes for fiber "improvement" in tetraploid cotton were contributed by the agronomically inferior D-genome diploid parent. PMID- 11256433 TI - Geographic patterning of variation in segment number in geophilomorph centipedes: clines and speciation. AB - Since their origin as a metameric group, arthropods have diversified considerably in their number of segments. Present-day geophilomorph centipedes provide a model system for investigating the evolutionary origins of this diversification, because they exhibit intraspecific variation in segment number. (This variation is, however, derived; it is not a plesiomorphic condition within the Chilopoda.) Previous studies have shown that there are significant differences in segment number between populations within several geophilomorph species. In one (arguably two) species, it has been demonstrated that there is a particular form of geographic patterning of the variation, namely a latitudinal cline, with the segment number decreasing with increasing distance north. Here, we provide additional data on four more species, all of which show evidence of a latitudinal cline in either one or both sexes. It is therefore becoming clear that this is a general phenomenon, applying widely (perhaps universally) across the Geophilomorpha, a group consisting of some 1,000 known species. It may be that latitudinal clines are a frequent part of the speciation cycle in this group. PMID- 11256434 TI - Etienne Geoffroy St.-Hilaire: father of "evo-devo"? AB - In the early decades of the nineteenth century, the most important disagreement among comparative anatomists was not evolution versus "special creation" but between advocates of "transcendental morphology" and those of teleological anatomy-form versus function. In France this dichotomy was represented by the 1830-1832 public debate between Geoffroy St.-Hilaire (form) and Cuvier (function). Geoffroy's aim was to establish links of homology (known to him as "analogies") between the four "embranchements" into which Cuvier had divided the animal kingdom. Despite the fanciful nature of some of his homologies, Geoffroy, who was guided by his "principe de connections," set in motion a school of morphology, some of whose conclusions, notably the homology of the dorsal surface of segmented invertebrates with the ventral surface of vertebrates, has been corroborated by recent studies in developmental genetics. PMID- 11256435 TI - Excitotoxic lesions of the pre- and parasubiculum disrupt object recognition and spatial memory processes. AB - Rats with bilateral ibotenic acid lesions centered on the pre- and parasubiculum and control rats were tested in a series of spatial memory and object recognition memory tasks. Lesioned rats were severely impaired relative to controls in both the reference and working memory versions of the water maze task and displayed a delay-dependent deficit in a delayed nonmatch to place procedure conducted in the T-maze. Lesioned rats also displayed reduced exploration in a novel environment, and performance was altered in an object recognition procedure as compared with the control group. These findings indicate that the pre- and parasubiculum plays an important role in the processing of both object recognition and spatial memory. PMID- 11256436 TI - Extracellular serotonin is enhanced in the striatum, but not in the dorsal hippocampus or prefrontal cortex, in rats subjected to an operant conflict procedure. AB - In rats trained in an operant fixed-interval-30-s schedule of food reward (FI 30s), acute exposure to contingent footshock resulted in a response suppression that was released by diazepam (DZP; 4 mg/kg ip) but not by buspirone (0.25 or 0.50 mg/kg ip). Compared with baseline, hippocampal and cortical extracellular levels of serotonin (5-HText) did not change, regardless of operant period (punished or nonpunished) and drug. In contrast, in the striatum, an increase of 5-HText levels (535%) occurred during the punished period, counteracted by DZP. This effect was observed only in rats that were low responders during both nonpunished and punished periods, that is, those that exerted an efficacious control over responding. Uncontrollable shocks or exposure to an unfamiliar open field did not modify striatal 5-HText. Together, these results suggest that an acute activation of 5-HT neurons afferent to the striatum allows the rats to efficiently block responses that are negatively reinforced. PMID- 11256438 TI - Attenuation of ethanol-induced conditioned taste aversion in mice sensitized to the locomotor stimulant effects of ethanol. AB - This study examined the effect of repeated ethanol (EtOH) injections that induced behavioral sensitization on subsequent acquisition of EtOH- and lithium chloride (LiCl)-induced conditioned taste aversion (CTA). CTA acquisition was assessed in independent groups of EtOH-sensitized and nonsensitized genetically heterogeneous female mice after injections of saline; 1, 2, or 4 g/kg EtOH; or 2 or 4 mEq/kg LiCl. Saline and 1 g/kg EtOH did not induce CTA. Four g/kg EtOH and 4 mEq/kg LiCl induced similar levels of CTA in EtOH-sensitized and nonsensitized groups. CTA induced by 2 g/kg EtOH and 2 mEq/kg LiCl was attenuated in EtOH-sensitized mice compared with nonsensitized counterparts. Thus, a sensitizing regimen of EtOH preexposure resulted in both a decrease in EtOH and LiCl aversion and an increase in EtOH locomotor sensitivity; such changes could ultimately contribute to enhanced EtOH intake and potentially to EtOH abuse. PMID- 11256437 TI - Critical periods for the effects of alcohol exposure on learning in rats. AB - Critical periods for alcohol-induced deficits in spatial navigation and passive avoidance learning were investigated with a rat model of fetal alcohol syndrome. Rats were exposed to alcohol prenatally (Gestational Days 1-10 or 11-22) or postnatally (Postnatal Days 2-10) or throughout all 3 periods. Offspring were tested in either a spatial navigation or an avoidance task as juveniles or adults. As juveniles, the combined exposure group took longer to learn the spatial navigation task compared with all other groups. This effect was not seen in adults. Passive avoidance performance was not affected. These results suggest that long-term exposure to alcohol during development has adverse effects on spatial learning. The lack of differences in the short-term exposure groups implies that there may not be 1 critical period of alcohol exposure, but that the adverse effects of alcohol during development may be cumulative on some behaviors. PMID- 11256439 TI - Recovery of associative function following early amygdala lesions in rats. AB - Adult rats with amygdala lesions made at either Postnatal Day (PND) 10 or PND40 were tested on a series of reversal tasks that tap the ability to form stimulus reward associations. PND40 rats were significantly impaired relative to both controls and PND10 rats on learning rate of the original discrimination and subsequent reversals. Analyses of discrete learning phases revealed that the impairment was specific to the postchance phase. The PND10 group was not impaired relative to controls on any measure. These results confirm prior findings that amygdala lesions sustained in adulthood impair the formation of stimulus-reward associations. They also demonstrate that substantial sparing or recovery of function is possible when the lesion is made during early development. Furthermore, the findings support the view that behavioral recovery may be more likely if the lesion is sustained near the time of peak synaptogenesis. PMID- 11256440 TI - Differential sensitivity to the wake-promoting actions of norepinephrine within the medial preoptic area and the substantia innominata. AB - Mapping studies were conducted to delineate the site(s) of action for the arousal enhancing actions of norepinephrine (NE) within the basal forebrain region encompassing the medial preoptic area (MPOA) and the substantia innominata (SI). Varying doses of NE, the beta-agonist, isoproterenol, or the alpha1-agonist, phenylephrine, were infused into the MPOA or SI in the resting rat. Infusions of NE (4 nmol, 16 nmo/150 nl), isoproterenol (15 nmol/150 nl), and phenylephrine (40 nmol/250 nl) into the MPOA elicited robust increases in waking. In contrast, neither isoproterenol or phenylephrine infusions into the SI altered behavioral state. NE infusions into the SI increased waking only at the highest dose, and at this dose there was an anatomical gradient for NE-induced waking, with infusions placed farther from the MPOA, producing smaller increases in waking. Thus, in contrast to the MPOA, the SI is relatively insensitive to the wake-promoting actions of NE. PMID- 11256441 TI - The contribution of adrenal and reproductive hormones to the opposing effects of stress on trace conditioning in males versus females. AB - Exposure to an acute stressful experience facilitates classical conditioning in male rats but impairs conditioning in female rats (T. J. Shors, C. Lewczyk, M. Paczynski, P. R. Mathew, & J. Pickett, 1998; G. E. Wood & T. J. Shors, 1998). The authors report that these effects extend to performance on the hippocampal dependent task of trace conditioning. The stress-induced impairment of conditioning in females was evident immediately, 24 hr and 48 hr after stress, depending on the stage of estrus. Moreover, the effect could be reactivated days later by reexposure to the stressful context. Corticosterone levels correlated with overall performance in males but not in females. Unlike the effect seen in males, adrenalectomy did not prevent the stress-induced effect on conditioning in females. These data indicate that exposure to the same experience can have opposite effects on learning in males versus females and that these opposing effects are mediated by differing hormonal systems. PMID- 11256442 TI - Dissociation of licking and volume intake controls in rats ingesting glucose and maltodextrin. AB - The volume of fluid that rats acquire with each lick was systematically varied across short-term tests with 12.5% glucose (Experiment 1) or 12.5% maltodextrin (Experiment 2). For glucose, rats increased the number of licks emitted as lick volume was reduced such that meal size remained remarkably stable across all (8, 4, and 2 microl) but the smallest (1 microl) lick volume conditions tested. Rats similarly compensated for lick volume reduction (8 to 4 microl) with maltodextrin by approximately doubling the number of licks emitted. Meal duration and a number of lick-microstructural parameters (initial ingestion rate, mean burst duration, terminal lick and ingestion rates, and burst duration) were not correlated with the intake outcome insofar as they varied significantly across conditions over which intake remained stable. Thus, in response to lick volume manipulation, rats demonstrated an impressive degree of behavioral flexibility in what may be regarded as a defense of meal size. PMID- 11256443 TI - Adult sex differences on a decision-making task previously shown to depend on the orbital prefrontal cortex. AB - Monkeys and children show sex differences on tasks that depend on the orbital prefrontal cortex. To determine whether similar sex differences exist across the life span, adults were tested on an orbital-dependent decision-making task, the Iowa Card Task, as well as on a control task, the California Weather Task. In addition, estradiol, progesterone, and testosterone were assayed. The 6 groups of participants were college-age men, older men, young low-hormone (menstruating) women, young high-hormone (midluteal) women, older postmenopausal women on estrogen replacement therapy (ERT), and older postmenopausal women not on ERT. Results showed a male superiority on the Iowa Card Task. Among college-age men there was a negative correlation between performance and testosterone levels. There were no significant differences among groups of women on the card task. There were no significant sex differences or hormone correlations on the California Weather Task. PMID- 11256444 TI - Conditioned activity to amphetamine in transgenic mice expressing an antisense RNA against the glucocorticoid receptor. AB - Glucocorticoids enhance the locomotion-stimulating and the rewarding properties of stimulant drugs. Amphetamine-induced conditioned activity was investigated in B6C3F1 (controls) and antisense transgenic mice. The latter expresses a neurofilament-promotor-driven antisense RNA complementary to a fragment of cDNA that codes for the mouse glucocorticoid receptor. This gene expression leads to approximately a 50% reduction in glucocorticoid receptor mRNA in the brain. Transgenic mice showed an increased novelty response when tested in an open field, in terms of both distance traveled and number of rearings. Moreover, they displayed enhanced amphetamine-induced conditioned activity. Behavioral sensitization was observed in controls, whereas behavioral tolerance developed in transgenic mice. These data support the concept of an enhanced stress response in these transgenic mice, rather than a general downregulation of the stress response because of impaired glucocorticoid receptor function. PMID- 11256445 TI - A two-platform task reveals a deficit in the ability of rats to return to the start location in the water maze. AB - The ability of rats to return to the start location was examined with a 4-arm radial water maze. The task required rats to find 2 hidden platforms in sequence. Rats were released from 1 of 3 arms and there was a platform located in the fourth arm. Once a rat found this platform, a 2nd platform was raised in another location, which was either the start location, for 1 group, or another fixed location, for a control group. Across 3 experiments, all rats learned the location of the 1st fixed platform in 80 to 120 trials. However, rats had difficulty finding a 2nd platform if it was at the start location. Control groups revealed that rats could learn 2 platform locations and that the difficulty in learning to return to the start location did not seem to be attributable to its aversive nature. In separate groups, exposure to the start location was increased by starting the rats from an initially stable platform. Rats still did not readily learn to return to the start location. The authors suggest that start location, when varied, cannot readily be used to define the location of a hidden platform. PMID- 11256446 TI - Spatial reference memory and neocortical neurochemistry vary with the estrous cycle in C57BL/6 mice. AB - Estrous cycle-related variations of spatial reference memory and neurochemistry in intact female mice were examined. Spatial reference memory was tested in cycling females, ovariectomized (OVX) females, and males by using a 1-day water maze protocol. Choline acetyltransferase (ChAT) and glutamic acid decarboxylase (GAD) activities were measured in the hippocampus and neocortex. Estrus females exhibited worse spatial acquisition and 30-min retention than did proestrus and metestrus females, higher neocortical ChAT activity than proestrus females, and higher neocortical GAD activity than OVX females and males. Neocortical, rather than hippocampal, neurochemistry was more sensitive to hormonal modulation, suggesting that hormonal mediation of neocortical function may play a critical role in regulating spatial reference memory in female mice. PMID- 11256447 TI - A single administration of testosterone induces cardiac accelerative responses to angry faces in healthy young women. AB - Recently, it was demonstrated how individuals with high levels of testosterone selectively attend toward angry faces. It was argued that this suggests that high levels of testosterone are associated with an aggressive, dominating personality style. In this study, the authors used a double-blind, placebo-controlled design to examine whether exogenous testosterone would induce cardiac acceleration in response to angry faces. Participants (healthy young women) were exposed to neutral, happy, or angry faces. Administration of a single dosage of testosterone (0.5 mg) induced an accelerative cardiac response to angry faces. It is argued that this effect is due to the encouragement of dominance behavior and the inclination toward aggression. Possible mechanisms behind testosterone-driven changes in behavior are discussed with relevance to steroid-responsive networks in the limbic system that drive and control motivational and physiological aspects of social behavior. PMID- 11256448 TI - Cannabinoid modulation of time estimation in the rat. AB - Cannabinoids have been implicated in a variety of cognitive processes in humans, including attention, learning, memory, and time estimation. However, studies of the effects of cannabinoids on rodent behavior have focused on motor, learning, and memory tasks. To assess cannabinoid effects on time perception, this study examined whether systemically administered cannabinoid receptor agonists and a cannabinoid receptor antagonist influenced rats' performance of a time interval estimation task based on a fixed-interval schedule (a "peak procedure"). Both cannabinoid agonists WIN 55,212-2 and delta9-tetrahydrocannabinol shortened the modal response time, and cannabinoid antagonist SR 141716A lengthened the modal response time. Secondary measures of the shape of the response distribution were not influenced by any of the drugs, suggesting that the response distribution shifts were not artifacts of drug side effects. Therefore, these experiments argue for the involvement of endogenous cannabinoids in time estimation. PMID- 11256449 TI - Sex differences, context preexposure, and the immediate shock deficit in Pavlovian context conditioning with mice. AB - The acquisition of context fear in rats is affected by variables such as the sex of the animal, the placement to shock interval (PSI), and preexposure to the context. The current experiments assessed the effects of these variables on context conditioning in mice (C57BL/6). In Experiment 1, mice were placed in a chamber and received a single shock 5s, 20 s, 40s, 60s, 180s, or 720s later. Increasing the PSI produced corresponding increases in conditional freezing during the context test. In addition, male mice acquired more context conditioning than female mice did but only at intermediate PSIs. In Experiment 2, preexposure to the context before training alleviated the sex difference found with an intermediate PSI. The results are discussed in terms of configural learning theory and are argued to be contrary to the predictions of scalar expectancy theory. PMID- 11256450 TI - Head direction, place, and movement correlates for cells in the rat retrosplenial cortex. AB - The retrosplenial cortex is strongly connected with brain regions involved in spatial signaling. To test whether it also codes space, single cells were recorded while rats navigated in an open field. As in earlier work (L. L. Chen, L. H. Lin, C. A. Barnes, & B. L. McNaughton, 1994; L. L. Chen, L. H. Lin, E. J. Green, C. A. Barnes, & B. L. McNaughton, 1994), the authors found head direction cells with properties similar to those in other areas. These cells were slightly anticipatory. Another cell type fired to particular combinations of location, direction, and movement, which suggested that they may fire whenever the rat approaches a particular location, using a particular locomotor behavior. The remaining cells could not be clearly categorized but also showed a significant correlation with one or more of the spatial-movement variables examined. The fact that the retrosplenial cortex contains spatial and movement-related signals and is connected with the motor cortex suggests that it may play a role in path integration or navigational motor planning. PMID- 11256451 TI - An fMRI study of personality influences on brain reactivity to emotional stimuli. AB - Functional imaging studies have examined which brain regions respond to emotional stimuli, but they have not determined how stable personality traits moderate such brain activation. Two personality traits, extraversion and neuroticism, are strongly associated with emotional experience and may thus moderate brain reactivity to emotional stimuli. The present study used functional magnetic resonance imaging to directly test whether individual differences in brain reactivity to emotional stimuli are correlated with extraversion and neuroticism in healthy women. Extraversion was correlated with brain reactivity to positive stimuli in localized brain regions, and neuroticism was correlated with brain reactivity to negative stimuli in localized brain regions. This study provides direct evidence that personality is associated with brain reactivity to emotional stimuli and identifies both common and distinct brain regions where such modulation takes place. PMID- 11256452 TI - Conditional discriminations based on external and internal cues in rats with cytotoxic hippocampal lesions. AB - Septal-hippocampal system lesions, mostly using aspiration techniques, have been reported to impair performance of conditional tasks. Rats with axon-sparing cytotoxic hippocampal lesions were therefore tested in a range of instrumental conditional paradigms. They did not differ from controls in their ability to choose the appropriate object in a conditional object discrimination cued by internal state (hunger or thirst) or on performance of conditional visuospatial object discriminations. Acquisition of a conditional visuospatial discrimination with black and white boxes as stimuli was also unimpaired. In contrast, lesioned rats were profoundly impaired on an open T-maze task when cued by either their internal state (reference memory task) or their previous response (working memory task). The results indicate that perception or use of spatial cues, rather than conditional responding per se, is impaired by cytotoxic hippocampal lesions. PMID- 11256453 TI - Effects of neonatal RU486 on adult sexual, parental, and fearful behaviors in rats. AB - Exposure to gonadal hormones during perinatal life influences later behavior. The finding that sex differences exist in progestin receptor expression in the perinatal rat brain suggests differential sensitivity of male and female brains to progesterone (C. K. Wagner, A. N. Nakayama, & G. J. De Vries, 1998). Because these sex differences are in neural sites that influence sexually differentiated sexual, parental, and fearful behaviors in adults, this study examined the effects of administering the progestin receptor antagonist RU486 for the first 10 days after birth on these behaviors in adulthood. Neonatal RU486 significantly reduced sexual behavior in males but did not impair reproduction in females. Neonatal RU486 did not affect parental responses of virgin rats exposed to pups (sensitization) but reduced fear in the elevated plus-maze in both sexes. Treatment of pups with RU486 affected neither mother-litter interactions nor plasma testosterone levels in males during or after treatment. These results suggest that neonatal exposure to progesterone, in addition to androgens and estrogens, influences behavioral development in rats. PMID- 11256454 TI - Comparison of the effects of infant handling, isolation, and nonhandling on acoustic startle, prepulse inhibition, locomotion, and HPA activity in the adult rat. AB - This study examined whether early isolation (EI), early handling (EH), or early nonhandling (NH) in infant rats alters (a) prepulse inhibition (PPI) of the acoustic startle response (ASR) or its disruption by apomorphine, (b) motor activity or its stimulation by amphetamine, or (c) corticosterone activity (because of its modulation of dopamine activity), in adulthood and in comparison with a normal-husbandry postnatal control environment. EI did not affect PPI, reduced PPI disruption by apomorphine in males, and increased amphetamine stimulated activity in males. NH increased the ASR, reduced activity in the open field, and increased corticosterone reactivity in males. In all paradigms, the effects of EH were similar to those of the control environment. This study provides an important contribution to the evidence on the relationship between postnatal experience and long-term neurobehavioral development in the rat and the relevance of this approach to animal models of neuropsychiatric disorder. PMID- 11256455 TI - Conditioned flavor preference and aversion: role of the lateral hypothalamus. AB - Food-restricted rats with ibotenic acid lesions of the lateral hypothalamus (LH) and sham controls were trained to associate flavored solutions with positive or negative postingestive consequences. The LH rats learned to prefer a flavor that was paired with concurrent intragastric infusions of maltodextrin. Unlike controls, the LH rats failed to learn a preference for a flavor paired with delayed maltodextrin infusions and showed an attenuated preference for a flavor paired with concurrent fat infusions. The LH rats did not differ from controls in learning to avoid flavors paired with concurrent or delayed infusions of lithium chloride. These data indicate that the LH is not essential for all types of flavor-postingestive consequence conditioning but is critical for learning to associate flavors with delayed nutrient feedback. PMID- 11256456 TI - The effect of excitotoxic lesions centered on the hippocampus or perirhinal cortex in object recognition and spatial memory tasks. AB - Rats with bilateral ibotenic acid lesions centered on the hippocampus (HPC) or perirhinal cortex (PRC) and sham-operated controls were tested in a series of object recognition and spatial memory tasks. Both HPC and PRC rats displayed reduced habituation in a novel environment and were impaired in an object location task. HPC rats were severely impaired in both the reference and working memory versions of the water maze and radial arm maze tasks. In contrast, although PRC rats displayed mild deficits in the reference memory version of the water maze and radial arm maze tasks, they were markedly impaired in the working memory version of both the tasks. These findings demonstrate that under certain conditions both the HPC and PRC play a role in the processing of spatial memory. Further investigation of these conditions will provide important new insights into the role of these structures in memory processes. PMID- 11256457 TI - Dosimetry bibliography. PMID- 11256458 TI - Cognitive-behavioral therapy: applications for the management of bipolar disorder. AB - OBJECTIVES: This paper reviews cognitive-behavioral therapy (CBT) for bipolar disorder (BD). Data on the poor outcome of about 50% of patients diagnosed with BD supports the addition of a psychosocial intervention for the treatment of this recurring disorder. The psychoeducational nature of CBT, the effectiveness of CBT in increasing compliance to pharmacological treatment, and the ability of CBT to prevent relapse in unipolar depression (UD) are well suited to the treatment of BD. METHOD: Psychosocial interventions for BD will be briefly reviewed. Individual and group CBT interventions (published and unpublished) will also be reviewed. The significance of comorbid anxiety disorders regarding response to treatment will also be discussed. A review of the treatment protocol with the specific cognitive-behavioral interventions as applied to BD will be presented. Finally, a case example will be presented to illustrate the application of CBT to BD. RESULTS: Preliminary results indicate that CBT may be an effective adjunctive, intervention for the treatment of BD. Specifically CBT may be helpful in increasing compliance, improving quality of life and functioning, help early symptom recognition, decrease relapse and decrease depressive symptomatology. CONCLUSIONS: Preliminary data on CBT for BD are promising but more rigorous randomized clinical trials are needed to confirm the efficacy of CBT for BD. An other area of research should be to pursue the understanding of cognitive processes in BD which would allow us to refine and develop CBT interventions unique to this disorder. PMID- 11256459 TI - Psychotherapy for bipolar depression: a phase-specific treatment strategy? AB - OBJECTIVES: The depressive phase of bipolar disorder is particularly difficult to treat. Pharmacologic strategies for bipolar depression are often inadequate. We therefore review the literature on the role of psychotherapy as an adjunct to medication in the treatment of bipolar depression. METHODS: With one exception, there are no descriptions of psychotherapies employed specifically for the treatment of bipolar depression. We therefore reviewed published reports of psychotherapy for bipolar disorder in general and extracted from these reports relevant data or impressions about the specific effects of the therapies on the depressive phase of the disorder. RESULTS: Described psychosocial approaches to bipolar disorder include psychoeducation, group therapy, cognitive-behavioral therapy, couples therapy, family therapy, and interpersonal psychotherapy. Only cognitive-behavioral therapy has been tested in a pilot study for the treatment of bipolar depression specifically. Results from randomized controlled trials of family therapy and interpersonal and social rhythm therapy suggest that these treatments may be more efficacious in the treatment and prevention of depression relative to mania. CONCLUSIONS: A limited number of well-designed studies and preponderance of case reports limit definitive conclusions about the role of psychotherapy in the treatment of bipolar depression. However, converging reports suggest that cognitive-behavioral therapy, family therapy, and interpersonal and social rhythm therapy may be particularly useful for bipolar depression. We propose a novel approach to the treatment of bipolar disorder that includes the use of phase-specific sequenced psychotherapies delivered in variable patterns and linked to fluctuating mood states. PMID- 11256460 TI - Ketoconazole in bipolar patients with depressive symptoms: a case series and literature review. AB - BACKGROUND: Data from several studies suggest that medications, such as ketoconazole, which lower cortisol levels, may be effective for major depressive disorder (MDD). As with MDD, the manic, depressive, and mixed phases of bipolar disorder are frequently associated with elevated cortisol levels. The literature on the use of cortisol-lowering strategies in mood disorders is reviewed, and a case series illustrating the use of ketoconazole in bipolar depression is presented. METHODS: For the review, the MEDLINE and PSYCHINFO databases were searched, as were the bibliographies of pertinent articles to find papers on the use of cortisol-lowering agents in patients with mood disorders. In our open label case series (n = 6), ketoconazole (up to 800 mg/day) as an add-on therapy was given to patients with treatment-resistant or intolerant bipolar I or II disorders with current symptoms of depression. RESULTS: Several case reports and small open studies suggest that cortisol-lowering agents may be useful for patients with depression. Two recent placebo-controlled trials of ketoconazole on patients with MDD report conflicting results. In our case series, all three patients who received a dose of at least 400 mg/day had substantial reductions in depressive symptoms. None had significant increases in mania. However, cortisol levels were not lowered in any of the subjects. CONCLUSIONS: The literature suggests that cortisol-lowering medications may be effective for a subset of depressed patients. Our preliminary findings suggest that ketoconazole may be useful in some patients with bipolar depression. Larger clinical trials are needed to confirm our observations. PMID- 11256462 TI - The clinical epidemiology of pure and mixed manic episodes. AB - INTRODUCTION: Few large clinical epidemiological studies have been undertaken comparing subjects meeting criteria for mixed and pure states of bipolar disorder. In part, the difficulty comparing these states emanates from confusion in their diagnostic separation. In the current report, we use a definition derived from receiver operating characteristic (ROC) curve analysis as an alternative to the DSM-IIIR/IV definition, and we compare the two subtypes of manic episodes. METHODS: Three hundred and sixty-six patients meeting DSM-IIIR criteria for bipolar disorder, manic or mixed, were categorized using newly described criteria for mixed states. The two subtypes were compared on demographic variables and clinical history variables, using multiple analysis of variance with post hoc univariate F tests. The same analyses were conducted using the DSM-IIIR-defined subtypes. RESULTS: Using the ROC criteria, 79 subjects (21.6%) were characterized as mixed, in contrast to 51 subjects (13.9%) using DSM IIIR criteria for bipolar disorder, mixed. The ROC-defined mixed manic group comprised more Caucasians and more females. Age of first psychiatric hospitalization was earlier and duration of illness longer in the mixed group. First episodes were unlikely to be categorized as mixed (< 5%). When the DSM-IIIR definition was employed, differences were not demonstrated. CONCLUSIONS: An earlier age of first psychiatric hospitalization and increased duration of illness, as well as a lower frequency of mixed subtype of manic episode during first hospitalization, are compatible with the view that mixed manic episodes occur more frequently later in the course of bipolar disorder. Moreover, differences in race, sex, and clinical histories of subjects in mixed episodes tend to support the separation of mixed mania as a diagnostic subtype of bipolar disorder. PMID- 11256461 TI - Transcranial magnetic stimulation in an amphetamine hyperactivity model of mania. AB - OBJECTIVES: Transcranial magnetic stimulation (TMS) of the brain has been reported to have therapeutic effects in mania, as well as depression. TMS has previously been reported to have effects similar to those of electroconvulsive shock in rat models of depression. METHODS: We, therefore, studied TMS in amphetamine-induced hyperactivity as a rat model of mania. RESULTS: While two and seven daily TMS sessions significantly reduced activity after amphetamine, twice daily TMS for 7 days enhanced activity after amphetamine. CONCLUSIONS: The results suggest that TMS treatment to rats interacts with the effects of amphetamine; the specific effects may be dependent on the schedule of treatment. PMID- 11256463 TI - Illness characteristics and their association with prescription patterns for bipolar I disorder. AB - INTRODUCTION: The present study explores the relationships among psychotropic medications, illness-related parameters, patient demography, suicidality, and levels of functioning in a voluntary bipolar case registry. METHODS: Four hundred and fifty-seven subjects with bipolar I disorder were selected from a voluntary registry for subjects with bipolar illness. Demographic characteristics, psychotropic medications, age at onset of illness, duration of illness, number of hospitalizations, the ability to live independently, employment and driving status as well as the history of suicidal attempts were obtained through a structured phone interview. RESULTS: Subjects treated with antidepressants had a shorter duration of illness, while patients treated with antipsychotic drugs had an earlier onset of illness. The number of hospitalizations for mania was fewer among patients taking a combination of lithium and carbamazepine as compared to patients not receiving them, while subjects taking neuroleptics had more hospitalizations as compared to subjects not receiving them. The number of psychotropic agents prescribed correlated positively with the number of hospitalizations for depressive episodes. Curiously, no correlations were found between the types of psychotropic agents prescribed and the levels of functioning or a history of suicidal attempts. Interestingly, our results suggest that more than half of the subjects were unable to live independently or to work due to their illness. Also, more than 50% of the subjects had at least one suicidal attempt, the majority occurred during depressive episodes. CONCLUSIONS: Our results suggest that subjects with bipolar I disorder have high rate of suicidal attempts and may have serious functional impairments. PMID- 11256464 TI - Valproate and the risk of hyperandrogenism. PMID- 11256465 TI - Gabapentin-induced anorgasmia as a cause of noncompliance in a bipolar patient. PMID- 11256467 TI - Ultrastructural changes in the pulmonary arterioles in acute hypoxic pulmonary hypertension in the rat. AB - This study was planned to obtain ultrastructural details of the early changes in intra-acinar arterioles in acute hypoxic pulmonary hypertension that could lead to understanding the mechanisms in the development of chronic hypoxic hypertension. In the anesthetized rat, using 5-10% normobaric O2, within minutes after hypoxia, there are changes in endothelial cells characteristic of activation: prominence of cell body and protuberance of the nucleus, electron dense membrane-bound bodies adluminally, increasing pseudopodia of the adluminal cell membrane, edema within (vacuoles) and beneath the endothelial cells with separation of the endothelial cells from the basal lamina. There is activation of platelets and leucocytes in the lumen and accumulation of platelets at the endothelium. Arteriolar wall edema rapidly increases, is excessive within 1 h, with dissection of the basal lamina and wall and cytolysis of wall components. At 24 h edema is reduced, the number of platelets is increased at the endothelium and fibroblasts are newly aligned within the arteriolar wall. At 48 h platelets further increase, a basal lamina develops in fibroblasts termed transitional cells and myofibrils occur subsequently to form smooth muscle. These findings suggest that activation of the endothelial cell is the initial event in a cellular cascade in the arteriolar hypoxic responses with fibroblast-to-smooth muscle transformation, which results in pulmonary arteriolar hyperplasia and vascular remodeling in hypoxic chronic pulmonary hypertension. PMID- 11256466 TI - Effects of a 21-day expedition to 6,194 m on human skeletal muscle SR Ca2+ ATPase. AB - We investigated the effects of a 21-day expedition to the summit of Mount Denali, Alaska (6,194 m) on selected Ca2+ sequestration properties of sarcoplasmic reticulum (SR) calcium pump in vastus lateralis muscle. Muscle samples were obtained by biopsy from 5 male climbers (peak oxygen consumption, VO2peak = 52.3 +/- 2.1 mL.kg(-1).min(-1)) approximately 7 days prior to (PRE) and 4 days following (POST) the expedition. A comparison of PRE versus POST measures of maximal Ca2+-ATPase activities (117 +/- 8.5 vs. 97.6 +/- 5.6 nmol.mg protein( 1).min(-1)) and Ca2+-uptake (204 +/- 15 vs. 161 +/- 11 nmol.mg protein(-1).min( 1)) measured in crude homogenates obtained from pre-exercised muscle, indicated only an effect (p < 0.05) of the expedition on Ca2+-uptake. The reduction in Ca2+ ATPase activity, representing 16.6%, was not significant (p = 0.089). The sarco endoplasmic reticulum calcium (SERCA)-ATPase isoforms, measured using Western blotting techniques, revealed a small reduction (p < 0.05) in SERCA 1 (-4.6 +/- 1.9%), but not in SERCA 2a (+2.0 +/- 1.4%). Prior to the expedition, both Ca2+ ATPase activity and Ca2+-uptake were reduced (p < 0.05) by approximately 34 and 18%, respectively, following 40 min of a two-step continuous cycling task (20 min at 59% VO2peak and 20 min at 74% VO2peak). The exercise-induced reduction in Ca2+ ATPase activity was independent of fiber type. Only in the case of Ca2+-uptake was a lower exercise response (p < 0.05) observed following the expedition, an effect that was due to the lower resting value. It is concluded that acclimatization as experienced during a mountaineering expedition induces changes in the properties of the SR Ca2+-pump, and particularly to Ca2+-sequestering function. PMID- 11256468 TI - Cold-induced peripheral vasodilation at high altitudes--a field study. AB - A significant reduction in cold-induced vasodilation (CIVD) is observed at high altitudes. No agreement is found in the literature about acclimatization effects on CIVD. Two studies were performed to investigate the effect of altitude acclimatization on CIVD. In the first study 13 male subjects immersed the distal phalanx of the left middle finger in water of 0 degrees C for 30 min to evoke CIVD. Five subjects were exposed to altitudes of 5,100 to 7,000 m for 45 days (A). Eight subjects were exposed to an altitude of 5,100 m for <3 days (NA). The groups did not differ in age, weight, and stature. No significant differences were observed between A and NA. However, the maximum finger skin temperature of group A tended to return to sea level values (6.9 +/- 3.2 degrees C at sea level vs. 6.0 +/- 0.7 degrees C at altitude), while a strong reduction was observed for the NA group (7.7 +/- 4.3 degrees C vs. 3.7 +/- 3.1 degrees C). This indicates that the CIVD response at altitude tended to be stronger for the acclimatized subjects. In a second study, nine males were followed in a longitudinal study. CIVD was measured before, during and after 7 days of exposure to 4,350 m. Maximum finger skin temperature before and after exposure did not differ (8.5 +/- 2.6 degrees C vs. 7.8 +/- 1.6 degrees C), and was reduced at altitude. There was no difference in maximum finger skin temperature between the 7 days at altitude (e.g., 5.3 +/- 2.7 degrees C at day 2 and 4.7 +/- 1.1 degrees C at Day 7). It can be concluded that no acclimatization effects of CIVD occur during the first 7 days of altitude exposure, but that differences may occur after altitude exposure of several weeks. PMID- 11256469 TI - The quality of sleep and periodic breathing in healthy subjects at an altitude of 3,200 m. AB - The medical risks of travel and stay at high altitude are well known. Many more people travel for recreation to lower but still significant altitudes. To investigate the quality of sleep and sleep-related breathing disorders (SRBD) at that altitude we performed full polysomnography in nine young volunteers at lowland (760 m above sea level) on the first and sixth night after ascent to 3,200 m. There have been few studies on such populations. The subjects were nonsmoking healthy males aged 20.3 +/- 3.5 years with normal spirometry and arterial blood gas measurements performed at low altitude. Although there was no statistically significant difference in the duration of stages and sleep quality between low altitude night and both nights at high altitude as assessed by percent of sleep spent in stage 1, 2, 3+4 NREM, and REM sleep, total sleep time (TST), and sleep efficiency; the number of arousals and awakenings doubled at high altitude. There was no periodic breathing (PB) during sleep, except in isolated central events of SRBD, at low altitude. PB appeared at altitude mostly during NREM sleep and its intensity remained stable throughout the study period. Individual variations of PB intensity were high, ranging from 0.1 to 24% of TST. There were also some episodes of obstructive apnea and hypopnea during sleep at high altitude (p < 0.001). Mean SaO2 was lower during the study nights at high altitude when compared with low altitude. There were some signs of ventilatory acclimatization as shown by a higher mean SaO2 during the sixth compared with the first night at altitude (p < 0.001). We conclude that the sleep quality at the altitude of 3,200 m remains satisfactory when compared to low altitude. There is high individual variability in intensity of PB at that altitude. PMID- 11256470 TI - Subarachnoid cyst and ascent to high altitude--a problem? AB - A 31-year-old man suffered diplopia and ataxia on two occasions when he ascended from sea level to 4,000 m. Evaluation revealed a moderate-sized subarachnoid cyst in the left frontal region, which did not communicate with the cerebral ventricles. The cyst might have acted as a space-occupying lesion, and caused symptoms on ascent due to hypoxic brain swelling, brain compression against the cyst, and elevated intracranial pressure. Subarachnoid cysts are common, and they should be considered in the differential diagnosis of neurological problems at high altitude. PMID- 11256471 TI - Antioxidant activities of buckwheat hull extract toward various oxidative stress in vitro and in vivo. AB - We have undertaken four basic in vitro studies and an animal experiment to obtain information about the antioxidant activities of buckwheat hull extract (BWHE). In the in vitro studies, BWHE scavenged super oxide anion produced in the xanthine/xanthine oxidase system (IC50=11.4 microg phenolic compound/ml), and strongly inhibited autoxidation of linoleic acid (IC50=6.2 microg phenolic compound/ml). Low-density lipoprotein (LDL) oxidation induced by Cu2+ ion was also protected by BWHE. In the animal experiment, ddY mice were fed a standard diet supplemented with 0.75% BWHE for 14 d. In blood, liver and brain of the mice TBARS and fluorescent substance concentration were significantly decreased compared with those of non-treated mice. SOD like activity in serum also significantly rose by BWHE treatment. BWHE was shown to be effective for protecting biological systems against various oxidative stresses in vitro, and to have antioxidant activity in vivo. PMID- 11256472 TI - Quantitative measurement of the novel human plasma protein, IHRP, by sandwich ELISA. AB - Inter-alpha-trypsin inhibitor family heavy chain-related protein (IHRP), which has a sequence similarity to the heavy chains of the inter-alpha-trypsin inhibitor (ITI) family, is a novel glycoprotein found in human plasma. We prepared two clones (1A4 and 6E11) of anti-IHRP mouse monoclonal antibody. Both of them recognized the C-terminal 35-kDa fragment which was produced by plasma kallikrein-digestion of IHRP. We developed a sandwich ELISA for measurement of the plasma IHRP concentration with the monoclonal antibody coated microtiter plate and the anti-N-terminal 57-kDa fragment of IHRP rabbit polyclonal antibody (anti-GP57). We found that the average concentration of IHRP in the plasma of healthy donors was 101.3+/-31.8 microg/ml (average+/-S.D.). The IHRP concentration in the plasma of patients with inflammatory disorders was slightly increased (137.5+/-40.2 microg/ml: average+/-S.D.). Together with the previous data indicating the induction of the porcine mRNA, which is thought to be the species counterpart of human IHRP mRNA, in the liver after resuscitation from cardiogenic shock, we propose that IHRP is a member of the acute-phase protein family. PMID- 11256473 TI - Hypocholesterolemic effect of bile acid sulfonate analogs in hamsters. AB - We studied the effects of bile acid sulfonate analogs, namely, 3alpha,7alpha,12alpha-trihydroxy-5beta-cholane-24-sulfonate (C-sul), 3alpha,7alpha-dihydroxy-5beta-cholane-24-sulfonate (CDC-sul), and 3alpha,7beta dihydroxy-5beta-cholane-24-sulfonate (UDC-sul), on serum and liver cholesterol levels, cholesterol 7alpha-hydroxylase activity, and biliary bile acid composition in hamsters fed cholesterol. Of the three analogs studied, UDC-sul slightly but significantly decreased free, esterified, and total cholesterol concentrations in the serum. UDC-sul and CDC-sul reduced liver total cholesterol levels by 25% and 18%, respectively, particularly in the esterified cholesterol fraction. Analysis of biliary bile acids showed the presence of the administered analogs, indicating that sulfonate analogs efficiently participate in enterohepatic cycling. The proportion of cholic acid was increased in all groups fed sulfonate analogs, but the ratio of glycine to taurine conjugated bile acids (G/T) was elevated only in UDC-sul feeding hamsters. There was no significant change in cholesterol 7alpha-hydroxylase activity in hamsters fed C-sul or CDC sul, while UDC-sul slightly stimulated the enzyme activity compared to the control. The UDC-sul induced decrease in serum and liver cholesterol concentrations may be secondary to enhanced bile acid synthesis. This is supported by the increased cholesterol 7alpha-hydroxylase activity and elevated G/T ratio in biliary bile acids observed following UDC-sul administration. PMID- 11256474 TI - Regulation mechanism of the serine protease activity of plasma hyaluronan binding protein. AB - The inhibitor for the serine protease activity of plasma hyaluronan binding protein (PHBP) was purified from human plasma by polyethylene glycol (PEG) fractionation, diethylaminoethyl (DEAE)-Sephacel ion-exchange chromatography, Phenyl Toyopearl 650M hydrophobic chromatography, Bio Gel A-0.5 m gel-filtration and hydroxyapatite chromatography. The serine protease activity of PHBP was measured with Boc-Phe-Ser-Arg-methylcoumarine amide (MCA) as the synthetic substrate of PHBP. The results of the amino acid sequence analyses of the purified PHBP inhibitor indicated that it was C1 inhibitor of the serpin family. C1 inhibitor formed a complex with PHBP, suggesting that it is the actual inhibitor of PHBP in human plasma. On the other hand, dextran sulfate and phosphatidylethanolamine enhanced the auto-fragmentation and the serine protease activity of pro-PHBP, but kaolin did not. These results suggested that the serine protease activity of PHBP was regulated in a similar manner to that of factor XII of the coagulation system. PMID- 11256475 TI - Inhibition of aminopeptidase N (AP-N) and urokinase-type plasminogen activator (uPA) by zinc suppresses the invasion activity in human urological cancer cells. AB - Zinc is an essential heavy metal and is more abundant in human prostate and kidney than in other tissues. The effects of zinc on the invasion activity of human prostate and renal cancer cell lines, PC-3, LNCaP and SKRC-1, were investigated in vitro using a Transwell cell-culture chamber and were compared with specific protease inhibitors for MMPs, uPA and AP-N, respectively. The invasion activity of PC-3 cells was effectively suppressed by zinc and by all protease inhibitors in a dose-dependent manner. The invasion activity of LNCaP cells was almost unaffected by these inhibitors. In SKRC-1 cells, the invasion activity was strongly suppressed by MP03, although a moderate inhibition by zinc and bestatin was observed. The purified AP-N activity was strongly inhibited by zinc at a concentration similar to that suppressing the invasion activity of PC-3 cells and this inhibition by zinc was apparently competitive. Although the purified uPA activity was also inhibited by zinc, this inhibition was uncompetitive. AP-N was expressed abundantly on the membrane fraction of PC-3 cells among these cells tested, while its expression on the membrane fraction of SKRC-1 cells was weaker than that of PC-3 cells. The expression of uPA was also highest on the membrane fraction of PC-3 cells. These results suggest that AP-N and uPA may be involved in the invasion of human prostate cancer cells and that zinc probably participates in the invasion and metastasis of cancer cells through the regulation of the enzymatic activity of AP-N and uPA in human cancerous prostate. PMID- 11256476 TI - Pur alpha protein implicated in dendritic RNA transport interacts with ribosomes in neuronal cytoplasm. AB - We have previously reported that pur alpha, known to be a regulator of DNA replication and transcription, links neural BC1 RNA to microtubules via dendrite targeting RNA motifs. Here we demonstrate the subcellular localization of pur proteins within the brain. Pur proteins were detected in neurons but not in glia. Immunohistochemical staining was prominent in perikarya and proximal dendrites and also extended into primary dendritic processes, but no significant signals were detected in the distal regions of dendrite. When homogenates of mouse brain were fractionated, pur alpha was most concentrated in the microsomal pellet. Consistently, pur alpha co-fractionated with free polysomes as well as with membrane-bound polysomes and the association with polysomes was mediated by binding ribosomal subunits. Levels of ribosomes with pur alpha progressively increased during postnatal development of the brain. PMID- 11256477 TI - Selective nonpeptidic inhibitors of herpes simplex virus type 1 and human cytomegalovirus proteases. AB - The proteases encoded by herpesviruses including herpes simplex virus type 1 (HSV 1) and human cytomegalovirus (HCMV) are attractive targets for antiviral drug development because of their important roles in viral replication. We randomly screened a chemical compound library for inhibitory activity against HSV-1 protease. 1,4-Dihydroxynaphthalene and three naphthoquinones were found to be potent inhibitors of HSV-1 protease with IC50 values of 6.4 to 16.9 microM. Inhibitory mode analysis of the compounds against HSV-1 protease suggested that, in spite of structural similarities, only 1,4-dihydroxynaphthalene was a competitive inhibitor, whereas the three naphthoquinones were noncompetitive inhibitors. Among all assayed dihydroxynaphthalene derivatives in the chemical compound library, 1,4-dihydroxynaphthalene proved to be the most potent inhibitor of HSV-1 protease. Therefore, the two hydroxyl groups located at positions 1 and 4 on the naphthalene structure seemed essential for exertion of a potent inhibitory activity against HSV-1 protease. In addition, we have found that these compounds are also potent inhibitors of HCMV protease with extremely low micromolar IC50 values. This differed from the results of inhibitory mode analysis of HSV-1 protease, 1,4-dihydroxynaphthalene was a noncompetitive inhibitor of HCMV protease, and three naphthoquinones were competitive inhibitors. These compounds showed no effective inhibitory activity against several mammalian serine proteases (trypsin, chymotrypsin, kallikrein, plasmin, thrombin and Factor Xa) at 100 microM. These results suggest that 1,4 dihydroxynaphthalene and three naphthoquinones may be useful in the development of nonpeptidic antiherpesvirus agents. PMID- 11256478 TI - The biological activities of 1alpha,25-dihydroxyvitamin D3 and its synthetic analog 1alpha,25-dihydroxy-16-ene-vitamin D3 in normal human osteoblastic cells and human osteosarcoma SaOS-2 cells are modulated by 17-beta estradiol and dependent on stage of differentiation. AB - We compared the effects of 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3] and its analog, 1alpha,25-dihydroxy-16-ene-vitamin D3 [1alpha,25(OH)2-16-ene-D3], as well as their interactions with 17-beta estradiol (E2) on osteoblastic function in our human normal (HOB) and osteosarcoma SaOS-2 cell models representing two different stages of differentiation, the more differentiated HOB+DEX cells and SaOS+DEX cells, and the corresponding less differentiated HOB-DEX and SaOS-DEX cells. The differential effects of 1alpha,25(OH)2D3 and 1alpha,25(OH)2-16-ene-D3 and the modulation by E2 on ALP activity in HOB-DEX and HOB+DEX cells were small but significant. The most significant effects were seen in SaOS+DEX cells, in which 1alpha,25(OH)2-16-ene-D3 was 100-fold more potent than 1alpha,25(OH)2D3, the maximal enhancement being exerted at 0.1 nM and 10 nM, respectively. E2 enhanced the stimulatory effects of both compounds, with ALP being increased 2 fold at 0.1 nM (p<0.001). Osteocalcin (OC) production in HOB-DEX cells was stimulated 1.3 to 1.4-fold by 1alpha,25(OH)2D3 and 1alpha,25(OH)2-16-ene-D3 at a concentration of 0.01 nM, with E2 inhibiting the effect of 1alpha,25(OH)2-16-ene D3. In SaOS-DEX and SaOS+DEX cells, 1alpha,25(OH)2D3 and 1alpha,25(OH)2-16-ene-D3 stimulated OC production 1.6-fold at 0.1 nM with E2 slightly enhancing the effect of 1alpha,25(OH)2D3. Western blot analysis of 1alpha,25(OH)2D3 receptor (VDR) levels showed that in SaOS+DEX cells, the effect of 1alpha,25(OH)2D3 was larger than that of 1alpha,25(OH)2-16-ene-D3. These results show that 1alpha,25(OH)2-16 ene-D3 is biologically active in human osteoblasts. PMID- 11256479 TI - Different generation of inhibitors against gallic acid-induced apoptosis produces different sensitivity to gallic acid. AB - Gallic acid (3,4,5-trihydroxybenzoic acid), a naturally occurring plant phenol, showed selective cytotoxicity against tumor cells with higher sensitivity than normal cells such as hepatocytes and keratinocytes. To elucidate the difference in sensitivity between normal and tumor cells to gallic acid, we studied whether the inhibitor of gallic acid-induced apoptosis existed or not. A serum-free conditioned medium, prepared from high density rat primary cultured hepatocytes and cytoplasm of hepatocytes, prevented gallic acid-induced apoptosis. In contrast, hepatomas and hepatic cell lines such as dRLh-84, PLC/PRF/5, HLE, and HUH and two other kinds of tumor cell, HeLa and KB, scarcely generated such an inhibitor in either their conditioned medium or their cells. Biochemical characterization of the inhibitors revealed that the inhibitor in the hepatocyte conditioned medium was completely inactivated by heating at 65 degrees C for 10 min. Its molecular weight was estimated at 150-250 kDa by gel filtration column chromatography, indicating that the inhibitor may be a protein-like substance. These results suggest that the generation of a large amount of the inhibitor may endow hepatocytes with insensitivity to gallic acid. In conclusion, the difference in the amount of the inhibitors generated by hepatocytes and tumor cells should contribute to the underlying mechanism in the difference in sensitivity of cells to gallic acid. PMID- 11256480 TI - Pharmacological effects of elenoside, an arylnaphthalene lignan. AB - Acute toxicity (24 h) and general behavior in mice of a lignan from Justicia hyssopifolia, a beta-D-glucoside (elenoside), was studied, and the cytotoxic activity was performed. Elenoside (arylnaphthalene lignan) in mice showed a moderate toxicity order (305 mg/kg) and central depressive properties at doses of 25, 50, and 100 mg/kg. It also displayed cytotoxic activity in a range of concentration of 10(-5)-10(-4) M when studied in the human tumor cell line panel of the US National Cancer Institute (NCI). The results indicated that elenoside has central depressant effects, and the cytotoxic activity of elenoside suggests that this compound and its genin derivatives merit further investigation as antitumoral drugs. PMID- 11256481 TI - Anti-androgenic activity of Myricae Cortex--isolation of active constituents from bark of Myrica rubra. AB - The aqueous ethanol extract of Myricae Cortex (bark of Myrica rubra Sieb. et Zucc., Myricaceae) showed in vitro testosterone 5alpha-reductase inhibitory activity and in vivo anti-androgenic activity using growth of flank organ in castrated Syrian hamsters and/or hair regrowth after shaving in testosterone treated C57Black/6CrSlc mice. Three constituents, myricanone, myricanol, and myricetin were identified as the main active principles. PMID- 11256482 TI - Study on anti-Oketsu activity of rhubarb II. Anti-allergic effects of stilbene components from Rhei undulati Rhizoma (dried rhizome of Rheum undulatum cultivated in Korea). AB - Methanol extract (RM-ext) obtained from the dried rhizome of Rheum undulatum was screened for activity in experimental models of type I allergy. RM-ext exhibited the inhibition on 48-h homologous passive cutaneous anaphylaxis (PCA) in rats and an antigen-induced histamine release from rat peritoneal mast cells. Among nine stilbenes isolated from RM-ext, seven inhibited the histamine release. Rhapontigenin (compound 1), piceatannol (2) and piceatannol 3'-beta-D-glycoside (6) with oral administration showed the inhibition on PCA. Compounds 1 and 2 exhibited the inhibitory effect on sheep red blood cell-induced delayed-type hyper sensitivity (SRBC-DTH) of type IV allergic model. These results indicated that the rhizome of Rheum undulatum inhibits the allergic reactions and that these inhibitory effects may be partially attributable to the stilbenes mentioned above. PMID- 11256483 TI - Receptor mediated endocytosis and cytotoxicity of transferrin-mitomycin C conjugate in the HepG2 cell and primary cultured rat hepatocyte. AB - Intracellular disposition and cytotoxicity of macromolecular conjugate of mitomycin C (MMC) with transferrin (TF) were examined in the human hepatoma cell line HepG2 cell and normal cultured rat hepatocyte. The conjugate (TF-MMC) was specifically bound to the HepG2 cell as well as TF. The number of the binding site and the association constant of TF-MMC in the HepG2 cell were 396000+/-31000 molecules/cell and 3.24 x 10(7)+/-0.58 x 10(7) M(-1), respectively. No difference in the binding parameters of TF-MMC and TF can be detected in the HepG2 cell. The association constant for the TF receptor was almost identical between HepG2 cell and hepatocyte, however, the numbers of the binding site of TF-MMC and TF in the HepG2 cell were from 40-times to 50-times greater than those in the hepatocyte. Furthermore, TF-MMC was internalized into the HepG2 cell and the hepatocyte as well as TF. The rates of internalization of TF-MMC and TF into the HepG2 cell were nearly identical to those into the hepatocyte. However, the levels of the internalization into the HepG2 cell were remarkably higher than those into the hepatocyte because the number of receptors in the HepG2 cell was larger than that in the hepatocyte, and the rate of release from the HepG2 cell was slower than that from the hepatocyte. TF-MMC inhibited the growth of the HepG2 cells. The 50% growth inhibition (GI50) of TF-MMC against the HepG2 cell was 0.9 microg MMC/ml, which was a little higher than that of MMC (GI50=0.5 microg/ml). These results indicated that the TF-MMC might be useful for delivery of MMC to the HepG2 cell. PMID- 11256484 TI - Pharmacokinetic differences between lansoprazole enantiomers and contribution of cytochrome P450 isoforms to enantioselective metabolism of lansoprazole in dogs. AB - The purpose of this study was to evaluate the pharmacokinetics of lansoprazole enantiomers and contribution of cytochrome P450 enzymes to enantioselective metabolism in dogs. The mean Cmax and area under the curve (AUC) values of (+) lansoprazole were 4-5 times greater than those of (-)-lansoprazole following oral administration of 30-mg racemic lansoprazole to dogs. The CLtot/F values of (+) lansoprazole were significantly smaller than those of (-)-lansoprazole (p<0.05). The mean unbound fraction of (-)-lansoprazole was significantly greater than that of the (+)-lansoprazole. The amount of (+)-lansoprazole remaining was significantly greater than that of the (-)-lansoprazole after incubation of racemic lansoprazole in dog liver microsomes. When the effects of ticlopidine or ketoconazole on the metabolism of lansoprazole were studied using dog liver microsomes, ticlopidine significantly inhibited the formation of 5 hydroxylansoprazole, but not another metabolite, lansoprazole sulfone; however ketoconazole significantly inhibited formation of both metabolites. When the amount of (+)- and (-)-enantiomers remaining was measured in the presence and absence of ticlopidine, the amount of (+)-lansoprazole was significantly greater than that of the (-)-lansoprazole. On the other hand, there was no significant difference between the amount of (+)- and (-)-enantiomers remaining in combination with ketoconazole. These results suggest that the enantioselective pharmacokinetics of lansoprazole enantiomers are probably ascribable to their enantioselective protein binding and/or metabolism, and among the cytochrome P450 enzymes, CYP3A contributed to the enantioselective metabolism of lansoprazole. PMID- 11256485 TI - Effect of lipophilicity on in vivo iontophoretic delivery. I. NSAIDs. AB - The effect of drug lipophilicity on in vivo iontophoretic transdermal absorption was evaluated. Non-steroidal anti-inflammatory drugs (NSAIDs) were selected as model drugs with a wide range of lipophilicity: salicylic acid (SA), ketoprofen (KP), naproxen (NP) and indomethacin (IM). Cathodal iontophoresis of NSAIDs was conducted in rats (0.625 mA/cm2; 90 min), and drug concentrations in skin, cutaneous vein and systemic vein were determined. Skin concentrations of NSAID were higher in the case of lipophilic drugs (SA=KP=NPKP=NP>IM). Additionally, the dependence of drug lipophilicity on systemic plasma concentration was similar to cutaneous plasma concentration. The transfer rate from skin to cutaneous vein (R(SC)) was calculated from the arterio-venous plasma concentration difference of drug in the skin. Normalized R(SC) by skin concentration (R(SC)/X(S)) yielded a negative correlation with the logarithm of n octanol/buffer partition coefficient (Log P at pH 7.4), suggesting that transfer of NSAIDs from skin to cutaneous vein decreased with increasing lipophilicity (SA>KP=NP>IM). This correlation means that drug partitioning between stratum corneum and viable epidermis might be a dominant step. PMID- 11256486 TI - A Kampo formulation: Byakko-ka-ninjin-to (Bai-Hu-Jia-Ren-Sheng-Tang) inhibits IgE mediated triphasic skin reaction in mice: the role of its constituents in expression of the efficacy. AB - We have demonstrated that oral administration of a Kampo formulation, Byakko-ka ninjin-to (Bai-Hu-Jia-Ren-Sheng-Tang), inhibited IgE-mediated triphasic skin reaction, including immediate phase response (IPR), late phase response (LPR) and very late phase response (vLPR), in passively sensitized mice with anti-DNP IgE antibody. Variant formulations of Byakko-ka-ninjin-to without Gypsum Fibrosum (Sekko), Glycyrrhizae Radix (Kanzo) or Oryzae Semen (Kobei) attenuated the inhibitory effect as compared with that of Byakko-ka-ninjin-to. The decreased effect of Byakko-ka-ninjin-to without Kanzo was restored by the addition of Kanzo to the variant formulations before oral administration, while the decreased effect of Byakko-ka-ninjin-to without Sekko could not be recovered by the addition of Sekko. Comparison of HPLC profiles of variant formulations without one crude drug with that of original Byakko-ka-ninjin-to revealed that some peaks could be detected only when five constituent crude drugs were simultaneously present during the preparation of Byakko-ka-ninjin-to formulation. Since elimination of Sekko from the Byakko-ka-ninjin-to constituents attenuated the efficacy although it did not show any activity per se, mutual interaction of Sekko with other constituents during the preparation may result in the production of new components. These findings suggest that the effect of Byakko-ka-ninjin-to formulation on cutaneous inflammatory disease can differ from the sum of the effect of the individual constituents. PMID- 11256487 TI - Bi-directional relationship of in vitro mast cell-nerve communication observed by confocal laser scanning microscopy. AB - Communication between nerves and mast cells is a prototypic demonstration of neuro-immune interaction. Recently, we used an in vitro co-culture approach comprising cultured murine superior cervical ganglia (SCG) and rat basophilic leukemia (RBL) cells to study this interaction. Previously, we concentrated mainly on the activation signal from neurites to mast cells (RBL). However, it is proposed that mast cell-nerve communication is not a one-sided relationship but a bi-directional one. In the present work, we studied the communication from mast cells to neurites. We observed that binding of anti-IgE receptor antibodies to mast cells increases calcium ion concentration [Ca2+]i in SCG neurites. This indicates that mast cell-nerve communication is bi-directional. Confocal fluorescence microscopic images indicated that [Ca2+]i in neurites increased after an increase of [Ca2+]i in mast cells. The lag-time of neurite activation was several times longer than that of mast cell activation. The correlation coefficient between the lag-times for mast cell and nerve activation was calculated to be 0.81. In addition, the fluorescence images showed that calcium signals in SCG neurites were able to extend to a long distance (100-200 microm) from the site where mast cells (RBL) attached to neurites. PMID- 11256488 TI - Influence of sugar chain on fibrin affinity of recombinant t-PA. AB - The role of the sugar chain on the fibrin affinity property of tissue plasminogen activator (t-PA) was investigated using two variants of wild type t-PA (WT t-PA I and WT t-PA II) and mutant type t-PA (mt-PA ; Gln117 t-PA I and Gln117 t-PA II), whose sugar chains have different structures. In terms of fibrin affinity, Gln117 t-PA was higher than WT t-PA ; moreover, Type II was higher than Type I. Bindings mediated via finger domain (F mode) and kringle 2 domain (K2 mode) were distinguished using epsilon-amino caproic acid (EACA). Consequently, F mode and K2 mode bindings were inhibited by the sugar chains at Asn117 and 184, respectively. These results were assumed to be due to the steric hindrance of the sugar chains. PMID- 11256489 TI - Cytotoxicity of the hinokitiol-related compounds, gamma-thujaplicin and beta dolabrin. AB - Gamma-thujaplicin and beta-dolabrin, the constituents of the wood of Thujopsis dolabrata Sieb. et Zucc. var. hondai showed strong in vitro cytotoxic effects against the human stomach cancer cell lines KATO-III and Ehrlich's ascites carcinoma. The cytotoxic effects of the two compounds against both tumor cell lines were clear when cell growth was measured by the 3-(4,5-dimethylthiazol-2 yl)-2,5-diphenyltetrazolium bromide (MTT) method. Gamma-thujaplicin and beta dolabrin at 0.32 microg/ml inhibited cell growth of human stomach cancer KATO-III by 85 and 67%, and Ehrlich's ascites carcinoma by 91 and 75%, respectively. There is no large difference in cytotoxicity between these compounds, but the activity of gamma-thujaplicin was slightly more potent than that of beta-dolabrin. On the other hand, hinokitiol acetate did not show a cytotoxic effect, suggesting that at least a part of the mechanism of the cytotoxic effect of hinokitiol-related compounds is due to metal chelation between the carbonyl group at C-1 and the hydroxyl group at C-2 in the tropolone skeleton of these molecules. The acute toxicities [50% lethal dose (LD50) value: intraperitoneal injection, Van der Waedem] of gamma-thujaplicin and beta-dolabrin in mice were 277 mg/kg and 232 mg/kg, respectively. PMID- 11256490 TI - Costunolide induces apoptosis by ROS-mediated mitochondrial permeability transition and cytochrome C release. AB - Costunolide is an active compound isolated from the root of Saussurea lappa Clarks, a Chinese medicinal herb, and is considered a therapeutic candidate for various types of cancers. Nevertheless, the pharmacological pathways of costunolide are still unknown. In this study, we investigate the effects of costunolide on the induction of apoptosis in HL-60 human leukemia cells and its putative pathways of action. Using apoptosis analysis, measurement of reactive oxygen species (ROS), and assessment of mitochondrial membrane potentials, we show that costunolide is a potent inducer of apoptosis, and facilitates its activity via ROS generation, thereby inducing mitochondrial permeability transition (MPT) and cytochrome c release to the cytosol. ROS production, mitochondrial alteration, and subsequent apoptotic cell death in costunolide treated cells were blocked by the antioxidant N-acetylcystein (NAC). Cyclosporin A, a permeability transition inhibitor, also inhibited mitochondrial permeability transition and apoptosis. Our data indicate that costunolide induces the ROS mediated mitochondrial permeability transition and resultant cytochrome c release. This is the first report on the mechanism of the anticancer effect of costunolide. PMID- 11256491 TI - Hematological studies on black cumin oil from the seeds of Nigella sativa L. AB - The methanol soluble portion of black cumin oil, which is prepared by compression of seeds of Nigella sativa L., showed inhibitory effects on arachidonic acid (AA) induced platelet aggregation and blood coagulation. By bioactive assay of AA induced platelet aggregation, the methanol soluble part was purified to isolate a new compound 2-(2-methoxypropyl)-5-methyl-1,4-benzenediol (1) and two known compounds, thymol (2), carvacrol (3), having very strong inhibitory activity. Further, we then examined the isolated compounds (1-3) and eight related compounds by the screening test for AA-induced platelet aggregation. Compounds possessing aromatic hydroxyl and acetoxyl group had more potent activity than aspirin, which is well known as a remedy for thrombosis. PMID- 11256492 TI - Antiviral amentoflavone from Selaginella sinensis. AB - Amentoflavone and three other flavonoids were isolated from the ethanol extract of Selaginella sinensis. Amentoflavone showed potent antiviral activity against respiratory syncytial virus (RSV), with an IC50 of 5.5 microg/ml. The contents of amentoflavone in nine species of Selaginella were determined by reversed-phase HPLC. S. sinensis showed a higher content of 1.13%. PMID- 11256493 TI - Assessment of gastric acidity of Japanese subjects over the last 15 years. AB - The gastric acidity of young to elderly Japanese subjects from 1989 to 1999 was assessed and compared with that obtained in 1984, using GA-Test capsules containing acid-dissolving granules of riboflavin. The percentage of achlorhydric subjects increased with age as observed before, however, an over all decrease in all age categories year by year was noted. The percentage of achlorhydric subjects aged 50 years in 1995-1999 was about 40%, which was lower than that (60%) in 1984. However, such a chronological change was not observed when the percentage of achlorhydric subjects was determined according to birth year, indicating that it is related to the birth year of subjects. The percentage of achlorhydric subjects correlated with infection by Helicobacter pylori. Considering the high percentage of achlorhydric elderly, bioavailability and bioequivalence studies should be performed taking into consideration the effects of gastric acidity on the in vivo performance of drug products. PMID- 11256494 TI - Anti-metastatic effect of the sialyl Lewis-X analog GSC-150 on the human colon carcinoma derived cell line KM12-HX in the mouse. AB - We investigated the inhibitory effect of the sialyl Lewis-X (sLeX) analog, GSC 150, on hepatic metastasis of the human colon carcinoma derived cell line, KM12 HX, which highly expresses sLeX antigen on the cell surface. The number of cancer nodules found in BALB/c nude mouse liver 6 weeks after intrasplenic injection of KM12-HX cells was significantly reduced by co-administration of GSC-150. The amount of [3H]thymidine-labeled KM12-HX cells distributed in liver was also significantly reduced by GSC-150 co-administration in lipopolysaccharide (LPS) treated mice at 48 h after administration of the tumor cells, while GSC-150 did not reduce the amount of HX cells distributed at 30 min. Considering our previous report that the initial phase of the distribution of KM12-HX cells in liver is governed by their being trapped in the hepatic microvessels because of their large size (Mizuno et al., J. Hepatol., 28, 865-877, 1998), these results suggest that GSC-150 does not inhibit this first-pass trapping by microvessels, but inhibits the subsequent process which is more directly related to final metastasis. GSC-150 inhibited the adhesion of KM12-HX cells to tumor necrosis factor-alpha (TNF-alpha)-activated human umbilical vein endothelial cells (HUVECs). These findings imply that the anti-metastatic effect of GSC-150 in vivo could be explained by its inhibition of cell-cell interactions between cancer and host cells. PMID- 11256495 TI - Biotreatment of hydrogen sulfide- and ammonia-containing waste gases by fluidized bed bioreactor. AB - Gas mixtures of H2S and NH3 are the focus of this study of research concerning gases generated from animal husbandry and treatments of anaerobic wastewater lagoons. A heterotrophic microflora (a mixture of Pseudomonas putida for H2S and Arthrobacter oxydans for NH3) was immobilized with Ca-alginate and packed into a fluidized bed reactor to simultaneously decompose H2S and NH3. This bioreactor was continuously supplied with H2S and NH3 separately or together at various ratios. The removal efficiency, removal rate, and metabolic product of the bioreactor were studied. The results showed that the efficiency remained above 95% when the inlet H2S concentration was below 30 ppm at 36 L/hr. Furthermore, the apparent maximum removal and the apparent half-saturation constant were 7.0 x 10(-8) g-S/cell/day and 76.2 ppm, respectively, in this study. The element sulfur as a main product prevented acidification of the biofilter, which maintained the stability of the operation. As for NH3, the greater than 90% removal rate was achieved as long as the inlet concentration was controlled below 100 ppm at a flow rate of 27 L/hr. In the NH3 inlet, the apparent maximum removal and the apparent half-saturation constant were 1.88 x 10(-6) g-N/cell/day and 30.5 ppm, respectively. Kinetic analysis showed that 60 ppm of NH3 significantly suppressed the H2S removal by Pseudomonas putida, but H2S in the range of 5-60 ppm did not affect NH3 removal by Arthrobacter oxydans. Results from bioaerosol analysis in the bioreactor suggest that the co-immobilized cell technique applied for gas removal creates less environmental impact. PMID- 11256496 TI - Air sampling and analysis of volatile organic compounds with solid phase microextraction. AB - Solid phase microextraction (SPME) presents many advantages over conventional analytical methods by combining sampling, preconcentration, and direct transfer of the analytes into a standard gas chromatograph (GC). Since its commercial introduction in the early 1990s, SPME has been successfully applied to the sampling and analysis of environmental samples. This paper presents an overview of the current methods for air sampling and analysis with SPME using both grab and time-weighted average (TWA) modes. Methods include total volatile organic compounds (TVOCs), formaldehyde, and several target volatile organic compounds (VOCs). Field sampling data obtained with these methods in indoor air were validated with conventional methods based on sorbent tubes. The advantages and challenges associated with SPME for air sampling are also discussed. SPME is accurate, fast, sensitive, versatile, and cost-efficient, and could serve as a powerful alternative to conventional methods used by the research, industrial, regulatory, and academic communities. PMID- 11256497 TI - Meteorological effects on the evolution of high ozone episodes in the greater Seoul area. AB - Three high O3 episodes--7 days in 1992 (July 3-July 9), 9 days in 1994 (July 21 July 29), and another 3 days in 1994 (August 22-August 24)--were selected on the basis of morning (7:00 a.m.-10:00 a.m.) average wind direction and speed and daily maximum O3 concentrations in the greater Seoul, Korea, of 1990-1997. To better understand their characteristics and life cycles, surface data from the Seoul Weather Station (SWS) and surface and 850-hPa wind field data covering northeast Asia around the Korean Peninsula were used for the analysis. In the July 1992 episode, westerly winds were most frequent as a result of the influence of a high-pressure system west of the Korean Peninsula behind a trough. In contrast, in the July 1994 episode, easterly winds were most frequent as a result of the effect of a typhoon moving north from the south of Japan. Despite different prevailing wind directions, the peak O3 concentrations for each episode occurred when a sea/land breeze developed in association with weak synoptic forcing. The August 1994 episode, which was selected as being representative of calm conditions, was another typical example in which a well-developed PMID- 11256498 TI - Air pollution and health: correlation or causality? The case of the relationship between exposure to particles and cardiopulmonary mortality. AB - Many epidemiologic studies have observed, in different contexts, a slight short term relationship between particles in air and cardiopulmonary mortality, even when air quality standards were respected. The causality of this relationship is important to public health because of the number of people exposed. Our aim was to make a critical assessment of the arguments used in 15 reviews of published studies. We explain the importance of distinguishing validity from causality, and we systematically analyze the various criteria of judgment within the context of ecologic time studies. Our conclusion is that the observed relationship is valid and that most of the causality criteria are respected. It is hoped that the level of exposure of populations to these particles be reduced. In Europe, acting at the root of the problem, in particular on diesel emissions, will also enable the reduction of levels of other pollutants that can have an impact on health. In the United States, the situation is more complicated, as particles are mainly secondary. It is also essential to continue with research to become better acquainted with the determinants of personal global exposures and to better understand the toxic role of the various physicochemical factors of the particles. PMID- 11256499 TI - Real-world vehicle emissions: a summary of the tenth coordinating research council on-road vehicle emissions workshop. AB - The Coordinating Research Council (CRC) held its tenth workshop in March 2000, focusing on results from the most recent real-world vehicle emissions research. In this paper, we summarize the presentations from researchers who are engaged in improving our understanding of the contribution of mobile sources to emission inventories. Participants in the workshop discussed efforts to improve mobile source emission models and emission inventories, results from gas- and particle phase emissions studies from spark-ignition and diesel-powered vehicles, new methods for measuring mobile source emissions, improvements in vehicle emission control systems (ECSs), and evaluation of motor vehicle inspection/maintenance (I/M) programs, as well as topics for future research. PMID- 11256501 TI - Remedial strategies for municipal solid waste management in China. AB - The purpose of this investigation is to evaluate the current status and to identify the problems of municipal solid waste (MSW) management in China to determine appropriate remedial strategies. This is the second of two papers proposed on this topic. Major problems or difficulties identified in MSW management in China include MSW land, air, and water pollution, commingled collection, poor administration, shortage of funds, lack of facilities, and problems of training and public awareness. In order to solve these problems and to improve MSW management in China, remedial strategies in three areas are recommended: institutional reform, technology development, and legislation and administrative improvement. The primary principle involved in institutional reform is unifying legislative responsibilities into one body and developing a market mechanism for handling MSW. Composting, landfills, and incineration should be equally developed in accordance with China's needs. The feasibility of developing technology to handle MSW in China is discussed. Also recommended is the establishment of sound regulatory systems, including a service fee system, a source separation system, and a training program. China is presently undergoing economic and institutional reform at the national and local levels. Results of this study will provide useful information on MSW management in China. PMID- 11256500 TI - Municipal solid waste characteristics and management in China. AB - The purpose of this investigation is to evaluate the current status and identify the problems of municipal solid waste (MSW) management in China in order to determine appropriate remedial strategies. This is the first of two papers, discussing primarily the general characteristics of MSW and its management in China. The second paper focuses on specific remedial strategies. MSW generation in China has increased rapidly in the past 20 years from 31.3 million tons in 1980 to 113.0 million tons in 1998. The annual rate of increase is 3-10%. The average generation per capita is 1.0 kg/day (0.38 t/year). Nearly one-half of the waste generated is dumped in the suburbs, where the accumulated quantity has reached 6 billion tons, which has caused heavy environmental pollution. This paper provides information on MSW management in China, such as MSW generation and its physical, chemical, and biological properties. Low calorific value and high moisture content characterize China's municipal waste. Other issues related to MSW management in China are also discussed, including the factors that influence MSW generation quantity and PMID- 11256502 TI - Ozone air quality over North America: part I--a review of reported trends. AB - Ozone and precursor trends can be used to measure the effectiveness of regulatory programs that have been implemented. In this paper, we review trends in the concentrations of O3 NOx, and HCs over North America that have been reported in the literature. Although most existing trend studies are confounded by meteorological variability, both the raw data trends and the trends adjusted for meteorology collectively indicate a general decreasing trend in O3 concentrations in most areas of the United States during 1985-1996. In Canada, mean daily maximum 1-hr O3 concentrations at urban sites show mixed trends with a majority of sites showing an increase from 1980 to 1993. Mean daily maximum 1-hr O3 at most regionally representative Canadian sites appears to decrease from 1985 to 1993 or shows no significant change. There are far fewer data and analyses of NOx and HC trends. Available studies covering various ranges of years indicate decreases in ambient NO and HC concentrations in Los Angeles, CA, decreases in HC concentrations in northeastern U.S. cities, and decreases in NOx concentrations in Canadian cities. Two key needs are long-term HC and NOx measurements, particularly at rural sites, and a systematic comparison of trend detection techniques on a reference data set. PMID- 11256503 TI - Ozone air quality over north america: part II--an analysis of trend detection and attribution techniques. AB - Assessment of regulatory programs aimed at improving ambient O3 air quality is of considerable interest to the scientific community and to policymakers. Trend detection, the identification of statistically significant long-term changes, and attribution, linking change to specific climatological and anthropogenic forcings, are instrumental to this assessment. Detection and attribution are difficult because changes in pollutant concentrations of interest to policymakers may be much smaller than natural variations due to weather and climate. In addition, there are considerable differences in reported trends seemingly based on similar statistical methods and databases. Differences arise from the variety of techniques used to reduce nontrend variation in time series, including mitigating the effects of meteorology and the variety of metrics used to track changes. In this paper, we review the trend assessment techniques being used in the air pollution field and discuss their strengths and limitations in discerning and attributing changes in O3 to emission control policies. PMID- 11256504 TI - Functional and regulatory characteristics of eukaryotic type II DNA topoisomerase. AB - DNA topoisomerases are ubiquitous nuclear enzymes that govern the topological interconversions of DNA by transiently breaking/rejoining the phosphodiester backbone of one (type I) or both (type II) strands of the double helix. Consistent with these functions, topoisomerases play key roles in many aspects of DNA metabolism. Type II DNA topoisomerase (topo II) is vital for various nuclear processes, including DNA replication, chromosome segregation, and maintenance of chromosome structure. Topo II expression is regulated at multiple stages, including transcriptional, posttranscriptional, and posttranslational levels, by a multitude of signaling factors. Topo II is also the cellular target for a variety of clinically relevant anti-tumor drugs. Despite significant progress in our understanding of the role of topo II in diverse nuclear processes, several important aspects of topo II function, expression, and regulation are poorly understood. We have focused this review specifically on eukaryotic DNA topoisomerase II, with an emphasis on functional and regulatory characteristics. PMID- 11256505 TI - Thermophilic adaptation of proteins. AB - Hyperthermophilic organisms optimally grow close to the boiling point of water. As a consequence, their macromolecules must be much more thermostable than those from mesophilic species. Here, proteins from hyperthermophiles and mesophiles are compared with respect to their thermodynamic and kinetic stabilities. The known differences in amino acid sequences and three-dimensional structures between intrinsically thermostable and thermolabile proteins will be summarized, and the crucial role of electrostatic interactions for protein stability at high temperatures will be highlighted. Successful attempts to increase the thermostability of proteins, which were either based on rational design or on directed evolution, are presented. The relationship between high thermo-stability of enzymes from hyperthermophiles and their low catalytic activity at room temperature is discussed. Not all proteins from hyperthermophiles are thermostable enough to retain their structures and functions at the high physiological temperatures. It will be shown how this shortcoming can be surpassed by extrinsic factors such as large molecular chaperones and small compatible solutes. Finally, the potential of thermostable enzymes for biotechnology is discussed. PMID- 11256506 TI - Characterization of the distribution of matter in hybrid liver support devices where cells are cultured in a 3-D membrane network or on flat substrata. AB - Bioreactors for liver assist tested on small animal models are generally scaled up to treat humans by increasing their size to host a given liver cell mass. In this process, liver cell function in different culture devices is often established based on the metabolite concentration difference between the bioreactor inlet and outlet irrespective of how matter distributes in the bioreactor. In this paper, we report our investigation aimed at establishing whether bioreactor design and operating conditions influence the distribution of matter in two bioreactors proposed for liver assist. We investigated a clinical scale bioreactor where liver cells are cultured around a three-dimensional network of hollow fiber membranes and a laboratory-scale bioreactor with cells adherent on collagen-coated flat substrata. The distribution of matter was characterized under different operating modes and conditions in terms of the bioreactor residence time distribution evaluated by means of tracer experiments and modeled as a cascade of N stirred tanks with the same volume. Under conditions recommended by the manufacturers, matter distributed uniformly in the clinical-scale bioreactor as a result of the intense backmixing (N=1) whereas axial mixing was negligible in the laboratory-scale bioreactor (N=8). Switching from recycle to single-pass operation definitely reduced axial mixing in the clinical-scale bioreactor (N=2). Increasing feed flow rate significantly enhanced axial mixing in the laboratory-scale bioreactor (N=4). The effects of design, operating mode and conditions on matter distribution in bioreactors for liver cell culture suggest that characterization of the distribution of matter is a necessary step in the scale-up of bioreactors for liver assist and when function of liver cells cultured in different bioreactors is evaluated and compared. PMID- 11256507 TI - The Type II Aachen-Keratoprosthesis in humans: case report of the first prolonged application. AB - PURPOSE: To improve the prognosis of corneal grafts in silicone-oil filled eyes of patients with severe ocular trauma by a prolonged application of the Type II Aachen-Keratoprosthesis (KPro). This application endeavors to improve post keratoplasty prognosis by avoiding corneal endothelial dystrophy in the aphakic eye due to contact with silicone oil. PATIENT AND PROCEDURES: The Aachen Keratoprosthesis' haptic was modified to allow tight contact with cells. The Type II Aachen-Keratoprosthesis was then implanted in an 18-year-old male, with previous management of bilateral corneal rupture. Rather than utilize the device as a temporary intraoperative tool, we extended the device's lifespan in the eye. MAIN FINDINGS: Following implantation, the patient could see hand movements up to 0.1 with best correction. After 8 weeks, vision decreased and a retroprosthetic membrane proliferated. Upon conjunctival retraction, 3 months after the initial surgery, we excised the prosthesis and performed a re-vitrectomy and corneal grafting. The silicone oil was removed. After eighteen postoperative months, the graft remained clear, the retina was completely attached, and the vision was stable: 0.1 best corrected. CONCLUSION: This case reports the prolonged implantation and prospect of the Type II Aachen-Keratoprosthesis to be utilized as a permanent device to restore vision in the near future. PMID- 11256508 TI - Multiorgan failure in a patient with HbS-HbC heterozygous erroneously submitted to plasma. PMID- 11256509 TI - Ake Senning in memoriam. PMID- 11256510 TI - From the heart-lung machine to the total artificial heart. PMID- 11256511 TI - Bidialysis: a new technique. AB - An increase in solute removal, a shorter dialysis session, the patient's well being and the reduction of global costs are the principal aims of the new hemodialysis methods. The simultaneous use of two hemodialyzers in hemodialysis has been experimented by other researchers. Our technique involves the use of two cuprophan hemodialyzers in sequence (double filter system: DFS), each one connected separately to fresh dialysate. Fifteen symptomatic large patients were treated with DFS and the results were compared to conventional hemodialysis (CHD). After the first hemodialyzer, modification of pH and electrolytes occurred in the plasma composition. In the second hemodialyzer, urea depuration occurred without further significant changes in hydroelectrolytic or acid-basic plasma patterns. The Kt/V increased from 1.10 to 1.29 (18%). The authors conclude that in DFS there was an advantage in urea clearance, osmolarity stability and reduction of side effects. PMID- 11256512 TI - Beneficial clinical effects of ozonated autohemotherapy in chronically dialysed patients with atherosclerotic ischemia of the lower limbs--pilot study. AB - Ozonated autohemotherapy is a controversial but successful method of treatment, used in particular in European countries. There are many fields in which medical ozone could be of value: treating different infections, immunodeficiency syndromes, neoplasms. Encouraging results have also been achieved in the treatment of atherosclerotic ischemia of the lower limbs. In this preliminary study, the influence of blood ozonation on the intensity of symptoms of ischemia of the lower extremities was analysed among dialysed patients with chronic renal failure. We examined 5 hemodialyzed patients and 7 patients treated with peritoneal dialysis immediately before and after 14 sessions of ozonated autohaemotherapy. Eleven patients (91.6%) reported a subjective decrease in perceived intensity of ischemic pains, or observed prolongation of intermittent claudication distance. During march tests performed on a treadmill, we found significant prolongation of intermittent claudication distance in all examined patients - 65.6% (mean value, p (< or =0.01). Patients treated with peritoneal dialysis achieved much greater improvement than did hemodialyzed patients (165% vs. 42%). We concluded that autohemotherapy with ozone, in a concentration of 34.4 mcg/ml of blood, is safe, easily applied and may be useful In the therapy of atherosclerotic ischemia of lower extremities among dialyzed patients. It could also be a complement to current treatment, especially in cases where the latter has failed. PMID- 11256513 TI - Oxygen metabolism under various bypass flow conditions during cardiopulmonary support in awake goats. AB - Despite its wide clinical application, patient recovery from cardiopulmonary support (CPS) is not necessarily satisfactory. To clarify what influence CPS has on organ perfusion, we investigated the oxygen metabolism under various bypass flow (BF) conditions in a series of chronic animal CPS experiments. The CPS system, which consists of a pulsatile ventricular assist device and a compact artificial lung was installed without anesthesia in 6 adult goats weighing 49-51 kg. BF was adjusted stepwise from 0% to 50%, 75%, 90%, and 100% of total systemic blood flow (TSF) by balancing the pulmonary arterial flow. The animals' TSF and oxygen delivery (DO2) were sufficiently maintained throughout the experiments. The oxygen consumption (VO2) and the oxygen extraction rate (ExO2) increased from 178+/-14 to 342+/-19 ml/min, and from 28+/-2% to 64+/-1%, respectively, in proportion to the increase of CPBF dependency from 0% to 100%. The blood lactate level did not change appreciably even at 90% BF from 5.7+/-0.3 to 11.2+/-1.2 mg/dl, but drastically elevated to 23.5+/-4.6 mg/dl at the total bypass. This indicates that CPS leads to a relative lack of oxygen and can induce organ dysfunction due to increasing VO2 and ExO2 in proportion to the increase of BF dependence even if TSF and DO2 are sufficiently maintained. PMID- 11256514 TI - Hemolysis and hematology profile during perfusion: inter-species comparison. AB - INTRODUCTION: Cardiopulmonary bypass components need to be tested on an animal model before their clinical application. Because their weight is similar to that of man, the calf and pig are often used. This study compares the impact of prolonged perfusion on hemolysis and hematology profile in both species. METHODS: Three calves (mean bodyweight: 77.2+/-4.4 kg) and three pigs (80+/-5.3 kg) were connected to an extracorporeal circulation circuit by jugular venous and carotid arterial cannulation, with a mean flow rate of 3.5L/min for 6h. After 7 days, the animals were sacrificed. A standard battery of blood samples was taken before, throughout, and 24h, 48h and 7 days after bypass. ANOVA was used for repeated measurements. RESULTS: Absolute values of red cell count were higher in the calf (p<0.001), while normalized values were higher in the pig (p<0.001). Absolute values of white cell count were higher in the pig, while normalized values diverged toward the end of the perfusion with an increase in the calf and a decrease in the pig (p<0.001). Free plasma Hb and LDH exhibited similar profiles in both groups. CONCLUSIONS: In the setting of prolonged perfusion, species type- bovine or porcine--has an impact on hematology profile, but not on hemolytic parameters. These findings should be taken into account when cardiopulmonary bypass components are tested. PMID- 11256515 TI - Hydrodynamic function of a biostable polyurethane flexible heart valve after six months in sheep. AB - Survival to six months for sheep with a non-biostable polyurethane mitral heart valve prosthesis has been reported previously, however, with surface degradation and accumulation of calcified fibrin/thrombus that impaired leaflet motion and compromised hydrodynamic function. Newly available biostable polyurethanes may overcome this problem. Six adult sheep with biostable polyurethane trileaflet heart valve prostheses of documented hydrodynamic performance, implanted in the mitral position, were allowed to survive for 6 months. Explanted valves were photographed, resubmitted to hydrodynamic function testing, and studied by light and electron microscopy. Explanted valves were structurally intact and differed little in appearance from their preimplant state. Hydrodynamic testing showed no deterioration in pressure gradient or energy losses compared with pre-implant values. Biostable polyurethanes demonstrated improved blood compatibility leaving leaflets flexible and valve function unimpaired. Biostable polyurethanes may thus improve prospects for prolonged function of synthetic heart valve prostheses. PMID- 11256516 TI - Current concepts in lymphocyte homing and recirculation. AB - The immune system consists of a complex collection of leukocytes and dendritic cells that surveys most tissues in the body for the appearance of foreign antigens. For an efficient immune response, the interaction and co-localization of antigen-presenting cells, costimulatory helper cells and effector cells are crucial parameters. Therefore, the migration routes of antigen-presenting cells and potential antigen-specific lymphocytes merge in secondary lymphoid organs in order to increase the likelihood and speed of a lymphocyte finding its cognate antigen. Additionally, antigen-primed effector cells are directed to the tissue where they are most likely to encounter their cognate antigen. This highly organized and efficient antigen encounter is based on a continuous recirculation of antigen-specific lymphocytes between blood, peripheral tissue, and secondary lymphoid organs. Moreover, the efficacy of the immune system is further increased by the ability of different lymphocyte subsets to recirculate only through distinct tissues. The scope of this review is to outline the concept and mechanisms of lymphocyte homing and recirculation and to discuss the significance for the immune defense. Current models in leukocyte homing and recirculation and the underlying molecular functions of implicated cell adhesion molecules, chemokines, and chemokine receptors are discussed. PMID- 11256517 TI - Recent advances in inflammatory bowel disease. AB - The last decade has seen tremendous advances in our knowledge, which has led to genuine improvements in our understanding of the pathogenesis and management of inflammatory bowel disease (IBD). The combined power of cellular and molecular biology has begun to unveil the enigmas of IBD, and, consequently, substantial gains have been made in the treatment of IBD. Refinements in drug formulation have provided the ability to target distinct sites of delivery, while enhancing the safety and efficacy of older agents. Simultaneous progress in biotechnology has fostered the development of new agents that strategically target pivotal processes in disease pathogenesis. This article addresses our current understanding of the pathogenesis of IBD, including the latest developments in animal models and covers agents currently used in the treatment of IBD as well as emerging therapies. PMID- 11256518 TI - Acute renal ischemia model in dogs: effects of metoprolol. AB - INTRODUCTION: To study the functional and histological alterations in dog kidneys submitted to total ischemia for thirty minutes and the possible metoprolol protective action. MATERIAL AND METHODS: Sixteen dogs anesthetized with sodium pentobarbital (SP) were studied and divided into two groups: G1-8 dogs submitted to left nephrectomy and right renal artery clamping for thirty minutes, and G2-8 dogs submitted to the same procedures of G1 and to the administration of 0.5 mg x kg(-1) metoprolol before ischemia. Attributes of renal function were studied. RESULTS: There was acute tubular necrosis and a decrease of renal blood flow and glomerular filtration, and a increase of renal vascular resistance in both groups. CONCLUSION: The thirty minute renal ischemia appears to have determined the alterations found in the renal function and histology in both groups. Metoprolol, used in G2, as to the time and dose applied didn't protect the kidney from the ischemic episode. PMID- 11256519 TI - Serum calcitonin in renal transplant patients. AB - We obtained blood samples from 60 renal transplant patients from our transplant clinic and from control subjects for biochemical analyses. Cyclosporin levels were measured in whole blood. Serum levels of calcitonin, calcium, phosphate, albumin, urea, creatinine, and activity of alkaline phosphatase were determined. Serum calcitonin levels were significantly higher in renal transplant patients. There was no correlation between serum calcitonin levels and activity of serum alkaline phosphatase, or levels of serum calcium, phosphate, albumin, urea, creatinine or cyclosporin. Serum calcitonin also showed no correlation with patient age or transplant age. PMID- 11256520 TI - Effects of nitric oxide synthesis inhibition on FK506-induced nephrotoxicity in rats. AB - This study was designed to evaluate the role of nitric oxide (NO) in FK506 induced nephrotoxicity by administering an inhibitor of NO synthesis, N omega nitro-L-arginine methyl ester (L-NAME) to rats treated with FK506. After one week of treatment with FK506 (3.2 mg/kg/day, intramuscularly) and/or L-NAME (5 mg/100 mL of L-NAME in the drinking water), the arterial pressure, urinary NOx, and parameters for renal function were measured, and histological analysis of the kidney was made. In the L-NAME without FK506 group, L-NAME administration effectively inhibited urinary NOx excretion and increased mean arterial pressure (MAP) without any change in renal function. In the FK506 without L-NAME group, FK506 treatment showed increase in urinary NOx excretion and mild renal dysfunction. In the FK506 with L-NAME group, urinary NOx excretion was decreased by L-NAME administration and renal function was significantly worsened than FK506 without L-NAME group. The plasma creatinine, BUN and urinary N-acetyl-beta-D glucosaminidase increased 2-, 3-, and 3-fold, respectively and the creatinine clearance was reduced by 50% as compared with that in the FK506 without L-NAME group. Histological analysis revealed severe interstitial fibrosis and tubular atrophy in the FK506 + L-NAME treatment group. Thus, results suggest that NO synthesis is enhanced in the kidney during FK506-induced nephrotoxicity and that NO synthesis inhibition aggravates FK506-induced nephrotoxicity. NO may play a protective role attributable to the balance of vasoactive substances in FK506 induced nephrotoxicity. PMID- 11256521 TI - Parathormon, calcium, phosphorus, and left ventricular structure and function in normotensive hemodialysis patients. AB - Clinical and experimental data suggest that Parathormon (PTH), calcium, and phosphorus participate in left ventricular hypertrophy (LVH) and affect myocardial contractility in end-stage renal disease. Cellular calcium overload and interstitial fibrosis induced by PTH may lead to impairment of left ventricular diastolic function. Hyperphosphatemia is an independent risk of cardiovascular mortality in dialysis patients. The aim of the study was to estimate the influence of PTH and calcium-phosphorus metabolism on left ventricular structure and function in hemodialysis patients, without hypertension and antihypertensive drug therapy (SBP = 126.2 +/- 11.1 DBP = 75.8 +/- 6.5 mmHg). Echocardiographic findings in a group of 22 normotensive HD patients had been compared to 43 hypertensive HD patients. Relationships between PTH, calcium phosphorus metabolism and echocardiography in normotensive group were then evaluated. Left ventricular mass index (LVMI) was lower in normotensive patients: 128.3 +/- 46.2 versus 165.8 +/- 46.7 (p < 0.01). The prevalence of LVH was 55% in normotensive HD patients compared to 86% in hypertensive group (p < 0.01). In normotensive group we found correlation between PTH and LVMI (r = 0.44; p < 0.05). There were also significant relationships between calcium and posterior wall thickness (r = -0.44; p < 0.05), phosphorus and LVMI (r = 0.47; p < 0.05). A significant correlation was observed between both phosphorus, calcium x phosphorus product and E/A ratio: r = -0.47 and r = -0.43, respectively (p < 0.05 both). Disturbances of calcium-phosphorus metabolism and secondary hyperparathyroidism contributes to left ventricular hypertrophy, and impaired left ventricular diastolic function in normotensive hemodialysis patients. PMID- 11256522 TI - The incorrect placement of hemodialysis catheters in veins. The necessity for urgent x-ray evaluation for its position. AB - We describe a 56-year-old woman with end stage renal failure, who has an unusual complication in relation to catheter placement. She started hemodialysis in 1992 and until October of 1996 had many arteriovenous fistulas in her upper arms and we placed a lot of grafts. Because she had no vascular access for hemodialysis, we decided to place a polyurethane catheter in the left internal jugular vein. The woman had been in good health for 15 months when the blood flow decreased rapidly, so we replaced the catheter with a new one stabilized with silicone in the same place. Because the blood flow again was insufficient for a good dialysis session we decided to check the catheter position angiographically, and were surprised to ascertain that the catheter was placed in the internal thoracic (mammalian) vein. We discuss this patient because she is the first in the international bibliography. PMID- 11256523 TI - Dialysis-related cervical amyloidoma presenting with quadriplegia. PMID- 11256524 TI - Thyrotoxic periodic paralysis in a patient abusing thyroxine for weight reduction. AB - Thyrotoxic periodic paralysis is a rare endocrine disorder most prevalent among individuals of Asian descent that presents as proximal muscle weakness, hypokalemia, and signs of hyperthyroidism. We present an unusual patient with previous normal thyroid function who had abused thyroxine as antiobesity pills and developed periodic paralysis affecting the upper and lower limbs. PMID- 11256525 TI - Repeated transient anuria following losartan administration in a patient with a solitary kidney. AB - We report the case of a 70-year-old hypertensive man with a solitary kidney and chronic renal insufficiency who developed two episodes of transient anuria after losartan administration. He was hospitalized for a myocardial infarction with pulmonary edema, treated with high-dose diuretics. Due to severe systolic dysfunction losartan was prescribed. Surprisingly, the first dose of 50 mg of losartan resulted in a sudden anuria, which lasted eight hours despite high-dose furosemide and amine infusion. One week later, by mistake, losartan was prescribed again and after the second dose of 50 mg, the patient developed a second episode of transient anuria lasting 10 hours. During these two episodes, his blood pressure diminished but no severe hypotension was noted. Ultimately, an arteriography showed a 70-80% renal artery stenosis. In this patient, renal artery stenosis combined with heart failure and diuretic therapy certainly resulted in a strong activation of the renin-angiotensin system (RAS). Under such conditions, angiotensin II receptor blockade by losartan probably induced a critical fall in glomerular filtration pressure. This case report highlights the fact that the angiotensin II receptor antagonist losartan can cause serious unexpected complications in patients with renovascular disease and should be used with extreme caution in this setting. PMID- 11256526 TI - Morphologic evaluation and integrin expression profile of renal tubular cells cultured from percutaneous renal biopsy specimen. AB - Kidney biopsy is an indispensible procedure for making a pathologic diagnosis of renal diseases by fixing and staining the biopsy specimen. However, it is not a routine procedure to culture the cells from a renal biopsy specimen directly, or to utilize the cultured cells for any kind of diagnostic or functional evaluation. In this study, primary culture of the renal tubular epithelial cells was tried from a piece of percutaneous kidney biopsy specimen. Successive passages of the cells were possible until fourth passage. With these cells, morphologic characteristics of the cultured cells and integrin expression profiles were investigated. On light and electron microscopy, these cells were characterized by the cobblestone-like growth, presence of microvilli and tight junction, and the preservation of polarity. Immunohistochemical studies demonstrated the epithelial nature of these cells and particularly their differentiation from renal tubular epithelial cells, of either proximal or distal nephronic segment. The integrin profile confirms the epithelial nature of the cell. We hope that our results facilitate the understanding of pathophysiology of renal tubular cells from the patient directly. PMID- 11256527 TI - Increased urinary uronic acid excretion in experimentally-induced renal papillary necrosis in rats. AB - We have evaluated the potential of urinary uronic acid measurement as an early indicator in the development of renal papillary necrosis (RPN). Urinary uronic acid was quantified with a range of other urinary biochemical parameters in rats given multiple doses of N-phenylanthranilic acid (NPAA) or mefenamic acid (MFA), each of which induces a dose-related papillary necrosis. In addition, histological examination was also carried out to confirm the development and presence of RPN. NPAA was administered to male wistar rats at p.o. doses of 100, 250, and 500 mg/kg and MFA at p.o. doses of 75, 150, and 300 mg/kg on days 1-4 and 8-11, and urine samples were collected for 16 hours each day. NPAA increased uronic acid excretion two-fold for both medium and high doses from day four. MFA increased uronic acid excretion to two and a half-fold by day 10 in the highest dose administered. Urinary creatinine was equally elevated in a dose-related manner following treatment with either NPAA or MFA. None of the other routine markers (urinary or serum) of nephrotoxicity showed any statistical changes. NPAA produced a dose- and time-related increase in excretion of uronic acid. Evidence of widespread papillary necrosis was seen histologically at the high doses of NPAA or MFA. The significant elevation of uronic acid in urine following treatment with either NPAA or MFA was well ahead of the development of RPN detectable by routine histology, suggesting that uronic acid measurement could serve as an early indicator of RPN. The assessment of urinary uronic acid may therefore provide a novel sensitive and selective marker of identifying the lesion earlier than is currently possible. An increase in urinary uronic acid following NPAA and MFA treatment supports the biochemical basis of these changes as a representative of acid mucopolysaccharides accumulation. PMID- 11256528 TI - Effect of cyclosporin A on nitric oxide production in cultured LLC-PK1 cells. AB - The effect of Cyclosporin A on nitric oxide production was studied in cultured LLC- PK1 cells. For this purpose the cells were incubated with vehicle (olive oil, 10 microg/ml in DMSO), Cyclosporin A (CsA, 10 microg/ml), tumor necrosis factor (TNF-alpha, 150 U/ml) + interferon (IFN-gamma, 500 U/ml) to upregulate NOS synthesis, and therefore NO production (used as a positive control), or CsA + TNF alpha + IFN-gamma. After 72 hours the culture medium was collected and nitrite was determined by the Griess method. The results were normalized to the protein harvested from these cells as measured by the Lowry method. Viability was determined by the exclusion of the fluorescent dyes (acridine orange and ethidium bromide). Intracellular calcium was measured spectrophotometrically using the fluorescent calcium indicator fura-2 AM. In CsA treated cells, the nitrite (pmoles/mg of protein) was decreased when compared to control (12.8 +/- 0.5 vs. 18.3 +/- 0.6; p < 0.05; both n = 8). TNF-alpha + IFN-gamma increased the nitrite synthesis (52.0 +/- 0.2; p < 0.05 vs. control; n = 6). This effect was decreased significantly by the simultaneous treatment with CsA (38.8 +/- 0.3; p < 0.05; n = 6). Cell viability in CsA group was decreased when compared to the control (84.7 +/- 0.2% vs. 93.6 +/- 0.1%; p < 0.05; both n = 10). TNF-alpha + IFN-gamma had no effect on viability (93.0 +/- 0.3%; n = 10). However, when combined with CsA, viability was decreased relative to the control (85.0 +/- 0.2%; p < 0.05; n = 10). Acute (1 h) or chronic (72 h) treatment of LLC- PK1 cells with CsA had no effect on basal calcium levels. Our results demonstrate a reduced level of nitric oxide production in LLC-PK1 cells treated with CsA. There was no effect of the drug on intracellular calcium levels, however CsA treatment did reduce cellular viability. We suggest that, in part, the decreased levels of NO production are a secondary consequence of direct cell damage. However, CsA may also be exerting direct effects on NO synthesis through its interactions with both iNOS and cNOS. These results also provide a dual mechanism of action for CsA induced nephrotoxicity, that is, direct cell damage and interference with the NO system within the nephron. PMID- 11256529 TI - Are urea and creatinine uremic toxins in the rat? AB - Urea and creatinine are not generally considered to be important uremic toxins despite evidence from dialysis experiments to the contrary, and despite striking elevations of these nitrogenous waste products in uremia. In order to study this problem in acute uremia, we used a new dietary method for prolonging the survival of bilaterally nephrectomized rats. Urea or creatinine were injected on three successive days starting one day after the inception of uremia. Urea or creatinine injections shortened the survival time of acutely uremic rats, and increased the involution of thymus and spleen. The extra urea, but not creatinine, increased the serum osmolality. These data indicate that urea and creatinine are toxic in the acutely uremic rat. Hypertonicity of the serum may contribute to the toxicity of urea. Additional mechanisms of toxicity and additional toxins are not excluded. PMID- 11256530 TI - Outcomes and APACHE II predictions for critically ill patients with acute renal failure requiring dialysis. AB - BACKGROUND: Despite the widespread availability of dialytic and intensive care unit technology, the probability of early mortality in critically ill patients with acute renal failure (ARF) is still high, and the evaluation of the patients' prognosis has been difficult. The Acute Physiology and Chronic Health Evaluation II (APACHE II) score is a reliable indicator of severity of illness and likelihood of survival in critically ill patients with ARF. We have attempted to determine whether the APACHE II scoring system can be used to predict prognosis. METHODS: A retrospective cohort study evaluated the medical records of 100 consecutive patients in intensive care units with acute renal failure who required dialysis from January 1997 through December 1998. RESULTS: Of the 100 patients studied, 65 were men and 35 were women. The mean age of survivors and nonsurvivors was 59.4 +/- 20.3 years and 58.3 +/- 20.0 years. The overall mortality rate was 71%. There were no significant differences between survivors and nonsurvivors in age, gender, or indication for dialysis. The cause of death in the majority of patients was related to higher APACHE II score during the 24 hours immediately preceding the initiation of acute hemodialysis, and carry mortality rates exceeding 85% with an APACHE II score of 24 or higher. CONCLUSION: We conclude that mortality rate for acute renal failure in intensive care unit patients continues to be high. The use of the APACHE II score determined at the time of initiation of dialysis for patients with ARF is a statistically significant predictor of patient survival. There is a significant trend with APACHE II score for outcome. PMID- 11256531 TI - Symptomatic atrial arrhythmias in hemodialysis patients. AB - BACKGROUND/AIMS: Cardiac arrhythmias are frequent in hemodialysis patients and can interrupt treatment. However, the frequency and risk factors have remained unclear because previous reports of arrhythmias in dialysis patients have usually been continuous-monitoring studies that looked at all cardiac ectopy regardless of its seriousness. METHODS: We reviewed retrospectively only symptomatic atrial arrhythmias in a population of 106 maintenance hemodialysis patients over three years, in order to estimate their actual frequency and any risk factors. RESULTS: Ten patients, seven men and three women, required treatment for atrial arrhythmias (9.4%): four for supraventricular tachycardia, three for atrial flutter, and three for atrial fibrillation. Their mean age was 53.7 +/- 6.1 years; five of them were < or = 40 years. Seven arrhythmias were new, three were recurrences. All but one occurred between 3 and 4 hours of hemodialysis, and dialysis had to be stopped in nine instances. There was no pattern of hypotensive episodes preceding the arrhythmias. Mean serum K+ drawn at the time of the arrhythmias was 3.8 +/- 0.2 mEq/L. Mean plasma intact parathormone was 1128 +/- 417 pg/mL, compared to 454 +/- 58 pg/mL for our entire hemodialysis population (p = .0036). Subsequent echocardiograms showed abnormalities in 9/10 patients: five had left ventricular hypertrophy, six had left atrial enlargement, five had valvular lesions (four mitral regurgitation; one aortic incompetence), and three had ejection fractions <50%. There were four deaths in these patients over the next 14 months, but probably only one was cardiac. CONCLUSIONS: Serious atrial arrhythmias are common in a hemodialysis population. Risk factors for symptomatic atrial arrhythmias in hemodialysis patients may include hyperparathyroidism and echocardiographic findings of chamber enlargement, valvular lesions, or ventricular dysfunction. PMID- 11256532 TI - Assessment of radiocontrast media induced renal vasoconstriction by color coded duplex sonography. AB - INTRODUCTION: Changes in renal hemodynamics are suspected to be one of the major pathogenetic correlates in radiocontrast media-induced nephrotoxicity. We investigated whether color-coded duplex sonography is an appropriate method to document changes in intrarenal vascular resistance, after intravenous injection of the low-osmolar contrast material lopamidol. METHODS: Intrarenal arterial doppler wave forms were analyzed every minute after intravenous injection of 100 mL lopamidol in 10 patients during a voiding cystourogram-procedure. The Resistive Index (RI) of each flow curve, reflecting intrarenal flow resistance, was calculated and compared to the mean of four RI measurements taken before contrast media application. RESULTS: One minute after injection of Iopamidol the RI remained unchanged compared to the baseline standard of 0.70. In measurements obtained 2, 3, 4, and 5 minutes after lopamidol injection a statistically significant rise was seen: (minute 2: 0.74, p < 0.001/minute 3: 0.75, p = 0.001/minute 4: 0.72, p =0.018/minute 5: 0.74, p = 0.031). During the further course, the resistive indices decreased progressively and showed no significant difference in comparison with the baseline standard value. CONCLUSION: Color coded duplex sonography is a simple method to detect changes in renal flow resistance after application of radiocontrast media. Based on our results, we believe that the analysis of intrarenal arterial doppler flow profiles constitutes an ideal method to investigate pathophysiologic mechanisms of radiocontrast media-induced nephrotoxicity, as well as pharmacological concepts in nephroprotectivity. PMID- 11256533 TI - A correlation between renal morphology and renal circulation in pediatric nephrotic syndrome. AB - A morphometric analysis of the renal biopsy specimens and intrarenal hemodynamic study were performed in 37 pediatric patients with idiopathic nephrotic syndrome. The study indicated an inverse correlation between intrarenal hemodynamics (renal plasma flow and peritubular capillary flow) and a relative area of renal cortical interstitium. In respect to the glomerular study, the incidence of glomerulosclerosis increased as the renal perfusion decreased, however, the correlation did not reach a statistical significance. PMID- 11256534 TI - Unmeasured cations: probable cause of relatively low anion gap in chronic renal failure. AB - It is commonly believed that the electrolyte pattern in the patients with chronic renal failure (CRF) is associated with high anion gap (AG) and low serum bicarbonate (HCO3). However it was seen in many clinical studies that the AG is normal or only minimally increased in such patients. It is also known that organic cations, in particular guanidines, also increase in the serum of patients with CRF. We thus postulated that the relatively small increase in AG could be, in part, explained by the coexistent increase in unmeasured cations. If this is true, one may expect that the serum osmolality measured directly will be higher than the estimated one, leading to an osmolar gap (OG). Previous studies have shown that indeed OG exists in patients with CRF. We proceeded to determine SMA 7, AG, and OG simultaneously in ambulatory, undialyzed CRF patients with serum creatinine between 4 and 12 mg/dL. These investigations were also done on nine patients, after dialysis, who went on to have dialysis. The patients were divided into the normal AG (AG < or = 14) and a high AG (AG > 14) groups. There was no correlation of serum bicarbonate with degree of renal dysfunction. Serum AG influenced HCO3 only in the patients with high AG group (bicarbonate = 23.85-0.69 (deltaAG), r2 = 0.45). In patients with normal anion gap there was a good correlation between deltaAG and OG (deltaAG = 3.4-0.15 OG, r = 0.46, r2 = 0.21, p < 0.05). Thus serum bicarbonate appears to be controlled by both AG and OG. Following dialysis, OG decreased from 15.5 +/- 1.06 to 6.08 +/- 1.71, p < 0.01. We conclude that OG must be made up of unmeasured cations of low molecular weight as it normalizes the AG, and gets cleared after dialysis. These low molecular weight substances could be guanidines, such as guanidosuccinic acid and methylguanidine, which are increased by one hundred fold in CRF. PMID- 11256535 TI - Does tuberculosis after kidney transplantation follow the trend of tuberculosis in general population? AB - Despite improvement in graft survival, infection continues to be an important cause of morbidity and mortality after kidney transplantation. We analyzed the clinical courses and outcomes of 16 transplanted patients with positive cultures for mycobacterium tuberculosis. In the course of a 20 year period, there were 13 cases of tuberculosis registered that developed in 456 patients who underwent kidney transplantation in our department, and in three refugees transplanted in other centers (a prevalence of 3.13%). Five of them developed tuberculous infections during 1997. Five patients had residual tuberculosis in preoperative chest X-ray, and specific pyelonephritis as an underlying kidney disease in two of them. All patients with treated with triple immunosuppressives. Before tuberculosis onset, 14 patients experienced one or more episodes of acute rejection and were treated with steroid pulses, ALG or OKT3. Tuberculosis was diagnosed after a period of 1.5 months to 10 years after transplantation. At the time of an infection, the graft function was normal in eight patients and chronic graft failure was evident in eight patients (sCr 210-700 micromol/L). The infection was pulmonary in 12 patients; urinary in two; disseminated in two; pulmonary and urinary, pulmonary and intestinal, and pancytopenia in one patient. All patients were treated with rifampicin and isoniazid in addition to ethambutol for the first two-month period. Treatment lasted from 1-22 months. With 14 patients favorable microbiological responses were registered. Two patients died within the first six months (both with disseminated disease), and the mortality rate was 14.3%. Throughout the followup period, the graft function remained stable and normal in eight patients who had normal graft function at the time of infection onset. Although six patients recovered, progressive graft failure developed and hemodialysis was restarted in one patient two months after antituberculous therapy introduction, and in two patients three years later. Four patients died 2-14 months after AT therapy withdrawal. The causes of death were severe liver failure, cerebrovascular insult and CMV. PMID- 11256536 TI - New insights in the pathophysiology of mitral and aortic regurgitation in pediatric age: role of angiotensin-converting enzyme inhibitor therapy. AB - This review has been focused on the new insights in the pathophysiology of mitral and aortic regurgitation and on the role of ACE-inhibitor therapy in children with chronic volume overload due to left-sided valvular lesions. Recent clinical studies show that these drugs have favorable effects when administered orally in chronic mitral and aortic regurgitation. Interestingly, the beneficial effects of ACE-inhibition regard the basic anatomic, hemodynamic and adaptive pathologic conditions related to volume overload, namely, the regurgitant orifice area and volume and ventricular remodeling. The heart is a plastic structure, constantly being altered in size, shape and composition in response to chronic volume overload. Thus, modulation of cardiac plasticity by ACE-inhibition raises the possibility of using new therapeutic strategies specifically designed to prevent and/or antagonize the mechanical disadvantages secondary to volume overload induced cardiac remodeling. The beneficial effects of ACE-inhibition have also been observed in growing children with asymptomatic valvular regurgitation; thus, it appears that the unloading therapy has the potential of influencing the natural history of both mitral and aortic regurgitation and possibly delays surgical valve repair or replacement. These data justify early inhibition of the renin-angiotensin system in children with left ventricular volume overload due to mitral and aortic regurgitation. PMID- 11256537 TI - "Natural histories" of mitral valve prolapse. Influence of patient selection on cardiovascular event rates. AB - BACKGROUND: In previous studies the reported incidence of cardiovascular events among mitral valve prolapse patients has differed more than 10 fold. We endeavored to determine the relation between the clinical features and mode of ascertainment of mitral valve prolapse and the resulting event rate. METHODS: Between January 1979 and August 1996, 275 patients (129-47% men, 146-53% women, mean age 43 +/- 19 years), were followed for a mean of 98 months after evaluation in a referral center for valvular heart disease. Comparative data were obtained from a separate, less selected population consisting of 316 patients. RESULTS: A total of 65 events occurred (2.9/100 patient-years): 46 (2.0/100 patient-years) mitral surgery, 12 cardiac deaths (0.5/100 patient-years), 6 neurologic ischemia (0.26/100 patient-years), and 1 infective endocarditis (0.04/100 patient-years). The overall event rate varied significantly according to demographic, clinical and echocardiographic variables (all p < 0.0001). It was higher among males (odds ratio-OR 2.1), subjects > or = 45 years of age (OR 14.7), those with a holosystolic murmur (OR 25.9), an enlarged left ventricle (OR 13.5) or left atrium (OR 34.9) and those with 3-4+ mitral regurgitation at color Doppler echocardiography (OR 40.0). It was lower in those with an audible mid-systolic click (OR 0.05). These ORs closely resembled those we reported previously in a less selected population. At multivariate analysis, male gender (p = 0.013), severe Doppler mitral regurgitation (p = 0.0048), and left atrial enlargement (p = 0.046) were all independent predictors of events. CONCLUSIONS: In a population of mitral valve prolapse patients, including many with significant mitral regurgitation at baseline, we identified similar predictors of events but an overall event rate nearly 3 times higher than that we previously reported for relatively unselected patients or family members in New York City (1/100 patient years). Therefore, the impact of patient selection on the prevalence of mitral regurgitation, older age and male gender strongly affects the adversity of the "natural history" of mitral valve prolapse. PMID- 11256538 TI - Transcatheter closure of patent foramen ovale in patients with cryptogenic stroke. AB - BACKGROUND: About 50% of patients with cryptogenic stroke have a patent foramen ovale (PFO). The recurrence rate of paradoxical embolism is higher if a PFO is detected. METHODS: Thirty-five patients with PFO and > or = 1 thromboembolic event due to paradoxical embolism were included in the study (23 males, 12 females, mean age 47.8 +/- 14 years, mean weight 75 +/- 15 kg). Twenty-three patients had a transient ischemic attack whereas 12 experienced an ischemic stroke. Twenty-nine patients had one thromboembolic event, 4 patients had two thromboembolic events, and 2 patients had three thromboembolic events. The implantation procedure was performed, as previously reported, under general anesthesia, fluoroscopic guidance and during transesophageal echocardiography. RESULTS: The implantation procedure was successful in all patients. There were no complications related to the procedure. Four different devices were implanted (Amplatzer 3 patients; Cardioseal 12 patients; Starflex 12 patients, PFO Star 8 patients). The procedure time and fluoroscopic time were 50 +/- 21.8 and 12.2 +/- 8.3 min respectively. At transesophageal echocardiography performed after the procedure, 11 patients had a trivial shunt. None of the patients had a residual shunt at 1 month of follow-up. The mean follow-up was 12.3 +/- 8 months (median 11.0 months, range 3-37 months). In no patient did recurrence of a thromboembolic event occur during follow-up. CONCLUSIONS: Percutaneous PFO closure is a feasible and safe technique for the prevention of recurrent paradoxical embolism. PMID- 11256539 TI - How cryptic is a cryptogenic stroke? PMID- 11256540 TI - Additional beneficial effects of canrenoate in patients with anterior myocardial infarction on ACE-inhibitor treatment. A pilot study. AB - BACKGROUND: Recent evidence suggests that, via the mineralocorticoid receptors present in cardiovascular tissues, aldosterone exerts profibrotic effects, and that partial aldosterone escape occurs during ACE-inhibitor treatment. METHODS: A double-blind, randomized study was performed in order to evaluate the feasibility, tolerability, and the effects of the administration of 25 mg/day of canrenoate plus captopril versus captopril alone to patients with anterior acute myocardial infarction (AMI) unsuitable for or not receiving thrombolytic treatment and to patients in whom such treatment resulted or did not result in reperfusion. One hundred eighty-seven patients with anterior AMI were included in the present study. In all cases serum creatinine concentrations and serum K concentrations were < 2.0 mg/dl and < 5.5 mmol/l respectively. The patients were randomized in two groups: the canrenoate group included 94 patients who received captopril and 25 mg i.v. of canrenoate (1 mg/hour for the first 72 hours and then orally 25 mg/day) whereas the placebo group (93 patients) received captopril and placebo. On admission and on days 10, 90 and 180 all patients underwent echocardiography in order to determine the end-systolic volume (ESV), the ejection fraction (EF), the end-diastolic diameter, the E/A ratio, the E deceleration time as well as the isovolumetric relaxation time (IVRT) and the E and A peak velocities. RESULTS: Unreperfused patients did not show patency of the infarct-related artery whereas in reperfused patients this vessel was patent (7 10 days after AMI). The two groups were similar in age, sex, incidence of diabetes, smoking habits, hypertension, creatine kinase enzymatic peak, adjuvant therapy, baseline EF, ESV, and incidence of coronary artery bypass grafting/coronary angioplasty. Following 10 days of treatment (canrenoate group), only 9 patients presented with serum K and creatinine concentrations respectively > 5.5 mmol/l and > 2.0 mg/dl. Among those patients receiving canrenoate, the mitral E/A ratio was higher compared to the placebo group (p = 0.001) whereas the ESV was significantly reduced (p < 0.05). The deceleration time for reperfused patients receiving canrenoate was higher than that observed for reperfused patients in the placebo group. The intragroup EF was significantly increased (p = 0.042). Compared to the placebo group, the IVRT was significantly higher for unreperfused patients receiving canrenoate than in the placebo group (p = 0.001). Serum creatinine, blood urea and K levels as well as the incidence and extent of vessel disease were similar for both groups. No side effects were observed during the study period. CONCLUSIONS: Our data suggest that the combination of captopril plus canrenoate is feasible for the initial treatment of patients presenting with AMI. Besides, compared to captopril alone it is more efficacious in improving the E/A ratio, the ESV, the EF, and the IVRT. PMID- 11256541 TI - Metoprolol-induced functional benefit in dilated cardiomyopathy is sustained over four years and favorably influences outcome. AB - BACKGROUND: Beta-blockers improve survival and ventricular function in patients with heart failure. We evaluated the long-term persistence of metoprolol-induced improvement and its impact on prognosis in idiopathic dilated cardiomyopathy. METHODS: Two hundred and four of 586 patients enrolled in a registry on the natural history of idiopathic dilated cardiomyopathy survived 4 years without transplantation; 98 of them were on standard heart failure treatment, whereas 106 took metoprolol in addition. We analyzed the effects of treatment using beta blockers in terms of changes in left ventricular ejection fraction (LVEF), NYHA functional class and left ventricular end-diastolic diameter index (LVEDDI) after 1, 2 and 4 follow-up years in order to elaborate an improvement score that was related to the subsequent outcome over 60 months after the 4-year follow-up visit. RESULTS: Greater LVEF increases and NYHA functional class and LVEDDI decreases were observed in patients submitted to metoprolol vs standard treatment at all stages of follow-up. Changes (delta vs baseline) for LVEF (p = 0.02), NYHA functional class (p = 0.0001) and LVEDDI (p = 0.004) were maximal during the first year (10 +/- 11 vs 6 +/- 12 units, -0.72 +/- 0.77 vs -0.23 +/- 0.81, -3.5 +/- 5 vs -1.6 +/- 3.5 mm), persisted at 2 (12 +/- 12 vs 8 +/- 12 units, -0.80 +/- 0.70 vs -0.37 +/- 0.87, -4.2 +/- 5 vs -2.3 +/- 4 mm) but showed a trend to decline at 4 years (11 +/- 12 vs 8 +/- 13 units, -0.54 +/- 0.90 vs -0.24 +/- 0.91, -4.3 +/- 5 vs -2.3 +/- 5 mm) of follow-up. Improvement at 4 years was associated with a better transplant-free survival (81 vs 52%, p = 0.0005, odds ratio 0.36, 95% confidence interval 0.18 to 0.74). CONCLUSIONS: In idiopathic dilated cardiomyopathy, the more significant improvement in symptoms and left ventricular function and size, that is observed following treatment using metoprolol, translates into a better outcome. These benefits peak within the first 2 years of start of treatment but may begin to fade thereafter. PMID- 11256542 TI - Continued cigarette smoking after coronary artery bypass surgery reduces endothelium-dependent vasodilation in internal thoracic artery grafts. AB - BACKGROUND: Cigarette smoking is known to promote endothelial dysfunction, thus it can be responsible for an impaired endothelium-dependent vasomotility in arterial grafts late after coronary surgery. METHODS: Twenty consecutive patients (mean age 64.5 years), previously submitted to coronary bypass surgery with the internal thoracic artery, underwent quantitative angiography of the implanted graft at long-term follow-up (mean time 2.5 years). To assess both endothelium dependent and independent vasomotility, angiograms were acquired before and after selective infusions of acetylcholine (10(-6) mmol/ml) and nitroglycerine (500 microg). The predictive value of risk factors, including previous and continued smoking, for an impairment in endothelium-dependent vasomotility was assessed. RESULTS: Continued smoking (p = 0.038), but not a previous history of smoking (p = 0.55) was the only predictor of a reduced endothelium-dependent vasodilation. While previous smokers and non-smokers showed a similar response to acetylcholine, current smokers showed a reduced endothelium-dependent vasodilation vs non-smokers (94.8 +/- 2.6 vs 99.6 +/- 2.3% of the maximal vasodilative capacity, p = 0.001). CONCLUSIONS: Although maintained, the vasodilative response to acetylcholine appears reduced in internal thoracic artery grafts of patients who continued smoking long term after coronary bypass surgery. Whether this could affect the long-term outcome of these patients has to be further investigated. PMID- 11256543 TI - Radiofrequency catheter ablation of atrioventricular nodal reentry tachycardia: selective approach to the slow pathway via the superior vena cava. AB - Selective radiofrequency catheter ablation of the slow atrioventricular nodal pathway is currently considered the first-line therapy for patients suffering from recurrent symptomatic atrioventricular nodal reentry tachycardia. In most cases slow pathway conduction may be selectively eliminated or modified by the application of radiofrequency current at the posterior portion of Koch's triangle. The ablation site is usually targeted by careful mapping of this area performed using an ablation catheter advanced via the inferior vena cava approach. In this report we describe 2 cases in which the conventional approach to the target site was either impossible owing to the presence of an atresic inferior vena cava (case 1), or contraindicated in view of a history of common femoral vein thrombosis, subsequently extended up to the inferior vena cava (case 2). In both patients a superior vena cava approach was utilized and the slow pathway was successfully ablated. In case of arrhythmias necessitating slow pathway mapping and ablation, such an approach may be considered as a feasible and safe alternative whenever, owing to the presence of anomalies and/or diseases of the inferior vena cava, the conventional approach cannot be employed. PMID- 11256544 TI - Accessory pathway potential recording in a case of permanent junctional reciprocating tachycardia with decremental conduction localized on the atrial site. AB - Permanent junctional reciprocating tachycardia (PJRT) is an uncommon form of atrioventricular reentrant tachycardia due to the presence of an accessory pathway characterized by slow and decremental retrograde conduction. We report a case of PJRT where we demonstrated the possibility of recording a distinct accessory pathway potential. Decremental retrograde conduction was evident using ventricular extrastimuli and it was also adenosine-sensitive. Delivering ventricular extrastimuli a prolongation of the accessory pathway potential-atrium interval was seen demonstrating that decremental conduction was located at the atrial insertion of the pathway. The accessory pathway was successfully ablated using the potential as the target of radiofrequency delivery. These electrophysiological findings seem to support the hypothesis that a nodal-like structure may be responsible for this arrhythmia. PMID- 11256545 TI - Images in cardiovascular medicine. Morphologic changes in left ventricular thrombus in a patient with acute anterior myocardial infarction. Assessment with contrast echocardiography. PMID- 11256546 TI - Evidence-based therapeutic strategies. There is the need to bridge the gap between simplified megatrials and individual prescriptions. PMID- 11256547 TI - The prognostic role of electrocardiography in chronic obstructive pulmonary disease. PMID- 11256548 TI - The molecular mechanisms of angiogenesis: a new approach to cardiovascular diseases. AB - Although the role of new blood vessel formation in cancer and its development have been well documented, the strategy to manipulate angiogenesis in order to restore blood flow in the ischemic myocardium, a novel form of therapy currently undergoing clinical trials, has received less attention. Recent advances in our understanding of the stimuli and of the control mechanisms regulating the development of new blood vessels in coronary heart disease have led to an improved picture of the compensatory healing process that accompanies myocardial ischemia and infarction. However, we have to remind that, together with life- and tissue-saving effects, the angiogenetic process might alter the natural course of the consequences and organ manifestation of arterial diseases as in the atherosclerotic plaque. The purpose of this review is to provide an overview on the molecular mechanisms involved in the angiogenetic process. Angiogenesis during ontogenesis, neoangiogenesis (adult new vessel formation), arteriogenesis and the related regulators will be analyzed. Moreover, the role of neoangiogenesis in plaque development and instability will be discussed. Due to the introductory nature of this review and the large number of studies on neovascularization in ischemic limbs this topic has been omitted. PMID- 11256549 TI - Myocardial viability in ischemic heart disease: new directions and perspectives. AB - In patients with ischemic heart disease detection of myocardial viability is of major clinical and prognostic importance and may significantly affect therapeutic decisions. Reversible left ventricular dysfunction may be due to different pathophysiological mechanisms, including myocardial hibernation and stunning, structural and ultrastructural myocardial changes and alterations in gene expression leading to myocardial cell dedifferentiation. Each of these mechanisms may have different importance related to the clinical history of the patient and severity and duration of left ventricular dysfunction and may significantly influence the extent and time course of functional recovery after myocardial revascularization. In the clinical arena detection of myocardial viability is currently based on the use of nuclear techniques, which show preserved tracer uptake and metabolism in viable myocardium and echocardiographic methods, which detect residual contractile reserve. Both techniques show a similar sensitivity in predicting functional recovery after revascularization, but dobutamine echocardiography has a higher specificity and therefore may be clinically more useful. Due to the limitations of current nuclear and echocardiographic methods in detecting myocardial viability, new developments are directed towards better quantification of viable myocardium and simultaneous assessment of myocardial metabolism, perfusion and function. Doppler tissue imaging, intravenous contrast echocardiography and ECG-gated SPECT with combined evaluation of metabolism and perfusion seem to be the most promising and cost-effective methods for a comprehensive assessment of myocardial viability. The major prognostic importance of myocardial viability in patients with severe left ventricular dysfunction is demonstrated by the fact that patients with a significant amount of viable myocardium have a marked survival benefit from revascularization and an improvement in left ventricular function and NYHA functional class compared with those without or only marginal viability. Thus, in patients with severe dysfunction preoperative quantification of viable myocardium is of utmost importance to identify patients who can benefit from revascularization. In patients with lesser amount of viable myocardium the possible beneficial effect of revascularization on survival, even in the absence of significant improvement in ventricular function, is yet to be demonstrated and should be assessed in future prospective clinical trials. PMID- 11256550 TI - Development of bronchial hyperresponsiveness during childhood. AB - Bronchial hyperresponsiveness (BHR) produces the characteristic pathological abnormalities seen in asthma and clearly plays a central role in the pathophysiology of asthma. The presence of BHR has been demonstrated in infants with asthma, as has the possibility of BHR persisting through the childhood period. The level of BHR may not only reflect the state of the airways, as a marker of airway dysfunction, but may also predict the persistent prognosis of the disease. Thus, measurement of BHR may provide important information about the symptoms and lung function in children with asthma. In view of multiple pathophysiological mechanisms, BHR does not seem to have a single cause. Many potential confounding variables, such as age, gender and genetic status, and some environmental factors, such as allergens, infections, and pollutants, could be responsible for the establishment of childhood BHR. There may be differences between the mechanisms that induce transient BHR and the mechanisms that induce persistent BHR. Also, there may be differences between the causes that induce BHR in the infantile period and the causes that maintain persistent BHR during childhood asthma. There is also disagreement as to the most suitable method to measure BHR in children, especially in infants. The assessment of BHR in young children has not been uniformly successful, and measurements of BHR changes over the childhood period (are associated with a number of problems. To resolve these problems, there may be two ways to study childhood BHR. One is to use age-matched specific techniques to clarify the precise BHR in each age group; the other is to use simple techniques that can be performed over the childhood period on a large number of subjects. In studies of infantile respirator, dysfunction the ultimate goal is to establish a simple, noninvasive method by which measurements of respiratory function may be obtained in infants. Further investigations and acceptable methods will be needed to clarify, the mechanisms involved in the establishment of asthma throughout the childhood period. PMID- 11256551 TI - Feasibility of a nurse-run asthma education program for urban African-Americans: a pilot study. AB - The objective of the study was to assess the feasibility of implementing and evaluating a culturally appropriate in-patient asthma education program specifically targeted for African-Americans. A consecutive sample of 28 African American patients ages 18-50 who were hospitalized for asthma were randomized to an intervention group, which received three one-on-one sessions on chronic asthma management, or a control group, which received the usual care. Data on symptom frequency, self-management behaviors, quality of life, depression, and health care resource use were collected at baseline and at 3 and 6 months. Although the time required to recruit our sample took longer than anticipated, 28 subjects agreed to be in the study (70% acceptance rate) and complete the baseline interview. We observed no statistically significant differences from baseline or changing trends in frequency of asthma symptoms, self-management behaviors, and health care resource use between the intervention and control groups at 3 and 6 months. However patients in the intervention group demonstrated a greater average increase in asthma-related quality of life and a greater average decrease in depression than the control group. Feasibility issues included shortened length of stay, which necessitated conducting all three self-management sessions together; multiple interruptions during the sessions, and retention issues at 3- and 6-month follow-ups. The lessons learned from this pilot study are invaluable in that they will enable us to make changes in our existing protocol to ensure the success of a larger clinical trial. PMID- 11256552 TI - A comparison of the individual best versus the predicted peak expiratory flow in patients with chronic asthma. AB - In the management of patients with asthma, peak expiratory flow (PEF) monitoring is used and based on the individual best PEF or the predicted PEE Recent international guidelines have recommended the use of the best PEF rather than the predicted PEF as an index, although there is little evidence to support which index is more appropriate. Therefore, we investigated the relationship between the best PEF and the predicted PEF in 166 consecutive asthmatic patients to see which value would be the better basis for their PEF monitoring. All eligible patients had undergone treatment for their asthma for over 6 months and were asked to measure their PEF four times a day. The best PEF was defined as the maximal PEF achieved at any time from all previous measurements. The predicted PEF was calculated based on a report on the standard PEF in normal Japanese subjects. The mean best PEF was significantly higher than the mean predicted PEF (p < 0.001). There was a strong correlation between the best PEF and the predicted PEF (r = 0.77, p < 0.001). However, in 72 patients (43%) the ratio of the best PEF to the predicted PEF was over 110%, and in 20 patients (12%) the ratio was lower than 90%. The best PEF was higher than the predicted PEF in 76 patients (46%) and lower in 22 patients (13%) by more than 50 L/min. These results suggest that when the predicted PEF was used as the index, pulmonary function was either underestimated or overestimated in over half of these patients. Therefore, the best PEF may be the better index for the management of patients with asthma. PMID- 11256553 TI - Cigarette smoking, but not sensitization to Alternaria, is associated with severe asthma in urban patients. AB - Hereditary susceptibility and allergen exposure have been identified as general risk factors for asthma. However, risk factors for severe asthma still remain to be identified. To further assess and quantify risk factors associated with severe asthma in adult patients apart from clinical exacerbations, 306 randomly selected subjects (mean age 40+/-17 years, 46% males) presenting to an inner city pulmonary practice between 1995 and 1996 were retrospectively investigated. Of these, 117 patients were atopic, 112 had current asthma, and 22 asthmatics had severe asthma. Risk factors associated with atopy were family history of atopy and any domestic pet ownership (OR: 3.1, 95% CI: 1.64-6.1). Asthma was generally associated with atopy (OR: 4.2, CI: 2.4-7.4) and pet ownership (OR: 2.4, CI: 1.2 4.6). Severe asthma was strongly associated with current smoking (OR: 4.8, CI: 1.3-18.3), and lung function was negatively correlated with the amount of consumed cigarettes per day (r = -0.61, p = 0.04). However, no association with sensitization to Alternaria was found in severe asthma. Cigarette smoking is an independent risk factor associated with severe asthma in urban patients, whereas sensitization to Alternaria is of less importance in these patients. PMID- 11256554 TI - Effects of pranlukast, a leukotriene receptor antagonist, on airway inflammation in mild asthmatics. AB - To determine the anti-inflammatory actions of pranlukast, a cysteinyl leuklotriene receptor antagonist, we measured exhaled nitric oxide (NO) concentrations and eosinophil ratio in induced sputum of three groups of mild asthmatics (n = 30): treated with bronchodilators alone, with bronchodilators and inhaled steroid (beclomethasone dipropionate; 400 microg/day), and bronchodilators and pranlukast (450 mg/day). Pranlukast (450 mg/day) reduced the eosinophil ratio in the induced sputum significantly (p < 0.01) without a major effect on the concentration of exhaled NO. Pranlukast also increased values of peak expiratory flow significantly (p < 0.05). Pranlukast may be useful for mild asthmatics, in part through its ability to suppress eosinophilic airway infiltration. PMID- 11256556 TI - Increasing U.S. asthma mortality rates: who is really dying? AB - Asthma mortality rates have been increasing since 1979, but rates of change among different demographic subgroups have not been examined in detail. This analysis identifies the demographic subgroups that are most responsible for the increase in asthma mortality rates in the United States between 1979 and 1996. The analysis is limited to those death certificates that specified asthma as the underlying cause of death. Blacks, females, and people aged 65 and older had the largest increases in age-adjusted asthma mortality rates between 1979 and 1996. When all three demographic variables are considered simultaneously, black females aged 65 years and older had the highest crude asthma mortality rates in 1996 and the largest increase in rates since 1979. However, white females aged 65 years and older contributed the most to the increase in age-adjusted rates between 1979 and 1996 because of their relatively larger population size. Overall, the increase in asthma mortality rates between 1979 and 1996 was due primarily to increased mortality rates in the population subgroup aged 65 years and older Even though the rapid increase in asthma mortality rates in those aged 65 years and older shows evidence of a slight reversal after 1989, efforts to develop strategies to reduce overall mortality from asthma should concentrate on middle aged and elderly women. PMID- 11256555 TI - Effects of therapeutic doses of albuterol on beta2-adrenergic receptor density and metabolic changes. AB - Beta2-agonist drugs at inhaled supratherapeutic doses or when given orally or parenterally alter peripheral lymphocyte beta2-adrenoceptor density (betaAR) and have demonstrable metabolic effects. However, it is not known whether these changes occur at therapeutic inhaled doses. We therefore studied the effects of therapeutic doses of inhaled albuterol in five asthmatic subjects (mean age 23.0+/-2.4 years) and six normal subjects (mean age 28.3+/-3.3 years). Subjects were studied in a randomized, double-blind protocol in which each subject received either inhaled albuterol (270 microg four times daily) for 2 weeks followed by placebo or vice versa in two sequential 2-week periods separated by a 2-week washout period. In the asthmatics, baseline FEV1 increased significantly (p < 0.05) after 2 weeks of inhaled albuterol treatment compared to the initial visit and after 2 weeks of placebo (mean FEV1: 3.2 L+/-0.7 L, 2.9 L+/-0.5 L, and 3.0 L+/-0. 7 L, respectively). Baseline peripheral lymphocyte betaAR was not significantly different (p > 0.05) between the asthmatic (mean: 757+/-176) and normal subjects (mean: 732+/-251). However, in neither group was there any significant change (p > 0.05) in betaAR or plasma potassium, insulin, or glucose, either acutely or after 2 weeks of albuterol therapy. The present study confirms that there is no difference in peripheral lymphocyte betaAR between asthmatic and normal subjects and also shows that at therapeutic doses of inhaled albuterol, there are no significant changes in betaAR or metabolic effects. PMID- 11256558 TI - The public health surveillance of asthma. AB - Asthma is a highly prevalent disease that affects the quality of life of many people in the United States. Yet there is limited descriptive epidemiological understanding of the disease, particularly at the state and local levels. Minimal surveillance of asthma is occurring across the country. Surveillance of a disease requires that public health workers have the ability to accurately identify cases, have access to needed data, and have adequate resources so that they can collect, assess, report, and use the data-all considerable challenges in the case of asthma. We consider four groups of questions that asthma surveillance should address: (1) How much asthma is there and what are the trends in asthma occurrence over time? (2) How severe is the asthma and what are the trends in asthma severity over time? (3) How well is asthma controlled and what are the trends in asthma management over time? (4) What is the cost of asthma? Because wise decision making in public health depends on the availability of appropriate data for program planning, implementation, and evaluation, we encourage increased surveillance of asthma in jurisdictions across the country. PMID- 11256557 TI - Content and outcomes of Dutch nurse clinics for children with asthma. AB - Dutch specialist asthma nurses run extramural and transmural nurse clinics for children with asthma. Extramural clinics are run under the responsibility and in the premises of a home care organization. Transmural clinics are run in an outpatient clinic in close collaboration and joint responsibility between home care organizations and hospitals. Effects of and differences between these clinics were determined by using a quasiexperimental design. Visiting a nurse clinic appears to result in a reduced information need and reduced use of health care services. Parents of asthmatic children visiting transmural nurse clinics appeared to have a lower information need than those attending extramural nurse clinics. PMID- 11256559 TI - A comparison of medication adherence indices to assess long-term inhaled corticosteroid medication use. AB - This study examined associations between select hypothesized adherence predictors in claims databases and long-term inhaled corticosteroid adherence. Medication adherence was measured by using medication possession indexes and a refill regularity measure in 1595 older adults using inhaled corticosteroids for 2 years. Medication possession indices were highly correlated with each other but not with the refill regularity measure. Additionally, the hypothesized predictors explained a larger percentage of the variance in the medication possession indices compared to the refill regularity measure. Although long-term retrospective medication utilization poses many measurability issues, the use of multiple indices gives a more complete picture of usage behavior. PMID- 11256560 TI - Parental self-efficacy and morbidity in pediatric asthma. AB - This study investigated the relationship between parental self-efficacy and asthma-related morbidity. Participants included 139 parents of children (ages 5 8) who were diagnosed with asthma and were primarily from lower-income and minority backgrounds. Parents completed a 22-item measure of self-efficacy; factor analysis was conducted on this measure, yielding two factors: learned helplessness and self-efficacy. Correlational analyses indicated that higher scores on the learned helplessness factor were significantly related to increased asthma-related morbidity for the majority of morbidity variables. The self efficacy factor was significantly related to days of school missed. Regression analyses conducted with the factor scores and the morbidity variables provide further support that the learned helplessness factor accounts for a significant amount of the variance in asthma morbidity for many of the variables studied, while the self-efficacy factor was related to only a few. Although improving health outcomes of children with asthma is a multifaceted process, the results of this study suggest that targeting parental self-efficacy, particularly with parents who are experiencing high levels of perceived learned helplessness, may be a helpful component of an intervention program with this population. PMID- 11256561 TI - Mechanisms of taurine antihypoxic and antioxidant action. AB - The study was undertaken to elucidate the effects of taurine on lipid peroxidation (LP) intensity and membrane Na+, K+-ATPase activity in a hypoxic rat model. It was shown that 3 intraperitoneal (i.p.) injections of 200 mg/kg of taurine prevented hypoxia-induced lactate accumulation and LP in brain, liver, and heart tissues and prevented the decrease of Na+, K+-ATPase activity in the liver. It is suggested that the effect of taurine on LP could be due to the taurine antiacidotic action as well as to its membrane stabilizing activity. PMID- 11256562 TI - Ketamine anesthesia at high altitude. AB - There is a clinical need for a safe and effective anesthetic technique in high altitude and remote areas. This report presents a series of 11 consecutive cases documenting the use of ketamine anesthesia in a remote hospital at an altitude of 3,900 m, by primary-care physicians without specialist training in anesthesia. The method of administration is fully described. At a low dose of 2.0 mg/kg, ketamine produces a dissociative anesthesia that does not depress the hypoxic drive, or interfere with the pharyngeal or laryngeal reflexes. Although supplemental oxygen is useful in the recovery phase for less acclimatized individuals, it is usually not required as reductions in oxygen saturation can be raised by physical stimulation that encourages the patient to breathe faster and deeper. The common side effect of emergent nightmares was avoided using midazolam as premedication and a quiet recovery area. This study offers the first available evidence that ketamine with midazolam offers a safe and effective means of anaesthesia at very high altitude, without the need for specialist equipment or training, by careful clinicians experienced in basic airway management. PMID- 11256563 TI - Ethnobotany and exchange of traditional medicines on the Southern Bolivian Altiplano. AB - Research conducted on the collection, use, and vending of traditional medicines by rural Bolivian women indicates that it is an important economic activity as well as having a place in the health system of high altitude inhabitants. The aim of this paper is to discuss the intersection of an approach that focuses on the exchange of traditional medicines with an ethnobotanical perspective that considers the medicines themselves. Women are the focus of this intersection because they are central to the enterprise of collecting and selling traditional medicines, which is an expanding business opportunity due in part to demands by urban consumers. In moving toward an ethnobotanical analysis of the plants themselves, it is important to consider how this focus will enhance our understanding of the marketing and use of traditional medicines and women's roles therein, but researchers must also understand the problems related to the potential use of ethnobotanical data to create new pharmaceuticals. PMID- 11256564 TI - Physiological effects of intermittent hypoxia. AB - Intermittent hypoxia (IH), or periodic exposure to hypoxia interrupted by return to normoxia or less hypoxic conditions, occurs in many circumstances. In high altitude mountaineering, IH is used to optimize acclimatization although laboratory studies have not generally revealed physiologically significant benefits. IH enhances athletic performance at sea level if blood oxygen capacity increases and the usual level of training is not decreased significantly. IH for high altitude workers who commute from low altitude homes is of considerable practical interest and the ideal commuting schedule for physical and mental performance is being studied. The effect of oxygen enrichment at altitude (i.e., intermittent normoxia on a background of chronic hypoxia) on human performance is under study also. Physiological mechanisms of IH, and specifically the differences between effects of IH and acute or chronic continuous hypoxia remains to be determined. Biomedical researchers are defining the molecular and cellular mechanisms for effects of hypoxia on the body in health and disease. A comparative approach may provide additional insight about the biological significance of these effects. PMID- 11256565 TI - Italian high altitude laboratories: past and present. AB - Italy is a mountainous country with a total of 88 huts and bivouacs at altitudes higher than 3,000 m. Starting in the 19th century a great deal of research in high altitude pathophysiology has been carried out in Italy and many Italian physicians have been involved in mountain medicine. Most of the Italian research has been carried out at two locations: the scientific laboratories "Angelo Mosso" on Monte Rosa (Capanna Regina Margherita and Laboratorio Angelo Mosso), and the "Pyramid" in Nepal. The Capanna Regina Margherita, located on the top of Punta Gnifetti (Monte Rosa, 4,559 m), was inaugurated in 1893. With the support of Queen Margherita of Savoy, an Observatory for scientific studies was built beside this hut in 1894. In 1980 the hut was completely rebuilt by the Italian Alpine Club. The Istituto Angelo Mosso at Col d'Olen, at the base of Monte Rosa (at 2,900 m) was inaugurated in 1907. The high altitude laboratory named the "Pyramid" was built in 1990. Made of glass and aluminium, this pyramid-shaped structure is situated in Nepal at 5,050 m. The scientific laboratories "Angelo Mosso" on Monte Rosa (mainly the Capanna Regina Margherita) and the Pyramid form a nucleus for high altitude research: the former is especially devoted to research regarding acute mountain sickness and the response to subacute hypoxia, whereas the latter is a unique facility for research responses to chronic hypoxia, the effect of exposure to very high altitude, and the study of the resident population living in the Himalayas for at least 25,000 years. PMID- 11256566 TI - The cover of the journal. PMID- 11256568 TI - Polymorphism and evolution of HLA class I and II genes and molecules. AB - Genes in the HLA complex, the human major histocompatibility complex (MHC), encode polymorphic HLA class I and II molecules that help T lymphocytes recognise and respond to foreign antigens. Certain HLA class I allotypes also regulate the response of natural killer cells. HLA class I and II molecules with little or no polymorphism contribute a variety of functions to the immune response, as do class I molecules coded by genes outside of the HLA region. Knowledge of the organisation of HLA class I and II genes, of the nucleotide sequences of their alleles, and the three-dimensional structures of their protein products, has facilitated analysis of the evolution and polymorphism of HLA class I and II genes and molecules. In turn, these analyses have provided insight into the mechanisms and selective forces driving change in the HLA complex. PMID- 11256567 TI - Hematological parameters in high altitude residents living at 4,355, 4,660, and 5,500 meters above sea level. AB - There have been a number of reports describing the hematological indicators of Andean residents living at altitudes above 4,000 m, but several confounding factors have made the published results difficult to interpret. To clear up the effect of hypoxia on hemoglobin concentration (Hb, g/dL), hematocrit (Hct, %) and red blood cell concentration (RBC, cells/microL), this publication describes and analyzes these variables in children, men, and women from three large and homogeneous populations living at 4,355 m (n = 151), 4,660 m (n = 400), and 5,500 m (n = 273) in the Southern Peruvian Andes. Hb, Hct, and RBC increase with age in men (p < 0.001), as well as in women (p < 0.001) at the three altitudes of the study. In children (boys and girls) living at 5,500, Hb increases 11% when compared with children living at 4,355 m, and in adults, Hb increases 9.6% when comparing the same altitudes. The maximum percentage increase in Hb with age was 5.6% at 5,500 m, in men and 3.2% at 4,355 m, in women. The average percentage of difference for the Hb concentration between adult men and women is 6.6% at 4,355 m, 9.8% at 4,660 m, and 11.6% at 5,500 m. The differences in Hb concentration between men and women can only be seen after puberty. Finally, Hb is higher in older than younger women, which confirms the role of menopause in the development of erythremia. The result of this analysis reinforces the notion that Hb and Hct seem to be stable and useful parameters for acclimatization only at moderate altitudes; with aging or with increasing altitude, they may become excessive and lose their efficiency to protect the venous oxygen pressure. PMID- 11256569 TI - Genetics and molecular genetics of the MHC. AB - The MHC is a well-characterised region of the human genome, containing a high diversity of genes and an apparent clustering of genes involved in the immune response. The genomic sequence of an 8 Mb section, containing the MHC and flanking regions and covering over 300 genes, will soon be available. Since this is a highly polymorphic region, the molecular genetics of the MHC and its relationship with disease have been studied in considerable detail. PMID- 11256570 TI - The human leucocyte antigens and clinical medicine: an overview. AB - The Major Histocompatibility Complex (MHC) occupies 4-6 megabases on the short arm of chromosome 6 and is the most intensively studied segment of the human genome. This region was first discovered through its influence on transplantation rejection and on antigen-specific immune responses. The most important genes for managing these functions encode the HLA molecules (human leucocyte antigens) which are highly polymorphic in human populations. HLA typing for these polymorphisms is widely used in clinical medicine when identifying optimal organ donors or recipients and in assessing the risk of diseases such as narcolepsy, hereditary hemochromatosis, ankylosing spondylitis and certain autoimmune disorders. As new genes are identified in the MHC, the clinical impact of this genetic region is likely to assume further importance as outlined in this review. PMID- 11256571 TI - MHC class I molecules, structure and function. AB - MHC class I molecules (MHC-I) are cell surface recognition elements expressed on virtually all somatic cells. These molecules sample peptides generated within the cell and signal the cell's physiological state to effector cells of the immune system, both T lymphocytes and natural killer (NK) cells. In addition, molecules structurally related to MHC-I, collectively known as MHC-Ib, are more specialized and, in some cases, interact with more limited subsets of lymphoid cells. Using the recently determined structure of the classical MHC-I molecule, H-2Dd, as a paradigm for structure and function, we review other MHC-I and MHC-Ib molecules, with an emphasis on how the same basic structural fold is employed by classical MHC-I molecules to bind specific peptides and T cell receptors, and is exploited by the MHC-Ib molecules in more stringent molecular interactions. It is instructive that structurally related molecules have evolved to perform a number of unique and distinct functions in immune and non-immune recognition. PMID- 11256572 TI - Structural principles of MHC class II antigen presentation. AB - Normal immune surveillance depends on the ability of MHC class II molecules to bind peptide antigens and carry them to the cell surface for display to T cells. To do this efficiently, class II molecules must be able to bind peptides from a broad array of antigen sequences and retain them at the cell surface long enough for T-cell recognition to occur. Class II molecules accomplish this task through a combination of clever structural biochemistry and the help of at least two different molecular chaperones: the class II-associated invariant chain (Ii); and a non-peptide binding class II molecule termed H2-DM in mouse and HLA-DM in man (DM). Here, we compare the existing 3-dimensional structures of class II-peptide complexes in order to review the general principles of peptide binding and presentation. We extend this analysis to include the structures of proteins known to interact with MHC class II, focusing primarily on the Ii chain and DM. PMID- 11256573 TI - Pathways of antigen processing and presentation. AB - CD8+ and CD4+ T lymphocytes recognise peptides stably bound to class I or class II MHC molecules, respectively. These complexes are assembled intracellularly during the biosynthesis and trafficking of MHC molecules. It is now clear that a number of different molecules and macromolecular complexes are drafted in to assist this process. Some of these are chaperones which appear to be dedicated to assisting MHC molecules capture peptides, whilst others may have additional cellular functions. Peptides form an integral part of the final MHC glycoprotein structure and their availability can regulate the kinetics and level of expression of MHC molecules on the cell surface. In vivo, significant time may elapse between generation of peptide/MHC complexes and their recognition by T cells. This requires that the complexes generated are stable and long-lived on the cell surface. Several mechanisms appear to contribute to the generation and display of long-lived complexes. Some pathogens have evolved mechanisms to evade and interfere with presentation of their own antigens. The strategies used are many and varied and are particularly well exemplified by the interaction of viral gene products with the MHC class I assembly pathway. Here, we provide an overview of what is currently known about the cellular biochemistry of antigen processing and the assembly of class I and class II MHC molecules. PMID- 11256574 TI - Emerging principles for T cell receptor recognition of antigen in cellular immunity. AB - The structural basis of antigen recognition in cellular immunity has been elucidated through the determination of crystal structures of major histocompatibility complex (MHC) molecules bound to antigenic peptides, T cell receptors (TCR), CD8 and CD4 co-receptors and, most recently, TCRs in complex with peptide-MHC (pMHC). The mechanisms that generate the diversity of the immune response to invading microorganisms were first realized at a genetic level and are necessary in order to cope with the enormous number of potential antigens. This diversity is manifested in the protein products of the genes which code for the components of the TCR signalling complex. The structure of the TCR reveals both striking similarities with and fundamental differences from its functional counterpart, the antibody, in the humoral immune system. The conserved manner in which the TCR recognizes and interacts with its peptide-MHC ligand allows the TCR great latitude in its potential to form productive interactions with antigen presenting cells that bear numerous ligands to which the TCR has not been previously exposed. This phenomenon of cross-, or alloreactivity arises from a combination of conserved structural features across all MHC molecules, both self and foreign, and some degree of molecular mimicry. Non-classical MHC ligands presenting either modified or specialized peptides, lipids, carbohydrates, or no ligand at all, are now thought to play increasingly important roles in cellular immunity. We review some of the recent structural results and our current state of knowledge about TCR structure, and how this relates to its function. PMID- 11256575 TI - MHC and T cell development. AB - The ability to discriminate self from non-self is a fundamental property of the immune system. In the case of T lymphocytes, the first level of this discrimination takes place in the thymus, where most lymphocytes carrying an alphabeta T cell receptor (TCR) become tolerant to self-epitopes represented within the thymic microenvironment and differentiate into CD4+ or CD8+ single positive thymocytes. In the periphery, these subsets correspond respectively to helper and cytolytic lymphocytes able to react to non-self antigens presented in the context of MHC class II and I molecules. Apart from an early phase, the development of alphabeta T cells is based on a TCR-MHC interaction which is allele-specific and, depending on its nature, leads to either protection from apoptosis and maturation (positive selection) or physical elimination of thymocytes (negative selection). Thus, these positive and negative selection processes concomitantly allow the rescue of the useful fraction and the elimination of the potentially harmful fraction of the TCR repertoire. Recent advances have provided important elements for the comprehension of the development of alphabeta T cells. In accordance with previous in vitro studies related to differentiation of CD8+ thymocytes, in vivo derived data have established that the positive selection of CD4+ thymocytes is a peptide-specific process: it is based on the intrathymic TCR recognition of self-peptide:self-MHC molecular complexes. Despite this fact, it is now clear that the TCR reactivity to non-self MHC molecules or alloreactivity--a major characteristic of the mature TCR repertoire--does not result from intrathymic T cell selection, but rather is an intrinsic property of germline-encoded TCR domains. Finally, a significant number of experiments indicate that, in secondary lymphoid organs, a repeated TCR MHC low affinity interaction is required to maintain the mature peripheral T cell pool and therefore the mature TCR repertoire. Such a TCR-MHC interaction-induced protection from apoptosis is remarkably reminiscent of the intrathymic positive selection phenomenon. Thus, the role of self-MHC recognition in TCR repertoire development and survival may account for the influence of MHC genotype on susceptibility to specific autoimmune diseases. PMID- 11256576 TI - Generation of mouse mutants as a tool for functional genomics. AB - As sequence information becomes available from the Human Genome Project, key developments include systematic methods for assigning function to each of the 100,000 or so genes. Strategies for coping with this sequence information, including microarray analysis and proteomics, will further our understanding of how genes function and interact. Ultimately, however, the simplest way to understand how a gene works is to examine the consequences of interference with its function: mutational analysis. The mouse represents the model organism of choice in the analysis of gene function; close enough to human to represent a satisfactory model organism, yet relatively easy to manipulate at a genetic level. Two complementary approaches, genotype- and phenotype-based, have been established in the mouse genetics and genomics communities to systematically generate new mouse mutations. Genotype-based approaches are advantageous in that molecular analysis of mutations is facilitated. Phenotypic analysis, however, is often assumed based on gene expression patterns, often leading to unexpected results. Phenotype-based approaches do not make prior assumptions about gene function. Often, however, it may be difficult to define the underlying genetic lesion. Progress in each of these approaches will be considered and situations in which they might be mutually beneficial will be investigated. PMID- 11256577 TI - Automated protein modelling--the proteome in 3D. AB - Functional analysis of the proteins discovered in fully sequenced genomes represent the next major challenge of life science research. Computational methods play an increasingly important role in this activity. Among them, comparative protein modelling will play a major role in this challenge, especially in the light of the Structural Genomics programmes about to be started around the world. In recent years, much progress has been made in automating these methods, enabling the production of models for genome scale problems. In this review we discuss how protein models can be applied to functional analysis, as well as some of the current issues and limitations inherent to these methods. PMID- 11256578 TI - The use of proteomics in ophthalmic research. AB - The goal of molecular ophthalmology is the early detection and therapeutic treatment of eye disease. Genomic technologies have profoundly enhanced the discovery of ocular disease candidate genes. Proteomics, the protein cognate of genomic technology, offers a means to monitor changes in the expression of a given ocular protein(s) and its post-translational modification, identify novel therapeutic targets and evaluate pharmacological effects on a given metabolic pathway. Using both tissue and cultured cells, numerous laboratories have begun to catalogue changes in ocular protein expression in normal, diseased and ageing subjects. Herein, we review published proteomic literature in the broad context of ophthalmic diseases involving various tissues of the eye. PMID- 11256579 TI - Expression databases--resources for pharmacogenomic R&D. AB - This review aims to outline the primary biological databases that are being generated to understand fundamental biology, identify new drug targets, and to look at compound profiling in a new light. We will give a brief overview of four of the main areas being studied in molecular biology: genomics, pharmacogenomics, pharmacogenetics and proteomics. Looking initially at each data set and some of its potential applications, we will go on to describe some of the potentially enormous advantages gained by fully integrating these data sets. PMID- 11256580 TI - Applications of biochip and microarray systems in pharmacogenomics. AB - A DNA microarray system is usually comprised of DNA probes formatted on a microscale on a glass surface (chip), plus the instruments needed to handle samples (automated robotics), to read the reporter molecules (scanners) and analyse the data (bioinformatic tools). Biochips are formed by in situ (on chip) synthesis of oligonucleotides or peptide nucleic acids (PNAs) or spotting of DNA fragments. Hybridisation of RNA- or DNA-derived samples on chips allows the monitoring of expression of mRNAs or the occurrence of polymorphisms in genomic DNA. Basic types of DNA chips are the sequencing chip, the expression chip and chips for comparative genomic hybridisation. Advanced technologies used in automated microarray production are photolithography, mechanical microspotting and ink jets. Bioelectronic microchips contain numerous electronically active microelectrodes with specific DNA capture probes linked to the electrodes through molecular wires. Several biosensors have been used in combination with biochips. PNA biosensors commonly rely on the immobilisation of a single-stranded DNA sequence (the 'probe') onto a transducer surface for hybridisation with the complementary ('target') strand to give a suitable electrical signal. Other sensors are cell-based immunobiosensors with engineered molecular recognition, integrated biosensors based on phototransistor integrated circuits and sensors based on surface plasmon resonance. Microarray technologies offer enormous savings in time and labour as compared to standard gel-based microsatellite methods. Reading of the information and its management by bioinformatics is necessary because of the enormous amount of data generated by the various technologies using microarrays. Standardised procedures are essential for compatible data production, quality control and analysis. Expression monitoring is the most biologically informative application of this technology at present. Microarray technology has important applications in pharmacogenomics: drug discovery and development, drug safety and molecular diagnostics. DNA chips will facilitate the integration of diagnosis and therapeutics, as well as the introduction of personalised medicines. PMID- 11256581 TI - The DRD2 gene in psychiatric and neurological disorders and its phenotypes. AB - The TaqIA D2 dopamine receptor (DRD2) minor (A1) allele was first associated with severe alcoholism a decade ago. Since then, studies both confirming and not confirmnning this finding were reported. However, a meta-analysis of a large number of Caucasian alcoholics (both more severe and less severe) and controls (both assessed and unassessed for substance use disorders) revealed a significantly higher frequency (p < 10(-6)) and prevalence (p < 10(-8)) of the DRD2 A1 allele in the alcoholics. Further analysis showed that the more severe alcoholics had a 3-fold higher prevalence of the DRD2 A1 allele than the assessed controls (p < 10(-10)), whereas no difference was found between the less severe alcoholics and the unassessed controls. DRD2 exonic or promoter mutations have not yet been associated with alcoholism, although two intronic variants at the TaqIB and intron 6 sites, which are in linkage disequilibrium with the TaqIA site, were associated with this disorder. Variants of the DRD2 gene have also been associated with cocaine, nicotine and opioid dependence, obesity and gambling. It is hypothesised that the DRD2 is a reinforcement or reward gene. Although less intensively studied than substance use disorders, the DRD2 gene has been implicated in Tourette's syndrome (TS), post-traumatic stress disorder (PTSD) and certain symptoms associated with affective disorders and schizophrenia. Further, DRD2 variants have been implicated in Parkinson's disease (PD) and in iatrogenically-induced movement disorders, as well as in certain migraineurs. Phenotypic differences have been associated with DRD2 variants. These include reduced D2 dopamine receptor numbers and diminished glucose metabolism in the brain of subjects who carry the DRD2 A1 allele. In addition, phenotypic differences have been found in neurocognitive and personality characteristics, and in treatment outcome of DRD2 variants. The involvement of the DRD2 gene in certain neuropsychiatric disorders opens up the potential of a targeted pharmacogenomic approach to the prevention and treatment of these disorders. PMID- 11256582 TI - Serotonin transporter polymorphism and response to SSRIs in major depression and relevance to anxiety disorders and substance abuse. AB - The selective serotonin re-uptake inhibitors (SSRIs) which modulate serotonergic activity are effective in the treatment of serotonin-related mental disorders, such as depression and anxiety. These agents bind to the serotonin transporter (5 HTT) and inhibit its capacity to transport serotonin (5-hydroxytryptamine; 5-HT). A functional polymorphism in the promoter region of 5-HTT (5-HTTLPR) has been described. The insertion variant of this polymorphism (long allele) is associated with higher expression of brain 5-HTT compared to the deletion variant (short allele). An association between the 5-HTTLPR polymorphism and mental disorders has been reported by some, but not all, investigators. In addition, the 5-HTT gene polymorphisms were found to be associated with a better and faster response to SSRIs with or without pindolol augmentation in depressed patients. Further studies are needed to clarify the relationship between the 5-HTT genotype, the susceptibility to mental disorders, the response to serotonergic agents and the side effect profile. PMID- 11256583 TI - Insulin receptor substrate polymorphisms and type 2 diabetes mellitus. AB - Insulin receptor substrate (IRS) molecules are key mediators in insulin signalling and play a central role in maintaining basic cellular functions, such as growth, survival and metabolism. They act as docking proteins for the insulin receptor and a complex network of intracellular signalling molecules containing Src homology 2 (SH2) domains. Four members (IRS-1, IRS-2, IRS-3 and IRS-4) of this family have been identified that differ in tissue distribution, subcellular localisation, developmental expression, binding to the insulin receptor and interaction with SH2 domain-containing proteins. Results from targeted disruption of the IRS genes in mice have provided important clues as to the functional differences among these related molecules and suggest that they play very different roles in vivo. The available data are consistent with the notion that both IRS-1 and IRS-2 are important for insulin action and glucose homeostasis in vivo, whereas IRS-and IRS-4 appear to play a redundant role in the IRS signalling system. Considering their key role in both insulin action and insulin secretion, IRS-1 and IRS-2 molecules have been considered plausible candidate genes involved in the pathogenesis of Type 2 diabetes. Several polymorphisms in the IRS genes have been identified, but only the Gly --> Arg72 substitution of IRS-1, acting with environmental factors, seems to have a pathogenic role in the development of Type 2 diabetes. In contrast, polymorphisms of the other IRS genes do not appear to contribute to Type 2 diabetes. PMID- 11256584 TI - Accelerating discoveries in the proteome and genome with MALDI TOF MS. AB - Recent developments in mass spectrometry (MS) provide scientists with an established analytical tool that addresses the demands for rapid, accurate and cost effective analyses of biomolecules. These advances clearly accelerate the rate and success of protein identification, genetic sequencing, determining biological variances and drug discovery. This review is intended to illustrate how matrix-assisted laser desorption/ionisation (MALDI) time-of-flight (TOF) MS is typically used to generate information about biomolecules under investigation. Additionally, examples will be used to describe the steps involved in preparing samples for MALDI TOF and obtaining answers through data management. PMID- 11256585 TI - Current pharmacogenomic approaches to clinical drug development. AB - Pharmacogenomics has recently become an integral part of the drug development process. The pharmacogenomics revolution comes at a time when pharmaceutical companies are faced with mounting pressures to lower the cost of drugs despite the continued rise in research and development spending needed to bring new drugs to market. Pharmaceutical companies want to avoid late stage failures or drugs labelled for restricted use following approval. More than twenty years of pharmacogenetic studies have established many of the genetic traits responsible for interindividual differences in the way patients metabolise drugs. The genetic polymorphisms found in the major drug metabolising enzymes (DMEs) and their associated phenotypes are well established. These monogenetic traits have a predictable influence on the pharmacokinetic and pharmacological effects of a large number of commonly prescribed drugs. This knowledge has been used to develop affordable, robust, clinical genotyping methods that can be used by pharmaceutical companies to screen patients prior to drug therapy. Prospective screening of Phase I volunteers for DME polymorphisms is done routinely at a number of pharmaceutical companies. As the pharmacogenomic initiatives at these companies evolve, more and more patients enrolled in Phase II-III clinical trials are genotyped to correlate efficacy with genetic markers that predict pharmacodynamic effects. There are a number of pharmacogenomic markers that provide useful diagnostic tools to prospectively evaluate treatment regimens, including the genetics of the host, cancerous tumours or infectious agents. The incorporation of pharmacogenomics into clinical drug development offers the opportunity for pharmaceutical companies to evaluate drugs with a better understanding of the effect that specific genetic variants will have on drug response. Prospective testing can ensure the inclusion of important phenotypic subgroups, thus impacting the efficiency of drug development. Ultimately this approach will validate the utility of genotyping prior to prescription, thereby ensuring that patients receive the right drug at the right dose the first time. PMID- 11256586 TI - Pharmacogenetic diagnostics of cytochrome P450 polymorphisms in clinical drug development and in drug treatment. AB - The current use and future perspectives of molecular genetic characterisation of cytochrome P450 enzymes (CYP) for drug development and drug treatment are summarised. CYP genes are highly polymorphic and the enzymes play a key role in the elimination of the majority of drugs from the human body. Frequent variants of some enzymes, CYP2A6, 2C9, 2C19 and 2D6, should be analysed in participants of clinical trials whenever these enzymes may play a role. It is suggested that a CYP genotype certificate is handed out to the volunteers or patients to avoid replicate analyses, and to allow that this information is available for future research and also for treatment with eventually needed drugs. Guidelines on what CYP alleles have to be analysed in drug development, as well as on analytical validation and CYP genotype data handling will be required. Treatment with several drugs may be improved by prior genotyping. The concepts and problems of CYP genotype-based clinical dose recommendations are presented and illustrated for selected drugs. The requirement for prospective trials on the medical and economic benefits of routine CYP genotyping is emphasised. Specific operationally defined recommendations dependent on genotype are a prerequisite for such studies and this review presents tentative CYP genotype-based dose recommendations systematically calculated from published data. Because of the multiplicity of factors involved, these doses will not be the optimal doses for each given individual, but should be more adequate than doses generally recommended for an average total population. Those CYP alleles and polymorphically metabolised drugs which are currently most interesting in drug development and drug treatment are reviewed, and more complete information is available from websites cited in this article. PMID- 11256588 TI - DNA microarray technology and antimicrobial drug discovery. AB - The genomics era is providing us with vast amounts of information derived from whole-genome sequencing. This will doubtlessly revolutionise biology and the way novel medicines will be discovered. To leverage this information efficiently, however, technologies in addition to high-throughput sequencing are required. DNA microarray technology is one technology that has already shown great potential for both basic research and drug discovery. With particular emphasis on antibacterial research we will summarise in this review the key technological aspects and most important applications of DNA microarrays demonstrated so far. PMID- 11256587 TI - The DD-ACE genotype and cardiovascular disease. AB - The renin-angiotensin-aldosterone system (RAS) plays a pivotal role in the cardiovascular system, and the therapeutic agents which interact with this pathway have a significant impact in both heart failure and following myocardial infarction (MI). Polymorphisms within the genes controlling this enzyme system may also contribute to the pathogenesis of cardiovascular disease. Over the last decade an association between a polymorphism of the angiotensin converting enzyme (ACE) gene (called the DD-ACE polymorphism) and phenotypic expression of cardiovascular disease, namely MI, has been reported. Since then, several small case-controlled studies have confirmed an association with manifestations of ischaemic heart disease or various other cardiac end points. However, in a large prospective study the ACE gene was found to confer no appreciable risk. This review article considers the evidence that links polymorphisms of the ACE gene with cardiovascular disease. The Medline database (1990 - 2000) was searched using the keywords myocardial infarction, ischaemic heart disease, angiotensin converting enzyme, polymorphisms (a search of the reference citations of relevant articles was also performed), and clinical studies on cardiovascular disease related to the ACE genotype were selected. Taken together, the available evidence supports the notion that the DD-ACE genotype adversely influences specific cardiovascular diseases, but appears to do so in specific geographical areas and in particular patient subgroups. It is not yet known whether it does this through an interaction with other genes or by as yet unexplained biochemical mechanisms. However, the impact of the DD-ACE genotype appears to be small and its clinical manifestations rather heterogeneous. This finding is not in contrast to the overall impact of the renin-angiotensin system in cardiovascular disease, given the fact that the ACE enzyme is only one component in the renin-angiotensin cascade and that one genetic variant cannot be expected to contribute more than a minor individual impact in genetically complex multifactorial cardiovascular disease. PMID- 11256589 TI - Human adipocyte proteomics--a complementary way of looking at fat. PMID- 11256590 TI - Qualitative gene profiling: a novel tool in genomics and in pharmacogenomics that deciphers messenger RNA isoforms diversity. AB - RNA splicing is a tightly regulated process. It is essential for gene expression and, therefore, intervenes in every biological phenomenon in mammals. RNA splicing regulation is cell type-specific in such a way that a cellular situation can be characterised by its repertoire of spliced events, the spliceome. Comparison of the splicing repertoire of two situations identifies alternative exons and introns. This regulation involves cis-acting sequences and transacting factors. Mutations, as well as modifications of signalling pathways, can alter the accuracy of splicing. Since deletion of exons or retention of introns within coding sequences modifies the corresponding proteins and functional domains of proteins are encoded by contiguous exons, identifying changes in the spliceome pinpoints functional domains, which are specifically regulated at the level of RNA splicing. We have developed a new method of gene profiling, qualitative gene profiling, that allows the comparative study of the repertoires of spliced events that characterise distinct physiopathological situations. We present in this review the different uses of this new genomic technique that can help each step of the R&D process in the pharmaceutical industry, and that represents a short cut towards functional genomics and pharmacogenomics. PMID- 11256591 TI - Phage display and pharmacogenomics. AB - Surface display of genetic diversity is a technology that can produce specific binding agents for almost any target molecule, and is especially well suited for making agents that bind specific proteins. Until now, pharmacogenomic studies have followed the response of cells to drugs and other agents by tracking the mRNAs that encode the proteins of interest, because it is relatively easy to produce nucleic acid ligands once the gene sequence is known. Nevertheless, in some cases, direct tracking of the proteins is preferred. Phage display of peptides and proteins (especially antibodies) is now able to provide the large number of specific binding agents needed to track proteins in pharmacogenomic studies. PMID- 11256592 TI - Therapeutic target discovery using Caenorhabditis elegans. AB - Use of the human genome sequence in disease therapy will require efficient identification of disease-causing and disease-associated genes with functions that are amenable to pharmacological manipulation. The validation and development of such genes as therapeutic targets requires information about both the genes' functions and the biochemical pathways in which they participate. One powerful means of obtaining such information is the study of homologous genes in model organisms amenable to laboratory manipulation. Among model organisms the nematode Caenorhabditis elegans offers several advantages, including well-established techniques for genetic and experimental manipulation and the first completed genome sequence for a multicellular organism. Molecular genetic experiments using C. elegans can contribute at several levels to drug discovery programs, from elucidation of genetic functions and pathways to the validation of candidate targets. Additionally, the ease of culture allows adaptation of the nematode for use in high-throughput chemical screens for the identification of lead compounds in drug development. PMID- 11256593 TI - Large-scale SNP scoring from unamplified genomic DNA. AB - Discoveries from the Human Genome Project (HGP) continue to spur changes in medical technology that will lead to new diagnostic procedures in the clinical lab. As more single nucleotide polymorphisms (SNPs) are discovered and correlated to human diseases, demands for genetic tests will increase. The enormity of the number of SNPs makes developing inexpensive and reliable high-throughput methods for SNP scoring imperative. High-throughput screening (HTS) means, at a minimum, a production rate of thousands of assays per day. Ideally, the technology will be easy, inexpensive and amenable to automation. The Invader assay offers a simple diagnostic platform to detect single nucleotide changes with high specificity and sensitivity from unamplified, genomic DNA. The Invader assay uses a structure specific 5' nuclease (or flap endonuclease) to cleave sequence-specific structures in each of two cascading reactions. The cleavage structure forms when two synthetic oligonucleotide probes hybridise in tandem to a target. One of the probes cycles on and off the target and is cut by the nuclease only when the appropriate structure forms. These cleaved probes then participate in a second Invader reaction involving a dye-labelled fluorescence resonance energy transfer (FRET) probe. Cleavage of this FRET probe generates a signal, which can be readily analysed by fluorescence microtitre plate readers. The two cascading reactions amplify the signal significantly; each original target molecule can lead to more than 10(6) cleaved signal probes in one hour. This signal amplification permits identification of single base changes directly from genomic DNA without prior target amplification. The sequences of the oligonucleotide components of the secondary reaction are independent of the target of interest and permit the development of universal secondary reaction components useful to identify any target. PMID- 11256594 TI - Approaches to allele frequency determination. AB - Hundreds of thousands of candidate single nucleotide polymorphisms (SNPs) are being identified as part of the human genome project. For these markers to be useful in any study their allele frequencies in the study population must be known, or much effort will be wasted when markers with the wrong characteristics are selected for studies involving large-scale genotyping of SNP markers. Because allele frequency estimations by genotyping a representative sample of a population is a costly endeavour, an obvious strategy to reduce the cost is to identify accurate and efficient approaches to allele frequency estimation using DNA pools. There are several allele frequency estimation approaches currently in use, including PCR-RFLP analysis, DNA sequencing, the Taqman assay and kinetic PCR. All these approaches give reasonably good allele frequency estimates. Accurate and efficient allele frequency estimation in DNA pools will reduce the cost and effort needed in genetic and association studies and opens up ways to perform pharmacogenomic and evolutionary studies efficiently. PMID- 11256595 TI - Does Europe exist? PMID- 11256596 TI - Our treacherous genes. The perils of an information explosion. PMID- 11256597 TI - Rewarding true innovation. Experimental use exemption and the trends in gene patenting. PMID- 11256598 TI - Genomics down under. A talk with John Mattick Director of the Australian Genome Research Facility, and Co-Director of the Institute for Molecular Bioscience. Interview by Holger Breithaupt. PMID- 11256599 TI - A meeting at the gene. Biodiversity and natural history. PMID- 11256600 TI - Would you buy a tomato from this man? How to overcome public mistrust in scientific advances. PMID- 11256601 TI - Bear market slashes at human genome. The dropping guesses about the number of human genes challenges researchers to explain human complexity with so few genes. PMID- 11256602 TI - Composing life. AB - Textbooks often assert that life began with specialized complex molecules, such as RNA, that are capable of making their own copies. This scenario has serious difficulties, but an alternative has remained elusive. Recent research and computer simulations have suggested that the first steps toward life may not have involved biopolymers. Rather, non-covalent protocellular assemblies, generated by catalyzed recruitment of diverse amphiphilic and hydrophobic compounds, could have constituted the first systems capable of information storage, inheritance and selection. A complex chain of evolutionary events, yet to be deciphered, could then have led to the common ancestors of today's free-living cells, and to the appearance of DNA, RNA and protein enzymes. PMID- 11256603 TI - ATP synthase. Is revolution effective? PMID- 11256604 TI - Aggresomes and Russell bodies. Symptoms of cellular indigestion? AB - All cells are equipped with a proteolytic apparatus that eliminates damaged, misfolded and incorrectly assembled proteins. The principal engine of cytoplasmic proteolysis, the 26S proteasome, requires that substrates be unfolded to gain access to the active site; consequently, it is relatively ineffective at degrading aggregated proteins. Cellular indigestion occurs when the production of aggregation-prone proteins exceeds the cell's (or organelle's) capacity to eliminate them. Cellular pathways that resolve this indigestion exist, but appear to have limited capacities. Russell bodies and aggresomes are manifestations of cellular indigestion in the endoplasmic reticulum and cytoplasmic compartments, respectively, and are often associated with disease. PMID- 11256605 TI - DNA motif associated with meiotic double-strand break regions in Saccharomyces cerevisiae. AB - Meiotic recombination in yeast is initiated by DNA double-strand breaks (DSBs) that occur at preferred sites, distributed along the chromosomes. These DSB sites undergo changes in chromatin structure early in meiosis, but their common features at the level of DNA sequence have not been defined until now. Alignment of 1 kb sequences flanking six well-mapped DSBs has allowed us to define a flexible sequence motif, the CoHR profile, which predicts the great majority of meiotic DSB locations. The 50 bp profile contains a poly(A) tract in its centre and may have several gaps of unrelated sequences over a total length of up to 250 bp. The major exceptions to the correlation between CoHRs and preferred DSB sites are at telomeric regions, where DSBs do not occur. The CoHR sequence may provide the basis for understanding meiosis-induced chromatin changes that enable DSBs to occur at defined chromosomal sites. PMID- 11256606 TI - Point mutation of bacterial artificial chromosomes by ET recombination. AB - Bacterial artificial chromosomes (BACs) offer many advantages for functional studies of large eukaryotic genes. To utilize the potential applications of BACs optimally, new approaches that allow rapid and precise engineering of these large molecules are required. Here, we describe a simple and flexible two-step approach based on ET recombination, which permits point mutations to be introduced into BACs without leaving any other residual change in the recombinant product. Introduction of other modifications, such as small insertions or deletions, is equally feasible. The use of ET recombination to achieve site-directed mutagenesis opens access to a powerful use of BACs and is extensible to DNA molecules of any size in Escherichia coli, including the E. coli chromosome. PMID- 11256607 TI - Mammalian Ku86 protein prevents telomeric fusions independently of the length of TTAGGG repeats and the G-strand overhang. AB - Ku86 together with Ku70, DNA-PKcs, XRCC4 and DNA ligase IV forms a complex involved in repairing DNA double-strand breaks (DSB) in mammals. Yeast Ku has an essential role at the telomere; in particular, Ku deficiency leads to telomere shortening, loss of telomere clustering, loss of telomeric silencing and deregulation of the telomeric G-overhang. In mammals, Ku proteins associate to telomeric repeats; however, the possible role of Ku in regulating telomere length has not yet been addressed. We have measured telomere length in different cell types from wild-type and Ku86-deficient mice. In contrast to yeast, Ku86 deficiency does not result in telomere shortening or deregulation of the G-strand overhang. Interestingly, Ku86-/- cells show telomeric fusions with long telomeres (>81 kb) at the fusion point. These results indicate that mammalian Ku86 plays a fundamental role at the telomere by preventing telomeric fusions independently of the length of TTAGGG repeats and the integrity of the G-strand overhang. PMID- 11256608 TI - The GAGA factor of Drosophila interacts with SAP18, a Sin3-associated polypeptide. AB - SAP18, a polypeptide associated with the Sin3-HDAC co-repressor complex, was identified in a yeast two-hybrid screen as capable of interacting with the Drosophila GAGA factor. The interaction was confirmed in vitro by glutathione S transferase pull-down assays using recombinant proteins and crude SL2 nuclear extracts. The first 245 residues of GAGA, including the POZ domain, are necessary and sufficient to bind dSAP18. In polytene chromosomes, dSAP18 and GAGA co localize at a few discrete sites and, in particular, at the bithorax complex where GAGA binds some silenced polycomb response elements. When the dSAP18 dose is reduced, flies heterozygous for the GAGA mutation Trl67 show the homeotic transformation of segment A6 into A5, indicating that GAGA-dSAP18 interaction contributes to the functional regulation of the iab-6 element of the bithorax complex. These results suggest that, through recruitment of the Sin3-HDAC complex, GAGA might contribute to the regulation of homeotic gene expression. PMID- 11256609 TI - BRCA1 can stimulate gene transcription by a unique mechanism. AB - Most familial breast and ovarian cancers have been linked to mutations in the BRCA1 gene. BRCA1 has been shown to affect gene transcription but how it does so remains elusive. Here we show that BRCA1 can stimulate transcription without the requirement for a DNA-tethering function in mammalian and yeast cells. Furthermore, the BRCA1 C-terminal region can stimulate transcription of the p53 responsive promoter, MDM2. Unlike many enhancer-specific activators, non-tethered BRCA1 does not require a functional TATA element to stimulate transcription. Our results suggest that BRCA1 can enhance transcription by a function additional to recruiting the transcriptional machinery to a targeted gene. PMID- 11256610 TI - Rapid caspase-3 activation during apoptosis revealed using fluorescence-resonance energy transfer. AB - Caspase-3 is a crucial component of the apoptotic machinery in many cell types. Here, we report the timescale of caspase-3 activation in single living cells undergoing apoptosis. This was achieved by measuring the extent of fluorescence resonance energy transfer within a recombinant substrate containing cyan fluorescent protein (CFP) linked by a short peptide possessing the caspase-3 cleavage sequence, DEVD, to yellow fluorescent protein (YFP; i.e. CFP-DEVD-YFP). We demonstrate that, once initiated, the activation of caspase-3 is a very rapid process, taking 5 min or less to reach completion. Furthermore, this process occurs almost simultaneously with a depolarization of the mitochondrial membrane potential. These events occur just prior to the characteristic morphological changes associated with apoptosis. Our results clearly demonstrate that, once initiated, the commitment of cells to apoptosis is a remarkably rapid event when visualized at the single cell level. PMID- 11256611 TI - A redox-dependent interaction between two electron-transfer partners involved in photosynthesis. AB - Ferredoxin:NADP+:reductase (FNR) catalyzes one terminal step of the conversion of light energy into chemical energy during photosynthesis. FNR uses two high energy electrons photoproduced by photosystem I (PSI) and conveyed, one by one, by a ferredoxin (Fd), to reduce NADP+ to NADPH. The reducing power of NADPH is finally involved in carbon assimilation. The interaction between oxidized FNR and Fd was studied by crystallography at 2.4 A resolution leading to a three-dimensional picture of an Fd-FNR biologically relevant complex. This complex suggests that FNR and Fd specifically interact prior to each electron transfer and disassemble upon a redox-linked conformational change of the Fd. PMID- 11256612 TI - Catalysis of serine oligopeptidases is controlled by a gating filter mechanism. AB - Proteases have a variety of strategies for selecting substrates in order to prevent uncontrolled protein degradation. A recent crystal structure determination of prolyl oligopeptidase has suggested a way for substrate selection involving an unclosed seven-bladed beta-propeller domain. We have engineered a disulfide bond between the first and seventh blades of the propeller, which resulted in the loss of enzymatic activity. These results provided direct evidence for a novel strategy of regulation in which oscillating propeller blades act as a gating filter during catalysis, letting small peptide substrates into the active site while excluding large proteins to prevent accidental proteolysis. PMID- 11256613 TI - The EXT1/EXT2 tumor suppressors: catalytic activities and role in heparan sulfate biosynthesis. AB - The D-glucuronyltransferase and N-acetyl-D-glucosaminyltransferase reactions in heparan sulfate biosynthesis have been associated with two genes, EXT1 and EXT2, which are also implicated in the inherited bone disorder, multiple exostoses. Since the cell systems used to express recombinant EXT proteins synthesize endogenous heparan sulfate, and the EXT proteins tend to associate, it has not been possible to define the functional roles of the individual protein species. We therefore expressed EXT1 and EXT2 in yeast, which does not synthesize heparan sulfate. The recombinant EXT1 and EXT2 were both found to catalyze both glycosyltransferase reactions in vitro. Coexpression of the two proteins, but not mixing of separately expressed recombinant EXT1 and EXT2, yields hetero oligomeric complexes in yeast and mammalian cells, with augmented glycosyltransferase activities. This stimulation does not depend on the membrane bound state of the proteins. PMID- 11256614 TI - Systematic subcellular localization of novel proteins identified by large-scale cDNA sequencing. AB - As a first step towards a more comprehensive functional characterization of cDNAs than bioinformatic analysis, which can only make functional predictions for about half of the cDNAs sequenced, we have developed and tested a strategy that allows their systematic and fast subcellular localization. We have used a novel cloning technology to rapidly generate N- and C-terminal green fluorescent protein fusions of cDNAs to examine the intracellular localizations of > 100 expressed fusion proteins in living cells. The entire analysis is suitable for automation, which will be important for scaling up throughput. For > 80% of these new proteins a clear intracellular localization to known structures or organelles could be determined. For the cDNAs where bioinformatic analyses were able to predict possible identities, the localization was able to support these predictions in 75% of cases. For those cDNAs where no homologies could be predicted, the localization data represent the first information. PMID- 11256615 TI - The ins and outs of protein phosphorylation. Workshop report: control of signaling by protein phosphorylation. PMID- 11256616 TI - A wheel invented three times. The molecular structures of the three carbonic anhydrases. PMID- 11256617 TI - The G x U wobble base pair. A fundamental building block of RNA structure crucial to RNA function in diverse biological systems. AB - The G x U wobble base pair is a fundamental unit of RNA secondary structure that is present in nearly every class of RNA from organisms of all three phylogenetic domains. It has comparable thermodynamic stability to Watson-Crick base pairs and is nearly isomorphic to them. Therefore, it often substitutes for G x C or A x U base pairs. The G x U wobble base pair also has unique chemical, structural, dynamic and ligand-binding properties, which can only be partially mimicked by Watson-Crick base pairs or other mispairs. These features mark sites containing G x U pairs for recognition by proteins and other RNAs and allow the wobble pair to play essential functional roles in a remarkably wide range of biological processes. PMID- 11256618 TI - Certain promise and uncertain peril. The debate on xenotransplantation. PMID- 11256619 TI - Curbing the nuclear activities of beta-catenin. Control over Wnt target gene expression. AB - Wnt molecules control numerous developmental processes by altering specific gene expression patterns, and deregulation of Wnt signaling can lead to cancer. Many Wnt factors employ beta-catenin as a nuclear effector. Upon Wnt stimulation, beta catenin heterodimerizes with T-cell factor (TCF) DNA-binding proteins to form a transcriptional activator complex. As the activating subunit of this complex, beta-catenin performs dual tasks: it alleviates repression of target gene promoters and subsequently it activates them. Beta-catenin orchestrates these effects by recruiting chromatin modifying cofactors and contacting components of the basal transcription machinery. Although beta-catenin and TCFs are universal activators in Wnt signaling, their target genes display distinct temporal and spatial expression patterns. Apparently, post-translational modifications modulate the interactions between TCFs and beta-catenin or DNA, and certain transcription factors can sequester beta-catenin from TCFs while others synergize with beta-catenin-TCF complexes in a promoter-specific manner. These mechanisms provide points of intersection with other signaling pathways, and contribute to the complexity and specificity of Wnt target gene regulation. PMID- 11256620 TI - An estimate of large-scale sequencing accuracy. AB - The accuracy of large-scale DNA sequencing is difficult to estimate without redundant effort. We have found that the mobile genetic element IS10, a component of the transposon Tn10, has contaminated a significant number of clones in the public databases, as a result of the use of the transposon in bacterial cloning strain construction. These contaminations need to be annotated as such. More positively, by defining the range of sequence variation in IS10, we have been able to determine that the rate of sequencing errors is very low, most likely surpassing the stated aim of one error or less in ten thousand bases. PMID- 11256622 TI - Going to the roots of the stem cell debate. The ethical problems of using embryos for research. PMID- 11256621 TI - The TGF-beta family member derriere is involved in regulation of the establishment of left-right asymmetry. AB - Although a number of genes that are involved in the establishment of left-right asymmetry have been identified, earlier events in the molecular pathway developing left-right asymmetry remain to be elucidated. Here we present evidence suggesting that the transforming growth factor-beta family member derriere is involved in the development of left-right asymmetry in Xenopus embryos. Ectopic expression of derriere on the right side can fully invert cardiac and visceral left-right orientation and nodal expression, and expression of a dominant negative form of derriere on the left side can partially randomize the left-right orientation and nodal expression. Moreover, while expression of the dominant negative derriere does not inhibit the activity of Vg1 directly, it can rescue the altered left-right orientation induced by Vg1. Vg1 can induce derriere in animal cap explants. These results suggest that derriere is involved in earlier molecular pathways developing the left-right asymmetry. PMID- 11256623 TI - Caenorhabditis elegans has a single pathway to target matrix proteins to peroxisomes. AB - All eukaryotes so far studied, including animals, plants, yeasts and trypanosomes, have two pathways to target proteins to peroxisomes. These two pathways are specific for the two types of peroxisome targeting signal (PTS) present on peroxisomal matrix proteins. Remarkably, the complete genome sequence of Caenorhabditis elegans lacks the genes encoding proteins specific for the PTS2 targeting pathway. Here we show, by expression of green fluorescent protein (GFP) reporters for both pathways, that the PTS2 pathway is indeed absent in C. elegans. Lack of this pathway in man causes severe disease due to mislocalization of PTS2-containing proteins. This raises the question as to how C. elegans has accommodated the absence of the PTS2 pathway. We found by in silico analysis that C. elegans orthologues of PTS2-containing proteins have acquired a PTS1. We propose that switching of targeting signals has allowed the PTS2 pathway to be lost in the phylogenetic lineage leading to C. elegans. PMID- 11256624 TI - Length recognition at the N-terminal tail for the initiation of FtsH-mediated proteolysis. AB - FtsH-mediated proteolysis against membrane proteins is processive, and presumably involves dislocation of the substrate into the cytosol where the enzymatic domains of FtsH reside. To study how such a mode of proteolysis is initiated, we manipulated N-terminal cytosolic tails of three membrane proteins. YccA, a natural substrate of FtsH was found to require the N-terminal tail of 20 amino acid residues or longer to be degraded by FtsH in vivo. Three unrelated sequences of this segment conferred the FtsH sensitivity to YccA. An artificially constructed TM9-PhoA protein, derived from SecY, as well as the SecE protein, were sensitized to FtsH by addition of extra amino acid sequences to their N terminal cytosolic tails. Thus, FtsH recognizes a cytosolic region of sufficient length (approximately 20 amino acids) to initiate the processive proteolysis against membrane proteins. Such a region is typically at the N-terminus and can be diverse in amino acid sequences. PMID- 11256625 TI - Prediction of structural domains of TAP reveals details of its interaction with p15 and nucleoporins. AB - Vertebrate TAP is a nuclear mRNA export factor homologous to yeast Mex67p. The middle domain of TAP binds directly to p15, a protein related to the nuclear transport factor 2 (NTF2), whereas its C-terminal domain interacts with various nucleoporins, the components of the nuclear pore complex (NPC). Here, we report that the middle domain of TAP is also similar to NTF2, as well as to regions in Ras-GAP SH3 domain binding protein (G3BP) and some plant protein kinases. Based on the known three-dimensional structure of NTF2 homodimer, a heterodimerization model of TAP and p15 could be inferred. This model was confirmed by site-directed mutagenesis of residues located at the dimer interface. Furthermore, the C terminus of TAP was found to contain a ubiquitin-associated (UBA) domain. By site directed mutagenesis we show that a conserved loop in this domain plays an essential role in mediating TAP-nucleoporin interaction. PMID- 11256626 TI - Masked antisense: a molecular configuration for discriminating similar RNA targets. AB - Antisense technology has great potential for the control of RNA expression, but there remain few successful applications of the technology. Expressed antisense RNA can effectively down-regulate expression of a gene over long periods, but cannot differentiate partly identical sequences, such as the mRNA of fusion genes or those with point mutants. We have designed a structured form of expressed antisense, which can discriminate between highly similar mRNA molecules. These 'masked' antisense RNAs have most of the antisense sequence sequestered within duplex elements, leaving a short single-stranded region to initiate binding to target RNA. After contacting the correct target, the structured RNA can unravel, releasing the masked antisense region to form a stable duplex with the mRNA. We demonstrate that suitable masked antisense RNA can discriminate between the two forms of BCR-ABL mRNA that result from the Philadelphia chromosomal translocations, as well as discriminating the normal BCR and ABL mRNA. PMID- 11256627 TI - Visualizing the spindle checkpoint in Drosophila spermatocytes. AB - The spindle assembly checkpoint detects defects in spindle structure or in the alignment of the chromosomes on the metaphase plate and delays the onset of anaphase until defects are corrected. Thus far, the evidence regarding the presence of a spindle checkpoint during meiosis in male Drosophila has been indirect and contradictory. On the one hand, chromosomes without pairing partners do not prevent meiosis progression. On the other hand, some conserved components of the spindle checkpoint machinery are expressed in these cells and behave as their homologue proteins do in systems with an active spindle checkpoint. To establish whether the spindle checkpoint is active in Drosophila spermatocytes we have followed meiosis progression by time-lapse microscopy under conditions where the checkpoint is likely to be activated. We have found that the presence of a relatively high number of misaligned chromosomes or a severe disruption of the meiotic spindle results in a significant delay in the time of entry into anaphase. These observations provide the first direct evidence substantiating the activity of a meiotic spindle checkpoint in male Drosophila. PMID- 11256628 TI - Flowering or wilting? Europe has to find a new way to continue basic research in plant science. PMID- 11256629 TI - Human Cdc25 A inactivation in response to S phase inhibition and its role in preventing premature mitosis. AB - The Cdc25 A phosphatase is required for the G1-S transition of the cell cycle and is overexpressed in human cancers. We found that it is ubiquitylated and rapidly degraded by the proteasome and that its levels increase from G1 until mitosis. By treating cells with the DNA synthesis inhibitor hydroxyurea, Cdc25 A rapidly decreased in abundance, and this was accompanied by an increase in Cdk2 phosphotyrosine content and a decrease in Cdk2 kinase activity. Cdc25 A overexpression altered the ability of cells to arrest in the presence of hydroxyurea, and caused them to undergo premature chromosome condensation. Cdc25 A overexpression could render tumor cells less sensitive to DNA replication checkpoints, thereby contributing to their genomic instability. PMID- 11256632 TI - World wide wisdom. Electronic publishing is moving ahead. PMID- 11256633 TI - Monitoring the ambient environment with diffusive samplers: theory and practical considerations. PMID- 11256631 TI - Coordinated response of mammalian Rad51 and Rad52 to DNA damage. AB - Biochemical analysis has shown that mammalian Rad51 and Rad52 interact and synergize in DNA recombination reactions in vitro, but these proteins have not been shown to function together in response to DNA damage in vivo. By analysis of murine cells expressing murine Rad52 tagged with green fluorescent protein (GFP) Rad52, we now show that DNA damage causes Rad51 and GFP-Rad52 to colocalize in distinct nuclear foci. Cells expressing GFP-Rad52 show both increased survival and an increased number of Rad51 foci, raising the possibility that Rad52 is limiting for repair. These observations provide evidence of coordinated function of Rad51 and Rad52 in vivo and support the hypothesis that Rad52 plays an important role in the DNA damage response in mammalian cells. PMID- 11256630 TI - Werner's syndrome protein (WRN) migrates Holliday junctions and co-localizes with RPA upon replication arrest. AB - Individuals affected by the autosomal recessive disorder Werner's syndrome (WS) develop many of the symptoms characteristic of premature ageing. Primary fibroblasts cultured from WS patients exhibit karyotypic abnormalities and a reduced replicative life span. The WRN gene encodes a 3'-5' DNA helicase, and is a member of the RecQ family, which also includes the product of the Bloom's syndrome gene (BLM). In this work, we show that WRN promotes the ATP-dependent translocation of Holliday junctions, an activity that is also exhibited by BLM. In cells arrested in S-phase with hydroxyurea, WRN localizes to discrete nuclear foci that coincide with those formed by the single-stranded DNA binding protein replication protein A. These results are consistent with a model in which WRN prevents aberrant recombination events at sites of stalled replication forks by dissociating recombination intermediates. PMID- 11256634 TI - Modular separation-based fiber-optic sensors for remote in situ monitoring. AB - A modular separation-based fiber-optic sensor (SBFOS) with an integrated electronically controlled injection device is described for potential use in remote environmental monitoring. An SBFOS is a chemical monitor that integrates the separation selectivity and versatility afforded by capillary electrophoresis with the remote and high sensitivity capabilities of fiber-optic-based laser induced fluorescence sensing. The detection module of the SBFOS accommodates all essential sensing components for dual-optical fiber, on-capillary fluorescence detection. An injection module, similar to injection platforms on micro-analysis chips, is also integrated to the SBFOS. The injection module allows for electronically controlled injection of the sample onto the separation capillary. The design and operational characteristics of the modular SBFOS are discussed in this paper. A micellar electrokinetic capillary chromatography mode of separation is employed to evaluate the potential of the sensor for in situ monitoring of neutral toxins (aflatoxins). The analytical figures of merit for the modular SBFOS include analysis times of between 5 and 10 min, separation efficiencies of approximately 10(4) theoretical plates, detection limits for aflatoxins in the mid-to-low nanomolar range, and controllable operation that results in sensor performance that is largely immune to sample matrix effects. PMID- 11256635 TI - Up close and personal. PMID- 11256636 TI - In situ laser-induced fluorescence (LIF) analysis of petroleum product contaminated soil samples. AB - General considerations of the calibrations of in situ measurements are presented and the concept of using an "average oil" with average analysability for calibration purposes is introduced. The in situ analysis of 30 petroleum product contaminated soil samples with laser-induced fluorescence (LIF) spectroscopy was performed. Compared to an uncontaminated laboratory reference (LR) soil, 23 soil samples exhibited significantly higher LIF signals, so that these soil samples were classified as contaminated. The repeatability and reproducibility of the in situ LIF analysis were investigated. For the calibration of the LIF data, two LR oils (a fuel oil and a crude oil) were employed. The degree of soil contamination with petroleum products ranged from the limit of detection (LOD) for LIF analysis (ca. 100 ppm), or below, to more than 10,000 ppm. The petroleum product concentrations determined with in situ LIF analysis reveal a reasonable correlation with the results of standard IR analysis after extraction of the contaminated soils. PMID- 11256637 TI - Isocyanates: measurement methodology, exposure and effects. PMID- 11256638 TI - The effect of oxidation and acidification on the speciation of heavy metals in sulfide-rich freshwater sediments using a sequential extraction procedure. AB - The speciation of metals in a contaminated, anoxic, sulfide-rich, freshwater sediment was determined experimentally, using a sequential extraction procedure based on the method of Tessier et al. Taking into account the advantages and disadvantages of sequential extractions, the applied methodology allowed the investigation of the influence of aeration and acidification on the distribution of various metals in the sediment. Aeration caused Zn and Cd to be released from sulfides. Carbonates were partly dissolved by the oxidation process, causing mobilisation of Ca. Fe became less mobile owing to a stronger binding to organic matter. The speciation of K, Al, Ni, Pb and Mn and to a lesser extent of Cu was not affected by aeration. As a result of acidification of the aerated sediment, Ca, Mn, Ni, Zn and Cd became more mobile owing to the dissolution of carbonates. PMID- 11256639 TI - Analysis of sediments from Shetland Island voes for polycyclic aromatic hydrocarbons, steranes and triterpanes. AB - A few days after the grounding of the oil tanker Braer on 5 January 1993, an Exclusion Zone was designated by Order under the Food and Environment Protection Act 1985, prohibiting the harvesting of farmed or wild shellfish within the Zone to prevent contaminated products reaching the market place. The order was progressively lifted for species that were found to be free of petrogenic taint and for which the polycyclic aromatic hydrocarbon (PAH) levels were within the range for reference samples. This Order, however, still remains in place for mussels (Mytilus edulis) as the PAH levels are higher than in reference mussels. To investigate the possible source of PAHs found in these mussels, sediments were collected from three reference and three Zone sites and their hydrocarbon compositions studied using the n-alkane composition and concentration, PAH composition and concentration and the sterane and triterpane composition. The reference site at Olna Firth was found to have the highest levels of 2-6-ring parent and branched PAHs, the highest concentration in one of the pooled sediments being 4,530 ng g(-1) dry weight. Values in the other two reference sites (Vaila Sound and Mangaster Voe) ranged from 248.7 to 902.2 ng g(-1) dry weight. PAH concentrations at the Zone sites (Sandsound Voe, Stromness Voe and Punds Voe) ranged from 641.0 to 2,766 ng g(-1) dry weight. The PAH data were normalised to the percentage of organic carbon and log-transformed prior to being analysed using principal component analysis. The mean total PAH concentrations for Zone sites were found not to be significantly different from the reference sites. The PAH concentration ratios were consistent with the main source of PAHs being pyrolysis. However, there was a petrogenic contribution, suggested by the presence of alkylated PAHs, with Punds Voe having the largest petrogenic hydrocarbon content. This was supported by the triterpane profiles and the presence of a UCM in the aliphatic chromatograms from Punds Voe sediments. PMID- 11256640 TI - Aquatic humus from an unpolluted Brazilian dark-brown stream: general characterization and size fractionation of bound heavy metals. AB - The main pool of dissolved organic carbon in tropical aquatic environments, notably in dark-coloured streams, is concentrated in humic substances (HS). Aquatic HS are large organic molecules formed by micro-biotic degradation of biopolymers and polymerization of smaller organic molecules. From an environmental point of view, the study of metal humic interactions is often aimed at predicting the effect of aquatic HS on the bioavailability of heavy metal ions in the environment. In the present work the aquatic humic substances (HS) isolated from a dark-brown stream (located in an environmental protection area near Cubatao city in Sao Paulo-State, Brazil) by means of the collector XAD-8 were investigated. FTIR studies showed that the carboxylic carbons are probably the most important binding sites for Hg(II) ions within humic molecules. 13C-NMR and 1H-NMR studies of aquatic HS showed the presence of constituents with a high degree of aromaticity (40% of carbons) and small substitution. A special five stage tangential-flow ultrafiltration device (UF) was used for size fractionation of the aquatic HS under study and for their metal species in the molecular size range 1-100 kDa (six fractions). The fractionation patterns showed that metal traces remaining in aquatic HS after their XAD-8 isolation have different distributions. Generally, the major percentage of traces of Mn, Cd and Ni (determined by ICP-AES) was preferably complexed by molecules with relatively high molecular size. Cu was bound by fractions with low molecular size and Co showed no preferential binding site in the various humic fractions. Moreover, the species formed between aquatic HS and Hg(II), prepared by spiking (determined by CVAAS), appeared to be concentrated in the relatively high molecular size fraction F1 (> 100 kDa). PMID- 11256641 TI - Remediation of a nonachloro biphenyl congener with zero-valent iron in subcritical water. AB - Dechlorination of a nonachloro biphenyl congener with zero-valent iron in water under high temperature and pressure was investigated over time. Temperature has the main influence on the speed of dechlorination. Determination of polychlorinated biphenyls (PCBs) according to the grade of chlorination was performed by gas chromatography with mass selective detection in single ion monitoring mode. Dechlorination results in a variety of lower chlorinated biphenyls. The level of chlorination decreases with time. The amount of PCB molecules decreases to one-third within 90 min at 250 degrees C and 100 atm. However, no increase of biphenyl could be detected over time. A first-order kinetic model fitted the data obtained. PMID- 11256643 TI - Phthalates: a ban too far? PMID- 11256642 TI - The changing nature of the 206Pb/207Pb isotopic ratio of lead in rainwater, atmospheric particulates, pine needles and leaded petrol in Scotland, 1982-1998. AB - The inductively coupled plasma-mass spectrometry (ICP-MS)-determined 206Pb/207Pb ratio of 145 samples of rainwater collected at 25 locations around Scotland during December 1997 and January 1998 and at three longterm monitoring stations in the northeast, central belt and southeast of the country from November 1997 to December 1998 averaged 1.144+/-0.017 (1 s). This represents a significant increase from the mean value of 1.120+/-0.016 recorded for the long-term sites in 1989 1991, only partly attributable to a concomitant increase in the 206Pb/207Pb ratio of leaded petrol from 1.075+/-0.013 to 1.088+/-0.007. The rainwater 206Pb/207Pb data for the late 1990s also contrast markedly with the lower 206Pb/207Pb ratios found for pine needle and atmospheric particulate samples from Scotland (e.g. Glasgow: 1.085+/-0.012 in 1985-1986, 1.099+/-0.007 in 1991-1992), England and Western Europe in this study for the period 1982-1992 when emissions of lead to the atmosphere from petrol-engined vehicles in the UK were approximately 2-9 times higher. The observed change in the lead isotopic signature of rainwater predominantly reflects the impact of measures, such as the introduction and growing uptake of unleaded petrol, to reduce car exhaust emissions of lead to the atmosphere in the UK. Based on the rainwater data, source apportionment calculations suggest a general decline in the contribution of leaded petrol to atmospheric lead in Scotland from 53-61% in 1989-1991 to 32 45% in 1997-1998, with a corresponding decline in the urban environment from 84 86% to 48-58%. PMID- 11256644 TI - Simultaneous monitoring of atmospheric methane and speciated non-methane hydrocarbon concentrations using Peltier effect sub-ambient pre-concentration and gas chromatography. AB - Sub-ambient trapping, used to pre-concentrate atmospheric samples for non-methane hydrocarbon (NMHC) analysis by gas chromatography, can also be used to measure ambient methane concentrations. Above a sample volume of 40 ml, a dynamic equilibrium is established between ambient and trapped methane allowing for simultaneous quantitative determinations of methane and NMHC. The temperature stability of the trap is critical for quantitative methane analysis and this can be achieved by Peltier effect cooling. Simultaneous measurements of methane and NMHC reduce the equipment required for field trips and can ease the interpretation and modelling of atmospheric data. The feasibility for deployment of the system in remote locations was demonstrated by running the apparatus virtually unattended for a 5-day period. The correlations between the concentrations of methane, ethane and ethene measured during this period are discussed. PMID- 11256645 TI - Chemical composition of individual aerosol particles in workplace air during production of manganese alloys. AB - Aerosol particle samples were collected at ELKEM ASA ferromanganese (FeMn) and silicomanganese (SiMn) smelters at Porsgrunn, Norway, during different production steps: raw material mixing, welding of protective steel casings, tapping of FeMn and slag, crane operation moving the ladles with molten metal, operation of the Metal Oxygen Refinement (MOR) reactor and casting of SiMn. Aerosol fractions were assessed for the analysis of the bulk elemental composition as well as for individual particle analysis. The bulk elemental composition was determined by inductively coupled plasma atomic emission spectrometry. For individual particle analysis, an electron microprobe was used in combination with wavelength dispersive techniques. Most particles show a complex composition and cannot be attributed to a single phase. Therefore, the particles were divided into six groups according to their chemical composition: Group I, particles containing mainly metallic Fe and/or Mn; Group II, slag particles containing mainly Fe and/or Mn oxides; Group III, slag particles consisting predominantly of oxidized flux components such as Si, Al, Mg, Ca, Na and K; Group IV, particles consisting mainly of carbon; Group V, mixtures of particles from Groups II, III and IV; Group VI, mixtures of particles from Groups II and III. In raw material mixing, particles originating from the Mn ores were mostly found. In the welding of steel casings, most particles were assigned to Group II, Mn and Fe oxides. During the tapping of slag and metal, mostly slag particles from Group III were found (oxides of the flux components). During movement of the ladles, most particles came from Group II. At the MOR reactor, most of the particles belonged to the slag phase consisting of the flux components (Group III). The particles collected during the casting of SiMn were mainly attributed to the slag phase (Groups III and V). Due to the compositional complexity of the particles, toxicological investigations on the kinetics of pure compounds may not be easily associated with the results of this study. PMID- 11256646 TI - Organic dust and gaseous contaminants at wood working shops. AB - Airborne dust bioaerosols, ammonia and formaldehyde levels were determined inside two different (ventilated and unventilated) wood working shops. Airborne dust was found at mean values of 4.3 and 3.01 mg m(-3). These levels were higher than that recommended by Egyptian environmental law [1 mg m(-3) indoor maximum allowable concentration (MAC) for hard wood]. The highest frequency of aerodynamic size distribution of airborne wood dust was detected at a diametre of 4.9 microm which was recorded during a machining operation. Total viable bacteria were recorded at a mean value of 10(4) colony-forming units (cfu) m(-3), whereas Gram-negative bacteria were found at very low counts (10(1) cfu m(-3)). Fungi levels were recorded at mean values of 10(3) and 10(2) cfu m(-3) in ventilated and unventilated shops, respectively. Penicillium, Aspergillus, Cladosporium and yeast species were dominant isolates. Moreover, actinomycetes were found at a mean value of 10(3) cfu m(-3) at both workshops. Ammonia was detected in relatively low concentrations (mean values of 457 and 623 microg m(-3)), whereas formaldehyde was found in relatively moderate concentrations (mean values of 0.42 and 0.64 ppm). PMID- 11256647 TI - Cathodic dissolution in the electrocoagulation process using aluminium electrodes. AB - All the authors working with aluminium electrodes in the electrocoagulation process have shown that a dissolution occurs at the cathode. This result cannot be explained by the electrochemical process in which only the anodes should be dissolved. The most probable reaction is a chemical attack by hydroxyl ions (generated during water reduction) on the aluminium cathode but nobody has proved it in the framework of the electrocoagulation process. So we are interested in determining what kind of reactions occurs at the cathode. For that, we have elaborated a batch pilot apparatus divided into two compartments, allowing measurement of gas formation taking place only in one compartment. The gases measurements were performed by mass spectrometry with helium as carrier gas. To validate our experimental protocol, the first experiments have been done with a stainless steel cathode: in this case, the results have indicated that the amount of created hydrogen is in good agreement with the values calculated using the second Faraday's law. The experiments realised with an aluminium cathode have shown that the hydrogen formation, in these conditions, was higher than those observed with the stainless steel cathode. All our investigations enable us to propose that with an aluminium cathode, hydrogen formation can be separated into two phenomena. The first one is due to an electrochemical reaction (water reduction), and the second one arises from a chemical reaction explaining the dissolution observed at the cathode. PMID- 11256648 TI - " Perspectives on lithium treatment of bipolar disorder" - by Mogens Schou: a commentary. PMID- 11256649 TI - " Perspecives on lithium treatment of bipolar disorder" - by Mogens Schou: a commentary. PMID- 11256650 TI - Discontinuing lithium maintenance treatment in bipolar disorders: risks and implications. AB - OBJECTIVE: To review research findings on clinical effects of discontinuing lithium maintenance treatment. METHODS: Data from studies reported since 1970 plus our recent findings were updated. RESULTS: Discontinuing lithium maintenance treatment led to marked increases of early affective morbidity and suicidal risk. Gradual discontinuation markedly reduced early recurrences of mania or depression, did so more in bipolar II than I disorder patients, and also tended to reduce suicidal risk. Similar effects were found in pregnant and nonpregnant women after lithium discontinuation. Long-term retreatment with lithium following discontinuation was only slightly less effective than in initial trials. CONCLUSIONS: Recurrences increased sharply soon after discontinuing lithium, but were markedly limited and not merely delayed, by slow discontinuation. Similar reactions may follow discontinuation of other drugs, evidently as responses to long-term pharmacodynamic adaptations. Discontinuing treatment is not equivalent to not-treating. Post-discontinuation relapse risk has implications for the design, management, and interpretation of protocols involving discontinuation of long-term treatments that should be considered in both clinical management and research. PMID- 11256651 TI - A two-illness model of bipolar disorder. AB - The current approach to mood disorders is that bipolar disorder, comprising both mania and depression, is a discreet illness distinct from unipolar depression. This formulation has profoundly influenced the approach to understanding the biology and etiology of these disorders, as well as the manner in which the various phases of bipolar disorder are treated. Our new model suggests that bipolar disorder comprises two distinct illnesses, mania and depression, and that bipolar depression is no different from unipolar depression. Studies of clinical syndromes, course of illness, family history and genetics, biological factors, and treatment response data directly or indirectly support this new model. PMID- 11256652 TI - 50 years of lithium treatment of mood disorders. PMID- 11256653 TI - "A two-illness model of bipolar disorder"--by RT Joffe, LT Young, and GM MacQueen: a commentary. PMID- 11256654 TI - "A two-illness model of bipolar disorder"--RT Joffe, LT Young, and GM MacQueen: a commentary. PMID- 11256655 TI - "A two-illness model of bipolar disorder"--RT Joffe, LT Young, and GM MacQueen: a commentary. PMID- 11256656 TI - Topiramate as add-on treatment for patients with bipolar mania. AB - OBJECTIVE: Anticonvulsant agents such as carbamazepine and valproate are alternatives to lithium in treating subjects with bipolar disorder. Topiramate (Topamax), a new antiepileptic agent, is a candidate drug for bipolar disorder. We evaluated topiramate as adjunctive treatment for bipolar patients. METHODS: Eighteen patients with DSM-IV bipolar I disorder [mania (n = 12), hypomania (n = 1), mixed episode (n = 5), and rapid cycling (n = 6)], and two subjects with schizoaffective disorder bipolar type, resistant to current mood-stabilizer treatment were initiated on topiramate, 25 mg/day, increasing by 25-50 mg every 3 7 days to a target dose between 100 and 300 mg/day, as other medications were held constant for 5 weeks. The Young Mania Rating Scale (Y-MRS), Hamilton Depression Rating Scale (Ham-D), and Clinical Global Impression-Bipolar Version Scale (CGI-BP) were used to rate subjects weekly. RESULTS: By 5 weeks, 12 (60%) subjects were responders, i.e., 50% reduction in the Y-MRS scores and a CGI of 'much' or 'very much improved'. Three subjects were 'minimally improved', four showed no change, and one was 'minimally worse'. Six subjects had parasthesia, three experienced fatigue, and two had 'word-finding' difficulties; in all cases, side effects were transient. All patients lost weight with a mean of 9.4 lb in 5 weeks, and a significant reduction in body mass index (BMI) occurred too. CONCLUSIONS: Topiramate appears to have efficacy for the manic and mixed phases of bipolar illness. Other preliminary data suggest antidepressant efficacy too. Among obese bipolar subjects, the weight loss potential of topiramate may be beneficial. If controlled trials confirm these initial results, topiramate may be a significant addition to the available treatments for bipolar disorder. PMID- 11256657 TI - Perspectives on lithium treatment of bipolar disorder: action, efficacy, effect on suicidal behavior. AB - OBJECTIVE: To review studies of (A) whether lithium has a prophylactic action in bipolar disorder, (B) the efficacy of prophylactic lithium treatment in comparison with the efficacy of treatment with anticonvulsant drugs, and (C) the effect of lithium treatment on suicidal behavior. METHODS: Analysis of all relevant publications. RESULTS: (A) The claim that a prophylactic action of lithium has never been satisfactorily demonstrated is based on wrong assumptions, biased selection of references, and unjustified generalizations. (B) In typical bipolar disorder lithium is significantly more efficacious than carbamazepine; in atypical bipolar disorder there is a non-significant trend for carbamazepine to be better than lithium. Valproate has not been proven prophylactically efficacious in typical bipolar disorder; in atypical bipolar disorder it may have an effect, but it has not been compared with that of lithium. (C) A significant association has been found between prophylactic lithium treatment, on the one hand, and reduced mortality and suicidal behavior, on the other. No such association has been reported for prophylactic treatment with other mood stabilizers. CONCLUSION: In bipolar disorder the choice of prophylactic drug must be based on a weighing of efficacy against tolerability, interactions, ease of management, use during pregnancy and lactation, and expense. Lithium should be the preferred prophylactic drug in patients with typical bipolar disorder and in patients who are at high risk of committing suicide, that is, patients with severe depressions or depressions combined with persistent suicidal ideas or with suicide attempts in the past. PMID- 11256658 TI - Delirious mania. AB - OBJECTIVES: To define the characteristics of delirious mania. METHODS: A list of patients exhibiting both delirium and mania admitted to an academic psychiatric treatment unit of a tertiary care medical center was maintained for 6 years. A literature review for the terms 'delirium' and 'bipolar disorder' was undertaken. RESULTS: Few articles identify the syndrome. Most cite Bell (On a form of disease resembling some advanced stages of mania and fever. Am J Insanity 1849; 6: 97 127) as the first observer and Bond (Recognition of acute delirious mania. Arch Gen Psychiatry 1980; 37: 553 554) as the most recent. Fourteen instances were identified in the case list. Delirious mania is a syndrome of the acute onset of the excitement, grandiosity, emotional lability, delusions, and insomnia characteristic of mania, and the disorientation and altered consciousness characteristic of delirium. Almost all patients exhibited signs of catatonia. Bond (1980) recommends lithium and a neuroleptic combination as the treatment. In the present series, electroconvulsive therapy was found to be safe and rapidly effective, with all cases responding within three treatments and requiring less than six treatments in the course. The rapidity of response is the same as that of patients with catatonia. CONCLUSION: Delirious mania warrants specific identification in the diagnostic nomenclature. The distinction between delirious mania and the excited or malignant forms of catatonia requires study. PMID- 11256659 TI - Gabapentin in the acute treatment of refractory bipolar disorder. AB - BACKGROUND: Gabapentin, a new anti-epileptic agent, has been anecdotally reported to be effective in the treatment of mania. We systematically assessed the response rate in bipolar patients being treated adjunctively with gabapentin for manic symptoms, depressive symptoms, or rapid cycling not responsive to standard treatments. METHOD: Twenty-eight bipolar patients experiencing manic (n = 18), depressive (n = 5), or rapid-cycling (n = 5) symptoms inadequately responsive to at least one mood stabilizer were treated in an open fashion with adjunctive gabapentin. Illness response was assessed using the Clinical Global Impression Scale modified for bipolar disorder (CGI-BP). A 'positive response' was operationalized as a CGI response of much or very much improved. RESULTS: Fourteen of the 18 (78%) treated for hypomania or mania had a positive response to a dosage range of 600-3,600 mg/day. Patients with hypomania responded fastest, with a positive response achieved in 12.7 +/- 7.2 days. Patients with classic mania had a mean time to positive response of 25 +/- 12 days, and in patients with mixed mania it was 31.8 +/- 20.9 days. All of the five patients treated for depression had a positive response within 21 +/- 13.9 days. Only one of five patients with rapid cycling had a positive response. Gabapentin was well tolerated by all patients, with the most common side-effect being sedation. CONCLUSIONS: Gabapentin appears to have acute anti-manic and anti-depressant properties as an adjunctive agent for refractory bipolar illness. Prospective double-blind studies are needed to further delineate its acute efficacy when used as monotherapy and its prophylactic efficacy as monotherapy or in conjuction with other mood stabilizers. PMID- 11256660 TI - Clinical validation of genetic tests. PMID- 11256661 TI - Heterotaxy: associated conditions and hospital-based prevalence in newborns. AB - PURPOSE: To provide insight into the possible etiology and prevalence of heterotaxy, we studied conditions associated with heterotaxy in a consecutive hospital population of newborns. METHODS: From 1972 to March, 1999 (except February 16, 1972 to December 31, 1978), 58 cases of heterotaxy were ascertained from a cohort of 201,084 births in the ongoing Active Malformation Surveillance Program at the Brigham and Women's Hospital. This registry includes livebirths, stillbirths, and elective abortions. Prevalence among nontransfers (i.e., patients whose mothers had planned delivery at this hospital) was calculated as approximately 1 per 10,000 total births (20 of 201,084). RESULTS: We analyzed a total of 58 patients consisting of 20 (34%) nontransfers and 38 (66%) transfers. Patients were categorized by spleen status as having asplenia (7 nontransfers, 25 total), polysplenia (8, 20), right spleen (4, 11), normal left (0, 1), and unknown (1, 0). Among the 20 nontransfer and 59 total heterotaxy patients, the following associated medical conditions were present: chromosome abnormality (1 nontransfer, 2 total), suspected Mendelian or chromosome microdeletion disorder (1 nontransfer, 6 total), and maternal insulin-dependent diabetes mellitus (1 nontransfer, 2 total). There were 6 twins (1 member each from 6 twin pairs including 1 dizygous, 4 monozygous, 1 conjoined; 2 were nontransfers). An associated condition occurred in 5 (25%) nontransfer and 16 (28%) total patients, or among 10 of 53 singleton births (19%). CONCLUSIONS: Although most cases of heterotaxy in this series were sporadic events, an associated condition was present in about one-fourth of the cases. Not all of these conditions would be considered causative etiologies. Based on this small series alone, maternal insulin-dependent diabetes cannot be viewed as a risk factor for heterotaxy. However, the specific association of diabetes with polysplenia with/without left atrial isomerism is noteworthy, and adds weight to animal and epidemiologic case control data. PMID- 11256662 TI - Dural ectasia in the Marfan syndrome: MR and CT findings and criteria. AB - PURPOSE: To create criteria for detecting dural ectasia on MR or CT images in adult Marfan patients. METHODS: Images were analyzed using a workstation. Parameters that predicted dural ectasia were included in our criteria. RESULTS: Major criteria include: (1) width of dural sac below L5 > width above L4; (2) anterior sacral meningocele. Minor criteria include: (1) L5 nerve root sleeve diameter > 6.5 mm and (2) S1 scalloping > 3.5. Dural ectasia exists if 1 major or 2 minor criteria are present. CONCLUSION: MR and CT diagnose dural ectasia with high specificity and sensitivity. Our criteria accurately diagnose dural ectasia in adult Marfan patients. PMID- 11256663 TI - Family history-taking in community family practice: implications for genetic screening. AB - PURPOSE: To identify characteristics of physicians, patients, and visits associated with obtaining family history information in community family practice. METHODS: Research nurses directly observed 4,454 patient visits to 138 family physicians and reviewed office medical records. RESULTS: Family history was discussed during 51% of visits by new patients and 22% of visits by established patients. Physicians' rates of family history-taking varied from 0% to 81% of visits. Family history was more often discussed at well care rather than illness visits. The average duration of family history discussions was <2.5 minutes. CONCLUSIONS: These data can form the basis for realistic interventions to increase the use of family history in primary care. PMID- 11256664 TI - On the use of population-based registries in the clinical validation of genetic tests for disease susceptibility. AB - PURPOSE: Many new genetic tests for susceptibility to adult-onset diseases are developed on the basis of selected and high-risk groups. Before such tests can be used in medical practice, however, epidemiologic studies must be conducted to evaluate their clinical sensitivity, specificity, and positive predictive value in the general population. For many common adult-onset diseases, this process may take decades of follow-up. METHOD: We illustrate how clinical validation of new predictive genetic tests can be done retrospectively using case-control studies that are derived from population-based registries of diseases. We use the examples of birth defects and cancer registries to illustrate a hypothetical process by which such tests can be clinically validated. RESULTS: We demonstrate how such epidemiologic studies can be successfully used to derive measures of a test's sensitivity, specificity, positive predictive value, negative predictive value, and of the population attributable fraction of disease due to the disease susceptibility genes. Under certain assumptions, data derived from population based case-control studies provide adequate estimates of lifetime risks for disease (penetrance) among people with specified genotypes. CONCLUSIONS: With adequate protections of human subjects, studies involving population-based registries of disease will increasingly become valuable in validating the numerous genetic tests that will emerge from advances in human genetic research and the Human Genome Project. PMID- 11256665 TI - Some ethical implications of the Human Genome Project. PMID- 11256666 TI - Challenges in communicating genetics: a public health approach. PMID- 11256667 TI - Adverse psychological effects, reproductive uncertainty, and risk perception in 52 "carrier by non-carrier" CF parental pairs. PMID- 11256668 TI - The virtues of the virtual world. PMID- 11256669 TI - Molecular cytogenetics: show me the colors. PMID- 11256670 TI - Utility of multicolor fluorescent in situ hybridization in clinical cytogenetics. AB - PURPOSE: Multicolor FISH (M-FISH) was introduced in 1996 to scan all 24 chromosomes in different fluorescent colors by use of a specific filter set and computer software. However, the clinical utility of M-FISH has been limited because of the lack of commercial availability of reagents and hardware. We have evaluated M-FISH for identification of markers, derivative chromosomes, and complex karyotypes. METHODS: We present our findings based on a representative sample of one normal and six abnormal cases from a variety of tissue types. The results of M-FISH were confirmed by other well-established FISH probes. RESULTS: M-FISH analyses were successful in all six cases. The derivative chromosomes, ring, and a complex karyotype were resolved. CONCLUSIONS: We find M-FISH to be an invaluable tool for a high degree of accuracy and efficiency for chromosome identification. The limitations similar to spectral karyotyping system (SKY) include the inability to detect intrachromosomal anomalies, abnormalities involving the p-arms of acrocentrics and areas rich in highly repetitive DNA. In addition, there are some concerns of misinterpretation due to overlap of fluorophore combinations of different chromosomes, especially for subtle insertional translocations. PMID- 11256671 TI - Genealogy construction in a historically isolated population: application to genetic studies of rheumatoid arthritis in the Pima Indian. AB - PURPOSE: Due to the characteristics of complex traits, many traits may not be amenable to traditional epidemiologic methods. We illustrate an approach that defines an isolated population as the "unit" for carrying out studies of complex disease. We provide an example using the Pima Indians, a relatively isolated population, in which the incidence and prevalence of Type 2 diabetes, gallbladder disease, and rheumatoid arthritis (RA) are significantly increased compared with the general U.S. population. A previous study of RA in the Pima utilizing traditional methods failed to detect a genetic effect on the occurrence of the disease. METHODS: Our approach involved constructing a genealogy for this population and using a genealogic index to investigate familial aggregation. We developed an algorithm to identify biological relationships among 88 RA cases versus 4,000 subsamples of age-matched individuals from the same population. Kinship coefficients were calculated for all possible pairs of RA cases, and similarly for the subsamples. RESULTS: The sum of the kinship coefficient among all combination of RA pairs, 5.92, was significantly higher than the average of the 4,000 subsamples, 1.99 (p < 0.001), and was elevated over that of the subsamples to the level of second cousin, supporting a genetic effect in the familial aggregation. The mean inbreeding coefficient for the Pima was 0.00009, similar to that reported for other populations; none of the RA cases were inbred. CONCLUSIONS: The Pima genealogy can be anticipated to provide valuable information for the genetic study of diseases other than RA. Defining an isolated population as the "unit" in which to assess familial aggregation may be advantageous, especially if there are a limited number of cases in the study population. PMID- 11256672 TI - Additional evidence of linkage between Crohn's disease and a putative locus on chromosome 12. AB - PURPOSE: The inflammatory bowel diseases (IBD), Crohn's disease (CD), and ulcerative colitis (UC) are chronic intestinal disorder of unknown etiology. Genetic factors play an important role in the pathogenesis of these diseases, but with a complex pattern of inheritance. A number of genome-wide scans have identified several putative susceptibility loci for both CD and UC, including a locus on chromosome 12 reported in a set of British families. We aim to evaluate the linkage between CD or UC and this chromosome 12 locus in an independent set of U.S. Caucasian families (36% being of Ashkenazi Jewish origin). METHODS: Microsatellite markers along chromosome 12 spaced at approximately 10 cm intervals were used to test the putative loci in CD only families (65 sib pairs from 46 families). Regions with positive linkage for CD were then tested in a panel of UC and mixed families (44 sib pairs from 29 families). Two point linkage analysis was performed with SIBPAL. Multipoint linkage analysis was carried out with MAPMAKER/SIBS. RESULTS: We observed evidence of linkage between the region on chromosome 12 and Crohn's disease, because there was a significant excess of allele sharing in CD sib pairs (pi = 0.62, p = 0.0004 from two-point linkage; and logarithm of the odds score (LOD) = 2.0 from multipoint linkage). However, we did not observe the same linkage in UC and mixed families (p = 0.48; not significant [ns]). CONCLUSION: Our data provided further evidence that the region on chromosome 12 is likely to contain a gene predisposing to CD. PMID- 11256673 TI - Cost and effectiveness of the California triple marker prenatal screening program. AB - PURPOSE: To report the utilization of services offered and pregnancy outcomes for a unique statewide prenatal triple marker screening program and to present a cost benefit analysis. A state population of 32 million with considerable ethnic and age distribution and with a wide variety of delivery systems providing prenatal care was considered. The entire pregnant population who appeared for care before 20 weeks gestation, approximately one-half million per year during the years of 1995 to 1997, was included in the study. METHODS: Mandatory offering of serum testing, using alpha-fetoprotein from 1986 to 1995, and the addition of human chorionic gonadotropin and unconjugated estriol in 1995, with systematic follow up of serum screen positives with ultrasound and amniocentesis. This study collected and analyzed the program data and reports of outcomes and collected similar information from the birth defects registry. RESULTS: Triple marker serum screening was accepted by 67.4% of the women eligible and yielded an initial positive rate of 7.3%. More than 90% of the initially screen positive pregnancies were seen at a prenatal diagnostic center. After correction of gestational age, 71.3% had amniocentesis. The overall amniocentesis rate among women screened was 2.6%. The Program's detection rate was predicted to be 85% for neural tube defects, and, based on Monte Carlo modeling, was theoretically calculated to be 62% for Down syndrome. In practice, detection rates were 75% for neural tube defects and 41% for Down syndrome due to lower than expected amniocentesis acceptance rate. Nevertheless, at a 5% discount rate, the screening program was cost beneficial at a ratio of 2.69:1. The cost per case detected was $35,365 and per case prevented was $110,741. CONCLUSION: It is possible to implement a cost effective population-based screening in compliance with quality standards in a diverse ethnic population with a variety of health-care providers. Triple marker screening in the second trimester is a cost beneficial program even if utilization of all services is less than ideal. PMID- 11256674 TI - The Online London Dysmorphology Database. AB - A prototype of the Online London Dysmorphology Database is now available for testing purposes. In this paper we describe the new online version of the London Dysmorphology Database (LDDB), which is accessible via the World-Wide Web. The system has been developed primarily to address the information lag associated with the stand-alone version of LDDB (currently 3-15 months) and therefore provides access to the latest data from the master database that resides on a web server. An additional benefit is that, because the Online LDDB is a web-based resource, it will be available to users of both Macintosh and Unix computers as well as PC users. PMID- 11256676 TI - Harland Sanders Award Statement. PMID- 11256675 TI - Analphoid marker chromosome in a patient with hyper-IgE syndrome, autism, and mild mental retardation. AB - Hyper-IgE syndrome with recurrent infections (HIES) is a primary immunodeficiency disease characterized by recurrent skin and lung abscesses and extreme elevations of serum IgE, but also involving dentition, bones, and connective tissue. Although the etiology of HIES is unknown, autosomal dominant inheritance has been observed in multiple kindreds. A 17 year old male with sporadic HIES, autism, and mild mental retardation was found to have a supernumerary marker chromosome in peripheral blood lymphocytes and skin fibroblasts. Microdissection and FISH analysis of the marker chromosome showed that it was derived from a small interstitial deletion of one homologue of chromosome 4q21. Lack of hybridization of probes specific for telomeres and alphoid centromeres, including a centromere 4 specific probe, established that the marker was an analphoid ring chromosome. Comparative genotyping of transformed B-cell subclones with (M+) and without (M-) the marker chromosome showed loss of the maternal alleles in M- cells between markers D4S1569 and D4S3010. FISH using YAC clones from 4q21 confirmed the size and location of the interstitial deletion. Thus our patient's phenotypes were associated with de novo formation of a marker chromosome containing 15-20 cM of DNA deleted from his maternally derived chromosome 4. Proximal chromosome 4q therefore is a candidate region for disease genes for both HIES and autism. Identification of genes disrupted or lost during the formation of the marker chromosome as well as linkage studies in kindreds with HIES or autism may help us to understand the etiology of these complex phenotypes. PMID- 11256677 TI - Antenatal treatment for classic 21-hydroxylase forms of congenital adrenal hyperplasia and the issues. PMID- 11256678 TI - Minimum guidelines for the delivery of prenatal genetics services. The evaluation of clinical services subcommittee, Great Lakes Regional Genetics Group. PMID- 11256679 TI - Letter to human genetics journals. PMID- 11256680 TI - American College of Medical Genetics position statement on gene patents and accessibility of gene testing. PMID- 11256681 TI - The mitochondrial hypothesis of bipolar disorder. PMID- 11256682 TI - Neuroimaging in bipolar disorder. AB - OBJECTIVE: The authors reviewed neuroimaging studies of bipolar disorder in order to evaluate how this literature contributes to the current understanding of the neurophysiology of the illness. METHOD: Papers were reviewed as identified, using the NIMH PubMed literature search systems that reported results of neuroimaging studies involving a minimum of five bipolar disorder patients compared with healthy comparison subjects. RESULTS: Structural neuroimaging studies report mixed results for lateral and third ventriculomegaly. Recent studies suggest subcortical structural abnormalities in the striatum and amygdala, as well as the prefrontal cortex. Proton spectroscopic studies suggest that abnormalities in choline metabolism exist in bipolar disorder, particularly in the basal ganglia. Additionally, phosphorous MRS suggests that there may be abnormalities in frontal phospholipid metabolism in bipolar disorder. Functional studies have identified affective state-related changes in cerebral glucose metabolism and blood flow, particularly in the prefrontal cortex during depression, but no clear abnormalities specific to bipolar disorder have been consistently observed. CONCLUSIONS: The current literature examining the neurophysiology of bipolar disorder using neuroimaging is limited. Nonetheless, abnormalities in specific frontal-subcortical brain circuits seem likely. Additional targeted studies are needed to capitalize on this burgeoning technology to advance our understanding of the neurophysiology of bipolar disorder. PMID- 11256683 TI - The use of nimodipine in the treatment of mood disorders. AB - Nimodipine, a dihydropyridine calcium entry blocker, has been shown to protect from neuronal damage due to ischemia by providing for increased postischemic perfusion. Further, it has also been demonstrated to have antiepileptic properties. These two properties--calcium channel blockade and anticonvulsant benefits have been applied with success to mood disorder treatment. Although found helpful nearly a decade ago for uncomplicated mania, nimodipine may have particular benefits for those diagnostic subclasses of bipolar disorder most resistant to therapy, e.g., ultra-rapid-cycling bipolars and brief recurrent depressions. PMID- 11256684 TI - Serum lithium levels and the outcome of maintenance therapy of bipolar disorder. AB - A literature review was conducted to locate studies that compared different serum lithium levels in the long-term treatment of patients with bipolar disorder and articles about factors that may affect serum lithium levels. Patients with bipolar disorder on long-term treatment with lithium are typically maintained at serum lithium concentrations between 0.6 and 1.0 mEq/L. Although there are individual exceptions, serum lithium levels below 0.6 mEq/L have been shown in controlled clinical trials to be less effective in preventing relapses than levels within this range, whereas levels much above 1.2 mEq/L can lead to toxicity. Differences in efficacy between levels within the accepted range have not been established. However, higher levels are associated with greater side effects, which can lead to poor compliance. Interindividual variation in pharmacokinetics and pharmacodynamics, as well as such external factors as diet and concomitant medications, can affect serum lithium levels. PMID- 11256685 TI - Mitochondrial dysfunction in bipolar disorder. AB - Mitochondrial dysfunction is implicated in bipolar disorder based on the following lines of evidence: 1) Abnormal brain energy metabolism measured by 31P magnetic resonance spectroscopy, that is, decreased intracellular pH, decreased phosphocreatine (PCr), and enhanced response of PCr to photic stimulation. 2) Possible role of maternal inheritance in the transmission of bipolar disorder. 3) Increased levels of the 4977-bp deletion in mitochondrial DNA (mtDNA) in autopsied brains. 4) Comorbidity of affective disorders in certain types of mitochondrial disorders, such as autosomal inherited chronic progressive external ophthalmoplegia and mitochondrial diabetes mellitus with the 3243 mutation. Based on these findings, we searched for mtDNA mutations/ polymorphisms associated with bipolar disorder and found that 5178C and 10398A polymorphisms in mtDNA were risk factors for bipolar disorder. The 5178C genotype was associated with lower brain intracellular pH. mtDNA variations may play a part in the pathophysiology of bipolar disorder through alteration of intracellular calcium signaling systems. The mitochondrial dysfunction hypothesis, which comprehensively accounts for the pathophysiology of bipolar disorder, is proposed. PMID- 11256687 TI - Does olanzapine have antidepressant properties? A retrospective preliminary study. AB - OBJECTIVE: Mood-stabilizing agents are ideally conceptualized as possessing antimanic and antidepressant properties. While research on olanzapine's antimanic effects is growing, data on its possible antidepressant properties are limited. We sought to determine if olanzapine is effective in the add-on treatment of major affective disorders, particularly depressive symptoms, in a naturalistic setting. METHODS: All charts of patients meeting DSM-IV criteria for bipolar disorder or unipolar major depressive disorder treated with olanzapine in a private psychiatric practice were reviewed and clinical response was assessed retrospectively using the Clinical Global Impression Scale for Improvement (CGI I). RESULTS: Olanzapine was moderately effective in 6/10 (60%) patients. Side effects were present in 8/10 (80%), most commonly weight gain. CONCLUSIONS: Olanzapine appears to be moderately effective in open add-on treatment in patients with mainly depressive symptoms. Accumulating evidence suggests that olanzapine, and atypical antipsychotics in general, possess mild to moderate adjunctive antidepressant properties. PMID- 11256686 TI - Lifetime prevalence of substance or alcohol abuse and dependence among subjects with bipolar I and II disorders in a voluntary registry. AB - OBJECTIVE: To evaluate the prevalence of substance abuse dependence and/or alcohol abuse dependence among subjects with bipolar I versus bipolar II disorder in a voluntary registry. METHOD: One hundred randomly selected registrants in a voluntary case registry for bipolar disorder were interviewed, using the Structured Clinical Interview for DSM-IV Axis I Disorders, to validate the diagnosis of this registry. Corroborative information was obtained from medical records, family members and the treating psychiatrist. Eighty-nine adults (18-65 years) met criteria for bipolar disorder (bipolar I = 71, bipolar II = 18) and were included in this analysis. RESULTS: Forty-one (57.8%) subjects with bipolar I disorder abused, or were dependent on one or more substances or alcohol, 28.2% abused, or were dependent on, two substances or alcohol, and 11.3% abused or were dependent on three or more substances or alcohol. Nearly 39% of bipolar II subjects abused or were dependent on one or more substances, nearly 17% were dependent on two or more substances or alcohol, and 11% were dependent on three or more substances or alcohol. Alcohol was the most commonly abused drug among either bipolar I or II subjects. CONCLUSIONS: Consistent with other epidemiologic and hospital population studies, this voluntary bipolar disorder registry suggests a high prevalence of comorbidity with alcohol and/or substance abuse dependence. Bipolar I subjects appear to have higher rates of these comorbid conditions than bipolar II subjects; however, as the number of bipolar II subjects was rather small, this suggestion needs confirmation. PMID- 11256688 TI - Li+/Mg2+ competition at therapeutic intracellular Li+ levels in human neuroblastoma SH-SY5Y cells. AB - OBJECTIVES: One proposed mechanism of lithium action in the treatment of bipolar disorder is that Li+ competes with Mg2+ for Mg2+ binding sites within the cell. In this study, we investigated this competition at therapeutic intracellular Li+ levels in human neuroblastoma SH-SY5Y cells. METHODS: We used fluorescence spectroscopy and a Mg2+ indicator, furaptra, to investigate this competition in human neuroblastoma SH-SY5Y cells. Atomic absorption spectrophotometry was used for determination of the intracellular Li+ levels. RESULTS: The neuroblastoma cells, incubated in 15 mM or 30 mM Li+-containing buffer, showed a significant increase in free intracellular Mg2+ levels [using a positive linear within-groups contrast t-test, the 15 mM condition produced t(2) = 5.0, one-tailed p < 0.02, and the 30 mM Li+-incubation conditions gave t(2) = 9.2, one-tailed p < 0.006] but did not significantly increase over time in the Li+-free condition [t(2) = 0.1, one-tailed p > 0.96]. At the earlier times during the incubation (1 or 10 min for the 15 mM or 30 mM Li+-containing buffers), the intracellular Li+ concentrations were 0.6-2.5 mM, values which are comparable to those reached in the brain of Li+-treated patients. CONCLUSION: We demonstrated that competition between Li+ and Mg2+ can occur at therapeutic intracellular Li+ levels. PMID- 11256690 TI - A new stilbene dimer--shegansu B from Belamcanda chinensis. AB - A new dimeric stilbene, named shegansu B (1) was isolated from the ethanolic extract of the roots of Belamtantida chinensis (L.) DC., along with the known compounds isorhapontigenin, resveratol, p-hydroxybenzoic acid, iridin, tectoridin, tectorigenin and daucosterol. The structures were elucidated by means of spectroscopic evidence including 2D-NMR studies. PMID- 11256689 TI - Structures of two diterpenoid dimers from bulbs of Fritillaria ebeiensis. AB - A new ent-kaurane diterpenoid dimer, fritillebinide C(1) together with one known diterpenoid dimer fritillebinide B (2) were isolated from the bulbs of Fritillaria ebeiensis G.D. Yu et G.Q. Ji. Compound 1 has been determined to be ent-3beta-acetoxy-kauran-16beta,17-acetal ent-16beta-kauran-17(S)-aldehyde(1) by means of spectral analysis and chemical evidence. PMID- 11256691 TI - Colebroside A, a new diglucoside of fatty acid ester of glycerin from Clerodendrum colebrookianum. AB - Colebroside A (1), a new diglucoside of fatty acid ester of glycerin, was isolated from the aerial parts of Clerodendrum colebrookianum Walp., along with nine known compounds (2-10). Their structures were elucidated by spectroscopic and chemical methods. Compounds 2, 3, 4, 5, 7, 8, 9 and 10 have been obtained from this plant for the first time. PMID- 11256692 TI - Three new triterpenoid saponins from the seeds of Vaccaria segetalis. AB - Three new triterpenoid saponins, named segetoside G(1), H(2) and I(3), have been isolated from the seeds of Vaccaria segetalis. On the basis of chemical reaction and spectral data, their structures have been established as: 28-O-beta-D xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-arabinofuranosyl (1-->3)]-beta-D-(4-O-acetyl)-fucopyranosyl-gypsogenin-3-O-beta-D-galactopyranosyl (1-->2)-beta-D-(6-O-butyl ester)-glucuronopyranoside (1), 28-O-beta-D xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-(5-O-acetyl) arabinofuranosyl-(1-->3)]-beta-D-(4-O-acetyl)-fucopyranosyl-gypsogenin-3-O-beta-D galactopyranosyl-(1-->2)-beta-D-glucuronopyranoside (2) and 28-O-beta-D xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-(5-O-acetyl) arabinofuranosyl-(1-->3)]-beta-D-(4-O-acetyl)-fucopyranosyl-quillaic acid-3-O beta-D-galactopyranosyl-(1-->2)-beta-D-glucuronopyranoside (3). PMID- 11256693 TI - Counter effects of higenamine and coryneine, components of aconite root, on acetylcholine release from motor nerve terminal in mice. AB - The counter effects of higenamine and coryneine, components of aconite root, on acetylcholine (ACh) release from motor nerve terminals in the mouse phrenic nerve diaphragm muscle preparation were studied by a radioisotope method. Both nerve evoked release and spontaneous release of [3H]-ACh from the preparation preloaded with [3H]-choline were measured. The change in the tetanic tension of muscle was simultaneously recorded in the same preparation. Higenamine (10 microM) augmented both the nerve-evoked and spontaneous ACh releases, and the muscle tension. The effects were inhibited by pretreatment with propranolol (10 microM), a beta adrenoceptor antagonist. Coryneine reduced the nerve-evoked release of ACh, accelerated the decay of tetanic tension (tetanic fade) at 30 microM, and it depressed the peak amplitude of tetanic tension at a higher concentration of 100 microM. These results suggest that of the two components contained in aconite root, higenamine increases ACh release via activation of beta-adrenoceptor, and conversely coryneine depresses ACh release by preferentially acting at motor nerve terminal. PMID- 11256694 TI - Hydrolyzable tannins and related polyphenols from Eucalyptus globulus. AB - Eucaglobulin (1), a new complex of gallotannin and monoterpene, was isolated from the leaves of Eucaloptus globulus. Its structure was elucidated on the basis of spectral data. Four known hydrolyzable tannins [tellimagrandin I (2), eucalbanin C (3), 2-O-digalloyl-1,3,4-tri-O-galloyl-beta-D-glucose (4), 6-O-digalloyl-1,2,3 tri-O-galloyl-beta-D-glucose (5)], as well as gallic acid (6) and (+)-catechin (7), were also isolated. The antibacterial effects of some of these compounds were examined. PMID- 11256695 TI - Structural elucidations of two ent-kaurane dimers from bulbs of Fritillaria ebeiensis var. purpurea. AB - A novel ent-kaurane diterpenoid dimer, fritillebinide B (1) together with one known diterpenoid dimer fritillebinide A (2) were isolated from the bulbs of Fritillaria eheiensis var. purpurea G.D. Yu et P. Li. Compound 1 has been established to be ent-3beta-acetoxy-kauran-16beta,17-acetal ent-16beta-kauran 17(R)-aldehyde (1) by means of spectral analysis and chemical evidence. PMID- 11256696 TI - A new isoflavone glucoside from Pterocarpus santalinus. AB - A new isoflavone glucoside (1) together with the known santal has been isolated from the heartwood of Pterocarpus santalinus. Based on spectral methods, the structure of the new compound was elucidated as 4',5-dihydroxy 7-O-methyl isoflavone 3'-O-beta-D-glucoside. PMID- 11256698 TI - Isolation and structure elucidation of novel gamma-lactones from Saccopetalum prolificum. AB - Two novel gamma-lactones, saccopetrin C(1) and saccopetrin D (2), were isolated from the roots of Saccopetalum prolificum. Their structures were established by spectroscopic and chemical methods. PMID- 11256697 TI - A new phytoecdysone from the roots of Rhaponticum uniflorum. AB - A new ecdysteroid named rhapontisterone R1 (1) together with two known phytoecdysones, rhapontisterone (2) and ecdysterone (3) were isolated from the roots of Rhaponticum uniflorum (L.) DC. The new compound was shown to be 2beta,3beta,11alpha,14alpha,20xi,22xi-hexahydroxy-stigma-7,24(28)-dien-6-oxo 28,25-carbolactone. The structure has been determined primarily on the basis of physico-chemical properties and spectral analysis. PMID- 11256699 TI - Rearranged abietane diterpenoids from Clerodendrum mandarinorum. AB - Five new abietane derivatives which have a commonly rearranged abietane skeleton contained a 17(15-->16),18(4-->3)-diabeo-abietane framework, mandarones D-H, were isolated from the stem of Clerodendrum nmantarinorum Diels (Verbenaceae). The structures were characterized as (16S)-12,16-epoxy-11-hydroxy-17(15-->16), 18(4- >3)-diabeo-abieta-3,5,8,11,13-pentaene-7-one (mandarone D, 1), 12,16-epoxy-11,14 dihydroxy-17(15-->16),18(4-->3)-diabeo-abieta-3,5,8,11, 13,15-hexaene-7-one (mandarone E, 2), 12,16-epoxy-6,11,14-trihydroxy-17(15-->16),18(4-->3)-diabeo abieta-3,5,8,11,13,15-hexaene-7-one (mandarone F, 3),12,16-epoxy-11,14-dihydroxy 6-methoxy-17(15-->16),18(4-->3)-diabeo-abieta-3,5,8,11,13,15-hexaene-2,7-dione (mandarone G, 4) and 12,16-epoxy-11,14-dihydroxy-17(15-->16),18(4-->3)-diabeo abieta-3,5,8,11,13,15-hexaene-1,7-dione (mandarone H, 5) respectively, mainly based on the spectral analysis and by comparison with those of closely related compounds. PMID- 11256700 TI - Monitoring hazardous waste sites: characterization and remediation considerations. PMID- 11256701 TI - Chemical composition of individual aerosol particles from working areas in a nickel refinery. AB - Individual aerosol particles (n = 1170) collected at work stations in a nickel refinery were analyzed by wavelength-dispersive electron-probe microanalysis. By placing arbitrary restrictions on the contents of sulfur and silicon, the particles could be divided into four main groups. Scanning electron images indicated that most of the particles examined were relatively small (< or = 2 microm, equivalent projected area diameter), and that their morphology suggested formation from a melt. There was an absence of well-defined phases and simple stoichiometries, indicating that exposures to pure substances such as nickel subsulfide or specific oxides appeared not to occur. Although the elemental composition of particles varied greatly, a rough association was evident with the known elemental content of the refinery intermediates. The implications of the findings for aerosol speciation measurements, toxicological studies and interpretation of adverse health effects are explored. PMID- 11256702 TI - Determination of n-alkanes and polycyclic aromatic hydrocarbons in atmospheric particulate and vapour phases in Oviedo, Spain, by GC-MS. AB - A GC-MS procedure for the determination of hydrocarbons in air samples from Oviedo, Spain, was developed. Air hydrocarbons were sampled with a high volume sampler equipped with a holder containing a glass fiber filter, to trap the particulate phase, and two polyurethane foams to capture hydrocarbons of the vapour phase. Compounds were extracted with CH2Cl2 by Soxhlet extraction and then fractionated using column chromatography with alumina silica. Analyses of the fractions were performed by GC-MS in the electron ionization mode. PAHs and n alkanes were the compounds examined in this work. Samples collected in the vicinity of the Faculty of Chemistry (a semi-urban area) were analysed. The total concentration of PAHs in the air samples analysed ranged from 28 to 76 ng m(-3). The total concentration of n-alkanes and PAHs in the vapour phase exceeded the concentration in the particulate phase in the samples analysed. PMID- 11256703 TI - Analysis of polycyclic aromatic hydrocarbons in the atmospheric particulate: focused microwaves for a faster extraction method. AB - In this paper, improvements obtained by using focused microwaves for extraction, in the analysis of polycyclic aromatic hydrocarbons (PAHs) adsorbed on particulate matter, are discussed. The method was tested on the following PAHs, which are considered to be among the most harmful with regard to carcinogenicity: benzo[a]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, indeno[1,2,3-cd]pyrene, dibenzo[a,h]anthracene. The extraction of PAHs and concentration of the sample can be performed in 3 h with a recovery of at least 70% and a maximum standard deviation of 4%. These steps are followed by clean-up on a SPE (solid-phase extraction) cartridge and analysis by GC-MS. Real samples collected in the urban area of Bari were analysed according to the proposed procedure. PMID- 11256704 TI - Application of a modified BCR sequential extraction (three-step) procedure for the determination of extractable trace metal contents in a sewage sludge amended soil reference material (CRM 483), complemented by a three-year stability study of acetic acid and EDTA extractable metal content. AB - This paper provides additional data on a sewage sludge amended soil certified reference material, CRM 483, which was certified in 1997 for its EDTA and acetic acid extractable contents of some trace metals, following standardised extraction procedures. The additional work aimed to test the long-term stability of the material and the applicability of an improved version of the BCR three-step sequential extraction procedure on the sewage sludge amended soil (CRM 483). The paper demonstrates the CRM 483 long-term stability for EDTA and acetic acid extractable contents of Cd, Cr, Cu, Ni, Pb and Zn and gives the results (obtained in the framework of an interlaboratory study) for the extractable contents of the same elements in the CRM 483, following the BCR three-step sequential extraction scheme. The aqua regia extractable contents following the ISO 11466 Standard are also given. The data are given as indicative (not certified) values. PMID- 11256705 TI - Column leaching and sorption experiments to assess the mobility of potentially toxic elements in industrially contaminated land. AB - Made-up ground collected from layers of a trial pit excavated on a former industrial site was treated with artificial rainwater in a series of column leaching and sorption experiments. Metal mobility and the ability of various layers of material obtained from the pit to act as sources or sinks of potentially toxic elements were assessed. Samples from different layers varied in their abilities to raise the pH of rainwater applied at pH 3.5 and 4.3, and this was reflected in the amounts of metals mobilised by the rainwater as it percolated through the soil column. Material from the top two layers of the pit released cadmium, copper, manganese, lead, nickel and zinc to the aqueous phase, but the lower layers, with higher buffering capacity, were able to resist acidification even when the equivalent of 12 months' rainfall (western UK) was applied. Column sorption experiments confirmed the ability of material from layer 4 (48-50 cm) to take up copper, manganese and zinc. Metals were determined in the leachates by flame and electrothermal atomic absorption spectrometry and principle anions by ion chromatography. PMID- 11256706 TI - Triethoxysilyl-substituted aminoethanethiol ligands for zinc and cadmium complexes and aminoethanethiol-modified silica gel. Evaluation of the corresponding supported molecular trap for metallic pollutant uptake (Cd2+, Hg2+ and Pb2+). AB - The reaction of 2-[3-(triethoxysilyl)propylamino]ethanethiol (LH, a) and 1-methyl 2-[3-(triethoxysilyl)propylamino]ethanethiol (LH, b) with ZnX2 and CdX2 (X = Cl, Br, I, NO3) in tetrahydrofuran (THF) or CH2Cl2 gives several complexes depending on the experimental conditions. Elemental analyses, IR, Raman, 13C[1H], 1H NMR and mass spectroscopies indicated the formation of mononuclear and dinuclear complexes. In the absence of NEt3 as proton quencher, the protonated ligands react in their zwitterionic form giving dinuclear [M(LH)X2]2 [M = Zn (1), Cd (2); LH = a, b; X = Cl, Br, I] or mononuclear M(NO3)2(LH)2 [M = Zn (5), Cd (6); LH = a] complexes. In both cases, coordination occurs through the S atoms, the ligands acting as terminal and bridging species. With NEt3, the deprotonated ligands are chelated through their N and S atoms and bridging occurs through the S atoms in [MLX]2 [M = Zn (3), Cd (4); LH = a; X = Cl, Br] complexes. The LH ligand is chemically grafted onto silica, the procedure optimized and the resulting material characterized by 13C and 29Si cross-polarization, magic-angle spinning (CP-MAS) NMR and DRIFT. This material is evaluated as a supported molecular trap for binding heavy metals (Cd2+, Hg2+, Pb2+) in aqueous solution. In both batch and column processes, it appears that Hg2+ and Pb2+ are trapped more than Cd2+, but in all cases values lower than those allowed were obtained. PMID- 11256707 TI - Avoiding pitfalls in the determination of halocarboxylic acids: the photochemistry of methylation. AB - Haloethanoic (haloacetic) acids are formed during chlorination of drinking water and are regulated by the Environmental Protection Agency (EPA). These compounds are normally quantified by gas chromatography with electron capture detection (GC ECD) as the methyl esters. EPA Method 552 uses diazomethane (CH2N2) for this purpose, but has only been validated by EPA for HAA6: chloro-, dichloro-, bromo-, dibromo-, bromochloro- and trichloroacetic acids. EPA Method 552.2 was developed and validated for all nine analytes (HAA9 = HAA6 + dibromochloro-, bromodichloro- and tribromoethanoic acids). Since the promulgation of Method 552.2, which uses acidic methanol, a debate has ensued over discrepancies observed by various laboratories when using diazomethane instead. In an effort to identify and eliminate potential sources for these discrepancies, a comparative study was undertaken for HAA9. Better accuracy and precision were observed for all HAA9 species by Method 552.2; recoveries were satisfactory in de-ionized and tap water. Method 552 remains satisfactory for HAA6. Systematic differences in instrumental response are observed for the two methods, but these are precise and may be accounted for using similarly treated standards and analyte-fortified (spiked) samples. That notwithstanding, Method 552 (CH2N2) was shown to be unsuitable for dibromochloro-, bromodichloro- and tribromoethanoic acids (HAA9 6). The primary problem appears to be a photoactivated reaction between diazomethane and the HAA9-6 analytes; however, side reactions were found to occur even in the dark. Analyte loss is most pronounced under typical laboratory lighting (white F40 fluorescent lamps + sunlight), but it is also observed under Philips gold F40 lamps (lambda > or = 520 nm), and in the dark. PMID- 11256708 TI - Ascorbic acid reduction of residual active chlorine in potable water prior to halocarboxylate determination. AB - In studies on the formation of disinfection byproducts (DBPs), it is necessary to scavenge residual active (oxidizing) chlorine in order to fix the chlorination byproducts (such as haloethanoates) at a point in time. Such research projects often have distinct needs from requirements for regulatory compliance monitoring. Thus, methods designed for compliance monitoring are not always directly applicable, but must be adapted. This research describes an adaptation of EPA Method 552 in which ascorbic acid treatment is shown to be a satisfactory means for reducing residual oxidizing chlorine, i.e., HOCl, ClO-, and Cl2, prior to determining concentrations of halocarboxylates. Ascorbic acid rapidly reduces oxidizing chlorine compounds, and it has the advantage of producing inorganic halides and dehydroascorbic acid as opposed to halogenated organic molecules as byproducts. In deionized water and a sample of chlorinated tap water, systematic biases relative to strict adherence to Method 552 were precise and could be corrected for using similarly treated standards and analyte-fortified (spiked) samples. This was demonstrated for the quantitation of chloroethanoate, bromoethanoate, 2,2-dichloropropanoate (dalapon), trichloroethanoate, bromochloroethanoate, and bromodichlorocthanoate when extracted, as the acids, into tert-butyl methyl ether (MTBE) and esterified with diazomethane prior to gas chromatography with electron capture detection (GC-ECD). Recoveries for chloroethanoate, bromoethanoate, dalapon, dichloroethanoate, trichloroethanoate, bromochloroethanoate, bromodichloroethanoate, dibromoethanoate, and 2 bromopropanoate at concentrations near the lower limit of detection were acceptable. Ascorbic acid reduction appears to be the best option presently available when there is a need to quench residual oxidants fast in a DBP formation study without generating other halospecies but must be implemented cautiously to ensure no untoward interactions in the matrix. PMID- 11256709 TI - Adsorption characteristics of a phenoxy acetic acid herbicide on activated carbon. AB - The adsorption of 2,4-dichlorophenoxyacetic acid (2,4-D) by two powdered coal activated carbons was studied in aqueous solution. The modelling of the adsorption equilibrium showed that the adsorption of 2,4-D fitted a Langmuir isotherm. Adsorption was influenced by the activated carbon type, adsorbent concentration and solution characteristics. The adsorption was found to decrease with an increase in pH over the range 1.5-9. Maximum adsorption occurred at pH approximately 2.5, which corresponds to the 2,4-D pKa value. The amount of 2,4-D adsorbed was also found to depend on the NaCl concentration. PMID- 11256710 TI - Detecting and characterising sources of persistent organic pollutants (PAHs and PCBs) in surface sediments of an industrialized area (harbour of Trieste, northern Adriatic Sea). AB - A sediment sampling based on a two-dimensional mapping was performed in the harbour of Trieste (northern Adriatic Sea), considering 28 sites exposed to pollutant inputs from harbour and industrial activities. Polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) were determined in surface sediments, because these very persistent pollutants seem to be responsible for the depletion of benthic populations observed in this area. The correlation matrix indicates that PAHs and PCBs are non-correlated, and probably have different sources. Both cluster analysis performed on the sampling sites and graphical drawing of the PAH sediment contents make it possible to locate along the shoreline a band of more polluted sediments, clustered around a site facing a steelmaking factory, to be considered as the main source point for PAHs. The evaluation of phenanthrene to anthracene (P/AN) and fluoranthene to pyrene (FL/PY) ratios permits the assessment of the pyrolytic, industrial origin of these PAHs, rejecting a second possible source of hydrocarbons (i.e., an oil pipeline terminal, situated near the steelmaking factory). Graphical drawing of the total PCB iso-concentrations reveals a different source-point for this other category of very persistent pollutants. PMID- 11256711 TI - Traceability from weathered oil spills in the marine environment to the original contamination source. A case study. AB - An algorithm for stepwise approximation to the most reliable source of contamination is proposed. The concept of the algorithm is based on discrimination rather than on similarity. Gradually, by means of more complicated and higher distinguishing power approaches, the differences between the spilled sample and the suspected source are checked. Capillary gas chromatography with flame-ionization and mass spectrometric detection was applied to gather the necessary data. Fingerprint, diagnostic parameter approaches and pattern recognition methods were used for reliably distinguishing the analytical results. If the compared samples are not discriminated by any means, they are considered as identical. PMID- 11256712 TI - Study of the relationships between bone lead levels and its variation with time and the cumulative blood lead index, in a repeated bone lead survey. AB - The study aims were to: (i) investigate long term human lead metabolism by measuring the change of lead concentration in the tibia and calcaneus; and (ii) assess whether improved industrial hygiene was resulting in a slow accumulation of lead in an exposed workforce. 109Cd excited K X-ray fluorescence was used to measure tibia lead and calcaneus lead concentrations in 101 workers in a secondary lead smelter. 51 subjects had had similar bone lead measurements 5 years previously. Most of the other subjects had been hired since the first survey. Measurements of whole blood lead were available for the large majority of subjects. Tibia lead concentrations fell significantly (p<0.001) in the 51 subjects with repeated bone lead measurements, from a mean of 39 microg Pb (g bone mineral)(-1) to 33 microg Pb (g bone mineral)(-1). The change correlated negatively with the initial tibia lead concentration, producing an estimate for an overall half-life of 15 years, with a 95% confidence interval of 9 to 55 years. Adding continuing lead exposure and recirculation of bone lead stores to the regression models produced half-life estimates of 12 and 9 years, respectively, for release of lead from the tibia. The repeat subjects showed no net change in calcaneus lead (64 microg Pb (g bone mineral)(-1) initially, 65 microg Pb (g bone mineral)(-1) 5 years later). Subjects not measured previously had average lead concentrations of 15 microg Pb (g bone mincral)(-1) in the tibia and 13 microg Pb (g bone mineral)(-1) in the calcaneus. The rate of clearance of lead from the tibia (9 to 15 years) is towards the more rapid end of previous estimates. The lack of a significant fall in the calcaneus lead was surprising. Attempts should be made to repeat this observation. If confirmed, it would have implications for models of lead metabolism. The relatively low lead concentrations in the non-repeat subjects are reassuring. However, observation after a longer period of employment would be desirable. PMID- 11256713 TI - A gas chromatography-quartz crystal microbalance for speciation of sulfur compounds in landfill gas. AB - A new method, based on separation with gas chromatography and detection with a quartz crystal microbalance, was used for the quantification of ethane-1-thiol, propane-1-thiol, ethyl methyl sulfide, propane-2-thiol, butane-2-thiol and butane 1-thiol. The analytical error, the analysis time and the general analytical performance are identical for gas chromatography-mass spectrometry (GC-MS) and gas chromatography-quartz crystal microbalance (GC-QCM). However, the GC-QCM method constitutes a low cost alternative to GC-MS for monitoring individual sulfur compounds in landfill gas. It is an easily assembled apparatus made with an injection device, a furnace, a column, a crystal cell and a quartz crystal. PMID- 11256714 TI - Analyst in the sky: satellite-based remote sensing. PMID- 11256715 TI - Can NRDAs take the environment back to the future? PMID- 11256716 TI - Does the mere presence of a pesticide residue in food indicate a risk? PMID- 11256717 TI - A checklist for critiquing treatment fidelity studies. AB - My assignment was to critique and integrate the previous papers. I have organized this paper as follows: (1) a description and rationale for a checklist which can be used in judging the quality of a fidelity measure; (2) a critique of the Schaedle and Epstein paper; (4) a critique of the Lucca paper; and (4) a review of the paper by Bond, Evans, Salyers, Williams, and Kim. PMID- 11256718 TI - Medicaid and African American outpatient mental health treatment. AB - The present study tested the hypothesis that Medicaid-financed African Americans would be more likely to receive outpatient mental health treatment than African Americans whose treatment was financed by private insurance. The hypothesis was confirmed: when compared with privately insured persons eligible for care under either fee-for-service or managed care, the Black-White gap in outpatient service use was significantly smaller under Medicaid. There was no racial difference in outpatient treatment rates among the uninsured. The often-noted difference between Blacks and Whites in the likelihood of receiving outpatient mental health treatment is confined largely to the privately insured. PMID- 11256720 TI - A clubhouse fidelity index: preliminary reliability and validity results. AB - Although core clubhouse components are well known, no instruments currently exist to measure clubhouse fidelity. In the current study, a short index was developed to assess program implementation of clubhouse components. Presence or absence of 15 objective clubhouse features was tabulated across 22 of 24 psychiatric rehabilitation programs throughout one New England state. Results supported the internal consistency, criterion-related validity, and convergent validity of this instrument. The clubhouse index was able accurately to differentiate programs at three levels of clubhouse fidelity, and results further indicated that programs with higher clubhouse fidelity scores also adhered more strongly to underlying principles of rehabilitation practice. These findings suggest the clubhouse index holds promise as a fidelity tool for researchers and administrators. PMID- 11256719 TI - Measurement of fidelity in psychiatric rehabilitation. AB - Until recently, most psychiatric rehabilitation models have been poorly defined and few have had systematic methods for measuring their implementation. We review the historical roots for the development of fidelity measures and describe recent applications in both research and practice. PMID- 11256721 TI - Specifying intensive case management: a multiple perspective approach. AB - "Intensive case management" (ICM) programs for people with serious mental illness are found widely throughout the United States. However, there is no standard definition or conceptualization of ICM. Despite these differences, ICM aspires to a set of common principles and core functions derived from the concept of continuity of care. This study attempted to identify the elements of ICM program theory by integrating information from the ICM literature with survey and focus group data reflecting the perspectives of three distinct ICM respondent groups (researchers/administrators, program managers, and case managers). The findings suggest a strong consensus about the structural dimensions of ICM, but a moderate consensus about their operationalization. More generally, the results support viewing ICM as more "client oriented," in contrast with conventional case management programs that are more "system driven." PMID- 11256722 TI - Correlates of adolescents' satisfaction with mental health services. AB - While there has been increased attention to consumers' satisfaction with mental health services as an indicator of quality of care, little is known about the construct of consumer satisfaction, especially for youth. The goal of this study was to examine potential correlates of adolescents' satisfaction with mental health services. One hundred eighty adolescents who had received out-patient mental health services completed a multidimensional satisfaction scale and measures of behavior problems, attitudes and expectations about treatment, perceived choice/motivation in seeking treatment, and service use history. Results indicate that the strongest unique correlates of satisfaction are severity of behavior problems, positive expectations about services, and perceived choice in seeking services. Satisfaction with services was also associated with service site, length of time in treatment, and reason for entering treatment. Demographic variables were not related to satisfaction. A discussion of the appropriateness of using satisfaction as an indicator of quality of care is included. PMID- 11256723 TI - Youths' access to mental health services: the role of providers' training, resource connectivity, and assessment of need. AB - This paper posits that providers with training in and knowledge of mental health resources are more likely to recognize youths' mental health problems, and provide youths with services. In 1994 and 1996, we interviewed 792 adolescents who were involved with St. Louis public health, juvenile justice, child welfare. or education service sectors. Two hundred eighty-two youths had received some services, listing 533 providers. We could identify 364 of those providers, and 61% (222) responded concerning service need, service use, and provider knowledge and behavior. Structural equation models demonstrate that provider assessment of youths' mental health problems is the largest and provider knowledge of service resources the second largest determinant of service provision. Youths' self reported mental health is not positively associated with increased services and is only minimally associated with provider assessment of their problems. Training (both professional and inservice) contributes to higher assessments of youths' problems and greater resource knowledge, which is associated with increased service provision. Providers from the mental health and child welfare sectors have more professional training in mental health and are more likely to receive inservice training. Inservice training should be offered to all who work with youths. PMID- 11256724 TI - Psychiatric hospitalizations, arrests, emergency room visits, and homelessness of clients with serious and persistent mental illness: findings from a randomized trial of two ACT programs vs. usual care. AB - OBJECTIVE: This investigation examined several adverse outcomes in clients with serious mental illness in a randomized trial of Assertive Community Treatment (ACT) versus usual care. METHOD: 163 subjects were randomized to one of two ACT experimental conditions (staffed by consumers or non-consumers) or usual community care. Conditions were compared on psychiatric hospitalization, emergency room visit, arrest, and homelessness, within the two-year study period. Demographic, program, and client variables were examined for significant associations with outcomes. RESULTS: Significant differences were found between ACT and usual care in time to first arrest, but not hospitalization, homelessness or ER visits. Shorter time to first hospitalization was associated with male gender, diagnoses other than schizophrenia, high psychiatric symptomatology and lower provider case load. ER visits were associated with increased client symptomatology. Shorter times to homelessness were predicted by poorer therapeutic alliance between case manager and clients. Shorter time to first arrest was predicted by client minority status and enrollment in usual care. CONCLUSIONS: The paucity of significant main effects may have been due to a prolonged "start-up" phase of the ACT programs, poor ACT implementation, restricted availability of psychiatric hospital beds, or changes in usual care services delivered over the study period. PMID- 11256725 TI - The impact of federal systems integration initiatives on services for mentally ill homeless persons. AB - Nearly everyone writing on homelessness over the past 15 years has called for comprehensive integrated systems of care to address the multiple and complex needs of people who become homeless, especially those with mental illness. What is often overlooked is that calls for systems integration are far from new. Although the names have changed over the years, the underlying concepts have not. The purposes of this paper are fourfold: (1) to clarify the distinction between services integration and systems integration; (2) to map the evolution of federal programs to demonstrate that most of these really have been focused on services integration rather than systems integration; (3) to assess the extent that data from these programs supports the idea of systems integration; and (4) to show how the current ACCESS demonstration for persons who are homeless and mentally ill is likely to provide answers that prior programs have not. Without these new data, systems integration, as one solution to the problems of homelessness, remains a theory without empirical evidence; albeit a theory with persuasive conceptual underpinnings. PMID- 11256726 TI - Patient, provider, and treatment factors associated with poor-quality care for schizophrenia. AB - Interventions are needed to improve the quality of care for schizophrenia. However, in designing these interventions it would be helpful to understand better which patients are at highest risk for poor-quality care and why care for this disorder is often of poor quality. We study the extent to which patient and treatment factors are associated with poor-quality care in 224 patients randomly sampled from two mental health clinics. Quality of medication management is evaluated using an established method based on national treatment recommendations. Multivariate regression is used to study the effect of patient and treatment factors on treatment quality, controlling for provider. Risk for poor-quality care was greater for patients who were more severely ill, older, and less compliant with treatment recommendations. There were trends toward poor management of symptoms in men and substance abusers, and poor management of side effects in Black patients. Provision of poor-quality care was associated with failure to document symptoms and side effects in the medical record. Interventions to improve care for schizophrenia should attend to the need for accurate clinical assessment and strategies for managing challenging clinical situations. PMID- 11256728 TI - Quality of care in services to family members of people with serious mental illnesses. AB - This paper discusses the current status, importance, and future directions of quality of care work regarding support and information services for family members of people with serious mental illnesses. In reviewing existing literature, it highlights the need for research that documents the services currently received by family members in more depth and detail, the importance of including family-member services in quality-of-care standards and evaluations, and the necessity of grappling with fundamental questions such as who defines "quality" and "optimal" care, whose outcomes are foregrounded in such inquiry, and the development of methodologies to advance this area of inquiry. PMID- 11256727 TI - Quality of public sector care for schizophrenia in Arkansas. AB - OBJECTIVE: Using process-of-care indicators, we examined the quality of care provided to 139 individuals receiving treatment for schizophrenia in public sector systems. METHODS: Longitudinal data on services use and medication management were abstracted from medical records. Medication adherence data were obtained by self- and informant reports. RESULTS: Overall, 39% of participants had less than monthly contact with community-based service (CBS) providers. When participants in day treatment or partial hospitalization programs were excluded, less than monthly CBS contact increased to 70%. Of participants, 40%-60% were prescribed medications outside guideline-recommended dose ranges. Up to half of participants reported taking half or less of prescribed antipsychotics. The adverse impact on patient outcomes of these practice patterns is well established. CONCLUSIONS: Public sector organizations face powerful challenges to the behavioral changes needed to sustain best practice care. Overcoming these challenges to assure high-quality care for schizophrenia will require tremendous creativity and commitment. PMID- 11256729 TI - History and evidence-based medicine: lessons from the history of somatic treatments from the 1900s to the 1950s. AB - This paper examines the early history of biological treatments for severe mental illness. Focusing on the period of the 1900s to the 1950s, I assess the everyday use of somatic therapies and the science that justified these practices. My assessment is based upon patient records from state hospitals and the contemporaneous scientific literature. I analyze the following somatic interventions: hydrotherapy, sterilization, malaria fever therapy, shock therapies, and lobotomy. Though these treatments were introduced before the method of randomized controlled trials, they were based upon legitimate contemporary science (two were Nobel Prize-winning interventions). Furthermore, the physicians who used these interventions believed that they effectively treated their psychiatric patients. This history illustrates that what determines acceptable science and clinical practice was and, most likely will, continue to be dependent upon time and place. I conclude with how this history sheds light on present-day, evidence-based medicine. PMID- 11256731 TI - Surveillance for incident HIV infection: new technology and new opportunities. AB - Although surveillance for HIV infection has traditionally focused on the incidence of AIDS and the prevalence of HIV, new diagnostic technologies that allow the estimation of incident HIV infection have become available. Number and distribution of new cases of HIV infection, rather than old cases, are the data most relevant to guide rational application of HIV prevention programs. Historically, incident HIV infection has been measured in longitudinal cohort studies, diagnosed clinically or since 1993 by detection of seroconverting patients (during the window period before appearance of HIV antibody) who are viremic as measured by p24 antigen or RNA-PCR. The sensitive-less sensitive EIA test (or serologic testing algorithm for recent HIV seroconversion [STAHRS]) has now made the serologic diagnosis of incident HIV infection in individual patients as well as the estimation of HIV incidence in populations possible. Examples of the public health application of this are studies of HIV incidence in anonymous test site attendees, sexually transmitted disease clinic patients, and in treatment injection drug users in San Francisco. These sorts of studies allow us not only to measure incidence cross-sectionally but also facilitate surveillance for HIV subtypes and primary antiretroviral resistance, targeting early antiretroviral therapy and partner notification, and understanding who is "failing" prevention. Having an HIV surveillance system that focuses on incident rather than prevalent infection should be our long-term goal. PMID- 11256730 TI - Effective detection of HIV. AB - Biomedical advances, new HIV testing technologies, and policy shifts in the last 15 years have created substantial new challenges and opportunities for service providers, policy makers, and researchers regarding broad scale identification of HIV-seropositive persons. Effective HIV testing will be achieved when we: (1) increase the number of high-risk persons tested; (2) decrease the time from HIV infection to detection; (3) increase testing acceptability; (4) increase the proportion of individuals tested who receive their results; and (5) increase the proportion of individuals tested seropositive who are linked to care. Strategies to enhance effectiveness include implementing new testing technologies and delivery modalities; expanding access to client-controlled testing; targeting providers' knowledge, attitudes, and behaviors regarding HIV testing; mainstreaming HIV testing as routine clinical care; targeting persons who engage in high-risk behaviors and those in high-risk groups; and implementing a national behavioral surveillance system. Addressing these challenges will improve HIV detection in the United States, which is vital to both HIV prevention and treatment. PMID- 11256732 TI - Surveillance, social risk, and symbolism: framing the analysis for research and policy. AB - Name-based surveillance for HIV, considered alone, is a useful public health measure; its benefits outweigh its direct costs. There is little evidence that name-based surveillance directly deters individuals at risk of HIV from being tested, or exposes them to significant social risks. Yet such surveillance is chronically controversial. Understood in a broader context of the social risks and symbolic politics of HIV, as subjectively experienced by people at risk, this opposition is both rational and instructive. Although often discussed, the social risks of HIV infection are poorly understood. To the extent these risks have been addressed by privacy and antidiscrimination laws, the solution has been less complete than many public health professionals appear to believe: developments in law and policy, including the increasing prevalence of criminal HIV transmission laws and proposed changes in HIV testing and counseling standards, are contextual factors that help explain the opposition to name-based surveillance. Rather than focusing piecemeal on specific "barriers" to testing and care, an appreciation of the surveillance debate in context suggests a positive undertaking in public health policy to provide the conditions of opportunity, information, motivation and confidence that people with HIV need to accept an effective program of early intervention. PMID- 11256733 TI - Voluntary counseling, testing, and referral for HIV: new technologies, research findings create dynamic opportunities. AB - Programs for voluntary counseling and testing (VCT) for HIV play an increasingly important role in comprehensive prevention and care strategies. New technological advancements and behavioral interventions can improve the effectiveness of VCT as a tool for preventing new HIV infections and helping HIV-positive individuals access appropriate care. With growing consensus that early access to HIV therapy increases its effectiveness, and that individuals diagnosed with HIV reduce risk behavior, VCT has become integral to the continuum of HIV primary care. However, federal funding of VCT has declined, with concomitant decreases in numbers of people being tested. An estimated 200,000 people in the United States remain unaware that they are HIV positive, and many at-risk individuals do not seek out standard HIV counseling and testing services. To increase the acceptability and effectiveness of VCT, the authors recommend that VCT programs employ outreach programs offering anonymous testing to reach those at heightened risk of HIV infection, and to make rapid use of new technologies and counseling strategies to improve the reach and efficacy. Given the important role that VCT can play in both prevention and early treatment, the authors recommend significant increases in federal support. PMID- 11256734 TI - Ethical issues in early detection of HIV infection to reduce vertical transmission. AB - Proposals to make prenatal HIV testing routine and universal dramatize ethical issues regarding early detection of HIV. These proposals would abolish pretest counseling and written informed consent for prenatal HIV testing. Ethical concerns include whether pregnant women are adequately informed that they may refuse such testing and whether patients have an opportunity to obtain more detailed information about the benefits and risks of HIV testing in this context. Several pertinent research questions need to be studied, including whether pregnant women find routine universal HIV testing acceptable and whether safeguards adequately protect women who receive testing. If analogous policies to enhance early detection of HIV are considered in other clinical contexts, the important clinical and ethical differences between vertical transmission and other situations of HIV transmission must be kept clearly in mind. PMID- 11256735 TI - Early detection of HIV: assessing the legislative context. AB - Early detection of HIV has important implications for both prevention and treatment. Promoting HIV testing, and thereby early detection, however, is a complicated task that must balance the interests of public health, personal privacy, and legislative efforts to curb transmission. This article assesses the legislative context within which public health officials must operate to promote early HIV identification. Specifically, the article reviews United States laws regarding HIV testing passed over the course of 3 years, 1997 to 1999, at the state-not the federal-level. The new laws demonstrate such major themes as limiting confidentiality of HIV test results, mandating name-based HIV reporting, partner notification and newborn testing, and criminalizing nondisclosure of HIV status in sexual and needle-sharing situations. The article evaluates these new laws and their potentially negative impact on early detection, and assesses implications for practices such as informed consent for HIV testing. Outcome evaluations of newly implemented state laws are recommended. Policy makers must be aware that these policy changes can either encourage or discourage HIV testing. PMID- 11256736 TI - Accessing HIV testing and care. AB - With the many recent improvements in the medical management of HIV, the benefits of early detection of the virus have increased. People found to be HIV-positive can be offered immediate referrals for medical care and a comprehensive continuum of services. However, it is estimated that, among the 650,000 to 900,000 seropositive persons in the United States, about one third are unaware of their serostatus. Many of those who are tested for HIV do not return for their results. Among those less likely to return for results are young people and black Americans. Many factors at the individual, system and societal levels negatively impact whether individuals at risk for HIV seek HIV testing in the first place, whether they return for their results, and whether they get appropriate care after they are found to be HIV-positive. Some solutions are offered to improve the identification of new HIV infections. These include social marketing campaigns to encourage individuals to be tested for HIV. Also, more use of the rapid HIV test, which will substantially increase the number of people obtaining their HIV results, is recommended. New computer technologies, such as telemedicine, also have the potential to improve linkages to care for newly diagnosed individuals. In addition, it is essential that HIV care continue to be readily available through the Ryan White Care Act. PMID- 11256737 TI - Early detection: the next steps. PMID- 11256738 TI - Early detection of HIV: implications for prevention research. Proceedings of a conference. PMID- 11256739 TI - HIV testing from a community perspective. AB - The communities of people impacted by AIDS in the United States are diverse and have divergent opinions about HIV antibody testing. Concerns over social issues like stigma, confidentiality, and disclosure, combined with a legacy of advocacy, have shaped testing policies and praxis since the introduction of HIV antibody testing in 1985. The continued impact of the HIV epidemic on the underserved and socially marginalized focuses these concerns and highlights the importance of linkages between testing and care and treatment. The development of new testing technologies challenges HIV communities to develop a new rhetoric promoting early detection of the disease. PMID- 11256740 TI - The context of HIV/AIDS surveillance. AB - HIV surveillance and diagnostic testing for HIV infection share elements in common, yet differ notably in context. Clinical testing provides vital information for individual medical and behavioral decisions, whereas surveillance, which focuses on populations, provides information to develop policy, direct resources, and plan services. HIV/AIDS surveillance has evolved over the course of the epidemic, reflecting changes in scientific knowledge, populations affected, and information needs. Likewise, the benefits of early diagnosis of HIV have become increasingly apparent with advances in HIV treatment. This article examines the changing context of HIV/AIDS surveillance and discusses the potential impact of HIV surveillance practices and policies on HIV testing behaviors. Special emphasis is placed on the importance of protecting the confidentiality of HIV/AIDS surveillance data and on the role of health department in monitoring the impact of surveillance policies on test-seeking patterns and behaviors. PMID- 11256741 TI - Toll and Toll-like proteins: an ancient family of receptors signaling infection. AB - Innate immunity is the first-line host defense of multicellular organisms that rapidly operates to limit infection upon exposure to microbes. It involves intracellular signaling pathways in the fruit-fly Drosophila and in mammals that show striking similarities. Recent genetic and biochemical data have revealed, in particular, that proteins of the Toll family play a critical role in the immediate response to infection. We review here the recent developments on the structural and functional characterization of this evolutionary ancient and important family of proteins, which can function as cytokine receptors (Toll in Drosophila) or pattern recognition receptors (TLR4 in mammals) and activate similar, albeit non identical signal transduction pathways, in flies and mammals. PMID- 11256742 TI - Mannose-binding lectin: structure, function, genetics and disease associations. AB - Mannose-binding lectin (MBL), a serum protein characterised by both collagenous regions and lectin domains, plays an important role in innate immune defence. It binds to the repeating sugar arrays on many microbial surfaces through multiple lectin domains and, following binding, is able to activate the complement system via an associated serum protease, MASP-2. Serum levels of MBL are influenced by three mutations clustered in exon 1 of the gene and are further modulated by various promoter region polymorphisms. The exon 1 mutations lead to secondary structural abnormalities of the collagenous triple helix and a failure to form biologically functional higher order oligomers. There is an increased incidence of infections in individuals with such mutations and an association with the autoimmune disorders SLE and rheumatoid arthritis. Nevertheless, MBL genotyping of various populations has led to the suggestion that there may be some biological advantage associated with absence of the protein. These and other findings suggest that the concept of MBL as a protein involved solely in first line defence is an oversimplification and the protein should rather be viewed as having a range of activities including disease modulation. PMID- 11256743 TI - Genetics of the complement system. AB - After a brief history of complement genetics, general considerations and applications to our understanding of immune function, evolution, population structure and migration and forensic medicine, selected topics in complement genetics are presented. For individual complement proteins, genetic polymorphisms and deficiency states are described, as are the molecular bases of some of them. The clinical abnormalities exhibited by some patients with complement deficiency states are discussed, as are possible pathophysiologic mechanisms for them. The chromosomal location and the close linkage and a sharing of structural features by groups of complement proteins, such as the complotypes of the major histocompatibility complex, the regulators of complement activation, Clr and Cls, and the terminal components C6, C7 and C9, are presented in some detail. From these facts, the broad outlines are drawn of the evolution of the classical and alternative complement pathways from the lectin pathway and the terminal pathway from a common progenitor. From markers within the complotype region, rough conclusions are delineated regarding the evolution of C2, factor B, C4A and C4B alleles. PMID- 11256744 TI - Chemokines and innate immunity. AB - Our environment contains a great variety of infectious microbes that may be potentially destructive and threaten our survival. As soon as microbes try to establish a site of infection, the host launches a complex defense system. Innate immunity is a non-specific response and serves as the first-line of defense where phagocytes, such as neutrophils and macrophages, and NK cells play central roles in neutralizing and clearing microorganisms. Thus, migration of cells into infectious foci and subsequent activation of these cells appear to be a critical step, enabling the host to achieve effective and efficient removal of microbes. Over the past decade, chemokines have been identified as chemotactic cytokines that attract and activate specific types of leukocyte populations in vitro. There is now evidence that the magnitude of chemokines' expression in infectious diseases is strongly associated with the severity of the inflammatory responses. Blocking chemokines or their receptors with neutralizing antibodies or gene targeting technology has allowed us to understand the pathological significance of chemokines in animal models of infectious diseases. Growing evidence suggests that chemokines play an important beneficial role in immune system development, homeostasis and in innate immunity, which may pave the way for new therapeutic strategies for the treatment of infectious diseases. PMID- 11256745 TI - Cytokines in innate and adaptive immunity. AB - Cytokines and chemokines are hormone-like messengers which act to regulate the development and expression of the broad array of immune responses that are mounted against a variety of pathogens. As such, they are critical determinants of the types of cells which will regulate and participate in innate and adaptive immune responses, they may act both in highly localised environments but also in a systemic manner, and they may, themselves, directly mediate antimicrobial effector activities. In this article, we will outline current concepts of the activities of cytokines and chemokines in the immune response and discuss the various cell types, including dendritic cells and other antigen-presenting cells, T cells and B cells, which both produce and respond to these potent regulatory molecules. PMID- 11256746 TI - Interferon activation and innate immunity. AB - The interferons are a family of cytokine mediators critically involved in alerting the cellular immune system to viral infection of host cells. Interferons not only exhibit important antiviral effects but also exert a key influence on the quality of the cellular immune responses and amplify antigen presentation to specific T cells. Type I interferon (IFN-alpha and IFN-beta) is secreted by virus infected cells while type II, immune or gamma interferon (IFN-gamma) is mainly secreted by T cells, natural killer (NK) cells and macrophages. Interferons interact with specific cellular receptors, which promote production of second messengers ultimately leading to expression of antiviral and immune modulatory genes. The IFN genes themselves are regulated by transcriptional and posttranscriptional mechanisms including modulation by a family of interferon regulatory factors (IRFs) synthesised by host cells. IFNs activate macrophages, induce B cells to switch immunoglobulin type, alter T helper response, inhibit cell growth, promote apoptosis and induce an antiviral state in uninfected cells. The therapeutic potential of the IFNs is currently the focus of intense attention in a number of virus-associated diseases, tumours and autoimmune disorders. PMID- 11256747 TI - NADPH oxidase, Nramp1 and nitric oxide synthase 2 in the host antimicrobial response. AB - Using highly conserved, complex enzyme systems, leukocytes utilize the toxic nature of free radical intermediates, derived from oxygen and nitrogen, to control microbial pathogens as part of the innate immune response. Upon activation, NADPH oxidase generates superoxide anion radicals, which in turn give rise to further reactive oxygen intermediates. Similarly, activated nitric oxide synthase 2 catalyses the production of nitric oxide radicals, which leads to the formation of reactive nitrogen intermediates. Nitrogen- and oxygen-centered reactive intermediates can interact to form further reactive species. In addition, presence of the cationic transporter, Nrampl, may exacerbate the effects of these toxic compounds on invading microbes. While each of these antimicrobial systems can operate independently, the combination of their activities is synergistic in the successful containment of almost all invading pathogens. These systems are activated and modulated by microbial products and a series of temporally expressed cytokines. They also feed directly into the initiation of the adaptive immune response, which culminates in lasting specific immunity. The effector molecules, generated in the early innate immune response, are not specific to the invading pathogen and may also cause damage to the host. It is the critical balance of these processes in the initial stages of infection that determines the outcome of infectious disease. PMID- 11256748 TI - CD1 proteins: targets of T cell recognition in innate and adaptive immunity. AB - The CD1 family consists of antigen presenting molecules encoded by genes located outside of the major histocompatibility complex. CD1 proteins are conserved among mammalian species and are expressed on the surface of cells involved in antigen presentation. The CD1 system has been shown to be involved in activation of cell mediated responses, and T cells specific for either CD1 molecules or antigens presented by CD1 have been isolated. Structural and biochemical analyses demonstrate that antigens presented by CD1 are nonpeptide lipid or glycolipid structures, including examples found in the cell walls of pathogenic mycobacteria. The hydrophobic part of these antigens most likely binds in the CD1 ligand-binding groove, whereas the polar headgroup of these antigens appears to make direct contact with the T cell receptor and determines specific recognition. Presentation of antigens by CD1 molecules requires uptake and intracellular processing by antigen presenting cells and can be achieved for both exogenous and endogenous antigens. T cells recognizing CD1 restricted antigens have a broad range of functional activities that suggest that the CD1 system is involved in both innate and adaptive immune responses against microbial infections. PMID- 11256749 TI - Natural killer cell recognition of HLA class I molecules. AB - Human NK cells express multiple receptors that interact with HLA class I molecules. These receptors belong to one of two major protein superfamilies, the immunoglobulin superfamily or the C type lectin superfamily. The killer cell immunoglobulin-like receptor (KIR) family predominantly recognise classical HLA class I molecules and different family members interact with discrete HLA class I allotypes. The solution of the crystal structure of KIR2DL2 in complex with its ligand, HLA-Cw3 has provided the molecular details of a KIR/class I interaction. The interaction site spans both the alpha1 and alpha2 helices of class I and the KIR makes direct contact with peptide residues 7 and 8. The allotype specificity of KIR2DL2 for HLA-Cw3 is the result of a single hydrogen bond from Lys44 of the KIR to Asn80 of HLA-C as all other HLA-C residues that contact KIR are conserved. The lectin-like CD94/NKG2 receptors specifically interact with the non-classical class I molecule, HLA-E. Cell surface expression of HLA-E is dependent on the expression of other class I molecules as they are the major source of HLA-E binding peptides in normal cells. Consequently recognition of HLA-E by the CD94/NKG2 receptors allows NK cells to indirectly monitor the expression of a broad array of class I molecules. While the molecular interactions underlying ligand recognition by both KIR and CD94/NKG2 receptors are likely to be distinct, recognition of class I by both families of receptors appears peptide dependent. This suggest that cells that lack class I and also those that are impaired in their ability to load class I molecules with peptide will become targets for NK mediated destruction. PMID- 11256751 TI - Apoptosis in mouse fetuses from dams exposed to T-2 toxin at different days of gestation. AB - T-2 toxin (2 mg/kg b.w.) was orally inoculated to pregnant mice at gestational day (GD) 8.5, 9.5, 10.5, 11.5, 12.5, 13.5, 14.5, 15.5 and GD 16.5, respectively, and the fetuses were examined 24 hours later. The number and region of pyknotic or karyorrhectic cells varied according to inoculation date. In the GD 13.5 subgroup, a moderate to high number of pyknotic or karyorrhectic neuronal cells were observed in the central nervous system, peri-ventricular zone to subventricular zone, and pyknosis or karyorrhexis were also observed in a small number of chondroblasts and chondrocytes. In the GD 16.5-subgroup, a moderate to high number of pyknotic or karyorrhectic cells were observed in the thymus and renal subcapsular parenchyma. The nuclei of these pyknotic or karyorrhectic cells were strongly stained by the terminal deoxy nucleotidyl transferase (TdT) mediated dUTP-digoxigenin nick end labeling method widely used for the in situ detection of apoptotic nuclei. In addition, a few fetuses from dams which were given T-2 toxin at GD 13.5 or GD 14.5 and killed at GD 17.5 showed skeletal abnormalities such as wavy ribs and short scapula. From the present findings and the well known fact that T-2 toxin readily crosses the rat placenta, it seems that T-2 toxin-induced apoptosis in the developing mouse fetuses might be a direct effect of T-2 toxin on fetuses. PMID- 11256750 TI - Tumors in long-term rat studies associated with microchip animal identification devices. AB - Tumors surrounding implanted microchip animal identification devices were noted in two separate chronic toxicity/oncogenicity studies using F344 rats. The tumors occurred at a low incidence rate (approximately 1 percent), but did result in the early sacrifice of most affected animals, due to tumor size and occasional metastases. No sex-related trends were noted. All tumors occurred during the second year of the studies, were located in the subcutaneous dorsal thoracic area (the site of microchip implantation) and contained embedded microchip devices. All were mesenchymal in origin and consisted of the following types, listed in order of frequency: malignant schwannoma, fibrosarcoma, anaplastic sarcoma, and histiocytic sarcoma. The following diagnostic techniques were employed: light microscopy, scanning electron microscopy, and immunohistochemistry. The mechanism of carcinogenicity appeared to be that of foreign-body induced tumorigenesis. PMID- 11256752 TI - Experimental hypoxia of STZ-diabetic rat myocardium and protective effects of Ginkgo biloba extract. II. Ultrastructural investigation of microvascular endothelium. AB - Four months after induction of diabetes by intraperitonal injection of 60 mg streptozotocin/kg body weight wistar rats were exposed to an acute respiratoric hypoxia of 20 min duration. One group of the rats received daily Ginkgo biloba extract EGb 761 (100 mg/kg body weight). By means of qualitative and quantitative electron microscopic analysis we compared the hypoxia-induced ultrastructural alterations of the myocardial microvascular endothelium in normal, diabetic, and EGb-protected rats. Aim of the study was to compare the hypoxia tolerance of myocardial microvessels of normal and diabetic rats and to test the possibility of antioxidative protection. The results revealed that only some ultrastructural microvascular parameters of diabetic rats were stronger altered after acute hypoxia than normal ones, e.g. capillary dilatation, number of lysosomes, frequency of vesicles and fused vesicles, endothelial swelling, and structural state of mitochondria. Other parameters exhibited less severe alterations than in healthy rats, as luminal blebbing and protrusions, endothelial vacuoles, mitochondrial swelling, and pericapillary debris. Protective effects of EGb could be demonstrated on endothelial swelling, blebbing, vesiculation and vesicular fusioning, partly on vacuolization, but not on mitochondrial parameters. The results were discussed on pathobiochemical background. EGb 761 was estimated to be protective against hypoxic damage on myocardial microvessels also in diabetic condition, but the study should be completed by inclusion of a reoxygenation interval. PMID- 11256753 TI - Cytochrome P450 (P450) isoforms expression, P450 concentration, monooxygenase activities, reactive oxygen species formation, lipid peroxidation, and glutathione content in wild catch carp and tench liver--influence of a two weeks exposure to phenobarbital. AB - Carps, both sexes, 3 years old, weighing about 1 kg, and tenches of both sexes, 6 years old, weight about 250 g, were caught from a Thuringian lake without industrial pollution in November 1995 (fish without food uptake, water temperature at about 10 degrees C) and kept for 2 weeks in basins with clean water and addition of 0, 0.1, 1.0 or 10.0 mg/l phenobarbital-Na (PB). The concentration of PB was controlled during and at the end of the exposure period. The animals were fed pellets, but no food uptake was observed. After 24-48 h in fresh water the fish were sacrificed and the following hepatic parameters were immediately determined biochemically: monooxygenase functions: cytochrome P450 (P450) content, ethylmorphine N-demethylation (EN), ethoxycoumarin O-deethylation (ECOD), ethoxyresorufin O-deethylation (EROD), 7-benzyloxy-4-methyl-coumarin O debenzylation (BCDB); oxidase function indicators: microsomal Fe2+/NADPH dependent hydrogen peroxide formation (H2O2), microsomal Fe2+/NADPH dependent luminol and lucigenin amplified chemiluminescence (LMCL, LCCL), microsomal Fe2+/NADPH dependent lipid peroxide formation (LPO); oxidative state: lipid peroxidation products (TBARS) and GSH and GSSG. Additionally, the expression of three P450 isoforms, 1A1, 2B and 3A, was assessed immunohistochemically in tissue samples from brain, gill, heart, spleen, liver, gut and ovary of both fish species and in kidney of tenches. PB did not influence body or liver weights, but increased liver P450 concentration in both species by 50-100%, though not significantly. Carp: PB increased both EN and EROD significantly, but not ECOD and BCDB; H2O2 and TBARS were enhanced significantly. LPO, LMCL and LCCL were not significantly influenced. Tench: PB increased all monooxygenase reactions (EN, ECOD, BCDB and EROD), though only significantly ECOD; H2O2 was elevated only after treatment with 0.1 mg/l PB, whereas LPO was decreased (!) after treatment by all three concentrations, though significantly only after 1.0 mg/l PB. LMCL was depressed (not significantly), but LCCL increased 5fold. TBARS were significantly enhanced. P450 1A1 subtype expression was concentration dependently elevated by PB in gill and liver of both fish and in the heart and kidney of tenches, P450 2B and 3A isoforms expression was induced in brain, gill, heart, liver and gut of both fish and in the kidney of tenches. In summary, the increased activities of the monooxygenase reactions tested and the elevated expression of all three P450 isoforms investigated in certain tissues indicate an induction of the P450 families 1, 2 and 3 by PB in fish. PMID- 11256754 TI - Light and electron microscopic immunohistochemical investigation on G and D cells in antral mucosa in Helicobacter pylori-related gastritis. AB - Recently, it has been recognized that Helicobacter pylori (H. pylori) infection is associated with an exaggeration of basal and meal gastrin secretion. We investigate whether there is a relationship between H. pylori-related chronic gastritis and G-cell and D-cell number and granule density index of G and D cells. - The number of antral G cells and D cells and granule density index of D and G cells are compared between thirty two patients with H. pylori-related chronic gastritis and twelve patients without H. pylori and inflammation. Antral mucosal biopsy specimens are examined using light and electron immunohistochemical techniques. - The number of G cells is the same in either infected or uninfected patients (98.40 +/- 11.39, 109.25 +/- 12.76 vs 101.17 +/- 7.72 for infected patients with non atrophic and with mild atrophic chronic gastritis and uninfected controls, respectively) except for the cases with moderate gastric mucosal atrophy, where G cells (58.22 +/- 5.63) decrease in number. The number of D cells is decreased in all patients with H. pylori-related gastritis. G cell granule density index is significantly (p < 0.05) increased in patients with H. pylori-related chronic gastritis than in controls (3.15 +/- 0.43 vs 2.528 +/- 0.01). D cell granule density index is similar between patients with H. pylori chronic gastritis and controls (3.18 +/- 0.05 vs 3.166 +/- 0.12). It is concluded that decreased D cells number in patients with H. pylori-related chronic gastritis might be one of the reasons for the existing hypergastrinaemia. PMID- 11256756 TI - Spectrum and age-related incidence of spontaneous tumours in a colony of Han:AURA hamsters. AB - One-hundred-and-forty-four male and 184 female untreated Syrian golden hamsters (strain Han:AURA) were kept for life under standard laboratory conditions. They were examined with regard to spontaneously occurring tumours in relation to their survival periods. The mean survival rate of the males was 106 +/- 26 weeks and that of the females 97 +/- 20 weeks. Tumours were found in 71% of males and 67% of females. Adenomas and carcinomas of the adrenal glands were the most frequently observed tumours in both sexes (male: 66%; female: 38%) and in the early stages of life. Malignant lymphoma (8%), adenomas and carcinomas of pancreatic islet-cells (8%) and papillomatous benign and malignant squamous cell tumours of the forestomach (7%) showed relatively high incidences in males, whilst in females, leiomyoma (10%) and endometrial adenocarcinoma (7%) of the uterus and adenomas and carcinomas in the pars distalis of the pituitary gland (9%) occurred frequently. PMID- 11256755 TI - Transplantation of fetal liver tissue suspension into the spleens of adult syngenic rats: effects of different cytotoxins on cytochrome P450 mediated monooxygenase functions and on oxidative state. AB - Syngenic fetal liver tissue suspensions were transplanted into the spleens of adult male Fisher 344 inbred rats. Four months after surgery, transplant recipients and age matched control rats were treated with different cytotoxins (allyl alcohol [AAL], bromobenzene [BBZ], carbon tetrachloride [CCl4], or thioacetamide [TAA]) or the respective solvents 24 or 48 hours before sacrifice. Effects of the cytotoxins on P450 mediated monooxygenase functions in liver and spleen 9,000 g supernatants were assessed by measuring the model reactions ethoxyresorufin O-deethylation (EROD), ethoxycoumarin O-deethylation (ECOD), pentoxyresorufin O-depentylation (PROD), and ethylmorphine N-demethylation (EMND). Additionally, the influence on the oxidative state was investigated by assessing the liver and spleen tissue content of lipid peroxidation (LPO) products and of reduced and oxidized glutathione (GSH;GSSG). The livers of both solvent treated transplant recipients and control rats displayed regular EROD, ECOD, PROD and EMND activities. After AAL treatment EROD and EMND activities within the livers were not affected, but ECOD and PROD activities were increased. BBZ administration caused a decrease in EROD and EMND activities, ECOD activity remained unaffected, and PROD activity was even increased. CCl4 and TAA administration caused a strong reduction in the activity of all four model reactions. Spleens of control rats displayed almost no P450 mediated monooxygenase functions, independent whether the rats had been treated with the cytotoxins or not. In the transplant containing spleens, however, significant EROD and ECOD, but hardly any PROD or EMND activities were seen. After AAL administration EROD activity was not affected in the transplant containing spleens, but ECOD activity was increased. BBZ treatment led to a decrease in EROD and an elevation in ECOD activity. CCl4 and TAA strongly reduced the activity of both of these model reactions. The tissue content of LPO products within livers and transplant containing spleens was significantly increased after BBZ and CCl4 treatment. An elevation in LPO products was also seen in the spleens of the control rats due to CCl4 administration. Tissue GSH and GSSG content in both livers and transplant containing spleens were strongly reduced after BBZ treatment. After CCl4 administration only a significant decrease in liver GSSG contents was seen. TAA treatment caused a reduction in the GSH and GSSG content in the spleens of both transplant recipients and control rats, but not in the livers. From these results it can be concluded, that the effects of cytotoxins like AAL, BBZ, CCl4 or TAA on P450 dependent monooxygenase functions and on oxidative state are exerted in the ectopic intrasplenic liver cell transplants in a similar way as in normal orthotopic liver. PMID- 11256757 TI - Valproate, but not lamotrigine, induces ovarian morphological changes in Wistar rats. AB - Valproate (VPA) medication is associated with development of polycystic ovaries, menstrual disorders and hormonal changes in women with epilepsy. We sought to determine if changes in the ovaries also occurred in an animal model without epilepsy, and whether this effect could be related to a carcinogenic effect expressed by overexpression of p53. A potentially alternative antiepileptic drug, lamotrigine (LTG), was evaluated simultaneously. To this end, female Wistar rats were fed perorally with VPA 400 mg/kg/day (n = 15), VPA 600 mg/kg/day (n = 20), LTG 10 mg/kg/day (n = 15) or control solution (n = 15) for 90-95 days. There was a significant, dose-dependent increase in the number of follicular cysts, reduction in the number of corpora lutea and reduction of ovarian weight in the VPA group. No ovarian pathology was observed in the LTG group. In neither of the groups were morphological changes seen in other organs, nor was there any overexpression of the tumor suppressor gene p53 found. An alternative antiepileptic drug, LTG, showed no ovarian pathology, and there were no light microscopic changes in other organs, or evidence of pathologic p53 overexpression in the LTG-treated animals. PMID- 11256758 TI - Kinetics of apoptosis-related genes mRNA expression in the dorsal skin of hypotrichotic WBN/ILA-ht rats after topical application of T-2 toxin. AB - The expression of apoptosis-related genes mRNAs was examined in the dorsal skin of hypotrichotic WBN/ILA-Ht rats topically applied with T-2 toxin (10 microl of 0.5 microg/microl solution). The total mRNA was obtained from skin biopsy samples from each rat at 3, 6, 12 and 24 hours after T-2 toxin treatment (HAT), and RT PCR was carried out with pairs of oligonucleotide primers corresponding to the cDNA sequences of rat p53, bcl-2, c-ki-ras, c-fos and c-jun oncogenes. The expression of c-fos mRNA markedly increased at 3 HAT, peaked at 6 HAT, and greatly decreased at 12 HAT. However it maintained a higher level, compared with the control level, even at 24 HAT. Although not prominent, the expression of c jun mRNA also showed significant elevation from 3 to 12 HAT. On the other hand, there were no changes in the expression of p53, bcl-2 and c-ki-ras mRNAs throughout the observation period. Judging from the present results and our previous report that epidermal cells developed apoptosis at 12 HAT (Histol Histopathol 1999; 14: 337-342), the induction of c-fos and perhaps of c-jun mRNAs may be associated with T-2 toxin-induced epidermal cell apoptosis. PMID- 11256759 TI - The influence of hypoxia on the myocardium of experimentally diabetic rats with and without protection by Ginkgo biloba extract. III: Ultrastructural investigations on mitochondria. AB - Completing our preceding ultrastructural studies on diabetes and additional acute hypoxia of rat myocardium and the protective effect of Ginkgo extract (EGb) we investigated specific ultrastructural-morphometric parameters of corresponding mitochondria. Aim of the study was to answer the question whether mitochondria of diabetic myocardium are more sensitive to hypoxia than in normal condition, and whether antioxidative protection by EGb is effective. Further we compared the ultrastructural reactions of mitochondria of different intracellular locations. Voluminal parameters of mitochondria indicated a moderate swelling after diabetes and a further slight swelling after additional hypoxia, which was slightly reduced after EGb pretreatment. Decrease of volume density of mitochondrial cristae was less expressed after diabetes and much stronger after additional hypoxia; slight protection by EGb was only visible after diabetes. Degenerative intramitochondrial areas increased significantly after diabetes and after hypoxia; EGb was protective only after additional hypoxia. The relative number of ATPase particles (F1-coupling factors) at the inner mitochondrial membranes was slightly but significantly reduced after diabetes and stronger reduced after additional hypoxia; only in the latter condition Ginkgo extract was slightly protective. The product of volume density of mitochondria x volume density of cristae x relative number of ATPase particles at the inner mitochondrial membrane (as structural equivalent of the myocardial capacity for ATP production) indicated better than single parameters the increasing mitochondrial damage after diabetes of 4 months duration and subsequent acute hypoxia of 20 min duration. After hypoxia this capacity amounted only to 46% of the normal and was improved by EGb to 53%. PMID- 11256760 TI - The psychology of action. AB - Actions are part of the way that the mind controls the body. Two fundamental psychological questions about actions are 'Where do they come from?' and 'How does the mind produce them?' These may be called the 'internal generation problem' and the 'information expansion problem' respectively. The importance of these questions was appreciated at the birth of the British Psychological Society (BPS) a century ago, though the experimental methods to study them were lacking. This article falls into two halves. The first half discusses some of the major epochs in the psychology of action over the last 100 years; the second half outlines some currently prominent research questions, and considers their historical antecedents. Finally, I offer some speculations regarding where future contributions to the psychology of action will be most fruitful. PMID- 11256761 TI - The language machine: psycholinguistics in review. AB - Psycholinguistics is the empirical and theoretical study of the mental faculty that underpins our consummate linguistic agility. This review takes a broad look at how the field has developed, from the turn of the 20th century through to the turn of the 21st. Since the linguistic revolution of the mid-1960s, the field has broadened to encompass a wide range of topics and disciplines. A selection of these is reviewed here, starting with a brief overview of the origins of psycholinguistics. More detailed sections describe the language abilities of newborn infants; infants' later abilities as they acquire their first words and develop their first grammatical skills; the representation and access of words (both spoken and written) in the mental lexicon; the representations and processes implicated in sentence processing and discourse comprehension; and finally, the manner in which, as we speak, we produce words and sentences. Psycholinguistics is as much about the study of the human mind itself as it is about the study of that mind's ability to communicate and comprehend. PMID- 11256762 TI - Is the mind a cauliflower or an onion? British insights into cognitive organization from the study of abnormal function. AB - Clinical and normal psychology have had a long tradition of close interaction in British psychology. The roots of this interplay may predate the development of the British Psychological Society, but the Society has encouraged and supported this line of research since its inception. One fundamental British insight has been to consider the evidence from pathology as a potential constraint on theories of normal function. In turn, theories of normal function have been used to understand and illuminate cognitive pathology. This review discusses some of the areas in which clinical contributions to cognitive theory have been most substantial. As with other contributions to this volume, attempts are also made to read the runes and anticipate future developments. PMID- 11256764 TI - Individual differences in cognition: British contributions over a century. AB - Research on individual differences in mental abilities is discussed from three viewpoints: the psychometric structure of ability differences, the predictive validity of mental test scores, and some putative causes of psychometric intelligence differences in terms of psychometric and cognitive components and biological indices. A hierarchical descriptive structure for mental ability differences, as it emerged in the 1980s and 1990s, prominently displays British discoveries and suggestions, especially those of Spearman, Burt and P. E. Vernon from the first half of the 20th century. Galton and Spearman's largely unproductive search for the origins of ability differences has seen new activity since the 1970s, and there are several replicable associations that are yet to be explained. Over the 20th century the emphasis has been on measuring mental ability differences; at its beginning there was an emphasis (largely British) on understanding psychometric intelligence. The new century is likely to see a continuation of this re-emphasis on explaining human ability differences. PMID- 11256763 TI - Human rationality and the psychology of reasoning: where do we go from here? AB - British psychologists have been at the forefront of research into human reasoning for 40 years. This article describes some past research milestones within this tradition before outlining the major theoretical positions developed in the UK. Most British reasoning researchers have contributed to one or more of these positions. We identify a common theme that is emerging in all these approaches, that is, the problem of explaining how prior general knowledge affects reasoning. In our concluding comments we outline the challenges for future research posed by this problem. PMID- 11256765 TI - Beatrice Edgell: an appreciation. AB - Beatrice Edgell's contribution to the development of psychology is assessed. Edgell was Head of the Department of Philosophy and Psychology at Bedford College, London, from 1898 to 1933. She did much to develop the status of psychology within the College and the University, and established one of the first psychological laboratories in Britain. She was the first British woman to gain a doctorate in psychology, the first woman Professor of Psychology in Britain and the first woman President of the British Psychological Society (as also of the Aristotelian Society, the Mind Association, and the Psychology Section of the British Association for the Advancement of Science). She made substantial contributions to research, both theoretical and empirical, including work on the Wheatstone-Hipp chronoscope and on memory, and trained a number of women who subsequently played a prominent role in the development of both academic and applied psychology in Britain. PMID- 11256766 TI - Social cognition: categorical person perception. AB - In attempting to make sense of others, perceivers regularly construct and use categorical representations (e.g. stereotypes) to streamline the person perception process. A debate that has dominated recent theorizing about the nature and function of these representations concerns the conditions under which they are activated in everyday life. The present article reviews this work and considers the automaticity of category activation in person perception. PMID- 11256767 TI - Cognitive development: no stages please--we're British. AB - British cognitive developmental psychology is characterized by its interest in philosophical questions, its preference for linking basic research to applied issues in education and cognitive disorders, and its willingness to learn both methodologically and theoretically from work in animal psychology and in physiology more generally. It has also been influenced profoundly by Jean Piaget's cognitive stage theory although in general British work has focused on demonstrating early strengths, rather than early deficits, in infant and child cognition. Following an overview of British work that encompasses past and present interests, issues and challenges for the future are highlighted. While the perspectives of the founding members of the British Psychological Society (BPS), as outlined by Edgell (1947), are still apparent in British research in cognitive developmental psychology today, it is argued that future cognitive work must become even more interdisciplinary and that the symbiotic relationship between research in adult cognition and in cognitive development needs greater recognition. PMID- 11256768 TI - The British Psychological Society. 1947. PMID- 11256769 TI - British memory research: a journey through the 20th century. AB - A century of research in memory has generated a wealth of knowledge encompassing theoretical developments within a number of distinct domains of memory. The aim of this article is to explore the progress made in memory research during the 20th century, to indicate critical influences on the direction of research, and to illustrate the important contribution made by British researchers. This article is confined to human memory research, and reviews research findings from the various psychological disciplines studied over the past 100 years. PMID- 11256770 TI - A selective review of selective attention research from the past century. AB - Research on attention is concerned with selective processing of incoming sensory information. To some extent, our awareness of the world depends on what we choose to attend, not merely on the stimulation entering our senses. British psychologists have made substantial contributions to this topic in the past century. Celebrated examples include Donald Broadbent's filter theory of attention, which set the agenda for most subsequent work; and Anne Treisman's revisions of this account, and her later feature-integration theory. More recent contributions include Alan Allport's prescient emphasis on the relevance of neuroscience data, and John Duncan's integration of such data with psychological theory. An idiosyncratic but roughly chronological review of developments is presented, some practical and clinical implications are briefly sketched, and future directions suggested. One of the biggest changes in the field has been the increasing interplay between psychology and neuroscience, which promises much for the future. A related change has been the realization that selection attention is best thought of as a broad topic, encompassing a range of selective issues, rather than as a single explanatory process. PMID- 11256771 TI - Surveying the seen: 100 years of British vision. AB - Perceptual phenomena and their interpretations have fashioned the course of psychology. This article surveys how theories of visual perception and methodologies have developed during the lifetime of the British Psychological Society. The experimental study of vision was instigated by British natural philosophers in the early nineteenth century but this impetus was not maintained thereafter. Not until the 1930s and 1940s did research on perception resume in earnest within British universities. The adoption of concepts (such as schema) potentially grounded in neural organization, particularly by Bartlett and Craik, accelerated experimental, theoretical and applied vision research. From mid century the influence of information processing models of perception became increasingly dominant, and they were often integrated with the rapidly expanding understanding of neurophysiological underpinnings. The epitome of these developments was Marr's model of vision which, in our view, marked the start of the modern era of vision research. Computers have transformed the nature of stimulus control and response measurement in perceptual experiments. More naturalistic stimuli can be presented and manipulated, and complex behavioural responses, such as patterns of eye movements, fractionated. Non-invasive recording of brain activity to visual stimulation has similarly been transformed with a variety of methods for imaging brain activity. Neuroimaging has been applied to localizing perceptual and cognitive functions and in studying patients with known deficits in visual recognition. However, the eagerness with which the computer has been adopted by perceptual psychologists is likely to be tempered by a growing awareness of the differences between viewing scenes and simulations of them. PMID- 11256772 TI - Dyslipidemia and coronary heart disease: management issues from Indian perspective. PMID- 11256773 TI - Coronary atherosclerosis: lessons learned from intravascular ultrasound. PMID- 11256774 TI - Platelet glycoprotein IIb/IIIa antagonists in percutaneous coronary interventions. PMID- 11256776 TI - Silent myocardial ischaemia in patients with type II diabetes mellitus and its relation with autonomic dysfunction. AB - Accelerated coronary and peripheral vascular atherosclerosis is one of the most common and chronic complications of diabetes mellitus. A relatively recently analysed aspect of coronary artery disease in this condition is its silent or asymptomatic nature. We studied silent/asymptomatic myocardial ischaemia in unselected consecutive middle aged asymptomatic diabetics and controls by 24-hour ambulatory electrocardiographic monitoring, treadmill test and coronary angiography. Also, a relationship was sought between silent myocardial ischaemia and autonomic dysfunction. Thirty asymptomatic diabetics between the ages 35-60 without any documented evidence of coronary artery disease and as many controls (matched for age, sex, smoking habits, blood pressure, serum cholesterol and body mass index) were studied. All the diabetics and controls were subjected to treadmill test and 24-hour ambulatory electrocardiographic monitoring. Coronary angiography was done in those who were positive in treadmill test or 24-hour ambulatory electrocardiographic monitoring. Also five simple bedside tests for autonomic dysfuncton i.e. heart rate response to valsalva, deep breathing and orthostatic variation and blood pressure response to orthostatic variation and sustained handgrip were done in all the subjects. Those with two or more abnormal tests were diagnosed as having autonomic dysfunction. ST segment depression indicating silent myocardial ischaemia was seen in 14 (46.7%) out of 30 diabetics and in 3 (10.0%) out of 30 controls on both Holter and treadmill test (p=0.002). Also, diabetics had higher heart rate and greater number of supraventricular and ventricular ectopics than controls. Coronary angiography done in patients with silent ischaemia revealed higher prevalence of multivessel involvement and diffuse disease in diabetics as compared to controls. Half the diabetics (50%) and none of the control had autonomic dysfunction. Autonomic dysfunction was present in 85.7 percent of diabetics with silent ischaemia compared to 18.7 percent diabetics without silent ischaemia (p=0.001). PMID- 11256775 TI - Low molecular weight heparin for treatment of acute myocardial infarction (FAMI): Fragmin (dalteparin sodium) in acute myocardial infarction. AB - The benefit of using subcutaneous low molecular weight heparin for the treatment of acute myocardial infarction is not known. The aim of this study was to determine the efficacy of a low molecular weight heparin (dalteparin sodium) for the treatment of acute myocardial infarction in patients not treated with thrombolytic therapy. Twenty-nine cardiological centres from leading hospitals in India participated in this prospective, multicentre, double-blind, placebo controlled study in two phases which included 1128 patients with acute myocardial infarction. In the acute phase (between day 1 and 3 of admission) all the patients received a weight-adjusted dose of subcutaneous dalteparin (120 IU/kg twice daily). In the second, double-blind phase of acute myocardial infarction, patients were randomised to receive a fixed dose of dalteparin (7,500 IU) or an identical placebo injection for 30 days. A composite primary endpoint of death, reinfarction, recurrence of angina and emergency revascularisation was used. All the 1128 patients with acute myocardial infarction were included in the trial. In the acute phase, the composite primary endpoint was observed in 58 (5.1%) patients. Of 1037 paients who were randomly assigned to receive a fixed dose of dalteparin (n=519) or placebo (n=518), the composite primary event rate was 6.7 percent and 7.0 percent, respectively (RR 0.97; 95% CI 0.62-1.52; p=0.90). To conclude, treatment with dalteparin administered subcutaneously in a weight adjusted dose of 120 IU/kg twice daily resulted in a lower than expected mortality during the acute phase of myocardial infarction. A lower fixed once daily dose of 7,500 IU during the chronic phase did not confer additional protection. PMID- 11256777 TI - Coronary artery ectasia: angiographic, clinical profile and follow-up. AB - Out of 3200 coronary angiograms we reviewed, there were 144 cases of coronary ectasia--an incidence of 4.5 percent. Among these, 122 were associated with atherosclerotic coronary artery disease, i.e. coronary stenosis more than 50 percent (group A) and 22 not associated with coronary artery disease (group B). The patients in groups A and B were compared with age- and sex-matched patients (group C) (n=100) who had coronary artery disease alone without ectasia. The incidence of ectasia was not increased in patients with thoracoabdominal aortic aneurysm i.e. 2/154 (1.8%) or in patients with peripheral occlusive vascular disease i.e. 5/161 (3.1%). Ectasia was typed according to a modified version of the criteria proposed by Markis et al. Type II was the commonest, followed by type I, III and IV. Right coronary artery was the most commonly involved vessel by ectasia followed by left circumflex, left anterior descending artery and left main coronary artery. Diffuse ectasia was seen more frequently in right coronary artery and localised ectasia in left anterior descending artery. Patients in groups A and B had similar epidemiological characteristics, though more patients with ectasia alone (group B) had better left ventricular function and negative stress tests. The patients in group A had a similar incidence of previous myocardial infarction, coronary risk factor profile, treadmill exercise test status and severity of coronary artery disease when compared to group C. On a mean follow-up of 3+/-1.2 years, all the three groups had similar event rates. PMID- 11256778 TI - Coronary stent thrombosis: time course and clinical outcome. AB - The current clinical practice of stent implantation has changed over the last few years. We analysed the incidence and time course of stent thrombosis in patients undergoing successful coronary angioplasty and stenting over the last three years. All the patients were treated with aspirin and ticlopidine. A total of 13 patients experienced stent thrombosis. The mean age was 52+/-12 years; 12 were smokers and 10 had a recent history of myocardial infarction. None of these patients had received abciximab. The median time from stent implantation to stent thrombosis was 10 hours, with all the stent occlusions occurring within 18 hours of stent implantation procedure. All the patients underwent a repeat intervention at a median time of 30 minutes after the clinical suspicion of stent occlusion. On follow-up of 1 to 24 months, three patients developed reocclusion. In the present era of coronary angioplasty and stenting, when interventional procedures are not pre-planned and patients are treated with aspirin and ticlopidine or clopidogrel at the time of stent implantation, the incidence of stent thrombosis is low; it is seen mainly in patients with recent myocardial infarction, majority of them being smokers, and occurs within 18 hours in all the patients. PMID- 11256779 TI - Haemostatic changes in children with cyanotic and acyanotic congenital heart disease. AB - This study was undertaken to screen children with congenital heart disease for coagulation abnormalities and to compare the groups of cyanotic and acyanotic children with congenital heart disease with respect to abnormalities of the coagulation system. Following investigations were done in all the patients: complete blood count, erythrocyte sedimentation rate, peripheral smear examination, bleeding time, prothrombin time, activated partial thromboplastin time, assay of fibrinogen, D-dimer, factors VII and VIII and antithrombin III. Red cell indices were determined in 12 control, 12 acyanotic and 20 cyanotic children. Twenty-five patients each, with echocardiographically proven cyanotic and acyanotic congenital heart disease under 12 years of age constituted the study group; as many children of the same age group were included as the control group. The results showed isolated abnormalities of laboratory tests with equal frequency (28%) in acyanotic and cyanotic groups but coexisting abnormalities of more than one test were seen in significantly larger number of cyanotic children (5/25 and 16/25, respectively). A significant association was noted between thrombocytopenia and a high haematocrit in cyanotic patients. It is concluded that laboratory abnormalities of tests of haemostasis are more common in cyanotic congenital heart disease patients. The patterns of laboratory abnormalities suggest a chronic compensated disseminated intravascular coagulation at a subclinical level, reduced synthesis of clotting factors and/or deranged platelet aggregation in different subgroups of patients. PMID- 11256780 TI - Recording of double atrial potentials as a marker for isthmus block during ablation of atrial flutter. AB - Radiofrequency ablation is an established method for treatment of type I atrial flutter. The assessment of creation of complete bidirectional isthmus block following linear ablation of the isthmus is an integral part of ablation procedure. Conventionally, bidirectional isthmus block is tested by pacing on either side of ablation line and looking for reversal of activation sequence in the right atrium. We looked at the feasibility of recording double potentials, separated by an isoelectric interval along the ablation line as an alternative method to demonstrate bidirectional isthmus block. An attempt was made to record the double potentials following linear ablation of the cavotricuspid isthmus. Following ablation, bidirectional isthmus block was also tested by pacing from the coronary sinus os and the low-lateral right atrium. We could demonstrate double potentials in 9 of the 11 patients in whom we attempted to record them following linear ablation of flutter. The presence of bidirectional block by pacing from coronary sinus os and low lateral right atrium could be demonstrated in 10 (91%) patients. Thus, double atrial potentials, separated by an isoelectric interval can be demonstrated following ablation of atrial flutter. Double potentials, if demonstrable on coronary sinus os and low lateral right atrium pacing, could serve as an alternative marker of isthmus block. PMID- 11256781 TI - Atrial fibrillation: how effectively can sinus rhythm be restored and maintained after balloon mitral valvotomy? AB - Atrial fibrillation, commonly associated with rheumatic mitral stenosis, worsens the prognosis. We studied the efficacy of achieving and maintaining sinus rhythm in patients with chronic atrial fibrillation who underwent a successful balloon mitral valvotomy. Fifty-four patients (26 men, 28 women; age 36+/-8 years) received amiodarone 200 mg thrice daily in the first week, and thereafter a maintenance dose of 200 mg once daily. Electrical cardioversion was attempted at 1 and 3 months and patients were followed up at 6, 12 and 18 months. At the end of 1, 3, 6, 12 and 18 months 81 percent, 72 percent, 60 percent, 54 percent and 49 percent of patients, respectively, were in sinus rhythm. Only one patient had a severe adverse effect (hypothyroidism). Univariate analysis revealed that lower age, shorter duration of atrial fibrillation and smaller left atrial size was associated with successful restoration to sinus rhythm. On multivariate analysis, the duration of atrial fibrillation was the only significant predictor of long term maintenance of sinus rhythm. Amiodarone seems safe and reasonably effective in restoration and maintenance of sinus rhythm in patients of atrial fibrillation with rheumatic heart disease. PMID- 11256782 TI - Hypothalamic digoxin and neural regulation of blood pressure and vascular thrombosis. AB - The isoprenoid pathway produces three key metabolites--digoxin (membrane sodium potassium ATPase inhibitor and regulator of neurotransmitter/aminoacid transport), dolichol (regulates N-glycosylation of proteins) and ubiquinone (free radical scavenger). This was assessed in patients with essential hypertension, familial hypotension, acute coronary artery disease and acute thrombotic strokes. The pathway was also assessed in patients with right hemispheric, left hemispheric and bihemispheric dominance for comparison. In patients with acute coronary artery disease, acute thrombotic stroke, essential hypertension and right hemispheric dominance, there was elevated digoxin synthesis, increased dolichol and glycoconjugate levels and low ubiquinone and high free radical levels. There was also an increase in tryptophan catabolites, reduction in tyrosine catabolites, increase in cholesterol-phospholipid ratio and a reduction in glycoconjugate level of RBC membrane in this group of patients as well as in those with right hemispheric dominance. In patients with familial hypotension and left hemispheric dominance, the patterns were reversed. The role of a dysfunctional isoprenoid pathway and endogenous digoxin in the pathogenesis of essential hypertension and familial hypotension and in thrombotic vascular disease in relation to hemispheric dominance is discussed. PMID- 11256783 TI - Studies on oxidative stress, serum iron and iron binding capacity in subjects prone to the risk of coronary artery disease. AB - Lipid peroxidation in vitro and in vivo has been postulated to be involved in the development of atherosclerosis. It is also known that free iron catalyses the lipid peroxidation. Therefore, we assessed the status of oxidative stress in smokers, hypertensives and non-insulin dependent subjects, who were prone to coronary artery disease. In addition, superoxide dismutase levels and iron binding capacity were also measured to know their antioxidant defences. One hundred seventy-five consecutive subjects below 60 years of age were examined; they were then divided into three groups: one with coronary artery disease, another without coronary artery disease and a healthy control group. The patients having either of the one risk factors for coronary artery disease i.e. smoking, hypertension and/or diabetes were studied. Serum lipid peroxides, superoxide dismutase, serum iron and iron binding capacity were estimated. Oxidative stress was highest in smokers with coronary artery disease (3.11+/-0.79 mmol/ml) as compared to hypertensives (2.69+/-0.20 mmol/nl) and non-insulin dependent diabetics (2.78+/-0.19 mmol/ml). Superoxide dismutase activity was also significantly decreased (p<0.001) in smokers with coronary artery disease as compared to hypertensives and non-insulin dependent diabetes mellitus. Final step of stepwise logistic regression based on malondialdehyde and superoxide dismutase correctly predicted coronary artery disease status in 90 percent smokers. Serum iron and total iron binding capacity were not significantly different in risk prone subjects. However, among all risk prone subjects, smokers with coronary artery disease showed highest serum iron levels and decreased iron binding capacity. PMID- 11256784 TI - Hyperhomocyst(e)inemia presenting as multisite vascular thrombosis. PMID- 11256785 TI - 'Ping-Pong' inside the heart. PMID- 11256787 TI - Aortocaval fistula: a rare complication of abdominal aortic aneurysm. PMID- 11256786 TI - Bacterial endocarditis caused by abiotrophia defectiva (nutritionally variant streptococci). PMID- 11256788 TI - Ebstein's anomaly of tricuspid valve: association with coarctation of aorta. PMID- 11256789 TI - The construct of a simple clinic questionnaire to assess physical activity and its relative validity. PMID- 11256790 TI - Current status of therapeutic angiogenesis. PMID- 11256791 TI - Blueprint for a quadrileaflet aortic valve prosthesis. PMID- 11256792 TI - Determination of human angiotensin converting enzyme (ACE) gene polymorphisms in erectile dysfunction: frequency differences of ACE gene polymorphisms according to the method of analysis. AB - The D polymorphism of angiotensin converting enzyme (ACE) gene has been found to be associated with various diseases, and ACE may also be involved in the pathogenesis of erectile dysfunction. On the other hand, interpretation of the data on the association of DD genotype with various diseases is controversial, due to methodological and technical variations in detection of the polymorphisms. We investigated a possible association between the DD genotype and erectile dysfunction in a Korean population, and compared the frequency of ACEgenotypes using our multiplexed PCR method with those based on the conventional PCR method in a sample of erectile dysfunctional and control subjects. There was significant difference in the distribution of ACE genotypes between the erectile dysfunctional (conventional PCR) and the control subjects (multiplexed PCR) (chi2=7.395, p<0.05), but there was no significant difference in the distribution of the genotypes between both groups (chi2=0.815, p>0.05) when our multiplexed PCR method was used. Therefore our results suggest that especially the conventional PCR method for ACE gene polymorphism may require careful control and may need repeated testing to verify the insertion deletion (ID) heterozygotes, and that a multiplexed PCR method can markedly increase the detection rate of the I allele in ID heterozygotes. No association was found between I/D polymorphism and erectile dysfunctional subjects in the Korean population studied. PMID- 11256793 TI - Simple gas chromatography analysis of faecal butyrate: application to patients at risk of pouchitis. AB - In spite of the fact that pouchitis is the most frequently occurring and troublesome complication found in patients treated by ileo-anal anastomosis for ulcerative colitis, no biological marker currently exists to monitor the outcome of the disease. Since it has been noted faecal butyrate is reduced in patients with pouchitis, we developed a simple gas chromatography method to quantify butyrate in faecal water. This test is based on diethyl ether extraction with the use of methacrylic acid as an internal standard. We demonstrated that butyrate was effectively measured when this technique was applied to eleven patients with ileal-pouch anal anastomosis within the first year after the closure of their ileostomy. We also observed a noticeable reduction in the concentration of butyrate in patients who went on to develop a pouchitis. PMID- 11256794 TI - Polymorphisms in the apolipoprotein E gene regulatory region in relation to coronary heart disease and their effect on plasma apolipoprotein E. AB - In a previous study which examined the distribution of apolipoprotein E genotypes and plasma levels in a sample of male coronary heart disease (CHD) patients and controls, we found a significant excess of the genotypes carrying APOE*4 allele in CHD men (18.2%) vs. controls (9.6%) and an association between the APOE*4 allele and the lowest concentrations of apoE. In the present investigation, we re examined in the same samples two recently identified polymorphisms in the promoter region of APOE, -491A/T and -427T/C, which may alter the level of apoE expression. No differences in the distributions of the -491A/T genotypes and alleles were observed between cases and controls (-491*A = 0.760 and 0.757 respectively). Polymorphism -427T/C showed in CHD patients an excess of -427*C allele (patients vs. controls = 0.123 vs. 0.074) and corresponding genotypes that was marginally significant. Stratification of the samples according to the presence/absence of APOE*4 showed that the excess of the -427*C allele concerned only CHD patients not carrying APOE*4 allele (patients vs. controls = 0.133 vs. 0.061; p=0.017). This result suggests that the presence of -427*C allele could represent a risk for developing CHD in subjects with E2/E2, E3/E2, and E3/E3 genotypes. Studies carried out on patients with Alzheimer's disease demonstrated that -491A/T and -427T/C polymorphisms affect the level of plasma apoE. In the present study, carried out on CHD patients and controls, the genetic variation at -427 and -491 sites of the APOE regulatory region had no apparent effect on apoE plasma concentration. PMID- 11256795 TI - Determination of glycerol in plasma by an automated enzymatic spectrophotometric procedure. AB - The plasma concentration of glycerol, the backbone of triglycerides and the end product of triacylglycerol breakdown, is considered to reflect lipolysis in adipose tissue. We evaluated an automated enzymatic procedure for the measurement of glycerol in plasma. The assay was linear up to 250 micromol/l. The detection limit was 8 micromol/l. Recovery averaged 94% from spiked plasma and 104% from diluted plasma. An extensive precision study performed according to the guidelines of the National Committee for Clinical Laboratory Standards showed within-run coefficients of variation between 14.8% and 2.6% for concentrations ranging from 28 micromol/l to 164 micromol/l. The reference range for fasting healthy adult men was 14-69 micromol/l. Glycerol levels were significantly correlated with free fatty acid levels. This automated enzymatic method is rapid and reliable, and provides greater sensitivity or convenience than previously described procedures. PMID- 11256797 TI - The clinical value of microtransferrinuria and microalbuminuria in the prediction of pre-eclampsia. AB - The aim of this study is to determine whether the presence of microtransferrinuria and microalbuminuria detected in pregnant women who are free of symptoms can predict the subsequent development of preeclampsia. One hundred fifty five pregnant women were successfully followed from 10 weeks gestation up till delivery. Pre-eclampsia developed in 31 women (17 mild and 12 severe pre eclampsia), and eclampsia developed in two cases, whereas 124 women remained normotensive (controls). First morning urine specimens were collected during 10 to 12 weeks gestation and analyzed for microalbuminuria by a specific immunochemical test strip method. Mid-trimester mean arterial blood pressure (MAP) was also measured. Urinary microtransferrin levels in pregnant women who subsequently developed severe pre-eclampsia and eclampsia were significantly higher than those of pregnant women who remained normotensive. Microtransferrinuria as a predictor for pre-eclampsia had a sensitivity 93.5%, specificity 65%, positive predictive value 83% and negative predictive value 98.4%, whereas these values for microalbuminuria were: 50%, 58%, 50% and 91%, respectively. Urinary microtransferrin levels were significantly elevated in women with elevated MAP and in women who delivered low birth weight and low Apgar score babies. In conclusion, microtransferrinuria is a potentially more sensitive predictor of pre-eclampsia than microalbuminuria. Moreover, microtransferrinuria in early pregnancy might be a negative marker of fetal outcome in pre-eclampsia. PMID- 11256796 TI - Clinical applications of the 2nd generation assay for anti-TSH receptor antibodies in Graves' disease. Evaluation in patients with negative 1st generation test. AB - Detection of autoantibodies to the thyrotropin receptor by radioreceptor assays is largely requested in clinical practice for the diagnosis of Graves' disease and its differentiation from diffuse thyroid autonomy. Additionally, thyrotropin receptor antibodies (TRAb) measurement during antithyroid drug treatment can be useful to evaluate the risk of relapse after discontinuation of the therapy. Nevertheless, some patients affected by Graves' disease are TRAb-negative when a 1st generation assay is used. In this study we evaluated the diagnostic performance of a newly developed 2nd generation TRAb assay in 46 patients with Graves' disease with negative 1st generation TRAb assay results. A control group of 50 Graves' disease patients with positive 1st generation TRAb assay results, 50 patients with Hashimoto's thyroiditis and 50 patients with nodular goiter were also examined. Forty one of 46 patients with Graves' disease with negative 1st generation TRAb assay results showed a positive 2nd generation test. No differences were seen in control groups. In conclusion, the 2nd generation TRAb assay is more sensitive than the 1st generation test and should be used in clinical practice. Long-term prospective studies are needed to evaluate the prognostic role of the 2nd generation TRAb assay in Graves' disease. PMID- 11256798 TI - A non-icteric cholecystectomized patient with recurrent attacks of right epigastric pain and dilated common bile duct--do liver function tests predict bile duct stones? AB - Cholecystectomized patients with recurrent attacks of right epigastric pain and with dilated common bile duct are a clinical challenge. In a number of these patients dilatation of the common bile duct is explained as a normal postoperative state following cholecystectomy and the recurrent pain attacks are of origin other than bile disorder, but in some cases dilatation of the common bile duct and attacks are caused by bile duct stones. The aim of the present work was to study the value of common plasma liver function tests in predicting bile duct stones in the group of non-icteric cholecystectomized patients with recurrent attacks of right epigastric pain and with dilated common bile duct. The study population comprised 24 consecutive non-icteric cholecystectomized patients admitted for elective endoscopic retrograde cholangiopancreatography because of attacks of right epigastric pain and dilated common bile duct in ultrasonography. All the liver function tests seemed to assist in separating patients with bile duct stones (n=11) from those without (n=13). Alanine aminotransferase levels were significantly higher (p=0.05) in patients with bile duct stones than in those without, but also alkaline phosphatase (p=0.07), gamma-glutamyl transferase (p=0.09) and bilirubin (p=0.09) levels seemed to be higher in patients with bile duct stones than in those without, although the differences in these values did not reach statistical significance. In conclusion, common plasma liver function tests assist in separating patients with bile duct stones from those without in this small but clinically important group of non-icteric cholecystectomized patients with recurrent attacks of right epigastric pain and with dilated common bile duct. However, the actual value of these measurements is limited in clinical decision making since overlapping of values occured. PMID- 11256799 TI - Serum lactate dehydrogenase isoenzyme 1 and prediction of death in patients with metastatic testicular germ cell tumors. AB - The International Germ Cell Cancer Collaborative Group study of patients with metastatic testicular germ cell tumors showed that catalytic concentration of serum lactate dehydrogenase (S-LD), serum alpha-fetoprotein concentration (S AFP), and serum human chorionic gonadotropin concentration (S-hCG) predicted death from tumor. The recent international TNM classification (T primary tumor, N lymph node metastasis, M distant metastasis) is based on these results. The aim of our study was to evaluate whether catalytic concentration of S-LD isoenzyme 1 (S-LD-1) was a better predictor than the criteria used for the international classification. In an evaluation series of 44 patients from Odense University Hospital, Denmark, a raised S-LD-1 (>1.0 x upper limit of reference values) had a predictive value for death from tumor in 5-years observation of 46%. The predictive value was 46% for S-LD, 25% for S-AFP, and 40% for S-hCG. A normal SLD 1 had a predictive value for survival over 5-years observation of 100%. It was 81% for S-LD, 75% for SAFP, and 77% for S-hCG. The fraction of the patients who died of tumor and had a raised tumor marker value was 100% for S-LD-1, 46% for S LD, 9% for S-AFP, and 18% for S-hCG. The fraction of patients with a normal serum tumor marker value among those who survived was 61% for S-LD-1, 81% for S-LD, 94% for SAFP, and 94% for S-hCG. A validation series of 37 patients treated at the University of Texas MD Anderson Cancer Center showed similar findings. Combining the patients in the two series, a raised value of SLD-1 classified more patients into a subgroup with an impaired survival (53%) than S-LD (35%), S-AFP (6%), or S hCG (11%), and the high risk subgroups based on the international classification (40%). The findings have implications for the staging and treatment of patients with metastatic testicular germ cell tumors. PMID- 11256800 TI - Annual rhythmic and non-rhythmic biological variation of magnesium and ionized calcium concentrations. AB - The annual inter- and intra-individual biological variation, including the circannual rhythmic variation, of the serum concentrations of magnesium and ionized calcium has been investigated in a group of 51 apparently healthy volunteers. Venous blood specimens were collected on intervals of once a month within a one-year period, using a standardized protocol. The inter-individual coefficients of variation were 5.12% for magnesium and 1.58% for ionized calcium. The medians of the intra-individual coefficients of variation were 1.93% for magnesium and 2.18% for ionized calcium. These data were used to determine the allowable imprecision, the allowable systematic error, the critical difference for significant change detection, and the usefulness of population reference values (index of individuality). Of the quantities studied, only the serum concentration of ionized calcium shows a significant annual rhythmic variation (amplitude 1.3%), although this result may be due to the between-run metrological variance, considering that the concentration of ionized calcium of the control material used during the study possesses a similar significant rhythmic variation. PMID- 11256801 TI - Unit-independent reporting of laboratory test results. AB - A variety of laboratory measurement units, assay systems, and reference values are presently being used in different clinical environments. There is therefore a challenge in provision of tools for presentation of laboratory data in harmonized forms that reduce the information load and reinforce clinical perception of test results. This paper describes the use of standard deviation (SD) units and logarithmic time graphical displays to improve presentation of laboratory test results. The SD concept employs strength of the change from normality as a diagnostic indicator, and the logarithmic time scale enables long-term overview of patient results. The suggested format is expected to promote effective transfer of test result information between laboratories, clinicians and hospitals. PMID- 11256802 TI - Analysis of large and small samples of biochemical and clinical data. AB - Statistical software often offers a list of various descriptive statistics of location and scale, but rarely selects an efficient estimate that is statistically adequate for an actual univariate sample. The sample interval estimate for a specified degree of uncertainty seems to be more meaningful if it covers an unknown value of the population parameter. The concept of an interval estimate in medicine is then used for medical decision-making. The proposed methodology, which uses the S-Plus algorithm for biochemical, biological and clinical data analysis contains the following steps: (i) Exploratory data analysis identifies basic statistical features and patterns of the data, the distributions of which are mostly non-normal, non-homogeneous and often corrupted by outliers. (ii) Sample assumptions about data, independence of sample elements, normality and homogeneity are examined. (iii) Power transformation and the Box Cox transformation to improve sample symmetry and stabilize the spread. (iv) Classical and robust statistics for both large (n>30) and medium-sized samples (156 glycosidic linkages. PMID- 11256810 TI - Base-type-selective high-resolution 13C edited NOESY for sequential assignment of large RNAs. AB - Extensive spectral overlap presents a major problem for the NMR study of large RNAs. Here we present NMR techniques for resolution enhancement and spectral simplification of fully 13C labelled RNA. High-resolution 1H-13C correlation spectra are obtained by combining TROSY-type experiments with multiple-band selective homonuclear 13C decoupling. An additional C-C filter sequence performs base-type-selective spectral editing. Signal loss during the filter is significantly reduced because of TROSY-type spin evolution. These tools can be inserted in any 13C-edited multidimensional NMR experiment. As an example we have chosen the 13C-edited NOESY which is a crucial experiment for sequential resonance assignment of RNA. Application to a 33-nucleotide RNA aptamer and a 76 nucleotide tRNA illustrates the potential of this new methodology. PMID- 11256811 TI - Chemical shifts in denatured proteins: resonance assignments for denatured ubiquitin and comparisons with other denatured proteins. AB - Chemical shift assignment is reported for the protein ubiquitin denatured in 8M urea at pH 2. The variations in 15N chemical shifts of three different proteins (ubiquitin, disulfide reduced, carboxymethylated lysozyme, all-Ala-alpha lactalbumin), all without disulfides and denatured in 8M urea at pH 2 are compared to 'random coil shifts' of small model peptides (Braun et al., 1994) and to the averaged native chemical shifts taken from the BMRB database. Both parameterizations show a remarkable agreement with the averaged measured 15N chemical shifts in the three denatured proteins. Detailed analysis of these experimental 15N chemical shifts provides an estimate of the influence of nearest neighbors and conformational preferences on the chemical shift and provides a direct means to identify non-random structural preferences in denatured proteins. PMID- 11256813 TI - A graphical method for the analysis of anisotropic rotational diffusion in proteins. AB - A graphical method for the analysis of relaxation data is presented. It allows a fast estimation of the range of values of the components of the axially symmetric rotational diffusion tensor that are compatible with the experimental relaxation data. The graphical method clearly shows the contribution of different experimental relaxation parameters to the measured anisotropy. In particular, for proteins with moderate anisotropy, data from at least two N-H bonds forming angles close to 0 degrees and 90 degrees with respect to the principal axis of the rotational diffusional tensor are needed. For very anisotropic systems, combination of different relaxation parameters from a single residue is enough to characterize the local anisotropy. PMID- 11256812 TI - 2D relayed anisotropy correlation NMR: characterization of the 13C' chemical shift tensor orientation in the peptide plane of the dipeptide AibAib. AB - An approach to the determination of the orientation of the carbonyl chemical shift (CS) tensor in a 13C-15N-1H dipolar coupled spin network is proposed. The method involves the measurement of the Euler angles of the 13C'-15N and 15N-1H dipolar vectors in the 13C' CS tensor principal axes system, respectively, via a 13C-15N REDOR experiment and by a 2D relayed anisotropy correlation of the 13C' CSA (omega2) and 15N-1H dipolar interaction (omega1). Via numerical simulations the sensitivity of the omega1 cross sections of the 2D spectrum to the Euler angles of the 15N-1H bond vector in the 13C' CSA frame is shown. Employing the procedure outlined in this work, we have determined the orientation of the 13C' CS tensor in the peptide plane of the dipeptide AibAib-NH2 (Aib = alpha aminoisobutyric acid). The Euler angles are found to be (chiCN, psiCN) = (34 degrees +/- 2 degrees, 88 degrees +/- 2 degrees) and (chiNH, psiNH) = (90 degrees +/- 10 degrees, 80 degrees +/- 10 degrees). From the measured Euler angles it is seen that the sigma33 and sigma22 components of the 13C' CS tensor approximately lie in the peptide plane. PMID- 11256814 TI - Complete 1H, 13C and 15N backbone assignments for the hepatitis A virus 3C protease. PMID- 11256815 TI - 1H, 15N and 13C resonance assignments of the N-terminal region of calponin. PMID- 11256816 TI - 1H, 13C and 15N resonance assignments and secondary structure of the c-Myc binding domain (MBD) and the SH3 domain of the tumor suppressor Bin1. PMID- 11256817 TI - Sequence-specific 1H, 13C and 15N resonance assignments of lymphocyte specific kinase unique and SH3 domains. PMID- 11256818 TI - 1H, 13C and 15N resonance assignments of the ERK2 binding domain of the MAPK phosphatase MKP-3. PMID- 11256820 TI - Research in sport and exercise science: the cutting edge. PMID- 11256819 TI - Sequential assignment and secondary structure of the triple-labelled carbohydrate binding domain of papG from uropathogenic E. coli. PMID- 11256821 TI - Electromyographic analysis of repeated bouts of eccentric exercise. AB - The repeated bout effect refers to the protective effect provided by a single bout of eccentric exercise against muscle damage from a similar subsequent bout. The aim of this study was to determine if the repeated bout was associated with an increase in motor unit activation relative to force production, an increased recruitment of slow-twitch motor units or increased motor unit synchronization. Surface electromyographic (EMG) signals were recorded from the hamstring muscles during two bouts of submaximal isokinetic (2.6 rad x s(-1)) eccentric (11 men, 9 women) or concentric (6 men, 4 women) contractions separated by 2 weeks. The EMG per unit torque and median frequency were analysed. The initial bout of eccentric exercise resulted in strength loss, pain and muscle tenderness, while the repeated eccentric bout resulted in a slight increase in strength, no pain and no muscle tenderness (bout x time effects, P < 0.05). Strength, pain and tenderness were unaffected by either bout of concentric exercise. The EMG per unit torque and median frequency were not different between the initial and repeated bouts of eccentric exercise. The EMG per unit torque and median frequency increased during both bouts of eccentric exercise (P < 0.01) but did not change during either concentric bout. In conclusion, there was no evidence that the repeated bout effect was due to a neural adaptation. PMID- 11256822 TI - Impact forces and neck muscle activity in heading by collegiate female soccer players. AB - Three soccer header types (shooting, clearing and passing) and two heading approaches (standing and jumping) were manipulated to quantify impact forces and neck muscle activity in elite female soccer players. The 15 participants were Division I intercollegiate soccer players. Impact forces were measured by a 15 sensor pressure array secured on the forehead. The electromyographic (EMG) activity of the left and right sternocleidomastoid and trapezius muscles was recorded using surface electrodes. Maximum impact forces and impulses as well as the EMG data were analysed with separate repeated-measures analyses of variance. Impact forces and impulses did not differ among the header types or approaches. Higher values were found for jumping versus standing headers in the mean normalized EMG for the right sternocleidomastoid. In addition, the integrated EMG was greater for the right sternocleidomastoid and right and left trapezius (P < 0.05). The sternocleidomastoid became active earlier than the trapezius and showed greater activity before ball contact. The trapezius became active just before ball contact and showed greater activity after ball contact. The increased muscle activity observed in the neck during the jumping approach appears to stabilize the connection between the head and body, thereby increasing the stability of the head-neck complex. PMID- 11256823 TI - Science and Gaelic football: a review. AB - This review focuses on Gaelic football and scientific reports of the characteristics of its players and the demands of the game. Anthropometric characteristics vary according to positional role, but top players display high muscularity and good all-round fitness at the peak of the competitive season. Match analysis indicates that exercise intensity is roughly equivalent to that for professional soccer. Average heart rates are approximately 160 beats x min( 1) during competitive matches, and average oxygen consumption is about 72% of maximum. Metabolic fatigue in active muscles is unlikely. Conventional biomechanical and electromyographic techniques are useful in gaining insight into individual games skills. Inadequate attention has been given to injury prevention and to psychological aspects of the game. Although possessing unique characteristics, Gaelic football has many similarities with other football codes, especially Australian Rules football where the ball is played by both hands and feet and where tackling is permitted. PMID- 11256824 TI - Anthropometric characteristics of elite female junior rowers. AB - During the 1997 Federation Internationale des Societes d'Aviron (FISA) World Junior Rowing Championships, the anthropometric characteristics of 245 female junior rowers aged 17.5 +/- 0.8 years (mean +/- s) were assessed. Twenty-seven body dimensions (body mass, 6 heights or lengths, 4 breadths, 10 girths and 6 skinfolds) were measured in total. The elite female junior rowers were taller (174.5 +/- 6.2 cm) and heavier (69.5 +/- 6.2 kg), with greater length, breadth and girth dimensions, but lower skinfold thicknesses than a representative sample of Flemish (Belgian) girls of the same chronological age. An anthropometric profile chart was constructed that was rowing-specific and norms were established. Compared with scullers, sweep rowers were heavier (+4.2 kg) and taller (+2.8 cm), with greater length, breadth (except for femur width) and girth dimensions (except for calf girth). Sweep rowers also had greater skinfold thicknesses (except for the thigh and calf skinfolds). Finalists were heavier (+3.6 kg) and taller (+3.9 cm), with greater length, breadth (except for femur width) and girth dimensions (except for calf girth) than non-finalists. No significant differences were found for skinfold thicknesses between finalists and non-finalists. PMID- 11256825 TI - Do generalized visual training programmes for sport really work? An experimental investigation. AB - We assessed the effectiveness of two generalized visual training programmes in enhancing visual and motor performance for racquet sports. Forty young participants were assigned equally to groups undertaking visual training using Revien and Gabor's Sports Vision programme (Group 1), visual training using Revien's Eyerobics (Group 2), a placebo condition involving reading (Group 3) and a control condition involving physical practice only (Group 4). Measures of basic visual function and of sport-specific motor performance were obtained from all participants before and immediately after a 4-week training period. Significant pre- to post-training differences were evident on some of the measures; however, these were not group-dependent. Contrary to the claims made by proponents of generalized visual training, we found no evidence that the visual training programmes led to improvements in either vision or motor performance above and beyond those resulting simply from test familiarity. PMID- 11256826 TI - Reliability of a 5-m multiple shuttle test. AB - The aim of the present study was to determine the reliability of a modified 5-m multiple shuttle test. The 'match-related fitness' of 23 female hockey players was assessed on four occasions within 4 weeks. The results of each test session and each shuttle were analysed using analysis of variance with repeated measures to determine the reliability of the test. The mean distance for each of the six shuttles decreased (121.2 +/- 7.5, 114.5 +/- 7.5, 112.2 +/- 7.5, 109.9 +/- 7.9, 108.4 +/- 8.1 and 108.7 +/- 8.3 m for shuttles 1-6, respectively; P < 0.001) similarly for each of the four sessions (P = 0.99). The total and peak distances covered during the tests were not significantly different (P = 0.99 and P = 0.12, respectively). The intra-class correlation coefficient (R) for these variables was 0.98 and 0.86, respectively. The delta distance and the fatigue index calculated post-test were significantly different (P = 0.001 and P = 0.006, respectively) between the four sessions. The intra-class correlation coefficient for both these variables was 0.74. Heart rate and rating of perceived exertion (RPE) were not significantly different between sessions (P = 0.42 and P = 0.095, respectively). The intra-class correlation coefficient for heart rate ranged from 0.65 to 0.97 and that for RPE from 0.85 to 0.91. We conclude that the 5-m multiple shuttle run test is a reliable measure of total and peak distances, heart rate and RPE response and is sufficiently reliabile to track changes in fitness over a season. The delta distance and fatigue index are not as reliable and should be interpreted with caution. PMID- 11256827 TI - Deposition and clearance of particles in the human lung. PMID- 11256828 TI - Experimental studies on the effects of diesel exhaust emissions. PMID- 11256830 TI - Human experimental studies on the effects of inhaled particles. PMID- 11256829 TI - Experimental studies on the effects of ultrafine particles. PMID- 11256831 TI - Epidemiologic studies on the health effects of ambient particulate air pollution. PMID- 11256832 TI - Particles in the ambient atmosphere. PMID- 11256833 TI - Health risk evaluation of particles in ambient air. PMID- 11256834 TI - Comparison of intraoperative frozen section analysis of sentinel node with preoperative positron emission tomography in the diagnosis of axillary lymph node status in breast cancer patients. AB - BACKGROUND: Although axillary lymph node status is an important prognostic factor and axillary dissection is regarded as the gold standard for staging, it requires radical surgery which is accompanied by considerable postoperative problems such as lymphedema. This study was carried out to evaluate the diagnostic accuracy of preoperative positron emission tomography (PET) and intraoperative frozen biopsy of sentinel lymphadenectomy (SLND) in detecting axillary lymph node metastasis. METHODS: We studied 18 patients who had preoperative PET and SLND for breast cancer in the Department of Surgery at Samsung Medical Center. They all had preoperative PET with a radiolabeled glucose analogue ([18F]FDG) to visualize primary tumors and metastatic nodes. Isosulphan blue dye was used for intraoperative SLND. Frozen and permanent biopsies were then compared after full axillary dissection. RESULTS: In 18 cases, six had positive metastatic nodes in the permanent biopsy of full axillary dissection but were negative in three cases by preoperative PET. There was one false negative result by frozen biopsy of SLND which was later shown to be positive by permanent biopsy. The sensitivity and specificity of SLND and PET for detecting axillary node metastasis were 83, 100% and 50, 100%, respectively. CONCLUSION: Although both methods are good for axillary nodal status, the intraoperative frozen biopsy result of SLND was superior to preoperative PET in our preliminary study. PMID- 11256835 TI - Serum immunosuppressive acidic protein as a potent prognostic factor for patients with metastatic renal cell carcinoma. AB - BACKGROUND: Estimation of survival probability of individual patients with metastatic renal cell carcinoma was difficult owing to diverse prognostic factors. We analyzed serum immunosuppressive acidic protein (IAP) levels and the cutoff value, then tested its validity for assessing patients' prognoses. METHODS: Serum IAP was measured longitudinally in 84 patients with metastatic disease. Before therapy, cutoff levels of IAP were tested every 20 microg/ml between 600 and 1200 microg/ml. The prognostic importance of IAP and its cutoff level was estimated. RESULTS: The cutoff level of IAP was set at 800 microg/ml for 40 patients who had metastatic disease with the primary tumor in situ and for 44 patients with recurrent disease. IAP was found to be a significant prognostic factor for both patient groups. CONCLUSIONS: Serum IAP is an important prognostic factor for patients with metastatic renal cell carcinoma. Stratification of patients according to prognosis is feasible using the cutoff level. PMID- 11256836 TI - Prospective and randomized comparison of combined androgen blockade versus combination with oral UFT as an initial treatment for prostate cancer. AB - OBJECTIVE: This prospective and randomized clinical study was initiated to compare the efficacy and safety of combined androgen blockade with combination with UFT in patients with untreated prostate cancer. METHODS: A total of 142 patients were entered in this study between April 1990 and December 1992. All patients received bilateral orchiectomy and 200 mg/day of diethylstilbestrol diphosphate. Of these patients, 70 patients were administered an additional 400 mg/day of UFT after randomization. Either treatment was continued for at least 1 year or until objective progression occurred if the initial response was equal to or better than no change. The endpoints of this study were progression-free survival, cancer-specific survival and change of QOL scores. RESULTS: A total of 136 patients were evaluable and 131 patients (96.3%) could be followed up with a median follow-up period of 1469 days. Both groups showed similar initial treatment response at 12 weeks, adverse effect and change of quality of life score during the first year after initiation of the treatment. There was a significantly longer progression-free survival and better but not significant cancer-specific survival in the endocrine chemotherapy group. The patients with earlier stage and initial serum prostate-specific antigen values <40 ng/ml showed a good indication for this endocrine chemotherapy. CONCLUSION: This endocrine chemotherapy was confirmed to be tolerable and significantly effective in the delay of disease progression, which leads to longer survival in patients with prostate cancer. PMID- 11256837 TI - Intravenous 5-fluorouracil versus oral doxifluridine as preoperative concurrent chemoradiation for locally advanced rectal cancer: prospective randomized trials. AB - BACKGROUND: Preoperative radiation treatment with concomitant intravenous infusion of 5-fluorouracil (5-FU) is known to be effective in shrinking and downstaging of tumors. However, chemotherapy has often been limited by its toxicity and poor patient compliance. Oral 5-FU is known to have several advantages over conventional intravenous 5-FU infusion such as lower toxicity and higher quality of life without compromising the efficacy of the treatment. The aim of this study was to compare intravenous 5-FU with oral doxifluridine with respect to tumor response, toxicity and quality of life. METHODS: Twenty-eight patients with rectal cancer, staged as over T3N1 or T4 by transrectal ultrasonography between July 1997 and December 1998, were included in this study. Intravenous 5-FU (450 mg/m2) and leucovorin (20 mg/m2) were given for five consecutive days during the first and fifth weeks of radiation therapy (50.4 Gy) (n = 14). Oral doxifluridine (700 mg/m2/day) and leucovorin (20 mg/m2) were given daily during radiation treatment (n = 14). Quality of life was scored according to 22 activity items (good, >77; fair, >58; poor, <57). Surgical resection was performed 4 weeks after completion of concurrent chemoradiation treatment. Tumor response was classified into CR (complete remission), PR (partial response; 50% diminution of tumor volume or downstaging ) and NR (no response). RESULTS: Tumor response was CR 3/14 (21.4%), PR 7/14 (50%) and NR 4/14 (28.6%) in the IV arm versus CR 2/14 (14.2%), PR 6/14 (42.9%) and NR 6/14 (42.9%) in the Oral arm (p = 0.16, 0.23, 0.24), respectively. The quality of life was poor (36.4% versus 33.3%), fair and good (63.6% versus 66.7%) between the IV arm and Oral arm, respectively. Gastrointestinal toxicity was 2/14 (14.3%) in the IV arm versus 5/14 (35.7%) in the Oral arm, respectively. Stomatitis was only observed in the IV arm (1/14, 7.1%). Hematological toxicity was 3/14 (21.4%) in the IV arm versus 4/14 (28.5%) in the Oral arm, respectively. Systemic recurrence during the follow up periods were 1/14 (7.1%) in the IV arm and 2/14 (14.3%) in the Oral arm, respectively (p = 0.307). One local recurrence was observed in the Oral arm. CONCLUSION: Even though the results were not entirely reliable owing to the small number of patients enrolled, oral doxifluridine-based chemotherapy as preoperative chemoradiation for advanced rectal cancer did not show any significant advantages over intravenous infusion. PMID- 11256838 TI - Possible clinical benefits of the use of peripheral blood stem cells over bone marrow in the allogeneic transplantation setting for the treatment of childhood leukemia. AB - BACKGROUND: The benefits of allogeneic peripheral blood stem/progenitor cell transplantation (PBSCT) over bone marrow transplantation (BMT), if any, have not been seriously evaluated in a pediatric population. We report here our experience with this procedure and demonstrate rapid engraftment to reduce procedure-related complications and enhanced allogeneic immune reaction to reduce leukemic relapse. METHODS: The feasibility of PBSCT was reviewed retrospectively. Four patients (2 AML and 2 ALL, aged 8-18 years) underwent allogeneic PBSCT for relapsed leukemia after primary allogeneic BMT (n = 2), for active hepatosplenic fungal abscess (n = 1) or for refractory relapse with conventional chemotherapy (n = 1). Four healthy donors (aged 10-49 years) received granulocyte colony-stimulating factor (G-CSF) 10 microg/kg/day by subcutaneous injection for 5 days. An individualized cytoreductive regimen was used before transplantation. RESULTS: No significant toxicities were observed in normal donors on G-CSF treatment or at collection of PBSC. After PBSCT, no significant acute toxicities were observed and the median duration to an absolute granulocyte count of 0.5 x 10(9)/l and a platelet count of 20 x 10(9)/l was 16 and 21 days, respectively. Although none of our patients developed acute graft-versus-host disease (GVHD), two developed chronic GVHD involving the liver and skin. Among those who developed chronic GVHD, one died of recurrent disease and another died of pneumonia 235 days after PBSCT. The two remaining patients have been alive without evidence of disease with follow-ups of 193 and 123 days, respectively. CONCLUSIONS: Allogeneic PBSCT can be a safe procedure in a pediatric population with fewer acute complications, although the potential risk of G-CSF treatment in normal donors should be seriously weighed against the existing risks of marrow aspiration under general anesthesia. The risk of chronic GVHD may need to be balanced against a possible graft-versus leukemia benefit in patients at higher risk of leukemic relapse. PMID- 11256839 TI - A 7 mm lung adenocarcinoma with mediastinal involvement and lymphangiosis carcinomatosa: a case report. AB - We report a case of a 46-year-old man with a 7 mm lung adenocarcinoma with mediastinal nodal involvement and lymphangiosis carcinomatosa. The resected right middle lobe contained a 7 mm well-differentiated papillary adenocarcinoma and lymphatic vessels towards the hilum were severely involved. The disease was pathologically diagnosed as T1N2M0. Six months after the operation, malignant pleural effusion and multiple bone metastases developed and he died 21 months after the operation. This case indicates that even a very small-sized lung cancer, 1 cm or smaller, could be biologically highly malignant. PMID- 11256840 TI - Uterine inversion caused by uterine sarcoma: a case report. AB - Uterine inversion caused by uterine sarcoma is a rare condition with 12 reported cases to date according to a MEDLINE search. We report two cases of this rare condition. A 71- and a 72-year-old woman presented with uterine sarcomas rapidly extruded into the vagina. In both cases, magnetic resonance imaging (MRI) scans showed U-shaped uterine cavities and the pedicles of these tumors were attached to the uterine fundi. Pathological examination confirmed a leiomyosarcoma and a heterologous carcinosarcoma. Uterine inversion can occur when uterine sarcoma rapidly increases in size and extrudes into the vagina. MRI should be performed in the diagnosis of this rare combination. PMID- 11256841 TI - Delayed recovery of neutrophil counts after peripheral stem cell transplantation which improved with administration of a minimal dose of G-CSF: a case report. AB - G-CSF administration may not affect the rate of engraftment unless sufficient numbers of stem cells are transplanted. However, there might be some cases in which neutrophil recovery is delayed after stem cell transplantation despite a relatively fast recovery of platelet counts and total white blood cell counts. This delayed engraftment might be observed in either autologous or allogeneic stem cell transplantation. Here we report four cases with delayed neutrophil engraftment which subsequently improved with administration of a minimal dose of G-CSF. As the response occurred on the day following G-CSF administration, a mobilization defect is suspected to be the mechanism of this delayed engraftment. PMID- 11256842 TI - Esophageal cancer mortality rates by prefectures in Japan. PMID- 11256843 TI - Phase I trial of gemcitabine in patients with advanced pancreatic cancer. AB - BACKGROUND: Gemcitabine is the most promising new agent currently being tested in pancreatic cancer. The present study was conducted to confirm the tolerability of a weekly schedule of gemcitabine at a dose of 1000 mg/m2 in Japanese patients with advanced pancreatic cancer. METHODS: The primary end-point was to evaluate the frequency of dose-limiting toxicity. Gemcitabine 1000 mg/m2 was administered over 30 min weekly in two schedules: gemcitabine x3 every 4 weeks (Schedule 1) and gemcitabine x7 followed by a week of rest and then gemcitabine x3 every 4 weeks thereafter (Schedule 2). At least three patients entered each schedule and three additional patients were treated in the presence of dose-limiting toxicity. RESULTS: Eleven chemo-naive patients with a good Karnofsky performance status of > or =80 points and distant metastasis were entered into this trial. In Schedule 1, no dose-limiting toxicity was observed in the three patients. In Schedule 2, the evaluation of dose-limiting toxicity was complete in six of the eight enrolled patients and two patients showed dose-limiting toxicity in this Schedule; one patient experienced both grade 4 leukocytopenia and grade 4 neutropenia, and both grade 4 neutropenia and grade 3 GOT/GPT increased in another patient. Two patients (18%) showed a partial response and a clinical benefit response was also achieved in two (29%) of the seven evaluable patients. CONCLUSION: Gemcitabine 1000 mg/m2 weekly x7 followed by a week of rest and weekly x3 every 4 weeks thereafter may be tolerated in Japanese patients with advanced pancreatic cancer. PMID- 11256844 TI - Statin therapy: where are we? Where do we go next? AB - Statin therapy reduces coronary artery disease morbidity and mortality in primary and secondary prevention trials including patients with elevated and average cholesterol levels. The association between reduction of total or low-density lipoprotein cholesterol and preventive benefit is well established. However, additional risk factors for coronary artery disease need to be incorporated into risk assessment to provide an accurate measure of global risk for use in lifestyle intervention and drug therapy guidelines. Assessment of outcomes in the Air Force/Texas Coronary Atherosclerosis Prevention Study primary prevention trial, which involved patients with average cholesterol levels and reduced high density lipoprotein cholesterol (HDL-C), suggests the importance of on-treatment values of apolipoproteins B and A-I in predicting first major events in such a population. Other data, including trials of fibrate therapy showing reduction in coronary artery disease events, support the importance of triglycerides and HDL-C in coronary artery disease risk. Challenges for future treatment guidelines include incorporation of emerging and novel risk factors into risk assessment, refinement of global risk measurement, and simplification for application to clinical practice. PMID- 11256845 TI - Economics of lipid lowering in primary prevention: lessons from the West of Scotland Coronary Prevention Study. AB - On the basis of the clinical benefit and virtual absence of adverse effects observed with statin treatment in major primary and secondary prevention trials, a case could be made for considering statin therapy for all patients meeting the eligibility criteria for these trials. Establishing a risk threshold for statin treatment based on cost is an urgent item on the agenda of modern healthcare systems. An economic analysis using West of Scotland Coronary Prevention Study (WOSCOPS) findings indicates that statin treatment would have prevented 318 events per 10,000 patients in a population similar to that in WOSCOPS (average 1.5% annual risk of a cardiovascular event) at a discounted cost per life-year gained of 20,375 pounds($31,818). Application of current European treatment guidelines to consider treatment in patients with risk >2% per year would result in treatment of approximately 40% of the WOSCOPS population; discounted cost per life-year gained in this case would be 13,995 pounds($21,855). In Scotland, an annual risk threshold of > or = 3% for treatment has been adopted, corresponding to only 8% of the WOSCOPS population; the discounted cost per life-year gained is 9,680 pounds ($15,116). Reductions in drug cost, improved compliance, and use of more potent statins could alter cost efficiencies and encourage use of statins in a greater proportion of the primary prevention population that has been shown to benefit from such treatment. PMID- 11256846 TI - Novel approaches to lipid lowering: what is on the horizon? AB - New approaches to lipid lowering include new uses of proven treatments and development of novel agents. Several large-scale clinical trials are assessing whether additional reduction of low-density lipoprotein cholesterol (LDL-C) levels with statin therapy results in additional benefit in coronary artery disease prevention. Statins with increased LDL-C-lowering potency, such as the new statin rosuvastatin (formerly known as ZD4522), have been developed and are in advanced-phase clinical trials. New cholesterol transport inhibitors, such as ezetimibe, have been found to produce significant reductions in intestinal cholesterol absorption, and new bile acid transport inhibitors are in development. Inhibitors of acyl coenzyme A:cholesterol acyltransferase, which can reduce cholesterol storage in macrophages and thereby in arterial lesions, have also been developed, with the agent avasimibe currently being evaluated in phase 2/3 trials. Combination approaches hold considerable promise, including combined use of statins with fibrates, niacin, and the new sterol absorption inhibitors. PMID- 11256847 TI - Preclinical and clinical pharmacology of Rosuvastatin, a new 3-hydroxy-3 methylglutaryl coenzyme A reductase inhibitor. AB - Rosuvastatin (formerly ZD4522) is a new 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor (statin) with distinct pharmacologic properties. Compared with most other statins, it is relatively hydrophilic, similar in this respect to pravastatin. Rosuvastatin has been shown to be a comparatively potent inhibitor of HMG-CoA reductase activity in a purified preparation of the catalytic domain of the human enzyme, as well as in rat and human hepatic microsomes. In rat hepatocytes, rosuvastatin was found to have significantly higher potency as an inhibitor of cholesterol synthesis than 5 other statins. Rosuvastatin was approximately 1,000-fold more potent in rat hepatocytes than in rat fibroblasts. Further studies in rat hepatocytes demonstrated that rosuvastatin is taken up into these cells by a high-affinity active uptake process. Rosuvastatin was also taken up selectively into the liver after intravenous administration in rats. Potent and prolonged HMG-CoA reductase inhibitory activity has been demonstrated after oral administration to rats and dogs. Pharmacokinetic studies in humans using oral doses of 5 to 80 mg showed that maximum plasma concentrations and areas under the concentration-time curve are approximately linear with dose. The terminal half-life is approximately 20 hours. Studies with human hepatic microsomes and human hepatocytes have suggested little or no metabolism via the cytochrome P-450 3A4 isoenzyme. On the basis of these observations, it is suggested that rosuvastatin has the potential to exert a profound effect on atherogenic lipoproteins. PMID- 11256848 TI - Pro and con: high-density lipoprotein, triglycerides, and other lipid subfractions are the future of lipid management. AB - Risk assessment limited to low-density lipoprotein cholesterol (LDL-C) levels fails to identify a substantial proportion of individuals at risk for coronary artery disease. Data from the Prospective Cardiovascular Munster (PRO-CAM) study indicate that triglyceride levels and high-density lipoprotein cholesterol levels are important determinants of risk irrespective of LDL-C level. Risk is most accurately predicted using an algorithm incorporating multiple risk factors, as shown by the multiple logistic function analysis using the independent risk variables identified on multivariate analysis in PROCAM. It remains unclear what effect statin therapy to decrease LDL-C has in individuals with risk associated with factors other than elevated LDL-C. Such individuals might derive preferential benefit from treatments targeting such risk factors. PMID- 11256849 TI - Statin therapy and reductions in low-density lipoprotein cholesterol: initial clinical data on the potent new statin Rosuvastatin. AB - The utility of statins with increased potency in reducing low-density lipoprotein cholesterol (LDL-C) is indicated by evidence that aggressive LDL-C lowering is associated with increased reduction in coronary artery disease risk, and the need for such agents is illustrated by the fact that many patients currently fail to achieve LDL-C target levels during treatment with available drugs. In dose ranging studies of patients with hypercholesterolemia, the new synthetic statin rosuvastatin (formerly ZD4522) produced significant, dose-dependent reductions in LDL-C compared with placebo across a range of doses. Reductions ranged from 34% at 1 mg per day to 65% at 80 mg per day, with linear regression analysis indicating an additional 4.5% reduction in LDL-C with each doubling of the rosuvastatin dose. Rosuvastatin treatment was well tolerated. Phase 3 clinical trials of this agent are under way. PMID- 11256850 TI - Pro and con: low-density lipoprotein cholesterol lowering is and will be the key to the future of lipid management. AB - A wealth of data demonstrate that reduction of cholesterol levels is associated with benefit in reducing coronary artery disease risk. The magnitude of benefit observed with statin treatment, which acts primarily to reduce low-density lipoprotein cholesterol (LDL-C), is greater than that observed with any other lipid-modifying intervention, and data from large statin trials indicate that this benefit is caused by LDL-C reduction. Statin treatment is highly cost effective compared with other accepted therapies, at least in the secondary prevention setting, and has a superior safety and tolerability profile. For the foreseeable future, LDL-C reduction will remain the goal of lipid-modifying therapy, and statins will remain the primary therapeutic modality for achieving that goal. PMID- 11256851 TI - Short-term effects of formaldehyde on peak expiratory flow and irritant symptoms. AB - The authors studied the respiratory effects of formaldehyde exposure among students who dissected cadavers in a gross anatomy laboratory. Peak expiratory flow and respiratory symptoms were measured before and after each weekly laboratory session. Each of 38 students was exposed to formaldehyde for 2.5 hr/wk for 14 wk. Individual, daily formaldehyde measurements averaged 1.1 ppm (standard deviation = 0.56 ppm). Multivariate models demonstrated two different time scales of effect of formaldehyde on peak expiratory flow: (1) exposure during the previous 2.5 hr reduced peak expiratory flow by -1.0% per ppm, and (2) average exposure during all preceding weeks reduced peak expiratory flow by an additional -0.5% per ppm of formaldehyde. However, the short-term exposure effect was diminished during the first 4 wk, suggesting at least partial acclimatization. Symptom reporting was also associated with exposure during the previous 2.5 hr, and similar evidence of acclimatization was observed. These results suggest that there are two different time scales of response to formaldehyde, and they emphasize the need for longitudinal studies, characterized by quantitative exposure characterization, and frequent measurements of outcome. PMID- 11256852 TI - Urinary 2-methoxy acetic acid accumulation in response to 2-methoxy ethanol exposure. AB - Urinary 2-methoxy acetic acid reportedly has a long half-life (77.1 hr) in humans. The authors studied urinary 2-methoxy acetic acid in a group of 18 workers exposed to 2-methoxy ethanol from Monday to Saturday following a 7-d cease in production. The weekly time-weighted average exposure concentration of 2 methoxy ethanol was 4.5 ppm. The urinary 2-methoxy acetic acid of exposed workers was increased significantly, from 18.5 microg/ml (10.6 mg/gm creatinine) on Monday (prior to work) to 48.4 microg/ml (46.5 mg/gm creatinine) on Friday (after work), to 51.2 microg/ml (45.6 mg/gm creatinine) on Saturday after work. These levels occurred, despite that fact that the daily mean time-weighted average 2 methoxy ethanol exposures were very consistent and were close to the current Taiwan Permissible Exposure Limit of 5 ppm. These urinary 2-methoxy acetic acid levels were much higher than levels that occur with inhalation only, and they demonstrate that skin absorption is a significant contributor to 2-methoxy ethanol exposure. The high background concentrations of 2-methoxy acetic acid in the preshift urine samples following a 7-d production halt confirm that there is a long half-life of 2-methoxy acetic acid in humans. The determination of urinary 2-methoxy acetic acid is recommended for exposure assessment of 2-methoxy ethanol. PMID- 11256853 TI - Maternal and umbilical cord blood lead levels: an Istanbul study. AB - Current mean lead concentrations in umbilical cords and in maternal blood in Istanbul were 1.69 +/- 0.91 (standard deviation) microg/dl and 2.37 +/- 0.89 microg/dl, respectively. These levels were much lower than values reported in previous years. Umbilical cord blood lead levels, which correlated significantly with mothers' blood lead levels, were approximately 70% of the latter. The drastic decrease in blood lead levels likely reflects the reduction in use of tetraalkyl lead in gasoline subsequent to 1989. PMID- 11256854 TI - Nasal lavage biomarkers: effects of water damage and microbial growth in an office building. AB - Selected nasal symptoms were studied in personnel who worked in a damp office building that had microbial growth (including Stachybotrys sp.) in mineral fiber insulation and gypsum board. There were also signs of dampness in the floor. Clinical examinations included nasal lavage and peak expiratory flow measurements in 12 subjects in the damp building; an additional 8 subjects in a control building (i.e., no signs of dampness or microbial growth) were also examined. Hygienic air measurements of microorganisms and volatile organic compounds were performed in both buildings. The concentrations of eosinophil cationic protein, myeloperoxidase, and albumin, and the number of subjects with eosinophils in lavage fluid, were higher among office workers in the damp building than among controls. The damp biiilding had greater amounts of total molds and bacteria in its construction than the building materials in nondamp buildings. In addition, an increase of 2-ethyl-1-hexanol in the indoor air was detected in the damp building-a sign of dampness-related alkaline degradation of diethyl-hexyl phthalate in polyvinyl chloride floor coatings. In conclusion, the results of this study indicate that exposures in a damp office building may cause an inflammatory nasal mucosal response. The results also support conclusions of earlier studies, indicating that building dampness is related to respiratory inflammation. PMID- 11256855 TI - Associations between outdoor air pollution and hospital admissions in Brisbane, Australia. AB - The authors investigated the effects of ambient air pollution on hospital admissions in Brisbane, Australia. The authors used the Air Pollution on Health: European Approach protocol to examine the effects of particles, ozone, sulfur dioxide, and nitrogen dioxide on daily hospital admissions for asthma and respiratory, cardiovascular, and digestive disorders (control diagnosis) that occurred during the period 1987-1994. Ozone was consistently associated with admissions for asthma and respiratory disease-with little evidence of a threshold. In two-pollutant models, the ozone effect was relatively unaffected by the control for high levels of other pollutants. Particulate pollution (measured by nephelometry) was associated positively with admissions for respiratory disease and admissions for asthma in summer, whereas a negative association was observed for cardiovascular admissions. Although sulfur dioxide was associated significantly with admissions for respiratory and cardiovascular disese, a significant association was also found for the control diagnosis of digestive disorders. No significant associations were found for nitrogen dioxide over the study period, although significantly positive seasonal interactions were found for asthma and respiratory disease in autumn, winter, and spring. It was concluded that current levels of ambient air pollution in Brisbane make a significant contribution to the variation in daily hospital admissions for asthma and respiratory disease. PMID- 11256856 TI - Mercury and methylmercury exposure in the New Jersey pregnant population. AB - Methylmercury is a known fetal developmental neurotoxicant. The only significant source of fetal exposure is maternal fish consumption; however, few recent data on exposure of the pregnant population are available. The authors undertook a study of methylmercury exposure in the New Jersey pregnant population to investigate the distribution of exposure and to identify predictors of elevated exposure. Mainly first-trimester pregnant women were recruited through six New Jersey obstetric practices. Hair and blood samples were analyzed for total mercury, and a subset was analyzed for methylmercury. A questionnaire on demographics, life style, and fish-consumption practices was also administered. Although 85-90% of the pregnant population had hair mercury levels that were less than 1.0 microg/gm, 1-2% had levels in a range of possible concern for adverse developmental effects (> 4.0 microg/gm). Regression analysis suggested that blacks and individuals with some college education experienced lower exposures to methylmercury. PMID- 11256857 TI - Effect of sulfur dioxide inhalation on erythrocyte antioxidant status, food intake, and lipid peroxidation during aging. AB - The effect of sulfur dioxide on red blood cell antioxidant status and lipid peroxidation was investigated in young (3 mo), middle-age (12 mo), and old (24 mo) male albino rats. Ten ppm of sulfur dioxide was administered to the rats in the sulfur dioxide groups in an exposure chamber. Exposure occurred 1 hr/d, 7 d/wk, for 6 wk; control rats were exposed to filtered air during the same time periods. Copper-zinc superoxide dismutase, glutathione peroxidase, catalase, glutathione, and glutathione-S-transferase activities were significantly decreased in the middle-aged and older groups, compared with the young group. Sulfur dioxide exposure significantly decreased copper-zinc superoxide dismutase activity in all experimental groups, compared with controls. Sulfur dioxide exposure significantly increased enzyme and glutathione activities. PMID- 11256858 TI - Ambient air pollution and chronic respiratory morbidity in Delhi. AB - The authors conducted a cross-sectional study among residents of Delhi to determine the role of ambient air pollution in chronic respiratory morbidity in Delhi. The authors selected a random, stratified sample (N = 4,171) of permanent residents who were 18+ y of age and who lived near 1 of the 9 permanent air quality monitoring stations in the city. Air-quality data for the past 10 y were obtained; data were based on the differences in total suspended particulates, and the study areas were categorized into lower- and higher-pollution zones. A standardized questionnaire was administered, clinical examination was carried out, and spirometry followed. The authors assessed chronic respiratory morbidity by (a) prevalence of chronic respiratory symptoms (i.e., chronic cough, phlegm, breathlessness, and wheezing) and airway diseases (i.e., chronic obstructive pulmonary disease/chronic bronchitis and bronchial asthma); and (b) lung function results in asymptomatic nonsmoking subjects in the two pollution zones. A multiple logistic regression identified the determinants of chronic symptoms. Smoking, male sex, increasing age, and lower socioeconomic status were strong independent risk factors for occurrence of chronic respiratory symptoms. In the comparison of nonsmoking residents of lower- and higher-pollution zones- stratified according to socioeconomic levels and sex--chronic cough, chronic phlegm, and dyspnea (but not wheezing) were significantly more common in the higher-pollution zone in only some of the strata. Furthermore, prevalence rates of bronchial asthma, chronic obstructive pulmonary disease, and chronic bronchitis among residents in the two pollution zones were not significantly different. Nonetheless, lung function of asymptomatic nonsmokers was consistently and significantly better among both male and female residents of the lower pollution zone. PMID- 11256859 TI - Lung mineral fibers of former miners and millers from Thetford-Mines and asbestos regions: a comparative study of fiber concentration and dimension. AB - Fiber dimension and concentration may vary substantially between two necropsy populations of former chrysotile miners and millers of Thetford-Mines and Asbestos regions. This possibility could explain, at least in part, the higher incidence of respiratory diseases among workers from Thetford-Mines than among workers from the Asbestos region. The authors used a transmission electron microscope, equipped with an x-ray energy-dispersive spectrometer, to analyze lung mineral fibers of 86 subjects from the two mining regions and to classify fiber sizes into three categories. The most consistent difference was the higher concentration of tremolite in lung tissues of workers from Thetford-Mines, compared with workers from the Asbestos region. Amosite and crocidolite were also detected in lung tissues of several workers from the Asbestos region. No consistent and biologically important difference was found for fiber dimension; therefore, fiber dimension does not seem to be a factor that accounts for the difference in incidence of respiratory diseases between the two groups. The greater incidence of respiratory diseases among workers of Thetford-Mines can be explained by the fact that they had greater exposure to fibers than did workers at the Asbestos region. Among the mineral fibers studied, retention of tremolite fibers was most apparent. PMID- 11256860 TI - Association of elevated blood levels of pentachlorophenol (PCP) with cellular and humoral immunodeficiencies. AB - It has long been suspected that pentachlorophenol (PCP) exerts a damaging influence on the immune system. In this study, the possible relationship between blood levels of PCP and immune function was studied in 190 patients who had been exposed for more than 6 mo to PCP-containing pesticides. The patients suffered from frequent respiratory infections and general fatigue. Lymphocyte subpopulations, in-vitro responses to mitogens, allogeneic stimulator cells, plasma neopterin, cytokines, soluble cytokine receptors, soluble adhesion molecules, and immunoglobulin autoantibodies were determined. A dose-response relationship between blood levels of PCP and cellular and humoral immune parameters was established. Blood levels of PCP were associated negatively with (a) total lymphocyte counts (p = .0002), CD4/CD8 ratios (p = .0015), and absolute counts of CD3+ (p < .0001), CD4+ (p < .0001), CD16+ (p < .0001), CD25+ (p = .0003), DR+ (p < .0001), CD8+/56+ (p = .020), and CD19+ cells (p = .092); (b) plasma levels of interleukin-2 (IL-2) (p < .0001), soluble IL-2R (p < .0001), IL 6 (p < .0001), IL-10 (p = .0039), interferon-gamma (IFN-gamma) (p < .0001), tumor necrosis factor-alpha (TNF-alpha) (p < .0001), transforming-growth factor-beta2 (p = .023), soluble IL-1 receptor antagonist (sIL-1 RA) (p < .0001), soluble intercellular adhesion molecule-1 (p = .0003); and (c) immunoglobulin (Ig) M-anti Fab type autoantibodies (p = .0353). PCP levels were associated positively with (a) number of impaired stimulation assays per patient (p = .041); (b) number of circulating CD11b+ monocytes (p = .0015); and (c) plasma levels of neopterin (p < .0001), IL-4 (p = .020), and sIL-6R (p = .020). Compared with patients who had PCP plasma levels that were less than or equal to 10 microg/l, patients with blood levels of PCP that exceeded 10 microg/l experienced the following more often: low numbers of total blood lymphocytes (p = .054), CD3+ (p = .0014), CD4+ (p = .0001), DR+ (p = .0003), CD16+ (p = .0033), and CD25+ cells (p = .0033). In addition, the same aforementioned patients experienced the following more frequently: undetectable plasma levels of IL-2 (p = .0057), IL-6 (p = .042), IL-8 (p = .038), IL-10 (p = .0001), TNF-alpha (p = .0062), and IFN-gamma (p = .016); and impaired in-vitro responses of lymphocytes (p = .071). The authors concluded that increased blood levels of PCP were associated significantly with cellular and humoral immunodeficiencies. Recurrent respiratory infections and general fatigue could originate from PCP-associated immunosuppression. PMID- 11256861 TI - Malignant mesothelioma in Japan: analysis of registered autopsy cases. AB - In the Annual of the Pathological Autopsy Cases in Japan, issued by the Japanese Society of Pathology from 1958 to 1996, a total of 1,846 (0.17%) malignant mesothelioma cases (1,287 male, 558 female, 1 unknown) were registered among 1,056,259 autopsy cases. The frequency of mesothelioma (number of cases/total autopsy cases) was 0.10% (461/440,334) for the term 1958-1979, 0.18% (716/390,124) for 1980-1989, and 0.30% (669/225,801) for 1990-1996; the frequency of cases increased significantly over the time periods (p < .0001). Among 1,785 cases for which tumor sites were ascertained, there were 1,213 pleural mesothelioma (68.0%), 431 peritoneal (24.1%), 108 pericardial (6.1%), 6 tunica vaginalis testis (0.3%), and 28 "others" (1.6%). Histological cell type was noted in 598 cases; 245 (41.0%) were epithelial, 168 (28.1%) were biphasic, and 185 (30.9%) were sarcomatous. Seventy-three (0.007%) cases of malignant mesothelioma with asbestosis were found during the entire 39-y period. The frequency of those with asbestosis (number of cases/total autopsy cases) was 0.001% (5/440,334) for the term 1958-1979, 0.006% (27/390,124) for 1980-1989, and 0.018% (41/225,801) for 1990-1996; this increase over time was statistically significant (p < .0001). Researchers expect that cases of asbestos-related mesothelioma will increase in Japan in the future. Tumor sites and histological cell types of mesothelioma with asbestosis did not differ from those in individuals without asbestosis. PMID- 11256863 TI - Generalization of delayed matching to sample following training at different delays. AB - Four groups of pigeons were trained to perform a delayed matching-to-sample task with a single delay of 0, 2, 4, or 6 s from the outset of training. The longer the training delay, the more sessions were required for all birds to reach the same level of response accuracy. Following initial training, five test sessions that included nonreinforced trials with delay intervals of 0, 2, 4, 6, 8, and 10 s were interspersed between training sessions. Unlike typical forgetting functions in which accuracy decreases monotonically with increasing delay, the forgetting functions from test sessions resembled generalization gradients with the peak of the functions occurring at the training delay. Following additional training for all birds with a 0-s delay, forgetting functions decreased monotonically with increasing delay. The results suggested that remembering can be trained at a specific delay interval, and generalizes to similar delay intervals. Generalization along the temporal dimension of delay may contribute to typical forgetting functions in which accuracy decreases from 0-s delay. PMID- 11256862 TI - Differences in lung function among school children in communities in Israel. AB - Differences in lung functions of school-age children who lived near two electrical power plants in the Ashkelon district of Israel were studied. Lung function tests were performed, and the American Thoracic Society questionnaire was administered in three study periods during the following years: (1) 1990, (2) 1994, and (3) 1997. Measurements of air pollutants (i.e., sulfur dioxide, nitric oxides, ozone) were also taken during the aforementioned study periods. Statistical analysis included an estimation of a series of fixed-effects regression models. A total of 2,455, 1,613, and 4,346 observations were included in the analyses for study years 1990, 1994, and 1997, respectively. The authors controlled for age, sex, height, weight, parents' education and smoking status, and being born out of Israel, and, consequently, substantial differences in lung function across the different communities and study periods were demonstrated in the study area. No robust association with air pollution was demonstrated. The cause of these differences in the respiratory health of children remains unknown. PMID- 11256864 TI - Behavioral momentum in computer-presented discriminations in individuals with severe mental retardation. AB - Behavioral momentum was examined in 2 individuals with severe mental retardation via within-subject manipulations of obtained reinforcer rates. Subjects performed self-paced discrimination problems presented on a touch screen computer monitor. Two different problems, Tasks A and B, alternated in blocks of 15 trials on a multiple schedule. Reinforcers were snack foods. The reinforcement schedule for Task A was continuous (fixed-ratio 1) and the schedule for Task B was continuous in some conditions and variable ratio in other conditions. Behavioral momentum was assessed in test sessions by prefeeding, presenting response-independent food, and making available alternatives to the tasks. When the obtained reinforcer rate for Task A was at least twice that for Task B, resistance to change was greater for Task A. When both reinforcer rates and response rates were a pproximately equal for the two tasks, resistance to change was approximately equal. These results are consistent with behavioral momentum effects. They extend previous findings with humans by examining momentum in self-initiated discrete trial discrimination tasks with ratio schedules, and by isolating relative reinforcer rates as a controlling variable via within-subject manipulations. PMID- 11256865 TI - Sensitivity to relative reinforcer rate in concurrent schedules: independence from relative and absolute reinforcer duration. AB - Twelve pigeons responded on two keys under concurrent variable-interval (VI) schedules. Over several series of conditions, relative and absolute magnitudes of reinforcement were varied. Within each series, relative rate of reinforcement was varied and sensitivity of behavior ratios to reinforcer-rate ratios was assessed. When responding at both alternatives was maintained by equal-sized small reinforcers, sensitivity to variation in reinforcer-rate ratios was the same as when large reinforcers were used. This result was observed when the overall rate of reinforcement was constant over conditions, and also in another series of concurrent schedules in which one schedule was kept constant at VI ached 120 s. Similarly, reinforcer magnitude did not affect the rate at which response allocation approached asymptote within a condition. When reinforcer magnitudes differred between the two responses and reinforcer-rate ratios were varied, sensitivity of behavior allocation was unaffected although response bias favored the schedule that arranged the larger reinforcers. Analysis of absolute response rates ratio sensitivity to reinforcement occurrred on the two keys showed that this invariance of response despite changes in reinforcement interaction that were observed in absolute response rates on the constant VI 120-s schedule. Response rate on the constant VI 120-s schedule was inversely related to reinforcer rate on the varied key and the strength of this relation depended on the relative magnitude of reinforcers arranged on varied key. Independence of sensitivity to reinforcer-rate ratios from relative and absolute reinforcer magnitude is consistent with the relativity and independence assumtions of the matching law. PMID- 11256866 TI - The long-term effect of high- and low-rate responding histories on fixed-interval responding in rats. AB - Tell rats were given extended lever-press training on a fixed-interval (FI) 30-s food reinforcement schedule from the outset or following exposure to one or two previous reinforcement schedules. For 4 rats the previots schedule was either fixed-ratio 20, which generated high response rates, or differential reinforcement-of-low-rate 20 s, which produced low response rates. For 4 additional rats the extended training on FI 30 s was preceded by experience with two schedules: fixed-ratio 20 followed by differential-reinforcement-of-low-rate 20 s; or the same two schedules in the reverse order. Fixed-interval response rates were initially affected by the immediately preceding schedule, but after 80 to 100 sessions, all traces of prior schedule history had disappeared. The results also showed no long-term effect of schedule history on the interfood interval patterns of responding on the FI 30-s schedule. These results support one of the most central tenets of the experimental analysis of behavior: control by the immediate consequences of behavior. PMID- 11256867 TI - Relations among equivalence, naming, and conflicting baseline control. AB - Three studies were conducted with different groups of 6 students each to explore the effects of training class-inconsistent relations and naming on demonstrations of emergent arbitrary stimulus relations. In all studies, two three-member equivalence classes of Greek symbols (A1B1C1 and A2B2C2) emerged as a result of training in conditional discriminations. Two new symbols were introduced (X and Y), and additional conditional discriminations were trained, whereby X was designated as the positive discriminative stimulus (S+) and Y was designated as the negative discriminative stimulus (S-) for A1 and B2. Conversely, Y was designated as the S+ and X as the S- for B1 and A2. This introduced conflicting sources of control within and between classes. In Study 1, subjects were not provided with names for the stimuli. In Study 2, the experimenter provided common names for the stimuli within each class. In Study 3, the subjects were required to use the common names during conditional discrimination training and test-trial blocks. In all experiments, equivalence responding with respect to the original classes was disrupted for some subjects subsequent to learning the new relations. Furthermore, in Studies 2 and 3, there were frequent examples of noncorrespondence between observed (listener or speaker) naming patterns and derived relations. These results support the view that demonstrations of equivalence are subject to control from a variety of sources rather than being fundamentally dependent on naming. PMID- 11256868 TI - Effects of cocaine on fixed-interval responding reinforced by the opportunity to run. AB - Rate-dependent drug effects have been observed for operant responding maintained by food, water, heat, light onset, electrical brain stimulation, shock-stimulus termination, and shock presentation. The present study sought to determine if the effects of cocaine on lever pressing maintained by the opportunity to run could also be described as rate dependent. Seven male Wistar rats were trained to respond on levers for the opportunity to run in a wheel. The schedule of reinforcement was fixed-interval 60 s, and the reinforcing consequence was the opportunity to run for 60 s. On this schedule, overall rates of responding were low, usually below six presses per minute, and pauses frequently exceeded the 60 s interval. Despite these differences, an overall scalloped pattern of lever pressing was evident for each rat. Doses of 1, 2, 4, 8, and 16 mg/kg cocaine were administered 10 min prior to a session. Only at the 16 mg/kg dose did the responding of the majority of rats change in a manner suggestive of a rate dependent drug effect. Specifically, lower response rates at the beginning of the intervals increased and higher rates at the end of the intervals decreased, as indicated by the fact that slopes from the regression of drug rates on control rates decreased. These data provide tentative support for the generalization of rate-dependent effects to operant responding maintained by wheel running. Differences in the baseline performance maintained by wheel running compared to those for food and water point to the need for further experimentation before this effect can be firmly established. PMID- 11256869 TI - Overmatching in rats: the barrier choice paradigm. AB - The barrier choice paradigm was used to impose a cost on rats' behavior of traveling between two levers: Pressing on two levers was reinforced with food on concurrent random-interval schedules, but rats had to climb over a barrier to move from one lever to another. The height of the barrier separating the levers was increased from 30.5 to 45.7 cm across two phases that involved various pairs of random-interval schedules. With the 30.5-cm barrier, the generalized matching law showed slopes equal to or slightly above 1.0 for response and time allocation. With the 45.7-cm barrier, the generalized matching law showed slopes above 1.2 for responses, indicating that sensitivity to reinforcement increased with increasing barrier height. For time allocation the slopes remained close to 1.0; sensitivity to reinforcement did not seem to increase with increasing barrier height. The role of locomotion effort in choice situations is discussed. PMID- 11256870 TI - Implications of the FMR1 gene in menopause: study of 147 Spanish women. AB - OBJECTIVE: To evaluate the relationship between the FMR1 premutation and premature ovarian failure (POF) in the Spanish population and the possible incorporation of this test in gynecological procedures for women with POF or early menopause (EM). DESIGN: Clinical and molecular genetic study. Ninety-eight premutated and six full-mutated carriers of fragile X syndrome and 43 women with POF were studied by polymerase chain reaction and Southern blot analysis for the CGG repeat expansion in the FMR1 gene. RESULTS: Among premutated carriers, 12.2% (12 of 98) presented with POF, and 15.3% (15 of 98) presented with EM. Neither POF nor EM was observed in any of the six full-mutated women. Two women of 43 from the POF population (4.65%) were carriers for the CGG premutation in the FMR1 gene. No correlation between CGG expansion size and age at menopause was found. A biased paternal origin of the premutation and a high twinning incidence was found in all premutated women, whether they had POF or not. CONCLUSIONS: Our data support the hypothesis that the FMR1 gene is one of the genes associated with POF and EM. Analysis of the CGG expansion in the FMR1 gene may be justified in women with POF and EM until the real role of the FMR1 premutation is determined. PMID- 11256871 TI - Factors related to sexual function in postmenopausal women with a history of breast cancer. AB - BACKGROUND: The normal life expectancy of survivors of early-stage breast cancer (BCS) underscores the need to address long-term quality of life issues in these women. Sexual dysfunction persists after breast cancer treatment, despite recovery in other domains. OBJECTIVE: To examine associations between a broad array of characteristics and sexuality in BCS. PARTICIPANTS: Sixty-one postmenopausal BCS who were participants in a randomized, controlled trial of nonhormonal interventions for menopause symptoms and who had a partnered, intimate relationship. METHODS: Cross-sectional analysis of baseline trial data. Outcomes were standardized scales of sexual interest, dysfunction, and satisfaction. Candidate predictors included demographic, anatomical, medical, psychological, sociocultural, and hormonal characteristics. Forward, stepwise regression was used. RESULTS: Relationship quality, vaginal discomfort, education, and hot flashes were each associated with two of the three domains of sexuality assessed. Ten other factors entered predictive models: age, time since diagnosis, breast conservation, comorbidity, urinary incontinence, perceived health, body image, bioavailable testosterone, luteinizing hormone, and sex hormone binding globulin. Each of these 10 factors was associated with only one sexuality domain. CONCLUSIONS: In this small sample of BCS, we found multiple correlates of sexuality. Most seem to impact uniquely on individual domains of sexual function. Several characteristics are modifiable and could be targets for intervention. PMID- 11256872 TI - Concomitant medication use in postmenopausal women using estrogen therapy. AB - OBJECTIVE: To determine whether long-term postmenopausal estrogen therapy is associated with use of other prescription medications. METHODS: Using computer pharmacy records from 1969 to 1973 for members of the Kaiser Permanente Medical Care Program in San Francisco, we identified the 215 most commonly used prescription medications in the pharmacy database and recorded their use by 232 postmenopausal long-term estrogen users and by 222 postmenopausal age-matched nonusers. These medications were grouped into 39 therapeutic classes. Classes of medications used by estrogen users and nonusers were compared. RESULTS: A statistically significant difference in use was seen for 21 of the 39 medication classes; of these 21 classes, 20 (95%) were used more frequently and 1 less frequently by estrogen users. Differences between estrogen users and nonusers were greatest for thyroid hormone preparations (estrogen user/nonuser multivariate odds ratio = 25.6, 95% confidence interval 5.9-112) and antimigraine preparations (11 recipients among estrogen users, none among nonusers). Postmenopausal women using estrogen were more likely than nonusers to use additional medications. CONCLUSION: Greater use of certain prescription medications by estrogen users than by nonusers should be considered in studying the health effects of estrogen replacement therapy. PMID- 11256873 TI - Frail older women's participation in a trial of hormone replacement therapy: perceived benefits and concerns. AB - OBJECTIVE: The aim of this study was to identify the reasons that were important to frail older women's decisions to participate or not participate in a clinical trial of hormone replacement therapy (HRT). DESIGN: We conducted a cross sectional study of a community-based sample of physically frail women > or = 75 years old, who were recruited to participate in an intervention trial of HRT. Participants were randomized 2:1 to either HRT or placebo, respectively. Questionnaires measured participants' reasons for participation and nonparticipants' reasons for declining. Five-point scaled responses to questionnaire items ranged from least to most important or least to most concerned. RESULTS: Sixty-nine women participated (84% white, 16% African American) in the trial. Nonparticipants (n = 41) were older, on average, than participants (83.8+/-4.2 vs. 82.2+/-3.6 years; p = 0.04). Important reasons for participation were reducing risk for Alzheimer's disease and osteoporosis, having more energy, improving self-care ability, and benefiting other women. Fear of cancer from postmenopausal estrogen was the predominant concern of 46% of nonparticipants and 78% of participants (p = 0.08). Recommendation against participation or use of estrogen by a woman's personal physician was the most prevalent additional reason given for nonparticipation. CONCLUSIONS: Disease prevention and improving self-care abilities were most important to participants. Fear of cancer was not a greater concern for nonparticipants than for participants. The role of the physician in older women's decision-making about use of postmenopausal estrogen seems to be important. PMID- 11256874 TI - Effects of hormone replacement on hemostasis in spontaneous menopause. AB - OBJECTIVE: To examine the effects of continuous combined estrogen-progesterone replacement therapy on coagulation and natural anticoagulant systems in spontaneous menopause. DESIGN: A randomized, double-blind, placebo-controlled study was conducted during a 6-month period to examine the effect of hormone replacement therapy (HRT) on blood coagulation parameters. One hundred-ten healthy postmenopausal women were randomized into two groups. Those in group 1 were given conjugated estrogen (0.625 mg/d, Premarin) and medroxyprogesterone acetate (5 mg/d, Farlutal), and those in group 2 were given identical tablets of placebo for 6 months. Serum levels of modified activated protein C resistance, antithrombin III, fibrinogen, factor VIIIa, factor VIII, factor IX, activated partial thromboplastin time, prothrombin time, thrombin time, and lipoprotein (a) were measured before and 6 months after the treatment and analyzed for changes in extrinsic and intrinsic coagulation parameters. RESULTS: At the end of the 6 month period, fibrinogen, lipoprotein (a), and activated protein C resistance levels were decreased significantly in the HRT group compared with the control group. Antithrombin III levels were increased, indicating antithrombin activity. Activated partial thromboplastin time, as a measure for intrinsic coagulation cascade, was prolonged in concert with decreased intrinsic coagulation factors, factor VIII, and factor IX (p < 0.05). In the extrinsic coagulation system, prothrombin time was significantly increased, although factor VIIa level was not changed (p > 0.05). CONCLUSION: Significant changes were observed in the coagulation parameters, which may further explain the cardioprotective effect of HRT. PMID- 11256875 TI - What is the impact of osteoporosis education and bone mineral density testing for postmenopausal women in a managed care setting? AB - OBJECTIVE: To assess whether osteoporosis education, with and without bone mineral density (BMD) testing, increases the initiation of lifestyle changes and pharmaceutical treatment to prevent osteoporosis. DESIGN: A total of 508 women, aged 54-65, from a large managed care organization who were not on osteoporosis prevention therapy participated in an intervention study. Participants were randomly assigned to either an education class on osteoporosis (n = 301) or education plus BMD (n = 207). A control group of 187 women receiving no intervention were also surveyed to serve as comparison. Group differences and differences based on BMD test result were compared 6 months after education regarding self-reported changes in health behaviors using chi2 tests and logistic regression analyses. RESULTS: Of the 508 intervention participants, 455 (90%) responded to the follow-up survey. Initiation of hormone replacement therapy was reported by 9%, with 5% reporting starting alendronate. More than half reported changes in diet, exercise, or calcium intake. Forty-three percent increased their vitamin D intake. There were no significant group differences in behavior except with regard to pharmaceutical therapy; subjects with education plus BMD were three times more likely than those receiving education only to report starting hormone replacement therapy (p = 0.004). Low BMD scores were associated with increasing vitamin D intake (p = 0.03) and starting medication (p = 0.001). Women in the intervention groups were significantly more likely to report modifying their diet (p < 0.001), calcium (p < 0.01), and vitamin D intake (p < 0.0001) than women in the control group, not exposed to education. CONCLUSION: Education regarding osteoporosis prevention seems to encourage women to make lifestyle changes. The inclusion of BMD testing enhances the likelihood that women will consider pharmaceutical therapy. PMID- 11256876 TI - Gynecologic malignancies accompanied by benign hormone-dependent diseases. AB - OBJECTIVE: To investigate the correlation between benign gynecologic diseases and hormone-dependent malignancies such as endometrial carcinoma in postmenopausal women. DESIGN: We retrospectively analyzed the prevalence of myoma uteri and adenomyosis uteri in 136 cases of endometrial carcinomas. We used 222 uterine prolapse cases as controls. RESULTS: The results showed that 21.6% and 9.9% of healthy postmenopausal women (control) had myoma uteri and adenomyosis uteri, respectively, after the cessation of menses. However, postmenopausal women with endometrial carcinomas had a 1.5- to 2-fold higher prevalence, respectively, for myoma uteri and adenomyosis uteri as compared with the postmenopausal control women. CONCLUSION: There was a higher prevalence of myoma uteri and adenomyosis uteri in postmenopausal patients with endometrial carcinomas than in the control population. PMID- 11256877 TI - Antiatherosclerotic effects of tibolone. PMID- 11256878 TI - Should women with premature menopause be screened for FMR-1 mutations? PMID- 11256879 TI - The role of calcium in peri- and postmenopausal women: consensus opinion of The North American Menopause Society. AB - OBJECTIVE: The North American Menopause Society (NAMS) established a goal to review the published medical data and develop an evidence-based consensus opinion regarding the role of calcium in peri- and postmenopausal women. DESIGN: In building this consensus opinion, NAMS followed the general principles established for evidence-based guidelines. As part of that process, NAMS appointed a panel of clinicians and researchers acknowledged to be experts in the field of calcium. Their advice was used to assist the NAMS Board of Trustees in developing this consensus opinion. RESULTS: Adequate calcium intake (in the presence of adequate vitamin D intake) has been shown to prevent bone loss and reduce fracture risk in peri- and postmenopausal women. Although calcium is not as effective as antiresorptive agents (e.g., estrogen, selective estrogen-receptor modulators, or bisphosphonates), it is an essential component of antiresorptive agent therapy for osteoporosis. Calcium has also been associated with beneficial effects in several nonskeletal disorders, primarily hypertension, colorectal cancer, obesity, and nephrolithiasis, although the extent of those effects and mechanisms involved have not been fully explored. Estimates of adequate intakes of calcium for peri- and postmenopausal women are based on evidence relating to osteoporosis prevention. At least 1,200 mg/day of calcium is required for most women; levels greater than 2,500 mg/day are not recommended. To ensure adequate calcium absorption, a daily intake of 400-600 IU of vitamin D is recommended, either through sun exposure or through diet or supplementation. Since no accurate test to determine calcium deficiency exists, clinicians should focus instead on ensuring that a woman consumes enough calcium to meet the recommended levels. CONCLUSION: Although the most definitive role for calcium in peri- and postmenopausal women is in bone health, it is clear that adequate calcium intake has implications that encompass a woman's overall health. Based on the available evidence, a strong statement can be made regarding the importance of ensuring adequate calcium intake in all women, particularly those in peri- or postmenopause. PMID- 11256880 TI - Comparison of the antiatherosclerotic effect of tibolone with that of estradiol and ethinyl estradiol in cholesterol-fed, ovariectomized rabbits. AB - OBJECTIVE: Tibolone is a synthetic steroid with tissue-specific estrogenic, progestogenic, and androgenic properties. The drug relieves climacteric symptoms and prevents osteoporosis but does not stimulate the endometrium. We have previously shown that in laboratory animals tibolone inhibits the atherogenesis induced by a high-cholesterol diet. Therefore, we compared the antiatherosclerotic effect of oral tibolone at different dose levels with that of oral 17beta-estradiol (E2) and ethinyl estradiol (EE). DESIGN: Atherosclerotic lesion formation (increase in vessel wall cholesterol deposition and fatty streak formation) was measured in ovariectomized rabbits after 20 weeks on an atherogenic diet (fed daily 80 g of a rabbit chow containing 0.4% cholesterol, 3.75% peanut oil, and 3.75% coconut oil) in eight groups: group 1, placebo (n = 35); group 2, control (n = 34) received normal rabbit chow; group 3, E2 group (E2 4 mg, n = 12); group 4, EE group (EE 60 microg, n = 10); and groups 5-8, tibolone (6 mg, n = 12; 2 mg, n = 13; 0.6 mg, n = 25; and 0.15 mg, n = 11, respectively). During the study, blood samples were obtained for the evaluation of plasma triglycerides, cholesterol, lipoproteins, and glutamate pyruvate transaminase. After 20 weeks, the animals were killed, and cholesterol concentration and the formation of fatty streaks in the wall of the aortic arch were evaluated. RESULTS: In the placebo group, the atherogenic diet induced a mean increase in total plasma cholesterol concentration from 1.1+/-0.1 mmol/L (control group) to 34.1+/-1.8 mmol/L (mean +/- SE). This resulted in an accumulation of cholesterol in the aortic arch from 48+/-4 (control group) to 608+/-44 nmol/mg protein and in the formation of fatty streaks (41.8+/-3.2% of the surface of the aortic arch was covered with fatty streaks). Tibolone had strong dose-dependent antiatherosclerotic effects. It reduced the accumulation of cholesterol in the aortic arch at doses of 6 to 0.15 mg by 99, 97, 87, and 57% and the formation of fatty streaks by 98, 97, 81, and 38%, respectively. E2 had only a marginal antiatherosclerotic effect, whereas EE showed an effect comparable to that of tibolone at doses of 2 to 0.6 mg. With EE, the accumulation of cholesterol in the vessel wall was reduced by 93% and the formation of fatty streaks by 73%. Mean plasma cholesterol concentrations were also reduced by tibolone (64, 70, 61, and 47%) and EE (57%). This reduction was mainly mediated via a reduction in beta very-low-density lipoprotein cholesterol. Analysis, however, indicated that the observed antiatherosclerotic effects of tibolone and EE, at least partly, are due to a direct effect on the vessel wall and independent of the changes in plasma cholesterol. At equipotent antiatherosclerotic doses, EE showed a stronger uterotropic effect (measured as the increase in uterine weight) than tibolone. EE increased uterine weight from 0.57 g/kg body weight (BW) (control group) to 3.5 g/kg BW; tibolone at doses of 6, 2, 0.6, and 0.15 mg increased uterine weight to 2.5, 2.8, 2.2, and 1.3 g/kg BW, respectively. CONCLUSION: Tibolone can protect the arterial vessel wall against atherosclerotic lesions induced by a hypercholesterolemic diet. However, it has much less estrogenic effects on the uterus compared with EE at equipotent doses, indicating tissue selectivity for tibolone. The clinical implications of these findings require investigation. PMID- 11256881 TI - Homocysteine-thiolactone induces caspase-independent vascular endothelial cell death with apoptotic features. AB - OBJECTIVE: Cell death is generally classified into two large categories: apoptosis, which represents active, physiological programmed cell death, and necrosis, which represents passive cell death without underlying regulatory mechanisms. Apoptosis plays an important role in tissue homeostasis and its role in endothelium integrity can be influenced by the functional status of endothelial cells. Homocysteine, a sulfated amino-acid product of methionine demethylation, is an independent risk factor for vascular disease (arterial and venous thombosis). Our goal was to investigate the thiol-derivatives effect on the endothelial cell apoptosis. METHODS: Three parameters were measured: mitochondrial membrane potential using DiOC6(3) as the probe, DEVDase activation, and phosphatidylserine exposure on the cell surface with fluorosceinated annexin V labeling which allows apoptosis to be distinguished from necrosis. RESULTS: Homocysteine-thiolactone induced endothelial cell apoptosis in a concentration dependent manner (range: 50-200 microM), independently of the caspase pathway. Only homocysteine-thiolactone, among the thiol derivatives tested, induced apoptosis. Apoptosis was not influenced by the serum concentration in culture medium, suggesting that the observed apoptotic process could occur in vivo. None of the inhibitors used (e.g., leupeptin, fumosinin B1, catalase, or z-VAD-fmk) was able to prevent homocysteine-induced apoptosis of vascular endothelial cells. CONCLUSION: The apoptosis of vascular endothelial cells induced by high concentration of homocysteine-thiolactone might be one step atherosclerotic cardiovascular disease, and contribute to its complication. PMID- 11256882 TI - Role of reactive oxygen species (ROS) in apoptosis induction. AB - Reactive oxygen species (ROS) and mitochondria play an important role in apoptosis induction under both physiologic and pathologic conditions. Interestingly, mitochondria are both source and target of ROS. Cytochrome c release from mitochondria, that triggers caspase activation, appears to be largely mediated by direct or indirect ROS action. On the other hand, ROS have also anti-apoptotic effects. This review focuses on the role of ROS in the regulation of apoptosis, especially in inflammatory cells. PMID- 11256883 TI - CD95/CD95L interactions and their role in autoimmunity. AB - CD95 (Fas/Apo-1) is a broadly expressed death receptor involved in a variety of physiological and pathological apoptotic processes. Since its discovery, defects in CD95/CD95L system have been proposed as major pathogenic factors responsible for impaired immunological tolerance to self antigens and autoimmunity. Later, analysis of altered sensitivity to CD95-induced apoptosis in cells targeted by the immune response has revealed an unexpected role for CD95 and CD95L in organ specific autoimmunity. CD95 has been shown to be expressed and functional in virtually all cell types that are target of the organ-specific autoimmune response. Here we review some of the major findings concerning the role of CD95 in autoimmunity, in dysfunctions due to increased or decreased CD95-induced apoptosis. PMID- 11256884 TI - Role of dysregulated apoptosis in atopic dermatitis. AB - Atopic dermatitis is a chronic inflammatory skin disease with a complex immune dysregulation and interplay of genetic, environmental and psychological factors. Activation and skin-selective homing of peripherial-blood T cells, and effector functions in the skin, represent sequential events in the pathogenesis. Dysregulated apoptosis in skin-homing T cells, eosinophils and keratinocytes contributes to the elicitation and persistence of atopic derrmatitis. PMID- 11256885 TI - Regulation of apoptosis and replicative senescence in CD8+ T cell following acute viral infection. AB - Viral infections are characterised by a large expansion of CD8+ effector T cells. Once generated, these T cells must be cleared and homeostasis re-established. In this review we describe two mechanisms, apoptosis and replicative senescence which are thought to play a vital role in this process. Apoptosis clears the unwanted T cells at the end of an immune response whereas replicative senescence prevents unlimited expansion of T cells. For successful memory to be produced T cells must avoid apoptosis and avoid replicative senescence. PMID- 11256886 TI - Regulation of apoptosis in mast cells. AB - Apoptosis is a physiological process of cell death that occurs in all multicellular organisms. Its dysregulation has been postulated as one of the main causes in the development of diseases such as cancer, AIDS, autoimmune diseases and allergy. Apoptosis has been mainly studied in the inflammatory cells that participate in the late and chronic stages of allergy (eosinophils, neutrophils, lymphocytes and macrophages) as a new way to elucidate the pathogenesis of this disease. Nevertheless, much less it is known about the regulation of apoptosis in the "initiators" of the allergic process: The Mast Cells. In normal conditions, mast cells are described as long-living cells that keep a constant number of cells in tissues. However, increased numbers of mast cells are observed in the late phase of asthma and in both the inflammatory and in the repair/remodeling stage of various inflammatory/fibrotic disorders. In this report, we discuss the possible mechanisms that regulate the apoptotic process in normal conditions and disease, such as survival factors and death receptors. A link between mast cell activation, during the early stages of the allergic process, and triggering of antiapoptotic signaling pathways is also suggested as an important contributor to the extended life of mast cells. PMID- 11256887 TI - Role of apoptosis in autoimmunity. AB - Apoptosis is a physiological form of cell death required to ensure that the rate of cell division is balanced by the rate of cell death in multicellular organisms. Dysregulation of apoptosis is associated with the pathogenesis of a wide array of diseases: cancer, neurodegeneration, autoimmunity, heart disease and others. In this review we collect arguments supporting a hypothesis of a dysregulated apoptosis leading to development of autoimmunity like systemic lupus erythematosus (SLE). This notion is supported by occurence of known autoantigens in apoptotic blebs, in vitro findings of an increased rate of apoptotic lymphoblasts despite optimal cytokine stimulation combined with a defective in vitro clearance of apoptotic bodies by SLE phagocytes. Moreover, we and others could generate histone-specific lymphocytic cell lines from cells after activation with autologous apoptotic material. These lymphocytes could stimulate autologous B-lymphocytes to produce of anti-dsDNA antibodies, a diagnostic hallmark for SLE. Finally, antibodies against phospholipids like phosphatidylserine are often associated with systemic autoimmunopathies like SLE and others. Phosphatidylserine is exposed on apoptotic cells as early sign of programmed cell death and serves as phagocyte recognition molecule for apoptotic cells. Formation of immune complexes and deposition in tissues might lead to organ damage and disease. This scenario will be discussed in this review in detail. PMID- 11256889 TI - Regulation of T cell apoptosis. AB - Proliferative expansion of lymphoid cells is required for effective immune responses against invading microorganisms, but after the infection is controlled, the expanded effector cells must be eliminated to prevent non-adaptive accumulation of cells. Higher vertebrates have developed extensive networks of signal transduction pathways to ensure controlled activation and expansion of cells during immune responses and apoptotic deletion of lymphoid cells that are no longer needed at the end of immune responses. Extracellular signals received by cell surface receptors that trigger intracellular signaling cascades are essential elements that control both processes. These signal transduction pathways converge to regulate cell fate at both transcriptional and post transcriptional levels. Here we review the role of pathways, especially those triggered by TNF receptor-related molecules, that determine the fate of T cells during development and activation. In addition, we introduce the possibility that these same pathways may be abnormally programmed and so lead to immune cell accumulation during inflammatory diseases such as asthma. PMID- 11256888 TI - Regulation of neutrophil apoptosis: a role for protein kinase C and phosphatidylinositol-3-kinase. AB - Neutrophils play a central role in host defense and are recruited in vast numbers to sites of infection where they phagocytose and kill invading bacterial pathogens. Neutrophils have a short half-life that is extended at the inflamed site by pro-inflammatory cytokines and contact with bacterial cell walls. Normal resolution of inflammation involves the removal of neutrophils and other inflammatory cells by the induction of apoptosis. Spontaneous neutrophil apoptosis does not require Fas ligation, but is mediated by caspases 3, 8 and possibly caspase 9 and also involves activation of protein kinase C-delta. With chronic inflammatory disease, neutrophil apoptosis is delayed by pro-inflammatory cytokines, leading to persistence of neutrophils at the inflamed site and non specific tissue damage. Here we discuss the evidence for inhibition of neutrophil apoptosis via signaling though PI-3-kinase and downstream pathways, including PDK 1 and PKB. Therapeutic strategies to resolve chronic inflammation could therefore usefully target neutrophil apoptosis and the PI-3-kinase or PKC-delta signaling pathways. PMID- 11256890 TI - Apoptosis and airway inflammation in asthma. AB - Asthma is a disease characterized by a chronic inflammation of the airways and by structural alterations of bronchial tissues, often referred to as airway remodelling. The development of chronic airway inflammation in asthma depends upon the continuous recruitment of inflammatory cells from the bloodstream towards the bronchial mucosa and by their subsequent activation. It is however increasingly accepted that mechanisms involved in the regulation of the survival and apoptosis of inflammatory cells may play a central role in the persistent inflammatory process characterizing this disease. Increased cellular recruitment and activation, enhanced cell survival and cell:cell interactions are therefore the key steps in the development of chronic airway inflammation in asthma, and represent the major causes for tissue damge, repair and remodelling. PMID- 11256891 TI - High-impact exercise and bones of growing girls: a 9-month controlled trial. AB - The maximum amount of bone a person can obtain during the first two decades of life is an important determinant of bone mass in later life, and an increase in peak bone mass has been associated with decreased risk for osteoporotic fractures. It is known that growth of bone and thus development of peak bone mass are strongly controlled by genetic factors, but information on the role of environmental factors, such as exercise and nutrition, (e.g., exercise) on growing bone is limited. We tested a hypothesis that in growing girls the benefit of mechanical loading on bone mineral mass and bone strength is better before rather than after the menarche. Sixty-four girls (25 premenarcheal, 39 postmenarcheal) carried out a supervised 9-month step-aerobic program (two sessions per week), each session complemented with additional jumps. Sixty-two girls (33 premenarcheal, 29 postmenarcheal) served as controls. Bone mineral content (BMC) at the lumbar spine and proximal femur was measured by dual-energy X-ray absorptiometry (DXA). In addition, the cortical density (CoD, mg/cm3) and cortical cross-sectional area (CoA, mm2) and the density-weighted polar section modulus (BSI, mm3) of the tibial midshaft were determined by peripheral quantitative tomography (pQCT). In the premenarcheal girls, BMC increased statistically significantly more in the trainees than controls at the lumbar spine (p = 0.012) (8.6% vs 5.3%) and femoral neck (p = 0.014) (9.3% vs 5.3%). In the tibial midshaft, the intergroup differences (CoD, CoA and BSI) were not significant. The postmenarcheal girls showed no significant post-training intergroup differences in any of the bone parameters (BMC increased in the lumbar spine 6.0% vs 4.9%; femoral neck 3.4% vs 3.2%; and trochanter 2.6% vs 3.5%). Although a large proportion of bone mineral increase in the growing girls of this study was attributable to growth itself, this 9-month exercise intervention showed that a clear and large additional bone gain could be obtained in exercising premenarcheal girls, but not in exercising postmenarcheal girls. In other words, exercise seemed more beneficial for additional bone mineral acquisition before menarche (i.e., during the growth spurt) rather than after it. PMID- 11256892 TI - Femoral neck and intertrochanteric fractures have different risk factors: a prospective study. AB - The aim of this study was to determine whether both types of hip fracture, femoral neck and intertrochanteric, have similar risk factors. A prospective cohort study was carried out on community-dwelling elderly women in four areas of the United States: Baltimore, MD; Pittsburgh, PA; Minneapolis, MN and Portland, OR. The participants were 9704 Caucasian women, 65 years and older, of whom 279 had fractured their femoral neck and 222 had fractured their trochanteric region of the proximal femur. The predictors used were the bone mass of the calcaneus and proximal femur, anthropometry, history of fracture (family and personal), medication use, functional status, physical activity and visual function. The main outcome measures were femoral neck and intertrochanteric fractures occurring during an average of 8 years of follow-up. In multivariate proportional hazards models, several risk factors increased the risk of both types of hip fracture; including femoral neck bone density and increased functional difficulty. In hazard regression models that directly compared risk factors for the two types of hip fracture, calcaneal bone mineral density (BMD) predicted femoral neck fractures more strongly than intertrochanteric fractures (OR = 1.16; 95% CI = 1.02-1.31). Steroid use and impaired functional status also predicted femoral neck fractures instead of intertrochanteric fractures. Poor health status (OR = 0.74; 95% CI = 0.55-1.00) predicted intertrochanteric fractures more strongly than femoral neck fractures. We conclude that femoral neck fractures are largely predicted by BMD and poor functional ability while aging and poor health status predispose to intertrochanteric fractures. PMID- 11256893 TI - Bone size and bone mass in 10-year-old Danish children: effect of current diet. AB - Lifestyle factors, such as diet, are believed to be involved in modifying bone health, although the results remain controversial, particularly in children and adolescents. The objective of the study was to identify associations between dietary factors and whole body bone measurements in 10-year-old children. The study was a cross-sectional analysis of a random sample of 105 healthy Danish children, aged 10 years (9.97+/-0.09). Whole body bone mineral content (BMC) and bone area (BA) were determined by dual-energy X-ray absorptiometry. The influence of diet (7 day food records) on BMC and BA were examined in bi- and multivariate analyses. The mean intakes of calcium, protein, phosphorus and sodium were 1226 mg, 78 g, 1523 mg and 3.3 g, respectively. In bivariate analyses, BMC and BA were strongly positively correlated with height (p<0.001) and weight (p<0.001), and with intakes of energy (p<0.005) and several nutrients. BMC was adjusted for size by including BA, height and weight in the multiple linear regression, and BA was adjusted for size by including height and weight in the multiple linear regression. In multivariate analyses, size-adjusted BMC was positively associated with calcium intake (p=0.02). Size-adjusted BA was positively associated with dietary protein (p=0.003), and negatively associated with intakes of sodium (p = 0.048) and phosphorus (p=0.01). In conclusion, calcium intake was positively associated with bone mineralization. There was a positive association between protein and BA, while for phosphorus and sodium the association was negative. PMID- 11256894 TI - The association between parathyroid hormone, vitamin D and bone mineral density in 70-year-old Icelandic women. AB - Parathyroid hormone (PTH) may be an important determinant of cortical bone remodeling in the elderly. Vitamin D status is one of the determining factors in this relationship. The aim of this study was to quantify the relationship between serum PTH, vitamin D and bone mineral density (BMD) in elderly women in Reykjavik (64 degrees N), where daily intake of cod liver oil is common and mean calcium intake is high. In PTH correlated inversely with 25(OH)D (r = -0.26, p<0.0 1). In multivariate analysis PTH correlated inversely with whole body BMD (mostly cortical bone) (R2 = 2.2%, p = 0.04) but not with the lumbar spine BMD, reflecting more cancellous bone. No association was found between 25(OH)D levels and BMD at any site in univariate or multivariate analysis. Osteocalcin, a measure of bone turnover, was negatively associated with BMD and this association remained significant when corrected for PTH levels. In summary, in this fairly vitamin D replete population with high calcium intake, PTH was negatively associated with total body BMD. We infer that suppression of PTH may reduce cortical bone loss, but other factors are likely to contribute to age-related bone remodeling and osteoporosis. PMID- 11256895 TI - Organochlorines and bone mineral density in Swedish men from the general population. AB - Persistent organochlorines (POCs), such as polychlorinated biphenyls (PCBs) and DDT, are present at relatively high concentrations in food and show estrogenic, anti-estrogenic or anti-androgenic activity in biological test systems. Because bone mineral density (BMD) in men is influenced by sex hormones, we looked for associations between BMD and serum concentrations of POCs in 115 men (mean age 63 years, range 40-75 years) from the general Swedish population. Ten PCB congeners, five DDT isomers, hexachlorobenzene, three hexachlorocyclohexane isomers, trans nonachlor and oxychlordane were analyzed by gas chromatography. Quantitative bone measurements were performed by dual-energy X-ray absorptiometry at three sites: whole body, the L2-L4 region of the lumbar spine, and the neck region of the proximal femur, as well as by quantitative ultrasound on the left os calcis (broadband ultrasound attenuation (BUA) and speed of sound (SOS)). After adjustment for confounding factors in linear regression analyses we found no strong association between serum concentrations of single POCs and the five BMD and ultrasound variables. When POCs were grouped according to hormonal activity (estrogenic, anti-estrogenic, anti-androgenic) and the study subjects were divided into organochlorine concentration quartiles, a weak association was indicated between increased serum concentrations of p,p'-DDE (antiandrogenic) and decreased BMD, BUA and SOS. This may suggest that p,p'-DDE could cause negative effects on bone density, but the findings might also be due to chance since multiple comparisons were made in the statistical analysis. Overall our results do not suggest that the studied POCs caused major effects on bone density in our study group. PMID- 11256896 TI - Bone mineral density and body composition in underweight and normal elderly subjects. AB - The importance of malnutrition as a risk factor in osteoporosis is emphasized by the evidence that patients with fractures of the proximal femur are often undernourished. In this study, nutritional status, bone mineral mass and its association with body composition were investigated in underweight and normal weight elderly subjects. Moreover the hypothesis that malnutrition in elderly is associated with a higher risk of osteoporosis was tested. The participants were 111 elderly subjects divided into two groups according to body mass index (BMI): 51 patients were underweight (BMI < 22 kg/m2) while in 60 subjects BMI ranged from 22 to 30 kg/m2. In all patients anthropometric parameters and blood indices of malnutrition and of bone turnover were measured. Fat-free soft mass (FFSM), fat mass (FM), bone mineral content (BMC) and bone mineral density (BMD) 'total body' and at the hip were obtained by dual-energy X-ray densitometry. Dietary intake was evaluated with the diet history method, while resting energy expenditure (REE) was measured by indirect calorimetry. Underweight subjects had other signs of malnutrition, such as low visceral proteins, sarcopenia, and an inadequate energy intake. Moreover they showed a significant reduction of BMC and BMD compared with normal subjects. In men with BMI <22 kg/m2, T-score was below 2.5 (-3 at femoral neck and -2.7 at total hip) while men in the control group had normal bone mineral parameters. T-score at different sites was lower in underweight women than in underweight men, always showing values under -3.5, with clear osteoporosis and a high fracture risk. In healthy women the T-score values indicated the presence of mild osteoporosis. In underweight subjects, low levels of albumin (< 35 g/l) were associated with higher femoral bone loss. Using a partial correlation model, BMC, adjusted for age, bone area, knee height and albumin showed a significant association with FM in women (r = 0.48; p < 0.01) and with FFSM in men (r = 0.48; p < 0.05). Albumin, when adjusted for other variables, was significantly correlated (r = 0.52; p < 0.05) with femoral neck BMC only in women. In conclusion, the underweight state in the elderly is associated with malnutrition and osteoporosis; other factors occurring in malnutrition, besides body composition changes, such as protein deficiency, could be involved in the association between underweight and osteoporosis. Moreover bone mineral status seems to be related to fat-free soft mass tissue in men while in women it is much more closely associated with total body fat. PMID- 11256897 TI - Comparison of six calcaneal quantitative ultrasound devices: precision and hip fracture discrimination. AB - Quantitative ultrasound (QUS) is now accepted as a useful tool in the management of osteoporosis. There are a variety of QUS devices clinically available with a number of differences among them, including their coupling methods, parameter calculation algorithms and sites of measurement. This study evaluated the abilities of six calcaneal QUS devices to discriminate between normal and hip fractured subjects compared with the established method of dual-energy X-ray absorptiometry (DXA). The short-term and mid-term precisions of these devices were also determined. Thirty-five women (mean age 74.5+/-7.9 years) who had sustained a hip fracture within the past 3 years, and 35 age-matched controls (75.8+/-5.6 years) were recruited. Ultrasound measurements were acquired using six ultrasound devices: three gel-coupled and three water-coupled devices. Bone mineral density was measured at the hip using DXA. Discrimination of fracture patients versus controls was assessed using logistic regression analysis (expressed as age- and BMI-adjusted odds ratios per standard deviation decrease with 95% confidence interval) and receiver operating characteristics (ROC) curve analysis. Measurement precision was standardized to the biological range (sCV). The sCV ranged from 3.14% to 5.5% for speed of sound (SOS) and from 2.45% to 6.01% for broadband ultrasound attenuation (BUA). The standardized medium-term precision ranged from 4.33% to 8.43% for SOS and from 2.77% to 6.91% for BUA. The pairwise Pearson correlation coefficients between different devices was highly significant (SOS, r = 0.79-0.93; BUA, r = 0.71-0.92). QUS variables correlated weakly, though significantly, with femoral BMD (SOS, r = 0.30-0.55; BUA, r = 0.35 0.61). The absolute BUA and SOS values varied among devices. The gel-coupled devices generally had a higher SOS than water-coupled devices. Bone mineral density (BMD) and BUA were weakly correlated with weight (r = 0.48-0.57 for BMD and r = 0.18-0.54 for BUA), whereas SOS was independent of weight. All the QUS devices gave similar, statistically significant hip fracture discrimination for both SOS and BUA measures. The odds ratios for SOS (2.1-2.8) and BUA (2.4-3.4) were comparable to those for femoral BMD (2.6-3.5), as were the area under the curve (SOS, 0.65-0.71; BUA, 0.62-0.71; BMD, 0.65-0.74) from ROC analysis. Within the limitation of the sample size all devices show similar diagnostic sensitivity. PMID- 11256898 TI - Peak bone mass. PMID- 11256899 TI - Autonomic functioning toxic nodule-therapeutic options. PMID- 11256900 TI - Efficacy of standard ten millicurie dose of radio-iodine in management of autonomously functioning toxic thyroid nodules. AB - Therapeutic effect of radio-iodine treatment on thyroid patients with autonomously functioning toxic thyroid nodule was evaluated. Fifty one patients were given a standard dose of 10 mci of radioiodine (I-131) and were followed up for 2-3 years. The failure rate (relapse after 10 mci of radioiodine) of this regime was 10%. It was found that the nodules less than or equal to 3 cms. in size were completely cured after a dose of 10 mic. of radio-iodine therapy, over a follow up period of next 6 months. Patients having nodules larger than 3 cms. relapsed after first dose of 10 mci of radio-iodine, but were cured completely after the second dose of 10 mci of radio-iodine therapy. Tri-iodothyronine (T3) and thyroxine (T4) values were both found to be high before giving treatment in all the cases. Only one case developed hypothyroidism after radioiodine therapy. PMID- 11256901 TI - Effect of intraperitoneal administration of sodium nitroprusside on the efficacy of peritoneal dialysis. AB - A prospective randomised trial of intraperitoneal sodium nitroprusside (SNP) administration on the efficacy of acute intermittent peritoneal dialysis was carried out in 40 adult patients of acute or acute on chronic renal failure. A total of 36 cycles of peritoneal dialysis (PD) with an exchange volume of 1 litre and duration of 1 hour per cycle were given to each patient. The 36 cycles of PD were divided into 12 clearance periods (I-XII) of 3 cycles each. SNP was added in a dosage of 4 mg/l of dialysate in clearance period II, IV, VI and VIII. Of 40 patients, 20 were subjected to standard PD (Gp A) while the other 20 received SNP added PD (Gp B). The peritoneal clearance of urea, creatinine, percentage fall of blood urea, serum creatinine and protein loss during the various clearance periods were compared in the two groups. It was observed that group B patients had significantly higher peritoneal clearance and the percentage reduction in the blood levels of urea and creatinine. Protein loss per clearance period was also significantly higher in group B patients. Twenty two cycles of SNP added PD were as effective as 36 cycles of standard PD. No systematic untoward effects of SNP were observed. It is therefore, concluded that intraperitoneal SNP administration is a safe and effective way of increasing the efficacy of PD thus reducing the duration of treatment. PMID- 11256902 TI - Renal failure in acute pancreatitis. AB - Twenty six (7.3%) of a total of 356 patients with acute renal failure were found to have acute pancreatitis as the primary disease. Seventeen (65.4%) of them were males. Their mean age was 35.6 years. Clinically epigastric pain and tenderness were seen in all (100%); nausea vomiting (73%), low grade fever (50%), left sided pleural effusion (38.4%), haemopericardium (26.9%), shock (26.9%), pseudocyst (19.3%) and adult respiratory distress syndrome (7.6%) were the other major presenting features. Serum amnylase (100%), lipase (53.8%), triglycerides (53.8%) and blood sugar (38.5%) were raised in majority whereas serum calcium was detected to be below normal in 46.2% patients. Blood urea and serum creatinine were raised in all and hyperkalacmia was found in 50% patients. CT scan and USG abdomen showed bilateral enlarged kidneys (100%), pancreatic oedema (80.7%), necrosis of pancreas (19.3%) and pseudocyst (19.3%). Management included repeated peritoneal dialysis in all (100%) and surgical intervention in 53.8% patients with severe necrotising and haemorrhagic pancreatitis. All patients recovered from acute renal failure, but 26.9% patients expired due to complications of acute pancreatitis other than acute renal failure. PMID- 11256903 TI - Categorisation of ventilatory pattern of response to bronchodilator therapy in airflow limitation. AB - Improvement in lung function studies following bronchodilator inhalation leads to different pattern of response in ventilatory parameters which are helpful in categorizing the patients into groups for correct interpretation of bronchodilator response and assessment of prognosis of the disease. PMID- 11256904 TI - Spleen size in health and disease: a sonographic assessment. AB - Ultrasonography was used to evaluate splenic size in normal patients as well as in patients with various clinical conditions. To express splenic size, a splenic volumetric index (SVI) was used. By grading the SVI on the basis of age and sex in normal patients, and in various diseases, characteristic distributions of SVI were obtained. Obtaining the SVI by the use of ultrasound appears to be a significant supplemental aid for evaluating spleen size, especially in patients whose spleens are not palpable. PMID- 11256905 TI - Effect of cigarette smoking on lipid profile in the young. AB - In view of the controversies existing regarding the atherogenic potential of smoking, this study was conducted in 40 healthy young male Cigarette smokers and 40 age and weight matched male non smokers, to find out the difference in the serum lipid profiles of both the groups. Subjects in both the groups were in the age range of 25 and 35 years having no history of alcohol abuse or diseases like diabetes mellitus or obesity. The mean serum total cholesterol (177.3 +/- 32.5 mg/dL) and LDL cholesterol (100.2 +/- 31.0 mg/dL) were significantly higher in smokers (p < 0.05) whereas mean serum HDL- Cholesterol was (43.2 +/- 5.8 mg/dL) was significantly lower (P < 0.05). Mean triglyceride (170.8 +/- 59.7 mg/dL) was significantly higher in smokers than in nonsmokers (p < 0.01). In the fed state the total serum cholesterol level and triglyceride level was increased by 10.4 mg/dL and 51.1 mg/dL respectively in smokers whereas the increase was 4.8 mg/dL and 24.3 mg/dL respectively in nonsmokers. There was less rise of HDL cholesterol (1.9 mg/dL) in smokers as compared to that in nonsmokers (3.4 mg/dL) and in LDL cholesterol (1.8 mg/dL) in smokers compared to nonsmokers (3.4 mg/dL) in fed state. PMID- 11256906 TI - Incidence of carrier state in treated patients of typhoid. PMID- 11256907 TI - Efficacy of low dose intradermal recombinant hepatitis B vaccine with and without immunomodulation in end stage renal disease patients on maintenance hemodialysis. PMID- 11256908 TI - Writer's cramp. PMID- 11256909 TI - Magnesium in cardiovascular therapy. PMID- 11256910 TI - Epidemiology of gallbladder cancer: present status. PMID- 11256911 TI - The idiopathic hypereosinophilic syndrome. PMID- 11256912 TI - Maturity onset diabetes of the young. PMID- 11256913 TI - Right sided infective endocarditis in an elderly non-addict female. PMID- 11256914 TI - Corticosteroid resistant haemolytic crisis in cold agglutinin disease: successful management by plasmapheresis and pulse cyclophosphamide. PMID- 11256915 TI - Right atrial myxoma. PMID- 11256916 TI - Idiopathic lateral sinus thrombosis. PMID- 11256917 TI - Intracranial tuberculoma manifesting as fever of unknown origin. PMID- 11256919 TI - Tuberculous pleurisy mimicking pleural mesothelioma. PMID- 11256918 TI - Cryptococcal meningitis in unimmunocompromised patients. PMID- 11256920 TI - Brain-stem involvement in thyroid fever. PMID- 11256922 TI - Facial palsy of uncommon aetiology (cephalic tetanus). PMID- 11256921 TI - Thymolipoma--a rare asymptomatic mediastinal tumour. PMID- 11256923 TI - Presentation at GOLD APICON. PMID- 11256924 TI - Phenformin induced lactic acidosis in an elderly NIDDM patient. PMID- 11256925 TI - Suspect more adrenocortical insufficiencies. PMID- 11256926 TI - Gall bladder involvement in acute viral hepatits. PMID- 11256927 TI - Neuro psychiatric presentations in a case of infective endocarditis. PMID- 11256928 TI - Disease modifying anti-rheumatic drugs (DMARDs) in rheumatoid disease. PMID- 11256929 TI - Dopa responsive dystonia and the Indian literature. PMID- 11256930 TI - Anti leishmanial therapy--the changing scene. PMID- 11256931 TI - Epilepsy: a common manifestation of neurocysticercosis. PMID- 11256932 TI - Early microscopy: history of fine needle aspiration (FNA) with particular reference to goitres. PMID- 11256934 TI - Adenocarcinoma of the uterine cervix compared with squamous cell carcinoma: a 12 year study in Southampton and South-west Hampshire. AB - In a 12-year study of the population of Southampton and South-west Hampshire (SSWH), there was no rise or fall in the incidence of adenocarcinoma of the uterine cervix, although the incidence of squamous cell carcinoma fell from 14 to 7.2 per 100000 women years and the overall fall in age-adjusted incidence of cervical carcinoma was commensurate with that of England and Wales. The majority (59%) of adenocarcinomas were seen in women aged less than 50, supporting the concept of a higher risk in young women. Screen-detected carcinomas accounted for 50% of adenocarcinomas and 41% of squamous cell carcinomas in women aged 20-64 (the difference was not significant). There were more screen-detected adenocarcinomas of less than 3 mm depth of invasion and 7 mm lateral extension during the third period of the study (1991-1993). The results are consistent with reports of an increased risk of cervical cancer in women born since 1940, and lesser effectiveness of screening in preventing adenocarcinoma compared with squamous cell carcinoma. High prevalence of early screen-detected carcinomas may have been a factor in recent reports of increased incidence of adenocarcinoma. PMID- 11256933 TI - Cytological grading of breast cancer in Giemsa-stained fine needle aspiration smears. AB - A potential cytological nuclear grading based on a semi-quantitative evaluation of three basic nuclear features, size of cell nuclei, anisonucleosis and the proportion of nucleoli-containing-nuclei, was tested on 74 Giemsa-stained fine needle aspiration of breast smears for its reliability in establishing the malignant potential of breast cancer. The prognostic impact of DNA-ploidy and S phase fraction was also assessed. A good correlation between the three basic nuclear features, DNA-ploidy, S-phase fraction, cytological nuclear grade and histological grade, was shown. Using the cytological nuclear grade proposed, correct classification of cases between low histological grade (HG I) and high histological grade (HG II + HG III) was achieved in 79.73%. A statistically significant difference in 5-year survival rate was also observed between low malignancy grade and high malignancy grade breast cancer patients, regardless of the grading method used. DNA-ploidy and S-phase fraction were not statistically significant in establishing the malignant potential of breast cancer. PMID- 11256935 TI - Cytological evaluation of angiogenesis in endometrial aspirates. AB - We conducted a comparative study of angiogenesis observed in endometrial aspirates according to histological type. Cytological specimens from 14 cases of proliferative phase endometrium, 21 cases of endometrial hyperplasia and 18 cases of well-differentiated endometrial adenocarcinoma were used in the investigation. Immunohistochemical staining was performed according to standard methods using CD34 monoclonal antibody, and new vessels were examined. New vessels were identified in all of the histological types, but no morphological differences were seen. New vessels were observed in more cases of adenocarcinoma than in cases of normal tissue or hyperplasia, and the differences were significant. The difference between the maximum and minimum rates of occurrence of cell clusters possessing new vessels tended to be greater in adenocarcinoma than in the other tissue types (P < 0.05). Based on the above findings, examination of new vessels appearing in aspirated endometrial specimens appeared to be of help in differential diagnosis, but it also seemed necessary to take changes due to the menstrual cycle etc. into consideration. PMID- 11256936 TI - Aspiration cytology from the pouch of Douglas at hysteroscopy. AB - Eighty-seven fine needle aspiration cytological samples from 29 patients suffering from irregular perimenopausal uterine bleeding were evaluated. Aspiration cytology was performed prior to hysteroscopy, after distension of the uterine cavity and finally after uterine curettage. In this paper, cytological examination of fluid from the pelvic content in women with benign endometrial findings is compared with that of patients with adenocarcinoma. Endometrial curettage influenced the cell content of the cytologica specimens. PMID- 11256937 TI - Perplexing findings in a cystic teratoma. PMID- 11256938 TI - Malignant mesothelioma mimicking squamous carcinoma in a pleural fluid aspirate. PMID- 11256939 TI - Deciduoid peritoneal mesothelioma. A report of the cytological appearances. PMID- 11256940 TI - Quantitative cytological assessment of a persistent oral lesion that underwent malignant transformation: a case report. PMID- 11256941 TI - The utility of transbronchial (Wang) fine needle aspiration in lung cancer diagnosis. AB - We evaluated our experience with transbronchial fine needle aspiration (TBNA) in cancer diagnosis over a period of 1 year. A total of 51 aspirates were performed by specialist chest physicians in the presence of a cytopathologist who made on spot evaluation of Diff-Quik smears for adequacy and guided the aspirator for additional sampling if necessary. Two clusters of at least 10 malignant cells were required on the Diff-Quik smears to render an on the spot positive diagnosis of malignancy. Aspirates showing atypical cells or few malignant cells not fulfilling the above criteria were placed in a suspicious category and additional material was requested. The TBNA results were correlated with the transbronchial biopsy when available. PMID- 11256942 TI - Recombinant aequorin and green fluorescent protein as valuable tools in the study of cell signalling. AB - Luminous proteins include primary light producers, such as aequorin, and secondary photoproteins that in some organisms red-shift light emission for better penetration in space. When expressed in heterologous systems, both types of proteins may act as versatile reporters capable of monitoring phenomena as diverse as calcium homoeostasis, protein sorting, gene expression, and so on. The Ca(2+)-sensitive photoprotein aequorin was targeted to defined intracellular locations (organelles, such as mitochondria, endoplasmic reticulum, sarcoplasmic reticulum, Golgi apparatus and nucleus, and cytoplasmic regions, such as the bulk cytosol and the subplasmalemmal rim), and was used to analyse Ca(2+) homoeostasis at the subcellular level. We will discuss this application, reviewing its advantages and disadvantages and the experimental procedure. The applications of green fluorescent protein (GFP) are even broader. Indeed, the ability to molecularly engineer and recombinantly express a strongly fluorescent probe has provided a powerful tool for investigating a wide variety of biological events in live cells (e.g. tracking of endogenous proteins, labelling of intracellular structures, analysing promoter activity etc.). More recently, the demonstration that, using appropriate mutants and/or fusion proteins, GFP fluorescence can become sensitive to physiological parameters or activities (ion concentration, protease activity, etc.) has further expanded its applications and made GFP the favourite probe of cell biologists. We will here present two applications in the field of cell signalling, i.e. the use of GFP chimaeras for studying the recruitment of protein kinase C isoforms and the activity of intracellular proteases. PMID- 11256943 TI - Organization of uroplakin subunits: transmembrane topology, pair formation and plaque composition. AB - The apical surfaces of urothelial cells are almost entirely covered with plaques consisting of crystalline, hexagonal arrays of 16 nm uroplakin particles. Although all four uroplakins, when SDS-denatured, can be digested by chymotrypsin, most uroplakin domains in native urothelial plaques are resistant to the enzyme, suggesting a tightly packed structure. The only exception is the C terminal, cytoplasmic tail of UPIII (UPIII) which is highly susceptible to proteolysis, suggesting a loose configuration. When uroplakins are solubilized with 2% octylglucoside and fractionated with ion exchangers, UPIa and UPII were bound as a complex by a cation exchanger, whereas UPIb and UPIII were bound by an anion exchanger. This result is consistent with the fact that UPIa and UPIb are cross-linked to UPII and UPIII, respectively, and suggests that the four uroplakins form two pairs consisting of UPIa/II and UPIb/III. Immunogold labelling using a new mouse monoclonal antibody, AU1, revealed that UPIII is present in all urothelial plaques, indicating that the two uroplakin pairs are not segregated into two different types of urothelial plaque and that all plaques must have a similar uroplakin composition. Taken together, these results indicate that uroplakins form a tightly packed structure, that the four uroplakins interact specifically forming two pairs, and that both uroplakin pairs are required for normal urothelial plaque formation. PMID- 11256944 TI - Identification of the G13 (cAMP-response-element-binding protein-related protein) gene product related to activating transcription factor 6 as a transcriptional activator of the mammalian unfolded protein response. AB - Eukaryotic cells control the levels of molecular chaperones and folding enzymes in the endoplasmic reticulum (ER) by a transcriptional induction process termed the unfolded protein response (UPR). The mammalian UPR is mediated by the cis acting ER stress response element consisting of 19 nt (CCAATN(9)CCACG), the CCACG part of which is considered to provide specificity. We recently identified the basic leucine zipper (bZIP) protein ATF6 as a mammalian UPR-specific transcription factor; ATF6 is activated by ER stress-induced proteolysis and binds directly to CCACG. Here we report that eukaryotic cells express another bZIP protein closely related to ATF6 in both structure and function. This protein encoded by the G13 (cAMP response element binding protein-related protein) gene is constitutively synthesized as a type II transmembrane glycoprotein anchored in the ER membrane and processed into a soluble form upon ER stress as occurs with ATF6. The proteolytic processing of ATF6 and the G13 gene product is accompanied by their relocation from the ER to the nucleus; their basic regions seem to function as a nuclear localization signal. Overexpression of the soluble form of the G13 product constitutively activates the UPR, whereas overexpression of a mutant lacking the activation domain exhibits a strong dominant-negative effect. Furthermore, the soluble forms of ATF6 and the G13 gene product are unable to bind to several point mutants of the cis-acting ER stress response element in vitro that hardly respond to ER stress in vivo. We thus concluded that the two related bZIP proteins are crucial transcriptional regulators of the mammalian UPR, and propose calling the ATF6 gene product ATF6alpha and the G13 gene product ATF6beta. PMID- 11256945 TI - Alpha-melanocyte-related tripeptide, Lys-d-Pro-Val, ameliorates endotoxin-induced nuclear factor kappaB translocation and activation: evidence for involvement of an interleukin-1beta193-195 receptor antagonism in the alveolar epithelium. AB - The potential anti-inflammatory role of alpha-melanocyte-stimulating hormone (alpha-MSH)-related tripeptide, lysine(11)-D-proline-valine(13) (KDPV), an analogue of interleukin (IL)-1beta(193-195) and an antagonist of IL 1beta/prostaglandin E(2), is not well characterized in the alveolar epithelium. In a model of foetal alveolar type II epithelial cells in vitro, we showed that lipopolysaccharide endotoxin (LPS) differentially, but selectively, induced the nuclear subunit composition of nuclear factor kappaB(1) (NF-kappaB(1)) (p50), RelA (p65) and c-Rel (p75), in parallel to up-regulating the DNA-binding activity (supershift indicating the presence of the p50-p65 complex). LPS accelerated the degradation of inhibitory kappaB-alpha (IkappaB-alpha), accompanied by enhancing its phosphorylation in the cytosolic compartment but not in the nucleus. KDPV suppressed, in a dose-dependent manner, the nuclear localization of p50, p65 and p75, an effect that led to the subsequent inhibition of NF-kappaB activation. Interleukin-1 receptor antagonist (IL-1ra) decreased the nuclear abundance of p50, p65 and p75, and subsequently depressed the DNA-binding activity induced by LPS. Analysis of the mechanism involved in the KDPV- and IL-1ra-mediated inhibition of NF-kappaB nuclear localization revealed a reversal in IkappaB-alpha phosphorylation and degradation, followed by cytosolic accumulation. LPS induced endogenous IL-1beta biosynthesis in a time-dependent manner; the administration of exogenous recombinant human interleukin 1 (rhIL-1) resulted in a dose dependent activation of NF-kappaB. KDPV and IL-1ra abrogated the effect of rhIL 1. Pretreatment with the non-steroidal anti-inflammatory drug (NSAID) indomethacin, an inhibitor of cyclo-oxygenase, blocked the LPS-induced activation of NF-kappaB. These results indicate the involvement of prostanoid-dependent (NSAID-sensitive) and IL-1-dependent (IL-1ra-sensitive) mechanisms mediating LPS induced NF-kappaB translocation and activation, a pathway that is regulated, in part, by a negative feedback mechanism transduced through IkappaB-alpha, the major cytosolic inhibitor of NF-kappaB. PMID- 11256946 TI - Kinetics of CO binding and CO photodissociation in Pseudomonas stutzeri cd(1) nitrite reductase: probing the role of extended N-termini in fast structural relaxation upon CO photodissociation. AB - cd(1) nitrite reductase from Pseudomonas stutzeri is a di-haem- containing enzyme, comprising a c-type haem and a d-type haem. Studies with the highly related cd(1) nitrite reductase of Pseudomonas aeruginosa have established that this enzyme undergoes fast (microsecond) and global structural relaxation upon CO photodissociation from the reduced enzyme. A key difference between the Ps. aeruginosa and Ps. stutzeri enzyme is the absence of a flexible N-terminal extension in the Ps. stutzeri enzyme. In Ps. aeruginosa cd(1) nitrite reductase the N-terminal extension wraps around the second subunit of the homodimer and with Tyr(10) stabilizing a water molecule co-ordinated to the d(1)-haem. Given the intimate association of the N-terminal extension with the d(1)-haem, we hypothesized that the presence of the N-terminal extension likely contributes to the fast structural reorganization seen during photodissociation of CO from the reduced enzyme. In the present study we have investigated the kinetics of CO association and CO photodissociation of Ps. stutzeri cd(1) nitrite reductase (which lacks the N-terminal arm seen in the Ps. aeruginosa enzyme) to probe the role and influence of the N-terminal arm in the fast global structural reorganization seen with Ps. aeruginosa. Surprisingly, we find that Ps. stutzeri cd(1) nitrite reductase also undergoes fast structural reorganization during CO photodissociation. We also show, in stopped-flow experiments, that the kinetics of CO binding and dissociation with reduced Ps. stutzeri cd(1) nitrite reductase are similar to those observed with Ps. aeruginosa enzyme, thus ruling out a major role for the N-terminal flexible arm found in Ps. aeruginosa in the kinetics of these processes. Our data indicate that global structural reorganization following CO photodissociation is an intrinsic property of the haem domains in cd(1) nitrite reductases. The absence of an N-terminal extension, as in the Ps. stutzeri cd(1) nitrite reductase, does not lead to loss of global structural reorganization following CO photodissociation. PMID- 11256947 TI - Characterization of the interaction between the transcription factors human polyamine modulated factor (PMF-1) and NF-E2-related factor 2 (Nrf-2) in the transcriptional regulation of the spermidine/spermine N1-acetyltransferase (SSAT) gene. AB - Polyamines and polyamine analogues have been demonstrated to modulate the transcription of various genes. Spermidine/spermine N(1)-acetyltransferase (SSAT) is transcriptionally regulated through the interaction of at least two trans acting transcription factors, NF-E2-related factor 2 (Nrf-2) and PMF-1 (polyamine modulated factor-1). Nrf-2 has previously been shown to regulate transcription of other genes through interactions between its C-terminal leucine zipper and the leucine-zipper region of other members of the small Maf protein family (the term "Maf" is derived from MusculoAponeurotic-Fibrosarcoma virus). Here it is demonstrated that the interaction between Nrf-2 and PMF-1 is mediated through the binding of the leucine-zipper region of Nrf-2 and a C-terminal coiled-coil region of PMF-1 that does not contain a leucine zipper. Mutations that interrupt either the leucine zipper of Nrf-2 or the coiled-coil region of PMF-1 are demonstrated to alter the ability of these factors to interact, thus their ability to regulate the transcription of the SSAT gene is lost. PMID- 11256948 TI - The Escherichia coli CcmG protein fulfils a specific role in cytochrome c assembly. AB - In Escherichia coli K-12, c-type cytochromes are synthesized only during anaerobic growth with trimethylamine-N-oxide, nitrite or low concentrations of nitrate as the terminal electron acceptor. A thioredoxin-like protein, CcmG, is one of 12 proteins required for their assembly in the periplasm. Its postulated function is to reduce disulphide bonds formed between correctly paired cysteine residues in the cytochrome c apoproteins prior to haem attachment by CcmF and CcmH. We report that loss of CcmG synthesis by mutation was not compensated by a second mutation in disulphide-bond-forming proteins, DsbA or DsbB, or by the chemical reductant, 2-mercaptoethanesulphonic acid. An anti-CcmG polyclonal antibody was used in Western-blot analysis to probe the redox state of CcmG in mutants defective in the synthesis of other proteins essential for cytochrome c assembly. The oxidized form of CcmG accumulated not only in trxA or dipZ mutants defective in the transfer of electrons from the cytoplasm for disulphide isomerization and reduction reactions in the periplasm, but also in ccmF and ccmH mutants. The requirement of both CcmF and CcmH for the reduction of the disulphide bond in CcmG indicates that CcmG functions later than CcmF and CcmH in cytochrome c assembly, rather than in electron transfer from the membrane associated DipZ (also known as DsbD) to CcmH. The data support a model proposed by others in which CcmG catalyses one of the last reactions specific to cytochrome c assembly. PMID- 11256949 TI - Selective recognition of inositol phosphates by subtypes of the inositol trisphosphate receptor. AB - Synthetic analogues of inositol trisphosphate (IP(3)), all of which included structures equivalent to the 4,5-bisphosphate of (1,4,5)IP(3), were used to probe the recognition properties of rat full-length type 1, 2 and 3 IP(3) receptors expressed in insect Spodoptera frugiperda 9 cells. Using equilibrium competition binding with [(3)H](1,4,5)IP(3) in Ca(2+)-free cytosol-like medium, the relative affinities of the receptor subtypes for (1,4,5)IP(3) were type 3 (K(d)=11+/-2 nM)>type 2 (K(d)=17+/-2 nM)>type 1 (K(d)=24+/-4 nM). (1,4,5)IP(3) binding was reversibly stimulated by increased pH, but the subtypes differed in their sensitivity to pH (type 1>type 2>type 3). For all three subtypes, the equatorial 6-hydroxy group of (1,4,5)IP(3) was essential for high-affinity binding, the equatorial 3-hydroxy group significantly improved affinity, and the axial 2 hydroxy group was insignificant; a 1-phosphate (or in its absence, a 2-phosphate) improved binding affinity. The subtypes differed in the extents to which they tolerated inversion of the 3-hydroxy group of (1,4,5)IP(3) (type 1>type 2>type 3), and this probably accounts for the selectivity of (1,4,6)IP(3) for type 1 receptors. They also differed in their tolerance of inversion, removal or substitution (by phosphate) of the 2-hydroxy group (types 2 and 3>type 1), hence the selectivity of (1,2,4,5)IP(4) for type 2 and 3 receptors. Removal of the 3 hydroxy group or its replacement by fluorine or CH(2)OH was best tolerated by type 3 receptors, and accounts for the selectivity of 3-deoxy(1,4,5)IP(3) for type 3 receptors. Our results provide the first systematic analysis of the recognition properties of IP(3) receptor subtypes and have identified the 2- and 3-positions of (1,4,5)IP(3) as key determinants of subtype selectivity. PMID- 11256951 TI - Long-lived glycosyl-enzyme intermediate mimic produced by formate re-activation of a mutant endoglucanase lacking its catalytic nucleophile. AB - The mutant E134A 1,3-1,4-beta-glucanase from Bacillus licheniformis, in which the catalytic nucleophilic residue has been removed by mutation to alanine, has its hydrolytic activity rescued by exogenous formate in a concentration-dependent manner. A long-lived alpha-glycosyl formate is detected and identified by (1)H NMR and matrix-assisted laser desorption ionization-time-of-flight-MS. The intermediate is kinetically competent, since it is, at least partially, enzymically hydrolysed, and able to act as a glycosyl donor in transglycosylation reactions. This transient compound represents a true covalent glycosyl-enzyme intermediate mimic of the proposed covalent intermediate in the reaction mechanism of retaining glycosidases. PMID- 11256950 TI - Xenobiotic induction of cytochrome P450 2B1 (CYP2B1) is mediated by the orphan nuclear receptor constitutive androstane receptor (CAR) and requires steroid co activator 1 (SRC-1) and the transcription factor Sp1. AB - The constitutive androstane receptor (CAR) activates the expression of a reporter gene attached to the phenobarbital-response element (PBRE) of the cytochrome P450 2B1 (CYP2B1) gene in response to the barbiturate phenobarbital and the plant product picrotoxin. The xenobiotic-mediated increase in transactivation occurs in transfected primary hepatocytes and in liver transfected by biolistic-particle mediated DNA transfer, but not in the transformed cell lines HepG2, CV-1 and HeLa, which support only constitutive activation of gene expression by CAR. Steroid co-activator 1 (SRC-1) enhances both constitutive and xenobiotic-induced CAR-mediated transactivation via the CYP2B1 PBRE in transfected primary hepatocytes. The nuclear receptor 1 (NR1) site of the PBRE is sufficient for CAR mediated transactivation, but additional sequences within the PBRE, and hence the proteins that bind to them, are required for the interaction of CAR with SRC-1. The NR2 site of the PBRE binds proteins other than CAR, including an unidentified nuclear receptor heterodimerized with retinoid X receptor alpha. By binding to the proximal promoter of CYP2B1, the transcription factor Sp1 increases both basal transcription and xenobiotic-induced expression via the PBRE. Thus induction of CYP2B1 expression by xenobiotics is mediated by the nuclear receptor CAR and, for optimal expression, requires SRC-1 and Sp1. PMID- 11256952 TI - Nitric oxide and cGMP activate Ca2+-release processes in rat parotid acinar cells. AB - We characterized the enzymic properties of ADP-ribosyl cyclase in rat parotid acinar cells by using a fluorescence technique. ADP-ribosyl cyclase is capable of synthesizing the Ca2+ -mobilizing nucleotide cADP-ribose (cADPR) from NAD(+) and has previously been shown to be regulated by cGMP via a cGMP-dependent protein kinase (G kinase). We therefore investigated whether NO/cGMP-activated pathways are present in rat parotid acinar cells and whether NO/cGMP signalling exerts control over cellular Ca2+ signalling processes. Our results showed that stimulation of acinar cells with adrenaline, isoproterenol, substance P and NO resulted in a rise in the [cGMP]. In addition, NO induced a release of Ca2+ from intracellular ryanodine-sensitive stores via a cGMP/G-kinase-mediated process. Thus our data reveal that a rise in [cGMP], caused by either neurotransmitter or NO activation, activates a G kinase, which in turn controls Ca2+ release from ryanodine-sensitive stores. Since parotid acinar cells possess ADP-ribosyl cyclase activity, we propose a model in which cADPR is the link between NO/cGMP signalling pathways and release of Ca2+ from ryanodine-sensitive stores. PMID- 11256953 TI - Enzymic characterization of epidermis-derived 12-lipoxygenase isoenzymes. AB - Substrate selectivity and other enzymic characteristics of two epidermis-derived lipoxygenases (LOXs), the epidermis-type (e) (12S)-LOX and (12R)-LOX, were compared with those of the platelet-type (p) (12S)-LOX. In contrast with p(12S) LOX, e(12S)-LOX and (12R)-LOX exhibited no or very low reactivity towards the customary substrates linoleic acid and arachidonic acid but metabolized the corresponding fatty acid methyl esters, which, in contrast, were not accepted as substrates by p(12S)-LOX. Other esters of arachidonic acid and linoleic acid, including propan-2-yl and cholesterol esters, 1-palmitoyl-2-arachidonyl-sn glycero-3-phosphocholine, 1-palmitoyl-2-linoleyl-sn-glycero-3 phosphoethanolamine, and ceramide 1 carrying an omega-linoleic acid ester, were not metabolized by these three LOX isoenzymes. Among various polyunsaturated fatty acids the isomeric eicosatrienoic acids were found to be oxygenated by e(12S)-LOX but not by (12R)-LOX. 4,7,10,13,16,19-Docosahexaenoic acid as a substrate was restricted to p(12S)-LOX. Variations in the pH and the Ca(2+) content of the incubation medium affected the catalytic potential only slightly. Whereas (12R)-LOX activity increased in the presence of Ca(2+) and with an acidic pH, Ca(2+) had no effect on p(12S)-LOX and e(12S)-LOX; an acidic pH decreased the catalytic activity of the latter two. However, the catalytic activity of the epidermis-type isoenzymes, but not of p(12S)-LOX, was found to be markedly increased in the presence of DMSO. Under these conditions, e(12S)-LOX and (12R) LOX oxygenated 4,7,10,13,16,19-docosahexaenoic acid to 14-hydroxy-4,7,10,12,16,19 docosahexaenoic acid and 13-hydroxy-4,7,10,14,16,19-docosahexaenoic acid respectively. In addition, (9R)-hydroxyoctadeca-10,12-dienoic acid methyl ester was generated from linoleic acid methyl ester by (12R)-LOX. Independently of the substrate, the catalytic activity of e(12S)-LOX and (12R)-LOX was always at most 2% of that of p(12S)-LOX with arachidonic acid as substrate. PMID- 11256954 TI - Differential regulation of expression of genes encoding uncoupling proteins 2 and 3 in brown adipose tissue during lactation in mice. AB - Thermogenic activity in brown adipose tissue (BAT) decreases during lactation; the down-regulation of the gene encoding uncoupling protein 1 (UCP1) is involved in this process. Our studies show that UCP2 mRNA expression does not change during the breeding cycle in mice. In contrast, UCP3 mRNA is down-regulated in lactation but it recovers after weaning, in parallel with UCP1 mRNA. This leads to a decrease in the content of UCP3 in BAT mitochondria during lactation. Lowering the energy-sparing necessities of lactating dams by decreasing litter size or feeding with a high-fat diet prevented the down-regulation of UCP1 mRNA and UCP3 mRNA. In most cases this resulted in a less marked decrease in UCP1 and UCP3 protein in BAT mitochondria owing to lactation. Fasting for 24 h caused a different response in UCP1 and UCP3 mRNA expression: it decreased UCP1 mRNA levels but had no effect on UCP3 mRNA abundance in virgin mice; it even increased UCP3 mRNA expression in lactating dams. These changes did not lead to modifications in UCP1 or UCP3 protein abundance. Whereas acute treatment with peroxisome-proliferator-activated receptor (PPAR)alpha and PPARgamma agonists increased UCP1 mRNA levels only in lactating dams, UCP3 mRNA expression was induced by both kinds of PPAR activator in lactating dams and by PPARalpha agonists in virgin mice. It is concluded that modifications of UCP2 mRNA levels are not part of the physiological adaptations taking place in BAT during lactation. In contrast, the down-regulation of UCP3 mRNA expression and mitochondrial UCP3 content is consistent with a role for the gene encoding UCP3 in the decrease in metabolic fuel oxidation and thermogenesis in BAT during lactation. PMID- 11256955 TI - Characterization of a novel phosphatidylinositol 3-phosphate-binding protein containing two FYVE fingers in tandem that is targeted to the Golgi. AB - We have identified a novel protein of predicted molecular mass 40 kDa that contains two FYVE domains in tandem and has therefore been named TAFF1 (TAndem FYVE Fingers-1). The protein is expressed predominantly in heart and binds to PtdIns3P specifically, even though the FYVE domains in TAFF1 lacks the first Arg of the consensus sequence R(K/R)HHCR, critical for the PtdIns3P binding of other FYVE domains identified so far. The first Arg is replaced by a Thr and Ser in the N-terminal and C-terminal FYVE domains of TAFF1 respectively. Mutational analysis indicates that both FYVE domains are required for high affinity binding to PtdIns3P. Cell localization studies using a green fluorescent protein fusion show that TAFF1 is localized to the Golgi, and that the Golgi targeting sequence is located within the N-terminal 187 residues and not in either FYVE domain. PMID- 11256956 TI - Proinsulin C-peptide rapidly stimulates mitogen-activated protein kinases in Swiss 3T3 fibroblasts: requirement of protein kinase C, phosphoinositide 3-kinase and pertussis toxin-sensitive G-protein. AB - It has been demonstrated that proinsulin C-peptide possesses several biological activities and that its specific binding sites are present on the surface of cell membranes. However, the molecular and cellular mechanisms of C-peptide actions are poorly known. In the present study we examined the possible involvement of the mitogen-activated protein kinase (MAPK) pathway in C-peptide effects. C peptide induced the phosphorylation of MAPK [p44 extracellular signal-regulated kinase 1 (ERK1) and p42 ERK2] in Swiss 3T3 and 3T3-F442A fibroblasts but not in 3T3-L1 fibroblasts and some other cell lines such as L(6)E(9) muscle cells. In Swiss 3T3 cells, C-peptide-induced phosphorylation of MAPK was dependent on time and concentration, being maximal at 1 min and at 1 nM C-peptide and was accompanied by an increase in MAPK activity and MAPK kinase (MEK) phosphorylation. The MAPK phosphorylation by C-peptide was abolished by treatment with pertussis toxin (PTX) and also with a MEK inhibitor, PD 98059. In addition, MAPK phosphorylation was attenuated by treatment with a phosphoinositide 3-kinase (PI-3K) inhibitor, wortmannin, and with a protein kinase C (PKC) inhibitor, GF109203X, and by down-regulation of PKC by prolonged treatment with PMA. Similar effects of the inhibitors and PTX were found on the MAPK phosphorylation induced by neuropeptide Y. These results suggest that C-peptide activates MAPK through a putative G(i)/G(o)-linked receptor for C-peptide and through PI-3K-dependent and PKC-dependent pathways. PMID- 11256957 TI - Protein-coenzyme interactions in adenosylcobalamin-dependent glutamate mutase. AB - Glutamate mutase catalyses an unusual isomerization involving free-radical intermediates that are generated by homolysis of the cobalt-carbon bond of the coenzyme adenosylcobalamin (coenzyme B(12)). A variety of techniques have been used to examine the interaction between the protein and adenosylcobalamin, and between the protein and the products of coenzyme homolysis, cob(II)alamin and 5' deoxyadenosine. These include equilibrium gel filtration, isothermal titration calorimetry, and resonance Raman, UV-visible and EPR spectroscopies. The thermodynamics of adenosylcobalamin binding to the protein have been examined and appear to be entirely entropy-driven, with DeltaS=109 J.mol(-1).K(-1). The cobalt carbon bond stretching frequency is unchanged upon coenzyme binding to the protein, arguing against a ground-state destabilization of the cobalt-carbon bond of adenosylcobalamin by the protein. However, reconstitution of the enzyme with cob(II)alamin and 5'-deoxyadenosine, the two stable intermediates formed subsequent to homolysis, results in the blue-shifting of two of the bands comprising the UV-visible spectrum of the corrin ring. The most plausible interpretation of this result is that an interaction between the protein, 5' deoxyadenosine and cob(II)alamin introduces a distortion into the ring corrin that perturbs its electronic properties. PMID- 11256958 TI - Control of Ca2+ influx in human neutrophils by inositol 1,4,5-trisphosphate (IP3) binding: differential effects of micro-injected IP3 receptor antagonists. AB - Neutrophils signal Ca2+ changes in response to occupancy of G-protein-linked receptors such as the formylated peptide receptor. This Ca2+ signal is composed of two parts, inositol 1,4,5-trisphosphate (IP3)-triggered release of Ca2+ from an intracellular store and Ca2+ influx. In order to probe the relationship between these events, cytosolic free Ca2+ changes in neutrophils were monitored after micro-injection of agents which inhibit IP3 binding. Micro-injection of heparin into neutrophils totally inhibited both formylmethionyl leucylphenylalanine-induced Ca2+ release and the subsequent Ca2+ influx. This effect was not due to prior depletion of Ca2+ stores. Furthermore, micro injection with anti-IP3-receptor antibody also inhibited Ca2+ release. However, anti-IP3-receptor antibody and another high-molecular-mass IP3-binding antagonist, heparin-albumin conjugate, failed to inhibit the accompanying Ca2+ influx. It was concluded that two IP3-binding sites exist in neutrophils: one accessible by both heparin and the high-molecular-mass inhibitors of IP3 binding and responsible for Ca2+ release, and another inaccessible to high-molecular-mass molecules and responsible for Ca2+ influx. PMID- 11256959 TI - Oxidative modification of H-ras: S-thiolation and S-nitrosylation of reactive cysteines. AB - The reactive cysteines in H-ras are subject to oxidative modifications that potentially alter the cellular function of this protein. In this study, purified H-ras was modified by thiol oxidants such as hydrogen peroxide (H(2)O(2)), S nitrosoglutathione, diamide, glutathione disulphide (GSSG) and cystamine, producing as many as four charge-isomeric forms of the protein. These results suggest that all four reactive cysteines of H-ras are potential sites of regulatory modification reactions. S-nitrosylated and S-glutathiolated forms of H ras were identified by protocols that depend on separation of alkylated proteins on electrofocusing gels. S-nitrosoglutathione could S-nitrosylate H-ras on four cysteine residues, while reduced glutathione (GSH) and H(2)O(2) mediate S glutathiolation on at least one cysteine of H-ras. Either GSSG or diamide S glutathiolated at least two cysteine residues of purified H-ras. Iodoacetic acid reacts with three cysteine residues. In intact NIH-3T3 cells, wild-type H-ras was S-glutathiolated by diamide. Similarly, cells expressing a C118S mutant or a C181S/C184S double mutant of H-ras were S-glutathiolated by diamide. These results suggest that H-ras can be S-glutathiolated on multiple thiols in vivo and that at least one of these thiols is normally lipid-modified. In cells treated with S-nitrosocysteine, evidence for both S-nitrosylated and S-glutathiolated H ras was obtained and S-nitrosylation was the predominant modification. These results show that oxidative modification of H-ras can be extensive in vivo, that both S-nitrosylated and S-glutathiolated forms may be important, and that oxidation may occur on reactive cysteines that are normally targeted for lipid modification reactions. PMID- 11256960 TI - Pectate lyase 10A from Pseudomonas cellulosa is a modular enzyme containing a family 2a carbohydrate-binding module. AB - Pectate lyase 10A (Pel10A) enzyme from Pseudomonas cellulosa is composed of 649 residues and has a molecular mass of 68.5 kDa. Sequence analysis revealed that Pel10A contained a signal peptide and two serine-rich linker sequences that separate three modules. Sequence similarity was seen between the 9.2 kDa N terminal module of Pel10A and family 2a carbohydrate-binding modules (CBMs). This N-terminal module of Pel10A was shown to encode an independently functional module with affinity to crystalline cellulose. A high sequence identity of 66% was seen between the 14.2 kDa central module of Pel10A and the functionally uncharacterized central modules of the xylan-degrading enzymes endoxylanase 10B, arabinofuranosidase 62C and esterase 1D, also from P. cellulosa. The 35.8 kDa C terminal module of Pel10A was shown to have 30 and 36% identities with the family 10 pectate lyases from Azospirillum irakense and an alkaliphilic strain of Bacillus sp. strain KSM-P15, respectively. This His-tagged C-terminal module of the Pel10A was shown to encode an independent catalytic module (Pel10Acm). Pel10Acm was shown to cleave pectate and pectin in an endo-fashion and to have optimal activity at pH 10 and in the presence of 2 mM Ca2+. Highest enzyme activity was detected at 62 degrees C. Pel10Acm was shown to be most active against pectate (i.e. polygalacturonic acid) with progressively less activity against 31, 67 and 89% esterified citrus pectins. These data suggest that Pel10A has a preference for sequences of non-esterified galacturonic acid residues. Significantly, Pel10A and the P. cellulosa rhamnogalacturonan lyase 11A, in the accompanying article [McKie, Vincken, Voragen, van den Broek, Stimson and Gilbert (2001) Biochem. J. 355, 167-177], are the first CBM-containing pectinases described to date. PMID- 11256961 TI - A new family of rhamnogalacturonan lyases contains an enzyme that binds to cellulose. AB - Pseudomonas cellulosa is an aerobic bacterium that synthesizes an extensive array of modular cellulases and hemicellulases, which have a modular architecture consisting of catalytic domains and distinct non-catalytic carbohydrate-binding modules (CBMs). To investigate whether the main-chain-cleaving pectinases from this bacterium also have a modular structure, a library of P. cellulosa genomic DNA, constructed in lambdaZAPII, was screened for pectinase-encoding sequences. A recombinant phage that attacked arabinan, galactan and rhamnogalacturonan was isolated. The encoded enzyme, designated Rgl11A, had a modular structure comprising an N-terminal domain that exhibited homology to Bacillus and Streptomyces proteins of unknown function, a middle domain that exhibited sequence identity to fibronectin-3 domains, and a C-terminal domain that was homologous to family 2a CBMs. Expression of the three modules of the Pseudomonas protein in Escherichia coli showed that its C-terminal module was a functional cellulose-binding domain, and the N-terminal module consisted of a catalytic domain that hydrolysed rhamnogalacturonan-containing substrates. The activity of Rgl11A against apple- and potato-derived rhamnogalacturonan substrates indicated that the enzyme had a strong preference for rhamnogalacturonans that contained galactose side chains, and which were not esterified. The enzyme had an absolute requirement for calcium, a high optimum pH, and catalysis was associated with an increase in absorbance at 235 nm, indicating that glycosidic bond cleavage was mediated via a beta-elimination mechanism. These data indicate that Rgl11A is a rhamnogalacturonan lyase and, together with the homologous Bacillus and Streptomyces proteins, comprise a new family of polysaccharide lyases. The presence of a family 2a CBM in Rgl11A, and in a P. cellulosa pectate lyase described in the accompanying paper [Brown, Mallen, Charnock, Davies and Black (2001) Biochem. J. 355, 155-165] suggests that the capacity to bind cellulose plays an important role in the activity of main-chain-cleaving Pseudomonas pectinases, in addition to cellulases and hemicellulases. PMID- 11256962 TI - Exonic Sp1 sites are required for neural-specific expression of the glycine receptor beta subunit gene. AB - The gene encoding the beta subunit of the inhibitory glycine receptor (GlyR) is widely expressed throughout the mammalian central nervous system. To unravel the elements regulating its transcription, we isolated its 5' non-coding and upstream flanking regions from mouse. Sequence analysis revealed significant differences between the 5' region of the beta subunit gene and the corresponding regions of the homologous GlyR alpha subunit genes; it also identified a novel exon (exon 0) that encodes most of the 5'-untranslated portion of the GlyR beta mRNA. Primer extension experiments disclosed multiple transcriptional start sites. Transfection experiments with luciferase reporter gene constructs showed that sequences encompassing 1.58 kb of upstream flanking region and 180 bp of exon 0 displayed high promoter activity in two neuroblastoma cell lines but not in non neural cells. Analysis of various deletion constructs showed that the 5' flanking region preceding the transcriptional start sites silences expression in non neural cells but is not essential for general promoter activity. In contrast, the deletion of sequences within exon 0 drastically decreased or abolished transcription; the removal of sequences harbouring Sp1 consensus sequences within exon 0 decreased expression specifically in a neuroblastoma cell line. Band-shift assays confirmed the binding of Sp1 to sites within the deleted sequence. Our results indicate that neural-specific expression of the GlyR beta subunit gene might depend on a direct interaction of Sp1 transcription factors with cis elements located downstream from transcription initiation sites. PMID- 11256963 TI - Sphingosylphosphocholine is a naturally occurring lipid mediator in blood plasma: a possible role in regulating cardiac function via sphingolipid receptors. AB - Blood plasma and serum contain factors that activate inwardly rectifying GIRK1/GIRK4 K+ channels in atrial myocytes via one or more non-atropine-sensitive receptors coupled to pertussis-toxin-sensitive G-proteins. This channel is also the target of muscarinic M(2) receptors activated by the physiological release of acetylcholine from parasympathetic nerve endings. By using a combination of HPLC and TLC techniques with matrix-assisted laser desorption ionization-time-of flight MS, we purified and identified sphingosine 1-phosphate (SPP) and sphingosylphosphocholine (SPC) as the plasma and serum factors responsible for activating the inwardly rectifying K+ channel (I(K)). With the use of MS the concentration of SPC was estimated at 50 nM in plasma and 130 nM in serum; those concentrations exceeded the 1.5 nM EC(50) measured in guinea-pig atrial myocytes. With the use of reverse-transcriptase-mediated PCR and/or Western blot analysis, we detected Edg1, Edg3, Edg5 and Edg8 as well as OGR1 sphingolipid receptor transcripts and/or proteins. In perfused guinea-pig hearts, SPC exerted a negative chronotropic effect with a threshold concentration of 1 microM. SPC was completely removed after perfusion through the coronary circulation at a concentration of 10 microM. On the basis of their constitutive presence in plasma, the expression of specific receptors, and a mechanism of ligand inactivation, we propose that SPP and SPC might have a physiologically relevant role in the regulation of the heart. PMID- 11256964 TI - Effect of polyamine depletion on caspase activation: a study with spermine synthase-deficient cells. AB - Activation of the caspase proteases represents a central point in apoptosis. The requirement for spermine for the processes leading to caspase activation has been studied in transformed embryonic fibroblasts obtained from gyro (Gy) mutant male mice. These cells lack spermine synthase activity and thus provide a valuable model to study the role of spermine in cell processes. Gy fibroblasts do not contain spermine and have a higher spermidine content. However, when compared with fibroblasts obtained from normal male littermates (N cells), Gy fibroblasts were observed to grow normally. The lack of spermine did not affect the expression of Bcl-2, and caspases 3 and 9 were activated by etoposide in both N and Gy cells, indicating that spermine is dispensable for caspase activation. Spermine deficiency did not significantly influence caspase activity in cells treated with etoposide, cycloheximide or staurosporine, but sensitized the cells to UV irradiation, which triggered significantly higher caspase activity in Gy cells compared with N cells. alpha-Difluoromethylornithine (DFMO), an inhibitor of polyamine synthesis that is able to deplete cells of putrescine and spermidine, but usually does not influence spermine content, was able to produce a more complete polyamine depletion in Gy cells. This depletion, which included spermine deficiency, dramatically increased caspase activation and cell death in Gy fibroblasts exposed to UV irradiation. On the other hand, in either N or Gy cells, DFMO treatment did not influence caspase activity triggered by staurosporine, but inhibited it when the inducers were cycloheximide or etoposide. In Gy cells depleted of polyamines by DFMO, polyamine replenishment with either spermidine or spermine was sufficient to restore caspase activity induced by etoposide, indicating that, in this model, polyamines have an interchangeable role in supporting caspase activation. Therefore, spermine is not required for such activation, and the effect and specificity of polyamine depletion on caspase activity may be very different, depending on the role of polyamines in the specific death pathways engaged by different stimuli. Some inducers of apoptosis, for example etoposide, absolutely require polyamines for caspase activation, yet the lack of polyamines, particularly spermine, strongly increases caspase activation when induced by UV irradiation. PMID- 11256965 TI - Enhancement of macrophage survival and DNA synthesis by oxidized-low-density lipoprotein (LDL)-derived lipids and by aggregates of lightly oxidized LDL. AB - Human atherosclerotic plaque contains a partially characterized range of normal and oxidized lipids formed mainly from free and esterified cholesterol and phospholipids, some of which can be located in macrophage-derived "foam" cells. Oxidation of low-density lipoprotein (LDL) is often considered as an important event leading to subsequent foam-cell development, which may also include enhanced cell survival and/or proliferation. The active component(s) in oxidized LDL (ox.LDL) causing macrophage proliferation is debated. We report here that the lipid component of ox.LDL can promote macrophage survival and DNA synthesis, the latter response showing a synergistic effect in the presence of low concentrations of macrophage colony-stimulating factor. 7-Ketocholesterol showed some stimulation of macrophage DNA synthesis whereas hypochlorite-oxidized (i.e. apolipoprotein B-oxidized) LDL did not. Plaque-derived lipids could enhance macrophage survival. It has not been proven that LDL in lesions is oxidized sufficiently to be the dominant source of sterols in vivo or to be able to induce macrophage growth in vitro or in vivo; it has been suggested that aggregation of modified LDL in vivo is an important step in the deposition of intracellular lipid. We found that aggregation of lightly oxidized LDL potentiated dramatically its ability to stimulate macrophage DNA synthesis, indicating that extensive oxidation of LDL is not required for this response in vitro and perhaps in vivo. PMID- 11256967 TI - Structural and functional similarities between the central eukaryotic initiation factor (eIF)4A-binding domain of mammalian eIF4G and the eIF4A-binding domain of yeast eIF4G. AB - The translation eukaryotic initiation factor (eIF)4G of the yeast Saccharomyces cerevisiae interacts with the RNA helicase eIF4A (a member of the DEAD-box protein family; where DEAD corresponds to Asp-Glu-Ala-Asp) through a C-terminal domain in eIF4G (amino acids 542-883). Mammalian eIF4G has two interaction domains for eIF4A, a central domain and a domain close to the C-terminus. This raises the question of whether eIF4A binding to eIF4G is conserved between yeast and mammalian cells or whether it is different. We isolated eIF4G1 mutants defective in eIF4A binding and showed that these mutants are strongly impaired in translation and growth. Extracts from mutants displaying a temperature-sensitive phenotype for growth have low in vitro translation activity, which can be restored by addition of the purified eIF4G1-eIF4E complex, but not by eIF4E alone. Analysis of mutant eIF4G(542-883) proteins defective in eIF4A binding shows that the interaction of yeast eIF4A with eIF4G1 depends on amino acid motifs that are conserved between the yeast eIF4A-binding site and the central eIF4A-binding domain of mammalian eIF4G. We show that mammalian eIF4A binds tightly to yeast eIF4G1 and, furthermore, that mutant yeast eIF4G(542-883) proteins, which do not bind yeast eIF4A, do not interact with mammalian eIF4A. Despite the conservation of the eIF4A-binding site in eIF4G and the strong sequence conservation between yeast and mammalian eIF4A (66% identity; 82% similarity at the amino acid level) mammalian eIF4A does not substitute for the yeast factor in vivo and is not functional in a yeast in vitro translation system. PMID- 11256966 TI - Recombinant soluble betaglycan is a potent and isoform-selective transforming growth factor-beta neutralizing agent. AB - Betaglycan is an accessory receptor of members of the transforming growth factor beta (TGF-beta) superfamily, which regulates their actions through ligand dependent interactions with type II receptors. A natural soluble form of betaglycan is found in serum and extracellular matrices. Soluble betaglycan, prepared as a recombinant protein using the baculoviral expression system, inhibits the actions of TGF-beta. Because of its potential use as an anti-TGF beta therapeutic agent, we have purified and characterized baculoviral recombinant soluble betaglycan. Baculoviral soluble betaglycan is a homodimer formed by two 110 kDa monomers associated by non-covalent interactions. This protein is devoid of glycosaminoglycan chains, although it contains the serine residues, which, in vertebrate cells, are modified by these carbohydrates. On the other hand, mannose-rich carbohydrates account for approximately 20 kDa of the mass of the monomer. End-terminal sequence analysis of the soluble betaglycan showed that Gly(24) is the first residue of the mature protein. Similarly to the natural soluble betaglycan, baculoviral soluble betaglycan has an equilibrium dissociation constant (K(d)) of 3.5 nM for TGF-beta1. Ligand competition assays indicate that the relative affinities of recombinant soluble betaglycan for the TGF-beta isoforms are TGF-beta2>TGF-beta3>TGF-beta1. The anti-TGF-beta potency of recombinant soluble betaglycan in vitro is 10-fold higher for TGF-beta2 than for TGF-beta1. Compared with a commercial pan-specific anti-TGF-beta neutralizing antibody, recombinant soluble betaglycan is more potent against TGF-beta2 and similar against TGF-beta1. These results indicate that baculoviral soluble betaglycan has the biochemical and functional properties that would make it a suitable agent for the treatment of the diseases in which excess TGF-beta plays a central physiopathological role. PMID- 11256968 TI - The mitochondrial consequences of uncoupling intact cells depend on the nature of the exogenous substrate. AB - In isolated mitochondria the consequences of oxidative phosphorylation uncoupling are well defined, whereas in intact cells various effects have been described. Uncoupling liver cells with 2,4-dinitrophenol (DNP) in the presence of dihydroxyacetone (DHA) and ethanol results in a marked decrease in mitochondrial transmembrane electrical potential (DeltaPsi), ATP/ADP ratios and gluconeogenesis (as an ATP-utilizing process), whereas the increased oxidation rate is limited and transient. Conversely, when DHA is associated with octanoate or proline, DNP addition results in a very large and sustained increase in oxidation rate, whereas the decreases in DeltaPsi, ATP/ADP ratios and gluconeogenesis are significantly less when compared with DHA and ethanol. Hence significant energy wastage (high oxidation rate) by uncoupling is achieved only with substrates that are directly oxidized in the mitochondrial matrix. Conversely in the presence of substrates that are first oxidized in the cytosol, uncoupling results in a profound decrease in mitochondrial DeltaPsi and ATP synthesis, whereas energy wastage is very limited. PMID- 11256970 TI - The efficiency of N-linked glycosylation of bovine DNase I depends on the Asn-Xaa Ser/Thr sequence and the tissue of origin. AB - Bovine DNase I contains two potential N-linked glycosylation sites with the sequences Asn(18)-Ala-Thr and Asn(106)-Asp-Ser. A previous report established that pancreatic DNase I has only one sugar chain at Asn(18) [Liao, Salnikow, Moore and Stein (1973) J. Biol. Chem. 248, 1489-1495]. We found, however, that bovine DNase I expressed in COS-1 cells was glycosylated about 70% at Asn(106) in addition to being completely glycosylated at Asn(18). Glycosylation of Asn(106) increased to 97% when Asp(107) was mutated to Glu or when Ser(108) was mutated to Thr. Mutation of Asp(107) to Trp had no effect, whereas a substitution with Pro at this position abolished glycosylation of Asn(106). Analysis of the state of glycosylation of DNase I purified from a variety of bovine tissues revealed that DNase I from spleen, submaxillary gland, lung and adrenal had two sugar chains, whereas enzyme from pancreas and kidney had only one sugar chain. These findings demonstrate a major difference in the ability of various tissues to utilize N linked glycosylation signals that contain suboptimal residues in the second and third positions. PMID- 11256971 TI - C-reactive protein: a central player in atherogenesis or an epiphenomenon? PMID- 11256969 TI - (S)-preferential detoxification of 4-hydroxy-2(E)-nonenal enantiomers by hepatic glutathione S-transferase isoforms in guinea-pigs and rats. AB - In guinea-pig liver cytosol, racemic 4-hydroxy-2(E)-nonenal (HNE), a reactive and highly toxic product released from biomembranes by lipid peroxidation, was detoxified (S)-preferentially by GSH conjugation mediated by glutathione S transferases (GSTs) and (R)-preferentially by NAD(+)-dependent oxidation mediated by aldehyde dehydrogenase (ALDH). The GST-mediated detoxification of the HNE enantiomers proceeded at much higher rates than that mediated by ALDH in guinea pig liver cytosol. All the major guinea-pig GSTs, A1-1, M1-1, M1-2 and M1-3*, isolated from guinea-pig liver cytosol also catalysed the (S)-preferential conjugation of the HNE enantiomers. The liver and other major tissues of guinea pigs had no immunologically detectable level of a putative GSTA4-4 orthologue, which exists as a minor GST protein in rat, mouse and human livers and exhibits extremely high catalytic activity towards HNE. All the hepatic rat GSTs, A1-1(2), A1-3, A4-4, M1-1, M1-2 and M2-2, also catalysed the (S)-preferential conjugation of HNE enantiomers. PMID- 11256972 TI - Roles of chemokines in the regulation of leucocyte recruitment. AB - In the search for new treatments for human inflammatory disease, antagonism of chemokine receptors by small molecules is an attractive goal. Although there are overlapping patterns of expression of chemokine receptors between leucocyte types, an investigation of the chemokine responsiveness of cells important in allergic inflammation, such as the eosinophil and the basophil, is beginning to uncover how selective recruitment may be regulated. The story of the eotaxin receptor, CCR3, and its central role in allergic inflammation illustrates that therapeutic antagonism of these pathways is imminently achievable. PMID- 11256973 TI - The bile acid taurocholate impairs rat cardiomyocyte function: a proposed mechanism for intra-uterine fetal death in obstetric cholestasis. AB - Obstetric cholestasis is a liver disease of pregnancy that can be complicated by sudden, hitherto unexplained, intra-uterine fetal death. Because intra-uterine death occurs suddenly, and because fetal heart rate abnormalities have been reported in obstetric cholestasis, we hypothesized that intra-uterine death is caused by impaired fetal cardiomyocyte function, resulting in fetal cardiac arrest. Obstetric cholestasis is associated with raised levels of maternal and fetal serum bile acids, and we propose that these may alter cardiomyocyte function. It was not possible to investigate the effects of bile acids on the intact human fetal heart at a cellular level. Therefore we used the closest available model of fetal myocardium at term: a primary culture of neonatal rat cardiomyocytes in which cells beat synchronously and develop pacemaker activity. The effect of the primary bile acid taurocholate (0.3 mM and 3 mM) on cultures of single cardiomyocytes, each with its own independent rate of contraction, was a reversible decrease in the rate of contraction and in the proportion of beating cells (P < 0.001). Addition of taurocholate to a network of synchronously beating cells caused a similar decrease in the rate of contraction. Furthermore, the integrity of the network was destroyed, and cells ceased to beat synchronously. Taurocholate also resulted in altered calcium dynamics and loss of synchronous beating. These data suggest that raised levels of the bile acid taurocholate in the fetal serum in obstetric cholestasis may result in the development of a fetal dysrhythmia and in sudden intra-uterine death. PMID- 11256974 TI - Relationship between C-reactive protein and intima-media thickness in the carotid and femoral arteries and to antibodies against oxidized low-density lipoprotein in healthy men: the Atherosclerosis and Insulin Resistance (AIR) study. AB - Results from several recent reports have linked high serum C-reactive protein (CRP) levels to atherosclerotic disease and its complications. The aims of the present study were to investigate the relationship between CRP levels and subclinical atherosclerosis, as measured by ultrasound in the carotid and femoral arteries; and also to examine whether CRP levels are associated with antibodies to oxidized low-density lipoprotein (Ox-LDL). The study group (n = 391) consisted of clinically healthy 58-year-old men recruited from the general population. CRP and antibody titres to Ox-LDL were measured by ELISA. The results showed an association between CRP and ultrasound-assessed subclinical atherosclerosis in the femoral artery (r = 0.14, P = 0.010), and also between CRP and systolic blood pressure, diastolic blood pressure, heart rate, triglycerides, high-density lipoprotein, body mass index, waist-to-hip ratio (WHR), blood glucose, cigarette years and antibody titres to ox-LDL (r = 0.19, P < 0.001). In this clinically healthy population of 58-year-old men, CRP levels were associated with both intima-media thickness and plaque occurrence in the femoral artery. The association between CRP and femoral atherosclerosis was not independent of smoking, serum LDL cholesterol, or systolic blood pressure. CRP levels were independently related to abdominal obesity measured as WHR, smoking and antibody titres to Ox-LDL. PMID- 11256975 TI - Improvement in intestinal permeability precedes morphometric recovery of the small intestine in coeliac disease. AB - It is often difficult to assess small bowel recovery in adults with coeliac disease on a gluten-free diet (GFD). This prospective study compares changes in intestinal permeability with changes in intestinal biopsy at various intervals after commencing a GFD. Intestinal permeability was measured by lactulose/rhamnose absorption from 1 week to 24 months after commencing a GFD. Intestinal morphometry was measured by villus area, crypt length and mitotic count per crypt at diagnosis and after commencing a GFD. Median intestinal permeability values decreased from 0.47 (n = 35) at diagnosis to 0.25 (n = 17) after 1 week and to 0.16 (n = 18) after 2 months of a GFD. Rhamnose absorption improved significantly at an early stage, from 6.6% (untreated) to 15.4% at 3 months of a GFD, whereas the decrease in lactulose permeation took longer: from 3.4% (untreated) to 0.8% after 12 months of a GFD. Mean villus area (n = 29) was reduced to 16% of control values at diagnosis, and improved to a maximum of 48% after 6 months on a GFD, but did not change thereafter. Mean crypt length and mitotic count per crypt were increased by 222% and 356% respectively at diagnosis, and these parameters remained elevated at 172% and 216% above control values after 6 months of a GFD. We conclude that intestinal permeability improves within 2 months after starting a GFD, but that measurable intestinal biopsy improvement requires ingestion of a GFD for at least 3-6 months, and even then remains incomplete. PMID- 11256976 TI - Nitric oxide modulation of blood vessel tone identified by arterial waveform analysis. AB - Traditionally, nitric oxide-mediated alteration in blood vessel tone has been inferred from changes in flow in response to physical and pharmacological interventions using plethysmographic or ultrasonic techniques. We hypothesized that alteration in pulsatile arterial function may represent a more sensitive measure to detect and monitor nitric oxide-mediated modulation of arterial smooth muscle tone. Healthy male volunteers (n = 15) had radial artery pressure pulse waveforms recorded using a calibrated tonometer device. A computer-based assessment of the diastolic pressure decay was employed to quantify changes in arterial waveform morphology in terms of altered pulsatile (arterial compliance) and steady-state (peripheral resistance) haemodynamics. N(G)-nitro-L-arginine methyl ester (L-NAME), a stereospecific inhibitor of nitric oxide synthesis, was infused intravenously in incrementally increasing doses of 0.25, 0.5 and 0.75 mg/kg for 8 min each. Subjects then received either L-arginine or D-arginine (200 mg/kg over 15 min) intravenously in a blinded fashion. On a separate day, subjects had radial artery pressure pulse waveforms recorded before and after the sublingual administration of glyceryl trinitrate, an exogenous donor of nitric oxide. Cardiac output and heart rate decreased and mean arterial blood pressure increased significantly (P < 0.01 for all) in response to the incremental intravenous infusion of L-NAME. Small artery compliance decreased, whereas systemic vascular resistance increased in response to nitric oxide synthesis inhibition (P < 0.01 for both). The intravenous infusion of L-arginine restored the pulsatile and steady-state haemodynamic parameters to pre-treatment values, whereas D-arginine had no effect. Sublingual glyceryl trinitrate decreased systemic vascular resistance by 11%, whereas large artery- and small artery compliance increased by 25% and 44% respectively. Pressure pulse contour analysis represents a sensitive and convenient technique capable of tracking changes in the pulsatile function of arteries accompanying nitric oxide-mediated alteration in arterial smooth muscle tone. PMID- 11256977 TI - A novel method for the delivery of nitric oxide therapy to the skin of human subjects using a semi-permeable membrane. AB - Nitric oxide (NO) is a mediator of essential biological processes, including vasodilatation, anti-microbial activity and wound healing. A chemical system using sodium nitrite and ascorbic acid has been developed which generates significant amounts of NO. The originally described system was messy and impractical, and the high acidity may cause pain and further tissue damage in ulcerated skin. To overcome this, a selectively permeable, hydrophilic polyester co-polymer membrane system (Sympatex ) has been identified that can be placed between the NO-generating chemicals and the skin. The aim of the present study was to determine whether NO derived from this chemical system was able to diffuse through the membrane and have a measurable vasodilatory effect on forearm skin in healthy volunteers. The Sympatex 10 microm membrane was found to be highly permeable to NO, while preventing passage of the constituents of the NO generation gel to the skin. The transmembrane NO-generation system had a vasodilatory effect comparable with that resulting from direct topical application. Additionally, the NO generated was effective in killing Staphylococcus aureus and Escherichia coli at doses lower than those required to increase skin blood flow. The vasodilatory and anti-microbial effects of this system may be useful as a patch-based topical therapy for skin ulceration, particularly when there is concomitant ischaemia and infection. PMID- 11256978 TI - Male pattern baldness is not associated with established cardiovascular risk factors in the general population. AB - A number of studies have shown an association between male pattern baldness (MPB) and cardiovascular disease. Few of these studies, however, have examined whether MPB is a novel risk factor, or is associated with abnormalities of established coronary risk factors. We have therefore performed an analysis of MPB and cardiovascular risk factors in the general population. A total of 1219 male participants aged 18-70 years from the Victorian Family Heart Study were surveyed using a validated questionnaire for degree and pattern of baldness. Carefully standardized measures of height, weight, blood pressure, pulse rate, total and high-density lipoprotein cholesterol, and plasma fibrinogen were made. Subjects were grouped according to the degree and pattern of baldness as: no baldness, frontal baldness and vertex baldness. Bald men were older than non-bald men (P < 0.0001). Age was also associated with increased levels of coronary risk factors (P < 0.0001). When multiple regression was used to adjust for age differences, the levels of coronary risk factors were not significantly different between the bald and non-bald groups. The lack of association between baldness and established coronary risk factors implies that baldness may predispose to coronary heart disease through novel mechanisms yet to be defined. PMID- 11256979 TI - Thrombosis in one coronary artery causes generalized coronary vasoconstriction in a dog model of unstable angina. AB - We investigated the effect of thrombosis in one coronary artery upon the vascular resistance of another coronary artery. In previous investigations, using an animal model of unstable angina, we have observed increased resistance downstream from thrombus within a left circumflex coronary artery (LCx) stenosis and vasoconstriction of collateral vessels from the left anterior descending artery (LAD) supplying the distal LCx vascular bed. In the present paper, we induced thrombosis within a stenosis of the LCx of 16 beagle dogs, and observed the changes in blood flow to the myocardium supplied by the LAD using the radioactive microsphere technique. This blood flow decreased with thrombosis (P = 0.005) in these animals, whereas it did not do so in three time-control experiments. The pressures across the coronary vascular bed, i.e. arterial pressure to coronary venous pressure (coronary sinus catheter), did not change. Thus the vascular resistance of the LAD bed increased significantly from 147 +/- ll.5 mmHg/ml/sec/g of tissue to 172 +/- 13.4 mmHg/ml/sec/g of tissue (P = 0.02). As the LAD territory is not perfused with blood from the artery containing thrombus, we conclude that the effect observed is caused either by release of vasoconstrictors from the thrombus into the general circulation, or by activation of a neural reflex vasoconstriction. The study suggests that unstable angina involving thrombosis in one coronary artery is a global coronary vascular disease. PMID- 11256980 TI - Mucosal protective effects of ecabet sodium: pepsin inhibition and interaction with mucus. AB - Pepsin, acid and Helicobacter pylori are major factors in the pathophysiology of peptic ulcer disease and reflux oesophagitis. Ecabet sodium reduces the survival of H. pylori in the stomach and inhibits pepsin activity in the gastric juice of experimental animals. Here we have investigated the effects of ecabet sodium on some of the factors involved in the dynamics of the mucosal barrier, i.e. pepsins and mucins. This study used gastric juice obtained from 12 non-symptomatic volunteers and nine patients with reflux oesophagitis. Ecabet sodium significantly inhibited pepsin activity in human gastric juice, with a maximum inhibition of 78%. Pepsin 1, the ulcer-associated pepsin, was inhibited to the greatest extent. The ability of gastric juice to digest mucin was significantly inhibited by ecabet. As with gastric juice proteolytic activity, the inhibitory effect of ecabet on mucolysis was greater in gastric juice from patients with reflux oesophagitis than in that from controls. Ecabet sodium showed a positive interaction with gastric mucin, as assessed by an increase in viscosity. Thus ecabet sodium may reduce the aggressive potential of gastric juice towards the mucosa, which may be relevant in the treatment of reflux oesophagitis and peptic ulcer disease. In addition, it may strengthen the mucus barrier in peptic ulcer disease and gastritis. PMID- 11256981 TI - Left atrial thrombin generation and prothrombin fragment 1+2: authors' reply. PMID- 11256982 TI - Follow-up study of the benefits of hormone replacement therapy on isometric muscle strength of adductor pollicis in postmenopausal women. PMID- 11256983 TI - Assessment of regional long-axis function during dobutamine echocardiography. AB - Echocardiographic analysis of regional left ventricular function is based upon the assessment of radial motion. Long-axis motion is an important contributor to overall function, but has been difficult to evaluate clinically until the recent development of tissue Doppler techniques. We sought to compare the standard visual assessment of radial motion with quantitative tissue Doppler measurement of peak systolic velocity, timing and strain rate (SRI) in 104 patients with known or suspected coronary artery disease undergoing dobutamine stress echocardiography (DbE). A standard DbE protocol was used with colour tissue Doppler images acquired in digital cine-loop format. Peak systolic velocity (PSV), time to peak velocity (TPV) and SRI were assessed off-line by an independent operator. Wall motion was assessed by an experienced reader. Mean PSV, TPV and SRI values were compared with wall motion and the presence of coronary artery disease by angiography. A further analysis included assessing the extent of jeopardized myocardium by comparing average values of PSV, TPV and SRI against the previously validated angiographic score. Segments identified as having normal and abnormal radial wall motion showed significant differences in mean PSV (7.9 +/- 3.8 and 5.9 +/- 3.3 cm/s respectively; P < 0.001), TPV (84 +/- 40 and 95 +/- 48 ms respectively; P = 0.005) and SRI (-1.45 +/- 0.5 and -1.1 +/- 0.9 s(-1) respectively; P < 0.001). The presence of a stenosed subtending coronary artery was also associated with significant differences from normally perfused segments for mean PSV (8.1+/-3.4 compared with 5.7+/-3.7 cm/s; P < 0.001), TPV (78 +/- 50 compared with 92 +/- 45 ms; P < 0.001) and SRI (-1.35 +/- 0.5 compared with -1.20 +/- 0.4 s(-1); P = 0.05). PSV, TPV and SRI also varied significantly according to the extent of jeopardized myocardium within a vascular territory. These results suggest that peak systolic velocity, timing of contraction and SRI reflect the underlying physiological characteristics of the regional myocardium during DbE, and may potentially allow objective analysis of wall motion. PMID- 11256985 TI - Clinical autonomic testing and posture: are we doing the right things? PMID- 11256984 TI - Influence of posture on the Valsalva manoeuvre. AB - The objective of the present study was to evaluate the influence of posture on the responses of blood pressure (BP) and heart rate (HR) to the Valsalva manoeuvre (VM). Neurohumoral activation, as well as changes in intravascular and intracardiac volumes and pressures, are well known effects of orthostatic stress. These changes are likely to have significant effects on cardiovascular reflexes, such as the response to the VM. However, the influence of posture on the VM has not been intensively evaluated, except for a few studies involving small sex- and age-selected case series. We therefore investigated the effects of posture on the VM in a larger non-selected group of healthy control subjects. In 19 healthy volunteers (ten female/nine male; age range 20-72 years, mean age 43 years), two reproducible VMs (40 mmHg; 15 s) were performed after 10 min of supine rest, 10 min of sitting and 10 min of standing. HR and BP were monitored continuously. End diastolic volume, total peripheral resistance and cardiac output were calculated at baseline for each position. We found that assuming an upright position resulted in increases in total peripheral resistance and HR, accompanied by decreases in end-diastolic volume and cardiac output. The fall in BP during early phase II and the BP overshoot during phase IV were clearly more pronounced with increasing orthostatic stress, whereas the rise in BP during late phase II remained unchanged; pulse pressure was more compressed during phase II, but higher during phase IV. The Valsalva ratio was not significantly affected, but baroreflex gain (calculated from early phase II) was significantly decreased in the upright position. While a reduced late phase II was observed on one occasion in each of the lying and sitting positions, three abnormal responses were observed during standing. We conclude that posture has a significant influence on BP responses to the VM, probably resulting from changes in the intrathoracic blood volume. Standing results in a lower rate of 'flat-top' responses, but also seems to reduce the specificity of this test. Sympathetic activation in the upright position seems to blunt baroreflexes, leading to similar HR responses in spite of larger changes in BP. PMID- 11256986 TI - Sodium/lithium countertransport, insulin resistance, insulin peptides and microalbuminuria in clinically healthy 58-year-old men. AB - The activity of the erythrocyte transport system, sodium/lithium countertransport (SLC), has been linked to the metabolic syndrome characterized by insulin resistance and compensatory hyperinsulinaemia. We measured SLC and insulin sensitivity with the euglycaemic hyperinsulinaemic clamp method in a patient sample (n = 93) randomly selected from a large clinically healthy group of 58 year-old men (n = 818). The lipid profile, blood pressure, body mass index (BMI) and insulin were also analysed. There was a significant difference (P < 0.001) in SLC between subjects with the metabolic syndrome (n = 19) and subjects without any components of this syndrome (n = 20). There was a highly significant correlation between SLC and BMI, waist/hip ratio, total body fat mass, serum triglycerides, plasma insulin, proinsulin split products and C-peptide in a univariate analysis. There was also a significant correlation between SLC and insulin sensitivity measured as insulin-mediated glucose uptake (P < 0.01). In multiple regression analysis, only two of the variables showing univariate significance were independently correlated to SLC, i.e. serum triglycerides (P < 0.001) and BMI (P < 0.01). The subjects with a SLC value in the highest tertile had a 6-fold higher prevalence of insulin resistance (low-insulin-mediated glucose uptake) as compared with those with a SLC value in the lowest tertile. We conclude that, in clinically healthy 58-year-old men from the general population, erythrocyte SLC is closely linked to metabolic syndrome, in particular to obesity, triglycerides and insulin resistance. PMID- 11256987 TI - beta2-Adrenoceptor desensitization in human alveolar macrophages induced by inhaled terbutaline in vivo is not counteracted by budesonide. AB - In vitro studies suggest that glucocorticoids may counteract beta-agonist-induced desensitization of beta-adrenoceptors by actions at the transcriptional level, but the clinical relevance of such findings is not clear. Oral terbutaline treatment decreases beta-adrenoceptor sensitivity in alveolar macrophages in vivo. This effect is not counteracted by inhaled or orally taken steroids. We therefore examined whether inhaled terbutaline elicited a similar effect on beta(2)-adrenoceptor sensitivity in alveolar macrophages, and if co-treatment with an inhaled steroid, budesonide, would prevent such down-regulation. Bronchoalveolar lavage (BAL) and lung function tests, including bronchodilator responses to inhaled terbutaline, were performed before and after 2 weeks of regular inhalation of terbutaline, 0.5 mg three times daily, and budesonide, 400 microg twice daily, or placebo, in 24 healthy volunteers. Four untreated subjects served as controls. A marked, approx. 90%, decrease in isoprenaline-induced cAMP accumulation in alveolar macrophages was found in both treatment groups after 2 weeks, with no difference between placebo and budesonide (P = 0.45). In the untreated control group, cAMP responses to both isoprenaline and prostaglandin E(1) tended to be lower on the second occasion. A limited, non-specific desensitization of adenylate cyclase activity thus contributed to the marked desensitization elicited by terbutaline inhalations. The bronchodilator response to inhaled terbutaline did not change after treatment in any of the three groups (F = 0.9, P = 0.50). In conclusion, inhalation of a beta-agonist induced marked down-regulation of beta(2)-adrenoceptor sensitivity in alveolar macrophages in vivo without influencing the bronchodilator response to a beta(2)-agonist in healthy subjects. Co-treatment with an inhaled steroid failed to counteract the desensitization of alveolar macrophage beta(2)-adrenoceptors. PMID- 11256988 TI - Effects of smoking and abstention from smoking on fibrinogen synthesis in humans. AB - Cigarette smoking and hyperfibrinogenaemia are both significant risk factors for the development of cardiovascular disease. Two studies are described here which aimed to establish the metabolic mechanism responsible for the raised plasma fibrinogen concentration observed in smokers. Chronic smokers had a significantly elevated absolute rate of fibrinogen synthesis (ASR) compared with non-smokers (22.7 +/- 1.3 mg/kg per day versus 16.0 +/- 1.3 mg/kg per day; means +/- S.E.M., P < 0.01), with plasma levels of fibrinogen significantly correlated with fibrinogen synthesis (r = 0.65, P = 0.04). Unlike fibrinogen, plasma albumin concentrations were lower in smokers than in non-smokers (45 +/- 0.4 versus 47 +/ 0.7 g/l, P < 0.05), but there was no difference in rates of albumin synthesis between the two groups. Two weeks cessation from smoking by previously chronic smokers was associated with a rapid and marked fall in plasma fibrinogen concentration (from 3.06 +/- 0.11 g/l to 2.49 +/- 0.14 g/l, P < 0.001), and a significant reduction in ASR (a 33% reduction, from 24.1 +/- 1.7 to 16.1 +/- 1.0 mg/kg per day, P < 0.001). These studies suggest a primary role for increased synthesis in producing the hyperfibrinogenaemia associated with smoking. Moreover, abstention from smoking for a period of only 2 weeks induces a significant decrease in the rate of fibrinogen synthesis by the liver, with a concomitant reduction in the plasma fibrinogen concentration. PMID- 11256991 TI - The postsynaptic NMDA-receptor--PSD-95 signaling complex in excitatory synapses of the brain. PMID- 11256992 TI - SH3 domains: complexity in moderation. AB - The SH3 domain is perhaps the best-characterized member of the growing family of protein-interaction modules. By binding with moderate affinity and selectivity to proline-rich ligands, these domains play critical roles in a wide variety of biological processes ranging from regulation of enzymes by intramolecular interactions, increasing the local concentration or altering the subcellular localization of components of signaling pathways, and mediating the assembly of large multiprotein complexes. SH3 domains and their binding sites have cropped up in many hundreds of proteins in species from yeast to man, which suggests that they provide the cell with an especially handy and adaptable means of bringing proteins together. The wealth of genetic, biochemical and structural information available provides an intimate and detailed portrait of the domain, serving as a framework for understanding other modular protein-interaction domains. Processes regulated by SH3 domains also raise important questions about the nature of specificity and the overall logic governing networks of protein interactions. PMID- 11256993 TI - Oligomerisation of G-protein-coupled receptors. AB - A range of approaches have recently provided evidence that G-protein-coupled receptors can exist as oligomeric complexes. Both homo-oligomers, comprising multiple copies of the same gene product, and hetero-oligomers containing more than one receptor have been detected. In several, but not all, examples, the extent of oligomerisation is regulated by the presence of agonist ligands, and emerging evidence indicates that receptor hetero-oligomers can display distinct pharmacological characteristics. A chaperonin-like role for receptor oligomerisation in effective delivery of newly synthesised receptors to the cell surface is a developing concept, and recent studies have employed a series of energy-transfer techniques to explore the presence and regulation of receptor oligomerisation in living cells. However, the majority of studies have relied largely on co-immunoprecipitation techniques, and there is still little direct information on the fraction of receptors existing as oligomers in intact cells. PMID- 11256994 TI - Characterization of residues and sequences of the carbohydrate recognition domain required for cell surface localization and ligand binding of human lectin-like oxidized LDL receptor. AB - Lectin-like oxidized low-density lipoprotein receptor (LOX-1) has been cloned from human aortic endothelial cells, and has a sequence identical to that from human lung. Previous studies showed that human LOX-1 can recognize modified LDL, apoptotic cells and bacteria. To further explore the relationship between the structure and function of LOX-1, a mutagenesis study was carried out. Our results showed that the carbohydrate recognition domain (CRD) was the ligand-binding domain of human LOX-1. We also investigated the sequences and residues in CRD that were essential for protein cell surface localization and ligand binding. LOX 1s carrying a mutation on each of six Cys in CRD resulted in a variety of N glycosylation and failed to be transported to the cell surface. This was strong evidence for the involvement of all six Cys in the intrachain disulfide bonds required for proper folding, processing and transport of LOX-1. The C-terminal sequence (KANLRAQ) was also essential for protein folding and transport, while the four final residues (LRAQ) were involved in maintaining receptor function. Both positive charged (R208, R209, H226, R229 and R231) and non-charged hydrophilic (Q193, S198, S199 and N210) residues were involved in ligand binding, suggesting that ligand recognition of LOX-1 is not merely dependent on the interaction of positively charged residues with negatively charged ligands. PMID- 11256995 TI - p110-related PI 3-kinases regulate phagosome-phagosome fusion and phagosomal pH through a PKB/Akt dependent pathway in Dictyostelium. AB - The Dictyostelium p110-related PI 3-kinases, PIK1 and PIK2, regulate the endosomal pathway and the actin cytoskeleton, but do not significantly regulate internalization of particles in D. discoideum. Bacteria internalized into delta ddpik1/ddpik2 cells or cells treated with PI 3-kinase inhibitors remained intact as single particles in phagosomes with closely associated membranes after 2 hours of internalization, while in control cells, bacteria appeared degraded in multi particle spacious phagosomes. Addition of LY294002 to control cells, after 60 minutes of chase, blocked formation of spacious phagosomes, suggesting PI 3 kinases acted late to regulate spacious phagosome formation. Phagosomes purified from control and drug treated cells contained equivalent levels of lysosomal proteins, including the proton pump complex, and were acidic, but in drug treated cells and delta ddpik1/ddpik2 cells phagosomal pH was significantly more acidic during maturation than the pH of control phagosomes. Inhibition of phagosomal maturation by LY294002 was overcome by increasing phagosomal pH with NH(4)Cl, suggesting that an increase in pH might trigger homotypic phagosome fusion. A pkbA null cell line (PKB/Akt) reproduced the phenotype described for cells treated with PI 3-kinase inhibitors and delta ddpik1/ddpik2 cells. We propose that PI 3-kinases, through a PKB/Akt dependent pathway, directly regulate homotypic fusion of single particle containing phagosomes to form multi-particle, spacious phagosomes, possibly through the regulation of phagosomal pH. PMID- 11256997 TI - Recruitment of beta-catenin to cadherin-mediated intercellular adhesions is involved in myogenic induction. AB - Cadherin-mediated cell adhesion is involved in muscle differentiation from early stages of myogenic induction to late stages of myoblast interaction and fusion. beta-Catenin is a major constituent of cadherin-based adherens junctions and also serves as a signal transduction molecule that regulates gene expression during development. In this study, we explored the involvement of beta-catenin in myogenic differentiation. We show here that shortly after a switch from growth to differentiation medium, beta-catenin translocates to cell-cell junctions and its levels increase. We further show that elevation of beta-catenin levels, induced either by inhibition of its breakdown, using LiCl, or by its overexpression, suppresses the formation of adherens junctions, resulting in a sharp decline in myogenin expression and an arrest of myogenic progression. Recruitment of beta catenin to adherens junctions after transfection with N-cadherin restores myogenin expression in the transfected cells. These results suggest that increased cadherin-mediated adhesion and translocation of beta-catenin to adherens junctions are involved in activating the early steps of myogenic differentiation. PMID- 11256996 TI - Role of aquaporin-4 water channel in the development and integrity of the blood brain barrier. AB - In this study, we have investigated the expression of aquaporin 4 during blood brain barrier development in the optic tectum of chick embryos and newly hatched chicks, by means of western-blot, reverse transcriptase-polymerase chain reaction, immunohistochemistry, and freeze-fracture and high-resolution immunogold electron microscopy. In the optic tecta of day-14 embryos, western blot analysis revealed an approx. 30 kDa band, immunoreactive for aquaporin-4, which was increased in day-20 embryos and in chicks. Semi-quantitative reverse transcriptase chain reaction experiments showed that there was already a high level of aquaporin-4 mRNA in day-9 embryos as well as in the subsequent stages and in newly hatched chicks. Immunohistochemically, reactivity for aquaporin-4 was detected in the optic tectum of day-14 embryos; similar results were obtained in telencephalon and cerebellum. Ultrastructurally, the microvessels of the tectum showed immunoreactivity for aquaporin-4 on the astroglial endfeet, which discontinuously surrounded endothelial cells joined by immature tight junctions. In the tectum, telencephalon and cerebellum of 20-day embryos and chicks, aquaporin-4 strongly labeled the ependymal cells and the subpial glial membranes, as well as the bodies and processes of astroglial cells. A continuous aquaporin-4 staining was found around the microvessel endothelial cells, which were sealed off from one another by extensive tight junctions. A complete astrocytic sheath, labeled by anti-aquaporin-4 gold particles, enveloped the endothelium-pericyte layer. Orthogonal arrays of particles were observed on fractured astrocytic membranes, starting from embryonic day 14 when the aquaporin-4 immunogold staining revealed clusters of gold particles, often forming square or rectangular clusters. The results showed that aquaporin-4 expression and organization of the intramembrane particles in orthogonal arrays followed the same temporal sequence. Finally, the lipopolysaccharide, a substance that induces blood-brain barrier disruption, determines a remarkable reduction in aquaporin-4 labeling, expressed by a few aquaporin-4 gold particles attached on swollen perivascular glial membranes. All these data show that aquaporin-4 expression occurs in the chick embryonic brain, in parallel with maturation and functioning of the blood-brain barrier and suggest that there is a close relationship between water transport regulation and brain development. PMID- 11256998 TI - Differential galactosylation of neuronal and haematopoietic signal regulatory protein-alpha determines its cellular binding-specificity. AB - Signal regulatory protein-alpha (SIRP alpha) is a member of the Ig superfamily selectively expressed by neuronal and myeloid cells. The molecule mediates functional interactions with CD47/integrin-associated protein. Here we provide evidence for the tissue-specific glycosylation of neuronal and haematopoietic SIRP alpha. We demonstrate a major difference in the galactosylation of N-linked glycans isolated from neuronal (i.e. brain-derived) SIRP alpha as compared to myeloid (i.e. spleen-derived) SIRP alpha, with neuronal SIRP alpha almost completely lacking galactose. beta 4-galactosyltransferase assays demonstrated that this is most likely due to a low galactosylation capacity of the brain. In order to investigate the role of galactosylation of SIRP alpha in cellular interactions, soluble recombinant SIRP alpha glycoforms containing galactose (SIRP alpha-Fc) or lacking galactose (SIRP alpha(Delta Gal)-Fc) were produced. Binding studies demonstrated superior binding of SIRP alpha(Delta Gal)-Fc to cerebellar neurons and isolated lymphocytes. In contrast, SIRP alpha-Fc bound relatively strong to macrophages. These data show that the galactosylation of SIRP alpha determines its cellular binding specificity. PMID- 11256999 TI - The GTPase Rac1 selectively regulates Salmonella invasion at the apical plasma membrane of polarized epithelial cells. AB - The bacterial pathogen Salmonella typhimurium colonizes its animal hosts by inducing its internalization into intestinal epithelial cells. This process requires reorganization of the actin cytoskeleton of the apical plasma membrane into elaborate membrane ruffles that engulf the bacteria. Members of the Rho family of small GTPases are critical regulators of actin structure, and in nonpolarized cells, the GTPase Cdc42 has been shown to modulate Salmonella entry. Because the actin architecture of epithelial cells is organized differently from that of nonpolarized cells, we examined the role of two Rho family GTPases, Cdc42 and Rac1, in invasion of polarized monolayers of MDCK cells by S. typhimurium. Surprisingly, we found that endogenous Rac1, but not Cdc42, was activated during bacterial entry at the apical pole, and that this activation required the bacterial effector protein SopE. Furthermore, expression of dominant inhibitory Rac1 but not Cdc42 significantly inhibited apical internalization of Salmonella, indicating that Rac1 activation is integral to the bacterial entry process. In contrast, during basolateral internalization, both Cdc42 and Rac1 were activated; however, neither GTPase was required for entry. These findings, which differ significantly from previous observations in nonpolarized cells, indicate that the host cell signaling pathways activated by bacterial pathogens may vary with cell type, and in epithelial tissues may further differ between plasma membrane domains. PMID- 11257000 TI - Rho and Rac but not Cdc42 regulate endothelial cell permeability. AB - Endothelial permeability induced by thrombin and histamine is accompanied by actin stress fibre assembly and intercellular gap formation. Here, we investigate the roles of the Rho family GTPases Rho1, Rac1 and Cdc42 in regulating endothelial barrier function, and correlate this with their effects on F-actin organization and intercellular junctions. RhoA, Rac1 and Cdc42 proteins were expressed efficiently in human umbilical vein endothelial cells by adenovirus mediated gene transfer. We show that inhibition of Rho prevents both thrombin- and histamine-induced increases in endothelial permeability and decreases in transendothelial resistance. Dominant-negative RhoA and a Rho kinase inhibitor, Y 27632, not only inhibit stress fibre assembly and contractility but also prevent thrombin- and histamine-induced disassembly of adherens and tight junctions in endothelial cells, providing an explanation for their effects on permeability. In contrast, dominant-negative Rac1 induces permeability in unstimulated cells and enhances thrombin-induced permeability, yet inhibits stress fibre assembly, indicating that increased stress fibre formation is not essential for endothelial permeability. Dominant-negative Cdc42 reduces thrombin-induced stress fibre formation and contractility but does not affect endothelial cell permeability or responses to histamine. These results demonstrate that Rho and Rac act in different ways to alter endothelial barrier function, whereas Cdc42 does not affect barrier function. PMID- 11257001 TI - Analysis of R-Ras signalling pathways. AB - R-Ras has a high degree of sequence homology to Ras and to other members of the Ras subfamily including Rap, TC21 and M-Ras. Activated versions of Ras and TC21 are highly transforming in a variety of cell lines and mutated forms of both proteins have been found in human tumours. R-Ras interacts with many of the same proteins as Ras and TC21, including c-Raf1, and can induce transformed foci, although this activity is weak compared to Ras and appears to be cell-type specific. Here, we have investigated R-Ras signalling pathways in a variety of cell types. We find that microinjection of activated R-Ras into quiescent fibroblasts stimulates cell cycle progression through G(1) phase and subsequent DNA synthesis. However, unlike Ras, R-Ras does not activate the ERK MAP kinase pathway nor does it activate the JNK or p38/Mpk2 MAP kinase pathways. Microinjection of R-Ras into PC12 cells does not induce terminal differentiation, but instead causes extensive cell spreading, consistent with R-Ras having a role in integrin activation. Finally, in a macrophage cell line, R-Ras activates the alpha(M)beta(2) integrin via the small GTPase Rap1, leading to phagocytosis of opsonized red blood cells, whereas Ras does not. These results indicate that R Ras has an important role in the regulation of cell growth and adhesion, but that this is mediated through downstream signals distinct from those used by Ras. PMID- 11257002 TI - Specific changes to the mechanism of cell locomotion induced by overexpression of beta-actin. AB - Overexpression of beta-actin is known to alter cell morphology, though its effect on cell motility has not been documented previously. Here we show that overexpressing beta-actin in myoblasts has striking effects on motility, increasing cell speed to almost double that of control cells. This occurs by increasing the areas of protrusion and retraction and is accompanied by raised levels of beta-actin in the newly protruded regions. These regions of the cell margin, however, show decreased levels of polymerised actin, indicating that protrusion can outpace the rate of actin polymerisation in these cells. Moreover, the expression of beta*-actin (a G244D mutant, which shows defective polymerisation in vitro) is equally effective at increasing speed and protrusion. Concomitant changes in actin binding proteins show no evidence of a consistent mechanism for increasing the rate of actin polymerisation in these actin overexpressing cells. The increase in motility is confined to poorly spread cells in both cases and the excess motility can be abolished by blocking myosin function with butanedione monoxime (BDM). Our observations on normal myoblasts are consistent with the view that they protrude by the assembly and cross linking of actin filaments. In contrast, the additional motility shown by cells overexpressing beta-actin appears not to result from an increase in the rate of actin polymerisation but to depend on myosin function. This suggests that the additional protrusion arises from a different mechanism. We discuss the possibility that it is related to retraction-induced protrusion in fibroblasts. In this phenomenon, a wave of increased protrusion follows a sudden collapse in cell spreading. This view could explain why it is only the additional motility that depends on spreading, and has implications for understanding the differences in locomotion that distinguish tissue cells from highly invasive cell types such as leucocytes and malignant cells. PMID- 11257003 TI - The Tem1 small GTPase controls actomyosin and septin dynamics during cytokinesis. AB - Cytokinesis in budding yeast involves an actomyosin-based ring which assembles in a multistepped fashion during the cell cycle and constricts during cytokinesis. In this report, we have investigated the structural and regulatory events that occur at the onset of cytokinesis. The septins, which form an hour-glass like structure during early stages of the cell cycle, undergo dynamic rearrangements prior to cell division: the hourglass structure splits into two separate rings. The contractile ring, localized between the septin double rings, immediately undergoes contraction. Septin ring splitting is independent of actomyosin ring contraction as it still occurs in mutants where contraction fails. We hypothesize that septin ring splitting may remove a structural barrier for actomyosin ring to contract. Because the Tem1 small GTPase (Tem1p) is required for the completion of mitosis, we investigated its role in regulating septin and actomyosin ring dynamics in the background of the net1-1 mutation, which bypasses the anaphase cell cycle arrest in Tem1-deficient cells. We show that Tem1p plays a specific role in cytokinesis in addition to its function in cell cycle progression. Tem1p is not required for the assembly of the actomyosin ring but controls actomyosin and septin dynamics during cytokinesis. PMID- 11257004 TI - BMP-2 augments FGF-induced differentiation of PC12 cells through upregulation of FGF receptor-1 expression. AB - When exposed to various neurotrophic factors, including fibroblast growth factors (FGF)-1 and -2, rat pheochromocytoma-derived PC12 cells differentiate into sympathetic neuron-like cells possessing elongated neurites. We found that while bone morphogenetic protein-2 (BMP-2) exerted little effect by itself on the differentiation of PC12 cells, in combination with FGF it strongly induced neurite outgrowth, even at subthreshold concentrations of FGF. Analysis of gene expression revealed that FGF receptor-1 (FGFR-1) mRNA was abundantly expressed in PC12 cells and that its expression was upregulated by pretreating the cells with BMP-2. Crosslinking the receptors with (125)I-FGF-2 and then immunoprecipitating them confirmed that expression of FGFR-1, but not other FGF receptor types, was enhanced by BMP-2. Furthermore, Scatchard analyses revealed that the numbers of FGF-2 binding sites were increased by approximately 40% after BMP-2 treatment. Pretreatment with BMP-2 also enhanced peak and sustained levels of FGF-induced ERK1/2 phosphorylation in PC12 cells. Finally, the augmentation of neurotrophic activity by BMP-2 was inhibited by SU5402, an FGFR-1 inhibitor. These findings indicate that BMP-2 augments FGF-induced differentiation of PC12 cells through selective upregulation of FGFR-1 expression, and suggest that BMP-2 and FGF act in concert to regulate cell differentiation in the nervous system. PMID- 11257005 TI - Cell-specific targeting of caveolin-1 to caveolae, secretory vesicles, cytoplasm or mitochondria. AB - In commonly used tissue culture cells, caveolin-1 is embedded in caveolae membranes. It appears to reach this location after being cotranslationally inserted into ER membranes, processed in the Golgi and shipped to the cell surface. We now report that caveolae are not the preferred location for caveolin 1 in all cell types. Skeletal muscle cells and keratinocytes target caveolin-1 to the cytosol while in exocrine and endocrine cells it accumulates in the secretory pathway. We also found that airway epithelial cells accumulate caveolin-1 in modified mitochondria. The cytosolic and the secreted forms appear to be incorporated into a soluble, lipid complex. We conclude that caveolin-1 can be targeted to a variety of intracellular destinations, which suggests a novel mechanism for the intracellular traffic of this protein. PMID- 11257006 TI - Alteration of the stability of Bag-1 protein in the control of olfactory neuronal apoptosis. AB - Normal apoptosis occurs continuously in the olfactory neuroepithelium of adult vertebrates, making it a useful model for studying neuronal apoptosis. Here we demonstrate that overexpression of the anti-apoptotic Bag-1 gene in olfactory neuronal cells confers a strong resistance to apoptosis. Conversely decreased levels of Bag-1 were found to precede a massive wave of olfactory neuronal apoptosis triggered by synaptic target ablation. We show that the decrease is brought about by ubiquitination and subsequent degradation of the Bag-1 protein. The ring finger protein Siah-2 is a likely candidate for the ubiquitination reaction since Siah-2 mRNA accumulated in lesioned olfactory neuroepithelium and overexpression of Siah-2 stimulated Bag-1 ubiquitination and degradation in transient expression assays. These results together identify destabilization of Bag-1 as a necessary step in olfactory neuronal apoptosis. PMID- 11257007 TI - Structural analysis of the role of the beta 3 subunit of the alpha V beta 3 integrin in IGF-I signaling. AB - The disintegrin echistatin inhibits ligand occupancy of the alpha V beta 3 integrin and reduces Insulin-like growth factor I (IGF-I) stimulated migration, DNA synthesis, and receptor autophosphorylation in smooth muscle cells. This suggests that ligand occupancy of the alpha V beta 3 receptor is required for full activation of the IGF-I receptor. Transfection of the full-length beta 3 subunit into CHO cells that have no endogenous beta 3 and do not migrate in response to IGF-I was sufficient for IGF-I to stimulate migration of these anchorage dependent cells. In contrast, transfection of either of two truncated mutant forms of beta 3 (terminating at W(715) or E(731)) or a mutant with substitutions for Tyr(747) Tyr(759) (YY) into either CHO or into porcine smooth muscle cells did not restore the capacity of these cells to migrate across a surface in response to IGF-I. This effect was not due to loss of IGF-I receptor autophosphorylation since the response of the receptor to IGF-I was similar in cells expressing either the full-length or any of the mutant forms of the beta 3 subunit. Echistatin reduced IGF-I receptor phosphorylation in cells expressing the full-length or the YY mutant forms of beta 3 subunit, but it had no effect in cells expressing either of two truncated forms of beta 3. A cell-permeable peptide homologous to the C-terminal region of the beta 3 subunit (amino acids 747-762) reduced IGF-I stimulated migration and receptor autophosphorylation of non-transfected porcine smooth muscle cells. These results demonstrate that the full-length beta 3 with intact tyrosines at positions 747 and 759 is required for CHO cells to migrate in response to IGF-I. Furthermore, a region of critical amino acids between residues 742-762 is required for echistatin to induce its regulatory effect on receptor phosphorylation. Since the IGF-I receptor does not bind to alpha V beta 3 the results suggest that specific but distinct regions of the beta 3 subunit interact with intermediary proteins to facilitate IGF-I stimulated cell migration and echistatin induced inhibition of IGF-I signal transduction. PMID- 11257008 TI - Riddle of biofilm resistance. PMID- 11257009 TI - Antifungal efficacy of GM237354, a sordarin derivative, in experimental oral candidiasis in immunosuppressed rats. AB - GM237354 is a novel sordarin derivative with a broad spectrum of potent activity against a wide range of fungi. The members of this new class of antifungal agents act as potent inhibitors of fungal protein synthesis. In this study, the therapeutic effects of GM237354 were investigated in a novel experimental oral Candida albicans infection model in immunosuppressed rats. The animals were immunosuppressed with dexamethasone in their drinking water and infected on three alternate days. GM237354 was given three times per day for seven consecutive days at 1.25, 2.5, 5, or 10 mg/kg of body weight per dose. In addition, to provide a preliminary idea of the correlation between regimen administration and therapeutic efficacy, GM237354 was administered to two additional groups of rats at 5 mg/kg once or twice a day for 7 days. The drug efficacy was assessed microbiologically, histologically, and by a morphometric study of lesions. Evident agreement was observed among results obtained by the different methods in all of the animals studied. Microbiologically, the efficacy of GM237354 was determined by measuring the number of C. albicans organisms in the oral cavities of rats in the middle (day 4) and at the end (day 7) of the treatment. GM237354 administered at 5, 7.5, 10, 15, or 30 mg/kg/day for 7 days significantly reduced the number of CFU in the oral cavities of treated rats compared with the number of CFU in the oral cavities of the untreated controls. A significant reduction was also observed when GM237354 was administered at 7.5, 10, 15, or 30 mg/kg/day for 4 days. Furthermore, C. albicans was not detected in oral swabs from any infected rats after 1 week of treatment when GM237354 was administered at 15 or 30 mg/kg/day or after 4 days of treatment at 30 mg/kg/day. Histologically, untreated control animals showed extensive colonization of the epithelium of the dorsal tongue by numerous hyphae. Animals treated with GM237354 at 7.5 mg/kg/day showed small areas with superficial hyphal penetration into the epithelium that produced intraepithelial microabscesses. However, animals treated with GM237354 at 15 mg/kg/day showed multiple regenerative areas of the covering epithelium, and only focalized zones of the tongue surface were occupied by hyphae. No hyphal colonization of the epithelium was seen in rats treated with GM237354 at 30 mg/kg/day and which showed extensive areas of epithelial regeneration of the tongue. The histopathology findings were confirmed by morphometry studies, and the percentage of epithelium occupied by C. albicans hyphae decreased from 17.5% in the control group to 4.8 and 0.1% in animals treated with GM237354 at 7.5 and 15 mg/kg/day, respectively. These results demonstrated that the sordarin derivative GM237354 was effective against experimental oral candidiasis in immunosuppressed rats, and further studies are needed to determine the potential of GM237354 for use in the treatment of this infection in humans. PMID- 11257010 TI - Xenograft model for identifying chemotherapeutic agents against papillomaviruses. AB - The report describes the establishment and characterization of a mouse xenograft transplantation model for the study of papillomavirus infection of bovine skin. Calf scrotal skin was inoculated with bovine papillomavirus type 2 before grafting it to the dorsum of severe combined immunodeficient mice. The grafted skin contained epidermis, dermis, and a thin layer of fat. After 5 months the induced warts not only showed histological features of papillomavirus infections but also tested positive for viral DNA and papillomavirus capsid antigen. The formation of infectious virions was demonstrated by inoculation of new transplants with crude extract from the induced warts as well as in a cell culture focus assay. Topical application of bromovinyl-2'-deoxyuridine led to a reduction in viral DNA content in the developing wart. This small-animal xenograft model should be useful for characterizing antiviral compounds and providing an understanding of the regulation of papillomavirus infections. PMID- 11257011 TI - Rapid screening technique for class 1 integrons in Enterobacteriaceae and nonfermenting gram-negative bacteria and its use in molecular epidemiology. AB - A screening technique for integrons in members of the family Enterobacteriaceae and nonfermenting gram-negative bacteria by real-time PCR is reported. A total of 226 isolates of gram-negative bacteria obtained from a variety of clinical specimens were screened for class 1 integrons by real-time PCR performed on a LightCycler instrument. This technique used a primer pair specific for a 300-bp conserved region at the 5' ends of class 1 integrons. The screening assay was evaluated by comparison with results obtained by the conventional, thermal-block PCR (long PCR) by using established conditions and primers for the detection of class 1 integrons, and the real-time PCR technique was thus shown to be both sensitive and specific. DNA from 50 of 226 (22%) isolates screened was identified as containing an integron by the screening PCR, and sequence data were obtained across the integron for 34 of 50 (68%) of these isolates. In an attempt to study the molecular epidemiology of antimicrobial resistance genes carried within integrons, a comparison of the types of gene cassettes carried by isolates from different patients was made. Adenyltransferase genes conferring resistance to streptomycin and spectinomycin were the predominant gene cassettes amplified in the study. Resistance to trimethoprim was also frequently found to be encoded within integrons. Furthermore, multiple bacterial isolates obtained from one patient over a 5-month period were all shown to carry an integron containing the same single adenyltransferase gene cassette, suggesting that these elements were relatively stable in this case. PMID- 11257012 TI - Efficacies of gel formulations containing foscarnet, alone or combined with sodium lauryl sulfate, against establishment and reactivation of latent herpes simplex virus type 1. AB - The influence of sodium lauryl sulfate (SLS) on the efficacies of gel formulations of foscarnet against herpes simplex virus type 1 (HSV-1) cutaneous lesions and on the establishment and reactivation of latent virus has been evaluated in a murine model of orofacial infection. Topical treatments were given twice daily for 3 days and were initiated at 6, 24, and 48 h after virus inoculation. The gel formulation that contained both 3% foscarnet and 5% SLS and that was administered within 48 h postinfection reduced the rate of development of herpetic skin lesions. This formulation also significantly decreased the viral content in skin tissues and in ipsilateral trigeminal ganglia when it was given within 24 and 6 h postinfection, respectively. A lower level of efficacy was observed for the gel formulation containing 3% foscarnet alone. Of prime interest, the gel formulation containing 5% SLS reduced significantly the mortality rate among mice in a zosteriform model of infection. Both formulations of foscarnet had no effect on the mean titers of reactivated virus in explant cultures of ipsilateral and contralateral trigeminal ganglia from latently infected mice. The use of a gel formulation containing combinations of foscarnet and SLS could represent an attractive approach for the treatment of herpetic mucocutaneous infections. PMID- 11257013 TI - Need for annual surveillance of antimicrobial resistance in Streptococcus pneumoniae in the United States: 2-year longitudinal analysis. AB - Although changing patterns in antimicrobial resistance in Streptococcus pneumoniae have prompted several surveillance initiatives in recent years, the frequency with which these studies are needed has not been addressed. To approach this issue, the extent to which resistance patterns change over a 1-year period was examined. In this study we analyzed S. pneumoniae antimicrobial susceptibility results produced in our laboratory with isolates obtained over 2 consecutive years (1997-1998 and 1998-1999) from the same 96 institutions distributed throughout the United States. Comparison of results revealed increases in resistant percentages for all antimicrobial agents studied except vancomycin. For four of the agents tested (penicillin, cefuroxime, trimethoprim sulfamethoxazole, and levofloxacin), the increases were statistically significant (P < 0.05). Resistance to the fluoroquinolone remained low in both years (0.1 and 0.6%, respectively); in contrast, resistance to macrolides was consistently greater than 20%, and resistance to trimethoprim-sulfamethoxazole increased from 13.3 to 27.3%. Multidrug resistance, concurrent resistance to three or more antimicrobials of different chemical classes, also increased significantly between years, from 5.9 to 11%. The most prevalent phenotype was resistance to penicillin, azithromycin (representative macrolide), and trimethoprim sulfamethoxazole. Multidrug-resistant phenotypes that included fluoroquinolone resistance were uncommon; however, two phenotypes that included fluoroquinolone resistance not found in 1997-1998 were encountered in 1998-1999. This longitudinal surveillance study of resistance in S. pneumoniae revealed that significant changes do occur in just a single year and supports the need for surveillance at least on an annual basis, if not continuously. PMID- 11257014 TI - Specific inhibition of coxsackievirus B3 translation and replication by phosphorothioate antisense oligodeoxynucleotides. AB - The 5' and 3' untranslated regions (UTRs) of coxsackievirus B3 (CVB3) RNA form highly ordered secondary structures that have been confirmed to play important regulatory roles in viral cap-independent internal translation initiation and RNA replication. We previously demonstrated that deletions in different regions of the 5' UTR significantly reduced viral RNA translation and infectivity. Such observations suggested strongly that viral RNA translation and replication could be blocked if highly specific antisense oligodeoxynucleotides (AS-ODNs) were applied to target crucial sites within the 5' and 3' UTRs. In this study, seven phosphorothioate AS-ODNs were synthesized, and the antiviral activity was evaluated by Lipofectin transfection of HeLa cells with AS-ODNs followed by infection of CVB3. Analysis by Western blotting, reverse transcription-PCR, and viral plaque assay demonstrated that viral protein synthesis, genome replication, and infectivity of CVB3 were strongly inhibited by the AS-ODNs complementary to different regions of the 5' and 3' UTRs. The most effective sites are located at the proximate terminus of the 5' UTR (AS-1), the proximate terminus of the 3' UTR (AS-7), the core sequence of the internal ribosome entry site (AS-2), and the translation initiation codon region (AS-4). These AS-ODNs showed highly sequence specific and dose-dependent inhibitory effects on both viral protein synthesis and RNA replication. It is noteworthy that the highest inhibitory activities were obtained with AS-1 and AS-7 targeting the termini of the 5' and 3' UTRs. The percent inhibition values of AS-1 and AS-7 for CVB3 protein VP1 synthesis and RNA replication were 70.6 and 79.6 for AS-1 and 73.7 and 79.7 for AS-7, respectively. These data suggest that CVB3 infectivity can be inhibited effectively by AS-ODNs. PMID- 11257015 TI - Peptide deformylase as an antibacterial drug target: assays for detection of its inhibition in Escherichia coli cell homogenates and intact cells. AB - An assay was developed to determine the activity of peptide deformylase (PDF) inhibitors under conditions as close as possible to the physiological situation. The assay principle is the detection of N-terminal [35S]methionine labeling of a protein that contains no internal methionine. If PDF is active, the deformylation of the methionine renders the peptide a substrate for methionine aminopeptidase, resulting in the removal of the N-terminal methionine label. In the presence of a PDF inhibitor, the deformylation is blocked so that the N-formylated peptide is not processed and the label is detected. Using this assay, it is possible to determine the PDF activity under near-physiological conditions in a cell-free transcription-translation system as well as in intact bacterial cells. PMID- 11257016 TI - Peptide deformylase as an antibacterial drug target: target validation and resistance development. AB - New inhibitors of peptide deformylase (PDF) which are very potent against the isolated enzyme and show a certain degree of antibacterial activity have recently been synthesized by our group. Several lines of experimental evidence indicate that these inhibitors indeed interfere with the target enzyme in the bacterial cell. (i) The inhibition of Escherichia coli growth could be counteracted by overexpression of PDF from different organisms, including E. coli, Streptococcus pneumoniae, and Haemophilus influenzae. Conversely, reduced expression of PDF in S. pneumoniae resulted in an increased susceptibility to the inhibitors. (ii) Proteome analysis on two-dimensional gels revealed a shift for many proteins towards lower pI in the presence of PDF inhibitors, as would be expected if the proteins still carry their N-formyl-Met terminus. (iii) PDF inhibitors show no antimicrobial activity against E. coli under conditions that make growth independent of formylation and deformylation. The antibacterial activity in E. coli was characterized as bacteriostatic. Furthermore, the development of resistance in E. coli was observed to occur with high frequency (10(-7)). Resistant mutants show a reduced growth rate, and DNA sequence analysis revealed mutations in their formyl transferase gene. Taking all these aspects into account, we conclude that PDF may not be an optimal target for broad-spectrum antibacterial agents. PMID- 11257018 TI - Penicillin pharmacodynamics in four experimental pneumococcal infection models. AB - Clinical and animal studies indicate that with optimal dosing, penicillin may still be effective against penicillin-nonsusceptible pneumococci (PNSP). The present study examined whether the same strains of penicillin-susceptible pneumococci (PSP) and PNSP differed in their pharmacodynamic responses to penicillin by using comparable penicillin dosing regimens in four animal models: peritonitis, pneumonia, and thigh infection in mice and tissue cage infection in rabbits. Two multidrug-resistant isolates of Streptococcus pneumoniae type 6B were used, one for which the penicillin MIC was 0.016 microg/ml and the other for which the penicillin MIC was 1.0 microg/ml. Two additional strains of PNSP were studied in the rabbit. The animals were treated with five different penicillin regimens resulting in different maximum concentrations of drugs in serum (C(max)s) and times that the concentrations were greater than the MIC (T(>MIC)s). The endpoints were bacterial viability counts after 6 h of treatment in the mice and 24 h of treatment in the rabbits. Similar pharmacodynamic effects were observed in all models. In the mouse models bactericidal activity depended on the T(>MIC) and to a lesser extent on the Cmax/MIC and the generation time but not on the area under the concentration-time curve (AUC)/MIC. Maximal bactericidal activities were similar for both PSP and PNSP, being the highest in the peritoneum and blood (approximately 6 log10 CFU/ml), followed by the thigh (approximately 3 log10 CFU/thigh), and being the lowest in the lung (approximately 1 log10 CFU/lung). In the rabbit model the maximal effect was approximately 6 log10 CFU/ml after 24 h. In the mouse models bactericidal activity became marked when T(>MIC) was > or =65% of the experimental time and C(max) was > or =15 times the MIC, and in the rabbit model bactericidal activity became marked when T(>MIC) was > or =35%, Cmax was > or =5 times the MIC, and the AUC at 24 h/MIC exceeded 25. By optimization of the Cmax/MIC ratio and T(>MIC), the MIC of penicillin for pneumococci can be used to guide therapy and maximize therapeutic efficacy in nonmeningeal infections caused by PNSP. PMID- 11257017 TI - Antiviral activity of beta-L-2',3'-dideoxy-2',3'-didehydro-5-fluorocytidine in woodchucks chronically infected with woodchuck hepatitis virus. AB - The L-nucleoside analog beta-L-2',3'-dideoxy-2',3'-didehydro-5-fluorocytidine (beta-L-Fd4C) was first shown to exhibit potent activity against hepatitis B virus (HBV) in tissue culture and then to significantly inhibit viral spread during acute infection in the duck HBV model (F. Le Guerhier et al., Antimicrob. Agents Chemother. 44:111-122, 2000). We have therefore examined its antiviral activity in a mammalian model of chronic HBV infection, the woodchuck chronically infected with woodchuck hepatitis virus (WHV). Side-by-side comparison of beta-L Fd4C and lamivudine administered intraperitoneally during short-term and long term protocols demonstrated a more profound inhibition of viremia in beta-L-Fd4C treated groups. Moreover, beta-L-Fd4C induced a marked inhibition of intrahepatic viral DNA synthesis compared with that induced by lamivudine. Nevertheless, covalently closed circular (CCC) DNA persistence explained the lack of clearance of infected hepatocytes expressing viral antigens and the relapse of WHV replication after drug withdrawal. Liver histology showed a decrease in the inflammatory activity of chronic hepatitis in woodchucks receiving beta-L-Fd4C. An electron microscopy study showed the absence of ultrastructural changes of hepatic mitochondria, biliary canaliculi, and bile ducts. However, a loss of weight was observed in all animals, whatever the treatment, as was a transient skin pigmentation in all woodchucks during beta-L-Fd4C treatment. There was no evidence that lamivudine or beta-L-Fd4C could prevent the development of hepatocellular carcinoma with the protocols used. These results indicate that beta-L-Fd4C exhibits a more potent antiviral effect than lamivudine in the WHV model but was not able to eradicate CCC DNA and infected cells from the liver at the dosage and with the protocol used. PMID- 11257019 TI - Circulating metabolites of the human immunodeficiency virus protease inhibitor nelfinavir in humans: structural identification, levels in plasma, and antiviral activities. AB - Nelfinavir mesylate (Viracept, formally AG1343) is a potent and orally bioavailable human immunodeficiency virus (HIV) type 1 (HIV-1) protease inhibitor (K(i) = 2 nM) and is being widely prescribed in combination with HIV reverse transcriptase inhibitors for the treatment of HIV infection. The current studies evaluated the presence of metabolites circulating in plasma following the oral administration of nelfinavir to healthy volunteers and HIV-infected patients, as well as the levels in plasma and antiviral activities of these metabolites. The results showed that the parent drug was the major circulating chemical species, followed in decreasing abundance by its hydroxy-t-butylamide metabolite (M8) and 3'-methoxy-4'-hydroxynelfinavir (M1). Antiviral assays with HIV-1 strain RF infected CEM-SS cells showed that the 50% effective concentrations (EC50) of nelfinavir, M8, and M1 were 30, 34, and 151 nM, respectively, and that the corresponding EC50 against another HIV-1 strain, IIIB, in MT-2 cells were 60, 86, and 653 nM. Therefore, apparently similar in vitro antiviral activities were demonstrated for nelfinavir and M8, whereas an approximately 5- to 11-fold-lower level of antiviral activity was observed for M1. The active metabolite, M8, showed a degree of binding to human plasma proteins similar to that of nelfinavir (ca. 98%). Concentrations in plasma of nelfinavir and its metabolites in 10 HIV positive patients receiving nelfinavir therapy (750 mg three times per day) were determined by a liquid chromatography tandem mass spectrometry assay. At steady state (day 28), the mean plasma nelfinavir concentrations ranged from 1.73 to 4.96 microM and the M8 concentrations ranged from 0.55 to 1.96 microM, whereas the M1 concentrations were low and ranged from 0.09 to 0.19 microM. In conclusion, the findings from the current studies suggest that, in humans, nelfinavir forms an active metabolite circulating at appreciable levels in plasma. The active metabolite M8 may account for some of the antiviral activity associated with nelfinavir in the treatment of HIV disease. PMID- 11257020 TI - Randomized prospective controlled trial of recombinant granulocyte colony stimulating factor as adjunctive therapy for limb-threatening diabetic foot infection. AB - Adult diabetic patients admitted to our Diabetes Center from September 1996 to January 1998 for severe, limb-threatening foot infection were consecutively enrolled in a prospective, randomized, controlled clinical study aimed at assessing the safety and efficacy of recombinant human granulocyte colony stimulating factor (G-CSF) (lenograstim) as an adjunctive therapy for the standard treatment of diabetic foot infection. Forty patients, all of whom displayed evidence of osteomyelitis and long-standing ulcer infection, were randomized 1:1 to receive either conventional treatment (i.e., antimicrobial therapy plus local treatment) or conventional therapy plus 263 microg of G-CSF subcutaneously daily for 21 days. The empiric antibiotic treatment (a combination of ciprofloxacin plus clindamycin) was further adjusted, when necessary, according to the results of cultures and sensitivity testing. Microbiologic assessment of foot ulcers was performed by both deep-tissue biopsy and swab cultures, performed at enrollment and on days 7 and 21 thereafter. Patients were monitored for 6 months; the major endpoints (i.e., cure, improvement, failure, and amputation) were blindly assessed at weeks 3 and 9. At enrollment, both patient groups were comparable in terms of both demographic and clinical data. None of the G-CSF-treated patients experienced either local or systemic adverse effects. At the 3- and 9-week assessments, no significant differences between the two groups could be observed concerning the number of patients either cured or improved, the number of patients displaying therapeutic failure, or the species and number of microorganisms previously yielded from cultures at day 7 and day 21. Conversely, among this small series of patients the cumulative number of amputations observed after 9 weeks of treatment appeared to be lower in the G-CSF arm; in fact, only three patients (15%) in this group had required amputation, whereas nine patients (45%) in the other group had required amputation (P = 0.038). In conclusion, the administration of G-CSF for 3 weeks as an adjunctive therapy for limb-threatening diabetic foot infection was associated with a lower rate of amputation within 9 weeks after the commencement of standard treatment. Further clinical studies aimed at precisely defining the role of this approach to this serious complication of diabetes mellitus appear to be justified. PMID- 11257021 TI - rho is not essential for viability or virulence in Staphylococcus aureus. AB - We have identified the gene for transcription termination factor Rho in Staphylococcus aureus. Deletion of rho in S. aureus reveals that it is not essential for viability or virulence. We also searched the available bacterial genomic sequences for homologs of Rho and found that it is broadly distributed and highly conserved. Exceptions include Streptococcus pneumoniae, Streptococcus pyogenes, Mycoplasma genitalium, Mycoplasma pneumoniae, Ureaplasma urealyticum, and Synechocystis sp. strain PCC6803, all of which appear not to possess a Rho homolog. Complementation studies indicate that S. aureus Rho possesses the same activity as Escherichia coli Rho and that the Rho inhibitor bicyclomycin is active against S. aureus Rho. Our results explain the lack of activity of bicyclomycin against many gram-positive bacteria and raise the possibility that the essentiality of rho may be the exception rather than the rule. PMID- 11257022 TI - Multiple mutations modulate the function of dihydrofolate reductase in trimethoprim-resistant Streptococcus pneumoniae. AB - Trimethoprim resistance in Streptococcus pneumoniae can be conferred by a single amino acid substitution (I100-L) in dihydrofolate reductase (DHFR), but resistant clinical isolates usually carry multiple DHFR mutations. DHFR genes from five trimethoprim-resistant isolates from the United Kingdom were compared to susceptible isolates and used to transform a susceptible control strain (CP1015). All trimethoprim-resistant isolates and transformants contained the I100-L mutation. The properties of DHFRs from transformants with different combinations of mutations were compared. In a transformant with only the I100-L mutation (R12/T2) and a D92-A mutation also found in the DHFRs of susceptible isolates, the enzyme was much more resistant to trimethoprim inhibition (50% inhibitory concentration [IC50], 4.2 microM) than was the DHFR from strain CP1015 (IC50, 0.09 microM). However, Km values indicated a lower affinity for the enzyme's natural substrates (Km for dihydrofolate [DHF], 3.1 microM for CP1015 and 27.5 microM for R12/T2) and a twofold decrease in the specificity constant. In transformants with additional mutations in the C-terminal portion of the enzyme, Km values for DHF were reduced (9.2 to 15.2 microM), indicating compensation for the lower affinity generated by I100-L. Additional mutations in the N-terminal portion of the enzyme were associated with up to threefold-increased resistance to trimethoprim (IC50 of up to 13.7 microM). It is postulated that carriage of the mutation M53-I-which, like I100-L, corresponds to a trimethoprim binding site in the Escherichia coli DHFR-is responsible for this increase. This study demonstrates that although the I100-L mutation alone may give rise to trimethoprim resistance, additional mutations serve to enhance resistance and modulate the effects of existing mutations on the affinity of DHFR for its natural substrates. PMID- 11257023 TI - Mdt(A), a new efflux protein conferring multiple antibiotic resistance in Lactococcus lactis and Escherichia coli. AB - The mdt(A) gene, previously designated mef214, from Lactococcus lactis subsp. lactis plasmid pK214 encodes a protein [Mdt(A) (multiple drug transporter)] with 12 putative transmembrane segments (TMS) that contain typical motifs conserved among the efflux proteins of the major facilitator superfamily. However, it also has two C-motifs (conserved in the fifth TMS of the antiporters) and a putative ATP-binding site. Expression of the cloned mdt(A) gene decreased susceptibility to macrolides, lincosamides, streptogramins, and tetracyclines in L. lactis and Escherichia coli, but not in Enterococcus faecalis or in Staphylococcus aureus. Glucose-dependent efflux of erythromycin and tetracycline was demonstrated in L. lactis and in E. coli. PMID- 11257024 TI - Quinolone resistance in Staphylococci: activities of new nonfluorinated quinolones against molecular targets in whole cells and clinical isolates. AB - The activity of three new, 8-methoxy-nonfluorinated quinolones (NFQs) against multiple-drug-resistant staphylococci was investigated. First, using Staphylococcus aureus strains containing point mutations in the serine 84-80 hot spots of the target genes (gyrA and grlA), cell growth inhibition potencies of the NFQs as a result of DNA gyrase and topoisomerase IV inhibition were estimated and compared with those of known fluoroquinolones. The NFQs and clinafloxacin showed higher affinities toward both the targets than ciprofloxacin, trovafloxacin and gatifloxacin. Furthermore, the ratio of the calculated affinity parameter for DNA gyrase to that for topoisomerase IV was lower in the case of the NFQs, clinafloxacin, and gatifloxacin than in the case of ciprofloxacin and trovafloxacin. These results suggest that the former group of quinolones is better able to exploit both the targets. Next, using clinical isolates of methicillin-resistant S. aureus (MRSA; n = 34) and coagulase-negative staphylococci (CoNS; n = 24), the NFQs and clinafloxacin were shown to be more potent (MIC at which 90% of the isolates are inhibited [MIC90] = 2 microg/ml for MRSA and 0.5 microg/ml for CoNS) than ciprofloxacin, trovafloxacin, and gatifloxacin (MIC90 = 16 to >64 microg/ml for MRSA and 4 to >32 microg/ml for CoNS). Bactericidal kinetics experiments, using two MRSA isolates, showed that exposure to the NFQs at four times the MIC reduced the bacterial counts (measured in CFU per milliliter) by > or =3 log units in 2 to 4 h. Overall, the NFQs and clinafloxacin were less susceptible than the other quinolones to existing mechanisms of quinolone resistance in staphylococci. PMID- 11257025 TI - In vivo monitoring of intracellular ATP levels in Leishmania donovani promastigotes as a rapid method to screen drugs targeting bioenergetic metabolism. AB - A method for the rapid screening of drugs targeting the bioenergetic metabolism of Leishmania spp. was developed. The system is based on the monitoring of changes in the intracellular ATP levels of Leishmania donovani promastigotes that occur in vivo, as assessed by the luminescence produced by parasites transfected with a cytoplasmic form of Phothinus pyralis luciferase and incubated with free membrane permeable D-luciferin analogue D-luciferin-[1-(4,5-dimethoxy-2 nitrophenyl) ethyl ester]. A significant correlation was obtained between the rapid inhibition of luminescence with parasite proliferation and the dissipation of changes in mitochondrial membrane potential (DeltaPsi(m)) produced by buparvaquone or plumbagin, two leishmanicidal inhibitors of oxidative phosphorylation. To further validate this test, a screen of 14 standard leishmanicidal drugs, using a 50 microM cutoff, was carried out. Despite its semiquantitative properties and restriction to the promastigote stage, this test compares favorably with other bioenergetic parameters with respect to time and cell number requirements for the screening of drugs that affect mitochondrial activity. PMID- 11257026 TI - Antibiotic susceptibility profiles of Escherichia coli strains lacking multidrug efflux pump genes. AB - The contribution of seven known and nine predicted genes or operons associated with multidrug resistance to the susceptibility of Escherichia coli W3110 was assessed for 20 different classes of antimicrobial compounds that include antibiotics, antiseptics, detergents, and dyes. Strains were constructed with deletions for genes in the major facilitator superfamily, the resistance nodulation-cell division family, the small multidrug resistance family, the ATP binding cassette family, and outer membrane factors. The agar dilution MICs of 35 compounds were determined for strains with deletions for multidrug resistance (MDR) pumps. Deletions in acrAB or tolC resulted in increased susceptibilities to the majority of compounds tested. The remaining MDR pump gene deletions resulted in increased susceptibilities to far fewer compounds. The results identify which MDR pumps contribute to intrinsic resistance under the conditions tested and supply practical information useful for designing sensitive assay strains for cell-based screening of antibacterial compounds. PMID- 11257027 TI - Identification of genes induced by a macrophage activator, S-28463, using gene expression array analysis. AB - S-28463 and imiquimod are imidazoquinoline compounds which stimulate microbicidal activity by inducing a local immune response at the site of application. Imiquimod-containing cream is an effective clinical treatment against cervical warts caused by human papillomavirus infection. Imiquimod also induces leishmanicidal activity both in vitro in macrophages and in vivo in a mouse model for cutaneous leishmaniasis. The major target cells of S-28463 and imiquimod are macrophages. To explore the molecular basis in which imidazoquinolines generate macrophage microbicidal activity, a cDNA gene array analysis was undertaken to identify genes induced by S-28463. Out of 588 genes screened in this assay, only 13 genes were significantly induced by S-28463. Remarkably, virtually all of the induced genes are involved in macrophage activation and inflammatory response. This experimental approach defines the mechanism of action of this clinically relevant compound in the induction of microbicidal activity in macrophages and also potentially identifies novel genes associated with microbicidal activity in this cell type. PMID- 11257028 TI - In vitro activity of a novel antimycobacterial compound, N octanesulfonylacetamide, and its effects on lipid and mycolic acid synthesis. AB - beta-Sulfonyl carboxamides have been proposed to serve as transition-state analogues of the beta-ketoacyl synthase reaction involved in fatty acid elongation. We tested the efficacy of N-octanesulfonylacetamide (OSA) as an inhibitor of fatty acid and mycolic acid biosynthesis in mycobacteria. Using the BACTEC radiometric growth system, we observed that OSA inhibits the growth of several species of slow-growing mycobacteria, including Mycobacterium tuberculosis (H37Rv and clinical isolates), the Mycobacterium avium complex (MAC), Mycobacterium bovis BCG, Mycobacterium kansasii, and others. Nearly all species and strains tested, including isoniazid and multidrug resistant isolates of M. tuberculosis, were susceptible to OSA, with MICs ranging from 6.25 to 12.5 microg/ml. Only three clinical isolates of M. tuberculosis (CSU93, OT2724, and 401296), MAC, and Mycobacterium paratuberculosis required an OSA MIC higher than 25.0 microg/ml. Rapid-growing mycobacterial species, such as Mycobacterium smegmatis, Mycobacterium fortuitum, and others, were not susceptible at concentrations of up to 100 microg/ml. A 2-dimensional thin-layer chromatography system showed that OSA treatment resulted in a significant decrease in all species of mycolic acids present in BCG. In contrast, mycolic acids in M. smegmatis were relatively unaffected following exposure to OSA. Other lipids, including polar and nonpolar extractable classes, were unchanged following exposure to OSA in both BCG and M. smegmatis. Transmission electron microscopy of OSA-treated BCG cells revealed a disruption in cell wall synthesis and incomplete septum formation. Our results indicate that OSA inhibits the growth of several species of mycobacteria, including both isoniazid-resistant and multidrug resistant strains of M. tuberculosis. This inhibition may be the result of OSA mediated effects on mycolic acid synthesis in slow-growing mycobacteria or inhibition via an undescribed mechanism. Our results indicate that OSA may serve as a promising lead compound for future antituberculous drug development. PMID- 11257029 TI - Novel carbapenem-hydrolyzing beta-lactamase, KPC-1, from a carbapenem-resistant strain of Klebsiella pneumoniae. AB - A Klebsiella pneumoniae isolate showing moderate to high-level imipenem and meropenem resistance was investigated. The MICs of both drugs were 16 microg/ml. The beta-lactamase activity against imipenem and meropenem was inhibited in the presence of clavulanic acid. The strain was also resistant to extended-spectrum cephalosporins and aztreonam. Isoelectric focusing studies demonstrated three beta-lactamases, with pIs of 7.2 (SHV-29), 6.7 (KPC-1), and 5.4 (TEM-1). The presence of bla(SHV) and bla(TEM) genes was confirmed by specific PCRs and DNA sequence analysis. Transformation and conjugation studies with Escherichia coli showed that the beta-lactamase with a pI of 6.7, KPC-1 (K. pneumoniae carbapenemase-1), was encoded on an approximately 50-kb nonconjugative plasmid. The gene, bla(KPC-1), was cloned in E. coli and shown to confer resistance to imipenem, meropenem, extended-spectrum cephalosporins, and aztreonam. The amino acid sequence of the novel carbapenem-hydrolyzing beta-lactamase, KPC-1, showed 45% identity to the pI 9.7 carbapenem-hydrolyzing beta-lactamase, Sme-1, from Serratia marcescens S6. Hydrolysis studies showed that purified KPC-1 hydrolyzed not only carbapenems but also penicillins, cephalosporins, and monobactams. KPC-1 had the highest affinity for meropenem. The kinetic studies also revealed that clavulanic acid and tazobactam inhibited KPC-1. An examination of the outer membrane proteins of the parent K. pneumoniae strain demonstrated that the strain does not express detectable levels of OmpK35 and OmpK37, although OmpK36 is present. We concluded that carbapenem resistance in K. pneumoniae strain 1534 is mainly due to production of a novel Bush group 2f, class A, carbapenem hydrolyzing beta-lactamase, KPC-1, although alterations in porin expression may also play a role. PMID- 11257030 TI - Comparison of the anti-influenza virus activity of RWJ-270201 with those of oseltamivir and zanamivir. AB - We have recently reported an influenza virus neuraminidase inhibitor, RWJ-270201 (BCX-1812), a novel cyclopentane derivative discovered through structure-based drug design. In this paper, we compare the potency of three compounds, RWJ 270201, oseltamivir, and zanamivir, against neuraminidase enzymes from various subtypes of influenza. RWJ-270201 effectively inhibited all tested influenza A and influenza B neuraminidases in vitro, with 50% inhibitory concentrations of 0.09 to 1.4 nM for influenza A neuraminidases and 0.6 to 11 nM for influenza B neuraminidases. These values were comparable to or lower than those for oseltamivir carboxylate (GS4071) and zanamivir (GG167). RWJ-270201 demonstrated excellent selectivity (>10,000-fold) for influenza virus neuraminidase over mammalian, bacterial, or other viral neuraminidases. Oral administration of a dosage of 1 mg/kg of body weight/day of RWJ-270201 for 5 days (beginning 4 h preinfection) showed efficacy in the murine model of influenza virus infection as determined by lethality and weight loss protection. RWJ-270201 administered intranasally at 0.01 mg/kg/day in the murine influenza model demonstrated complete protection against lethality, whereas oseltamivir carboxylate and zanamivir at the same dose demonstrated only partial protection. In the delayed treatment murine influenza model, oral administration of a 10-mg/kg/day dose of RWJ-270201 or oseltamivir (GS4104, a prodrug of GS4071) at 24 h postinfection showed significant protection against lethality (P < 0.001 versus control). However, when the treatment was delayed for 48 h, no significant protection was observed in either drug group. No drug-related toxicity was observed in mice receiving 100 mg/kg/day of RWJ-270201 for 5 days. These efficacy and safety profiles justify further consideration of RWJ-270201 for the treatment and prevention of human influenza. PMID- 11257031 TI - DNA transformation of Leishmania infantum axenic amastigotes and their use in drug screening. AB - Protocols for DNA electroporation in Leishmania promastigote cells are well established. More recently, in vitro culture of axenic Leishmania amastigotes became possible. We have established conditions for DNA transformation of axenically grown Leishmania infantum amastigotes. Parameters for DNA electroporation of Leishmania axenic amastigotes were systematically studied using luciferase-mediated transient transfection. Cell lines expressing stable luciferase activity were then selected, and their ability to be used in an in vitro drug screening procedure was determined. A model was established, using axenic amastigotes expressing luciferase activity, for rapidly determining the activity of drugs directly against both axenic and intracellular amastigotes. For intracellular amastigotes, the 50% effective concentrations of pentamidine, sodium stibogluconate (Pentostam), meglumine (Glucantime), and potassium antimonyl tartrate determined with the luciferase assay were 0.2 microM (0.12 microg/ml), 55 microg/ml, 95 microg/ml, and 0.12 microg/ml, respectively; these values are in agreement with values determined by more labor-intensive staining methods. We also showed the usefulness of luciferase-expressing parasites for analyzing drug resistance. The availability of luciferase-expressing amastigotes for use in high-throughput screening should facilitate the search for new antileishmanial drugs. PMID- 11257032 TI - Role of ATP-binding-cassette transporter genes in high-frequency acquisition of resistance to azole antifungals in Candida glabrata. AB - Candida glabrata has been often isolated from AIDS patients with oropharyngeal candidiasis treated with azole antifungal agents, especially fluconazole. We recently showed that the ATP-binding-cassette (ABC) transporter gene CgCDR1 was upregulated in C. glabrata clinical isolates resistant to azole antifungal agents (D. Sanglard, F. Ischer, D. Calabrese, P. A. Majcherczyk, and J. Bille, Antimicrob. Agents Chemother. 43:2753-2765, 1999). Deletion of CgCDR1 in C. glabrata rendered the null mutant hypersusceptible to azole derivatives and showed the importance of this gene in mediating azole resistance. We observed that wild-type C. glabrata exposed to fluconazole in a medium containing the drug at 50 microg/ml developed resistance to this agent and other azoles at a surprisingly high frequency (2 x 10(-4) to 4 x 10(-4)). We show here that this high-frequency azole resistance (HFAR) acquired in vitro was due, at least in part, to the upregulation of CgCDR1. The CgCDR1 deletion mutant DSY1041 could still develop HFAR but in a medium containing fluconazole at 5 microg/ml. In the HFAR strain derived from DSY1041, a distinct ABC transporter gene similar to CgCDR1, called CgCDR2, was upregulated. This gene was slightly expressed in clinical isolates but was upregulated in strains with the HFAR phenotype. Deletion of both CgCDR1 and CgCDR2 suppressed the development of HFAR in a medium containing fluconazole at 5 microg/ml, showing that both genes are important mediators of resistance to azole derivatives in C. glabrata. We also show here that the HFAR phenomenon was linked to the loss of mitochondria in C. glabrata. Mitochondrial loss could be obtained by treatment with ethidium bromide and resulted in acquisition of resistance to azole derivatives without previous exposure to these agents. Azole resistance obtained in vitro by HFAR or by agents stimulating mitochondrial loss was at least linked to the upregulation of both CgCDR1 and CgCDR2. PMID- 11257034 TI - Potentiation of inhibition of wild-type and mutant human immunodeficiency virus type 1 reverse transcriptases by combinations of nonnucleoside inhibitors and d- and L-(beta)-dideoxynucleoside triphosphate analogs. AB - Combinations of reverse transcriptase (RT) inhibitors are currently used in anti human immunodeficiency virus therapy in order to prevent or delay the emergence of resistant virus and to improve the efficacy against viral enzymes carrying resistance mutations. Drug-drug interactions can result in either positive (additive or synergistic inhibition) or adverse (antagonistic interaction, synergistic toxicity) effects. Elucidation of the nature of drug interaction would help to rationalize the choice of antiretroviral agents to be used in combination. In this study, different combinations of nucleoside and nonnucleoside inhibitors, including D- and L-(beta)-deoxy- and -dideoxynucleoside triphosphate analogues, have been tested in in vitro RT assays against either recombinant wild-type RT or RT bearing clinically relevant nonnucleoside inhibitor resistance mutations (L100I, K103N, Y181I), and the nature of the interaction (either synergistic or antagonistic) of these associations was evaluated. The results showed that (i) synergy of a combination was not always equally influenced by the individual agents utilized, (ii) a synergistic combination could improve the sensitivity profile of a drug-resistant mutant enzyme to the single agents utilized, (iii) L-(beta)-enantiomers of nucleoside RT inhibitors were synergistic when combined with nonnucleoside RT inhibitors, and (iv) inter- and intracombination comparisons of the relative potencies of each drug could be used to highlight the different contributions of each drug to the observed synergy. PMID- 11257033 TI - Amphotericin B lipid complex or amphotericin B multiple-dose administration to rabbits with elevated plasma cholesterol levels: pharmacokinetics in plasma and blood, plasma lipoprotein levels, distribution in tissues, and renal toxicities. AB - The purpose of the present study was to determine if a relationship exists between the plasma cholesterol concentration, the severity of amphotericin B (AmpB)-induced renal toxicity, and the pharmacokinetics of AmpB in plasma in hypercholesterolemic rabbits administered multiple doses of amphotericin B (AmB) deoxycholate (Doc-AmB) and AmB lipid complex (ABLC). After 7 days of administration of a cholesterol-enriched diet (0.50% [wt/vol]) or a regular rabbit diet, each rabbit was administered a single intravenous bolus of Doc-AmB (n = 8) or ABLC (n = 10) (1.0 mg/kg of body weight) daily for 7 consecutive days (a total of eight doses). Blood samples were obtained daily before and 24 h after the administration of each dose and serially thereafter following the administration of the last dose for the assessment of pharmacokinetics in plasma, kidney toxicity, plasma lipoprotein levels, and drug distribution in tissue. The pharmacokinetics of AmB in blood following the administration of ABLC were also determined in rabbits fed cholesterol-enriched and regular diets (n = 3 each group). Before drug treatment, cholesterol-fed rabbits demonstrated marked increases in total, low-density lipoprotein (LDL), and triglyceride-rich lipoprotein (TRL) cholesterol levels in plasma compared with the levels in rabbits on a regular diet. No significant differences in total plasma triglyceride levels were observed. Significant increases in plasma creatinine levels were observed in rabbits fed a cholesterol-enriched diet (P < 0.05) and rabbits fed a regular diet (P < 0.05) when administered AmB. However, the magnitude of this increase was twofold greater in rabbits fed a regular diet than in rabbits fed a cholesterol-enriched diet. An increase in plasma creatinine levels was observed only in rabbits on a cholesterol-enriched diet administered ABLC. The pharmacokinetics of AmB were significantly altered in rabbits on a cholesterol-enriched diet administered Doc-AmB or ABLC compared to those in rabbits on a regular diet administered each of these compounds. The pharmacokinetics of AmB in blood were significantly different following ABLC administration but not following Doc-AmB administration in both rabbits fed cholesterol-enriched diets and rabbits fed regular diets compared to their corresponding pharmacokinetics in plasma. An increased percentage of AmB was recovered in the TRL fraction when Doc-AmB was administered to rabbits fed a cholesterol-enriched diet than when it was administered to rabbits fed a regular diet. Furthermore, an increased percentage of AmB was recovered in the LDL and TRL fractions when ABLC was administered to rabbits fed a cholesterol-enriched diet rabbits fed a regular diet. These findings suggest that an increase in plasma cholesterol levels modifies the pharmacokinetics of AmB and renal toxicity following the administration of multiple intravenous doses of Doc-AmB and ABLC. PMID- 11257035 TI - Combination treatment with intralesional cidofovir and viral-DNA vaccination cures large cottontail rabbit papillomavirus-induced papillomas and reduces recurrences. AB - We used the cottontail rabbit papillomavirus (CRPV) New Zealand White rabbit model to test a combination treatment of large established papillomas with intralesional cidofovir and DNA vaccination to cure sites and reduce recurrences. Intralesional 1% (wt/vol) (0.036 M) cidofovir treatment of rabbit papillomas led to elimination, or "cure," of the papillomas over a 6- to 8-week treatment period (N. D. Christenson, M. D. Pickel, L. R. Budgeon, and J. W. Kreider, Antivir. Res. 48:131-142, 2000). However, recurrences at periods from 1 to 8 weeks after treatment cessation were observed at approximately 50% of cured sites. DNA vaccinations with CRPV E1, E2, E6, and E7 were initiated either after or at the time of intralesional treatments, and the recurrence rates were observed. When DNA vaccinations were started after intralesional cures, recurrence rates were similar to those of vector-vaccinated rabbits. A small proportion of recurrent sites subsequently regressed (4 out of 10, or 40%) in the vaccinated group versus no regression of recurrences in the vector-immunized group (0 out of 19, or 0%), indicating partial effectiveness. In contrast, when DNA vaccinations were conducted during intralesional treatments, a significant reduction of recurrences (from 10 out of 19, or 53%, of sites in vector-immunized rabbits to 3 out of 20, or 15%, of sites in viral-DNA-immunized rabbits) was observed. DNA vaccination without intralesional treatments had a minimal effect on preexisting papillomas. These data indicated that treatment with a combination of antiviral compounds and specific immune stimulation may lead to long-term cures of lesions without the ensuing problem of papilloma recurrence. PMID- 11257036 TI - Potent anti-Trypanosoma cruzi activities of oxidosqualene cyclase inhibitors. AB - Trypanosoma cruzi is the protozoan agent that causes Chagas' disease, a major health problem in Latin America. Better drugs are needed to treat infected individuals. The sterol biosynthesis pathway is a potentially excellent target for drug therapy against T. cruzi. In this study, we investigated the antitrypanosomal activities of a series of compounds designed to inhibit a key enzyme in sterol biosynthesis, oxidosqualene cyclase. This enzyme converts 2,3 oxidosqualene to the tetracyclic product, lanosterol. The lead compound, N (4E,8E)-5,9, 13-trimethyl-4,8, 12-tetradecatrien-1-ylpyridinium, is an electron poor aromatic mimic of a monocyclized transition state or high-energy intermediate formed from oxidosqualene. This compound and 27 related compounds were tested against mammalian-stage T. cruzi, and 12 inhibited growth by 50% at concentrations below 25 nM. The lead compound was shown to cause an accumulation of oxidosqualene and decreased production of lanosterol and ergosterol, consistent with specific inhibition of the oxidosqualene cyclase. The data demonstrate potent anti-T. cruzi activity associated with inhibition of oxidosqualene cyclase. PMID- 11257037 TI - Efficacy of zanamivir against avian influenza A viruses that possess genes encoding H5N1 internal proteins and are pathogenic in mammals. AB - In 1997, an avian H5N1 influenza virus, A/Hong Kong/156/97 (A/HK/156/97), caused six deaths in Hong Kong, and in 1999, an avian H9N2 influenza virus infected two children in Hong Kong. These viruses and a third avian virus [A/Teal/HK/W312/97 (H6N1)] have six highly related genes encoding internal proteins. Additionally, A/Chicken/HK/G9/97 (H9N2) virus has PB1 and PB2 genes that are highly related to those of A/HK/156/97 (H5N1), A/Teal/HK/W312/97 (H6N1), and A/Quail/HK/G1/97 (H9N2) viruses. Because of their similarities with the H5N1 virus, these H6N1 and H9N2 viruses may have the potential for interspecies transmission. We demonstrate that these H6N1 and H9N2 viruses are pathogenic in mice but that their pathogenicities are less than that of A/HK/156/97 (H5N1). Unadapted virus replicated in lungs, but only A/HK/156/97 (H5N1) was found in the brain. After three passages (P3) in mouse lungs, the pathogenicity of the viruses increased, with both A/Teal/HK/W312/97 (H6N1) (P3) and A/Quail/HK/G1/97 (H9N2) (P3) viruses being found in the brain. The neuraminidase inhibitor zanamivir inhibited viral replication in Madin-Darby canine kidney cells in virus yield assays (50% effective concentration, 8.5 to 14.0 microM) and inhibited viral neuraminidase activity (50% inhibitory concentration, 5 to 10 nM). Twice daily intranasal administration of zanamivir (50 and 100 mg/kg of body weight) completely protected infected mice from death. At a dose of 10 mg/kg, zanamivir completely protected mice from infection with H9N2 viruses and increased the mean survival day and the number of survivors infected with H6N1 and H5N1 viruses. Zanamivir, at all doses tested, significantly reduced the virus titers in the lungs and completely blocked the spread of virus to the brain. Thus, zanamivir is efficacious in treating avian influenza viruses that can be transmitted to mammals. PMID- 11257038 TI - Anti-human immunodeficiency virus activity of YK-FH312 (a betulinic acid derivative), a novel compound blocking viral maturation. AB - Betulinic acid, a triterpenoid isolated from the methyl alcohol extract of the leaves of Syzigium claviflorum, was found to have a potent inhibitory activity against human immunodeficiency virus type 1 (HIV-1). Betulinic acid derivatives were synthesized to enhance the anti-HIV activity. Among the derivatives, 3-O (3',3'-dimethylsuccinyl) betulinic acid, designated YK-FH312, showed the highest activity against HIV-induced cytopathic effects in HIV-1-infected MT-4 cells. To determine the step(s) of HIV replication affected by YK-FH312, a syncytium formation inhibition assay in MOLT-4/HIV-1(IIIB) and MOLT-4 coculture, a multinuclear-activation-of-galactosidase-indicator (MAGI) assay in MAGI-CCR5 cells, electron microscopic observation, and a time-of-addition assay were performed. In the syncytium formation inhibition assay or in the MAGI assay for de novo infection, the compound did not show inhibitory effects against HIV replication. Conversely, no virions were detected in HIV-1-infected cell cultures treated with YK-FH312 either by electron microscopic observation or by viral yield in the supernatant. In accordance with a p24 enzyme-linked immunosorbent assay of culture supernatant in the time-of-addition assay, YK-FH312 inhibited virus expression in the supernatant when it was added 18 h postinfection. However, Western blot analysis of the cells in the time-of-addition assay revealed that the production of viral proteins in the cells was not inhibited completely by YK-FH312. These results suggest that YK-FH312 might affect the step(s) of virion assembly and/or budding of virions, and this is a novel mechanism of action of an anti-HIV compound. PMID- 11257039 TI - Antiviral activity of lovastatin against respiratory syncytial virus in vivo and in vitro. AB - Respiratory syncytial virus (RSV) is an important human pathogen that can cause severe and life-threatening respiratory infections in infants and immunocompromised adults. We have recently shown that the RSV F glycoprotein, which mediates viral fusion, binds to RhoA. One of the steps in RhoA activation involves isoprenylation at the carboxy terminus of the protein by geranylgeranyltransferase. This modification allows RhoA to be attached to phosphatidyl serine on the inner leaflet of the plasma membrane. Treatment of mice with lovastatin, a drug that inhibits prenylation pathways in the cell by directly inhibiting hydroxymethylglutaryl coenzyme A reductase, diminishes RSV but not vaccinia virus replication when administered up to 24 h after RSV infection and decreases virus-induced weight loss and illness in mice. The inhibition of replication is not likely due to the inhibition of cholesterol biosynthesis, since gemfibrozil, another cholesterol-lowering agent, did not affect virus replication and serum cholesterol levels were not significantly lowered by lovastatin within the time frame of the experiment. Lovastatin also reduces cell-to-cell fusion in cell culture and eliminates RSV replication in HEp 2 cells. These data indicate that lovastatin, more specific isoprenylation inhibitors, or other pharmacological approaches for preventing RhoA membrane localization should be considered for evaluation as a preventive antiviral therapy for selected groups of patients at high risk for severe RSV disease, such as the institutionalized elderly and bone marrow or lung transplant recipients. PMID- 11257040 TI - Multicenter survey of the changing in vitro antimicrobial susceptibilities of clinical isolates of Bacteroides fragilis group, Prevotella, Fusobacterium, Porphyromonas, and Peptostreptococcus species. AB - In vitro surveys of antimicrobial resistance among clinically important anaerobes are an important source of information that can be used for clinical decisions in the choice of empiric antimicrobial therapy. This study surveyed the susceptibilities of 556 clinical anaerobic isolates from four large medical centers using a broth microdilution method. Piperacillin-tazobactam was the only antimicrobial agent to which all the isolates were susceptible. Similarly, imipenem, meropenem, and metronidazole were highly active (resistance, <0.5%), whereas the lowest susceptibility rates were noted for penicillin G, ciprofloxacin, and clindamycin. For most antibiotics, blood isolates were less susceptible than isolates from intra-abdominal, obstetric-gynecologic, and other sources. All isolates of the Bacteroides fragilis group were susceptible to piperacillin-tazobactam and metronidazole, while resistance to imipenem and meropenem was low (<2%). For these same isolates, resistance rates (intermediate and resistant MICs) to ampicillin-sulbactam, cefoxitin, trovafloxacin, and clindamycin were 11, 8, 7, and 29%, respectively. Among the individual species of the B. fragilis group, the highest resistance rates were noted among the following organism-drug combinations: for clindamycin, Bacteroides distasonis and Bacteroides ovatus; for cefoxitin, Bacteroides thetaiotaomicron, B. distasonis, and Bacteroides uniformis; for ampicillin-sulbactam, B. distasonis, B. ovatus, and B. uniformis; and for trovafloxacin, Bacteroides vulgatus. For the carbapenens, imipenem resistance was noted among B. fragilis and meropenem resistance was seen among B. fragilis, B. vulgatus, and B. uniformis. With few exceptions all antimicrobial agents were highly active against isolates of Prevotella, Fusobacterium, Porphyromonas, and Peptostreptococcus. These data further establish and confirm that clinically important anaerobes can vary widely in their antimicrobial susceptibilities. Fortunately most antimicrobial agents were active against the test isolates. However, concern is warranted for what appears to be a significant increases in resistance to ampicillin-sulbactam and clindamycin. PMID- 11257041 TI - Activities of the combination of quinupristin-dalfopristin with rifampin in vitro and in experimental endocarditis due to Staphylococcus aureus strains with various phenotypes of resistance to macrolide-lincosamide-streptogramin antibiotics. AB - We evaluated the activities of quinupristin-dalfopristin (Q-D), alone or in combination with rifampin, against three strains of Staphylococcus aureus susceptible to rifampin (MIC, 0.06 microg/ml) and to Q-D (MICs, 0.5 to 1 microg/ml) but displaying various phenotypes of resistance to macrolide lincosamide-streptogramin antibiotics: S. aureus HM1054 was susceptible to quinupristin and dalfopristin (MICs of 8 and 4 microg/ml, respectively); for S. aureus RP13, the MIC of dalfopristin was high (MICs of quinupristin and dalfopristin for strain RP13, 8 and 32 microg/ml, respectively); and S. aureus HM1054R was obtained after conjugative transfer of macrolide-lincosamide streptogramin B constitutive resistance to HM1054, and the MIC of quinupristin for this strain was high (MICs of quinupristin and dalfopristin, 64 and 4 microg/ml, respectively). In vitro time-kill curve studies showed an additive effect [corrected] between Q-D and rifampin, at a concentration of four times the MIC, against the three strains. Rabbits with aortic endocarditis were treated 4 days with Q-D, rifampin, or their combination. In vivo, the combination was highly bactericidal and synergistic against strains susceptible to quinupristin (HM1054 and RP13) and sterilized 94% of the animals. In contrast, the combination was neither synergistic nor bactericidal against the quinupristin-resistant strain (HM1054R) and did not prevent the emergence of mutants resistant to rifampin. We conclude that the in vivo synergistic and bactericidal activity of the combination of Q-D and rifampin against S. aureus is predicted by the absence of resistance to quinupristin but not by in vitro combination studies. PMID- 11257042 TI - In70 of plasmid pAX22, a bla(VIM-1)-containing integron carrying a new aminoglycoside phosphotransferase gene cassette. AB - An Achromobacter xylosoxydans strain showing broad-spectrum resistance to beta lactams (including carbapenems) and aminoglycosides was isolated at the University Hospital of Verona (Verona, Italy). This strain was found to produce metallo-beta-lactamase activity and to harbor a 30-kb nonconjugative plasmid, named pAX22, carrying a bla(VIM-1) determinant inserted into a class 1 integron. Characterization of this integron, named In70, revealed an original array of four gene cassettes containing, respectively, the bla(VIM-1) gene and three different aminoglycoside resistance determinants, including an aacA4 allele, a new aph-like gene named aphA15, and an aadA1 allele. The aphA15 gene is the first example of an aph-like gene carried on a mobile gene cassette, and its product exhibits close similarity to the APH(3')-IIa aminoglycoside phosphotransferase encoded by Tn5 (36% amino acid identity) and to an APH(3')-IIb enzyme from Pseudomonas aeruginosa (38% amino acid identity). Expression of the cloned aphA15 gene in Escherichia coli reduced the susceptibility to kanamycin and neomycin as well as (slightly) to amikacin, netilmicin, and streptomycin. Characterization of the 5' and 3' conserved segments of In70 and of their flanking regions showed that In70 belongs to the group of class 1 integrons associated with defective transposon derivatives originating from Tn402-like elements. The structure of the 3' conserved segment indicates the closest ancestry with members of the In0-In2 lineage. In70, with its array of cassette-borne resistance genes, can mediate broad-spectrum resistance to most beta-lactams and aminoglycosides. PMID- 11257043 TI - Biochemical characterization of the FEZ-1 metallo-beta-lactamase of Legionella gormanii ATCC 33297T produced in Escherichia coli. AB - The bla(FEZ-1) gene coding for the metallo-beta-lactamase of Legionella (Fluoribacter) gormanii ATCC 33297T was overexpressed via a T7 expression system in Escherichia coli BL21(DE3)(pLysS). The product was purified to homogeneity in two steps with a yield of 53%. The FEZ-1 metallo-beta-lactamase exhibited a broad spectrum activity profile, with a preference for cephalosporins such as cephalothin, cefuroxime, and cefotaxime. Monobactams were not hydrolyzed. The beta-lactamase was inhibited by metal chelators. FEZ-1 is a monomeric enzyme with a molecular mass of 29,440 Da which possesses two zinc-binding sites. Its zinc content did not vary in the pH range of 5 to 9, but the presence of zinc ions modified the catalytic efficiency of the enzyme. A model of the FEZ-1 three dimensional structure was built. PMID- 11257044 TI - Transposons Tn1696 and Tn21 and their integrons In4 and In2 have independent origins. AB - The first 13.6 kb of the mercury and multidrug resistance transposon Tn1696, which includes the class 1 integron In4, has been sequenced. In4 is 8.33 kb long and contains the 5'-conserved segment (5'-CS) and 2.24 kb of the 3'-conserved segment (3'-CS) flanking four integrated cassettes. The 3'-CS region is followed by one full copy and an adjacent partial copy of the insertion sequence IS6100 flanked, in inverse orientation, by two short segments (123 and 152 bp) from the outer right-hand end of class 1 integrons. This structure is representative of a distinct group of class 1 integrons that differs from In2, found in Tn21, and other related class 1 integrons. In4 does not include transposition genes but is bounded by characteristic 25-bp inverted repeats and flanked by a direct duplication of 5 bp of the target sequence, indicating that it was inserted by a transpositional mechanism. In4 lies between the resII and resI sites of a backbone mercury resistance transposon which is >99.5% identical to Tn5036. Although Tn21 and Tn1696 are both classified as members of the Tn21 subfamily of the Tn3 transposon family, the backbone mercury resistance transposons are only 79 to 96% identical. Tn21 also contains a region of about 0.7 kb not found in Tn1696. The integrons In2 and In4 carrying the antibiotic resistance genes have been inserted at different locations into distinct ancestral mercury resistance transposons. Thus, Tn21 and Tn1696 have independent histories and origins. Other transposons (Tn1403 and Tn1412) that include a class 1 integron also have independent origins. In all except Tn21, the integron is located within the res region of the backbone transposon. PMID- 11257046 TI - New TEM variant (TEM-92) produced by Proteus mirabilis and Providencia stuartii isolates. AB - The sequences of the bla(TEM) genes encoding TEM-92 in Proteus mirabilis and Providencia stuartii isolates were determined and were found to be identical. Except for positions 218 (Lys-6) and 512 (Lys-104), the nucleotide sequence of bla(TEM-92) was identical to that of bla(TEM-20), including the sequence of the promoter region harboring a 135-bp deletion combined with a G-162-->T substitution. The deduced amino acid sequence of TEM-92 differed from that of TEM 52 by the presence of a substitution (Gln-6-->Lys) in the peptide signal. PMID- 11257045 TI - In vitro generation of novel pyrimethamine resistance mutations in the Toxoplasma gondii dihydrofolate reductase. AB - Pyrimethamine is a potent inhibitor of dihydrofolate reductase and is widely used in the treatment of opportunistic infections caused by the protozoan parasite Toxoplasma gondii. In order to assess the potential role of dhfr sequence polymorphisms in drug treatment failures, we examined the dhfr-ts genes of representative isolates for T. gondii virulence types I, II, and III. These strains exhibit differences in their sensitivities to pyrimethamine but no differences in predicted dhfr-ts protein sequences. To assess the potential for pyrimethamine-resistant dhfr mutants to emerge, three drug-sensitive variants of the T. gondii dhfr-ts gene (the wild-type T. gondii sequence and two mutants engineered to reflect polymorphisms observed in drug-sensitive Plasmodium falciparum) were subjected to random mutagenesis and transfected into either wild type T. gondii parasites or dhfr-deficient Saccharomyces cerevisiae under pyrimethamine selection. Three resistance mutations were identified, at amino acid residues 25 (Trp-->Arg), 98 (Leu-->Ser), and 134 (Leu-->His). PMID- 11257047 TI - In vitro and in vivo combinations of cefotaxime and minocycline against Aeromonas hydrophila. AB - The activities of cefotaxime and minocycline against Aeromonas hydrophila were investigated. Cefotaxime (4 times the MIC) plus minocycline (0.75 times the MIC) elicited an inhibitory effect for 48 h in a time-kill study, and more infected mice treated with both drugs survived (91%) than survived after treatment with cefotaxime (9%) or minocycline (44%) alone, suggesting that cefotaxime and minocycline act synergistically against A. hydrophila. PMID- 11257048 TI - Hypersusceptibility of the Pseudomonas aeruginosa nfxB mutant to beta-lactams due to reduced expression of the ampC beta-lactamase. AB - The Pseudomonas aeruginosa nfxB mutant lacking mexAB-oprM showed hypersusceptibility to 9 out of 24 beta-lactams tested. This hypersusceptibility was found for the nfxB mutant lacking mexAB-oprM-mexXY (N108) but not for the nfxB mutant lacking both mexAB-oprM-mexXY and ampC. The level of the AmpC beta lactamase induction was reduced in N108. Thus, the reduced AmpC induction must be the cause of the hypersusceptibility. PMID- 11257049 TI - In vitro and in vivo activities of amikacin, cefepime, amikacin plus cefepime, and imipenem against an SHV-5 extended-spectrum beta-lactamase-producing Klebsiella pneumoniae strain. AB - The in vitro and in vivo effectiveness of amikacin, cefepime, and imipenem was studied using a high inoculum of an extended-spectrum beta-lactamase-producing Klebsiella pneumoniae strain. An in vitro susceptibility test at the standard inoculum predicted the in vivo outcome of amikacin or imipenem while it did not do so for cefepime due to the inoculum effect. PMID- 11257050 TI - Combination effect of vancomycin and beta-lactams against a Staphylococcus aureus strain, Mu3, with heterogeneous resistance to vancomycin. AB - We tested the combined activity of vancomycin and seven beta-lactam antibiotics against Staphylococcus aureus clinical strain Mu3, which displays heterogeneous resistance to vancomycin. When combined with vancomycin, four of the seven tested beta-lactams exhibited an additive effect at or near their MICs, while all showed an antagonistic effect at lower, sub-MIC levels. This study implicated the unpredictable nature of combination therapy of beta-lactams and vancomycin against S. aureus with reduced susceptibility to vancomycin. PMID- 11257051 TI - In vitro activities of linezolid against multiple Nocardia species. AB - Linezolid was tested by broth microdilution against 140 clinical Nocardia isolates belonging to seven species. The MIC at which 50% of the strains are inhibited (MIC50) and MIC90 for all species other than Nocardia farcinica were 2 and 4 microg/ml. Linezolid is the first antimicrobial agent demonstrated to be active against all Nocardia species. PMID- 11257052 TI - Bactericidal activities of milk lipids. AB - The bactericidal capacity of digestion products of bovine milk triglycerides and membrane lipids was tested in vitro using Escherichia coli O157:H7, Salmonella enteritidis, Campylobacter jejuni, Listeria monocytogenes, and Clostridium perfringens. C10:0 and C12:0 fatty acids and digestion products of sphingolipids appeared to be effective bactericidal agents, whereas digestion products of phosphoglycerides were moderately bactericidal. Thus, milk fat sphingolipids and triglycerides, particularly those containing C10:0 and C12:0 fatty acids, may protect against food-borne gastroenteritis. PMID- 11257053 TI - Inhibition of isoniazid-induced expression of Mycobacterium tuberculosis antigen 85 in sputum: potential surrogate marker in tuberculosis chemotherapy trials. AB - Mycobacterium tuberculosis antigen 85 is induced in vitro by isoniazid (INH); its sustained induction in sputum during tuberculosis (TB) therapy predicts relapse. In this trial, rifampin or rifalazil inhibited the induction of sputum antigen 85 by INH in a dose-dependent fashion. This approach may facilitate the evaluation of new TB drugs. PMID- 11257054 TI - Multidrug resistance is mediated by large plasmids carrying a class 1 integron in the emergent Salmonella enterica serotype [4,5,12:i:-]. AB - A multidrug-resistant Salmonella enterica serotype [4,5,12:i:-] clone carried a class 1 integron harboring dfrA12 and aadA2 gene cassettes and bla(TEM-1), aac(3) IV, cmlA1, and tetA genes located in large plasmids of about 140 kb (carrying spv) or 120 kb (lacking spv). Several segregants, lacking multidrug resistance, contained a plasmid smaller than the parental one and no longer hybridized with probes for the lost resistances. The genes mediating resistance to ampicillin, chloramphenicol, and tetracycline in the [4,5,12:i:-] clone are different from those found in the pentadrug-resistant serotype Typhimurium DT104 clone. PMID- 11257055 TI - Diversity of Tn1546 elements in clinical isolates of glycopeptide-resistant enterococci from Scottish hospitals. AB - The Tn1546-related elements of 48 Van glycopetide-resistant enterococci were compared. Ten distinct Tn1546 types were identified with variation primarily due to IS1542 and IS1216V-like insertions. Clonal isolates frequently differed in their Tn1546 type, indicating instability of Tn1546-related elements. A putative hybrid promoter was identified, generated upstream of vanR by the insertion of IS1542. The presence of this hybrid promoter was associated with constitutive expression of the van genes and elevated teicoplanin resistance. PMID- 11257056 TI - Blocking the receptor for interleukin 10 protects mice from lethal listeriosis. AB - High doses of Listeria monocytogenes overcome the ability of a normal mouse to control the infection, due to massive bacterial replication. Treatment with an anti-interleukin 10 (IL-10) receptor monoclonal antibody prevented the fatal course of infection with high doses of bacteria. This work shows that blocking the receptor for IL-10 may have useful therapeutic applications. PMID- 11257057 TI - Latex sensitization in health care workers and in the US general population. AB - Sensitization to natural rubber latex is a prerequisite to type I immediate hypersensitivity reactions (urticaria, angioedema, anaphylaxis, and allergic rhinitis) that result from subsequent latex exposure. This study examines occupations in which latex glove use is common to determine whether it is associated with increased prevalence odds of latex sensitization (measured by latex-specific immunoglobulin E antibodies) by using data from 5,512 adults aged 17--60 years from the Third National Health and Nutrition Examination Survey (1988--1991). After other factors associated with latex sensitization were controlled for, there was a nonsignificant association between longest-held jobs in health care and latex sensitization (odds ratio (OR) = 1.49, 95 percent confidence interval (CI): 0.92, 2.40). For current occupations, latex sensitization was not associated with health care work in which gloves were used (OR = 1.17, 95 percent CI: 0.51, 2.65) or with other occupations in which latex glove use is common (OR = 1.01, 95 percent CI: 0.49, 2.07) compared with other occupations. Current health care workers who reported not using gloves were at increased risk of latex sensitization, both among those without a history of childhood atopy (OR = 2.30, 95 percent CI: 1.04, 5.13) and those with such a history (OR = 28.04, 95 percent CI: 3.64, 215.97). This odds ratio heterogeneity suggests that subjects with childhood atopy may be at high risk of latex sensitization. PMID- 11257058 TI - Invited commentary: assessing latex sensitization using data from NHANES III. PMID- 11257060 TI - Association of transposition of the great arteries in infants with maternal exposures to herbicides and rodenticides. AB - The Baltimore-Washington Infant Study, a case-control study of congenital heart defects in liveborn infants conducted in 1981--1989, interviewed parents about a wide range of environmental exposures that occurred during and before the pregnancy. In the period 1987--1989, the questionnaire was expanded to include a detailed inquiry about exposures to pesticides. An analysis of these latter data revealed an association of maternal exposure to any pesticides during the first trimester with transposition of the great arteries in their infants (TGA; n = 66 infants), relative to 771 control infants, with an odds ratio of 2.0 (95% confidence interval (CI): 1.2, 3.3). No other heart defects were associated with pesticides. When analyzed by type of pesticide and adjusted for covariates, there were associations of TGA with maternal exposures to herbicides (odds ratio (OR) = 2.8; 95% CI: 1.3, 7.2) and to rodenticidal chemicals (OR = 4.7; 95% CI: 1.4, 12.1) but not to insecticides (OR = 1.5; 95% CI: 0.9, 2.6). No data were collected on specific chemicals or brand names. These results raise new questions about the possible epidemiologic association of TGA with some classes of pesticides and warrant new, carefully targeted investigations. PMID- 11257061 TI - Relation of alleles of the sodium-potassium adenosine triphosphatase alpha 2 gene with blood pressure and lead exposure. AB - Lead is associated with elevated blood pressure, although the mechanism of action is unknown. Genetic differences in sodium-potassium adenosine triphosphatase (Na(+)-K(+)ATPase) could explain some of the variation in the strength of the blood pressure-blood lead relation that has been observed in previous studies. In 1996-1997, the authors studied the association of blood pressure, hypertension prevalence, and polymorphisms in the gene for the alpha 2 subunit of Na(+) K(+)ATPase (ATP1A2) among 220 former organolead manufacturing workers from New Jersey. Subjects were genotyped for a restriction fragment length polymorphism (RFLP) on the ATP1A2 gene. The association between blood lead and blood pressure was stronger among persons who were homozygous for the variant allele. Genotype was also associated with hypertension (adjusted odds ratio = 7.7; 95% confidence interval: 1.9, 31.4). Finally, the variant allele was 1.8 times more common among African Americans than among Caucasians. The RFLP may indicate susceptibility to the effect of lead on blood pressure. Moreover, the alpha 2 gene (or a closely linked gene) may contribute to the pathophysiology of hypertension. However, because the number of subjects (especially African Americans) with the susceptible genotype in this study was small, these observations should be considered preliminary. PMID- 11257062 TI - Analysis of 13 (32)P-DNA postlabeling studies on occupational cohorts exposed to air pollution. AB - Industrial and urban workers may be exposed to significant levels of air pollutants resulting from the incomplete combustion of organic matter. The authors performed a meta-analysis of 13 DNA-adduct studies ((32)P-DNA postlabeling technique) on occupational cohorts exposed to air pollution. The association between levels of DNA adducts and air pollution exposure was significant both in heavily exposed industrial workers and in less severely exposed urban workers. Moreover, in an analysis using the seven studies that reported measuring levels of benzo[a]pyrene (B(a)P), a typical marker of exposure, DNA adduct levels in exposed workers (versus those in referents) were significantly correlated with air levels of B(a)P. The relation between DNA adducts and B(a)P was found to be linear at low doses and sublinear at high doses, indicating that DNA adduct formation tends to reach some kind of saturation point at higher levels of exposure to the chemical mixtures present in fumes. When the authors examined the efficiency of DNA adduct production associated with increasing air pollution exposures, the production of DNA adducts per unit of exposure was significantly decreased at higher B(a)P exposure levels. These findings suggest that linear downward extrapolations based on DNA adduct levels associated with B(a)P concentrations of > or =20 ng/m(3) might be affected by underestimation bias. PMID- 11257063 TI - Skin cancer risk in relation to toenail arsenic concentrations in a US population based case-control study. AB - Arsenic is a known carcinogen specifically linked to skin cancer occurrence in regions with highly contaminated drinking water or in individuals who took arsenic-containing medicines. Presently, it is unknown whether such effects occur at environmental levels found in the United States. To address this question, the authors used data collected on 587 basal cell and 284 squamous cell skin cancer cases and 524 controls interviewed as part of a case-control study conducted in New Hampshire between 1993 and 1996. Arsenic was determined in toenail clippings using instrumental neutron activation analysis. The odds ratios for squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) were close to unity in all but the highest category. Among individuals with toenail arsenic concentrations above the 97th percentile, the adjusted odds ratios were 2.07 (95% confidence interval (CI): 0.92, 4.66) for SCC and 1.44 (95% CI: 0.74, 2.81) for BCC, compared with those with concentrations at or below the median. While the risks of SCC and BCC did not appear elevated at the toenail arsenic concentrations detected in most study subjects, the authors cannot exclude the possibility of a dose-related increase at the highest levels of exposure experienced in the New Hampshire population. PMID- 11257064 TI - Association of perforation of the appendix with female tubal infertility. AB - Although perforation of the appendix is considered a risk factor for female tubal infertility, the epidemiologic evidence supporting this relation is inconsistent. Risk factors for tubal infertility were compared for 121 women with documented primary tubal infertility attending in vitro fertilization clinics in Toronto, Canada, from July to December 1998 and 490 controls who were pregnant during the same time period. Self-administered questionnaires and review of medical records were used to assess exposures. The authors found that neither history of acute appendicitis nor perforation of the appendix was a statistically significant risk factor for tubal infertility. The crude odds ratio for perforated appendicitis was 3.4 (95% confidence interval (CI): 0.9, 12.9), and the adjusted odds ratio was 1.4 (95% CI: 0.3, 6.2). In addition to increased age and annual income, cigarette smoking (odds ratio (OR) = 2.0, 95% CI: 1.2, 3.2), history of endometriosis (OR = 6.0, 95% CI: 2.8,12.8), and history of pelvic inflammatory disease (OR = 6.0, 95% CI: 2.8, 12.8) were significantly associated with tubal infertility in multivariate analysis. These data do not provide substantial evidence that perforation of the appendix is an important risk factor for female tubal infertility. PMID- 11257065 TI - Plasma concentrations of carotenoids, retinol, and tocopherols in preeclamptic and normotensive pregnant women. AB - This case-control study was conducted in Lima, Peru, from June 1997 through January 1998 to assess whether plasma concentrations of carotenoids (alpha carotene, beta-carotene, lycopene, lutein, zeaxanthin, beta-cryptoxanthin), retinol, and tocopherols (alpha-tocopherol and gamma-tocopherol) are decreased in women with preeclampsia. A total of 125 pregnant women with preeclampsia and 179 normotensive pregnant women were included. Plasma concentrations of antioxidants were determined using high performance liquid chromatography. After adjusting for maternal demographic, behavioral, and reproductive characteristics and total plasma lipid concentrations, the authors found a linear increase in risk of preeclampsia with increasing concentrations of alpha-tocopherol (odds ratio of the highest quartile = 3.13; 95% confidence interval: 1.06, 9.23, with the lowest quartile as the reference group; p value of the test of linear trend = 0.040). The risk of preeclampsia decreased across increasing quartiles of concentrations for retinol (odds ratio of the highest quartile = 0.32; 95% confidence interval: 0.15, 0.69, with the lowest quartile as the reference group; p value of the test of linear trend = 0.001). Some of these results are inconsistent with the prevailing hypothesis that preeclampsia is an antioxidant-deficient state. Preliminary findings confirm an earlier observation of increased plasma concentrations of alpha-tocopherol among women with preeclampsia as compared with normotensive pregnant women. PMID- 11257066 TI - Life-course predictors of ultrasonic heel measurement in a cross-sectional study of immigrant women from Southeast Asia. AB - Few studies address chronic disease risk for Southeast Asians in the United States. In 1999, the authors conducted a cross-sectional study of bone mineral density (BMD) estimated from ultrasonic calcaneal measurements in women born in Southeast Asia who then lived in Chicago, Illinois. The study addressed three questions: Do Southeast-Asian women have relatively low BMD? What factors before and after immigration are associated with BMD? Are factors that reflect the childhood/adolescent environment equally associated with BMD for postmenopausal and premenopausal women? An interviewer-administered bilingual questionnaire collected immigration, reproductive, and lifestyle data from 213 women (aged 20- 80 years) born in Vietnam, Cambodia, or Laos. The authors found that the estimated mean BMD of postmenopausal Southeast-Asian women was lower than the reference values for White women. Four summary indicators of childhood/adolescent environment were predictive of higher BMD: more years of education, earlier age at menarche, lower height, and coastal birth; these indicators were more strongly associated with BMD for premenopausal (multiple-partial R(2) = 0.21) than postmenopausal (R(2) = 0.06) women. Young-adult exposures (e.g., early first pregnancy and age at immigration) and proximal lifestyle factors (e.g., smoking, physical inactivity, vegetarian diet, and betel nut use) were also assessed as potential predictors of BMD. PMID- 11257068 TI - Cross-sectional and prospective study of exercise and depressed mood in the elderly : the Rancho Bernardo study. AB - This study examined cross-sectional and prospective associations of exercise with depressed mood in a community-based sample of older men and women (aged 50--89 years in 1984--1987) in southern California. Regular strenuous exercise and exercise > or =3 times per week were reported; depressed mood was assessed by using the Beck Depression Inventory (BDI). After exclusion of persons with categorical depression and those rating themselves largely or extremely physically limited during the previous month, data on 932 men and 1,097 women were available for cross-sectional analysis. Exercise and depressed mood were reassessed for 404 men and 540 women in 1992--1995; these data were the focus of prospective analyses. In 1984--1987, exercise rates were high (>80%), and average BDI scores were low. Cross-sectional analyses indicated that before and after adjustment for covariates, exercise was significantly associated with less depressed mood. However, prospective analyses of the 944 persons who attended both clinic visits indicated no association between baseline exercise and either follow-up BDI score (p > 0.10) or change in BDI score between baseline and follow up (p > 0.10). Results confirm that exercisers have less depressed mood. However, exercise does not protect against future depressed mood for those not clinically depressed at baseline. PMID- 11257067 TI - Risk factors for proximal humerus, forearm, and wrist fractures in elderly men and women: the Dubbo Osteoporosis Epidemiology Study. AB - Fractures of the proximal humerus, forearm, and wrist account for approximately one third of total osteoporotic fractures in the elderly. Several risk factors for these fractures were evaluated in this prospective study of 739 men and 1,105 women aged > or =60 years in Dubbo, Australia. During follow-up (1989-1996), the respective incidences of humerus and of forearm and wrist fractures, per 10,000 person-years, were 22.6 and 33.8 for men and 54.8 and 124.6 for women. Independent predictors of humerus fracture were femoral neck bone mineral density (FNBMD) (relative risk (RR) = 2.3, 95% confidence interval (CI): 1.2, 4.5) in men and FNBMD (RR = 2.4, 95% CI: 1.7, 3.5) and height loss (RR = 1.1, 95% CI: 1.0, 1.2) in women. For forearm and wrist fractures, risk factors were FNBMD (men: RR = 1.5, 95% CI: 1.0, 2.3; women: RR = 1.5, 95% CI: 1.2, 1.9) and height loss (men: RR = 1.2, 95% CI: 1.0, 1.3; women: RR = 1.1, 95% CI: 1.0, 1.2). In addition, dietary calcium (men: RR = 2.0, 95% CI: 1.0, 3.6) and a history of falls (women: RR = 1.9, 95% CI: 1.4, 2.6) were also significant. These data suggest that elderly men and women largely share common risk factors for upper limb fractures and that FNBMD is the primary risk factor. PMID- 11257069 TI - Multiple chemical sensitivity and chronic fatigue syndrome in British Gulf War veterans. AB - The objective of this study was to measure the prevalence of multiple chemical sensitivity (MCS) and chronic fatigue syndrome (CFS) in British Gulf War veterans and to investigate their association with reported exposures and psychologic morbidity. In 1997--1998, the authors undertook a cross-sectional survey of three cohorts of British military personnel comprising Gulf veterans (n = 3,531), those who had served in Bosnia (n = 2,050), and those serving during the Gulf War but not deployed there (Era cohort, n = 2,614). MCS and CFS were defined according to operational criteria. The prevalence of MCS in the Gulf, Bosnia, and Era cohorts was 1.3%, 0.3%, and 0.2%, respectively. For CFS, the prevalence was 2.1% (Gulf cohort), 0.7% (Bosnia cohort), and 1.8% (Era cohort). In Gulf veterans, MCS was strongly associated with exposure to pesticides (adjusted odds ratio = 12.3, 95% confidence interval: 5.1, 30.0). Both syndromes were associated with high levels of psychologic morbidity. These findings suggest that CFS and MCS account for some of the medically unexplained illnesses reported by veterans after deployment to the Gulf. MCS was particularly associated with Gulf deployment and self reported exposure to pesticides, findings that merit further exploration given the controversial status of this diagnosis and the potential for recall bias in a questionnaire survey. PMID- 11257070 TI - Incidence of Guillain-Barre syndrome following infection with Campylobacter jejuni. AB - Evidence of recent or ongoing Campylobacter jejuni infection has been found in approximately one out of every four cases of Guillain-Barre syndrome (GBS). It is increasingly accepted that C. jejuni infection is an important causal factor for GBS. However, the likelihood of GBS' occurring following an episode of C. jejuni gastroenteritis has not been measured. The authors measured the incidence of GBS in a large cohort of persons with laboratory-confirmed C. jejuni infection. Cases of C. jejuni infection were derived from the Swedish national laboratory reporting system for the years 1987--1995. Follow-up for GBS was carried out using the Swedish national hospital inpatient register. Nine cases of GBS were detected in the cohort, which comprised 29,563 cases of C. jejuni infection--a rate of 30.4 per 100,000 (95% confidence interval: 13.9, 57.8). This compares with an expected incidence of 0.3 per 100,000 in a 2-month period in the general population. GBS is an important but rare complication of C. jejuni infection. The risk of developing GBS during the 2 months following a symptomatic episode of C. jejuni infection is approximately 100 times higher than the risk in the general population. PMID- 11257071 TI - Re: "Are children living near high-voltage power lines at increased risk of acute lymphoblastic leukemia?". PMID- 11257072 TI - Re: "Use of two-segmented logistic regression to estimate change-points in epidemiologic studies". PMID- 11257073 TI - Testing platelet activation with a shear-dependent platelet function test versus aggregation-based tests: relevance for monitoring long-term glycoprotein IIb/IIIa inhibition. AB - BACKGROUND: Tests developed to monitor glycoprotein (GP) IIb/IIIa blockade do not properly reflect platelet function in vivo and need a baseline (pretreatment) value. Because GP IIb/IIIa is essential in platelet aggregation and thrombosis under shear conditions, a flow-dependent approach to monitor its inhibition can be used. METHODS AND RESULTS: We compared a test based on flow-dependent platelet deposition, the Cone and Platelet Analyzer (CPA), with in vitro platelet aggregometry and the Rapid Platelet Function Assay (RPFA) on platelet function after GP IIb/IIIa inhibition. In vitro, increasing concentrations of abciximab (0% to 100% receptor occupancy) were tested. Ex vivo, platelet function was monitored with the CPA and with aggregometry for up to 1 week after abciximab administration. The CPA was better correlated with the percentage of free GP IIb/IIIa receptors than was aggregometry or the RPFA. Only the RPFA, when expressed as a ratio over baseline (pretreatment), was comparable to the CPA. Ex vivo, the CPA, but not aggregometry, showed prolonged platelet inhibition with gradual recovery from GP IIb/IIIa receptor blockade in the first week after abciximab administration. CONCLUSIONS: Platelet function assessment by shear induced deposition is a reliable test to monitor a wide range of GP IIb/IIIa inhibition. Its accuracy does not require a baseline reference. The effects of GP IIb/IIIa blockade on platelet function should be examined under high shear conditions. PMID- 11257074 TI - Regression of left ventricular hypertrophy after nonsurgical septal reduction therapy for hypertrophic obstructive cardiomyopathy. AB - BACKGROUND: Hypertrophic obstructive cardiomyopathy (HOCM) is characterized by left ventricular hypertrophy (LVH) in the absence of increased external load. Recently, nonsurgical septal reduction therapy (NSRT) with intracoronary ethanol has been introduced to treat severely symptomatic patients with outflow tract obstruction. Its long-term effects on LV mass, however, are unknown. METHODS AND RESULTS: The LV size, function, and outflow tract gradient of 26 HOCM patients (53+/-15 years old) who underwent NSRT were assessed by echocardiography at baseline and 1 and 2 years after the procedure. LVH was evaluated by wall thickness of individual myocardial segments, planimetered myocardial area, and mass. The outflow gradient decreased from 36+/-6 mm Hg before NSRT to 0+/-3 mm Hg at 2 years (P<0.001), with patients experiencing symptomatic improvement (P<0.05). LV end-diastolic and end-systolic dimensions increased significantly at both 1 and 2 years (P<0.001). All parameters of LVH showed evidence of regression. LV mass decreased (301+/-78 g at baseline, 223+/-5 g at 1 year, and 190+/-58 g at 2 years; P<0.01), with the 2-year reduction in mass related to infarct size and the acute reduction in outflow tract gradient (r=0.48, P<0.05 and r=0.63, P<0.01, respectively). CONCLUSIONS: NSRT results in LV remodeling that is characterized by an increase in LV size and a decrease in the extent of LVH. PMID- 11257075 TI - Tamoxifen effects on endothelial function and cardiovascular risk factors in men with advanced atherosclerosis. AB - BACKGROUND: Tamoxifen and its analogues act as selective estrogen receptor modulators (SERMs) in women, with estrogen-like activities on some plasma cardiovascular risk factors (eg, lipoproteins). Effects of SERMs on men with coronary artery disease (CAD) have not been reported. METHODS AND RESULTS: Thirty one men with angiographically proven CAD were recruited; 16 were treated with tamoxifen (40 mg/d) for 56 days, and 15 were untreated. All the CAD patients were medicated with aspirin and an HMG-CoA reductase inhibitor for >/=6 weeks before entering the study. Ten men with angina-like symptoms but normal coronary arteries by angiography (NCA group) were also treated with tamoxifen. Blood samples were collected at days -7, 0, 7, 14, 21, 28, and 56 of treatment. Endothelium-dependent flow-mediated dilatation (ED-FMD) of the brachial artery was measured by high-resolution ultrasound at 5 visits. Tamoxifen caused an increase in %ED-FMD maximal at 28 days in the CAD group (2.1+/-0.3% to 7.5+/ 0.7%; P<0.0001) and the NCA group (3.8+/-0.4% to 7.9+/-1.0%; P<0.0001), with no significant change in the untreated group. Tamoxifen also caused decreases in several plasma cardiovascular risk factors, including total cholesterol, triglycerides, lipoprotein(a), and fibrinogen. Except for the triglyceride response, these effects were similar to those reported for postmenopausal women treated with tamoxifen. CONCLUSIONS: Tamoxifen substantially increased ED-FMD in men with CAD who were taking conventional medication. Together with the effects on risk factors, the data strongly support clinical evaluation of SERMs for the treatment of men with CAD. PMID- 11257076 TI - Independent and joint effects of antibodies to human heat-shock protein 60 and Chlamydia pneumoniae infection in the development of coronary atherosclerosis. AB - BACKGROUND: Studies have suggested that the prevalence of antibodies against heat shock proteins (HSPs), Chlamydia pneumoniae (CPN), and cytomegalovirus (CMV) is associated with coronary artery disease (CAD), but the independent or joint effects of human (h) HSP60 antibodies and these pathogens in patients have not been fully elucidated. METHODS AND RESULTS: A total of 405 subjects (276 patients with CAD and 129 control individuals) were tested for serum antibodies to hHSP60, CPN, and CMV immediate-early-1 (IE1) antigens. Patients were also assessed for serum cholesterol, triglyceride levels, and smoking habit. Significantly elevated levels of antibodies to hHSP60 and CPN but not to CMV-IE1 antigens were documented in CAD patients. Multiple logistic regression analysis and subanalyses of selected subjects showed that these associations were independent of age, sex, smoking, and serum lipid levels. Antibodies to hHSP60 and CPN did not correlate quantitatively; however, the relative risk of disease development was substantially increased in subjects with high antibody levels to both hHSP60 and CPN:, reaching an odds ratio of 82.0 (95% CI 10.6 to 625.0). CONCLUSIONS: High levels of antibodies to hHSP60 and CPN: are independent risk factors for coronary atherosclerosis, but their simultaneous presence substantially increases the risk for disease development. PMID- 11257077 TI - Molecular fingerprint of interferon-gamma signaling in unstable angina. AB - BACKGROUND: Activation of circulating monocytes in patients with acute coronary syndromes may reflect exposure to bacterial products or stimulation by cytokines such as IFN-gamma. IFN-gamma induces phosphorylation and nuclear translocation of transcription factor STAT-1, which initiates a specific program of gene induction. To explore whether monocyte activation is IFN-gamma driven, patients with unstable (UA) or stable angina (SA) were compared for nuclear translocation of STAT-1 complexes and upregulation of IFN-gamma-inducible genes CD64 and IP-10. METHODS AND RESULTS: Peripheral blood mononuclear cells were stained for expression of CD64 on CD14(+) monocytes and analyzed by PCR for transcription of IP-10. Expression of CD64 was significantly increased in patients with UA. Monocytes from UA patients remained responsive to IFN-gamma in vitro, with accelerated transcriptional competency of CD64. IP-10-specific sequences were spontaneously detectable in 82% of the UA patients and 15% of SA patients (P<0.001). Most importantly, STAT-1 complexes were found in nuclear extracts prepared from freshly isolated monocytes of patients with UA, which provides compelling evidence for IFN-gamma signaling in vivo. CONCLUSIONS: Monocytes from UA patients exhibit a molecular fingerprint of recent IFN-gamma triggering, such as nuclear translocation of STAT-1 complexes and upregulation of IFN-gamma inducible genes CD64 and IP-10, which suggests that monocytes are activated, at least in part, by IFN-gamma. IFN-gamma may derive from stimulated T lymphocytes, which implicates specific immune responses in the pathogenesis of acute coronary syndromes. PMID- 11257078 TI - Human tissue valves in aortic position: determinants of reoperation and valve regurgitation. AB - BACKGROUND: Human tissue valves for aortic valve replacement have a limited durability that is influenced by interrelated determinants. Hierarchical linear modeling was used to analyze the relation between these determinants of durability and valve regurgitation measured by serial echocardiography. METHODS AND RESULTS: In adult patients, 218 cryopreserved aortic allografts were implanted with the subcoronary (85) or the root replacement technique (133), and 81 patients had root replacement with a pulmonary autograft. Mean follow-up was 4.2 years (SD 2.7; range, 0 to 10.5). Patient age, operator experience with subcoronary implantation, and allograft diameter were independent predictors for reoperation. With repeated color Doppler echocardiography, the severity of aortic regurgitation was assessed by the jet length method and the jet diameter ratio. Multilevel hierarchical linear modeling was used to estimate initial aortic regurgitation (intercept), its change over time (slope), and the effect of 11 potential determinants of durability on aortic regurgitation. With the jet length method, the intercept was 0.94 grade and the slope was 0.11 grade per year. With the jet diameter ratio, the intercept was 0.34 and the annual increase was 0.01. Subcoronary implanted valves had more initial aortic regurgitation, but progression of aortic valve regurgitation did not differ from root replacement. At midterm follow-up, recipient age <40 years was the only independent predictor of aortic regurgitation. CONCLUSIONS: Subcoronary implantation has a learning curve, resulting in more initial aortic regurgitation and early reoperation compared with root replacement. In both techniques, progression of aortic regurgitation over time is small but accelerated in young adults. PMID- 11257080 TI - Abdominal aortic calcific deposits are an important predictor of vascular morbidity and mortality. AB - BACKGROUND: The impact of abdominal arterial calcific deposits on the prediction of cardiovascular disease (CVD) over a long follow-up interval deserves greater scrutiny. METHODS AND RESULTS: Lateral lumbar radiographs were studied as a predictor of incident coronary heart disease (CHD), CVD, and CVD mortality in 1049 men and 1466 women (mean age, 61 years) who were followed from 1967 to 1989. Anterior and posterior wall calcific deposits in the aorta at the level of the first through fourth lumbar vertebrae were graded according to increasing severity using a previously validated rating scale for abdominal aortic calcium (AAC) that ranges from 0 to 24 points. There were 454 cases of CHD, 709 cases of CVD, and 365 CVD deaths. Proportional hazards logistic regression was used to test for associations between AAC and later events after adjustment for age, cigarette use, diabetes mellitus, systolic blood pressure, left ventricular hypertrophy, body mass index, cholesterol, and HDL cholesterol. In comparisons with the lowest AAC tertile, the multivariate age-adjusted relative risks (RR) for CVD were increased in tertile 2 (men: RR, 1.33; 95% confidence interval [CI], 1.02 to 1.74; women: RR, 1.25; 95% CI, 0.95 to 1.65) and tertile 3 (men: RR, 1.68; 95% CI, 1.25 to 2.27; women: RR, 1.78; 95% CI, 1.33 to 2.38). Similar results were obtained with CHD and CVD mortality. CONCLUSIONS: AAC deposits, detected by lateral lumbar radiograms, are a marker of subclinical atherosclerotic disease and an independent predictor of subsequent vascular morbidity and mortality. PMID- 11257079 TI - Bone formation and inflammation in cardiac valves. AB - BACKGROUND: For nearly a century, the mechanical failure of calcified heart valves was attributed to a passive degenerative process. Recently, several case reports described bone formation in surgically excised heart valves and suggested an unexpected process of tissue repair. METHODS AND RESULTS: We studied the prevalence and pathology of heterotopic ossification in 347 surgically excised heart valves (256 aortic, 91 mitral) in 324 consecutive patients (182 men, 142 women; mean age 68 years) who underwent cardiac valve replacement surgery between 1994 and 1998. The valves were examined microscopically to determine the prevalence and features of bone formation and remodeling. Two hundred eighty eight valves (83%) had dystrophic calcification. Mature lamellar bone with hematopoietic elements and active bone remodeling were present in 36 valves (13%) with dystrophic calcification. Endochondral bone formation, similar to that seen in normal fracture repair, was identified in 4 valves. Microfractures were present in 92% of all valves with ossification. Neoangiogenesis was found in all valves with ossification. Bone morphogenetic proteins 2 and 4 (BMP 2/4), potent osteogenic morphogens, were expressed by myofibroblasts and preosteoblasts in areas adjacent to B- and T-lymphocyte infiltration in valves where ossification was identified. Mast cells were present in calcified and ossified valves and were especially prominent in atheromatous regions. CONCLUSIONS: Heterotopic ossification consisting of mature lamellar bone formation and active bone remodeling is a relatively common and unexpected finding in end-stage valvular heart disease and may be associated with repair of pathological microfractures in calcified cardiac valves. PMID- 11257081 TI - Prognosis after aortic valve replacement with a bioprosthesis: predictions based on meta-analysis and microsimulation. AB - BACKGROUND: Bioprostheses are widely used as an aortic valve substitute, but knowledge about prognosis is still incomplete. The purpose of this study was to provide insight into the age-related life expectancy and actual risks of reoperation and valve-related events of patients after aortic valve replacement with a porcine bioprosthesis. METHODS AND RESULTS: We conducted a meta-analysis of 9 selected reports on stented porcine bioprostheses, including 5837 patients with a total follow-up of 31 874 patient-years. The annual rates of valve thrombosis, thromboembolism, hemorrhage, and nonstructural dysfunction were 0.03%, 0.87%, 0.38%, and 0.38%, respectively. The annual rate of endocarditis was estimated at 0.68% for >6 months of implantation and was 5 times as high during the first 6 months. Structural valve deterioration was described with a Weibull model that incorporated lower risks for older patients. These estimates were used to parameterize, calibrate, and validate a mathematical microsimulation model. The model was used to predict life expectancy and actual risks of reoperation and valve-related events after implantation for patients of different ages. For a 65 year-old male, these figures were 11.3 years, 28%, and 47%, respectively. CONCLUSIONS: The combination of meta-analysis with microsimulation enabled a detailed insight into the prognosis after aortic valve replacement with a bioprosthesis for patients of different ages. This information will be useful for patient counseling and clinical decision making. It also could serve as a baseline for the evaluation of newer valve types. PMID- 11257082 TI - Isolated mitral valve replacement with St. Jude medical prosthesis: long-term results: a follow-up of 19 years. AB - BACKGROUND: In this retrospective study, approximately 440 patients received mitral valve replacements with the St Jude Medical prosthesis. The last patient was operated on 10 years before the beginning of the follow-up. The extended follow-up was 19 years. METHODS AND RESULTS: Four hundred forty patients (sex ratio, 1.32 [men to women]; age, 60+/-11.4 years; age range, 7 to 75 years) were operated on from 1979 to 1987. All patients underwent isolated mitral valve replacement. Tricuspid plasty was the only associated procedure. The follow-up at 19 years was 98% complete. The overall actuarial survival rate was 63+/-3.3% at 19 years, and the actuarial survival rate (only valve related) was 83+/-2.7%. The operative mortality rate (0 to 30 days) was 4.09%. We found that 89.4% of the patients alive at 19 years were in NYHA class I/II. Multivariate analysis showed that age and sex were significantly correlated with valve-related mortality and that age, sex, NYHA class, and atrial fibrillation were significantly correlated with overall mortality. The linearized rates (percent patient-years) of thromboembolism, thrombosis, and hemorrhage were 0.69, 0.2, and 1, respectively. At 19 years, freedom from endocarditis and reoperation was 98.6+/-1% and 90+/-3%, respectively. CONCLUSIONS: In this study, the very-long-term results confirm the excellent durability of the St Jude Medical prosthesis in the mitral position and show the difficulty of adjusting the anticoagulation protocol, even after long term treatment. PMID- 11257084 TI - Age-dependent and hypoxia-related differences in myocardial protection during pediatric open heart surgery. AB - BACKGROUND: Current cardioplegic protection techniques used in pediatric cardiac surgery do not take into consideration age and cyanotic differences. The aim of the present work was to address this question by monitoring clinical outcome, myocardial metabolism, and reperfusion injury in pediatric patients protected by cold-crystalloid cardioplegia. METHODS AND RESULTS: Fifty-eight patients (31 children and 27 infants) with or without hypoxic stress (cyanosis) undergoing open heart surgery with cold-crystalloid cardioplegia were included in the study. Clinical outcome measures assessed included inotropic and ventilatory support, intensive care, and hospital stay. Ischemia-induced changes in metabolism (adenine nucleotides, purines, lactate, and amino acids) were determined in ventricular biopsies collected at the beginning and end of ischemic time (cross clamp time). Reperfusion injury was assessed by measuring postoperative serial release of troponin I. Evidence was observed of ischemic stress during cardioplegic arrest in children and infants as shown by significant changes in cellular metabolites. Compared with infants, children had significantly less reperfusion injury and better clinical outcome, and these factors were related to duration of ischemic time. Cyanosis did not influence outcome in infants, but cyanotic children showed worse reperfusion injury and clinical outcome than acyanotic children. CONCLUSIONS: Extent of myocardial protection with cold crystalloid cardioplegia in pediatric open heart surgery is dependent on age and degree of cyanosis. PMID- 11257083 TI - Effects of nonlipid risk factors on atherosclerosis in youth with a favorable lipoprotein profile. AB - BACKGROUND: The strong association between coronary heart disease and dyslipoproteinemia has often overshadowed the effects of the nonlipid risk factors-smoking, hypertension, obesity, and diabetes and impaired glucose tolerance-and even led to questioning the importance of these risk factors in the presence of a favorable lipoprotein profile. METHODS AND RESULTS: A cooperative multicenter study, the Pathobiological Determinants of Atherosclerosis in Youth (PDAY), examined the relation of the nonlipid risk factors to atherosclerosis in 629 men and 227 women 15 to 34 years of age who died of external causes and who had a favorable lipoprotein profile (non-HDL cholesterol <4.14 mmol/L [<160 mg/dL] and HDL cholesterol >/=0.91 mmol/L [>/=35 mg/dL]). In the abdominal aorta, smokers had more extensive fatty streaks and raised lesions than nonsmokers, and hypertensive blacks had more raised lesions than normotensive blacks. In the right coronary artery, hypertensive blacks had more raised lesions than normotensive blacks, obese men (body mass index >/=30 kg/m(2)) had more extensive fatty streaks and raised lesions than nonobese men, and individuals with impaired glucose intolerance had more extensive fatty streaks. Obese men had more severe lesions (American Heart Association grade 2 through 5) of the left anterior descending coronary artery. CONCLUSIONS: These substantial effects of the nonlipid risk factors on the extent and severity of coronary and aortic atherosclerosis, even in the presence of a favorable lipoprotein profile, support the need to control all cardiovascular risk factors. PMID- 11257085 TI - Endothelin-1 has a unique oxygen-saving effect by increasing contractile efficiency in the isolated rat heart. AB - BACKGROUND: The effect of endothelin (ET)-1 on cardiac energetics is not fully understood. METHODS AND RESULTS: In isolated, coronary-perfused rat hearts, we measured left ventricular contractility index (E(max)), pressure-volume area (PVA), and myocardial oxygen consumption (MVO(2)) before and after administration of ET-1 (1x10(-)(9) mol/L). ET-1 increased E(max) by 48+/-16% (P<0.01) and the total MVO(2) by 24+/-11% (P<0.01). The MVO(2)-PVA relations were linear both before and after ET-1 (r>0.99). ET-1 shifted MVO(2)-PVA upward, increasing the MVO(2) intercept by 24+/-13%. At the same time, ET-1 decreased the slope (S), with 1/S (contractile efficiency) being 46+/-5% before and 56+/-5% after ET-1 (P<0.01). ET-1-induced increases in E(max) and in contractile efficiency were abolished by an ET(A) receptor blocker (S-0139) but not by an ET(B) blocker (BQ 788). Although high [Ca(2+)] perfusion increased E(max) and the intercept to the same extent as ET-1, it did not change S. N(G)-Nitro-L-arginine (an inhibitor of nitric oxide synthase) increased the coronary perfusion pressure as much as ET-1, but S again remained unchanged. Dimethylamyloride (Na(+)/H(+) exchanger inhibitor) partially blocked the positive inotropic effect of ET-1 but not the ET 1-induced increase in the contractile efficiency. CONCLUSIONS: Agonistic effects of ET-1 on the ET(A) receptor economized the chemomechanical conversion efficiency of the left ventricular unit myocardium by a mechanism independent of the Na(+)/H(+) exchanger. This unique oxygen-saving effect of ET-1 may play an adaptive role in the failing myocardium, in which local accumulation of ET-1 is present. PMID- 11257086 TI - Serial magnetic resonance imaging of microvascular remodeling in the infarcted rat heart. AB - BACKGROUND: Alterations in the coronary circulation are important determinants of myocardial function. Few data are available, however, about microvascular changes in reactive hypertrophy. With MRI, serial determination of myocardial microcirculation after myocardial infarction (MI) is feasible. METHODS AND RESULTS: We quantitatively determined myocardial perfusion and relative intracapillary blood volume using an MRI technique. Infarct size, myocardial mass, and left ventricular volumes were determined with cine MRI. Rats were investigated at 8, 12, and 16 weeks after MI (mean MI size 24.1+/-2.0%) or sham operation. Vasodilation was induced by adenosine. In the infarcted group, maximum perfusion decreased significantly from 8 to 16 weeks (5.6+/-0.3 versus 3.5+/-0.2 mL. g(-1). min(-1), P<0.01) compared with sham animals (5.5+/-0.3 versus 5.0+/ 0.2 mL. g(-1). min(-1), P=0.17). Myocardial mass increased significantly (559.1+/ 20.8 mg at 8 weeks versus 690.9+/-42.7 mg at 16 weeks, P<0.05) compared with sham operated animals (516.3+/-41.7 versus 549.2+/-32.3 mg). Basal relative intracapillary blood volume increased significantly to 15.7+/-0.5 vol% at 8 weeks after MI and remained elevated (16.8+/-0.6 vol%) at 16 weeks compared with 12.1+/ 0.3 vol% (P<0.01) in sham-operated rats. CONCLUSIONS: Our results indicate that significant microvascular changes occur during cardiac remodeling. Hypoperfusion in the hypertrophied myocardium is related to an increase in vascular capacity, suggesting a compensatory vasodilatory response at the capillary level. These microvascular changes may therefore contribute to the development of heart failure. PMID- 11257087 TI - High-energy phosphate metabolism and creatine kinase in failing hearts: a new porcine model. AB - BACKGROUND: This study aimed to create a pig model of heart failure secondary to severe aortic stenosis and to examine the relationship between the alterations in myocardial high-energy phosphate (HEP) metabolism and protein expression of creatine kinase (CK) isoforms. METHODS AND RESULTS: Sixteen pigs with left ventricular hypertrophy (LVH) secondary to ascending aortic banding and 10 normal pigs (N) were studied. Myocardial protein levels of CK isoforms (Western blot), HEP levels, and CK kinetics ((31)P MR spectroscopy) were measured under basal conditions. Nine of the 16 animals with LVH developed congestive heart failure (CHF), as evidenced by ascites (100 to 2000 mL). LV weight/body weight ratio (g/kg) was 2.18+/-0.15 in N hearts, 3.04+/-0.14 in hearts with LVH (P<0.01), and 4.23+/-0.36 in hearts with CHF (P<0.01 versus LVH). Right ventricle weight/body weight ratio and LV end-diastolic pressure were significantly higher in hearts with CHF (each P<0.01 versus N or LVH). Myocardial phosphocreatine/ATP ratios and the CK forward flux rates were decreased in LVH hearts, most severely in hearts with CHF. CK-M/beta-actin ratios were 2.21+/-12 (N), 1.69+/-0.15 (LVH), and 1.39+/-0.27 (CHF, P<0.05 versus N). CK-mitochondria (CK-Mt)/beta-actin ratios were 1.40+/-0.09 (N), 1.24+/-0.09 (LVH), and 1.02+/-0.08 (CHF, P<0.05 versus N or LVH). The severity of the reduction of CK flux rate was linearly related to the severity of the decrease of CK-Mt/beta-actin (r=0.68, P<0.01). CONCLUSIONS: In this new model of heart failure/hypertrophy, the abnormal myocardial HEP metabolism is related to the decreased CK-Mt protein level, which in turn is related to the severity of the hypertrophy. PMID- 11257088 TI - Decreased sarcoplasmic reticulum calcium content is responsible for defective excitation-contraction coupling in canine heart failure. AB - BACKGROUND: Altered excitation-contraction (E-C) coupling in canine pacing induced heart failure involves decreased sarcoplasmic reticulum (SR) Ca uptake and enhanced Na/Ca exchange, which could be expected to decrease SR Ca content (Ca(SR)) and may explain the reduced intracellular Ca (Ca(i)) transient. Studies in other failure models have suggested that the intrinsic coupling between L-type Ca current (I:(Ca,L)) and SR Ca release is reduced without a change in SR Ca load. The present study investigates whether Ca(SR) and/or coupling is altered in midmyocardial myocytes from failing canine hearts (F). METHODS AND RESULTS: Myocytes were indo-1-loaded via patch pipette (37 degrees C), and Ca(i) transients were elicited with voltage-clamp steps applied at various frequencies. I(Ca,L) density was not significantly decreased in F, but steady-state Ca(i) transients were reduced to 20% to 40% of normal myocytes (N). Ca(SR), measured by integrating Na/Ca exchange currents during caffeine-induced release, was profoundly decreased in F, to 15% to 25% of N. When Ca(SR) was normalized in F by preloading in 5 mmol/L external Ca before a test pulse at 2 mmol/L Ca, a normal amplitude Ca(i) transient was elicited. E-C coupling gain was dependent on Ca(SR) but was affected similarly in both groups, indicating that intrinsic coupling is unaltered in F. CONCLUSIONS: A decrease in Ca(SR) is sufficient to explain the diminished Ca(i) transients in F, without a change in the effectiveness of coupling. Therefore, therapeutic approaches that increase Ca(SR) may be able to fully correct the Ca handling deficit in heart failure. PMID- 11257089 TI - Left ventricular hypertrophy decreases slowly but not rapidly activating delayed rectifier potassium currents of epicardial and endocardial myocytes in rabbits. AB - BACKGROUND: Delayed rectifier K(+) currents are critical to action potential (AP) repolarization. The present study examines the effects of left ventricular hypertrophy (LVH) on delayed rectifier K(+) currents and their contribution to AP repolarization in both epicardial (Epi) and endocardial (Endo) myocytes. METHODS AND RESULTS: VH was induced in rabbits by a 1-kidney removal, 1-kidney vascular clamping method. Slowly (I(Ks)) and rapidly (I(Kr)) activating delayed rectifier K(+) currents were recorded by the whole-cell patch-clamp technique, and APs were recorded by the microelectrode technique. In normal rabbit left ventricular myocytes, I(Ks) densities were larger in Epi than in Endo (1.1+/-0.1 versus 0.43+/-0.07 pA/pF), whereas I(Kr) density was similar between Epi and Endo (0.31+/-0.05 versus 0.36+/-0.07 pA/pF) at 20 mV. LVH reduced I(Ks) density to a similar extent (approximately 40%) in both Epi and Endo but had no significant effect on I(Kr) in either Epi or Endo. Consequently, I(Kr) was expected to contribute more to AP repolarization in LVH than in control. This was confirmed by specific I(Kr) block with dofetilide, which prolonged AP significantly more in LVH than in control (31+/-3% versus 18+/-2% in Epi; 53+/-6% versus 32+/-4% in Endo at 2 Hz). In contrast, L-768,673 (a specific I(Ks) blocker) prolonged AP less in LVH than in control. The very small I(Ks) density in Endo with LVH is consistent with the greater incidence of early afterdepolarizations induced in this region by dofetilide. CONCLUSIONS: LVH induces a decrease in I(Ks) density and increases the propensity to develop early afterdepolarizations, especially in Endo. PMID- 11257090 TI - Impaired conduction in the bundle branches of mouse hearts lacking the gap junction protein connexin40. AB - BACKGROUND: Connexin (Cx)40 and Cx45 are the major protein subunits of gap junction channels in the conduction system of mammals. To determine the role of Cx40, we correlated cardiac activation with Connexin distribution in normal and Cx40-deficient mice hearts. METHODS AND RESULTS: Epicardial and septal activation was recorded in Langendorff-perfused adult mice hearts with a 247-point compound electrode (interelectrode distance, 0.3 mm). After electrophysiological measurements, hearts were prepared for immunohistochemistry and histology to determine Connexin distribution and fibrosis. In both wild-type and Cx40 deficient animals, epicardial activation patterns were similar. The right and left ventricular septum was invariably activated from base to apex. Histology revealed a continuity of myocytes from the common bundle to the septal myocardium. Within this continuity, colocalization was found of Cx43 and Cx45 but not of Cx40 and Cx43. Both animals showed similar His-bundle activation. In Cx40 deficient mice, the proximal bundle branches expressed Cx45 only. The absence of Cx40 in the proximal bundles correlated with right bundle-branch block. Conduction in the left bundle branch was impaired as compared with wild-type animals. CONCLUSIONS: Our data show that (1) in mice, a continuity exists between the common bundle and the septum, and (2) Cx40 deficiency results in right bundle branch block and impaired left bundle-branch conduction. PMID- 11257091 TI - Direct inhibition of expressed cardiac l- and t-type calcium channels by igg from mothers whose children have congenital heart block. AB - BACKGROUND: Congenital heart block (CHB) is a disease that affects the offspring of mothers with autoimmune diseases. We recently reported that maternal sera containing antibodies against SSA/Ro and SSB/La ribonucleoproteins (positive IgG) inhibited L-type Ca current in isolated cardiac myocytes and induced sinus bradycardia in a murine model of CHB. The direct interaction of positive IgG with L-type Ca channel proteins and the possible inhibition of T-type Ca current that could account for the sinus bradycardia remain unknown. METHODS AND RESULTS: The 2-electrode voltage-clamp technique was used to record currents via L-type (I(Ba) alpha(1C) or I(Ba)-alpha(1C)+beta(2a)+alpha(2)/delta) and T-type (I(Ba) alpha(1H)) Ca channels, Na channels (I(Na)-hH1), and K channels (I(Ks) minK+KvLQT1) expressed in Xenopus oocytes. Positive IgG (350 microgram/mL) inhibited I(Ba)-alpha(1C) by 50.6+/-4.7% (P<0.01) and I(Ba) alpha(1C)+beta(2a)+alpha(2)/delta by 50.9+/-4.2% (P<0.01); I(Ba)-alpha(1H) was reduced by 18.9+/-1.0% (P<0.01). Immunoblot data show cross-reactivity of positive IgG with alpha(1C) subunit. Pretreatment of oocytes with atropine (1 micromol/L) or acetylcholine (10 micromol/L) did not affect the inhibitory effect of IgG on I(Ba)-alpha(1C) and I(Ba)-alpha(1C)+beta(2a)+alpha(2)/delta (P<0.05). Positive IgG had no effect, however, on either I(Na)-hH1 or I(Ks)-minK+KvLQT1. CONCLUSIONS: Positive IgG inhibited expressed L-type I:(Ba) and cross-reacted with the alpha(1C) subunit in Xenopus oocytes, providing strong evidence that maternal antibodies interact directly with the pore-forming alpha(1)-subunit of Ca channels. In addition, we show for the first time that positive IgG also inhibited T-type I(Ba) but not I(Na)-hH1 or I(Ks)-minK+KvLQT1. This could provide, in part, the ionic basis of sinus bradycardia reported in animal models of CHB and clinically in humans. PMID- 11257092 TI - Apical hypertrophic cardiomyopathy developing at a relatively advanced age. PMID- 11257093 TI - Percutaneous stenting of a vertebral artery supplying the entire brain. PMID- 11257094 TI - Institute of medicine report seeks overhaul of health system. PMID- 11257095 TI - The molecular genetics of cancer: down the rabbit hole. PMID- 11257096 TI - Tissue microarray technology for high-throughput molecular profiling of cancer. AB - Tissue microarray (TMA) technology allows rapid visualization of molecular targets in thousands of tissue specimens at a time, either at the DNA, RNA or protein level. The technique facilitates rapid translation of molecular discoveries to clinical applications. By revealing the cellular localization, prevalence and clinical significance of candidate genes, TMAs are ideally suitable for genomics-based diagnostic and drug target discovery. TMAs have a number of advantages compared with conventional techniques. The speed of molecular analyses is increased by more than 100-fold, precious tissues are not destroyed and a very large number of molecular targets can be analyzed from consecutive TMA sections. The ability to study archival tissue specimens is an important advantage as such specimens are usually not applicable in other high throughput genomic and proteomic surveys. Construction and analysis of TMAs can be automated, increasing the throughput even further. Most of the applications of the TMA technology have come from the field of cancer research. Examples include analysis of the frequency of molecular alterations in large tumor materials, exploration of tumor progression, identification of predictive or prognostic factors and validation of newly discovered genes as diagnostic and therapeutic targets. PMID- 11257097 TI - Genome and genetic resources from the Cancer Genome Anatomy Project. AB - The Cancer Genome Anatomy Project (CGAP) is a collaborative network of cancer researchers with a common goal: to decipher the genetic changes that occur during cancer formation and progression. The project brings together several recent technologies capable of high-throughput analysis to help achieve this goal. Automated sequencing of cDNA libraries is a primary focus and is geared towards providing a comprehensive and annotated set of human and mouse transcribed sequences. This effort includes full-length transcript sequence generated by CGAP's new Mammalian Gene Collection initiative. Single nucleotide polymorphisms (SNPs) within human gene sequences (Genetic Annotation Initiative) and chromosomal rearrangements within cancer cells (Cancer Chromosome Aberration Project) are also being cataloged as part of CGAP. Finally, to help determine gene expression patterns related to cancer, CGAP provides a quantitative catalog of data through its SAGEmap initiative. The genome and genetic analysis tools listed in this review are all freely distributed by CGAP (http://cgap.nci.nih.gov/) without restriction. PMID- 11257098 TI - Unraveling human cancer in the mouse: recent refinements to modeling and analysis. AB - The ability to manipulate the mouse genome has made the mouse the primary mammalian genetic model organism. It has been possible to model human cancer in the mouse by overexpressing oncogenes or inactivating tumor suppressor genes, and these experiments have provided much of our in vivo understanding of cancer. However, these transgenic approaches do not always completely and accurately model human carcinogenesis. Recent developments in transgenic and knockout approaches have improved the accuracy of modeling somatic cancer in the mouse and analyzing the genomic instability that occurs in murine tumors. It is possible to use retroviral gene delivery, chromosome engineering and inducible transgenes to selectively manipulate the genome in a more precise spatial and temporal pattern. In addition, the development of powerful cytogenetic tools such as spectral karyotyping, fluorescence in situ hybridization and comparative genome hybridization have improved our ability to detect chromosomal rearrangements. Finally, global patterns of gene expression can be determined by microarray analysis to decipher complex gene patterns which occur in cancers. Several of these advances in mouse modeling of human cancer are discussed in this review. PMID- 11257099 TI - Telomerase and cancer. AB - Telomerase, a eukaryotic ribonucleoprotein (RNP) complex, contains both an essential RNA and a protein reverse transcriptase subunit. By reverse transcription, the telomerase RNP maintains telomere length stability in almost all cancer cells. Over the past few years there has been significant progress in identifying the components of the telomerase holoenzyme complex and the proteins that associate with telomeres, in order to elucidate mechanisms of telomere length regulation. This review covers recent advances in the field including the use of telomerase in cancer diagnostics and an overview of anti-telomerase cancer therapeutic approaches. PMID- 11257100 TI - Aberrant patterns of DNA methylation, chromatin formation and gene expression in cancer. AB - Gene function in cancer can be disrupted either through genetic alterations, which directly mutate or delete genes, or epigenetic alterations, which alter the heritable state of gene expression. The latter events are mediated by formation of transcriptionally repressive chromatin states around gene transcription start sites and an associated gain of methylation in normally unmethylated CpG islands in these regions. The genes affected include over half of the tumor suppressor genes that cause familial cancers when mutated in the germline; the selective advantage for genetic and epigenetic dysfunction in these genes is very similar. The aberrant methylation can begin very early in tumor progression and mediate most of the important pathway abnormalities in cancer including loss of cell cycle control, altered function of transcription factors, altered receptor function, disruption of normal cell-cell and cell-substratum interaction, inactivation of signal transduction pathways, loss of apoptotic signals and genetic instability. The active role of the aberrant methylation in transcriptional silencing of genes is becoming increasingly understood and involves a synergy between the methylation and histone deacetylase (HDAC) activity. This synergy can be mediated directly by HDAC interaction with DNA methylating enzymes and by recruitment through complexes involving methyl cytosine binding proteins. In the translational arena, the promoter hypermethylation changes hold great promise as DNA tumor markers and their potentially reversible state creates a target for cancer therapeutic strategies involving gene reactivation. PMID- 11257101 TI - Transcriptional control at regulatory checkpoints by histone deacetylases: molecular connections between cancer and chromatin. AB - Cancer cells exhibit a set of unique properties that distinguish them from their normal counterparts. Among these features are increased growth rates, loss of differentiation, escape from cell death pathways, evasion of anti-proliferative signals, a decreased reliance on exogenous growth factors and escape from replicative senescence. Acquisition of these features by malignant cells requires impairment of normal cellular control mechanisms. Over the past few years, it has become increasingly apparent that an important subset of the molecular changes commonly found in cancer cells involves inappropriate regulation of gene expression. This review will address regulatory pathways whose disruption contributes to the malignant phenotype. The failure to deacetylate and thus repress transcription by the Class I histone deacetylases HDAC1 and HDAC2 due to disruption of the Rb family of proteins has been firmly established as a mechanism leading to increases in growth rate and cellular proliferation. Recent data suggest that this regulatory circuit also executes G(1) checkpoint arrest downstream of DNA damage, cellular senescence and contact inhibition. In contrast to this failure to deacetylate, it now seems probable that changes in differentiation status may result in part from inappropriate deacetylation and concomitant transcriptional repression mediated by the Class II histone deacetylases. This inappropriate deacetylation by HDAC4, HDAC5 and HDAC6 follows their relocalization from the cytoplasm to the nucleus. Thus, multiple classical features of cancer cells can be manifested by improper histone deacetylation. PMID- 11257102 TI - The Rb/E2F pathway and cancer. AB - Over the past decade, studies focusing on the mechanisms controlling cellular proliferation have converged with equally intensive efforts directed at the analysis of oncogenic pathways associated with human cancer. These convergent studies have revealed the central role played by the pathway that controls the activity of the retinoblastoma tumor suppressor protein (Rb), which in turn regulates the E2F transcription factor. In particular, it is now clear that the Rb/E2F pathway is critical in regulating the initiation of DNA replication. It is also clear that the control of the pathway is disrupted in virtually all human cancers. Questions remain, however, as to the specific role played by individual activities within the pathway in the control of cell growth and their participation in the development of cancer. PMID- 11257103 TI - BRCA1 and BRCA2 and the genetics of breast and ovarian cancer. AB - Germline mutations in the tumor suppressor genes BRCA1 and BRCA2 predispose individuals to breast and ovarian cancers. Progress in determining the function of BRCA1 and BRCA2 suggests that they are involved in two fundamental cellular processes: DNA damage repair and transcriptional regulation. We evaluate current knowledge of BRCA1 and BRCA2 functions to explain why mutations in BRCA1 and BRCA2 lead specifically to breast and ovarian cancer. The BRCA1 and BRCA2 genes contain unusually high densities of repetitive elements. These features of the BRCAs genomic regions contribute to chromosomal instability of these genes. We propose that somatic alterations of BRCA1 and BRCA2 are common and driven by rearrangements between repetitive elements. Inherited and somatic mutations occur in BRCA1 and BRCA2; virtually all somatic mutations are the result of large genomic rearrangements. What are the consequences of such large somatic mutations of BRCA1 and BRCA2 in women with or without inherited mutations? The breast and ovary are estrogen-responsive tissues. Beginning in puberty, the breast epithelium proliferates rapidly in response to fluctuating levels of estrogen. We present a genetic model outlining how BRCA-deficient cells may gain uncontrolled proliferation leading to tumor formation. Central to this model of BRCA-mediated tumorigenesis are estrogen-mediated proliferation of breast and ovarian epithelium and the distinctive genomic context of the BRCA genes. PMID- 11257104 TI - "Other" breast cancer susceptibility genes: searching for more holy grail. AB - While germline mutations in BRCA1 and BRCA2 account for most, if not all families with autosomal dominant transmission of susceptibility to both breast and ovarian cancer, it has become clear that together these genes only account for a small proportion of hereditary site-specific breast cancer susceptibility. However, difficulties due to genetic heterogeneity, reduced penetrance and perhaps gene mutation frequency complicate ongoing efforts to identify additional susceptibility genes. Therefore, multiple approaches are being used to identify additional high and low penetrance genes. Families with three or more breast cancer cases are being used in traditional linkage studies, which are expected to yield only moderate or high penetrance susceptibility genes. Breast cancer case control studies are being used to look for genetic variants or polymorphisms that confer an increased risk of breast cancer in a wide variety of cellular pathways, ranging from the detoxification of environmental carcinogens to steroid hormone metabolism, DNA damage repair and immune surveillance, an approach useful primarily to identify low penetrance susceptibililty genes. However, neither approach has yielded convincing results to date. A third approach, using BRCA1 and BRCA2 mutation carriers to identify genes that are associated with modification of breast cancer risk has met with some limited success, perhaps because effects on breast cancer risk in BRCA1 and BRCA2 mutation carriers are more readily detected in smaller studies, given the much higher number of events in these cohorts at very high risk of breast cancer. Clearly, hereditary breast cancer susceptibility is a complex phenomenon, in which multiple genes may play a role. It will be necessary to use all of these approaches, as well as more comprehensive genomic studies, to identify additional breast cancer-related genes. PMID- 11257105 TI - The ABC of APC. AB - Familial adenomatous polyposis (FAP) is an autosomal dominant inherited disease characterized by the presence of adenomatous polyps in the colon and rectum, with inevitable development of colorectal cancer if left untreated. FAP is caused by germline mutations in the adenomatous polyposis coli (APC) gene. Somatic mutations in the APC gene are an early event in colorectal tumorigenesis, and can be detected in the majority of colorectal tumours. The APC gene encodes a large protein with multiple cellular functions and interactions, including roles in signal transduction in the wnt-signalling pathway, mediation of intercellular adhesion, stabilization of the cytoskeleton and possibly regulation of the cell cycle and apoptosis. The fact that APC is an integral part of so many different pathways makes it an ideal target for mutation in carcinogenesis. This review deals with our understanding to date of how mutations in the APC gene translate into changes at the protein level, which in turn contribute to the role of APC in tumorigenesis. PMID- 11257106 TI - Deficient DNA mismatch repair: a common etiologic factor for colon cancer. AB - Hereditary non-polyposis colon cancer (HNPCC), the most common form of hereditary colon cancer, is a syndrome of deficient DNA mismatch repair (MMR). Five, possibly six, human MMR genes have been identified that, when mutated in the germline, cause susceptibility to this syndrome. To date, more than 300 different predisposing mutations are known, mainly affecting the MMR genes MLH1 ( approximately 50%), MSH2 ( approximately 40%) and MSH6 ( approximately 10%). Genetically predisposed individuals carry a defective copy of an MMR gene in every cell. Somatic inactivation of the remaining wild-type copy in a target tissue, typically colon, gives rise to a profound repair defect, progressive accumulation of mutations and cancer. Instability at short tandem repeat sequences, microsatellites, is a typical manifestation of MMR deficiency and apart from HNPCC tumors, occurs in approximately 15% of sporadic colon and other tumors. The majority of the latter cases are attributable to one particular MMR gene, MLH1, and unlike HNPCC, an epigenetic rather than a genetic mechanism plays an important role in the inactivation of this gene. The present review provides an update of the genetics of HNPCC and more generally, of cancer development driven by deficient MMR. Recent discoveries suggest that apart from post replication repair, MMR proteins have several other functions that are highly relevant to carcinogenesis. Knowledge of the complex interplay between the MMR system and other cellular pathways allows us to better understand the phenotypic manifestations of HNPCC and other cancers with deficient MMR. PMID- 11257107 TI - DNA helicase deficiencies associated with cancer predisposition and premature ageing disorders. AB - Deficiency in a helicase of the RecQ family is found in at least three human genetic disorders associated with cancer predisposition and/or premature ageing. The RecQ helicases encoded by the BLM, WRN and RECQ4 genes are defective in Bloom's, Werner's and Rothmund-Thomson syndromes, respectively. Cells derived from individuals with these disorders in each case show inherent genomic instability. Recent studies have demonstrated direct interactions between these RecQ helicases and human nuclear proteins required for several aspects of chromosome maintenance, including p53, BRCA1, topoisomerase III, replication protein A and DNA polymerase delta. Here, we review this network of protein interactions, and the clues that they present regarding the potential roles of RecQ family members in DNA repair, replication and/or recombination pathways. PMID- 11257108 TI - The neurofibromatoses: when less is more. AB - The study of cancer predisposition syndromes presents unique opportunities to gain insights into the genetic events associated with tumor pathogenesis. Individuals with two inherited cancer syndromes, neurofibromatosis 1 (NF1) and neurofibromatosis 2 (NF2), develop both benign and malignant tumors. The corresponding genes mutated in these two disorders encode tumor suppressor proteins, termed neurofibromin (NF1) and merlin (NF2), which function in novel ways to regulate cell growth and differentiation. Neurofibromin inhibits cell proliferation, at least in part, by modulating mitogenic pathway signaling through inactivation of p21-ras. In contrast, merlin may act as a membrane associated molecular switch that regulates cell-cell and cell-matrix signals transduced by cell surface receptors. Significant progress in our understanding of the genetics and biology of NF1 and NF2 has elucidated the roles of these genes in tumor initiation and progression. PMID- 11257109 TI - The hedgehog pathway and basal cell carcinomas. AB - Developmental pathways first elucidated by genetic studies in the fruit fly, Drosophila melanogaster, are conserved in vertebrates, and disruption of these pathways has been associated with various human congenital anomalies. Many developmental genes continue to play an important role in regulation of cell growth and differentiation after embryogenesis, and mutations in some of these genes can result in cancer. Basal cell carcinoma (BCC) of the skin is the most common type of cancer in humans. Although most BCCs are sporadic, in rare cases, individuals have a hereditary disease, Gorlin syndrome, that predisposes to multiple skin tumors as well as a variety of birth defects. Mutations in the human homolog of a Drosophila gene, patched, underlie Gorlin syndrome. Genetic studies in Drosophila show that patched is part of the hedgehog signaling pathway, important in determining embryonic patterning and cell fate in multiple structures of the developing embryo. Human patched is mutated in sporadic as well as hereditary BCCs, and inactivation of this gene is probably a necessary if not sufficient step for tumor formation. Delineation of the biochemical pathway in which patched functions may lead to rational medical therapy for skin cancer and possibly other tumors. PMID- 11257110 TI - Genotype-phenotype correlation in von Hippel-Lindau syndrome. AB - The von Hippel-Lindau (VHL) syndrome (OMIM 193300) is an autosomal dominant disorder caused by deletions or mutations in a tumor suppressor gene on human chromosome 3p25. It is characterized clinically by vascular tumors including benign hemangioblastomas of the cerebellum, spine, brain stem and retina. Clear cell renal cell carcinoma is a frequent cause of death, occurring in up to 70% of patients with VHL. Pheochromocytomas occur in association with specific alleles (usually mutations as opposed to deletions), therefore a family history of pheochromocytoma in association with VHL is an indication for thorough surveillance for pheochromocytoma in affected family members. The VHL gene coding sequence contains three exons. Two isoforms of mRNA exist, reflecting the presence or absence of exon 2. Tumors arise following the loss or inactivation of the wild-type allele in a cell. In initial studies approximately 20% of patients had large germline mutations detectable by Southern blot analysis, 27% had missense mutations and 27% had nonsense or frameshift mutations. Advances in mutation analysis now allow for a 100% mutation detection rate in families with definite VHL. Families may be characterized by the presence [type 2 (7-20% of families)] or absence (type 1) of pheochromocytomas. Most type 2 families are affected by missense mutations, whereas most type 1 families have deletions or premature termination mutations. The prognosis for the lifetime risk of pheochromocytoma can be estimated by determination of the underlying mutation even if there is no family history of VHL. PMID- 11257111 TI - Oncogenes and tumor suppressors in the molecular pathogenesis of acute promyelocytic leukemia. AB - Acute promyelocytic leukemia (APL) is associated with reciprocal chromosomal translocations always involving the retinoic acid receptor alpha (RARalpha) gene on chromosome 17 and variable partner genes (X genes) on distinct chromosomes. RARalpha fuses to the PML gene in the vast majority of APL cases, and in a few cases to the PLZF, NPM, NuMA and Stat5b genes, respectively, leading to the generation of RARalpha-X: and X:-RARalpha fusion genes. Both fusion proteins can exert oncogenic functions through their ability to interfere with the activities of X and RARalpha proteins. Here, it will be discussed in detail how an extensive biochemical analysis as well as a systematic in vivo genetic approach in the mouse has allowed the definition of the multiple oncogenic activities of PML RARalpha, and how it has become apparent that this oncoprotein is able to impair RARalpha at the transcription level and the tumor suppressive function of the PML protein. PMID- 11257112 TI - Potential of gene therapy for brain tumors. AB - Brain tumors comprise a broad spectrum of biological and clinical entities making it unlikely for any single therapeutic approach to be universally applicable. In particular, malignant glioblastoma multiforme have defied all current therapeutic modalities. Gene therapy offers the potential to augment current neurosurgical, radiation and drug treatments with little increase in morbidity. Many therapeutic transgenes have shown efficacy in experimental models, including generation of toxic compounds, enzymatic activation of pro-drugs, expression of tumor suppressor or apoptotic proteins, inhibition of angiogenesis and enhancement of immune responses to tumor antigens. Vectors have been used as gene delivery vehicles and as cytotoxic agents in their own right by selective replication and lysis of tumor cells, thereby also generating vectors on-site. Brain tumors appear to offer some "Achilles' heels" in that they are usually contained within the brain and represent a unique dividing cell population there. However, the heterogeneous and invasive characteristics of these tumor cells, as well as sequestration of tumor antigens within a relatively immune privileged location present serious problems for effective therapy. This review will focus on current transgene/vector strategies, including novel therapeutic genes, combinational therapies and new delivery modalities, the latter of which appears to be the rate limiting factor for gene therapy of brain tumors in humans. PMID- 11257114 TI - UGO1 encodes an outer membrane protein required for mitochondrial fusion. AB - Membrane fusion plays an important role in controlling the shape, number, and distribution of mitochondria. In the yeast Saccharomyces cerevisiae, the outer membrane protein Fzo1p has been shown to mediate mitochondrial fusion. Using a novel genetic screen, we have isolated new mutants defective in the fusion of their mitochondria. One of these mutants, ugo1, shows several similarities to fzo1 mutants. ugo1 cells contain numerous mitochondrial fragments instead of the few long, tubular organelles seen in wild-type cells. ugo1 mutants lose mitochondrial DNA (mtDNA). In zygotes formed by mating two ugo1 cells, mitochondria do not fuse and mix their matrix contents. Fragmentation of mitochondria and loss of mtDNA in ugo1 mutants are rescued by disrupting DNM1, a gene required for mitochondrial division. We find that UGO1 encodes a 58-kD protein located in the mitochondrial outer membrane. Ugo1p appears to contain a single transmembrane segment, with its NH(2) terminus facing the cytosol and its COOH terminus in the intermembrane space. Our results suggest that Ugo1p is a new outer membrane component of the mitochondrial fusion machinery. PMID- 11257115 TI - Diacylglycerol kinase zeta regulates Ras activation by a novel mechanism. AB - Guanine nucleotide exchange factors (GEFs) activate Ras by facilitating its GTP binding. Ras guanyl nucleotide-releasing protein (GRP) was recently identified as a Ras GEF that has a diacylglycerol (DAG)-binding C1 domain. Its exchange factor activity is regulated by local availability of signaling DAG. DAG kinases (DGKs) metabolize DAG by converting it to phosphatidic acid. Because they can attenuate local accumulation of signaling DAG, DGKs may regulate RasGRP activity and, consequently, activation of Ras. DGK zeta, but not other DGKs, completely eliminated Ras activation induced by RasGRP, and DGK activity was required for this mechanism. DGK zeta also coimmunoprecipitated and colocalized with RasGRP, indicating that these proteins associate in a signaling complex. Coimmunoprecipitation of DGK zeta and RasGRP was enhanced in the presence of phorbol esters, which are DAG analogues that cannot be metabolized by DGKs, suggesting that DAG signaling can induce their interaction. Finally, overexpression of kinase-dead DGK zeta in Jurkat cells prolonged Ras activation after ligation of the T cell receptor. Thus, we have identified a novel way to regulate Ras activation: through DGK zeta, which controls local accumulation of DAG that would otherwise activate RasGRP. PMID- 11257116 TI - Rac mediates cytoskeletal rearrangements and increased cell motility induced by urokinase-type plasminogen activator receptor binding to vitronectin. AB - The urokinase-type plasminogen activator receptor (uPAR) is involved in the regulation of cell motility in a variety of cell types. We show here that expression of human uPAR in growing murine fibroblasts leads to a dramatic reorganization of the actin cytoskeleton. uPAR expression induces multiple rapidly advancing protrusions that resemble the leading edge of migrating cells. The cytoskeletal changes are independent of uPA and activation of the RGD-binding activity of integrins but require uPAR binding to vitronectin (VN). The actin reorganization is blocked by coexpression of dominant negative versions of either Rac (N17Rac) or p130Cas, but not by inhibitors of Cdc42 or Rho, and is accompanied by a Rac-dependent increase in cell motility. In addition, a fourfold increase in the level of activated Rac is induced by uPAR expression. We conclude that uPAR interacts with VN both to initiate a p130Cas/Rac-dependent signaling pathway leading to actin reorganization and increased cell motility and to act as an adhesion receptor required for these responses. This mechanism may play a role in uPAR-mediated regulation of cell motility at sites where VN and uPAR are co expressed, such as malignant tumors. PMID- 11257117 TI - An acidic motif retains vesicular monoamine transporter 2 on large dense core vesicles. AB - The release of biogenic amines from large dense core vesicles (LDCVs) depends on localization of the vesicular monoamine transporter VMAT2 to LDCVs. We now find that a cluster of acidic residues including two serines phosphorylated by casein kinase 2 is required for the localization of VMAT2 to LDCVs. Deletion of the acidic cluster promotes the removal of VMAT2 from LDCVs during their maturation. The motif thus acts as a signal for retention on LDCVs. In addition, replacement of the serines by glutamate to mimic phosphorylation promotes the removal of VMAT2 from LDCVs, whereas replacement by alanine to prevent phosphorylation decreases removal. Phosphorylation of the acidic cluster thus appears to reduce the localization of VMAT2 to LDCVs by inactivating a retention mechanism. PMID- 11257118 TI - A role for syndecan-1 in coupling fascin spike formation by thrombospondin-1. AB - An important role of cell matrix adhesion receptors is to mediate transmembrane coupling between extracellular matrix attachment, actin reorganization, and cell spreading. Thrombospondin (TSP)-1 is a modulatory component of matrix expressed during development, immune response, or wound repair. Cell adhesion to TSP-1 involves formation of biochemically distinct matrix contacts based on stable fascin spikes. The cell surface adhesion receptors required have not been identified. We report here that antibody clustering of syndecan-1 proteoglycan specifically transduces organization of cortical actin and fascin bundles in several cell types. Transfection of COS-7 cells with syndecan-1 is sufficient to stimulate cell spreading, fascin spike assembly, and extensive protrusive lateral ruffling on TSP-1 or on syndecan-1 antibody. The underlying molecular mechanism depends on glycosaminoglycan (GAG) modification of the syndecan-1 core protein at residues S45 or S47 for cell membrane spreading and on the VC2 region of the cytoplasmic domain for spreading and fascin spike formation. Expression of the VC2 deletion mutant or GAG-negative syndecan-1 showed that syndecan-1 is necessary in spreading and fascin spike formation by C2C12 cells on TSP-1. These results establish a novel role for syndecan-1 protein in coupling a physiological matrix ligand to formation of a specific matrix contact structure. PMID- 11257119 TI - Atypical protein kinase C is involved in the evolutionarily conserved par protein complex and plays a critical role in establishing epithelia-specific junctional structures. AB - We have previously shown that during early Caenorhabditis elegans embryogenesis PKC-3, a C. elegans atypical PKC (aPKC), plays critical roles in the establishment of cell polarity required for subsequent asymmetric cleavage by interacting with PAR-3 [Tabuse, Y., Y. Izumi, F. Piano, K.J. Kemphues, J. Miwa, and S. Ohno. 1998. Development (Camb.). 125:3607--3614]. Together with the fact that aPKC and a mammalian PAR-3 homologue, aPKC-specific interacting protein (ASIP), colocalize at the tight junctions of polarized epithelial cells (Izumi, Y., H. Hirose, Y. Tamai, S.-I. Hirai, Y. Nagashima, T. Fujimoto, Y. Tabuse, K.J. Kemphues, and S. Ohno. 1998. J. Cell Biol. 143:95--106), this suggests a ubiquitous role for aPKC in establishing cell polarity in multicellular organisms. Here, we show that the overexpression of a dominant-negative mutant of aPKC (aPKCkn) in MDCK II cells causes mislocalization of ASIP/PAR-3. Immunocytochemical analyses, as well as measurements of paracellular diffusion of ions or nonionic solutes, demonstrate that the biogenesis of the tight junction structure itself is severely affected in aPKCkn-expressing cells. Furthermore, these cells show increased interdomain diffusion of fluorescent lipid and disruption of the polarized distribution of Na(+),K(+)-ATPase, suggesting that epithelial cell surface polarity is severely impaired in these cells. On the other hand, we also found that aPKC associates not only with ASIP/PAR-3, but also with a mammalian homologue of C. elegans PAR-6 (mPAR-6), and thereby mediates the formation of an aPKC-ASIP/PAR-3-PAR-6 ternary complex that localizes to the apical junctional region of MDCK cells. These results indicate that aPKC is involved in the evolutionarily conserved PAR protein complex, and plays critical roles in the development of the junctional structures and apico-basal polarization of mammalian epithelial cells. PMID- 11257120 TI - The high mobility group (HMG) boxes of the nuclear protein HMG1 induce chemotaxis and cytoskeleton reorganization in rat smooth muscle cells. AB - HMG1 (high mobility group 1) is a ubiquitous and abundant chromatin component. However, HMG1 can be secreted by activated macrophages and monocytes, and can act as a mediator of inflammation and endotoxic lethality. Here we document a role of extracellular HMG1 in cell migration. HMG1 (and its individual DNA-binding domains) stimulated migration of rat smooth muscle cells in chemotaxis, chemokinesis, and wound healing assays. HMG1 induced rapid and transient changes of cell shape, and actin cytoskeleton reorganization leading to an elongated polarized morphology typical of motile cells. These effects were inhibited by antibodies directed against the receptor of advanced glycation endproducts, indicating that the receptor of advanced glycation endproducts is the receptor mediating the HMG1-dependent migratory responses. Pertussis toxin and the mitogen activated protein kinase kinase inhibitor PD98059 also blocked HMG1-induced rat smooth muscle cell migration, suggesting that a G(i/o) protein and mitogen activated protein kinases are required for the HMG1 signaling pathway. We also show that HMG1 can be released by damage or necrosis of a variety of cell types, including endothelial cells. Thus, HMG1 has all the hallmarks of a molecule that can promote atherosclerosis and restenosis after vascular damage. PMID- 11257121 TI - Enhanced expression of the alpha 7 beta 1 integrin reduces muscular dystrophy and restores viability in dystrophic mice. AB - Muscle fibers attach to laminin in the basal lamina using two distinct mechanisms: the dystrophin glycoprotein complex and the alpha 7 beta 1 integrin. Defects in these linkage systems result in Duchenne muscular dystrophy (DMD), alpha 2 laminin congenital muscular dystrophy, sarcoglycan-related muscular dystrophy, and alpha 7 integrin congenital muscular dystrophy. Therefore, the molecular continuity between the extracellular matrix and cell cytoskeleton is essential for the structural and functional integrity of skeletal muscle. To test whether the alpha 7 beta 1 integrin can compensate for the absence of dystrophin, we expressed the rat alpha 7 chain in mdx/utr(-/-) mice that lack both dystrophin and utrophin. These mice develop a severe muscular dystrophy highly akin to that in DMD, and they also die prematurely. Using the muscle creatine kinase promoter, expression of the alpha 7BX2 integrin chain was increased 2.0-2.3-fold in mdx/utr(-/-) mice. Concomitant with the increase in the alpha 7 chain, its heterodimeric partner, beta 1D, was also increased in the transgenic animals. Transgenic expression of the alpha 7BX2 chain in the mdx/utr(-/-) mice extended their longevity by threefold, reduced kyphosis and the development of muscle disease, and maintained mobility and the structure of the neuromuscular junction. Thus, bolstering alpha 7 beta 1 integrin-mediated association of muscle cells with the extracellular matrix alleviates many of the symptoms of disease observed in mdx/utr(-/-) mice and compensates for the absence of the dystrophin- and utrophin-mediated linkage systems. This suggests that enhanced expression of the alpha 7 beta 1 integrin may provide a novel approach to treat DMD and other muscle diseases that arise due to defects in the dystrophin glycoprotein complex. A video that contrasts kyphosis, gait, joint contractures, and mobility in mdx/utr(-/-) and alpha 7BX2-mdx/utr(-/-) mice can be accessed at http://www.jcb.org/cgi/content/full/152/6/1207. PMID- 11257122 TI - The NC1/endostatin domain of Caenorhabditis elegans type XVIII collagen affects cell migration and axon guidance. AB - Type XVIII collagen is a homotrimeric basement membrane molecule of unknown function, whose COOH-terminal NC1 domain contains endostatin (ES), a potent antiangiogenic agent. The Caenorhabditis elegans collagen XVIII homologue, cle-1, encodes three developmentally regulated protein isoforms expressed predominantly in neurons. The CLE-1 protein is found in low amounts in all basement membranes but accumulates at high levels in the nervous system. Deletion of the cle-1 NC1 domain results in viable fertile animals that display multiple cell migration and axon guidance defects. Particular defects can be rescued by ectopic expression of the NC1 domain, which is shown to be capable of forming trimers. In contrast, expression of monomeric ES does not rescue but dominantly causes cell and axon migration defects that phenocopy the NC1 deletion, suggesting that ES inhibits the promigratory activity of the NC1 domain. These results indicate that the cle 1 NC1/ES domain regulates cell and axon migrations in C. elegans. PMID- 11257123 TI - Oligomerization-dependent regulation of motility and morphogenesis by the collagen XVIII NC1/endostatin domain. AB - Collagen XVIII (c18) is a triple helical endothelial/epithelial basement membrane protein whose noncollagenous (NC)1 region trimerizes a COOH-terminal endostatin (ES) domain conserved in vertebrates, Caenorhabditis elegans and Drosophila. Here, the c18 NC1 domain functioned as a motility-inducing factor regulating the extracellular matrix (ECM)-dependent morphogenesis of endothelial and other cell types. This motogenic activity required ES domain oligomerization, was dependent on rac, cdc42, and mitogen-activated protein kinase, and exhibited functional distinction from the archetypal motogenic scatter factors hepatocyte growth factor and macrophage stimulatory protein. The motility-inducing and mitogen activated protein kinase-stimulating activities of c18 NC1 were blocked by its physiologic cleavage product ES monomer, consistent with a proteolysis-dependent negative feedback mechanism. These data indicate that the collagen XVIII NC1 region encodes a motogen strictly requiring ES domain oligomerization and suggest a previously unsuspected mechanism for ECM regulation of motility and morphogenesis. PMID- 11257124 TI - Angiomotin: an angiostatin binding protein that regulates endothelial cell migration and tube formation. AB - Angiostatin, a circulating inhibitor of angiogenesis, was identified by its ability to maintain dormancy of established metastases in vivo. In vitro, angiostatin inhibits endothelial cell migration, proliferation, and tube formation, and induces apoptosis in a cell type-specific manner. We have used a construct encoding the kringle domains 1--4 of angiostatin to screen a placenta yeast two-hybrid cDNA library for angiostatin-binding peptides. Here we report the identification of angiomotin, a novel protein that mediates angiostatin inhibition of migration and tube formation of endothelial cells. In vivo, angiomotin is expressed in the endothelial cells of capillaries as well as larger vessels of the human placenta. Upon expression of angiomotin in HeLa cells, angiomotin bound and internalized fluorescein-labeled angiostatin. Transfected angiomotin as well as endogenous angiomotin protein were localized to the leading edge of migrating endothelial cells. Expression of angiomotin in endothelial cells resulted in increased cell migration, suggesting a stimulatory role of angiomotin in cell motility. However, treatment with angiostatin inhibited migration and tube formation in angiomotin-expressing cells but not in control cells. These findings indicate that angiostatin inhibits cell migration by interfering with angiomotin activity in endothelial cells. PMID- 11257125 TI - Budding yeast chromosome structure and dynamics during mitosis. AB - Using green fluorescent protein probes and rapid acquisition of high-resolution fluorescence images, sister centromeres in budding yeast are found to be separated and oscillate between spindle poles before anaphase B spindle elongation. The rates of movement during these oscillations are similar to those of microtubule plus end dynamics. The degree of preanaphase separation varies widely, with infrequent centromere reassociations observed before anaphase. Centromeres are in a metaphase-like conformation, whereas chromosome arms are neither aligned nor separated before anaphase. Upon spindle elongation, centromere to pole movement (anaphase A) was synchronous for all centromeres and occurred coincident with or immediately after spindle pole separation (anaphase B). Chromatin proximal to the centromere is stretched poleward before and during anaphase onset. The stretched chromatin was observed to segregate to the spindle pole bodies at rates greater than centromere to pole movement, indicative of rapid elastic recoil between the chromosome arm and the centromere. These results indicate that the elastic properties of DNA play an as of yet undiscovered role in the poleward movement of chromosome arms. PMID- 11257126 TI - Cyclin E uses Cdc6 as a chromatin-associated receptor required for DNA replication. AB - Using an in vitro chromatin assembly assay in Xenopus egg extract, we show that cyclin E binds specifically and saturably to chromatin in three phases. In the first phase, the origin recognition complex and Cdc6 prereplication proteins, but not the minichromosome maintenance complex, are necessary and biochemically sufficient for ATP-dependent binding of cyclin E--Cdk2 to DNA. We find that cyclin E binds the NH(2)-terminal region of Cdc6 containing Cy--Arg-X-Leu (RXL) motifs. Cyclin E proteins with mutated substrate selection (Met-Arg-Ala-Ile-Leu; MRAIL) motifs fail to bind Cdc6, fail to compete with endogenous cyclin E--Cdk2 for chromatin binding, and fail to rescue replication in cyclin E--depleted extracts. Cdc6 proteins with mutations in the three consensus RXL motifs are quantitatively deficient for cyclin E binding and for rescuing replication in Cdc6-depleted extracts. Thus, the cyclin E--Cdc6 interaction that localizes the Cdk2 complex to chromatin is important for DNA replication. During the second phase, cyclin E--Cdk2 accumulates on chromatin, dependent on polymerase activity. In the third phase, cyclin E is phosphorylated, and the cyclin E--Cdk2 complex is displaced from chromatin in mitosis. In vitro, mitogen-activated protein kinase and especially cyclin B--Cdc2, but not the polo-like kinase 1, remove cyclin E- Cdk2 from chromatin. Rebinding of hyperphosphorylated cyclin E--Cdk2 to interphase chromatin requires dephosphorylation, and the Cdk kinase-directed Cdc14 phosphatase is sufficient for this dephosphorylation in vitro. These three phases of cyclin E association with chromatin may facilitate the diverse activities of cyclin E--Cdk2 in initiating replication, blocking rereplication, and allowing resetting of origins after mitosis. PMID- 11257127 TI - Internal modification of U2 small nuclear (sn)RNA occurs in nucleoli of Xenopus oocytes. AB - U2 small nuclear (sn)RNA contains a large number of posttranscriptionally modified nucleotides, including a 5' trimethylated guanosine cap, 13 pseudouridines, and 10 2'-O-methylated residues. Using Xenopus oocytes, we demonstrated previously that at least some of these modified nucleotides are essential for biogenesis of a functional snRNP. Here we address the subcellular site of U2 internal modification. Upon injection into the cytoplasm of oocytes, G capped U2 that is transported to the nucleus becomes modified, whereas A-capped U2 that remains in the cytoplasm is not modified. Furthermore, by injecting U2 RNA into isolated nuclei or enucleated oocytes, we observe that U2 internal modifications occur exclusively in the nucleus. Analysis of the intranuclear localization of fluorescently labeled RNAs shows that injected wild-type U2 becomes localized to nucleoli and Cajal bodies. Both internal modification and nucleolar localization of U2 are dependent on the Sm binding site. An Sm-mutant U2 is targeted only to Cajal bodies. The Sm binding site can be replaced by a nucleolar localization signal derived from small nucleolar RNAs (the box C/D motif), resulting in rescue of internal modification as well as nucleolar localization. Analysis of additional chimeric U2 RNAs reveals a correlation between internal modification and nucleolar localization. Together, our results suggest that U2 internal modification occurs within the nucleolus. PMID- 11257128 TI - Glial growth factor/neuregulin inhibits Schwann cell myelination and induces demyelination. AB - During development, neuregulin-1 promotes Schwann cell proliferation and survival; its role in later events of Schwann cell differentiation, including myelination, is poorly understood. Accordingly, we have examined the effects of neuregulin-1 on myelination in neuron-Schwann cell cocultures. Glial growth factor (GGF), a neuregulin-1 isoform, significantly inhibited myelination by preventing axonal segregation and ensheathment. Basal lamina formation was not affected. Treatment of established myelinated cultures with GGF resulted in striking demyelination that frequently began at the paranodes and progressed to the internode. Demyelination was dose dependent and accompanied by dedifferentiation of Schwann cells to a promyelinating stage, as evidenced by reexpression of the transcription factor suppressed cAMP-inducible POU; a significant proportion of cells with extensive demyelination also proliferated. Two other Schwann cell mitogens, fibroblast growth factor-2 and transforming growth factor-beta, inhibited myelination but did not cause demyelination, suggesting this effect is specific to the neuregulins. The neuregulin receptor proteins, erbB2 and erbB3, are expressed on ensheathing and myelinating Schwann cells and rapidly phosphorylated with GGF treatment. GGF treatment of myelinating cultures also induced phosphorylation of phosphatidylinositol 3-kinase, mitogen activated protein kinase, and a 120-kD protein. These results suggest that neuronal mitogens, including the neuregulins, may inhibit myelination during development and that activation of mitogen signaling pathways may contribute to the initial demyelination and subsequent Schwann cell proliferation observed in various pathologic conditions. PMID- 11257129 TI - Oncogenic Ras-induced proliferation requires autocrine fibroblast growth factor 2 signaling in skeletal muscle cells. AB - Constitutively activated Ras proteins are associated with a large number of human cancers, including those originating from skeletal muscle tissue. In this study, we show that ectopic expression of oncogenic Ras stimulates proliferation of the MM14 skeletal muscle satellite cell line in the absence of exogenously added fibroblast growth factors (FGFs). MM14 cells express FGF-1, -2, -6, and -7 and produce FGF protein, yet they are dependent on exogenously supplied FGFs to both maintain proliferation and repress terminal differentiation. Thus, the FGFs produced by these cells are either inaccessible or inactive, since the endogenous FGFs elicit no detectable biological response. Oncogenic Ras-induced proliferation is abolished by addition of an anti-FGF-2 blocking antibody, suramin, or treatment with either sodium chlorate or heparitinase, demonstrating an autocrine requirement for FGF-2. Oncogenic Ras does not appear to alter cellular export rates of FGF-2, which does not possess an NH(2)-terminal or internal signal peptide. However, oncogenic Ras does appear to be involved in releasing or activating inactive, extracellularly sequestered FGF-2. Surprisingly, inhibiting the autocrine FGF-2 required for proliferation has no effect on oncogenic Ras-mediated repression of muscle-specific gene expression. We conclude that oncogenic Ras-induced proliferation of skeletal muscle cells is mediated via a unique and novel mechanism that is distinct from Ras-induced repression of terminal differentiation and involves activation of extracellularly localized, inactive FGF-2. PMID- 11257130 TI - Importin beta-mediated nuclear import of fibroblast growth factor receptor: role in cell proliferation. AB - Although growth factor receptors are generally thought to carry out their role in signal transduction at the cell surface, many of these transmembrane proteins translocate to the nucleus after ligand stimulation. Here, we show that the nuclear translocation of fibroblast growth factor receptor (FGFR)1 occurs via a mechanism distinct from classical nuclear import but dependent on importin beta, a component of multiple nuclear import pathways. Furthermore, we show that nuclear FGFR1 induces c-Jun and is involved in the regulation of cell proliferation. These data are the first description of a nuclear import pathway for transmembrane growth factor receptors and elucidate a novel signal transduction pathway from the cell surface to the nucleus. PMID- 11257131 TI - The Caenorhabditis elegans unc-78 gene encodes a homologue of actin-interacting protein 1 required for organized assembly of muscle actin filaments. AB - Assembly and maintenance of myofibrils require dynamic regulation of the actin cytoskeleton. In Caenorhabditis elegans, UNC-60B, a muscle-specific actin depolymerizing factor (ADF)/cofilin isoform, is required for proper actin filament assembly in body wall muscle (Ono, S., D.L. Baillie, and G.M. Benian. 1999. J. Cell Biol. 145:491--502). Here, I show that UNC-78 is a homologue of actin-interacting protein 1 (AIP1) and functions as a novel regulator of actin organization in myofibrils. In unc-78 mutants, the striated organization of actin filaments is disrupted, and large actin aggregates are formed in the body wall muscle cells, resulting in defects in their motility. Point mutations in unc-78 alleles change conserved residues within different WD repeats of the UNC-78 protein and cause less severe phenotypes than a deletion allele, suggesting that these mutations partially impair the function of UNC-78. UNC-60B is normally localized in the diffuse cytoplasm and to the myofibrils in wild type but mislocalized to the actin aggregates in unc-78 mutants. Similar Unc-78 phenotypes are observed in both embryonic and adult muscles. Thus, AIP1 is an important regulator of actin filament organization and localization of ADF/cofilin during development of myofibrils. PMID- 11257132 TI - Hold that line. Angiomotin regulates endothelial cell motility. PMID- 11257134 TI - A dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin (DC-SIGN)-related protein is highly expressed on human liver sinusoidal endothelial cells and promotes HIV-1 infection. AB - The discovery of dendritic cell (DC)-specific intercellular adhesion molecule (ICAM)-3-grabbing nonintegrin (DC-SIGN) as a DC-specific ICAM-3 binding receptor that enhances HIV-1 infection of T cells in trans has indicated a potentially important role for adhesion molecules in AIDS pathogenesis. A related molecule called DC-SIGNR exhibits 77% amino acid sequence identity with DC-SIGN. The DC SIGN and DC-SIGNR genes map within a 30-kb region on chromosome 19p13.2-3. Their strong homology and close physical location indicate a recent duplication of the original gene. Messenger RNA and protein expression patterns demonstrate that the DC-SIGN-related molecule is highly expressed on liver sinusoidal cells and in the lymph node but not on DCs, in contrast to DC-SIGN. Therefore, we suggest that a more appropriate name for the DC-SIGN-related molecule is L-SIGN, liver/lymph node-specific ICAM-3-grabbing nonintegrin. We show that in the liver, L-SIGN is expressed by sinusoidal endothelial cells. Functional studies indicate that L SIGN behaves similarly to DC-SIGN in that it has a high affinity for ICAM-3, captures HIV-1 through gp120 binding, and enhances HIV-1 infection of T cells in trans. We propose that L-SIGN may play an important role in the interaction between liver sinusoidal endothelium and trafficking lymphocytes, as well as function in the pathogenesis of HIV-1. PMID- 11257133 TI - Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) contributes to interferon gamma-dependent natural killer cell protection from tumor metastasis. AB - Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is expressed by in vitro activated natural killer (NK) cells, but the relevance of this observation to the biological function of NK cells has been unclear. Herein, we have demonstrated the in vivo induction of mouse TRAIL expression on various tissue NK cells and correlated NK cell activation with TRAIL-mediated antimetastatic function in vivo. Expression of TRAIL was only constitutive on a subset of liver NK cells, and innate NK cell control of Renca carcinoma hepatic metastases in the liver was partially TRAIL dependent. Administration of therapeutic doses of interleukin (IL)-12, a powerful inducer of interferon (IFN) gamma production by NK cells and NKT cells, upregulated TRAIL expression on liver, spleen, and lung NK cells, and IL-12 suppressed metastases in both liver and lung in a TRAIL-dependent fashion. By contrast, alpha-galactosylceramide (alpha-GalCer), a powerful inducer of NKT cell IFN-gamma and IL-4 secretion, suppressed both liver and lung metastases but only stimulated NK cell TRAIL mediated function in the liver. TRAIL expression was not detected on NK cells from IFN-gamma-deficient mice and TRAIL-mediated antimetastatic effects of IL-12 and alpha-GalCer were strictly IFN-gamma dependent. These results indicated that TRAIL induction on NK cells plays a critical role in IFN-gamma-mediated antimetastatic effects of IL-12 and alpha-GalCer. PMID- 11257135 TI - Pivotal role of signal transducer and activator of transcription (Stat)4 and Stat6 in the innate immune response during sepsis. AB - Signal transducer and activator of transcription (Stat)4 and Stat6 are transcription factors that provide type 1 and type 2 response, respectively. Here, we explored the role of Stat4 and Stat6 in innate immunity during septic peritonitis. Stat4-/- and Stat6-/- mice were resistant to the lethality compared with wild-type (WT) mice. At the mechanistic level, bacterial levels in Stat6-/- mice were much lower than in WT mice, which was associated with increased peritoneal levels of interleukin (IL)-12, tumor necrosis factor (TNF)-alpha, macrophage-derived chemokine (MDC), and C10, known to enhance bacterial clearance. In Stat4-/- mice, hepatic inflammation and injury during sepsis were significantly ameliorated without affecting local responses. This event was associated with increased hepatic levels of IL-10 and IL-13, while decreasing those of macrophage inflammatory protein (MIP)-2 and KC. Sepsis-induced renal injury was also abrogated in Stat4-/- mice, which was accompanied by decreased renal levels of MIP-2 and KC without altering IL-10 and IL-13 levels. Thus, Stat6 /- and Stat4-/- mice appeared to be resistant to septic peritonitis by enhancing local bacterial clearance and modulating systemic organ damage, respectively, via balancing cytokine responses. These results clearly highlight an important role of local type 1 and systemic type 2 cytokine response in protective immunity during sepsis, which can be regulated by Stat proteins. PMID- 11257136 TI - Evidence that gammadelta versus alphabeta T cell fate determination is initiated independently of T cell receptor signaling. AB - Two types of T cells, alphabeta and gammadelta, develop in vertebrates. How these two T cell lineages arise from a common thymic T progenitor is poorly understood. Differentiation of alphabeta lineage T cells requires the surrogate alpha chain (pTalpha), which associates with the T cell receptor (TCR) beta chain to form the pre-TCR. gammadelta lineage development does not appear to involve an obligatory surrogate chain, but instead requires productive rearrangement and expression of both TCR gamma and delta genes. It has been proposed that the quality of signals transmitted by the pre-TCR and gammadelta TCR are distinct and that these "instructive" signals determine the lineage fate of an uncommitted progenitor cell. Here we show that the thymic T progenitor cells (CD25(+)CD44(+)c-kit(+)CD3( )CD4(-)CD8(-) thymocytes, termed pro-T cells) from young adult mice that have yet to express TCRs can be subdivided based on interleukin 7 receptor (IL-7R) expression. These subsets exhibit differential potential to develop into gammadelta versus alphabeta lineage (CD4+CD8+ cells) in the thymus. Upon intrathymic injection, IL-7R(neg-lo) pro-T cells generated a 13-fold higher ratio of alphabeta lineage to gammadelta lineage cells than did IL-7R(+) pro-T cells. Much of this difference was due to a fivefold greater potential of IL-7R(+) pro-T cells to develop into TCR-gammadelta T cells. Evidence indicates that this biased developmental potential is not a result of enhanced TCR-gamma gene rearrangement/expression in IL-7R(+) pro-T cells. These results indicate that the pro-T cells are heterogeneous in developmental potential before TCR gene rearrangement and suggest that in some precursor cells the initial lineage commitment is independent of TCR-mediated signals. PMID- 11257137 TI - Insertion of phosphoglycerine kinase (PGK)-neo 5' of Jlambda1 dramatically enhances VJlambda1 rearrangement. AB - Gene-targeted mice were generated with a loxP-neomycin resistance gene (neo(r)) cassette inserted upstream of the Jlambda1 region and replacement of the glycine 154 codon in the Clambda1 gene with a serine codon. This insertion dramatically increases Vlambda1-Jlambda1 recombination. Jlambda1 germline transcription levels in pre-B cells and thymus cells are also greatly increased, apparently due to the strong housekeeping phosphoglycerine kinase (PGK) promoter driving the neo gene. In contrast, deletion of the neo gene causes a significant decrease in VJlambda1 recombination to levels below those in normal mice. This reduction is due to the loxP site left on the chromosome which reduces the Jlambda1 germline transcription in cis. Thus, the correlation between germline transcription and variable (V), diversity (D), and joining (J) recombination is not just an all or none phenomenon. Rather, the transcription efficiency is directly associated with the recombination efficiency. Furthermore, Jlambda1 and Vlambda1 germline transcription itself is not sufficient to lead to VJ recombination in T cells or early pre-B cells. The findings may suggest that in vivo: (a) locus and cell type specific transactivators direct the immunoglobulin or T cell receptor loci, respectively, to a "recombination factory" in the nucleus, and (b) transcription complexes deliver V(D)J recombinase to the recombination signal sequences. PMID- 11257138 TI - Absence of monocyte chemoattractant protein 1 in mice leads to decreased local macrophage recruitment and antigen-specific T helper cell type 1 immune response in experimental autoimmune encephalomyelitis. AB - Monocyte chemoattractant protein (MCP)-1 plays a critical role in innate immunity by directing the migration of monocytes into inflammatory sites. Recent data indicated a function for this chemokine in adaptive immunity as a regulator of T cell commitment to T helper cell type 2 (Th2) effector function. Studies in a Th1 dependent animal model, experimental autoimmune encephalomyelitis (EAE), showed that MCP-1 was highly expressed in the central nervous system (CNS) of affected rodents, and MCP-1 antibodies could block relapses of the disease. Mice deficient for the major MCP-1 receptor, CC chemokine receptor (CCR)2, did not develop EAE after active immunization but generated effector cells that could transfer the disease to naive wild-type recipients. We analyzed EAE in mice deficient for MCP 1 to define the relevant ligand for CCR2, which responds to murine MCP-1, MCP-2, MCP-3, and MCP-5. We found that C57BL/6 MCP-1-null mice were markedly resistant to EAE after active immunization, with drastically impaired recruitment of macrophages to the CNS, yet able to generate effector T cells that transferred severe disease to naive wild-type recipients. By contrast, adoptive transfer of primed T cells from wild-type mice into naive MCP-1-null recipients did not mediate clinical EAE. On the SJL background, disruption of the MCP-1 gene produced a milder EAE phenotype with diminished relapses that mimicked previous findings using anti-MCP-1 antibodies. There was no compensatory upregulation of MCP-2, MCP-3, or MCP-5 in MCP-1-null mice with EAE. These results indicated that MCP-1 is the major CCR2 ligand in mice with EAE, and provided an opportunity to define the role of MCP-1 in EAE. Compared with wild-type littermates, MCP-1-/- mice exhibited reduced expression of interferon gamma in draining lymph node and CNS and increased antigen-specific immunoglobulin G1 antibody production. Taken together, these data demonstrate that MCP-1 is crucial for Th1 immune responses in EAE induction and that macrophage recruitment to the inflamed CNS target organ is required for primed T cells to execute a Th1 effector program in EAE. PMID- 11257139 TI - Colony-stimulating factor 1 promotes progression of mammary tumors to malignancy. AB - In human breast carcinomas, overexpression of the macrophage colony-stimulating factor (CSF-1) and its receptor (CSF-1R) correlates with poor prognosis. To establish if there is a causal relationship between CSF-1 and breast cancer progression, we crossed a transgenic mouse susceptible to mammary cancer with mice containing a recessive null mutation in the CSF-1 gene (Csf1(op)) and followed tumor progression in wild-type and null mutant mice. The absence of CSF 1 affects neither the incidence nor the growth of the primary tumors but delayed their development to invasive, metastatic carcinomas. Transgenic expression of CSF-1 in the mammary epithelium of both Csf1(op)/Csf1(op) and wild-type tumor prone mice led to an acceleration to the late stages of carcinoma and to a significant increase in pulmonary metastasis. This was associated with an enhanced infiltration of macrophages into the primary tumor. These studies demonstrate that the growth of mammary tumors and the development to malignancy are separate processes and that CSF-1 selectively promotes the latter process. CSF-1 may promote metastatic potential by regulating the infiltration and function of tumor-associated macrophages as, at the tumor site, CSF-1R expression was restricted to macrophages. Our data suggest that agents directed at CSF-1/CSF 1R activity could have important therapeutic effects. PMID- 11257141 TI - Neonatally induced inactivation of the vascular cell adhesion molecule 1 gene impairs B cell localization and T cell-dependent humoral immune response. AB - Vascular cellular adhesion molecule (VCAM)-1 is a membrane-bound cellular adhesion molecule that mediates adhesive interactions between hematopoietic progenitor cells and stromal cells in the bone marrow (BM) and between leukocytes and endothelial as well as dendritic cells. Since VCAM-1-deficient mice die embryonically, conditional VCAM-1 mutant mice were generated to analyze the in vivo function of this adhesion molecule. Here we show that interferon-induced Cre loxP-mediated deletion of the VCAM-1 gene after birth efficiently ablates expression of VCAM-1 in most tissues like, for example, BM, lymphoid organs, and lung, but not in brain. Induced VCAM-1 deficiency leads to a reduction of immature B cells in the BM and to an increase of these cells in peripheral blood but not in lymphoid organs. Mature recirculating B cells are reduced in the BM. In a migration assay, the number of mature B cells that appears in the BM after intravenous injection is decreased. In addition, the humoral immune response to a T cell-dependent antigen is impaired. VCAM-1 serves an important role for B cell localization and the T cell-dependent humoral immune response. PMID- 11257140 TI - Conditional vascular cell adhesion molecule 1 deletion in mice: impaired lymphocyte migration to bone marrow. AB - We generated vascular cell adhesion molecule (VCAM)-1 "knock-in" mice and Cre recombinase transgenic mice to delete the VCAM-1 gene (vcam-1) in whole mice, thereby overcoming the embryonic lethality seen with conventional vcam-1 deficient mice. vcam-1 knock-in mice expressed normal levels of VCAM-1 but showed loss of VCAM-1 on endothelial and hematopoietic cells when interbred with a "TIE2Cre" transgene. Analysis of peripheral blood from conditional vcam-1 deficient mice revealed mild leukocytosis, including elevated immature B cell numbers. Conversely, the bone marrow (BM) had reduced immature B cell numbers, but normal numbers of pro-B cells. vcam-1-deficient mice also had reduced mature IgD+ B and T cells in BM and a greatly reduced capacity to support short-term migration of transferred B cells, CD4+ T cells, CD8+ T cells, and preactivated CD4+ T cells to the BM. Thus, we report an until now unappreciated dominant role for VCAM-1 in lymphocyte homing to BM. PMID- 11257142 TI - Induction of a diverse T cell receptor gamma/delta repertoire by the helix-loop helix proteins E2A and HEB in nonlymphoid cells. AB - During specific stages of thymocyte development, the T cell receptor (TCR) locus is assembled from variable (V), diversity (D), and joining (J) gene segments. Proper TCR gamma and delta V(D)J rearrangement during thymocyte development requires the presence of the E2A proteins. Here we show that E2A and a closely related protein, HEB, in the presence of recombination activating gene (RAG)1 and RAG2, each have the ability to activate TCR gamma and delta rearrangement in human kidney cells. The coding joints are diverse, contain nucleotide deletions, and occasionally show the presence of P nucleotides. Interestingly, only a subset of V, D, and J segments are targeted by the E2A and HEB proteins. Thus, E2A and HEB permit localized accessibility of the TCR gamma and delta loci to the recombination machinery. These data indicate that a distinct but diverse TCR repertoire can be induced in nonlymphoid cells by the mere presence of the V(D)J recombinase and the transcriptional regulators, E2A and HEB. PMID- 11257143 TI - The inhibition of arginase by N(omega)-hydroxy-l-arginine controls the growth of Leishmania inside macrophages. AB - Polyamine synthesis from l-ornithine is essential for Leishmania growth. We have investigated the dependence of Leishmania infection on arginase, which generates l-ornithine, in macrophages from BALB/c, C57BL/6, and nitric oxide synthase II (NOS II)-deficient mouse strains. We have found that N(omega)-hydroxy-l-arginine (LOHA), a physiological inhibitor of arginase, controls cellular infection and also specifically inhibits arginase activity from Leishmania major and Leishmania infantum parasites. The effect was proportional to the course of infection, concentration dependent up to 100 microM, and achieved without an increase in nitrite levels of culture supernatants. Moreover, when the l-arginine metabolism of macrophages is diverted towards ornithine generation by interleukin 4-induced arginase I, parasite growth is promoted. This effect can be reversed by LOHA. Inhibition of NOS II by N(G)-methyl-l-arginine (LNMMA) restores the killing obtained in the presence of interferon (IFN)-gamma plus lipolysaccharide (LPS), whereas the nitric oxide scavenger 2-(4-carboxyphenyl)-4,4,5,5, tetramethylimidazoline-3-oxide-1-oxyl (carboxy-PTIO) was without effect. However, exogenous l-ornithine almost completely inhibits parasite killing when added in the presence of LOHA to macrophages from NOS II-deficient mice or to BALB/c infected cells activated with IFN-gamma plus LPS. These results suggest that LOHA is an effector molecule involved in the control of Leishmania infection. In addition, macrophage arginase I induction by T helper cell type 2 cytokines could be a mechanism used by parasites to spread inside the host. PMID- 11257145 TI - The 'zero patient' design to compare the prevalences of rare diseases. PMID- 11257146 TI - Mortality associated with cervical spine disorders: a population-based study of 1666 patients with rheumatoid arthritis who died in Finland in 1989. AB - OBJECTIVE: To evaluate the mortality associated with cervical spine deformities in rheumatoid arthritis (RA) based on national data. METHODS: The role of rheumatoid disorders of the cervical spine as a cause of death was studied in 1666 subjects who died in Finland in 1989 and had been entitled under the national sickness insurance scheme to receive reimbursed medication for RA. Death certificates and certificates for drug reimbursement of these 1666 patients and the clinical files of 853 patients were examined for the mention of cervical spine disorders. Thereafter, the cervical spine radiographs and detailed clinical histories of patients with diagnosed cervical spine disorder were evaluated separately. RESULTS: According to the official death certificates, cervical spine disorder was not an underlying, contributory or immediate cause of death in any of these patients. Cervical spine abnormalities had been diagnosed only in 38/853 (4.5%) patients. Cervical spine radiographs from 33 patients were available for examination, and in 17 patients cervical spine deformities were found to be severe enough to be a potential cause of fatal complications. Among these 17 cases, four sudden and four postoperative deaths were recorded (one after cervical spine operation) and three patients were suffering from quadriparesis or paraparesis at the time of death. Among the other 16 patients with cervical spine radiographs, the cervical deformities were less severe and their death histories differed from those of the group with more severe deformities. CONCLUSIONS: Cervical spine disorders in RA should be diagnosed early and treated actively to prevent severe and potentially fatal complications. Deaths caused by these disorders are rare, but they should be remembered when the death certificates are written. PMID- 11257144 TI - Inflammatory cells and cancer: think different! PMID- 11257147 TI - Evaluating the cutaneous involvement in scleroderma: torsional stiffness revisited. AB - OBJECTIVE: The pace, progression and extent of the skin lesions in scleroderma may parallel the risk of new internal organ involvement and the progression of existing internal lesions. Accurate assessment of cutaneous change permits an evaluation of patient prognosis and the response to therapy. The aim of this study was to assess a simple device for measuring skin stiffness for its ability to measure sclerodermatous skin in a quantitative and reproducible manner. Materials and methods. Torsional skin stiffness was measured in 56 normal subjects and 42 scleroderma patients (31 of whom had the limited form and nine the diffuse form, and two had mixed connective tissue disease). Data for the scleroderma patients were compared with data obtained by the use of the modified Rodnan clinical skin scoring technique. Intraclass correlation coefficients (ICCs) were calculated as a measure of intraobserver and interobserver variability. RESULTS: For the left and right hands respectively, the ICCs for intraobserver variability were 0.908 and 0.906 and those for interobserver variability were 0.871 and 0.628. There was a significant difference in mean angular rotation obtained by normal subjects compared with scleroderma patients (15.1 vs 11.3 degrees, P<0.001). There was a significant difference in the angular rotation with increasing severity of skin involvement (skin score 0, median rotation 16.3 degrees; score 1, 10.5 degrees; score 2, 8.5 degrees; score 3, 8.0 degrees; P<0.00001). CONCLUSIONS: The measurements obtained with the skin stiffness device are highly reproducible and are consistent with the current clinical method of assessment of skin involvement. The significant difference in angular rotation obtained by normal subjects and scleroderma patients indicates that the device can distinguish normal from sclerodermatous skin. The torsional stiffness measurements derived from the device may also be useful in longitudinal studies. PMID- 11257149 TI - Increased urinary pyridinoline cross-link compounds of collagen in patients with systemic sclerosis and Raynaud's phenomenon. AB - OBJECTIVE: To study concentration changes in collagen degradation markers in patients with diffuse and limited cutaneous systemic sclerosis and patients with scleroderma-related diseases. METHODS: Pyridinoline cross-link compounds were analysed in urine samples using high-performance liquid chromatography. Samples were analysed for pyridinoline (Pyr), deoxypyridinoline (Dpyr) and soft-tissue pyridinoline (stPyr) in patients with diffuse cutaneous systemic sclerosis (dcSSc, n=23) and limited cutaneous systemic sclerosis (lcSSc, n=48) and in patients with scleroderma-related diseases such as primary Raynaud's phenomenon (pRP, n=16) and secondary Raynaud's phenomenon (sRP, n=14). Healthy controls (n=18) and patients with post-menopausal osteoporosis (OP, n=35) were also investigated. RESULTS: Urinary Pyr, Dpyr and stPyr concentrations were significantly higher in patients with Raynaud's phenomenon and systemic sclerosis than in healthy controls. The highest concentrations (two to three times greater than in healthy controls) were found in patients with dcSSc. The stPyr concentration was significantly higher in patients with dcSSc than in those with lcSSc, sRP and pRP. No significant difference in stPyr concentration was found between the healthy controls and the OP group, suggesting that stPyr is derived from soft tissues rather than bone. The extent and severity of skin involvement, measured as a skin score, significantly correlated with the concentrations of stPyr and Pyr, whereas no such correlation was found for Dpyr. CONCLUSIONS: Increased urinary concentrations of piridinoline cross-links reflect alterations in collagen turnover in both Raynaud's phenomenon and systemic sclerosis. The close correlation between stPyr concentration and the extent of skin involvement in systemic sclerosis suggests that this parameter may be useful in monitoring ongoing fibrosis in this disease. PMID- 11257148 TI - Evidence for a protective role of the human leukocyte antigen class II region in early rheumatoid arthritis. AB - OBJECTIVE: To analyse the distribution of predisposing DQ3 (DQB1*03(*04)/DQA1*03) and DQ5 (DQB1*0501/DQA1*01), shared epitope encoding and protective DRB1 alleles in patients with early rheumatoid arthritis (RA) and undifferentiated arthritis (UA). METHODS: Consecutive patients enrolled in an early arthritis clinic were DNA-typed for HLA-DQ and DR. RA patients (n=195), UA patients (n=160) and controls from the same region (n=306) were sorted according to their DQ-DR phenotypes: DQ3 vs DQ5 and the presence or absence of a protective DERAA-positive DRB1 molecule. The three groups were also sorted according to the shared epitope (SE) hypothesis. RESULTS: We observed the association of both DQ3 and DQ5 with RA. DQ3/3 homozygous individuals had the highest risk of developing disease [odds ratio (OR)=20.00]. Conversely DQ5, but not DQ3, was associated with undifferentiated arthritis (OR=2.15 vs 1.25). Consistent with these differences, DQ3-positive individuals had significantly more active disease at the first visit at the outpatient clinic than DQ5-positive patients. DRB1 alleles encoding a DERAA motif in their third hypervariable region provided a strong dominant protection against RA among DQ5-positive individuals and decreased arthritis activity among DQ3-positive patients. Individuals with SE-positive DR1, DR4 and DR10 alleles were also predisposed to RA, DR4/4 homozygous individuals having the highest risk of developing RA (OR=11.00). CONCLUSION: The DQ3-DR4/9 haplotypes are associated with RA. The DQ5-DR1/10 haplotypes are associated with less active disease, i.e. UA, and DERAA encoding DRB1 alleles modulate either predisposition to or the severity of RA. We propose that HLA polymorphism influences not only the initiation or perpetuation of RA but also protection against the disease. PMID- 11257150 TI - Are antineutrophil cytoplasmic antibodies a marker predictive of relapse in Wegener's granulomatosis? A prospective study. AB - OBJECTIVES: To investigate the predictive value of testing for antineutrophil cytoplasmic antibodies (ANCA) in 55 patients with systemic Wegener's granulomatosis (WG) included in a randomized, prospective trial comparing corticosteroids and oral or pulse cyclophosphamide. METHODS: All 55 patients received corticosteroids. A cyclophosphamide pulse of 0.7 g/m2 was given at the time of diagnosis. After the first pulse, the patients were assigned at random to receive either pulse or oral cyclophosphamide (2 mg/kg/day), independently of ANCA results. ANCA were sought using an immunofluorescence assay and an attempt was made to correlate them with relapse of WG. ANCA were monitored throughout the study. RESULTS: At the time of diagnosis, ANCA were detected in 48 (87%) patients, with a cytoplasmic labelling pattern in 44 and a perinuclear pattern in four. ANCA follow-up was available for 50 patients. ANCA disappeared in 34 patients and persisted in nine. For 79% of the patients, the clinical course improved with the disappearance of ANCA and deteriorated with their persistence or increased titre. Among the patients who were initially ANCA-positive, 23 relapses occurred. Relapses were more frequent when ANCA remained positive or reappeared [13/19 ANCA-positive patients vs 3/29 ANCA-negative patients (P<0.01)]. Nine relapses (39%) occurred in patients with persistent ANCA, and ANCA reappearance preceded relapse in eight (35%). The mean time between inclusion and relapse did not differ between the patients who became ANCA negative and those who were persistently ANCA-positive (14.6+/-13.2 vs 14.4+/-8.2 months). The mean time to ANCA disappearance was similar for the patients who relapsed and those who did not. Corticosteroids and pulse or oral cyclophosphamide did not significantly modify the time to ANCA disappearance. Throughout the study, seven patients were ANCA-negative. CONCLUSION: Although ANCA positivity was associated with relapse, discordance between cytoplasmic ANCA and disease activity was not unusual. In the absence of clinical manifestations, ANCA titres alone can serve as a warning signal but not indicate whether to adjust or initiate treatment. PMID- 11257151 TI - Human parvovirus B19 infection in patients with systemic lupus erythematosus. AB - OBJECTIVE: The clinical significance of the presence of B19 DNA in patients with SLE was studied. METHODS: Sera from 72 patients with systemic lupus erythematosus (SLE), 23 patients with rheumatoid arthritis (RA), 18 patients with Sjogren's syndrome (SS), eight patients with Raynaud's phenomenon (RP), five patients with primary biliary cirrhosis (PBC), five patients with polymyositis (PM), four patients with erythema infectiosum (EI) and 22 normal controls were examined for parvovirus B19 (B19) infection by serological assays, nested PCR and Southern blotting. RESULTS: Parvovirus B19 DNA was detected in 17 of 72 patients with SLE and in three of four patients with EI, but not in patients with other systemic rheumatic diseases. Of the 17 patients with B19 DNA, only one had IgG anti-B19 antibody and two had IgM anti-B19 antibodies, whereas IgG and IgM anti-B19 antibodies were detected in 27 (49.1%) and 21 (38.2%) of 55 SLE patients without B19 DNA respectively. All sera from the patients with EI contained both IgG and IgM anti-B19 antibodies. B19 DNA was found more commonly in sera from SLE patients without anti-B19 antibodies than in those with anti-B19 antibodies (P<0.05). CONCLUSIONS: B19 infection in patients with SLE may be due to lack of anti-B19 antibodies because of either the immunocompromised nature of the host or the use of immunosuppressive drugs. There was a higher prevalence of hypocomplementaemia and RP in patients with parvovirus B19 viraemia than in those without parvovirus B19 viraemia. PMID- 11257152 TI - Morphological analysis of knee synovial membrane biopsies from a randomized controlled clinical study comparing the effects of sodium hyaluronate (Hyalgan) and methylprednisolone acetate (Depomedrol) in osteoarthritis. AB - OBJECTIVE: The study was part of a randomized open-label clinical trial designed to evaluate the effects of intra-articular injections of hyaluronan (Hyalgan) (HY) in osteoarthritis (OA) of the human knee. Data were compared with those obtained after treatment with methylprednisolone acetate (Depomedrol) (MP). METHODS: Synovial membranes from patients with OA of the knee, primary or secondary to a traumatic event and classified according to the American College of Rheumatology criteria, were examined by arthroscopy and by light and electron microscopy before and 6 months after local injection of HY (2 ml of 500-730 000 MW hyaluronan, 10 mg/ml in saline, one injection per week for 5 weeks) or MP (1 ml of methylprednisolone acetate, 40 mg/ml, one injection per week for 3 weeks). RESULTS: Arthroscopy revealed a significant decrease in inflammatory score after both treatments. Histology showed that HY treatment was effective (P< or =0.05) in reducing the number and aggregation of lining synoviocytes, as well as the number and calibre of the vessels. MP treatment significantly reduced the number of mast cells in primary OA. Both treatments tended to decrease the number of hypertrophic and to increase the number of fibroblast-like lining cells, to decrease the numbers of macrophages, lymphocytes, mast cells and adipocytes, and to decrease oedema, especially in primary OA, and to increase the number of fibroblasts and the amount of collagen. These phenomena were evident throughout the thickness of the synovial tissue. CONCLUSION: At least in the medium term, both HY and MP modified a number of structural variables of the synovial membrane of the osteoarthritic human knee towards the appearance of that of normal synovium. The effect was more evident in primary OA than in OA secondary to a traumatic event. This is the first evidence that local hyaluronan injections modify the structural organization of the human knee synovium in OA. PMID- 11257153 TI - The sternoclavicular syndrome: experience from a district general hospital and results of a national postal survey. AB - OBJECTIVE: To report our local experience of the sternoclavicular syndrome and sample the experience of other rheumatologists in the UK. METHODS: We studied case records of 23 patients referred to the Southend rheumatology clinic and data obtained from a postal questionnaire survey of British rheumatologists. RESULTS: We describe 58 cases (20 males and 38 females, mean age 47.2 yr). The disease was unilateral in 40 patients. Shoulder and/or arm pain (38 cases) with limitation of shoulder movements was an important presenting feature; other presenting features were anterior chest wall pain (14 cases) and neck pain (15 cases). Peripheral joint involvement was seen in 12 cases. Skin rash was reported in 12 cases (psoriasis, 6; acne, 2; none had pustulosis). No patients had symptoms or signs of sacroiliitis, and HLA-B27 was negative in 22 out of 23 patients. 99Technetium scintiscanning showed increased uptake in the sternoclavicular region in 31/34 patients (91.1%), but not in the sacroiliac areas. Plain radiographs were abnormal in 18 cases (sclerosis, 9; erosions, 2; soft tissue swelling, 2; bony expansion, 5). CT and/or MRI scans (available in 27 cases) showed erosions in 12 and osteitis in 18. Available histology showed a variable picture, including inflammation, bone erosion, sterile osteomyelitis and fibrosis. The majority of patients (45) were treated with non-steroidal anti-inflammatory drugs: 12 received steroids and 10 received disease-modifying anti-rheumatic drugs (methotrexate, 4; sulphasalazine, 6). Follow-up information was available for 38 patients, of whom 14 became asymptomatic and 24 had chronic disease with intermittent flares. CONCLUSION: Sternoclavicular disease is not uncommon in the UK. It can present with pain in the shoulder, neck or anterior chest wall, and may be underdiagnosed. Our results do not show a link with acne or pustulosis. Features of spondyloarthropathies, such as sacroiliitis and HLA-B27 positivity, were rare in this survey. PMID- 11257154 TI - Prevalence and spectrum of rheumatic diseases associated with proteinase 3 antineutrophil cytoplasmic antibodies (ANCA) and myeloperoxidase-ANCA. AB - OBJECTIVES: To evaluate the prevalence and association of antineutrophil cytoplasmic antibodies (ANCA) and their subtypes [proteinase 3 (PR3)-ANCA, myeloperoxidase (MPO)-ANCA] with distinct clinical features in various clinicopathological syndromes. METHODS: All consecutive ANCA-positive patients seen at the combined unit for rheumatology for Bad Bramstedt and the University of Lubeck between 1989 and 1999 were analysed. ANCA were detected by an immunofluorescence technique and ANCA subspecificities were determined by ELISA. Clinical features at presentation and diagnoses were recorded according to standardized procedures. RESULTS: Among 4620 patients tested, 333 were cytoplasmic ANCA-positive and 291 were perinuclear ANCA-positive. cANCA/PR3-ANCA were strongly associated with Wegener's granulomatosis (WG), whereas pANCA/MPO ANCA were associated with a diverse disease spectrum. Further investigation of PR3-ANCA-positive (n=80) and MPO-ANCA-positive patients (n=40) revealed a greater extent of disease [disease extent index (DEI); median 8 vs 5, P<0.01] and more frequent involvement of the upper/lower respiratory tract and the eyes in PR3 ANCA-positive than in MPO-ANCA-positive patients. Fewer than 5% of WG patients were MPO-ANCA-positive. Compared with matched PR3-ANCA-positive WG patients, the MPO-ANCA-positive WG patients had a lower DEI (median 5 vs 8) and had a lower frequency of peripheral neuropathy. CONCLUSIONS: ANCA testing is useful due to its high sensitivity and specificity, especially for cANCA/PR3-ANCA in WG. We found a divergence in the disease spectrum between PR3- and MPO-ANCA-positive patients, characterized by higher DEI and extrarenal manifestations in the PR3 ANCA group. MPO-ANCA was rarely found in WG and was associated with less organ involvement. PMID- 11257156 TI - Anticardiolipin antibodies in Behcet's disease: a reassessment. AB - OBJECTIVE: To assess the frequency and clinical relevance of anticardiolipin antibodies (aCL) in Behcet's disease (BD). METHODS: IgG, IgM and IgA aCL isotypes were investigated by ELISA in 128 patients with BD, 143 healthy controls and 20 systemic lupus erythematosus (SLE) patients. RESULTS: The IgA binding index (BI) was slightly elevated in BD compared with healthy controls (120+/-53 vs 107+/-46, P=0.02), whereas IgG and IgM aCL levels were not significantly different (IgG, BD 2.5+/-2.4 G phospholipid (GPL), healthy controls 2.8+/-3.6 GPL, P=0.6; IgM, BD 0.7+/-0.9 M phospholipid (MPL), healthy controls 0.9+/-1.3 MPL, P=0.6). The frequency of aCL positivity was 7% in BD (IgG 0.8%, IgM 1.6%, IgA 4.6%), 50% in SLE and 5.6% in healthy controls. IgA BI was elevated in the HLA-B5-negative group compared with HLA-B5-positive patients (P<0.005). In a literature review, the frequency of aCL was found to be 9.5% in studies from Turkey compared with 25.5% in other series (P<0.0001). CONCLUSION: These results do not suggest a primary role for aCL in BD. A significantly lower frequency of aCL in Turkish BD patients than in other series indicate that regional determinants, whether environmental or genetic, might also play a role in controlling aCL production in BD. PMID- 11257155 TI - Non-steroidal anti-oestrogens inhibit the differentiation of synovial macrophages into dendritic cells. AB - BACKGROUND: Dendritic cells (DC) have been suggested to play an important role in the pathogenesis of rheumatoid arthritis (RA). Agents that inhibit DC differentiation and function may have a therapeutic value in the treatment of RA. OBJECTIVE: To examine the effect of the non-steroidal anti-oestrogens toremifene and tamoxifen on the differentiation of synovial fluid (SF) macrophages into DC. METHODS: SF macrophages from patients with RA were cultured with interleukin (IL) 4 and granulocyte/macrophage colony-stimulating factor (GM-CSF) in the presence or absence of anti-oestrogens. The expression of cell surface markers on SF antigen-presenting cells (APC) was studied by flow cytometry. The capacity of SF APC to stimulate allogeneic T cells was studied using the mixed lymphocyte reaction. The production of tumour necrosis factor-alpha, IL-10 and transforming growth factor-beta1 was studied using ELISA. RESULTS: Anti-oestrogens inhibited the differentiation of SF macrophages into DC and the capacity of SF macrophage derived DC to stimulate allogeneic T cells. CONCLUSIONS: By inhibiting the differentiation of SF macrophages into DC, non-steroidal anti-oestrogens may have beneficial effects in RA. PMID- 11257157 TI - A high low-density lipoprotein cholesterol to high-density lipoprotein cholesterol ratio as a potential risk factor for corticosteroid-induced osteonecrosis in rabbits. AB - OBJECTIVE: This study was designed to determine the potential risk factors for corticosteroid-induced osteonecrosis (ON) based on lipid metabolism, using a rabbit ON model. METHODS: Blood samples were obtained from 38 rabbits, which then received a single intramuscular injection of 20 mg/kg methylprednisolone acetate. Four weeks after the injection, the femora and humeri were examined histopathologically for the presence of ON, and the sizes of the bone marrow fat cells were also measured. RESULTS: Rabbits with and without ON differed significantly in the ratio of low-density lipoprotein cholesterol to high-density lipoprotein cholesterol (LDL/HDL cholesterol ratio), which is considered to be a serological marker of lipid transport (P=0.026). The marrow fat cells were significantly larger in the rabbits with ON than in those without ON (P<0.0001). CONCLUSION: A higher LDL/HDL cholesterol ratio was significantly associated with the development of ON, and such an elevated ratio may partly contribute to the increased size of marrow fat cells. PMID- 11257158 TI - Pro-inflammatory interleukins in the synovial fluid of rheumatoid arthritis associated with joint hypermobility. AB - BACKGROUND: Joint hypermobility (JH) is frequently seen in rheumatology; in some cases, such as rheumatoid arthritis (RA), it may represent a worsening of disease evolution. The aim of our study was to evaluate the influence of joint hypermobility on RA synovial fluid (SF) inflammation. Patients and methods. One hundred consecutive adult patients with RA and joint effusion of the knee were examined for the presence of JH. In the SF we evaluated volume, the number of white blood cells (WBC) and the levels of interleukin (IL)-1beta, IL-6 and IL-8 and prostaglandin E2 (PGE2). RESULTS: JH was associated with RA (JH-RA) in 18 patients, all of whom were female. Compared with non-JH RA, all the SF indices found in JH-RA were higher, although significant differences were observed only for volume, IL-8 and PGE2. CONCLUSION: In JH-RA, increased joint mobility seems to be associated with a more severe local inflammatory response, which may contribute to the more erosive evolution observed in our patients. PMID- 11257160 TI - A reappraisal of amyloidosis in Behcet's syndrome. AB - OBJECTIVE: To evaluate the clinical features and outcome of patients with Behcet's syndrome (BS) and amyloidosis and to assess the associated risk factors. METHOD: A chart review was done to determine the frequency of amyloidosis in BS among 4000 patients. Data from 14 BS patients with amyloidosis were compared with data obtained from 718 patients with BS without amyloidosis. Multiple stepwise logistic regression analysis was used to assess the risk factors. RESULTS: All patients with amyloidosis presented with the nephrotic syndrome or significant proteinuria. The mean time to the onset of amyloidosis was 8.1 yr (range 3-15 yr). The mean duration of follow-up after amyloidosis was 3.4 yr (range 1-11 yr). Seven out of 14 patients were alive at the time of the evaluation. Peripheral and pulmonary arterial involvement and arthritis were associated with amyloidosis (P<0.05). CONCLUSION: Amyloidosis in BS is rare and has a 50% mortality rate at 3.4 yr (range 1-11 yr). Peripheral and pulmonary arterial involvement and arthritis seem to be the strongest predictors of amyloidosis in BS. PMID- 11257159 TI - Sustained improvement in rheumatoid arthritis following a protocol designed to deplete B lymphocytes. AB - OBJECTIVES: An open study of B-lymphocyte depletion was undertaken in rheumatoid arthritis (RA) patients to test the hypothesis that B lymphocytes may be essential to disease perpetuation. METHODS: Five patients with refractory RA were treated with a monoclonal anti-CD20 antibody, cyclophosphamide and prednisolone and followed for 12-17 months. Patient 2 received further treatments at 8 and 12 months and patient 4 at 11 months. RESULTS: At 26 weeks all patients satisfied the American College of Rheumatology ACR50 and patients 1-3 the ACR70 criteria of improvement, without further therapy. Patients 1, 3 and 5 achieved ACR70 at 1 yr and rheumatoid factor (RF) levels fell to normal. In patients 3 and 5, B lymphocytes returned without relapse. Patient 2 relapsed at 28 weeks and patient 4 at 38 weeks, coincident with the return of B lymphocytes in the presence of raised RF levels. Both achieved ACR70 on retreatment. Adverse events were limited to respiratory episodes (two patients) and marginal thrombocytopenia (one patient). CONCLUSIONS: These findings are consistent with the concept that RA is critically dependent on B lymphocytes and suggest that B-lymphocyte depletion may be a safe and effective therapy. PMID- 11257161 TI - Pulmonary involvement in juvenile dermatomyositis: a two-year longitudinal study. AB - OBJECTIVE: To investigate the prevalence and features of asymptomatic pulmonary involvement in juvenile dermatomyositis (JDM). METHODS: Twelve JDM patients underwent pulmonary function tests at baseline, 12 and 24 months. Disease activity, duration, serum lactate dehydrogenase (LDH) values and antinuclear antibody (ANA) titres were also evaluated. RESULTS: Five patients showed lung impairment at baseline and four at 12 and 24 months. Forced expiratory volume in 1s, forced vital capacity (FVC), carbon monoxide diffusing capacity (DLCO) and alveolar volume were the most frequently altered variables, indicating a restrictive pattern and impairment of diffusion. The prevalence and features of pulmonary alterations did not change during follow-up. FVC values were significantly lower in active JDM patients and were inversely related to LDH. DLCO values were significantly lower in ANA-positive patients. About half of the patients of this small case series of JDM had asymptomatic lung disease. CONCLUSIONS: We suggest that lung function should be evaluated at disease onset and regularly during follow-up, as pulmonary function tests can detect otherwise unpredictable pulmonary involvement. PMID- 11257162 TI - Uveitis in sibling pairs with juvenile idiopathic arthritis. AB - OBJECTIVE: To ascertain the occurrence and characteristics of uveitis in sibling pairs affected with juvenile idiopathic arthritis (JIA). METHODS: The sibling series comprised 80 JIA patients from 37 families with two or three JIA children, seen at the paediatric department of the Rheumatism Foundation Hospital in Heinola, Finland. An ophthalmologist examined the children for uveitis two to four times a year and the course of the condition was recorded during the follow up. RESULTS: Uveitis was diagnosed in 21 of the 80 patients (26%). Three pairs (3.4 pairs expected) were concordant for the presence of asymptomatic uveitis. Two patients with enthesitis-related arthritis had acute unilateral uveitis. Among the remaining cases, uveitis was chronic and continuously active at the end of follow-up in 13 instances, but in spite of this only one patient had impaired vision. HLA allele B27 occurred more frequently in patients with uveitis than in those without uveitis (52 vs 30%, P=0.073) and all six subjects in the pairs concordant for chronic uveitis carried this allele. CONCLUSIONS: The observed concordance rate for uveitis did not differ from that expected. Although the uveitis was chronic in most instances, its course was usually mild. PMID- 11257163 TI - Outcome status in children with sustained polyarticular and systemic juvenile idiopathic arthritis. PMID- 11257164 TI - Familial erosive arthritis associated with seroreactivity to human intracisternal retroviral particle type I (HIAP-I). PMID- 11257165 TI - Survival of Mexican patients with systemic lupus erythematosus. PMID- 11257166 TI - Histopathological assessment and pathological significance of matrix degeneration in supraspinatus tendons. PMID- 11257167 TI - Methotrexate misadventure: a case for counselling. PMID- 11257168 TI - Spontaneous hydropneumothorax in a man with rheumatoid arthritis. PMID- 11257169 TI - Microscopic polyangiitis in a patient with relapsing polychondritis. PMID- 11257170 TI - Three-dimensional computed tomography for visualization of carotid bypasses in Takayasu arteritis. PMID- 11257171 TI - Control of unremitting rheumatoid arthritis by the prolactin antagonist cabergoline. PMID- 11257172 TI - Fibromyalgia... I mistake your shape. PMID- 11257173 TI - Robin goodfellow. PMID- 11257174 TI - The origin and evolution of hepatitis viruses in humans. AB - The spread and origins of hepatitis C virus (HCV) in human populations have been the subject of extensive investigations, not least because of the importance this information would provide in predicting clinical outcomes and controlling spread of HCV in the future. However, in the absence of historical and archaeological records of infection, the evolution of HCV and other human hepatitis viruses can only be inferred indirectly from their epidemiology and by genetic analysis of contemporary virus populations. Some information on the history of the latter may be obtained by dating the time of divergence of various genotypes of HCV, hepatitis B virus (HBV) and the non-pathogenic hepatitis G virus (HGV)/GB virus-C (GBV-C). However, the relatively recent times predicted for the origin of these viruses fit poorly with their epidemiological distributions and the recent evidence for species-associated variants of HBV and HGV/GBV-C in a wide range of non-human primates. The apparent conservatism of viruses over long periods implied by these latter observations may be the result of constraints on sequence change peculiar to viruses with single-stranded genomes, or with overlapping reading frames. Large population sizes and intense selection pressures that optimize fitness may be the factors that set virus evolution apart from that of their hosts. PMID- 11257175 TI - Evolutionarily conserved RNA secondary structures in coding and non-coding sequences at the 3' end of the hepatitis G virus/GB-virus C genome. AB - Hepatitis G virus (HGV)/GB virus C (GBV-C) causes persistent, non-pathogenic infection in a large proportion of the human population. Epidemiological and genetic evidence indicates a long-term association between HGV/GBV-C and related viruses and a range of primate species, and the co-speciation of these viruses with their hosts during primate evolution. Using a combination of covariance scanning and analysis of variability at synonymous sites, we previously demonstrated that the coding regions of HGV/GBV-C may contain extensive secondary structure of undefined function (Simmonds & Smith, Journal of Virology 73, 5787 5794, 1999 ). In this study we have carried out a detailed comparison of the structure of the 3'untranslated region (3'UTR) of HGV/GBV-C with that of the upstream NS5B coding sequence. By investigation of free energies on folding, secondary structure predictive algorithms and analysis of covariance between HGV/GBV-C genotypes 1-4 and the more distantly related HGV/GBV-C chimpanzee variant, we obtained evidence for extensive RNA secondary structure formation in both regions. In particular, the NS5B region contained long stem-loop structures of up to 38 internally paired nucleotides which were evolutionarily conserved between human and chimpanzee HGV/GBV-C variants. The prediction of similar structures in the same region of hepatitis C virus may allow the functions of these structures to be determined with a more tractable experimental model. PMID- 11257176 TI - Interferon-alpha inhibits hepatitis C virus subgenomic RNA replication by an MxA independent pathway. AB - Hepatitis C virus (HCV) persists in the majority of infected individuals and is a major cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. Chronic hepatitis C is currently treated with interferon (IFN)-alpha or with a combination of IFN-alpha and ribavirin. The availability of an HCV replicon system (Lohmann et al., SCIENCE: 285, 110-113, 1999) allowed the investigation of the effects of IFN on genuine HCV replication in cultured cells. It is shown here that IFN-alpha inhibits subgenomic HCV RNA replication in HuH-7 human hepatoma cells. Immunofluorescence, Western blot and Northern blot analysis revealed that levels of both HCV protein and replicon RNA were reduced after treatment with IFN alpha in a dose-dependent manner. In further experiments, it was investigated whether MxA plays a role in the inhibition of HCV. The human MxA protein is an IFN-induced GTPase that has antiviral activity against various RNA viruses. However, HCV RNA replication was not affected in transfected HuH-7 cells that transiently overexpressed MxA. Moreover, a dominant-negative mutant of MxA did not interfere with the antiviral activity of IFN-alpha against HCV RNA replication. Taken together, these results demonstrate that IFN-alpha inhibits HCV replicons via an MxA-independent pathway. PMID- 11257177 TI - A small region of the dengue virus-encoded RNA-dependent RNA polymerase, NS5, confers interaction with both the nuclear transport receptor importin-beta and the viral helicase, NS3. AB - The dengue virus RNA-dependent RNA polymerase, NS5, and the protease/helicase, NS3, are multidomain proteins that have been shown to interact both in vivo and in vitro. A hyperphosphorylated form of NS5 that does not interact with NS3 has been detected in the nuclei of virus-infected cells, presumably as the result of the action of a functional nuclear localization sequence within the interdomain region of NS5 (residues 369-405). In this study, it is shown by using the yeast two-hybrid system that the C-terminal region of NS3 (residues 303-618) interacts with the N-terminal region of NS5 (residues 320-368). Further, it is shown that this same region of NS5 is also recognized by the cellular nuclear import receptor importin-beta. The interaction between NS5 and importin-beta and competition by NS3 with the latter for the same binding site on NS5 were confirmed by pull-down assays. The direct interaction of importin-beta with NS5 has implications for the mechanism by which this normally cytoplasmic protein may be targetted to the nucleus. PMID- 11257178 TI - The early pathogenesis of foot-and-mouth disease in pigs infected by contact: a quantitative time-course study using TaqMan RT-PCR. AB - Foot-and-mouth disease (FMD) is a highly contagious, economically important virus disease of cloven-hoofed animals. The objective of the present study was to examine the early pathogenesis of FMD in pigs by a quantitative time-course study. Under experimental conditions, recipient pigs were infected by contact with donor pigs affected by FMD. Every 24 h from day 1 to day 4 after exposure, two recipient pigs were selected randomly, killed and necropsied. A range of tissues were analysed by a quantitative TaqMan RT-PCR method and by titration of FMD virus on primary bovine thyroid cells. The titres of virus determined by assay in cell culture and calculated from the quantitative TaqMan data correlated strongly (r>0.9), thereby establishing the validity of the TaqMan calculations. The data indicated that the replication of virus in the lungs contributes only in small part to airborne virus excretion. Sites in the pharynx, trachea and nasal mucosa are probably more important in that regard. The sites of earliest virus infection and possibly replication in recipient pigs appeared to be in the pharynx (soft palate, tonsil and floor of pharynx). The data indicated that FMD virus replication in pigs is rapid and that the majority of virus amplification occurs in the skin. A model for the progression of infection is proposed, indicating initial spread from the pharyngeal region, through regional lymph nodes and via the blood to epithelial cells, resulting in several cycles of virus amplification and spread. PMID- 11257179 TI - Functional interaction of translation initiation factor eIF4G with the foot-and mouth disease virus internal ribosome entry site. AB - In the life-cycle of picornaviruses, the synthesis of the viral polyprotein is initiated cap-independently at the internal ribosome entry site (IRES) far downstream from the 5' end of the viral plus-strand RNA. The cis-acting IRES RNA elements serve as binding sites for translation initiation factors that guide the ribosomes to an internal site of the viral RNA. In this study, we show that the eukaryotic translation initiation factor eIF4G interacts directly with the IRES of foot-and-mouth disease virus (FMDV). eIF4G binds mainly to the large Y-shaped stem-loop 4 RNA structure in the 3' region of the FMDV IRES element, whereas stem loop 5 contributes only slightly to eIF4G binding. Two subdomains of stem-loop 4 are absolutely essential for eIF4G binding, whereas another subdomain contributes to a lesser extent to binding of eIF4G. At the functional level, the translational activity of stem-loop 4 subdomain mutants correlates with the efficiency of binding of eIF4G in the UV cross-link assay. This indicates that the interaction of eIF4G with the IRES is crucial for the initiation of FMDV translation. A model for the interaction of initiation factors with the IRES element is discussed. PMID- 11257180 TI - Proteolytic processing of Semliki Forest virus-specific non-structural polyprotein by nsP2 protease. AB - The RNA replicase proteins of Semliki Forest virus (SFV) are translated as a P1234 polyprotein precursor that contains two putative autoproteases. Point mutations introduced into the predicted active sites of both proteases nsP2 (P2) and nsP4 (P4), separately or in combination, completely abolished virus replication in mammalian cells. The effects of these mutations on polyprotein processing were studied by in vitro translation and by expression of wild-type polyproteins P1234, P123, P23, P34 and their mutated counterparts in insect cells using recombinant baculoviruses. A mutation in the catalytic site of the P2 protease, C(478)A, (P2(CA)) completely abolished the processing of P12(CA)34, P12(CA)3 and P2(CA)3. Co-expression of P23 and P12(CA)34 in insect cells resulted in in trans cleavages at the P2/3 and P3/4 sites. Co-expression of P23 and P34 resulted in cleavage at the P3/4 site. In contrast, a construct with a mutation in the active site of the putative P4 protease, D(6)A, (P1234(DA)) was processed like the wild-type protein. P34 or its truncated forms were not processed when expressed alone. In insect cells, P4 was rapidly destroyed unless an inhibitor of proteosomal degradation was used. It is concluded that P2 is the only protease needed for the processing of SFV polyprotein P1234. Analysis of the cleavage products revealed that P23 or P2 could not cleave the P1/2 site in trans. PMID- 11257181 TI - Mapping the domains on the phosphoprotein of bovine respiratory syncytial virus required for N-P and P-L interactions using a minigenome system. AB - The interaction of bovine respiratory syncytial virus (BRSV) phosphoprotein (P) with nucleocapsid (N) and large polymerase (L) proteins was investigated using an intracellular BRSV-CAT minigenome replication system. Coimmunoprecipitation assays using P-specific antiserum revealed that the P protein can form complexes with N and L proteins. Deletion mutant analysis of the P protein was performed to identify the regions of P protein that interact with N and L proteins. The results indicate that two independent N-binding sites exist on the P protein: an internal region of 161-180 amino acids and a C-terminal region of 221-241 amino acids. The L-binding site was mapped to a region of P protein encompassing amino acids 121-160. The data suggest that N and L protein binding domains on the P protein do not overlap. PMID- 11257182 TI - Activity of Toscana and Rift Valley fever virus transcription complexes on heterologous templates. AB - A transcription system for Toscana virus (TOSV) (a member of the family BUNYAVIRIDAE:, genus PHLEBOVIRUS:) was constructed. For in vivo expression, the TOSV transcription system uses the viral N and L proteins and an S-like RNA genome containing the chloramphenicol acetyltransferase reporter gene in the antisense orientation flanked by the viral genomic 5'- and 3'-terminal S sequences. It was found that the N and L proteins represent the minimal protein requirement for an active transcription complex. To investigate the possibility of reassortment between TOSV and Rift Valley fever virus (RVFV), the activity of their polymerase complexes was tested on their heterologous S-like RNA genomes and this showed that both virus complexes were active. Moreover, hybrid transcriptase complexes with protein components originating from the two viruses were tested on both virus templates and only the combination RVFV L + TOSV N on RVFV S-like RNA was found to be active in this assay. These results suggest that virus reassortants might be generated whenever the two viruses infect the same host. PMID- 11257183 TI - Infection kinetics, prostacyclin release and cytokine-mediated modulation of the mechanism of cell death during bluetongue virus infection of cultured ovine and bovine pulmonary artery and lung microvascular endothelial cells. AB - Bluetongue virus (BTV) infection causes a haemorrhagic disease in sheep, whereas BTV infection typically is asymptomatic in cattle. Injury to the endothelium of small blood vessels is responsible for the manifestations of disease in BTV infected sheep. The lungs are central to the pathogenesis of BTV infection of ruminants; thus endothelial cells (ECs) cultured from the pulmonary artery and lung microvasculature of sheep and cattle were used to investigate the basis for the disparate expression of bluetongue disease in the two species. Ovine and bovine microvascular ECs infected at low multiplicity with partially purified BTV were equally susceptible to BTV-induced cell death, yet ovine microvascular ECs had a lower incidence of infection and produced significantly less virus than did bovine microvascular ECs. Importantly, the relative proportions of apoptotic and necrotic cells were significantly different in BTV-infected EC cultures depending on the species of EC origin and the presence of inflammatory mediators in the virus inoculum. Furthermore, BTV-infected ovine lung microvascular ECs released markedly less prostacyclin than the other types of ECs. Results of these in vitro studies are consistent with the marked pulmonary oedema and microvascular thrombosis that characterize bluetongue disease of sheep but which rarely, if ever, occur in BTV-infected cattle. PMID- 11257184 TI - Complete sequence characterization of the genome of the St Croix River virus, a new orbivirus isolated from cells of Ixodes scapularis. AB - An orbivirus identified as St Croix River virus (SCRV) was isolated from cells of Ixodes scapularis ticks. Electron microscopy showed particles with typical orbivirus morphology. The SCRV genome was sequenced completely and compared to previously characterized orbivirus genomes. Significant identity scores (21-38%) were detected between proteins encoded by segments S1, S2, S4, S5, S6, S8, S9 and S10 of SCRV and those encoded by segments S1, S3, S4, S5, S6, S7, S9 and S10, respectively, of Bluetongue virus (BTV), the prototype orbivirus species. The protein encoded by SCRV genome segment 3 (VP3) is thought to be the equivalent of VP2 of BTV. Segment 7 encodes a protein homologous to non-structural protein NS2(ViP) of BTV. Analysis of VP1(Pol) (segment 1) shows that SCRV is an orbivirus, distantly related to the other sequenced species. Blot hybridizations and sequence comparisons of the conserved protein encoded by genome segment 2 (the T2 subcore shell protein) with previously identified orbiviruses confirm that SCRV is a distinct orbivirus species, unrelated to another tick-borne species, Great Island virus. The presence of SCRV in cells prepared from tick eggs suggests that transovarial transmission of SCRV may occur in ticks. PMID- 11257185 TI - Expression of VP5 of infectious pancreatic necrosis virus strain VR299 is initiated at the second in-frame start codon. AB - Infectious pancreatic necrosis virus (IPNV), a member of the BIRNAVIRIDAE: with two double-stranded RNA genome segments, encodes five proteins designated VP1 to VP5. To study the function of the 17 kDa nonstructural protein VP5 during virus replication several mutated IPNV genome segments A were constructed and included in a reverse genetics system for IPNV to obtain recombinant virus. Mutations between nt 68 and 85 or nt 94 and 103 in the noncoding region failed to yield viable virus. Only mutations located between nt 86 and 92 and downstream of nt 104 were tolerated, and viable virus could be generated. All IPNV generated showed no difference in replication compared with the wild-type IPNV, indicating that the absence of expression of VP5 did not influence virus growth in vitro. Furthermore, the results presented here indicate that initiation of translation of VP5 occurs at position 113, the second in-frame start codon. PMID- 11257186 TI - An immunodominant neutralization epitope on the 'thumb' subdomain of human immunodeficiency virus type 1 reverse transcriptase revealed by phage display antibodies. AB - An antibody phage display library was produced from the splenocytes of mice immunized with an infectious vaccinia virus recombinant (WRRT) expressing the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1). The library was panned against HIV-1 RT. Two clones, 5F and 5G, which produced Fab fragments specific for RT, were isolated. Surprisingly, both 5F and 5G Fab fragments were capable of strongly inhibiting the RNA-dependent DNA polymerase activity of HIV-1 RT. A hybridoma cell line that produces the monoclonal antibody 7C4, which strongly inhibits RT activity, was established previously using splenocytes from mice immunized with WRRT by the same immunization protocol. The epitope recognized by 7C4 exists in the region of the template primer-binding sites (or the 'helix clump') of RT. By epitope mapping and competitive ELISA analysis, it was shown that the 5F and 5G Fab fragments were directed against the same, or a very closely related, epitope that is recognized by 7C4. The neutralizing activities of the 5F, 5G and 7C4 Fab fragments correlated with their affinities for HIV-1 RT. DNA sequencing indicated that the immunoglobulin genes of the heavy chains of 5G and 7C4, as well as those of the light chains of 5F and 5G, had the same origin. These results suggest that the neutralizing epitope, which is recognized by these antibodies, becomes immunodominant after repeated immunization of mice with WRRT. This unique epitope, HIV-1 RT-specific and immunodominant neutralizing epitope (HRSINE), is a logical target for new types of HIV-1 RT inhibitors and gene therapy. PMID- 11257187 TI - Analysis of the molecules involved in human T-cell leukaemia virus type 1 entry by a vesicular stomatitis virus pseudotype bearing its envelope glycoproteins. AB - Cellular entry of human T-cell leukaemia virus type 1 (HTLV-1) was studied by a quantitative assay system using vesicular stomatitis virus (VSV) pseudotypes in which a recombinant VSV (VSVDeltaG*) containing the gene for green fluorescent protein instead of the VSV G protein gene was complemented with viral envelope glycoproteins in trans. Most of the cell lines tested showed susceptibility to VSVDeltaG* complemented with either HTLV-1 envelope glycoproteins (VSVDeltaG* Env) or VSV G protein (VSVDeltaG*-G), but not to VSVDeltaG* alone, indicating that cell-free HTLV-1 could infect many cell types from several species. High concentration pronase treatment of cells reduced their susceptibility to VSVDeltaG*-Env, while trypsin treatment, apparently, did not. Treatment of the cells with sodium periodate, heparinase, heparitinase, phospholipase A2 or phospholipase C reduced the susceptibility of cells to VSVDeltaG*-Env, but not to VSVDeltaG* complemented with measles virus (Edmonston strain) H and F proteins (VSVDeltaG*-EdHF), which was used as a control. Purified phosphatidylcholine also inhibited the infectivity of VSVDeltaG*-Env, but not VSVDeltaG*-G. These findings indicated that, in addition to cell surface proteins, glycosaminoglycans and phospholipids play an important role in the process of cell-free HTLV-1 entry. PMID- 11257188 TI - Vascular cell adhesion molecule-1 induced by human T-cell leukaemia virus type 1 Tax protein in T-cells stimulates proliferation of human T-lymphocytes. AB - Human T-cell leukaemia/lymphotropic virus type 1 (HTLV-1), aetiologically linked to lymphoproliferative as well as inflammatory diseases, infects and activates CD4(+) helper T-cells and thus alters immunoregulatory pathways. The viral regulatory Tax protein has been shown previously to induce the expression of vascular cell adhesion molecule-1 (VCAM-1) by T-cells. To determine the functional role of this adhesion molecule, Jurkat T-cells stably expressing either Tax or both Tax and Rex (another viral regulatory protein) were used in binding and coculture assays performed with either control Jurkat cells or primary human T-lymphocytes. Evidence was provided that VCAM-1 acting in synergy with leucocyte function-associated antigen-3 promotes T-cell-T-cell interactions and increases T-cell proliferation. Interestingly, Rex was found to modulate these events. These data establish that VCAM-1 induced by Tax on T-cells thus contributes to the immunopathological process triggered by HTLV-1 infection. PMID- 11257189 TI - Porcine endogenous retroviruses: in vitro host range and attempts to establish small animal models. AB - Using transgenic pigs as the source of cells or organs for xenotransplantation is associated with the risk of porcine endogenous retrovirus (PERV) transmission. Multiple proviruses are integrated into the genome of all pigs, and virus particles, some of which are able to infect human cells, are released from normal pig cells. In order to evaluate the potential risk posed by the transmission of PERVs, in vitro infection studies were performed as a basis for small animal as well as non-human primate models. In vitro infectivity was demonstrated for permanent cell lines and primary cells from a wide range of species. Productive infection was shown using reverse transcriptase (RT) assays and RT-PCR for mink, feline and human kidney cell lines, primary rhesus peripheral blood mononuclear cells (PBMCs), and baboon spleen cells and PBMCs as well as for different human lymphoid and monocyte cell lines and PBMCs. In an attempt to establish a small animal model, naive guinea pigs, non-immunosuppressed rats, rats immunosuppressed by cyclosporin-A and immunosuppressed rats treated with cobra venom factor were inoculated with PERVs produced from porcine kidney PK-15 cells, infected human 293 kidney cells and mitogen-stimulated porcine PBMCs. Animals were also inoculated with PERV-producing PK-15 and 293 cells. No antibodies against PERV and no provirus integration were observed in any of the treated animals. This suggests that productive infection of these animals did not occur in this experimental setting. PMID- 11257190 TI - Differential roles of B cells and IFN-gamma-secreting CD4(+) T cells in innate and adaptive immune control of genital herpes simplex virus type 2 infection in mice. AB - The role of B, CD4(+) T and CD8(+) T cells in both primary genital infection with attenuated herpes simplex virus type 2 (HSV-2) and development of protective immunity to a later challenge with virulent HSV-2 using lymphocyte-deficient mice has been elucidated. Following primary inoculation with attenuated thymidine kinase-deficient (TK(-)) HSV-2, B cell-deficient (microMT) mice developed a local viraemia and transient genital inflammation, suggesting a role for B cells in the innate control of local infection and inflammation. Natural antibodies are implicated in this process, as passive transfer of normal serum into microMT mice significantly reduced HSV-2 TK(-) shedding in the vaginal lumen, although it did not affect subsequent inflammation. Protection against lethal HSV-2 challenge was noted in HSV-2-vaccinated wild-type, CD8(+) T cell-deficient and microMT mice and was characterized by strong virus-specific IFN-gamma responses in vitro and delayed type hypersensitivity (DTH) responses in vivo. In contrast, CD4(+) T cell deficient (CD4(-/-)) mice had impaired HSV-2-specific IFN-gamma production and DTH responses and succumbed rapidly to genital HSV-2 challenge. However, protective responses to HSV-2 could be induced in HSV-2-vaccinated CD4(-/-) mice by treatment with recombinant IFN-gamma. Taken together, these results suggest that CD4(+) T cells secreting IFN-gamma are critical for immune protection against lethal genital HSV-2 re-infection, whereas B cells/natural antibodies have anti-viral and -inflammatory effects in the innate control of a primary infection. PMID- 11257191 TI - Human telomerase reverse transcriptase-immortalized MRC-5 and HCA2 human fibroblasts are fully permissive for human cytomegalovirus. AB - MRC-5 cells are a well-characterized human diploid fibroblast cell line approved for vaccine production and favoured for the routine propagation of human cytomegalovirus (HCMV). Ectopic expression of telomerase in fibroblasts is capable of overcoming replicative senescence induced by telomere shortening. Following delivery of the hTERT gene to MRC-5 cells using a retrovirus vector three clones were generated that (i) expressed functional telomerase activity, (ii) exhibited telomere extension and (iii) were sustained for >100 population doublings. Immortalized MRC-5-hTERT and also HCA2-hTERT human fibroblasts were both fully permissive for HCMV as determined by plaque assay, studies of virus growth kinetics and measurement of virus yields. Furthermore, telomerase immortalized HCA2 cells proved capable of supporting the stable maintenance of an EBV-based episomal vector with efficient transgene expression when driven by the HCMV immediate early promoter. An indicator cell line suitable for the efficient detection of HCMV infection was also generated using an episome containing a reporter gene (lacZ) under the control of the HCMV beta-2.7 early promoter. Telomerase immortalization of human fibroblasts will thus facilitate the growth and detection of HCMV and also the generation of helper cell lines for the propagation of HCMV deletion mutants. Immortalization of fibroblasts by telomerase does not affect cell morphology or growth characteristics. The MRC-5 hTERT clones may therefore be suitable for additional applications in virology, cell biology, vaccine production and biotechnology. PMID- 11257192 TI - Evidence for interspecies transmission of oyster herpesvirus in marine bivalves. AB - Since 1991, numerous herpesvirus infections associated with high mortality have been reported around the world in various marine bivalve species. In order to determine whether these infections are due to ostreid herpesvirus-1 (OsHV1), a previously characterized pathogen of the Japanese oyster (Crassostrea gigas), PCR analysis was carried out on 30 samples of larvae collected from four bivalve species (C. gigas, Ostrea edulis, Ruditapes decussatus and Ruditapes philippinarum), most exhibiting mortality prior to collection. All samples were shown to be infected by OsHV1. Viral genomes in three samples of C. gigas and three of R. philippinarum that originated from the same hatchery were unusual in bearing a deletion of at least 2.8 kbp in an inverted repeat region. The results demonstrate that OsHV1 is capable of infecting several bivalve species, and this raises the possibility that interspecies transmission may be promoted by intensive rearing in modern hatcheries. PMID- 11257193 TI - Intracellular localization of the hepatitis B virus HBx protein. AB - The hepatitis B virus (HBV) X protein (HBx) was originally suggested to be a viral transcriptional activator, but its functional mechanisms are still unclear. In this study we have analysed the intracellular localization of HBx in transfected cells and demonstrate that its compartmentalization is dependent on overall expression levels. HBx was exclusively or predominantly localized in the nuclei in weakly expressing cells. However, elevated cellular levels correlated with its accumulation in the cytoplasm, suggesting that the capacity of HBx for nuclear compartmentalization might be limited. Cytoplasmic HBx was detected either as punctate granular staining or in dispersed, finely granular patterns. We have further analysed the detailed cytoplasmic compartmentalization, using confocal microscopy, and show no association with the endoplasmic reticulum, plasma membrane or lysosomes, but a substantial association of HBx with mitochondria. However, a major fraction of cytoplasmic HBx did not localize in mitochondria, indicating the presence of two distinctly compartmentalized cytoplasmic populations. Furthermore, high levels of HBx expression led to an abnormal mitochondrial distribution, involving clumping and organelle aggregation, which was not observed at lower expression levels. The data presented here provide novel insights into the compartmentalization of HBx and may prove important for future evaluations of its functions, both in the viral life-cycle and in the pathology of HBV-related liver disease. PMID- 11257195 TI - Analysis of two genomic variants of orang-utan hepadnavirus and their relationship to other primate hepatitis B-like viruses. AB - We recently described orang-utan hepadnavirus (OuHV) (Warren et al., Journal of Virology, 73, 7860-7865, 1999). Phylogenetic analyses indicated that the various isolates of OuHV can be divided into two genomic variants. Two representatives from each genomic cluster were analysed both molecularly and phylogenetically. Their genome organization was highly similar to other hepadnaviruses of apes and humans. The complete genome sequences of the two OuHV types had an overall 5% sequence difference. Research on 25 seropositive Bornean orang-utans showed that, of the 19 animals infected with one variant, 12 originated from East Kalimantan. Phylogenetic analysis was performed using the full-length genomes of various primate hepadnaviruses. The tree topology revealed one cluster of Old World hepadnaviruses that is divided into two subclusters, one consisting of the ape viruses, and the other comprising the human genotypes A-E. These data suggest that the great apes and gibbons have been infected with a common ancestor hepadnavirus. PMID- 11257194 TI - A novel variant genotype C of hepatitis B virus identified in isolates from Australian Aborigines: complete genome sequence and phylogenetic relatedness. AB - There have been no reports of DNA sequences of hepatitis B virus (HBV) strains from Australian Aborigines, although the hepatitis B surface antigen (HBsAg) was discovered among them. To investigate the characteristics of DNA sequences of HBV strains from Australian Aborigines, the complete nucleotide sequences of HBV strains were determined and subjected to molecular evolutionary analysis. Serum samples positive for HBsAg were collected from five Australian Aborigines. Phylogenetic analysis of the five complete nucleotide sequences compared with DNA sequences of 54 global HBV isolates from international databases revealed that three of the five were classified into genotype D and were most closely related in terms of evolutionary distance to a strain isolated from a healthy blood donor in Papua New Guinea. Two of the five were classified into a novel variant genotype C, which has not been reported previously, and were closely related to a strain isolated from Polynesians, particularly in the X and Core genes. These two strains of variant genotype C differed from known genotype C strains by 5.9-7.4% over the complete nucleotide sequence and 4.0-5.6% in the small-S gene, and had residues Arg(122), Thr(127) and Lys(160), characteristic of serotype ayw3, which have not been reported previously in genotype C. In conclusion, this is the first report of the characteristics of complete nucleotide sequences of HBV from Australian Aborigines. These results contribute to the investigation of the worldwide spread of HBV, the relationship between serotype and genotype and the ancient common origin of Australian Aborigines. PMID- 11257196 TI - Progressive multifocal leukoencephalopathy in human immunodeficiency virus type 1 infected patients: absence of correlation between JC virus neurovirulence and polymorphisms in the transcriptional control region and the major capsid protein loci. AB - Progressive multifocal leukoencephalopathy (PML) is a rapidly fatal demyelinating disease of the central nervous system related to JC polyomavirus (JCV) replication in oligodendrocytes. PML usually occurs in immunocompromised individuals, especially in the setting of AIDS. Administration of highly active anti-retroviral therapy (HAART) may improve survival prognosis in some, but not all, patients with AIDS-related PML. This observation might be explained by the outgrowth of some JCV variants of increased fitness. To evaluate this hypothesis, two subgroups of five patients with AIDS-related PML, started on HAART after PML diagnosis, were analysed. The non-responder (NR) patients died rapidly despite HAART, while responders (R) had a positive outcome and were still alive. JCV DNA was extracted from cerebrospinal fluid biopsies and two regions of the genome were analysed, the transcriptional control region (TCR) and the major capsid protein gene (VP1). Both regions show different degrees of polymorphism and are recognized as evolving independently. Sequence analysis demonstrated that (i) extensive TCR rearrangements were present in both subgroups of patients, (ii) VP1 sequence polymorphisms could be identified in the BC loop, suggesting the absence of immune selection, and (iii) no genomic marker for JCV specific neurovirulence could be identified in the TCR and VP1 loci. PMID- 11257197 TI - Avian polyomavirus agnoprotein 1a is incorporated into the virus particle as a fourth structural protein, VP4. AB - Agnoproteins, encoded by the 5'-region of the late bicistronic mRNA of some polyomaviruses, are small proteins with largely unknown functions. In avian polyomavirus (APV)-infected cells, mRNAs of seven putative agnoproteins have been observed. Recently, it has been shown that agnoprotein 1a and its truncated variant agnoprotein 1b, encoded by the predominant mRNA species, are essential for APV replication. Here, the presence of agnoprotein 1a is demonstrated in the nucleus of APV-infected cells and in purified APV particles. Interaction between agnoprotein 1a and the major structural protein, VP1, was demonstrated by co immunoprecipitation experiments using lysates of recombinant baculovirus-infected insect cells. With proteins expressed in E. coli, binding to double-stranded DNA in a sequence-unspecific manner was shown for agnoprotein 1a, whereas agnoprotein 1b failed to bind. A leucine zipper-like motif present in agnoprotein 1a is considered to be involved in DNA binding. Due to the absence of any structural or functional homologies between APV agnoprotein 1a and the agnoproteins of mammalian polyomaviruses, it is suggested that this protein should be renamed VP4, indicating its function as a fourth structural protein of APV. PMID- 11257198 TI - Construction and initial characterization of an infectious plasmid clone of a newly identified hamster parvovirus. AB - The construction and characterization of a full-length infectious plasmid clone of the newly identified hamster parvovirus (HaPV) are described. Following transfection of hamster BHK cells with the infectious clone, pHaPV, the specific intracellular DNA replicative forms, RNA transcripts and viral proteins that were expected for this rodent parvovirus were generated. Infected cells were lysed and progeny virus was produced, demonstrating that pHaPV could generate a productive virus infection. The complete sequences of both hairpin termini, which had not been previously determined, were obtained. Preliminary host-range studies, which compared virus production and macromolecular synthesis in various cell lines following either HaPV infection or pHaPV transfection, demonstrated an early block of infection of HaPV in both monkey COS-1 and murine A9 cells. The availability of an HaPV infectious clone will facilitate its genetic analysis and allow the elucidation of the determinants important in host range, tissue tropism and pathogenicity of this newly identified rodent parvovirus. PMID- 11257200 TI - Towards a protein interaction map of potyviruses: protein interaction matrixes of two potyviruses based on the yeast two-hybrid system. AB - A map for the interactions of the major proteins from Potato virus A (PVA) and Pea seed-borne mosaic virus (PSbMV) (members of the genus POTYVIRUS:, family POTYVIRIDAE:) was generated using the yeast two-hybrid system (YTHS). Interactions were readily detected with five PVA protein combinations (HC-HC, HC CI, VPg-VPg, NIa-NIb and CP-CP) and weak but reproducible interactions were detected for seven additional combinations (P1-CI, P3-NIb, NIaPro-NIb, VPg-NIa, VPg-NIaPro, NIaPro-NIa and NIa-NIa). In PSbMV, readily detectable interactions were found in five protein combinations (HC-HC, VPg-VPg, VPg-NIa, NIa-NIa and NIa NIb) and weaker but reproducible interactions were detected for three additional combinations (P3-NIa, NIa-NIaPro and CP-CP). The self-interactions of HC, VPg, NIa and CP and the interactions of VPg-NIa, NIa-NIaPro and NIa-NIb were, therefore, common for the two potyviruses. The multiple protein interactions revealed in this study shed light on the co-ordinated functions of potyviral proteins involved in virus movement and replication. PMID- 11257199 TI - Geographically distant isolates of the crinivirus Cucurbit yellow stunting disorder virus show very low genetic diversity in the coat protein gene. AB - The population structure and genetic variation of Cucurbit yellow stunting disorder virus (CYSDV) isolates were estimated by single-strand conformation polymorphism and nucleotide sequence analyses of the CYSDV coat protein gene. Analysis of 71 isolates collected from Spain, Jordan, Turkey, Lebanon, Saudi Arabia and North America showed that, from a genetic viewpoint, these isolates could be divided into two diverged subpopulations: an Eastern subpopulation composed of Saudi Arabian isolates and a Western subpopulation containing the rest of the CYSDV isolates. The genetic variation within the Western subpopulation was very small (nucleotide identity >99%) in spite of the extensive and discontinuous geographical distribution and different years of collection. We also estimated the within-isolate genetic structure and variation of three CYSDV isolates by analysing 30 clones per isolate. Our results showed that these CYSDV isolates had a quasispecies structure. PMID- 11257201 TI - A subpopulation of RNA 1 of Cucumber mosaic virus contains 3' termini originating from RNAs 2 or 3. AB - Tobacco plants transgenic for RNA 1 of Cucumber mosaic virus and inoculated with transcript of RNAs 2 and 3 regenerated viral RNA 1 from the transgenic mRNA, and the plants became systemically infected by the reconstituted virus. cDNA fragments corresponding to the 3' non-coding region (NCR) of viral RNA 1 were amplified, cloned and sequenced. In some clones the termini of the 3' NCR corresponded to those of viral RNAs 2 or 3. This suggested that in some cases RNA 1 may have been regenerated during replication by a template switching mechanism between the inoculated transcript RNAs and the mRNA. However, encapsidated, recombinant RNA 1 with the 3' NCR ends originating from RNAs 2 or 3 also was found in virus samples that had been passaged exclusively through non-transgenic plants. Thus, these chimeras occur naturally due to recombination between wild type viral RNAs, and they are found encapsidated in low, but detectable amounts. PMID- 11257202 TI - Recognition of cis-acting sequences in RNA 3 of Prunus necrotic ringspot virus by the replicase of Alfalfa mosaic virus. AB - Alfalfa mosaic virus (AMV) and Prunus necrotic ringspot virus (PNRSV) belong to the genera ALFAMOVIRUS: and ILARVIRUS:, respectively, of the family BROMOVIRIDAE: Initiation of infection by AMV and PNRSV requires binding of a few molecules of coat protein (CP) to the 3' termini of the inoculum RNAs and the CPs of the two viruses are interchangeable in this early step of the replication cycle. CIS: acting sequences in PNRSV RNA 3 that are recognized by the AMV replicase were studied in in vitro replicase assays and by inoculation of AMV-PNRSV RNA 3 chimeras to tobacco plants and protoplasts transformed with the AMV replicase genes (P12 plants). The results showed that the AMV replicase recognized the promoter for minus-strand RNA synthesis in PNRSV RNA 3 but not the promoter for plus-strand RNA synthesis. A chimeric RNA with PNRSV movement protein and CP genes accumulated in tobacco, which is a non-host for PNRSV. PMID- 11257203 TI - The molecular characterization of 16 new sequence variants of Hop stunt viroid reveals the existence of invariable regions and a conserved hammerhead-like structure on the viroid molecule. AB - At present isolates of Hop stunt viroid (HSVd) are divided into five groups: three major groups (plum-type, hop-type and citrus-type) each containing isolates from only a limited number of isolation hosts and two minor groups that were presumed to derive from recombination events between members of the main groups. In this work we present the characterization of 16 new sequence variants of HSVd obtained from four Mediterranean countries (Cyprus, Greece, Morocco and Turkey) where this viroid had not previously been described. Molecular variability comparisons considering the totality of the sequence variants characterized so far revealed that most of the variability is found in the pathogenic and variable domains of the viroid molecule whereas both the terminal right (T(R)) and left (T(L)) domains are regions of low or no variability, respectively, suggesting the existence of constraints limiting the heterogeneity of the sequence variants. Phylogenetic analyses revealed that sequence variants belonging to the two minor recombinant subgroups are more frequent than previously thought. When the cruciform structure alternative to the typical rod-like conformation was considered it was observed that the upper part of this structure (hairpin I) was strictly conserved whereas in the lower part a reduced variability was found. The existence of a covariation in this lower part was notable. Interestingly, a hammerhead-like sequence was found within the T(R) domain of HSVd and it was strictly conserved in all the sequence variants. The evolutionary implications of the presence of this motif on the HSVd are discussed. PMID- 11257204 TI - Persistent expression of a newly characterized Hyposoter didymator polydnavirus gene in long-term infected lepidopteran cell lines. AB - An Hyposoter didymator ichnovirus (HdIV) gene was stably maintained and efficiently transcribed in lepidopteran cell lines more than 3 years after HdIV infection. This K-gene had two introns and the fully spliced cDNA, named K19, comprised a short open reading frame and a long 3'-untranslated region with 13 imperfectly repeated sequences (44 to 102 nt). Transcripts related to the K-gene were detected in several long-term infected cell lines (Sf9, Spodoptera littoralis haemocytes, Trichoplusia ni). Conversely, no transcripts related to seven other viral cDNAs were detected, suggesting that the K-related DNA is selectively retained in long-term infected Sf9 cells. The function of the K-gene product and its association with stably transformed insect cell lines remains to be investigated. PMID- 11257206 TI - Molecular diagnosis of Lyme disease: review and meta-analysis. AB - The diagnosis of Lyme disease is difficult because tests that reflect active disease or have reasonable sensitivity and specificity are lacking or not timely. Molecular methods are controversial because of differences in assays, gene targets, and limited clinical validation. This review summarizes published assays for Lyme disease diagnosis using skin, plasma, synovial fluid, cerebrospinal fluid (CSF), and urine. Meta-analyses show the strengths and weaknesses of these methods. Overall, assays for skin and synovial fluid (68% and 73%, respectively) have high sensitivity and uniformity. The low test sensitivity of CSF (18%) and plasma (29%), variable sensitivities among CSF and urine assays, and persistence of Borrelia burgdorferi DNA in urine and synovial fluid even with therapy and convalescence make these unsuitable for primary diagnosis. Molecular assays for Lyme disease are best used with other diagnostic methods and only in situations in which the clinical probability of Lyme disease is high. PMID- 11257207 TI - Extracting functional information from tissues. PMID- 11257208 TI - Multisite phosphotyping of the ErbB-2 oncoprotein in human breast cancer. AB - BACKGROUND: Overexpression of the ErbB-2 (HER2/neu) receptor tyrosine kinase is one of the most common molecular changes in human cancer, but the functional significance of this phenotype remains uncertain. METHODS AND RESULTS: Using phosphorylation-specific antibodies recognizing different ErbB-2 functional states, we assessed the phosphorylation status of ErbB-2 in 102 human breast cancer specimens. Quantitative ErbB-2 immunoblotting intensity correlated directly with that of immunohistochemistry (r = 0.84). Widely varying phosphorylation profiles were evident in 65 ErbB-2-positive carcinomas, suggesting different ErbB-2 functions in different tumors. In a subset of patients for whom clinical data were obtainable, mortality trends were strongly associated with the quantitative signal intensities of ErbB-2 phosphoantibodies (P < or =.02), but not with those of conventional antibodies to ErbB-2 (P = .147), epidermal growth factor receptor (P = .44), or phosphotyrosine (P = .94). CONCLUSION: Although requiring corroboration in larger prospective clinical studies, these findings suggest that immunophenotyping using phosphorylation specific antibodies may enable more accurate prediction of cancer behavior than is currently obtainable using conventional reagents. PMID- 11257209 TI - Comparative genomic hybridization of hepatocellular carcinoma: correlation with fluorescence in situ hybridization in paraffin-embedded tissue. AB - BACKGROUND: Proto-oncogene MYC, mapped to chromosomal band 8q24 and the genes for hepatocyte growth factor (HGF at 7q21) and its receptor, MET, at chromosomal band 7q31, have an important role in the biology and growth of normal and neoplastic liver. Comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH) studies have reported frequent abnormalities of chromosomes 1 and 8 in hepatocellular carcinomas (HCCs) of various clinical and pathological stages. Chromosome 7 involvement is reported to be less frequent. MATERIALS AND METHODS: Frozen tissue from 17 HCCs was used for CGH analysis and sections of corresponding formalin-fixed, paraffin-embedded HCC tissue were used for dual color FISH with locus-specific (LSI-cMYC for chromosome 8q24 and LSI D7S486 for chromosome 7q31) and centromeric probes, CEP8 (8p11.1-q11.2) and CEP7 (7p11.1 q11.2) (Vysis, Inc, Downers Grove, IL). This study intended to determine the pattern of chromosomal aberrations in early-stage (incidental) HCC and large surgically resected HCC, and also compared the efficiency and usefulness of the two cytogenetic methods. RESULTS: CGH showed abnormalities on chromosomes 1q, 5q, 7q, 8q, 9, 10, 13q, 15, 16, 17p, 18q, 19, 20, 21, 22, and X. Gains of 8q were noted in 50% of the HCCs, including five cases of incidental HCCs by CGH. Increase in copy numbers of MYC detected by FISH was noted in 25% of tumors that had shown 8q gains by CGH and in five cases with no chromosome abnormalities noted by CGH. Three cases with 7q31 copy number abnormalities were found by FISH in addition to those detected by CGH. CONCLUSION: Combined use of CGH and FISH may provide important information about early and/or primary genetic changes in the development of HCC. PMID- 11257210 TI - Real-time PCR-based fluorescent assay for quantitation of human papillomavirus types 6, 11, 16, and 18. AB - BACKGROUND: Quantitation of human papillomavirus (HPV) DNA in clinical samples may yield important clinical information. METHODS AND RESULTS: We developed a 5' exonuclease fluorescent probe assay for HPV quantitation that uses real-time PCR. The assay was optimized for HPV types 6 (HPV-6), -11, -16, and -18. A multiplex format was developed to quantify a cellular target of known iteration simultaneously with HPV quantitation, which controls for the amount of input DNA. Dilution series of target and heterologous templates were used to verify the assay. The assay was successfully used on fresh and PreservCyt-fixed cell lines, as well as cervical samples. The linear range of the assay is from 10 to 10 million copies. Intraclass correlations for HPV, actin, and globin assays ranged from 0.95 to 0.99, indicating the analytic precision of repeated measures. CONCLUSION: The method is accurate over a large copy number range, reproducible, type specific, normalized for input DNA quantity, and applicable to PreservCyt fixed material. PMID- 11257211 TI - Is the P25L a "real" VHL mutation? AB - BACKGROUND: The von Hippel-Lindau (VHL) gene has two translational initiation sites separated by 53 codons. Both proteins have been detected in cells and have equivalent activity. A mutation in the first 53 codons of the open reading frame has no effect on the structure of the smaller protein. As expected, the vast majority of VHL mutations are downstream of the second initiation site and alter both proteins. However, several candidate mutations have been found in the first 53 codons, including a substitution of leucine for proline at position 25 (P25L) of the larger protein. METHODS AND RESULTS: DNA sequence analysis showed two VHL gene mutations, P25L and P86R, in an individual with a clinical diagnosis of VHL disease. Both mutations have been reported previously. P25L alters only the upstream protein, whereas P86R alters both VHL proteins. Based on the positions of the mutations, P86R is more likely to be pathogenically significant than the P25L mutation. A survey of anonymized DNAs for P25L, using allele-specific PCR, revealed that it is a variant with an allele frequency of approximately 0.5%. CONCLUSION: P25L is a rare variant of the VHL gene and cannot be considered a cause of VHL disease. However, this work does not prove that P25L is entirely innocuous. PMID- 11257213 TI - A seminested PCR test for simultaneous detection of two common mutations (35delG and 167delT) in the connexin-26 gene. AB - BACKGROUND: Several mutations described in the connexin-26 gene cause nonsyndromic autosomal recessive deafness (NARD). The prevalence of two frame shift mutations, known as 35delG and 167delT, was relatively high in patients with NARD from different populations. METHODS AND RESULTS: A seminested PCR test has been developed for simultaneous detection of two common mutations in the connexin-26 gene. The test is based on PCR amplification of a 285-bp DNA fragment that covers both the 35delG and 167delT mutations. The latter mutation destroys a Pst I site and is easily detected by Pst I digestion of the 285-bp DNA fragment. However, the 35delG mutation does not destroy or create a restriction site. To create a site, we designed a mismatched primer that generated an EcoN I site in an 87-bp DNA fragment, but only if the 35delG mutation was present. The test was validated using five DNA samples previously characterized for the connexin-26 mutations. After validation, we screened 45 unrelated patients with NARD and 280 healthy Omani subjects for the presence or absence of the 35delG and 167delT mutations. Neither mutation was found to be present in patients or control subjects. CONCLUSION: We developed a seminested PCR test for the simultaneous detection of both common mutations in the connexin-26 gene. In our analysis of 45 patients and 280 control subjects, the 35delG and 167delT mutations were absent in both groups. PMID- 11257212 TI - Analysis of the factor V Leiden mutation using the READIT Assay. AB - BACKGROUND: A variety of methods exist for the detection of single-nucleotide polymorphisms (SNPs) present in amplified segments of genomic DNA. We show the application of a novel SNP scoring tool for analysis of the factor V Leiden mutation. METHODS AND RESULTS: We have developed a novel method for analyzing SNPs. The luciferase-based technique, known as the READIT Technology (Promega Corp, Madison, WI), was used to analyze 510 residual human samples sent for factor V Leiden testing from three independent testing laboratories. A blinded retrospective analysis of the factor V Leiden mutation was used to determine the accuracy and throughput capabilities of the technology. One hundred percent concordance was observed between the READIT Assay and genotype assignments made in the testing laboratories. In addition, greater than 6 SDs of separation were observed between the means of wild-type and heterozygote sample populations. Repetitive sample measurements with representative wild-type, heterozygote, and mutant samples showed that greater than 9 SDs separated the means of heterozygote and homozygote sample populations. Confidence intervals based on the means of wild-type, heterozygote, and mutant sample populations were determined. CONCLUSION: Perfect concordance using the READIT Assay showed its effectiveness as a SNP scoring tool. The design of the factor V READIT Assay was straightforward, requiring the design of two unmodified oligonucleotides that differ at the 3' penultimate position to form perfect hybrids with the wild-type or Leiden form of the factor V sequence. The use of previously published amplification primers and conditions minimized the time needed to optimize and validate the assay. The READIT Calculator supplied with the assay allowed automated genotype assignments and statistical analysis from the READIT Assay data. Confidence-interval analysis validated the ability to distinguish between wild-type, heterozygote, and mutant samples using the READIT Assay. PMID- 11257214 TI - Clinical news update. PMID- 11257215 TI - Nuclear visions: functional flexibility from structural instability. PMID- 11257216 TI - Protein mobility within the nucleus--what are the right moves? PMID- 11257217 TI - Positions of potential: nuclear organization and gene expression. PMID- 11257218 TI - The control of mammalian DNA replication: a brief history of space and timing. PMID- 11257219 TI - Lamins and disease: insights into nuclear infrastructure. PMID- 11257220 TI - X inactivation of the OCNC1 channel gene reveals a role for activity-dependent competition in the olfactory system. AB - The organization of neuronal systems is often dependent on activity and competition between cells. In olfaction, the X-linked OCNC1 channel subunit is subject to random inactivation and is essential for odorant-evoked activity. Reporter-tagged OCNC1 mutant mice permit the visualization of OCNC1-deficient olfactory neurons and their projections. In heterozygous females, X inactivation creates a mosaic with two populations of genetically distinct neurons. OCNC1 deficient neurons are slowly and specifically depleted from the olfactory epithelium and display unusual patterns of projection to the olfactory bulb. Remarkably, this depletion is dependent on odorant exposure and is reversed by odorant deprivation. This suggests that odorants and the activity they evoke are critical for neuronal survival in a competitive environment and implicate evoked activity in the organization and maintenance of the olfactory system. PMID- 11257221 TI - A chemosensory gene family encoding candidate gustatory and olfactory receptors in Drosophila. AB - A novel family of candidate gustatory receptors (GRs) was recently identified in searches of the Drosophila genome. We have performed in situ hybridization and transgene experiments that reveal expression of these genes in both gustatory and olfactory neurons in adult flies and larvae. This gene family is likely to encode both odorant and taste receptors. We have visualized the projections of chemosensory neurons in the larval brain and observe that neurons expressing different GRs project to discrete loci in the antennal lobe and subesophageal ganglion. These data provide insight into the diversity of chemosensory recognition and an initial view of the representation of gustatory information in the fly brain. PMID- 11257222 TI - Inducible and reversible enhancement of learning, memory, and long-term potentiation by genetic inhibition of calcineurin. AB - The threshold for hippocampal-dependent synaptic plasticity and memory storage is thought to be determined by the balance between protein phosphorylation and dephosphorylation mediated by the kinase PKA and the phosphatase calcineurin. To establish whether endogenous calcineurin acts as an inhibitory constraint in this balance, we examined the effect of genetically inhibiting calcineurin on plasticity and memory. Using the doxycycline-dependent rtTA system to express a calcineurin inhibitor reversibly in the mouse brain, we find that the transient reduction of calcineurin activity facilitates LTP in vitro and in vivo. This facilitation is PKA dependent and persists over several days in vivo. It is accompanied by enhanced learning and strengthened short- and long-term memory in several hippocampal-dependent spatial and nonspatial tasks. The LTP and memory improvements are reversed fully by suppression of transgene expression. These results demonstrate that endogenous calcineurin constrains LTP and memory. PMID- 11257223 TI - EGF receptor and Notch signaling act upstream of Eyeless/Pax6 to control eye specification. AB - The Drosophila compound eye is specified by the concerted action of seven nuclear factors that include Eyeless/Pax6. These factors have been called "master control" proteins because loss-of-function mutants lack eyes and ectopic expression can direct ectopic eye development. However, inactivation of these genes does not cause the presumptive eye to change identity. Surprisingly, we find that several of these eye specification genes are not coexpressed in the same embryonic cells-or even in the presumptive eye. We demonstrate that the EGF Receptor and Notch signaling pathways have homeotic functions that are genetically upstream of the eye specification genes, and show that specification occurs much later than previously thought-not during embryonic development but in the second larval stage. PMID- 11257224 TI - The EGF receptor defines domains of cell cycle progression and survival to regulate cell number in the developing Drosophila eye. AB - The number of cells in developing organs must be controlled spatially by extracellular signals. Our results show how cell number can be regulated by cell interactions controlling proliferation and survival in local neighborhoods in the case of the Drosophila compound eye. Intercellular signals act during the second mitotic wave, a cell cycle that generates a pool of uncommitted cells used for most ommatidial fates. We find that G1/S progression to start the cell cycle requires EGF receptor inactivity. EGF receptor activation is then required for progression from G2 to M phase of the same cells, and also prevents apoptosis. EGF receptor activation depends on short-range signals from five-cell preclusters of photoreceptor neurons not participating in the second mitotic wave. Through proliferation and survival control, such signals couple the total number of uncommitted cells being generated to the neural patterning of the retina. PMID- 11257225 TI - Conserved role of a complement-like protein in phagocytosis revealed by dsRNA knockout in cultured cells of the mosquito, Anopheles gambiae. AB - We characterize a novel hemocyte-specific acute phase glycoprotein from the malaria vector, Anopheles gambiae. It shows substantial structural and functional similarities, including the highly conserved thioester motif, to both a central component of mammalian complement system, factor C3, and to a pan-protease inhibitor, alpha2-macroglobulin. Most importantly, this protein serves as a complement-like opsonin and promotes phagocytosis of some Gram-negative bacteria in a mosquito hemocyte-like cell line. Chemical inactivation by methylamine and depletion by double-stranded RNA knockout demonstrate that this function is dependent on the internal thioester bond. This evidence of a complement-like function in a protostome animal adds substantially to the accumulating evidence of a common ancestry of immune defenses in insects and vertebrates. PMID- 11257226 TI - Nongenotropic, sex-nonspecific signaling through the estrogen or androgen receptors: dissociation from transcriptional activity. AB - The relationship of the classical receptors and their transcriptional activity to nongenotropic effects of steroid hormones is unknown. We demonstrate herein a novel paradigm of sex steroid action on osteoblasts, osteocytes, embryonic fibroblasts, and HeLa cells involving activation of a Src/Shc/ERK signaling pathway and attenuating apoptosis. This action is mediated by the ligand binding domain and eliminated by nuclear targeting of the receptor protein; ERalpha, ERbeta, or AR can transmit it with similar efficiency irrespective of whether the ligand is an estrogen or an androgen. This antiapoptotic action can be dissociated from the transcriptional activity of the receptor with synthetic ligands, providing proof of principle for the development of function-specific-as opposed to tissue-selective-and gender-neutral pharmacotherapeutics. PMID- 11257227 TI - Arginine methylation of STAT1 modulates IFNalpha/beta-induced transcription. AB - Transcriptional induction by interferons requires the tyrosine and serine phosphorylation of STAT transcription factors. The N-terminal region is highly homologous among the STAT proteins and surrounds a completely conserved arginine residue. Here we demonstrate arginine methylation of STAT1 by the protein arginine methyl-transferase PRMT1 as a novel requirement for IFNalpha/beta induced transcription. Methyl-thioadenosine, a methyl-transferase inhibitor that accumulates in many transformed cells, inhibits STAT1-mediated IFN responses. This inhibition arises from impaired STAT1-DNA binding due to an increased association of the STAT inhibitor PIAS1 with phosphorylated STAT1 dimers in the absence of arginine methylation. Thus, arginine methylation of STAT1 is an additional posttranslational modification regulating transcription factor function, and alteration of arginine methylation might be responsible for the lack of interferon responsiveness observed in many malignancies. PMID- 11257228 TI - The BARD1-CstF-50 interaction links mRNA 3' end formation to DNA damage and tumor suppression. AB - The mRNA polyadenylation factor CstF interacts with the BRCA1-associated protein BARD1, and this interaction represses the nuclear mRNA polyadenylation machinery in vitro. Given the suspected role of BRCA1/BARD1 in DNA repair, we tested whether inhibition of mRNA processing is linked to DNA damage. Strikingly, we found that 3' cleavage in extracts from cells treated with hydroxyurea or ultraviolet light was strongly, but transiently, inhibited. Although no changes were detected in CstF, BARD1, and BRCA1 protein levels, increased amounts of a CstF/BARD1/BRCA1 complex were detected. Supporting the physiological significance of these results, a previously identified tumor-associated germline mutation in BARD1 (Gln564His) reduced binding to CstF and abrogated inhibition of polyadenylation. Together these results indicate a link between mRNA 3' processing and DNA repair and tumor suppression. PMID- 11257229 TI - Structural analyses of DNA recognition by the AML1/Runx-1 Runt domain and its allosteric control by CBFbeta. AB - The core binding factor (CBF) heterodimeric transcription factors comprised of AML/CBFA/PEBP2alpha/Runx and CBFbeta/PEBP2beta subunits are essential for differentiation of hematopoietic and bone cells, and their mutation is intimately related to the development of acute leukemias and cleidocranial dysplasia. Here, we present the crystal structures of the AML1/Runx-1/CBFalpha(Runt domain) CBFbeta(core domain)-C/EBPbeta(bZip)-DNA, AML1/Runx-1/CBFalpha(Runt domain) C/EBPbeta(bZip)-DNA, and AML1/Runx-1/CBFalpha(Runt domain)-DNA complexes. The hydrogen bonding network formed among CBFalpha(Runt domain) and CBFbeta, and CBFalpha(Runt domain) and DNA revealed the allosteric regulation mechanism of CBFalpha(Runt domain)-DNA binding by CBFbeta. The point mutations of CBFalpha related to the aforementioned diseases were also mapped and their effect on DNA binding is discussed. PMID- 11257230 TI - Structural basis of caspase-7 inhibition by XIAP. AB - The inhibitor of apoptosis (IAP) proteins suppress cell death by inhibiting the catalytic activity of caspases. Here we present the crystal structure of caspase 7 in complex with a potent inhibitory fragment from XIAP at 2.45 A resolution. An 18-residue XIAP peptide binds the catalytic groove of caspase-7, making extensive contacts to the residues that are essential for its catalytic activity. Strikingly, despite a reversal of relative orientation, a subset of interactions between caspase-7 and XIAP closely resemble those between caspase-7 and its tetrapeptide inhibitor DEVD-CHO. Our biochemical and structural analyses reveal that the BIR domains are dispensable for the inhibition of caspase-3 and -7. This study provides a structural basis for the design of the next-generation caspase inhibitors. PMID- 11257231 TI - Structural basis of caspase inhibition by XIAP: differential roles of the linker versus the BIR domain. AB - The inhibitor of apoptosis proteins (IAPs) represent the only endogenous caspase inhibitors and are characterized by the presence of baculoviral IAP repeats (BIRs). Here, we report the crystal structure of the complex between human caspase-7 and XIAP (BIR2 and the proceeding linker). The structure surprisingly reveals that the linker is the only contacting element for the caspase, while the BIR2 domain is invisible in the crystal. The linker interacts with and blocks the substrate groove of the caspase in a backward fashion, distinct from substrate recognition. Structural analyses suggest that the linker is the energetic and specificity determinant of the interaction. Further biochemical characterizations clearly establish that the linker harbors the major energetic determinant, while the BIR2 domain serves as a regulatory element for caspase binding and Smac neutralization. PMID- 11257232 TI - Structural basis for the inhibition of caspase-3 by XIAP. AB - The molecular mechanism(s) that regulate apoptosis by caspase inhibition remain poorly understood. The main endogenous inhibitors are members of the IAP family and are exemplified by XIAP, which regulates the initiator caspase-9, and the executioner caspases-3 and -7. We report the crystal structure of the second BIR domain of XIAP (BIR2) in complex with caspase-3, at a resolution of 2.7 A, revealing the structural basis for inhibition. The inhibitor makes limited contacts through its BIR domain to the surface of the enzyme, and most contacts to caspase-3 originate from the N-terminal extension. This lies across the substrate binding cleft, but in reverse orientation compared to substrate binding. The mechanism of inhibition is due to a steric blockade prohibitive of substrate binding, and is distinct from the mechanism utilized by synthetic substrate analog inhibitors. PMID- 11257233 TI - A.E. Bennett Research Award. Prenatal rubella, premorbid abnormalities, and adult schizophrenia. AB - BACKGROUND: Premorbid neurocognitive, neuromotor, and behavioral function tends to be disturbed in schizophrenia. We previously demonstrated that a birth cohort clinically and serologically documented with prenatal rubella evidenced a marked increase in risk of nonaffective psychosis. In our study, we examined whether rubella-exposed subjects destined to develop schizophrenia and other schizophrenia spectrum disorders (SSD), compared with exposed control subjects, had greater impairment in several premorbid functions. METHODS: Subjects were interviewed using a direct, comprehensive research assessment and diagnosed by consensus. We compared the degree of IQ decline, as well as premorbid neuromotor and behavioral dysfunction, between rubella-exposed subjects who developed schizophrenia spectrum psychosis (SSP) and exposed control subjects from the cohort. We also compared the gestational timing of rubella infection between the cases and control subjects. RESULTS: This rubella-exposed birth cohort evidenced a markedly increased risk of SSD (20.4% or 11/53). Rubella-exposed SSP cases, compared with rubella-exposed control subjects, demonstrated a decline in IQ from childhood to adolescence, and increased premorbid neuromotor and behavioral abnormalities. Moreover, it appears that early gestational rubella exposure may represent a period of increased vulnerability for SSD. CONCLUSIONS: These findings link a known prenatal exposure, a deviant neurodevelopmental trajectory in childhood and adolescence, and SSP in adulthood within the same individuals. PMID- 11257234 TI - Longitudinal study of brain morphology in first episode schizophrenia. AB - BACKGROUND: Beginning with Kraepelin, schizophrenia has been viewed as a progressive disorder. Although numerous studies of the longitudinal course of schizophrenia have demonstrated the clinical deterioration that occurs predominantly in the early stages of the illness, the pathophysiology of this clinical phenomenon has not been established. This aspect of the illness may be of critical importance to understanding the pathogenesis of schizophrenia and determining preventive therapeutic strategies. Abnormalities in brain morphology have been consistently described in schizophrenia, but it is not known when in the natural history of the illness they arise and whether they are progressive. Previous studies of brain morphology have been inconclusive, in part because of the variability of methods for image acquisition and analysis, assessment of patients already at chronic stages of their illness with extensive prior treatment exposure, and inadequate periods of follow-up. METHODS: To address these questions we examined 107 patients in their first episode of schizophrenia or schizoaffective disorder and 20 healthy volunteers using high resolution magnetic resonance imaging (MRI) and clinical assessments of psychopathology and treatment outcome for periods of up to 6 years. Fifty-one patients and 13 control subjects had MRIs after at least 12 months of follow-up. RESULTS: Results confirm the findings of ventricular enlargement and anterior hippocampal volume reductions in first episode schizophrenia patients that have been previously reported. In addition, we found changes in selected structures over time in relation to treatment outcome, including increases in ventricular volume that were associated with poor outcome patients. Contrary to our hypothesis, there were no significant reductions in cortical and hippocampal volumes over time. CONCLUSIONS: The finding of progressive ventricular enlargement in patients with poor outcome schizophrenia is consistent with the hypothesis that persistent positive and negative symptoms result in progressive brain changes in the form of ventricular enlargement, possibly due to neurodegeneration rather than the confounding effects of treatment. Future studies of first episodes of schizophrenia should utilize higher resolution imaging techniques that compare clinically well characterized patients with and without poor outcome and recurrent symptoms to control subjects who are well matched to patients for age and gender. There is also a need to control for treatment effects of typical antipsychotic medication on brain structure. PMID- 11257235 TI - Cortical instability and the mechanism of mania: a neural network simulation and perceptual test. AB - BACKGROUND: A previous neural network simulation suggested that manic states arise from excessive levels of noise that destabilize neural representations. The Necker cube stick figure provides a simple perceptual task that assesses stability of gestalt-type representations. METHODS: A neural network was developed that included a simulation of the Necker cube task. Noise was added to induce maniclike jumps from one representation to another. A parallel study of Necker cube perception was conducted with 16 patients diagnosed with manic spectrum disorder, 18 patients with schizophrenia, and 19 normal control subjects. Cognitive speed and rate of indiscriminate responses were assessed using an auditory continuous performance task. RESULTS: During processing of the "Necker cube" stimulus, the reversal rate of the noise-destabilized "manic" network was increased by 30%. In the human subject study, the median score of Necker cube reversal rates for manic-spectrum patients was roughly twice that of normal control subjects and patients with schizophrenia. Accelerated reversal rates in the manic-spectrum group were not attributable to excessive cognitive speed or higher rates of indiscriminate responses. CONCLUSIONS: The two studies, considered together, support the hypothesis that excessive cortical noise destabilizes neural representations in manic-spectrum patients. PMID- 11257236 TI - Significantly reduced docosahexaenoic and docosapentaenoic acid concentrations in erythrocyte membranes from schizophrenic patients compared with a carefully matched control group. AB - BACKGROUND: Fatty acid research in schizophrenia has demonstrated an altered cell membrane phospholipid metabolism. Erythrocyte membrane phospholipid composition closest reflects that of neuronal membranes. METHODS: (Poly)(un)saturated fatty acid concentrations were measured in the erythrocyte membranes of 19, consecutively admitted, medicated young schizophrenic patients and then compared with matched control subjects. Psychiatric symptomatology was rated with the Positive and Negative Symptom Scale and Montgomery-Asberg Depression Rating Scale. Because diet, hormones, and cannabis influence fatty acid metabolism, we included these factors in our study. RESULTS: The most distinctive findings concerned the omega-3 series: C22:5 omega-3, C22:6 omega-3 (docosahexaenoic acid), and the sum of omega-3 fatty acids were significantly decreased. Interestingly, C20:4 omega-6 (arachidonic acid) was not lowered. In the omega-9 series, higher levels of C22:1 omega-9 and lower levels its elongation product, C24:1 omega-9 (nervonic acid), were found. Interestingly, the other arm of the desaturation-elongation sequence of C18:1 omega-9, C20:3 omega-9, was lower in patients. The total omega-9 fatty acid levels were also lower in patients. CONCLUSIONS: Significant differences in erythrocyte fatty acid composition were found. The differences were not due to diet or hormonal status and could not be explained by the medication or cannabis use. No consistent pattern emerged from the different fatty acid abnormalities and the clinical symptom scores. PMID- 11257237 TI - Increased tachykinin NK(1) receptor immunoreactivity in the prefrontal cortex in schizophrenia. AB - BACKGROUND: Changes in levels of substance P and substance P-binding sites have been implicated in schizophrenia. However, no studies have used receptor-specific antibodies to directly investigate the substance P (neurokinin 1) receptor in schizophrenia. METHODS: We used an antibody directed against the human neurokinin 1 receptor to compare the distribution of neurokinin-1 receptors in the prefrontal cortices from six subjects with schizophrenia and six control subjects, matched for age, gender, and postmortem interval. RESULTS: In control tissue, dots of neurokinin-1 receptor immunoreactivity were observed in layer I to upper/mid layer III only. In contrast, dots of neurokinin-1 receptor immunoreactivity were observed in all layers of the prefrontal cortex in subjects with schizophrenia, and the density of dots was significantly greater than in control subjects. CONCLUSIONS: This is the first report of increased neurokinin-1 receptor immunoreactivity in the prefrontal cortex in subjects with schizophrenia. These changes may have implications for understanding the pathophysiology of the prefrontal cortex in schizophrenia and for the treatment of this disorder. PMID- 11257238 TI - Delayed onset of enhanced MK-801-induced motor hyperactivity after neonatal lesions of the rat ventral hippocampus. AB - BACKGROUND: Abnormalities in the glutamatergic system, glutamate/dopamine/gamma aminobutyric acid interactions, and cortical development are implicated in schizophrenia. Moreover, patients with schizophrenia show symptom exacerbation in response to N-methyl-D-aspartate (NMDA) antagonist drugs. Using an animal model of schizophrenia, we compared the impact of neonatal and adult hippocampal lesions on behavioral responses to MK-801, a noncompetitive NMDA antagonist. METHODS: Neonatal rats were lesioned on postnatal day 7. Their motor activity in response to MK-801 was tested at a juvenile age, in adolescence, and in adulthood. We also measured binding of [(3)H]MK-801 and the expression of NR1 messenger RNA (mRNA) in the medial prefrontal cortex and nucleus accumbens. Adult rats received similar lesions and were tested 4 and 8 weeks after the lesion. RESULTS: As juveniles, neonatally lesioned rats did not differ from control rats in responsiveness to MK-801, whereas in adolescence and adulthood they showed more pronounced hyperactivity than control rats. The adult lesion did not alter behaviors elicited by MK-801. Neonatally lesioned rats showed no apparent changes in [(3)H]MK-801 binding or expression of the NR1 mRNA. CONCLUSIONS: These results suggest that an early lesion of the ventral hippocampus affects development of neural systems involved in MK-801 action without changes at the NMDA receptor level, and they show that the behavioral changes manifest first in early adulthood. PMID- 11257239 TI - A quantitative MRI study of posterior fossa development in velocardiofacial syndrome. AB - BACKGROUND: Velocardiofacial syndrome (VCFS) has been identified as a risk factor for developing schizophrenia. Qualitative neuroimaging studies indicated that VCFS was frequently associated with abnormal development of structures in the posterior fossa of the brain. The objective of this investigation was to identify the specific structures affected in the posterior fossa and investigate the association of these neuroanatomic variations with behaviors potentially related to later-onset psychiatric disorders. METHODS: Twenty-four children and adolescents with VCFS individually matched for age and gender with 24 control subjects received magnetic resonance imaging scans. Analysis of covariance models were used to investigate regional brain differences. Association between brain areas and behaviors measured on the Child Behavior Checklist (CBCL) were assessed using simple regression models. RESULTS: Children with VCFS had significantly smaller size of vermal lobules VI--VII and the pons after adjusting for overall brain size. There were no significant associations between scores on the CBCL and measures of neuroanatomic variation within the VCFS group. CONCLUSIONS: Structural alterations of the posterior fossa in VCFS are specifically limited to cerebellar vermis lobules VI--VII and pons. Previous literature has suggested that the vermis is involved in social cognition, and alteration of lobules VI- VII could therefore partially explain the neurobehavioral profile associated with VCFS. PMID- 11257240 TI - Increased glucocorticoid production and altered cortisol metabolism in women with mild to moderate Alzheimer's disease. AB - BACKGROUND: Abnormalities at several levels of the hypothalamic-pituitary-adrenal axis, which may promote glucocorticoid dependent neurodegeneration, have been reported in patients with Alzheimer's disease. In this study we have explored the possibility that peripheral cortisol metabolism is enhanced and glucocorticoid production is increased compensatory in patients with mild to moderate Alzheimer's disease. METHODS: Urinary excretion of cortisol and its principal conjugated metabolites was studied in ten women with mild to moderate Alzheimer's disease, seven healthy elderly women and seven young women. RESULTS: There was an age-related fall in glucocorticoid production (median, 2009 [range 1828--4201] vs. 9315 [range 3613--16,244] microg/24 hours in elderly vs. young control subjects). A-ring metabolism (i.e., the irreversible inactivation of cortisol by 5 alpha- and 5 beta-reductases) was reduced in old age. However, patients with Alzheimer's disease showed persistence of cortisol metabolite excretion. Glucocorticoid production was significantly increased in patients with Alzheimer's disease versus healthy elderly control subjects (median, 7269 [range 6005-15,335] vs. 2009 [range 1828--4201] microg/24 hours), and patients with Alzheimer's disease had increased A-ring reduction of cortisol. CONCLUSIONS: An increased glucocorticoid production is an early feature of Alzheimer's disease and may be secondary to enhanced metabolic clearance of cortisol by A-ring reduction. PMID- 11257241 TI - Clinical outcomes following cocaine infusion in nontreatment-seeking individuals with cocaine dependence. AB - BACKGROUND: In this study we explored if laboratory-based cocaine administration to human subjects was associated with long-term adverse outcomes. METHODS: Twenty one non--reatment seeking individuals with cocaine dependence were evaluated at baseline and again 5 and 10 months following cocaine infusion in a brain imaging study. Outcomes included computer-driven multidimensional clinical assessments and radioimmunoassay of hair. For comparison, identical data were collected from 19 cocaine-dependent subjects who did not receive the infusion. RESULTS: The infused and noninfused groups did not differ on frequency of cocaine use (corroborated by radioimmunoassay of hair), Addiction Severity Index drug composite score, or Hamilton Rating Scale for Depression score at both follow-up time points. In a time-related trend analysis, both groups showed significant reductions in frequency of cocaine use. CONCLUSIONS: Laboratory-based cocaine administration can be a safe paradigm even in individuals who are not engaged in treatment. PMID- 11257242 TI - Comparison of the lipoprotein, carbohydrate, and hemostatic effects of phasic oral contraceptives containing desogestrel or levonorgestrel. AB - Desogestrel (DSG) is a less-androgenic progestogen than levonorgestrel (LNG). This difference in androgenicity may be responsible for observed differences in metabolic effects between oral contraceptive (OC) formulations containing almost equivalent estrogen doses but with either DSG or LNG as a progestogen. To test the hypothesis, a prospective 9-month randomized comparison of plasma lipids, glucose, insulin, hemostasis, and sex hormone binding globulin (SHBG) was conducted in 66 healthy women using phasic formulations of OCs containing either DSG (DSG-OC) or LNG (LNG-OC). The study results showed that SHBG increased 3-fold with DSG-OC and 2-fold with LNG-OC. DSG-OC increased HDL-C, HDL(2)-C and HDL(3) C; LDL-C decreased transiently. LNG-OC decreased HDL(2)-C and increased HDL(3)-C; HDL-C was unchanged and LDL-C decreased transiently. Both formulations increased VLDL-C and triglycerides, more with DSG-OC, but apolipoprotein B levels increased equally. Apo A-I and A-II increased more with DSG-OC than with LNG-OC. Neither formulation altered Lp(a) or fasting glucose and insulin levels. Postprandially, both formulations decreased glucose and increased insulin responses, but to an equivalent degree. Both OCs slightly enhanced procoagulant and profibrinolytic parameters to the same extent except for internally compensating decreases in Factor V and protein S with DSG-OC. In summary, at almost equivalent estrogen doses, a phasic OC containing DSG compared with LNG has a less androgenic effect on lipoproteins and SHBG, similar effects on hemostatic parameters with lower protein S and factor V activity and equivalent effects on carbohydrate metabolism. The lipoprotein, SHBG, and protein S and factor V differences are likely due to the lesser androgenicity of DSG allowing for a greater expression of the dose of estrogen. PMID- 11257243 TI - The possible role of enterohepatic cycling on bioavailability of norethisterone and gestodene in women using combined oral contraceptives. AB - Using steady-state conditions we aimed to test if administration of oral activated charcoal affects the bioavailability of norethisterone acetate (NET Ac) and gestodene (GEST) by inhibiting their enterohepatic recirculation. Thirteen volunteers received, in a randomized order, Minulet (75 microg GEST and 30 microg ethinylestradiol [EE(2)]) and Econ/30 (1 mg NET Ac and 30 microg EE(2)), each for 4 months. Serum GEST and norethisterone (NET) levels were evaluated with respect to C(max,) t(max) and 24-h area under the curve (AUC(0-24h)) in the middle of the control (3rd) cycle and the charcoal treatment (4th) cycle during both pill treatments. No statistically significant difference was seen in any of the aforementioned variables between the control and charcoal treatment cycles of either pill. Neither was a difference seen in the bioavailability of GEST and NET as evaluated by the ratios of two 24-h AUCs calculated in the control and charcoal cycles of each pill treatment (p = 0.29). The results suggest that enterohepatic circulation of GEST and NET is not of clinical importance. We conclude that women on oral contraceptives can take activated charcoal for the treatment of diarrhea when administered 3 h after and at least 12 h before pill intake. PMID- 11257244 TI - Lack of effect of alcohol on ethinylestradiol in premenopausal women. AB - An acute elevation in estradiol during alcohol intake has been reported in postmenopausal women on estrogen replacement therapy. The objective of the present study was to investigate the acute and long-term effect of alcohol on ethinylestradiol, the estrogen component found in most oral contraceptives. Nine healthy premenopausal women with regular use of an oral contraceptive containing 30 microg ethinylestradiol and 75 microg gestodene were challenged with alcohol (0.4 g/kg p.o., approximately 2-3 standard drinks) 2 h after intake of the oral contraceptive pill at menstrual cycle day 14. Blood samples were taken at 0, 2, 3, 4, 5, and 6 h from intake of alcohol. The challenge was repeated after a 7-day period of controlled alcohol intake (0.8 g/kg/day) at cycle day 21. The same experiments were carried out during placebo conditions. At day 21 an increase in the alcohol elimination rate was observed compared with day 14. No significant acute or long-term effect of alcohol on ethinylestradiol was found. The lack of an acute effect comparable to that reported for estradiol may be due to the protection of the ethinyl group at the 17-position of ethinylestradiol. PMID- 11257245 TI - Randomized crossover trial comparing the eZ.on plastic condom and a latex condom. AB - This randomized crossover trial compared the breakage and slippage rates, safety, and acceptability of the recently developed polyurethane bi-directional eZ.on condom with a marketed latex condom. Three hundred sixty couples were asked to use 4 eZ.on condoms and 4 latex condoms. Like several other non-latex condoms tested to date, the eZ.on condom had a higher clinical breakage rate than its latex comparator, while the slippage rates were similar. The clinical breakage rate for the eZ.on condom was 5.6%, compared with 0.9% for the latex condom (difference = 4.76%, with upper 95% confidence bound on the difference = 6.26%). Thus, based on an a priori definition of a 2% clinically acceptable difference, the study failed to conclude equivalence relative to clinical breakage. The complete slippage rate for eZ.on was 1.6%; compared to 0.7% for latex (difference = 0.87%, with upper 95% confidence bound = 1.55%). Thus, based on an a priori definition of a 2% difference we concluded equivalence relative to complete slippage. The safety profile of the eZ.on condom was good and similar to the latex condom. The eZ.on was also found to be easier to don and remove than the latex condom. Although no overall preference existed for either condom, nearly 30%women and men strongly preferred the eZ.on condom to the latex condom. The eZ.on condom may be an acceptable alternative for couples unable or unwilling to use latex condoms. PMID- 11257246 TI - Structural integrity of the female condom after multiple uses, washing, drying, and re-lubrication. AB - Establishing the safety of re-using the female condom could significantly increase women's access to barrier methods especially in poorer countries. In this study, the structural integrity of female condoms was tested (n = 295) after multiple acts of vaginal intercourse. Fifty women were recruited to the study. Each woman re-used one condom up to eight times and washed, dried, and re lubricated between each use. Structural integrity was measured using standard quality control testing; water-leakage, air-burst, and seam tensile strength. All results were compared to the United States Food and Drug Administration (US FDA) standards for an unused female condom. The results of the structural integrity tests for all cycles were above the FDA minimum standards for seam strength and burst tests. There was no deterioration detected in condoms used 8 times when compared to new female condoms in these tests. Five holes were detected by the water leakage test across all cycles, of which three were detected by the subjects themselves and reported to the investigators, therefore, giving a breakage rate of 1.7%. The holes were not associated with increased number of uses. This study provides further evidence that suggests the structural integrity of the female condom after multiple use is still within FDA minimum standards, although random holes resulting from handling occur infrequently with the re-use procedure. PMID- 11257247 TI - Intrauterine device and maternal copper metabolism during lactation. AB - The effects of intrauterine device (IUD) on maternal copper (Cu) metabolism during breastfeeding was studied in 95 volunteer mothers who chose to use non hormonal contraceptive methods. They were divided into two groups that were inserted with the IUD-Cu 380A (n = 33), IUD-Cu 200B (n = 29), and a third group that did not use any IUDs served as control (n = 33). Endometrial biopsies, blood, and milk samples were collected before (at 10 weeks postpartum) and 6 weeks after insertion of device for the determination of metabolites associated with copper metabolism, namely, serum ceruloplasmin, and copper concentrations in breast milk and endometrium. Endometrial copper concentration increased in women using IUDs, but was statistically significant (p = 0.001) only in the IUD-Cu 380A group. The increase in endometrial copper concentration did not affect serum ceruloplasmin or milk copper concentrations. PMID- 11257248 TI - Evaluation of active and passive transport mechanisms in genital tracts of IUD bearing women with radionuclide hysterosalpingoscintigraphy. AB - The objective of this study was to evaluate the active and passive transport mechanisms in the genital tracts of copper T-200 intrauterine device (IUD) bearing women. (Tc-99m)HMPAO-labeled spermatozoa and (Tc-99m)-labeled albumin macrospheres were placed into the vagina at midcycle. Serial scintigraphic images were obtained over a period of 2 h. Migration of spermatozoa and particles in the genital tract and the direction of transport related to dominant follicle were evaluated. While active sperm migration was greatly inhibited, the passive transport of the particles was not affected in IUD-bearing women. The direction of radiolabeled particles and spermatozoa was toward the dominant follicle side. Passive transport was not affected, whereas active transport of spermatozoa was strongly inhibited in the genital tract by the presence of the IUD. However, the direction of active and passive transport related to dominant follicle side was unchanged in IUD-bearing women and was preferentially toward the tube ipsilateral to the dominant follicle. PMID- 11257249 TI - Optimization of contraceptive dosage regimen of Centchroman. AB - Centchroman (Ormeloxifene), a non-steroidal oral contraceptive, is used at a dose of 30 mg once a week. To prevent failures in the beginning of the therapy, it is recommended that a dose of 30 mg twice a week for 12 weeks be administered to build up adequate blood levels. The present study was undertaken to simplify the dosing schedule without sacrificing the purpose of twice a week dosing regimen, using modeling and measurement approaches. The drug was given to 60 female volunteers who were divided into seven groups: group I, 30 mg weekly; group II, 30 mg twice a week; group III, 30 mg twice a week for 12 weeks followed by 30 mg weekly; group IV, 30 mg twice a week for 6 weeks followed by 30 mg weekly; group V, 60 mg weekly; and groups VI and VII, single 60 mg loading dose followed by 30 mg weekly doses. The blood samples were collected and analyzed by HPLC. In group I, mean trough concentrations of centchroman and its active metabolite, 7 desmethyl centchroman, were comparable to the steady-state trough concentrations in groups III, IV, VI, and VII. The metabolite to parent drug ratio remained constant in all the groups. The pharmacokinetic parameters in group VII were comparable to those reported after a single 30 mg dose. Dosage regimen VI was more convenient and provided better pregnancy protection (Pearl index 1.18; unpublished report) than regimen III, which is currently on the market and, thus, could be effectively used for contraception. PMID- 11257250 TI - Absence of interceptive effect in rats treated with Solanum lycocarpum (St. Hil). AB - The interceptive effect of the powder of Solanum lycocarpum (St. Hil) (Solanaceae) fruit, used as a hypoglycemic agent by diabetic patients in Minas Gerais state (Brazil), was evaluated to observe possible effects upon zygote and pre-embryo transport in rats, since it contains solamargine and solasonine from which a 3beta-acetoxipregna-5,16-dien-20-one is obtained as well as an alkaloid with stereospecific configuration to the synthesis of steroid hormones. Inseminated rats received an aqueous suspension of 100 mg of the lobeira powder/kg of body weight, by oral gavage, from the 1st to the 4th day of pregnancy. A control group received 5 mL of distilled water in the same schedule. The pregnant rats were weighed at the beginning of treatment and on sacrifice day. Animals were killed on the 5th day of pregnancy. The oviducts and uterine horns were removed and flushed with saline solution to count expanded blastocysts. It was concluded that administration of lobeira did not cause maternal toxicity, alteration of the pre-embryo transport or reduction of the number of expanded blastocysts. PMID- 11257251 TI - Genetics and atherosclerosis: broadening the horizon. PMID- 11257252 TI - Polymorphisms in endothelial nitric oxide synthase and atherogenesis: John French Lecture 2000. PMID- 11257253 TI - Apolipoprotein E genotype and cardiovascular disease in the Framingham Heart Study. AB - BACKGROUND: Apolipoprotein (apo) E is a constituent of lipoproteins with considerable variation due to cysteine-arginine exchanges. The apo E4 (Arg112 Cys) polymorphism has been associated with dementia and hypercholesterolemia. We investigated the relation of APOE genotype to cardiovascular disease (CVD) in the Framingham Offspring Study. METHODS AND RESULTS: DNA was isolated from 3413 study participants and APOE genotypes were determined utilizing the polymerase chain reaction and restriction isotyping. In the entire group of subjects, 20.7% had apo E4/4 or E3/4 (Group E4); 14.1% had apo E2/2 or E2/3 (Group E2) and 63.9% had the apo E3/3 genotype (Group E3). Subjects with E2/4 (1.3%) were excluded. Period prevalence of CVD between examinations 1 and 5 (1971-1994) (366 events) was related to APOE genotype. Age adjusted period prevalence of CVD in men was 18.6% for Group E4, 18.2% for Group E2 and 12.7% for Group E3 (P=0.004); while in women these rates were 9.9, 4.9, and 6.6%, respectively (P=0.037). After adjustment for non-lipid risk factors the relative odds for CVD in Group E2 men was 1.79 (P=0.0098) and in Group E4 it was 1.63 (P=0.0086) compared with the Group E3; while in Group E4 women it was 1.56 (P=0.054). After adjustment for all CVD risk factors, the relative odds in Group E2 men was 1.94 (P=0.004) and in Group E4 men it was 1.51 (P=0.0262). CONCLUSIONS: The presence of the apo E2 or apo E4 alleles in men is associated with significantly greater CVD risk. This genotypic information may help to identify individuals at increased risk for CVD events. PMID- 11257254 TI - Effect of apolipoprotein E3/4 phenotype on postprandial triglycerides and retinyl palmitate metabolism in plasma from hyperlipidemic subjects in Japan. AB - In a previous study it was shown that postprandial lipid metabolism is delayed in individuals with intra-abdominal visceral fat accumulation. Population studies have shown that as compared with individuals with apolipoprotein (apo) E3/3, those with phenotype apo E3/4 phenotype have higher plasma and low density lipoprotein (LDL)-cholesterol (C) concentration and increased susceptibility to coronary heart disease. The aim of the present study is to determine how apo E4 affects postprandial lipid metabolism by comparing individuals with apo E3/4 to those with apo E3/3 phenotype matched for abdominal visceral fat. Sixty-two Japanese subjects (41 male, 21 female) [average age 48+/-14 years; mean body mass index (BMI) 25+/-5.6 kg/m2] were recruited for this study. The subjects were divided into two groups: those with apo E3/3 (n=43) and those with apo E3/4 phenotype (n=19), as determined by isoelectric focusing (IEF). Visceral fat accumulation was analyzed as area of fat deposition by computerized tomography at the umbilicus level. After a 12-h overnight fasting, an oral vitamin A and a fatty meal were administered to these subjects. The plasma triglyceride (TG) increased significantly hours after fat loading in both groups but the levels of TG were significantly higher in apo E3/4 than in apo E3/3 phenotype at 2, 4 and 6 h after fat loading. Plasma retinyl palmitate (RP) levels were also significantly higher in individuals with apo E3/4 than in those with apo E3/3 phenotype at 2, 4 and 6 h after fat loading. This investigation was then conducted in both genders separately, and found that these associations were statistically significant in men. Furthermore, after matching men for fasting TG levels, these associations did not persist for plasma TG levels at any time point, while plasma RP levels were still significantly higher in apo E3/4 group at 2 and 6 h after fat loading. These results indicate that in Japanese population especially for men apo E phenotype E3/4 is associated with an impaired postprandial TG-rich lipoprotein metabolism relative to apo E3/3 phenotype when matched for intra-abdominal visceral fat accumulation, which has a substantial effect on the metabolism of plasma TG-rich lipoproteins. PMID- 11257255 TI - Apoprotein E genotype and the response of serum cholesterol to dietary fat, cholesterol and cafestol. AB - Previous studies on the effect of apoprotein E (APOE) polymorphism on the response of serum lipids to diet showed inconsistent results. We therefore studied the effect of apoprotein E polymorphism on responses of serum cholesterol and lipoproteins to various dietary treatments. We combined data on responses of serum cholesterol and lipoproteins to saturated fat, to trans-fat, to dietary cholesterol, and to the coffee diterpene cafestol with newly obtained data on the apoprotein E polymorphism in 395 mostly normolipidemic subjects. The responses of low-density lipoprotein (LDL-) cholesterol to saturated fat were 0.08 mmol/l larger in subjects with the APOE3/4 or E4/4 genotype than in those with the APOE3/3 genotype (95% confidence interval: -0.01-0.18 mmol/l). In contrast, responses of LDL-cholesterol to cafestol were 0.11 mmol/l smaller in subjects with the APOE3/4 or E4/4 genotype than in those with the APOE3/3 genotype (95% confidence interval: -0.29-0.07 mmol/l). Responses to dietary cholesterol and trans-fat did not differ between subjects with the various APOE genotypes. In conclusion, the APOE genotype may affect the response of serum cholesterol to dietary saturated fat and cafestol in opposite directions. However, the effects are small. Therefore, knowledge of the APOE genotype by itself may be of little use in the identification of subjects who respond to diet. PMID- 11257256 TI - Fh-Souassi: a founder frameshift mutation in exon 10 of the LDL-receptor gene, associated with a mild phenotype in Tunisian families. AB - Familial hypercholesterolemia (FH) has a higher prevalence in central Tunisia together with a milder clinical expression than in western countries. The molecular basis of FH in Tunisia remains unknown. Our aim was to identify FH causing mutations in three unrelated families (21 subjects) from the area of Souassi (central Tunisia). In probands with a presentation of homozygous FH, the promoter and 18 exons of the low density lipoprotein (LDL)-receptor gene were sequenced in both orientations. A novel complex frameshift mutation was identified in exon 10, nucleotides 1477-1479 (TCT) at Serine 472 were replaced by an insertion of seven nucleotides (AGAGACA), producing a premature termination codon 43 amino acids downstream. Binding of 125I-labelled LDL at 4 degrees C to cultured fibroblasts from two probands showed <2% normal LDL-receptor activity. AvaII digestion of PCR amplified genomic DNA identified this unique mutation in all families; homozygotes n=11, heterozygotes n=10. All mutation carriers shared the same haplotype (7 RFLPs), suggesting that they had a common ancestor. Despite high plasma LDL levels (m=16.0+/-3.0 mmol/l) and extravascular cholesterol deposits, most homozygotes were diagnosed after puberty and had a delayed onset of cardiovascular complications. Moreover, most heterozygotes were free of clinical signs and had plasma LDL cholesterol in the normal range (4.7+/-1.3 mmol/l) without taking any lipid-lowering medication. This mild clinical phenotype which contrasted with the severity of the mutation, could not be explained by specific apolipoprotein E or lipoprotein lipase alleles. PMID- 11257257 TI - Sequencing of the coding exons of the LRP1 and LDLR genes on individual DNA samples reveals novel mutations in both genes. AB - Five coding polymorphisms in de LRP1 gene, i.e. A217V, A775P, D2080N, D2632E and G4379S were discovered by sequencing its 89 exons in three test-groups of 22 healthy individuals, 29 Alzheimer patients and 18 individuals with different clinical and molecularly uncharacterized lipid metabolism problems. No genetic defect was evident in the LRP1 gene of any of the Alzheimer's disease (AD) patients, further excluding LRP1 as a major genetic problem in AD. Lipoprotein receptor related protein (LRP) A217V (exon 6) was clearly present in all groups as a polymorphism, while D2632E was observed only once in a healthy volunteer. On the other hand, LRP1 alleles A775P, D2080N, and G4379 were encountered only in patients with FH or with undefined problems of lipid metabolism. This finding forced one to also analyze the LDL receptor (LDLR) gene, for which a method was devised to sequence the entire region comprising LDLR exons 2-18. The resulting sequence contig of 33567 nucleotides yielded finally an exact physical map that corrects published and listed LDLR gene maps in many positions. In addition, next to known mutations in LDLR that cause FH, four novel LDLR defects were defined, i.e. del e7-10, exon 9 mutation N407T, a 20 bp insertion in exon 4, and a double mutation C292W/K290R in exon 6. No evidence for pathology connected to the LRP1 'mutations' was obtained by subsequent screening for the five LRP1 variants in larger groups of 110 FH patients and 118 patients with molecularly undefined, clinical problems of cholesterol and/or lipid metabolism. In three individuals with a mutant LDLR gene a variant LRP1 allele was also present, but without direct, obvious clinical compound effects, indicating that the variant LRP1 alleles must, for the present, be considered polymorphisms. PMID- 11257259 TI - A new DNA polymorphism in the 5' untranslated region of the human SREBP-1a is related to development of atherosclerosis in high cardiovascular risk population. AB - Sterol-regulatory element binding proteins (SREBPs) are ubiquitous transcription factors that regulate the genes encoding key proteins in the control of cholesterol homeostasis. We looked for mutations or polymorphisms within the sequences of the SREBP-1a gene critical for the synthesis and/or activity of the protein in 204 asymptomatic men. A single G deletion at base pair -36 of the translation initiation site (designated G-) was found using single-strand conformation polymorphism (SSCP), in addition to three rare variants. This new marker was then assessed for its influence on the lipid parameters of 812 men at high cardiovascular risk, and on the presence of echographic atherosclerotic plaque in their peripheral arteries. The allelic frequency of the -36delG polymorphism was 0.58. At least one plaque was found in the carotid in 24% of subjects, in the femoral arteries of 48%, and in the aorta of 25%. There were significant associations between the -36delG polymorphism and mean total cholesterol (p=0.02) and LDL-cholesterol (P=0.02). There was a graded relationship between the G- allele and the presence of carotid plaque (r=0.084, P=0.02). In addition, there was a statistically significant interaction between the -36delG genotype and the apoE phenotype for plasma LDL-cholesterol (P=0.04) and apoB (P=0.05), suggesting a gene-gene interaction. Stepwise multiple regression analysis for lipid traits, risk factors, and apoE phenotype showed an independent association between carotid plaque and the -36delG polymorphism (beta=0.311, P=0.03). Thus, we have identified a new polymorphism in the 5' untranslated region of the SREBP-1a gene, and demonstrated its association with an atherogenic lipid profile and echographic plaques. PMID- 11257258 TI - Variable association between genetic variation in the CYP7 gene promoter and plasma lipoproteins in three Canadian populations. AB - The promoter sequence variant -278A in the CYP7 gene, which encodes cholesterol 7 alpha hydroxylase, was previously reported to be associated with reduced plasma low density lipoprotein (LDL) cholesterol concentration. We tested for association of CYP7-278A with plasma lipoprotein traits in samples taken from three distinct Canadian populations: 594 Alberta Hutterites, 325 Ontario Oji-Cree and 190 Keewatin Inuit. The CYP7-278A allele frequencies in these three groups were 0.708, 0.466 and 0.490, respectively. The frequencies of CYP7-278A/A homozygotes were 0.481, 0.215 and 0.247, respectively. In the Hutterites, CYP7 278A was associated with reduced plasma HDL-cholesterol and apolipoprotein AI concentration. In the Oji-Cree, CYP7-278A was not significantly associated with any plasma lipoprotein trait. In the Inuit CYP7-278A was associated with elevated plasma total and LDL-cholesterol. There was no consistent relationship between the population mean plasma LDL-cholesterol concentration and the population CYP7 278A frequency. Our findings suggest that the common -278A promoter variant of CYP7 was inconsistently associated with variation in plasma LDL- and HDL cholesterol in samples from three independent populations. The inconsistencies could be due to differences in genetic background or to unspecified environmental or genetic factors. PMID- 11257260 TI - A point mutation in ABC1 gene in a patient with severe premature coronary heart disease and mild clinical phenotype of Tangier disease. AB - The proband is a 50 year-old woman born from a consanguineous marriage. She has been suffering from angina pectoris since the age of 38 and underwent coronary bypass surgery for three-vessel disease at 48. The presence of low plasma levels of total cholesterol and high density lipoprotein (HDL) cholesterol (2.4 and 0.1 mmol/l) and apo AI (<15 mg/dl), associated with corneal lesions and a mild splenomegaly suggested the diagnosis of Tangier disease. However, none of the other features of Tangier disease, including hepatomegaly, anemia and peripheral neuropathy, were present. The analysis of the dinucleotide microsatellites located in chromosome 9q31 region demonstrated that the proband was homozygous for the alleles of D9S53, D9S1784 and D9S1832. The mother and son of the proband, both with low levels of HDL cholesterol, shared one of the proband's haplotypes, whereas neither of these haplotypes was present in the normolipidemic proband's sister. The sequence of ATP-binding cassette transporter 1 (ABC1-1) cDNA obtained by reverse transcription-PCR (RT-PCR) of total RNA isolated from cultured fibroblasts showed that the proband was homozygous for a C>T transition in exon 13, which caused a tryptophane for arginine substitution (R527W). This mutation was confirmed by direct sequencing of exon 13 amplified from genomic DNA. It can be easily screened, as the nucleotide change introduces a restriction site for the enzyme Afl III. R527W substitution occurs in a highly conserved region of the NH2 cytoplasmic domain of ABC1 protein. R527W co-segregates with the low HDL phenotype in the family and was not found in 200 chromosomes from normolipidemic individuals. PMID- 11257261 TI - Common variants in the gene encoding ATP-binding cassette transporter 1 in men with low HDL cholesterol levels and coronary heart disease. AB - HDL cholesterol (HDL-C) deficiency is the most common lipid abnormality observed in patients with premature coronary heart disease (CHD). Recently, our laboratory and others demonstrated that mutations in the ATP-binding cassette transporter 1 (ABCA1) gene are responsible for Tangier disease, a rare genetic disorder characterized by severely diminished plasma HDL-C concentrations and a predisposition for CHD. To address the question of whether common variants within the coding sequence of ABCA1 may affect plasma HDL-C levels and CHD risk in the general population, we determined the frequencies of three common ABCA1 variants (G596A, A2589G and G3456C) in men participating in the Veterans Affairs Cooperative HDL Cholesterol Intervention Trial (VA-HIT), a study designed to examine the benefits of HDL raising in men having low HDL-C (< or =40 mg/dl) and established CHD, as well as in CHD-free men from the Framingham Offspring Study (FOS). Allele frequencies (%) in VA-HIT were 31, 16, and 4 for the G596A, A2589G, and G3456C variants, respectively, versus 27, 12, and 2 in FOS (P<0.03). None of the variants were significantly associated with plasma HDL-C concentrations in either population; however, in VA-HIT, the G3456C variant was associated with a significantly increased risk for CHD end points, suggesting a role for this variant in the premature CHD observed in this population. PMID- 11257262 TI - Enhanced fractional catabolic rate of apo A-I and apo A-II in heterozygous subjects for apo A-I(Zaragoza) (L144R). AB - We have recently reported a new apolipoprotein (apo) A-I variant (apo A I(Zaragoza) L144R) in a Spanish family with HDL-C levels below the 5th percentile for age and sex and low apo A-I concentrations. All the apo A-I(Zaragoza) subjects were heterozygous and none of them showed evidence of coronary artery disease (CAD). Mean plasma HDL-C, apo A-I, and apo A-II levels were lower in apo A-I(Zaragoza) carriers as compared to control subjects (40, 60, and 50%, respectively). Lipid composition analysis revealed that apo A-I(Zaragoza) carriers had HDL particles with a higher percentage of HDL triglyceride and a lower percentage of HDL esterified cholesterol as compared to those of control subjects. Lecithin:cholesterol acyltransferase (LCAT) activity and cholesterol esterification rate of apo A-I(Zaragoza) carriers were normal. Apo A-I and apo A II metabolic studies were performed on two heterozygous apo A-I(Zaragoza) carriers and on six control subjects. We used a primed constant infusion of [5,5,5-2H3]leucine and HDL apo A-I and apo A-II tracer/tracee ratios were determined by gas chromatography mass spectrometry and fitted to a monoexponential equation using SAAM II software. Both subjects carrying apo A I(Zaragoza) variant showed mean apo A-I fractional catabolic rate (FCR) values more than two-fold higher than mean FCR values of their controls (0.470+/-0.0792 vs. 0.207+/-0.0635 x day(-1), respectively). Apo A-I secretion rate (SR) of apo A I(Zaragoza) subjects was slightly increased compared with controls (17.32+/-0.226 vs. 12.76+/-3.918 mg x kg(-l) x day(-1), respectively). Apo A-II FCR was also markedly elevated in both subjects with apo A-I(Zaragoza) when compared with controls (0.366+/-0.1450 vs. 0.171+/-0.0333 x day(-1), respectively) and apo A-II SR was normal (2.31+/-0.517 vs. 2.1+/-0.684 mg x kg(-l) x day(-1), respectively). Our results show that the apo A-I(Zaragoza) variant results in heterozygosis in abnormal HDL particle composition and in enhanced catabolism of apo A-I and apo A II without affecting significantly the secretion rates of these apolipoproteins and the LCAT activation. PMID- 11257263 TI - Hepatic lipase promoter activity is reduced by the C-480T and G-216A substitutions present in the common LIPC gene variant, and is increased by Upstream Stimulatory Factor. AB - The common -216G-->A and -480C-->T substitutions in the promoter region of the human hepatic lipase (LIPC) gene show high allelic association, and are correlated with decreased hepatic lipase activity and increased high-density lipoprotein cholesterol levels. To test the functionality of these substitutions, CAT-reporter assays were performed in HepG2 cells. LIPC (-650/+48) but not ( 650/+61) promoter constructs showed transcriptional activity. LIPC (-650/+48) constructs with both -216A and -480T exhibited significantly lower promoter activity (-45%) than the wild-type form. Activities of -289/+48 constructs were not significantly affected by the -216G-->A substitution. The -480C/T site lies within a binding region for Upstream Stimulatory Factor (USF). Gel-shift assays showed that the binding affinity of USF protein for HL specific oligonucleotides was decreased four-fold by the -480C-->T substitution. However, promoter activity of the -650/+48 constructs was not significantly affected by the -480C-->T substitution alone. Co-transfection of HepG2 cells with USF(43) cDNA yielded a similar dose-dependent increase in activity of all -650/+48 constructs; the absolute difference in promoter activity increased but the relative difference between the variant promoter forms was maintained. Our studies demonstrate that the common LIPC promoter variation is functional, which explains the association of the -480T allele with a lower hepatic lipase activity in man. PMID- 11257264 TI - A prospective study of paraoxonase gene Q/R192 polymorphism and severity, progression and regression of coronary atherosclerosis, plasma lipid levels, clinical events and response to fluvastatin. AB - Human serum paraoxonase (PON1) is a high-density lipoprotein (HDL)-associated enzyme that is responsible for the protective effect of HDL against oxidation of low-density lipoprotein (LDL). PON1 has a Glu to Arg polymorphism at codon 192 (CGA-->CAA) which is designated R/Q192. The R/Q192 polymorphism has been associated with coronary artery disease (CAD) in several, but not all, case control studies. We prospectively studied the association of the Q/R192 genotypes with the severity, progression and regression of CAD, plasma lipid levels, clinical events and response to treatment with fluvastatin in a well characterized cohort. Genotypes were determined by polymerase chain reaction (PCR) and restriction mapping with AlwI enzyme in 356 subjects in the Lipoprotein and Coronary Atherosclerosis Study (LCAS). Fasting plasma lipids were measured and quantitative coronary angiograms were obtained at baseline and 2.5 years following randomization to fluvastatin or placebo. A total of 177 (50%), 142 (40%) and 37 (10%) subjects had Q/Q, Q/R and R/R genotypes, respectively. Baseline and final plasma levels of HDL, LDL, triglyceride and other lipoproteins, lesion-specific minimum lumen diameters (MLD), mean MLD, number of coronary lesions and total occlusions at baseline and follow-up and clinical event rates were not significantly different among the genotypes. There was no genotype-treatment interaction with respect to plasma lipid levels and angiographic indices of CAD. The Q/R192 variants of PON1 are not associated with severity, progression or regression of coronary atherosclerosis, plasma lipid levels, clinical events, or response to treatment with fluvastatin. Thus, the Q/R192 polymorphism is not a major risk factor in susceptibility to CAD in the LCAS population. PMID- 11257265 TI - PON2 gene variants are associated with clinical manifestations of cardiovascular disease in familial hypercholesterolemia patients. AB - Paraoxonase is an enzyme associated with the high-density lipoprotein (HDL) particle. It catalyses the hydrolysis of organophosphates and protects LDL from oxidative modification in vitro by hydrolyzing lipid peroxides, suggestive of a role for paraoxonase in the development of atherosclerosis. Two frequent mutations at the paraoxonase gene locus (PON1) underlie the leucine (Leu allele) -> methionine (Met allele) and the glutamine(Gln allele) --> arginine(Arg allele) aminoacid substitutions at residues 55 and 192, respectively. These polymorphisms have been associated with increased risk for cardiovascular disease (CVD) in several studies, while others have not found this association. Recently, another member of the PON gene family designated PON2 has been identified. While the PON2 gene product is expressed ubiquitously, its physiological role is unknown. A common polymorphism at codon 311 (Cys-->Ser) in the PON2 gene has been described. In our study we assessed the frequency and genotype distribution of the PON1 and PON2 polymorphisms in 197 patients with familial hypercholesterolemia (FH), to determine the possible association between these mutations and susceptibility for CVD. The FH cohort group was divided into subjects with (n=83) and without (n=114) definite clinical manifestations of CVD (FH-Symptomatic and FH Asymptomatic respectively). The control population consisted of 201 healthy normolipidemic blood donors. All subjects in this study were of Caucasian background. Genotypes were identified by PCR based analysis. With regard to the PON1 polymorphisms 55 and 192, no different distributions of allele frequencies were found between the groups studied. However, we did show an association between the PON2 311 polymorphism and CVD. The frequencies of PON2 Ser311 carriers (Ser/Ser and Cys/Ser) between FH-Symptomatic and both FH-Asymptomatic and controls did show a significant difference (P=0.01 and P=0.02 respectively). In the FH-Symptomatic population, surprisingly, no subjects were homozygous for PON2 Cys311, whereas in the FH-Asymptomatic population nine persons (7.9%) and in the control group 12 persons (6.0%) were homozygous. Our data indicate that the common PON2 polymorphism is associated with clinical manifestations of CVD in FH patients. While PON2 Ser311 carriers seem to be at risk, subjects with the Cys/Cys311 genotype are likely to be protected against the development of premature CVD. PMID- 11257266 TI - Identification of six methylenetetrahydrofolate reductase (MTHFR) genotypes resulting from common polymorphisms: impact on plasma homocysteine levels and development of coronary artery disease. AB - Although three common MTHFR polymorphisms (C677T, A1298C, T1317C) have been reported, only polymorphism C677T has been investigated intensively as a risk factor for coronary artery disease (CAD). We investigated polymorphism frequencies, allelic associations and the effect of the resulting MTHFR genotypes on total plasma homocysteine (tHcy) levels and on coronary risk in a case-control study with 1000 angiographically confirmed Middle-European CAD patients and 1000 matched controls. Three out of four theoretically possible MTHFR haplotypes were detected: *1 (677C, 1298A), *2 (677T, 1298A), and *3 (677C, 1298C). The frequencies were *1: 36.4 and 34.4%; *2: 30.8 and 32.3%; and *3: 32.8 and 33.3%, in cases and controls, respectively. Only one patient was heterozygous for 1317C. None of the six resulting genotypes showed significant influence on tHcy levels. Moreover, there was no significant association with CAD risk or with disease severity or early disease manifestation. In the subgroup presenting with acute coronary syndromes, MTHFR genotypes *2/*3 and *3/*3 were surprisingly underrepresented (relative risk of *3/*3, 0.40; 95% confidence interval 0.20 0.79, P=0.009). We conclude from our genotype-based analysis that, in this well fed Middle-European population, the observed common allelic variants of the MTHFR gene have no significant influence on tHcy levels or on the chronic process of CAD development. PMID- 11257267 TI - The effect of the C677T and A1298C polymorphisms in the methylenetetrahydrofolate reductase gene on homocysteine levels in elderly men and women from the British regional heart study. AB - Total blood levels of homocysteine (tHcy) have been shown to depend on both environmental and genetic factors, and to be associated with the risk of developing atherosclerosis with its complications of coronary heart disease (CHD) and stroke. In this study, 408 men and 346 women from two towns, Dewsbury and Maidstone were examined for tHcy levels and genotyped for the C677T and the A1298C polymorphisms in the methylenetetrahydrofolate reductase (MTHFR) gene. Blood tHcy was significantly higher in men from the CHD high risk town of Dewsbury (12.7 micromol/l) than in the low CHD risk town of Maidstone (11.5 micromol/l) P<0.001, but not in women (10.7 vs. 10.5 micromol/l), with women in both towns, thus, showing significantly lower tHcy than men. There was no difference between towns in folate or vitamin B12 levels but the conventional inverse relationship with tHcy was seen. Smoking men and women from both towns had significantly higher tHcy and lower folate levels than non-smoking individuals (P<0.001). The frequency of the 677T allele in Dewsbury was 0.35 (95% CI; 0.32-0.39) compared with 0.29 (95% CI; 0.26-0.32) in Maidstone (P<0.01). Similar frequency difference of borderline statistical significance was seen both for men (P=0.054) and women (P=0.048) in both the towns, suggesting a true regional frequency difference. The effect of the 677T on tHcy was highly significant in the group as a whole with the most profound effect seen in men (12.0 micromol/l for CC vs. 14.1 micromol/l for TT, P<0.001). By contrast, there was no significant effect of the A1298C polymorphism on tHcy, folate or vitamin B12 levels, with no evidence for an interaction with the C677T genotype. The regional differences in tHcy levels were still present after the adjustment for folate and vitamin B12 levels, smoking and the effect of the C677T polymorphism. This suggests that there may be other unidentified factors, either environmental or genetic, affecting tHcy levels, and thus potentially having an impact on the risk of developing hyperhomocysteinaemia and CHD. These observations may have a bearing on regional differences in tHcy levels and the variation in CHD risk between regions in the UK. PMID- 11257268 TI - Influence of a methionine synthase (D919G) polymorphism on plasma homocysteine and folate levels and relation to risk of myocardial infarction. AB - Methionine synthase (MS) encodes an enzyme that catalyzes the remethylation of homocysteine to methionine using a methyl group donated by 5 methyltetrahydrofolate, which is the major circulating form of folate in the body. Functional genetic variants of the MS may alter total homocysteine (tHcy) as well as folate levels which are independent risk factors for vascular disease. The influence of a common genetic polymorphism (2756A-->G, D919G) of the MS gene on plasma tHcy and folate levels and its relation to the risk of myocardial infarction (MI) in a prospective study of male physicians in the US was investigated. A nested case-control study was conducted within the Physicians' Health Study which was originally designed as a double-blind trial of aspirin and beta-carotene among 22071 US male physicians, aged 40-84 years in 1982. Sixty eight percent of participants also donated a blood sample. The study included 387 incident MI case and 767 controls matched on age, smoking status, and time from randomization in 6-month intervals. Individuals with GG genotype had a non significant reduction of MI risk (RR 0.51, 95% CI 0.17-1.16) compared to individuals with DD genotype after adjusting for MI risk factors. The MS polymorphism was associated with decreased tHcy (10.55, 9.87 and 9.57 nmol/ml for DD, DG and GG genotypes, respectively) and increased folate levels (3.95, 3.78, 7.31 ng/ml for DD, DG and GG genotypes, respectively) only among controls but not cases. It was concluded that influence of the MS (D919G) polymorphism on the plasma tHcy and folate levels is at most moderate, but should be further investigated in other large prospective studies. PMID- 11257269 TI - Angiotensin-converting enzyme gene polymorphism in a cohort of coronary angiography patients. AB - An association between a polymorphism of the angiotensin-converting enzyme (ACE) gene and myocardial infarction (MI) in men has been previously reported. The present study examines the association between ACE genotype, atherosclerosis, MI, hypertension and other cardiovascular risk factors in Caucasian men (n=576) and women (n=124) who have undergone coronary angiography. Gene frequencies are also reported for African-American men (n=56). Genotype determination was based on the presence (allele I) or absence (allele D) of a 287 nucleotide Alu sequence in intron 16 of the ACE gene. Genotype frequencies for DD, ID and II were: 30.9, 47.7, 21.4% for Caucasian men; 28.2, 48.4, 23.4% for Caucasian women; and 30.4, 46.4, 23.2% for African-American men. There were no statistically significant associations between ACE genotype and number of plaques (> or =10% obstruction), lipid variables, or body mass index (BMI) for Caucasian men. Caucasian women with the DD genotype had on average fewer plaques, but this was accounted for by their younger ages. In Caucasian males, the DD genotype independently contributed to the presence of hypertension (odds ratio=1.8, 95% CI 1.1-2.9) after adjusting for age and BMI. In Caucasian males with total cholesterol levels less than 200 mg/dl (n=237), the DD (odds ratio=2.5, 95% CI 1.2-5.4) and ID genotypes (odds ratio=2.2, 95% CI 1.1-4.4) were associated with a history of MI. PMID- 11257270 TI - Familial and genetic determinants of systemic markers of inflammation: the NHLBI family heart study. AB - Inflammation is thought to play a central role in the etiology and outcome of atherosclerosis. Animal studies as well as in vitro and in vivo human studies suggest that host factors modulate the magnitude and extent of inflammatory responses. We investigated familial aggregation of three systemic markers of inflammation (C-reactive protein (CRP), white blood cell count (WBC), and albumin) in a large, cross-sectional study conducted in four US communities. We found evidence of substantial heritability (35-40%) for CRP levels as well as for WBC and albumin levels. Negligible spouse correlations suggested little influence of shared household environment on these traits. The combination of sociodemographic factors (age, center, education), behavioral and lifestyle factors (cigarette smoking, alcohol intake, hormone replacement therapy), obesity and fat patterning, and prevalent diabetes explained 13-30% the interindividual variability of these traits. There was no evidence that these inflammation phenotypes were linked to a microsatellite marker in the interleukin-1 gene cluster on chromosome 2q, a region that includes several candidate genes for chronic inflammatory diseases. Our findings suggest that CRP levels, albumin levels, and WBC are determined at least partially by genetic factors. Further efforts to identify gene loci affecting these traits are warranted. PMID- 11257271 TI - Polymorphisms within the tumor necrosis factor locus and prevalence of coronary artery disease in middle-aged men. AB - Tumor necrosis factor (TNF) is an important cytokine in the inflammation process of atherosclerosis and is also involved in lipid metabolism. Two biallelic polymorphisms within TNF gene locus-TNFA at the position -308 in the promoter region of the TNF gene and TNFB in the first intron of the lymphotoxin-alpha (LT alpha) have been reported to be associated with TNF production and with susceptibility to inflammatory diseases. We studied the association of these polymorphisms within the major histocompatibility complex (MHC) III region with coronary atherosclerosis and its manifestations. The autopsy series comprised 700 Caucasian Finnish men, aged 33-70 years (The Helsinki Sudden Death Study). Coronary stenosis and surface area of atherosclerotic changes (fatty streaks, fibrous plaques, complicated lesions and calcification) were measured and the presence of myocardial infarction and coronary thrombosis recorded. TNFA and TNFB genotypes were determined by the PCR-RFLP technique. The allele frequencies were TNFA1/TNFA2=0.88/0.12 and TNFB1/TNFB2=0.30/0.70. There was a strong linkage disequilibrium between the two polymorphisms. There were no differences in coronary stenosis and in the frequency of old or recent myocardial infarction or coronary thrombosis between men with different genotype status in either locus. Men with TNFA22 or TNFB11 genotype tended to have more fibrous lesions and calcification in their coronary arteries. TNFA and TNFB polymorphisms are unlikely to contribute to progression of atherosclerosis in a way clinically important. PMID- 11257272 TI - A prospective evaluation of the CD14 C(-260)T gene polymorphism and the risk of myocardial infarction. AB - The T allele at position -260 of the CD14 lipopolysaccharide receptor gene (CD14) has recently been hypothesized to be a risk factor for myocardial infarction (MI). However, no prospective data relating this polymorphism to risk of future MI are available. In the physicians' health study (PHS), 14916 apparently healthy men were followed over a 12-year period for incident MI. Employing a nested case control study design, the CD14 C(-260)T polymorphism was evaluated among 387 study participants who developed MI (cases) and among an equal number of age- and smoking-matched study participants who remained free of vascular diseases during follow-up (controls). All observed genotype frequencies were in Hardy-Weinberg equilibrium. However, the allele and genotype distributions of the CD14 polymorphism were similar among cases and controls, both in the total cohort and in all subgroups evaluated. Furthermore, no evidence of association was observed assuming additive, dominant, or recessive mode of inheritance. For example, the relative risk of future MI in a comparison of homozygous mutants to homozygous wild types was 1.00 (95% CI=0.7-1.5; P=0.9). In this large prospective study, the CD14 C(-260)T gene polymorphism was not associated with risks of future MI. Thus, in contrast to prior studies, these data indicate that screening for CD14 C( 260)T genotypes is unlikely to be a useful tool for risk assessment. PMID- 11257273 TI - Polymorphisms in the thrombopoietin gene are associated with risk of myocardial infarction at a young age. AB - Five polymorphisms in the thrombopoietin (TPO) gene were identified, one in the 5' untranslated region (UTR) (C1796T), two within intron 5 (C4830A and A4877C), and two in the 3' UTR (A5713G and A6160T). The allele frequencies were determined in a group of 450 healthy middle aged men from the UK and found to be 0.46 for 1796T, 0.38 for 4830A, 0.004 for 4877C, 0.47 for 5713G and 0.07 for 6160T. Genotypes for the three common polymorphisms were determined in a group of 176 young male Swedish survivors of a myocardial infarction (MI) and 186 age-matched controls and a group of 156 young Italian survivors of an MI and 147 age and sex matched controls. In both the Swedish and the Italian studies polymorphisms were found to be associated with increased risk of MI. In the Swedish sample the frequency of 4830A was significantly higher in controls (0.40) compared with patients (0.29) (P=0.003), with an odds ratio for AA homozygotes of 0.48 (0.25 0.92; P=0.03) compared with CC homozygotes. In the Italian sample the frequency of 5713G was significantly lower in controls (0.31) compared with cases (0.40) (P=0.03), with an odds ratio for GG homozygotes of 2.29 (1.08-4.89; P=0.03) compared with AA homozygotes. These risk associations are consistent since 4830A and 5713A show strong allelic association. After adjusting for other measured risk factors the effect on risk was still significant in the Italian sample 2.39 (1.02-5.58), but not in the Swedish sample 0.46 (0.16-1.32). The observation of frequency differences between cases and controls in two independent samples strongly suggests that the TPO gene is involved as a risk factor for developing MI at a young age, but the identified polymorphisms are probably acting as markers for an unidentified functional mutation elsewhere in the gene locus. PMID- 11257274 TI - Functional mutation in the promoter region of thrombomodulin gene in relation to carotid atherosclerosis. AB - Thrombomodulin is an important endothelial anticoagulant protein that decreases thrombin activity and activates protein C. Our recent study has shown that the G 33A promoter mutation of thrombomodulin gene is associated with coronary artery disease. This study was conducted to determine whether the G-33A mutation in the promoter region of thrombomodulin gene is a genetic risk factor for ischemic stroke or carotid atherosclerosis. The functional significance of this mutation was also evaluated. We recruited 333 patients (mean age 64 years, 59% male) with ischemic stroke and 257 age- and sex-matched controls. In all study participants, carotid atherosclerosis was assessed by Duplex scanning, and thrombomodulin G-33A promoter mutation was detected by single-strand conformation polymorphism. Luciferase reporter gene assay was used to assess the influence of this mutation on thrombomodulin promoter activity. There was no significant difference in the thrombomodulin G-33A mutation frequency (GA+AA genotypes) between the stroke and the control groups (18.3 vs. 24. 1%, P=0.105). The G-33A mutation frequency was also similar between the study participants with and without carotid atherosclerosis (22.2 vs. 19.8%, P=0.550). When only younger subjects (age 0.05) in attractive force between the Fe-36 at%Pt alloy specimens and the stainless steel keepers for both magnets. A definite correlation between Pt percentage and the value of saturation magnetization was also found (r(2)=-1.000). SIGNIFICANCE: The Fe-Pt alloys with less than Fe-39.5 at%Pt produced high saturation magnetization values and great attractive force to the magnet, and thus, they have the potential to serve as magnetic attachment keepers. Of the Fe-Pt alloys tested, Fe-36 at%Pt seemed to be the best composition for making magnetic attachment keepers. PMID- 11257293 TI - A survey of the use of dentin-bonding systems in Denmark. AB - OBJECTIVES: Dentin-bonding systems, which usually involve multistep procedures, are relatively new in dentistry. This study was conducted to survey dental practitioners in Denmark concerning their use of dentin-bonding systems. METHODS: A questionnaire regarding dentin-bonding systems was distributed to dentists at an annual dental conference, and 462 dentists were included in the survey. RESULTS: Dentin-bonding systems were used by 99% of the dentists. Twenty-one different dentin-bonding systems were being used, with six of the systems being used by 78% of the dentists. Of the 456 dentists who used a dentin-bonding system, 77% recalled a clinical procedure for their dentin-bonding system that was in accordance with the written instructions of the manufacturer. The degree to which the dentists complied with the instructions for use, was influenced by the number of operating steps involved for a given dentin-bonding system, by the frequency with which the dentin-bonding system was used, and by the degree to which the dentist was satisfied with the instructions for use provided by the manufacturer. The frequency with which dentin-bonding systems were used, was influenced by year of graduation, place of work, and gender of the dentist. SIGNIFICANCE: Further studies, which focus on the adherence of dental practitioners to instruction manuals are warranted as are investigations of the potential clinical consequences of incorrect use of dentin-bonding systems. PMID- 11257292 TI - Mode of failure in the dentin-adhesive resin-resin composite bonded joint as determined by strength-based (muTBS) and fracture-based (CNSB) mechanical testing. AB - OBJECTIVE: To determine the failure mode between dentin-adhesive resin-resin composite bonded joint produced with a chevron-notch short-bar (CNSB) and microtensile test methods. METHODS: Forty teeth were randomly selected for microtensile and forty for CNSB specimen fabrication and stored in 0.5% chloramine T at 37 degrees C until respective static load to failure testing at 30 and 180days. Failure modes were categorized by SEM and tested with Fisher's exact test. Within respective mechanical testing methods the probability of failure curve distributions being significantly different were analyzed by the Wald chi-square statistic. RESULTS: The characteristic fracture toughness at 30- and 180-day storage was 0.82 and 0.87MPam(1/2), while the Weibull Modulus (m) for the failure distributions, was 4.60 and 4.56, respectively. No significant difference was demonstrated in the failure distributions between these groups (p=0.45). The characteristic tensile strength (muTBS(o)) at 30- and 180-day storage was 52.53 and 14.71MPa with an m of 3.04 and 1.56, respectively. Failure distributions for muTBS groups were significantly different (p<0.001). K(IvM) failure modes, regardless of storage time, were within the adhesive joint with 30 day debonds primarily through the top region of the hybrid layer (THL) and after 180-days involving the bottom of the hybrid layer (BHL). The 30-day muTBS group demonstrated a propensity to debond in dentin or resin composite substrates but after 180-days storage debonds again involved the BHL. SIGNIFICANCE: The weak links in the dentin-adhesive resin-resin composite bonded joint may be the interphase regions between the THL and the adhesive resin and the BHL and dentin. PMID- 11257294 TI - In vitro corrosion behavior of cast iron-platinum magnetic alloys. AB - OBJECTIVE: The objective of this study was to investigate the corrosion resistance of cast Fe-Pt alloys of varying compositions for use as attachment keepers and to make a comparison with the corrosion resistance of magnetic stainless steel. METHODS: The corrosion behavior of cast Fe-Pt alloy keepers (Fe 40 at%Pt, Fe-38 at%Pt, Fe-37 at%Pt and Fe-36 at%Pt) was evaluated by means of an immersion test and an anodic polarization test. The solutions used were a 1.0% lactic acid aqueous solution (pH=2.3) (10 ml) and 0.9% NaCl solution (pH=7.3) (10 ml). As a control, the corrosion resistance of a magnetic stainless steel keeper (SUS 447J1: HICOREX) was also measured. RESULTS: Chromium and platinum ions were not detected in either the 1.0% lactic acid or 0.9% NaCl solutions. The only released ions detected were the Fe ions in the 1.0% lactic acid solution. The amounts of Fe ions released from the Fe-40 at%Pt and Fe-38 at%Pt alloys were significantly (p<0.05) lower than from the Fe-37at%Pt, Fe-36 at%Pt and SUS 447J1 alloys. In the anodic polarization test, the potentials at the beginning of passivation for the four Fe-Pt alloys were higher than for the SUS 447J1 alloy in both solutions. SIGNIFICANCE: The Fe-Pt alloys, especially the alloys with higher Pt percentages (Fe-40 and 38 at%Pt), indicated a high corrosion resistance compared to the magnetic stainless steel keeper. A reduction in the Pt percentage may decrease the corrosion resistance in the oral environment. PMID- 11257295 TI - Primary cyclization in the polymerization of bis-GMA and TEGDMA: a modeling approach to understanding the cure of dental resins. AB - An optimal dental restorative polymeric material would have a homogeneous cross linking density giving it consistent mechanical strength throughout the material. When multifunctional monomers are polymerized, a pendant double bond can react intramolecularly with the radical on its propagating chain to form a loop, which results in a primary cyclization reaction. Primary cyclization does not contribute to overall network structure, causes microgel formation, and leads to heterogeneity in the polymer. Knowledge of how cure conditions control the degree of primary cyclization and cross-linking in the polymer is important in developing better dental materials. To gain more understanding about the evolving polymer network, the photopolymerization of a typical dental resin (75/25 wt% bis GMA/TEGDMA) is modeled using a first principals approach. The overall polymerization rate behavior of 75/25 wt% bis-GMA/TEGDMA is predicted using experimentally obtained propagation and termination kinetic rate constants. The effect of chain stiffness and light intensity on the polymerization kinetics is also explored. Furthermore, the model predicts the extent of cross-linking and primary cyclization in the growing polymer network. At 45% conversion, the fraction of bis-GMA and TEGDMA pendant double bonds created that have cycled is 11 and 33%, respectively. The model shows that using a stiff monomer, like bis GMA, in dental resins diminishes the extent of cyclization and increases the cross-linking density of the polymer. Therefore, better mechanical properties are obtained than if more flexible monomers were used. PMID- 11257296 TI - Response of human pulps capped with a self-etching adhesive system. AB - OBJECTIVE: The aim of this study was to evaluate the human pulp response following direct pulp capping with a current self-etching bonding agent and calcium hydroxide (CH). METHODS: Thirty-three sound human premolars had their pulp tissue mechanically exposed. Sterile distilled water was used to control the hemorrhage and exudation from the pulp exposure site. The pulps were capped with Clearfil Liner Bond 2 (CLB-2) or CH and the cavities were filled with a resin composite (Z-100) according to the manufacturer's instructions. After 5, 30 and 120-300 days, the teeth were extracted and processed for microscopic examination. RESULTS: At short-term, CLB-2 elicited a mild to moderate inflammatory pulp response with dilated and congested blood vessels adjacent to pulp exposure site. With time, macrophages and giant cells engulfing globules and particulates of resinous material displaced into the pulp space were observed. This chronic inflammatory pulp response triggered by fragments of bonding agent displaced into the pulp space did not allow pulp repair interfering with the dentin bridging. On the other hand, pulps capped with CH exhibited an initial organization of elongated pulp cells underneath the coagulation necrosis. Pulp repair and complete dentin bridge formation was observed at long-term evaluation. SIGNIFICANCE: The present study demonstrated that CH remains the pulp capping agent of choice for mechanically exposed human pulps. CLB-2 did not allow complete connective tissue repair adjacent to the pulp exposure site. Consequently, this bonding agent cannot be recommended for pulp therapy of sound human teeth. PMID- 11257297 TI - A predictive formula of the contraction stress in restorative and luting materials attending to free and adhered surfaces, volume and deformation. AB - OBJECTIVES: To find a predictive formula of stress, considering the surfaces (free, adhered) involved, the volume and characteristics of material and the deformation of the measuring system. MATERIALS AND METHODS: 231 samples of five chemically cured restoratives (Silar (SIL, 23), Clearfil F2 (CLE, 39), P10 (P10, 33), Concise (CON, 30), Isopast (ISO, 28)) and four luting (3M Experimental 241 (EXM, 20), Variolink II (VAR, 13), Vitremer LC (VTM, 20) and Dyract Cem (DYR, 25)) materials were allowed to polymerize until they reached a maximum tension (T(max), 25 min) between six pairs (null 5.81, 8.5, 11.26, 12.42, 17.02, 23.14 mm) of polished metallic discs (range of distances: 0.02-5.9 mm) mounted in a tension machine. The deformation of the measuring system was measured for the recorded forces. RESULTS: A descriptive non-linear formula T(max)=KVol( 3.267)FS(3.283)AS(0.642)Def(0.561) was found that individualizes the material's characteristics (K) that considers volume (Vol), free (FS) and adhered (AS) surfaces and deformation (Def) of the system for each force. This formula renders good correlation (material K (r(2) coefficient)): SIL 0.9998 (0.995), CLE 1.0062 (0.989), P10 1.0224 (0.990), CON 0.9908 (0.992), ISO 0.9648 (0.974), EXM 1.0083 (0.991), VAR 0.9777 (0.996), VTM 0.9925 (0.993), DYR 0.9971 (0.997) between actual T(max) and calculated Tension. There are statistically significant differences (p=0.002) between K values of both (restorative and luting) groups. SIGNIFICANCE: Predictive parameters have influence in a different way to what is actually considered, if the system is allowed to have deformation, as occurs naturally and volume and material's characteristics are considered. PMID- 11257299 TI - Polymerization contraction stress in light-cured packable composite resins. AB - OBJECTIVE: Determination of the polymerization contraction stress of packable composites (ALERT, Surefil, Solitaire, Solitaire 2) and a packable ORMOCER material (Definite) in comparison with a conventional hybrid composite (Tetric Ceram). METHODS: Contraction force generated by the test materials (10 replications each) was measured by polymerizing the composites filled in a plastic tray between two aluminum attachments mounted in a Stress-Strain-Analyzer testing machine (specimen size: 4x4x2 mm, C-factor=0.33). Contraction force was recorded for 300s under a standard exposure condition (40s, 800mW/cm(2)). Maximum contraction stress (MPa), force rate (N/s), relative force rate (%/s) of each material were statistically analyzed by ANOVA (alpha=0.05) and post-hoc Tukey's test. RESULTS: Maximum contraction stresses of the packable materials were 4.60 +/- 0.32MPa (ALERT), 4.16 +/- 0.18MPa (Definite), 3.36 +/- 0.08MPa (Solitaire 2), 3.33 +/- 0.23MPa (Solitaire) and 3.13 +/- 0.18MPa (Surefil), which were significantly higher than that of Tetric Ceram (2.51 +/- 0.14MPa). Tetric Ceram exhibited the significantly lowest force rate. Force/time curves were S-shaped. Solitaire especially showed a longer pre-gelation phase before contraction force was recorded. SIGNIFICANCE: High contraction stress and rapid contraction force development can lead to failure of bond to tooth structure. This study suggested that, packable composite resins are less capable of reducing the contraction stress during the early setting stage, thus not superior in maintaining the bond with cavity walls to conventional hybrid composite Tetric Ceram. PMID- 11257298 TI - Measurement of the setting and thermal expansion of dental investments used for the superplastic forming of dental implant superstructures. AB - OBJECTIVES: The study examined the setting and thermal expansions of seven dental casting investments intended for use in the superplastic forming (SPF) of Ti6A14V for dental implant superstructures. The total expansion is compared with the coefficient of thermal expansion of the alloy. METHODS: The unrestricted setting expansion of the investment was measured at room temperature using a lined brass trough and dial gauge. The thermal expansion was measured at a heating rate of 5 degrees C/min up to 900 degrees C. The thermal expansion of Ti6A14V was measured at a heating rate of 15 degrees C/min. RESULTS: The Selevest investment had a small shrinkage on setting. The largest setting expansions were observed with Rema Exakt and Fujivest. Selevest DM and Fujivest had the greatest thermal expansions. Rema Exakt had the greatest overall expansion. The total expansion of Rematitan most closely matched the thermal expansion of Ti6A14V when the manufacturer's recommended special liquid to water ratio for crowns and bridges was used. SIGNIFICANCE: SPF is a novel technique which is being applied to the manufacture of dental prostheses. As dental casting investments are used to make the dies used for SPF the dimensional changes associated with these materials should be matched to the dimensional changes of the Ti6A14V alloy used in order to achieve a passive fit. This study has identified Rematitan as an investment whose total expansion may enable an accurately fitting SPF implant superstructure to be made. PMID- 11257300 TI - Flexural strength of Cerec 2 machined and jointed InCeram-Alumina and InCeram Zirconia bars. AB - OBJECTIVE: The flexural strength of Cerec 2 InCeram-Alumina and InCeram-Zirconia bars is evaluated. The focus of the in vitro study is to identify a jointing procedure for InCeram which may be used for producing full-ceramic fixed-partial denture frameworks. METHODS: Six groups (n=15) of machined and jointed InCeram Alumina (T1-T5) and InCeram-Zirconia (T6) bars (3x4x13mm(3)), respectively, were examined using a 3-point-bending test. InCeram-Alumina joint-free controls were: machined (C1), slip cast (C2, C3) and cut from the block (C4) bars. Machined joint-free InCeram-Zirconia bars were used as controls (C5). InCeram-Alumina slip was used for jointing T1-T5 and InCeram-Zirconia slip for bars T6. Bars were jointed in groups T1 and T2 using butt joint (S1), in T3 and T4 oblique (S2, S3) and in T5 and T6 rounded (S4) joint shapes. RESULTS: Two-way analysis of variance showed significant differences between materials (p<0.001) and jointing shapes (p<0.001). The rounded (S4) shape showed the highest flexural strength of 434 (65) MPa of InCeram-Alumina (T5) and 475 (54) MPa of InCeram-Zirconia (T6) bars, respectively but machined/joint-free InCeram-Alumina (511 (59) MPa, C1) and machined/joint-free InCeram-Zirconia (624 (58) MPa, C5) were significantly (p<0.01/p<0.001) stronger. No significant differences (p>0.05) were found between machined/jointed InCeram-Zirconia (475 (54) MPa, T6), joint-free InCeram-Alumina slip cast (498 (125) MPa, C2) and joint-free InCeram-Alumina machined bars (511 (59) MPa, C1). SIGNIFICANCE: Compared to conventional slip cast InCeram-Alumina the flexural strength of machined/jointed InCeram-Zirconia appears to be adequate for fixed-partial-denture frameworks. PMID- 11257301 TI - The influence of water storage and C-factor on the dentin-resin composite microtensile bond strength and debond pathway utilizing a filled and unfilled adhesive resin. AB - OBJECTIVE: To test the elastic wall concept utilizing adhesive resins of varying stiffness in a low- and high-C-factor cavity design after short- and long-term water storage. METHODS: A flat and box-shaped cavity was restored on occlusal dentin with a resin composite using a filled and unfilled adhesive resin from which microtensile specimens with a 0.5mm(2) cross-sectional area were formed. After storage for 30- and 150-days the microtensile bond strength (muTBS) was determined in a Zwick materials testing machine and the subsequent debond pathway was examined under scanning electron microscopy. Fisher's exact test was used to determine differences in joint and substrate failure modes and a Weibull regression model with gamma frailties was used to test for differences between failure distributions. Tests for three-way and two-way interactions were also completed for storage time, C-factor and adhesive. All tests were at 95% confidence levels. RESULTS: The characteristic strength (TBS degrees ) for the Optibond FL adhesive applied on a flat cavity was 47.57 and 20.90MPa and a box shaped cavity was 49.26 and 17.49MPa for short- and long-term storage, respectively, while the corresponding TBS degrees for the unfilled Optibond adhesive on the flat cavity design was 36.93 and 32.68MPa and in a box-shaped cavity was 32.84 and 15.46MPa. Combining all groups according to storage time revealed a three-fold increase in the debond pathway including the bottom of the hybrid layer. SIGNIFICANCE: Evidence suggests that the durability of the bonded joint is threatened by hydrolysis and the most susceptible region is the bottom half of the hybrid layer and in low C-factor cavity designs a more flexible adhesive resin liner was more durable. PMID- 11257302 TI - Control of the classical and the MBL pathway of complement activation. AB - The activation of complement via the mannan-binding lectin (MBL) pathway is initiated by the MBL complex consisting of the carbohydrate binding molecule, MBL, two associated serine proteases, MASP-1 and MASP-2, and a third protein, MAp19. In the present report we used an assay of complement activation specifically reflecting the physiological activity of the MBL complex to identify biological and synthetic inhibitors. Inhibitor activity towards the MBL complex was compared to the inhibition of the classical pathway C1 complex and to a complex of MBL and recombinant MASP-2. A number of synthetic inhibitors were found to differ in their activities towards complement activation via the MBL pathway and the classical pathway. C1 inhibitor inhibited both pathways whereas alpha2-macroglobulin (alpha2M) inhibited neither. C1 inhibitor and alpha2M were found to be associated with the MBL complex. Upon incubation at 37 degrees C in physiological buffer, the associated inhibitors as well as MASP-1, MASP-2, and MAp19 dissociated from MBL, whereas only little dissociation of the complex occurred in buffer with high ionic strength (1 M NaCl). The difference in sensitivity to various inhibitors and the influence of high ionic strength on the complexes indicate that the activation and control of the MBL pathway differ from that of the classical pathway. MBL deficiency is linked to various clinical manifestations such as recurrent infections, severe diarrhoea, and recurrent miscarriage. On the other hand, impaired control of complement activation may lead to severe and often chronically disabling diseases. The results in the present report suggests the possibility of specifically inhibiting of the MBL pathway of complement activation. PMID- 11257303 TI - Structural analysis of two HLA-DR-presented autoantigenic epitopes: crucial role of peripheral but not central peptide residues for T-cell receptor recognition. AB - Specific and major histocompatibility complex (MHC)-restricted T-cell recognition of antigenic peptides is based on interactions of the T-cell receptor (TCR) with the MHC alpha helices and solvent exposed peptide residues termed TCR contacts. In the case of MHC class II-presented peptides, the latter are located in the positions p2/3, p5 and p7/8 between MHC anchor residues. For numerous epitopes, peptide substitution studies have identified the central residue p5 as primary TCR contact characterized by very low permissiveness for peptide substitution, while the more peripheral positions generally represent auxiliary TCR contacts. In structural studies of TCR/peptide/MHC complexes, this has been shown to be due to intimate contact between the TCR complementarity determining region (CDR) three loops and the central peptide residue. We asked whether this model also applied to two HLA-DR presented epitopes derived from an antigen targeted in type 1 diabetes. Large panels of epitope variants with mainly conservative single substitutions were tested for human leukocyte antigen (HLA) class II binding affinity and T cell stimulation. Both epitopes bind with high affinity to the presenting HLA-DR molecules. However, in striking contrast to the standard distribution of TCR contacts, recognition of the central p5 residue displayed high permissiveness even for non-conservative substitutions, while the more peripheral p2 and p8 TCR contacts showed very low permissiveness for substitution. This suggests that intimate TCR interaction with the central peptide residue is not always required for specific antigen recognition and can be compensated by interactions with positions normally acting as auxiliary contacts. PMID- 11257304 TI - Analysis of diversity of nucleotide and amino acid distributions in the VD and DJ joining regions in Ig heavy chains. AB - Nucleotide fill-in between the germ line V, D and J genes in the H3 loop of immunoglobulins contributes to the diversity of the antibody repertoire. This fill-in process is mediated by terminal deoxynucleotidyl transferase (TdT), which has been widely believed to insert nucleotides in a random fashion. Using a database of 2443 immunoglobulin sequences, we identified the regions of nucleotide fill-in between the V-D and D-J gene regions. We translated the fill in nucleotides and measured the diversity within the two regions both at the nucleotide and amino acid level. We found that the nucleotide and amino acid distributions that resulted from nucleotide fill-in were in fact not random. Examination of the synonymous substitution rates of nucleotides revealed evidence suggesting that TdT plays a less significant role in generating antibody diversity than previously thought. We observed preferences for polar residues, which are more likely to encourage interaction with ligand than non-polar residues and are often found in loop regions in general. We also observed a preference for the insertion of smaller residues versus larger residues of similar biochemical properties, aiding in loop flexibility. We interpret these findings to reflect the significant influence of biochemical (i.e. folding) constraints and/or binding affinity constraints (at the cellular/selectional level) on the sequence diversity in the H3 region. These constraints act as a filter on the randomness generated by nucleotide addition by TdT, as well as other diversity generating processes such as recombination of VDJ gene segments and somatic mutation. The results of this study suggest that the antibody repertoire might be reduced from what is traditionally believed. PMID- 11257305 TI - Human and murine immunoglobulin expression vector cassettes. AB - We describe the construction of new immunoglobulin (Ig) expression vectors and their use in the production of recombinant chimeric Ig molecules in transfected mammalian cells. The vectors contain the cDNA encoding the constant regions of human (mu, alpha1, gammal, gamma2, gamma3, gamma4, kappa) and murine (mu, gamma2a, kappa) Ig heavy and light chains. Unique restriction sites flanking the Ig variable region allow for replacement of variable regions generated by PCR. The CMV promoter allows for the transfection and expression of Ig in non-lymphoid cells. Distinct drug selection markers for heavy chain and light chain expression vectors allows for sequential or co-transfection of the vectors. We show that secretion of recombinant Ig can reach 1.2 microg/ml per million cells per day for transfected B cells. Replacement of the variable region results in the production of functional Ig retaining antigen specificity. PMID- 11257306 TI - Kinetic and genetic bases for the heteroclitic recognition of mouse cytochrome c by mouse anti-pigeon cytochrome c monoclonal antibodies. AB - The B lymphocyte response to pigeon cytochrome c (CYT) in BALB/c mice was previously shown to initiate as a heteroclitic response specific for the self antigen mouse CYT. As the immune response progressed, the mAb that were produced became less heteroclitic and often bound pigeon CYT with higher affinity than they bound mouse CYT [Minnerath, J. et al., 1995. Proc. Natl. Acad. Sci. USA 92, 12379-12383]. To study the basis for heterocliticity and its loss in this system, the H and L chain amino acid sequences of anti-pigeon CYT mAb obtained from the primary and secondary Ab responses were first compared. The most frequent somatic mutations and Ig gene joints were then introduced into an engineered single-chain Fv (scFv) that expressed the germline-encoded V(H) and V(L) amino acid sequences. The effects of those changes on the on- rate, off-rate, and affinity constants in binding both mouse and pigeon CYT were determined by surface plasmon resonance. In this system, the heterocliticity of primary mAb was largely due to a decreased on-rate constant for binding pigeon CYT relative to mouse CYT. Various combinations of the three frequently occurring H chain somatic mutations (H31, H56, and H58) increased the on-rate constant to maximal levels. Only one of the mutations (H58) decreased the off-rate constant when in combination with the other mutations and the effect of H58 was greater for scFv binding mouse CYT than pigeon CYT. Consequently, the mutated scFv and many secondary mAb remained heteroclitic, although their affinities for pigeon CYT increased. Secondary mAb that were no longer heteroclitic expressed non-canonical amino acid sequences in the V(H)-D-J(H) joint in the context of the canonical V genes or expressed different V genes. In addition to providing insight into the molecular basis for heterocliticity, our findings confirm that both faster on-rate and slower off rate constants are favored during affinity maturation of the Ab response. PMID- 11257307 TI - Human CD1d associates with prolyl-4-hydroxylase during its biosynthesis. AB - Recent studies have shown that the CD1 family of proteins present various glycolipid antigens to subsets of T cells. CD1d is expressed on human intestinal epithelial cells (IEC) and exists in two biochemical forms: 37-kDa, beta2 microglobulin (beta2m) independent, nonglycosylated, and 47-kDa, beta2m dependent, glycosylated forms. The biosynthetic pathways and the mechanisms of generation of these two biochemically distinct forms of CD1d in human IEC are unknown. Using a human colonic cell line, T84, transfected with CD1d, the biosynthesis of CD1d was investigated. Pulse-chase metabolic labeling studies of T84 transfected with wild type CD1d demonstrated that CD1d was a stable protein over a 4-day chase period. During the first 24 h of the chase, a novel 65-kDa glycoprotein was co-immunoprecipitated with CD1d. Microsequencing of this protein identified the glycoprotein as the alpha and beta subunits of the resident endoplasmic reticulum protein, prolyl-4-hydroxylase (P4H), an enzyme responsible for hydroxyl modification of proline residues. To study if either one or both biochemical forms of CD1d contained hydroxyproline residues, amino acid composition analysis of the 37 and 48 kDa was performed, and demonstrated that only the 37-kDa, but not the 48-kDa form of CD1d, contained hydroxyproline residues. These studies demonstrate that CD1d exhibits a prolonged association with P4H and that the 37-kDa form contains hydroxyproline residues. This suggests that P4H association with CD1d during its biosynthesis results in a novel post translational modification of CD1d. PMID- 11257308 TI - Agouti-related protein functions as an inverse agonist at a constitutively active brain melanocortin-4 receptor. AB - Agouti-related protein (AGRP) is one of two naturally occurring antagonists of G Protein coupled receptors (GPCRs) identified to date, and has been physiologically implicated in regulating food intake, body weight, and energy homeostasis. AGRP has been identified in vitro, as competitively antagonizing the brain melanocortin-4 (MC4R) and melanocortin-3 (MC3R) receptors, and when over expressed in transgenic mice, results in an obese phenotype. Emerging data propose that AGRP has additional targets in the hypothalamus and/or physiologically functions via a mechanism in addition to competitive antagonism of alpha-MSH at the brain melanocortin receptors. We report data herein supporting an alternative mechanism for AGRP involvement in feeding behavior. A constitutively active MC4R has been generated which possess EC(50) values for melanocortin agonists (alpha-MSH, NDP-MSH, and MTII) and a pA2 value for the synthetic peptide antagonist SHU9119 identical to the wildtype receptor, but increases basal activity to 50% maximal response. AGRP possesses inverse agonist activity at this constitutively active MC4R. These data support the hypothesis for an additional physiological mechanism for AGRP action in feeding behavior and energy homeostasis. PMID- 11257309 TI - The effect of calcitonin gene-related peptide on pancreatic blood flow and secretion in conscious dogs. AB - The effects of human alpha-calcitonin gene-related peptide (alpha-CGRP) and beta CGRP on pancreatic arterial (PA), superior mesenteric (SMA) and left gastric arterial (LGA) blood flows were studied by ultrasound transit-time blood flow meters in five conscious dogs. Intravenous injections of alpha-CGRP and beta-CGRP (5-200 pmol/kg) induced a dose-related increase in PA flow and a dose-related decrease in its resistance. At lower doses, alpha-CGRP was more potent than beta CGRP, but their maximal responses were similar. The blood flow responses to alpha CGRP (200 pmol/kg) were 153% of the basal flow in LGA, 313% in PA, and 534% in SMA, while those to VIP (100 pmol/kg) were 467% in LGA, 953% in PA and 163% in SMA. Somatostatin reduced blood flow in all arteries. alpha-CGRP, but not beta CGRP, at higher doses induced gastric contractions and pancreatic protein-rich secretion, which were blocked by atropine. These results suggest that CGRP in perivascular nerves in the pancreas may regulate pancreatic blood flow in dogs but its physiological function remains to be studied. PMID- 11257310 TI - Plasma levels of N-terminal peptide of pro-atrial natriuretic peptides in myoskeletal injuries. AB - To determine whether concentrations of the N-terminal peptide of pro-atrial natriuretic peptide (proANP) and of alpha atrial natriuretic peptide 1-28 (aANP) releases are affected by myoskeletal injuries, samples from 24 patients with muscle injuries were therefore collected within 48 h of injury. The mean age of patients was 65; range: 17-90 years. These were compared with 18 non-injured subjects with a mean age of 40; range: 17-80 years. A specific enzyme immunoassay (EIA) method suitable for the determination of proANP and aANP was used. aANP required plasma extraction and no extraction was needed for proANP determination.ProANP level was significantly higher in patients on admission and this level was maintained 24 h after admission (p < 0.05) compared to controls. However, aANP 1-28 level remained statistically unchanged in the patients samples. The level of proANP was over 10 times greater than the levels obtained with aANP. N-terminal peptide of proANP may be a supplementary tool in the study of early phase of myoskeletal injuries in human. PMID- 11257311 TI - Atrial natriuretic peptide inhibits tumor necrosis factor-alpha production by interferon-gamma-activated macrophages via suppression of p38 mitogen-activated protein kinase and nuclear factor-kappa B activation. AB - We investigated whether the atrial natriuretic peptide (ANP) might have an inhibitory effect on inflammatory cells. Treatment of RAW264.7 macrophages with interferon-gamma (IFN- gamma) caused a significant increase in tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO) production. Activation of p38 mitogen-activated protein (MAP) kinase was observed 30 to 120 min after IFN gamma, and transcription factor nuclear factor-kappa B (NF-kappaB) was activated about 7 to 9 times of the basal activity. Human ANP(99-126) and a specific p38 MAP kinase inhibitor SB203580 inhibited the IFN-gamma-induced TNF-alpha production in a dose-dependent manner without affecting NO production. ANP inhibited the IFN-gamma-induced p38 MAP kinase activation, and ANP and SB203580 inhibited NF-kappaB activation. To study the involvement of oxidative stress in this system, the effects of allopurinol and acetovanillone, inhibitors of xanthine oxidase and NADPH oxidase, respectively, were studied. Allopurinol or acetovanillone did not inhibit the IFN-gamma-induced production of TNF-alpha or NO, suggesting little involvement of oxidative stress in this system. This is the first evidence in vitro that ANP has an anti-inflammatory activity on IFN-gamma activated macrophages by suppressing signal transduction pathway leading to p38 MAP kinase and NF-kappaB activation. PMID- 11257312 TI - Acute and chronic alterations in blood pressure variability following experimental subarachnoid haemorrhage. AB - This study examined the role of the renin-angiotensin and vasopressin systems on systolic blood pressure (SBP) variability following subarachnoid haemorrhage (SAH) in conscious rats. Animals received no treatment, the angiotensin II AT1 receptor antagonist, losartan, or the vascular vasopressin receptor antagonist, AVPX. SAH resulted in a transient sympathetic activation as estimated from the increase in the mid-frequency oscillations of SBP (3.2 +/- 0.8 mm Hg2, 3 hours after the injury vs. 1.3 +/- 0.3 mm Hg2 in control conditions, p < 0.01). On the second and fourth day following SAH, a marked elevation in the low-frequency component of SBP was observed (7.1 +/- 1.0 mm Hg2 on day 2 vs. 2.6 +/- 0.3 mm Hg2 in control conditions, p < 0.001 and 6.3 +/- 1.1 mm Hg2 on day 4 vs. 2.6 +/- 0.3 mm Hg2 in control conditions, p < 0.01). Pre-treatment with losartan prevented the acute rise in the mid-frequency oscillations in SBP and partially reduced the low-frequency component observed at 2 and 4 days. Administration of AVPX on the second and fourth day following SAH normalised the elevated low-frequency oscillations in SBP. This study indicates that the modifications in SBP variability observed in the early and delayed stage after subarachnoid haemorrhage involve angiotensin II. Vasopressin seems to be implicated in the delayed development of low-frequency fluctuations of SBP. PMID- 11257313 TI - Effect of leptin on insulin sensitivity in the Otsuka Long-Evans Tokushima Fatty rat. AB - Leptin has been proposed to be a sensor of energy storage in adipose tissues, and is capable of mediating a feedback signal to the hypothalamus, which is involved in the regulation of energy homeostasis and body weight. In order to investigate the issue of whether resistance to the activity of leptin on insulin sensitivity is observed in young Otsuka Long-Evans Tokushima Fatty (OLETF) rats at 8 weeks of age, leptin (50 nmol/kg/h) was administered intravenously for 16 h to OLETF and Long-Evans Tokushima Otsuka (LETO) (lean controls) rats, followed by a measurement of insulin-stimulated glucose uptake in hindlimb muscles during hyperinsulinemic euglycemic clamp technique. In the case of LETO rats, the administration of leptin significantly decreased plasma insulin levels prior to the clamp test, but did not change plasma glucose levels. Furthermore, leptin led to an increase in insulin-stimulated glucose uptake in hindlimb muscles. However, in the case of OLETF rats, leptin administration changed neither plasma insulin levels nor insulin-stimulated glucose uptake. These data demonstrate that OLETF rats at 8 weeks of age have already become resistant to high concentration of peripheral leptin. PMID- 11257314 TI - Hormone-specific combinations of isoforms of adenylyl cyclase and phosphodiesterase in the rat liver. AB - Since many isoforms of adenylyl cyclase and adenosine 3', 5'-monophosphate (cAMP) phosphodiesterase have been cloned, it is likely that receptors of each hormone have a specific combination of these isoforms. Types I, III and VIII adenylyl cyclases are reported to be stimulated by Ca(2+)-calmodulin, type I phosphodiesterase by Ca(2+)-calmodulin, but types IV and VII (cAMP-specific) phosphodiesterases by Co2+. In the present study, we examined different effects of Ca2+ and Co2+ on hormone-induced cAMP response in the isolated perfused rat liver.The removal of Ca2+ from the perfusion medium (0 mM CaCl(2 ) + 0.5 mM EGTA) did not affect glucagon (0.1 nM)-responsive cAMP but reduced secretin (1 nM)-, vasoactive intestinal polypeptide (VIP, 1-10 nM)- and forskolin (1 microM) responsive cAMP considerably. The addition of 1 mM CoCl2 reduced glucagon- and secretin-responsive cAMP considerably, forskolin-responsive cAMP partly, did not affect 1 nM VIP-responsive cAMP, but enhanced 10 nM VIP-responsive cAMP. Forskolin- and VIP-responsive cAMP was greater in the combination (0 mM CaCl(2) + 0.5 mM EGTA + 3 mM CoCl2) than in the Ca(2+)-free perfusion alone. These results suggest that secretin, VIP1 and VIP2 receptors are linked to Ca(2+)-calmodulin sensitive adenylyl cyclase; glucagon receptor to Ca(2+)-calmodulin-insensitive adenylyl cyclase; VIP1 receptor to Ca(2+)-calmodulin-dependent phosphodiesterase; glucagon, secretin and VIP2 receptors to cAMP-specific phosphodiesterase, respectively, in the rat liver. PMID- 11257315 TI - Autoradiographic localization of angiotensin II receptors in developing rat cerebellum and brainstem. AB - The role of Angiotensin II (Ang II) as a growth promoting or modulating factor has recently become a field of intensive research. A central issue in developmental neurobiology is the understanding of mechanisms governing the formation of spatially ordered connections. In this study, we show the localization of Ang II receptor subtypes by autoradiography in 2-week-old rat hindbrains confronting these data with membrane binding assays. Competition studies done on membrane preparations evidence no major changes on the relative affinities for both receptor subtypes between 2-week-old and adult rat tissues. By autoradiography, we found that all the areas (1-10) of the 2-week-old cerebellum showed both receptor subtypes present in complementary adjacent layers. Areas expressing a high level of AT2 receptors follow: inferior colicullus (IC), dorso tegmental nucleus, central (DTgC), subcoeruleus, alpha, sensory root of the trigeminal nerve, principal sensory root trigeminal nucleus (Pr5, Pr5VL) supragenual nucleus, genu facial nerve, facial nucleus, cerebellar peduncles, vestibular and lateral nuclei. Spinal trigeminal, (oral) and Raphe nuclei express AT1 receptor subtype. The high level of Ang II AT2 receptors present in the cerebellar peduncles might have a meaning on the establishment of the olivo-cerebellar connection. The high expression of Ang II AT2 receptors on 2 week-old rat hindbrains, a critical age on development, as well as its disappearance in the adult, strongly suggests a probable role of these receptors in cell migration and neuronal synaptogenesis. PMID- 11257316 TI - Stimulation of rat pancreatic exocrine secretion by cocaine- and amphetamine regulated transcript peptide. AB - Cocaine- and amphetamine-regulated transcript (CART) peptide is a recently described neuropeptide that has been localized to areas of the central and peripheral nervous systems. CART has been shown to be involved in feeding behavior when injected centrally, however, its effects upon peripheral tissues have not been studied. This report describes the effects of CART peptide on rat pancreatic exocrine secretion. Infusion of CART peptide caused four-fold increases in amylase secretion from anesthetized rats that had been fashioned with a bile-pancreatic duct cannula. CART peptide-induced increases in pancreatic secretion appear to involve pathways that are sensitive to both acetylcholine (ACh) and cholecystokinin (CCK) since pre-treatment with atropine (ACh receptor antagonist) or L-364,718 (CCK-A receptor antagonist) inhibited the effects of CART peptide on amylase secretion. Pre-treatment with a combination of atropine and L-364,718 abolished the effects of CART peptide. When isolated rat pancreatic acini were exposed to varying doses of CART peptide, no increase in amylase secretion was observed. The results of the present study suggest that CART peptide has stimulatory effects upon pancreatic exocrine secretion. CART peptide induced increases in pancreatic secretion appear to be indirectly mediated as no direct effect upon pancreatic acini was shown. CART peptide likely acts upon either peripheral or central regulators of pancreatic secretory function that are distant from the acinar unit. PMID- 11257317 TI - Alternative medicine: to teach or not to teach. PMID- 11257318 TI - Pharmacokinetic-pharmacodynamic modeling: why? AB - At present, pharmacokinetic-pharmacodynamic (PK-PD) modeling has emerged as a major tool in clinical pharmacology to optimize drug use by designing rational dosage forms and dosage regimes. Quantitative representation of the dose concentration-response relationship should provide information for prediction of the level of response to a certain level of drug dose. Several mathematical approaches can be used to describe such relationships, depending on the single dose or the steady-state measurements carried out. With concentration and response data on-phase, basic models such as fixed-effect, linear, log-linear, E(MAX), and sigmoid E(MAX) can be sufficient. However, time-variant pharmacodynamic models (effect compartment, acute tolerance, sensitization, and indirect responses) can be required when kinetics and response are out-of-phase. To date, methodologies available for PK-PD analysis barely suppose the use of powerful computing resources. Some of these algorithms are able to generate individual estimates of parameters based on population analysis and Bayesian forecasting. Notwithstanding, attention must be paid to avoid overinterpreted data from mathematical models, so that reliability and clinical significance of estimated parameters will be valuable when underlying physiologic processes (disease, age, gender, etc.) are considered. PMID- 11257319 TI - Inhibition of platelet aggregation by putrescine, spermidine, and spermine in hypercholesterolemic rabbits. AB - BACKGROUND: Hypercholesterolemia causes alterations in platelet function. Platelet hyperaggregation is considered a predisposing factor for atherosclerosis. In this paper, the antiaggregating effect of the polyamines putrescine, spermidine, and spermine was studied on platelets of normal and hypercholesterolemic rabbits. METHODS: New Zealand rabbits were fed with a cholesterol-enriched diet for 10 weeks. Lipids and glucose were determined in serum. The assays of platelet aggregation were carried out using platelet-rich plasma (PRP) obtained from both control and cholesterol-fed rabbits. We used 2.5 micromol /mL ADP and 2 microg/mL collagen as inductors of platelet aggregation. In addition, arginase activity and L-arginine content were determined in PRP. RESULTS: Serum total cholesterol and LDL-cholesterol concentrations were increased from 26.3 +/- 8.1 to 1,485.0 +/- 26.8 mg/dL and from 15.9 +/- 5.9 to 1,383.8 +/- 58.9 mg/dL, respectively, whereas triglyceride concentration increased from 88.3 +/- 35.6 to 411.0 +/- 154.5 mg/dL upon cholesterol feeding. Seventy-five percent of platelet aggregation inhibition was observed with 10 microM of polyamines in PRP of normal rabbits. Spermine inhibited platelet aggregation by 54% in PRP of hypercholesterolemic rabbits when ADP was used as agonist. The order of polyamine action was spermine > spermidine > putrescine. In addition, we found that platelet arginase activity and L-arginine content were unaltered upon hypercholesterolemia. CONCLUSIONS: These results show that the polyamines putrescine, spermidine, and spermine have antagonist action in platelet aggregation and suggest a key role of polyamines in platelet aggregation under normal and hypercholesterolemic conditions. PMID- 11257320 TI - Effects of graft placement site on the survival of adrenal medulla transplants into the brain and its relation with the recovery of motor function. AB - BACKGROUND: Because of their lack of long-term viability, adrenal tissue transplants have shown limited success in alleviating the motor disturbances associated with experimental and pathologic striatal dopamine denervation. In this study, we examined how the graft placement site influences adrenal medulla transplant survival and its relation with the reduction of motor deficits in rats bearing unilateral 6-OHDA lesion. METHODS: One or 5 microL of fetal adrenal medullar tissue was grafted either inside the striatal parenchyma or into the lateral ventricle in contact with the dopamine-denervated striatum. Motor disturbances, as assessed by apomorphine-induced rotation, were correlated to the graft morphologic survival features. RESULTS: Apomorphine-induced rotation showed a marginal reduction of 11% in all groups independently of graft survival features or placement site. Intrastriatal transplants showed limited viability characterized by a substantial loss of graft initial volume as well as fewer and smaller chromaffin cells compared to ventricular grafts, which had a reduced loss of graft initial volume and more and larger chromaffin cells. CONCLUSIONS: Although the lateral ventricle may favor adrenal medulla transplant viability, their induced motor outcome is comparable to that induced by less viable intrastriatal grafts, suggesting that the implanted dopamine-producing cells may interact and influence striatal neurons better when placed in close proximity. PMID- 11257321 TI - Sarcoplasmic reticulum Ca2+ depletion by caffeine and changes of [Ca2+](i) during refilling in bovine airway smooth muscle cells. AB - BACKGROUND: In airway smooth muscle (ASM), Ca2+ influx in response to the Ca2+ depletion of the sarcoplasmic reticulum (SR) seems to play a role in the regulation of intracellular free Ca2+ concentrations ([Ca2+](i)). This study evaluates some possible Ca2+ entry pathways activated during SR-Ca2+ depletion induced by 10 mM caffeine. METHODS: Enzymatically dispersed bovine ASM cells were loaded with Fura-2/AM to permit measurement of [Ca2+](i) changes in single cells. RESULTS: Caffeine (10 mM) induced a transient increase in [[Ca2+](i) that depleted SR-Ca(2)+ content. After caffeine washout, a decrease in basal [Ca2+](i) (undershoot) was invariably observed, followed by a slow recovery. This phenomenon was inhibited by cyclopiazonic acid (5 microM). External Ca(2)+ removal in depolarized and nondepolarized cells induced a decrease in basal [Ca2+](i) that continued until depletion of the SR-Ca2+ content. The decrease in [Ca2+](i) induced by Ca2+-free physiological saline solution (PSS) was accelerated in caffeine-stimulated cells. Recovery from undershoot was not observed in Ca2+-free PSS. Depolarization with KCl and addition of D600 (30 microM) did not modify recovery. Similar results were obtained when the Na(+)/Ca2+ exchanger was blocked by substituting NaCl with KCl in normal PSS (Na(+)-free PSS) or by adding benzamil amiloride (25 microM). CONCLUSIONS: SR Ca2+ content plays an important role in the Ca2+ leak induced by Ca2+-free medium, and does not depend on membrane potential. Additionally, recovery from undershoot after caffeine depends on extracellular Ca2+, and neither voltage dependent Ca2+ channels nor the Na(+)/Ca2+ exchanger are involved. PMID- 11257322 TI - A fortifier comprising protein, vitamins, and calcium-glycerophosphate for preterm human milk. AB - BACKGROUND: The infant's own mother's milk, fortified with proteins, minerals, and vitamins, is considered the best food for low-birth-weight infants. This paper describes the process to obtain a liquid human milk fortifier. METHODS: The fortifier comprises a protein concentrate, calcium, phosphate, and zinc salts, as well as vitamins A and D. A powdered whey protein extracted from bovine milk was concentrated from 31.5-76.8 g/100 g using repetitive dialysis. The protein concentrate was dissolved in a 0.2 M phosphate buffer pH 7.4 and mixed with calcium-glycerophosphate and calcium-gluconate, vitamins A and D, folic acid, and zinc. Each 10 mL of this liquid fortifier has 0.78 g protein, 53 mg calcium, 36 mg phosphate, and 0.93 mg zinc. RESULTS: Repetitive dialysis did not modify the protein structure as demonstrated by electrophoresis. A total of 95% of lactose content was discarded. Enriching human milk using this human milk fortifier increased the concentration per deciliter of all added nutrients; proteins increased from 1.68-2.35 g, calcium from 26-90 mg, and phosphorus, from 15-51 mg. CONCLUSIONS: A liquid human milk fortifier was successfully manufactured using a noncomplex procedure. An intake of 180-200 mL/kg/day of the fortified human milk by the premature infant would satisfy the infant's nutritional requirements and achieve expected growth and development. PMID- 11257323 TI - The short-term effect of a switch from glibenclamide to metformin on blood pressure and microalbuminuria in patients with type 2 diabetes mellitus. AB - BACKGROUND: Renal hyperfiltration and albuminuria have a deleterious effect on kidney function. Therefore, we studied the effect of metformin on blood pressure, renal hemodynamics, and microalbuminuria in type 2 diabetic patients. METHODS: A clinical trial was designed in type 2 diabetic patients with incipient nephropathy. All patients were below the age of 65, normotensive, and without evidence of malignant, hepatic, or cardiovascular disorders. They were randomly allocated to receive glibenclamide or metformin. At baseline and 12 weeks thereafter we measured body mass index (BMI), serum insulin, blood glucose, lipid profile, glycosylated hemoglobin, blood pressure, glomerular filtration rate, renal plasma flow, and urine albumin. RESULTS: We studied 23 patients in the glibenclamide group and 28 in the metformin group. There was no difference in baseline variables between the groups. Metabolic control was obtained in both groups. In the metformin group, all the following variables decreased: microalbuminuria was reduced by a mean of 24.2 mg/day (p = 0.008); systolic and diastolic blood pressure by a mean of 5.3 mmHg (p = 0.002) and 3.93 mmHg (p = 0.009), respectively; insulin levels by an average of 11.8 microIU/mL (p = 0.001), and total cholesterol levels and triglycerides by an average of 0.45 and 0.18 mmol/L, respectively. Insulin resistance measured by the homeostasis model decreased more in the metformin group than in the glibenclamide group. Patients treated with glibenclamide had an increase in HDL cholesterol of 0.082 mmol/L (p = 0.01). CONCLUSIONS: Metformin significantly decreased the urine albumin excretion rate with none of the expected changes in renal hemodynamics, probably due to its favorable effects on blood pressure, lipid profile, metabolic control, and insulin resistance. PMID- 11257324 TI - A rating system for prompt clinical diagnosis of ischemic stroke. AB - BACKGROUND: When a CT scan is not available, an early accurate clinical diagnosis of ischemic stroke is essential to initiate prompt therapy. Our objective was to construct a clinical index that is easy to use when stroke patients are first evaluated at the hospital, to identify those who probably are experiencing an acute ischemic episode. The study was conducted at a university-affiliated medical referral center and two community general hospitals in Mexico. METHODS: Clinical records were reviewed for 801 patients with sudden onset of a focal or global neurologic dysfunction, presumably of vascular origin lasting more than 24 h. Eligibility criteria for this study were admission to the hospital within the first 24 h after symptomatic onset, CT scan diagnosis between 24 and 72 h, and age >45 years. Ischemic stroke included cases of arterial brain infarction, while nonischemic stroke included subarachnoid or intraparenchymatous hemorrhage, mass lesion, venous infarction, and in cases without a CT scan evidence that could explain the clinical manifestations. Data excerpted for analysis were age, sex, history of diabetes mellitus or previous stroke/transient ischemic attack (TIA), time of onset of symptoms, presence of headache, vomiting, neck stiffness, hemiplegia, leukocytosis or atrial fibrillation, diastolic blood pressure, and Glasgow coma scale (GCS) rating. Two multivariable analyses were used: 1) step wise multiple logistic regression (SMLR), and 2) conjunctive consolidation (CC). RESULTS: After appropriate exclusions, the study proceeded with 83 ischemic and 42 nonischemic stroke patients. With SMLR, six variables were selected as predictive for ischemic stroke, including neck stiffness, diastolic blood pressure, previous history of stroke/TIA, hemiplegia, GCS, and atrial fibrillation. An appropriate sum of weighted ratings had a positive predictive value (PPV) of 100% for ischemic stroke. With consolidated categories, the PPV was 97% when patients had the following: no neck stiffness; no atrial fibrillation but history of stroke/TIA and GCS > or =12, or no neck stiffness but atrial fibrillation. CONCLUSIONS: Among patients with acute stroke, clinical data can be used to identify a group with a high probability of ischemic stroke. There are slightly different results between both methods; while SMLR includes the four variables selected by CC, the latter included neither diastolic blood pressure nor hemiplegia/hemiparesia. However, CC results seem easier to understand and interpret than with SMLR. PMID- 11257325 TI - Arylsulfatase A levels in patients with chronic alcoholism. AB - BACKGROUND: The goal of this study was to find the association between low arylsulfatase A (ASA) activity and psychiatric disorders in chronic alcoholic patients. METHODS: The study was carried out in 30 chronic alcoholic patients (27 male, 3 female); age range was 25-65 years. There were 20 normal controls (18 males, 2 females), and age range was 24-67 years. ASA and routine aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activity laboratory tests were measured in blood serum from all patients and control subjects. RESULTS: Alcoholic patients with psychiatric disorders have ASA average values of 68.25 nmol/mL/4 h. This is less than averages found in the alcoholics without psychiatric disorders group (82.48 nmol/mL/4 h) and the control group (90.8 nmol/mL/4 h). There were no statistically significant differences among the three groups studied. Alcoholic subjects with elevated activity of AST and ALT (n = 10) have ASA activity average values of 134.82 nmol/mL/4 h), which is 48.8% higher than the control group (90.6 nmol/mL/4 h). These means show statistically significant differences (p <0.05). CONCLUSIONS: Results indicate an association between low serum ASA activity and alcoholism. The appearance of psychiatric manifestations could be related to the low activity of this enzyme in chronic alcoholic patients. Alcoholic patients with elevated enzyme activity of AST and ALT in sera also have elevated sera arylsulfatase A (ASA) activity. We consider that these findings may be useful for evaluating the psychiatric state as a prognosis in chronic alcoholic patients, and should be a routine laboratory test in alcoholic patients. PMID- 11257326 TI - The seasonal conception of lethal congenital malformations. AB - BACKGROUND: The objective of the study was to investigate the possibility of the effect of seasonal temperature on the incidence of lethal congenital malformations in a retrospective study. METHODS: At the National Institute of Perinatology in Mexico City, perinatal deaths due to congenital malformations were compared with the the remainder of perinatal deaths over a period of 3 years in relation to the average temperature of the months in which the fifth week of gestation occurred. RESULTS: The division between the average temperature of the individual months was 18 degrees C, as there were no months with an average between 17.9 and 18.8 degrees C. An average of 0.86 perinatal deaths with lethal malformations had spent their embryonic life in a cold month, whereas an average of 1.54 had spent their embryonic life in a hot month; in contrast, an average of 10.24 perinatal deaths with lethal malformations had spent their embryonic life in a cold month, and a smaller number (9.23) in a hot month. CONCLUSIONS: The perinatal deaths with lethal malformations therefore showed a significant tendency to have spent their embryonic lives during the hotter months in comparison with the other perinatal fetal deaths (p = 0.04). Further studies should be made with larger numbers of cases, maintaining careful attention to early perinatal data and local temperatures. PMID- 11257327 TI - Clinical outcome of invasive infections in children caused by highly penicillin resistant Streptococcus pneumoniae compared with infections caused by penicillin susceptible strains. AB - BACKGROUND: In this report based on data from the Institutional Surveillance System during 1994-1998, we document the continuing emergence of drug-resistant Streptococcus pneumoniae strains at the Hospital Infantil de Mexico Federico Gomez in Mexico City. METHODS: We evaluate the clinical course of 49 invasive pneumococcal infection outside the central nervous system (CNS) by a number of factors including the site, severity, and place where the infection was acquired, the underlying health of the patient, and the adequacy of antimicrobial therapy. RESULTS: An underlying illness was present in 21 of 49 (43%) patients, 37 (75%) patients had taken previous antimicrobial therapy, and 25% of the infections were nosocomially acquired. Overall, 25 of 49 (51%) of the pneumococcal strains tested were pencillin-resistant; strains with the highest resistance to penicillin were also resistant to cephalosporins. Twenty-two percent of all strains were considered to be multidrug-resistant. Eleven of 25 penicillin-resistant strains were identified as multidrug-resistant, i.e., to erythromycin, TMP/SMX, and chloramphenicol. Ten serotypes accounted for 88% of the isolates, the most frequent serotypes being 23F, 14, 19V, 6A, and 6B. The overall case-fatality rate was 37% (18 of 49), with most deaths occurring within 3-5 days after antibiotic therapy was initiated. There was no difference in the case fatality rate between children with penicillin-nonsusceptible and penicillin-susceptible pneumococcal infections; instead; case-fatality rate correlated with severity of illness on admission and presence of underlying disease. CONCLUSIONS: Characterizing groups at risk for invasive pneumococcal disease could aid in the development of preventive programs and increase the benefits from wide use of future conjugated vaccines. PMID- 11257328 TI - Natural fertility in northeastern Mexico. AB - BACKGROUND: The aims of this study were as follows: 1) to describe the fertility of a sample of Mexican women (> or =45 years of age, married, not using any family planning methods, and residing in the Mexican state of Nuevo Leon); 2) to determine whether or not the distribution of completed family size fits the negative binomial distribution, as in other populations studied in the world, and 3) to assess the association between fertility and 10 explanatory variables. METHODS: A sample of 410 women was interviewed at and selected from seven medical units of the Instituto Mexicano del Seguro Social (IMSS). The women were grouped by their year of birth (1896-1925 and 1926-1955) and birthplace [persons whose four grandparents were born in northeastern Mexico (NE) and outside northeastern Mexico (Not-NE)]. A binomial negative distribution analysis was assessed. Multiple linear regression was used to assess association between fertility (transformed by the use of inverse hyperbolic sines) and 10 explanatory variables, including age at marriage, heterozygosity, individual admixture, wife's education, husband's education, wife's occupation, husband's occupation, and couple's residence zone, birth year, and birthplace. RESULTS: Completed fertility was only associated with age at marriage. This population showed a fertility pattern similar to those described in Venezuelan and Brazilian populations in 1950 and 1940, respectively. CONCLUSIONS: We conclude that before worldwide family planning programs, fertility was determined mainly by natural selection forces. PMID- 11257329 TI - Evagination of Taenia solium cysticerci: a histologic and electron microscopy study. AB - BACKGROUND: After a human being ingests a cysticercus, the larval stage of Taenia solium, the cysticercus gradually develops toward the adult parasite. In this paper, we describe the sequential progress of evagination of cysticerci. METHODS: Intact cysticerci were obtained from swine muscle, and incubated in bovine bile to stimulate evagination. Dissecting, light, and electron microscopy of whole parasites and histologic sections were used for photographic registers. RESULTS: The first event was the widening of the opening of the bladder wall for the scolex and neck to emerge. The two chambers that conform the cysticercus were identified. Histologic sections provided explanation for the conformation of the spiral canal. The scolex uncoils during evagination but does not turn inside out. CONCLUSIONS: The scolex and the neck comprise a different structure from the bladder wall, although they are contiguous. PMID- 11257330 TI - Bisalbuminemia in two Croatian families. AB - BACKGROUND: Bisalbuminemia is a dysproteinemia characterized by the occurrence of two albumin fractions on serum protein separation by electrophoresis on cellulose acetate sheets. Bisalbuminemia may occur as a hereditary trait or as analytical interference with some drugs, especially penicillin. METHODS: Two patients with the finding of bisalbuminemia are presented. Both patients (patient 1 was a 4-1/2 month-old male infant, and patient 2 was a 15-year-old boy) were admitted for respiratory infection. RESULTS: Bisalbuminemia was detected by serum protein electrophoresis and confirmed by isoelectric focusing in pH gradient gel (pH range 4.0-6.5). This finding was supported by simultaneous detection of abnormal albumin in the mother of patient 1, while the father had normal albumin. The abnormal fast albumin in both patients had an increased relative mobility of 1.08 when measured from the sample application position. CONCLUSIONS: The results presented are the first description of albumin mutations in Croatia (that according to the CISMEL group could be classified as ZC/HZ), and present the first step in identification prior to determination of structural change and amino acid sequence in the albumin molecule. PMID- 11257332 TI - Recent progress in vaccines development and new trends in immunisation. PMID- 11257333 TI - Vaccines--a wonderful tool for equity in health. AB - Equity is a word representing one of the most important concerns for planners, politicians and the academia. It comes as an answer to the permanent rediscovery of inequity in the benefits of progress among the peoples of the world. Inequity in the distribution of risks for health and disease is a finding that comes on and on again in real life and in the literature. Inequity in disease burden is clearly related to poverty and the eradication of poverty is the mission for every government and international assistance organisations. Development is also not only a precondition for justice but for peace and stability. Health is seen more and more as preceding development and not only as a consequence of wealth. Therefore many things can and must be done in health to achieve development and of course, equity. This concept has been crucial in the new orientation of WHO and our agency is working hard on its promotion at every level. Vaccines are with us as a simple and effective tool for two centuries. The research and development of new vaccines and all in the underlining knowledge has been much faster than its utilization and there is a huge gap to fill in accessibility and actual coverage. World Health Organization (WHO) has been involved all its life in the promotion of disease control and eradication through the use of immunisation techniques. Presently we are part of GAVI and our Vaccines and Biologicals Department has undergone an important modernisation in order to achieve better performance in pursuing three main objectives: Innovation, Immunisation Systems improvement and Accelerated Disease Control. Therefore in order to walk at a faster pace in the better utilisation of vaccines as a tool for equity, we must join forces and participate actively in all the initiatives that are underway. PMID- 11257334 TI - The future of vaccines, immunisation concepts and practice. AB - Vaccines have prevented more deaths, disability and suffering than any other medical discovery or intervention. Recent breakthroughs in immunology and genomics offer the prospect of the development of many new prophylactic and therapeutic vaccines not only against infectious diseases but also for use in conditions such as allergy, autoimmunity and carcinogenesis where malfunction of the immune system undoubtedly plays a role. These hopeful perspectives are however dimmed by several counterproductive societal trends that include the spreading-although unjustified-belief that vaccines are not safe and may even be unnecessary, escalating costs of vaccine research, development, production and control that are exacerbated by political pressure on selling prices and expensive lawsuits by 'victims' of vaccination who claim excessive compensation. Negative media coverage of vaccine issues is adversely affecting acceptance of vaccination. In spite of these negative trends, vaccines should have a bright future, because it is increasingly being realised that prevention is not only better than cure but it is often also more cost-effective. A better understanding of the dynamics of microbial transmission in populations is leading to more rational immunisation practices on a global scale that could lead to eradication of several pathogens. Attention is being given to making vaccines more user friendly through the development of combined vaccines and the introduction of less invasive inoculation techniques. PMID- 11257335 TI - Current progress in the development of human immunodeficiency virus vaccines: research and clinical trials. AB - In spite of extensive prevention programs, the HIV pandemic is still spreading worldwide, particularly in developing countries. AIDS is the leading cause of death in Africa and the fourth cause worldwide. WHO estimates that there are 16000 new cases of HIV infection daily and that 100 million individuals will be infected during the next decade. In spite of the spectacular results of triple therapy, the best strategy for controlling the HIV epidemics remains the development of an efficacious prophylactic vaccine. However, the development of such a vaccine remains a formidable challenge to both the industry and the scientific community (Esparza J. Bhamarapravati N. Accelerating the development and future availability of HIV-1 vaccines: why, when, where, and how? Lancet 2000; 355: 2061-6 [1]). PMID- 11257336 TI - Influenza vaccines: a review and rationale for use in developed and underdeveloped countries. AB - Multiple studies have demonstrated that influenza infection results in considerable morbidity and mortality, as well as other economic consequences, such as school and work absenteeism. Influenza vaccine has been shown to be both cost-effective and cost-saving. Despite this, the influenza vaccine appears to be under-utilized throughout the world, with significant variations both between countries and within countries over time. Data will be discussed that provide a rationale for the use of the influenza vaccine to protect the public health. Recommendations for the use of the influenza vaccine in various countries and guidelines for influenza vaccine use worldwide will be proposed. PMID- 11257337 TI - Safety, efficacy and effectiveness of the influenza virus vaccine, trivalent, types A and B, live, cold-adapted (CAIV-T) in healthy children and healthy adults. AB - Influenza is a major cause of illness. We have assessed the safety, efficacy, and effectiveness of CAIV-T vaccine. A two year, multicenter, double-blind, placebo controlled, efficacy field trial in pre-school aged children was conducted; 1602 enrolled in Year One and 1358 (85%) returned in Year Two. In both study years combined, the overall vaccine efficacy against culture-confirmed influenza was 92% (95% CI: 88, 94). The vaccine efficacy was 95% (95% CI: 62, 99) against lower respiratory illness, 94% (95% CI: 90, 96) against febrile illness and 96% (95% CI: 88, 99) against otitis media associated with culture-confirmed influenza. A multicenter, double-blind, placebo-controlled, effectiveness field trial was conducted in 4561 working adults aged 18 to 64 years. Episodes and days of febrile illness (FI), severe febrile illness (SFI), febrile upper respiratory illness (FURI), work loss, and health care use were assessed. Vaccination significantly reduced the numbers of SFI, 18.8% reduction (95% CI: 7, 29), and FURI, 26.3% reduction (95% CI: 13, 33); and led to fewer days of illness (22.9% reduction for FI, 27.3% reduction for SFI), fewer days of work lost (17.9% reduction for SFI, 28.4% for FURI), and fewer days of health care provider visits (24.8% for SFI, 40.9% for FURI). Prescription antibiotics and over-the-counter medications were also reduced. The vaccine was generally safe and well tolerated with no vaccine related serious adverse events. LAIV represents an additional important option for the control of influenza. PMID- 11257338 TI - Immunoprophylactic strategies against respiratory influenza virus infection. AB - The objective of this report is to evaluate the prophylactic efficacy of liposome mediated immunotherapy for prevention of respiratory influenza virus infection in mice. Antiviral antibody, interferon-gamma and poly (ICLC) were encapsulated in liposomes and they were evaluated for their ability to induce protective immunity against lethal influenza infection. Passive immunization using liposome encapsulated antiviral antibody was found to offer complete protection against the virus challenge. However, this pretreatment must be administered within 24 h prior to virus challenge to be protective. Pretreatment with liposome encapsulated interferon-gamma was found to stimulate cellular immune responses, but the protection is partial. Immunoprophylaxis using liposome-encapsulated double-stranded (ds) RNA poly (ICLC) provided complete and longer-lasting protection against influenza infection. These results suggest liposome-mediated immunoprophylactic approaches are effective in the prevention of respiratory influenza virus infection. PMID- 11257339 TI - Pneumococcal infection and vaccination in the elderly. AB - The burden of pneumococcal disease among elderly people is not widely known. In most countries, the annual incidence of invasive pneumococcal disease in elderly people is most probably >50 cases/100000 persons. Over the past 20 years, only the United States has used pneumococcal vaccine to any appreciable extent, yet among all vaccine-preventable diseases, pneumococcal infections account for the largest proportion of still unprevented illness in Western Europe. Retrospective studies show that vaccination prevents invasive pneumococcal disease. Pneumococcal vaccination appears to be more cost-effective than any other medical intervention commonly used in the elderly. As new data confirm the benefits of vaccination of the elderly against pneumococcal infections, vaccine use has increased in recent years associated with a decline in invasive disease. PMID- 11257340 TI - New strategies for closing the gap of measles susceptibility in infants: towards vaccines compatible with current vaccination schedules. AB - Peptides representing epitopes of the measles virus glycoproteins have been designed to induce neutralizing and protective antibodies. Those that escape recognition by passively acquired anti-whole virus antibodies could potentially be used as components of a 'pre-vaccine' that could be given during early childhood irrespective of persisting maternal antibodies. Unlike vaccines based on recombinant proteins, epitope-based vaccines can be designed to be compatible with a subsequent boost with the standard life attenuated vaccine. Although synthetic peptides may induce only short-term immunity they have the potential to close in young infants the gap of vulnerability until the standard live attenuated vaccine can be given at 9 or 15 months of age. PMID- 11257341 TI - Monitoring of measles elimination using molecular epidemiology. AB - The different measles virus genotypes are confined to more or less distinct geographic regions. Molecular characterization of virus isolates has been successfully used to determine epidemiological links between cases and the geographic origin of imported viruses. In Europe, indigenous measles has been eliminated in some countries, but in others the disease is still endemic. Intra outbreak variability can be used to differentiate between sporadic endemic cases and a 'pseudo-outbreak' of unrelated imported cases. The interruption of virus circulation by mass vaccination campaigns could be demonstrated by comparing the variability of pre-campaign viruses with post-campaign isolates. Simplified tools are being developed that could bring genotyping within reach of laboratories that do not have the possibility of sequencing. PMID- 11257342 TI - Studying experimental measles virus vaccines in the presence of maternal antibodies in the cotton rat model (Sigmodon hispidus). AB - The inhibition of vaccine-induced seroconversion after vaccination is one of the problems associated with measles virus (MV) immunization. In cotton rats, after transfer of human MV specific antibodies, vaccine-induced seroconversion is inhibited. With this model, it was shown that plasmid immunization (although successful in seronegative animals) was inhibited by maternal antibodies. In contrast, immunization via a mucosal surface with a vesicular stomatitis virus expressing the MV hemagglutinin induced seroconversion in the presence of maternal antibodies and subsequent protection. PMID- 11257343 TI - Prevention of measles in Sudan: a prospective study on vaccination, diagnosis and epidemiology. AB - Despite the availability of safe and effective live attenuated vaccines, measles continues to be endemic in many developing countries. Control and elimination of measles will be especially difficult in East Africa, because of its limited infrastructure and political instability. We have studied diagnostic and epidemiological aspects of measles in suburban Khartoum, Sudan. Prospective studies were carried out in a cohort of clinically diagnosed measles cases and in a cohort of newborns, which were both followed up for 2 years. The studies intended to provide a rational basis for improvement of measles vaccination strategies, and strengthen measles research infrastructure in Khartoum. PMID- 11257344 TI - Measles elimination: old and new challenges? AB - Life attenuated measles vaccines have dramatically reduced measles morbidity and mortality world-wide. Despite high vaccination coverage, measles outbreaks continue to occur both in developed and developing countries. While secondary vaccine failure may be responsible for disease in some seroconverted individuals, evidence suggests that many more vaccinees who are protected against disease may not be fully protected against virus infection. In low-income developing countries protection by maternal antibodies seems to erode faster than previously estimated especially in infants who were born to vaccinated mothers. Problems of infectivity and susceptibility of vaccinees will be compounded in case wild-type viruses become less sensitive to vaccine induced immunity. These observations suggest that elimination may be more easily achieved as long as large proportions of populations are protected by wild-type virus-induced immunity. PMID- 11257345 TI - Oral poliomyelitis vaccine: time to change? AB - For 3 decades, vaccination against poliomyelitis has rested mainly on the use of the oral attenuated vaccine (OPV). In countries where wild type poliomyelitis has been successfully controlled by OPV, the rare cases of poliomyelitis that can still be identified occur in vaccinees or their contacts and are caused by vaccine related strains. Over years, data indicating that the inactivated vaccine (IPV) also has the potential to control poliomyelitis and that there are no known risks associated with the use of this vaccine have accumulated. The reasons for changes in vaccine policy in industrialised countries and the situation of the global effort of poliomyelitis immunisation are described. Some of the issues and challenges for the future are reviewed. PMID- 11257346 TI - Inactivated poliovirus vaccine and the final stages of poliovirus eradication. AB - The use of the inactivated poliovirus vaccine (IPV) will increase before and probably also after the global eradication of the wild type poliovirus. Before eradication, the switch from the use of oral poliovirus vaccine (OPV) to IPV has been due to the better safety record of IPV. Introduction of IPV in the regular immunisation schedules is made easier by the development of several combination vaccines, including IPV. Maternal antibodies and young age, often considered problematic for early initiation of IPV schedules, did not compromise optimal maintenance of seropositivity during infancy or long-term persisting antibody levels in our studies. OPV-derived, potentially pathogenic and transmissible poliovirus strains, excreted by some individuals for years, may present a problem for a blunt stopping of all polio immunisations after eradication. Our recent results suggest that locally excreted IgA might have a role in the elimination of poliovirus infection in the intestinal tissues. PMID- 11257347 TI - The incurable wound revisited: progress in human rabies prevention? AB - Rabies is the most important viral zoonosis from a global perspective. Modern human postexposure prophylaxis consists of potent vaccines and local infiltration of rabies immune globulins (RIGs), but the latter biologicals are not widely available or affordable. Monoclonal antibodies (Mabs) offer several theoretical advantages over RIGs. To this end, several human and equine RIGS, alone or in combination with vaccine, were investigated for postexposure efficacy in a Syrian hamster model, compared with a single neutralizing murine Mab. Preliminary results suggest that: (1) animal models continue to provide utility as human surrogates in the demonstration of product efficacy against rabies; (2) RIG preparations differ substantially in experimental effectiveness and clearance; and (3) relevant alternatives, such as Mabs, should be pursued for future improvements to human rabies prevention. PMID- 11257348 TI - Clinical relevance of lower Hib response in DTPa-based combination vaccines. AB - Combination vaccines are essential to enable administration of all the required antigens in routine infant immunisation schedules at any single visit. Some combinations of diphtheria-tetanus-acellular pertussis (DTPa) with Haemophilus influenzae type b (Hib) conjugate vaccines have been shown to result in lower Hib titres than when Hib is administered separately. While confirming that a primary series with a DTPa-HBV-IPV/Hib combination gives lower antibody levels than separate Hib conjugates, we show that the nature (isotype and IgG subclasses) and function (avidity and opsonic activity) of the antibodies are the same, and immunologic memory is induced. It is likely therefore that the DTPa-HBV-IPV/Hib combination will be efficacious against Hib disease. PMID- 11257349 TI - Progress in vaccination against Helicobacter pylori. AB - This review discusses recent progress in the development of a vaccine against Helicobacter pylori. This progress includes demonstration that: effective immunisation is independent of antibodies but dependent upon CD4+ T helper cells, although their role remains unknown; the immunisation regime can be improved to increase efficacy; successful immunisation against H. pylori is possible using a live vector; a strain of H. pylori suitable for experimental infection of humans has been developed. Important issues that remain to be addressed include incomplete protection, non-availability of suitable mucosal adjuvants and post immunisation gastritis. Significantly, commercial development of products for clinical trial is underway. PMID- 11257350 TI - Safety and immunogenicity of an intranasal Pseudomonas aeruginosa hybrid outer membrane protein F-I vaccine in human volunteers. AB - A hybrid protein [Met-Ala-(His)(6) OprF(190-342)-OprI(21-83)] consisting of the mature outer membrane protein I (OprI) and amino acids 190-342 of OprF of Pseudomonas aeruginosa was expressed in Escherichia coli and purified by Ni(2+) chelate-affinity chromatography. After several studies in healthy volunteers, as well as in patients, had proven the tolerability and immunogenicity of the the OprF-OprI vaccine, after intra-muscular application, we developed an emulgel for intranasal immunization. For this purpose we combined a highly concentrated OprF I with sodium dodecylsulfate as vehicle and the gel matrix natriumlauryl sulfate. After safety and pyrogenicity evaluations in animals, eight healthy adult human volunteers received the OprF-I gel intranasally three times at 2-week intervals. The vaccination was well tolerated and no side effects were observed. An antibody induction (IgG and IgA) could be detected in the sera. These data support continued clinical investigation of the protection against infections in cystic fibrosis patients and patients prone to P. aeruginosa infections. PMID- 11257351 TI - Tuberculosis subunit vaccine development: on the role of interferon-gamma. AB - Tuberculosis (TB) remains a major global health problem and subunit vaccines for the control of the disease are presently under development. This vaccine strategy requires an in vitro correlate of protection for the identification of relevant vaccine candidate antigens and for monitoring the induction of a protective cell mediated immune response after vaccination. New studies of experimental vaccines in the mouse model of TB support interferon-gamma as a relevant marker for the induction of a protective immune response. In contrast, searching for immunodominant antigens capable of inducing strong interferon-gamma responses in PPD positive healthy or TB infected individuals may not identify all relevant candidate antigens for inclusion in a novel TB subunit vaccine. PMID- 11257352 TI - The prevention of Lyme disease with vaccine. AB - Lyme disease is a potentially serious and debilitating infection caused by Borrelia burgdorferi that is endemic in North America, Europe, and Asia. Personal protective and environmental measures have not significantly impacted its increasing incidence. An adjuvanted recombinant vaccine (LYMErix) has been approved in the United States for the prevention of Lyme disease in adults, and has demonstrated both safety and efficacy. A clinical trial of over 10000 adults showed 76% efficacy following the third dose of a 0, 1, 12 schedule. Accelerated schedules demonstrate equivalent levels of protective antibody. Up to 100% of children 2-14 years of age achieve seroprotective levels of antibody. Booster doses induced protective levels of antibody in more than 96% of recipients when administered at months 12 and 24. Only mild or moderate, transient vaccine associated adverse events have been reported after immunization. The vaccine is a safe and effective method of preventing Lyme disease. PMID- 11257353 TI - Transmission blocking malaria vaccines. AB - Transmission blocking vaccines (TBVs) against malaria are intended to induce immunity against the stages of the parasites which infect mosquitoes so that TBV immunised individuals cannot transmit malaria. As malarial infections are transmitted mainly within a few hundreds of meters from an infectious human source, TBVs used within in a community would protect the immediate neighbourhood of the vaccinated individuals. TBVs against the two major species of human malaria, Plasmodium falciparum and P. vivax, are under development. Candidate TBV constructs for both Plasmodium species have been successfully tested in animal systems and testing is in progress with clinical grade material in humans. PMID- 11257354 TI - Genomic tools for gene and protein discovery in malaria: toward new vaccines. AB - Advances in malaria vaccine and drug development have been hindered in part by the complex multistage life cycle of the parasite, much of which is inaccessible to study, and by a large genome encoding over 5000 genes. Two human models of immunity to malaria, however, suggest that the development of an effective vaccine is within reach. We have outlined a strategy to identify the expression of hundreds to thousands of potential vaccine targets employing recently developed technologies for gene and protein expression. Combined with the exciting developments of malaria DNA vaccine technologies, these approaches form the basis for malaria subunit vaccines that may mimic the protective efficacy of our human model systems and provide the foundation for novel approaches to vaccine development for a range of pathogens. PMID- 11257355 TI - Conjugates and reverse vaccinology to eliminate bacterial meningitis. AB - Bacterial meningitis is caused mainly by Haemophilus influenzae, Streptococcus pneumoniae and Neisseria meningitidis. Haemophilus influenzae type b vaccines have been extremely successful in eradicating the disease from those countries where the vaccine has been introduced. The recent licensure of the conjugated pneumococcal vaccine suggests that this pathogen also will be soon controlled. Consequently, if we succeed in developing effective vaccines against meningococcus, this will enable us to eliminate bacterial meningitis. The global elimination of bacterial meningitis is a goal which, if appropriate resources are applied, can be reached within the first fifteen years of the 21st century. PMID- 11257356 TI - Peptide and recombinant antigens for protection against bacterial middle ear infection. AB - Passive immunization of chinchillas with serum specific for either LB1 or for LPD LB1 (f)(2,1,3) prior to challenge with heterologous NTHI isolates (relative to diversity in region three of P5-fimbrin), significantly inhibited the signs and incidence of otitis media (P < or = 0.01) induced by any of the challenge isolates. The ability of these antisera to induce total eradication of NTHI from the nasopharynx was not however equivalent among challenged cohorts. The data thus suggested that while early, complete eradication of NTHI from the nasopharynx was highly protective, reduction of the bacterial load to below a critical threshold level appeared to be similarly effective. Both immunogens thus remain strong vaccine candidates. PMID- 11257357 TI - Recombinant measles viruses expressing heterologous antigens of mumps and simian immunodeficiency viruses. AB - We have genetically engineered a panel of recombinant measles viruses (rMVs) that express from various positions within the MV genome either the HN or F surface glycoproteins of mumps virus (MuV) or the env, gag or pol proteins from simian immunodeficiency virus (SIV). All rMVs were rescued from the respective antigenomic plasmid constructs; progeny viruses replicated comparably to the progenitor Edmonston B MV, but showed slight propagation retardation, which was dependent on the size and nature of the expressed proteins and on the genomic position of the inserts. All transgenes except that encoding mumps F glycoprotein were faithfully maintained and expressed even after virus amplification by 10(20). Our results suggest possible applications of rMVs as live-attenuated, multivalent vaccines against retroviruses such as SIV and HIV as well as other pathogens more distantly related to MV than MuV. PMID- 11257358 TI - An efficacious recombinant subunit vaccine against the salmonid rickettsial pathogen Piscirickettsia salmonis. AB - Piscirickettsia salmonis is the aetiological agent of salmonid rickettsial septicaemia, an economically devastating rickettsial disease of farmed salmonids. Infected salmonids respond poorly to antibiotic treatment and no effective vaccine is available for the control of P. salmonis. Bacterin preparations of P. salmonis were found to elicit a dose-dependent response in coho salmon (Oncorhynchus kisutch), which varied from inadequate protection to exacerbation of the disease. However, an outer surface lipoprotein of P. salmonis, OspA, recombinantly produced in Escherichia coli elicited a high level of protection in vaccinated coho salmon with a relative percent survival as high as 59% for this single antigen. In an effort to further improve the efficacy of the OspA recombinant vaccine, T cell epitopes (TCE's) from tetanus toxin and measles virus fusion protein, that are universally immunogenic in mammalian immune systems, were incorporated tandemly into an OspA fusion protein. Addition of these TCE's dramatically enhanced the efficacy of the OspA vaccine, reflected by a three-fold increase in vaccine efficacy. These results represent a highly effective monovalent recombinant subunit vaccine for a rickettsia-like pathogen, P. salmonis, and for the first time demonstrate the immunostimulatory effect of mammalian TCE's in the salmonid immune model. These results may also be particularly pertinent to salmonid aquaculture in which the various subspecies are outbred and of heterologous haplotypes. PMID- 11257359 TI - Identification and characterisation of multiple linear B cell protectopes in the respiratory syncytial virus G protein. AB - Respiratory syncytial virus (RSV) is an important respiratory pathogen in man, against which no vaccine is available. However, recent evidence suggests that antibodies to the RSV F and G proteins may play an important role in disease prevention. We previously demonstrated that BBG2Na, a subunit vaccine candidate including residues 130-230 of the Long strain G protein, protects rodents against RSV challenge. Using a panel of monoclonal antibodies (MAb) and synthetic peptides, five linear B cell epitopes were identified that mapped to residues 152 163, 165-172, 171-187 (two over-lapping epitopes) and 196-204. Antibody passive transfer and peptide immunisation studies revealed that all were protective. Pepscan analyses of anti-RSV-A and BBG2Na murine polyclonal sera suggested stronger immunogenicity of some protective epitopes (protectopes) in the context of BBG2Na compared with live virus. However, all the identified murine B cell protectopes were conserved in RSV seropositive humans. Should these protectopes correspond with protection in humans, BBG2Na may constitute a very interesting vaccine candidate against RSV. PMID- 11257360 TI - Characterisation of a protective linear B cell epitope against feline parvoviruses. AB - Monoclonal antibody 3C9 was the starting material in the definition of the epitope that led to the synthesis of the first efficient peptide vaccine against a viral disease (canine parvovirus) in the natural host (dog). In this report, we have analysed the specificity of the antibody at the single amino acid level and the contribution of each residue to the binding, using multiple length analysis. Moreover, a replacement analysis allowed determining those critical residues for the binding. Finally, in an attempt to optimise the production cost of the vaccine, we have determined that the minimal dose required for induction of protective antibodies can be as low as 0.5 microg of peptide. Also, KLH can be replaced as a carrier for a much cheaper alternative such as ovalbumine. All these findings implicate a substantial reduction in the cost of the vaccinal dose. PMID- 11257361 TI - Exploiting molecular mimicry to broaden the immune response to carbohydrate antigens for vaccine development. AB - Peptide mimetics of carbohydrates represent an alternative approach to induce anti-carbohydrate responses. Depending on their formulation, peptide mimetics can mediate T-independent or T-dependent responses. Multivalent peptide mimeotopes can induce high IgM/IgG ratios, as non-conjugated carbohydrates do. Here we observe that immunization with multivalent peptide mimeotope conjugated to BSA enhances carbohydrate reactive antibodies in Balb/c mice and xid mice, with IgG1 greater than IgG2a, in xid mice. These results suggest that mimeotope-conjugate formulations might augment carbohydrate-specific immune responses in immuno compromised hosts. PMID- 11257362 TI - Pitfalls of reductionism in the design of peptide-based vaccines. AB - It is widely believed that all biological phenomena can be reduced to chemistry and physics. Such a reductionist view disregards the fact that complex biological systems have relational (also called emergent) properties that their constituents lack and that cannot be deduced or predicted from the properties of the isolated components. When the individual components of the immune system are studied in isolation, many interconnections are lost and it is not possible to understand how the system functions at the level of the organism as a whole. Our increasing knowledge of the antigenic structure of viral proteins has also been of little help for improving the immunogenicity of individual viral epitopes and for enhancing their capacity to elicit a protective immune response against viral infection. When molecular design principles are used to optimize the binding properties of a synthetic peptide epitope with respect to one neutralizing monoclonal antibody, this does not ensure that the peptide, when used as immunogen, will be able to induce neutralizing antibodies that protect against disease. A reductionist approach does not provide the information required for designing peptide immunogens that will elicit neutralizing rather than non neutralizing antibody responses. PMID- 11257363 TI - Hepatitis B: vaccination programmes in Europe--an update. AB - In the eight years since the Global Advisory Group of the Expanded Program on Immunisation set 1997 as the target for integrating hepatitis B (HB) vaccination into national immunisation programs world-wide, more than 116 countries have included HB vaccine as part of their routine infant or adolescent immunisation programs. Meanwhile, many countries have performed economic evaluation studies, while others have initiated sero-epidemiological studies to generate input data for burden of disease calculation. These studies have indicated that epidemiological and economic arguments cannot be used to delay the implementation of universal hepatitis B vaccination. Some countries have improved their surveillance system and included viral hepatitis in the surveillance programs. Other have put hepatitis B vaccination on the political agenda. By the year 2000, following countries of the WHO European Region (51 countries) have implemented a universal hepatitis B immunisation programme: Andorra, Albania, Austria, Belarus, Belgium, Bulgaria, Estonia, France, Germany, Greece, Italy, Israel, Kazakhstan, Kyrgyzstan, Latvia, Lithuania, Luxembourg, Malta, Moldova, Monaco, Poland, Portugal, parts of the Russian Federation, Romania, Slovakia, Slovenia, San Marino, Spain, Switzerland, Turkey and Uzbekistan. The Netherlands and some other European countries are seriously studying the issues or are making budgetary provisions for introduction of HB vaccine into their vaccination programme. Most of the European countries, which now use the vaccine routinely, have started with adolescent or infant immunisation. Belgium (1999), France (1994) and Italy (1991) have begun with both adolescent and infant HB immunisation. France continues since 1st October 1998 with the infant immunisation programme only. The rewards of effective implementation of the programmes in these countries are becoming apparent; and their success offers an exemplary model for other countries. The deadline was 1997. Globally, work still remains to be done to support and implement interventions that will bring us closer to the WHO goal and to control, eliminate and eradicate hepatitis B in the coming generations at large. If all the 145 million infants born in 1991 had been vaccinated in this way, the number of chronic carriers would have been reduced by 7.5 million, and 1.8 million deaths prevented. PMID- 11257364 TI - Update on hepatitis B vaccination in Italy 10 years after its implementation. AB - Since the beginning of the Italian program of immunization against hepatitis B, vaccine has been given to more than 9 million children, with an outstanding record of safety and efficacy. The coverage rate is globally around 94%, with differences between Northern and Southern regions, the latter having the lower acceptance rate. According to the National Surveillance System (SEIEVA), the incidence of acute hepatitis B per10(5) inhabitants declined from 5.4 in 1990 to 2.9 in 1998. The reduction was even greater among 15-24-year-old individuals, where the incidence rate per 10(5) decreased from 17.3 to 4.2 in the same period. In parallel with the decline of hepatitis B, Delta hepatitis has also dropped significantly. We expect that by the year 2003 (12 years after the beginning of the program) this vaccination strategy will have led to the protection of all Italians aged 0-24 years, who are those at the higher risk for acquiring hepatitis B virus (HBV) and for developing the chronic carrier state. PMID- 11257365 TI - Vaccination against hepatitis viruses in Poland. AB - At present, two etiologic varieties of viral hepatitis (VH) can be directly vaccine-preventable: VH type A and VH type B. In addition, VH type D can be prevented indirectly through vaccination against VH-B. The first commercially available vaccine against VH-B appeared in 1981 and was human plasma-derived. After several years, it has generally been replaced by a recombinant type of vaccines. The obvious benefits of vaccination against VH-B prompted its introduction into the national immunization program in Poland in 1989. At that time, vaccination was offered free of charge to high-risk groups: newborns of HBsAg-carrier mothers, health-care workers, students: at medical schools, nursing schools, medical technology schools, and caretakers at institutions for mentally retarded persons. However, similarly to the experiences of other countries, observations in Poland indicated that such a targeted strategy fails to induce major epidemiological changes. In 1989 and in 1993, the incidence of VH-B per 100000 was 40.3 and 34.6, respectively. In addition, during these years, the incidence of V-B per 100000 children aged 0-4 years was 20.0 and 38.4, respectively. It has been decided that vaccination against VH-B will be obligatory for all newborns beginning from 1993. Due to financial constraints, it has been introduced in three phases, and since 1996, all newborns in Poland have been vaccinated. Already in 1993, three additional risk groups have been offered vaccination: patients with chronic diseases, patients awaiting planned surgery, and persons coming into close contact with acute VH-B or chronically HBV-infected individuals. In 1999, the incidence of VH-B per 100000 was 9.1/100000, and it may be assumed that vaccination helped to decrease the incidence of VH-B in Poland. The country experience with vaccination against VH-A is still limited. At present, it is recommended for children and adolescents and people dealing with food distribution, as well as for several other groups of people, such as travellers or long-term visitors (soldiers, missionaries, diplomats) to the endemic regions of the world. It has also been recommended in connection with natural disasters such as floods occurring in a large area of Poland in 1997. PMID- 11257366 TI - Vaccination against hepatitis B in health care workers. AB - Hepatitis B is the most important infectious occupational disease for health care workers. The high risk of being infected is the consequence of the prevalence of virus carriers in the assisted population, the high frequency of exposure to blood and other body fluids and the high contagiousness of hepatitis B virus (HBV). Vaccination is able to prevent the most threatening consequences of the infection (acute disease and chronic carriage) in responders, even after loss of detectable antibodies. Non-responders to the primary series may benefit from administration of up to three more doses of vaccine (40-70% of initial non responders show seroconversion to the new series). However, newly developed vaccines that seem more immunogenic are presently under evaluation and should further decrease the number of non-immune workers in the near future. In the mean time, coverage with standard vaccines should be improved also by supplying complete information on the risks of hepatitis B and on the safety and efficacy of active immunisation. PMID- 11257367 TI - Immunotherapy of chronic hepatitis B by anti HBV vaccine: from present to future. AB - Chronic liver disease and hepatocellular carcinoma associated with chronic hepatitis B virus (HBV)-infection are among the most serious human health problems in highly endemic regions. Current therapeutic approaches to control chronic hepatitis such as interferon-alpha and lamivudine are unsatisfactory. Vaccination would be the therapeutic procedure with the lowest cost and the potentially greatest benefit. The immunogenicity of selected HBV envelope- or capsid-based vaccine formulations for the induction or the broadening of T and B cell responses, deficient in HBV chronic carriers, are currently under study in animal models and in clinical trials. PMID- 11257368 TI - A prophylactic hepatitis B vaccine with a novel adjuvant system. AB - Studies with recombinant hepatitis B vaccines show seroprotection rates varying between 91 and 100%. Thus, a limited risk may remain for non-responding populations (e.g. non-responders, haemodialysis patients, elderly) who could benefit from a more immunogenic hepatitis B vaccine. One strategy to enhance the immune response is the use of novel adjuvants. SmithKline Beecham has developed a new adjuvant system containing alum and 3-deacylated monophosphoryl lipid A: SBAS4 (SmithKline Beecham Adjuvant System 4). Pilot studies showed that SBAS4 improved in vivo humoral and in vitro cellular immune responses compared to the response to classical recombinant hepatitis B vaccines and was safe and well tolerated. Several studies assessed the profile of the HBsAg/SBAS4 vaccine in a healthy population, non-responders or elderly. In general the HBsAg/SBAS4 vaccine was well tolerated. Compared to an established recombinant hepatitis B vaccine, we observed an increased local reactogenicity but few symptoms were reported as severe. The HBsAg/SBAS4 vaccine elicits a strong immune response: subjects are protected faster and the GMTs are usually much higher. HBsAg/SBAS4 thus has the potential to protect those subjects who fail to be protected by well established hepatitis B vaccines. PMID- 11257369 TI - Vaccination policy against hepatitis A in Italy. AB - In Italy, improved sanitation and living conditions have led to a decline in the rate of hepatitis A infection among children generating an increasing proportion of adults susceptible to this virus. Shellfish consumption is a major source of infection while person to person transmission is important in the spread of infection and in the maintenance of outbreaks. Thus prevention of secondary HAV infection is a crucial point. A randomised controlled trial of hepatitis A vaccine in household contacts of people with sporadic HAV infection in Italy has shown a protective efficacy of 82% (CI 20-96%). The two secondary infections in the vaccine group were symptomless, suggesting that the disease expression may be weaker in vaccinated subjects. PMID- 11257370 TI - Combining hepatitis A and B vaccination in children and adolescents. AB - Hepatitis A and B are common infections worldwide and their severity is related to the individual's age upon initial infection. Furthermore, when hepatitis B infection occurs in infants, the risk of becoming a chronic carrier is 90%. For hepatitis A, the lower incidence of disease arising from an improvement in living conditions leaves a greater number of children, adolescents and young adults susceptible to residual circulating virus. Consequently, initial infection occurs later in life when clinical illness is more frequent and the rate of morbidity and mortality higher. Although both viruses differ greatly, including their modes of transmission, the overlap in their epidemiology warrants the combination of hepatitis A and B vaccination. The immune response elicited by the combined hepatitis A and B vaccine following a three-dose schedule compares well with the anti-hepatitis A virus (HAV) and anti-hepatitis B sero-responses obtained with monovalent vaccines. In addition, it was shown that the seroprotection rate for anti-hepatitis B increased more rapidly with the administration of the combined vaccine, with values of more than 80% within 1 month after the first two doses (schedule, 0, 1 and 6 months). Currently, according to the World Health Organization recommendations, more than 116 countries are vaccinating their infants and/or adolescents against hepatitis B. Recently, several countries were considering or have decided to begin mass vaccination against HAV (more than fifteen states in the US, Spain (Catalonia), Italy (Puglia)). For these countries, the combination of hepatitis A and B antigens in one single vaccine offers the following advantages: fewer injections for protection against two infections, better compliance, lower implementation costs, and fewer missed vaccination opportunities. Further simplification of the schedule, by reducing the number of doses, would improve the compliance rate as well as being more convenient for the vaccinee. This should translate into a reduction in costs associated with vaccine administration. In some recent vaccine studies, the immunogenicity and safety profile of a two-dose schedule (0 and 6 months) of the adult formulation of the combined hepatitis A and B vaccine was investigated in children aged 1-11 years, as well as in adolescents aged 12-15 years. Current results indicate that this two-dose schedule of the adult formulation could be considered a viable alternative for immunization of children and adolescents. PMID- 11257371 TI - Australian experience in general medical practice immunization--achievements and problems. AB - There are three essential components in the vaccination equation model: the vaccine giver, the vaccine receiver and the process facilitator. All three components must be considered if an immunization programme is to achieve its maximum potential. The author uses this model to explain the rationale behind strategies used in the Immunize Australia Campaign which began in 1997. The article discusses options used to overcome obstacles from the perspective of a general medical practitioner. PMID- 11257373 TI - Adverse events and vaccination-the lack of power and predictability of infrequent events in pre-licensure study. AB - The recent setback in the development of a safe and effective rotavirus vaccine illustrates an important problem regarding prelicensure testing and its ability to identify rare vaccine-related adverse effects. It is our contention that the possibility of a rare but serious vaccine adverse effect is difficult to detect in prelicensure testing. In this paper, we review the history regarding the testing and eventual studies that led to the permanent withdrawal of that vaccine. The post-licensure discovery of a serious adverse event associated with the rotavirus vaccine is not unique among vaccines, but represents a recurrent phenomenon that in fact is mathematically predictable. Prelicensure studies examine thousands of subjects and not hundreds of thousands. A sample size of 10,000 subjects may provide excellent estimates of efficacy, but cannot provide an adequate denominator to rule out rare adverse events. It lacks the power. Just as with the rotavirus vaccine, only after hundreds of thousands of doses of vaccines are distributed, will such rare events appear often enough to permit detection. For that reason, we must depend upon the modern post-licensure surveillance programs that we already have in place. PMID- 11257372 TI - Making vaccines more acceptable--methods to prevent and minimize pain and other common adverse events associated with vaccines. AB - The growing abundance of highly immunogenic vaccines has arrived with a burden of pain, distress, and common adverse reactions that in turn may interfere with parental compliance and aggravate anti-vaccine sentiment. In a study of 150 children in each of 2 age-groups, we found that approximately 20% of the subjects suffered serious distress or worse. During the procedural phase, approximately 90% of the 15-to-18 month old children and 45% of the 4-to-6 year old children showed serious distress or worse. To address non-adherence with pediatric vaccine schedules, we must consider all of the possible issues that might prevent a parent from taking a child to a health care provider for vaccination. In that same study we identified useful predictors for both preparatory and procedural distress - predictors that might be used in identifying children who might benefit from preventive interventions. Vaccine providers might consider a variety of interventions. Attitude, empathy, instruction, and practice have all been shown to have a salutatory effect upon pain and anxiety with medical procedures in general and specifically with vaccinations. Distraction has also been found to be an effective method for distress and pain prevention in children. More formal methods of clinical hypnosis which combine a deep state of relaxation with focused imagery and suggestion have also been found to be effective in helping children and adolescents prepare for, cope with, and tolerate the pain and anxiety associated with medical procedures. So-called 'sugar nipples' delivering small amounts of sucrose orally at the time of a painful procedure in an infant has been not been shown to decrease vaccination pain and studies on refrigerant topical anesthetics are mixed. Studies have found a eutectic mixture of 2.5% lidocaine and 2.5% prilocaine (EMLA) effective in providing adequate local anesthesia in children, but it suffers from problems in practical application. Studies with various injection techniques have not identified ready solutions, and although jet injection appears to provoke less anxiety and cause less immediate pain, studies also indicate a somewhat greater incidence of delayed local reactogenicity including soreness and edema. Other measures to prevent or rapidly treat other common adverse events have been shown effective and should be considered as well. PMID- 11257374 TI - Twin studies of immunogenicity--determining the genetic contribution to vaccine failure. AB - CONTEXT: Estimating the magnitude of the genetic contribution to the overall variation of antibody levels among individuals should help clarify the role of genetic association in the biological mechanism of vaccine response and failure. This, in turn, should help guide the design of improved vaccines with enhanced efficacy. OBJECTIVE: To explore the magnitude of genetic influence on antibody levels following measles, mumps and rubella vaccines. DESIGN: Cross-sectional survey study. SETTING: Olmsted County, Minnesota. PARTICIPANTS: Healthy twin pairs. Of the 100 twin-pairs enrolled, 45 were monozygotic. INTERVENTIONS: Determinations of zygosity, vaccine status, and quantitative IgG to measles, mumps, and rubella. MAIN OUTCOME MEASURE: Heritability (ratio of genetic variance to total variance). RESULTS: The number of vaccine-doses, the age at initial immunization, and the time between immunization and sampling did not differ between monozygotic and dizygotic twin pairs. The genetic variance - the variance in antibody levels presumably due to genetic effects - was 0.49 for measles, 0.54 for mumps, and 0.13 for rubella. Heritability, the ratio of genetic variance to total variance, was 88.5% for measles, with the lower bound of a one-sided 95% confidence interval equal to 52.4%. The heritability was, for mumps, 38.8% with a lower bound of 1.60%. The heritability for rubella was 45.7% with a lower bound of 4.94%. CONCLUSION: Our data support the concept that genetic influences play a substantial role in the variation of antibody levels following immunization against measles and, to a lesser extent, mumps and rubella. PMID- 11257375 TI - Understanding those who do not understand: a brief review of the anti-vaccine movement. AB - Vaccines and the ability to prevent morbidity and mortality due to infectious diseases have been one of the greatest public health success stories. On a global level, it is one of the few cost-effective medical measures that result in universal benefit. Despite this, there is evidence of a growing anti-vaccine movement. In turn, this has, in some cases, resulted in major disruptions in vaccine programs, with resultant needless morbidity and mortality. Of interest are the factors that seem to contribute to the current trend of anti-vaccine sentiment. This paper will examine the current anti-vaccine movement and provide current examples. Finally, a review of suggestions for dealing with the anti vaccine movement will be presented. PMID- 11257376 TI - Vaccine safety: risk communication--a global perspective. AB - The complexity of risks connected with both vaccine-preventable diseases as well as with immunization will be discussed and used as the basis for conclusions: what should be done to avoid damage to current and future immunization programs and -how to improve risk communication. The need for more complete data on true, perceived and unknown immunization risks necessitates strengthening our research capabilities as well as surveillance and vaccine safety programs, and to critically examine the factors influencing public sentiments, taking into account that public perceptions of risk vary depending on the characteristics of risk. A concerted effort is needed to improve benefit and risk communication at all levels. The medical community should play a key role for improved communication. PMID- 11257377 TI - Vaccine safety--improving monitoring. AB - The growing numbers of vaccines and vaccine combinations make monitoring of vaccine safety increasingly complex. At the same time, health agencies and the public request better information on the safety of vaccines. The following steps should be taken to improve monitoring of vaccine safety: bar code labelling of vaccines to improve accuracy and completeness of information on administered vaccines, establishing immunisation registers to provide numerator data on exposure to vaccines, to facilitate long term follow-up and to do linked database studies. Development of improved case definitions to increase comparability between studies. Organising systematic assessments of all information on suspect or alleged vaccine reactions. PMID- 11257378 TI - DNA vaccination against respiratory influenza virus infection. AB - DNA vaccination using plasmid encoding the hemagglutinin (HA) gene of influenza A/PR/8/34 virus to induce long-lasting protective immunity against respiratory infection was evaluated in this study. Using liposomes as carriers, the efficacy of DNA vaccines was determined using a lethal influenza infection model in mice. Mice immunized intranasally or intramuscularly with liposome-encapsulated pCI plasmid encoding HA (pCI-HA10) were completely protected against an intranasal 5 LD(50) influenza virus challenge. Mice immunized with liposome-encapsulated pCI HA10, but not naked pCI-HA10, by intranasal administration were found to produce high titers of serum IgA. These results suggest DNA vaccines encapsulated in liposomes are efficacious in inducing complete protective immunity against respiratory influenza virus infection. PMID- 11257379 TI - Post-exposure DNA vaccination protects mice against rabies virus. AB - Post-exposure anti-rabies vaccination for individuals who have not previously been immunized against rabies includes a cell culture-derived vaccine and a one time injection of rabies immune globulin. Recent studies have shown DNA vaccinations to be highly effective in rabies pre-exposure experiments, but post exposure protection has not been achieved. This failure is likely due to the slow onset of DNA vaccine induced antibody production. In an attempt to accelerate the onset of the antibody response, we manipulated variables, such as the route of vaccination and booster frequency. Anti-rabies virus antibody was detected 5 days after the initial DNA vaccination. Using this vaccination protocol and a single non-protective dose of anti-rabies immune serum, we questioned whether mice injected 6 h previously with rabies virus would be protected if a DNA vaccine was substituted for the cell culture-derived human diploid cell vaccine (HDCV). The DNA vaccine protected 87% of the mice (P = 0.00005, compared with unvaccinated control mice). Some 75% of mice receiving HDCV were protected (P = 0.00097, compared with unvaccinated control mice). Mice receiving only anti-rabies immune serum were not protected (P > 0.05 compared to unvaccinated control mice). Thus, post-exposure therapy, substituting a DNA vaccine for HDCV, did not compromise protection against rabies virus. PMID- 11257381 TI - Evaluation of genetic vaccine against classical swine fever. AB - Classical swine fever is important diseases affecting pigs. It results in great losses in their population and in limitations in the commercial international trade of pigs. The aim of the study was the preparation of the genetic vaccine against CSF and the estimation of its safety, protection value and immunogenicity. Clinical observations, body temperature and the immune response (haematological and FACS analyses) were monitored. Pigs vaccinated with the DNA vaccine were protected from the challenge, however, 2 days fever > 40 degrees C was registered. Slight activity of B and T cells was noted in those animals. PMID- 11257380 TI - Immunization of livestock with DNA vaccines: current studies and future prospects. AB - Early studies using DNA immunization suggest the potential benefits of this form of immunization including: long-lived immunity, a broad spectrum of immune responses (both cell-mediated immunity and humoral responses) and the simultaneous induction of immunity to a variety of pathogens through the use of multivalent vaccines. Using marine and cow models, we studied methods to enhance and direct the immune response to polynucleotide vaccines. We demonstrated the ability to modulate the magnitude and direction of the immune response by co administration of plasmid encoded cytokines and antigen. Also, we clearly demonstrated that the cellular components (cytosolic, membrane-anchored, or extracellular) to which the expressed antigen is delivered determines the types of immune responses induced. Since induction of immunity at mucosal surfaces (route of entry for many pathogens) is critical to prevent infection, various methods of delivering polynucleotide vaccines to animals including mucosal surfaces have been attempted and are described as future prospects for improving immune responses by DNA vaccination. PMID- 11257382 TI - Expansion of HBV-specific memory CTL primed by dual HIV/HBV genetic immunization during SHIV primary infection in rhesus macaques. AB - We have previously shown the induction of humoral and cytotoxic responses specific for human immunodeficiency virus (HIV) and hepatitis B virus (HBV) antigens, following genetic immunization of rhesus macaques with a plasmid encoding both the third variable domain of the HIV-1 external envelope glycoprotein and the pseudo-viral particle of hepatitis B surface antigen (HBsAg) as presenting molecules. The DNA-immunized primates and two control animals were then challenged with a chimeric simian/human immunodeficiency virus (SHIV). They were all infected. Significant frequencies of SHIV specific cytotoxic T lymphocyte precursors (CTLp) were detected early in peripheral blood. But, in all DNA-immunized macaques, HBV envelope specific CTLp were detected during the primary infection, and they were correlated with the peak of SHIV viremia. Furthermore, HBV or SHIV specific cytotoxicity corresponded in part to CD8(+) T cells presenting a memory phenotype. Several mechanisms could account for this cellular response. But our results suggest that an expansion of memory cytotoxic CD8(+) cells, not restricted to SHIV specific effectors, could occur in peripheral blood during SHIV primary infection. PMID- 11257383 TI - Modulation of antigen-specific cellular immune responses to DNA vaccination in rhesus macaques through the use of IL-2, IFN-gamma, or IL-4 gene adjuvants. AB - Extensive experiments have shown DNA vaccines' ability to elicit immune responses in vivo in a safe and well-tolerated manner in several model systems, including rodents and non-human primates. As the DNA-based vaccine and immunotherapy approaches are being explored in humans, significant efforts have also been focused on further improving the immune potency of this technology. One strategy to enhance immune responses for DNA vaccines is the use of molecular or genetic adjuvants. These molecular adjuvant constructs (which encodes for immunologically important molecules such as cytokines) can be co-administered along with DNA vaccine constructs. Once delivered, these adjuvants have shown to modulate the magnitude and direction (humoral or cellular) of the vaccine-induced immune responses in rodent models. To date, however, there has been very little data reported from studies in primates. In this study, we examined the effects of cytokine gene adjuvants to enhance the level of cell-mediated immune responses in rhesus macaques. We co-immunized rhesus macaques with expression plasmids encoding for IL-2, IFN-gamma or IL-4 cytokines along with the DNA vaccine constructs encoding for HIV env/rev (pCEnv) and SIV gag/pol (pCSGag/pol) proteins. We observed that coadministration of IL-2 and IFN-gamma cDNA resulted in enhancement of antigen-specific T cell-mediated immune responses. PMID- 11257384 TI - Gram-positive and Gram-negative bacteria as carrier systems for DNA vaccines. AB - Vaccination by intradermal or intramuscular injection of eukaryotic antigen expression vectors (so-called DNA vaccines) elicits strong cellular and humoral immune responses. A novel approach employs attenuated mutant strains of Gram positive and Gram-negative intracellular bacteria as carriers for the delivery of DNA vaccines. This strategy allows the administration of the DNA vaccines via mucosal surfaces and a direct delivery of the plasmid DNA to professional antigen presenting cells (APC), such as macrophages and dendritic cells (DC). In this work, we have found that several Gram-negative bacteria are capable of delivering plasmid vectors to human DC. In addition, we tested the suitability of the Gram positive bacterium Listeria monocytogenes as a vaccine carrier for the immunization of fish. PMID- 11257385 TI - Vaccines and mucosal immunisation. AB - The earliest attempts to protect humans against infectious diseases and toxins were by administering foreign substances to mucosal membranes, predominantly by the oral route. In the late 1880s, significant attention was given to the concept of 'local' immunisation, and the discipline of mucosal immunology was born in the early 1900s. However, despite the early enthusiasm, progress has been slow, with few mucosal vaccines being efficacious. The complexities of mucosal immune regulation and the lack of appropriate antigen delivery systems which can access mucosal inductive sites, have remained substantial obstacles. Recent studies demonstrating compartmentalisation of the common mucosal immune system create further challenges for the development of organ-specific vaccines. In the 21st century, our knowledge of mucosal immunoregulatory mechanisms, coupled with new technology for antigen delivery and immunomodulation will provide the necessary know-how to see the development and widespread use of mucosal vaccines for both preventative and therapeutic use. PMID- 11257386 TI - A therapeutic vaccine for mucosal candidiasis. AB - Persistent and recurrent infection of mucosal surfaces with Candida albicans is common, ranging from a nuisance to a life threatening clinical problem. No effective prophylactic or therapeutic vaccine has been developed. We have studied a mouse model of oral candida infection to identify regulatory and effector molecules of T cell activation as parameters of induced immunity, and here describe the use of this model to determine an optimal immunisation strategy. Oral immunisation with the blastospore yeast form (but not subcutaneous immunisation) induced clinical immunity, with a shift in parameters of cytokine response characterised by an early and sustained production of both IFN-gamma and IL-4 from antigen-stimulated cervical node T lymphocytes. PMID- 11257387 TI - Programmed inflammatory processes induced by mucosal immunisation. AB - Inflammation is essential to repair tissue damaged by physical, microbial or allergic mechanisms. Inappropriately zealous responses lead to destructive pathology or chronic disease cycles, whereas ideal outcomes are associated with complete and rapid restoration of tissue structure and function. The establishment of a rodent model investigating the different immune responses to non-typeable Haemophilus influenzae infection in both the lung and the ear indicate an ability to clear bacteria and reduce inflammation following mucosal immunisation. Lung histochemistry, upregulaion of macrophages and polymorphonuclear neutrophils, recruitment of gammadelta(+) and CD8(+) T cells, cytokine levels and depletion studies all support the hypothesis that mucosal immunisation facilitates control of the immune response resulting in enhanced bacterial clearance and programming of inflammation which limits damage and promotes the rapid restoration of structural normality. PMID- 11257388 TI - Mucosal immunity in the lung and upper airway. AB - The mucosal surfaces of the lungs and upper airways are common sites for infection. Extensive studies of the mechanisms associated with immune responses in the respiratory tract have found that understanding the system is challenging and involves many complex interactions to prevent and eliminate infection. Immune protection against diseases transmitted through the respiratory tract requires an understanding of the important aspects associated with beneficial, detrimental or ineffective immune responses. Two critical aspects of an immune response against a pathogen are that of the inductive stage, either induced by vaccination or primary infection, and the effector stage, the ability to recognise, respond to and eliminate the infection without detriment to the host. An immunisation strategy must not only have a measure of the induced antigen specific response, but this response must also be protective. PMID- 11257389 TI - Mucosal vaccines: non toxic derivatives of LT and CT as mucosal adjuvants. AB - Most vaccines are still delivered by injection. Mucosal vaccination would increase compliance and decrease the risk of spread of infectious diseases due to contaminated syringes. However, most vaccines are unable to induce immune responses when administered mucosally, and require the use of strong adjuvant on effective delivery systems. Cholera toxin (CT) and Escherichia coli enterotoxin (LT) are powerful mucosal adjuvants when co-administered with soluble antigens. However, their use in humans is hampered by their extremely high toxicity. During the past few years, site-directed mutagenesis has permitted the generation of LT and CT mutants fully non toxic or with dramatically reduced toxicity, which still retain their strong adjuvanticity at the mucosal level. Among these mutants, are LTK63 (serine-to-lysine substitution at position 63 in the A subunit) and LTR72 (alanine-to-arginine substitution at position 72 in the A subunit). The first is fully non toxic, whereas the latter retains some residual enzymatic activity. Both of them are extremely active as mucosal adjuvants, being able to induce very high titers of antibodies specific for the antigen with which they are co administered. Both mutants have now been tested as mucosal adjuvants in different animal species using a wide variety of antigens. Interestingly, mucosal delivery (nasal or oral) of antigens together with LTK63 or LTR72 mutants also conferred protection against challenge in appropriate animal models (e.g. tetanus, Helicobacter pylori, pertussis, pneumococci, influenza, etc). In conclusion, these LTK63 and LTR72 mutants are safe adjuvants to enhance the immunogenicity of vaccines at the mucosal level, and will be tested soon in humans. PMID- 11257390 TI - The B cell targeted adjuvant, CTA1-DD, exhibits potent mucosal immunoenhancing activity despite pre-existing anti-toxin immunity. AB - We recently developed a novel immunomodulating gene fusion protein, CTA1-DD, that combines the ADP-ribosylating ability of cholera toxin (CT) with a dimer of an Ig binding fragment, D, of Staphylococcus aureus protein A. The CTA1-DD adjuvant was found to be non-toxic and greatly augmented T cell dependent and independent responses. Following injection it binds to both naive and memory B cells and up regulates co-stimulatory molecules as well as prevents apoptosis of activated B cells. Here we show that CTA1-DD is a potent mucosal adjuvant administered intranasally. A dose-response analysis revealed that the adjuvant effect of CTA1 DD given intranasally was equally strong to that observed after systemic immunizations. The adjuvant effect was independent of any possible contamination with endotoxin as indicated by the similar enhancing effects of CTA1-DD in C3H/HeN and the LPS-insensitive C3H/HeJ mice. Contrary to many other adjuvants CTA1-DD induces an immune response to itself. However, despite the presence of high serum titers of pre-existing anti-CTA1 antibodies we observed no reduction of the adjuvant function of CTA1-DD when given either intranasally or systemically. These results support the notion that the CTA1-DD adjuvant can repeatedly be used in the clinic without loss of efficacy even when pre-existing anti-CTA1 antibody levels are high. PMID- 11257391 TI - Clinical studies of human papilloma vaccines in pre-invasive and invasive cancer. AB - Cervical cancer is the second most common cause of cancer death in women worldwide. It is almost invariably associated with infection with human papilloma virus (HPV) particularly types 16 and 18. The ubiquitous expression of E6 and E7 oncogene products has been recognised as an attractive target for CTL-mediated immunotherapy. In-vivo expansion of an HPV oncogene product specific MHC class 1 restricted response has been demonstrated using intradermally administered live vaccinia virus HPV 16 and 18 E6/E7 gene construct (TA-HPV, Cantab Pharmaceuticals). Responses have been seen in 1/3 evaluable patients with advanced cervical cancer, and 3/12 CIN3 volunteers, and in 4/29 patients with early invasive cervical cancer (Rankin et al. Proceedings of 91st AACR Meeting, San Francisco, April 2000). In addition, the adoptive transfer of ex vivo HPV 16 or 18 positive autologous tumour lysate pulsed dendritic cells is currently being tested as an alternative means of expanding HPV specific CTL in advanced cervical cancer patients. So far an HLA-A*O201 restricted CD8 T cell response has been recorded in the single HLA-A*O201 patient whose tumour was shown to be HPV16 positive. It appears therefore feasible to induce HPV specific CTL responses in patients with cervical cancer using several vaccine strategies. However, further clinical trials are needed to determine the full anti-tumour potential of this vaccine based immunotherapy. PMID- 11257392 TI - The organotypic culture of HPV-transformed keratinocytes: an effective in vitro model for the development of new immunotherapeutic approaches for mucosal (pre)neoplastic lesions. AB - The purpose of this study is to develop a reliable in vitro human model to test new immunotherapeutic approaches for squamous cell carcinoma that develop on mucosal surfaces. The organotypic (raft) culture permits cells to proliferate and differentiate at an air-liquid interface on a dermal equivalent support. Normal keratinocytes stratify and fully differentiate in a manner similar to the normal squamous epithelial tissues, while human papillomavirus-immortalized and established squamous carcinoma cell lines exhibit dysplastic morphologies similar to (pre)neoplastic lesions seen in vivo. We have demonstrated the ability of these organotypic cultures to be manipulated by altering the epithelial stratification with cytokines (interferon-gamma and tumor necrosis factor-alpha) and by integrating activated lymphocytes or dendritic cells into the in vitro formed epithelial sheet. This model may provide a useful tool to investigate the factors contributing to the presence and function of immunocompetent cells within a neoplastic epithelium that develops on a mucosal surface. PMID- 11257393 TI - Autologous, hapten-modified vaccine as a treatment for human cancers. AB - We have devised a novel approach to active immunotherapy based on modification of autologous cancer cells with the hapten, dinitrophenyl (DNP). The treatment program consists of multiple intradermal injections of DNP-modified autologous tumor cells mixed with BCG. Administration of DNP-vaccine to patients with metastatic melanoma induces a unique reaction - the development of inflammation in metastatic masses. Histologically, this consists of infiltration of T lymphocytes, most of which are CD8+. These T cells usually produce gamma interferon in situ. Moreover, they represent expansion of T cell clones with novel T cell receptor structures. Occasionally, administration of DNP-vaccine results in partial or complete regression of measurable metastases. The most common site of regression has been small lung metastases. Administration of DNP vaccine to patients in the post-surgical adjuvant setting produces a more striking clinical effect. We have treated 214 patients with clinically evident stage III melanoma who had undergone lymphadenectomy. With a median follow-up time of 4.4 years (1.8-10.4 years) the 5-year overall survival (OS) rate is 47% (one nodal site = 51%, two nodal sites = 33%). These results appear to be comparable to those obtained with high dose interferon. More recent studies suggest that this therapeutic approach is also applicable to ovarian cancer. There appear to be no insurmountable impediments to applying this approach to much larger numbers of patients or to developing it as a standard cancer treatment. PMID- 11257395 TI - Adjuvant active specific immunotherapy of stage II and stage III colon cancer with an autologous tumor cell vaccine: first randomized phase III trials show promise. AB - We performed three multi-institutional, prospectively randomized, controlled clinical trials, assessing the therapeutic effect of post-resection adjuvant active specific immunotherapy in patients with stage II and stage III colon cancer. In each study four outcomes were considered: time-to-disease recurrence, overall survival intervals, disease-free survival intervals, and recurrence-free survival intervals using the Kaplan-Meir method for generating curves and the log rank test used to compare efficacy distributions. In addition, a meta-analysis of the three phase III trials was performed since the trials had proven homogeneity. Two main analyses were performed: (1) the intent-to-treat colon cancer patients from all three studies; and (2) analyzable colon cancer patients in all three studies. The conclusion of these analyses is that adjuvant active specific immunotherapy provided significant clinical benefits in patients with stage II colon cancer and appears to be an important new adjuvant treatment for these patients. PMID- 11257394 TI - Therapeutic vaccines against melanoma and colorectal cancer. AB - Our overall strategy is to develop multivalent recombinant vaccines capable of eliciting broad immune responses in patients with malignant melanoma or colorectal cancer. We report herein results from initial studies conducted in cancer patients to evaluate the effect of intratumoral administration of recombinant canarypox viruses carrying cytokine genes. Our current focus is on the induction of tumor-specific T-cell responses using a prime/boost immunization schedule with a unique vector system derived from the canary pox virus called ALVAC, in which we incorporate genes encoding Tumor Associated Antigens (TAAs) of interest. Clinical studies in colorectal cancer evaluating an ALVAC CEA candidate vaccine have shown that this approach is safe and can induce tumor-specific T cell responses. Additional clinical studies evaluating candidate vaccines against melanoma and colorectal cancer, targeting either the gp100, Mage 1, Mage 3 or p53 molecules are ongoing. PMID- 11257397 TI - Heat shock proteins: the 'Swiss Army Knife' vaccines against cancers and infectious agents. AB - The ability of heat shock proteins to: (a) chaperone peptides, including antigenic peptides; (b) interact with antigen presenting cells through a receptor; (c) stimulate antigen presenting cells to secrete inflammatory cytokines; and (d) mediate maturation of dendritic cells, makes them a one-stop shop for the immune system. These properties also permit the utilization of heat shock proteins for development of a new generation of prophylactic and therapeutic vaccines against cancers and infectious diseases. PMID- 11257396 TI - Therapeutic potential of protein and adjuvant vaccinations on tumour growth. AB - Over 90% of cervical cancers are associated with HPV infection, the commonest being the HPV-16 subtype. Two early viral genes, E6 and 7, play major roles in the development and maintenance of the malignant phenotype. The vaccine potential of a recombinant HPV16 E7 protein was examined in two murine models of E7 expressing tumours. Formulations including the immunostimulants MPL and QS21 induced therapeutically active immune responses leading to regression of pre established TC1 tumour lesions, associated with induction of IgG antibodies, lymphoproliferation and CTL. Our data provide a clear incentive to investigate the clinical application of this approach in cancer immunotherapy. PMID- 11257398 TI - Vaccination with Her-2/neu DNA or protein subunits protects against growth of a Her-2/neu-expressing murine tumor. AB - The present study utilizes an in vivo murine tumor expressing human Her-2/neu to evaluate potential Her-2/neu vaccines consisting of either full length or various subunits of Her-2/neu delivered in either protein or plasmid DNA form. Our results demonstrate that protective immunity against Her-2/neu-expressing tumor challenge can be achieved by vaccination with plasmid DNA encoding either full length or subunits of Her-2/neu. Partial protective immunity was also observed following vaccination with the intracellular domain (ICD), but not extracellular domain (ECD), protein subunit of Her-2/neu. The mechanism of protection elicited by plasmid DNA vaccination appeared to be exclusively CD4 dependent, whereas the protection observed with ICD protein vaccination required both CD4 and CD8 T cells. PMID- 11257399 TI - Using in silico transcriptomics to search for tumor-associated antigens for immunotherapy. AB - Immunotherapy approaches to fight cancer are based on the principle of mounting an immune response against a self-antigen expressed by the tumor cells. In order to reduce potential autoimmunity side-effects, the antigens used should be as tumor-specific as possible. A complementary approach to experimental tumor antigen discovery is to screen the human genome in silico, particularly the databases of "Expressed Sequence Tags" (ESTs), in search of tumor-specific and tumor-associated antigens. The public databases currently provide a massive amount of ESTs from several hundreds of cDNA tissue libraries, including tumoral tissues from various types. We describe a novel method of EST database screening that allows new potential tumor-associated genes to be efficiently selected. The resulting list of candidates is enriched in known genes, described as being expressed in tumor cells. PMID- 11257400 TI - Generation of auto-antibodies towards Alzheimer's disease vaccination. AB - We developed a novel procedure to evoke anti-aggregating beta-amyloid (Abeta) antibodies, using filamentous phages displaying only four amino acids EFRH of the beta-amyloid peptide (AbetaP). This epitope was found to be the main regulatory site for fibril formation. For the first time, effective auto-antibodies have been obtained in guinea-pigs, which exhibit human identity in the AbetaP. Immunization through a phage-carrying epitope was found to be long-lasting, and no toxic effect caused by autoimmune response was detected in the challenged animal sections. These antibodies performed similarly to site-directed monoclonal antibodies and to antibodies raised against fibrillar Abeta in disaggregation of plaques, and may serve as the basis for developing an anti-Abeta vaccine. PMID- 11257401 TI - Recombinant attenuated bacteria for the delivery of subunit vaccines. AB - Using attenuated intracellular bacteria as carriers, we have developed two different approaches for the delivery of subunit vaccines encoding heterologous antigens. The first system is based on the direct secretion of the heterologous antigens in Gram-negative bacteria via the hemolysin secretion system of Escherichia coli into either phagosome or cytosol of infected cells. The second approach is based on the transport of eukaryotic antigen expression vectors by intracellular bacteria like Listeria and Salmonella into the host cell and here, preferably, into the cytosolic compartment. After release of the plasmid DNA from the bacteria, the plasmid-encoded antigens can be expressed directly by the host cell. Finally, we combined both types of subunit vaccines in one live vector - we equipped Salmonella strains with a phagosomal escape function by utilization of the hemolysin secretion system and used this recombinant vaccine strain for the delivery of a eukaryotic antigen expression vector into the cytosol of macrophages. PMID- 11257402 TI - A powder formulation of measles vaccine for aerosol delivery. AB - Both the mortality rate for measles and the risks associated with injection continue to be high in the developing world. In response to the need for safe, cost-effective vaccine delivery technologies, a powder formulation of measles vaccine has been developed to test the feasibility of administering measles vaccine as an aerosol. The first challenge in aerosol formulation development is to produce fine particles without damaging the activity of the virus or inducing physical changes. In this study, live attenuated measles vaccine is micronized by jet milling to generate particle sizes appropriate for pulmonary delivery (1-5 microm). Milling does not induce detectable physical changes and significant viral potency is maintained. Potency retention of milled vaccine ranges from 31 to 89%, demonstrating that the standard dose of vaccine can easily be achieved. Following size reduction, particles are blended with an inert carrier to improve handling and aerosol dispersion. The measles vaccine formulation is dispersable, as shown by laser light particle size analysis of vaccine aerosols. Thus, evaluation of both the potency retention and the aerosol characteristics of the current formulation clearly demonstrates the feasibility of delivering measles vaccine as a powder aerosol for immunization. PMID- 11257403 TI - The efficacy of oral vaccination of mice with alginate encapsulated outer membrane proteins of Pasteurella haemolytica and One-Shot. AB - The goal of this study was to examine the efficacy of oral delivery of alginate encapsulated outer membrane proteins (OMP) of Pasteurella haemolytica and a commercial One-Shot vaccine in inducing protection in mice against lethal challenge with virulent P. haemolytica. We examined two alginate microsphere formulations and compared them with oral unencapsulated and subcutaneously administered vaccines. Alginate microspheres were made by the emulsion-cross linking technique. They were examined for size, hydrophobicity, and antigen loading efficiency before they were used in the study. Mice were vaccinated by administering 200 microg of antigens in 200 microl of microspheres suspension orally or subcutaneously. One group of mice received blank microspheres and a second group was given unencapsulated antigen orally. A third and a fourth group received different formulations of alginate encapsulated antigens by oral administration. Three groups received subcutaneous inoculations (alginate encapsulated, non-adjuvanted and unencapsulated antigens, and adjuvanted One Shot), and one group received water (naive group). Mice were vaccinated orally for four consecutive days and challenged with P. haemolytica 5 weeks after the first vaccination. Weekly serum and feces samples were assayed for antigen specific antibodies. The number of dead mice in each group 4 days post challenge was used to compare the efficacy of the various vaccination groups. The mean volume sizes of blank alginate microsphere formulations A, and AA were 15.9, 16 and 9.2 microm, respectively. Hydrophobicity of the microspheres was evaluated by measuring contact angle on a glass slide coated with the microspheres. The contact angles on A and AA were 37.8 and 74.3 degrees, respectively. Antigen concentration in a 1:1 w/w suspension of microspheres in water was 0.9 mg/ml. Rate of death for the blank group was 42.8% whereas for groups vaccinated with antigens encapsulated in A and AA the death rates were 40 and 33.33%, respectively. The death rate in mice vaccinated with unencapsulated antigens was 55.6%. Groups vaccinated by subcutaneous inoculation showed the lowest death rate. These results show that encapsulating OMP and One-Shot in alginate microspheres improves their performance as an oral vaccine. PMID- 11257404 TI - Genetic adjuvants for DNA vaccines. AB - The relatively low efficacy of DNA vaccines in inducing immune responses, especially in large animal species and humans, has impaired their practical use. Despite considerable effort expended on improving DNA vaccine delivery, only minute amounts of Ag are available for immune induction following DNA vaccination. Two complementary strategies have been used to improve and modulate the immune response induced by DNA vaccines: (i) supplementing DNA vaccines with plasmids encoding cytokines and (ii) targeting the Ag encoded by DNA vaccine through genetically fusing the Ag to molecules binding cell surface receptors. This paper reviews recent progress in these two areas and possible mechanisms responsible for the observed effects. PMID- 11257405 TI - The potential of oligodeoxynucleotides as mucosal and parenteral adjuvants. AB - Synthetic oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs (CpG ODN) are potent adjuvants in mice when delivered by parenteral (intramuscular, subcutaneous) and mucosal (intranasal, oral and intrarectal) routes. We have recently shown that with mucosal delivery non-CpG ODN can also have immunostimulatory properties which, in contrast to the Th1-bias characteristic of CpG ODN, are predominantly Th2-like. Herein, using hepatitis B surface antigen (HBsAg) and tetanus toxoid (TT) as model antigens in BALB/c mice, we have examined a number of different ODN (CpG, non-CpG, poly-T, poly-CG) to determine their effects on immune responses after mucosal (oral) and parenteral (IM) immunizations. Our findings demonstrate that with mucosal delivery, there is a Th2-biased immunostimulatory effect that is associated with non-CpG ODN, and that the presence of CpG motifs can shift this towards a Th1 response. The adjuvant effect of non-CpG ODN was much less evident after parenteral immunization. PMID- 11257406 TI - Immune responses to ISCOM formulations in animal and primate models. AB - ISCOMs are typically 40 nm cage-like structures comprising antigen, saponin, cholesterol and phospholipid. ISCOMs have been shown to induce antibody responses and activate T helper cells and cytolytic T lymphocytes in a number of animal species, including non-human primates. Recent clinical studies have demonstrated that ISCOMs are also able to induce antibody and cellular immune responses in humans. This review describes the current understanding of the ability of ISCOMs to induce immune responses and the mechanisms underlying this property. Recent progress in the characterisation and manufacture of ISCOMs will also be discussed. PMID- 11257407 TI - Adjuvants designed for veterinary and human vaccines. AB - Adjuvants play an important role in the efficacy of vaccines as the antigens become more and more purified. Indeed recombinant proteins or synthetic peptides are safer than crude inactivated micro-organism, but less immunogenic. This can be balanced by specific adjuvants. But there is no universal adjuvants and their action is not yet clear and rely on different mechanisms. Then, they must be adapted according to several criteria, like the target species, the antigens, the type of immune response, the route of inoculation, or the duration of immunity. For this purpose different type of emulsions have been developed. Water in oil (W/O) emulsions induce a strong and long term immune response. Those based on mineral oils are known to be very efficient but can sometimes induce local reactions with reactive antigens. Non mineral oils are well tolerated but less efficient with poor immunogens. Multiphasic (W/O/W) emulsions can induce short and long term immune responses with various antigens and oil in water (O/W) emulsions are well tolerated and induce a short term immune response. New generation of adjuvants are based on a new concept called 'immunosol' and stem from the association of nanoparticles with a new immunostimulant. They can be used when emulsions are not suitable to obtain a good balance between safety and immunogenicity. PMID- 11257408 TI - The adjuvanted influenza vaccines with novel adjuvants: experience with the MF59 adjuvanted vaccine. AB - Elderly people and subjects with underlying chronic diseases are at increased risk for influenza and related complications. Conventional influenza vaccines provide only limited protection in the elderly population. In order to enhance the immune response to influenza vaccines, several adjuvants have been evaluated. Among these, an oil in water adjuvant emulsion containing squalene, MF59, has been combined with subunit influenza antigens and tested in clinical trials in comparison with non-adjuvanted conventional vaccines. Data from a clinical database of over 10000 elderly subjects immunised with this adjuvanted vaccine (Fluad, Chiron Vaccines, Siena, Italy) demonstrate that, although common postimmunisation reactions are more frequent in recipients of the adjuvanted vaccine, this vaccine is well tolerated, also after re-immunisation in subsequent influenza seasons. Immunogenicity analyses demonstrate a consistently higher immune response with statistically significant increases of postimmunisation geometric mean titres, and of seroconversion and seroprotection rates compared to non-adjuvanted subunit and split influenza vaccines, particularly for the A/H3N2 and the B strains. The higher immunogenicity profile of the MF59-adjuvanted vaccine is maintained also after subsequent immunisations. An even higher adjuvant effect was shown in subjects with low pre-immunisation titre and in those affected by chronic underlying diseases. In conclusion, the addition of MF59 to subunit influenza vaccines enhances significantly the immune response in elderly subjects without causing clinically important changes in the safety profile of the influenza vaccine. PMID- 11257409 TI - Technologies for the design, discovery, formulation and administration of vaccines. AB - This paper summarizes major technologies, with emphasis on applications to preventive vaccines for infectious diseases. A limited number of examples of each technology are provided. PMID- 11257410 TI - Reverse vaccinology, a genome-based approach to vaccine development. AB - The conventional approach to vaccine development requires cultivation of the pathogenic microorganism and its dissection using biochemical, immunological, and microbiological methods in order to identify the components important for immunity. This method, while successful in many cases, failed to provide a solution for many of those pathogens for which a vaccine is not yet available. Today, the possibility of using genomic information allows us to study vaccine development in silico, without the need of cultivating the pathogen. This approach, which we have named 'reverse vaccinology', reduces the time required for the identification of candidate vaccines and provides new solutions for those vaccines which have been difficult or impossible to develop. The potential of this new approach is illustrated by the use of reverse vaccinology for the development of a vaccine against serogroup B meningococcus. The application of reverse vaccinology to other fields, including viral vaccines is discussed. PMID- 11257411 TI - The role of mass spectrometry in vaccine development. AB - For the most part, vaccine development to date has been empiric. While sometimes successful, such a strategy is 'hit or miss', and fails to advance the basic science of vaccine development. Preferable would be tools that allow for a more directed development of vaccines at either the population or sub-population level. Characteristics of useful tools in vaccine development should include the ability to identify and characterize the spectrum of antigenic peptides presented by MHC molecules to which the immune system responds by the development of protective immune responses. In addition, because the explosion in human genomics allows the ability to understand MHC haplotypes at the population level, as well as an enhanced understanding of MHC binding motifs, new tools might further allow for an understanding of which vaccine antigens are capable of being bound and presented to the immune system by MHC molecules. New mass spectrometry technology fulfils these criteria, and may well lead to a revolution in the design of new vaccines. This paper will review the basics of mass spectrometry techniques as applied to the identification and characterization of vaccine peptide antigens, and discusses how these tools can be applied to vaccine development. PMID- 11257412 TI - Transcutaneous immunization: T cell responses and boosting of existing immunity. AB - Transcutaneous immunization (TCI) is a novel immunization strategy by which antigen and adjuvant are applied topically to intact, hydrated skin to induce potent antibody and cell-mediated immune responses specific for both the antigen and the adjuvant. Using tetanus toxoid as a model antigen, we examined the T cell response to tetanus toxoid after topical immunization with a variety of adjuvants. TCI readily induced systemic antigen specific T cell responses with a mixed Th1/Th2 phenotype but with a Th2 bias. We also investigated whether priming by the intramuscular route, which is known to induce T cell memory, could be followed by a boosting immunization on the skin to induce secondary responses. TCI could augment existing immunity, but interestingly, this strategy induced potent responses only if the antibody titer was low at the time of TCI boosting. These and previous observations suggest that TCI follows known immunological principles that govern other routes of vaccine delivery. Furthermore, booster immunization using tetanus toxoid may provide a useful model for further development of important patch and formulation concepts for TCI, and act as an early candidate for validating product feasibility of TCI in humans. PMID- 11257413 TI - The bare skin and the nose as non-invasive routes for administering peptide vaccines. AB - Among the different technologies currently tested for the development of novel vaccines, synthetic peptides represent a promising option, since they are chemically pure and induce immune responses of predetermined specificity. Furthermore, they can be replaced with pseudopeptides or peptide mimetics that contain changes in the amide bond, resulting in more stable and immunogenic molecules. Administration of peptide vaccines via non-invasive routes, such as the nose or the bare skin, allows the efficient uptake of antigen by antigen presenting cells, which are abundant in the associated lymphoid tissues, ensuring the induction of effective systemic and mucosal immune responses. Using non invasive routes could be advantageous for vaccination programs in third-world countries, since vaccine administration is simple, painless and economical. In this review, we discuss and present some preliminary data on the advantages of synthetic peptides and peptidomimetics as candidate vaccines, and their potential for administration via the skin and the nose. PMID- 11257414 TI - The human cell line PER.C6 provides a new manufacturing system for the production of influenza vaccines. AB - Influenza viruses for vaccine production are currently grown on embryonated eggs. This manufacturing system conveys many major drawbacks such as inflexibility, cumbersome down stream processing, inability of some strains to replicate on eggs to high enough yields, and selection of receptor-binding variants with reduced antigenicity. These limitations emphasize the need for a cell line-based production system that could replace eggs in the production of influenza virus vaccines in a pandemic proof fashion. Here we present the efficient propagation of influenza A and B viruses on the fully characterized and standardized human cell line PER.C6. PMID- 11257415 TI - Herd immunity after vaccination: how to quantify it and how to use it to halt disease. AB - In comparison to unvaccinated individuals, vaccinated individuals have fewer clinical symptoms, reduced susceptibility and reduced infectivity. The first two effects of vaccination can mean that each vaccinated individual is protected against clinical symptoms. From experiments and field trials, the extent of individual protection can be determined by a statistical analysis of the resulting data. In addition, there is an effect of the vaccination on the populations in which one or more individuals are vaccinated. This effect on the population is due to the effects of vaccination on susceptibility and infectivity of the vaccinated individuals. The population effect is called herd immunity and is observed as a reduction in chance of becoming infected when being part of a population with some of the individuals vaccinated. Note that the protection by herd immunity applies to vaccinated individuals as well as to unvaccinated individuals. Thus, protection against disease can be achieved not only by vaccinating the individuals that have to be protected but also by vaccinating other individuals in the same population. Such an application of herd immunity is especially important in protecting farm animals. To plan and evaluate vaccination at the population level, the herd immunity needs to be quantified. It will be illustrated that it is possible, not only theoretically but also practically, to quantify herd immunity among farm animals with data from small-scale experiments as well as with data from field trials. PMID- 11257416 TI - An in vitro approach in quality control of toxoid vaccines. AB - For routine immunogenicity testing of traditionally produced vaccines, animal tests are required by regulatory authorities, with potency estimated in International Units. A new concept focuses on assuring immunogenicity by monitoring batch-to-batch consistency in production. This concept is used for well-defined biologicals such as hormones. Through the use of immunochemical and bio- and physiochemical techniques the traditional products can be characterised as completely as possible. Developments in in vitro methodologies offer opportunities for immunogenicity testing in vitro. This study describes the possibilities for applying the consistency concept to the traditional products, tetanus and diphtheria toxoids. The sources of variation in these products were studied by flocculation time, SDS-PAGE, biosensor analysis, gel permeation chromatography and in vitro cytokine production studies. Batch-to-batch variation was shown using these in vitro techniques. Results indicate that it is possible to apply the consistency concept in the quality control of traditional vaccines like tetanus and diphtheria toxoids. PMID- 11257417 TI - The green revolution: plants as heterologous expression vectors. AB - Agrobacterium tumefaciens mediated gene transfer into the plant genome laid the groundwork for new procedures aimed at crop improvement, including resistance to pathogens, increased product yield, modified oil content, and resistance to environmental stress conditions. New developments in molecular plant virology have led to the generation of plant-based systems for transient expression of foreign sequences using plant virus vectors. In the last decade both transgenic plants and plant virus vectors have been used increasingly to produce a wide range of biomedical reagents, including vaccine antigens, in a safe and economically feasible manner. These new plant-based technologies have enormous potential for a variety of applications, including the oral delivery of vaccine antigens. PMID- 11257418 TI - Plant-based vaccines: unique advantages. AB - Numerous studies have shown that viral epitopes and subunits of bacterial toxins can be expressed and correctly processed in transgenic plants. The recombinant proteins induce immune responses and have several benefits over current vaccine technologies, including increased safety, economy, stability, versatility and efficacy. Antigens expressed in corn are particularly advantageous since the seed can be produced in vast quantities and shipped over long distances at ambient temperature, potentially allowing global vaccination. We have expressed the B subunit of Escherichia coli heat-labile enterotoxin and the spike protein of swine transmissible gastroenteritis virus at high levels in corn, and demonstrate that these antigens delivered in the seed elicit protective immune responses. PMID- 11257420 TI - Production of antibodies and antibody fragments in plants. AB - Our current knowledge allows the generation of transgenic plants that efficiently produce heterologous proteins from plant, bacterial, fungal or animal origin. Among all types of recombinant proteins, antibodies are particularly attractive because of their ability to specifically recognize and bind virtually any type of antigen. Plants show several advantages as a large-scale antibody production system: they can be grown easily and inexpensively in large quantities that can be harvested, stored and processed by using existing infrastructures. Isolation and purification of plant-made antibodies, if necessary, allow fundamental, industrial, and therapeutical applications. In the past, we and others have successfully generated antibody-producing plants. The maximal accumulation levels of antibodies and antibody fragments that we observed are 1-5% of the extracted proteins. Currently, several biotechnological companies grow field crops to produce antibodies for ex planta applications on an industrial scale. PMID- 11257419 TI - Oral immunisation of naive and primed animals with transgenic potato tubers expressing LT-B. AB - The efficacy of edible vaccines produced in potato tubers was examined in mice. Transgenic plants were developed by Agrobacterium tumefaciens-mediated transformation. The antigen selected was the non-toxic B subunit of the Escherichia coli enterotoxin (recLT-B). A synthetic gene coding for recLT-B was made and optimised for expression in potato tubers and accumulation in the endoplasmic reticulum. Introduction of this gene under control of the tuber specific patatin promoter in potato plants resulted in the production of functional, i.e. Gm1-binding, recLT-B pentamers in tubers. Selected tubers containing about 13 microg of recLT-B per gram fresh weight were used for immunisation. Subcutaneous immunisation with an extract of recLT-B tubers yielded high antibody titres in serum that were similar to those obtained with bacterial recLT-B. The efficacy of oral administration of recLT-B tubers was determined by measuring mucosal and systemic immune responses in naive and primed mice. Animals were primed by subcutaneous injection of an extract of recLT-B tuber plus adjuvant. Naive and primed mice were fed 5 g of tubers ( approximately 65 microg of recLT-B) or were intubated intragastrically with 0.4 ml of tuber extract ( approximately 2 microg of recLT-B). In naive mice, feeding recLT-B tubers or intubation of tuber extract did not induce detectable anti-LT antibody titres. In primed animals, however, oral immunisation resulted in significant anti-LT IgA antibody responses in serum and faeces. Intragastric intubation of tuber extract revealed higher responses than feeding of tubers. These results indicate clearly that functional recLT-B can be produced in potato tubers, that this recombinant protein is immunogenic and that oral administration thereof elicits both systemic and local IgA responses in parentally primed, but not naive, animals. PMID- 11257421 TI - Uptake and efflux of the peptidic delta-opioid receptor agonist. AB - P-glycoprotein (P-gp) and organic anion transporting polypeptides (Oatp) are expressed at the blood-brain barrier (BBB). There is little functional evidence for Oatp-mediated transport at the BBB. The peptidic delta opioid-receptor agonist [D-penicillamine(2,5)]-enkephalin (DPDPE) is a substrate of mdr1a P-gp and Oatp2. The present study evaluated the influence of these transporters on brain uptake of DPDPE by in situ perfusion in mice. Brain uptake was increased approximately 12-fold in mice lacking P-gp in the BBB, but the P-gp inhibitor dexverapamil did not increase uptake in P-gp-competent mice. In P-gp-deficient mice, DPDPE uptake was saturable (K(m) approximately 24 mM), and was inhibited by dexverapamil and the Oatp2 substrates digoxin, estradiol-17beta-glucuronide and fexofenadine. These results confirm P-gp-mediated efflux of DPDPE, and suggest functional uptake transport of DPDPE by Oatp, at the murine BBB. PMID- 11257422 TI - Presence of different ATP receptors on rat midbrain single synaptic terminals. Involvement of the P2X(3) subunits. AB - Adenosine 5'-triphosphate (ATP) stimulates a [Ca(2+)](i) increase via specific ionotropic receptors, termed P2X receptors, in rat midbrain presynaptic terminals. A microfluorimetric technique enabled study of the [Ca(2+)](i) increase in isolated single synaptic terminals, showing that 33.4+/-2.5% of them responded to ATP. Immunological studies carried out, after functional studies, with specific anti-P2X receptor subunit antibodies showed only positive labelling with anti-P2X(3) antibodies in 23.5+/-1.7% of the terminals. All positively P2X(3) labelled synaptic terminals responded to ATP. Nevertheless, not all of them responded to alpha,beta-meATP, these representing 6.7+/-1.5% of the total. In addition, 9.8+/-2.3% of the terminals responded to ATP but exhibit negative P2X(3)-labelling. These results demonstrate the existence of a heterogeneous population of ionotropic ATP receptors at the presynaptic level. PMID- 11257423 TI - Enlargement of the fornix in early-onset schizophrenia: a quantitative MRI study. AB - Abnormalities of temporal lobe structure and frontal lobe function occur in schizophrenia. There have been few studies of young people with schizophrenia and little is known about temporo-frontal connectivity in the disease. Therefore, the cross-sectional area of the body of the fornix was measured on MR images from 17 young people with schizophrenia, nine with other serious psychiatric illnesses and eight without illness. The mean age of each group was 16-17 years. The mean cross-sectional fornix area in subjects with schizophrenia was significantly larger than that in subjects without illness ( approximately 40%) and psychiatric controls ( approximately 26%). There was no such significant difference between subjects without illness and psychiatric controls. The nature of the larger fornix in early-onset schizophrenia, whether it persists and whether it occurs in schizophrenia presenting in adulthood, remain to be elucidated. PMID- 11257424 TI - Determination of 14-3-3 protein levels in cerebrospinal fluid from Creutzfeldt Jakob patients by a highly sensitive capture assay. AB - The level of 14-3-3gamma protein was determined in the cerebrospinal fluid (CSF) from patients with Creutzfeldt-Jakob disease (CJD) and non-CJD patients applying a new and fast microplate assay (14-3-3 protein capture assay), based on the binding to a peptide comprising a phosphorylated recognition motif of 14-3-3 protein. The levels of the gamma-isoform of 14-3-3 protein in CSF samples from CJD patients (n=41) were significantly higher than those observed in patients with non-CJD dementias (n=36) suggesting that this capture assay is a reliable method in the diagnosis of CJD. Since this assay allows the direct measurement of 14-3-3 protein in the CSF without prior concentration it is an easy and simple alternative to the conventionally applied immunoblotting procedures. PMID- 11257425 TI - Energetic metabolism in mouse cerebral cortex during chronic hypoxia. AB - We measured the activities of Na(+)K(+) ATPase and of enzymes of the glycolytic pathway, Krebs cycle, and the respiratory chain in cerebral cortex of mice exposed to chronic hypoxia for three weeks and compared their values with those of sea level controls. There were no differences in Na(+)K(+) ATPase activity or in the activity of glycolytic enzymes. In the Krebs cycle, a 66% increase of succinate dehydrogenase activity was found due to a lower Km. In contrast, respiratory chain cytochrome oxidase activity was reduced by 12% in mice exposed to hypoxia. This suggested that the metabolic demand would be satisfied despite the respiratory chain depression (cytochrome oxidase), probably due to anaerobic energy production within the mitochondria (succinate dehydrogenase). PMID- 11257426 TI - Regional distribution of ethanolamine plasmalogen in the hippocampal CA1 and CA3 regions and cerebral cortex of the gerbil. AB - Although ethanolamine plasmalogens (EtnPm) are the predominant phospholipids in neural tissue, their physiological role has not been clarified. The biophysical conformation of EtnPm in the proteoliposome enhances the activity of the sodium calcium exchanger, which has been proposed to induce intracellular calcium ion accumulation during ischemia and early reperfusion. The levels of EtnPm in the areas of the gerbil brain selectively vulnerable to ischemia, namely the hippocampal CA1 and CA3 regions and the cerebral cortex, were measured by high performance thin-layer chromatography and gas-liquid chromatography. The concentration of EtnPm in the CA1 region, which is the most vulnerable to ischemic and anoxic stress, was 2.6- and 2.7-fold higher than that in the CA3 region and cerebral cortex, respectively. The significantly higher concentration of EtnPm in the hippocampal CA1 region may enhance sodium-calcium exchanger activity and play an important role in the vulnerability of this region to ischemia. PMID- 11257427 TI - The additivity of the auditory feature analysis in the human brain as indexed by the mismatch negativity: 1+1 approximately 2 but 1+1+1<3. AB - The automatic auditory feature analysis in the human brain was investigated using the mismatch-negativity (MMN) component of the event-related potential. MMNs were recorded in ignore and attend conditions to stimuli deviating from the repetitive standard stimuli simultaneously either in one, two or three features (frequency, intensity, stimulus-onset asynchrony). If the processing of each of these three features is independent of the others, the MMNs to the double and triple deviants should equal to the sum of the MMNs elicited by the corresponding single deviants. The double-deviant MMNs were found to be additive but the triple deviant MMN was clearly underadditive. The results suggest complex interactions between brain processes involved in analyzing several simultaneous deviant features. PMID- 11257428 TI - Induction of cysteine string protein after chronic antidepressant treatment in rat frontal cortex. AB - We have previously identified 204 partial cDNA fragments (ADRG1-204) as antidepressant related genes/expressed sequence tags. Then, we developed our original cDNA microarrays, on which the 194 clones out of ADRG1-204 were spotted. With this ADRG microarray, we found that the expression of a spot, ADRG55, which representing cysteine string protein (CSP), was significantly increased in rat brain after chronic treatment with a selective serotonin reuptake inhibitor, sertraline. In the present study, reverse transcription-polymerase chain reaction analysis confirmed the induction of CSP at mRNA levels in rat frontal cortex after chronic treatment with two different classes of antidepressants, imipramine or sertraline. Western blot analysis also revealed that CSP-immunoreactivity was increased after antidepressant treatment. In conclusion, our data suggest that CSP is one of the common functional molecules induced after chronic antidepressant treatment. PMID- 11257430 TI - Functional modulation of gamma-aminobutyric acid(A) receptors by etifoxine and allopregnanolone in rodents. AB - We looked for an interaction between etifoxine and the neurosteroid allopregnanolone at central gamma-aminobutyric acid (GABA(A)) receptors. Etifoxine (2 microM) did not affect the affinity of allopregnanolone (IC(50)=108 nM) for its site in preparations of Sprague-Dawley rat cerebral cortex membranes, as determined by the inhibition of [(35)S] t-butylbicyclophosphorothionate binding, a specific ligand of the GABA(A) receptor chloride channel site. Etifoxine and allopregnanolone were anticonvulsants, blocking the clonic convulsions induced by bicuculline (an antagonist of the GABA(A) receptor) in CD1 mice. A combination of subactive doses of the two compounds showed additive anticonvulsant effects. These results suggest that etifoxine and allopregnanolone bind to distinct putative recognition sites at or near the chloride channel site. Functionally, their binding may have an additive effect by enhancing GABA(A) inhibitory transmission. PMID- 11257429 TI - A new method for measuring the conduction velocities of Abeta-, Adelta- and C fibers following electric and CO(2) laser stimulation in humans. AB - The conduction velocities of Abeta-, Adelta- and C-fibers of a peripheral nerve of the upper limb in normal subjects were measured by a combination of conventional electric stimulation, painful CO(2) laser stimulation and non painful CO(2) laser stimulation of a tiny skin surface area, respectively. The values obtained were 69.1+/-7.4 m/s, 10.6+/-2.1 and 1.2+/-0.2 m/s, respectively. These findings demonstrated that the combined methods are useful for experimental and clinical exploration of the physiological function and pathophysiological role of Abeta-, Adelta- and C-fibers of a given peripheral nerve. PMID- 11257431 TI - Induction of Fos-like immunoreactivity in the ventral thalamus after electrical or chemical stimulation of various subcortical centres of rats. AB - We have examined the patterns of Fos-like immunoreactivity in the ventral thalamus (thalamic reticular nucleus (Rt), zona incerta (ZI) and ventral lateral geniculate nucleus (LGv)) after electrical or chemical stimulation of nuclei in either the brainstem (midbrain reticular nucleus), basal forebrain (substantia innominata) or dorsal thalamus (parafascicular nucleus). Sprague-Dawley rats were anaesthetised with Halothane or Ketamil/Rompun and the above mentioned centres were stimulated either electrically or chemically (using kainic acid). Brains were then processed for Fos-like immunocytochemistry using standard methods. We detected no major differences in the labelling patterns after either electrical or chemical stimulation or after using Halothane or Ketamil/Rompun anaesthesia. After brainstem or dorsal thalamic stimulations, many Fos-like immunoreactive cells were seen within the rostral pole of the Rt, the dorsal sector of the ZI and the parvocellular lamina of the LGv. After basal forebrain stimulations, many Fos-like immunoreactive cells were seen in the rostral pole of the Rt and rostral sector of the ZI, but very few were apparent in the LGv. Overall, our results show that distinct groups of cells in the ventral thalamus show increased levels of Fos-like immunoreactivity after stimulation of different subcortical centres. These activated cells of the ventral thalamus, are in turn, in a position to influence particular thalamocortical pathways through their dorsal thalamic projections. PMID- 11257432 TI - Elevated plasma amyloid beta-peptide 1-42 and onset of dementia in adults with Down syndrome. AB - We compared levels of plasma amyloid beta-peptides Abeta1-42 and Abeta1-40 in 108 demented and nondemented adults with Down syndrome (DS) and 64 adults from the general population. Abeta1-42 and Abeta1-40 levels were significantly higher in adults with DS than in controls (P=0.0001). Compared to nondemented adults with DS, Abeta1-42 levels in demented adults with DS were selectively increased by 26% (28.2 pg/ml vs. 22.4 pg/ml, P=0.004). In addition, mean plasma levels of Abeta1 42 were 22% higher in DS cases with the apolipoprotein varepsilon4 allele than in DS subjects without an varepsilon4 allele (25.9 pg/ml vs. 21.2 pg/ml, P=0.01), while mean plasma levels of Abeta1-40 did not vary by APOE genotype. These results support the hypothesis that Abeta1-42 plays an important role in the pathogenesis of dementia associated with DS, as it does in Alzheimer's disease, and that variations in plasma levels may be related to disease progression. PMID- 11257433 TI - Fluid-phase uptake and transit in axenic Dictyostelium cells. AB - The main route for fluid-phase uptake in Dictyostelium is macropinocytosis, a process powered by the actin cytoskeleton. Nutrients within the endocytosed fluid are digested and resorbed, disposal of remnants follows by exocytosis. Along the endocytic pathway, membrane fusion and fission events take place at multiple steps. The regulator and effector molecules involved in uptake and transit are largely conserved between higher and lower eukaryotes. This feature, together with its accessibility by molecular genetics, recommend Dictyostelium as a valuable model system for mammalian cells. PMID- 11257434 TI - Regulation of phagocytosis and endo-phagosomal trafficking pathways in Dictyostelium discoideum. AB - Phagocytosis, a critically important process employed by leukocytes against invading pathogens, is an actin-dependent clathrin-independent process that results in the internalization of particles >0.5 microm in diameter. Phagocytosis consists of a number of stages, including the binding of particles to the cell surface via interaction with a receptor, engulfment of the particle by pseudopod extension, and fission and fusion reactions to form phago-lysosomes. Much remains to be learned concerning the molecular mechanisms that regulate particle internalization and phagosome maturation. Dictyostelium is a genetically tractable professional phagocyte that has proven useful in determining the molecular steps involved in these processes. We will summarize, in this chapter, what we currently understand concerning the molecular mechanisms that regulate the process of phagocytosis in Dictyostelium, and we will compare and contrast this body of information with that available describing phagocytosis in higher organisms. We will also present current information that suggests that macropinocytosis, a process morphologically similar to phagocytosis, utilizes a different signaling pathway than phagocytosis. Finally, we will discuss the process of maturation of phagosomes, which requires membrane trafficking events, and we will summarize data that support the use of Dictyostelium as a model to determine how intracellular pathogens survive. PMID- 11257435 TI - The regulation of actin polymerization and cross-linking in Dictyostelium. AB - It is clear that the polymerization and organization of actin filament networks plays a critical role in numerous cellular processes. Inhibition of actin polymerization by pharmacological agents will completely prevent chemotactic motility, macropinocytosis, endocytosis, and phagocytosis. Recently there has been great progress in understanding the mechanisms that control the assembly and structure of the actin cytoskeleton. Members of the Rho family of GTPases have been identified as major players in the signal transduction pathway leading from a cell surface signal to actin polymerization. The Arp2/3 complex has been added to the list of means by which new actin filaments can be nucleated. However, it is clear that actin polymerization by Arp2/3 complex is not the whole story. In principle, the final structures formed by actin filaments will depend on factors such as: the length of actin filaments, the degree of branching, how they are cross-linked and the tensions imparted on them. In addition, the means by which actin polymerization generates protrusion of membranes is still controversial. A phagosome, filopodium and a lamellipodium all require polymerization of new actin filaments, but each has a characteristic morphology and cytoskeletal structure. In the following chapter, we will discuss actin polymerization and filament organization, especially as it relates to the machinery of phagocytosis in Dictyostelium. PMID- 11257436 TI - Dictyostelium discoideum: a genetic model system for the study of professional phagocytes. Profilin, phosphoinositides and the lmp gene family in Dictyostelium. AB - Profilin is a key regulator of actin polymerization, and plays a pivotal role at the interface of the phosphoinositide signal transduction pathway and the cytoskeleton. Recent evidence suggests the involvement of profilin in the regulation of phagocytosis and macropinocytosis, and the transport along the endosomal pathway. Disruption of profilin leads to a complex phenotype that includes abnormal cytokinesis, a block in development and defects in the endosomal pathway. Macropinocytosis, fluid phase efflux and secretion of lysosomal enzymes were reduced, whereas the rate of phagocytosis was increased as compared to wild-type cells. The lmpA gene, a homolog of the CD36/LIMPII family, was identified as a suppressor for most of the profilin-minus defects. This gene encodes an integral membrane protein, it localizes to lysosomes and macropinosomes, and binds to phosphoinositides. Even though phosphatidylinositol lipids constitute only a small fraction of total lipids in the membranes of eukaryotic cells, they play an important role in vesicle transport, signal transduction and cytoskeletal regulation. Disruption of lmpA in wild-type cells resulted in defects in fluid phase efflux and macropinocytosis, but not in phagocytosis. The discovery and initial characterization of two additional members of the CD36/LIMPII family in Dictyostelium, lmpB and lmpC, that exhibit intriguing differences in developmental regulation and their putative sorting signals, suggests that a set of lysosomal integral membrane proteins contribute to the crosstalk between vesicles and cytoskeletal proteins. PMID- 11257438 TI - Regulation of Dictyostelium myosin I and II. AB - Dictyostelium expresses 12 different myosins, including seven single-headed myosins I and one conventional two-headed myosin II. In this review we focus on the signaling pathways that regulate Dictyostelium myosin I and myosin II. Activation of myosin I is catalyzed by a Cdc42/Rac-stimulated myosin I heavy chain kinase that is a member of the p21-activated kinase (PAK) family. Evidence that myosin I is linked to the Arp2/3 complex suggests that pathways that regulate myosin I may also influence actin filament assembly. Myosin II activity is stimulated by a cGMP-activated myosin light chain kinase and inhibited by myosin heavy chain kinases (MHCKs) that block bipolar filament assembly. Known MHCKs include MHCK A and MHCK B, which have a novel type of kinase catalytic domain joined to a WD repeat domain, and MHC-protein kinase C (PKC), which contains both diacylglycerol kinase and PKC-related protein kinase catalytic domains. A Dictyostelium PAK (PAKa) acts indirectly to promote myosin II filament formation, suggesting that the MHCKs may be indirectly regulated by Rac GTPases. PMID- 11257437 TI - Molecular motors and membrane traffic in Dictyostelium. AB - Phagocytosis and membrane traffic in general are largely dependent on the cytoskeleton and their associated molecular motors. The myosin family of motors, especially the unconventional myosins, interact with the actin cortex to facilitate the internalization of external materials during the early steps of phagocytosis. Members of the kinesin and dynein motor families, which mediate transport along microtubules (MTs), facilitate the intracellular processing of the internalized materials and the movement of membrane. Recent studies indicate that some unconventional myosins are also involved in membrane transport, and that the MT- and actin-dependent transport systems might interact with each other. Studies in Dictyostelium have led to the discovery of many motors involved in critical steps of phagocytosis and membrane transport. With the ease of genetic and biochemical approaches, the established functional analysis to test phagocytosis and vesicle transport, and the effort of the Dictyostelium cDNA and Genome Projects, Dictyostelium will continue to be a superb model system to study phagocytosis in particular and cytoskeleton and motors in general. PMID- 11257439 TI - Dictyostelium: an ideal organism for genetic dissection of Ras signalling networks. AB - Signalling pathways based on the small GTPase Ras regulate a multitude of cellular events in eukaryotic cells. Dictyostelium expresses a large and varied family of Ras proteins. It also uses a range of known Ras regulators, in particular RasGEFs, and effectors. The genetic tractability of Dictyostelium, together with the wide range of Ras proteins and regulators, make it an ideal model for the genetic dissection of Ras pathways. This review highlights the recent advances in our understanding of Ras function in Dictyostelium, and considers the implications of these findings for our understanding of eukaryotic signal transduction. PMID- 11257441 TI - Sortilin/neurotensin receptor-3: a new tool to investigate neurotensin signaling and cellular trafficking? AB - The identification of gp95sortilin, a sorting protein, as being the 100 kDa neurotensin (NT) receptor, a non-G-protein coupled receptor, constitutes a new and interesting but intriguing step in the neuropeptide signaling as well as in cellular trafficking. The isolation of the same protein by three different experimental approaches sum up the complexity for researchers involved in the functional significance of the so-called sortilin/neurotensin receptor 3 (NTR3). This review will concentrate on the putative physiological and cellular roles of sortilin/NTR3 as most results so far have proposed hypothetical conclusions rather than concrete evidence. PMID- 11257442 TI - Protein phosphatase 2A: the Trojan Horse of cellular signaling. AB - Dynamic phosphorylation and dephosphorylation of proteins are fundamental mechanisms utilized by cells to transduce signals. Whereas transduction by protein kinases has been a major focus of studies in the last decade, protein phosphatase 2A (PP2A) enzymes emerge in this millenium as the most fashionable players in cellular signaling. Viral proteins target specific PP2A enzymes in order to deregulate chosen cellular pathways in the host and promote viral progeny. The observation that a variety of viruses utilize PP2A to alienate cellular behavior emphasizes the fundamental importance of PP2A in signal transduction. This review will primarily focus on discussing the uniqueness of PP2A regulation and uncovering the critical role played by protein-protein interactions in the modulation of PP2A signaling. Moreover, the place of PP2A in signaling pathways and its functional significance for human diseases will be discussed. PMID- 11257443 TI - U73122 inhibits the dephosphorylation and translocation of cofilin in activated macrophage-like U937 cells. AB - Cofilin, an actin-binding protein, plays an important role in the migration, phagocytosis, and superoxide production of activated phagocytes through cytoskeletal reorganization. In unstimulated phagocytes, cofilin is a major phosphoprotein. However, upon activation, the phosphoprotein is dephosphorylated and translocated from cytosol to plasma membranes. Only the unphosphorylated form of cofilin is an active form that binds actin, whereas the regulatory mechanisms of cofilin have not been elucidated. We found that 1-[6-[[17beta-3-methoxyestra 1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione (U73122), an inhibitor of phospholipase C (PLC), suppressed both opsonized zymosan (OZ)-induced dephosphorylation and translocation of cofilin in macrophage-like U937 cells at 4 microM concentration. OZ triggered an increase in inositol 1,4,5-trisphosphate (IP3), and U73122 inhibited it. 1-[6-[[17beta-3-Methoxyestra-1,3,5(10)-trien-17 yl]amino]hexyl]-1H-pyrrole-2,5-pyrrodione-dione (U73343), which was employed as an inactive analogue, had no such inhibitory activities as did U73122. Furthermore, herbimycin A, an inhibitor of src-type tyrosine kinase, also inhibited OZ-triggered IP3 formation. These results suggest that the activity and localization of cofilin are regulated by PLC at the downstream of src-family tyrosine kinase. PMID- 11257444 TI - Hypoxia activates Akt and induces phosphorylation of GSK-3 in PC12 cells. AB - Akt is a serine/threonine kinase that has been shown to play a central role in promoting cell survival and opposing apoptosis. We evaluated the effect of hypoxia on Akt in rat pheochromocytoma (PC12) cells. PC12 cells were exposed to varying levels of hypoxia, including 21%, 15%, 10%, 5%, and 1% O(2). Hypoxia dramatically increased phosphorylation of Akt (Ser(473)). This effect peaked after 6 h exposure to hypoxia, but persisted strongly for up to 24 h. Phosphorylation of Akt was paralleled with a progressive increase in phosphorylation of glycogen synthase kinase-3 (GSK-3), one of its downstream substrates. The effect of hypoxia on phosphorylation of Akt was completely blocked by pretreatment of the cells with wortmannin (100 nM), indicating that this effect is mediated by phosphatidylinositol 3-kinase (P13K). In contrast, whereas hypoxia also strongly induced phosphorylation of the transcription factors CREB and EPAS1, these effects persisted in the presence of wortmannin. Thus, hypoxia regulates both P13K-dependent and P13K-independent signaling pathways. Furthermore, activation of the P13K and Akt signaling pathways may be one mechanism by which cells adapt and survive under conditions of hypoxia. PMID- 11257445 TI - Inactivation of JNK activity by mitogen-activated protein kinase phosphatase-2 in EAhy926 endothelial cells is dependent upon agonist-specific JNK translocation to the nucleus. AB - We have investigated the termination of agonist-stimulated mitogen-activated protein (MAP) kinase activity in EAhy926 cells by MAP kinase phosphatase-2 (MKP 2). In cells expressing either wild-type (WT) or catalytically inactive (CI)-MKP 2, there was no significant differences in TNFalpha-stimulated JNK or p38 MAP kinase activity, however hydrogen peroxide (H2O2)-stimulated JNK activity was substantially reduced in WT-MKP-2 expressing clones and enhanced in cells expressing CI-MKP-2. Consistent with these findings, we observed substantial nuclear translocation of JNK occurred in response to H2O2 but not TNFalpha. Using a phosphospecific anti-JNK antibody, we found that TNFalpha-stimulated JNK activity was associated principally with the cytosol while in response to H2O2, JNK activity was found within the nucleus. These results show that the role of MKP-2 in terminating JNK activity is determined by the translocation of JNK to the nucleus, which is under agonist-specific regulation and not a universal cellular response to stimulation. PMID- 11257446 TI - Pancreastatin, a chromogranin A-derived peptide, activates Galpha(16) and phospholipase C-beta(2) by interacting with specific receptors in rat heart membranes. AB - Pancreastatin (PST) is one of the chromogranin A (CGA)-derived peptides with known biological activity. It has a general inhibitory effect on secretion in many exocrine and endocrine systems including the heart atrium. Besides, a role of PST as a counter-regulatory peptide of insulin action has been proposed in the light of its effects on glucose and lipid metabolism in the liver and adipose tissue, where receptors and signaling have been described. Galpha(q/11) pathway seems to mediate PST action. Since PST has been shown to function as a typical calcium-dependent hormone, and increased plasma levels have been found in essential hypertension correlating with catecholamines, we sought to study its possible interaction and signaling in heart membranes. Here, we are characterizing specific PST binding sites and signaling in rat heart membranes. We have found that PST receptor has a K(d) of 0.5 nM and a B(max) of 34 fmol/mg of protein. The PST binding is inhibited by guanine nucleotides, suggesting the functional coupling of the receptor with GTP binding proteins (G proteins). Moreover, PST dose-dependently increases GTP binding to rat heart membranes. Finally, we have studied PST signaling-effector system by measuring phospholipase C (PLC) activity using blocking antibodies against different G proteins and PLC isoforms. We have found that PST stimulates PLCbeta(2)>PLCbeta(1)>PLCbeta(3) by activating Galpha(16) in rat heart membranes. These data suggest that PST may modulate the cardiac function. PMID- 11257447 TI - Grb2 promotes reinitiation of meiosis in Xenopus oocytes. AB - The adaptor protein Grb2 plays a central role in cell proliferation and/or cell cycle progression. In this study, we investigate the role of Grb2 in signalling pathways involved in meiotic reinitiation. For that purpose, Xenopus Grb2 cRNA and its mutated forms or human Grb2 protein was microinjected into immature Xenopus oocytes. Reinitiation of meiosis was seen in unstimulated oocytes. Induction of the meiosis was time dependent and Ras dependent, and the presence in Grb2 of SH2 and SH3 domains was required. Several tyrosine phosphorylated proteins were solely detected in oocytes responsive to Grb2 injection. Our results are in favour of an unusual recruitment and initiation of the Grb2 transduction cascade independent of a receptor tyrosine kinase (RTK) stimulation. PMID- 11257448 TI - Inhibition of the type 1 inositol 1,4,5-trisphosphate-sensitive Ca2+ channel by calmodulin antagonists. AB - This study describes the effects of a number of calmodulin antagonists on the cerebellar type 1 inositol 1,4,5-trisphosphate (InsP3) receptor. All the antagonists tested (trifluoperazine, fluphenazine, chlorpromazine and calmidazolium) inhibited the extent of InsP3-induced Ca2+ release (IICR) with similar IC(50) values (between 60 and 85 microM). They did not affect the efficacy of InsP3 to release Ca2+, since the concentrations of InsP3 required to cause half-maximal release was little affected in the presence of these agents. In addition, these agents did not affect InsP3 binding to its receptor. Stopped flow studies to determine the rate constants of IICR showed this process to be biphasic with a fast and slow component. All the calmodulin antagonists appeared to reduce the rate constants for Ca2+ release in a phase-specific manner, preferentially reducing the fast phase component. Chlorpromazine (75 microM) appeared to have the most potent effect on the fast phase rate constant, reducing it from 1.0 to 0.08 s(-1), while only reducing the rate constant for the slow phase about twofold (0.2-0.08 s(-1)). The fact that calmodulin itself inhibits both IICR and InsP3 binding, while these calmodulin antagonists also reduce Ca2+ release and do not affect InsP3 binding, suggests that the mechanism of action of these agents is unlikely to be due to the reversal of the modulatory action of calmodulin on this receptor. PMID- 11257449 TI - Regulation of Pyk2 expression by p56(Lck) in Jurkat T lymphocytes. AB - Lysates from the Jurkat T lymphocyte cell line were immunoblotted with anti-Pyk2, and two major forms of Pyk2 were identified. When lysates from the p56(Lck) negative (J.CaM1/Rep3) and CD45 negative Jurkat cell line derivatives were immunoblotted with anti-Pyk2, only the lower mobility form of Pyk2 was predominant. Transfection of J.CaM1 cells with p56(Lck) restored expression of the multiple forms of Pyk2. Using RT-PCR, we found that both species of the alternatively spliced mRNA for Pyk2 were present in all of the lines regardless of their ability to express CD45 or p56(Lck) protein. When p56(Lck) immunoprecipitates were immunoblotted with anti-Pyk2, only the higher mobility form of Pyk2 immunoprecipitated with p56(Lck). These data demonstrate that certain members of the Src family of kinases interact preferentially with the different isoforms of Pyk2 and may have a role in the regulation of the Pyk2 protein in lymphocytes. PMID- 11257451 TI - Receptor activity modifying proteins. AB - Our understanding of G protein-coupled receptor (GPCR) function has recently expanded to encompass novel protein interactions that underlie both cell-surface receptor expression and the exhibited phenotype. The most notable examples are those involving receptor activity modifying proteins (RAMPs). RAMP association with the calcitonin (CT) receptor-like receptor (CRLR) traffics this receptor to the cell surface where individual RAMPs dictate the expression of unique phenotypes. A similar function has been ascribed to RAMP interaction with the CT receptor (CTR) gene product. This review examines our current state of knowledge of the mechanisms underlying RAMP function. PMID- 11257452 TI - LPS induction of gene expression in human monocytes. AB - Lipopolysaccharide (LPS [endotoxin]) is the principal component of the outer membrane of Gram-negative bacteria. Recent studies have elucidated how LPS is recognized by monocytes and macrophages of the innate immune system. Human monocytes are exquisitely sensitive to LPS and respond by expressing many inflammatory cytokines. LPS binds to LPS-binding protein (LBP) in plasma and is delivered to the cell surface receptor CD14. Next, LPS is transferred to the transmembrane signaling receptor toll-like receptor 4 (TLR4) and its accessory protein MD2. LPS stimulation of human monocytes activates several intracellular signaling pathways that include the IkappaB kinase (IKK)-NF-kappaB pathway and three mitogen-activated protein kinase (MAPK) pathways: extracellular signal regulated kinases (ERK) 1 and 2, c-Jun N-terminal kinase (JNK) and p38. These signaling pathways in turn activate a variety of transcription factors that include NF-kappaB (p50/p65) and AP-1 (c-Fos/c-Jun), which coordinate the induction of many genes encoding inflammatory mediators. PMID- 11257453 TI - Activation of Ras, Raf-1 and protein kinase C in differentiating human neuroblastoma cells after treatment with phorbolester and NGF. AB - The human neuroblastoma cell line SH-SY5Y/TrkA differentiates in vitro and acquires a sympathetic phenotype in response to phorbolester (activator of protein kinase C, PKC) in the presence of serum or growth factors, or nerve growth factor (NGF). We have now investigated to what extent phorbolester and NGF cause activation of Ras and Raf-1 and the involvement of PKC in this response in differentiating SH-SY5Y/TrkA cells. NGF stimulated increased accumulation of Ras GTP and a threefold activation of Raf-1. In contrast, 12-O-tetradecanoylphorbol 13-acetate (TPA) had no effect on the amount of Ras-GTP but led to a smaller activation of Raf-1. NGF caused a limited increase in phosphorylation of Raf-1 compared with TPA, and NGF-induced Raf activity was independent of PKC. Analysis of phosphorylation of the endogenous PKC substrate myristoylated alanine-rich C kinase substrate (MARCKS), and of subcellular distribution of PKC-alpha, -delta, and -epsilon revealed that NGF only caused a very small activation of PKC in SH SY5Y/TrkA cells. The results identify Raf-1 as a target for both TPA- and NGF induced signals in differentiating SH-SY5Y/TrkA cells and demonstrate that signalling to Raf-1 was mediated via distinct mechanisms. PMID- 11257454 TI - Prostaglandin F(2)alpha enhances glucose consumption through neither adipocyte differentiation nor GLUT1 expression in 3T3-L1 cells. AB - Arachidonic acid (AA) at 0.2 mM enhances glucose uptake through increased levels of glucose transporter (GLUT) 1 protein in 3T3-L1 adipocytes. Since AA is a precursor of prostaglandins (PGs), we investigated the effect of PGs on glucose consumption in 3T3-L1 cells. Among several PGs, only prostaglandin F(2)alpha (PGF(2)alpha) enhanced glucose consumption in 3T3-L1 cells treated with dexamethasone (DEX), 3-isobutyl-1-methyl-xanthine (IBMX), and insulin. To study the mechanism of PGF(2)alpha-enhanced glucose consumption, we investigated the effect of PGF(2)alpha on glycerol-3-phosphate dehydrogenase (GPDH) activity, triglycerides (TGs) content, and the expression of GLUT1 protein. PGF(2)alpha suppressed GPDH activity and did not increase the expression of GLUT1 protein in 3T3-L1 cells treated with DEX, IBMX, and insulin. These results suggest that AA stimulated glucose uptake is not through the effect of PGF(2)alpha. Our results indicate that PGF(2)alpha is a unique regulator of adipocyte differentiation (suppression) and glucose consumption (enhancement) in 3T3-L1 cells. PMID- 11257455 TI - Redistribution of cPLA(2) in rat renal tubular cell cultures in response to PCBs. AB - The influence of different polychlorinated biphenyls (PCBs) upon the cytosolic phospholipase A(2) (cPLA(2)) redistribution to the particulate fraction has been investigated in rat renal proximal tubule culture cells. Treatment with Aroclor 1248 increased PLA(2) activity in the particulate fraction in a concentration dependent manner using two radioactive substrates. However, the activity of PLA(2) in the cytosolic fraction decreased. This work also shows that 2,2',4,4',5,5'-hexachlorobiphenyl (HCB) (a di-ortho-substituted nonplanar congener) can increase the activity of PLA(2) in the particulate fraction and decrease the enzyme activity in the cytosolic fraction. The exposure of cell cultures to 3,3',4,4'-tetrachlorobiphenyl (TCB) (a non-ortho-subtituted planar congener) does not alter PLA(2) activity. These results suggest that PCBs, depending on their planar or nonplanar structures, cause a translocation of the enzyme from the cytosol to membranes. To evaluate this possibility, the contents of immunoreactive cPLA(2) were examined by immunoblot analysis in the high-speed supernatant and the particulate fraction of treated cell cultures. The increases/decreases in the amounts of cPLA(2) protein agree with the increases/decreases of PLA(2) activity previously cited. These data demonstrate that the PCB-stimulated redistribution of cPLA(2) to membranes is associated, at least in part, with the changes detected in the activity of the enzyme. PMID- 11257457 TI - Overexpression of EXTL3/EXTR1 enhances NF-kappaB activity induced by TNF-alpha. AB - EXTL3/EXTR1 is a member of the EXT gene family, which may represent a class of glycosyltransferases involved in heparan sulfate biosynthesis. It is known that heparan sulfate interacts with a variety of proteins and is therefore implicated in various cellular responses. Here, we examined the effect of EXTL3 on nuclear factor-kappaB (NF-kappaB) activity stimulated by tumor necrosis factor-alpha (TNF alpha), one of heparin-binding cytokine. The luciferase assay demonstrated that overexpression of EXTL3 enhanced TNF-alpha-induced NF-kappaB activity. This is confirmed with an electrophoretic mobility shift assay. However, EXTL3 did not affect the CD40-mediated NF-kappaB activation. The EXTL3 mutants lacking the amino terminus region failed to enhance the activity. The fluorescence of enhanced green fluorescent protein (EGFP)-fused EXTL3 was observed at the perinuclear region, whereas, the amino terminus-truncated mutant was found in a diffuse cytoplasmic region. These results suggest that EXTL3 may modulate NF kappaB mediated by TNF-alpha. PMID- 11257456 TI - C1q arrests the cell cycle progression of fibroblasts in G(1) phase: role of the cAMP/PKA-I pathway. AB - C1q may participate in the loss of connective tissue occurring in chronic inflammatory lesions. The hypothesis of a detrimental role of C1q on cell proliferation was tested on primary cultures of human fibroblasts (HFs). C1q suppressed the DNA synthesis of HF in response to platelet-derived growth factor (PDGF) with an IC(50) of 20 microg/ml, and blocked 78% of the cycling cells in G(1) phase. The C1q block did not involve production of inhibitory prostaglandin by the cells. Given that C1q elicits signals of the adenylyl cyclase pathway in HF, we examined cAMP-dependent mechanisms to understand how C1q inhibited the PDGF response. Whereas the C1q block was enhanced by agonist dibutyryl-adenosine 3', 5'-cyclic mono-phosphate (db-cAMP), antagonist adenosine 3', 5'-cyclic monophosphorotioate triethylammonium salt (Rp-cAMP) minimized it. C1q increased the level of cAMP-dependent protein kinase I (PKA-I) 4.5-fold, without altering the activation of the extracellular-regulated protein kinase (ERK) pathway. These results demonstrate that the interactions of C1q with HF cause growth arrest at the G(1) phase through mechanisms associated with a PKA-I dependent pathway. PMID- 11257458 TI - PTH stimulates PLCbeta and PLCgamma isoenzymes in rat enterocytes: influence of ageing. AB - We previously reported that in rat duodenal cells (enterocytes), parathyroid hormone (PTH [1-34]: PTH) stimulates the hydrolysis of polyphosphoinositides by phospholipase C (PLC), generating the second messengers inositol trisphosphate (IP(3)) and diacylglycerol (DAG) and that this mechanism is severely altered in old animals. In the present study, we show that PTH [1-34]-dependent IP(3) release in young rats was blocked to a great extent by an antibody against guanine nucleotide binding protein Galphaq/11, indicating that the hormone activates a beta isoform of PLC coupled to the alpha subunit of Gq/11. In addition, PTH rapidly (within 30 s, with maximal effects at 1 min) stimulated tyrosine phosphorylation of PLCgamma in a dose-dependent fashion (10(-10)-10(-7) M). The hormone response was specific as PTH [7-34] was without effects. The tyrosine kinase inhibitors, genistein (100 microM) and herbimycin (2 microM), suppressed PTH-dependent PLCgamma tyrosine phosphorylation. Stimulation of PLCgamma tyrosine phosphorylation by PTH [1-34] greatly decreased with ageing. PP1 (10 microM), a specific inhibitor of the Src family of tyrosine kinases, completely abolished PLCgamma phosphorylation. The hormone-induced Src tyrosine dephosphorylation, a major mechanism of Src activation, an effect that was blunted in old animals. These results indicate that in rat enterocytes PTH generates IP(3) mainly through G-protein-coupled PLCbeta and stimulates PLCgamma phosphorylation via the nonreceptor tyrosine kinase Src. Impairment of PTH activation of both PLC isoforms upon ageing may result in abnormal hormone regulation of cell Ca(2+) and proliferation in the duodenum. PMID- 11257460 TI - Studies on the effect of diadenlyated nucleotides on calcium mobilization and prostacyclin synthesis in bovine aortic endothelial cells. AB - Extracellular adenine dinucleotides are modulators of blood vessel tone. We have previously demonstrated that Ap(2)A and Ap(4)A induce the synthesis of nitric oxide (NO) from bovine aortic endothelial cells (BAEC) while Ap(3)A and Ap(5)A do not [FEBS Lett. 427 (1998) 320; Arch. Biochem. Biophys. 364 (1999) 280.]. In this communication we determine the effect of Ap(x)As (x=2-5) on prostacyclin (PGI(2)) synthesis and Ca(2+) mobilization in BAEC. Ap(2)A and Ap(4)A significantly enhanced the synthesis of PGI(2) while Ap(3)A and Ap(5)A do not. These data support the notion that Ap(2)A and Ap(4)A are vasodilators. All four dinucleotides significantly enhanced Ca(2+) mobilization over basal levels. Ap(5)A and Ap(3)A enhanced 2.0 and 1.6 times more Ca(2+) release than Ap(4)A, respectively. Since neither Ap(5)A nor Ap(3)A enhanced the synthesis of either PGI(2) or NO but did mobilize Ca(2+), these data support the hypothesis that in BAEC Ca(2+) release is localized or compartmentalized. PMID- 11257459 TI - EGF receptor crosstalks with cytokine receptors leading to the activation of c Jun kinase in response to UV irradiation in human keratinocytes. AB - Ultraviolet (UV) irradiation causes photoageing through induction of matrix degrading metalloproteinases (MMP), which are upregulated by activator protein-1 (AP-1) (Jun/Fos). The c-Jun kinase activity proves to be critically important in the regulation of AP-1 activity. Our previous studies showed that UV irradiation activates epidermal growth factor receptor (EGFR) and cytokine receptors leading to the activation of c-Jun kinase in cultured human skin keratinocytes in vitro and in human skin in vivo. However, the mechanism of UV-induced cell surface receptor activation and the crosstalk among growth factor receptor and cytokine receptors were not fully investigated. This study showed that UV (30 mJ/cm(2)) induced EGFR tyrosine phosphorylation in a manner similar to EGF (100 ng/ml), or IL-1beta (10 ng/ml) in cultured human keratinocytes. In all cases, EGFR tyrosine phosphorylation was completely inhibited by pretreatment of PD153035 (100 nM, 1 h). Also observed was that UV induced autophosphorylation of interleukin 1 receptor associated kinase (IRAK) in a manner analogous to IL-1beta or EGF. In both UV and EGF cases, the phosphorylation of IRAK was inhibited by pretreatment of PD153035. However, IL-1beta-induced IRAK activation was not affected by PD153035. In vitro kinase assay using GST-c-Jun as a substrate revealed that pretreatment of PD153035 completely inhibited UV- and IL-1-induced c-Jun kinase activity in cultured keratinocytes. Taken together, the above data suggest that EGFR plays dominant role in the crosstalk among growth factor receptor and cytokine receptors leading to the activation of c-Jun kinase upon UV irradiation, and that EGFR could be one of the targets for clinical and cosmetical prevention of UV-induced skin aging. PMID- 11257461 TI - Changes in UCP mRNA expression levels in brown adipose tissue and skeletal muscle after feeding a high-energy diet and relationships with leptin, glucose and PPARgamma. AB - Brown adipose tissue and skeletal muscle are known to be important sites for nonshivering thermogenesis. In this context, it is accepted that uncoupling proteins (UCPs) are involved in such process, but little is known about the physiological regulation of these proteins as affected by the intake of a high energy (cafeteria) diet inducing fat deposition. In this study, the UCP messenger RNA (mRNA) expression in interscapular brown adipose tissue (iBAT) and skeletal muscle was assessed to evaluate the influence of a dietary manipulation on energy homeostasis regulation. We report a statistically significant increase in mRNA levels of iBAT UCP1 and UCP3 and a statistical marginal rise in skeletal muscle UCP3 mRNA expression after feeding a high-energy diet, whereas no changes in UCP2 expression were found in either tissue. Furthermore, significant positive associations between iBAT UCP1 and UCP3 mRNA levels with serum leptin were found. Although the expression of the beta(3) adrenoceptor (beta(3)AR) was about 50% in the lean controls compared with the obese group in iBAT, no statistically significant changes were observed concerning peroxisome proliferator-activated receptor gamma2 (PPARgamma2) mRNA levels in muscle or iBAT. We conclude that feeding a diet inducing weight and fat gain produces different outcomes on iBAT and skeletal muscle UCP mRNA expression, revealing a tissue-dependent response for the three UCPs. Results suggest that the regulation of UCP expression in both tissues under these specific dietary conditions may be related to leptin circulating levels. PMID- 11257462 TI - Oxidative status of plasma and muscle in rabbits supplemented with dietary vitamin E. AB - Thirty New Zealand white rabbits, mean weight 2 kg, were divided into three equal groups balanced for body weight and randomly assigned to a diet containing 60 (C), 150 (T1) or 375 (T2) mg/kg of all-rac-alpha-tocopheryl acetate. After 29 days, the animals were slaughtered. alpha-Tocopherol was assayed in muscle (longissimus dorsi) and plasma; triglycerides and cholesterol (total, high density lipoprotein, low density lipoprotein) were analysed in plasma; reactive oxygen metabolites (ROMs) were analysed in serum; and thiobarbituric acid reactive substances (TBARS) were analysed in muscle. There were no body weight and food intake differences between the groups. The plasma vitamin E and vitamin E:lipid ratio were significantly higher in groups T1 and T2 than in C, but increases were not linearly related to dietary levels. Muscle alpha-tocopherol concentrations in the treated groups were significantly higher than in C, and linearly related (R =.67) to the vitamin E:lipid ratio. ROM and vitamin E levels in blood were inversely related (R =.74), with ROMs significantly lower in the treated groups than in C. The 60-mg/kg dose of C recommended by the National Research Council was unable to control ROM production. Lipid oxidation in muscle was significantly lower in T2 than in the other groups, and TBARS correlated significantly with muscle vitamin E (R =.61) and serum ROM (R =.73). These data suggest that vitamin E supplemented at 375 mg/kg diet can effectively control ROM production and improve muscle lipostability. ROM assay provides a useful indirect estimate of the oxidative status of muscle in vivo. PMID- 11257463 TI - Characterization of antioxidants present in hawthorn fruits. AB - Hawthorn fruit extract has been shown to have many health benefits including being cardiovascular protective, hypotensive and hypocholesterolemic. The present study was carried out to characterize further the antioxidants of hawthorn fruit and their effect on the oxidation of human low density lipoprotein (LDL) and alpha-tocopherol. The dry hawthorn fruit was extracted successively with ether, ethyl acetate, butanol and water. The ethyl acetate fraction was only effective in inhibition of Cu(+2)-mediated LDL oxidation. The column chromatographic separation led to isolation of eight pure compounds; namely, ursolic acid, hyperoside, isoquercitrin, epicatechin, chlorogenic acid, quercetin, rutin and protocatechuic acid. All of these phenolic compounds, except ursolic acid, were protective to human LDL from Cu(+2)-mediated LDL oxidation. They were also effective in preventing the peroxy free radical-induced oxidation of alpha tocopherol in human LDL. The inhibitory effect of these compounds on oxidation of LDL and alpha-tocopherol was dose-dependent at concentrations ranging from 5 to 40 uM. In addition, supplementation of 2% hawthorn fruit powder significantly elevated serum alpha-tocopherol by 18-20% in rats fed a 30% polyunsaturated canola oil diet, as compared with the control. The present results suggest that part of the mechanism for cardiovascular protective effects of hawthorn fruit might also involve the direct protection to human LDL from oxidation or indirect protection via maintaining the concentration of alpha-tocopherol in human LDL. PMID- 11257465 TI - Fibrinogen is an efficient antioxidant. AB - Fibrinogen has been included among the risk factors for vascular disease. Fibrinogen belongs with albumin, ceruloplasmin and transferrin to an acute phase protein group in the plasma. Albumin, ceruloplasmin and transferrin are already recognized as natural antioxidants. In the present study we used three different oxygen generating systems in order to test whether fibrinogen is able to act as an antioxidant in an in vitro system. We used 1) pyrogallol auto-oxidation, 2) the reaction catalysed by xanthine oxidase coupled with the reduction of ferricytochrome c and 3) chemiluminescence. We found that in a dose-dependent manner fibrinogen inhibited superoxide generation (pyrogallol and xanthine xanthine oxidase reactions), ferrous ion oxidation and hydroxyl radical dependent degradation (of deoxyribose). Fibrinogen also inhibited LDL oxidation (copper and azo compound-induced), hydrogen peroxide oxidation and chemiluminescence produced by polymorphonuclear leukocytes. Fibrinogen, albumin, ceruloplasmin and transferrin act as a supplementary antioxidant defense mechanism against oxidative stress arising from inflammatory conditions. PMID- 11257464 TI - Lipolytic effect of BRL 35 135, a beta3 agonist, and its interaction with dietary lipids on the accumulation of fats in rat body. AB - The type of intaked fat and fat uptake mechanisms such as adrenergic-induced lipolysis affect patterns of fat accumulation in animal body. In this study, in vitro lipolytic effect of BRL 35135, a selectivebeta3 agonist, and its interaction with different dietary fats on fat accumulation in animal body (in vivo) were studied. For in vitro study, adipocytes isolated from epididymal fat were incubated with 10(-5) M -10(-9) M of either BRL 35135 or isoproterenol, a non-selectivebeta-agonist. In animal study, two groups of SD-rats, i.e., BRL35135 intaked (dosed at 0.5 mg/kg/day in diet) and control, were divided into 4 sub groups and fed diets containing 12% of either beef tallow (BT), canola oil (CO), olive oil (OO) or safflower oil(SO) for 6 weeks. In vitro study showed that BRL 35135 was 10 times more potent than isoproterenol in increasing the lipolysis in rat adipocytes. In animal study, inclusion of BRL35135 reduced daily weight gain in CO and SO groups (P < 0.05). Abdominal fat weight in BRL35135-intaked group was significantly lower than control in all dietary sub-groups (CO, OO and SO) except BT (P < 0.05). In BT group, abdominal fat contained significantly higher amount of total saturated fatty acids (SFAs) compared to CO, OO or SO. It was concluded that, although BRL 35135 was very potent in increasing lipolysis in the isolated adipocytes of rat, its preventive effect on lipid accumulation in animal body through the lipolysis could be affected by the type of dietary fat and was lesser when rats fed fats rich in SFAs. PMID- 11257466 TI - Iron status and the risk of coronary heart disease: an example of the use of nutritional epidemiology in chronic disease research. PMID- 11257467 TI - Different effects of conjugated linoleic acid isomers on lipoprotein lipase activity in 3T3-L1 adipocytes. AB - Conjugated linoleic acids (CLAs) are the positional and geometric isomers of linoleic acid. In the present study the effects of cis-9, trans-11 CLA (c9,t11 CLA) and trans-10, cis-12 CLA (t10,c12 CLA ) on intracellular and heparin releasable (HR-) lipoprotein lipase (LPL) activity in 3T3-L1 adipocytes were investigated. Cells were exposed to the two CLA isomers and linoleic acid, which were bound to bovine serum albumin (BSA). In the adipocytes insulin up-regulated and tumor necrosis factor alpha (TNFalpha) down-regulated HR-LPL activity, which corresponds with the findings in vivo. The experimental fatty acids at low concentrations (<30 umol/L) moderately increased intracellular and HR-LPL activity. At a concentration of 100 umol/L, c9,t11 CLA and t10,c12 CLA suppressed HR-LPL activity to 20 and 24% below the BSA control level, respectively, while linoleic acid had no effect unless its concentration was as high as 1000 umol/L. Insulin abolished the inhibitory effect of c9,t11 CLA, but not of t10,c12 CLA. In the presence of insulin, t10,c12 CLA inhibited HR-LPL activity by 41% compared to BSA control. In contrast to TNFalpha, which suppressed both intracellular LPL and HR-LPL activity, CLAs suppressed HR-LPL activity without decreasing intracellular LPL activity. Additionally, t10,c12 CLA (100 umol/L) partially prevented TNFalpha induced decrease of intracellular LPL activity. These results indicate that CLAs differ from linoleic acid in regulating HR-LPL activity, and t10,c12 CLA appeared to be more effective than c9,t11 CLA. PMID- 11257468 TI - Novel dystrophin mutations revealed by analysis of dystrophin mRNA: alternative splicing suppresses the phenotypic effect of a nonsense mutation. AB - The complete dystrophin mRNA sequence has been analyzed in 20 Duchenne muscular dystrophy and Becker muscular dystrophy patients. In 13 cases, deletions in mRNA were detected using reverse transcription-polymerase chain reaction and in another seven cases, point mutations were found using the protein truncation test. Sixteen patients diagnosed with Duchenne muscular dystrophy showed the presence of deletions or of nonsense point mutations. From four patients with the Becker muscular dystrophy phenotype, three cases were associated with deletions conserving the translational frame and one was associated with a nonsense mutation E1110X. In the case of the E1110X mutation, an alternative splicing of dystrophin mRNA (3485-3640del) was detected in this patient which included the E1110X mutation site (nucleotide 3536) and did not change the translation reading frame. Individual nonsense point mutations were characterized by sequence analysis, which showed five novel mutations with respect to those reported in the Cardiff Human Gene Mutation Database http://uwcm.web.cf.ac.uk/uwcm/mg/hgmd0.html and the Leiden muscular dystrophy pages http://www.dmd.nl/. PMID- 11257469 TI - A new dysferlin gene mutation in two Japanese families with limb-girdle muscular dystrophy 2B and Miyoshi myopathy. AB - We found a new dysferlin gene mutation in two Japanese families, one with limb girdle muscular dystrophy 2B and the other with Miyoshi myopathy. All patients in the limb-girdle muscular dystrophy 2B family showed apparent proximal dominant muscle atrophy and weakness, whereas a patient with Miyoshi myopathy in the second family showed distal muscle involvement at an early stage. The common clinical feature of all patients in both families was preferential involvement of calf muscles rather than the tibialis anterior muscle, which was confirmed by muscle computed tomography scan. All patients in both families shared the same homozygous alleles for chromosome 2p13 markers, and dysferlin gene analysis revealed a novel missense mutation, a G to A transition at nt 5882, which changed aspartic acid to asparagine at codon 1837. Allele-specific polymerase chain reaction analysis was used for confirmation of the mutation and for genotype analysis of the family members. PMID- 11257470 TI - Abnormalities in the expression of nebulin in chromosome-2 linked nemaline myopathy. AB - Nemaline myopathy is clinically and genetically heterogeneous. The most common autosomal recessive form affecting infants (NEM2) links to chromosome 2q, and is caused by mutations in the gene for nebulin. We have examined the immunocytochemical expression of nebulin in skeletal muscle in 11 cases of nemaline myopathy, from ten families, with linkage compatible to chromosome 2q.22, the locus for nebulin. Mutations in the gene for nebulin have been found in eight of these cases. Immunolabelling with polyclonal antibodies to C-terminal regions of nebulin was compared with antibodies to fibre-type-specific myofibrillar proteins, including myosin heavy chain isoforms and alpha-actinin isoforms. No cases showed a complete absence of C-terminal nebulin, and no enhancement of labelling of the rods was seen with conventional fluorescence microscopy. In control muscle an antibody to the M176-181 repeat region of nebulin showed higher expression in fibres with slow myosin, while ones to the serine-rich domain and to the SH3 domain showed uniform expression. In some cases of nemaline myopathy differences in these patterns were observed. Two siblings with a homozygous mutation in exon 185, that produces a stop codon, showed an absence of labelling only with the SH3 antibody, and other cases showed uneven labelling with this antibody or some fibres devoid of label. Fibre type correlations also showed differences from controls, as some fibres had a fast isoform of one protein but a slow isoform of another. These results indicate that analysis of nebulin expression may detect abnormalities in some cases linked to the corresponding locus and may help to direct molecular analysis. In addition, they may also be relevant to studies of fibre type plasticity and diversity in nemaline myopathy. PMID- 11257471 TI - Nebulin expression in patients with nemaline myopathy. AB - Nemaline myopathy is a structural congenital myopathy which may show both autosomal dominant and autosomal recessive inheritance patterns. Mutations in three different genes have been identified as the cause of nemaline myopathy: the gene for slow alpha-tropomyosin 3 (TPM3) at 1q22-23, the nebulin gene (NEB) at 2q21.1-q22, and the actin gene (ACTA1) at 1q42. The typical autosomal recessive form appears to be the most common one and is caused by mutations in the nebulin gene. We have studied the pattern of nebulin labeling, in patients with the typical congenital form (ten patients), the severe congenital form (two patients) or the mild, childhood-onset form (one patient), using antibodies against three different domains of nebulin. A qualitative and quantitative nebulin analysis in muscle tissue showed the presence of nebulin in myofibers from all patients. Some differences relating to the rod structure were observed. The majority of the largest subsarcolemmal rods were not labeled with the N2 nebulin antibody (I-band epitope) and showed an indistinct pattern with the two antibodies directed to the Z-band portion of nebulin (epitopes M176-181 and serine-rich domain). Diffuse rods were not revealed using the three antibodies. A discordant pattern of nebulin N2 epitope labeling was found in two affected sisters with a mutation in the nebulin gene, suggesting that modifications in nebulin distribution inside the rods might occur with the progression of the disease. Western blot analysis showed no direct correlation with immunofluorescence data. In nine patients, the band had a molecular weight comparable to the normal control, while in one patient, it was detected with a higher molecular weight. Our results suggest that presence/absence of specific nebulin Z-band epitopes in rod structures is variable and could depend on the degree of rod organization. PMID- 11257472 TI - Hyper-CK-aemia revisited. PMID- 11257473 TI - Longitudinal data analysis: an application to construction of a natural history profile of Duchenne muscular dystrophy. AB - A 30-month prospective study of 27 Scandinavian boys with confirmed diagnosis of Duchenne muscular dystrophy was carried out to construct profiles of the natural history of the disease. Assessments which included measures of voluntary muscle strength and function were done at 3 monthly intervals except for the first and second which were separated by 1 month. Recently developed statistical methods for analysis of longitudinal data with repeated observations on the same individual were used avoiding the problem of induced serial correlations. This allowed for the construction of both reference and prediction profiles for the variables %MRC, motor ability, walking time for 10 m and the sum of myometry of seven muscle groups. PMID- 11257474 TI - A standardized method for the evaluation of respiratory muscle endurance in patients with Duchenne muscular dystrophy. AB - The aim of the study was to develop a standardized method using controlled breathing to quantify respiratory muscle endurance in children with Duchenne muscular dystrophy (DMD) and to test its reproducibility. In 10 DMD patients, all between 10 and 14 years (mean age, 11.5 +/- 1.5 years), except for two patients of 20 and 22 years, and 10 healthy children (mean age, 12 +/- 1 years), we measured the maximal time (Tlim) that a threshold load fixed at 35% of the individual maximal inspiratory pressure (Pimax) could be tolerated. We asked the children to maintain their rest breathing pattern until exhaustion using visual feedback and an auditory signal. The mean Tlim in the DMD children was 4.45 +/- 1.45 min and values were reproducible. All healthy children were able to obtain Tlim values greater than 30 min. The respiratory muscles of DMD children are more susceptible to fatigue than those of healthy subjects. This method should be satisfactory for estimating the effect of treatment and for the specific training of respiratory muscles in DMD patients without significant learning disability. PMID- 11257475 TI - Evaluation of cardiac and respiratory involvement in sarcoglycanopathies. AB - Sarcoglycanopathies constitute a subgroup of limb-girdle recessive muscular dystrophies due to defects in sarcoglycan complex that comprises five distinct transmembrane proteins called alpha-, beta-, gamma-, delta-and epsilon sarcoglycans. As it is well known that sarcoglycans are expressed both in heart and in skeletal muscles and a complete deficiency in delta-sarcoglycan is the cause of the Syrian hamster BIO.14 cardiomyopathy, we studied cardiac and respiratory involvement in 20 patients with sarcoglycanopathies by clinical, electrocardiographic, echocardiographic, scintigraphic and spirometric assessments. A normal heart function was found in 31.3% of all patients; a preclinical cardiomyopathy in 43.7%; an arrhythmogenic cardiomyopathy in 6.3% and initial signs of dilated cardiomyopathy in 18.7%. In one patient the data were examined retrospectively. No correlation was found between cardiac and skeletal muscle involvement. With reference to the type of sarcoglycanopathy, signs of hypoxic myocardial damage occurred in beta-, gamma- and delta sarcoglycanopathies, while initial signs of a dilated cardiomyopathy in gamma- and delta-sarcoglycanopathies were found. A normal respiratory function was observed in 23.5% of all patients, a mild impairment in 35.4%, a moderate impairment in 29.4%, and a severe impairment in 11.7%. PMID- 11257476 TI - Cardiac abnormalities and skeletal muscle weakness in carriers of Duchenne and Becker muscular dystrophies and controls. AB - Cardiac abnormalities, cardiomyopathy and skeletal muscle weakness have been described in female carriers of the Xp21 (Duchenne and Becker) muscular dystrophies (J Neurol 1975;209(4):279-285; Br Med J 1969;2:418-420; J AmMed Assoc 1996;275(17):1335-1338; Neurology 1980;30(5):497-501; Neuromusc Disord 1999;9:347 351; Arch Neurol 1989;46:673-675). We have screened volunteers from our Xp21 genetics register and found the prevalence of previously unrecognized, clinically relevant, abnormalities in this group to be less than previously reported. We studied 91 women (56 carriers and 35 controls), aged between 18 and 69 years, from our local population known to the Oxford Regional Genetics Register. Our study included controls, with the investigators being blind to the subject's genetic status. The prevalence of previously unrecognised cardiac abnormalities on echocardiogram and ECG was 18% (10/56). Seven percent (4/56) of carriers had cardiomyopathy, defined by significant LV dilatation and decreased shortening fraction. In most cases, subjects with abnormal cardiac findings were asymptomatic. Echocardiography was more frequently abnormal than electrocardiography, but in many subjects the measurements of left ventricular dimensions were only just outside the normal ranges. The prevalence of skeletal muscle weakness was 12% (7/56). It was usually recognized by the individual, although not previously volunteered, but was mild and did not substantially affect activities of daily living. PMID- 11257477 TI - Force and power output of diaphragm muscle strips from mdx and control mice after clenbuterol treatment. AB - Based on its anabolic properties, treatment with the beta(2)-adrenoceptor agonist, clenbuterol, has been proposed as a strategy for ameliorating the symptoms of muscular dystrophy. In the dystrophic mdx mouse, only the diaphragm muscle exhibits progressive and severe degeneration in muscle structure and function similar to that observed in Duchenne muscular dystrophy. We tested the hypothesis that 20 weeks of clenbuterol treatment ( approximately 1.5-2 mg kg( 1)day(-1)) would increase the force and power output of diaphragm muscle strips of 6-month-old mdx and control mice. At this age, the diaphragm muscles of mdx mice show extensive degeneration and impaired contractility compared with control mice. Clenbuterol treatment did not increase the normalized force or power output of diaphragm strips from either mdx or control mice. The degeneration and necrosis within the diaphragm muscle of mdx mice was also not ameliorated by clenbuterol treatment. The results indicate that clenbuterol treatment does not improve the structure or function of diaphragm muscles from mdx mice. PMID- 11257479 TI - Therapeutic options in ocular myasthenia gravis. AB - The term ocular myasthenia gravis refers to the disease clinically restricted to extrinsic ocular muscles. It can be disabling as ptosis, and to a greater extent diplopia, both interfere with daily activities. Although ocular disturbances are the most frequent initial complaints in myasthenic patients, symptoms usually progress to generalized disease and only 15% of patients complain of purely ocular weakness for the entire course of their illness. Secondary generalization occurs with the highest frequency in the first 2 years from the onset. Both the severity of symptoms and the risk of generalization should be taken into account when devising a therapeutic plan for these patients. Anticholinesterases are of limited efficacy and a considerable proportion of patients require additional therapy. Corticosteroid therapy, generally prednisone on an alternate-day schedule, is very effective, but a reason for concern is represented by the frequent need for long-term administration with increased risk of severe complications. In patients unresponsive to prednisone or requiring too high dosages, immunosuppressive drugs like azathioprine should be used with the same criteria applied in generalized myasthenia. As corticosteroids and immunosuppressants reduce the chance of generalization, their use is justified in patients with recent-onset disabling disease. In long-standing cases with low risk of generalization, treatment is aimed at the relief of symptoms and pharmacological therapy should be reduced to the minimum effective dosage. The indication for thymectomy in ocular myasthenia remains highly controversial and should be reserved for disabled patients in the early stages of the disease. PMID- 11257478 TI - Extraocular muscle is spared despite the absence of an intact sarcoglycan complex in gamma- or delta-sarcoglycan-deficient mice. AB - Models of the dystrophin-glycoprotein complex do not reconcile the novel sparing of extraocular muscle in muscular dystrophy. Extraocular muscle sparing in Duchenne muscular dystrophy implies the existence of adaptive properties in these muscles that may extend protection to other neuromuscular diseases. We studied the extraocular muscle morphology and dystrophin-glycoprotein complex organization in murine targeted deletion of the gamma-sarcoglycan (gsg(-/-)) and delta-sarcoglycan (dsg(-/-)) genes, two models of autosomal recessive limb girdle muscular dystrophy. In contrast to limb and diaphragm, the principal extraocular muscles were intact in gsg(-/-) and dsg(-/-) mice. However, central nucleated, presumptive regenerative, fibers were seen in the accessory extraocular muscles (retractor bulbi, levator palpebrae superioris) of both strains. Skeletal muscles of gsg(-/-) mice exhibited in vivo Evans Blue dye permeability, while the principal extraocular muscles did not. Disruption of gamma-sarcoglycan produced secondary displacement of alpha- and beta-sarcoglycans in the extraocular muscles. The intensity of immunofluorescence for dystrophin and alpha- and beta dystroglycan also appeared to be slightly reduced. Utrophin localization was unchanged. The finding that sarcoglycan disruption was insufficient to elicit alterations in extraocular muscle suggests that loss of mechanical stability and increased sarcolemmal permeability are not inevitable consequences of mutations that disrupt the dystrophin-glycoprotein complex organization and must be accounted for in models of muscular dystrophy. PMID- 11257480 TI - Successful in vitro growth of rat two-cell embryos to blastocysts using a simple chemically defined medium. AB - The aim of the present study was to formulate a simple chemically defined medium for the in vitro growth of rat two-cell embryos to blastocysts. Embryos from day 2 pregnant rats were retrieved and placed in paraffin oil-covered droplets of "rat two-cell embryo culture medium" (R2ECM) containing combinations of various serum supplements, glucose, L-glutamine, and cultured up to 96 h in a CO(2) incubator. Embryos cultured in the basic medium (R2ECM), as well as those supplemented either with fetal bovine serum (FBS) or male rat serum (MRS) did not develop beyond the two- to four-cell stage. In R2ECM with 0.3% bovine serum albumin (BSA) and 7.5 mM glucose, 44% of embryos reached the blastocyst stage by 96 h in culture, and the blastulation rate increased to about 83% when 1 mM of L glutamine was added. To evaluate the effects of varying doses of glucose, two cell embryos were cultured in R2ECM supplemented with 0.3% BSA, 1 mM L-glutamine, and 2.5, 5.0, or 7.5 mM of glucose. The percentage of embryos reaching the blastocyst stage for 2.5, 5.0, and 7.5 mM glucose was 64.6%, 65.3%, and 82.9%, respectively. The present study showed that the modified medium (R2ECM) is a simple chemically defined medium that is capable of supporting in vitro growth of rat two-cell embryos to blastocysts in high proportion (greater than 80%) without the need for change of medium within 96 h of culture. PMID- 11257481 TI - Determination of epithelial magnesium transport with stable isotopes. AB - The inability to adequately determine Mg(2+) flux rates with radiotracer studies has stymied our efforts to understand how magnesium is transported by epithelial cells. To evaluate epithelial Mg(2+) transport, a stable 25Mg isotope was used to measure magnesium uptake into normal and Mg(2+)-depleted Madin-Darby canine kidney (MDCK) cells. 25Mg entry rates were significantly increased in Mg(2+) depleted cells relative to those cultured in normal magnesium media, 0.5 mM. 25Mg uptake was inhibited by external La(3+) but not Ca(2+) in both normal and Mg(2+) depleted cells suggesting a specific entry pathway. These results with 25Mg were the same as with microfluorescence determinations using mag-fura-2. We have shown that Mg(2+) entry into epithelial cells reflects transepithelial transport; accordingly, increased Mg(2+) uptake in Mg(2+)-depleted cells provides an important intrinsic control of renal magnesium absorption. Furthermore, these studies indicate that cellular Mg(2+) transport may be quantitated with the use of stable isotopes that may be successfully applied to cells other than epithelia. PMID- 11257482 TI - A transition metal enhanced luminol chemiluminescence in the presence of a chelator. AB - We have investigated the chemiluminescence signal of luminol and hydrogen peroxide in the presence of a transition metal (Co(II), Cu(I), Fe(II), Fe(III)) and of a chelator (EDTA, citric acid) in pH 8.5, 9 and 10 borate buffer solutions. We observed that the chemiluminescence intensities of these systems reached a plateau, where they remained stable for a period of 2-30 s. We also observed linearity between the intensity of chemiluminescence and the hydrogen peroxide concentration. The combination of Co(II) and EDTA at pH 9 was found to give the optimum signal with reference to time stability, intensity and reproducibility. Thus, compared to previous chemiluminescence applications, the present results permit us to propose a simple, enzyme-free and time-independent technique for the detection and quantification of hydrogen peroxide. PMID- 11257483 TI - Use of a single-pass in situ perfused rat hindlimb to study tissue distribution kinetics. Method development and experiences with Evans blue. AB - A single-pass in situ rat hindlimb preparation has been developed as an alternative to the isolated perfused preparation to study tissue drug distribution kinetics with minimal disturbance of the animal physiology. Evans blue (EB), a vascular marker, was administered (in saline or plasma) as a bolus into the femoral artery, and the total outflow blood was collected at timed intervals from the corresponding femoral vein. Donor blood was infused through the jugular vein at a rate that matched the loss. No changes in the blood flow or development of edema were observed. The outflow profile was characterized using statistical moments. Regardless of the injected solution, the estimated vascular volume (10% of the hindlimb wet weight) was higher than that reported for the isolated rat hindlimb (4.1%) but closer to the in vivo blood volume (6.8-8.1% of body weight for 250-g rat) and sum of vascular volumes of its composite tissues (6.9% of tissue weights). The large normalized variance (CV(2)) of the outflow (2.2+/-0.1) confirms the known heterogeneity of the hindlimb. Entrapment in the limb (approximately 4%) and escape to the rest of the body could explain the incomplete recovery (79-89%) of the dye. PMID- 11257484 TI - Lidocaine and surgical modification reduces mortality in a rat model of cardiac failure induced by coronary artery ligation. AB - Coronary artery ligation in the rat provides a useful experimental model of cardiac failure; however, this procedure carries with it a high mortality rate (50%). In this study, we used lidocaine (10 mg/kg, i.m.) before coronary artery ligation and 2 h after surgery to minimise the incidence of ventricular fibrillation (VF) that leads to sudden death in this model. We found that coronary artery ligation, using lidocaine in conjunction with a modified surgical procedure, had a mortality rate of 15%, much lower than reported in previous studies using this model. These modifications allow for the production of larger infarcts with 29% of animals having an infarct size > 50% of the epicardial surface. Infarct size in our myocardial infarction (MI) group varied between 5% and 75% of the left ventricular (LV) surface area resulting in a mean infarct size of 41.3 +/- 1.3% for the epicardial surface and 40.0 +/- 1.3% for the endocardial surface. PMID- 11257485 TI - Determination of the adsorption of tramadol hydrochloride by activated charcoal in vitro and in vivo. AB - Although tramadol is one of the most widely used centrally acting analgesics worldwide, no literature is available regarding adsorption of tramadol HCl powder or tablets (Ultram; 50 mg tramadol HCl per tablet) by activated charcoal (AC) for use as potential adjunct treatment of overdose. The present study incorporated a novel combination of in vitro and in vivo methods to investigate this question. Based on a binding curve of tramadol UV absorbance (UV(a); 225 nm) plotted against the amount of AC, the ratio of amount of tramadol completely adsorbed by AC was 0.05 mg/mg. Also based on UV(a), no tramadol was detected in filtrate of slurries in which up to 62 tablets of Ultram were mixed with 50 g AC; 4.6% of unbound tramadol was detected when 100 tablets of Ultram were mixed with AC. The ratio of amount of tramadol completely adsorbed by AC in this test was 0.10. In vivo, co-administration of 0.1 g/ml of AC produced a 13- to 14-fold rightward shift in tramadol's antinociceptive dose-response curve and a 1.6-fold rightward shift in tramadol's lethality dose-response curve. PMID- 11257486 TI - Continuous assessment of multiple vital physiological functions in conscious freely moving rats using telemetry and a plethysmography system. AB - The general pharmacologist in the pharmaceutical industry is challenged to generate physiologically relevant data on possible safety liabilities or on secondary therapeutic uses as early as possible in drug development. This implies the need for efficient use of usually only small supplies of test article. For this reason, we have developed a new animal model combining various elements to provide a broad spectrum of data focussing on the so-called vital physiological functions: cardiovascular, respiratory, and central nervous system. This system uses rats chronically implanted with transmitters for the measurement of arterial pressure, ECG, and body temperature. Modification of the transmitters also allows for the simultaneous assessment of locomotor activity. Studies are performed with these rats in plethysmographs placed directly over the antenna units thus allowing for the additional assessment of respiratory function in the same studies. Using this system, we can generate simultaneously a wide range of relevant physiological parameters in conscious rats with a modest requirement for test article. Such an approach is highly useful for getting early safety readouts of potential drug development candidates as well as for detecting possible secondary therapeutic actions of a drug. PMID- 11257487 TI - Inflammation induced by latex of Calotropis procera--a new model to evaluate anti inflammatory drugs. AB - Latex of Calotropis procera was studied for its inflammatory reactions using pedal oedema and air pouch models of inflammation in rats. Subcutaneous injection of aqueous solution (0.1 ml of 1%) of dry latex (DL) into the plantar surface of paw produced significant inflammation. Maximum inflammatory response was obtained 1 h after the injection and was maintained for a further 1 h. The inflammatory response was accompanied by an increase in vascular permeability that reached its maximum within 15 min. Inflammation was also induced in the 6-day-old rat air pouch by injecting a 2.5% solution of DL. The latter model was characterized for the exudate volume and its protein concentration, and wet and dry weights of granuloma. A time-course study indicated that both the exudate volume and the weight of granuloma were at maximum on day 5 after DL injection while the protein concentration peaked on the third day. Further, the two models were also studied for the anti-inflammatory effect of various drugs. It was observed that in the pedal oedema model, phenylbutazone was more effective than prednisolone while almost complete inhibition was produced by mepyramine and cyproheptadine. On the other hand, in the air pouch model, prednisolone was more effective than phenylbutazone in inhibiting the inflammation. Thus, the DL-induced inflammation in different models could be used to evaluate anti-inflammatory drugs. PMID- 11257488 TI - Effect of perfusate albumin on organ viability and vascular responses in the in vitro dual-perfused rat liver. AB - Inclusion of albumin in the perfusate has been previously shown to be detrimental to liver function, but its effect on hepatic vascular reactivity remains unknown. The aim of this study was to determine the effects of albumin on hepatic arterial vascular reactivity and liver viability in the isolated dual-perfused rat liver. A total of 12 rat livers were perfused with Krebs-Bulbring buffer without (Group 1) and with (Group 2) addition of 1% bovine serum albumin (BSA) through the hepatic artery and portal vein for up to 5 h. Hepatic arterial responses to acetylcholine and sodium nitroprusside were studied at 30-min intervals. Liver viability was assessed by bile volume production, release of aspartate serine aminotransferase (AST) and lactic acid dehydrogenase (LDH), and histological examination. Hepatic arterial responses to acetylcholine were significantly attenuated in Group 2. No significant differences in sodium nitroprusside responses were noted. However, bile volume production in Group 2 was significantly decreased compared to Group 1. Effluent AST and LDH release increased significantly in Group 1 but not in Group 2. Histological results showed that sinusoidal endothelial cells and hepatocytes were well preserved without significant deterioration in either group, although there was a marked decrease in vasodilatation to acetylcholine in Group 2. This data suggested that the presence of albumin in the perfusate did not improve retention of smooth muscle reactivity and reduced endothelium-dependent vasodilatation and bile volume production during perfusion. However, improved liver parenchymal cell function was observed. PMID- 11257489 TI - A docking model of key components of the DISC complex: death domain superfamily interactions redefined. AB - Apoptosis is mediated by a highly regulated signal transduction cascade that eventually leads to precisely directed cell death. The death-inducing signaling complex (DISC), composed of Fas, FADD, and caspase-8, is an apical signaling complex that mediates receptor-induced apoptosis. We have docked the experimentally determined structures of the Fas and FADD death domains into a model of a partial DISC signaling complex. The arrangement of Fas and FADD was determined using the interaction modes of the two heterodimer crystal structures determined to date, Pelle/Tube and Apaf-1/procaspase-9. The proposed model reveals that both interactions can be accommodated in a single multimeric complex. Importantly, the model is consistent with reported site-directed mutagenesis data indicating residues throughout the domain are critical for function. These results imply that members of the death domain superfamily have the potential for multivalent interactions, offering novel possibilities for regulation of apoptotic signaling. PMID- 11257490 TI - The molecular peculiarities of catalase-peroxidases. AB - In developing ideas of how protein structure modifies haem reactivity, the activity of Class I of the plant peroxidase superfamily (including cytochrome c peroxidase, ascorbate peroxidase and catalase-peroxidases (KatGs)) is an exciting field of research. Despite striking sequence homologies, there are dramatic differences in catalytic activity and substrate specificity with KatGs being the only member with substantial catalase activity. Based on multiple sequence alignment performed for Class I peroxidases, we present a hypothesis for the pronounced catalase activity of KatGs. In their catalytic domains KatGs are shown to possess three large insertions, two of them are typical for KatGs showing highly conserved sequence patterns. Besides an extra C-terminal copy of the ancestral hydroperoxidase gene resulting from gene duplication, these two large loops are likely to control the orientation of both the haem group and of essential residues in the active site. They seem to modulate the access of substrates to the prosthetic group at the distal side as well as the flexibility and character of the bond between the proximal histidine and the ferric iron. The hypothesis presented opens new possibilities in the rational engineering of peroxidases. PMID- 11257491 TI - The structure of glutamate transporters shows channel-like features. AB - Neuronal and glial glutamate transporters remove the excitatory neurotransmitter glutamate from the synaptic cleft and thus prevent neurotoxicity. The proteins belong to a large family of secondary transporters, which includes transporters from a variety of bacterial, archaeal and eukaryotic organisms. The transporters consist of eight membrane-spanning alpha-helices and two pore-loop structures, which are unique among secondary transporters but may resemble pore-loops found in ion channels. Another distinctive structural feature is the presence of a highly amphipathic membrane-spanning alpha-helix that provides a hydrophilic path through the membrane. The unusual structural features of the transporters are discussed in relation to their function. PMID- 11257492 TI - Anticoagulants and inhibitors of platelet aggregation derived from leeches. AB - Increased life expectancy is associated with aging populations in the developed countries, and we can expect an increased incidence of cardiovascular and inflammatory diseases and cancers. A priority for medical research is to reduce such morbidity. Leeches have been demonstrated to be a useful source of drugs to treat cardiovascular diseases, as they have evolved highly specific mechanisms to feed on their hosts by blocking blood coagulation. Powerful molecules acting at different points in the coagulation cascade or in the inhibition of platelet aggregation have been purified from these animals. Moreover, clinical trials confirm their potential to treat cardiovascular diseases. PMID- 11257493 TI - The TIM-barrel fold: a versatile framework for efficient enzymes. AB - Recent studies on triosephosphate isomerase (TIM)-barrel enzymes highlight the remarkable versatility of the TIM-barrel scaffold. At least 15 distinct enzyme families use this framework to generate the appropriate active site geometry, always at the C-terminal end of the eight parallel beta-strands of the barrel. Sequence and structure comparisons now suggest that many of the TIM-barrel enzymes are evolutionarily related. Common structural properties of TIM-barrel enzymes are discussed. PMID- 11257494 TI - Protein kinase B (PKB/Akt)--a key regulator of glucose transport? AB - The serine/threonine kinase protein kinase B (PKB/Akt) has been shown to play a crucial role in the control of diverse and important cellular functions such as cell survival and glycogen metabolism. There is also convincing evidence that PKB plays a role in the insulin-mediated regulation of glucose transport. Furthermore, states of cellular insulin resistance have been shown to involve impaired PKB activation, and this usually coincides with a loss of glucose transport activation. However, evidence to the contrary is also available, and the role of PKB in the control of glucose transport remains controversial. Here we provide an overview of recent findings, discuss the potential importance of PKB in the regulation of glucose transport and metabolism, and comment on future directions. PMID- 11257495 TI - A new family of small, palmitoylated, membrane-associated proteins, characterized by the presence of a cysteine-rich hydrophobic motif. AB - We recently cloned the CHIC2 gene (previously BTL) by virtue of its involvement in a chromosomal translocation t(4;12)(q11;p13) occurring in acute myeloid leukemias. In this study we show that CHIC2 is a member of a highly conserved family of proteins characterized by the presence of a striking cysteine-rich hydrophobic (CHIC) motif. Our data illustrate that cysteines in this central CHIC motif are palmitoylated and that CHIC2 is associated with vesicular structures and the plasma membrane. The CHIC proteins thus resemble the cysteine string proteins, which function in regulated exocytosis. PMID- 11257496 TI - Corticosterone-induced rapid phosphorylation of p38 and JNK mitogen-activated protein kinases in PC12 cells. AB - The present study showed that corticosterone (B) could induce a rapid activation of p38 and c-Jun NH(2)-terminal protein kinase (JNK) in PC12 cells. The dose response and time-response curves were bell-shaped with maximal activation at 10( 9) M and at 15 min. RU38486 had no effect, and bovine serum albumin-coupled B could induce the activation. Genistein failed to block the phosphorylation, suggesting the pathway was not involved in tyrosine kinase activity. Phorbol 12 myristate 13-acetate could mimic, while Go6976 could abolish the actions. These results demonstrated that B might act via a putative membrane receptor to activate p38 and JNK rapidly through a protein kinase C-dependent pathway. PMID- 11257497 TI - Scar/WAVE is localised at the tips of protruding lamellipodia in living cells. AB - Cell motility entails the extension of cytoplasmic processes, termed lamellipodia and filopodia. Extension is driven by actin polymerisation at the tips of these processes via molecular complexes that remain to be characterised. We show here that a green fluorescent protein (GFP) fusion of the Wiskott-Aldrich syndrome protein family member Scar1/WAVE1 is specifically recruited to the tips of lamellipodia in living B16F1 melanoma cells. Scar1-GFP was recruited only to protruding lamellipodia and was absent from filopodia. The localisation of Scar was facilitated by the finding that the formerly described inhibition of lamellipodia formation by ectopical expression of Scar, could be overcome by the treatment of cells with aluminium fluoride. These findings show that Scar is strategically located at sites of actin polymerisation specifically engaged in the protrusion of lamellipodia. PMID- 11257498 TI - Tau-tubulin kinase phosphorylates tau at Ser-208 and Ser-210, sites found in paired helical filament-tau. AB - Hyperphosphorylated tau protein is known to be a major component of the paired helical filaments (PHFs) that accumulate in the brain of Alzheimer's patients. The kinase that phosphorylated Ser-208 and Ser-210 in PHF-tau had remained unknown. We used anti-pS208 and anti-pS210 antibodies and Western blots to confirm that the tau-tubulin kinase (TTK) phosphorylates tau at Ser-208 and at Ser-210. Using partial amino acid sequences of purified bovine brain TTK, a mouse cDNA of TTK was isolated and the sequence was determined. Its 963 bp coding region is composed of 320 amino acids and encodes a 36 kDa protein indistinguishable in size from authentic bovine brain TTK. Our immunoblot analysis demonstrated that TTK is ubiquitously distributed in the rat tissues, and that it is developmentally regulated in the rat brain. PMID- 11257499 TI - The C-terminus of dUTPase: observation on flexibility using NMR. AB - The dynamics of the C-terminus of the dUTPases from Escherichia coli and equine infectious anaemia virus (EIAV) were studied by 1H-(15)N nuclear magnetic resonance spectroscopy. The two enzymes differ with regard to flexibility in the backbone of the 15 most C-terminal amino acid residues, some of which are conserved and essential for enzymic activity. In the bacterial enzyme, the residues closest to the C-terminus are highly flexible and display a correlation time in the nanosecond time range. No similar high flexibility could be detected for the C-terminal part of EIAV dUTPase, indicating a different time range of flexibility. PMID- 11257500 TI - A cis-dominant cyclic nucleotide-dependent regulatory domain in the 3' untranslated region of Na(+)/glucose cotransporter (SGLT1) mRNA. AB - A 122 nt uridine-rich sequence (URE) in the Na(+)/glucose cotransporter (SGLT1) mRNA 3'-untranslated region is critical for cAMP-dependent message stabilization. Its function was investigated in LLC-PK(1) cells stably expressing beta-globin reporter transcripts. Insertion of the SGLT1 URE downstream from an unrelated destabilizing sequence, the c-fos ARE, evoked cAMP-dependent message stabilization. Stabilization was blocked by a substitution mutation within the SLGT1 URE. These observations indicate that the SGLT1 URE is sufficient to transmit cAMP-dependent, cis-dominant mRNA stabilization in the presence of appropriate trans-acting factors and appears to function independently of the nature of the destabilizing domain. PMID- 11257501 TI - Nucleotides U28-A42 and A37 in unmodified yeast tRNA(Trp) as negative identity elements for bovine tryptophanyl-tRNA synthetase. AB - Wild-type bovine and yeast tRNA(Trp) are efficiently aminoacylated by tryptophanyl-tRNA synthetase both from beef and from yeast. Upon loss of modified bases in the synthetic transcripts, mammalian tRNA(Trp) retains the double recognition by the two synthetases, while yeast tRNA(Trp) loses its substrate properties for the bovine enzyme and is recognised only by the cognate synthetase. By testing chimeric bovine-yeast transcripts with tryptophanyl-tRNA synthetase purified from beef pancreas, the nucleotides responsible for the loss of charging of the synthetic yeast transcript have been localised in the anticodon arm. A complete loss of charging akin to that observed with the yeast transcript requires substitution in the bovine backbone of G37 in the anticodon loop with yeast A37 and of C28-G42 in the anticodon stem with yeast U28-A42. Since A37 does not prevent aminoacylation of the wild-type yeast tRNA(Trp) by the beef enzyme, a negative combination apparently emerges in the synthetic transcript after unmasking of U28 by loss of pseudourydilation. PMID- 11257502 TI - Aspartic protease in leaves of common bean (Phaseolus vulgaris L.) and cowpea (Vigna unguiculata L. Walp): enzymatic activity, gene expression and relation to drought susceptibility. AB - Four cultivars of related species, common bean and cowpea, which exhibit different degrees of drought resistance, were submitted to water stress by withholding irrigation. Drought induced an increase in endoproteolytic activity, being higher in susceptible cultivars (bean) than in tolerant ones (cowpea). An aspartic protease activity was found to be strongly induced especially in bean. From a cowpea leaf cDNA library, a full length aspartic protease precursor cDNA was obtained. Transcript accumulation in response to water stress indicated that the expression of the gene was constitutive in cowpea and transcriptionally up regulated in bean. The results showed that drought-tolerant and drought susceptible bean plants differ regarding aspartic protease precursor gene expression. PMID- 11257503 TI - Cross-talk between IL-6 and TGF-beta signaling in hepatoma cells. AB - Interleukin-6 (IL-6) is a multifunctional cytokine that plays important roles in the immune system, hematopoiesis, and acute phase reactions. Transforming growth factor-beta (TGF-beta) also has pleiotropy including the production of acute phase proteins in hepatocytes. To elucidate the cross-talk between IL-6 and TGF beta signaling pathways in hepatic cells, we investigated the effects of TGF-beta on IL-6-induced signal transducer and activator of transcription-3 (STAT3) activation in a human hepatoma cell line, Hep3B. IL-6-induced activation of STAT3 activity and STAT3-mediated gene expression were augmented by TGF-beta in Hep3B cells. We provide evidence that these activities were due to physical interactions between STAT3 and Sma- and MAD-related protein-3, bridged by p300. These results demonstrate a molecular mechanism of a cross-talk between STAT3 and TGF-beta signaling pathways in hepatocytes. PMID- 11257504 TI - Spontaneous subunit exchange in porcine liver fructose-1,6-bisphosphatase. AB - No evidence to date suggests the possibility of subunit exchange between tetramers of mammalian fructose-1,6-bisphosphatase. An engineered fructose-1,6 bisphosphatase, with subunits of altered electrostatic charge, exhibits spontaneous subunit exchange with wild-type enzyme in the absence of ligands. The exchange process reaches equilibrium in approximately 5 h at 4 degrees C, as monitored by non-denaturing gel electrophoresis and anion exchange chromatography. Active site ligands, such as fructose 6-phosphate, abolish subunit exchange at the level of the monomer, but permit dimer-dimer exchanges. AMP, alone or in the presence of active site ligands, abolishes all exchange processes. Exchange phenomena may play a role in the kinetic mechanism of allosteric regulation of fructose-1,6-bisphosphatase. PMID- 11257505 TI - Relationship of sequence and structure to specificity in the alpha-amylase family of enzymes. AB - The hydrolases and transferases that constitute the alpha-amylase family are multidomain proteins, but each has a catalytic domain in the form of a (beta/alpha)(8)-barrel, with the active site being at the C-terminal end of the barrel beta-strands. Although the enzymes are believed to share the same catalytic acids and a common mechanism of action, they have been assigned to three separate families - 13, 70 and 77 - in the classification scheme for glycoside hydrolases and transferases that is based on amino acid sequence similarities. Each enzyme has one glutamic acid and two aspartic acid residues necessary for activity, while most enzymes of the family also contain two histidine residues critical for transition state stabilisation. These five residues occur in four short sequences conserved throughout the family, and within such sequences some key amino acid residues are related to enzyme specificity. A table is given showing motifs distinctive for each specificity as extracted from 316 sequences, which should aid in identifying the enzyme from primary structure information. Where appropriate, existing problems with identification of some enzymes of the family are pointed out. For enzymes of known three-dimensional structure, action is discussed in terms of molecular architecture. The sequence-specificity and structure-specificity relationships described may provide useful pointers for rational protein engineering. PMID- 11257506 TI - Molecular enzymology of carnitine transfer and transport. AB - Carnitine (L-3-hydroxy-4-N-trimethylaminobutyric acid) forms esters with a wide range of acyl groups and functions to transport and excrete these groups. It is found in most cells at millimolar levels after uptake via the sodium-dependent carrier, OCTN2. The acylation state of the mobile carnitine pool is linked to that of the limited and compartmentalised coenzyme A pools by the action of the family of carnitine acyltransferases and the mitochondrial membrane transporter, CACT. The genes and sequences of the carriers and the acyltransferases are reviewed along with mutations that affect activity. After summarising the accepted enzymatic background, recent molecular studies on the carnitine acyltransferases are described to provide a picture of the role and function of these freely reversible enzymes. The kinetic and chemical mechanisms are also discussed in relation to the different inhibitors under study for their potential to control diseases of lipid metabolism. PMID- 11257507 TI - Active site structure of the catalase-peroxidases from Mycobacterium tuberculosis and Escherichia coli by extended X-ray absorption fine structure analysis. AB - The catalase-peroxidase encoded by katG of Mycobacterium tuberculosis is a more effective activator of the antibiotic isoniazid than is the equivalent enzyme from Escherichia coli. The environment of the heme iron was investigated using X ray absorption spectroscopy to determine if differences in this region were associated with the differences in reactivity. The variation in the distal side Fe-ligand distances between the two enzymes was the same within experimental error indicating that it was not the heme iron environment that produced the differences in reactivity. Analysis of variants of the E. coli catalase peroxidase containing changes in active site residues Arg102 and His106 revealed small differences in Fe-water ligand distance including a shorter distance for the His106Tyr variant. The Arg102Leu variant was 5-coordinate, but His106Cys and Arg102Cys variants showed no changes within experimental error. These results are compared with those reported for other peroxidases. PMID- 11257508 TI - Structural determinants outside the PXDLS sequence affect the interaction of adenovirus E1A, C-terminal interacting protein and Drosophila repressors with C terminal binding protein. AB - C-Terminal binding protein (CtBP) interacts with a highly conserved amino acid motif (PXDLS) at the C terminus of adenovirus early region 1A (AdE1A) protein. This amino acid sequence has recently been demonstrated in the mammalian protein C-terminal interacting protein (CtIP) and a number of Drosophila repressors including Snail, Knirps and Hairy. In the study described here we have examined the structures of synthetic peptides identical to the CtBP binding sites on these proteins using NMR spectroscopy. It has been shown that peptides identical to the CtBP binding site in CtIP and at the N terminus of Snail form a series of beta turns similar to those seen in AdE1A. The PXDLS motif towards the C terminus of Snail forms an alpha-helix. However, the motifs in Knirps and Hairy did not adopt well-defined structures in TFE/water mixtures as shown by the absence of medium range NOEs and a high proportion of signal overlap. The affinities of peptides for Drosophila and mammalian CtBP were compared using enzyme-linked immunosorbent assay. CtIP, Snail (N-terminal peptide) and Knirps peptides all bind to mammalian CtBP with high affinity (K(i) of 1.04, 1.34 and 0.52 microM, respectively). However, different effects were observed with dCtBP, most notably the affinity for the Snail (N-terminal peptide) and Knirps peptides were markedly reduced (K(i) of 332 and 56 microM, respectively) whilst the Hairy peptide bound much more strongly (K(i) for dCtBP of 6.22 compared to 133 microM for hCtBP). In addition we have shown that peptides containing identical PXDLS motifs but with different N and C terminal sequences have appreciably different affinities for mammalian CtBP and different structures in solution. We conclude that the factors governing the interactions of CtBPs with partner proteins are more complex than simple possession of the PXDLS motif. In particular the overall secondary structures and amino acid side chains in the binding sites of partner proteins are of importance as well as possible global structural effects in both members of the complex. These data are considered evidence for a multiplicity of CtBPs and partner proteins in the cell. PMID- 11257509 TI - Hydrophobicity of the NADPH binding domain of camel lens zeta-crystallin. AB - Interaction of camel lens zeta-crystallin with the hydrophobic probe 1 anilinonaphthalene-8-sulfonic acid (ANS) enhanced the ANS fluorescence and quenched the protein fluorescence. Both of these events were concentration dependent and showed typical saturation curves suggesting specific ANS-zeta crystallin binding. Quantitative analysis indicated that 1 mole zeta-crystallin bound at most 1 mole ANS. NADPH but not 9,10-phenanthrenequinone (PQ) was able to displace zeta-crystallin-bound ANS. These results suggested the presence of a hydrophobic domain in zeta-crystallin, possibly at the NADPH binding site. alpha Crystallin as well as NADPH protected zeta-crystallin against thermal inactivation suggesting the importance of this site for enzyme stability. The NADPH:quinone oxidoreductase activity of zeta-crystallin was inhibited by ANS with NADPH as electron donor and PQ as electron acceptor. Lineweaver-Burk plots indicated mixed-type inhibition with respect to NADPH, with a K(i) of 2.3 microM. Secondary plots of inhibition with respect to NADPH indicated a dissociation constant (K'I) of 12 microM for the zeta-crystallin-NADPH-ANS complex. The K(i) being smaller than K'I suggested that competitive inhibition at the NADPH binding site was predominant over non-competitive inhibition. Like ANS-zeta-crystallin binding, inhibition was dependent on ANS concentration but independent of incubation time. PMID- 11257510 TI - Different domains in the third intracellular loop of the GLP-1 receptor are responsible for Galpha(s) and Galpha(i)/Galpha(o) activation. AB - It has previously been shown that the GLP-1 receptor is primarily coupled to the adenylate cyclase pathway via activation of Galpha(s) proteins. Recent studies have shown that the third intracellular loop of the receptor is important in the stimulation of cAMP production. We have studied the effect of three synthetic peptide sequences derived from the third intracellular loop of the GLP-1 receptor on signal transduction in Rin m5F cell membranes. The whole third intracellular loop strongly stimulates both pertussis toxin and cholera toxin-sensitive G proteins, while the N-terminal half exclusively stimulates cholera toxin sensitive G proteins and the C-terminal half only stimulates pertussis toxin sensitive G-proteins as demonstrated by measurements of GTPase activity. These data confirm that the principal stimulatory G-protein interaction site resides in the third intracellular loop, but also suggest that the GLP-1 receptor is not only coupled to the Galpha(s) but also to the Galpha(i)/Galpha(o) type of G proteins and that distinct domains within the third intracellular loop are responsible for the activation of the different G-protein subfamilies. PMID- 11257511 TI - Urea equilibrium unfolding of the major core protein of the retrovirus feline immunodeficiency virus and its tryptophan mutants. AB - Circular dichroism and fluorescence spectroscopy have been employed to study the urea unfolding mechanism of a recombinant form of the major core protein of feline immunodeficiency virus (FIV-rp24) and its native tryptophan mutants. The equilibrium denaturation curves indicate the existence of two transitions. The first unfolding transition most likely reflects the denaturation of the carboxy terminal region of FIV-rp24. Consequently, the second transition, where the changes in fluorescence are produced, should reflect the denaturation of the amino-terminal region. If the intermediate observed upon urea denaturation is an on-pathway species, the data described herein can reflect the sequential and independent loss of structure of the two domains that this type of proteins possesses. PMID- 11257512 TI - X-Ray study on an artificial mung bean inhibitor complex with bovine beta-trypsin in neat cyclohexane. AB - The active trypsin inhibiting component, SPC1, was obtained during the synthesis of a 22-residue peptide with three disulfide bridges according to the mimic mung bean Bowman-Birk type inhibitor. The K(i) value of SPC1 is 1.2x10(-7) M. In order to determine the topological structure of SPC1, X-ray diffraction studies were carried out on the complex of SPC1 with bovine beta-trypsin. Only the binding loop of SPC1 resolved at 2.2 A resolution due to conformational flexibility of the other residues [1]. The amino acid sequence was re-determined and electrospray mass spectroscopy was also performed to ensure that no cleaving occurred on SPC1 and the primary sequence of SPC1 is correct. Because the protein is more rigid in nonaqueous medium as has been proved by others [2], we treated the complex of SPC1 with neat cyclohexane and then subjected it to X-ray diffraction analysis, and the result showed that all the 22 residues of SPC1 were located in the electron density map. So the topological structure of SPC1 has been determined, suggesting that crystal treatment with cyclohexane may be used as a method to determine the conformation of the disordered regions in protein crystal structures. PMID- 11257513 TI - Expression of Pleurotus eryngii aryl-alcohol oxidase in Aspergillus nidulans: purification and characterization of the recombinant enzyme. AB - Aryl-alcohol oxidase (AAO) is an extracellular flavoenzyme involved in lignin biodegradation by some white-rot fungi. The enzyme catalyzes the extracellular oxidation of aromatic alcohols to the corresponding aldehydes. The electron acceptor is molecular oxygen yielding H(2)O(2) as the product. Herein we describe, for the first time, the expression of AAO from Pleurotus eryngii in the ascomycete Aspergillus nidulans. The activity of the recombinant enzyme in A. nidulans cultures is much higher than found in the extracellular fluid of P. eryngii. The recombinant enzyme showed the same molecular mass, pI and catalytic properties as that of the mature protein secreted by P. eryngii. The enzymic properties are also similar to those reported from other Pleurotus and Bjerkandera species. PMID- 11257514 TI - Plasticity of secondary structure in the N-terminal region of beta-dystroglycan. AB - The secondary structure content of the N-terminal extracellular domain of beta dystroglycan (a recombinant fragment extending from positions 654 to 750) has been quantitatively determined by means of CD and FTIR spectroscopies. The elements of secondary structure, namely an 8-10 residue long alpha-helix (10%) and two beta-strands (24%) have been assigned to specific amino acid sequences by means of a GOR constrained prediction method. The remaining 66% of the whole sequence is classified as turns or unordered. The temperature dependence of CD and FTIR spectra has been investigated in detail. A reversible, non-cooperative thermal transition is observed with both CD and FTIR spectroscopies up to 95 degrees C. The profile of the transition is typical of the unfolding of isolated peptides and corresponds to the progressive loss of the secondary structure elements of the protein with no evidence for collapsing phenomena, typical of globular proteins, upon heating. PMID- 11257515 TI - Structural and functional studies of alpha-helix 5 region from Bacillus thuringiensis Cry1Ab delta-endotoxin. AB - The crystal insecticidal proteins from Bacillus thuringiensis are modular proteins comprised of three domains connected by single linkers. Domain I is a seven alpha-helix bundle, which has been involved in membrane insertion and pore formation activity. Site-directed mutagenesis has contributed to identify regions that might play an important role in the structure of the pore-forming domain within the membrane. There are several evidences that support that the hairpin alpha4-alpha5 inserts into the membrane in an antiparallel manner, while other helices lie on the membrane surface. We hypothesized that highly conserved residues of alpha5 could play an important role in toxin insertion, oligomerization and/or pore formation. A total of 15 Cry1Ab mutants located in six conserved residues of Cry1Ab, Y153, Y161, H168, R173, W182 and G183, were isolated. Eleven mutants were located within helix alpha5, one mutant was located in the loop alpha4-alpha5 and three mutants, W182P, W182I and G183C, were located in the loop alpha5-alpha6. Their effect on binding, K(+) permeability and toxicity against Manduca sexta larvae was analyzed and compared. The results provide direct evidence that some residues located within alpha5 have an important role in stability of the toxin within the insect gut, while some others also have an important role in pore formation. The results also provide evidence that conserved residues within helix alpha5 are not involved in oligomer formation since mutations in these residues are able to make pores in vitro. PMID- 11257516 TI - Evidence of dimerisation among class D beta-lactamases: kinetics of OXA-14 beta lactamase. AB - OXA-14 enzyme, a class D beta-lactamase, gave biphasic kinetics with all penicillin and cephalosporin substrates tested, such that the catalytic rate declined more swiftly than was explicable by substrate depletion. This biphasic behaviour was independent of temperature or extraneous protein but was lost if the enzyme was diluted to occupy almost the total assay volume before addition of a small amount of concentrated substrate. The presence of substrate could partially protect the enzyme against conversion to the less active form, with protection greatest at substrate concentration above the K(m). These observations are compatible with the hypothesis that the biphasic kinetics depended on the enzyme existing as a highly active dimer at high concentration and as a less active monomer at low concentration. Direct evidence supporting this hypothesis came from the observation that gel exclusion chromatography indicated a higher molecular weight for concentrated enzyme than for dilute. Biphasic kinetics are not so universal for different substrates amongst beta-lactamases (OXA-10, -11, 13, -16 and -17) that differ from OXA-14 by only one to two amino acid substitutions. It may be that the monomer:dimer equilibrium is more rapidly achieved with these enzymes than with OXA-14, or that the kinetic properties of the dimers and monomers of these enzymes are similar, masking any biphasic trait. PMID- 11257517 TI - Assessment of P450 induction in the marmoset monkey using targeted anti-peptide antibodies. AB - The identity and expression of hepatic P450 enzymes in marmosets was investigated using a panel of anti-peptide antibodies originally targeted against human P450 enzymes. In immunoblotting, of 12 antibodies examined, 10 bound specifically to bands in marmoset liver microsomal fraction corresponding to P450 enzymes. It is proposed that these represent marmoset CYP1A1, CYP1A2, CYP2A, CYP2B, CYP2C forms (CYP2C-1 and CYP2C-2), CYP2D19, CYP3A21 and another CYP3A form (CYP3A-m). The antibodies, together with an anti-marmoset CYP2E1 antibody, were used to investigate the expression of 10 P450 enzymes in marmosets treated with P450 inducing chemicals. Treatment with phenobarbitone caused CYP2B, CYP2C-2 and CYP3A21 levels to increase, rifampicin caused increases in CYP2B and CYP2C-1 and a decrease in CYP3A21 levels, whereas dioxin caused CYP1A1 and CYP1A2 levels to increase and CYP2E1 levels to decrease. Clofibric acid did not induce any P450. P450 enzyme activities were assessed using 8 different substrates and increases were found after treatment with phenobarbitone, rifampicin, and dioxin. However, due to species differences in substrate selectivity, it proved difficult to ascribe these changes to individual P450 enzymes. Thus, the use of anti-peptide antibodies provides a more informative way of assessing the levels of specific P450 enzymes than enzyme activity measurements. PMID- 11257518 TI - Extensive interactions between HIV TAT and TAF(II)250. AB - The HIV transactivator, Tat, has been shown to be capable of potent repression of transcription initiation. Repression is mediated by the C-terminal segment of Tat, which binds the TFIID component, TAF(II)250, although the site(s) of interaction were not defined previously. We now report that the interaction between Tat and TAF(II)250 is extensive and involves multiple contacts between the Tat protein and TAF(II)250. The C-terminal domain of Tat, which is necessary for repression of transcription initiation, binds to a segment of TAF(II)250 that encompasses its acetyl transferase (AT) domain (885-1034 amino acids (aa)). Surprisingly, the N-terminal segment of Tat, which contains its activation domains, also binds to TAF(II)250 and interacts with two discontinuous segments of TAF(II)250 located between 885 and 984 aa and 1120 and 1279 aa. Binding of Tat to the 885-984 aa segment of TAF(II)250 requires the cysteine-rich domain of Tat, but not the acidic or glutamine-rich domains. Binding by the N-terminal domain of Tat to the 1120-1279 aa TAF(II)250 segment does not involve the acidic, cysteine- or glutamine-rich domains. Repression of transcription initiation by Tat requires functional TAF(II)250. We now demonstrate that transcription of the HIV LTR does not depend on TAF(II)250 which may account for its resistance to Tat mediated repression. PMID- 11257519 TI - Osmolytes protect mitochondrial F(0)F(1)-ATPase complex against pressure inactivation. AB - We have previously reported that carbohydrates and polyols protect different enzymes against thermal inactivation and deleterious effects promoted by guanidinium chloride and urea. Here, we show that these osmolytes (carbohydrates, polyols and methylamines) protect mitochondrial F(0)F(1)-ATPase against pressure inactivation. Pressure stability of mitochondrial F(0)F(1)-ATPase complex by osmolytes was studied using preparations of membrane-bound submitochondrial particles depleted or containing inhibitor protein (IP). Hydrostatic pressure in the range from 0.5 to 2.0 kbar causes inactivation of submitochondrial particles depleted of IP (AS particles). However, the osmolytes prevent pressure inactivation of the complex in a dose-dependent manner, remaining up to 80% of hydrolytic activity at the highest osmolyte concentration. Submitochondrial particles containing IP (MgATP-SMP) exhibit low ATPase activity and dissociation of IP increases the hydrolytic activity of the enzyme. MgATP-SMP subjected to pressure (2.2 kbar, for 1 h) and then preincubated at 42 degrees C to undergo activation did not have an increase in activity. However, particles pressurized in the presence of 1.5 M of sucrose or 3.0 M of glucose were protected and after preincubation at 42 degrees C, showed an activation very similarly to those kept at 1 bar. In accordance with the preferential hydration theory, we believe that osmolytes reduce to a minimum the surface of the macromolecule to be hydrated and oppose pressure-induced alterations of the native fold that are driven by hydration forces. PMID- 11257520 TI - Analysis of the physicochemical interactions between Clostridium difficile toxins and cholestyramine using liquid chromatography with post-column derivatization. AB - A potential therapy for antibiotic-associated pseudomembranous colitis is to bind Clostridium difficile toxins A and B using cholestyramine, a hydrophobic anion exchange medium. Frontal analysis in isotonic phosphate buffer was studied using post-column derivatization with o-phthalaldehyde, which gave a highly sensitive (> or =30 ng) flow-through analysis. Following load (1.5-3.0 microg toxin/3.6 mg), toxin A was bound at a slightly higher capacity than B, due to slower kinetics. A salt gradient eluted roughly 20% of bound toxin A with 0.6 M NaCl and toxin B with 1.1 M NaCl, hence toxin A showed weaker electrostatic affinity. The remainder of toxin A (65%) and some of toxin B (10% out of 50%) were eluted using a subsequent gradient to 60% acetonitrile in normal saline, which measured predominantly hydrophobic binding. Low and high affinity populations of both toxins were observed. Glycocholic acid or amino acids were competitive binders, although these components had little effect on the toxin A population bound primarily through ionic interactions. Competitive protein constituents in hamster cecal contents were also profiled. These results help to explain the variable clinical response in using cholestyramine to treat colitis. Using quaternary amine-polyhydroxymethacrylate (PHM) ion exchange chromatography, a trend for increased binding at higher pH was observed, especially for toxin A. Binding to strong cation exchange resins (sulfonate-PHM) was not observed. A range of reversed phase media retained both toxins, although recovery was very poor relative to protein standards. Size exclusion chromatography with light scattering detection showed that toxin B exists in different aggregation states, while toxin A remains monomeric. PMID- 11257521 TI - Tryptophan environment in adenosine deaminase. I. Enzyme modification with N bromosuccinimide in the presence of adenosine and EHNA analogues. AB - Adenosine deaminase from bovine cerebral hemisphere (white and gray matter) and spleen was treated with N-bromosuccinimide, a reagent known to oxidize selectively tryptophan residues in proteins. Spectrally observable tryptophan modification was accompanied by enzyme inactivation. Tsow graphics revealed that two Trps are essential for the activity of enzyme from both tissues. Enzyme inhibitors and substrate analogues, derivatives of erythro-9-(2-hydroxy-3 nonyl)adenine (EHNA) and adenosine, were able to protect Trp against modification, and this effect correlated in general with the enzyme activity protection. In the presence of adenosine deaza analogues (the noninhibitor tubercidin among them) only two Trps were modified in the fully inactivated enzyme. In the presence of EHNA and its deaza analogues, full inactivation of the enzyme was accompanied by the modification of four Trps. The obtained data confirm the previous hypothesis about the presence on the enzyme of different binding sites for adenosine and EHNA derivatives that are responsible for the different effects on the enzyme conformation elicited by the corresponding derivatives. Moreover, these data allow us to suggest that Trp residues, still unidentified by X-ray analysis, are essential for the functioning of the enzyme. PMID- 11257522 TI - Importance of the N-terminal sequence of the extrinsic 23 kDa polypeptide in Photosystem II in ion retention in oxygen evolution. AB - The function of the extrinsic 23 kDa polypeptide (OEC23) in Photosystem II (PS II) is to retain Ca(2+) and Cl(-) during the S-state turnover of the Mn cluster in photosynthetic oxygen evolution. Recombinant OEC23s from several plant species were produced in Escherichia coli to characterize the molecular mechanism of the OEC23 function then used in reconstitution experiments. One tobacco isoform, OEC23 (2AF), had much less oxygen-evolving activity than the spinach and cucumber OEC23s when PS II activities were reconstituted in salt-washed spinach PS II particles. The fact that the OEC23s had similar binding affinities for PS II particles suggests different ion-retention capacities for the individual OEC23s: The chimeric OEC23s produced between spinach and 2AF and those produced between cucumber and 2AF show that 58 N-terminal amino acid residues are important for PS II activity. Further dissection of the sequence and site-directed mutagenesis indicated the importance of 20 N-terminal amino acid residues for activity, in particular the asparagine at the 15th position. In spinach the N15D mutation decreased PS II activity, whereas in 2AF the D15N mutation increased it. This shows the importance of the N-terminal sequence of OEC23 in ion retention during the water-splitting process. PMID- 11257523 TI - Differential lipid specificities of the repeated domains of annexin IV. AB - The roles of the four domains of annexin IV in binding to phospholipids and glycolipids were assessed by analyzing the binding of a group of mutant annexins IV in which one or more of the four domains was inactivated by replacing a critical amino residue(s) (Asp or Glu) with the neutral residue Ala. The data reveal that individual annexin domains may have characteristic affinities for different lipids. In particular, inactivation of the fourth domain inhibits the binding to phosphatidylserine (PS) and phosphatidylinositol (PI) but not to phosphatidylglycerol (PG), suggesting that this domain specifically can accommodate the larger head groups of PS and PI whereas the other three domains may form more restricted binding pockets. In order to block binding to PG, domain 1, or both domains 2 and 3 must be inactivated in addition to domain 4, suggesting that all four domains may be able to accommodate the headgroup of PG to some extent. Binding to acidic glycolipids (sulfatides) was also sensitive to inactivation of domain 4. However, in the case of sulfatides the nature of the binding reaction is fundamentally different compared with the binding to phospholipids since the interaction with sulfatides was highly sensitive to an increase in ionic strength. The binding to sulfatides may depend therefore on charge-charge interactions whereas the binding to phospholipid may involve a more specific interaction between the lipid headgroup and the protein surface, and/or interaction of the protein with the hydrophobic portion of a lipid bilayer. PMID- 11257524 TI - Activation of phosphatidylinositol transfer protein alpha and beta isoforms from inclusion bodies. AB - Fully active phosphatidylinositol transfer protein (PI-TP) isoforms alpha and beta have been obtained from Escherichia coli inclusion bodies. Folding and activation of PI-TPalpha was achieved in the presence of DiC7:0 phosphatidylcholine-Triton X-114 (PtdCho-TX114) mixed micelles. Replacement of DiC7:0-PtdCho with the natural ligands of PI-TPalpha, i.e. long-chain PtdCho and phosphatidylinositol, did not stimulate activation. Efficient activation of PI TPalpha required a low temperature (4 degrees C), the presence of dithiothreitol, and was achieved at a relatively high protein concentration (i.e. up to 500 microg ml(-1)). The inclusion bodies yielded 10 mg homogeneous PI-TPalpha per liter of E. coli culture. Conditions for full activation of PI-TPbeta were similar to those for PI-TPalpha except that long-chain PtdCho-TX114 mixed micelles and a very low protein concentration (i.e. 10 microg ml(-1)) were required. In contrast to PI-TPalpha, PI-TPbeta lost its lipid transfer activity within a few days. This inactivation could be prevented by addition of beta alanine. In summary, despite 94% sequence similarity, PI-TPalpha and PI-TPbeta display a striking difference both in their preference for the PtdCho acyl chain length required for activation, and in their conformational stability after folding. PMID- 11257525 TI - Structural and immunological similarities between high molecular weight zinc ion dependent p-nitrophenylphosphatase and fructose-1,6-bisphosphate aldolase from bovine liver. AB - High molecular weight zinc ion-dependent acid p-nitrophenylphosphatase (HMW ZnAPase) was purified from bovine liver to homogeneity as judged by native and sodium dodecyl sulfate polyacrylamide gel electrophoresis. The partial sequence of the purified enzyme electroblotted on PVDF membrane reveals a 95% sequence homology with human and bovine liver fructose-1,6-bisphosphate aldolase isozyme B (FALD B). FALD B was isolated from bovine liver using an affinity elution from phosphocellulose column. FALD B from bovine liver shows a native and subunit molecular weight that is indistinguishable from that of HMW-ZnAPase. In addition, an affinity purified antiserum raised in rabbits against purified HMW-ZnAPase cross-reacts with bovine liver FALD B and rabbit muscle isozymes. Despite these similarities, HMW-ZnAPase does not show FALD activity and bovine liver FALD does not display any zinc ion-p-nitrophenylphosphatase activity. These results suggested the existence of structural and immunological similarities between bovine liver HMW-ZnAPase and FALD B. Differences in some amino acid residues in enzyme activity indicate that they may be involved in different biochemical functions. PMID- 11257526 TI - Synthesis of (di)adenosine polyphosphates by non-ribosomal peptide synthetases (NRPS). AB - In response to nutritional stress conditions, Bacillus brevis produces the cyclodecapeptide antibiotic tyrocidine via tyrocidine synthetase, a multifunctional non-ribosomal peptide synthetase. The apo-form of tyrocidine synthetase 1 forms adenosine (5')tetraphospho(5')adenosine, when incubated with MgATP(2-), amino acid and inorganic pyrophosphatase. The synthesis is an intrinsic property of the adenylation domain, is strictly dependent upon the amino acid, and proceeds from a reverse reaction of adenylate formation involving a second ATP molecule. In the presence of tri- or tetrapolyphosphate preferential synthesis of adenosine 5'-tetraphosphate and adenosine 5'-pentaphosphate occurs, respectively. A potential involvement of adenosine (5')-n-phospho(5')adenosine in the regulation of the biosynthetic process has been suggested. PMID- 11257527 TI - Kinetics of phosphoenolpyruvate carboxylase from Zea mays leaves at high concentration of substrates. AB - At low concentrations of phosphoenolpyruvate and magnesium, the substrate of phosphoenolpyruvate carboxylase (PEPC) from Zea mays leaves is the MgPEP complex and free phosphoenolpyruvate (fPEP) is an allosteric activator [A. Tovar-Mendez, R. Rodriguez-Sotres, D.M. Lopez-Valentin, R.A. Munoz-Clares, Biochem. J. 332 (1998) 633-642]. To further the understanding of this photosynthetic enzyme, we have re-investigated its kinetics covering a 500-fold range in fPEP and free Mg(2+) (fMg(2+)) concentrations. Apparent V(max) values were dependent on the concentration of the fixed free species, suggesting that these species are substrates of the PEPC-catalyzed reaction. However, when substrate inhibition was taken into account, similar V(max) values were obtained in all saturation curves for a given varied free species, indicating that MgPEP is indeed the reaction substrate. As substrate inhibition may be the result of the rise in ionic strength of the assay medium, we studied its effects on the kinetics of the enzyme. Mixed inhibition against MgPEP was found, with apparent K(ic) and K(iu) values of 36 and 1370 mM, respectively. Initial velocity patterns determined at constant ionic strength, 600 mM, were consistent with MgPEP being the true PEPC substrate, fPEP an allosteric activator, and fMg(2+) a weak, non-competitive inhibitor, thus confirming the kinetic mechanism determined previously at low concentrations of PEP and Mg(2+), and indicating that apparent substrate inhibition by MgPEP in maize leaf PEPC is caused by inhibition by high magnesium and ionic strength. PMID- 11257528 TI - Exploring the catalytic center of TaqI endonuclease: rescuing catalytic activity by double mutations and Mn2+. AB - TaqI is a metal-dependent endonuclease that recognizes T(downward arrow)CGA, with the arrow indicating the cleavage site. Mutations at K158 render the enzyme inactive and mutations at K157 significantly reduce DNA cleavage activity (W. Cao and F. Barany (1998) J. Biol. Chem. 273, 33002-33010). Aspartate, glutamate, and histidine substitutions were made at K158 in the wild-type and K157S mutant TaqI endonuclease to understand the functional organization of the active site. None of the mutants was active with Mg(2+), but the DNA cleavage activities were partly rescued by Mn2+ for K157S-K158E and K157S-K158H mutants. The rescuing effects were observed with Mn2+ but not with other divalent cations. K157S-K158E required higher Mn2+ concentrations than the wild-type enzyme for DNA cleavage activity, suggesting that a Mn2+ ion is weakly bound at the 158 position. The need to neutralize K157 to recover the catalytic activity of K158E and K158H indicates that K158 and K157 may interact functionally. In analogy with EcoRV, Ca2+ stimulated Mn2+-mediated cleavage for the wild-type TaqI, suggesting the existence of at least two metal ions at the catalytic center. A catalytic mechanism involving two metal ions and the K157-K158 pair is proposed for TaqI endonuclease. PMID- 11257531 TI - Single molecule DNA sequencing in submicrometer channels: state of the art and future prospects. AB - We demonstrate a new method for single molecule DNA sequencing which is based upon detection and identification of single fluorescently labeled mononucleotide molecules degraded from DNA-strands in a cone shaped microcapillary with an inner diameter of 0.5 microm. The DNA was attached at an optical fiber via streptavidin/biotin binding and placed approximately 50 microm in front of the detection area inside of the microcapillary. The 5'-biotinylated 218-mer model DNA sequence used in the experiments contained 6 fluorescently labeled cytosine and uridine residues, respectively, at well defined positions. The negatively charged mononucleotide molecules were released by addition of exonuclease I and moved towards the detection area by electrokinetic forces. Adsorption of mononucleotide molecules onto the capillary walls as well as the electroosmotic (EOF) flow was prevented by the use of a 3% polyvinyl pyrrolidone (PVP) matrix containing 0.1% Tween 20. For efficient excitation of the labeled mononucleotide molecules a short-pulse diode laser emitting at 638 nm with a repetition rate of 57 MHz was applied. We report on experiments where single-stranded model DNA molecules each containing 6 fluorescently labeled dCTP and dUTP residues were attached at the tip of a fiber, transferred into the microcapillary and degraded by addition of exonuclease I solution. In one experiment, the exonucleolytic cleavage of 5-6 model DNA molecules was observed. 86 photon bursts were detected (43 Cy5-dCMP and 43 MR121-dUMP) during 400 s and identified due to the characteristic fluorescence decay time of the labels of 1.43+/-0.19 ns (Cy5 dCMP), and 2.35+/-0.29 ns (MR121-dUMP). The cleavage rate of exonuclease I on single-stranded labeled DNA molecules was determined to 3-24 Hz under the applied experimental conditions. In addition, the observed burst count rate (signals/s) indicates nonprocessive behavior of exonuclease I on single-stranded labeled DNA. PMID- 11257530 TI - Data registration and selective single-molecule analysis using multi-parameter fluorescence detection. AB - A general strategy to identify and quantify sample molecules in dilute solution employing a new spectroscopic method for data registration and specific burst analysis denoted as multi-parameter fluorescence detection (MFD) was recently developed. While keeping the experimental advantage of monitoring single molecules diffusing through the microscopic open volume element of a confocal epi illuminated set-up as in experiments of fluorescence correlation spectroscopy, MFD uses pulsed excitation and time-correlated single-photon counting to simultaneously monitor the evolution of the four-dimensional fluorescence information (intensity, F; lifetime, tau; anisotropy, r; and spectral range, lambda(r)) in real time and allows for exclusion of extraneous events for subsequent analysis. In this review, the versatility of this technique in confocal fluorescence spectroscopy will be presented by identifying freely diffusing single dyes via their characteristic fluorescence properties in homogenous assays, resulting in significantly reduced misclassification probabilities. Major improvements in background suppression are demonstrated by time-gated autocorrelation analysis of fluorescence intensity traces extracted from MFD data. Finally, applications of MFD to real-time conformational dynamics studies of fluorescence labeled oligonucleotides will be presented. PMID- 11257532 TI - Fluorescently labeled model DNA sequences for exonucleolytic sequencing. AB - We describe here the enzyme-catalyzed, low-density labeling of DNAs with fluorescent dyes. Firstly, for "natural" template DNAs, dNTPs were partially substituted in the labeling reactions by the respective fluorophore-bearing analogs. The DNAs were labeled by PCR using Taq DNA polymerase. The covalent incorporation of dye-dNTPs decreased in the following order: rhodamine-green-5 dUTP (Molecular Probes, the Netherlands), tetramethylrhodamine-4-dUTP (FluoroRed, Amersham Pharmacia Biotech), Cy5-dCTP (Amersham Pharmacia Biotech). Exonucleolytic degradation by the 3'-->5' exonuclease activity of T7 DNA polymerase (wild type) in the presence of excess reduced thioredoxin proceeded to complete breakdown of the labeled DNAs. The catalytic cleavage constants determined by fluorescence correlation spectroscopy were between 0.5 and 1.5 s( 1) at 16 degrees C, normalized for the covalently incorporated dye-nucleotides. Secondly, rhodamine-green-X-dUTP (Roche Diagnostics), tetramethylrhodamine-6-dUTP (Roche Diagnostics), and Cy5-dCTP were covalently incorporated into the antisense strand of "synthetic" 218-b DNA template constructs (master sequences) at well defined positions, starting from the primer binding site, by total substitution for the naturally occurring dNTPs. The 218-b DNA constructs were labeled by PCR with a thermostable 3'-->5' exonuclease deficient mutant of the Tgo DNA polymerase which we have selected. The advantage of the special, synthetic DNA constructs as compared to natural DNAs lies in the possibility of obtaining tailor-made nucleic acids, optimized for testing the performance of exonucleolytic sequencing. The number of incorporated fluorescent nucleotides determined by complete exonucleolytic degradation and fluorescence correlation spectroscopy were six out of six possible incorporations for rhodamine-green-X dUTP and tetramethylrhodamine-6-dUTP, respectively. Their covalent and base specific incorporations were confirmed by the novel analysis methodology of re sequencing (i.e. mobility-shift gel electrophoresis, reversion-PCR and re sequencing) first developed in the paper Foldes-Papp et al. (2001) and in this paper. This methodology was then used by other groups within the whole sequencing project. PMID- 11257533 TI - Highly efficient single molecule detection in microstructures. AB - For DNA single molecule sequencing, the complete detection of all dye-labeled monomers which are cleaved off during the sequencing reaction is an essential requirement. In this work we address the feasibility of single molecule detection in microstructures with a confocal multi element set-up. We present statistical data on single molecule recognition and explain a refined data evaluation technique for single molecule burst analysis. From these data the signal-to-noise ratio in microstructures is evaluated as well as the overall detection efficiency. So far, detection efficiencies of single molecule events of up to 60% have been shown in microstructures. PMID- 11257534 TI - Fluorescent high-density labeling of DNA: error-free substitution for a normal nucleotide. AB - The enzymatic incorporation of deoxyribonucleoside triphosphates by a thermostable, 3'-->5' exonuclease deficient mutant of the Tgo DNA polymerase was studied for PCR-based high-density labeling of 217-bp "natural" DNA in which fluorescent-dUTP was substituted completely for the normal dTTP. The amplified DNA carried two different sorts of tethered dye molecules. The rhodamine-green was used for internal tagging of the DNA. Since high-density incorporation of rhodamine-green-X-dUTP led to a substantial reduction (quenching) of the rhodamine-green fluorescence, a second "high" quantum yield label, Cy5, was inserted via a 5'-tagged primer in order to identify the two-color product. A theoretical concept of fluorescence auto- and cross-correlation spectroscopy developed here was applied to quantify the DNA sequence formed in terms of both the number of two-color fluorescent molecules and the number of covalently incorporated rhodamine-green-X-dUMP residues. The novel approach allowed to separate optically the specific DNA product. After complete, exonucleolytic degradation of the two-color DNA we determined 82-88 fluorescent U* labels incorporated covalently out of 92 maximum possible U* incorporations. The heavily green-labeled DNA was then isolated by preparative mobility-shift electrophoresis, re-amplified in a subsequent PCR with normal deoxyribonucleoside triphosphates, and re-sequenced. By means of this novel methodology for analyzing base-specific incorporations that was first developed here, we found that all fluorescent nucleotides and the normal nucleotides were incorporated at the correct positions. The determined labeling efficiency of 0.89-0.96 indicated that a fraction of the substrate analog was not bearing the fluorophore. The results were used to guide developments in single-molecule DNA sequencing. The labeling strategy (principal approach) for PCR-based high-density tagging of DNA, which included an appropriate thermostable DNA polymerase and a suitable fluorescent dye-dNTP, was developed here. PMID- 11257535 TI - Towards a general procedure for sequencing single DNA molecules. AB - In this paper we report on the latest technical advances towards single molecule sequencing, a useful method currently developed especially for fast and easy de novo sequencing. Different approaches for complete labeling of DNA with fluorescent dyes are described. In addition, the experimental set-up for the sequencing process is shown. We demonstrate the ability to purify the buffer and enzyme solutions. Inorganic buffers were purified down to at least 20 fM of remaining fluorescent impurities. The exonuclease buffer solution could be cleaned down to 0.8 pM whereby its full activity was kept. Finally, we show a selection procedure for beads and present the data of a model experiment, in which immobilized DNA is degraded by an exonuclease within a polymethylmethacrylate (PMMA) microstructure. Furthermore, the mathematical processing of the obtained raw data is described. A first complete experimental cycle is shown, combining all preparatory steps which are necessary for single molecule sequencing in microstructures. PMID- 11257536 TI - Fluorescent DNA: the development of 7-deazapurine nucleoside triphosphates applicable for sequencing at the single molecule level. AB - 7-Deaza-2'-deoxyadenosine and -guanosine phosphoramidite building blocks as well as corresponding 5'-triphosphate derivatives are described carrying in position 7 substituents such as iodo, hexyn-1-yl or 5-aminopentyn-1-yl residues. The phosphoramidites were used to synthesize a series of modified oligodeoxynucleotides. A systematic study of the thermal stabilities of these oligonucleotide duplexes demonstrated that the 7-substituents are well accommodated in the major groove of B-DNA. The 7-(aminoalkyn-1-yl)-7-deazapurine 2'-deoxynucleoside triphosphates were labeled with bulky fluorophores such as Rhodamine Green(R) or tetramethylrhodamine. PMID- 11257537 TI - Polymer support for exonucleolytic sequencing. AB - Different kinds of particles were investigated for their potential use as supports for exonucleolytic sequence analysis. Composite beads composed of an unreactive polystyrene "core" and a "shell" of functionalized silica nanoparticles were found to best fulfill the various prerequisites. The biotin/streptavidin system was used for attachment of DNA to composite beads of 6 microm diameter. Applying M13 ssDNA in extremely high dilution (approximately 1 molecule versus 100 beads) with internal fluorescent labels, only a small fraction of beads was found to be associated with fluorescent entities, which likely correspond to a very small number of bound DNA molecules per particle. For better selection and transfer of DNA-containing beads into microstructures for exonuclease degradation the loading experiments were repeated with composite beads of 2.3 microm diameter. In this case a covalent bond was formed between carboxylate-functionalized beads and amino-terminated oligonucleotides, which were detected through external labelling with fluorescent nanoparticles interacting with biotinylated segments of the complementary strand. PMID- 11257538 TI - Progress towards single-molecule sequencing: enzymatic synthesis of nucleotide specifically labeled DNA. AB - The enzymatic incorporation of modified dNTPs into a growing DNA strand has intensively been studied. Modifications were detectable reporter groups such as digoxigenin or biotin, fluorochromes or aliphatic side chains covalently attached to the base. Incorporation efficiencies were determined with several DNA polymerases using linear primer-extension reactions followed by denaturing PAGE as a high-resolution detection system. We describe the enzymatic synthesis of DNA consisting of modified nucleotides exclusively. A defined template-primer system allows us to trace incorporation: (1) in up to 18 neighboring positions for several dUTP-derivatives; or (2) in stretches of DNA of up to 40 bases in length with complete substitution of all four natural dNTPs by differently modified counterparts. Synthesized DNA molecules are shown to particularly exhibit dramatically altered physico-chemical properties by contrast with native DNA. These results provide a fundamental data set for probe generation in single molecule DNA sequencing (SMS). PMID- 11257539 TI - Epigenetic regulation of carnitine metabolising enzymes in Proteus sp. under aerobic conditions. AB - Proteus sp. is able to catalyse the reversible transformation of crotonobetaine into L(-)-carnitine during aerobic growth. Contrary to other Enterobacteriaceae no reduction of crotonobetaine into gamma-butyrobetaine could be detected in the culture supernatants. Activities of L(-)-carnitine dehydratase, carnitine racemasing system and crotonobetaine reductase could be determined enzymatically in cell-free extracts of Proteus sp. Small amounts of gamma-butyrobetaine were found in cell-free extracts, indicating that it accumulates in the cell and inhibits the crotonobetaine reductase. Crotonobetaine and L(-)-carnitine were able to induce enzymes of carnitine metabolism. gamma-Butyrobetaine and glucose repress carnitine metabolism in Proteus sp. Other betaines are neither inducers nor repressors. Monoclonal antibodies against purified CaiA from Escherichia coli O44K74 recognise an analogous protein in cell-free extract of Proteus sp. No cross-reactivity could be detected with monoclonal antibodies against purified CaiB and CaiD from E. coli O44K74. PMID- 11257540 TI - Restricted growth of ent(-) and tonB mutants of Salmonella enterica serovar Typhi in human Mono Mac 6 monocytic cells. AB - Monocytes and macrophages are an important host defense in humans infected with Salmonella enterica serovar Typhi. Bacterial ability to survive in these cells is therefore a crucial virulence characteristic of this pathogen. In this study, we demonstrate that growth of a Salmonella enterica serovar Typhi enterochelin synthesis mutant and a tonB mutant in the human monocyte cell line Mono Mac 6 is restricted compared to that of the parental wild-type Ty2 strain. These results suggest that enterochelin- and TonB-mediated iron uptake plays a role in S. enterica serovar Typhi pathogenesis, and also suggest that mutations in iron uptake may attenuate S. enterica serovar Typhi strains for human beings. PMID- 11257541 TI - A mutant of the cyanobacterium Anabaena variabilis ATCC 29413 lacking cyanophycin synthetase: growth properties and ultrastructural aspects. AB - The gene cphA encoding cyanophycin synthetase was interrupted in Anabaena variabilis ATCC 29413 by insertional mutagenesis. The mutant lacked cyanophycin granules and the polar nodules of heterocysts. The mutant grew as fast as the wild-type irrespective of the nitrogen source at low light intensity whereas growth on N(2) was somewhat reduced in high light. It is concluded that cyanophycin metabolism and polar nodules are not essential for aerobic N(2) fixation. PMID- 11257542 TI - The immunological dissection of the Escherichia coli molecular chaperone DnaJ. AB - In Escherichia coli, one of the main molecular chaperones is DnaJ (hsp40), which mediates in a variety of highly conserved cellular processes including protein folding reactions and the assembly/disassembly of protein complexes. DnaJ is characterised by the presence of four distinct domains: the J-domain, glycine/phenylalanine-rich (G/F), cysteine-rich (Zn-finger) and C-terminal regions. Truncated DnaJ polypeptides (DnaJ 1-108, DnaJ Delta1-108, DnaJ Delta1 199) representing these domains were over-produced and used as a source of immunogens for the generation of sequence-specific polyclonal antibodies. Epitope mapping was achieved by Western blotting, which demonstrated the presence of antibodies directed against these domains. These characterised affinity-purified antibodies were then used to assess the role of DnaJ in the protection of firefly luciferase from irreversible heat-inactivation. In this study we have demonstrated the involvement of J-, G/F and Zn-finger domains in the protection of luciferase from heat-inactivation. The C-terminal region had only partial involvement in luciferase protection. PMID- 11257543 TI - Role of SspA in the density-dependent expression of the transcriptional activator AarP in Providencia stuartii. AB - The AarP protein in Providencia stuartii encodes a small transcriptional activator which activates the chromosomal aminoglycoside acetyltransferase aac(2')-Ia gene. In addition, AarP activates genes involved in a multiple antibiotic resistance (Mar) phenotype. Expression of an aarP-lacZ fusion increased in a density-dependent manner and reached peak levels at stationary phase. The expression of an aarP-lacZ fusion could be prematurely activated in cells at early to mid-exponential phase by the addition of spent culture supernatants from stationary phase cultures or by ethyl acetate extracts of these supernatants. Nutrient starvation had a negligible effect on aarP expression. In a search for mutations that block aarP activation at stationary phase, a mini Tn5Cm insertion has been identified within a gene whose product was 77% identical to SspA, a regulatory protein involved in stationary phase gene expression and virulence. An unmarked sspA null allele (sspA2) was created by allelic replacement to further examine the role of sspA in P. stuartii. The sspA2 allele resulted in substantial decrease in aarP mRNA accumulation at various phases of growth. Furthermore, in an sspA mutant background, the aarP-lacZ fusion was no longer activated by an extracellular signal. PMID- 11257544 TI - Isolation and diagnostic potential of ISMav2, a novel insertion sequence-like element from Mycobacterium avium subspecies paratuberculosis. AB - A novel Mycobacterium avium ssp. paratuberculosis (M. paratuberculosis) specific insertion sequence has been identified by representational difference analysis and designated as ISMav2. ISMav2 has no similarity to known mycobacterial IS elements but shows more than 50% identity to a non-composite transposon of Streptomyces coelicolor at the DNA and protein level. ISMav2 is present in at least three copies on the genome as assessed by Southern blot analysis and its potential value as a diagnostic tool was confirmed by PCR analyses on 79 M. paratuberculosis field isolates, nine M. avium ssp. avium isolates, and the reference strains of nine other mycobacterial species. PMID- 11257545 TI - Effects of N(G)-nitro-L-arginine methyl ester on endotoxin-induced leakage of lactate dehydrogenase and cytotoxicity in J774A.1 cells. AB - In the present study, we investigated the effects of the nitric oxide (NO) synthase inhibitor N(G)-nitro-L-arginine-methyl ester (L-NAME) on tissue injury or cytotoxicity caused by endotoxin challenge by assaying lactate dehydrogenase (LDH) isozymes and cell viability in J774A.1 cells. In mice treated with L-NAME (10 mg kg(-1), i.v.), the activity of LDH in serum 18 h after endotoxin (6 mg kg( 1), i.p.) injection was not significantly different from that in mice treated with endotoxin alone. Mice injected with endotoxin exhibited leakage of LDH isozymes 3 and 5, but L-NAME did not protect against endotoxin-induced acute leakage of LDH isozymes. Treatment with L-NAME (10-1000 microM) significantly inhibited NO generation by endotoxin (1 microg ml(-1))-activated J774A.1 cells. However, L-NAME (10-1000 microM) did not affect endotoxin-induced cytotoxicity in J774A.1 cells. These findings suggested that endotoxin-induced NO formation may not contribute to tissue injury or cytotoxicity caused by endotoxin. PMID- 11257546 TI - Evidence of calcium influx across the plasma membrane depends upon the initial rise of cytosolic calcium with activation of IP(3) in rat enterocytes by heat stable enterotoxin of Vibrio cholerae non-O1. AB - In response to heat-stable enterotoxin of Vibrio cholerae non-O1, the initial rise of cytosolic Ca(2+) occurred with activation of IP(3). Chelation of extracellular Ca(2+) with EGTA and suspension of cells in Ca(2+) free buffer both demonstrated the involvement of internal stores in the rise of [Ca(2+)]i. Cells pretreated with dantrolene resulted in decrease of [Ca(2+)]i response which suggested that the rise of intracellular level of Ca(2+) was mostly due to the mobilization from IP(3) sensitive stores. When the cytosolic Ca(2+) was chelated by loading the cells with BAPTA, NAG-ST could not induce Ca(2+) entry to the cell as assessed by Mn(2+) quenching of fura-2 fluorescence which suggested that calcium influx across the plasma membrane depends upon initial rise of this bivalent cation that maintained the sustained phase of [Ca(2+)]i response. Addition of toxin to the fura-2-loaded cells, preincubated with lanthanum chloride, resulted in reduction of [Ca(2+)]i level with a short duration of irregular sustained phase further suggesting that the influx of Ca(2+) across the plasma membrane might be through the calcium channel. PMID- 11257547 TI - Molecular and biochemical characterisation of Mycobacterium smegmatis alcohol dehydrogenase C. AB - The gene encoding of an alcohol dehydrogenase C (ADHC) from Mycobacterium smegmatis was cloned and sequenced. The protein encoded by this gene has 78% identity with Mycobacterium tuberculosis and Mycobacterium bovis BCG ADHC. The M. smegmatis ADHC was purified from M. smegmatis and the kinetic parameters of this enzyme showed that using NADPH as electron donor it has a strong preference for aliphatic and aromatic aldehyde substrates. Like the M. bovis BCG ADHC, this enzyme is more likely to act as an aldehyde reductase than as an alcohol dehydrogenase. The discovery of such an ADHC in a fast-growing, and easily engineered mycobacterial species opens the way to the utilisation of this M. smegmatis enzyme as a convenient model for the study of the physiological role of this alcohol dehydrogenase in mycobacteria. PMID- 11257548 TI - Yersinia HPI in septicemic Escherichia coli strains isolated from diverse hosts. AB - High pathogenicity islands (HPIs), first identified in various Yersinia species, encode an iron uptake system. We have studied the occurrence of HPIs in septicemic strains of Escherichia coli isolated from a variety of hosts. The results presented in this communication indicate that most septicemic strains tested contained HPI sequences even though they already have the aerobactin encoding genes. We have also observed two types of HPI deletions, suggesting genetic instability of this element. Notable exceptions are several strains isolated from septicemia in sheep that lacked both iron acquisition systems. PMID- 11257549 TI - Incidence of the cblA major subunit pilin gene amongst Burkholderia species. AB - PCR-amplification has been used to screen 75 isolates of the Burkholderia cepacia complex for the cblA pilin gene. PCR-amplified products of the correct size (664 bp) were cloned and sequenced and the sequences compared. Apart from in the control, epidemic cystic fibrosis (CF)-associated B. cepacia lineage we also identified, for the first time, cblA genes in a unique, non-CF clinical isolate from France and a plant (onion) pathogenic isolate from Italy. The sequence of the cblA gene amplified from the clinical isolate was more diverged from the epidemic lineage than that amplified from the onion pathogenic isolate. PMID- 11257550 TI - Stable carbon isotope fractionations of the hyperthermophilic crenarchaeon Metallosphaera sedula. AB - The stable carbon isotopic compositions of the inorganic carbon source, bulk cell material, and isoprenoid lipids of the hyperthermophilic crenarchaeon Metallosphaera sedula, which uses a 3-hydroxypropionate-like pathway for autotrophic carbon fixation, have been measured. Bulk cell material was approximately 3 per thousand enriched in 13C relative to the dissolved inorganic carbon, and 2 per thousand depleted in 13C relative to isoprenoid membrane lipids. The isotope data suggested that M. sedula uses mainly bicarbonate rather than CO(2) as inorganic carbon source, which is in accordance with a 3 hydroxypropionate-like carbon fixation pathway. To the best of our knowledge this is the first report of 13C fractionation effects of such a hyperthermophilic crenarchaeon. PMID- 11257551 TI - Comparative study on the selective chalcopyrite bioleaching of a molybdenite concentrate with mesophilic and thermophilic bacteria. AB - This study evaluates different bioleaching treatments of a molybdenite concentrate using mesophilic and thermophilic bacterial cultures. Further studies on the chemical leaching and the electrochemical behavior of the MoS(2) concentrate were carried out. Bioleaching tests showed a progressive removal of chalcopyrite from the molybdenite concentrate with an increase in temperature. Chemical leaching tests support the idea of an indirect attack of the concentrate. Electrochemical tests indicate that chalcopyrite dissolution is favored when molybdenite is present. Therefore, this type of bioleaching treatment could be applied to purify molybdenite flotation concentrates by selectively dissolving chalcopyrite. PMID- 11257552 TI - Synthesis of the Streptomyces lividans maltodextrin ABC transporter depends on the presence of the regulator MalR. AB - During growth with maltotriose or amylose, Streptomyces lividans and Streptomyces coelicolor A3(2) synthesize a maltodextrin uptake system with highest specificity for maltotriose. The transport activity is absent in mutants of S. coelicolor A3(2) lacking a functional MalE binding protein. Cloning and sequencing data suggest that the mal operon of S. coelicolor A3(2) corresponds to the one of S. lividans and that the deduced S. lividans Reg1 amino acid sequence is identical to that of MalR from S. coelicolor A3(2). It can be concluded that both strains have the same ABC transport system for maltodextrins. The S. lividans malR was cloned in Escherichia coli in frame with six histidine-encoding codons. The resulting, purified 6HisMalR(SI) was shown to bind to two motifs within the S. lividans malR-malE intergenic region and to dissociate in the presence of maltopentaose. PMID- 11257554 TI - Nutritional assessment of the lung transplant patient: body mass index as a predictor of 90-day mortality following transplantation. AB - BACKGROUND: It is well documented that malnourished and/or obese surgical patients have increased morbidity and mortality post-operatively. Only a few studies investigating the effect of nutritional status on mortality are available pertaining to the transplant population. Since limited data are available on the nutritional status and its effects on mortality in the lung transplant population, we sought to ascertain whether there is an association between mortality and preoperative nutritional status. METHODS: We examined mortality during the first 3 months after transplantation. Patients were grouped by body mass index (BMI) categories as < 17 kg/m(2), 17 to < 20 kg/m(2), 20 to 25 kg/m(2) (reference group), > 25 to 27 kg/m(2), and > 27 kg/m(2). Additional risk factors retrieved from the pre-transplant records included age, gender, diagnosis, energy requirements, protein requirements, protein and caloric intake, and weight history. Logistic regression for univariate and multivariate analysis for mortality used recipient age, gender, disease category, pre-transplant cytomegalovirus (CMV) serology, transplant type (single or bilateral), and donor age, gender, and CMV serology. RESULTS: The likelihood estimates or odds ratios (ORs) of the risk of death within 90 days of lung transplantation for the BMI categories compared to the reference group were 3.7 for BMI < 17 kg/m(2) (p = 0.085), 1.6 for BMI < 17 to 20 kg/m(2) (p = 0.455), 3.5 for BMI > 25 to 27 kg/m(2) (p = 0.069), and 5.0 for BMI > 27 kg/m(2) (p = 0.003). CONCLUSIONS: In patients with a pre-transplant BMI < 17 kg/m(2) or > 25 kg/m(2) the risk of dying within 90 days post-transplant was increased. In patients with a pre-transplant BMI of > 27 kg/m(2) the risk was significantly higher in than the reference group. PMID- 11257553 TI - Death after rejection with severe hemodynamic compromise in pediatric heart transplant recipients: a multi-institutional study. AB - BACKGROUND: Rejection with severe hemodynamic compromise results in high mortality in adult transplant patients. This study determines the incidence, outcome and risk factors for rejection with severe hemodynamic compromise in a multi-institutional study of pediatric heart transplant recipients. METHODS: Data from 847 patients transplanted between 1/1/93 and 12/31/98 at 18 centers in the Pediatric Heart Transplant Study were analyzed. Rejection with severe hemodynamic compromise was defined as a clinical event occurring beyond 1 week postoperatively, which led to augmentation of immunosuppression and use of inotropic therapy. Actuarial freedom from such rejection and death after rejection were determined and risk factors sought. RESULTS: Among 1,033 rejection episodes in 532 patients, 113 (11%) episodes were associated with severe hemodynamic compromise in 95 patients. The highest risk for severe rejection was in the first year. Risk factors were older recipient age (p >.05) and non-white race (p >.001). Survival after an episode was poor (60%), and biopsy score did not affect outcome. Deaths were due to rejection (n = 14), other cardiac causes (n = 17), infection (n = 5), lymphoma (n = 2), pulmonary causes (n = 2), and thrombosis (n = 1). CONCLUSIONS: Rejection with severe hemodynamic compromise occurs in 11% of pediatric patients, irrespective of age, sex or biopsy score, and mortality is high. Non-white race and older recipient age are independent risk factors for rejection with severe hemodynamic compromise. Aggressive treatment and close surveillance should be crucial components of protocols aimed at reducing the high mortality. PMID- 11257555 TI - Cardiopulmonary transplantation for congenital heart disease in the adult. AB - BACKGROUND: Patients surviving into adulthood with congenital heart disease (CHD) often succumb to progressive cardiopulmonary dysfunction. For these patients transplantation is often considered. METHODS: We performed a retrospective review of 69 adults (age >18 years) with CHD transplanted between 1984 and 1999. RESULTS: We evaluated 31 heart-lung (HLTxp), 30 lung (LTxp), and 8 heart (HTxp) transplants performed in 22 men and 47 women with CHD. Mean age was 37 +/- 10 years with a mean follow-up of 3.1 +/- 3.5 years. A concomitant cardiovascular procedure was performed in 1 HLTxp, 23 LTxp, and 2 HTxp. Early mortality (>30 days) was 26% (8/31) for HLTxp, mostly due to bleeding. Early LTxp mortality was 23% (7/30), largely due to graft failure. One and 3-year survival was similar in adults transplanted for CHD and adults transplanted for other disease. Early mortality among HTxp recipients was 50% (4/8) from rejection or technical complications. Survival for patients undergoing HLTxp versus LTxp with cardiac repair was similar. When examined by era, the survival of patients transplanted for CHD between 1992 and 1999 was greater than that of patients transplanted between 1984 and 1991. CONCLUSIONS: Adults undergoing HLTxp and LTxp for CHD can expect survival comparable to that of non-CHD adults. In the presence of a reparable cardiac lesion, LTxp with cardiovascular repair for CHD is an attractive option, optimizing organ allocation. Specific technical concerns are discussed. Survival of adults undergoing cardiopulmonary transplantation for CHD has improved over time. PMID- 11257556 TI - Pre-transplant corticosteroid use and outcome in lung transplantation. AB - BACKGROUND: The early experience of lung transplantation was plagued with airway anastomotic complications. The use of corticosteroids in the pre-transplant period has been implicated as a major contributing factor in bronchial dehiscence, and many patients have been denied transplantation on the basis of corticosteroid use. We conducted the current study to assess the risks associated with pre-transplant corticosteroid use. METHODS: We analyzed records of 73 single and bilateral-single lung transplant recipients who had chronic obstructive pulmonary disease or alpha(1)-antitrypsin deficiency as their underlying disease from 1986 to 1996. Twenty-six patients (steroid group) received daily corticosteroid therapy (prednisone, 1.5 to 40 mg/day) up to the time of transplantation, whereas 47 patients did not receive chronic corticosteroids and had no corticosteroid therapy within 3 months of transplantation (non-steroid group). RESULTS: The demographic profiles of the 2 groups were comparable. We noted no statistical significances in length of hospital stay, duration of intensive care, and post-operative pulmonary function. The rates of cytomegalovirus infection, acute rejection, bronchiolitis obliterans syndrome, and survival were also similar. The non-steroid group seemed to have a higher rate of bronchial stenosis at 3 years (29% vs 6%, p = 0.03). Bronchial dehiscence did not occur in either study group. CONCLUSIONS: Pre-transplant use of corticosteroids does not adversely affect outcome following lung transplantation. PMID- 11257557 TI - Effective treatment of hyperhomocysteinemia in heart transplant recipients with and without renal failure. AB - BACKGROUND: Elevated total plasma homocysteine (tHcy) levels have been associated with vascular disease and higher mortality in patients with coronary artery disease. Graft coronary disease is a major cause of mortality in long-term survivors of heart transplantation, and hyperhomocysteinemia may be one of its causes. The objectives of our study were to establish the effectiveness of a 3 stage homocysteine-lowering algorithm in a group of 84 heart transplant (HTx) patients and to evaluate the effect of renal function on the response to homocysteine-lowering therapy. METHODS: Prospective treatment of 84 Htx patients (64 male; mean age, 48 +/- 13 years) with tHcy > 75th percentile consisted of a 3 stage treatment algorithm: Stage 1, folic acid (FA) 2 mg + vitamin (vit) B(12) 500 mcg daily; Stage 2, addition of vit B(6) 100 mg daily; Stage 3, increase FA to 15 mg daily. Serum creatinine (Cr) and tHcy levels were measured before treatment and 21 +/- 19 weeks after each stage of treatment. RESULTS: All 3 stages of treatment significantly lowered mean tHcy from 22.4 +/- 16.3 (mean +/- SD) micromol/liter to 16.3 +/- 6.7 micromol/liter (p < 0.00001), from 17.6 +/- 6.1 micromol/liter to 15.2 +/- 5.3 micromol/liter (p < 0.0001), and from 16.8 +/- 5.2 micromol/liter to 15.6 +/- 5.3 micromol/liter (p < 0.05), respectively. The average reduction from baseline was 38%. Creatinine levels did not change significantly during the study period. Total plasma homocysteine levels decreased below the 75th percentile in 55% of patients, with Cr levels significantly lower in this group of patients (126 +/- 36 micromol/liter vs 182 +/- 65 micromol/liter, p < 0.00001). However, we found no significant relationship between % change in tHcy and baseline Cr. CONCLUSIONS: In a group of 84 heart transplant patients with tHcy levels >75th percentile, treatment with FA and vit B(6) and B(12) according to a 3-stage algorithm resulted in statistically significant declines in mean tHcy levels. Overall, tHcy levels decreased 38%, with target tHcy levels <75th percentile achieved in 55% of the patients. The % change in tHcy was not related to Cr. Further studies are needed to correlate treatment of hyperhomocysteinemia with clinical endpoints, such as the time to development of transplant vasculopathy and long-term survival, and to define the most appropriate targets for therapy. PMID- 11257558 TI - Plasmapheresis and cyclophosphamide in the treatment of humoral rejection after heart transplantation. AB - BACKGROUND: Clinical reports on humoral rejection after heart transplantation showed that these episodes were often more severe than those mediated through T lymphocytes and that the patient's prognosis was significantly worsened. METHODS: To evaluate the impact of plasmapheresis on the course of humoral rejection with hemodynamic compromise (HRHC) episodes, we retrospectively investigated the records of 1,108 heart transplant patients. All patients received triple-drug immunosuppression (cyclosporine a, azathioprine, prednisone) and cytolytic antibodies for induction. Between April 1986 and December 1990, HRHC episodes were treated with cortisone boli and cytolytic antibodies for at least 3 days (Group A). Between January 1991 and April 1999, HRHC episodes were treated with cortisone boli, cytolytic antibodies, and plasmapheresis for at least 3 days (Group B). All patients who survived their first HRHC episode received cyclophosphamide instead of azathioprine as maintenance immunosuppression. RESULTS: Altogether we observed 29 HRHC episodes. In 11 cases, no therapy could be administered or the therapy regimen did not correspond to either Protocol A or B. In the remaining 18 HRHC episodes, 7 episodes in 7 patients were treated without plasmapheresis (Group A), but only 2 patients survived, whereas in 11 HRHC episodes in 6 patients, therapy included plasmapheresis (Group B) and all patients survived (p = 0.002). Four of 6 patients who received cyclophosphamide after their first HRHC episode experienced at least 1 further HRHC episode. CONCLUSIONS: Plasmapheresis seems to improve outcomes in HRHC. However, cyclophosphamide as a maintenance immunosuppressive drug failed to prevent further humoral rejection episodes. PMID- 11257559 TI - Effects of human tissue plasminogen gene transfer on allograft coronary atherosclerosis. AB - BACKGROUND: Transplant coronary atherosclerosis is a major limiting factor to successful long-term cardiac transplantation. The depletion of tissue plasminogen activator (tPA) in the arteriolar smooth muscle cells has been associated with a higher incidence of accelerated graft atherosclerosis. In vivo overexpression of tPA may inhibit accelerated graft atherosclerosis and improve the long-term results of heart transplantation. We evaluated the feasibility, distribution, and effects of intracoronary transfer of the human tPA (htPA) gene in a rabbit heterotopic cardiac transplant model, using a novel cationic liposome compound designed for improved delivery to vascular endothelium. METHODS: Human tPA cDNA under the control of the SV40 promoter (100 microg) was complexed with the novel cationic liposome (+/-)-N-(3-aminopropyl)-N,N-dimethyl-2,3-bis(dodecyloxy)-1 propanaminium bromide (GAP: DLRIE) (50 microg), and delivered ex vivo to the donor heart by slow intracoronary infusion. Control hearts received an "empty" liposome preparation. Grafts were then implanted into recipient rabbits in the heterotopic cervical position. For the analysis of gene expression, beating donor hearts were collected at 4 days. To examine the effects of htPA expression on graft atherosclerosis, animals received a 0.5% cholesterol diet for 30 days posttransplant, as well as 10 mg/kg cyclosporine A daily. Beating hearts were collected at 30 days posttransplant and analyzed for the development of transplant atherosclerosis by image analysis. RESULTS: Northern blot analysis for the htPA messenger RNA (mRNA) transcripts showed significantly higher counts in hearts receiving the htPA gene as compared to controls. The distribution of these transcripts favored the left ventricle (LV) and septal regions over the right ventricle (RV). Scintillation analysis of specimens stained by immunoflourescence showed expression of htPA throughout the perivascular myocardium that was significantly higher in grafts transduced with the htPA gene than in control or native hearts. Expression in the vascular wall was also significantly enhanced. Scintillation counts per x 200 field were 262 +/- 145 in htPA-transduced hearts and 20 +/- 27 in controls (p = 0.001), and mean luminescence was 83.7 +/- 12.5 in htPA-transduced hearts and 62.9 +/- 12.8 in controls (p = 0.01). Intimal hyperplasia was assessed by mean percent luminal stenosis in small- and medium sized arteries and was 31.12 +/- 23.5% in htPA-transduced hearts and 86.59 +/- 17.5% in control hearts (p < 0.0001). These results demonstrate that expression of the htPA gene can be induced by ex vivo intracoronary gene transfer at the time of allograft preservation. Liposome-mediated delivery of the htPA gene at the time of transplantation results in significant early transgene expression, and significantly inhibits the development of graft coronary atherosclerosis. PMID- 11257560 TI - RAD in stable lung and heart/lung transplant recipients: safety, tolerability, pharmacokinetics, and impact of cystic fibrosis. AB - BACKGROUND: RAD is a novel macrolide with potent immunosuppressive and antiproliferative activities. This study characterizes the safety, tolerability, and pharmacokinetics of two different single oral doses of RAD in stable lung and heart/lung transplant recipients with and without cystic fibrosis (CF). METHODS: This was a Phase I, multicenter, randomized, double-blind, two-period, two sequence, crossover study. Single doses of RAD capsules at doses of 0.035 mg/kg (2.5 mg maximum) or 0.10 mg/kg (7.5 mg maximum) were administered with cyclosporine (Neoral [cyclosporine, USP] modified), steroids, and azathioprine on Day 1. The alternate dose was administered on Day 16. Laboratory assessments, vital signs, and adverse events were recorded throughout the study. RAD pharmacokinetic profiles were assessed over a 7-day period following each dose. Steady-state cyclosporine (CsA) profiles were assessed at baseline and with each RAD dose; RAD and CsA trough concentrations were obtained throughout the study period. RESULTS: Of the 20 patients randomized, 8 had CF and 12 did not. Single doses of RAD were safe and well tolerated. Headache was the most common side effect. RAD produced a mild, dose-dependent, reversible decrease in platelet and leukocyte counts. Cholesterol and triglycerides were minimally affected. At both doses, CF patients had significantly lower peak concentrations of RAD than did non-CF patients (p = 0.03); however, overall exposure (area under the curve/dose) was not different between the groups (p = 0.63). At the higher dose, there was a clinically minor under-proportionality in AUC, averaging -11%. Steady-state pharmacokinetics of CsA were not affected by RAD co-administration.RAD was safe and well tolerated by stable lung and heart/lung transplant recipients with and without CF. The presence of CF did not influence the extent of RAD exposure. Single doses of RAD did not affect the pharmacokinetics of CsA. Ongoing studies are assessing the long-term safety and efficacy of RAD in lung and heart/lung transplantation. PMID- 11257562 TI - Myocardial dysfunction associated with brain death: clinical, echocardiographic, and pathologic features. AB - BACKGROUND: The sequelae of severe brain injury include myocardial dysfunction. We sought to describe the prevalence and characteristics of myocardial dysfunction seen in the context of brain-injury-related brain death and to compare these abnormalities with myocardial pathologic changes. METHODS: We examined the clinical course, electrocardiograms, head computed tomography scans, and echocardiographic data of 66 consecutive patients with brain death who were evaluated as heart donors. In a sub-group of patients, we compared echocardiographic findings with pathologic findings. RESULTS: Echocardiographic systolic myocardial dysfunction was present in 28 (42%) of 66 patients and was not predicted by clinical, electrocardiographic, or head computed tomographic scan characteristics. Ventricular arrhythmias were more common in the patients with, compared to those without, myocardial dysfunction (32% vs 0%; p < 0.001). Myocardial dysfunction was segmental in all 8 patients with spontaneous subarachnoid or intracerebral hemorrhage. In these patients, the left ventricular apex was often spared. Myocardial dysfunction was either segmental or global in 17 patients who suffered head trauma and in 3 patients who died of other central nervous system illnesses. In 11 autopsied hearts, we found poor correlation between echocardiographic dysfunction and pathologic findings. CONCLUSIONS: Systolic myocardial dysfunction is common after brain-injury-related brain death. After spontaneous subarachnoid or intracerebral hemorrhage, the pattern of dysfunction is segmental, whereas after head trauma, it may be either segmental or global. We found poor correlation between the echocardiographic distribution of dysfunction and light microscopic pathologic findings. PMID- 11257561 TI - Acute rejection and cardiac graft vasculopathy in the absence of donor-derived ICAM-1 or P-selectin. AB - BACKGROUND: ICAM-1 and P-selectin are molecules that facilitate adhesion of circulating leukocytes to vessel walls. We have investigated the role of donor derived ICAM-1 and P-selectin in acute and chronic cardiac allograft rejection. METHODS: C57BL/6J (H-2(b)) mice were used as donors for heterotopic heart transplantation into CBA/Ca (H-2(k)) recipients. The donors were wild-type or homozygous for gene mutations of ICAM-1 or P-selectin. We measured acute rejection in non-immunosuppressed recipients by daily palpation and sacrificed mice at Days 2, 4, and 6 for immunohistochemical analysis. For chronic rejection, recipients received monoclonal antibody against CD4+ T cells. We removed hearts at Days 60 to 62 for histologic assessment of vasculopathy using quantitative morphometry to measure intimal thickening. RESULTS: Time (days) to rejection was 7.1 +/- 0.57 for wild-type (n = 10), 7.0 +/- 0.71 for ICAM-1 -/- (not significantly different, n = 7) and 6.1 +/- 0.33 (p = 0.001) for P-selectin -/- donors. ICAM-1 deficiency was associated with delayed infiltrate at Day 4 compared with wild-type. In the model of chronic rejection, elastin-positive vessels showed a mean occlusion of 34% +/- 3% in transplanted wild-type hearts; vessels were divided into those showing 0% to 20%, 20% to 50%, and 50% to 100% occlusion. We observed no difference in the number of affected vessels or the amount of vascular thickening in donors lacking ICAM-1 or P-selectin compared with wild-type controls. CONCLUSIONS: The absence of ICAM-1 or P-selectin in donor tissues neither lengthens the time of allograft survival nor inhibits the vascular lesions associated with chronic rejection. Indeed, the absence of P selectin may exacerbate alloimmune injury. PMID- 11257564 TI - Experimental heterotopic heart transplantation without cardiopulmonary bypass: auxiliary support for the recipient heart. AB - BACKGROUND: Auxiliary cardiac support using heterotopic heart transplant is of considerable interest, but the outcome is not known. To investigate technical feasibility and the possibility of using auxiliary support from heterotopic heart transplantation without cardiopulmonary bypass, we evaluated hemodynamics including the pressure-volume relationship in experimental animals. METHODS: In heterotopic heart transplantation, we tailored the donor heart by removing the pulmonary and tricuspid valves, and by wide removal of the inter-atrial septum. Next, we anastomosed the descending aorta and left atrium of the donor heart to the descending aorta and left atrium of the recipient, without using cardiopulmonary bypass. Consequently, declamping the recipient's descending aorta allowed the donor heart to fill with blood and to start beating. We performed hemodynamic assessments including the effects of adrenergic stimulation. We measured the pressure and volume relationship of the recipient heart by closing and opening inflow of the donor left atrium to change the pre-load of the donor left ventricle. RESULTS: The donor left ventricle produced a systolic blood pressure that was augmented by the recipient blood pressure and responded to adrenergic stimulation. When inflow of the donor left atrium was opened, the pressure-volume loop of the recipient heart shifted to the left and pressure volume area decreased. Simultaneously, the mechanical efficiency and E(max) (the slope of the end-systolic pressure-volume relationship) of the recipient heart increased when inflow of the donor left atrium was opened. CONCLUSIONS: This transplant model, without cardiopulmonary bypass, is feasible and can be applied to transplant investigations as a working heart model on the basis of the response of adrenergic stimulation. The increased pre-load of the donor left atrium from the recipient left atrium resulted in a recipient leftward shift of the pressure-volume relationship, suggesting that this transplant model with adequate pre-load acts as auxiliary assistance in the recipient intrathoracic cavity. PMID- 11257563 TI - Impact of PAF antagonist BN 52021 (Ginkolide B) on post-ischemic graft function in clinical lung transplantation. AB - BACKGROUND: Platelet activating factor (PAF) is associated with ischemia/reperfusion injury (I/R) after lung transplantation. Following promising experimental results, this prospective trial investigated the potential effect of PAF antagonist BN 52021 (ginkolide B) on clinical Euro-Collins (EC)-based lung preservation. METHODS: We analyzed 8 double-lung transplant patients in each of 3 groups. In the low-dose group (LDG), donor lungs were perfused with EC containing 2 mg/kg BN 52021, whereas we used 10 mg/kg in the high-dose group (HDG) and placebo in the control group (CG). Before reperfusing the first lung, we administered intravenously 120 mg BN 52021 (LDG), 600 mg BN 52021 (HDG), or placebo (CG). Hemodynamics in terms of pulmonary arterial pressure, pulmonary vascular resistance and serial determinations of the alveolo-arterial oxygen difference (AaDO(2)) were recorded. We measured blood levels of PAF pre operatively and post-operatively, after 10 minutes and after 3, 8, 24, 48, and 144 hours. RESULTS: Within 32 hours, we noted a tendency toward better AaDO(2) in the LDG and the HDG compared with the CG (p > 0.05). We observed a significant improvement of AaDO(2) after 3 hours (HDG, p = 0.033) and 8 hours (LDG, p = 0.024), with poorest values in the CG. The PAF concentrations were lowest in the HDG, with significant deterioration 10 minutes after reperfusion. In contrast, placebo led to higher PAF levels. We measured significantly lower PAF concentrations (HDG vs CG) at 10 minutes and at 6 days post-operatively. CONCLUSIONS: Use of high-dose PAF antagonist BN 52021 can easily be combined with clinical preservation methods and may help optimize pulmonary function with reduced PAF levels, in the early post-ischemic period. PMID- 11257565 TI - A retrospective assessment of safety, efficacy, and pharmacoeconomics of generic azathioprine in heart-transplant recipients. AB - Although a generic formulation of azathioprine (AZA) has been available since 1996, safety, efficacy and pharmacoeconomic implications following conversion from Imuran (AZA) to generic AZA in heart-transplant patients remains to be determined. A retrospective, safety and efficacy assessment, in addition to a cost comparison, was performed in 30 heart-transplant patients who had been switched from Imuran to generic AZA. In heart-transplant patients converted from Imuran to generic AZA, no compromise in safety and efficacy, as measured by white blood cell (WBC) count, infections, rejections, malignancies, and hospitalizations was observed. Generic substitution of Imuran results in an annual cost savings of $318 per patient. PMID- 11257566 TI - Recurrence of giant cell myocarditis in cardiac allograft. AB - BACKGROUND: Giant cell myocarditis causes essentially irreversible fulminant left ventricular dysfunction with associated conduction abnormalities and congestive failure. Response to immunosuppressive therapy is poor and cardiac transplantation is the only viable treatment option. The histologic hallmarks of giant cell myocarditis include a polymorphous inflammatory response with numerous multinucleated giant cells and extensive myocyte necrosis in a geographic pattern. There were 38 patients who received a cardiac transplant for giant cell myocarditis in the Giant Cell Myocarditis Registry. Among these patients, there were 9 recurrences of disease in the allograft. Concern has been expressed that recurrence of giant cell myocarditis in the allograft might be a contraindication for cardiac transplantation in the future. METHODS: In our single-center analysis we describe the clinical and histologic findings of 5 patients transplanted for giant cell myocarditis at the Cleveland Clinic. RESULTS: All but 1 of the patients were New York Heart Association (NYHA) class 4 with an average cardiac index (CI) of 1.52 liters/min x m(2). Of the 5 patients transplanted, 1 developed recurrent giant cell myocarditis. Routine right ventricular endomyocardial biopsy at 1 week exhibited severe multifocal myocardial fibrosis in addition to mild acute vascular rejection and mild grade 1A cellular rejection. Follow-up biopsy in this patient indicated grade IIIA moderate acute rejection in addition to multinucleated giant cells. Two distinct inflammatory processes were noted consisting of foci of T-cell inflammation identified by immunohistochemistry to be consistent with rejection, and a second inflammatory process with few mononuclear cells staining for macrophage or T-cell markers with eosinophils and myocyte necrosis consistent with giant cell myocarditis. Follow-up right ventricular endomyocardial biopsies (RVBXs) in this patient have subsequently demonstrated improvement in the degree of inflammatory infiltrate without vascular or significant cellular rejection. Vascular rejection was noted in 1 of the remaining 4 patients and was treated successfully with muramab-CD3 and plasmapheresis. CONCLUSIONS: Giant cell myocarditis should be expected to recur in the allograft and often does so concurrently with rejection. However, the disease in the allograft responds to therapy in a favorable manner, which differs dramatically from that in the native heart. This might be the result of detection of the disease at an earlier stage than in the native heart, or the immunosuppression milieu in the allograft. The favorable response to therapy suggests that the likelihood of recurrence of giant cell myocarditis should not be considered a barrier to transplantation. PMID- 11257567 TI - Pathologic regression of primary pulmonary hypertension in left native lung following right single-lung transplantation. PMID- 11257569 TI - Heart failure: running to the rescue. PMID- 11257568 TI - Gene patenting: what is implied for the future of R&D? PMID- 11257570 TI - Vaccine stops blood supply. PMID- 11257571 TI - Genome sequences reveal key genetic element in PD. PMID- 11257574 TI - The human medicine project has started: place your bets now. PMID- 11257575 TI - Is there a future for GMO medicines in New Zealand? PMID- 11257576 TI - What does the human genome sequence mean to you? PMID- 11257578 TI - Rhodopsin crystal structure: provides information on GPCR-ligand binding in general? PMID- 11257577 TI - Education: the chemistry between academia and industry. AB - The Discussion Forum provides a medium for airing your views on any issues related to the pharmaceutical industry and obtaining feedback and discussion on these views from others in the field. You can discuss issues that get you hot under the collar, practical problems at the bench, recently published literature, or just something bizarre or humorous that you wish to share. Publication of letters in this section is subject to editorial discretion and company promotional letters will be rejected immediately. Furthermore, the views provided are those of the authors and are not intended to represent the views of the companies they work for. Moreover, these views do not reflect those of Elsevier, Drug Discovery Today or its editorial team. Please submit all letters to Rebecca Lawrence, News & Features Editor, Drug Discovery Today, e-mail: Rebecca.Lawrence@current-trends.com PMID- 11257579 TI - The optimal fragmentation principle - Reply. PMID- 11257580 TI - Strategic and technical challenges for drug discovery. PMID- 11257581 TI - In vivo pharmacokinetics and pharmacodynamics in drug development using positron emission tomography. AB - Positron-emission tomography (PET) is a sensitive technique that can be used to measure drug pharmacokinetics and pharmacodynamics non-invasively in target tissues of patients. Here we focus on the application of this technology to address some of the bottlenecks in drug development, including: elucidation of pathophysiology, evaluation of pharmacokinetics, proof of principle of mechanism, and assessment of efficacy and/or response to therapy. PMID- 11257582 TI - The cellular delivery of antisense oligonucleotides and ribozymes. AB - The design and development of antisense oligonucleotides and ribozymes for the treatment of diseases arising from genetic abnormalities has become a real possibility over the past few years. Improvements in oligonucleotide chemistry have led to the synthesis of nucleic acids that are relatively stable in the biological milieu. However, advances in cellular targeting and intracellular delivery will probably lead to more widespread clinical applications. This review looks at recent advances in the in vitro and in vivo delivery of antisense oligodeoxynucleotides and ribozymes. PMID- 11257583 TI - Impact of human genome sequencing for in silico target discovery. AB - The year 2000 stands as a landmark in modern biology: the first draft of the human genome sequence has been completed. For the pharmaceutical industry, this achievement provides tremendous opportunities because the genomic sequence exposes all human drug targets for therapeutic intervention. The challenge for the pharmaceutical companies is to exploit this definitive resource for the identification of potential molecular targets, rapid characterization of their function and validation of their involvement in disease pathology. Bioinformatics approaches provide increasingly crucial tools to systematically support this exploratory target drug discovery activity. PMID- 11257584 TI - Monitor: molecules and profiles. AB - Monitor provides an insight into the latest developments in drug discovery through brief synopses of recent presentations and publications together with expert commentaries on the latest technologies. There are two sections: Molecules summarizes the chemistry and the pharmacological significance and biological relevance of new molecules reported in the literature and on the conference scene; Profiles offers commentary on promising lines of research, emerging molecular targets, novel technology, advances in synthetic and separation techniques and legislative issues. PMID- 11257585 TI - Combinatorial chemistry. PMID- 11257587 TI - Erratum. PMID- 11257586 TI - Drug delivery. PMID- 11257588 TI - [Role of the ETS transcription factors in the control of endothelial-specific gene expression and in angiogenesis]. AB - The transcription factors of the ETS family are involved in the control of the endothelial-specific expression of genes that are important for the formation of new blood vessels. The analysis of the expression pattern of ets1, the gene inactivation of tel and fli1, the in vitro analysis of potential target genes of ETS factors in endothelial cells, the in vivo studies of the promoter regions of endothelial-specific genes all demonstrate a role for ETS factors in this specificity. However, the precise role of individual ETS factors in the endothelial identity and in angiogenesis in general remains difficult to understand in vivo. PMID- 11257589 TI - [Acute myeloid leukemia in the elderly: a review]. AB - The diagnosis and management of acute myeloid leukemia (AML) in the elderly is reviewed, including the basic aspects of epidemiology, cytogenetics, and prognostic factors. AML at higher ages is associated with several unique biologic and clinical characteristics. It generally arise from an early level of hematopoietic stem cells, and has a particularly high incidence of poor prognostic karyotypes. Effective treatment of the elderly patient with AML remains a challenging task. The importance of therapeutic approaches and promising new drugs is summarized. Prospective randomized studies clearly demonstrate that elderly patients benefit from intensive induction therapy. Hematopoietic growth factors accelerate the recovery from treatment-induced neutropenia and may improve the outcome. In patients not eligible for intensive chemotherapy, prospective studies testing different palliative or moderately intensive treatments are needed to improve quality of life and survival. New treatment strategies need to be developed to further improve on the therapeutic perspectives for elderly patients with AML. PMID- 11257590 TI - [Intra-operative radiation therapy in tumors of the digestive tract]. AB - All the retrospective and prospective studies concerning IORT in tumors of the digestive tract tend to substantiate a significant improvement in local control without a significant increase in survival. Improvements in the results of IORT can be expected in the near future, and may occur on several fronts: - Technological improvements: advances in the development of IORT machines, with the construction of electron accelerators specifically designed for IORT; greater precision in the systems of collimation (asymmetric collimator, multiple leaves, computerization of the control of collimation); and increased adaptability of the localizers to each clinical situation and to each patients. - Increase in the biological effects of IORT: determination of the exact role of IORT in relation to other therapeutic methods, its integration into the global therapeutic strategy for cancer, and the optimization of IORT doses should all be studied in phase II and III trials. Interesting results are expected from the combination of different methods of preoperative, intraoperative, and postoperative radiotherapy with chemotherapy; the cumulative effect of radiosensitization and cytotoxicity can bring about both local control and treatment of the general disease. In addition, the combination of hypoxic cell radiosensitizers and IORT, a source of important cellular hypoxia as a result of single doses, appears promising. - Lastly, randomized studies in a larger number of patients with objectives and methodologies to be perfected will document the actual contribution of IORT to an increase in survival as part of an overall treatment strategy for digestive tumors. At present, the prognosis remains significantly related to systemic metastatic evolution; this can only be influenced by chemotherapy, whose efficacy remains to be demonstrated. As means for better control of systemic disease are discovered, the clear benefits of local control via IORT will assume increased importance. PMID- 11257592 TI - [Conservative surgery for T2 > 3 cm, T3 N0 N1 M0 breast cancer after induction chemotherapy]. AB - Induction chemotherapy (IC) provides in more than 20% of cases a complete shrinkage of the tumor. This down staging is a new challenge for the surgeons for breast conservative procedure. Although, IC has become the standard of care for breast cancer T2 > 3 cm T3 N0 N1 M0. No guidelines have devoted attention to the surgical problems due to this down staging after IC. Location and size of the tumor before IC have to be studied and outlined by the surgeon himself. During surgery, the residual tumor volume and how much mammary gland must be removed are very difficult to determine. The maximum volume of mammary gland to be removed after IC around the primary site of the tumor before IC is the volume which permits a good cosmetic reconstruction of the breast. After IC, in spite of an important downstaging, an axillary clearance must be done. For N0 patients, sentinel lymph node biopsy could be performed before IC. If the sentinel node is p N0, axillary clearance could be avoided. PMID- 11257591 TI - [Prevention and treatment of chemotherapy-induced digestive toxicities]. AB - Digestive complications are frequent dose-limiting side-effects of chemotherapy. The incidence and the severity of these toxicities have to be systematically evaluated in order to provide specific curative and preventive treatments. Quick resolution of these complications is important to improve quality of life, and also to prevent hospitalization. Nausea, vomiting, mucositis, and diarrhea are the main digestive toxicities of chemotherapy. This review emphasizes the recent therapeutic approaches which could lead to a decrease of the incidence or the severity of these toxicities. Although marked progress has been accomplished these last years, a substantial degree of further improvement is needed to improve the effectiveness of specific treatments. Furthermore, research might focus on new strategies which may prevent the development of overall toxicity, by increasing the selectivity of chemotherapy on tumor cells. PMID- 11257593 TI - [Standards, Options and Recommendations for the surgical management of carcinoma of the endometrium]. AB - CONTEXT: The "Standards, Options and Recommendations" (SOR) project, started in 1993, is a collaboration between the Federation of the French Cancer Centres (FNCLCC), the 20 French Cancer Centres and specialists from French Public Universities, General Hospitals and Private Clinics. The main objective is the development of clinical practice guidelines to improve the quality of health care and outcome for cancer patients. The methodology is based on literature review and critical appraisal by a multidisciplinary group of experts, with feedback from specialists in cancer care delivery. OBJECTIVES: To develop clinical practice guidelines according to the definitions of the Standards, Options and Recommendations project for the surgical management of carcinoma of the endometrium. METHODS: Data were identified by searching Medline and personal reference lists of members of the expert groups. Once the guidelines were defined, the document was submitted for review to independent reviewers, and to the medical committees of the 20 French Cancer Centres. RESULTS: The main recommendations for the surgical management of carcinoma of the endometrium are: 1) where-ever possible, surgery is the primary treatment of both localised and advanced disease; 2) surgery is performed according to the stage of the cancer and the status of the patient; 3) surgery for stages I and II disease entails total abdominal hysterectomy and bilateral salpingo-oophorectomy. A modified radical hysterectomy is undertaken in cases of macroscopic cervical involvement. An omenectomy is recommended for serous papillary types. Pelvic lymphadenectomy for the purposes of precise staging is undertaken if the patient is of good performance status and without bad pronostic factors. Para-aortic lymphadenectomy can be undertaken to determine involvement of para-aortic nodes; 4) surgery for stages III and IV: radical surgery must be undertaken if at all possible with additional treatment as indicated. In the case of advanced disease, debulking surgery is indicated. PMID- 11257594 TI - [Xeroderma pigmentosum: report of 6 cases in Dakar]. AB - We report 6 cases of black Senegalese boys with xeroderma pigmentosum. They were between 2 and 16 year-old and presented features of hypersensitivity to UV (keratosis, lentigines, poikilodermia and photophobia). Our cases were remarkable by the early occurrence of squamous cell and basal cell carcinoma located in photoexposed sites causing the death of 5 of them. Xeroderma pigmentosum must be considered as the first preneoplastic genodermatosis. PMID- 11257595 TI - [Chemotherapy with cisplatinum, carboplatin and 5FU-folinic acid, followed by concomitant chemo-radiotherapy in unresectable esophageal carcinomas]. AB - The best chemotherapeutic regimen for advanced carcinoma of the esophagus remains to be determined. We have evaluated a combination of carboplatin, cisplatin and 5FU modulated by folinic acid. Patients. Twenty-seven patients (median age 57 yrs) with an unresectable carcinoma of the esophagus were included in this trial: 9 patients with a local relapse after surgery, 6 patients with a locally advanced (T4) tumor, and 12 patients with metastasis. Treatment schedule. Initial chemotherapy : carboplatine IV d1, AUC4; 5FU: bolus injection of 400 mg/m2 d1, followed by a continuous infusion of 600 mg/m2/24 h, d1 and d2; folinic acid (200 mg/m2) IV, before the 5FU bolus, d1 and d2; cisplatine 80 mg/m2, d3; on d15 and d16, 5FU and folinic acid were repeated with the same schedule. The second cycle began on d28. Concomitant chemo-radiotherapy with 5FU (1,000 mg/m2 d1 to d3), cisplatine (50 mg/m2 d1 and d2) and external irradiation (20 Gy in 10 fractions from d1 to d12) was then performed, for three cycles (until a total dose of 60 Gy). Results. TOXICITY: neutropenia grade 3-4 (32%), thrombopenia grade 3-4 (18%). More important, a lymphopenia (< 500/mm3) was noted in 12 patients (43%). Accordingly, 4 serious infectious complications were observed, with three toxic deaths. Objective response rate: 44% after initial chemotherapy; 75% after chemoradiotherapy, with 8 complete responses (38%). Median survival was 7.4 months, with a one- and two-year survival of 33% and 17,8%, respectively. Conclusion. This association of cisplatin, carboplatin, and 5FU did not offer a better response rate than the classical 5FU-cisplatinum association. But serious infectious complications occurred during the trial. We do not recommended further evaluation of this biplatinum therapy with 5FU in advanced esophageal carcinomas. PMID- 11257596 TI - [Clinical coding in oncology]. PMID- 11257597 TI - [Cancers and oncologists from the GP's point of view: opinion and comments]. PMID- 11257598 TI - [About standard: from rule to patient. Breast cancer as an example]. PMID- 11257599 TI - Modeling of the enzymatic kinetic synthesis of cephalexin--influence of substrate concentration and temperature. AB - During enzymatic kinetic synthesis of cephalexin, an activated phenylglycine derivative (phenylglycine amide or phenylglycine methyl ester) is coupled to the nucleus 7-aminodeacetoxycephalosporanic acid (7-ADCA). Simultaneously, hydrolysis of phenylglycine amide and hydrolysis of cephalexin take place. This results in a temporary high-product concentration that is subsequently consumed by the enzyme. To optimize productivity, it is necessary to develop models that predict the course of the reaction. Such models are known from literature but these are only applicable for a limited range of experimental conditions. In this article a model is presented that is valid for a wide range of substrate concentrations (0 490 mM for phenylglycine amide and 0-300 mM for 7-ADCA) and temperatures (273-298 K). The model was built in a systematic way with parameters that were, for an important part, calculated from independent experiments. With the constants used in the model not only the synthesis reaction but also phenylglycine amide hydrolysis and cephalexin hydrolysis could be described accurately. In contrast to the models described in literature, only a limited number (five) of constants was required to describe the reaction at a certain temperature. For the temperature dependency of the constants, the Arrhenius equation was applied, with the constants at 293 K as references. Again, independent experiments were used, which resulted in a model with high statistic reliability for the entire temperature range. Low temperatures were found beneficial for the process because more cephalexin and less phenylglycine is formed. The model was used to optimize the reaction conditions using criteria such as the yield on 7-ADCA or on activated phenylglycine. Depending on the weight of the criteria, either a high initial phenylglycine amide concentration (yield on 7-ADCA) or a high initial 7 ADCA concentration (yield on phenylglycine amide) is beneficial. PMID- 11257600 TI - Kinetic resolution of chiral amines with omega-transaminase using an enzyme membrane reactor. AB - A kinetic resolution process for the production of chiral amines was developed using an enzyme-membrane reactor (EMR) and a hollow-fiber membrane contactor with (S)-specific omega-transaminases (omega-TA) from Vibrio fluvialis JS17 and Bacillus thuringiensis JS64. The substrate solution containing racemic amine and pyruvate was recirculated through the EMR and inhibitory ketone product was selectively extracted by the membrane contactor until enantiomeric excess of (R) amine exceeded 95%. Using the reactor set-up with flat membrane reactor (10-mL working volume), kinetic resolutions of alpha-methylbenzylamine (alpha-MBA) and 1 aminotetralin (200 mM, 50 mL) were carried out. During the operation, concentration of ketone product, i.e., acetophenone or alpha-tetralone, in a substrate reservoir was maintained below 0.1 mM, suggesting efficient removal of the inhibitory ketone by the membrane contactor. After 47 and 32.5 h of operation using 5 U/mL of enzyme, 98.0 and 95.5% ee of (R)-alpha-MBA and (R)-1 aminotetralin were obtained at 49.5 and 48.8% of conversion, respectively. A hollow-fiber membrane reactor (39-mL working volume) was used for a preparative scale kinetic resolution of 1-aminotetralin (200 mM, 1 L). After 133 h of operation, enantiomeric excess reached 95.6% and 14.3 g of (R)-1-aminotetralin was recovered (97.4% of yield). Mathematical modeling of the EMR process including the membrane contactor was performed to evaluate the effect of residence time. The simulation results suggest that residence time should be short to maintain the concentration of the ketone product in EMR sufficiently low so as to decrease conversion per cycle and, in turn, reduce the inhibition of the omega-TA activity. PMID- 11257601 TI - Metabolic control of recombinant protein N-glycan processing in NS0 and CHO cells. AB - Chinese hamster ovary and murine myeloma NS0 cells are currently favored host cell types for the production of therapeutic recombinant proteins. In this study, we compared N-glycan processing in GS-NS0 and GS-CHO cells producing the same model recombinant glycoprotein, tissue inhibitor of metalloproteinases 1. By manipulation of intracellular nucleotide-sugar content, we examined the feasibility of implementing metabolic control strategies aimed at reducing the occurrence of murine-specific glycan motifs on NS0-derived recombinant proteins, such as Galalpha1,3Galbeta1,4GlcNAc. Although both CHO and NS0-derived oligosaccharides were predominantly of the standard complex type with variable sialylation, 30% of N-glycan antennae associated with NS0-derived TIMP-1 terminated in alpha1,3-linked galactose residues. Furthermore, NS0 cells conferred a greater proportion of terminal N-glycolylneuraminic (sialic) acid residues as compared with the N-acetylneuraminic acid variant. Inclusion of the nucleotide-sugar precursors, glucosamine (10 mM, plus 2 mM uridine) and N acetylmannosamine (20 mM), in culture media were shown to significantly increase the intracellular pools of UDP-N-acetylhexosamine and CMP-sialic acid, respectively, in both NS0 and CHO cells. The elevated UDP-N-acetylhexosamine content induced by the glucosamine/uridine treatment was associated with an increase in the antennarity of N-glycans associated with TIMP-1 produced in CHO cells but not N-glycans associated with TIMP-1 from NS0 cells. In addition, elevated UDP-N-acetylhexosamine content was associated with a slight decrease in sialylation in both cell lines. The elevated CMP-sialic acid content induced by N acetylmannosamine had no effect on the overall level of sialylation of TIMP-1 produced by both CHO and NS0 cells, although the ratio of N-glycolylneuraminic acid:N-acetylneuraminic acid associated with NS0-derived TIMP-1 changed from 1:1 to 1:2. These data suggest that manipulation of nucleotide-sugar metabolism can promote changes in N-glycan processing that are either conserved between NS0 and CHO cells or specific to either NS0 cells or CHO cells. PMID- 11257602 TI - Improved oligosaccharide synthesis by protein engineering of beta-glucosidase CelB from hyperthermophilic Pyrococcus furiosus. AB - Enzymatic transglycosylation of lactose into oligosaccharides was studied using wild-type beta-glucosidase (CelB) and active site mutants thereof (M424K, F426Y, M424K/F426Y) and wild-type beta-mannosidase (BmnA) of the hyperthermophilic Pyrococcus furiosus. The effects of the mutations on kinetics, enzyme activity, and substrate specificity were determined. The oligosaccharide synthesis was carried out in aqueous solution at 95 degrees C at different lactose concentrations and pH values. The results showed enhanced synthetic properties of the CelB mutant enzymes. An exchange of one phenylalanine to tyrosine (F426Y) increased the oligosaccharide yield (45%) compared with the wild-type CelB (40%). Incorporation of a positively charged group in the active site (M424K) increased the pH optimum of transglycosylation reaction of CelB. The double mutant, M424K/F426Y, showed much better transglycosylation properties at low (10-20%) lactose concentrations compared to the wild-type. At a lactose concentration of 10%, the oligosaccharide yield for the mutant was 40% compared to 18% for the wild-type. At optimal reaction conditions, a higher ratio of tetrasaccharides to trisaccharides was obtained with the double mutant (0.42, 10% lactose) compared to the wild-type (0.19, 70% lactose). At a lactose concentration as low as 10%, only trisaccharides were synthesized by CelB wild-type. The beta-mannosidase BmnA from P. furiosus showed both beta-glucosidase and beta-galactosidase activity and in the transglycosylation of lactose the maximal oligosaccharide yield of BmnA was 44%. The oligosaccharide yields obtained in this study are high compared to those reported with other transglycosylating beta-glycosidases in oligosaccharide synthesis from lactose. PMID- 11257603 TI - Comparison of Bcl-2 to a Bcl-2 deletion mutant for mammalian cells exposed to culture insults. AB - Apoptosis has been found to occur in bioreactors as a result of environmental stresses. The overexpression of bcl-2 is a widely used strategy to limit the induction of apoptosis in mammalian cell cultures. In this study, the effectiveness of wild-type Bcl-2 was compared to a Bcl-2 mutant lacking the nonstructured loop domain in two commercially prominent cell lines, Chinese hamster ovary (CHO) and baby hamster kidney (BHK) cells. The generation of a DNA "ladder" and condensation of chromatin indicated that apoptosis occurred in these cell lines following Sindbis virus infection and serum deprivation. When cells were engineered to overexpress the bcl-2 mutant, cell death due to Sindbis virus was inhibited in a concentration-dependent manner. Furthermore, the Bcl-2 mutant provided increased protection as compared to wild-type Bcl-2 following two model insults, Sindbis virus infection and serum deprivation. Total production for a heterologous protein encoded on the Sindbis virus was increased in cell lines expressing the Bcl-2 variants compared to the parental cell line. In order to understand the reasons for the improved anti-apoptosis properties of the mutant, wild-type Bcl-2 and mutant Bcl-2 were examined by Western blot following each model insult. Wild-type Bcl-2 was observed to degrade into a 23 kDa fragment following both Sindbis virus infection and serum withdrawal in both cell lines, while the mutant Bcl-2 protein was not degraded during the same period. The processing of Bcl-2 was found to correlate with reduced cell viabilities following the two external insults to suggest that Bcl-2 degradation may limit its ability to inhibit apoptosis. These studies indicate that the cells regulate anti-apoptosis protein levels and these processing events can limit the effectiveness of cell death inhibition strategies in mammalian cell culture systems. PMID- 11257604 TI - Avoiding acetate accumulation in Escherichia coli cultures using feedback control of glucose feeding. AB - An automated glucose feeding strategy that avoids acetate accumulation in cultivations of Escherichia coli is discussed. We have previously described how a probing technique makes it possible to detect and avoid overflow metabolism using a dissolved oxygen sensor. In this article these ideas are extended with a safety net that guarantees that aerobic conditions are maintained. The method is generally applicable, as no strain-specific information is needed and the only sensor required is a standard dissolved oxygen probe. It also gives the highest feed rate possible with respect to limitations from overflow metabolism and oxygen transfer, thus maximizing bioreactor productivity. The strategy was implemented on three different laboratory-scale platforms and fed-batch cultivations under different operating conditions were performed with three recombinant strains, E. coli K-12 UL635, E. coli BL21(DE3), and E. coli K-12 UL634. In spite of disturbances from antifoam and induction of recombinant protein production, the method reproducibly gave low concentrations of acetate and glucose. The ability to obtain favorable cultivation conditions independently of strain and operating conditions makes the presented strategy a useful tool, especially in situations where it is important to get good results on the first attempt. PMID- 11257605 TI - Variations in the enantioselectivity of salt-activated subtilisin induced by lyophilization. AB - Including excess salt during lyophilization has been shown to increase the activity of freeze-dried subtilisin Carlsberg (SC) in anhydrous media by over 20,000-fold [Ru et al. (1999) Biotechnol Bioeng 63:233-241]. In the present study, salt-activated SC (KCl-SC) showed a 30% enhancement in enantioselectivity compared to the salt-free enzyme in a variety of organic solvents. Activity toward both enantiomers of N-acetyl-phenylalanine methyl ester (APME) increased in tandem by 2-3 orders of magnitude in all solvents, indicating that the mechanism of salt activation is inherent to the enzyme and does not strongly favor one enantiomer over the other. However, activity and enantioselectivity of salt-activated SC could be manipulated through changes in the lyophilization conditions. Variations in lyophilization time, initial KCl concentration, and initial lyophilization volume altered enantioselectivity over 2-fold. The changes in enantioselectivity reflected the activity for the L enantiomer, while the activity toward the D enantiomer was mostly unaffected. The results indicate that the lyophilization time and final water content of the KCl-SC are important determinants of enzyme activity for the L enantiomer, suggesting that the favored reaction is more sensitive to the structural integrity of the salt-activated enzyme. PMID- 11257606 TI - The metabolic burden of the PGK1 and ADH2 promoter systems for heterologous xylanase production by Saccharomyces cerevisiae in defined medium. AB - Five recombinant S. cerevisiae strains were cultivated under identical conditions to quantify the molecular basis of the metabolic burden of heterologous gene expression, and to evaluate mechanisms for the metabolic burden. Two recombinant S. cerevisiae strains, producing Trichoderma reesei xylanase II under control of either the PGK1 or ADH2 promoters, were compared quantitatively with three references strains, where either the heterologous xylanase II (XYN2) gene, or the heterologous gene and the promoter and terminator were omitted from the recombinant plasmid. Neither the replication of multiple copies of the 2-microm based YEp352 plasmid nor the replication the foreign XYN2 gene represented a metabolic burden to the cell, as the growth of the host organism was not affected. The inclusion of a glycolytic promoter on the recombinant plasmid, however, reduced the maximum specific growth rate (12% to 15%), biomass yield on glucose (8% to 11%), and specific glucose consumption rate (6% to 10%) of the recombinant strains. The presence of the heterologous XYN2 gene on the recombinant plasmid caused a further reduction in the maximum specific growth rate (11% to 14%), biomass yield (4%), and specific glucose consumption rate (12%) of the host strain during active gene expression, which was dictated by the regulatory characteristics of the promoter utilized. The metabolic effect of foreign gene expression was disproportionally large, with respect to on the amount of heterologous protein produced. This was most likely due to an increased energetic demand for the expression of a foreign gene and/or a competition for limiting amounts of transcription or translation factors, biosynthetic precursors or metabolic energy. PMID- 11257607 TI - Improving biomaterial properties of collagen films by chemical modification. AB - Films of bovine collagen were chemically modified with the goal of improving their biomaterial properties. The modified films were investigated with respect to their affinity to fibroblast and endothelial cells, as well as their antibacterial properties tested by adhesion of Staphylococcus aureus. Modifications that only change the net charge of collagen, such as acetylation, succinylation, and treatment with glutaraldehyde (all increase the negative charge), and amination with ethylenediamine (EDA), N,N-dimethyl-EDA (DMEDA), or butylamine (all increase the positive charge), did not dramatically alter the mammalian cell attachment to the film. In contrast, derivatization of collagen using methoxypoly(ethylene glycol) (PEG) diminished the attachment of fibroblasts by 98 +/- 1% and of endothelial cells by more than 99% compared to unmodified collagen. Moreover, the rate of growth of fibroblasts dropped by 97 +/- 1% and that of endothelial cells by 88 +/- 3% as a result of PEGylation of collagen. Adhesion of S. aureus cells also plummeted by 93 +/- 2% as a result of this PEGylation. With these antifouling properties, PEG-collagen may be a promising coating material for coronary stents. Subsequent derivatization of PEG-collagen with EDA or DMEDA abolished its mammalian cell-repelling ability, whereas bacterial cell repulsion was partially retained: for example, DMEDA-modified PEG collagen exhibits up to a 5-fold lower bacterial adhesion than collagen. It is worth noting that a material that allows mammalian cell attachment but reduces bacterial adhesion could be useful as an implant or coating. PMID- 11257608 TI - One-step acclimatization of plantlets using a mist reactor. AB - A mist reactor was used to grow and acclimatize carnation plants in vitro without using ex vitro acclimatization techniques. The acclimatization protocol in the reactor consisted of altering the mist-on period during the course of the culture period and a stepwise reduction in the relative humidity surrounding the plants from 98% to 70% relative humidity (RH) during the final week of in vitro growth. After transfer and further growth in a greenhouse for 5 weeks, survival was 91% for plants grown in reactors, 81% from vented boxes, and 50% from unvented boxes. Ex vitro survival directly correlated with increased in vitro rooting and decreased hyperhydration. In vitro rooting also correlated with high-quality plants, but did not significantly correlate with low hyperhydration, as normal plants often lacked roots. After 5 weeks in the greenhouse, the quantity of mid- and high-quality plants obtained from reactors and ventilated boxes was similar. Conditions in the mist reactor can be manipulated to produce plants that are readily acclimatized and are equal or better in quality and yield than plants produced using conventional methods. PMID- 11257609 TI - Laser ablation reveals regulation of ciliary activity by serotonergic neurons in molluscan embryos. AB - Early in embryonic development, the pond snail Helisoma trivolvis exhibits a rotational behavior that is generated by beating of cilia in the dorsolateral and pedal bands. Although previous anatomical and pharmacological studies provided indirect evidence that a pair of serotonergic neurons, Embryonic Neurons C1 (ENC1s), is involved in regulating embryonic rotation, direct evidence linking ENC1 to ciliary function is still lacking. In the present study, we used laser microbeams to perturb ENC1 in vivo while monitoring ciliary activity in identified ciliary bands. A laser treatment protocol to specifically ablate ENC1 without damaging the surrounding cells was established. Unilateral laser treatment of ENC1 caused transient increases in the activity of the pedal and ipsidorsolateral cilia, lasting 30-50 min. In contrast, activity of cilia that were not anatomically associated with ENC1 was unaffected by laser treatment. Mianserin, an effective serotonin antagonist in Helisoma ciliated cells, decreased the overall CBF of pedal and dorsolateral cilia by reducing the occurrence of spontaneous CBF surges in these cilia. Finally, the cilioexcitatory action of ENC1 laser treatment was mimicked by serotonin and reduced in the presence of mianserin. These results suggest that laser treatment provokes a release of serotonin from ENC1, resulting in a prolonged elevation of activity in the target ciliary cells. We conclude that, in addition to their previously established role in regulating neurodevelopment, ENC1s also function as serotonergic motor neurons to regulate ciliary activity, and therefore the rotational behavior of early embryos. PMID- 11257610 TI - Drosophila neuropeptide F mediates integration of chemosensory stimulation and conditioning of the nervous system by food. AB - The conserved neuropeptide Y (NPY) signaling pathway has been strongly implicated in the stimulation of food uptake in vertebrates as well as in the regulation of food conditioned foraging behaviors of Caenorhabditis elegans. Using in situ RNA hybridization and immunocytochemistry, we report the neuronal network of Drosophila neuropeptide F (dNPF), a human NPY homologue, in the larval central nervous system and its food-dependent modifications. We provide indications that gustatory stimulation by sugar, but not its ingestion or metabolism, is sufficient to trigger long-term, dose-dependent alterations of the dNPF neuronal circuit through both dnpf activation and increased synaptic transmission. Our results strongly suggest that the dNPF neuronal circuit is an integral part of the sensory system that mediates food signaling, providing the neural basis for understanding how invertebrate NPY regulates food response. PMID- 11257611 TI - Calmodulin and profilin coregulate axon outgrowth in Drosophila. AB - Coordinated regulation of actin cytoskeletal dynamics is critical to growth cone movement. The intracellular molecules calmodulin and profilin actively regulate actin-based motility and participate in the signaling pathways used to steer growth cones. Here we show that in the developing Drosophila embryo, calmodulin and profilin convey complimentary information that is necessary for appropriate growth cone advance. Reducing calmodulin activity by expression of a dominant inhibitor (KA) stalls axon extension of pioneer neurons within the CNS, while a partial loss of profilin function decreases extension of motor axons in the periphery. Yet, surprisingly, when calmodulin and profilin are simultaneously reduced, the ability of both CNS pioneer axons and motor axons to extend beyond the choice points is restored. In the CNS, at the time when growth cones must decide whether to cross or not to cross the midline, a reduction in calmodulin and/or roundabout signaling causes axons to cross the midline inappropriately. These inappropriate crossings are suppressed when profilin activity is simultaneously reduced. Interestingly, the mutual suppression of calmodulin and profilin activity requires a minimal level of profilin. In KA combinations with profilin null alleles, defects in axon extension and midline guidance are synergistically enhanced rather than suppressed. Together, our data indicate that the growth cone must coordinate the activity of both calmodulin and profilin in order to advance past selected choice points, including those dictating midline crossovers. PMID- 11257612 TI - Phosphatidylinositol-3-OH kinase regulatory subunits are differentially expressed during development of the rat cerebellum. AB - Recent evidence implicates a central role for PI3K signalling in mediating cell survival during the process of neuronal differentiation. Although PI3K activity is stimulated by a wide range of growth factors and cytokines in different cell lines and tissues, activation of this pathway by insulin-like growth factor I (IGF-I) most likely represents the main survival signal during neuronal differentiation. IGF-I is highly expressed during development of the central nervous system, and thus is a critical factor for the development and maturation of the cerebellum. Upon ligand binding, the IGF-I receptor phosphorylates tyrosine residues in SHC and insulin receptor substrates (IRSs) initiating two main signalling cascades, the MAP kinase and the phosphatidylinositol 3-kinase (PI3K) pathways. Activated PI3K is composed of a catalytic subunit (p110alpha or beta) associated with one of a large family of regulatory subunits (p85alpha, p85beta, p55gamma, p55alpha, and p50alpha). To evaluate the contributions of these various regulatory subunits to neuronal differentiation, we have used antibodies specific for each of the PI3K subunits. Using these antisera, we now demonstrate that PI3K subunits are differentially regulated in cerebellar development, and that the expression level of the p55gamma regulatory subunit reaches a maximum during postnatal development, decreasing thereafter to low levels in the adult cerebellum. Furthermore, our studies reveal that the distribution of the various PI3K regulatory subunits varies during development of the cerebellum. Interestingly, p55gamma is expressed in both glial and neuronal cells; moreover, in Purkinje neurones, this subunit colocalises with the IGF-IR. PMID- 11257613 TI - Postembryonic proliferation in the spiny lobster antennular epithelium: rate of genesis of olfactory receptor neurons is dependent on molt stage. AB - Olfactory systems undergo continuous growth and turnover in many animals. Many decapod crustaceans, such as lobsters and crayfish, have indeterminate growth, and in these animals, turnover of both peripheral and central components of the olfactory system occurs continuously throughout life. In this study, we examine the dynamics of olfactory receptor neuron (ORN) proliferation in the antennule of the Caribbean spiny lobster, Panulirus argus, using in vivo incorporation of the cell proliferation marker BrdU. We show that addition of ORNs occurs in a "proximal proliferation zone" (PPZ), which exists on the proximo-lateral margin of the existing ORN population. The PPZ is spatially and temporally dynamic in that it travels as a wave in the proximal and lateral directions in the antennule. This wave results in continuous addition of ORNs throughout the molt cycle. The rate of proliferation, as measured by the size and shape of the PPZ, changes depending on the animal's molt stage. The rate is highest during premolt and lowest during intermolt. ORNs are the most prominent cell-type produced in the PPZ, but other cell types, including glia, are also produced. Patches of proliferating epithelial cells occur immediately proximal to the PPZ, suggesting that neuronal and glial precursors reside in this region. Possible mechanisms for peripheral and central modulation of ORN development are discussed. PMID- 11257614 TI - RISK-1: a novel MAPK homologue in axoplasm that is activated and retrogradely transported after nerve injury. AB - Sensory neurons (SNs) of Aplysia are widely used to study the molecular correlates of learning. Among these is the activation of an Aplysia (ap) MAPK that phosphorylates the transcription factor apC/EBPbeta. Because crushing the axons of the SNs induces changes similar to learning, we tested the hypothesis that apMAPK is a point of convergence on the pathways for learning and injury. One event in common is long-term hyperexcitability (LTH), and LTH was induced in the SNs after intrasomatic injection of active vertebrate extracellular signal regulated kinase 1 (ERK1; as an apMAPK surrogate). Nerve crush activated an axoplasmic kinase at the site of injury that phosphorylated apC/EBPbeta. Surprisingly, this was not apMAPK, but a kinase that was recognized by antibodies to vertebrate ERKs and to doubly phosphorylated, activated ERKs. The activated kinase was transported to the cell body and nucleus and its arrival was concurrent with an injury-induced increase in apC/EBPbeta mRNA and protein. We call this retrogradely transported kinase RISK-1. RISK-1 initiated the binding of apC/EBPbeta to the ERE enhancer site in vitro and an increase in ERE-binding was detected in injured neurons containing active RISK-1. Thus, Aplysia neurons contain two MAPK homologues, one of which is a late acting retrogradely transported injury signal. PMID- 11257616 TI - Refined diagnostic criteria for implants associated with ovarian atypical proliferative serous tumors (borderline) and micropapillary serous carcinomas. AB - Characterization of invasive peritoneal implants from patients with noninvasive serous ovarian tumors has important prognostic and treatment implications, but the criteria for distinguishing invasive and noninvasive implants vary among investigators and can be difficult to apply. The authors studied 148 implants from 60 patients, 33 with primary atypical proliferative serous tumor, and 27 with primary noninvasive micropapillary serous carcinoma, with a mean follow-up of 62 months (median follow-up, 52 months). Previously reported and newly proposed histologic features for implant classification were evaluated and correlated with clinical outcome. Three criteria were applied for the diagnosis of "invasive" implants: invasion of underlying normal tissue, micropapillary architecture, and solid epithelial nests surrounded by clefts. Implants displaying any one of these three features were classified as "invasive," whereas those lacking all three features were classified as "noninvasive." Sixty-six implants were invasive and 82 were noninvasive. Of the 31 patients with invasive implants, six were dead of disease (DOD), 13 were alive with progressive disease (AWPD), and 12 were alive with no evidence of disease (NED). Of the 29 patients with noninvasive implants, two were DOD, one was dead of uncertain causes, one was AWPD, and 25 were alive with NED. Eighty-nine percent of invasive implants had a micropapillary architecture and 83% had solid epithelial nests surrounded by clefts. A minority of invasive implants (14% of those with underlying normal tissue) demonstrated invasion of normal underlying tissue. Nuclear atypia, mitoses, calcification, necrosis, and identification of individual cells "infiltrating" the stroma did not correlate with implant type. The proposed criteria permitted recognition of implants that correlated strongly with adverse outcome. Sixty-one percent of patients with implants displaying any one of the three features used to diagnose invasive implants were AWPD or DOD compared with 10% of patients whose implants lacked these features (p = 0.00001). Because implants associated with an adverse outcome can be identified before they invade underlying normal tissue, the term invasive implant to describe them is inaccurate and misleading. These implants resemble patterns of growth in micropapillary serous carcinoma of the ovary and the recurrent tumor that is obvious carcinoma. Accordingly, we propose that these extraovarian lesions be designated "well-differentiated serous carcinoma." PMID- 11257617 TI - The spectrum of metanephric adenofibroma and related lesions: clinicopathologic study of 25 cases from the National Wilms Tumor Study Group Pathology Center. AB - The authors report nine new metanephric adenofibroma (MAFs; previously termed nephrogenic adenofibroma) and 16 related tumors from the files of the National Wilms Tumor Study Group Pathology Center (NWTSGPC). All tumors contained a variable amount of a bland spindle cell stroma, which is essentially identical to the recently described metanephric stromal tumor (MST). Features that distinguish this stroma from congenital mesoblastic nephroma (CMN) include intratumoral angiodysplasia, concentric cuffing of entrapped tubules ("onion skinning"), and heterologous differentiation. The epithelial components of these lesions spanned a wide range of appearances. All tumors contained at least focally an inactive embryonal epithelium identical morphologically to metanephric adenoma (MA), and hence each case could be classified as containing MAF. The epithelium of nine tumors had this appearance throughout, and hence these were considered usual MAFs. The epithelium of four tumors demonstrated increased mitotic activity but was otherwise similar to MA. The epithelial component of seven tumors spanned a morphologic spectrum from inactive MA to malignant epithelial predominant Wilms tumor (WT), with gradual transitions noted in several cases. Five other tumors contained a carcinomatous component distinct from these lesions but identical morphologically to papillary renal cell carcinoma (PRCC). In one of these cases, this component had metastasized to the regional lymph nodes at the time of diagnosis. No tumor recurred during follow-up, although almost all patients received adjuvant therapy for WT regardless of their tumor's histology and NWTSGPC diagnosis. In conclusion, MAF is a biphasic tumor that spans the morphologic spectrum between benign pure stromal (MST) and pure epithelial (MA) lesions, and can merge with the morphology of WT, supporting the concept that these are all related lesions. A relationship to PRCC is also evident. PMID- 11257618 TI - Endometriosis of the intestinal tract: a study of 44 cases of a disease that may cause diverse challenges in clinical and pathologic evaluation. AB - Endometriosis of the intestinal tract may mimic a number of diseases both clinically and pathologically. The authors evaluated 44 cases of intestinal endometriosis in which endometriosis was the primary pathologic diagnosis, and evaluated them for a variety of gross and histologic changes. Cases with preneoplastic or neoplastic changes were excluded specifically because they were the subject of a previous study. The patients ranged in age from 28 to 56 years (mean age, 44 years), and presenting complaints included abdominal pain (n = 15), an abdominal mass (n = 12), obstruction (n = 8), rectal bleeding (n = 2), infertility (n = 3), diarrhea (n = 2), and increasing urinary frequency (n = 1). The clinical differential diagnoses included diverticulitis, appendicitis, Crohn's disease, tubo-ovarian abscess, irritable bowel syndrome, carcinoma, and lymphoma. Forty-two patients underwent resection of the diseased intestine and two patients underwent endoscopic biopsies. In 13 patients there were predominantly mural masses, which were multiple in two patients (mean size, 2.6 cm). In addition, 11 cases had luminal stenosis or strictures, six had mucosal polyps, four had submucosal masses that ulcerated the mucosa (sometimes simulating carcinoma), three had serosal adhesions, one had deep fissures in the mucosa, and one was associated with appendiceal intussusception. Involvement of the lamina propria or submucosa was identified in 29 cases (66%) and, of these, 19 had features of chronic injury including architectural distortion (n = 19), dense lymphoplasmacytic infiltrates (n = 7), pyloric metaplasia of the ileum (n = 1), and fissures (n = 1). Three cases had features of mucosal prolapse (7%), ischemic changes were seen in four (9%), and segmental acute colitis and ulceration were seen in four and six cases (9% and 13%) respectively. In 14 patients, endometriosis formed irregular congeries of glands involving the intestinal surface epithelium, mimicking adenomatous changes. Mural changes included marked concentric smooth muscle hyperplasia and hypertrophy, neuronal hypertrophy and hyperplasia, and fibrosis of the muscularis propria with serositis. Follow-up of 20 patients (range, 1-30 years; mean, 7.8 years) revealed that only two patients had recurrent symptoms. None of the patients developed inflammatory bowel disease. Endometriosis can involve the intestinal tract extensively, causing a variety of clinical symptoms, and can result in a spectrum of mucosal alterations. Because the endometriotic foci may be inaccessible to endoscopic biopsy or may not be sampled because of their focality, clinicians and pathologists should be aware of the potential of this condition to mimic other intestinal diseases. PMID- 11257619 TI - h-Caldesmon expression effectively distinguishes endometrial stromal tumors from uterine smooth muscle tumors. AB - Distinction of endometrial stromal neoplasms from cellular smooth muscle tumors of the uterus is sometimes difficult. Immunohistochemistry is often not helpful because muscle actins and desmin are expressed in both neoplasms. This study's goal was to determine whether h-caldesmon, a smooth muscle-specific isoform of a calcium, calmodulin, and actin binding protein, could effectively distinguish endometrial stromal tumors from uterine smooth muscle tumors. The authors analyzed immunohistochemical expression in 24 endometrial stromal neoplasms (21 sarcomas and three nodules), 29 leiomyosarcomas, 32 leiomyomas (10 "usual," 14 cellular leiomyoma, and eight "highly cellular" types), 40 myometria, and 25 endometria. h-Caldesmon was diffusely positive in all myometria, leiomyomata, and leiomyosarcomas. Of note, 16 leiomyosarcomas (55%) were positive for h-caldesmon in more than 50% of tumor cells. In five "highly cellular" leiomyomas, h caldesmon expression was markedly decreased or absent in areas morphologically resembling endometrial stromal tumors, raising the possibility that these tumors may be mixed smooth muscle-endometrial stromal neoplasms. In contrast, h caldesmon expression was absent in all endometria and endometrial stromal neoplasms apart from accompanying small vessels. Desmin was diffusely positive in all myometria and leiomyomata. The fraction of cells expressing desmin was greater than that of h-caldesmon in only 10% of leiomyosarcomas. Focal desmin expression was also present in eight of 25 (32%) endometria and 12 of 24 (50%) endometrial stromal neoplasms. h-Caldesmon appears to be a more sensitive and specific marker of smooth muscle differentiation in the uterus than desmin and may be a useful tool for distinguishing and classifying uterine mesenchymal tumors. PMID- 11257621 TI - Significance of MIB-1 staining indices in meningiomas: comparison of two counting methods. AB - The authors evaluated the predictability of MIB-1 immunohistochemistry for growth and recurrences of meningiomas using two different counting methods: 1) in the area of the highest MIB-1 labeling (HL method) and (2) in randomly selected fields (RS method). The MIB-1 staining indices (SIs) determined by the HL method were approximately twice as high as those by the RS method, and the correlation coefficient between them was high (R = 0.86) in 139 meningiomas when transformed logarithmically. The differences in SIs in histologic grades were significant with either method. Tumor doubling time (Td) was calculated in 22 meningiomas from serial radiologic examinations. The RS method yielded a slightly higher correlation coefficient between log Td and log SI than the HL method. When the authors examined the predictability of recurrence in 112 totally removed meningiomas, the RS method distinguished the recurrent group more definitively. Several benign meningiomas with low SIs by the RS method exhibited focal accumulation of MIB-1-positive cells. Although they were assigned high MIB-1 values by the HL method, these meningiomas did not recur, and therefore obscured the prognostic importance of the MIB-1 value with the HL method. Focal accumulation of MIB-1-positive cells in meningiomas is not likely to correlate with their biologic aggressiveness. PMID- 11257620 TI - Microcystic adnexal carcinoma: an immunohistochemical study including markers of proliferation and apoptosis. AB - Microcystic adnexal carcinoma (MAC) is the prototype for a subset of locally aggressive adnexal carcinomas (LAACs). Ultraviolet radiation (UVR) and UVB signature p53 mutations are implicated in the etiology of the most common cutaneous carcinomas. However in MACs, the role of UVR and p53 mutations is unknown. In addition, controversy still exists regarding the patterns of differentiation within these tumors. The objective of this study was to determine the expression patterns of immunohistochemical markers for p53, Ki-67, c-erbB-2, and Bcl-2 in MACs, and to compare these patterns with two MAC histologic stimulants: sclerosing type basal cell carcinomas (sBCCs) and desmoplastic trichoepitheliomas (dTEs). Other objectives were to compare expression patterns of cytokeratin (CK) AE1/AE3, CK7, CD20, endothelial membrane antigen (EMA), Ber EP4, CD34, alpha-smooth muscle actin (SMA), and S-100 protein in MACs with its histologic simulators, and to determine the usefulness of all the immunohistochemical studies in diagnosis. Immunohistochemical markers were performed on 10 MACs, 10 sBCCs, and four dTEs. They included p53, Ki-67, c-erbB 2, Bcl-2, CK AE1/AE3, CK7, CD20, EMA, Ber-EP4, CD34, S-100 protein, and alpha SMA. MACs expressed p53 in less than 25% of the tumor cells in only two cases (20%), and both cases showed only moderately intense staining, whereas 80% of the sBCCs were positive and showed intense staining, and all dTEs were negative. In MACs, less than 5% of the tumor cells were Ki-67 positive, whereas the sBCCs showed 20% to 40% Ki-67-positive tumor cells and dTEs showed rare Ki-67-positive cells. Bcl-2 was expressed focally in MACs, diffusely in sBCCs, and in scattered cells in dTEs. All tumors were negative for c-erbB-2. CD34, CK7, EMA, Ber-EP4, S 100 protein, and alpha-SMA all showed a distinctive pattern of staining in MACs. Although MACs arise commonly in chronically sun-exposed skin, increased expression of p53 is not found frequently. Overexpression of c-erbB-2 does not appear to be a factor in the development and progression of these adnexal tumors. Bcl-2 is expressed in MACs, but not diffusely as in sBCCs. The low level of Ki-67 supports a low proliferative rate, and other immunohistochemical markers support divergent patterns of adnexal differentiation in MACs. Immunohistochemical studies may help to differentiate MAC from sBCCs and dTEs. PMID- 11257622 TI - Prognostic significance of pulmonary lymphangioleiomyomatosis histologic score. AB - Correlations were made between clinical and follow-up data and histopathologic findings in 105 women (mean age +/- standard deviation, 38.3 +/- 9.0 years) with pulmonary lymphangioleiomyomatosis (LAM). The actuarial survival (to pulmonary transplantation or death) of the patients from the time of lung biopsy was 85.1% and 71.0% after 5 and 10 years respectively. The histologic severity of LAM, graded as a LAM histologic score (LHS), was determined on the basis of semiquantitative estimation of the percentage of tissue involvement by the two major features of LAM: the cystic lesions and the infiltration by abnormal smooth muscle cells (LAM cells) in each case: LHS-1, <25%; LHS-2, 25% to 50%; and LHS-3, >50%. Analysis using the Kaplan-Meier method revealed significant differences in survival for patients with LHS-1, -2, and -3 (p = 0.01). The 5-and 10-year survivals were 100% and 100% for LHS-1, 81.2% and 74.4% for LHS-2, and 62.8% and 52.4% for LHS-3. Increased degrees of accumulation of hemosiderin in macrophages also were associated with higher LHS scores (p = 0.029) and a worse prognosis (p = 0.0012). Thus, the current study suggests that the LHS may provide a basis for determining the prognosis of LAM. PMID- 11257623 TI - Reticular perineurioma: a distinctive variant of soft tissue perineurioma. AB - Soft tissue perineurioma is a relatively recently characterized, uncommon tumor composed of perineurial cells exhibiting immunoreactivity for epithelial membrane antigen (EMA). These lesions occur preferentially in adults and may arise in a wide variety of anatomic sites. We report the clinicopathologic, immunohistochemical, and ultrastructural features of six cases of a poorly recognized morphologic variant of soft tissue perineurioma, characterized by a highly distinctive reticular growth pattern. Four of the patients were women, two were men (age range, 34-61 yrs; median, 43 yrs). Four of the cases arose in the subcutis of the upper extremity; three were located distally (thumb, finger, palm), whereas one was situated more proximally near the elbow region. One case each was located in the gingiva and subcutaneous tissue of the inguinal region, respectively. In those cases in which clinical information was available (n = 5), the lesions were asymptomatic and had been present from 4 months to 10 years before resection. Tumor size ranged from 1.5 cm to 10 cm (median size, 4.25 cm). Microscopically the lesions demonstrated a predominantly lace-like or reticular growth pattern composed of anastomosing cords of fusiform cells with bipolar cytoplasmic processes and palely eosinophilic cytoplasm. Nuclei were centrally placed, ovoid to fusiform in shape, and no mitoses were seen. Transition to more cellular areas was focally present in all cases. The stroma was variably collagenous to myxoid. Immunohistochemically all six cases stained positively for EMA but not for S-100 protein. Two cases demonstrated focal positive cytoplasmic staining for cytokeratin, whereas one case was focally desmin positive. Ultrastructural examination of two tumors showed typical features of perineurial cells. Follow up (available in only two cases) showed no evidence of recurrence. Reticular perineurioma of soft tissue represents an unusual morphologic variant within the perineurioma group, which should be distinguished from myoepithelial tumors, extraskeletal myxoid chondrosarcoma, and myxoid synovial sarcoma. PMID- 11257624 TI - Dysembryoplastic neuroepithelial tumor-like neoplasm of the septum pellucidum: a lesion often misdiagnosed as glioma: report of 10 cases. AB - The authors report a series of 10 low-grade neoplasms arising in the midline anteriorly in the region of the septum pellucidum with many of the histologic features of dysembryoplastic neuroepithelial tumor (DNT). The patients (five female, five male) ranged in age from 6 to 35 years (mean age, 21.5 years). The most common presenting symptoms were headache, nausea and vomiting, and visual disturbances. Radiographically, the tumors extended into the lateral ventricles from the septal region and obstructed the foramen of Monro. Varying degrees of hydrocephalus were present. The lesions were lobular, well-delineated, hypointense to brain on T1-weighted magnetic resonance imaging, and hyperintense on T2-weighted images. They were uniformly nonenhancing or showed only minimal peripheral enhancement. The tumors, in aggregate, had the histologic features of DNT. These included a mucin-rich background, oligodendrocyte-like cells, "floating neurons," and a "specific glioneuronal element." Seven patients underwent gross total resection and two underwent subtotal resection. No patients received adjuvant chemotherapy or radiotherapy. On follow-up (n = 6; median, 14 months), all tumors had either not recurred or were radiologically stable. On the basis of both neuroimaging and histopathology, DNT-like lesions should be considered in the differential diagnosis of midline intraventricular tumors in children and young adults. Distinction from more aggressive neoplasms is essential because these tumors appear to behave in a benign fashion. PMID- 11257625 TI - Hyperplastic polyps of the stomach: associations with histologic patterns of gastritis and gastric atrophy. AB - Hyperplastic polyps are common gastric lesions characterized by hyperplastic foveolae with variable amounts of inflamed stroma. Their pathogenesis is unknown, but they have been reported to occur in association with various forms of chronic gastritis, particularly autoimmune gastritis and Helicobacter pylori gastritis. Comprehensive histologic evaluation of the background mucosal pathology in patients with hyperplastic polyps has not been previously performed. We studied 160 patients with gastric hyperplastic polyps and characterized endoscopic and histologic features of the polyps (i.e., location, multiplicity, and presence of dysplasia and adenocarcinoma) and the background gastric mucosa (i.e., intestinal metaplasia, dysplasia, carcinoma, and presence and classification of gastritis). Hyperplastic polyps were most common in the antrum (60%) and were multiple in 20% of patients. Focal intestinal metaplasia of the polyp was present in 16% and dysplasia in 4% of patients. Only one patient (0.6%) had adenocarcinoma within the polyp. Evaluation of the surrounding gastric mucosa showed at least focal intestinal metaplasia in 37% of patients, adenoma or low-grade flat epithelial dysplasia in 2%, and synchronous or metachronous adenocarcinoma in 6%. Eighty five percent of patients had inflammatory mucosal pathology, most commonly active chronic H. pylori gastritis (25%), reactive or chemical gastropathy (21%), and metaplastic atrophic gastritis of the autoimmune (12%) or environmental (8%) type. These results indicate a strong association between various forms of gastritis and the development of hyperplastic polyps and further emphasize the importance of biopsy of the nonpolypoid gastric mucosa during endoscopic examination. PMID- 11257626 TI - The role of Yersinia enterocolitica and Yersinia pseudotuberculosis in granulomatous appendicitis: a histologic and molecular study. AB - Granulomatous appendicitis is an enigmatic entity. Purported causes include Crohn's disease, foreign body reactions, sarcoidosis, and infectious agents; however, most cases remain idiopathic. Yersinia enterocolitica (YE) and Y. pseudotuberculosis (YP) have been implicated as causes of appendicitis, ileocolitis, and mesenteric adenitis. The authors examined the potential role of YE and YP in granulomatous appendicitis using histologic and molecular methods. Forty cases of granulomatous appendicitis were evaluated for histologic features including transmural inflammation, number and character of granulomas, and mucosal changes. Twort Gram, Grocott methenamine-silver (GMS), and Ziehl-Neelsen stains were evaluated, and polymerase chain reaction (PCR) analysis was performed to identify pathogenic YP and YE. Twenty-five percent (10 of 40) of the cases were positive for pathogenic Yersinia by PCR (four YE, four YP, and two with both species). Prominent histologic features included epithelioid granulomas with lymphoid cuffing, transmural inflammation with lymphoid aggregates, mucosal ulceration, and cryptitis. One Yersinia-positive case contained mural Gram negative bacilli; fungal and acid-fast bacilli stains were all negative. Except for one culture-negative case, serologies and cultures were not done or results were unavailable. Two Yersinia-positive patients were diagnosed subsequently with Crohn's disease, suggesting a possible relationship between the two entities. No other patients developed significant sequelae. YE and YP are important causes of granulomatous appendicitis, and Yersinia infection may mimic Crohn's disease. No histologic features distinguish reliably between Yersinia species, or between Yersinia-positive and Yersinia-negative cases. Because special stains and cultures are often not diagnostic, PCR analysis is an excellent technique for the diagnosis of Yersinia. PMID- 11257627 TI - Lymphoepithelioma-like cholangiocarcinoma: an Epstein-Barr virus-associated tumor. AB - Epstein-Barr virus (EBV) has been linked to carcinomas of several body sites, especially of the nasopharynx, salivary gland, lung, and stomach. We present five cases of lymphoepithelioma-like cholangiocarcinoma, including one that had been previously reported. Two patients were men and three were women. Their ages ranged from 42 to 66 years. Histologically, all five tumors were composed of variable proportions of undifferentiated epithelial cells and glandular components in a lymphocyte-rich stroma. EBV was detected in all five tumors by in situ hybridization for EBER-1 in both lymphoepithelioma-like carcinoma (LELC) and glandular parts, but not in 36 cases of cholangiocarcinoma without the LELC component. Taken together, these observations indicate that lymphoepithelioma like cholangiocarcinoma is strongly linked to EBV. The LELC type of cholangiocarcinoma, like LELC of other body sites, may be more common in areas with endemic EBV infection. PMID- 11257628 TI - Juvenile xanthogranuloma of peripheral nerve: a report of two cases. AB - As a rule, juvenile xanthogranuloma (JXG) is a cutaneous lesion most often occurring in infancy. An inflammatory process of unknown etiology, it is self limited and benign in nature. The spectrum of JXG has expanded to include adult examples, multifocal lesions, and ones arising at extracutaneous locations. Although a variety of extracutaneous sites may be affected, few reported lesions have involved cranial or peripheral nerves. Solitary examples have been reported in trigeminal nerve and spinal nerve root; affected individuals were children or adolescents. An optic nerve lesion has also been described. We describe two additional cases of JXG of nerve. One patient developed multiple dorsal nerve root lesions, as well as skin involvement. The other case featured isolated involvement of the left radial nerve. Both patients were adults with no known underlying systemic disorder. These cases further expand the spectrum of extracutaneous JXG, and underscore its consideration in the differential of nerve "tumors." PMID- 11257629 TI - Pathologists and the judicial process: how to avoid it. AB - This review article covers the full range of issues concerning malpractice as it relates to pathologists. Following a brief summary as to the incidence and general statistics on the outcome of lawsuits as well as common pathology misdiagnoses resulting in lawsuits, the definition of malpractice is discussed. These include duty, breech of standard of care, proximal cause, and damage. Details are provided as to what a pathologist should do from the initial threat of a lawsuit, to the initial lawsuit, and through the initial physician/lawyer meeting. An in-depth analysis as to how pathologists should handle themselves through the discovery process and, in particular, deposition is provided. Plaintiff attorneys' goals at deposition are covered in depth. These goals include: 1) education about the pathologist's case and strategies; 2) impeachment of the pathologist's credibility; and 3) judgment as to how effective a witness the pathologist will be at trial. Various types of plaintiff's attorney at deposition are summarized. Also discussed is the post-deposition meeting with the legal representative, whether to settle, and specific issues relating to trial. Finally, general tips on how to avoid a lawsuit in pathology are reviewed. PMID- 11257630 TI - Molecular genetic characterization of primary cutaneous B-cell lymphomas. PMID- 11257631 TI - Clinical and pathological overlap in nonsteroidal anti-inflammatory drug-related small bowel diaphragm disease and the neuromuscular and vascular hamartoma of the small bowel. PMID- 11257632 TI - The histologic spectrum and clinical outcome of refractory and unclassified sprue. PMID- 11257633 TI - Signet-ring cells associated with pseudomembranous colitis. PMID- 11257634 TI - Use of the Van Nuys Ductal Carcinoma In Situ classification. PMID- 11257635 TI - Thyroid transcription factor-1 is of limited value in practical distinction between pulmonary and extrapulmonary small cell carcinomas. PMID- 11257636 TI - Deciduoid mesothelioma of the pleura after radiation therapy for Hodgkin's disease presenting as a mediastinal mass. PMID- 11257638 TI - Are desmoid tumors kit positive? PMID- 11257637 TI - Benign and malignant mesothelial proliferations. PMID- 11257639 TI - Mucin production in squamous intraepithelial lesions (CIN) of the cervix. PMID- 11257640 TI - Current bladder tumor tests: does their projected utility fulfill clinical necessity? AB - PURPOSE: We reviewed currently available bladder cancer tests in the context of the clinical expectations of a noninvasive bladder cancer test. MATERIALS AND METHODS: We reviewed the literature on bladder cancer tests that are commercially available or have shown clinical usefulness and examined how each test compares with standard methods of bladder cancer diagnosis. RESULTS: The clinical necessity for a noninvasive test for bladder cancer is 2-fold, including the early detection of high grade bladder tumors before muscle invasion and monitoring tumor recurrence or new onset. An ideal noninvasive test should be sensitive, specific, rapid, technically simple and have low intra-assay and interassay variability. Urine cytology has high specificity but limited applicability due to its relatively low sensitivity and subjective nature. Hematuria detection by Hemastix dipstick is sensitive but not specific for detecting bladder cancer. Molecules associated with bladder tumor growth and progression may serve as a basis for designing noninvasive diagnostic tests. The Food and Drug Administration approved BTA Stat and BTA TRAK tests, which detect human complement factor H and a related protein in urine, have 60% to 80% sensitivity and 50% to 70% specificity (lower in symptomatic patients) for bladder cancer. The Food and Drug Administration approved NMP22 test, which measures the level of nuclear mitotic apparatus protein in urine, has 50% to 100% sensitivity and 60% to 90% specificity. Accu-Dx detects fibrin degradation products, fibrin and fibrinogen in urine, although this test is no longer commercially available. The Immunocyt test combines cytology with an immunofluorescence technique to improve the sensitivity of cytology for detecting low grade tumors. The telomeric repeat amplification protocol assay for telomerase in exfoliated cells has 70% to 86% sensitivity and 60% to 90% specificity for bladder cancer. However, the low stability of telomerase in urine affects its sensitivity. The hyaluronic acid and hyaluronidase (HA-HAase) test, which measures the urinary level of hyaluronic acid and hyaluronidase, has 90% to 92% sensitivity and 80% to 84% specificity for bladder cancer. Quanticyt karyometry evaluates nuclear shape and DNA content of exfoliated cells to detect bladder cancer. The list of bladder tumor markers is growing rapidly and large multicenter trials are essential to assess their usefulness. CONCLUSIONS: Although currently noninvasive bladder cancer tests cannot replace cystoscopy, some have shown a promise of being clinically useful. One or a combination of these tests-markers may prove to be a prostate specific antigen for bladder cancer provided that patients and, more importantly, clinicians accept it. PMID- 11257641 TI - Robotic assisted, laparoscopic pelvic lymph node dissection in humans. AB - PURPOSE: We evaluate the feasibility and efficacy of robotic assisted, laparoscopic pelvic lymph node dissection for locally advanced prostate cancer staging. MATERIALS AND METHODS: Robotic assisted, laparoscopic pelvic lymph node dissection was performed in 10 consecutive patients with mainly T3 M0 prostatic carcinoma (robotic group). Operative, postoperative and pathological parameters were compared with the results of the last 10 patients undergoing conventional, laparoscopic pelvic lymph node dissection performed with similar indications by the same operator (laparoscopy group). RESULTS: All operations were performed according to the established protocol with no specific intraoperative or postoperative complications. No conversion was required, and no technical incidents were observed in the robotic group. Mean operating time plus or minus standard deviation for the robotic group was 125 +/- 57 minutes (range 75 to 215), significantly longer than that for the laparoscopy group, which was 60 +/- 15 minutes (p = 0.0013). In the robotic group 2 patients presented with postoperative lymphoceles revealed in 1 by deep venous thrombosis and in the second by obturator pain. In the laparoscopy group 1 patient presented with acute urinary retention. The histological results concerning the number of lymph nodes removed were similar in both groups (p = 0.5). CONCLUSIONS: We show the technical feasibility of robotic assisted, laparoscopic pelvic lymph node dissection in humans. Although the benefit of this technique has not yet been established, predictable technological improvements would suggest the development of telesurgery and an improved precision of surgical procedure. PMID- 11257642 TI - Prospective comparison of nonenhanced helical computerized tomography and Doppler ultrasonography for the diagnosis of renal colic. AB - PURPOSE: We evaluate the accuracy of nonenhanced helical computerized tomography (CT) and Doppler ultrasonography for the diagnosis of renal colic. MATERIALS AND METHODS: Our study includes 109 patients, with 218 kidneys, who presented with unilateral flank pain. All patients underwent nonenhanced helical CT, Doppler ultrasonography and excretory urography (IVP). CT was evaluated for the presence of ureteral stones and manifestation of ureteral obstruction. For Doppler ultrasonography the renal resistive index was measured for the left and right kidneys in each patient, and change in resistive index between ipsilateral and contralateral kidneys was calculated and considered positive for ureteral obstruction with values 0.04 or greater. As a reference standard, absence of obstruction was considered if IVP was negative and the cause of flank pain was confirmed not to be urological. Obstruction was diagnosed not only by positive IVP, but also by patient followup until passage or retrieval of ureteral stones. Results of CT and change in resistive index were compared with those of the reference standard. RESULTS: Unilateral ureteral obstruction was confirmed in 52 patients, while no obstruction was found in 57. Of the 57 patients without ureterolithiasis the change in resistive index results was negative in all patients with a specificity of 100%, while CT was negative in 55 with a specificity of 96%. Of the 52 patients with ureteral obstruction CT was positive in 50, and change in resistive index was positive in 47 with a sensitivity of 96% and 90%, respectively, with a difference of no significant value. CONCLUSIONS: Nonenhanced helical CT and change in resistive index are sensitive and specific tests that can contribute significantly to the diagnosis of acute unilateral renal obstruction. They can replace IVP, particularly in situations in which it is undesirable. PMID- 11257643 TI - Correlation of unilateral urolithiasis with sleep posture. AB - PURPOSE: Recurrent stone formers commonly present with calculi on the same side and the etiology of recurrent unilateral urolithiasis is unclear. Despite comprehensive metabolic evaluations, many patients will not be readily categorized into a treatable group. Data from the literature support that sleep posture may result in alterations of renal hemodynamics. We investigate the correlation of sleep posture with unilateral urinary stone formation. MATERIALS AND METHODS: A prospective study of 110 patients with recurrent unilateral nephrolithiasis was conducted. A questionnaire was used to evaluate patient sleep posture. Right or left side down and rotisserie-like sleep postures were defined. The side of stone formation was correlated with sleep posture using chi-square test. RESULTS: Of the patients 93 slept consistently with 1 side in a dependent position and the side of stone was identical to the dependent sleep side in 76% (p = 0.008). The positive predictive values of right and left side down sleep posture for formation of ipsilateral calculi were 82% and 70%, respectively. CONCLUSIONS: Although the exact pathophysiology of the association between sleep posture and recurrent unilateral stone disease remains to be elucidated, sleep posture may alter renal hemodynamics during sleep and promote stone formation. This observation needs further investigation and should be factored into the evaluation and prevention of unilateral urinary stone disease. PMID- 11257644 TI - Percutaneous nephrostomy versus ureteral stents for diversion of hydronephrosis caused by stones: a prospective, randomized clinical trial. AB - PURPOSE: Urinary diversion with percutaneous nephrostomy or ureteral stent is indicated by symptoms, such as persistent colic, high temperature and uremia, of hydronephrosis caused by stones. We evaluate which of these 2 methods is superior concerning the course of procedure, relief of accompanying symptoms and quality of life in regard to patient age and sex. MATERIALS AND METHODS: A total of 40 patients with stone induced hydronephrosis were randomized into either percutaneous nephrostomy or stent insertion groups. These patients were then evaluated by procedure (use of analgesics, x-ray exposure, success of insertion), relief of accompanying symptoms (duration of diversion, intravenous administration of antibiotics for high temperature) and quality of life (questionnaire immediately and 2 to 4 weeks postoperatively). RESULTS: Two comparable groups of patients were formed, with an average age of 55 versus 49 years and a male-to-female ratio of 12:8 versus 9:11 for those who underwent percutaneous nephrostomy versus those who received a stent, respectively. Percutaneous nephrostomy was successfully completed in 100% of patients and stents were successful in 80%, with a 20% conversion to percutaneous nephrostomy. The x-ray exposure was shorter in the percutaneous nephrostomy group (p = 0.052). Administration of analgesics was more frequent in the stent group (p = 0.061). Percutaneous nephrostomy indwelling time was shorter (50% less than 2 weeks) than that of stents (25% less than 2 weeks, p = 0.043). Antibiotics were administered for greater than 5 days in 0% of patients who underwent percutaneous nephrostomy versus 64% in those with stents (p = 0.174). Reduction in quality of life was moderate but more pronounced in patients with stents compared to those who underwent percutaneous nephrostomy, and was more distinct in males and younger patients. The quality of life progressively improved in the course of diversion with percutaneous nephrostomy but deteriorated with stents. CONCLUSIONS: Our results demonstrated that percutaneous nephrostomy is superior to ureteral stents for diversion of hydronephrosis caused by stones, especially in patients with a high temperature, as well as in males and juveniles. PMID- 11257645 TI - Laparoscopic bilateral synchronous nephrectomy for autosomal dominant polycystic kidney disease: the initial experience. AB - PURPOSE: We report our experience with laparoscopic bilateral synchronous nephrectomy for giant symptomatic autosomal dominant polycystic kidney disease (ADPKD) and compare outcome data with open bilateral nephrectomy. MATERIALS AND METHODS: Since March 1998, 10 patients underwent bilateral synchronous laparoscopic nephrectomy for giant symptomatic ADPKD. A 3 port retroperitoneal laparoscopic approach was used to secure the renal hilum and mobilize the kidney. Intact specimen extraction was performed through a midline infraumbilical extraperitoneal incision. The patient was then repositioned for the contralateral retroperitoneoscopic nephrectomy, with the second specimen also delivered through the same infraumbilical incision. Data were retrospectively compared with 10 patients who had undergone bilateral synchronous open nephrectomy for ADPKD between 1981 and 1992. RESULTS: Patients in the laparoscopic and open groups were comparable in regard to age (53 versus 47 years, p = 0.54) and Anesthesiologist Society of America class (3 versus 3, p = 0.84) but patients in the laparoscopic group were significantly more obese (body mass index 35.9 versus 23.8, p = 0.02). For comparable total specimen weights (3 versus 3 kg, p = 0.69) surgical time was longer in the laparoscopic group (4.4 versus 3.8 hours, p = 0.007). However, the laparoscopic group was superior in regard to blood loss (150 versus 325 cc, p = 0.05), postoperative requirement of nasogastric tube (10% versus 100%, p = 0.0001), narcotic analgesics (34.2 versus 120.4 mg. morphine sulfate equivalent, p = 0.03) and hospital stay (1.5 versus 9 days, p = 0.004). Complications occurred in 5 patients (50%) in the laparoscopic group and 4 (40%) in the open group (p = 0.66). No laparoscopic case was converted to open surgery. CONCLUSIONS: Synchronous bilateral retroperitoneal laparoscopic nephrectomy for giant symptomatic adult polycystic kidney disease is feasible, safe and efficacious, and can be performed either before or after renal transplantation. Compared to open surgery, the laparoscopic approach results in significantly shorter hospital stay, decreased morbidity and quicker recovery. Laparoscopy is currently our technique of choice in this setting. PMID- 11257646 TI - Retroperitoneoscopy assisted live donor nephrectomy: the Yonsei experience. AB - PURPOSE: Retroperitoneoscopy assisted live donor nephrectomy has become standard based on our experience with 103 consecutive cases operated on between January 1993 and May 2000. We describe the advantages of retroperitoneoscopy assisted compared to laparoscopic live donor nephrectomy. MATERIALS AND METHODS: After performing more than 1,200 cases of open live donor nephrectomy (S. C. Y.), we combined our experience with open and laparoscopic surgery to develop a specific technique of minilaparotomy live donor nephrectomy. Operations were performed by 1 senior surgeon and 1 assistant, with the help of specially designed piercing abdominal and peritoneal retractors. A 5 to 7 cm. transverse pararectal skin incision is made at the level of 10th rib and the abdominal muscles are split without division. A 10 mm. port is placed at the lower abdomen to allow for the telescope. The procedure is performed extraperitoneally, combining open and laparoscopic instruments under direct vision. Renal pedicles and ureters are ligated using laparoscopic clips and sutures. The kidney is removed via laparotomy and the wound is closed. RESULTS: Average operating time for the 103 live donor nephrectomies was 130 minutes (range 85 to 210), and there was no case of kidney loss, open surgical conversion or blood transfusion. Mean warm ischemia time was 2.3 +/- 1.2 minutes and average incision length was 6.5 cm. (range 5.1 to 7.0). Postoperative pain was minimal and analgesics were generally not required by postoperative day 2. Patients were fully ambulatory a mean 1.5 days (range 1 to 3.5) postoperatively. CONCLUSIONS: Retroperitoneoscopy assisted live donor nephrectomy is not only feasible, but reproducible. Any surgeon with previous experience with conventional open live donor nephrectomy can perform this hybrid, minimally invasive procedure. PMID- 11257647 TI - The impact of a 4 cm. cutoff point for stratification of T1N0M0 renal cell carcinoma after radical nephrectomy. AB - PURPOSE: The 1997 TNM classification defines T1 tumors as those smaller than 7 cm. Recently, a cutoff point of 4 cm. has been proposed to create a subclass of T1 tumors. We evaluated the validity of this cutoff point by assessing the pathological findings and prognoses of patients with T1N0M0 renal cell carcinoma following radical nephrectomy. MATERIALS AND METHODS: We reviewed the hospital charts of 333 patients with T1N0M0 tumors, followed as long as 282 months (median 63) after radical nephrectomy. The validity of tumor size cutoff point for predicting survival outcome was tested in relation to other prognostic factors, including patient age, tumor position, nuclear grade, tumor histopathology and degree of microscopic venous invasion. RESULTS: During followup 32 patients (9.6%) had tumor recurrence and 21 (6.3%) died of renal cell carcinoma. A 5 cm. cutoff point maximized the differences in cancer specific survival rates and a 4 cm. cutoff point maximized the differences in disease-free survival rates. Tumor size was directly related to microscopic venous invasion and nuclear grade, which are significant prognostic factors, and a 4 cm. cutoff point enhanced these relationships. CONCLUSIONS: Tumor size is an important prognostic factor for patients with T1N0M0 renal cell carcinoma. A cutoff point of 4 cm. is practical for dividing the T1N0M0 classification into T1a and T1b subclasses. PMID- 11257648 TI - Botulinum toxin urethral sphincter injection to restore bladder emptying in men and women with voiding dysfunction. AB - PURPOSE: Botulinum toxin injection into the external urinary sphincter in spinal cord injured men with detrusor-sphincter dyssynergia has been reported. We expand the clinical use of botulinum toxin for a variety of bladder outlet obstructions and to decrease outlet resistance in patients with acontractile detrusor but who wish to void by the Valsalva maneuver. MATERIALS AND METHODS: Prospective treatment was performed for voiding dysfunction in 8 men and 13 women 34 to 74 years old. The reasons for voiding dysfunction included neurogenic detrusor sphincter dyssynergia in 12 cases, pelvic floor spasticity in 8 and acontractile detrusor in 1 patient with multiple sclerosis who wished to void by the Valsalva maneuver. Using a rigid cystoscope and a collagen injection needle, a total of 80 to 100 units of botulinum A toxin (Botox) were injected into the external sphincter at the 3, 6, 9 and 12 o'clock positions. RESULTS: Preoperatively 19 of 21 patients were on indwelling or intermittent catheterization. After botulinum A injection all but 1 patient were able to void without catheterization. No acute complications, such as general paralysis or respiratory depression, occurred and none of the patients had dribbling or stress urinary incontinence. Postoperative post-void residual decreased by 71% and voiding pressures decreased on average 38%. Of the 21 patients 14 (67%) reported significant subjective improvement in voiding. Followup ranges from 3 to 16 months, with a maximum of 3 botulinum A injections in some patients. CONCLUSIONS: Urethral sphincter botulinum injection should be considered for complex voiding dysfunction. Encouraging improvement without complications were seen in most of our patients. We have expanded the use of botulinum toxin to treat pelvic floor spasticity and also women. PMID- 11257650 TI - Mechanisms of prostatic stromal invasion in patients with bladder cancer: clinical significance. AB - PURPOSE: We assess the pathological mechanisms of silent prostatic stromal invasion in patients with bladder cancer for early detection and treatment. MATERIALS AND METHODS: Between August 1998 and January 1999, 10 patients with clinically organ confined transitional cell carcinoma of the bladder and known prostatic stromal invasion on transurethral biopsy or who were high risk for prostatic involvement due to tumor location near the bladder neck were studied for histological patterns of prostatic invasion. There were 5 cystectomy specimens distended for 24 hours with formalin via a Foley catheter, then step sectioned longitudinally at 3 mm. intervals through the bladder neck and prostate. Standard hematoxylin and eosin staining methods were used and sections were analyzed by 2 pathologists. RESULTS: There were 3 separate patterns of prostatic stromal invasion elucidated, including 2 previously described methods of extravesical or intraurethral invasion into the prostatic stroma and a third one through the bladder neck directly into the prostatic stroma. The third pattern was not grossly evident on endoscopy or urethral biopsy before cystectomy. CONCLUSIONS: Longitudinal sectioning of the bladder neck and prostate of cystectomy specimens suggests tumors at the bladder neck may directly invade the prostatic stroma without histological evidence of extravesical or intraurethral spread. Such direct silent tumor invasion of the prostate by superficial or endoscopically inapparent tumor is difficult to detect clinically by current biopsy methods. New methods of detection are necessary. PMID- 11257649 TI - Cystectomy for bladder cancer: a contemporary series. AB - PURPOSE: To validate the current TNM staging system, we analyzed our contemporary experience with 300 cystectomies. MATERIALS AND METHODS: The pathological material and medical records of 300 patients treated with cystectomy were reviewed, and the new TNM classification was adopted. RESULTS: The median followup of patients with no evidence of disease was 65 months, and overall survival rate was 45% with a median survival of 50 months. In a Cox regression analysis only patient age, pT stage and neoadjuvant chemotherapy were significant factors for survival. The disease specific survival was 67% with a median survival of 94 months. In a multiple proportional hazards analysis only pT stage and previous chemotherapy were significant factors of disease specific survival. A significant difference was seen in the overall and disease specific survival between patients with organ confined and nonorgan confined tumors. We did not observe a difference in the survival rate among patients with pT4a to pT3 tumors. Significant differences were not seen in survival rates between sexes or among patients of different age groups. Transitional cell carcinoma was the predominant histological type, and no significant difference was found in patient outcome among the different histological subtypes. CONCLUSIONS: Bladder cancer can be categorized into organ confined and nonorgan confined tumors. This dichotomous grouping is better suited for evaluating adjuvant clinical trials. The pT stage of the bladder and prostate should be prospectively analyzed together to better define the clinical implications of prostatic involvement. In our opinion the histological subtypes do not affect outcome. PMID- 11257651 TI - Fluorescence endoscopy with 5-aminolevulinic acid reduces early recurrence rate in superficial bladder cancer. AB - PURPOSE: Several investigators have demonstrated an approximately 20% higher tumor detection rate by 5-aminolevulinic acid (ALA) fluorescence endoscopy compared to standard white light cystoscopy, and suggested a reduction in tumor recurrences when fluorescence endoscopy was performed as standard procedure during transurethral resection. We test this hypothesis. MATERIALS AND METHODS: In a prospective randomized multicenter study 102 patients underwent transurethral resection of bladder tumor(s) either with white light or ALA fluorescence assisted endoscopy. A second look transurethral resection with ALA fluorescence endoscopy was performed 6 weeks after the initial operation. RESULTS: At second look transurethral resection tumor was detected in 20 of 51 patients (39%) in the white light group and in 8 of 51 (16%) in the ALA fluorescence endoscopy group. This difference was statistically significant (p = 0.005). CONCLUSIONS: ALA fluorescence endoscopy is an innocuous and inexpensive diagnostic procedure that significantly improves bladder tumor detection rates compared to standard white light endoscopy. In our controlled study ALA fluorescence endoscopy reduced the residual tumor detection rate at second look transurethral resection by 59%. PMID- 11257652 TI - Stage progression in Ta papillary urothelial tumors: relationship to grade, immunohistochemical expression of tumor markers, mitotic frequency and DNA ploidy. AB - PURPOSE: We studied 363 patients with stage Ta bladder tumors during long-term followup who were classified according to the 1998 WHO and International Society of Urological Pathology consensus classifications. We determine whether various immunohistochemical and molecular markers could predict tumor progression. MATERIALS AND METHODS: A total of 680 patients in western Sweden with a first diagnosis of bladder carcinoma in 1987 and 1988 were registered and followed for at least 5 years. There were 363 (53%) tumors that were papillary stage pTa. The tumors were classified as papillary urothelial neoplasm of low malignant potential in 95 patients, low grade papillary urothelial carcinoma in 160 and high grade carcinoma in 108. Of the patients in the latter group 95 were subdivided into WHO grade 2 and 13 into WHO grade 3. Tissue from the primary tumors that progressed in stage during followup was further analyzed with immunohistochemical methods (p21, p53, Ki67 and pRb), DNA ploidy and mitotic frequency. The results were compared with those in matched controls (nonprogressors). RESULTS: Recurrence developed in 35% of patients with papillary urothelial neoplasm of low malignant potential compared to 71% with low grade urothelial carcinoma and 73% with high grade carcinoma (p <0.0001). No papillary urothelial neoplasm of low malignant potential progressed in stage. Disease progressed in 4% of patients with low grade compared to 23% with high grade carcinoma (p <0.0001). Of the patients with WHO grade 3 disease progressed in 45% compared to grade 2 in 20% (p <0.0011). At first diagnosis p53 score was significantly higher (p <0.0022) among patients with WHO grade 2 carcinoma which later progressed compared to that in matched controls but there was no significant difference regarding the other markers. In contrast to grade 2 most grade 3 carcinoma was aneuploid, had high mitosis frequency, high p53 and Ki67 scores as well as loss of retinoblastoma gene expression. CONCLUSIONS: The 1988 WHO and International Society of Urological Pathology consensus classifications divide noninvasive papillary bladder tumors into 3 subgroups with different clinical behavior, which seems to be an advantage compared with the 1973 WHO classification. A disadvantage is that the high grade carcinoma group contains 2 subgroups with different progression rates and immunohistochemical marker profiles, corresponding to the 1999 WHO grades 2 and 3. Grade 2 tumors in patients that progressed in stage years later seem to have different immunohistochemical and molecular marker profiles compared to those in matched controls. PMID- 11257653 TI - Substitution urethroplasty with buccal mucosal-free grafts. AB - PURPOSE: Buccal mucosal grafts and the Barbagli technique are recent developments in the treatment of urethral strictures. MATERIALS AND METHODS: We reviewed the results of and experience with urethroplasty using buccal mucosal graft in 128 patients. RESULTS: The re-stricture rate was 11% for patch grafts and 45% for tube grafts. There were no other complications. CONCLUSIONS: Buccal mucosal graft is at least as good as any other material for substitution urethroplasty with fewer complications. The 2-stage is more reliable than the stage 1 approach for circumferential reconstruction of the urethra. PMID- 11257654 TI - Urethral recurrence after radical radiotherapy for bladder cancer. AB - PURPOSE: Following cystectomy for bladder cancer, orthotopic reconstruction may result in a decreased risk of urethral recurrence compared to cutaneous diversion. We evaluate the rate of urethral recurrence after radical external beam radiotherapy. MATERIALS AND METHODS: We reviewed the records of 214 men who had received radical radiotherapy at a single center from 1990 to 1995. Patients treated with chemotherapy were excluded from study. RESULTS: A total of 214 men (median age 69 years, range 39 to 86) underwent radical radiotherapy for cure. Tumor stages were T1 in 7%, T2 in 41%, T3 in 42% and T4a in 10% of the patients. Median followup was 32 months (range 1 month to 8.4 years) and 5-year survival rate was 30%. Urethral recurrence developed in 7 (3.2%) cases and was detected within 18 months (median 10 months, range 3 months to 5 years) of followup in 5. In 2 of these 7 cases recurrence developed in the prostatic urethra, and when these 2 cases were excluded from analysis the recurrence rate decreased to 2.3%. A total of 64 men completed 5-year followup, with a 4.7% rate of urethral recurrence (3.1% excluding prostatic urethral recurrence). Multifocal disease, bladder neck involvement, prostatic disease and cis were possible risk factors for urethral recurrence. CONCLUSIONS: The risk of urethral recurrence after radical radiotherapy for transitional cell carcinoma of the bladder is comparable with that reported after orthotopic reconstruction. It is not possible to exclude completely that men at higher risk were offered cystectomy, but the data are consistent with the suggestion that continued contact with urine may be protective. PMID- 11257655 TI - Tumor stage, vascular invasion and the percentage of poorly differentiated cancer: independent prognosticators for inguinal lymph node metastasis in penile squamous cancer. AB - PURPOSE: We determine if histopathological factors of the primary penile tumor can stratify the risk of the development of inguinal lymph node metastases. MATERIALS AND METHODS: Clinical records of 48 consecutive patients with squamous cell carcinoma of the penis who underwent resection of the primary lesion and either inguinal lymph node dissection or were observed for signs of recurrence (median followup 59 months) were reviewed. Parameters examined included pathological tumor stage, quantified depth of invasion and tumor thickness, histological and nuclear grade, percentage of poorly differentiated cancer in the primary tumor, number of mitoses and presence or absence of vascular invasion. Variables were compared in 18 lymph node positive and 30 lymph node negative cases. RESULTS: Pathological tumor stage, vascular invasion and presence of greater than 50% poorly differentiated cancer were the strongest predictors of nodal metastasis on univariate and multivariate regression analyses. None of 15 pT1 tumors exhibited vascular invasion or lymph node metastases. Of 33 patients with pT2 or greater tumors 21 (64%) had vascular invasion and 18 (55%) had metastases. Only 4 of 25 patients (15%) with 50% or less poorly differentiated cancer in the penile tumor had metastases compared with 14 of 23 patients (61%) with greater than 50% poorly differentiated cancer (p = 0.001). No other variables tested were significantly different among the patient cohorts. CONCLUSIONS: Pathological stage of the penile tumor, vascular invasion and greater than 50% poorly differentiated cancer were independent prognostic factors for inguinal lymph node metastasis. Prophylactic lymphadenectomy in compliant patients with pT1 lesions without vascular invasion and 50% or less poorly differentiated cancer does not appear warranted. PMID- 11257656 TI - Predictive value of total and percent free prostate specific antigen in high grade prostatic intraepithelial neoplasia lesions: results of the Tyrol Prostate Specific Antigen Screening Project. AB - PURPOSE: We evaluate the predictive values of total and percent free prostate specific antigen (PSA) in regard to high grade intraepithelial lesions in volunteers who participated in the Tyrol PSA Screening Project. MATERIALS AND METHODS: Between June 1995 and December 1998, 1,474 patients undergoing transrectal biopsy of the prostate were evaluated. The primary detection rates of prostate cancer and high grade intraepithelial lesions were evaluated. In addition, the rate of prostate cancer detected on biopsy in patients diagnosed with high grade prostatic intraepithelial neoplasia on the previous biopsy was assessed. Mean total PSA values and mean percent free PSA levels were determined for each study group and compared using the Mann-Whitney U test. RESULTS: A total of 1,077 (73.1%) volunteers had benign prostatic hyperplasia or prostatitis, and 327 (22.2%) had prostate cancer. The primary detection rate for high grade intraepithelial lesions was 4.7% (70 patients) and on repeat biopsy was 38.6% (27). Mean total PSA for the benign prostatic hyperplasia, prostate cancer, high grade and intraepithelial cancer groups were 6.0, 8.7, 5.9 and 5.2 ng./ml., respectively. Mean percent free PSA values for the various groups were 21.9, 12.1, 15.0 and 12.0, respectively. In regard to total PSA there was a statistically significant difference between the prostate cancer and high grade prostatic intraepithelial neoplasia groups (p = 0.016), as well as the prostate cancer and intraepithelial cancer groups (p = 0.028). However, the high grade and intraepithelial cancer groups did not differ significantly. In regard to percent free PSA there were statistically significant differences between the prostate cancer and high grade prostatic intraepithelial neoplasia groups (p = 0.0001), and the high grade and intraepithelial cancer groups (p = 0.013). CONCLUSIONS: In regard to percent free PSA our data indicate a significant difference between high grade intraepithelial lesion and intraepithelial cancer. Due to a substantial overlap in percent free prostate specific antigen between the 2 groups, a clinically useful cutoff point could not be established. Therefore, we recommend repeat biopsy in all patients with high grade intraepithelial lesions regardless of the percent free PSA. PMID- 11257657 TI - Defining prostate specific antigen progression after radical prostatectomy: what is the most appropriate cut point? AB - PURPOSE: The most appropriate definition of biochemical progression after radical prostatectomy and radiation therapy is uncertain. We analyzed the effect of using various prostate specific antigen (PSA) end point definitions for defining biochemical progression after radical prostatectomy and attempted to determine the best PSA cut point to use. Aspects of the American Society for Therapeutic Radiology and Oncology (ASTRO) definition of biochemical failure after radiation therapy are also analyzed in our radical prostatectomy cases. MATERIALS AND METHODS: A total of 2,782 men with clinically localized prostate cancer (cT1-T2) who had undergone radical prostatectomy between 1987 and 1993 were reviewed. All patients had regular PSA determinations from surgery through followup. Analysis was limited to patients who did not receive adjuvant treatment within 90 days of radical prostatectomy. Biochemical, PSA progression-free percent after radical prostatectomy was determined by the Kaplan-Meier method using several PSA cut points, including 0.2, 0.3, 0.4 and 0.5 ng./ml. or greater, as well as 0.4 ng./ml. or greater and increasing. Progression-free percent was also assessed using the ASTRO definition, which is 3 increases in PSA. To determine which PSA level was most appropriate to define progression after radical prostatectomy, the percentage of patients with a continued PSA increase after reaching each cut point was determined. The relationship between the maximum PSA within 3 years of surgery and subsequent development of clinical disease was also assessed. RESULTS: Progression-free percent was dependent on the PSA cut point used. Biochemical progression-free percentages for cut points 0.2, 0.3, 0.4 and 0.5 ng./ml. or greater were 62%, 72%, 76% and 78% at 5 years, and 43%, 54%, 59% and 61% at 10 years, respectively. A subsequent increase in PSA was noted in 49%, 62% and 72% of patients who had PSA 0.2, 0.3 and 0.4 ng./ml., respectively. Subsequent clinical progression (local or systemic) was directly related to the maximum PSA attained within 3 years of radical prostatectomy (p=0.0001). Progression-free percent for definitions requiring multiple increases in PSA were dependent on when the event was said to occur. Backdating of events at or before the first PSA (ASTRO definition) resulted in poorer, short-term progression-free percent (78% at 5 years), with little apparent likelihood of long-term failure (78% at 10 years). Coding the event at the last PSA increase when all event criteria had been met resulted in more realistic progression-free percent estimates (85% at 5 and 59% at 10 years). CONCLUSIONS: Biochemical, PSA progression rates vary markedly depending on the method used to define PSA failure. Methods that require multiple increasing PSA values, for example the ASTRO definition, give misleading results, especially if the event time is backdated. Standards for defining PSA progression would allow more consistent and comparable progression estimates after radical prostatectomy. PSA 0.4 ng./ml. or greater may be the most appropriate cut point to use since a significant number of patients with lower PSA do not have a continued increase in it. PMID- 11257658 TI - The Willet F. Whitmore, Jr., Lectureship: blockade of epidermal growth factor receptors as anticancer therapy. PMID- 11257659 TI - Burn-out of urologists in the county of Schleswig-Holstein, Germany: a comparison of hospital and private practice urologists. AB - PURPOSE: Mental and physical burden of physicians, especially surgeons, is high. The degree of burn-out was estimated among urologists in the German federal county of Schleswig-Holstein, with special emphasis on differences related to age, qualification and hospital versus private practice. MATERIALS AND METHODS: The Maslach Burnout Inventory was mailed to all urologists and urologists in training registered in the county of Schleswig-Holstein to determine the frequency and intensity of the 3 burn-out subscales of emotional exhaustion, depersonalization and low personal accomplishment, together with a questionnaire covering demographic data. RESULTS: Of 128 urologists 75 (58.6%) replied. Levels of burn-out in the subscales of emotional exhaustion and depersonalization were increased among hospital urologists, urologists in training and urologists younger than 45 years. Urologists in private practices, fully trained urological specialists and urologists older than 45 years showed a low degree of burn-out, corresponding to normal values, whereas young urologists in training and working in hospitals had the highest risk of burn-out. The personal accomplishment level was generally high in all groups. CONCLUSIONS: The constellation of being a urologist in private practice and older than 45 years appears to provide some protection against burn-out that might be due to a more personal relationship to the patients, lesser hierarchical situation and workload related income. In times of increasing pressure on the health care system, the personal situation of physicians in training should not be overlooked. PMID- 11257660 TI - Renal artery embolization for benign obstructive uropathy. AB - PURPOSE: We demonstrate safety of renal artery embolization as an alternative to nephrectomy in symptomatic patients with obstructed, poorly functioning kidneys who are unfit for surgery. MATERIALS AND METHODS: We performed renal artery embolization in 2 medically unfit patients with benign obstructive uropathy and poorly functioning kidneys. The affected kidney was initially drained with a nephrostomy tube and the kidney was then selectively embolized using absorbable gelatin sponge particles and Gianturco coils. The nephrostomy tube was removed before hospital discharge. RESULTS: Urine production by the affected kidney ceased and symptoms resolved. CONCLUSIONS: Renal artery embolization with antegrade drainage is a safe alternative to surgery in unfit symptomatic patients with benign obstructive uropathy and poorly functioning kidneys. PMID- 11257661 TI - The use of a lithoclast probe for ureterorenoscopic coagulation of bleeding ureteral cancer. AB - PURPOSE: We describe use of the lithoclast device, normally used for electrohydraulic stone fragmentation, for safe ureteroscopic intraureteral coagulation of a bleeding tumor. MATERIALS AND METHODS: Intraureteral ureteroscopic coagulation of a bleeding transitional cell carcinoma was performed in an 86-year-old high risk patient. A 0.8 mm. probe of the lithoclast device was connected to high frequency current. To avoid short circuit to the ureteroscope the probe was covered with a standard 5Fr ureteral catheter, which served as a perfect isolator. RESULTS: The bleeding intraureteral tumor was coagulated successfully under direct vision. CONCLUSIONS: The lithoclast probe in combination with a standard ureteral catheter can be used for intraureteral coagulation of bleeding tumors. It is an inexpensive device that is often available in nonuniversity departments. PMID- 11257662 TI - Post-cesarean cervicovesical fistula: technique of laparoscopic repair. AB - PURPOSE: We describe the technique of laparoscopic repair of post-cesarean cervicovesical fistula. MATERIALS AND METHODS: Two patients 30 and 35 years old, respectively, presented with urinary incontinence, amenorrhea and menouria. Both patients had undergone previously a second cesarean section. Diagnosis was post cesarean cervicovesical fistula, which was repaired laparoscopically. RESULTS: Laparoscopic repair was successfully completed in 1 patient. In the other case, after complete dissection of the fistula and preparation of the edges for suturing, surgery had to be converted to an open procedure due to inability to maintain a pneumoperitoneum. CONCLUSIONS: Laparoscopy is a viable option for the repair of post-cesarean cervicovesical fistula with all its attendant advantages. PMID- 11257663 TI - A novel technique for ventriculovesical shunting of congenital hydrocephalus. AB - PURPOSE: Hydrocephalus is typically treated with a ventriculoperitoneal or ventriculoatrial shunt. However, shunt malfunction, recurrent infection or other co-morbidities occasionally make these shunts inappropriate. As early as 1925 a ureterodural anastomosis was used to divert cerebrospinal fluid into the urinary system. Since then techniques for cerebrospinal fluid urinary diversion have improved. We designed a new technique to drain cerebrospinal fluid into the urinary system surgically while averting some of the problems encountered with previous methods. MATERIALS AND METHODS: We describe a new technique to perform a ventriculovesicular shunt, which we performed on a 17-year-old woman with congenital hydrocephalus. To put this unique operation into perspective we briefly review the history of cerebrospinal fluid urinary shunting. RESULTS: Our results were promising, with no evidence of distal shunt malfunction or infection postoperatively. CONCLUSIONS: Our novel technique of a ventriculovesical shunt with a polyester cuff that provides stabilization and infection control holds promise for patients with hydrocephalus who are not candidates for a ventriculoperitoneal or ventriculoatrial shunt. PMID- 11257664 TI - The use of an artificial urinary sphincter in women with type III incontinence and a negative Marshall test. AB - PURPOSE: We evaluate the efficacy of the AMS 800 artificial urinary sphincter in women with type III incontinence. MATERIALS AND METHODS: We enrolled 207 women with genuine stress incontinence due to intrinsic sphincter deficiency. Primary inclusion criterion was a negative Marshall test. A modified surgical procedure was used to implant the AMS 800 through an abdominal approach, with placement of the cuff around the bladder neck between the periurethral fascia and vagina. Followup data were available for 206 women, including 179 with nonneurogenic and 27 with neurogenic bladders (mean followup 3.9 years). RESULTS: There were 12 (5.9%) explantations due to the prosthesis either through an erosion, extrusion or both that were necessary. The only significant risk factor for explantation was perioperative injury. This injury resulted in 8 explantations in 49 patients compared with 4 in 155 who did not have such injuries (p = 0.0016). Of the 190 patients with working devices continence was achieved in 88.7% (49 of 168) and 81.8% (18 of 22) of those with nonneurogenic and neurogenic bladders, respectively. Social continence (slight leakage but no pad use) was reported by 7.7% (13 of 168) and 9.1% (2 of 22) of patients in the nonneurogenic and neurogenic groups, respectively. The remaining patients reported leakage and pad use. CONCLUSIONS: The AMS 800 can be used successfully to treat women with genuine stress incontinence due to intrinsic sphincter deficiency. The modified surgical approach resulted in fewer perioperative injuries and, consequently, a low explantation rate. Women with genuine stress incontinence, a low urethral closure pressure and negative Marshall test indicating severe intrinsic sphincter deficiency are potential candidates for artificial urinary sphincter implantation. PMID- 11257665 TI - Randomized comparison of local versus epidural anesthesia for tension-free vaginal tape operation. AB - PURPOSE: We determine the difference between local anesthesia and epidural blockade for the tension-free vaginal tape operation. MATERIALS AND METHODS: Between November 1995 and November 1997, 73 women who had genuine stress incontinence in the absence of pelvic prolapse underwent a prospective randomized study. The study was conducted using a standardized protocol for different types of anesthesia for the tension-free vaginal tape procedure. A formal pain scale was used to determine the pain score for the patients during the operation. Additionally an anxiety scale was used to measure the anxiety level of the subjects immediately after admission to the ward and before discharge from the hospital. RESULTS: One woman was excluded from study due to loss at followup. The comparisons of pain score, duration of procedure and anxiety level of the 2 different types of anesthesia were not significantly different in the 72 study subjects. There was no significant difference in the amount of blood loss, while initial spontaneous voiding occurred significantly earlier (3.5 +/- 2.3 versus 5.8 +/- 0.1 hours, p <0.01), the number of patients in whom initial spontaneous voiding occurred more than 6 hours postoperatively was fewer (2 versus 10, p =0.01), amount of post-void residual during hospitalization was significantly less (98 +/- 63 versus 155 +/- 56 ml., p <0.01) and length of hospital stay was significantly shorter (3.4 +/- 1.4 versus 5.5 +/- 1.6 days, p <0.01) in the local anesthesia compared to epidural group. Subjective and objective success rates were not significantly different in these 2 groups. CONCLUSIONS: Both anesthetic methods can be equally effectively used for the tension-free vaginal tape operation. Local may be better than epidural anesthesia but its clinical significance needs to be proved by further study. PMID- 11257666 TI - Measurement of vaginal and minor labial oxygen tension for the evaluation of female sexual function. AB - PURPOSE: Female sexual dysfunction is a new, rapidly expanding area of sexual medicine. Female sexual arousal disorder may, in part, be due to decreased pelvic blood flow. Therefore, we developed a simple noninvasive reproducible technique to measure vaginal and minor labial blood flow. MATERIALS AND METHODS: The study included 12 healthy young women able to have orgasm through self-stimulation. Observations at orgasm were recorded in the 12 subjects after self-stimulation. Measurements were obtained intravaginally and on the minor labia using a modified Clark oxygen electrode to obtain partial oxygen pressure (pO(2)). RESULTS: Mean basal vaginal value was 3.8 +/- 0.9 mm Hg and mean basal pO(2) on the minor labia was 18.3 +/- 3.7 mm. Hg. As soon as self-stimulation was initiated an increase in oxygen tension occurred and continued during sexual stimulation. Just before orgasm a further increase was noted with peak values measured immediately after the orgasm began (pO(2) 28.6 +/- 3.1 mm Hg intravaginally and 47.3 +/- 4.1 labial). Labial pO(2) measurement decreased relatively rapidly soon after orgasm. The time to return to basal vaginal values after orgasm varied from 20 to 30 minutes. CONCLUSIONS: Previously, changes in female sexual arousal responses have been difficult to evaluate and quantify clinically. We developed a simple noninvasive reproducible technique to measure vaginal and minor labial blood flow. Age based and cycle dependent normograms now can be produced for vaginal and labial blood flow using this method. PMID- 11257667 TI - Urological assistance during therapeutic pubectomy. AB - PURPOSE: A technique is described to optimize exposure during female pubectomy to minimize injury to associated urological structures. MATERIALS AND METHODS: Three females with diagnosed osteomyelitis pubis underwent pubectomy. Before resection of the bone, the patients underwent a formal combined transvaginal and retropubic dissection. This dissection allowed complete freeing of the urethra and bladder from areas of orthopedic resection and optimized surgical exposure. RESULTS: All operations were completed successfully with no incidence of intraoperative urological structure injury and no postoperative pelvic instability. None of the patients required intraoperative or postoperative blood transfusions. Pelvic pain resolved in all 3 patients. CONCLUSIONS: Using a combined transvaginal and retropubic technique, the urologist may assist the orthopedic surgeon at the time of pubectomy. This technique potentially minimizes the incidence of vesical and urethral injury. PMID- 11257668 TI - Test-retest variation of pressure flow parameters in men with bladder outlet obstruction. AB - PURPOSE: We assess short-term (5 to 10 minutes) and long-term (24 weeks) test retest changes of repeated pressure flow examinations. MATERIALS AND METHODS: The pressure flow charts of 84 patients with benign prostatic enlargement and bladder outlet obstruction who had received either androgen suppressive therapy or placebo were reviewed retrospectively. Pressure flow examinations were performed at baseline, and at weeks 24 and 48. Each pressure flow session included 3 sequential voids. RESULTS: Median detrusor opening pressure, maximum detrusor pressure, detrusor pressure at maximum flow rate and minimum voiding pressure decreased statistically significantly from void 1 to 2, ranging from 9.5% to 15.8%. From void 2 to 3 during the same pressure flow session there was a further reduction in obstruction parameters. Median Abrams/Griffiths number was 10.7% lower at void 2 compared to void 1 (p <0.0001) and the urethral resistance algorithm was 3.2% lower (p <0.0001). Long-term test-retest changes from baseline to week 24 and from week 24 to week 48 for the pressure flow parameters studied were negligible. CONCLUSIONS: Changes in pressure flow parameters at short-term test-retesting are considerable and probably of clinical significance. The standard pressure flow nomograms, which are based on single void pressure flow studies, might need modification when applied to repeat void studies. PMID- 11257669 TI - Percutaneous afferent neuromodulation for the refractory overactive bladder: results of a multicenter study. AB - PURPOSE: More than 20 million Americans have an overactive bladder, the predominant symptoms being frequency, urgency, urge incontinence and pelvic pain. While the etiology is not completely understood, most investigators believe the causes to be many and the pelvic floor to be intimately related. Whatever the etiology, traditional therapies, including dietary manipulation, bladder drill, medications and physical therapy, are often poorly tolerated and/or ineffective. We report a prospective, multicenter clinical trial that was undertaken to determine the safety and efficacy of percutaneous peripheral afferent nerve stimulation for treatment of refractive overactive bladder and/or pelvic floor dysfunction. MATERIALS AND METHODS: A total of 53 patients with overactive bladders, in whom all traditional therapy failed, were enrolled in 1 of 5 sites within the United States. Patients received weekly percutaneous electrical stimulations via a 34 gauge needle placed near the tibial nerve 3 finger breadths above the ankle. Urodynamic studies, detailed voiding diaries, quality of life surveys, and incontinence impact questionnaires were completed before, during and after the study. RESULTS: Of the patients with a mean age of 57.4 years 89% (47 of 53) completed the 12-week study. A total of 71% of patients were classified as treatment successes by the investigators and were started on long-term treatment. On average patients noticed a 25% reduction in mean daytime and 21% reduction in mean nighttime voiding frequencies (p <0.05). Urge incontinence was reduced by an average of 35% (p <0.05). Statistically significant improvements were noted in selective pain and quality of life indexes. No significant adverse events related to treatment were noted in any patients. CONCLUSIONS: Percutaneous peripheral afferent nerve stimulation offers a safe, minimally invasive and effective treatment for managing refractive overactive bladder and/or pelvic floor dysfunction. PMID- 11257670 TI - Malignant composite pheochromocytoma of the adrenal gland in a patient with von Recklinghausen's disease. PMID- 11257671 TI - Spontaneous subcapsular renal hematoma secondary to anticoagulant therapy. PMID- 11257672 TI - Multimodal therapy for stage IV adult Wilms tumor. PMID- 11257673 TI - Management of diaphragmatic injury during laparoscopic nephrectomy. PMID- 11257674 TI - Partial nephrectomy for renal cell carcinoma in an allograft kidney 15 years after transplantation. PMID- 11257675 TI - Renal cell carcinoma recurrence in the renal fossa after nephrectomy. PMID- 11257676 TI - von Hippel-Lindau disease: renal tumors less than 3 cm. can metastasize. PMID- 11257677 TI - Allogeneic peripheral blood stem cell transplantation for metastatic renal cell carcinoma. PMID- 11257678 TI - Renal autotransplantation for complete ureteral avulsion following lumbar disk surgery. PMID- 11257679 TI - Ureterouterine and vesicoureterovaginal fistulas as a complication of cesarean section. PMID- 11257680 TI - Successful conservative management of perforated ileal neobladder. PMID- 11257681 TI - Missed female urethral injury complicated by myonecrosis of the thigh. PMID- 11257682 TI - Myositis and myonecrosis of the thigh: an unusual complication of a testicular thigh pouch. PMID- 11257683 TI - Intracavernosal injection of sildenafil citrate: misapplication of the drug. PMID- 11257684 TI - Testicular infarction associated with protein S deficiency. PMID- 11257685 TI - Transitional cell carcinoma in a prostatic remnant 10 years after radical cystectomy. PMID- 11257686 TI - Re: Renal cell carcinoma: prognostic significance of incidentally detected tumors. PMID- 11257687 TI - Re: Penile fracture in Kermanshah, Iran: report of 172 cases. PMID- 11257688 TI - Re: Simultaneous irradiation for prostate cancer: intermediate results with modern techniques. PMID- 11257690 TI - Retroperitoneal laparoscopic access in children using a direct vision technique. AB - PURPOSE: Retroperitoneal procedures were initiated in 1992 by balloon dissection of the retroperitoneum. More recently a new type of retroperitoneal access has been obtained by directly entering the retroperitoneum using the Visiport visual trocar. We present our initial experience with direct visual access to the retroperitoneum in the pediatric population. MATERIALS AND METHODS: A total of 31 children underwent retroperitoneal laparoscopy, including renal biopsy in 22, varicocelectomy in 5, renal cyst ablation in 3 and pyelolithotomy for a staghorn stone in 1. Patients were placed in the full flank position. A maximum of 3 ports was used and the initial trocar was placed under direct vision. The laparoscope was then used to dissect bluntly a working space in the retroperitoneum. RESULTS: All procedures were successful. Blood loss was minimal. Operative time was 4 hours for pyelolithotomy and less than 1 for the other procedures. Mean hospital stay was 1.5 days and all patients returned to normal activity at a mean of 6 days. Two minor complications developed. The peritoneum was inadvertently entered in 1 case, in which no further treatment was necessary and convalescence was uneventful and short. In another case severe arrhythmia developed, resulting in an aborted procedure. CONCLUSIONS: This technique is simple, safe and does not require extensive laparoscopic experience. PMID- 11257691 TI - Intravesical Jackson-Pratt drain for urinary diversion after augmentation cystoplasty. AB - PURPOSE: Augmentation cystoplasty has become a primary form of bladder management in children with a noncompliant bladder. Excellent urinary drainage is required for anastomotic healing and the removal of mucous buildup. Suprapubic drainage traditionally involves a Malecot catheter, although poor irrigation and dislodgment of this type of catheter are well-known complications. We report the placement of an intravesical Jackson-Pratt drain for urinary diversion in augmented bladders. MATERIALS AND METHODS: We reviewed our use of an intravesical Jackson-Pratt drain for urinary diversion between 1995 and 1999 in 17 patients. Postoperative catheter drainage and irrigation characteristics were assessed as well as catheter related complications. RESULTS: Average patient age was 13 years (range 3 to 27). The majority of patients underwent ileal (11) or sigmoid (4) cystoplasty and 1 each underwent composite and ureteral cystoplasty. Drains remained in place an average of 27 days (range 6 to 57). All patients had excellent drainage during the postoperative period. Irrigation was subjectively easier than with a Malecot catheter. Average cost of a latex-free Malecot catheter was 2.7-fold that of a Jackson-Pratt drain. No catheters became nonfunctional before removal, although 1 was inadvertently pulled during patient transfer. CONCLUSIONS: A Jackson-Pratt drain provides excellent urinary drainage in patients undergoing augmentation cystoplasty. Multiple openings along the tube seem to improve irrigation in contrast to the single opening in a Malecot catheter, which often aspirates a region of the augmented bladder. The ready availability, decreased cost, ease of irrigation, increased pliability with decreased chance of dislodgment and lack of latex make an intravesical Jackson Pratt drain a superior choice for augmented neurogenic bladder. PMID- 11257689 TI - Outcome of patients with prenatally detected duplex system ureterocele; natural history of those managed expectantly. AB - PURPOSE: We assessed the outcome of patients treated for prenatally detected duplex system ureterocele with particular reference to those treated expectantly. MATERIALS AND METHODS: We reviewed the records of 52 consecutive patients treated between 1984 and 1999 with a median followup of 8 years (range 1 to 16.2). RESULTS: Of the 38 patients who underwent surgical treatment 13 subsequently required unplanned secondary procedures. A total of 14 cases satisfying currently defined criteria, including less than 10% upper renal pole function, an unobstructed lower pole (absent nonrefluxing hydroureteronephrosis), lower pole vesicoureteral reflux not exceeding grade III and unobstructed bladder outflow, were managed expectantly with a median followup of 8 years (range 1.6 to 12.8). In this group of patients prophylactic antibiotics were routinely prescribed until the completion of toilet training or age 5 years in those with persistent reflux on repeat cystography. None has required surgery or had symptoms or urinary infection. In 6 cases followup ultrasonography showed substantial resolution of upper pole hydronephrosis with a collapsed ureterocele. Furthermore, 7 of the 38 patients who underwent surgical treatment early in our series would have been treated expectantly had the current criteria been applied. CONCLUSIONS: In 14 of the 52 patients (approximately 27%) with prenatally detected duplex system ureterocele the natural history of the complaint is essentially benign within the currently available followup. PMID- 11257692 TI - The Giessen-Mainz-Frankfurt procedure: a new method for complex pelvic reconstruction for bladder exstrophy. AB - PURPOSE: In bladder exstrophy primary reconstruction remains the gold standard worldwide. Despite various types of osteotomies the permanent correction of pubic diastasis remains a challenge. In maxillofacial surgery callus distraction is a routine treatment for hypoplastic mandibles. Originally described by Ilizarov, this method provides stable and true bone lengthening after gradual distraction of an osteotomy site as long as the periosteum remains intact. In cooperation with the departments of maxillofacial surgery and orthopedics we used this technique to correct pubic diastasis and facilitate phallic reconstruction in a 4 1/2-year-old boy with bladder exstrophy who had previously undergone continent diversion. MATERIALS AND METHODS: Three-dimensional computerized tomography was used to create a stereolithography model and mock surgery was performed. Based on this model bilateral osteotomies of the superior and inferior segments of the pubic bones were done, preserving the periosteum. Pins were inserted and a multidirectional external fixation device was mounted. Distraction was started on day 5 postoperatively. The distraction rate was 1 mm. daily and immobilization time was 28 days. The distraction progress was monitored by sonography. The device was removed 6 weeks postoperatively. RESULTS: Radiography of the pelvis 2 years postoperatively revealed that the distance between the pubic bones had decreased from 6 to 3 cm. (50%). Simultaneously the slanting angle normalized from 24 to 35 degrees due to upward rotation of the inferior pubic rami. Mineralization in the newly formed bones was excellent. Visible penile length had increased significantly. CONCLUSIONS: To our knowledge we describe the first use of the basic Ilizarov principle of callus distraction for permanent complex pelvic reconstruction for bladder exstrophy in a 4 1/2-year-old boy. After subperiostial osteotomies approximation of the symphysis and rotation of the inferior pubic rami were achieved with a device commonly used in maxillofacial surgery. Approximation of 1 mm. daily for 28 days resulted in significant penile lengthening. At a followup of 2 years there were stable pelvic ring reconstruction and normal mineralization of the newly formed bones. In contrast to the standard techniques of osteotomies for correcting pubic diastasis, the Giessen-Mainz-Frankfurt procedure provides true bone growth with a stable decrease in diastasis. Successful penile reconstruction was facilitated 1 year postoperatively. This method may also be useful in primary and secondary bladder reconstruction. PMID- 11257693 TI - Congenital posterior urethral membrane: variable morphological expression. AB - PURPOSE: We assessed the variable morphological expression of posterior urethral membrane by reviewing video recorded cystoscopy. MATERIALS AND METHODS: Between December 1990 and July 2000, 86 males newborn to 15 years old undergoing cystoscopy for urethral anomalies were identified with a posterior urethral membrane. Recorded cystoscopy was reviewed and membrane degree was graded as minimal, moderate or severe. RESULTS: Of the 86 boys with a membranous lesion in the posterior urethra the condition was considered severe in 40, moderate in 21 and minimal in 21. Four patients on whom data were too inadequately recorded to be properly classified were excluded from study. CONCLUSIONS: This study demonstrates that congenital posterior urethral membrane represents a spectrum of lesions and may vary in the degree of obstruction. PMID- 11257694 TI - Molecular analysis of hypospadias in a boy with dicentric Y chromosome. PMID- 11257696 TI - Results of preputial reconstruction in 77 boys with distal hypospadias. AB - PURPOSE: There is growing interest in preputial reconstruction combined with hypospadias repair. We retrospectively analyzed its results for future developments and patient information. MATERIALS AND METHODS: We evaluated 77 boys who underwent distal hypospadias repair combined with preputial reconstruction to determine complications, risks and failures. RESULTS: At a mean followup of 2.5 years 52 patients had an anatomically normal penis with a normal retractable foreskin, while 25 (33%) presented with a complication. The most common complications were partial dehiscence, and fistula of the prepuce and urethra. There was a complication of the reconstructed foreskin only in 16 cases (21%), a combined problem with the reconstructed foreskin and reconstructed urethra in 7 (9%), and a problem with the reconstructed urethra in 2 (3%). Of the 25 patients with complications 19 underwent reoperation with closure of the fistula or dehiscence and 5 were circumcised, while in 1 the parents accepted the minor cosmetic problem and refused reoperation. CONCLUSIONS: Preputial repair combined with hypospadias repair may lead to anatomically correct reconstruction of the penis at the cost of a 33% complication rate. Parents are informed about this risk and to date in 15% of all boys with distal hypospadias the parents have elected preputial reconstruction. PMID- 11257695 TI - Etiological studies of severe or familial hypospadias. AB - PURPOSE: Hypospadias is a congenital anomaly occurring in 1250 to 1830 live male births, of which 20% involve a severe type. The recurrence risk in families is high. In the majority of cases the underlying etiology remains unknown, which hampers further management based on the specific requirements associated with a specific etiology. MATERIALS AND METHODS: In a single center study 63 unselected cases of severe hypospadias were studied for all presently known causes of hypospadias using clinical as well as molecular biological techniques. Also, 16 families with hypospadias were analyzed for possible androgen receptor gene mutations. RESULTS: In 31% of cases of severe hypospadias the underlying etiology was identified. Of these 31% of cases 17% were due to complex genetic syndromes, 9.5% were due to chromosomal anomalies, and 1 involved the vanishing testes syndrome, the androgen insensitivity syndrome and 5alpha-reductase type 2 deficiency, respectively. Based on hormone stimulation tests Leydig cell hypoplasia and disorders of testosterone biosynthesis were suspected in some patients but not confirmed by mutation analysis of the respective genes. Familial hypospadias was due to androgen insensitivity in only 1 family but no other etiologies were identified in this group. CONCLUSIONS: Using patient history, physical examination, karyotyping, hormonal evaluation, including human chorionic gonadotropin testing in prepubertal cases and additional biochemical and molecular genetic evaluation, an etiological diagnosis was made in 31% of cases of severe hypospadias. This diagnosis has implications for further patient treatment. In addition, familial hypospadias is rarely due to the androgen insensitivity syndrome. PMID- 11257697 TI - The histopathology of iatrogenic cryptorchid testis: an insight into etiology. AB - PURPOSE: Iatrogenic undescended testis may develop after inguinal hernia repair, presumably as a result of mechanical tethering of the testis or cord in scar tissue. Because some true cryptorchid testes appear to be completely descended at birth and later ascend during childhood, some iatrogenic undescended testes may be low lying undescended testes. To determine whether iatrogenic undescended testes may be unrecognized cryptorchid testes at herniorrhaphy we examined biopsies of iatrogenic undescended testes and the corresponding contralateral descended testis. MATERIALS AND METHODS: Between 1985 and 1999 bilateral testis biopsies were obtained at orchiopexy in 37 boys 1.5 to 11.8 years old who previously underwent inguinal hernia correction. Histomorphometric analysis of germ cell counts was performed on the undescended and contralateral descended testes, and compared to the count in bilateral biopsies of 37 age and position matched patients with true unilateral cryptorchidism. RESULTS: There were no significant differences in volume or total and differential germ cell counts in the undescended and contralateral descended testes in the study groups and age matched controls with primary unilateral cryptorchidism. The mean number of germ cells per tubule in the undescended testis in patients with a greater than 5-year interval from herniorrhaphy to orchiopexy was significantly decreased compared to those with an operative interval of less than 5 years (0.27 +/- 0.33 versus 0.93 +/- 1.4, p = 0.026). CONCLUSIONS: Some patients with iatrogenic undescended testis may have an unrecognized low cryptorchid testis. Careful physical examination before and after inguinal surgery is recommended. The early repair of iatrogenic undescended testis is warranted to prevent further damage. PMID- 11257698 TI - Familial Mediterranean fever as an unusual cause of acute scrotum. PMID- 11257699 TI - Early stage of urolithiasis formation in experimental hyperparathyroidism. AB - PURPOSE: We have previously noted marked acceleration in the proliferative activity of parathyroid cells in rats with spontaneous hypercholesterolemia and secondary hyperparathyroidism. Using this proliferative potential we investigated whether transplantation of these enlarged parathyroids into normal rats would induce hyperparathyroidism and renal stones. MATERIALS AND METHODS: We used 26 week-old male rats with spontaneous hypercholesterolemia as donors, and 5-week old normal male Sprague-Dawley rats and rats with spontaneous hypercholesterolemia as recipients. Enlarged parathyroid glands were transplanted into group 1--Sprague-Dawley rats with no treatment, group 2--Sprague-Dawley rats that received FK-506 as an immuno-suppressor, group 3--rats with spontaneous hypercholesterolemia rats that underwent parathyroidectomy plus FK-506 administration and group 4--Sprague-Dawley rats that underwent parathyroidectomy plus FK-506 administration. Parathyroidectomy was performed in recipients before transplantation to ensure a low calcium condition. RESULTS: Grafts were rejected within 11 and 15 weeks in groups 1 and 2, respectively. In group 3, 78% of the grafts were successful even after 19 weeks. In group 4 graft survival was 30% at 15 weeks with complete rejection at 19 weeks. In group 3 gradually elevated serum parathyroid hormone was observed as well as stone plaques containing calcium oxalate and calcium phosphate in renal tubules located mainly in the corticomedullary junction. An increased number of plaques was associated with higher parathyroid hormone. CONCLUSIONS: Our study shows that transplanted parathyroid glands function with an immunosuppressive agent and the maintenance of hypocalcemic conditions, and they secrete sufficient parathyroid hormone to demonstrate hyperparathyroidism. Plaque in these kidneys indicates an early stage of urolithiasis caused by hyperparathyroidism. PMID- 11257700 TI - Treatment of human renal cell carcinoma by a conditionally replicating herpes vector G207. AB - PURPOSE: Surgical removal remains the only potentially curative therapy for renal cell carcinoma. In this study we evaluated the inhibitory effect of the replication competent engineered herpes simplex virus type 1, G207, for renal cell carcinoma in vitro and in vivo. MATERIALS AND METHODS: The nature of G207 enables it to replicate within cancer cells, thus, causing cytolysis, but replication is restricted within normal cells. The susceptibility of the human renal cancer cell lines ACHN and A498 to G207 at a multiplicity of infection of 0.1 was examined. In addition, the growth characteristics of G207 was assessed. In vivo athymic mice bearing subcutaneous tumors were inoculated with 1 x 10(7) plaque forming units of G207 intra-neoplastically. For pathological analysis subcutaneous tumors were stained with X-gal. RESULTS: Two cell lines were efficiently destroyed by G207 within 1 week. The viral yields of G207 increased in a time dependent manner. In vivo the intra-neoplastic inoculation of G207 caused significantly decreased tumor growth in athymic mice harboring subcutaneous human renal cancer cells. On day 14 the mean growth ratio of ACHN and A498 lesions was significantly inhibited in G207 treated compared to control tumors (p <0.005 and <0.0001, respectively). CONCLUSIONS: These results suggest that G207 should be considered another potential therapeutic agent for renal cell carcinoma. PMID- 11257701 TI - Intravital microscopic assessment of pressure induced microcirculatory changes after enterocystoplasty in rats. AB - PURPOSE: Chronic over distention may lead to enterocystoplasty rupture. It is hypothesized that pressure induced microvascular derangement and subsequent ischemia of the enterocystoplasty intestinal patch may have a role in this process. We describe distention induced microcirculatory alterations in a chronic rat model of enterocystoplasty using intravital video microscopy. MATERIALS AND METHODS: Microcirculation in the muscle layer of the intact bladder and intestine, and in the enterocystoplasty 90 days after surgery were examined at greater and less than urethral sphincter closure pressure. Microcirculatory changes were recorded during stepwise increments of intraluminal pressure up to 80 mm. Hg or when 20 mm. Hg was continuously maintained for 60 minutes. RESULTS: The enterocystoplasty components of intestine and bladder displayed baseline microcirculatory characteristics similar to those observed in the intact organs. As evidenced by microcirculatory flow and functional capillary density measurements in the intact intestine and the ileal portion of enterocystoplasty, intraluminal pressure elevation to greater than 25 or 30 mm. Hg significantly compromised capillary perfusion by approximately 50% and 75%, respectively. Lower intraluminal pressure did not cause microcirculatory disturbance even when maintained for a longer period. In the intact bladder and bladder portion of enterocystoplasty only pressure increases to greater than 80 mm. Hg affected tissue perfusion. CONCLUSIONS: Intravital microscopy in the augmented rat bladder is a sensitive and suitable means of assessing clinically relevant microcirculatory changes. These experiments demonstrate the significance of distention induced microcirculatory impairment in the intact bowel and the intestinal site of enterocystoplasty even at less than urethral closure pressure. PMID- 11257702 TI - Mesenchymal-epithelial interactions in bladder smooth muscle development: effects of the local tissue environment. AB - PURPOSE: We have previously shown that mesenchymal-epithelial interactions are necessary for the development of bladder smooth muscle. Specifically without fetal or adult urothelium embryonic rat bladder mesenchyma does not differentiate into smooth muscle. The mechanism responsible for this interaction is not known, although it is postulated that diffusable growth factors have a role. Our hypothesis is that diffusable factors within adult rat bladders influence smooth muscle differentiation. MATERIALS AND METHODS: Chimeric bladders were created by surgically implanting 14-day embryonic rat bladder mesenchyma before smooth muscle differentiation into the detrusor space of adult syngeneic hosts to test whether the host urothelium would induce smooth muscle differentiation without being in direct contact with fetal bladder mesenchymal tissue. Sub-detrusor pockets were created between the serosa and smooth muscle layer, between the smooth muscle layer and lamina propria, and between the lamina propria and urothelium in direct contact with urothelium. Controls consisted of intact 14-day embryonic rat bladders with the urothelium not removed, and 14-day embryonic bladder mesenchyma recombined with urothelium (direct contact) placed within the sub-detrusor space of the bladder and under the renal capsule. RESULTS: Immunohistochemical staining with antibodies directed against smooth muscle alpha actin and urothelium (cytokeratin 7) revealed smooth muscle differentiation in intact embryonic bladders and bladder mesenchyma plus urothelium recombinants in contrast to bladder mesenchyma alone, which had no alpha-actin staining (morphometric smooth muscle analysis p = 0). There was no alpha-actin staining in chimeric bladders even when bladder mesenchymal grafts were placed directly in contact with host urothelium. In addition, bladder mesenchyma plus urothelial recombinants within the host bladder had less alpha-actin staining than their counterparts placed under the renal capsule (p = 0.001). CONCLUSIONS: A diffusable factor most likely exists within adult rat bladders that inhibits smooth muscle differentiation. PMID- 11257703 TI - Up-regulation of a gene homologous to the human tumor necrosis factor receptor associated factor 6 gene in the obstructed rabbit bladder determined by differential display polymerase chain reaction. AB - PURPOSE: We identified differentially expressed genes in the rabbit bladder after partial outlet obstruction. MATERIALS AND METHODS: Differential display polymerase chain reaction (PCR) was performed on smooth muscle tissue from normal, 2 and 6-week obstructed rabbit bladders. Semiquantitative reverse transcriptase PCR, Western and RNA blot analysis were done to confirm messenger RNA and protein up-regulation. RESULTS: A signal transducing protein human tumor necrosis factor receptor associated factor 6 (TRAF6)-like protein was identified on differential display PCR. TRAF6-like protein was up-regulated in rabbit bladders after 2 weeks of partial outlet obstruction. Reverse transcriptase PCR demonstrated TRAF6-like protein in bladder muscle tissue and semiquantitative analysis confirmed up-regulation in 2-week obstructed tissue. These findings were confirmed by RNA and Western blot analysis. CONCLUSIONS: TRAF6-like protein is up regulated during the early phase of bladder outlet obstruction in rabbits. To our knowledge involvement of this gene in bladder outlet obstruction has not been described previously. TRAF6 may have a role in the regulation of molecular changes during the early bladder response to outlet obstruction, such as the up regulation of growth factors and proto-oncogenes. Further understanding of this signaling pathway and its role in bladder outlet obstruction may open new avenues for treating detrusor dysfunction. PMID- 11257704 TI - Morphological aspects of the female pig bladder neck and urethra: quantitative analysis using computer assisted 3-dimensional reconstructions. AB - PURPOSE: To investigate whether the pig is a suitable model for studies of lower urinary tract function and dysfunction, we sought to determine the morphology of the female pig bladder neck and urethra. Computer assisted 3-dimensional (D) reconstructions from step serial histological sections were used for visualization of the spatial relationships between neighboring urethral wall components, and the quantification of these components in the bladder neck and along the urethra. MATERIALS AND METHODS: Step serial histological paraffin sections from the bladder neck and urethra of 6 female pigs, stained with Masson's trichrome, were used to generate computer assisted 3-D reconstructions using MacStereology (Ranfurly MicroSystems Ltd., Airdrie, United Kingdom) as the 3-D software package. RESULTS: The bladder neck and urethral anatomy revealed well defined smooth and striated muscle layers that varied in location, regional distribution and orientation. Circular smooth muscle was maximally developed in the mid urethra, at which point maximal urethral pressure was observed. The longitudinal smooth muscle layer appeared continuous with the detrusor, implicating a possible role in urethral shortening at the onset of voiding. A small circular and longitudinal striated muscle component was present in the distal urethra. CONCLUSIONS: Anatomical differences exist between the female pig and human bladder neck and urethra, which were successfully highlighted using computer assisted 3-D reconstructions from step serial histological paraffin sections. PMID- 11257705 TI - The use of hydrogen peroxide to enhance the efficacy of doxorubicin hydrochloride in a murine bladder tumor cell line. AB - PURPOSE: We determined whether the cytotoxicity of doxorubicin hydrochloride would be enhanced by adding hydrogen peroxide as a source of oxygen free radicals. MATERIALS AND METHODS: Mouse bladder tumor cells (MBT-2) were grown in RPMI 1640 medium and treated with various concentrations of doxorubicin hydrochloride for 2 hours. Protein content was assayed as a measure of cell growth. A similar set of experiments was done with cells exposed to hydrogen peroxide only and combined doxorubicin and hydrogen peroxide. Protein content was again assayed as a measure of cell growth. Cells were also assayed for glutathione peroxidase and malonyl dialdehyde, a product of lipid peroxidation, to determine the mechanism of cell damage. Furthermore, MBT-2 cells were incubated with 100 M. alpha-tocopherol, a free radical scavenger, before exposure to hydrogen peroxide to determine whether the effects of hydrogen peroxide could be reversed. RESULTS: We observed a dose dependent inhibition of MBT-2 cell growth after exposure to doxorubicin hydrochloride. Exposure to doxorubicin and hydrogen peroxide resulted in greater cell growth inhibition than exposure to either agent alone. The effects of hydrogen peroxide on cell proliferation were reversed by pre-incubation with alpha-tocopherol. CONCLUSIONS: As a source of oxygen free radicals, hydrogen peroxide enhances the antiproliferative effect of doxorubicin hydrochloride on a mouse bladder tumor cell line. Thus, hydrogen peroxide may be a relatively inexpensive, nontoxic method of augmenting the cytotoxicity of doxorubicin hydrochloride. Further studies are warranted to determine whether these observations may have clinical application. PMID- 11257706 TI - Nontraumatic urethral dyssynergia in neonatally estrogenized male rats. AB - PURPOSE: Bladder outlet obstruction develops in estrogen treated males. Because of the lack of electromyography recordings, earlier studies have not clarified the failure mechanisms of voiding. We simultaneously recorded electromyography activity of the proximal rhabdosphincter in neonatally estrogenized rats with transvesical cystometry and urethral flow, followed by morphometric analysis of the urethral structure. MATERIALS AND METHODS: Rats treated neonatally with 10 microg. diethylstilbestrol daily on days 1 to 5 after birth were used in urodynamics and morphological studies at ages 5 to 6.5 months. Using anesthesia the bladder, anterior surface of the proximal rhabdosphincter and distal urethra were exposed to record simultaneously the high frequency oscillations of intraluminal bladder pressure, and the rates of intermittent flow from the distal urethra and electromyography activity of the proximal rhabdosphincter with a suction electrode. RESULTS: Neonatally estrogenized rats had higher mean maximal bladder pressure plus or minus standard deviation (42.1 +/- 6.4 versus 37.7 +/- 4.9 mm. Hg, p = 0.01), decreased mean flow (2.3 +/- 0.1 versus 4.1 +/- 1.6 ml. per minute, p < 0.0001) and mean increment of proximal rhabdosphincter electromyography depolarization amplitude (3.0 +/- 0.78 versus 2.6 +/- 0.87 mV., p = 0.02) compared with controls, while mean transient repolarization was absent or highly decreased (-0.3 +/- 0.61 versus 0.3 +/- 0.9 mV., p = 0.04). Morphologically the proximal rhabdosphincter was atrophied with increased connective tissue. CONCLUSIONS: Alterations in the structure and electromyography activity of the urethral musculature imply that neonatal exposure to diethylstilbestrol predisposes male rats to urethral atrophy and dyssynergia, evident as altered electromyography activity of the proximal rhabdosphincter. PMID- 11257707 TI - Vascular endothelial growth factor restores corporeal smooth muscle function in vitro. AB - PURPOSE: The therapeutic use of vasculogenic growth factors has been successfully demonstrated in models of organ ischemia. We determined whether vascular endothelial growth factor (VEGF) would reverse corporeal smooth muscle dysfunction in the hypercholesterolemic rabbit model of erectile dysfunction. MATERIALS AND METHODS: A total of 36 New Zealand White rabbits were fed a normal (12) or 1% cholesterol (24) diet and treated after 6 weeks with 0.9 mg. VEGF or vehicle. At 6 weeks 24 rabbits received a single intracavernous dose and 12 received a single intravenous bolus of either drug. Ten days after injection corporeal smooth muscle function was analyzed after relaxation to acetylcholine and sodium nitroprusside using isometric tension studies. Corporeal sections were assessed for smooth muscle content with f-actin staining and VEGF expression by immunohistochemical study and enzyme-linked immunosorbent assay. RESULTS: Endothelium dependent (acetylcholine) and nitric oxide mediated (sodium nitroprusside) smooth muscle relaxation were impaired in cholesterol fed animals (p = 0.021 and 0.003, respectively). Intracavernous VEGF treatment restored sodium nitroprusside mediated relaxation to normal (p = 0.015) and intravenous VEGF restored acetylcholine and sodium nitroprusside mediated relaxation (p = 0.014 and 0.018, respectively). Decreased smooth muscle content was noted in cholesterol fed animals versus normal diet controls (p = 0.008), which was not affected by VEGF treatment (p = 0.450). Corporeal endothelial cell content was increased after intracavernous but not intravenous VEGF treatment (p = 0.001 and 0.385, respectively). VEGF expression was augmented after treatment with recombinant VEGF (p <0.001). CONCLUSIONS: VEGF administration variably mitigated the impairment of corporeal smooth muscle relaxation in the hypercholesterolemic rabbit model of erectile dysfunction. PMID- 11257708 TI - The oral efficacy of vardenafil hydrochloride for inducing penile erection in a conscious rabbit model. AB - PURPOSE: Inhibiting cyclic guanosine monophosphate metabolism may induce penile erection during concomitant nitric oxide production. Vardenafil hydrochloride is a new, highly selective, potent cyclic guanosine monophosphate phosphodiesterase 5 inhibitor. We determined the oral effectiveness of vardenafil in a simple and quantitative conscious rabbit model. MATERIALS AND METHODS: Vardenafil was given orally to conscious rabbits. Erection was assessed in a time dependent manner by measuring the length of the uncovered penile mucosa. Erection was evaluated in the absence and presence of intravenous sodium nitroprusside, a nitric oxide donor. RESULTS: Vardenafil induced dose dependent penile erection in conscious rabbits after the oral administration of 1 to 30 mg./kg. The efficacy of vardenafil was potentiated and effective doses were significantly reduced by the simultaneous administration of sodium nitroprusside. CONCLUSIONS: The effect of vardenafil on penile erection after oral administration was clearly demonstrated in the conscious rabbit model. The time course and early onset of activity indicate that it may be useful for treating erectile dysfunction. Potentiation of the effect by the nitric oxide donor sodium nitroprusside implies that it would have enhanced activity during sexual arousal, when nitric oxide is produced endogenously. The clinical development of this product for erectile dysfunction is proceeding. PMID- 11257709 TI - Insulin-like growth factor binding protein-3 mediates 1 alpha,25-dihydroxyvitamin d(3) growth inhibition in the LNCaP prostate cancer cell line through p21/WAF1. AB - PURPOSE: We determined that insulin-like growth factor binding protein 3 (IGFBP 3) induction by 1,25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)) is a necessary component of 1,25-(OH)(2)D(3) mediated growth inhibition of the LNCaP human prostate cancer cell line. In addition, induction of the cyclin dependent kinase inhibitory protein p21/WAF/CIP1 by 1,25-(OH)(2)D(3) is mediated by IGFBP-3. MATERIALS AND METHODS: Induction of IGFBP-3 by 1,25-(OH)(2)D(3) was determined by enzyme-linked immunosorbent assay for IGFBP-3 protein and by Northern blot analysis for IGFBP-3 messenger (m) RNA. Growth assays for LNCaP cells were determined by measuring DNA content. The contribution of IGFBP-3 toward 1,25 (OH)(2)D(3) mediated growth inhibition was determined by adding either antisense oligonucleotides or immuno-neutralizing antibodies to culture media of growth assays. Regulation of p21/WAF/CIP1 was determined by Western blot analysis. RESULTS: Adding 1,25-(OH)(2)D(3) to LNCaP prostate cancer cells demonstrated that 1,25-(OH)(2)D(3) significantly up-regulated IGFBP-3 at the mRNA and protein levels in these cells approximately 3-fold over control levels. Also, adding IGFBP-3 protein to LNCaP cell growth medium inhibited LNCaP cell growth. Interestingly adding IGFBP-3 antisense oligonucleotides or antibodies directed toward IGFBP-3 abolished the growth inhibitory actions of 1,25-(OH)(2)D(3), indicating that this effect is IGFBP-3 dependent. Furthermore, to connect the mechanisms of IGFBP-3 and 1,25-(OH)(2)D(3) mediated growth inhibition we demonstrated that IGFBP-3 up-regulates the expression of p21/WAF1 protein to approximately 2-fold over the control level. Adding an IGFBP-3 immuno neutralizing antibody completely prevented the 1,25-(OH)(2)D(3) induced up regulation of p21/WAF1. CONCLUSIONS: 1,25-(OH)(2)D(3) up-regulates IGFBP-3 in the LNCaP cell line at the mRNA and protein levels. The growth inhibitory action of 1,25-(OH)(2)D(3) on LNCaP cells depends on active IGFBP-3, as evidenced by the loss of growth inhibition induced by IGFBP-3 antisense oligonucleotide and immuno neutralization experiments. A possible connection between IGFBP-3 and 1,25 (OH)(2)D(3) lies in the cyclin dependent kinase inhibitory protein p21/WAF1 since IGFBP-3 and 1,25-(OH)(2)D(3) each up-regulate this protein and both inhibit LNCaP cell growth. Therefore, we hypothesize that the mechanism of action by which IGFBP-3 and 1,25-(OH)(2)D(3) induce growth inhibition is the induction of p21/WAF1 because IGFPB-3 immuno-neutralizing antibodies completely abrogate the 1,25-(OH)(2)D(3) mediated up-regulation of p21/WAF1 and growth inhibition. PMID- 11257710 TI - Scatter factor-hepatocyte growth factor elevation in the serum of patients with prostate cancer. AB - PURPOSE: Scatter factor (SF), also known as hepatocyte growth factor (HGF), has been shown to induce proliferation, scattering and invasiveness in human prostate cancer cell lines. In this study we determined the serum level of SF-HGF in men with metastatic prostate cancer compared to those with localized prostate cancer and without prostate cancer. MATERIALS AND METHODS: Serum samples were obtained from men with biopsy proved adenocarcinoma of the prostate and radiographic evidence of metastatic disease, those with biopsy proved adenocarcinoma of the prostate and clinically localized disease, and those with negative sextant prostate biopsies. Serum SF-HGF was determined using a commercially available enzyme-linked immunosorbent assay kit. RESULTS: Of the 108 men enrolled in our study 52 had negative sextant biopsies, 36 had clinically localized cancer and 20 had metastatic disease. The serum level in men with metastatic disease was significantly elevated (mean 2,117 pg./ml., range 820 to 6,403) compared to that in men with localized cancer and without prostate cancer (mean 974 pg./ml., range 437 to 2,132 and 700, range 272 to 1,875, respectively, p = 9.5 x 10(-15)). Logistic regression analysis demonstrated that the association of ln (SF-HGF) with prostate cancer persisted after controlling for patient age and ln (prostate specific antigen) (p = 3.1 x 10(-4)). CONCLUSIONS: Serum SF-HGF is increased in men with metastatic prostate cancer. SF-HGF levels are associated with metastatic prostate cancer independent of the prostate specific antigen level and patient age. These data imply that SF-HGF may be an important serum marker for prostate cancer. PMID- 11257711 TI - Expression studies and mutational analysis of the androgen regulated homeobox gene NKX3.1 in benign and malignant prostate epithelium. AB - PURPOSE: The NKX-3.1gene is an androgen regulated prostate specific homeobox gene that is believed to have a vital role in normal prostate development. In mice the homologue NKx3.1 is exclusively expressed in prostate epithelium. In humans NKX3.1 expression is also restricted to the prostate but to our knowledge the cellular location has not been described. Furthermore, since NKX3.1 maps to chromosomal band 8p21, a region with high loss of heterozygosity in prostate cancer, the gene has been proposed to have tumor suppressor function. In this study we demonstrate that in human prostates NKX3.1 is expressed exclusively in secretory epithelial cells and the level of NKX3.1 expression remains invariant in normal tissue and in tissue showing various grades of prostate cancer. In the 19 cases examined the DNA sequences of the NKX3.1 gene were identical and no mutation was detected. MATERIALS AND METHODS: Frozen tissue from patients who underwent radical prostatectomy was used for this study. For in situ hybridization experiments a 377 bp fragment corresponding to a portion of the 3' untranslated region of the NKX3.1 gene was amplified by polymerase chain reaction and cloned into the pCRII plasmid vector Invitrogen. Antisense or sense [33P] uridine triphosphate labeled RNA probes were generated with SP6 or T7 RNA polymerase and hybridized to the tissue sections. Slides were exposed to photographic emulsion and visualized on autoradiography. Laser capture microdissection was performed to procure pure populations of malignant epithelium. DNA was isolated by digesting samples in proteinase K buffer. Polymerase chain reaction and direct sequencing was performed using standard protocols. RESULTS: In vitro hybridization showed that NKX3.1 expression was restricted to secretory epithelial cells within benign prostate glands. No expression was detected in stroma or infiltrating lymphocytes. NKX3.1 was expressed in all grades of malignant epithelium in all 25 cases examined. Direct sequencing of the coding region of NKX3.1 revealed the wild-type sequence in all 18 microdissected cancers analyzed. CONCLUSIONS: Based on our studies we propose that NKX3.1 gene expression is restricted to benign and malignant secretory epithelium within the prostate but NKX3.1 does not appear to be a classic tumor suppressor gene responsible for prostate cancer initiation. These findings are consistent with the role of NKX3.1 in the development of normal prostate epithelium and maintenance of normal secretory function. Thus, NKX3.1 may represent a useful molecular marker for benign and malignant prostate epithelium. PMID- 11257712 TI - Expression analysis of Y chromosome genes in human prostate cancer. AB - PURPOSE: We hypothesized that alterations in Y chromosome gene expression may be associated with prostate cancer. To test this hypothesis we analyzed the expression of 19 Y chromosome genes in benign and malignant prostate tissue. MATERIALS AND METHODS: To study the expression of Y chromosome genes RNA was extracted from prostate cancer and benign prostatic hyperplasia (BPH) tissue as well as from prostate cancer cell lines. RNA was reverse transcribed and polymerase chain reaction amplified using specific primers. These primers were designed for each gene sequence obtained from the gene data bank. We analyzed 19 Y chromosome genes using 6 cell lines, 7 BPH and 7 prostate cancer tissues. Normal testis RNA served as a positive control. RESULTS: Of the 19 genes analyzed in cell lines BPH-1 cells expressed the RPS4Y, USP9Y, TMSB4Y and DBY genes; DUPro expressed RPS4Y, USP9Y, TMSB4Y, DBY and UTY; DU145 expressed DAZ, RPS4Y, USP9Y, TMSB4Y, DBY, EIAFIY, PRKY and SMCY; LNCaP expressed TSPY, SRY, BPY1, PRY, DAZ, RBMIH, RPS4Y, DBY, EIAFIY, PRKY and SMCY; ND1 expressed DAZ, RPS4Y, USP9Y, TMSB4Y, DBY, EIAFIY, PRKY and SMCY; and PC3 expressed RPS4Y, USP9Y and DBY. BPH tissue expressed the SRY, PRY, DBY, PRKY, RPS4Y, TMSB4Y, USP9Y and ZFY genes. Prostate cancer tissue expressed the PRY, TSPY, USP9Y, UTY, DBY, SMCY, ZFY, EIAFIY, TMSB4Y and RPS4Y genes. CONCLUSIONS: The differential expression of Y chromosome genes in prostate cancer, BPH tissue and prostate cancer cell lines indicates that they may have a role in prostate cancer. PMID- 11257713 TI - Interaction of bladder glycoprotein GP51 with uropathogenic bacteria. AB - PURPOSE: A major component of bladder surface mucin is a glycoprotein GP51 (molecular weight 51 kD.). GP51, which has previously been isolated from rabbit mucosa, appears to function as part of the defense mechanism in an in vivo infection model. GP51 coats the epithelium and is secreted into the urine, as detected by immunohistochemical testing and enzyme-linked immunosorbent assay (ELISA). Increased urinary GP51 occurs during urinary tract infection. To elucidate the role of GP51 as a component of the primary defense mechanism we studied interactions with uropathogenic bacterial isolates and urine from symptomatic patients with urinary tract infection. MATERIALS AND METHODS: ELISA was performed to demonstrate the binding of GP51 and various uropathogens. Immunochemical studies were done using monoclonal antibodies to GP51 to determine the interaction of GP51 with certain uropathogenic isolates, including Escherichia coli, Enterobacter cloacae, Klebsiella pneumoniae, Proteus vulgaris, Pseudomonas aeruginosa, Serratia marcescens, Staphylococcus aureus, S. epidermidis and Streptococcus faecalis. Infected urinary sediments and uropathogenic bacterial cultures were examined by immunocytochemical testing to localize GP51. Antigen inhibition ELISA was done to quantitate urinary GP51 in the urine of 17 normal controls and 19 patients with urinary tract infection. RESULTS: ELISA revealed that GP51 binds to a wide spectrum of gram-positive and gram-negative uropathogens in semiquantitative fashion. Immunochemical methods confirmed that purified GP51 binds to bacteria, encapsulating and aggregating the bacteria. Clinical specimens showed GP51 localized to bacteria and uroepithelial cells. We observed a significant increase in urinary GP51 in urinary tract infection compared to uninfected urine (p = 0.0003). CONCLUSIONS: These studies suggest that GP51, a component of bladder mucin, may be a strategic factor in the primary defense mechanism of the bladder. PMID- 11257716 TI - Over-the-counter (OTC) cough remedies. PMID- 11257717 TI - Direct-to-consumer advertisements for Glucophage XR. PMID- 11257714 TI - The presence of the virulence island containing the usp gene in uropathogenic Escherichia coli is associated with urinary tract infection in an experimental mouse model. AB - PURPOSE: A putative virulence island commonly noted in the genome of uropathogenic Escherichia coli strains has recently been reported. We have observed that the island includes a gene consisting of a protein designated uropathogenic specific protein (usp) and 3 small open reading frames (orfU1-3). In our current study we assessed the importance of the genes located in the putative virulence island in the pathogenesis of urinary tract infection using a mouse pyelonephritis model. MATERIALS AND METHODS: A total of 427 E. coli strains isolated from the urine of 194, 76 and 107 subjects suffering from cystitis, pyelonephritis and prostatitis, respectively, and 50 isolates from the feces of healthy individuals were examined for genotypes and serotypes. In addition, several recombinant E. coli strains possessing usp and/or orfU1 to 3 were constructed for evaluating the significance of these genes using an experimental pyelonephritis mouse model. RESULTS: The usp was significantly more often associated with uropathogenic E. coli strains (79.4% from cystitis, 93.4% from pyelonephritis and 88.8% from prostatitis) than with fecal E. coli strains from healthy individuals (24%). Furthermore, usp was frequently associated with all common serotypes of uropathogenic E. coli (71.7% to 100%). In challenge experiments using the mouse urinary tract infection model the vector possessing usp significantly enhanced the infectibility of the E. coli host cell, whereas the 3 small proteins at the downstream of usp failed to show the effect. CONCLUSION: Our results indicate that usp may contribute to the causation of urinary tract infection and may be considered a major virulence determinant of uropathogenic E. coli. PMID- 11257718 TI - Once-a-week alendronate (Fosamax). PMID- 11257719 TI - Improving stroke patients' care: a patient held record is not enough. AB - BACKGROUND: Stroke patients' care in hospital tends to be poorly organised, with poor communication and a lack of information being frequent sources of complaint. The purpose of this study was to evaluate whether a patient-held record (PHR) would result in greater patient satisfaction and better care planning for stroke patients. METHODS: A time series control (6 months) - intervention (8 months) - control (6 months) was used among London teaching hospital general medical and geriatric medicine inpatient wards. All stroke patients admitted to the wards during the intervention phase received a PHR and were instructed in its use. Demographic, stroke severity, social factors and outcomes were collected from all stroke patients during all phases of the study. RESULTS: Of 252 stroke patients aged 46 to 98 years entered into the study, by six months after admission 118 (46.8%) had died. PHR and control group patients were well matched in terms of socio-demographic characteristics and pre-stroke ability. At six months after admission, 119 (97%) patients responded to the questionnaire. Just over half (56%, 13) of intervention group patients recalled receiving a PHR. Of those patients, 59% reported reading the PHR, 27% had lost their PHR, and two-thirds said they had difficulties encouraging staff to write in the PHR. Half felt that possession of the PHR was more trouble than it was worth. PHR group patients were more satisfied with the recovery they had made (79% vs. 59%, p=0.04), but felt less able to talk to staff about their problems (61% vs. 82%, p=0.02). PHR group patients reported receiving fewer explanations about their condition (18% vs. 33%, p=0.12) and treatment (26% vs. 45%, p=0.07), and were more afraid of asking doctors questions (21% vs. 4%, p=0.01) than controls. PHR group patients were no better prepared for hospital discharge than control group patients, and both groups were ill-informed about services and benefits that might have helped after discharge from hospital. CONCLUSIONS: Stroke patients received poor information and explanations regardless of whether they received a PHR. A PHR did not appear to improve patient satisfaction or discharge planning, and may have reduced opportunities for communication and explanation. PMID- 11257721 TI - Post-embedding double-gold labeling immunoelectron microscopic co-localization of neurotransmitters in the rat brain. AB - BACKGROUND: In this report, we describe a new quantitative electron microscopic protocol based on the use of double colloidal-gold post-embedding immunostaining procedure as markers to analyze the subcellular distribution of enkephalin (ENK) and GABA neurotransmitters simultaneously in the same ultrathin tissue sections in the periaqueductal gray area (PAG) of the rat brain. MATERIAL AND METHODS: Double gold particle signals were significantly improved using a number of technical adjustments for the immunogold electron microscopic co-localization technique of ENK and GABA. The GABA-like neuronal elements were immuno-reacted with 20 nm gold particles and the enkephalin-like immunoreactive neurons were labeled with 10 nm gold particles. This double labeling was more apparent in tissue sections that were deactivated for the gold staining of the first antibody. Excellent double labeling was obtained when we blocked antigenicity of the first antiserum with hot (80 degrees C) paraformaldehyde fumes. To minimize the clumping of the second gold particles around the first gold particles used against the first antibody, we tried different staining order for the neurotransmitters tested in this study. It was necessary to use a detergent (Triton X-100) at very low concentration (0.1%) instead of etching to expose the antigenic determinants of the neurotransmitters and at the same time to reduce the deleterious effects on the morphology of the tissue sections. Furthermore, the high-glutaraldehyde fixation and the decrease in the interval between cutting and labeling of the ultrathin sections significantly improved the results obtained in this study. RESULTS: Double labeling of sections with ENK and GABA produced co-localization in 23.1% and 1.2% of the immunoreactive axonal terminals and dendrites, respectively. Most of the double-labeled terminals contained more GABA-like than ENK-like immunolabeling. Half of the axon terminals [51%] and dendrites [56%] in the ventrolateral PAG were not labeled with either of GABA or ENK immunoreactivity. CONCLUSIONS: This procedure was found to be completely compatible with good double immunolabeling and ultrastructural preservation. PMID- 11257720 TI - Influence of the diet made by its family ancestors in the hereditary component of an individual: study in six generations of rats. AB - BACKGROUND: The prevalence of the deficiency of enzyme Lactase/phlorizin hydrolase (LPH) varies widely between different countries, with a low of 1-3% in Scandinavia and close to 100% in most of South-East Asia. Various carbohydrates are capable of enhancing LPH mRNA levels in the small intestine, and that transcriptional control plays a major role in the carbohydrate-induced alterations of LPH mRNA expression. MATERIAL AND METHODS: A generation of rats was randomized and assigned to two groups; those that were fed a high carbohydrate diet and, those that were fed standard rodent meal. The sixth generation of these animals was fed standard rodent meal. Transfection experiments using Caco-2 cells were performed using sperm samples of the sixth generation of rats. RESULTS: This study suggests that the feeding with a high carbohydrate diet during five generations of rats increases the capacity of production of enzyme LPH, LPH mRNA levels and the transcription rate of the LPH gene in the sixth generation of these animals, and this fact happens independently of the diet that this generation of rats had. Transfection experiments show that this influence has had to take place necessarily within the hereditary component which the last generation of rats received from its family ancestors. CONCLUSION: The feeding received by the ancestors of a generation of rats could influence within hereditary component received by them. PMID- 11257722 TI - Effect of different drugs on the level of DNA-hydrolyzing polyclonal IgG antibodies in sera of patients with Hashimoto's thyroiditis and nontoxic nodal goiter. AB - BACKGROUND: DNA-hydrolyzing IgG antibodies have been detected recently in the sera of patients with several autoimmune diseases. MATERIAL AND METHODS: The relative activity of DNA-hydrolyzing IgG from the sera of patients with Hashimoto's thyroiditis and with non-toxic nodal goiter as a function of the patient's condition were measured. The effect of different drugs on the level of DNA-hydrolyzing IgG, functional activity of thyroid gland, and improvement of clinical condition of two groups of patients were analyzed. RESULTS: We demonstrate here for the first time that IgG from peripheral blood of patients with Hashimoto's thyroiditis (65%) and non-toxic nodal goiter (38%) possesses DNAse activity. The relative level-specific activity of IgGs in hydrolysis of DNA increases with the enhancement of the relative amount of antibodies against thyroglobulin and all patients with hypothyroidism (namely a reduced concentration of thyroxine and triiodothyronine and enhanced level of thyrotropic hormone) are characterized by a high level of catalytic antibodies. CONCLUSIONS: The very widely used therapy of patients with thyroxine led only to a temporary change of the hormone concentration in the blood but did not affect the level of DNA-hydrolyzing antibodies. However, treatment with an immunosuppressive drug plaquenil (7-chloro-4(beta-diethylamine-alpha-methylbutylamie)quinoline), significantly decreased the DNA-hydrolyzing activity of Abs, which correlated with enhancement of thyroid hormone concentrations, elevation of functional activity of the thyroid gland, and improvement of the clinical state of the patients. PMID- 11257723 TI - Effect of sevoflurane on intracranial pressure and cardiovascular function in rabbits with experimental intracerebral haematoma. AB - BACKGROUND: Sevoflurane is one of the newest volatile anaesthetic agents. The effect of sevoflurane on ICP in conditions of intracranial pathology is essential from the clinical point of view but still not sufficiently clear. The aim of the study was to examine the effects of 1 MAC, 2 MAC, 3 MAC sevoflurane on intracranial pressure (ICP), mean arterial blood pressure (MABP) and heart rate (HR) in rabbits with experimental intracerebral haematoma (ICH). MATERIAL AND METHODS: The experiments were performed in 13 adult rabbits, 3.5-4.0 kg weight. The rabbits were randomly allocated to two different groups. In group I, (n = 7), sevoflurane was administered in stepwise increasing concentrations of 2.2 vol%, 4.4 vol% and 6.6 vol%, each for a period of 15 minutes. In group II (n = 6), intracerebral haematoma was produced and subsequently sevoflurane was administered in the same manner as in group I. Ventilation parameters, inspiratory and end-tidal sevoflurane concentration, end-tidal CO2 concentration (ETCO2), HR, MABP, ICP and body temperature, measured in the nasopharynx, were monitored throughout the experiment. RESULTS: Mean values of ETCO2 and temperature in the nasopharynx were not significantly different in both groups and remained stable in the whole observation period. In group II in all cases the evidence of intraventricular haematoma was observed. In this group mean values of ICP, MABP and HR after haematoma production were significantly higher than those in group I. Statistically significant increase of ICP was observed in 30th minute in group I, while in 35th minute in group II. In both groups a statistically significant decrease in MABP was observed from 20th minute of observation. A significant decrease in HR in both groups from 25th minute of experiment was also observed. CONCLUSION: In conclusion it should be stressed that sevoflurane, in doses not exceeding 1 MAC, shows no significant effect on ICP and cardiovascular function in the course of intracranial haematoma. PMID- 11257724 TI - Nitric oxide synthase type 1 expression in human lung cancer and its relation to p53. AB - BACKGROUND: The nitric oxide synthases (NOS) have been observed in human tumour cell lines as well as in solid tumours. Neuronal isoform of NOS (NOS1) was particularly abundant in low-differentiated gynaecological, breast and central nerve system tumours, suggesting that it may characterize poorly differentiated tumours. So far, little is known about expression of the neuronal NOS isoform in non-small cell lung cancer. Our aim was to compare NOS1 expression in non-small lung carcinomas with respect to tumor staging and p53 protein expression. MATERIAL AND METHODS: Thirty-two cases of non-small lung carcinomas of all grades of malignancy and ten control lung specimens with neoplastic lesions were examined. Paraffin-embedded tissues were stained with hematoxylin and eosin, or with antibodies to NOS1 and p53, and evaluated under light microscope. RESULTS: No statistical correlation between expression of p53, NOS1 and degree od tumour differentiation was observed. CONCLUSION: Expression of NOS1 can not serve as a marker for highly malignant tumours in the non-small cell lung carcinomas. PMID- 11257725 TI - Search for enterotoxin gene in Bacteroides fragilis strains isolated from clinical specimens in Poland, Great Britain, The Netherlands and France. AB - BACKGROUND: Bacteroides fragilis is a member of normal human flora and well known pathogenic agent. This bacterium produces many virulence factors. In 1984 new virulence factor--enterotoxin was described. The aim of the study was to search for enterotoxin gene in B. fragilis strains isolated from clinical specimens. MATERIAL AND METHODS: Strains isolated in Poland, Great Britain, France and the Netherlands were cultured on BBE medium. For DNA isolation Genomic DNA PREP PLUS isolation kit manufactured by A&A Biotechnology (Poland) was used. In order to detect enterotoxin (fragilysin) gene, polymerase chain reaction (PCR) was applied utilizing the following primers: 404 (GAG CGG AAG ACG GTG TAT GTG ATT TGT) and 407 (TGC TCA GCG CCC AGT ATA TGA CCT AGT). DNA obtained from bacterial cells was amplified in thermocycler Techne. The amplification products were detected by the electrophoresis in 1% agarose gel. RESULTS: Among 65 investigated B. fragilis strains, the enterotoxin gene was detected in DNA isolated from 12 strains. CONCLUSION: The enterotoxin producing B. fragilis strains were detected among strains isolated from different clinical specimens in Poland, Great Britain, the Netherlands and France. PMID- 11257726 TI - Regional cerebral glucose metabolism of patients with malignant diseases in different clinical phases. AB - BACKGROUND: Psychological and psychiatric aspects of cancer patients have not been studied well in terms of functional neuroimaging. The aim of this pilot study was to investigate the relationship between regional cerebral metabolism and different clinical phases. MATERIAL AND METHODS: Relative cerebral glucose metabolism of 77 Japanese patients with various types of malignant diseases was studied by positron emission tomography with 18F-fluorodeoxyglucose. They were subgrouped into the 1) pre-treatment, 2) post-treatment, 3) recurrence and 4) terminal patient groups. These subgroups were compared to the control group of 17 in-patients with benign diseases, using voxel-based Statistic Parametrical Mapping software (SPM). RESULTS: Relative reduction in the regional cerebral metabolism was detected in the prefrontal and basolateral (inferolateral) prefrontal cortex, orbitofrontal cortex, anterior and posterior cingulate gyrus, insula, basal ganglia, hippocampus and thalamus in the pre-treatment group. The anterior and posterior cingulate gyri were hypometabolic only in the pre treatment group. Hypometabolic areas were detected only in the basolateral prefrontal cortex, orbitofrontal cortex, ventral part of cingulate gyrus and insula in the post-treatment group. In the terminal group, the hypometabolic pattern seemed very close to that in the pre-treatment group. The results seemed to suggest that hypometabolic findings before treatments could reverse after treatments but became prominent again in terminal stages. These results do not contradict with previous epidemiological findings that high incidence of adjustment disorders was seen in earlier stages and more brain organic syndromes were seen in terminal stages. CONCLUSION: It could be speculated that decreased metabolism in early stages reflects state-dependent changes and that decreased metabolism in the terminal stages reflects organic brain damages. brain of cancer patients show a fluctuation in the regional metabolism. This finding might give a suggestion that functional imaging could be used as a supplementary method for psychological evaluation of patients with severe diseases. PMID- 11257727 TI - Vitamin d-receptor gene polymorphisms and vertebral bone density in men with idiopathic hypogonadotrophic hypogonadism. AB - BACKGROUND: Optimal peak bone mass is closely related to sufficient and appropriately timed androgen release. However, attainment of peak bone mass in men, as in women, is under genetic control, as well as being subject to hormonal and mutational effects. With increasing recognition of osteoporosis and related fractures in men, it is of interest to consider whether there is relationship between bone density and vitamin D receptor (VDR) polymorphisms, as described in women. MATERIAL AND METHODS: To assess the influence of allelic variation in the VDR gene on vertebral bone density in men with idiopathic hypogonadrotrophic hypogonadism (IHH), 27 patients (mean age 21.4 +/- 0.4 yrs) and 25 age-and-BMI matched healthy males (mean age 21.2 +/- 0.3) were genotyped using three restriction enzymes (Apa I, Bsm I, and Taq I). Vertebral bone mineral density was measured using dual energy X-ray absorptiometry (DEXA). RESULTS: As expected, vertebral bone density was reduced significantly in patients with IHH (p < 0.001). Despite weak evidence for an association between Apa I polymorphism and vertebral bone density in the IHH group (r = 0.454, p = 0.017 and r2 = 0.20), VDR genotype was not associated with vertebral bone density in either group. When analyzing homozygous haplotypes, the probability of carrying the favorable BAt haplotype was greater in the control group (OR = 2.000 vs. 0.500). CONCLUSION: We conclude that VDR genotype has no influence on vertebral bone density in men with IHH. Thus, allelic variation in the VDR cannot help define those at increased risk for osteoporosis and related fractures among such patients. PMID- 11257728 TI - Molecular analysis of SRY gene in Brazilian 46,XX sex reversed patients: absence of SRY sequence in gonadal tissue. AB - BACKGROUND: The importance of the Y chromosome in male determination has been well established for a long time. The presence of a translocation of chromosomal material encoding the Testis-Determining Factor from Y to another chromosome has been one of the hypothesis to explain testicular development in XX sex-reversed patients. MATERIAL AND METHODS: In the present study, we searched for SRY sequence in genomic DNA isolated from peripheral leukocytes in eleven 46,XX true hermaphrodites and four 46,XX males (only one with ambiguous genitalia). We also analyzed the presence of SRY sequence in fresh gonadal tissues from two 46,XX true hermaphrodites. RESULTS: SRY sequence was absent in DNA blood samples of all true hermaphrodites and in testicular and ovarian tissues of two cases studied. Of the four 46,XX males, two with normal male external genitalia were SRY positive. CONCLUSIONS: We did not identify the SRY gene in 46,XX true hermaphrodites and 46,XX males with ambiguous genitalia, therefore SRY translocation to X chromosome or autosome is unlikely. Hidden Y mosaicism in gonadal tissues was also ruled out in two cases, suggesting that cryptic SRY mosaicism in gonadal tissues is not the usual mechanism responsible for testicular development in patients with 46,XX true hermaphroditism. However, SRY gene was identified in two 46,XX males with male external genitalia showing that SRY gene determined their male phenotype. Despite the recent advances in the knowledge of the role of several genes involved in sexual determination we are still unable to explain the cause of most of Y-chromosome-negative 46,XX sex reversed patients. PMID- 11257729 TI - Detection of etiological agent for cholera by PCR protocol. AB - BACKGROUND: PCR protocol for Vibrio cholerae, the causative agent of the diarrheal disease cholera has been described in this report. We report the detection of Vibrio species in drinking water samples in a duplex PCR reaction. The target loci used in the study were ctxA and tcpA. The sensitivity and efficiency of detection of this protocol can be applied in epidemic conditions, wherein monitoring of target organisms is very crucial. MATERIAL AND METHODS: Single step thermocycling programs have been reported for amplification of specific target loci. We have demonstrated that a gradient multi-step thermocycling program is more efficient in improving the sensitivity of detection by PCR. The method for preparation of template DNA from environmental sample uses filtration of water followed by harvesting the possible bacterial residue. This was further treated with proteinase K and heat to yield DNA compatible for PCR. The protocol was optimized for amplification of target loci from standard strains as well as from environmental water samples. RESULTS: The method can simultaneously detect two different loci of Vibrio cholerae in a single reaction. The sensitivity of detection achieved for the pathogen was 100 cells per reaction. The specificity of the primers was demonstrated by spiking the reaction with non-specific template. The developed protocol was successfully extended to environmental samples. CONCLUSIONS: The developed duplex PCR protocol allows the simultaneous detection of two genetic loci of the target pathogen from water samples. This enhances the efficiency of detection and provides a sensitive tool for the rapid detection of the pathogen that can be useful in epidemic conditions where the time factor involved in the identification of target pathogen is very crucial. PMID- 11257731 TI - Expression of Fas antigen on T cell subpopulations in peripheral blood of patients with relapsing-remitting multiple sclerosis. AB - BACKGROUND: During the relapse of multiple sclerosis, the activation of T cells, autoreactive to myelin antigens in blood, enhanced and maintained as a result of anomalous mechanisms of their earlier elimination, leads on para- and autocrine basis to the activation of antigen- non-specific cells of immune system. In consequence, activated cells secrete a range of proinflammatory cytokines and display activation antigen expression on their surface, which results in blood brain barrier damage. The differentiation of lymphocytes into effector cells in blood during MS relapse is to increase the number of cells supporting inflammatory reactions and simultaneously to reduce the number of cells which play a role of suppressors. Fas antigen is present among activation antigens found on T cells. Once this antigen has been combined with the ligand, it transmits apoptic signal to the cell. The presence of Fas antigen on activated peripheral blood T cells may enable us to estimate their activation and it may also indicate a potential to eliminate those cells from blood. The aim of the study was to provide a quantitative assessment of the subpopulations of CD3, CD4 and CD8 lymphocytes in peripheral blood and to investigate Fas antigen expression on these subsets in patients with relapsing-remitting multiple sclerosis, in relation to clinical activation of the disease. MATERIAL AND METHODS: Thirty-five patients participated in the study, including 14 patients finding themselves in clinical relapse of the disease and 21 patients in the state of remission. Additionally, 21 healthy subjects were included. Quantitative assessment of individual subpopulations and Fas co-expression was carried out with the use of monoclonal antibodies anti CD3, CD4 and CD8 as well as anti CD95 antibodies, and flow cytometer Pas/Dako Galaxy. RESULTS: The differences in the percentage of particular lymphocytes between 3 groups proved insignificant. Patients in the relapse of the disease showed significantly greater Fas expression on subpopulations CD3 and CD4 when compared to the results obtained from remission patients and control subjects. This difference was not observed for Fas expression on subset CD8. CONCLUSIONS: The investigation of Fas receptor expression may be useful in order to monitor clinical course of the disease, which is characterised by the periods of exacerbation and remission. PMID- 11257730 TI - Search for mitochondrial A3243G tRNA(Leu) mutation in Polish patients with type 2 diabetes mellitus. AB - BACKGROUND: The influences of genetic and environmental factors form a clinical picture of type 2 diabetes mellitus. Genetic studies of type 2 diabetes mellitus become increasingly important. The knowledge of the molecular background of type 2 diabetes has been growing rapidly over recent years. One of the forms of the disease defined on the molecular level is maternally inherited type 2 diabetes mellitus. This diabetes, which is frequently accompanied by hearing impairment of deafness (maternally inherited diabetes with deafness-MIDD), was linked with sequence differences in mitochondrial DNA. The most frequent cause of MIDD is A3243G substitution in a mitochondrial tRNA(Leu) gene. While this mutation was identified in different races in several populations, it is still important and valuable to evaluate its prevalence in various ethnic groups. The aim of the project was to determine the prevalence of A3243G substitution in a mitochondrial tRNA(Leu) gene among Polish diabetic subjects. MATERIAL AND METHODS: In total 129 individuals, with type 2 diabetes and 12 with gestational diabetes were selected for this study. Two techniques based on restriction fragment length polymorphism (RFLP) method were used to screen for A3243G mutation. In the first approach, non radioactive PCR reactions of mitochondrial DNA region of interest were performed using DNA of the study participants. This was followed by Apa I restriction enzyme digestion of the PCR product. Subsequently an electrophoretic separation was done on 2% agarose gel with ethidium bromide staining. In the second, more sensitive, modification of RFLP, [alpha 32P]dCTP was used for internal primer labeling and the electrophoresis was done on acrylamide gel. A positive sample was used to control the quality of the genotyping. RESULTS: In both approaches none of the samples, except for the positive control, showed the evidence of the G variant. CONCLUSIONS: In summary, the A3243G mutation in mitochondrial tRNA(Leu) gene is not a frequent cause of diabetes in the Polish population. Further screening of enlarging study group is necessary to fully determine the prevalence of this mutation in our population. This, together with the search for other mitochondrial mutations, should allow to fully determine the prevalence of MIDD and its specific molecular background in the Polish population. PMID- 11257732 TI - Thrombomodulin--endothelial thrombin receptor in blood of patients with unstable angina pectoris. AB - BACKGROUND: Thrombomodulin (TM) a membrane receptor for thrombin generated in blood in vivo, is present on the surface of vascular endothelium cells. The aim of our work was the determination of thrombomodulin in the blood of patients with unstable angina pectoris. MATERIAL AND METHODS: In the study took part 87 patients with unstable angina pectoris (40 women and 47 men) at age 41-79 years. Thrombomodulin was determined in citrated blood plasma with the use of enzyme immunoassay ELISA with the diagnostic kit manufactured by American Diagnostica. RESULTS: Statistically significant higher thrombomodulin concentration (sTM) was found in the patients when compared with the values recorded in control group. Elevated sTM levels depended on patients age and they were significantly higher in patients over 50 years old. There were no statistically significant differences with respect to patients gender and coexistent risk factors such as arterial hypertension, diabetes or smoking, but significantly higher sTM concentration was observed in patients with high serum cholesterol level. CONCLUSION: Elevated sTM concentrations result from the damage to vascular endothelium cells by the atheromatous process manifested in unstable angina pectoris. PMID- 11257733 TI - Computer analysis of normal and basal cell carcinoma mast cells. AB - BACKGROUND: Basal cell carcinoma (BCC) is the most frequently observed neoplasm of the human skin. Mast cells are regarded as important cellular elements of all organs and tissues of the human body. They are involved in many physiological and pathological processes, including inflammation and fibrotic changes observed in BCC. The aim of our study was to characterise in detail mast cells observed in BCC and normal skin on the basis of computer analysis techniques. MATERIAL AND METHODS: Series of photographs obtained from histopathological biopsies of basal cell carcinomas from 18 patients, stained specifically for mast cells with Astra blue and nuclear red, were employed for the study. Control group consisted of series of photographs from normal skin obtained from 18 persons who underwent plastic surgery operations. All the photographs underwent digital computer analysis by means of a number of subroutines written in Delphi developed at the Technical University of Lodz. RESULTS: Specific staining allowed for thresholding the image and subsequently mast cells were visualised as black objects on a white background. Those objects were then analysed in detail i.e. their geometrical features (area, perimeter, maximal diameter) were calculated. The employed image analysis techniques allowed for the differentiation and detailed characteristics of mast cells stained with Astra blue and nuclear red. CONCLUSIONS: We observed that mast cells in BCC gathered mainly in fibrous stroma and were more numerous, had considerably larger area, longer perimeter and maximal diameter than normal skin mast cells. PMID- 11257734 TI - Effect of fungal colonization of gastric mucosa on the course of gastric ulcers healing. AB - BACKGROUND: The aim of the study was to evaluate the effect of fungal colonization on the course of gastric ulcer disease with particular regard to regression of clinical symptoms and reduction in ulcer niche size. MATERIAL AND METHODS: The study was performed on the group of 293 patients, aged 20-80, with clinical symptoms of peptic ulcer disease, who attended Gastroenterology Outpatient Department of Collegium Medicum of Krakow Jagiellonian University. Endoscopy of the upper gastrointestinal tract was performed before and following a-4-week period of a standard anti-ulcerous treatment. During the examination aspirate of the stomach contents and surface brushing were collected. Moreover, biopsy specimens from the bottom of the ulcer or inflamed gastric mucosa were taken for mycological and histopathological examinations. According to contemporary recommendations eradication treatment of Helicobacter pylori was introduced in urease-positive patients with endoscopy-proved gastric ulcer. RESULTS AND CONCLUSIONS: The results of our research showed that fungal colonization of gastric ulcers impairs the course of ulcer healing. Moreover, it results in clinical symptoms maintenance as compared with ulcers with non significant fungal cells count. PMID- 11257735 TI - Omeprazole therapy and salivary flow rate in duodenal ulcer patients. AB - BACKGROUND: Previous reports have shown that in some reflux-oesophagitis patients omeprazole therapy alters salivary secretion. The aim of the study was to examine this effect in duodenal ulcer patients. MATERIAL AND METHODS: Thirty nine Helicobacter pylori positive subjects of both sexes, predominantly men, were recruited for the study. They were taking for two weeks only omeprazole (n = 17), or omeprazole in combination with either amoxycillin or amoxycillin and tinidazole (n = 22). Salivary secretion was assessed before and at the end of the treatment, both in basal conditions and during a gastric secretory test. Gastric secretion was monitored concurrently with salivary flow rate. Additionally gastritis score and serum gastrin levels were assessed. RESULTS: Basal salivary secretions remained unchanged in patients on omeprazole monotherapy, but decreased in five of eight saliva collection periods in patients on eradication regimens. During the gastric secretory test, salivary secretions fell in both groups, but only after pentagastrin stimulation (in one collection period in patients on omeprazole, and in three collection periods in patients on eradication therapy). The observed changes in salivary secretion were inversely related to the pre-treatment gastric pH values. CONCLUSION: The influence of omeprazole and omeprazole-based eradication therapies on salivary flow rate is presumably secondary to changes in gastric pH values and is likely to be related to oesophago-salivary reflex generation. PMID- 11257736 TI - Pentoxyfilline treatment does not influence the plasma levels of IL-2 and sIL-2R in limited scleroderma patients. AB - BACKGROUND: In this study we investigated whether plasma level of IL-2 or sIL-2R shows the clinical disease status. MATERIAL AND METHODS: Plasma levels of IL-2 and sIL-2R were measured by ELISA in 23 patients with limited scleroderma (1SSc acrosclerosis) and compared with 12 healthy women. RESULTS: Plasma levels of sIL 2R in the group of patients with severe internal organs involvement (3446.1 +/- 1329.7 pg/ml) were significantly higher compared with those with minimal internal organs involvement (1606.4 +/- 507.3 pg/ml). Plasma level of IL-2 in the group of patients with severe internal organs involvement (8.12 +/- 7.77 pg/ml) did not differ much from that of the patients with minimal internal organs involvement (7.42 +/- 9.28 pg/ml). Treatment with pentoxyfilline did not influence the plasma levels of IL-2 and sIL-2R. CONCLUSIONS: We concluded that elevated plasma levels of IL-2 and sIL-2R are markers of internal organ involvement in scleroderma. PMID- 11257738 TI - Reversibility of asthma-like symptoms and lung function growth over two-year follow-up in preadolescent children. AB - BACKGROUND: The interesting feature of childhood asthma is the great reversibility of symptoms. The main purpose of this study was to assess the effect of persistent and reversed asthma-like symptoms on lung function growth in preadolescent children. MATERIAL AND METHODS: The follow-up respiratory health survey has been conducted in the sample of 1129 children aged 9 yrs over two years follow-up. The basic health end-points was the occurrence of asthma-like symptoms and the slower lung function growth (SLFG). RESULTS: Adjusted OR for SLFG(FVC) was significantly higher only in the children having the continued symptoms(OR = 3.40; 95% CI: 1.32-8.77). There was a consistent trend of adjusted ORs for SLFG(FEV1) with the category of symptoms, where OR was 1.46 (95% CI: 1.02 2.10) in children with recent new symptoms; 2.06; (95% CI: 0.93-4.58) in those with symptoms remitted, while 3.46; (95% CI: 1.43-8.40) in children who had persistent symptoms. The corresponding ORs for SLFG(FEF25-75%) were 2.03; (95% CI: 0.97-4.28), 2.63; (95% CI: 1.38-4.99), and 5.84; (95% CI: 2.53-13.50). CONCLUSIONS: The consistent association between reversibility of asthma-like symptoms and lung function gain in preadolescent children confirmed the clinical significance of the symptoms in question. The observed slower lung function gain in preadolescence may have implications for the development of chronic lung disease later in adulthood. PMID- 11257737 TI - Are antibiotics necessary in nonperforated appendicitis in children? A double blind randomized controlled trial. AB - BACKGROUND: The use of antibiotics in uncomplicated appendicitis in children, remains the area of controversy. The aim of the study was to assess the necessity of antibiotic administration in nonperforated appendicitis in children. MATERIAL AND METHODS: The design of the study was a double blind randomized controlled trial, with a follow-up of 4 to 20 months. SETTING: Surgical Department in a University Pediatric Hospital. PATIENTS: One hundred and eighty seven out of 249 children subjected to emergency appendectomies met the inclusion criteria, with 35 eligible but not included in the study. The remaining 152 patients were randomized; 41 had complicated appendicitis, 3 other diagnosis, 108 were analyzed within 3 study groups: 1 (n = 31) no antibiotic, 2 (n = 41) one dose, 3 (n = 36) 5-day course. Open appendectomy was a surgical procedure and Ceftriaxone 1.0 g i.v. was administered. Investigated parameters were: body temperature, WBC, bowel sounds, wound healing, recovery and morbidity. RESULTS: Valid outcome data were available for 90 of 108 randomized patients. Protocols of 18 children due to fever > 39 degrees C, upper airway infection or allergy were disclosed. In the remaining 90 children, there were no differences in WBC and oral feeding between groups 1 (n = 24), 2 (n = 35) and 3 (n = 31). Group 1 and 2 had a higher mean temperature on day 1 post-op, without any clinical significance. A higher mean temperature was noted on day 5 post-op in group 1, due to wound infection in one patient. There were no intraabdominal abscesses. The only other complications were 2 adhesion small bowel obstructions (in groups 1 and 2 each). CONCLUSION: Routine use of antibiotics in nonperforated appendicitis in children is not necessary. PMID- 11257739 TI - Case of spinocerebellar ataxia type 1 showing high intensity lesions in the frontal white matter on T2-weighted magnetic resonance images. AB - We report a case of genetically confirmed spinocerebellar ataxia type 1 (SCA1) in which magnetic resonance imaging (MRI) demonstrated a high signal intensity on T2 weighted images in the white matter of the frontal lobes. The patient was a 60 year-old Japanese man who complained of gait instability and speech difficulties. He was diagnosed as having spinocerebellar ataxia at the age of 46. A CAG repeat number of the patient was 48/26. Brain MRI showed marked atrophy of the cerebellum and brain stem. The high-signal intensity lesions on T2-weighted MRI in the white matter of the frontal lobes were evident in the periventricular regions. Such MRI abnormalities have not been described in SCA1 previously. PMID- 11257740 TI - Pituitary tumor in a woman with a 47,XXX karyotype--case report. AB - BACKGROUND: The 47,XXX karyotype is a rare sex chromosome anomaly. This karyotype is usually not associated with a characteristic physical phenotype. CASE REPORT: In presented case a 47 triple X women with pituitary tumor and premature ovarian failure is identified. Diagnosis of a 47,XXX individual remains difficult because specific clinical criteria used to identify this condition are not available. CONCLUSIONS: The case described should attract attention to how difficult it is to diagnose properly a genetic disease in young women with correct phenotype. PMID- 11257742 TI - Verification of selected anatomic landmarks used as reference points for universal goniometer positioning during knee joint mobility range measurements. AB - The study was concerned with verification of the selection of reference points used for knee joint mobility range goniometry. The verification was based on photometric and electrogoniometric methods of femorotibial angle measurements. The material for measurements were three knee joint preparations. The obtained data were subjected to descriptive analysis; photographic documentation of geometric relations between the reference points was also prepared. Considerable divergence of measurement results was observed with respect to the actual knee joint flexion angle, amounting even to 16 degrees. The differences are due to the selection of anatomic landmarks, and depend in particular on the distance of the point to which the goniometer axis is applied from the approximate location of the mechanical joint axis. The corrections of knee joint flexion angle measurements were also estimated on the basis of data obtained from the photographs and the derived mathematical formula. PMID- 11257741 TI - Massive fetomaternal transplacental hemorrhage as a perinatology problem, role of ABO fetomaternal compatibility--case studies. AB - BACKGROUND: Massive fetomaternal transplacental hemorrhage is not simply a problem of possible alloimunization in Rh incompatibility but also endangers the fetus (newborn) by massive anemization. Bleeding from placental vessels can occur after small trauma to the gravid uterus with mild or no clinical signs (bleeding or spotting, pain, hypertonus). The rupture of anchoring villi related to early uterine contractions is also possible. In the case of slow blood loss, the fetus reacts by adequate or inadequate compensatory reactions (hydrops fetus). Rapid and massive blood loss is followed by perinatal hypoxic damage and finally death. Our goal was to map out the diagnostic and therapeutic possibilities in regard to specific neonatal care. CASE REPORT: We evaluated four cases of fetomaternal transfusion during a 2-year period with special regard to postpartum adaptation of the newborn and the perinatal outcome. The incidence of adverse outcomes following massive fetomaternal transplacental hemorrhage was 50% (2 of 4). There was one perinatal death and one infant was affected by spastic quadriplegia. CONCLUSIONS: For diagnosis, it is possible to use cardiotocography (decreased variability, sinusoid pattern), ultrasound (biophysical profile) and special hematological tests for quantitative determination of fetal erythrocytes in the maternal blood. For the treatment of such cases one should consider premature termination of pregnancy or intraumbilical transfusion. PMID- 11257743 TI - Injury in vascular surgery--the intimal hyperplastic response. AB - Intimal hyperplasia is extensively studied in order to improve arterial reconstruction outcome. The mechanisms leading to stenosis or restenosis may vary according to the technique used for arterial reconstruction. Lesions are mostly made of an accumulation of smooth muscle cells and fibroblasts, with only sparse inflammatory cells. The accumulated material reduces the graft lumen and ultimately induces thrombosis. Intimal hyperplasia with smooth muscle cell and matrix accumulation is the prominent feature in all these situations with evidences of intense cell proliferation and cell death. The purpose of this review is to present the biology of intimal hyperplastic response based on the recently published data. Experiments in the rabbits have shown that the vein wall thickening is mainly regulated by the tangential wall stress which is applied transversely to the vein wall as a blood pressure. Experiments in the rat carotid artery balloon injury suggested that heparin could be used as a treatment to prevent intimal hyperplasia. Treatments for preventing restenosis after angioplasty or stenoses development in bypasses have been disappointing clinical evaluation suffers from insufficient prospective randomized studies. Intimal hyperplasia is the major cause of failure after arterial reconstruction. The biology of intimal hyperplasia is complex, and treatment disappointing. Some types of hyperplasia may need to be preserved in order to prevent functional atrophy and aneurysmal dilatation of vein grafts. PMID- 11257744 TI - CD52 antigen--a review. AB - Human CD52 is a glycosylphosphatidylinositol (GPI)--anchored antigen expressed on the all lymphocytes as well as within the male genital tract, where it can be produced by epithelial cells of the distal epididymis and duct deferens. CD52 can be shed into seminal plasma and then acquired by sperm cells during their passage through the genital tract, thus it is detectable on the surface of epididymal sperm and in the ejaculate but not on either spermatogenetic cells or testicular spermatozoa. Protein core of the sperm and lymphocyte CD52 is identical and it is a product of a single copy gene located on chromosome 1. However, N-linked carbohydrate side chains (attached to Asn-3) and GPI-anchor structure are different. Physiological role of CD52 on lymphocytes is unclear, although antibody directed to CD52, CAMPATH-1, is capable of complement activation and it can be clinically useful to treat lympho-proliferative disorders and to deplete lymphocytes in bone marrow transplants. Monoclonal antibodies directed against sperm CD52 may cause sperm immobilisation and agglutination and affect hamster oocyte penetration suggesting an association of this molecule with fertilisation. PMID- 11257745 TI - New data on toxic metal intoxication (Cd, Pb, and Hg in particular) and Mg status during pregnancy. AB - The technological revolution we witness today poses a threat to the homo sapiens species, and its biological results are unpredictable. Excess toxic metals in the environment and the deficiency of bio-elements are particularly harmful for developing organisms. Long-term fetal exposure during pregnancy to even lower concentrations of toxic metals, which have the ability to accumulate, often leads to irreversible developmental disorders, On the basis of accessible literature, the paper presents transplacental transmission of cadmium, lead and mercury to the fetus. The disadvantageous effects of cadmium and lead on ionic transmission, functional potential and submicroscopic amnion structure as well as the interdependence between the unfavorable effects of these two metals on the amniotic membrane and the competitive antagonistic activity of Mg ions are emphasized. This paper presents a hypothesis suggesting the involvement of cadmium in the etiopathogenesis of eclampsia based on the literature. It also considers the present state of knowledge of the toxic effects of Cd, Pb and Hg on the course of pregnancy and fetal development. Magnesium--an intracellular cation second in importance to potassium plays a significant biological role, though it has not been fully explored yet. The concentration of Mg in the placental and fetal tissues increases during pregnancy. The requirements for this element in a pregnant woman's organism generally exceed its supply; hence, pregnancy should be considered a condition of 'physiological hypomagnesemia'. The accessible data concerning the content of Mg during pregnancy in the blood as well as in the uterine muscular wall in physiological and pathological pregnancies are diverse. The prevailing opinion is that oral supplementation of magnesium during pregnancy makes up for its deficit in the organism of the pregnant woman and also positively influences fetal development. It is recommended to administer magnesium with food in the form of magnesium salts at the dose 5 mg/kg body mass daily. In clinical obstetric practice magnesium therapy is necessary in cases of imminent preterm birth and preeclampsia. This paper discusses the mechanism and therapeutic effectiveness of magnesium sulfate as used in complications of pregnancy. The contamination of the pregnant woman's organism by toxic metals- cadmium, lead and mercury--poses a serious risk of the same quantitative degree of contaminating the organism of the child developing in her womb. Qualitative changes may be much more serious in the fetus as they affect young structures, intensively developing, with no well-formed defense mechanisms. It is also worth mentioning that the complications in the course of pregnancy may result from toxic metal concentrations lower than those leading to fetal necrosis or premature termination of pregnancy. PMID- 11257746 TI - Environmental health services in Europe 5. Guidelines for evaluation of environmental health services. PMID- 11257747 TI - [Tennis elbow]. PMID- 11257748 TI - [Risk of cancer after intake of uranium]. PMID- 11257749 TI - [Lateral humeral epicondylitis--"tennis elbow". I. Epidemiology, clinical picture and pathophysiology]. AB - Lateral epicondylitis (LE) is a common overuse syndrome of unknown etiology. Studies imply that it is mostly a degenerative condition implying that the term "epicondylitis" is a misnomer. LE often occurs in middle-aged persons in connection with acute or chronic strain. Symptoms are mostly related to occupational tasks. Other conditions can cause pain in the lateral elbow region, and it is important to know the differential diagnosis in order to start the proper treatment. The diagnosis is mainly based on clinical findings but may be substantiated by other diagnostic techniques, such as magnetic resonance imaging. However, the specificity and the sensitivity of these techniques are unclarified. PMID- 11257750 TI - [Lateral humeral epicondylitis--"tennis elbow"]. AB - There is a great variety of potential therapies in the treatment of lateral epicondylitis, surgical intervention being the most invasive. The most frequently applied therapies are corticosteroid injections, pain-relieving medication (NSAID) and physical therapy. Though many studies have been conducted, a "gold standard" in the treatment of LE is not available. Perhaps this is explained by a lack of methodological quality of many of these studies. PMID- 11257751 TI - [Comparison of two different treatments of lateral humeral epicondylitis--"tennis elbow". A randomized controlled trial]. AB - INTRODUCTION: Many therapeutic interventions have been used in the treatment of lateral epicondylitis (LE), but to date, no specific intervention has proved universally efficacious. The aim of this study was to compare local corticosteroid injections versus splinting in a randomized controlled design. MATERIAL AND METHODS: Sixteen patients were treated with injections, and fourteen with a wrist splint. The patients were evaluated at inclusion and after six weeks with grip strength and pain scores. The patient's global assessment was used as a main indicator of improvement. RESULTS: No differences were noticed between the two groups. Most patients improved. A questionnaire after > 1 year showed that some patients still had pain complaints. DISCUSSION: We conclude that injections were as effective as splinting in LE. Splinting is recommended in the early stages of the disorder because of its lack of adverse effects. In the long view diagnostics regarding LE have to be refined in order to differentiate patients who are expected to benefit from different treatments. PMID- 11257752 TI - [Cervical lymphatic metastases from occult primary tumor. A nation-wide 20-year study from the Danish society of head and neck oncology]. AB - INTRODUCTION: The management of patients with cervical lymph node metastases from unknown primary tumours is a major challenge in oncology. This study presents data collected from all five oncology centres in Denmark. MATERIALS AND METHODS: Of the 352 consecutive patients with squamous cell or undifferentiated tumours seen from 1975 to 1995, a total of 277 (79%) were treated with radical intent. Most patients received radiotherapy to both sides of the neck as well as elective irradiation of the mucosal sites in nasopharynx, oropharynx, hypopharynx and larynx (81%). Irradiation of the ipsilateral neck only was done in 26 patients (10%). Radical surgery was the only treatment in 23 N1-N2 patients (9%). RESULTS: The five-year estimates of neck control, disease-specific survival and overall survival for radically treated patients were 51%, 48% and 36%, respectively. The emergence of the occult primary was observed in 66 patients (19%). About half of the emerging primaries were within the head and neck region with oropharynx, hypopharynx and oral cavity being the most common sites. Emerging primaries outside the head and neck region were primarily located in the lung (19 patients) and oesophagus (five patients). The most important factor for neck control was nodal stage (5-year estimates 69% [N1], 58% [N2] and 30% [N3]). Other important parameters for neck control and disease-specific survival included haemoglobin, gender and overall treatment time. Patients treated with ipsilateral radiotherapy had a relative risk of recurrence in the head and neck region of 1.9 compared to patients treated at both neck and mucosa. At five years, the estimated control rates were 27% (ipsilateral) and 51% (bilateral; p = 0.05). The 5-year disease specific survival estimates were 28% and 45%, respectively (p = 0.10). DISCUSSION: Extensive irradiation to both sides of the neck and the mucosa in the entire pharyngeal axis and larynx resulted in significantly fewer loco-regional failures compared to patients treated with ipsilateral techniques, but only a trend towards better survival. Determination of the optimal strategy in terms of loco-regional control, survival and morbidity requires a prospective randomized trial. PMID- 11257753 TI - [Rupture of the symphysis after spontaneous delivery. Surgical treatment of four cases]. AB - Rupture of the symphysis pubis is a rare, but known, complication during labour. We describe four cases of rupture of the symphysis, for which surgical treatment was chosen shortly after labour (2-30 days). Internal fixation was done in two cases and external fixation in the other two. There were no postoperative complications. The patients were mobile soon after the operation, and had no pain and normal function at follow-up 6-12 months later. PMID- 11257754 TI - [Should we advice pregnant women not to intake coffee and other products containing caffeine?]. PMID- 11257755 TI - [Does Danish food lack vitamins and minerals?]. PMID- 11257756 TI - [On the cutting edge of development. Thrombolytic therapy for myocardial infarction]. PMID- 11257757 TI - [Concerning evidence-based surgery]. PMID- 11257758 TI - [Influence of age on serial change in TL/BMIPP dual isotope SPECT images after direct PTCA in patients with acute myocardial infarction]. AB - The purpose of this study was to investigate the influence of age on serial change in 201TlCl (TL) and 123I-BMIPP (BMIPP) dual isotope single photon emission computed tomography (SPECT) images after direct PTCA in patients (pts) with acute myocardial infarction (MI). Dual SPECT with TL and BMIPP at rest, radionuclide ventriculography for left ventricular ejection fraction (LVEF), and two dimensional echocardiography for wall motion analysis were performed in 26 pts at the subacute and chronic phases after direct PTCA for acute MI. A defect score (DS) for SPECT images was interpreted as normal: 0, mildly decreased: 1, moderately or severely decreased: 2, complete defect: 3. The difference in DS between TL and BMIPP was defined as the mismatch score (MS). DS in BMIPP was greater than that in TL at the subacute phase in all pts. Significant improvement in the wall motion score was recognized in pts who showed TL/BMIPP discrepancy at the subacute phase. Pts were classified by age into two groups; group I: younger than 65 years old (n = 18); group II: 65 years and older (n = 8). Improvement of MS from the subacute to chronic phase was significant in group I (5.2 +/- 1.9 to 3.2 +/- 1.9, p = 0.0001), whereas not significant in group II (6.2 +/- 2.9 to 6.1 +/- 2.9, NS). There was a significant negative correlation between relative MS (ratio of subacute MS to chronic MS) and age (r = -0.78, p < 0.0001). No significant correlation was observed between age and improvement in LVEF. These results indicate that disordered myocardial fatty acid metabolism, reflected by TL/BMIPP discrepancy, persist longer in elderly pts than younger pts after acute MI. PMID- 11257759 TI - [Development of neck filters for reducing artifact in cervical bone SPECT]. AB - In cervical bone scintillation SPECT studies using TEW and OS-EM methods, we have observed an artifact that may interfere with evaluation of the image; higher accumulation in cervical vertebra compared with in the head and thoracic vertebra. As the neck is smaller in diameter than in the thorax and head, gamma ray absorption is lower. In addition, as the distance between the neck and the detector is greater, scattered gamma rays are increased, interfering with imaging and causing artifact. To overcome these problems, we have developed special absorbers (neck filter) to make the relative absorption level of the neck comparable to that of the head and thorax and have employed these cervical filters in bone scintillation SPECT studies in combination with TEW scatter correction and OS-EM method. Our results showed that artifacts were significantly reduced and satisfactory images were obtained. PMID- 11257760 TI - [Left ventricular systolic wall motion after exercise stress and myocardial perfusion in patients with ischemic heart disease--investigation by ECG gated myocardial tomography]. AB - To investigate regional left ventricular (LV) wall motion (WM) after recovery from myocardial ischemia, we performed ECG-gated myocardial perfusion tomography with 99mTc-MIBI (G-SPECT) in patients with ischemic heart disease (IHD). In addition, we compared the left ventricular (LV) systolic function obtained by G SPECT at rest with that obtained by contrast left ventriculography (LVG). We performed G-SPECT at 30 minutes after exercise stress (Ex-30) and 3 hours after exercise (rest). LVWM and LV ejection fractions (EF) were analyzed by the QGS (quantitative gated SPECT) program. The LV was divided into 9 segments and regional WM (RWM) was analyzed quantitatively. In addition, myocardial perfusion was assessed quantitatively. In 64 patients with several different types of heart disease, EF obtained by G-SPECT correlated well with LVG-EF (r = 0.907, p < 0.001), and RWM of G-SPECT coincided well with that of LVG (kappa value 0.67, p < 0.01). Eighty patients with suspected IHD were divided according to Ex-Rest myocardial perfusion. In 83% of patients with Ex-induced perfusion abnormalities disappeared completely at rest, and in 58% of patients with Ex-induced abnormalities disappeared incompletely, RWM abnormalities which were observed at Ex-30 improved at rest and as did EF. In 79% of patients with a fixed defect (FD), RWM abnormalities and EF at Ex-30 did not differ with those at rest, but in 12% of the patients, the RWM abnormality of Ex-30 improved at rest. In most myocardial segments that had recovered from transient ischemia, RWM abnormalities persisted at least 30 minutes after Ex (stunning). In a small portion of the myocardial segments regarded as having myocardial necrosis because of a fixed perfusion abnormality, RWM abnormalities at Ex-30 improved at rest. These segments were supposed to contain viable myocardium. In conclusion, G-SPECT is a powerful method for clarifying the relation between the regional systolic function and myocardial perfusion. PMID- 11257762 TI - [A case of orbital MALT lymphoma in which 67Ga scintigraphy was useful for evaluating the radiation therapy response]. AB - A 67-year-old man was hospitalized at our institution complaining with epiphora and exophthalmos on the left side. Magnetic resonance imaging (MRI) showed an ill demarcated retrobulbar mass in the left orbit. 67Ga scintigraphy revealed avid uptake in the left orbital region. The patient was treated with radiation therapy. One month after the radiation therapy, the size of the mass decreased remarkably on MRI. 67Ga planar imaging after treatment showed no uptake, but 67Ga SPECT showed slightly increased uptake in the left orbital region. One year after the radiation therapy, MRI showed residual mass in the left orbital region. Both 67Ga planar imaging and SPECT showed no uptake in the left orbital region. 1.8 years after the radiation therapy, MRI showed the residual mass with no interval change in size. Both 67Ga planar imaging and SPECT showed no uptake in the left orbital region. The patient remains well with no evidence of local recurrence. 67Ga scintigraphy is useful in assessing the response to radiation therapy of MALT lymphoma in this case. PMID- 11257761 TI - [Evaluation of time dependency of the acetazolamide effect on cerebral hemodynamics as measured by 99mTc-ECD single-photon emission computed tomography]. AB - PURPOSE: Kuwabara et al. have examined the cerebral artery dilation with acetazolamide (ACZ) challenge test using PET. And, they reported that ACZ reaction came out time dependently. We have developed a unique SPECT's method using Technetium-99m ethyl cysteinate dimer (99mTc-ECD) to verify the results obtained by Kuwabara et al. METHOD: 1000 MBq of 99mTc-ECD was exactly divided into three syringes. Each of which was intravenous infused (i.v.) at rest, 7.5, and 20 minutes after ACZ administration. Data collection was started using dynamic SPECT immediately after 99mTc-ECD i.v. at rest. Using necessary data only, SPECT images representing each of the three 99mTc-ECD i.v. was reconstructed. SPECT counts were obtained by the ROI method from each images to calculate relative CBF from rest to 7.5 and 20 minutes after ACZ administration. RESULT: The 18 hemispheres of nine patients in the negative control group in whom ACZ was not loaded. CBF was stable during the three evaluation. The measurement error our method was estimated as small. The 18 hemispheres of nine patients in the positive control group who has normal vasodilatory reserve, CBF was increased by 26.2 +/- 8.1% at 7.5 minutes and 29.3 +/- 13.1% at 20 minutes after ACZ administration. Seven patients with and chronic stage unilateral internal carotid artery severe stenosis and/or occlusion were evaluated as the test group. Case of unaffected side, CBF was increased by 17.6 +/- 6.9% at 7.5 minutes and 24.8 +/- 11.3% 20 minutes after ACZ administration. And, increase rate of CBF in the affected side was 2.8 +/- 1.6% at 7.5 minutes and 17.3 +/- 5.0% at 20 minutes after ACZ administration. In the affected side, timing of the maximum CBF increase caused by ACZ was remarkably delayed. CONCLUSION: Our method based on 99mTc-ECD SPECT also revealed delayed cerebral artery dilation in the affected side. It was suggested that ACZ reaction came out time dependently, as reported by Kuwabara et al. PMID- 11257763 TI - [Sentinel node detection of patients with breast cancer by radionuclide method: consideration of radiation safety]. AB - Sentinel node was detected by 99mTc labeled nanocolloid in five patients with breast cancer. Surgery of breast cancer was done at 16 hours after the administration of 74 MBq of 99mTc labeled nanocolloid. Sentinel node was searched by scintigraphy prior to surgery and by gamma-probe during surgery. Radioactivity of injected site, sentinel nodes, blood contaminated gauze, and other garbage was measured by GM detector. Radiation to medical staffs was monitored by a pocket radiation detector and film batches. Sentinel nodes were successfully detected both by scintigraphy and gamma-detector. More than 70% of radioactivity remained in the administered site at 16 hours. Small amount of radioactivity was detectable in the sentinel node. Almost no radioactivity was detectable in blood contaminated gauze and other garbage. Radiation dose to the main surgeon was 4 to 6 microSv per surgery by a pocket radiation detector. Radiation dose to the assistant surgeon was 2 microSv per surgery. Radiation dose by labeling or injection was 0 to 1 microSv per procedure. No detectable radiation was measured by film batches. It is concluded that the detection of sentinel node by 99mTc labeled nanocolloid is a safe procedure from the point of radiation safety consideration. PMID- 11257764 TI - Combined grafting of thoracic aortic aneurysm and cardiac repair using continuous cold-blood coronary perfusion. AB - OBJECTIVE: For patients diagnosed with combined thoracic aortic aneurysms and cardiac lesions, we conduct a 1-stage operation for ascending and aortic arch grafting. We studied surgical outcome comparatively with patients undergoing aortic grafting alone. For descending and thoracoabdominal aortic grafting, we choose a 2-stage operation. SUBJECTS AND METHODS: Subjects were 80 patients undergoing ascending and aortic arch aneurysm repair between June 1994 and March 1999. Group 1 consisted of 30 undergoing simultaneous cardiac repair. Concomitant cardiac procedures involved 21 valvular, 5 coronary arterial, and 4 valvular and coronary arterial surgeries. Group 2 consisted of 50 undergoing aortic grafting alone. We used crystalloid cardioplegia and additional antegrade continuous cold blood coronary perfusion in Group 1, and crystalloid cardioplegia alone in Group 2. RESULTS: Hospital mortality was 10% in Group 1 and 2% in Group 2. Surgery length, cardiopulmonary bypass time, and aortic cross-clamping time in Group 1 were significantly longer than Group 2. Myocardial ischemic time did not differ significantly. Postoperative ICU stay, mechanical ventilation time and catecholamine support time did not differ significantly. Actuarial survival was 66.9 +/- 13.1% at 52 months in Group 1 and 87.2 +/- 4.8% at 57 months in Group 2 (p = 0.2918). CONCLUSION: Simultaneous cardiac repair and ascending and aortic arch aneurysm repair were conducted using continuous cold-blood coronary perfusion. Hospital mortality and mid-term survival did not differ significantly between groups. PMID- 11257765 TI - Clinical pathway for impalpable or small lung lesions treated with coil marking and thoracoscopy. AB - OBJECTIVES: Advances in computed tomography are detecting increasingly impalpable or small pulmonary lesions. We propose a clinical pathway for managing such lesions. METHODS: We conducted a retrospective study in a community teaching hospital to describe the hospital schedules of 18 patients having 19 lesions 10 mm or less and ground glass attenuation. Under computed tomography, a coil (Complex Helical Fibered Platinum Coil-18) was placed at the proximal side of the lesion. Using thoracoscopy and radiographic fluorography, we conducted partial lung resection targeting the coil the next day, adding lobectomy, if required. RESULTS: Final diagnosis included primary and metastatic lung cancer (n = 14), atypical adenomatous hyperplasia (n = 1), and benignancy (n = 4). Patients were admitted 2* days before surgery (*Numbers are medians). On postoperative day 3, chest tubes were removed. Epidural analgesia was continued for 5 days. On postoperative day 7, patients were discharged. Their admission charge was a total of yen 979,610. CONCLUSIONS: The hospital course above may be applied to the clinical pathway for managing impalpable or small lung lesions. PMID- 11257766 TI - Acute pulmonary thromboembolism with a floating right-heart thrombus. 4 surgical cases. AB - OBJECTIVE: We determined the efficacy of surgery for acute pulmonary thromboembolism with a floating right-heart thrombus. METHODS AND RESULTS: Thrombi were diagnosed by transthoracic echocardiography and electron beam computed tomography in 4 patients with acute pulmonary thromboembolism with a floating right-heart thrombus, and thromboembolectomy was done in all patients. Surgical procedure included intermittent deep hypothermic circulatory arrest. One patient died of endobronchial hemorrhage and 3 survived. CONCLUSION: Because acute pulmonary thromboembolism with a right-heart thrombus is life-threatening, immediate thromboembolectomy is required to decrease mortality. PMID- 11257767 TI - Combined coronary intervention in heart-transplant patient with rapidly accelerated cardiac allograft vasculopathy. AB - A 46-year-old man accepted for heart transplantation due to persistent cardiac failure from dilated cardiomyopathy underwent a transplant in Germany on July 13, 1995. The donor heart was suspected of coronary artery disease at explantation, but he could wait no longer because of his rapidly deteriorating hemodynamics. Postoperative coronary angiography revealed 25% stenosis of the left descending artery. He showed several episodes of minimal or moderate rejection postoperatively, and coronary angiography 15 months postoperatively showed rapidly accelerated cardiac allograft vasculopathy demonstrating triple vessel disease with multiple lesions. Percutaneous transluminal coronary angioplasty was successful on 2 coronary vessels, but immediately recurrent stenosis and new lesions involving the left main trunk occurred 6 weeks thereafter. Since he was financially unable to afford a second heart transplantation, quadruple coronary artery bypass grafting was conducted October 25, 1996. A biventricular assist device was used when he could not be weaned from cardiopulmonary bypass. He died of multiple organ failure 3 days after surgery. PMID- 11257768 TI - Mitral valve replacement in a patient with a collapsed lung. AB - A 67-year-old man, with a history of pulmonary tuberculosis since 18 years old, presented shortness of breath because of severe mitral regurgitation. Magnetic resonance imaging showed that the heart was displaced into the left thoracic cavity and rotated clockwise around its long axis. The forced expiratory volume per second was 1.06 l (46.7% of the predicted value) and the vital capacity was 2.48 l (72.1% of predicted value). Surgery was performed through a median sternotomy. An internal mammary artery harvest retractor was used to obtain operative exposure. Extensive pericardial suspension was used to push the over inflated right lung across the midline. Extracorporeal circulation was established. The mitral valve was replaced with a mechanical prosthesis. The patient was weaned easily from extracorporeal circulation and was extubated on the day of surgery. If preoperative respiratory function is adequate, cardiac surgery can be performed safely in a patient with only one functional lung. PMID- 11257769 TI - Revascularization of left subclavian artery for coronary subclavian steal syndrome. AB - We have experienced 2 patients with coronary subclavian steal syndrome which progressed each to a different prognosis. Both cases received percutaneous transluminal angioplasty for subclavian artery stenosis after coronary artery bypass grafting. Although one case is doing well without any symptoms, the other case required axilloaxillary artery bypass grafting for the subclavian artery restenosis. PMID- 11257770 TI - Mitral regurgitation after myocardial infarction. Coronary artery bypass grafting and mitral valve replacement with chordae preservation. AB - A patient with acute ischemic mitral regurgitation after acute myocardial infarction required emergency coronary artery bypass grafting and mitral valve replacement with chordae preservation. For severe mitral regurgitation and heart failure due to myocardial infarction and ischemic papillary muscle dysfunction, mitral valve replacement with chordae preservation was effective. Here, we discuss the etiology of ischemic mitral regurgitation and the operative method for valve repair or replacement. PMID- 11257771 TI - Aortobiventricular fistulas associated with pseudoaneurysm of the ascending aorta 12 years after patch repair of supravalvular aortic stenosis. AB - Fistula formation between the aorta and cardiac chamber is a rare complication of the ascending aortic aneurysm. A 27 year-old man undergoing successful patch aortoplasty for supravalvular aortic stenosis 12 years before admission had a high fever, infectious signs in blood laboratory data, and congestive heart failure. Transthoracic and transesophageal echocardiography revealed a pseudoaneurysm of the aortic root and aortobiventricular fistulas. Detachment of the proximal end of the aortic patch was thought to be the cause of the pseudoaneurysm. Debridement of necrotic tissue surrounding fistulas produced large defects in the anterior wall of the right ventricle, interventricular septum, and ascending aorta. A modified Konno operation effectively reconstructed the outflow tracts of both ventricles and the aorta. PMID- 11257772 TI - Coronary arteriovenous fistula with annuloaortic ectasia and aortic regurgitation. AB - Cardiac catheterization in a 57-year-old man disclosed a left coronary artery coronary sinus fistula with annuloaortic ectasia and severe aortic regurgitation. During surgery, the fistula vessel was exposed and clamped before infusing cardioplegic solution for cardiac arrest. The proximal orifice of the fistula was closed directly and the distal orifice doubly ligated. A modified Bentall's procedure was conducted without difficulty. The postoperative course was uneventful. PMID- 11257773 TI - Pulmonary thromboembolism after one and a half ventricle repair. Successful catheter-directed thrombolysis. AB - Acute pulmonary thromboembolism in a patient who had undergone bidirectional Glenn anastomosis was treated by percutaneous selective intravascular thrombolysis. A 20-year-old woman was diagnosed with pulmonary stenosis and right ventricular hypoplasia, complete occlusion of the left pulmonary artery secondary to a Blalock-Taussig shunt, and atrial septal defect. The patient developed thromboembolism of the subsegmental branches of the right pulmonary artery resulting in critical hemodynamic deterioration 2 weeks after undergoing one and a half ventricle repair (bidirectional Glenn shunt). The patient was treated with tissue plasminogen activator administered directly into the right pulmonary artery via an intravascular catheter. Progressive recanalization of the obstruction began immediately. Pulmonary angiography 3 months after thrombolytic therapy demonstrated patent subsegmental vessels. Early detection of the pulmonary thromboembolism and prompt intervention are crutial to relieving this fatal complication after a Fontan operation. PMID- 11257774 TI - Surgical management of multidrug-resistant tuberculosis. AB - We report surgical resections in 3 patients with active multidrug-resistant tuberculosis. All cases involved strains of Mycobacterium tuberculosis resistant to at least isoniazid and rifampin and patients who were poor candidates for medical therapy alone. We conducted pulmonary resections (partial resection in case 1, lobectomy in case 2, and segmentectomy in case 3). The optimum multiple drug regimen, based on drug susceptibility studies, was used preoperatively and postoperatively. In all cases, sputum smears and cultures yielded negative results postoperatively, and continue to be negative for Mycobacterium tuberculosis to date. It is recommended that, if localized disease is present and medical treatment is likely to fail, pulmonary resection be conducted for multidrug-resistant Mycobacterium tuberculosis. PMID- 11257775 TI - Extrapleural pneumonectomy for diffuse malignant pleural mesothelioma. A treatment option in selected cases? AB - OBJECTIVE: There is no standard treatment for diffuse malignant pleural mesothelioma. Sporadic extrapleural pneumonectomy for the disease in off-protocol set has been performed when the disease was judged as being confined to a hemithorax. SUBJECT: To evaluate the role of extrapleural pneumonectomy in the treatment for the disease, cases that underwent surgery in a single institution are reported with descriptions of the stages based on the system from the International Mesothelioma Interest Group. METHODS: Patients with diffuse malignant pleural mesothelioma treated with extrapleural pneumonectomy from 1989 to 1995 at the National Cancer Center Hospital East were reviewed. RESULTS: Five patients underwent extrapleural pneumonectomy. All were male. Their ages ranged from 48 to 68 years. The disease was right-sided in two and left-sided in three. The histologic subtype was epithelial in three, and mixed epithelial and sarcomatous in the other two. Post-surgical pathologic stage was Ib (T1bN0M0) in three, III (T3N1M0) in one, and IV (T4N2M0) in one. There was no treatment related death. Two cases with epithelial histology were alive with local recurrence on the chest wall at 75 months and at 51 months after surgery, respectively. A case with mixed epithelial and sarcomatous histology died of an esophageal cancer at 25 months after surgery. The two other cases died of the progressed disease at ten months and at four months after surgery, respectively. CONCLUSION: Extrapleural pneumonectomy for diffuse malignant pleural mesothelioma was concluded to be a treatment option in stage Ib cases with epithelial histology. To evaluate precisely the efficacy of surgery for the disease, a randomized control trial must be organized multi-institutionally. PMID- 11257776 TI - Alternate venous drainage and return of warmed blood combined with continuous hypothermic visceral perfusion. A new adjunct of thoracoabdominal aortic aneurysm repair. AB - OBJECTIVE: As a new adjunct to surgical repair for a thoracoabdominal aortic aneurysm, we have devised and used visceral perfusion in combination with alternate venous drainage and return of warmed blood in 4 patients. METHODS: Surgical repair of a thoracoabdominal aortic aneurysm of Crawford type III (n = 1) or type IV (n = 3) was performed. During visceral branch reconstruction, hypothermic blood (30-32 degrees C) was perfused continuously to each visceral and renal artery at a total flow rate of 300 ml/min, in combination with alternate venous drainage and return of warmed blood to the inferior vena cava. RESULTS: The visceral and renal perfusion time was 115 +/- 55 (with a range from 52 to 190) minutes. All 4 patients recovered uneventfully. CONCLUSIONS: Alternate venous drainage and return of warmed blood combined with continuous hypothermic visceral perfusion were a useful adjunct to thoracoabdominal aneurysm repair during reconstruction of visceral and renal arteries. PMID- 11257777 TI - Cardiac surgery in patients with end-stage renal disease. Utility of continuous ambulatory peritoneal dialysis. AB - OBJECTIVES: The number of patients with end-stage renal disease undergoing open heart surgery continues to grow. We evaluated continuous ambulatory peritoneal dialysis and the extracorporeal ultrafiltration method during cardiopulmonary bypass in the management of these difficult patients. METHODS: These 2 methods were used in 4 patients with renal failure who underwent open heart surgery between July 1997 and March 1999. Preoperative continuous ambulatory peritoneal dialysis was conducted using standard protocols. Extracorporeal ultrafiltration method was used only during cardiopulmonary bypass. Continuous ambulatory peritoneal dialysis was initiated upon arrival at the intensive care unit. Mean follow-up was 12 months. RESULTS: Postoperative blood urea nitrogen and creatinine concentrations were lower than preoperative concentrations. No patients required hemodialysis. All 4 patients were discharged to their homes. No deaths occurred. CONCLUSIONS: Continuous ambulatory peritoneal dialysis and extracorporeal ultrafiltration method are combined to treat patients with end stage renal disease who require open heart surgery. This combination is simple, and does not require specialized personnel, and obviates the hemodynamic instability associated with hemodialysis. PMID- 11257778 TI - [Resistance to Fas-mediated apoptosis in human papillomavirus type 16 immortalized human laryngeal epithelial cells after tumorigenesis]. AB - Apoptosis plays an important role in such biological processes as multistep carcinogenesis. Human papillomavirus type 16(HPV16)-immortalized human epithelial cell lines are, for the most part, nontumorigenic in nude mice and useful for studying mechanisms involved in multistep carcinogenesis. We previously reported that HPV16-immortalized human laryngeal epithelial cell line HLEC16 formed tumors after treatment with 4-(methyl-nitrosamine)-1-(3-pyridyl)-1-butanone, and derived one tumorigenic cell line, HLEC16T. We compared sensitivity to Fas receptor mediated apoptosis of nontumorigenic HLEC16 and tumorigenic HLEC16T. HLEC16 and HLEC16T expressed Fas protein (Fas) but not Fas-ligand (FasL) mRNA. We applied an anti-Fas monoclonal, antibody, CH11, to HLEC16 and HLEC16T, and monitored cell death. HLEC16T was found to be significantly less sensitive to CH11-mediated cell death that HLEC16. Western blot analysis showed no significant difference in levels of apoptosis-inducing protein, Bax, between HLEC16 and HLEC16T. Levels of apoptosis-inhibiting proteins Bcl-2 and Bcl-XL increased in HLEC16T. These results suggest that the inhibition of Fas-mediated apoptosis through apoptosis inhibiting protein overexpression may promote tumorigenicity in HLEC16T. PMID- 11257779 TI - [Clinical and bacteriological significance of the Streptococcus milleri group in deep neck abscesses]. AB - Streptococcus constellatus, S. intermedius, and S. anginosus, the three species of the S. milleri group, form part of the normal flora most commonly found in the mouth, throat, gastrointenstinal tract, and genital tract. The S. milleri group has become known as an important pathogen in abscess disease, but little attention has been paid to their role in deep neck abscesses. We have treated 9 patients with deep neck abscesses relating to the S. milleri group since 1991, and regarded this group as an important pathogen also in these abscesses. We studied the frequency of the S. milleri group isolated from deep neck abscesses in our cases and from the literature and discuss clinical significance and bacteriological pathogenesis. Cases numbered 27 treated at our facility since 1991 and 200 cases reported in the Japanese literature since 1990. Of our 9 cases, 4 originated from acute pharyngitis, 3 from peritonsillar abscesses, and 2 from odontogenic infection. Serious complications such as mediastinitis, cervical necrotizing fasciitis, sepsis accompanied by disseminated intravascular coagulation, and spondylitis of the cervical vertebrae were seen in 4 cases. Among organisms isolated, the S. milleri group appeared to be a pathogen contributing to abscess formation and to serious complications. The genus Streptococcus was most frequently isolated both in our 27 cases (66.7%) and the 200 in the literature (45.5%). Among species of the genus Streptococcus, the S. milleri group numbered the highest in our cases at 33.3% but only 8.5% in the literature. Cases in the literature, however, contained many unknown species of Streptococci--31.5% vs. 18.5% in our cases. alpha-streptococcus was frequently reported in the literature among unknown species of Streptococci--36 of 63. Culture-negative cases were also numbered more in the literature than in our case -29.0% vs. 18.5%. Special conditions and procedures are required to suitably isolate and detect the S. milleri group. Since not all facilities use identical techniques in routine bacteriological examination, a considerable number of the S. milleri group could be missed in unknown species of Streptococci or alpha streptococcus and culture-negative cases. The detailed pathogenesis of the S. milleri group remains to be clarified. Infection by normal flora on mucosa is thought to occur due to an imbalance between organisms and host defense in deep neck abscesses. Some strains of the S. milleri group have been reported to produce many tissue-destroying enzymes such as collagenase and hyaluronidase. The co-existence of the S. milleri group with some anaerobe strains has also been suggested to accelerate inflammation. We discuss the mechanism inducing the massive release of cytokines through T cell response to certain exotoxins produced by S. milleri group, as reported in toxic shock-like syndrome due to the group A beta-streptococcus and in alpha-streptococcal shock syndrome due to viridans streptococci (alpha-streptococci). PMID- 11257780 TI - [Clinical study on prognostic factors in thyroid carcinoma]. AB - We treated 227 patients, 45 men and 182 women, with thyroid carcinoma at our hospital from 1984 to 1998. Of these, 177 had papillary carcinoma and 50 follicular carcinoma. The extent of resection was based on tumor size in papillary carcinoma but not follicular carcinoma, and 70% of carcinoma patients underwent hemithyroidectomy. Neck lymph nodes were resected in 93.2% of papillary carcinoma patients, with D1 neck dissection in 45.7% and D2 or D3 neck dissection in 47.5%. In contrast, 70% of follicular carcinoma patients with lymph node resection had D1 dissection. Locoregional recurrence was noted in 22 patients and distant metastasis in 6 cases. Nonsurvivors numbered 17, 12 papillary and 5 follicular carcinoma patients, died of their primary disease. Almost all deaths were in patients with advanced disease, pT3 in 3, pT4 in 10, N1a in 3 and N1b in 8. The prognostic factors for papillary carcinoma were extracapsular spread, age, and distant metastasis, while the only factor for follicular carcinoma was distant metastasis. The 5-year survival for patients with papillary carcinoma was 93.0% and 10-year survival 88.8%, compared to 5-year survival for 93.5% of follicular carcinoma patients and 10-year survival for 93.5%. PMID- 11257781 TI - [Anthropometric study on normal human auricle in Japan]. AB - OBJECTIVE: Japanese people are physically larger and live longer than in the past. In light of these changes, we studied anthropometric auricular morphology in Japanese people in all age groups. METHODS: Subjects were 1,958 healthy Japanese people with no ear disease, 966 males aged 0-94 years and 992 females aged 0-99 years. They were classified at 5-year intervals into 18 age groups, and each group consisted of 50-72 persons. Bilateral size of auricles (ear length, ear width, length of ear attachment, auricular cartilage length, and auricular lobe length) were measured in the usual manner. All measurements were made with calipers by a single observer. RESULTS: Larger values were obtained in males than in females in almost all age groups. Rapid growth was observed until late teenage, and significant growth continued thereafter until advanced age. Auricular size was found to be greater than that in Japanese people in the past. CONCLUSIONS: In addition to changes in auricular size believed attributable to growth until late teenage, age-associated changes appear to continue during adulthood. PMID- 11257783 TI - [A case of anti-GQ1b-positive atypical Fisher syndrome with internal ophthalmoplegia but without external ophthalmoplegia]. AB - We report a 21-year-old man who developed an atypical form of Fisher syndrome. One week after having a common cold, he was admitted to our hospital because of a gait disturbance. Neurological examination revealed a somnolent state, cerebellar ataxia, areflexia, limb muscle weakness, and numbness in a glove and stocking like distribution. The patient had internal ophthalmoplegia but did not have external ophthalmoplegia. Brain MRI showed no abnormality in the orbital and the pretegmental brain regions. The protein level in the cerebrospinal fluid was 57 mg/dl and the cell count was 5 mononuclear cells/mm3. His serum titer of anti GQ1b IgG antibody was markedly elevated. There have been only two previous reports of isolated internal ophthalmoplegia with elevated anti-GQ1b antibody. The present case suggests that anti-GQ1b antibody play an important role in the pathogenesis of patients who present with internal ophthalmoplegia but without external ophthalmoplegia. PMID- 11257782 TI - [Analysis of ischemic stroke in patients aged up to 50 years]. AB - BACKGROUND AND PURPOSE: Ischemic stroke in young adults has not fully been studied in Japan. The purpose of this study is to clarify the clinical features and pathogenetic mechanisms of ischemic stroke in young adults. METHODS: From January 1990 to June 2000, 133 (7.1%) of 1,862 consecutive patients with ischemic stroke were aged 16 to 50 years (92 men and 41 women, aged of 42.6 +/- 7.7 years). We divided the patients into three groups according to the age of stroke onset; 16 to 40 (group A, n = 38), 41 to 45 (group B, n = 33), and 46 to 50 years (group C, n = 62). Sex, vascular risk factors including hypertension, diabetes mellitus, hyperlipidemia and smoking, cerebrovascular lesions, potential cardiac sources of emboli (emboligenic cardiac diseases), and clinical categories of stroke were examined. We compared them among the three groups. RESULTS: Men predominated in the all groups. Patients in the group C had more frequently hypertension (p < .0005) and diabetes mellitus (p < .05) than those in the group A and B. The vertebrobasilar infarcts were often revealed in the group A but not in the group B and C (p < .01). No significant difference was observed about the incidence of emboligenic cardiac diseases among the three groups. Out of 46 emboligenic cardiac diseases, the most frequent one was patent foramen ovale (15 patients), whereas nonvalvular atrial fibrillation was documented only in five. Fourteen of 20 patients with significant atherosclerotic arterial lesions were found in the group C, whereas 13 of 16 patients with nonatherosclerotic vasculopathies in the group A. Ulcerated atheroma in the aortic arch was detected in two patients, both in the group C. Overall, clinical categories of stroke were identified as lacunar (Lac) in 34 patients (26%), atherothrombotic (AT) in 16 (12%), cardioembolic (CE) in 44 (33%), miscellaneous (Misc) in 27 (20%), and undetermined in 12 (9%). The most frequent cause of Misc (nine patients) was arterial dissection, occurring mostly in the vertebrobasilar system (eight of nine patients). Misc was most frequent in the group A (p < .0001), and Lac was so in the group C (p < .001). Seventy-six percent of patients in the group A and 61% of those in the group B were attributed to nonatherosclerotic causes. In contrast, 61% of patients in the group C had atherosclerotic causes. CONCLUSIONS: Incidence of athero- and arteriosclerosis as causes of ischemic stroke was apparently increased beyond 45 years of age. Patients < or = 45 years are likely to have various and unique etiologies for ischemic stroke. A specific diagnostic approach is strongly recommended for young patients with ischemic stroke. PMID- 11257784 TI - [A case of agnosia for streets and houses unaccompanied by prosopagnosia of familiar faces due to the right occipital lobe infarction]. AB - We reported a patient who showed agnosia for streets and homes unaccompanied by prosopagnosia of familiar faces following infarction in the right occipital lobe. A 70-years-old right-handed man admitted to our department because of sudden development of visual impairment. He had left hemianopsia, left unilateral spatial neglect and slight visual memory disturbance but no other neurological abnormalities. His verbal memory was maintained intact. He was, however, unable to distinguish the sceneries of buildings and streets regardless of their familiarities and often got lost in the hospital. His topographical abilities in map-sketching and route-description were partially impaired depending upon the familiarity of targets; the disability was observed toward unfamiliar targets. His ability in facial recognition was also partially impaired depending upon the familiarity of persons. The patient was unable to distinguish faces of unfamiliar persons, such as nurses and doctors whom he met following stroke, whereas he was able to distinguish faces of familiar persons, such as his family and friends. The brain MRI demonstrated infarction in the right medial occipital lobe including parahippocampal gyrus, lingual gyrus and fusiform gyrus. In general, the manifestation of agnosia for streets and houses is associated with prosopagnosia. The present case, however, exhibited only the former in association with the partial manifestation of the latter. The fact suggests that those two are independent syndromes. The present case also showed a dissociation in the abilities of topographical and facial recognition according to the familiarity of targets. The process for the retrieval and reference of acquired information and that for the acquirement of new visual information may work in an independent manner. PMID- 11257785 TI - [X-Linked Charcot-Marie-Tooth disease with a new mutation (Thr191Ala) in the connexin32]. AB - We report a 59-year-old man with X-linked Charcot-Marie-Tooth (CMT) disease and a new point mutation in the connexin32 gene. The patient first noticed mild gait disturbance five years previously. On admission, he exhibited muscle atrophy and weakness in the distal part of both legs, mild muscle atrophy of both hands without weakness, and a minimal reduction of touch sensation in the right dorsal foot. Nerve conduction velocity of the peripheral nerves was diffusely reduced. Electromyography exhibited high-amplitude, long-duration, polyphasic motor unit potentials in the muscles of the extremities. Fibrillation potential and positive sharp wave were present in the affected muscles. Cerebrospinal fluid protein was slightly elevated. The polyneuropathy did not respond to high-dose corticosteroid treatment, and showed very slow progression. His parents were not consanguineous. His father and two sons were healthy, but similar illness (more severe) was suspected in his younger brother. Gene analysis (Southern hybridization) did not reveal any duplication or deletion in the CMT 1 A-REP region. However, a novel mutation (Thr191Ala) was detected in the connexin32. Although more than 160 mutations in the connexin 32 gene have been identified worldwide, approximately ten mutations have so far been reported in Japan. In comparison with X-linked CMT patients with other connexin32 mutations, the present case was characterized by late onset and mild neurological manifestation. Gene analysis provides a useful tool for diagnosing cases with slowly progressive, motor dominant polyneuropathy of unknown origin. PMID- 11257786 TI - [A case of sarcoid meningoencephalitis with an isolated supratentorial lesion]. AB - A rare case of sarcoid meningoencephalitis with no systemic lesion is reported here. A 58-year old man was admitted experiencing dull headache and speech disturbance. He had never received a diagnosis of systemic sarcoidosis. On admission, neurological examination revealed dysarthria, a defect of the right side visual field and accelerated right Achilles tendon reflex. A T2-weighted MRI showed a high-intensity signal in the white matter of the left parieto-occipital lobe surrounded by severe brain edema with a mass effect. The meninges around the lesion were enhanced by gadolinium, but no enhancement was observed in the basal portion. Angiotensin-converting enzyme (ACE) activities of cerebrospinal fluid (CSF) and serum were within normal range. The level of interleukin-6 in the CSF was slightly elevated. Chest X ray films and chest CT revealed no abnormal lesions. Whole body gallium scanning showed a hot region only in the intracranial lesion. A brain biopsy was performed. Histological examination revealed typical granuloma of sarcoidosis accompanied by microvasculitis and epithelioid cell granuloma without caseous necrosis. Oral administration of prednisolone improved all symptoms and MRI findings. These observations suggest that release of cytokines from macrophages and epithelioid cells, as well as disruption of the blood-brain barrier due to microvasculitis, are involved in the mechanism responsible for producing lesions of sarcoid meningoencephalitis. PMID- 11257787 TI - [A case of amyotrophic lateral sclerosis presenting with circulatory collapse during artificial respiration]. AB - A 71-year-old man developed dysarthria and difficulty of swallowing in December 1997. He was diagnosed as having the bulbar type of amyotrophic lateral sclerosis (ALS). In November 1998, he was admitted to our hospital to undergo treatment for bulbar palsy and respiratory discomfort. In January 1999, ventilatory support (synchronous intermittent mandatory ventilation) during sleep at night was initiated. Severe progressive hypotension and loss of consciousness were observed soon after the start of artificial respiration, and both symptoms disappeared after artificial respiration was discontinued. This phenomenon was observed consistently during ventilatory support, while unpleasant stimuli such as bronchoscopy and replacement of the cannula tube induced severe hypertension. To clarify the mechanism of underlying these abnormal changes in blood pressure, autonomic function tests were performed while awake during the daytime. Ventilatory support induced a drop in blood pressure accompanied by a decrease in influx speed to the right ventriculum, the latter of which suggested a reduction in venous return. These values returned to the baseline following detachment of the ventilator. A 60 degrees head-up tilt (HUT) angle and standing from a supine position produced orthostatic hypotension, the latter of which was accompanied by a compensatory increase in pulse rate. The basal supine plasma noradrenaline (NA) level was high and the HUT showed a slight elevation of NA. The basal supine plasma arginine vasopressin (AVP) level was within the normal range, whereas the AVP level did not increase during HUT. Urinary secretion rates of NA and 3 methoxy-4-hydroxy-phenylglycol were elevated. A cold pressor test demonstrated reflex hypertension. The oculovagal reflex, coefficient of variation of R-R intervals. (CVR-R) and increase in pulse rate in response to atropine administration were within the normal range. The combination of midodrine, L dihydroxyphenylserine (DOPS) and increasing intravascular volume via continuous intravenous drip infusion relieved the circulatory collapse during artificial respiration. In conclusion, the present case of ALS had sympathetic hyperactivity, somatosympathetic reflex and dysregulation of the baroreflex arc. Degeneration of central autonomic network, including the hypothalamus and the central nucleus of the amygdala, which has been shown in some ALS patients, might underlie the autonomic abnormalities in this patient. PMID- 11257788 TI - [A family with oculopharyngeal muscular dystrophy with (GCG)9 expansion in which a sister had neck as well as proximal and her brother proximal lower limb muscle weakness]. AB - We report a 58-year-old woman (patient 1) and her 60-year-old brother (patient 2) with autosomal dominant oculopharyngeal muscular dystrophy. Patient 1 first noticed blepharoptosis and neck weakness at age 55. On neurological examination, she showed bilateral blepharoptosis and weakness in the neck and upper proximal limbs. Serum creatine kinase (CK) level was slightly elevated. Her older brother first noticed blepharoptosis and lower limb weakness at age 51. On neurological examination, he showed bilateral blepharoptosis, slight ophthalmoparesis and bilateral iliopsoas muscle weakness. Serum CK level was normal. Esophageal fluoroscopy disclosed dysfunction of the constrictor pharyngeal muscles. Muscle biopsy of them showed myopathic changes with rimmed vacuoles. The (GCG)9 mutation in the poly (A) binding protein 2 gene was identified, which was the same as seen in the large French-Canadian kindred in Quebec in Canada. The clinical phenotype in patient 2 is similar to that of French-Canadian patients but it in patient 1 is different in distribution of muscle weakness. PMID- 11257789 TI - [Muscle involvement of Stormorken's syndrome]. AB - We described two patients, a mother and daughter, of Stormorken's syndrome. The syndrome is characterized clinically by autosomal dominant inheritance, congenital miosis, thrombocytopenia, asplenia and muscle weakness. Both patients had bleeding tendency, ichthyosis of arms, and muscle weakness. The daughter additionally had short stature (146 cm), low body weight (32 kg) and muscle cramp. Neurological findings of the patients included migraine-like headache, cognitive dysfunction, limitation of upward and lateral gaze, and amydriasis. Femoral muscle MRI of the daughter demonstrated decreased volume with patchy high intensity areas in the hamstrings. A muscle biopsy from the daughter showed myogenic changes with muscle fiber necrosis and regeneration, variation in fiber size, tubular aggregates in approximately 5% of fibers, and fibrous tissue proliferation. Dystrophin, dystrophin-associated proteins and dysferlin were normally expressed. Although both patients had elevated creatine kinase levels and generalized muscle wasting, muscle weakness was mild with slow progression. A certain membrane defect in the platelet and muscle fiber might be responsible for the pathogenesis of this syndrome. PMID- 11257790 TI - [A case of neuro-Behcet's disease with HLA B54 and predominant cerebral white matter lesions]. AB - We report a 55-year-old woman with neuro-Behcet's disease with HLA B54 and predominant cerebral white matter lesions. She showed a cryptogenic high fever and cerebral cortical symptoms such as perseveration, limbkinetic apraxia and dementia. CSF study showed an increase of cell count and protein and a decrease of sugar. MRI showed diffuse T2-high signal intensity mainly in the subcortical white matter of left parieto-occipital lobes and basal ganglia. Her clinical signs greatly improved after administration of prednisolone. Her HLA type was not B51 but B54. Though our patient did not completely satisfy clinical criteria for neither neuro-Behcet's disease nor Sweet's syndrome, she showed partial features of both Behcet's disease and Sweet's syndrome. PMID- 11257791 TI - [A case of Gerstmann-Straussler-Scheinker syndrome (P102L) accompanied by optic atrophy]. AB - We report a patient with Gerstmann-Straussler-Scheinker syndrome (GSS102) who developed optic atrophy. He had been complaining of slowly progressive postural unsteadiness and pain in both legs for 3 years. Visual acuity subacutely worsened in the last half year. His father and two aunts, who already died, had been diagnosed to have dementia. It is uncertain whether they had optic atrophy or not. He was alert but apathetic. Neurological examination revealed cerebellar ataxia, painful dysesthesia and loss of deep tendon reflexes in the lower limbs. Fundoscopic examination revealed bilateral optic atrophy without retinal degeneration, which has never been reported in GSS. A brain MRI showed mild atrophy of cerebellar hemispheres without signal abnormalities of optic nerves. DNA analysis of prion gene revealed point mutation at codon 102 (P102L), which was relatively common mutation in GSS. Other mutations were not found. Only two patients of Creutzfeldt-Jakob disease with optic atrophy have been reported. This case seems to be important to investigate why optic tracts are generally spared in prion disease. PMID- 11257792 TI - [Japanese encephalitis presenting with left hemiplegia and thalamic neglect--a case report]. AB - This report concerns a 51-year-old right-handed man with Japanese encephalitis, showing left hemiplegia and left hemispatial neglect. On admission, he had a slight fever, mild consciousness disturbance, left hemiplegia, and left hemispatial neglect but no neck stiffness, headache nor nausea. He was treated on the basis of cerebral infarction, but his fever and consciousness disturbance worsened. We found pleocytosis (145/mm3) in the cerebrospinal fluid (CSF) and right thalamic edema on a brain CT scan obtained 4 days later. He was finally diagnosed as having Japanese encephalitis on the basis of an increase in anti viral antibodies observed in paired CSF and serum samples. In the exacerbation phase, 123I-IMP single photon emission CT (SPECT) demonstrated a marked decrease in cerebral perfusion in the right hemisphere, while a brain MRI revealed irregular lesions localized the right thalamus (mainly posterior and medial parts), showing low intensity on T1-weighted and high intensity on T2-weighted images. In the recovery phase, asymmetrical perfusion was no longer observed on SPECT and the symptoms including the left hemispatial neglect had improved. These findings suggest that the left hemispatial neglect in this patient might been caused by the right thalamic lesion resulting in damage to the activating system of the right hemisphere. This case thus shows that acute onset of hemispatial neglect could be caused by cerebral encephalitis. PMID- 11257793 TI - [Overloading to neck extensor muscles is an aggravating factor to induce further neck drop in isolated neck extensor myopathy (Katz). A case report]. AB - A 78-year-old woman was hospitalized because of progressive anterior neck drop over 4 months prior to admission. She was normal except for mild weakness of her neck, trapezius and biceps brachii muscles. EMG revealed mild myopathic changes in the neck extensors, trapezius, deltoid and sternocleidomastoid muscles. Bilateral splenius capitis muscles had high intensities on T2-weighted and STIR pulse-sequenced MRI. However, there were no inflammatory changes in the right splenius muscle biopsy. Accordingly, the abnormal MRI finding seems not to result from an inflammatory process but from an physiological increase of intracellular water content due to sustained muscle contraction. Because apparent neuromuscular diseases responsible for neck drop were excluded, her clinical features met the criteria of isolated neck extensor myopathy (INEM, Katz). After strict bed-rest for one month, her neck drop improved dramatically. When she returned to the previous life style after discharge, her symptoms of the neck drop reappeared. Although the cause of INEM remains unclear, the present case indicates that the condition is reversible at least in the early stage of the disease, and the overloading to the neck extensor muscles is an aggravating factor of the neck drop in INEM. PMID- 11257794 TI - [A case of chronic hypertrophic cranial pachymeningitis causing cranial polyneuropathy and unilateral central retinal vein occlusion]. AB - We presented a 70-year-old woman who developed unilateral visual loss due to central retinal vein occlusion caused by chronic hypertrophic cranial pachymeningitis. She had developed right blindness with optic atrophy due to chronic intracranial pachymeningitis one year before admission. In June 1999, she noticed visual loss of the left eye. On admission, neurological examination revealed left visual loss and sensory impairment in the first branch area of bilateral trigeminal nerves. Ophthalmological examination revealed central retinal vein occlusion of the left eye. Brain MRI showed dural thickening with gadolinium enhancement of the cavernous sinus near the left optic nerve and dilatation of the left supraorbital vein. There was no compression of the left optic nerve. We suggest that central retinal vein occlusion may be caused by compression of the supraorbital vein by dural thickening. This is the first case report of central retinal vein occlusion associated with chronic hypertrophic cranial pachymeningitis. PMID- 11257796 TI - A measure of satisfaction for the assisted-living industry. AB - A self-administered satisfaction survey instrument for the assisted-living industry was developed and validated. The survey contains 45 Likert-type items that measure residents' and family members' satisfaction with the most central aspects of housing and care. The scale covers six key service dimensions: activities, personnel, dining, apartment, facility, and management. Internal consistency tests indicate high reliability. Multiple tests of validity confirm the scale's effectiveness in measuring residents' and family members' satisfaction with the six dimensions. Overall, residents are less satisfied with assisted-living programs than their family members are, but they may feel inhibited about expressing criticism in the presence of family. A priority index highlights service areas in which performance improvement efforts should be made to obtain the greatest increases in satisfaction while making the most efficient use of limited resources. Managerial implications of the tool are discussed. PMID- 11257795 TI - Redesigning clinical office practices to improve performance levels in an individual practice association model HMO. AB - Independent Health, a large health maintenance organization in western New York, is participating in a national initiative to redesign the way healthcare is delivered in the clinical office setting. The Idealized Design of Clinical Office Practices (IDCOP) project aims to initiate profound change in the ways physicians deliver care and treatment in the office; the focus is on demonstrating improvements in access to care, patient care, patient and staff satisfaction, and financial performance. Two clinical prototype test sites have successfully implemented an "open access policy," which enables most patients to see their physician on the day an appointment is requested. The two test sites also found improved clinical indicators (for diabetes and coronary heart disease) that positively affected the health of patients; reduced waiting time, enriching the overall experience of the office visit; and improved patient and staff satisfaction with the operational and care delivery modifications. This article describes the findings of the IDCOP initiative at Independent Health. PMID- 11257797 TI - Utility of UB-92 data for monitoring emergency department performance improvement. AB - The Universal Billing Code of 1992 (UB-92) is a standard database used by hospitals to generate itemized charges for patient visits. This study examined the use of UB-92 information to monitor emergency department performance improvement projects. UB-92 data were used to determine whether urine tests had been ordered for emergency department patients. A population of patients at low risk for requiring a urine culture was defined as discharged female patients between 16 and 60 years of age undergoing a urinalysis as part of their emergency department treatment during a 10-month period. Based on UB-92 data, only a total of 2,138 patients were identified who met the study's low-risk criteria. Recommendations for the optimum use of these tests were presented to the emergency physicians as part of departmental performance improvement activities. Additional logistical problems associated with the procedure for ordering this test were identified and corrected as part of this project. After an additional 5 month period, a second analysis of the entire 15 months of UB-92 data was performed. Prior to physician notification, 41.6% of low-risk patients underwent both a urinalysis and a urine culture and sensitivity in the emergency department. In the 5-month follow-up period, the percentage of patients undergoing both tests decreased 18% to only 23%. PMID- 11257798 TI - Improved cholesterol management in coronary heart disease patients enrolled in an HMO. AB - The purpose of the study was to describe the effect of physician reminders on the measurement of low-density lipoprotein cholesterol (LDL-C) levels and treatment to achieve an LDL-C goal of < or = 100 mg/dL in coronary heart disease (CHD) patients. After reminders were initiated, the number of CHD patients without a documented LDL-C was reduced from 30% to 18%, between January 1997 and July 1998, and the percentage of CHD patients achieving the LDL-C goal improved from 10% to 27%. Thus, reminders can be an effective tool in improving cholesterol management of CHD patients. In contrast, a cholesterol-lowering clinic made available to some physicians, in addition to the reminders, was rarely used. PMID- 11257799 TI - A quality measurement framework for managed care organizations. AB - This article presents a framework for a quality measurement system and its corresponding principles for quality improvement within a managed care environment. The quality measurement system comprises four modes: quality assessment, comparison, improvement, and system evaluation. If these modes are to be effective, the measurement system must incorporate the following five principles: (a) maintain and continually improve the data systems, (b) develop and utilize sound methodology and valid indicators, (c) build a competent measurement team, (d) develop effective strategies for implementation, and (e) protect member confidentiality. This model is helpful in the ongoing development and testing of new measurement systems. PMID- 11257800 TI - Initiating a pediatric office-based quality improvement program. AB - This article describes a pediatric primary care network's initiative to implement an office-based quality improvement (QI) program. First, network physicians determined their priorities for quality improvement. Then primary care practitioners on the Quality Committee (QC), representing each local group practice, were educated about a physician-led QI process. The four steps of this process are as follows: (1) Set quality aims, (2) determine measurements of improvements, (3) generate ideas for change, and (4) test changes. Next, the QC selected two initial projects--office preparedness for pediatric emergencies and asthma management--and corresponding subcommittees were formed. The pediatricians identified QI practice changes they believed could be implemented successfully to make a measurable difference in children's healthcare. PMID- 11257802 TI - Inhibition of platelet aggregation of a mutant proinsulin molecule engineered by introduction of a native Arg-Gly-Asp sequence. AB - A 13 amino acid sequence, CRVARGDWNDNYC, originated from disintegrin eristostatin, was introduced into an inactive human proinsulin molecule between the B29 and A2 sites to replace proinsulin C-peptide by molecular cloning techniques. The constructed Arg-Gly-Asp (RGD)-proinsulin gene was cloned into a temperature-inducible vector pBV220 and expressed in Escherichia coli. The expressed RGD-proinsulin was refolded and purified by Sephadex G50 and DEAE Sephadex A25 separations. The chemical identity was confirmed by both amino acid composition and mass spectrometry analyses. This RGD-proinsulin showed an inhibitory activity of adenosine 5'-diphosphate-induced human platelet aggregation with an IC50 value of 200 nM. Its insulin receptor binding activity remained as low as 0.03% with native insulin as a control, and its insulin immune activity retained 27.6% compared with proinsulin. PMID- 11257804 TI - Spectroscopic and electrochemical investigation of ternary complexes of D- or L aspartic acid containing polyacrylamides-Cu(2+)-bovine serum albumin and their radio-stability. AB - D- or L-aspartic acid containing polyacrylamides were synthesized. Binary and ternary complex formation between these polymers with copper and bovine serum albumin was studied by spectroscopic and electrochemical measurement. Depending on the ratio of the polymer/copper and protein/polymer, the mixture exhibited water-soluble and insoluble character. A hypothetical structural scheme for the formation of ternary complexes is proposed. The effect of radiation on these complexes was also investigated. PMID- 11257803 TI - Direct antigen capture by soluble scFv antibodies. A method for detection, characterization, and determination of affinity. AB - For ease of detection, soluble forms of phage-displayed scFv antibodies are usually expressed with a tag, e.g., c-myc or His (Histidine). The binding is then assayed by a monoclonal antibody to the tag. In this article, we describe the use of biotinylated antigen for detecting soluble scFv antibodies without utilizing the peptide tag detection system. The scFv antibodies were against the oncoplacental antigen heat-stable alkaline phosphatase (HSAP). The method essentially consisted of either reverse Western or antigen capture enzyme-linked immunosorbent assay (ELISA). In the reverse Western, periplasmic extract was electrophoresed, and binding to biotinylated antigen was detected by the detection system based on streptavidin-horseradish peroxidase. The antigen capture ELISA utilized the binding of periplasmic extract to a polystyrene plate. We have also demonstrated the use of antigen capture ELISA for studying specificity and affinity of the selected clones. Although these techniques have been developed for antibodies to HSAP, they have general utility for phage expression systems without a peptide tag. PMID- 11257805 TI - An improved method for extraction of beta-carotene from Blakeslea trispora. AB - An improved method for the extraction of beta-carotene from Blakeslea trispora is described. The fermentation broth was steamed at 121 degrees C for 15 min, and the liquid was centrifuged at 5000 g for 20 min. beta-Carotene was removed from the biomass by extraction with absolute ethanol at a ratio of 1:100 at 30 degrees C for 2 h in a rotary shaker incubator at 300 rpm. The carotenoid pigment was completely removed from the cells after three repeated extractions. The removal of beta-carotene from B. trispora was higher during the first stage (75%) whereas in the other stages it was very slow. PMID- 11257806 TI - Comparative studies on a mesophilic and a thermophilic alpha-amylase. AB - A comparative study was performed on thermal stability of mesophilic and thermophilic alpha-amylases, and the effects of various denaturing agents, organic solvents, and stabilizers were investigated. As expected, the thermophilic enzyme showed higher resistance toward denaturation in water as its natural medium, but such a difference could not be detected in nonaqueous environments. Furthermore, stability of these molecules was improved by including various stabilizing agents. Of the compounds tested, sorbitol provided the highest degree of protection, which was found to be owing to its effect on increasing Tm and its ability in totally preventing deamidation of amino acid residues in the protein molecules. PMID- 11257807 TI - GpdA-promoter-controlled production of glucose oxidase by recombinant Aspergillus niger using nonglucose carbon sources. AB - The gpdA-promoter-controlled exocellular production of glucose oxidase (GOD) by recombinant Aspergillus niger NRRL-3 (GOD3-18) during growth on glucose and nonglucose carbon sources was investigated. Screening of various carbon substrates in shake-flask cultures revealed that exocellular GOD activities were not only obtained on glucose but also during growth on mannose, fructose, and xylose. The performance of A. niger NRRL-3 (GOD3-18) using glucose, fructose, or xylose as carbon substrate was compared in more detail in bioreactor cultures. These studies revealed that gpdA-promoter-controlled GOD synthesis was strictly coupled to cell growth. The gpdA-promoter was most active during growth on glucose. However, the unfavorable rapid GOD-catalyzed transformation of glucose into gluconic acid, a carbon source not supporting further cell growth and GOD production, resulted in low biomass yields and, therefore, reduced the advantageous properties of glucose. The total (endo- and exocellular) specific GOD activities were lowest when growth occurred on fructose (only a third of the activity that was obtained on glucose), whereas utilization of xylose resulted in total specific GOD activities nearly as high as reached during growth on glucose. Also, the portion of GOD excreted into the culture fluid reached similar high levels (approximately equal to 90%) by using either glucose or xylose as substrate, whereas growth on fructose resulted in a more pelleted morphology with more than half the total GOD activity retained in the fungal biomass. Finally, growth on xylose resulted in the highest biomass yield and, consequently, the highest total volumetric GOD activity. These results show that xylose is the most favorable carbon substrate for gpdA-promoter-controlled production of exocellular GOD. PMID- 11257808 TI - Nonproteolytic cleavage of aspartyl proline bonds in the cellulosomal scaffoldin subunit from Clostridium thermocellum. AB - Previous work from our group [Morage (Morgenstern), E., Bayer, E. A., and Lamed, R. (1991), Appl. Biochem. Biotechnol. 30, 129-136] has demonstrated an anomalous electrophoretic mobility pattern for scaffoldin, the 210-kDa cellulosome integrating subunit of Clostridium thermocellum. Subsequent evidence [Morag, E., Bayer, E. A., and Lamed, R. (1992), Appl. Biochem. Biotechnol. 33, 205-217] indicated that the effect could be attributed to a nonproteolytic fragmentation of the subunit into a defined series of lower-molecular-weight bands. In the present work, a recombinant segment of the scaffoldin subunit was employed to determine the site(s) of bond breakage. An Asp-Pro sequence within the cohesin domain was identified to be the sensitive peptide bond. This sequence appears quite frequently in the large cellulosomal proteins, and the labile bond may be related to an as yet undescribed physiological role in the hydrolysis of cellulose by cellulosomes. PMID- 11257809 TI - Quaternized wood as sorbent for hexavalent chromium. AB - The potential of quaternized wood (QW) chips in removing hexavalent chromium from synthetic solution and chrome waste under both batch and continuous-flow conditions was investigated. Sorption was found to be dependent on pH, metal concentration, and temperature. QW chips provide higher sorption capacity and wider pH range compared with untreated wood chips. The equilibrium data could be fitted into the Langmuir isotherm model, and maximum sorption capacities were calculated to be 27.03 and 25.77 mg/g in synthetic chromate solution and chrome waste, respectively. The presence of sulfate in high concentration appeared to suppress the uptake of chromium by QW chips. Column studies showed that bed depth influenced the breakthrough time greatly whereas flow rate of influent had little effect on its sorption on the column. PMID- 11257810 TI - [Surgery for a recently symptomatic severe carotid stenosis, indicated or not? Clinical application of 'evidence-based' medicine]. AB - Two trials have shown that carotid surgery reduced the risk of stroke or death for patients with a recently symptomatic severe carotid stenosis. Nevertheless, it remains difficult to advise in individual cases. The risk of a stroke is about 20% in the next 3 years on medical treatment alone. This means that surgery is of no value in about 80% of the patients. With the help of prediction rules (developed by Rothwell and Warlow) it is possible to further individualize the decision whether or not to operate, by incorporating factors such as the angiographic roughness of the stenosis lining, the time since the cerebrovascular symptoms appeared, the extent of the stenosis, increased operation risk, sex, peripheral vascular disease and systolic blood pressure. If these prediction rules prove to be correct, doctors will be able to better advise their individual patients. PMID- 11257812 TI - [Development and the developmental disorders of human brain. I. Early development of the cerebrum]. AB - The recent discovery of many genes that regulate brain development is revolutionizing our knowledge of neuroembryology and, moreover, our understanding of how gene defects cause human birth defects. The first 8 weeks of the development of the cerebrum can be subdivided into 23 stages, with early development of mostly the spinal cord and the brain stem. Regionalization of the brain has been related to genes that play a part in it. A characteristic developmental disorder for this early phase in the development of the forebrain is holoprosencephaly, a brain patterning disorder. Numerous genes play a part in its occurrence; abnormal function of signal factors as well as of transcription factors may lead to holoprosencephaly. PMID- 11257811 TI - [Evaluation of the therapeutic effects in individual patients with Alzheimer disease]. AB - Rivastigmine was recently licensed for treatment of Alzheimer's disease on the basis of large double-blind placebo-controlled clinical trials. However, it is difficult to determine the clinical relevance for individual patients from the results of this type of clinical trial. With the help of standardised measurements in individual patients the clinical relevance can be better established. In order to avoid extra burden on patients, caregivers and doctors, these measurements should be simple. A combination of three clinimetrical scales for cognition, ability to perform daily activities and behaviour, which takes about 15 min to complete, appears to be efficient. Comparison of these data with data from a group of untreated Alzheimer's patients can give an impression of the efficacy of the medication. With the use of goal-attainment scaling, measurements can be individualised even more. This approach allows the clinician, in consultation with the caregiver and the patient, to make an informed decision about whether or not to continue treatment. PMID- 11257813 TI - [Optimizing antibiotics policy in the Netherlands. VI. SWAB advice: no selective decontamination of intensive care patients on mechanical ventilation]. AB - The Working Party on Antibiotic Policy (Dutch acronym is SWAB) has issued a guideline in which the pro and cons of the routine use of selective decontamination (SD) in patients in intensive care (IC) on mechanical ventilation are compared in order to decide whether SD is indicated. The effectiveness of SD in IC patients was evaluated in 28 prospective, randomized studies. In most studies a significant reduction in the incidence of pneumonia was demonstrated. The incidence of pneumonia in the control groups varied from 5 to 85%. The reduction in the incidence of pneumonia seems to have no effect on duration of mechanical ventilation and IC unit stay or the use of antibiotics. No effect on IC mortality was demonstrated. However, only major reductions could have been demonstrated with the size of the studies carried out so far. A significant reduction of about 20% was suggested in two meta-analyses. The validity of these meta-analyses is questionable. Based on the data available, it is not possible to reach the conclusion that SD will be cost-effective. The size of the studies is too small and the study duration too short to prove that the use of SD, if applied on a large scale, might not eventually lead to development of resistance. Selection of micro-organisms that are already intrinsically resistant or had already acquired resistance to one of the agents used, has been demonstrated. In the absence of clearly demonstrated advantages (decrease in mortality, reduction in the use of antibiotics, cost-effectiveness), the routine use of SD in IC patients on mechanical ventilation is not recommended. PMID- 11257814 TI - [Diagnostic image (26). Pneumopericardium due to leaking drainage system]. AB - A 55-year-old man with pleuropericarditis carcinomatosa was treated with pericardiocentesis and drainage. Because of leakage in the drainage system a pressure pneumopericardium developed. PMID- 11257815 TI - [From gene to disease; from Notch3 to cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy]. AB - Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy' (CADASIL) is an autosomal dominant inherited arteriopathy leading to brain infarcts and dementia at middle age with extensive cerebral white matter changes on MRI. CADASIL is caused by mutations in the Notch3 gene on chromosome 19. PMID- 11257817 TI - [Low rate of recurrence at a follow-up study of vaginal repair of enterocele]. AB - OBJECTIVE: Evaluation of short and longterm follow-up after vaginal repair of enterocele. DESIGN: Retrospective medical record investigation, questionnaire investigation and outpatient clinic follow-up examination. METHODS: In the Department of Gynaecology, Ikazia and Haven Hospital, Rotterdam, the Netherlands, 66 patients were treated because of enterocele by vaginal repair in 1989-1998. Follow-up results were gathered from medical records, a written questionnaire and recent gynaecological examination. RESULTS: The questionnaire response was 49/66 (74%); of the 17 non-respondents 6 had died. Of the 29 patients who had been routinely examined gynaecologically more than one year before, 25 consented to an additional gynaecological examination. The median follow-up until the questionnaire or the outpatient clinic examination was 3 years and 7 months (range: 1 month to 9 years and 7 months). A recurrent enterocele had been found in 4 patients; 3 of them underwent a repeat repair. A serious short-term complication was one rectovaginal fistula with a temporary artificial outlet. No clear relation of dysfunction of voiding and defaecation with the operation was found. However, problems with coitus were new in 19% of the sexually active group. CONCLUSION: The vaginal repair of enterocele showed good results with a low recurrence rate of 6% in a complex group of patients. PMID- 11257816 TI - [Great discrepancies between European, Dutch and Belgian criteria for the use of statins in the prevention of primary cardiovascular disease in family practice]. AB - OBJECTIVE: To determine the differences in prescribing advice for statines in primary cardiovascular prevention, applying different protocols, in a first-line setting. METHOD: In February-March 2000, at the general practice 'Medicine for the People' in Deurne-Antwerp, Belgium, all contacts with patients known with at least one cardiovascular risk factor and no signs of cardiovascular disease were included in the study. The absolute risk of developing cardiovascular disease in the next 10 years was calculated per patient according to the protocols of the 'European Society of Cardiology' (ESC) and the Dutch College of General Practitioners (NHG) and it was determined whether these protocols advised prescription of statines. It was also determined if the Belgian criteria for repayment of statines, developed by the Rijksinstituut voor Ziekte- en Invaliditeitsverzekering (RIZIV) were met. RESULTS: The study group comprised 143 patients with a mean age of 66 years, of which 51 (36%) were men. According to the RIZIV criteria 75 (52%) patients of these 143 were eligible for the repayment of statines. The NHG protocol advised to prescribe statines for 4 (3%) patients and the ESC protocol for 69 (48%) patients. Of the 75 patients who were considered for repayment, 34 (45%) according to the ESC protocol and 74 (99%) according to the NHG protocol did not need statines. Also, in the whole study population, 28 (20%) patients needed statines according to ESC and 3 (2%) patients according to NHG, but they could not get repayment for statines according to the RIZIV. The ESC protocol estimated the risk per patient on average 8.5% (95% confidence interval: 7.1-9.8; p < 0.0001) higher than the NHG protocol. CONCLUSION: The NHG protocol estimated the risk significantly and markedly lower than the European protocol, although they are both based on the same Framingham data. There also existed a weak concordance between both protocols. The RIZIV criteria were not 'evidence-based'. They incited to an irrational and wasting prescribing behaviour. There is a need for an integrated guideline for primary cardiovascular prevention and for the adjustment of the RIZIV criteria. PMID- 11257818 TI - [Hypercalcemia due to adult T-cell lymphoma in a man from Surinam]. AB - A 48-year-old male patient from Surinam presented with anorexia, nausea and weight loss. An extreme hypercalcaemia of 5.08 mmol/l was found. Further diagnostic investigations showed that this patient had a HTLV-1 positive adult T cell leukaemia/lymphoma (ATL/L). This is often associated with multilobularly nucleated lymphocytes, bone destruction and hypercalcaemia. Skin localisations are frequently observed. The combination of cytomorphology, immunophenotyping, HTLV-1 seropositivity and clinical findings points to the diagnosis. The patient was treated with 6 courses of chemotherapy consisting of cyclophosphamide, doxorubicin, teniposide, prednisone, vincristine and bleomycin. Upon inquiry it appeared that he had died elsewhere. Chemotherapy induces a short-lived remission in a minority of ATL/L patients. Antiviral therapy (alpha interferon, zidovudine) might offer new possibilities. PMID- 11257819 TI - [Waiting times in specialist medical care]. AB - Waiting times in specialist medical care are difficult to reduce owing to the fast-growing demand with supply lagging behind. These waiting times were the subject of a conference of this Journal, where experts from different backgrounds assessed the problems and discussed promising ways of coping with them at micro, meso and macro level. In the first category, a system developed in Leiden University Medical Centre was presented that provides insight into the expected waiting time per disease category, elucidates the bottlenecks in practice and supports the quality of care and the planning of patient flows. At the meso level, the discussion addressed how the differences within and between institutions and within and between regions may be reduced; this may be done, for instance, by better spread of the work load. This offers a better contribution to a structural solution than extra-regular initiatives. The conference finally discussed the importance of the current shift of important (control) tasks from the government to insurers. Those present expected that stimulation of regional initiatives of hospitals and health insurers by means of more money and latitude, allocated by the government and under its control (inspection), offers the best opportunities to shortening of the waiting lists and improvement of the quality of care. PMID- 11257820 TI - [Care for body materials obtained from autopsy]. AB - During every autopsy, small tissue samples are taken from each organ for microscopic examination. Organs or part of organs are retained for a longer period if (a) the organ concerned is so small that the entire organ is needed for the microscopic examination; (b) the organ exhibits complicated abnormalities which can only be diagnosed following fixation or a special treatment; (c) the organ must be fixed before the examination can take place, for example, the brain. This means that the corpse is not buried or cremated in its entirety. Next of kin are not always aware of this procedure even if they have given their consent for an autopsy. Changes within society, especially the handling of aborted foetal material and foetal material from intra-uterine deaths, require new agreements and rules for the retention of organs. Next of kin must be able to have confidence in the provision of information and the careful management of body material retained during autopsies. PMID- 11257822 TI - The effect of wall losses in the numerical simulation of microwave heating problems. AB - A study is made into the numerical modeling of wall losses for a microwave heating application. It makes use of a surface integral term for both a frequency and time domain finite edge element formulation in order to model the wall impedance of the enclosed microwave cavity. The paper describes how the surface element matrix of the complex wall impedance is combined with the matrix formulation. The results are checked against analytical expressions for a single mode resonant cavity. An analysis on the effect of lossy walls is provided using four low-loss material insertions over a range of surface conductivities. PMID- 11257821 TI - [AIDS: the future of world is in 'their' hands, but comes from 'our' pockets; an impression from the World Aids Conference in South Africa]. PMID- 11257823 TI - The influence of microwave heating on the characteristics of polyelectrolytes. AB - Results obtained by microwave (MW) and simultaneous microwave and electron beam (MW + EB) application in the chemistry of acrylamide and acrylic acid co-polymers (PA type polyelectrolytes) are presented. Comparative results concerning the molecular weight (Mw) and Huggins' constant for the acrylamide copolymers obtained by classical heating, MW heating, EB irradiation and simultaneous MW + EB treatment are reported. MW heating produces high PA water solubility but median Mw values; EB irradiation gives high Mw values but associated with a cross linked structure (poor water solubility) while MW energy addition to EB energy gives simultaneously high Mw values and high PA water solubility. A MW installation of 2.45 GHz and 2.5 kW, designed to provide small-scale commercial production of PA polyelectrolytes, is described. PMID- 11257824 TI - Modification of a household microwave oven for continuous temperature and weight measurements during drying of foods. AB - The main objective of this study was to modify a conventional household microwave oven for recording temperature and weight measurements continuously during microwave drying of foods. A household microwave oven with digital control was equipped with an electronic balance and a set of standard thermocouples that were connected to a PC for continuous data collection. The efficiency of the modified thermocouples was tested against the built-in probe of the oven. It was found that this microwave oven dryer set-up could be used for drying kinetics studies of foods since both the temperature and weight of the sample could be recorded continuously during the microwave drying operation. PMID- 11257825 TI - Crossed waveguide directional couplers for high power applications. AB - The small aperture coupling theory of the broad wall to broad wall (Moreno) coupler is reviewed and extended. It is shown that a lower limit on the directivity is given by the ratio of magnetic polarizability to electric polarizability for the coupling apertures. Hence directivity may be improved by using very narrow crossed slots for coupling. The results of coupling measurements on a Moreno coupler are presented to demonstrate the accuracy of the small aperture theory. A small aperture design procedure has also been developed for the narrow wall to broad wall coupler, which takes into account the effect of wall proximity on slot coupling. The measured results for the manufactured coupler are in excellent agreement with this theory. PMID- 11257826 TI - Dielectric properties of dates at 2.45 GHz determined with a tunable single-mode resonant cavity. AB - A tunable TM012-mode resonant cavity with an additional tuning mechanism and working at 2.45 GHz has been designed, fabricated and tested for determination of dielectric properties of dates. The cavity has a Q-factor > 5000, and a tuning mechanism which gives it more flexibility and controllability. The cavity has been used for determining the dielectric properties of Rezaiz, the most common type of dates used in the production of data juice in the Eastern Province of the Kingdom of Saudi Arabia. The dielectric constant for this type of dates was 4.6 +/- 0.16, and the loss factor was 0.21 +/- 0.03, at 8.75% moisture. These results are comparable with the dielectric properties of some other fruits, of similar composition, at the same moisture level. PMID- 11257827 TI - Improving electric field distribution in a microwave heating device. AB - The authors describe a microwave reheater for food products in which two magnetrons and a magic tee are used for decoupling the waves emitted and for obtaining good impedance matching. Symmetric and asymmetric parts of the electric field pattern can be separately adjusted and optimized for best results. PMID- 11257828 TI - Tobacco and laryngeal pathology. PMID- 11257829 TI - Smoking and cardiovascular disease. PMID- 11257830 TI - Smoking and lung disease. PMID- 11257831 TI - Chronic obstructive pulmonary disease. PMID- 11257832 TI - Second-hand cigarette smoke is a major contributor to asthma in children. PMID- 11257833 TI - Smoking and bladder cancer. PMID- 11257834 TI - Smoking and renal cell carcinoma. PMID- 11257835 TI - Pregnant women should not smoke: no ifs, ands, nor butts. PMID- 11257837 TI - Tobacco is killing (and costing) us. PMID- 11257836 TI - Treating nicotine dependence. AB - To explore the biology of nicotine addiction and to discuss the latest effective treatments for nicotine dependence. Research indicates that the most effective methods for treating nicotine dependence are nicotine replacement therapy (NRT) and bupropion SR (BUP). We conclude that pharmacological adjuncts (NRT & BUP) achieve their greatest success when combined with behavioral counseling. PMID- 11257838 TI - A comprehensive tobacco control program critical for West Virginia. PMID- 11257839 TI - Possible savings from new efforts to reduce smoking in West Virginia. PMID- 11257840 TI - Tobacco & taxes. PMID- 11257841 TI - Benefits from a West Virginia cigarette tax increase. PMID- 11257842 TI - Youth tobacco use and control efforts in West Virginia. PMID- 11257843 TI - WVSMA's health programs focus on reducing tobacco use. PMID- 11257844 TI - WVSMA alliance: an integral part of West Virginia's tobacco control efforts. PMID- 11257845 TI - PEIA and Medicaid offer tobacco cessation benefit. PMID- 11257846 TI - Big tobacco's fight against smoking regulations in Harrison County. PMID- 11257848 TI - Mexico and beyond. PMID- 11257847 TI - Tobacco cancer of the oral cavity and pharynx. AB - Cancer of the oral cavity and pharynx affects a significant number of individuals worldwide. The most important risk factor for development of oral cavity cancer is cigarette smoking. Tobacco in other forms and other types of smoking are also thought to be major risk factors. Co-factors, particularly alcohol consumption, are also important factors in oral cancer. Management of oral cancer requires a multidisciplinary team and has major implications for patient quality of life and for public health. Efforts are underway in many countries to reduce the incidence of oral cancer. These efforts always emphasize cessation of tobacco use and cancer screening. Local smoking cessation programs should be supported to improve the future of health care in West Virginia. PMID- 11257849 TI - Strengthening environmental and educational nutrition programmes in worksite cafeterias and supermarkets in The Netherlands. AB - The purpose of this study was to assess conditions for the adoption and continued implementation of different healthy nutrition programmes in worksite cafeterias and supermarkets, i.e. an educational programme and two environmental programmes (a food labelling programme and a food supply programme). Twenty semi-structured interviews were conducted with representatives of worksite cafeterias and supermarkets. Concepts of theories of diffusion were used as a framework for the study. Questions were formulated about the attributes of the innovation, and organizational and personal characteristics that might influence programme adoption and implementation. Results indicated that educational and environmental programmes in both worksite cafeterias and supermarkets should meet specific requirements regarding programme design, methods and materials in order to be adopted and implemented. Besides, some important implementation strategies of the educational and environmental programmes were identified. It is concluded that it seems feasible to conduct educational and environmental intervention programmes in worksite cafeterias and supermarkets, but that certain conditions for adoption and continued implementation have to be met. Based on the implications of this study, the development of an educational programme, a labelling programme and a food supply programme was completed. PMID- 11257850 TI - Mexico conference on health promotion: open letter to WHO Director General, Dr Gro Harlem Brundtland. PMID- 11257851 TI - Community involvement at what cost?--local appraisal of a pan-European nutrition promotion programme in low-income neighbourhoods. AB - In the UK, government has committed itself to improving health and reducing inequalities in health. For the first time, issues such as food poverty will be addressed by tackling the causes of poverty and wider determinants of ill health. The time has never been better, therefore, for health and local authorities to work collaboratively to promote and improve health. Community involvement is also paramount to sustainable programmes. However, such a dramatic shift in policy and greater emphasis on public health requires health professionals themselves to adopt a different approach. The World Health Organization (WHO) recommends a health promotion approach as a framework for action. But despite the existence of this framework there is little evidence that a wider understanding of health promotion and the necessary practical experience has been achieved. This has weakened the potential impact of health promotion and has possibly encouraged inappropriate use of health promotion principles in practice. The European Food and Shopping Research Project (SUPER project) was established under the WHO European network of Healthy Cities to help local projects implement the principles of health promotion (WHO, 1986). This paper describes the SUPER project and its implementation in Liverpool (1989-1997), where levels of unemployment, deprivation and ill health are amongst the highest in the UK. Participation in SUPER is appraised to identify the various benefits and obstacles involved and to identify links with progress at the local level. This appraisal is discussed and the use, and potential misuse, of participatory appraisal techniques to elicit information and mobilize communities is examined. PMID- 11257852 TI - Development of a national injury prevention/safe community programme in Vietnam. AB - The aim of this study is to describe the initiation of a national programme on injury prevention/safe community (IP/SC). Market economy, Doi Moi, was introduced in Vietnam in 1986, and since then the injury pattern has been reported to have changed. The number of traffic injury deaths has increased three-fold from 1980 to 1996 and traffic injuries more than four-fold. Injuries are now the leading cause of mortality in hospitals. There are difficulties in obtaining a comprehensive picture of the injury pattern from official statistics and, in conjunction with the work initiated by the Ministry of Health, a number of local reporting systems have already been developed. Remarkable results have been achieved within the IP/SC in a very short time, based on 20 years of experience. An organizational construction system has been built from province to local community areas. Management is based on administrative and legislative documents. IP/SC implementation is considered the duty of the whole community, local authorities and people committees, and should be incorporated into local action plans. The programme is a significant contribution towards creating a safe environment in which everybody may live and work, allowing the stability for society to develop. Implementation of the programme in schools is a special characteristic. The programme will be developed in 800 schools with a large number of pupils (25% of the population). This model for safer schools is considerably concerned and is a good experience to disseminate. The recommendations are that more pilot models of IP/SC should be conducted in other localities and that the programme should be expanded to a national scale. Furthermore, co-operation between sectors and mass organizations should be encouraged and professional skills of key SC members at all levels should be raised. PMID- 11257853 TI - Why do Swedish-speaking Finns have longer active life? An area for social capital research. AB - We performed ecological and individual register studies to compare disability free life expectancies and disability pensions among Swedish-speaking and Finnish speaking Finns residing on the western coast of Finland. The study was conducted to establish our assumption that the Swedish-speaking ethnic minority has a longer active life than the Finnish-speaking majority and to show that this disparity can be seen in a limited geographical area with similar socio-economic and health service structures. We suggest that the observed disparities in active life and in mortality depend on differences in the extent of social capital. A detailed characterization of the social capital and its impact on the health of the Swedish-speaking individuals is in progress. PMID- 11257854 TI - Smoking trends in adolescence: report on a Greek school-based, peer-led intervention aimed at prevention. AB - This article presents a school-based, peer-led programme aiming at smoking prevention among Greek adolescents. The intervention was based on the social influence approach, and utilized 28 students' personal sensitization for the development of audio-visual material, which was presented to the remaining students of the two experimental schools (n = 440). The effectiveness of the intervention was assessed by means of experimental and control (n = 217) students' completion of a questionnaire prior to, immediately after and 3 months after the intervention. The Repeated Measures Analysis of Variance (MANOVAR) showed declines in the experimental group's smoking behaviour and intention to smoke immediately after, but not 3 months after the intervention, and more lasting changes in this group's knowledge of addiction and anti-smoking attitude. These effects were not observed for the control group. Programme evaluation and the implementation of health education programmes in the Greek school curriculum are discussed in the light of these findings. PMID- 11257855 TI - Health promotion is fantastic, fascinating and fun--personal reflections. PMID- 11257856 TI - Does reported health promotion activity neglect people with ill-health? AB - Considering health as an alternative to ill-health ignores the multidimensionality of both concepts and invites neglect of health promotion as a multidimensional activity in persons with known ill-health. Drawing on the Ottawa Charter and Maori perspectives of health, we interpret (ill) health according to people's ability to function in their environment by developing physical, psychological, social and spiritual resources for living. We use this framework to test empirically our hypothesis that although the concept of health promotion has always included people with ill-health, the practice of health promotion has continued to neglect them. Our exploratory review of articles published during 1989-99 and indexed on three electronic databases suggests widespread omission of people with ill-health from research on interventions for health promotion. Of 881 citations, approximately three-quarters included people without ill-health in any dimension. This finding could reflect a failure to include these people in health promotion, to describe activity to improve their health as health promotion, or both. Supporting the latter interpretation is uncertainty over the meaning of health, and the targeting of health promotion at groups at high risk of ill-health and 'all' persons. We need therefore to enable health promotion activity to include ill people explicitly. PMID- 11257857 TI - Health impact assessment: a tool for healthy public policy. AB - Healthy Public Policy is one of the key health promotion actions. Advancement of Healthy Public Policy requires that the health consequences of policy should be correctly foreseen and that the policy process should be influenced so that those health consequences are considered. Health Impact Assessment is an approach that could assist in meeting both requirements. Policies often produce health impacts by multiple indirect routes, which makes prediction difficult. Prediction in Health Impact Assessment may be based on epidemiological models or on sociological disciplines. Health Impact Assessment must be based on an understanding of, and aim to add value to, the policy-making process. It must therefore conform to policy-making timetables, present information in a form that is policy relevant and fit the administrative structures of policy makers. Health Impact Assessment may be used to inform health advocacy but is distinct from it. There is a danger that Health Impact Assessment could be misunderstood as health imperialism. PMID- 11257858 TI - Health literacy in health systems: perspectives on patient self-management in Israel. AB - Health systems will face new challenges in this millennium. Striking the balance between the best quality of care and optimal use of dwindling resources will challenge health policy makers, managers and practitioners. Increasingly, improvements in the outcomes of interventions for both acute and chronic patients will depend on partnerships between health service providers, the individual and their family. Patient education that incorporates self-management and empowerment has proven to be cost-effective. It is essential that health care providers promote informed decision making, and facilitate actions designed to improve personal capacity to exert control over factors that determine health and improve health outcomes. It is for these reasons that promoting health literacy is a central strategy for improving self-management in health. The different types of health literacy--functional, interactive and critical health literacy--are considered. The potential to improve health literacy at each of these levels has been demonstrated in practice among diabetics and other chronic disease patients in Clalit Health Services (CHS) in Israel is used as an example to demonstrate possibilities. The application of all three types of health literacy is expressed in: (i) developing appropriate health information tools for the public to be applied in primary, secondary and tertiary care settings, and in online and media information accessibility and appropriateness using culturally relevant participatory methods; (ii) training of health professionals at all levels, including undergraduate and in-service training; and (iii) developing and applying appropriate assessment and monitoring tools which include public/patient participatory methods. Health care providers need to consider where their patients are getting information on disease and self-management, whether or not that information is reliable, and inform their patients of the best sources of information and its use. The improved collaboration with patient and consumer groups, whose goals are to promote rights and self-management capabilities and advocate for improved health services, can be very beneficial. PMID- 11257859 TI - Barriers to employment-related healthy public policy in Canada. AB - The Ottawa Charter for Health Promotion calls for building healthy public policy, that is for '[putting] health on the agenda of policy makers in all sectors and at all levels, directing them to be aware of the health consequences of their decisions and to accept their responsibilities for health'. The objective of this study was to assess the past and potential future influence of information about the health consequences of unemployment and job insecurity on policy making and to identify the barriers to the use of such information in policy making. We conducted telephone interviews with 38 policy makers in the health and employment sectors of all three levels of Canadian government, as well as the executive directors of 10 Canadian non-governmental organizations that are active on employment issues. The interviews included both numerical ratings of the influence of this information and semi-structured questions about how this information could be used in policy making. Using an interpretive approach grounded in the political science literature, we identified barriers to using this information in their responses to these questions. Respondents rated the potential future influence of this information (mean 4.2 and median 5 on a seven point Likert scale) higher than its past influence (mean 3.5 and median 3 on a seven-point Likert scale). Barriers related to the information itself or more commonly to the values of those who could respond to the information (i.e. idea related barriers) were cited more frequently than either barriers related to how decisions are made (i.e. institution-related barriers) or barriers related to who would win and who would lose if the information were acted upon (i.e. interest related barriers). We concluded that to build employment-related healthy public policy, these barriers would have to be overcome. Policy makers in health departments could, for example, frame information about health consequences in language that fits more easily with the values of other departments and advocate for institutional innovations that establish cross-departmental or cross governmental accountability for health. PMID- 11257860 TI - Statement by Dr Gro Harlem Brundtland, Director General WHO, to the Fifth Global Conference on Health Promotion, Mexico City, 5 June 2000. PMID- 11257861 TI - Letter from Canada: paradigms, politics and principles. An end of the millennium update from the birthplace of the Healthy Cities movement. PMID- 11257862 TI - Atmospheric water vapour processor designs for potable water production: a review. AB - Atmospheric water vapour processing (AWVP) technology is reviewed. These processors are machines which extract water molecules from the atmosphere, ultimately causing a phase change from vapour to liquid. Three classes of machines have been proposed. The machines either cool a surface below the dewpoint of the ambient air, concentrate water vapour through use of solid or liquid desiccants, or induce and control convection in a tower structure. Patented devices vary in scale and potable water output from small units suitable for one person's daily needs to structures as large as multi-story office buildings capable of supplying drinking water to an urban neighbourhood. Energy and mass cascades (flowcharts) are presented for the three types of water vapour processors. The flowcharts assist in classifying designs and discussing their strengths and limitations. Practicality and appropriateness of the various designs for contributing to water supplies are considered along with water cost estimates. Prototypes that have been tested successfully are highlighted. Absolute humidity (meteorological normals) ranges from 4.0 g of water vapour per cubic metre of surface air in the atmosphere (Las Vegas, Nevada, USA) to 21.2 g m 3 (Djibouti, Republic of Djibouti). Antofagasta, Chile has a normal absolute humidity of 10.9 g m-3. A 40% efficient machine in the vicinity of Antofagasta requires an airflow of 10 m3 s-1 to produce 3767 l of water per day. At a consumption of 50 l per person per day, 75 people could have basic water requirements for drinking, sanitation, bathing, and cooking met by a decentralized and simplified water supply infrastructure with attendant economic and societal benefits. PMID- 11257864 TI - Kinetic modeling of pyruvic acid ozonation in aqueous solutions catalyzed by Mn(II) and Mn(IV) ions. AB - The ozonation of pyruvic acid (2-ketopropionic acid) in aqueous solutions, catalyzed by Mn(II) and Mn(IV) ions, has been studied at three different pH values (pH = 1.1, 2.0 and 3.0). A mathematical model of the unsteady operation of the experimental reactor has been developed, which takes into account the reactions occurring in the liquid phase and the ozone mass transfer from the gas bubbles. Those reactions have been described with two alternative kinetic models, both made out of four elementary steps. The two kinetic models correlate the experimental data with a fair accuracy, respectively at the lowest and at the highest pH examined. In particular, at pH = 3.0, the ozonation results are inhibited by the acetate ions produced by the reaction itself. This effect has been correctly described in the terms of a complex formed with the low oxidation state manganese, which successively reacts with the dissolved ozone. PMID- 11257863 TI - Removal of chlorine dioxide disinfection by-products by ferrous salts. AB - Chlorine dioxide when used as an effective disinfectant forms undesirable disinfection by-products, i.e. chlorite and chlorate ions. The aim of this research was to study the removal of these ions by ferrous ions in the presence or absence of oxygen. The efficiency of Fe+2 for ClO2- and ClO3- removal was followed by a determination of their initial and final concentrations, pH and delta Fe+2 consumed/delta ClO2- removed ratios. The optimal weight ratio of delta Fe+2 consumed/delta ClO2- removed for complete ClO2 removal was found to be close to the theoretical calculated value of 3.31. It was proved that ferrous salts can reduce chlorite ions to harmless Cl- ions. This method can be recommended as a part of ClO2 disinfection to ensure safe drinking water, with no harm to water consumers and to the environment. PMID- 11257865 TI - Polycyclic musk fragrances in different environmental compartments in Berlin (Germany). AB - The aim of this study was to obtain data about the contamination of different environmental compartments (102 surface water samples, 59 sediment samples and 165 eel samples) by polycyclic musks (HHCB, AHTN, ADBI, AHMI, and ATII) within the framework of an exposure monitoring program. Results for HHCB (Galaxolide) gave the following mean values in areas strongly polluted with sewage: surface water 1.59 micrograms l-1; sediment 0.92 mg kg-1 d.w. and eel 1513 micrograms kg 1 f.w. (in the edible portion) (6471 micrograms kg-1 lipid). The following average concentrations were found in waters hardly contaminated with sewage: surface water 0.07 microgram l-1, sediment < 0.02 mg kg-1 and eel 52 micrograms kg-1 f.w. (445 micrograms kg-1 lipid). Mean concentrations of 6.85 micrograms l-1 (maximum: 13.3 micrograms/l) could be measured at sewage treatment plants' outlets. It could be shown that these polycyclics are highly suited to use as indicators of the degree of contamination of waters with organic substances originating from sewage. A mean bioconcentration factor (BCF) on wet weight of 862 (HHCB) and 1069 (AHTN) for the transfer from water to eel under natural conditions could be calculated. The corresponding BCF-values based on the lipid content of eel were 3504 (HHCB) and 5017 (AHTN). PMID- 11257866 TI - Influence of transient substrate overloads on the proliferation of filamentous bacterial populations in an activated sludge pilot plant. AB - Using oligonucleotide probes directed at the rRNA of filamentous bacteria, this study looks at the influence of the components of transient substrate overloads on the growth of the dominant filamentous bacteria of activated sludge fed by a synthetic substrate. By dissociating the massive input of organic matter from the oxygen shortage that the latter generally induces, it is revealed that each of these factors applied alone, induces only transitory, small-scale growth of the filaments Nostocoida limicola, Haliscomenobacter hydrossis. Thiothrix and of type 021N. In contrast, combining them during a reconstituted transient substrate overload with an artificially created oxygen deficit, induces very fast growth of H. hydrossis which is responsible for establishing major proliferation. This massive proliferation was easily reduced by chlorination. PMID- 11257867 TI - Photocatalytic reduction of Cr(VI) in aqueous solutions by UV irradiation with the presence of titanium dioxide. AB - The reduction of Cr(VI) in aqueous solution by UV/TiO2 reduction process was studied under various solution pH values, TiO2 dosages, light intensities, dissolved oxygen levels and other operating conditions. The reduction rates of Cr(VI) by photocatalytic-induced elections were significantly higher for acidic solutions than those for alkaline solutions. Increasing the light intensity would drastically increase the reduction rate of Cr(VI), but was ultimately influenced by the amount of TiO2 present in solutions. The presence of dissolved oxygen had minimum effect on the reduction of Cr(VI) by UV/TiO2 process in acidic solutions. The presence of ethanol might act as scavenger for holes and promoted the photocatalytic reduction of Cr(VI) by electrons. The Cr(VI) adsorbed on the surface of TiO2 particles was observed to be photoreduced to Cr(III) almost completely. PMID- 11257868 TI - Transfer of organic matter between wastewater and activated sludge flocs. AB - The organic matter of wastewater was fractionated into settleable (i.e., particulate) and non-settleable (i.e., colloidal + soluble) fractions by settling followed by 0.22 micron filtration. Particulate, colloidal and soluble proportions were found to be relatively constant (45, 31 and 24% of the total COD, respectively). Transfer of soluble fraction always occurred from the wastewater to the activated sludge flocs, whereas bidirectional transfer occurred for the colloidal fraction. The transfer of soluble and colloidal matter reached a steady state after 40 min-mixing and 20 min-mixing, respectively. Desorption of a part of the colloidal organic matter pre-sorbed on the activated sludge flocs was evidenced. The biosorption capacity of activated sludge was around 40-100 mgCODg-1TSS. The biosorbable fraction of wastewater represented on average 45% of the non-settleable fraction. PMID- 11257870 TI - Hybrid reactor for priority pollutant-trichloroethylene removal. AB - The present study was initiated to explore the potential of a hybrid biological reactor, combining trickling filter (TF) and activated sludge process (ASP), to treat wastewater containing trichloroethylene (TCE) at ambient temperature at different hydraulic retention time (HRT). The biofilm acclimation was achieved in 55-60 days with gradual increase in TCE concentration from 1 mg/l to 100 mg/l with a parallel increase in the concentration of substrate sodium acetate and other nutrients. COD and TCE concentration were taken as prime parameters for monitoring the growth of biofilm. During acclimation COD removal varied between 54.6-97.5% while TCE was removed 72.6-99.9%. HRT study was performed after acclimation. The removal efficiency increased with decreasing flow rate with maximum TCE removal (99.99%) at 6 l/d corresponding to an HRT of 28 h (TF 18 h + ASP 10 h). This was followed by a C:N:P ratio study. A ratio of 100:20:1 led to the sustenance of maximum TCE removal. Maximum TCE removal (99.99%) was observed at a substrate:cosubstrate ratio of 100:1. A pH of 7.4 +/- 0.2 was found to be optimum for degradation. Finally, volatilization losses were estimated to be 18.5%. A mass balance gave an efficiency of 81.51% for biological removal of TCE. PMID- 11257869 TI - Controlled struvite crystallisation for removing phosphorus from anaerobic digester sidestreams. AB - Enhanced biological phosphorus removal wastewater treatment plants that use anaerobic digesters for sludge treatment, have high phosphorus concentrations in the sidestreams from their sludge dewatering equipment. To remove phosphorus from such sidestreams controlled struvite crystallisation can be used. Struvite (or MAP) is a naturally occurring crystal of magnesium, ammonium and phosphate. We present operational results obtained with a continuously operated pilot-scale MAP reactor. The pilot-scale reactor (143 l) was an air agitated column reactor with a reaction and a settling zone, based on the Phosnix process of Unitika Ltd., Japan. The influent to the MAP reactor was centrate from the centrifuge that dewaters anaerobically digested sludge at the Oxley Creek wastewater treatment plant in Brisbane. We used a 60% magnesium hydroxide slurry to add the required magnesium to the process and to obtain the alkaline pH value required. The pilot scale MAP process achieved an ortho-P removal ratio of 94% from an average influent ortho-P concentration of 61 mg/l. The reactor was operated at a pH of around 8.5. Insufficient dosing of magnesium reduced the P removal performance. There was no influence of the hydraulic residence time on the process in the range of 1-8 h. The dry MAP product had cadmium, lead and mercury concentrations well below the legal limits for fertilisers in Queensland, Australia and can be reused as a valuable slow-release fertiliser. PMID- 11257871 TI - Pore distribution effect of activated carbon in adsorbing organic micropollutants from natural water. AB - Adsorption isotherms of organic micropollutants in coexistence with natural organic matter (NOM) were analyzed to evaluate the impacts of pore size distribution of activated carbon (AC) on the competition effects of the NOM. Single solute adsorption experiments and simultaneous adsorption experiments with NOM contained in a coagulation-pretreated surface water were performed for four agricultural chemicals and three coal-based activated carbons (ACs) having different pore distributions. The results showed that, for all the carbons used, the adsorption capacity of the chemicals was reduced distinctly in the presence of NOM. Such a reduction was more apparent for AC with a larger portion of small pores suitable for the adsorption of small organic molecules and for the agricultural chemicals with a more hydrophilic nature. Ideal adsorbed solution theory (IAST) incorporated with the Freundlich isotherm expression (IAST Freundlich model) could not interpret the impact of NOM on the adsorption capacity of the chemicals unless a pore blockage effect caused by the adsorption of NOM was also considered. By taking into account this effect, the adsorption isotherm of the chemicals in the presence of NOM was well described, and the capacity reduction caused by the NOM was quantitatively assessed from the viewpoints of the site competition and the pore blockage. Analytical results clearly indicated that pore blockage was an important competition mechanism that contributed to 10-99% of the total capacity reductions of the chemicals, the level depended greatly on the ACs, the chemicals and the equilibrium concentrations, and could possibly be alleviated by broadening the pore size distributions of the ACs to provide a large volume percentage for pores with sizes above 30 A. PMID- 11257872 TI - A titrimetric respirometer measuring the nitrifiable nitrogen in wastewater using insensor-experiment. AB - Measurement of nitrifiable nitrogen contained in wastewater by combining the existing respirometric and titrimetric principles is reported. During an in sensor-experiment using nitrifying activated sludge, both the dissolved oxygen (DO) and pH in the mixed liquor were measured, and the pH was controlled at a set point through titration of base or acid. A combination of the oxygen uptake rate (OUR), which was obtained from the measured DO signal, and the titration data allowed calculation of the nitrifiable nitrogen and the short-term biological oxygen demand (BOD) of the wastewater sample that was initially added to the sludge. The calculation was based solely on stoichiometric relationships. The approach was preliminarily tested with two types of wastewaters using a prototype sensor. Good correlation was obtained. PMID- 11257873 TI - Combined phosphate and nitrogen removal in a sequencing batch reactor using the aerobic denitrifier, Microvirgula aerodenitrificans. AB - A phosphate removal sludge was bioaugmented with the aerobic denitrifier, Microvirgula aerodenitrificans in order to reduce the nitrate produced during the aerobic nitrifying-phosphate uptake phase. Fluorescent in situ hybridization (FISH) was used to follow the fate of the added strain. In order to maintain the pure strain in the complex ecosystem, diverse physiological and kinetic based strategies of bioaugmentation were tested under the sequencing batch reactor (SBR) type culture. The nature of the M. aerodenitrificans inoculum (adapted to nitrate-aerobic conditions or to anoxic one) had no influence on the SBR performances and did not enhance aerobic denitrifying performances. The optimum quantity of the added strain (10% of the total biomass) seemed to have much more positive influence on the long term maintenance of the pure strain than on the SBR performances. A small but daily supply of M. aerodenitrificans gave exactly the same result than a massive and 1-day supply, i.e. no enhancement of performances and no amelioration of the length of maintenance. A continuous supply of carbon during the first hour of the aerobic phase combined to a 10% supply of M. aerodenitrificans gave the best compromise in terms of phosphate removal, nitrification and aerobic denitrification performances. It was accompanied too by a decreased number of the ammonia and nitrite-oxidizing bacteria and a modification of the nitrite-oxidizing floc structure. FISH on M. aerodenitrificans revealed that (i) before bioaugmentation, the strain was already present in the phosphate removal sludge and (ii) the added bacteria almost disappeared from the reactor after 16 HRT. In a last experiment, M. aerodenitrificans embedded in alginate beads allowed enhancement of both aerobic denitrifying performances and length of strain maintenance. PMID- 11257875 TI - Non-steady state simulation of BOM removal in drinking water biofilters: applications and full-scale validation. AB - A biofilter model called "BIOFILT" was used to simulate the removal of biodegradable organic matter (BOM) in full-scale biofilters subjected to a wide range of operating conditions. Parameters that were varied included BOM composition, water temperature (3.0-22.5 degrees C), and biomass removal during backwashing (0-100%). Results from biofilter simulations suggest a strong dependence of BOM removal on BOM composition. BOM with a greater diffusivity or with faster degradation kinetics was removed to a greater extent and also contributed to shorter biofilter start-up times. In addition, in simulations involving mixtures of BOM (i.e. readily degradable and slowly degradable components), the presence of readily degradable substrate significantly enhanced the removal of slowly degradable material primarily due to the ability to maintain greater biomass levels in the biofilters. Declines in pseudo-steady state BOM removal were observed as temperature was decreased from 22.5 to 3 degrees C and the magnitude of the change was significantly affected by BOM composition. However, significant removals of BOM are possible at low temperatures (3-6 degrees C). Concerning the impact of backwashing on biofilter performance, BOM removal was not affected by backwash resulting in biomass removals of 60% or less. This suggests that periodic backwashing should not significantly impact biofilter performance as observed biomass removals from full scale biofilters were negligible. In general, the simulation results were in good qualitative and quantitative agreement with experimental results obtained from full-scale biofilters. PMID- 11257874 TI - Non-steady state simulation of BOM removal in drinking water biofilters: model development. AB - A numerical model was developed to simulate the non-steady-state behavior of biologically-active filters used for drinking water treatment. The biofilter simulation model called "BIOFILT" simulates the substrate (biodegradable organic matter or BOM) and biomass (both attached and suspended) profiles in a biofilter as a function of time. One of the innovative features of BIOFILT compared to previous biofilm models is the ability to simulate the effects of a sudden loss in attached biomass or biofilm due to filter backwash on substrate removal performance. A sensitivity analysis of the model input parameters indicated that the model simulations were most sensitive to the values of parameters that controlled substrate degradation and biofilm growth and accumulation including the substrate diffusion coefficient, the maximum rate of substrate degradation, the microbial yield coefficient, and a dimensionless shear loss coefficient. Variation of the hydraulic loading rate or other parameters that controlled the deposition of biomass via filtration did not significantly impact the simulation results. PMID- 11257876 TI - Optimisation of the storage of natural freshwaters before organotin speciation. AB - The speciation of organotin compounds is essential due to the species-dependent toxicity, especially in natural waters. Precautions have to be taken during sampling and storage of waters in order to prevent degradations and losses. Experimental design methodology has been used to study the conditions of stability of organotins after water sampling in rivers. The modelling of results allows the determination of optimal conditions of preservation. After acidification at pH = 4 with nitric acid, the storage in polyethylene containers at 4 degrees C in the dark is suitable to preserve the most degradable trisubstituted (butyl- and phenyl-) species over 1 month. These conditions of sampling and storage are applied to two different freshwaters. The rate of species decomposition appears to be only dependent on the water nature, whatever the organotin concentrations in the sample. Speciation could be so preserved between 1 and 3 months. PMID- 11257877 TI - Ozonation of NOM and algal toxins in four treated waters. AB - The occurrence of cyanobacteria (blue-green algae) blooms and the possibility of production of cyanotoxins (algal toxins) have become major concerns for drinking water providers worldwide. Ozone has been shown to be effective for the destruction of some classes of toxins under specific conditions, although most researchers agree that the dose and contact time required will depend on water quality. The clarification of the relative effects of water quality parameters such as dissolved organic carbon concentration and character, and alkalinity, has not been previously attempted. In this study the cyanotoxins microcystin LR and LA and anatoxin-a were ozonated at a range of ozone doses in four treated waters with very different water quality. For both the toxins, 100% destruction was related to a residual ozone concentration present after 5 min. This was, in turn, related to the water quality and indicated that a direct reaction with molecular ozone could be responsible for the destruction. The results confirmed that both the toxins would be destroyed under conditions usually utilised for ozonation prior to granular activated carbon (GAC) filtration. This will apply under a range of water quality conditions but not necessarily a range of temperatures. The saxitoxin class of compounds was very resistant to oxidation by ozone and would require further treatment such as GAC filtration. PMID- 11257878 TI - Photolysis of polycyclic aromatic hydrocarbons in water. AB - The decomposition of benzo[a]pyrene (BAP), chrysene (CHR) and fluorene (FLU) in an aqueous solution by means of photolysis has been studied. The influence of initial polycyclic aromatic hydrocarbons' (PAHs) concentration, pH of the reaction mixture, temperature, presence of oxygen and tert-butyl alcohol (t-BuOH) on the degradation rate has been observed. BAP and CHR are decomposed by a mechanism different than FLU. Quantum yields of the photolytic decomposition of BAP, CHR and FLU were determined and equal 0.014, 0.0031 and 0.0038, respectively. PMID- 11257879 TI - Chlorination of pure bacterial cultures in aqueous solution. AB - The fate and distribution of chlorine in aqueous solutions containing four pure bacterial cultures was studied. Solutions were subjected to chlorination at different initial free chlorine concentrations. Resulting concentrations of residual chlorine were determined by both DPD/FAS titration and membrane introduction mass spectrometry (MIMS). In all cases, false-positive breakpoint chlorination curves, probably attributable to the formation of chloroorganic-N compounds, were observed by DPD/FAS titration, while little or no inorganic residual chloramine was found by MIMS. Free chlorine was observed in similar quantities by both methods after chlorine demand by bacterial cellular materials in solution was satisfied. These results indicated the residual chloramines existed in the form of organic chloramines; these compounds are generally recognized as being poor antimicrobial agents. Further investigation confirmed that the bacterial cells were the source of organic-N compounds. The kinetics of chlorination of pure bacterial suspensions was also studied. The pattern of residual chlorine decay following chlorination of the bacterial suspensions indicated rapid initial free chlorine consumption, followed by slow free chlorine consumption, with trace quantities of inorganic chloramine being formed. PMID- 11257880 TI - Effect of lipids and oleic acid on biomass development in anaerobic fixed-bed reactors. Part I: Biofilm growth and activity. AB - Two similar anaerobic fixed-bed bioreactors which allowed the biomass to be periodically withdrawn were run in parallel. After feeding each digester with synthetic dairy wastes of different lipid content (Period I), both digesters were fed with increasing sodium oleate concentrations with skim milk as co-substrate (Period II) and oleate as the sole carbon source (Period III). In Period I, the digester fed with lipids was more efficient and exhibited lower levels of volatile fatty acids than the digester fed without lipids. The biofilm built up in the presence of lipids was thinner, but more resistant to the presence of oleate than the biofilm formed in the absence of lipids, which lost 53% of its solids after contacting with oleic acid. The specific methanogenic activity with butyrate as substrate was enhanced in the presence of lipids, but no significant effect was detected on the acetoclastic and hydrogenophilic activities, which remained similar for both digesters along the trial period. PMID- 11257881 TI - Effects of lipids and oleic acid on biomass development in anaerobic fixed-bed reactors. Part II: Oleic acid toxicity and biodegradability. AB - Oleic acid toxicity and biodegradability were followed during long-term operation of two similar anaerobic fixed-bed units. When treating an oleate based effluent, the sludge from the bioreactor that was acclimated with lipids during the first operation period, showed a higher tolerance to oleic acid toxicity (IC50 = 137 mg/l) compared with the sludge fed with a non-fat substrate (IC50 = 80 mg/l). This sludge showed also the highest biodegradation capacity of oleic acid, achieving maximum methane production rates between 33 and 46 mlCH4(STP)/gVS.day and maximum percentages of methanization between 85 and 98% for the range of concentrations between 500 and 900 mg oleate/l. When oleate was the sole carbon source fed to both digesters, the biomass became encapsulated with organic matter, possibly oleate or an intermediate of its degradation, e.g. stearate that was degraded at a maximum rate of 99 mlCH4(STP)/gVS.day. This suggests the possibility of using adsorption-degradation cycles for the treatment of LCFA based effluents. Both tolerance to toxicity and biodegradability of oleic acid were improved by acclimatization with lipids or oleate below a threshold concentration. PMID- 11257882 TI - Zooplankton assemblages and biomass during a 4-period survey in a northern Mediterranean coastal lagoon. AB - The authors proposed to examine zooplankton biomass at three stations inside (T and Z) and outside (M) a coastal lagoon of the north-western Mediterranean Sea. Station T represented the lagoon central area, and station Z was positioned in a shellfish farming sector, while the seaside station (M) served as a reference of marine environment. Analyses were designed to outline the net zooplankton assemblages (taxonomic structures and length distributions) in different environmental conditions, including the farming activity. A discriminant analysis of environmental variables determined that temperature, salinity and phytoplankton implied mainly in spatial pattern of the samples. An ordination of taxa biomasses showed two main factors which might contribute to the organisation of the zooplankton assemblages: the geographical position and the thermal period. The geographical position integrated the lagoon-sea water exchange under forcing parameters (habitat, tides and winds). The thermal period reflected both the populations development cycles and the environmental constraints (temperature, salinity, trophic resources). The resulting effects appeared in structured zooplankton assemblages in space and time. The number of 50 microns interval length classes and of taxa decreased from the seaside and the lagoon central area free of farming activity to the shallower farming zone. But the biomass-length distribution profiles did not closely follow such an expected opposition between opened and confined areas: more extended profiles were observed at station Z. Biomass dominant size classes concerned the range up to 300 microns. This size category appeared to collapse in terms of biomass from the seaside or central area of the lagoon towards the farming area, similarly to zooplankton global biomass fluctuations. Difference between biomass levels and between biomass structures suggested that net zooplankton partly acted as food competitors of macro-filtering organisms, and as preys for farming shellfish and associated epifauna. This impact mainly concerned microzooplankton populations. PMID- 11257884 TI - Anaerobic biodegradation of naphthalene, phenanthrene, and biphenyl by a denitrifying enrichment culture. AB - In previous results [Rockne and Strand (1998) Environ. Sci. Technol. 32, 2962 3967], anaerobic biodegradation of the polycyclic aromatic hydrocarbons, naphthalene, phenanthrene, and biphenyl in a fluidized bed reactor (FBR) enrichment was demonstrated. In this paper, re-feeding and mineralization experiments with sub-cultures of the nitrate-reducing enrichment are described. The subcultures continued to remove the PAHs after three feedings. PAH biodegradation ceased when nitrate was depleted and resumed when the enrichment was fed nitrate, demonstrating that PAH biodegradation was dependent upon nitrate reduction. Tests with radiolabeled PAH confirmed that PAH was mineralized, although the extent of mineralization differed greatly with different PAHs. Only partial mineralization (17% of initial carbon) was observed when the culture was fed naphthalene, whereas nearly complete mineralization (96%) was observed with phenanthrene. PAH carbon was incorporated into cell mass and mineralized after complete biodegradation of the PAHs, with 78-102% recoveries of radiolabel for naphthalene and phenanthrene, respectively. PAH carbon incorporation into biomass also varied considerably. Minor assimilation of biphenyl or phenanthrene was observed in the culture, whereas extensive assimilation of naphthalene carbon (57%) was observed when the culture was challenged with naphthalene. PAH degradation was approximately stoichiometric with the amount of nitrate consumed. Headspace analysis showed production of N2O, suggesting the enrichment coupled the biodegradation of PAH to denitrification. PMID- 11257883 TI - Chalk as the carrier for nitrifying biofilm in a fluidized bed reactor. AB - A fluidized bed reactor for nitrification with chalk as the biomass carrier and the sole buffer agent was studied. Chalk dissolution in the reactor was found to follow the stoichiometric ratio of 1 mole of CaCO3 dissolved for each mole of NH4+ oxidized. Three batches of chalk, each one having a different dissolution rate, were used to replace the dissolved chalk. The three dissolution rates resulted in three different steady state pH levels in the reactor (4.7-6.6) and nitrification rates. Nitrification was found to be limited by either the chalk dissolution rate or dissolved oxygen concentration depending on the type of chalk used. A maximal nitrification rate of 1.44 g NH4(+)-N/l reactor.d was observed. The average cell yield was 0.1 g cells/g N oxidized, similar to the cell yield during reactor start-up when the pH was 7. The specific ammonium oxidation rates varied between 0.08 and 0.15 mg NH4(+)-N oxidized/mg protein.h, values which are in the reported range for nitrification at pH 7 to 8. Oxygen update rate (OUR) results indicated that the major mechanism responsible for the high nitrification rate observed in the reactor operating at low pH seems to be the favorable microenvironment provided by the chalk. PMID- 11257885 TI - Immobilization of heavy metals from aqueous solutions using polyacrylamide grafted hydrous tin (IV) oxide gel having carboxylate functional groups. AB - A new adsorbent containing a carboxylate group has been prepared by the surface modification of a polyacrylamide grafted hydrous tin (IV) oxide gel. The product exhibits a very high adsorption potential for Pb(II), Hg(II) and Cd(II). The effect of initial metal ion concentration, adsorbent dose, pH, concentration of light metal ions, and temperature on metal removal has been studied. The process follows a first-order rate kinetics. The intraparticle diffusion of metal ions through pores in the adsorbent was shown to be the main rate limiting step. The equilibrium data fit well with the Langmuir adsorption isotherm. The selectivity order of the adsorbent is Pb(II) > Hg(II) > Cd(II). Adsorption rate constants and thermodynamic parameters were also presented to predict the nature of adsorption. The method was applied on synthetic wastewaters. Acid regeneration has been tried for several cycles with a view to recover the adsorbed metal ions and also to restore the sorbent to its original state. PMID- 11257886 TI - Hydrodynamical effects on spatial distribution of enteric bacteria in the Jordan River-Lake Kinneret contact zone. AB - The aim of this study was to determine under what hydrodynamic conditions the change in the number of enteric bacteria in the water of the River Jordan--Lake Kinneret contact zone was due to sedimentation and under what conditions the change was due to dilution. The data were then utilized to build a conceptual model explaining the distribution of biological pollutants (bacteria) in the river-lake contact zone of a shallow tropical lake. The study uses, as an example, the microbial communities of the River Jordan--Lake Kinneret contact zone. The changes in numbers of three groups of bacteria (fecal coliforms, Escherichia coli and Klebsiella pneumoniae) along the jet flow agree well with changes in the concentration of suspended particulate matter, caused by the sedimentation of particles. PMID- 11257887 TI - Metal uptake in a coastal diatom influenced by major nutrients (N, P, and Si). AB - The influence of major nutrient additions on trace metal uptake (Cd, Se, and Zn) in a coastal diatom Thalassiosira pseudonana was investigated. A short-term exposure (5 h) was employed to measure the concentration factor (ratio of metal concentration in the cells to metal concentration in the medium) which was used as a kinetic parameter to quantify the relative rate of metal uptake in the cells. Following an initial rapid surface sorption, a linear pattern of uptake over time was typical for Cd, Se and Zn, indicating that these metals may have been transported intracellularly during the short-term exposure period. N addition significantly increased the rates of Cd uptake in the cells. Although the concentration factor of Zn increased with N addition, statistical analysis indicated that N addition did not significantly affect the rate of Zn uptake in the cells. Se uptake in the cells was independent of N additions, but was lowered with increasing Si concentration. Si addition did not significantly affect Cd and Zn uptake. Similarly, P addition did not influence the rates of metal uptake in the cells. There was a significant correlation between the rate of uptake of Cd and Zn and the cell growth rate. This study demonstrated that water quality brought about by a change in nutrient condition could considerably influence the uptake of metals by marine phytoplankton. PMID- 11257888 TI - Regeneration of surfactant-modified zeolite after saturation with chromate and perchloroethylene. AB - Surfactant-modified zeolites (SMZ) have drawn recent attention as sorbents due to their removal of multiple types of contaminants and low material cost. However, like most sorbents, SMZ has a finite sorption capacity for different contaminants. The potential applications, economics, and efficiency of SMZ as a sorbent are related to the ability to regenerate the material. This paper reports several methods to regenerate chromate- and perchloroethylene-saturated SMZ. Regeneration of chromate-saturated SMZ was achieved by flushing with a sodium carbonate/sodium hydroxide solution. However, this high-pH solution increased the counterion competition for chromate sorption sites and decreased the chromate sorption capacity of the regenerated SMZ. As an alternative regeneration method, chromate sorbed to SMZ was reduced to Cr(III) in situ using sodium dithionite solution. Although reduction with dithionite restored the chromate sorption maximum, the chromate sorption intensity was lowered, possibly due to the competition by sulfate (generated from oxidation of dithionite) for chromate sorption sites. Carbonate-regenerated SMZ showed no loss of sorption affinity for perchloroethylene (PCE) compared to virgin SMZ. Air sparging of PCE-saturated SMZ fully regenerated the SMZ. The results show that it is possible to regenerate and re-use SMZ following saturation with anionic or nonpolar organic contaminants. PMID- 11257889 TI - Evaluation of a second derivative UV/visible spectroscopy technique for nitrate and total nitrogen analysis of wastewater samples. AB - A method for nitrate analysis based on second derivative UV/Visible spectroscopy was developed by Simal et al. (1985: Simal J., Lage M. A., and Iglesias I. (1985) Second derivative ultraviolet spectroscopy and sulfamic acid method for determination of nitrates in water. J. Assoc. Analyt. Chem. 68, 962-964) and Suzuki and Kuroda (1987: Suzuki, N. and Kuroda R. (1987) Direct simultaneous determination of nitrate and nitrite by ultraviolet second-derivative spectrophotometry. Analyst 112, 1077-1079), and later modified for the analysis of total nitrogen in aqueous samples of varying nitrate:organic nitrogen ratios (Crumpton et al., 1992: Crumption W. G., Isenhart T. M. and Mitchell P. D. (1992) Nitrate and organic N analyses with second-derivative spectroscopy. Limnol. Oceanogr. 37, 907-913). The procedure uses the second derivative of the absorption spectrum for nitrate (NO3-), which has a peak at approximately 224 nm that is proportional to the NO3- concentration. Samples for total N analysis are first oxidized to NO3- by persulfate digestion. The objectives of this study were to: (1) test the accuracy and precision of the second derivative method through the use of NIST-traceable wastewater check samples; (2) determine whether the second derivative method for nitrate analysis can be used for wastewater samples and whether the method compares favorably with other currently used nitrate analysis methods; and (3) use the method to analyze wastewater samples containing a range of nitrate and total nitrogen concentrations. Our results indicated that the method needed to be modified to include a longer digestion time (60 min) and dilution of samples prior to digestion (if needed). With the modified method, nitrogen recoveries were not significantly different (P > or = 0.05) from samples with known N concentrations. In addition, nitrate concentrations in constructed wetland and wastewater samples analyzed by both second derivative spectroscopy and ion chromatography were not significantly different. Total nitrogen concentrations in wastewater samples also compared favorably to the same samples analyzed by Kjeldahl digestion. The method is faster, simpler, requires smaller sample volumes, and generates less waste than many EPA-approved methods of N analysis, and may offer a suitable alternative to current methods for analysis of nitrate and total N in wastewater samples. PMID- 11257890 TI - Removal of DDD and DDE from wastewater using bagasse fly ash, a sugar industry waste. AB - Bagasse fly ash, a waste from the sugar industry, was converted into an effective adsorbent and was used for the removal of DDD [2,2-Bis(4-chlorophenyl)-1,1 dichloroethane] and DDE [2,2-Bis(4-chlorophenyl)-1,1-dichloroethene] pesticides from wastewater. The DDD and DDE are removed by the developed adsorbent up to 93% at pH 7.0, with the adsorbent dose of 5 g/l of particle size 200-250 microns at 30 degrees C. The removal of these two pesticides was achieved up to 97-98% in column experiments at a flow rate of 0.5 ml/min. The adsorption was found to be exothermic in nature. The bagasse fly ash system has been used for the removal of DDD and DDE from the wastewater. The developed system is very useful, economic, and reproducible. PMID- 11257891 TI - Effect of detector wavelength on the determination of the molecular weight of humic substances by high-pressure size exclusion chromatography. AB - High-pressure size exclusion chromatography (HPSEC) has proven to be an effective method for determining the molecular weights (MW) of humic substances (HS) from a variety of aquatic and terrestrial environments. HPSEC systems often use a variable wavelength UV-vis detector, which detects the analytes based upon their chromophoric composition. HS contain a range of moieties with chromophores having unique molar absorptivities (for a given wavelength), and the calculated MW may be dependent upon the wavelength chosen for the analysis. As a consequence, the choice of wavelength becomes an important parameter for the reliable determination of MW by HPSEC. The effect of UV-vis detector wavelength on the determination of the MW distribution of selected humic and fulvic acids by HPSEC is examined in this paper. For the HS examined, both the number (Mn) and weight average (Mw) MW increased with increasing wavelength. The relative increase in MW was most pronounced for Lake Fryxell fulvic acid, with a 63 and 21% increase in Mn and Mw, respectively, between 220 and 380 nm. The increases observed for Suwannee River humic and fulvic acids were less pronounced. Mn was more sensitive to changes in detector wavelength than Mw, and as a result the target HS appeared to be less polydisperse at higher wavelengths. Within the range of wavelengths commonly used for the determination of MW of HS by HPSEC (i.e., 220-280 nm), the magnitude of the increases in MW was not significant compared to variability in MW that results from changes to other operational parameters in HPSEC. PMID- 11257892 TI - Inactivation of Cryptosporidium parvum oocysts with ozone and monochloramine at low temperature. AB - The rate of Cryptosporidium parvum inactivation decreased with decreasing temperature (1-20 degrees C) for ozone and for monochloramine applied alone as well as after pre-treatment with ozone. Synergy was observed at all temperatures studied for the ozone/monochloramine sequential disinfection scheme. The synergistic effect was found to increase with decreasing temperature. The inactivation rate with monochloramine after ozone pre-treatment was 5 times faster at 20 degrees C and 22 times faster at 1 degree C than the corresponding post-lag phase rates of inactivation with monochloramine at these temperatures when no ozone pre-treatment was applied. The CT required for achieving 2-logs of inactivation ranged from 11,400 mg min l-1 at 20 degrees C to 64,600 mg min l-1 at 1 degree C when monochloramine was applied alone. In contrast, the CT required for an overall sequential inactivation of 2-logs ranged from 721 mg min l-1 at 20 degrees C to 1350 mg min l-1 at 1 degree C when applying monochloramine after ozone pre-treatment. The presence of excess ammonia in the monochloramine solutions was not responsible for the synergy observed in ozone/monochloramine sequential disinfection. PMID- 11257893 TI - Thermal pyrolysis characteristics of polymer flocculated waste activated sludge. AB - Polyelectrolyte conditioning is a common practice in wastewater management. This paper experimentally elucidated the thermal pyrolysis characteristics of waste activated sludge at a temperature range of 300-900 K (27-627 degrees C) using thermogravimetric analysis (TGA) in inert atmosphere, with especial attention on the effect of polyelectrolyte flocculation (using cationic polyacrylamide). On the pyrolysis rate vs temperature plot two maxima were noted. At the heating rate of 8 degrees C/min, polyelectrolyte does not influence the pyrolysis process. As higher heating rates (14 and 20 degrees C/min), on the other hand, flocculation to charge neutralization point would enhance the rate of thermal pyrolysis. A simple two parallel-reaction kinetic model is applied to interpret the experimental data. Possible roles of flocculant on sludge pyrolysis are discussed on the basis of change in sludge structures and the hindrance of surface reactions of sludge particles. PMID- 11257895 TI - An algal biosensor for the monitoring of water toxicity in estuarine environments. AB - An algal biosensor for toxicity assessment of estuarine waters is proposed. The sensor was obtained by coupling a suited algal bioreceptor (the cyanobacterium Spirulina subsalsa) to an amperometric gas diffusion electrode. The analytical device allows the monitoring of the evolution of photosynthetic O2 and the detection of alterations due to toxic effects caused by environmental pollutants present in the medium. Four chemical species representative of three main different classes of pollutants (heavy metals, triazinic herbicides, carbamate insecticides) were tested at different concentrations using a standardized natural water as experimental medium. In all the cases a toxic response was detected (i.e. a dose-related inhibition of photosynthetic activity was recorded) with good reproducibility. PMID- 11257894 TI - Design parameters for an electrochemical cell with porous electrode to treat metal-ion solution. AB - An electrochemical reactor was designed and operated to treat the solution containing copper ions. Design parameters for the electrochemical reactor using the porous cathode and RuO2/IrO2/Ti anode were investigated. The porous cathode was prepared by the electroless nickel deposition on polyurethane. The apparent current, the gap between cathode and anode, and hydraulic retention time (HRT) were selected as design parameters. The copper removal rate increased with an increase in apparent current. It was not affected by the gap between the cathode and the anode. A reduction in hydraulic retention time stimulated the mass transfer toward the electrode and increased the cathodic current efficiency. Dimensional analysis was conducted to obtain the design equation for scale-up of the electrochemical reactor. The dependence of Sh on Re, Sc, and characteristic lengths, DC/A/L and DW/C/L, was described in the form of a power series. The coefficients were obtained from experimental data. PMID- 11257896 TI - Ultrafiltration of wastewater: effects of particles, mode of operation, and backwash effectiveness. AB - The effects that wastewater quality and mode of operation have on the performance of an asymmetric, hollow fiber, polysulfone, ultrafiltration (UF) membrane with a molecular weight cutoff of 100,000 Daltons were investigated. Performance was assessed through monitoring membrane flux, transmembrane pressure, effluent biochemical oxygen demand, and operational cost of the experimental system while treating filtered secondary, secondary, and filtered primary effluents. Fluxes achieved for filtered secondary (129-173 l/m2 h), secondary (101-158 l/m2 h), and filtered primary (20-41 l/m2 h) effluents were compared to those obtained at three other locations where similar UF systems were operated. A conceptual model of the impact of an insufficient backwash and of operating the UF system at constant flux on membrane performance is presented to explain the differences in fluxes. Employing pre-membrane granular filtration to remove a portion of the problematic particles in secondary effluent prior to UF led to optimal operational conditions. The costs associated with the operation of pre-membrane granular filtration were offset by the increase in production achieved. Although the use of recirculation could increase maintainable flux when treating a concentrated feed (e.g., filtered primary effluent), the associated costs were high. Improved UF performance was found to result from allowing flux to decline naturally, rather than using a constant flux mode of operation. The effluents produced when filtered secondary and secondary effluents were the feeds would be equivalent to an oxidized, coagulated, clarified, and filtered wastewater as per Title 22 California Wastewater Reclamation Criteria. PMID- 11257897 TI - The immunochemical detection of stress proteins in activated sludge exposed to toxic chemicals. AB - The heat shock protein, GroEL, was found to be induced in activated sludge cultures exposed to perturbations of chemicals (cadmium, pentachlorophenol, and acetone) or heat stress. In laboratory activated sludge reactors, GroEL was rapidly induced (within minutes) in the presence of 5 mg/l or greater total cadmium. At 5 mg/l cadmium, however, moderate to insignificant changes in activated sludge process performance indicators [effluent suspended solids concentration, chemical oxygen demand (COD) removal, and specific oxygen uptake rate] were observed. As total cadmium concentrations increased above 5 mg/l, there was a significant and consistent increase in effluent volatile suspended solids concentrations from activated sludge sequencing batch reactors relative to unstressed controls. These results indicate that stress proteins may serve as sensitive and rapid indicators of mixed liquor toxicity which can adversely impact treatment process performance, but that GroEL may not be a good candidate protein for this purpose. PMID- 11257898 TI - The dawning of the Age of Pisces. PMID- 11257899 TI - Down the ataxin-1 track. PMID- 11257900 TI - Out of the loop and on the move. PMID- 11257901 TI - A smoother path to LIS1. PMID- 11257902 TI - Spiny bottlenecks for calcium. PMID- 11257903 TI - Gating by osmosis. PMID- 11257904 TI - Accessory factors in clathrin-dependent synaptic vesicle endocytosis. AB - Clathrin-mediated endocytosis is a special form of vesicle budding important for the internalization of receptors and extracellular ligands, for the recycling of plasma membrane components, and for the retrieval of surface proteins destined for degradation. In nerve terminals, clathrin-mediated endocytosis is crucial for synaptic vesicle recycling. Recent structural studies have provided molecular details of coat assembly. In addition, biochemical and genetic studies have identified numerous accessory proteins that assist the clathrin coat in its function at synapses and in other systems. This review summarizes these advances with a special focus on accessory factors and highlights new aspects of clathrin mediated endocytosis revealed by the study of these factors. PMID- 11257905 TI - Rho GTPases in neuronal morphogenesis. AB - The Rho family of small GTPases act as intracellular molecular switches that transduce signals from extracellular stimuli to the actin cytoskeleton and the nucleus. Recent evidence implicates Rho GTPases in the regulation of neuronal morphogenesis, including migration, polarity, axon growth and guidance, dendrite elaboration and plasticity, and synapse formation. Signalling pathways from membrane receptors to Rho GTPases and from Rho GTPases to the actin cytoskeleton are beginning to be discovered. Mutations in these signalling pathways have been reported in human neurological diseases, which underscores their importance in the development and function of the nervous system. PMID- 11257906 TI - Dendritic integration of excitatory synaptic input. AB - A fundamental function of nerve cells is the transformation of incoming synaptic information into specific patterns of action potential output. An important component of this transformation is synaptic integration--the combination of voltage deflections produced by a myriad of synaptic inputs into a singular change in membrane potential. There are three basic elements involved in integration: the amplitude of the unitary postsynaptic potential; the manner in which non-simultaneous unitary events add in time (temporal summation), and the addition of unitary events occurring simultaneously in separate regions of the dendritic arbor (spatial summation). This review discusses how passive and active dendritic properties, and the functional characteristics of the synapse, shape these three elements of synaptic integration. PMID- 11257907 TI - Neural consequences of environmental enrichment. AB - Neuronal plasticity is a central theme of modern neurobiology, from cellular and molecular mechanisms of synapse formation in Drosophila to behavioural recovery from strokes in elderly humans. Although the methods used to measure plastic responses differ, the stimuli required to elicit plasticity are thought to be activity-dependent. In this article, we focus on the neuronal changes that occur in response to complex stimulation by an enriched environment. We emphasize the behavioural and neurobiological consequences of specific elements of enrichment, especially exercise and learning. PMID- 11257908 TI - Multiple reward signals in the brain. AB - The fundamental biological importance of rewards has created an increasing interest in the neuronal processing of reward information. The suggestion that the mechanisms underlying drug addiction might involve natural reward systems has also stimulated interest. This article focuses on recent neurophysiological studies in primates that have revealed that neurons in a limited number of brain structures carry specific signals about past and future rewards. This research provides the first step towards an understanding of how rewards influence behaviour before they are received and how the brain might use reward information to control learning and goal-directed behaviour. PMID- 11257909 TI - Memory traces revisited. PMID- 11257910 TI - Strengthening the shaky trace through retrieval. PMID- 11257911 TI - Memory involves far more than 'consolidation'. AB - The observation that retrieval returns a stable memory into a labile state cannot be readily explained by any simple version of consolidation theory. This finding has been interpreted as evidence for the need to reconsolidate a memory after reactivating it. However, as we discuss in this commentary, other behavioural observations indicate that even this modification to consolidation theory may be insufficient to describe the dynamic properties of memory. PMID- 11257912 TI - The labile nature of consolidation theory. AB - 'Consolidation' has been used to describe distinct but related processes. In considering the implications of our recent findings on the lability of reactivated fear memories, we view consolidation and reconsolidation in terms of molecular events taking place within neurons as opposed to interactions between brain regions. Our findings open up a new dimension in the study of memory consolidation. We argue that consolidation is not a one-time event, but instead is reiterated with subsequent activation of the memories. PMID- 11257913 TI - Reliability and validity of a psycho-social problem inventory for childhood epilepsy. AB - In response to the need for formal psychosocial assessment in childhood epilepsy, a 25-question inventory was constructed from raw items based on multidisciplinary inputs and tested for practicality, and psychometric attributes. Weighted inter rater and test-retest reliability estimates were 61.8% and 63.9% respectively. A measure of concurrent validity was satisfactory though a modest discrepancy was observed between two raters (P = 0.019 and 0.0064). Discriminant validity was very satisfactory (average P 0.001) based on compared means of ratings from two groups of subjects with disparate prognosis. The inventory should be useful in the assessment of similar chronic childhood disabilities but interpretations of data should be in a wider clinical context including inputs of informant and school authorities as indicated. PMID- 11257914 TI - Noise-induced hearing loss among textile workers in Lagos metropolis. AB - A cross-sectional sample of 204 textile workers was selected randomly from all sections (including the non-production areas), and was initially screened to exclude subjects with pathological middle ear as well as those currently working in sections classified as non-noise but who had been exposed to excessive noise in the past. A total of 17 workers were thus excluded leaving 187 who were then assessed by means of air-conduction and bone-conduction audiograms obtained by the use of a manual (medicor-SA 3L), pure-tone audiometer. All audiometric tests were preceded by an otological examination to rule out the presence of any significant aural pathological conditions. The subjects were divided into 3 groups, based on the noise levels observed at their worksites using a Bruel and Kjaer type 2225 (integrating) sound level meter viz: the noise-exposed group (noise levels > 90 dBA); the less-noise-exposed group (noise levels 85-90 dBA); and the non-noise-exposed group (noise levels < 85 dBA). A comparative analysis of the data on hearing threshold levels of the 3 groups showed that the noise exposed group had significantly (P < 0.05) elevated hearing threshold levels at all frequencies and in each age group, although the maximal threshold shifts were observed at the 4000 Hz frequency. Also, the hearing threshold levels for the noise-exposed group increased with the duration of noise exposure. The study clearly showed the deleterious effects of uncontrolled occupational noise exposure on unprotected workers. A very high prevalence rate of noise-induced hearing loss (79.8%) was recorded for the noise-exposed group. The less-noise exposed group recorded a comparatively low figure of 11.3%. The weaving section alone recorded the highest prevalence rate of 84.5%, followed very closely by the spinning section with 71.0%. The lowest prevalence rate (2.9%) was recorded for the administrative staff. PMID- 11257915 TI - Socio demographic profile and career aspirations of medical students in a new medical school. AB - All fifty-two pioneer medical students in a new medical school (The College of Medicine of Lagos State University), participated in a cross sectional survey to identify their socio-demographic profile and career intentions. A self administered structured questionnaire was used to collect data. The students were observed to be mature. Their mean age was 23.6 years +/- 3.8. A third were graduates with at least a first degree, while 23% had parents with medical background. Factors influencing their choice of medicine as a career were desire to serve mankind (91%), primary interest in the profession (90%) and parental influence (84%). Sixty-seven per cent of them would want to specialize while the most frequently chosen fields were obstetrics and gynaecology (28.6%) and surgery (25.7%). The main reasons influencing specialisation were primary interest in the specialty (91%), service to humanity (91%) and job satisfaction (85.7%). A desire to specialize was evident even in these medical students who were just starting medical school. PMID- 11257916 TI - Experience with ambulatory anaesthesia in gynaecological patients. AB - Ambulatory surgical care is becoming a common feature in Nigeria. Reports on the role of anaesthesia on outcome of surgery in ambulatory setting are rather scanty. In a 24-month period all patients who had gynaecological operations on ambulatory setting were identified and their hospital records reviewed. Sociodemographic characteristics, intraoperative variables, postoperative outcome and postoperative pain management were studied. A total of 28 patients had anaesthesia for various gynaecological procedures. All the patients were in the age range of 20-41 years and American Society of Anaesthesiologists physical status 1. Majority of the patients (71.4%) arrived for day care admission. General anaesthesia with relaxant technique of anaesthesia was the most commonly used technique of anaesthesia (75.0%). Late completion of surgery accounted for 70% (n = 7) of late discharges. Postdural puncture headache (PDPH) was the only adverse outcome seen in our study. Ambulatory anaesthesia is safe, efficient and promising for gynaecological day care procedures. PMID- 11257917 TI - Knowledge of and attitudes to sickle cell disease and sickle carrier screening among new graduates of Nigerian tertiary educational institutions. AB - Six hundred and ten new graduates of Nigerian tertiary institutions were studied for their knowledge of and attitude to sickle cell disorders. A questionnaire was administered to assess knowledge and attitudes. Then a two-hour educational seminar on the basic genetics, transmission, and implications for the affected individuals and their families, of sickle cell disease (SCD) and carrier states was conducted. Sickle carrier screening was undertaken by cellulose acetate haemoglobin electrophoresis. A sickle carrier frequency of 21.6% was found and the questionnaires revealed severely deficient knowledge of the transmission of SCD among the 20-32 year old graduates. After the seminar there was eagerness among the graduates to know their sickle status. It is concluded that unmarried youths in, or graduating from, higher educational institutions may be a most suitable target for information, carrier detection and genetic counselling in the prevention and control of sickle cell disorders. PMID- 11257918 TI - The examination of the jugular venous pressure (JVP): variation of technique amongst Nigerian resident doctors. AB - Resident doctors in Internal Medicine Faculty and General Medical Practice from accredited residency programmes from all over Nigeria were surveyed to investigate the existence of variation in the technique of clinical examination. Using a 10 item self administered questionnaire, the doctors were required to answer questions to test knowledge and skill of examining the jugular venous pressure. Data from 70 Internal Medicine (IMR) and 30 General Medical Practice Residents (GMP) were analysed. For both groups, years of post medical graduation and period spent in residency programme were comparable. On methodology of examination of JVP, there was significant inter- and intra group variation. Sixty per cent (60%) of IMR and only 10% of family practice doctors will use the internal jugular vein alone for assessment, whilst a comparable proportion (30% of IMR and 25% of GMP) will use both internal and external jugular veins. Whilst the sternal angle was the choice of reference point in the majority (87% of IMR versus 80% of GMP), the patient placement angle of 45 degrees was 93% for IMR and 100% for GMP. No respondent chose 60 degrees. Only 40% of both IMR and GMP will always use confirmatory maneuvers, especially the hepatojugular reflux. No GMP doctor and 25% of internal medicine will sometimes use posture and respiratory maneuvers to enhance their clinical decisions. Only 33% IMR and 40% of GMP indicated a correct value and/or unit of measurement for normal jugular venous pressure (JVP). Comparing the internal medicine and General Medical Practice Residents, there was marked inter- and intra-group variation, with more variation noted amongst the IMR. Most importantly, less than 50% of residents knew the correct value of an abnormal JVP. This variability of methodology and poor knowledge will impact on performance in the residency programme, patient care and medical education. PMID- 11257919 TI - Colorectal cancer in Lagos: a critical review of 100 cases. AB - This study critically analysed the clinical presentation, diagnosis, treatment and outcome in 100 patients with colorectal cancer seen over a twelve year period (1988 to 1999). Comparing our findings with those obtained three decades ago at our institution allowed for determination of time trends. There were 48 males and 52 females giving a sex ratio of approximately 1:1. On the average our patients were 10 to 15 years younger than their Caucasian counterparts and one third were 40 years in age or below. In almost two thirds (61%) of patients, the tumours were in the rectum and sigmoid colon. The tumours were resected in 58 (58%) patients, non-resectional surgery was offered to 28 patients, while 14 patients were inoperable, not fit or refused surgery. Overall 34 (34%) patients had distant metastases. The commonest (91.5%) histological type was adenocarcinoma. The postoperative complication rate was 40%, postoperative mortality was 23.3% and the crude one year survival rate was 64%. The clinicopathologic features of colorectal cancer have not changed over a period of three decades at our institution. Public enlightenment campaigns on cancer and facilities to adequately treat patients with colorectal cancer are required in our subregion. PMID- 11257920 TI - Perineal burns from sitz bath in children: case reports. AB - This is a report of 2 children aged < 18 months who sustained perineal and gluteal burns from sitz bath prescribed for fissure-in-ano and perianal pruritus from ascaris infestation respectively at peripheral hospitals. The management included regular cleaning of the burns wounds and open dressing with 1% silver sulphadiazine and administration of broad-spectrum antibiotics and tetanus prophylaxis. This natal cleft was parted regularly to avoid adhesion and anal stenosis. A plea is made for appropriate investigation and treatment of anorectal conditions in children. Arbitrary use of sitz bath is to be discouraged. PMID- 11257922 TI - Oral health status and treatment needs of pregnant women in Lagos State. AB - The oral health status and treatment needs of 250 pregnant women attending the antenatal clinic at Randle Health Centre was investigated. A coded questionnaire was administered to the pregnant women followed by their oral examination in the dental clinic. The mean oral hygiene index score increased progressively throughout pregnancy viz 1st trimester 0.72, second trimester 1.06 and third trimester 1.23. Community Periodontal Index of Treatment Needs (CPITN) revealed that 50% required scale and polish and oral hygiene instruction, 13.60% required oral hygiene instruction only and 32.2% did not require any treatment. Decayed Missing and Filled (DMF) recorded was 1.54. 51.72% of the pregnant women required amalgam fillings, 23.27% required extraction due to caries and 16.38% required partial dentures. PMID- 11257921 TI - Prevalence of hepatitis B surface antigen among patients suspected of liver diseases in a Nigerian hospital. AB - Hepatitis B virus (HBV) infection has been reported to be the aetiological factor for hepatocellular carcinoma (HCC), hepatitis and liver cirrhosis. This study was therefore carried out to determine the seroprevalence of hepatitis B surface antigen (HBsAg), a specific marker of HBV infection in patients with suspected liver diseases. The investigation was carried out among patients attending the University of Maiduguri Teaching Hospital (UMTH), Nigeria with clinical symptoms suggestive of liver diseases and others with non-specific clinical features during the period from 1990-1995. A total of 197 (38%) of 517 patients tested positive for HBsAg, 81 (49%) out of 144 with symptoms suggestive of hepatitis, 75 (50%) of 149 HCC and 10 (56%) of 18 liver cirrhosis were HBsAg positive. Comparison of the yearly total prevalence values of HBsAg for the six years under study showed no significant difference. Similarly, yearly prevalence values amongst patients with suspected liver diseases showed no significant differences. Nevertheless, significant difference, (P < 0.05) between the prevalence of HBsAg among suspected cases of liver diseases and others with non-specific signs and symptoms was observed. Similarly, HBsAg was statistically significantly higher among males than females. The high prevalence rate of HBsAg in our environment may be associated with the suspected cases of liver diseases, which are equally prevalent in our locality. Health education to prohibit traditional practices that could predispose individuals to HBV infections is emphasised. The incorporation of HBV vaccination in the national programme on immunization currently in use in Nigeria is highly recommended. This would be an effective method of preventing HBV infection from childhood. PMID- 11257923 TI - Gene expression analysis as an aid to the identification of drug targets. AB - Many drug targets are components of complex signalling pathways, and in order to understand the true biological consequences of modulating these targets it is necessary to understand the biology of the system in great detail. Genomics research can contribute some of the tools to achieve this, for example through the use of cDNA microarrays. Since activation of signalling pathways leads to mRNA expression, microarray technology can be used to provide a detailed quantitative assessment of the consequences of this activation, often providing a completely new biological perspective on well established cellular systems. This review will discuss some of the results obtained using mRNA profiling of yeast and mammalian cells to analyse signalling pathways of relevance to inflammation and cancer, and will point towards the future applications of this exciting approach to drug target evaluation. PMID- 11257924 TI - Applications of proteomics in oncology. AB - Genomics has expanded the field of molecular oncology, and proteomics is complementing genomics in the fields of elucidation of pathophysiology, gene function, molecular diagnosis and anticancer drug discovery. This trend is reflected in the establishment of the Human Tumour Gene Index by the National Cancer Institute (NCI), which is now followed by the Tissue Proteomics Initiative. Laser capture microdissection (LCM) provides an ideal method for extraction of cells from specimens in which the exact morphologies of both the captured cells and the surrounding tissue are preserved. Proteomic technologies can be applied for the further characterisation and analysis of proteins. LCM can also be combined with the protein chip technology. Proteomic technologies have been used for the study of cancer of various organs including the liver, prostate, breast, bladder and oesophagus. Some of the anticancer strategies are directed against proteases that facilitate several steps in cancer progression. Proteomic mapping of blood vessels in normal and malignant tissues can be used to identify tissue-specific markers on the endothelium that serve as potential targets for in vivo drug delivery. Studies of global protein expression in human tumours have led to the identification of various polypeptide markers, potentially useful as diagnostic tools. Genes that encode proteins that are overexpressed in tumours are being identified. Demonstration of tissue or cell type specific expression of some nuclear matrix proteins has led to the search for tumour specific nuclear matrix proteins. There is considerable activity in the commercial sector to develop diagnostic tests, as well as to facilitate anticancer drug discovery using proteomic technologies. Continued refinement of techniques and methodologies to determine the abundance and status of proteins in vivo holds great promise for future study of normal cells and associated neoplasms. PMID- 11257925 TI - Recent trends in protein biochip technology. AB - This article presents current trends and advances in protein biochip technologies that rely upon extraction and retention of target proteins from liquid media. Analytical strengths as well as technical challenges for these evolving platforms are presented with particular emphasis on selectivity, sensitivity, throughput and utility in the post-genome era. A general review of protein biochip technology is provided, which delineates approaches for protein biochip format and operation, as well as protein detection. A focused discussion of three protein biochip technologies, Biomolecular Interaction Analysis (Biacore, Uppsala, Sweden), Surface Enhanced Laser Desorption/Ionisation (SELDI) ProteinChip Arrays (Ciphergen Biosystems, Fremont, CA, USA) and Fluorescent Planar Wave Guide (Zeptosens, Witterswil, Switzerland), follows along with examples of relevant applications. PMID- 11257926 TI - Progress and potential of Drosophila protein interaction maps. AB - Protein-protein interactions mediate many important cellular processes and are central to the mechanisms by which most proteins function. Charting the interactions among the proteins involved in a process has been an essential step in characterising the function of proteins and pathways. The yeast two-hybrid system is one approach to detecting protein interactions that can now be scaled up to assay large sets of proteins systematically, such as those being identified from genome sequencing efforts. The system has already been extensively used to acquire data that have enabled construction of large protein interaction maps (PIMs). When combined with other data, including data being generated by other functional genomics approaches, PIMs help assign function to new proteins and delineate functional networks. Hypotheses generated in such a manner often must be tested by additional experimentation, preferably in vivo. The model organism Drosophila melanogaster has a wealth of genetic and bioinformatic tools available for such analyses. The proteome predicted from the recently sequenced Drosophila genome indicates that humans have more genes in common with Drosophila than with any other invertebrate model organism characterised to date. Thus, the construction and characterisation of Drosophila PIMs will help define the functions of many conserved genes and pathways, and will provide the pharmaceutical research industry with invaluable data to assist with drug target identification and validation. PMID- 11257927 TI - Transgenic gene knock-outs: functional genomics and therapeutic target selection. AB - The completion of the first draft of the human genome presents both a tremendous opportunity and enormous challenge to the pharmaceutical industry since the whole community, with few exceptions, will soon have access to the same pool of candidate gene sequences from which to select future therapeutic targets. The commercial imperative to select and pursue therapeutically relevant genes from within the overall content of the genome will be particularly intense for those gene families that currently represent the chemically tractable or 'drugable' gene targets. As a consequence the emphasis within exploratory research has shifted towards the evaluation and adoption of technology platforms that can add additional value to the gene selection process, either through functional studies or direct/indirect measures of disease alignment e.g., genetics, differential gene expression, proteomics, tissue distribution, comparative species data etc. The selection of biological targets for the development of potential new medicines relies, in part, on the quality of the in vivo biological data that correlates a particular molecular target with the underlying pathophysiology of a disease. Within the pharmaceutical industry, studies employing transgenic animals and, in particular, animals with specific gene deletions are playing an increasingly important role in the therapeutic target gene selection, drug candidate selection and product development phases of the overall drug discovery process. The potential of phenotypic information from gene knock-outs to contribute to a high-throughput target selection/validation strategy has hitherto been limited by the resources required to rapidly generate and characterise a large number of knock-out transgenics in a timely fashion. The offerings of several companies that provide an opportunity to overcome these hurdles, albeit at a cost, are assessed with respect to the strategic business needs of the pharmaceutical industry. PMID- 11257929 TI - [Message of the editors]. PMID- 11257928 TI - 3D-1D threading methods for protein fold recognition. AB - The threading approach to protein fold recognition attempts to evaluate how well a query sequence fits into an already-solved fold. 3D-1D threaders rely on matching 1-dimensional strings of 3-dimensional information predicted from the query sequence with corresponding features of the target structure. In many cases this is combined with a sequence comparison. The combination of sequence and structure information has been shown to improve the accuracy of fold recognition, relative to the exclusive use of sequence or structure. In this paper, we review progress made since the introduction of threading methods a decade ago, highlighting recent advances. We focus on two emerging methods that are unconventional 3D-1D threaders: proximity correlation matrices and parallel cascade identification. PMID- 11257930 TI - [Saccadic eye movements in extrapyramidal disorders and particularly in Huntington's disease]. PMID- 11257931 TI - MRI atrophy parameters related to cognitive and motor impairment in Parkinson's disease. AB - BACKGROUND: Patients with Parkinson's disease (PD) show specific neuropsychological deficits in attention, memory, visuospatial or frontal lobe functions, which can arise from degeneration of different cerebral structures. OBJECTIVE: The aim of the present study was to analyze the role of focal degeneration (basal ganglia and substantia nigra) and diffuse cerebral atrophy (ventricular enlargement) in motor/cognitive impairment in PD. PATIENTS AND METHODS: We administered to 14 patients with advanced PD the following tests: Purdue Pegboard, Rey's Auditory-Verbal Learning test (RAVLT), Benton's Line Orientation, Trail Making, phonemic verbal fluency and Stroop test. Ventricular system, caudate and putamen nuclei and pars compacta of the substantia nigra were quantitatively measured by magnetic resonance imaging. Correlation analyses were carried out. RESULTS: The results showed that ventricular enlargement is negatively correlated with the performance on RAVLT and Stroop test. No relationship was found between caudate atrophy and cognitive deficits. Degeneration of putamen nucleus was found to be associated with motor deficits. CONCLUSION: Memory and frontal impairment are related to diffuse cerebral degeneration and the motor deficit is related to degeneration of the putamen nucleus. PMID- 11257932 TI - [Speed of ocular saccades in Huntington disease. Prospective study]. AB - BACKGROUND: Oculomotor abnormalities, especially slow saccades, have long been recognized in Huntington's disease (HD). OBJECTIVES AND METHODS: To study prospectively horizontal saccade velocity by videonystagmography in 21 patients with genetically confirmed HD. The study included a baseline analysis and a second evaluation after 18.8 +/- 7.1 months. We included a control group of 15 subjects. RESULTS: HD group exhibited decreased saccade velocity when compared with that from a control group (for predictive and unpredictive target). HD patients showed decreased saccade velocity with the passage of time (for predictive target, p < 0.01). Finally we found statistical significant correlation between saccade velocity and triplet length. CONCLUSIONS: The measurement of saccade velocity might be an objective method to study the natural evolution of HD, and thus evaluate the effectiveness of future therapies. PMID- 11257933 TI - [Stereotaxic radiosurgery. Review of their indications and limitations]. AB - In this review we set up the indications of treatment of vascular malformations (arteriovenous malformations, arteriovenous fistulas and cavernous angiomas), brain tumors (meningiomas, neurinomas, pituitary adenomas, craneofaryngiomas, metastases, gliomas, chondromas, craniopharingiomas and hemangioblastomas) with stereotaxic radiosurgery, and the application of this technique to functional neurosurgery. We assess the limitations of stereotaxic radiosurgery, basically those regarding to volume, proximity to critical anatomical structures and radiosensibility. The review of the literature has allowed us making an approach on efficacy and complications of what with no-doubts we consider an excellent therapeutic alternative. PMID- 11257934 TI - [The Parks-Bielschowsky test. Diagnosis of paresis of the greater oblique muscle]. PMID- 11257935 TI - [Motor impairment, in patients with severe Parkinson's disease, associated with dopaminergic hyperstimulation (entacapone)]. AB - OBJECTIVE: [corrected] Entacapone was given to try to improve the motor complications in eight patients with Parkinson's disease (PD) treated chronically with levodopa, with daily severe motor fluctuations and dyskinesias. PATIENTS AND METHODS: We introduced entacapone (200 mg added to every dose of levodopa) to 8 parkinsonian patients (mean age: 68.25 +/- 2.3; range: 68-72; mean PD duration: 10.4 +/- 2.7 years) treated with oral levodopa, plus a dopa-decarboxylase inhibitor (mean dose: 706.25 +/- 2.3 mg/day; mean period of levodopa-treatment; 9 +/- 2.3 years). Dyskinesias were present in all patients (chorea: 8 patients; "off"--dystonia: 4; byphasic dyskinesias: 3). The type and duration (time "on" and "off") of fluctuations was categorized on the "on-off" charts drawn up by the patients or their relatives, and observation by the investigators after the introduction of entacapone. One patient, with severe impairment with entacapone, was evaluated (motor response) during i.v. apomorphine infusion (40 mg, during 3 hours). RESULTS: The combination of levodopa and entacapone was associated with a net increase in "off" time in all patients (from 5.8 +/- 1.2 h to 12.4 +/- 4.4 h) without change in the URPD. In the patient studied with i.v. apomorphine, "off" periods appeared during the infusion. CONCLUSION: These findings suggest that increased daily levodopa consumption may reduce striatal responsiveness to dopaminergic stimulation in severe parkinsonian patients. These data should be considered when planning the treatment strategy of complex parkinsonian patients. PMID- 11257936 TI - [Progressive encephalomyelitis with rigidity. Clinical and electrophysiological aspects]. AB - A 72-year-old man presented with a chronic illness constituted by muscle rigidity affecting his lower limbs, trunk and neck, either spontaneous or triggered by stimuli, together with a spastic paraparesis, manual amyotrophy and pseudobulbar syndrome. The electrophysiologic study showed continuous motor unit activity integrated by normal motor unit potentials. Biochemical and imaging results were normal. These data suggest the diagnosis of idiopathic progressive encephalomyelitis with rigidity. Following administration of gabapentin (2000 mg daily), muscle rigidity and electromyographic continuous motor unit activity were suppressed. Transient drowsiness was the only side effect. The authors have tried to relate these findings to those found in the literature. PMID- 11257937 TI - Focal stiff-person syndrome. AB - Stiff-person syndrome (SPS) is a disorder of motor function characterized by rigidity of axial musculature and fluctuating painful spasms, which are often induced by startle or emotional stimuli. Neurophysiological studies have demonstrated the presence of continuous motor unit activity in muscle at rest, with abnormally enhanced extereoceptive reflexes. Although criteria for the diagnosis of SPS were proposed, several variants of this syndrome have been described before. In this communication, we report the case of a patient with a focal form of SPS. A 39-year-old woman developed progressive instability in her gait, spasms and stiffness restricted to both legs. The electromyographic examination showed continuous motor unit activity of the affected muscles at rest. Moreover, high anti-GAD antibodies titers were found in CSF and serum. Clinical symptoms, electrophysiological and immunological profiles suggest a focal form of SPS. Clinical and immunological findings indicate that SPS is a heterogeneous disease, suggesting the need to redefine its diagnostic criteria. Definition of the range of clinical expression and immunological profiles could be important for the clinical management of these patients. PMID- 11257938 TI - [Sleep attacks and pergolide mesylate]. PMID- 11257940 TI - The management of pediatric neuropsychiatric disorders and the human genome. PMID- 11257939 TI - [The future of neurology and neurologic health care in the public Spanish framework]. PMID- 11257941 TI - The "medicalization" of pediatric mental illness. PMID- 11257942 TI - Childhood depression: diagnosis and treatment strategies in general pediatrics. PMID- 11257943 TI - Antipsychotic medications for children and adolescents. PMID- 11257944 TI - Medication for the management of anxiety disorders in children and adolescents. PMID- 11257945 TI - Obsessive-compulsive disorder and related conditions. PMID- 11257946 TI - An update on assessment and treatment of complex attention-deficit hyperactivity disorder. PMID- 11257948 TI - [Do oral free communications or posters have a different scientific value in international congresses of anesthesiology]. PMID- 11257947 TI - Resident's column: searching for answers to attention-deficit hyperactivity disorder. PMID- 11257949 TI - [Intraoperative rupture of the laryngeal mask]. PMID- 11257950 TI - [Apneic oxygenation: a new tool in difficult intubations?]. PMID- 11257951 TI - [Multicenter study on the usefulness of the NasOral system for the denitrogenation and apneic oxygenation in anesthesia]. AB - OBJECTIVES: To study the usefulness of the NasOral system for denitrogenation prior to anesthetic induction for improving pulmonary oxygen storage that maintains SpO2 within the normal range during induced apnea and facilitates apneic oxygenation. MATERIAL AND METHODS: To establish the study population of 125, five hospitals of the Valencian Community (Spain) enrolled patients scheduled for elective procedures under general anesthesia. The patients were preoxygenated using the NasOral system (denitrogenation). For two minutes, the patients inhaled oxygen through the nose (FiO2 1) at a flow rate of 8-10 l/min (never less than the patient's own minute volume) and exhaled orally through a unidirectional valve. We measured time of apnea with SpO2 > or = 96% to assess the usefulness of the device for denitrogenation. We also measured PetCO2 after endotracheal intubation and after maximum time of apnea (< or = 10 minutes) to assess use of the device for apneic oxygenation during laryngoscopy. RESULTS: We found no significant differences with regard to age, sex, ASA or Mallampati classification among patient groups enrolled at the participating hospitals. Cox's regression analysis was used to determine relative risk of SpO2 < 96%. At 10 minutes post-apnea, 88.8% of all patients maintained SpO2 > or = 96%. However, SpO2 > 96% was maintained by 94.1% in the Mallampati I group and by 84.1% in the Mallampati II group. SpO2 fell below the cut-off (< 96%) in 33.3% of obese patients and in 7.5% of non-obese patients. Analysis of the likelihood of SpO2 < 96% associated with the variables of obesity, sex, age, ASA and Mallampati classification was significant only for obesity, for which a risk of 1.95 was calculated relative to non-obesity (95% CI 1.14-3.35). The NasOral system allows performance of direct laryngoscopy for oral tracheal intubation, maintaining oxygen flow through the permeable airway to facilitate apneic oxygenation. CONCLUSIONS: The NasOral system facilitates denitrogenation before induction of anesthesia in all patients with permeable nasal fossae as well as apneic oxygenation during laryngoscopy. PMID- 11257952 TI - [Bupivacaine in continuous epidural infusion using a portable mechanical devise for postoperative analgesia after surgery for hernia of the lumbar disk]. AB - OBJECTIVES: To determine the optimum concentration of bupivacaine administered by continuous epidural infusion, using a Baxter Single Day elastomeric infusor at a rate of 2 ml/h, to treat pain during the first 24 h after lumbar laminectomy. PATIENTS AND METHODS: Sixty patients undergoing elective repair of a herniated lumbar disk were randomly assigned to three homogeneous groups of 20 each. Group I received epidural infusion of 0.0625% bupivacaine, group II received 0.125% bupivacaine and group III received 0.25% bupivacaine. After surgery each patient was given a 4 mL solution of the local anesthetic being studied, followed by an infusion of the same through an elastomeric infusor at a rate of 2 ml/h throughout the first 24 h after surgery. Ketorolac was delivered through a device for patient controlled analgesia after surgery. Pain was assessed on a visual analog scale (VAS) at rest and during movement. Pain relief was assessed on a simple descriptive scale. RESULTS: Significantly less ketorolac was required during epidural infusion of 0.125% and 0.25% bupivacaine than when the 0.0625% concentration was being infused (29 +/- 16 and 28 +/- 13 mg, respectively, versus 110 +/- 35 mg; p < 0.001). VAS scores were significantly lower during infusion of 0.125% and 0.25% bupivacaine than with 0.0625% bupivacaine. No instances of motor blockade or infection related to catheter insertion were observed in any of the patients. CONCLUSIONS: Continuous epidural infusion of 0.125% and 0.25% bupivacaine through an elastomeric infusor gives excellent analgesia during the first 24 h after surgery. Administration of 0.25% bupivacaine is associated with a higher incidence of urinary retention. We therefore think that the most recommendable concentration of bupivacaine for infusion is 0.125%. PMID- 11257953 TI - [Prevention of postoperative nausea and vomiting in gynecologic surgery with 3 fixed doses of metoclopramide, droperidol or placebo]. AB - OBJECTIVES: To compare the efficacy and side effects of three doses of metoclopramide, droperidol or placebo administered every 8 h to prevent nausea and vomiting during the first 24 h after surgery. MATERIAL AND METHODS: Prospective, double blind study of 104 patients scheduled for major intraabdominal gynecological surgery under general anesthesia. The patients were randomly assigned to three groups: group M received 10 mg of metoclopramide, group D received 1.25 mg of droperidol and group P received a saline solution. The patients were premedicated with oral diazepam. All patients were anesthetized using similar techniques, with fentanyl, thiopental, vecuronium, oxygen/nitrogen protoxide and isoflurane. Muscle relaxation was reversed with atropine and neostigmine. Postoperative analgesia was given with endovenous morphine and metamizol. Immediately after surgery each patient received an endovenous dose of the assigned antiemetic drug. Patients were monitored for 24 h and observations were recorded every hour on the following scale: 0, for no emetic symptoms, 1 for nausea and 2 for vomiting. RESULTS: Fifteen patients (42.9%) in group D, 21 (60% in group M and 19 (54.3%) in group P experienced nausea during the 24 h after surgery, with no significant differences. However, the incidence of vomiting was significantly lower in group D, with 7 patients (20%) vomiting in group D versus 11 patients (31.43%) in group M and 17 (50%) in group P. Side effects were mild and required no treatment. CONCLUSIONS: Droperidol at a dose of 1.25 mg every 8 h is effective and safe for preventing postoperative nausea and vomiting and has minimal side effects. Metoclopramide at a dose of 10 mg every 8 h, in our study, was no better for the same purpose than placebo. PMID- 11257954 TI - [General considerations for the reform of the anesthesiology and critical care training system in Spain]. PMID- 11257955 TI - [Intravenous nicardipine: a new calcium antagonist for perioperative use]. AB - Nicardipine is a calcium antagonist belonging to the dihydropyridine group of molecules. An intravenous formulation has recently become available, offering certain advantages over other drugs in this group, given that it lacks their negative inotropic effects and its vasodilator effect favors coronary and cerebral artery beds. Nicardipine lowers arterial pressure in proportion to plasma levels by means of a reduction in systemic vascular resistance produced by relaxation of arteriolar smooth muscle. It has been suggested that nicardipine has a protective effect on the myocardium. PMID- 11257956 TI - [Malignant hyperthermia in a pig anesthesized with desflurane]. AB - A large-white pig that had not been genetically selected to develop malignant hyperthermia (MH) during anesthesia nevertheless suffered an episode of severe MH after repeated exposure to increasing concentrations of desflurane. MH is a hypermetabolic alteration that may develop in susceptible patients who have inhaled certain drugs or agents that act as triggers. Early identification and appropriate treatment are essential to reduce the likelihood of death associated with this severe alteration. We report a case of late-developing MH triggered by low concentrations of inhaled desflurane. PMID- 11257957 TI - [Fastrach laryngeal mask, sevoflurane and remifentanil: an anesthetic alternative for the myasthenic patient]. AB - A 46-year-old myasthenic man diagnosed two months earlier and experiencing nocturnal dyspnea was scheduled for transsternal thymectomy. The patient was premedicated with midazolam in the operating room. Anesthetic induction and maintenance were with inhaled sevoflurane and an intravenous infusion of remifentanil, with no need for neuromuscular relaxants. Airway management was achieved by inserting a Fastrach laryngeal mask (LM-Fastrach), through which an endotracheal tube could be inserted easily. The tube was withdrawn through the mask at the end of surgery and the mask was removed in the operating room 6 minutes later. Anesthesia in patients with myasthenia gravis is one of the greatest challenges in clinical anesthesiology. The interest of this case lies mainly in that the anesthetic technique chosen allows neuromuscular relaxants to be avoided. Moreover, airway access through the Fastrach laryngeal mask is highly useful for transsternal thymectomy of the patient with myasthenia gravis, providing immobility and adequate hemodynamic stability during sternotomy as well as facilitating safe and rapid postanesthetic recovery. PMID- 11257958 TI - [Paraplegia after left thoracotomy in a patient carrying an epidural catheter]. AB - A 61-year-old man undergoing left thoracotomy, in whom a lumbar epidural catheter had been inserted, experienced paraplegia in the immediate recovery period. Diagnostic tests and clinical examinations ruled out neurological causes attributable to the catheter, indicating that the cause was surgical. Paraplegia after thoracotomy is rare but has been described in the literature. We discuss ways that injuries may produce medullary lesions related to thoracic surgery. PMID- 11257959 TI - [Pregnancy and partum in the woman with a spinal cord lesion in chronic phase]. AB - A 34-year-old pregnant paraplegic woman with a T12 medullary lesion in chronic phase underwent cesarean delivery in the thirty-seventh week due to pelvic cephalic disproportion. After failure of epidural anesthesia related to technical difficulties, general anesthesia was provided. A hypertensive crisis developed during surgery but was resolved within minutes after administration of hydralazine. No further complications arose. Pregnancy in a patient with medullary lesion in chronic phase is considered high risk, requiring special care due to extraordinary changes in pathophysiology caused by the lesion in addition to changes directly related to gestation. The main complications that arise are decreased respiratory volume and arterial pressure, increased incidence of thromboembolic events, anemia, urinary tract infections, premature birth, unusual progression of delivery and autonomic hyperreflexia, which is the most serious. When a hypertensive peak develops in such patients, the anesthesiologist must first rule out autonomic hyperreflexia, which has an incidence of 85% in lesions over T7 and has also been described in patients with lower lesions. PMID- 11257960 TI - [Fiberoptic bronchoscopy through the laryngeal mask in a patient with Goldenhar syndrome]. PMID- 11257961 TI - [Persistent left superior vena cava: an infrequent localization of the central venous catheter]. PMID- 11257962 TI - [Anesthesia in a case of Bardet-Biedl syndrome]. PMID- 11257963 TI - Public perception of science. Part 2. PMID- 11257964 TI - Nonhomogeneous resolution of images of natural scenes. AB - The aim of this research is to model and simulate the loss of visual resolution as a function of retinal eccentricity in the perception of natural scenes. The model of visual resolution is based on a space-variant low-pass filter, having a variable convolution kernel according to retinal eccentricity. The parameters of the model are computed from psychophysical measures of visual acuity as a function of retinal eccentricity. The implementation of the model allowed us to generate images of scenes with nonhomogeneous space-variant resolution, simulating the filtering executed by the eye. These scenes are used to test and optimise the model by means of experiments in static vision (through tachistoscopic presentations) and in dynamic vision where the resolution of the scene is computed, in real-time, as a function of the location of gaze. PMID- 11257965 TI - Testing the aesthetic significance of the golden-section rectangle. AB - The aesthetic significance of the golden-section rectangle was tested in two studies designed to obviate some of the criticisms of earlier experiments. In the first, employing the method of use, the mean sides-ratios of samples of paintings from five subject-matter categories (landscape, still life, head-and-shoulders portrait, upper-body portrait, full-length portrait) provided no evidence for the significance of the golden section. However, the sides ratio of portraits varied between categories in ways that were consistent with the requirements of the proportions of the subject matter. In the second study, using the method of production, participants produced the most pleasing four-sided shape, under four instruction conditions. Under a 'portrait painting' condition and a 'landscape painting' condition, the mean sides-ratios differed significantly from the golden section. Under two 'context free' geometric shape conditions--horizontal rectangle and vertical rectangle--the mean sides-ratio approximated the golden section. The results are discussed in terms of the methodological requirements for a valid test of the aesthetic significance of the golden section and the possibility that this ratio may indeed have special significance. PMID- 11257966 TI - Velocity constancy in a virtual reality environment. AB - During everyday life the brain is continuously integrating multiple perceptual cues in order to allow us to make decisions and to guide our actions. In this study we have used a simulated (virtual reality--VR) visual environment to investigate how cues to speed judgments are integrated. There are two sources that could be used to provide signals for velocity constancy: temporal-frequency or distance cues. However, evidence from most psychophysical studies favours temporal-frequency cues. Here we report that two depth cues that provide a relative object--object distance--disparity and motion parallax--can provide a significant input to velocity-constancy judgments, particularly when combined. This result indicates that the second mechanism can also play a significant role in generating velocity constancy. Furthermore, we show that cognitive factors, such as familiar size, can influence the perception of object speed. The results suggest that both low-level cues to spatiotemporal structure and depth, and high level cues, such as object familiarity, are integrated by the brain during velocity estimation in real-world viewing. PMID- 11257967 TI - On binocular alternation. AB - Diaz-Caneja (1928) made some prescient observations about binocular rivalry. Being in French, however, his paper remained largely unknown to the broader research community. His findings are similar to those reported very recently by contemporary researchers who had independently observed similar phenomena. Using concentric circles and parallel lines as stimuli, Diaz-Caneja presented half of each form to opposite eyes to provoke binocular rivalry. He observed periods in the ensuing binocular alternations in which rivalry occurred between the good Gestalt forms, despite the fact that they were distributed between the eyes. He proposed that each half of a good form generates synchronised oscillations in the visual system, and that this synchronisation enables the dichoptically viewed halves of the one form to be perceived as a whole. PMID- 11257968 TI - An analogy between colour and spatial coding. AB - The early stages of colour coding are well established in that the trichromatic receptor stage is followed by a set of opponent colour channels. One interpretation of the sequence is that opponent channels carry unrelated aspects of the colour stimulus, unlike the cone channels. The overlap of the cone channels can be removed by decorrelating their spectral-sensitivity functions, and this procedure has been found to give opponent colour channels which match those found psychophysically. Since the known spatial-frequency channels also show considerable overlap, the question arises which aspects of the spatial stimulus are captured by decorrelating the spatial-frequency channels. The results of decorrelating the spatial-frequency channels are that the first decorrelated spatial filter acts as a broad bandpass filter which has a peak sensitivity at 7.9 cycles deg-1, and that the second decorrelated spatial filter acts as an opponent spatial-frequency channel, with a minimum output at a low (4.1 cycles deg-1) spatial frequency and a maximum output at a high (15.1 cycles deg-1) spatial frequency. The characteristics of the first decorrelated filter closely resemble the properties of the foveal perceptive field which have been used to explain the Hermann grid illusion. Thus, the decorrelation analysis produces a model for the functional organisation of the channel implementation at the neural and psychophysical levels, but which directly relates to the subjective appearance of the visual stimuli. PMID- 11257969 TI - Imposed vibration influences perceived tactile smoothness. AB - According to the duplex theory of tactile texture perception, detection of cutaneous vibrations produced when the exploring finger moves across a surface contributes importantly to the perception of fine textures. If this is true, a vibrating surface should feel different from a stationary one. To test this prediction, experiments were conducted in which subjects examined two identical surfaces, one of which was surreptitiously made to vibrate, and judged which of the two was smoother. In experiment 1, the vibrating surface was less and less often judged smoother as the amplitude of (150 Hz) vibration increased. The effect was comparable in subjects who realized the surface was vibrating and those who did not. Experiment 2 showed that different frequencies (150-400 Hz) were equally effective in eliciting the effect when equated in sensation level (dB SL). The results suggest that vibrotaction contributes to texture perception, and that, at least within the Pacinian channel, it does so by means of an intensity code. PMID- 11257970 TI - Large systematic deviations in visual parallelism. AB - The visual environment is distorted with respect to the physical environment. Luneburg [1947, Mathematical Analysis of Binocular Vision (Princeton, NJ: Princeton University Press)] assumed that visual space could be described by a Riemannian space of constant curvature. Such a space is described by a metric which defines the distance between any two points. It is uncertain, however, whether such a metric description is valid. Two experiments are reported in which subjects were asked to set two bars parallel to each other in a horizontal plane. The backdrop consisted of wrinkled black plastic sheeting, and the floor and ceiling were hidden by means of a horizontal aperture restricting the visual field of the subject vertically to 10 deg. We found that large deviations (of up to 40 degrees) occur and that the deviations are proportional to the separation angle: on average, the proportion is 30%. These deviations occur for 30 degrees, 60 degrees, 120 degrees, and 150 degrees reference orientations, but not for 0 degree and 90 degrees reference orientations; there the deviation is approximately 0 degree for most subjects. A Riemannian space of constant curvature, therefore, cannot be an adequate description. If it were, then the deviation between the orientation of the test and the reference bar would be independent of the reference orientation. Furthermore, we found that the results are independent of the distance of the bars from the subject, which suggests either that visual space has a zero mean curvature, or that the parallelity task is essentially a monocular task. The fact that the deviations vanish for a 0 degree and 90 degrees orientation is reminiscent of the oblique effect reported in the literature. However, the 'oblique effect' reported here takes place in a horizontal plane at eye height, not in a frontoparallel plane. PMID- 11257971 TI - Perception of partly occluded figures by baboons (Papio papio). AB - Comparative literature provides conflicting findings whether animals experience amodal completion. Five experiments were conducted to verify if baboons perceive partly occluded objects as complete. The first three experiments used a go/no-go procedure and a video monitor for stimulus presentation. These experiments failed to reveal amodal completion, suggesting that the stimuli were processed as 2-D images rather than 3-D objects. In contrast, completion was demonstrated in a fourth experiment with cardboard stimuli in a two-alternative forced-choice (2AFC) discrimination task presented in a Wisconsin General Test Apparatus. Although in experiment 5 the same 2AFC procedure was used as in experiment 4, completion was absent when the stimuli were shown with a computer graphic system. The results suggest that baboons share with humans the ability for amodal completion, but also underline some procedural factors that might affect the elicitation of this capacity. PMID- 11257973 TI - Multiple visual worlds. PMID- 11257972 TI - Last but not least. Positive scotomata of the blind spots during abrupt changes between darkness and twilight. PMID- 11257974 TI - Distance perception within near visual space. AB - We investigated the perception of distance of visual targets with constant size and luminance presented between 20 and 120 cm from subjects' eyes. When retinal disparity cues were present, the subjects could reproduce very accurately the distance of a seen reference in this area. When only extraretinal information was available, distance perception was still correct for distances of 40 cm or less. However, distances beyond 60 cm were underestimated. When forced to evaluate the distance between a reference and themselves, e.g. when evaluating the absolute distance or half the distance or twice the distance of a reference, subjects used an egocentric plane of reference located on average 10.4 cm in front of their eyes. Measurements of binocular eye movements indicated a clear relationship between vergence angle and target distance. The egocentric plane of reference at 10.4 cm also corresponds to the maximum achievable vergence. These results suggest that ocular convergence can be used as a reliable cue for distance within the arm's reaching space. PMID- 11257975 TI - The pew illusion--a real-world example of misperceived slant. PMID- 11257976 TI - Efficient search for size targets on a background texture gradient: is detection guided by discontinuities in the retinal-size gradient of items? AB - Six visual search experiments were carried out to investigate the processing of size information in early vision. The apparent size of display items was manipulated independently of their retinal size by placing items on a textured surface which altered the perceived distance in depth of the items. Overall, these experiments demonstrate that a target item differing from non-target items in terms of apparent size can be detected efficiently. However, the pattern of results indicates that, rather than deriving apparent-size information, target detection is guided by discontinuities in the 'retinal-size gradient' of items, in particular between items at the same 'depth'. Although the arrangement of items on the texture surface strongly influenced search, this was largely due to the retinal size of items and the retinal separation between items. The implications of these experiments for the nature of the pre-attentive representation of size are discussed. PMID- 11257977 TI - Spatially parallel processing of within-dimension conjunctions. AB - Within-dimension conjunction search for red-green targets amongst red-blue, and blue-green, nontargets is extremely inefficient (Wolfe et al, 1990 Journal of Experimental Psychology: Human Perception and Performance 16 879-892). We tested whether pairs of red-green conjunction targets can nevertheless be processed spatially in parallel. Participants made speeded detection responses whenever a red-green target was present. Across trials where a second identical target was present, the distribution of detection times was compatible with the assumption that targets were processed in parallel (Miller, 1982 Cognitive Psychology 14 247 279). We show that this was not an artifact of response-competition or feature based processing. We suggest that within-dimension conjunctions can be processed spatially in parallel. Visual search for such items may be inefficient owing to within-dimension grouping between items. PMID- 11257978 TI - Brain areas sensitive to coherent visual motion. AB - Detection of coherent motion versus noise is widely used as a measure of global visual-motion processing. To localise the human brain mechanisms involved in this performance, functional magnetic resonance imaging (fMRI) was used to compare brain activation during viewing of coherently moving random dots with that during viewing spatially and temporally comparable dynamic noise. Rates of reversal of coherent motion and coherent-motion velocities (5 versus 20 deg s-1) were also compared. Differences in local activation between conditions were analysed by statistical parametric mapping. Greater activation by coherent motion compared to noise was found in V5 and putative V3A, but not in V1. In addition there were foci of activation on the occipital ventral surface, the intraparietal sulcus, and superior temporal sulcus. Thus, coherent-motion information has distinctive effects in a number of extrastriate visual brain areas. The rate of motion reversal showed only weak effects in motion-sensitive areas. V1 was better activated by noise than by coherent motion, possibly reflecting activation of neurons with a wider range of motion selectivities. This activation was at a more anterior location in the comparison of noise with the faster velocity, suggesting that 20 deg s-1 is beyond the velocity range of the V1 representation of central visual field. These results support the use of motion-coherence tests for extrastriate as opposed to V1 function. However, sensitivity to motion coherence is not confined to V5, and may extend beyond the classically defined dorsal stream. PMID- 11257979 TI - Configural features in the context of upright and inverted faces. AB - When faces are turned upside down, recognition is known to be severely disrupted. This effect is thought to be due to disruption of configural processing. Recently, Leder and Bruce (2000, Quarterly Journal of Experimental Psychology A 53 513-536) argued that configural information in face processing consists at least partly of locally processed relations between facial elements. In three experiments we investigated whether a local relational feature (the interocular distance) is processed differently in upside-down versus upright faces. In experiment 1 participants decided in which of two sequentially presented photographic faces the interocular distance was larger. The decision was more difficult in upside-down presentation. Three different conditions were used in experiment 2 to investigate whether this deficit depends upon parts of the face beyond the eyes themselves; displays showed the eye region alone, the eyes and nose, or the eyes and nose and mouth. The availability of additional features did not interact with the inversion effect which was observed strongly even when the eyes were shown in isolation. In experiment 3 all eyes were turned upside down in the inverted face condition as in the Thatcher illusion (Thompson, 1980 Perception 9 483-484). In this case no inversion effect was found. These results are in accordance with an explanation of the face-inversion effect in which the disruption of configural facial information plays the critical role in memory for faces, and in which configural information corresponds to spatial information that is processed in a way which is sensitive to local properties of the facial features involved. PMID- 11257980 TI - Developmental changes in the effect of inversion: using a picture book to investigate face recognition. AB - A novel child-oriented procedure was used to examine the face-recognition abilities of children as young as 2 years. A recognition task was embedded in a picture book containing a story about two boys and a witch. The story and the task were designed to be entertaining for children of a wide age range. In eight trials, the children were asked to pick out one of the boys from amongst eight distractors as quickly as possible. Response-time data to both upright and inverted conditions were analysed. The results revealed that children aged 6 years onwards showed the classic inversion effect. By contrast, the youngest children, aged 2 to 4 years, were faster at recognising the target face in the inverted condition than in the upright condition. Several possible explanations for this 'inverted inversion effect' are discussed. PMID- 11257981 TI - Features derived from first-order motion mechanisms predict anomalies in motion perception. AB - Current dominant hypotheses of how humans detect the movement of patterns assume that the pattern is divided into one-dimensional sinusoidally varying luminance patterns, referred to as gratings (first-order components). The speed of these gratings is independently encoded from predominantly spatial and temporal frequency information, and their direction is encoded from orientation information. This paper addresses the problem of how the individually encoded grating information is combined to give perceived pattern direction, given that real moving objects are generally made up of more than one component. More specifically, further evidence is presented for a combination based on the use of a feature derived from first-order components--'first-order feature hypothesis'. This hypothesis essentially implements a constraint on pattern direction called the intersection of constraints (IOC) proposed by Adelson and Movshon [1982, Nature 300 523-525]. A simulation of the model is used to make three new predictions about a perceived motion reversal reported by Derrington et al (1992, Vision Research 32 699-707); these predictions are tested and found to be consistent with the first-order feature hypothesis. PMID- 11257982 TI - Perceptual learning without feedback and the stability of stereoscopic slant estimation. AB - Subjects were examined for practice effects in a stereoscopic slant-estimation task involving surfaces that comprised a large portion of the visual field. In most subjects slant estimation was significantly affected by practice, but only when an isolated surface (an absolute disparity gradient) was present in the visual field. When a second, unslanted, surface was visible (providing a second disparity gradient and thereby also a relative disparity gradient) none of the subjects exhibited practice effects. Apparently, stereoscopic slant estimation is more robust or stable over time in the presence of a second surface than in its absence. In order to relate the practice effects, which occurred without feedback, to perceptual learning, results are interpreted within a cue interaction framework. In this paradigm the contribution of a cue depends on its reliability. It is suggested that normally absolute disparity gradients contribute relatively little to perceived slant and that subjects learn to increase this contribution by utilizing proprioceptive information. It is argued that--given the limited computational power of the brain--a relatively small contribution of absolute disparity gradients in perceived slant enhances the stability of stereoscopic slant perception. PMID- 11257983 TI - How Canadian legislators view health promotion: does party affiliation matter? PMID- 11257985 TI - Alcohol-related policy measures in Ontario: who supports what and to what degree? AB - Using 1998 provincial survey data (n = 1,205), the authors examine responses to 7 items concerning public opinion on alcohol-related policy in Ontario. The purpose of the study is to get a sense of overall public opinion on certain topical policy-related measures and to see whether this opinion is predicted by demographic characteristics of respondents (sex, age and self-reported drinking pattern). Cross-tabulations of opinion items with demographic variables revealed strong majority support for the status quo with regard to number of liquor and beer stores, beer and liquor store hours, and prohibition of the sale of alcohol in corner stores. A somewhat less robust majority also supported the status quo for alcohol taxes and legal drinking age. Among the demographic groups, high-risk heavy drinkers stood out for their greater support of relaxation of controls and this finding was confirmed by means of logistic regression. The majority of all groups, except frequent bar-goers, liked the idea of warning labels on alcoholic beverage containers. The authors conclude that, according to these survey data, policy initiatives towards greater access to alcohol, such as extended liquor store hours and sale of alcohol in corner stores, are not mandated by the majority of the population of Ontario. PMID- 11257984 TI - The use of population health and health promotion research by health regions in Canada. AB - This study examined the use of population health and health promotion (PH&HP) research by health regions in Canada. An 11-item survey was faxed to 137 (of 140) health regions. Eighty-three completed questionnaires were returned (60.8%). Results indicate that while research, in general, plays more than a moderate role in the majority of participating health regions, PH&HP research is not used frequently. The most frequent uses of PH&HP research include the development of health goals and objectives, the development of programs and services, and resource allocation. Health regions most frequently obtain PH&HP research from their own staff and from government departments. University-based researchers are not a commonly used source. This study provides a descriptive overview of health regions' engagement in evidence-based decision making related to PH&HP issues, and points to a number of strategies that both health regions and researchers can employ to enhance the use of PH&HP research by health regions. PMID- 11257986 TI - Assessing the efficacy of a school-based asthma education program for children: a pilot study. AB - BACKGROUND: Asthma diminishes the health-related quality of life for many school aged children. This study sought to explore the effect of a School-Based Asthma Education Program (SBAEP) on quality of life. METHODS: Children with asthma who attended grades 1-5 at two selected schools were requested to participate in this pilot study. Participants at one school were provided with a SBAEP, those at another school (control group) were provided with written educational material about asthma. The children completed the Paediatric Asthma Quality of Life Questionnaire (PAQLQ) before and one month after the educational interventions. RESULTS: There were clinically important improvements in the SBAEP group in quality of life, specifically in the symptom subdomain. CONCLUSIONS: The "Air Force" SBAEP appears to result in a favourable trend in quality of life for children. A larger scale trial is required following revisions to the program. PMID- 11257988 TI - Tobacco industry litigation and the role of government: a public health perspective. PMID- 11257987 TI - Completing the picture: adolescents talk about what's missing in sexual health services. AB - This qualitative study was conducted to learn adolescents' opinions about sexual health services and strategies to improve their delivery. Sixteen 1.5-hour, same sex focus groups were conducted in one rural and one urban high school in each of two Ontario regions. In total, 83 students (49 females and 34 males) participated in the study. Topics were: sources and quality of sexual health information, knowledge and use of sexual health services, gender differences, factors that influence sexual behaviour, and suggestions for improving sexual health services. The adolescents reported that sex education focussed too much on "plumbing" and was often provided by teachers with whom they felt uncomfortable discussing sexual issues. Peers and media were their main sources of information although they acknowledged that these were not always accurate. The participants had limited knowledge of the services available. Many of their comments reflected traditional gender differences. Peers, and for females, partners and parents influenced sexual decision-making. The participants made numerous suggestions for improving sexual health services. PMID- 11257989 TI - Mothers' perceptions of childhood immunizations in First Nations communities of the Sioux Lookout Zone. AB - OBJECTIVE: Low uptake of childhood immunizations is a problem in many First Nations communities. This article describes the results of a study that examined mothers' perceptions of childhood immunizations and the factors that influence uptake. METHOD: Person-centred interviews focussing on childhood immunizations and child health were conducted with 28 mothers of young children in two First Nations communities in the Sioux Lookout Zone. Content analysis was applied to the interview data and patterns and themes were developed. RESULTS: Data analysis identified four key factors as negatively influencing immunization uptake: knowledge barriers, the influence of others, vaccine barriers, and missed opportunities. CONCLUSIONS: Further research with Elders and community members along with culturally sensitive education initiatives are required to address low immunization uptake. Changes in health professionals' behaviours may serve to reduce missed opportunities. PMID- 11257990 TI - Mothers' efforts to protect their infants from environmental tobacco smoke. PMID- 11257991 TI - Cancer prevention in the community: a survey of community residents. AB - Lifestyle exposures account for the greatest proportion of risk factors for cancer, yet these exposures have proven most difficult to alter. Despite intensive intervention efforts, many behaviour change programs are ill suited to the community. This research was undertaken to increase our understanding of prevention activities of interest to a sample of residents in two Ontario communities. 248 (62.3%) adult residents responded to a semi-structured self administered questionnaire including open-ended questions on health issues, exposures and prevention activities of interest. While some of the beliefs expressed by respondents might have been anticipated (e.g., cigarette smoking and family history increase risk of cancer), others were not (e.g., only between 40 and 75% of respondents thought a high fat diet increased risk). Furthermore, many of those with personal health concerns expressed an interest in prevention. This process is proposed as a first step in launching more appropriate and sustainable community-based health promotion programs for cancer prevention. PMID- 11257992 TI - The new international trade regime: problems for publicly funded healthcare in Canada? PMID- 11257993 TI - Three top Canadian and personal health concerns of a random sample of Nova Scotian women. AB - We sought to understand the subjective reports of women's health concerns. A randomly dialled telephone survey was conducted resulting in a sample of 458 women (Caucasian/European = 302, Native/Aboriginal = 81, Black = 75), aged 18-81. Women were asked in an open-ended format to list their three top health concerns for themselves and then for Canadian women. Responses were recorded verbatim and categorized into one of nine mutually exclusive health concern categories. The three main health concerns for Canadian women were: Psychosocial Issues, Other Specific Illnesses, and Cancer. The three most important personal health concerns were Psychosocial Issues, Other Specific Illnesses, and Heart and Related Diseases. Few ethnic differences were noted. Results suggest that it is important to recognize and attempt to alleviate health concerns about stress and depression, which are not usually considered as being major health problems by health care professionals. PMID- 11257994 TI - [Emotional, physical and social consequences of breast cancer: viability and utilization of a clinical questionnaire]. AB - The aim of this study was to evaluate the reliability of a French Canadian version of the "Psychological Consequences Questionnaire" (PCQ) among 306 Quebec women between the ages of 50 and 69. The internal consistencies of the emotional, physical and social dimensions are respectively 0.91, 0.86 and 0.72 (Cronbach's alpha). The temporal stability was greater than 68% for each question. Meanwhile, analysis showed that 4 of the 12 questions were highly predictive of the results of the whole questionnaire (R2 = 0.91). In addition, these 4 questions are in concordance with the diagnostic criteria for adaptive disorder, anxiety disorder and general anxiety. The French Canadian version of PCQ was found to be easy to use and reliable. We would suggest a shorter version of the questionnaire, including only four questions. This version would be even easier to use and more effective to control the emotional, physical and social consequences over the different stages of breast cancer screening. PMID- 11257995 TI - Descriptive analysis of endemic cryptosporidiosis cases reported in Ontario, 1996 1997. AB - Endemic cryptosporidiosis in Ontario was studied using notifiable disease data from the Ontario Ministry of Health for the years 1996-1997 inclusive. For this study period, 451 endemic cases were identified, corresponding to a provincial mean annual age- and sex-adjusted incidence rate of 2.13 cases per 100,000. Children under five years of age had the highest incidence of disease. Males had a higher incidence than females, except for those 15-19 years of age. Five percent of cases were reported as HIV-positive or having AIDS. The proportion of cases occurring between July and November inclusive (63%) was significantly higher than expected (42%) assuming no seasonal variation (p < 0.01). The proportion of rural cases observed (29%) was significantly higher than expected (17%) based on the Ontario population (p < 0.01). Travel to or prior residence in an endemic area was identified in 22% of the cases where a risk setting was reported (n = 265). PMID- 11257996 TI - A serological survey of rural dogs and cats on the southwestern Canadian prairie for zoonotic pathogens. AB - A survey for antibodies against agents of plague, tularemia, and Rocky Mountain spotted fever (RMSF), and against Sin Nombre hantavirus (SNV), Bartonella henselae and B. clarridgeiae was conducted in the summer of 1995 using serum from rural dogs and cats living in the vicinity of four public parks in southeastern Alberta and southwestern Saskatchewan. Antibodies to all pathogens were detected in all survey areas. Overall prevalence rates were 0.075 for Yersinia pestis, 0.089 for Francisella tularensis, 0.025 for Rickettsia rickettsii (dogs only), and 0.029, 0.178 and 0.186 for SNV, B. henselae and B. clarridgeiae, respectively (cats only). This serological survey of rural dogs and cats was more sensitive and efficient than previous surveys based on collection and culture of rodents and ectoparasites. All six pathogens appear endemic to the region. Surveillance for plague, tularemia, RMSF and SNV, and management of associated public risks should be done in endemic regions. PMID- 11257997 TI - Some reflections on the relationship of risk, harm and responsibility in recent tobacco lawsuits, and implications for public health. PMID- 11257998 TI - Work-related population health indicators. AB - OBJECTIVE: To provide evidence-based recommendations for work-related population health indicators. METHODS: Drawing on a framework of work-related experiences, we systematically reviewed studies that assess the association between these experiences and health and reviewed related measures at the population level that could be used as indicators. RESULTS: We recommend (and grade the strength of evidence supporting our recommendation for) the following indicators for which data are already routinely collected: unemployment rate (strong), long-term unemployment rate (limited), and permanent lay-off rate (limited). As well, we recommend and grade our support for the following new indicators: insecurity associated with pending job loss (limited), with possible major organizational change (limited), and with actual major organizational change (limited); and job strain (medium). CONCLUSION: These evidence-based indicators can be used to monitor work-related determinants of health and thus to inform the conceptualization, development, and evaluation of policies and programs related to these determinants. PMID- 11258000 TI - Two-year-old children's sensitivity to the referential (in)efficacy of their own pointing gestures. AB - In three studies, two-year-old children communicated to a parent which of two out of-reach objects contained a sticker. Across trials, the objects were positioned in different configurations so that it was possible or impossible for a child's pointing gesture to unambiguously specify one object. In Study 1, the objects used were two boxes distinguished by a different picture of a vehicle on the front, and children (n = 16; mean age 2;8) were significantly more likely to name the box's picture on trials where pointing alone could not unambiguously specify the box than on trials where it could. In Studies 2 and 3, the stickers were hidden inside different animal figures. Older two-year-olds (n = 16, mean age 2;9), but not younger two-year-olds (n = 16, mean age 2;4), showed an ability to recognize the referential (in)efficacy of their pointing gestures and to adapt their communication accordingly. PMID- 11257999 TI - Controlling the ischemic heart disease epidemic: strategies for the 21st century. PMID- 11258001 TI - The role of performance limitations in the acquisition of verb-argument structure: an alternative account. AB - This study investigates the role of performance limitations in children's early acquisition of verb-argument structure. Valian (1991) claims that intransitive frames are easier for children to produce early in development than transitive frames because they do not require a direct object argument. Children who understand this distinction are expected to produce a lower proportion of transitive verb utterances early in development in comparison with later stages of development and to omit direct objects much more frequently with mixed verbs (where direct objects are optional) than with transitive verbs. To test these claims, data from nine children aged between 1;10.7 and 2;0.25 matched with Valian's subjects on MLU were examined. When analysed in terms of abstract syntactic structures Valian's findings are supported. However, a detailed lexical analysis of the data suggests that the children were not selecting argument structure on the basis of syntactic complexity. PMID- 11258002 TI - The acquisition of control crosslinguistically: structural and lexical factors in learning to licence PRO. AB - Rules for interpreting empty category (EC) subjects of complement clauses vary crosslinguistically across structural and lexical dimensions. In adult Greek, a distinction is made between the verbs meaning WANT and TRY, the former but not the latter permitting the EC subject of its subjunctive complement to refer outside the sentence. The EC is pro for WANT and PRO for TRY. In adult Spanish, both the verbs meaning WANT and TRY require the EC subject (pro) to refer outside when the complement is in the subjunctive, and require the EC subject (PRO) to refer to the main clause subject when the complement is in the infinitive. Twenty three Greek-speaking four- to five-year-olds and 10 adults, 29 Spanish-speaking four- to five-year-olds, 18 six- to seven-year-olds and eight adults took part in act-out experiments. The results indicate an awareness of language-particular distinctions governing the interpretation of EC complement subjects. However, child speakers of both languages experience difficulty in giving sentence external reference, leading to error in the case of subjunctive sentences for Spanish-speaking children. We argue that the data overall is most compatible with children having access to the empty category PRO by age four, and that failure to give external reference of an EC when required can plausibly be treated as performance error. A picture verification task produced less clear results, but points to the need for data from younger children to establish whether there is an early stage in which lexical semantics dominates children's interpretation of ECs. PMID- 11258004 TI - Cantonese consonantal development: towards a nonlinear account. AB - Descriptions of the development of prosodic and segmental tiers of children's phonological systems have been derived from investigations of the development of English. This paper provides a preliminary description of phonological tier development in Cantonese-speaking children. Eight children, (two each at 1;7, 2;6, 3;5, and 4;2 years) named 95 pictures. The data were analysed for word, syllable, onset-rime, skeletal, and segmental tiers. The results suggested a developmental order in the acquisition of hierarchical features. Decreasing order of accuracy of the tiers was word = syllable > onset-rime = skeletal > segmental. A model of feature geometry was adopted to describe the acquisition of features. An interesting finding is the way the laryngeal feature (aspiration) was combined with place contrasts one at a time rather than all at once. PMID- 11258003 TI - Development of sentence interpretation strategies by typically developing and late-talking toddlers. AB - Three studies, designed to examine use of word order and animacy for interpretation of sentences by 21 typically-developing two-year-old, 23 typically developing two-and-one-half-year-old, 16 typically-developing three-year-old, 17 language-delayed two-and-one-half-year-old and 19 language-delayed three-year-old children were carried out. Results indicated that typically-developing two-year olds used neither cue consistently to interpret sentences. Typically-developing two-and-one-half-year olds, on the other hand used a coalition of word order and animacy cues and language-delayed two-and-one-half-year olds used neither cue. At three years of age both groups of children used word order exclusively to interpret sentences. PMID- 11258005 TI - Filler syllables: what is their status in emerging grammar? AB - Although it has long been observed that some children incorporate unglossable syllables into their early utterances, it has been difficult to integrate these 'fillers' into theories of language acquisition. Because they straddle boundaries between phonology and morphosyntax, and between pragmatics and lexicon, they do not fit neatly into linguists' notions about 'modules' of language. Fillers have been reported in quite an array of languages, and yet they seem to be more common among learners of some languages than others. Even when language is held constant, children seem to vary immensely as to whether they produce fillers at all. With more researchers reporting fillers in more languages, it seems time to (1) review what we now know about fillers; (2) propose a reasonably unified set of criteria for identifying them; and (3) suggest an approach that will promote their further study. PMID- 11258006 TI - Discriminating between constructivist and nativist positions: fillers as evidence of generalization. PMID- 11258007 TI - A prosodic approach to filler syllables. PMID- 11258008 TI - On the possible rise and inevitable fall of fillers. PMID- 11258009 TI - Fillers across languages and language abilities. PMID- 11258010 TI - Early fillers: undoubtedly more than phonological stuffing. PMID- 11258011 TI - Fillers: how much do they generalize? PMID- 11258012 TI - The importance of studying filler-producing children. PMID- 11258013 TI - More crosslinguistic evidence on fillers in the late single-word period. PMID- 11258014 TI - Maternal responses to word approximations in Japanese children's transition to language. AB - Verbal/vocal interactions of three Japanese mother-child dyads were examined when the children were 1;0, 1;2 to 1;3, 1;6 to 1;7, and 1;8 to 1;9 to determine whether mothers provide information which may facilitate the elaboration of child lexical forms during the transition from the prelinguistic to the linguistic period. Mothers were likely to reproduce only the child's word-like utterances, both well- and ill-formed. This provided an opportunity for the child's ill formed word-like utterance to be contrasted with an immediate maternal response. This finding, along with results showing within-child variability of lexical forms, suggested that maternal contrastive replies 1) signal errors to the child (cf. Saxton, 1997), and 2) may promote the child's selection and stabilization of production alternatives which are more accurate. Maternal reproductive responding presumably originated in their tendency to seek content-oriented communication, as was reflected in mother's growing inclination to continue verbal interactions following the child's non-word-like vocalizations. PMID- 11258015 TI - Mixing and pragmatic parental strategies in early bilingual acquisition. AB - This paper investigates the relationship between a child's degree of bilingualism and features of parental input. It seeks to demonstrate that parental discourse strategies have a direct bearing on the levels of mixing present in the child's utterances in his weaker language, English. It is based on the longitudinal study of a Catalan/English bilingual child from 1;3 to 4;2 years old. The strategies adopted by both parents in response to their child's mixing are examined following Lanza's (1992, 1997) categorization of parental discourse strategies. Whereas the Catalan-speaking mother negotiates a bilingual context of interaction with her son, as of the child's third year, the English-speaking father endeavours to impose a monolingual context. Such a change of strategy clearly favours the child's increasing use of the minority language, which entails a sharp decline in rates of mixing. It appears that parents' pragmatic choices may have an impact on the development of productive family bilingualism. PMID- 11258017 TI - The need for institutional humanism and pride. PMID- 11258016 TI - Function as a criterion for the extension of new words. AB - Four experiments examine the child's ability to draw on functional information to predict the similarity of function across exemplars and to extend new words from an initial exemplar to one of two others. Experiments 1 and 2 tested 16 three- and four-year-olds each (mean ages: 4;5 and 4;4, in Experiments 1 and 2, respectively) and reveal that three- and four-year-olds are able to predict similarity of function and to extend new words on the basis of function if the function is based on shape, but not if the function is based on material makeup. Experiment 3 tested 53 older children, at three age groups (mean ages: 7;2, 8;3, 9;10) and suggests that seven-year-olds have more difficulty in judging similarity of function on the basis of material makeup than eight- and nine-year olds, and that nine-year-olds are more likely than seven- and eight-year-olds to use functional information to extend new words. Experiment 4 tested 108 children in two age groups (with mean ages of 7;9 and 9;8) and reveals that when function is pitted against syntax as criteria for categorization under a new name, seven year-olds are more attentive to syntax than nine-year-olds, while nine-year-olds are more attentive to function than seven-year-olds. The cognitive difficulties associated with judgements concerning material function are discussed in relation to additional factors that could lead children under age nine to perform in a non adult-like fashion in the extension of new names. PMID- 11258018 TI - Perception of harm and hidden benefits from human cloning by somatic cell nuclear transfer. PMID- 11258019 TI - Liberty, equality, and genetic selection. PMID- 11258020 TI - The search for a nonaddicting opioid. PMID- 11258021 TI - Francois Rabelais: Renaissance man, philosopher, author, and physician. PMID- 11258022 TI - The many domains of humanism in the care of patients. PMID- 11258023 TI - Nursery songs. PMID- 11258024 TI - Professional obligations and abortion referral. PMID- 11258025 TI - Blalock and cigarettes. PMID- 11258026 TI - Hellen Taussig--a heroine. PMID- 11258027 TI - Marriage and residency--incompatible! PMID- 11258028 TI - [Defect coverage of the hand and forearm with a free scapula-parascapula flap]. AB - BACKGROUND: Complex defects of the forearm and hand are associated with the loss of important structures. Single-stage reconstruction of these defects requires composite tissue transplantations. The subscapular region offers various components for the coverage of complex defects. The scapular and the parascapular flaps can be used each as cutaneous, fasciocutaneous and osteocutaneous or as a combined flap as well. The purpose of this study was to present our experience with the combined scapular-parascapular free flap for defect coverage of the forearm and hand in 13 patients. PATIENTS AND METHOD: Evaluation of 12 patients who underwent coverage of forearm and hand defects with the combined scapular parascapular free flap during a five-year period. Other treatment options are discussed. RESULTS: Average age of the patients was 39 years, there were ten male and two female patients. Average follow-up was 20 months. Eleven patients suffered from massive trauma, one patient from severe infection of the forearm and hand. The defect size varied from 12 x 8 cm to 45 x 20 cm. In nine cases, a cutaneous and/or fasciocutaneous scapular-parascapular flap was used, two patients underwent defect coverage with an osteocutaneous scapular-parascapular flap, and in one patient, a "four-flap-mega-flap" was transplanted. One flap developed a partial necrosis. Eight patients developed a good or very good functional outcome with their hand and forearm, two patients have a moderate degree of disability. Two patients can use their hand as a supporting hand. CONCLUSION: This study demonstrates that the combined scapular-parascapular free flap is a reliable treatment choice for early coverage of defects of the forearm and hand. The advantages are the long, large and consistent vascular pedicle, the possibility of combination with other flaps and "custom-tailoring", including whatever component is necessary to close the particular defect. PMID- 11258029 TI - [A new classification for standardizing nomenclature in wound closure by microvascular flap-plasty]. AB - The profusion of terms currently used to describe microvascular flap wound closure according to the time of reconstruction makes reliable comparisons of outcomes between institutions difficult if not impossible. To address the issue, a consistent terminology applicable to microvascular flap wound closure in general was formulated with respect to our experience with a total of 197 microvascular tissue transplantations. The nomenclature presented divides microvascular flap closure into three categories: "primary microvascular flap closure" (within 24 hours). "delayed primary microvascular flap closure" (two to seven days), and "secondary microvascular flap closure" (after seven days). This is consistent with known biological, microbiological, and surgical principles of wound closure in general and should provide a simple basis for classifying microvascular flap wound closure. Sample cases are selected to illustrate the categories within this new classification scheme. PMID- 11258030 TI - [Gradual wound closure by dermatotraction of the hand]. AB - There are various treatments for skin defects. In our study we used a skin stretching device for the closure of six skin defects of the hand and wrist (Sure Closure System, Life Medical Sciences, Inc., Princeton, N.J.). Three defects were closed completely. The size of the other three wounds was reduced by 50 to 70%. We saw no complications. The skin-stretching device helped to close skin defects with local sensate tissue. PMID- 11258031 TI - [Hand injury caused by air bag trauma. Case report and review of the literature]. AB - Although the airbag is designed to protect people from injury, it sometimes causes injuries. A case with open dislocation of the first carpometacarpal joint and fractures of the ulna and of several ribs due to an airbag is presented. The treatment is reported and the prevention of this injury is discussed with the literature reviewed. PMID- 11258033 TI - [Stener lesion yes or no? Diagnosis by ultrasound]. AB - 25 patients with clinical signs for ulnar collateral ligament injury to the metacarpophalangeal joint of the thumb were examined by ultrasonography before operation to verify Stener's lesion. The results were compared with intraoperative findings. In 18 patients (74%), ultrasound and surgical exploration yielded equivalent results. Seven patients showed signs of Stener's lesion in ultrasound that could not be verified during surgery. No Stener's lesion was missed. PMID- 11258032 TI - [Is mini-screw osteosynthesis a suitable procedure in surgical treatment of intra articular distal phalanx fractures?]. AB - In a retrospective clinical study, the results of operative treatment of intraarticular fractures of the distal phalanx were studied. From February 1992 to December 1998, 75 patients were treated operatively, 47 were examined. 14 patients were treated with Lengemann wire and temporary transfixation of the DIP joint, 25 patients were treated with screw fixation and in eight patients another procedure (wire suture, Kirschner wire, "Hakendraht", combination of screw and wire) was used. All patients had little or no pain. All patients had no or only little limitation in activities of daily living. Two patients after screw fixation had serious limitations. Four patients after Lengemann wire fixation had an extension lag in the DIP joint of up to 10 degrees, four patients between 11 and 20 degrees and one patient 25 degrees. 19 patients after screw fixation had an extension lag of the DIP joint of up to 10 degrees, five patients between 11 and 20 degrees and one patient of 25 degrees. Arthrosis of the DIP joint was seen in five patients after Lengemann wire fixation and in two patients after screw fixation. Screw fixation in the treatment of intraarticular fractures of the distal phalanx is a good, but demanding procedure with the potential for exact reconstruction of the joint surface, since implant removal is not necessary. PMID- 11258034 TI - [Klippel-Trenaunay syndrome as a rare cause of carpal tunnel syndrome--case report]. AB - The case of a 12-year-old girl with Klippel-Trenaunay syndrome and the signs of median nerve entrapment is reported. A cavernous haemangioma surrounding the median nerve is thought to be responsible for the carpal tunnel syndrome through mechanical compression and ischemic damage to the nerve due to the changed haemodynamic situation in the carpus. PMID- 11258035 TI - [Measuring grip strength of the hand with a sensor glove by gripping with submaximal and maximal strength]. AB - Grip strength measurements with the aid of dynamometers are dependent on the cooperation and motivation of the individual examined. Recognising a simulated loss of grip is a hitherto unresolved problem. With a sensor glove developed at the Berlin University of Technology, the force distribution pattern of any form of grip can be captured via ten pressure sensors and evaluated by computer. In five subjects, the force distribution patterns at submaximum and maximum force of five different grips were determined in three successive measurements each. The data were recorded in force-time diagrams. For each act of grasping, the measurements produced a curve with near-constant measuring signals within a defined period (plateau). The difference in the plateau heights of successive grips and the structures of the individual plateaux (oscillation, standard deviation) were used as distinguishing criteria. The dynamics of the measuring signals showed statistically significant differences between grips with submaximum and maximum force (p < 0.0001). PMID- 11258036 TI - [Reproducibility of measuring the finger joint angle with a sensory glove]. AB - Joint angles are measured using goniometers according to the neutral-zero method. These measurements are dependent on the examiner and are thus subject to great variation. With a newly developed sensor glove, the grip patterns of the hand can be captured with a computer-assisted technique. In a comparative study involving five subjects, the joint angles during power grip around a cylindrical body and while clenching a fist were determined by nine examiners at three different times, using the conventional technique according to the neutral-zero method. In the same experimental design, the joint angles were measured with the sensor glove. As a criterion for the reproducibility of the measurements, the intraclass correlation coefficient (ICC) was calculated by means of an analysis of variance. The ICC is a value which approximates 1 if the reliability of the measurements is high. At 0.94, the ICC of the data determined with the sensor glove was markedly higher than the mean ICC for all examiners of 0.50 using the conventional method. Besides the measurement of the finger joint angles, the sensor glove allows the dynamic recording of joint movements of all finger joints during different grip patterns of the hand. PMID- 11258037 TI - [E-mail in plastic surgery--a useful addition to current communication possibilities?]. AB - The internet is a global network of computers with a broad variety of services and options. It offers completely new ways of communication and investigation. At present, this new medium seems to be the final technical revolution in communication. It presents new dimensions which were utopia so far. Electronic mail (e-mail) is the most frequently used service within the internet. For a plastic surgeon it seems to be the fastest and the most cost-effective way of transferring data of any kind. The aim of this paper is to critically assess the transfer of confidential data via e-mail as well as to present general conditions for the use of e-mails. Therefore, our own experiences made with this technology are described to point to the possibilities, but also to indicate problems and weaknesses of this medium. Some rules should be obeyed for the transfer of confidential patient data via e-mail in order to satisfy the demand for data security standards. Also medicolegal aspects must be considered. These recommendations or rules and aspects will be discussed to offer a guideline to plastic surgeons for their e-mail applications. PMID- 11258038 TI - Homodigital reconstruction of the digits--the perspective of one unit. AB - Over a period of ten years, a strategy of reconstruction of digital injuries has been evolved in our unit which now allows us to reconstruct most defects of the digits with more predictable and, hopefully, better results than in the 1980s. This policy includes much of the philosophy and techniques of earlier and contemporary surgeons, with local modifications which we believe are improvements of the earlier techniques and a few new procedures. Wherever possible, the extraordinary capacity of the digital skin to regenerate has been exploited and homodigital techniques of reconstruction are used to limit the total injury of the trauma and reconstruction to the damaged digit. The techniques which we currently use are summarised in this article. PMID- 11258039 TI - Effects of an onion-olive oil maceration product containing essential ingredients of the Mediterranean diet on blood pressure and blood fluidity. AB - Twenty-four patients with arterial hypertension (WHO class I) received either 4 capsules of an onion-olive oil maceration product, containing essential ingredients of the Mediterranean diet, or placebo daily over a period of one week. In order to investigate the acute effect on arterial blood pressure, measurements were performed before and 5 h after the administration of the first dose of 4 capsules verum or placebo, respectively. For the evaluation of the long term effect further blood pressure measurements were performed after one week's treatment with a daily dose of 4 capsules. After a wash-out phase of 2 weeks followed by a crossover, the second study phase, which was identical in design, was carried out. In addition patients were instructed to measure their blood pressure 4 times daily at home for the whole study period. Haemorheological parameters (platelet aggregation, erythrocyte aggregation, plasma viscosity and haematocrit) were also determined at the measuring points mentioned above. The onion-olive oil maceration product led to a significant decrease in systolic blood pressure. There was also a trend towards a decrease in diastolic blood pressure. The improved blood fluidity observed resulted from a decrease in haematocrit. All effects could be shown immediately and after one week's administration. PMID- 11258040 TI - [Drug interactions--their impact on safe drug therapy in the example of the new thiazolidinedione group (glitazone)]. AB - Troglitazone (CAS 97322-87-7), rosiglitazone (CAS 155141-29-0) and pioglitazone (CAS 111025-46-8) represent novel agents for the treatment of diabetes mellitus. Very often such patients receive several drugs at the same time and consequently their interaction potential needs to be known, especially as troglitazone was recently withdrawn from the market partly because it inhibited and induced drug metabolism. PMID- 11258041 TI - Synthesis, characterization and analgesic activity of new 4-arylhydrazono-3 methoxymethyl-2-pyrazolin-5-ones. AB - New 4-arylhydrazono-3-methoxymethyl-2-pyrazolin-5-ones (5a-j) and 4-arylhydrazono 3-methoxymethyl-1-phenyl-2-pyrazolin-5-ones (6a-j) were synthesized by the reaction of ten novel methyl 2-arylhydrazono-4-methoxy-3-oxobutanoates (4a-j) with hydrazine hydrate and phenylhydrazine, respectively. Treatment of 5a with acetic anhydride yielded 1-acetyl-4-arylhydrazono-3-methoxymethyl-2-pyrazolin-5 one (7). The structures of the tautomerically dynamic compounds were established by spectral data (IR, 1H-NMR, 13C-NMR, EIMS and CIMS) and elemental analysis. The analgesic activity of the title compounds was determined by the modified Koster's test. The most active compound was 4-[(4-chlorophenyl)hydrazono]-3-methoxymethyl 1-phenyl-2-pyrazolin-5-one (6c) demonstrating twice the activity (60%) exerted by the reference drug acetylsalicylic acid (ASA, 27%). The majority of the tested compounds were found to be more effective than ASA. PMID- 11258042 TI - Tissue distribution of tolterodine, a muscarinic receptor antagonist, and transfer into fetus and milk in mice. AB - Tolterodine ((R)-N,N-diisopropyl-3-(2-hydroxy-5-methyl-phenyl)-3 phenylpropanamine, CAS 124937-51-5) is an antimuscarinic agent developed specifically for the treatment of the overactive bladder. In this study, the extent and profile of tissue distribution of 14C-tolterodine, after single and repeat oral dosing, was investigated in the mouse. Overall, distribution of radioactivity in tissues was rapid, and there were no gender-specific differences. The concentration of radioactivity in most tissues was similar to, or exceeded, that in blood. Highest concentrations were measured in gall bladder, urinary bladder, liver, kidneys and lungs, while the lowest concentration (10 times lower than in plasma) was seen in the brain. The distribution pattern after repeat oral dosing was similar to that after a single dose, although the decline in tissue concentrations was slower. Studies in pregnant mice showed that the distribution of radioactivity differed between dams and fetuses. Radioactivity was low and showed homogeneous distribution in the fetus, while a heterogeneous pattern was seen in the dam. Highest concentrations were seen in the fetal liver, brain and spinal cord. Some accumulation was observed in the choroid plexus. Placental concentrations of radioactivity were generally higher than those in the fetus, with some accumulation in the yolk sac. Studies in suckling mouse pups showed low levels of exposure to drug-related radioactivity (around 0.2% of the dose); the milk:plasma concentration ratio was 0.0-0.7. In conclusion, tolterodine and its metabolites are rapidly distributed into tissues following oral administration of radiolabelled drug in the mouse, with many tissues (including the fetus) reaching similar concentrations to that observed in blood. In other tissues, especially the eliminating organs, radioactivity levels were much higher than in blood. Penetration of the central nervous system was low, suggesting that the risk of deleterious effects on cognitive function may be lower with tolterodine than with more lipophilic antimuscarinic drugs. PMID- 11258043 TI - Pharmacokinetics of tolterodine, a muscarinic receptor antagonist, in mouse, rat and dog. Interspecies relationship comparing with human pharmacokinetics. AB - Tolterodine ((R)-N,N-diisopropyl-3-(2-hydroxy-5-methyl-phenyl)-3 phenylpropanamine, CAS 124937-51-5) is an antimuscarinic agent developed specifically for the treatment of the overactive bladder. In this study, the pharmacokinetics of tolterodine were investigated in the mouse, rat and dog, following which allometric scaling was performed to predict oral pharmacokinetics in man. The intestinal absorption of tolterodine after oral dosing was almost complete in all three species, with peak serum concentrations observed within 1 hour post-dose. Bioavailability varied between 2-20% in rodents and 58-63% in the dog. A high volume of distribution in all three species was consistent with extravascular distribution. Tolterodine was extensively metabolised in all the animal models, but the profile of metabolism differed in the rat compared to the mouse and dog, the latter having similar metabolism routes as man. Limitation of metabolism capacity caused a non-linear increase of tolterodine concentrations with dose (repeat-dose study in the mouse), and changed the relative metabolite concentration pattern. The results suggest that the hydroxylation of tolterodine is a high affinity, low capacity pathway, while N-dealkylation follows a low affinity, high capacity pathway. The elimination of tolterodine from serum was rapid, with a half-life of less than 2 h in all species. A very high clearance in the mouse and rat (10-15 l/h.kg), and in the dog (1.4 l/h.kg), indicated additional non-metabolic clearance mechanisms for tolterodine (shown to be attributed to biliary excretion). Urinary excretion of compound-related radioactive substance was around 15%, 45% and 50%, respectively, in the rat, mouse and dog. Allometric scaling allowed a good prediction of clearance and volume of distribution to be extrapolated for comparison with tolterodine pharmacokinetics in man. In conclusion, the pharmacokinetics of tolterodine are similar in the mouse and dog, and correlate with that in man. Although the rat has a different metabolic profile, clearance fits into the allometric relationship between species, enabling prediction of total clearance of tolterodine in man from preclinical data. PMID- 11258044 TI - Bioequivalence study of finasteride. Determination in human plasma by high pressure liquid chromatography coupled to tandem mass spectrometry. AB - Two different finasteride (CAS 98319-26-7) tablet formulations were evaluated for their relative bioavailability (Flaxin tablets 5 mg, as the test formulation vs reference formulation, tablets 5 mg) in 23 healthy male volunteers who received a single 5 mg oral dose of each preparation. The study was open, randomized with a two-period crossover design and a 7-day washout period. Plasma samples were obtained over a 48-h interval. The finasteride concentrations were determined by high-pressure liquid chromatography (HPLC) coupled to tandem mass spectrometry (LC-MS-MS). The analytical method developed has a limit of quantitation (LOQ) of 0.50 ng/ml in plasma. For the quality control the measured concentration was 2.05 +/- 0.14 ng/ml (mean +/- SD, n = 30) with a precision of 6.9% and an accuracy of 2.55% at a concentration of the starting solution of 2.00 ng/ml, while with 20.00 ng/ml starting solution the measured concentrations were 20 +/- 0.80 ng/ml (n = 30) with a precision of 3.81% and an accuracy of 0.09%. From the plasma finasteride concentration vs time curves the following pharmacokinetics parameters were obtained: AUC0-48, AUC0-infinity, Cmax, Cmax/AUC0-48, Ke, elimination half-life and tmax. Geometric mean test/reference formulations individual percent ratio was 95.71 for AUC0-48 h and 88.70% for Cmax. The 90% confidence interval for the geometric mean of the individual ratio test/reference formulations was 95.70-120.20% for AUC0-48 h, 94.60-121.30 for AUC0-infinity and 88.70-108% for Cmax. Since for both Cmax or AUC the 90% Cl values are within the interval proposed by the Food and Drug Administration, the test formulation is bioequivalent to the reference formulation for both the rate and extent of absorption after single dose administration. PMID- 11258045 TI - Intestinal absorption of fluoride at high luminal concentration of fluoride. AB - The paper reports that high fluoride concentrations in the intestinal lumen hinders the absorption of this anion. This conclusion was verified with three different experimental models. Pharmacokinetic experiments done in human volunteers revealed that the bioavailability of fluoride from sodium fluoride (NaF, CAS 7681-49-4) enteric coated tablets was 33% of that of plain (immediate release) tablets. The latter findings were confirmed in rats receiving 1 ml of NaF solutions (40, 80 or 160 mmol/l) by gavage. The greatest AUC (area under the curve of fluoremia as a function of time) was obtained with an oral dose of 80 mumol of NaF. This parameter was significantly greater (p < 0.01) with 80 mumol than with 40 mumol NaF, but similar to that observed with 160 mumol. Fecal fluoride excretions (in the 24 h following a single dose of NaF) and the bone fluoride contents (found at the end of 30 days of treatment with 40, 80 or 160 mumol NaF/day), agreed with the AUC values. The rate of fluoride absorption (v, mumol/10 min) through the intestinal wall was investigated with perfused, isolated rat duodenum in vivo. Fluoride absorption increased between 0 and 10 mmol/l luminal fluoride and decreased with higher concentrations. Oxygen consumption of duodenal-tissue decreased exponentially between zero (1.12 mumol O2 min-1 g-1) and 10 mmol/l fluoride (0.45 mumol O2 min-1 g-1). PMID- 11258047 TI - In vitro antimalarial activity of novel trifluoromethyl- and bis(trifluoromethyl)quinoline derivatives. AB - The in vitro antimalarial activity of a series of 2- and 8-trifluoromethyl- and 2,8-bis(trifluoromethyl)quinoline-4-(5-pyrimidino) and N4-ethyl-5 nitroimidazolo)methylene ketones was assessed against the chloroquine-sensitive strain (D10) of Plasmodium falciparum. Although the in vitro antimalarial activity of these compounds is more or less of the same order of magnitude, derivatives containing two trifluoromethyl groups achieve a slightly higher in vitro activity than compounds with one trifluoromethyl group, with 2,8 bis(trifluoromethyl) quinoline-4-(N4-ethyl-5-nitroimidazolo) methylene and 2,8 bis(trifluoromethyl) quinoline-4-(5-pyrimidino) ketones showing IC50 of 4.8 and 5.2 micrograms/ml, respectively. These compounds seem to bind to DNA by intercalation. PMID- 11258046 TI - Synthesis, antiretroviral and antioxidant evaluation of a series of new benzo[b]furan derivatives. AB - The antiretroviral and anti-oxidant profile of a series of new C-2 and C-7 substituted benzo[b]furans was explored by employing well established antiviral and antioxidant protocols. The most potent antioxidant compound tested was analog 7, which bears an OH at C-7 and a benzoyl group at C-2. In the influenza A type H3N2 virus screens analog 8a was almost five-fold more active than its counterparts and equipotent to rimantadine and amantadine. In the influenza B screening all of the new compounds tested were at least ten-fold more active than the control drug amantadine. The anti-HIV screening, using acutely infected MT-4 cells, showed that compound 8f (n = 4), was fifteen-fold more active than its monomer congeners, 8a and 8c, d and almost five-fold more potent than monomer 8b and dimer 8f (n = 3). PMID- 11258048 TI - Radioligand binding assays in the drug discovery process: potential pitfalls of high throughput screenings. AB - Radioligand binding assays evaluating directly the ability of a drug to interact with a defined molecular target is part of the drug discovery process. The need for a high throughput rate in screening drugs is actually leading to simplified experimental schemes that increase the probability of false negative results. Special concern involves voltage-gated ion channel drug discovery where a great care is required in designing assays because of frequent multiplicity of (interacting) binding sites. To clearly illustrate this situation, three different assays used in the academic drug discovery program of the authors were selected because they are rich of intrinsic artifacts: (I) (20 mmol/l caffeine almost duplicated [3H]ryanodine binding (89% higher than control) to rat heart microsomes at 0.3 mumol/l free calcium but did not exert any effect when using a high (107 mumol/l) free calcium, as mostly used in ryanodine binding assays; (II) An agonist for the ionotropic glutamate receptor of the kainate type can distinctly affect [3H]kainate binding to chicken cerebellum membranes depending on its concentration: unlabelled kainic acid per se either stimulated about 30% (at 50-100 nmol/l), had no effect (at 200 nmol/l) or even progressively decreased (at 0.3-2 mumol/l) the binding of 5 nmol/l [3H]kainate, emphasizing the risk of using a single concentration for screening a drug; (III) in a classical [3H]flunitrazepam binding assay, the stimulatory effect of a GABA (gamma aminobutyric acid) agonist was only observed when using extensively washed rat brain synaptosomes (10 mumol/l GABA increased flunitrazepam binding by 90%). On the other hand, the inhibitory effect of a GABA antagonist was only observed when using crude synaptosomes (10 mumol/l bicuculine reduced [3H]flunitrazepam binding by 40%). It can be concluded that carefully designed radioligand assays which can be performed in an academic laboratory are appropriate for screening a small number of drugs, especially if these are potential hits because of their rational design. Therefore, the low throughput rate could be partially balanced by a higher performance when compared to what is done in a robotic high throughput screening where simplification of assay conditions can lead to false negative results. PMID- 11258049 TI - Influence of a co-stimulation of human leucocytes with an Escherichia coli preparation and fixed immunoglobulins on cytokine release in the presence of hydrocortisone. AB - Pharmaceuticals of biological origin consisting of bacterial culture suspensions (BCS) as active ingredients have long been used for the treatment of hemorrhoidal diseases and chronic anal pruritogenic eczemas. However, some of these pharmaceuticals often contain glucocorticoids such as hydrocortisone as an anti inflammatory supplement. Therefore, the question arises whether the claimed immunostimulatory capacity of the bacterial culture suspension might be altered by the steroid. Up to now, numerous reports support the evidence that the stimulation of the different Fc-receptor subtypes on leucocytes result in profound immunoregulatory activities influencing phagocytosis and antigen processing, antibody-dependent cytotoxicity or secretory functions thereby enhancing the overall activities of the immune system towards foreign antigens/pathogens. With these findings in mind it was investigated whether the immunomodulatory capacity(s) of the BCS in the presence of hydrocortisone will be modified by solid-phase bound immunoglobulins (Igs). For this purpose freshly prepared human peripheral blood leucocytes (PBLs) were incubated with different concentrations of the BCS (0.1, 1, 10 micrograms/ml), either with or without fixed human immunoglobulins in the presence of increasing concentrations of hydrocortisone. As a parameter of PBL activation the secretion of different cytokines was measured, e.g. tumor necrosis factor alpha (TNF-alpha), interleukin 10 (IL-10) and granulocyte-macrophage colony stimulating factor (GM-CSF). Cytokines were determined with specific sandwich ELISAs. With this modified cell culture system it was demonstrated that the immunosuppressive activities, normally caused by hydrocortisone, were partially antagonized by the combination of BCS plus fixed Igs. TNF-alpha and GM-CSF were significantly more produced, even in the presence of hydrocortisone, whereas the synthesis of IL-10 was diminished by fixed Igs. However, this effect could be reversed with increasing concentrations of hydrocortisone. These results raise the possibility that in the natural environment, e.g. the rectal mucosa, antigens derived from the BCS are bound by specific Igs, thereby modifying secretory and effector functions of locally present leucocytes in another way as free antigens. The biological relevance of these in vitro data with respect to the therapeutic benefit of the BCS preparations with hydrocortisone will be discussed considering recent findings in the literature. PMID- 11258050 TI - Determination of the antiepileptics vigabatrin and gabapentin in dosage forms and biological fluids using Hantzsch reaction. AB - A selective and sensitive method was developed for the determination of the anticonvulsants vigabatrin (I) (CAS 60643-86-9) and gabapentin (II) (CAS 60142-96 3). The method is based on the condensation of the drugs through their amino groups with acetylacetone and formaldehyde according to Hantzsch reaction yeilding the highly fluorescent dihydropyridine derivatives. The yellowish-orange color was also measured spectrophotometrically at 410 nm and 415 nm for I and II, respectively. The absorbance-concentration plots were rectilinear over the ranges 10-70 micrograms/ml and 20-140 micrograms/ml for I and II, respectively. As for the fluorescence-concentration plots, they were linear over the ranges 0.5-10 micrograms/ml and 2.5-20 micrograms/ml with minimum detection limits (S/N = 2) of 0.05 microgram/ml (approximately 2.1 x 10(-8) mol/l) and 0.1 microgram/ml (approximately 5.8 x 10(-7) mol/l) for I and II, respectively. The spectrophotometric method was applied to the determination of I and II in their tablets. The percentage recoveries +/- SD (n = 6) were 99.45 +/- 0.13 and 98.05 +/- 0.53, respectively. The spectrofluorimetric method was successfully applied to the determination of I and II in spiked human urine and plasma. The % recoveries +/- SD (n = 5) were 98.77 +/- 0.29 and 98.39 +/- 0.53 for urine and 99.32 +/- 0.74 and 98.90 +/- 0.96 for plasma, for I and II, respectively. No interference was encountered with the co-administered drugs: valproic acid (CAS 99-66-1), diphenylhydantoin (CAS 57-41-0), phenobarbital (CAS 50-06-6), carbamazepine (CAS 298-46-4), clonazepam (CAS 1622-61-3), clobazam (CAS 22316-47 8) or cimetidine (CAS 51481-61-9). A proposal of the reaction pathway is suggested. The advantages of the proposed methods over existing method are discussed. PMID- 11258051 TI - [Treatment of malaria: prevention of resistance by combinations of antimalarial agents]. PMID- 11258052 TI - [Human and wild mammal parasitosis in French Guiana]. PMID- 11258053 TI - [Health support for a military company while traveling in the equatorial forest]. PMID- 11258055 TI - [Time to evaluate]. PMID- 11258054 TI - [East Timor: a difficult transition to independence]. PMID- 11258056 TI - [Drug resistance of Plasmodium falciparum in coastal regions of Madagascar]. AB - Chloroquine is still the drug of choice for first-line treatment of uncomplicated malaria in Madagascar. However development and spread of chloroquine-resistance could compromise this therapeutic strategy in the future. The purpose of this 1997 study was to compare the efficacy of combined treatment using sulfadoxine and pyrimethamine and single-agent treatment using chloroquine for management of uncomplicated malaria. Study data were collected at four sites in coastal areas of Madagascar where transmission of malaria is perennial. Prevalence of malaria ranged from 15 p. 100 to 22 p. 100 in school children and from 24 p. 100 to 72 p. 100 in outpatient consulting spontaneously at community health centers. All four Plasmodium species affecting man were identified. Plasmodium falciparum was involved in 83 p. 100 of cases. In vivo testing of the susceptibility of Plasmodium falciparum to chloroquine was performed in 149 patients according to the standard simplified 7-day protocol of the WHO. The 35 tests in school children demonstrated no evidence of resistance. However type R1 + R2 resistance was noted in 17 of the 114 tests performed on outpatients, i.e. 14.9 p. 100. In vitro testing demonstrated chloroquine resistance in four of the 90 specimens tested, i.e. 4.4 p. 100. With regard to combined sulfadoxine/pyrimethamine treatment, 45 of 46 in vivo tests in outpatients showed no evidence of resistance. Combination treatment was more effective than single-agent treatment (p = 0.02) and could offer an effective alternative for future use. PMID- 11258057 TI - [Malaria and pregnancy: attitude of health care personnel during prenatal care in Cotonou, Benin]. AB - The purpose of this study was to evaluate the state of knowledge and use of drug in pregnant women diagnosed with malaria at the first prenatal examination. Survey data were conducted of physicians and paramedical staff working the public and private sector in Cotonou, Benin from November 1996 to January 1997. A questionnaire including items on the type of antimalarials used, therapeutic regimens and compliance with the treatment posology and duration guidelines as defined by the WHO and recommended by the National Program against Malaria. A study of informational supports allowed validation of responses obtained during the interview on the day of survey. A random two-degree, stratified sample of 208 prescribers including 109 physicians, 51 midwives, and 48 nurses was interviewed. Data analysis showed that chemoprophylaxis was prescribed by all midwives, 87.5 p. 100 of nurses, and 8.2 p. 100 of physicians. Midwives prescribed this strategy in all pregnant women. For 5.5 p. 100 of physicians and 6.3 p. 100 of nurses, primigravid women were considered as a target group. Chloroquine was the most widely used antimalarial. It was sometimes used in association with proguanil. Of the other drugs used for prevention, quinine was prescribed by 2.4 p. 100 of nurses and 1.1 p. 100 of physicians. The therapeutic regimes and posologies used by the health care workers were not in compliance with the policies of the National Program for Control of Malaria. The findings warrant the creation of an organization to promote consultation and communication between health care authorities and workers. PMID- 11258058 TI - [Prenatal care in a semiurban area of Central African Republic: frequency, influential factors, maternal and neonatal prognosis]. AB - This 9-month longitudinal study was carried out in a cohort of 287 women hospitalized for various obstetrical reasons in Bouar, the third largest city in Central Africa. A total of 225 of these women gave birth to 229 children. The purpose of study was to evaluate attendance at prenatal examinations, risk factors for obstetrical morbidity, and maternal and neonatal prognosis. Results showed an overall absence rate at scheduled examinations of 38 p. 100. Only 8.4 p. 100 of women enrolled for prenatal care complied fully with the examination schedule. The factors associated poor attendance were age older than 19 years, distance greater than 5 kilometers between the woman's home and care facility, and illiteracy. The incidence of complicated pregnancy was significantly higher in refractory women. Reported complications included eclampsia, extrauterine pregnancy, and death of the mother. A total of 43.5 p. 100 of home deliveries involved women that did not attend prenatal care clinics. A total of 23.1 p. 100 of newborns without prenatal care presented complications requiring intensive neonatal care with an Apgar score below 7. Overall the perinatal death rate was 12.7 p. 100 (n = 29). The stillbirth rate was 10.1 p. 100 (n = 16). The findings of this study demonstrate the favorable impact of prenatal care on pregnancy outcome for both mother and child. Prenatal care should be made available to all pregnant women. PMID- 11258059 TI - [Buruli ulcer: risk of bone involvement! Apropos of 33 cases observed in Benin]. AB - Skin lesions are the best known manifestations of Mycobacterium ulcerans infection. However osteomyelitis is not uncommon with incidences as high as 14 p. 100 having been reported. The literature contains little specific data on bone lesions. This report describes a series of 33 patients presenting bacteriologically and/or histologically confirmed osteomyelitis induced by Mycobacterium ulcerans. In 64 p. 100 of cases, bone involvement occurred within the first year of the disease. In 70 p. 100 of cases, the site of involvement was on extremities. The median number of sites per patient was two (range, 1 to 5) and the number of bones was not correlated with the duration of disease. Overall, 41 p. 100 of bone sites occurred locally beneath skin lesions and 59 p. 100 were distant metastases. Osteomyelitis was associated with other germs in only 16 p. 100 of cases and superinfection was not a prerequisite for development of bone involvement. Clinical manifestations were usually low-grade with non inflammatory, slightly painful local mass. The patient's general health status was unaffected in 79 p. 100. Histological examination allowed identification of the type of bone lesions induced by Mycobacterium ulcerans. For the first time, bone biopsy specimens were cultured. Based on bacteriological and histological findings, several postulations can be made about underlying pathophysiological mechanisms. The role of Buruli toxin is dealt with in the discussion. PMID- 11258060 TI - [Acute renal failure during severe malaria: physiopathology and therapeutic management. Apropos of 2 cases]. AB - Renal failure secondary to acute tubular necrosis is a common complication of severe Plasmodium falciparum malaria. The purpose of this report is to describe two cases of severe malaria featuring acute renal failure observed in young patients who had failed to comply with chemoprophylaxis. Occurrence of renal failure was delayed four to seven days in relation to the beginning of the malaria attack. Hemodialysis was required in one case. Both patients were successfully treated by quinine perfusion. The main pathophysiology mechanisms underlying acute tubular necrosis are obstruction of capillaries and post capillary venules by infected red blood cells and activation of monocytes that release cytokines such as tumor necrosis factor. Other nonspecific mechanisms may come into play including hypovolemia, release of catecholamines and subsequent activation of the rennin-angiotensin system, complement activation, and rhabdomyolysis. Acute tubular necrosis is the main renal complication of Plasmodium falciparum malaria but latent forms of acute glomerulonephritis have also been documented. Prognosis is usually favorable depending mainly on early diagnosis and prompt treatment. PMID- 11258061 TI - [Hydatidosis: apropos of several uncommon locations]. AB - Most hydatid cysts are found in the liver or lungs but occurrence in other locations is possible. The purpose of this report is to describe three cases involving cysts locate in the ovary, joint, and thyroid. Clinical findings associated with these uncommon locations were poorly specific and diagnosis required histological examination after surgical exploration. Imaging and serology were useful to differentiate hydatid cyst from benign or malignant tumor disease but often failed to achieve definitive differential diagnosis. Thorough investigation is necessary to rule out more common locations in the liver or lung. Continued surveillance is needed for early detection of recurrence. Preventive campaigns are necessary in endemic areas, such as Morocco. PMID- 11258062 TI - [First confirmed case of laryngeal diphtheria in Djibouti]. AB - The first bacteriologically confirmed case of laryngeal diphtheria in Djibouti was reported in 1998. It involved a three-year-old native-born infant who had been vaccinated during the first year of life with three doses of a combined vaccine against diphtheria, tetanus, poliomyelitis, and pertussis. A rapid clinical improvement was observed under erythromycin treatment. Other cases of laryngeal diphtheria have been observed. It is important to reverse decreasing vaccinal coverage in Djibouti and to warn incoming travelers of the need to be adequate immunized against diphtheria. Enhanced epidemiologic surveillance of this disease is also needed. PMID- 11258063 TI - [Analysis of the prevention of post-ivermectin Loa loa encephalopathy by administration of initial low dose]. AB - A number of cases of Loa-encephalopathy have been reported following ivermectin (Mectizan) treatment for onchocerciasis in patients with high Loa microfilaraemia. A possible explanation for these severe reactions is the formation of micro-emboli in small brain vessels as a result of massive paralysis of Loa microfilariae in the blood. This suggests that encephalopathy might be prevented by giving an initial low dose of ivermectin to induce a more gradual action on the Loa microfilariae. To test this hypothesis, a trial was conducted on 23 adult patients in Cameroon. Patients were divided into two groups. One group received the recommended dose of 150 micrograms/Kg. The other group received a lower dose of 50 micrograms/Kg (one 3 mg tablet of Mectizan). Blood smears were made daily from day 1 to 7 after treatment and then on days 15 and 30. Results showed no difference in the effect of the dosage level on Loa microfilaraemia. This finding suggests that an initial low dose of 3 mg Mectizan will not prevent encephalopathy following treatment for onchocerciasis. PMID- 11258064 TI - [Imported dengue hemorrhagic fever: aprops of 1 case presenting with signs of acute alithiasic cholecystitis]. AB - Dengue is prevalent in all subtropical areas. Hemorrhagic forms of the disease were first described in southeast Asia but have now been observed on several continents. Travelers are at risk for infection and the likelihood of imported dengue has grown in relation to volume of air traffic. In developed countries, dengue usually presents in the benign form, but sudden aggravation is always possible. The purpose of this report is to describe a case of imported dengue hemorrhagic fever associated with abdominal pain in a traveler returning from Asia. Radiological findings were suggestive of nonlithiasic cholecystitis. Similar ultrasound feature have been reported by pediatric groups during dengue outbreaks in Asia. Previous findings have shown that bladder involvement is a predictive sign of severe disease and impending shock. Surgery is contraindicated in these patients. Close clinical and laboratory surveillance is necessary due to the high risk of aggravation. The pathogenesis of this severe life-threatening form of the disease is unclear. A possible explanation is involvement of a more virulent strain of virus. Dengue should always be considered after malaria in the differential diagnosis of returning travelers patients presenting fever. PMID- 11258065 TI - [Systemic tropical mycoses]. AB - Systemic tropical mycoses have been emerging since the beginning of the AIDS epidemic. The incidence of these infections is probably even underestimated since most cases occur in populations with poor access to medical care in regions where modern diagnostic methods are unavailable. In Europe, this pathology is sometimes observed in returning travelers, aid workers, and immigrants. Differential diagnosis of imported systemic tropical mycosis may be difficult for uniformed physicians. The purpose of this review of the literature is to provide up-to-date epidemiological, clinical, diagnostic, and therapeutic data on the six main systemic tropical mycoses with disseminated forms. The six mycoses described in this study are histoplasmosis, coccidioidomycosis, paracoccidioidomycosis, blastomycosis, sporotrichosis, and penicilliosis due to Penicillium marneffei. Strictly superficial and subcutaneous mycoses are not covered. PMID- 11258066 TI - [The control of lymphatic filariasis]. AB - Recent advances in the diagnosis, treatment, and pathophysiology of lymphatic filariasis have raised hopes for eradication. Development of an easy-to-use, rapid-format diagnostic tool has facilitated precise location of infectious zones thus allowing quick intervention. Two new drugs, i.e., ivermectin and albendazole, have been shown to be highly effective in the management of microfilariasis due to Wuchereria bancrofti, Brugia malayi, and Brugia timori thus greatly expanding a therapeutic arsenal once limited to diethylcarbamazine. The current eradication strategy is designed to break the chain of transmission in populations exposed to the risk of infection by administering a single dose of two drugs yearly for five to six years. Combined treatment using albendazole plus ivermectin or albendazole plus diethylcarbamazine has resulted in near-zero microfilaremia levels for at least one year. Based on these new developments, the World Health Assembly adopted a resolution calling on member states to work for the elimination of lymphatic filariasis as a public health problem. Thanks to the donation of albendazole and ivermectin by SmithKline Beecham and Merck & Co. respectively, the WHO has started a worldwide program and set up task force including a wide-range of actors in different fields for eradication of this disease affecting 120 million people. PMID- 11258068 TI - [Cholera in the Central University Hospital of Brazzaville, Congo]. PMID- 11258069 TI - [Ebola and Marburg virus: entomologic hypothesis to confirm]. PMID- 11258067 TI - [Epidemics of cutaneous leishmaniasis in military personnel working in French Guiana]. AB - Cutaneous leishmaniasis transmitted by phlebotomine sandflies is endemic in the rain forests of French Guyana. The 3rd Regiment Etranger d'Infanterie, based in Kourou carries out numerous operations in the Amazonian areas. In 1998 two outbreaks of cutaneous leishmaniasis occurred: one during an exercise at the training center in the equatorial forest of Regina (10 patients) and the other during a mission in Saint Elie (21 patients). Clinical findings were variable and diagnosis was confirmed by skin smear tests. Patients were treated by two intramuscular injections of pentmidine isethionate (Pentacarinate). Recurrence was observed in two patients who were retreated using the same agent. Persistent lesions were treated by intralesional injection of meglumine antimoniate (Glucantime). Both outbreaks were characterized by high attack rates (91 p. 100 and 84 p. 100) and were facilitated by non-observance of standard procedures because of training or operational requirements at the beginning of the leishmaniasis season. Strict planning of activities, wearing protective clothing, deployment of insecticide treated bed nets, and of candles rather than electrical lamps for lighting are key preventive measures. Greater emphasis is needed on the use of insect repellents. PMID- 11258070 TI - [A case of furuncular myiasis from Cordylobia anthropophaga in Morocco]. PMID- 11258071 TI - [Scorpion sting and acute interstitial nephropathy: Apropos of 1 case]. PMID- 11258072 TI - [Spontaneous post-obstetrical vesicovaginal fistula and bladder catheterization ]. PMID- 11258073 TI - [Human dirofilariasis in France: new data confirming the human transmission of Dirofilaria repens to the north of the 46 degree north latitude]. PMID- 11258074 TI - Declaration of independence. PMID- 11258075 TI - Composite vs. amalgam. PMID- 11258076 TI - Now's the time. PMID- 11258077 TI - Spreading the news. PMID- 11258078 TI - Surgeon general's report. PMID- 11258079 TI - Minority dentists. PMID- 11258080 TI - Initial carious lesions. PMID- 11258081 TI - Air abrasion devices. PMID- 11258082 TI - Air/particulate abrasion. PMID- 11258083 TI - Temporal arteritis. PMID- 11258084 TI - JADA online coming. PMID- 11258085 TI - Have you or a member of your staff ever sustained an injury that is unequivocally related to the provision of dental care? PMID- 11258086 TI - The impact of universal access to dental care on disparities in caries experience in children. AB - BACKGROUND: The authors investigated the association between socioeconomic status and the severity of dental caries in 6- and 7-year-old children who had had access to dental care throughout their lives. The children had lived since birth in Nova Scotia, Canada, a province with a universal publicly financed dental care program. METHODS: The authors selected a representative sample of first-grade children using a stratified multistage sampling method of primary schools (n = 1,614). The response rate was 78.8 percent. Two dentists were trained to diagnose dental caries using modified World Health Organization criteria. Intra- and interexaminer reliability was excellent (kappa > or = 0.88). Of the children who were examined (n = 1,271), 955 were lifelong residents of Nova Scotia, Canada, and so were included in this analysis. Data were weighted and adjusted for clustering (design) effects. RESULTS: Only 8.4 percent of the children had visited a dental office before the age of 2 years, and 88.5 percent of the children had their first dental visit between the ages of 2 and 5 years. Children whose parents had completed a university education had a significantly lower mean number of decayed, missing and filled surfaces, or dmfs, in their primary teeth than did children whose parents had a lower education level. A Poisson regression model indicated that parents' high education status, optimal fluoride concentration in schools' water supplies, daily toothbrushing and dental visits for checkup were significantly associated with low dmfs scores. CONCLUSION: Having access to a universal publicly financed dental insurance program since birth did not eliminate the disparities in caries experience. PRACTICE IMPLICATIONS: This analysis of a highly utilized universal dental insurance program suggests that disparities in oral health status cannot be reduced solely by providing universal access to dental care. Focused efforts by professional and governmental organizations should be directed toward understanding the socioeconomic, behavioral and community determinants of oral health disparities. PMID- 11258087 TI - Widespread pain and the effectiveness of oral splints in myofascial face pain. AB - BACKGROUND: The research literature reaches inconsistent conclusions about the efficacy of oral splints for treating myofascial face pain. This investigation hypothesizes that their effectiveness varies as a function of the presence or absence of widespread pain. METHODS: In a randomized, controlled clinical trial, 63 women with myofascial face pain were assigned to use of either an active, maxillary, flat-plane, hard acrylic splint or a palatal splint that did not interfere with occlusion. Participants also were classified according to the presence or absence of widespread pain throughout the body. After six weeks, groups were compared regarding pain on palpation, self-reported pain and functional outcome. RESULTS: Overall, the findings showed a modest tendency for subjects receiving the active vs. the palatal splint to exhibit improvement on self-reported pain and functional outcome. On further division of the sample into subjects with local vs. widespread pain, the general pattern showed that patients with widespread pain who received an active splint did not experience improvement, while patients with local pain who received the active splint did. CONCLUSIONS: The presence or absence of widespread pain may help to define the specific circumstances under which oral splints should be prescribed for patients with myofascial face pain. CLINICAL IMPLICATIONS: Clinicians should screen patients with myofascial face pain for the presence of widespread pain, since this comorbid symptom pattern may be a contraindication for the use of oral splints. PMID- 11258088 TI - Is use of exogenous estrogen associated with temporomandibular signs and symptoms? AB - BACKGROUND: Studies of historical data suggest a link between exogenous estrogen use and referral for treatment for temporomandibular disorders, or TMDs. The purpose of the authors' study was to determine the association between exogenous estrogen use and signs and symptoms of TMD assessed by direct physical examination in a randomly selected community sample of primarily postmenopausal women. METHODS: A calibrated clinical examiner examined a stratified random sample of 510 women aged 37 to 82 years using the Craniomandibular Index, or CMI. All medications that subjects were taking at the time of the examination were identified by interview and examination of subjects' medication containers on two occasions. One hundred seventy-four subjects were taking medications containing estrogen, and 336 were taking no such medications. RESULTS: The muscle and joint signs and symptoms of women taking and not taking estrogen were not significantly different after the authors controlled for sociocultural, demographic and health care utilization variables. Estrogen use also failed to distinguish women receiving relatively high and low scores on the CMI. CONCLUSION: Estrogen replacement therapy does not place women at increased risk of developing TMDs. CLINICAL IMPLICATIONS: Clinicians need not be concerned that patients taking oral contraceptives or replacement estrogens are at increased risk of developing TMDs. PMID- 11258089 TI - Evaluating, documenting and following up oral pathological conditions. A suggested protocol. AB - BACKGROUND: Many textbooks and articles are available to assist dentists in examining patients, establishing diagnoses for oral lesions and understanding the techniques of biopsy. There is little guidance in the literature, however, on when and how to follow up lesions that have a low index of clinical suspicion, or for which the pathological diagnosis does not demonstrate any overt signs of malignancy or premalignancy. TYPES OF STUDIES REVIEWED: The authors reviewed the literature, talked to numerous clinicians and sought legal opinions regarding how a reasonable and prudent dentist should manage patients with clinically evident oral lesions that do not suggest any adverse long-term effects on the health and safety of the patient. RESULTS: The few guidelines available in the literature, coupled with the observations of the authors and others, allow logical and reasonable interim recommendations to be proposed regarding the frequency of examinations, the timing of invasive procedures and medicolegally prudent documentation guidelines. Future studies are needed to refine these recommendations. CLINICAL IMPLICATIONS: Some dentists have been sued for alleged failure to monitor patients, document cases or refer patients with oral lesions. The recommendations provided here can help dentists manage these patients, but they should not be construed as being rigid guidelines or legal standards that apply to all clinical situations. In some cases, the judgment and experience of clinicians may indicate the need to deviate from these recommendations. Refinements of these guidelines may emerge on the basis of future studies. PMID- 11258090 TI - A method for removing a fractured resin post and core pattern. PMID- 11258091 TI - Selecting sleep-disordered-breathing appliances. Biomechanical considerations. AB - BACKGROUND: Dentists who wish to provide sleep-disordered-breathing therapy have many different mandibular advancement devices, or MADs, from which to select. Documented research directly about the variations in MADs is sparse. TYPES OF STUDIES REVIEWED: The author reviewed dental and medical literature dealing with biological and mechanical principles affecting the function of MADs. RESULTS: The author found that MADs vary in four major areas: freedom of mandibular movement, amount and rigidity of dental coverage, amount of mandibular advancement and amount of bite opening. Each of these areas appears to affect the appliance's efficacy, safety or both. The main potential detrimental effect of MADs is occlusal shifting. The author presents biological and mechanical considerations in an attempt to determine the optimum parameters for each of the MAD variation areas. The MAD must be constructed in a manner and with material that secures the mandible in its optimum position. The optimum mandibular position needs to be captured and transferred to the articulator with an accurate construction bite. CLINICAL IMPLICATIONS: MAD therapy may last a lifetime. Therefore, dentists must consider the efficacy and the safety of an MAD when selecting an appliance. Since occlusal shifting appears to be the main potential detrimental effect, dentists should consider all available means to monitor and minimize these changes. PMID- 11258092 TI - The amalgam controversy. An evidence-based analysis. AB - BACKGROUND: There are a number of patients and health care professionals who believe dental amalgam restorations are a factor in a host of diseases and conditions. They have been influenced by anecdotal case reports in the medical and dental literature, research published in the refereed literature and media stories concerning the alleged dangers of amalgam restorations. METHODS: The author uses an evidence-based approach in analyzing the data both supporting and condemning the continued use of amalgam restorations. He reviewed the articles from both peer-reviewed and non-peer-reviewed sources and evaluated their relevance, research design and statistical analysis, as well as whether the conclusions follow from the data. CONCLUSIONS: There are numerous logical and methodological errors in the anti-amalgam literature. The author concludes that the evidence supporting the safety of amalgam restorations is compelling. CLINICAL IMPLICATIONS: Amalgam restorations remain safe and effective. Dentists should educate patients and other health care professionals who may be mistakenly concerned about amalgam safety. PMID- 11258093 TI - The ethics of health care professionals' opinions for hire. AB - BACKGROUND: Serving as a forensic consultant or medical expert witness is a professional duty and social responsibility. While objectivity is a hallmark of ethical health care evaluation, conflict arises when medical experts exhibit bias and serve the hiring party's interests instead of the public's. METHODS: The authors define different types of expert witnesses and examine the means for improving the quality of testimony. They evaluate professional association codes of ethics, health care education, neutral medical experts and the field of bioethics. RESULTS: Incorporating case-based analysis of medical expert testimony in the education of health care professionals may elevate the caliber of testimony. Court-appointed neutral experts may overcome some of the problems inherent in professional and product liability litigation. Neutral bioethicists may play a constructive role in mediating medicolegal disputes. Adopting strong professional association codes of ethics requires the consent of all members, and loss of membership for ethics violations may not be a powerful enough deterrent to biased testimony. CONCLUSION: Biased testimony contributes to scientifically unfounded liability verdicts. The public ultimately pays for huge monetary settlements via higher costs for all goods and services. Financial incentives in the current professional and product liability system will make it difficult to institute court-appointed neutral expert panels on a widespread basis. PRACTICE IMPLICATIONS: Dentists should view themselves as clinicians and evaluators. Rendering expert opinions on forensic matters is an ethical part of dental practice. By doing so with honesty and objectivity, dentists serve as a bridge to justice and fulfill a social responsibility. PMID- 11258095 TI - Restorative dentistry for pediatric teeth. State of the art 2001. PMID- 11258096 TI - Courts divided on AWP laws. PMID- 11258094 TI - Evaluation of a bioadhesive device for the management of aphthous ulcers. AB - BACKGROUND: Aphthous ulcers are common and painful. Current treatments are palliative and focused on pain reduction. This article reports on the clinical trials of a novel, bioadhesive treatment modality. METHODS: Formulations of 2 octyl cyanoacrylate, or 2-OCA, tissue adhesive were tested in two blinded, sham controlled studies. A total of 200 patients with a single, painful aphthous ulcer were entered. In the first study, the investigators applied the tissue adhesive to the aphthous ulcers; in the second trial, the subjects themselves applied the tissue adhesive to their ulcers. The authors evaluated the safety, pain reduction and healing times associated with the bioadhesive. RESULTS: The bioadhesives were found to be safe with no significant adverse events. The short- and long-term pain reduction achieved with an investigator-applied adhesive was significant compared with that achieved with a sham device (P = .024 and P = .036, respectively). The investigator-applied adhesive also demonstrated a significant reduction in healing time over the sham device (P = .021). In the definitive trial, in which the subjects themselves applied the tissue adhesive, pain reduction with a predicate device approved by the U.S. Food and Drug Administration and with the bioadhesive was significantly better than with a sham application (P < .05). The active devices were not statistically different from each other (P = .37). No difference in healing time was evident between devices and the sham. CONCLUSIONS: The formulations of 2-OCA tissue adhesives tested were safe and demonstrated statistically significant pain reduction when applied by either the investigators or the subjects. CLINICAL IMPLICATIONS: Our clinical trials indicate that these novel tissue adhesives could be used as nonprescription, over-the-counter devices to provide significant pain relief for patients suffering from aphthous ulcers. PMID- 11258097 TI - Getting to the root of endodontic (root canal) treatments. The goal: preserving the tooth. PMID- 11258098 TI - Pasteur, Pastorians, and the dawn of immunology: the importance of specificity. AB - Throughout his career, the problems that attracted Louis Pasteur almost invariably involved considerations of specificity of structure and/or of action. Thus, his work on asymmetric crystals showed that chemical form not only specifies crystalline structure, but affects the affinity of ferments as well. In his studies of diseases of silkworms, of beer, and of wine, he could unerringly distinguish with the microscope the specific agents of disease. From this emerged his concept of the specificity of species and against the nonspecificity of spontaneous generation, whence the germ theory of disease. It was in the new field of immunology, however, where the manifestations of an exquisite specificity were most clearly seen. Here, Pasteur's vaccines worked because he chose the specific pathogen in order to induce a specific immunity, and he succeeded each time. But the two most prominent Pastorian successors in immunology, Elie Metchnikoff and Jules Bordet, were not equally successful. Although each contributed significantly to the birth of immunology, each advanced a theory that neglected the principle of specificity and paid a price in consequence. Metchnikoff's phagocytic theory of immunity could not survive the demonstrable specificity of humoral antibodies, while Bordet's physical adsorptive concept of the antibody-cell interaction quickly fell to Paul Ehrlich's demonstration of the stereochemical determination of immunological specificity. PMID- 11258099 TI - The heuristic function of 'error' in the scientific methodology of Louis Pasteur: the case of the silkworm diseases. AB - With the aid of the Cahiers de laboratoire, the Correspondence and, of course, the Oeuvre de Pasteur, this work reconstructs the extraordinary scientific undertakings of the great French scientist in his study of silkworm diseases. The focus of this study consists in the attempt to explain the initial perplexing behaviour of Pasteur, even in the presence of correct interpretations regarding the causes of these diseases (cfr. the results obtained by Bechamp); for a good three years he insisted on maintaining that the aetiology of silkworm diseases could not be attributed to pathogenetic germs from outside. And this was in spite of the fact that previously (through fermentation and spontaneous generation) he had been able to demonstrate the importance of microorganisms in biological processes. Finally it is intended to highlight the extraordinary methodological depth of that initial 'error', which was capable of paving the way for the future conquests of Pasteur in the field of aetiology and the prevention of infectious diseases. PMID- 11258100 TI - Linking cause and disease in the laboratory: Robert Koch's method of superimposing visual and 'functional' representations of bacteria. AB - Robert Koch based his claim that specific microorganisms cause particular diseases on laboratory studies. This paper examines how Koch set up a plausible line of argument by using special methods of representing bacteria. One kind of representation consisted in making the bacteria visible; the other mode of representation was based on disease phenomena. Using a range of techniques of isolating and controlling microorganisms, Koch combined these different modes of representation in a way that made his claims convincing. Thus, the microorganism as a specific cause of disease emerged through a chain of repeated processes of selection and representation in the laboratory. PMID- 11258102 TI - Pasteur, Koch and American bacteriology. AB - This study traces American awareness of the work of Louis Pasteur and Robert Koch from the 1860s to the 1890s. In the years before the Civil War, American interest in germ theories had appeared at times of epidemics and persisted to a limited extent among physician-microscopists. Discussions of Pasteur's work occurred primarily in the context of spontaneous generation and antisepsis. Few Americans imitated his work on immunology or studied with Pasteur, but his work on immunity influenced their faith in the potential of bacteriology as a solution to problems of infectious disease. Koch's discoveries of the bacterial agents of tuberculosis and cholera stimulated American medical and public health interest in bacteriology in a more practical way. Americans learned Koch's methods by taking his courses and imported them directly into their own laboratories. A context of enthusiasm for science, educational reform, and problems of infectious disease associated with urbanization and changes in agriculture aided the growth of bacteriology in the American context. PMID- 11258101 TI - Money and microbes: Robert Koch, tuberculin and the Foundation of the Institute for Infectious Diseases in Berlin in 1891. AB - Starting from an assessment of how far Robert Koch's bacteriology had developed in the late 1880s this paper attempts to analyse different aspects of the process that led to the foundation of the Berlin Institute for Infectious Diseases in 1891. With the development of his supposed cure against tuberculosis, tuberculin, Koch attempted to give his research a new direction, earn a fortune with the profits and become more independent of Prussian government officials who, up to that point, had had a major influence on his career. In the period following the presentation of the cure in autumn 1890, however, it became clear that tuberculin's value in treatment was at most dubious. Thus, the failure of tuberculin meant that Koch had to drop his own plans and accommodate those of the Prussian Ministry of Culture. As a result he assumed directorship of the newly founded Institute for Infectious Diseases in Berlin. Even though this was definitely a prestigious position it reaffirmed Koch's dependency on Prussian government officials and was by no means the kind of institution he had aimed for at the outset. PMID- 11258103 TI - Marketplace. Doctors subject to malpractice cases even when the person they treat isn't a patient. PMID- 11258104 TI - [Modern pacemaker therapy]. PMID- 11258105 TI - [Prevention of atrial arrhythmias by pacing]. AB - BACKGROUND: Atrial fibrillation is the most frequent arrhythmia. It can impair quality of life considerably. Due to thromboembolic complications it contributes to the patients' morbidity and mortality and to the costs for their medical treatment. PREVENTION: In chronic atrial fibrillation there is a need for adequate anticoagulation and heart rate control. In paroxysmal and intermittent atrial fibrillation it should be sought to prevent its progression to chronic atrial fibrillation. Since atrial fibrillation initiates negative processes of remodeling within the atrial myocardium, it has the tendency to perpetuate itself. From a theoretical point of view, it can be expected that all means which prevent episodes of atrial fibrillation or which terminate it immediately after its onset, are able to prevent or at least to delay the progression to chronic atrial fibrillation. Pharmacologic treatment is usually used to prevent recurrences of atrial fibrillation. Based on the actual data it can also be expected that pacemakers with special preventive pacing algorithms are able to reduce the atrial arrhythmic burden. Besides consequent overdrive pacing, more sophisticated algorithms like "suppression of premature atrial contractions", "post exercise response", "automatic rest rate" or "post mode-switch pacing" have been developed. They can be applied either alone or in combination with special lead positions (interatrial septal pacing or pacing of the triangle of Koch) or special stimulation configurations like dual site right atrial pacing or biatrial pacing. These pacing strategies cover the most relevant onset mechanisms of atrial fibrillation. Furthermore, there are algorithms to treat atrial tachyarrhythmias actively by antitachycardia pacing (ATP). First clinical results have shown that about 2/3 of the diagnosed atrial tachyarrhythmias could be terminated by these means immediately after their onset. ONGOING TRIALS: This article gives an overview over the principles of pacing in the management of atrial arrhythmias and ongoing clinical trials in this field. Before a definite judgement on the clinical relevance of these new preventive and therapeutic pacing strategies can be given, the results of these ongoing controlled clinical studies have to be analyzed. PMID- 11258106 TI - [Mode-switching algorithms: programming and usefulness]. AB - BACKGROUND: Automatic mode switching is defined as the ability of a pacemaker to reprogram itself from tracking to non-tracking mode in response to atrial tachyarrhythmias, and to regain tracking mode as soon as the tachyarrhythmia terminates. In contrast to upper rate behavior, mode switching does not only limit atrial tracking at a certain rate but actively drives the ventricular pacing rate back to lower rate or sensor rate as long as the atrial tachyarrhythmia persists. In contrast to DDD with mode switch, AV synchrony may be lost in DDIR mode if the sinus rate exceeds the sensor rate. DDD pacing with mode switching represents a valuable option in patients with AV block and paroxysmal atrial tachyarrhythmias. It may prevent the transition from paroxysmal to permanent atrial fibrillation after AV node ablation to a higher extent than VVI(R) pacing. On the other hand, patients with sinus node disease and normal AV conduction may benefit from DDIR mode with long AV interval. Mode switching should provide a rapid, sensitive and specific detection of atrial tachyarrhythmias, fast switch to non-tracking mode without ventricular pacing at the upper rate limit, adequate ventricular rate during the atrial tachyarrhythmia, rapid, sensitive and specific detection of conversion to sinus rhythm and fast switch back to tracking mode. In addition, oscillations between DDD and DDI mode with sudden ventricular rate changes should be avoided. MODE SWITCHING ALGORITHMS: To achieve these aims, different mode-switching algorithms have been developed which all show specific disadvantages: reliable but slow response to atrial tachyarrhythmias, fast but unspecific switch to non-tracking mode, mode oscillations, inclination to inadequate mode-switching due to ventricular far-field sensing, failure to perform modeswitching during atrial flutter or intermittent atrial undersensing. Some of these problems can be avoided by careful atrial lead implantation providing atrial signals above 2 mV and avoiding ventricular far-field signals. Programming of mode-switching related parameters (e.g. atrial rate and number of fast beats required for mode switch), atrial blanking times, and atrial sensitivity can solve some of the problems with mode switching. Clinical results show a strong influence of device programming and atrial undersensing on mode-switching performance. Some data suggest a superiority of fast mode-switching algorithms with regard to clinical symptoms. However, loss of AV synchrony during sinus rhythm due to premature or inadequate mode switching may limit the benefit of fast mode switching. FURTHER DEVELOPMENTS: Improved performance may be achieved by a combination of different mode-switching algorithms (e.g. one algorithm for detection of atrial fibrillation, another one for detection of atrial flutter). In addition, programmability of several algorithms (e.g. mean atrial rate, beat-to-beat, x out of y) within the same device and atrial cycle-dependent sensitivity adjustment similar to automatic gain control in implantable defibrillators may further increase the clinical use of automatic mode switching. PMID- 11258107 TI - Improved detection and analysis of sensed and paced events in dual chamber pacemakers with extended memory function. A prospective multicenter trial in 626 patients. AB - PATIENTS AND METHODS: This prospective study analyzed the incidence of atrial arrhythmias in a population of 626 patients in 173 medical centers of eleven European countries and Japan with indication for a dual chamber pacemaker system. The accuracy of the new Automatic Interpretation for Diagnosis Assistance (AIDA) program which is included in Chorus pacemakers was evaluated and the AIDA analysis was compared to and proven with Holter monitoring. Data stored in the pacemakers' memories for the first 24 hours (D1) were compared with simultaneously recorded 24-hour surface electrocardiograms, and data stored over the following 28 days (D28) were examined against reported intercurrent symptoms. RESULTS: At D1, atrial arrhythmias were detected by AIDA in 60 of 626 patients (12%), consisting of atrial fibrillation (n = 29), atrial flutter (n = 4), and miscellaneous arrhythmias (n = 17), and closely corroborated by Holter monitoring (sensitivity 93.7%, specificity 94.9%). At D28, 149 of 386 patients (49%) had had episodes of automatic mode switch prompted by atrial arrhythmias. Symptoms were reported by 81 patients (54%), 92 (62%) had no histories of atrial arrhythmias, and 57 patients (38%) were neither symptomatic nor had histories of atrial arrhythmias. An inverse relationship was found between the number of atrial paced events and the occurrence of atrial arrhythmias (p < 0.001). A history of atrial arrhythmias and older age were associated with a higher risk of atrial arrhythmias (p < 0.05). In contrast, gender, hypertension, concomitant heart disease, or type of atrial lead fixation system were not related with the occurrence of atrial arrhythmias. CONCLUSION: AIDA allowed to confirm, or disprove, the occurrence of atrial arrhythmias as a source of symptoms reported during long-term follow-up. It could also be used to examine the efficacy of antiarrhythmic therapy, and be of assistance when weighing the needs for anticoagulation in patients experiencing asymptomatic atrial arrhythmias. PMID- 11258108 TI - [Usefulness of a VDD defibrillation electrode in recording atrial electrograms during atrial flutter and atrial fibrillation]. AB - BACKGROUND: Monitoring of atrial signals improves the accuracy in identifying supraventricular tachyarrhythmias to prevent inappropriate therapies in patients with implantable cardioverter-defibrillators (ICD). Since complications due to the additional atrial lead were found in dual chamber ICD systems with 2 leads, we designed a single-pass VDD-lead for use with dual chamber ICDs. PATIENTS AND METHODS: After promising animal experiments in a German multicenter study a prototype VDD lead (single-coil defibrillation electrode with 2 additional fractally coated rings for bipolar sensing in the atrium) was temporarily used in 20 patients. Atrial and ventricular signals were recorded during sinus rhythm, atrial flutter, atrial fibrillation and ventricular tachycardia or ventricular fibrillation. Terminations of ventricular arrhythmias were performed by internal DC shock. RESULTS: The implantation of the electrode was successful in 18 of 20 patients. Mean atrial pacing threshold was 2.45 +/- 0.9 V/0.5 ms, mean atrial impedance was 215 +/- 31 Ohm. Atrial amplitudes were greater during sinus rhythm (2.7 +/- 1.6 mV) than during atrial flutter (1.36 +/- 0.28 mV, p < 0.05) or atrial fibrillation (0.92 +/- 0.29 mV, p < 0.01). During ventricular fibrillation atrial "sinus"-signals had significantly (p < 0.01) lower amplitudes than during sinus rhythm. Mean ventricular sensing was 13.3 +/- 7.9 mV, mean ventricular impedance was 577 +/- 64 Ohm. Defibrillation was successful with 20 J shock. 99.6% of P waves could be detected in sinus rhythm and 85 +/- 9.9% of flutter waves during atrial flutter. During atrial fibrillation 55% of atrial signals could be detected without modification of the signal amplifier. CONCLUSIONS: A new designed VDD dual chamber electrode provides stable detection of atrial and ventricular signals during sinus rhythm and atrial flutter. For reliable detection of atrial fibrillation modifications of the signal amplifier are necessary. PMID- 11258109 TI - [Heart rate variability preceding the onset of atrial fibrillation]. AB - BACKGROUND: The occurrence of paroxysmal atrial fibrillation is related to changes in autonomic tone. Vagally mediated atrial fibrillation predominantly occurs at night in young male patients without history of structural heart disease. In contrast, sympathetically mediated atrial fibrillation is typically triggered by stress. ANALYSIS OF HEART RATE VARIABILITY: Heart rate variability usually measured by Holter monitoring can determine changes in autonomic tone immediately preceding the onset of atrial fibrillation. Studies on this topic found divergent results on the incidence of atrial fibrillation mediated by changes in autonomic tone. Dependent on the results of different studies, night time episodes of idiopathic atrial fibrillation either are vagally or sympathetically mediated. A sympathetic predominance is found in patients after coronary bypass surgery, concordantly. The inconsistency of these findings points to the fact that not only one mechanism, but more complex changes in autonomic tone are responsible for the occurrence of atrial fibrillation in many cases. In modern pacemakers and implantable cardioverter-defibrillators, PP or RR intervals can be stored automatically before the onset of an arrhythmia. By use of these stored intervals, time-domain parameters of heart rate variability can be calculated. We determined changes in short-term heart rate variability (10-second intervals) by analyzing 26 episodes of PP intervals sampled over the last 2 minutes before onset of atrial fibrillation by a modern dual chamber pacemaker. We observed a significant increase of the standard deviation of PP intervals (SDNN10s) as well as of the mean square route of the squared PP intervals (rMSSD10s) within the last 10-second interval before onset of atrial fibrillation (p < 0.05). This finding points to changes in autonomic tone-immediately preceding the onset of atrial fibrillation. CONCLUSION: The preliminarity of these findings and the use of yet not validated short intervals for determination of heart rate variability does not allow to draw pathogenetic or even therapeutic conclusions from these findings. PMID- 11258110 TI - [A new dual-chamber defibrillation system: duration, frequency and circadian variation of recurrent supraventricular tachyarrhythmias]. AB - BACKGROUND: The treatment of concomitant atrial tachyarrhythmias in patients with malignant ventricular tachyarrhythmias is a major challenge for new defibrillator devices. Atrial fibrillation is not only responsible for inappropriate ventricular therapies, but also reduced left ventricular performance, especially in patients with heart failure and severely depressed left ventricular function. Furthermore, it is a strong risk factor for the development of thromboembolism. NEW SYSTEM: A new dual-chamber implantable defibrillator is capable of tiered atrial therapies for both regular and irregular atrial tachyarrhythmias. In first investigations a high sensitivity and specificity could be shown as well as a promising therapy efficacy of atrial antitachycardia ramp and burst pacing for the treatment of atrial tachycardias. Atrial ramp pacing has shown to be successful for regular atrial tachyarrhythmias in up to 60 to 70% of all episodes. The results have supported a programming of a high first shock energy for treatment of atrial fibrillation. The incidence of atrial fibrillation in patients with a history of atrial fibrillation or without is much higher in the present investigated patient populations than expected. CONCLUSION: The more complicated and subtle new dual-chamber detection algorithm has proven to be safe and effective both for the detection of ventricular tachycardia but also in terms of an increase of specificity and a reduction of inappropriate ventricular therapies for atrial tachyarrhythmias. PMID- 11258111 TI - Microgenerators for energy autarkic pacemakers and defibrillators: fact or fiction? AB - BACKGROUND: Implantable devices for medical use like permanent pacemakers, defibrillators, and fluid pumps depend on an energy provided by batteries. Unfortunately, the battery usually determines the duration of life of these devices, while technical problems occur infrequent. Device replacement for battery exhaustion requires surgical procedures and account for up to 1/3 of all pacemakers sold. Attempts to provide unlimited power support using radio transmission, nuclear energy etc. did not gain clinical acceptance. METHOD: We therefore evaluated the potential role of a microgenerator (designed for use in wrist watches) to recharge pacemaker batteries. We used the Epson-Seiko Caliber 5M22 that uses a "Gold-Cap" for energy storage. The mass of the actuator is 1.6 g and an angle of > 10 degrees is needed to overcome friction. Output at a rotor frequency of 200 Hz is 1.8 mWatt To measure the power provided, various experiments were made with the microgenerator taped to the chest of a normal person working in an office. Range of 11 experiments over 8 hours each was 0.2 to 3.1 microWatt (median 0.5 microWatt). Therefore, the power generated was 10 to 100 times less than the calculated power needed to recharge a typical pacemaker battery. A second type of generator (Mondaine, Zurich, Switzerland) with less mechanical parts, available in a "black box" version only, generated not more power. CONCLUSION: Thus, commercially available, yet not optimized microgenerators provided only between 1 to 10% of the power requirements of a pacemaker. However, modifications in design and mainly the orientation and weight of the actuator to generate more power from the G-forces during walking, would result in a more meaningful energy output. PMID- 11258112 TI - [Support of spontaneous atrioventricular conduction in patients with DDR(R) pacemakers: effectiveness and safety]. AB - AIM: In a prospective and randomized multicenter study using a cross-over protocol we compared the efficacy and the safety of the ELA medical mode-switch algorithm (DDD/AMC = DDD to AAI) to conventional DDD stimulation in patients with spontaneous AV conduction. PATIENTS AND METHOD: Forty-eight patients with a mean age of 67 +/- 13 years were included. Underlying heart disease was present in 54%. Pacemaker indications were paroxysmal AV block (21%), sick-sinus syndrome (46%), paroxysmal AV block + sick-sinus syndrome (31%) and tachycardia bradycardia syndrome (8%). Patients were excluded from the study in case of a permanent 1st to 3rd degree AV block, a right bundle-branch block with QRS > 120 ms, severe coronary heart disease or idiopathic cardiomyopathy. The programming of the pacemaker was randomized to either DDD/AMC or DDD and was crossed over after 1 month. The AV interval (AVI) which was programmed in conventional DDD pacing was calculated as AVI = PR (or AR) + 30 ms at rest or as AVI = PR (or AR) 50 ms during exercise. When the DDD/AMC mode was programmed, the AV interval was calculated automatically. We analyzed the AV interval, the frequency of ventricular pacing, the number of pacemaker-induced tachycardias, the number of atrial tachyarrhythmias, and the final programming which was left to the physician's choice. RESULTS: The AV interval after conventional DDD stimulation was 201 +/- 38 ms vs 195 +/- 28 ms with DDD/AMC (p = ns). Ventricular stimulation was significantly less often in the DDD/AMC mode than in the DDD mode (15 +/- 17% vs 48 +/- 37%, p < 0.001). Thereby the DDD/AMC algorithm led to a 69% reduction of ventricular pacing which means an approximately 5.5 months prolongation of the battery lifetime. There was no significant difference in the incidence of pacemaker-induced tachycardias. At the end of the study 77% of the physicians programmed the DDD/AMC mode. CONCLUSION: The analyzed DDD/AMC mode-switch algorithm leads to a significant reduction of ventricular pacing in patients with spontaneous AV conduction or with only paroxysmal AV block. Thereby the battery lifetime is prolonged and the incidence of complications due to ventricular pacing can be reduced. PMID- 11258113 TI - [Sensor-induced rate adaptation: sensor systems and tuning. Manual versus automatic tuning]. AB - BACKGROUND: Rate adaptive stimulation represents an established therapeutic option in the management of cardiac rhythm disturbance. To realize a physiologic heart rate in patients with chronotopic incompetence several different sensor systems were developed. Due to the requirements of a physiologic pacemaker with all algorithms and therapeutic functions complex pacemaker systems were created. The realization of an automatic rate optimization was given by self adjusting algorithm or new sensor systems (closed loop systems). Comparing data respecting automatic vs manual sensor adjustment are not present. The automatic sensor adjustment was examined only in small patient groups. CONCLUSION: This studies indicated that most patients could be adjusted by the automatic function. Nevertheless, out of technical reasons a small group of patients could not be adjusted. The verification of the correct individual rate response in all patients treated with rate response pacemakers belongs to the physicians obligation. PMID- 11258114 TI - [Experiences with a new transvenous electrode for left ventricular stimulation]. AB - BACKGROUND: Left ventricular and biventricular pacing has recently been introduced as a new therapy for chronic heart failure in selected patients. We report our initial experience with a new electrode for transvenous left epicardial pacing via tributaries of the coronary sinus. PATIENTS AND METHOD: Inclusion criteria were: chronic heart failure NYHA > or = II, QRS-duration > 120 ms, left ventricular ejection fraction < 35%. Dual chamber pacemakers (CPI Contak TR) or defibrillators (CPI Contak CD) designed for atrial triggered biventricular stimulation were implanted in conjunction with the CPI Easytrak-lead for left ventricular pacing in a coronary vein. Lead placement was achieved via a subclavian vein access and a preformed guiding catheter for coronary sinus insertion. RESULTS: In 13 of 16 patients (81%) the left ventricular lead was implanted successfully in a mid to distal posterior or anterolateral vein. Lead insertion could not be achieved in 2 patients with significant cardiomegaly and right atrial enlargement (12.5%), while 1 patient with a history of myocardial infarction and small anterior ventricular aneurysm had inacceptable high left ventricular pacing thresholds intraoperatively. The implantation was well tolerated by all patients without complications. There was no case of lead dysfunction (mean follow-up time: 142 +/- 126 days). Intraoperative electrode measurements and chronic parameters (> or = 3 months, n = 8) are given in Table 1. CONCLUSION: In the past left ventricular pacing has mainly been achieved by epicardially placed electrodes after thoracotomy with conventional electrodes. This new approach for chronic left ventricular pacing uses the familiar transvenous over-the-wire technique in combination with a newly developed guiding catheter and electrode for pacing in left epicardial veins. Lead placement was shown to be safe and success rate was higher than in previous reports with standard electrodes. We conclude that left epicardial lead placement with the over-the-wire technique and a preformed guiding catheter for coronary sinus access presents as a safe and maybe more efficient method for left ventricular pacing. PMID- 11258115 TI - [Cardiac resynchronization therapy in terminal heart failure: current status and prospects]. AB - BACKGROUND: With regard to epidemiological aspects, heart failure has been shown an increasing incidence in constrast to coronary artery disease which counts decreasingly due to secondary and primary prevention. The present data show an incidence for heart failure of 2% per year. 4-5 million people are newly affected by the disease. The prognosis is limited after diagnosis is confirmed. According to the US Framingham study, the median life expectancy is 3.2 and 5.4 after diagnosis for male and female, respectively. For patients in an advanced stage of the disease the mortality rate is 27% within 3 years. AV SEQUENTIAL PACING: The introduction of AV sequential pacing by the Austrian group of Margarete Hochleitner in 1992 showed an improved left ventricular systolic function, an improved clinical benefit in terms of NYHA classification, an enhanced left ventricular ejection fraction, an improved systolic and diastolic blood pressure, a reduction of the heart-chest relationship as well as a reduction of the resting hart rate and the echocardiographic dimension parameters. STUDIES: First experimental approach for biventricular stimulation, which means the simultaneous activation of the right and the left chamber, relied on the observation of a distorted ventricular activation due to the presence of a bundle branch block. The bundle branch block is a result of the dilatation of the myocardial fibers, death of myocardial cells which are replaced by fibrous tissue. Resynchronization of both ventricles was associated with an improved left ventricular function and improved diastolic relaxation. Clinical studies of patients with heart failure, severe left ventricular systolic dysfunction, and left bundle branch block have shown that systolic function can be improved by electrically stimulating the site of late activation. The magnitude of the improvement seems to be associated with the duration of the intrinsic surface QRS complex and whether the ventricle ipsilateral with the conduction defect is stimulated. The effect of ventricular resynchronization therapy was optimized by timing of atrioventricular activation associated with a decrease in both systolic and diastolic mitral regurgitation. CONCLUSION: The prognosis of patients with end-stage heart failure is limited by two determinants: myocardial pump failure and sudden (arrhythmogenic) cardiac death. Due to the fact that the incidence for sudden cardiac death is considerably high in patients with end-stage heart failure it is reasonable to include the implantation of cardioverters/defibrillaters (ICD) in the concept of biventricular stimulation. PMID- 11258116 TI - [Ambulatory non-center occupation-related rehabilitation of patients with coronary disease: an alternative in rural areas, too?]. PMID- 11258117 TI - Haptic after-effect of successively touched curved surfaces. AB - A flat surface is more often judged to be convex after the touching of a concave surface than after the touching of a convex surface. This haptic after-effect increases with the time of contact with the curved surface till it saturates, and it decreases with the time-lapse between the touching of the first surface and the next one. In this paper, the haptic after-effect of two successively touched spherically curved surfaces is investigated. It is found that both surfaces contribute to the after-effect, but the after-effect is not additive. The time course of the after-effect of two successive surfaces can be described by a first order integrator with a single time constant of about 7 s and an amplitude equal to the difference between the saturation levels of the after-effects of the two surfaces when measured in isolation. The new saturation level is therefore equal to that of the after-effect of the most recently touched surface. PMID- 11258118 TI - The role of internal reference prices in consumers' willingness to pay judgments: Thaler's Beer Pricing Task revisited. AB - Alternative reference prices, either displayed in the environment (external) or recalled from memory (internal) are known to influence consumer judgments and decisions. In one line of previous research, internal reference prices have been defined in terms of general price expectations. However, Thaler (Marketing Science 4 (1985) 199; Journal of Behavioral Decision Making 12 (1999) 183) defined them as fair prices expected from specific types of seller. Using a Beer Pricing Task, he found that seller context had a substantial effect on willingness to pay, and concluded that this was due to specific internal reference prices evoked by specific contexts. In a think aloud study using the same task (N = 48), we found only a marginal effect of seller context. In a second study using the Beer Pricing Task and seven analogous ones (N = 144), general internal reference prices were estimated by asking people what they normally paid for various commodities. Both general internal reference prices and seller context influenced willingness to pay, although the effect of the latter was again rather small. We conclude that general internal reference prices have a greater impact in these scenarios than specific ones, because of the lower cognitive load involved in their storage and retrieval. PMID- 11258119 TI - Temporal-spatial memory: retrieval of spatial information does not reduce recency. AB - Factors influencing the shape of serial position curves in non-verbal serial short-term memory were examined, using a task testing memory for the position of dots. Similar recency slopes were found when both position and order were recalled (Experiment 1A) and when order only was required (Experiment 1B). This observation was confirmed and tested further in conditions requiring the same encoding but different amounts of spatial information at retrieval (Experiment 2). However, Experiment 2 also revealed an effect of spatial information retrieval on the overall level of memory for recency items. Overall, the results indicate that spatial items produce bow-shaped serial positions curves in tasks requiring the maintenance of order information and that recency is affected by the demand on spatial information retrieval in terms of the overall level of performance but not in terms of the recency slope. These findings are contrary to what is found in the literature on serial verbal recall when both item and order information are required. PMID- 11258120 TI - Propositional reasoning and working memory: the role of prior training and pragmatic content. AB - Working memory involvement in propositional reasoning was explored after different kinds of training. The training conditions aimed to reduce the impact of non-analytic heuristics and to enhance analytic inference processes according to mental logic theories, the mental model theory, and the theory of pragmatic reasoning schemata. Following an initial training phase, secondary task interference was investigated using concurrent spatial tapping (Experiment 1), random number generation (Experiment 2), and articulatory suppression (Experiment 3). A training condition practicing the construction and use of mental models via a truth table task increased the disruption of reasoning performance by random number generation and articulatory suppression, whereas the other training conditions did not affect susceptibility to secondary task interference. The results corroborate implications of the mental model theory of reasoning. PMID- 11258121 TI - [Human lens cells in culture. I. Isolation of adult lens epithelial cells from lens capsule preparations and reactivation of nucleus-containing fiber cells]. AB - BACKGROUND: Bovine lens epithelial cells in culture revealed a high sensitivity against micromolar concentrations of linoleic acid. To prove the assumption that unsaturated free fatty acids are risk factors for cataractogenesis, human lens cell lines are needed. Furthermore, the reactivation of nucleus-containing fiber cells to mitotic growth may hint at their role in after cataract genesis. MATERIAL AND METHODS: Epithelium-capsule-preparations obtained by capsulorhexis were cultured in serum containing medium. Subculturing of these adult human lens epithelial cells was done by trypsinization. Fiber cell bundles from the equator region of a fetal human lens were transferred into culture medium. Aggregates of nucleus containing fiber cells were isolated from floating fiber cell bundles by trypsinization. Subculturing and cryoconservation of suitable cell lines. RESULTS: Primary culture of epithelium-capsule-preparations results in flattening, migration and proliferation of adult human lens epithelial cells. Nucleus containing fiber cells were reactivated to mitotic growth after adhesion to a suitable substratum. Established cell lines were received from adult human lens epithelial cells and fetal human fiber cells after repeated subculturing. CONCLUSIONS: Lens-capsule-preparations available from cataract surgery are well suited for the isolation of human lens cell lines, which were needed for testing cytotoxicity of drugs and for tracing of cataractogenic risk factors. The finding that nucleus containing fiber cells from the equator of human lenses can be reactivated to proliferating cells let us suppose, that these cells, which can not be removed easily from the posterior lens capsule, contribute to the after cataract formation. PMID- 11258122 TI - [Effect of intraocular lens design on posterior capsule opacification: an in vitro model]. AB - BACKGROUND: The development of posterior capsule opacification (PCO) is one of the commonest complications of modern cataract surgery. The various designs of intraocular lenses (IOL) seem to exert a barrier effect on the proliferation and migration of lens epithelial cells and the following development of PCO. METHODS: We set up a cell culture model (advanced 3D capsular bag model) and investigated six differently designed IOL made of different materials as to their effect on cell proliferation. Proliferation and migration of the cells were analysed and documented over a period of 28 days. A cell viability test using the LIVE/DEAD kit (Molecular Probes) was carried out at the end of the investigation. RESULTS: In all tests, lens epithelial cells adhered to and migrated onto the capsular bag. During the culture period, lens epithelial cells migrated only to the optical rim of two of the implanted IOL. On the other four, lens epithelial cells migrated further and covered the whole optical area of the IOL. CONCLUSIONS: Certain lens designs seem to have a reducing effect on the development of PCO. Our advanced in vitro capsular bag model is a suitable cell culture model for the investigation of the reducing effect of various IOL on the development of PCO. PMID- 11258123 TI - [Vitamins C and E protect cultures of bovine lens epithelium from the damaging effects of blue light (430 nm) and UVA light (300-400 nm)]. AB - BACKGROUND: In 1992 Radlmaier showed that catalase can protect bovine lens epithelial cultures from blue light. The following experiments have the aim to examine, if vitamin C and E have protective functions, also. MATERIAL AND METHODS: 367 cultures of bovine lens epithelial cells were incubated in Medium 199 added by 20% foetal calf serum FCS, by 125 I.U./ml penicillin, by 125 mg/ml Streptomycin, by 0.31 microgram/ml amphotericin B, by 2% L-Glutamin (200 mM), and by 1.25% Hepesbuffer. The incubation temperature was kept at 36 degrees C at a pCO2 of 5%. In order to avoid secondary morphologic alterations, we experimented only on the second to the third subculture. Light exposure followed three days after addition of the substances such as Cebion 500 (R) ad injection (6 mg/dl) and E-Vicotrat (R) ad injection (0.5 mg/dl). The blue light (420-430 nm, Draeger) was applied in a baby bed at 36 degrees C for four hours (1.1 mW/cm). The UV-A lamp was constructed and described by Heller [7]; at a wave length of 300-400 nm, the irradiation time was 10 minutes at 100 mW/cm2. In morphologic evaluation we looked for criteria such as cell diameter, cellular wall alterations, cellular inclusions and vacuoles. The cell count was done after staining with the vital dye trypan-blue in Neubauer's chamber. In statistic evaluation we used the pair comparison of Tuckey and Kruskall-Wallis-Test. RESULTS: After addition of 0.5 mg/ml vitamin E: In 89 cultures, morphologic evaluation and cell count showed a significant protection against light toxicity: After Uv-A-irradiation we counted 50% more living cells and after blue light 30% more. After addition of 6 mg/ml Cebion vitamin C: Morphologic evaluation and cell count of 223 cultures showed them also to be significantly protected: after irradiation by UV-A we counted 25% more and after exposition to blue light 15% more living cells. CONCLUSION: The demonstrated experiments significantly showed, that vitamin C and vitamin E can protect lens epithelial cultures from toxic stress by blue and by UV-A light and might delay cataract formation in man. PMID- 11258124 TI - [Early childhood cataract in hereditary UDP-galactose-4-epimerase deficiency--a case report]. AB - BACKGROUND: Increased plasma galactitol levels may lead to development of bilateral pediatric cataract. PATIENT: A 3-year-old boy was found to suffer from a bilateral zonular cataract. Extracapsular lensectomy with posterior capsulotomy, transpupillar anterior vitrectomy and posterior chamber lens implantation were performed during a 4-month-interval. RESULTS: The epimerase activity in red cells of the index patient was found to be significantly decreased (11.2 mumol/h/g Hb; normal range; 19-35). From other family members, such as the brother (16.8), the father (16.0) and the grandfather (15.6), a diminished red cell activity was observed. The mother whose epimerase activity was considerably lower than that of the above mentioned family members (13.3) showed also a zonular bilateral cataract. CONCLUSIONS: Investigation of enzymes and polyols of galactose metabolism as well as consultation of the concerned families are recommended for clarification of cataract development. PMID- 11258125 TI - [Scheimpflug photographic imaging following implantation of anterior and posterior chamber phakic intraocular lenses: preliminary results]. AB - BACKGROUND: For the correction of refractive errors lenticular procedures are increasingly used in addition to corneal refractive surgery. One of those techniques is the implantation of intraocular lenses into phakic eyes (pIOL). Due to the close neighborhood of the implant to delicate intraocular structures, exact positioning and high postoperative stability are required. Scheimpflug photography has been shown to be a suitable instrument for the biometry of the anterior eye segment and the examination of IOL position. PATIENTS AND METHODS: Four anterior chamber phakic IOLs (pIOLs) (Bausch & Lomb NuVita) and 7 posterior chamber pIOLs (Staar ICL) were examined 1 week, 1 month and 3-6 months following implantation. At each examination 1 Scheimpflug slit image and 1 infrared retroillumination image were taken using the anterior eye segment analysis system EAS-1000 (Nidek Co., Gamagori, Japan). Evaluation of the images was performed with a personal computer and the software provided by the manufacturer. The distance of the pIOL to cornea and human lens was calculated and incidence and amount of pIOL rotation around the optical axis and potential crystalline lens opacification were assessed. RESULTS: The distance between the anterior chamber pIOL and the cornea 1 week after implantation was 1.61 +/- 0.10 mm. The distances between the myopic posterior chamber pIOL and the human lens were 0.34 +/- 0.11 mm and between the hyperopic posterior chamber pIOL and the human lens 0.26 and 0.29 mm, respectively. The values were constant over a period of 3-6 months. The pIOL showed no movement or change of position around the optical axis. There was no detectable cataract formation in the human lens. CONCLUSIONS: All implanted phakic anterior and posterior chamber IOLs showed a stable position in the eye within the observation period. Scheimpflug photography is proved to be a useful technique for the postoperative evaluation of the positioning of phakic IOLs. PMID- 11258126 TI - [Can fixation disparity be detected reliably by measurement techniques according to H. J. Haase?]. PMID- 11258127 TI - [What is behind the fixation disparity?]. PMID- 11258128 TI - [Image analysis and Sheimpflug photography of anterior segment of the eye--a review]. AB - BACKGROUND: Light scattering properties of the cornea and lens change in relation to age. This process is influenced by noxious factors which may induce the formation of cataracts. Four different camera types based on the Scheimpflug principle have been developed for documentation and evaluation of light scattering in the eye. Topcon SL-45. Zeiss SLC, Oxford CASE 2000, Nidek EAS 1000. Only the SL-45 and EAS 1000, however, have found a wider application in clinical and experimental studies. MATERIAL AND METHODS: Documentation of the eye with a Scheimpflug system is performed in a dark room after mydriasis induction. The rotating axis of the camera is aligned to the optical axis of the eye with fixation devices. Four meridians have been found appropriate for routine documentation. Image analysis is based on peak height and distance evaluation of 2 layers in the cornea and 10 layers in the lens. Standardization devices allow correction of technical differences in the recording process. RESULTS: Evaluation of the ageing properties of the lens has evidenced typical density developments for each individual layer, the cortex increasing in density earlier than the nucleus. Cataract classification using a Scheimpflug camera provides an objective characterization of cataract type and progression. The outcome of surgical procedures changing the refractive power of the eye can be evaluated with a Scheimpflug camera as well. Its application in studies dealing with the ocular effects of Quinolones and HMG-CoA-reductase inhibitors, has demonstrated that Scheimpflug photography is the method of choice for establishment of a safety profile for a new drug. CONCLUSIONS: Scheimpflug photography has proven to be ideal for objective documentation of the anterior eye segment in human and animal eyes, although both documentation and image analysis demand technical expertise to be reproducible. PMID- 11258129 TI - [Characteristics and frequency of cortical cataracts at an early stage (Reykjavik Eye Study in Iceland)]. AB - BACKGROUND: The frequency and characteristics of cortical cataract localization within a dilated pupillary area were investigated in a population-based cataract epidemiological survey performed in the city of Reykjavik in Iceland. MATERIALS AND METHODS: Among 1045 randomly selected individuals, the right eyes of 277 persons with mild cortical lens opacification (Grade I) were selected for our study. The localization of cortical cataracts was examined using retroillumination images, which were divided into 56 circular and radial sections, and calculations were made of the percentages for each opacified area. A questionnaire was used to record the number of hours spent outdoors during week days, the time of the day and whether the subjects wore spectacles, sunglasses or hats when outside. RESULTS: The percentage of cortical opacification was significantly higher in the lower nasal quadrant than in all other quadrants (p < 0.05). Furthermore, the percentage of opacification localized in the lower nasal quadrant was higher in individuals with a longer history of outdoor activity than those without (p < 0.05). The percentage of opacification in individuals, who have had the habit of wearing sunglasses for outdoor activities lasting longer than half an hour during weekdays in their 20's and 30's was lower in all quadrants and statistically significantly lower in the upper temporal quadrant (p < 0.05) compared to those not wearing sunglasses. CONCLUSION: This suggests that wearing sunglasses is effective in preventing the development and/or progression of cortical cataracts. PMID- 11258130 TI - [Concentrations of diclofenac in aqueous humor and in plasma after i.v. injection in 59 patients undergoing cataract surgery--a prospective study]. AB - BACKGROUND: The mean diclofenac level in the aqueous humor after i.v.-injection should be examined as an example for other systemic applied drugs. The clinical effect is to be evaluated with regard to perioperative anti-inflammatory therapy in patients undergoing cataract surgery. PATIENTS AND METHODS: In this prospective clinical study 59 patients (age 41-86 years) scheduled for phacoemulsification by the tunnel technique, received 75 mg diclofenac-sodium by short time i.v.-infusion (30 min) within an interval between 30 min and 7 h before surgery. Aqueous humor samples and one blood sample were taken at the beginning, and a second blood sample was taken at different time intervals before surgery. RESULTS: The plasma concentration of diclofenac one minute after infusion was 13,961 +/- 4277 ng/ml. The concentration decreased rapidly during the distribution phase (1305 +/- 324 ng/ml after 40 min) and reached after 336 min 419 +/- 277 ng/ml. A concentration of 83.9 ng/ml diclofenac was found for the last sampling point after 353 min in the elimination phase. The mean diclofenac concentration in the aqueous humor reached after about 60 min the highest level (21.7 +/- 12.7 ng/ml). This level decreases to 4.4 +/- 0.4 ng/ml (120 min) and 3.1 +/- 1.3 ng/ml (300 min), respectively. CONCLUSIONS: The calculated mean residence time of diclofenac molecules in aqueous humor of 123 min matched the mean disposition residence time of diclofenac in the body (119 min). PMID- 11258131 TI - [Effect of retinal coagulation status on oxidative metabolite and VEGF in 208 patients with proliferative diabetic retinopathy]. AB - BACKGROUND: Oxidative metabolites and different cytokines are believed to be involved in the pathogenesis of (proliferative) diabetic retinopathy. It was the aim of this study to analyze vitreous body and proliferations of diabetic patients for oxidative metabolites and VEGF und to correlate these values to the retinal coagulation status. PATIENTS AND METHODS: The study was performed in 208 patients vitrectomized for diabetic retinopathy (Type I: n = 114, Type II: n = 94). Grouping of patients was performed according to retinal coagulations status: (1) no or minimal preoperative coagulation [mean coagulation area, CA: 156 mm2], (2) coagulation (scatter laser coagulation und/oder cryopexy) < 3 months before surgery [CA: 589 mm2]. (3) coagulation > 3 months before surgery [CA: 546 mm2]. In the vitreous body and, if present, in the fibrovascular proliferations (Type I: n = 83; Type II: n = 73) the level of lipid peroxides (LPO, measured with two methods) and VEGF was determined. RESULTS: In the vitreous body LPO in group 1 were significantly (P < 0.01 (Type I und II)) higher as compared to other groups. In group 3 LPO were significantly lower as compared to group 2 (P < 0.01 (Typ I) and P < 0.05 (Typ II)). Similar results were observed in the proliferations. In Type I patients VEGF values were significantly (P < 0.01 for group 1 vs. 2 and groups 1/2 vs. 3) reduced following coagulation and coagulation + 3 months. In Type II patients only group 3 was significantly (P < 0.01) different from group 1. In proliferations groups 2 and 3 were significantly different from group 1 (P < 0.05 for Typ I and Type II patients). CONCLUSIONS: The time course of the values leads to the conclusion that oxidative metabolites are able to directly modulate growth activity and to exert this effect via induction of VEGF. This hypothesis has to be confirmed in vitro and by means of a prospective study. PMID- 11258132 TI - [Effect of serum and osmotically active substances on metabolism in 262 tissue cultures of pig's corneas--improved preservation of donor tissue for keratoplasty]. AB - BACKGROUND: The storage time of corneal organ cultures in limited in the closed system mainly used in European eye banks. In addition, swelling during storage reduces the quality of the donor material for keratoplasty. Therefore, dextrane was added in concentrations of 2.5%, 5%, 7.5% and 10% to the culture medium, and the energy producing metabolism was investigated. MATERIALS AND METHODS: The experiments were carried out with 262 pig corneas. MEM with some supplements was used at 31 degrees C. No serum was added, because glucose consumption, lactate production and ATP levels proved to be the same or better than with serum supplement. After 6 and 12 days, the corneas from organ cultures were extracted with perchloric acid. The levels of glucose and lactate in the stroma and of ATP and ADP in the epithelium were analysed with enzymatic-optical tests. RESULTS: Dextrane inhibited glycolysis and the production of ATP in corneal organ cultures during twelve days. With 7.5% and 10% dextrane in the medium, lactate levels in the cornea decreased from 6.09 to 4.5 and 4.6 microM/g H2O instead of increasing. At the same time, glucose increased paradoxical from 1.3 to 3.4 and 2.2 microM/g H2O, respectively. With 5% dextrane, glycolysis operated sufficiently producing an increase of lactate levels from 6.0 to 8.8 microM/g H2O and consuming glucose from levels of 1.27 down to 0.57 microM/g H2O. After 12 days with 7.5% and 10.0% dextrane, ATP levels were reduced, from 4.54 to 0.98 and 0.49 microM/g dry weight, and ATP/ADP ratios from 1.9 to 1.1 and 0.7 respectively. With 2.5% dextrane, the ATP was diminished from 4.5 to 2.2 microM/g dry weight. When 5% dextrane were added to the culture medium, the hydration was at optimum by 4.1, and ATP levels were reduced only from 4.5 to 2.6 microM/g dry weight. Moreover, the ATP/ADP ratios were at 2.1 as good as without dextrane. CONCLUSIONS: From the results it was concluded, that serum free medium may be used, and that permanent dewelling proved to be beneficial for the energy producing metabolism of the cornea in organ culture. Such conditions may improve morphological and metabolic quality of donor material. From previous publications, it was recommended, to use instead of toxic dextran the well tolerated HES (hydroxyethyl starch) and to apply a storage temperature of 21 degrees C, which slowed down glucose consumption without impairing the energy producing metabolism. PMID- 11258133 TI - [Irregularities of the eye: exact correction]. PMID- 11258134 TI - [Anti-ulcer drug pirenzepin: new use as an aid for prevention of myopia?]. PMID- 11258135 TI - [Magnetization transfer contrast (MTC): optimizing off-resonance and on-resonance frequency MTC methods at 0.5 and 1.5 T]. AB - AIM: To compare technical parameters and clinically relevant aspects of on- and off-resonance MTC sequences in mid- and high-field MR systems. MATERIAL AND METHODS: Both on- and off-resonance techniques were combined with an FFE sequence using 0.5 and 1.5 Tesla superconducting systems. Parameters were systematically measured by scanning a cadaveric knee joint. Signal-to-noise ratios and MT ratios for fat, cartilage and reference solution (copper sulphate) were determined. Minimal TR and the energy absorption rate were also compared. RESULTS: The MT effect at 1.5 T was more pronounced. However, using optimized parameters, clinically adequate MT contrast was achieved with both techniques and both MT units. The most important parameters for the off-resonance method are pulse angle and off-centre frequency; for the on-resonance method, pulse angle and number of composite pulse elements. Energy absorption was below 2 W/kg. Minimal TR was prolonged by up to 400%. CONCLUSION: In order to produce MTC images, optimized parameters should be applied. Using optimized pulse parameters, adequate MTC imaging is achievable with mid- and high-field systems using on- and off resonance techniques. To ensure comparability of MTC studies, the pulse parameters need to be given, and, ideally, standardized. PMID- 11258136 TI - [Is CNS activity modified by pulsed electromagnetic fields?]. AB - In the present pilot study we examined the effects of electromagnetic fields on the biological organism. The study was prompted by discussions on the possible effects on the nervous system and cognitive processes of fields produced by mobile phones. The experiments were performed in an electrophysiological laboratory. Eleven volunteers were exposed to pulsed electromagnetic fields (GSM standard). The psychophysiological method of assessing the order threshold (Ordnungsschwelle = OS) was used to examine cognitive performance. Under the test conditions, nine of the subjects showed a loss of mental regeneration, as reflected by an increase in the OS, in comparison with the field-free situation. The results of our experiments suggest an influence of pulsed fields on the cognitive regeneration process. PMID- 11258137 TI - Passive autofocusing for digital endoscopic imaging systems. AB - An autofocus is a desirable feature of an endoscope, because it relieves the user from performing a task which can be automated and thereby prevents unnecessary interruptions in the work to be performed. Autofocusing is in general best achieved by an active system, i.e., on the basis of a distance measurement. Yet, in handheld medical endoscopes such a method is unsuited because of the added weight associated with the necessary electromechanical components. Autofocusing should rather be performed passively, furthermore, in applications which are particularly critical with respect to safety, e.g., in the eye, a stable and reliable operation in real time and without interruption is necessary. Passive autofocus strategies applied to date and known to the authors lead however to algorithms which are either too slow for a real time implementation and/or are influenced by the structure of the object which is to be brought into focus. Accordingly, a new autofocus procedure has been developed which exhibits a stable and reliable operation in real time under all circumstances of interest. It is based on the squared differences of the intensity of adjacent points in both dimensions of a plane image (Square Plane Sum Modulus Difference, SPSMD) and as such particularly suitable for digital camera systems and real-time needs (typically, 30 evaluations per second on an image of 1024 x 1024 pixels). The SPSMD criterion is more sensitive, has a larger SNR than other focus criteria known to the authors and exhibits in particular no secondary extrema which could adversely affect proper focusing. As it includes intensity differences in both (perpendicular) directions in the image plane, it is essentially independent of image structures. PMID- 11258138 TI - Source localization of induced cortical oscillations during tactile finger stimulation. AB - We investigated the EEG beta event-related synchronization (ERS) after tactile finger stimulation in three subjects. Prior studies from our group using electrical stimulation and self-paced movement showed a beta rebound within one second after stimulation respectively movement offset. As the tactile-stimulation data showed a similar ERS behaviour, we extracted the cortical sources for this beta rebound by the linear estimation method in order to see whether the representation areas of different fingers were distinguishable (as is possible with MEG data). Although realistic head models of two subjects were used for the calculations the fingers could not be spatially distinguished. However, regarding the whole spatio-temporal pattern of the ERS for different fingers clear differences can be observed. PMID- 11258139 TI - [Influence of cartilage cap modelling on FEM simulation of femoral head stress]. AB - The present study reports on the finite element analysis (FEA) of the femoral head in a process of preparation for a program for the realistic simulation of correctional osteotomies of the proximal femur. While the material properties have been studied extensively, only few publications consider the influence of the cartilage layer geometry on FE stimulation of the hip joint. Various models of the femoral head with and without the cartilage layer are generated and analysed. On looking at the maximum surface stresses, we found a strong influence of the cartilage layer and the subchondral osseous layer on the magnitude of the von Mises equivalent stress. The model with an anatomically realistic cartilage layer and compact bone shows stresses of between 4 and 5.5 MPa, depending on the position of the joint, while the model with a concentric cartilage layer has a maximum von Mises stress of 0.8 MPa. Only on simulation of a "realistic" cartilage layer, with a maximum thickness at the "pole" and minimum thickness at the "equator" do the changes in stress distribution--determined by changes in the position of the femoral head--become visible. Owing to major artefacts and the inability to create a realistic cartilage layer, voxel-based models of the femur are not suitable for the simulation of the femoral head surface. PMID- 11258140 TI - Prior-authorization requirements for patient referrals to specialists. PMID- 11258141 TI - Assessing safety of new technology: conference looks at evaluation process. PMID- 11258142 TI - Physician perceptions of a national formulary. AB - OBJECTIVE: To assess the perceptions of US Department of Veterans Affairs (VA) physicians regarding effects of a National Formulary (NF) on patient care, access to drugs, physician workload, and resident training approximately 1 year after it was implemented. STUDY DESIGN: Cross-sectional survey. METHODS: A questionnaire was sent to attending physicians working within the VA healthcare system. Participants included general internists (n = 2824), neurologists (n = 238), psychiatrists (n = 997), general surgeons (n = 429), and urologists (n = 152). The response rate was 45%. RESULTS: Most physicians (63%) thought that they could prescribe needed drugs; 65% agreed that patients could obtain needed nonformulary drugs. One third disagreed that access to prescription pharmaceuticals had increased; 29% stated the NF impinged on providing quality care to their own patients, and 21% thought it did so to patients from other VA facilities. Thirty eight percent of physicians perceived the NF to be more restrictive than private sector formularies; 16% thought that the NF diminished the ability to train residents for managed care. Forty percent thought that the NF added to workload. Generalists more often perceived that the NF improved their ability to provide care compared with neurologists (27% vs 18%, P = .046), psychiatrists (27% vs 22%, P = .027), and internal medicine subspecialists (27% vs 18%, P = .001). Physicians with more clinic time were more likely to perceive that the NF increased workload. CONCLUSION: Although differences of opinions among physicians were noted, most responding VA physicians did not perceive that the NF adversely affected patient care, access to pharmaceuticals, physician workload, or resident training. PMID- 11258143 TI - Specialty of principal care physician and Medicare expenditures in patients with coronary artery disease: impact of comorbidity and severity. AB - OBJECTIVE: To explore differences in expenditures for elderly patients with acute and chronic coronary artery disease according to the specialty of the principal care physician. STUDY DESIGN: Retrospective analysis of Medicare claims. PATIENTS AND METHODS: A total of 250,514 patients with coronary artery disease (International Classification of Diseases, Ninth Revision, Clinical Modification [ICD-9-CM] codes 410-414) were drawn from a national random sample of 1992 Medicare expenditures. Patients were classified by the physician type with the highest number of Medicare Part B outpatient claims into a cardiologist group and a generalist group. The outcome was mean total expenditures, stratifying (1) by comorbidity as measured by the modified Charlson Index and (2) by severity defined as the proportion of patients with acute myocardial infarction or unstable angina. RESULTS: Those patients in the cardiologist group had lower comorbidity and higher severity than those in the generalist group. Overall mean expenditures were significantly higher for the cardiologist group than for the generalist group ($7658 vs $6047; P < .001). These differences in mean expenditures were evident at each level of comorbidity. However, when stratified by severity of diagnosis, differences were seen predominantly in those with acute diagnoses. For those with either acute myocardial infarction or unstable angina, the mean expenditures were higher for the cardiologist group than for the combined generalist group ($15,378 vs $12,260; P < .001); however, the mean expenditures for those with only chronic conditions were similar ($4856 vs $4745; P = .53). CONCLUSION: Expenditures were higher when cardiologists were the principal care physicians treating patients with acute disease but not chronic disease. PMID- 11258144 TI - Clinical and economic outcomes of an ambulatory urinary tract infection disease management program. AB - OBJECTIVE: To evaluate the effectiveness of a urinary tract infection disease management program. STUDY DESIGN: A pre-post design was used. One year of data before and after promoting the treatment guideline were compared. PARTICIPANTS AND METHODS: A 300,000-member managed care organization introduced an antibiotic treatment guideline designed to change the antibiotic prescribing practices of community physicians. The study intervention was the promotion of a treatment guideline through mailings and face-to-face interventions by 2 disease management specialists. A relational database was created to measure changes in healthcare resources, use of antibiotics, and health event profiles. RESULTS: The study demonstrated that prescribing patterns could be modified through treatment guideline distribution and face-to-face discussions. The study also found similar success rates across a range of antibiotics. Average health event costs decreased by 36% for kidney infections (P = .696) and by 7% for bladder infections (P < .05) after the treatment guideline was implemented; however, when controlling for patient age, sex, and comorbidities, the econometric model did not find a reduction in health event costs for either kidney or bladder infections. Fluoroquinolones were a cost driver compared with other antibiotics used to treat kidney and bladder infections. CONCLUSIONS: Consideration should be given to expanding the number of well-established antibiotics on the treatment guideline. Also, fluoroquinolones should be reserved for patients with sulfa allergies or failures with initial antibiotic treatment. In addition, it is recommended that future costs and outcomes be assessed after changes are made to the treatment guideline. PMID- 11258145 TI - Renal effects of angiotensin-converting enzyme inhibitors that result in cost savings and improved patient outcomes. AB - BACKGROUND: In some patients with renal disease, use of angiotensin-converting enzyme (ACE) inhibitors is thought to improve renal function, whereas in others their use leads to worsening. Many questions remain about the categories of patients that benefit from ACE inhibitor use. OBJECTIVE: To clarify the use of ACE inhibitors in patients with renal disease. STUDY DESIGN: A literature review focusing on various renal diseases, ACE inhibitors, and criteria of cost effectiveness was performed. RESULTS: Almost 100 clinical studies were reviewed. Treatment with ACE inhibitors seems to have beneficial effects in type 1 and type 2 diabetes mellitus with nephropathy, AIDS nephropathy, and other nondiabetic renal diseases. Use of these agents in these diseases decreases the progression of renal disease and the need for dialysis, resulting in potential cost savings and improved quality of life. Data supporting goal blood pressures indicate the need to aggressively decrease this risk factor. Use of ACE inhibitors is hazardous in bilateral renal artery stenosis, particularly with volume depletion, but may be valuable in patients with unilateral stenosis. In African Americans, ACE inhibitor treatment is likely to be of benefit, although required doses may be higher than for whites, and caution must be exercised in certain situations. The potential efficacy of angiotensin receptor blockers and other new drugs that affect the renin-angiotensin system is assessed. CONCLUSIONS: Use of ACE inhibitors has benefit in renal disease states characterized by increased glomerular perfusion pressure, their use in other renal disease states, particularly those characterized by reduced glomerular perfusion pressure, may be risky. The benefits conferred by ACE inhibitor therapy are so dramatic in terms of cost savings and improved quality of life that their use in certain clinical situations should be strongly encouraged in managed care and other practice settings. PMID- 11258146 TI - Are patients' chief complaints generally specific to one organ system? AB - BACKGROUND: The coordinator of care function is one of the most important roles played by primary care physicians. This role is essential for efficient delivery of healthcare to patients with unfocused medical problems. OBJECTIVES: To identify which chief complaints are unfocused and to determine how often visits to office-based physicians are for unfocused chief complaints. STUDY DESIGN: Retrospective review of National Ambulatory Medical Care Survey data. METHODS: We defined an unfocused chief complaint as one for which fewer than 95% of the office visits for the top 10 diagnoses associated with that chief complaint were related to a single organ system or specialty area. We analyzed data from the 1990-1994 National Ambulatory Medical Care Survey to determine the frequency of new patient visits to physicians for different chief complaints and to determine the frequency with which common chief complaints yield diagnoses in a single organ system. RESULTS: The 3 most common chief complaints in each of 12 symptom categories accounted for 80 million (32%) of the 250 million new patient office visits made during the survey period. Unfocused conditions accounted for 26% of visits for these chief complaints. The unfocused chief complaints included musculoskeletal conditions (back pain, knee pain, low back pain), mental/nervous system conditions (anxiety/nervousness, smoking problems, headaches, vertigo/dizziness), abnormal pulsations, swollen glands, and abdominal pain. CONCLUSIONS: Patients' chief complaints and the resulting diagnoses are often within the same organ system. We found that a coordinator of care role for primary care physicians is appropriate for common neurologic, rheumatologic, and general complaints. A coordinator of care is not needed for specific specialty areas, including ophthalmology, dermatology, obstetrics/gynecology, urology, and otolaryngology, because patients typically can accurately self-refer to these specialists. Our study did not address reasons to use primary care physicians as coordinators of care, such as preventive care, patient preference, or cost effectiveness of care. PMID- 11258147 TI - Vascular and metabolic abnormalities in patients with angina pectoris and normal coronary angiograms. PMID- 11258148 TI - Glucagon-like peptide-1. Gastrointestinal function and possible mechanism of action. PMID- 11258149 TI - Major reproductive health characteristics in male Gulf War Veterans. The Danish Gulf War Study. AB - INTRODUCTION: The male reproductive system could have been affected by various hazardous agents and exposures during and in the aftermath of the Persian Gulf War scenario. We tested the hypothesis that, compared to controls, male Danish Gulf War Veterans would have adverse sex hormone levels, decreased fertility, and a larger proportion of adverse pregnancy outcomes including spontaneous abortions, congenital diseases and malformations. MATERIAL AND METHODS: A cross sectional study was performed during the period January 1997 to January 1998 which included 661 male subjects who had been deployed in the Persian Gulf within the period August 2 1990 until December 31 1997. A control group of 215 Danish military men, not deployed in the Gulf region, was selected with random matching by age and type of work. All participants underwent clinical and paraclinical examinations, and had an interview based on a previously completed comprehensive questionnaire. A venous blood sample was drawn to determine serum concentrations of the follicle-stimulating hormone (FSH), the luteinizing hormone (LH), testosterone, serum hormone binding globulin (SHBG), and inhibin B. The free androgen index was calculated from testosterone and SHBG levels. RESULTS: No differences were found between Gulf War Veterans and controls with respect to any of the reproductive hormones measured, nor with respect to fertility or the prevalence of spontaneous abortions, congenital diseases or malformations among the offspring. Also cohabitational characteristics were similar. CONCLUSION: Based on the results of this study we conclude that the biological reproductive health of male Danish Gulf War Veterans seemed to be unaffected by their engagement in the post war peace-keeping mission. PMID- 11258151 TI - Survey of cash balance conversions. AB - By examining data on actual conversions to cash balance pension plans, the author challenges the validity of major criticisms faced by employers sponsoring cash balance plans and their advisors. The data admonish against using broad generalizations to assess the total impact on employers and employees of a conversion to a cash balance plan. PMID- 11258150 TI - Use and validation of public data files for identification of the diabetic population in a Danish county. AB - INTRODUCTION: This study aims to describe the process of identifying people known to have diabetes through public data files, to validate this method, and to describe models for optimization of such identification processes. PATIENTS AND METHODS: In a study population of 303,250 citizens, the diabetics were identified by combining information from public data files with information from general practitioners. Data validity was checked by comparing the results of data searches in public data files against information from general practitioners and a random sample of diabetics. Two models were defined to optimize the use of public data files for identification of diabetics. In model A the minimum number of parameters needed to obtain a sensitivity as high as possible was identified. In model B the optimal combination of parameters needed to obtain a high positive predictive value combined with a high sensitivity was identified. RESULTS: A total of 5449 diabetics were identified. Of those 4438 (81%) were classified as Type 2 diabetics and 1011 (19%) were classified as Type 1 diabetics. The data validation revealed that one person was misclassified as a diabetic and 93 persons were misclassified as non-diabetics. In model A the identification parameters included: "prescription", "HbA1c", "chiropodist service" and "glucose service". In model B the optimal combination of parameters was identified as: minimum two HbA1c measurements, minimum one visit to a chiropodist, minimum one prescription or minimum one abnormal HbA1c during one year. CONCLUSION: Public data files are suitable for identification of both Type 1 and Type 2 diabetics. Models have been developed to identify diabetics and to promote the possibilities of long-term follow-up and quality assessment in an unselected diabetic population in a region. PMID- 11258152 TI - Legal issues in cash balance pension plan conversions. AB - Replacing a traditional pension with a cash balance plan raises a number of complicated and unsettled legal issues, including the protection of accrued benefits, the rate of benefit accrual, age discrimination and notice requirements. This article discusses those issues and concludes that routine conversions to cash balance plans appear to be legal both currently and into the foreseeable future. PMID- 11258153 TI - Cash balance plans: helping employers meet today's staffing needs. AB - The author examines how cash balance plans better meet employers' staffing needs than traditional pension plans. He asserts that out-of-date pension laws, rather than employers, are responsible for creating the very "abuses" that so many are complaining about with regard to cash balance plan conversions. PMID- 11258154 TI - Cash balance pension plans--accounting and business implications. AB - This article illustrates the accounting and disclosure implications of converting from traditional pensions to cash balance plans. That information is followed by a description of issues that have encouraged employers to initiate those conversions. PMID- 11258155 TI - Rate-of-return guarantees for defined contribution plans. AB - This article explores the conceptual basis for an employer-backed minimum rate-of return guarantee as an option under defined contribution plans. Expanding an earlier analysis, the author presents a prototype model of how the guarantee would work and discusses how it might be used for a mandatory Social Security defined contribution plan. PMID- 11258156 TI - The costs and benefits of the Form 5500 annual report. AB - The author suggests that although the Form 5500 annual report may help to reduce legal violations, it may do so in an unnecessarily expensive way that serves the economic interests of some professional groups. In particular, his investigation suggests that the CPA audit provides questionable benefits and amounts to a special interest group's extraction of rents through the regulatory process. PMID- 11258157 TI - Administration--delayed benefits payment; interest. Fotta v. Trustees of the United Mine Workers of America, Health and Retirement Fund of 1974, 165 F.3d.209 (3rd Cir. 1998). AB - In a case of first impression, the Third Circuit holds that a beneficiary of an ERISA plan may sue the plan for interest on delayed benefits payments under Section 502(a)(3)(B) regardless of whether he also seeks to recover unpaid benefits. A beneficiary is not made whole when a plan pays benefits after delay since he has lost the time value of the money. Interest for delayed benefits payments is appropriate equitable relief that prevents the plan from being unjustly enriched at the beneficiary's expense. Since interest is money damages that is restitutive in nature, it is a form of equitable relief. Interest is not prohibited as "extracontractual" since it is compensation that is part of the contractual obligation. PMID- 11258158 TI - Adopting hybrid pension plans: financial and communication issues. AB - This article provides a systematic framework for the evaluation of the movement toward hybrid pension plans by examining the reasons given by firms for converting their existing pension plans to hybrid plans, illustrating the impact of plan changes on expected pension benefits, and identifying winners and losers. PMID- 11258159 TI - Public plans--Americans with Disabilities Act--discrimination-disparate treatment of mental disabilities. Rogers v. Department of Health and Environmental Control, 174 F.3d 431 (4th Cir.1999). AB - Title II of the Americans with Disabilities Act of 1990 (ADA), which prohibits discrimination in the services, programs or activities of public entities, does not require a state's long-term disability plan to provide the same level of benefits for mental and physical disabilities. By placing mental disabilities in different risk classifications from physical disabilities, South Carolina's long term disability plan did not violate a South Carolina statute that prohibits discrimination "between insureds of the same class and risk involving the same hazards." PMID- 11258160 TI - Age discrimination--disability benefits; public sector. Arnett v. California Public Employees Retirement System, No. 98-15574, __F.3d __(9th Cir.1999). AB - Under the statutory formula for determining disability retirement benefits for California public safety employees, the amount of benefits differs solely on the basis of the retiree's age when hired. This formula may violate the Age Discrimination in Employment Act (ADEA) under a disparate treatment theory if the employees can prove discriminatory motive. While a formula for determining benefits that is based upon actual years of service does not violate ADEA, a formula based upon potential years of service as determined with reference to a presumed retirement age may violate the act. The disparate impact theory may be used to prove violation of ADEA in the Ninth Circuit. PMID- 11258161 TI - Out-patient parenteral antibiotic therapy (OPAT): clinical outcomes and adverse events. AB - A chart review was conducted of patients in a program featuring self directed, home infusion of antibiotics for serious infections utilizing an out-patient medical office for teaching, mixing of drugs, and monitoring of patients. 302 courses of out-patient parenteral antibiotic therapy (OPAT) were administered to 221 patients. Therapy was successful in 94% of the episodes. Objective adverse events were noted in 25% of patients. To maximize the chance for a successful outcome, treatment plans should be individualized and structured to include systematic monitoring for adverse effects. PMID- 11258162 TI - Substance abuse and dependence in a public hospital: Hawaii. AB - A pilot study of the prevalence of substance abuse disorders was completed on the only open unit of the only state psychiatric hospital in Hawaii to address the following questions: 1) What were the substances of abuse and dependence in this population? 2) What was the pattern of abuse and/or dependence in this sample? 3) How did these patterns compare to the patterns observed in the published literature? 4) What was the predominant Stage of Change of these patients? 5) Was the staff perception that these patients were in denial an accurate perception? METHODOLOGY: All patients admitted to the unit between 1st June and 31st August 1999 comprised the sample, N = 35. Each patient was assigned diagnoses based upon the DSM-IV criteria and level of change was assessed. RESULTS: The demographic characteristics of the sample follow: 60% were ages 20-39, 89% were male; 92% were currently single, 71% had an education of high school or college, and 66% had a diagnosis of schizophrenia or schizoaffective disorder. The racial composition reflected the diversity of Hawaii. Although 20% of the sample had no substance abuse problem, 66% of the remaining patients were multiply dependent upon alcohol, cannabis, crystal methamphetamine, or cocaine with 48% of these patients in the Precontemplative Stage of Change (denial). The patterns of multiple substances of abuse and dependence were higher than in the reported literature. Further studies are needed. PMID- 11258163 TI - Barriers to good end-of-life care: a physician survey. AB - Surveyed about barriers to good end-of-life care were 804 Hawaii physicians in specialties most likely to care for dying patients. Responses were received by 367 (46%). The majority attended terminally ill patients within the past year and felt that the physician should be the first to tell a patient that he/she is dying. Yet 86% identified barriers to talking about end-of-life preferences and 94% identified barriers to providing good end-of-life care. Perceived as major barriers were family conflict about the best course of action, patient/family discomfort with or fear of death, and cultural/religious beliefs of the patient or family. Since relatively few respondents supported the concepts of physician assisted suicide (32%) or physician-assisted death (18%), the alternative is for physicians to join with other concerned entities to help overcome the attitudinal, behavioral, educational, and economic barriers to providing appropriate, humane, and compassionate care for the dying. PMID- 11258164 TI - New pain standards offer key role for pharmacy. PMID- 11258165 TI - HCFA, states work at increasing prescription drug access in Medicare, Medicaid. PMID- 11258166 TI - Thrombin inhibitor approved for anticoagulation during angioplasty. PMID- 11258167 TI - Recap of FDA product approvals--2000. PMID- 11258168 TI - JCAHO adds patient safety standards for hospitals. PMID- 11258169 TI - Wisconsin coalition recommends steps for reducing medication errors. PMID- 11258170 TI - Improved glycemic control results in short-term cost savings, researchers find. PMID- 11258172 TI - Editing in an age of troubled writing. PMID- 11258171 TI - Infliximab licensing expands to include halting arthritis progression. PMID- 11258173 TI - Moxifloxacin: clinical efficacy and safety. AB - The activity, pharmacokinetics, pharmacodynamics, efficacy, safety, drug interactions, and dosage and administration of moxifloxacin are reviewed. Moxifloxacin is an oral 8-methoxyquinolone antimicrobial approved in December 1999 for use in the treatment of acute bacterial sinusitis, acute bacterial exacerbations of chronic bronchitis, and community-acquired pneumonia. This fluoroquinolone is active against common community-acquired respiratory pathogens (Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis), atypical pathogens, and many anaerobes. Moxifloxacin has an absolute bioavailability of 90% after oral administration and a mean elimination half-life of 12 hours. The drug is not a substrate or inhibitor of the hepatic cytochrome P 450 isoenzyme system thereby avoiding many potential drug interactions. Moxifloxacin has limited phototoxic potential. In clinical trials, moxifloxacin had clinical success rates of 88-97% and bacteriologic eradication rates of 90 97%. Reported adverse effects were primarily gastrointestinal (nausea, diarrhea) and were mild to moderate in severity. Moxifloxacin prolongs the QT interval by a mean + S.D. of 6 +/- 26 milliseconds above baseline and should be used with caution in patients with proarrhythmic conditions and avoided in patients receiving antiarrhythmia agents, such as quinidine, procainamide, amiodarone, and sotalol. The standard oral dosage is 400 mg once a day. Dosage adjustment is unnecessary in patients with renal dysfunction or mild to moderate hepatic dysfunction. Moxifloxacin is a safe and effective antimicrobial that will be useful for treating acute sinusitis, acute bacterial exacerbations of chronic bronchitis, and community-acquired pneumonia. PMID- 11258174 TI - Community pharmacist knowledge and behavior in collecting drug copayments from Medicaid recipients. AB - Community pharmacists' knowledge and behavior regarding the collection of copayments for prescription drugs from Medicaid recipients were studied. In fall 1998 a questionnaire was mailed to a random sample of 1465 community pharmacists (one pharmacist per drugstore) in Maryland, Pennsylvania, and West Virginia. The objectives were to determine the extent to which these pharmacists waived copayments for prescription drugs for Medicaid recipients, to document the pharmacists' knowledge of federal policies on Medicaid copayments, and to evaluate the factors associated with pharmacist copayment collection and knowledge of federal copayment policies. A total of 543 pharmacists (37%) responded. Most respondents indicated that they collected copayments for over 90% of drugs dispensed to Medicaid patients subject to copayment policies. Pharmacists most likely to waive Medicaid copayments practiced in drugstores with a high volume of Medicaid-related prescriptions and a large percentage of customers who were elderly Medicaid recipients. Pharmacists least likely to waive copayments believed that doing so would have a negative financial impact on the pharmacy. Nearly three fourths of the pharmacists exhibited fair or good knowledge of federal Medicaid copayment policies, but this varied widely by state. Many said that they would collect copayments in at least some situations even if this opposed federal policy. Pharmacists in Maryland, Pennsylvania and West Virginia had highly variable behavior patterns and knowledge with respect to the collection of drug copayments from Medicaid recipients. PMID- 11258175 TI - Gluten in pharmaceutical products. PMID- 11258176 TI - Verapamil intoxication after substitution of immediate-release for extended release verapamil. PMID- 11258177 TI - Strategic management of pharmaceutical expenses. PMID- 11258179 TI - Achieving blood lipid goals in patients with coronary artery disease. PMID- 11258180 TI - The formulary of a large health maintenance organization in Israel. PMID- 11258181 TI - Defining home care. PMID- 11258183 TI - A Jehovah's Witness with oversized hands. PMID- 11258184 TI - Understanding abdominal pain. PMID- 11258185 TI - Malabsorption in children. PMID- 11258186 TI - Casebook: dyspepsia. PMID- 11258187 TI - Diagnosing gastroenteritis and travellers' diarrhoea. PMID- 11258188 TI - Management of abnormal liver biochemistry. PMID- 11258189 TI - Gastroenterological diseases and the mouth. PMID- 11258190 TI - A GP guide to inflammatory bowel disease. PMID- 11258191 TI - Nutrition and dietary advice in diabetes. PMID- 11258193 TI - The relative power of SNPs and haplotype as genetic markers for association tests. AB - Identifying the polymorphisms that contribute to disease predisposition and drug response is a major goal of the post-genome era. Single nucleotide polymorphisms (SNPs) in disease-related genes are often used as candidates in the search for causative variations. Association tests based on haplotypes have also been suggested and, at times, have provided greater statistical power than tests based on the underlying SNPs. Here we review the statistical model traditionally used to describe association studies for complex traits and derive novel results for the relative power of SNP-based and haplotype-based tests of association. In the model, a set of independent SNP-based variations, some of which contribute to a measured phenotype, may be used as markers directly or may be organised into haplotype markers. Provided that the marker set includes all the causative SNPs, we find a simple rule for the relative power of SNP and haplotype markers: SNP based tests have greater power when the number of causative SNPs (a subset of the total set of SNPs) is smaller than the total number of haplotypes. Furthermore, we find that regression tests for the simple main effect of each haplotype are generally more powerful than ANOVA tests applied to haplotype pairs. A review of recent literature supports our findings. PMID- 11258194 TI - Cluster analysis and promoter modelling as bioinformatics tools for the identification of target genes from expression array data. AB - Expression arrays yield enormous amounts of data linking genes, via their cDNA sequences, to gene expression patterns. This now allows the characterisation of gene expression in normal and diseased tissues, as well as the response of tissues to the application of therapeutic reagents. Expression array data can be analysed with respect to the underlying protein sequences, which facilitates the precise determination of when and where certain groups of genes are expressed. More recent developments of clustering algorithms take additional parameters of the experimental set-up into account, focusing more directly on co-regulated set of genes. However, the information concerning transcriptional regulatory networks responsible for the observed expression patterns is not contained within the cDNA sequences used to generate the arrays. Regulation of expression is determined to a large extent by the promoter sequences of the individual genes (and/or enhancers). The complete sequence of the human genome now provides the molecular basis for the identification of many regulatory regions. Promoter sequences for specific cDNAs can be obtained reliably from genomic sequences by exon mapping. In the many cases in which cDNAs are 5'-incomplete, high quality promoter prediction tools can be used to locate promoters directly in the genomic sequence. Once sufficient numbers of promoter sequences have been obtained, a comparative promoter analysis of the co-regulated genes and groups of genes can be applied in order to generate models describing the higher order levels of transcription factor binding site organisation within these promoter regions. Such modules represent the molecular mechanisms through which regulatory networks influence gene expression, and candidates can be determined solely by bioinformatics. This approach also provides a powerful alternative for elucidating the functional features of genes with no detectable sequence similarity, by linking them to other genes on the basis of their common promoter structures. PMID- 11258195 TI - Genomics and large scale phenotypic databases. AB - Progress in the development of molecular genetic tools and in sequencing the human genome will accelerate the understanding of complex genetic diseases. However, phenotypic clinical data needs to be obtained and recorded to a similar degree of precision in order to match the wealth of molecular genetic data. To achieve this goal, large scale phenotypic databases of complex genetic diseases are under construction. LURIC (the LUwigshafen Risk and Cardiovascular Health Study) is such a project, aiming to identify new genetic and environmental risk factors or markers for cardiovascular disease in order to better understand the pathophysiology of complex genetic disease. It should also allow the determination of the prognostic role of new markers by studying functional genomics, (the association between a gene variant and phenotype), and pharmacogenomics (the influence of genetic variation on the response to therapeutic agents). PMID- 11258196 TI - The human genome project: a historical perspective. AB - Efforts in genomics over the last decade have created a stream of opportunities for drug discovery. High-throughput DNA sequencing has forced a re-definition of the paradigm for identification and validation of targets for drug development. One purpose of this review is to delineate the different approaches to sequence data generation and to establish their various uses for the definition of gene function. There still remain crucial dilemmas for the pharmaceutical industry. The multitude of potential targets can each absorb enormous validation costs and the vast majority are likely to prove academically interesting but useless for drug development. An additional dimension arises from the importance of sequence variation between different individuals. These differences can determine response to therapy and must inform both the drug development process and healthcare delivery. This presents great challenges and opportunities for drug companies, their customers and society as a whole. I will review the technological aspects in some detail and give my view of the legal and social aspects. The field of bioinformatics is at the core of functional and pharmacogenomics and advances will depend on the continuing evolution of tools to interpret data. For the most part this evolution is reviewed in the context of specific application areas rather than as a discrete field, in recognition of its all-pervasive effects. PMID- 11258197 TI - ABC drug transporters: hereditary polymorphisms and pharmacological impact in MDR1, MRP1 and MRP2. AB - Transport by ATP-dependent efflux pumps, such as P-glycoprotein (PGP) and multi drug resistance related proteins (MRPs), influences bioavailability and disposition of drugs. These efflux pumps serve as defence mechanisms and determine bioavailability and CNS concentrations of many drugs. However, despite the fact that substantial data have been accumulated on the structure, function and pharmacological role of ABC transporters and even though modification of PGP function is an important mechanism of drug interactions and adverse effects in humans, there is a striking lack of data on variability of the underlying genes. This review focuses on the human drug transporter proteins PGP (MDR1) and the multi-drug resistance proteins MRP1 and MRP2. An overview is provided of pharmacologically relevant genetic, structural and functional data as well as on hereditary polymorphisms, their phenotypical consequences and pharmacological implications. PMID- 11258199 TI - Notes from the SNP vs. haplotype front. AB - Single nucleotide polymorphisms (SNPs) and haplotypes are commonly used genetic markers in clinical studies. We provide some broad guidelines for deciding which of the two is most appropriate in particular circumstances. Molecular haplotyping techniques are also briefly reviewed and contrasted with electronic approaches. PMID- 11258198 TI - Pharmacogenetics of human androgens and prostatic diseases. AB - Prostate cancer (PCa) and benign prostatic hypertrophy (BPH) are two common and growing public health problems in the Western world. We review here the recent biochemical and pharmacogenetic literature related to these two prostatic disorders. We focus first on constitutional ('germline') single nucleotide polymorphism (SNPs) at the steroid 5 alpha-reductase (SRD5A2) locus, which encodes the human prostatic (or Type II) steroid 5 alpha-reductase enzyme. The investigations reviewed point to several uses of personalised medicine at the SRD5A2 locus. In addition, we report on recent identification of somatic pharmacogenetic alterations at the androgen receptor (AR) locus, which encodes the human androgen receptor, suggesting that this also may be a fruitful field of investigation, with important clinical applications. Pharmacogenomic investigation of constitutional and somatic DNA changes in human genes predisposing to cancer may lead to significant advances in chemoprevention, presymptomatic diagnosis and improved treatment of PCa. PMID- 11258200 TI - Cambridge Healthtech Institute's Third Annual Conference on Lab-on-a-Chip and Microarrays. 22-24 January 2001, Zurich, Switzerland. AB - Cambridge Healthtech Institute's Third Annual Conference on Lab-on-a-Chip and Microarray technology covered the latest advances in this technology and applications in life sciences. Highlights of the meetings are reported briefly with emphasis on applications in genomics, drug discovery and molecular diagnostics. There was an emphasis on microfluidics because of the wide applications in laboratory and drug discovery. The lab-on-a-chip provides the facilities of a complete laboratory in a hand-held miniature device. Several microarray systems have been used for hybridisation and detection techniques. Oligonucleotide scanning arrays provide a versatile tool for the analysis of nucleic acid interactions and provide a platform for improving the array-based methods for investigation of antisense therapeutics. A method for analysing combinatorial DNA arrays using oligonucleotide-modified gold nanoparticle probes and a conventional scanner has considerable potential in molecular diagnostics. Various applications of microarray technology for high-throughput screening in drug discovery and single nucleotide polymorphisms (SNP) analysis were discussed. Protein chips have important applications in proteomics. With the considerable amount of data generated by the different technologies using microarrays, it is obvious that the reading of the information and its interpretation and management through the use of bioinformatics is essential. Various techniques for data analysis were presented. Biochip and microarray technology has an essential role to play in the evolving trends in healthcare, which integrate diagnosis with prevention/treatment and emphasise personalised medicines. PMID- 11258201 TI - Cambridge Healthtech Institute's Third Annual Conference on human genetic variation. 16-18 October 2000, Philadelphia, Pennsylvania, USA. AB - A major goal of pharmacogenomics is to identify the human genetic variation that influences susceptibility to complex diseases. Recently, theoretical statistical analyses have suggested that genes for complex diseases may be found by linkage disequilibrium (i.e., association). Single nucleotide polymorphism (SNP) susceptibility alleles for common diseases can occur at high frequencies in various populations and, thus, have a major impact on morbidity and mortality. To be successful, SNP mapping studies require successful teamwork, integrating clinicians, epidemiologists, molecular genetics experts, laboratory automation engineers, bioinformatics and database experts. New statistical methods are also developing rapidly and promise to further increase the power of these studies. A recent conference on human genetic variation provided an opportunity for experts in all of these disciplines to exchange ideas. At present, great technological challenges need to be overcome in order to increase the throughput greatly while lowering cost and still maintaining high accuracy for SNP genotyping. Although this approach is relatively new (at least on the scale now being contemplated), the large payoffs anticipated to accrue from the successful mapping of SNPs in disease genes has led the area to be very strongly supported by both public and private funding sources. The potential payoff for improving disease diagnosis and therapeutic efficacy, with better avoidance of adverse events based on SNP associations, is providing a tremendous incentive to move this effort forward at an ever-accelerating pace. PMID- 11258202 TI - Pharmacogenomics WebWatch. PMID- 11258204 TI - Some natural skepticism about the Natural Health Products Directorate. PMID- 11258203 TI - Rationale and design of the LURIC study--a resource for functional genomics, pharmacogenomics and long-term prognosis of cardiovascular disease. AB - BACKGROUND AND AIM: Coronary artery disease (CAD), arterial hypertension and Type 2 diabetes mellitus are common polygenetic disorders which have a major impact on public health. Disease prevalence and progression to cardiovascular complications, such as myocardial infarction (MI), stroke or heart failure, are the product of environment and gene interaction. The LUdwigshafen RIsk and Cardiovascular Health (LURIC) study aims to provide a well-defined resource for the study of environmental and genetic risk factors, and their interactions, and the study of functional relationships between gene variation and biochemical phenotype (functional genomics) or response to medication (pharmacogenomics). Long-term follow-up on clinical events will allow us to study the prognostic importance of common genetic variants (polymorphisms) and plasma biomarkers. SETTING: Cardiology unit in tertiary care medical centre in south-west Germany. STUDY DESIGN: Prospective cohort study of individuals with and without cardiovascular disease at baseline. PATIENTS AND METHODS: LURIC is an ongoing prospective study of currently > 3300 individuals in whom the cardiovascular and metabolic phenotypes CAD, MI, dyslipidaemia, hypertension, metabolic syndrome and diabetes mellitus have been defined or ruled out using standardised methodologies in all study participants. Inclusion criteria for LURIC were: German ancestry (limitation of genetic heterogeneity) clinical stability (except for acute coronary syndromes [ACSs]) availability of a coronary angiogram (this inclusion criterium was waived for family members provided that they met all other inclusion and exclusion criteria) Exclusion criteria were: any acute illness other than ACSs any chronic disease where non-cardiac disease predominated a history of malignancy within the past five years. Exclusion criteria were pre specified in order to minimise the impact of concomitant non-cardiovascular disease on intermediate biochemical phenotypes or on clinical prognosis (limitation of clinical heterogeneity). A standardised personal and family history questionnaire and an extensive laboratory work-up (including glucose tolerance testing in non-diabetics and objective assessment of smoking exposure by determination of cotinine plasma levels) was obtained from all individuals after informed consent. A total of 115 ml of fasting venous blood was sampled for the determination of a pre-specified wide range of intermediate biochemical phenotypes in serum, plasma or whole blood, for leukocyte DNA extraction and immortalisation of B-lymphocytes. Biochemical phenotypes measured included markers of endothelial dysfunction, inflammation, oxidative status, coagulation, lipid metabolism and flow cytometric surface receptor expression of lympho-, mono and thrombocytes. In addition, multiple aliquots of blood samples were stored for future analyses. RESULTS: A total of 3500 LURIC baseline measurements were performed in 3316 individuals between July 1997 and January 2000. The baseline examination was repeated within a median of 35 days in 5% of study participants (n = 166, including a third examination in 18 after a median of 69 days) for pharmacogenomic assessment of lipid-lowering therapy and for quality control purposes. A five-year follow-up on major clinical events (death, any cardiovascular event including MI, stroke and revascularisation, malignancy and any hospitalisation) is ongoing. The clinical phenotypes prevalent at baseline in the cohort of 2309 men (70%) with a mean age of 62 +/- 11 years and 1007 women (30%), mean age 65 +/- 10 years, were angiographically-documented CAD in 2567 (79%), MI in 1368 (41%), dyslipidaemia in 2050 (62%) with hypercholesterolaemia > or = 240 mg/dl (27%), hypertriglyceridaemia > or = 150 mg/dl (44%) and HDL cholesterol < or = 35 mg/dl (38%) in individuals not treated with lipid-lowering agents, systemic hypertension in 1921 (58%), metabolic syndrome in 1591 (48%), Type 2 diabetes in 1063 (32%) and obesity defined by body mass index > or = 30 kg/m2 in 770 (23%). Control patients in whom CAD had been ruled out angiographically were five years younger than those with CAD (59 +/- 12 and 64 +/ 10 years, respectively; p < 0.001), twice as often females (48% compared to 25% females in the CAD group, p < 0.001) and had significantly less cardiovascular risk factors than individuals with CAD. The prevalence of specific cardiovascular risk subsets in LURIC, such as the elderly (> or = 75 years), was 375 (11%), while 213 (6%) were young adults (< 45 years) and 904 (27%) were postmenopausal women (90% of all females). A low risk status (< or = 1 out of the four traditional risk factors: dyslipidaemia, smoking, hypertension and diabetes mellitus) was identified in 314 (9%) individuals of the entire cohort (5% in CAD and 26% in controls, p < 0.001) and 97 (3%) carried none of the four risk factors (1% in CAD and 9% in controls, p < 0.001). (ABSTRACT TRUNCATED) PMID- 11258205 TI - To treat or not to treat: managing bacteriuria in elderly people. PMID- 11258206 TI - Patient-controlled analgesia. PMID- 11258207 TI - Phenylpropanolamine, stroke and hypertension. PMID- 11258208 TI - Double trouble: impact of inappropriate use of asthma medication on the use of health care resources. AB - BACKGROUND: There is considerable controversy about the regular use of short acting beta-agonists for the treatment of asthma. Although case-control studies have suggested that excessive use of these drugs may worsen asthma control and increase the risk of fatal or near-fatal asthma, the controversy remains unresolved because of the confounding that exists among disease control, disease severity and the use of short-acting beta-agonists. Whatever the cause-and-effect relation between the use of short-acting beta-agonists and disease severity, we hypothesized that their excessive use, in conjunction with underuse of inhaled corticosteroids, would be a marker for poorly controlled asthma and excessive use of health care resources. METHODS: To characterize the pattern of health services utilization among asthmatic patients taking various doses of inhaled beta agonists and corticosteroids in British Columbia, we linked the relevant health administrative databases. All patients between 5 and 50 years of age for whom a prescription for a short-acting beta-agonist was filled in 1995 and whose prescription data were captured through the provincial drug plan were included in a retrospective analysis of prescriptions for asthma drugs, physician prescribing patterns and health services utilization. Patients' use of asthma medication was classified as appropriate (low doses of short-acting beta-agonist and high doses of inhaled corticosteroid) or inappropriate (high doses of short-acting beta agonist and low doses of inhaled corticosteroid), and the 2 resulting groups were compared, by means of logistic, Poisson and gamma regression, for differences in prescribing patterns, physician visits and use of hospital resources. RESULTS: A total of 23,986 patients were identified as having filled a prescription for a short-acting beta-agonist (for inhalation) in 1995. Of these, 3069 (12.8%) filled prescriptions for 9 or more canisters of beta-agonist, and of this group of high dose beta-agonist users, 763 (24.9%) used no more than 100 micrograms/day of inhaled beclomethasone. On average, those with inappropriate use of beta-agonists visited significantly more physicians for their prescriptions (1.8 v. 1.4), and each of these physicians on average wrote significantly more prescriptions for asthma medications per patient than the physicians who prescribed to appropriate users (5.2 v. 2.5 prescriptions). Patients with inappropriate use were more likely to be admitted to hospital (adjusted relative risk [RR] 1.68, 95% confidence interval [CI] 1.25-2.26), were admitted to hospital more frequently (adjusted RR 1.81, 95% CI 1.41-2.32) and were more likely to require emergency admission (adjusted RR 1.93, 95% CI 1.35-2.77). INTERPRETATION: Despite the widespread distribution of guidelines for asthma pharmacotherapy, inappropriate use of asthma medications persists (specifically excessive use of inhaled short acting beta-agonists combined with underuse of inhaled corticosteroids). Not only are patients who use medication inappropriately at higher risk for fatal or near fatal asthma attacks, but, as shown in this study, they use significantly more health care resources than patients with appropriate medication use. PMID- 11258210 TI - Guidelines do matter. PMID- 11258209 TI - Guidelines as rationing tools: a qualitative analysis of psychosocial patient selection criteria for cardiac procedures. AB - BACKGROUND: Cardiac procedure guidelines often include psychosocial criteria for selecting patients that potentially introduce social value judgements into clinical decisions and decisions about the rationing of care. The aim of this study was to investigate the terms and justifications for and the meanings of psychosocial patient characteristics used in cardiac procedure guidelines. METHODS: We selected English-language guidelines published since 1990 and chapters in textbooks published since 1989. These guidelines amalgamated multiple sources of evidence and expertise and made recommendations regarding patient selection for specific procedures. A multidisciplinary team of physicians and social scientists extracted passages regarding psychosocial criteria and developed categories and conceptual relationships to describe and interpret their content. RESULTS: Sixty-five papers met the criteria for inclusion in the study. Forty-five (69%) mentioned psychosocial criteria as procedure indications or contraindications. The latter fell into several categories, including behavioural and psychological issues, relationships with significant others, financial resources, social roles and environmental circumstances. INTERPRETATION: Psychosocial characteristics are portrayed as having 2 roles in patient selection: as risk factors intrinsic to the candidate or as indicators of need for special intervention. Guidelines typically simply list psychosocial contraindications without clarifying their specific nature or providing any justification for their use. Psychosocial considerations can help in the evaluation of patients for cardiac procedures, but they become ethically controversial when used to restrict access. The use of psychosocial indications and contraindications could be improved by more precise descriptions of the psychosocial problem at issue, explanations regarding why the criterion matters and justification of the characteristic using a biological rationale or research evidence. PMID- 11258211 TI - Psychosocial patient selection criteria in clinical practice guidelines: an ethical basis for rationing? PMID- 11258212 TI - Clinical practice guidelines for the care and treatment of breast cancer: adjuvant systemic therapy for node-positive breast cancer (summary of the 2001 update). The Steering Committee on Clinical Practice Guidelines for the Care and Treatment of Breast Cancer. PMID- 11258213 TI - Problems for clinical judgement: 1. Eliciting an insightful history of present illness. AB - This article presents the results of a review of studies of psychology that describe how ordinary human reasoning may lead patients to provide an unreliable history of present illness. Patients make errors because of mistakes in comprehension, recall, evaluation and expression. Comprehension of a question changes depending on ambiguities in the language used and conversational norms. Recall fails through the forgetting of relevant information and through automatic shortcuts to memory. Evaluation can be mistaken because of shifting social comparisons and faulty personal beliefs. Expression is influenced by moods and ignoble failures. We suggest that an awareness of how people report current symptoms and events is an important clinical skill that can be enhanced by knowledge of selected studies in psychology. These insights might help clinicians avoid mistakes when eliciting a patient's history of present illness. PMID- 11258214 TI - Malaria deaths in visitors to Canada and in Canadian travellers: a case series. AB - Over the last decade there has been a marked increase in case of drug-resistant and severe malaria in Canadian travellers. We report 7 deaths due to falciparum malaria that occurred in Canada or in Canadian travellers. Risks for malaria infection include inappropriate recommendations for malaria prevention by health care providers and lack of knowledge about or adherence to appropriate recommendations by the travelling public. Risks for death include delays in seeking medical attention, delays in diagnosis and inadequate care by Canadian physicians and hospitals, and lack of access to parenteral therapy for severe malaria. Malaria infections and deaths are preventable. Better education of health care providers and travellers about the risks of malaria and appropriate prevention and treatment measures may decrease this unnecessary burden on the Canadian health care system. PMID- 11258216 TI - Family sues after negligence settlement against MD goes unpaid. PMID- 11258217 TI - A 550-year-old microbiology lesson emerges from BC glacier. PMID- 11258218 TI - Eating disorders present tricky diagnosis. PMID- 11258219 TI - Fewer antibiotics prescribed to children. PMID- 11258220 TI - All quiet on the flu front in 2001. PMID- 11258221 TI - Invest in children and reap the rewards: UNICEF. PMID- 11258222 TI - A crisis aborning in maternity and newborn care? PMID- 11258215 TI - The role of angiogenesis in prostate and other urologic cancers: a review. AB - Angiogenesis is a process critical to both tumour growth and metastasis. It is a dynamic integrated process involving basement membrane degradation, endothelial cell proliferation and migration, and capillary tubule formation. Under normal circumstances, the microvasculature is maintained in a quiescent state. The acquisition of the angiogenic phenotype depends on the outcome of stimulatory and inhibitory regulation by the tumour and its microenvironment. There are markers of angiogenesis that potentially could provide prognostic information in addition to that gained from conventional clinicopathologic data, and antiangiogenic therapy for urologic cancers has potential advantages over current therapeutic strategies. Promising preclinical studies have led to the initiation of phase I studies of antiangiogenic therapy in combination with chemotherapy, which may lead to novel treatments for urologic malignant tumours and may identify new intermediate markers for the response to therapy. PMID- 11258223 TI - Feds announce plan to reduce domestic use of pesticides. PMID- 11258224 TI - The Internet and genetic disease. PMID- 11258225 TI - Beyond bone marrow: a new source of stem cells. PMID- 11258226 TI - Ebola erupts again. PMID- 11258227 TI - Separated twins and the genetics of personality differences: a critique. AB - This article discusses studies of separated twins, with special emphasis on the Minnesota Study of Twins Reared Apart (MISTRA), to determine whether they support the existence of an important genetic component in behavioral and personality differences. The methods and conclusions of the MISTRA team are discussed in the context of earlier studies of separated identical twins. I argue that volunteer based studies are biased toward greater twin similarity. In addition, the MISTRA research team did not publish or share raw data and case history information. Reared-together and reared-apart monozygotic twins share important environmental similarities not controlled for by comparing personality correlations. I propose an alternative control group consisting of biologically unrelated pairs of strangers matched on all environmental factors common to pairs of separated monozygotic twins. I conclude that the evidence from studies of twins reared apart does not support the role of genetic factors in personality and behavioral differences. PMID- 11258228 TI - Effects of familiarity level and repetition on recognition accuracy. AB - Theories of recognition memory based on signal detection theory posit that a recognition decision is similar to a psychophysical judgment. Like a judgment of stimulus brightness or loudness, a recognition judgment is based on the value of a unidimensional signal computed for the test item. This signal has been called the strength or familiarity value. One prediction of these models is that the ability to discriminate between a studied and nonstudied test item depends on the ability to detect the difference in their familiarities. This ability in turn is influenced by the items' familiarity levels; discrimination should become more difficult as the familiarity of both items increases. This prediction was supported in 3 experiments using a forced-choice procedure. Also, accuracy was higher when the list contained repeated items rather than a comparable number of distinct items. PMID- 11258229 TI - A constraint on eliminating semantic priming by repeating a prime. AB - Repeated semantic primes have been shown to generate an additive semantic priming effect when those primes are different words, but repetition priming of the prime seems to eliminate semantic priming of lexical decisions for reasons that are not clear. These studies replicated this effect but also showed that repetition priming of the prime does not eliminate semantic priming with a longer lag between the repeated primes. The implications of these results for understanding the basic effect are discussed. PMID- 11258230 TI - Memory of psychodiagnostic information: biases and effects of expertise. AB - Problem-solving expertise has been associated with enhanced memory of domain specific information. This enhanced memory is thought to play an important role in expert decisions. Meanwhile, research on psychodiagnostic decision making has found consistent limitations in experienced clinicians' ability to make optimal decisions. To what extent are these limitations associated with suboptimal memory processes? We compared memories of expert clinicians and novice graduate students for information learned while viewing a videotaped psychodiagnostic interview. Results of 3 tests suggest that expert clinicians exhibit enhanced memory that is flexible, selective, and accurate but with limitations that might contribute to poor decisions. Experts exhibited superior memory of personal criteria and disconfirmatory information. However, a framing manipulation induced performance in experts consistent with suboptimal decision making, and both groups needed exhaustive prompts for optimal memory search. Implications of these findings for expertise models are discussed. PMID- 11258231 TI - The roles of attention and awareness in the false recognition effect. AB - Jacoby and Whitehouse (1989) reported that masked words can unconsciously bias memory judgments. Others suggest that these results cannot be taken as evidence that words are perceived without awareness, as Jacoby and Whitehouse claimed. In the present experiments, participants' ability to attend to briefly presented context words was manipulated by masking the context words and by an instructional manipulation. In Experiment 1, masked context words increased false recognitions. Different from Jacoby and Whitehouse's results, nonmasked context words showed a similar pattern. In Experiment 2, these effects were replicated and an attended condition was added in which participants read the context words aloud and tried to remember them. False recognition was increased in both masked and unmasked conditions and decreased in the attended condition. These results suggest that the condition for increase or decrease in false recognition is not whether a stimulus is seen or not but whether a stimulus is attended. They suggest that the qualitative difference criterion proposed by Jacoby and Whitehouse is insufficient for determining whether participants are aware of masked presentations. PMID- 11258232 TI - Healthcare information security. The threats and the safeguards--and how to manage them. AB - As more and more hospitals have begun storing patient information in electronic form, data security has become a hot topic. With the passage of the Health Insurance Portability and Accountability Act (HIPAA) in the United States, as well as other security legislation across the globe, even more attention has been focused on the subject. In this article, we discuss the various types of security threats faced by healthcare institutions, from external attacks via the Internet to internal violations resulting from malice or simple carelessness. We also discuss what you can do to maintain the privacy and integrity of your electronic records--not only the computer-based safeguards that are available, but also the internal systems and procedures you need to have in place. We tell you how to go about setting up (or beefing up) your institutional security system. And we describe the security efforts being made around the world by governments and standard-setting organizations. This article also includes an extensive glossary of computer-security terms and a listing of useful data-security Web sites. PMID- 11258233 TI - Independence 3000 IBOT Transporter. A major advance in powered wheelchairs. PMID- 11258235 TI - Hazard report. Agilent Information Center (formerly Hewlett-Packard VIC) monitoring systems: are your speaker plugs secure? PMID- 11258234 TI - DICOM reference guide. AB - The Digital Imaging and Communications in Medicine standard, known as DICOM, exists to facilitate communications between imaging devices and systems. By dictating specific data and interface requirements, DICOM helps ensure that devices--particularly devices made by different suppliers--can communicate with one another. Unfortunately, because different devices can implement the standard in different ways, not all devices that comply with the standard will be compatible. Thus, it's not enough for healthcare facilities to determine whether a device complies with the DICOM standard; they must also analyze how each device complies with the standard. This Reference Guide is intended to provide readers with the background information they need to perform such analyses. The Guide includes the following articles: THE BASICS: This article provides a brief overview of DICOM for the benefit of readers unfamiliar with the standard. THE DETAILS: This article focuses on the specific DICOM components that readers will need to understand when working with the standard. EVALUATING DICOM CONFORMANCE STATEMENTS: This article describes how to determine whether, and to what degree, two devices will support communication with one another. In addition to offering detailed, step-by-step guidance, we include a DICOM Compatibility Matching Form to help readers organize the data they'll be analyzing during the evaluation process. PMID- 11258236 TI - User experience network. Erroneous date/time annotation for Hewlett-Packard VIC monitoring systems. PMID- 11258238 TI - Disease management over the Internet reduces hospitalization. PMID- 11258237 TI - HIPAA may produce better cash flow for hospitals. PMID- 11258239 TI - Recruiting effective hospital board members is becoming more difficult. PMID- 11258240 TI - Spending more on Medicare is top priority for 33% in pre-Bush poll. PMID- 11258241 TI - Retention may be key to nurse shortage. PMID- 11258242 TI - AHA, WEDI, consultants, HHS offer HIPAA resources on the Web. PMID- 11258243 TI - Interstudy report shows providers give larger discounts to national PPOs. PMID- 11258244 TI - [Ventricular cancer--still a problem]. PMID- 11258245 TI - [Mechanisms in osteoporosis]. PMID- 11258246 TI - [Curative treatment of ventricular cancer]. AB - Extended lymphadenectomy on connection with the surgical treatment of gastric cancer is gaining access in western centres especially since Japanese centres have shown an ever increasing rate of survival over several decades, coupled with the fact that operative procedures have become more sophisticated. The latest prospective studies in the west seem to confirm the value of lymphadenectomy in some patients. Furthermore, correct staging demands extended lymphadenectomy. For patients with gastric cancer, adjuvant preoperative chemotherapy is probably an asset. PMID- 11258247 TI - [Intervention for apoplexy patients discharged from hospital. Psychosocial support: a literature review]. AB - Psychosocial support has been suggested as a way of easing stroke survivors' and their carers' adjustment to a life with disability. The literature on psychosocial support services following primary rehabilitation in hospital was reviewed. Eleven controlled studies evaluating the effect of psychosocial support interventions after discharge from hospital were identified. The studies differed widely with respect to design, intervention and evaluation methods. The results suggest that psychosocial support after discharge can improve psychological well being and quality of life for stroke survivors and their families and improve the social activity of patients. The effect was achieved by using different types of intervention such as providing information, counselling and support from stroke clubs. Psychosocial support for carers was effective as well. Future research should elucidate this area, including evaluation of psychosocial support as a tertiary prevention strategy. PMID- 11258248 TI - [Intervention for apoplexy patients discharged from hospital. Physical training: a literature review]. AB - Stroke is among the leading causes of disability in Denmark. Rehabilitation services are in the process of being reorganised into dedicated stroke units. There is a general tendency toward reduction of length of in-patient treatment. The literature on outpatient rehabilitation services following primary rehabilitation on an inpatient basis was reviewed. The results of 16 randomised studies indicate that: 1) Continued rehabilitation after discharge can improve functional capacity of disabled stroke survivors; 2) Home-based rehabilitation is as effective as hospital-based outpatient rehabilitation; 3) Early supported discharge (ESD) services can reduce length of hospital stay but the relative advantages and drawbacks remain unclear. Ongoing rehabilitation by teams specialised in stroke rehabilitation seems to be crucial. More research, including evaluation of home-based rehabilitation services, is called for and existing outpatient rehabilitation services should be evaluated scientifically. PMID- 11258249 TI - [Surgical treatment of stomach cancer at a university hospital. A retrospective survey of the period 1990-1995]. AB - AIM: A retrospective study of 69 cases of gastric cancer seen during the period from 1/1-1990 to 31/12-1994 treated in a University Hospital. The aim of the study was to describe morbidity, mortality and identify independent prognostic variables for mortality. METHOD: Patient data were recovered from the hospital's central database. Mortality was chosen as end-parameter. Univariate log-rank-test identified statistically significant variables which were then analysed by Cox backward stepwise regressional analysis. MATERIAL: Sixty-nine patients were available for analysis, median age 73 years. Fifty-one patients underwent operation. Eighteen patients did not have a surgical procedure due to disseminated disease. The overall postoperative morbidity was 25% and postoperative mortality 10%. The overall five-year survival rate was 8%, 12% for operated patients, 35% after radical and 0% after non-radical or omitted surgery. Age, radicality of operation, type of operation, Borrmann's tumour classification, and degree of depth of local infiltration were identified as significant factors for survival. Cox's analysis identified type of operation (p = 0.0002) and Borrmann's tumour classification (p = 0.001) as independent variables. DISCUSSION: The overall five-year survival is low and has not changed over two decades in Denmark, whereas mortality and morbidity rates have improved. It should be recommended that: The treatment of gastric cancer must be centralised in order to develop preoperative examinations, operative technique and the necessary routine for the surgeons. All gastric ulcers must be considered malignant and biopsies taken accordingly. PMID- 11258250 TI - [Danish endocrinologists' examination and treatment of non-toxic multinodular goiter. A questionnaire study]. AB - AIM: To assess the attitudes towards the management of the non-toxic multinodular goitre by means of a questionnaire concerning on case history, which was circulated to all Danish endocrinologists ("A 42-year-old woman with an irregular, non-tender, bilaterally enlarged thyroid of 50-80 g and no clinical suspicion of malignancy"). DESIGN: Eleven variations of the (basic) case report were proposed in order to evaluate the impact on the management of each alteration. In the index case, serum TSH was the routine choice of 100%, serum T4/free T4-index and T3/free T3-index were measured by 83% and 79%, serum TPOab and serum calcitonin by 49% and 4%, respectively. RESULTS: The median number of blood tests was three (range 1-7). Thyroid scintigraphy was used by 96% and ultrasound by 94%, both methods were employed by 90% with scintigraphy most often having the first priority. Fine-needle aspiration biopsy was not routinely used, unless a scintigraphically dominant "cold" area was present; 70% used ultrasound guidance. Radioiodine was the preferred treatment of 51%, surgery 7%, L-T4 4%, and no treatment 38%. DISCUSSION: In the case of young patients or factors predisposing to thyroid cancer, treatment preferences changed to surgery. The prevailing use of radioiodine in Denmark contrasts with the situation in most other European countries, but Danish endocrinologists also disagree about the optimum management of multinodular non-toxic goitre. PMID- 11258251 TI - [Comparison of monolayer specimens and conventional smears]. AB - INTRODUCTION: Monolayer specimens were compared with conventional smears in a split sample study. The quality of the specimens, sensitivity, time consumption, and costs were elucidated. METHOD: Conventional smears and fixated residual material from 1701 women were available. RESULTS: The number of inadequate/less adequate specimens was reduced significantly (p < 0.001). Diagnostic agreement was found in 1531 (90%) of the 1701 cases (kappa = 0.52, SE (kappa) = 0.026). Dysplasia (NOS) was diagnosed in seven monolayer specimens (0.4%) versus 33 smears (1.9%) and ratio atypia/low grade was reduced by 33%, which indicates greater accuracy in diagnosis in monolayers. Histological follow-up showed sensitivities of 95% (monolayers) and 94% (smears). The time consumed (laboratory work, screening) on a smear versus a monolayer specimen was 11.5 minutes versus 9.3 minutes. Utensil costs of a smear are kr. 10.69. compared to kr. 26.50 for a monolayer specimen. CONCLUSION: The higher costs should be set against the saving in significantly improved specimen quality, improved diagnostic accuracy, and shorter time consumption. The use of the monolayer technique (direct to vial) is recommended to replace the conventional smear. PMID- 11258252 TI - [Workload, gender and age among personnel with reported sudden lifting injuries]. AB - INTRODUCTION: In 1996 the Danish Parliament passed a new Act on industrial injuries, including the so-called sudden lifting injuries. This type of lifting injury had been experienced in some industries in connection with lifting situations causing sudden pain in the back. Less frequently, other regions of the body may be affected. This study is a distribution of a sample of reported industrial injuries of this type in groups according to types of lifts. The study also includes a comparison with parameters from the Danish Work Environment Cohort Study. MATERIAL AND METHODS: The sample consisted of a total of 482 cases of back injuries. Of these, 346 cases were distributed according to six categories of manual lifting work and to age and gender. The rest, 136 cases, were excluded for technical reasons. RESULTS: In the distribution, 8% of the cases were in jobs with many heavy lifts, 66% of the cases were in jobs with moderate daily lifted weights or in jobs with only few or light lifts. Approximately 25% of cases concerned employees performing patient transfers. The distribution contained more men than the Danish Work Environment Cohort Study, and the persons in the cases were slightly older than this reference population. DISCUSSION: Altogether, this study points towards an occurrence of sudden lifting injuries among employees performing certain types of manual lifting work. The industrial injuries cases may have been subject to some selection. PMID- 11258253 TI - [Utilization of health care system by patients with chronic pain who apply for disability pensions. A registry study]. AB - INTRODUCTION: Patients with chronic pain may be awarded a disability pension if their working capacity is reduced as a result of an illness. Patients with chronic pain very often consult several specialists in order to obtain proof of their illness. The aim of this study was to investigate how an application for disability pension affected the utilization of health care services by patients with chronic pain. METHOD: Patients with chronic pain who applied for a disability pension in 1989 and 1990 were entered. The study period was divided into three: the year preceding the submission of the application for a disability pension, the time from submission of the application to the making of the decision, and the year following the decision of the health authorities. The patients were divided into four subgroups according to whether the disability pension was awarded or rejected, and whether the patients accepted or appealed the decision. The total costs of care in the primary sector were calculated on the number of and charges for visits to GPs and the total costs of hospital care were calculated on the number of bed days, visits to outpatients clinics, operations, and investigations. RESULTS: Patients with chronic pain had a significantly lower health care utilisation after the case has been closed. Those who did not get a disability pension and those who were not satisfied with the level of the disability pension, continued their utilisation of health care after the decision. The mean health care used by patients who appealed the level of the disability pension was three times higher than that used by patients who accepted the level of the disability pension. CONCLUSION: The study indicates that lack of or insufficient economic compensation from the social system in patients with chronic pain may contribute to inexpedient behaviour leading to increased costs to the health care sector. PMID- 11258254 TI - [Work-related stress behind sudden death cases? Two young Swedes hit by a known Japanese phenomenon]. PMID- 11258255 TI - [Primary cardiac lymphoma]. AB - Primary cardiac non-Hodgkin lymphoma is very rare. Results recently published suggest that the prognosis is good, if the lymphoma is diagnosed early. The symptoms are nevertheless unspecific and a clinical investigation is often inconclusive. We report a case of a woman with symptoms of severe dyspnoea at rest, chest pain, and fatigue. The ECG showed a complete atrioventricular block. Magnetic resonance imaging (MRI) revealed a tumour in the right atrium and ventricle. A myocardial biopsy showed malignant non-Hodgkin lymphoma of the diffuse, large cell B-type. The patient was treated with chemotherapy and control MRI after four treatments showed complete regression of the tumour. PMID- 11258256 TI - [In the middle of the influenza time]. PMID- 11258257 TI - [Survey of vaccination complications]. PMID- 11258258 TI - [Non-disposable acupuncture needles are potential hazards of transmission of prion diseases]. PMID- 11258259 TI - [Center for epidemiologic basic research]. PMID- 11258260 TI - [Life style diabetes]. PMID- 11258261 TI - [Old knowledge about obesity and diabetes]. PMID- 11258262 TI - [Evaluation of research environment--how?]. PMID- 11258263 TI - [Wrong again!]. PMID- 11258264 TI - [Treatment of obesity and type 2 diabetes: New knowledge?]. PMID- 11258265 TI - Staying current on healthcare regulations. PMID- 11258266 TI - HIPAA's challenge to finance. PMID- 11258267 TI - "B-u-s-h" does not spell "relief" from federal investigations. PMID- 11258268 TI - Finance and quality go hand in hand. PMID- 11258269 TI - Strategic facility planning improves capital decision making. AB - A large, Midwestern IDS undertook a strategic facility-planning process to evaluate its facility portfolio and determine how best to allocate future investments in facility development. The IDS assembled a facility-planning team, which initiated the planning process with a market analysis to determine future market demands and identify service areas that warranted facility expansion. The team then analyzed each of the IDS's facilities from the perspective of uniform capacity measurements, highest and best use compared with needs, building condition and investment-worthiness, and facility growth and site development opportunities. Based on results of the analysis, the strategy adopted entailed, in part, shifting some space from inpatient care to ambulatory care services and demolishing and replacing the 11 percent of facilities deemed to be in the worst condition. PMID- 11258270 TI - Effective strategies for managing pharmacy benefits. AB - Prescription drug costs are among the fastest-growing healthcare costs. Effective plan design and benefit management strategies can help pharmacy benefit plans manage costs while maintaining quality and customer satisfaction. These strategies include using formulary management, intervention techniques, and cost sharing to encourage the use of generic drugs; employing a mail-service pharmacy benefit for maintenance medications; and implementing concurrent and retrospective review programs to ensure eligibility and plan compliance, identify practice patterns, and encourage appropriate drug selection. PMID- 11258271 TI - The bribery statute: a new weapon against Medicare fraud. AB - A May 2000 U.S. Supreme Court decision determining when a Federal bribery statute can be used to fight Medicare fraud has ramifications for healthcare providers. In Fischer v. United States, the Court concluded that healthcare providers that participate in Medicare are considered to receive benefits as set forth in the bribery statute and thus can be prosecuted for fraudulent activities against the government under the statute. The statute mandates a fine, imprisonment for up to 10 years, or both for anyone convicted under it. Provider organizations that receive Medicare payments and business associates of such organizations should be aware that the government may step up its use of the bribery law in prosecuting fraudulent activity. In addition, although the case pertained specifically to healthcare providers that participate in Medicare, providers that do not participate in Medicare may wish to evaluate the advisability of accepting other Federal funding because of the possible reach of the bribery statute. PMID- 11258272 TI - Designing and maintaining an effective chargemaster. AB - The chargemaster is the central repository of charges and associated coding information used to develop claims. But this simple description belies the chargemaster's true complexity. The chargemaster's role in the coding process differs from department to department, and not all codes provided on a claim form are necessarily included in the chargemaster, as codes for complex services may need to be developed and reviewed by coding staff. In addition, with the rise of managed care, the chargemaster increasingly is being used to track utilization of supplies and services. To ensure that the chargemaster performs all of its functions effectively, hospitals should appoint a chargemaster coordinator, supported by a chargemaster review team, to oversee the design and maintenance of the chargemaster. Important design issues that should be considered include the principle of "form follows function," static versus dynamic coding, how modifiers should be treated, how charges should be developed, how to incorporate physician fee schedules into the chargemaster, the interface between the chargemaster and cost reports, and how to include statistical information for tracking utilization. PMID- 11258273 TI - The FASB explores accounting for future cash flows. AB - The FASB's Statement of Financial Accounting Concepts No. 7, Using Cash Flow Information and Present Value in Accounting Measurements (Statement No. 7), presents the board's views regarding how cash-flow information and present values should be used in accounting for future cash flows when information on fair values is not available. Statement No. 7 presents new concepts regarding how an asset's present value should be calculated and when the interest method of allocation should be used. The FASB proposes a present-value method that takes into account the degree of uncertainty associated with future cash flows among different assets and liabilities. The FASB also suggests that rather than use estimated cash flows (in which a single set of cash flows and a single interest rate is used to reflect the risk associated with an asset or liability), accountants should use expected cash flows (in which all expectations about possible cash flows are used) in calculating present values. PMID- 11258274 TI - Meeting the challenge of a group practice turnaround. AB - Many healthcare organizations that acquired group practices to enhance their market share have found that the practices have not met their financial goals. Turning around a financially troubled, hospital-owned group practice is challenging but not impossible for healthcare organizations that take certain basic actions. Direction, data, desire, dedication, and drive must be present to effect the financial turnaround of a group practice. The healthcare organization needs to evaluate the practice's strategy and operations and identify the issues that are hindering the practice's ability to optimize revenues. Efforts to achieve profitable operations have to be ongoing. PMID- 11258275 TI - HIPAA's privacy standards require understanding and action. PMID- 11258276 TI - Networking is key to CFO career development. PMID- 11258277 TI - Data trends. Comprehensive data improves interpretation of indicators. PMID- 11258278 TI - Patient safety--financial & cultural implications. PMID- 11258279 TI - What Works Awards 2000. Maimonides Medical Center, Brooklyn, NY. PMID- 11258280 TI - What Works Awards 2000. High Desert Medical Group, Lancaster, CA. PMID- 11258281 TI - Safer, cheaper, smarter. Computerized physician order entry promises to streamline and improve healthcare delivery. PMID- 11258282 TI - Technology triumphs. PMID- 11258283 TI - Leaving money on the table? How comprehensive coding can raise practice revenues by $40,000 per physician per year. PMID- 11258284 TI - Market resource guide 2001. PMID- 11258285 TI - What Works Awards 2000. Brooke Army Medical Center, Fort Sam Houston, TX. PMID- 11258286 TI - Roadblock to reform. Democrats fight Medicare revamp that would give bigger role to private sector. PMID- 11258287 TI - A second chance. New public comment period gives industry a shot at reining in HIPAA. PMID- 11258288 TI - Failed mission. AMA loses members, revenue despite ambitious recruiting drive. PMID- 11258289 TI - Breaking up. After six years, Catholic hospital system and HCA decide to call it quits. PMID- 11258290 TI - Justice jumps on Tenet. Feds join kickback suit against Fla. hospital; case to test enforcement policies. PMID- 11258291 TI - Hospitals facing new assault from Washington. PMID- 11258292 TI - Guaranteed by Uncle Sam. Obscure FHA office backs hospital mortgages, providing big savings. PMID- 11258293 TI - IOM strikes again. Second report calls for transforming failing industry; money is big question. PMID- 11258294 TI - Budgeting the future. Bush plan proposes no Medicare payment cuts, puts aside money for drug benefit. PMID- 11258295 TI - Military matters. PMID- 11258296 TI - What's so unusual? AB - You're probably asking yourself, "What's so different about this case?" The patient was on the side of the road, explained what she was doing there, denied any injuries and refused treatment and transport. Sounds like a routine call- what's the big deal? But this case did not end after the patient signed a treatment refusal form and was asked by law enforcement to get back in her car and drive on. The EMS providers returned to their station, only to get called back 30 minutes later to the same spot they had just responded to, again for a "woman down." Upon this arrival, they found the same car on the side of the road, and a parked semi tractor trailer in a traffic [figure: see text] lane approximately 25 yards ahead of the vehicle. There was no damage to either vehicle, and providers noticed a familiar dog inside the woman's car. A police officer informed the providers that the woman, the driver of the vehicle, had jumped in front of the semi, as described by the truck's driver, and was hit at a very high rate of speed. The victim was deceased on the scene. Upon assessing the scene, providers confirmed that the victim was the same woman they'd responded to earlier. Several days after the incident, more information about the patient's medical history was released. She had been released on the morning of the accident from a mental health institution, where she'd been diagnosed as a manic depressive. Manic depression is a condition characterized by mood swings. For example, a patient might rapidly go from a state of euphoria to experiencing debilitating depression. Although this patient appeared fine on the exterior, she had been diagnosed with a severe psychological disorder. She presented as being alert, oriented and non-threatening moments before taking actions that led to her death. This case posed a significant challenge for the EMS providers because there was no clinical evidence that there was anything wrong with the patient. Moreover, the patient denied any past medical history. It was a lesson to all involved with this case: There is no such thing as a routine call. PMID- 11258297 TI - Maintenance matters. PMID- 11258298 TI - Dopamine made easy. PMID- 11258299 TI - Spinal immobilization. PMID- 11258300 TI - From provider to patient: injured in the line of duty. PMID- 11258301 TI - So you want to be a paramedic. PMID- 11258302 TI - Strategies for becoming a flight crew member. PMID- 11258303 TI - Offshore medicine. AB - Offshore life can be refreshing for medics who are looking for a little change of pace; however, it is not for everyone. Working offshore can be the easiest or most boring job you'll ever have. It takes a specific type of medic to fit this mold. So, if you are considering a career in the offshore field, take all of the above into consideration. You are not just making a change in jobs, but a change in lifestyle. Once you become accustomed to this lifestyle, it will be hard to go back to the everyday hustle and bustle of the streets. For more information about working offshore, contact Acadian Contract Services at 800/259-333, or visit www.acadian.com. PMID- 11258304 TI - Emergency assistance for the interviewee. PMID- 11258305 TI - Prehospital management of hypothermia. PMID- 11258306 TI - Student field performances: how valid are your preceptors' evaluations? AB - Developing relevant performance standards and rating criteria and preparing preceptors for the role of evaluator are critical to establishing valid student field performance evaluations and providing assurance to educational institutions that graduating students truly are competent and ready to assume an entry-level position. Using scripted, pre-scored video case studies in conjunction with written guidelines for rating the performance standards and providing specific feedback to explain rationale can significantly improve interrater agreement between preceptors. PMID- 11258307 TI - The burn wheel: a guide to burn resuscitation. PMID- 11258308 TI - Hearts out, hands up to the helpless of Honduras. PMID- 11258309 TI - 9-1-1: the Achilles' heel of EMS. PMID- 11258310 TI - [Idiopathic portal hypertension]. AB - The authors present a report of 14 patients with the syndrome of portal hypertension without liver cirrhosis and with recurring esophagogastric bleedings. The cause of the alterations in portal hemodynamics remains unknown. Operative treatments (splenorenal shunts in 9 cases, 3 splenectomies and 1 ligation of the splenic artery) were successful. Two patients in whom splenectomy had been performed in combination with omentohepatopexy died 6 and 10 years after operation due to recurrent hemorrhages. The other patients did not have recurrent bleedings, but in 6 patients 6-10 years after splenorenal shunts there appeared other diseases (encephalopathy, nephrolithiasis, arterial hypertension, duodenal ulcer). The authors consider that indications for shunting operations for idiopathic portal hypertension, especially when using renal veins, should be determined more carefully, phlebosclerotic therapy and transsection of the esophagus being recommended as alternative interventions. PMID- 11258311 TI - [The 3d Congress of the Russian Association of Endoscopic Surgery (Moscow, 24-25 February 2000)]. PMID- 11258312 TI - [V.I. Kolesov: a coryphaeus of Russian surgery]. PMID- 11258313 TI - [Diagnosis and surgical treatment of mediastinal neoplasms]. AB - The authors share their experiences with examination and surgical treatment of 163 patients with various neoplasms of the mediastinum. The first place among the neoplasms is occupied by tumors of the lymphatic apparatus (33%), the second--by tumors of the thymus (21%). The video-thoracoscopic technique was used in 17 of 131 operations performed. Complications after the operations took place in 6 patients (4.5%), two patients died (1.5%). A conclusion is made that the patients of this category must be treated not only by thoracic surgeons but also by hematologists, neurologists, neurosurgeons and specialists in radiation therapy. PMID- 11258314 TI - [Methods of restoration of patency of the main respiratory pathways]. PMID- 11258315 TI - [Preoperative embolization of the portal vein in liver tumors]. PMID- 11258316 TI - [Use of bacteriophage therapy in surgical practice]. PMID- 11258317 TI - [Use of cytochrome C in the prevention of myocardial reperfusion injury during heart valve prosthesis implantation under conditions of extracorporeal circulation]. AB - Cytochrome C used at the postischemic period causes a rapid reestablishment of coronary circulation, hemodynamic parameters, prevents activation of lipid peroxidation in reoxygenation of the heart after prolonged total myocardial ischemia in patients with acute bacterial endocarditis in whom the prostheses of heart valves had been fulfilled. Parallel with the positive inotropic effect, Cytochrome C reduces the postloading and, as a result, transfers the cardiac muscle to a more profitable regimen. At the end of the myocardial ischemia period Cytochrome C provides rapid and effective recovery of the bioelectrical function of the heart, improves its pumping function, thus allowing the dose and duration of inotropic stimulation of the myocardium at the postischemic period to be reduced, lowers pulmonary resistance and finally leads to less postoperative intrahospital lethality from acute heart failure. PMID- 11258318 TI - [Diagnosis and surgical treatment of postbulbar ulcers of the duodenum]. AB - The method of surgical treatment of postbulbar ulcers of the duodenum based on the data of their localization, complications, functional state of the stomach and duodenum, technical skills of the surgeon includes radical ablation of the ulcer, reestablishment of the gastroduodenal continuity with the formation of the functionally active anastomosis or selective proximal vagotomy. The method allows to considerably lessen the amount of complications at the early postoperative period and to obtain good functional results at late terms of observations. PMID- 11258319 TI - [Minimally invasive technologies in the surgical treatment of patients with mechanical jaundice and suppurative cholangitis]. AB - Minimally invasive technologies were used in treatment of 89 patients with cholelithiasis complicated by mechanical jaundice and purulent cholangitis. Advantages of the two-stage operative treatment of such patients were shown. The first stage is decompression of the bile duct, the second stage is a radical operation either by laparoscopic or minilaparotomic methods. It allows to avoid extensive and traumatic operations and secures better immediate and long-term results of treatment. PMID- 11258320 TI - [Long-term results of the use of different modifications of the partial ileal shunt operation in the treatment of patients with atherogenic dyslipoproteinemias]. AB - The article presents long-term (up to 20 years) results of performing the classical H. Buchwald operations and its various modifications in 145 patients. Creation of ileoileoanastomoses 5 cm from the ileocecal valve and a standard shunt of 250 cm of the ileum allowed to obtain a necessary and considerable (27% at an average) hypocholesterolemic effect and to eliminate diarrhea at the postoperative period. PMID- 11258321 TI - [Prognostic factors influencing results of the surgical treatment of pancreatic cancer]. AB - In patients with carcinoma of the pancreas 128 gastropancreatoduodenal resections (GPDR), 15 distal resections of the pancreas and 3 pancreatectomies were performed. After GPDR 5-year survival was 12%, the survival median was 24.3 months. Only one patient is living 6 years after left-sided resection and pancreatectomy. Long-term results of the operative treatment for carcinoma of the pancreas depended on the amount of regional metastases, degree of differentiation of the tumor, its size and invasion into the vessels. The long-term results were considerably worse if the tumor was localized in the uncinate process, body and tail of the pancreas. The 5-year survival was noted mainly in patients with the 0 and I stages of the disease. It shows the early diagnostics to be necessary. PMID- 11258322 TI - [Differential diagnosis and surgical treatment of colorectal nonepithelial tumors]. AB - An analysis of results of the examination and treatment of 92 patients with nonepithelial colorectal tumors has shown that the correct diagnosis can be made in 82% of the cases on the basis of using complex X-ray, ultrasonic and endoscopic methods. Of decisive significance in determination of the tumor tissue character is the morphological investigation. Extensive surgical interventions were performed due to suspected malignization on 21 patients with benign nonepithelial tumors. In other cases (n = 49) the organ-saving operations were fulfilled. The decision between the volumes of the surgery for malignant nonepithelial tumors (n = 24) was not difficult in most cases and depended on the spread of the oncological process. The 5-year survival in this group of patients was 57%. No recurrences were noted after ablation of benign tumors. PMID- 11258323 TI - [Specialized surgical care of severe mechanical multiple trauma]. AB - The authors share their experiences with diagnosing and surgical treatment of 1792 patients with mechanical combined traumas. The average age of the patients was (31.6 +/- 1.4) years. All the patients were divided into two groups according to the medico-diagnostic methods. The traditional methods of diagnosis and treatment were used in 811 patients of the first group. Complications took place in 39.2%, lethality was 28.4%. The diagnosis in 981 patients of the second group included an ultrasound investigation, computed tomography, videothoraco- and laparoscopy. The surgical treatment was based on the objective multifactorial assessment of the trauma severity and determination of the prognosis criteria. Complications developed in 24.8% of the patients. Lethality was 19.5%. PMID- 11258324 TI - [Indications and results of artificial pacemaker implantation in children]. AB - Since 1989 till 1999 operations were performed on 74 children aged from 13 months to 15 years. There were 130 operations, 56 reoperations included. Iatrogenic atrioventricular blockades were diagnosed in 47 children (63.5%), congenital and postmyocarditic alterations in the rhythm and cardiac conductivity--in 27 children (36.5%). In 48 patients the primary implantation of electrocardiostimulators was fulfilled using myocardial electrodes, in 26 cases the endocardial method was used. Three children died at the postoperative period. The causes of death were the growing heart failure, hypoxic brain edema, ventricular fibrilation. ECG, Holter monitoring, echocardiography with dopplerography were made in the children in order to solve the question of the implantation of artificial pace-makers. At the postoperative period the electrocardiostimulators' parameters were selected individually under control of echocardiography. PMID- 11258326 TI - [Development of navigation technologies for ambulatory surgery]. AB - The article considers the main directions in the development of navigation surgical technologies allowing to decrease traumatic effects of certain operative interventions and to fulfill them in "one day" hospitals. The historical aspects of navigation surgery are given as well as a classification of the navigation interventions and examples of the newest of them used at the present time in the Medical Center of Bank of Russia. PMID- 11258325 TI - [Total esophagoplasty in esophageal strictures]. AB - An analysis of immediate and long-term results of surgical treatment of 170 patients with esophageal obstruction is presented. Esophagoplasty with different fragments of the gastrointestinal tract was used (87--ileocolon, 16--small intestine, 23--left half of the colon, 44--a stalk from the greater curvature of the stomach). The greater amount of complications due to incompetence of the esophageal anastomosis sutures was noted in patients after esophagoplasty with a fragment of the small (50%) and right half of the large (59.7%) intestine, more rarely-after plasty with a gastric stalk (15.9%) and left half of the colon (26.1%). There was no necrosis of the transplant from the left half of the colon. Severe ischemic lesions made their appearance in the fundal portion of the gastric stalk (2.2%), necrosis of the small intestine part of ileocolon developed in 16.1% of the patients. In the patients with less amount of postoperative complications one-stage esophagoplasty was performed. Reconstructive operations on the artificial esophagus were necessary in 43.6% of the patients. The main indication for it was mechanical dysphagia. PMID- 11258328 TI - [Iatrogenic corpora aliena]. AB - For 9 years (1975-1983) in republic Karelia 95,515 operations were performed. For this period there were 8 cases when foreign bodies were left in the abdominal and thoracic cavities and in soft tissues of the anterior abdominal wall: Bujalski spatula, a hemostatic forcepts, five gauze drapes, two rubber drainage tubes and two gauze globules. The time of staying of the foreign bodies in the cavities or tissues varied from 1-14 days to 33 years. Out of all the operated patients who had those foreign bodies one patients died due to a wound of the heart (during thoracotomy a gauze drape was left in the pleural cavity). PMID- 11258327 TI - [Evaluation of the effectiveness of drainage of the abdominal cavity]. AB - In order to generalize the indications for drainage of the abdominal cavity the authors have systematized and subdivided them into one-type groups depending on the aims, (medical, prophylactic) and tasks, which on their opinion can allow the surgeons to better realize the reasons of using this method of treatment and to critically assess it. An analysis of clinical material was made in order to find out the frequency and localization of intraabdominal abscesses and to assess the measures taken for their prevention. It was found that in spite of the abdominal cavity drainage after 1568 operations on emergency indications, abscesses developed in 43 patients. An analysis of the investigation has shown that the causes, frequency and localization of postoperative abscesses are not predictable, prophylactic measures have low efficiency due to using passive glove tube drainage and to too frequent using this not safe method without special need. PMID- 11258329 TI - [Concept of epidural and associated combined spinal-epidural anesthesia concerning prevention of a neurodystrophic component of intra- and postoperative complications in patients subjected to thoracic and abdominal surgery]. AB - The stimulation of reflexogenic zones and interoreceptors in experiments and in patients during operations on the heart, lungs and organs of the gastrointestinal tract induces hyperactivation of the sympatho-adrenal system and associated with it neurogenic dystrophic alterations of the internal organs (markedly decreased level of noradrenaline, destruction of the cell ultrastructure, mitochondria in particular) and energy metabolism. The use of high epidural anesthesia as the main component of narcosis in operations on the heart (mitral commissurotomy) and on the lung prevents the development of intra- and postoperative complications, the above mentioned dystrophic alterations in particular. Similar protecting effects in the abdominal operations are obtained when using associated combined spinal-epidural anesthesia. PMID- 11258330 TI - [Prognostication of the outcome of heart valve prosthesis implantation by the method of nonlinear chaos dynamics of the heart rate]. AB - An examination of 39 patients with heart diseases was performed before and after operations by the method of geometrical analysis of the nonlinear chaos (fractal) variability of the cardiac rhythm on the basis of the apparatus-programmed complex "Poly-spectrum". Five-minute-long registrations of ECG were carried on. On the basis of the data of examination of 195 healthy volunteers the reference norm of this method parameters was determined. Reliably lower parameters were found in patients with the acquired valvular disease. A dynamic investigation of the geometrical nonlinear structure of the cardiac rhythm has shown the possibility to make a prognosis of early complications after prosthetics of the heart valves. PMID- 11258331 TI - [New approach to correction of ventricular septal defects complicated by critical pulmonary hypertension]. AB - The authors have studied the effect of preoperative administration of Vazaprostan 20 into the pulmonary artery of patients with a defect of the interventricular septum (DIVS) and high pulmonary hypertension. By the end of the second day the patients examined had lower pressure in the pulmonary artery from 18% to 38%, higher intracardiac output in patients with the III A and III B degrees of pulmonary hypertension. In all the patients plasty of the DIVS was fulfilled under conditions of hypothermal extracorporeal circulation. The degree of residual pulmonary hypertension in the group under study was 42% at an average, and in the control group it was 50%. The preoperative administration of Vazaprostan-20 decreased the operative risk and facilitated the postoperative period. It allowed to make operations on the wider group of patients whose possibility to be operated on was doubtful. The patients of the group under study were discharged from the hospital in a satisfactory state. PMID- 11258332 TI - [20-year experience with diagnosis and treatment of different forms of purulent mediastinitis]. AB - An experience of many years has shown that the most severe clinical course is observed in patients with mediastinitis due to injuries of the esophagus, and with odontogenous and tonsillogenous mediastinitis. Using the ultrasound method of investigation and computed tomography made the possibility to early diagnose this disease considerably wider. Treatment of patients with mediastinitis included surgical sanitation of the purulent focus, antibacterial, detoxicating and immune therapy, the central role being given to active drainage of the mediastinum with lavage and aspiration. In patients with mediastinitis resulting from rupture of the inferiorthoracic portion of the esophagus the isolation of the injured wall of the esophagus from the aggressive influence of gastric juice is an important measure. This problem can be solved by ligation of the abdominal part of the esophagus or by Nissen's fundoplication. Severe complications of mediastinitis such as empyema of the chest, purulent peritonitis and arrosive bleedings require active surgical strategy. PMID- 11258333 TI - [Intratracheal lymphotropic ozone therapy in erosive-ulcerous tracheitis]. AB - Under observation there were 452 patients with chronic stenosis of hollow organs of the neck having cannulas during the period from 3 months to several years. During the tracheobronchoscopic examination it was found that 35 patients had trachea ulcers, 46 patients had erosive tracheobronchitis. The ulcers were localized on the anterior wall of the thoracic part of the trachea. Their diameter was from 1 to 2.5 cm. The ulcers were accompanied by diffuse bilateral bronchitis of the II-III degree of the inflammation intensity. 2-3 ml of ozonated sodium chloride solution with the concentration of ozone in it 5 mg/l were introduced into the ulcer edges, i.e. lymphotropically, into the submucous membrane. The same solution (40-60 ml) was used for daily sanitation of the tracheobronchial tree. Complete epithelization of the ulcers and cleansing of the bronchial tree took 3-4 curative bronchoscopies. PMID- 11258335 TI - [Cancer of the large intestine complicated by inflammatory process and perforation]. AB - Based on an analysis of treatment of 293 patients with carcinoma of the colon complicated by the inflammatory process and perforation the data are presented concerning the frequency, causes of appearance, differences in clinical manifestations and strategy of treatment depending on the degree of the inflammatory-destructive process. The differentiated approach to treatment for different forms of inflammatory complications allowed lethality to be decreased to 13%. PMID- 11258334 TI - [Use of sodium hypochlorite and intravascular laser irradiation of blood in complex treatment of emergency surgical patients]. AB - Results of treatment of 50 patients with urgent pathology of organs of the abdominal cavity are presented. Indirect electro-chemical detoxication of blood against the background of the antioxidant protection of organism was applied with the purpose of detoxication. The problems of a possible side effect of the indirect electro-chemical detoxication of blood, the optimum dosage of sodium hypochlorite in the intravenous injection were investigated. PMID- 11258336 TI - [Multiple stenting of the right coronary artery in a patient with ischemic heart disease]. PMID- 11258337 TI - [Association of 2 autoimmune diseases with unusual clinical-laboratory manifestations and benign tumors of the liver]. PMID- 11258338 TI - Proceedings of the 3rd Symposium on Molecular Diagnostics in Laboratory Medicine 3-6 May 2000. PMID- 11258339 TI - Growth failure and decreased ultrasound parameters of bone density in children with inflammatory bowel disease. PMID- 11258340 TI - Bibliography. Current World Literature. Neuro-ophthalmology and neuro-otology. PMID- 11258341 TI - Bibliography. Current world literature. Cerebrovascular disease. PMID- 11258342 TI - Reasons for the slow implementations of CHART for patients with non-small cell lung cancer in clinical practice. PMID- 11258343 TI - Accelerated radiotherapy vs. chemoradiation in non-small cell lung cancer: quantifying the hazards. PMID- 11258344 TI - Smoking cessation: still looking for a miracle. PMID- 11258345 TI - Offering CF carrier screening: who set the goal, and what is the goal? PMID- 11258346 TI - ACMG's continuing medical education program: a new initiative. PMID- 11258348 TI - Structural anomalies revealed by neuroimaging studies in the brains of patients with neurofibromatosis type 1 and large deletions. AB - PURPOSE: The basis for cognitive problems in patients with neurofibromatosis type 1 (NF1) is unknown. A subset of NF1 patients with deletion of the entire NF1 gene has severe learning problems or mental retardation. We have reviewed neuroimaging studies (CT and MRI) in five such patients to determine whether structural anomalies in the brain are present and might explain the impaired cognitive function. METHODS: Five patients with NF1 and deletion of the entire gene were identified by FISH studies. A retrospective review was conducted of CT and MRI images, as well as of data from developmental assessments. RESULTS: All five patients had severe developmental impairment. None had been exposed to chemotherapy or radiation therapy. All had multiple regions of bright T2 signal intensity. Structural anomalies were seen in three of the five patients and included callosal dysgenesis in one, septum cavum vergae and pellucidum in two, mega cisterna magna in one, and Chiari I malformation with severe hydrocephalus in one patient. CONCLUSION: Individuals with NF1 and large gene deletions have an increased frequency of structural anomalies of the brain not usually seen in NF1 patients. This suggests that the mental retardation in these individuals is due, at least in part, to abnormal brain development rather than a defect in brain function due to haplosufficiency of the NF1 gene product. PMID- 11258349 TI - Analysis of imprinted genes in subjects with Prader-Willi syndrome and chromosome 15 abnormalities. AB - PURPOSE: To determine gene expression of five imprinted genes or transcripts from the 15q11-q13 chromosome region using reverse transcription polymerase chain reaction (RT-PCR) in a relatively large survey of Prader-Willi syndrome (PWS) and control subjects with several different chromosome 15 abnormalities. METHODS: RT PCR was undertaken on mRNA isolated from tissue (e.g., mostly lymphoblasts) from 38 PWS and 10 control subjects. DNA primers were used for five imprinted genes or transcripts (ZNF127, SNRPN, PAR5, IPW, and PAR1) from 15q11-q13 and fibrillin, a control gene from 15q21. RESULTS: One PWS subject with maternal disomy 15 showed weak but detectable expression of PAR1, whereas SNRPN expression was detected in two PWS subjects [one with the 15q11-q13 deletion and one with a t(15;15) karyotype and maternal disomy 15], and the remaining typical PWS subjects showed no expression of the imprinted genes or transcripts. CONCLUSION: No obvious clinical differences were identified in those PWS subjects with weak expression of genes compared with those showing no expression. Although the reason(s) for weak expression is unknown, possible explanations include relaxation of imprinting caused by failure to reset the imprinted genes or transcripts in the maternal germ line or by postzygotic gene expression or undetected chromosome 15 mosaicism in the deletion PWS subjects. The timing, tissue source, and other factors relating to partial expression of genes that are thought to be imprinted may play a role in clinical variability and allow for a better understanding of molecular mechanisms in PWS and other abnormalities of proximal chromosome 15q. PMID- 11258347 TI - Issues in implementing prenatal screening for cystic fibrosis: results of a working conference. AB - PURPOSE: To summarize a conference convened to examine how cystic fibrosis screening might appropriately be introduced into routine prenatal practice. METHODS: Participants included experts from various relevant disciplines. Systematic reviews and data from individual trials were presented; issues were identified and discussed. RESULTS: Judged by published criteria, prenatal cystic fibrosis screening is suitable for introduction. Screening can be performed cost effectively by identifying racial/ethnic groups at sufficient risk and then using either of two models for delivering laboratory services. Validated educational materials exist. Ethical issues are not unique. CONCLUSIONS: Once adequate facilities for patient and provider education, testing, counseling, quality control, and monitoring are in place, individual programs can begin prenatal screening for cystic fibrosis. PMID- 11258350 TI - Long-term outcome in treated combined methylmalonic acidemia and homocystinemia. AB - PURPOSE: To describe the clinical and biochemical features and long-term outcome of a cohort of eight patients with methylmalonic acidemia and homocystinuria (cblC). METHODS: Documentation of clinical features at birth and longitudinal follow-up of the biochemical and clinical response to treatment with daily oral carnitine and intramuscular hydroxocobalamin observed during continuous follow-up for an average of 5.7 years. RESULTS: Our patients had an increased incidence of congenital malformations including microcephaly (<5%) at birth (2 of 8), congenital heart disease (2 of 8), dysmorphic facial features (1 of 8), and thyroglossal duct cyst (1 of 8). Postnatal hydrocephalus (2 of 8) and hip dislocation caused by ligament laxity (1 of 8) were also noted. One patient had profound visual impairment before 6 months of age secondary to cblC retinopathy, and two patients had abnormal retinal pigmentation with normal visual function. All patients presented with poor growth, feeding problems, and/or seizures. No patients had acute acidotic crises before or after treatment. All patients had dramatic reduction of plasma free homocystine and urine methylmalonic acid excretion after initiation of therapy with carnitine, intramuscular (IM) hydroxocobalamin (OHcbl) and, in two cases, oral betaine. Growth was significantly improved in most cases after the initiation of therapy, and microcephaly was resolved in one patient. All patients were developmentally delayed regardless of age of treatment onset, although two patients had relatively mild developmental delay. CONCLUSION: cblC patients may have an increased incidence of congenital malformations suggesting prenatal effects of abnormal cbl metabolism. Treatment with IM OHcbl and carnitine successfully corrects the biochemical abnormalities and improves growth. Developmental delay of variable severity is always present regardless of age at diagnosis or treatment onset. PMID- 11258351 TI - Southwestern Athabaskan (Navajo and Apache) genetic diseases. AB - PURPOSE: Four apparently unique disorders are known among the Southwestern Athabasan Amerindians, i.e., the Navajo and Apache; they are Athabaskan severe combined immunodeficiency, Navajo neuropathy, Navajo poikiloderma, and Athabaskan brainstem dysgenesis. This study reviews background information on Athabaskan groups and clinical descriptions of these recessive disorders. METHODS: The major clinical findings of these four disorders are reviewed. In addition, the findings of epidemiological surveys are included where available. RESULTS: Although the importance of genetic bottlenecks in increasing the frequency of rare, sometimes unique, autosomal recessive disorders is known for a number of populations, similar phenomena among Native Americans seem to be less well known. CONCLUSION: As many more Native Americans move off the Reservation, the awareness of susceptibility to particular genetic diseases needs to be more widely disseminated. PMID- 11258352 TI - Emerging approaches toward the treatment of neurofibromatoses. PMID- 11258353 TI - What's in a name? PMID- 11258354 TI - Duty to re-contact. PMID- 11258355 TI - Paradoxical neutrophil activation and coagulopathy during the recovery phase after endotoxemia. PMID- 11258356 TI - Low-dose prostacyclin reverses endotoxin-induced intestinal vasoconstriction: potential for the prevention of bacterial translocation in early sepsis. PMID- 11258357 TI - Epinephrine and gut lactate production. PMID- 11258359 TI - Primum [non nocere] lavate manus. PMID- 11258358 TI - Cerebral ischemia in humans after traumatic brain injury. PMID- 11258360 TI - Hepatitis C in clinical practice. AB - Hepatitis C virus infection is now one of the most important causes of chronic liver disease. Primary care physicians play an important role in the diagnosis and initial work-up of patients infected with this virus. Understanding which patients may be at risk is the first step. By understanding the correct use of hepatitis C virus diagnostic testing and the risk and benefits of antiviral therapy, providers will be better equipped to screen and counsel their patients. PMID- 11258361 TI - From fact to recommendation: explicit value premises make the conclusion more convincing. AB - In this article we describe one way to use value premises, i.e. conceptions of what is right or wrong, good or bad, just or unjust. They may, in combination with the results from surveys on attitudes, be used to defend normative conclusions, for example changes in professional guidelines. Several ethical principles may be relevant when discussing an end-of-life decision. We use the preferences of those involved in or affected by that decision (the principle of autonomy) and the obligation to maximize benefits and minimize harm (the principle of beneficence) as value premises. To illustrate the ethical, empirical and logical issues relevant to assess normative conclusions, we present a Swedish survey of the attitudes of health care workers and the general public. In this survey, the physicians stated that they do not want the family to be the sole decision-maker, whilst the public did not want the physician to be the sole decision-maker. As a third option we propose joint decision-making regarding end of-life decisions. This normative conclusion may be rationally discussed, not only by questioning scientific aspects of the survey, but also by critically assessing value premises (autonomy and beneficence) and the logic of the argument. PMID- 11258362 TI - Geographical differences in the incidence of acute myocardial infarction in Sweden. Analyses of possible causes using two parallel case-control studies. AB - OBJECTIVES: To analyse differences in myocardial infarction incidence between two Swedish counties and to evaluate the importance of major risk factors for the observed differences. DESIGN: The incidence of first myocardial infarction was studied using information from registers. For a number of risk factors of myocardial infarction, the prevalence as well as the relative risk was estimated from population controls of case-control studies in the two areas. SUBJECTS: Men and women aged 45-64 years in Stockholm and Vasternorrland County 1993-94. MAIN OUTCOME MEASURES: Relative risks (RRs) and impact fractions were used to evaluate the importance of differences in risk factor prevalence for differences in myocardial infarction incidence between the two areas. RESULTS: The incidence of first myocardial infarction was higher in Vasternorrland than in Stockholm amongst both men (RR = 1.23; 95% CI = 1.08-1.40) and women (RR = 1.41; 95% CI = 1.11-1.79). Obesity and increased levels of blood serum lipids were more prevalent in Vasternorrland than in Stockholm amongst men with impact fractions of 6 and 9-11%, respectively. Amongst women, corresponding differences were not seen, but job strain and shift work tended to be more common in the more northern area. Current smoking was more frequent in Stockholm, particularly for women. CONCLUSIONS: The incidence of first myocardial infarction was higher in Vasternorrland than in Stockholm in both genders. A higher prevalence of obesity and elevated blood serum lipids may explain, in part, this excess incidence amongst men, but amongst women the causes of the higher incidence in the more northern area remain largely unclear. PMID- 11258364 TI - Taking lessons from the new philanthropists. PMID- 11258365 TI - Fracas over 5 million dollars Gulf syndrome grant. PMID- 11258366 TI - Plan for medical research base secures future of UK lab. PMID- 11258367 TI - Postdocs call for better pay and conditions. PMID- 11258368 TI - Gene sequencers hope to put the bite on mosquitoes. PMID- 11258369 TI - Biomedical philanthropy, Silicon Valley style. PMID- 11258370 TI - A question of intent: when is a 'schematic' illustration a fraud? PMID- 11258372 TI - Weighing the Universe. PMID- 11258371 TI - Conservation should be a high priority in Singapore. PMID- 11258373 TI - Ageing. Yeast longevity gene goes public. PMID- 11258374 TI - Condensed-matter physics. First glimpse of the orbiton. PMID- 11258375 TI - Evolutionary biology. What's in a baboon's behind? PMID- 11258376 TI - Materials science. Salting the surface. PMID- 11258377 TI - Prions. The shape of a species barrier. PMID- 11258378 TI - Condensed-matter physics. Superconducting plastic. PMID- 11258379 TI - Crackling noise. AB - Crackling noise arises when a system responds to changing external conditions through discrete, impulsive events spanning a broad range of sizes. A wide variety of physical systems exhibiting crackling noise have been studied, from earthquakes on faults to paper crumpling. Because these systems exhibit regular behaviour over a huge range of sizes, their behaviour is likely to be independent of microscopic and macroscopic details, and progress can be made by the use of simple models. The fact that these models and real systems can share the same behaviour on many scales is called universality. We illustrate these ideas by using results for our model of crackling noise in magnets, explaining the use of the renormalization group and scaling collapses, and we highlight some continuing challenges in this still-evolving field. PMID- 11258380 TI - Noise to order. AB - Patterns in natural systems abound, from the stripes on a zebra to ripples in a riverbed. In many of these systems, the appearance of an ordered state is not unexpected as the outcome of an underlying ordered process. Thus crystal growth, honeycomb manufacture and floret evolution generate regular and predictable patterns. Intrinsically noisy and disordered processes such as thermal fluctuations or mechanically randomized scattering generate surprisingly similar patterns. Here we discuss some of the underlying mechanisms believed to be at the heart of these similarities. PMID- 11258381 TI - Supercooled liquids and the glass transition. AB - Glasses are disordered materials that lack the periodicity of crystals but behave mechanically like solids. The most common way of making a glass is by cooling a viscous liquid fast enough to avoid crystallization. Although this route to the vitreous state-supercooling-has been known for millennia, the molecular processes by which liquids acquire amorphous rigidity upon cooling are not fully understood. Here we discuss current theoretical knowledge of the manner in which intermolecular forces give rise to complex behaviour in supercooled liquids and glasses. An intriguing aspect of this behaviour is the apparent connection between dynamics and thermodynamics. The multidimensional potential energy surface as a function of particle coordinates (the energy landscape) offers a convenient viewpoint for the analysis and interpretation of supercooling and glass-formation phenomena. That much of this analysis is at present largely qualitative reflects the fact that precise computations of how viscous liquids sample their landscape have become possible only recently. PMID- 11258382 TI - Exploring complex networks. AB - The study of networks pervades all of science, from neurobiology to statistical physics. The most basic issues are structural: how does one characterize the wiring diagram of a food web or the Internet or the metabolic network of the bacterium Escherichia coli? Are there any unifying principles underlying their topology? From the perspective of nonlinear dynamics, we would also like to understand how an enormous network of interacting dynamical systems-be they neurons, power stations or lasers-will behave collectively, given their individual dynamics and coupling architecture. Researchers are only now beginning to unravel the structure and dynamics of complex networks. PMID- 11258383 TI - Synchronization and rhythmic processes in physiology. AB - Complex bodily rhythms are ubiquitous in living organisms. These rhythms arise from stochastic, nonlinear biological mechanisms interacting with a fluctuating environment. Disease often leads to alterations from normal to pathological rhythm. Fundamental questions concerning the dynamics of these rhythmic processes abound. For example, what is the origin of physiological rhythms? How do the rhythms interact with each other and the external environment? Can we decode the fluctuations in physiological rhythms to better diagnose human disease? And can we develop better methods to control pathological rhythms? Mathematical and physical techniques combined with physiological and medical studies are addressing these questions and are transforming our understanding of the rhythms of life. PMID- 11258387 TI - Philadelphia chromosome-positive acute lymphocytic leukemia. AB - On a molecular and cellular level, Ph+ ALL seems to be a heterogeneous disease. Unfortunately, the unifying theme of Ph positivity is the poor outcome associated with its presence. Further characterization of molecular subtypes of Ph+ ALL may in the future distinguish those few patients with a potentially good outcome from the majority who face inevitable relapse. Also, novel targeted biologic therapy especially in combination with aggressive, early chemotherapy, may soon be able to temper the disease. Most patients who obtain a remission would be best served by transplantation during remission. For those without a donor, following the disease by PCR-based techniques may detect early relapse. For relapsed patients without the option of transplantation, investigative studies are appropriate. PMID- 11258388 TI - Modularity and dissociation in the evolution of gene expression territories in development. AB - Modularity is a salient feature of development and crucial to its evolution. This paper extends modularity to include the concept of gene expression territory, as established for sea urchin embryos. Territories provide a mechanism for partitioning of the cells of a rapidly developing embryo into functional units of a feeding larva. Territories exhibit the characteristics of modules. The paper asks if the embryo and the nonfeeding larva of the direct-developing sea urchin Heliocidaris erythrogramma are organized into gene expression territories, and if its territories correspond to the canonical territories of the pluteus. An analysis of cell lineage and gene expression data for H. erythrogramma shows that skeletogenic cell, coelomic, and vegetal plate gene expression territories are conserved, although they arise from cell lineages distinct from those of the pluteus, and the overall morphology of the larva differs from that of a pluteus. The ectoderm, as in indirect developers, is divided into territories. However, the oral ectodermal territory characteristic of the pluteus is absent in H. erythrogramma. Oral ectoderm is restored in hybrids of H. erythrogramma eggs fertilized by Heliocidaris tuberculata sperm. This indicates that embryonic modules evolve by changes in expression of dominant regulatory genes within territories and that entire modules can be eliminated in evolution of embryos. PMID- 11258389 TI - Adaptation and constraint in the evolution of Drosophila melanogaster wing shape. AB - We have taken advantage of parallel instances of natural selection on body size in Drosophila melanogaster to investigate constraints and adaptation affecting wing shape. Using recently developed techniques for statistical shape analysis, we have examined variation in wing shape in similar body size clines on three continents. Gender-related shape differences were constant among all populations, suggesting that gender differences represent a developmental constraint on wing shape. In contrast, the underlying shape varied significantly between continents and shape change within each cline (i.e., between small and large body size populations) also varied between continents. Therefore, variation at these two levels presumably results from either drift or natural selection. Functional considerations suggest that shape variation between the continents is unlikely to be adaptive. However, cline-related shape change, which we show has a significant allometric component, may be adaptive. The overall range of wing shape variation, across a large range of wing size, is extremely small, and the possibility that wing shape is subject to stabilizing selection (or canalization) is discussed. PMID- 11258390 TI - Editorial: From the horseman's mouth. PMID- 11258391 TI - The rocky road to Mendel's play. PMID- 11258392 TI - The scale independence of evolution. AB - In this paper, I argue that the ultimate causes of morphological, and hence developmental, evolution are scale independent. In other words, micro- and macroevolutionary patterns show fundamental similarities and therefore are most simply explained as being caused by the same kinds of evolutionary forces. I begin by examining the evolution of single lineages and argue that dynamics of adaptive evolution are the same for bacteria in test-tube evolution experiments and fossil lineages. Similarly, I argue that the essential features of adaptive radiations large and small can be attributed to conventional forces such as mutation and diversifying natural selection due to competition. I then address recent claims that the molecular features of metazoan development are the result of clade-level selection for evolvability, and suggest that these features can be more easily explained by conventional individual-level selection for the suppression of deleterious pleiotropic effects. Finally, I ask what must be known if we are to understand the ultimate causes of molecular and developmental diversity. PMID- 11258393 TI - Macroevolution is more than repeated rounds of microevolution. AB - Arguments over macroevolution versus microevolution have waxed and waned through most of the twentieth century. Initially, paleontologists and other evolutionary biologists advanced a variety of non-Darwinian evolutionary processes as explanations for patterns found in the fossil record, emphasizing macroevolution as a source of morphologic novelty. Later, paleontologists, from Simpson to Gould, Stanley, and others, accepted the primacy of natural selection but argued that rapid speciation produced a discontinuity between micro- and macroevolution. This second phase emphasizes the sorting of innovations between species. Other discontinuities appear in the persistence of trends (differential success of species within clades), including species sorting, in the differential success between clades and in the origination and establishment of evolutionary novelties. These discontinuities impose a hierarchical structure to evolution and discredit any smooth extrapolation from allelic substitution to large-scale evolutionary patterns. Recent developments in comparative developmental biology suggest a need to reconsider the possibility that some macroevolutionary discontinuites may be associated with the origination of evolutionary innovation. The attractiveness of macroevolution reflects the exhaustive documentation of large-scale patterns which reveal a richness to evolution unexplained by microevolution. If the goal of evolutionary biology is to understand the history of life, rather than simply document experimental analysis of evolution, studies from paleontology, phylogenetics, developmental biology, and other fields demand the deeper view provided by macroevolution. PMID- 11258394 TI - Characterization of amphioxus AmphiWnt8: insights into the evolution of patterning of the embryonic dorsoventral axis. AB - The full-length sequence and developmental expression of an amphioxus Wnt gene (AmphiWnt8) are described. In amphioxus embryos, the expression patterns of AmphiWnt8 suggest patterning roles in the forebrain, in the hindgut, and in the paraxial mesoderm that gives rise to the muscular somites. Phylogenetic analysis indicates that a single Wnt8 subfamily gene in an ancestral chordate duplicated early in vertebrate evolution into a Wnt8 clade and a Wnt8b clade. Coincident with this gene duplication, the functions of the ancestral AmphiWnt8-like gene appear to have been divided between vertebrate Wnt8b (exclusively neurogenic, especially in the forebrain) and vertebrate Wnt8 (miscellaneous, especially in early somitogenesis). Amphioxus AmphiWnt8 and its vertebrate Wnt8 homologs probably play comparable roles in the early dorsoventral patterning of the embryonic body axis. PMID- 11258395 TI - The A/P axis in echinoderm ontogeny and evolution: evidence from fossils and molecules. AB - Even though echinoderms are members of the Bilateria, the location of their anterior/posterior axis has remained enigmatic. Here we propose a novel solution to the problem employing three lines of evidence: the expression of a posterior class Hox gene in the coeloms of the nascent adult body plan within the larva; the anatomy of certain early fossil echinoderms; and finally the relation between endoskeletal plate morphology and the associated coelomic tissues. All three lines of evidence converge on the same answer, namely that the location of the adult mouth is anterior, and the anterior/posterior axis runs from the mouth through the adult coelomic compartments. This axis then orients the animal such that there is but a single plane of symmetry dividing the animal into left and right halves. We tentatively hypothesize that this plane of symmetry is positioned along the dorsal/ventral axis. These axis identifications lead to the conclusion that the five ambulacra are not primary body axes, but instead are outgrowths from the central anterior/posterior axis. These identifications also shed insight into several other evolutionary mysteries of various echinoderm clades such as the independent evolution of bilateral symmetry in irregular echinoids, but do not elucidate the underlying mechanisms of the adult coelomic architecture. PMID- 11258396 TI - Treatment of hepatocellular carcinoma: a pivotal role for nuclear medicine? PMID- 11258397 TI - Intra-tumoural injection of 90Y microspheres into an animal model of hepatoma. AB - BACKGROUND: Intrahepatic arterial injection of 90Y glass microspheres (90Y microspheres) is a useful therapeutic modality for inoperative liver tumour. Recently, a new concept of interstitial radiotherapy in the treatment of hepatic malignancies has been carried out with even more encouraging results. However, information regarding this technique is still very rare. The purpose of this study was to analyse the kinetics and biodistribution of 90Y microspheres in rats with hepatic tumours following intra-tumoural injection. METHODS: Twenty male Sprague-Dawley rats with hepatoma were killed at 1 h, 24 h, 48 h and 72 h (five rats each time) after intra-tumoural injection of approximately 7.4 MBq of 90Y microspheres. Samples of various organs were obtained and used to calculate the tissue concentrations and radiation doses. RESULTS: Our data showed that the radioactivity in the tumour was very high throughout this study. The lung was the only organ other than the tumour which showed high radioactivity. The concentrations of radioactivity in other organs, such as normal liver, muscle, spleen, bone, testis, and whole blood were quite low throughout the study. CONCLUSION: Direct intra-tumoural injection of 90Y microspheres is extremely attractive as a clinical therapeutic alternative in hepatoma patients. PMID- 11258398 TI - Dynamic 99Tcm-HIDA SPET: non-invasive measuring of intrahepatic bile flow. Description of the method and a study in primary sclerosing cholangitis. AB - In this study dynamic 99Tcm-HIDA single photon emission tomography (SPET) was performed in patients with primary sclerosing cholangitis and normal test subjects. The method offers the possibility of functional analysis of individual liver segments. After injection of 120 MBq of 99Tcm-HIDA, 12 consecutive SPET examinations were performed at 6-min intervals. The segmental borders of liver segments as seen on computed tomography or magnetic resonance examinations were superimposed on the scintigraphic images allowing placement of regions of interest (ROIs) in specific liver segments. Sampling from the same ROIs in consecutive SPET images enabled creation of time-activity curves for individual liver segments. A range of normal values was created by quantitative analysis of normal volunteer studies. Results of the studies in patients correlated well with cholangiographic extent of disease, liver function tests and histological stage. The technique may have particular value in diseases that affect the liver in a nonhomogenous or segmental fashion. Giving an indication of bile clearance from individual liver segments, it can quantify the functional importance of radiologically detected strictures. Percutaneous liver biopsy can be directed to the worst affected parts of the liver, making biopsy more representative. Sequential studies may allow monitoring of disease progression, aiding in selection and timing of therapeutic procedures. PMID- 11258399 TI - The classical stroop interference task as a prefrontal activation probe: a validation study using 99Tcm-ECD brain SPECT. AB - This study investigated the feasibility of brain single photon emission computed tomography (SPECT) functional imaging in a neuropsychological test setting, following a single-day protocol with a split-dose paradigm. The Stroop Color Word Test (SCWT) is an example of a well-documented prefrontal activation task. In a split-dose protocol, ten right-handed healthy volunteers were injected twice with 370 MBq 99Tcm-ethyl cysteinate dimer while performing consecutively both series of card-reading of the SCWT. Images were reconstructed using filtered back projection and normalized to a standard template in Talairach coordinates. Statistical Parametric Mapping (SPM96) was used to determine voxelwise significant changes. A first activation cluster was found in the left medial prefrontal cortex, consisting of the gyrus cinguli anterior and the gyrus frontalis medius and superior. A second activation cluster included the right gyrus frontalis dorsalis and medius. These findings confirm to a large extent the results of previous functional magnetic resonance imaging, positron emission tomography studies of Stroop-like tasks. The choice and validity of various methodological characteristics of the experimental design leading to these results is critically discussed. It is concluded that brain SPECT activation with the Stroop Color Word Test under standard neuropsychological conditions in healthy volunteers, is both technically and practically feasible. PMID- 11258400 TI - SPECT imaging with [123I]-beta-CIT in Parkinsonism: comparison of SPECT images obtained by a single-headed and a three-headed gamma camera. AB - Single photon emission computed tomography (SPECT) imaging of dopamine transporters by using the cocaine derivative [123I]-(1R)-2-beta-carbomethoxy-3 beta-(4-iodophenyl)-tropane ([123I]-beta-CIT) has been shown to be useful in patients with Parkinsonism. The aim of this study was to compare beta-CIT imaging with single-headed (SHS) and three-headed gamma camera systems (THS). In 17 patients with Parkinsonism, SPECT imaging with an SHS and a THS was performed 24 h after injection of 180 MBq of [123I]-beta-CIT. The SPECT studies were evaluated by visual assessment of the caudate nucleus (CN) and the putamen (PT) and the calculation of the striatal/cerebellar (S/C) ratios (with additional comparison to clinical symptoms measured by the Unified Parkinson's Disease Rating Scale (UPDRS)). The S/C ratios measured by the SHS and THS showed highly significant correlation (two-tailed P < 0.01) with Spearman correlation coefficients (SCCs) of 0.864 for the right side, 0.676 for the left side, and 0.761 for both sides. By the SHS, a sufficient visual differentiation between the CN and the PT could not be achieved. A significantly better distinction could be achieved by using the THS (Wilcoxon P<0.05). The S/C ratios of the THS only showed a significant (P < 0.05) SCC of -0.514 comparing to the UPDRS. Pathological alterations in the beta-CIT uptake pattern could be identified by using the SHS, but a significantly better differentiation of CN and the PT was possible by using the THS. The significant correlation of the S/C ratios measured by THS only emphasizes the value of THS in beta-CIT imaging. PMID- 11258401 TI - The optimal imaging time for [99Tcm]TRODAT-1/SPET in normal subjects and patients with Parkinson's disease. AB - Imaging of dopamine transporters (DATs) in the brain using [99Tcm]TRODAT-1 showed excellent pharmacokinetics for estimation of transporter concentrations. It has been reported that there may be differences in the binding kinetics of DAT radiotracers to DATs between normal subjects and patients with Parkinson's disease (PD). The aim of this study was to determine an optimal time point for (99Tcm]TRODAT-1 brain single photon emission tomography (SPET) acquisition that provides stable target to non-target ratios reflecting the DAT concentration in the brain. Serial [99Tcm]TRODAT-1 brain SPET images 2, 3 and 4 h after intravenous injection of [99Tcm]TRODAT-1 (925 MBq) were performed in five healthy subjects and nine PD patients. Regions of interests were drawn, and caudate/occipital (C/O) and putamen/occipital (P/O) specific to non-specific [99Tcm]TRODAT-1 binding ratios were calculated. The C/O and P/O ratios in healthy subjects showed consistent increases with time, but in PD patients, the C/O and P/O ratios of [99Tcm]TRODAT-1 reached a stable level at 3 h post-injection. There was a statistically significant difference (P < 0.001) between PD and normal subjects at 4 h post-injection for both the C/O and the P/O ratios. In conclusion, we recommend the acquisition of [99Tcm]TRODAT-1 SPET images at 4 h post-injection, as at this time point the C/O and P/O ratios can be used to discriminate between PD patients and healthy subjects. PMID- 11258402 TI - Quantitative analysis of 99Tcm-sestamibi myocardial perfusion SPECT using a three dimensional reference heart: a comparison with experienced observers. AB - BACKGROUND: Quantification of myocardial perfusion single photon emission computed tomography (SPECT) may improve scintigraphic analysis. Recently, a fully operator independent technique for the quantification of myocardial perfusion SPECT was described, based on a normal three-dimensional averaged reference heart. The purpose of this study was to compare the automated SPECT quantification technique with experienced observers. METHODS: A total of 43 patients, 36 with one-vessel coronary artery disease (CAD) and seven with a low likelihood of CAD, underwent 99Tcm-sestamibi SPECT (99Tcm-MIBI SPECT). Three experienced observers and a panel (composed of the three observers), blinded to the clinical and angiographic data, analysed the size and severity of perfusion defects and the relation to the distribution areas of the coronary arteries. Inter-observer agreement was calculated by using kappa (kappa) statistics. RESULTS: The inter-observer agreement between the human observers and the automated quantitative analysis, for severity and size of perfusion abnormality, was moderate (kappa range 0.38-0.68), while this was fair between three individual observers (kappa range 0.36-0.87) and good between the individual observers and the panel (kappa range 0.63-0.89). There were no differences between the quantitative analysis and the panel in the allocation of perfusion abnormalities to the affected coronary artery. CONCLUSIONS: The operator independent quantification method showed a moderate agreement with individual observers and a panel analysis for size and severity of perfusion abnormalities. The automatic quantification has a similar ability to assign perfusion abnormalities to the diseased coronary artery as compared to an expert panel. PMID- 11258403 TI - Detection of myocardial viability using rest-redistribution thallium-201 imaging in a stress 99Tcm-tetrofosmin/rest thallium-201 dual-isotope protocol. AB - This study investigated the utility of optional thallium-201 (201Tl) imaging for detecting myocardial viability in the stress 99Tcm-tetrofosmin/rest 201Tl dual isotope protocol. Seventy-nine patients with old myocardial infarction and 25 patients with acute myocardial infarction underwent acquisition of three consecutive 201Tl images (early, intermediate and delayed) using the dual-isotope protocol. A polar map was created and defect scores (extent and severity) were determined by comparison with normal control data. Fluorodeoxyglucose positron emission tomography was also performed in 16 patients with old myocardial infarction. In patients with old infarction, the severity score decreased significantly from the early to the intermediate images, and decreased further on the delayed images. In patients with acute infarction, the score increased from the early to the intermediate images, but not on the delayed images. Regional uptake on the delayed images showed a better correlation with the fluorodeoxyglucose images than that on the early images. Redistribution on the delayed images was exclusively observed in the myocardial segments with less uptake than that estimated by fluorodeoxyglucose. In conclusion, addition of delayed 201Tl imaging to the dual-isotope protocol could improve the sensitivity for detecting myocardial viability. PMID- 11258404 TI - Detection of residual wall motion after sustained myocardial infarction by gated 99Tcm-tetrofosmin SPECT: a comparison with echocardiography. AB - The differentiation of residual viability from necrotic myocardium in patients with a previously sustained myocardial infarction is important in deciding indications for revascularization. Myocardial viability can be assessed by studying perfusion and regional wall motion. With gated single photon emission computed tomography (SPECT), it is possible to augment SPECT perfusion data with ventricular functional data both at a global and regional level. The aim of the study was to analyse the concordance between wall motion score derived by gated SPECT and echocardiography. Furthermore, the agreement between myocardial perfusion and left ventricular wall motion was analysed with both techniques. We studied a homogenous group of 25 consecutive patients with a previous myocardial infarction (MI) using both gated SPECT 99Tcm-tetrofosmin myocardial perfusion imaging and two-dimensional echocardiography. Echocardiography was performed within 2 weeks of the gated SPECT study. Both for gated SPECT and for echocardiography the left ventricle was divided into seven regions per patient. For comparison, the gated SPECT regions were matched to the echocardiographic regions, resulting in a total of 175 regions. Prevalence of abnormal wall motion (akinetic or dyskinetic) was 23% (39/171) for echocardiography and 21% (36/175) for gated SPECT (P = NS). There was a high agreement in wall motion score between echocardiography and gated SPECT of 80% (136/171). The agreement between myocardial perfusion and myocardial wall motion was 82% (143/175) for gated SPECT and 76% (130/171) for echocardiography (P = NS). Nineteen (34%) of the 56 regions with severely diminished or absent myocardial perfusion showed normal or hypokinetic wall motion both by gated SPECT and echocardiography suggesting residual myocardial viability in malperfused regions. Our results suggest that, gated SPECT imaging is a reliable tool for the assessment of regional wall motion in post myocardial infarction patients. Furthermore, in patients with a previous myocardial infarction gated SPECT imaging has the potential to detect preserved wall motion in regions with fixed perfusion defects, which might be indicative of residual myocardial viability. PMID- 11258405 TI - Use of 99Tcm-MIBI scintigraphy in the evaluation of perfusion improvement after myocardial revascularization with the use of the left internal thoracic artery. AB - In spite of successful revascularization, in a significant group of patients myocardial ischaemia is present after surgery. The final effect of surgery depends on preoperative left ventricular function, initial coronary artery status, completeness of revascularization, the use of arterial or venous grafts, and many other factors. The aim of our 99Tcm-MIBI scintigraphy study was to examine the improvement of perfusion in the left anterior descending artery (LAD) vascular territory after revascularization with the use of the left internal thoracic artery (LITA), with respect to the LAD diameter and use of additional venous graft to diagonal artery. The study group consisted of 45 subjects (42 male, three female) aged 34-68 years (mean age 50.9+/-8.3 years) recruited from patients in whom LITA was grafted into LAD. The operation and postoperative period was uneventful in all patients. Two weeks before, and 3-4 months after surgery, dipyridamole-rest sestamibi SPECT were performed. The revascularization significantly improved both stress (deltaPI = 0.77+/-0.66; P < 0.001) and rest (deltaPI = 0.32+/-0.60; P < 0.001) perfusion of the LAD territory. The improvement was slightly better in patients who received two grafts (deltaPI = 1.42+/-0.91) for the LAD territory in comparison to the group revascularized only with LITA (deltaPI = 0.80+/-0.69; P = patients who received an arterial bypass to the LAD artery the perfusion was abnormal in all eight patients after anterior myocardial infarction and in 39% of patients without a history of infarction. The perfusion improvement was the best when the diameter of LAD was > or = 1.5 mm (deltaPI = 0.88+/-0.95). The independent predictors of perfusion improvement were the number of segments with reversible perfusion defect within the revascularized area (beta = 0.84, P < 0.001), the diameter of revascularized artery (beta = 0.17, P = 0.03) and the presence of pathological Q wave at preoperative ECG (beta = -0.20, P = 0.02). We conclude that the degree of perfusion improvement in the LAD territory after revascularization with the use of LITA depends on the diameter of bypassed coronary artery, completeness of revascularization and the reversibility of preoperative perfusion defect. PMID- 11258406 TI - Prognostic value of 201Tl myocardial scintigraphy after coronary artery bypass grafting. AB - BACKGROUND: 201Tl myocardial scintigraphy (201Tl SPECT) is of strong prognostic value in various populations with suspected or known coronary artery disease. However, its value in patients with coronary artery bypass grafting (CABG) is not fully assessed. METHODS: We examined 115 consecutive patients to determine the relation between clinical data/stress 201Tl SPECT performed 5+/-3 years after CABG, and subsequent cardiac events. RESULTS: Thirteen patients (11%) had stress induced angina, 22 (19%) had electrical positivity, and 97 (84%) had abnormal scintigraphy, including 62 (54%) with reversible defects. During follow-up (35+/ 22 months), there were nine cardiac deaths, seven myocardial infarctions, and 20 revascularization procedures. Multivariate Cox analysis identified the delay between CABG and scintigraphy (P<0.01, relative risk (RR) = 1.01), the extent of stress 201Tl defects (P = 0.04, RR = 1.18), and increased stress 201Tl lung uptake (P = 0.03, RR = 3.56) as significant predictors of cardiac deaths/infarctions. Delay between CABG and scintigraphy (P < 0.001, RR = 1.01), the extent of stress 201Tl defects (P = 0.03, RR = 1.15), and that of reversible defects (P = 0.05, RR = 1.13) were the only significant predictors of total events. CONCLUSIONS: Besides the delay between CABG and scintigraphy, the scintigraphic parameters were the only significant and additive predictors of cardiac events in 115 patients with CABG. PMID- 11258407 TI - Scintigraphic imaging of uveal melanoma with 99Tcm-glutathione. AB - A prospective study with a new tumour-seeking agent, 99Tcm-glutathione (GSH), was performed on 17 patients with choroidal melanoma. Planar and SPECT images using 99Tcm-GSH clearly demonstrated melanotic melanoma but failed to show amelonotic melanomas. Following confirmation of our results by concurrent ultrasonography and magnetic resonance imaging or computed tomography, patients were managed by either 125I plaque brachytherapy, diode laser transpupillary thermotherapy or enucleation depending on the site and location. In combination with other diagnostic tests, 99Tcm-GSH scintigraphy may play a role in the detection of uveal melanoma and its possible distant metastases. PMID- 11258408 TI - Different chelators and different peptides together influence the in vitro and mouse in vivo properties of 99Tcm. AB - Relatively few studies comparing different methods of labelling peptides with 99Tcm have been reported. In this investigation, we evaluated the influence of three chelators on the in vitro and in vivo properties of two small, similar peptides (HNE2 and HNE4) labelled with 99Tcm. Both peptides were labelled with hydrazinonicotinamide (HYNIC) (tricine) at pH 5-6 and with diethylenetriaminepentaacetic acid (DTPA) and mercaptoacetyltriglycine (MAG3) at both pH 5-6 and 7-8. All ten preparations were brought to pH 7.2 immediately after labelling. Each preparation labelled well and control labelling showed each label to be attached specifically at chelation sites. Analysis of 37 degrees C human serum incubates showed little evidence of label instability but high protein binding in several cases. The stability of 99Tcm to cysteine challenge for labelled DTPA- and MAG3-peptides was similar but lower than that for the HYNIC-peptides. Reverse phase HPLC of the DTPA-peptides, but not the MAG3 peptides, showed different 99Tcm species depending on labelling pH. The 3 h biodistributions in normal mice were generally independent of labelling pH for both MAG3-peptides but were heavily influenced by labelling pH for both DTPA peptides. While significant differences in biodistribution for the same labelling method were evident between peptides, as expected, far larger differences in the case of both peptides resulted from changing chelators and, in the case of DTPA, changing the labelling method. In summary, the chelators and labelling methods influenced the biodistribution of 99Tcm in a characteristic fashion common to both peptides. Differences in biodistribution due to the different peptides were relatively small and generally lost in the much larger differences due to chelator and labelling method. In conclusion, it may be important to compare chelators and labelling methods before selecting a 99Tcm labelling method for any particular peptide. PMID- 11258409 TI - Surgical correction of vesicoureteral reflux: 5-year follow-up with 99Tcm-DMSA scintigraphy. AB - AIM: To evaluate kidney function before and after surgical correction of vesicoureteral reflux. The long-term effect was measured with quantitative nephro scintigraphy using 99Tcm labelled dimercaptosuccinic acid (99Tcm-DMSA). METHODS: Forty-five children with a history of urinary tract infections due to vesicoureteral reflux (VUR) were studied. VUR grade was determined with contrast voiding cystourethrography. Planar scintigraphy was performed with 99Tcm-DMSA and uptake measured as a percentage of injected dose. Kidney function was evaluated at baseline and 5 years after corrective surgery. RESULTS: Three months after surgery, persistent mild reflux was found in eight of 76 treated renal units. Kidney uptake at 5-year follow-up was unchanged in the majority of children, indicating preservation of renal function found at baseline. The split renal function showed an excellent correlation (r = 0.99) between baseline and follow up studies (regression slope 1.01). Percentage uptake had a regression slope of 0.89 significantly different from unity (P<0.05). Empirical kidney-depth correction techniques were compared. The scintigraphic pattern worsened in six kidneys, indicative of increased scarring in a minority of children. CONCLUSION: Planar nephro-scintigraphy with 99Tcm-DMSA was well tolerated in our paediatric population, and appeared appropriate to evaluate kidney function in time. After surgical correction of VUR, the baseline function was maintained in 94% of kidneys. PMID- 11258410 TI - The utility of SPECT in determining the relationship between radiation dose and salivary gland dysfunction after radiotherapy. AB - Salivary gland scintigraphy (SGS) is used to depict salivary gland dysfunction after radiotherapy (RT). The aim of this study was to investigate the utility of SGS combined with single photon emission computed tomography (SPECT). Twenty-one patients with a carcinoma of head and neck underwent SGS before and 1 month after RT. After injection of 370 MBq 99Tcm-pertechnetate, a biplanar dynamic acquisition (12 x 1 min) was started, followed by a SPECT acquisition during 4 min. Carbachol was then injected and a second dynamic study (16 x 1 min) was performed, again followed by a SPECT acquisition. The salivary excretion fraction (SEF) was calculated both from the geometric mean planar image for each parotid and from the SPECT data for each transverse plane through the parotids. The RT induced changes in the SEF (dSEF) were correlated with the mean radiation dose calculated using tomography-based dosimetry. The mean radiation dose to the parotids was 44 Gy (range 4.4-68.1 Gy). The mean range of the variation in radiation dose to the transverse slices within the parotids of a patient was 24 Gy (range 6.2-51.9 Gy). Considering all transverse planes through the parotids in all patients, a linear correlation was found between the dSEF calculated using SGS-SPECT and the radiation dose (r=0.45, P=0.0001). Thirteen patients had a variation in radiation dose within the parotids of more than 20 Gy. In nine of these a significant intra-individual correlation between radiation dose and the dSEF of the transverse parotid slices was found (r range 0.55-0.97; P value range 0.037-0.0001). In conclusion, SGS-SPECT can be used for monitoring radiation induced parotid gland dysfunction. It offers the unique possibility for the assessment of intra-individual dose-dysfunction curves in patients with large variations in the radiation dose within the parotids. PMID- 11258411 TI - Ictal ECD-SPECT differentiates between temporal and extratemporal epilepsy: confirmation by excellent postoperative seizure control. AB - We investigated whether ictal single photon emission computed tomography (SPECT) with 99Tcm-ethyl cysteinate dimer (ECD) could differentiate between temporal (TE) and extratemporal epilepsy (ETE) in 30 consecutive patients (n = 21 for TE, n = 9 for ETE), all of whom had excellent postoperative seizure control (class I according to Engel's classification). Ictal SPECT showed isolated temporal hyperperfusion in 90% (19 out of 21) of the TE patients and normal perfusion in two patients. All the ETE patients had ictal SPECT findings consistent with extratemporal seizure onset. The sensitivity of ictal ECD-SPECT for correct localization of the seizure onset zone was 80% in all patients, 86% in TE patients and 66% in ETE patients. Although ictal ECD-SPECT has a lower sensitivity in ETE than in TE, it can be used to clearly distinguish between TE and ETE. It provides non-invasive imaging information for use in further diagnostic and treatment strategies in patients with drug-resistant focal epilepsy. PMID- 11258412 TI - Long-term results of radiation synovectomy: a clinical follow-up study. AB - Radiation synovectomy by intra-articular injection of beta-emitting radionuclides is a reliable and easy-to-perform therapy without harmful side effects for the treatment of inflammatory rheumatoid as well as degenerative joint diseases. The indication for radiation synovectomy is based on both clinical symptoms and on proven hyperperfusion, with active synovitis being seen on a pre-therapeutic three-phase bone scan. In this study, the clinical response after 6-18 months, evaluated by a standardized questionnaire, was compared with the reduction of synovitis seen on three-phase bone scintigraphy after treatment of 475 joints in 151 patients. The best clinical results were obtained in cases of true rheumatoid arthritis (73.4%), with less in other kinds of arthritis (48.8%) such as psoriatic or reactive arthritis. Because of the inflamed synovium being the main target tissue, clinical results in osteoarthritis with severe bone destruction are poorer (33.9%). However, synovitis can be markedly reduced (in approximately 70%), regardless of the underlying diagnosis, as shown by post-therapeutic three phase bone scanning. Radiation synovectomy can be recommended in all kinds of arthritis. It should also be considered in cases of osteoarthritis as a last therapeutic option prior to joint replacement. PMID- 11258413 TI - Correlation of marrow dose estimates based on serial pretreatment radiopharmaceutical imaging and blood data with actual marrow toxicity in anti CD20 yttrium-90 monoclonal antibody radioimmunotherapy of non-Hodgkin's B-cell lymphoma. AB - The purpose of this study was to investigate whether marrow radiation absorbed dose estimates predict haematotoxicity following radioimmunotherapy with an yttrium-90 labelled anti-CD20 monoclonal antibody in non-Hodgkin's B-cell lymphoma (NHL). Radiopharmaceutical data from 12 NHL radioimmunotherapy patients were analysed retrospectively using three methods of marrow radiation absorbed dose estimation based on serial pretreatment indium-111 labelled anti-CD20 monoclonal antibody activity versus time data (0-144 h): (i) lumbar spine (LS) image counts; (ii) blood clearance (BL); and (iii) whole body (WB) activity. Linear regressions were performed between the methods, and between each method and the 0-6 month post-treatment platelet and white blood cell count nadir and absolute drop in count (ADC). For the range of yttrium-90 activities (740-1547 MBq), absorbed dose estimates (mean +/- sigma) were: LS, 142+/-50 cGy (range 62 233 cGy); BL, 89+/-21 cGy (range 63-140 cGy); and WB, 54+/-10 cGy (range 36-63 cGy). The LS and BL marrow estimates differed significantly (P <0.003), with a correlation coefficient r of 0.36 (P = NS), while WB correlated significantly with both LS (r = 0.50, P < 0.05) and BL (r = 0.58, P < 0.05). The range of r with platelet nadir and ADC was -0.20 < or = r < or = 0.01, except for WB with ADC (r = 0.38) (all P = NS). Values of r for white blood cell nadir were unexpectedly positive, being 0.13 for BL and 0.29 for LS (P = NS), and 0.60 for WB (P < 0.025). Values of r for white blood cell ADC were 0.36 for BL and -0.26 for LS (P = NS), and 0.50 for WB (P < 0.05). These results indicate that different commonly employed methods of estimating marrow radiation absorbed dose may yield significantly differing results, which may not correlate with actual radiation toxicity. Therefore, caution must be exercised in relying on these results to predict haematotoxicity. PMID- 11258414 TI - Does simple estimation of 131I-metaiodobenzylguanidine uptake in patients with neural crest tumours correlate with clinical outcome? AB - A retrospective study was undertaken in six patients (three male and three female) with neural crest tumours who received therapeutic doses of 131I-meta iodobenzylguanidine (131I-MIBG) (6.7-10.5 GBq). The age range of the patients was 13-65 years (mean 36 years). Quantification of tumour uptake was obtained from images acquired with a large-field-of-view gamma camera on a single occasion between 2 and 10 days post-treatment. Tumour uptake was calculated to be 0.1% and 3.2% of the administered dose, corresponding to uptakes of 6.7-142.8 MBq. Tumour volume was assessed from computed tomography (CT) or magnetic resonance (MR) images and estimates of tumour dose made from the Medical Internal Radiation Dosimetry scheme (MIRD) tables. Estimated doses were between 7 and 113 Gy. Most significantly, our findings indicate that high tumour uptake did not always correlate with good clinical response. PMID- 11258415 TI - Antigen specific T lymphocyte activation. AB - Following recognition of foreign antigens, the T cell antigen receptor (TCR) transduces signals leading to clonal expansion and differentiation into effector cells. Engagement of the TCR results first in the activation of cytosolic protein tyrosine kinases, followed by initiation of multiple signaling cascades. Recently, it has been shown that regulation and integration of these signaling pathways requires formation of multimolecular complexes and the recruitment of these complexes to specific regions within the cell. This review focuses on the events leading to T cell activation including the signaling cascades and the role of adapter molecules in coordination of these pathways. In addition, the mechanism by which TCR ligation can lead to T cell removal following elimination of an antigenic challenge is discussed. PMID- 11258416 TI - CD28 and T cell co-stimulation. AB - Over the last decade the concept of T cell co-stimulation has emerged to take a central role in the process of T cell activation. However, the exact definition of co-stimulation is still unclear. In this review, we re-examine the concept of co-stimulation. We suggest that while co-stimulation is important, there is little evidence to link co-stimulation with T cell anergy. We then suggest a framework for studying co-stimulation. Focusing on recent advances in our understanding of CD28, we discuss four areas of T cell activation where co stimulation may play a role. PMID- 11258417 TI - Signaling through the B cell antigen receptor in developing B cells. AB - Signaling events arising from the B cell antigen receptor (BCR) complex are critical for the normal progression of a B cell through its stages of maturation. These stages are characterized by the generation and expression of BCR components. Initially, the heavy chain is formed and expressed along with the surrogate light chain and VpreB. This pre-BCR may initiate events that induce the cell to generate the light chain molecules. Proper expression of these molecules triggers the cell to develop into a mature B cell. If a key signal is absent at any stage of B cell development, maturation ceases, and either the defects are corrected or the cell undergoes apoptosis. A deficiency in the expression of several intracellular proteins required for these signaling events leads to the arrest of B cell development. Advancement to specific stages of maturation relies on a particular intensity of signal through pathways leading to the activation of a set of transcription factors. These pathways include the calcium and MAPK family pathways. Factors such as the concentration and avidity of the antigen, the coligation of co-receptors, and the formation of signaling complexes dictate the intensity of these signals and thereby the fate of the B cell at each stage of development. PMID- 11258418 TI - Paired inhibitory and activating receptor signals. AB - The immunological literature has become inundated with reports regarding paired inhibitory receptors. Paired inhibitory receptor systems are highly conserved families that contain receptors involved in either cellular inhibition or activation. In most cases the paired putative biochemical antagonists are co expressed on a given cell and thought to bind similar, if not identical, ligands making their biological role difficult to understand. Examples of these systems include immunoglobulin (Ig)-like receptors (Killer Ig Receptors, Immunoglobulin like Transcripts/Leukocyte Ig-like Receptors/Monocyte Macrophage Ig Receptors, and Paired Ig-like Receptors), and type II lectin-like receptor systems (NKG2 and Ly49). General characteristics of these inhibitory receptors include a cytoplasmic immunoreceptor tyrosine-based inhibitory motif (ITIM). The ITIM is phosphorylated upon engagement and recruits protein tyrosine phosphatases that dephosphorylate cellular substrates that would otherwise mediate activation. In contrast, the activating receptors of these pairs use charged residues within their transmembrane domains to associate with various signal transduction chains including the gamma chain of the receptor for the Fc portion of IgE, DAP12 or DAP10. Once phosphorylated, these chains direct the signal transduction cascade resulting in cellular activation. Here we review the signaling of several paired systems and present the current models for their signal transduction cascades. PMID- 11258419 TI - Jak-STAT signaling pathways in cells of the immune system. AB - Many, if not most, cytokines important for immune responses utilize the Jak-STAT signaling pathway. Jaks are receptor-associated protein tyrosine kinases, and STATs are latent cytoplasmic transcription factors that are activated by tyrosine phosphorylation. STATs play critical, nonredundant roles in mediating cellular transcriptional responses to cytokines, and in cell activation, survival and proliferation. The roles of Jaks and STATs in immune responses have been elucidated by analysis of induction of STAT target genes, and of mice rendered deficient in Jak and STAT genes. Cytokine signaling is modulated by crosstalk between the Jak-STAT pathway and pathways triggered by other major immune receptors, such as antigen receptors and receptors for inflammatory cytokines. Tight regulation of cytokine signaling is required for homeostasis, and several constitutive and inducible mechanisms for downregulation of Jak-STAT signaling have been described. PMID- 11258420 TI - Genetics of primary immunodeficiency diseases. AB - The primary immunodeficiencies are a heterogeneous group of disorders that affect either the development or the function of the immune system. In the last ten years, the genes responsible for many of the most common and best studied immunodeficiencies have been identified. As might be expected, the expression of most of these genes is limited to the hematopoietic system. Although most are members of gene families, their association with disease indicates that they do not perform redundant functions. Some immunodeficiencies involve the effector functions of the immune system, for example the NADPH oxidase system or perforin; however, a striking number of the disorders involve signal transduction pathways. These include defects in ligands, transmembrane receptors, kinases, adaptor proteins and transcription factors. Mutations for each disorder tend to be highly variable and the specific mutation in a gene is only one of the factors that influence the clinical phenotype. Polymorphic variations in susceptibility genes may also contribute to the disease phenotype. Not all genes responsible for immunodeficiency have been identified. As many as 20 to 30% of patients with clinical and laboratory evidence of single gene defects of the immune system do not fit any well described clinical disorder. PMID- 11258421 TI - Molecular biology of the Wiskott-Aldrich syndrome. AB - The Wiskott-Aldrich syndrome (WAS) is an X-linked primary immunodeficiency associated with thrombocytopenia, bloody diarrhea, eczema, recurrent infections, and a high incidence of malignancies. X-linked thrombocytopenia (XLT) is a milder form with predominant platelet abnormalities. Both are caused by mutations of the cytoplasmic WAS protein (WASP). To date, mutations of WASP have been identified in over 340 families and consist of missense and nonsense mutations, deletions and insertions, and splice site mutations. There is a striking correlation between phenotype and genotype. The complex gene product of WASP has multiple functional domains that contribute to actin polymerization, cell motility, intracellular signaling, and apoptosis. Understanding the molecular basis of WAS/XLT not only explains the highly variable clinical phenotype, but also affects the medical management of this serious congenital disorder. PMID- 11258422 TI - X-linked lymphoproliferative disease: genetics and biochemistry. AB - Primary immunodeficiencies comprise a broad group of disorders due to germline mutations in genes regulating lymphocyte development and function. One of these genes, DSHP (also known as SH2D1A, SAP), is mutated in X-linked lymphoproliferative syndrome (XLP), an inherited immunodeficiency characterized by increased susceptibility to primary Epstein-Barr virus (EBV) infection, hypogammaglobulinenia, and lymphoma. Expressed primarily in T and NK cells, DSHP consists of a single SH2 domain and short carboxyl-terminal tail. The presence of a single SH2 domain, without other functional motifs, suggests that DSHP may be a physiologic competitor of other SH2 domain-containing proteins whose binding to phosphotyrosine controls lymphocyte activation and/or function. DSHP binds to the cytoplasmic domains of CDw150 (Signaling Lymphocyte Activation Molecule, SLAM) and 2B4, and may regulate signals transmitted by these receptors in T and NK cells, respectively. Unlike other SH2 domain-containing proteins, DSHP associates with both phosphorylated and non-phosphorylated tyrosine residues, and crystallography studies have revealed novel properties of the DSHP SH2 domain. Future studies exploring the function of DSHP during lymphocyte proliferation and activation should improve our ability to diagnose and treat XLP and possibly other human diseases associated with EBV. PMID- 11258423 TI - Molecular genetics of the Bare lymphocyte syndrome. AB - Major Histocompatibility Complex class II (MHC-II) molecules play a pivotal role in the adaptive immune system because they direct the development, activation and homeostasis of CD4+ T helper cells. Hereditary defects leading to the absence of MHC-II expression result in a severe autosomal recessive immunodeficiency disease called the Bare Lymphocyte Syndrome (BLS), also referred to as MHC-II deficiency. The genetic lesions responsible for BLS do not lie within the MHC-II locus itself, but reside instead in genes encoding transcription factors controlling MHC-II expression. Mutations in four different MHC-II regulatory genes are known to lead to BLS. These genes encode CIITA, RFXANK, RFX5 and RFXAP. CIITA (Class II Transactivator) is a transcriptional coactivator that functions as a master control factor dictating the cell type specificity, induction and level of MHC-II expression. RFXANK, RFX5 and RFXAP are the three subunits of RFX (regulatory factor X), a DNA-binding complex that binds to a conserved cis-acting sequence, the X box, present in the promoters of all MHC-II genes. Elucidation of the molecular defects underlying BLS has led to major advances in our understanding of the mechanisms regulating expression of MHC-II genes. PMID- 11258424 TI - Growth regulation of activated lymphocytes: defects in homeostasis lead to autoimmunity and/or lymphoma. AB - Homeostasis within the immune system is complicated by the need to selectively force the survival of potentially useful lymphocytes in the central lymphoid organs and of antigen-reactive cells in the periphery. Coupled with this requirement, is the need to delete strongly autoreactive cells in the thymus and bone marrow and downsize the foreign antigen-reactive cells following elimination of the pathogen. Homeostasis is achieved by coupling the fate of the cell to the integration of signals received through the antigen receptor, co-stimulatory receptors and cytokine receptors as well as members of the tumor necrosis factor receptor family that are highly specialized to promote survival or death of a cell. In this review, we briefly discuss how well-defined pathways that promote cell survival PI-3 kinase, Akt, Bcl-2 family and inhibitors of apoptosis (IAPs) function within the cell. We discuss how cell death stimuli signal either the intrinsic, mitochondrial pathway of apoptosis or kill the cell through one of the six death receptors such as Fas (APO-1/CD95). Finally, the consequences of spontaneous and genetically engineered mutations within survival and death pathways are discussed in the context of predisposition to autoimmune disease and cancer. PMID- 11258425 TI - Searching for a balance when applying immunosuppression after liver transplantation. PMID- 11258426 TI - Defective fasL reveals immunregulation after costimulation blockade. PMID- 11258427 TI - Extension of ischemic tolerance of porcine livers by cold preservation including postconditioning with gaseous oxygen. AB - BACKGROUND: Transplantation of organs from nonheartbeating donors was recommended to reduce organ shortage. In vitro experiments with rat livers have shown that the warm ischemic tolerance of the liver may be extended by persufflation with gaseous oxygen during cold storage. The qualification of this method for procurement of livers harvested after cardiac arrest was tested in an in vivo approach with pigs. METHODS: Livers from 15 donor pigs were explanted, heparinized, flushed with and stored in University of Wisconsin solution for 4 hr at 4 degrees C, and then implanted into 15 recipients. The organs were dissected immediately after cardiac arrest (group 1) or after 60 min of warm ischemia (groups 2 and 3). Group 2 livers received 75,000 IU of superoxide dismutase together with the flush solution and were persufflated with gaseous oxygen via the venous vascular system during cold storage. Main end point was survival after 5 days. Additionally, metabolic, functional and inflammatory criteria were measured in the blood. RESULTS: All animals of the groups 1 and 2 survived, all animals of group 3 died within 3 hr after reperfusion. In all groups the blood parameters reflected significant damage of the livers. However, the ischemic damage was comparable in the groups 1 and 2 whereas the livers of group 3 exhibited significantly higher levels of aspartate alanine aminotransferase and lactate dehydrogenase, and a significantly elongated partial thromboplastin time 1 hr after reperfusion (P=0.016). CONCLUSIONS: Venous systemic oxygen persufflation in combination with antioxidative medication is a promising new method of resuscitating ischemically altered livers from nonheartbeating donors for successful transplantation. PMID- 11258428 TI - Fulminant hepatitis by Fas-ligand expression in MRL-lpr/lpr mice grafted with Fas positive livers and wild-type mice with Fas-mutant livers. AB - BACKGROUND: Fulminant hepatitis in mice could be induced by gene-transfection of Fas ligand (FasL). However, the mechanisms of this event still remain controversial as to whether it is mediated by direct Fas/FasL interaction and/or neutrophil migration. To investigate the role of exogenous FasL-expression, we established a simple but clear mouse model on which we performed liver transplantation between Fas-mutant mice (MRL-lpr/lpr) and wild-type mice (MRL+/+). METHODS: The controls were nontransplanted wild-type (group 1) and MRL lpr/lpr (group 2) mice. We obtained recipients with a Fas defect only in the liver (group 3; MRL-lpr/lpr liver graft in wild-type mice) and Fas-defected recipients with Fas-positive livers (group 4; wild-type graft in MRL-lpr/lpr). We successfully expressed FasL in the liver by cotransfection of two types of adenoviral vectors, AxCALNFasL and AxCANCre, with a Cre-loxP switching system. RESULTS: FasL-expression in the livers in groups 3 and 4 resulted in animal death due to fulminant hepatitis within 48 hr after administration of the vectors. We obtained similar findings in group 1, whereas the mice in group 2 survived without any evidence of hepatitis. Immune staining revealed a marked infiltration of CD11b-positive cells in group 1 and group 3. Despite the number of apoptotic cells, a few infiltration of CD11b-positive cells were seen in group 4. We observed no remarkable findings in the FasL-expressed livers in group 2. CONCLUSION: The results indicated that exogenous FasL-expression induces hepatocyte apoptosis both by direct interaction with Fas and by recruiting Fas positive inflammatory cells. These findings are important for generating a new strategy to prevent hepatitis as well as for understanding the role of the Fas/FasL interaction in the pathophysiology of hepatitis. PMID- 11258429 TI - A novel management strategy of steroid-free immunosuppression after liver transplantation: efficacy and safety of tacrolimus and mycophenolate mofetil. AB - BACKGROUND: Corticosteroids have been used traditionally for immunosuppression after solid organ transplantation. The variety of modern immunosuppressive agents offers the chance to replace drugs with an unfavorable risk-benefit ratio. The objective of this prospective pilot study was to investigate a novel steroid-free immunosuppressive regimen after clinical liver transplantation. METHODS: 30 adult liver graft recipients were included in an intent-to-treat analysis. Dual induction immunosuppression consisted of tacrolimus and mycophenolate mofetil. Prophylactic steroids were not given. Efficacy and safety parameters analyzed were patient and graft survival, incidence and severity of rejection, and adverse events in correlation to immunosuppressive drug levels. RESULTS: Patient and graft survival at 2 years was 86.7 and 83.9%, respectively. Acute rejection occurred in 26.2%, and was associated with subtherapeutic tacrolimus blood levels and diarrhea. All rejections were completely reversible by temporary addition of steroids. Acute renal failure was seen in 10/30 patients, and was related to high tacrolimus blood levels together with primary liver graft dysfunction. 43% of all patients never received any steroids, and 73% were on a steroid-free maintenance regimen. CONCLUSIONS: These results confirm that corticosteroids can be completely avoided from the beginning after liver transplantation. Double drug immunosuppression with tacrolimus and mycophenolate mofetil is effective and safe in terms of patient and graft survival as well as incidence and severity of rejection. In order to avoid under- or over-immunosuppression, which may be caused by impaired absorption or metabolism, close drug monitoring is advised. PMID- 11258430 TI - Effect of histological damage on long-term kidney transplant outcome. AB - BACKGROUND: Chronic renal allograft failure remains a major challenge to overcome. Factors such as donor quality, delayed graft function (DGF), acute rejection, and immunosuppression are known to affect long-term outcome, but their relationship to histological damage to graft outcome is unclear. METHODS: Protocol kidney biopsies (n=112) obtained at 3 months after transplantation yielded 102 with adequate tissue. Histology was scored by the Banff schema, and compared with implantation biopsies (n=91), repeat 12-month histology (n=39), decline in serum creatinine and serial isotopic glomerular filtration rate, onset of chronic allograft nephropathy (CAN), and actuarial graft survival censored for death with a functioning graft. RESULTS: At a median follow-up of 9.3 years, 20 patients had graft failure and 26 died with a functioning graft. Banff chronic nephropathy was present in 24% of 3-month biopsies, and was predicted by microvascular disease in the donor, cold ischemia, DGF, and acute vascular rejection (P<0.001). Acute glomerulitis at 3 months correlated with segmental glomerulosclerosis at 12 months, subsequent recurrent glomerulonephritis, and graft failure (P<0.01). Subclinical rejection at 3 months occurred in 29% of biopsies, correlated with prior acute rejection and HLA mismatch, and led to chronic histological damage by 12 months (r=0.25-0.67, P<0.05-0.001). Subclinical rejection, arteriolar hyalinosis, and tubulitis present at 3 months had resolved by 12 months. The 10-year survival rates for Banff chronic nephropathy were 90.4% for grade 0, 81.0% grade 1, and 57.9% for grades 2 or greater (P<0.01). Early tubulointerstitial damage at 3 months profoundly influenced graft survival beyond 10 years. CAN was predicted by kidney ischemia, 3-month chronic intimal vascular thickening, tubular injury, proteinuria, and late rejection. Chronic fibrointimal thickening of the small arteries and chronic interstitial fibrosis at 3 months independently predicted graft loss and decline in renal function (P<0.05-0.001). CONCLUSIONS: Early transplant damage occurs in the tubulointerstitial compartment from preexisting donor kidney injury and discrete events such as vascular rejection and DGF. Subsequent chronic damage and graft failure reflect accumulated previous injury and chronic interstitial fibrosis, vascular impairment, subclinical rejection, and injury from late rejection. CAN may be conceptualized as the sequelae of incremental and cumulative damage to the transplanted kidney. The duration of graft survival is dependent and predicted by the quality of the transplanted donor kidney combined with the intensity, frequency, and irreversibility of these damaging insults. PMID- 11258431 TI - Energy and substrate metabolism in patients with chronic extensive graft-versus host disease. AB - BACKGROUND: Allogeneic stem cell transplantation is frequently complicated by graft-versus-host disease (GVHD). Weight loss is one of the characteristic features of GVHD. The etiology of weight loss in GVHD is not completely understood. METHODS: We measured resting energy expenditure (REE) and substrate oxidation rates by indirect calorimetry in patients with stable chronic extensive GVHD under immunosuppressive therapy (n=13) and sex-, age-, height-, and weight matched healthy controls (n=13) in order to evaluate metabolic changes in these patients. Measurements were done on day 518+/-261 after allogeneic stem cell transplantation in the postabsorptive state. Serum concentrations of glucagon, norepinephrine, tumor necrosis factor-alpha, interleukin-6, and free fatty acids were determined. RESULTS: Patients showed a maximum weight loss of 22% during their course of GVHD; nevertheless, they regained 15% of total body weight (TBW) during successful treatment of GVHD. Indirect calorimetry showed an increase in REE per kilogram of TBW (patients, 21.8+/-3.1 kcal/kg TBW/day; controls, 19.9+/-2 kcal/kg TBW/day; P<0.05). Respiratory quotient (patients, 0.79+/-0.04, controls, 0.86+/-0.04; P<0.005) and non-protein respiratory quotient (0.78+/-0.05 and 0.87+/-0.05, respectively; P<0.005) were decreased in patients. GVHD patients had elevated serum glucagon and norepinephrine concentrations, whereas tumor necrosis factor-alpha and interleukin-6 were in the normal range. CONCLUSIONS: Patients with chronic extensive GVHD show an increase in REE and alterations in fat and carbohydrate oxidation rates. These changes seem to be the result of increased action of glucagon and norepinephrine. PMID- 11258432 TI - Endogenous nitric oxide and exogenous nitric oxide supplementation in hepatic ischemia-reperfusion injury in the rat. AB - BACKGROUND: Although nitric oxide (NO) is thought to be beneficial in hepatic ischemia-reperfusion (I/R), the mechanisms for this effect are not well established. METHODS: To investigate the effects of endogenous NO and exogenous NO supplementation on hepatic I/R injury and their pathogenic mechanisms, serum ALT and hyaluronic acid (endothelial cell damage), and hepatic malondialdehyde and H2O2 (oxidative stress), myeloperoxidase activity (leukocyte accumulation), and endothelin (vasoconstrictor peptide opposite to NO) were determined at different reperfusion periods in untreated rats and rats receiving L-NAME, L NAME+L-arginine, and spermine NONOate (exogenous NO donor). RESULTS: After reperfusion every parameter increased in untreated animals. Endogenous NO synthesis inhibition by L-NAME increased hepatocyte and endothelial damage as compared to untreated rats, which was reverted and even improved by the addition of L-arginine. Spermine NONOate also improved this damage. However, different mechanisms account for the beneficial effect of endogenous and exogenous NO. Oxidative stress decreased by both L-NAME and L-NAME+L-arginine, but remained unmodified by spermine NONOate. Myeloperoxidase increased by L-NAME and this effect was reverted by the addition of L-arginine, whereas no change was observed with spermine NONOate. Endothelin levels were not modified by L-NAME and L-NAME+L arginine, but decreased with spermine NONOate. CONCLUSIONS: These results suggest that, although both endogenous and exogenous NO exert a protective role in experimental hepatic I/R injury, the mechanisms of the beneficial effect of the two sources of NO are different. PMID- 11258433 TI - FasL is important in costimulation blockade-resistant skin graft rejection. AB - BACKGROUND: Simultaneous blockade of the CD40 and CD28 costimulatory pathways is effective in prolonging allograft survival in murine and primate models. Recent data suggest that intact apoptotic pathways are crucial for the induction of hyporesponsiveness by costimulation blockade. We have studied the impact of fas/fasL signaling, an important T cell apoptotic pathway, on the effects of costimulation blockade. Methods. Wild type, lpr (fas deficient), and gld (fasL deficient), mice were used as donors and recipients in the murine skin graft model. Allograft survival was compared in untreated and costimulation blockade (500 microg anti-CD40L and 500 microg CTLA4-Ig, days 0, 2, 4, 6) treated recipients. In some recipients, CD4+ T cells were depleted using rat anti-murine CD4 (100 microg day -3, -2, -1, and weekly). RESULTS: gld mice treated with costimulation blockade enjoy a significantly greater increase in skin allograft survival than do wild-type mice. This effect is not replicated using lpr donors or recipients. Experiments in which CD4+ cells were depleted demonstrate that fasL is not necessary for CD8-mediated allograft rejection, and that depletion of CD4+ cells eliminates some of the survival advantage induced by costimulation blockade. CONCLUSIONS: FasL is not required for the establishment of costimulation blockade induced hyporesponsiveness, but rather appears to be required for normal costimulation blockade resistant rejection. Fas expression is not critical for costimulation blockade resistant rejection, suggesting that fasL may be interacting with other receptors. Further, it appears that CD4+ cells are important in the maintenance of allograft protection induced by costimulation blockade in this model. PMID- 11258434 TI - Reprogramming of gene expression in cultured cardiomyocytes and in explanted hearts by the myosin ATPase inhibitor butanedione monoxime. AB - BACKGROUND: Butanedione monoxime (BDM) is a reversible myosin ATPase inhibitor. Its use in transplantation medicine may be of benefit in the preservation of hearts. As little is known about its ability to prevent stress and metabolic deregulation, we wanted to investigate the genomic response in cultured cardiomyocytes and explanted, preserved hearts at the transcriptional level. METHODS: We thus investigated the gene expression of the transcription factors GATA-4, Nkx2.5, MEF-2c, and Oct-1 and of the downstream target genes atrial and brain natriuretic peptide, alpha- and beta-myosin heavy chain, alpha-cardiac actin, and alpha-skeletal actin. Additionally, lactate dehydrogenase and creatine kinase enzyme activities were measured as markers for membrane integrity and metabolic deregulation of cardiomyocytes. RESULTS: In untreated cardiomyocyte cultures, expression of GATA-4 and Nkx2.5 was increased 7- and 4-fold, 72 hr after isolation, but the gene expression of MEF-2c and Oct-1 was reduced to 10% and 70%, at day 3 in culture. We show atrial natriuretic peptide and brain natriuretic peptide gene expression to be maximal 24 and 72 hr after isolation, the level being 3- and 2-fold, when compared with freshly isolated cells. The gene expression of alpha- and beta-myosin heavy chain was reduced to approximately 30% at day 3 in culture and similar observations were made for alpha-cardiac and alpha-skeletal actin, which declined to approximately 20% and 10% of control values, 72 hr after isolation. BDM prevented at the transcriptional level enhanced expression of markers for stress and metabolic deregulation, and the activities of lactate dehydrogenase and creatine kinase were highly significantly reduced. Similar results were obtained when explanted hearts were stored in BDM-containing organ preservation solution. CONCLUSIONS: Preservation of metabolic function in donor organs is of critical importance in transplantation medicine, and we show gene markers for stress and metabolic deregulation in cultures of cardiomyocytes and explanted hearts to be significantly reduced by BDM. Reprogramming of gene expression of nuclear transcription factors and downstream target genes may prolong the acceptable storage time between explantation and transplantation. PMID- 11258435 TI - Transferrin receptor-mediated gene transfer to the corneal endothelium. AB - BACKGROUND: The application of gene therapy to prevent allograft rejection requires the development of noninflammatory vectors. We have therefore investigated the use of a nonviral system, transferrin-mediated lipofection, to transfer genes into the cornea with the aim of preventing corneal graft rejection. METHODS: Rabbit and human corneas were cultured ex vivo and transfected with either lipofection alone or in conjunction with transferrin. The efficiency of transfection, localization, and kinetics of marker gene expression were determined. Strategies to increase gene expression, using chloroquine and EDTA, were investigated. In addition to a marker gene, a gene construct encoding viral interleukin 10 (vIL-10) was transfected and its functional effects were examined in vitro. RESULTS: Transferrin, liposome, and DNA were demonstrated to interact with each other, forming a complex. This complex was found to deliver genes selectively to the endothelium of corneas resulting in gene expression. Treatment of corneas with chloroquine and EDTA increased the transfection efficiency eight-fold and threefold, respectively. We also demonstrated that constructs encoding vIL-10 could be delivered to the endothelium. Secreted vIL-10 was shown to be functionally active by inhibition of a mixed lymphocyte reaction. CONCLUSIONS: Our data indicate that transferrin-mediated lipofection is a comparatively efficient nonviral method for delivering genes to the corneal endothelium. Its potential for use in preventing graft rejection is shown by the ability of this system to induce vIL-10 expression at secreted levels high enough to be functional. PMID- 11258437 TI - Changes in the composition of serum bile acid as a predictor of small bowel allograft rejection in rats. AB - Composition of bile acid was studied as a noninvasive rejection marker after small bowel transplantation (SBT). Orthotopic SBT were performed in rats, and they were divided into four groups: group 1, sham-operated rats; group 2, recipients with isografts; group 3, recipients with allografts treated with FK506; and group 4, recipients with untreated allografts. On postoperative days (POD) 5 and 7, the graft histology, intraluminal bacterial overgrowth, individual bile acids concentration of the recipient serum and bile were examined. On POD 5, early histological findings of acute rejection were observed in group 4, and the ratio of secondary bile acids of this group were significantly higher than the other groups. The bile acid changes were enhanced by bacterial overgrowth on POD 7. The ratio of secondary bile acids changed in relation to acute rejection after SBT, suggesting that they can be useful for early diagnosis of acute SBT allograft rejection. PMID- 11258436 TI - Induction of hyporesponsiveness to fully allogeneic cardiac grafts by intratracheal delivery of alloantigen. AB - BACKGROUND: Soluble protein delivered through the mucosal surface can induce immunological unresponsiveness. The purpose of this study was to determine if prior exposure to alloantigen via the trachea could modulate the immune response to subsequent cardiac allografts. METHODS: Hearts from C57BL/10(H2b) mice were transplanted into CBA(H2k) recipients. Recipient mice were given donor 1x10(7) splenocytes into the trachea with or without antibody specific for mouse CD80 (1G10) and/or CD86 (GL1) (100 microg each) 7 days before transplantation. RESULTS: All grafts survived in recipients treated with intratracheal delivery of alloantigen for over 35 days (mean survival time [MST], 56 days), whereas naive control mice and mice treated with syngeneic antigen rejected grafts acutely (MST, 8 and 7 days, respectively). Interestingly, when 1G10, GL1, or both of them were combined with the protocol, the majority of grafts were rejected within 21 days after grafting (MST, 7, 15, and 17 days, respectively). CONCLUSION: Intratracheal delivery of alloantigen induced significantly prolonged survival of fully mismatched cardiac allografts and the effect was abrogated by the blockade CD80 and/or CD86 pathway. PMID- 11258438 TI - Progressive functional adaptation of segmental bowel graft from living related donor. AB - We report a patient with short gut syndrome successfully treated with living related bowel transplantation. A 27-year-old Caucasian man was referred after traumatic loss of almost the entire bowel from the third portion of duodenum to the sigmoid colon. His HLA-identical sister volunteered as a donor. A 200-cm segment of ileum was successfully transplanted under tacrolimus-based immunosuppression. The posttransplant course was uneventful, without rejection or infectious complication. Total parenteral nutrition was discontinued 1 week posttransplant. At 6 months the patient had returned to his preinjury weight. Water and D-xylose absorption as well as fecal fat studies were markedly abnormal 1 month posttransplant but normalized by 6 months. The donor recovery was uneventful. A well-matched segmental ileal graft from living donor can provide complete rehabilitation for patients with short gut syndrome. We documented a progressive functional adaptation of the ileal graft, resulting in normal absorption by 5 months posttransplantation. PMID- 11258439 TI - Role of Doppler echocardiography in the assessment of portopulmonary hypertension in liver transplantation candidates. AB - BACKGROUND: Portopulmonary hypertension is a severe complication of liver cirrhosis that carries a high risk for posttransplantation mortality. We aimed at evaluating the utility of Doppler echocardiography in screening for portopulmonary hypertension in liver transplantation candidates. METHODS: One hundred seven cirrhotic patients candidates for liver transplantation were studied by Doppler echocardiography and subsequently, by cardiac catheterization at transplantation. Two parameters were estimated by Doppler: systolic pulmonary arterial pressure (SPAP) derived from tricuspid regurgitation and the pulmonary acceleration time. Portpulmonary hypertension was suspected when SPAP was > or = 40 mm Hg and/or pulmonary acceleration time < 100 ms. RESULTS: Portpulmonary hypertension was suspected by Doppler study in 17 patients (15%). However, portopulmonary hypertension (mean pulmonary arterial pressure > or = 25 mm Hg and pulmonary vascular resistance > 120 dynes.s/cm5) was confirmed by the hemodynamic study in five patients (4.7%). Sensitivity and specificity of Doppler echocardiography for detecting portopulmonary hypertension was 100 and 88%, respectively, with a positive predictive value of 30%. The diagnostic accuracy of pulmonary acceleration time alone (96%) was better than pulmonary arterial pressure alone (90%). CONCLUSIONS: Doppler echocardiography, and especially the determination of pulmonary acceleration time, is a useful screening method for portopulmonary hypertension in patients with liver cirrhosis who are candidates for liver transplantation. PMID- 11258440 TI - Improved phagocyte response by co-amoxiclav in renal transplant recipients. AB - BACKGROUND: Infectious diseases are a major source of morbidity and mortality for immunosuppressed transplant recipients and the antimicrobial chemotherapy can be often less effective in these individuals, because the contribution of underlying host defenses is absent. METHODS: The influence of co-amoxiclav on the functions of polymorphonuclear granulocytes (PMNs) from renal transplant recipients were investigated. RESULTS: PMNs from renal transplant recipients showed a diminished phagocytic activity with reduced phagocytosis and bactericidal activity against intracellular Klebsiella pneumoniae, compared to that seen with PMNs from healthy subjects. Co-amoxiclav significantly elicited the functions of PMNs from uremic patients, resulting in an increased percentage of ingested klebsiellae and in a higher bactericidal effect (98-99%), compared with the drug-free control system. When PMNs were collected from renal transplant recipients treated with co amoxiclav a significant high increase in both phagocytosis and killing activity were detected, showing the co-amoxiclav capability of "restoring" even in vivo the depressed primary functions of PMNs. CONCLUSIONS: The interesting beneficial properties of co-amoxiclav, which result in restoring the phagocyte-dependent response in renal transplant patients both in vitro and in vivo, may make this drug more suitable for the treatment of infections in patients with defects of phagocyte functions. PMID- 11258442 TI - Toward the first legally binding European text on the ethics of transplantation. PMID- 11258441 TI - Prevalence of donor-specific anti-HLA antibodies during episodes of renal allograft rejection. AB - BACKGROUND: Recent studies suggest that the appearance of anti-HLA antibodies in the early posttransplant period is associated with an increased incidence of acute and chronic rejection months later. However, very little is known about the prevalence of anti-HLA antibodies at the time that the rejection episodes are diagnosed. The purpose of this study was to analyze retrospectively 420 sera from 263 renal allograft recipients who were readmitted to the hospital for any reason between 1989 and 1998 in order to determine if a correlation existed between the presence of donor-specific anti-HLA antibodies and graft rejection. METHODS: Sera were assayed for IgG HLA class I and II antibodies by ELISA. The ELISA results were analyzed using contingency tables with Fisher's exact test and compared with mismatched antigens in the donor. RESULTS: Antibodies to donor HLA class I molecules in the posttransplant sera were extremely rare, occurring in only 6 of the 420 sera (1.4%) analyzed. Antibodies to donor class II antigens were slightly more common, occurring in 25 of the 420 sera (6%). In 21 of these 25 cases (84%), the presence of donor-specific HLA class II antibodies was associated with episodes of either acute (n=14) or chronic rejection (n=7). Five patients had antibodies to both class I and class II donor antigens, and all five of them lost their grafts to rejection. CONCLUSION: Although the presence of donor-specific HLA antibodies presented a significant risk for acute or chronic rejection, 77% of all acute and chronic rejections occurred in patients without detectable HLA antibodies. PMID- 11258443 TI - A limitation of fluorescence in situ hybridization (FISH) assays targeting sex chromosomes for monitoring of early relapse after transplantation. PMID- 11258444 TI - Hyperacute xenograft rejection is not consistent after pig to primate solid organ transplantation. PMID- 11258445 TI - Transforming growth factor-beta1 plasma levels in stable renal allograft recipients under different immunosuppression. PMID- 11258446 TI - Molecular cloning of horse Hsp90 cDNA and its comparative analysis with other vertebrate Hsp90 sequences. AB - Heat shock protein 90 (Hsp90), a molecular chaperone, is ubiquitous and involved in numerous cellular processes. To contribute to the relatively small collection of vertebrate Hsp90 sequences in the gene data bank, we cloned and sequenced horse (Equus caballus) Hsp90 alpha and beta cDNAs. This enabled identification of horse-specific primers for development of a convenient PCR-based method that could monitor horse stress tolerance. We analyzed the sequence data comparatively and phylogenetically with other Hsp90 cDNA sequences, and identified vertebrate specific and isoform-specific conserved regions to facilitate future molecular investigations of Hsp90 functions. We found 4 highly conserved regions to vertebrate Hsp90 exclusively and 27 amino acids conserved among but differing between Hsp90 alpha and Hsp90 beta sequences. Protein-based phylogenetic trees revealed high conservation between mammal species within Hsp90 alpha and beta clusters. Comparison of nucleotide and amino acid substitution levels suggests that horse Hsp90 beta has undergone strong purifying selection, while rat Hsp90 beta and hamster Hsp90 alpha have been positively selected. Surprisingly, fish Hsp90 alpha genes clearly clustered with Hsp90 beta genes, and no distinct placement of fish Hsp90 alpha protein was found. The Hsp90 alpha isoform is apparently the result of beta gene duplication. Our results highlight the importance of organism- and isoform-specific Hsp90 functional analyses in describing the role of Hsp90 in cells. PMID- 11258447 TI - Changes in quantitative profile of extracellular matrix components in the kidneys of rats with adriamycin-induced nephropathy. AB - Extracellular matrix components (ECMs) in histological sections of the kidney cortex from the rats with adriamycin (ADR)-induced nephropathy (5 mg/kg, i.v.) were quantified by an immunohistochemical micromethod. Changes in kidney histopathology and urine and blood biochemistry were investigated. Enlarged kidneys were granular on the surface and pale in color in ADR-treated rats, and these rats had kidneys with glomeruli with expanded mesangial area and with capillary aneurysm. Severe albuminuria, hypoalbuminemia, hypercholesterolemia and disorders in other nephrotic parameters were observed in ADR-treated rats. Type I and IV collagens, fibronectin and laminin contents in the renal cortex of ADR treated rats at 10 weeks were 329, 317, 263 and 295%, respectively, higher than in each vehicle control, and those at 28 weeks were 1,211, 930, 1,057 and 1,012%, respectively. The glomerular sclerotic abnormalities progressed in a time dependent manner. Moreover, there was a strong correlation between the ECM levels and serum creatinine and blood urea nitrogen levels. In conclusion, microquantification provided useful information for accurate diagnosis and prognosis of nephrotic lesions and is a good tool to assess the advancement of renal disorders in patients with nephropathy. PMID- 11258448 TI - Negative inotropic effect of diazepam in isolated guinea pig heart. AB - The inotropic effect of diazepam, a benzodiazepine derivative, and its mechanism of action were examined using guinea pig heart and single ventricular cell preparations. In Langendorff hearts and right ventricular free-wall preparations, diazepam (10 to 100 microM) produced a monophasic negative inotropic effect in a concentration dependent manner. Neither a central type (flumazenil 1 microM) nor a peripheral type (PK11195 10 microM) of benzodiazepine receptor antagonist antagonized the monophasic negative inotropic effects of diazepam. Diazepam (10 to 100 microM) shortened action potential duration of papillary muscle in a concentration dependent manner. In isolated single ventricular cells, diazepam (30 and 100 microM) inhibited the calcium current (I(Ca)) in a concentration dependent manner. Diazepam produced a significant decrease in I(Ca) elicited by first depolarizing pulses, however, the decrease of I(Ca) was not augmented during a train of depolarizing pulses. Thus, diazepam appears to produce a tonic block of cardiac calcium channels and the mode of inhibition is clearly different from the use-dependent block of verapamil. From these results, it was concluded that diazepam produces a monophasic negative inotropic effect that is independent of the benzodiazepine receptor, and is probably mediated through an inhibition of I(Ca) in guinea pig heart preparations. PMID- 11258449 TI - A mucinous histochemical study on malignancy of aberrant crypt foci (ACF) in rat colon. AB - The relationship between malignancy and number of crypts (crypt multiplicity) comprising aberrant crypt foci (ACF) was investigated, by studying changes in the mucous nature of ACF with 5 crypts or less, ACF with 6-13 crypts, adenomas and invasive adenocarcinomas induced by 1,2-dimethylhydrazine in distal colon of rats. A paradoxical Con A-staining was performed for goblet cell mucins. Of the sulfomucin-dominant ACF with 1-3 crypts, 82.6% had labile class III mucin, similar to the distal colon in the normal rats. However, in most of the goblet cell mucin produced by the ACF with 4-5 crypts with an indicated relation to colorectal carcinoma or the sialomucin (SiM) -dominant ACF with 1-3 crypts, mucin types other than class I were rarely present. The incidence of class I mucin decreased with the increase in crypt multiplicity of ACF or in the degree of histological malignancy, with the lowest incidence of 40% in adenocarcinomas. In contrast, the incidence of class II mucin increased markedly with the increase in crypt multiplicity of ACF or in the degree of histological malignancy, with the highest incidence in adenocarcinomas (95%). The ACF with 6-13 crypts had a mucous profile similar to that of adenomas. These results suggested that malignancy of ACF related to the crypt multiplicity. In the ACF with 1-3 crypts, SiM-dominant ACF had the potential to progress to malignant lesions. PMID- 11258450 TI - Seasonal changes of testicular weight, sperm production, serum testosterone, and in vitro testosterone release in Korean ring-necked pheasants (Phasianus colchicus karpowi). AB - To study the biology of reproduction of male Korean ring-necked pheasants kept under natural conditions of temperature and photoperiod, testicular weight, serum testosterone concentrations, testosterone release from the luteinizing hormone (LH)-stimulated testis in vitro and sperm production were measured. Significant changes associated with seasonal cycles were found. Testis weight decreased dramatically in August, remained low until February, rapidly increased from March to high levels to June, and decreased subsequently. Serum testosterone concentrations remained little from August until February, but increased sharply in March to reach the highest levels in April. Thereafter, the concentrations decreased significantly from June. The testosterone release was low from August to February, increased abruptly in March to reach the highest levels in May, and showed rapid decrease thereafter. Sperm production decreased to nondetectable levels from August to February, increased markedly in March, reached a peak in May, and sharply decreased thereafter. Thus, the pheasants breed from late March to late June. These results indicate that the Korean ring-necked pheasant, under natural conditions, exhibits characteristics of a seasonal cycle in reproduction. PMID- 11258451 TI - Clinical trial of a feline pheromone analogue for feline urine marking. AB - Thirty-six cases of feline urine marking problem were collected through the cooperation of veterinary practitioners in the Kanto, Chubu, and Kansai areas in Japan, for an assessment of the clinical effect of treatment with a synthetic analogue of a feline cheek gland pheromone-like product. The mean frequency of urine marking was 14.2 times/week (median, 10; range, 1-77) at pre-treatment week (preW), and decreased significantly from the first week of treatment, dropping to 4.2 times/week (median, 2; range, 0-44) at the fourth week of treatment. This effect continued until the fourth week after cessation of treatment. These 36 cases were divided into 3 groups based on the effectiveness of treatment as demonstrated in the fourth week of treatment; 37% was categorized as the totally eliminated group (urine marking was not seen), 40% as the reduced group (the frequency of urine marking was equal to or less than 50% that of the preW), and 23% as the unchanged group (the frequency of urine marking was more than 50% that of the preW). Effectiveness of treatment in these groups was 38%, 24%, and 38% at the fourth week after the cessation of treatment, respectively. The decreasing rate of urine marking was compared between cats with and without intercat aggression, and it was revealed that the frequency of marking was sustained at high level in cats with intercat aggression. These results suggest that this pheromone treatment is as effective in Japan as has been reported in other countries for solving feline urine marking problems. PMID- 11258452 TI - Ultrastructure of aortic elastic fibers in copper-deficient Sika deer (Cervus nippon Temminck). AB - Light microscopic and transmission and scanning electron microscopic observations were performed on the aortas of two 4- and 6-year-old deer affected with cervine ataxia and two 6-month- and 4-year-old healthy deer. Examination of the aortas from affected deer by transmission electron microscopy revealed the absence of distinct elastic laminae in the internal elastic lamina and tunica media, but discontinuous and irregular clumps of elastin were present. Scanning electron microscopy disclosed immature architecture of elastic fibers in the aortas from the copper-deficient deer, and the architecture was similar to that of a 6-month old healthy deer. PMID- 11258453 TI - Serum growth hormone and insulin-like growth factor-1 concentrations in Japanese black cattle with growth retardation. AB - Serum concentrations of growth hormone (GH) and insulin-like growth factor-1 (IGF 1) were determined in 5 calves in the same lineage with growth retardation. They had normal appetites, activities, body proportion, and laboratory test results. Calves with growth retardation had higher serum GH concentrations and lower serum IGF-I concentrations. These findings suggested defects in the GH-IGF-1 axis, such as in the GH-receptor. PMID- 11258454 TI - Detection of antibodies against Pasteurella multocida using immunohistochemical staining in an outbreak of rabbit pasteurellosis. AB - To detect serum antibody against Pasteurella multocida (P. multocida) in infected rabbits. a modified immunoperoxidase assay was applied. An outbreak of P. multocida infection in rabbits started from sudden death. The infected rabbits had severe fibrinous and purulent pneumonia with hemorrhage, and a large number of P. multocida (A:12) was isolated from the trachea and lungs of the animals. Antibodies of IgM and IgG to P. multocida were assessed by immunohistochemical staining using the sera of the animals as primary antibodies and applying them to formalin-fixed, paraffin-embedded sections of P. multocida attached to calf fibrin. IgM antibodies to P. multocida were first detected 7 days after the onset of the disease. IgG antibodies began to rise on the 7th or 14th day. These results suggested that the modified immunoperoxidase assay could detect antibodies against P. multocida. PMID- 11258455 TI - Changes in serum thyroid hormone levels in newborn calves as a diagnostic index of endemic goiter. AB - Maximum serum thyroxine (T4) and triiodothyronine (T3) levels of healthy calves were seen at 1 day after birth, and thereafter rapidly decreased until 5 days after birth. They stabilized until 2 weeks after birth, then gradually decreased until 4 weeks after birth. Serum T4 levels of calves with endemic goiter tended to be lower than those of healthy ones, but showed similar levels to those of adult cows. T3 levels of calves with goiter were similar to those of healthy ones, but showed higher variation. T4/T3 ratio of calves with goiter were significantly lower than those of healthy ones and adult cows. While individual levels of serum T4 and T3 at just after birth could not be considered as a diagnostic index, the T4/T3 ratio could be adopted as a diagnostic index of endemic goiter. PMID- 11258456 TI - Hemodynamic alterations in dogs with shock induced by intravenous injection of heartworm extract. AB - To elucidate one way of the shock mechanisms, the hemodynamic alterations were examined in 7 dogs with heartworm (HW) extract-induced shock. The first alteration observed after injection of HW extract was a decrease in right ventricular end-diastolic pressure (RVEDP). After that, left ventricular (LV) end diastolic pressure, LV systolic pressure, and LV dp/dt fell significantly, followed by a decrease in the cardiac output of all dogs to below the detectable level (1.00 l/min). Since RVEDP depends on blood flow into the right ventricle, the decrease in RVEDP means a reduction in venous return. Therefore, this study showed that the first trigger of a decrease in blood pressure in HW extract induced shock is the reduction in venous return. PMID- 11258457 TI - Pituitary chromophobe carcinoma with a low level of serum gonadotropin and an aspermatogenesis in a dog. AB - A 5-year-old male Shiba dog with progressive neurologic signs was examined by computed tomography (CT). A CT image of the brain disclosed a large, spherical high-density lesion in the thalamus and diencephalon. Serum LH, FSH and testosterone levels were all low. Macroscopically the large mass was connected with the sella turcia, and it was histopathologically diagnosed as a pituitary chromophobe carcinoma. An aspermatogenesis was observed in the testes. Therefore, it was suggested that the low levels of gonadotropin secretion from the pituitary gland due to the pituitary tumor resulted in the failure of maturation of spermatozoa and spermatids. PMID- 11258459 TI - Diffuse bilateral hemangiosarcoma of the uterus in a dog. AB - A 15-year-old female mongrel dog showed abdominal swelling, marked hemorrhagic ascites and vulvar discharge, and ovariohysterectomy was performed. Grossly, the uterus was enlarged bilaterally without apparent mass formation. Histologically, the uterine muscular wall was composed of proliferated sinusoidal vessels. In some areas, irregular and small vessels proliferated markedly, while in others, pleomorphic and atypical tumor cells forming irregular vascular structures were predominant. From these findings, the case was diagnosed as diffuse bilateral hemangiosarcoma of the uterus that invaded to the ovary and broad ligament. The relationship between the tumor and angiomatosis was discussed. PMID- 11258458 TI - Reduction of the infectivity of Toxoplasma gondii and Eimeria stiedai sporozoites by treatment with bovine lactoferricin. AB - Sporozoites of Toxoplasma gondii preincubated with lactoferricin showed decreased activity in penetration of mouse embryonal cells. Mice inoculated with 10(5) sporozoites preincubated with lactoferricin showed a higher survival rate than those inoculated with the same number of untreated sporozoites. Likewise, sporozoites of Eimeria stiedai preincubated with lactoferricin also showed decreased activity in penetration of rabbit hepatobiliary cells. Rabbits inoculated with 10(5) sporozoites preincubated with lactoferricin shed fewer oocysts than those inoculated with the same number of untreated sporozoites. These results indicate that lactoferricin is effective to reduce the infectivity of sporozoites of Toxoplasma gondii and Eimeria stiedai. PMID- 11258460 TI - Comparison of various reproductive status in Sika deer (Cervus nippon) using fecal steroid analysis. AB - The feasibility of fecal steroid analysis for pregnancy diagnosis and sex determination were tested in sika deer (Cervus nippon). Feces were collected from captive sika deer in June (non-breeding season and late-pregnancy period) and October (breeding season), and also from the rectum of 24 female sika deer (19 pregnant and 5 non-pregnant females) shot as part of programs for population control in February and March (mid-pregnancy period). In mid- and late-pregnancy periods, fecal progesterone concentrations were significantly higher in pregnant female than in male and non-pregnant female deer. In October, fecal testosterone concentrations were higher in adult male deer, and no difference was found between young males and females. These results suggest that fecal steroid analysis would be a useful means for estimating pregnancy status and for detecting adult male among wild deer. PMID- 11258461 TI - Giant cell hepatitis in two young cats. AB - A very rare case of the liver lesion characterized by formation of multinucleated giant hepatocytes with inflammatory cell infiltration were observed in two young (1.5 years and 2 years old) cats bearing thymic malignant lymphoma. Histopathological features of this liver lesion were very similar to giant cell hepatitis (GCH) in human neonates and infants. Therefore, the lesion was diagnosed as feline GCH. PMID- 11258462 TI - Isolation of Encephalitozoon cuniculi using primary tissue culture techniques from a rabbit in a colony showing encephalitozoonosis. AB - Encephalitozoon spores were isolated in a primary tissue culture of the kidneys from an encephalitozoonosis-suspected rabbit in a municipal zoo in Hokkaido. The isolated spores were morphologically characteristic of microsporidial ones in chromotrope stain, and proven to be E. cuniculi by a polymerase chain reaction (PCR) with a species-specific primer set and by direct DNA sequencing of the PCR products. PMID- 11258463 TI - Complete nucleotide sequence of the cytochrome b gene of channel catfish Ictalurus punctatus and comparison of sequence homology among channel catfish and other fishes. AB - To clarify the phylogenic relationship of channel catfish with other fishes, the cytochrome b (Cyt b) gene of the catfish was cloned and sequenced. Channel catfish (Ictalurus punctatus) belonging to the family Ictaluridae in the order Siluriformes showed a 78.4-87.4% similarity to all but one fish of the family Cyprinidae and river loach Crossostoma lacustre of the family Balitoridae in the order Cypriniformes in which genes had already been sequenced, and a 97.2% similarity to the goldfish (Carassius auratus) belonging to the family Cyprinidae. Within the family Cyprinidae, a 78.8-89.2% similarity to one another was recorded. In addition, the similarity rate between the family Cyprinidae and the family Balitoridae reached a value of 77.8-79.9% in the order Cypriniformes. Furthermore, in an unrooted phylogenetic tree consisting of four branches among eight fishes, channel catfish and goldfish appeared in the same branch. These results suggested that the Cyt b gene of the channel catfish in the order Siluriformes was closely related to that of a goldfish in the order Cypriniformes. The results were not agreement with the morphological classification. Genetic reclassification of the fishes may be necessary to identify the ancestor. This is the first report on the cloning and complete sequencing the Cyt b gene of the channel catfish which may contribute to the genetic reclassification of catfishes belonging to the order Siluriformes. PMID- 11258464 TI - Molecular cloning of feline hepatocyte growth factor (HGF) cDNA. AB - Hepatocyte growth factor (HGF) is a pleiotropic cytokine responsible for regeneration, development and maintenance of various organs, and growth, invasion and metastasis of tumor cells. A full-length feline HGF cDNA was cloned and sequenced by RT-PCR from cat liver. Feline HGF consists of 728 amino acid and contains alpha- and beta-chains encoded in a single open reading frame. The predicted amino acid sequence of feline HGF showed 93.2, 93.3 and 93.3% homology with those of human, mouse and rat HGF, respectively. The putative proteolytic processing site, all cysteine residues, and four potential glycosylation sites are conserved in all species. Therefore, feline HGF is expected to have a similar three-dimensional structure to human, mouse and rat HGF. PMID- 11258465 TI - Isolation of monoclonal antibodies that inhibit the binding of infectious bursal disease virus to LSCC-BK3 cells. AB - Three hybridoma cell lines producing monoclonal antibodies (MAbs) against LSCC BK3 cells which are susceptible to infectious bursal disease virus (IBDV) infection were produced and characterized. The MAbs, designated T7, Q11 and Q13, inhibited the attachment of IBDV to LSCC-BK3 cells. Furthermore, these MAbs bound to LSCC-BK3 but not to nonpermissive cells in flow cytometry. MAb T7 detected a 110-kDa membrane protein of LSCC-BK3 cells, whereas Q11 and Q13 reacted with membrane proteins of molecular weights 58-, 85-, 90- and 110-kDa. These observations imply that the 110-kDa protein recognized by all the MAbs is associated with IBDV binding. The MAbs established in this study are useful for studying the interaction between IBDV and its target cell. PMID- 11258466 TI - Detection of cell membrane proteins that interact with virulent infectious bursal disease virus. AB - To detect the molecules that interact with infectious bursal disease virus (IBDV), the chicken B lymphoblastoid cell line, LSCC-BK3, which is permissive for virulent IBDV infection was investigated. The sodium dodecyl sulfate-solubilized plasma membrane fraction from the cells was subjected to a virus overlay protein binding assay. The IBDV specifically bound to proteins in LSCC-BK3 plasma membranes with molecular weights of 70, 82 and 110 kDa. This is the first report to demonstrate cellular molecules that interact with virulent IBDV. PMID- 11258467 TI - Imaging of the lectin-labeled cell surface of human lymphocytes by the use of atomic force microscope. AB - The atomic force microscope (AFM) is a new useful tool to examine the surface structure of specimens with a higher resolution than the conventional scanning electron microscope. In the present study, we used the AFM to observe the surface of paraformaldehyde-fixed human lymphocytes processed for histochemistry using a biotinylated lectin, wheat germ agglutinin, followed by colloidal gold and silver enhancement method. Before the treatment, no particles were attached to the cell surface. After treatment, many particles about 100 to 150 nm in diameter were visualized on it. Since we could observe the same cells on the slide glass before and after treatment, the AFM has the advantage to enable us the repeated imaging of samples treated with various kinds of histochemistries. PMID- 11258468 TI - Back to Darwin? PMID- 11258469 TI - Comment on the interview with Philippe Busquin in EMBO reports, August 2000. PMID- 11258470 TI - Reprogenetics: third millennium speculation. The consequences for humanity when reproductive biology and genetics are combined. PMID- 11258471 TI - Vaccine cornucopia. Transgenic vaccines in plants: new hope for global vaccination? PMID- 11258472 TI - A voice for European life scientists. An interview with Kai Simons, President of the European Life Science Organisation and Director of the Max Planck Institute for Molecular Cell Biology and Genetics in Dresden. PMID- 11258473 TI - Can they think for themselves? The coming generation of scientists must learn more in school than simple fact memorising. PMID- 11258474 TI - The discovery of functional genomics. EU announces new funding approach for European research. PMID- 11258475 TI - Preparing for the worst. The USA and Japan's preparations for a terrorist attack with chemical or biological weapons. PMID- 11258476 TI - Light, flies and cell division. Workshop: regulation of cell division in Drosophila. PMID- 11258477 TI - A new era for the RNA world. Conference: RNA 2000. PMID- 11258478 TI - The atypical protein kinase Cs. Functional specificity mediated by specific protein adapters. AB - Since its discovery more than 10 years ago, the atypical PKC (aPKC) subfamily has attracted great interest. A number of reports have shown that the kinases of this subfamily play critical roles in signaling pathways that control cell growth, differentiation and survival. Recently, several investigators have identified a number of aPKC-interacting proteins whose characterization is helping to unravel the mechanisms of action and functions of these kinases. These interactors include p62, Par-6, MEK5 and Par-4. The details of how these adapters serve to link the aPKCs to different receptor signaling pathways and substrates in response to specific stimuli are crucial not only for developing an understanding of the roles and functions of the aPKCs themselves, but also for more generally establishing a view of how specificity in signal transduction is achieved. PMID- 11258479 TI - Protein unfolding by mitochondria. The Hsp70 import motor. AB - Protein unfolding is a key step in the import of some proteins into mitochondria and chloroplasts and in the degradation of regulatory proteins by ATP-dependent proteases. In contrast to protein folding, the reverse process has remained largely uninvestigated until now. This review discusses recent discoveries on the mechanism of protein unfolding during translocation into mitochondria. The mitochondria can actively unfold preproteins by unraveling them from the N terminus. The central component of the mitochondrial import motor, the matrix heat shock protein 70, functions by both pulling and holding the preproteins. PMID- 11258480 TI - Finding nuclear localization signals. AB - A variety of nuclear localization signals (NLSs) are experimentally known although only one motif was available for database searches through PROSITE. We initially collected a set of 91 experimentally verified NLSs from the literature. Through iterated 'in silico mutagenesis' we then extended the set to 214 potential NLSs. This final set matched in 43% of all known nuclear proteins and in no known non-nuclear protein. We estimated that >17% of all eukaryotic proteins may be imported into the nucleus. Finally, we found an overlap between the NLS and DNA-binding region for 90% of the proteins for which both the NLS and DNA-binding regions were known. Thus, evolution seemed to have used part of the existing DNA-binding mechanism when compartmentalizing DNA-binding proteins into the nucleus. However, only 56 of our 214 NLS motifs overlapped with DNA-binding regions. These 56 NLSs enabled a de novo prediction of partial DNA-binding regions for approximately 800 proteins in human, fly, worm and yeast. PMID- 11258481 TI - Genome-wide insertional mutagenesis in human cells by the Drosophila mobile element Minos. AB - The development of efficient non-viral methodologies for genome-wide insertional mutagenesis and gene tagging in mammalian cells is highly desirable for functional genomic analysis. Here we describe transposon mediated mutagenesis (TRAMM), using naked DNA vectors based on the Drosophila hydei transposable element Minos. By simple transfections of plasmid Minos vectors in HeLa cells, we have achieved high frequency generation of cell lines, each containing one or more stable chromosomal integrations. The Minos-derived vectors insert in different locations in the mammalian genome. Genome-wide mutagenesis in HeLa cells was demonstrated by using a Minos transposon containing a lacZ-neo gene trap fusion to generate a HeLa cell library of at least 10(5) transposon insertions in active genes. Multiple gene traps for six out of 12 active genes were detected in this library. Possible applications of Minos-based TRAMM in functional genomics are discussed. PMID- 11258482 TI - An enhanceosome containing the Jun B/Fra-2 heterodimer and the HMG-I(Y) architectural protein controls HPV 18 transcription. AB - Recent studies have reported new mechanisms that mediate the transcriptional synergy of strong tissue-specific enhancers, involving the cooperative assembly of higher-order nucleoprotein complexes called enhanceosomes. Here we show that the HPV18 enhancer, which controls the epithelial-specific transcription of the E6 and E7 transforming genes, exhibits characteristic features of these structures. We used deletion experiments to show that a core enhancer element cooperates, in a specific helical phasing, with distant essential factors binding to the ends of the enhancer. This core sequence, binding a Jun B/Fra-2 heterodimer, cooperatively recruits the architectural protein HMG-I(Y) in a nucleoprotein complex, where they interact with each other. Therefore, in HeLa cells, HPV18 transcription seems to depend upon the assembly of an enhanceosome containing multiple cellular factors recruited by a core sequence interacting with AP1 and HMG-I(Y). PMID- 11258483 TI - Two distinct action mechanisms of immunophilin-ligand complexes for the blockade of T-cell activation. AB - The immunosuppressive effects of cyclosporin A (CsA) and FK506 are mediated through binding to immunophilins. Here we show that FK506-FKBP complex suppresses the activation of JNK and p38 pathways at a level upstream of mitogen-activated protein kinase (MAPK) kinase kinase (MAPKK-K) besides the calcineurin-NFAT pathway. A238L, a viral gene product that binds to immunophilin, also blocks activation of both pathways. In contrast, direct inhibitors of calcineurin, Cabin 1 and FR901725, suppress the activation of NFAT but not the JNK or p38 pathway. We further demonstrate that co-expression of a constitutively active NFAT and a constitutively active MEKK1 renders the interleukin-2 promoter in Jurkat T lymphocytes resistant to CsA and FK506, whereas Jurkat cells expressing a constitutively active NFAT alone are still sensitive to CsA or FK506. Therefore, CsA and FK506 exert their immunosuppressive effects through targeting both the calcineurin-dependent NFAT pathway and calcineurin-independent activation pathway for JNK and p38. PMID- 11258484 TI - One RNA polymerase serving two genomes. AB - The land plant Arabidopsis thaliana contains three closely related nuclear genes encoding phage-type RNA polymerases (RpoT;1, RpoT;2 and RpoT;3). The gene products of RpoT;1 and RpoT;3 have previously been shown to be imported into mitochondria and chloroplasts, respectively. Here we show that the transit peptide of RpoT;2 possesses dual targeting properties. Transient expression assays in tobacco protoplasts as well as stable transformation of Arabidopsis plants demonstrate efficient targeting of fusion peptides consisting of the N terminus of RpoT;2 joined to green fluorescent protein to both organelles. Thus, RpoT;2 might be the first RNA polymerase shown to transcribe genes in two different genomes. RNA polymerase activity of recombinant RpoT;2 is uneffected by the inhibitor tagetin, qualifying the gene product of RpoT;2 as a phage-type polymerase. PMID- 11258485 TI - The class 2 selenophosphate synthetase gene of Drosophila contains a functional mammalian-type SECIS. AB - Synthesis of monoselenophosphate, the selenium donor required for the synthesis of selenocysteine (Sec) is catalyzed by the enzyme selenophosphate synthetase (SPS), first described in Escherichia coli. SPS homologs were identified in archaea, mammals and Drosophila. In the latter, however, an amino acid replacement is present within the catalytic domain and lacks selenide-dependent SPS activity. We describe the identification of a novel Drosophila homolog, Dsps2. The open reading frame of Dsps2 mRNA is interrupted by an UGA stop codon. The 3'UTR contains a mammalian-like Sec insertion sequence which causes translational readthrough in both transfected Drosophila cells and transgenic embryos. Thus, like vertebrates, Drosophila contains two SPS enzymes one with and one without Sec in its catalytic domain. Our data indicate further that the selenoprotein biosynthesis machinery is conserved between mammals and fly, promoting the use of Drosophila as a genetic tool to identify components and mechanistic features of the synthesis pathway. PMID- 11258487 TI - Our fight continues. PMID- 11258486 TI - Genetic disruption of mineralocorticoid receptor leads to impaired neurogenesis and granule cell degeneration in the hippocampus of adult mice. AB - To dissect the effects of corticosteroids mediated by the mineralocorticoid (MR) and the glucocorticoid receptor (GR) in the central nervous system, we compared MR-/- mice, whose salt loss syndrome was corrected by exogenous NaCI administration, with GR-/- mice having a brain-specific disruption of the GR gene generated by the Cre/loxP-recombination system. Neuropathological analyses revealed a decreased density of granule cells in the hippocampus of adult MR-/- mice but not in mice with disruption of GR. Furthermore, adult MR-/- mice exhibited a significant reduction of granule cell neurogenesis to 65% of control levels, possibly mediated by GR due to elevated corticosterone plasma levels. Neurogenesis was unaltered in adult mice with disruption of GR. Thus, we could attribute long-term trophic effects of adrenal steroids on dentate granule cells to MR. These MR-related alterations may participate in the pathogenesis of hippocampal changes observed in ageing, chronic stress and affective disorders. PMID- 11258488 TI - Pulmonary manifestations in malaria. AB - Malaria, a major killer of mankind, apart from classical ague presentation, may present with respiratory manifestations. This may be misdiagnosed and important time may be lost in instituting antimalarials leading to higher morbidity and mortality. Present work was undertaken to study the clinical presentations of malaria with special reference to respiratory system and to evaluate the effect of antimalarials to such atypical presentation. One hundred slide positive cases of malaria were taken and detailed for respiratory involvement. Response to antimalarials was seen in these cases and associated complications (if any) were looked for. Mean age of the cases was 29.3 years with a male predominance. Positivity of peripheral smear read as: P vivax(53%), P falciparum (36%) and mixed infection (11%). Twenty-six patients had presented with respiratory manifestations-bronchitis (15), pneumonia (4), asthmatic bronchitis (1), adult respiratory distress syndrome (ARDS) (4) and pulmonary tuberculosis (2). Of these 26 cases, presenting symptoms noticed were cough (77%), dyspnoea (32%), expectoration (29%) and chest pain (15%). Twenty-five (96%) of these 26 patients were positive for P falciparum. Response to antimalarials was not significantly different in these 26 patients as compared to the rest (74 cases). All patients developing ARDS expired. The present study concludes that malarial atypical respiratory presentations are far higher in incidence than reported in literature. Peripheral smear examination in all patients of high grade fever with chills and rigors and having respiratory manifestations may unmask malarial infection and warrant early antimalarial treatment resulting in decreased morbidity and mortality. PMID- 11258489 TI - Review of trends of malaria in an urban community of Calcutta during 1984-1997. AB - The trends in the malaria situation over the last 14 years in an urban community of Calcutta with a population of 1,04,000 have been analysed with particular reference to the malaria epidemic which took place in Calcutta in November December, 1995. Annual and monthwise data on malariometric indices pertaining to 1984-1997 were collected from health centre records. The total number of blood slides examined every year has steadily increased from around 8,000 in 1984 to more than 23,000 in 1996 with some decline in 1997. About 95% of the confirmed malaria cases were of the benign tertian type, while remaining 5% were of the malignant tertian type. The slide vivax rate has steadily declined over the years 1984-1997 from around 50 per cent in 1984 to between 20 to 30 per cent during 1993-1997. The slide falciparum rate over the same period varied between 0.5 to 2 per cent and did not show any clear secular trend. The annual parasite incidence (API) which varied cyclically between 40 to 60 per thousand during 1984-87 increased (with a cyclical trend superimposed on a secular increasing trend) to vary between 50 to 70 per thousand during 1994-97. The 1995 winter epidemic of malaria was characterised by an increased occurrence of both P vivax and P falcipanrum malaria. PMID- 11258490 TI - Severe falciparum malarial complications in Ukhrul, Manipur. AB - Development of complications is very common among the patients suffering from Plasmodium falciparum infection. A total of 64 patients of Plasmodium falciprum infections were admitted to the District Hospital, Ukhrul, during the period of 1st May 1996 to 15th June 1999; 9.37% patients do not develop complication while the rest 90.63% developed one or more complications. The most common complication is anaemia accounting for 76.56% followed by cerebral malaria (59.38%). Other lesser complications were leucopenia (15.63%), thrombocytopenia (26.56%), adult respiratory distress syndrome (6.25%). There is no single record of blackwater fever, 12.5% died due to development of multiple complications like severe haemolytic anaemia, haemolytic jaundice, cerebral malaria and acute renal failure. This study confirms presence of severe and complicated falciparum malaria in this part of the country. PMID- 11258491 TI - Laboratory diagnosis of malaria. AB - Malaria particularly falciparum malaria is a major public health problem in India. Its correct and early diagnosis is very important for prompt treatment as a preventive and control measure. Microscopy is the traditional method for laboratory diagnosis of malaria, which is used widely. However, it is time consuming, needs expertism and the detection limit is 10-20 parasites/ml blood in thick film and 100 parasites/ml blood in thin film. Quantitative buffy coat technique (QBC) is highly sensitive method but expensive equipment is needed for this test. The serological methods involving antibody detection give information regarding exposure to malaria but do not differentiate between present and past infections. Genetic probes and PCR are highly sensitive methods but require expensive equipments. Parasite antigen detection tests are useful in field and PHC level for rapid diagnosis of P falciparum malaria.LDH based test for diagnosing malaria is sensitive but can not differentiate between species. For monitoring of drug resistance or follow-up of patients, methods which can quantify parasitaemia are needed. Simply microscopy is the best solution at present. PMID- 11258492 TI - Guideline in management of severe malaria. AB - Severe malaria remains a major cause of mortality in the world. Malaria can mimic many diseases and there is no absolute diagnostic clinical features. High index of suspicion is clue for clinical diagnosis. Previous travel history to endemic area should be elicited in all, and in particular, febrile patients. Management of severe malaria needs potent antimalarial drug and intensive care. Artemisinin derivatives can be of altemative use to quinine. Dexamethasone and mannitol have no beneficial value in the management of cerebral malaria. In pulmonary oedema patients whose hydration assessments are difficult to monitor, central venous pressure evaluation may be useful. Acute renal failure patients may need dialysis until uraemic syndrome subsides or patients can void urine. Most severe malaria patients have thrombocytopenia; however, platelet concentrate transfusion is indicated only in patients with systemic bleeding. Morbidity and mortality will be reduced in severe malaria patients with early diagnosis and prompt treatment. PMID- 11258493 TI - Malaria--an overview. AB - Starting from the history, clinical features, diagnosis, preventive and public health dimensions of malaria the paper ends in treatment aspect and feasibility of anti-malarial vaccine. An overview on malaria can be appreciated from the article. PMID- 11258494 TI - Virulent resurgence of malaria in Calcutta, West Bengal. AB - The reasons of virulent resurgence of malaria and the measures adopted for its control in a metropolitan city by Calcutta Municipal Corporation is discussed in detail along with proper references. Publicity compaign, especially among school students, has been stressed. PMID- 11258495 TI - Hypomagnesaemia. AB - In the management of electrolyte abnormalities along with hypokalaemia and hypocalcaemia, etc, presence of hypomsgenesaemia should be thought of even when serum magnesium is within normal limit as magneium is a predominantly intracellulr ion. Two cases of hypomagnesaemia as one of the factors of electrolyte distrubances are communicated in this write-up. PMID- 11258496 TI - Say no to indiscriminate use of newer antimalarials. PMID- 11258497 TI - Phenolphthalein: an unacceptable laxative. PMID- 11258499 TI - Eradication of leprosy. PMID- 11258501 TI - Comparative trial to evaluate the efficacy and tolerability of cefuroxime 250mg with probenecid 250mg with cefuroxime 500mg in the management of community acquired pneumonia, acute bronchitis and acute exacerbation of chronic bronchitis. PMID- 11258502 TI - Osteoporosis epidemiology review and panacea osteoporosis evaluation study. PMID- 11258504 TI - What causes smear-negative pulmonary tuberculosis in Malawi, an area of high HIV seroprevalence? AB - SETTING: The Central Hospital and the District Tuberculosis (TB) Registry in Lilongwe, the capital of Malawi. In this setting smear-negative pulmonary tuberculosis (PTB) is diagnosed using clinical and radiographic criteria for TB, and mycobacterial cultures are not routinely available. OBJECTIVE: To determine the proportion of patients being registered for smear-negative PTB treatment in Lilongwe who have TB that can be confirmed microbiologically. DESIGN: Prospective cohort study of patients about to start treatment under operational conditions for smear-negative PTB in Lilongwe between October 1997 and June 1998. Patients referred to the study team underwent a detailed clinical re-assessment, testing for human immunodeficiency virus (HIV), repeat sputum smear microscopy for acid fast bacilli and mycobacterial cultures of sputum and blood. Bronchoscopy and bronchoalveolar lavage (BAL) were performed and BAL fluid was examined for TB, Pneumocystis carinii and other fungi. RESULTS: Of 352 smear-negative PTB suspects assessed, the diagnosis of TB was confirmed in 137 (39%) cases. Eighty-nine per cent of patients assessed were HIV-positive, of whom 81% met the expanded case definition for the acquired immune-deficiency syndrome (AIDS). CONCLUSION: TB was the most commonly confirmed diagnosis amongst patients about to start treatment for smear-negative PTB in an area of high background HIV seroprevalence. PMID- 11258503 TI - Tuberculosis control in the era of the HIV epidemic: risk of tuberculosis infection in Tanzania, 1983-1998. AB - SETTING: In Tanzania, a national tuberculosis programme (NTP) was established in 1979 based on the principles currently known as the World Health Organization DOTS strategy. From the period 1983-1987 to 1994-1998, notification rates of smear-positive tuberculosis increased from 32 to 69 per 100,000 population, mainly due to the human immunodeficiency virus (HIV) epidemic. OBJECTIVES: To estimate the trend in the annual risk of tuberculosis infection and to establish to what extent the opposing forces of improved tuberculosis control and HIV have had an impact on tuberculosis transmission. METHODS: Three national surveys were conducted in Tanzania among primary school children at 5-year intervals. The annual risk of tuberculosis infection and its trend were determined by tuberculin skin testing. RESULTS: The annual risk of infection in children without BCG scar using the criterion '17 mm + 2 x 18 mm' or more was estimated at 1.1% in 1983 1987, 1.0 in 1988-1992, and 0.9% in 1993-1998. There appears to have been little change in the annual risk of infection over the study period, either when using other criteria to define infection or in children with a BCG scar. The estimated number of infections per notified case decreased over time from 36 to 19. CONCLUSIONS: Despite strongly increased tuberculosis notification rates in adults, associated with the HIV epidemic, the risk of tuberculosis infection in children appears to have been stable over the past 15 years in Tanzania. This remarkable achievement is probably due to the impact of the NTP on tuberculosis transmission. PMID- 11258505 TI - TB prevention in HIV clinics in New York City. AB - SETTING: Ten hospital-based human immunodeficiency virus (HIV) clinics in New York City. OBJECTIVE: To evaluate tuberculosis (TB) prevention in HIV clinics based on the prevalence and incidence of TB and the efficacy of preventive therapy with isoniazid (INH). DESIGN: The medical records of 2393 HIV-infected patients with a first clinic visit in 1995 were reviewed retrospectively. Deaths and TB cases through December 1997 were ascertained through a match with the TB and AIDS registries. RESULTS: At first visit, 92 patients (4%) had a history of TB, 98 (4%) were being treated for TB, and six (<1%) were diagnosed with TB. During follow-up, 23 cases were diagnosed, an incidence of 0.53 per 100 person years (py) (95%CI 0.34-0.77). Among 439 tuberculin skin test (TST) positive patients, the incidence of TB/100 py was 1.63 (95%CI 0.27-5.02) in patients with no INH, 1.28 (95%CI 0.40-2.98) in patients with <12 months of INH, and 1.06 (95%CI 0.38-2.28) in patients with 12 months of INH. The incidence/100 py was 0.0 (95%CI 0.0-0.78) in TST-negative patients and 0.37 (95%CI 0.09-0.95) in anergic patients. The relative risk of TB was 0.65 (95%CI 0.14-4.56) in TST-positive patients with 12 months of INH (vs. none). CONCLUSIONS: The benefits of TB prevention efforts in these HIV clinics from 1995 to 1997 were limited because most TB occurred before the first clinic visit. Methods for reaching HIV-infected patients earlier should be identified. PMID- 11258506 TI - Low failure rate in standardised retreatment of tuberculosis in Nicaragua: patient category, drug resistance and survival of 'chronic' patients. AB - SETTING: IUATLD collaborative programme, Nicaragua. OBJECTIVE: To analyse reported trends in the retreatment failure rate (2SRHZE/1RHZE/5R3H3E3), and assess demographic characteristics, drug resistance and survival in patients who fail retreatment. DESIGN: A retrospective, descriptive study. Reports from 1988 1996 were analysed and records of 69 patients who failed retreatment were reviewed. RESULTS: The treatment success rate in new cases improved from 71% in 1988-1991 to 79% in 1992-1996, the default rate decreased from 16% to 10%, and the failure rate remained stable at 2-3%. The proportion of previously treated patients among all smear-positives decreased from 20% to 15%. In retreatment patients the failure rate declined from 6.6% to 4.3% and the average annual number of failures from 24 to 13. In 1992-1996, 64 patients, 0.8% of new smear positive cases treated during this period, failed retreatment; the corresponding figures for 1988-1991 are 95 and 1.6%. Among 69 retreatment failure cases reviewed, there was male predominance and increasing age during the study period. Drug susceptibility results were available for 38, of whom 89% were resistant to isoniazid and rifampicin. The median survival of patients after failure was 3.9 years. CONCLUSION: Treatment results improved over the study period. The proportion of patients on retreatment out of all smear positives treated decreased, as did the absolute number of failures and the retreatment failure rate. Development of multidrug resistance has been largely prevented in Nicaragua; the low failure rate justifies the continued use of the standardised retreatment regimen. PMID- 11258507 TI - Treatment of tuberculosis in a rural area of Haiti: directly observed and non observed regimens. The experience of H pital Albert Schweitzer. AB - SETTING: Artibonite Valley, a rural area in Haiti. OBJECTIVE: To evaluate a tuberculosis control program in rural Haiti and to compare two strategies for treatment implemented in two areas that were not chosen at random: treatment delivered at the patients' homes observed by former tuberculosis patients (DOT), and non observed treatment (non-DOT). DESIGN: Retrospective analysis of the clinical records of adult patients diagnosed with tuberculosis at H pital Albert Schweitzer in Deschapelles, Haiti, during 1994-1995. RESULTS: There were 143 patients in the non-DOT group and 138 patients in the DOT group. The results of treatment were significantly different: in the non-DOT group 29% defaulted, 12% died and 58% had a successful outcome; in the DOT group 7% defaulted (P < 0.01), 4% died (P = 0.01) and 87% had a successful outcome (P < 0.01). These differences are also significant when considering only human immunodeficiency virus (HIV) infected patients (defaulted P < 0.01; died P = 0.09; successful outcome P < 0.01). CONCLUSION: Delivering treatment in patients' homes with direct observation by former tuberculosis patients can achieve good results, even in an area of extreme poverty and high rates of HIV infection. In this population the number of patients who are able to complete their treatment without observed administration is far from optimal. PMID- 11258508 TI - Trends in the prevalence and incidence of tuberculosis in south India. AB - OBJECTIVE: To study trends in the prevalence and incidence of tuberculosis in south India. METHODS: In 1968-1970, about 100,000 subjects were surveyed for tuberculosis and followed thereafter for 15 years, mainly by repeat survey once every 2.5 years. New entrants were inducted at every repeat survey. Radiographic examination of subjects aged 5 years or more and sputum smear and culture examinations of those with an abnormal shadow were undertaken; tuberculin tests were done initially on all, and at 4, 10 and 15 years in selected samples of those aged 1-9 years. RESULTS: The prevalence of culture-positive tuberculosis decreased by 1.4% per annum to 694/100,000, while that of smear-positive tuberculosis showed no significant decrease from 457/100,000. The annual incidence of culture-positive tuberculosis decreased by 4.3%/annum to 189/100,000 and that of smear-positive tuberculosis decreased by 2.3%/annum to 113/100,000. Decreases in incidence occurred exclusively in those with abnormal radiographic findings suggestive of tuberculosis at the start of the period. The annual risk of tuberculosis infection (ARTI) was initially 2%, and showed no sign of decline over the period. CONCLUSION: The prevalence of tuberculosis and ARTI showed little or no decrease over the 15-year period. A significant decrease in incidence occurred, but exclusively in those with abnormal radiograph suggestive of tuberculosis at the start of the period. PMID- 11258509 TI - A study of the variation in tuberculosis incidence and possible influential variables in Manchester, Liverpool, Birmingham and Cardiff in 1991-1995. AB - SETTING AND OBJECTIVE: The reversal of the decline in United Kingdom tuberculosis rates has sparked a resurgence of interest in the epidemiology and prevention of tuberculosis in the UK. In this paper we quantify the primary factors explaining the variability in the electoral ward level relative risk of tuberculosis in Manchester, Liverpool, Birmingham and Cardiff. DESIGN: Ecological analysis of the incidence of tuberculosis in 397 wards using hospital admissions data as a proxy for tuberculosis incidence. Admissions were evaluated from the financial years 1991/1992 to 1994/1995. Ward level covariates included measures of country of birth, ethnicity and various socio-economic measures. RESULTS: Separate analyses were carried out for pulmonary and non-pulmonary tuberculosis. For pulmonary tuberculosis the final model included measures of the ward population born in India and Pakistan, overcrowded housing and not-owner-occupied housing. For non pulmonary tuberculosis the covariates were ward population born in India and Pakistan, overcrowded housing and the proportion of households with no car. CONCLUSIONS: The country of birth of the ward population is the single most influential explanatory factor in the variability of the ward rates for both pulmonary and non-pulmonary tuberculosis in these four cities. Measures of poverty are of secondary importance. PMID- 11258510 TI - Risk for tuberculosis infection among internally displaced persons in the Republic of Georgia. AB - OBJECTIVE: To determine the prevalence of tuberculosis (TB) infection and disease among internally displaced persons residing in Tbilisi, Republic of Georgia. DESIGN: Residents of eight refugee hostels were screened for TB infection using a tuberculin skin test (TST) and a symptom questionnaire. Participation was voluntary. TST-positive individuals were referred for chest radiography. Subjects with cough, fever, or night sweats of > 2 weeks duration provided sputum for acid fast bacilli (AFB) microscopy and culture. RESULTS: Of approximately 4000 potential subjects (internally displaced persons), 988 (24.7%) participated in the screening program. Of these 988, 931 (94.2%) who had a TST placed returned at 48-72 hours to have the skin test examined; 447 (48.0%) were TST-positive (> or = 10 mm induration). In multivariate analysis, risk factors for a positive TST included male sex, ever having received BCG, history of close contact with a case of active tuberculosis, and living in one specific refugee hostel. Risk for a positive TST was greater among subjects > 20 years old, but there was no difference between age groups over the age of 20 years. Five patients with active TB were identified through the screening program, giving a case rate of 537 per 100,000 population. CONCLUSION: Tuberculosis infection and disease were common in this group of internally displaced persons. Screening was a useful mechanism of case finding among this high prevalence population. PMID- 11258511 TI - Tuberculosis as an occupational hazard for health care workers in Estonia. AB - SETTING: Tuberculosis incidence has been increasing in the Baltic states since the 1990s, accompanied by the emergence of drug resistance, including multidrug resistance (MDR). In this changing situation, the potential threat of nosocomial spread of tuberculosis to other patients and health care workers (HCW) has remained unrecognised. OBJECTIVE: To investigate the risk of tuberculosis in health care workers in Estonia. DESIGN: Cases of tuberculosis registered among HCWs from 1994 to 1998 were evaluated. The case records were analysed retrospectively and combined with bacteriological data including data on drug resistance. RESULTS: Sixty-seven HCWs (23 physicians, 23 nurses and seven laboratory technicians, 12 assistant nurses and two cleaners), all of whom tested negative for human immunodeficiency virus, were diagnosed as having active tuberculosis. The incidence of tuberculosis among HCWs (mean 91/100,000/year) was 1.5 to three times higher than in the general population. In a chest hospital in charge of regional tuberculosis care, the incidence was 30 to 90 times higher, and was highest among physicians. In 49 HCWs tuberculosis was confirmed by culture. Among these, drug resistance was detected in 23 (49%), 18 (38%) of whom had MDR tuberculosis. CONCLUSIONS: Health care workers, especially those working in a chest hospital where tuberculosis patients were treated, were found to be at an elevated risk of tuberculosis. MDR tuberculosis poses a particular threat which is difficult to combat. PMID- 11258512 TI - Tuberculosis risk factors in a silicotic cohort in Hong Kong. AB - SETTING: Hong Kong silicotic patients are followed regularly at the Pneumoconiosis Clinic. OBJECTIVE: To quantify the incidence of tuberculosis disease that occurs after the date of diagnosis of silicosis (DOD), and to identify the risk factors for its development. DESIGN: Retrospective analysis of a Hong Kong silicotic cohort with DOD from 1 January 1988 to 31 December 1993. RESULTS: Of 718 subjects whose records were identified, 11 were excluded from the study. The incidence of tuberculosis (TB) after DOD was 3019 patients per 100,000 population, approximately nine times that of the local population matched for age and sex. Twelve factors were subject to univariate analysis followed by logistic regression. Four TB risk factors were identified: 1) no anti-tuberculosis treatment before DOD (relative risk [RR] 4.51, 95% confidence interval [CI] 2.46 8.24), 2) progressive massive fibrosis (PMF) (RR 3.78, 95%CI 2.25-6.36), 3) small opacities exceeding 1.5 mm (RR 2.17, 95%CI 1.38-3.42), and 4) caisson work (RR 1.56, 95%CI 1.01-2.41). Relative risks were calculated for patient subgroups stratified according to TB risk factors. CONCLUSION: This study has reaffirmed the strong association of tuberculosis and silicosis, and has identified several TB risk factors with a logistic regression model. PMID- 11258513 TI - Serial evaluation of serum neopterin in HIV seronegative patients treated for tuberculosis. AB - OBJECTIVE: To delineate the course of serum neopterin (s-neo) concentrations in patients with pulmonary tuberculosis who are on anti-tuberculosis therapy. DESIGN: S-neo concentrations were measured by high performance liquid chromatography (HPLC) in 39 patients treated for pulmonary tuberculosis at pretreatment, at one month and at end of treatment. It was also measured in 11 relapse cases and their matched controls at the above time points and at the time of relapse. The results were correlated with bacteriological and radiological findings. RESULTS: All patients had elevated levels of s-neo at pretreatment which had declined at 1 month and were near normal at the end of treatment. The decline was more significant in patients with moderate lesions, suggesting that immune activation is maximum in this group of patients. The mean decrease was 37% at one month and 66% at the end of treatment. The corresponding decreases were 11% and 56% in patients with limited lesions and 11% and 45% in those with extensive lesions. It continued to fall after completion of therapy in patients who did not relapse, whereas an increase after completion of therapy was associated with bacteriologically proven relapse. CONCLUSIONS: The measurement of s-neo concentration could be of help in evaluating response to therapy. This study provides a rational basis for the association between s-neo concentration and relapse. PMID- 11258514 TI - Validity of the IUATLD (1986) questionnaire in the EGEA study. International Union Against Tuberculosis and Lung Disease. Epidemiological study on the Genetics and Environment of Asthma, bronchial hyperresponsiveness and atopy. AB - SETTING: No validity study of the IUATLD asthma-like questions has been performed in a group of well-defined, clinically-based asthmatics. OBJECTIVES: To assess the validity of the questions regarding asthmatics included in a case-control study, and to assess their validity as regards bronchial hyperresponsiveness in population-based subjects. DESIGN: Data from the case-control Epidemiological study on the Genetics and Environment of Asthma, bronchial hyperresponsiveness and atopy (EGEA) were used. The analysis concerned 201 adult asthmatic cases recruited in chest clinics through a standardised protocol and 284 population based controls. RESULTS: The analysis of the case-control study regarding asthma showed a sensitivity of 0.56, 0.68 and 0.86 for nocturnal symptoms of cough, shortness of breath, and chest tightness, and a specificity of 0.72, 0.98 and 0.89. As regards bronchial hyperresponsiveness (PD20 < or = 4 mg) in the control group, specificity was high (0.77, 0.99 and 0.90), but sensitivity was markedly lower (0.36, 0.11 and 0.20). CONCLUSION: Asthma-like symptoms assessed by the IUATLD questionnaire have good validity, both for specificity and sensitivity, for asthma patients recruited in chest clinics. In general populations, questions have a high specificity, an important criterion in aetiological epidemiological research, and they were designed in that perspective. However their moderate sensitivity limits their usefulness as a screening test. PMID- 11258515 TI - Multicenter study of incidence of Mycobacterium marinum in humans in Spain. AB - A retrospective survey was carried out on the epidemiological monitoring of the isolation incidence of Mycobacterium marinum in 21 laboratories, in an attempt to gain an insight into the frequency of its isolation in Spain. From 1991 to 1998, 39 cases (22 male and 17 female) of bacteriologically confirmed M. marinum infection were accumulated. The majority of the cases (35) were fish related. The clinical presentation usually suggested sporotrichosis, with the majority of skin eruptions appearing on the hand (32 cases). For treatment, minocycline was recommended as the drug of choice in 12 cases, rifampicin in eight cases and clarithromycin and ethambutol in seven cases. PMID- 11258516 TI - The irreversible cost of delayed diagnosis of tuberculosis in HIV co-infected persons in sub-Saharan Africa. PMID- 11258517 TI - Why the Indian TB control programme must stop ignoring private practitioners. PMID- 11258518 TI - An efficient batch preparation of high specific activity. AB - The increasing demand for radiolabeled metaiodobenzylguanidine (mIBG) prompted the need to obtain the radiopharmaceutical by a reliable, routine and simple synthetic method for batch production. The production of mIBG labeled with either 123I or 124I has been optimized by modifying literature methods that involve solid-state exchange reaction on "cold" mIBG facilitated by ammonium sulfate. The radiochemical yield and purity of radioiodinated mIBG generally exceeded 80 and 98%, respectively, with specific activity of > 50 mCi/mg. PMID- 11258519 TI - Radioisotope studies of root activity and root-level interactions in tree-based production systems: a review. AB - This review focuses on the radioisotope methods used for studying the root activity patterns of woody perennials and root competition in multi-species cropping systems. 32P soil-injection is by far the most widely used method. The method is unique as it enables one to delineate the lateral and vertical spread of the absorbing roots. A system of classification of crop plants based on the concept of 'effective foraging space' has been developed. The scope and applicability of the radioisotope methods in different cropping situations as well as from the point of view of radiological safety were discussed. PMID- 11258520 TI - Analysis of arsenic pollution in groundwater aquifers by X-ray fluorescence. AB - The serious contamination of groundwater in the southeastern plain of the province of Cordoba (Argentina), a phenomenon mentioned in the literature for over 80 years, has given rise to this initial hydrologic study covering an area over 250 km2. This study analyzes a rural area near a little town called La Francia, and is motivated by the existence of an important pollution with arsenic in the first-aquifer groundwater of the region. This phenomenon has been mentioned for a long time and evidenced by the high incidence of diseases associated with this element in the local population. By means of the X-ray fluorescence (XRF) technique, and using an energy-dispersive spectrometer, 50 samples of groundwater of the rural zone of La Francia from about 100 m deep (second aquifer), were analyzed. The samples were excited with a 3 kW X-ray tube and measured using a reflecting geometry with 45 of incident and take-off directions. Preconcentration techniques for the preparation of the samples were employed in order to obtain an adequate signal-to-noise ratio. The As concentration in water was obtained using calibration curves and the internal standard method for quantification. A high percentage of the analyzed samples showed concentrations lesser than or equal to 0.05 mg l(-1). This value corresponds to the maximum pollutant level for humans. The maximum measured value reaches 3 mg l(-1) in samples collected in perforations of first-aquifer wells and in some second-aquifer isolated wells. PMID- 11258521 TI - Preparation of the thicker americium targets by molecular plating. AB - Electrodeposition of americium from the mixture of isopropyl alcohol and dilute nitric acid was studied. The 241,243Am targets with thickness between 600 microg/cm2 and 1.2 mg/cm2 had been prepared on thin aluminum foil (7 microm) by molecular plating at one time. PMID- 11258523 TI - Self-radioiodination of iodogen. AB - Iodogen (1,3,4,6-tetrachloro-3alpha,6alpha-diphenylglucoluril) is commonly used for the radioiodination of proteins as an oxidative agent. The oxidative character of iodogen is not clear, but the two carbonyl groups in its structure probably have an essential role in its oxidizing character. In this study, the self-radioiodination of iodogen has been examined. It was observed that about 10 20% of the initial iodine radioactivity was consumed for the self-radioiodination of iodogen itself. On the other hand, the radioiodinated iodogen removed by ethyl alcohol from the iodogen-coated tubes showed clearly that no thyroid uptake was observed and that it was rapidly cleared out from the whole body of a rabbit administered with the radioiodinated iodogen by injection via the ear vein. PMID- 11258522 TI - Synthesis and biodistribution of a new 99mTc nitrido complex for brain imaging. AB - Bis(N-isobutyl-dithiocarbamato) nitrido technetium-99m complex [99mTcN(IBDTC)2] (IBDTC: N-isobutyl dithiocarbamato) was synthesized by the reduction of 99mTcO4- into [99mTc = N]2+ with stannous chloride in the presence of succinic dihydrazide and propylenediamine tetraacetic acid, followed by the addition of sodium N isobutyl dithiocarbamate dihydrate. The radiochemical purity of the complex was over 90% as measured by thin layer chromatography. It was stable over 6h at room temperature. Its partition coefficient indicated that it is a good lipophilic complex. Biodistribution in mice demonstrated that the complex accumulated in the brain with high uptake and good retention. The brain uptake (ID%/g) was 6.22, 5.45 and 3.88 and the brain/blood ratio was 1.51, 2.24, 1.84 at 5, 30 and 60 min post-injection, respectively. These results suggest potential usefulness of the complex as a brain perfusion imaging agent. PMID- 11258524 TI - A water target with beam sweep for routine fluorine-18 production. AB - Fluorine-18 (18F) production with 18O-enriched water targets is well established world-wide. Heat transfer is still a critical point, however, when high beam currents are used to increase the amount of radioactivity per batch within an adequate irradiation period. The technique of beam sweep formerly used in Hannover for all targets has been adapted for this water target again. A titanium target with a beam sweep of up to 3 cm in the horizontal plane was designed to keep the total amount of enriched water low (2.06 ml), to allow beam currents of 30 microA for 1 h on a daily schedule and thereby to extend maintenance periods to more than 6 months. The design had to take into account that the target must be mounted on a target ladder with five additional positions and that the beam is carried by a 5.50 m beam line from a variable energy, multi-particle cyclotron (mc35, Scanditronix) to the target. The target has been in use for more than 130 consecutive runs with beam currents of about 30 microA at a target pressure of up to 15 bar without any maintenance. The mean value of the recovered radioactivity has reached 91.9+/-7.7% of the theoretically expected value at the end of bombardment (EOB). PMID- 11258525 TI - HPGe detector photopeak efficiency calculation including self-absorption and coincidence corrections for Marinelli beaker sources using compact analytical expressions. AB - Direct mathematical methods to calculate total and full-energy peak (photopeak) efficiencies, coincidence correction factors and the source self-absorption of a closed end coaxial HPGe detector for Marinelli beaker sources have been derived. The source self-absorption is determined by calculating the photon path length in the source volume. The attenuation of photons by the Marinelli beaker and the detector cap materials is also calculated. In the experiments gamma aqueous sources containing several radionuclides covering the energy range from 60 to 1836 keV were used. By comparison, the theoretical and experimental full-energy peak efficiency values are in good agreement. PMID- 11258526 TI - Coincidence summing corrections for the natural decay series in gamma-ray spectrometry. AB - Using a Monte Carlo code and a Markov formalism to describe the decay schemes, coincidence-summing correction factors can be calculated with a suitable accuracy. For two different measuring geometries and an HPGe detector, calculated and experimental correction factors have been shown to closely agree for 152Eu. The simulation method has subsequently been applied in assessing the need for coincidence-summing corrections for members of the uranium, thorium and actinium series measurable by gamma-spectrometry. Correction factors were calculated for predominant gamma emissions significantly affected by coincidence-summing effects and the correctness of our calculations tested for environmental samples. The test makes it evident that in order to obtain reliable and unbiased activity values for some natural radionuclides coincidence summing cannot be neglected in environmental measurements at small source-detector distances. PMID- 11258527 TI - Corrections for self-attenuation in gamma-ray spectrometry of bulk samples. AB - On the basis of the measurement of over 70 radioactive standard bulk sources with different matrix density and different shapes, the gamma-ray self-attenuation corrections needed in activity determination by means of gamma-ray spectrometry are evaluated. The full-energy peak efficiency dependence on the density, and the self-attenuation correction dependence on the photon energy are described. PMID- 11258528 TI - Comparison of measured and calculated emission probabilities for K and L X-rays following radioactive disintegration processes. AB - The program EMISSION has been developed at the Physikalisch-Technische Bundesanstalt (PTB). It allows emission probabilities to be calculated for K and L X-rays following radioactive disintegration processes. This program was already applied to the calculation of total K X-ray emission probabilities for six radionuclides. The results are found to be in reasonable agreement with measured values. In order to check the validity of this program also for the individual components of K and L X-rays, calculated emission probabilities of K X-ray components (139Ce, 203Hg) and L X-ray components (139Ce, 203Hg, 241Am) were compared with remeasured (139Ce, 203Hg) and already available (241Am) emission probabilities, measured earlier at the PTB. In most cases there is an agreement within the standard uncertainties of the measured and calculated values, usually within 1-5%. Larger deviations, not covered by three times the standard uncertainty have been found for some weak lines. PMID- 11258529 TI - Dependencies of the radiation sensitivity of human tooth enamel in EPR dosimetry. AB - The EPR dose response of tooth enamel was determined for human molars collected in Egypt. The influence of age, gender and residence of the tooth donors as well as tooth position and sample preparation on EPR sensitivity and its variability over the enamel samples was investigated. The EPR sensitivity and its variability were found to depend only on the sample preparation procedure. The variability in EPR sensitivity of enamel from Egyptian teeth was maximally 10% and the mean sensitivity was in good agreement with that of German teeth. PMID- 11258530 TI - Practical aspects of Compton scatter densitometry. AB - A technique for determining the physical density distribution within an object using Compton scattered radiation is described. The technique is based on measuring the variation with the depth of 90 scattered radiation flux from an external monoenergetic photon source using a high-resolution solid-state detector. The measured scattered signal from an isolated volume element of the interrogated object is proportional to its electron density. For a definite region within the scanned object, the variation of the scattered yield with depth, along the direction of the incident beam, will give a counting curve with slope proportional to the density of the examined region. The spatial resolution of the system, determined by the collimator width, is 2 mm. The mathematical aspects of the technique are presented and the experimental data obtained in line scans of representative tissue equivalent materials are also shown to demonstrate the validity of the method. PMID- 11258531 TI - Characterization and modeling of the metal diffusion from deep ultraviolet photoresist and silicon-based substrate. AB - The radioactive tracer technique was applied to investigate the out-diffusion of the transition metals (Cu, Fe and Co) from deep ultraviolet (DUV) photoresist into underlying substrate. Two important process parameters, viz., baking temperatures and substrate types (i.e., bare silicon, polysilicon, silicon oxide and silicon nitride), were evaluated. Results indicate that the out-diffusion of Co is insignificant, irrespective of the substrate type and baking temperature. The out-diffusion of Cu is significant for substrates of bare silicon and polysilicon but not for silicon oxide and nitride; for Fe, the story is reversed. The substrate type appears to strongly affect the diffusion, while the baking temperature does not. Also, the effect of solvent evaporation was found to play an important role in impurity diffusion. Using the method of numerical analysis, a diffusion profile was depicted in this work to describe the out-diffusion of metallic impurities from photoresist layer under various baking conditions. In addition, the effectiveness of various wet-cleaning recipes in removing metallic impurities such as Cu, Fe and Co was also studied using the radioactive tracer technique. Among the six cleaning solutions studied, SC2 and SPM are the most effective in impurity removal. An out-diffusion cleaning model was first proposed to describe the cleaning process. A new cleaning coefficient, h(T), was suggested to explain the cleaning effect. The cleaning model could explain the tracer results. PMID- 11258532 TI - Role of the activator in the performance of scintillators used in X-ray imaging. AB - The aim of this study was to investigate how the activator type affects the performance of X-ray scintillators. To this aim the behavior of scintillator materials was modeled under X-ray excitation conditions, similar to those used in imaging techniques. The model describes the light emission efficiency, the spectral compatibility with optical detectors (films, photodiodes, and photocathodes), and the imaging capabilities of a scintillating layer. Using the model equations the role of the activator type in scintillator performance was examined. Activators affect some important properties of materials, like the intrinsic X-ray to light conversion efficiency, the spectrum of the emitted light, and the light attenuation coefficients. The performances of a high efficiency material (Gd2O2S) combined with either Tb3+ or Eu3+ activators were compared. Results showed that the terbium-activated material exhibited high emission efficiency (number of emitted photons per incident X-ray) and modulation transfer function (spatial resolution and image contrast) while the europium activated material showed slightly better signal-to-noise ratio properties at low spatial frequencies. Both materials were found to exhibit high spectral compatibility with currently used modern optical detectors. In conclusion, the choice of activator may improve spectral compatibility, but care must be taken because it may also alter emission efficiency and image quality. PMID- 11258533 TI - Reconstruction of individual absorbed doses by tooth enamel on the base of non linear simulation of their EPR-spectra. AB - Model of separate spectral lines composing EPR-spectra of tooth enamel can be described with fine accuracy by integral of convolution of three distribution functions: Lorenzian, Gaussian and anisotropy. Simulation of spectra was done by the method of optimisation of non-linear parameters combined with the Gauss method of exclusion for linear parameters to obtain the minimum of the sum of the squares of the differences between the experimental EPR-spectrum and its model. The final result was the deconvolution of complex EPR-spectrum into its components (the background and radiation-induced signals) with the following reconstruction of the individual absorbed dose. PMID- 11258534 TI - Optimisation of cyclotron production parameters for the 209Bi(alpha, 2n) 211At reaction related to biomedical use of 211At. AB - The cyclotron alpha beam production of 211At and of the contaminant 210At related to beam energy were studied. Radiochemical purification of 211At from the other main contaminant, 210Po, by an extraction procedure was also evaluated. To avoid impurities 28MeV has previously been considered as a maximum beam energy, but by using instead 29.1 MeV as a limit a large increase in EOB yield and sufficient radiochemical purity of extracted 211At were obtained. More cyclotrons could thereby deliver quantities useful for clinical cancer trials. PMID- 11258535 TI - Asymptotic solutions of neutron transport equation and the limits of correct use of diffusion approximation for rocks. AB - The diffusion approximation solution for neutron transport has been used in well logging geophysics for calculating tool responses in boreholes, sometimes with success. The problem of the dimension of different materials to which it can be applied with success is important for the borehole environment. The results obtained show that the diffusion approximation can be used for distances greater than a few millimetre in some rock types. For iron, barium, and other highly absorbing media the use of the diffusion approximation is inappropriate even for large distances. PMID- 11258536 TI - A comparison of theoretical solutions of the three-layer coaxial diffusion approximation of the borehole with measurements at the Zielona Gora calibration facility. AB - Czubek has obtained a practically successful solution of a two-layer borehole geometry using the neutron diffusion approximation. Czubek and Woznicka solved the more realistic three-layer borehole model, which includes a middle layer consisting of mud-cake or steel tubing, in this approximation. The comparison with experimental data for 15-mm thick steel tubes shows some discrepancies. By using an exact Monte Carlo solution of the Boltzmann equation for calculating the migration length, measurements agree more closely with the "general calibration curve". This opens opportunities for cased borehole logging. PMID- 11258537 TI - Heavy metal pollution of some Danube Delta lacustrine sediments studied by neutron activation analysis. AB - Using neutron activation analysis and radiometric measurements, the vertical distribution of six possible pollutant elements (Zn, Cr, Co, As, Sb and Br), two trace elements of natural origin (Sc and Hf) and radioactive 137Cs were determined in three lakes, Furtuna, Lung and Mesteru, located in an active sedimentary zone of the Danube Delta. The accumulation rate of recent sediments was estimated from radiocesium profiles. Zn, As, Sb and Br were identified as possible pollutants. The vertical profiles of their concentrations reflect recent historic inputs, showing a gradual increase until around 1990, followed by a continuous decrease. PMID- 11258538 TI - Natural gamma-emiting radionuclides in Pakistani Portland cement. AB - Studies of the natural gamma-emitting radionuclides in Portland cement manufactured in the North West Frontier Province (NWFP) of Pakistan and the various raw materials which compose the product have been carried out using gamma spectrometric techniques. For data acquisition a high-purity germanium detector (HPGe) was used. The range of the total specific activity (minimum and maximum values) due to all the three radionuclides (40K, 226Ra and 232Th) were found to be 187.8+/-63.5-573.2+/-73.1 Bq kg(-1) (Portland cement); 54.5+/-16.1-183.9+/ 31.4 Bq kg(-1) (limestone); 87.1+/-30.7-297.1+/-64.4 Bq kg(-1) (gypsum); 696.4+/ 79.1-1043.9+/-85.0 Bq kg(-1) (slate); and 490.9+/-54.5-570.2+/-59.8 Bq kg(-1) (latrite). The average specific activities due to 40K in Portland cement and all the raw materials were found to be higher when compared with 226Ra and 232Th. Such materials do not pose any excess radiological health problem. PMID- 11258539 TI - Ellagic acid inhibited 2-aminofluorene and p-aminobenzoic acid acetylation by mononuclear leucocytes from Sprague-Dawley rats. AB - Following exposure of rats to the arylamine carcinogen 2-aminofluorene, DNA carcinogen adducts were found in the liver and bladder target tissues, and also in circulating leucocytes. This work investigated the effect of ellagic acid on arylamine (2-aminofluorene and p-aminobenzoic acid) acetylations in rat leucocytes. Evidence is presented that rat mononuclear leucocytes are capable of acetylating 2-aminofluorene and p-aminobenzoic acid. Both lymphocytes and monocytes were able to acetylate arylamines during 18 h of culture. Cultured lymphocytes produced about twice as much N-acetyl-2-aminofluorene from 2 aminofluorene and 2.2-fold as much N-acetyl-p-aminobenzoic acid from p aminobenzoic acid as monocytes. After cotreatment with ellagic acid the lymphocyte and monocyte cultures indicated that ellagic acid reduced 2 aminofluorene acetylation. PMID- 11258540 TI - Silver staining of nucleolar organizer regions (NOR) in some species of Hymenoptera (bees and parasitic wasp) and Coleoptera (lady-beetle). AB - Adaptations of the nucleolar organizer regions (NOR) banding technique using precipitation of silver salts significantly improved the NOR characterization of some species of hymenopterans and one coleopteran. The bee Melipona marginata (2n = 18) showed one metacentric pair of chromosomes with a NOR in the pericentromeric position. The parasitic wasp Mellitobia australica (2n = 12) also showed one metacentric pair with a strongly Ag-positive NOR. The male lady-beetle Cycloneda sanguinea (2n = 18 + Xy(p)) displayed a NOR on a pair of acrocentric autosomes. In the male Euglossa sp. (a haplodiploid species) (n = 21) the NOR were multiple, and occurred in five chromosomes. In the bee Plebeia sp. 1 (2n = 34) the NOR seemed restricted to one of the homologues of a metacentric pair. The systematic advances brought out by using this technique in the context of current theories of karyotypic evolution of these taxa are described and discussed. PMID- 11258542 TI - Genotypic diversity in species of Scilla. AB - Two cytotypes of Scilla indica and three other species, namely S. nervosa, S. siberica and S. vindobonensis were investigated. The estimation of nuclear DNA content was utilized as a measure of genetic diversity at inter- and intraspecific levels and as an index of the trends of evolution. S. siberica with a very high DNA value was regarded as indicative of a primitive state. The heavy intraspecific difference in DNA content in S. indica has been attributed to amplification of the DNA. PMID- 11258541 TI - Modulation of the renin-angiotensin system may alter the adrenocortical regeneration. AB - The effect of the renin-angiotensin system on adrenocortical regeneration has been studied in rats subjected to left adrenal enucleation combined with contralateral adrenalectomy. It was found that angiotensin II stimulated both proliferation and the steroidogenic capacity of the regenerating adrenal cortex cells by 6 days after operation. The stimulatory effect of angiotensin was prevented by losartan, a type 1 angiotensin (AT1) receptor antagonist, and by nifedipine, a calcium channel blocker. Losartan and nifedipine also depressed the adrenocortical regeneration when given alone. Enalapril, an angiotensin converting enzyme blocker, inhibited both the proliferation and steroidogenesis of the regenerating adrenal cortex, but this effect could be prevented by angiotensin II. PMID- 11258543 TI - White and UV light effects on cell nuclei in the aurea genotype of Lycopersicum esculentum L. AB - The effects of white light and UV light on chromosome endoreduplication, chromatin conformation and RNA synthesis were analysed in the hypocotyl cortical cylinder of the cv UC-105 of tomato and its isogenic mutant aurea, deficient in photoactive phytochrome A at the etiolated stage. Short white light or UV irradiations were administered to 4-day-old seedlings grown in the dark. White light had no effect on the mean ploidy level in the cv UC-105, but it increased the ploidy level in the aurea mutant. This is explained by hypothesizing that phytochrome inhibits endoreduplication, while cryptochrome stimulates it. UV light produced a higher ploidy in both genotypes, possibly because of residual action of cryptochrome at this wavelength or of specific UV-responsive photoreceptor. White light or UV light transiently increased heterochromatin amounts in the diploid nuclei of cv UC-105, and produced higher levels of RNA transcription than continuous dark. It is suggested that these responses were mediated by phytochrome, because they were lacking in the aurea mutant. PMID- 11258544 TI - Role of S-adenosyl-L-methionine in potentiating cadmium mobilization by diethylenetriamine penta acetic acid in mice. AB - The beneficial effects of S-adenosyl-L-methionine (SAM) in potentiating the mobilization of cadmium by cadmium trisodium diethylenetriamine penta acetic acid (DTPA) from the major target organs and restoration of depleted tissue glutathione (GSH), zinc and copper concentration, were determined in cadmium exposed mice. The results indicated a significant depletion of cadmium concentration from the blood in DTPA plus SAM treated animals compared with DTPA or SAM alone treated groups. The treatment with SAM alone was also effective in correcting the zinc and GSH concentrations. The results indicated few beneficial effects of concomitant SAM administration during chelation of cadmium with DTPA. PMID- 11258545 TI - The zero-stress state of the gastrointestinal tract: biomechanical and functional implications. AB - The stresses and strains that remain in an organ when the external load is removed (the no-load state) are called residual stresses and strains. They can be relieved by cutting up the organ to obtain the zero-stress configuration. This phenomenon was demonstrated more than 15 years ago in cardiovascular research but until recently it was not realized by the gastrointestinal research community. The function of the gastrointestinal tract is to propel food by peristaltic motion, which is a result of the interaction of the tissue forces in the wall and the hydrodynamic forces in the food bolus. To understand the tissue forces, it is necessary to know the stress-strain relationships of the tissues that must be measured in reference to the zero-stress state. It has become clear that the zero stress configuration of the gastrointestinal tract is very different from that of the no-load condition and that the zero-stress state is sensitive to structural and mechanical remodeling. The purpose of this review is to describe the basic theory and experiments of residual stress and to explore its physiological and pathophysiological implications in the gastrointestinal system. PMID- 11258546 TI - Evidence for a Na+-H+ exchange across human colonic basolateral plasma membranes purified from organ donor colons. AB - The mechanism(s) of electrolyte transport across the human colonic contraluminal domain is not well understood. Current studies were undertaken to develop a technique for the isolation and purification of the human colonic basolateral membrane vesicles (BLMV) and to examine the presence of a Na+-H+ exchange process in these membranes. BLMV were purified from mucosal scrapings of organ donor proximal colons utilizing a Percoll density gradient centrifugation technique, and Na+ transport was examined utilizing a rapid filtration, technique. Our data demonstrate that purified basolateral membranes were enriched 10- to 11-fold in Na+, K+-ATPase activity compared to crude homogenate. Results consistent with the Na+-H+ exchange in BLMV are as follows: (1) an outwardly directed H+ gradient stimulated 22Na uptake; (2) 22Na uptake was markedly inhibited by EIPA and amiloride; (3) H+-gradient-stimulated 22Na uptake was not inhibited by bumetanide, SITS, DIDS, acetazolamide, phenamil and benzamil; (4) 22Na uptake was voltage insensitive; (5) 22Na uptake demonstrated saturation kinetics; (6) 22 Na uptake was markedly inhibited by Na+ and Li+ but was unaffected by N-methyl glucamine+, choline+, and NH4+. Immunoblotting studies demonstrated this Na+-H+ exchanger isoform to be represented by NHE1. In conclusion, a technique has been established for the purification of functional human proximal colonic BLMV, and an electroneutral Na+-H+ exchange process has been demonstrated in these membranes. PMID- 11258547 TI - Gastrointestinal tract antigenic profile of cotton-top tamarin, Saguinus oedipus, is similar to that of humans with inflammatory bowel disease. AB - As an animal model for human inflammatory bowel disease and colorectal cancer, the cotton-top tamarin remains controversial. Demonstration of antigenic similarity to the human would enhance its validity. Using colonic extracts and washings, we compared binding of seven monoclonal antibodies reactive with bowel and cancer antigens in both tamarins and humans with inflammatory bowel disease. Additionally, telomerase activity was tested for. Expression of a mucin antigen specific to human cancer was increased in tamarin colonic washings as well as aminoproteoglycans and EGFR in tamarin extracts, as compared to those of humans with inflammatory bowel disease (P < 0.005). An adenoma-associated antigen and k ras p21 protein were negative in the tamarin. A trend to greater telomerase activity exists in tamarins. The antigenic similarity validates this model for human inflammatory bowel disease and colorectal cancer. A trend to increased telomerase activity in tamarins is consistent with the greater predisposition to cancer in these animals. PMID- 11258548 TI - Oral administration of avian tumor necrosis factor antibodies effectively treats experimental colitis in rats. AB - Tumor necrosis factor (TNF) is implicated in the pathogenesis of inflammatory bowel disease. Clinical trials indicate that intravenous infusion of anti-TNF antibody is an effective therapy for Crohn's disease. An oral anti-TNF therapy may be a preferred approach, reducing systemic side effects and eliminating the inconvenience and expense of administering infusions. We tested oral avian anti TNF antibodies in the acute and chronic phases of a rodent colitis model. Efficacy was compared to sulfasalazine and dexamethsone. Rats with chemically induced colitis were treated orally with anti-TNF antibody, placebo, or comparator. Efficacy was assessed by change in colonic weight, morphology, histology, and tissue myeloperoxidase activity. Oral anti-TNF antibody, in both the acute and chronic phases of the model, significantly decreased all inflammatory end points and proved to be more effective than sulfasalazine and dexamethasone. Oral delivery of avian anti-TNF antibodies is an effective treatment of experimental colitis and may provide advantages to current parenteral anti-TNF antibodies. PMID- 11258549 TI - Dose-effect relationship of BB-10010/MIP-1 alpha on proliferation in murine small intestinal epithelium: single and double administration protocols. AB - BB10010/MIP-1 alpha reduces the number of proliferating cells in the small intestine, strongly suggesting a radioprotective potential in this organ. This study was designed to optimize BB10010 administration for maximal radioprotection. In single administration protocols 1 or 4 mg/kg of BB10010 was injected into mice 2, 4 or 10 hr before death. In double administration protocols an initial dose of either 0.4 or 200 microg/kg, and a second dose (2.5 hr apart) of 200 microg/kg 4 hr before death were administered. The number of vincristine arrested metaphases were counted on individually microdissected crypts from the midpoint of the small intestine. When compared to the smaller doses of BB 10010 used in our previous studies, the higher doses used in these experiments did not result in any further reduction in the number of proliferating cells under any of the protocols assessed. Furthermore, some values were found to be above not only those observed with the smaller doses, but also above untreated controls. It is concluded that a single dose of 200 microg/kg of BB10010 offers the most consistent reduction of mitotic cells, and is, therefore, considered optimal for assessment of radioprotection. PMID- 11258550 TI - Administration of bacterial lipopolysaccharide to rats induces heme oxygenase-1 and formation of antioxidant bilirubin in the intestinal mucosa. AB - Heme oxygenase (HO)-1, the rate-limiting enzyme in heme degradation, is induced by oxidative stress and its major end product, bilirubin, is a potent physiological antioxidant. We studied the induction of HO-1 and bilirubin production in intestinal mucosa using a rat model of sepsis. E. coli lipopolysaccharide was administered intraperitonealy to male Wistar rats and intestinal mucosa was harvested. Intestinal lipid peroxides increased significantly at 1 hr and peaked at 170% of the control value at 5 hr. GSH significantly decreased at 3 hr, reaching the nadir of 50% of the control value at 5 hr. HO-1 mRNA was maximally induced fivefold at 3 hr and HO-1 protein maximally increased to 10 times the control value at 7.5 hr. Both bilirubin and bilirubin oxidative metabolites were maximally increased at 10 hr, to 4.3 and 3.7 times the control value, respectively. These data suggest that oxidative stress in sepsis quickly induces HO-1 in intestinal mucosa and that subsequent production of bilirubin works as an antioxidant. The small intestinal mucosa is an active participant in the general response to sepsis. PMID- 11258551 TI - Dextran sulfate sodium-induced colitis in immunodeficient rats. AB - To examine the morphology of colitis and study the role of the immune system in colitis, we compared colitis in immunocompetent Wistar-Kyoto rats with that in spontaneously hypertensive rats, known to have T-cell dysfunction. Rats were treated with 3% dextran sulfate in drinking water for periods ranging from 3 to 60 days. Diarrhea developed earlier and was associated with a more severe weight loss in Wistar-Kyoto rats than spontaneously hypertensive rats. The morphologic findings (flattening of the gland epithelium, gland dropout and ulceration) in spontaneously hypertensive rats were milder than in Wistar-Kyoto rats. Only spontaneously hypertensive rats survived 60 days of treatment; the findings included ulceration, crypt distortion, and inflammatory pseudopolyp formation. Immunostaining for B-cell, T-cell, and macrophage markers showed no difference in the distribution of these cells in the mucosa of Wistar-Kyoto rats and spontaneously hypertensive rats. Spontaneously hypertensive rats with T-cell dysfunction develop dextran sulfate sodium-induced colitis. PMID- 11258552 TI - In vitro and in vivo effects of gliotoxin, a fungal metabolite: efficacy against dextran sodium sulfate-induced colitis in rats. AB - Gliotoxin is a fungal metabolite that has immunosuppressive properties. First, we determined if gliotoxin could inhibit cytokine production from macrophage and colonic epithelial cell lines, as well as whether it inhibited nuclear factor kappa B in these same cell types. Second, we evaluated whether gliotoxin could reduce dextran sodium sulfate-induced colitis in rats. A disease activity index, myeloperoxidase activity, and cytokine levels were evaluated on either day 7 or 21. In both cell lines, gliotoxin dose dependently inhibited cytokine production and nuclear factor-kappa B. On day 21, gliotoxin significantly reduced disease activity (diarrhea and bloody stools) in rats. On day 7, gliotoxin treatment significantly improved various indices of colitis, including colonic cytokine levels. Decreased food consumption and weight gain was evident with a larger dose of gliotoxin. In summary, gliotoxin, a nuclear factor-kappa B inhibitor, effectively reduced dextran sodium sulfate-induced colitis in rats. However, gliotoxin exhibited a narrow therapeutic to toxicity ratio in these rats. PMID- 11258553 TI - Corticosteroid pretreatment prevents small intestinal mucosal lesion induced by acetic acid-perfusion model in rats. AB - One of the important problems in experimentally induced small intestinal lesions is that there is no reproducible model of diffuse and stable mucosal lesion. In this paper, we studied in detail the effects of continuous perfusion of various concentrations of acetic acid on the rat small intestinal mucosa. In order to evaluate its applicability for screening of the preventive effect of drugs on gut damage, we also evaluated the efficacy of corticosteroid pretreatment in preventing acetic acid-induced mucosal lesion. Male Sprague-Dawley rats were fasted for 12 hr, and the small intestinal lumen was perfused with 1%, 1.5%, 2.5%, 3%, 3.75% (pH 2.4-2.6) acetic acid or saline (control) at 1 ml/min for 15 min. In separate experiments, the effect of preadministration of budesonide (0.5 or 0.75 mg/kg/day) and prednisone (0.75 mg/kg/day) on 1.5% acetic acid-induced mucosal damage was investigated. Macroscopic and microscopic lesions occurred diffusely in a concentration-dependent fashion. Histological findings revealed signs of transmural inflammation characterized by mucosal-submucosal edema, ulceration, and neutrophil infiltration. Mucosal-submucosal height had an inverse relation with the acetic acid concentrations perfused. Myeloperoxidase activity levels increased several-fold in the acetic acid-perfused groups. Corticosteroid pretreatment prevented microscopic damage and was associated with reduction of MPO activity levels in 1.5% acetic acid-perfused rats. We conclude that this simple and reproducible model could be applied for the screening of new drugs in the gastrointestinal tract in which large numbers of animals are taken into account. PMID- 11258554 TI - Hyperhomocysteinemia in Greek patients with inflammatory bowel disease. AB - In recent years hyperhomocysteinemia has been established as a new risk factor for arterial and venous thrombosis. Since patients with inflammatory bowel disease (IBD) frequently suffer from thromboembolic events, we studied the prevalence and clinical significance of hyperhomocysteinemia in Greek patients with ulcerative colitis (UC) and Crohn's disease (CD). In 108 consecutive fasting IBD patients (53 UC and 55 CD) and 74 healthy controls (HC), a standard record of various clinical thrombotic risk factors was completed by interview, and fasting serum concentrations of total homocysteine (tHcy), folate, cobalamin, creatinine, cholesterol, HDL, LDL, and triglycerides were measured. The concentration (mean +/- SD) of serum tHcy was significantly higher in UC (15.9 +/- 10.3 micromol/liter) and CD patients (13.6 +/- 6.5) than in controls (9.6 +/- 3.4, P < 0.05). Both UC and CD patients had lower levels of folate than HC (P < 0.05). Covariance analysis of age, gender, and all clinical variables indicated that the differences in homocysteine levels between IBD patients and HC remain significant even after adjustment for these covariates. In conclusion, mild hyperhomocysteinemia is common in Greek IBD patients and may account for the increased thrombotic risk of these patients. PMID- 11258555 TI - Short- and long-term effect of glyceryl trinitrate (GTN) ointment 0.2% and 2% on anal canal pressure in patients with chronic anal fissures. AB - Controversial data are available on the duration of action of glyceryl trinitrate after acute and chronic application on anal canal pressure. Our aim was to assess the effect of glyceryl trinitrate at 0.2% and 2% on anal canal pressure before and after eight weeks of treatment. Anal canal pressure was evaluated in 12 patients with chronic anal fissures with an electronic probe with three recording sites before and after the application of glyceryl trinitrate, 120 mg on the external anal verge. Six patients received glyceryl trinitrate at 0.2% and six at 2%. Glyceryl trinitrate 0.2% and 2% equally reduce basal anal canal pressure in all three recording sites (P < 0.001) with major effect on the inner site of the canal toward the rectum, for a 60-min period. Eight weeks after application, the effect of glyceryl trinitrate was unchanged. In conclusion, glyceryl trinitrate ointment at 0.2% and 2%, equally reduces anal canal pressure for 60 min and this effect is kept unchanged after eight weeks of application. PMID- 11258556 TI - Coincidental malabsorption of lactose, fructose, and sorbitol ingested at low doses is not common in normal adults. AB - Normal subjects may incompletely absorb either lactose, fructose, or sorbitol and may therefore have abdominal symptoms. The frequency of coincidental malabsorption of these sugars is not known. This is clinically important, since we often ingest them during the same day and malabsorption may cause abdominal symptoms. To shed light on this issue we studied 32 normal subjects. Volunteers drank in random order the following solutions: 20 g lactulose, 50 g sucrose, 50 and 25 g lactose, 50 and 25 g fructose, 20 and 10 g sorbitol. Semiquantitative carbohydrate malabsorption was estimated with lactulose standards. Frequency of 50-g lactose (69%), 50-g fructose (81%), and 20-g sorbitol (84%) malabsorption was not significantly different (P = 0.3). The estimated median fraction of the ingested high dose malabsorbed was 42, 19, and 68% for lactose, fructose, and sorbitol, respectively. At low challenging doses, 63% of the volunteers absorbed two of three or all three sugars, and 88% were asymptomatic to two or all three sugars. In conclusion, the frequency of coincidental malabsorption of lactose, fructose, and sorbitol and intolerance to these sugars is not common, when normal adults ingest them at low doses. PMID- 11258557 TI - Giant cell arteritis and intestinal angina. PMID- 11258558 TI - Urachal hernia: an unusual intra-abdominal hernia caused by incarceration into a urachal cyst. PMID- 11258559 TI - Symptomatic gastroesophageal reflux as a risk factor for esophageal adenocarcinoma. AB - A population-based, case-control study to investigate the possible association between gastroesophageal reflux (GER) and adenocarcinoma of the esophagus and gastric cardia was performed. It demonstrated an odds ratio of 7.7 for esophageal adenocarcinoma in patients with GER symptoms. The frequency, severity, and duration of symptoms correlated with an increased risk of esophageal adenocarcinoma. A weaker association was noted for GER and adenocarcinoma of the gastric cardia. No association surfaced between GER and squamous cell carcinoma of the esophagus. PMID- 11258560 TI - Hepatocellular carcinoma presenting with chronic inflammatory demyelinating polyradiculoneuropathy. PMID- 11258561 TI - Electrophysiological differences in normal colon mucosa from diverticular disease vs cancer. PMID- 11258562 TI - Differential effect of prostaglandins on gallstone-free and gallstone-containing human gallbladder. AB - Nonsteroidal antiinflammatory drugs, inhibitors of prostaglandin synthesis, have different effects on gallbladder contractility in normal and diseased human gallbladders in vivo. We investigated this differential effect by comparing the effects of prostaglandins PGE2 and PGF2alpha, the thromboxane A2 mimetic U46619, and PGI2 on in vitro contractility in gallstone-free and gallstone-containing human gallbladders. Isometric tension was measured in gallbladder muscle strips mounted in organ baths. EC50 was calculated for each agonist. The rank order of potency in gallstone-free gallbladders was PGE2 > CCK > U46619 > PGF2alpha and in gallstone-containing gallbladders was U46619 > PGE2 > CCK > PGF2alpha. PGI2 produced contraction of gallstone-free gallbladder and relaxation of gallstone containing gallbladder in the basal state. Further, PGI2 produced no relaxation in gallstone-free muscle strips precontracted with CCK, but significant relaxation in CCK precontracted gallstone-containing strips. PGE2, PGF2alpha, and U46619 are potent contractors of gallstone-free and gallstone-containing gallbladders, whereas PGI2 relaxes only gallstone-containing gallbladders. Since gallbladders containing cholesterol-supersaturated bile produce increased PGI2, this PGI2-induced relaxation may be a determinant of the impaired gallbladder motility of gallstone disease. PMID- 11258563 TI - Gallbladder dysfunction enhances physical density but not biochemical metastability of biliary vesicles. AB - The gallbladder role in cholesterol gallstone pathogenesis occurs through modulation of bile cholesterol metastability. The present study characterized the effects of concentrating bile on cholesterol crystallization through vesicle transformation, crystal habits, and potentiation of effector substances. Supersaturated model biles with total lipid concentrations of 12, 9, 6, and 3 g/dl were prepared with identical molar ratios (taurocholate-egg yolk phosphatidylcholine-cholesterol: 71:18:11). Bile metastability was assessed spectrophotometrically, and morphology of vesicle and crystal was sequentially scanned by video-enhanced differential contrast microscopy. The effects of replacing 30% of egg yolk phosphatidylcholine with soy bean phosphatidylcholine, 30% of taurocholate with taurodeoxycholate or tauroursodeoxycholate, and addition of concanavalin A-binding glycoprotein on each model bile were examined. By lowering total lipid concentration, cholesterol crystallization was retarded with less fusion and aggregation of vesicles. The effects of substances promoting cholesterol crystallization were enhanced with lesser bile. By replacing 30% of taurocholate with tauroursodeoxycholate, cholesterol crystallization was markedly inhibited in all concentrations, forming stable liquid-crystals. Impaired water absorption by the gallbladder may stabilize vesicles and inhibit rapid cholesterol crystallization, but the potential of cholesterol crystallization effector substances must be modified to alter bile cholesterol metastability. PMID- 11258564 TI - Epidemiology of gallstone disease in Argentina: prevalences in the general population and European descendants. AB - To assess gallstone disease prevalence in Argentina, a random sample of the Rosario City population was studied, considering already known associated factors, and analyzing ethnic groups living in the city. A total of 1,173 participants (69% response), both sexes, 20 years and older were studied. Each subject underwent an abdominal ultrasound examination, a blood test, and a standardized questionnaire. It was seen that gallstone disease prevalence (gallstones or cholecystectomy), overall was 20.5% (23.8% in women and 15.5% in men; (P = 0.0005), and was associated with age and body mass index in both sexes, and with pregnancy number and hypertriglyceridemia in women. As regards ancestors' nationalities, Italian and Spanish descendants presented higher prevalence rates for all age groups than those described in Italy and Spain. Thus far, in a subsample of 78% of nonparticipants submitted to a new home visit, presence of cholecystectomy or symptoms did not differ from participants, supporting the validity of our results. PMID- 11258565 TI - Herpes simplex-induced fulminant hepatitis in adults: a call for empiric therapy. AB - Herpes simplex-induced fulminant hepatitis is an infrequently reported cause of hepatitis in adults. Pregnant females and patients with impaired cellular immunity may be at increased risk, although healthy adults have been affected. The diagnosis may be underrecognized due to nonspecific presenting symptoms and lack of typical cutaneous herpes lesions. We present three cases of fatal herpes simplex fulminant hepatitis. Our review of case reports of herpes simplex hepatitis in adults demonstrates improved survival with intravenous acyclovir therapy. We believe that empiric use of acyclovir should be considered while the diagnostic evaluation of non-acetaminophen-induced fulminant hepatitis is underway. Recognition of characteristic liver function abnormalities seen with fulminant herpes simplex hepatitis include marked elevation of transaminases with AST > ALT and a mild hyperbilirubinemia (anicteric hepatitis), and they should prompt acyclovir therapy. This is especially true when there are no obvious risk factors for other forms of hepatitis. PMID- 11258566 TI - Pulmonary expression of iNOS and HO-1 protein is upregulated in a rat model of prehepatic portal hypertension. AB - Portal hypertension is associated with a wide range of pulmonary pathophysiologies, ranging from portopulmonary hypertension to hepatopulmonary syndrome. Although the clinical and pathological features of pulmonary dysfunction in this setting have been extensively characterized, the underlying biology is not well understood. Specifically, the role of mediators that regulate mesenteric vascular hemodynamics in portal hypertension, such as nitric oxide and endothelin, have not been studied in the lung. Using a rat model of prehepatic portal hypertension with preserved hepatic function, we examined pulmonary elaboration of endothelial nitric oxide synthase (NOS), inducible NOS, heme oxygenase- 1 (HO-1), heme oxygenase-2 (HO-2), endothelin-1 mRNA, and protein. In comparison to sham controls, portal hypertensive animals exhibited significantly increased pulmonary iNOS and HO-1 mRNA and protein. Cyclic GMP was significantly increased in portal hypertensive lung tissue, suggesting activation of guanylyl cyclase by the endproducts of iNOS and/or HO-1 activity. Using immunohistochemical analysis, iNOS expression was localized to the vascular endothelium, while HO-1 localized to bronchiolar epithelium and macrophages. These results suggest that production of nitric oxide and carbon monoxide may contribute to the pulmonary pathology associated with portal hypertension. PMID- 11258567 TI - Improvement of hepatic protoporphyrin accumulation after antibiotic treatment. PMID- 11258568 TI - High viral eradication with a daily 12-week natural interferon-beta treatment regimen in chronic hepatitis C patients with low viral load. IFN-beta Research Group. AB - Virological sustained response (SR) is achieved in 31-49% of patients with chronic hepatitis C with combination therapy using interferon (IFN)-alpha and ribavirin for 24-48 weeks. However, as a period of 24-48 weeks is a burden for patients, we investigated the effect of daily intravenous administration of natural IFN-beta for 12 weeks in this study. In all, 112 patients were enrolled and received a daily administration of 6 MU of natural IFN-beta intravenously for 12 weeks. Serum HCV-RNA before treatment was assessed by the competitive reverse transcription polymerase chain reaction assay. The patients were divided into two groups according to pretreatment viral load: the low viral load group (N = 25, <6.3 x 10(5) copies/ml), and the high viral load group (N = 87, > or =6.3 x 10(5) copies/ml) who were additionally administered IFN-beta thrice weekly for subsequent 14 weeks at the patients' request. Virological SR was obtained in 37% (41/112) of all the patients; 88% of those with a low viral load, and 22% of patients with a high viral load. Virological SR was attained in 21% of patients with HCV subtype 1, and in 67% of those with subtype 2a. In patients with HCV subtype 1b, virological SR was obtained in patients with the mutant-type (> or =4 amino acid mutations in the NSSA2209-48) having a low viral load (4/4), but not in those having a high viral load (0/3). The results suggest that a daily intravenous administration of natural IFN-beta for 12 weeks showed high therapeutic efficacy in patients with a low viral load despite the shorter treatment period and that the therapeutic efficacy depends on viral load rather than on the number of NS5A2209-48 amino acid mutations. PMID- 11258569 TI - PCR-SSCP analysis of 5'-nontranslated region of hepatitis A viral RNA: comparison with clinicopathological features of hepatitis A. AB - The recent development of the sensitive reverse transcription-polymerase chain reaction (RT-PCR) method allowed us to detect the presence of hepatitis A virus (HAV) RNA in sera from hepatitis A patients. To determine whether differences in HAV are related to the wide range of clinical severity, we used PCR-single strand conformation polymorphism (SSCP) to analyze the amplified product of the 5' nontranslated region (5'NTR), where relatively homologous sequences are reported and the internal ribosomal entry site is considered to exist, from these various levels of hepatitis A. Twenty-seven patients admitted to Chiba University Hospital (between 1988 and 1997) were examined for HAV RNA in their sera by RT PCR with primers located at 5'NTR of HAV RNA. The nucleotide (nt) sequence of a central part (nt 277-551) of 5'NTR was amplified and the product was examined by PCR-SSCP. HAV RNA was detected in all 27 hepatitis A patients examined: in 3 with fulminant hepatitis, 2 with severe acute hepatitis, and 22 with self-limited acute hepatitits. The amplified sequence revealed by PCR-SSCP showed that samples from fulminant and severe acute hepatitis patients had similar mobility in the gel, whereas that from acute hepatitis patients demonstrated a considerable variety in mobility patterns. From the analysis of the amplified product of 5'NTR of HAV, the bands are likely to be similar in the more severe forms of hepatitis A compared to those from uncomplicated self-limited acute hepatitis. PMID- 11258570 TI - HCV RNA values. PMID- 11258571 TI - Reduced interleukin-12, interleukin-18, and interferon-gamma production with prolonged rat hepatic allograft survival after donor-specific blood transfusion. AB - Interferon-gamma is a key immunoregulatory cytokine involved in acute graft rejection. Immunologic unresponsiveness to organ allografts has been induced by pretransplantation donor-specific blood transfusion, both experimentally and clinically. We investigated interferon-gamma production and intragraft gene expression of type-1 T-helper cytokines such as interleukin-12 and -18 and type-2 T-helper cytokines such as interleukin-10 and transforming growth factor-beta in rats receiving hepatic allografts after such transfusions. The animals were divided into four groups: group I received isografts; group II received allografts; group III received allografts after donor-specific transfusion; and group IV received allografts and was treated with FK 506. Donor blood given seven days prior to transplantation significantly prolonged allograft survival. The serum interferon-gamma concentrations in group II increased, peaking on day 5 and then decreasing. Serum interferon-gamma concentrations in groups I, III, and IV were significantly lower than those observed in group II, as were levels of interleukin-12 and interleukin-18 mRNA in the graft. Transforming growth factor beta and interleukin-10 mRNA levels in grafts in transfused animals were significantly greater than those in the untreated allograft group. Interleukin-12 and -18 mRNA transcripts in an allogeneic mixed lymphocyte reaction were inhibited by interleukin-10 and transforming growth factor-beta. These results suggest that interleukin-12 and -18 expression in hepatic allografts is inhibited in the immunologically unresponsive state induced by donor-specific transfusion. PMID- 11258572 TI - Upregulated cyclooxygenase-2 inhibits apoptosis of human gastric epithelial cells infected with Helicobacter pylori. AB - Helicobacter pylori induces apoptosis and alters the proliferation of gastric mucosal epithelial cells. Cyclooxygenase-2 (COX-2), the inducible form of prostaglandin (PG) synthesis, is known to cause alteration in epithelial cell growth. The goal of this study was to determine whether COX-2 gene expression by H. pylori infection could influence gastric epithelial cell apoptosis. Expression of COX-2 mRNA and proteins was up-regulated in Hs746T gastric epithelial cell lines infected with H. pylori, when assessed by quantitative RT-PCR and western blot. Inhibition of COX-2 expression using NS-398, a specific COX-2 inhibitor, showed a significant increase of gastric epithelial cell apoptosis and caspase-3 activation in Hs746T cells infected with H. pylori. Moreover, the effect of NS 398 on H. pylori-induced apoptosis was reversed by the addition of PGE2. These results suggest that up-regulated COX-2 expression by H. pylori infection can inhibit apoptosis of gastric epithelial cells. PMID- 11258573 TI - Specific Helicobacter pylori antigens unable to distinguish nonucler dyspepsia or peptic ulcer cases from asymptomatic seropositive controls: a nested case-control study in employees of a large company. AB - We investigated in a large industrial population the antibody response to specific H. pylori antigens (CagA and seven others) in relation to peptic ulcer and nonulcer dyspepsia (NUD). The two groups consisted of 37 and 39 employees, respectively, with endoscopically proven peptic ulcer and NUD. Age- and gender matched controls were H. pylori seropositive employees without abdominal complaints or history of ulcer disease. IgG antibodies against CagA and other antigens were analyzed by western immunoblot. Relative percentages of CagA positive individuals were 89 and 76% for ulcer cases and their controls (P = 0.22) and 77% and 74% for NUD cases and their controls. The corresponding percentages of VacA-positive individuals were 87 and 76% for ulcer cases and controls and 64% and 77% for NUD cases and controls, respectively. Analysis of other H. pylori-specific antigens was not particularly helpful in discriminating between symptomatic and asymptomatic seropositive individuals for either disease group. In conclusion, assessment of IgG response against specific H. pylori antigens was not predictive of peptic ulcer or NUD case status in this active employee population and would not appear to be useful in routine clinical practice. PMID- 11258574 TI - Cardioesophageal reflexes: an invasive human study. AB - Neural reflex arcs from the esophagus and heart have been shown in both animals and man. The purpose of this study was to further investigate these pathways in individuals undergoing cardiac catheterization. A total of 298 patients undergoing cardiac catheterization were asked to participate in the protocol. Thirty patients were able to complete the study. Esophageal manometry and pH were monitored throughout the cardiac procedure. Afterwards, esophageal provocation with ice water, hydrochloric acid, and balloon inflation was performed with observation of cardiac rate and rhythm. Twelve patients with normal coronary arteries developed diffuse esophageal spasm on either esophageal or cardiac provocation. In one patient with abnormal coronary arteries, coronary angioplasty precipitated diffuse esophageal spasm. Esophageal acid sensitivity was increased in patients with normal coronaries as compared to those that were abnormal. The esophageal pain threshold was significantly lower in patients undergoing angioplasty versus those undergoing coronary angiography alone. There was no significant change in esophageal pH during invasive cardiac maneuvers and manipulations. In conclusion, cardiac manipulation can induce esophageal motility abnormalities, but not gastroesophageal reflux. Coronary angioplasty is associated with esophageal hyperalgesia. PMID- 11258575 TI - Expression of immediate early genes, HSP70, and COX-2 mRNAs in rat stomach following ethanol ingestion. AB - The aim of this study was to show the temporal and spatial molecular responses in the rat stomach that follow absolute ethanol-induced acute mucosal injury. Intense signals for immediate early genes (IEG)/transcriptional factors such as c fos, c-jun, and nerve growth factor-induced gene-A (NGFI-A) mRNAs were observed in the superficial mucosa and in the blood vessels from 15 min to 6 hr after administration, peaking at 15-30 min. Signals for heat shock protein (HSP) 70 mRNA were also detected in the superficial mucosa, in the fibroblasts around gastric erosions, and in the blood vessels from 15 min to 6 hr (peak at 1-2 hr). The signals for cyclooxygenase-2 (COX-2) mRNA were up-regulated in the surface mucous cells that surround the erosions from 30 min to 6 hr (peak at 60-90 min). These findings suggest that IEG, HSP70, and COX-2 are involved in gastric mucosal restitution in different ways. PMID- 11258577 TI - Implications of mechanical stretch on wound repair of gastric smooth muscle cells in vitro. AB - Gastric smooth muscle cells continually receive repetitive physical stretching by food storage, peristalsis and fasting contraction; therefore mechanical stretch can not be disregarded in gastric events. The aim of this study was to clarify the effects of mechanical stretch on wound repair using a rabbit gastric smooth muscle cell sheet. Mechanical stretch was imposed on adherent cells on a flexible membrane in order to increase elongation by an average of 5% and 10%, respectively, at 5 cycles per minute after artificial wounding. Adherent cells not subjected to mechanical stretch served as controls. The restoration process was monitored by measuring wound size for 48 h. Proliferation was assessed by BrdU staining and the influence on the cytoskeletal system was assessed by actin staining. The speed of restoration was highest in controls and lowest in the 10% stretch groups. Proliferation was almost equal to that of controls in the stretch groups. Under the condition of mechanical stretch, stress fibers appeared weakened and the direction of fibers was not consistent but random. In conclusion, mechanical stretch inhibited the migration of gastric smooth muscle cells, leading to cytoskeletal dysfunction. It is suggested that physical stretch, such as mechanical stretch, might be an important factor in the process of gastric wound repair. PMID- 11258576 TI - Analysis of ambulatory duodenogastroesophageal reflux monitoring. AB - Some methodological in vitro observations concerning bile reflux monitoring (Bilitec) suggested that Bilitec monitoring is underestimating reflux in an acid environment. Moreover, other studies showed that the area above the cutoff level of bilirubin absorbance would provide an adequate quantitative marker for reflux of duodenal contents. Our aim was to study whether correction for intraesophageal acidity and the area above cutoff during Bilitec monitoring affects the results and the correlation with pH measurement and esophageal lesions. In 84 patients (46 men; mean age 46 +/- 2.7 years) evaluated for suspected gastroesophageal reflux disease, we performed ambulatory 24-hr esophageal pH and Bilitec monitoring after an upper gastrointestinal endoscopy. We obtained total area, percent total time, and correction by computer software. The correction factor for bilirubin absorbance was based on literature data for acidified bile (0.06 for pH < 3.6; 0.21 for pH < 2.6). Endoscopy revealed esophagitis grade 1-2 (E1-2) and 3-4 (E3-4) in 23 and 16 patients, respectively. A progressive increase of mixed (acid + bile) reflux occurred with increasing severity of endoscopic lesions (E3-4 vs no esophagitis, P < 0.05). A pathologic Bilitec monitoring result was present in the same 35 patients before and after correction and the correlation between the pH measurement and percent time of bile reflux was not improved by correction for intraesophageal pH (r = 0.386 and r = 0.391; P < 0.05). The total area of bilirubin absorbance above 0.14 (abs x min) was 7.8 +/- 2.2 in patients without esophagitis, and 11.7 +/- 4.4 and 17.0 +/- 4.2 in the E1 2 and E3-4 groups, respectively (E3-4 vs no esophagitis, P < 0.05). The correlation between the Bilitec monitoring and pH measurement regarding percent (r = 0.427, P < 0.01) or area of time below 4 (r = 0.280, P < 0.05) was not improved by considering the area of bilirubin absorbance above the cutoff level. Correction for intraesophageal pH has only a minor effect on the results of ambulatory Bilitec monitoring. Taking into account the surface rather than the percent of time above the cutoff level for bilirubin absorbance does not improve the correlation of Bilitec with acid reflux and with esophageal lesions. PMID- 11258578 TI - A system for recording of auditory evoked responses. AB - A system for recording of evoked potentials from auditory stimulation was developed. The system consists of a PC equipped with an audio bandwidth board with analog input and output channels. The sound stimulus signal is generated in the computer, D/A converted, and via audio amplifier fed to earphones on the test subject. Auditory evoked potentials in response to sound stimuli are recorded via electrodes, amplified and filtered in an EEG recording system and fed to an A/D converter. The signal is analysed in the PC. The modular design of the program makes it a flexible system where stimulus and recording parameters can easily be modified and new applications can be added to standard clinical measurements. Three applications that are not possible with commercially available systems were developed and evaluated. a) A diagnostic procedure to verify hydrops in patients with Meniere's disease. b) Intraoperative recordings of auditory evoked potentials during neurootological surgery. c) Recording of mismatch negativity (MMN) potentials in evaluation of central auditory functions. PMID- 11258579 TI - Hemodynamic evaluation with TURBO BRISK--a rapid phase contrast angiography technique. AB - Hemodynamic imaging by phase contrast angiography was significantly accelerated by selective interpolation and segmentation in k-space using TURBO BRISK. The method was tested in vitro on three independent flowfields, representative of human blood rheology: a straight tube simulating the descending aorta, a curved tube simulating the aortic arch and a two-chamber orifice flow model simulating valvular regurgitation. The results were compared to data obtained by Laser Doppler Velocimetry (LDV) and showed good agreement. For the straight tube, the flow velocity obtained by five TURBO BRISK methods with increasing segmentation factors and corresponding time savings showed good agreement with LDV. For the curved tube, the velocity showed good general agreement with some differences in the decelerating part of the cycle, and in the low-velocity secondary flow structures. The orifice flow evaluation, the most time consuming case, was performed by the control volume method. It showed good agreement with actual flows through the orifice. Data acquisitions for TURBO-4 BRISK could be performed in 20s for each velocity component. The method shows promise for breath-hold acquisitions in clinical applications, including calculation of blood flow volumes through diseased arteries, measurement of blood backflow volumes through dysfunctional heart valves to time valve replacement operations, and evaluation of arterial wall shear stress, an important factor in the genesis of atherosclerosis. PMID- 11258580 TI - Fixation of the acetabular cup in cemented total hip replacement: improving the anchorage hole profile using finite element method. AB - Long-term studies have shown that failure of the acetabular component in total hip replacement increases exponentially ten years following surgery and occurs mostly at the bone-cement interface. During the cemented fixation of the acetabular cup, straight anchorage holes, 3-15 mm diameter and 3-20 mm deep, are drilled in the acetabulum in order to increase torsional resistance at the bone cement interface. The aim of this paper is to provide guidelines for improving the profile of anchorage holes. Results from our finite element models show that the efficiency of anchorage holes may be improved if they are drilled perpendicularly to the acetabulum floor and if they have chamfered necks. A 10 degree inclination of the anchorage hole increases Von Mises stress in the cement mantle by 6% while creating chamfered anchorage holes, instead of straight holes, decreases it by 14%. Increasing depth of anchorage holes does not improve efficiency. PMID- 11258581 TI - Foreign body sarcoma: effects of foreign DNA, beta-carotene and paprika applied to the implant surface. AB - Sarcoma arises extremely rarely on foreign bodies in man, but is aggressive and often lethal. A coating for implants which would further reduce the risk in man is desirable. The incidence in mice is much greater, and responds to chemical treatment of the implant surface. Coating with histones increases tumour yield. Accordingly, related substances, foreign DNA, DNase and a mixture of the two, were tested for anticancer activity by application to 25 mm nitrocellulose filters in groups of 30-45 BALB/c mice, in comparison with untreated filters. Other substances reported to influence neoplasia, paprika, beta-carotene, rhodamine and tuftsin; and substances expected to be neutral, oxyprenolol, liquid paraffin, iodine, and adenosine diphosphate were similarly tested against concurrent untreated controls for comparison. Bovine DNA (p = 0.01) and DNA/DNase mixture (p = 0.04) and DNase fomented tumour growth by 55, 45 and 59% respectively. Paprika and beta-carotene did so by 70% (p = 0.05). The other substances were inert. None were candidates for an anti-sarcoma coating. PMID- 11258582 TI - Cardiac arrhythmia classification using neural networks. AB - In this paper, ECG arrhythmia classification using principal component analysis is proposed. Hebbian neural networks are used for computing the principal components of an ECG signal. This provides an unsupervised feature extraction, dimension reduction and an improved computing efficiency. Results from 14 pathological records obtained from the MIT ECG database demonstrate the capability of this method in differentiating between five different types of arrhythmia despite the variations in signal morphology. An average value for classification sensitivity and positive predictivity were found to be Se% = 98.1% and +P% = 94.7% respectively. PMID- 11258583 TI - Problems and solutions for artificial kidney. AB - The uremic syndrome is the prototype of a slowly progressive endogenous intoxication, when a detoxifying organ (in this case the kidney) fails. It is characterized by the gradual retention of a host of metabolites, which is in part corrected by dialysis, allowing survival with an acceptable quality of life. This paper reviews the main problems of hemodialysis today, and possible solutions. Adequacy of dialysis is estimated currently from the concentration of urea, which is used as a marker molecule. The problem is that urea is not really toxic by itself. Other markers with known toxicity, such as middle molecules (300-12,000 D) and protein bound compounds should be considered. The question then arises whether the classical dialytic concept based on diffusion should be modified. Adsorptive systems may be strong binders of protein bound solutes. Other concepts that are now arising, and that may add to toxin removal, are slow and daily dialysis. Another question that could be raised is whether it would not be possible to support toxin removal, by administering peroral sorbants. Dialysis patients are prone to vascular disease and die early from cardio-vascular complications. One of the solutions for this problem could be to bring the blood of dialyzed patients into contact with antioxidants (e.g. vitamin C or E). The risk for perdialytic hemodynamic instability is increased in many dialysis patients. The ideal solution would be to develop an "intelligent" dialysis system, whereby blood volume and plasma osmolality are sensed continuously, and ultrafiltration and dialysate sodium concentration are adapted in function of this evolution. An adequate vascular access is indispensable to perform adequate dialysis, but thrombotic/stenotic complications are frequent. This could be prevented by molecular biological modification of vascular grafts, whereby genetic information is entered into the cells, blocking the natural chain of events that otherwise unavoidably leads to neointimal hyperplasia and atherosclerosis. Another old dream is to develop a wearable artificial kidney, whereby patients can move around, and be treated 24 hours per 24 hours, in stead of being treated intermittently at a specific location by the dialysis machine. According to some authors, part of the natural renal function could be replaced by cultured renal tubular cells, which are brought in contact with the blood of the patients. It is concluded that thrilling improvements lie ahead in the future, but the following questions arise: 1) What is the cost of all these improvements? 2) Will it remain possible to reimburse all this? 3) What is going to happen in transplantation, mainly regarding improvements in immunosuppression and the development of xenotransplantation? PMID- 11258584 TI - Oxygen saturation increases during childhood and decreases during adulthood among high altitude native Tibetians residing at 3,800-4,200m. AB - This report describes age differences in oxygen saturation throughout the life cycle in a sample of high altitude native Tibetans residing in villages at 3,800 4,200 m altitude in the Tibet Autonomous Region, China. Oxygen saturation of 3,812 Tibetans was measured by pulse oximetry and a subsample of 1,582 healthy, nonpregnant, nonsmokers from 1 week to 80 years of age was selected for analyses. Infants under 1 year of age had 5-6% lower oxygen saturation than the peak of 89.8% attained at 11 years of age. There was a steady increase in mean oxygen saturation-for-age during the first decade of life, but not during the second decade. Adult males exhibited a slight decrease starting in the 20-29 year age range. Adult females maintained the peak oxygen saturation through the 40-49 year age range, exhibiting a decrease in oxygen saturation beginning in the 50-59 year age range and as a result had higher oxygen saturation than males during the female reproductive span. Thus, developmental factors during infancy and childhood, but not adolescence, enhanced oxygen transfer in this high altitude native resident Tibetan sample. The age of onset of aging processes detrimental to oxygen transfer differed for females and males. PMID- 11258585 TI - Cerebral blood flow and oxygen delivery at high altitude. AB - During acclimatization to moderate altitudes, a simple calculation from data of others shows that the rise in cerebral blood flow (CBF) is sufficient that oxygen delivery to brain (DaO2) is constant as arterial oxygen content (CaO2) falls. This balance occurs on average even though the hypocapnia caused by hypoxic hyperventilation causes cerebral vasoconstriction, conflicting with hypoxic cerebral vasodilation. The relative strengths of the ventilatory and cerebral vascular sensitivities may affect this balance in individual subjects. There is no evidence for a mechanism to detect or respond directly to DaO2. Hypoxic cerebral vasodilation is believed to depend upon tissue and capillary PO2 and content, not arterial. Despite these reservations it is of interest that the average resultant DO2 remains constant. I speculate here that this match may relate to the well-known local hyperemic response to neuronal activity which now has been shown to initially overcompensate, in that tissue PO2 and pH rise in the first few seconds after neural activity. Analysis of results from the paper by Severinghaus et al. (Circ. Res. 1966;19:274-282) shows that in their subjects, despite approximately 20% reductions in arterial oxygen content at 3,810 m altitude, the data does not show any significant fall in DaO2 as a result of increased cerebral blood flows. PMID- 11258586 TI - Does hypoxia impair ovarian function in Bolivian women indigenous to high altitude? AB - Fertility appears to be reduced in at least some high altitude populations relative to their counterparts at lower elevations. Inferring from the difficulties with reproduction of newcomers to high altitude and from animal experiments, it has been hypothesized that this apparent reduction is the result of hypoxia acting to reduce fecundity and/or increase fetal loss. In humans, however, several behavioral as well as biological factors may affect fertility levels. These many factors have been organized by demographers into a framework of seven proximate determinants that includes fecundability (the monthly probability of conception) of which successful ovulation is one component. To test whether ovarian function is impaired in women indigenous to high altitude, we measured salivary progesterone (P) in a sample (n = 20) of Quechua women (aged 19-42 years) residing at 3,100 m. It was found that mean luteal P = 179 pmol/L and mean midluteal P = 243 pmol/L, levels that fall about midway in the range of known values for several populations and are higher than some lower altitude populations. These findings suggest that hypoxia does not appear to significantly impair ovarian function in those with lifelong residence at high altitude. There are, however, several factors common to many high altitude populations that may act to reduce fecundability and fertility including intercourse patterns (affected by marriage and migration practices), prolonged lactation, dietary insufficiency, and hard labor. PMID- 11258587 TI - Six percent oxygen enrichment of room air at simulated 5,000 m altitude improves neuropsychological function. AB - Cognitive and motor function are known to deteriorate with the hypoxia accompanying high altitude, posing a substantial challenge to the efficient operation of high altitude industrial and scientific projects. To evaluate the effectiveness of enriching room air oxygen by 6% at 5,000 m altitude in ameliorating such deficits, 24 unacclimatized subjects (16 males, 8 females; mean age 37.8, range 20 to 47) underwent neuropsychological testing in a specially designed facility at 3,800 m that can simulate an ambient 5,000 m atmosphere and 6% enrichment at 5,000 m. Each subject was tested in both conditions in a randomized, double-blinded fashion. The 2-h test battery of 16 tasks assessed various aspects of motor and cognitive performance. Compared with simulated breathing air at 5,000 m, oxygen enrichment resulted in higher arterial oxygen saturations (93.0 vs. 81.6%), quicker reaction times, improved hand-eye coordination, and more positive sense of well-being (on 6 of 16 scales), each significant at the p < 0.05 level. Other aspects of neuropsychological function were not significantly improved by 6% additional oxygen. PMID- 11258588 TI - Altered structure and function of the carotid body at high altitude and associated chemoreflexes. AB - The ventilatory response to hypoxia is complex. First contact with hypoxia causes an increase in ventilation within seconds that reaches full intensity within minutes because of an increase in carotid sinus nerve (CSN) input to the brain stem. With continued exposure, ventilation increases further over days (ventilatory acclimatization). Initially, it was hypothesized that ventilatory acclimatization arose from a central nervous system (CNS) mechanism. Compensation for alkalosis in the brain and restoration of pH in the vicinity of central chemoreceptors was believed to cause the secondary increase in ventilation. However, when this hypothesis could not be substantiated, attention was turned to the peripheral chemoreceptors. With the lowering of arterial PO2 at high altitude, there is an immediate increase in firing of afferents from chemoreceptors in the carotid body. After peaking over the next few minutes, the firing rate of afferents begins to rise again within hours until a steady state is reached. This secondary increase occurs along with increase in neurotransmitter synthesis and release and altered gene expression followed by hypertrophy of carotid body glomus cells. Further exposure to hypoxia eventually leads to blunting of the CSN output and ventilatory response in some species. This mini review is about the altered structure and function of the carotid body at high altitude and the associated blunting of the chemoreceptor and ventilatory responses observed in some species. PMID- 11258589 TI - Scientific activities of Academician Nikolai Sirotinin. PMID- 11258590 TI - Elevated plasma cholecystokinin at high altitude: metabolic implications for the anorexia of acute mountain sickness. AB - The aims of the present study were to measure the satiety neuropeptide cholecystokinin (CCK) in humans at terrestrial high altitude to investigate its possible role in the pathophysiology of anorexia, cachexia, and acute mountain sickness (AMS). Nineteen male mountaineers aged 38 +/- 12 years participated in a 20 +/- 5 day trek to Mt. Kanchenjunga basecamp (BC) located at 5,100 m, where they remained for 7 +/- 5 days. Subjects were examined at rest and during a maximal exercise test at sea-level before/after the expedition (SL1/SL2) and during the BC sojourn. There was a mild increase in Lake Louise AMS score from 1.1 +/- 1.2 points at SL1 to 2.3 +/- 2.3 points by the end of the first day at BC (P < 0.05). A marked increase in resting plasma CCK was observed on the morning of the second day at BC relative to sea-level control values (62.9 +/- 42.2 pmol/L(-1) vs. SL1: 4.3 +/- 8.3 pmol/L(-1), P < 0.05 vs. SL2: 26.5 +/- 25.2 pmol/L(-1), P < 0.05). Maximal exercise increased CCK by 78.5 +/- 24.8 pmol/L( 1), (P < 0.05 vs. resting value) during the SL1 test and increased the plasma concentration of non-esterified fatty acids and glycerol at BC (P < 0.05 vs. SL1/SL2). The CCK response was not different in five subjects who presented with anorexia on Day 2 compared with those with a normal appetite. While there was no relationship between the increase in CCK and AMS score at BC, a more pronounced increase in resting CCK was observed in subjects with AMS (> or =3 points at the end of Day 1 at BC) compared with those without (+98.9 +/- 1.4 pmol/L(-1) vs. +67.6 +/- 37.2 pmol/L(-1), P < 0.05). Caloric intake remained remarkably low during the stay at BC (8.9 +/- 1.4 MJ.d(-1)) despite a progressive decrease in total body mass (-4.5 +/- 2.1 kg after 31 +/- 13 h at BC, P < 0.05 vs. SL1/SL2), which appeared to be due to a selective loss of torso adipose tissue. These findings suggest that the satiogenic effects of CCK may have contributed to the observed caloric deficit and subsequent cachexia at high altitude despite adequate availability of palatable foods. The metabolic implications of elevated CCK in AMS remain to be elucidated. PMID- 11258591 TI - Preparing for the revolution--pharmacogenomics and the clinical lab. AB - Pharmacogenomics seeks to apply the field of genomics to improve the efficacy and safety of therapeutics. Sinmply put, pharmacogenomics is genetic-based testing to determine patient therapy. Interestingly, the clinical lab has rarely been discussed within the context of pharmacogenomics. Since clinical labs fill a key role in drug development it is important that they are included in pharmacogenomic discussions. Currently, clinical labs assist pharmaceutical sponsors in preclinical pharmacogenetic testing. In the future, clinical labs will be looked to for genetic test development and validation, and high throughput genotyping of patients in clinical trials and routine testing. Clinical labs are an essential link in the chain of pharmacogenomic drug development, a fact which must be recognised by both the labs themselves and the industry as a whole. PMID- 11258592 TI - Suppression subtractive hybridisation: application in the discovery of novel pharmacological targets. AB - Suppression subtractive hybridisation (SSH) is a recently developed technology designed for identifying differentially expressed genes. Total cellular RNA isolated from two sources of tissue (such as diseased versus normal) is reverse transcribed to generate cDNA and subtracted. One critical feature of this technique is the suppression polymerase chain reaction (PCR) in combination with subtraction. To optimnise the subtraction the cDNA is digested with a restriction enzyme to generate small DNA fragments (approximately 500 nucleotides in length). Specially designed DNA adapters are ligated onto one of the cDNA pools (usually the diseased one), which is followed by two rounds of hybridisation and PCR. Since this method is PCR-based, it is sensitive and efficient to identify genes of interest. The discovery of disease-related genes is an important step for the identification of novel therapeutic targets. PMID- 11258593 TI - The predictive power of haplotypes in clinical response. AB - A variety of approaches have been proposed to find genetic markers that can be used in a clinical setting. Single nucleotide polymorphisms (SNPs) are the basis of the most commonly used approaches. Here we describe an approach using gene based haplotypes, which are collections of SNPs located throughout the ftinctional regions of candidate genes, and organised as they occur separately on an individual's two chromosomes. The main point of this review is that the haplotype has greater power than any individual SNP to track an unobsenrved, but evolutionarily linked, variable site. PMID- 11258594 TI - Mining for SNPs: putting the common variants--common disease hypothesis to the test. AB - Classical molecular genetic strategies have succeeded in identifying mutations responsible for numerous rare diseases with Mendelian patterns of inheritance, but have been largely unsuccessful in unravelling the (genetic basis of complex medical conditions like cardiovascular disease' diabetes and mental illness. These common disorders are shaped by multiple genes that exert weak allelic effects in the setting of confounding environmental variables. Association study designs provide statistical povwer to reveal the modest contributions of weak alleles, and evidence is mounting that common genetic polymorphisms play a role in complex diseases. Cataloguing genetic variation in human populations is a prerequisite for further validation of the 'common variants-common disease' hypothesis, and polymorphism discovery has begun in earnest in the academic and private sector. We will review several strategies for high-throughput polymorphism discovery and discuss the implications of early results from polymorphism screens for future genetic studies. PMID- 11258595 TI - The use of single nucleotide polymorphisms in the isolation of common disease genes. AB - The numerous successes using positional cloning to identify genes mutated in monogenic disorders has galvanised geneticists to start using similar techniques to tackle common complex diseases such as asthma, osteoarthritis, depression and early onset heart disease. The technology is currently at an intermediate stage in which linkage in family studies is being supplemented with locus-specific association studies in populations, enabling accurate localisation of the disease causing or susceptibility gene. These studies are often labour and time intensive unless focus is placed on biological candidate genes. In general, most candidate gene studies for common diseases have been unrewarding. However, single nucleotide polymorphisin (SNP) mapping has accelerated complex disease gene localisation, providing a tool to narrow the linkage region by the detection of multiple SNPs associated with the disease in a relatively small linkage disequilibriuln (LD) region. Identification of susceptibility genes will enable a better understanding of the mechanisms of the disease processes and will facilitate the discovery of new and more efficacious medicines. Whole genome SNP maps will also allow abbreviated SNP profiles to be developed for pharmacogenetic applications, enabling physicians to tailor therapeutic regimens (i.e., identify patients likely to receive therapeutic benefit and not suffer adverse reactions). PMID- 11258596 TI - Insulin receptor variant forms and type 2 diabetes mellitus. AB - Type 2 diabetes is a polygenic and heterogeneous disease resulting from interaction of genetic factors with environmental influences. Numerous candidate genes have been investigated, but no single major susceptibility gene for Type 2 diabetes has been identified. The insulin receptor was considered a plausible candidate gene. The insulin receptor exists in two isoforms differing by the absence (Ex11-) or presence (Ex11+) of 12 amino acids in the C-terminus of the alpha-subunit due to alternative splicing of exon 11.Ex11- binds insulin with two fold higher affinity than Ex11+. This difference is paralleled by a decreased sensitivity for metabolic actions of insulin. Some, but not all, studies have reported that expression of the low-affinity Exll+ is increased in Type 2 diabetes, suggesting that alterations in abundance of the two isoforms mnight contribute to insulin resistance. Insulin and Type 1 insulin-like growth factor (IGF) receptors have been shown to form hybrid receptors in tissues co-expressing both molecules. Hybrid receptors bind IGF-I, but not insulin, with high affinity, and behave as IGF-I receptors rather than insulin receptors in terms of receptor autophosphorylation and hormone internalisation. It has been shown that the abundance of hybrid receptors is increased in skeletal muscle and fat from Type 2 diabetic patients, and is negatively correlated with in vivo insulin sensitivity. Mutations in the insulin receptor gene were identified in studies which examined an appropriately sized population of Type 2 diabetic patients. The prevalence of mutations in the insulin receptor gene ranged from 0.4 to 7.8%. PMID- 11258597 TI - Pharmacogenomics to predict drug response. AB - From theory to proof-of-concept, pharmacogenomics promises to improve future general healthcare in a number of ways. By identifying individuals who will respond to a particular drug treatment compared to those who have a low probability of response, pharmacogenomic test development hopes to aid the physician in prescribing the optimal medication for each patient. This approach promises faster relief from symptoms, a lowering of side effect risks and a reduction in healthcare costs. Pharmacogenomic tests used by the pharmaceutical companies themselves can be used to help identify suitable subjects for clinical trials, aid in interpretation of clinical trial results, find new markets for current products and speed up the development of new treatments and therapies. This type of approach should also see fewer compounds failing during later phases of development. The questions we are faced with as we enter the new millennium, however, are if and when the promises of pharmacogenomnics in improving healthcare will be fulfilled. Currently, there are only a handful of pharmacogenomic tests and associated products which are commercially available and it remains to be seen what impact these will have on the market and on healthcare in general. PMID- 11258598 TI - Genome polymorphism and alcoholism. AB - Different gene variants have been identified as risk or protective factors in alcoholism. The genes coding for dopamine receptors, serotonin transporters, and dehydrogenases represent susceptibility loci for addictive behaviour. However, alcoholism represents a complex psychiatric symptomatology which is caused by multiple factors, both genetic and environmental. Furthermore, there are probably different subtypes of alcoholism each with a distinct pathophysiology, and thus a different genetic background. Genetic research can help to identify such subtypes, which may require different therapeutic approaches. However, gene polymorphisms are not only responsible for a predisposition to alcoholism, but also for personality traits which influence the likelihood of developing addictive behaviour. Moreover, genetic polymorphisms are probably involved in the way an individual responds to treatment. Also, the severity of secondary diseases resulting from chronic alcohol uptake may depend on the genetic makeup of an individual. New treatment strategies focusing on genes contributing towards drug and alcohol dependence (such as gene therapy) are already under examination in animal models. However, further research is required before these developments will considerably change today's clinical handling of alcoholism. PMID- 11258599 TI - Genetic variation in coronary heart disease and myocardial infarction: methodological overview and clinical evidence. AB - The precise molecular mechanisrms that lead to coronary artery disease (CAD) and myocardial infarction (MI) are not understood, despite a wealth of knowledge on predisposing risk factors and pathomechanisms. CAD and MI are complex genetic diseases; neither the environment alone nor a single gene cause disease, but a mix of environmental and genetic factors lead to atherosclerosis of the coronary arteries and subsequent manifestation of clinical disease. The biological complexity of atherosclerotic disease results from unknown or unpredictable interactions of many genetic and environmental factors which, by themselves, have only been partially identified. According to current knowledge, genetic variations in causative or susceptihility genes form the basis of molecular mechanisms that, together with environmental impact, lead to CAD/MI and determine its clinical course. Linkage analysis, which follows 'disease' alleles in families, or genetic association in a population of unrelated individuals are tools used in the search for chromosomal loci and candidate genes that are involved in these complex diseases. Progress in sequencing and mapping of the human genorne and efforts to identify all of the expected one million single nucleotide polymorphisms (SNPs) expected to be present in mankind will allow new approaches such as genome-wide association studies. The contribution of the current state of knowledge on genetic variation in man towards the dissection of CAD/MI as complex traits is sobering. Raised expectations with regard to the power of molecular genetic studies as compared to the traditional pathophysiological experimental approaches, lack of precise clinical phenotyping, lack of functional characterisation of gene variants, and the vast number of yet undetected genes may provide some explanation. Except for certain polymorphisms in lipid genes (i.e., apolipoprotein E [apo E]) or rare genetic variations (i.e., LDL receptor), which have a causal effect on both the intermediate (LDL cholesterol level in plasma) and the clinical phenotypes (CAD/MI), the role of most gene polymorphisms is controversial or unknown. Despite the enormous progress in sequencing the human genome and in molecular genetic and bioinformatic techniques during the past decade, the progress in mapping and identifying genes responsible for complex traits such as CAD/MI has been modest and presents a formidable challenge to medical research in the 21st century. PMID- 11258600 TI - High-throughput genotyping assay approaches. AB - High-throughput genotyping approaches are being developed to meet the demands of pharmacogenomnics, where numerous individuals are studied with thousands of single nucleotide polymorphism (SNP) markers. All non-gel-based genotyping approaches achieve allelic discrimination by one of four mechanisms: allele specific hybridisation, allele-specific primer extension, allele-specific oligonucleotide ligation and allele-specific cleavage of a flap probe. By combining one of these allelic discrimination mechanisms with either a homogeneous or solid-phase reaction format and a detection method such as fluorescence intensity, fluorescence polarisation or mass spectrometry, a number of viable high-throughput genotyping methods have been developed and are being readied for routine use. With the biochemistry for robust genotyping in place, good engineering solutions are needed to make high-throughput genotyping a reality. PMID- 11258601 TI - Therapeutic apheresis: further development in the new century. PMID- 11258602 TI - Large volume apheresis of autologous plasma and preparation of autologous fibrin glue from the plasma. AB - Autologous blood transfusion (ABT) is useful for prevention of undesirable effects of allogeneic blood transfusion. In our hospital, not only autologous whole blood but also autologous red blood cells, autologous fresh frozen plasma (Auto-FFP), and autologous fibrin glue (Auto-FG) are routinely produced for surgical patients. The Auto-FG is prepared from plasma which is separated from manually collected whole blood. However, when a large volume of Auto-FG is required, the plasma obtained by an apheresis method may be useful. Therefore, a pilot study was conducted to determine whether a collection of 2 U (160 ml) of red blood cells (RBCs) and 400 ml of plasma at 1 apheresis is acceptable. We first performed the apheresis on healthy donors, and then applied for autologous blood donation. The apheresis is safe. The collected plasma is used for the production of Auto-FFP and Auto-FG. The remaining RBCs also are used for ABT. The preparation of Auto-FG is simple, and it is effective for the reduction of allogeneic fibrin glue. PMID- 11258603 TI - Status of platelet collection and platelet transfusion. AB - Platelet product derived from single donor plateletpheresis is required to reduce the risks of adverse reactions by blood transfusion. The objectives of this study are to evaluate the status of platelet collection and its efficacy by various kinds of plateletpheresis equipment and to assess the achievement of platelet transfusion by platelet product derived from a single donor. Since the blood centers have introduced some kinds of efficient plateletpheresis equipment, large units of platelet products have been supplied mainly for the patients. Amicus and CCS might be preferable plateletpheresis machines because of their collection efficiencies and wider indication for donors. The average number of donors of platelet product per patient has recently reached nearly 1.0, and around 90% of patients have received platelet product derived from a single donor in the recent several years. However, platelet transfusion derived from a single donor has not yet been completely achieved. Each regional blood center should seriously consider the efficacy of each plateletpheresis equipment and arrange the equipment to collect platelets more effectively to achieve platelet transfusion from a single donor. PMID- 11258604 TI - Acute changes in plasma levels of hepatocyte growth factor during low-density lipoprotein apheresis. AB - Hepatocyte growth factor (HGF) is a mesenchyme-derived pleiotropic factor, and angiogenesis is included in a variety of its functional effects. HGF levels were measured in 5 sessions of low-density lipoprotein (LDL) apheresis in 3 patients with severe hypercholesterolemia. Blood was collected at the start (T0) and at 1,000 ml (T1), 2,000 ml (T2), and 3,000 ml (T3) plasma treatments. During LDL apheresis, HGF levels increased from 1.59 +/- 0.78 (mean +/- SE, n = 5) ng/ml at T0 to 6.64 +/- 0.97 at T1, 6.28 +/- 0.97 at T2, and 5.20 +/- 0.94 at T3. In one apheresis session, HGF increased immediately at the 100 ml plasma treatment stage. HGF was adsorbed completely by a dextran-sulfate (DS) column. Despite the adsorption by the DS column, HGF in the patient blood increased to the levels with functional effects. The improvement of ischemic symptoms due to LDL apheresis may be related to the angiogenic activities of HGF. PMID- 11258605 TI - Successful delivery in a pregnant woman with lupus anticoagulant positive systemic lupus erythematosus treated with double filtration plasmapheresis. AB - The case was a 29 year old female who has suffered from systemic lupus erythematosus (SLE) since 15 years of age. The activity of SLE was low, and she took prednisolone orally. Her first pregnancy failed after 14 weeks. In the second pregnancy, she had thrombocytopenia, prolonged activated partial thromboplastin time (APTT), positive lupus anticoagulant (LAC) and thus was diagnosed with antiphospholipid antibody syndrome (APS). Combination therapy with steroids and aspirin was started, and she underwent treatment of double filtration plasmapheresis (DFPP) in the early stage of pregnancy. Her platelet count increased, and the value of APTT has normalized with DFPP treatment. She delivered successfully on the 32nd week of pregnancy. We think that DFPP is an effective and safe treatment in patients with an LAC positive pregnancy. PMID- 11258606 TI - The severity of hyperdynamic circulation may predict the effects of direct hemoperfusion with the adsorbent column using polymyxin B-immobilized fiber in patients with gram-negative septic shock. AB - It has been reported that direct hemoperfusion with the adsorbent column using polymyxin B-immobilized fiber (DHP with PMX-F column) ameliorates hyperdynamic circulation in septic shock and improves survival rate. However, the clinical characteristics of patients with an improvement of septic shock after DHP with PMX-F column have not been evaluated. To clarify this issue, the clinical profiles of 46 patients who were suggested to have gram-negative septic shock and treated using DHP with PMX-F column were analyzed retrospectively. Of 46 patients, 31 were diagnosed with gram-negative septic shock (G group). Mean arterial pressure (MAP) just before DHP with PMX-F column was not different between the G and the non-G group. As compared with the non-G group, the G group had a higher cardiac index (CI) and a lower systemic vascular resistance (SVR). Significant increases in MAP and SVR with a significant decrease in CI were observed after DHP with PMX-F column in the G group. In the non-G group, MAP was significantly increased after the DHP therapy, but systemic hemodynamics were unchanged. Patients in the G group who fulfilled the following criteria were considered as the effective group: MAP was elevated more than 10 mm Hg or 125% of the basal MAP and/or the dose of vasopressors was reduced after DHP with PMX-F column. Twenty-one patients (67.8%) were in the effective group. In comparison with the effective group, the noneffective group was characterized by a significant increase in CI before DHP with PMX-F column. All patients with a CI less than 6 L/min/m2 were in the effective group. These data suggest that DHP with PMX-F column was useful for patients with gram-negative septic shock who did not have severe hyperdynamic circulation. PMID- 11258607 TI - Improved outcome prediction for patients with multiple organ failure undergoing continuous hemodiafiltration. AB - A number of patients with multiple organ failure (MOF) regardless of accompanying acute renal failure have been treated with continuous hemodiafiltration (CHDF). However, despite its high cost, the costs/benefits of CHDF for MOF patients still need to be evaluated. Although many scoring systems were established to predict the outcome of MOF, their predictive powers were not estimated in MOF patients undergoing CHDF. Therefore, using 52 Japanese patients with MOF treated with CHDF for more than 1 week, we estimated the predictive powers of multiple organ dysfunction (MOD) scores and acute physiology and chronic health evaluation (APACHE) III scores, retrospectively. The patients were divided into 2 groups according to outcome at Day 28 after the initiation of CHDF. In both scoring systems, the median values at Day 0 were not significantly different between the survival (n = 19) and the nonsurvival (n = 33) groups. In contrast, at Day 3, the median values of MOD scores was 4 (0-14) in the survival group and 9 (1-12) in the nonsurvival group (p = 0.0035). The median value of APACHE III scores were 37 (19-97) and 87 (16-150) at Day 3, respectively (p < 0.0001). In the survival group, APACHE III scores significantly decreased from the median value of 64 (32 89) to 37 (p = 0.0269), and in the nonsurvival group, it increased significantly from the median value of 70 (29-103) to 87 (p = 0.0116). In contrast, no significant changes were observed in the MOD scores. In conclusion, the MOD score and the APACHE III score systems had less power to predict the outcome of MOF patients undergoing CHDF at Day 0. However, rescoring at Day 3 of each index was much more powerful to accurately predict the outcome of such patients. PMID- 11258608 TI - Changes of cytokines in direct endotoxin adsorption treatment on postoperative multiple organ failure. AB - Multiple organ failure (MOF) is a serious condition that involves simultaneous or consecutive functional failure of several important organs. Furthermore, sepsis is known to play an important role in the occurrence of MOF. Hemoadsorption therapy with the endotoxin adsorption column containing polymyxin B immobilized fibers by direct hemoperfusion (PMX-DHP) is reportedly effective in the treatment of septic shock. This study examined the changes induced on cytokines upon PMX DHP treatment in 25 patients who underwent emergency abdominal surgery and were immediately started on a postoperative regimen of continuous hemodiafiltration (CHDF) and PMX-DHP. Postoperative MOF was observed in these patients with a mean APACHE II SCORE of 25.5. Eighty percent of patients survived for more than 1 month. We were able to reduce the necessary dose of dopamine in 85.7% of patients because hemodynamic stability improved after administration of PMX-DHP. Interleukin 6 blood levels did not change significantly before or after PMX-DHP treatment in either the surviving or nonsurviving patients. Blood interleukin 1 receptor antagonist levels decreased in both groups. Intercellular adhesion molecular-1, NOx, and thrombomodulin did not change significantly during the course of treatment in either group. Decreased blood levels of PAI-1 levels were found in the surviving patients whereas increased levels of PAI-1 were found in the non-surviving patients. In conclusion, PMX-DHP treatment may be limited clinically in its ability to remove inflammatory cytokines and humoral mediators. However, PMX-DHP treatment is useful for hemodynamic stabilization, which prevents development of MOF. PMID- 11258609 TI - Donor leukocyte infusion for Japanese patients with relapsed leukemia after allogeneic bone marrow transplantation: indications and dose escalation. AB - To clarify the role of dose escalation of donor leukocyte infusion (DLI) in the treatment of relapsed leukemia after allogeneic bone marrow transplant (BMT), data from 100 patients were collected from 46 facilities in Japan and analyzed with respect to indications and infused cell dose. Complete remission (CR) was achieved in 11 of 12 (91%) patients with relapsed chronic myelogenous leukemia (CML) in the chronic phase, 3 of 11 (27%) with CML in the acute phase, 8 of 21 (38%) with acute myelogenous leukemia (AML), 6 of 23 (25%) with acute lymphoblastic leukemia (ALL), and 5 of 11 (45%) with myelodysplastic syndrome (MDS). The probability of remaining in CR at 3 years was 82% in CML patients in the chronic phase, but 0% in those with CML in the acute phase, 7% in those with AML, 0% with ALL, and 33% with MDS. Acute graft-versus-host disease (GVHD) (> or = 2) developed in 31 of 89 (34%) patients with human leukocyte antigen identical related donors and was fatal for 7 (7%). A leukocyte dose of 1 x 10(7)/kg of recipient body weight with CML in the chronic phase, 3 x 10(7)/kg of recipient body weight with MDS, and 1 x 10(8)/kg of recipient body weight with acute leukemia appeared to be optimal as an initial dose of DLI. However, the minimal dose of leukocyte developing fatal GVHD was 7 x 10(7)/kg of recipient body weight. These suggest that a relatively small dose of DLI ranging from 1 x 10(7)/kg to 5 x 10(7)/kg of recipient body weight should be administered initially then the infused escalating dose 2 or 3 months later in patients with CML in the chronic phase and MDS. However, a large number of leukocytes around 1 x 10(8)/kg are needed to induce graft versus leukemia effects in patients with acute leukemia despite a 7% fatality in GVHD. PMID- 11258610 TI - Concentration of progenitor cells collected from bone marrow fluid using a continuous flow cell separator. AB - In ABO major incompatibility on bone marrow transplantation (BMT), red cells must be removed from collected marrow fluid to prevent hemolysis. We report the concentration of progenitor cells collected using a continuous flow cell separator (Cobe Spectra). The average volume of concentrated bone marrow was 132 +/- 47 ml and that of red cells included was 5.1 +/- 2.4 ml. The red cell removal rate was 97.6%. The recovery rate was 40.6% for total nuclear cells, 77.9% for mononuclear cells, 100% for CD34+ cells, and 93.9% for colony forming unit granulocyte-macrophage. Eighteen patients undergoing allogeneic BMT showed no signs of fever or hemolysis during concentrated marrow fluid transfusion. The recovery rate of progenitor cells was high, indicating sufficient recovery of hemopoiesis. This technique is applicable in ABO-incompatible BMT and in frozen storage stem cells. PMID- 11258611 TI - A case of thrombotic thrombocytopenic purpura refractory to plasma exchange. AB - We experienced a case of thrombotic thrombocytopenic purpura (TTP) finally relieved after 74 sessions of plasma exchange (PE). The patient was a 56-year-old male. In August 1999, he was examined in emergency because of brown urine and a lowered level of consciousness. As TTP was suspected according to the laboratory findings of abnormally high lactate dehydrogenase and total bilirubin, decreased platelet counts, and numerous fragmented erythrocytes, he was admitted to the ICU of our hospital. Immediately after admission, PE was started consecutively. Upon concomitant use of antiplatelet drugs and prostacyclin, the level of platelet counts recovered to 100,000/microl once, but decreased again. Thus, in addition to the PE, prednisolone and vincristine were administrated, which elevated the level of platelet counts to 200,000 to 300,000/microl. Since the erythrocyte fragmentation was noted frequently, PE was continued twice a week. From the 60th day of admission onward, however, his body temperature rose above 40 degrees C with a rapid increase of C-reactive protein. A blood culture detected methicillin resistant Staphylococcus aureus (MRSA) which derived from a left lung abscess. During the course of anti-MRSA treatment, he presented acute renal failure and acute hepatic dysfunction, but survived because of the combined therapy. He was discharged on the 180th day of admission. These results suggest that a combined therapy of steroid and vincristine is effective to treat TTP refractory to PE, but careful attention should be paid to the complications caused by immunosuppression. PMID- 11258612 TI - Allogeneic peripheral blood stem cell transplantation for severe aplastic anemia. AB - Allogeneic peripheral blood stem cell transplantation (PBSCT) is rarely applied for the treatment of severe aplastic anemia (SAA) because of questionable durability of engraftment and increased risk of graft versus host disease (GVHD). We performed allogeneic PBSCT in 3 SAA patients from their human leukocyte antigen (HLA)-identical siblings. One received bone marrow after conditioning with cyclophoshamide (Cy) plus antithymocyte globulin. He had a second transplant with peripheral blood stem cells from the original donor because of a graft failure (GF). Two other patients received PBSCT as a first option, with Cy as the only conditioning drug. The 3 patients received short-term methotrexate and cyclosporine as a postgrafting immunosupression. In the latter 2 cases, no GF has been observed, and a successful and complete hematological recovery was achieved and maintained for 28 and 25 months, respectively. In conclusion, PBSCT provides a quick and complete hematological recovery in SAA patients. PMID- 11258613 TI - A new method for removal of albumin-binding uremic toxins: efficacy of an albumin dialysate. AB - It is difficult for conventional hemodialysis to remove albumin-binding uremic toxins (ABUTs) even though they are low molecular weight substances. We investigated the efficiency of albumin-dialysate (AD) for removal of ABUT. Phenols and indoxyl sulfate were selected as ABUT. In vitro dialysis was performed for 2 h in the closed circuit with ABUT containing plasma as a test solution using conventional dialysate (CD) or AD. By the use of CD, the ABUT concentration in the test solution only was reduced by 20 to 30%. On the other hand, AD caused a marked reduction and an increase in test solution and dialysate concentration of ABUT, respectively. ABUT in AD could be adsorbed effectively by activated-charcoal column; accordingly, the ABUT concentration in the test solution continued to decrease throughout the study period. These results suggest that AD could remove ABUT more efficiently than CD, and AD may be useful for reducing accumulated ABUT levels. PMID- 11258614 TI - Plasma exchange in stiff-man syndrome. AB - Stiff-man syndrome (STS) is a rare neurological disorder characterized by involuntary axial and proximal limb rigidity and continuous motor unit activity on electromyography (EMG). Autoantibodies to glutamic acid decarboxylase (GAD) present in 60% of the patients are implicated. We report on the use of plasma exchange (PE) in 2 patients with STS whose serum and cerebrospinal fluid were negative for GAD autoantibodies. One patient showed minimal clinical improvement following PE while the second reported subjective improvement, but not any different from that with medications. Based on the results of PE in our patients, it seems that those who are autoantibody negative are less likely to respond. Whether a more aggressive approach to PE will be beneficial remains speculative. PMID- 11258615 TI - Three cases of C-ANCA-positive vasculitis treated with immunoadsorption: possible benefit in early treatment. AB - Wegener's granulomatosis is a vasculitic disease predominantly affecting the upper respiratory tract, lungs, and kidneys. Three patients with Wegener's granulomatosis and rapidly progressive glomerulonephritis were treated with an intensified regimen of immunoadsorption (IA) (Excorim or Therasorb) in addition to cyclophosphamide (CYC) and methylprednisolone (PRE). Patient A had been in remission under oral CYC/PRE. The first exacerbation was treated successfully with 4 IA treatments without changing medication. Patient B experienced 3 flares within 1 year, which were treated with 28 IA (3-7 IAs/course), intravenous CYC after each course, and PRE. A fall of creatinine levels from 120 to 190 micromol/L to 100 micromol/L was noted after IA and before administration of CYC. Patient C presented in uremia. Autoantibodies were eliminated by 11 IA treatments parallel to CYC/PRE therapy. They remained within a normal range for >1 year's follow-up; however, kidney function did not return. In conclusion, the observations in Patients A and B suggest a beneficial therapeutic effect of early IA in WG. PMID- 11258616 TI - Experience with artificial liver support in 16 living related liver transplant recipients. AB - This study was a retrospective investigation about the indication and efficacy of artifical liver support for liver transplant recipients. Apheresis was performed in 16 of 41 patients subjected to living related liver transplantation (LRLTx) as articial liver support, including plasmapheresis (PP) in 13 cases, continuous hemodiafiltration (CHDF) in 7 cases, and plasma adsorption (PA) in 2 cases. One patient with cryptogenic liver cirrhosis was subjected to PP before the LRLTx, and the result was satisfactory. On the contrary, the results of PP and CHDF for graft, respiratory, or cardiac failure were not acceptable. Only 1 patient survived despite multiple organ failure. Both PP and PA for patients with hyperbilirubinemia were effective and improved their critical conditions. We conclude that apheresis for liver transplant patients is effective to treat hyperbilirubinemia, but it is not indicated for respiratory and cardiac failure nor for hepatic failure. PMID- 11258617 TI - Transdermal delivery of antisense oligonucleotides can induce changes in gene expression in vivo. AB - The potential for using antisense compounds as therapeutic agents has generated great enthusiasm. Strategies for delivery of these compounds are, therefore, of great interest. Transdermal iontophoresis has been used successfully as an enhancement technique for the transdermal delivery of these compounds in vitro. The effectiveness of using percutaneous penetration as a means to deliver therapeutic levels of these compounds in vivo, however, remains to be demonstrated. The purpose of this work was to demonstrate the ability of iontophoretically delivered compounds to alter enzyme levels in the intact rat. A C5 propyne-modified phosphorothioate oligonucleotide (PS-ODN) targeted to the cytochrome p450-3A2 (CYP3A2) mRNA translational start site and the reverse sequence, used as a control, were synthesized. A patch containing either an oligonucleotide or a buffer control was placed on the animal's back, and an iontophoretic current of 0.5 mA/cm2 was applied for 3.5 hours. Twenty-four hours later, CYP3A2 levels were measured noninvasively using the midazolam-induced sleeping rat model. Liver and small intestinal microsomes were made after completion of sleep studies and assayed for CYP3A2, CYP1A1/2, CYP2B1/2, and CYP2E1. Midozolam-treated animals with antisense to CYP3A2 slept significantly longer than did the controls (p < 0.05). CYP3A2 levels were significantly lower in liver microsomes from antisense-treated animals than in either buffer control (p < 0.001) or reverse sequence animals (p < 0.05). The reverse sequence was also significantly different from the buffer control (p < 0.01), indicating a nonspecific effect of the PS background. Nontarget cytochrome levels were not altered by treatment. There were no significant differences in small intestine CYP3A2 levels between treatment groups. These data demonstrate that transdermally delivered PS-ODN can reach concentrations sufficient to induce changes in specific target enzymes in vivo. Further studies are warranted to investigate potential uses for these molecules. PMID- 11258618 TI - Tissue distribution and physiologically based pharmacokinetics of antisense phosphorothioate oligonucleotide ISIS 1082 in rat. AB - The aim of this study was to develop a whole body physiologically based model of the pharmacokinetics (PBPK) of the phosphorothioate oligonucleotide (PS-ODN) ISIS 1082 in vivo. Rats were administered an intravenous (i.v.) bolus dose of ISIS 1082 (10 mg/kg plus 3H tracer), and arterial blood and tissues were taken at specific times up to 72 hours. Radioactivity was measured in all samples. The parent compound was determined specifically in blood and tissues at 90 minutes and in liver and kidney also at 24 hours, using capillary gel electrophoresis (CGE). A whole body PBPK model was fitted to the combined blood and tissue radioactivity data using nonlinear regression analysis. CGE analysis indicated that the predominant species in plasma and all tissues is ISIS 1082, together with some n-1 and n-2 metabolites. Total radioactivity primarily reflects these species. The whole body model successfully described temporal events in all tissues. However, to adequately model the experimental data, all tissues had to be partitioned into vascular and extravascular spaces to accommodate the relatively slow distribution of ISIS 1082 out of blood because of a permeability rate limitation. ISIS 1082 distributes extensively into tissues, but the relative affinity varies enormously, being highest for kidney and liver and lowest for muscle and brain. A whole body PBPK model with a permeability rate limited tissue distribution was developed that adequately described events in both blood and tissue for an oligonucleotide. This model has the potential not only to characterize the events in individual tissues throughout the body for such compounds but also to scale across animal species, including human. PMID- 11258619 TI - Introduction of an RNA stability element at the 5'-end of an antisense RNA cassette increases the inhibition of target RNA translation. AB - This communication describes improvement strategies used on a previously described two-unit antisense RNA cassette system. This cassette system encodes RNA with noncontiguous regions of complementarity to a bacterial target RNA, lacI mRNA. One of the units of complementarity was contained within an RNA stem-loop resembling that of the very efficient, naturally occurring antisense RNA CopA. As relatively low inhibitory activity was obtained previously, we tested variants in which several stem-loops were combined within one RNA, each of them directed against a different stretch of target RNA. One to four stem-loop RNAs were tested and found to be relatively ineffective, likely because of low metabolic stability. To increase the intracellular stability of these and other antisense RNAs, a stabilizer element (stem-loop derived from gene 32 mRNA of phage T4) was inserted at their 5'-ends. The results indicate that addition of this element indeed increased antisense RNA efficiency in vivo. As expected, this effect was primarily due to a longer antisense RNA half-life, as shown by RNA abundance (Northern analysis) and decay rates (rifampicin runout experiments). In summary, the results reported indicate that rational design of antisense RNA is feasible, but that the degree of inhibition (approximately 75% maximum inhibition) accomplished here could still be improved. PMID- 11258620 TI - Development of ribozymes that target stathmin, a major regulator of the mitotic spindle. AB - Stathmin is a major cytosolic phosphoprotein that plays an important role in the control of cellular proliferation by regulating the dynamics of the microtubules that make up the mitotic spindle. Because stathmin is expressed at high levels in all human cancers, it is an attractive molecular target for anticancer interventions. We had shown previously that antisense stathmin inhibition results in marked abrogation of the transformed phenotype of leukemic cells in vitro and in vivo. Unlike the antisense approach, ribozymes can catalytically cleave several molecules of target RNA. This may provide a more efficient strategy for downregulating genes, such as stathmin, that are expressed at very high levels in cancer cells. We designed several antistathmin hammerhead ribozymes and tested their cleavage activity against short synthetic stathmin RNA substrates. In vitro cleavage studies demonstrated site-specific cleavage of stathmin RNA that was dependent on ribozyme concentration and duration of exposure to ribozyme. The most active antistathmin ribozyme was capable of cleaving >90% stathmin RNA in a catalytic manner, cleaving multiple substrate molecules per ribozyme molecule. We also demonstrated that the designed antistathmin ribozymes are capable of selectively cleaving native stathmin RNA in a mixture of total RNA isolated from leukemic cells. These antistathmin ribozymes may provide a novel and effective form of gene therapy that may be applicable to a wide variety of human cancers. PMID- 11258621 TI - Complex host cell responses to antisense suppression of ACHE gene expression. AB - 3'-End-capped, 20-mer antisense oligodeoxynucleotides (AS-ODN) protected with 2' O-methyl (Me) or phosphorothioate (PS) substitutions were targeted to acetylcholinesterase (AChE) mRNA and studied in PC12 cells. Me-modified AS-ODN suppressed AChE activity up to 50% at concentrations of 0.02-100 nM. PS-ODN was effective at 1-100 nM. Both AS-ODN displayed progressively decreased efficacy above 10 nM. In situ hybridization and confocal microscopy demonstrated dose dependent decreases, then increases, in AChE mRNA. Moreover, labeling at nuclear foci suggested facilitated transcription or stabilization of AChE mRNA or both under AS-ODN. Intracellular concentrations of biotinylated oligonucleotide equaled those of target mRNA at extracellular concentrations of 0.02 nM yet increased only 6-fold at 1 microM ODN. Above 50 nM, sequence-independent swelling of cellular, but not nuclear, volume was observed. Our findings demonstrate suppressed AChE expression using extremely low concentrations of AS-ODN and attribute reduced efficacy at higher concentrations to complex host cell feedback responses. PMID- 11258623 TI - Improved intracellular delivery of oligonucleotides by square wave electroporation. AB - Prior studies have shown that electroporation is a simple and effective method for the introduction of oligonucleotides (ODN) into cells. In ex vivo bone marrow purging models, electroporation of ODN into cells has been associated with selective killing of human neoplastic cells while sparing hematopoietic stem cells. Prior studies used conventional electroporation methods (i.e., exponential decay) to introduce ODN into cells. Square wave electroporation allows the delivery of a more defined and regulated electrical pulse and is associated with high transfection efficiencies in a variety of systems. The current study was undertaken to determine whether square wave electroporation was more effective than exponential decay electroporation for the delivery of ODN into hematopoietic cells. Using fluorescein-tagged ODN and K562, chronic myelogenous leukemia (CML) cells, higher transfection rates were observed after square wave electroporation. In addition, c-myc antisense ODN were more effective in reducing c-myc protein when introduced by square wave electroporation, as compared with introduction by exponential decay electroporation. Square wave electroporation is thus identified as the optimal method for delivering ODN into hematopoietic cells. PMID- 11258622 TI - Impact of a phosphorothioate oligodeoxynucleotide MCP-1 on NF-kappaB, AP-1, SP1 and NF-kappaB, and AP-1 subunit composition in human pulmonary endothelial cells. AB - Phosphorothioate oligodeoxynucleotides (PS-ODN) are widely used prototypic antisense oligomers for sequence-specific suppression of normal and diseased gene expression. As polyanionic molecules, however, PS-ODN may also evoke nonsequence specific side effects. The objective of the present study was to evaluate the impact of PS-ODN treatment of human pulmonary artery endothelial cells (HPAEC) and microvascular endothelial cells of the lung (HMVEC-L) on the cellular pool of the transcription factors nuclear factor-kappaB (NF-kappaB) and activator protein 1 (AP-1) as well as Sp1, using gel shift assays. In addition, by performing supershift assays, we investigated whether antisense treatment of endothelial cells affected the subunit composition of NF-kappaB and AP-1. Our data show that pretreatment of HPAEC and HMVEC-L with PS-ODN doses ranging from 50 to 5000 nM did not affect the total NF-kappaB, AP-1, or Sp1 pool in tumor necrosis factor alpha (TNF-alpha)-activated endothelial cells (EC) or the subunit composition of the transcription factors NF-kappaB and AP-1. These findings suggest that putative nonsequence-specific effects of PS-ODN are not due to interactions of these oligomers with the transcription factors NF-kappaB, AP-1, or Sp1, at least in the EC type, a common target in transfection studies. PMID- 11258624 TI - In search of the Holy Grail: NF1 mutation analysis and genotype-phenotype correlation. PMID- 11258625 TI - Exon 10b of the NF1 gene represents a mutational hotspot and harbors a recurrent missense mutation Y489C associated with aberrant splicing. AB - PURPOSE: To analyze the spectrum and frequency of NF1 mutations in exon 10b. METHODS: Mutation and sequence analysis was performed at the DNA and cDNA level. RESULTS: We identified nine exon 10b mutations in 232 unrelated patients. Some mutations were recurrent (Y489C and L508P), others were unique (1465-1466insC and IVS10b+2delTAAG). Surprisingly, at the RNA level, Y489C causes skipping of the last 62 nucleotides of exon 10b. Another recurrent mutation, L508P, is undetectable by the Protein Truncation Test. CONCLUSION: As exon 10b shows the highest mutation rate yet found in any of the 60 NF1 exons, it should be implemented with priority in mutation analysis. PMID- 11258627 TI - Transmission of Angelman syndrome by an affected mother. AB - PURPOSE: To determine: 1) If a 15q11-13 deletion was transmitted from a female with Angelman syndrome to her fetus, and 2) If the UBE3A gene was functionally imprinted in fetal eye. METHODS: Individuals were genotyped by microsatellite analysis. DNA methylation imprints were assessed by Southern blot analysis and methylation-specific PCR. Expression was analyzed by RT-PCR. RESULTS: The mother and fetus inherited large deletions of maternal 15q11-13 and demonstrated paternal-only DNA methylation imprints along 15q11-13. UBE3A was paternally expressed in eye tissue from the fetus with Angelman syndrome. CONCLUSIONS: We show that females with Angelman syndrome are fully capable of reproduction and that UBE3A is not imprinted in fetal eye. PMID- 11258626 TI - Molecular refinement of karyotype: beyond the cytogenetic band. AB - PURPOSE: Illustrate the use of molecular methodologies to delineate subtle, de novo, chromosome aberrations and determine the presence, or absence, of known genes, allowing improved predictions of long-term phenotypic effect. METHOD: High resolution chromosome analysis followed by FISH and microsatellite analysis to determine the extent and parental origin of the abnormalities. RESULTS: Four de novo deletions involving chromosomes 5q, 10q, and 16p were delineated molecularly. Specific genes were shown to be, or not to be, involved in each aberration, refining karyotype-genotype correlation. CONCLUSION: Molecular characterization of subtle chromosomal aberrations can provide information to assist in predicting clinical outcome in cases involving genes known to have an effect due to haploinsufficiency or aberrant gene dosage. PMID- 11258628 TI - Population-based studies reveal differences in the allelic frequencies of two functionally significant human interleukin-4 receptor polymorphisms in several ethnic groups. AB - PURPOSE: The presence of functionally significant human interleukin-4 receptor sequence variants, Gln551Arg and Ile50Val, was examined in four anonymous New York State populations defined by ethnic origin. These variants were studied because they are associated with atopy or atopic asthma whose prevalence varies in different populations. METHODS: PCR/RFLP (Ile50Val) and PCR/allele-specific oligonucleotide hybridization (Gln551Arg) assays were developed to detect both polymorphisms in 855 newborn screening specimens. RESULTS: Arg551 was most frequently found in Blacks (allele frequency of 68%). However, the Ile50 allele was most common in Whites (allele frequency, 87%). Significantly more Blacks had chromosomes bearing both of the "enhanced signaling" variants (Ile50/Arg551). CONCLUSIONS: Enhanced IL-4R signaling is associated with increased IgE production (atopy). Therefore, our data suggest that the African American population may be at increased risk for diseases, including asthma, which are associated with atopy. These data also emphasize the importance of determining the frequencies of single nucleotide polymorphisms in different populations before drawing conclusions from allele association studies, since the background allele frequencies may be disparate between different populations. PMID- 11258629 TI - Genetic services for common complex disorders: surveys of health maintenance organizations and academic genetic centers. AB - PURPOSE: To learn the extent to which HMOs and academic genetic centers (1) are involved in predictive genetic tests for common, complex disorders and (2) interact with each other. METHODS: Surveys of HMO medical directors and directors of U.S. academic genetic centers. RESULTS: In 1996, approximately 28% of HMOs were covering predictive tests for breast and colon cancer, but 75% of all medical directors said their HMO would consider policies regarding predictive testing in the next 5 years. Approximately 80% of directors of academic genetic centers said they provided genetic counseling services for common adult-onset disorders for patients covered by managed care organizations (MCOs), but they ranked the volume of services they provide for pediatric and prenatal indications much higher. Most academic genetic centers (72%) have contracts with MCOs. CONCLUSION: Although genetic services are being provided by academic genetic centers to patients who are members of managed care organizations, many patients with whom genetic testing for adult onset disorders is discussed may never see a geneticist. Academic genetic centers should educate nongeneticist professionals about the use of tests for common disorders. PMID- 11258630 TI - Inconsistencies in genetic counseling and screening for consanguineous couples and their offspring: the need for practice guidelines. AB - PURPOSE: To determine current practices of genetic counseling and screening for consanguineous couples, their pregnancies and children, and to compare these practices to recommendations in the literature. METHODS: A questionnaire was mailed to 1,582 board certified genetic counselors and medical geneticists in the United States. RESULTS: The return rate was 20% (n = 309). There was wide variation in the risk figures quoted to consanguineous couples to have offspring with birth defects and mental retardation (1% to 75% for incest between first degree relatives, and 0.25% to 20% for first cousin unions). Suggested screening practices differed for consanguineous unions before conception, during pregnancy, following birth, and for children placed for adoption. Most respondents recommended screening based on ethnicity, yet disagreed as to which genetic disorders to include. CONCLUSIONS: To standardize genetic services, guidelines for screening the offspring of consanguineous unions are needed. A consensus should be reached as to the empirical risks for genetic disorders, birth defects, and mental retardation that may impair the offspring of consanguineous unions, with definition as to what these disorders are, and if the data applies to global populations. Guidelines should consider costs, the sensitivity and specificity of DNA and biochemical testing, and current practices of prenatal and newborn screening. Consideration should be given to screening based on ethnicity, particularly in populations where consanguineous unions are common, while remaining sensitive to cultural belief systems. Recommendations for screening healthy children from consanguineous unions to be placed for adoption pose ethical challenges. PMID- 11258631 TI - Medium-chain acyl-CoA dehydrogenase deficiency: sudden and unexpected death of a 45 year old woman. PMID- 11258632 TI - A new age in the genetics of deafness. AB - Recent advancements have been made in understanding, diagnosing, and treating deafness. In particular, much has been learned from the discovery of a small fraction of the genes responsible for deafness. This understanding will doubtless increase as additional genes are cloned and their functions elucidated. Trailing close behind these achievements will be more clinical advancements facilitating diagnosis of the etiologies of deafness. Integrating these genetic and clinical perspectives is critical to the development of better treatments and interventional strategies for deafness and its associated difficulties. Although opinions toward these advancements are likely to vary between the hearing population and the Deaf community, a growing understanding of the hearing process and how genetic variations result in deafness is ultimately likely to offer benefits to both groups. PMID- 11258633 TI - Southwestern Athabaskan genetic diseases. PMID- 11258634 TI - Population-based surveillance of services provided to counseling and prenatal clients in a multi-state region by state health departments: a proposal. PMID- 11258635 TI - Rescue lifting system (RLS) might help to prevent death after rescue from immersion in cold water. AB - OBJECTIVE: In order to prevent sudden death after rescue from immersion in cold water, victims should be handled carefully avoiding additional cardiovascular stress. In this study we investigated if a new double-sling rescue system ("Rescue Lifting System-RLS) was superior to conventional single-sling techniques. METHODS: We studied 14 healthy male subjects in good physical condition aged 21 to 40 years. They were lifted up from the ground with the new RLS and two conventional techniques ("Lifesling" and a navy rescue system used in SAR helicopters). Heart rate was determined by QRS detection (Polar Precision Performance device; Polar Electro Oy, Kempele, Finland) and blood pressure by sphygmomanometry. RLS and "Lifesling" were tested under conditions of dry land and immersion in 18 degrees C water. RESULTS: Rescue with RLS induced only moderate heart rate changes which were significantly lower (about 30 bpm) than with conventional techniques. These findings could be reproduced under "wet" condition. DISCUSSION: RLS enables rescue in a supine position avoiding extensive orthostatic stress. It might therefore be favourable in preventing sudden death after rescue from immersion in cold water. PMID- 11258636 TI - Catecholamines response of high performance wheelchair athletes at rest and during exercise with autonomic dysreflexia. AB - Autonomic dysreflexia presents a special situation in high-lesion spinal cord injury, however, intentionally or self-induced autonomic dysreflexia directly before or during competition to increase performance, so called 'boosting', is also being reported. In order to examine the influence of autonomic dysreflexia on plasma catecholamines, cardiocirculatory and metabolic parameters, 6 spinal cord injured wheelchair athletes with high-level lesions underwent wheelchair ergometry without (ST1) and with (ST2) autonomic dysreflexia. At the point of exhaustion significantly higher values for norepinephrine and epinephrine were observed in ST2 than in ST1. During autonomic dysreflexia a significantly higher peak performance (77.5 vs. 72.5 watt), higher peak heart rate (161 vs. 149 x min( 1)), and peak oxygen consumption (1.96 vs. 1.85 l x min(-1)), with comparable peak lactate (7.11 vs. 7.00 mmol x l(-1)) were reached on average. The blood pressure values in ST2 were partially hypertensive and higher than in ST1. In conclusion, autonomic dysreflexia, as a sympathetic spinal reflex, leads to a higher release of catecholamines during exercise. This results in higher peak performance, peak heart rate, peak oxygen consumption, and higher blood pressure values. The peak lactate, as an indicator of the anaerobic lactate metabolism, was unchanged. However, autonomic dysreflexia presents an unpredictable risk, caused predominantly by hypertensive blood pressure values, for high-lesion spinal cord injured persons at rest and more so during exercise; it is seen as a prohibited manipulation by the doping guidelines of the International Paralympic Committee. PMID- 11258637 TI - The effect of prior high-intensity cycling exercise on the VO2 kinetics during high-intensity cycling exercise is situated at the additional slow component. AB - In previous studies conclusions about the effect of prior exercise on VO2 kinetics of subsequent high-intensity exercise are generally based on observed changes in the overall VO2 response without considering the effects on the VO2 fast and slow component. The aim of the present study was to examine the effect on the VO2 fast and slow component separately. Therefore 10 subjects performed an exercise protocol consisting of an initial 3 min period of unloaded cycling followed by two constant-load work bouts at a work rate corresponding to 90% VO2peak, separated by 3 min of rest and 3 min of unloaded cycling. VO2 was measured on a breath-by-breath basis, and the response curves were analysed by a biexponential model. To increase signal-to-noise ratio, subjects performed four repetitions of the exercise protocol, each separated by at least one day. There was no significant alteration in VO2 kinetic parameters of the primary, fast component after high-intensity exercise. However, there was a significant effect of prior high-intensity exercise on the VO2 kinetic parameters of the slow component. The time constant and the amplitude of the slow component were reduced by respectively 44% (from 231.0 +/- 111.7 s to 130.1 +/- 50.4 s) and 49% (from 824 +/- 270 ml x min(-1) to 417 +/- 134 ml x min(-1)). The results of this study indicate that the effect of high-intensity exercise on the VO2 kinetics of a subsequent high-intensity exercise is probably limited to an effect on the slow component. PMID- 11258638 TI - Relationship between run times to exhaustion at 90, 100, 120, and 140% of vVO2max and velocity expressed relatively to critical velocity and maximal velocity. AB - The aim of the present study was to explain the inter-individual variability in running time to exhaustion (tlim) when running speed was expressed as a percentage of the velocity, associated with maximal oxygen uptake (vVO2max). Indeed for the same percentage of vVO2max the anaerobic contribution to energy supply is different and could be dependent on the critical velocity (Cv) and also on the maximal running velocity (vmax). Ten subjects ran four tlim at 90, 100, 120, and 140% of vVO2max; mean and standard deviation for tlim were 839 +/- 236 s, 357 +/- 110 s, 122 +/- 27 s, and 65 +/- 17s, respectively. Each velocity was then expressed 1) as a percentage of the difference between vVO2max and Cv (%AeSR); 2) as a percentage of the difference between vmax and Cv (%MSR); 3) as a percentage of the difference between vmax and vVO2max (%AnSR). Highest correlations were found between tlim90 and tlim100 and velocity expressed as %MSR (r = -0.82, p < 0.01 and r = -0.75, p < 0.01), and between tlim120 and tlim140 and velocity expressed as %AnSR (r = -0.83, p < 0.01 and r = -0.94, p < 0.001). These results show that the same intensity relative to aerobic contribution did not represent the same absolute intensity for all and could partly explain variability in tlim. Therefore expressing intensity as a percentage of MSR for sub-maximal and maximal velocities and as a percentage of AnSR for supra-maximal velocities allows individual differences in anaerobic work capacity to be taken into account and running times to exhaustion to be predicted accurately. PMID- 11258639 TI - Comparison of two questionnaires with a tri-axial accelerometer to assess physical activity patterns. AB - Two physical activity questionnaires were evaluated against cardiorespiratory fitness (VO2 peak), and a tri-axial accelerometer for movement registration (Tracmor) in 166 men, aged 40 years, within the framework of the Leuven Longitudinal Study on Lifestyle, Fitness, and Health. Tracmor data were obtained during four successive days. Besides the work index, the sport index, and the total activity index from the Baecke questionnaire, the subjective activity score, calculated energy expenditure during work, work index, and the total activity index from the Tecumseh Community questionnaire showed significant correlation coefficients with the mean Tracmor output (r = 0.26-0.47, p < 0.01). The questionnaire submeasures and the Tracmor output as generated in the same physical activity dimension showed the same relationships (r = 0.22-0.50, p < 0.01). Multiple stepwise regression and stepwise discriminant analyses showed the Baecke questionnaire as the best indicator of the subject's physical activity level. Extra information about the physical activity level was given by two Tecumseh submeasures, e.g. energy expenditure during work and sleeping time. The results indicated that the Baecke questionnaire is superior in large-scale studies because of simplicity. However, the Tecumseh questionnaire can give detailed information about physical activity patterns and energy expenditure. PMID- 11258640 TI - Isokinetic strength testing in patients with chronic heart failure--a reliability study. AB - Measurement of skeletal muscle strength gains increasing importance as outcome parameter in patients with chronic heart failure. This study aimed at establishing short-term reliability of isokinetic strength measurements of knee extensor and flexor muscles in 38 patients with chronic heart failure. Strength tests were performed on the Cybex 6000 dynamometer. Trunk fixation was restricted to pelvic fixation. Two bouts of strength testing were performed on day 1 and one on day 5. Each isokinetic bout consisted of 3 reciprocal knee extension and flexion movements with an angular speed of 60 degrees per second. Isometric strength was measured at 30 degree knee angulation. Intraclass correlations ranged between 0.96 and 0.99 for isokinetic and isometric peak torque of knee extensor muscles and 0.82-0.96 for flexor muscles. Analysis of repeated measurements showed significant differences among the values of flexor peak torque in the isokinetic mode and between all measurements in the isometric mode (p < 0.05). Pearson's correlation coefficients for isokinetic and isometric extensor peak torques ranged between 0.95 and 0.99, for flexor peak torques between 0.81 and 0.85 (all p < 0.0001). Measurement of isokinetic knee extensor and flexor peak torque is a reliable method to assess muscle strength in patients with chronic heart failure even with altered trunk fixation. PMID- 11258642 TI - Immunological effects of competitive versus recreational sports in cross-country skiing. AB - For a period of two months during the competitive season the effects of endurance training in cross-country skiers were evaluated in order to compare the adaptive and innate immune systems between 10 competitive athletes, 10 moderately trained athletes and 10 untrained healthy controls. The main results were as follows: the peripheral T-lymphocyte count of the competitive athletes was decreased. In contrast the number of peripheral blood NK cells was increased in this group. These data imply a diminution of the adaptive immune system due to repeated bouts of intense exercise and contemporaneous reinforcement of the innate immune response. Moreover the inducible IL-12-expression following monocyte stimulation was significantly decreased in competitive athletes. Compared with the other two groups, the moderately trained athletes showed a significantly increased production of IFN-gamma upon T-cell stimulation. These data suggest that the immune system may profit from moderate endurance training by an increased capacity to generate IFN-gamma while the immune situation following repeated exhausting exercise of competitive athletes tends to deteriorate through downregulation of IFN-gamma and IL-12. PMID- 11258641 TI - Isokinetic strength and anaerobic power of elite, subelite and amateur French soccer players. AB - Information about the influence of different practice levels on physical characteristics of a large number of soccer players is lacking. Therefore we assessed muscular strength and anaerobic power of elite, subelite and amateur soccer players to clarify what parameters distinguish the top players from the less successful. We tested 95 soccer players from the French first division (elite), second division (subelite), and amateurs and determined the isokinetic strength of the knee extensor and flexor muscles at angular velocities from -120 degrees x s(-1) to 300 degrees x s(-1). Vertical jump, 10 m sprint, 30 m sprint and maximum ball speed during shooting were also measured. The elite players had higher knee flexor torque than the amateurs at all angular velocities (p < 0.05), except at 300 degrees x s(-1). The hamstring/quadriceps ratios proposed with two different methods were significantly lower in the amateur group than in the elite group (p < 0.05), except at 300 degrees x s(-1). Maximum ball speed during shooting and speed over 30 m sprint were not different between elite, subelite, and amateur players while speed over a 10 m sprint was significantly slower in amateur players and faster in the elite group (p < 0.05). Although performance in soccer is not determined only by measurable variables, professional players differ from amateurs in terms of knee flexor muscle strength and short-distance sprinting speed. Based on these findings we conclude that hamstring strength is extremely important in soccer players for joint stabilization during various tasks, notably in eccentric action. Further, short-sprinting performance may mirror actual game situations at high level and could be an important determinant of match-winning actions. PMID- 11258643 TI - Imaging overuse injury of the elbow in professional team handball players: a bilateral comparison using plain films, stress radiography, ultrasound, and magnetic resonance imaging. AB - The purpose of this study was to compare the manifestations of elbow stress due to repetitive valgus forces between the dominant and the non-dominant elbow in 40 uninjured elite team handball players using plain films, stress radiographs, ultrasound, and MRI examination. On comparative plain films generalized bony hypertrophy manifested by increased humeral diameter, and cortical hypertrophy of the humeral shaft of the dominant extremity was observed in all players. A significantly greater difference in medial joint space opening between stressed and unstressed elbows was measured in the dominant elbow compared with the non dominant elbow (0.41 +/- 0.59 mm). The ultrasonographic findings showed statistically significant bilateral differences in the thickness of the flexor pronator tendon (0.90 +/- 0.56 mm), extensor tendon (0.96 +/- 0.50 mm), triceps tendon (0.69 +/- 0.27 mm), and medial collateral ligament (0.47 +/- 0.24 mm): the values were systematically higher on the dominant side. US examination showed intra-articular effusions in 67% and small loose bodies in 33.3% of the players, exclusively in dominant elbows. MRI showed joint effusion in the same subjects as US, but loose bodies were only detected in half of the cases found by ultrasound. This study demonstrates that repetitive stress on the dominant extremities of handball players is responsible for physiologic and pathologic changes in the dominant elbow. PMID- 11258644 TI - Muscle soreness and damage parameters after prolonged intermittent shuttle running following acute vitamin C supplementation. AB - Exercise-induced free-radical production may be partly responsible for muscle soreness and damage following demanding exercise. A number of studies have investigated the effect of antioxidant supplementation although there is a paucity of information regarding vitamin C. Therefore the aim of the present study was to investigate the effect of vitamin C supplementation on exercise induced muscle soreness and damage. Nine habitually active males consumed a 1 g dose of vitamin C 2 h before exercise, and on another occasion consumed an identical placebo. The exercise comprised a 90 min intermittent shuttle-running test, which was designed to simulate the multiple-sprint sports. Vitamin C supplementation increased plasma concentrations of vitamin C before exercise, and plasma concentrations continued to increase during the shuttle-run to reach a peak of approximately 200 micromol x l(-1) immediately after exercise. However, muscle soreness, and markers of both muscle damage (creatine kinase and aspartate aminotransferase) and lipid peroxidation (malondialdehyde) were elevated to an equal extent after exercise in placebo and supplemented trials. Therefore acute supplementation with vitamin C had no beneficial effects although it is possible that such short-term vitamin C supplementation was ineffective because it occurred at an inappropriate time. PMID- 11258645 TI - Creatine loading does not impact on stroke performance in tennis. AB - The effect of acute creatine supplementation on stroke quality was investigated during simulated match play. Well-trained tennis players reported to the test center on two occasions. On each occasion they performed the Leuven Tennis Performance Test (LTPT) and a 70 m shuttle run (SHR). During 5 days prior to each test session they received in random order and according to a double-blind cross over study design either oral creatine supplements (4 x 5 g per day) or placebo. The two experimental periods were separated by a 5-week washout period. Stroke quality was evaluated during the LTPT by means of registration of error rate and measurement of ball velocity and precision of lateral and longitudinal ball placement. Compared with placebo, creatine supplementation did not significantly impact on either power or precision of first and second services, baseline strokes in neutral and defensive rallies, and volleys. Shuttle run time was 19.87 +/- 0.30 sec during placebo versus 19.85 +/- 0.27 sec during creatine treatment. Acute creatine supplementation does not enhance stroke performance or sprint power in match-like conditions in elite tennis players. PMID- 11258646 TI - Individual anaerobic threshold: methodological aspects of its assessment in running. AB - The present study was designed in order to examine the objectivity and reliability of the individual anaerobic threshold (IAT) as well as its resistance against several interfering factors: missing exhaustion, preliminary exercise, longer step duration, and lower speed increment. 87 male and 24 female runners and triathletes were examined. They performed both the original test procedure (IAT(3/2); 3 min step duration, 2 km x h(-1) increment until volitional exhaustion) and either a retest or one of several alternative test procedures (submaximal, preliminary exercise, 5 min step duration, 1 km x h(-1) increment) to be compared with IAT(3/2). The graphic determination of the IAT is characterized by a low inter-observer-variability without significant differences between 4 independent blinded examiners. Both the lactate performance curve (p = 0.07) as well as the heart rate performance curve (p = 0.05) show a slight shift to higher velocities during the retest. The IAT shows an identical running velocity during the retest, the heart rate tends to be lower, and the lactate concentration is significantly (by 0.26 mmol x l(-1); p < 0.05) lower than during the first test. Both a low degree of exhaustion (3-6 mmol x l(-1) lactate; IAT(3/2) shortened by 180 s) and an extension of the step duration from 3 to 5 min do not lead to significantly different velocities at the IAT. Moderate preliminary exercises (approximately 4 mmol x l(-1) lactate) do not influence the velocity at the IAT, the heart rate is significantly higher. A reduction of the speed increment from 2 to 1 km x h(-1) significantly increases the velocity at the IAT by 6%. It is concluded that the determination of the IAT is highly objective, reliable and insensitive to changes of the incremental graded testing protocol, such as a previous warm-up, an extension of the step duration from 3 to 5 min as well as a lower degree of exhaustion. Significant differences may only arise from changes in the speed increment. PMID- 11258647 TI - Comparison of critical swimming velocity and velocity at lactate threshold in elite triathletes. Int J Sports Med 2000; 21: 366-368 re: Martin L. Whyte GP. PMID- 11258648 TI - Effects of topical budesonide treatment on glucocorticoid receptor mRNA down regulation and cytokine patterns in nasal polyps. AB - The effects of a topically applied corticosteroid, budesonide, on the expression of glucocorticoid receptor (GR) mRNA and regulation of pro-inflammatory cytokine patterns in patients with nasal polyps were evaluated. All patients were eligible for surgical polypectomy, and a majority of them had been treated with nasal steroids. Patients were given 400 microg b.i.d. (group A, n = 11), 200 microg b.i.d. (group B, n = 10), or no treatment (group C, n = 15) during two months before polypectomy. Morning serum cortisol was analyzed on the day of surgery. Surgically removed polyps were taken for analysis of GR mRNA expression by solution hybridization. Remaining tissue was cryostat-sectioned, whereafter quantification of the cytokines interleukin 1beta, interleukin 2, interleukin 4, interleukin 5, interleukin 6, interleukin 10, tumor necrosis factor alpha, and interferon gamma was made by immunohistochemistry and digitized image analysis. No significant differences among the three groups were found for any of the parameters investigated. CONCLUSION: nasal polyps do not respond with down regulation of GR mRNA or cytokines following topical corticosteroid treatment. The proposed corticosteroid resistance may be inherent, or induced by a change of local tissue bioavailability. PMID- 11258649 TI - Use of the Rhinosinusitis Disability Index (RSDI) in rhinologic disease. AB - Chronic rhinosinusitis has been shown to have an adverse impact on the quality of life in those afflicted as determined by a number of standardized outcome tools. We have utilized a standardized, statistically validated, disease-specific tool, the Rhinosinusitis Disability Index (RSDI), to investigate the disability suffered by 292 consecutive patients with nine unique rhinologic diagnoses, including chronic rhinosinusitis. Physical, functional, and emotional domains were assessed. We have found that individuals with rhinologic disease in general have lower physical scores, followed by functional scores and emotional scores. Individuals with chronic rhinosinusitis and allergic rhinitis have the greatest level of disability, while those with aspirin triad are least affected. PMID- 11258650 TI - Safety and efficacy of endoscopic repair of CSF leaks and encephaloceles: a survey of the members of the American Rhinologic Society. AB - The purpose of this article is to review the endoscopic management of cerebrospinal fluid (CSF) leaks and encephaloceles, with particular emphasis on safety and efficacy, by retrospective assessment utilizing the results of a mailed questionnaire. Surveys were mailed to members of the American Rhinologic Society with practices in both academic centers and/or private settings. Survey results were then assessed and tabulated. There were 635 mailings, with 197 responses (31%). Seventy-two (36% of respondents) indicated that they performed endoscopic management of CSF leaks and encephaloceles, while 125 (64% of respondents) did not. Respondents reported approximately 522 cases of CSF leaks and approximately 128 cases of encephaloceles managed by endoscopy. Success rates after a single procedure were estimated at 90% for CSF leaks and 93% for encephaloceles. Success rates after a secondary procedure were estimated at 86% and 97%, respectively; 29% of respondents have, at some point, made a referral to neurosurgery. A total of 13 complications related to endoscopic repairs were reported (2.5%). For CSF leak repair, complications included seizures, 0.2%; meningitis, 1.1%; and one reported case each of cavernous sinus thrombosis, temporary visual problems, sinusitis, and intracranial hypertension/bleed. There was only one reported death in the approximately 522 cases. Eleven complications following encephalocele repairs (8.5%) included seizures, 3.1%; meningitis, 2.3%; and one reported case each of brain abscess, sinusitis, false aneurysm of middle cerebral artery, and mild dizziness. No deaths following encephalocele repair were reported. The endoscopic management of CSF leaks and encephaloceles has become increasingly popular and has proven to have low morbidity and mortality with high success. Overall, our results confirm that in the hands of the skilled endoscopist, endoscopic management of CSF leaks and encephaloceles is highly efficacious and has a very low incidence of significant complication. PMID- 11258651 TI - Meningiomas of the paranasal sinuses. AB - Extracranial meningiomas are rare tumors, comprising approximately 2% of all meningiomas. Previously reported sites include the orbit, parapharyngeal space, and rarely, the paranasal sinuses. A retrospective chart review of patients with meningiomas was performed over the last 25 years, and three patients were identified with meningiomas involving the paranasal sinuses. The locations included the frontal sinus, the ethmoid sinus, and the sphenoid sinus. Presenting symptoms included facial pain and nasal obstruction; two patients noted facial swelling. Diagnosis was established via endoscopic transnasal biopsy in two patients. Computed tomographic (CT) guided biopsy was utilized to confirm the diagnosis in the third patient. Surgical extirpation was successfully performed with tumors arising from the ethmoid and frontal sinuses. The patient with neoplasm in the sphenoid sinus underwent radiation therapy. Extracranial meningiomas of the paranasal sinuses are rare tumors that may present a diagnostic and therapeutic challenge. We present three cases and discuss the clinical presentation, radiographic findings, diagnostic evaluation, and treatment options. PMID- 11258652 TI - Dilated pupil during endoscopic sinus surgery: what does it mean? AB - Endoscopic sinus surgery has become the standard of care for the surgical management of chronic sinus disease. Sinus disease and its surgical treatment carry the risk of orbital complications, irrespective of the approach. Orbital complications associated with sinus surgery include nasolacrimal duct damage, extraocular muscle injury, intraorbital hemorrhage/emphysema, and direct optic nerve damage, resulting in blindness. The finding of an unequal pupil at the end of a procedure would be a cause of considerable concern, but it is most likely due to the topical contamination of the eye with a mydriatic pharmacological agent commonly used in endoscopic sinus surgery. PMID- 11258653 TI - Detection of the nasal cycle. AB - The nasal cycle is a fluctuation of nasal patency due to the stages of congestion and decongestion of the nasal mucosa on both the right and left nasal conchae. We compared the effectiveness of the rhinostereometer in detecting the presence of a nasal cycle with the acoustic rhinometer whose effectiveness we have demonstrated in previous studies. The rhinostereometer measures the horizontal range of the most anterior portion of the inferior turbinate. The acoustic rhinometer measures the volume and various cross-sectional areas of the nostril using a pulse emitted from a sound tube. Among some of the subjects tested, it was found that rhinostereometer and acoustic rhinometer measurements of nasal patency correlated reasonably well with r values up to 0.78. The overall correlation between rhinostereometry and acoustic rhinometry was not as strong at r = 0.36. Observed variations between rhinostereometry and acoustic rhinometry could be a result of certain confounding variables that may have altered the nasal cycle between measurements. PMID- 11258654 TI - Elevation of nasal mucosal temperature increases the ability of the nose to warm and humidify air. AB - The nose functions to warm and humidify inspired air. The factors that influence these functions have been studied to a limited degree. We have developed a method for measuring the temperature and relative humidity of the air before and after nasal conditioning to study nasal function. In this experiment we studied the effects of raising the mucosal surface temperature by immersion of the feet in warm water. Six subjects (avg. age = 27.0 years) were randomized to immersion of the feet in 30 degrees C and 40 degrees C water. The nasal mucosal temperature increased significantly from the 32.2+/-1.3 degrees C during immersion in the 30 degrees C water to the 33.1+/-1.2 degrees C during immersion in 40 degrees water (p < 0.05). No significant difference in nasal volume was noted between the 30 degrees (17.8+/-4.5 cc) and the 40 degrees (17.7+/-5.3 cc) immersions. There was a significant increase in the conditioning capacity of the nose (as measured by total water content of inspired air) in response to cold-air challenge during the 40 degrees immersion (1669+/-312 mg water) when compared to the 30 degrees immersion (1324+/-152 mg water). From these data we deduce that warming of the nasal mucosa improves the ability of the nose to condition inspired air without a significant change in the volume of the nasal cavity. PMID- 11258655 TI - Leukotriene B4 levels in rabbit maxillary sinusitis: limitations of the current model. AB - Since the late 1980s, the rabbit model for sinusitis has been widely used for experimental studies on sinusitis; however, the clinical relevance of these experimental data has been questioned. To elucidate the role of leukotrienes in the pathogenesis of sinusitis, leukotriene B4 (LTB4) levels were determined in acute Streptococcus pneumoniae sinusitis in this model. The rabbit model for acute maxillary sinusitis was utilized. Briefly, the right maxillary ostium of each New Zealand white rabbit was occluded with cyanoacrylate under general anesthesia. Twenty-four hours after occlusion, the occluded sinus received an inoculation of 10(8) Streptococcus pneumoniae (ATCC 10813) or a sham inoculation of saline alone. Rabbits were then sacrificed one week later, and the maxillary sinus mucosae were harvested. Leukotriene B4 levels were determined by ELISA assay. LTB4 levels in the sinuses inoculated with bacteria tended to be higher; however, statistical analysis did not reveal significant differences between the experimental and control groups. It is possible to reliably assess leukotriene B4 levels in this model of sinusitis. Although the data suggest a trend for elevated LTB4 levels, statistical analysis did not support this conclusion. The study also demonstrated significant limitations in the current rabbit model for sinusitis; that is, the standard human sinus bacterial pathogens are minimally pathogenic in rabbit sinuses and the small size of the sinus limits the material available for assay. Further modifications of the model are necessary. After such adjustments, the role of leukotrienes in sinusitis may be further explored. PMID- 11258656 TI - Efficacy of endoscopic sinus surgery in the management of patients with asthma and chronic sinusitis. AB - An association between chronic sinusitis and asthma has been noted for many years, although the precise nature of the relationship is poorly understood. Earlier studies, using traditional surgical techniques, have demonstrated subjective improvement in asthmatic complaints. Reports demonstrating improvement following endoscopic sinus surgery for chronic sinusitis are rare. To report our experience with endoscopic sinus surgery and asthmatics, we reviewed the charts of 75 consecutive patients with asthma and chronic sinusitis who underwent endoscopic sinus surgery between 1994 and 1996. Study criteria included the following: chronic sinusitis, one year preoperative and one year postoperative follow-up from endoscopic sinus surgery, and asthma requiring inhaled steroids and oral prednisone for control. Many patients required prednisone bursts for control of asthma. Number of days and total dose of oral prednisone were used as objective measures of asthma control. Number of weeks of antibiotics was used as a relative measure of sinusitis. Fourteen of the 15 patients meeting study criteria decreased their postoperative prednisone requirement by total number of days (preoperative 84 versus postoperative 63 days [p < 0.0001]). Postoperatively, patients required an average of 1300 mg less oral prednisone (p < 0.033). Antibiotic use also decreased, with an average use of antibiotic nine weeks preoperatively versus seven weeks postoperatively (p < 0.045). This study provides corroborative objective evidence that, at least in the short term, endoscopic sinus surgery is efficacious in the management of patients with chronic sinusitis and asthma. PMID- 11258657 TI - Effects of long-term induced ostial obstruction in the rabbit maxillary sinus. AB - One of the widely proposed theories for mucocele formation is sinus ostial obstruction. Accordingly, this study was undertaken to investigate the long-term effects of ostial obstruction in the rabbit maxillary sinus and its potential role in the pathogenesis of mucoceles. Maxillary sinus ostial obstruction was induced on one side in eight Pasteurella-free White New Zealand rabbits using Histoacryl. The rabbits were housed in a Pasteurella-free zone for 24 weeks. At re-exploration, only three of the eight maxillary sinuses where ostial obstruction was induced showed pressure recording consistent with ostial obstruction. Mucociliary clearance activity was assessed using India ink. Swabs for culture were taken from the infected maxillary sinuses. Mucosal specimens for histopathological examination were harvested from one of the maxillary sinuses with obstructed ostium as well as from another sinus with nonobstructed ostium. The three maxillary sinuses with obstructed ostia showed gross evidence of infection and deranged mucociliary clearance, but no mucocele formation. Based on the findings of this study it is concluded that long-term ostial obstruction indeed plays a role in the pathogenesis of chronic sinusitis, but it did not induce mucocele formation in the rabbit maxillary sinus. PMID- 11258658 TI - Substance P immunoreactive sensory axons as a subset of the total axonal population in the maxillary sinus of the rabbit: a characterization of normal and infected mucosa. AB - Substance P (SP), one of the neuropeptides released from sensory nerves, is thought to mediate neurogenic inflammation. Although SP immunoreactive axons have been described in the sinus mucosa, no attempt has been made to characterize SP fibers as a subset of all axons present in the sinus mucosa. In addition, no study to date has characterized the changes in infected sinus mucosa. The maxillary sinus mucosa of New Zealand white rabbits was harvested from control animals and in animals with induced maxillary sinusitis. Immunohistochemical staining of the sinus mucosa for both Protein Gene Product 9.5 (PGP), a nonspecific marker for all nerves, and for SP was performed on 11 animals: 3 controls and 8 infected. In sinus mucosa from the control rabbits, <50% of all axons labeled by PGP were immunoreactive for SP. In infected mucosa, the absolute number of axons found by PGP staining decreased and nearly all of these remaining fibers were also immunoreactive for SP. We conclude that the phenotypical labeling of nerve fibers seen in normal mucosa is altered by bacterial-induced infection. PMID- 11258659 TI - Nasal polyposis and allergy: is there a correlation? AB - Nasal polyposis (NP) is a chronic inflammatory disease of the nasal mucosa. The etiology and formation of NP are still not elucidated and have been debated for many years. The objective of the present study was to investigate the role of nasal allergy in the development of NP. The following aspects were analyzed: age, sex, and patient's symptoms; correlation between asthma, aspirin intolerance, and NP; serum immunoglobulin levels and eosinophilia; and concentration of interleukins 1beta, 3, and 4 in NP. Thirty-nine patients with NP were selected, 13 of them allergic and 26 non-allergic. A control group of 11 individuals was also studied. The concentrations of interleukins 1beta, 3, and 4 were measured by enzyme-linked immunosorbent assay (ELISA). There was a higher incidence of NP after the fourth decade of life and among men. We found no correlation of asthma or aspirin intolerance with the presence or absence of allergy. Serum levels of IgE and eosinophils were significantly higher in patients with allergy and NP and the concentrations of interleukins 3 and 4 were positively correlated with NP. There was no difference in interleukin 3 and 4 concentration between the non allergic group with NP and the control group, suggesting that these interleukins do not play an important role in the etiology and formation of NP. These results suggest that the immunologic pathway involved in the etiology of NP is differentfrom the one correlated with allergy (IgE-mediated). PMID- 11258660 TI - Synthetic peptides and fluorogenic substrates related to the reactive site sequence of Kunitz-type inhibitors isolated from Bauhinia: interaction with human plasma kallikrein. AB - We have previously described Kunitz-type serine proteinase inhibitors purified from Bauhinia seeds. Human plasma kallikrein shows different susceptibility to those inhibitors. In this communication, we describe the interaction of human plasma kallikrein with fluorogenic and non-fluorogenic peptides based on the Bauhinia inhibitors' reactive site. The hydrolysis of the substrate based on the B. variegata inhibitor reactive site sequence, Abz-VVISALPRSVFIQ-EDDnp (Km 1.42 microM, kcat 0.06 s(-1), and kcat/Km 4.23 x 10(4) M(-1) s(-1)), is more favorable than that of Abz-VMIAALPRTMFIQ-EDDnp, related to the B. ungulata sequence (Km 0.43 microM, kcat 0.00017 s(-1), and kcat/Km 3.9 x 10(2) M(-1) s(-1)). Human plasma kallikrein does not hydrolyze the substrates Abz-RPGLPVRFESPL-EDDnp and Abz-FESPLRINIIKE-EDDnp based on the B. bauhinioides inhibitor reactive site sequence, the most effective inhibitor of the enzyme. These peptides are competitive inhibitors with Ki values in the nM range. The synthetic peptide containing 19 amino acids based on the B. bauhinioides inhibitor reactive site (RPGLPVRFESPL) is poorly cleaved by kallikrein. The given substrates are highly specific for trypsin and chymotrypsin hydrolysis. Other serine proteinases such as factor Xa, factor XII, thrombin and plasmin do not hydrolyze B. bauhinioides inhibitor related substrates. PMID- 11258661 TI - Identification and characterization of an aromatic amino acid decarboxylase from the filarial nematode, Dirofilaria immitis. AB - A novel secreted aromatic amino acid decarboxylase-like molecule was identified in the excretory/secretory products of L3/L4 larvae as well as in an extract of adult Dirofilaria immitis. The secretion of the enzyme was developmentally regulated. Peak enzyme activities were detected in the culture medium before and after the molting of L3 larvae in vitro. The enzyme was purified from D. immitis adult extracts and the excretory/secretory products of L3/L4 larvae using different chromatographic methods followed by isoelectric focusing and SDS-PAGE. The enzyme has a molecular mass of 48 kDa and a pI of 5.6, and shows a specific enzymatic activity towards the aromatic amino acid substrates phenylalanine, tyrosine and tryptophan. The enzyme's activity did not show an absolute requirement for exogenous pyridoxal-5-phosphate. However, addition of pyridoxal-5 phosphate at 5 microM in the reaction increased the enzyme activity greatly. The enzyme had the ability to catalyze the formation of dopamine from L-dopa. Studies on the effects of inhibitors on the enzyme activity showed that the enzyme was sensitive to Pefabloc and p-chloromercuribenzoic acid, but not to diisopropyl flurophosphate. The Km values of the enzyme for H-Phe-AMC, H-Tyr-AMC and H-Trp AMC were calculated to be 32.1 microM, 35.1 microM and 29.1 microM, respectively. PMID- 11258662 TI - Molecular cloning and pharmacological characterization of the canine B1 and B2 bradykinin receptors. AB - The dog is a valuable animal model in the study of the physiological role of both the B1 and B2 bradykinin receptors. To more thoroughly characterize the pharmacological properties of the canine kinin receptors we isolated the cDNA sequence encoding the B1 and B2 bradykinin receptor subtypes and overexpressed them in Chinese hamster ovary (CHO) cells. The cDNA sequence of the canine B1 bradykinin receptor encodes a protein comprised of 350 amino acids that is 76% identical to the human B1 bradykinin receptor. The cDNA sequence of the canine B2 bradykinin receptor encodes a protein of 392 amino acids that is 81% identical to the human B2 bradykinin receptor. The amino acid sequence of the canine B1 and B2 receptors are 35% identical. Pharmacological studies of the cloned receptors revealed that the agonist affinity of the dog B1 receptor is similar to the rodent B1 receptors, and differs from the human form in that there is no preference for the presence of the N-terminal Lys residue of [des-Arg10]Lys bradykinin. Significantly, the B1 receptor antagonist [des-Arg9,Leu8]BK behaves as partial agonist on the cloned dog B1 receptor. The dog B2 receptor exhibits the 'classical' pharmacological properties of this receptor subtype. PMID- 11258663 TI - Ligand-mediated regulation of kinin receptors in the rabbit. AB - The kinin B1 receptors (B1Rs), stimulated by des-Arg9-kinins, are completely inducible, notably following treatment of animals with bacterial lipopolysaccharide. Several studies based on cultured cells have suggested a form of autoregulation of kinin receptors, because B2 receptor (B2R) or B1R stimulation could transcriptionally upregulate B1R expression. The B2R may rather be downregulated in inflammatory conditions. A rabbit B2R-green fluorescent protein (GFP) conjugate stably expressed in HEK 293 cells was rapidly internalized in response to the agonist bradykinin. Ligand-induced receptor cycling was documented applying confocal microscopy. The results confirm agonist induced B2R endocytosis, but extensive recycling to the cell membrane does not support agonist-induced downregulation. PMID- 11258664 TI - Activation of sphingosine kinase by the bradykinin B2 receptor and its implication in regulation of the ERK/MAP kinase pathway. AB - Sphingosine kinase phosphorylates sphingosine to generate sphingosine 1 phosphate, a phospholipid that has been implicated in signaling by a number of transmembrane receptors and was recently shown to act as a ligand for a specific class of G protein-coupled receptors. Here we show that the G protein-coupled bradykinin B2 receptor activates sphingosine kinase leading to a time- and dose dependent elevation of cellular sphingosine 1-phosphate levels that was blocked by the sphingosine kinase inhibitor dihydrosphingosine. Furthermore, increasing doses of this inhibitor partially affected the bradykinin-mediated ERK/MAP kinase activation and fully blocked the protein kinase C-independent component of the signaling pathway from the B2 receptor to the ERK/MAP kinase cascade. Overexpression of sphingosine kinase did not additionally increase the bradykinin induced ERK/MAP kinase activity, indicating a permissive rather than activating role of sphingosine 1-phosphate in B2 receptor-mediated mitogenic signaling. PMID- 11258665 TI - Novel roles of kallistatin, a specific tissue kallikrein inhibitor, in vascular remodeling. AB - We have purified, cloned and characterized kallistatin, a tissue kallikrein binding protein (KBP) in humans and rodents. Kallistatin is a unique serine proteinase inhibitor (serpin) with Phe-Phe residues at the P2 and P1 positions. Structural and functional analysis of kallistatin by site-directed mutagenesis and protein engineering indicate that wild-type kallistatin is selective for tissue kallikrein. Kallistatin is expressed and localized in endothelial and smooth muscle cells of blood vessels and has multiple roles in vascular function independent of the tissue kallikrein-kinin system. First, kallistatin induces vasorelaxation of isolated aortic rings and reduces renal perfusion pressure in isolated rat kidneys. Transgenic mice overexpressing rat kallistatin are hypotensive, and adenovirus-mediated gene delivery of human kallistatin attenuates blood pressure rise in spontaneously hypertensive rats. Second, kallistatin stimulates the proliferation and migration of vascular smooth muscle cells in vitro and neointima formation in balloon-injured rat arteries. Third, kallistatin inhibits the proliferation, migration and adhesion of endothelial cells in vitro and angiogenesis in the rat model of hindlimb ischemia. These results demonstrate novel roles of kallistatin in blood pressure regulation and vascular remodeling. PMID- 11258666 TI - Signal transduction from bradykinin, angiotensin, adrenergic and muscarinic receptors to effector enzymes, including ADP-ribosyl cyclase. AB - Muscarinic acetylcholine receptors in NG108-15 neuroblastoma x glioma cells, and beta-adrenergic or angiotensin II receptors in cortical astrocytes and/or ventricular myocytes, utilize the direct signaling pathway to ADP-ribosyl cyclase within cell membranes to produce cyclic ADP-ribose (cADPR) from beta-NAD+. This signal cascade is analogous to the previously established transduction pathways from bradykinin receptors to phospholipase Cbeta and beta-adrenoceptors to adenylyl cyclase via G proteins. Upon receptor stimulation, the newly-formed cADPR may coordinately function to upregulate the release of Ca2+ from the type II ryanodine receptors as well as to facilitate Ca2+ influx through voltage dependent Ca2+ channels. cADPR interacts with FK506, an immunosuppressant, at FKBP12.6, FK506-binding-protein, and calcineurin, or ryanodine receptors. cADPR also functions through activating calcineurin released from A-kinase anchoring protein (AKAP79). Thus, some G(q/11)-coupled receptors can control cADPR dependent modulation in Ca2+ signaling. PMID- 11258667 TI - Kinins 1925-2000. PMID- 11258668 TI - Classification of kinin receptors. AB - This minireview is divided into three parts: the first part refers to the characterization and classification of kinin receptors using agonists and antagonists in isolated tissues (classical pharmacology). Two kinin receptors have been considered on the basis of their distinct pharmacology, namely the B1 receptor of the rabbit aorta (rank order of potency of agonists: LysdesArg9BK > desArg9BK > or = LysBK > BK; apparent affinities of antagonists Lys[Leu8]desArg9BK (pIC50 8.4) > [Leu8]desArg9BK (pIC50 7.4) >>> HOE 140, a B2 receptor antagonist, pIC50<5.0), and the B2 receptor of the rabbit jugular vein (potency of agonists: LysBK = BK >>> LysdesArg9BK = desArg9BK and HOE 140 (pIC50 9.0) >>> Lys[Leu8]desArg9BK, pIC50<5.0). The second part describes species related B1 receptor subtypes, demonstrated by different pharmacological profiles of agonists and antagonists: human, rabbit and pig subtypes (LysdesArg9BK >> desArg9BK and Lys[Leu8]desArg9BK > [Leu8]desArg9BK) and dog, rat, mouse and hamster B1 receptors (desArg9BK = LysdesArg9BK and [Leus]desArg9BK = Lys[Leu8]desArg9BK). Affinities of agonists and antagonists in some species (man, rabbit, pig) are significantly increased (at least 10-fold) by the presence of a Lys at their N-terminus. The last part describes species-related B2 receptor subtypes supported by results obtained with non-peptide receptor agonists (FR 190997) and antagonists (FR 173657). While BK acts as a full agonist in man, rabbit and pig, FR 190997 behaves as a full agonist on human, as partial agonist on rabbit, and as pure antagonist on pig B2 receptors. Various hypotheses are considered to interpret these findings. PMID- 11258669 TI - Metabolism-resistant bradykinin antagonists: development and applications. AB - Bradykinin plays many roles in normal and pathological physiology, but rapid enzymatic degradation made elucidation of its functions extremely difficult. Development of effective degradation-resistant antagonists made it possible to delineate these roles and to open the way for development of new drugs to control pathology due to excess production of bradykinin. Presently available peptide bradykinin antagonists are extremely potent, are completely resistant to enzymatic degradation, and are orally available. Non-peptide bradykinin antagonists have also been discovered. Development of bradykinin antagonists as drugs for cancer, inflammation and trauma is anticipated. PMID- 11258670 TI - Kinins, receptors, kininases and inhibitors--where did they lead us? AB - Based on studies presented here and other published experiments performed with surviving tissue preparations, with transfected cells and with cells that constitutively express the human angiotensin I converting enzyme ACE and B2 receptors, we concluded the following: ACE inhibitors and other endogenous peptides that react with the active site of ACE potentiate the effect of bradykinin and its ACE resistant peptide congeners on the B2 receptor. They also resensitize receptors which had been desensitized by the agonist. ACE and bradykinin receptors have to be sterically close, possibly forming a heterodimer, for the ACE inhibitors to induce an allosteric modification on the receptor. When ACE inhibitors augment bradykinin effects, they reduce the phosphorylation of the B2 receptor. The primary actions of bradykinin on the receptor are not affected by protein kinase C or phosphatase inhibitors, but the potentiation of bradykinin or the resensitization of the receptor by ACE inhibitors are abolished by the same inhibitors. The results with protein kinase C and phosphatase inhibitors indicate that another intermediate protein may be involved in the processes of signaling induced by ACE inhibitors, and that ACE inhibitors affect the signal transduction pathway triggered by bradykinin on the B2 receptor. PMID- 11258671 TI - Bradykinin signalling to MAP kinase: cell-specific connections versus principle mitogenic pathways. AB - Mitogenic signalling pathways from G protein-coupled receptors (GPCRs) to the mitogen-activated protein kinase (MAPK) cascade may involve alpha- or betagamma subunits of heterotrimeric G proteins, receptor or non-receptor tyrosine kinases, adaptor molecules, phosphoinositide 3-kinases, protein kinase C, and probably other proteins. The majority of models describing the connection of different signalling proteins within a mitogenic pathway are based on experimental data obtained by co- and overexpression of epitope-tagged MAPK together with the respective GPCR and other signalling proteins of interest in transfectable cell lines. Here the link of the bradykinin B2 receptor (B2R) to MAPK in the COS-7 cell expression system is compared with mitogenic signalling pathways of bradykinin in various tumour cell lines. It becomes evident that in natural or tumour cells expressing individual amounts and different isoforms of signalling proteins completely other relations between B2R and MAPK may exist than in COS-7 cells, suggesting a high degree of cellular specificity in mitogenic signalling. PMID- 11258672 TI - The expanded human kallikrein (KLK) gene family: genomic organisation, tissue specific expression and potential functions. AB - The tissue kallikreins are serine proteases encoded by highly conserved multi gene families. The rodent kallikrein (KLK) families are particularly large, consisting of 13-26 genes clustered in one chromosomal locus. It has been recently recognised that the human KLK gene family is of a similar size (15 genes) with the identification of another 12 related genes (KLK4-KLK15) within and adjacent to the original human KLK locus (KLK1-3) on chromosome 19q13.4. The structural organisation and size of these new genes is similar to that of other KLK genes except for additional exons encoding 5' or 3' untranslated regions. Moreover, many of these genes have multiple mRNA transcripts, a trait not observed with rodent genes. Unlike all other kallikreins, the KLK4-KLK15 encoded proteases are less related (25-44%) and do not contain a conventional kallikrein loop. Clusters of genes exhibit high prostatic (KLK2-4, KLK15) or pancreatic (KLK6-13) expression, suggesting evolutionary conservation of elements conferring tissue specificity. These genes are also expressed, to varying degrees, in a wider range of tissues suggesting a functional involvement of these newer human kallikrein proteases in a diverse range of physiological processes. PMID- 11258673 TI - Kinins and epithelial ion transport in the alimentary tract. AB - Kinin effects on epithelial electrogenic ion transport are reviewed, with reference to the alimentary tract. The transported ion is usually chloride, but some epithelia also transport bicarbonate. The key components of the transport system are the sodium-potassium-chloride cotransporter, Na+-K+ ATPase (both located basolaterally) and the CFTR chloride channel (located apically). Activation of K+-channels in both membranes may secondarily affect the anion transport mechanism. The types of kinin receptors that cause chloride secretion, the second messengers involved and the possible functional responsibilities of the kinin-activated secretory mechanism are discussed. PMID- 11258675 TI - Role of the light chain of high molecular weight kininogen in adhesion, cell associated proteolysis and angiogenesis. AB - Cleavage of high molecular weight kininogen (HK) by plasma kallikrein results in a light chain and a heavy chain (HK). The light chain has two domains: D6, which binds (pre)kallikrein, and D5, which binds to anionic surfaces, including heparin as well as zinc. Initially, HK was thought to be important for surface-activated coagulation. HKa or D5 binds to the urokinase receptor on endothelial cells, thereby enhancing the conversion of prourokinase to urokinase by kallikrein, and, thus, cell-associated fibrinolysis. HKa or D5 is antiadhesive by competing with vitronectin binding to the urokinase receptor and/or forming a complex with vitronectin. D5 inhibits endothelial cell migration, proliferation, tube formation and angiogenesis, thus modulating inflammation and neovascularization. PMID- 11258674 TI - Role of the renal kallikrein-kinin system in the development of salt-sensitive hypertension. AB - The role of the renal kallikrein-kinin system in the development of salt sensitive hypertension was studied using mutant kininogen-deficient Brown-Norway Katholiek (BN-Ka) rats, which generate no kinin in their urine, and other hypertensive rat models. It was found that ingestion of a low sodium diet or infusion of NaCl in doses slightly above 0.15 M caused hypertension and sodium accumulation in erythrocytes and the cerebrospinal fluid of kininogen-deficient BN-Ka rats. Development of hypertension in the deoxycorticosterone-acetate-salt model was completely prevented by administration of a newly discovered inhibitor, ebelactone B, of carboxypeptidase Y-like exopeptidase (an urinary kininase). The urinary kallikrein excretion of spontaneously hypertensive rats was lower than that of Wistar Kyoto rats at 4 weeks of age and did not increase by administration of furosemide, a diuretic agent, although approximately 50% of the diuretic action of this agent was dependent upon the renal kallikrein-kinin system in normal rats. In conclusion, the renal kallikrein-kinin system works as a safety valve for excess sodium intake. PMID- 11258676 TI - Activation of the kinin-forming cascade on the surface of endothelial cells. AB - Activation of the plasma kallikrein-kinin forming cascade takes place upon incubation with human umbilical vein endothelial cells. The mechanism by which initiation occurs is uncertain. Zinc-dependent binding of plasma proteins to gC1qR, cytokeratin 1, and perhaps u-PAR is requisite for activation to take place. We demonstrate here that during a 2 hour incubation time plasma deficient in either factor XII or high molecular weight kininogen (HK) fails to activate, as compared to normal plasma, but with more prolonged incubation, factor XII deficient plasma gradually activates while HK-deficient plasma does not. Our data support both factor XII-dependent (rapid) and factor XII-independent (slow) mechanisms; the latter may require a cell-derived protease to activate prekallikrein and the presence of zinc ions and HK. PMID- 11258677 TI - Kallikrein and kinin receptor expression in inflammation and cancer. AB - The kallikrein family of serine proteases has been investigated in many inflammatory disorders as molecular mapping, gene characterisation and cloning of kinin receptor genes have unfolded experimentally. In the molecular events of the inflammatory response the kallikrein cascade plays a significant role, since it is considered to initiate and maintain systemic inflammatory responses and immune modulated disorders. A primary event is the chemotactic attraction of neutrophils which deliver the kallikrein-kinin cascade to sites of cellular injury and carcinogenic transformation of cells. The present study establishes the casual involvement of the kallikrein cascade in infection, inflammatory joint disease, acute transplant rejection, renal glomerular diseases, angiogenesis and carcinoma. We provide strong evidence for new or enhanced expression of kinin B1 receptors in inflammation, and additionally the induction of kallikrein genes in angiogenesis and carcinoma. The results provide insights into possible roles of kallikrein inhibitors and kinin receptor antagonists. PMID- 11258678 TI - Altered neutrophil homeostasis in kinin B1 receptor-deficient mice. AB - The kallikrein-kinin system is activated during inflammation and plays a major role in the inflammatory process. One of the main mechanisms of kinin action includes the modulation of neutrophil function employing both receptors for kinins, B1 and B2. In this report we show by the use of B1 receptor-deficient mice that neutrophil migration in inflamed tissues is dependent on kinin B1 receptors. However, there is no change in circulating leukocyte number and composition after genetic ablation of this receptor. Furthermore, apoptosis of neutrophils necessary for the resolution of persistent inflammatory processes is impaired in mice lacking the B1 receptor. We also show that this receptor is expressed on neutrophils, thus it may be directly involved in the induction of apoptosis in these cells after prolonged activation at inflamed sites. In conclusion, our data show that the kinin B1 receptor modulates migration and the life span of neutrophils at sites of inflammation and may be therefore an important drug target in the therapy of inflammatory diseases. PMID- 11258680 TI - Ragweed pollinosis: answers awaiting explanations. PMID- 11258679 TI - Cystatins as calpain inhibitors: engineered chicken cystatin- and stefin B kininogen domain 2 hybrids support a cystatin-like mode of interaction with the catalytic subunit of mu-calpain. AB - Within the cystatin superfamily, only kininogen domain 2 (KD2) is able to inhibit mu- and m-calpain. In an attempt to elucidate the structural requirements of cystatins for calpain inhibition, we constructed recombinant hybrids of human stefin B (an intracellular family 1 cystatin) with KD2 and deltaL110 deletion mutants of chicken cystatin-KD2 hybrids. Substitution of the N-terminal contact region of stefin B by the corresponding KD2 sequence resulted in a calpain inhibitor of Ki = 188 nM. Deletion of L110, which forms a beta-bulge in family 1 and 2 cystatins but is lacking in KD2, improved inhibition of mu-calpain 4- to 8 fold. All engineered cystatins were temporary inhibitors of calpain due to slow substrate-like cleavage of a single peptide bond corresponding to Gly9-Ala10 in chicken cystatin. Biomolecular interaction analysis revealed that, unlike calpastatin, the cystatin-type inhibitors do not bind to the calmodulin-like domain of the small subunit of calpain, and their interaction with the mu-calpain heterodimer is completely prevented by a synthetic peptide comprising subdomain B of calpastatin domain 1. Based on these results we propose that (i) cystatin-type calpain inhibitors interact with the active site of the catalytic domain of calpain in a similar cystatin-like mode as with papain and (ii) the potential for calpain inhibition is due to specific subsites within the papain-binding regions of the general cystatin fold. PMID- 11258681 TI - School readiness for children with food allergies. PMID- 11258682 TI - Asthma severity and prevalence in adolescents--is there a change afoot? PMID- 11258683 TI - Problems encountered in improving care for inner-city children with asthma. PMID- 11258684 TI - Interpreting atmospheric pollen counts for use in clinical allergy: allergic symptomology. AB - BACKGROUND: Allergists generally consider atmospheric pollen counts to be an estimate of the antigenic challenge confronting allergic individuals. The nature of this challenge depends on the particular pollen types found in the atmosphere and also the airborne concentration of these pollen types. Both clinical experience and clinical investigations support these assumptions; however, a coherent system for relating pollen counts and allergic symptomology does not exist. OBJECTIVE: This review article will systematically review the medical and technical literature concerning the clinical significance of atmospheric pollen counts. DATA SOURCES: This review article will consider three independent bodies of literature: 1) data contrasting human exposure patterns with rooftop pollen counts; 2) data concerning dose-response relationships between atmospheric pollen counts and allergic symptomology; and 3) data concerning methods for indexing atmospheric pollen counts based on a pollen type's in vivo allergenicity and terminal velocity. RESULTS: Three principal results emerged. First, rooftop pollen counts imperfectly approximate human exposure to atmospheric pollen. Differences in both the concentration and type of pollen encountered by humans can be expected to differ from samples obtained on rooftops. Second, allergic symptomology is positively correlated with atmospheric pollen counts. Investigations involving Betula (birch) pollen offer quantitative dose-response models. Complex, nonlinear relationships that seem to reflect both the priming effect and late-phase reactions exist. Last, atmospheric pollen counts can be indexed based on a contemporary application of Thommen's postulates. This system provides allergists with a means to estimate the clinical significance of various pollen types by combining data concerning in vivo allergenicity and terminal velocity. CONCLUSIONS: These conclusions should allow allergists to judge the clinical significance of atmospheric pollen counts with greater sophistication than was previously possible. PMID- 11258685 TI - Hourly variation of airborne ragweed pollen in Kansas City. AB - INTRODUCTION: Ragweed pollen is one of the major causes of allergic rhinitis in the midwest United States. Previous studies have demonstrated that ragweed pollen production begins after sunrise and airborne pollen levels peak several hours later. Variations in this pattern that may be of clinical importance within a small region and the effect of weather on these variations have not been investigated. METHODS: Volumetric pollen collectors were stationed at four sites around the metropolitan area. Ten-minute grab samples were taken at each location every 2 hours for continuous 24-hour periods during the 1997 ragweed season. The downtown location had a weather station that logged meteorological conditions at hourly intervals during this time. Ragweed pollen grains were counted microscopically at 400 x. Uninterrupted data covering at least 20 days during the season were evaluated for each collection site. RESULTS: The lowest ragweed pollen counts occur at 6:00 AM and the highest occur at noon for both suburban and urban sites. Rainfall was seen to be the most influential weather-related factor. Significant rainfall events effectively removed pollen grains from the air. CONCLUSIONS: Though ragweed pollen emission begins at 6:00 AM, peak pollen exposure occurs at midday. This observation is in agreement with many other studies. The myth that highest ragweed exposure occurs in the early morning hours is not supported by this or previous studies. PMID- 11258686 TI - School readiness for children with food allergies. AB - BACKGROUND: Food allergy reactions and anaphylaxis may occur in children while at school. However, information regarding school readiness for children with food allergies is unknown. OBJECTIVE: To identify school education and prevention and treatment policies for food-allergic children in Michigan. METHODS: A questionnaire assessing food allergy awareness and avoidance and treatment strategies was mailed to a randomized sample (n = 273) of 2,082 public elementary school principals. RESULTS: One hundred four responses representing 109 schools were collected. From school estimates of 66,598 children, there was a 1.7% self reported prevalence of food allergy. The most common allergens were milk and peanut, followed by tree nuts, shellfish, egg, and wheat. Affected children were identified primarily through office records, with few reporting individual emergency plans or designated classrooms, teachers, or lunch tables. Methods of food allergy education included parents of students and in-services. Avoidance strategies, food substitution, and "no-sharing" policies were common, whereas other measures such as food-label-teaching were uncommon. A minority of schools had epinephrine immediately accessible, either in the student's classroom, carried by the student, or passed by teachers. Principals, nurses, and teachers were most often trained to administer epinephrine. CONCLUSIONS: There appears to be a need for schools to formally educate staff on food allergy, provide information on prevention measures such as reading of food labels, establish immediate accessibility to emergency epinephrine, and train staff for appropriate epinephrine use. PMID- 11258687 TI - Prevalence of asthma and other allergic diseases in an adolescent population: association with gender and race. AB - OBJECTIVES: To estimate the prevalence and severity of asthma in an adolescent population by sex and race. METHODS: Cross-sectional, population-based survey of school children. SETTING: Midwestern city experiencing damage from the 1993 Mississippi River flood. PARTICIPANTS: 2,693 children attending grades 7 to 12. MEASUREMENTS: Questions from the International Study of Asthma and Allergies in Childhood (ISAAC). RESULTS: Two thousand six hundred ninety-three children were surveyed, for a response rate of 90%. In this population, 16.4% reported having ever been diagnosed with asthma; 25% reported wheezing in the last 12 months; 32% reported symptoms of rhinitis in the last 12 months; and 22% reported ever having hay fever. The prevalence rate for current asthma was 12.6%. Female students had significantly greater prevalence rates for current asthma (16.4% vs 9.0%, OR = 1.85); ever-diagnosed asthma (18.5% vs 14.3%, OR = 1.36); wheezing > or = 4 times in the last 12 months (12.0% vs 5.6%, OR = 1.95); current rhinitis (38.7% vs 25.4%, OR = 1.73); and hay fever (26.4% vs 18.4%, OR = 1.57). All associations with sex remained significant, except ever-diagnosed asthma, after controlling for other known risk factors in logistic regression. African-Americans had higher prevalence rates than other races with differences reaching statistical significance for ever-diagnosed asthma and current asthma; however, these relationships did not remain significant after controlling for other known risk factors in logistic regression. CONCLUSIONS: Our prevalence rates were similar to those reported by other studies that used the ISAAC questionnaire. Female students reported significantly more asthma, wheezing, rhinitis, and hay fever than male students. Female students also reported more severe symptoms and a greater number of emergency room and hospital admissions. PMID- 11258688 TI - A comprehensive inner-city asthma program reduces hospital and emergency room utilization. AB - OBJECTIVE: To evaluate the efficacy of a comprehensive asthma program on emergency department (ED) visits and hospital admission rates in an inner-city pediatric population. DESIGN: A12-month prospective randomized trial. METHODS: Three hundred asthma patients, ages 2 to 17 years, were recruited and randomized in an inner-city pediatric ED, to obtain asthma care in a specialty clinic or to continue receiving care by other health resources. The specialty clinic provided intensive medical and environmental control, education, close monitoring, and 24 hour availability. For the prospective study, monthly questionnaires were sent to the caregivers of these children to evaluate use of hospital facilities for asthma care. For the retrospective study, use of hospital resources by the study participants was analyzed using a hospital database. RESULTS: One hundred twenty nine patients (60 in the treatment group and 69 in the control group) were included in the final analysis. Asthma severity index was significantly higher for the patients in the treatment versus the control group (35% versus 16.2%, P = .05). Fewer patients in the treatment group visited the ED at least once during the first study year, 32 versus 46, (P = .11), and they made fewer visits, 73 versus 269. The mean number of ED visits of the patients who used the ED was 0.1 versus 0.326 for the control group (P = .01). There were also fewer admissions in the treatment group, 22 versus 29 (P < .59). The 53 patients remaining in the treatment group in the second study year made fewer visits to the ED versus the control group (P < .03). In comparison to the first year, fewer patients in the treatment group visited the ED or were hospitalized in the second year (P = .007 and P = .04, respectively). CONCLUSIONS: A comprehensive asthma care program is efficacious in reducing hospital utilization. PMID- 11258689 TI - A community-based study of near-fatal asthma. AB - BACKGROUND: The purpose of this study was to describe the community-based impact of near-fatal asthma within the District of Columbia (Washington, DC). METHODS: The design was a prospective cohort study. Subjects included all persons in 1993 who presented to Washington, DC hospitals alive, requiring intubation for respiratory failure (including subjects who subsequently died in the hospital). Washington, DC hospitals were contacted on a biweekly basis to identify subjects. Patients were contacted by mail, followed by an interview with the subject or proxy. RESULTS: Of the 35 case subjects identified, 31 (88.6%) were interviewed. Sixty-one percent of the subjects were female; 84% were African-American; and 45.2% were less than 18 years old. Forty-five percent had asthma for 10 or more years. Twenty-three percent reported the emergency department as their usual source of health care, and 32% saw a provider on a weekly basis. Fifty-two percent were taking four or more prescription medications, and 29% were taking no anti-inflammatory medications. In the 24 hours before the event, 77% reported difficulty breathing, but only 64% reported contacting a health care provider. CONCLUSIONS: Community-based investigation of near-fatal asthma may lead to a better characterization of risk factors associated with this event. Findings from this study suggest that some of the factors associated with near-fatal events may be different from those associated with fatal asthma and that up to one third of the events may have been preventable. PMID- 11258690 TI - Efficacy of a single diagnostic test for sensitization to common inhalant allergens. AB - BACKGROUND: UniCAP Phadiatop is a single laboratory test designed to determine the presence or absence of specific IgE to a variety of common inhalants. Its purpose is to aid in the differentiation of patients with symptoms attributable to allergic disease from other common causes. METHODS: Consecutive children and adolescent patients (n = 145) at two centers were examined by having their history and physical examination performed by two board certified allergists. Their conclusions along with skin prick tests and specific IgE measurements regarding seven common inhalants (mite, oak, ragweed, grass, dog, cat, Alternaria) were compared with UniCAP Phadiatop test results. This was done using concordance of all test results. Attempts to resolve test discrepancies, when found, included specific RAST inhibitions, total IgE values, and physicians' judgment after testing. RESULTS: All patients with resolved diagnoses (143 of 145, 103 positive and 40 negative) were identified correctly by the UniCAP Phadiatop test. Skin test results and specific IgE measurements correlated well, but neither correlated well with the history by itself, suggesting a minimal false-positive component of the history of 23%. UniCAP Phadiatop results demonstrated a quantitative relationship between the patient's score and the amount of IgE specific to these individual allergens. CONCLUSIONS: The UniCAP Phadiatop test was shown to be highly sensitive and specific in differentiating individuals who are sensitized to common inhalants from those who are not. This test is recommended to all physicians as an aid in diagnostic and referral decisions for patients suspected of having an inhalant allergic diathesis. PMID- 11258691 TI - Mometasone furoate: efficacy and safety in moderate asthma compared with beclomethasone dipropionate. AB - BACKGROUND: Mometasone furoate (MF) is a new inhaled glucocorticoid administered by dry powder inhaler (DPI). OBJECTIVE: MF-DPI was evaluated for safety and efficacy and compared with placebo DPI and beclomethasone dipropionate (BDP) administered by metered dose inhaler (MDI) in the treatment of patients with moderate persistent asthma. METHODS: Eligible patients (n = 227), 13 to 75 years of age, maintained on inhaled glucocorticoids before entering the trial, were randomized to receive: MF-DPI, 100 microg, twice daily, MF-DPI, 200 microg, twice daily, BDP MDI, 168 microg, twice daily, or placebo in a 12-week, multicenter, double-blind study. RESULTS: At endpoint, FEV1 (primary efficacy variable) significantly improved for all three active treatments compared with placebo (P < .01, all comparisons). The response to MF-DPI, 200 microg, twice daily treatment was approximately twice as large as the response to MF-DPI, 100 microg, twice daily or BDP MDI treatment, although the differences between these groups did not reach statistical significance. Secondary efficacy variables including PEFR, asthma symptoms, nocturnal awakenings, and albuterol use showed similar trends. The MF-DPI, 100 microg, twice daily and BDP MDI, 168 microg, twice daily treatment groups produced comparable results for all efficacy variables. CONCLUSIONS: MF-DPI, 100 microg and 200 microg, twice daily were well-tolerated and significantly improved lung function and symptom control in the treatment of patients with moderate persistent asthma. In this study, MF-DPI, 200 microg, twice daily seemed to be the most effective dosage. PMID- 11258692 TI - Assessing the quality of asthma care provided to Medicaid patients enrolled in managed care organizations in Connecticut. AB - BACKGROUND: Many states have enrolled Medicaid beneficiaries in managed care organizations (MCOs). Few assessments of the quality of asthma care provided by these new programs are available. OBJECTIVE: To describe the quality of care provided to asthmatic Medicaid children enrolled in MCOs. METHODS: For this cross sectional survey, a chart abstraction tool was developed to evaluate fulfillment of key performance measures chosen from a national guideline for asthma diagnosis and management. These measures were prescription of an inhaled anti-inflammatory medication, accomplishment of patient education, evaluation of exposure to environmental triggers of asthma, and administration of influenza vaccination. From State of Connecticut administrative databases, a random sampling of Medicaid children, ages 5 to 18 years, enrolled in four MCOs was selected. Chart entries from July 1, 1996 to June 30, 1997 were reviewed using the abstraction tool. Accomplishment of performance measures was evaluated for the total sample and for children who were high utilizers of medical services (at least one ED visit or hospitalization during the study period). RESULTS: For 80 high utilizers among 315 children, completion of performance measures was suboptimal: 46% were prescribed inhaled steroids; an action plan was outlined for 43%; evaluation of patient or family tobacco use was documented for 56%; evaluation of the presence of a pet for 43% or mite exposure for 19%; and allergy skin testing or RAST was accomplished for 15%. CONCLUSIONS: This information suggests that opportunities exist to improve the quality of care for these children. PMID- 11258693 TI - Patients receiving immunotherapy report it is effective as assessed by the rhinitis outcomes questionnaire (ROQ) in a private practice setting. AB - BACKGROUND: Outcomes measurement is a proficient method in determining the effectiveness of medical therapy. Currently, there are no easy-to-use and inexpensive questionnaires available to evaluate the effectiveness of immunotherapy by allergists in private practice. OBJECTIVE: To evaluate the effectiveness of immunotherapy in the treatment of patients with allergic rhinitis in a private practice setting using the rhinitis outcomes questionnaire (ROQ). METHODS: One hundred seventy-five patients were randomly chosen from three private practices nationwide. They were surveyed regarding global systemic problems and nasal, eye, and chest symptoms, as well as their medical treatment history. This questionnaire was administered twice in one sitting, with the first a recall of symptoms before immunotherapy treatment, and the second an evaluation of current symptoms. RESULTS: The data revealed that 81% of the patients believed immunotherapy worked, with 19% unsure. Patients experienced a 67% decrease in antibiotic use, a 68% decrease in emergency room visits, a 75% decrease in days lost from work or school, and a 79% decrease in hospital admissions. The average symptom score reduction with immunotherapy was 52%. CONCLUSIONS: This study demonstrates that the user-friendly ROQ can be effectively and inexpensively used in a private practice setting and that immunotherapy is effective in the treatment of allergic rhinitis. PMID- 11258694 TI - Validation of the rhinitis outcomes questionnaire (ROQ). AB - BACKGROUND: Currently, there is no easy-to-use and comprehensive questionnaire that measures the effectiveness of treatment of rhinitis-related symptoms in a private practice setting. OBJECTIVE: To validate a brief, effective, self administered rhinitis symptoms questionnaire that can easily be used in a private practice setting for tracking treatment outcomes. METHODS: One hundred seventy five patients were randomly chosen from three private practices nationwide and were surveyed regarding systemic problems, nasal, eye, and chest symptoms, as well as medical treatment obtained. This survey was administered twice in one sitting with the first administration a recall of symptoms before immunotherapy, and the second administration an evaluation of current symptoms. Appropriate statistical methods were used to evaluate the validity, reliability, and responsiveness of the questionnaire presented. The validation of this instrument included content, statistical, construct, and predictive validity. RESULTS: Both the content and statistical validity were very good in that the questionnaire was easily understood and covered required domains. Construct validity revealed that the items were sensibly related to the domains that contained them and had less association with items from different domains. The reliability of the scales was very good (Cronbach's alpha = 0.80) to outstanding (Cronbach's alpha = 0.92). CONCLUSIONS: Functionality was established for the rhinitis outcomes questionnaire (ROQ) for use in a private practice setting. This allows private practice allergists to collect reliable data with the ROQ. PMID- 11258695 TI - Cytologic distinctions between clinical groups using curette-probe compared to cytology brush. AB - BACKGROUND: We had previously used curette-probe (Rhinoprobe; Arlington Scientific, Springville, UT) to study nasal cytology in various types of patients. Because of the potential sampling ease of using a brush, we sought to compare cytological results obtained with a curette-probe with those obtained using a cytology brush (Cytobrush Plus; Medscand, Malmo, Sweden). OBJECTIVE: To compare the ability of samples of nasal leukocytes obtained with a curette-probe versus a cytology brush to distinguish clinical categories of patients attending an allergy clinic. METHODS: Adult allergy clinic patients were studied by both curette-probe and cytology brush sampling. Quantitation of eosinophils and total leukocytes was performed on samples. Comparisons of cell quantities for each sampling method were made in patients classified into clinical groups. Patients with rhinitis complaints and abnormalities of nasal mucosal appearance with or without aeroallergy were compared with other patients. The adjustment of leukocyte quantities for the numbers of epithelial cells observed was also analyzed. Sampling methods were also compared for receiver operating characteristics. RESULTS: Curette-probe sample leukocyte quantities distinguished patients with symptoms of rhinitis (SR) with abnormal nasal appearance from other patients. This between-group distinction was more significant for leukocyte numbers normalized for the number of epithelial cells. SR patients with both abnormal nasal appearance and aeroallergy had significantly more eosinophils and less goblet cells than other patients. Greater than five curette-probe eosinophils were only observed in patients with SR. Brush samples did not show differences between patients stratified in these ways, and eosinophils were observed in patients without SR. Receiver operating characteristics favored curette-probe samples in terms of leukocyte or eosinophil increases characterizing their respective symptomatic patient subgroups. CONCLUSIONS: Curette-probe-obtained nasal samples allow for leukocyte and eosinophil quantitations which characterize rhinitis patients better than brush-obtained samples. Total leukocyte quantitations obtained by curette-probe may represent a marker of inflammatory nasal disease in adults undergoing allergy evaluation and treatment for rhinitis. PMID- 11258696 TI - Genetic polymorphisms as biomarkers of sensitivity to inhaled sulfur dioxide in subjects with asthma. AB - BACKGROUND: Individuals with asthma are sensitive to inhaled sulfur dioxide (SO2); decrements in pulmonary function occur after exposure to low concentrations even for a short duration of time. There is a great amount of interindividual variation in response to SO2. OBJECTIVE: It was our objective to determine whether one of the following polymorphism locations linked with asthma is associated with the bronchial hyperresponsiveness to SO2 observed in some asthmatic patients: the beta2-adrenergic receptor, interleukin-4 (IL-4) receptor alpha subunit, Clara cell secretory protein (CC16), TNF-alpha gene promoter, and first intron of the lymphotoxin alpha (LT-alpha) gene. METHODS: Subjects were volunteers with physician-diagnosed asthma requiring regular asthma medication. Spirometry was performed before and after a 10-minute exposure to 0.5 ppm SO2. Subjects were classified as SO2 responders if forced expiratory volume in 1 second (FEV1) decreased > or = 12%. DNA obtained from buccal cell samples was analyzed for genetic polymorphisms. RESULTS: Of the 62 subjects (21 male and 41 female), 13 had a 12% or greater decrement in FEV1 after SO2 exposure (range + 19% to -49%). Response to SO2 was associated with the wild-type allele of the TNF alpha promoter polymorphism (12 of 12 SO2 responders versus 28 of 46 nonresponders; P < .05) but with no other polymorphisms. Medication category and atopic status showed no association with SO2 sensitivity. CONCLUSIONS: The wild type allele of the TNF-alpha promoter polymorphism may be associated with mechanisms of asthmatic sensitivity to inhaled SO2. PMID- 11258698 TI - Latex/allergy. PMID- 11258697 TI - Bee pollen-induced anaphylactic reaction in an unknowingly sensitized subject. AB - BACKGROUND: The food supplement bee pollen has been previously found to cause anaphylactic reactions. It has been proposed as useful for "everything from bronchitis to hemorrhoids." OBJECTIVE: This study describes an atopic patient who experienced a non-life-threatening anaphylactic reaction upon her initial ingestion of bee pollen. Microscopic examination of the pollen sample and ELISA inhibition assays were performed. RESULTS: The patient had a 7 mm/28 mm wheal/erythema reaction to bee pollen at 1 mg/mL concentration. Bee pollen caused 52% inhibition of IgE binding to short ragweed and 55% to ryegrass. Microscopic analysis revealed ragweed, Alternaria, Cladosporium, honeysuckle (Lonicera sp), privet shrub (Ligustrum sp), and vetch (Vicia sativa). CONCLUSIONS: An unknowingly sensitized atopic patient experienced an anaphylactic reaction after ingestion of a small quantity of bee pollen that contained pollens and fungi. Previously administered allergen immunotherapy that had reduced rhinitis symptoms did not prevent this allergic reaction. PMID- 11258699 TI - American elm (Ulmus americana) is a native tree that has had a wide range from the entire eastern states through the Central Plains. PMID- 11258700 TI - Molecular and cytological analysis of a 5.5 Mb minichromosome. AB - Mammalian artificial chromosomes (MACs) provide a new tool for the improvement of our knowledge of chromosome structure and function. Moreover, they constitute an alternative and potentially powerful tool for gene delivery both in cultured cells and for the production of transgenic animals. In the present work we describe the molecular structure of MC1, a human minichromosome derived from chromosome 1. By means of restriction and hybridization analysis, satellite-PCR, in situ hybridization on highly extended chromatin fibres, and indirect immunofluorescence, we have established that: (i) MC1 has a size of 5.5 Mb; (ii) it consists of 1.1 Mb alphoid, 3.5 Mb Sat2 DNA, and telomeric and subtelomeric sequences at both ends; (iii) it contains an unusual region of interspersed Sat2 and alphoid DNAs at the junction of the alphoid and the Sat2 blocks; and (iv) the two alphoid blocks and the Sat2-alphoid region bind centromeric proteins suggesting that they participate in the formation of a functional kinetochore. PMID- 11258701 TI - Chromosomal DNA demethylation specified by protein binding. AB - In the present study, we utilize the well-characterized Escherichia coli lac repressor/operator system to demonstrate that protein binding can lead to demethylation at the binding sites in the chromosome. Similar to the findings using the episome, we found that the presence of LacI in the cells can lead to demethylation of methylated lacO in the chromosome and the LacI inhibitor, isopropyl-beta-D-thiogalactopyranoside (IPTG), can prevent demethylation of the methylated lacO. The lacO sites become progressively more demethylated over time with the presence of LacI, supporting the role of protein occupancy in demethylation targeting. These results validate our earlier conclusions using a stable episomal system, and establish for the first time that protein binding can specify sites of demethylation in the chromosome. PMID- 11258702 TI - The histone H4 acetyltransferase MOF uses a C2HC zinc finger for substrate recognition. AB - Site-specific acetylation of histone H4 by MOF is central to establishing the hyperactive male X chromosome in Drosophila. MOF belongs to the MYST family of histone acetyltransferases (HATs) characterized by an unusual C2HC-type zinc finger close to their HAT domains. The function of these rare zinc fingers is unknown. We found that this domain is essential for HAT activity, in addition to the established catalytic domain. MOF uses its zinc finger to contact the globular part of the nucleosome as well as the histone H4 N-terminal tail substrate. Point mutations that leave the zinc-finger structure intact nevertheless abolish its interaction with the nucleosome. Our data document a novel role of the C2HC-type finger in nucleosome binding and HAT activity. PMID- 11258703 TI - MCM3AP, a novel acetyltransferase that acetylates replication protein MCM3. AB - The MCM proteins are essential for the initiation of DNA replication. We have isolated an MCM3-associated protein (MCM3AP) in a two-hybrid screen using MCM3. Here we demonstrate that MCM3AP is an acetyltransferase which acetylates MCM3 and that chromatin-bound MCM3 is acetylated in vivo. The MCM3 acetylase, MCM3AP, is also chromatin-bound. This study also indicates that MCM3AP contains putative acetyl CoA binding motifs conserved within the GCN5-related N-acetyltransferase superfamily. Mutation of those motifs significantly inhibits the MCM3 acetylase activity. Over-expression of MCM3AP inhibits DNA replication, whereas mutation of the acetylase motifs abolishes this effect, suggesting that acetylation plays a role in DNA replication. Taken together, we suggest that MCM3 acetylation is a novel pathway which might regulate DNA replication. PMID- 11258704 TI - An activation-independent role of transcription factors in insulator function. AB - Chromatin insulators are defined as transcriptionally neutral elements that prevent negative or positive influence from extending across chromatin to a promoter. Here we show that yeast subtelomeric anti-silencing regions behave as boundaries to telomere-driven silencing and also allow discontinuous propagation of silent chromatin. These two facets of insulator activity, boundary and silencing discontinuity, can be recapitulated by tethering various transcription activation domains to tandem sites on DNA. Importantly, we show that these insulator activities do not involve direct transcriptional activation of the reporter promoter. These findings predict that certain promoters behave as insulators and partition genomes in functionally independent domains. PMID- 11258705 TI - The archaeal TFIIEalpha homologue facilitates transcription initiation by enhancing TATA-box recognition. AB - Transcription from many archaeal promoters can be reconstituted in vitro using recombinant TATA-box binding protein (TBP) and transcription factor B (TFB)- homologues of eukaryal TBP and TFIIB--together with purified RNA polymerase (RNAP). However, all archaeal genomes sequenced to date reveal the presence of TFE, a homologue of the alpha-subunit of the eukaryal general transcription factor, TFIIE. We show that, while TFE is not absolutely required for transcription in the reconstituted in vitro system, it nonetheless plays a stimulatory role on some promoters and under certain conditions. Mutagenesis of the TATA box or reduction of TBP concentration in transcription reactions sensitizes a promoter to TFE addition. Conversely, saturating reactions with TBP de-sensitizes promoters to TFE. These results suggest that TFE facilitates or stabilizes interactions between TBP and the TATA box. PMID- 11258706 TI - Inhibition of p53-dependent transcription by BOX-I phospho-peptide mimetics that bind to p300. AB - The N-terminal BOX-I domain of p53 containing a docking site for the negative regulator MDM2 and the positive effector p300, harbours two recently identified phosphorylation sites at Thr18 or Ser20O whose affect on p300 is undefined. Biochemical assays demonstrate that although MDM2 binding is inhibited by these phosphorylations, p300 binding is strikingly stabilized by Thr18 or Ser20 phosphorylation. Introducing EGFP-BOX-I domain peptides with an aspartate substitution at Thr18 or Ser20 induced a significant inhibition of endogenous p53 dependent transcription in cycling cells, in irradiated cells, as well as in cells transiently co-transfected with p300 and p53. In contrast an EGFP-wild-type BOX-I domain peptide stimulated p53 activity via inhibition of MDM2 protein binding. These results suggest that phosphorylation of p53 at Thr18 or Ser20 can activate p53 by stabilizing the p300-p53 complex and also identify a class of small molecular weight ligands capable of selective discrimination between MDM2- and p300-dependent activities. PMID- 11258707 TI - p19ARF-independent induction of p53 and cell cycle arrest by Raf in murine keratinocytes. AB - In tumorigenesis of the skin, activated Ras co-operates with mutations that inactivate the tumour suppressor p53, but the molecular basis for this co operation remains unresolved. Here we show that activation of the Raf/MAP kinase pathway in primary mouse keratinocytes leads to a p53 and p21Cip1-dependent cycle arrest and to terminal differentiation. Raf activation in keratinocytes lacking p53 or p21Cip1 genes leads to expression of differentiation markers, but the cells do not cease to proliferate. Thus, loss of p53 or p21Cip1 function is necessary to disable growth-inhibitory Raf/MAP kinase signalling. Activation of oncogenes, including Ras, has been reported to stabilize and activate p53 via induction of the tumour suppressor p19ARF. However, the response to Raf in p19ARFI-/- keratinocytes was indistinguishable from wild-type controls. Thus, p19ARF is not essential for Raf-induced p53 induction and cell cycle arrest in keratinocytes, indicating that oncogenes engage p53 activity via multiple mechanisms. PMID- 11258708 TI - A role for chemokine receptor transactivation in growth factor signaling. AB - Complex cell responses require the integration of signals delivered through different pathways. We show that insulin-like growth factor (IGF)-I induces specific transactivation of the Gi-coupled chemokine receptor CCR5, triggering its tyrosine phosphorylation and Galpha recruitment. This transactivation occurs via a mechanism involving transcriptional upregulation and secretion of RANTES, the natural CCR5 ligand. CCR5 transactivation is an essential downstream signal in IGF-I-induced cell chemotaxis, as abrogation of CCR5 function with a transdominant-negative KDELccr5A32 mutant abolishes IGF-I-induced migration. The relevance of this transactivation pathway was shown in vivo, as KDELccr5A32 overexpression prevents invasion by highly metastatic tumor cells; conversely, RANTES overexpression confers built-in invasive capacity on a non-invasive tumor cell line. Our results suggest that this extracellular growth factor-chemokine network represents a general mechanism connecting tumorigenesis and inflammation. PMID- 11258710 TI - Money in scientists' pockets. PMID- 11258709 TI - EGF receptor/Rolled MAP kinase signalling protects cells against activated Armadillo in the Drosophila eye. AB - beta-catenin/Armadillo are transcriptional co-activators that mediate Wnt signalling in normal development. Activated forms of beta-catenin are oncogenic. We have constructed mutant forms of Drosophila Armadillo which correspond to common human oncogenic mutations, and find them to activate Armadillo constitutively. When expressed in the Drosophila eye, these eventually induce apoptosis in all cell types. Intriguingly, cells in the eye are resistant to the effects of activated Armadillo for a long period prior to the onset of cell death at the mid-pupal stage. This latency is conferred by EGF receptor (EGFR)/MAP kinase signalling, which prevents activated Armadillo from inducing apoptosis; when EGFR signalling naturally ceases, the cells rapidly die. Nemo, the Drosophila homologue of NLK in mice and LIT-1 in Caenorhabditis elegans, does not antagonize activated Armadillo, suggesting that the Nemo-like MAP kinases may not generally interact with Armadillo/beta-catenin. Thus, our results show that activated Armadillo is subject to a specific negative control by EGFR/Rolled MAP kinase signalling. PMID- 11258711 TI - Comment on the editorial 'The secret ways of scientists' in EMBO reports, December 2000. PMID- 11258712 TI - Comment on the editorial 'Back to Darwin?' in EMBO reports, November 2000. PMID- 11258713 TI - An old enemy, a new battle plan. Perspective on combating drug-resistant malaria. PMID- 11258715 TI - The Nobel Prizes in the new century. An interview with Ralf Pettersson, Director of the Stockholm Branch of the Ludwig Institute for Cancer Research, the Karolinska Institute, and former chairman of the Nobel Prize Committee for Physiology/Medicine. Interview by Holger Breithaupt. PMID- 11258714 TI - Clinical practice in the new era. A fusion of molecular biology and classical medicine is transforming the way we look at and treat diseases. PMID- 11258716 TI - Brave small world. Biotechnology and nanotechnology may give rise to a completely new industry. PMID- 11258717 TI - What is patent-worthy? Due to differing legal standards for patentability, applications for gene patents face different outcomes in various countries. PMID- 11258718 TI - The 'double lives' of membrane lipids. Workshop: Anno 2000. A lipid milestone. PMID- 11258720 TI - Applications to veterinary school--a touch of class? PMID- 11258719 TI - JAB1/CSN5 and the COP9 signalosome. A complex situation. AB - The Jun activating binding protein (JAB1) specifically stabilizes complexes of c Jun or JunD with AP-1 sites, increasing the specificity of target gene activation by AP-1 proteins. JAB1 is also known as COP9 signalosome subunit 5 (CSN5), which is a component of the COP9 signalosome regulatory complex (CSN). Over the past year, JAB1/CSN5 has been implicated in numerous signaling pathways including those that regulate light signaling in plants, larval development in Drosophila, and integrin signaling, cell cycle control, and steroid hormone signaling in a number of systems. However, the general role of the CSN complex, and the specific role of JAB1/CSN5, still remain obscure. This review attempts to integrate the available data to help explain the role of JAB1/CSN5 and the COP9 signalosome in regulating multiple pathways (in this review, both JAB1 and CSN5 terminologies are used interchangeably, depending on the source material). PMID- 11258721 TI - Clinical signs, histopathology and genetics of experimental transmission of BSE and natural scrapie to sheep and goats. AB - This paper compares the dinical signs, histopathology, detection of PrPSc protein and PrP genetics of the transmission of BSE to sheep and goats, with the effects of the transmission of natural scrapie from a brain homogenate from a single sheep. After intracerebral and oral inoculations there were similarities in the clinical signs due to the two sources of infection, but there were differences in pathology at the end stage of disease and in the genotypes of the sheep which succumbed to the challenges. The incubation period of BSE was associated with the sheep PrP codon 171 genotype, but the natural scrapie source, despite inducing disease only in known susceptible genotypes, showed no clear association with PrP genotype. PMID- 11258722 TI - Acute phase protein responses to uterine bacterial contamination in cattle after calving. AB - Repeated ultrasonographic examinations and collections of blood samples and uterine lumenal swabs between seven and 28 days after calving were used to examine the relative effects of bacterial contamination and involution of the uterus on the concentrations of acute phase proteins in the blood of 26 dairy cows. The severity of bacterial contamination, as determined by the total bacterial growth score, was a significant variable for the concentrations of the acute phase proteins alpha1-acid glycoprotein (P < 0.0001), haptoglobin (P < 0.05) and ceruloplasmin (P < 0.0001). In addition, the concentrations of alpha1 acid glycoprotein and ceruloplasmin were increased in the cows from which Escherichia coli (P < 0.0001) and Arcanobacterium pyogenes (P < 0.05), respectively, were isolated from the uterine lumen. Uterine involution, as determined by the decreasing diameter of the previously gravid uterine horn, was associated with a decrease in the concentrations of alpha1-acid glycoprotein (P < 0.005), haptoglobin (P < 0.05) and ceruloplasmin (P < 0.01). However, the response of the acute phase proteins to bacterial contamination was independent of the day on which the samples were collected, indicating that their concentrations were increased by bacterial contamination in addition to the changes associated with uterine involution. PMID- 11258723 TI - Scapulohumeral osteoarthritis in 20 Shetland ponies, miniature horses and falabella ponies. AB - This paper describes the clinical and diagnostic features of 20 cases of scapulohumeral osteoarthritis in Shetland ponies, miniature horses and falabella ponies. The history and clinical signs were similar in all the cases Radiographically they all had consistent changes which consisted predominantly of articular osteophytes and periarticular enthesiophytes. Six of the cases had radiographic evidence of dysplasia of the scapulohumeral joint, although it was uncertain whether this was a primary or a secondary finding. No specific treatment appeared to be advantageous. At follow up, six of the ponies had to be euthanased owing to continuing severe lameness; the other 14 ponies remained lame, but were maintained at pasture by the occasional use of oral non-steroidal anti-inflammatory drugs. No definitive aetiology for the condition was identified, but it is proposed that an underlying dysplasia, or lack of collateral support may predispose the scapulohumeral joint of miniature horse breeds to the disease. PMID- 11258724 TI - Thoracic sympathetic chain ganglion neuronal abnormalities that may explain some of the clinical signs of grass sickness. PMID- 11258725 TI - Suilysin production by Streptococcus suis strains isolated from diseased and healthy carrier pigs in Spain. PMID- 11258726 TI - Arcanobacterium/Corynebacterium-like bacterial isolates from sheep. PMID- 11258727 TI - Privilege to dispense. PMID- 11258728 TI - Bulk tank milk failures. PMID- 11258729 TI - Closure of the Thurso veterinary investigation centre. PMID- 11258730 TI - Representation on RCVS Council. PMID- 11258731 TI - Proposals for new RCVS certificates. PMID- 11258732 TI - FLUTD/feline idiopathic cystitis study. PMID- 11258734 TI - Trimming and shoeing of horses' hooves. PMID- 11258733 TI - Coenurosis in a pet rabbit. PMID- 11258735 TI - Elderly companion animal healthcare. PMID- 11258736 TI - Neosporosis of cattle. PMID- 11258737 TI - Early Trombicula autumnalis infection. PMID- 11258738 TI - Efficacy and effectiveness of child and adolescent psychotherapy and pharmacotherapy. AB - Decades of intervention research have produced a rich body of evidence on the effects of psychotherapies and pharmacotherapies with children and adolescents. Here we summarize and critique that evidence. We review findings bearing on the efficacy of psychosocial treatments and medications under controlled experimental conditions. We also report evidence, where available, on the effectiveness of both classes of treatment with clinically referred youth treated in real-world clinical contexts. In general, the large body of evidence on efficacy contrasts sharply with the small base of evidence on effectiveness. Addressing this gap through an enriched research agenda could contribute importantly to linking scientific inquiry and clinical practice-to the benefit of both ventures. This is one element of a multifaceted agenda for future research and for synthesis of research, which will require the interplay of multiple disciplines related to child and adolescent mental health. PMID- 11258739 TI - A comparison of public and privatized approaches to managed behavioral health care for persons with serious mental illness. AB - This article compares public and privatized approaches to managed behavioral health care for persons with serious mental illness in Massachusetts. Data from the Department of Mental Health (DMH) for 247 patients receiving care managed by DMH and 312 in a Medicaid carve-out were compared. Repeated measures multivariate analysis of variance models were used to examine adjusted changes in number of admissions, bed days, and facilities used from a baseline year before program implementation in 1992 through two follow-up years. Results were comparable for the two programs with similar reductions in the number of people receiving inpatient care but increases in admissions and bed days. Possible problems with continuity of care, indicated by individuals using multiple facilities, were identified for both. Given the evidence of comparable results, the choice between the two approaches is likely to be dictated by various pragmatic and subjective factors other than their demonstrated effectiveness. PMID- 11258740 TI - Multisystemic treatment of substance-abusing and dependent delinquents: outcomes, treatment fidelity, and transportability. AB - The effectiveness and transportability of multisystemic therapy (MST) were examined in a study that included 118 juvenile offenders meeting DSM-III-R criteria for substance abuse or dependence and their families. Participants were randomly assigned to receive MST versus usual community services. Outcome measures assessed drug use, criminal activity, and days in out-of-home placement at posttreatment (T2) and at a 6-month posttreatment follow-up (T3); also treatment adherence was examined from multiple perspectives (i.e., caregiver, youth, and therapist). MST reduced alcohol, marijuana, and other drug use at T2 and total days in out-of-home placement by 50% at T3. Reductions in criminal activity, however, were not as large as have been obtained previously for MST. Examination of treatment adherence measures suggests that the modest results of MST were due, at least in part, to difficulty in transporting this complex treatment model from the direct control of its developers. Increased emphasis on quality assurance mechanisms to enhance treatment fidelity may help overcome barriers to transportability. PMID- 11258741 TI - Cost-shifting in manged care. AB - We conducted a study of the change from fee-for-service to managed care for mental health services in the Massachusetts Medicaid program, which occurred in fiscal year 1993. We estimated the effect of managed care on total public expenditures over both the short and the long term. Per person expenditures were lower by 24% in the first year of managed care but only lower by 5% in the second and third years. We also tested for cost-shifting by estimating expenditures for five specific services paid by three public agencies. Expenditures on services paid by the managed care vendor decreased, expenditures paid by Medicaid increased, and expenditures paid by the Department of Mental Health decreased. We discuss the implications for both cost-shifting and quality of care improvements. The results from two-part expenditure models indicate that some cost-shifting may be related to quality improvement. The effects are generally stronger for the beneficiaries in the highest quartile of expenditures. PMID- 11258742 TI - Particle electrophoresis for quality assurance and process control. AB - Process control is an increasingly important issue as life science companies world-wide strive for recognition of their manufacturing and product development quality measures according to International Standards Organization (ISO) or good manufacturing practices (GMP) standards. Analytical particle electrophoresis (APE) has the potential for significant contributions, not just to basic research, but also in process development and control in manufacturing environments. An important feature of colloidal (small) particles, which controls their behavior, is their surface charge. Optimization of life science products and process conditions involving small particles (>100 nm) may be approached by a variety of strategies based upon direct measurements of the charge properties of process particles or "reporter" particles. The availability of increasingly powerful instruments and control particle preparations (National Institute of Standards and Technology ((NIST) and others) for validation of instrument operation make the method more attractive than ever. We summarize highly flexible electrophoretic strategies for assessing process consistency both from the perspective of particles being processed as well as the processing environment and describe principles for the use of polymer microspheres both as control particles for validation of instrument operation as well as for probes of the assay medium. PMID- 11258743 TI - One-step separation of endocytic organelles, Golgi/trans-Golgi network and plasma membrane by density gradient electrophoresis. AB - Many different methods for the fractionation of subcellular organelles have been reported. However, no protocol for rapid separation of plasma membrane, Golgi/trans-Golgi network (TGN) and endosomes is available to date. Such a method is a prerequisite for a quantitative biochemical analysis of vesicular transport from the Golgi/TGN compartment to plasma membrane and endosomes. Here a density gradient electrophoresis protocol is described that allows the fractionation of these organelles in one step. This protocol requires only low-cost instrumentation available in most biochemical laboratories. PMID- 11258744 TI - Electroelution of proteins from bands in gel electrophoresis without gel sectioning for the purpose of protein transfer into mass spectrometry: elements of a new procedure. AB - Electroelution of protein bands from a gel has advantages over the competitive common technique requiring gel sectioning with respect to yield, speed and the potential for computer-controlled application to multicomponent two-dimensional (2-D) gels. The electroelution design for the commercial high-performance gel electrophoresis (HPGE) apparatus represented the most advanced technique to date until the recent discontinuation of its production. The present report serves to summarize the necessary design elements for the purpose of renewing and further developing the electroelution technique. A rudimentary technique is presented by which the electroeluate is collected in a glass tube superimposed on a reversibly stained gel band and connected to an anolyte reservoir. Although the stain used is insufficiently sensitive, the technique allowed for the qualitative verification of its usefulness in the transfer of the electroeluate into mass spectrometry. PMID- 11258745 TI - Green-light transilluminator for the detection without photodamage of proteins and DNA labeled with different fluorescent dyes. AB - The excitation spectra of Nile red and SYPRO red, two currently used dyes for the fluorescent staining of protein bands in sodium dodecyl sulfate (SDS) polyacrylamide gels, show an excitation peak in the UV region and another in the visible region (maximum at about 550 nm). Ethidium bromide and other intercalating dyes, e.g. propidium iodide, ethidium dimers, and benzoxazolium-4 quinolinium dimer-3 (YOYO), used for the fluorescent staining of DNA bands in agarose gels also show an excitation peak in the same region of the visible spectrum. We have designed and constructed a green-light transilluminator with an emission maximum at 542 nm. This visible transilluminator allows the detection of protein bands stained with Nile red and SYPRO red with the same sensitivity obtained with a 300 nm UV transilluminator. The green-light transilluminator also allows the detection of about 2 ng of DNA per band in gels stained with ethidium bromide and the other intercalating dyes indicated above. In contrast to the UV transilluminators, the green-light transilluminator does not produce photodamage of DNA even after long exposures (10 min). This makes this transilluminator very useful for preparative work. Furthermore, the green-light transilluminator does not require UV safety equipment and, consequently, it can be very convenient for teaching laboratories. PMID- 11258746 TI - The use of polyacrylamide gel electrophoresis in animal systematics, phylogeny, and ecophysiology. AB - In the 1980s, the use of polyacrylamide gel electrophoresis (PAGE) was popular. But more recent developments in other electrophoretic techniques have resulted in this method being less widely used. However, it is adequate for the needs of naturalists for issues which do not require high-performance methods, such as in systematics, phylogeny, studies of intraspecific or clinal variability, ecology and ecophysiology. We illustrate the application of PAGE by the results of a research program on marine and terrestrial invertebrates which was conducted from 1979 to 1992, but which is still used to initiate graduate students to the research. Thus, issues faced by the zoologist can be clarified by this method, not yet obsolete and still useful in natural history despite considerable advances in other electrophoretic methods. PMID- 11258747 TI - The electrophoretic mobility of DNA three-way junctions is affected by the sequence of overhanging single-stranded ends. AB - The folding of three- and four-way DNA junctions is often assessed by comparing the electrophoretic mobility of restriction enzyme fragments, using the long short arm assay. We have compared the mobility of synthetic three-way junctions that contain identical branch point sequences, but different restriction sites in the arms. We show that the mobility of fragments is affected by the sequence of the overhanging ends. In general, GC-rich overhangs produce fragments with anomalous mobilities. These anomalies can be prevented by treating the cleaved junctions with mung bean endonuclease, elevating the electrophoresis temperature or using blunt cleaving restriction endonucleases. PMID- 11258748 TI - A novel technique for detecting single nucleotide polymorphisms by analyzing consumed allele-specific primers. AB - We present a simple and rapid polymerase chain reaction (PCR)-based technique, termed consumed allele-specific primer analysis (CASPA), as a new strategy for single nucleotide polymorphism (SNP) analysis. The method involves the use of labeled allele-specific primers, differing in length, with several noncomplementary nucleotides added in the 5'-terminal region. After PCR amplification, the amounts of the remaining primers not incorporated into the PCR products are determined. Thus, nucleotide substitutions are identified by measuring the consumption of primers. In this study, the CASPA method was successfully applied to ABO genotyping. In the present method, the allele specific primer only anneals with the target polymorphic site on the DNA, so it is not necessary to analyze the PCR products. Therefore, this method is only little affected by modification of the PCR products. The CASPA method is expected to be a useful tool for typing of SNPs. PMID- 11258749 TI - Single-molecule immunoassay and DNA diagnosis. AB - Many assays relevant to disease diagnosis are based on electrophoresis, where the migration velocity is used for distinguishing molecules of different size or charge. However, standard gel electrophoresis is not only slow but also insensitive. We describe a single-molecule imaging procedure to measure the electrophoretic mobilities of up to 100000 distinct molecules every second. The results correlate well with capillary electrophoresis (CE) experiments and afford confident discrimination between normal (16.5 kbp) and abnormal (6.1 kbp) mitochondrial DNA fragments, or beta-phycoerythrin-labeled digoxigenin (BP-D) and its immunocomplex (anti-D-BP-D). This demonstrates that virtually all electrophoresis diagnostic protocols from slab gels to CE should be adaptable to single-molecule detection. This opens up the prossibility of screening single copies of DNA or proteins within single biological cells for disease markers without performing polymerase chain reaction (PCR) or other biological amplification. PMID- 11258750 TI - Detection of avocado sunblotch viroid variants using fluorescent single-strand conformation polymorphism analysis. AB - A specific reverse transcription-polymerase chain reaction (RT-PCR) protocol has been developed for routine detection of avocado sunblotch viroid (ASBVd). Modifications in this diagnostic technique were made to enable fluorescent detection and variant identification using automated capillary electrophoresis (CE) and fluorescent single-strand conformation polymorphism (SSCP) analysis. Sixteen sequence variants characterized in a previous study were analyzed using CE-SSCP on two ABI 310 Genetic Analyzers. Significant differences were detected between data obtained from the two ABI 310 Genetic Analyzers indicating that an internal control must be run concurrently with the samples. The 16 variants could be classified into 11 groups based on the SSCP patterns. The statistical analysis of the migration rate data provided support for the visual differences in SSCP patterns. The use of SSCP in the ASBVd assay is easily accomplished and gives an estimate of the number of variants in crude samples extracted from infected avocado plants. PMID- 11258751 TI - Genotyping Cryptosporidium parvum by single-strand conformation polymorphism analysis of ribosomal and heat shock gene regions. AB - Polymerase chain reaction (PCR)-coupled single-strand conformation polymorphism (SSCP) approaches utilizing nuclear DNA regions of the small subunit (SSU) of ribosomal RNA and heat shock protein 70 gene (HSP70) were established for genotyping Cryptosporidium parvum. The regions were amplified (individually or in a multiplex reaction) by PCR from DNA extracted from oocysts from ruminant or human hosts, then denatured and subjected to electrophoresis in a mutation detection enhancement (nondenaturing) gel matrix. Single-strand profiles produced in SSCP allowed the unequivocal identification/differentiation of the two common (human, 1 and cattle, 2) genotypes of C. parvum and the direct display of sequence variability within some samples, reflecting population variation. As these are considered among the most closely related genotypes (based on SSU and HSP70 sequence data), these findings and other preliminary results for C. felis (from cat) C. serpentis (from snake) and C. baileyi (from bird) indicate that the SSCP approaches established could be employed to identify any of the currently recognised genotypes and species of Cryptosporidium. PMID- 11258752 TI - The use of tat and env sequences from human immunodeficiency virus 1 in phylogenetic epidemiological studies. AB - Using nucleotide sequences from the first exon of the tat gene of the human immunodeficiency virus 1 (HIV-1), we tested the hypothesis that a Florida dentist (a common source) infected five of his patients in the course of dental procedures against the null hypothesis that the dentist and each individual of the dental group independently acquired the virus within the local community. This novel approach of analyzing the tat gene region was used because it may, in some circumstances, be more informative for phylogenetic epidemiology than the more commonly used C2-V3 envelope gene region. The first exon of the tat gene was polymerase chain reaction (PCR)-amplified and directly sequenced from uncultured peripheral blood mononuclear cells. Patient's sequences were compared with sequences from six HIV-1 infected heterosexual couples unrelated to the dentist or the five patients, but from the same general geographic area. In addition, a sixth infected dental patient, previously inferred to have acquired HIV-1 from a source other than the dentist, was included. Multiple phylogenetic analyses demonstrated that the sequences of the five patients were significantly more closely related to each other than to sequences of the controls. Our results using tat sequences, combined with envelope sequence data, strongly support a common phylogenetic epidemiological relationship among these five patients, and the HIV-1 infected dentist who treated them. Correct recovery of known epidemiological relationships among couples included in the analysis further strengthens this conclusion. PMID- 11258753 TI - Evidence for the relation of herpes simplex virus type 1 to Down syndrome and Alzheimer's disease. AB - The peripheral and central nervous system are harbouring herpes simplex virus type 1 (HSV-1) and this virus has been proposed to be implicated in the aetiology of Alzheimer's disease (AD). We tested whether the HSV-1 genome is found indeed in the brain of controls, patients with AD and Down syndrome (DS) and whether HSV 1 infectious proteins in brain were induced. Moreover, we tested whether interleukin (IL)-6, a marker for neuroinflammation, is found in brains of AD and DS. HSV-1 glycoprotein D gene, as well as viral phosphoprotein and glycoprotein were detected in all brain samples. IL-6 was detectable in seven out of the eight AD and all of the eight DS patients, but only three out of ten controls in the frontal cortex. IL-6 in cerebellum was detectable in all AD and DS patients, but only three out of nine controls. In conclusion, we propose that the detection of HSV-1 genome and HSV-1 inducible protein IL-6 not only shows the presence in human brain, but may indicate a role for HSV-1 in the process of neuroinflammation and apoptosis, known to occur in both neurodegenerative disorders, AD and DS. PMID- 11258754 TI - Spatial open-network formed by mixed triblock copolymers as a new medium for double-stranded DNA separation by capillary electrophoresis. AB - A mixture of two polyoxybutylene-polyoxyethylene-polyoxybutylene (BEB) triblock copolymers (B6E46B6 and B10E271B10, respectively) was used as a new separation medium for separating double-stranded DNA (dsDNA) fragments by capillary electrophoresis (CE). The two block copolymer mixtures were designed to form mixed flower-like micelles in dilute solution and a homogeneous gel-like open network with hydrophobic clusters as cross-linking points at higher polymer concentrations. Being a polyoxyalkylene block copolymer gel, the separation medium has some special advantages, including the temperature-dependent sol-gel transition that makes sample injection easy, and the self-coating of the inner capillary wall that makes experimental procedures simple and reproducible. Furthermore, it can shorten the elution time and further improve the separation resolution, especially for small dsDNA fragments, when compared with EPE-type separation media, e.g., F127 (E99P69E99, with P being polyoxypropylene) block copolymer gels formed by the closed packing of spherical micelles. Single base pair resolution can be achieved by using the new separation medium for dsDNA fragments up to over 100 base pairs. PMID- 11258755 TI - Effects of electrolyte modification and capillary coating on separation of glycoprotein isoforms by capillary electrophoresis. AB - The capillary electrophoresis (CE) running electrolyte composition was optimized for the separation of selected glycoproteins. A good separation of the ovalbumin (OV) and alpha-acid glycoprotein (AAG) isoforms was achieved in 20 mmol x L(-1) N (2-hydroxyethyl)piperazine-2'-(2-ethanesulfonic acid) (HEPES) at pH 7.0, in 20 mmol x L(-1) phosphate, pH 7.0, or in 25 mmol x L(-1) borate, pH 7.9. Various ways of suppression of the glycoprotein adsorption onto the capillary wall were compared. Alpha, omega-diamine alkanes and bis(aminoalkyl) amines were added to the CE buffers, the optimized concentration being 1 mmol x L(-1) in 20 mmol x L( 1) phosphate buffer. The OV and AAG isoforms could be separated using all the alpha,omega-diamine alkanes or bis(2-aminoethyl)amine. The length of the alkyl chain in the diaminoalkane did not influence the separation. The separation of the isoforms of pollen allergens was also tested. The effects of modification of the capillary wall by succinyl-poly-L-lysine and hydrophilic CElect-P1 capillary were compared. A decrease in the glycoprotein and protein adsorption resulted in an improved separation of the isoforms. PMID- 11258756 TI - Determination of pKa values of anthraquinone compounds by capillary electrophoresis. AB - In this paper, the dissociation constants (pKa1, pKa2) of five anthraquinones were determined from the relation between the effective mobility at different pH values and the buffer pH value, which was derived from the basic electrophoresis theory and the dissociation equilibrium of a binary acid. In addition, the changes of pKa values of the five compounds were also investigated when organic modifiers were added to the buffer system. PMID- 11258757 TI - Separation of multicomponent mixtures of 2,4-dinitrophenyl labelled amino acids and their enantiomers by capillary zone electrophoresis. AB - The use of capillary zone electrophoresis (CZE) for the separation of a group of 33 2,4-dinitrophenyl labeled amino acids (DNP-AA), including DNP-AA racemates, DNP-L-AA enantiomers and achiral DNP-AAs, was investigated. Alpha-, beta- and gamma-cyclodextrins (CDs) and their derivatives (hydroxypropyl derivatives of alpha-, beta- and gamma-CDs, polymeric beta-CD and 6A-methylamino-beta cyclodextrin (MA-beta-CD)) served as complexing agents and chiral selectors in this investigation. Although native alpha- and gamma-CDs and their derivatives influenced the effective mobilities of the studied DNP-AAs in different ways, they generally failed to resolve enantiomers of the individual DNP-AAs. On the other hand, beta-CD and all of its derivatives were found to be effective in this respect. Of these, the best results were achieved with a positively charged MA beta-CD and this chiral selector resolved enantiomers of ten DNP-AA racemates available for this study. However, a complete resolution of these enantiomers in one CZE run required that the effect of the chiral selector be complemented by complexing effects of polyvinyl pyrrolidone (PVP) or gamma-CD. Complexing and chiral recognition capabilities of MA-beta-CD combined with complexing effects of gamma-CD and PVP provided separating conditions suitable for the CZE separations of multicomponent mixtures of DNP-AAs with preserved resolutions of the enantiomers. For example, a mixture consisting of 43 DNP-AA constituents was resolved using an MA-beta-CD/gamma-CD combination with three peak overlaps. PMID- 11258758 TI - Capillary zone electrophoretic chiral discrimination using a cationic cyclodextrin derivative: determination of velocity and association constants of each enantiomer of the amino acid derivative with 6-trimethylammonio-deoxy-beta cyclodextrin. AB - A positively charged beta-cyclodextrin possessing a trimethylammonio group, 6 trimethylammonio-6-deoxy-beta-cyclodextrin chloride 1 was prepared by the reaction of an amino derivative 3 with methyl iodide under very mild conditions. The cyclodextrin derivative discriminated between enantiomers of acetylphenylalanine 2 on capillary zone electrophoresis (CZE) analysis, which was possibly due to an electrostatic interaction between the trimethylammonio group of 1 and the carboxylate group of 2, and also due to the inclusion of 2 in a hydrophobic molecular cavity of 1. Double data normalization based on relative electrophoretic velocity of peaks due to indirect response of 1 and also water in an injected sample may be effective for elimination of variation and fluctuation of physical parameters of medium, such as viscosity and ionic strength, in order to determine intrinsic association constants and velocity of the complexes. PMID- 11258759 TI - Determination of nonsteroidal anti-inflammatory drugs in biological fluids by automatic on-line integration of solid-phase extraction and capillary electrophoresis. AB - A new, automatic method for the clean-up, preconcentration, separation, and quantitation of nonsteroidal anti-inflammatory drugs (NSAIDs) in biological samples (human urine and serum) using solid-phase extraction coupled on-line to capillary electrophoresis is proposed. Automatic pretreatment is carried out by using a continuous flow system operating simultaneously with the capillary electrophoresis equipment, to which it is linked via a laboratory-made mechanical arm. This integrated system is controlled by an electronic interface governed via a program developed in GWBasic. Capillary electrophoresis is conducted by using a separation buffer consisting of 20 mM NaHPO4, 20 mM beta-cyclodextrin and 50 mM SDS at pH 9.0, an applied potential of 20 kV and a temperature of 20 degrees C. The analysis time is 10 min and the detection limits were between 0.88 and 1.71 microg mL(-1). Automatic clean-up and preconcentration is accomplished by using a C-18 minicolumn and 75% methanol as eluent. The limit of detection of NSAIDs can be up to 400-fold improved when using sample clean-up. The extraction efficiency for these compounds is between 71.1 and 109.7 microg mL(-1) (RSD 2.0-7.7%) for urine samples and from 77.2 to 107.1 microg mL(-1) (RSD 3.5-7.1%) for serum samples. PMID- 11258760 TI - Nonaqueous capillary electrophoresis-mass spectrometry for separation of venlafaxine and its phase I metabolites. AB - Aqueous and nonaqueous capillary electrophoresis (NACE) were investigated for separation of venlafaxine, a new second-generation antidepressant, and its three phase I metabolites. Working at basic pH, around the venlafaxine pKa value, was effective in resolving the investigated drugs, but created considerable peak tailing. To overcome electrostatic interactions between analytes and silanol groups, investigations were also carried out at acidic pH. However, despite the addition of up to 50% v/v of organic solvents (e.g., methanol or acetonitrile), complete separation of the studied compounds was not possible. NACE was found to be an appropriate alternative to resolve venlafaxine and its metabolites simultaneously. Using a conventional capillary (fused-silica, 64.5 cm length, 50 microm inner diameter), and a methanol-acetonitrile mixture (20/80 v/v) containing 25 mM ammonium formate and 1 M formic acid, complete resolution of these closely related compounds was performed in less than 3.5 min. Selectivity, efficiency and separation time were greatly affected by the organic solvent composition. As the electric current generated in nonaqueous medium was very low, the electric field was further increased by reducing the capillary length. This allowed a baseline resolution of venlafaxine and its three metabolities in 0.7 min. Selectivity was compared in aqueous and nonaqueous media in relation to the acid-base properties of the analytes as well as to the solvation degree. Finally, the method successfully coupled on-line to mass spectrometry with electrospray ionization interface allowed significant sensitivity enhancement. PMID- 11258761 TI - Analysis of metacycline by capillary electrophoresis. AB - The development and validation of an optimized capillary electrophoresis method for the determination of metacycline in the presence of its related substances by capillary electrophoresis is shown. The influence of methanol as organic modifier, buffer pH, buffer concentration, capillary length, column temperature, Triton X-100 and methyl-beta-cyclodextrin was investigated. A central composite design was performed in order to optimize the method. The optimal separation conditions were: uncoated fused-silica capillary (39 cm total length, 31 cm effective length, 50 microm ID); as background electrolyte a solution of 160 mM sodium carbonate and 1 mM EDTA (pH 10.35)/methanol (89:13 v/v); temperature, 15 degrees C; voltage, 12 kV. The method showed good selectivity, repeatability, linearity, and sensitivity. The limits of detection and quantitation are 0.024% and 0.06%, respectively, relative to a 2.5 mg/mL solution. Six commercial samples were analyzed quantitatively. PMID- 11258762 TI - Modelling of migration behaviour of inorganic anions in ion-exchange capillary electrochromatography. AB - A theoretical model to explain the observed mobility of inorganic anions in capillary electrochromatography (CEC) using ion-exchange (IE) stationary phases has been derived. The model divides contributions to the observed mobility of an analyte ion into capillary electrophoretic (CE) and IE components. The CE component includes the influence of varying the ionic strength of the background electrolyte on the electrophoretic mobility of the analyte, while the IE component accounts for the variation in retention of the analyte ion caused by changing the composition of the background electrolyte. The model was verified using a mixture of UV-absorbing inorganic ions in electrolytes of differing eluotropic strength in both packed and open-tubular CEC systems, with excellent agreement (r2 > 0.98) for both systems. Values of constants in the model equation determined by nonlinear regression were used to estimate the relative strengths of the interactions of different analytes with the stationary phase and these were found to agree well with elution orders observed in conventional IE chromatography. PMID- 11258763 TI - Capillary electrochromatography of hydrophobic amines on continuous beds. AB - The capillary electrochromatographic separation performance of hydrophobic amines and a related quaternary ammonium compound on continuous beds based on polymers of acrylamide has been studied. The chromatographic bed is polymerized in situ and the character of the polymers with regard to hydrophobicity and charge has been systematically changed by regulating its content of isopropyl and sulfonate ligands, respectively. The best performance was obtained for columns with a molar ratio of 1:80 for the sulfonate and isopropyl groups, and resulted in efficiencies up to 200000 plates per meter. The effects on retention, resolution and elution order by ionic strength, pH, and content of acetonitrile in the mobile phase have been investigated. The quaternary ammonium compound was always the least retained irrespective of pH. By increasing the pH, a reversal of the migration order between the tertiary and secondary amine was obtained. The results indicate a complex migration/retention mechanism where ion-exchange, adsorption and electrophoretic mobilities play a role. The concentration limit of detection could be lowered from 1.3 microg/mL to 50 pg/mL by using a high content of 2-propanol (96%) in the sample compared to dissolving the analytes in the mobile phase. PMID- 11258764 TI - Enantiomer separation by strong anion-exchange capillary electrochromatography with dynamically modified sulfated beta-cyclodextrin. AB - A novel mode of capillary electrochromatography (CEC) based on a dynamically modified stationary phase was presented for chiral separation. The capillary column was packed with strong anion-exchange (SAX) stationary phase packing; the sulfated beta-cyclodextrin (S-CD), which was added to the mobile phase, was dynamically adsorbed to the packing surface. Separation of enantiomers was achieved by their different abilities to form an inclusion complex with the adsorbed S-CD. The enantiomers of tryptophan, praziquantel, atropine, metoprolol, and verapamil were successfully separated in this system with a column efficiency of 36000-412000 plates/m. The resolution value obtained for atropine was as high as 11.23. The superiority of CEC with a dynamically modified stationary phase over that with a physically adsorbed stationary phase was demonstrated. The influence of ionic strength, S-CD concentration, and methanol content on separation was also studied. PMID- 11258765 TI - Determination of alkylphenol ethoxylates by micellar electrokinetic chromatography with bile salts. AB - Octyl- and nonylphenol ethoxylates (OPEs and NPEs) with different numbers of ethoxy units (average values: n = 10 and N = 40 for OPEs, and n = 10 for NPEs) were separated by micellar electrokinetic chromatography under positive polarity using an 80 mM borate buffer of pH 8.5 containing sodium deoxycholate (SDC) or sodium cholate (SC). When sodium dodecyl sulfate (SDS) was added to the background electrolyte (BGE) in the absence of the bile salt, a single peak at a migration time longer than that of the EOF was obtained. Substituting the SDS by a bile salt, the homologues were resolved. At the same bile salt concentration, resolution between the homologues was higher with SDC than using SC. Optimum resolution between consecutive homologues was obtained with 50 mM SDC. In the presence of low or moderate amounts of acetonitrile or n-propanol, the background line improved significantly, whereas resolution may increase or decrease slightly. We propose a procedure for the determination of OPEs and NPEs with optimum resolution between the homologues as well as a modified procedure with improved selectivity for the single-run determination of other absorbing nonionic, cationic, and anionic (such as linear alkylbenzene sulfonates) surfactants in industrial and household cleaning products and its application to a variety of samples. The detection limit was ca. 28 microg x mL(-1) of total NPE (n = 10), and peak area repeatabilities at 50 microg x mL(-1) were 1.7% (intraday) and 5.6% (interday). PMID- 11258766 TI - Use of vancomycin silica stationary phase in packed capillary electrochromatography I. Enantiomer separation of basic compounds. AB - Chiral separation of basic compounds was achieved by using 75 or 100 microm ID fused-silica capillaries packed with a vanoomycin-modified diol silica stationary phase. The capillary was firstly packed for about 12 cm with a slurry mixture composed of diolsilica (3:1) then with the vancomycin modified diol-silica (3:1) (23 cm), and finally with diol-silica (3:1) for about 2 cm. Frits were prepared by a heating wire at the two ends of the capillary; the detector window was prepared at 8.5 cm from the end of the capillary where vancomycin was not present. The influence of the mobile phase composition (pH and concentration, organic modifier type and concentration) on the velocity of the electroosmotic flow, chiral resolution and enantioselectivity was studied. Good enantiomeric resolution was achieved for atenolol, oxprenolol, propranolol, and venlafaxine using a mobile phase composition of 100 mM ammonium acetate solution (pH 6)/water/acetonitrile (5:5:90 v/v/v) while for terbutaline a mixture of 5:15:80 v/v/v provided the best separations. The use of methanol instead of acetonitrile caused a general increase of enantiomer resolution of the studied compounds together with a reduction of efficiency and detector response. However, the combination of acetonitrile and methanol in the mobile phase (as, e.g., 10% methanol and 80% acetonitrile) allowed to improve the enantiomer resolution with satisfactory detector response. PMID- 11258767 TI - Separation of basic, acidic and neutral compounds by capillary electrochromatography using uncharged monolithic capillary columns modified with anionic and cationic surfactants. AB - A mode of capillary electrochromatography (CEC), based on the dynamical adsorption of surfactants on the uncharged monolithic stationary phases has been developed. The monolithic stationary phase, obtained by the in situ polymerization of butyl methacrylate with ethylene dimethacrylate, was dynamically modified with an ionic surfactant such as the long-chain quaternary ammonium salt of cetyltrimethylammonium bromide (CTAB) and long-chain sodium sulfate of sodium dodecyl sulfate (SDS). The ionic surfactant was adsorbed on the surface of polymeric monolith by hydrophobic interaction, and the ionic groups used to generate the electroosmotic flow (EOF). The electroosmotic mobility through these capillary columns increased with increasing the content of ionic surfactants in the mobile phase. In this way, the synthesis of the monolithic stationary phase with binary monomers can be controlled more easily than that with ternary monomers, one of which should be an ionic monomer to generate EOF. Furthermore, it is more convenient to change the direction and magnitude of EOF by changing the concentration of cationic or anionic surfactants in this system. An efficiency of monolithic capillary columns with more than 140000 plates per meter for neutral compounds has been obtained, and the relative standard deviations observed for to and retention factors of neutral solutes were about 0.22% and less than 0.56% for ten consecutive runs, respectively. Effects of mobile phase composition on the EOF of the column and the retention values of the neutral solutes were investigated. Simultaneous separation of basic, neutral and acidic compounds has been achieved. PMID- 11258768 TI - A combination of chemical derivatisation and improved bioinformatic tools optimises protein identification for proteomics. AB - The identification of individual protein species within an organism's proteome has been optimised by increasing the information produced from mass spectral analysis through the chemical derivatisation of tryptic peptides and the development of new software tools. Peptide fragments are subjected to two forms of derivatisation. First, lysine residues are converted to homoarginine moieties by guanidination. This procedure has two advantages, first, it usually identifies the C-terminal amino acid of the tryptic peptide and also greatly increases the total information content of the mass spectrum by improving the signal response of C-terminal lysine fragments. Second, an Edman-type phenylthiocarbamoyl (PTC) modification is carried out on the N-terminal amino acid. The renders the first peptide bond highly susceptible to cleavage during mass spectrometry (MS) analysis and consequently allows the ready identification of the N-terminal residue. The utility of the procedure has been demonstrated by developing novel bioinformatic tools to exploit the additional mass spectral data in the identification of proteome proteins from the yeast Saccharomyces cerevisiae. With this combination of novel chemistry and bioinformatics, it should be possible to identify unambiguously any yeast protein spot or band from either two-dimensional or one-dimensional electropheretograms. PMID- 11258769 TI - The effect of protease inhibitors on the two-dimensional electrophoresis pattern of red blood cell membranes. AB - The last few years have brought dramatic improvements for sample preparation and solubilization of protein for electrophoretic analyses. The use of reagents such as thiourea and novel sulfobetaine surfactants increases the total number of proteins able to be visualized from a complex mixture such as a cell lysate and also allows more hydrophobic membrane proteins to be resolved. As the red blood cell (RBC) contain no organelles, it is an ideal source of relatively pure plasma membrane for protein solubilization studies. In addition, there are a number of diseases related to abnormalities of RBCs proteins, thus it is of medical relevance as well as a test sample for technology development. However, the procedure for purifying RBC membranes is rather time-consuming and is normally carried out under almost physiological conditions, which can be conducive to proteolytic degradation of the membrane proteins. Significant differences in two dimensional (2-D) patterns with and without protease inhibitors in sample preparation are demonstrated. In addition, is shown that preparation of RBC membranes with sodium carbonate, pH 11, leads to multimeric complexes of hemoglobin and causes hemoglobin to be irreversibly attached to the membrane. When using immobilized pH gradients (IPG) as the first dimension, it is demonstrated that the spectrins (large, filamentous proteins of 280 kDa) are lost from the 2-D map unless active, instead of passive, sample hydration into the IPG strip is adopted. PMID- 11258770 TI - Characterization of differently processed forms of enolase 2 from Saccharomyces cerevisiae by two-dimensional gel electrophoresis and mass spectrometry. AB - Two-dimensional gel electrophoresis, bioinformatics, and mass spectrometry are key analysis tools in proteome analysis. The further characterization of post translational modifications in gel-separated proteins relies fully on data obtained by mass spectrometric analysis. In this study, stress-induced changes in protein expression in Saccharomyces serevisiae were investigated. A total of eleven spots on a silver-stained two-dimensional (2-D) gel were identified by matrix-assisted laser desorption/ionization (MALDI) peptide mass mapping to represent C and/or N-terminal processed forms of enolase 2. The processing sites were determined by MALDI peptide mass mapping using a variety of proteolytic enzymes, by optimizing the sample preparation procedure and by specific labeling of all C-termini derived from in-gel digestion using a buffer containing 16O:18O (1:1). Out of eleven processed forms of enolase 2, six were fully characterized and the approximate processing sites identified for the remaining five. PMID- 11258771 TI - Expression of the small tyrosine phosphatase (Stp1) in Saccharomyces cerevisiae: a study on protein tyrosine phosphorylation. AB - Small tyrosine phoshatase 1 (Stp1) is a Schizosaccharomyces pombe low-molecular mass phosphotyrosine-phosphatase 50% identical to Saccharomyces cerevisiae Ltp1. In order to investigate the role of Stp1 in yeast, a mutant was generated having the characteristic of a dominant negative molecule. Changes in protein tyrosine phosphorylation in S. cerevisiae proteome in response to Stp1 or its dominant negative mutant expression were analyzed by high-resolution two-dimensional (2-D) electrophoresis. The most remarkable result is the modification by phosphorylation on tyrosine of several proteins involved in carbohydrate metabolism. Twelve proteins were identified on the basis of their positions in the anti-phosphotyrosine immunoblot of the 2-D electrophoresis. Ten of these present tyrosyl residues that are within the consensus sequence for protein kinase CK2 (casein kinase-2). These data open the possibility for the identification of Stp1 substrates in yeast and provide hints about the nature of tyrosine phosphorylating agents in yeast and in other organisms where bona fide tyrosine kinases are lacking. PMID- 11258772 TI - Proteome analysis of cultivar-specific interactions between Rhizobium leguminosarum biovar trifolii and subterranean clover cultivar Woogenellup. AB - Proteome analysis was used to identify proteins that are involved in the early stages of nodulation between the subterranean clover cultivar Woogenellup and the Rhizobium leguminosarum bv. trifolii strains ANU843 and ANU794. Strain ANU843 induces nitrogen-fixing nodules whereas strain ANU794 forms aberrant nodules on the roots of cv. Woogenellup that fail to develop beyond an early stage. Our aim was to identify proteins that might be involved in the early stages of nodulation over a 48 h period and to identify proteins that are differentially displayed during the interactions between the host and the two microbes. Proteome maps from control Woogenellup roots and inoculated roots were generated and compared at 24 and 48 h post inoculation. Over 1500 spots were resolved on all gels. Of the 16 protein spots that were differentally displayed or developmentally regulated, 10 were assigned putative identities. These included an alpha-fucosidase, several ethylene-induced proteins, a Cu/Zn superoxide dismutase, a hypothetical 16.5 kDa protein, tubulin alpha-chain, chaperonin 21 precursor and triosephosphate isomerase. Of the 22 constitutively expressed proteins spots examined, eight spots were assigned putative protein homologies through N-terminal sequencing and included several pathogenesis and stress-related proteins. The result may suggest that ethylene levels are upregulated during the early stages of infection but that this does not result in the induction of common pathogenesis-related proteins. The specific induction of alpha-fucosidase by ANU794 may be important in the nodulation failure phenotype of strain ANU794. PMID- 11258773 TI - Long-term results of a prospective surgical trial comparing 2 cm vs. 4 cm excision margins for 740 patients with 1-4 mm melanomas. AB - BACKGROUND: The Intergroup Melanoma Surgical Trial began in 1983 to examine the optimal surgical margins of excision for primary melanomas of intermediate thickness (i.e., 1-4 mm). There is now a median 10-year follow-up. METHODS: There were two cohorts entered into a prospective multi-institutional trial: (1) 468 patients with melanomas on the trunk or proximal extremity who randomly received a 2 cm or 4 cm radial excision margin and (2) 272 patients with melanomas on the head, neck, or distal extremities who received a 2 cm radial excision margin. RESULTS: A local recurrence (LR) was associated with a high mortality rate, with a 5-year survival rate of only 9% (as a first relapse) or 11% (anytime) compared with an 86% survival for those patients who did not have a LR (P < .0001). The 10 year survival for all patients with a LR was 5%. The 10-year survival rates were not significantly different when comparing 2 cm vs. 4 cm margins of excision (70% vs. 77%) or comparing the management of the regional lymph nodes (observation vs. elective node dissection). The incidences of LR were the same for patients having a 2 cm vs. 4 cm excision margin regardless of whether the comparisons were made as first relapse (0.4% vs. 0.9%) or at anytime (2.1% vs. 2.6%). When analyzed by anatomic site, the LR rates were 1.1% for melanomas arising on the proximal extremity, 3.1% for the trunk, 5.3% for the distal extremities, and 9.4% for the head and neck. The most profound influence on LR rates was the presence or absence of ulceration; it was 6.6% vs. 1.1% in the randomized group involving the trunk and proximal extremity and was 16.2% vs. 2.1% in the non-randomized group involving the distal extremity and head and neck (P < .001). A multivariate (Cox) regression analysis showed that ulceration was an adverse and independent factor (P = .0001) as was head and neck melanoma site (P = .01), while the remaining factors were not significant (all with P > .12). CONCLUSION: For this group of melanoma patients, a local recurrence is associated with a high mortality rate, a 2-cm margin of excision is safe and ulceration of the primary melanoma is the most significant prognostic factor heralding an increased risk for a local recurrence. PMID- 11258774 TI - Risk factors for nodal recurrence after lymphadenectomy for melanoma. AB - BACKGROUND: The risk and outcome of regional failure after elective and therapeutic lymph node dissection (ELND/TLND) for microscopically and macroscopically involved lymph nodes without adjuvant radiotherapy were evaluated. METHODS: Retrospective melanoma database review of 338 patients (ELND 85, TLND 253) from 1970 to 1996 with pathologically involved lymph nodes. RESULTS: Regional recurrence occurred in 14% of patients treated with ELND (n = 12) and 28% of patients treated with TLND (n = 72; P = .009). Risk factors associated with nodal recurrence were advanced age, primary lesion in the head and neck region, depth of the primary lesion, number of involved lymph nodes, and extracapsular extension (ECE). For each nodal basin, the ELND group had a lower incidence of recurrence than the TLND group. The TLND group had larger lymph nodes, greater number of involved lymph nodes, and a higher incidence of ECE. The 10-year disease-specific survival was 51% vs. 30% for ELND and TLND, respectively (P = .0005). Nodal basin failure was predictive of distant metastasis, with 87% developing distant disease compared with 54% of patients without nodal recurrence (P < .0001). Of six patients who underwent a second dissection after isolated nodal recurrence, five patients have had a median disease-free interval of 79 months. CONCLUSIONS: After ELND or TLND, patients who have a large tumor burden (thick primary melanoma, multiply involved lymph nodes, ECE), advanced age, and a primary lesion located in the head and neck have a significantly increased likelihood of relapse and a decreased survival. Few patients present with an isolated nodal recurrence, but the majority can be salvaged by a second dissection. PMID- 11258775 TI - Significance of plasma cytokine levels in melanoma patients with histologically negative sentinel lymph nodes. AB - INTRODUCTION: Although sentinel lymph node (SLN) status is the most powerful predictor of prognosis in patients with clinically localized melanoma, a proportion of melanoma patients with histologically negative SLNs will still recur. It is hypothesized that tumor response may be altered or mediated by specific cytokines. We therefore investigated whether levels of IL-4, IL-6, IL 10, TNF-alpha, or IFN-gamma would predict disease recurrence in melanoma patients with histologically negative SLNs. METHODS: This prospective cohort study involved 218 patients with clinically localized melanoma who underwent a histologically negative SLN biopsy. Preoperative plasma cytokine levels were determined by enzyme-linked immunosorbent assay on these patients, as well as on 90 healthy controls. Kaplan-Meier life tables were constructed, and Cox proportional hazards analyses were performed to assess predictors of disease-free survival (DFS). RESULTS: At a median follow-up of 43 months, 33 of 218 patients (15%) had suffered disease recurrence. Melanoma patients had significant elevations of IL-4, IL-6, and IL-10 compared to healthy controls; levels of IFN gamma were less elevated in melanoma patients compared to controls. Despite this, melanoma patients with detectable IFN-gamma levels were at significantly higher risk for recurrence compared to patients with undetectable levels (5-year DFS 70% vs. 86%, P = .03). On multivariate analysis including standard melanoma prognostic factors, only tumor thickness (P = .004) and the presence of detectable IFN-gamma levels (P = .05) were significant independent prognostic factors for disease-free survival. CONCLUSIONS: Among melanoma patients with clinically localized disease who have undergone a histologically negative SLN biopsy, presence of a detectable plasma level of IFN-gamma is an independent predictor of disease recurrence. Elevated levels of IFN-gamma may identify a group of early-stage melanoma patients who are more likely to have recurrence of disease and who may benefit from adjuvant therapies, including immunotherapies. PMID- 11258776 TI - Neoadjuvant chemoradiotherapy for adenocarcinoma of the pancreas: treatment variables and survival duration. AB - BACKGROUND: For patients with potentially resectable pancreatic cancer, the poor outcome associated with resection alone and the survival advantage demonstrated for combined-modality therapy have stimulated interest in preoperative chemoradiotherapy. The goal of this study was to analyze the effects of different preoperative chemoradiotherapy schedules, intraoperative radiation therapy, patient factors. and histopathologic variables on survival duration and patterns of treatment failure in patients who underwent pancreaticoduodenectomy for adenocarcinoma of the pancreatic head. METHODS: Data on 132 consecutive patients who received preoperative chemoradiation followed by pancreaticoduodenectomy for adenocarcinoma of the pancreatic head between June 1990 and June 1999 were retrieved from a prospective pancreatic tumor database. Patients received either 45.0 or 50.4 Gy radiation at 1.8 Gy per fraction in 28 fractions or 30.0 Gy at 3.0 Gy per fraction in 10 fractions with concomitant infusional chemotherapy (5 fluorouracil, paclitaxel, or gemcitabine). If restaging studies demonstrated no evidence of disease progression, patients underwent pancreaticoduodenectomy. All patients were evaluated with serial postoperative computed tomography scans to document first sites of tumor recurrence. RESULTS: The overall median survival from the time of tissue diagnosis was 21 months (range 19-26, 95%CI). At last follow-up, 41 patients (31%) were alive with no clinical or radiographic evidence of disease. The survival duration was superior for women (P = .04) and for patients with no evidence of lymph node metastasis (P = .03). There was no difference in survival duration associated with patient age, dose of preoperative radiation therapy, the delivery of intraoperative radiotherapy, tumor grade, tumor size, retroperitoneal margin status, or the histologic grade of chemoradiation treatment effect. CONCLUSION: This analysis supports prior studies which suggest that the survival duration of patients with potentially resectable pancreatic cancer is maximized by the combination of chemoradiation and pancreaticoduodenectomy. Furthermore, there was no difference in survival duration between patients who received the less toxic rapid-fractionation chemoradiotherapy schedule (30 Gy, 2 weeks) and those who received standard fractionation chemoradiotherapy (50.4 Gy, 5.5 weeks). Short-course rapid fractionation preoperative chemoradiotherapy combined with pancreaticoduodenectomy, when performed on accurately staged patients, maximizes survival duration and is associated with a low incidence of local tumor recurrence. PMID- 11258777 TI - A new radiocolloid for sentinel node detection in breast cancer. AB - BACKGROUND: The optimal radioactive tracer and technique for sentinel lymph node localization in breast cancer is yet to be determined. The dilemma of small particle size with dispersion to second echelon nodes versus failure of migration of larger radiocolloids needs to be resolved. A new radiocolloid preparation with particle size under 0.1 micron was developed with excellent primary/post lymphatic entrapment ratio. OBJECTIVE: To assess the feasibility of a new 99mTc radiocolloid cysteine-rhenium colloid in sentinel lymph node (SLN) localization for breast cancer. METHODS: Forty-seven patients with newly diagnosed T1 or T2 breast cancer underwent injection of 99mTc-labeled cysteine-rhenium colloid followed by lymphoscintigraphy. Same day SLN biopsy with patent blue dye and intraoperative gamma probe to identify SLNs were performed. RESULTS: SLN mapping and intraoperative localization were successful in 46/47 (98%) of patients. The blue dye radioactive tracer concordance was 94%. There was one false-negative in a patient with a nonpalpable tumor that underwent ultrasound-guided peritumoral radiocolloid injection. CONCLUSIONS: 99mTc-cysteine-rhenium colloid is highly effective in identifying SLNs. It has the advantage of smaller particle size than sulfur colloid with easier lymphatic migration. It has a more neutral pH with less pain on injection and does not require filtration, thereby minimizing radiation exposure to technologists. PMID- 11258778 TI - Breast cancer after augmentation mammoplasty. AB - BACKGROUND: It is thought that implants interfere with breast cancer diagnosis and that cancers in women who have had breast augmentation carry a worse prognosis. METHODS: A prospective breast cancer database was reviewed, comparing augmented and nonaugmented patients for details of histology, palpability, tumor size, nodal status, mammographic status, receptor status, nuclear grade, stage, and outcome. RESULTS: Ninety-nine cancers in augmented women and 2857 cancers in nonaugmented women were identified. Among these women, mammography was normal in 43% of those who had had augmentation and in 5% of those who had not. Augmented women were more likely to have palpable cancers (83% vs. 59%) and nodal involvement (48% vs. 36%), and less likely to have ductal carcinoma in situ (DCIS) (18% vs. 28%). When comparing only women younger than 50, the differences in invasiveness and nodal status lost significance. Cancers diagnosed in the 1990s were more likely to be nonpalpable and noninvasive than those diagnosed in the 1980s. This trend was more pronounced in the augmented population. CONCLUSIONS: Augmented patients were more likely to have palpable cancers, although the overall stage and outcome were similar to those of nonaugmented women. Although there have been significant improvements in our ability to diagnose early breast cancer over the past two decades, mammography continues to be suboptimal in augmented women. PMID- 11258779 TI - Lymphovascular invasion enhances the prediction of non-sentinel node metastases in breast cancer patients with positive sentinel nodes. AB - BACKGROUND: Fifty percent of patients with sentinel lymph node (SLN) metastases have no metastatic disease in non-SLNs on axillary lymph node dissection (ALND). The goal of this study is to determine which patients have metastatic disease limited to the SLN, and, therefore, may not require completion ALND. METHODS: Of the first 1000 patients undergoing SLN biopsy at Memorial Sloan-Kettering Cancer Center, using a combined blue dye and isotope technique, 231 (26%) had positive SLN. Of these, 206 underwent completion ALND. They are the study group for this report. RESULTS: The likelihood of non-SLN metastasis was inversely related to three clinicopathologic variables: tumor size < or = 1.0 cm; absence of lymphovascular invasion (LVI); and SLN micrometastases (< or = 2 mm). None of 24 patients with all three predictive factors had non-SLN metastases, whereas 58% of patients with none of the factors had disease in the non-SLN. CONCLUSION: Patients with small breast cancers, no LVI, and SLN micrometastases have a low risk of non-SLN metastases, and may not require completion ALND. PMID- 11258780 TI - Validation of lymphatic mapping in colorectal cancer: in vivo, ex vivo, and laparoscopic techniques. AB - BACKGROUND: The use of lymphatic mapping (LM) is being investigated to improve the staging of colorectal cancer (CRC) and thereby identify patients who might benefit from adjuvant chemotherapy. This study evaluated in vivo, laparoscopic, and ex vivo approaches for the ultrastaging of CRC. METHODS: Seventy-five CRC patients were enrolled in a study of LM with peritumoral injection of isosulfan blue dye. LM was undertaken during open colon resection (OCR) in 64 patients, during laparoscopic colon resection (LCR) in 9 patients, and after specimen removal (ex vivo) in 2 patients. Ex vivo LM was also undertaken in 6 patients after unsuccessful in vivo LM. All nodes were examined by hematoxylin and eosin (H&E) staining; in addition, sentinel lymph nodes (SNs) were multisectioned and examined by immunohistochemical staining with cytokeratin (CK-IHC). RESULTS: At least one SN was identified in 72 patients (96%). In vivo LM identified SNs in 56 of 64 (88%) patients undergoing OCR and in 9 of 9 (100%) patients undergoing LCR. Ex vivo LM was undertaken as the initial mapping procedure in 2 cases of intraperitoneal colon cancer and after in vivo LM had failed in 6 cases of extraperitoneal rectal carcinoma; an SN was identified in 7 of the 8 cases. Focused examination of the SN correctly predicted nodal status in 53 of 56 OCR cases, 9 of 9 LCR cases, and 6 of 7 ex vivo cases. Multiple sections and CK-IHC identified occult micrometastases in 13 patients (17%), representing 10 OCR, 1 LCR, and 2 ex vivo cases. CONCLUSIONS: LM of drainage from a primary CRC can be accurately performed in vivo during OCR or LCR. Ex vivo LM can be applied when in vivo techniques are unsuccessful and may be useful for rectal tumors. During LCR, colonoscopic injection can be used to mark the primary tumor and define the lymphatic drainage so that adequate resection margins are obtained. These LM techniques improve staging accuracy in CRC. PMID- 11258781 TI - Detection and prediction of micrometastasis in the lymph nodes of patients with pN0 gastric cancer. AB - BACKGROUND: The clinicopathologic significance of micrometastasis (MM) and tumor cell microinvolvement (TCM) in regional lymph nodes as identified by immunohistochemical staining for cytokeratin expression was evaluated in patients with node-negative gastric cancer. METHODS: MM was defined as tumor cells with stromal reaction, and TCM was defined as individual tumor cells without stromal reaction. We investigated 1761 lymph nodes obtained from 67 gastric cancer patients whose diagnosis showed no lymph node metastasis by routine histological examination. The depth of tumor invasion was T1 (submucosa) in 33 patients and T2 (muscularis propria and subserosa) in 34 patients. The lymph nodes were examined immunohistochemically for the presence of tumor cells using anti-cytokeratin AE1/AE3 monoclonal antibody. Both the biopsy tumor specimens obtained prior to surgery and the resected primary tumors were immunostained with E-cadherin (E cad) monoclonal antibody. RESULTS: Thirty (1.5%) of the 1761 lymph nodes showed MM and/or TCM. MM with or without TCM was found in 10 patients, and TCM alone was found in 4 patients; 6 (18.2%) of the 33 patients with T1 tumor and 8 (23.5%) of the 34 patients with T2 tumor had occult lymph node metastasis. The 5-year survival rate was worse among those with MM with or without TCM, than among those without MM. Nearly all of the patients with MM and/or TCM had reduced or negative E-cad expression in the primary tumor. CONCLUSIONS: We demonstrated that the incidence of MM and/or TCM in the lymph nodes of patients with gastric cancer is quite high, and that such metastasis is associated with the prognosis of patients with pN0. Examination of E-cad expression in biopsy tumor specimens may be useful for predicting MM and/or TCM. PMID- 11258782 TI - Distal margin requirements after preoperative chemoradiotherapy for distal rectal carcinomas: are < or = 1 cm distal margins sufficient? AB - BACKGROUND: Sphincter-sparing alternatives to abdominoperineal resection (APR) in the treatment of rectal cancer often are underused out of concern for inadequate distal margins and local failure. The present study addresses whether sphincter sparing techniques with distal margins < or = 1 cm adversely influence oncological outcome in patients given preoperative chemoradiotherapy. METHODS: Thirty-seven patients with rectal cancer < or = 8 cm from the anal verge were enrolled in the study. Preoperative external beam radiotherapy (5400 Gy) was administered together with continuous infusion of 5-fluorouracil (300 mg/m2/day). Surgical resection was performed in 36 patients with pathological assessment of tumor response and margins. Patients with sphincter-sparing resection and distal margins > 1 cm or < or = 1 cm and those who underwent APR were compared. RESULTS: Thirty-six patients completed preoperative chemoradiotherapy, with successful sphincter-preservation in 28 patients. At a median follow-up of 33 months, there were 12 recurrences overall, which included 11 distant failures and four pelvic failures. Disease-free survival (DFS) was not different between those who had an APR compared with sphincter-sparing resection with distal margins < or = 1 cm. DFS was worse (P < .02) when radial margins were < or = 3 mm compared with > 3 mm. CONCLUSIONS: Sphincter preservation is feasible in more than 75% of patients with tumors < or = 8 cm from the anal verge after preoperative chemoradiotherapy. Sphincter-sparing surgery with distal margins < or = 1 cm can be used without adversely influencing local recurrence or DFS. Limited radial margins (< or = 3 mm), however, are associated with increased disease recurrence. PMID- 11258783 TI - Lymph node micrometastases in patients with early gastric cancer: experience with 139 patients. AB - BACKGROUND: Although lymph node metastases in patients with early gastric cancer (EGC) is an important prognostic factor, the prognostic relevance of lymph node micrometastases is still uncertain. METHODS: The authors studied 1488 lymph nodes, which were histologically confirmed as pN0, dissected from 139 patients who were treated for EGC between 1976-1994. Micrometastases were defined as a single or small cluster of neoplastic cells identifiable only by immunohistochemical methods. RESULTS: Lymph node micrometastases was observed in 24 of the 139 patients (17%). No significant correlation was observed between micrometastases and other clinicopathological characteristics. Analysis of overall survival showed no significant difference between the micrometastases positive and negative groups. CONCLUSION: The results of our study show that the presence of lymph node micrometastases in EGC does not have an influence on patient prognosis. PMID- 11258784 TI - VATS port site recurrence: a technique dependent problem. AB - OBJECTIVES: Video-assisted thoracic surgery (VATS) has become an accepted approach for the diagnosis and treatment of thoracic malignancies. Port site tumor recurrence is a reported complication of VATS. However, the true incidence of this problem is unknown. To try to determine the incidence of port site recurrence, we analyzed our experience with patients undergoing VATS wedge resection for malignancy. METHODS: Data were obtained from our prospective VATS database. The analysis was confined to patients undergoing VATS wedge resection for malignancy, excluding those having a pleural biopsy only. Parameters analyzed included demographic factors, surgical technique, and port site recurrences identified by physical examination, CT scan, or both. RESULTS: From 1992 to 1996, 410 patients (182 men, 228 women; median age = 61 years) underwent a VATS wedge resection for malignancy. The procedure was performed for diagnosis or staging in 90% of cases. Access incisions plus port sites were used in 97 (24%) patients; port sites only were used in 313 (76%) patients. Conversion to thoracotomy was necessary in 102 patients (25%) either for definitive resection (58 patients) or because VATS was not technically adequate (44 patients). Specimens were retrieved via access incisions or port sites with or without a specimen bag. The operative mortality was 0.25%. With long-term follow-up (median = 25 months) available for 374 patients (91%), only one port site recurrence was identified (0.26%). CONCLUSION: Our experience confirms the safety of VATS wedge resection in cancer patients. The incidence of port site tumor recurrence is low when oncologic principles are respected. In our institution, these principles include performing VATS wedge resection only for lesions that can be widely removed; converting to thoracotomy for definitive or extensive cancer operation; and using meticulous technique for the extraction of specimens from the pleural space. PMID- 11258785 TI - Treatment of nosocomial postoperative pneumonia in cancer patients: a prospective randomized study. AB - BACKGROUND: Nosocomial pneumonia continues to be associated with high morbidity and mortality in cancer patients. METHODS: In an attempt to find an optimal treatment for this infection, nonneutropenic cancer patients with postoperative nosocomial pneumonia were randomized to receive either piperacillin/tazobactam (P/T) 4.5 g i.v. every 6 hours (30 patients) or clindamycin (Cl) 900 mg plus aztreonam (Az) 2 g i.v. every 8 hours (22 patients). Amikacin 500 mg i.v. every 12 hours was given to all patients for the first 48 hours. RESULTS: The two groups were comparable for the characteristics of pneumonia that included gram negative etiology and duration of intubation. Response rates were 83% for patients who received P/T and 86% for those who received Cl/Az (P > .99). There were no serious adverse events; however, at our center the cost of the P/T regimen was $73.86 compared with $99.15 for the Cl/Az regimen. CONCLUSIONS: The two regimens had comparable high efficacy, and P/T had a slight cost advantage. Either of these antibiotic regimens combined with an aminoglycoside could lead to favorable outcome in cancer patients at high risk for nosocomial pneumonia. PMID- 11258786 TI - The thyroid nodule. PMID- 11258787 TI - Establishing a standard of care for the patient with melanoma. PMID- 11258788 TI - Role of fine-needle aspiration biopsy and frozen section analysis in the surgical management of thyroid tumors. AB - INTRODUCTION: The role of fine-needle aspiration (FNA) and frozen section (FS) in the management of thyroid neoplasms continues to generate considerable controversy. We reviewed our recent experience to determine the clinical utility of FNA and FS in our surgical management and intraoperative decision-making. METHODS: All patients who had operations for thyroid disease between January 1996 and June 1999 were identified in our prospective database. Completion and incidental thyroidectomies were excluded. Data obtained from the pathology files included FNA, FS, and the final histologic diagnosis. RESULTS: Five hundred sixty four patients, including 409 women (73%), with a median age of 50 years (range, 6 94) were identified, of whom 293 (52%) had cancer diagnosed on permanent sections. Three hundred twenty-nine patients (58%) had evaluable FNA, of which 91 (28%) were benign, 94 were malignant (28%), and 144 (44%) were suspicious (46% of these were malignant on final). Frozen section was performed in 397 (70%) patients; of these samples, 170 (43%) were found to be benign, 106 (27%) were malignant, and 121 (30%) were deferred (46% malignant on final). Fine-needle aspiration positively identified 51% of confirmed malignancies; 13% of patients with malignancy had a benign FNA result. Total thyroidectomy was performed in 64% of malignant tumors and 29% of benign thyroid disease (P < .001). Logistic regression revealed no association of extent of surgery with FNA results. A frozen section positive for malignancy was associated with total thyroidectomy (P < .001, RR 6 [CI 3-10]), and a negative frozen section report was associated with lobectomy (P < .05, RR 0.5 [CI 0.3-0.96]). Frozen sections results altered the preoperative plan in only 29 patients (5%). CONCLUSION: Results of preoperative FNA had no direct impact on the selection of the surgical procedure in this selected cohort. Intraoperative FS added very little to surgical management. The majority of thyroid operations at this institution are planned and performed based on known prognostic factors and intraoperative findings. PMID- 11258789 TI - CD58/LFA-3 and IL-12 provided by activated monocytes are critical in the in vitro expansion of CD56+ T cells. AB - A small proportion of human CD3+ T lymphocytes are known to co-express CD56, an antigen usually restricted in its expression to natural killer (NK) cells. Whereas the in vivo function of CD3+ CD56+ T cells remains unknown, we and others have previously shown that both in vitro and in vivo, these cells can mediate a significantly greater degree of MHC-unrestricted cytotoxicitv against a variety of human tumor cells when compared to either CD3+ CD56- T cells or lymphokine activated killer (LAK) cells. While the mechanismns regulating the in vivo expansion of CD56+ T cells are not known, here we demonstrate the importance of CD2-mediated IL-12-dependent signals in the in vitro expansion of CD56+ T cells. Specifically, we show that activated monocytes provide a contact dependent factor (CD58/LFA-3) and a soluble factor (IL-12), both critical for the in vitro expansion of CD56+ T cells. The biological and therapeutic implications of these findings are discussed. PMID- 11258790 TI - Effects of granulocyte-colony-stimulating factor and granulocyte/macrophage colony-stimulating factor administration on T cell proliferation and phagocyte cell-surface molecules during hematopoietic reconstitution after autologous peripheral blood progenitor cell transplantation. AB - Thirty-four ovarian and breast cancer patients received autologous peripheral blood progenitor cell transplantation after high-dose myeloablative chemotherapy and either granulocyte-colony-stimulating factor (G-CSF) or granulocyte/macrophage-colony-stimulating fictor (GM-CSF) in the immediate post transplant period. The recovery of T cell functionality was monitored by a three color flow-cytometric approach using carboxyfluorescein diacetate succinimidyl ester, a probe the fluorescence intensity of which halves at each round of cell replication, in conjunction with CD3 and CD25 monoclonal antibodies. There was no significant difference between the two treatments on days 12, 20, and 40, T cell proliferation always being considerably lower than that of control cultures from healthy donors. At day 80, a significantly higher proportion of mitogen stimulated T cells from GM-CSF-treated patients expressed interleukin-2 receptor, and a higher proportion of these T cells were actively proliferating. This phenomenon did not reflect any difference in the relative proportion of various lymphocyte subsets (T cells, CD4 and CD8+ T cells, CD45RA+ and CD45RO- T cells, and natural killer cells). At the end of follow-up (1-1.5 years) T cell proliferation had returned to values typically observed in healthy individuals in both groups of patients. Soon after transplantation (day 12), neutrophils from G CSF-treated patients had a more elevated Fcgamma receptor I density and monocytes from GM-CSF-treated patients had a more elevated Fcgamma receptor II and MHC class II molecules density. The up-modulation of Fcgamma receptor II was maintained until day 40. Thus, administering G-CSF and GM-CSF in the post transplant period affects T lymphocyte proliferation and phagocyte membrane molecules differently. PMID- 11258791 TI - A murine B cell lymphoma expressing human HER2 / neu undergoes spontaneous tumor regression and elicits antitumor immunity. AB - In the present study we describe a novel murine tumor model in which the highly malignant murine B cell lymphoma 38C13 has been transduced with the cDNA encoding human tumor-associated antigen HER2/neu. This new cell line (38C13-HER2/neu) showed stable surfiace expression but not secretion of human HER2/neu. It also maintained expression of the idiotype (Id) of the surface immunoglobulin of 38C13, which serves as another tumor-associated antigen. Surprisingly, spontaneous tumor regression was observed following s.c. but not i.v. injection of 38C13-HER2/neu cells in immunocompetent syngeneic mice. Regression was more frequently observed with larger tumor cell challenges and was mediated through immunological mechanisms because it was not observed in syngeneic immunodeficient mice. Mice that showed complete tumor regression were immune to challenge with the parental cell line 38C13 and V1, a variant of 38C13 that does not express the Id. Immunity could be transferred with sera, suggesting that an antibody response mediated rejection and immunity. Continuously growing s.c. tumors as well as metastatic tumors obtained after the i.v. injection of 38C13-HER2/neu maintained expression of human HER2/neu, which can serve as a target for active immunotherapy. As spontaneous tumor regression has not been observed in other human murine models expressing human HER2/neu, our results illustrate the enormous differences that can exist among different murine tumors expressing the same antigen. The present model provides a useful tool for the study of the mechanisms of protective immunity to B cell lymphoma and for the evaluation of different therapeutic approaches based on the stimulation or suppression of the immune response. PMID- 11258792 TI - 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone modulation of cytokine release in U937 human macrophages. AB - The nicotine-derived N-nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1 butanone (NNK), is one of the most abundant and potent carcinogens found in tobacco smoke. NNK induces lung tumors in rodents and is most likely involved in lung carcinogenesis in humans. Studies on the metabolism and carcinogenicity of NNK have been extensive. However, its effects on the immune system have not been investigated thoroughly. Considering that tobacco smoking partially suppresses the immune response in humans, and that immune surveillance plays a critical role in cancer development, we examined the effects of NNK on the production of selected cytokines. In a previous study, we observed an inhibition of NK cell activity and IgM secretory cell number in NNK-treated A/J mice [Rioux and Castonguay (1997) J Natl Cancer Inst 89: 874]. In this study, we demonstrate that U937 human macrophages activate NNK to alkylating intermediates by alpha-carbon hydroxylation and detoxify NNK by N-oxidation. We observed that NNK, following activation, induces the release of soluble tumor necrosis factor (TNF), but inhibits interleukin(IL)-10 synthesis. We also report that 4 (acetoxymethylnitrosamino)-1-(3-pyridyl)- -butanone, and nitroso(acetoxymethyl)methylamine, which generate the same alkylating intermediates as NNK, have similar effects on TNF and IL-10. This suggests that pyridyloxobutylating and methylating intermediates generated from NNK are potent modulators of the immune response. The levels of IL-6, granulocyte/macrophage colony-stimulating factor and macrophage chemotactic protein 1 were also decreased in supernatants of NNK-treated U937 macrophages. In contrast, IL-2 synthesis in Jurkat cells was inhibited by NNK treatment. This is the first study demonstrating that NNK, via its alkylating intermediates, alters the cytokine synthesis profile in human cells. Modulation of cytokine synthesis by NNK might partially explain the immunosuppresion observed in smokers. Inhibition of immune functions, resulting from NNK activation to alkylating agents, may facilitate lung tumor development. PMID- 11258793 TI - The self peptide annexin II (208-223) presented by dendritic cells sensitizes autologous CD4+ T lymphocytes to recognize melanoma cells. AB - Annexin II is known to be over-expressed in different types of tumours. We show here that annexin II protein is expressed by melanoma cell lines in various amounts, consistent with previous findings that an annexin II (208-223) peptide could be eluted from isolated HLA-DR molecules of a constitutively MHC class II positive melanoma line. T cells sensitized to annexin II (208-223) in vitro using peptide-pulsed autologous dendritic cells responded only to the lines which overexpressed annexin II, in a peptide-specific, HLA-DR-restricted fashion. These CD4+ T cells proliferated strongly and secreted large amounts of type 1 cytokines in response to annexin II (208-223) peptide or annexin II protein-positive melanoma cell lines. These results demonstrate that the annexin II (208-223) peptide, corresponding to a non-mutated sequence of a normal protein, induces antigen-specific T cells which can respond to melanoma cells over-expressing the annexin II molecule. This peptide may therefore be useful in immunotherapy for recruiting CD4+ type 1 helper cells active locally in the tumour environment. PMID- 11258794 TI - Production and validation of the pharmacokinetics of a single-chain Fv fragment of the MGR6 antibody for targeting of tumors expressing HER-2. AB - The HER-2 antigen, which is overexpressed in many breast carcinomas, is an ideal target for monoclonal antibodies due to its low expression in normal tissue and its homogeneous distribution in the tumor mass. We have developed and characterized the murine MAb MGR6 against HER-2, which is able to inhibit proliferation of tumor cells overexpressing HER-2. On the basis of these preclinical results, phase I studies in breast carcinoma patients were conducted and radiolocalization data indicated an antibody half life which directly paralleled that of other whole antibodies and thus resulting in a limited in vivo diagnostic capacity. To obtain a smaller reagent with possibly improved in vivo properties, a single chain variable fragment (scFv) of the original MGR6 producing hybridoma was generated by phage display technology. Biologically active MGR6 scFv was purified rapidly and at high yield by metal affinity chromatography. Competition FACS and ELISA analyses identified an epitope on the HER-2 extracellular domain that was shared by the scFv and the parental MAb. BlAcore analysis indicated a Koff of 9.3 x 10(-4) s(-1), similar to that of the intact MGR6 MAb. Distribution and elimination half-lives of MGR6 scFv, calculated from in vivo preclinical evaluations, were much faster (13 min and 6.2 h, respectively) than previously published results for the intact MAb (mean t1/2beta of 46 h). This represents a theoretical improvement in pharmacokinetics with respect to the parental murine MAb and points to the potential for utilizing this fragment in redirecting therapeutic agents, such as radioisotopes, to different human carcinomas overexpressing HER-2. PMID- 11258795 TI - Identification of novel transcribed sequences on human chromosome 22 by expressed sequence tag mapping. AB - To identify sequences on the human genome that are actually transcribed, we mapped expressed sequence tags (ESTs) of long cDNAs ranging from 4 kb to 7 kb along a 33.4-Mb sequence of human chromosome 22, the first human chromosome entirely sequenced. By the EST mapping of 30,683 long cDNAs in silico, 603 cDNA sequences were found to locate on chromosome 22 and classified into 169 clusters. Comparison of the genomic loci of these cDNA sequences with 679 genes already annotated on chromosome 22q revealed that 46 clusters represented newly identified transcribed sequences. To further characterize these sequences, we sequenced 12 cDNAs in their entirety out of 46 clusters. Of these 12 cDNAs, 6 were predicted to include a protein-coding region while the remaining 6 were unlikely to encode proteins. Interestingly, 3 out of the 12 cDNAs had the nucleotide sequences of the opposite strands of the genes previously annotated, which suggested that these genomic regions were transcribed bi-directionally. In addition to these newly identified 12 cDNAs, another 12 cDNAs were entirely sequenced since these cDNAs were likely to contain new information about the predicted protein-coding sequences previously annotated. In the cases of KIAA1670 and KIAA1672, these single cDNA sequences covered two separately annotated transcribed regions. For example, the sequence of a clone for KIAA1670 indicated that the CHKL and CPT1B genes were co-transcribed as a contiguous transcript without making both the protein-coding regions fused. In conclusion, the mapping of ESTs derived from long cDNAs followed by sequencing of the entire cDNAs provided indispensable information for the precise annotation of genes on the genome together with ESTs derived from short cDNAs. PMID- 11258796 TI - Complete genome sequence of enterohemorrhagic Escherichia coli O157:H7 and genomic comparison with a laboratory strain K-12. AB - Escherichia coli O157:H7 is a major food-borne infectious pathogen that causes diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Here we report the complete chromosome sequence of an O157:H7 strain isolated from the Sakai outbreak, and the results of genomic comparison with a benign laboratory strain, K-12 MG1655. The chromosome is 5.5 Mb in size, 859 Kb larger than that of K-12. We identified a 4.1-Mb sequence highly conserved between the two strains, which may represent the fundamental backbone of the E. coli chromosome. The remaining 1.4-Mb sequence comprises of O157:H7-specific sequences, most of which are horizontally transferred foreign DNAs. The predominant roles of bacteriophages in the emergence of O157:H7 is evident by the presence of 24 prophages and prophage like elements that occupy more than half of the O157:H7-specific sequences. The O157:H7 chromosome encodes 1632 proteins and 20 tRNAs that are not present in K 12. Among these, at least 131 proteins are assumed to have virulence-related functions. Genome-wide codon usage analysis suggested that the O157:H7-specific tRNAs are involved in the efficient expression of the strain-specific genes. A complete set of the genes specific to O157:H7 presented here sheds new insight into the pathogenicity and the physiology of O157:H7, and will open a way to fully understand the molecular mechanisms underlying the O157:H7 infection. PMID- 11258797 TI - A genome-wide analysis of transcriptional effect of Gal11 in Saccharomyces cerevisiae: an application of "mini-array hybridization technique". AB - The Gal11 protein is a subunit of the Mediator complex. Biochemical as well as genetic studies have strongly suggested that Gal11 is a positive global regulator of transcription. Some reports argue that Gal11 is a negative regulator, however. Here we have adopted the "Mini-array membrane hybridization" to analyze the effect of Gal11 in a genome-wide fashion. This technique has been demonstrated to be reliable to identify genes whose expression is controlled by a specific set of genetic and/or physiological signals. Our experiments indicate that this technique is applicable to profile the gene expression in yeast grown in rich medium. Thus mRNAs of 40% of significantly expressed genes are reduced more than two fold in gal11null yeast, in which only 3% of mRNAs are increased more than two fold. These results strongly suggest that Gal11 functions globally as a positive regulator in vivo. PMID- 11258798 TI - Characterization of equine microsatellites and microsatellite-linked repetitive elements (eMLREs) by efficient cloning and genotyping methods. AB - We performed efficient cloning and genotyping methods for isolation of a large number of polymorphic microsatellites. The methods contain the time-efficient cloning method of constructing microsatellite-enriched libraries and the economic genotyping method of fluorescent labeling of PCR products. Eighty novel equine microsatellites cloned were efficiently isolated from the enrichment library and analyzed for genotype polymorphism. Of these, 72 microsatellites were analyzed with a good resolution. The average heterozygosity of all loci was 0.52, and the number of alleles ranged from one to 9 with an average of 4.5 alleles. The other eight loci showed multiple bands of PCR products, suggesting the occurrence of microsatellites in a repetitive element, in which the number of microsatellite repeats varies among different members of the repetitive element. We found five homologous groups at flanking regions in comparison with the flanking regions of microsatellites from DNA databases. One of them showed homology to equine repetitive element-2. In the other four homologous groups, the two groups were named equine microsatellite-linked repetitive element-1 (eMLRE-1) and equine microsatellite-linked repetitive element-2 (eMLRE-2) as novel equine repetitive elements identified from equine genome. These data should help the analysis of equine DNA sequences and the design of equine genome markers. PMID- 11258800 TI - Antigen-induced bronchial hyperreactivity and bronchopulmonary inflammation in rats: effect of TNF receptor binding protein. AB - The effect of a receptor binding protein for tumor necrosis factor (TNFrbp) on cell infiltration, bronchial hyperreactivity, and release of inflammatory mediators were studied following antigen challenge in sensitized rats. A 3-fold increase in total cell number, mainly neutrophils and eosinophils, was noted in bronchoalveolar lavage (BAL) fluid 8 hours after antigen challenge. Antigen challenge also induced a significant hyperreactivity of the lower bronchus to carbachol and serotonin, but did not affect the reactivity of the trachea and upper bronchus. This increased responsiveness of the lower bronchus was transient, being detected 8 hours but not 24 hours after antigen challenge. Thromboxane B2 (TxB2), prostaglandin E2 (PGF2), and nitric oxide (NO) levels increased in the BAL fluid of sensitized rats 8 hours after antigen challenge by 197%, 172%, and 173%, respectively. TNFrbp treatment reduced by 83% the antigen induced cell infiltration, with neutrophils being the cells most affected. The bronchial hyperreactivity induced by antigen challenge was also significantly inhibited by TNFrbp, whereas TxB2, PGE2, and NO levels in the BAL fluid were not affected. In our animal model, the cell infiltration and bronchial hyperreactivity appear to be mediated to some extent by TNF, but not by prostanoids nor NO. PMID- 11258801 TI - Developmental expression of catenins and associated proteins during submucosal gland morphogenesis in the airway. AB - Although lymphoid enhancer binding factor-1 (Lef-1) plays an obligatory role in airway submucosal gland (SMG) development, its expression alone is not an adequate signal for initiating gland morphogenesis. Because Lef-1 forms a bipartite transcription factor with beta-catenin to mediate wnt pathway signaling, we investigated the expression of beta-catenin and associated proteins during SMG development with both in situ hybridization and immunocytochemistry. Unexpectedly, high levels of E-cadherin mRNA were expressed by cells in developing gland buds from the earliest stages through subsequent differentiation into mature glands. In contrast, a decreased level of E-cadherin immunoreactivity in stage I gland bud cells suggested that post-translational modulation of E cadherin protein levels may play a critical role in early stages of gland morphogenesis. Adenomatous polyposis coli (APC) mRNA was expressed relatively weakly in the developing ferret trachea, but higher levels of protein staining were observed throughout the cytoplasm of gland buds and surface epithelial cells. B-Catenin mRNA was abundantly expressed throughout the tracheal epithelium and at the highest levels in primordial gland buds. B-Catenin protein localized to the basolateral membranes of all airway epithelial cell types. However, no detectable increases in nuclear or cytoplasmic staining were associated with gland buds, as would be expected if beta-catenin served as a transcriptional cofactor for Lef-1 in gland morphogenesis. Additional studies demonstrated the gamma-catenin distribution to be remarkably similar to that of beta-catenin, whereas alpha-catenin staining was more diffuse in the cytoplasm of airway epithelial and gland bud cells. These descriptive results do not rule out a role for wnt signaling in SMG development , but provide no evidence that beta-catenin, or gamma-catenin, is a cofactor in Lef-1 regulation of SMG development. PMID- 11258802 TI - Modulation of transalveolar fluid absorption by endogenous aldosterone in adult rats. AB - We examined whether transalveolar fluid transport is modulated by aldosterone in adult rats. Because colocalization of mineralocorticoid receptors (MR) with 11beta-hydroxysteroid dehydrogenase type 2 (11betaHSD2) is important for aldosterone specific action, we first determined the immunohistochemical distribution of MR and 11betaHSD2 in the lung. We found that alveolar epithelial cells express both MR and 11betaHSD2. Reverse transcriptase polymerase chain reaction (RT-PCR) demonstrated that rat alveolar type II epithelial cells express both MR and 11betaHSD2. We then measured alveolar fluid clearance in rats treated with chronic low-sodium diet. A low-sodium diet (0.1% NaCl for 12 to 14 days) caused hyperaldosteronism accompanied by hypokalemia, whereas serum corticosterone and adrenaline levels remained normal. We found that hyperaldosteronism was associated with significantly higher alveolar fluid clearance and that this increase was related to the amiloride-sensitive component. In addition, the increase in alveolar fluid clearance was inhibited by spironolactone. Our results show that aldosterone is able to stimulate Na+ channels of alveolar epithelial cells. We conclude that alveolar epithelium is a physiological target tissue for aldosterone and transalveolar fluid absorption could in part be modulated by endogenous aldosterone acting via MR. PMID- 11258803 TI - Infection of human lung fibroblasts with Epstein-Barr virus causes increased IL 1beta and bFGF production. AB - An association between Epstein-Barr virus (EBV) infection and fibroblast proliferation in the interstitial spaces of the lung has been suggested in idiopathic interstitial pneumonia. In this study we show that EBV can infect human lung fibroblasts in vitro. A primary-cultured human lung fibroblast cell line, designated CCD-32Lu, expressed EBV nuclear antigen 1 after coculture with lethally irradiated EBV producing cells. The infection further induced CCD-32Lu cells to produce the fibrogenic cytokines basic fibroblast growth factor (bFGF) and interleukin-1beta. These findings indicate that lung fibroblasts may be a target for EBV infection and suggest that EBV may play a role in increased production of these cytokines and induce fibroblast proliferation in idiopathic interstitial pneumonia. PMID- 11258805 TI - Interactions between sediments and water: summary of the Eighth International Symposium. PMID- 11258804 TI - Inflammatory cytokines modulate eotaxin release by human lung fibroblast cell line. AB - Eotaxin, a potent eosinophil-specific chemotactic factor, is increased in the lower respiratory tract of asthma patients. Recently, lung fibroblasts have been reported to produce eotaxin and their activation can be modulated by inflammatory cytokines. To test the hypothesis that inflammatory cytokines modulate the eotaxin release from lung fibroblasts, we investigated the potential of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), or interferon-gamma (IFN-gamma) to induce the release of eotaxin and eotaxin mRNA by the human fetal lung fibroblast cell line, HFL-1, was evaluated. HFL-1 released eotaxin constitutively without stimulation, but IL-1beta or TNF-alpha stimulated eotaxin release in a dose- and time-dependent manner. IL-1beta or TNF-alpha treatment of HFL-1 also resulted in the augmented expression of eotaxin mRNA. Although IFN-gamma alone had negligible effect on eotaxin release and mRNA expression, IFN-gamma induced a significant, concentration-dependent attenuation of eotaxin release and eotaxin mRNA expression from HFL-1 stimulated with IL 1beta or TNF-alpha. These findings are consistent with the concept that lung fibroblast-derived eotaxin may in part be responsible for the eosinophil infiltration observed in the airways of asthmatic patients and that network of cytokines may modulate the eosinophil recruitment to the airways by stimulation of fibroblasts to release eotaxin. PMID- 11258806 TI - Sediment nutrient characteristics and aquatic macrophytes in lowland English rivers. AB - Aquatic macrophytes play an important role in the nutrient dynamics of streams. As a result, there is much interest in their use as trophic indicators. However, the relationship between aquatic macrophytes and the trophic status of rivers is a complex one, partly because of the effects of a wide range of environmental variables and partly because submerged, rooted macrophytes can absorb nutrients from the river sediments and/or the water column. Experiments which have tried to establish the relative importance of sediments or water as sources of nutrients are inconclusive and further work is needed to establish how sediment nutrient characteristics vary within and among rivers (spatially and temporally) and the inter-relationships between sediment nutrients, water column chemistry and macrophytes. This paper presents the initial findings from a study of 17 lowland rivers in southern England which is exploring the spatial variability of sediment characteristics (total and inorganic phosphorus, total nitrogen, organic carbon, silt-clay fraction and organic matter content) and the relationship with aquatic macrophytes. The preliminary analysis indicates that although sediment characteristics are highly variable within 100-m river reaches, the variability across the 17 rivers is even greater; this is despite the limited geographic and trophic range of the study sites. The results presented in this paper also give some indication of the sediment characteristics associated with five macrophyte species but it is too early to ascribe sediment preferences for particular species. PMID- 11258807 TI - Controls on the nutrient content of suspended sediment transported by British rivers. AB - Recent studies of nutrient fluxes from river basins have emphasised the importance of sediment-associated transport. In order to develop an improved understanding of sediment-associated nutrient transport in UK rivers and of contrasts between individual rivers, attention has been directed to spatial and temporal variations in the nutrient (N and P) content of suspended sediment transported by four rivers, which embrace a representative range of British conditions. Bulk samples of suspended sediment were collected during storm events and these were analysed for both total N and P content and for individual fractions of these nutrients. Significant temporal and spatial variability in these various measures of the nutrient content of suspended sediment has been documented. Spatial variability has been linked to catchment characteristics, which in turn influence sediment sources. Patterns of temporal variability, which are reasonably consistent among the different rivers, have been related both to variations in other sediment properties and to hydrometeorological conditions. PMID- 11258808 TI - Downward fluxes of particulate carbon, nitrogen and phosphorus in the north western Adriatic sea. AB - Downward fluxes of particles, organic carbon, total nitrogen and total phosphorus and the composition of the settled particulate matter were determined in the north-western Adriatic Sea at two coastal sites influenced by the outflows of the Po and Adige rivers and one offshore site. Vertical fluxes were strongly influenced by resuspension processes in addition to the primary flux and advection. The resuspended material contributed on average 34-43% of the total matter sedimented in the near bottom traps in coastal waters. Net annual vertical fluxes (due to primary flux and advection) of organic carbon, total nitrogen and phosphorus in the coastal stations were: 71-97 g C m(-2) year(-1), 8-14 g N m(-2) year(-1) and 2.1-2.3 g P m(-2) year(-1), with the highest values recorded at the station off the Po river delta. The offshore site was characterised by net annual fluxes of particulates, C, N and P approximately one order of magnitude lower than the above. The carbon export to the bottom was limited in the warm seasons when it constituted only 2-9% of primary production, due to high recycling and utilisation in the upper layer of the water column, increasing up to 8-18% in winter because of the instability of the water column and low biological utilisation. PMID- 11258809 TI - Estimates of benthic fluxes of nutrients across the sediment-water interface (Guarapiranga reservoir, Sao Paulo, Brazil). AB - Concentration profiles of nutrients (dissolved organic carbon, nitrate, nitrite, ammonium and soluble reactive phosphorus) were determined in pore waters from sediment from the Guarapiranga reservoir (Sao Paulo, Brazil). Redox potential and acid volatile sulfide measurements on bulk sediment samples were determined in the field and laboratory, respectively. The sediment redox potential ranged from 170 to -220 mV at 0-1 cm and increased to somewhat higher values at 20 cm. The acid volatile sulfide (AVS) profile had a bimodal pattern with concentration peaks at 3 cm (27-55 mg kg(-1)) and 14 cm (70-110 mg kg(-1)). Dissolved organic carbon (DOC) concentrations increased from the surface (4.7-5.6 mg l(-1)) to 20 cm (values up to 12 mg l(-1)). The concentration of ammonium increased significantly with depth, with maximum concentrations occurring at 15 cm; nitrate nitrite concentrations only increased appreciably at 10 cm. The SRP profiles increased in concentration from the surface to approximately 10-cm depth, with a maximum value of 1200 microg H2PO4- l(-1). Benthic fluxes from the sediment into the pore water ranged from 278 to 339 mg cm(-2) year(-1) for ammonium ions and from 8 to 18 mg cm(-2) year(-1) for SRP. These upward diffusive fluxes correspond to 47-70% and to 10-24% of the total deposition of N and P measured in the reservoir, respectively. The burial rates for N and P in these sediments are 30 54% and 76-89%, respectively. PMID- 11258810 TI - Anoxic mineralization of biogenic debris in near-shore marine sediments (Gulf of Trieste, northern Adriatic). AB - Anoxic degradation of sedimentary biogenic debris using closed incubation experiments was studied at two sampling stations in the Gulf of Trieste (northern Adriatic). Production rates of dissolved inorganic carbon (DIC), NH4+, PO(4)3- and dissolved Si (dSi), and reduction rates of SO(4)2- were measured and anoxic mineralization rates were modeled using a first order G-model and multi-G approach. The depth profiles of these rates revealed an exponential decrease indicating that the largest fraction of mineralization of biogenic debris and SO(4)2- reduction occurs in the surficial sediment layer and on the sediment surface. Comparing the depth-integrated anoxic mineralization rates at both stations with benthic fluxes of DIC, NH4+, PO(4)3- and dSi measured at the in situ temperature in the dark, it appears that the DIC and PO(4)3- fluxes are higher because the mineralization mostly occurs at the sediment-water interface, and that besides SO(4)2- reduction, other electron acceptors are involved in the organic matter decomposition pathway in these surficial sediments. The NH4+ production was higher than the benthic fluxes because of NH4+ oxidation. The production of dSi was in good agreement with benthic fluxes implying that temperature is the main factor of dSi production and benthic fluxes in these sediments. PMID- 11258811 TI - Sediment dynamics in Rangoon river, Myanmar. AB - The behavior of fine-grained sediment in Rangoon River depends on seasonal variations in freshwater discharge and tidal amplitudes that range from 2 to 5 m. During the monsoon, freshwater with sediment concentrations of 1 g/l, or less, causes unidirectional, seaward flow. In the dry season, salinities reach 20% and sediment concentrations rise to 6 g/l. The saline intrusion advects large quantities of sediment landward from seaward sources. Near-surface sediment concentrations are very low during neap tides, while a layer of 'fluid mud' rests on the bottom. Current speeds greater than 0.2 m/s are needed to entrain sediment into the upper layer. Layered suspensions occur most commonly during decelerating phases of tidal flow and are dispersed by rapidly accelerating flow. When current speeds exceed 0.6 m/s, no 'fluid mud' forms, and sediment concentrations as great as 6-8 g/l extend through the water column. PMID- 11258813 TI - The use of an ultrafiltration technique for measurement of orthophosphate in shallow wetlands. AB - The overestimation of orthophosphate by filterable reactive phosphorus (FRP) measurement techniques has long been accepted. The aim of this study was to quantify that overestimation in 17 wetlands over time. Specifically an ultrafiltration technique was used prior to the application of the molybdenum blue phosphorus detection method to quantify orthophosphate concentrations. Samples were collected over a 6-month period and analysed for total filterable (< 0.50 microm) phosphorus (TFP), filterable reactive phosphorus (FRP < 0.50 microm) and ultrafiltered (< 10(3) Da) reactive phosphorus (PO4). FRP correlated well with PO4, however, FRP overestimated PO4 particularly with increasing colloidal phosphorus concentration. The ratio of DOC to TFP (C:P) influenced the fate of PO4 in the water, implying that DOC was forming complexes with phosphorus. The PO4 concentrations decreased with increasing C:P ratios in some of the wetlands over the 6-month monitoring period. PMID- 11258812 TI - A novel sediment gas sampler and a subsurface gas collector used for measurement of the ebullition of methane and carbon dioxide from a eutrophied lake. AB - A novel sediment bubble gas sampler and a subsurface bubble gas collector were designed to measure the ebullition of gases from profundal sediments of aquatic ecosystems. The sediment gas sampler was constructed to collect bubble gas samples directly from the uppermost sediment layers for gas composition analysis. The floating subsurface gas collector, designed to trap the bubbles released naturally from sediments, permitted the measurement of both the volume and the composition of the bubble gas. Due to its low cost, light weight and rapid sampling capability, the gas collector is ideal for studies requiring many replicate collectors. These devices were used for measurement of the ebullition of methane (CH4) and carbon dioxide (CO2) during an open water period from hypereutrophic Lake Postilampi, situated within the midboreal zone in Finland. The bubble gas obtained from the sediment with the sediment gas sampler had higher concentrations of CH4 and CO2 than the bubbles trapped in the gas collectors. This indicated that the bubble gas composition changed, either naturally during the migration of the bubbles from the sediment through the water column to the gas collectors, and/or during their storage in the collectors prior to sampling. The mean CH4 ebullition from Lake Postilampi was estimated to be in the range from 36 to 46 mg m(-2 d(-1), based on the bubble gas CH4 concentrations measured from the gas collectors and sediment, respectively. The bubbles contained only 0.02-0.57% of CO2 and thus, the ebullition had no significance in the release of CO2 from the lake. PMID- 11258814 TI - Diagenesis of magnetic minerals in the intertidal sediments of the Yangtze Estuary, China, and its environmental significance. AB - In this study, diagenesis of iron oxides in intertidal sediments of the Yangtze Estuary, China, has been investigated by combined environmental magnetic and geochemical methods. The results indicated that the magnetic properties of the sediments were dominated by ferrimagnetic magnetite. The content of Fe, DCB- and AOD-extractable iron oxides correlated positively with the concentration of fine grained magnetite near the superparamagnetic/stable single domain (SP/SSD, approximately 0.03 microm) boundary, and with anti-ferromagnetic minerals (hematite/goethite). The magnetic parameters for core SDK indicated a substantial decrease in magnetite concentration from a depth of approximately 20 cm toward the surface, together with a shift in the grain-size distribution of magnetic minerals toward the coarse end, suggesting selective dissolution of fine grained magnetite under reducing conditions. The reduction of iron oxides inferred from magnetic measurements was supported by the similar decrease in the concentration of Fe and Mn and a lower ratio of Mn/Fe. Magnetic measurements on another core from elsewhere also indicated substantial reductive dissolution of iron oxides. In conjunction with the results of heavy metal analysis, it was suggested that the dissolution of iron oxides had a direct effect on the cycling of heavy metals. Therefore, magnetic measurements may provide useful information as to early diagenesis within intertidal sediments, which greatly influences the behavior of heavy metals in coastal environments. PMID- 11258815 TI - Downstream changes in the transport and storage of sediment-associated contaminants (P, Cr and PCBs) in agricultural and industrialized drainage basins. AB - Samples of suspended, floodplain and channel bed sediment have been used to examine downstream changes in ediment-associated contaminant transport and storage in contrasting rivers in Yorkshire, UK. The concentrations of hosphorus, chromium and selected PCBs associated with sediment in the River Aire and its main tributary, the River Calder, which drain an urbanized and industrialized catchment, are considerably higher than those in the relatively unpolluted River Swale, which drains an agricultural catchment. Concentrations of sediment associated contaminants in the Aire/Calder system increase downstream, reflecting the location of urban and industrial areas in the middle and lower reaches, and the location of point source inputs, such as sewage treatment works. The ontaminant concentrations associated with floodplain and channel bed sediment in the Rivers Aire and Calder are high, particularly in the lower reaches. This, combined with measurements of sediment storage on the floodplain and channel bed, indicate that significant storage of sediment-associated contaminants occurs in the Rivers Aire and Calder. PMID- 11258816 TI - Floodplain lakes as sinks for sediment-associated contaminants--a new source of proxy hydrological data? AB - Lake and reservoir sediments often contain valuable records of sediment yield history and sediment-associated nutrient and contaminant transport spanning timescales from decades to millennia. Nevertheless, there have been few attempts to evaluate floodplain lakes as a source of proxy hydrological data for reconstructing short-term trends in sediment-associated nutrient and contaminant transport. Results from an analysis of floodplain lake sediment cores suggested good preservation of the 137Cs record, which provided an absolute dating control. Changes in the concentration of sediment-associated heavy metals and phosphorus were observed downcore and the analysis of mineral magnetic properties and particle size were used to identify the influx of sediment associated with high magnitude, low frequency flood events. Although floodplain lake-sediments only preserve a partial record, they may provide a valuable source of proxy hydrological data for reconstructing trends in sediment and sediment-associated contaminant transport where long-term sediment monitoring programmes are not available. PMID- 11258817 TI - Contaminants in sediments: remobilisation and demobilisation. AB - In this study, the contaminated anoxic sediment of the Mulde reservoir (Saxony, Germany) was investigated. Several sediment cores were analysed for heavy metals and organic chemicals such as chlorobenzenes and DDTs. The comparison between anoxic and oxidised sediment cores showed the potential danger for heavy metal (Zn and Cd) remobilisation from sediment due to bioturbation or resuspension by flooding. Chemical sequential extraction was used to describe partitioning of heavy metals among different mineralogical components in the sediments. Results showed remobilisation of Zn and Cd from the sediments. The stable fraction (organic/sulfidic-bound) of Zn and Cd decreased from 10 to 3%, and from 35 to 5%, respectively. Simultaneously, the carbonate fraction increased from 3 to 12% for Cd and from 10 to 22% for Zn. Furthermore, the simulation of the diffusion of organic pollutants showed remobilisation of 1,4-dichlorobenzene. The results confirmed the necessity of sediment remediation in the reservoir. Capping seems to be a promising approach for a low-cost remediation. PMID- 11258818 TI - Sulfate reduction in Lake Agmon, Israel. AB - Lake Agmon, a newly reflooded water body in the southern part of the Hula valley is characterized by a clear phase period in winter with almost no blooms of phytoplankton. Dense macrobenthos and algal blooms form in the lake during summer and autumn. High primary production and chlorophyll-a concentrations were measured in April and during the summer of 1997. Fresh organic matter in the sediments from the degradation and decomposition of the blooms together with high sulfate concentrations, allowed microbial sulfate reduction to occur. Sediment cores taken from different sites (peat and marl) during various seasons in 1997, showed high sulfate reduction rates in June in the marl region, and in September in the peat region (842 and 2834 nmol SO(4)2- reduced ml(-1) day(-1), respectively). In February, corresponding to the development and decline of macrophytes and algal blooms, lower rates of sulfate reduction were recorded (11 nmol SO(4)2- reduced ml(-1) day(-1)). Sulfate reduction is limited by the supply of organic matter and is one of the major processes contributing to the mineralization of organic matter in this lake. PMID- 11258819 TI - Comparison of iron accumulation in lakes using sediment core and mass balance calculations. AB - The accumulation of iron (Fe) in several lakes in Ontario, Canada was determined by two independent approaches. First, Fe accumulation was calculated in cores collected from several sites in each lake by integrating Fe concentration profiles with sediment accumulation rates determined from Pb210 dating. These site-specific accumulation rates were corrected for sediment focussing so that whole-lake Fe accumulation values could be derived. Using this approach, recent whole-lake Fe accumulation in eight lakes ranged between approximately 750 and 4000 mg/m2 per year. Second, whole-lake Fe accumulation was estimated from lake mass budgets, which were measured over a maximum of 14 years. Accumulation measured using the mass balances ranged from 10 to 1330 mg/m2 per year. Comparison of the two approaches indicated that retentions calculated from the sediment cores were much greater than those estimated from the mass balances. The most likely explanation for this difference is that, in the two decades since the cores were collected, there has been a substantial decline in Fe retention (in mass units but not percent) in the study lakes, principally as a result of reduced inputs of Fe from the catchments. PMID- 11258820 TI - The degradation of xenobiotic branched carboxylic acids in anaerobic sediment of the Pearl River in Southern China. AB - Xenobiotic branched carboxylic acids (BCAs) discharged by industries are often persistent in biological wastewater treatment systems and end up in water and sediments. In this study, the degradation of 12 typical BCAs in an anaerobic environment of river sediment was studied in vitro using enrichment shake-flask cultures. The anaerobic consortium taken from the river sediment, comprising BCA degrading and methane-producing genera, degraded BCAs with tertiary carbons through beta-oxidation, followed by methanogenesis mechanisms. The maximum cell densities in the cultures using BCAs as the sole carbon source ranged between 5.0 and 6.0 x 10(5) cells/ml. The maximum degradation rates were between 5.0 and 8.5 x 10(-3) mmol/h. The consortium could not degrade BCAs with quaternary carbons. The degree of branching at the alpha or beta position along the carbon chain interfered with the beta-oxidation mechanisms. These BCAs would accumulate in the sediment and significantly affect the cycling of organic carbon and nutrients. PMID- 11258821 TI - Retention of conservative and sorptive solutes in streams--simultaneous tracer experiments. AB - The effective transport velocity of solutes in rivers and streams is governed by transient storage in hyporheic zones in which the longitudinal advection velocity is small relative to the main stream flow velocity. Results from a simultaneous tracer experiment using a non-reactive (tritium, 3H2O) and a sorptive tracer [chromium, 51Cr(III)] have formed the basis of a more accurate interpretation of the retention characteristics of solutes in streams than previously has been possible. By using a simultaneous injection of these two tracers, it was possible to distinguish between their different behaviours. Based on estimations of fluxes, the retained mass of chromium in the storage zones along the 30-km-long study-reach was 76% after 150 h. Independent observations in the bed sediment indicated that the loss of chromium observed in the water was mainly a result of uptake into the bed sediment. To describe the transport in the stream, a model concept including solute sorption kinetics in the bed sediment was proposed. Evaluation of parameters in the model, indicated that the uptake of chromium in the bed sediment is controlled by sorption kinetics. PMID- 11258822 TI - Impact of frontal systems on estuarine sediment and pollutant dynamics. AB - In this paper, a brief description of frontal systems, their modes of occurrence and impact on the estuarine environment, is presented. Previous studies of estuarine fronts have largely focused on the water surface and within the water column. New observations in the Tay Estuary, Scotland have shown that the presence of fronts within the water column may be marked, not only by surface foam bands, but also by abrupt (i.e. non-gradational) changes in the underlying bedform morphology and/or sediment facies, as detected using side-scan sonar. This preliminary evidence suggests that fronts may exert a control, not only on the surface and intra-water column sediment and pollutant partitioning, but also on the distribution and persistence of bedload transport pathways. PMID- 11258823 TI - Effect of sorption kinetics on the transport of solutes in streams. AB - To provide an appropriate description of the transport of a reactive substance in a stream, it is important to include a kinetic description of sorption in a transport model. In this study, first-order sorption kinetics was taken into account in both the transient storage zone and the stream water, and analytical expressions for relative error in statistical moments of the residence time PDF, resulting from disregarding sorption kinetics, were derived. The sorption rate coefficient in the water was found to influence the error in the expected value, and the error was found to approach infinity as the travel distance or sorption rate coefficient approaches zero. The sorption rate coefficient in the storage zone influences only higher-order moments. For sufficiently long distances, the error in the variance was found to be more pronounced when sorption kinetics in the storage zone was disregarded, than when sorption kinetics in the stream water was disregarded. Parameter values from a tracer experiment with 51Cr revealed that the relative error in the variance could be more than 100%, if sorption kinetics in the storage zone is disregarded. PMID- 11258824 TI - Erosion investigation and sediment quality measurements for a comprehensive risk assessment of contaminated aquatic sediments. AB - In this paper, an assessment strategy is introduced which allows one to evaluate the ecological hazard of contaminated sediments in connection with the risk of in stream erosion. Special techniques for sediment sampling, non-intrusive density profiling, and depth related measurement of erosion are presented, which, in combination with ecological aspects, lead to a comprehensive risk assessment of fluvial sediments. The strategy was applied to a lock-regulated reach of the River Neckar in Germany. The spatial pattern of contamination in the river reservoir was found to be remarkably heterogeneous. At some sites, very high heavy metal concentrations were detected at the sediment surface. A sudden increase in contamination with depth at other sites could be attributed to an erosional unconformity. The critical shear stress of erosion for old contaminated sediments is higher than for recently deposited material. Nevertheless, during major flood events, bottom shear stress in the river exceeds the critical shear stresses of erosion of all sediments. Accordingly, there is a substantial risk that old contaminated sediment can be mobilised from the reservoir and transported downstream. PMID- 11258826 TI - Detoxification of tributyltin contaminated sediments by an electrochemical process. AB - Preliminary experiments have shown that dibutyl and tributyltin can be decomposed by the electrochemical treatment of sediment. Two different process pathways have been described and compared. A slurry electrolysis of the suspended sediment seemed to be more efficient than column leaching followed by electrolysis. Tributyltin was destroyed under oxidising as well as under reducing process conditions. The detoxification mechanism seemed to be stepwise removal of the butyl groups. A partial debutylation of tri- and dibutyltin could be achieved, although monobutyltin was not affected. This technique is promising, but further investigation is necessary to improve the experimental conditions and to characterise the real potential of these process pathways. PMID- 11258825 TI - Radium migration through clay liners at waste disposal sites. AB - The migration of 226Ra through the bottom compacted clay liner of the wastewater disposal reservoirs of an industrial plant that processes uranium ore was evaluated. An instrumental method for 226Ra analysis in soils, consisting of detector calibration, the determination of detector counting efficiency, cumulative counting of both background and soil samples in regular counting intervals, and photo-peak smoothing was developed. The 226Ra was analyzed by means of its granddaughter 214Bi, at a photo-peak of 609 keV. The results showed that most of the 226Ra which diffused from the solution into the soil was retained in the upper layer of the sample, and that just a small percentage migrated to the subjacent layers. This methodology is adequate for the assessment of the migration of radionuclides through soil layers and for environmental impact studies related to contamination of soils by radionuclides. PMID- 11258827 TI - The impact of a hydroelectric power plant on the sediment load in downstream water bodies, Svartisen, northern Norway. AB - When the Svartisen hydroelectric power plant was put into operation, extensive sediment pollution was observed in the downstream fjord area. This paper discusses the impact of the power plant and the contribution from various sources of sediment. Computation of the sediment load was based on samples collected one to four times per day. Grain size distribution analyses of suspended sediments were carried out and used as input in a routing model to study the movement of sediments through the system. Suspended sediment delivered to the fjord before the power station was constructed was measured as 8360 metric tons as an annual mean for a 12-year period. During the years 1995-1996 when the power plant was operating, the total suspended load through the power station was measured as 32609 and 30254 metric tons, respectively. Grain size distribution analyses indicate a major change in the composition of the sediments from 9% clay before the power plant was operative to 50-60% clay afterwards. This change, together with the increase in sediment load, is believed to be one of the main causes of the drastic reduction in secchi depths in the fjord. The effect of the suspended sediment load on the fjord water turbidity was evaluated by co-plotting secchi depth and power station water discharge. Measurements during 1995 and 1996 showed that at the innermost of these locations the water failed to attain the minimum requirement of 2 m secchi depth. In later years secchi depths were above the specified level. In 1997 and 1998 the conditions improved. At the more distal locality, the conditions were acceptable with only a few exceptions. A routing model was applied to data acquired at a location 2 km from the power station in order to calculate the contributions from various sediment sources. This model indicated that the contribution from reservoir bed erosion dominated in 1994 but decreased significantly in 1995. Future operation of the power station will mostly take place with a high water level in the reservoir and is likely to result in acceptable water quality in the fjord. However, during periods of low drawdown, sediment pollution may again become a problem. PMID- 11258828 TI - Sedimentological research as a basis for environmental management: the Oresund fixed link. AB - A fixed 16 km link (immersed tunnel, artificial island and peninsula, bridges) to connect the Scandinavian countries of Denmark and Sweden is being constructed during the period 1995-2000. The link crosses Oresund, one of the three sounds connecting the brackish Baltic Sea to the oceanic North Sea. Dredging of approximately 8 x 10(6) m3 of limestone bedrock and clay till is being carried out during the construction phase. Environmental planning and management when constructing the fixed link were heavily dependent on extensive sedimentological and biological research prior to construction. Base-line turbidity and seabed sediments were investigated, and sediment spill parameters were measured during test dredging and test reclamation, in addition to the environmental impact. Now, when dredging and reclamation have been finalised in Oresund, it is clear that the environmental management approach used during construction of the fixed link has safeguarded the marine environment in Oresund. At the same time it has allowed the dredging and reclamation works to be carried out within the planned time period and expenses. A detailed base-line monitoring of the regional turbidity regime and the regional sedimentation regime in Oresund made post dredging evaluation of the regional effects possible. The measurements in Oresund showed that the construction caused no permanent regional change in turbidity and sedimentation, as the base-line conditions did not change. PMID- 11258829 TI - Modeling the ecological impact of heavy metals on aquatic ecosystems: a framework for the development of an ecological model. AB - In this paper, an ecological model is proposed to predict the effects of heavy metals on aquatic ecosystems. The bioavailable concentration of metals and a concept of toxicity strength (TS) are combined. The integrated ecological model relates the transport, distribution and speciation of heavy metals and their toxicity, and the effect of environmental variability on metal toxicity. It also emphasizes the link between physical and chemical processes of heavy metals in rivers and ecological effects. Based on the data obtained from research in the CERP project (Co-operative Ecological Research Project), the ecological impact of heavy metals on the aquatic ecosystem of the Le An River (polluted by heavy metals from a copper mine) was predicted. The results show that the estimated values of toxicity strength for surface water are in agreement with the percentage inhibition for the test organism (P. phosphoreum) and that the predicted ecological effect of polluted sediment is consistent with natural variability in aquatic ecosystems. PMID- 11258830 TI - A preliminary model for predicting heavy metal contaminant loading from an urban catchment. AB - The toxicity of heavy metals to biota in urban catchments has been regarded as a very important non-point source pollution issue. Numerous studies on heavy metal pollution in urban receiving waters have found that metal transport by surface runoff is closely correlated to the partitioning of the metal forms between dissolved and particulate phases, where sediment plays an important role in the transport process. Sediment cycling on urban streets, metal binding form, and rainfall character in the catchment area are considered to be the key factors for metal transport. A preliminary model is developed based on these considerations. Starting from classical build-up and wash-off processes for the suspended sediment (SS) on the urban impervious surface, the model links the transport of suspended sediment to the transport of metal species. Monitoring data from a small highway catchment were used in the model development. A total of 47 rain events over 1 year were monitored intensively at short time intervals (5-10 min) for hydrological data, rainfall intensity, and stormwater quality. In developing the model, lead was used for the metal load prediction, as it has been a common fuel additive for urban transportation. Agreement between model results and monitoring data indicates that the model can be used in predicting metal load from impervious urban areas, such as streets and roadways, on a long-term basis. PMID- 11258831 TI - Mapping and monitoring contaminated-sediment geometry and stability. AB - Environment Canada's National Water Research Institute (NWRI) conducts research on freshwater contaminated sediments, much of which is focused on designated areas of concern in the Great Lakes and their connecting channels. This paper reviews new acoustic and video equipment and procedures developed to map the geometry and stability of the sediments, and describes their applications. A RoxAnn acoustic seabed-classification system is used for mapping bottom-sediment types and locating the deposits of fine-grained sediments with which contaminants are associated. The system uses the acoustic properties of sediments to distinguish textural types ranging from mud to boulders, and displays the data as they are collected. The sediment thickness is measured with a weighted video acoustic tripod which is lowered into the sediments to refusal, and which recorded penetration with a video camera or an echosounder transducer. The stability of the contaminated sediments was monitored with a bottom-mounted, high precision echo sounder-digitizer, which logs changes in the position of the sediment-water interface produced by erosion or deposition. The same procedure can be used in capping or dredging projects to track bottom changes as they occur, or they can be measured by pre- and post-project mapping of bathymetry and morphology with sweep-sonar or side-scan sonar equipment. The new equipment and procedures have been successfully applied to a number of areas of concern in the Great Lakes basin. They provide a faster and more detailed characterization of sediment properties and geometry than was previously available, and have been particularly effective in optimizing sampling surveys and monitoring remediation projects. PMID- 11258832 TI - Effects of sediment bulk properties on erosion rates. AB - Considerable work has been done recently on the effects of sediment bulk properties on erosion rates. From this it is known that erosion rates depend on at least the following parameters: bulk density, average particle size, particle size distribution, mineralogy, organic content, volume of gas in the sediment, salinity of the pore waters, and time after deposition. This work is reviewed and discussed here with the purpose of presenting a quantitative overview of the effects of each of these parameters. This information is then used to demonstrate a procedure for estimating erosion rates of sediments based on a knowledge of their bulk properties. PMID- 11258833 TI - Parameter estimation for suspended sediment transport processes under random waves. AB - This paper presents a parameter estimation method for suspended sediment transport processes subject to random wave environments. An objective function was constructed based on measurements of suspended sediment concentration profiles and the governing equation of sediment transport. The Chebyshev least square method was employed to approximate the process parameters, i.e. vertical eddy diffusivity (epsilon) and net vertical velocity (w). The objective function of sediment transport processes with Chebyshev's parameters is well posed and does not require boundary conditions. First, second, and third order epsilon and w Chebyshev orthogonal functions were determined for monochromatic (MONO), narrow banded (NBR) and broad-banded (BBR) random wave conditions. In the BBR and NBR conditions, the best fit Chebyshev approximations of epsilon and w were 2nd degree, while the best approximations in the MONO condition were 2nd degree for epsilon and 1st degree for w. The Chebyshev method provides a quick, accurate and direct estimation of two prime parameters in sediment transport dynamics. PMID- 11258834 TI - Modelling sedimentation processes in a constructed stormwater wetland. AB - The design and operation of constructed wetlands for the treatment of stormwater relies heavily on promoting sedimentation, and being able to predict accurately the expected effectiveness of the pond in removing material from the inflows. A study of sediment behaviour has been carried out in a stormwater wetland in Adelaide, Australia where computer predictions, based on solving the hydrodynamic equations and the transport equation, have been compared to deposition patterns observed in the field. The long-term residence time distribution has been shown to be useful in predicting overall sediment removal rates. Comparisons between the model and field observations indicate generally good agreement. Sources of potential error identified include the variable nature of the runoff concentrations entering the pond and the sediment size distribution. The importance of the transient nature of the flow events was highlighted by the spread of sediment throughout the whole pond. PMID- 11258836 TI - The effect of depositional history on contaminated bed sediment stability. AB - Experiments were conducted in an annular flume using a commercially available kaolinite clay as well as contaminated bed sediment from Hamilton Harbour (Ontario) to assess their stability against erosion. Critical shear stress for erosion was measured under different conditions of bed formation (quiescently deposited beds and shear deposited beds) as well as with and without the presence of a biostabilized bed. Results suggest that a biostabilized bed and a bed formed under a flowing condition, similar to a river scenario, will be more resistant against erosion than will a non-biostabilized bed and a bed formed under quiescent conditions. Up to three cycles of erosion and flocculation/deposition were observed to occur within one experiment. These results suggest that the depositional history and biostabilization of river bed sediments need to be seriously considered within sediment and contaminant transport models if meaningful estimates of sediment and contaminant source, fate and effect are to be generated and used for the management of our aquatic ecosystems. PMID- 11258835 TI - The pattern of particle flux variability in Swedish and Swiss lakes. AB - Particle settling flux in the upper and lower water column, measured with cylindrical sediment traps in 11 Swedish and nine Swiss lakes showed high temporal and spatial variability within and among lakes, ranging from 0.1 to 385 g m(-2) day(-1). Such high variability reflects the large differences in lake morphology and trophic state. However, despite these differences, the particle flux variability followed a consistent pattern resulting in a significant relationship between minimum, mean and maximum particle flux. The mean particle flux is significantly related to the dynamic ratio (square root of area divided by mean depth) of the lake, provided that the lake is shallow (mean lake depth < or = 9 m), its dynamic ratio is > or = 0.15 and allochthonous particulate matter input is < or = 10%. In such lakes, typically found in Sweden, the described relationships may be used to predict the variability of particle settling flux without developing complicated models. PMID- 11258837 TI - Phosphorus characteristics of settling and suspended particles in Lake Erken. AB - The aim of the study was to determine proportions of phosphorus forms as well as total phosphorus content in suspended and settling particles during spring, summer and autumn in order to improve the understanding of particle composition and mineralization processes in moderately eutrophic, temperate lakes with summer stratification. The highest phosphorus content was found in epilimnetic suspended matter with maximum mean concentration in summer. There was a gradual decrease in total P content from suspended matter to settling particles and surficial sediment. Phosphorus extracted with sodium hydroxide was the dominant fraction showing organic phosphorus to be most important. Labile phosphorus was a significant fraction and contributed more in epilimnetic suspended matter and in material from the uppermost traps in comparison to hypolimnetic material and surficial sediment. There was a seasonal variation in P forms with maximal shares of labile P and organic P in summer. The findings clearly indicate a dominance of phytoplankton and detritus in the epilimnetic particulate matter during summer, while inorganic and resuspended material is more important during mixing periods. The P composition of settling particles and suspended matter show similar trends, but the total content and shares of labile and organic P are significantly higher in the latter. PMID- 11258838 TI - Evaluation of iron-phosphate as a source of internal lake phosphorus loadings. AB - Biological, physical and chemical characteristics of the water column of a shallow (Zmax = 9.2 m), small (surface area 3.8 km2) residential and recreational lake near Prince George, British Columbia, indicated that the system was being loaded internally with phosphorus (P) from the sediments. The abundance of P released from the fine glaciolacustrine, and organic rich sediments was resulting in excess algal and weed growth. It was postulated that iron-phosphate reduction at redox potentials below approximately 200 mV and/or bacterially mediated orthophosphate (PO4-P) releases could be occurring. The development of an appropriate nutrient management strategy required that the process associated with the sediment P release be determined. The MINTEQA2 geochemical model was used to predict the release of orthophosphate (PO4-P) into the interstitial water with the assumption that P is present alternately as strengite, variscite and hydroxyapatite. The predicted release of PO4-P from these P containing minerals was compared to the concentration of PO4-P and total phosphorus (TP) in the overlying hypolimnion. In order to improve the accuracy of the model prediction, the proportion of the sediment present as iron-bound phosphate was estimated. A significant correlation between the observed hypolimnetic TP and interstitial PO4 P concentrations as predicted from iron-bound P dissolution (r2 = 0.59) was found. Total phosphorus release rates to the hypolimnion were also found to be strongly correlated to the iron-bound P component of the sediment (r2 = 0.88). Multivariate regression analyses showed significant relationships between hypolimnetic PO4-P and sediment iron-bound P, Eh, and interstitial Fe (r2 = 0.76). These results provided sufficient evidence to conclude that PO4-P in the system is predominantly bound to Fe-containing minerals and therefore could be managed using treatment techniques that address iron-bound phosphates. PMID- 11258839 TI - Particulate phosphorus transport by sub-surface drainage from agricultural land in the UK. Environmental significance at the catchment and national scale. AB - Phosphorus (P) is the key limiting nutrient in most UK freshwater systems. With increased legislation controlling point source inputs, dissolved (DP) and particulate P (PP) derived from diffuse sources are making a more significant contribution to the total P loading of surface waters. Recent research has focused on pathways linking diffuse sources to the fluvial system and sub-surface field drains have been shown to transport both sediment and P rapidly to watercourses. Preliminary results are presented from an ongoing study using environmental tracers to identify the source of the drain sediment and its potential as a carrier of PP. These results suggest that the majority of sediment in drains is topsoil derived, but the significance of P loss via this pathway in a regional or UK context has yet to be evaluated. A protocol to study the potential problem at a regional/national scale is discussed and initial data presented. PMID- 11258840 TI - Collaborative exercise on the use of FISH chromosome painting for retrospective biodosimetry of Mayak nuclear-industrial personnel. AB - PURPOSE: To investigate within the framework of a multilaboratory study the suitability of FISH chromosome painting to measure so-called stable translocations in peripheral lymphocytes of Mayak nuclear-industrial workers (from the Southern Urals) and their use for retrospective biodosimetry. MATERIALS AND METHODS: Chromosime analyses were carried out from 69 workers who had received protracted occupational radiation exposures (0.012-6.065 Gy) up to approximately 40 years before blood sampling. Twenty-one unexposed people living in the same area were controls. A multicolour FISH-painting protocol with the target chromosomes 1, 4 and 8 simultaneously with a pancentromeric probe was used to score potentially transmissible chromosome-type aberrations (reciprocal translocations 2B and related 'one-way' patterns I-III according to the S&S classification). RESULTS: Individual biodosimetry estimates were obtained in terms of these potentially long-term surviving aberration types based on the linear component of a low dose-rate gamma-ray calibration curve produced using identical staining and scoring protocols. For comparison, the workers personal and total background doses were converted to red bone marrow doses. The estimated doses were mainly lower than would be predicted by the calibration curve, particularly at accumulated higher dose levels. CONCLUSIONS: Owing to the limited life-time of circulating T-lymphocytes, the long-term persistence of translocations in vivo requires the assumption of a clonal repopulation of these naturally senescing cells from the haemopoietic stem cell compartments. Obviously such a replacement cannot be fully achieved, leading to a temporal decline even of the yield of transmissible aberrations types. Assuming further a highly selective capacity of stem cells against any type of chromosomal damage and the fact that one must rely on partial genome findings, the potential of FISH chromosome painting for retrospective dose reconstruction is probably limited to a decade or so after high-level protracted radiation exposure. PMID- 11258841 TI - Evaluation of ionizing radiation sensitivity markers in a panel of lymphoid cell lines. AB - PURPOSE: To examine the possible associations between radiation sensitivity to doses 2 Gy, and such features of lymphoid cell responses as apoptosis, expression of apoptosis regulatory proteins (Bcl-2 family) and cell cycle progression in relation to biological dosimetry. MATERIALS AND METHODS: The cell lines examined were: Epstein Barr virus transformed lymphoid ataxia-telangiectasia (AT) cell lines, GM00717C, homozygous, and GM00736A, heterozygous, for ATM; human pro-B lymphoblastic leukaemia, Reh; murine L5178Y lymphoma sublines, LY-R and LY-S. Assays performed following X-irradiation with doses from 0.1 to 2 Gy were: terminal deoxyribonucleotidyl transferase (TdT) assay to measure apoptotic fraction, DNA content analysis by flow cytometry to assess cell cycle distribution, trypan blue exclusion test to determine cell viability, cytochalasin block micronucleus assay to assess cytogenetic damage, and Western blotting to detect proteins from the Bcl-2 family. RESULTS: The cell lines in the study were of different but rather high radiation sensitivity, which was unrelated to their propensity to undergo apoptosis or micronucleus frequency. The expression of apoptotic regulatory proteins from the Bcl-2 family (constitutive and expressed 4 or 24 h after irradiation) was not related to radiation sensitivity. CONCLUSION: None of the simple predictive tests used in the study, alone or evaluated together was suitable for detection of radiation hypersensitivity although cells known to be hypersensitive (LY-S and GM00717C) were included in the analysis. PMID- 11258843 TI - Relationships between clonogenic cell survival, DNA damage and chromosomal radiosensitivity in nine human cervix carcinoma cell lines. AB - PURPOSE: To compare clonogenic cell survival, DNA damage and chromosomal radiosensitivity in nine cervix carcinoma cell lines. MATERIALS AND METHODS: Initial and residual (after 24h repair) radiation-induced DNA damage was evaluated using pulsed field gel electrophoresis. Chromosome damage was measured by micronucleus (MN) induction in cytochalasin-B-induced binucleate cells. RESULTS: Significant differences between the cell lines were obtained in the induced levels of initial damage, residual damage and MN. Values for SF2 for the nine cell lines ranged from 0.36 to 0.92. No correlation was found between clonogenic measurements of radiosensitivity and initial DNA damage dose response slopes. However, borderline significant correlations were seen between clonogenic radiosensitivity data and the levels of residual DNA damage. There was no correlation between clonogenic radiosensitivity and the levels of radiation induced MN. Cell lines with high levels of initial damage had high yields of MN induced by radiation and the correlation seen was significant. CONCLUSIONS: The poor correlation between the different endpoints precludes their use in a clinical setting on primary tumour samples in vitro. It may be that tumour cell lines in vitro are a poor model for tumours in vivo. Studies aimed at assessing assays for measuring tumour radiosensitivity therefore should employ clinical samples. In vitro cell line work should concentrate on unravelling the complex mechanisms involved in determining a radiosensitive or radioresistant phenotype. PMID- 11258842 TI - Redox reactivity of guanyl radicals in plasmid DNA. AB - PURPOSE: It has been previously argued that gamma-irradiation of plasmid DNA in the presence of thiocyanate ions produces products recognized by the E. coli base excision-repair endonuclease formamidopyrimidine-DNA N-glycosylase (FPG), and there that derive from an intermediate guanyl radical species. The wish was to characterize the reactivity of this intermediate with reducing agents. MATERIALS AND METHODS: Aqueous solutions of plasmid DNA containing either bromide or thiocyanate (10(-3) to 10(-1) mol dm(-3)) and also one of six other additives (azide, ferrocyanide, iodide, nitrite, promethazine, tryptophan, 10(-7) to 10(-3) mol dm(-3)) were subjected to 137Cs gamma-irradiation (662 keV). After irradiation, the plasmid was incubated with FPG. Strand break yields before and after incubation were determined by agarose gel electrophoresis under neutral conditions. RESULTS: The very high yields of FPG-sensitive sites in the presence of SCN- or Br- decreased significantly with increasing concentrations of all of the six additives, with promethazine and tryptophan being the most efficient additives, and azide and iodide the least. CONCLUSIONS: From the results it is possible to estimate values of the rate constants for the reduction of the DNA guanyl radical (5 x 10(5), 2 x 10(5), 10(7) and 10(7) dm3 mol(-1) s(-1) for ferrocyanide, nitrite, promethazine and tryptophan respectively). PMID- 11258844 TI - Overexpression of the DNA-binding domain of poly(ADP-ribose) polymerase inhibits rejoining of ionizing radiation-induced DNA double-strand breaks. AB - PURPOSE: To assess the influence of trans-dominant inhibition of poly(ADP ribosyl)ation on the rejoining kinetics of radiation-induced DNA double-strand breaks (DSB). MATERIALS AND METHODS: Stable transfectants of the SV40-transformed hamster cell line CO60 were used: COM3 cells contain a construct to overexpress the poly(ADP-ribose) polymerase (PARP-1) DNA-binding domain (DBD) when induced by dexamethasone, as well as a construct for the constitutive overexpression of the human glucocorticoid receptor (Hg0). COR3 are control cells containing only the Hg0 plasmid. DSB induction and rejoining in X-irradiated cells was assessed by DNA pulsed-field electrophoresis. RESULTS: DSB induction was identical in both cell lines and independent of the presence of dexamethasone. DSB rejoining kinetics was independent of dexamethasone in COR3 cells and identical to COM3 cells without dexamethasone. However, in COM3 cells treated with dexamethasone to induce PARP-1 DBD overexpression, the fast component of the rejoining kinetic was largely reduced, and residual fragmentation increased concomitant with the increased damage fraction in slow rejoining. CONCLUSIONS: The results indicate that inhibition of cellular PARP-1 does not affect the rate-limiting step of either fast or slow DSB rejoining. Rather, it appears that absence of poly(ADP ribosyl)ation due to dominant negative PARP-1 expression induces a shift from rapid to slow DSB rejoining and by this mechanism PARP inhibition may increase the risk of repair failures. PMID- 11258845 TI - High-LET radiation induces apoptosis in lymphoblastoid cell lines derived from atazia-telangiectasia patients. AB - PURPOSE: To investigate and compare the propensity of Epstein-Barr virus (EBV) transformed lymphoblastoid cell lines (LCL), derived from ataxia-telangiectasia (A-T) patients and from unaffected healthy individuals (controls), to undergo apoptosis after exposure to high-linear energy transfer (LET) radiation. MATERIALS AND METHODS: Four A-T (ARO, BMA, CSA and RJO) and two control (JAC and KKB3) LCL were exposed to doses of up to 4Gy of accelerated nitrogen ions (32-45 MeV/u, 8-12Gy/min). For comparative purposes X-ray irradiation (1.36 Gy/min) was also performed. The induction of apoptosis was studied 0-48 h after irradiation with the use of two methods: (1) monitoring of high molecular weight (HMW) DNA fragments by field inversion pulse gel electrophoresis (FIGE); and (2) morphological characterization ofapoptotic cells after fluorescent staining. In parallel, cell-cycle distribution, monitored by DNA flow cytometry, as well as measurements of p53/p21(WAF1) protein levels by Western blots, were investigated in these cells. RESULTS: High-LET radiation-induced apoptosis and G2/M-arrest in both A-T and control LCL. No significant increase in the amount of p53/p21(WAF1) proteins preceded apoptosis in control or in A-T LCL after high-LET irradiation. However, low-LET radiation did induce significant enhanced levels of p53 proteins in control but not in A-T LCL. CONCLUSIONS: LCL from both A-T homozygous and unaffected healthy individuals undergo apoptosis without accumulation of p53/p21(WAF1) proteins after exposure to high-LET radiation. In contrast, low-LET radiation induces apoptosis and significantly increases levels of p53 protein in control but not in A-T LCL. PMID- 11258846 TI - Cell cycle progression and radiation survival following prolonged hypoxia and re oxygenation. AB - PURPOSE: To investigate cell cycle progression and radiation survival following prolonged hypoxia and re-oxygenation. MATERIALS AND METHODS: NHIK 3025 human cervical carcinoma cells were exposed to extremely hypoxic conditions (<4ppm O2) for 20 h and then re-oxygenated. The subsequent cell cycle progression was monitored by analysing cell cycle distribution at different time-points after re oxygenation using two-dimensional flowcytometry. The clonogenic survival after a 3.6 Gy X-ray dose was also measured at each of these time-points. The measured radiation survival was compared with theoretical predictions based on cell cycle distribution and the radiation age response of the cells. RESULTS: Following re oxygenation the cells resumed cell cycle progression, completed S-phase, and then accumulated in G2. Non-clonogenic cells remained permanently arrested in G2, while the remainder of the cells completed mitosis after a few hours delay. The radiation survival of the hypoxia-pretreated cell population remained lower than for an exponentially growing control population for the investigated 50h of re oxygenation. However, following 7 h of re-oxygenation, the radiation survival of the hypoxia-treated cell population correlated well with theoretically predicted values based on cell cycle distribution and radiation age response. CONCLUSIONS: The work demonstrates that prolonged hypoxia followed by re-oxygenation results in a G2 delay similar to that observed after DNA damage. Furthermore, chronic hypoxia results in decreased radiation survival for at least 50h following the reintroduction of oxygen. The hypoxia-induced radiosensitization following 7 h of re-oxygenation could in large part be explained by the synchronous cell cycle progression that occurred. PMID- 11258847 TI - A higher micronucleus yield in B-versus T-cells after low-dose gamma-irradiation is not linked with defective Ku86 protein. AB - PURPOSE: To elaborate the B-cell micronucleus (MN) response in the low-dose region in detail and to investigate the postulated deficiency in DNA-PK in B cells. MATERIALS AND METHODS: Lymphocytes of five healthy volunteers were irradiated with low LET gamma-rays and high LET fast neutrons with doses ranging between 0.01 and 2 Gy. After post-irradiation incubation, B- and T-cells were isolated via CD3 and CD19 immunomagnetic microbeads. MN were analysed in both subpopulations. To study the underlying mechanism of chromosomal radiosensitivity, cell extracts prepared from purified B- and T-cells were subjected to SDS-electrophoresis and electroblotting using antibodies directed against the DNA-PK repair enzymes Ku70/86 and DNA-PKcs. Activity measurements were performed using the SignaTECT DNA-dependent protein kinase assay. DNA double strand break (DSB) induction and rejoining was determined using constant-field gel electrophoresis. RESULTS: For low LET gamma-rays a higher MN yield was observed in B-cells than in T-cells, but only in those samples exposed to doses < 1 Gy. For 1 Gy, the MN yields were comparable and for 2Gy even lower in B-cells compared with T-cells. After high LET neutron irradiation no significant differences in MN yields were observed between both subsets. The results of the DNA-PK experiments demonstrate that there is no difference between T- and B-cells in the basal expression and activity of DNA-PK repair proteins. No differences in DNA DSB induction and rejoining were found between T- and B-cells using constant field gel electrophoresis. CONCLUSIONS: From the results, it was concluded that the enhanced chromosomal radiosensitivity in B-cells is restricted to low doses (<1 Gy) of low LET radiation and that the chromosomal behaviour of B-cells to low LET radiation cannot be attributed to aberrant forms of the DNA-PK components. A type of chromosomal induced radioresistance (IRR) may be a possible explanation for the observed effect. PMID- 11258848 TI - Modification of oral mucositis by keratinocyte growth factor: single radiation exposure. AB - PURPOSE: The aim was to quantify the effect of recombinant human keratinocyte growth factor (rhKGF) on acute oral mucositis induced by a single radiation dose, simulating accidental radiation exposure. MATERIAL AND METHODS: Tongue epithelium of the C3H/Neu mouse was irradiated with graded single doses of 25 kV X-rays to a 3 x 3 mm2 area in the centre of the lower tongue surface. Acute mucosal ulceration, as a clinically relevant reaction, was used as the quantal endpoint for dose response analyses by probit analysis. As a secondary endpoint the time course, i.e. time to first diagnosis of ulcer (latent time) and individual ulcer duration, was analysed. KGF was applied before, after or in combination before and after radiation exposure. RESULTS: Administration of KGF in all protocols resulted in a significant reduction of the incidence of oral mucosal ulceration, as illustrated by an increase in iso-effective dose from 10.9 to 24.9 Gy; the corresponding dose-modification factors ranged between 1.7 and 2.3. The effect was most pronounced when KGF was applied after irradiation. In all protocols where KGF was given after irradiation, a significant shortening of the latent time to ulceration from 11 to 6-8 days was observed. CONCLUSIONS: The mechanisms underlying the amelioration of the oral mucosal response to single-dose irradiation remain unclear. However, KGF represents a promising approach for the effective management of acute radiation reactions in oral, gastrointestinal and cutaneous epithelia after radiation exposure. PMID- 11258849 TI - Absence of protective role of afferent nerves in early intestinal mucosal alterations induced by abdominal irradiation in rats. AB - PURPOSE: To assess the early effects of primary afferent nerve suppression by systemic treatment with the neurotoxin capsaicin in an acute model of abdominal irradiation in rats (10Gy, gamma). MATERIALS AND METHODS: Changes in myeloperoxidase (MPO) activity, calcitonin gene-related peptide (CGRP) tissue content, number of mast cells and apoptotic cells were determined in jejunum and ileum in four groups of rat male Wistar (vehicle sham-irradiated, vehicle irradiated, capsaicin sham-irradiated and capsaicin irradiated) at 1 and 3 days post-irradiation. RESULTS: In vehicle irradiated rats, CGRP was significantly increased from the first day after irradiation in jejunal mucosa; MPO activity increased in both segments at day 3 but not at day 1 after irradiation; the number of detectable mucosal mast cells dropped to nearly zero on days 1 and 3, while the apoptotic cells in the intestinal mucosa were significantly increased at day 1. Similar results were obtained for mast cells and apoptosis in capsaicin irradiated rats as compared to capsaicin sham-irradiated rats, while MPO activity was significantly increased and CGRP concentration in jejunal mucosa significantly decreased from the first day in these rats in comparison with capsaicin sham-irradiated rats. CONCLUSIONS: Intestinal sensory innervation seems not to have a major protective role against a radiation-induced intestinal inflammatory reaction. PMID- 11258850 TI - Role of cell cycle control in radiosensitization of mouse spermatogonial stem cells. AB - PURPOSE: To investigate the possible role of cell cycle arrest in the radiosensitization of mouse spermatogonial stem cells due to small conditioning X ray exposures. MATERIALS AND METHODS: A 24 h fractionation interval between conditioning (1 Gy) and challenging (8 or 9 Gy) exposures was used. Two approaches were followed: the first in the Swiss random-bred wild-type mouse of the radiation-induced cell cycle arrest-evading agents 3-aminobenzamide (3-AB) and caffeine; and, second, using the C57BL/6 mouse of different p53 status. As biological parameter stem cell survival was analysed by the repopulation index (RI) method and chromosomal translocations were recorded using spermatocyte analysis at appropriate posttreatment periods. RESULTS: In the Swiss wild-type mouse, the application of 3-AB or caffeine significantly suppressed the sensitization of stem cells towards killing or translocation induction. In the C57BL/6 mouse, somewhat more variability in response was observed but no significant differences in sensitization between the p53 +/+, +/- or -/- mouse were recorded, suggesting no involvement of p53 in this process. CONCLUSIONS: The results indicate that p53-independent cell cycle regulation plays an important role in the radiosensitization of mouse spermatogonial stem cells. PMID- 11258851 TI - Role of stress responses in human cell survival following exposure to ultraviolet C radiation. AB - PURPOSE: To investigate in human skin and other cells the role of tyrosine kinase and protein kinase-C (PKC) in eliciting cell-signalling responses to UV radiation (UVR) that affect the survival of irradiated cells. MATERIALS AND METHODS: The survival of HeLa S3 cells, NCTC 2544 human keratinocytes and A431 human epidermal carcinoma cells was measured following incubation with various tyrosine kinase or PKC inhibitors and exposure to UVC (254nm) radiation. In addition, Western blotting measured PKC isozyme expression in human keratinocytes following UVC exposure. RESULTS: It was confirmed that inhibition of tyrosine kinase activation reduces the survival of UV-irradiated HeLa S3 cells. However, no effect was seen on the survival of either NCTC 2544 human keratinocytes or A431 human epidermal carcinoma cells. In contrast, specific inhibition of PKC reduced the survival of UV-irradiated keratinocytes but had no effect on HeLa cells. Comparison of the effects of different inhibitors in keratinocytes suggested that this effect was mediated mostly through PKCmu and PKClambda/iota. In addition, keratinocyte exposure to UVC induced large and temporally distinct increases in PKCmu and PKClambda/iota. CONCLUSIONS: The survival of NCTC 2544 keratinocytes, but not HeLa S3 cells, following UVC exposure is mediated by signalling through PKC, mostly PKCmu and PKClambda/iota. Further study is required to confirm these results in normal human keratinocytes. PMID- 11258852 TI - Comparative biokinetics of tritium in rats during continuous ingestion of tritiated water and tritium-labeled food. AB - PURPOSE: The biokinetics of tritium during continuous ingestion of tritiated water and tritiated wheat were investigated to estimate the radiation dose rates at the end of two modes of chronic exposure. MATERIALS AND METHODS: Wistar strain male rats continuously ingested tritiated water as drinking water and tritiated wheat as food for 14 weeks. Urine and tissue samples were obtained and total tritium in the fresh wet samples and organically bound tritium (OBT) in the freeze-dried samples were determined. RESULTS: The biokinetics of tritium was different between the two modes of exposure. The concentration of total tritium in the tissues exposed to tritiated water attained a steady-state condition by 2 3 weeks. The steady-state condition in the case of exposure to tritiated wheat was not observed for 10 weeks after the start of exposure in the majority of tissues. The relatively efficient and prolonged OBT formation during chronic exposure to tritiated wheat resulted in relatively high incorporation and retention of tritium in the tissues compared with those for exposure to the same activity of tritiated water. CONCLUSION: Radiation dose rates estimated at the end of continuous ingestion showed that tritiated wheat gave higher dose rates than tritiated water by a factor of 1.3 to 4.5, but the factors were within 2.0 in the majority of tissues except for small intestine and adipose tissue. PMID- 11258853 TI - Effect of external irradiation on the gastrointestinal absorption of uranium and neptunium in rats. AB - PURPOSE: The gastrointestinal absorption and systemic distribution of uranium and neptunium were determined after external gamma irradiation. MATERIALS AND METHODS: Rats were exposed to a single whole-body dose of gamma radiation (6Gy; 0.75Gy.min(-1)). Three days after irradiation they were orally and/or intravenously contaminated with 100 microg.kg(-1) uranium or 3kBq.kg(-1) neptunium. The gastrointestinal absorption and organ distribution of both radionuclides were measured 6 days after irradiation. RESULTS: External irradiation increased the intestinal transit time of uranium and neptunium but had no effect on their gastrointestinal absorption. The average fractional absorption was determined to be 0.93 and 0.98% (uranium) and 4.7 and 4.8% (neptunium) for the irradiated and non-irradiated rats respectively. The excretion of uranium and neptunium was not affected by the irradiation. CONCLUSION: A 6 Gy whole-body irradiation (gamma; 0.75Gy.min(-1)) did not affect the absorption of uranium and neptunium after oral intake. PMID- 11258854 TI - Radiation-induced acute intestinal inflammation differs following total-body versus abdominopelvic irradiation in the ferret. AB - PURPOSE: The studies were designed to investigate the differences in the intestinal inflammatory response following abdominopelvic or total-body irradiation in a ferret model. MATERIALS AND METHODS: Ferrets were exposed either to total-body or to abdominopelvic gamma-radiation (5 Gy) and various parameters of inflammation studied in the jejunum, ileum and colon 2 and 7 days later. RESULTS: Abdominopelvic and, to a greater extent, total-body irradiation caused weight loss by 7 days. White blood cell counts were reduced in both groups, but more so following total-body irradiation. Myeloperoxidase activity was significantly increased in the ileum 2 days after abdominopelvic irradiation, but it was reduced after total-body irradiation. Total-body irradiation increased tissue prostaglandin E2 levels in all regions at 2 days and decreased jejunal leukotriene B4 levels in the jejunum at both time points. Ileal prostaglandin E2 levels were increased 2 days after abdominopelvic irradiation. Expression of inducible nitric oxide synthase was not altered by either irradiation protocol. CONCLUSIONS: The data show that there are regional differences in the intestinal response to irradiation, depending on whether it was delivered to the whole body or locally to the abdominopelvic region. In particular, the ileum exhibited an acute increase in myeloperoxidase activity following abdominopelvic but not total body irradiation. PMID- 11258855 TI - Response to the "In my opinion article". PMID- 11258856 TI - Infection and exposure control in North American dental schools. AB - The results of a survey sent to all U.S. and Canadian dental schools clearly indicate that several substantial changes have occurred in infection control and exposure control in the past fifteen to twenty years. Predominant among these are that the responsibility for instrument preparation and sterilization in most schools has passed from the student to trained staff, the routine practice of universal precautions has eliminated the need to treat patients known to carry bloodborne diseases in a special area, and pre-admission and enrollment vaccination and health screening requirements have changed significantly. Other important changes result from the fact that the majority of U.S. schools responding to the survey are now, to a great extent, in compliance with the OSHA Bloodborne Pathogens Standard or equivalent requirements. PMID- 11258857 TI - An association perspective: responding to the American Dental Association's future of dentistry project. AB - In response to the American Dental Association's (ADA) Future of Dentistry Project, the American Dental Education Association (ADEA) provided its perspective on the most critical issues facing the dental profession. ADEA responded in six areas, each corresponding to the areas of focus in the ADA project. The comments in this Association Report reflect those provided to the six panels conducting the project. PMID- 11258858 TI - Report of the ADEA president's task force on the Surgeon General's report on oral health. American Dental Education Association. AB - In response to the first-ever Surgeon General's report on oral health, released on May 25, 2000, ADEA President Rowland A. Hutchinson, D.D.S., M.S., appointed a task force to study the report from the perspective of dental education. The task force was charged with making recommendations to the ADEA Board of Directors as to the Association's message to members and the general public, the Association's role in addressing oral health disparities, the legislative and policy implications of the report, and areas of collaboration between ADEA and others in the dental and health professions. The task force reviewed the report and made five recommendations, including increasing public awareness of the report's messages, promoting collaborative activities with a goal of improving America's oral health, and providing leadership in the drive to promote the incorporation of new science in dental education. The task force also identified numerous ADEA initiatives that address issues related to the Surgeon General's report. PMID- 11258859 TI - Reflections on changes in geriatric dentistry. AB - The development of geriatric dental education programs in the United States and at the University of Iowa over the last twenty years is reviewed. The program at Iowa evolved from a didactic elective program taught by a single faculty person to required didactic and clinical programs that include a special care clinic in the dental school and a mobile unit with portable equipment serving ten area nursing homes with comprehensive care. Factors influencing the curriculum development are identified and discussed, and as no dental schools are the same, some general applications are suggested from the Iowa experience. PMID- 11258860 TI - PAT Program report: background and current status. Proficiency Analytical Testing (PAT) program. PMID- 11258862 TI - AIHA consultants listing. PMID- 11258861 TI - ELPAT Program report: background and current status. Environmental Lead Proficiency Analytical Testing. PMID- 11258863 TI - Dermal absorption of neat liquid solvents on brief exposures in volunteers. AB - The dermal absorption of liquid 1,1,1-trichloroethane (111TRI), trichloroethene (TRI), tetrachloroethene (TETRA), toluene (TOL), and m-xylene (XYL) was studied in volunteers. The solvents were applied for 3 min on the volar forearm over an area of 27 cm2. An inhalation exposure with a known input rate served as a reference exposure. Using the linear system dynamics method, permeation rates were calculated from exhaled air concentration courses measured after both inhalation and dermal exposure. The permeation time courses of the solvents showed two different patterns. TRI, TOL, and 111TRI in three subjects showed fast increase in permeation, reaching maximal permeation rates a few minutes after initiation of exposure. Slower permeation was seen in the other three subjects exposed to 111TRI and in all subjects exposed to TETRA and XYL with the time of maximal permeation rates of 15-25 min. These differences in the permeation may partly be explained by the irritation of the skin observed in all subjects showing fast permeation kinetics. The flux into the skin averaged over the exposure period amounted to 56, 430, 69, 223, and 46 nmol/cm2/min for 111TRI, TRI, TETRA, TOL, and XYL, respectively. Comparing the dermal uptake with the respiratory uptake at the TLV, all solvents showed substantial skin absorption, although at present only TOL has a skin indication in the American Conference of Governmental Industrial Hygienists threshold limit value list. PMID- 11258864 TI - Effect of impactor inlet efficiency on the measurement of wood dust size distribution. AB - Two designs of cascade impactor inlets were evaluated experimentally to determine their particle sampling differences. One was the standard shrouded inlet provided with the low flow rate (2 L/min) Marple Personal Cascade impactor (Marple). The other was a simple vertical tube used with medium flow rate (15-30 L/min) impactors used as area samplers (e.g., Microorifice Uniform Deposit Impactor, Andersen, Sierra, Berner, or University of Washington [UW] impactor). When two impactors (Marple and UW) were used side-by-side to measure particle size distributions in the wood products industry, they often produced very different size distributions. In the laboratory the particle aspiration efficiencies of both impactor inlets were determined using monodisperse solid particles with aerodynamic diameters (Da) ranging from 5 to 68 microm at wind speeds of 0.55 and 1.1 m/sec. For particles with Da greater than 10 microm, the sample obtained using a simple vertical inlet tube was more representative of the environmental concentration than that obtained using the shielded inlet provided with the Marple impactor. Tests conducted using the Marple impactor inlet without a visor produced aspiration efficiencies that varied with inlet orientation and wind speed. The vertical sampling tube was found to be a better sampler inlet for indoor aerosol sampling. PMID- 11258865 TI - Design and evaluation of a breath-analysis system for biological monitoring of volatile compound. AB - To ensure the health and safety of workers, integrated industrial hygiene methodologies often include biological monitoring of the workers to help understand their exposure to chemicals. To this end, a field-portable breath analysis system was developed and tested to measure selected solvents in exhaled air. The exhaled breath data were evaluated using a physiologically based pharmacokinetic (PBPK) model to relate exposure to tissue dose. The system was designed to monitor workers every time they entered or left a work environment--a vast improvement over current 8-hour integrated monitoring strategies. The system combines (1) chemical dosimeters to measure airborne contaminant levels (analyzed in the field/ workplace); (2) real-time breath analysis to quantitate exposure; and 3) PBPK models to estimate internal target tissue dose. To evaluate the system, field tests were conducted at two locations: (1) at an incinerator in Tennessee monitoring benzene and toluene exposures; and (2) a waste repackaging facility in Washington State where hexane, trimethylbenzene, and methylene chloride was monitored. Exhaled breath was sampled and analyzed before and after each specific job task, which ranged from 15 min to 8 hours in duration. In both field studies several volunteers had posttask breath levels higher than pretask levels. The greatest increase corresponded to 573 ppb for methylene chloride and 60 ppb for toluene. Compared with breath analysis, the chemical dosimeters underpredicted the dosimetry, particularly for longer sampling intervals when the volume of air sampled may have diluted exposures. The results of the field studies illustrate the utility of monitoring workers for exposures throughout the day, particularly when job-specific tasks may indicate a potential for exposure. PMID- 11258866 TI - Respiratory protection as a function of respirator fitting characteristics and fit-test accuracy. AB - The fitting characteristics of particulate respirators are no longer assessed in the National Institute for Occupational Safety and Health respirator certification program. It is important for respirator program administrators to understand the implications of that change and the additional burden it may impose. To address that issue, a typical respirator fit-testing program is analyzed using a mathematical model that describes the effectiveness of a fit testing program as a function of the fitting characteristics of the respirator and the accuracy of the fittesting method. The model is used to estimate (1) the respirator assignment error, the percentage of respirator wearers mistakenly assigned an ill-fitting respirator; (2) the number of fit-test trials necessary to qualify a group of workers for respirator use; and (3) the number of workers who will fail the fit-test with any candidate respirator model and thereby fail to qualify for respirator use. Using data from previous studies, the model predicts respirator assignment errors ranging from 0 to 20%, depending on the fitting characteristics of the respirator models selected and the fit-testing method used. This analysis indicates that when respirators do not necessarily have good fitting characteristics, respirator program administrators should exercise increased care in the selection of respirator models and increased care in fit-testing. Also presented are ways to assess the fitting characteristics of candidate respirator models by monitoring the first-time fit-testing results. The model demonstrates that significant public health and economic benefits can result when only respirators having good fitting characteristics are purchased and respirators are assigned to workers using highly accurate fit-testing methods. PMID- 11258867 TI - Quantitative level of protection offered to workers by ACGIH threshold limit values occupational exposure limits. AB - The details of the example or modeling methodologies used herein are not critical to the general point of this article, which advises the estimation of residual risk at the OEL by using some quantitative modeling structure. Specifically, the authors believe that an explicit attempt to gauge the level of residual risk at the OEL based on conceptual stochastic models with transparent and testable assumptions could be seen as an important enhancement to the process. This is especially true in sharing the OEL deliberations and explaining OEL decisions to the stakeholders. Indeed, if this approach is used, it is critically important to understand and continually communicate that this "cloud of uncertainty" represents model estimates in which the true risk would most likely be less than worst case estimates and could possibly be zero. It is also possible but highly unlikely that it could be higher than the worst case upper-bound estimate. The above quantitative estimation scheme represents a possible improvement that could provide a reasoned attempt on the part of the risk assessors to use rational science (i.e., conceptual models with transparent and testable assumptions) to inform all of the OEL users and stakeholders of their meaning. PMID- 11258868 TI - Subchronic inhalation exposures to aerosols of three petroleum lubricants. AB - Subchronic inhalation studies were performed with three petroleum lubricants: generic cutting oil (GCO), generic gear oil (GO), and generic commercial engine oil (CEO). Each formulation had a mineral oil base. Sprague-Dawley rats were exposed 6 hours/day, 5 days/week for 13 weeks to aerosol concentrations of 0 (untreated controls), 0 (sham-exposed controls), approximately 50, 150, or 400 520 mg/m3. At necropsy, 15 rats/sex/group were sampled for serum chemistry (18 parameters), hematology, and weights of 13 organs. Testis and epididymis of males in the control and high-dose group were used for number of spermatids and morphology of epididymal sperm. Histopathological slides were evaluated for 22 or more organs. Pulmonary function tests were done on 10 additional males/group. Pulmonary hydroxyproline was measured in these rats for GCO and GO. Residual oil in the lungs was determined for GCO. The primary organ affected by exposures to these three formulations was the lung; the main observed effects were accumulation of foamy macrophages in pulmonary alveoli and alveolar walls, very mild thickening of alveolar walls due to foamy macrophages and a mixed cell infiltrate, and subtle epithelial hyperplasia. The foamy macrophages tended to group together in aggregates, and the aggregates seemed responsible for plaques seen visibly on the surface of the lung. These histological changes were accompanied by concentration-related increases in lung weight and pulmonary hydroxyproline, whereas pulmonary function tests were generally unaffected. Effects distal to the lung were more limited. These results indicated low toxicity of these aerosols in this model. PMID- 11258869 TI - "Total" and respirable dust exposures in the U.S. carbon black manufacturing industry. AB - This article describes an industrywide exposure assessment study of "total" and respirable carbon black dust exposures among U.S. carbon black manufacturing workers conducted between 1993 and 1995. In addition to updating a 1979-1980 industrywide "total" dust study conducted among all major U.S. carbon black producers, this research was the first comprehensive evaluation of respirable dust exposures in the U.S. industry. A total of 2060 samples were collected (n = 1,004 "total," n = 1,056 respirable). The distributions of both dust fractions were lognormal with an overall "total" dust mean concentration, represented as the maximum variance unbiased estimator of 0.59 mg/m3 (range = 0.01-13.25 mg/m3), and an overall respirable dust mean concentration of 0.15 mg/m3 (range = 0.01 2.62 mg/m3). The fraction of "total" dust exposures greater than the current Occupational Safety and Health Administration permissible exposure limit of 3.5 mg/m3 was less than 3%. Material handling jobs experienced the highest "total" and respirable dust exposures, a finding consistent with the 1979-1980 study. The highest mean exposure for an individual job title was observed for product baggers/packers/sackers at 2.30 mg/m3 for "total" dust and 0.48 mg/m3 as respirable dust. Overall, mean "total" dust exposures had decreased significantly (up to 70%) for the majority of job classifications since the 1979-80 study. To evaluate the relationship between "total" and respirable dust fractions, 680 matched pairs of respirable and "total" samples were analyzed with a resulting mean ratio of 0.37 (respirable fraction to "total" dust). A log-transformed regression equation was obtained that provides a predictive relationship between "total" and respirable carbon black dust concentrations that may be applied to estimate the respirable fraction of historical "total" dust data collected under similar environmental and manufacturing conditions. PMID- 11258870 TI - Interchangeability of gas detection tubes and hand pumps. AB - Users of gas detection tubes occasionally seek the convenience of using a single hand pump with different brands of tubes, to avoid the need to carry more than one pump. Several professional organizations recommend against such interchange. However, these recommendations appear to be based on a single study of pump designs that mostly are no longer in use. The present study was undertaken to determine if current hand pumps are interchangeable. Both piston-type and bellows type hand pumps were evaluated by comparing pump flow profiles and test gas measurements with a variety of tubes. The results demonstrate that three piston hand pumps in common use (Sensidyne/Gastec GV/100, RAE Systems LP-1200, and Matheson-Kitagawa 8104-400A) are fully interchangeable. Two bellows pumps (Draeger Accuro and MSA Kwik-Draw) also are interchangeable with each other. Mixing of bellows and piston systems is often possible, but there are enough exceptions to conclude that such practice should be discouraged because it can give inaccurate readings. It is recommended that technical standards be adopted, such as total volume and an initial pump vacuum or a pump flow curve, to assess hand pump interchangeability. When two manufacturers' pumps meet the same standard and routine leak tests are conducted, interchangeability is scientifically valid and poses no risk to the end user while offering greater convenience. PMID- 11258871 TI - Airborne concentrations of ethyl and methyl cyanoacrylate in the workplace. AB - A survey was conducted of persons who manufacture, mix, bottle, and package methyl 2-cyanoacrylate (MCA) and ethyl 2-cyanoacrylate (ECA). Airborne concentrations of these cyanoacrylates also were measured during waste-handling operations. During a 1-week period, 162 personal and area samples were collected. About 90% of the samples were analyzed for ECA (the predominant adhesive being manufactured at the facility). About 50% of the samples were collected during periods of 15 min or less, the remainder for 15 to 240 min. Some 8-hour time weighted average (TWA) samples also were collected. Samples were collected using Tenax tubes with subsequent analysis by high-performance liquid chromatography. Most samples were collected where highest exposure was likely (e.g., during mixing, bottling, and packaging). Peak concentrations of exposure (duration of 15 min or less), measured during a variety of tasks, ranged from 0.003 to 1.5 ppm. In particular, personal mean short-term airborne concentrations of ECA for the mixing operators ranged from 0.039 ppm to 0.650 ppm, while various 10-min to 1 hour activities were performed, with a TWA concentration of 0.07 ppm. Personal short-term airborne concentrations of ECA for bottling and packaging workers (n = 60) were 0.040 ppm +/- 0.016 ppm (mean +/- standard deviation), with similar 8 hour TWA concentrations due to the continuous nature of bottling and packaging. Other personal samples were not significantly different. The area samples were normally within a factor of 2 of the peak personal sampling results. These data indicate that, when handled at room temperature and relative humidity ranging from 40-69%, both MCA and ECA produce airborne concentrations that are nearly always less than about 0.1 ppm, which is less than the threshold of irritation. PMID- 11258872 TI - Ion mobility spectrometric monitoring of phosdrin from foliage in greenhouse. AB - The monitoring of Phosdrin (mevinphos; insecticide) from foliage and foliage extracts was achieved by an aspiration-type ion mobility spectrometer. This technique is based on ion mobility, which is proportional to the molecular weight, shape, and charge. The operation principle of the ion mobility spectrometer is to measure mobility distribution changes of product and reactant ions. This technique can measure positive and negative ion clusters at the same time in six different measuring electrodes. Each measuring electrode detects a different portion of the ion mobility distribution formed within the cell's radioactive source. The pattern recognition used is based on differences in the gas profiles for different compounds. This study shows that an ion mobility spectrometer can be used to monitor Phosdrin from foliage without the need for any time-consuming extraction procedure. The responses for Phosdrin-containing and background (control) samples were easily separated from each other. The responses declined as a function of time in the positive and sum response channels. In addition, the sum of the absolute values of signals at six measuring channels (sum response) were linearly proportional to the concentration of Phosdrin. Just before application (i.e., in background), this value was 41 bits, whereas these values were 10-fold, 11-fold, 8-fold, 6-fold, 5-fold, and 3.5-fold at the time points 4, 8, 11, 24, 50, and 72 hours after the spraying of Phosdrin. PMID- 11258873 TI - Evaluation of respiratory and cutaneous doses and urinary excretion of alkylphosphates by workers in greenhouses treated with omethoate, fenitrothion, and tolclofos-methyl. AB - This research evaluated exposure pathways across work tasks for three organophosphate pesticides in a group of greenhouse workers. During reentry in ornamental plant greenhouses, five male workers were monitored for five consecutive days. Skin contamination (excluding hands) was evaluated with nine pads of filter paper placed on the skin. Hand contamination was assessed by washing with 95% ethanol. Respiratory exposure was evaluated by personal air sampling. The respiratory dose was based on a lung ventilation of 20 L/min. The doses absorbed were estimated assuming 10% skin penetration and 100% lung retention. Urinary alkylphosphates were assayed in the 24-hour urine samples of the days on which exposure was evaluated. Respiratory exposure was usually less than skin contamination, being 4.5 +/- 8.4%, 9.9 +/- 10.0%, and 49.5 +/- 26.6% (mean +/- standard deviation) of total exposure for omethoate, tolclofos-methyl, and fenitrothion, respectively. Multiple regression analysis showed that urinary alkylphosphate (nmol/24 hours) (y) was significantly correlated (r = 0.716, p < 0.001) with the respiratory doses of the three active ingredients absorbed the same day (x1) and with the cutaneous dose absorbed the previous day (x2). The relationship was expressed by the equation y = 0.592x2 + 0.117x, + 156.364. The doses of omethoate absorbed by one worker were more than 45 times the acceptable daily intake (ADI) of 1.41 nmol/kg body weight (b.w.) The ADI for fenitrothion and tolclofos-methyl (10.8 and 212.6 nmol/kg body weight, respectively) were never exceeded. High absorption by one worker underlines the importance of correct use of protective clothing. In this study the hands were always a source of contact with the pesticides. Greater precautions should be taken to reduce contamination (clean gloves, constant use of gloves). PMID- 11258874 TI - Workplace protection factors--supplied air hood. AB - Several organizations list assigned protection factors. For supplied air hoods, the value of the assigned protection factors varies from <10 to 2,000 depending on the organization. Workplace protection factors (WPFs) of a supplied air hood were measured during aircraft sanding and painting operations on several types of aircraft to evaluate whether the American National Standard Z88.2 (1992) assigned protection factor of 1,000 was realistic. The primary contaminant during these activities is strontium chromate. Samples collected inside the hood show that employees during sanding and painting operations were not exposed to strontium. The respirator performed adequately. This study is consistent with other simulated and WPF studies in that the ANSI Z88.2 WPF of 1,000 is supported. PMID- 11258875 TI - Identification of hydrogen bonds between Escherichia coli DNA polymerase I (Klenow fragment) and the minor groove of DNA by amino acid substitution of the polymerase and atomic substitution of the DNA. AB - DNA polymerases replicate DNA with high fidelity despite the small differences in energy between correct and incorrect base pairs. X-ray crystallographic and structure-activity kinetic experiments have implicated interactions with the minor groove of the DNA as being crucial for catalysis and fidelity. The current hypothesis is that polymerases check the geometry of the base pairs through hydrogen bonds and steric interactions with the minor groove of the DNA. The mechanisms by which these interactions are related to catalysis and fidelity are not known. In this manuscript, we have studied these interactions using a combination of site-specific mutagenesis of Escherichia coli DNA polymerase I (Klenow fragment) and atomic substitution of the DNA. Crystal structures have predicted hydrogen bonds from Arg668 to the terminal base on the primer (P1) and Gln849 to its base pair partner (T1). Kinetic studies, however, have implicated the minor groove of the primer terminus but not its base pair partner as being important to catalysis and fidelity. Hydrogen bonds between Arg668 and Gln849 to the DNA were probed with the site specific mutants, R668A and Q849A. Hydrogen bonds from the DNA were probed with three oligodeoxynucleotides which have a guanine or 3-deazaguanine (3DG) at P1, T1, or T2. We found that the pre-steady state parameter k(pol) was decreased with R668A (40-fold) and Q849A (150-fold) or with 3DG at P1 (300-fold) or T2 (25-fold). When R668A was combined with 3DG at P1 the decrease in rate was only 80-fold, consistent with a hydrogen bond between Arg668 and P1. In contrast, when the 3DG substitution at P1 was combined with Q849A the rate reduction was 15000-fold. Similar reactions between R668A or Q849A and T2 showed that there are interactions between these sites although the interactions are not as strong as between P1 and R668. PMID- 11258876 TI - Cofactor-induced refolding: refolding of molten globule carbonic anhydrase induced by Zn(II) and Co(II). AB - The stability versus unfolding to the molten globule intermediate of bovine carbonic anhydrase II (BCA II) in guanidine hydrochloride (GuHCl) was found to depend on the metal ion cofactor [Zn(II) or Co(II)], and the apoenzyme was observed to be least stable. Therefore, it was possible to find a denaturant concentration (1.2 M GuHCl) at which refolding from the molten globule to the native state could be initiated merely by adding the metal ion to the apo molten globule. Thus, refolding could be performed without changing the concentration of the denaturant. The molten globule intermediate of BCA II could still bind the metal cofactor. Cofactor-effected refolding from the molten globule to the native state can be summarized as follows: (1) initially, the metal ion binds to the molten globule; (2) compaction of the metal-binding site region is then induced by the metal ion binding; (3) a functioning active center is formed; and (4) finally, the native tertiary structure is generated in the outer parts of the protein. PMID- 11258877 TI - Putative disulfide-forming pathway of porcine insulin precursor during its refolding in vitro. AB - Although the structure of insulin has been well studied, the formation pathway of the three disulfide bridges during the refolding of insulin precursor is ambiguous. Here, we reported the in vitro disulfide-forming pathway of a recombinant porcine insulin precursor (PIP). In redox buffer containing L arginine, the yield of native PIP from fully reduced/denatured PIP can reach 85%. The refolding process was quenched at different time points, and three distinct intermediates, including one with one disulfide linkage and two with two disulfide bridges, have been captured and characterized. An intra-A disulfide bridge was found in the former but not in the latter. The two intermediates with two disulfide bridges contain the common A20-B19 disulfide linkage and another inter-AB one. Based on the time-dependent formation and distribution of disulfide pairs in the trapped intermediates, two different forming pathways of disulfide bonds in the refolding process of PIP in vitro have been proposed. The first one involves the rapid formation of the intra-A disulfide bond, followed by the slower formation of one of the inter-AB disulfide bonds and then the pairing of the remaining cysteines to complete the refolding of PIP. The second pathway begins first with the formation of the A20-B19 disulfide bridge, followed immediately by another inter-AB one, possibly nonnative. The nonnative two disulfide intermediates may then slowly rearrange between CysA6, CysA7, CysA11, and CysB7, until the native disulfide bond A6-A11 or A7-B7 is formed to complete the refolding of PIP. The proposed refolding behavior of PIP is compared with that of IGF-I and discussed. PMID- 11258878 TI - Structural characterization of n-butyl-isocyanide complexes of cytochromes P450nor and P450cam. AB - Alkyl-isocyanides are able to bind to both ferric and ferrous iron of the heme in cytochrome P450, and the resulting complexes exhibit characteristic optical absorption spectra. While the ferric complex gives a single Soret band at 430 nm, the ferrous complex shows double Soret bands at 430 and 450 nm. The ratio of intensities of the double Soret bands in the ferrous isocyanide complex of P450 varies, as a function of pH, ionic strength, and the origin of the enzyme. To understand the structural origin of these characteristic spectral features, we examined the crystallographic and spectrophotometric properties of the isocyanide complexes of Pseudomonas putida cytochrome P450cam and Fusarium oxysporum cytochorme P450nor, since ferrous isocyanide complex of P450cam gives a single Soret band at 453 nm, while that of P450nor gives one at 427 nm. Corresponding to the optical spectra, we observed C-N stretching of a ferrous iron-bound isocyanide at 2145 and 2116 cm(-1) for P450nor and P450cam, respectively. The crystal structures of the ferric and ferrous n-butyl isocyanide complexes of P450cam and P450nor were determined. The coordination structure of the fifth Cys thiolate was indistinguishable for the two P450s, but the coordination geometry of the isocyanide was different for the case of P450cam [d(Fe-C) = 1.86 A, angleFe-C-N = 159 degrees ] versus P450nor [d(Fe-C) = 1.85 A, angleFe-C-N = 175 degrees ]. Another difference in the structures was the chemical environment of the heme pocket. In the case of P450cam, the iron-bound isocyanide is surrounded by some hydrophobic side chains, while, for P450nor, it is surrounded by polar groups including several water molecules. On the basis of these observations, we proposed that the steric factors and/or the polarity of the environment surrounding the iron-bound isocyanide significantly effect on the resonance structure of the heme(Fe)-isocyanide moiety and that differences in these two factors are responsible for the spectral characteristics for P450s. PMID- 11258879 TI - Probing erectile function: S-(2-boronoethyl)-L-cysteine binds to arginase as a transition state analogue and enhances smooth muscle relaxation in human penile corpus cavernosum. AB - The boronic acid-based arginine analogue S-(2-boronoethyl)-L-cysteine (BEC) has been synthesized and assayed as a slow-binding competitive inhibitor of the binuclear manganese metalloenzyme arginase. Kinetic measurements indicate a K(I) value of 0.4-0.6 microM, which is in reasonable agreement with the dissociation constant of 2.22 microM measured by isothermal titration calorimetry. The X-ray crystal structure of the arginase-BEC complex has been determined at 2.3 A resolution from crystals perfectly twinned by hemihedry. The structure of the complex reveals that the boronic acid moiety undergoes nucleophilic attack by metal-bridging hydroxide ion to yield a tetrahedral boronate anion that bridges the binuclear manganese cluster, thereby mimicking the tetrahedral intermediate (and its flanking transition states) in the arginine hydrolysis reaction. Accordingly, the binding mode of BEC is consistent with the structure-based mechanism proposed for arginase as outlined in Cox et al. [Cox, J. D., Cama, E., Colleluori D. M., Pethe, S., Boucher, J. S., Mansuy, D., Ash, D. E., and Christianson, D. W. (2001) Biochemistry 40, 2689-2701.]. Since BEC does not inhibit nitric oxide synthase, BEC serves as a valuable reagent to probe the physiological relationship between arginase and nitric oxide (NO) synthase in regulating the NO-dependent smooth muscle relaxation in human penile corpus cavernosum tissue that is required for erection. Consequently, we demonstrate that arginase is present in human penile corpus cavernosum tissue, and that the arginase inhibitor BEC causes significant enhancement of NO-dependent smooth muscle relaxation in this tissue. Therefore, human penile arginase is a potential target for the treatment of sexual dysfunction in the male. PMID- 11258880 TI - Mechanistic and metabolic inferences from the binding of substrate analogues and products to arginase. AB - Arginase is a binuclear Mn(2+) metalloenzyme that catalyzes the hydrolysis of L arginine to L-ornithine and urea. X-ray crystal structures of arginase complexed to substrate analogues N(omega)-hydroxy-L-arginine and N(omega)-hydroxy-nor-L arginine, as well as the products L-ornithine and urea, complete a set of structural "snapshots" along the reaction coordinate of arginase catalysis when interpreted along with the X-ray crystal structure of the arginase-transition state analogue complex described in Kim et al. [Kim, N. N., Cox, J. D., Baggio, R. F., Emig, F. A., Mistry, S., Harper, S. L., Speicher, D. W., Morris, Jr., S. M., Ash, D. E., Traish, A. M., and Christianson, D. W. (2001) Biochemistry 40, 2678-2688]. Taken together, these structures render important insight on the structural determinants of tight binding inhibitors. Furthermore, we demonstrate for the first time the structural mechanistic link between arginase and NO synthase through their respective complexes with N(omega)-hydroxy-L-arginine. That N(omega)-hydroxy-L-arginine is a catalytic intermediate for NO synthase and an inhibitor of arginase reflects the reciprocal metabolic relationship between these two critical enzymes of L-arginine catabolism. PMID- 11258881 TI - Cathepsins X and B can be differentiated through their respective mono- and dipeptidyl carboxypeptidase activities. AB - Several new cysteine proteases of the papain family have been discovered in the past few years. To help in the assignment of physiological roles and in the design of specific inhibitors, a clear picture of the specificities of these enzymes is needed. One of these novel enzymes, cathepsin X, displays a unique specificity, cleaving single amino acid residues at the C-terminus of substrates very efficiently. In this study, the carboxypeptidase activities and substrate specificity of cathepsins X and B have been investigated in detail and compared. Using quenched fluorogenic substrates and HPLC measurements, it was shown that cathepsin X preferentially cleaves substrates through a monopeptidyl carboxypeptidase pathway, while cathepsin B displays a preference for the dipeptidyl pathway. The preference for one or the other pathway is about the same for both enzymes, i.e., approximately 2 orders of magnitude, a result supported by molecular modeling of enzyme-substrate complexes. Cleavage of a C-terminal dipeptide of a substrate by cathepsin X can become more important under conditions that preclude efficient monopeptidyl carboxypeptidase activity, e.g., nonoptimal interactions in subsites S(2)-S(1). These results confirm that cathepsin X is designed to function as a monopeptidyl carboxypeptidase. Contrary to a recent report [Klemencic, I., et al. (2000) Eur. J. Biochem. 267, 5404 5412], it is shown that cathepsins X and B do not share similar activity profiles, and that reagents are available to clearly distinguish the two enzymes. In particular, CA074 was found to inactivate cathepsin B at least 34000-fold more efficiently than cathepsin X. The insights obtained from this and previous studies have been used to produce an inhibitor designed to exploit the unique structural features responsible for the carboxypeptidase activity of cathepsin X. Although of moderate potency, this E-64 derivative is the first reported example of a cathepsin X-specific inhibitor. PMID- 11258882 TI - High resolution X-ray structures of different metal-substituted forms of phosphotriesterase from Pseudomonas diminuta. AB - Phosphotriesterase, isolated from the soil-dwelling bacterium Pseudomonas diminuta, catalyzes the detoxification of organophosphate-based insecticides and chemical warfare agents. The enzyme has attracted significant research attention in light of its possible employment as a bioremediation tool. As naturally isolated, the enzyme is dimeric. Each subunit contains a binuclear zinc center that is situated at the C-terminal portion of a "TIM" barrel motif. The two zincs are separated by approximately 3.4 A and coordinated to the protein via the side chains of His 55, His 57, His 201, His 230, Asp 301, and a carboxylated Lys 169. Both Lys 169 and a water molecule (or hydroxide ion) serve to bridge the two zinc ions together. Interestingly, these metals can be replaced with cadmium or manganese ions without loss of enzymatic activity. Here we describe the three dimensional structures of the Zn(2+)/Zn(2+)-, Zn(2+)/Cd(2+)-, Cd(2+)/Cd(2+)-, and Mn(2+)/Mn(2+)-substituted forms of phosphotriesterase determined and refined to a nominal resolution of 1.3 A. In each case, the more buried metal ion, referred to as the alpha-metal, is surrounded by ligands in a trigonal bipyramidal ligation sphere. For the more solvent-exposed or beta-metal ion, however, the observed coordination spheres are either octahedral (in the Cd(2+)/Cd(2+)-, Mn(2+)/Mn(2+) , and the mixed Zn(2+)/Cd(2+)-species) or trigonal bipyramidal (in the Zn(2+)/Zn(2+)-protein). By measuring the anomalous X-ray data from crystals of the Zn(2+)/Cd(2+)-species, it has been possible to determine that the alpha-metal ion is zinc and the beta-site is occupied by cadmium. PMID- 11258883 TI - Direct comparison of experimental and calculated folding free energies for hydrophobic deletion mutants of chymotrypsin inhibitor 2: free energy perturbation calculations using transition and denatured states from molecular dynamics simulations of unfolding. AB - Previous molecular dynamics (MD) simulations of thermal denaturation of chymotrypsin inhibitor 2 (CI2) have provided transition-state models in good agreement with experiment. Unfortunately, however, the comparisons have been necessarily indirect. The simulations have provided detailed structural information but not energetics, while from experiment, structure is inferred from a ratio of free energy changes upon mutation (Phi values). Here, direct comparison with experimental free energies is obtained by performing free energy perturbation calculations of hydrophobic deletion mutants of CI2 using transition and denatured-state structures from various denaturation MD simulations. The agreement between the calculated and experimental DeltaDeltaG and Phi values is quite good (R = 0.8-0.9). In addition, given the availability of realistic atomic models for the denatured protein, the common approach of using small peptides to represent the denatured state in stability calculations can now be evaluated. To this end, two different extended tripeptide models were used: one using the sequence from the protein with the residue to be mutated in the center and the other with this residue surrounded by Ala residues. The results for the two peptides agree neither with one another nor with the different full-length denatured-state models, which do provide results in good agreement with experiment. This finding is noteworthy because the denatured state of CI2 is very disrupted with little residual structure, such that the peptides might have been expected to serve as reasonable models. Overall the calculations presented here validate our previous MD-generated transition- and denatured-state models and therefore the simulated unfolding pathways and their relevance to refolding. PMID- 11258884 TI - Structural studies of duck delta 1 and delta 2 crystallin suggest conformational changes occur during catalysis. AB - Duck delta1 and delta2 crystallin are 94% identical in amino acid sequence, and while delta2 crystallin is the duck orthologue of argininosuccinate lyase (ASL) and catalyzes the reversible breakdown of argininosuccinate to arginine and fumarate, the delta1 isoform is enzymatically inactive. The crystal structures of wild type duck delta1 and delta2 crystallin have been solved at 2.2 and 2.3 A resolution, respectively, and the refinement of the turkey delta1 crystallin has been completed. These structures have been compared with two mutant duck delta2 crystallin structures. Conformational changes were observed in two regions of the N-terminal domain with intraspecies differences between the active and inactive isoforms localized to residues 23-32 and both intra- and interspecies differences localized to the loop of residues 74-89. As the residues implicated in the catalytic mechanism of delta2/ASL are all conserved in delta1, the amino acid substitutions in these two regions are hypothesized to be critical for substrate binding. A sulfate anion was found in the active site of duck delta1 crystallin. This anion, which appears to mimic the fumarate moiety of the argininosuccinate substrate, induces a rigid body movement in domain 3 and a conformational change in the loop of residues 280-290, which together would sequester the substrate from the solvent. The duck delta1 crystallin structure suggests that Ser 281, a residue strictly conserved in all members of the superfamily, could be the catalytic acid in the delta2 crystallin/ASL enzymatic mechanism. PMID- 11258885 TI - Local structural plasticity of the prion protein. Analysis of NMR relaxation dynamics. AB - A template-assisted conformational change of the cellular prion protein (PrP(C)) from a predominantly helical structure to an amyloid-type structure with a higher proportion of beta-sheet is thought to be the causative factor in prion diseases. Since flexibility of the polypeptide is likely to contribute to the ability of PrP(C) to undergo the conformational change that leads to the infective state, we have undertaken a comprehensive examination of the dynamics of two recombinant Syrian hamster PrP fragments, PrP(29-231) and PrP(90-231), using (15)N NMR relaxation measurements. The molecular motions of these PrP fragments have been studied in solution using (15)N longitudinal (T(1)) and transverse relaxation (T(2)) measurements as well as [(1)H]-(15)N nuclear Overhauser effects (NOE). These data have been analyzed using both reduced spectral density mapping and the Lipari-Szabo model free formalism. The relaxation properties of the common regions of PrP(29-231) and PrP(90-231) are very similar; both have a relatively inflexible globular domain (residues 128-227) with a highly flexible and largely unstructured N-terminal domain. Residues 29-89 of PrP(29-231), which include the copper-binding octarepeat sequences, are also highly flexible. Analysis of the spectral densities at each residue indicates that even within the structured core of PrP(C), a markedly diverse range of motions is observed, consistent with the inherent plasticity of the protein. The central portions of helices B and C form a relatively rigid core, which is stabilized by the presence of an interhelix disulfide bond. Of the remainder of the globular domain, the parts that are not in direct contact with the rigid region, including helix A, are more flexible. Most significantly, slow conformational fluctuations on a millisecond to microsecond time scale are observed for the small beta-sheet. These results are consistent with the hypothesis that the infectious, scrapie form of the protein PrP(Sc) could contain a helical core consisting of helices B and C, similar in structure to the cellular form PrP(C). Our results indicate that residues 90-140, which are required for prion infectivity, are relatively flexible in PrP(C), consistent with a lowered thermodynamic barrier to a template-assisted conformational change to the infectious beta-sheet-rich scrapie isoform. PMID- 11258886 TI - The role for an invariant aspartic acid in hypoxanthine phosphoribosyltransferases is examined using saturation mutagenesis, functional analysis, and X-ray crystallography. AB - The role of an invariant aspartic acid (Asp137) in hypoxanthine phosphoribosyltransferases (HPRTs) was examined by site-directed and saturation mutagenesis, functional analysis, and X-ray crystallography using the HPRT from Trypanosoma cruzi. Alanine substitution (D137A) resulted in a 30-fold decrease of k(cat), suggesting that Asp137 participates in catalysis. Saturation mutagenesis was used to generate a library of mutant HPRTs with random substitutions at position 137, and active enzymes were identified by complementation of a bacterial purine auxotroph. Functional analyses of the mutants, including determination of steady-state kinetic parameters and pH-rate dependence, indicate that glutamic acid or glutamine can replace the wild-type aspartate. However, the catalytic efficiency and pH-rate profile for the structural isosteric mutant, D137N, were similar to the D137A mutant. Crystal structures of four of the mutant enzymes were determined in ternary complex with substrate ligands. Structures of the D137E and D137Q mutants reveal potential hydrogen bonds, utilizing several bound water molecules in addition to protein atoms, that position these side chains within hydrogen bond distance of the bound purine analogue, similar in position to the aspartate in the wild-type structure. The crystal structure of the D137N mutant demonstrates that the Asn137 side chain does not form interactions with the purine substrate but instead forms novel interactions that cause the side chain to adopt a nonfunctional rotamer. The results from these structural and functional analyses demonstrate that HPRTs do not require a general base at position 137 for catalysis. Instead, hydrogen bonding sufficiently stabilizes the developing partial positive charge at the N7-atom of the purine substrate in the transition-state to promote catalysis. PMID- 11258887 TI - Structural bases for inhibitor binding and catalysis in polyamine oxidase. AB - Polyamine oxidase (PAO) carries out the FAD-dependent oxidation of the secondary amino groups of spermidine and spermine, a key reaction in the polyamine catabolism. The active site of PAO consists of a 30 A long U-shaped catalytic tunnel, whose innermost part is located in front of the flavin ring. To provide insight into the PAO substrate specificity and amine oxidation mechanism, we have investigated the crystal structure of maize PAO in the reduced state and in complex with three different inhibitors, guazatine, 1,8-diaminooctane, and N(1) ethyl-N(11)-[(cycloheptyl)methyl]-4,8-diazaundecane (CHENSpm). In the reduced state, the conformation of the isoalloxazine ring and the surrounding residues is identical to that of the oxidized enzyme. Only Lys300 moves away from the flavin to compensate for the change in cofactor protonation occurring upon reduction. The structure of the PAO.inhibitor complexes reveals an exact match between the inhibitors and the PAO catalytic tunnel. Inhibitor binding does not involve any protein conformational change. Such lock-and-key binding occurs also in the complex with CHENSpm, which forms a covalent adduct with the flavin N5 atom. Comparison of the enzyme complexes hints at an "out-of-register" mechanism of inhibition, in which the inhibitor secondary amino groups are not properly aligned with respect to the flavin to allow oxidation. Except for the Glu62 Glu170 pair, no negatively charged residues are involved in the recognition of substrate and inhibitor amino groups, which is in contrast to other polyamine binding proteins. This feature may be exploited in the design of drugs specifically targeting PAO. PMID- 11258888 TI - Linked folding and anion binding of the Bacillus subtilis ribonuclease P protein. AB - Ribonuclease P (RNase P) is the endoribonuclease responsible for the 5' maturation of precursor tRNA transcripts. In bacteria, RNase P is composed of a catalytic RNA subunit and an associated protein subunit that enhances the substrate specificity of the holoenzyme. We have initiated a study of the biophysical properties of the protein subunit from Bacillus subtilis RNase P (P protein) toward the goal of understanding the thermodynamics of RNase P holoenzyme assembly. The P protein is predominantly unfolded in 10 mM sodium cacodylate at neutral pH based on circular dichroism and NMR studies and therefore has several characteristics typical of "intrinsically unstructured" proteins. Furthermore, the P protein folds to its native alpha/beta structure upon addition of various small molecule anions. Anion-induced folding is best attributed to the binding of these anions to the folded state of the protein, and a model is presented which describes the observed tightly coupled folding and binding phenomena. The P protein also undergoes a cooperative folding transition upon addition of the osmolyte trimethylamine N-oxide (TMAO). The equilibrium constant of folding (K(fold)) at 37 degrees C for the P protein was determined to be 0.0071 +/- 0.0005 using a two-state folding model to describe the TMAO titration data. Thus, the folding and binding equilibria observed in the anion induced folding of the P protein can be uncoupled to determine the intrinsic binding affinities (K(a)'s) of the anionic ligands. Evidence that the osmolyte induced and the ligand-induced folded conformations of the P protein are structurally similar is also presented. PMID- 11258889 TI - Stabilization of coiled-coil peptide domains by introduction of trifluoroleucine. AB - Substitution of leucine residues by 5,5,5-trifluoroleucine at the d-positions of the leucine zipper peptide GCN4-p1d increases the thermal stability of the coiled coil structure. The midpoint thermal unfolding temperature of the fluorinated peptide is elevated by 13 degrees C at 30 microM peptide concentration. The modified peptide is more resistant to chaotropic denaturants, and the free energy of folding of the fluorinated peptide is 0.5-1.2 kcal/mol larger than that of the hydrogenated form. A similarly fluorinated form of the DNA-binding peptide GCN4 bZip binds to target DNA sequences with affinity and specificity identical to those of the hydrogenated form, while demonstrating enhanced thermal stability. Molecular dynamics simulation on the fluorinated GCN4-p1d peptide using the Surface Generalized Born implicit solvation model revealed that the coiled-coil binding energy is 55% more favorable upon fluorination. These results suggest that fluorination of hydrophobic substructures in peptides and proteins may provide new means of increasing protein stability, enhancing protein assembly, and strengthening receptor-ligand interactions. PMID- 11258890 TI - Structural and functional analysis of the HIV gp41 core containing an Ile573 to Thr substitution: implications for membrane fusion. AB - The envelope glycoprotein of HIV-1 consists of the surface subunit gp120 and the transmembrane subunit gp41. Binding of gp120 to target cell receptors induces a conformational change in gp41, which then mediates the fusion of viral and cellular membranes. A buried isoleucine (Ile573) in a central trimeric coiled coil within the fusion-active gp41 ectodomain core is thought to favor this conformational activation. The role of Ile573 in determining the structure and function of the gp120-gp41 complex was investigated by mutating this residue to threonine, a nonconservative substitution in HIV-1 that occurs naturally in SIV. While the introduction of Thr573 markedly destabilized the gp41 core, the three dimensional structure of the mutant trimer of hairpins was very similar to that of the wild-type molecule. A new hydrogen-bonding interaction between the buried Thr573 and Thr569 residues appears to allow formation of the trimer-of-hairpins structure at physiological temperature. The mutant envelope glycoprotein expressed in 293T cells and incorporated within pseudotyped virions displayed only a moderate reduction in syncytium-inducing capacity and virus infectivity, respectively. Our results demonstrate that the proper folding of the gp41 core underlies the membrane fusion properties of the gp120-gp41 complex. An understanding of the gp41 activation process may suggest novel strategies for vaccine and antiviral drug development. PMID- 11258891 TI - Backbone dynamics of a module pair from the ligand-binding domain of the LDL receptor. AB - The ligand-binding domain of the LDL receptor consists of seven contiguous LDL-A modules. The fifth of these ligand-binding modules is absolutely required for recognition of both LDL and beta-VLDL particles. A four-residue linker of variable sequence connects each pair of modules, except for modules four and five, which are connected by a 12-residue linker. To provide a more detailed understanding of the structural relationship in a typical pair of functionally important LDL-A repeats of the LDLR, we investigated the backbone dynamics of repeats five (LR5) and six (LR6) alone and in the context of the covalently connected LR5-6 pair. Our results reveal substantial flexibility in the four residue linker connecting the two repeats in the LR5-6 pair. The intrinsic dynamic behavior of each repeat is essentially unchanged when the repeats are covalently connected. These observations indicate that the relative orientation of repeats in LR5-6 is not fixed. Modeled in an extended conformation, the linker can separate LR5 and LR6 by up to 15 A, a distance that would allow substantial freedom of motion of each repeat with respect to the other in the pair. PMID- 11258892 TI - Semisynthetic derivatives of concanamycin A and C, as inhibitors of V- and P-type ATPases: structure-activity investigations and developments of photoaffinity probes. AB - V-type ATPases are inhibited by the plecomacrolides bafilomycin and concanamycin, which exert their inhibitory potential at nanomolar concentrations. In addition, some P-type ATPases are inhibited at micromolar concentrations. We initiated intensive structure-activity investigations with semisynthetic concanamycin derivatives to approach the following two questions: (i) What is the pharmacophor, the structural key element, of the plecomacrolides that leads to their inhibitory potential against V- and P-type ATPases? (ii) Where is the binding site within these two different types of ATPases? In a first step, we examined where chemical modifications (O-acylations, substitutions, eliminations) could be placed without seriously affecting the inhibitory potential of the macrolides. In a second step, we used the knowledge of these structure-activity investigations to introduce traceable elements (fluorescent or radioactive) or nitrene-generating azido or carbene-generating diazirine-groups able to bind the inhibitors to their target covalently. These studies led finally to the synthesis of two photoaffinity probes that were used in labeling experiments with the purified plasma membrane V-type ATPase of Manduca sexta (described in a following paper, Huss, M., Gassel, M., Ingenhorst, G., Drose, S., Zeeck, A., Altendorf, K., Wieczorek, H., manuscript submitted). PMID- 11258893 TI - Interaction of the N-terminal domain of apolipoprotein E4 with heparin. AB - Apolipoprotein E (apoE) is an important lipid-transport protein in human plasma and brain. It has three common isoforms (apoE2, apoE3, and apoE4). ApoE is a major genetic risk factor in heart disease and in neurodegenerative disease, including Alzheimer's disease. The interaction of apoE with heparan sulfate proteoglycans plays an important role in lipoprotein remnant uptake and likely in atherogenesis and Alzheimer's disease. Here we report our studies of the interaction of the N-terminal domain of apoE4 (residues 1-191), which contains the major heparin-binding site, with an enzymatically prepared heparin oligosaccharide. Identified by its high affinity for the N-terminal domain of apoE4, this oligosaccharide was determined to be an octasaccharide of the structure DeltaUAp2S(1-->[4)-alpha-D-GlcNpS6S(1-->4)-alpha-L-IdoAp2S(1-->](3)4) alpha-D-GlcNpS6S by nuclear magnetic resonance spectroscopy, capillary electrophoresis, and polyacrylamide gel electrophoresis. Kinetic analysis of the interaction between the N-terminal apoE4 fragment and immobilized heparin by surface plasmon resonance yielded a K(d) of 150 nM. A similar binding constant (K(d) = 140 nM) was observed for the interaction between immobilized N-terminal apoE4 and the octasaccharide. Isothermal titration calorimetry revealed a K(d) of 75 nM for the interaction of the N-terminal apoE fragment and the octasaccharide with a binding stoichiometry of approximately 1:1. Using previous studies and molecular modeling, we propose a binding site for this octasaccharide in a basic residue-rich region of helix 4 of the N-terminal fragment. From the X-ray crystal structure of the N-terminal apoE4, we predicted that binding of the octasaccharide at this site would result in a change in intrinsic fluorescence. This prediction was confirmed experimentally by an observed increase in fluorescence intensity with octasaccharide binding corresponding to a K(d) of approximately 1 microM. PMID- 11258894 TI - Hepatitis B virus protein pX enhances the monomer assembly pathway of bZIP.DNA complexes. AB - The rapid and correct assembly of dimeric transcription factors on target DNA is essential for accurate transcriptional regulation. Here we ask how a viral accessory factor, hepatitis B virus X protein (pX), influences the rate and identity of the assembly pathway followed by members of the basic region leucine zipper (bZIP) transcription factor family. A combination of fluorescence polarization and fluorescence resonance energy transfer (FRET) experiments demonstrates unequivocally that pX does not increase the concentration of properly folded bZIP dimers in solution. Rather, fluorescence polarization and gel mobility shift experiments reveal that pX interacts directly with the basic spacer segment of the bZIP peptide and stabilizes the complex composed of this monomer and target DNA. By stabilizing the intermediate formed along the monomer assembly pathway but not the one formed along the dimer pathway, pX enhances the equilibrium stability of a bZIP.DNA complex without changing the molecular mechanism used for complexation. Additional experiments reveal that pX decreases the kinetic specificity of certain bZIP proteins. To the extent that it is reflected at the transcriptional level, this loss in specificity could have far reaching consequences for the host cell. PMID- 11258895 TI - Conformational exchange on the microsecond time scale in alpha-helix and beta hairpin peptides measured by 13C NMR transverse relaxation. AB - 13C-NMR relaxation experiments (T(1), T(2), T(1)(rho), and NOE) were performed on selectively enriched residues in two peptides, one hydrophobic staple alpha-helix forming peptide GFSKAELAKARAAKRGGY and one beta-hairpin-forming peptide RGITVNGKTYGR, in water and in water/trifluoroethanol (TFE). Exchange contributions, R(ex), to spin-spin relaxation rates for (13)C(alpha) and (13)C(beta) groups were derived and were ascribed to be mainly due to peptide folding-unfolding. To evaluate the exchange time, tau(ex), from R(ex), the chemical shift difference between folded and unfolded states, Deltadelta, and the populations of these states, p(i), were determined from the temperature dependence of (13)C chemical shifts. For both peptides, values for tau(ex) fell in the 1 micros to 10 micros range. Under conditions where the peptides are most folded (water/TFE, 5 degrees C), tau(ex) values for all residues in each respective peptide were essentially the same, supporting the presence of a global folding-unfolding exchange process. Rounded-up average tau(ex) values were 4 micros for the helix peptide and 9 micros for the hairpin peptide. This 2-3-fold difference in exchange times between helix and hairpin peptides is consistent with that observed for folding-unfolding of other small peptides. PMID- 11258896 TI - Light induces destabilization of photoactive yellow protein. AB - To understand the effect of visible light on the stability of photoactive yellow protein (PYP), urea denaturation experiments were performed with PYP in the dark and with PYP(M) under continuous illumination. The urea concentrations at the midpoint of denaturation were 5.26 +/- 0.29 and 3.77 +/- 0.19 M for PYP and PYP(M), respectively, in 100 mM acetate buffer, and 5.26 +/- 0.24 and 4.11 +/- 0.12 M for PYP and PYP(M), respectively, in 100 mM citrate buffer. The free energy change upon denaturation (DeltaG(D)(H2O)), obtained from the denaturation curve, was 11.0 +/- 0.4 and 7.6 +/- 0.2 kcal/mol for PYP and PYP(M), respectively, in acetate buffer, and 11.5 +/- 0.3 and 7.8 +/- 0.1 kcal/mol for PYP and PYP(M), respectively, in citrate buffer. Even though the DeltaG(D)(H2O) value for PYP(M) is almost identical in the two buffer systems, the urea concentration at the midpoint of denaturation is lower in acetate buffer than in citrate buffer. Although their CD spectra indicate that the protein conformations of the denatured states of PYP and PYP(M) are indistinguishable, the configurations of the chromophores in their denatured structures are not necessarily identical. Both denatured states are interconvertible through PYP and PYP(M). Therefore, the free energy difference between PYP and PYP(M) is 3.4-3.7 kcal/mol for the protein moiety, plus the additional contribution from the difference in configuration of the chromophore. PMID- 11258897 TI - Biological activities of glucagon-like peptide-1 analogues in vitro and in vivo. AB - Studies support a role for glucagon-like peptide 1 (GLP-1) as a potential treatment for diabetes. However, since GLP-1 is rapidly degraded in the circulation by cleavage at Ala(2), its clinical application is limited. Hence, understanding the structure-activity of GLP-1 may lead to the development of more stable and potent analogues. In this study, we investigated GLP-1 analogues including those with N-, C-, and midchain modifications and a series of secretin class chimeric peptides. Peptides were analyzed in CHO cells expressing the hGLP 1 receptor (R7 cells), and in vivo oral glucose tolerance tests (OGTTs) were performed after injection of the peptides in normal and diabetic (db/db) mice. [D Ala(2)]GLP-1 and [Gly(2)]GLP-1 showed normal or relatively lower receptor binding and cAMP activation but exerted markedly enhanced abilities to reduce the glycemic response to an OGTT in vivo. Improved biological effectiveness of [D Ala(2)]GLP-1 was also observed in diabetic db/db mice. Similarly, improved biological activity of acetyl- and hexenoic-His(1)-GLP-1, glucagon((1-5)-, glucagon((1-10))-, PACAP(1-5)-, VIP(1-5)-, and secretin((1-10))-GLP-1 was observed, despite normal or lower receptor binding and activation in vitro. [Ala(8/11/12/16)] substitutions also increased biological activity in vivo over wtGLP-1, while C-terminal truncation of 4-12 amino acids abolished receptor binding and biological activity. All other modified peptides examined showed normal or decreased activity in vitro and in vivo. These results indicate that specific N- and midchain modifications to GLP-1 can increase its potency in vivo. Specifically, linkage of acyl-chains to the alpha-amino group of His(1) and replacement of Ala(2) result in significantly increased biological effects of GLP 1 in vivo, likely due to decreased degradation rather than enhanced receptor interactions. Replacement of certain residues in the midchain of GLP-1 also augment biological activity. PMID- 11258898 TI - Thyroid hormone promotes serine phosphorylation of p53 by mitogen-activated protein kinase. AB - L-Thyroxine (T(4)) nongenomically promotes association of mitogen-activated protein kinase (MAPK) and thyroid hormone receptor TRbeta1 (TR) in the cell nucleus, leading to serine phosphorylation of the receptor. The oncogene suppressor protein, p53, is serine phosphorylated by several kinases and is known to interact with TRbeta1. We studied whether association of p53 and TR is modulated by T(4) and involves serine phosphorylation of p53 by MAPK. TR-replete 293T human kidney cells were incubated with a physiological concentration of T(4) for 10-90 min. Nuclear fractions were immunoprecipitated and the resulting proteins separated and immunoblotted for co-immunoprecipitated proteins. Activated MAPK immunoprecipitates of nuclei from T(4)-treated cells accumulated p53 in a time-dependent manner; T(4) and T(4)-agarose were more effective than T(3). T(4)-induced nuclear complexing of p53 and MAPK was inhibited by PD 98059 (PD) and U0126, two MAPK kinase (MEK) inhibitors, and was absent in cells treated with MEK antisense oligonucleotide and in dominant negative Ras cells. T(4) also caused nuclear co-immunoprecipitation of TRbeta1 and p53, an effect also inhibited by PD. Nuclear complexing of p53 and MAPK also occurred in HeLa cells, which lack functional TR. Constitutively activated MAPK caused phosphorylation of a recombinant p53-GST fusion protein in vitro; thus, p53 is a substrate for MAPK. An indicator of p53 transcriptional activity, accumulation of the immediate-early gene product, c-Jun, was inhibited by T(4). This T(4) effect was reversed by PD, indicating that the transcriptional activity of p53 was altered by T(4)-directed MAPK-p53 interaction. PMID- 11258899 TI - Difference in molecular structure of rod and cone visual pigments studied by Fourier transform infrared spectroscopy. AB - To investigate the local structure that causes the differences in molecular properties between rod and cone visual pigments, we have measured the difference infrared spectra between chicken green and its photoproduct at 77 K and compared them with those from bovine and chicken rhodopsins. In contrast to the similarity of the vibrational bands of the chromophore, those of the protein part were notably different between chicken green and the rhodopsins. Like the rhodopsins, chicken green has an aspartic acid at position 83 (D83) but exhibited no signals due to the protonated carboxyl of D83 in the C=O stretching region, suggesting that the molecular contact between D83 and G120 through water molecule evidenced in bovine rhodopsin is absent in chicken green. A pair of positive and negative bands due to the peptide backbone (amide I) was prominent in chicken green, while the rhodopsins exhibited only small bands in this region. Furthermore, chicken green exhibited characteristic paired bands around 1480 cm(-1), which were identified as the imide bands of P189 using site-directed mutagenesis. P189, situated in the putative second extracellular loop, is conserved in all the known cone visual pigments but not in rhodopsins. Thus, some region of the second extracellular loop including P189 is situated near the chromophore and changes its environment upon formation of the batho-intermediate. The results noted above indicate that differences in the protein parts between chicken green and the rhodopsins alter the changes seen in the protein upon photoisomerization of the chromophore. Some of these changes appear to be the pathway from the chromophore to cytoplasmic surface of the pigment and thus could affect the activation process of transducin. PMID- 11258900 TI - Polyphosphoprotein from the adhesive pads of Mytilus edulis. AB - Achieving a satisfactory biochemical explanation for the opportunistic underwater adhesion of marine invertebrates such as mussels and barnacles requires a detailed characterization of proteins extracted from holdfast structures produced by these organisms. Mefp-5 is an adhesive protein derived from the foot of the common mussel, Mytilus edulis, and deposited into the byssal attachment pads. Purification and primary structure of mefp-5 was determined by peptide mapping and cDNA sequencing. The protein is 74 residues long and has a mass of about 9500 Da. Mefp-5 composition shows a strong amino acid bias: aromatic amino acids, lysine, and glycine represent 65 mol % of the composition. More than a third of all the residues in the protein are posttranslationally modified by hydroxylation or phosphorylation. The conversion of tyrosine to 3, 4-dihydroxyphenyl-L-alanine (DOPA) and serine to O-phosphoserine accounts for the hydroxylation and phosphorylation, respectively. Neither modification is complete since variations in the extent of phosphorylation and hydroxylation can be detected by mass spectrometry. More than 75% of the DOPA is adjacent to basic residues, e.g., Lys DOPA and DOPA-Lys. Phosphoserine occurs in sequences strikingly reminiscent of acidic mineral-binding motifs that appear in statherin, osteopontin, and others. This may be an adaptation for adhesion to the most common substrata for mussels, i.e., calcareous materials. PMID- 11258901 TI - Alkaline denaturation of the light-harvesting complex II from the purple bacterium Ectothiorhodospira sp.: kinetic evidence of the existence of the 780 nm upper exciton component of the B850 bacteriochlorophylls. AB - The light-harvesting complex II of the purple bacteria has two strong near infrared electronic absorption bands around 800 (B800) and 850 (B850) nm, arising from the Q(y)() transitions of the bacteriochlorophyll a. In the present work, high concentrations of NaOH were used to study the destabilization of the complex of the Ectothiorhodospira sp. The majority of the bacteriochlorophylls were monomerized within 90 min of treatment. However, the kinetic patterns of the two near-infrared bands were remarkably different. After an instantaneous blue shift from 853 to 828 nm, B850 showed a first-order monomerization with a rate constant of -0.016 min(-1). This instantaneous blue shift was previously attributed to the deprotonation of a lysine and was independent of the monomerization process. The observed native B800 is in fact composed of two bands, one at 796 nm and the other at 780 nm. The band absorbing at 780 nm red shifted also instantaneously to 786-788 nm and then disappeared in a first-order process as B850. The other band absorbing at 796 nm has a two-step process of monomerization; after a rapid conversion a slower first-order process occurred with a rate constant of -0.025 min(-1). The similarity between the kinetic behaviors of B850 and the 780 nm band indicated a strong relationship between these two bands. Our interpretation of the results considers the 780 nm band as the upper exciton component of the B850 bacteriochlorophylls. PMID- 11258902 TI - Binding of XPA and RPA to damaged DNA investigated by fluorescence anisotropy. AB - The proteins XPA and RPA are assumed to be involved in primary damage recognition of global genome nucleotide excision repair. XPA as well as RPA have been each reported to specifically bind DNA lesions, and ternary complex formation with damaged DNA has also been shown. We employed fluorescence anisotropy measurements to study the DNA-binding properties of XPA and RPA under true equilibrium conditions using damaged DNA probes carrying a terminal fluorescein modification as a reporter. XPA binds with low affinity and in a strongly salt-dependent manner to DNA containing a 1,3-d(GTG) intrastrand adduct of the anticancer drug cisplatin or a 6-nt mismatch (K(D) = 400 nM) with 3-fold preference for damaged vs undamaged DNA. At near physiological salt conditions binding is very weak (K(D) > 2 microM). RPA binds to damaged DNA probes with dissociation constants in the range of 20 nM and a nearly 15-fold preference over undamaged DNA. The presence of a cisplatin modification weakens the affinity of RPA for single stranded DNA by more than 1 order of magnitude indicating that binding to the lesion itself is not a driving force in damage recognition. Our fluorescence anisotropy assays also show that the presence of XPA does not enhance the affinity of RPA for damaged DNA although both proteins interact. In contrast, cooperative binding of XPA and RPA is observed in EMSA. Our results point to a damage-sensing function of the XPA-RPA complex with RPA mediating the important DNA contacts. PMID- 11258903 TI - Thermodynamic basis for sequence-specific recognition of ssDNA by an autoantibody. AB - 11F8 is a sequence-specific DNA binding monoclonal autoantibody previously isolated from an autoimmune lupus-prone mouse [Stevens, S. Y., and Glick, G. D. (1999) Biochemistry 38, 560-568]. This antibody, like many other lupus anti-DNAs, localizes to kidney tissue and eventually leads to renal damage through a process that first involves the binding of DNA antigens. A series of experiments were conducted to investigate the thermodynamic and structural basis by which this antibody discriminates between specific, noncognate, and nonspecific sequences. Sequence-specific binding occurs with a minimal dependence on the polyelectrolyte effect along with a favorable binding enthalpy reflecting the presence of base stacking and contacts to DNA bases. This favorable binding enthalpy apparently is derived from desolvation at the binding interface and is consistent with recent models of the nonclassical hydrophobic effect. Noncognate recognition is also driven by the nonclassical hydrophobic effect, but is accompanied by highly unfavorable entropies that are responsible for reduced affinity relative to the high-affinity consensus sequence. Nonspecific recognition is driven completely by the polyelectrolyte effect involving extensive electrostatic interactions with the phosphate backbone. Collectively, the data demonstrate the ability of 11F8 to adapt its mode of binding to the available DNA surface and provide a thermodynamic model for sequence-specific recognition of single-stranded DNA. The salient features of this model employ the paradigms invoked to explain protein.dsDNA, protein.RNA, and antibody.antigen binding. PMID- 11258904 TI - Hierarchy of DNA damage recognition in Escherichia coli nucleotide excision repair. AB - DNA damage recognition plays a central role in nucleotide excision repair (NER). Here we present evidence that in Escherichia coli NER, DNA damage is recognized through at least two separate but successive steps, with the first focused on distortions from the normal structure of the DNA double helix (initial recognition) and the second specifically recognizing the type of DNA base modifications (second recognition), after an initial local separation of the DNA strands. DNA substrates containing stereoisomeric (+)- or (-)-trans- or (+)- or ( )-cis-BPDE-N(2)-dG lesions in DNA duplexes of known conformations were incised by UvrABC nuclease with efficiencies varying by up to 3-fold. However, these stereoisomeric adducts, when positioned in an opened, single-stranded DNA region, were all incised with similar efficiencies and with enhanced rates (by factors of 1.4-6). These bubble substrates were also equally and efficiently incised by UvrBC nuclease without UvrA. Furthermore, removal of the Watson-Crick partner cytosine residue (leaving an abasic site) in the complementary strand opposite a (+)-cis-BPDE-N(2)-dG lesion led to a significant reduction in both the binding of UvrA and the incision efficiency of UvrABC by a factor of 5. These data suggest that E. coli NER features a dynamic two-stage recognition mechanism. PMID- 11258905 TI - Multiconnection of identical zinc finger: implication for DNA binding affinity and unit modulation of the three zinc finger domain. AB - Cys(2)-His(2)-type zinc finger proteins have a tandemly repeated array structure consisting of independent finger modules. They are expected to elevate the DNA binding affinity and specificity by increasing the number of finger modules. To investigate the relation between the number and the DNA binding affinity of the zinc finger, we have designed the two- to four-finger peptides by connecting the central zinc finger (finger 2) of Sp1 with the canonical linker sequence, Thr-Gly Glu-Lys-Pro. Gel mobility shift assays reveal that the cognate three- and four finger peptides, Sp1(zf222) and Sp1(zf2222), strongly bind to the predicted target sequences, but the two-finger peptide, Sp1(zf22), does not. Of special interest is the fact that the dissociation constant for Sp1(zf2222) binding to the target DNA is comparable to that for Sp1(zf222). The methylation interference, DNase I and hydroxyl radical footprintings, and circular permutation analyses demonstrate that Sp1(zf2222) binds to its target site with three successive zinc fingers and the binding of the fourth zinc finger is inhibited by DNA bending induced by the binding of the three-finger domain. The present results strongly indicate that the zinc finger protein binds to DNA by the three-finger domain as one binding unit. In addition, this information provides the basis for the design of a novel multifinger protein with high affinity and specificity for long DNA sequences, such as chromosomal DNAs. PMID- 11258906 TI - A RecA homologue in Ustilago maydis that is distinct and evolutionarily distant from Rad51 actively promotes DNA pairing reactions in the absence of auxiliary factors. AB - Two RecA homologues have been identified to date in Ustilago maydis. One is orthologous to Rad51 while the other, Rec2, is structurally quite divergent and evolutionarily distant. DNA repair and recombination proficiency in U. maydis requires both Rec2 and Rad51. Here we have examined biochemical activities of Rec2 protein purified after overexpression of the cloned gene. Rec2 requires DNA as a cofactor to hydrolyze ATP and depends on ATP to promote homologous pairing and DNA strand exchange. ATPgammaS was found to substitute for ATP in all pairing reactions examined. With superhelical DNA and a homologous single-stranded oligonucleotide as substrates, Rec2 actively promoted formation and dissociation of D-loops. When an RNA oligonucleotide was substituted it was found that R-loops could also be formed and utilized as primer/template for limited DNA synthesis. In DNA strand exchange reactions using oligonucleotides, we found that Rec2 exhibited a pairing bias that is opposite that of RecA. Single-stranded oligonucleotides were activated for DNA strand exchange when attached as tails protruding from a duplex sequence due to enhanced binding of Rec2. The results indicate that Rec2 is competent, and in certain ways even better than Rad51, in the ability to provide the fundamental DNA pairing activity necessary for recombinational repair. We propose that the emerging paradigm for homologous recombination featuring Rad51 as the essential catalytic component for strand exchange may not be universal in eukaryotes. PMID- 11258907 TI - The role of copper in topa quinone biogenesis and catalysis, as probed by azide inhibition of a copper amine oxidase from yeast. AB - All known copper amine oxidases (CAOs) contain 2,4,5-trihydroxyphenylalanine quinone (TPQ) as a redox cofactor. TPQ is derived posttranslationally from a specific tyrosine residue within the protein itself, and is utilized by the enzyme to oxidize amines to aldehydes. Several oxidative mechanisms for both turnover and the biogenesis of the cofactor have been proposed in recent years, which differ mainly in the nature of the interaction of oxygen with the enzyme. In this study, azide is used to probe the role of copper in catalysis and biogenesis, especially with respect to potential interactions between the metal and oxygen. During turnover, it is found that azide is a noncompetitive inhibitor with respect to O(2), most consistent mechanistically with oxygen binding off the metal prior to reaction. During biogenesis, it is found that azide likely prohibits ligation of the precursor tyrosine to the copper, thus preventing the formation of this key intermediate. This result is consistent with previous proposals, where the copper-tyrosine unit is the species that undergoes reaction with O(2). In addition, it is found that oxygen consumption is kinetically uncoupled from TPQ formation; this leads to an expanded kinetic model for biogenesis, with important implications for previous results. PMID- 11258908 TI - Regiospecificity assignment for the reaction of kanamycin nucleotidyltransferase from Staphylococcus aureus. AB - Aminoglycoside nucleotidyltransferases catalyze the transfer of a nucleoside monophosphoryl group from a nucleotide to a hydroxyl group of an aminoglycoside antibiotic. Kanamycin nucleotidyltransferase [ANT (4',4' ')-I] from Staphylococcus aureus confers resistance to numerous aminoglycosides with a 4' or 4' ' hydroxyl group in the equatorial position. The synthesis of m-nitrobenzyl triphosphate, a new substrate of kanamycin nucleotidyltransferase, is reported. The kanamycin nucleotidyltransferase catalyzed reaction of kanamycin A with m nitrobenzyl triphosphate is 2 orders of magnitude slower than that with ATP. The MALDI-TOF spectra of the purified products of both reactions revealed that kanamycin A was modified only at one position. The regiospecificity of the reaction catalyzed by kanamycin nucleotidyltransferase of kanamycin A with either ATP or m-nitrobenzyl triphosphate was determined directly by one- and two dimensional hetero- and homonuclear NMR techniques. The site of the modification was unambiguously assigned to the 4' hydroxyl of kanamycin A; thus, the products formed are 4'-(adenosine-5'-phosphoryl)-kanamycin A and 4'-(m-nitrobenzyl phosphoryl)-kanamycin A. This eliminates the uncertainty concerning the point of modification since this could not be determined from the crystal structure of the enzyme with bound MgAMPCPP and kanamycin A [Pedersen, L. C., Benninig, M. M., and Holden, H. M. (1995) Biochemistry 34, 13305-13311]. PMID- 11258909 TI - Characterization of the transition-state structure of the reaction of kanamycin nucleotidyltransferase by heavy-atom kinetic isotope effects. AB - The transition-state structure for the reaction catalyzed by kanamycin nucleotidyltransferase has been determined from kinetic isotope effects. The primary (18)O isotope effects at pH 5.7 (close to the optimum pH) and at pH 7.7 (away from the optimum pH) are respectively 1.016 +/- 0.003 and 1.014 +/- 0.002. Secondary (18)O isotope effects of 1.0033 +/- 0.0004 and 1.0024 +/- 0.0002 for both nonbridge oxygen atoms were measured respectively at pH 5.7 and 7.7. These isotope effects are consistent with a concerted reaction with a slightly associative transition-state structure. PMID- 11258910 TI - Glycosylated polyproline II rods with kinks as a structural motif in plant hydroxyproline-rich glycoproteins. AB - Hydroxyproline-rich glycoproteins (HRGPs) are the major proteinaceous components of higher plant walls and the predominant components of the cell wall of the green alga Chlamydomonas reinhardtii. The GP1 protein, an HRGP of the C. reinhardtii wall, is shown to adopt a polyproline II helical configuration and to carry a complex array of arabinogalactoside residues, many branched, which are necessary to stabilize the helical conformation. The deduced GP1 amino acid sequence displays two Ser-Pro-rich domains, one with a repeating (SP)(x)() motif and the other with a repeating (PPSPX)(x)() motif. A second cloned gene a2 also carries the PPSPX repeat, defining a novel gene family in this lineage. The SP repeat domains of GP1 form a 100-nm shaft with a flexible kink 28 nm from the head. The gp1 gene encodes a PPPPPRPPFPANTPM sequence at the calculated kink position, generating the proposal that this insert interrupts the PPII helix, with the resultant kink exposing amino acids necessary for GP1 to bind to partner molecules. It is proposed that similar kinks in the higher plant HRGPs called extensins may play a comparable role in wall assembly. PMID- 11258911 TI - Structural changes of mitochondrial creatine kinase upon binding of ADP, ATP, or Pi, observed by reaction-induced infrared difference spectra. AB - Structural modifications of rabbit heart mitochondrial creatine kinase induced by the binding of its nucleotide substrates and Pi were investigated. Reaction induced difference spectra (RIDS), resulting from the difference between infrared spectra recorded before and after the photorelease of a caged ligand, allow us to detect very small variations in protein structure. Our results indicated that the protein secondary structure remained relatively stable during nucleotide binding. Indeed, this binding to creatine kinase affected only a few amino acids, and caused small peptide backbone deformations and alterations of the carbonyl side chains of aspartate or glutamate, reflecting modifications within preexisting elements rather than a net change in secondary structure. Nonetheless, MgADP and MgATP RIDS were distinct, whereas the MgPi RIDS presented some similarities with the MgATP one. The difference between MgADP and MgATP RIDS could reflect a distinct configuration of the two metal-nucleotide complexes inducing a different positioning and/or a distinct binding mode to the creatine kinase active site. Comparison of the MgATP and MgPi RIDS suggests that Pi binding took place at the same binding site as the gamma-phosphoryl group of ATP. Thus, the difference between MgADP and MgATP RIDS would mainly be due to the effect of the gamma-P of ATP. The differences observed when comparing the RIDS resulting from the binding of nucleotides to octameric mitochondrial creatine kinase or dimeric cytosolic isoform could reflect the distinct oligomerization states and physicochemical or kinetic properties of the two isoenzymes. PMID- 11258912 TI - Absorption changes induced by the binding of triazines to the QB pocket in reaction centers of Rhodobacter capsulatus. AB - Inhibitors which block electron transfer from the primary (Q(A)) to the secondary (Q(B)) quinone of the bacterial reaction center are competing with the pool ubiquinones for binding at the Q(B) pocket. Due to the much greater stability of the semiquinone state Q(B)(-) compared with fully oxidized or reduced quinone, a displacement of the inhibitors takes place after one flash from state Q(A)(-)I to state Q(A)Q(B)(-). This process can be monitored from near-IR absorption changes which reflect local absorption shifts specific to Q(A)(-) and Q(B)(-). An anomalous behavior was observed when using triazines in chromatophores of R. capsulatus: the IR absorption change reflecting the formation of Q(B)(-) after one flash was absent. A normal transient decay of this signal was, however, triggered by a second flash, followed by a rapid return to the baseline. We show that this phenomenon is due to an absorption change induced by inhibitor binding (thus present in the dark baseline), with a spectrum close to that of Q(B)(-), so that the Q(B)(-) changes are canceled out during the inhibitor displacement process. On the second flash, one monitors the destruction of the semiquinone, leading transiently to the Q(A)Q(B) state, followed by inhibitor rebinding. This allows a direct measurement of the binding kinetics. This behavior was observed both in chromatophores and in isolated reaction centers from R. capsulatus, but not in R. sphaeroides. PMID- 11258913 TI - Equilibrium binding studies of a tryptophan-shifted mutant of phosphofructokinase from Bacillus stearothermophilus. AB - A tryptophan-shifted mutant of phosphofructokinase (PFK) from Bacillus stearothermophilus has been constructed. This mutant, which is functionally similar to wild-type, provides the opportunity to examine the allosteric properties of PFK under equilibrium conditions. The unique fluorescence properties of the tryptophan-shifted mutant enzyme, W179F/F230W, have been utilized to deduce the thermodynamics of ligand binding and the allosteric perturbations in the absence of catalytic turnover. Specifically, phospho(enol)pyruvate (PEP) and MgADP binding to the mutant PFK can be directly observed using tryptophan fluorescence, and dissociation constants for these ligands have been measured to be equal to 2.71 +/- 0.04 and 90.4 +/- 3.5 microM, respectively. In addition, the homotropic couplings for the allosteric ligands have been assessed for the first time. PEP binds cooperatively with a Hill number of 2.9 +/- 0.3, while MgADP binding is not cooperative. The equilibrium couplings between these ligands and the substrate fructose 6-phosphate (Fru-6-P) have also been determined and follow the same trends with temperature observed under steady state kinetic assay conditions using wild-type PFK, indicating that the presence of bound MgATP has little influence on the allosteric interactions. Like wild type PFK, the coupling free energies for the mutant result from largely compensating enthalpy and entropy components at 25 degrees C. Furthermore, the sign of each coupling free energy, which signifies the nature of the allosteric effect, is opposite that of the enthalpy contribution and is therefore due to the larger absolute value of the associated entropy change. This characteristic stands in direct contrast to the thermodynamic basis of the allosteric response in the homologous PFK from E. coli in which the sign of the coupling free energy is established by the sign of the coupling enthalpy. PMID- 11258914 TI - Multivalent protein-carbohydrate interactions. A new paradigm for supermolecular assembly and signal transduction. AB - Many biological recognition processes involve the binding and clustering of ligand-receptor complexes and concomitant signal transduction events. Such interactions have recently been observed in human T cells in which binding and cross-linking of specific glycoprotein counter-receptors on the surface of the cells by an endogenous bivalent carbohydrate binding protein (galectin-1) leads to apoptosis [Pace, K. E., et al. (1999) J. Immunol. 163, 3801-3811]. Importantly, different counter-receptors associated with specific phosphatase or kinase activities were shown to form separate clusters on the surface of the cells as a result of galectin-1 binding to the carbohydrate moieties of the respective glycoproteins. This suggests that the unique separation and organization of signaling molecules that results from galectin-1 binding is involved in delivering the signal to die. The ability of galectin-1 to induce the separation of specific glycoprotein receptors was modeled on the basis of molecular and structural studies of the binding of multivalent carbohydrates to lectins that result in the formation of specific two- and three-dimensional cross linked lattices. These latter studies have been recently highlighted by X-ray crystallographic results showing that a single tetravalent lectin forms distinct cross-linked complexes with four different bivalent oligosaccharides [Olsen, L. R., et al. (1997) Biochemistry 36, 15073-15080]. In this report, binding and cross-linking of multivalent carbohydrates with multivalent lectins is shown to be a new paradigm for supermolecular assembly and signal transduction in biological systems. PMID- 11258915 TI - The antibacterial peptide pyrrhocoricin inhibits the ATPase actions of DnaK and prevents chaperone-assisted protein folding. AB - Recently, we documented that the short, proline-rich antibacterial peptides pyrrhocoricin, drosocin, and apidaecin interact with the bacterial heat shock protein DnaK, and peptide binding to DnaK can be correlated with antimicrobial activity. In the current report we studied the mechanism of action of these peptides and their binding sites to Escherichia coli DnaK. Biologically active pyrrhocoricin made of L-amino acids diminished the ATPase activity of recombinant DnaK. The inactive D-pyrrhocoricin analogue and the membrane-active antibacterial peptide cecropin A or magainin 2 failed to inhibit the DnaK-mediated phosphate release from adenosine 5'-triphosphate (ATP). The effect of pyrrhocoricin on DnaK's other significant biological function, the refolding of misfolded proteins, was studied by assaying the alkaline phosphatase and beta-galactosidase activity of live bacteria. Remarkably, both enzyme activities were reduced upon incubation with L-pyrrhocoricin or drosocin. D-Pyrrhocoricin, magainin 2, or buforin II, an antimicrobial peptide involved in binding to bacterial nucleic acids, had only negligible effect. According to fluorescence polarization and dot blot analysis of synthetic DnaK fragments and labeled pyrrhocoricin analogues, pyrrhocoricin bound with a K(d) of 50.8 microM to the hinge region around the C terminal helices D and E, at the vicinity of amino acids 583 and 615. Pyrrhocoricin binding was not observed to the homologous DnaK fragment of Staphylococcus aureus, a pyrrhocoricin nonresponsive strain. In line with the lack of ATPase inhibition, drosocin binding appears to be slightly shifted toward the D helix. Our data suggest that drosocin and pyrrhocoricin binding prevents the frequent opening and closing of the multihelical lid over the peptide-binding pocket of DnaK, permanently closes the cavity, and inhibits chaperone-assisted protein folding. The biochemical results were strongly supported by molecular modeling of DnaK-pyrrhocoricin interactions. Due to the prominent sequence variations of procaryotic and eucaryotic DnaK molecules in the multihelical lid region, our findings pave the road for the design of strain-specific antibacterial peptides and peptidomimetics. Far-fetched applications of the species-specific inhibition of chaperone-assisted protein folding include the control of not only bacteria but also fungi, parasites, insects, and perhaps rodents. PMID- 11258916 TI - A helical turn motif in Mss4 is a critical determinant of Rab binding and nucleotide release. AB - Monomeric Rab GTPases function as ubiquitous regulators of intracellular membrane trafficking. Mss4, an evolutionarily conserved Rab accessory factor, promotes nucleotide release from exocytic but not endocytic Rab GTPases. Here we describe the results of a high-resolution crystallographic and mutational analysis of Mss4. The 1.65 A crystal structure of Mss4 reveals a network of direct and water mediated interactions that stabilize a partially exposed structural subdomain derived from four highly conserved but nonconsecutive sequence elements. The conserved subdomain contains the invariant cysteine residues required for Zn2+ binding as well as the residues implicated in the interaction with Rab GTPases. A strictly conserved DPhiPhi motif, consisting of an invariant aspartic acid residue (Asp 73) followed by two bulky hydrophobic residues (Met 74 and Phe 75), encodes a prominently exposed 3(10) helical turn in which the backbone is well ordered but the side chains of the conserved residues are highly exposed and do not engage in intramolecular interactions. Substitution of any of these residues with alanine dramatically impairs nucleotide release activity toward Rab3A, indicating that the DPhiPhi motif is a critical element of the Rab interaction epitope. In particular, mutation of Phe 75 results in a defect as severe as that observed for mutation of Asp 96, which is located near the zinc binding site at the opposite end of the conserved subdomain. Despite severe defects, however, none of the mutant proteins is catalytically dead. Taken together, the results suggest a concerted mechanism in which distal elements of the conserved Rab interaction epitope cooperatively facilitate nucleotide release. PMID- 11258917 TI - Ras catalyzes GTP hydrolysis by shifting negative charges from gamma- to beta phosphate as revealed by time-resolved FTIR difference spectroscopy. AB - FTIR difference spectroscopy has been used to determine the molecular GTPase mechanism of the small GTP binding protein Ras at the atomic level. The reaction was initiated by the photolysis of caged GTP bound to Ras. The addition of catalytic amounts of the GTPase activating protein (GAP) reduces the measuring time by 2 orders of magnitude but has no influence on the spectra as compared to the intrinsic reaction. The reduced measuring time improves the quality of the data significantly as compared to previously published data [Cepus, V., Scheidig, A., Goody, R. S., and Gerwert, K. (1998) Biochemistry 37, 10263-10271]. The phosphate vibrations are assigned using 18O-labeled caged GTP. In general, there is excellent agreement with the results of Cepus et al., except in the nu(a)(alpha-PO2-) vibration assignments. The assignments reveal that binding of GTP to Ras induces vibrational uncoupling into mainly individual vibrations of the alpha-, beta-, and gamma-phosphate groups. In contrast, for unbound GTP, the phosphate vibrations are highly coupled and the corresponding absorption bands are broader. This result indicates that binding to Ras forces the flexible GTP molecule into a strained conformation and induces a specific charge distribution different from that in the unbound case. The binding causes an unusual frequency downshift of the GTP beta-PO2- phosphate vibration, whereas the alpha-PO2- and gamma-PO3(2-) phosphate vibrations shift to higher wavenumbers. The frequency downshift indicates a lowering of the bond order of the nonbridged P-O bonds of the beta-phosphate group of GTP and GDP. The bond order changes can be explained by a shift of negative charges from the gamma- to the beta-oxygens. Thereby, the GTP charge distribution becomes more like that in GDP. The charge shift appears to be a key factor contributing to catalysis by Ras in addition to the correct positioning of the attacking water. Ras appears to increase the negative charge at the pro-R beta-oxygen mainly by interaction of Mg(2+) and at the pro-S beta oxygen mainly by interactions of the backbone NHs of Lys 16, Gly 15, and Val 14. The correct positioning of the backbone NHs of Lys 16, Gly 15, and Val 14, and especially the Lys 16 side chain, of the structural highly conserved phosphate binding loop relative to beta-phosphate therefore seems to be important for the catalysis provided by Ras. PMID- 11258918 TI - The "catalytic" triad of isocitrate dehydrogenase kinase/phosphatase from E. coli and its relationship with that found in eukaryotic protein kinases. AB - The isocitrate dehydrogenase kinase/phosphatase (IDHK/P) of E. coli is a bifunctional enzyme responsible for the reversible phosphorylation of isocitrate dehydrogenase (IDH) on a seryl residue. As such, it belongs to the serine/threonine protein kinase family. However, only a very limited homology with the well-characterized eukaryotic members of that family was identified so far in its primary structure. In this report, a new region of amino acids including three putative residues involved in the kinase activity of IDHK/P was identified by sequence comparison with eukaryotic protein kinases. In IDHK/P, these residues are Asp-371, Asn-377, and Asp-403. Their counterpart eukaryotic residues have been shown to be involved in either catalysis (former residue) or magnesium binding (the two latter residues). Site-directed mutagenesis was performed on these three IDHK/P residues, and also on the Glu-439 residue equivalent to that of the Ala-Pro-Glu motif found in the eukaryotic protein kinases. Mutations of Asp-371 into either Ala, Glu, or Gln residues drastically lowered the yield and the quality of the purification. Nevertheless, the recovered mutant enzymes were barely able to phosphorylate IDH either in vitro or after expression in an aceK (-) mutant strain. In contrast, mutation of either Asn-377, Asp-403, or Glu-439 into an Ala residue altered neither the yield of purification nor the maximal phosphorylating capacity of the enzyme. However, when IDH was phosphorylated in the presence of increasing concentrations of magnesium ions, the two former mutants displayed a much lower affinity for this cation, with a K(m) value of 0.6 or 0.8 mM, respectively, as compared to 0.1 mM for the wild-type enzyme. On the other hand, the Glu439Ala mutant has an affinity for magnesium essentially unaffected. Therefore, and in contrast to the current opinion, our results suggest that the catalytic mechanism of IDHK/P exhibits some similarities with that found in the eukaryotic members of the protein kinase family. PMID- 11258919 TI - Mutagenesis of two acidic active site residues in human muscle creatine kinase: implications for the catalytic mechanism. AB - Creatine kinase (CK) catalyzes the reversible phosphorylation of the guanidine substrate, creatine, by MgATP. Although several X-ray crystal structures of various isoforms of creatine kinase have been published, the detailed catalytic mechanism remains unresolved. A crystal structure of the CK homologue, arginine kinase (AK), complexed with the transition-state analogue (arginine-nitrate-ADP), has revealed two carboxylate amino acid residues (Glu225 and Glu314) within 2.8 A of the proposed transphosphorylation site. These two residues are the putative catalytic groups that may promote nucleophilic attack by the guanidine amino group on the gamma-phosphate of ATP. From primary sequence alignments of arginine kinases and creatine kinases, we have identified two homologous creatine kinase acidic amino acid residues (Glu232 and Asp326), and these were targeted for examination of their potential roles in the CK mechanism. Using site-directed mutagenesis, we have made several substitutions at these two positions. The results indicate that of these two residues the Glu232 is the likely catalytic residue while Asp326 likely performs a role in properly aligning substrates for catalysis. PMID- 11258920 TI - Hemoglobin of the Antarctic fishes Trematomus bernacchii and Trematomus newnesi: structural basis for the increased stability of the liganded tetramer relative to human hemoglobin. AB - Hemoglobins extracted from fishes that live in temperate waters show little or no dissociation even in the liganded form, unlike human hemoglobin (HbA). To establish whether cold adaptation influences the tendency to dissociate, the dimer-tetramer association constants (L(2,4)) of the carbonmonoxy derivatives of representative hemoglobins from two Antarctic fishes, Trematomus newnesi (Hb1Tn) and Trematomus bernacchii (Hb1Tb), were determined by analytical ultracentrifugation as a function of pH in the range 6.0-8.6 and compared to HbA. HbA is more dissociated than fish hemoglobins at all pH values and in particular at pH 6.0. In contrast, both fish hemoglobins are mostly tetrameric over the whole pH range studied. The extent of hydrophobic surface area buried at the alpha(1)beta(2) interface upon association of dimers into tetramers and the number of hydrogen bonds formed are currently thought to play a major role in the stabilization of the hemoglobin tetramer. These contributions were derived from the X-ray structures of the three hemoglobins under study and found to be in good agreement with the experimentally determined L(2,4) values. pH affects oxygen binding of T. bernacchii and T. newnesi hemoglobins in a different fashion. The lack of a pH effect on the dissociation of the liganded proteins supports the proposal that the structural basis of such effects resides in the T (unliganded) structure rather than in the R (liganded) one. PMID- 11258921 TI - Large-scale domain movements and hydration structure changes in the active-site cleft of unligated glutamate dehydrogenase from Thermococcus profundus studied by cryogenic X-ray crystal structure analysis and small-angle X-ray scattering. AB - Here we describe the large-scale domain movements and hydration structure changes in the active-site cleft of unligated glutamate dehydrogenase. Glutamate dehydrogenase from Thermococcus profundus is composed of six identical subunits of M(r) 46K, each with two distinct domains of roughly equal size separated by a large active-site cleft. The enzyme in the unligated state was crystallized so that one hexamer occupied a crystallographic asymmetric unit, and the crystal structure of the hexamer was solved and refined at a resolution of 2.25 A with a crystallographic R-factor of 0.190. In that structure, the six subunits displayed significant conformational variations with respect to the orientations of the two domains. The variation was most likely explained as a hinge-bending motion caused by small changes in the main chain torsion angle of the residue composing a loop connecting the two domains. Small-angle X-ray scattering profiles both at 293 and 338 K suggested that the apparent molecular size of the hexamer was slightly larger in solution than in the crystalline state. These results led us to the conclusion that (i) the spontaneous domain motion was the property of the enzyme in solution, (ii) the domain motion was trapped in the crystallization process through different modes of crystal contacts, and (iii) the magnitude of the motion in solution was greater than that observed in the crystal structure. The present cryogenic diffraction experiment enabled us to identify 1931 hydration water molecules around the hexamer. The hydration structures around the subunits exhibited significant changes in accord with the degree of the domain movement. In particular, the hydration water molecules in the active-site cleft were rearranged markedly through migrations between specific hydration sites in coupling strongly with the domain movement. We discussed the cooperative dynamics between the domain motion and the hydration structure changes in the active site of the enzyme. The present study provides the first example of a visualized hydration structure varying transiently with the dynamic movements of enzymes and may form a new concept of the dynamics of multidomain enzymes in solution. PMID- 11258922 TI - Structure of a serine protease proteinase K from Tritirachium album limber at 0.98 A resolution. AB - X-ray diffraction data at atomic resolution to 0.98 A with 136 380 observed unique reflections were collected using a high quality proteinase K crystals grown under microgravity conditions and cryocooled. The structure has been refined anisotropically with REFMAC and SHELX-97 with R-factors of 11.4 and 12.8%, and R(free)-factors of 12.4 and 13.5%, respectively. The refined model coordinates have an overall rms shifts of 0.23 A relative to the same structure determined at room temperature at 1.5 A resolution. Several regions of the main chain and the side chains, which were not observed earlier have been seen more clearly. For example, amino acid 207, which was reported earlier as Ser has been clearly identified as Asp. Furthermore, side-chain disorders of 8 of 279 residues in the polypeptide have been identified. Hydrogen atoms appear as significant peaks in the F(o) - F(c) difference electron density map accounting for an estimated 46% of all hydrogen atoms at 2sigma level. Furthermore, the carbon, nitrogen, and oxygen atoms can be differentiated clearly in the electron density maps. Hydrogen bonds are clearly identified in the serine protease catalytic triad (Ser-His-Asp). Furthermore, electron density is observed for an unusual, short hydrogen bond between aspartic acid and histidine in the catalytic triad. The short hydrogen bond, designated "catalytic hydrogen bond", occurs as part of an elaborate hydrogen bond network, involving Asp of the catalytic triad. Though unusual, these features seem to be conserved in other serine proteases. Finally there are clear electron density peaks for the hydrogen atoms associated with the Ogamma of Ser 224 and Ndelta1 of His 69. PMID- 11258924 TI - Characterization of the maturation of human pro-apolipoprotein A-I in an in vitro model. AB - The reaction conditions and the protein structural features involved in the maturation of pro-apolipoprotein A-I (cleavage of pro-peptide) were investigated in an in vitro model. ProapoA-I, mutants and wild type, were expressed in the PGEX/E. coli expression system as fusion proteins with glutathione S-transferase (GST). Use of GST-proapoA-I and truncated forms of proapoA-I enabled quantitation of the amount of GST and apoA-I formed as a result of cleavage following incubation with human serum. Deletion of the pro-peptide (GST-apoA-I) resulted in complete inhibition of the reaction. Truncation of proapoA-I to residues 222, 150, 135, and 25 as well as substitution of residues -6, -5, and -4 with alanine did not affect the reaction. Substitution of residues -1, -2, 1, 3, and 4 with alanine either completely blocked or substantially inhibited cleavage of the pro peptide. The reaction was inhibited by addition of EDTA, o-phenanthroline, dithiothreitol, and beta-mercaptoethanol and to a lesser extent by p chloromercuriphenylsulfonic acid, but not by leupeptin, N-ethylmaleimide, PMSF, pepstatin A, or trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane. Calcium was essential for the activation of the cleavage enzyme, but it had a biphasic effect on the cleavage, activating it at concentrations below 1.5 mM and inhibiting at concentrations above 1.75 mM. Manganese alone was not essential for activation of the enzyme nor did it modify the effect of low concentration of calcium. However, a high concentration of manganese partially reverted the inhibitory effect of a high calcium concentration. Thus, residues within -2 to +4 are involved in forming the cleavage site for the maturation enzyme. The reaction of maturation is inhibited by metalloprotease inhibitors and is dependent upon calcium. PMID- 11258923 TI - C2 domains from different Ca2+ signaling pathways display functional and mechanistic diversity. AB - The ubiquitous C2 domain is a conserved Ca2+ triggered membrane-docking module that targets numerous signaling proteins to membrane surfaces where they regulate diverse processes critical for cell signaling. In this study, we quantitatively compared the equilibrium and kinetic parameters of C2 domains isolated from three functionally distinct signaling proteins: cytosolic phospholipase A2-alpha (cPLA2 alpha), protein kinase C-beta (PKC-beta), and synaptotagmin-IA (Syt-IA). The results show that equilibrium C2 domain docking to mixed phosphatidylcholine and phosphatidylserine membranes occurs at micromolar Ca2+ concentrations for the cPLA2-alpha C2 domain, but requires 3- and 10-fold higher Ca2+ concentrations for the PKC-beta and Syt-IA C2 domains ([Ca2+](1/2) = 4.7, 16, 48 microM, respectively). The Ca2+ triggered membrane docking reaction proceeds in at least two steps: rapid Ca2+ binding followed by slow membrane association. The greater Ca2+ sensitivity of the cPLA2-alpha domain results from its higher intrinsic Ca2+ affinity in the first step compared to the other domains. Assembly and disassembly of the ternary complex in response to rapid Ca2+ addition and removal, respectively, require greater than 400 ms for the cPLA2-alpha domain, compared to 13 ms for the PKC-beta domain and only 6 ms for the Syt-IA domain. Docking of the cPLA2-alpha domain to zwitterionic lipids is triggered by the binding of two Ca2+ ions and is stabilized via hydrophobic interactions, whereas docking of either the PKC-beta or the Syt-IA domain to anionic lipids is triggered by at least three Ca2+ ions and is maintained by electrostatic interactions. Thus, despite their sequence and architectural similarity, C2 domains are functionally specialized modules exhibiting equilibrium and kinetic parameters optimized for distinct Ca2+ signaling applications. This specialization is provided by the carefully tuned structural and electrostatic parameters of their Ca2+ and membrane-binding loops, which yield distinct patterns of Ca2+ coordination and contrasting mechanisms of membrane docking. PMID- 11258925 TI - Characterization of the glycosylation sites in cyclooxygenase-2 using mass spectrometry. AB - Cyclooxygenase is involved in the biosynthesis and function of prostaglandins. It is a glycoprotein located in the endoplasmic reticulum and in the nuclear envelope, and it has been found to have two isoforms termed COX-1 and COX-2. This paper reports on the glycosylation site analysis of recombinant COX-2 using matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) and nanoelectrospray (nanoESI) quadrupole-TOF (Q-TOF) MS. The nanoESI MS analysis of COX-2 revealed the presence of three glycoforms at average molecular masses of 71.4, 72.7, and 73.9 kDa. Each glycoform contained a number of peaks differing by 162 Da indicating heterogeneity and suggesting the presence of high-mannose sugars. The masses of the glycoforms indicate that oligosaccharides occupy two to four sites and a single N-acetylglucosamine (GlcNAc) residue occupied up to two sites. The MALDI MS analysis of a tryptic digest of the protein showed a number of potential glycopeptides. The peptides differed by 162 Da which further suggested high-mannose sugars. Nanoelectrospray MS/MS experiments confirmed glycosylation at the Asn 53 and Asn 130 sites and confirmed the presence of the peptides Asn 396-Arg 414 + GlcNAc and Thr 576-Arg 587 + GlcNAc containing Asn 580. It was not possible to conclusively determine whether the Asn 396 site was glycosylated via an MS/MS experiment, so the tryptic digest was deglycosylated to confirm the presence of the glycopeptides. Finally, a non-glycosylated tryptic peptide was observed containing the Asn 592. PMID- 11258926 TI - Determination of an optimal potential window for catalysis by E. coli dimethyl sulfoxide reductase and hypothesis on the role of Mo(V) in the reaction pathway. AB - Protein film voltammetry (PFV) of Escherichia coli dimethyl sulfoxide (DMSO) reductase (DmsABC) adsorbed at a graphite electrode reveals that the catalytic activity of this complex Mo-pterin/Fe-S enzyme is optimized within a narrow window of electrode potential. The upper and lower limits of this window are determined from the potential dependences of catalytic activity in reducing and oxidizing directions; i.e., for reduction of DMSO (or trimethylamine-N-oxide) and oxidation of trimethylphosphine (PMe(3)). At either limit, the catalytic activity drops despite the increase in driving force: as the potential is lowered below 200 mV (pH 7.0-8.9), the rate of reduction of DMSO decreases abruptly, while for PMe(3), an oxidative current is observed that vanishes as the potential is raised above +20 mV (pH 9.0). Analysis of the waveshapes reveals that both activity thresholds result from one-electron redox reactions that arise, most likely, from groups within the enzyme; if so, they represent "switches" that reflect the catalytic mechanism and may be of physiological relevance. The potential window of activity coincides approximately with the appearance of the Mo(V) EPR signal observed in potentiometric titrations, suggesting that crucial stages of catalysis are facilitated while the active site is in the intermediate Mo(V) oxidation state. PMID- 11258927 TI - Small angle neutron scattering and gel filtration analyses of neutrophil NADPH oxidase cytosolic factors highlight the role of the C-terminal end of p47phox in the association with p40phox. AB - The NADPH oxidase of phagocytic cells is regulated by the cytosolic factors p47(phox), p67(phox), and p40(phox) as well as by the Rac1-Rho-GDI heterodimer. The regulation is a consequence of protein-protein interactions involving a variety of protein domains that are well characterized in signal transduction. We have studied the behavior of the NADPH oxidase cytosolic factors in solution using small angle neutron scattering and gel filtration. p47(phox), two truncated forms of p47(phox), namely, p47(phox) without its C-terminal end (residues 1-358) and p47(phox) without its N-terminal end (residues 147-390), and p40(phox) were found to be monomeric in solution. The dimeric form of p67(phox) previously observed by gel filtration experiments was confirmed. Our small angle neutron scattering experiments show that p40(phox) binds to the full-length p47(phox) in solution in the absence of phosphorylation. We demonstrated that the C-terminal end of p47(phox) is essential in this interaction. From the comparison of the presence or absence of interaction with various truncated forms of the proteins, we confirmed that the SH3 domain of p40(phox) interacts with the C-terminal proline rich region of p47(phox). The radii of gyration observed for p47(phox) and the truncated forms of p47(phox) (without the C-terminal end or without the N terminal end) show that all these molecules are elongated and that the N-terminal end of p47(phox) is globular. These results suggest that the role of amphiphiles such as SDS or arachidonic acid or of p47(phox) phosphorylation in the elicitation of NADPH oxidase activation could be to disrupt the p40(phox) p47(phox) complex rather than to break an intramolecular interaction in p47(phox). PMID- 11258928 TI - Factors that control the reactivity of the interface cysteine of triosephosphate isomerase from Trypanosoma brucei and Trypanosoma cruzi. AB - The amino acid sequences and X-ray structures of homodimeric triosephosphate isomerase from the pathogenic parasites Trypanosoma brucei (TbTIM) and Trypanosoma cruzi (TcTIM) are markedly similar. In the two TIMs, the side chain of the only interface cysteine (Cys14) of one subunit docks into loop 3 of the other subunit. This portion of the interface is also markedly similar in the two enzymes. Nonetheless, Cys14 of TcTIM is nearly 2 orders of magnitude more susceptible to the thiol reagent methylmethane thiosulfonate (MMTS) than Cys14 of TbTIM. The causes of this difference were explored by measuring the second-order rate constant of inactivation by MMTS (k(2)) under various conditions. At pH 7.4, k(2) in TcTIM is 70 times higher than in TbTIM. The difference decreases to 30 when the amino acid sequence of loop 3 and adjoining residues of TbTIM are conferred to TcTIM (triple mutant). The pK(a) values of the thiol group of the interface cysteine of TcTIM and the triple mutant were 0.7 pH unit lower than in TbTIM. Because this difference could account for the different sensitivity of the enzymes to thiol reagents, we determined the k(2) of inactivation at equal levels of ionization of their interface cysteines. Under these conditions, the difference in k(2) between TcTIM and TbTIM became 8-fold, whereas that of the triple mutant to TbTIM was 1.5 times. The substrate analogue phosphoglycolate did not modify the pK(a) of the thiol group of the interface, albeit it diminished the rate of its derivatization by MMTS. In the presence of phosphoglycolate, under conditions in which the interface cysteines of the enzymes had equal levels of protonation, the difference in k(2) of TcTIM and TbTIM became smaller, whereas k(2) of the triple mutant was almost equal to that of TbTIM. Thus, from measurements of the reactivity of the interface cysteine in various conditions, it was possible to obtain information on the factors that control the dynamics of a portion of the dimer interface. PMID- 11258929 TI - Secondary structure and position of the cell-penetrating peptide transportan in SDS micelles as determined by NMR. AB - Transportan is a 27-residue peptide (GWTLN SAGYL LGKIN LKALA ALAKK IL-amide) which has the ability to penetrate into living cells carrying a hydrophilic load. Transportan is a chimeric peptide constructed from the 12 N-terminal residues of galanin in the N-terminus with the 14-residue sequence of mastoparan in the C terminus and a connecting lysine. Circular dichroism studies of transportan and mastoparan show that both peptides have close to random coil secondary structure in water. Sodium dodecyl sulfate (SDS) micelles induce 60% helix in transportan and 75% helix in mastoparan. The 600 MHz (1)H NMR studies of secondary structure in SDS micelles confirm the helix in mastoparan and show that in transportan the helix is localized to the mastoparan part. The less structured N-terminus of transportan has a secondary structure similar to that of the same sequence in galanin [Ohman, A., et al. (1998) Biochemistry 37, 9169-9178]. The position of mastoparan and transportan relative to the SDS micelle surface was studied by adding spin-labeled 5-doxyl- or 12-doxyl-stearic acid or Mn2+ to the peptide/micelle system. The combined results show that the peptides are for the most part buried in the SDS micelles. Only the C-terminal parts of both peptides and the central segment connecting the two parts of transportan are clearly surface exposed. For mastoparan, the secondary chemical shifts of the amide protons were found to vary periodically and display a pattern almost identical to those reported for mastoparan in phospholipid bicelles [Vold, R., et al. (1997) J. Biomol. NMR 9, 329-335], indicating similar structures and interactions in the two membrane-mimicking environments. PMID- 11258930 TI - An N-terminal three-helix fragment of the exchangeable insect apolipoprotein apolipophorin III conserves the lipid binding properties of wild-type protein. AB - Apolipophorin III (apoLp-III) from the greater wax moth Galleria mellonella is an exchangeable insect apolipoprotein that consists of five amphipathic alpha helices, sharing high sequence identity with apoLp-III from the sphinx moth Manduca sexta whose structure is available. To define the minimal requirement for apoLp-III structural stability and function, a C-terminal truncated apoLp-III encompassing residues 1-91 of this 163 amino acid protein was designed. Far-UV circular dichroism spectroscopy revealed apoLp-III(1-91) has 50% alpha-helix secondary structure content in buffer (wild-type apoLp-III 86%), increasing to essentially 100% upon interactions with dimyristoylphosphatidylcholine (DMPC). Guanidine hydrochloride denaturation studies revealed similar stability properties for wild-type apoLp-III and apoLp-III(1-91). Resistance to denaturation for both proteins increased substantially upon association with phospholipid. In the absence of lipid, wild-type apoLp-III was monomeric whereas apoLp-III(1-91) partly formed dimers and trimers. Discoidal apoLp-III(1-91)-DMPC complexes were smaller in diameter (13.5 nm) compared to wild-type apoLp-III (17.7 nm), and more molecules of apoLp-III(1-91) associated with the complexes. Lipid interaction revealed that apoLp-III(1-91) binds to modified spherical lipoprotein surfaces and efficiently transforms phospholipid vesicles into discoidal complexes. Thus, the first three helices of G. mellonella apoLp-III contain the basic features required for maintenance of the structural integrity of the entire protein. PMID- 11258931 TI - Investigations of the alkaline and acid transitions of umecyanin, a stellacyanin from horseradish roots. AB - The effect of pH on Cu(I) and Cu(II) umecyanin (UCu), a phytocyanin obtained from horseradish roots, has been studied by electronic and NMR spectroscopy and using direct electrochemical measurements. A pK(a) value of approximately 9.5-9.8 is observed for the alkaline transition in UCu(II), and this leads to a slightly altered active site structure, as indicated by the changes in the paramagnetic 1H NMR spectrum. Electrochemical studies show that the pK(a) value for this transition in UCu(I) is 9.9. The alkaline transition is caused by the deprotonation of a surface lysine residue, with Lys96 being the most likely candidate. The isotropically shifted resonances in the (1)H NMR spectrum of UCu(II) also shift upon lowering the pH (pK(a) 5.8), and this can be assigned to the protonation of the surface (noncoordinating) His65 residue. This histidine titrates in UCu(I) with a pK(a) of 6.3. The reduction potential of the protein in this range is also dependent on pH, and pK(a) values matching those from NMR, for the two oxidation states of the protein, are obtained. There is no evidence for either of the active site histidines (His44 and His90) titrating in UCu(I) in the pH range studied (down to pH 3.7). Also highlighted in these studies are the remarkable active site similarities between umecyanin and the other phytocyanins which possess an axial Gln ligand. PMID- 11258932 TI - S100A7, S100A10, and S100A11 are transglutaminase substrates. AB - S100 proteins are a family of 10-14 kDa EF-hand-containing calcium binding proteins that function to transmit calcium-dependent cell regulatory signals. S100 proteins have no intrinsic enzyme activity but bind in a calcium-dependent manner to target proteins to modulate target protein function. Transglutaminases are enzymes that catalyze the formation of covalent epsilon-(gamma glutamyl)lysine bonds between protein-bound glutamine and lysine residues. In the present study we show that transglutaminase-dependent covalent modification is a property shared by several S100 proteins and that both type I and type II transglutaminases can modify S100 proteins. We further show that the reactive regions are at the solvent-exposed amino- and carboxyl-terminal ends of the protein, regions that specify S100 protein function. We suggest that transglutaminase-dependent modification is a general mechanism designed to regulate S100 protein function. PMID- 11258933 TI - The closed/open model for lipase activation. Addressing intermediate active forms of fungal enzymes by trapping of conformers in water-restricted environments. AB - The behavior of prototypic fungal lipases in a water-restricted environment has been investigated by exploiting the reported experimental strategy that allows the trapping (freeze-drying) of the enzyme in the conformation present in aqueous solution and to subsequently assay it in nonaqueous media [Mingarro, I., Abad, C., and Braco, L. (1995) Proc. Natl. Acad. Sci. U.S.A. 92, 3308-3312]. We now report, using simple esterification as well as acidolysis (triglycerides as substrates) as nonaqueous model reactions, that the presence of a detergent (n octyl-beta-glucopyranoside) in the freeze-drying buffer, at concentrations below the critical micellar concentration, generates different catalytically active (kinetically trapped) conformational states of the enzyme. These activated forms exquisitely discriminate between short- and long-chain fatty acids, suggesting that they can be correlated with intermediate conformations of the protein sufficiently open to permit the access of relatively small but not large substrates. Additional data obtained from aqueous solution activity measurements in the presence of detergent revealed that the fungal lipase retains an active conformation induced by high detergent concentration (30 mM) for a long period of time, a 'memory effect', which is stabilized in the absence of a well-defined interface by few detergent molecules. Together these results provide support to a model of lipase action involving several equilibrium states (closed, intermediate, and open), which can be modulated by the composition of the microenvironment, i.e., by the detergent concentration. PMID- 11258934 TI - Helix packing in the lactose permease of Escherichia coli: distances between site directed nitroxides and a lanthanide. AB - By exploiting substrate protection of Cys148 in lactose permease, a methanethiosulfonate nitroxide spin-label was directed specifically to one of two Cys residues in a double-Cys mutant, followed by labeling of Cys148 with a thiol reactive chelator that binds Gd(III) quantitatively. Distances between bound Gd(III) and the nitroxide spin-label were then studied by electron paramagnetic resonance. The results demonstrate that the Gd(III)-induced relaxation effects on nitroxides at positions 228, 226 (helix VII), and 275 (helix VIII) agree qualitatively with results obtained by studying spin-spin interactions [Wu, J., Voss, J., et al. (1996) Proc. Natl. Acad. Sci. U.S.A. 93, 10123-10127]. Thus, a nitroxide attached to position 228 (helix VII) is closest to the lanthanide at position 148 (helix V), a nitroxide at position 275 (helix VIII) is further away, and the distance between positions 226 (helix VII) and 148 is too long to measure. However, the Gd(III)-spin-label distances are significantly longer than those estimated from nitroxide-nitroxide interactions between the same pairs due to the nature of the chelator. Although the results provide strong confirmation for the contention that helix V lies close to both helices VII and VIII in the tertiary structure of lactose permease, other methods for binding rare earth metals are discussed which do not involve the use of bulky chelators with long linkers. PMID- 11258935 TI - Calmodulin binding to the C-terminus of the small-conductance Ca2+-activated K+ channel hSK1 is affected by alternative splicing. AB - We identified three splice variants of hSK1 whose C-terminal structures are determined by the independent deletion of two contiguous nucleotide sequences. The upstream sequence extends 25 bases in length, is initiated by a donor splice site within exon 8, and terminates at the end of the exon. The downstream sequence consists of nine bases that compose exon 9. When the upstream sequence (hSK1(-)(25b)) or both sequences (hSK1(-)(34b)) are deleted, truncated proteins are encoded in which the terminal 118 amino acids are absent. The binding of calmodulin to these variants is diminished, particularly in the absence of Ca2+ ions. The first 20 amino acids of the segment deleted from hSK1(-)(25b) and hSK1( )(34b) contain a 1-8-14 Ca2+ calmodulin binding motif, and synthetic oligopeptides based on this region bind calmodulin better in the presence than absence of Ca2+ ions. When the downstream sequence (hSK1(-)(9b)) alone is deleted, only the three amino acids A452, Q453, and K454 are removed, and calmodulin binding is not reduced. On the basis of the relative abundance of mRNA encoding each of the four isoforms, the full-length variant appears to account for most hSK1 in the human hippocampus, while hSK1(-)(34b) predominates in reticulocytes, and hSK1(-)(9b) is especially abundant in human erythroleukemia cells in culture. We conclude that the binding of calmodulin by hSK1 can be modulated through alternative splicing. PMID- 11258936 TI - Structure and interaction with membrane model systems of a peptide derived from the major epitope region of HIV protein gp41: implications on viral fusion mechanism. AB - The HIV-1 gp41 envelope protein mediates entry of the virus into the target cell by promoting membrane fusion. With a view toward possible new insights into viral fusion mechanisms, we have investigated by infrared, fluorescence, and nuclear magnetic resonance spectroscopies and calorimetry a fragment of 19 amino acids corresponding to the immunodominant region of the gp41 ectodomain, a highly conserved sequence and major epitope. Information on the structure of the peptide both in solution and in the presence of model membranes, its incorporation and location in the phospholipid bilayer, and the modulation of the phase behavior of the membrane has been gathered. Here we demonstrate that the peptide binds and interacts with negatively charged phospholipids, changes its conformation in the presence of a membraneous medium, and induces leakage of vesicle contents as well as a new phospholipid phase. These characteristics might be important for the formation of the fusion-active gp41 core structure, promoting the close apposition of the two viral and target-cell membranes and therefore provoking fusion. PMID- 11258937 TI - Human flap endonuclease-1: conformational change upon binding to the flap DNA substrate and location of the Mg2+ binding site. AB - Human flap endonuclease-1 (FEN-1) is a member of the structure-specific endonuclease family and is a key enzyme in DNA replication and repair. FEN-1 recognizes the 5'-flap DNA structure and cleaves it, a specialized endonuclease function essential for the processing of Okazaki fragments during DNA replication and for the repair of 5'-end single-stranded tails from nicked double-stranded DNA substrates. Magnesium is a cofactor required for nuclease activity. We have used Fourier transform infrared (FTIR) spectroscopy to better understand how Mg2+ and flap DNA interact with human FEN-1. FTIR spectroscopy provides three fundamentally new insights into the structural changes induced by the interaction of FEN-1 with substrate DNA and Mg2+. First, FTIR difference spectra in the amide I vibrational band (1600-1700 cm(-1)) reveal a change in the secondary structure of FEN-1 induced by substrate DNA binding. Quantitative analysis of the FTIR spectra indicates a 4% increase in helicity upon DNA binding or about 14 residues converted from disordered to helical conformations. The observation that the residues are disordered without DNA strongly implicates the flexible loop region. The conversion to helix also suggests a mechanism for locking the flexible loop region around the bound DNA. This is the first direct experimental evidence for a binding mechanism that involves a secondary structural change of the protein. Second, in contrast with DNA binding, no change is observed in the secondary structure of FEN-1 upon Mg2+ binding to the wild type or to the noncleaving D181A mutant. Third, the FTIR results provide direct evidence (via the carboxylate ligand band at 1535 cm(-1)) that not only is D181 a ligand to Mg2+ in the human enzyme but Mg2+ binding does not occur in the D181A mutant which lacks this ligand. PMID- 11258938 TI - Significance of nucleobase shape complementarity and hydrogen bonding in the formation and stability of the closed polymerase-DNA complex. AB - DNA polymerases insert a dNTP by a multistep mechanism that involves a conformational rearrangement from an open to a closed ternary complex, a process that positions the incoming dNTP in the proper orientation for phosphodiester bond formation. In this work, the importance and relative contribution of hydrogen-bonding interactions and the geometric shape of the base pair that forms during this process were studied using Escherichia coli DNA polymerase I (Klenow fragment, 3'-exonuclease deficient) and natural dNTPs or non-hydrogen-bonding dNTP analogues. Both the geometric fit of the incoming nucleotide and its ability to form Watson-Crick hydrogen bonds with the template were found to contribute to the stability of the closed ternary complex. Although the formation of a closed complex in the presence of a non-hydrogen-bonding nucleotide analogue could be detected by limited proteolysis analysis, a comparison of the stabilities of the ternary complexes indicated that hydrogen-bonding interactions between the incoming dNTP and the template increase the stability of the complex by 6-20 fold. Any deviation from the Watson-Crick base pair geometry was shown to have a destabilizing effect on the closed complex. This degree of destabilization varied from 3- to 730-fold and was found to be correlated with the size of the mismatched base pair. Finally, a stable closed complex is not formed in the presence of a ddNTP or rNTP. These results are discussed in relation to the steric exclusion model for the nucleotide insertion. PMID- 11258939 TI - Proposed secondary structure of eukaryote specific expansion segment 15 in 28S rRNA from mice, rats, and rabbits. AB - The expansion segments in eukaryotic ribosomal RNAs are additional RNA sequences not found in the RNA core common to both prokaryotes and eukaryotes. These regions show large species-dependent variations in sequence and size. This makes it difficult to create secondary structure models for the expansion segments exclusively based on phylogenetic sequence comparison. Here we have used a combination of experimental data and computational methods to generate secondary structure models for expansion segment 15 in 28S rRNA in mice, rats, and rabbits. The experimental data were collected using the structure sensitive reagents DMS, CMCT, kethoxal, micrococcal nuclease, RNase T(1), RNase CL3, RNase V(1), and lead(II) acetate. ES15 was folded with the computer program RNAStructure 3.5 using modification data and phylogenetic similarities between different ES15 sequences. This program uses energy minimization to find the most stable secondary structure of an RNA sequence. The presented secondary structure models include several common structural motifs, but they also have characteristics unique to each organism. Overall, the secondary structure models showed indications of an energetically stable but dynamic structure, easily accessible from the solution by the modification reagents, suggesting that the expansion segment is located on the ribosomal surface. PMID- 11258940 TI - Trapping conformational intermediate states in the reaction center protein from photosynthetic bacteria. AB - In protein, conformational changes are often crucial for function but not easy to observe. Two functionally relevant conformational intermediate states of photosynthetic reaction center protein (RCs) are trapped and characterized at low temperature. RCs frozen in the dark do not allow electron transfer from the reduced primary quinone, Q(A)(-), to the secondary quinone, Q(B). In contrast, RCs frozen under illumination in the product (P(+)Q(A)Q(B)(-)) state, with the oxidized electron donor, P(+), and reduced Q(B)(-), return to the ground state at cryogenic temperature in a conformation that allows a high yield of Q(B) reduction. Thus, RCs frozen under illumination are found to be trapped above the ground state in a conformation that allows product formation. When the temperature is raised above 120 K, the protein relaxes to an inactive conformation which is different from the RCs frozen in the dark. The activation energy for this change is 87 +/- 8 meV, and the active and inactive states differ in energy by only 16 +/- 3 meV. Thus, there are several conformational substates along the reaction coordinate with different transition temperatures. The ground state spectra of the RCs in active and inactive conformations report differences in the intraprotein electrostatic field, demonstrating that the dipole or charge distribution has changed. In addition, the electrochromic shift associated with the Q(A)(-) to Q(B) electron transfer at low temperature was characterized. The electron-transfer rate from Q(B)(-) to P(+) was measured at cryogenic temperature and is similar to the rate at room temperature, as expected for an exothermic, electron tunneling reaction in RCs. PMID- 11258942 TI - The ribosome at atomic resolution. AB - The publication of atomic resolution crystal structures for the large ribosomal subunit from Haloarcula marismortui and the small ribosomal subunit from Thermus thermophilus has permanently altered the way protein synthesis is conceptualized and experiments designed to address its unresolved issues. The impact of these structures on RNA biochemistry is certain to be no less profound. The background and substance of these developments are reviewed here. PMID- 11258943 TI - Substitution of aspartic acid for methionine-306 in factor VIIa abolishes the allosteric linkage between the active site and the binding interface with tissue factor. AB - The enzyme factor VIIa (FVIIa) triggers the blood coagulation cascade upon association with tissue factor (TF). The TF-induced allosteric enhancement of FVIIa's activity contributes to the procoagulant activity of the complex, and Met 306 in the serine protease domain of FVIIa participates in this event. We have characterized FVIIa variants mutated in position 306 with respect to their ability to be stimulated by TF. The amidolytic activity of FVIIa mutants with Ser, Thr, and Asn in position 306 was stimulated 9-, 12-, and 7-fold, respectively, by soluble TF as compared to 22-fold for wild-type FVIIa. In contrast, the activity of Met306Asp-FVIIa only increased about 2-fold and that of Met306Asp/Asp309Ser-FVIIa increased about 1.5-fold. Modeling suggests that Asp in position 306 prevents the TF-induced stimulation of FVIIa by disrupting essential intermolecular hydrogen bonds. The ability of the FVIIa variants to catalyze factor X activation and the amidolytic activity were enhanced to a similar extent by soluble TF. This indicates that factor X does not promote its own activation through interactions with exosites on FVIIa made accessible upon FVIIa-TF assembly. Met306Asp-FVIIa binds soluble TF with a dissociation constant of 13 nM (about 3-fold higher than that of FVIIa), and, in sharp contrast to FVIIa, its binding kinetics are unaltered after inactivation with D-Phe-Phe-Arg chloromethyl ketone. We conclude that a single specific amino acid replacement, substitution of Asp for Met-306, virtually prevents the TF-induced allosteric changes which normally result in dramatically increased FVIIa activity and eliminates the effect of the active site inhibitor on TF affinity. PMID- 11258944 TI - Identification of a stability determinant on the edge of the Tet repressor four helix bundle dimerization motif. AB - Isofunctional tetracycline repressor (TetR) proteins isolated from different bacteria show a sequence identity between 38 and 88% of the residues. Their active state is a homodimer formed by a four-alpha-helix bundle as the main interaction motif. We utilize this sequence variation of isofunctional proteins to determine residues contributing to the stability of the four-helix bundle. The thermodynamic stabilities of two TetR proteins with 63% sequence identity were determined by urea-induced reversible denaturation followed by fluorescence and circular dichroism. Both methods yield identical results. The deltaG(o)U (H2O) values are 60 and 75 kJ x mol(-1). We have constructed TetR hybrid proteins derived from these wild types to identify the determinant leading to the 15 kJ x mol(-1) stability difference. Successive size reduction of the exchanged portion yielded two single residues affecting the overall protein stability. The P184Q exchange leads to a more stable protein, whereas the G181D exchange located at the solvent's exposed edge of the four-helix bundle is solely responsible for the reduced stability. Additional mutants based on crystal structures of TetR do not reveal any hint for steric interference of the Asp181 side chain with neighboring residues. Thus, this is an example for the role played by surface-exposed turn residues for the stability of four-helix bundles. We assume that the larger conformational flexibility of Gly and the reduction of the negative surface charge could favor formation of the turn on the edge of the four-helix bundle. PMID- 11258945 TI - Changes in Ca2+ affinity upon activation of Agkistrodon piscivorus piscivorus phospholipase A2. AB - Changes in the affinity of calcium for phospholipase A2 from Agkistrodon piscivorus piscivorus during activation of the enzyme on the surface of phosphatidylcholine vesicles have been investigated by site-directed mutagenesis and fluorescence spectroscopy. Changes in fluorescence that occur during lipid binding and subsequent activation have been ascribed to each of the three individual Trp residues in the protein. This was accomplished by generating a panel of mutant proteins, each of which lacks one or more Trp residues. Both Trp21, which is found in the interfacial binding region, and Trp119 show changes in fluorescence upon protein binding to small unilamellar zwitterionic vesicles or large unilamellar vesicles containing sufficient anionic lipid. Trp31, which is near the Ca2+ binding loop, exhibits little change in fluorescence upon lipid bilayer binding. A change in the fluorescence of the protein also occurs during activation of the enzyme. These changes arise from residue Trp31 as well as residues Trp21 and Trp119. The calcium dependence of the fluorescence change of Trp31 indicates that the affinity of the enzyme for calcium increases at least 3 orders of magnitude upon activation. These studies suggest either that a change in conformation of the enzyme occurs upon activation or that the increase in calcium affinity reflects formation of a ternary complex of calcium, enzyme, and substrate. PMID- 11258946 TI - Cl- channel inhibitors of the arylaminobenzoate type act as photosystem II herbicides: a functional and structural study. AB - The Cl- channel blocker NPPB (5-nitro-2-(3-phenylpropylamino) benzoic acid) inhibited photosynthetic oxygen evolution of isolated thylakoid membranes in a pH dependent manner with a K(i) of about 2 microM at pH 6. Applying different electron acceptors, taking electrons either directly from photosystem II (PS II) or photosystem I (PS I), the site of inhibition was localized within PS II. Measurements of fluorescence induction kinetics and thermoluminescence suggest that the binding of NPPB to the QB binding site of PS II is similar to the herbicide DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea). The effects of different arylaminobenzoate derivatives and other Cl- channel inhibitors on photosynthetic electron transport were investigated. The structure--activity relationship of the inhibitory effect on PS II shows interesting parallels to the one observed for the arylaminobenzoate block of mammalian Cl- channels. A molecular modeling approach was used to fit NPPB into the QB binding site and to identify possible molecular interactions between NPPB and the amino acid residues of the binding site in PS II. Taken together, these data give a detailed molecular picture of the mechanism of NPPB binding. PMID- 11258947 TI - Ultra-high-field MAS NMR assay of a multispin labeled ligand bound to its G protein receptor target in the natural membrane environment: electronic structure of the retinylidene chromophore in rhodopsin. AB - 11-Z-[8,9,10,11,12,13,14,15,19,20-(13)C10]Retinal prepared by total synthesis is reconstituted with opsin to form rhodopsin in the natural lipid membrane environment. The 13C shifts are assigned with magic angle spinning NMR dipolar correlation spectroscopy in a single experiment and compared with data of singly labeled retinylidene ligands in detergent-solubilized rhodopsin. The use of multispin labeling in combination with 2-D correlation spectroscopy improves the relative accuracy of the shift measurements. We have used the chemical shift data to analyze the electronic structure of the retinylidene ligand at three levels of understanding: (i) by specifying interactions between the 13C-labeled ligand and the G-protein-coupled receptor target, (ii) by making a charge assessment of the protonation of the Schiff base in rhodopsin, and (iii) by evaluating the total charge on the carbons of the retinylidene chromophore. In this way it is shown that a conjugation defect is the predominant ground-state property governing the molecular electronics of the retinylidene chromophore in rhodopsin. The cumulative chemical shifts at the odd-numbered carbons (Delta(sigma)odd) of 11-Z protonated Schiff base models relative to the unprotonated Schiff base can be used to measure the extent of delocalization of positive charge into the polyene. For a series of 11-Z-protonated Schiff base models and rhodopsin, Delta(sigma)odd appears to correlate linearly with the frequency of maximum visible absorption. Since rhodopsin has the largest value of Delta(sigma)odd, the data contribute to existing and converging spectroscopic evidence for a complex counterion stabilizing the protonated Schiff base in the binding pocket. PMID- 11258948 TI - ADP ribosylation of Arg41 of Rap by ExoS inhibits the ability of Rap to interact with its guanine nucleotide exchange factor, C3G. AB - ExoS is a bifunctional type III cytotoxin that is secreted by Pseudomonas aeruginosa. The N-terminal domain comprises a RhoGAP activity, while the C terminal domain comprises a ADP-ribosyltransferase activity. Previous studies showed that ExoS ADP ribosylated Ras at Arg41 which interfered with the ability of Ras to interact with its guanine nucleotide exchange factor. Rap and Ras share considerable primary amino acid homology, including Arg41. In this study, we report that ExoS ADP ribosylates Rap1b at Arg41 and that ADP ribosylation of Arg41 inhibits the ability of C3G to stimulate guanine nucleotide exchange. The mechanism responsible for this inhibition is one in which ADP-ribosylated Rap binds inefficiently to C3G, relative to wild type Rap. This identifies a second member of the Ras GTPase subfamily that can be ADP ribosylated by ExoS and indicates that ExoS can inhibit both Ras and Rap signaling pathways in eukaryotic cells. PMID- 11258949 TI - Interactions of the 8-kDa domain of rat DNA polymerase beta with DNA. AB - Interactions between the isolated 8-kDa domain of the rat DNA polymerase beta and DNA have been studied, using the quantitative fluorescence titration technique. The obtained results show that the number of nucleotide residues occluded in the native 8-kDa domain complex with the ssDNA (the site size) is strongly affected by Mg2+ cations. In the absence of Mg2+, the domain occludes 13 +/- 0.7 nucleotide residues, while in the presence of Mg2+ the site size decreases to 9 +/- 0.6 nucleotides. The high affinity of the magnesium cation binding, as well as the dramatic changes in the monovalent salt effect on the protein-ssDNA interactions in the presence of Mg2+, indicates that the site size decrease results from the Mg2+ binding to the domain. The site size of the isolated domain ssDNA complex is significantly larger than the 5 +/- 2 site size determined for the (pol beta)5 binding mode formed by an intact polymerase, indicating that the intact enzyme, but not the isolated domain, has the ability to use only part of the domain DNA-binding site in its interactions with the nucleic acid. Salt effect on the intrinsic interactions of the domain with the ssDNA indicates that a net release of m approximately 5 ions accompanies the complex formation. Independence of the number of ions released upon the type of anion in solution strongly suggests that the domain forms as many as seven ionic contacts with the ssDNA. Experiments with different ssDNA oligomers show that the affinity decreases gradually with the decreasing number of nucleotide residues in the oligomer. The data indicate a continuous, energetically homogeneous structure of the DNA-binding site of the domain, with crucial, nonspecific contacts between the protein and the DNA evenly distributed over the entire binding site. The DNA binding site shows little base specificity. Moreover, the domain has an intrinsic affinity and site size of its complex with the dsDNA conformation, similar to the affinity and site size with the ssDNA. The significance of these results for the mechanistic role of the 8-kDa domain in the functioning of rat pol beta is discussed. PMID- 11258950 TI - RNA species that replicate with DNA-dependent RNA polymerase from Escherichia coli. AB - An RNA that replicates with core RNA polymerase from E. coli and the substrates ATP, CTP, ITP, and UTP, was selected from a random poly(A,U,I,C) library and named EcorpI. Another replicating RNA, EcorpG, was obtained by template-free incubation of holo RNA polymerase and the substrates ATP, CTP, GTP, and UTP. Both RNA species showed typical autocatalytic RNA amplification profiles with replication rates in the range of other RNA replicons. The replication products were heterogeneous in length; the different lengths appeared to be different replication intermediates. Both RNA were single-stranded with much internal base pairing but low melting points. Their sequences were composed by permutations of certain sequence motives in both polarities separated by short oligo(A) and oligo(U) clusters. There was evidence for 3'-terminal elongation on an intramolecular template. No double-stranded RNA was found, even though base pairing is certainly the underlying basis of the replication process. The reaction was highly sensitive: a few RNA strands were sufficient to trigger an amplification avalanche. PMID- 11258951 TI - Stimulation of topoisomerase II-mediated DNA damage via a mechanism involving protein thiolation. AB - The breakage/reunion reaction of DNA topoisomerase II (TOP2) can be interrupted by DNA intercalators (e.g., doxorubicin), enzyme binders (e.g., etoposide), or DNA lesions (e.g., abasic sites) to produce TOP2-mediated DNA damage. Here, we demonstrate that thiol alkylation of TOP2 can also produce TOP2-mediated DNA damage. This conclusion is supported by the following observations using purified TOP2: (1) Thiol-reactive quinones were shown to induce TOP2-mediated DNA cleavage. (2) Thiol-reactive compounds such as N-ethylmaleimide (NEM), disulfiram, and organic disulfides [e.g., 2,2'-dithiobis(5-nitropyridine)] were also shown to induce TOP2-mediated DNA cleavage with similar reaction characteristics as thiol-reactive quinones. (3) TOP2-mediated DNA cleavage induced by thiol-reactive quinones was completely abolished using mutant yeast TOP2 with all cysteine residues replaced with alanine (cysteineless TOP2). These results suggest the possibility that cellular DNA damage could occur indirectly through thiolation of a nuclear protein, TOP2. The implications of this reaction in carcinogenesis and apoptotic cell death are discussed. PMID- 11258952 TI - Conformational changes of the 120-kDa Na+/Ca2+ exchanger protein upon ligand binding: a Fourier transform infrared spectroscopy study. AB - The 120-kDa Na+/Ca2+ exchanger was purified and reconstituted into lipid vesicles. The secondary structure composition of the exchanger was 39% alpha helices, 20% beta-sheets, 25% beta-turns, and 16% random coils, as analyzed by Fourier transform infrared attenuated total reflection spectroscopy. The secondary structure composition of the COOH-terminal portion of the protein was compatible with a topology model containing 4-6 transmembrane segments. Furthermore, the secondary structure of the NH2-terminal portion of the cytoplasmic loop was analyzed and found to be different from that of the COOH terminal portion. Ca2+ and/or the exchange inhibitory peptide (XIP) failed to affect the secondary structure of the 120-kDa protein. Tertiary structure modifications induced by Ca2+ and XIP were analyzed by monitoring the hydrogen/deuterium exchange rate for the reconstituted exchanger. In the absence of ligand, 51% of the protein was accessible to solvent. Ca2+ decreased accessibility to 40%, implicating the shielding of at least 103 amino acids. When both Ca2+ and XIP were added, accessibility increased to 66%. No modification was obtained when XIP was added alone. Likewise, in the presence of Ca2+, XIP failed to modify the tertiary structure of the 70-kDa protein, suggesting that XIP acts at the level of the COOH-terminal portion of the intracellular loop. The present data describe, for the first time, conformational changes of the Na+/Ca2+ exchanger induced by Ca2+ and XIP, compatible with an interaction model where regulatory Ca2+ and inhibitory XIP bind to distinct sites, and where XIP binding requires the presence of Ca2+. PMID- 11258953 TI - Electron paramagnetic resonance analysis of different Azotobacter vinelandii nitrogenase MoFe-protein conformations generated during enzyme turnover: evidence for S = 3/2 spin states from reduced MoFe-protein intermediates. AB - Rapid-freezing experiments elicited two transient EPR signals, designated 1b and 1c, during Azotobacter vinelandii nitrogenase turnover at 23 degrees C and pH 7.4. The first of the signals to form, signal 1b, exhibited g values of 4.21 and 3.76. Its formation was at the expense of the starting EPR signal (signal 1a with g values of 4.32, 3.66, and 2.01). The second signal to arise, signal 1c, with a characteristic g value of 4.69, formed very slowly and was always of low intensity. Both signals occurred independently of the substrate being reduced. Increased electron flux through the MoFe protein caused these signals to form more rapidly. Moreover, after a MoFe-protein solution had been pretreated (using conditions of extremely low electron flux) to set up an equimolar mixture of its resting state and one-electron reduced state, these signals appeared even more rapidly when this mixture was exposed to an excess of the Fe protein. We have simulated the kinetics of formation of these EPR features using the published kinetic model for nitrogenase catalysis [Lowe, D. J., and Thorneley, R. N. F. (1984) Biochem. J. 224, 887-909] and propose that they arise from reduced states of the MoFe protein and reflect different conformations of the FeMo cofactor with different protonation states. PMID- 11258954 TI - Hydrogen peroxide formation during iron deposition in horse spleen ferritin using O2 as an oxidant. AB - The reaction of Fe2+ with O2 in the presence of horse spleen ferritin (HoSF) results in deposition of FeOH3 into the hollow interior of HoSF. This reaction was examined at low Fe2+/HoSF ratios (5-100) under saturating air at pH 6.5-8.0 to determine if H2O2 is a product of the iron deposition reaction. Three methods specific for H2O2 detection were used to assess H2O2 formation: (1) a fluorometric method with emission at 590 nm, (2) an optical absorbance method based on the reaction H2O2 + 3I- + 2H+ = I3- + 2H2O monitored at 340 nm for I3- formation, and (3) a differential pulsed electrochemical method that measures O2 and H2O2 concentrations simultaneously. Detection limits of 0.25, 2.5, and 5.0 microM H2O2 were determined for the three methods, respectively. Under constant air-saturation conditions (20% O2) and for a 5-100 Fe2+/HoSF ratio, Fe2+ was oxidized and the resulting Fe3+ was deposited within HoSF but no H2O2 was detected as predicted by the reaction 2Fe2+ + O2 + 6H2O = 2Fe(OH)3 + H2O2 + 4H+. Two other sets of conditions were also examined: one with excess but nonsaturating O2 and another with limiting O2. No H2O2 was detected in either case. The absence of H2O2 formation under these same conditions was confirmed by microcoulometric measurements. Taken together, the results show that under low iron loading conditions (5-100 Fe2+/HoSF ratio), H2O2 is not produced during iron deposition into HoSF using O2 as an oxidant. This conclusion is inconsistent with previous, carefully conducted stoichiometric and kinetic measurements [Xu, B., and Chasteen, N. D. (1991) J. Biol. Chem. 266, 19965], predicting that H2O2 is a quantitative product of the iron deposition reaction with O2 as an oxidant, even though it was not directly detected. Possible explanations for these conflicting results are considered. PMID- 11258955 TI - Modifying Mg2+ binding and exchange with the N-terminal of calmodulin. AB - To follow Mg2+ binding to the N-terminal of calmodulin (CaM), we substituted Phe in position 19, which immediately precedes the first Ca2+/Mg2+ binding loop, with Trp, thus making F19WCaM (W-Z). W-Z has four acidic residues in chelating positions, two of which form a native Z-acid pair. We then generated seven additional N-terminal CaM mutants to examine the role of chelating acidic residues in Mg2+ binding and exchange with the first EF-hand of CaM. A CaM mutant with acidic residues in all of the chelating positions exhibited Mg2+ affinity similar to that of W-Z. Only CaM mutants that had a Z-acid pair were able to bind Mg2+ with physiologically relevant affinities. Removal of the Z-acid pair from the first EF-hand produced a dramatic 58-fold decrease in its Mg2+ affinity. Additionally, removal of the Z-acid pair led to a 1.8-fold increase in the rate of Mg2+ dissociation. Addition of an X- or Y-acid pair could not restore the high Mg2+ binding lost with removal of the Z-acid pair. Therefore, the Z-acid pair in the first EF-hand of CaM supports high Mg2+ binding primarily by increasing the rate of Mg2+ association. PMID- 11258956 TI - Evidence that Myc isoforms transcriptionally repress caveolin-1 gene expression via an INR-dependent mechanism. AB - The c-Myc oncoprotein contributes to oncogenesis by activating and repressing a repertoire of genes involved in cellular proliferation, metabolism, and apoptosis. Increasing evidence suggests that the repressor function of c-Myc is critical for transformation. Therefore, identifying and characterizing Myc repressed genes is imperative to understanding the mechanisms of Myc-induced tumorigenesis. Here, we employ NIH 3T3 cell lines harboring c-Myc-ER or N-Myc-ER to dissect the relationship between Myc activation and caveolin-1 expression. In this well-established inducible system, treatment with estrogen like molecules, such as tamoxifen, leads to activation of Myc, but in a tightly controlled fashion. Using this approach, we show that Myc activation induces the repression of caveolin-1 expression at the transcriptional level. We also provide two independent lines of evidence suggesting that caveolin-1 is a direct target of Myc: (i) the effect of Myc activation on caveolin-1 expression is independent of new protein synthesis, as revealed through the use of cycloheximide; and (ii) Myc mediated repression of the caveolin-1 promoter is dependent on an intact INR sequence. Moreover, we show that expression of caveolin-1, via an adenoviral vector approach, can suppress cell transformation that is mediated by Myc activation. In support of these observations, treatment with an adenoviral vector harboring anti-sense caveolin-1 specifically potentiates transformation induced by Myc activation. Taken together, our results indicate that caveolin-1 is a direct target of Myc repression, and they also provide evidence for an additional mechanism by which Myc repression can elicit a malignant phenotype. PMID- 11258957 TI - Correlation between processing efficiency for ribonuclease P minimal substrates and conformation of the nucleotide -1 at the cleavage position. AB - It is demonstrated that acceptor stem duplexes derived from native tRNAs which contain a three-nucleotide extension at the 5'-terminus of mature tRNA are minimal substrates for ribonuclease P from both Escherichia coli and Bacillus subtilis. Variants with a cytidine at position -1 are most efficiently processed whereas the G -1 variant represents a comparatively poor substrate. An A -1 acceptor stem variant is a slightly better substrate than the G -1 variant though generally distinctly less efficient than the C -1 duplex. This is in qualitative agreement with the frequency of the occurrence of the corresponding nucleotides at position -1 in natural substrates, which is highest for pyrimidines and least for G. NMR analyses of the corresponding acceptor stems reveal that the conformation of the nucleotides at position -1 correlates with the substrate preferences of Ribonuclease P: Whereas C -1 adopts a conformation characterized by a glycosidic angle in the anti range (close to high-anti), the G -1 is clearly in syn conformation, and that of A -1 is intermediate between high-anti and syn. The riboses of nucleotides -1 are in all cases predominantly 2'-endo puckered. PMID- 11258958 TI - Solution structure of the DNA-binding domain of TraM. AB - The solution structure of the DNA-binding domain of the TraM protein, an essential component of the DNA transfer machinery of the conjugative resistance plasmid R1, is presented. The structure has been determined using homonuclear 2 dimensional NMR spectroscopy as well as 15N labeled heteronuclear 2- and 3 dimensional NMR spectroscopy. It turns out that the solution structure of the DNA binding domain of the TraM protein is globular and dominantly helical. The very first amino acids of the N-terminus are unstructured. PMID- 11258960 TI - Kinetic and mechanistic studies of the NO*-mediated oxidation of oxymyoglobin and oxyhemoglobin. AB - The second-order rate constants for the reactions between nitrogen monoxide and oxymyoglobin or oxyhemoglobin, determined by stopped-flow spectroscopy, increase with increasing pH. At pH 7.0 the rates are (43.6 +/- 0.5) x 10(6) M(-1) x s(-1) for oxymyoglobin and (89 +/- 3) x 10(6) M(-1) x s(-1) for oxyhemoglobin (per heme), whereas at pH 9.5 they are (97 +/- 3) x 10(6) M(-1) x s(-1) and (144 +/- 3) x 10(6) M(-1) x s(-1), respectively. The rate constants for the reaction between oxyhemoglobin and NO* depend neither on the association grade of the protein (dimer/tetramer) nor on the concentration of the phosphate buffer (100-1 mM). The nitrogen monoxide-mediated oxidations of oxymyoglobin and oxyhemoglobin proceed via intermediate peroxynitrito complexes which were characterized by rapid scan UV/vis spectroscopy. The two complexes MbFe(III)OONO and HbFe(III)OONO display very similar spectra with absorption maxima around 500 and 635 nm. These species can be observed at alkaline pH but rapidly decay to the met-form of the proteins under neutral or acidic conditions. The rate of decay of MbFe(III)OONO increases with decreasing pH and is significantly larger than those of the analogous complexes of the two subunits of hemoglobin. No free peroxynitrite is formed during these reactions, and nitrate is formed quantitatively, at both pH 7.0 and 9.0. This result indicates that, as confirmed from protein analysis after reacting the proteins with NO* for 10 times, when peroxynitrite is coordinated to the heme of myoglobin or hemoglobin it rapidly isomerizes to nitrate without nitrating the globins in physiologically significant amounts. PMID- 11258959 TI - Kinetic analysis of the slow ionization of glutathione by microsomal glutathione transferase MGST1. AB - An important aspect of the catalytic mechanism of microsomal glutathione transferase (MGST1) is the activation of the thiol of bound glutathione (GSH). GSH binding to MGST1 as measured by thiolate anion formation, proton release, and Meisenheimer complex formation is a slow process that can be described by a rapid binding step (K(GSH)d = 47 +/- 7 mM) of the peptide followed by slow deprotonation (k2 = 0.42 +/- 0.03 s(-1). Release of the GSH thiolate anion is very slow (apparent first-order rate k(-2) = 0.0006 +/- 0.00002 s(-)(1)) and thus explains the overall tight binding of GSH. It has been known for some time that the turnover (kcat) of MGST1 does not correlate well with the chemical reactivity of the electrophilic substrate. The steady-state kinetic parameters determined for GSH and 1-chloro-2,4-dinitrobenzene (CDNB) are consistent with thiolate anion formation (k2) being largely rate-determining in enzyme turnover (kcat = 0.26 +/- 0.07 s(-1). Thus, the chemical step of thiolate addition is not rate-limiting and can be studied as a burst of product formation on reaction of halo-nitroarene electrophiles with the E.GS- complex. The saturation behavior of the concentration dependence of the product burst with CDNB indicates that the reaction occurs in a two-step process that is characterized by rapid equilibrium binding ( = 0.53 +/- 0.08 mM) to the E.GS- complex and a relatively fast chemical reaction with the thiolate (k3 = 500 +/- 40 s(-1). In a series of substrate analogues, it is observed that log k3 is linearly related (rho value 3.5 +/- 0.3) to second substrate reactivity as described by Hammett sigma- values demonstrating a strong dependence on chemical reactivity that is similar to the nonenzymatic reaction (rho = 3.4). Microsomal glutathione transferase 1 displays the unusual property of being activated by sulfhydryl reagents. When the enzyme is activated by N-ethylmaleimide, the rate of thiolate anion formation is greatly enhanced, demonstrating for the first time the specific step that is activated. This result explains earlier observations that the enzyme is activated only with more reactive substrates. Taken together, the observations show that the kinetic mechanism of MGST1 can be described by slow GSH binding/thiolate formation followed by a chemical step that depends on the reactivity of the electrophilic substrate. As the chemical reactivity of the electrophile becomes lower the rate determining step shifts from thiolate formation to the chemical reaction. PMID- 11258961 TI - Structural alterations and inhibition of unisite and multisite ATP hydrolysis in soluble mitochondrial F1 by guanidinium chloride. AB - The effect of guanidinium chloride (GdnHCl) on the ATPase activity and structure of soluble mitochondrial F1 was studied. At high ATP concentrations, hydrolysis is carried by the three catalytic sites of F1; this reaction was strongly inhibited by GdnHCl concentrations of <50 mM. With substoichiometric ATP concentrations, hydrolysis is catalyzed exclusively by the site with the highest affinity. Under these conditions, ATP binding and hydrolysis took place with GdnHCl concentrations of >100 mM; albeit at the latter concentration, the rate of hydrolysis of bound ATP was lower. Similar results were obtained with urea, although nearly 10-fold higher concentrations were required to inhibit multisite hydrolysis. GdnHCl inhibited multisite ATPase activity by diminishing the V(max) of the reaction without significant alterations of the Km for MgATP. GdnHCl prevented the effect of excess ATP on hydrolysis of ATP that was already bound to the high-affinity catalytic site. With and without 100 mM GdnHCl and 100 microM [3H]ATP in the medium, F1 bound 1.6 and 2 adenine nucleotides per F1, respectively. The effect of GdnHCl on some structural features of F1 was also examined. GdnHCl at concentrations that inhibit multisite ATP hydrolysis did not affect the exposure of the cysteines of F1, nor its intrinsic fluorescence. With 100 mM GdnHCl, a concentration at which unisite ATP hydrolysis was still observed, 0.7 cysteine per F1 became solvent-exposed and small changes in its intrinsic fluorescence of F1 were detected. GdnHCl concentrations on the order of 500 mM were required to induce important decreases in intrinsic fluorescence. These changes accompanied inhibition of unisite ATP hydrolysis. The overall data indicate that increasing concentrations of GdnHCl bring about distinct and sequential alterations in the function and structure of F1. With respect to the function of F1, the results show that at low GdnHCl concentrations, only the high affinity site expresses catalytic activity, and that inhibition of multisite catalysis is due to alterations in the transmission of events between catalytic sites. PMID- 11258962 TI - Oligomerization of NhaA, the Na+/H+ antiporter of Escherichia coli in the membrane and its functional and structural consequences. AB - Recently, a two-dimensional crystal structure of NhaA, the Na+/H+ antiporter of Escherichia coli has been obtained [Williams, K. A., Kaufer, U. G., Padan, E., Schuldiner, S. and Kuhlbrandt, W. (1999) EMBO J., 18, 3558-3563]. In these crystals NhaA exists as a dimer. Using biochemical and genetic approaches here we show that NhaA exists in the native membrane as a homooligomer. Functional complementation between the polypeptides of NhaA was demonstrated by coexpression of pairs of conditional lethal (at high pH in the presence of Na+) mutant alleles of nhaA in EP432, a strain lacking antiporters. Physical interaction in the membrane was shown between the His-tagged NhaA polypeptide which is readily affinity purified from DM-solubilized membranes with a Ni2+-NTA column and another which is not; only when coexpressed did both copurify on the column. The organization of the oligomer in the membrane was studied in situ by site-directed cross-linking experiments. Cysteine residues were introduced--one per NhaA--into certain loops of Cys-less NhaA, so that only intermolecular cross-linking could take place. Different linker-size cross-linkers were applied to the membranes, and the amount of the cross-linked protein was analyzed by mobility shift on SDS PAGE. The results are consistent with homooligomeric NhaA and the location of residue 254 in the interface between monomers. Intermolecular cross-linking of V254C caused an acidic shift in the pH profile of NhaA. PMID- 11258963 TI - In vitro and in vivo interactions of bisphenol A and its metabolite, bisphenol A glucuronide, with estrogen receptors alpha and beta. AB - The estrogenic activities of bisphenol A (BPA) and its major metabolite BPA glucuronide (BPA-G) were assessed in a number of in vitro and in vivo assays. BPA competed with [3H]-17beta-estradiol (E2) for binding to mouse uterine cytosol ER, a glutathione S-transferase (GST)-human ER D, E, and F domain fusion protein (GST hERalphadef) and full-length recombinant hERbeta. The IC(50) values for E2 were similar for all three receptor preparations, whereas BPA competed more effectively for binding to hERbeta (0.96 microM) than to either mouse uterine cytosol ER (26 microM) or GST-hERalphadef (36 microM). In contrast, BPA-G did not competitively displace [3H]E2 from any of the ER preparations. In MCF-7 cells transiently transfected with Gal4-hERalphadef or Gal4-hERbetadef, BPA induced reporter gene activity with comparable EC(50) values (71 and 39 microM, respectively). No significant induction of reporter gene activity was seen for BPA-G. Cotreatment studies showed that concentrations of (10 microM) BPA and BPA G did not antagonize E2-induced luciferase mediated through either Gal4 hERalphadef or Gal4-hERbetadef. In vivo, the uterotropic effect of gavage or subcutaneous (sc) administration of 0.002-800 mg of BPA/kg of body weight/day for three consecutive days was examined in immature rats. Dose-related estrogenic effects on the rat uterus were observed at oral doses of 200 and 800 mg/kg and at sc doses of 10, 100, and 800 mg/kg. These results demonstrate that BPA competes more effectively for binding to ERbeta, but induces ERalpha- and ERbeta-mediated gene expression with comparable efficacy. In contrast, BPA-G did not exhibit any in vitro estrogenic activity. In addition, there was a clear route dependency on the ability of BPA to induce estrogenic responses in vivo. PMID- 11258964 TI - Characterization of DNA damage at purine residues in oligonucleotides and calf thymus DNA induced by the mutagen 1-nitrosoindole-3-acetonitrile. AB - N-Nitrosoindoles can efficiently transfer the nitroso group to nucleophilic targets in isolated purine nucleotides, causing depurination, deamination, and the formation of a novel guanine analogue, oxanine [Lucas, L. T., Gatehouse, D., and Shuker, D. E. G. (1999) J. Biol. Chem. 274, 18319-18326]. To determine the likely biological relevance of these modification pathways, the reactivity of 1 nitrosoindole-3-acetonitrile (NIAN), a model 3-substituted N-nitrosoindole, with oligonucleotides and calf thymus DNA was examined at physiological pH and temperature. Reaction of NIAN with single-stranded oligonucleotides containing various guanine motifs resulted in the production of single-strand break products at guanine sites due to the formation of alkali-labile lesions. The number of lesions increased with NIAN concentration and incubation time. Modification of calf thymus DNA by NIAN resulted in depurination, which gave the corresponding purine bases, deamination coupled with depurination, which gave xanthine, and the formation of oxanine. The former pathway was clearly the most important, and all reaction products exhibited a dose-response relationship. Cytosine and thymine residues were inactive toward NIAN. Further studies revealed an additional product in NIAN-treated duplex DNA containing a CCGG motif that was characterized as an interstrand cross-link, the yield of which increased with increasing NIAN concentration. These results indicate that the transnitrosating ability of NIAN to modify purine residues is preserved at the macromolecular level, with guanine residues appearing to be a primary site of reaction. All of these modification processes are potentially mutagenic events if they occur in vivo. PMID- 11258965 TI - Synthesis, characterization, and comparative 32P-postlabeling efficiencies of 2,6 dimethylaniline-DNA adducts. AB - 2,6-Dimethylaniline (2,6-diMeA) is a ubiquitous environmental pollutant that is used in industry as a synthetic intermediate. It is also found in tobacco smoke and as a major metabolite of lidocaine. Although the potential carcinogenicity of 2,6-diMeA in humans is presently uncertain, this aromatic amine has been classified as a rodent carcinogen. In addition, it is known to form hemoglobin adducts in humans, which indicates a profile of metabolic activation similar to that of typical arylamine carcinogens. Like other aromatic amines, 2,6-diMeA has been shown to yield N-(deoxyguanosin-8-yl)-2,6-dimethylaniline (dG-C8-2,6-diMeA) as a major DNA adduct in vitro. In this study, we show that 2,6-diMeA yields an unusual pattern of DNA adducts. In addition to dG-C8-2,6-diMeA, we have isolated two new adducts, 4-(deoxyguanosin-N(2)-yl)-2,6-dimethylaniline (dG-N(2)-2,6 diMeA) and 4-(deoxyguanosin-O(6)-yl)-2,6-dimethylaniline (dG-O(6)-2,6-diMeA), from the reaction of N-acetoxy-2,6-dimethylaniline with deoxyguanosine. A similar reaction conducted with deoxyadenosine yielded 4-(deoxyadenosin-N(6)-yl)-2,6 dimethylaniline (dA-N(6)-2,6-diMeA). All four adducts were detected in DNA reacted with N-acetoxy-2,6-dimethylaniline, with the relative yields being 46% for dA-N(6)-2,6-diMeA, 22% for dG-N(2)-2,6-diMeA, 20% for dG-O(6)-2,6-diMeA, and 12% for dG-C8-2,6-diMeA. This product profile contrasts markedly with the usual pattern of adducts obtained with aromatic amines, where C8-substituted deoxyguanosine products typically predominate. We further analyzed the kinetics of the T(4) polynucleotide kinase (PNK)-catalyzed phosphorylation of the C8 and N(2) deoxyguanosine 3'-phosphate adducts from 2,6-diMeA. The kinetic parameters obtained with these two structurally different adducts are compared to those determined with the parent nucleotide (dG3'p), and with (+/-)-anti-10 (deoxyguanosin-N(2)-yl)-7,8,9-trihydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene 3' phosphate, the major adduct derived from the environmental pollutant benzo[a]pyrene. The results indicate that all the adducts were labeled with lower efficiencies than dG3'p, stressing the likely underestimation of adduct levels in typical 32P-postlabeling protocols. Nonetheless, the N(2) adducts derived from 2,6-diMeA and benzo[a]pyrene were both labeled with higher efficiencies than the C8 adduct derived from 2,6-diMeA, with the benzo[a]pyrene adduct being the best substrate for PNK. Thus, the data suggest that N(2) adducts from dG3'p are intrinsically better substrates than their C8 analogues for PNK, and that bulkier aromatic fragments may favor the enzyme-substrate interaction during the labeling step. PMID- 11258967 TI - Metabonomic characterization of genetic variations in toxicological and metabolic responses using probabilistic neural networks. AB - Current emphasis on efficient screening of novel therapeutic agents in toxicological studies has resulted in the evaluation of novel analytical technologies, including genomic (transcriptomic) and proteomic approaches. We have shown that high-resolution 1H NMR spectroscopy of biofluids and tissues coupled with appropriate chemometric analysis can also provide complementary data for use in in vivo toxicological screening of drugs. Metabonomics concerns the quantitative analysis of the dynamic multiparametric metabolic response of living systems to pathophysiological stimuli or genetic modification [Nicholson, J. K., Lindon, J. C., and Holmes, E. (1999) Xenobiotica 11, 1181-1189]. In this study, we have used 1H NMR spectroscopy to characterize the time-related changes in the urinary metabolite profiles of laboratory rats treated with 13 model toxins and drugs which predominantly target liver or kidney. These 1H NMR spectra were data reduced and subsequently analyzed using a probabilistic neural network (PNN) approach. The methods encompassed a database of 1310 samples, of which 583 comprised a training set for the neural network, with the remaining 727 (independent cases) employed as a test set for validation. Using these techniques, the 13 classes of toxicity, together with the variations associated with strain, were distinguishable to >90%. Analysis of the 1H NMR spectral data by multilayer perceptron networks and principal components analysis gave a similar but less accurate classification than PNN analysis. This study has highlighted the value of probabilistic neural networks in developing accurate NMR based metabonomic models for the prediction of xenobiotic-induced toxicity in experimental animals and indicates possible future uses in accelerated drug discovery programs. Furthermore, the sensitivity of this tool to strain differences may prove to be useful in investigating the genetic variation of metabolic responses and for assessing the validity of specific animal models. PMID- 11258966 TI - Identification of a reactive metabolite of terbinafine: insights into terbinafine induced hepatotoxicity. AB - Oral terbinafine treatment for superficial fungal infections of toe and fingernails is associated with a low incidence (1:45000) of hepatobiliary dysfunction. Due to the rare and unpredictable nature of this adverse drug reaction, the mechanism of toxicity has been hypothesized to be either an uncommon immunological or metabolically mediated effect. However, there is little evidence to support either mechanism, and toxic metabolites of terbinafine have not been identified. We incubated terbinafine with both rat and human liver microsomal protein in the presence of GSH and were able to trap an allylic aldehyde, 7,7-dimethylhept-2-ene-4-ynal (TBF-A), which corresponds to the N dealkylation product of terbinafine. TBF-A was also prepared synthetically and reacted with excess GSH to yield conjugates with HPLC retention times and mass spectra identical to those generated in the microsomal incubations. The major GSH conjugate, characterized by (1)H NMR, corresponds to addition of GSH in a 1,6 Michael fashion. There remains a second electrophilic site on this metabolite, which can bind either to a second molecule of GSH or to cellular proteins via a 1,4-Michael addition mechanism. Moreover, we demonstrated that the formation of the GSH conjugates was reversible. We speculate that this allylic aldehyde metabolite, formed by liver enzymes and conjugated with GSH, would be transported across the canalicular membrane of hepatocytes and concentrated in the bile. The mono-GSH conjugate, which is still reactive, could bind to hepatobiliary proteins and lead to direct toxicity. Alternatively, it could modify canalicular proteins and lead to an immune-mediated reaction causing cholestatic dysfunction. PMID- 11258968 TI - Detection and quantification of depurinated benzo[a]pyrene-adducted DNA bases in the urine of cigarette smokers and women exposed to household coal smoke. AB - Polycyclic aromatic hydrocarbons (PAH) are metabolized to electrophiles that can bind to DNA bases and destabilize the N-glycosyl bond, causing rapid depurination of the adducted bases. Recent studies support depurination of DNA as a mechanism central to the genesis of H-ras mutations in PAH-treated mouse skin. Depurinating adducts account for 71% of all DNA adducts formed in mouse skin treated with benzo[a]pyrene (BP). This study analyzed urine of cigarette smokers, coal smoke exposed women, and nonexposed controls for the presence and quantities of the depurinated BP-adducted DNA bases, 7-(benzo[a]pyren-6-yl)guanine (BP-6-N7Gua) and 7-(benzo[a]pyren-6-yl)adenine (BP-6-N7Ade). Since these adducted bases originate from reaction of the BP radical cation with double-stranded DNA and not with RNA or denatured DNA, their presence in urine is indicative of DNA damage. Urine samples were fractionated by a combination of SepPak extraction and reverse-phase HPLC, and then analyzed by tandem mass spectrometry and capillary electrophoresis with laser-induced fluorescence. BP-adducted bases were detected in the urine from three of seven cigarette smokers and three of seven women exposed to coal smoke, but were not detected in urine from the 13 control subjects. Concentrations were estimated to be 60-340 and 0.1-0.6 fmol/mg of creatinine equivalent of urine for coal smoke-exposed women (maximum possible BP intake of ca. 23 000 ng/day) and cigarette smokers (BP intake of ca. 800 ng/day), respectively, exhibiting a sensitive response to BP exposures. BP-6-N7Gua was present at ca. 20-300 times the concentration of BP-6-N7Ade in the urine of coal smoke-exposed women, but was not detected in the urine of cigarette smokers. This difference may be due to the remarkably different BP exposures experienced by the two groups of PAH-exposed individuals. These results justify more extensive studies of depurinated BP-adducted DNA bases as potential biomarkers of PAH associated cancer risk. PMID- 11258969 TI - Oxidations of N(omega)-hydroxyarginine analogues and various N-hydroxyguanidines by NO synthase II: key role of tetrahydrobiopterin in the reaction mechanism and substrate selectivity. AB - Oxidations of L-arginine 2, homo-L-arginine 1, their N(omega)-hydroxy derivatives 4 and 3 (NOHA and homo-NOHA, respectively), and four N-hydroxyguanidines, N(omega)-hydroxynor-L-arginine 5 (nor-NOHA), N(omega)-hydroxydinor-L-arginine 6 (dinor-NOHA), N-(4-chlorophenyl)-N'-hydroxyguanidine (8), and N-hydroxyguanidine (7) itself, by either NOS II or (6R)-5,6,7,8-tetrahydro-L-biopterin (BH4)-free NOS II, have been studied in a comparative manner. Recombinant BH4-free NOS II catalyzes the oxidation of all N-hydroxyguanidines by NADPH and O2, with formation of NO2(-) and NO3(-) at rates between 20 and 80 nmol min(-1) (mg of protein)(-1). In the case of compound 8, formation of the corresponding urea and cyanamide was also detected besides that of NO2(-) and NO3(-). These BH4-free NOS II-dependent reactions are inhibited by modulators of electron transfer in NOS such as thiocitrulline (TC) or imidazole (ImH), but not by Arg, and are completely suppressed by superoxide dismutase (SOD). They exhibit characteristics very similar to those previously reported for microsomal cytochrome P450 catalyzed oxidation of N-hydroxyguanidines. Both P450 and BH4-free NOS II reactions appear to be mainly performed by O2(.-) derived from the oxidase function of those heme proteins. In the presence of increasing concentrations of BH4, these nonselective oxidations progressively disappear while a much more selective monooxygenation takes place only with the N-hydroxyguanidines that are recognized well by NOS II, NOHA, homo-NOHA, and 8. These monooxygenations are much more chemoselective (8 being selectively transformed into the corresponding urea and NO) and are inhibited by Arg but not by SOD, as expected for reactions performed by the NOS Fe(II)-O2 species. Altogether, these results provide a further clear illustration of the key role of BH4 in regulating the monooxygenase/oxidase ratio in NOS. They also suggest a possible implication of NOSs in the oxidative metabolism of certain classes of xenobiotics such as N hydroxyguanidines, not only via their monooxygenase function but also via their oxidase function. PMID- 11258971 TI - The lethal interaction and formation of a lipophilic ternary complex between 2,4,5-trichlorophenol and the Cu(II)-bis(1,10-phenanthroline) complex. AB - When nonlethal levels of 2,4,5-trichlorophenol (0.2 mM) and the Cu(II)-bis(1,10 phenanthroline) complex [Cu(II)(OP)2] (0.1 microM) were combined, a remarkable synergistic cytotoxicity was observed as measured by the extent of bacterial inactivation. In contrast, no such synergism was observed for the combination of 2,4,5-trichlorophenol with the Cu(II)-bis(bathophenanthroline disulfonate) complex [Cu(II)(BPS)2] which has a chemical structure similar to Cu(II)(OP)2, except for the net charge. The synergism observed for 2,4,5-trichlorophenol and Cu(II)(OP)2 was found to be due to the neutralization of their opposite charge and formation of a lipophilic ternary complex which facilitated copper transport into the bacterial cells. PMID- 11258970 TI - Metabolism of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline in human hepatocytes: 2-amino-3-methylimidazo[4,5-f]quinoxaline-8-carboxylic acid is a major detoxification pathway catalyzed by cytochrome P450 1A2. AB - Metabolic pathways of the mutagen 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) remain incompletely characterized in humans. In this study, the metabolism of MeIQx was investigated in primary human hepatocytes. Six metabolites were characterized by UV and mass spectroscopy. Novel metabolites were additionally characterized by 1H NMR spectroscopy. The carcinogenic metabolite, 2-(hydroxyamino)-3,8-dimethylimidazo[4,5-f]quinoxaline, which is formed by cytochrome P450 1A2 (P450 1A2), was found to be transformed into the N(2)-glucuronide conjugate, N(2)-(beta-1-glucosiduronyl)-2-(hydroxyamino)-3,8 dimethylimidazo[4,5-f]quinoxaline. The phase II conjugates N(2)-(3,8 dimethylimidazo[4,5-f]quinoxalin-2-yl)sulfamic acid and N(2)-(beta-1 glucosiduronyl)-2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, as well as the 7 oxo derivatives of MeIQx and N-desmethyl-MeIQx, 2-amino-3,8-dimethyl-6-hydro-7H imidazo[4,5-f]quinoxalin-7-one (7-oxo-MeIQx), and 2-amino-6-hydro-8-methyl-7H imidazo[4,5-f]quinoxalin-7-one (N-desmethyl-7-oxo-MeIQx), thought to be formed exclusively by the intestinal flora, were also identified. A novel metabolite was characterized as 2-amino-3-methylimidazo[4,5-f]quinoxaline-8-carboxylic acid (IQx 8-COOH), and it was the predominant metabolite formed in hepatocytes exposed to MeIQx at levels approaching human exposure. IQx-8-COOH formation is catalyzed by P450 1A2. This metabolite is a detoxication product and does not induce umuC gene expression in Salmonella typhimurium strain NM2009. IQx-8-COOH is also the principal oxidation product of MeIQx excreted in human urine [Turesky, R., et al. (1998) Chem. Res. Toxicol. 11, 217-225]. Thus, P450 1A2 is involved in both the metabolic activation and detoxication of this procarcinogen in humans. Analogous metabolism experiments were conducted with hepatocytes of untreated rats and rats pretreated with the P450 inducer 3-methylcholanthrene. Unlike human hepatocytes, the rat cell preparations did not produce IQx-8-COOH but catalyzed the formation of 2-amino-3,8-dimethyl-5-hydroxyimidazo[4,5-f]quinoxaline as a major P450 mediated detoxication product. In conclusion, our results provide evidence of a novel MeIQx metabolism pathway in humans through P450 1A2-mediated C(8)-oxidation of MeIQx to form IQx-8-COOH. This biotransformation pathway has not been detected in experimental animal species. Considerable interspecies differences exist in the metabolism of MeIQx by P450s, which may affect the biological activity of this mutagen and must be considered when assessing human health risk. PMID- 11258972 TI - Accumulation of the 1-methyl-4-phenylpyridinium ion in suncus (Suncus murinus) brain: implication for flavin-containing monooxygenase activity in brain microvessels. AB - The metabolism of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was examined in an effort to evaluate the role of flavin-containing monooxygenase (FMO) expressed in the brain of suncus (Suncus murinus) and rats. MPTP was metabolized to generate both 1-methyl-4-phenylpyridinium ion (MPP(+)) and MPTP N oxide by brain homogenates from rats. Although the level of MPP(+)-producing activity was similar in suncus and rats, a remarkable difference was found between the animal species in MPTP N-oxygenase activity, which was not detectable in brain homogenates from suncus. The concentrations of MPP(+) in suncus brain after a single ip administration of MPTP were markedly higher than that in rats, probably because of the lack of FMO activity in the suncus brain. The MPTP N oxygenase activity of microvessel homogenates of rat brain was 21-fold greater than that of whole brain homogenates. These results suggest that FMO(s) plays a significant role in the detoxification of MPTP in cerebral endothelial cells. PMID- 11258973 TI - Nitrogen dioxide as an oxidizing agent of 8-oxo-7,8-dihydro-2'-deoxyguanosine but not of 2'-deoxyguanosine. AB - The redox reactions of guanine and its widely studied oxidation product, the 8 oxo-7,8-dihydro derivative, are of significant importance for understanding the mechanisms of oxidative damage in DNA. Employing 2'-deoxyguanosine 5' monophosphate (dGMP) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) in neutral aqueous solutions as model systems, we have used nanosecond laser flash photolysis to demonstrate that neutral radicals, dGMP(-H)(*), derived by the one electron oxidation and deprotonation of dGMP, can oxidize nitrite anions (NO2(-)) to the nitrogen dioxide radical (*)NO2. In turn, we show that (*)NO2 can give rise to a one-electron oxidation of 8-oxo-G, but not of dGMP. The one-electron oxidation of dGMP was initiated by a radical cation generated by the laser pulse induced photoionization of a pyrene derivative with enhanced water solubility, 7,8,9,10-tetrahydroxytetrahydrobenzo[a]pyrene (BPT). The dGMP(-H)(*) neutral radicals formed via deprotonation of the dGMP(*)(+) radical cations and identified by their characteristic transient absorption spectrum (lambda(max) approximately 310 nm) oxidize nitrite anions with a rate constant of (2.6 +/- 0.3) x 10(6) M(-1) s(-1). The 8-oxo-dG is oxidized by (*)NO2 with a rate constant of (5.3 +/- 0.5) x 10(6) M(-1) s(-1). The 8-oxo-dG(-H)(*) neutral radicals thus generated are clearly identified by their characteristic transient absorption spectra (lambda(max) approximately 320 nm). The rate constant of 8-oxo-dG oxidation (k(12)) by the (*)NO2 one-electron oxidant (the (*)NO2/NO2(-) redox potential, E degrees approximately 1.04 V vs NHE) is lower than k(12) for a series of oxidizing aromatic radical cations with known redox potentials. The k(12) values for 8-oxo-dG oxidation by different aromatic radical cations derived from the photoionization of their parent compounds depend on the redox potentials of the latter, which were in the range of 0.8-1.6 V versus NHE. The magnitude of k(12) gradually decreases from a value of 2.2 x 10(9) M(-1) s(-1) (E degrees = 1.62 V) to 5.8 x 10(8) M(-1) s(-1) (E degrees = 1.13 V) and eventually to 5 x 10(7) M(-1) s(-1) (E degrees = 0.91 V). The implications of these results, including the possibility that the redox cycling of the (*)NO2/NO2(-) species can be involved in the further oxidative damage of 8-oxo-dG in DNA in cellular environments, are discussed. PMID- 11258974 TI - p53 Signaling and cell cycle checkpoints. PMID- 11258975 TI - Novel 2,5-hexanedione analogues. Substituent-induced control of the protein cross linking potential and oxidation susceptibility of the resulting primary amine derived pyrroles. AB - The neurotoxic gamma-diketone, 2,5-hexanedione (2,5-HD), induces neurofilamentous swellings at prenodal sites in proximal axons as a consequence of pyrrolation of lysine epsilon-amino groups on neurofilament proteins. However, there is disagreement as to whether pyrrole formation and the associated alteration of noncovalent interactions is sufficient to cause neurofilament accumulation, or whether pyrrole autoxidation and subsequent protein-protein cross-linking is an obligatory event. To investigate gamma-diketones that might form pyrroles inert to autoxidative-induced cross-linking, we synthesized 1,1,1-trifluoro-2,5 hexanedione, 3-(trifluoromethyl)-2,5-hexanedione (3-TFMHD), and two 3 (dialkylaminocarbonyl)-2,5-diketones and assessed their rates of pyrrole formation with amines, the oxidation susceptibility of the resulting pyrroles, and the protein cross-linking potential in vitro, relative to those of 3-methyl 2,5-hexanedione. 1,1,1-Trifluoro-2,5-hexanedione does not form pyrroles, but the three 2,5-HD analogues with an electron-withdrawing 3-substituent all rapidly formed pyrroles that were inert to autoxidation. Although 3-TMFHD nonetheless still induced cross-linking of ribonuclease A, by a nonoxidative mechanism independent of the pyrrole, the two 3-(dialkylaminocarbonyl)-2,5-diketones did not exhibit any protein cross-linking. As these two gamma-diketones possess aqueous-organic partitioning properties similar to those of 2,5-HD, they should serve as useful mechanistic probes in further studies. PMID- 11258976 TI - Nicotine and cotinine adducts of a melanin intermediate demonstrated by matrix assisted laser desorption/ionization time-of-flight mass spectrometry. AB - Pigmentation is a major factor in the incorporation of many drugs into hair. In an attempt to elucidate potential mechanisms of drug-melanin interaction, melanin was synthesized in vitro in the presence of nicotine, which we have shown to have a substantial interaction with melanin, and cotinine, a primary nicotine metabolite. L-DOPA, a precursor of eumelanin, was oxidized and oligomerized with tyrosinase. Nicotine, cotinine, and/or their deuterated analogues were added to the oligomerization reaction mixture in a 10:1 L-DOPA:drug ratio. A black precipitate formed within 60 min. Aliquots were removed from the incubation mixture at 60, 120, and 360 min. MALDI-TOF MS determinations were carried out on each sample to provide a mean and standard error for the masses of interest. Internal calibration allowed accurate mass measurement of the products. A careful comparison of the spectra of samples prepared both with and without drug indicated the presence of masses corresponding to the protonated drug, melanin oligomers, and nicotine or continine adducts of the monomeric melanin intermediate dopaquinone (DOPAQ). Additional support for the presence of drug melanin adducts was provided by employing deuterated analogues of nicotine and L DOPA in the reaction and observing that the masses shifted accordingly. Structures of the adducts were further confirmed by select ion gating and postsource decay analysis. PMID- 11258977 TI - Structure-activity relationships for a large diverse set of natural, synthetic, and environmental estrogens. AB - Understanding structural requirements for a chemical to exhibit estrogen receptor (ER) binding has been important in various fields. This knowledge has been directly and indirectly applied to design drugs for human estrogen replacement therapy, and to identify estrogenic endocrine disruptors. This paper reports structure-activity relationships (SARs) based on a total of 230 chemicals, including both natural and xenoestrogens. Activities were generated using a validated ER competitive binding assay, which covers a 10(6)-fold range. This study is focused on identification of structural commonalities among diverse ER ligands. It provides an overall picture of how xenoestrogens structurally resemble endogenous 17beta-estradiol (E(2)) and the synthetic estrogen diethylstilbestrol (DES). On the basis of SAR analysis, five distinguishing criteria were found to be essential for xenoestrogen activity, using E(2) as a template: (1) H-bonding ability of the phenolic ring mimicking the 3-OH, (2) H bond donor mimicking the17beta-OH and O-O distance between 3- and 17beta-OH, (3) precise steric hydrophobic centers mimicking steric 7alpha- and 11beta substituents, (4) hydrophobicity, and (5) a ring structure. The 3-position H bonding ability of phenols is a significant requirement for ER binding. This contributes as both a H-bond donor and acceptor, although predominantly as a donor. However, the 17beta-OH contributes as a H-bond donor only. The precise space (the size and orientation) of steric hydrophobic bulk groups is as important as a 17beta-OH. Where a direct comparison can be made, strong estrogens tend to be more hydrophobic. A rigid ring structure favors ER binding. The knowledge derived from this study is rationalized into a set of hierarchical rules that will be useful in guidance for identification of potential estrogens. PMID- 11258978 TI - Development and application of a sensitive and rapid immunoassay for the quantitation of N7-methyldeoxyguanosine in DNA samples. AB - N7-Methyldeoxyguanosine (N7-MedG) in DNA is a biomarker of exposure to environmental and endogenous methylating agents and may be of use in epidemiological studies. To quantitate N7-MedG in human samples, a sensitive assay system that uses only small quantities of DNA (<10 microg) is required. To this end, polyclonal antibodies against the imidazole ring-opened form of N7-MedG have been used to develop a highly sensitive immunoslot blot (ISB) assay. The limit of detection of the assay is 0.10 micromol of N7-MedG/mol of deoxyguanosine (dG) using 1 microg of DNA per analysis. The method was optimized using in vitro methylated calf thymus DNA and then applied to a study of DNA methylation in liver and brain tissues of mice following a single iv dose of the antitumor agent Temozolomide. The amount of N7-MedG in both tissues was strictly proportional to dose over a range of 10-200 mg of Temozolomide/kg of body weight. The ISB assay was then validated using pyloric DNA of rats treated with N-methyl-N'-nitro-N nitrosoguanidine and DNA samples from human bladder tumors, for both of which N7 MedG levels had already been quantitated by an HPLC/(32)P-postlabeling method previously described. The results showed a high degree of correlation (r = 0.98) between the two assays. The ISB assay was then applied to a range of human samples. A series of peripheral blood mononuclear cell DNA samples from cancer patients following treatment with Temozolomide had levels of N7-MedG ranging from 0.22 to 320 micromol/mol of dG. DNA samples from colon carcinoma and normal colorectal mucosa from individuals not known to be exposed to methylating agents contained levels of 0.11-1.34 micromol of N7-MedG/mol of dG. The ISB assay offers the potential for the rapid and high-throughput analysis of DNA obtained from routine biopsies and blood samples, thus enabling the determination of the extent of human exposure to environmental and endogenous sources of methylating agents in large-scale biomonitoring studies. PMID- 11258979 TI - A kinetic study into the hydrolysis of the ochratoxins and analogues by carboxypeptidase A. AB - The hydrolyses of the ochratoxins and analogues by carboxypeptidase A were assessed. This was done by measuring the amount of phenylalanine formed with liquid chromatography coupled to tandem electrospray mass spectrometry. The kinetic data of ochratoxin A, ochratoxin B, and the synthetic bromo-ochratoxin B were compared to the values of a number of synthesized structure analogues, namely, ochratoxin A methyl ester, ochratoxin B methyl ester, N-(2 hydroxybenzoyl)phenylalanine, N-(5-chloro-2-hydroxybenzoyl)phenylalanine, N-(5 bromo-2-hydroxybenzoyl)phenylalanine, and N-(5-fluoro-2 hydroxybenzoyl)phenylalanine. The halogen-containing analogues had lower turnovers than their des-halo analogues. There are no substantial differences in the kinetic data between the different halogen-containing analogues. PMID- 11258980 TI - Arsenicals inhibit thioredoxin reductase in cultured rat hepatocytes. AB - Thioredoxin reductase (TR), an NADPH-dependent flavoenzyme that catalyzes the reduction of many disulfide-containing substrates, plays an important role in the cellular response to oxidative stress. Trivalent arsenicals, especially methyl As that contains trivalent arsenic (MAs(III)), are potent noncompetitive inhibitors of TR purified from mouse liver. Because MAs(III) is produced in the biomethylation of As, it was postulated that the extent of inhibition of TR in cultured rat hepatocytes would correlate with the intracellular concentration of methyl As. Exposure of cultured hepatocytes to inorganic As(III) (iAs(III)), MAs(III), or aurothioglucose (ATG, a competitive inhibitor of TR activity) for 30 min caused a concentration-dependent reduction in TR activity. The estimated IC(50) was >>100 microM for iAs(III), approximately 10 microM for ATG, and approximately 3 microM for MAs(III). In hepatocytes exposed to 1 microM MAs(III) for up to 24 h, the inhibition of TR activity was maximal ( approximately 40%) after exposure for 15 min. After exposure for 3 h [when most MAs(III) has been converted to dimethyl As (DMAs)], TR activity in these cells had returned to control levels. Notably, exposure of the cell to 50 microM DMAs(III) did not affect TR activity. In hepatocytes exposed to 10 microM iAs(III) for up to 24 h, the inhibition of TR activity was progressive; at 24 h, activity was reduced approximately 35%. Following exposure to iAs(III) or MAs(III), the extent of inhibition of TR activity correlated strongly with the intracellular concentration of MAs. Taken together, these results suggest that arsenicals formed in the course of cellular metabolism of As are potent inhibitors of TR activity. In particular, MAs(III), an intermediate in the metabolic pathway, is an especially potent inhibitor of TR. Hence, the capacity of cells to produce or consume the intermediates in the pathway for As methylation may be an important determinant of susceptibility to the toxic effects of As. PMID- 11258982 TI - Phase 2 metabolites of N-hydroxylated amidines (amidoximes): synthesis, in vitro formation by pig hepatocytes, and mutagenicity testing. AB - A pig hepatocyte culture system was used for phase 2 biotransformation studies in vitro. The viability of the cultured hepatocytes was characterized daily during cultivation by lactate dehydrogenase release into the supernatant and albumin synthesis of the cells. The metabolic activity of the hepatocyte cultures was measured by 7-ethoxycoumarin (ECOD) and 7-ethoxyresorufin O-deethylation (EROD). The viability and metabolic activity of these pig hepatocytes were preserved for several days by culturing the cells in a monolayer culture system. Besides the known reduction of N-hydroxylated benzamidine (benzamidoxime) (2) to benzamidine (1), glucuronidation and, to a much smaller extent, sulfation of 2 to benzamidoxime O-glucuronide (3) and benzamidoxime O-sulfate (4) by cultured pig hepatocytes were found. The analyses were performed using HPLC and LC/MS studies. For unequivocal assignment, the hitherto unknown metabolites 3 and 4 were synthesized and characterized by spectroscopic techniques. Examination of benzamidoxime O-glucuronide and benzamidoxime O-sulfate for mutagenicity by means of the Ames test revealed that both phase 2 conjugates have no mutagenic effects in the TA98 and TA100 strains. So the phase 2 conjugation of benzamidoxime is significant in terms of detoxification. PMID- 11258981 TI - Reactive nitrogen oxygen species metabolize N-acetylbenzidine. AB - A close association has been reported for certain types of cancers influenced by aromatic amines and infection/inflammation. Reactive nitric oxygen species (RNOS), components of the inflammatory response, are bactericidal and tumoricidal, and contribute to the deleterious effects attributed to inflammation on normal tissues. This study assessed the possible transformation of the aromatic amine N-acetylbenzidine (ABZ) by RNOS. RNOS were generated by various conditions to react with ABZ, and samples were evaluated by HPLC. Conditions which generate nitrogen dioxide radical (NO(2)(-) + myeloperoxidase + H(2)O(2), ONOO(-), and NO(2)(-) + HOCl) produced primarily a single new product termed 3' nitro-ABZ. The myeloperoxidase-catalyzed reaction with 0.3 mM NO(2)(-) was completely inhibited by 1 mM cyanide, and not effected by 100 mM chloride with or without 1 mM taurine. In contrast, conditions which generate N(2)O(3), such as spermine NONOate, did not produce 3'-nitro-ABZ, but rather two compounds termed 4'-OH-AABP and AABP. (1)H NMR and mass spectrometry identified 3'-nitro-ABZ as 3' nitro-N-acetylbenzidine, 4'-OH-AABP as 4'-OH-4-acetylaminobiphenyl, and AABP as 4 acetylaminobiphenyl. Human polymorphonuclear neutrophils incubated with [(3)H]ABZ and stimulated with beta-phorbol 12-myristate 13-acetate produced 3'-nitro-ABZ in the presence of NO(2)(-) (0.1-1 mM). Neutrophil 3'-nitro-ABZ formation was verified by mass spectrometry and was consistent with myeloperoxidase oxidation of NO(2)(-). The results demonstrate that ABZ forms unique products in the presence of nitrosating and nitrating RNOS, which could influence the carcinogenic process and serve as biomarkers for these reactive species. PMID- 11258983 TI - Simultaneous quantitation of N(2),3-ethenoguanine and 1,N(2)-ethenoguanine with an immunoaffinity/gas chromatography/high-resolution mass spectrometry assay. AB - We have previously described an immunoaffinity/gas chromatography/electron capture negative chemical ionization high-resolution mass spectrometry (IA/GC/ECNCI-HRMS) assay for quantitation of the promutagenic DNA adduct N(2),3 ethenoguanine (N(2),3-epsilonGua) in vivo. Here we present an expanded assay that allows simultaneous quantitation of its structural isomer, 1,N(2)-ethenoguanine (1,N(2)-epsilonGua), in the same DNA sample. 1,N(2)-epsilonGua and N(2),3 epsilonGua were purified together from hydrolyzed DNA using two immobilized polyclonal antibodies. GC/ECNCI-HRMS was used to quantitate the 3,5 bis(pentafluorobenzyl) (PFB) derivative of each adduct against an isotopically labeled analogue. Selected ion monitoring was used to detect the [M - 181](-) fragments of 3,5-(PFB)(2)-N(2),3-epsilonGua and 3,5-(PFB)(2)-[(13)C(4),(15)N(2)] N(2),3-epsilonGua and the [M - 201](-) fragments of 3,5-(PFB)(2)-1,N(2) epsilonGua and 3,5-(PFB)(2)-[(13)C(3)]-1,N(2)-epsilonGua. The demonstrated limits of quantitation in hydrolyzed DNA were 7.6 fmol of N(2),3-epsilonGua and 15 fmol of 1,N(2)-epsilonGua in approximately 250 microg of DNA, which corresponded to 5.0 N(2),3-epsilonGua and 8.7 1,N(2)-epsilonGua adducts/10(8) unmodified Gua bases, respectively. 1,N(2)-epsilonGua was found to be the predominant ethenoguanine adduct formed in reactions of lipid peroxidation products with DNA. The respective ratios of 1,N(2)-epsilonGua to N(2),3-epsilonGua were 5:1 and 38:1 when calf thymus DNA was treated with ethyl linoleate or 4-hydroxynonenal, respectively, under peroxidizing conditions. Only N(2),3-epsilonGua was detected in DNA treated with the vinyl chloride (VC) metabolite 2-chloroethylene oxide and in hepatocyte DNA from rats exposed to 1100 ppm VC for 4 weeks (6 h/day for 5 days/week). These data suggest that 1,N(2)-epsilonGua plays a minor role relative to N(2),3-epsilonGua in VC-induced carcinogenesis, but that 1,N(2)-epsilonGua may be formed to a larger extent from endogenous oxidative processes. PMID- 11258993 TI - Synthesis, crystal structure, spectral studies, and catechol oxidase activity of trigonal bipyramidal Cu(II) complexes derived from a tetradentate diamide bisbenzimidazole ligand. AB - A new benzimidazole-based diamide ligand-N,N'-bis(glycine-2- benzimidazolyl)hexanediamide (GBHA)-has been synthesized and utilized to prepare Cu(II) complexes of general composition [Cu(GBHA)X]X, where X is an exogenous anionic ligand (X = Cl(-), NO(3)(-), SCN(-)). The X-ray structure of one of the complexes, [Cu(GBHA)Cl]Cl.H(2)O.CH(3)OH, has been obtained. The compound crystallizes in the monoclinic space group C2/c with unit cell dimensions a = 26.464(3) A, b = 10.2210(8) A, c = 20.444(2) A, alpha = 90 degrees, beta = 106.554(7) degrees, gamma = 90 degrees, V= 5300.7(9) A(3), and Z = 8. To the best of our knowledge, the [Cu(GBHA)Cl]Cl.H(2)O.CH(3)OH complex is the first structurally characterized mononuclear trigonal bipyramidal copper(II) bisbenzimidazole diamide complex having coordinated amide carbonyl oxygen. The coordination geometry around the Cu(II) ion is distorted trigonal bipyramidal (tau = 0.59). Two carbonyl oxygen atoms and a chlorine atom form the equatorial plane, while the two benzimidazole imine nitrogen atoms occupy the axial positions. The geometry of the Cu(II) center in the solid state is not preserved in DMSO solution, changing to square pyramidal, as suggested by the low temperature EPR data g( parallel) > g( perpendicular) > 2.0023. All the complexes display a quasi-reversible redox wave due to the Cu(II)/Cu(I) reduction process. E(1/2) values shift anodically from Cl(-) < NO(3)(-) < SCN(-), indicating that the bound Cl(-) ion stabilizes the Cu(II) ion while the N-bonded SCN(-) ion destabilizes the Cu(II) state in the complex. When calculated against NHE, the redox potentials turn out to be quite positive as compared to other copper(II) benzimidazole bound complexes (Nakao, Y.; Onoda, M.; Sakurai, T.; Nakahara, A.; Kinoshita, L.; Ooi, S. Inorg. Chim. Acta 1988, 151, 55. Addison, A. W.; Hendricks, H. M. J.; Reedijk, J.; Thompson, L. K. Inorg. Chem. 1981, 20 (1), 103. Sivagnanam, U.; Palaniandavar, M. J. Chem. Soc., Dalton Trans. 1994, 2277. Palaniandavar, M.; Pandiyan, T.; Laxminarayan, M.; Manohar, H. J. Chem. Soc., Dalton Trans. 1995, 457. Sakurai, T.; Oi, H.; Nakahara, A. Inorg. Chim. Acta 1984, 92, 131). It is therefore concluded that binding of amide carbonyl oxygen destabilizes the Cu(II) state. The complex [Cu(II)(GBHA)(NO(3))](NO(3)) could be successfully reduced by the addition of dihydroxybenzenes to the corresponding [Cu(I)(GBHA)](NO(3)). (1)H NMR of the reduced complex shows slightly broadened and shifted (1)H signals. The reduction of the Cu(II) complex presumably occurs with the corresponding 2e(-) oxidation of the quinol to quinone. Such a conversion is reminiscent of the functioning of a copper-containing catechol oxidase from sweet potatoes and the met form of the enzyme tyrosinase. PMID- 11258999 TI - Methylation of (2-methylethanethiol-bis-3,5-dimethylpyrazolyl)methane zinc complexes and coordination of the resulting thioether: relevance to zinc containing alkyl transfer enzymes. AB - A series of zinc complexes using a new tripodal, N(2)S, heteroscorpionate ligand (L3SH) that is isostructural and isoelectronic with the well-known N(3) trispyrazolylborates have been methylated in solution and the coordination properties of the resulting thioether examined. This system models the reactivity of zinc-containing enzymes involved in alkyl group transfers such as the DNA repair protein Ada from E. coli, or farnasyl transferase where it has been shown that the thioether resulting from alkyl group transfer remains in the coordination sphere of the zinc. The following complexes have been structurally characterized: [(L3S)ZnI] (1), [(L3SCH(3))ZnI(2)] (2), [(L3SCH(3))ZnI]BF(4) (3), [(L3SCH(3))Zn-mu-bis-acetato-mu-hydroxo-Zn(L3SCH(3))]BF(4) (5), [(L3SCH(3))ZnSPh(F5)]ClO(4) (7), and [(L3SCH(3))(2)Zn](BF(4))(2) (8). Complexes 3, 4, 5, 7, and 8 all display thioether coordination. Thus in the absence of superior anionic ligands, thioethers are reasonably good donors to zinc in either a tetrahedral or octahedral geometry. The methylation of the complex [(L3S)ZnSPh(F5)], which contains two different thiols, produces a single product, 7, where only the aliphatic thiol has been alkylated. This observation validates the suggestion that reactivity in enzymes with multiple zinc-bound thiols could be controlled by differences in thiol pK(a) (Hammes, B. S.; Warthen, C. R.; Crans, D.; Carrano, C. J. J. Biol. Inorg. Chem. 2000, 6, 82. Compound 7 is also of interest in that it resembles the metal ion-binding site of the blue copper protein, azurin. PMID- 11259012 TI - Cs(2) fixation by carbonic anhydrase model systems-a new substrate in the catalytic cycle. AB - The conversion of CS(2) with common carbonic anhydrase model systems has been studied using Hartree-Fock and density-functional theory methods employing the 6 311+G basis set. The calculated geometries and energetical parameters for [L(3)ZnOH](+)/CS(2) model systems (L = NH(3), imidazole) are compared with those obtained previously for the CO(2) hydration. While the same reaction mechanism applies for both heterocumulenes, the hypothetical conversion of CS(2) to give [L(3)ZnSC(O)SH](+) is characterized by a higher barrier and is much more exothermic than the corresponding CO(2) reaction cascade. Due to the increased number of heteroatoms, additional intermediates and product structures (compared with those involved in the CO(2) conversion) must be taken into account and have been analyzed in detail. The smaller electrophilicity of CS(2) is the reason for the higher activation energies, while the significantly increased exothermicity is due to the strong zinc(II)/sulfur interaction. The reversibility and therefore the existence of a catalytic cycle which could allow comparable CS(2) transformations must be questioned. Nevertheless, an interesting field of stoichiometric zinc-mediated CS(2) transformations is conceivable. PMID- 11259020 TI - Phthalazine-based dinucleating ligands afford dinuclear centers often encountered in metalloenzyme active sites. PMID- 11259026 TI - Synthesis and stereochemical elucidation of a 14-membered ring phosphonate. AB - [structure: see text]. We report the synthesis and stereochemical elucidation of a 14-membered ring phosphonate. The key step in the synthesis of the macrolide phosphonate was the cyclization of the acyclic precursor using the Mitsunobu reaction, a mild reaction for the preparation of mixed phosphonates. PMID- 11259027 TI - Approach to the synthesis of antitumor quassinoids from labdane diterpenes: an efficient synthesis of a picrasane-related intermediate. AB - [structure: see text]. The tetracyclic ketal 24, a suitable intermediate for the synthesis of antitumor pentacyclic quassinoids, has been efficiently prepared from communic acids (5a-c), via methyl ketone 9. The synthetic sequence from 9 to 24 consists of 15 steps in 12% overall yield. PMID- 11259028 TI - A novel palladium-mediated coupling approach to 2,3-disubstituted benzo(b)thiophenes and its application to the synthesis of tubulin binding agents. AB - [structure: see text]. Flexible, convergent access to 2,3-disubstituted benzo[b]thiophenes has been developed. The most concise approach involves sequential coupling of o-bromoiodobenzenes with benzylmercaptan and zinc acetylides to give benzyl o-ethynylphenyl sulfides which react with iodine to give 3-iodobenzo[b]thiophenes in a 5-endo-dig iodocyclization. These iodides can be further elaborated using palladium-mediated coupling and/or metalation techniques. This method has been applied to the synthesis of some novel tubulin binding agents. PMID- 11259029 TI - Unusually short distances between the carbonyl oxygen and the tin atom in RCOSMR'3 (M = Ge, Sn, Pb): the importance of intramolecular n(O) --> sigma*(MS) orbital interactions. AB - [structure: see text]. We found that the C=O.Sn distance in RCOSSnR'3 was shorter than the C=O.Ge distance in RCOSGeR'3 and theoretically confirmed that the orbital interactions between the nonbonding orbitals on the carbonyl oxygen (n(O)) and the sigma*(SnS) orbitals were important in regard to the shortness. PMID- 11259030 TI - Convenient synthesis and diversification of dehydroalaninyl phosphinic peptide analogues. AB - [structure: see text]. Dehydroalaninyl phosphinic dipeptide analogues were synthesized, via an efficient tandem Arbuzov addition/allylic rearrangement, in high yields. The susceptibility of the conjugate system to 1,4 nucleophilic additions was investigated. C-Elongation of the dipeptides was performed, and the efficiency of 1,4 addition to the resulting acrylamidic moiety was evaluated. Derivatization of such phosphinic templates is a powerful approach for rapid access to large number of phosphinic pseudopeptides bearing various side chains in the P1' position. PMID- 11259031 TI - High enantioselectivities in an (E)-alkene epoxidation by catalytically active chromium salen complexes. Insight into the catalytic cycle. AB - [structure: see text]. The epoxidation of (E)-beta-methylstyrene mediated by an oxochromium salen complex yields the epoxide in 92% ee in stoichiometric mode, the highest ee yet reported for a metal-mediated epoxidation of an (E)-alkene. The effect of added donor ligands, previously substantial, has reached a ceiling with this complex. In catalytic mode a slightly reduced ee and higher yield is obtained, indicating both the presence of a second oxidation cycle and that the major oxidant reacts with its reduced form. PMID- 11259032 TI - Stereocontrolled syntheses of epimeric 3-aryl-6-phenyl-1-oxa-7 azaspiro(4.5)decane NK-1 receptor antagonist precursors. AB - [structure: see text]. Complementary stereoselective syntheses of individual C3 epimers of the NK-1 receptor antagonist precursor 1 have been developed. Both diastereomers were derived from the common intermediate 3; introduction of the 3S stereocenter in 1a was achieved through hydrogenation of an arylated dihydrofuran, whereas the corresponding stereogenic center in 1b was installed using a stereo- and regioselective alkene hydroarylation. PMID- 11259033 TI - A double ring closing metathesis reaction in the rapid, enantioselective synthesis of NK-1 receptor antagonists. AB - [structure: see text]. The NK-1 receptor antagonist 1 has been prepared in seven steps from phenylglycine methyl ester. The key steps are a double ring closing metathesis reaction of tetraene 7 to prepare spirocycle 6 and a reductive Heck reaction to introduce the aryl moiety. This latter reaction discriminates the olefins of compound 6 and proceeds in a highly regio- and stereoselective manner. PMID- 11259034 TI - Stereoselective synthesis of substituted gamma-butyrolactones by the [3+2] annulation of allylic silanes with chlorosulfonyl isocyanate: enantioselective total synthesis of (+)-blastmycine. AB - [structure: see text]. A stereoselective synthesis of gamma-butyrolactones by the [3 + 2] annulation of allylic silanes with N-chlorosulfonyl isocyanate (CSI) was developed. An enantioselective total synthesis of (+)-blastmycinone was accomplished using this annulation as the key step. PMID- 11259035 TI - Enantio- and diastereocontrolled synthesis of (-)-iridolactone and (+) pedicularis-lactone. AB - [structure: see text]. Two iridoid lactones, (-)-iridolactone and (+)-pedicularis lactone, have been synthesized in an enantio- and diastereocontrolled manner starting from a tricyclic chiral building block serving as a synthetic equivalent of chiral 3-(hydroxymethyl)cyclopenta-2,4-dien-1-ol. PMID- 11259036 TI - Efficient asymmetric epoxidation of alpha,beta-unsaturated ketones using a soluble triblock polyethylene glycol-polyamino acid catalyst. AB - [reaction: see text]. Polyethylene glycol (PEG)-bound poly-L-leucine acts as a THF-soluble catalyst for the Julia-Colonna asymmetric epoxidation of enones. Excellent enantioselectivities may be obtained even with short chain length polyleucine. FT-IR investigations have determined that the catalytically active polyleucine components of these copolymers have an alpha-helical structure. PMID- 11259037 TI - Intramolecular glycosylation under neutral conditions for synthesis of 1,4-linked disaccharides. AB - [structure: see text]. A new method for intramolecular glycosylation, in which the donor and acceptor were linked via a 3,5-dinitrosalicylic acid derivative, was developed. Simply dissolving the tethered glycoside in CH3NO2 and warming to 40-60 degrees C led to formation of 1,4-linked disaccharides under neutral, hence, exceptionally mild, conditions. PMID- 11259038 TI - Synthesis, crystal structure, and thermolysis of the first tetracoordinate 1lambda4,2-selenazetidines: aziridine formation reaction from a four-membered heterocycle bearing highly coordinate selenium. AB - [structure: see text]. The first tetracoordinate 1lambda4,2-selenazetidines were synthesized by taking advantage of the Martin ligand and characterized by X-ray crystallographic analysis. The selenazetidines gave the corresponding aziridine and the cyclic selenenate on their thermolysis, which indicates the possibility that a highly coordinate 1,2-heterachalcogenetane may provide the corresponding heteracyclopropane regardless of the heteroatoms. PMID- 11259039 TI - Solution structure of (+)-discodermolide. AB - [structure: see text]. The solution structure of (+)-discodermolide (1) has been determined via 1- and 2-D NMR techniques in conjunction with Monte Carlo conformational analysis. Taken together, the results demonstrate that in solution (+)-discodermolide occupies a helical conformation remarkably similar to the solid state conformation. PMID- 11259040 TI - Snapshots of titanium BINOLate complexes with diverse solid state structures. AB - [structure: see text]. Many important asymmetric reactions are catalyzed by (BINOLate)Ti species with unknown structures. Reported here are three structures of BINOLate titanium complexes that show an interesting aggregation of (BINOLate)Ti(OiPr)2 with itself and with titanium tetraisopropoxide. These complexes are potential intermediates in the asymmetric addition of alkyl groups to aldehydes. PMID- 11259042 TI - Total synthesis of the immunosupressant (-)-pironetin (PA48153C). AB - [reaction: see text]. Total synthesis of the immunosuppresant pironetin has been achieved by a synthetic route in which the connections between starting materials and the desired structure are readily discerned. Key steps include a diastereoselective Lewis acid mediated crotylstannane aldehyde addition, a highly selective Lewis acid promoted Mukaiyama aldol reaction, an anti-selective SmI2 reduction of a beta-hydroxyketone, and finally a lactone annulation reaction. PMID- 11259041 TI - An approach to the skeleton of the securinega alkaloids. The total synthesis of (+/-)-securinine. AB - [structure: see text]. The concise total synthesis of securinine in nine steps from readily available starting materials is described. Key steps of the synthesis include an addition of a silyloxyfuran to an in situ generated iminium ion and a novel ring closing metathesis reaction. PMID- 11259043 TI - Asymmetric syntheses of fused bicyclic compounds by conjugate additions of allylic organolithium species to activated olefins and subsequent cyclizations. AB - [reaction: see text]. Addition of the configurationally stable organolithium species produced by enantioselective deprotonation of N-Boc-N-(p-methoxyphenyl) allylamines to alpha,beta-unsaturated carbonyl compounds affords 1,4-addition products in good yields with high diastereomeric and enantiomeric ratios. Further transformations of these compounds provide [3.3.0]-, [4.3.0]-, [5.3.0]-, and [5.4.0]-carbocycles and heterocycles with high stereoselectivities. PMID- 11259044 TI - Electronic probing of ketone catalysts for asymmetric epoxidation. Search for more robust catalysts. AB - [structure: see text]. Ketones with a fused oxazolidinone were synthesized and investigated to determine the electronic effect of the substituents at alpha positions of ketone catalysts on the Baeyer-Villiger oxidation and catalytic properties for asymmetric epoxidation. These new ketones give high yields and ee's with only 1-5 mol % catalyst. The current studies further show that the electronic effect is very important for the ketone-catalyzed epoxidation. PMID- 11259045 TI - A novel, selective, and efficient route to carotenoids and related natural products via Zr-catalyzed carboalumination and Pd- and Zn-catalyzed cross coupling. AB - [structure: see text]. A highly efficient and stereoselective protocol for the syntheses of symmetrical and unsymmetrical carotenoids involving Zr-catalyzed carboalumination of conjugated oligoenynes and Pd- and Zn-catalyzed alkenyl alkenyl coupling has been developed and applied to the syntheses of beta- and gamma-carotene and vitamin A. gamma-Carotene of > or =99% isomeric purity was prepared in three linear steps (five steps overall) from beta-ionone, enyne 8, (E)-ICH=CHBr, and (E)-Me3SiC triple bond CCH=CHBr in 32% overall yield. PMID- 11259046 TI - Tandem hetero diels-alder reaction: synthesis of oxygenated macrocycles. AB - [structure: see text]. C2-symmetric, oxygenated macrocycles have been synthesized from simple acyclic precursors in a single step by a tandem hetero Diels-Alder reaction. Thermolysis and Lewis acid catalysis can effect this novel transformation. The tandem reaction product is formed in preference to the intramolecular cycloaddition product, and an explanation for this result is proposed. PMID- 11259047 TI - One-pot direct conversion of 2,3-epoxy alcohols into enantiomerically pure 4 hydroxy-4,5-dihydroisoxazole 2-oxides. AB - [structure: see text]. A new methodology for the one-pot direct conversion of 2,3 epoxy alcohols into enantiomerically pure 4-hydroxy-4,5-dihydroisoxazole 2-oxides 1 has been found. The reaction works at room temperature and can be run at the 5 10 g scale. The mixture of 4,5-cis and 4,5-trans isomers obtained can be separated as such or as the bis-TDS ethers. A preliminary example of reductive cleavage of 1 to the corresponding amino polyol is also reported. PMID- 11259048 TI - Stereoselective transformations on D-glucose-derived eight-membered ring carbocycles. AB - [structure: see text]. The cyclooctenol derivative 1 can be transformed into the nine-membered ring lactone 3, as well as the amino-containing carbocycles 4 and 5. The corresponding ketone 2 gives access to the conformationally locked azasugar 6. PMID- 11259049 TI - Ruthenium complex-catalyzed anti-Markovnikov hydration of terminal alkynes. AB - [reaction: see text]. Highly regioselective, efficient, and substituent-tolerant anti-Markovnikov hydration of terminal alkynes occurs to give n-aldehyde by use of a catalytic amount of easily available cyclopentadienylruthenium complexes bearing appropriate bidentate or monodentate phosphine ligands. Typically, RuCpCl(dppm) (1 mol %) catalyzes the addition of water to 1-hexyne at 100 degrees C to give hexanal in 95% yield: 2-hexanone is not detected at all. PMID- 11259050 TI - Enantioselective synthesis of epigallocatechin-3-gallate (EGCG), the active polyphenol component from green tea. AB - [reaction: see text]. Enantioselective synthesis of epigallocatechin-3-gallate (EGCG, 3b), the active polyphenol component from green tea, has been achieved by using a stereospecific cyclization of the Sharpless asymmetric dihydroxylation product 7c as the key step. PMID- 11259051 TI - Mesyloxy-group migration as the stereoselective preparation method of various functionalized olefins and its reaction mechanism. AB - [structure: see text]. It was demonstrated that mesylation of appropriate gamma,gamma-difluorinated allylic alcohols under usual conditions furnished the corresponding alpha,alpha-difluorinated allylic mesylates, possibly by way of 1,3 mesyloxy-group migration after formation of the expected "normal" intermediates, gamma,gamma-difluorinated allylic mesylates. This rearrangement was conveniently applied to the construction of trisubstituted allylic alcohols, alpha,beta unsaturated esters, amides, or ketones in good to excellent chemical yields with exclusive E selectivities. PMID- 11259052 TI - A new phenoxyacetate-based linker system for the solid-phase synthesis of oligosaccharides. AB - [structure: see text]. A novel linker system has been designed, and its first application to solid-phase oligosaccharide synthesis is described. The use of the highly reactive o-nitro-phenoxyacetate linker allows a fast and quantitative cleavage using mild basic conditions. This method combined with the trichloroacetimidate glycosylation exhibits highly promising results as demonstrated for the synthesis of tetrasaccharide 1 (n = 3) containing glucose beta(1 --> 4) and beta(1 --> 6) linkages. PMID- 11259053 TI - Studies toward gymnodimine: development of a single-pot Hua reaction for the synthesis of highly hindered cyclic imines. AB - [structure: see text]. In studies directed toward gymnodimine and related marine toxins, a single-pot variation of the Hua cyclic imine synthesis has been developed. The reaction involves generation of N-trimethylsilyl lactams in situ followed by alkyllithium addition leading directly to cyclic imines. Importantly, this reaction proceeds efficiently with highly hindered alpha,alpha-dialkyl lactams, provided 1,2-dimethoxyethane (DME) is used as solvent, leading to stable cyclic imines. Overall, this transformation allows a one-pot coupling of an alkyliodide and a lactam to give a cyclic imine. PMID- 11259054 TI - Efficient stereochemical relay en route to leucascandrolide A. AB - [structure: see text]. A complete relay of the initial stereochemical information is central to the efficient and highly stereocontrolled construction of the C1 C15 fragment of the marine macrolide leucascandrolide A. Cyclic silane 3, assembled via Pt-catalyzed hydrosilylation, was designed to serve as a temporary template for the installation of the C12 stereogenic center. The strategy features a highly convergent C10-C11 bond construction via 1,5-anti-selective aldol reaction and rapid assembly of the trisubstituted pyran subunit via Prins desymmetrization. PMID- 11259055 TI - Masked oxo sulfinimines (N-sulfinyl imines) in the asymmetric synthesis of proline and pipecolic acid derivatives. AB - [structure: see text]. On addition of Et2AlCN/i-PrOH, masked oxo sulfinimines give alpha-amino nitriles that afford oxo alpha-amino acids on hydrolysis. These amino acids cyclize and are reduced to cis proline and cis pipecolic acids derivatives in high ee and good yield. This new procedure avoids many of the limitations related to the preparation of oxo amino acids from proteinogenic amino acids. PMID- 11259056 TI - Addition reaction of dialkyl disulfides to terminal alkynes catalyzed by a rhodium complex and trifluoromethanesulfonic acid. AB - [reaction: see text]. Addition of dialkyl disulfides to terminal alkynes is catalyzed by a rhodium-phosphine complex and trifluoromethanesulfonic acid giving (Z)-bis(alkylthio)olefins stereoselectively. PMID- 11259057 TI - Total synthesis of FR901483. AB - [structure: see text]. The total synthesis of FR901483, a structurally novel immunosuppressant, has been accomplished by the use of technology recently developed in this laboratory for the oxidative cyclization of phenolic oxazolines to spirolactams. Our approach may reflect the biosynthetic pathway leading to the natural product. PMID- 11259058 TI - Synthesis and reactivity of N-acyl-5- (1-hydroxyalkyl)-2,3-dihydro-4-pyridones. AB - [structure: see text]. The Nozaki-Hiyama-Kishi reaction was used to prepare the 5 (1-hydroxyalkyl)-2,3-dihydro-4-pyridones 3. Reduction, oxidation, and substitution reactions of 3 were examined. PMID- 11259059 TI - Asymmetric synthesis of beta-substituted alpha-methyl-beta-amino esters by Mannich reaction of (S,S)-(+)-pseudoephedrine acetamide derived enolate with imines. AB - [reaction: see text]. The reaction of an (S,S)-(+)-pseudoephedrine acetamide based enolate with several imines afforded smoothly and with full stereochemical control a series of beta-substituted alpha-methyl-beta-aminoamides that upon hydrolysis/esterification afforded the corresponding beta-aminoester derivatives in good yields and in almost enantiomerically pure form. PMID- 11259060 TI - Transfer of stereochemical information in a minimal self-replicating system. AB - [structure: see text]. The rational design, synthesis, and characterization of a minimal self-replicating system based on a 1,3-dipolar cycloaddition between a nitrone and a maleimide is presented. The importance of molecular recognition in this system is demonstrated using a competitive inhibitor. Doping experiments demonstrate that only one of the two diastereoisomeric products of the cycloaddition reaction is capable of acting as an efficient template for its own formation, accelerating the reaction between the nitrone and maleimide and controlling the stereochemical outcome of the reaction. PMID- 11259061 TI - Improved solid-phase peptide synthesis method utilizing alpha-azide-protected amino acids. AB - [structure: see text]. Pure alpha-azido acids were prepared using an efficient diazo transfer method followed by buffered workup. These building blocks were used to prepare small peptides on Wang resin by two approaches. Peptides prone to diketopiperazine formation were prepared in good yields by coupling acids to resin bound iminophosphoranes during Fmoc-Wang synthesis. The iminophosphoranes can also be hydrolyzed under neutral conditions to provide unprotected amines ready for further coupling. PMID- 11259062 TI - C-H and C-C bond activation of primary amines through dehydrogenation and transimination. AB - [structure: see text]. Dehydrogenation and subsequent transimination of primary amines offer a new pathway for C-H bond activation, ortho-alkylation, and C-C bond activation to afford a variety of ketones in the reaction of 1-alkene by a cocatalyt system of Rh(I) and 2-amino-3-picoline. PMID- 11259063 TI - GaCl3-promoted ethenylation of silylated beta-dicarbonyl compound with silylethyne. Synthesis of ethenylmalonate. AB - [reaction: see text]. In the presence of GaCl3, silyl enol ethers derived from alpha-substituted beta-ketoesters or malonates are ethenylated at the alpha carbon atom with trimethylsilylethyne in high yields. Ethenylmalonates can also be synthesized by this method. PMID- 11259064 TI - An efficient synthesis of 3-hetero-13,14-dihydro prostaglandin F1alpha analogues. AB - [structure: see text]. A new class of 3-hetero-13,14-dihydro prostaglandin F(1)(alpha) analogues was synthesized from a common intermediate. The latter was constructed via a two-step, three-component process. The lower chain, containing the 15-(phenoxymethyl) group, was synthesized in enantiopure form using Jacobsen's (salen)Co-catalyzed kinetic resolution of a terminal epoxide with phenol. PMID- 11259065 TI - Special issue on gay, lesbian and bisexual youth. PMID- 11259066 TI - A critique of research on sexual-minority youths. AB - Developmental scientists should seriously reconsider traditional empirical and theoretical paradigms that narrowly define sexual-minority adolescents in terms of those who adopt a culturally defined sexual identity label. A broader consideration of youth populations who have same-sex desires but who might not necessarily identify as gay, lesbian or bisexual, lead one to a very different understanding of sexual-minority youths than is apparent in most published studies. First, they are in most regards just like all other adolescents with similar developmental needs and concerns. Second, they are not a homogeneous group but vary among themselves in predictable ways. Third, this expanded definition allows us to conclude that same-sex attraction per se does not lead to pathology or to problematic behavior such as drug abuse, suicide, prostitution or HIV infection. Indeed, researchers and clinicians should focus on the resiliency that often characterizes sexual-minority youths. PMID- 11259067 TI - The quest for modern manhood: masculine stereotypes, peer culture and the social significance of homophobia. AB - This paper explores the use of homophobic terms by boys and young men and the meanings they invoke when using them. Highly detailed interviews were conducted with young men from diverse backgrounds about their own experiences while growing up and their observations of schools, teachers, family and peers. Homophobia was found to be more than a simple prejudice against homosexuals. Homophobic terms like "poofter" and "faggot" have a rich developmental history and play a central role in adolescent male peer-group dynamics. Homophobic terms come into currency in primary school. When this happens, words like poofter and faggot rarely have sexual connotations. Nevertheless, far from being indiscriminate terms of abuse, these terms tap a complex array of meanings that are precisely mapped in peer cultures, and boys quickly learn to avoid homophobia and to use it decisively and with great impact against others. Significantly, this early, very powerful use of homophobic terms occurs prior to puberty, prior to adult sexual identity and prior to knowing much, if anything, about homosexuality. An effect of this sequence is that early homophobic experiences may well provide a key reference point for comprehending forthcoming adult sexual identity formation (gay or not) because powerful homophobic codes are learned first. PMID- 11259068 TI - Naming the "outsider within": homophobic pejoratives and the verbal abuse of lesbian, gay and bisexual high-school pupils. AB - Few studies have looked explicitly at the use of homophobic pejoratives among young high-schoolers and-not always an easy group to access, nor a comfortable subject to discuss. In this study, 377 14 and 15 year olds listed the pejoratives they heard at school and identified the ones they considered most taboo. As some of the most vitriolic items reported, homophobic pejoratives accounted for 10 per cent of the 6000 items generated. Significantly, however, homophobic verbal abuse was rated much less seriously than either racist abuse or other taboo slang. Boys reported more homophobic pejoratives than girls, but rated them more seriously. As further evidence of the increasingly well-documented daily assault on the psychological health of young homosexual people, this study confirms the prevalence of homophobic verbal abuse in high schools, its particularly aggressive nature, and the relative disregard with which it is used. As a contribution from Language and Communication Research, directions are offered for both sex(uality) education and language education. PMID- 11259069 TI - Suicide and gay/lesbian/bisexual youth: implications for clinicians. AB - The research indicating the incidence rates and specific risks for suicide in the gay, lesbian, bisexual, and questioning (GLBQ) adolescent population is reviewed. An ecological model of suicide risk assessment for GLBQ youth is presented based on Bronfenbrenner's model of human development. The model argues for individual, micro, and macro levels of assessment to increase clinical judgement and accuracy in determining high risk GLBQ adolescents. The model also delineates both primary and secondary intervention strategies which could be utilized to prevent GLBQ youth suicide. PMID- 11259070 TI - "I was terrified of being different": exploring gay men's accounts of growing-up in a heterosexist society. AB - This study examined retrospective accounts of gay identity formation during adolescence. Twenty in-depth interviews were conducted with working-class gay men from a small town in the north of England. Themes salient to understanding their adolescent experiences of identity formation were identified: "defined by difference", "self-reflection and inner conflict", "alienation and isolation", "living a lie", "telling others", and "wholeness and integrity". We illustrate how the socio-cultural context of compulsory heterosexuality is central in understanding accounts of both reported minority stress and identity construction. The implications of the research for future interventions designed to tackle homophobia and heterosexism are discussed. PMID- 11259071 TI - Gendered (s)explorations among same-sex attracted young people in Australia. AB - This paper seeks to import a more complex understanding of gendered subjectivity into discussions of young people and homosexuality, and is based on an Australian national survey (n=749) of same-sex attracted youth (SSAY) aged between 14 and 21. Results revealed significant gender differences with regard to patterns of sexual attraction, behaviour and identity labels among participants. For the young men in the study, there was more congruence between feelings of gender a typicality, same-sex attractions and same-sex behaviours. Overall, young women displayed more fluidity with regard to their sexual feelings, behaviours and identities. Young women were more likely to be engaged in private explorations of lesbianism, concurrent with participation in heterosexual sex and relationships. Young women were also grappling with more limited and emotionally risky opportunities for sex with other girls who were already known to them as friends. The invisibility of lesbianism as an identity or practice led to confusion about what feelings meant for the future in the arena of lived experience. The paper concludes that more research is needed into the impact of gender on the development of young people's experiences of homosexuality, particularly the manner in which invisibility and lack of social acceptance of a full spectrum of sexual diversity may disadvantage young women's emotional health and well-being. PMID- 11259072 TI - Benefits of cross-sexual orientation friendships among adolescent females. AB - The present study investigates benefits of cross-sexual orientation friendships in adolescent girls. Interviews were conducted with 20 participants in close friendships pairs. Participants included 10 heterosexual, five lesbian and five bisexual individuals. Participants ranged in age from 19-25 at the time of the interview and 12-23 at the onset of their friendship. Results of this study indicate that cross-sexual orientation friendships function similarly in comparison to other friendships, and that issues of sexual orientation can impact friendship development in positive ways. Overall benefits of cross-sexual orientation friendships are discussed, as are benefits that are unique to both heterosexual and lesbian/bisexual youth. PMID- 11259073 TI - Heterosexism and homophobia in Scottish school sex education: exploring the nature of the problem. AB - This paper focuses on instances of heterosexism and homophobia, as well as examples of good practice in sex education, observed in 25 Scottish schools. Possible reasons for why good practice is not more widespread are explored. Qualitative and quantitative data are presented. Various constraints to good practice are identified by teachers, and are discussed in the context of survey data on pupil attitudes towards same-sex sexual relationships, and on teacher confidence in teaching about homosexuality. PMID- 11259074 TI - School outcomes of sexual minority youth in the United States: evidence from a national study. AB - Using data from the Add Health Study, the first nationally representative study of adolescents in the U.S. to include information on same-sex romantic attraction, we examine school outcomes (school troubles, attitudes, and performance) of same-sex attracted youth within the context of four relational domains: family, teacher, social, and peer. Results indicate that each domain plays a role in the negative attitudes about school held by these sexual minority youth. However, sexual minority youths' feelings about their teachers play an important role in explaining school troubles. PMID- 11259075 TI - Playing it safe: addressing the emotional and physical health of lesbian and gay pupils in the U.K. AB - Compared to young people in general, young lesbians and gay men can face specific challenges to their physical and emotional well-being. These include discrimination, victimization, homophobic bullying and an elevated suicide risk. Relative to initiatives which attempt to address bullying in general, little has been done in schools in the U.K. to address physical and verbal homophobic bullying. This paper reports on an exploratory study to examine teachers' perceptions of homophobic bullying, the responses made to this form of bullying, and the factors which impact on the provision of education and support to lesbian and gay pupils. Findings suggested that teachers were aware of homophobic bullying but were confused, unable or unwilling to address the needs of lesbian and gay pupils. Implications for policy, practice and research are discussed. While current U.K. Government policy promoting Healthy Schools and Citizenship education offers hope for the future, much remains to be done to ensure that such initiatives are inclusive of all pupils. PMID- 11259076 TI - It's time for psychoneuroimmunology. AB - It is intuitively obvious that the mind and the body are joined in ways that are not yet understood. The mission of the PsychoNeuroImmunology Research Society (PNIRS) is to delineate these relationships, to try to understand their connections at the molecular level and to use this knowledge to prevent and relieve human pain and suffering. Members of our Society have already made substantial and important contributions toward accomplishing these goals. For example, regulation of the neuroendocrine system by proinflammatory cytokines and development of the concept of sickness behavior have now become established and well-accepted tenets in psychoneuroimmunology. Although we possess some of the research tools that are needed to accomplish our goals, we need more. We must continue to apply new information that is constantly being generated in the biological sciences, such as what may be found in the recently completed mapping and sequencing of the human genome. There will always be fundamental discoveries that can and should be used to advance the field of psychoneuroimmunology and to help us accomplish our mission. Our research is needed to minimize human afflictions and to learn how patients can better participate in their own health management. That is why the time for psychoneuroimmunology is now. PMID- 11259077 TI - Cytokine-induced sickness behavior: where do we stand? AB - Sickness behavior refers to the coordinated set of behavioral changes that develop in sick individuals during the course of an infection. At the molecular level, these changes are due to the effects of proinflammatory cytokines, such as interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNFalpha), in the brain. Peripherally released cytokines act on the brain via a fast transmission pathway involving primary afferent nerves innervating the body site of inflammation and a slow transmission pathway involving cytokines originating from the choroid plexus and circumventricular organs and diffusing into the brain parenchyma by volume transmission. At the behavioral level, sickness behavior appears to be the expression of a central motivational state that reorganizes the organism's priorities to cope with infectious pathogens. There is clinical and experimental evidence that activation of the brain cytokine system is associated with depression, although the exact relationship between sickness behavior and depression is still elusive. PMID- 11259078 TI - Behavioral effects of infection with IL-6 adenovector. AB - The onset of autoimmunity in lupus-prone mice is accompanied by a constellation of behavioral deficits, termed Autoimmunity-Associated Behavioral Syndrome (AABS). In particular, a spontaneous increase in serum interleukin-6 (IL-6) levels in five-week old MRL-lpr mice coincides temporally with blunted responsiveness to sucrose and excessive immobility in the forced swim test. These relationships, along with evidence that sucrose intake drops after systemic IL-6 overexpression is induced in healthy mice, have led to the hypothesis that sustained elevation in serum IL-6 also induces other aspects of AABS. This hypothesis is tested by comparing the behavioral profiles of healthy mice infected with Ad5mIL6 adenovirus (2 x 10(8) pfu of virus/mouse i.p.) with those of animals infected with control Ad5 virus. This methodology was used to achieve high circulating levels of IL-6, to overcome the problem of its short half-life, and to avoid the stressful effects of repeated injections. The Ad5mIL6 infection (known to induce excessive IL-6 levels over five days) transiently reduced food, water, and sucrose intake, as well as rectal temperature in MRL +/+ and AKR/J mice. Although the level of locomotor activity did not decline, Ad5mIL6-infected AKR/J mice demonstrated less novel object exploration. Performance in the step down, plus-maze, and spontaneous alternation tests were disturbed to various degrees in all infected animals. The present results suggest that prolonged exposure to circulating IL-6 primarily impairs ingestive behavior, likely reflecting enhanced catabolism. The inability of circulating IL-6 to alter other aspects of behavior supports the hypothesis that multiple immuno-neuroendocrine mechanisms contribute to the pathogenesis of AABS. PMID- 11259079 TI - Involvement of central, but not placental corticotropin releasing hormone (CRH) in heat stress induced immunosuppression during pregnancy. AB - To clarify whether corticotropin releasing hormone (CRH) and beta-endorphin (betaEP) system mediate maternal immunosuppression in pregnant rats exposed to heat through central or placental pathway, we examined the effects of intravenous (iv) (100 or 500 microg) or intracerebroventricular (icv) (5 microg) administration of CRH receptor antagonist alpha-helical CRH (9-41) on splenic natural killer cell activity (NKCA) as well as betaEP in blood, pituitary lobes, and placenta in pregnant rats at 15 to 16 days gestation. Two-way ANOVA revealed that heat reduced NKCA and elevated blood and pituitary betaEP but did not change placental betaEP. Iv administered 500 microg and icv administered alpha-helical CRH reversed the reduced NKCA and the elevated pituitary betaEP, while iv administration of 100 microg alpha-helical CRH did not. The increased blood betaEP was reversed by iv 100 and 500 microg alpha-helical CRH and icv administration. Both iv and icv administrations reduced placental betaEP independent of heat exposure. Thus, the response of placental betaEP to iv administration of alpha-helical CRH seemed to be stronger than that of pituitary betaEP. These results indicate that alpha-helical CRH which acts on pituitary betaEP antagonizes heat-induced immunosuppression during pregnancy, suggesting that immunosuppression produced by heat stress during pregnancy is mediated by the central CRH system. The placental CRH-betaEP system seems unlikely to be involved in the immunosuppression. Physiologic roles of placental CRH and opioid system should be clarified by future in vitro experiments using placenta specimen including placental immunocyte. PMID- 11259080 TI - Stress-induced changes in peripheral natural killer cell cytotoxicity in pigs may not depend on plasma cortisol. AB - The study examined cortisol (COR) involvement in stress-related changes in natural killer cell cytotoxicity (NKCC). The relationship between blood COR level, phasic changes in NKCC, and the number of large granular lymphocytes (LGL) was examined in pigs during the course of 4-h immobilization stress (IMB) and for 6 days after its termination. NKCC was determined using 18-h 51Cr-release assay, LGL number was assessed with a standard hematological method, and plasma COR level was measured by radioimmunoassay. The blood level of COR was increasing during IMB (max 446Delta% at the second hour) and decreased after its termination (max -59Delta% on day 2). Changes in NKCC level and LGL number were biphasic; i.e., an initial increase in both measures (NKCC max 24Delta%, LGL max 18Delta%) in an early phase of stress (0-1h) was followed by their subsequent decrease (NKCC max -35Delta%, LGL max -41Delta%) in the late phase (3-4 h) of stress, which persisted for several days after termination of IMB. Thus, in the early phase of stress, there was a positive correlation between NKCC, LGL number, and COR levels (all elevated); a positive correlation between the measures also occurred after termination of IMB (all decreased). A negative correlation between COR and NKCC, which might be indicative of COR-related immunosuppression, was found only in the late (3-4 h) phase of stress. It is concluded that COR may be only one of multiple factors (possibly antagonistic) determining an actual immune response during stress. PMID- 11259081 TI - Central CRH suppresses specific antibody responses: effects of beta-adrenoceptor antagonism and adrenalectomy. AB - Central corticotropin releasing hormone (CRH)-induced activation of the sympathetic nervous system and/or the pituitary-adrenal axis is hypothesized to mediate suppression of in vivo specific antibody responses. To test whether beta adrenergic receptor activation is involved in the immunosuppressive effects of central CRH, rats were pretreated with propranolol or saline before intracerebroventricular infusion of CRH and immunization with keyhole limpet hemocyanin (KLH). KLH (3 microg/kg) immunization induced significant increases in circulating levels of antigen-specific IgM and IgG. Central infusion of CRH (200 pmol) suppressed both IgM and IgG responses. Pretreatment with propranolol (20 mg/kg IP) reversed CRH-induced suppression of IgG responses but had no effect on IgM levels. To test whether adrenal activation also plays a role in the effects of KLH on specific antibody responses, a separate group of animals underwent adrenalectomy prior to CRH infusion and immunization with KLH. As compared to nonadrenalectomized control rats, adrenalectomized rats showed a reduction of antibody responses, and CRH failed to induce a further suppression of IgM or IgG responses in adrenalectomized animals. Collectively, these data suggest that beta adrenoceptors mediate the suppression of primary antibody responses induced by central CRH. Moreover, the adrenals may promote optimal primary antibody responses after exposure to physiological levels of antigen. PMID- 11259082 TI - Aberrant nuclear expression of AP-1 and NFkappaB in lymphocytes of women stressed by the experience of breast biopsy. AB - We have investigated the expression of AP-1 and NFkappaB in peripheral blood lymphocytes of women scheduled for breast biopsy. Samples were collected when women were informed of the need for biopsy (prebiopsy, T1, 5-7 days prior to the actual biopsy) and 7-10 days after they learned the result of their biopsy (postbiopsy, T2). At the time of blood collection, psychological stress was evaluated using Speilberger's State Trait Anxiety Inventory (STAI) and the Profile of Mood States (POMS). Women scheduled to undergo breast biopsy reported significant increases in anxiety (STAI) and mood disturbance (POMS). Gel shift mobility assays showed that mitogen stimulated peripheral blood lymphocytes of these women were less capable of the nuclear expression of AP-1 or NFkappaB at T1. Similar assessments, 7-10 days after the women learned of the results of their breast biopsy, showed these same women to have a marked reduction in anxiety and mood disturbance and an increased nuclear translocation of AP-1 and NFkappaB. These results show a significant decrease in nuclear AP-1 and NFkappaB expression during the period of emotional distress prior to biopsy with a return of nuclear transcription activity to normal levels when distress was relieved. Several studies have correlated increased psychological stress with decreased immune function. The results of this study suggest that psychological stress may mediate immunosuppression by altering the expression of the transcription factors, AP-1 and NFkappaB. PMID- 11259083 TI - Altered functional responses with preserved morphology of gonadotrophic cells in congenitally athymic mice. AB - Neonatal thymectomy or congenital absence of the thymus induces severe reproductive deficiencies in female mice, which are associated with reduced levels of circulating and pituitary gonadotropins. In contrast, the reproductive function is well preserved in nude males. It was therefore of interest to assess gonadotrophic cell morphology and function in congenitally athymic male mice. Circulating gonadotropins were measured under basal and stressful conditions, taking as a reference their haired counterparts. Adult normal (+/+), heterozygous nude (nu/+), and homozygous (nu/nu) CD-1 mice were subjected to 1-h immobilization stress. Serum levels of luteinizing hormone (LH) and follicle stimulating hormone (FSH) were assessed by RIA at 0, 30, and 60 min poststress. Athymic animals showed significantly lower basal levels of serum LH and FSH than their heterozygous littermates. Immunohistochemical assessment of LH and FSH cell populations revealed a normal morphology and cell number in the athymic animals compared to their normal littermates. Immobilization stress induced a significant reduction in gonadotrophin levels, particularly LH, in normal mice but had only a weak effect in athymic animals. It is concluded that congenital athymia in the adult male mouse is associated with decreased basal levels of serum LH and FSH, in the presence of a normal gonadotroph number and morphology. The anomalous responses of athymic mice to stress do not appear to be due to primary hypopituitarism but, rather, to an altered modulation of pituitary hormone secretion. . PMID- 11259084 TI - Peripheral blood natural killer cell cytotoxicity after damage to the limbic system in the rat. AB - The present work was aimed at examining the possible involvement of different parts of the septal area (dorsal, medial, lateral, and septohypothalamic nucleus), the basolateral amygdala, and the bed nucleus of the stria terminalis (BNST) in the regulation of the cytotoxic activity of NK cells (NKCC). The experimental approach included performing electrolytic (or sham) lesions in the tested brain areas and to measuring the peripheral blood NKCC (chromium-51 release assay), the number of leukocytes and lymphocytes, and the plasma corticosterone levels both before and at different time points after the lesion. Lesions were also induced in the three extralimbic structures: the paraventricular hypothalamic nucleus (PVN), the dorsal caudate-putamen, and the cerebellum. To test for a possible effect on NKCC of stress associated with blood collection, anesthesia, cranial surgery, and passing electric current through the brain the proper control experiments were also performed. Lesions of the medial septum and BNST caused gradual depression of NKCC, which peaked on the 10th day after the lesion, followed by a recovery to the baseline on days 21 (medial septum) and 42 (BNST) postinjury. In the respective sham-lesioned groups, mere insertion of electrodes into the medial septum and BNST evoked transient enhancement of NKCC (on the 3rd postlesion day), probably resulting from mechanical stimulation of the nervous tissue. Destruction of the other limbic and extralimbic structures appeared ineffective. After PVN lesions NKCC remained unchanged, despite an approximately 60% decrease in the basal corticosterone level. No adverse effects of the experimental and surgical procedures on NKCC, leukocyte and lymphocyte number, and corticosterone level were found, indicating that electrolytic lesions and other stereotaxic techniques can be safely used to study the brain-immune system interactions. The results obtained raise the question about the interrelationship between the medial septum and the hippocampal formation, BNST, the medial amygdala, and the hypothalamus (both medial and lateral) as a possible circuit involved in the regulation of cellular immune functions. PMID- 11259085 TI - Determination of p53 genotypes in oral cancer patients from India. AB - The p53 tumour suppressor gene is inactivated in various types of human cancers, and has been implicated as an early event in several cancers. A p53 Pro/Arg polymorphism at exon 4 codon 72, has been suggested to be involved in susceptibility to cancers as well. Hence, in the current study, we investigated p53 exon 4 codon 72 polymorphism using Proline or Arginine specific primers from the peripheral blood cells (PBC) representing constitutional DNA from 72 oral cancer patients. PBC from 153 normal healthy individuals were used to determine the frequency of the p53 genotypes, Pro/Pro, Arg/Arg and Pro/Arg, in the Indian population. The frequency of distribution of genotypes in the normal healthy individuals was, Pro/Pro - 0.20 (31/153), Arg/Arg -- 0.14 (22/153) and Pro/Arg -- 0.65 (100/153); and in the oral cancer patients was, Pro/Pro -- 0.19 (14/72), Arg/Arg -- 0.08 (6/72) and Pro/Arg -- 0.72 (52/72). Thus, we observed an equidistribution of the genotypes in normal control and oral cancer patients (chi(2)= 1.77, df = 2, 0.3 A > B in ICC; in the cases with a single HPV16 E6 sequence, coexisting ICC, CIN and normal epithelium in the same patient shared the E6 variant; and 4 cases of ICC had double/multiple E6 variants. The results did not show any importance of E6 variants for ICC progression in Russian women. The results also indicated that the original HPV16 variant persisted during ICC progression, and that at a low frequency, double infections and/or mutation of variants might occur. PMID- 11259094 TI - Addition of high-risk HPV testing improves the current guidelines on follow-up after treatment for cervical intraepithelial neoplasia. AB - We assessed a possible role for high-risk human papillomavirus (HPV) testing in the policy after treatment for cervical intraepithelial neoplasia (CIN) 2 or 3 (moderate to severe dysplasia). According to the Dutch guidelines follow-up after treatment consists of cervical cytology at 6, 12 and 24 months. Colposcopy is only performed in case of abnormal cervical cytology. In this observational study 184 women treated for CIN 2 or 3 were prospectively monitored by cervical cytology and high-risk HPV testing 3, 6, 9, 12 and 24 months after treatment. Post-treatment CIN 2/3 was present in 29 women (15.8%). A positive high-risk HPV test 6 months after treatment was more predictive for post-treatment CIN 2/3 than abnormal cervical cytology (sensitivity 90% and 62% respectively, with similar specificity). At 6 months the negative predictive value of a high-risk HPV negative, normal smear, was 99%. Largely overlapping, partly different groups of women with post-treatment CIN 2/3 were identified by HPV testing and cervical cytology. Based on these results we advocate to include high-risk HPV testing in monitoring women initially treated for CIN 2/3. In case of a high-risk HPV positive test or abnormal cervical cytology, colposcopy is indicated. All women should be tested at 6 and 24 months after treatment and only referred to the population-based cervical cancer screening programme when the tests are negative on both visits. PMID- 11259095 TI - Decrease of breast cancer cell invasiveness by sodium phenylacetate (NaPa) is associated with an increased expression of adhesive molecules. AB - Sodium phenylacetate (NaPa), a non-toxic phenylalanine metabolite, has been shown to induce in vivo and in vitro cytostatic and antiproliferative effects on various cell types. In this work, we analysed the effect of NaPa on the invasiveness of breast cancer cell (MDA-MB-231, MCF-7 and MCF-7 ras). Using the highly invasive breast cancer cell line MDA-MB-231, we demonstrated that an 18 hour incubation with NaPa strongly inhibits the cell invasiveness through Matrigel (86% inhibition at 20 mM of NaPa). As cell invasiveness is greatly influenced by the expression of urokinase (u-PA) and its cell surface receptor (u PAR) as well as the secretion of matrix metalloproteinases (MMP), we tested the effect of NaPa on these parameters. An 18-hour incubation with NaPa did not modify u-PA expression, either on MDA-MB-231 or on MCF-7 and MCF-7 ras cell lines, and induced a small u-PA decrease after 3 days of treatment of MDA-MB-321 with NaPa. In contrast, an 18 h incubation of MDA-MB-231 increased the expression of u-PAR and the secretion of MMP-9. As u-PAR is a ligand for vitronectin, a composant of the extracellular matrix, these data could explain the increased adhesion of MDA-MB-231 to vitronectin, while cell adhesivity of MCF-7 and MCF-7 ras was unmodified by NaPa treatment. NaPa induced also an increased expression of both Lymphocyte Function-Associated-1 (LFA-1) and Intercellular Adhesion Molecule-1 (ICAM-1), which was obvious from 18 hour incubation with NaPa for the MDA-MB-231 cells, but was delayed (3 days) for MCF-7 and MCF-7 ras. Only neutralizing antibodies against LFA-1 reversed the decreased invasiveness of NaPa treated cells. Therefore we can conclude that the strong inhibition of MDA-MB-231 invasiveness is not due to a decrease in proteases involved in cell migration (u PA and MMP) but could be related both to the modification of cell structure and an increased expression of adhesion molecules such as u-PAR and LFA-1. PMID- 11259096 TI - Chromogranin A and the alpha-subunit of glycoprotein hormones in medullary thyroid carcinoma and phaeochromocytoma. AB - Using specific immunoradiometric assays, we evaluated the clinical usefulness of chromogranin A and the alpha-subunit of glycoprotein hormones in neuroendocrine tumours of neuroectodermic origin. The serum alpha-subunit of glycoprotein hormones was only slightly increased in 2 out of 44 medullary thyroid carcinoma or phaeochromocytoma patients with increased calcitonin or 24-hour urinary metanephrine levels. Serum chromogranin A was increased in 12 of 45 (27%) medullary thyroid carcinoma patients with an elevated calcitonin level and in 4 of 16 medullary thyroid carcinoma patients (25%) with an undetectable calcitonin level, in 5 of 7 phaeochromocytoma patients with increased urinary catecholamine and metabolite excretion, and in 2 of 3 patients with a non-functioning phaeochromocytoma. During follow-up, the course of chromogranin A was found to parallel that of tumour burden and/or 24-hour urinary metanephrine in 5 phaeochromocytoma patients. We conclude that chromogranin A measurement is not recommended for the diagnosis of medullary thyroid carcinoma patients. It may be useful in patients with functioning and non-functioning phaeochromocytomas as a follow-up marker. In neuroendocrine tumour patients with elevated calcitonin secretion, the serum alpha-subunit of glycoprotein hormone measurement may help differentiate medullary thyroid carcinoma or phaeochromocytoma patients from other endodermal-derived neuroendocrine tumour patients in whom it is frequently elevated. PMID- 11259097 TI - Development of pulmonary bronchiolo-alveolar adenocarcinomas in transgenic mice overexpressing murine c-myc and epidermal growth factor in alveolar type II pneumocytes. AB - Transgenic mouse models were established to study tumorigenesis of bronchiolo alveolar adenocarcinomas derived from alveolar type II pneumocytes (AT-II cells). Transgenic lines expressing the murine oncogene c- myc under the control of the lung-specific surfactant protein C promoter developed multifocal bronchiolo alveolar hyperplasias, adenomas and carcinomas respectively, whereas transgenic lines expressing a secretable form of the epidermal growth factor (IgEGF), a structural and functional homologue of transforming growth factor alpha (TGF alpha), developed hyperplasias of the alveolar epithelium. Since the oncogenes c- myc and TGF alpha are frequently overexpressed in human lung bronchiolo-alveolar adenocarcinomas, these mouse lines are useful as models for human lung bronchiolo alveolar adenocarcinomas. The average life expectancies of hemizygous and homozygous c- myc transgenics were 14.25 months and 9.2 months, respectively, suggesting that a dosage effect of c- myc caused an accelerated bronchiolo alveolar adenocarcinoma formation. First analyses of double transgenics, hemizygous for both c- myc and IgEGF, show that these mice develop bronchiolo alveolar adenocarcinomas at the average age of 9 months, indicating that these oncogenes cooperate during the lung cancer formation. Our results demonstrate that c- myc and EGF are directly involved and cooperate with one another during formation of bronchiolo-alveolar adenocarcinomas in the lung. PMID- 11259098 TI - Lectin binding to cutaneous malignant melanoma: HPA is associated with metastasis formation. AB - Changes in protein glycosylation of tumour cells, as detected by lectin histochemistry, have been associated with metastasis formation in several human malignancies. This study analysed the association between lectin binding and metastasis in cutaneous malignant melanoma. In a 10-year retrospective study, sections of 100 primary cutaneous malignant melanomas were histochemically stained for the following 5 lectins: HPA, SNA-I, MAA, WGA and PHA-L, differing in their carbohydrate specificity. Since differences in the results of HPA binding depending on methodology have been reported, an indirect and a biotinylated method were employed for HPA. Kaplan-Meier analysis of time to first metastasis revealed a positive correlation between HPA binding and metastasis for both methods, with the biotinylated HPA method (P< 0.0001) being superior to the 'indirect' method (P = 0.0006). Cox regression analysis demonstrated that even after adjustment for stage, HPA positivity is an independent predictor for metastasis. The results of the present study indicate that N -acetyl galactosamine/-glucosamine residues, recognized by HPA, are linked to metastasis in malignant melanoma. In contrast, beta1-6 branched oligosaccharides or sialic acid residues, both of which were correlated with metastasis in other malignancies, are of no functional importance for metastasis formation in malignant melanoma. Thus, HPA proved to be a useful and independent prognostic marker for the metastatic phenotype of melanoma. PMID- 11259099 TI - Tumour-amplified kinase BTAK is amplified and overexpressed in gastric cancers with possible involvement in aneuploid formation. AB - Our recent analysis of gastric cancers using comparative genomic hybridization (CGH) revealed a novel high frequent copy number increase in the long arm of chromosome 20. Tumour-amplified kinase BTAK was recently cloned from breast cancers and mapped on 20q13 as a target gene for this amplification in human breast cancers. In the study presented here, we analysed BTAK copy-number and expression, and their relation to the ploidy pattern in 72 primary gastric cancers. Furthermore, wild-type BTAK and its deletion mutants were transfected to gastric cancers to examine changes in cell proliferation and DNA ploidy pattern. Evaluation of 72 unselected primary gastric cancers found BTAK amplification in 5% and overexpression in more than 50%. All four clinical samples with BTAK amplification showed aneuploidy and poor prognosis. Transfection of BTAK in near diploid gastric cancers induced another aneuploid cell population. In contrast, the c-terminal-deleted mutant of BTAK induced no effect in DNA ploidy pattern and inhibited gastric cancer cell proliferation. These results suggest that BTAK may be involved in gastric cancer cell aneuploid formation, and is a candidate gene for the increase in the number of copies of the 20q, and thus may contribute to an increase in the malignant phenotype of gastric cancer. PMID- 11259100 TI - Combretastatin-A4 disrupts neovascular development in non-neoplastic tissue. AB - Combretastatin-A4 phosphate (cis-CA-4) is a tubulin-binding agent currently undergoing clinical trials as an anti-tumour drug. We have investigated whether CA-4 functions as a tumour-specific anti-vascular agent using the hyperplastic thyroid as a novel in vivo model of neovascularization. CA-4 elicited pathological changes in normal tissue, manifested as the induction of multiple, discrete intravascular thrombi. These vascular-damaging effects indicate that CA 4P does not function as a tumour-specific agent but targets neovasculature irrespective of the primary angiogenic stimulus. PMID- 11259101 TI - Inhibition of erythropoietin signalling destroys xenografts of ovarian and uterine cancers in nude mice. AB - We have recently shown that malignant tumours from the ovary and uterus expressed erythropoietin (Epo) and its receptor (EpoR), and that deprivation of Epo signal in tumour blocks induced death of malignant cells and capillary endothelial cells in vitro (Yasuda et al, submitted). These in vitro results prompted us to examine the effect of Epo-signal withdrawal on tumours in vivo. RT-PCR analysis demonstrated the expression of mRNAs for Epo and EpoR in the transplants of uterine and ovarian tumours in nude mice. Then we injected locally anti-Epo antibody or soluble form of EpoR into the transplants. At 12 h, 1, 7 or 14 days after the injection, all transplants were resected and examined macro- and microscopically. Tumour size was reduced in Epo signal-deprived transplants. Immunohistochemical examinations revealed destruction of Epo-responding malignant and capillary endothelial cells through apoptotic death. The degree of tumour regression correlated well with the dose and frequency of the injections. Control xenografts with saline injection or needle insertion showed well-developed tumour masses. This Epo response pathway will have profound implications for our understanding of the development and progression of malignant tumours and for the use of Epo-signal deprivation as an effective therapy. PMID- 11259102 TI - EGCG, a major component of green tea, inhibits tumour growth by inhibiting VEGF induction in human colon carcinoma cells. AB - Catechins are key components of teas that have antiproliferative properties. We investigated the effects of green tea catechins on intracellular signalling and VEGF induction in vitro in serum-deprived HT29 human colon cancer cells and in vivo on the growth of HT29 cells in nude mice. In the in vitro studies, (-) epigallocatechin gallate (EGCG), the most abundant catechin in green tea extract, inhibited Erk-1 and Erk-2 activation in a dose-dependent manner. However, other tea catechins such as (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECG), and (-)-epicatechin (EC) did not affect Erk-1 or 2 activation at a concentration of 30 microM. EGCG also inhibited the increase of VEGF expression and promoter activity induced by serum starvation. In the in vivo studies, athymic BALB/c nude mice were inoculated subcutaneously with HT29 cells and treated with daily intraperitoneal injections of EC (negative control) or EGCG at 1.5 mg day( 1)mouse(-1)starting 2 days after tumour cell inoculation. Treatment with EGCG inhibited tumour growth (58%), microvessel density (30%), and tumour cell proliferation (27%) and increased tumour cell apoptosis (1.9-fold) and endothelial cell apoptosis (3-fold) relative to the control condition (P< 0.05 for all comparisons). EGCG may exert at least part of its anticancer effect by inhibiting angiogenesis through blocking the induction of VEGF. PMID- 11259103 TI - Squamous cell carcinoma antigen suppresses radiation-induced cell death. AB - Previous study has demonstrated that squamous cell carcinoma antigen (SCCA) 1 attenuates apoptosis induced by TNF alpha, NK cell or anticancer drug. In this study, we have examined the effect of SCCA2, which is highly homologous to SCCA1, but has different target specificity, against radiation-induced apoptosis, together with that of SCCA1. We demonstrated that cell death induced by radiation treatment was remarkably suppressed not only in SCCA1 cDNA-transfected cells, but also in SCCA2 cDNA-transfected cells. In these transfectants, caspase 3 activity and the expression of activated caspase 9 after radiation treatment were suppressed. Furthermore, the expression level of phosphorylated p38 mitogen activated protein kinase (p38 MAPK) was suppressed compared to that of the control cells. The expression level of upstream stimulator of p38 MAPK, phosphorylated MKK3/MKK6, was also suppressed in the radiation-treated cells. Thus, both SCCA1 and SCCA2 may contribute to survival of the squamous cells from radiation-induced apoptosis by regulating p38 MAPK pathway. PMID- 11259104 TI - Optimal timing and dosage of chemotherapy as a combined treatment with androgen withdrawal in the human prostate LNCaP tumour model. AB - Although numerous chemotherapeutic agents have been evaluated for patients with advanced prostate cancer, none have demonstrated improved survival benefits. Here, in order to determine whether the efficacy of chemotherapy can be enhanced by changing the regimen, we evaluated the effect of the varied timing and dosage of chemotherapy in combination with androgen withdrawal on time to androgen independent (AI) progression in the human androgen-dependent LNCaP tumour model. We first demonstrated the synergistic effect of mitoxantrone on induction of apoptosis in LNCaP cells maintained in the steroid hormone-depleted charcoal stripped media (CSM) compared to those in the standard media. In addition, this synergy was most remarkable during the simultaneous treatment of LNCaP cells with mitoxantrone and CSM compared to the pre- or post-use of mitoxantrone with CSM. LNCaP tumour growth in athymic nude mice and their increase in serum PSA levels were significantly inhibited by the simultaneous injection of mitoxantrone with castration, compared to the administration of mitoxantrone 2 weeks before or after castration. The TUNEL staining revealed that apoptotic cell death was extensively induced in LNCaP tumours treated with simultaneous castration and mitoxantrone compared to tumours treated with the other schedules. Furthermore, nude mice bearing LNCaP tumours were injected with a total of 0.5 mg mitoxantrone divided into 2, 5 and 10 days, with the most significant therapeutic effect and delayed AI progression observed in mice given injection of mitoxantrone for 2 days. These findings suggest that this might be the optimal way to perform cytotoxic chemotherapy and androgen withdrawal simultaneously in patients with advanced prostate cancer and to administer chemotherapeutic agents at high dosage within short intervals. PMID- 11259105 TI - NK4, a four-kringle antagonist of HGF, inhibits spreading and invasion of human pancreatic cancer cells. AB - Because of the highly aggressive behaviour, i.e. invasive, disseminative and metastatic properties, the outcome for patients with pancreatic cancer is morbid. A better understanding and interference with the malignant behaviour of pancreatic cancer may provide new directions for treatment. We report here the induction of highly motile and invasive properties in human pancreatic cancer cells by hepatocyte growth factor (HGF) and blockage of these properties by NK4, a newly identified antagonist for HGF. In all of eight human pancreatic cancer cell lines we used (AsPC-1, BxPC-3, H-48N, KP-1N, KP-2, KP-3, MIA PaCa-2 and SUIT 2 cells), the c-Met/HGF receptor was expressed at varying levels. Although weak mitogenic activity of HGF was seen only in SUIT-2 and KP-3 cells, HGF strongly stimulated migration and invasion of these pancreatic cancer cells, except for BxPC-3 and MIA PaCa-2 cells. In contrast, migration and invasion potently induced by HGF in KP-1N, KP-3 and SUIT-2 cells were inhibited by NK4. The invasion of SUIT-2 cells was also potently stimulated with the influence of cocultured pancreatic fibroblasts and by ascitic fluid obtained after pancreatic cancer resection, however, invasiveness of the cancer cells in such conditions was practically abolished by NK4. Consistently, the ascitic fluid in patients who had undergone pancreatic cancer surgery contained high levels of HGF. These findings mean that HGF is probably involved in invasion, dissemination, and metastasis of pancreatic cancer, particularly through tumour-stromal interaction and after resection of the pancreatic cancer. NK4, an effective antagonist of HGF, may prove to have the potential for anti-invasion/metastasis. PMID- 11259106 TI - Error monitoring in speech production: a computational test of the perceptual loop theory. AB - A theory of speech monitoring, proposed by Levelt (1983), assumes that the quality of one's speech is checked by the speech comprehension system. This system inspects one's own overt speech but would also inspect an inner speech plan ("the inner loop"). We have elaborated and tested this theory by way of formalizing it as a computational model. This model includes a new proposal concerning the timing relation between planning the interruption and the repair: the proposal that these two processes are performed in parallel. We attempted to simulate empirical data about the distribution of error-to-cutoff and cutoff-to repair intervals and the effect of speech rate on these intervals (these intervals are shorter with faster speech). The main questions were (1) Is an inner monitor that utilizes the speech perception system fast enough to simulate the timing data? (2) Can the model account for the effects of speech rate on these intervals? We conclude that including an inner loop through the speech comprehension system generates predictions that fit the empirical data. The effects of speed can be accounted for, given our proposal about the time course of planning interruption and repair. A novel prediction is that the error-to cutoff interval decreases with increasing position in the phrase. PMID- 11259107 TI - To Matrix, Network, or Hierarchy: That Is the Question. AB - The authors present a structural analysis of three spatial diagrams-matrices, networks, and hierarchies-that specifies 10 properties on which these diagrammatic representations are hypothesized to differ: global structure, building block, number of sets, item/link constraints, item distinguishability, link type, absence of a relation, linking relations, path, and traversal. Each property has a "value" for each diagram, and these property values constitute the applicability conditions for the representations. Twenty-three college students (computer science majors and math educators) selected the type of diagram they thought would be most efficient for organizing the information in each of 18 short scenarios and verbally justified the reasons for their selections. The verbal protocols were coded with respect to the structural analysis. Both the representation selection and verbal justification data provided strong support for the structural analysis. Additionally, a factor analysis of students' justifications indicated that the organization of their knowledge is consistent with the structural analysis. Students' use of the structural properties to select appropriate representations and to justify those selections indicates that the structural analysis has psychological force. PMID- 11259108 TI - Beta-amyloid plaques induce neuritic dystrophy of nitric oxide-producing neurons in a transgenic mouse model of Alzheimer's disease. AB - A causative role for nitric oxide has been postulated in a number of neurodegenerative diseases. Using histochemical and immunohistochemical methods, we examined the effect of beta-amyloid plaques on nitric oxide-producing cells in transgenic mice which overexpress a mutant human amyloid precursor protein (APP). In 14-month-old animals, nitric oxide synthase (NOS)-positive dystrophic neurites were observed frequently in the cerebral cortex and hippocampus of all of 16 plaque-bearing transgenic animals and in none of 16 wild-type animals. Double labeling of NOS and beta-amyloid revealed that 90% of beta-amyloid plaques were associated with NOS-containing dystrophic neurites. In 7-month-old animals, beta amyloid plaques were very rare, but those present were frequently associated with NOS-positive neuritic dystrophy. We conclude that beta-amyloid plaques induce neuritic dystrophy in cortical neurons containing NOS in this model of AD, and hypothesize that this finding may be relevant to the mechanism of beta-amyloid neurotoxicity in human AD. PMID- 11259109 TI - Delayed glial cell death following wallerian degeneration in white matter tracts after spinal cord dorsal column cordotomy in adult rats. AB - The devastating consequences of spinal cord injury (SCI) result primarily from damage to long tracts in the spinal white matter. To elucidate the secondary injury processes occurring after SCI, we investigated the relationship between apoptosis and Wallerian degeneration in spinal white matter tracts. In the rat spinal cord, the corticospinal tract (CST) and the dorsal ascending tract (DAT) are separated from each other in the dorsal column and relay information in opposite directions. A dorsal column cordotomy at the eighth thoracic (T8) level simultaneously induces Wallerian degeneration in the CST caudal to and in the DAT rostral to the injury. Using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) method, we demonstrate that apoptosis occurred in areas of Wallerian degeneration in both tracts throughout the length of the cord segments studied (from T3 to T12). This delayed cell death, more apparent in the DAT, began at 7 days after injury and peaked at 14 days for the DAT and 28 days for the CST. Although a few TUNEL+ cells, slightly above the noninjury control level, were found in intact areas of both tracts, statistically significant differences in the number of TUNEL+ cells were found between the intact and the lesioned tract segments (CST, F < 0.01; DAT, F < 0.001). Within a particular spinal segment, a mean number of 64 and 939 TUNEL+ cells in the degenerating CST and DAT, respectively, were estimated stereologically at 14 days postinjury. TUNEL+ cells in degenerating tracts outnumber their intact counterparts by 3.8:1 in the CST and 4.1:1 in the DAT, although a statistically significant difference between the two was only found in the DAT at this time point (P < 0.05). Finally, we demonstrated that oligodendrocytes, the myelin-forming cells in the central nervous system, constitute at least a portion of the cells undergoing apoptosis within areas of Wallerian degeneration. PMID- 11259110 TI - Herpes simplex virus vector-mediated expression of Bcl-2 protects spinal motor neurons from degeneration following root avulsion. AB - Proximal axotomy in adult animals results in delayed death of motor neurons. Features characteristic of both necrosis and apoptosis have been described in motor neurons of the spinal cord following proximal avulsion of the ventral roots. We have previously demonstrated that a genomic herpes simplex virus (HSV) based vector expressing the anti-apoptotic peptide Bcl-2 protects dopaminergic neurons of the substantia nigra from neurotoxin-induced apoptotic cell death and preserves the neurotransmitter phenotype of those cells. In this study we examined whether the same vector could protect adult rat lumbar motor neurons from cell death following proximal ventral root avulsion. Injection of the Bcl-2 expressing vector 1 week prior to root avulsion increased the survival of lesioned motor neurons, determined by retrograde Fluorogold labeling, by 50%. The Bcl-2-expressing vector did not preserve choline acetyltransferase neurotransmitter phenotype of the lesioned cells. These results shed light on the mechanism of cell death following axonal injury, and have implications for developing an effective treatment for the clinical problem of proximal root avulsion. PMID- 11259111 TI - Changes in the gene expression of GABA(A) receptor alpha1 and alpha2 subunits and metabotropic glutamate receptor 5 in the basal ganglia of the rats with unilateral 6-hydroxydopamine lesion and embryonic mesencephalic grafts. AB - By using an animal model of parkinsonism, we examined the expression of GABA(A) receptor (R) and metabotropic glutamate receptor (mGluR) 5 in the basal ganglia after transplantation with dopamine-rich tissue. The adult rats were unilaterally lesioned by the injection of 6-hydroxydopamine to their left medial forebrain bundles. At 5-10 weeks following the dopaminergic denervation, the levels of GABA(A)R in the left caudate-putamen and globus pallidus were about 20 and 16% lower than that of the right intact (control) sides, as shown by [3H]flunitrazepam binding autoradiography on the brain sections. However, the receptor density increased to around 132 and 130% of control levels in the entopeduncular nucleus and substantia nigra pars reticulata of the lesioned sides. Furthermore, in situ hybridization analysis exhibited parallel trends of changes in the levels of the GABA(A)R alpha1 and alpha2 subunit and mGluR5 mRNAs in the neurons of the brain regions with that of the proteins detected by the binding assay. A number of the rats 5 weeks postlesion were transplanted with the ventral mesencephalon of the embryonic rat into their left striata. Five weeks later, the changes in the [3H]flunitrazepam binding seemed to be recovered by approximately 50-63% on the grafted sides of the areas. Moreover, the transplantation appeared to produce a nearly complete reversal of the lesion induced alterations in the levels of the mRNAs. Thus, the data indicate the mechanism of gene regulation for the modified expression of the receptors and could implicate the participation of the receptors in the pathogenesis of Parkinson's disease. PMID- 11259112 TI - Turnover of rat brain perivascular cells. AB - Brain perivascular spaces harbor a population of cells which exhibit high phagocytic capacity. Therefore, these cells can be labeled by intraventricular injection of tracers. Such perivascular cells at the interface between blood and brain are believed to belong to the monocyte/macrophage lineage and to be involved in antigen presentation. Currently, it is unclear whether these cells undergo a continuous turnover by entering and leaving the bloodstream. Using bone marrow-chimeric animals, migration of donor macrophages into brain perivascular spaces has been reported. On the other hand, following intracerebral injection of india ink into nontransplanted animals, ink-labeled perivascular cells were still found 2 years after injection, suggesting a high stability of this cell pool. Thus, the turnover of perivascular cells observed in chimeras might be a result of bone marrow transplantation rather than a physiological occurrence. To address this issue, we monitored de novo invasion of macrophages into perivascular spaces of apparently healthy adult rats by applying techniques other than bone marrow transplantation, (i) consecutive injections of different tracers and (ii) ex vivo isolation of macrophages from the blood, cell labeling, and reinjection into the same animal to avoid MHC mismatch. Both approaches revealed vivid de novo invasion of macrophages into perivascular spaces, but not into brain parenchyma, rendering untenable the concept of perivascular cells forming a stable population of macrophages in the brain. Thus, brain perivascular spaces are under permanent immune surveillance of blood borne macrophages in normal adult rats. PMID- 11259113 TI - Overexpression of HuD accelerates neurite outgrowth and increases GAP-43 mRNA expression in cortical neurons and retinoic acid-induced embryonic stem cells in vitro. AB - The neuron-specific RNA-binding protein HuD binds to a U-rich regulatory element of the 3' untranslated region (3' UTR) of the GAP-43 mRNA and stabilizes the mRNA. We have previously shown that overexpression of HuD in PC12 cells increases GAP-43 protein expression and induces the spontaneous formation of multiple neurites (K. D. Anderson et al. 2000. J. Neurochem. 75: 1103-1114). In this study, we examined the effects of HuD overexpression on the initial stages of neurite outgrowth and on GAP-43 gene expression using two in vitro systems: E19 rat cortical neurons and retinoic acid (RA)-induced embryonic stem (ES) cells. Normal neurite outgrowth of cortical neurons in vitro occurs over a 3-day period with a concomitant increase in GAP-43 and HuD expression. Cortical cells were infected with a replication-deficient HSV-1 vector containing the HuD cDNA in the sense orientation (HSV-HuD). Overexpression of HuD accelerated the formation of neurites. Immunocytochemical analysis showed that excess HuD resulted in a threefold increase in the number of GAP-43-positive cells undergoing morphological differentiation after 24 h of treatment. Using in situ hybridization, we found that the increased HuD expression resulted in a twofold increase in the levels of GAP-43 mRNA. Similarly, overexpression of HuD in RA induced embryonic stem cells was found to increase the number of GAP-43-positive cells undergoing process outgrowth. In conclusion, our results demonstrate that HuD functions in the initiation of neurite outgrowth in a manner due, at least in part, to its regulation of GAP-43 expression. PMID- 11259114 TI - Depletion of taurine in experimental diabetic neuropathy: implications for nerve metabolic, vascular, and functional deficits. AB - In diabetes, increased oxidative stress, disruption of signal transduction pathways, and endothelial dysfunction have been critically implicated in the pathogenesis of experimental diabetic neuropathy (EDN). The development of nerve conduction slowing in diabetes is accompanied by depletion of the beta-amino acid taurine. Since taurine functions as an antioxidant, calcium modulator, and vasodilator, taurine depletion may provide a pathogenetic link between nerve metabolic, vascular, and functional deficits complicating diabetes. The mechanism(s) of nerve taurine depletion, the localization of critical taurine deficits, and its pathophysiological significance in EDN are however unknown. This study explored the pathophysiological effects of selective nerve taurine replacement in streptozotocin-diabetic (STZ-D) rats. A polyclonal human taurine transporter (TT) antibody was also generated in order to determine potential loci of critical taurine depletion. Two weeks of STZ-D reduced sciatic motor nerve conduction velocity (NCV) by 23% (P < 0.01), decreased composite nerve blood flow by 38% (P < 0.01), and reduced nerve taurine content by 29% (P < 0.05). In STZ-D rats, a 1% taurine diet corrected nerve taurine depletion, prevented motor NCV slowing, and partially attenuated composite nerve blood flow deficits. After 6 weeks of STZ-D, a 1% taurine diet ameliorated motor NCV slowing and endoneurial nutritive blood flow deficits, prevented digital sensory NCV slowing, and reduced ouabain-sensitive nerve (Na,K)-ATPase activity. Immunohistochemical studies localized taurine and the TT to the vascular endothelium and Schwann cells of the sciatic nerve. In conclusion, taurine depletion in the vascular endothelium and Schwann cells of the sciatic nerve may contribute to the neurovascular and metabolic deficits in EDN. PMID- 11259115 TI - Temporal-spatial pattern of acute neuronal and glial loss after spinal cord contusion. AB - The secondary loss of neurons and glia over the first 24 h after spinal cord injury (SCI) contributes to the permanent functional deficits that are the unfortunate consequence of SCI. The progression of this acute secondary cell death in specific neuronal and glial populations has not previously been investigated in a quantitative manner. We used a well-characterized model of SCI to analyze the loss of ventral motoneurons (VMN) and ventral funicular astrocytes and oligodendrocytes at 15 min and 4, 8, and 24 h after an incomplete midthoracic contusion injury in the rat. We found that both the length of lesion and the length of spinal cord devoid of VMN increased in a time-dependent manner. The extent of VMN loss at specified distances rostral and caudal to the injury epicenter progressed symmetrically with time. Neuronal loss was accompanied by a loss of glial cells in ventral white matter that was significant at the epicenter by 4 h after injury. Oligodendrocyte loss followed the same temporal pattern as that of VMN while astrocyte loss was delayed. This information on the temporal spatial pattern of cell loss can be used to investigate mechanisms involved in secondary injury of neurons and glia after SCI. PMID- 11259116 TI - Relationship of altered glutamate receptor subunit mRNA expression to acute cell loss after spinal cord contusion. AB - Alterations in the expression of ionotropic glutamate receptors (GluR) contribute to neuronal loss after brain ischemia and epilepsy. In order to determine whether altered expression of GluR subunits might contribute to cell loss after spinal cord injury (SCI), we performed a time course study of subunit mRNA expression using quantitative in situ hybridization. Expression was studied in ventral horn motor neurons (VMN) and glia in adjacent ventral white matter at 15 min and 4, 8, and 24 h after SCI in tissue sections 4 mm rostral and caudal to the injury epicenter. We found that the AMPA subunit GluR2 was significantly down-regulated in VMN at 24 h, but not at the earlier times examined, although half the loss of VMN in these locations occurs by 8 h after injury. No changes in the normal expression of GluR2 or GluR4 were found in white matter where glial loss occurs after SCI. NMDA subunits NR1 and NR2A were significantly and rapidly up-regulated in VMN after SCI, but only caudal to the lesion site, while VMN loss is similar rostral and caudal to the epicenter. Thus, the temporal pattern of AMPA and the spatial pattern of NMDA subunit expression changes were distinct from the pattern of VMN loss after SCI. We conclude that altered GluR subunit expression after SCI is unlikely to be involved in secondary cell loss and instead may be involved with plasticity and reorganization of the injured spinal cord. PMID- 11259117 TI - Selective cell-cycle arrest and induction of apoptosis in proliferating neural cells by ganglioside GM3. AB - Control of cell proliferation and cell survival is critical during development of the vertebrate central nervous system (CNS). Much of the cell death seen during early stages of CNS development occurs through apoptosis; however, the factors that induce this early apoptosis are not clearly understood. Gangliosides, sialylated glycosphingolipids, are expressed in the CNS and have been proposed to regulate cell growth and differentiation. Here we show that the simple ganglioside GM3 selectively inhibits the proliferation of and induces apoptosis of actively dividing astrocyte precursors and other neural progenitors. The inhibition of astrocyte precursor proliferation by GM3 appears to be mediated in part by the cyclin-dependent kinase (Cdk) inhibitor p27(Kip1). During neonatal development there is extensive cell proliferation and little apoptosis in the ventricular and subventricular zones of the CNS. This proliferation was dramatically inhibited and the degree of apoptosis dramatically increased following intraventricular administration of GM3. These data suggest that GM3, a simple ganglioside, may regulate cell proliferation and death in the CNS and as such may have potential for brain tumor therapy. PMID- 11259118 TI - GM3 as a novel growth regulator for human gliomas. AB - The simple ganglioside GM3 inhibits proliferation and induces apoptosis in proliferating immature rodent CNS cells. To determine whether GM3 influenced the expansion of human neural tumors the effects of GM3 treatment on primary human brain tumors were assayed. Here we demonstrate that GM3 treatment dramatically reduces cell numbers in primary cultures of high-grade human glioblastoma multiforme (GBM) tumors and the rat 9L cell gliosarcoma cell line. By contrast, GM3 treatment had little effect on cell number in cultures of normal human brain. A single injection of GM3 3 days after intracranial implantation of 9L tumor cells in a murine xenograft model system resulted in a significant increase in the symptom-free survival period of host animals. The effects of GM3 were not restricted to GBMs and 9L cells. Cultures of high-grade ependymomas, mixed gliomas, astrocytomas, oligodendrogliomas, and gangliogliomas were all susceptible to GM3 treatment. These results suggest that GM3 may have considerable value as a selectively toxic chemotherapeutic agent for human high grade gliomas. PMID- 11259119 TI - Radial glial cell line C6-R integrates preferentially in adult white matter and facilitates migration of coimplanted neurons in vivo. AB - C6-R is a cell line derived from C6 glioma cells that exhibits key properties of radial glia including the ability to support neuronal migration in culture. To explore its potential use in promoting neuronal migration in vivo, we analyzed the behavior of C6-R cells in the intact and injured adult rat CNS. At 6-11 days postimplantation at the splenium of the corpus callosum, green fluorescent protein-labeled C6-R cells were observed primarily in either the corpus callosum or the hippocampus in the brain, and in the spinal cord they migrated more extensively in the white matter than in the grey matter. To determine whether C6 R cells retain their ability to promote neuronal migration in vivo, they were coinjected with labeled neurons into adult brain. When rat embryonic neurons were coimplanted with C6-R cells, the neurons and C6-R cells comigrated through a much larger volume than neurons alone or neurons coimplanted with fibroblasts. In brains preinjured with ibotenic acid, C6-R cells as well as coimplanted neurons distributed widely within the lesion site and migrated into adjacent brain tissue, while transplants with neurons alone were restricted primarily to the lesion site. The results suggest that radial glial cell lines can serve as a scaffold for neuronal migration that may facilitate development of experimental models for neural transplantation and regeneration. PMID- 11259120 TI - Delayed increase in neuronal nitric oxide synthase immunoreactivity in thalamus and other brain regions after hypoxic-ischemic injury in neonatal rats. AB - We examined the response of neuronal nitric oxide synthase (nNOS)-containing CNS neurons in rats exposed to a unilateral hypoxic-ischemic insult at 7 days of age. Animals were sacrificed at several time points after the injury, up to and including 7 days (Postnatal Day 14). Brain regions ipsilateral to the injury (including cerebral cortex, caudate-putamen, and thalamus) exhibited delayed, focal increases in nNOS immunoreactivity. The increase in nNOS immunoreactive fiber staining was prominent in areas adjacent to severe neuronal damage, especially in the cortex and the thalamus, regions that are also heavily and focally injured in term human neonates with hypoxic-ischemic encephalopathy. In cerebral cortex, these increases occurred despite modest declines in nNOS catalytic activity and protein levels. Proliferation of surviving nNOS immunoreactive fibers highlights regions of selective vulnerability to hypoxic ischemic insult in the neonatal brain and may also contribute to plasticity of neuronal circuitry during recovery. PMID- 11259121 TI - Astrocyte-targeted expression of interleukin-3 and interferon-alpha causes region specific changes in metallothionein expression in the brain. AB - Transgenic mice expressing IL-3 and IFN-alpha under the regulatory control of the GFAP gene promoter (GFAP-IL3 and GFAP-IFNalpha mice) exhibit a cytokine-specific, late-onset chronic-progressive neurological disorder which resemble many of the features of human diseases such as multiple sclerosis, Aicardi-Goutieres syndrome, and some viral encephalopathies including HIV leukoencephalopathy. In this report we show that the metallothionein-I+II (MT-I+II) isoforms were upregulated in the brain of both GFAP-IL3 and GFAP-IFNalpha mice in accordance with the site and amount of expression of the cytokines. In the GFAP-IL3 mice, in situ hybridization analysis for MT-I RNA and radioimmunoassay results for MT-I+II protein revealed that a significant upregulation was observed in the cerebellum and medulla plus pons at the two ages studied, 1-3 and 6-10 months. Increased MT I RNA levels occurred in the Purkinje and granular layers of the cerebellum, as well as in its white matter tracts. In contrast to the cerebellum and brain stem, MT-I+II were downregulated by IL-3 in the hippocampus and the remaining brain in the older mice. In situ hybridization for MT-III RNA revealed a modest increase in the cerebellum, which was confirmed by immunohistochemistry. MT-III immunoreactivity was present in cells that were mainly round or amoeboid monocytes/macrophages and in astrocytes. MT-I+II induction was more generalized in the GFAP-IFNalpha (GIFN12 and GIFN39 lines) mice, with significant increases in the cerebellum, thalamus, hippocampus, and cortex. In the high expressor line GIFN39, MT-III RNA levels were significantly increased in the cerebellum (Purkinje, granular, and molecular layers), thalamus, and hippocampus (CA2/CA3 and especially lacunosum molecular layers). Reactive astrocytes, activated rod like microglia, and macrophages, but not the perivenular infiltrating cells, were identified as the cellular sources of the MT-I+II and MT-III proteins. The pattern of expression of the different MT isoforms in these transgenic mice differed substantially, demonstrating unique effects associated with the expression of each cytokine. The results indicate that the MT expression in the CNS is significantly affected by the cytokine-induced inflammatory response and support a major role of these proteins during CNS injury. PMID- 11259122 TI - Differential expression and distribution of alpha-, beta-, and gamma-synuclein in the developing human substantia nigra. AB - Although the functions of alpha-, beta-, and gamma-synuclein (alphaS, betaS, gammaS, respectively) are unknown, these synaptic proteins are implicated in the pathogenesis of Parkinson's disease (PD) and related disorders. For example, alphaS forms Lewy bodies (LBs) in substantia nigra (SN) neurons of PD. However, since it is not known how these hallmark PD lesions contribute to the degeneration of SN neurons or what the normal function of alphaS is in SN neurons, we studied the developing human SN from 11 weeks gestational age (GA) to 16 years of age using immunohistochemistry and antibodies to alphaS, betaS, gammaS, other synaptic proteins, and tyrosine hydoxylase (TH). SN neurons expressed TH at 11 weeks GA and alphaS, betaS, and gammaS appeared initially at 15, 17, and 18 weeks GA, respectively. These synucleins first appeared in perikarya of SN neurons after synaptophysin, but about the same time as synaptotagmin and synaptobrevin. Redistribution of alphaS from perikarya to processes of SN neurons occurred by 18 weeks GA in parallel with synaptophysin, while betaS and synaptotagmin were redistributed similarly between 20 and 28 weeks GA and this also occurred with gammaS and synaptobrevin between 33 weeks GA and 9 months postnatal. These data suggest that alphaS, betaS, and gammaS may play a functional role in the development and maturation of SN neurons, but it remains to be determined how sequestration of alphaS as LBs in PD contributes to the degeneration of SN neurons. PMID- 11259123 TI - Transgenic ALS mice show increased vulnerability to the mitochondrial toxins MPTP and 3-nitropropionic acid. AB - The pathogenesis of neurodegenerative diseases may involve a genetic predisposition acting in concert with environmental toxins. To test this hypothesis we examined whether transgenic mice with the G93A mutation in Cu,Zn superoxide dismutase show increased vulnerability to either 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine (MPTP) or 3-nitropropionic acid (3-NP). Compared to littermate controls G93A transgenic mice showed a greater loss of striatal dopamine, DOPAC, and HVA at 50, 70, and 120 days of age following administration of MPTP; however, cell loss in the substantia nigra was not greater. The G93A transgenic mice showed significantly increased vulnerability to striatal lesions produced by 3-NP compared with littermate controls at 120 days of age. The finding that G93A mice show increased vulnerability to mitochondrial toxins further implicates mitochondrial dysfunction in the pathogenesis of neuronal death in these mice. The findings support the hypothesis that a genetic defect can increase susceptibility to environmental toxins and that this may play a role in the pathogenesis of neurodegenerative diseases. PMID- 11259124 TI - Ischemia-induced degeneration of CA1 pyramidal cells decreases seizure severity in a subgroup of epileptic gerbils and affects parvalbumin immunoreactivity of CA1 interneurons. AB - Mongolian gerbils are epilepsy-prone animals. In adult gerbils two major groups can be differentiated according to their seizure behavior: Highly seizure sensitive gerbils exhibit facial and forelimb clonus or generalized tonic-clonic seizures from the first test on, while kindled-like gerbils are seizure free for the first three to six consecutive tests, later develop forelimb myoclonus, and eventually progress to generalized tonic-clonic seizures. In the hippocampus, seizure history of the individual animal is mirrored in the intensity in which GABAergic neurons are immunostained for the calcium-binding protein parvalbumin: they lose parvalbumin with increasing seizure incidence. In a first step to clarify the influence of hippocampal projection neurons on spontaneous seizure behavior and related parvalbumin expression, we induced degeneration of the CA1 pyramidal cells by transient forebrain ischemia. This results in a decreased seizure sensitivity in highly seizure-sensitive gerbils. The kindling-like process, however, is not permanently blocked by the ischemic nerve cell loss, suggesting that an intact CA1 field is not a prerequisite for the development of seizure behavior. The seizure-induced loss of parvalbumin from the ischemia resistant interneurons recovers after ischemia. Thus, changes in parvalbumin content brought about by repeated seizures are not permanent but can rather be modulated by novel stimuli. PMID- 11259125 TI - Therapeutic effects of environmental enrichment on cognitive function and tissue integrity following severe traumatic brain injury in rats. AB - Postinjury environmental enrichment (EE) has been shown to alter functional and anatomical outcomes in a number of injury paradigms, including traumatic brain injury (TBI). The question of whether EE alters functional outcome following TBI in a model which produces overt histopathological consequences has not been addressed. We investigated this question using the severe, parasagittal fluid percussion injury (FPI) model. Rats (n = 7 per group, enriched and standard for behavior; n = 15 per group for histology) underwent severe (2.2-2.6 atm) FPI, with sham-operated rats (n = 7 per group, enriched and standard for behavior; n = 6 enriched, n = 3 standard for histology) serving as controls. Animals were allowed to recover for 11 days either in standard single housing or together (injured and sham) in an enriched environment consisting of a 92 x 61 x 77-cm ferret cage filled with various stimulatory objects. Consistent with earlier reports, injured animals recovering in the enriched environment showed significantly (P < 0.05) shorter latencies to find the platform in a Morris Water Maze task versus injured/standard animals on day 12 post-TBI. However, both injured groups showed significant deficits versus sham groups (P < 0.05). There were no differences between the sham/enriched and sham/standard groups. No significant group differences in swim speed were observed. At 14 days post-TBI, enriched animals had approximately twofold smaller lesion areas in regions of the cerebral cortex posterior to the injury epicenter (-4.5, -5.8, -6.8 mm relative to bregma; P < 0.05) compared to injured/standard animals. In addition, overall lesion volume for the entire injured cortical hemisphere was significantly smaller in animals recovering in the enriched environment. These results indicate that noninvasive environmental stimulation is beneficial in attenuating cognitive deficits and preserving tissue integrity in a TBI model which causes cerebral contusion and cell death. PMID- 11259126 TI - Mechanisms of estrogenic protection against gp120-induced neurotoxicity. AB - gp120, an HIV coat glycoprotein that may play a role in AIDS-related dementia complex (ADC), induces neuronal toxicity characterized by NMDA receptor activation, accumulation of intracellular calcium, and downstream degenerative events including generation of reactive oxygen species and lipid peroxidation. We have previously demonstrated estrogenic protection against gp120 neurotoxicity in primary hippocampal cultures. We here characterize the mechanism of protection by blocking the classical cytosolic estrogen receptors and by measuring oxidative end points including accumulation of extracellular superoxide and lipid peroxidation. Despite blocking ERalpha and ERbeta with 1 microM tamoxifen, we do not see a decrease in the protection afforded by 100 nM 17 beta-estradiol against 200 pM gp120. Additionally, 17alpha-estradiol, which does not activate estrogen receptors, protects to the same extent as 17beta-estradiol. 17beta-Estradiol does, however, decrease gp120-induced lipid peroxidation and accumulation of superoxide. Together the data suggest an antioxidant mechanism of estrogen protection that is independent of receptor binding. PMID- 11259127 TI - Enhancement of laminar FosB expression in frontal cortex of rats receiving long chronic clozapine administration. AB - The frontal cortex (FrC) and cingulate cortex (CgC) are critical sites for normal cognitive function, as well as cognitive dysfunction in schizophrenia. Thus, modulation of synaptic transmission within these cortical areas may, in part, account for the therapeutic actions of antipsychotic drugs such as haloperidol and clozapine. FosB and DeltaFosB are immediate-early gene (IEG) products sensitive to changes in response to chronic neuroleptic drug administration. We quantitatively examine whether there are light microscopic regional and/or laminar variations in FosB or DeltaFosB in the FrC or CgC of normal adult rats, or animals receiving 6 months administration of either drinking water clozapine, or depot haloperidol. Only animals receiving chronic haloperidol developed vacuous chewing movements, the equivalent of tardive dyskinesia in humans. In control animals, the deep and superficial layers of the FrC showed a higher area density of FosB, but not DeltaFosB immunoreactive cells than the medial layers of FrC or any of the CgC layers. In animals receiving clozapine, but not haloperidol there was increase in the area density of FosB immunoreactive neurons in all FrC layers, but the major increase occurs in medial layers. These findings suggest that FosB expression identifies those FrC neurons that are most active during normal waking behaviors and are further activated following chronic administration of atypical neuroleptics without motor side effects. The results also indicate that the actions of clozapine are attributed in large part to modulation of the output of frontal cortical pyramidal neurons residing in the medial layers. PMID- 11259128 TI - PP2A mRNA expression is quantitatively decreased in Alzheimer's disease hippocampus. AB - Since abnormal tau phosphorylation may play a role in neurofibrillary tangle (NFT) formation in aging and Alzheimer's disease (AD), we probed the distribution and abundance of protein phosphatase 2A (PP2A) catalytic (Calpha) and regulatory (PR55alpha and gamma, PR61varepsilon and delta) subunit mRNA in control and AD hippocampus using in situ hybridization. Quantitation of grain density per neuron area of PP2A subunits and beta-actin was determined for the CA3 region of hippocampus and cerebellum, while a qualitative assessment was performed for CA1, CA4, and dentate gyrus. All subunits are expressed in neurons, while PR55gamma and PR55alpha mRNA are also evident in glia. The expression levels of Calpha, all PP2A regulatory subunits studied, and beta-actin were similar in control and AD cerebellum. beta-Actin mRNA was, however, reduced in AD hippocampus. In addition to the generalized reduction of mRNA, as indicated by decreased beta-actin signal, there was a significant loss of Calpha, PR55gamma, and PR61epsilon mRNA in the CA3 hippocampus of AD. This study delineates the distribution of critical PP2A mRNAs and reveals a neuron- and subunit-specific reduction in PP2A catalytic and regulatory mRNA in AD hippocampus. This could result in decreased protein expression and phosphatase activity, leading to the hyperphosphorylation of tau and the formation of NFTs, as well as neuron degeneration in AD. PMID- 11259129 TI - Clinic-based cases with frontotemporal dementia show increased cerebrospinal fluid tau and high apolipoprotein E epsilon4 frequency, but no tau gene mutations. AB - Frontotemporal dementia (FTD) belongs to a group of neurodegenerative disorders known as tauopathies, characterized by intracellular aggregation of hyperphosphorylated tau protein in the brain. Some tauopathies, like Alzheimer's disease (AD), consistently show increased levels of tau protein in cerebrospinal fluid (CSF). However, similar studies in FTD populations have shown variable results, although mutations in the tau gene are identified as causes of disease in certain FTD families. In the present study, a Swedish clinic-based FTD population was investigated with respect to CSF tau levels, apolipoprotein E (APOE) genotype distribution and occurrence of mutations in the tau gene. CSF tau levels were significantly increased among FTD patients (534 +/- 235 pg tau/ml, P < 0.001) (n = 47) compared to controls (316 +/- 137 pg tau/ml) (n = 51). Furthermore, a strong increase in the APOE epsilon4 allele frequency was found in the FTD population, as 52% were epsilon4 carriers, compared to 21% of the controls. However, no mutations in the tau gene were identified. These findings support the present notion of a common pathogenic pathway in the disease processes for several tauopathies, with both APOE epsilon4 and CSF tau being a pathological link between the different disorders. Furthermore, we conclude that mutations in the tau gene are a rare cause of FTD. . PMID- 11259130 TI - Effects of an inhibitor of poly(ADP-ribose) polymerase, desmethylselegiline, trientine, and lipoic acid in transgenic ALS mice. AB - The development of transgenic mouse models of amyotrophic lateral sclerosis (ALS) allows the testing of neuroprotective agents. We evaluated the effects of five agents in transgenic mice with the G93A Cu,Zn superoxide dismutase mutation. A novel inhibitor of poly(ADP-ribose) polymerase showed no effects on survival. Desmethylselegiline and CGP3466 are agents that exert antiapoptotic effects in vitro by preventing nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase. They had no significant effects on survival in the G93A mice. Trientine, a copper chelator, produced a modest significant increase in survival. Similarly administration of lipoic acid in the diet produced a significant improvement in survival. These results therefore provide evidence for potential therapeutic effects of copper chelators and lipoic acid in the treatment of ALS. PMID- 11259131 TI - Retrograde tracing techniques influence reported death rates of adult rat nigrostriatal neurons. AB - Injury often causes loss of neuronal markers and prior retrograde labeling can circumvent this problem of identification. We have previously used a time consuming protocol for labeling all dopaminergic substantia nigra pars compacta neurons in adult rats by injecting the fluorescent tracer DiI into six sites throughout each neostriatum. Here, 2 weeks after injection of DiI into two central locations, only half of these nigrostriatal neurons were labeled. With six sites, more medial and lateral neurons were labeled, and also more in the midportion along the medial-lateral extent of the pars compacta. Less than 0.5% of the contralateral neurons were labeled. Two injections of Fluorogold also labeled fewer neurons, but their morphology was clearer. Two to 4 weeks after injection of the neurotoxin 6-OHDA into the two neostriatal sites, the total number of surviving neurons appeared greater with six sites of DiI than with two. However, within the middle region of the nigra, survival was lower with the six sites. This suggests that neurons that project outside the two central striatal tracer and 6-OHDA injection regions may be spared initially, but that those in the midportion that project to the central region are more vulnerable with the six-site protocol. Some reports suggest that Fluorogold prelabeling increases neuronal death. Here, survival after 6-OHDA or axotomy was similar with DiI or Fluorogold. These results suggest that because of a complex projection pattern of the nigrostriatal neurons, detailed quantification of neuronal survival should rely on extensive labeling. However, for drug screening purposes, faster labeling with Fluorogold using two sites is more suitable and should provide reliable data. PMID- 11259132 TI - Selective inhibition of MAO-B through chronic low-dose (R)-deprenyl treatment in C57BL/6 mice has no effect on basal neostriatal dopamine levels. AB - C. Thiffault, L. Lamarre-Theroux, R. Quirion, and J. Poirier (1997, Mol. Brain Res. 44: 238-244) recently reported that chronic treatment of young (12 week old) C57BL/6 mice with (R)-deprenyl, a mechanism-based inactivator of monoamine oxidase B (MAO-B), leads to a more than fourfold increase in neostriatal dopamine levels. Such an effect could complicate the interpretation of results obtained from mechanistic studies designed to evaluate the putative neuroprotective effects of (R)-deprenyl in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) lesioned mice. In contrast to the results of Thiffault et al., we have found that neostriatal dopamine levels in mature (32 week old) C57BL/6 mice were unaltered by chronic (R)-deprenyl treatment even though brain monoamine oxidase B activity was reduced by more than 80%. Neostriatal dopamine levels also were unaltered in both young and mature mice when the (R)-deprenyl treatment period was doubled compared to that reported by Thiffault et al. Consequently, studies on the putative neuroprotective properties of (R)-deprenyl in MPTP-lesioned mice are unlikely to be complicated by the possibility that inhibition of MAO-B alone will lead to an increase in neostriatal dopamine levels. PMID- 11259133 TI - Progesterone receptors as neuroendocrine integrators. AB - Intracellular progesterone receptors (PRs) are ligand-inducible transcription factors that mediate the majority of the effects of progesterone (P) on neuroendocrine functions. During the past decade, evidence has accumulated which suggest that PRs can also be activated independently of P, by signals propagated through membrane-bound receptors to the interior of cells. The activation of PRs by this type of "cross-talk" mechanism has been implicated in the physiological regulation of several important neuroendocrine processes, including estrous behavior and periovulatory hormone secretions. We review evidence that both ligand-dependent and ligand-independent activation of PRs occurs in central neurons and in anterior pituitary cells and that the convergence and summation of these signals at the PR serves to integrate neural and endocrine signals which direct several critically important neuroendocrine processes. An integrative function for PRs is reviewed in several physiological contexts, including the display of lordosis behavior in female rodents, the neurosecretion of gonadotropin-releasing hormone surges, secretion of preovulatory gonadotropin surges, and release of periovulatory follicle stimulating hormone surges. The weight of evidence indicates that cross talk at the intracellular PR is an essential component of the integrative mechanisms that direct each of these neuroendocrine events. The recurrence of PR's integrative actions in several different physiological contexts suggests that other intracellular steroid receptors similarly function as integrators of neural and endocrine signals in other neuroendocrine processes. PMID- 11259134 TI - Neurobiological bases underlying the control of the onset of puberty in the rhesus monkey: a representative higher primate. AB - The purpose of this article is to discuss our understanding of the neurobiological mechanisms that govern the timing of the onset of puberty in the rhesus monkey, a representative higher primate, and, whenever possible, to place findings obtained from studies of this macaque in perspective with those for the human situation. Specifically, the dynamics in the postnatal ontogeny of hypothalamic GnRH gene expression and release are described, and the roles of neuropeptide Y and gamma-aminobutyric acid in imposing the restraint on pulsatile GnRH release during juvenile development are examined. Finally, the hypothesis that circulating leptin provides the signal that times the reaugmentation of pulsatile GnRH release at the termination of juvenile development, and therefore triggers the onset of primate puberty, is discussed. PMID- 11259135 TI - Implications of multiple phenotypes observed in prolactin receptor knockout mice. AB - The development of a mouse line deficient in the PRL receptor (PRLR) would be an ideal means to better understand the multiple functions of prolactin. We were worried initially that removal of the PRLR from the mouse genome might be lethal and were surprised to find this not to be the case. We identified numerous deficiencies in PRLR knockout (KO) animals. Female homozygous mice are completely infertile and lack normal mammary development, while hemizygotes are unable to lactate following their first pregnancy. PRLR KO males and females have markedly elevated (30- to 100-fold) serum prolactin levels and in some instances pituitary hyperplasia is present. Maternal behavior is severely affected in both hemizygous and heterozygous animals. Bone formation is reduced in young animals and adults (males and females). Recently, we noticed that older KO animals show a slight reduction in body weight which appears to be due to reduced abdominal fat deposition. PMID- 11259136 TI - Good medicines for bad genes. PMID- 11259137 TI - A collective failure of medical practice? PMID- 11259138 TI - Mortality prediction in the elderly. PMID- 11259139 TI - Management of patients with heart failure: are internists as good as cardiologists? PMID- 11259140 TI - Catheter ablation of accessory pathways is associated with an excellent long-term prognosis. PMID- 11259141 TI - A novel strategy to maximize the efficacy of left ventricular assist devices as a bridge to recovery. PMID- 11259142 TI - Update on atrial remodelling owing to rate; does atrial fibrillation always 'beget' atrial fibrillation? PMID- 11259143 TI - Lifestyle and risk factor management and use of drug therapies in coronary patients from 15 countries; principal results from EUROASPIRE II Euro Heart Survey Programme. AB - AIMS: The principal aim of the second EUROASPIRE survey was to determine in patients with established coronary heart disease whether the Joint European Societies' recommendations on coronary prevention are being followed in clinical practice. METHODS: This survey was undertaken in 1999-2000 in 15 European countries: Belgium, Czech Republic, Finland, France, Germany, Greece, Hungary, Ireland, Italy, the Netherlands, Poland, Slovenia, Sweden, Spain and the U.K., in selected geographical areas and 47 centres. Consecutive patients, men and women < or =70 years were identified retrospectively with the following diagnoses: coronary artery bypass graft, percutaneous transluminal coronary angioplasty, acute myocardial infarction and myocardial ischaemia. Data collection was based on a review of medical records and interview and risk assessment at least 6 months after hospital admission. RESULTS: 8181 medical records (25% women) were reviewed and 5556 patients (adjusted participation rate 76%) interviewed. Recording of risk factor history and risk factor measurement in hospital notes was incomplete, particularly for discharge documents. At interview (median time 1.4 years after hospital discharge), 21% of patients smoked cigarettes, 31% were obese, 50% had raised blood pressure (systolic blood pressure > or =140 mmHg and/or diastolic blood pressure > or =90 mmHg), 58% had elevated serum total cholesterol (total cholesterol > or =5 mmol x l(-1)) and 20% reported a medical history of diabetes. Glucose control in these diabetic patients was poor with 87% having plasma glucose >6.0 mmol x l(-1)and 72% > or =7.0 mmol x l(-1). Among the patients interviewed the use of prophylactic drug therapies on admission, at discharge and at interview was as follows: aspirin or other antiplatelets drugs 47%, 90% and 86%; beta-blockers 44%, 66% and 63%; ACE inhibitors 24%, 38% and 38%; and lipid-lowering drugs 26%, 43% and 61%, respectively. With the exception of antiplatelet drugs, wide variations in the use of prophylactic drug therapies exist between countries. CONCLUSIONS: This European survey of coronary patients shows a high prevalence of unhealthy lifestyles, modifiable risk factors and inadequate use of drug therapies to achieve blood pressure and lipid goals. There is considerable potential throughout Europe to raise the standard of preventive cardiology through more effective lifestyle intervention, control of other risk factors and optimal use of prophylactic drug therapies in order to reduce coronary morbidity and mortality. PMID- 11259144 TI - Cardiovascular risk factors and 10-year all-cause mortality in elderly European male populations; the FINE study. Finland, Italy, Netherlands, Elderly. AB - BACKGROUND: This study aims to examine cardiovascular risk factors in relation to all-cause mortality in elderly populations of different European countries. METHODS: Men aged 65--84 years from defined administrative areas were enrolled in Finland (rural areas of east and west Finland; n=716), in the Netherlands (the town of Zutphen; n=887), and in Italy (the rural areas of Crevalcore and Montegiorgio; n=682). Ten-year all-cause mortality was studied in relation to measurements taken at entry: age, systolic blood pressure, HDL- and non-HDL cholesterol, body mass index, heart rate and smoking habits. Univariate and multivariate analyses were performed with all-cause mortality as the end-point. RESULTS: Ten-year death rates from all causes were higher in Finland (574 per 1000), lower in the Netherlands (475 per 1000), and Italy (466 per 1000). Age, heart rate and smoking in all three countries were independently associated with 10-year all-cause mortality. Non-HDL-cholesterol was not related with all-cause mortality. The observed associations between HDL-cholesterol, systolic blood pressure, body mass index and all-cause mortality were dependent on the in- or exclusions of early death. CONCLUSION: In these elderly men only age, smoking habits and heart rate were consistently associated with all-cause mortality. PMID- 11259145 TI - Indirect evidence for a role of a subpopulation of activated neutrophils in the remodelling process after percutaneous coronary intervention. AB - AIM: Leukocytes have been implicated in restenosis following percutaneous transluminal coronary angioplasty. We investigated the link between the activated status of circulating neutrophils and restenosis after angioplasty. METHODS AND RESULTS: The population of 108 patients with single, de novo lesions located in native coronary arteries were treated with elective balloon angioplasty (n=44) or stenting (n=64). Pre-, post-procedure and 6-month follow-up, angiograms were analysed by an independent core laboratory. Blood samples were collected immediately before treatment and the antigen CD66, which is specifically expressed by activated neutrophils, was measured. Overall, the average expression of CD66 was 6.4+/-3.6 of mean fluorescence intensity. In the stepwise linear regression model, which included biological, clinical and angiographic variables, absolute gain showed a direct association (P<0.001) with relative late loss (relative late loss=absolute late loss/pre-procedure reference diameter), whereas CD66 expression was inversely associated with relative late loss (P=0.004). CD66 expression also showed an inverse association with relative late loss in the balloon angioplasty treated patients (P=0.002, beta=-0.49). In the stent subgroup, only reference vessel diameter and acute gain were independent predictors of relative late loss. CONCLUSION: Our results confirm the beneficial role of activated neutrophils pre-procedure in the restenotic process after balloon angioplasty. The lack of a relationship between CD66 expression by neutrophils and relative late loss after stenting suggests that this leukocyte may be involved in the remodelling process. PMID- 11259146 TI - Statin therapy is associated with reduced restenosis rates after coronary stent implantation in carriers of the Pl(A2)allele of the platelet glycoprotein IIIa gene. AB - Aims Platelets play a central role in the restenosis process by inducing neointimal proliferation after coronary interventions. Glycoprotein IIb/IIIa Pl(A2)polymorphism has been associated with the occurrence of acute coronary syndromes and increased restenosis rates. Statins have been shown to exert potent antiproliferative, antiinflammatory and antithrombotic properties, thereby potentially interfering with the major processes of in-stent restenosis. Therefore, we sought to find out whether statin therapy interferes with restenosis and clinical outcome at 6 months following successful coronary stent implantation in the presence or absence of the Pl(A2)allele. Methods and Results Six hundred and fifty consecutive patients were followed for 6 months after coronary stent insertion. Carriers of the Pl(A2)allele demonstrated a significantly increased restenosis rate, which was abrogated by statin therapy (50.9% vs 28.6%, P=0.01). Moreover, statin therapy was associated with a significant reduction (28.2% vs 49.3%, P<0.01) in the occurrence of major adverse coronary events (myocardial infarction, cardiac death, target vessel revascularization) in the 6 months after the intervention in patients with the Pl(A2)allele. Conclusion Statin therapy reduces increased stent restenosis rates and improves clinical outcome following coronary stent implantation in patients bearing the Pl(A2)allele, suggesting that statins interfere with the functional consequence of a genetically determined platelet-mediated risk factor associated with Pl(A2)polymorphism. PMID- 11259147 TI - Specialty-related differences in the epidemiology, clinical profile, management and outcome of patients hospitalized for heart failure; the OSCUR study. Oucome dello Scompenso Cardiaco in relazione all'Utilizzo delle Risore. AB - AIMS: This study was designed to identify potential specialty-related differences in the epidemiology, clinical profile, management and outcome of patients hospitalized for congestive heart failure in departments of cardiology or internal medicine. METHODS AND RESULTS: From 1 July to 31 December 1998, we prospectively recorded epidemiological and clinical data from patients with congestive heart failure consecutively admitted to 11 departments of cardiology and 12 departments of internal medicine in Liguria, a northern area of Italy. The overall study population included 749 patients; 22% were treated by cardiologists and 78% by internists (P<0.0001). Patients managed by cardiologists were more likely to undergo echocardiography (92% vs 37%), Holter monitoring (25% vs 3%) and exercise stress testing (20% vs 0.5%) than those managed by internists (P=0.001). At discharge, patients treated by cardiologists were more likely to be prescribed beta-blockers (41% to 4%) and ACE inhibitors (100% to 74%) than those treated by internists (P<0.0001), and the latter medication at higher dosages by cardiologists than internists. In addition, patients followed by cardiologists were younger (70+/-9 to 79+/-1 years;P<0.0001), more likely to be male (61% to 50%;P=0.011) and to have coronary artery disease (57% to 45%;P<0.006) than those followed by internists. Conversely, patients followed by internists were more likely to have diabetes, chronic obstructive pulmonary disease, atrial fibrillation and renal failure (P<0.03). In the overall study population, 53 patients (7%) died during hospitalization. Patients treated by cardiologists had a mortality not significantly different from that of patients treated by internists (10% and 6%, respectively;P=0.067), although congestive heart failure was more severe on admission in patients treated by cardiologists. CONCLUSION: Cardiologists follow published guidelines for congestive heart failure more strictly than internists, but treat a smaller number of patients who are younger, have more severe congestive heart failure and fewer co-morbidities than those managed by internists. PMID- 11259148 TI - Late clinical outcome after successful radiofrequency catheter ablation of accessory pathways. AB - AIMS: To evaluate the long-term clinical results of patients who underwent successful radiofrequency catheter ablation of a symptomatic drug-resistant accessory-pathway-mediated tachycardia. METHODS AND RESULTS: Clinical follow-up was done by direct contact with the patients and their physicians. One hundred and eighty consecutive patients (113 males, 67 females) were followed during a median period of 48.1 months. There were seven procedure related complications (4%). During the follow-up period, 79% of the patients remained asymptomatic; 14% complained of short bouts of palpitations due to isolated or short runs of atrial or ventricular premature beats; 7% had sustained palpitations due either to accessory pathway recurrence (4%) or supraventricular tachyarrhythmias not associated with an accessory pathway (3%). Symptoms due to accessory pathway recurrence appeared either in the first month following the ablation or at least later than 3 months when sustained supraventricular arrhythmias occurred related to another cause. CONCLUSIONS: Initially successful radiofrequency catheter ablation has a low, long-term recurrence rate (4%). Recurrence of accessory pathway-mediated tachycardia is observed during the first month while later symptoms suggest supraventricular arrhythmias from another cause. PMID- 11259149 TI - Sudden cardiovascular death in correlation with sexual activity -- results of a medicolegal postmortem study from 1972--1998. PMID- 11259150 TI - Arginine intake and 25-year CHD mortality: the Seven Countries Study. PMID- 11259151 TI - Men with coronary artery disease have lower levels of androgens than men with normal coronary angiograms. PMID- 11259153 TI - Comment on the ESC/ACC redefinition of myocardial infarction by a consensus dissenter. PMID- 11259155 TI - Very short telomeres in the peripheral blood of patients with X-linked and autosomal dyskeratosis congenita. AB - Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome in which patients undergo premature ageing and have a predisposition to malignancy. X linked and autosomal (dominant and recessive) forms of the disease are recognized. The gene responsible for X-linked DC (DKC1) encodes a highly conserved protein called dyskerin that is believed to be essential in ribosome biogenesis and may also be involved in telomerase RNP assembly. Here we show that in X-linked DC, peripheral blood cells have dramatically reduced telomere lengths but normal levels of telomerase activity. We also find that subjects with autosomal DC have significantly shorter telomeres than age-matched normal controls suggesting that both forms of the disease are associated with rapid telomere shortening in hemopoietic stem cells. The further characterization of these genes will not only lead to a better understanding of the biology of DC but may also provide further insights into the maintenance of telomeres and the biology of aplastic anemia, ageing, and cancer. PMID- 11259156 TI - Viral infections and phenotypic heterogeneity in Gaucher disease. AB - Gaucher disease, the most common lysosomal storage disorder, is remarkable for its tremendous phenotypic heterogeneity even among patients with the same genotype. Beyond mutations at the 1q21 locus, there may be other genetic and environmental factors that impact on the natural course of Gaucher disease and indeed may trigger symptoms and signs. Among candidate events are viral infections such as the Epstein-Barr virus (EBV) or cytomegalovirus (CMV). The purpose of this study was to ascertain if indeed prior infection with EBV or CMV in patients homozygous for the most common mutation, N370S (1226G), is predictive of a more severe phenotype. Evidence for an EBV virus was IgG and IgM antibodies to early antigen and IgG anti-EBNA. For CMV infection, IgG and IgM antibodies were sought. This study failed to demonstrate any correlation between prior EBV or CMV infection and clinical course of Gaucher disease in patients homozygous for the N370S (1226G) mutation. The only positive finding was a higher level of anti-EBNA antibodies among patients with moderate/severe disease. In conclusion, other than a small subset of patients who showed a pattern comparable to immunosuppression, there was no association between severity of Gaucher disease and prior EBV or CMV infection. PMID- 11259157 TI - Population genetics of factor V Leiden in Europe. AB - We have analyzed 5971 control individuals originating from 26 geographically defined populations in Europe and neighboring countries for the presence of factor V Leiden, an important genetic risk factor in venous thromboembolism. The mean frequency of the mutation in the populations studied is 0.027%, with a peak value incidence of more than 12% in Cyprus; the mutation is absent in Inuits and is at low incidence in French Basques. A west to east increasing cline of allele factor V Leiden prevalence (r = 0.479, p < 0.02) was observed in Europe, together with a decreasing south to north cline (r = -0.801, p < 0.001) of these values but in this last situation only when southwest populations are excluded from the analysis. Based on these findings, an interpretation is proposed of the history of factor V Leiden expansion in Europe during the Neolithic period, from a probable Anatolian center of origin in Turkey. PMID- 11259158 TI - AML1 and AML1 fusion protein AML1-ETO in myeloid gene regulation and leukemogenesis. PMID- 11259159 TI - Disparate regulation of human fetal erythropoiesis by the microenvironments of the liver and bone marrow. AB - The liver and the bone marrow (BM) are the major organs that support hematopoiesis in the human fetus. Although both tissues contain the spectrum of hematopoietic cells, erythropoiesis dominates the liver. Previous studies suggested that a unique responsiveness of fetal burst-forming units erythroid (BFU-E) to erythropoietin (EPO) obviates the need for cytokines with burst promoting activity (BPA) in fetal erythropoiesis. This potential regulatory mechanism whereby fetal erythropoiesis is enhanced was further investigated. Fluorescence-activated cell sorting was used to isolate liver and BM progenitors based on their levels of CD34 and CD38 expression. The most mature population of CD34+ lineage (Lin-) cells was also the most prevalent of the three subpopulations and contained BFU-E responsive to EPO alone under serum-deprived conditions. Kit ligand (KL) also strongly synergized with EPO in stimulating the growth of these BFU-E. An intermediate subset of CD34++CD38+Lin- cells contained erythroid progenitors responsive to EPO alone, but also displayed synergism between EPO and KL, granulocyte-macrophage colony-stimulating factor (GM-CSF), or interleukin (IL)-3, demonstrating that erythroid progenitors that respond to cytokines with BPA do exist in fetal tissues as in the adult BM. Candidate stem cells (CD34++CD38-Lin- cells) did not respond to EPO. Synergisms among KL, GM CSF, and IL-3, and to a lesser extent granulocyte colony-stimulating factor (G CSF) and FLK-2/FLT-3 ligand (FL), supported the growth of primitive multipotent progenitors that became responsive to EPO. These data define the limits of EPO activity in fetal erythropoiesis to cells that express CD38 and demonstrate the potential for various cytokine interactions to be involved in regulating fetal erythropoiesis. Furthermore, a comparison of the responses of liver and BM erythroid progenitors revealed similarity in their responses to cytokines but a difference in the frequency of BFU-E among the three subpopulations examined. A higher frequency of BFU-E among the intermediate and late progenitor subsets in the liver indicates that regulatory factors acting on stem cells and their immediate progeny are partially responsible for the high content of erythropoiesis in the liver. These data implicate a critical role for the microenvironments of the liver and BM in regulating the disparate levels of erythropoiesis in these tissues. PMID- 11259160 TI - Effect of tetracaine chlorhydrate on the mechanical properties of the erythrocyte membrane. AB - Pharmacologically active agents that locate in the cell membrane are useful tools to investigate the interactions taking place between its molecular components. In the present work, the effect of tetracaine chlorhydrate (Tc) on the membrane mechanical properties of intact and desialated erythrocytes was studied. Our results evince the complex interaction between the drug and the membrane structures. The effect of Tc on erythrocyte shape suggests that this drug locates in the inner hemilayer of the lipid bilayer. Since Tc also modifies osmotic fragility and mechanical properties ascribed to the cytoskeleton, it can be inferred that the lipid bilayer has an effect on the rheology of the membrane, in a direct or indirect way, in this case through the close interaction with the structural proteins. Moreover, our results support the hypothesis of a second localization of the drug in the membrane, i.e., as monovalent cations intercalated among the glycocalix sialic endings, where it generates an effect superimposed on that produced from its typical site in the lipid bilayer. PMID- 11259161 TI - Increase in band 3 density and aggregation in hereditary spherocytosis. AB - PURPOSE: Red cells in hereditary spherocytosis are characterized by a reduced surface area/volume ratio. The mechanisms leading to the loss of membrane material and subsequent elimination of the cells have still not been clarified. It was the aim of the present study to analyze band 3 distribution in the red cell membrane and its putative role in red cell elimination. METHODS/RESULTS: Immunogold histochemistry was performed to detect band 3 in red cell membranes. Band 3 density and distribution were visualized by electron microscopy. Unsplenectomized spherocytosis patients (n = 12) showed reduced band 3 density and aggregation compared to controls (n = 15) (density: 1.2 +/- 0.1 gold particles/microm circumference of red cell membrane vs 1.5 +/- 0.07 gold particles/microm, x +/- SEM; P < 0.05; aggregation: 0.26 +/- 0.02 aggregates/microm vs 0.3 +/- 0.02 aggregates/microm). By contrast, band 3 density and aggregation were increased in spherocytosis patients who had undergone splenectomy (density: 2.8 +/- 0.1 gold particles/microm vs 2.0 +/- 0.1 gold particles/microm; P < 0.05; aggregation: 1.5 +/- 0.1 aggregates/microm vs 0.5 +/- 0.03 aggregates/microm; P < 0.01). Artificial ageing of red cells from healthy controls (n = 6) led to a significant increase in band 3 aggregation (2.06 +/- 0.2 aggregates/microm vs 0.33 +/- 0.1 aggregates/microm; P(Wilcoxon) < 0.01) but no change in band 3 density. In hereditary spherocytosis (n = 6), both band 3 density and aggregation increased significantly after artificial ageing of the red cells. The elevated band 3 aggregation was associated with a stimulated erythrophagocytosis in vitro. CONCLUSION: Band 3 aggregation characterizes the red cells in hereditary spherocytosis. It may be the cause of selective splenic phagocytosis of both spherocytes and senescent erythrocytes. PMID- 11259162 TI - The Second International Workshop on Myb Genes: foreword. PMID- 11259163 TI - Identification and validation of candidate Myb target genes. AB - While a considerable number of candidate Myb target genes have been reported to date, most of these are likely to play little or no role in transformation by myb oncogenes. Here we have used a conditionally myb-transformed myeloid cell line (ERMYB) to further examine Myb regulation of one candidate target gene--c-myc- that has the potential to affect cell proliferation. It was found that the major influence on c-myc expression was the presence of cytokine (GM-CSF) rather than Myb activity. We also describe the application of PCR-based subtractive hybridization and low-density cDNA array screening, in conjunction with the ERMYB line, to the identification of additional Myb target genes. Preliminary identification of a number of candidates is reported; these include myeloperoxidase, which is known to have essential Myb-binding sites in its regulatory region. PMID- 11259164 TI - Regulation of the cell cycle by B-Myb. AB - B-Myb is a cell-cycle-regulated member of the Myb transcription factor and, like c-Myb, has been implicated in regulation of hematopoietic cell proliferation and differentiation. In this study we have examined the mechanisms by which B-Myb regulates the cell cycle. We found that the ability of B-Myb both to promote Saos 2 cells into the S phase of the cell cycle and to overcome G1 arrest mediated by overexpression of the retinoblastoma-related p107 protein was correlated with the capacity of B-Myb to form an in vivo complex with p107, but was independent of its transactivation function. Further experiments using a B-Myb dominant-negative protein suggested that transcriptional activation of genes regulated through Myb DNA-binding sequences was required for cell proliferation. Our experiments suggest, therefore, that B-Myb influences cell cycle progression at two distinct levels: by inhibiting p107 and by inducing transcription of specific target genes. PMID- 11259165 TI - Phosphorylation-dependent conformation and proteolytic stability of c-Myb. AB - The c-Myb oncoprotein is a critical regulator of hematopoietic cell proliferation and differentiation. Normal c-Myb is rapidly degraded by the ubiquitin-26S proteasome pathway, and instability determinants have been localized within the negative regulatory domain in the carboxyl terminus. Our recent work has shown that, in myeloid cells, inhibition of cellular Ser/Thr protein phosphatases with okadaic acid (OA) causes a rapid increase in c-Myb phosphorylation and 26S proteasome-dependent breakdown [J. Bies, S. Feikova, D. P. Bottaro, and L. Wolff (2000) Oncogene 19, 2846-2854]. Furthermore, phosphoamino acid analyses revealed that the increase in phosphorylation was mainly on threonine residues. Here we investigated the ability of c-Myb to bind DNA following phosphorylation. Our results suggest that the hyperphosphorylated form of c-Myb binds to DNA with affinity very similar to the hypophosphorylated form. Therefore, the increased proteolytic instability of the former cannot be explained by a difference in DNA binding capacity. Conformational changes in the carboxyl terminus were proposed previously to be a consequence of phosphorylation because we observed phosphorylation-induced alterations in gel electrophoresis mobilities and alterations in recognition by specific monoclonal antibodies. Further support for this notion has come from this study, in which we have detected new degradation products in electrophoretic mobility shift assays, as well as an increased rate of in vitro proteolysis, following OA treatment. We speculate that these alterations in the conformation of the negative regulatory domain expose epitopes on the surface of c-Myb, which in turn can serve as recognition signal(s) for ubiquitin-26S proteasome proteolytic machinery. PMID- 11259166 TI - An ex vivo model to study v-Myb-induced leukemogenicity. AB - The v-myb(AMV) oncogene transforms myelomonocytic cells in vitro and induces acute monoblastic leukemia in chickens. We analyzed the activity of the evolutionarily conserved PEST-like domain (P1 domain) for biochemical and biological activities of v-Myb in ex vivo cultures and in vivo. Deletion of the P1 domain did not affect v-Myb transcriptional activity, intracellular stability, or subcellular localization. However, it resulted in subtle yet important changes in biological activities. Although the mutant DeltaP1 v-Myb protein blocked the terminal differentiation of the monocyte/macrophage lineage as efficiently as the wild type (wt) in ex vivo cultures, it failed to induce the acute phase of monoblastic leukemia, with its fatal consequences, in vivo. Interestingly, in DeltaP1 v-myb-infected animals large numbers of monoblasts, comparable to those induced by wt v-myb, were present in the bone marrow but very few were found in the peripheral blood. The comparison of ex vivo wt- and DeltaP v-Myb bone marrow cells revealed several important features of v-Myb transformation: (i) the proliferation of transformed monoblasts is not an apparent consequence of the differentiation block with these processes being at least in part independent; (ii) the P1 domain is required for proliferation of v-Myb-mediated transformed monoblasts; (iii) the mechanism which renders transformed cells growth factor independent does not involve activation of an autocrine growth factor loop; and (iv) deletion of the P1 domain affects self-adhesion properties of v-myb transformed monoblasts as well as their interaction with bone marrow stromal cells. These data indicate that the DeltaP1 v-myb mutant and ex vivo bone marrow cell cultures represent a valuable tool for studies on the mechanisms of leukemia formation. PMID- 11259167 TI - Functional evolution of the Myb oncogene family. AB - Three Myb-related genes (A-Myb, B-Myb, and c- Myb) have been found in all vertebrates examined thus far including mammals, birds, and amphibians. Two invertebrates, the sea urchin and the fruit fly, have only one Myb-related gene. Our laboratory has used Drosophila as a model system to explore the function of its sole Myb gene. We have also reintroduced the three different vertebrate Myb genes into Drosophila in order to begin to understand how their different functions may have arisen following gene duplication during evolution. PMID- 11259168 TI - The conserved DNA binding domain mediates similar regulatory interactions for A Myb, B-Myb, and c-Myb transcription factors. AB - The vertebrate A-Myb, B-Myb, and c-Myb proteins comprise a family of related transcription factors that share a highly conserved DNA binding domain. Although all three proteins are capable of binding the same sites in DNA, they have distinct, but overlapping patterns of expression and are presumed to be regulated independently. Here we show that the transcriptional activity of all three vertebrate Myb proteins can be severely inhibited by coexpression of a dominant negative allele of p100, a coactivator protein that interacts with Myb DNA binding domains. Thus, the conserved Myb domains mediate interactions with common sites in DNA, as well as common regulators, suggesting that the proteins provide alternative or complementary responses to common upstream signaling pathways. PMID- 11259169 TI - Characterization of direct readout contacts of the Myb DNA-binding domain. AB - The Myb protein contacts its recognition sequence by means of direct protein-DNA interactions. We used site-directed mutagenesis in order to substitute amino acids crucial for these contacts and probed the mutant proteins for their DNA binding and transactiving activities. We could show that amino acids involved in direct readout contacts do not contribute equivalently in the recognition process. PMID- 11259170 TI - Defective stem cell factor expression in c-myb null fetal liver stroma. AB - High levels of c-Myb are observed in immature precursor myeloid and lymphoid cells, while downregulation of c-myb accompanies terminal differentiation to a mature phenotype. This has established c-Myb as a crucial transcription factor for hematopoiesis. Further evidence for this is the embryonic death of the c-myb homozygous mutant mouse at ED15 due to defective fetal liver erythropoiesis. Cells from fetal liver of wild-type and c-myb-/- embryos were examined in detail for their hematopoietic potential and the capacity of the stroma to support wild type hematopoiesis. The c-myb-/- fetal liver was shown to harbor sevenfold fewer spleen focus-forming cells and a similarly lower number of cells with long-term repopulating capacity (high proliferative potential cells). However, shorter term repopulating cells were not substantially reduced. c-myb-/- stromal cells were unable to support the proliferation of wild-type bone marrow lineage-negative cells. This was found to be partly due to a decrease in stem cell factor (SCF) expression while partial rescue of the stromal cell cultures was achieved through the addition of exogenous SCF. DNA binding studies for two sites within the SCF promoter demonstrated an in vitro interaction between the SCF promoter and c-Myb and transient transfection studies demonstrated that c-Myb could substantially transactivate the SCF promoter in HEK293 cells. These data explain why the c-myb /- embryos are so impaired in their ability to establish hematopoiesis. PMID- 11259171 TI - Three genes with different functions in transformation are regulated by c-Myb in myeloid cells. AB - The proto-oncogene c-myb is constitutively expressed in murine leukemia virus induced myeloid leukemia (MML) due to the integration of virus at this locus. Our recent focus has been the determination of genes regulated by this transcription factor that may be involved in transformation. Data presented here, using conditional expression of Myb in myeloid cells, show that c-Myb directly transactivates the endogenous c-myc and Bcl-2 genes, which explains at least in part how c-Myb regulates proliferation and survival. In addition, c-Myb prevents expression at the RNA level of the tumor suppressor INK4b gene. This gene encodes a cyclin-dependent kinase inhibitor, p15INK4b, that is normally upregulated at the mRNA level during myeloid differentiation and promotes growth arrest. The MMLs are generally characterized as differentiated monocytic tumors and possess the phenotype that is normally associated with p15INK4b expression. c-Myb inhibits expression of this gene, however, and therefore acts to promote a pathway which is abnormal in mature cells. This activity of c-Myb collaborates with its maintenance of c-myc expression to promote growth. PMID- 11259172 TI - Identification and characterization of a novel mutation c.1090G>T (G325W) and nine common mutant alleles leading to Gaucher disease in Spanish patients. AB - BACKGROUND: Gaucher disease is an autosomal recessive disorder resulting from mutations in the glucocerebrosidase gene (GBA). The lack of full genotype/phenotype correlation complicates counseling regarding clinical outcome and treatment recommendations. SUBJECTS AND METHODS: Several mutations in the human beta-glucosidase gene associated with Gaucher disease in 16 Spanish families were identified utilizing a combination of methods: enzymatic restriction, PCR-SSCP, and sequence analyses. Expression studies were performed following the introduction of the mutagenized human acid beta-glucosidase cDNA into COS-1 cells, and the residual enzyme activities of the mutant protein were measured and compared with the normal cDNA. RESULTS: The identified mutations and corresponding residual enzyme activities of the expressed protein are as follows: c.517A>C (T134P), 1%; c.721G>A (G202R), 17%; c.1090G>T (G325W), 13.9%; c.1093G>A (E326K), 26%; c.1208G>A (S364N), 4.1%; c.1226A>G (N370S), 17,8%; c.1246G>A (G377S), 17.6%; c.1289C>T (P391L), 8.5%; c.1448T>C (L444P), 3%; and c.1504C>T (R463C), 24.5%. CONCLUSIONS: Site-directed mutagenesis and expression in COS-1 cells are useful methods to increase our understanding of causality in Gaucher disease and the correlation between disease severity, gene defects, and residual enzyme activity. Our study demonstrates the functional consequences of the identified human beta-glucosidase mutations (T134P, S364N, G377S, P391L, and G325W) and provide evidence for the molecular and biochemical basis of Gaucher disease, among patients of Spanish ancestry. PMID- 11259173 TI - Elevating the terms of the GM food debate. PMID- 11259174 TI - Safety evaluation of phosphodiesterase produced from Penicillium citrinum: summary of toxicological data. AB - The toxicity of Enzyme RP-1, an enzyme preparation used to hydrolyze yeast RNA to produce flavor enhancers for use in the food industry, was evaluated in a series of studies. A 5-week dietary toxicity study in Wistar rats was conducted in which animals received Enzyme RP-1 in feed at concentrations of 0, 500, 2000, or 8000 mg/kg body wt/day. A 13-week dietary toxicity study in Sprague-Dawley rats was conducted in which animals received RP-1 concentrate at 0, 0.125, 0.5, or 2% in their diets. At the highest dose levels in both studies, submandibular glands in the oral cavity were enlarged, an effect attributed to protease activity of the enzyme preparation. The no-observed-effect level in rats in the 13-week study was 0.5%, equivalent to 317 mg/kg body wt/day for males and 346 mg/kg body wt/day for females. Based on estimated dietary exposure to the enzyme preparation, the margin of exposure is estimated to be greater than 38,000. Lack of genotoxic potential was demonstrated by an in vitro reverse mutation assay in Salmonella typhimurium strains TA98, TA100, TA1535, and TA1537 and in Escherichia coli strain WP2uvr and by an in vitro chromosome aberration test in CHL/IU cells derived from fibroblasts from the lungs of Chinese hamsters. Finally, the particular strain of Penicillium citrinum, the fungal strain used to prepare Enzyme RP-1, was shown to have low pathogenicity upon a single injection into the tail vein of rats of viable spores at doses up to 2.8x10(5) colony-forming units per animal. The results of these studies demonstrate that the enzyme preparation may be considered safe to workers and consumers when employed in the production of flavor enhancers from yeast. PMID- 11259175 TI - Physiological-model-based derivation of the adult and child pharmacokinetic intraspecies uncertainty factors for volatile organic compounds. AB - The intraspecies uncertainty factor (UF(HH)=10x) is used in the determination of the reference dose or reference concentration and accounts for the pharmacokinetic and pharmacodynamic heterogeneity within the human population. The Food Quality Protection Act of 1996 mandated the use of an additional uncertainty factor (UF(HC)=10x) to take into account potential pre- and postnatal toxicity and lack of completeness of the data with respect to exposure and toxicity to children. There is no conclusive experimental or theoretical justification to support or refute the magnitude of the UF(HH) and UF(HC) nor any conclusive evidence to suggest that a factor of 100 is needed to account for intrahuman variability. This study presents a new chemical-specific method for estimating the pharmacokinetic (PK) component of the interspecies uncertainty factor (UF(HH-PK) and UF(HC-PK)) for volatile organic compounds (VOCs). The approach utilizes validated physiological-based pharmacokinetic (PBPK) models and simplified physiological-model-based algebraic equations to translate ambient exposure concentration to tissue dose in adults and children the ratio of which is the UF(HH-PK) and UF(HC-PK). The results suggest that: (i) the UF(HH-PK) and UF(HC-PK) are chemical specific; (ii) for the chemicals used in this study there is no significant difference between UF(HH-PK) and UF(HC-PK); (iii) the magnitude of UF(HH-PK) and UF(HC-PK) varies between 0.033 and 2.85 with respect to tissue and blood concentrations; (iv) the body weight, the rate of ventilation, the fraction of cardiac output flowing to the liver, the blood : air partition coefficient, and the hepatic extraction ratio are the only parameters that play a critical role in the variability of tissue and blood doses within species; and (v) the magnitude of the UF(HH-PK) and UF(HC-PK) obtained with the simplified steady-state equations is essentially the same with that obtained with PBPK models. Overall, this study suggests that no adult-children differences in the parent chemical concentrations of the VOCs are likely to be observed during inhalation exposures. The physiological-model-based approaches used in the present study to estimate the UF(HH-PK) and UF(HC-PK) provide a scientific basis for their magnitude. They can replace the currently used empirical default approaches to provide chemical-specific UF(HH-PK) in future risk assessments. PMID- 11259176 TI - Evaluation of potential neurotoxic effects of occupational exposure to (L) lactates. AB - Organo psycho syndrome (OPS) or chronic toxic encephalopathy (CTE) is a neurotoxic condition reported following long-term exposure to paints containing organic solvent and to other solvents. Lactate esters are finding wider use as solvents. Lactate esters have been well studied in standard toxicity tests, but specific neurotoxicity studies have not been conducted. No clinical signs of chronic neurotoxicity have been observed in standard toxicity tests. Lactate esters are rapidly hydrolyzed in the body to lactic acid and the corresponding alcohol. Alcohols have been reported to have acute neurotoxic effects, usually following high levels of ingestion. The literature on alcohols was reviewed to establish the no-observed-adverse-effect level (NOAEL) for acute neurotoxicity and to look for any evidence of chronic neurotoxicity from the alcohols produced by hydrolysis of the lactate esters. The NOAELs were compared with the potential amounts of alcohol produced by hydrolysis of different lactate esters at 200 mg//m(3) (the NOAEL for most of the lactate esters). In all cases neither acute nor chronic neurotoxicity would be expected based on the amounts of alcohol produced by hydrolysis of the lactate esters at their NOAELs. L-Lactic acid is a normal metabolite in the body and is not considered neurotoxic. Based on this information there is no evidence to suggest that L-lactate esters can cause any chronic neurotoxicity, OPS, or CTE. PMID- 11259177 TI - How useful is linear regression analysis in detecting the existence of dose response relationships in large-scale epidemiologic studies when only a fraction of the population is sensitive? The case of methylmercury. AB - The existence of a dose response in epidemiologic studies is generally determined from the linear regression slope after controlling for covariates. This approach assumes the entire population is equally sensitive to the toxicant and that response is a function only of dose and a random error function. However, sensitive subpopulations have been identified for a variety of toxicants possibly including methylmercury (MeHg). The study of MeHg exposure in the Seychelles Islands has failed to find significant effects (dose-response slope not significantly different from zero) while other studies have found such effects. Using data on the error function in developmental test scores and MeHg exposure distributions from that study, and assuming plausible dose-response relationships for sensitive subpopulations, we conducted Monte-Carlo simulations of the power of linear regression analysis to detect a dose-response relationship from the total sample (n=700), and to compare dose-response slopes in the total and sensitive populations. Linear regression did not reliably detect a dose-response relationship for most scenarios when sensitives were 5% of the total and for some scenarios when sensitives were 10% of the total. We also found that the dose response slope for the total population underestimated the sensitive dose response slope in all cases by about an order of magnitude. These findings may have important implications for detection and quantification of dose-response relationships from epidemiologic studies. PMID- 11259178 TI - Evaluation of subchronic toxicity data using the benchmark dose approach. AB - We used the benchmark dose (BMD) methodology devised by Crump (Fundam. Appl. Toxicol. 4, 854-871, 1984) to estimate BMDs for 90-day toxicological data and several fabricated data sets. From a toxicological perspective, dose-response modeling offers certain advantages over using a point estimate, such as the currently used no-observable-adverse-effect level (NOAEL) approach. However, there are many variables associated with the BMD that could be set to produce unreasonable BMD estimates. Some of these variables and decisions are examined in this study. BMDs were calculated for discrete and continuous endpoints using a variety of different variables (e.g., maximum likelihood estimates [MLEs], lower confidence limits [LCLs], and different risk levels). In addition, the fabricated data sets were manipulated (i.e., dose groups eliminated) and the BMDs recalculated. This process tested how the BMD estimates varied using different forms of the data. For the 90-day toxicological studies, the BMDs were typically within an order of magnitude of the NOAEL for discrete endpoints. For the discrete endpoints, the MLEs were typically greater than the NOAEL and the LCLs were typically less than the NOAEL. The BMD was insensitive to changes in the data points one to two dose groups beyond the NOAEL/LOAEL. With the continuous data, the ratios of MLEs and LCLs to the NOAEL were highly variable, and no general trend could be determined. The BMD methodology offers potential improvements in the risk assessment process since dose-response characteristics are used to calculate the BMD. Depending upon how the BMD is defined, i.e., the form of the dose-response model, and how the BMD is used in the risk assessment process, BMD estimates may produce reference doses/concentrations that are more or less conservative than the NOAEL approach. Active involvement in discussions with regulatory agencies is needed to ensure that inappropriate models and unreasonable BMDs are not used. In addition, further discussions on how BMDs should be used in the risk assessment process are needed. PMID- 11259179 TI - Development of a metabolism factor for polycyclic aromatic hydrocarbons for use in multipathway risk assessments of hazardous waste combustion facilities. AB - As a state authorized to permit hazardous waste combustion facilities, the Texas Natural Resource Conservation Commission (TNRCC) has conducted several site specific, multipathway risk assessments in support of permitting actions. These risk assessments were carried out in accordance with guidance provided in U.S. Environmental Protection Agency's Human Health Risk Assessment Protocol for Hazardous Waste Combustion Facilities, Peer Review Draft (HHRAP). This protocol uses a prospective, screening-level, risk assessment paradigm and incorporates conservative default assumptions. In conducting such risk assessments, staff of the Toxicology and Risk Assessment Section have found that modeled emissions of polycyclic aromatic hydrocarbons (PAHs) at concentrations near or below detectable levels can produce unacceptable risk in the "farmer" exposure scenario. In accordance with recommendations in HHRAP, the TNRCC followed a tiered approach, whereby additional site-specific information is incorporated in place of standard default assumptions in cases where the initial screening-level risk assessment yielded unacceptable risk or hazard. Sensitivity analyses indicated that one of the key parameters affecting the risk estimates for PAHs in the farmer exposure scenario was the way in which PAHs were modeled up through the food chain. Therefore, refinement of the model focused on the development of a PAH-specific metabolism factor, as described in the article. PMID- 11259180 TI - Safety evaluation of dietary aluminum. AB - Aluminum is a nonessential metal to which humans are frequently exposed. Aluminum in the food supply comes from natural sources, water used in food preparation, food ingredients, and utensils used during food preparations. The amount of aluminum in the diet is small, compared with the amount of aluminum in antacids and some buffered analgesics. The healthy human body has effective barriers (skin, lungs, gastrointestinal tract) to reduce the systemic absorption of aluminum ingested from water, foods, drugs, and air. The small amount of aluminum (<1%) that is systemically absorbed is excreted principally in the urine and, to a lesser extent, in the feces. No reports of dietary aluminum toxicity to healthy individuals exist in the literature. Aluminum can be neurotoxic, when injected directly into the brains of animals and when accidentally introduced into human brains (by dialysis or shrapnel). A study from Canada reports cognitive and other neurological deficits among groups of workers occupationally exposed to dust containing high levels of aluminum. While the precise pathogenic role of aluminum in Alzheimer's disease (AD) remains to be defined, present data do not support a causative role for aluminum in AD. High intake of aluminum from antacid for gastrointestinal ailments has not been reported to cause any adverse effects and has not been correlated with neurotoxicity or AD. Foods and food ingredients are generally the major dietary sources of aluminum in the United States. Cooking in aluminum utensils often results in statistically significant, but relatively small, increases in aluminum content of food. Common aluminum-containing food ingredients are used mainly as preservatives, coloring agents, leavening agents, anticaking agents, etc. Safety evaluation and approval of these ingredients by the Food and Drug Administration indicate that these aluminum-containing compounds are safe for use in foods. PMID- 11259181 TI - Evaluation of health aspects of kojic acid in food. AB - Kojic acid is a fungal metabolite commonly produced by many species of Aspergillus, Acetobacter, and Penicillium. The Aspergillus flavus group has traditionally been used in the production of a number of foods, including miso (soybean paste), shoyu (soy sauce), and sake. Kojic acid is widely used as a food additive for preventing enzymatic browning, and in cosmetic preparations as a skin-lightening or bleaching agent. Because kojic acid is often produced during the fermentation of historically used dietary staples, it has a long history of consumption. Various types of compounds, such as glucose, sucrose, acetate, ethanol, arabinose, and xylose, have been used as carbon sources for kojic acid production. Different Aspergillus species are known to produce variable amounts of kojic acid. The mechanism of action of kojic acid is well defined and it has been shown to act as a competitive and reversible inhibitor of animal and plant polyphenol oxidases, xanthine oxidase, and D- and some L-amino acid oxidases. The structure of kojic acid indicates a relatively simple route of metabolism much like dietary hexoses. Acute or subchronic toxicity resulting from an oral dose has not been reported, but convulsions may occur if kojic acid is injected. Results of mutagenicity studies are mixed, but in the in vivo mammalian dominant lethal assay, kojic acid was proven negative. Continuous administration of high doses of kojic acid in mice resulted in induction of thyroid adenomas in both sexes. Kojic acid reversibly affects thyroid function primarily by inhibiting iodine uptake, leading to decreases in T3 and T4 and increase in TSH. Increased TSH from pituitary gland in turn stimulates thyroid hyperplasia. Several lines of evidence indicate that the proliferative effects of kojic acid on thyroid are not related to a genotoxic pathway. The risk of functional inhibition of iodine uptake and its metabolism (organification) and thyroid tumor induction by kojic acid in humans appears to be extremely low. Based on the literature reviewed and discussed here, consumption of kojic acid at levels normally found in food does not present a concern for safety. PMID- 11259182 TI - Risk assessment for glass wool exposure. PMID- 11259184 TI - Absence of genetic diversity reduction in the HIV-1 integrated proviral LTR sequence population during successful combination therapy. AB - We report the integrated proviral LTR sequence variation in four patients on highly active antiretroviral therapy (HAART). Integrated proviral fragments of LTR taken from four time points were PCR amplified from PBMCs and 10 to 12 individual clones were sequenced for each time point. Intrasample genetic distances and phylogenetic reconstruction of all LTR sequences demonstrated that 1-2 years of successful HAART did not significantly reduce the genetic repertoire of the integrated reservoir of HIV-1. PMID- 11259185 TI - Characterization of less pathogenic infectious molecular clones derived from acute-pathogenic SHIV-89.6p stock virus. AB - For a better understanding of the acute pathogenicity of SHIV-89.6P stock virus, which induces prominent CD4 cell loss within a month after inoculation in monkeys, we have constructed four infectious molecular clones (cl 18, cl 64, cl 69, and cl 71). Cl 64, cl 69, and cl 71, like the parental virus, showed a high in vitro replication ability and a pathogenic-like effect (CD4 downmodulation) in a monkey CD4(+) cell line, whereas cl 18 showed a lower replication ability and could not downmodulate CD4. Cl 64, which has characteristics similar to those of the parental virus in vitro, was inoculated into four rhesus monkeys. All monkeys showed a plasma viral load similar to that of the parental virus with a peak at 2 weeks after inoculation. However, the viral load gradually decreased and the virus failed to cause an AIDS-like disease in infected monkeys, but it induced a strong antiviral antibody response. These results demonstrate the polyclonal nature of the parental SHIV-89.6P virus stock and demonstrate that cl 64, aside from its high replicability, may differ qualitatively from the parental virus. PMID- 11259186 TI - Molluscum contagiosum virus inhibitors of apoptosis: The MC159 v-FLIP protein blocks Fas-induced activation of procaspases and degradation of the related MC160 protein. AB - Molluscum contagiosum virus contains two open reading frames, MC159 and MC160, that encode proteins with death effector domains resembling those of cellular regulators of apoptosis. Previous transfection analyses indicated that the MC159 protein binds to cellular FADD and inhibits Fas-induced cytolysis. For further studies, we inserted the MC159 or MC160 gene into the genome of vaccinia virus that had its own major anti-apoptosis gene deleted. The MC159-expressing virus blocked Fas-induced activation of caspase-3 and -8, degradation of PARP, and cleavage of DNA, whereas the parental vaccinia virus did not. The MC159 protein bound to procaspase-8, in addition to FADD, and was included in a complex with Fas upon receptor activation. Although the MC160 protein associated with FADD and procaspase-8 in co-immunoprecipitation studies, no protection against morphological or biochemical changes associated with Fas-induced apoptosis were discerned and the MC160 protein itself was degraded. Co-expression of MC159, as well as other caspase inhibitors, protected the MC160 protein from degradation, suggesting a functional relationship between the two viral proteins. PMID- 11259187 TI - Hepatitis C virus protein expression induces apoptosis in HepG2 cells. AB - The mechanisms of hepatocyte death and the events that lead to a high rate of chronic liver disease in patients infected with hepatitis C virus are not known. We established a HCV replication system in HepG2 cell culture and utilized this model to address the effect of HCV proteins on HepG2 cell growth and viability. After transfection of HepG2 cells with full-length RNA, a truncated RNA, or an antisense RNA, cell proliferation and cell viability were analyzed by thymidine uptake and the trypan blue exclusion method, respectively. Full-length RNA transfected HepG2 cells showed a decrease in cell proliferation and viability compared to cells transfected with HCV truncated RNA and antisense RNA control. A subset of cells expressing HCV proteins underwent apoptosis as documented by morphological studies, ultrastructural analysis, cell cycle analysis by flow cytometry, terminal transferase enzyme mediated end labeling of DNA, and DNA laddering. This study suggests that expression of HCV proteins can lead to cell death by apoptosis, which may be an important event in the pathogenesis of chronic hepatitis C virus infection in humans. PMID- 11259188 TI - Pathogenicity of a natural recombinant associated with ageratum yellow vein disease: implications for geminivirus evolution and disease aetiology. AB - Yellow vein disease of Ageratum conyzoides is caused by a viral DNA complex consisting of the genomic component (DNA A) of the monopartite begomovirus Ageratum yellow vein virus (AYVV, family: Geminiviridae) and a small satellite like DNA beta component. AYVV DNA A is unable to induce symptoms in this host alone but can systemically infect A. conyzoides in which it accumulates to low levels. Here, we demonstrate that the yellow vein phenotype can also be produced by co-inoculating A. conyzoides with AYVV DNA A and recDNA-Abeta17, a naturally occurring recombinant of approximately the same size as DNA beta that contains sequences from both DNA A and DNA beta. Symptoms induced by DNA A and recDNA Abeta17 in A. conyzoides and Nicotiana glutinosa are qualitatively similar to those associated with DNA A and DNA beta although milder. Recombination between DNA A and DNA beta to produce a chimera resembling recDNA-Abeta17 was observed after whitefly transmission of the disease in A. conyzoides. Hence, such recombination events are likely to occur frequently, implying that recombinants will normally be associated with this type of disease complex in the field. Possible implications of these findings for the evolution of begomoviruses and the aetiology of their diseases are discussed. PMID- 11259189 TI - Syncytium formation amplifies apoptotic signals: a new view on apoptosis in HIV infection in vitro. AB - Infection of CD4+ cells with HIV in vitro causes extensive cytopathology. The mechanism that underlies this process is unclear and conflicting data exist regarding whether cytotoxicity is due to necrosis or apoptosis. It was previously reported and is shown here that the coculture of HIV glycoprotein-expressing cells with CD4+ cells results in apoptosis within several hours. This study demonstrates that apoptosis did not occur in single cells and was mediated neither by CD4 nor by coreceptor signaling, indicating that apoptosis was not induced by intra- or intercellular glycoprotein-receptor interaction. Detection of apoptosis required cell-to-cell fusion and undetectable levels of apoptotic cell death were substantially amplified upon syncytium formation. Similar results were obtained with syncytium-forming cultures of measles virus glycoprotein expressing cells. These findings indicate that the apoptotic cell death observed in cultures of HIV and other syncytium-forming viruses is primarily due to amplification of background apoptosis in the wake of cell-to-cell fusion. PMID- 11259190 TI - Induction of herpes simplex virus gB-specific cytotoxic T lymphocytes in TAP1 deficient mice by genetic immunization but not HSV infection. AB - Loading of most endogenous peptides on major histocompatibility complex class I molecules is conditional on their transport into the endoplasmic reticulum (ER) by the peptide transporter TAP. We describe an HSV-2/1 cross-reactive cytotoxic T cell (CTL) epitope that is processed in a TAP1-independent manner in vivo following immunization of TAP1-/- mice with naked DNA or a recombinant vaccinia virus. These data indicated that TAP1-independent processing of endogenous proteins is sufficient to prime CTLs in vivo. TAP1-independent processing of this epitope was not due to ER targeting by signal sequences and exogenous loading of MHC-I molecules and was not influenced by the amino acids flanking this epitope. In contrast, TAP1-/- mice infected with HSV-2 or HSV-2 mutants did not mount a CTL response against this epitope. PMID- 11259191 TI - Neurological diseases and viral dynamics in the brains of neonatally borna disease virus-infected gerbils. AB - Borna disease virus (BDV) is a noncytolytic, neurotropic RNA virus that causes a chronic neurological disease in a wide variety of animal species. To develop a better understanding of the correlation between neurological disorders caused by BDV infection and virus distribution in the brain, we investigated viral dynamics in the central nervous system (CNS) of neonatally BDV-infected gerbils during the late stage of infection. Despite the severe symptoms and aggressive proliferation of BDV in the infected gerbils, no apparent neuroanatomical abnormalities or neuronal cell loss was observed in the infected gerbil brain. Furthermore, no or only minimal infiltration was observed in the infected gerbil brain. By in situ hybridization and real-time PCR analyses, we demonstrated that the predominant area of expression of BDV mRNA, as well as the protein, was shifted in the brain in association with progression of disease. In nondiseased gerbils, the virus replication was predominantly detected in the cerebral cortex and hippocampus of the CNS. On the other hand, diseased animals showed a high level of expression in the lower brain stem and cerebellum, especially in Purkinje cell neurons. These observations suggested that significant replication of the virus in specific areas of the CNS is critical for development of the neurological disorders in BDV infected neonatal gerbils. PMID- 11259192 TI - An inducible packaging cell system for safe, efficient lentiviral vector production in the absence of HIV-1 accessory proteins. AB - Lentiviral vectors based on human immunodeficiency virus type 1 (HIV-1) possess the ability to deliver exogenous genes to both dividing and nondividing cells and to subsequently establish a stable provirus in these target cells, which can allow long-term expression of the transferred gene. Herein we describe a stable packaging cell line that is devoid of HIV-1 tat, vif, vpr, vpu, and nef. In order to avoid any risk of cytotoxicity associated with constitutive expression of HIV 1 protease or the VSV-G envelope protein, transcription of the packaging and envelope constructs was tightly controlled by employing the ecdysone-inducible system. Using this cell line, we have been able to consistently generate concentrated pseudotyped vector virus stocks with titers in the range of 10(8) IU/ml, which can efficiently transduce actively dividing and growth-arrested cells in vitro. This novel packaging cell line for lentiviral vectors facilitates the production of high-titer virus stocks in the absence of replication-competent virus and provides us with an important tool for use in future gene transfer studies. PMID- 11259193 TI - The 3' end of hepatitis E virus (HEV) genome binds specifically to the viral RNA dependent RNA polymerase (RdRp). AB - Hepatitis E virus (HEV) is the major cause of acute epidemic and sporadic hepatitis in the developing world. It is a positive-strand RNA virus with a genome length of about 7.2 kb. The replication mechanism of this virus is virtually unexplored. Identification of the regulatory elements involved in initiation of replication may help in designing specific inhibitors for therapy. In the positive-stranded RNA viruses the initiation of replication requires interaction of the 3' end of genome with its RNA-dependent RNA polymerase (RdRp) and possibly host-derived cofactors for synthesis of the minus-strand replicative intermediate. Secondary structure prediction of the conserved 3' end of the infectious HEV genome was carried out to identify possible stem-loop structures necessary for RNA-protein interaction and the model was confirmed by structure probing experiments. Electrophoretic mobility-shift assays showed specific binding of purified and refolded recombinant HEV RdRp protein to the 3' end of its RNA genome containing the poly(A) stretch. Mutations at the 3' end, in which the stem-loop structures were partially or completely destroyed or recreated revealed that the two stem-loop structures SL1 and SL2 at the 3' end and the poly(A) stretch are necessary for this binding. The interacting nucleotides in such an interaction were further identified by generating footprints of the complex by Pb(II)-induced hydrolysis. This specific binding of viral RdRp to the 3' end of HEV RNA directs the synthesis of complementary-strand RNA and thus such a binding domain might assume the role of a possible cis-acting element as a potential site for the initiation of replication. PMID- 11259194 TI - Binding properties, cell delivery, and gene transfer of adenoviral penton base displaying bacteriophage. AB - The penton base of adenovirus mediates viral attachment to integrin receptors and particle internalisation, properties that can be exploited to reengineer prokaryotic viruses for the infection of mammalian cells. We report that filamentous phage displaying either the full-length penton base gene or a central region of 107 amino acids on their surface were able to bind, internalise, and transduce mammalian cells expressing integrin receptors. Both phage bound alphavbeta3, alphavbeta5, alpha3beta1, and alpha5beta1 integrin subtypes. Cell binding was shown by electron microscopy; internalisation was investigated by immunofluorescence and confirmed by micropanning. As it has been described for adenovirus, pharmacologic disruption of phosphoinositide-30H kinase, but not of myosin light-chain kinase, inhibited phage internalisation. Recombinant phage encoding an eukaryotic expression cassette was able to mediate gene expression in mammalian cells. Taken together, these data open insights for the exploit of recombinant phage for integrin-targeted gene delivery. PMID- 11259195 TI - Phenotypic and functional characteristics of FIV infection in the bone marrow stroma. AB - Human (HIV) and feline (FIV) immunodeficiency virus has been reported to infect bone marrow (BM) and stroma, followed by a loss in normal hematopoiesis. However, the magnitude and nature of HIV and FIV pathogenesis of the BM/stromal network are still unclear. In the current studies, pathogenesis of stromal cells was evaluated using the FIV model. Fourteen specific-pathogen-free cats inoculated with the four different strains (FIV(UK8), FIV(Bang), FIV(Shi), or FIV(Pet)) were monitored for FIV infection in the peripheral blood mononuclear cells (PBMC), BM cells, and stromal cells. All inoculated cats became positive for FIV in the PBMC by 7 weeks p.i. and 13 of 14 cats had FIV in the BM cells by 7-13 weeks p.i. FIV was detected in macrophages and stromal fibroblasts from FIV(UK8)-, FIV(Bang)-, and FIV(Shi)-infected cats but not from FIV(Pet)-infected cats and only transiently in cells from FIV(Shi)-infected cats. The ability of the supernatants from FIV-infected stromal cells to sustain the growth of uninfected BM cells was decreased 35-46% when compared to the supernatants from uninfected stromal cells. These results suggest that the FIV infection of the stroma alters normal hematopoietic function(s) and that the infected stromal cells can also serve as a reservoir for FIV infection. PMID- 11259196 TI - A simian human immunodeficiency virus with a nonfunctional Vpu (deltavpuSHIV(KU 1bMC33)) isolated from a macaque with neuroAIDS has selected for mutations in env and nef that contributed to its pathogenic phenotype. AB - Previous studies have shown that passage of nonpathogenic SHIV-4 through a series of macaques results in the selection of variants of the virus that are capable of causing rapid subtotal loss of CD4(+) T cells and AIDS within 6-8 months following inoculation into pig-tailed macaques. Using a pathogenic variant of SHIV-4 known as SHIV(KU-1bMC33), we reported that a mutant of this virus with the majority of the vpu deleted was still capable of causing profound CD4(+) T cell loss and neuroAIDS in pig-tailed macaques (McCormick-Davis et al., 2000, Virology 272, 112-116). In this study, we have analyzed the tissue-specific changes in the env and nef in one macaque that developed neuroAIDS (macaque 50 O) and in three macaques that developed only a moderate or no significant loss of CD4(+) T cells and no neurological disease (macaques 50 Y, 20220, 20228) following inoculation with DeltavpuSHIV(KU-1bMC33). Sequence analysis of the gp120 region of env isolated from lymphoid tissues (lymph node and spleen) of macaques 50 Y, 20220, and 20228 revealed no consensus amino acid substitutions. In contrast, analysis of the gp120 sequences isolated from lymphoid and CNS tissues (parietal cortex, basal ganglia, and pons) of macaque 50 O revealed numerous amino acid substitutions. The significance of the amino acid substitutions in gp120 was supported by neutralization assays which showed that the virus isolated from the lymph node of macaque 50 O was neutralization resistant compared to the parental SHIV(KU-1bMC33). Analysis of changes in the nef gene from macaque 50 O revealed in-frame deletions in Nef that ranged from 4 to 13 amino acids in length, whereas the nef genes isolated from the other three macaques revealed no deletions or consensus amino acid substitutions. Inoculation of the virus isolated from the lymph node of the macaque which developed neuroAIDS, SHIV(50OLNV), into four pig tailed macaques resulted in a severe loss of the circulating CD4(+) T cells within 2 weeks postinoculation, which was maintained for up to 20 weeks postinoculation, confirming that this virus had indeed become more pathogenic in pig-tailed macaques. Taken together, these observations suggest that DeltavpuSHIV(KU-1bMC33) has a low pathogenic phenotype in macaques but that individual pig-tailed macaques can select for additional mutations within the Env and Nef which can compensate for the lack of an intact Vpu and ultimately increase its pathogenicity. PMID- 11259197 TI - The effect of human bcl-2 and bcl-X genes on dengue virus-induced apoptosis in cultured cells. AB - Infection of dengue viruses (DENs) can cause human dengue fever, hemorrhagic fever, or shock syndrome. Although DEN-induced apoptosis has been implicated in pathogenesis of the DEN-related diseases, the underlying mechanism remains largely unexplored. In this study, we investigated the effect of ectopic expression of human bcl-2 and bcl-X genes on DEN-induced apoptosis in cultured cells. We employed a human isolate of DEN serotype 2 (DEN-2), PL046, which not only caused cell-cycle arrest in the G1 phase but also induced apoptosis in infected baby hamster kidney (BHK-21) cells, murine neuroblastoma N18 cells, and human neuronal NT-2 cells. Our results reveal that overexpression of bcl-2 in fibroblast-like BHK-21 cells, although not inhibiting virus yields, delayed the process of DEN-induced apoptosis, thereby permitting surviving cells to become persistently infected. In contrast, stable bcl-2 expression in neuronal N18 cells failed to block DEN-induced apoptosis. On the other hand, Bcl-X(L), expressed predominantly in the nervous system, appeared to delay DEN's killing effect in neuronal N18 cells but not in fibroblast-like BHK-21 cells. In addition, inducible expression bcl-X(s), despite its proapoptotic property in other reported system, was found to merely accelerate cell death in DEN-infected N18 but not in infected BHK-21 cells. Thus, through studying the effect of human bcl 2-related genes, our results suggest that DEN infection may trigger target cells to undergo morphologically similar but biochemically distinct apoptotic pathways in a cell-specific manner. PMID- 11259198 TI - Generation of replication-defective helper-free vectors based on simian immunodeficiency virus. AB - A systematic study on generating simian immunodeficiency virus (SIV)-based vectors was carried out. The goal was to generate helper-free, replication defective SIVmac-based vectors at high titers. The general approach was to cotransfect into human 293T cells a plasmid carrying the vector construct along with two helper plasmids that together expressed the SIVmac virion proteins. Initial vectors carried the bacterial beta-galactosidase gene (beta-gal). These vectors had a technical difficulty: "pseudotransduction" of beta-gal protein produced during the 293T cell transfections. As a result, infection of cultures with these vector stocks also resulted in passive transfer into, and X-gal staining of, cells that had not actually been infected by the vector. A second generation of vectors expressing the enhanced jellyfish green fluorescence protein (EGFP) was not subject to this artifact. A systematic study of the SIVmac based EGFP vectors was carried out. Helper-free vector stocks were obtained when helper plasmids lacking the SIVmac packaging signals were used. By employing envelope helper plasmids derived from different SIVmac isolates, it was possible to generate SIVmac-based vectors pseudotyped with envelope proteins of different cell tropism. Optimization of vector and helper plasmid structures, transfection conditions, and infection procedures ultimately yielded vector titers in excess of 10(6)/ml. PMID- 11259199 TI - Monoclonal antibodies against different epitopes of nonstructural protein sigmaNS of avian reovirus S1133. AB - Ten monoclonal antibodies (MAbs) were prepared against the nonstructural protein sigmaNS of avian reovirus S1133. Eight of them were selected for two-way competitive binding assay after coupling with horseradish peroxidase. The results allowed the definition of three epitopes, designated A, B, and C. Blocking assay of poly(A)-Sepharose binding activity of sigmaNS with MAbs indicated that MAb recognizing epitope B was able to block poly(A) oligomer binding, suggesting that epitope B is involved in ssRNA binding of sigmaNS. An immuno-dot binding assay was used to analyze the effect of denaturation on antibody recognition of the epitopes. All MAbs bound to protein sigmaNS in its native form. After denaturation by boiling in SDS and 2-mercaptoethanol, the binding of MAbs recognizing epitopes B and C was not affected. The reactivity of MAbs recognizing epitope A was fully abolished by denaturation. These results suggest that the binding of MAbs directed against epitope A is conformation-dependent; however, the recognition by MAbs of epitopes B and C is not conformation-dependent. In addition, the results from the cross-reactivity of MAbs to heterologous avian reovirus strains suggest that the three epitopes are highly conserved among these virus strains. PMID- 11259200 TI - Interaction of human immunodeficiency virus type 1 Vpr with the HHR23A DNA repair protein does not correlate with multiple biological functions of Vpr. AB - The virion-associated Vpr protein of human immunodeficiency virus type 1 (HIV-1) alters cell cycle progression from the G2 phase, influences the virus in vivo mutation rate, and participates in the nuclear translocation of viral DNA. While many Vpr-interacting proteins have been identified, the functional relevance of these interactions remains to be thoroughly documented. We have explored the contribution of the interaction of HIV-1 Vpr with HHR23A, a cellular protein implicated in DNA repair, to the known phenotypes of Vpr. The association of Vpr with HHR23A required the core region of Vpr, which encompasses the two alpha helical structures of the protein. No binding of HHR23A was detected with the Vpr and Vpx proteins of other primate lentiviruses. HIV-1 Vpr variants containing single amino acid substitutions in each alpha-helix and deficient for binding to HHR23A were isolated. The functional characterization of these Vpr variants indicated that binding to HHR23A did not correlate with the ability of Vpr to induce cell cycle arrest, even though it was previously proposed that HHR23A is a mediator of the Vpr-induced G2 arrest. Also, the Vpr-HHR23A interaction did not influence the HIV-1 in vivo mutation rate. Finally, Vpr and HHR23A are both localized in the nucleus, but no correlation was observed between the nuclear targeting of Vpr and the interaction with HHR23A. Further analysis is needed to determine the functional role(s) of the Vpr-HHR23A association during the HIV-1 life cycle. PMID- 11259201 TI - Packaging cell line DNA contamination of vector supernatants: implication for laboratory and clinical research. AB - Investigators conducting retroviral gene therapy trials are required to monitor for the presence of replication-competent retrovirus (RCR). The required testing utilizes a combination of biologic assays and molecular tests using PCR. In the course of a human clinical gene therapy trial, we detected 4070A viral envelope sequences in CD34(+) peripheral blood stem cells 2 days after transduction using a PCR-based assay, suggesting the presence of RCR. The supernatant and producer cells used for vector generation had been negative in extensive screening using the extended S(+)/L(-) assay. The presence of a replication-competent virus was subsequently excluded by a combination of biologic and PCR analyses. The source of the 4070A viral envelope sequences was determined to be packaging cell line DNA in the vector supernatant. The analysis of a variety of vector supernatants by quantitative real-time PCR revealed 4070A envelope DNA sequences from the packaging cell line in concentrations equivalent to approximately 50-500 focus forming units per milliliter of wild-type 4070A virus. When PCR was performed after reverse transcriptase treatment of supernatant (i.e., assessing both RNA and DNA content), 4070A envelope sequence concentrations ranged from 10(2) to 3.5 x 10(3) focus-forming units per milliliter of wild-type 4070A virus. Our data indicate that PCR should not be used to analyze transduced cells for RCR within the first 2 weeks of vector exposure. Furthermore, investigators using PCR to analyze transduction efficiency shortly after vector exposure may experience false-positive findings. PMID- 11259202 TI - Marek's disease virus down-regulates surface expression of MHC (B Complex) Class I (BF) glycoproteins during active but not latent infection of chicken cells. AB - Infection of chicken cells with three Marek's disease virus (MDV) serotypes interferes with expression of the major histocompatibility complex (MHC or B complex) class I (BF) glycoproteins. BF surface expression is blocked after infection of OU2 cells with MDV serotypes 1, 2, and 3. MDV-induced T-cell tumors suffer a nearly complete loss of cell surface BF upon virus reactivation with 5 bromo-2'-deoxyuridine (BUdR). The recombinant virus (RB1BUS2gfpDelta) transforming the MDCC-UA04 cell line expresses green fluorescent protein (GFP) during the immediate early phase of viral gene expression. Of the UA04 cells induced to express the immediate early GFP, approximately 60% have reduced levels of BF expression. All of the reactivated UA04 and MSB1 tumor cells expressing the major early viral protein pp38 display reduced levels of BF. Thus, BF down regulation begins in the immediate early phase and is complete by the early phase of viral gene expression. The intracellular pool of BF is not appreciably affected, indicating that the likely mechanism is a block in BF transport and not the result of transcriptional or translational regulation. PMID- 11259203 TI - The basic loop of the RNase H domain of MLV RT is important both for RNase H and for polymerase activity. AB - Escherichia coli RNase H has a basic extension that is involved in binding nucleic acid substrates. This basic extension is present in the RNase H of Moloney murine leukemia virus reverse transcriptase (MLV RT), but has been deleted from the RNase H of HIV-1 RT. Previous work showed that removing the basic loop from MLV RT (the mutant is called DeltaC) blocked viral replication; however, DeltaC MLV RT retained RNase H activity in an in situ gel assay. We prepared recombinant DeltaC MLV RT and compared its activity to wild-type MLV RT. The DeltaC mutant is impaired in both polymerase and RNase H activity; the pattern of defects suggests that the basic loop is involved in the binding of MLV RT to a heteropolymeric template-primer. PMID- 11259204 TI - Genetic association studies of alcoholism--problems with the candidate gene approach. AB - In recent years, progress has been made in the identification of causative factors in most single gene disorders and those with genes of major effect. In comparison, no genes contributing to a complex disorder have been unambiguously identified. A number of reasons for this have been previously presented in theoretical papers. Alcoholism is such a complex illness and genetic studies into its underlying genetic causes have suffered from lack of power due to small subject numbers, poor selection of control subjects, and over-emphasis on markers with low prior probability of involvement. PMID- 11259205 TI - A comparison of rating scales for the alcohol-withdrawal syndrome. AB - This paper reviews the literature on the use of rating scales within the treatment of the alcohol-withdrawal syndrome. A computer-assisted literature search identified trials of therapy for and rating scales used in alcohol withdrawal states. Eighteen rating scales were identified. There is a wide variation in symptom items included in these scales. Scales also vary in their length and ease of application. We conclude that it is important to use validated and reliable assessment scales in research if proper comparisons of treatments for the alcohol-withdrawal syndrome are to be made. PMID- 11259206 TI - Cochrane Drugs and Alcohol Group: the development of systematic reviews of treatment outcome. AB - - The aim of the Cochrane Collaboration is to promote review processes which address all aspects of health care and which can be viewed by clinicians to guide their day-to-day clinical practice. Recently, a Cochrane review group on drugs and alcohol has been developed. The Cochrane Review Group Editorial base is in Rome, Italy. There is an international editorial board with editors in the UK, Italy, France, Australia and the USA. So far, the group has published five reviews addressing treatment for opioid, cocaine and alcohol dependence. Additional reviews and protocols are in progress. A growing number of titles are registered with the group. Interested readers and potential reviewers and/or referees can contact the Cochrane Drugs and Alcohol Group Coordinator in Rome at the e-mail address: dacochrane@asplazio.it PMID- 11259207 TI - Alcohol use inventory: screening and assessment of first-time driving-while impaired offenders. I. Reliability and profiles. AB - This study evaluated the use of the Alcohol Use Inventory (AUI) in a drink driving offender population court-mandated to attend a screening programme. We compared offenders' scale scores, reliability statistics and profiles to those from two clinical populations on which the AUI was normed. Among offenders, males and females had similar levels of involvement with alcohol, and Native Americans had higher scale scores than other ethnic groups. Comparisons with the normative population revealed lower mean scale scores and lower reliability scores among offenders. Differences between the offender and normative populations were most pronounced for the primary scales. We also found inconsistencies in offenders' responses to certain questions. To address this, we recommend that, when using the AUI for screening offenders: (1) screeners place more emphasis on second- and third-order scales than primary scales; (2) lower cut-off points be used for identifying problem drinkers; (3) counsellors conduct in-person interviews with clients to develop rapport and encourage self-disclosure. PMID- 11259208 TI - Alcohol use inventory: screening and assessment of first-time driving-while impaired offenders. II. Typology and predictive validity. AB - This study evaluated the use of Alcohol Use Inventory (AUI) for driving-while impaired (DWI) screening, by determining whether DWI offenders (n = 1644), grouped according to their reported alcohol involvement on the AUI, would have different rates of recidivism in a 5-year follow-up. Cluster analysis using the six second-order scales produced six groups (clusters 1-6) described as the Low Profile (50%), Alcohol-Preoccupation (14%), Enhanced (22%), Enhanced-Disrupt (9%), Anxious-Disrupt (3%), and High-Profile (1%) types. They were characterized by different sociodemographic profiles. Members of cluster 4 were associated with the highest DWI recidivism rate (40%), committing one or more further DWI, and clusters 5 and 6 were associated with the highest rate of committing two or more DWIs. Rates of subsequent traffic convictions and crashes were, however, not statistically different among the clusters. Predictors of DWI recidivism included male gender, young age, less-educated, high blood-alcohol concentration at arrest, and clusters of 3 and 4. Different typologies indicated that the needs for treatment might be different. Evaluators should keep in mind the strength of AUI, use risk factors identified in the study, and take measures of test-taking defensiveness to enhance overall predictive validity. PMID- 11259209 TI - Gin Lane: did Hogarth know about fetal alcohol syndrome? AB - Medical historians have searched for evidence that the characteristics of fetal alcohol syndrome (FAS) were recognized long before its modern description in 1973. This search has often focused on the 'gin epidemic' in 18th century London, and especially William Hogarth's Gin Lane, which some authors allege reflects an awareness of the facial characteristics of the syndrome. While the 'gin epidemic' undoubtedly resulted in the increased birth of weak and sickly children, claims about Hogarth's awareness of the stigmata of the FAS are unfounded. The birth of weak and sickly children, and the high infant mortality rates associated with this period, long preceded the 'gin epidemic' and were primarily due to disease, starvation, exposure, and deliberate infanticide. PMID- 11259210 TI - Exploring attitude and belief correlates of adhering to the new guidelines for low-risk single-occasion drinking: an application of the theory of planned behaviour. AB - The present study explores the correlates of adhering to the recent low-risk single-occasion drinking (LRSOD) guidelines. This was achieved by exploring key beliefs and attitudes underlying adherence to these guidelines within the framework of the theory of planned behaviour (TPB). Female students (n = 173) provided information about their LRSOD and beliefs and attitudes pertaining to LRSOD. Analyses of the resultant data showed the TPB to be significantly predictive of LRSOD, accounting for 27% of the variance, with normative beliefs, behavioural beliefs, and attitude emerging as significant predictors in the regression analysis. The implications of the study findings are discussed in terms of the current utility of the LRSOD limits for reducing alcohol-related harm. PMID- 11259211 TI - Opinions on alcohol-related issues among professionals in primary, occupational, and specialized health care. AB - The objective of this study was to analyse differences in health care personnel's knowledge, skills, and attitudes in relation to alcohol-related matters by a postal questionnaire between primary, occupational, and specialized health care. Heavy drinking was considered to be common among patients at all health care levels, and particularly in specialized health care. However, early recognition and treatment of heavy drinkers was considered more appropriate in primary and occupational health care, than in specialized health care. Alcohol consumption was found to be an easy subject to discuss at all health care levels. In addition, 90% (165/183) of the respondents thought that patients had a positive or neutral attitude towards questions on their alcohol consumption. Of the respondents, 32% (58/182) considered discussing alcohol-related matters unacceptable and 81% (121/149) believed that they could not influence patients' drinking using brief intervention; there was no significant difference between different settings. Additionally, motivational skills of doctors and nurses were found to be poor at all health care levels. Our study shows that, although discussing alcohol consumption is easy, better motivational skills and more positive attitudes are needed in primary, occupational, and specialized health care. Professionals need further education at all health care levels, but particularly in specialized health care. PMID- 11259212 TI - Measuring the facial phenotype of individuals with prenatal alcohol exposure: correlations with brain dysfunction. AB - The purpose of this report is to demonstrate how to measure the magnitude of expression of the fetal alcohol syndrome (FAS) facial phenotype using the new 4 Digit Diagnostic Code and the previously developed D-score and to demonstrate how these two measures of the FAS facial phenotype correlate with brain function and structure; correlations that fail to be identified by the older gestalt method of facial measurement. The D-score and the facial component of the 4-Digit Diagnostic Code quantitatively measure the magnitude of expression of the FAS facial phenotype using three facial features (palpebral fissure length, philtrum smoothness, and upper lip thinness). These facial measurement systems were developed by the Washington State FAS Diagnostic and Prevention Network (FAS DPN) of clinics and are used to screen and diagnose the facial component of FAS for all patients evaluated in the network of clinics (1500 to date). The 4-Digit Diagnostic Code is a comprehensive diagnostic system developed by the FAS DPN in 1997 to diagnose the full spectrum of outcomes among patients with prenatal alcohol exposure. The four digits reflect the magnitude of expression of the four key diagnostic features of FAS in the following order: (1) growth deficiency; (2) the FAS facial phenotype; (3) brain dysfunction; (4) gestational alcohol exposure. The 4-Digit Diagnostic Code was developed to overcome the subjective, highly variable gestalt method of diagnosis that has been used as the standard to date, worldwide. Prior to the development of the 4-Digit Diagnostic Code, the first 445 patients evaluated in the FAS DPN were diagnosed using the gestalt method. For research purposes, their gestalt diagnoses were transformed into 4 Digit Diagnostic Codes, presenting a unique opportunity to directly compare the two diagnostic methods. When the facial phenotype was measured using the 4-Digit Diagnostic Code or D-score, the magnitude of expression of the FAS facial phenotype was significantly correlated with structural, neurologic, and functional measures of brain damage, and the phenotype of those receiving a 4 Digit Diagnosis of FAS showed little variability. When the gestalt method of diagnosis was used, the magnitude of expression of the FAS facial phenotype did not correlate with structural, neurologic and functional measures of brain damage, and the facial phenotype of those receiving a gestalt diagnosis of FAS was highly variable. The 4-Digit Diagnostic Code and D-score thus provide more precise and accurate measures of the FAS facial phenotype and reveal important correlations with brain structure and function, suggesting that intermediate expressions of the FAS facial phenotype may serve as important risk factors for brain damage caused by prenatal alcohol exposure. PMID- 11259213 TI - Health care professionals referred for treatment of alcohol and drug problems. AB - This study reports on 62 health care professionals referred to a specialist drug and alcohol treatment service. Most patients used more than one type of substance. Health problems were common, but were seldom reasons for referral. Self-referral was infrequent. Referral was often subsequent to intoxication at work or persistent absenteeism. Just over half of admissions completed treatment. Multiple drug use was a poor prognostic indicator with fewer multiple drug users engaging with, or completing, treatment. PMID- 11259214 TI - Sweet liking and family history of alcoholism in hospitalized alcoholic and non alcoholic patients. AB - The present study was designed to test the hypothesis that preference for stronger sweet solutions may be associated with the genetic risk for alcoholism. Thirty-two male patients with alcohol dependence admitted for alcoholism in patient treatment and 25 non-alcoholic control subjects were used in the study. Hedonic response to sweets was evaluated using the sweet preference test. Family history of alcoholism was evaluated using a Russian version of the Michigan Alcoholism Screening Test modified for the assessment of the alcohol-related behaviour of the subject's biological father. Similar to our previous findings, alcoholics were far more likely to prefer the highest offered sucrose concentration (0.83 M), compared to non-alcoholic controls. Such preference was determined by two factors: positive family history of alcoholism and alcoholic status. Statistically, these factors contributed to the likelihood of preferring sweet solutions independently. Therefore, the effects of these factors may enhance each other. These findings support the hypothesis that preference for a stronger sweet solution is associated with a paternal history of alcohol dependence and may reflect a genetic predisposition to alcoholism. PMID- 11259215 TI - Visual performance and recovery in recently detoxified alcoholics. AB - In order to assess the impact of chronic alcohol misuse on basic visual functions, we investigated motion perception, visual short-term memory, and visual divided attention in recently detoxified patients and matched controls by means of visual psychophysical tasks. Subjects were tested twice within the first 3 weeks of detoxification in order to assess the potential recovery of visual performance. Patients demonstrated significant impairments in visual perception of coherent motion for slow, but not faster, speeds, and in speed discrimination as assessed by random dot kinematograms. Visual short-term memory tested with a delayed vernier discrimination task, on the other hand, was not significantly affected in patients. When processing hierarchical letters, a divided attention task, detoxified patients showed neither impairments in overall attentional capacity nor attentional allocation, but slightly enhanced interference of global information on local target processing. The results of the visual divided attention task contradict the predictions of the 'right hemisphere' hypothesis of alcoholism: global target information - mediated by the right hemisphere - was not only accessible to detoxified patients, but seemed to exert an even greater influence on local processing during early detoxification, than in matched controls. Limited recovery within the first 3 weeks was seen only in visual speed discrimination. Recently detoxified patients revealed deficits similar to intoxicated social drinkers in identical tests of visual perception of motion, but not visual short-term memory. PMID- 11259217 TI - The value of oral thiamine. PMID- 11259218 TI - Outpatient services for children. PMID- 11259219 TI - Stamps in paediatrics: Poliomyelitis. PMID- 11259220 TI - Investigation and treatment of facial paralysis. PMID- 11259221 TI - The "How!" sign--a central palmar blister induced by overplaying on a Nintendo console. PMID- 11259222 TI - New pneumococcal vaccines for children. PMID- 11259223 TI - Nutritional support at home and in the community. AB - Technical developments in feeding, together with the growth of support structures in the community has lead to a steady increase in the number of children receiving home enteral tube feeding and home parenteral nutrition. In many cases the adverse nutritional consequences of disease can be ameliorated or prevented, and long term parenteral nutrition represents a life saving intervention. Careful follow up of children receiving home nutritional therapy is necessary to establish the ratio of risks to benefits. A considerable burden is sometimes placed on family or other carers who therefore require adequate training and ongoing support. The respective responsibilities of different agencies relating to funding and support tasks require more clear definition. PMID- 11259224 TI - Social paediatrics and child public health--a European perspective. PMID- 11259226 TI - Mental health and foster carer training. AB - AIMS: To evaluate the impact of training foster carers on children's emotional and behavioural functioning. METHODS: In a randomised controlled trial in 17 Scottish local council areas, with immediate and nine month follow up, 182 children and their foster families were randomly allocated to either standard services alone or standard services plus extra training for foster carers on communication and attachment. Main outcome measures were child psychopathology, attachment disorder, self esteem, and cost of foster care. RESULTS: Over 60% of children had measurable psychopathology at baseline. The training was perceived as beneficial by participants. Scores for parent reported psychopathology and attachment disorders decreased by around 5%, self esteem increased by 2%, and costs by 22% in the intervention group. Results were non-significant. CONCLUSIONS: Despite being well received by foster carers, the training was not sufficient to make a useful impact on the high level of psychopathology. This group may warrant more intensive interventions. PMID- 11259227 TI - Social deprivation and the causes of stillbirth and infant mortality. AB - AIMS: To investigate the relation between social deprivation and causes of stillbirth and infant mortality. METHODS: Stillbirths and infant deaths in 6347 enumeration districts in Wales were linked with the Townsend score of social deprivation. In 1993-98 there were 211 072 live births, 1147 stillbirths, and 1223 infant deaths. Poisson regression analysis was used to estimate the magnitude of effect for associations between the Townsend score and categories of death by age and the causes of death. The relative risk of death between most and least deprived enumeration districts was derived. RESULTS: Relative risk of combined stillbirth and infant death was 1.53 (95% CI 1.35 to 1.74) in the most deprived compared with the least deprived enumeration districts. The early neonatal mortality rate was not significantly associated with deprivation. Sudden infant death syndrome showed a 307% (95% CI 197% to 456%) increase in mortality across the range of deprivation. Deaths caused by specific conditions and infection were also associated with deprivation, but there was no evidence of a significant association with deaths caused by placental abruption, intrapartum asphyxia, and prematurity. CONCLUSIONS: Collaborative public health action at national and local level to target resources in deprived communities and reduce these inequalities in child health is required. Early neonatal mortality rates and deaths from intrapartum asphyxia and prematurity are not significantly associated with deprivation and may be more appropriate quality of clinical care indicators than stillbirth, perinatal, and neonatal mortality rates. PMID- 11259229 TI - Work, family socioeconomic status, and growth among working boys in Jordan. AB - AIMS: To describe the work, family socioeconomic characteristics, and growth of a representative sample of working children in Jordan. METHODS: In a cross sectional survey of growth and health, 135 working children (aged 10-16 years) were studied in the areas of Irbid, Jarash, and North Jordan Valley. The children and their parents were interviewed and data collected on length of working week, income earned by the child, duration of work in years, age of starting work, type of work, child's smoking status, and family socioeconomic status. RESULTS: The mean age of the children was 13.3 years; 14.8% had started work before the age of 10 and 12.6% had been working for more than four years. Mean income was 34 Jordanian Dinars but 6.7% were unwaged; 34% were working more than 60 hours per week, and 85.9% more than 40 hours. Monthly income and working hours were positively correlated with the age of the child. There was no correlation between age and smoking status; 37.8% smoked more than five cigarettes per day. Mean height and weight z scores were -0.365 and -0.081 of the UK standard respectively. Packed cell volume was within the anaemic range in 34.1% of children. CONCLUSIONS: In Jordan many children start work at an early age and work long hours for little or no income. Stunting and anaemia are common and many are established smokers. Relevance of these findings for social policy and health care of working children in Jordan and elsewhere is discussed. PMID- 11259230 TI - Developing sustainable international partnerships in child health and paediatric care. PMID- 11259231 TI - Evaluation of urinary tract calculi in children. PMID- 11259232 TI - An audit of RCP guidelines on DMSA scanning after urinary tract infection. AB - AIM: To assess the outcome of imaging investigations carried out in children with urinary tract infection (UTI), to compare the investigations with national guidelines, and to assess the impact on management. METHODS: Retrospective review of inpatients and outpatients, aged 0-12 years, referred to the University Hospital of Wales Healthcare Trust between February 1997 and January 1998 with UTI. All children without bacterial evidence of UTI and children previously investigated for antenatal urological anomalies, major congenital anomalies, or UTI were excluded. RESULTS: A total of 164 children (51 boys, 113 girls) were included. Thirteen of 56 infants (23%) and 82/108 older children (76%) were diagnosed at home over one year. The prevalence of dilatation on ultrasound was 8%, renal scarring on dimercaptosuccinic acid (DMSA) scan was 11%, and vesicoureteric reflux (VUR) was 34% when investigations were carried out following guidelines published by the Royal College of Physicians. In children aged 1-6 years, the prevalence of scarring was 1/54 (2%) in those treated at home and 6/18 (33%) in inpatients. CONCLUSION: The low yield of positive results and lack of evidence of impact on management indicate that DMSA scanning, with all the implications of isotope exposure, intravenous injection, staff time, psychological trauma, and expense, could be omitted in children over 1 year with first simple UTI not sufficiently ill to be admitted to hospital. The low rate of detection of UTI in primary care in infants may represent under diagnosis. PMID- 11259233 TI - Faecal candida and diarrhoea. AB - BACKGROUND: Candida species are frequently isolated from stools of children with diarrhoea but are not proven enteropathogens. It is hypothesised that faecal candida causes diarrhoea. AIMS: To determine the prevalence of faecal candida in childhood diarrhoea and the relation between faecal yeasts and diarrhoea. METHODS: Comparison of clinical and laboratory data, including quantitative stool culture for yeasts from 107 children hospitalised with diarrhoea and 67 age matched controls without diarrhoea. RESULTS: Yeast species, predominantly candida, were identified in the stools of 43 children (39%) with diarrhoea and 26 (36%) without diarrhoea. The concentration of candida was positively associated with recent antibiotic use (p = 0.03) and with the presence of another enteric pathogen (p < 0.005), but not with patient age, nutritional status, or duration of diarrhoea. CONCLUSION: Candida species do not cause childhood diarrhoea in well nourished children. PMID- 11259234 TI - Procalcitonin in children admitted to hospital with community acquired pneumonia. AB - AIMS: To assess the sensitivity, specificity, and predictive value of procalcitonin (PCT) in differentiating bacterial and viral causes of pneumonia. METHODS: A total of 72 children with community acquired pneumonia were studied. Ten had positive blood culture for Streptococcus pneumoniae and 15 had bacterial pneumonia according to sputum analysis (S pneumoniae in 15, Haemophilus influenzae b in one). Ten patients had Mycoplasma pneumoniae infection and 37 were infected with viruses, eight of whom had viral infection plus bacterial coinfection. PCT concentration was compared to C reactive protein (CRP) concentration and leucocyte count, and, if samples were available, interleukin 6 (IL-6) concentration. RESULTS: PCT concentration was greater than 2 microg/l in all 10 patients with blood culture positive for S pneumoniae; in eight of these, CRP concentration was above 60 mg/l. PCT concentration was greater than 1 microg/l in 86% of patients with bacterial infection (including Mycoplasma and bacterial superinfection of viral pneumonia). A CRP concentration of 20 mg/l had a similar sensitivity but a much lower specificity than PCT (40% v 86%) for discriminating between bacterial and viral causes of pneumonia. PCT concentration was significantly higher in cases of bacterial pneumonia with positive blood culture whereas CRP concentration was not. Specificity and sensitivity were lower for leucocyte count and IL-6 concentration. CONCLUSIONS: PCT concentration, with a threshold of 1 microg/l is more sensitive and specific and has greater positive and negative predictive values than CRP, IL-6, or white blood cell count for differentiating bacterial and viral causes of community pneumonia in untreated children admitted to hospital as emergency cases. PMID- 11259235 TI - A novel scheme for the reporting of adverse drug reactions. AB - BACKGROUND: The safety of medicines used in children is of considerable public interest, yet available data to monitor the safety of medicines in children is limited. AIMS: To raise awareness and stimulate reporting of adverse drug reactions (ADRs) in children in the Trent region. METHODS: A pilot Paediatric Regional Monitoring Centre (PRMC) has been established in the Trent region. The scheme operates as an extension of the UK's spontaneous reporting scheme, the Yellow Card Scheme run by the Medicines Control Agency and the Committee on Safety of Medicines. Proactive interventions including a monthly reminder letter and presentations to staff in the identified hospitals have been made. RESULTS: During the first year of the PRMC, 95 reports were received from the Trent region compared to 40 for the previous year. Twenty four of these reports were for medicines used "off label". The 95 reports involved 105 drugs and 171 suspected ADRs. Twenty six of the ADRs (15%) were considered medically significant. CONCLUSIONS: The number of ADR reports from the Trent region has increased considerably in the first year of the scheme. The results show that intensive education and promotion of ADR reporting can result in a major increase in reporting. This initiative will increase our knowledge about the safety of medicines used to treat children and so help protect public health. PMID- 11259236 TI - Final height of short subjects of low birth weight with and without growth hormone treatment. AB - AIM: To compare final height in two groups of low birth weight children examined for short stature: the first group untreated because of normal growth hormone (GH) secretion, the second treated with human growth hormone (hGH) because of abnormal secretion. METHODS: A total of 49 subjects born at term of birth weight below the 10th centile were consecutively examined for idiopathic short stature. The first group of subjects (n = 20) with normal GH peaks after pharmacological tests (>8 microg/l) spontaneously reached final height. The second group (n = 29) with abnormal secretion were treated with hGH (20 U/m(2)/week) for 36-84 months. At diagnosis the two groups were of similar height for chronological age and bone age, and had similar target height. RESULTS: In both groups final height was significantly lower than target height (-0.65 (SEM 0.20) in untreated cases, 0.61 (0.18) in treated cases). Fewer than one third of subjects had a final height above target height. Final height data of untreated and treated cases were not different. In the treated group the best results were obtained by those subjects who improved their height for bone age after three years of therapy. CONCLUSIONS: Our subjects with birth weight below the 10th centile remained as short adults with final height below target height. Treatment with hGH 20 U/m(2)/week in those diagnosed as deficient was not effective, with final results overlapping those of untreated subjects. PMID- 11259238 TI - Eosinophilic cystitis. AB - We describe four cases of eosinophilic cystitis in whom no specific cause could be found, and review the literature. Complaints at presentation included urgency, frequency, abdominal pain, and haematuria. In three patients the symptoms and ultrasound pictures suggested a bladder tumour. One patient was treated with anticholinergics and corticosteroids without relief of symptoms; a localised eosinophilic tumour was excised in one patient who remained symptom free; and two patients were managed conservatively with spontaneous resolution of bladder pathology and symptoms. One case was identified by random bladder biopsy in 150 consecutive patients with unexplained irritable micturition complaints. Eosinophilic cystitis is rare in children. After biopsy, we consider a wait and see policy is justified as symptoms tend to disappear spontaneously. Routine bladder biopsies in children with unexplained bladder symptoms is not justifiable. PMID- 11259239 TI - Crushed prednisolone tablets or oral solution for acute asthma? AB - In a randomised trial, treatment with prednisolone in two formulations (oral solution or crushed tablets) was compared in 78 young children with acute asthma. Prednisolone oral solution was better tolerated than crushed tablets (less vomiting, superior taste); clinical resolution was similar. PMID- 11259240 TI - Extent of fussing and colic type crying preceding atopic disease. AB - In a prospective follow up of 116 high risk infants, a 24 hour behavioural chart on seven consecutive days was analysed at seven and 12 weeks of age. Of children who manifested atopic disease at 2 years, 44/116 (38%), had shown significantly more fussing during the seventh, and colic type cry during the twelfth week than those who remained healthy (72/116, 62%). PMID- 11259241 TI - The incidence and distribution of Legg-Calve-Perthes' disease in Liverpool, 1982 95. AB - AIMS: To determine the incidence and distribution of Legg-Calve-Perthes' disease in Liverpool, in the period 1982-95. METHODS: Examination of information in a register, analysing the patients' addresses by indices of deprivation. RESULTS: A total of 122 white children were diagnosed as having Perthes' disease during the study, whereas black and minority groups form 5.8% of the population. The incidence rate in inner Liverpool had decreased to 10.5 in the period 1990-95. Simple Spearman correlations revealed an association between the disease incidence in electoral wards and deprivation. Regression analysis showed that for the period 1990-95 the most powerful effects on incidence were increases in ward deprivation since 1976, the percentage free school meals in 1986, the ward Health Index in 1981, and the percentage low birth weight in 1981. CONCLUSIONS: We suggest that environmental influences may come into play some years before a child presents with pain in the hip. There may be a genetic predisposition to the disease. PMID- 11259243 TI - Fertility preservation for children treated for cancer (2): ethics of consent for gamete storage and experimentation. PMID- 11259242 TI - Fertility preservation for children treated for cancer (1): scientific advances and research dilemmas. PMID- 11259244 TI - Normal bone mineral density in cystic fibrosis. AB - BACKGROUND: Osteoporosis has been reported as a complication of cystic fibrosis (CF). AIMS: To measure bone mineral density (BMD) in non-acutely ill adults and bone mineral content (BMC) in children with CF. METHODS: We analysed data from 28 adults and 13 children with CF. Corticosteroid use was minimal for the year prior to study in both groups. Dual x ray absorptiometry was used to measure total body and regional bone mineral density in adults. In children, whole body BMC was measured. Lean tissue mass (LTM) was also measured in all subjects. There were two control groups: A (matched for LTM and height, in addition to age and gender); and B (matched for age and gender only). RESULTS: There was no difference in whole body or regional BMD density between adult CF patients and control A subjects. Both whole body and regional BMD were significantly lower in adult CF patients than in control B subjects. Total body BMD was correlated with body mass index, LTM, and percent fat in both CF and control subjects. There was no significant correlation between total body BMD or regional BMD and either NIH clinical status scores, or pulmonary function tests in adults. There was no difference in total body BMC between CF children and control A subjects. Total body BMC was significantly lower in CF children than in control B subjects. There was no correlation between pulmonary function results and BMC in children. CONCLUSION: Osteopenia and osteoporosis in CF may be caused more by malnutrition and chronic use of intravenous or oral corticosteroids than by a CF related inherent defect in BMD. Appropriate "normal" data should be selected when determining whether or not osteoporosis is present in a CF patient. PMID- 11259245 TI - Increased cerebrospinal fluid concentrations of soluble Fas (CD95/Apo-1) in hydrocephalus. AB - BACKGROUND AND AIMS: The ventricular enlargement observed in children with chronically raised intracranial pressure (ICP) causes a secondary loss of brain tissue. In animal studies of hydrocephalus, programmed cell death (apoptosis) has been found as a major mechanism of neuronal injury. One of the regulators of the apoptotic cell death programme is the receptor mediated Fas/Fas ligand interaction. METHODS: The apoptosis regulating cytokines soluble Fas (sFas) and soluble Fas ligand (sFasL) were studied in the cerebrospinal fluid (CSF) of 31 hydrocephalic children undergoing shunt surgery for symptomatic hydrocephalus and 18 controls. RESULTS: High concentrations of sFas were observed in children with hydrocephalus (median 252 ng/ml); in controls sFas was below the detection limit (0.5 ng/ml). sFasL was undetectable in all but one sample. CONCLUSION: High concentrations of sFas in the CSF of children with hydrocephalus suggest intrinsic sFas production, potentially antagonising pressure mediated Fas activation. PMID- 11259246 TI - Glucocorticoid regulation of human and ovine parturition: the relationship between fetal hypothalamic-pituitary-adrenal axis activation and intrauterine prostaglandin production. AB - Birth in many animal species and in humans is associated with activation of hypothalamic-pituitary-adrenal function in the fetus and the increased influence of glucocorticoids on trophoblast cells of the placenta and fetal membranes. We suggest that in ovine pregnancy glucocorticoids directly increase fetal placental prostaglandin production, and indirectly increase prostaglandin production by maternal uterine tissues through the stimulation of placental estradiol synthesis. The events of ovine parturition are compared with those of human parturition. In the latter, we suggest similar direct effects of glucocorticoids on prostaglandin synthesis and metabolism in fetal membranes and similar indirect effects mediated by glucocorticoid-stimulated increases in intrauterine corticotropin-releasing hormone expression. PMID- 11259247 TI - Angiogenesis in the placenta. AB - The mammalian placenta is the organ through which respiratory gases, nutrients, and wastes are exchanged between the maternal and fetal systems. Thus, transplacental exchange provides for all the metabolic demands of fetal growth and development. The rate of transplacental exchange depends primarily on the rates of uterine (maternal placental) and umbilical (fetal placental) blood flows. In fact, increased uterine vascular resistance and reduced uterine blood flow can be used as predictors of high risk pregnancies and are associated with fetal growth retardation. The rates of placental blood flow, in turn, are dependent on placental vascularization, and placental angiogenesis is therefore critical for the successful development of viable, healthy offspring. Recent studies, including gene knockouts in mice, indicate that the vascular endothelial growth factors represent a major class of placental angiogenic factors. Other angiogenic factors, such as the fibroblast growth factors or perhaps the angiopoietins, also may play important roles in placental vascularization. In addition, recent observations suggest that these angiogenic factors interact with the local vasodilator nitric oxide to coordinate placental angiogenesis and blood flow. In the future, regulators of angiogenesis that are currently being developed may provide novel and powerful methods to ensure positive outcomes for most pregnancies. PMID- 11259248 TI - Prostaglandin F(2alpha) receptor in the corpus luteum: recent information on the gene, messenger ribonucleic acid, and protein. AB - The prostaglandin (PG) F(2alpha) receptor (FPr) in the corpus luteum is essential for maintaining normal reproductive cyclicity in many species. Activation of this seven-transmembrane spanning receptor at the end of the cycle leads to a decrease in progesterone and the demise of the corpus luteum (luteolysis). Recently, the gene structure of the FPr in three mammalian species has been elucidated; however, promoter regulation of the gene is still poorly understood. The FPr mRNA is extremely low in steroidogenic follicular cells (theca or granulosa) but is expressed at high levels in the corpus luteum, particularly in the large luteal cells. Treatment with PGF(2alpha) decreased FPr mRNA expression in luteal cells in most species that have been studied. Key amino acids have been suggested to be critical for binding of FPr to PGF(2alpha) based on three-dimensional modeling and comparisons with other G-protein-coupled receptors. Moieties of the PGF(2alpha) molecule that are essential for binding or specificity of binding to the FPr have been identified by radioreceptor binding studies. In this article, recent information is reviewed on the structure of the FPr gene, regulation of luteal FPr mRNA, and receptor/ligand interaction requirements for the FPr protein. PMID- 11259249 TI - An ovarian progastricsin is present in the trout coelomic fluid after ovulation. AB - An up-regulated cDNA fragment was isolated using a differential display polymerase chain reaction between ovulatory and postovulatory brook trout ovarian tissues. Using this fragment as a probe, a full-length cDNA of 1783 base pairs was obtained from an ovarian cDNA library. The cDNA presumably codes for a 383 amino acid protein with strong sequence similarity to an aspartic protease, progastricsin (EC 3.4.23.3), also known as pepsinogen C. On Northern blots of ovarian tissue, the trout progastricsin cDNA hybridized with a 1.8-kilobase transcript that was strongly up-regulated 4-6 days after ovulation. Of all other tissues tested, a transcript was only detected in the stomach. A recombinant trout progastricsin protein was produced and used to raise an antibody. On Western blots of ovarian tissue, the progastricsin antibody recognized a single 39-kDa protein that was present in the ovary only following ovulation. On Western blots of coelomic fluid, the 39-kDa protein was strongly detected 4-10 days after ovulation. The trout progastricsin was immunocytochemically localized to the granulosa cells of postovulatory follicles, suggesting that it is released from this tissue into the coelomic fluid following ovulation. Progastricsin has been found in the stomach, prostate, seminal vesicle, seminal fluid, and pancreas of vertebrates; however, this is the first report of a progastricsin in an animal ovary. PMID- 11259250 TI - Influences of age and ovarian follicular reserve on estrous cycle patterns, ovulation, and hormone secretion in the Long-Evans rat. AB - This study examined the influences of aging and reduced ovarian follicular reserve on estrous cyclicity, estradiol (E(2)) production, and gonadotropin secretion. Young virgin and middle-aged (MA) retired breeder female rats were unilaterally ovariectomized (ULO) or sham operated (control). Unilateral ovariectomy of young rats reduced the ovarian follicular reserve by one-half, to a level similar to that found in MA controls. Unilateral ovariectomy of MA females reduced the follicular pool further, to one half of MA controls. The incidence of regular cyclicity was significantly lower in MA ULO females than in young controls, with intermediate cycle frequency in young ULO and MA controls. Among cyclic rats, the magnitude of the proestrous LH surge was highest in young controls, intermediate in young ULO rats and MA controls, and lowest in MA ULO females. Similarly, ovulation rates were highest in young controls, intermediate in young ULO rats and MA controls, and lowest in MA ULO females. While young ULO rats exhibited augmented secondary FSH surges on estrous morning, middle-aged ULO females displayed secondary FSH levels comparable to young controls. The effects of age and reduced follicle number on estrous cyclicity and gonadotropin secretion were not due to altered E(2) secretion, as preovulatory E(2) levels were similar among all groups. Thus, experimental reduction in the follicular reserve exerts acute effects on the preovulatory LH surge, ovulation rate, and estrous cyclicity in both young and MA rats. However, decreased follicle number increases FSH levels only in young rats, indicating aging-related alterations in the feedback regulation of FSH. PMID- 11259251 TI - Long-term cortisol treatment inhibits pubertal development in male common carp, Cyprinus carpio L. AB - The onset and regulation of puberty is determined by functional development of the brain-pituitary-gonad (BPG) axis. Stress has been shown to interfere with reproduction and the functioning of the BPG axis. The response to chronic and severe stress may require much energy and force the organism to make adaptive choices. Energy that is normally available for processes like growth, immune response, or reproduction will be channeled into restoration of the disturbed homeostasis. Cortisol plays a key role in the homeostatic adaptation during or after stress. In the present study, immature common carp were fed with cortisol containing food pellets covering the pubertal period. We showed that cortisol caused an inhibition of pubertal development, by affecting directly or indirectly all components of the BPG axis. The salmon GnRH content of the brain was decreased. Luteinizing hormone- and FSH-encoding mRNA levels in the pituitary and LH plasma levels were diminished by long-term cortisol treatment, as was the testicular androgen secretion. Testicular development, reflected by gonadosomatic index and the first wave of spermatogenesis, was retarded. PMID- 11259252 TI - Organization of the human gene encoding the epididymis-specific EP2 protein variants and its relationship to defensin genes. AB - The EP2 gene codes for at least nine message variants that are all specifically expressed in the epididymis. These variants putatively encode small secretory proteins that differ in their N- and C-termini, resulting in proteins that can have little or no sequence similarity to each other. We have isolated and sequenced the human EP2 gene to determine the molecular origin of these variants. The EP2 gene has two promoters, eight exons, and seven introns. Exons 3 and 6 encode protein sequences homologous to beta-defensins, a family of antimicrobial peptides. This sequence homology and the arrangement of promoters and defensin encoding exons suggest that the EP2 gene originated from two ancestral beta defensin genes arranged in tandem, each contributing a promoter and two exons encoding a leader sequence and a defensin peptide. The proposed evolutionary relationship between the EP2 gene and defensin genes is supported by the observation that the EP2 gene is located on chromosome 8p23 near the defensin gene cluster and is separated by 100 kilobases or less from DEFB2, the gene for beta-defensin-2. While the EP2 gene transcribes beta-defensin-like message variants, most of the known message variants code for nondefensin proteins or proteins containing only a partial defensin peptide sequence. We suggest that, during its evolution, the EP2 gene has acquired new functions that may be important for sperm maturation and/or storage in the epididymis. PMID- 11259253 TI - Apolipoprotein E is a putative autocrine regulator of the rat ovarian theca cell compartment. AB - Apolipoprotein (apo) E inhibits androgen production by ovarian theca cells. We found that apo E, as a synthetic peptide mimicked the full-size protein, induced theca and interstitial cell (TIC) apoptosis indicated by pyknotic cell morphology, increased DNA end-labeling (TUNEL), and DNA ladders. None of the low density lipoprotein (LDL) receptor superfamily members were involved because the universal antagonist of these receptors, receptor-associated protein (RAP), did not block apo E-induced apoptosis. Furthermore, several apo E synthetic peptides that do not bind the LDL receptor did induce TIC apoptosis. Similar to apo E, apoptogenic agents such as ceramide and LY 294002, a phosphatidylinositol (PI) 3 kinase inhibitor, induced apoptosis and suppressed androstenedione production. However, apoptosis alone was not responsible for apo E suppression of androstenedione production because both insulin and IGF-I prevented apo E-induced apoptosis, but neither restored androstenedione production. Theca cells of atretic follicles express the greatest apo E mRNA, and here we show that cultured TIC produce apo E. When considered with the observation of TUNEL-positive theca cells in atretic follicles these results support our hypothesis that intraovarian apo E controls theca cell production of androgen as well as limiting the size of the theca cell compartment. PMID- 11259254 TI - Molecular characterization and high expression during oocyte development of a shrimp ovarian cortical rod protein homologous to insect intestinal peritrophins. AB - Penaeoid shrimp oocytes nearing the completion of oogenesis are enveloped in an acellular vitelline envelope and possess extracellular cortical rods (CRs) that extended into the cortical cytoplasm. These cortical specializations are precursors of the jelly layer (JL) of the egg. In searching for highly expressed mRNAs during oogenesis in the marine shrimp (Penaeus semisulcatus), two related cDNAs have been isolated that encode a mature protein of 250 amino acid residues. The deduced amino acid sequences revealed the presence of repeated cysteine-rich domains that are related to the chitin-binding domains of insect intestinal peritrophins. Similar cysteine-rich domains were reported in insect intestinal mucin, crustacean tachycitin, and invertebrate chitinases. The shrimp ovarian peritrophin (SOP) is glycosylated and can bind chitin when extracted from CRs. Its apparent molecular mass in SDS-PAGE is 29-35 kDa and 33-36 kDa, under nonreducing or reducing conditions, respectively. SOP is a major protein of CRs and the JL, and was immunodetected in ovaries; purified CRs; fertilized eggs that were surrounded by a JL matrix; and in the cloudy, whitish flocculent material appearing in sea water immediately after spawning. Immunolocalization in tissue sections determined that SOP was present in oocyte cytoplasm and in extraoocytic CRs. Shrimp expressed SOP mRNA in ovaries at all oocyte developmental stages, whereas expression in the hepatopancreas was restricted to vitellogenic stages. SOP mRNA was abundant in the shrimp ovary and was detected before the presence of the corresponding protein. This is the first demonstration that a protein with similar features to insect intestinal peritrophins is a component of CRs and is therefore a main precursor of the JL of spawned shrimp eggs. PMID- 11259255 TI - Mating induces gonadotropin-releasing hormone neuronal activation in anosmic female ferrets. AB - In females of both spontaneously and induced ovulating species, pheromones from male conspecifics can directly stimulate GnRH neuronal activity, thereby inducing pituitary LH secretion and stimulating the onset of estrus. However, whether pheromones contribute to the steroid- or mating-induced preovulatory activation of GnRH neurons is less clear. Previous studies in the ferret, an induced ovulator, raised the possibility that olfactory cues contribute to the ability of genital-somatosensory stimulation to activate GnRH neurons in the mediobasal hypothalamus (MBH). In the present study the percentage of GnRH neurons colabeled with Fos-immunoreactivity (IR), used as a marker for neuronal activation, was investigated in the MBH of mated gonadectomized, estradiol-treated female ferrets in which both nares were occluded. In addition, the percentage of GnRH neurons colabeled with Fos-IR was examined in the MBH of gonadectomized, estradiol treated female ferrets exposed to male bedding. Bilateral nares occlusion successfully blocked mating or odor-induced increments in Fos-IR in central olfactory regions, including the cortical and medial amygdala. By contrast, the percentage of GnRH neurons expressing Fos-IR did not differ between mated nares- and sham-occluded females. Exposure to male bedding alone failed to induce Fos-IR in MBH GnRH neurons. Thus, the mating-induced preovulatory activation of GnRH neurons in the female ferret's MBH appears to rely solely on genital somatosensory as opposed to olfactory inputs. PMID- 11259256 TI - Phosphatidylinositol 3-kinase in cumulus cells and oocytes is responsible for activation of oocyte mitogen-activated protein kinase during meiotic progression beyond the meiosis I stage in pigs. AB - The roles of phosphatidylinositol 3-kinase (PI 3-kinase) during meiotic progression beyond the meiosis I (MI) stage in porcine oocytes were investigated. PI 3-kinase exists in cumulus cells and oocytes, and the PI 3-kinase inhibitor, LY294002, suppressed the activation of mitogen-activated protein (MAP) kinase in denuded oocytes during the beginning of the treatment. However, in denuded oocytes cultured with LY294002, the MAP kinase activity steadily increased, and at 48 h of cultivation MAP kinase activity, p34(cdc2) kinase activity, and proportion of oocytes that had reached the meiosis II (MII) stage were at a similar level to those of oocytes cultured without LY294002. In contrast, LY294002 almost completely inhibited the activation of MAP kinase, p34(cdc2) kinase activity, and meiotic progression to the MII stage in oocytes surrounded with cumulus cells throughout the treatment. Treating cumulus oocyte complexes (COCs) with LY294002 produced a significant decrease in the phosphorylation of connexin-43, a gap junctional protein, in cumulus cells compared with that in COCs cultured without LY294002. These results indicate that PI 3-kinase activity in cumulus cells contributes to the activation of MAP kinase and p34(cdc2) kinase, and to meiotic progression beyond the MI stage. Moreover, gap junctional communications between cumulus cells and oocytes may be closed by phosphorylation of connexin-43 through PI 3-kinase activation in cumulus cells, leading to the activation of MAP kinase in porcine oocytes. PMID- 11259257 TI - Evaluation of the 5'-flanking regions of murine glutathione peroxidase five and cysteine-rich secretory protein-1 genes for directing transgene expression in mouse epididymis. AB - Based on strong epididymal expression of the mouse glutathione peroxidase 5 (GPX5) and cysteine-rich secretory protein-1 (CRISP-1) genes, we evaluated whether the 5.0-kilobase (kb)-long GPX5 and 3.8-kb-long CRISP-1 gene 5'-flanking regions could be used to target expression of genes of interest into the epididymis in transgenic mice. Of the two candidate promoters investigated, the CRISP-1 promoter-driven enhanced green fluorescent protein (EGFP) reporter gene was highly expressed in the tubular compartment of the testis in all stages of the seminiferous epithelial cycle between pachytene spermatocytes at stage VII to elongated spermatids at step 16. In contrast to CRISP-1, the 5.0-kb 5' region of the mouse GPX5 gene directed EGFP expression to the epididymis. In the various GPX5-EGFP mouse lines, strongest expression of EGFP mRNA was found in the epididymis, but low levels of reporter gene mRNA were detected in several other tissues. Strong EGFP fluorescence was found in the principal cells of the distal caput region of epididymis, and few fluorescent cells were also detected in the cauda region. No EGFP fluorescence was detected in the corpus region or in the other tissues analyzed. Hence, it is evident that the 5.0-kb 5'-flanking region of GPX5 promoter is suitable for directing the expression of structural genes of interest into the caput epididymidis in transgenic mice. PMID- 11259258 TI - Ovarian tumorigenesis in mice transgenic for murine inhibin alpha subunit promoter-driven Simian Virus 40 T-antigen: ontogeny, functional characteristics, and endocrine effects. AB - We previously reported formation of ovarian granulosa cell tumors with 100% penetration in a transgenic mouse model with murine inhibin alpha subunit promoter-driven (inhalpha)/Simian Virus 40 T-antigen (Tag). The tumor-bearing inhalpha/Tag mice showed highly elevated serum levels of immunoreactive inhibin. To investigate the onset of tumorigenesis and related endocrine consequences, 6-8 female mice of two inhalpha/Tag lines and their mating control littermates were killed monthly between 1 and 6 mo of age. We also investigated tumorigenesis related fertility aspects of these two mouse lines. The ontogeny and progression of tumors could be monitored in both inhalpha/Tag lines by alterations of ovarian weights and serum hormone levels. Serum progesterone levels increased in both inhalpha/Tag lines in an age-dependent manner as ovarian tumorigenesis progressed, and a reciprocal decrease occurred in serum LH and FSH. Neither serum estradiol (E(2)) nor uterine weights were significantly altered during tumorigenesis, suggesting that the ovarian tumors represented late stages of granulosa cell differentiation. In conclusion, the present findings show in the inhalpha/Tag TG mice a relation between endocrine consequences of granulosa cell tumorigenesis, and a connection of onset of tumor formation with aberrant steroidogenesis and gonadotropin secretion. These findings indicate that tumors are endocrinologically active and able to exert enhanced negative feedback effects on pituitary function. The tumors provide a good model for endocrinologically active hormone-dependent tumors. PMID- 11259259 TI - Effects of gestational age and labor on expression of prostanoid receptor genes in baboon uterus. AB - We determined the effect of gestational age and labor on the regional expression of prostanoid receptor genes in baboon myometrium. Cesarean hysterectomy was performed on 15 pregnant baboons of known gestational age in the last third of pregnancy, five of them during spontaneous term labor. Expression of prostanoid receptor genes was studied using Northern blot analysis. Transcripts of similar size to the human were detected for prostanoid EP(1), EP(2), EP(3), EP(4), IP, FP, and TP receptor genes using Northern blot analysis. There were no gestational age-related changes in expression of these genes. Expression of EP(1), EP(3), and IP receptor RNA mRNA was significantly higher in myometrium from the fundus (compared with the lower segment), whereas EP(2) gene expression was significantly lower in the fundus. Labor was associated with a reduction in the regional variation of both EP(2) and IP receptor gene expression, but not EP(1) and EP(3) expression. Labor was also associated with an overall lower level of expression of EP(2) receptor mRNA. We conclude that regional and labor-related variation in myometrial expression of prostanoid receptor genes may have a key role in primate parturition. PMID- 11259260 TI - Spermatid-specific expression of Iba1, an ionized calcium binding adapter molecule-1, in rat testis. AB - Postnatal development of mammalian seminiferous tubules can be divided into three phases: spermatogonial mitosis, spermatocyte meiosis, and a postmeiotic phase in which drastic morphological changes occur in spermatids (spermiogenesis). In an attempt to elucidate the molecular mechanisms involved in spermiogenesis, we have applied a differential display method to identify genes that are developmentally up-regulated during rat testis development. One of the cDNA fragments isolated by differential display turned out to be iba1, an ionized calcium binding adapter molecule-1, that contains two EF hand-like motifs. Expression of iba1 mRNA in the rat testis was detected first at 4 wk in postnatal development and then increased up to adulthood. Using the antibody against a synthetic peptide corresponding to the N-terminal Iba1 protein, we discovered that Iba1 protein was not detectable by immunohistochemistry in spermatogonia, spermatocytes, and round spermatids in adult rat testis but was specifically expressed in the cytoplasm of elongate spermatids (steps 10-19) as well as in residual bodies that are ultimately engulfed by Sertoli cells. In situ hybridization, on the other hand, revealed that iba1 mRNA is present in round spermatids as well as early elongate spermatids (steps 1-12) but not in late spermatids, suggesting that iba1 mRNA undergoes post-transcriptional regulation. Because Iba1 protein is specifically expressed in the cytoplasm of elongate spermatids, which is finally engulfed as residual bodies into Sertoli cells, we suggest that Iba1 may be involved in the final stage of spermiogenesis (i.e., in elimination of the residual cytoplasm from spermatids). PMID- 11259261 TI - Purified and refolded recombinant bonnet monkey (Macaca radiata) zona pellucida glycoprotein-B expressed in Escherichia coli binds to spermatozoa. AB - Bonnet monkey (Macaca radiata) zona pellucida glycoprotein-B (bmZPB), excluding the N:-terminal signal sequence and the C:-terminus transmembrane-like domain, has been expressed in Escherichia coli as polyhistidine fusion protein. A requirement of 4 M urea to maintain the purified protein in soluble state rendered it unsuitable for biological studies. Purification of refolded r-bmZPB without urea and devoid of lower molecular weight fragments was achieved by following an alternate methodology that involved purification of inclusion bodies to homogeneity and solubilization in the presence of a low concentration of chaotropic agent (2 M urea) and high pH (pH 12). The solubilized protein was refolded in the presence of oxidized and reduced glutathione. The circular dichroism spectra revealed the presence of both alpha helical and beta sheet components in the secondary structure of the refolded r-bmZPB. The binding of the refolded r-bmZPB to the spermatozoa was evaluated by an indirect immunofluorescence assay and also by direct binding of the biotinylated r-bmZPB. The binding was restricted to the principal segment of the acrosomal cap of capacitated bonnet monkey spermatozoa. In the acrosome-reacted spermatozoa a shift in the binding pattern of r-bmZPB was observed and it bound to the equatorial segment, postacrosomal domain, and midpiece region. Binding of biotinylated r-bmZPB was inhibited by cold r-bmZPB as well as by monoclonal and polyclonal antibodies generated against r-bmZPB. These results suggest that nonglycosylated bmZPB binds to capacitated as well as acrosome-reacted spermatozoa in a nonhuman primate and may have a functional role during fertilization. PMID- 11259262 TI - Effect of bcl-2 on the primordial follicle endowment in the mouse ovary. AB - Little is known about the embryonic factors that regulate the size of the primordial follicle endowment at birth. A few studies suggest that members of the B-cell lymphoma/leukemia-2 (bcl-2) family of protooncogenes may be important determinants. Thus, the purpose of this study was to test whether bcl-2 regulates the size of the primordial follicle pool at birth. To test this hypothesis, three lines of transgenic mice (c-kit/bcl-2 mice) were generated that overexpress human bcl-2 in an effort to reduce prenatal oocyte loss. The overexpression was targeted to the ovary and appropriate embryonic time period with the use of a 4.8 kilobase c-kit promoter. This promoter provided two to three times more expression of bcl-2 in the ovaries with minimal or no overexpression in most nongonadal tissues. On Postnatal Days 8-60, ovaries were collected from homozygous c-kit/bcl-2 and nontransgenic littermates (controls) and processed for histological evaluation of follicle numbers. All lines of c-kit/bcl-2 mice were born with significantly more primordial follicles than control mice (P < or = 0.05). By Postnatal Days 30-60, however, there were no significant differences in follicle numbers between c-kit/bcl-2 and control mice. These results indicate that bcl-2 overexpression increases the number of primordial follicles at birth, but that the surfeit of primordial follicles is not maintained in postnatal life. These data suggest that it is possible that the ovary may contain a census mechanism by which excess numbers of primordial follicles at birth are detected and removed from the ovary by adulthood. PMID- 11259263 TI - Fos-induction in gonadotropin-releasing hormone neurons receiving vasoactive intestinal polypeptide innervation is reduced in middle-aged female rats. AB - A hallmark of reproductive aging in rats is a delay in the initiation and peak, and a decrease in the amplitude, of both proestrous and steroid-induced surges of LH and a decrease in the number of GnRH neurons that express Fos during the surge. The altered timing of the LH surge and the decline in Fos expression in GnRH neurons may be due to changes in the rhythmic expression of vasoactive intestinal polypeptide (VIP), a neuropeptide that carries time-of-day information from the circadian pacemaker, located in the suprachiasmatic nuclei (SCN), to GnRH neurons. The goals of our study were to determine if aging alters 1) the innervation of GnRH neurons by VIP and 2) the ability of VIP to activate GnRH neurons by examining the effects of aging on the number of GnRH neurons apposed by VIP fibers and the number of GnRH neurons that receive VIP input that express Fos. Immunocytochemistry for GnRH and VIP; or GnRH, VIP, and Fos was performed on tissue sections collected from young (2-4 mo), regularly cycling females and middle-aged (10-12 mo) females in constant estrus. The number of GnRH neurons, GnRH neurons apposed by VIP fibers, and GnRH neurons that express Fos and apposed by VIP fibers were counted in both age groups. Our results clearly demonstrate that aging does not alter the number of GnRH neurons that receive VIP innervation. However, the number of GnRH neurons that receive VIP innervation and coexpress Fos decreases significantly. We conclude that the age-related delay in the timing of the LH surge is not due to a change in VIP innervation of GnRH neurons, but instead may result from a decreased sensitivity of GnRH neurons to VIP input. PMID- 11259264 TI - Expression of heparin/heparan sulfate interacting protein/ribosomal protein l29 during the estrous cycle and early pregnancy in the mouse. AB - Using a variety of approaches, we have examined the expression of the heparin/heparan sulfate (Hp/HS) interacting protein/ribosomal protein L29 (HIP/RPL29) in mouse uteri during the estrous cycle and early pregnancy. HIP/RPL29 selectively binds heparin and HS and may promote HS-dependent embryo adhesion. HIP/RPL29 was prominently expressed in both luminal and glandular epithelia under almost all conditions, including the phase of embryo attachment. In contrast, differences were noted in HIP/RPL29 expression in the stromal compartment both during the estrous cycle and during early pregnancy. Most notably, HIP/RPL29 accumulated in decidua, where it displayed a pattern complementary to that of pericellular deposition of the HS proteoglycan, perlecan. HIP/RPL29 protein was detected in implanted embryos at both initial and later stages of implantation; however, embryonic HIP/RPL29 mRNA accumulation was more pronounced at later stages (Day 7.5 postcoitum). In situ hybridization revealed similar spatial changes for HIP/RPL29 mRNA during these different physiological states. Whereas differences in the spatial pattern of HIP/RPL29 protein and mRNA expression were demonstrable, little change was detected in the level of HIP/RPL29 mRNA or protein in total endometrial extracts. Mouse blastocysts attached, but did not outgrow, on surfaces coated with recombinant murine HIP/RPL29. Surprisingly, soluble glycosaminoglycans including heparin, low molecular weight heparin, or chondroitin sulfate were not able to inhibit embryo attachment to HIP/RPL29-coated surfaces. These latter observations indicate that embryonic cell surface components other than HS proteoglycans can promote binding to HIP/RPL29 expressed by uterine cells. PMID- 11259265 TI - Annual ovarian cycles in an aseasonal breeder, the springbok (Antidorcas marsupialis). AB - The springbok is an arid-adapted antelope inhabiting the desert and semidesert regions of southern Africa. Because it thrives in these sparsely vegetated areas, the springbok is of potential agricultural importance and the prospect of domestication has been speculated for many years. However, apart from observational studies on its breeding in the wild, suggesting it is an aseasonal breeder, little is known about the underlying reproductive endocrinology of this species. In this study, biweekly peripheral blood samples were collected from eight captive springbok ewes from October 1995 until September 1998 and analyzed for progesterone. At the start of the study, six ewes were prepubertal and cycling commenced spontaneously between November 1995 and June 1996. Cycling had already commenced in two ewes. At the end of November 1996, estrous cycles ceased abruptly in all ewes and restarted in April 1997. Cycling ceased again between December 1997 and February 1998 and restarted in June 1998 in six ewes; there was no cessation of estrous cycles in two ewes. Thus, although some individuals cycle continuously, there is a clear endocrine anestrus of between 4 and 5 mo in springbok, the timing and duration of which is synchronized between some individuals but the time of onset and cessation is variable from year to year. To ensure that the fluctuations we observed in progesterone levels were reliable indicators of changes in the estrous cycle, blood samples were collected every 6 h for 16 days in August 1998. A surge in LH secretion was observed in all ewes 55 +/- 5 h after the fall in progesterone. Progesterone levels increased again 45 +/ 8 h after the surge. A final study showed that the pattern of melatonin release in springbok exhibits a normal day/night profile, and thus photoperiodic information is transformed into an endocrine code to springbok but does not appear to affect reproduction. Rather, our data raise the possibility that the prevailing ambient temperature may influence the onset of ovarian activity in this species. PMID- 11259266 TI - E-cadherin-mediated cell contact prevents apoptosis of spontaneously immortalized granulosa cells by regulating Akt kinase activity. AB - The present studies were designed to determine the role that homophilic E cadherin binding plays in preventing apoptosis of spontaneously immortalized granulosa cells (SIGCs). Although the levels of E-cadherin were similar to serum control levels, the amount of E-cadherin at the plasma membrane was dramatically reduced by 5 h after serum withdrawal. To determine whether disrupting homophilic E-cadherin binding leads to apoptosis, SIGCs were cultured in serum in the presence of either EGTA or an E-cadherin antibody. Treatment with either EGTA, which disrupts all calcium-dependent contacts, or E-cadherin antibody, induced apoptosis. Exposure to EGTA reduced MEK and Akt kinase activity, whereas E cadherin antibody only attenuated Akt kinase activity. Because Akt kinase controls caspase-3 activity, an important activator of apoptosis, caspase-3 activity was monitored. Caspase-3 activity increased after serum depletion, or EGTA or E-cadherin antibody treatment. Time-series analysis of caspase-3 activity within single cells revealed that during apoptosis cell contact was disrupted then caspase-3 activity was detected. Finally, the caspase inhibitor, Z-VAD-FMK, blocked apoptosis. These data taken together are consistent with the concept that E-cadherin-mediated cell contact, either directly or indirectly, promotes Akt kinase activity, which in turn, inhibits caspase-3 activation and thereby maintains SIGC viability. PMID- 11259267 TI - Nuclear receptor Dax-1 represses the transcriptional cooperation between GATA-4 and SF-1 in Sertoli cells. AB - A crucial step in mammalian sex differentiation is the regression of the Mullerian ducts in males. This is achieved through the action of Mullerian inhibiting substance (MIS), a key hormone produced by fetal Sertoli cells. Proper spatiotemporal expression of the MIS gene requires the concerted action of several transcription factors that include Sox9, SF-1, WT-1, GATA-4, and Dax-1. Indeed, SF-1 contributes to MIS gene expression by transcriptionally cooperating with other factors such as GATA-4 and WT-1. Dax-1 is coexpressed with SF-1 in many tissues, including the gonads, where it acts as a negative modulator of SF-1 dependent transcription. We now report that Dax-1 can repress MIS transcription in Sertoli cells by disrupting transcriptional synergism between GATA-4 and SF-1. Dax-1-mediated repression of GATA-4/SF-1 synergism did not involve direct repression of GATA-dependent transactivation, but rather, it occurred through a direct protein-protein interaction with DNA-bound SF-1. It is interesting that SF 1, Dax-1, and GATA factors are coexpressed in several tissues such as the pituitary, the adrenals, and the gonads. Because we have shown that other GATA family members also have the ability to synergize with SF-1, Dax-1 repression of GATA/SF-1 synergism may represent an important mechanism for fine-tuning the regulation of SF-1-dependent genes in multiple target tissues. PMID- 11259268 TI - Macrophage migration inhibitory factor in the human endometrium: expression and localization during the menstrual cycle and early pregnancy. AB - Macrophage migration inhibitory factor (MIF) was discovered as an activated T lymphocyte-derived protein that inhibits the random migration of macrophages in vitro. Subsequently, knowledge of the physiological actions of MIF was extended to include its role as a proinflammatory cytokine that affects several functions of macrophages and lymphocytes. Previous reports have suggested an involvement of MIF in reproduction. However, no data are currently available on the presence of this cytokine in the human endometrium. In this study, the expression and tissue localization of MIF was evaluated in specimens of cycling endometrium, first trimester placenta bed biopsy, and isolated endometrial glands by Western blot analysis, immunohistochemistry, ELISA, and reverse transcription-polymerase chain reaction. The results demonstrated that MIF is expressed in human endometrium across the menstrual cycle and in early pregnancy. Immunohistochemical localization identified the protein in glandular epithelium, in stromal and predecidualized stromal cells of cycling endometrium, as well as in the decidua of first-trimester placenta. The proinflammatory features and specific actions of MIF on lymphoid cells suggest its potential involvement in several aspects of endometrial physiology. PMID- 11259269 TI - Granulocyte-macrophage colony-stimulating factor promotes glucose transport and blastomere viability in murine preimplantation embryos. AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) secretion from epithelial cells lining the female reproductive tract is induced during early pregnancy by ovarian steroid hormones and constituents of seminal plasma. In this study we have investigated the influence of GM-CSF on development of preimplantation mouse embryos. Blastocyst-stage embryos were found to specifically bind (125)I-GM-CSF and analysis of GM-CSF mRNA receptor expression by reverse transcriptase-polymerase chain reaction indicated expression of the low-affinity alpha subunit of the GM-CSF receptor, but not the affinity converting beta subunit (beta(c)), or GM-CSF ligand. GM-CSF receptor mRNA was present in the fertilized oocyte and all subsequent stages of development, and in blastocysts it was expressed in both inner cell mass and trophectoderm cells. In vitro culture of eight-cell embryos in recombinant GM-CSF accelerated development of blastocysts to hatching and implantation stages, with a maximum response at a concentration of 2 ng/ml (77 pM). Blastocysts recovered from GM-CSF-null mutant (GM-/-) mice on Day 4 of natural pregnancy or after superovulation showed retarded development, with the total cell number reduced by 14% and 18%, respectively, compared with GM+/+ embryos. Blastocysts generated in vitro from two-cell GM-/- and GM+/+ embryos were larger when recombinant GM-CSF was added to the culture medium (20% and 24% increases in total cell numbers in GM+/+ and GM-/ blastocysts, respectively). Incubation of blastocysts with recombinant GM-CSF elicited a 50% increase in the uptake of the nonmetabolizable glucose analogue, 3 O-methyl glucose. In conclusion, these data indicate that GM-CSF signaling through the low-affinity GM-CSF receptor in blastocysts is associated with increased glucose uptake and enhanced proliferation and/or viability of blastomeres. Together, the findings implicate a physiological role for maternal tract-derived GM-CSF in targeting the preimplantation embryo, and suggest that defective blastocyst development contributes to compromised pregnancy outcome in GM-CSF-null mutant mice. PMID- 11259270 TI - Exposure of neonatal female rats to p-tert-octylphenol disrupts afternoon surges of luteinizing hormone, follicle-stimulating hormone and prolactin secretion, and interferes with sexual receptive behavior in adulthood. AB - The present study investigated the effects of exposure of neonatal female rats to p-tert-octylphenol (OP) on estrogen-induced afternoon surges of LH, FSH, and prolactin (PRL) secretion, and on sexual behavior in adulthood. After birth, one group of female Wistar rat pups received s.c. injections of OP (100 mg/kg body weight [BW]; OP group) dissolved in DMSO, while the control group received DMSO only (DMSO group). In order to make a qualitative comparison, a third group was injected with estradiol-17beta (500 microg/kg BW; estradiol group) dissolved in DMSO. Injections were given on Days 1, 3, 5, 7, 9, 11, 13, and 15 of age. The rats from the OP and estradiol groups that were used for subsequent experiments were in persistent vaginal estrus. Spontaneous LH surge measured at Postnatal Days (PND) 78-81 was observed only in the DMSO group on the afternoon of the day of proestrus. At PND 115, randomly selected rats from each of three treatment groups were bilaterally ovariectomized (ovx), and 8 days later, Silastic capsules containing estradiol-17beta were implanted under the skin. Estrogen implants stimulated afternoon surges of LH, FSH, and PRL for two consecutive days in the DMSO group, but not in the OP and estradiol groups. Rats from the OP and DMSO groups underwent ovx at PND 186, and 6 days later they were treated with a combination of estradiol benzoate s.c. (15 microg/kg BW) and progesterone s.c. (2 mg/kg BW) to test the lordosis reflex. In response to this hormone treatment and mounting stimulus delivered by the stud male rats, the OP-treated rats were less receptive compared with control DMSO-treated rats, and thus the lordosis quotient and lordosis rating were significantly (P < 0.05) reduced in the OP group compared with the DMSO group. Analysis of the area of the sexually dimorphic nucleus of the preoptic area of the brain revealed that the area of this nucleus was larger in the OP group than it was in control DMSO rats. We conclude that the exposure of neonatal female rats to higher doses of OP disrupts the cyclic release of LH, FSH, and PRL, and interferes with the display of sexual receptive behavior in adulthood. PMID- 11259271 TI - Highly prolific Booroola sheep have a mutation in the intracellular kinase domain of bone morphogenetic protein IB receptor (ALK-6) that is expressed in both oocytes and granulosa cells. AB - The Booroola fecundity gene (FecB) increases ovulation rate and litter size in sheep and is inherited as a single autosomal locus. The effect of FecB is additive for ovulation rate (increasing by about 1.6 corpora lutea per cycle for each copy) and has been mapped to sheep chromosome 6q23-31, which is syntenic to human chromosome 4q21-25. Bone morphogenetic protein IB (BMP-IB) receptor (also known as ALK-6), which binds members of the transforming growth factor-beta (TGF beta) superfamily, is located in the region containing the FecB locus. Booroola sheep have a mutation (Q249R) in the highly conserved intracellular kinase signaling domain of the BMP-IB receptor. The mutation segregated with the FecB phenotype in the Booroola backcross and half-sib flocks of sheep with no recombinants. The mutation was not found in individuals from a number of sheep breeds not derived from the Booroola strain. BMPR-IB was expressed in the ovary and in situ hybridization revealed its specific location to the oocyte and the granulosa cell. Expression of mRNA encoding the BMP type II receptor was widespread throughout the ovary. The mutation in BMPR-IB found in Booroola sheep is the second reported defect in a gene from the TGF-beta pathway affecting fertility in sheep following the recent discovery of mutations in the growth factor, GDF9b/BMP15. PMID- 11259272 TI - Expression of messenger ribonucleic acids for fibroblast growth factors 7 and 10, hepatocyte growth factor, and insulin-like growth factors and their receptors in the neonatal ovine uterus. AB - In sheep, uterine development begins during fetal life but is only completed postnatally with proliferation and branching morphogenetic differentiation of the endometrial glandular epithelium (G) from the luminal epithelium (L) between birth or Postnatal Day (PND) 0 and PND 56. In other epithelial-mesenchymal organs, fibroblast growth factor (FGF)-7 and FGF-10, hepatocyte growth factor (HGF), and insulin-like growth factor (IGF)-I and IGF-II play essential roles in ductal branching morphogenesis. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and in situ hybridization analyses were used to study temporal and spatial alterations in expression of mRNAs for growth factors (FGF 7, FGF-10, HGF, IGF-I, IGF-II) and their respective receptors (FGF receptor or FGFR2IIIb, c-met, and IGF-IR) in the developing neonatal ovine uterus. The RT-PCR analyses indicated that expression of FGF-10, HGF, IGF-I, and IGF-II mRNAs increased in the neonatal uterus between PND 1 and 56. In situ hybridization analyses indicated that FGFR2IIIb and c-met mRNAs were expressed solely in uterine L and developing G, whereas IGF-IR was expressed in all uterine cell types, with highest levels in L and developing G. Both IGF-I and IGF-II mRNAs were expressed in the endometrial stroma and myometrium, with IGF-I predominantly in the intercaruncular endometrial stroma. The highest levels of IGF-I and IGF-II mRNA expression were detected in the intercaruncular endometrial stroma surrounding the nascent and proliferating glands. Immunohistochemistry revealed that phosphorylated extracellular regulated kinases-1 and -2 were most abundantly expressed in the nascent and proliferating glands of the developing neonatal uterine wall. These results implicate FGF-7, FGF-10, HGF, IGF-I, IGF-II, and their epithelial receptors in epithelial-mesenchymal interactions regulating endometrial gland morphogenesis in the neonatal sheep uterus. PMID- 11259273 TI - An unusual subcellular localization of GLUT1 and link with metabolism in oocytes and preimplantation mouse embryos. AB - Although mouse oocytes and cleavage-stage embryos prefer pyruvate and lactate for metabolic fuels, they do take up and metabolize glucose. Indeed, presentation of glucose during the cleavage stages is required for subsequent blastocyst formation, which normally relies on uptake and metabolism of large amounts of glucose. Expression of the facilitative glucose transporter GLUT1 was examined using immunohistochemistry and Western blotting, and in polyspermic oocytes, metabolism of glucose was measured and compared with that of pyruvate and glutamine. GLUT1 was observed in all oocytes and embryos, and membrane and vesicular staining was present. Additionally, however, in polyspermic oocytes, the most intense staining was in the pronuclei, and this nuclear staining persisted in cleaving normal embryos. Furthermore, GLUT1 expression appeared to be up-regulated both in nuclei and plasma membranes following culture of oocytes in the absence of glucose. In polyspermic oocytes, the metabolism of glucose, but not of pyruvate or glutamine, was directly proportional to the number of pronuclei formed. After compaction, nuclear staining diminished, and GLUT1 localized to basolateral membranes of the outer cells and trophectoderm. In blastocysts, a weak but uniform staining of inner-cell-mass plasma membranes was apparent. The results are discussed in terms of potential roles for GLUT1 in pronuclei of oocytes and zygotes, nuclei of cleavage-stage embryos, and a transepithelial transport function for GLUT1, probably coupled with GLUT3, in compacted embryos and blastocysts. PMID- 11259274 TI - Dynamic changes of connexin-43, gap junctional protein, in outer layers of cumulus cells are regulated by PKC and PI 3-kinase during meiotic resumption in porcine oocytes. AB - Mammalian oocytes are surrounded by numerous layers of cumulus cells, and the loss of gap junctional communication in the outer layers of cumulus cells induces meiotic resumption in oocytes. In this study, we investigated the dynamic changes in the gap junctional protein connexin-43 in cumulus cells during the meiotic resumption of porcine oocytes. The amount of connexin-43 in all layers of cumulus cells recovered from cumulus-oocyte complexes was increased after 4-h cultivation. However, at 12-h cultivation, the positive signal for connexin-43 immunoreactivity was markedly reduced in the outer layers of cumulus cells. When these reductions of connexin-43 were blocked by protein kinase C (PKC) or phosphatidylinositol (PI) 3-kinase inhibitor, networks of filamentous bivalents (i.e., advanced chromosomal status) were undetectable in the germinal vesicle of the oocyte. After 28-h cultivation, when the majority of oocytes were reaching the metaphase I (MI) stage, the connexin-43 in the inner layers of cumulus cells was phosphorylated, regardless of mitogen-activated protein (MAP) kinase activation. These results suggest that the initiation of meiotic resumption, namely, the formation of networks of filamentous bivalents in germinal vesicle, is associated with the reduction of gap junctional protein connexin-43 in the outer layers of cumulus cells via the PKC and/or PI 3-kinase pathway. Moreover, the connexin-43 in the inner layers of cumulus cells is phosphorylated during meiotic progression beyond the MI stage, regardless of MAP kinase activation in cumulus cells surrounding the oocyte. PMID- 11259275 TI - X-linked inhibitor of apoptosis protein expression and the regulation of apoptosis during human placental development. AB - In this study, we have examined the expression and potential role of X-linked inhibitor of apoptosis protein (XIAP), Fas, and Fas ligand (FasL) in the regulation of apoptosis throughout placental development. Protein expression was determined by Western blot analysis and immunohistochemistry, whereas apoptotic cell death was assessed by DNA fragmentation analysis and TUNEL. The XIAP was present in trophoblast throughout placental development, but its content significantly decreased during late pregnancy, when apoptosis was maximal. The FasL content was low during early placental development but increased coincidentally to the decrease in XIAP during the third trimester. Our data also suggest that placental apoptosis is the culmination of the relative expression of these cell-death and -survival proteins, a phenomenon that is cell type-specific and dependent on cytodifferentiation and the stage of placental development. Moreover, the induction of syncytiotrophoblast apoptosis may involve the concomitant up-regulation of FasL for Fas activation and the removal of downstream inhibition of the apoptotic cascade by XIAP. PMID- 11259276 TI - Continuously proliferative stem germ cells partially repopulate the aged, atrophic rat testis after gonadotropin-releasing hormone agonist therapy. AB - Aging in the male human is accompanied by testicular atrophy, although relatively little is known about the mechanisms underlying germ cell loss. Testicular atrophy in the aged Brown Norway rat, an animal model for studies of aging in the human, has been attributed to a loss of spermatogonial stem cells. However, examination of testicular cross-sections from 27-mo-old Brown Norway rats indicated that approximately 14% of type A spermatogonia were stem cells. Furthermore, using bromodeoxyuridine labeling, we found that approximately 47% of these stem cells were actively dividing, with a cell cycle time of approximately 12.6 days. Both serum and testicular interstitial fluid testosterone levels were depressed in the aged rat. Therapy with the GnRH agonist, leuprolide, which has been empirically shown to reverse testicular atrophy in other models of germ cell loss, also partially restored spermatogenesis in the aged Brown Norway rat. The extent of testicular atrophy varied considerably, not only within the control and leuprolide-treatment groups but also between the left and right testes of the same animals. No significant difference was found between the mean percentage of populated tubules in 31-mo-old control animals (16.2 +/- 28%, mean +/- SD) and 31 mo-old leuprolide-treated animals (20.9 +/- 19.8%), but categorical comparisons showed that significantly fewer leuprolide-treated animals and testes contained < or = 1% populated tubules, indicating that GnRH agonist therapy stimulates differentiation of type A spermatogonia. An increase in the ratio of soluble to membrane stem cell factor mRNA levels was present in aged rats and partially reversed following leuprolide therapy. PMID- 11259277 TI - Modulation of primary radical pair kinetics and energetics in photosystem II by the redox state of the quinone electron acceptor Q(A). AB - Time-resolved photovoltage measurements on destacked photosystem II membranes from spinach with the primary quinone electron acceptor Q(A) either singly or doubly reduced have been performed to monitor the time evolution of the primary radical pair P680(+)Pheo(-). The maximum transient concentration of the primary radical pair is about five times larger and its decay is about seven times slower with doubly reduced compared with singly reduced Q(A). The possible biological significance of these differences is discussed. On the basis of a simple reversible reaction scheme, the measured apparent rate constants and relative amplitudes allow determination of sets of molecular rate constants and energetic parameters for primary reactions in the reaction centers with doubly reduced Q(A) as well as with oxidized or singly reduced Q(A). The standard free energy difference DeltaG degrees between the charge-separated state P680(+)Pheo(-) and the equilibrated excited state (Chl(N)P680)* was found to be similar when Q(A) was oxidized or doubly reduced before the flash (approximately -50 meV). In contrast, single reduction of Q(A) led to a large change in DeltaG degrees (approximately +40 meV), demonstrating the importance of electrostatic interaction between the charge on Q(A) and the primary radical pair, and providing direct evidence that the doubly reduced Q(A) is an electrically neutral species, i.e., is doubly protonated. A comparison of the molecular rate constants shows that the rate of charge recombination is much more sensitive to the change in DeltaG degrees than the rate of primary charge separation. PMID- 11259278 TI - Electrical-to-mechanical coupling in purple membranes: membrane as electrostrictive medium. AB - In this paper, we present acousto-electrical measurements performed on dry films of purple membranes (PM) of Halobacterium salinarium. The purpose of these measurements is to determine the relation between mechanical and electrical phenomena in bacteriorhodopsin and to define the role of the protein in the proton transfer process. Electrical-to-mechanical coupling in PMs manifests itself as direct and inverse piezoelectric effects. Measurements performed on the samples with different degrees of PM orientation and at various values of the externally applied cross-membrane electric field indicate that piezoelectric phenomena in PMs arise from the electric asymmetry of the membranes, i.e., they originate from electrostriction. Experiments with samples made of oriented PMs allow estimation of the value of the intrinsic cross-membrane electric field, which is approximately 10(8) V/m. A hypothetical model of PM is presented where the electrical-to-mechanical coupling is suggested to be the main driving force for the proton translocation against the Coulomb forces acting in the membrane. PMID- 11259279 TI - Effects of excluded surface area and adsorbate clustering on surface adsorption of proteins. II. Kinetic models. AB - Models for equilibrium surface adsorption of proteins have been recently proposed (Minton, A. P., 2000. Biophys. Chem. 86:239-247) in which negative cooperativity due to area exclusion by adsorbate molecules is compensated to a variable extent by the formation of a heterogeneous population of monolayer surface clusters of adsorbed protein molecules. In the present work this concept is extended to treat the kinetics of protein adsorption. It is postulated that clusters may grow via two distinct kinetic pathways. The first pathway is the diffusion of adsorbed monomer to the edge of a preexisting cluster and subsequent accretion. The second pathway consists of direct deposition of a monomer in solution onto the upper (solution-facing) surface of a preexisting cluster ("piggyback" deposition) and subsequent incorporation into the cluster. Results of calculations of the time course of adsorption, carried out for two different limiting models of cluster structure and energetics, show that in the absence of piggyback deposition, enhancement of the tendency of adsorbate to cluster can reduce, but not eliminate, the negative kinetic cooperativity due to surface area exclusion by adsorbate. Apparently noncooperative (Langmuir-like) and positively cooperative adsorption progress curves, qualitatively similar to those reported in several published experimental studies, require a significant fraction of total adsorption flux through the piggyback deposition pathway. According to the model developed here and in the above-mentioned reference, the formation of surface clusters should be a common concomitant of non-site-specific surface adsorption of proteins, and may provide an important mechanism for assembly of organized "protein machines" in vivo. PMID- 11259280 TI - Molecular dynamics simulation of the structure of dimyristoylphosphatidylcholine bilayers with cholesterol, ergosterol, and lanosterol. AB - Five molecular dynamics computer simulations were performed on different phospholipid:sterol membrane systems in order to study the influence of sterol structure on membrane properties. Three of these simulated bilayer systems were composed of a 1:8 sterol:phospholipid ratio, each of which employed one of the sterol molecules: cholesterol, ergosterol, and lanosterol. The two other simulations were of a bilayer with a 1:1 sterol:phospholipid ratio. These simulations employed cholesterol and lanosterol, respectively, as their sterol components. The observed differences in simulations with cholesterol and lanosterol may have their implication on the form of the phospholipid/sterol phase diagram. PMID- 11259281 TI - Brownian dynamics simulations of interaction between scorpion toxin Lq2 and potassium ion channel. AB - The association of the scorpion toxin Lq2 and a potassium ion (K(+)) channel has been studied using the Brownian dynamics (BD) simulation method. All of the 22 available structures of Lq2 in the Brookhaven Protein Data Bank (PDB) determined by NMR were considered during the simulation, which indicated that the conformation of Lq2 affects the binding between the two proteins significantly. Among the 22 structures of Lq2, only 4 structures dock in the binding site of the K(+) channel with a high probability and favorable electrostatic interactions. From the 4 candidates of the Lq2-K(+) channel binding models, we identified a good three-dimensional model of Lq2-K(+) channel complex through triplet contact analysis, electrostatic interaction energy estimation by BD simulation and structural refinement by molecular mechanics. Lq2 locates around the extracellular mouth of the K(+) channel and contacts the K(+) channel using its beta-sheet rather than its alpha-helix. Lys27, a conserved amino acid in the scorpion toxins, plugs the pore of the K(+) channel and forms three hydrogen bonds with the conserved residues Tyr78(A-C) and two hydrophobic contacts with Gly79 of the K(+) channel. In addition, eight hydrogen-bonds are formed between residues Arg25, Cys28, Lys31, Arg34 and Tyr36 of Lq2 and residues Pro55, Tyr78, Gly79, Asp80, and Tyr82 of K(+) channel. Many of them are formed by side chains of residues of Lq2 and backbone atoms of the K(+) channel. Thirteen hydrophobic contacts exist between residues Met29, Asn30, Lys31 and Tyr36 of Lq2 and residues Pro55, Ala58, Gly79, Asp80 and Tyr82 of the K(+) channel. These favorable interactions stabilize the association between the two proteins. These observations are in good agreement with the experimental results and can explain the binding phenomena between scorpion toxins and K(+) channels at the level of molecular structure. The consistency between the BD simulation and the experimental data indicates that our three-dimensional model of Lq2-K(+) channel complex is reasonable and can be used in further biological studies such as rational design of blocking agents of K(+) channels and mutagenesis in both toxins and K(+) channels. PMID- 11259282 TI - An electrochemical model of the transport of charged molecules through the capillary glycocalyx. AB - An electrochemical theory of the glycocalyx surface layer on capillary endothelial cells is developed as a model to study the electrochemical dynamics of anionic molecular transport within capillaries. Combining a constitutive relationship for electrochemical transport, derived from Fick's and Ohm's laws, with the conservation of mass and Gauss's law from electrostatics, a system of three nonlinear, coupled, second-order, partial, integro-differential equations is obtained for the concentrations of the diffusing anionic molecules and the cations and anions in the blood. With the exception of small departures from electroneutrality that arise locally near the apical region of the glycocalyx, the model assumes that cations in the blood counterbalance the fixed negative charges bound to the macromolecular matrix of the glycocalyx in equilibrium. In the presence of anionic molecular tracers injected into the capillary lumen, the model predicts the size- and charge-dependent electrophoretic mobility of ions and tracers within the layer. In particular, the model predicts that anionic molecules are excluded from the glycocalyx at equilibrium and that the extent of this exclusion, which increases with increasing tracer and/or glycocalyx electronegativity, is a fundamental determinant of anionic molecular transport through the layer. The model equations were integrated numerically using a Crank Nicolson finite-difference scheme and Newton-Raphson iteration. When the concentration of the anionic molecular tracer is small compared with the concentration of ions in the blood, a linearized version of the model can be obtained and solved as an eigenvalue problem. The results of the linear and nonlinear models were found to be in good agreement for this physiologically important case. Furthermore, if the fixed-charge density of the glycocalyx is of the order of the concentration of ions in the blood, or larger, or if the magnitude of the anionic molecular valence is large, a closed-form asymptotic solution for the diffusion time can be obtained from the eigenvalue problem that compares favorably with the numerical solution. In either case, if leakage of anionic molecules out of the capillary occurs, diffusion time is seen to vary exponentially with anionic valence and in inverse proportion to the steady-state anionic tracer concentration in the layer relative to the lumen. These findings suggest several methods for obtaining an estimate of the glycocalyx fixed-charge density in vivo. PMID- 11259283 TI - The formation and dynamics of proton wires in channel environments. AB - By virtue of an accurate interaction model, the equilibrium and dynamical properties of an excess proton in aqueous systems are studied, in which the water and excess proton are confined to hydrophobic cylindrical channels. Solvation structures of the excess proton and its mobility along the channel are considered as a function of the channel radius. It is found that when the aqueous proton systems are sufficiently constricted there is a substantial increase in the diffusion of the excess proton charge accompanied by a decrease in the diffusion of water molecules along the channel. Such systems present clear evidence for the possible existence of "proton wires." PMID- 11259284 TI - Theory of tunable pH-sensitive vesicles of anionic and cationic lipids or anionic and neutral lipids. AB - The design of vesicles that become unstable at an easily tuned value of pH is of great interest for targeted drug delivery. We present a microscopic theory for two forms of such vesicles. A model of lipids introduced by us previously is applied to a system of ionizable anionic lipid and permanently charged cationic lipid. We calculate the pH at which the lamellar phase becomes unstable with respect to an inverted hexagonal one, a value that depends continuously on the system composition. Identifying this instability with that displayed by unilamellar vesicles undergoing fusion, we obtain very good agreement with the recent experimental data of Hafez, Ansell, and Cullis, (2000, Biophys. J. 79:1438 1446) on the pH at which fusion occurs versus vesicle composition. We explicate the mechanism in terms of the role of the counterions. This understanding suggests that a system of a neutral, nonlamellar-forming lipid stabilized by an anionic lipid would serve equally well for preparing tunable, pH-sensitive vesicles. Our calculations confirm this. Further, we show that both forms of vesicle have the desirable feature of exhibiting a regime in which the pH at instability is a rapidly varying function of the vesicle composition. PMID- 11259285 TI - Predicted profiles of ion concentrations in olfactory cilia in the steady state. AB - The role of ciliary geometry for transduction events was explored by numerical simulation. The changes in intraciliary ion concentrations, suspected to occur during transduction, could thus be estimated. The case of a single excised cilium, having a uniform distribution of membrane channels, voltage clamped to 80 mV, was especially investigated. The axial profile of membrane voltage was that of a leaky cable. The Ca(2+) concentration profile tended to show a maximum in proximal segments, due to a preponderance of Ca(2+) inflow over Ca(2+) export at those locations. The local increase in Ca(2+) concentration activated Cl(-) channels. The resulting current caused a local drop in Cl(-) concentration, especially at the tip of the cilium and in distal segments, accompanied by a drop in ciliary K(+) concentration. In consequence, the membrane Cl(-) current was low in distal segments but stronger in proximal segments, where resupply was sufficient. The model predicts that the Cl(-) depletion will codetermine the time course of the receptor potential or current and the ciliary stimulus-response curve. In conclusion, when modeling with transduction elements presently known to participate, the ciliary geometry has large effects on ion distributions and transduction currents because ciliary ion transport is limited by axial electrodiffusion. PMID- 11259286 TI - Effect of contact time and force on monocyte adhesion to vascular endothelium. AB - In this study we examined whether monocytic cell attachment to vascular endothelium was affected by elevating shear stress at a constant shear rate. Contact time, which is inversely related to the shear rate, was fixed and viscosity elevated with dextran to increase the shear stress (and hence the net force on the cell) independently of shear rate. At a fixed contact time, tethering frequencies increased, rolling velocities decreased, and median arrest durations increased with increasing shear stress. Rolling and short arrests (< 0.2 s) were well fit by a single exponential consistent with adhesion via the formation of a single additional bond. The cell dissociation constant, k(off), increased when the shear stress was elevated at constant shear rate. Firmly adherent cells arresting for at least 0.2 s were well fit by a stochastic model involving dissociation from multiple bonds. Therefore, at a fixed contact time and increasing shear stress, bonds formed more frequently for rolling cells resulting in more short arrests, and more bonds formed for firmly arresting cells resulting in longer arrest durations. Possible mechanisms for this increased adhesion include greater monocyte deformation and/or more frequent penetration of microvilli through steric and charge barriers. PMID- 11259287 TI - Particle diameter influences adhesion under flow. AB - The diameter of circulating cells that may adhere to the vascular endothelium spans an order of magnitude from approximately 2 microm (e.g., platelets) to approximately 20 microm (e.g., a metastatic cell). Although mathematical models indicate that the adhesion exhibited by a cell will be a function of cell diameter, there have been few experimental investigations into the role of cell diameter in adhesion. Thus, in this study, we coated 5-, 10-, 15-, and 20-microm diameter microspheres with the recombinant P-selectin glycoprotein ligand-1 construct 19.ek.Fc. We compared the adhesion of the 19.ek.Fc microspheres to P selectin under in vitro flow conditions. We found that 1) at relatively high shear, the rate of attachment of the 19.ek.Fc microspheres decreased with increasing microsphere diameter whereas, at a lower shear, the rate of attachment was not affected by the microsphere diameter; 2) the shear stress required to set in motion a firmly adherent 19.ek.Fc microsphere decreased with increasing microsphere diameter; and 3) the rolling velocity of the 19.ek.Fc microspheres increased with increasing microsphere diameter. These results suggest that attachment, rolling, and firm adhesion are functions of particle diameter and provide experimental proof for theoretical models that indicate a role for cell diameter in adhesion. PMID- 11259288 TI - Traction force microscopy of migrating normal and H-ras transformed 3T3 fibroblasts. AB - Mechanical interactions between cell and substrate are involved in vital cellular functions from migration to signal transduction. A newly developed technique, traction force microscopy, makes it possible to visualize the dynamic characteristics of mechanical forces exerted by fibroblasts, including the magnitude, direction, and shear. In the present study such analysis is applied to migrating normal and transformed 3T3 cells. For normal cells, the lamellipodium provides almost all the forces for forward locomotion. A zone of high shear separates the lamellipodium from the cell body, suggesting that they are mechanically distinct entities. Timing and distribution of tractions at the leading edge bear no apparent relationship to local protrusive activities. However, changes in the pattern of traction forces often precede changes in the direction of migration. These observations suggest a frontal towing mechanism for cell migration, where dynamic traction forces at the leading edge actively pull the cell body forward. For H-ras transformed cells, pockets of weak, transient traction scatter among small pseudopods and appear to act against one another. The shear pattern suggests multiple disorganized mechanical domains. The weak, poorly coordinated traction forces, coupled with weak cell-substrate adhesions, are likely responsible for the abnormal motile behavior of H-ras transformed cells. PMID- 11259289 TI - Direct measurements of multiple adhesive alignments and unbinding trajectories between cadherin extracellular domains. AB - Direct measurements of the interactions between antiparallel, oriented monolayers of the complete extracellular region of C-cadherin demonstrate that, rather than binding in a single unique orientation, the cadherins adhere in three distinct alignments. The strongest adhesion is observed when the opposing extracellular fragments are completely interdigitated. A second adhesive alignment forms when the interdigitated proteins separate by 70 +/- 10 A. A third complex forms at a bilayer separation commensurate with the approximate overlap of cadherin extracellular domains 1 and 2 (CEC1-2). The locations of the energy minima are independent of both the surface density of bound cadherin and the stiffness of the force transducer. Using surface element integration, we show that two flat surfaces that interact through an oscillatory potential will exhibit discrete minima at the same locations in the force profile measured between hemicylinders covered with identical materials. The measured interaction profiles, therefore, reflect the relative separations at which the antiparallel proteins adhere, and are unaffected by the curvature of the underlying substrate. The successive formation and rupture of multiple protein contacts during detachment can explain the observed sluggish unbinding of cadherin monolayers. Velocity-distance profiles, obtained by quantitative video analysis of the unbinding trajectory, exhibit three velocity regimes, the transitions between which coincide with the positions of the adhesive minima. These findings suggest that cadherins undergo multiple stage unbinding, which may function to impede adhesive failure under force. PMID- 11259290 TI - Arg(615)Cys substitution in pig skeletal ryanodine receptors increases activation of single channels by a segment of the skeletal DHPR II-III loop. AB - The effect of peptides, corresponding to sequences in the skeletal muscle dihydropyridine receptor II-III loop, on Ca(2+) release from sarcoplasmic reticulum (SR) and on ryanodine receptor (RyR) calcium release channels have been compared in preparations from normal and malignant hyperthermia (MH)-susceptible pigs. Peptide A (Thr(671)-Leu(690); 36 microM) enhanced the rate of Ca(2+) release from normal SR (SR(N)) and from SR of MH-susceptible muscle (SR(MH)) by 10 +/- 3.2 nmole/mg/min and 76 +/- 9.7 nmole/mg/min, respectively. Ca (2+) release from SR(N) or SR(MH) was not increased by control peptide NB (Gly(689) Lys(708)). AS (scrambled A sequence; 36 microM) did not alter Ca (2+) release from SR(N), but increased release from SR(MH) by 29 +/- 4.9 nmoles/mg/min. RyR channels from MH-susceptible muscle (RyR(MH)) were up to about fourfold more strongly activated by peptide A (> or =1 nM) than normal RyR channels (RyR(N)) at -40 mV. Neither NB or AS activated RyR(N). RyR(MH) showed an approximately 1.8 fold increase in mean current with 30 microM AS. Inhibition at +40 mV was stronger in RyR(MH) and seen with peptide A (> or = 0.6 microM) and AS (> or = 0.6 microM), but not NB. These results show that the Arg(615)Cys substitution in RyR(MH) has multiple effects on RyRs. We speculate that enhanced DHPR activation of RyRs may contribute to increased Ca(2+) release from SR in MH-susceptible muscle. PMID- 11259291 TI - Cell volume kinetics of adherent epithelial cells measured by laser scanning reflection microscopy: determination of water permeability changes of renal principal cells. AB - The water channel aquaporin-2 (AQP2), a key component of the antidiuretic machinery in the kidney, is rapidly regulated by the antidiuretic hormone vasopressin. The hormone exerts its action by inducing a translocation of AQP2 from intracellular vesicles to the cell membrane. This step requires the elevation of intracellular cyclic AMP. We describe here a new method, laser scanning reflection microscopy (LSRM), suitable for determining cellular osmotic water permeability coefficient changes in primary cultured inner medullary collecting duct (IMCD) cells. The recording of vertical-reflection-mode x-z-scan section areas of unstained, living IMCD cells proved useful and valid for the investigation of osmotic water permeability changes. The time-dependent increases of reflection-mode x-z-scan section areas of swelling cells were fitted to a single-exponential equation. The analysis of the time constants of these processes indicates a twofold increase in osmotic water permeability of IMCD cells after treatment of the cells both with forskolin, a cyclic AMP-elevating agent, and with Clostridium difficile toxin B, an inhibitor of Rho proteins that leads to depolymerization of F-actin-containing stress fibers. This indicates that both agents lead to the functional insertion of AQP2 into the cell membrane. Thus, we have established a new functional assay for the study of the regulation of the water permeability at the cellular level. PMID- 11259292 TI - Oxidatively modified calmodulin binds to the plasma membrane Ca-ATPase in a nonproductive and conformationally disordered complex. AB - Oxidation of either Met(145) or Met(146) in wheat germ calmodulin (CaM) to methionine sulfoxide prevents the CaM-dependent activation of the plasma membrane (PM) Ca-ATPase (D. Yin, K. Kuczera, and T. C. Squier, 2000, Chem. Res. Toxicol. 13:103-110). To investigate the structural basis for the inhibition of the PM-Ca ATPase by oxidized CaM (CaM(ox)), we have used circular dichroism (CD) and fluorescence spectroscopy to resolve conformational differences within the complex between CaM and the PM-Ca-ATPase. The similar excited-state lifetime and solvent accessibility of the fluorophore N-1-pyrenyl-maleimide covalently bound to Cys(26) in unoxidized CaM and CaM(ox) indicates that the globular domains within CaM(ox) assume a native-like structure following association with the PM Ca-ATPase. However, in comparison with oxidized CaM there are increases in the 1) molar ellipticity in the CD spectrum and 2) conformational heterogeneity between the opposing globular domains for CaM(ox) bound to the CaM-binding sequence of the PM-Ca-ATPase. Furthermore, CaM(ox) binds to the PM-Ca-ATPase with high affinity at a distinct, but overlapping, site to that normally occupied by unoxidized CaM. These results suggest that alterations in binding interactions between CaM(ox) and the PM-Ca-ATPase block important structural transitions within the CaM-binding sequence of the PM-Ca-ATPase that are normally associated with enzyme activation. PMID- 11259293 TI - Localization of the extracellular end of the voltage sensor S4 in a potassium channel. AB - The opening and closing of the pore of voltage-gated ion channels is the basis for the nervous impulse. These conformational changes are triggered by the movement of an intrinsic voltage sensor, the fourth transmembrane segment, S4. The central problem of how the movement of S4 is coupled to channel opening and where S4 is located in relation to the pore is still unsolved. Here, we estimate the position of the extracellular end of S4 in the Shaker potassium channel by analyzing the electrostatic effect of introduced charges in the pore-forming motif (S5-S6). We also present a three-dimensional model for all transmembrane segments. Knowledge of this structure is essential for the attempts to understand how voltage opens these channels. PMID- 11259294 TI - Covalently linked gramicidin channels: effects of linker hydrophobicity and alkaline metals on different stereoisomers. AB - The direct role of the dioxolane group on the gating and single-channel conductance of different stereoisomers of the dioxolane-linked gramicidin A (gA) channels reconstituted in planar lipid bilayers was investigated. Four different covalently linked gA dimers were synthesized. In two of them, the linker was the conventional dioxolane described previously (SS and RR channels). Two gAs were covalently linked with a novel modified dioxolane group containing a retinal attachment (ret-SS and ret-RR gA dimers). These proteins also formed ion channels in lipid bilayers and were selective for monovalent cations. The presence of the bulky and hydrophobic retinal group immobilizes the dioxolane linker in the bilayer core preventing its rotation into the hydrophilic lumen of the pore. In 1 M HCl the gating kinetics of the SS or RR dimers were indistinguishable from their retinal counterparts; the dwell-time distributions of the open and closed states in the SS and ret-SS were basically the same. In particular, the inactivation of the RR was not prevented by the presence of the retinal group. It is concluded that neither the fast closing events in the SS or RR dimers nor the inactivation of the RR are likely to be a functional consequence of the flipping of the dioxolane inside the pore of the channel. On the other hand, the inactivation of the RR dimer was entirely eliminated when alkaline metals (Cs(+) or K(+)) were the permeating cations in the channel. In fact, the open state of the RR channel became extremely stable, and the gating characteristics of both the SS and RR channels were different from what was seen before with permeating protons. As in HCl, the presence of a retinal in the dioxolane linker did not affect the gating behavior of the SS and RR in Cs(+)- or K(+)-containing solutions. Alternative hypotheses concerning the gating of linked gA dimers are discussed. PMID- 11259295 TI - Lateral organization and domain formation in a two-component lipid membrane system. AB - The thermodynamic phase behavior and lateral lipid membrane organization of unilamellar vesicles made from mixtures of 1,2-dimyristoyl-sn-glycero-3 phosphocholine (DMPC) and 1,2 distearoyl-sn-glycero-3-phosphocholine (DSPC) were investigated by fluorescence resonance energy transfer (FRET) as a function of temperature and composition. This was done by incorporating a headgroup-labeled lipid donor (NBD-DPPE) and acceptor (N-Rh-DPPE) in low concentrations into the binary mixtures. Two instances of increased energy transfer efficiency were observed close to the phase lines in the DMPC/DSPC phase diagram. The increase in energy transfer efficiency was attributed to a differential preference of the probes for dynamic and fluctuating gel/fluid coexisting phases. This differential preference causes the probes to segregate (S. Pedersen, K. Jorgensen, T. R. Baekmark, and O. G. Mouritsen, 1996, Biophys. J. 71:554-560). The observed increases in energy transfer match with the boundaries of the DMPC/DSPC phase diagram, as measured by Fourier transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC). We propose that the two instances of probe segregation are due to the presence of DMPC-rich and DSPC-rich domains, which form a dynamic structure of gel/fluid coexisting phases at two different temperatures. Monitoring the melting profile of each lipid component independently by FTIR shows that the domain structure is formed by DMPC-rich and DSPC-rich domains rather than by pure DMPC and DSPC domains. PMID- 11259296 TI - Voltage-induced nonconductive pre-pores and metastable single pores in unmodified planar lipid bilayer. AB - Electric fields promote pore formation in both biological and model membranes. We clamped unmodified planar bilayers at 150-550 mV to monitor transient single pores for a long period of time. We observed fast transitions between different conductance levels reflecting opening and closing of metastable lipid pores. Although mean lifetime of the pores was 3 +/- 0.8 ms (250 mV), some pores remained open for up to approximately 1 s. The mean amplitude of conductance fluctuations (approximately 500 pS) was independent of voltage and close for bilayers of different area (40,000 and 10 microm(2)), indicating the local nature of the conductive defects. The distribution of pore conductance was rather broad (dispersion of approximately 250 pS). Based on the conductance value and its dependence of the ion size, the radius of the average pore was estimated as approximately 1 nm. Short bursts of conductance spikes (opening and closing of pores) were often separated by periods of background conductance. Within the same burst the conductance between spikes was indistinguishable from the background. The mean time interval between spikes in the burst was much smaller than that between adjacent bursts. These data indicate that opening and closing of lipidic pores proceed through some electrically invisible (silent) pre-pores. Similar pre pore defects and metastable conductive pores might be involved in remodeling of cell membranes in different biologically relevant processes. PMID- 11259297 TI - Structural studies of the HIV-1 accessory protein Vpu in langmuir monolayers: synchrotron X-ray reflectivity. AB - Vpu is an 81 amino acid integral membrane protein encoded by the HIV-1 genome with a N-terminal hydrophobic domain and a C-terminal hydrophilic domain. It enhances the release of virus from the infected cell and triggers degradation of the virus receptor CD4. Langmuir monolayers of mixtures of Vpu and the phospholipid 1,2-dilignoceroyl-sn-glycero-3-phosphocholine (DLgPC) at the water air interface were studied by synchrotron radiation-based x-ray reflectivity over a range of mole ratios at constant surface pressure and for several surface pressures at a maximal mole ratio of Vpu/DLgPC. Analysis of the x-ray reflectivity data by both slab model-refinement and model-independent box refinement methods firmly establish the monolayer electron density profiles. The electron density profiles as a function of increasing Vpu/DLgPC mole ratio at a constant, relatively high surface pressure indicated that the amphipathic helices of the cytoplasmic domain lie on the surface of the phospholipid headgroups and the hydrophobic transmembrane helix is oriented approximately normal to the plane of monolayer within the phospholipid hydrocarbon chain layer. At maximal Vpu/DLgPC mole ratio, the tilt of the transmembrane helix with respect to the monolayer normal decreases with increasing surface pressure and the conformation of the cytoplasmic domain varies substantially with surface pressure. PMID- 11259298 TI - Dipole potentials indicate restructuring of the membrane interface induced by gadolinium and beryllium ions. AB - The dipole component of the membrane boundary potential, phi(d), is an integral parameter that may report on the conformational state of the lipid headgroups and their hydration. In this work, we describe an experimental approach to measurements of the dipole potential changes, Deltaphi(d), and apply it in studies of Be(2+) and Gd(3+) interactions with membranes composed of phosphatidylserine (PS), phosphatidylcholine (PC), and their mixtures. Deltaphi(d) is determined as the difference between the changes of the total boundary potential, phi(b), measured by the IFC method in planar lipid membranes and the surface potential, phi(s), determined from the electrophoretic mobility of liposomes. The Gouy-Chapman-Stern formalism, combined with the condition of mass balance, well describes the ion equilibria for these high-affinity cations. For the adsorption of Be(2+) and Gd(3+) to PC membranes, and of Mg(2+) to PS membranes, the values of Deltaphi(b) and Deltaphi(s) are the same, indicative of no change of phi(d). Binding of Gd(3+) to PS-containing membranes induces changes of phi(d) of opposite signs depending on the density of ionized PS headgroups in the bilayer. At low density, the induced Deltaphi(d) is negative (-30 mV), consistent with the effect of dehydration of the surface. At maximal density (pure PS, neutral pH), adsorption of Be(2+) or Gd(3+) induces an increase of phi(d) of 35 or 140 mV, respectively. The onset of the strong positive dipole effect on PS membranes with Gd(3+) is observed near the zero charge point and correlates with a six-fold increase of membrane tension. The observed phenomena may reflect concerted reorientation of dipole moments of PS headgroups as a result of ion adsorption and lipid condensation. Their possible implications to in-vivo effects of these high-affinity ions are discussed. PMID- 11259299 TI - Rapid compression transforms interfacial monolayers of pulmonary surfactant. AB - Films of pulmonary surfactant in the lung are metastable at surface pressures well above the equilibrium spreading pressure of 45 mN/m but commonly collapse at that pressure when compressed in vitro. The studies reported here determined the effect of compression rate on the ability of monolayers containing extracted calf surfactant at 37 degrees C to maintain very high surface pressures on the continuous interface of a captive bubble. Increasing the rate from 2 A(2)/phospholipid/min (i.e., 3% of (initial area at 40 mN/m)/min) to 23%/s produced only transient increases to 48 mN/m. Above a threshold rate of 32%/s, however, surface pressures reached > 68 mN/m. After the rapid compression, static films maintained surface pressures within +/- 1 mN/m both at these maximum values and at lower pressures following expansion at < 5%/min to > or = 45 mN/m. Experiments with dimyristoyl phosphatidylcholine at 37 degrees C produced similar results. These findings indicate that compression at rates comparable to values in the lungs can transform at least some phospholipid monolayers from a form that collapses readily at the equilibrium spreading pressure to one that is metastable for prolonged periods at higher pressures. Our results also suggest that transformation of surfactant films can occur without refinement of their composition. PMID- 11259300 TI - New ordered metastable phases between the gel and subgel phases in hydrated phospholipids. AB - Formation of low-temperature ordered gel phases in several fully hydrated phosphatidylethanolamines (PEs) and phosphatidylcholines (PCs) with saturated chains as well as in dipalmitoylphosphatidylglycerol (DPPG) was observed by synchrotron x-ray diffraction, microcalorimetry, and densitometry. The diffraction patterns recorded during slow cooling show that the gel-phase chain reflection cooperatively splits into two reflections, signaling a transformation of the usual gel phase into a more ordered phase, with an orthorhombic chain packing (the Y-transition). This transition is associated with a small decrease (2-4 microl/g) or inflection of the partial specific volume. It is fully reversible with the temperature and displays in heating direction as a small (0.1 0.7 kcal/mol) endothermic event. We recorded a Y-transition in distearoyl PE, dipalmitoyl PE (DPPE), mono and dimethylated DPPE, distearoyl PC, dipalmitoyl PC, diC(15)PC, and DPPG. No such transition exists in dimyristoyl PE and dilauroyl PE where the gel L(beta) phase transforms directly into subgel L(c) phase, as well as in the unsaturated dielaidoyl PE. The PE and PC low-temperature phases denoted L(R1) and SGII, respectively, have different hydrocarbon chain packing. The SGII phase is with tilted chains, arranged in an orthorhombic lattice of two-nearest neighbor type. Except for the PCs, it was also registered in ionized DPPG. In the L(R1) phase, the chains are perpendicular to the bilayer plane and arranged in an orthorhombic lattice of four-nearest-neighbor type. It was observed in PEs and in protonated DPPG. The L(R1) and SGII phases are metastable phases, which may only be formed by cooling the respective gel L(beta) and L(beta') phases, and not by heating the subgel L(c) phase. Whenever present, they appear to represent an indispensable intermediate step in the formation of the latter phase. PMID- 11259301 TI - Fabrication of nanometer-sized protein patterns using atomic force microscopy and selective immobilization. AB - A new methodology is introduced to produce nanometer-sized protein patterns. The approach includes two main steps, nanopatterning of self-assembled monolayers using atomic force microscopy (AFM)-based nanolithography and subsequent selective immobilization of proteins on the patterned monolayers. The resulting templates and protein patterns are characterized in situ using AFM. Compared with conventional protein fabrication methods, this approach is able to produce smaller patterns with higher spatial precision. In addition, fabrication and characterization are completed in near physiological conditions. The adsorption configuration and bioreactivity of the proteins within the nanopatterns are also studied in situ. PMID- 11259302 TI - Coiled-coil unwinding at the smooth muscle myosin head-rod junction is required for optimal mechanical performance. AB - Myosin II has two heads that are joined together by an alpha-helical coiled-coil rod, which can separate in the region adjacent to the head-rod junction (Trybus, K. M. 1994. J. Biol. Chem. 269:20819-20822). To test whether this flexibility at the head-rod junction is important for the mechanical performance of myosin, we used the optical trap to measure the unitary displacements of heavy meromyosin constructs in which a stable coiled-coil sequence derived from the leucine zipper was introduced into the myosin rod. The zipper was positioned either immediately after the heads (0-hep zip) or following 15 heptads of native sequence (15-hep zip). The unitary displacement (d) decreased from d = 9.7 +/- 0.6 nm for wild type heavy meromyosin (WT HMM) to d = 0.1 +/- 0.3 nm for the 0-hep zip construct (mean +/- SE). Native values were restored in the 15-hep zip construct (d = 7.5 +/- 0.7 nm). We conclude that flexibility at the myosin head-rod junction, which is provided by an unstable coiled-coil region, is essential for optimal mechanical performance. PMID- 11259303 TI - Photolytic release of MgADP reduces rigor force in smooth muscle. AB - Photolytic release of MgADP (25-300 microM) from caged ADP in permeabilized tonic (rabbit femoral artery-Rfa) and phasic (rabbit bladder-Rbl) smooth muscle in high tension rigor state, in the absence of Ca(2+), caused an exponential decline (approximately 1.5% in Rfa and approximately 6% in Rbl) of rigor force, with the rate proportional to the liberated [MgADP]. The apparent second-order rate constant of MgADP binding was estimated as approximately 1.0 x 10(6) M(-1) s(-1) for both smooth muscles. In control experiments, designed to test the specificity of MgADP, photolysis of caged ADP in the absence of Mg(2+) did not decrease rigor force in either smooth muscle, but rigor force decreased after photolytic release of Mg(2+) in the presence of ADP. The effects of photolysis of caged ADP were similar in smooth muscles containing thiophosphorylated or non-phosphorylated regulatory myosin light chains. Stretching or releasing (within range of 0.1-1.2% of initial Ca(2+)-activated force) did not affect the rate or relative amplitude of the force decrease. The effect of additions of MgADP to rigor cross-bridges could result from rotation of the lever arm of smooth muscle myosin, but this need not imply that ADP-release is a significant force-producing step of the physiological cross-bridge cycle. PMID- 11259304 TI - The effects of exogenous calcium buffers on the systolic calcium transient in rat ventricular myocytes. AB - The aim of this work was to characterize the effects that two commonly used "caged" calcium buffers (NP-EGTA and nitr-5) have on the amplitude and time course of decay of the calcium transient. We made quantitative measurements of both free and total calcium using the measured buffering properties of the cell. Intracellular calcium concentration ([Ca(2+)](i)) was measured with fluo-3 in rat ventricular myocytes. Incorporation of the buffer NP-EGTA decreased both the amplitude and rate of decay of the caffeine response. The slowing could be quantitatively accounted for by the measured increased buffering. These effects were removed by photolysis of NP-EGTA. Similar results were obtained with nitr-5 except that the effects were not completely removed by photolysis. This was shown to be due to the persistence of a component of the increased buffering after photolysis. Both buffers decreased the amplitude of the systolic calcium transient. However, although nitr-5 produced a simple slowing of the decay, NP EGTA resulted in an initial rapid phase of decay. This rapid phase of decay is attributed to calcium binding to NP-EGTA. This work represents the first quantitative analysis of the effects that extra buffering by a fast and a slow calcium chelator may have on the calcium transient. PMID- 11259305 TI - Binding of dystrophin's tandem calponin homology domain to F-actin is modulated by actin's structure. AB - Dystrophin has been shown to be associated in cells with actin bundles. Dys-246, an N-terminal recombinant protein encoding the first 246 residues of dystrophin, includes two calponin-homology (CH) domains, and is similar to a large class of F actin cross-linking proteins including alpha-actinin, fimbrin, and spectrin. It has been shown that expression or microinjection of amino-terminal fragments of dystrophin or the closely related utrophin resulted in the localization of these protein domains to actin bundles. However, in vitro studies have failed to detect any bundling of actin by either intact dystrophin or Dys-246. We show here that the structure of F-actin can be modulated so that there are two modes of Dys-246 binding, from bundling actin filaments to only binding to single filaments. The changes in F-actin structure that allow Dys-246 to bundle filaments are induced by covalent modification of Cys-374, proteolytic cleavage of F-actin's C terminus, mutation of yeast actin's N-terminus, and different buffers. The present results suggest that F-actin's structural state can have a large influence on the nature of actin's interaction with other proteins, and these different states need to be considered when conducting in vitro assays. PMID- 11259306 TI - Entropy and heat capacity of DNA melting from temperature dependence of single molecule stretching. AB - When a single molecule of double-stranded DNA is stretched beyond its B-form contour length, the measured force shows a highly cooperative overstretching transition. We have measured the force at which this transition occurs as a function of temperature. To do this, single molecules of DNA were captured between two polystyrene beads in an optical tweezers apparatus. As the temperature of the solution surrounding a captured molecule was increased from 11 degrees C to 52 degrees C in 500 mM NaCl, the overstretching transition force decreased from 69 pN to 50 pN. This reduction is attributed to a decrease in the stability of the DNA double helix with increasing temperature. These results quantitatively agree with a model that asserts that DNA melting occurs during the overstretching transition. With this model, the data may be analyzed to obtain the change in the melting entropy DeltaS of DNA with temperature. The observed nonlinear temperature dependence of DeltaS is a result of the positive change in heat capacity of DNA upon melting, which we determine from our stretching measurements to be DeltaC(p) = 60 +/- 10 cal/mol K bp, in agreement with calorimetric measurements. PMID- 11259307 TI - DNA folding: structural and mechanical properties of the two-angle model for chromatin. AB - We present a theoretical analysis of the structural and mechanical properties of the 30-nm chromatin fiber. Our study is based on the two-angle model introduced by Woodcock et al. (Woodcock, C. L., S. A. Grigoryev, R. A. Horowitz, and N. Whitaker. 1993. Proc. Natl. Acad. Sci. USA. 90:9021-9025) that describes the chromatin fiber geometry in terms of the entry-exit angle of the nucleosomal DNA and the rotational setting of the neighboring nucleosomes with respect to each other. We analytically explore the different structures that arise from this building principle, and demonstrate that the geometry with the highest density is close to the one found in native chromatin fibers under physiological conditions. On the basis of this model we calculate mechanical properties of the fiber under stretching. We obtain expressions for the stress-strain characteristics that show good agreement with the results of recent stretching experiments (Cui, Y., and C. Bustamante. 2000. Proc. Natl. Acad. Sci. USA. 97:127-132) and computer simulations (Katritch, V., C. Bustamante, and W. K. Olson. 2000. J. Mol. Biol. 295:29-40), and which provide simple physical insights into correlations between the structural and elastic properties of chromatin. PMID- 11259308 TI - Solution structure of the SL1 RNA of the M1 double-stranded RNA virus of Saccharomyces cerevisiae. AB - The 20-nucleotide SL1 VBS RNA, 5'-GGAGACGC[GAUUC]GCGCUCC (bulged A underlined and loop bases in brackets), plays a crucial role in viral particle binding to the plus strand and packaging of the RNA. Its structure was determined by NMR spectroscopy. Structure calculations gave a precisely defined structure, with an average pairwise root mean square deviation (RMSD) of 1.28 A for the entire molecule, 0.57 A for the loop region (C8-G14), and 0.46 A for the bulge region (G4-G7, C15-C17). Base stacking continues for three nucleotides on the 5' side of the loop. The final structure contains a single hydrogen bond involving the guanine imino proton and the carbonyl O(2) of the cytosine between the nucleotides on the 5' and 3' ends of the loop, although they do not form a Watson Crick base pair. All three pyrimidine bases in the loop point toward the major groove, which implies that Cap-Pol protein may recognize the major groove of the SL1 loop region. The bulged A5 residue is stacked in the stem, but nuclear Overhauser enhancements (NOEs) suggest that A5 spends part of the time in the bulged-out conformation. The rigid conformation of the upper stem and loop regions may allow the SL1 VBS RNA to interact with Cap-Pol protein without drastically changing its own conformation. PMID- 11259309 TI - Optical measurement of transverse molecular diffusion in a microchannel. AB - Quantitative analysis of molecular diffusion is a necessity for the efficient design of most microfluidic devices as well as an important biophysical method in its own right. This study demonstrates the rapid measurement of diffusion coefficients of large and small molecules in a microfluidic device, the T-sensor, by means of conventional epifluorescence microscopy. Data were collected by monitoring the transverse flux of analyte from a sample stream into a second stream flowing alongside it. As indicated by the low Reynolds numbers of the system (< 1), flow is laminar, and molecular transport between streams occurs only by diffusion. Quantitative determinations were made by fitting data with predictions of a one-dimensional model. Analysis was made of the flow development and its effect on the distribution of diffusing analyte using a three-dimensional modeling software package. Diffusion coefficients were measured for four fluorescently labeled molecules: fluorescein-biotin, insulin, ovalbumin, and streptavidin. The resulting values differed from accepted results by an average of 2.4%. Microfluidic system parameters can be selected to achieve accurate diffusion coefficient measurements and to optimize other microfluidic devices that rely on precise transverse transport of molecules. PMID- 11259310 TI - Molecular heterogeneity of O-acetylserine sulfhydrylase by two-photon excited fluorescence fluctuation spectroscopy. AB - O-acetylserine sulfhydrylase, a homo-dimeric enzyme from Salmonella typhimurium, covalently binds one pyridoxal 5'-phosphate molecule per subunit as a fluorescent coenzyme. Different tautomers of the Schiff base between the coenzyme and lysine 41 generate structured absorption and fluorescence spectra upon one-photon excitation. We investigated the protein population heterogeneity by fluorescence correlation spectroscopy and lifetime techniques upon two-photon excitation. We sampled the fluorescence intensity from a small number of molecules (approximately 10) and analyzed the distribution of photon counts to separately determine the number and the fluorescence brightness of excited protein molecules. The changes in the average number of molecules and in the fluorescence brightness with the excitation wavelength indicate the presence of at least two fluorescent species, with two-photon excitation maxima at 660 and 800 nm. These species have been identified as the enolimine and ketoenamine tautomers of the protein-coenzyme internal aldimine. Their relative abundance is estimated to be 4:1, whereas the ratio of their two-photon cross sections is reversed with respect to the single-photon excitation case. Consistent results are obtained from the measurement of the lifetime decays, which are sensitive to the excited state heterogeneity. At least two components were detected, with lifetimes of approximately 2.5 and 0.5 ns. The lifetimes are very close to the values measured in bulk solutions upon one-photon excitation and attributed to the ketoenamine tautomer and to a dipolar species formed upon proton dissociation in the excited state. PMID- 11259311 TI - Study of the chaperoning mechanism of bovine lens alpha-crystallin, a member of the alpha-small heat shock superfamily. AB - We have studied the interaction between lysozyme, destabilized by reducing its -S S- bonds, and bovine eye lens alpha-crystallin, a member of the alpha-small heat shock protein superfamily. We have used gel filtration, photon correlation spectroscopy, and analytical ultracentrifugation to study the binding of lysozyme by alpha-crystallin at 25 degrees C and 37 degrees C. We can conclude that alpha crystallin chaperones the destabilized protein in a two-step process. First the destabilized proteins are bound by the alpha-crystallin so that nonspecific aggregation of the destabilized protein is prevented. This complex is unstable, and a reorganization and inter-particle exchange of the peptides result in stable and soluble large particles. alpha-Crystallin does not require activation by temperature for the first step of its chaperone activity as it prevents the formation of nonspecific aggregates at 25 degrees C as well as at 37 degrees C. The reorganization of the peptides, however, gives rise to smaller particles at 37 degrees C than at 25 degrees C. Indirect evidence shows that the association of several alpha-crystallin/substrate protein complexes leads to the formation of very large particles. These are responsible for the increase of the light scattering. PMID- 11259312 TI - High-sensitivity fluorescence anisotropy detection of protein-folding events: application to alpha-lactalbumin. AB - An experimental procedure has been devised to record simultaneously fluorescence intensity and fluorescence anisotropy. A photoelastic modulator on the excitation beam enables the anisotropy signal to be recorded in one pass using a single photomultiplier tube and eliminates the need for a polarizer on the emission path. In conjunction with a stopped-flow mixer, providing a time-resolved capability, this procedure was used to study the refolding of apo alpha lactalbumin following dilution from guanidinium chloride. Although the fluorescence intensity does not change detectably, the fluorescence anisotropy was found to resolve the conformational changes occurring between the initial unfolded state and the molten globule state formed either kinetically during refolding at pH 7.0 or at equilibrium at pH 2.0 (A-state). This result provides further evidence that fluorescence anisotropy is a valuable probe of protein structural transitions and that the information it provides concerning the rotational mobility of a fluorophore can be complementary to the information about the local environment provided by fluorescence intensity. PMID- 11259313 TI - Streptavidin tetramerization and 2D crystallization: a mean-field approach. AB - A mean-field theoretical approach is applied to streptavidin tetramerization and two-dimensional (2D) crystallization. This theory includes, in particular, solvent-residue interactions following the inhomogeneous Flory-Huggins model for polymers. It also takes into account residue-residue interactions by using tabulated pair interaction parameters. This theory allows one to explicitly calculate the entropy of the inhomogeneous system. We show that hydrophobic interactions are responsible for the stability of tetramerization. Within the present theory, the equilibrium distance between the two dimers is the same as that determined experimentally. The free energy of tetramerization (i.e., dissociation of the two dimers) is 50 k(B)T. Unlike tetramerization, hydrophobic interactions alone are not sufficient to stabilize the 2D crystal C(222), but solvent-mediated residue-residue interactions give the most important contribution. PMID- 11259314 TI - Trehalose effect on low temperature protein dynamics: fluctuation and relaxation phenomena. AB - We performed spectral diffusion experiments in trehalose-enriched glycerol/buffer glass on horseradish peroxidase where the heme was replaced by metal-free mesoporphyrin IX, and compared them with the respective behavior in a pure glycerol/buffer-glass (Schlichter et al., J. Chem. Phys. 2000, 112:3045-3050). Trehalose has a significant influence: spectral diffusion broadening speeds up compared to the trehalose-free glass. This speeding up is attributed to a shortening of the correlation time of the frequency fluctuations most probably by preventing water molecules from leaving the protein interior. Superimposed to the frequency fluctuation dynamics is a relaxation dynamics that manifests itself as an aging process in the spectral diffusion broadening. Although the trehalose environment speeds up the fluctuations, it does not have any influence on the relaxation. Both relaxation and fluctuations are governed by power laws in time. The respective exponents do not seem to change with the protein environment. From the spectral dynamics, the mean square displacement in conformation space can be determined. It is governed by anomalous diffusion. The associated frequency correlation time is incredibly long, demonstrating that proteins at low temperatures are truly nonergodic systems. PMID- 11259315 TI - Direct imaging of dehydrogenase activity within living cells using enzyme dependent fluorescence recovery after photobleaching (ED-FRAP). AB - Reduced nicotine adenine dinucleotide (NADH) is a key metabolite involved in cellular energy conversion and many redox reactions. We describe the use of confocal microscopy in conjunction with enzyme-dependent fluorescence recovery after photobleaching (ED-FRAP) of NADH as a topological assay of NADH generation capacity within living cardiac myocytes. Quantitative validation of this approach was performed using a dehydrogenase system, in vitro. In intact cells the NADH ED FRAP was sensitive to temperature (Q(10) of 2.5) and to dehydrogenase activation by dichloroacetate or cAMP (twofold increase for each). In addition, NADH ED-FRAP was correlated with flavin adenine dinucleotide (FAD(+)) fluorescence. These data, coupled with the cellular patterns of NADH ED-FRAP changes with dehydrogenase stimulation, suggest that NADH ED-FRAP is localized to the mitochondria. These results suggest that ED-FRAP enables measurement of regional dynamics of mitochondrial NADH production in intact cells, thus providing information regarding region-specific intracellular redox reactions and energy metabolism. PMID- 11259316 TI - Highly nonlinear photodamage in two-photon fluorescence microscopy. AB - Two-photon fluorescence excitation is being increasingly used in laser scan microscopy due to very low photodamage induced by this technique under normal operation. However, excitation intensity has to be kept low, because nonlinear photodamage sets in when laser power is increased above a certain threshold. We studied this kind of damage in bovine adrenal chromaffin cells, using two different indicators of damage: changes in resting [Ca(2+)] level and the degranulation reaction. In agreement with previous studies, we found that, for both criteria, damage is proportional to the integral (over space and time) of light intensity raised to a power approximately 2.5. Thus, widening the laser pulse shape at constant average intensity both in time and in focal volume is beneficial for avoiding this kind of damage. Both measures, of course, reduce the two-photon fluorescence excitation. However, loss of signal can be compensated by increasing excitation power, such that, at constant damaging potential, signals may be even larger with long pulses and large focal volumes, because the exponent of the power law of damage is higher (mu approximately 2.5) than that of the two photon signal (mu approximately 2). PMID- 11259317 TI - Cellular distribution and handling of liver-targeting preparations in human livers studied by a liver lobe perfusion. AB - We developed and tested a novel method for perfusing parts of human liver to study uptake and handling of drug-targeting preparations. These preparations, designed for the treatment of liver fibrosis in man, have been extensively studied in animals, but little is known about the uptake and handling by human livers. Human liver tissue was obtained from livers procured from multiorgan donors and from cirrhotic livers of patients. To assess tissue viability, perfusate glutamate-oxalacetate-transaminase (GOT), glutamate-pyruvate transaminase (GPT), and lactate dehydrogenase (LDH) levels were determined. To assess tissue functionality, the uptake of taurocholic acid and phase I and II metabolism of lidocaine and 7-hydroxycoumarin were determined. Uptake of a drug targeting preparation was studied with Dexa10-HSA, which is designed for targeting of dexamethasone to nonparenchymal cells in the liver. During a 90-min perfusion period, no elevation of either GOT, GPT, or LDH was found. Both healthy control livers and cirrhotic livers showed phase I and II drug metabolism and functional taurocholic acid uptake. Studies with Dexa10-HSA revealed that 60 min after administration, 40% of the dose had been taken up by control livers and only 5% by cirrhotic livers. In control livers, Kupffer and endothelial cells had taken up Dexa10-HSA, whereas in cirrhotic livers only Kupffer cells were responsible for the uptake. Viability parameters and liver function tests clearly showed the applicability of this method. In the perfusion set-up, we showed uptake of the drug-targeting preparation Dexa10-HSA by healthy and cirrhotic human liver tissue, although the distribution patterns differed. This demonstrates the need to study new concepts in (diseased) human tissue. PMID- 11259318 TI - Substrate inhibition kinetics for cytochrome P450-catalyzed reactions. AB - Most cytochrome P450 (P450 or CYP)-catalyzed reactions are adequately described by classical Michaelis-Menten kinetic parameters (e.g., Km and Vmax), which are usually determined by a saturation profile of velocity of product formation versus substrate concentration. In turn, these parameters may be used to predict pharmacokinetics. However, some P450 enzymes exhibit atypical or non-Michaelis Menten kinetics, due largely to substrate inhibition at higher concentrations of substrate. Although the mechanism of substrate inhibition is unknown, ignoring it and truncating the data can lead to erroneous estimates of kinetic parameters. In the present study, 13 P450 marker substrates were examined with 10 recombinant P450 proteins, and 6 were found, to varying degrees, to exhibit substrate inhibition. To understand the nature of the inhibition, a kinetic model was proposed (assuming that two binding sites exist on the enzyme) and used to fit the experimental data. The derived data indicated that 1) the K(I) values (substrate inhibition) were approximately 1.2- to 10-fold greater than the respective K(S) values; 2) both K(S) and K(I) values may be affected by the interaction of the two bound substrates within the enzyme, exhibited by a factor alpha (alpha = 5.1-23.3); and 3) enzyme activity was inhibited markedly (39-97%) at excess concentrations of the substrates (beta = 0.03-0.61). These findings suggest that substrates have access to both the inhibitory site and catalytic site simultaneously (K(I) > K(S)). Furthermore, the two sites, in the presence of substrate, can interact with each other. Therefore, the degree of inhibition of the enzyme is dependent on the concentration of the substrate (usually >K(I)) that sufficiently occupies the inhibitory site. PMID- 11259319 TI - S-naproxen-beta-1-O-acyl glucuronide degradation kinetic studies by stopped-flow high-performance liquid chromatography-1H NMR and high-performance liquid chromatography-UV. AB - Acyl-migrated isomers of drug beta-1-O-acyl glucuronides have been implicated in drug toxicity because they can bind to proteins. The acyl migration and hydrolysis of S-naproxen-beta-1-O-acyl glucuronide (S-nap-g) was followed by dynamic stopped-flow HPLC-1H NMR and HPLC methods. Nine first order rate constants in the chemical equilibrium between six species (S-nap-g, its alpha/beta-2-O-acyl, alpha/beta-3-O-acyl, alpha/beta-4-O-acyl, and alpha-1-O-acyl migration isomers, and S-naproxen aglycone) were determined by HPLC-UV studies in 25 mM potassium phosphate buffer, pH 7.40, 25 mM potassium phosphate buffer in D2O pD 7.40, and 25 mM potassium phosphate buffer in D2O pD 7.40/MeCN 80:20 v/v (HPLC-1H NMR mobile phase). In the 25 mM potassium phosphate buffer (pH 7.40) the acyl-migration rate constants (h(-1)) were 0.18 (S-nap-g-alpha/beta-2-O-acyl isomer), 0.23 (alpha/beta-2-O-acyl-alpha-1-O-acyl), 2.6 (alpha-1-O-acyl alpha/beta-2-O-acyl), 0.12 (alpha/beta-2-O-acyl-alpha/beta-3-O-acyl), 0.048 (alpha/beta-3-O-acyl-alpha/beta-2-O-acyl), 0.059 (alpha/beta-3-O-acyl-alpha/beta 4-O-acyl), and 0.085 (alpha/beta-4-O-acyl-alpha/beta-3-O-acyl). The hydrolysis rate constants (h(-1)) were 0.025 (hydrolysis of S-nap-g) and 0.0058 (hydrolysis of all acyl-migrated isomers). D2O and MeCN decreased the magnitude of all nine kinetic rate constants by up to 80%. The kinetic rate constants for the degradation of S-nap-g in the mobile phase used for HPLC-1H NMR determined using HPLC-UV could predict the results obtained by the dynamic stopped-flow HPLC-1H NMR experiments of the individual acyl-migrated isomers. It is therefore recommended that beta-1-O-acyl glucuronide degradation kinetics be investigated by HPLC-UV methods once the identification and elution order of the isomers have been established by HPLC-1H NMR. PMID- 11259320 TI - Metabolism of dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2' dicarboxylate (DDB) by human liver microsomes: characterization of metabolic pathways and of cytochrome P450 isoforms involved. AB - Metabolic fate of DDB and identification of P450 isozymes involved in the metabolism of DDB were investigated in human liver microsomes. DDB was rapidly metabolized to five different metabolites, and the structures of each metabolite were characterized based on UV, mass, and NMR spectral analyses. The major metabolic pathways of DDB in human liver microsomes were identified as O demethylation of the carboxymethyl moiety (M4) and demethylenation of the methylenedioxyphenyl group (M2). The intramolecular lactonization between the hydroxyl group at the C6 and carboxymethyl group at the C2' of M2 resulted in the generation of M5, which was either hydrolyzed to its hydrolyzed derivative (M1) or further metabolized to the O-demethylated derivative (M3). The interconversion of M1, M2, and M5 took place nonenzymatically depending on the solvent condition. M5 was predominantly detected at the acidic condition, whereas M1 was preferentially detected at the basic environment. Cytochrome P450 (P450) isoform(s) involved in the metabolism of DDB was identified using several in vitro approaches. Chemical inhibition using isoform-selective P450 inhibitors, correlation of DDB metabolites formation with several isoform-specific P450 activities in a panel of liver microsomes, metabolism by microsomes derived from P450 cDNA-expressed B-lymphoblastoid cells, and immunoinhibition by isoform specific anti-P450 antibodies collectively indicated that CYP1A2, CYP2C9, and CYP3A4 are responsible for the metabolism of DDB. O-Dealkylation of the carboxymethyl group was preferentially catalyzed by CYP1A2, whereas demethylenation of the methylenedioxyphenyl moiety was catalyzed by CYP3A4 and CYP2C9. PMID- 11259321 TI - Mechanism-based inhibition of human cytochrome P450 1A1 by rhapontigenin. AB - Recently we reported that resveratrol (trans-3,4',5-trihydroxystilbene) showed selective inhibition of recombinant human cytochrome P450 (P450) 1A1 in a concentration-dependent manner. The inhibition of recombinant human P450 1A1, 1A2, or 1B1 by various hydroxystilbene compounds having a similar structure to resveratrol was investigated using bacterial membranes from a human P450/NADPH P450 reductase bicistronic expression system to find new candidates for cancer chemopreventive agents. Of seven compounds tested, rhapontigenin (3,3',5 trihydroxy-4'-methoxystilbene) exhibited a potent and selective inhibition of human P450 1A1 with an IC50 value of 0.4 microM. Rhapontigenin showed 400-fold selectivity for P450 1A1 over P450 1A2 and 23-fold selectivity for P450 1A1 over P450 1B1. Rhapontigenin did not show any significant inhibition of ethoxyresorufin O-deethylation (EROD) activity in human liver microsomes, the other human P450s such as P450 2E1, P450 3A4, P450 2D6, P450 2C8, and P450 2C9, or human NADPH-P450 reductase. We have further investigated the inhibition kinetics of P450 1A1 by rhapontigenin. Rhapontigenin inhibited EROD activity of expressed human P450 1A1 in a competitive manner. The loss of EROD activity was time- and concentration-dependent. The values for K(i) and k(inactivation) were 0.09 microM and 0.06 min(-1), respectively. The loss was not blocked by the trapping agents glutathione, N-acetylcysteine, or dithiothreitol. These results suggest that rhapontigenin is a potent mechanism-based inactivator of human P450 1A1 and may be considered as a good candidate for a cancer chemopreventive agent in humans. PMID- 11259322 TI - Carrier-mediated hepatobiliary transport of a novel antifolate, N-[4-[(2,4 dianninopteridine-6-yl)methyl]-3,4-dihydro-2H-1,4-benzothiazin-7-yl]carbonyl-L homoglutamic acid, in rats. AB - The hepatic uptake and biliary excretion of a novel methotrexate derivative, N-[4 [(2,4-diamminopteridine-6-yl)methyl]-3,4-dihydro-2H-1,4-benzothiazin-7 yl]carbonyl-L-homoglutamic acid (MX-68), were examined in rats in vitro using isolated hepatocytes and bile canalicular membrane vesicles (CMVs), respectively. The uptake of MX-68 by isolated rat hepatocytes showed a saturable component (Km = 2.15 microM and Vmax = 2.34 pmol/min/mg of protein) and was inhibited by ATP depletors and anionic compounds such as taurocholate and probenecid. [3H]MX-68 uptake was also inhibited by folate analogs such as methotrexate and 5CH3 tetrahydrofolate, but the effect of these compounds was slightly less than that of unlabeled MX-68. On replacing Na+ with choline, MX-68 uptake remained unchanged, whereas the methotrexate uptake was reduced. Uptake of MX-68 increased as the extracellular pH fell from 7.5 to 5.5. These results suggest that MX-68 is taken up via active transport systems. The uptake of MX-68 by CMVs prepared from normal rats exhibited clear ATP dependence, whereas ATP had only a minimal effect on the uptake by CMVs from Eisai-hyperbilirubinemic rats with a hereditary deficiency in canalicular multispecific organic anion transporter (cMOAT). The initial uptake rate of ATP-dependent MX-68 transport showed saturation with kinetic parameters similar to those of methotrexate. MX-68 inhibited the ATP dependent transport of 2,4-dinitrophenyl-S-glutathione, a typical substrate for cMOAT, the inhibition constant (162 microM) being comparable with the Km of ATP dependent MX-68 transport. These results suggest that the biliary excretion of MX 68 via the bile canalicular membrane is mediated mainly by cMOAT. In conclusion, active transport systems are involved in membrane penetration of MX-68 both at sinusoidal and canalicular sides in the liver, the latter being mainly involved with methotrexate (MTX) whereas the former differs partially from that for MTX. PMID- 11259323 TI - Methemoglobin oxidation of N-acetylbenzidine to form a sulfinamide. AB - Aromatic amine sulfinamide adducts of hemoglobin are biomarkers of exposure and evidence for cytochrome P-450 N-hydroxylation. The possible peroxidatic formation of an N-acetylbenzidine (ABZ) sulfinamide adduct by methemoglobin was examined. Following addition of H2O2, 0.06 mM [3H]ABZ was metabolized by methemoglobin. With 0.3 mM glutathione, a new peak was observed, ABZ-SG, representing 17% of the total radioactivity. N'-Hydroxy-N-acetylbenzidine and 4'-nitro-4 acetylaminobiphenyl were not detected. Optimal ABZ-SG formation was observed with 3 uM methemoglobin, 0.1 to 0.3 mM glutathione, and pH 5.5. Higher concentrations of glutathione were inhibitory. Without glutathione, an H2O2-to-ABZ molar ratio of 1:1 resulted in complete metabolism of ABZ. This ratio increased to greater than 2:1 with 0.3 mM glutathione. Nearly complete inhibition of ABZ-SG formation by cyanide (10 mM), ascorbic acid (0.1 mM), 5,5-dimethyl-1-pyrroline N-oxide (50 mM), thiourea (1 mM), and azide (0.3 mM), and the lack of inhibition by mannitol (50 mM) and superoxide dismutase (2 microg) is consistent with a methemoglobin mediated peroxidatic reaction, which does not involve hydroxyl radical or superoxide. ABZ-SG was identified by electrospray ionization/mass spectrometry as N'-(glutathion-S-yl)-N-acetylbenzidine S-oxide. Conjugate was hydrolyzed by 0.1 N HCl and NaOH, was relatively stable at pH 5.5 and 7.4, and was susceptible to gamma-glutamyltranspeptidase treatment. Formation of an ABZ sulfinamide conjugate with hemoglobin was demonstrated. The results demonstrate that methemoglobin can catalyze the peroxidatic formation of an ABZ sulfinamide adduct, perhaps by a diimine monocation intermediate. PMID- 11259324 TI - In vitro glucuronidation of the cyclin-dependent kinase inhibitor flavopiridol by rat and human liver microsomes: involvement of UDP-glucuronosyltransferases 1A1 and 1A9. AB - The metabolism of flavopiridol (FLAP), a novel anticancer drug currently undergoing clinical development, was investigated in rat and human liver microsomes. In the presence of uridine 5'-diphosphoglucuronic acid, two biotransformation products (M1 and M2) could be detected. Formation of metabolite M1 and M2 in terms of enzymatic efficacy (Vmax/K(M)) was about 50- and 5-fold higher in rat (1.58 +/- 2.23 and 7.22 +/- 1.17 microl/min/mg) as compared with human liver microsomes (0.032 +/- 0.016 and 1.52 +/- 0.93 microl/min/mg), indicating species-related differences in FLAP glucuronidation. Incubation in the presence of human recombinant UDP-glucuronosyltransferases (UGTs) demonstrated that M1 is almost exclusively catalyzed by UGT1A1, whereas M2 is formed by UGT1A9 and only to a minor extent by UGT1A1 and UGT1A10. Chemical inhibition experiments further prove the involvement of UGT1A1 and UGT1A9 in the formation of M1 and M2, as the UGT1A1 substrate bilirubin preferably inhibited M1 over M2 (K(i): 36 and 258 microM, respectively), whereas the UGT1A9 substrate propofol showed a more pronounced decrease in M2 but not in M1 formation (K(i): 47 and 142 microM, respectively). Both conjugates were purified from rat liver microsomes and analyzed by mass spectrometry, NMR, and UV experiments. On the basis of these results, M1 was identified as 5-O-beta-glucopyranuronosyl-flavopiridol and M2 as 7-O-beta-glucopyranuronosyl-flavopiridol. In conclusion, our results elucidate the enzymatic pathways of FLAP in rat and human liver, which must be considered during cancer therapy of patients. PMID- 11259325 TI - Pharmacokinetics and metabolism of nateglinide in humans. AB - The pharmacokinetics and metabolism of nateglinide were studied in six healthy male subjects receiving a single oral (120 mg) and intravenous (60 mg) dose of [14C]nateglinide in randomized order. Serial blood and complete urine and feces were collected for 120 h post dose. Nateglinide was rapidly (approximately 90%) absorbed, with peak blood and plasma concentrations at approximately 1 h post dose. The maximal plasma concentrations of radioactivity (6360 ngEq/ml) and nateglinide (5690 ng/ml) were comparable, and plasma radioactivity concentrations were about twice those of blood at all times. Oral bioavailability was 72%, indicating only a modest first-pass effect. After either dose, plasma nateglinide concentrations declined rapidly with elimination half-lives of 1.5 to 1.7 h and plasma clearance of 7.4 l/h. Plasma radioactivity was eliminated more slowly with half-lives of 52 and 35 h in plasma and blood, respectively, after the oral dose. The contribution of this more slowly eliminated component to the AUC(0-infinity) was minor. Nateglinide was extensively metabolized, with excretion predominantly (84-87%) in urine. Only approximately 16% of the dose was excreted unchanged in urine after either dosing route. The major metabolites were the result of oxidative modifications of the isopropyl group. Three of these were monohydroxylated, two of which appeared to be diastereoisomers. Additionally, one metabolite with an unsaturation in the isopropyl group and two diol-containing isomers were identified. Glucuronic acid conjugates resulting from direct glucuronidation of the carboxylic acid were also present. The major metabolite in plasma and urine was the result of hydroxylation of the methine carbon of the isopropyl group. PMID- 11259326 TI - A preclinical pharmacokinetic study of the bioreductive drug AQ4N. AB - AQ4N (1,4-bis-[[2-(dimethylamino-N-oxide)ethyl]amino]5,8-dihydroxyanthracene-9,10 dione) is in a class of bioreductive agents incorporating the aliphatic N-oxide functionality and is well documented as a very effective enhancer of radiotherapy and chemotherapy. The compound is shortly to enter Phase I clinical trials in the United Kingdom, and this study describes the preclinical pharmacokinetics and metabolism of AQ4N in mice. AQ4N was administered by i.v. injection at doses of 200, 100, and 20 mg/kg and was quantified by high-performance liquid chromatography and liquid chromatography/mass spectroscopy. There was a linear increase in the maximum plasma concentration (Cmax) proportional to dose with a Cmax of 1171 microg/ml at the maximum tolerated dose of 200 mg/kg. The area under plasma concentration versus time curve (AUC) increased disproportionately with dose from 14.1 microg/h/ml at 20 mg/kg to 247 microg/h/ml at 200 mg/kg with a subsequent decrease in clearance. Terminal elimination half-lives ranged from 0.64 to 0.83 h. The spectra of the two major metabolites matched those from authentic standards with the molecular ions [M + H]+ being detected at m/z 445.4 (AQ4N), m/z 429.5 (AQ4 mono-N-oxide) and m/z 413.5 (AQ4). Only low concentrations of the toxic metabolite (AQ4) were detected in plasma at all three doses, with the AUC and Cmax at 200 mg/kg being 3.54 microg/h/ml and 3.7 microg/ml, respectively, representing <2% of AQ4N. Concentrations of the intermediate AQ4 M represented 8, 10, and 18% of those for AQ4N at the doses of 20,100, and 200 mg/kg. The concentrations necessary for a therapeutic response in vivo have been described in this pharmacokinetic study. PMID- 11259327 TI - Decreases in phenytoin hydroxylation activities catalyzed by liver microsomal cytochrome P450 enzymes in phenytoin-treated rats. AB - Phenytoin, 5,5-diphenylhydantoin, is a widely used anticonvulsant agent with a variety of toxicities, including drug interactions. The formation of four oxidative metabolites, 4'-hydroxylated (4'-HPPH), 3'-hydroxylated (3'-HPPH), a catechol (3',4'-diHPPH), and the 3',4'-dihydrodiol form of phenytoin was examined in rat liver microsomes. In 11 cDNA-expressed rat P450 enzymes tested, CYP2C6 had the highest activities in 4'- and 3'-HPPH formation from phenytoin, followed only by CYP2C11. In contrast, CYP2C11 had high activity for 3',4'-diHPPH formation from 4'-HPPH, followed by CYP2C6. The rates of 4'-HPPH and 3',4'-diHPPH formation from phenytoin in liver microsomes in the presence of NADPH were significantly decreased by oral administration of phenytoin (300 mg/kg for 20 days) to rats, despite the increase in P450 contents. However, the cumene hydroperoxide supported formation of 3',4'-dihydrodiol and 4'-HPPH from phenytoin was induced by phenytoin administration. Hydrogen peroxide formation in reaction mixtures with NADPH was induced by the administration of phenytoin; however, the coupling ratio of phenytoin oxidation was decreased in phenytoin-induced liver microsomal P450 systems. These results suggested that phenytoin could not stimulate its own apparent oxidative metabolism by liver P450s induced with phenytoin administration. The increase of unmetabolized phenytoin and byproducts of oxygen generated in the phenytoin-induced liver microsomal P450 system may be involved in phenytoin-related drug toxicity. PMID- 11259328 TI - The disposition of gemifloxacin, a new fluoroquinolone antibiotic, in rats and dogs. AB - Gemifloxacin is a fluoroquinolone antibacterial compound with enhanced affinity for bacterial topoisomerase IV and is being developed for the treatment of respiratory and urinary tract infections. The disposition and metabolic fate of this antibiotic was studied in the rat and the dog, the animal species used in its toxicological evaluation. The investigations were carried out following oral and intravenous administration of gemifloxacin mesylate. Gemifloxacin is a racemic compound; therefore, the pharmacokinetics of its individual (+) and (-) enantiomers were characterized using a chiral high-performance liquid chromatography/tandem mass spectrometry assay. In both rat and dog, the pharmacokinetic profiles of the (+) and (-) enantiomers were essentially identical. The enantiomers were rapidly absorbed following oral administration of racemic gemifloxacin mesylate. They distributed rapidly beyond total body water, and their blood clearance values were approximately equal to one quarter of the hepatic blood flow in each species. Terminal phase elimination half-lives were ca. 2 h in the rat and 5 h in the dog. Gemifloxacin was metabolized to a limited extent following oral and intravenous administration of [14C]gemifloxacin mesylate, and all metabolites formed were relatively minor. The principal metabolites formed were the E-isomer (4-6% of dose) and the acyl glucuronide of gemifloxacin (2-6% of dose) in both species and N-acetyl gemifloxacin (2-5% of dose) in the rat. Data obtained following intravenous administration indicated that gemifloxacin-related material is eliminated from the body via urinary excretion, biliary secretion, and gastrointestinal secretion. Material was eliminated approximately equally by the three routes in the dog, whereas a slightly higher proportion of the dose was eliminated in the urine (46%) and a lower proportion in the bile (12%) of rats. PMID- 11259329 TI - Prediction of midazolam-CYP3A inhibitors interaction in the human liver from in vivo/in vitro absorption, distribution, and metabolism data. AB - The extent of decreases in apparent hepatic clearance and intrinsic hepatic clearance of midazolam (MDZ) after intravenous administration of MDZ with concomitant oral administration of cimetidine (CIM), itraconazole (ITZ), or erythromycin (EM) was predicted using plasma unbound concentrations and liver unbound concentrations of inhibitors. When MDZ was concomitantly administered with CIM, the observed increase in MDZ concentration was successfully predicted using inhibition constants assessed by human liver microsome and liver-to-plasma unbound concentration ratios in rats. However, the extent of interaction with ITZ or EM was still underestimated even taking into account the concentrative uptake of inhibitors into liver. We could predict the degree of "mechanism-based" inhibition by EM on the hepatic metabolism of MDZ, after repeated administration of EM, by a physiological model incorporating the amount of active enzyme as well as the concentration of inhibitor. The maximum inactivation rate constant and the apparent inactivation constant of EM on MDZ metabolism were 0.0665 min(-1) and 81.8 microM, respectively. These kinetic parameters for the inactivation of the enzyme were applied to the physiological model with pharmacokinetic parameters of EM and MDZ obtained from published results. Consequently, we estimated that cytochrome P450 3A4 in the liver after repeated oral administration of EM was inactivated, resulting in 2.6-fold increase in the plasma concentration of MDZ. The estimated extent of increase in MDZ concentration in our study correlated well with the observed value based on metabolic inhibition by EM from published results. PMID- 11259330 TI - Uptake of teicoplanin by isolated rat hepatocytes: comparison with in vivo hepatic distribution. AB - Tissue distribution of teicoplanin, a large glycopeptide antibiotic, is slow but at equilibrium its whole body distribution volume is relatively large (Vss = 1.18 2.78 liter/kg), despite a high binding to plasma albumin. In vivo distribution into liver is extensive. Previous in vitro homogenate studies suggest that teicoplanin binds to cell membranes but only enters some cells. This possibility was investigated with isolated hepatocytes incubated for 4 h with [14C]teicoplanin alone and in the presence of additional teicoplanin (1 and 100 microg/ml). Uptake was determined after separating the cells by rapid centrifugation through a dibutyl phthalate layer and assessing viability by the trypan blue exclusion test. Teicoplanin cell uptake curves, initially rapid followed by slower distribution (which agrees with in vivo findings), were adequately described by a closed two-compartment model. Whereas entry into hepatocytes was independent of drug concentration, binding to the cell exterior membrane was concentration-dependent. The equilibrium distribution ratio (Kpu(c) +/- S.D.; 42 +/- 10) was somewhat smaller than estimated in vivo (106 +/- 9), but similar to that reported previously in vitro using liver homogenates (54 +/- 11). Also, the estimated membrane permeability-surface area product was larger in vitro than in vivo (PSu +/- S.D.; 5.5 +/- 2.9 versus 0.74 +/- 0.10 ml/min per whole liver). The most likely explanation for this difference is that in vivo only a small fraction of the total cell surface area is exposed to the perisinusoidal space, where exchange occurs. PMID- 11259331 TI - Flunitrazepam metabolism by cytochrome P450S 2C19 and 3A4. AB - We have identified CYP2C19 and CYP3A4 as the principal cytochrome P450s involved in the metabolism of flunitrazepam to its major metabolites desmethylflunitrazepam and 3-hydroxyflunitrazepam. Human CYP2C19 and CYP3A4 mediated the formation of desmethylflunitrazepam with Km values of 11.1 and 108 microM, respectively, and 3-hydroxyflunitrazepam with Km values of 642 and 34.0 microM, respectively. In human liver microsomes (n = 4) formation of both metabolites followed biphasic kinetics. Desmethylflunitrazepam formation was inhibited 31% by S-mephenytoin and 78% by ketoconazole, suggesting involvement of both CYP2C19 and CYP3A4. Formation of 3-hydroxyflunitrazepam was also significantly inhibited by ketoconazole (94%) and S-mephenytoin (18%). In support of these chemical inhibition data, antibodies directed against CYP2C19 and CYP3A4 selectively inhibited formation of desmethylflunitrazepam by 26 and 45%, respectively, while anti-CYP3A4 antibodies reduced 3-hydroxyflunitrazepam formation by 80%. Our data also suggest that CYP1A2, -2B6, -2C8, -2C9, -2D6, and 2E1 are not involved in either of these metabolic pathways. We estimate that the relative contributions of CYP2C19 and CYP3A4 to the formation of desmethylflunitrazepam in vivo are 63 and 37%, respectively, at therapeutic flunitrazepam concentrations (0.03 microM). We conclude that the polymorphic enzyme CYP2C19 importantly mediates flunitrazepam demethylation, which may alter the efficacy and safety of the drug, while CYP3A4 catalyzes the formation of 3 hydroxyflunitrazepam. PMID- 11259332 TI - Werner Kalow commemorative issue. ASPET/AACC conference: New directions in pharmacogenetics and ecogenetics, genetic defenses against environmental impacts, and responses to infections, foods, and environmental toxicants. October 3-6, 1999, Ann Arbor, Michigan, USA. PMID- 11259333 TI - Pharmacogenetics in perspective. PMID- 11259334 TI - From Bcg/Lsh/Ity to Nramp1: three decades of search and research. PMID- 11259335 TI - The sole gateway to endotoxin response: how LPS was identified as Tlr4, and its role in innate immunity. AB - Tlr4 has emerged as a specific conduit for the bacterial lipopolysaccharide (LPS) response. The fact that such a protein exists, and furthermore, the fact that it is one member of a family of proteins expressed by mononuclear cells, yields considerable insight into the mechanism by which phagocytes "see" the microbial universe. It cannot yet be assumed that all the Tlrs have specificity comparable to that of Tlr4, but it is probable that they do, given the molecular constraints to which all proteins are subject. Indeed, it is remarkable that Tlr4 is able to sense so diverse an array of LPS molecules as it does. The total number of Tlr proteins is not yet known. Although approximately 30 leucine-rich proteins bearing Toll-like cytoplasmic domains might be anticipated based on a survey of the genes in Drosophila, far fewer Toll-like genes have been found in mammals to date, although approximately 2 million expressed sequence tag sequences are now archived, and much of the genome has been covered. Some of the Toll-like proteins are, in fact, cytokine receptors. Ten leucine-rich Tlrs have been reported so far. Even a small number of receptors might be sufficient to confer recognition of most pathogens, be they fungi, bacteria, or protozoa. Some such receptors may also play developmental roles. The mutational deletion of Tlr genes alone and in combination with one another may help to establish the functions of each member of this newly emergent family of proteins. PMID- 11259336 TI - Genetics of susceptibility to infectious diseases: tuberculosis and leprosy as examples. PMID- 11259337 TI - Genetics of parasitic infections. AB - Parasites cause much suffering mainly in countries of the southern hemisphere. Hundreds of millions of individuals are infected by schistosomes, leishmanias, plasmodiums, trypanosomes, and various other parasites, and severe clinical disease occurs in a sizable fraction of the infected population causing death and severe sequelae. The outcome, asymptomatic, subclinical or clinical disease, of an infection depends mostly on the parasite and on its host. Several groups analyzing the genetics of human susceptibility to parasites have began to identify the critical steps of the pathogenic mechanisms in a few parasitic infections such as malaria and schistosomiasis. The present article, which is not meant to be an exhaustive review of the field, illustrates the progresses made in this field from pioneer studies in animals to works in endemic populations using modern strategies of human genetics. PMID- 11259338 TI - Pharmacogenetics of alcohol response and alcoholism: the interplay of genes and environmental factors in thresholds for alcoholism. AB - Recent advances in neuroscience and genetics have enabled a better understanding of genetically influenced differences in ethanol ("alcohol")-related responses and differential vulnerability to alcohol dependence at the cellular and molecular levels. Heritability studies reveal that the role of genetic factors in alcoholism is largely substance-specific, with the exception of nicotine. One focus of genetic research in alcoholism is the study of functional polymorphisms influencing alcohol metabolism, such as the aldehyde dehydrogenase type 2 Glu487Lys and alcohol dehydrogenase type 2 His47Arg polymorphisms, which affect vulnerability to alcoholism via pharmacokinetic mechanisms, and cross-population studies have begun to reveal important gene-environment interactions. The other focus is on functional genetic variants of proteins involved in the neuronal response to alcohol, including alcohol sensitivity, reward, tolerance, and withdrawal. Studies on the roles of GABA(A) alpha6-amino acid substitutions in rodents in alcohol and benzodiazepine sensitivity, and potential roles in human alcohol and benzodiazepine sensitivity are reviewed. These studies, together with recently developed knowledge on a GABA(A) receptor gene cluster at a quantitative trait loci for alcohol withdrawal on mouse chromosome 11, indicate that research investigation of variation at GABA(A) neurotransmission is a promising area in the pharmacodynamics of alcohol and in differential susceptibility to alcoholism. Genes for proteins involved in alcohol-mediated reward include genes for transporters and receptors for dopamine, serotonin, opioids, and GABA. These genes and their functional variants also represent important targets for understanding alcohol's effects in humans. Identification of genes for alcoholism vulnerability is important in the near future, not only for prevention, but also for development and targeting treatments. PMID- 11259339 TI - Genetics of iron storage and hemochromatosis. AB - The regulation of total body iron is important to all organisms. In mammals, the iron content of the body is controlled almost entirely through regulation of absorption. The precise mechanism by which iron is absorbed and the manner in which the absorption is regulated is unknown, but a number of different proteins that are involved either in the transport process itself or its regulation have been identified. These include HFE, a class 1 HLA molecule involved in hereditary hemochromatosis, the divalent metal transporter (DMT-1), hephaestin, the transferrin receptor, and mobilferrin. Iron overload occurs in a number of hereditary disorders including atransferrinemia, aceruloplasminemia, X-linked hereditary sideroblastic anemia, thalassemia major, congenital dyserythropoietic anemia, and various red cell enzyme deficiencies. In Europeans, most cases of hereditary hemochromatosis are due to mutations of the HFE gene. There are two major mutations of this gene c.845G-->A (C282Y) and c.187C-->G (H63D). These mutations have extraordinarily high prevalence in northern Europe and approximately five in a thousand Europeans are homozygous for the 845A mutation. The penetrance of even the homozygous state for the 845A mutation is very low and that for the compound heterozygote 845A/187G, which is also associated with hemochromatosis, is even lower. The reason for the markedly variable penetrance that exists in this disorder remains unknown. PMID- 11259340 TI - Molecular genetics of salt-sensitivity and hypertension. AB - For the past decade, hypertension research has shifted strongly in the direction of molecular genetics. The success stories are the monogenic hypertensive syndromes. Classic linkage analyses have located the responsible genes for glucocorticoid-remediable aldosteronism, Liddle syndrome, and apparent mineralocorticoid excess. Furthermore, a recent gain-of-function mutation has recently been described in the gene for the mineralocorticoid receptor. These genes have been cloned and their functions elucidated. Other monogenic syndromes are currently being intensively studied. However, in the area of primary hypertension, the successes have relied on the candidate gene approach. Allelic variants in the genes for angiotensinogen, alpha-adducin, the beta2-adrenergic receptor, the G-protein beta3-subunit, and the T594M mutation in the beta-subunit of the epithelial sodium channel have been identified; however, the importance of these allelic variants to primary hypertension as a whole is not yet clear. Recently, an association approach was employed to implicate the mineralocorticoid receptor gene in salt-sensitivity. Linkage approaches have been attempted and the beta-subunit of the epithelial sodium channel has been linked to hypertension and to blood pressure as a quantitative trait locus. New approaches are necessary to elucidate salt-sensitive hypertension. The analysis of multiple genes simultaneously in terms of a metabolic control analysis may provide a more promising approach. PMID- 11259341 TI - Pharmacogenetics of the vitamin D receptor and osteoporosis. AB - Osteoporosis is a major health care problem internationally with important implications for health care costs, morbidity, and mortality. Bone density, an important predictor of osteoporotic fracture risk, is affected by hormonal and environmental factors. However, in twin and family studies most of its age specific variance is genetically determined. Common allelic variations in the vitamin D receptor (VDR) gene were the first to be linked to bone density. Recently, other candidate genes, notably oestrogen receptor, collagen 1alphaI, and PTH receptor genes and a chromosome 11 locus, have been associated with bone density and fracture. Polymorphisms in adjacent regulatory regions may be important mechanisms since functional coding region mutations have not been defined. For example, the polymorphic region in the collagen 1alphaI gene alters a SpI binding site and may alter collagen gene expression. At the pharmacogenetic level, VDR alleles predict differences in gut calcium absorption and long-term bone density response to calcium intake and active vitamin D analog treatment. Understanding the mechanisms underlying these allelic differences in relation to diet and lifestyle factors as well as response to therapy could aid selection of optimal therapy for osteoporosis prevention and treatment. PMID- 11259342 TI - Intolerance to lactose and other dietary sugars. AB - Intolerance of dietary carbohydrate and sugars can result from a variety of genetically determined enzyme and transporter deficiencies. This article reviews this topic and discusses in more detail the current state of our own research on lactase. PMID- 11259343 TI - Trimethylaminuria: the fish malodor syndrome. AB - The fish malodor syndrome (also known as the fish odor syndrome and trimethylaminuria) is a metabolic disorder characterized by the presence of abnormal amounts of the dietary-derived tertiary amine, trimethylamine, in the urine, sweat, expired air, and other bodily secretions. Trimethylamine itself has the powerful aroma of rotting fish, and this confers upon the sufferer a highly objectionable body odor, which can be destructive to the personal, social, and work life of the affected individual. In recent years, much progress has been made at all levels-clinical, epidemiological, biochemical, and genetic-in our understanding of this unfortunate condition. The present article summarizes this progress, draws attention to the different types of fish malodor syndrome, and highlights the current needs in the treatment of such patients. PMID- 11259344 TI - Interactions between dietary chemicals and human sulfotransferases-molecular mechanisms and clinical significance. AB - Sulfation plays a major role in the detoxication of xenobiotics as well as in modulating the biological activity of numerous important endogenous chemicals. In contrast to this "chemical defense" function, sulfation is also a key step in the bioactivation of a host of pro-mutagens and pro-carcinogens. These reactions are catalyzed by an expanding family of sulfotransferase (SULT) enzymes, which transfer a sulfuryl moiety from the universal donor 3'-phosphoadenosine 5' phosphosulfate. Here, we discuss current knowledge of the human sulfotransferase enzyme family, of which at least 11 members have been identified to date, including regulation of expression by endogenous compounds and xenobiotics as well as the molecular basis of polymorphisms in members of the SULT1A (phenol sulfotransferase) family. We also present new data on the inhibition of SULT1A enzymes by dietary chemicals, showing that compounds to which we are exposed regularly, such as epigallocatechin gallate and epicatechin gallate are extremely potent inhibitors of phenol sulfotransferases (K(i) in the nanomolar range for SULT1A1). We found that the mechanism of inhibition by these chemicals varied depending on the individual isoform involved, showing uncompetitive inhibition of SULT1A1 whereas with SULT1A2 and -1A3 they demonstrated mixed type inhibition. Thus, genetic-environmental interactions may play an important role in modulating sulfotransferase activity and in determining individual response to chemicals metabolized by these important enzymes. PMID- 11259345 TI - Food-derived heterocyclic amine mutagens: variable metabolism and significance to humans. AB - The cooking of meat has been found to generate compounds that possess extreme mutagenicity when examined in short term tests. This observation led to the isolation and identification of a family of mutagenic chemicals, all of which are heterocyclic amines. These amines are potent bacterial and eukaryotic cell mutagens, and all of those tested have been found to induce tumors in laboratory animals. Metabolic activation of the heterocyclic amines predominantly involves CYP1-mediated N-hydroxylation and then O-esterification by phase II enzymes. In contrast, carbon oxidation, glucuronidation, and sulfation reactions at sites other than the hydroxylamine yield detoxication metabolites. In humans, the activities of these pathways are known to vary between individuals and are likely to influence susceptibility to the genetic toxicity of the heterocyclic amines. Clearly, accurate determination of human exposure to the heterocyclic amines and identification of the key enzyme systems involved and their regulation will be required for rational assessment of the risk and will help devise strategies to reduce such risk. PMID- 11259346 TI - Genetic taste markers and food preferences. AB - Sensitivity to the bitter taste of 6-n-propylthiouracil (PROP) is an inherited trait. Although some people find PROP to be extremely bitter, others cannot distinguish PROP solutions from plain water. In a series of studies, greater PROP sensitivity was linked with lower acceptability of other bitter compounds and with lower reported liking for some bitter foods. Women, identified as "super tasters" of PROP, had lower acceptance scores for grapefruit juice, green tea, Brussels sprouts, and some soy products. Many of these foods contain bitter phytochemicals with reputed cancer-protective activity. These include flavonoids in citrus fruit, polyphenols in green tea and red wine, glucosinolates in cruciferous vegetables, and isoflavones in soy products. Consumer acceptance of these plant-based foods may depend critically on inherited taste factors. This review examines the role of genetic taste markers in determining taste preferences and food choices. PMID- 11259347 TI - Food idiosyncrasies: beetroot and asparagus. AB - Anecdotal observations scattered throughout the literature have often provided clues to underlying variations in humans' ability to handle dietary chemicals. Beetroot, the red root of the garden beet used extensively as a food source, is known to produce red urine in some people following its ingestion, whereas others appear to be able to eat the vegetable with impunity. Asparagus, a vegetable whose young shoots have been eaten as a delicacy since the times of the Roman Empire, has been associated with the production of a malodorous urine smelling like rotten cabbage. Those who produce this odor assume that everyone does, and those who do not produce it have no idea of its potential olfactory consequences. These two examples, where the population appears divided in its ability to process food products or more precisely the chemicals contained within them, are reviewed in detail in this article. PMID- 11259348 TI - Identification of novel glutathione transferases and polymorphic variants by expressed sequence tag database analysis. AB - The human expressed sequence tag (EST) database can be searched by different sequence alignment strategies to identify new members of gene families and allelic variants. To illustrate the value of database analysis for gene discovery, we have focused on the glutathione S-transferase (GST) super family, an approach that has led to the identification of the Zeta class. The Zeta class GSTs catalyze the glutathione-dependent biotransformation of alpha-haloacids and the isomerization of maleylacetoacetic acid to fumarylacetoacetic acid, an essential step in the catabolism of tyrosine. Allelic variants of the GST Z1 and GST A2 genes have also been identified by EST database analysis. One GST Z1 variant (GST Z1A) has significantly higher activity with dichloroacetic acid as a substrate than other GST Z1 isoforms. This variant may be important in the clinical treatment of lactic acidosis where dichloroacetic acid is prescribed. Our experience with the application of EST database searching methods suggests that it may be productively applied to other gene families of pharmacogenetic interest. PMID- 11259349 TI - Variable CYP2A6-mediated nicotine metabolism alters smoking behavior and risk. AB - Nicotine is the psychoactive substance responsible for tobacco dependence; smokers adjust their cigarette consumption to maintain brain nicotine levels. In humans, 70 to 80% of nicotine is metabolized to the inactive metabolite cotinine by the enzyme CYP2A6. CYP2A6 can also activate tobacco smoke procarcinogens [e.g., NNK, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone]. In initial studies we found that there was an under-representation of individuals carrying defective CYP2A6 alleles in a tobacco-dependent population, and that among smokers, those with deficient nicotine metabolism smoked fewer cigarettes. We have since reproduced this data in a prospective smoking study (400 male and female, heavy and light smokers) examining the role of the CYP2A6 genotype on carbon monoxide levels, plasma and urine nicotine and cotinine levels, and cigarette counts. We have also recently identified deletion and duplication variants in the CYP2A6 gene locus and have examined their impact on smoking. These data provide the impetus to examine how inhibition of CYP2A6 activity might be useful in a therapeutic context. Both kinetic and behavioral experiments in human smokers demonstrated that inhibiting CYP2A6 in vivo decreased nicotine metabolism and smoking behavior. This article summarizes the preliminary results from our studies. PMID- 11259350 TI - Epithelial sodium channels and hypertension. AB - Hypertension is a major risk factor for heart attacks, stroke, and kidney failure. It is estimated to cause as many as 25% of all deaths in the United States, particularly for African Americans, in whom the disease is both more common and more severe. Essential hypertension is a multifactorial disorder influenced by both genetic and environmental factors. Physiological studies have shown that the kidneys play an important role in the maintenance of sodium balance, extracellular fluid volume, and long-term control of blood pressure. The sodium transporters in the kidney affect the amount of sodium and water reabsorption in the nephron and thus control extracellular fluid volume and blood pressure. Of the renal sodium transporters, the amiloride-sensitive epithelial sodium channels (ENaC), which are responsible for the rate-limiting step of sodium reabsorption in the distal nephron, are therefore important candidates in the development of hypertension. Moreover, mutations in this channel have been shown to cause a rare form of heritable hypertension (Liddle's syndrome), and genetic linkage studies show that the beta- and gamma-subunits are linked to systolic blood pressure. Several polymorphisms have been identified in the beta- and gamma-subunits of this channel, of which the beta-T594M variant is of particular interest. This variant is found in individuals of African American descent and not in Caucasians and may be associated with hypertension in some populations of African descent. Lymphocytes from individuals with this variant channel show an increased sodium conductance in response to cAMP in vitro. Studying the polymorphic variants in the various subunits of ENaC may further our understanding of the mechanisms that underlie sodium balance in mammals. These variants will provide an avenue to identify molecular targets for new diagnostic and therapeutic tools in the clinical treatment of hypertension. PMID- 11259351 TI - Genetic variation in beta-adrenergic receptors and their relationship to susceptibility for asthma and therapeutic response. AB - Our early work focused on quantitative variation in traits related to beta adrenergic signaling: lymphocyte levels of cAMP and the binding properties of the beta-adrenergic receptor. We confirmed the work of others that the major histocompatibility locus had an effect on cAMP levels in mice and in man. The latter result was presumably due to the influence of beta-adrenergic receptors, since it was the isoproterenol-stimulated lymphocyte cAMP level that was studied. We went on to directly study beta-adrenergic receptor levels in mouse hepatic plasma membranes using dihydroalprenolol binding. We discovered a strain variation, apparently controlled by a single locus, which determined a magnesium influence on measurable levels of dihydroalprenolol binding. More recently, our group's work has focused on the human single nucleotide polymorphisms (SNPs) in the human beta2-adrenergic receptor. We found that variation at amino acid position 16 had a very marked effect on the response of subjects to albuterol. Similar trends were observed for asthmatic and nonasthmatic children, and the results were independent of baseline lung function. Our results, and those of other groups relating SNPs in the beta-adrenergic receptor to a number of responses, mostly related to asthma, are reviewed in this article. PMID- 11259352 TI - ADH2 and CYP2E1 genetic polymorphisms: risk factors for alcohol-related birth defects. AB - Considerable variation in offspring outcome occurs following intrauterine ethanol exposure. The mechanism underlying this varying susceptibility may involve genetic differences in ethanol metabolism catalyzed by alcohol dehydrogenase (ADH) and cytochrome P450 2E1 (CYP2E1). A recent population study demonstrated a protective role for the ADH-beta(3) isoform, which is encoded by ADH2*3, an allele unique to African Americans. Drinking during pregnancy was associated with lower scores on the Bayley Scales of Infant Developmental Mental Index (MDI), but only in the offspring of mothers without an ADH2*3 allele. Lower MDI scores were associated with the three-way interaction among increasing ethanol intake and maternal and offspring absence of the ADH2*3 allele (p < 0.01, analysis of variance, model r(2) = 0.09). The protection afforded by this allele is likely secondary to its encoding of the high K(m), high V(max) ADH-beta3 isoenzyme, which would provide more efficient ethanol metabolism at high blood ethanol concentrations. However, the small amount of variance accounted for by the ADH2 polymorphism suggests that other genetic and/or environmental factors are also determinants of offspring risk. We recently described a 96-bp insertion polymorphism in the CYP2E1 regulatory region that is associated with enhanced CYP2E1 metabolic ability in the presence of ethanol intake or obesity, conditions associated with CYP2E1 induction (p < 0.01, both). The frequency of the insertion varies across ethnic groups, occurring in about 30% of African Americans and 7% of Caucasians (p < 0.01), and is sufficiently common to impact susceptibility to alcohol-related birth defects. Thus, genetic differences in ADH and CYP2E1 are likely determinants of offspring risk. PMID- 11259353 TI - Serum paraoxonase (PON1) isozymes: the quantitative analysis of isozymes affecting individual sensitivity to environmental chemicals. AB - In a recent study on Gulf War veterans who developed delayed neurotoxicity symptoms, we found their levels of serum paraoxonase (PON1) isozyme type Q to be significantly lower than in the control, unaffected veteran group. These results were obtained in 25 ill veterans and 20 well control subjects, of which 10 were deployed and 10 were nondeployed battalion members who remained in the United States during the Gulf War. The blood samples were also assayed for serum butyrylcholinesterase in our laboratory, and more recently in Dr. C. Broomfield's laboratory for somanase and sarinase activities. The cholinesterase activities showed no significant correlation with the PON1 isozyme levels or the severity of the clinical symptoms, but the somanase and sarinase levels ran parallel to the PON1 type Q isozyme concentrations. Although there is no direct evidence that these Gulf War veterans were directly exposed to or encountered either of these nerve gases, they may have been exposed to some environmental or chemical toxin with a similar preference for hydrolysis by the PON1 type Q isozyme. The number of subjects is relatively small, but the results should encourage other investigators to examine both the individual phenotypes and the levels of PON1 isozymes in other groups exhibiting neurological symptoms. PMID- 11259354 TI - Implications of polymorphic cytochrome p450-dependent drug metabolism for drug development. AB - The main part of human cytochrome P450-dependent drug metabolism is carried out by polymorphic enzymes that can cause abolished, quantitatively or qualitatively altered, or enhanced drug metabolism. Ultrarapid metabolism is due to stable duplication, multiduplication, or amplification of active genes. Several examples exist where subjects carrying certain alleles suffer from a lack of drug efficacy due to ultrarapid metabolism or, alternatively, adverse effects from the drug treatment due to the presence of defective alleles. The polymorphic enzymes create a problem for the drug industry because of the extensive interindividual variability in the metabolism of candidate drugs that are substrates for such enzymes. The new area for lead generation has a more preclinical emphasis and involves combinatorial chemistry in conjunction with high-throughput-based analysis of thousands of substances with respect to their absorption, metabolism, and excretion characteristics. The outcome is that companies drop substrates for polymorphic enzymes at an early stage in development, which will of course create fewer problems with polymorphic enzymes in the future. The risk is that very valuable candidates, which cannot be replaced easily, never come out on the market. The alternative, however, of using the patient's genotype as a basis for individualized drug treatment constitutes, in light of rapid methodological developments, a very feasible approach to safer and more efficient drug therapies. PMID- 11259355 TI - Genotype and severity of long QT syndrome. AB - Sudden cardiac death occurs in the United States with an incidence greater than 300,000 persons per year. The underlying cause of death is commonly considered to be due to primary or secondary arrhythmias. In cases in which no structural heart disease can be identified, the long QT syndromes (LQTS) are now commonly considered as likely causes. Multiple genes causing LQTS have been identified thus far, all encoding cardiac ion channels. These include two potassium channel alpha-subunits (KVLQT1, HERG), two potassium channel beta-subunits (minK, MiRP1), and one sodium channel gene (SCN5A). The purpose of this review is to describe the current understanding of the molecular genetics of LQTS and the resultant phenotypes. PMID- 11259356 TI - Moving toward genetic profiling in patient care: the scope and rationale of pharmacogenetic/ecogenetic investigation. PMID- 11259357 TI - Utilization of new technologies in drug trials and discovery. AB - It has become widely accepted that individual genetic variation is a prime determinant in both disease susceptibility and toxic response to therapeutic agents and xenobiotics. Emerging genetic sequence data and phenotype association studies are expected to enable disease risk prediction and guide subsequent therapeutic approaches in individual cases. However, making a good match between an individual genetic profile, disease risk prediction, and appropriate therapeutic intervention will require genotyping many polymorphic sites in large numbers of genes or single nucleotide polymorphism sites throughout the genome. Additionally, each polymorphism will have to be associated with a phenotype. Presumably, a composite phenotype may be predicted by integrating anticipated contributions from each polymorphism contributing to the complex genotype. Methods for executing such large-scale genotyping studies are rapidly evolving and becoming available. DNA microarray technology applied in hybridization-based genotyping assays is particularly well suited to respond to the accelerating pace of polymorphism discovery and the associated demand for highly parallel genotyping capability. PMID- 11259358 TI - Pharmacogenetic application in drug development and clinical trials. AB - Pharmacogenetics examines the genetic characteristics of individuals to understand variations in response to therapeutics. This approach has the potential to significantly affect the development of new medicines. The application of pharmacogenetic principles could yield significant time and resource savings within the drug development process. In preclinical drug development, pharmacogenetics could be applied to compound screening and identifying potential side effects before entering full clinical testing. Subpopulations of patients with different drug responses and underlying genetic markers could be stratified in clinical trials by analyzing their genotype. These data can improve clinical trial design and offer the possibility of optimized drug prescription based on patient genotype. Pharmacogenetics can guide the development of therapeutic interventions by identifying nonresponder patient groups. Advances in high-throughput genotyping technologies have added potential by facilitating the technical hurdles and improving drug development strategies, clinical trial design, and postmarket pharmaco-vigilance. Pharmacogenetics, thus, impacts all phases of drug development and will fundamentally change the practice of medicine in the near future. PMID- 11259359 TI - Pharmacogenetics of anticancer agents: lessons from amonafide and irinotecan. AB - Amonafide and irinotecan are anticancer drugs representative of the clinical relevance of N-acetyltransferase (NAT) and uridine diphosphate glucuronosyltransferase (UGT) polymorphisms in cancer chemotherapy, respectively. Amonafide, a substrate for the polymorphic NAT2, has an active metabolite, N acetyl-amonafide. Using caffeine as a probe, slow and rapid acetylators of amonafide were identified. Fast acetylators experienced greater myelosuppression than did slow acetylators, and a reduced dose of amonafide for fast acetylators has been recommended. A pharmacodynamic model based on acetylator phenotype, pretreatment white blood cell count, and gender has been proposed for dose individualization. The strategy adopted for amonafide is a model for future investigations in pharmacogenetics, although amonafide is no longer in clinical development. SN-38, the active metabolite of irinotecan, is glucuronidated to the inactive SN-38 glucuronide by UGT1A1, the isoform catalyzing bilirubin glucuronidation. Genetic defects in UGT1A1 determine Crigler-Najjar and Gilbert's syndromes characterized by unconjugated hyperbilirubinemia. Gilbert's syndrome often remains undiagnosed and occurs in up to 19% of individuals. Gilbert's syndrome is due to a homozygous TA insertion in the TATAA promoter of UGT1A1, leading to the mutated (TA)(7) allele. Irinotecan toxicity depends on the individual glucuronidation rate of SN-38. Decreased SN-38 glucuronidating activity has been found in livers obtained from individuals carrying the (TA)(7) allele. A phenotyping procedure for UGT1A1 has not been identified and genotyping of the UGT1A1 promoter in patients receiving irinotecan may identify patients at increased risk of toxicity. A clinical trial at the University of Chicago is ongoing to demonstrate the predictive significance of UGT1A1 genotyping for irinotecan pharmacodynamics. PMID- 11259360 TI - Thiopurine pharmacogenetics: clinical and molecular studies of thiopurine methyltransferase. AB - Thiopurine drugs are used to treat patients with neoplasia and autoimmune disease as well as transplant recipients. These agents are metabolized, in part, by S methylation catalyzed by thiopurine methyltransferase (TPMT). The discovery nearly two decades ago that levels of TPMT activity in human tissues are controlled by a common genetic polymorphism led to one of the best examples of the potential importance of pharmacogenetics for clinical medicine. Specifically, it is now known that patients with inherited very low levels of TPMT activity are at greatly increased risk for thiopurine-induced toxicity such as myelosuppression when treated with standard doses of these drugs, while subjects with very high activity may be undertreated. Furthermore, recent reports indicate that TPMT may be the target for clinically significant drug interactions and that this common genetic polymorphism might be a risk factor for the occurrence of therapy-dependent secondary leukemia. In parallel with these clinical reports, the molecular basis for the TPMT polymorphism has been determined as a result of cloning and characterization of the human TPMT cDNA and gene. Those advances led to the description and characterization of a series of single nucleotide polymorphisms that result in low levels of enzyme activity as well as a polymorphic variable number tandem repeat within the 5'-flanking region of the TPMT gene that may "modulate" level of enzyme activity. As a result of these observations, the TPMT genetic polymorphism represents a model system for the way in which basic pharmacogenetic information is developed and applied to clinical medicine. PMID- 11259361 TI - Interethnic variability in human drug responses. AB - The scientific study of interethnic differences in responses to drugs has been extant for 80 years. Many of these differences have been described at the phenotypic level, and some have been explained by genetic factors. However, it is frequently difficult to disentangle accurately the hereditary and environmental influences in phenotypic comparisons. This is where the recent developments in knowledge of the genes responsible for drug receptors are starting to make a big impact. The beta 2 adrenoceptor is described; it has three genetic polymorphisms. The different genotypes influence responses to agonists such as albuterol (Salbutamol). New gene frequency data including those for Saudi Arabians, Indians, and Africans are shown. The expanding body of knowledge about genetic (and interethnic) variability in drug receptors is likely to be important in clinical medicine. PMID- 11259362 TI - Prospects for pharmacogenetics and ecogenetics in the new millennium. AB - Genetics and genomics are certain to have a large impact in drug development and proper pharmaceutical treatment of subgroups of patients with many specific diseases. We should be able to increase the therapeutic margin for many agents. Genetic variation will also be important in refining estimates of risk from all kinds of environmental agents and in choosing more effective and more cost effective risk reduction strategies. The linkage of information about genetic variation and information about environmental, nutritional, behavioral, metabolic, medical, and healthcare factors will be necessary to interpret the variation in clinical and public health terms. However, there is a great risk that present federal and state efforts to protect confidentiality and privacy of individual genetic information may make such research infeasible. In Michigan, a Governor's Commission has sought to strike an appropriate balance. PMID- 11259363 TI - A short peptide domain of platelet factor 4 blocks angiogenic key events induced by FGF-2. AB - Platelet factor 4 (PF-4) is a CXC-chemokine with strong anti-angiogenic properties. We have shown previously that PF-4 inhibits angiogenesis by associating directly with fibroblast growth factor 2 (FGF-2), inhibiting its dimerization, and blocking FGF-2 binding to endothelial cells. We now have characterized a small peptide domain (PF-447-70) derived from the C-terminus of PF-4, which conserves anti-angiogenic effects of the parent protein. PF-447-70 inhibited internalization of 125I-FGF-2 by endothelial cells in a time-dependent manner. The peptide reduced FGF-2-stimulated cell migration to control levels in wounded monolayers of bovine capillary endothelial cells. PF-447-70 also reduced FGF-2 induced phosphorylation of MAP kinases ERK-1 and ERK-2, which are essential for migration and survival of endothelial cells. In a serum-free ex vivo angiogenesis assay, the peptide blocked microvessel outgrowth by 89%. A single amino acid substitution within PF-447-70 abolished all inhibitory activities. To simulate a real anti-angiogenic treatment situation, we administered PF-447-70 systemically to mice implanted subcutaneously with FGF-2 containing gelatin sponges with the result of sparse, scattered, and immature vessel growth. The small peptide fragment derived from the angio-inhibitory CXC-chemokine PF-4 might be used as a starting point to develop anti-angiogenic designer drugs for angiogenesis-dependent pathologies such as cancer, diabetic retinopathy, and rheumatoid arthritis. PMID- 11259364 TI - Regulation of Smad signaling by protein kinase C. AB - Cross talk between transforming growth factor b(TGF-b) serine/threonine kinase receptor signaling and tyrosine kinase receptor signaling modulates cell responsiveness to polypeptide growth factors regulating cell proliferation, differentiation, and apoptosis. Here we provide a mechanism through which Smad dependent TGF-b signaling is modulated by protein kinase C (PKC). PKC, for example, is activated downstream of tyrosine kinase receptors. We show that PKC directly phosphorylates receptor-regulated Smad proteins. This phosphorylation abrogates the ability of Smad3 to bind directly to DNA, which leads to subsequent inability to mediate transcriptional responses dependent on the direct binding of Smad3 to DNA. Interference with PKC regulation of Smad functions increased cell sensitivity to transformation by the tumor promoter phorbol 12-myristate 13 acetate (PMA). PKC-dependent phosphorylation of Smad3 was found also to be a key event in the PMA-dependent inactivation of TGF-b-stimulated cell death. Thus, PKC dependent phosphorylation of Smad3 leads to down-regulation of the growth inhibitory and apoptotic action of TGF-b. PMID- 11259365 TI - c-Fos associates with the endoplasmic reticulum and activates phospholipid metabolism. AB - c-Fos, a transcription factor that constitutes DNA-binding AP-1 complexes, regulates gene expression that promotes long-lasting cellular changes. We show that, in addition to its transcription factor activity, c-Fos regulates the metabolism of phospholipids cytoplasmically by an AP-1-independent activity. Two waves of c-Fos expression that promote subsequent waves of stimulation of 32P orthophosphate incorporation into phospholipids are evidenced in quiescent cultured fibroblasts induced to re-enter the cell cycle. The first wave of c-Fos expression peaks at 7.5 min and returns to control levels by 15 min. The second wave starts by 30 min and remains elevated at 120 min. In the first wave, the lipids that incorporate 32P are predominantly second-messenger polyphosphoinositides (PIP, PIP2, PIP3); whereas in the second wave, membrane biogenesis-related lipids (PI, PE, PA), become radioactive. Both waves of phospholipid activation depend on c-Fos expression. It is interesting that a peptide that blocks AP-1 nuclear import does not affect phospholipid activation. Immunocytochemical examination showed c-Fos immunoreactivity associated to the endoplasmic reticulum. We conclude that c-Fos, rapidly induced upon cell stimulation, associates to the endoplasmic reticulum where it first regulates the synthesis/ replenishment of phospholipids required for signal transduction pathways and subsequently regulates enzymes involved in the genesis of new membrane necessary for cell growth. PMID- 11259366 TI - Small proteoglycans in human diabetic nephropathy: discrepancy between glomerular expression and protein accumulation of decorin, biglycan, lumican, and fibromodulin. AB - Small leucine-rich proteoglycans (SLRPs), for example, decorin, biglycan, fibromodulin, and lumican, are extracellular matrix organizers and binding partners of TGF-b. Decorin is also involved in growth control and angiogenesis. Hence, these proteoglycans are likely of importance in the pathogenesis of diabetic glomerulosclerosis. In normal kidney, SLRPs were preferentially expressed in the tubulointerstitium. Weak expression occurred in the mesangial matrix. Biglycan was expressed by glomerular endothelial cells and, together with fibromodulin, by distal tubular cells and in collecting ducts. In all stages of diabetic nephropathy, there was a marked up-regulation of the proteoglycans in tubulointerstitium and glomeruli. Decorin and lumican became expressed in tubuli. However, in glomeruli, overexpression was not mirrored by local proteoglycan accumulation except in advanced nephropathy. In severe glomerulosclerosis, increased decorin concentrations were found in plasma and urine, and urinary TGF b/decorin complexes could be demonstrated indirectly. The failure to detect an increased glomerular proteoglycan quantity during the development of nephropathy could be explained by assuming that they are secreted into the mesangial matrix, but cleared via the vasculature or the urinary tract, in part as complexes with TGF-b. They could thereby counteract the vicious circle being characterized by increased TGF-b production and increased matrix deposition in diabetic nephropathy. PMID- 11259367 TI - Proteolysis-inducing factor regulates hepatic gene expression via the transcription factors NF-(kappa)B and STAT3. AB - A novel protein, proteolysis-inducing factor (PIF), has been isolated from the urine of patients with pancreatic cancer and is capable of inducing muscle proteolysis in vitro. Only adult skeletal muscle and liver exhibit substantial binding of PIF. We have investigated the effect of PIF on hepatic gene expression. Primary cultures of human hepatocytes and the human cell line HepG2 were incubated in the presence of PIF to assess its effects on hepatic transcription factors, proinflammatory cytokine production, and acute phase proteins. PIF activates both the transcription factors NF-kB and STAT3, which result in the increased production of IL-8, IL-6, and C-reactive protein and the decreased production of transferrin. The function of PIF, beyond muscle degradation, is unknown but here we show that it is involved in hepatic gene expression, and is thus likely to be involved in the proinflammatory response observed in cachexia. These results may also suggest a potential role for PIF during embryonic development. The expression of PIF peaks during the embryonic period E8 to E9, a stage that is crucial in the development of skeletal muscle and liver and during which both NF-kB and STAT3 activation can also be observed. PMID- 11259368 TI - Distinct Ca2+ thresholds determine cytochrome c release or permeability transition pore opening in brain mitochondria. AB - In diseases associated with neuronal degeneration, such as Alzheimer's or cerebral ischemia, the cytosolic Ca2+ concentration ([Ca2+]cyt) is pathologically elevated. It is still unclear, however, under which conditions Ca2+ induces either apoptotic or necrotic neuronal cell death. Studying respiration and morphology of rat brain mitochondria, we found that extramitochondrial [Ca2+] above 1 M causes reversible release of cytochrome c, a key trigger of apoptosis. This event was NO-independent but required Ca2+ influx into the mitochondrial matrix. The mitochondrial permeability transition pore (PTP), widely thought to underlie cytochrome c release, was not involved. In contrast to noncerebral tissue, only relatively high [Ca2+] (is approximately equal to 200 M) opened PTP and ruptured mitochondria. Our findings might reflect a fundamental mechanism to protect postmitotic neuronal tissue against necrotic devastation and inflammation. PMID- 11259369 TI - Protective effects of anti-C5a peptide antibodies in experimental sepsis. AB - We evaluated antibodies to different peptide regions of rat C5a in the sepsis model of cecal ligation and puncture (CLP) for their protective effects in rats. Rabbit polyclonal antibodies were developed to the following peptide regions of rat C5a: amino-terminal region (A), residues 1-16; middle region (M), residues 17 36; and the carboxyl-terminal region (C), residues 58-77. With rat neutrophils, the chemotactic activity of rat C5a was significantly inhibited by antibodies with the following rank order: anti-C > anti-M >> anti-A. In vivo, antibodies to the M and C (but not A) regions of C5a were protective in experimental sepsis, as determined by survival over a 10-day period, in a dose-dependent manner. The relative protective efficacies of anti-C5a preparations (in descending order of efficacy) were anti-C > anti-M >> anti-A. In CLP rats, a delay in infusion of antibodies, which were injected at 6 or 12 h after CLP, still resulted in significant improvement in survival rates. These in vivo and in vitro data suggest that there are optimal targets on C5a for blockade during sepsis and that delayed infusion of anti-C5a antibody until after onset of clinical evidence of sepsis still provides protective effects. PMID- 11259370 TI - Nitric oxide induces MIP-2 transcription in rat renal mesangial cells and in a rat model of glomerulonephritis. AB - Nitric oxide is a crucial mediator of several forms of glomerulonephritis. We examined the effects of NO on the mRNA expression pattern in glomerular mesangial cells by using a low-stringency reverse transcriptase-polymerase chain reaction method and detected a cDNA fragment that was induced by interleukin 1b (IL-1b) and further up-regulated by the NO donor diethylenetriamine-nitric oxide (DETA NO). Each respective cDNA fragment was found to match with the cDNAs of rat macrophage inflammatory protein 2 (MIP-2) and GRO/cytokine-induced neutrophil chemoattractant 2b (CINC-2b). Further characterization of MIP-2 regulation by Northern blot analysis confirmed an NO- and IL-1b-dependent increase in MIP-2 mRNA levels. Moreover, inhibition of IL-1b-induced endogenous NO formation by the NO-synthase (NOS) inhibitor L-NMMA markedly attenuated MIP-2 protein expression. We cloned 770 bp of the 5'-flanking region of rat MIP-2 and fused this fragment to a luciferase reporter gene. Transfection of the construct into mesangial cells resulted in a 3.5-fold increase in luciferase activity in cells treated with DETA NO when compared to controls, suggesting a transcriptional mechanism for NO induced MIP-2 expression. Deletion and mutational analysis identified critical nuclear factor (NF)-kB and NF-IL-6 binding sites required for NO regulation of MIP-2. In vivo, inhibition of NO synthesis in the Thy-1.1 model of mesangioproliferative glomerulonephritis by the specific inducible-NOS inhibitor L-NIL resulted in a marked reduction of MIP-2 mRNA expression. Furthermore, infiltration of neutrophils into the glomerulus was dramatically attenuated in L NIL-treated rats. PMID- 11259371 TI - Portal hypertensive gastric mucosa has reduced activation of MAP kinase (ERK2) in response to alcohol injury: a key to impaired healing? AB - Portal hypertensive (PHT) gastric mucosa has increased susceptibility to injury and impaired mucosal healing. Because our previous study showed that ulcer induced activation of mitogen-activated protein (MAP) kinase (ERK) plays a pivotal role in gastric mucosal healing, we investigated whether ERK activation is altered in PHT gastric mucosa following alcohol injury. We studied ERK2 phosphorylation and activity and expression of MAP kinase phosphatase-1 (MKP-1) in gastric mucosa of PHT and sham-operated (SO) normal rats both at baseline and following alcohol injury. In SO gastric mucosa, ERK2 phosphorylation and activity were significantly increased time-dependently following alcohol injury: by 221% and 137%, respectively at 24 h vs. baseline. In contrast, in PHT gastric mucosa following alcohol injury, neither ERK2 phosphorylation nor activity was increased versus baseline. In PHT gastric mucosa, MKP-1 mRNA and protein expression were increased at baseline versus SO rats and were increased further following alcohol injury with values higher by 20%-40% at each study time versus SO rats. Because ERK2 is crucial for mucosal healing, reduced ERK2 activation resulting from the overexpression of MKP-1 might be the basis for the impaired mucosal healing in PHT gastric mucosa. PMID- 11259372 TI - Molecular and in silico characterization of a promoter module and C/EBP element that mediate LPS-induced RANTES/CCL5 expression in monocytic cells. AB - The chemokine RANTES/CCL5 is a proinflammatory agent produced by a variety of tissues in response to specific stimuli. In human monocytes, RANTES/CCL5 transcription is up-regulated rapidly and transiently in response to LPS. We describe here two regions that help control LPS-driven transcription from the human RANTES/CCL5 promoter in monocytic cells. These sites were analyzed by using DNase I footprinting, transient transfection assays, site-directed mutagenesis, and EMSA. RANTES site E (R(E), -125/-99) constitutively binds C/EBP proteins in monocytic Mono Mac 6 cells. Mutation of region R(E) led to a significant (40% 50%) reduction in LPS-induced promoter reporter activity. Region R(AB) is composed of tandem kB-like elements R(A) and R(B) (-73/-34). These sites working in concert act as an LPS-responsive promoter module. R(A) constitutively binds Sp1, and Rel p50/p65 following LPS stimulation. Either factor can mediate transcriptional effects at R(A). Induced Rel p50/p50 binding to site R(B) is required for LPS regulation of RANTES/CCL5 transcription. A series of computer models based on the RANTES/CCL5 promoter were generated to represent the organization of these functional elements. The models could identify LPS regulated promoters in human, other vertebrate, and viral sequences in various databases. PMID- 11259373 TI - Human CD38 and its ligand CD31 define a unique lamina propria T lymphocyte signaling pathway. AB - CD38, a nonlineage-restricted surface glycoprotein, is an ecto-enzyme (ADP ribosyl cyclase/cADPR hydrolase/EC 3.2.2.6) that regulates cytoplasmic Ca2+ and cell-cell interactions. The molecule also delivers trans-membrane signals, despite a structural ineptitude to the scope. To reconcile these issues in a unitarian model, we compared the effects of CD38 signaling in circulating and residential T lymphocytes, the latter represented by those colonizing the intestinal lamina propria. Results are as follows: 1) LP T cells express an enzymatically active form of CD38, characterized by a modified ratio between cyclase and hydrolase functions; 2) LP T cells do not mobilize Ca2+ upon CD38 ligation, as seen in PB T cells (this condition is due to a lack in activation of PLC- g, constantly observed in PB T lymphocytes); 3) The early steps of CD38 signaling involve activation of lck, syk, and LAT; 4) Late events include synthesis and release of IL-2, IL-4, IL-5, IL-10, IFN-g and GM-CSF; 5) The uniqueness of the CD38 pathway in LP T cells is not caused by impaired interactions with the CD31 ligand. The differences observed concern the signaling machinery that CD38 exploits for its own use and not the interplay with its ligand. PMID- 11259374 TI - Protection by nitric oxide against liver inflammatory injury in animals carrying a nitric oxide synthase-2 transgene. AB - The effect of pre-existent hepatic NO synthesis on liver injury induced by lipopolysaccharide was studied in animals carrying a nitric oxide synthase-2 (NOS 2) transgene under the control of the phosphoenolpyruvate carboxykinase (PEPCK) promoter. These animals expressed NOS-2 in liver cells under fasting conditions. Lipopolysaccharide-induced liver injury in D-galactosamine-conditioned mice, which enhanced notably the effect of the endotoxin on the liver, was impaired in animals expressing NOS-2. This protection against inflammatory liver damage was dependent on NO synthesis and was caused by an inhibition of nuclear factor kB (NF-kB) activity and an impairment of the synthesis of the proinflammatory cytokines tumor necrosis factor a and interleukin 1b. These data indicate that intrahepatic synthesis of NO protects liver by inhibiting the release of cascades of proinflammatory mediators and suggest a beneficial role for local delivery of NO in the control of liver injury. PMID- 11259375 TI - High-efficiency electrotransfection of human primary hematopoietic stem cells. AB - A major obstacle to gene transfer into hematopoietic stem cells, a key step for many gene therapy and tissue replacement applications, is its low efficiency. High cell mortality is responsible for the low efficiency of electrotransfection when this technique is applied to certain 'refractory' cell types such as hematopoietic stem cells. Using human primary CD-34+ cells from peripheral blood as a model, we found that transfection-induced apoptosis and, to a lesser extent, postpulse colloidal-osmotic swelling are two main factors for the poor transfection of these cells. By applying caspase inhibitors (B-D-Fluomethyl Ketone and Z-VAD-FMK) to reduce apoptosis, and by using the postpulse pelleting method to suppress colloidal-osmotic swelling, we achieved a transfection efficiency of ~20%, regardless of the presence of cytokines in the suspension medium. This effort brings the ex vivo electrotransfection efficiency within the reach of therapeutic applications. PMID- 11259376 TI - Pro-apoptotic function of calsenilin/DREAM/KChIP3. AB - Apoptotic cell death and increased production of amyloid b peptide (Ab) are pathological features of Alzheimer's disease (AD), although the exact contribution of apoptosis to the pathogenesis of the disease remains unclear. Here we describe a novel pro-apoptotic function of calsenilin/DREAM/KChIP3. By antisense oligonucleotide-induced inhibition of calsenilin/DREAM/KChIP3 synthesis, apoptosis induced by Fas, Ca2+-ionophore, or thapsigargin is attenuated. Conversely, calsenilin/DREAM/KChIP3 expression induced the morphological and biochemical features of apoptosis, including cell shrinkage, DNA laddering, and caspase activation. Calsenilin/DREAM/KChIP3-induced apoptosis was suppressed by caspase inhibitor Z-VAD and by Bcl-XL, and was potentiated by increasing cytosolic Ca2+, expression of Swedish amyloid precursor protein mutant (APPSW) or presenilin 2 (PS2), but not by a PS2 deletion lacking its C-terminus (PS2/411stop). In addition, calsenilin/DREAM/KChIP3 expression increased Ab42 production in cells expressing APPsw, which was potentiated by PS2, but not by PS2/411stop, which suggests a role for apoptosis-associated Ab42 production of calsenilin/DREAM/KChIP3. PMID- 11259377 TI - Induction of apoptosis in cervical carcinoma cells by peptide aptamers that bind to the HPV-16 E7 oncoprotein. AB - Human papillomaviruses (HPV) of the high-risk type are causally involved in human tumors, in particular cervical carcinoma. Expression of the viral oncogenes E6 and E7 is maintained in HPV-positive tumors, and it was shown that E6 and E7 of HPV-16 can immortalize human keratinocytes, the natural host cells of the virus. Expression of the viral genes is also required for maintenance of the transformed phenotype. The oncogenic activity of the E6 and E7 oncoproteins is mediated by their physical and functional interaction with cellular regulatory proteins. To knock out the function of the E7 protein in living cells, we have developed peptide aptamers with high specific binding activity for the E7 protein of HPV 16. We show here that E7-binding peptide aptamers induce programmed cell death (apoptosis) in E7-expressing cells, whereas E7-negative cells are not affected. Furthermore, E7-binding peptide aptamers induce apoptosis in HPV-16-positive tumor cells derived from cervical carcinoma. The data suggest that E7-binding peptide aptamers may be useful tools to specifically eliminate HPV-positive tumors. PMID- 11259378 TI - Inverse, protean, and ligand-selective agonism: matters of receptor conformation. AB - Concepts regarding the mechanisms by which drugs activate receptors to produce physiological response have progressed beyond considering the receptor as a simple on-off switch. Current evidence suggests that the idea that agonists produce only varying degrees of receptor activation is obsolete and must be reconciled with data to show that agonist efficacy has texture as well as magnitude. Thus, agonists can block system constitutive response (inverse agonists), behave as positive and inverse agonists on the same receptor (protean agonists), and differ in the stimulus pattern they produce in physiological systems (ligand-selective agonists). The molecular mechanism for this seemingly diverse array of activities is the same, namely, the selective microaffinity of ligands for different conformational states of the receptor. This paper reviews evidence for the existence of the various types of agonism and the potential therapeutic utility of different agonist types.-Kenakin, T. Inverse, protean, and ligand-selective agonism: matters of receptor conformation. PMID- 11259379 TI - Reactive oxygen-dependent production of novel photochemotherapeutic agents. AB - The reactive nature of species derived from oxygen, such as singlet oxygen and hydrogen peroxide, has been exploited in the clinical setting for targeting bacteria, viruses, and tumor cells by photodynamic excitation of a variety of chromophores. This modality, termed photodynamic therapy (PDT), is currently being used to treat some forms of cancer. However, the applicability of conventional PDT is limited due to the absolute dependence on simultaneous exposure of the target to the photoactive compound and light. In 1990, we demonstrated that the need for simultaneous exposure of the biological target to light and photosensitizer could be circumvented by prior exposure (activation) of the sensitizer molecule to light and its subsequent use as any other anti-cancer or anti-viral drug. By dint of the nature of the protocol, this process was termed preactivation. Since then, the generation of biologically active molecules in vitro by preactivation has been validated using a variety of chromophores, such as merocyanine 540, Photofrin II, and naphthalimide. Here we briefly review the role of reactive oxygen species in the photodynamic effect, and provide an explanation for the mechanism of preactivation. We propose that photo-oxidation not only provides a novel means for the generation of biologically active molecules, but could also explain, at least in part the mechanism of conventional PDT. It is likely that the light-dependent breakdown of the chromophore to generate novel active compounds, in addition to reactive oxygen species, also contributes to the photodynamic damage observed on simultaneous exposure of the chromophore and target tissue to light during PDT.-Pervaiz, S. Reactive oxygen dependent production of novel photochemotherapeutic agents. PMID- 11259380 TI - Endothelin-1 protects astrocytes from hypoxic/ischemic injury. AB - Under pathological conditions such as ischemia (I), subarachnoid hemorrhage, and Alzheimer's disease, astrocytes show a large increase in endothelin (ET) -like immunoreactivity. However, it is not clear whether ET is protective or destructive to these cells during brain injury. Using astrocytes from ET-1 deficient mice, we determined the effect of ET-1 on these cells under normal, hypoxic (H), and hypoxic/ischemic (H/I) conditions. Under normal culture conditions, astrocytes from wild-type and ET-1-deficient mice showed no difference in their morphology and cell proliferation rates. ET-3 and ETA receptor mRNAs were up-regulated whereas ETB receptor mRNA was down-regulated in ET-1-deficient astrocytes, suggesting that ET-1 and ET-3 may complement each other's functions and that the expressions of these endothelins and their receptors are regulated by a complex feedback mechanism. Under H and H/I conditions, ET-1 peptide and mRNA were up-regulated in wild-type astrocytes, and the astrocytes without ET-1 died faster than the wild-type astrocytes, as indicated by greater efflux of lactate dehydrogenase. The present study suggests that astrocytes without ET-1 are more vulnerable to H and H/I injuries and that the up-regulation of astrocytic ET-1 is essential for the survival of astrocytes. PMID- 11259381 TI - Longevity and heavy metal resistance in daf-2 and age-1 long-lived mutants of Caenorhabditis elegans. AB - In the nematode Caenorhabditis elegans, dauer formation, stress resistance, and longevity are determined in part by DAF-2 (insulin receptor-like protein), AGE-1 (phosphatidylinositol-3-OH kinase catalytic subunit), and DAF-16 (forkhead transcription factor). Mutations in daf-2 and age-1 result in increased resistance to heat, oxidants, and UV. We have discovered that daf-2 and age-1 mutations result in increased Cd and Cu ion resistance in a 24 h toxicity assay. Lethal concentration (LC50) values for Cd and Cu ions in daf-2 and age-1 mutants were significantly (P<0.001) higher than in wild-type nematodes. However, LC50 values in daf-16;age-1 mutants were not significantly different, implying that metal resistance is influenced by a DAF-16-related function. As metallothionein (MT) proteins play a major role in metal detoxification, we examined the expression of MT genes both under noninducing conditions and after exposure to sublethal and acute heavy metal stress. MT1 mRNA levels were significantly (P<0.05) higher in daf-2 mutants compared to age-1 mutants and wild-type C. elegans under basal conditions. After 10 mM Cd treatment, induction of MT1 and MT2 mRNA was three- and twofold higher, respectively, in daf-2 mutant worms than in wild-type. However, a sublethal concentration of Cd (0.1 mM) resulted in even higher (three- to sevenfold) levels of both MT mRNAs in all strains. Cu did not induce MT1 or MT2 mRNAs. These results are consistent with a model in which the insulin-signaling pathway determines life span through regulation of stress protein genes PMID- 11259382 TI - Direct influence of the p53 tumor suppressor on mitochondrial biogenesis and function. AB - Mitochondrial localization of p53 has been observed in several cell systems, but an understanding of its organelle-based physiological activity remains incomplete. The purpose of the present study was to investigate the mitochondrial DNA genomic response to dominant-negative p53 mutant miniprotein (p53DD) fused to a mitochondrial import signal. Constructs were generated to express mitochondrial targeted enhanced green fluorescent protein (mEGFP) or dominant-negative mutant p53 miniprotein (m53DD) by in-frame fusion to the signal peptide sequence of murine Cox8l. Control cytosolic vectors (cEGFP, c53DD) had the signal sequence placed in antisense orientation. NIH 3T3 cells were transiently transfected with these vectors in various combinations. Mitochondrial 16S ribosomal RNA (16S rRNA) expression and fluorochrome staining with Mitotracker Red CMXRos (DeltaPsim) were decreased in cells expressing m53DD. Both alterations were specific for mitochondrial import competence (e.g., m53DD vs. c53DD) as well as the passenger protein (e.g., m53DD vs. mEGFP). The normal functional state of mitochondria was restored with PK11195, a specific ligand of the mitochondrial peripheral-type benzodiazepine receptor. Negative dominance of m53DD on 16S rRNA expression and CMXRos staining, and rescue of these parameters with PK11195, imply a direct positive effect of p53 on mitochondrial biogenesis and function. PMID- 11259383 TI - SCF/c-kit signaling is required for cyclic regeneration of the hair pigmentation unit. AB - Hair graying, an age-associated process of unknown etiology, is characterized by a reduced number and activity of hair follicle (HF) melanocytes. Stem cell factor (SCF) and its receptor c-kit are important for melanocyte survival during development, and mutations in these genes result in unpigmented hairs. Here we show that during cyclic HF regeneration in C57BL/6 mice, proliferating, differentiating, and melanin-producing melanocytes express c-kit, whereas presumptive melanocyte precursors do not. SCF overexpression in HF epithelium significantly increases the number and proliferative activity of melanocytes. During the induced hair cycle in C57BL/6 mice, administration of anti-c-kit antibody dose-dependently decreases hair pigmentation and leads to partially depigmented (gray) or fully depigmented (white) hairs, associated with significant decreases in melanocyte proliferation and differentiation, as determined by immunostaining and confocal microscopy. However, in the next hair cycle, the previously treated animals grow fully pigmented hairs with the normal number and distribution of melanocytes. This suggests that melanocyte stem cells are not dependent on SCF/c-kit and when appropriately stimulated can generate melanogenically active melanocytes. Therefore, the blockade of c-kit signaling offers a fully reversible model for hair depigmentation, which might be used for the studies of hair pigmentation disorders. PMID- 11259384 TI - Calcium-magnesium interactions in pancreatic acinar cells. AB - Although the role of calcium (Ca2+) in the signal transduction and pathobiology of the exocrine pancreas is firmly established, the role of magnesium (Mg2+) remains unclear. We have characterized the intracellular distribution of Mg2+ in response to hormone stimulation in isolated mouse pancreatic acinar cells and studied the role of Mg2+ in modulating Ca2+ signaling using microspectrofluorometry and digital imaging of Ca2+- or Mg2+-sensitive fluorescent dyes as well as Mg2+-sensitive intracellular microelectrodes. Our results indicate that an increase in intracellular Mg2+ concentrations reduced the cholecystokinin (CCK) -induced Ca2+ oscillations by inhibiting the capacitive Ca2+ influx. An intracellular Ca2+ mobilization, on the other hand, was paralleled by a decrease in [Mg2+]i, which was reversible upon hormone withdrawal independent of the electrochemical gradients for Mg2+, Ca2+, Na+, and K+, and not caused by Mg2+ efflux from acinar cells. In an attempt to characterize possible Mg2+ stores that would explain the reversible, hormone-induced intracellular Mg2+ movements, we ruled out mitochondria or ATP as potential Mg2+ buffers and found that the CCK-induced [Mg2+]i decrease was initiated at the basolateral part of the acinar cells, where most of the endoplasmic reticulum (ER) is located, and progressed from there toward the apical pole of the acinar cells in an antiparallel fashion to Ca2+ waves. These experiments represent the first characterization of intracellular Mg2+ movements in the exocrine pancreas, provide evidence for possible Mg2+ stores in the ER, and indicate that the spatial and temporal distribution of intracellular Mg concentrations profoundly affects acinar cell Ca2+ signaling. PMID- 11259385 TI - A low-molecular-weight fraction of human seminal plasma activates adenylyl cyclase and induces caspase 3-independent apoptosis in prostatic epithelial cells by decreasing mitochondrial potential and Bcl-2/Bax ratio. AB - The majority of elderly men are affected by benign and malign diseases of the prostate that are governed by endocrine factors and local stromal/epithelial and luminal/epithelial interactions. Prostate epithelial cells secrete numerous factors into the seminal plasma (SMP) that are thought to be responsible for nutrition, accurate pH, and ionic environment of sperm. Our hypothesis assumes that prostatic factors responsible for optimal fertility might have retrograde influences on epithelial cell growth, differentiation, and function. SMP was analyzed for proteins and other biologically active substances by size exclusion high-performance liquid chromatography. Each fraction was investigated for its effect on cell growth and death. A low molecular mass fraction (2-4 kDa) was responsible for inducing apoptosis in proliferating prostate epithelial cells. Signal transduction was mediated by the production of cAMP; no significant changes in tyrosine phosphorylation of membrane receptors were observed. Mechanisms of apoptosis, i.e., caspase- and mitochondria-dependent pathways, were investigated in prostate epithelial cells by caspase activity assays, annexin/propidium iodide staining, changes in mitochondrial potential, p53, Par 4, Bax, and Bcl-2 protein levels. SMP induced p53- and Bcl-2-dependent apoptosis without activation of caspase-3. Obviously, SMP contains protective factors that help eliminate degenerated cells and control epithelial renewal. Age-related changes in the composition of SMP or the susceptibility of epithelial cells might, therefore, contribute to proliferative prostatic diseases PMID- 11259386 TI - Characterization of control and immobilized skeletal muscle: an overview from genetic engineering. AB - To elucidate the molecular basis of muscle atrophy, we have performed the serial analysis of gene expression (SAGE) method with control and immobilized muscles of 10 rats. The genes that expressed >0.5% in muscle are involved in the following three functions: 1) contraction (troponin I, C and T; myosin light chain 1-3; actin; tropomyosin; and parvalbumin), 2) energy metabolism (cytochrome c oxidase I and III, creatine kinase, glyceraldehyde-3-phosphate-dehydrogenase, phosphoglycerate mutase, ATPase 6, and aldolase A), and 3) housekeeping (lens epithelial protein). Muscle atrophy appears to be caused by changes in mRNA levels of specific regulators of proteolysis, protein synthesis, and contractile apparatus assembling, such as polyubiquitin, elongation factor 2, and nebulin. Immobilization has produced a decrease more than threefold in gene expression of enzymes involved in energy metabolism, especially ATPase, cytochrome c oxidase, NADH dehydrogenase, and protein phosphatase 1. Differential gene expressions of selenoprotein W and uroporphyrinogen decarboxylase, which can be involved in oxidative stress, were also observed. Other genes with various functions, such as cholesterol metabolism and growth factors, were also differentially expressed. Moreover, novel genes regulated by immobilization were discovered. Thus, the current study allows a better understanding of global muscle characteristics and the molecular mechanisms of sedentarity and sarcopenia. PMID- 11259387 TI - Gene expression profiles of proliferating vs. G1/G0 arrested human leukemia cells suggest a mechanism for glucocorticoid-induced apoptosis. AB - Glucocorticoids (GC) have pronounced effects on metabolism, differentiation, proliferation, and cell survival (1). In certain lymphocytes and lymphocyte related malignancies, GC inhibit proliferation and induce apoptotic cell death, which has led to their extensive use in the therapy of malignant lymphoproliferative disorders (2). Most of these effects result from regulation of gene expression via the GC receptor (GR), a ligand-activated transcription factor (3). Although hundreds of genes are regulated by GC (1), how certain biological GC effects relate to individual gene regulation remains enigmatic. To address this question with respect to GC-induced cell cycle arrest and apoptosis, we applied DNA chip technology (4, 5) to determine gene expression profiles in proliferating and G1/G0-arrested (by conditional expression of the CDK inhibitor p16/INK4a) acute lymphoblastic T cells undergoing GC-induced apoptosis. Of 7074 genes tested, 163 were found to be regulated by dexamethasone in the first 8 h in proliferating cells and 66 genes in G1/G0-arrested cells. An almost nonoverlapping set of genes (i.e., only eight genes) was coordinately regulated in proliferating and arrested cells. Analysis of the regulated genes supports the concept that GC-induced apoptosis results from positive GR autoregulation entailing persistent down-regulation of metabolic pathways critical for survival PMID- 11259388 TI - Oxidative stress in the aging rat heart is reversed by dietary supplementation with (R)-(alpha)-lipoic acid. AB - Oxidative stress has been implicated as a causal factor in the aging process of the heart and other tissues. To determine the extent of age-related myocardial oxidative stress, oxidant production, antioxidant status, and oxidative DNA damage were measured in hearts of young (2 months) and old (28 months) male Fischer 344 rats. Cardiac myocytes isolated from old rats showed a nearly threefold increase in the rate of oxidant production compared to young rats, as measured by the rates of 2,7-dichlorofluorescin diacetate oxidation. Determination of myocardial antioxidant status revealed a significant twofold decline in the levels of ascorbic acid (P = 0.03), but not alpha-tocopherol. A significant age-related increase (P = 0.05) in steady-state levels of oxidative DNA damage was observed, as monitored by 8-oxo-2'-deoxyguanosine levels. To investigate whether dietary supplementation with (R)-alpha-lipoic acid (LA) was effective at reducing oxidative stress, young and old rats were fed an AIN-93M diet with or without 0.2% (w/w) LA for 2 wk before death. Cardiac myocytes from old, LA-supplemented rats exhibited a markedly lower rate of oxidant production that was no longer significantly different from that in cells from unsupplemented, young rats. Lipoic acid supplementation also restored myocardial ascorbic acid levels and reduced oxidative DNA damage. Our data indicate that the aging rat heart is under increased mitochondrial-induced oxidative stress, which is significantly attenuated by lipoic acid supplementation. PMID- 11259389 TI - Paradoxical actions of exogenous and endogenous parathyroid hormone-related protein on renal vascular smooth muscle cell proliferation: reversion in the SHR model of genetic hypertension. AB - In previous studies, added parathyroid hormone-related protein (PTHrP) inhibits whereas transfected PTHrP stimulates the proliferation of A10 aortic smooth muscle cells by nuclear translocation of the peptide. In the present studies, we asked whether these paradoxical trophic actions of PTHrP occur in smooth muscle cells (SMC) cultured from small intrarenal arteries of, and whether they are altered in, 12-wk-old spontaneously hypertensive rats (SHR) as compared to normotensive Wistar-Kyoto (WKY) rats. SHR cells grew faster than WKY cells. PTHrP transcript was increased in SHR-derived cells whereas PTH1 receptor (PTH1R) transcripts were similar in both cell lines. In both strains of cells, stable transfection with human PTHrP(1-139) cDNA did not further induce proliferation, suggesting maximal effect of endogenous PTHrP in wild cells. In contrast, transfection with antisense hPTHrP(1-139) cDNA, which abolished PTHrP mRNA, decreased WKY but increased SHR cell proliferation. Added PTHrP(1-36) (1-100 pM) decreased WKY and increased SHR cell proliferation. Additional studies indicated that the preferential coupling of PTH1-R to G-protein Gi was responsible for the proliferative effect of exogenous PTHrP in SHR cells. Moreover, PTHrP was detected in the nucleolus of a fraction of WKY and SHR renal SMC, in vitro as well as in situ, suggesting that the nucleolar translocation of PTHrP might be involved in the proliferative effects of endogenous PTHrP. In renovascular SMC, added PTHrP is antimitogenic, whereas endogenously produced PTHrP is mitogenic. These paradoxical effects of PTHrP on renovascular SMC proliferation appear to be reversed in the SHR model of genetic hypertension. A new concept emerges from these results, according to which a single molecule may have opposite effects on VSMC proliferation under physiological and pathophysiological conditions. PMID- 11259390 TI - Ceramide glycosylation potentiates cellular multidrug resistance. AB - Ceramide glycosylation, through glucosylceramide synthase (GCS), allows cellular escape from ceramide-induced programmed cell death. This glycosylation event confers cancer cell resistance to cytotoxic anticancer agents [Liu, Y. Y., Han, T. Y., Giuliano, A. E., and M. C. Cabot. (1999) J. Biol. Chem. 274, 1140-1146]. We previously found that glucosylceramide, the glycosylated form of ceramide, accumulates in adriamycin-resistant breast carcinoma cells, in vinblastine resistant epithelioid carcinoma cells, and in tumor specimens from patients showing poor response to chemotherapy. Here we show that multidrug resistance can be increased over baseline and then totally reversed in human breast cancer cells by GCS gene targeting. In adriamycin-resistant MCF-7-AdrR cells, transfection of GCS upgraded multidrug resistance, whereas transfection of GCS antisense markedly restored cellular sensitivity to anthracyclines, Vinca alkaloids, taxanes, and other anticancer drugs. Sensitivity to the various drugs by GCS antisense transfection increased 7- to 240-fold and was consistent with the resumption of ceramide-caspase-apoptotic signaling. GCS targeting had little influence on cellular sensitivity to either 5-FU or cisplatin, nor did it modify P glycoprotein expression or rhodamine-123 efflux. GCS antisense transfection did enhance rhodamine-123 uptake compared with parent MCF-7-AdrR cells. This study reveals that GCS is a novel mechanism of multidrug resistance and positions GCS antisense as an innovative force to overcome multidrug resistance in cancer chemotherapy. PMID- 11259391 TI - Apoptosis and adaptive responses to oxidative stress in human endothelial cells exposed to cyclosporin A correlate with BCL-2 expression levels. AB - Treatment of transplanted patients with cyclosporin A (CSA) may cause adverse effects such as nephrotoxicity and hypertension. As CSA is known to induce oxidative stress in several tissues, it may cause vascular problems by triggering oxidative stress in endothelial cells (EC). However, oxidative stress has been reported for acute exposure to CSA concentrations exceeding its clinical range, whereas immunosuppression requires life-long treatment with therapeutic concentrations. We therefore compared the effects of 21 h pharmacological (200 microM) vs. 8 days clinical (0.5-2.5 microM) doses of CSA on cultured human EC. Pharmacological doses of CSA cause a decrease in cell density via apoptosis and a down-regulation of the antiapoptotic protein Bcl-2. However, these effects are independent of CSA-induced oxidative stress. In contrast, therapeutic concentrations of CSA cause Bcl-2 up-regulation and modification of EC morphology, both effects blocked by antioxidants. Therefore, a low level of oxidants may act in EC as second messengers that up-regulate Bcl-2, thus promoting survival of impaired EC. Our data suggest that the oxidative stress induced by clinical concentrations of CSA may be involved in the adverse effects of the drug on the vascular system of transplanted patients via an adaptive response involving Bcl-2 up-regulation rather than an apoptotic process PMID- 11259392 TI - Reactive oxygen species (ROS) mediates the mitochondrial-dependent apoptosis induced by transforming growth factor (beta) in fetal hepatocytes. AB - Treatment of fetal rat hepatocytes with transforming growth factor beta (TGF beta) is followed by apoptotic cell death. Analysis of radical oxygen species (ROS) content and mitochondrial transmembrane potential (Deltapsim), using specific fluorescent probes in FACScan and confocal microscopy, showed that TGF beta mediates ROS production that precedes the loss of Deltapsim, the release of cytochrome c, and the activation of caspase 3. TGF-beta induces a decrease in the protein and mRNA levels of bcl-xL, an antiapoptotic member of the Bcl-2 family. In contrast, there is no change in the expression and/or translocation of Bax, a proapoptotic member of the same family. EGF maintains Bcl-xL, preventing Deltapsim collapse and release of cytochrome c. The presence of radical scavengers blocks the decrease in bcl-xL levels, Deltapsim collapse, cytochrome c release, and activation of caspase 3; in contrast, the presence of glutathione synthesis inhibitors such as BSO accentuated the effect. The incubation of fetal hepatocytes in the presence of ter-butyl-hydroperoxide alone produces a decrease in bcl-xL. These results indicate that during the apoptosis mediated by TGF-beta in fetal hepatocytes, ROS may be responsible for the decrease in bcl-xL mRNA levels that precedes the loss of Deltapsim, the release of cytochrome c, and the activation of caspase 3, culminating in cell death. PMID- 11259393 TI - Increased DNA alterations in atherosclerotic lesions of individuals lacking the GSTM1 genotype. AB - Reduced glutathione (GSH) plays a critical role as an intracellular defense system providing detoxification of a broad spectrum of reactive species and their excretion as water-soluble conjugates. Conjugation of GSH with electrophiles is catalyzed by GSH S-transferases (GST), which constitute a broad family of phase II isoenzymes. Two of the GST encoding genes, GSTM1 (mu) and GSTT1 (theta), have a null genotype due to their homozygous deletion that results in lack of active protein. Polymorphisms within GSTT1 and especially GSTM1 have often been associated with cancer in various organs as well as with elevated levels of DNA adducts in various cell types. We recently demonstrated that DNA adducts are consistently detectable in smooth muscle cells (SMC) of human abdominal aorta affected by atherosclerotic lesions. Here we provide evidence that levels of adducts to SMC DNA from atherosclerotic lesions are consistently increased in individuals having the null GSTM1 genotype, whereas no association was established with the GSTT1 polymorphism. The influence of GSTM1 deletion was better expressed in never-smokers and ex-smokers than in current smokers. These findings bear relevance to the epidemiology of atherosclerosis and suggest that metabolic polymorphisms may contribute to the interindividual variability in susceptibility not only to carcinogens, but also to DNA binding atherogens. PMID- 11259394 TI - Dominant cell death induction by extramitochondrially targeted apoptosis-inducing factor. AB - The complete AIF cDNA comprising the amino-terminal mitochondrial localization sequence (MLS) and the oxidoreductase domain has been fused in its carboxyl terminus to enhanced green fluorescent protein (GFP), thereby engineering an AIF GFP fusion protein that is selectively targeted to the mitochondrial intermembrane space. Upon induction of apoptosis, the AIF-GFP protein translocates together with cytochrome c (Cyt-c) to the extramitochondrial compartment. Microinjection of recombinant AIF leads to the release of AIF-GFP and Cyt-c-GFP, indicating that ectopic AIF can favor permeabilization of the outer mitochondrial membrane. These mitochondrial effects of AIF are caspase independent, whereas the Cyt-c-microinjection induced translocation of AIF-GFP and Cyt-c-GFP is suppressed by the pan-caspase inhibitor Z-VAD.fmk. Upon prolonged culture, transfection-enforced overexpression of AIF results in spontaneous translocation of AIF-GFP from mitochondria, nuclear chromatin condensation, and cell death. These effects are caspase independent and do not rely on the oxidoreductase function of AIF. Spontaneous AIF-GFP translocation and subsequent nuclear apoptosis can be retarded by overexpression of a Bcl-2 protein selectively targeted to mitochondria, but not by a Bcl-2 protein targeted to the endoplasmic reticulum. Overexpression of a mutant AIF protein in which the MLS has been deleted (AIF Delta 1-100) results in the primary cytosolic accumulation of AIF. AIF Delta 1-100-induced cell death is suppressed by neither Z-VAD.fmk or by Bcl-2. Thus, extramitochondrially targeted AIF is a dominant cell death inducer. PMID- 11259395 TI - Immunoglobulin and cytokine expression in mixed lymphocyte cultures is reduced by disruption of gap junction intercellular communication. AB - Connexins (Cx), the protein subunits assembled into gap junction intercellular communication channels, are expressed in primary lymphoid organs and by circulating leukocytes. Human tonsil-derived T and B lymphocytes express Cx40 and 43; circulating human T, B, and NK lymphocytes express Cx43 and directly transfer between each other a low molecular dye indicative that functional gap junctions exist. We now identify specific properties in the immune system underwritten by gap junctions. Mixed lymphocytes cultured in the presence of two reagents with independent inhibitory action on gap junction communication, a connexin mimetic peptide and 18-alpha-glycyrrhetinic acid, markedly reduced the secretion of IgM, IgG, and IgA. The secretion of these immunoglobulins by purified B cells was also reduced by the two classes of gap junction inhibitors. Complex temporal inhibitory effects on the expression of mRNA encoding interleukins, especially IL 10, were also observed. The results indicate that intercellular signaling across gap junctions is an important component of the mechanisms underlying metabolic cooperation in the immune system. PMID- 11259396 TI - Progestins block cholesterol synthesis to produce meiosis-activating sterols. AB - The resumption of meiosis is regulated by meiosis-preventing and meiosis activating substances in testes and ovaries. Certain C29 precursors of cholesterol are present at elevated levels in gonadal tissue, but the mechanism by which these meiosis-activating sterols (MAS) accumulate has remained an unresolved question. Here we report that progestins alter cholesterol synthesis in HepG2 cells and rat testes to increase levels of major MAS (FF-MAS and T-MAS). These C29 sterols accumulated as a result of inhibition of Delta24-reduction and 4alpha-demethylation. Progesterone, pregnenolone, and 17alpha-OH-pregnenolone were potent inhibitors of Delta24-reduction in an in vitro cell assay and led to the accumulation of desmosterol, a Delta5,24 sterol precursor of cholesterol. A markedly different effect was observed for 17alpha-OH-progesterone, which caused the accumulation of sterols associated with inhibition of 4alpha-demethylation. The flux of 13C-acetate into lathosterol and cholesterol was decreased by progestins as measured by isotopomer spectral analysis, whereas newly synthesized MAS accumulated. The combined evidence that MAS concentrations can be regulated by physiological levels of progestins and their specific combination provides a plausible explanation for the elevated concentration of MAS in gonads and suggests a new role for progestins in fertility. PMID- 11259397 TI - beta1 integrin and organized actin filaments facilitate cardiomyocyte-specific RhoA-dependent activation of the skeletal alpha-actin promoter. AB - Activation of RhoA GTPase causes actin filament bundling into stress fibers, integrin clustering, and focal adhesion formation through its action on actin cytoskeleton organization. RhoA also regulates transcriptional activity of serum response factor (SRF). Recent studies in NIH 3T3 fibroblasts have shown that SRF activation by RhoA does not require an organized cytoskeleton and may be regulated by G-actin level. In cardiac myocytes, the organization of actin fibers into myofibrils is one of the primary characteristics of cardiac differentiation and hypertrophy. The primary purpose of this study was to examine if RhoA regulates SRF-dependent gene expression in neonatal cardiomyocytes in a manner different from that observed in fibroblasts. Our results show that RhoA-dependent skeletal alpha-actin promoter activation requires beta1 integrin and a functional cytoskeleton in cardiomyocytes but not in NIH 3T3 fibroblasts. Activation of the alpha-actin promoter by RhoA is greatly potentiated (up to 15-fold) by co expression of the integrin beta1A or beta1D isoform but is significantly reduced by 70% with a co-expressed dominant negative mutant of beta1 integrin. Furthermore, clustering of beta1 integrin with anti-beta1 integrin antibodies potentiates synergistic RhoA and beta1 integrin activation of the alpha-actin promoter. Cytochalasin D and latrunculin B, inhibitors of actin polymerization, significantly reduced RhoA-induced activation of the alpha-actin promoter. Jasplakinolide, an actin polymerizing agent, mimics the synergistic effect of RhoA and beta1 integrin on the actin promoter. These observations support the concept that RhoA regulates SRF-dependent cardiac gene expression through cross talk with beta1 integrin signal pathway via an organized actin cytoskeleton. PMID- 11259398 TI - Molecular mechanisms of TGF-(beta) antagonism by interferon (gamma) and cyclosporine A in lung fibroblasts. AB - Lung fibrosis is a fatal condition of excess extracellular matrix (ECM) deposition associated with increased transforming growth factor beta (TGF-beta) activity. Although much is known about its pathological features, our understanding of the signal transduction pathways resulting in increased ECM and collagen deposition in response to TGF-beta is still incompletely defined. We have previously reported that a JunD homodimer of the transcription factor AP-1 is specifically activated by TGF-beta in lung fibroblasts. Here we demonstrate that JunD is also specifically required for TGF-beta-induced effects. Antisense against JunD, but not c-fos or c-jun, significantly inhibited collagen deposition in response to TGF-beta in primary human lung fibroblasts. We then investigated the ability of pharmacological agents to inhibit TGF-beta-induced signaling and collagen deposition. Cs-A and IFN-gamma, but not glucocorticoids, cyclophosphamide, or azathioprine, inhibited TGF-beta-induced signaling, as assessed by luciferase reporter gene assays, and collagen deposition. TGF-beta antagonism by Cs-A was associated with direct inhibition of JunD activation, as demonstrated by electrophoretic mobility shift analyses. In contrast, the effects of IFN-gamma required signal transducer and activator of transcription (STAT)-1. We thus identify the JunD isoform of AP-1 as an essential mediator of TGF-beta induced effects in lung fibroblasts. TGF-beta-induced signaling and collagen deposition are efficiently antagonized by Cs-A and IFN-gamma treatment, both of which exhibit distinct molecular mechanisms of action. These observations therefore offer novel targets for future therapy of fibrotic lung disease. PMID- 11259399 TI - Ceramide induces aSMase expression: implications for oxLDL-induced apoptosis. AB - Sphingomyelinase (SMase) stimulation and subsequent ceramide generation are suggested to be involved in signal transduction of stress-induced apoptosis. We now show that apoptosis of human macrophages (MPhi) and fibroblasts initiated by oxidized low density lipoproteins (minimally modified LDL, mmLDL) is associated with an increase in acid SMase (aSMase, E.C. 3.1.4.12) expression and ceramide concentration. Application of a novel, potent, and specific inhibitor of aSMase expression (NB6) diminished the effects of mmLDL and C6-ceramide treatment by inhibiting transcription via Sp1 and AP-2. Moreover, apoptosis was abolished after mmLDL and C6-ceramide treatment of hereditary aSMase-deficient fibroblasts (from Niemann-Pick patients). We suggest that in mmLDL-initiated apoptosis 1) enhanced ceramide generation via aSMase appears to be required as well as 2) a positive feedback control of aSMase expression by the increase in intracellular ceramide concentration. PMID- 11259400 TI - Overpassing an aberrant V(kappa) gene to sequence an anti-idiotypic abzyme with (beta)-lactamase-like activity that could have a linkage with autoimmune diseases. AB - A monoclonal antibody 9G4H9 that exhibits a beta-lactamase-like activity was previously obtained in accordance with the idiotypic network theory. This abzyme presents the most catalytic efficiency in amidase activity described in literature (kcat = 0.9 min-1). Some reports have demonstrated that functionality as complex as catalysis may be mimicked in this way. Comparison of the catalytic properties of both enzyme and abzyme previously allowed us to obtain better knowledge about 9G4H9 abzymatic machinery. In attempt to characterize this abzyme, the variable regions of kappa and heavy chain were cloned. We present a 'universal' method to clone the correct Vkappa gene to bypass aberrant Vkappa (abVkappa) produced by MOPC-21-derived hybridomas. Sequences obtained are compared in the GenBank database. The VH and Vkappa genes present some important sequence homology with autoantibodies suggesting a direct relationship between catalytic anti-idiotypic antibody and autoimmunity. PMID- 11259401 TI - O2 dependence of K+ transport in sickle cells: the effect of different cell populations and the substituted benzaldehyde 12C79. AB - The molecular basis of sickle cell disease (SCD) is well known but the pathophysiology is poorly understood. It remains intractable to therapy. Hyperactivity of several membrane transport systems, including the K+-Cl- cotransporter (termed KCC), cause HbS-containing red cells (termed HbS cells) to dehydrate and sickle, leading to the development of sickle cell crises (SCCs). Contrary to normal red cells (HbA cells), KCC in HbS cells is active at low O2 tensions (PO2s), remaining responsive to low pH or urea. Since these stimuli are usually encountered in hypoxic regions, the abnormal O2 dependence increases the contribution of KCC to dehydration, and hence development of SCCs. These differences with HbA cells may be due to the younger population of cells or to polymerization of HbS. We used 86Rb+ as a K+ congener to investigate the activity of KCC at different PO2s, and density gradient separation to investigate different red cell fractions. We found no correlation of O2 dependence with cell fractions. We also used the substituted benzaldehyde 12C79 to increase the O2 affinity of HbS and found that its effect on HbS O2 saturation and cell sickling correlated with that on both Cl--independent and Cl--dependent K+ transport, implying that, at low PO2s, KCC activity correlated with HbS polymerization. The importance of these results to understanding the pathophysiology of SCD, and for the design of chemotherapeutic agents to ameliorate or prevent SCC, is discussed. PMID- 11259403 TI - Neural network model of gene expression. AB - Many natural processes consist of networks of interacting elements that, over time, affect each other's state. Their dynamics depend on the pattern of connections and the updating rules for each element. Genomic regulatory networks are networks of this sort. In this paper we use artificial neural networks as a model of the dynamics of gene expression. The significance of the regulatory effect of one gene product on the expression of other genes of the system is defined by a weight matrix. The model considers multigenic regulation including positive and/or negative feedback. The process of gene expression is described by a single network and by two linked networks where transcription and translation are modeled independently. Each of these processes is described by different network controlled by different weight matrices. Methods for computing the parameters of the model from experimental data are discussed. Results computed by means of the model are compared with experimental observations. Generalization to a 'black box' concept, where the molecular processes occurring in the cell are considered as signal processing units forming a global regulatory network, is discussed.-Vohradsky, J. Neural network model of gene expression. PMID- 11259402 TI - Uncoupling protein 3 transcription is regulated by peroxisome proliferator activated receptor (alpha) in the adult rodent heart. AB - Relatively little is known concerning the regulation of uncoupling proteins (UCPs) in the heart. We investigated in the adult rodent heart 1) whether changes in workload, substrate supply, or cytokine (TNF-alpha) administration affect UCP 2 and UCP-3 expression, and 2) whether peroxisome proliferator-activated receptor alpha (PPARalpha) regulates the expression of either UCP-2 or UCP-3. Direct comparisons were made between cardiac and skeletal muscle. UCP-2, UCP-3, and PPARalpha expression were reduced when cardiac workload was either increased (pressure overload by aortic constriction) or decreased (mechanical unloading by heterotopic transplantation). Similar results were observed during cytokine administration. Reduced dietary fatty acid availability resulted in decreased expression of both cardiac UCP-2 and UCP-3. However, when fatty acid (the natural ligand for PPARalpha) supply was increased (high-fat feeding, fasting, and STZ induced diabetes), cardiac UCP-3 but not UCP-2 expression increased. Comparable results were observed in rats treated with the specific PPARalpha agonist WY 14,643. The level of cardiac UCP-3 but not UCP-2 expression was severely reduced (20-fold) in PPARalpha-/- mice compared to wild-type mice. These results suggest that in the adult rodent heart, UCP-3 expression is regulated by PPARalpha. In contrast, cardiac UCP-2 expression is regulated in part by a fatty acid dependent, PPARalpha-independent mechanism. PMID- 11259404 TI - Pex19p dampens the p19ARF-p53-p21WAF1 tumor suppressor pathway. AB - We isolated a 33-kDa protein, Pex19p/HK33/HsPXF, as a p19ARF-binding protein in a yeast two-hybrid screen. We demonstrate here that Pex19p interacts with p19ARF in the cell cytoplasm and excludes p19ARF from the nucleus, leading to a concurrent inactivation of p53 function. Down-regulation of Pex19p by its antisense expression resulted in increased levels of p19ARF, increased p53 function, and a p53/p21WAF1-mediated senescence-like cell cycle arrest. The data demonstrated a novel mechanism of down-regulation of the p19ARF-p53 pathway. PMID- 11259405 TI - DNA methyltransferase inhibition in normal human fibroblasts induces a p21 dependent cell cycle withdrawal. AB - Maintenance of methylation patterns in the mammalian genome by DNA (cytosine-5) methyltransferases (DNAMeTase) is required for normal cell and tissue function. Inhibition of DNAMeTase in cultured cells induces the expression of p21, a cyclin dependent kinase (Cdk) inhibitor critical for cells to enter replicative senescence. We investigated the effects of DNAMeTase inhibition in normal human fibroblasts and found that it induces an irreversible growth arrest. Cells arrested by DNAMeTase inhibition became enlarged and had a flat morphology, exhibited an increased expression of collagenase and p21, and the DNA synthesis block could be overcome by the introduction of the SV40 large T antigen, all characteristics of senescent cells. In contrast, normal human fibroblasts lacking a functional p21 gene fail to undergo cell cycle arrest following DNAMeTase inhibition, indicating that p21 is an essential component of this arrest. Furthermore, DNAMeTase activity was reduced as cells approached the end of their proliferative potential. These data suggest that DNAMeTase could be an integral part of the mechanisms by which cells count the number of cell divisions completed and initiate a signaling cascade that ultimately results in the senescent phenotype. PMID- 11259406 TI - Independent repression of a GC-rich housekeeping gene by Sp1 and MAZ involves the same cis-elements. AB - The transcription factors Sp1 and MAZ (Myc-associated zinc finger protein) contain several zinc finger motifs, and each functions as both a positive and a negative regulator of gene expression. In this study, we characterized the extremely GC-rich promoter of the human gene for MAZ, which is known as a housekeeping gene. Unique symmetrical motifs in the promoter region (nucleotides 383 to -334) were essential for the expression of the gene for MAZ, whereas an upstream silencer element (nucleotides -784 to -612) was found to act in a position-dependent but orientation-independent manner. Sp1 and MAZ bound to the same cis-elements in the GC-rich promoter, apparently sharing DNA-binding sites. The relative extent of binding of Sp1 and MAZ to these cis-elements corresponded to the extent of negative regulation of the expression of the gene for MAZ in various lines of cells. Furthermore, novel repressive domains in both Sp1 (amino acids 622-788) and MAZ (amino acids 127-292) were identified. Suppression by Sp1 and suppression by MAZ were independent phenomena; histone deacetylases were involved in the autorepression by MAZ itself, whereas DNA methyltransferase 1 was associated with suppression by Sp1. Our results indicate that both deacetylation and methylation might be involved in the regulation of expression of a single gene via the actions of different zinc finger proteins that bind to the same cis elements. PMID- 11259407 TI - The epithelium-specific ETS protein EHF/ESE-3 is a context-dependent transcriptional repressor downstream of MAPK signaling cascades. AB - Exon trapping and cDNA selection procedures were used to search for novel genes at human chromosome 11p13, a region previously associated with loss of heterozygosity in epithelial carcinomas. Using these approaches, we found the ESE 2 and ESE-3 genes, coding for ETS domain-containing transcription factors. These genes lie in close proximity to the catalase gene within a approximately 200 kilobase genomic interval. ESE-3 mRNA is widely expressed in human tissues with high epithelial content, and immunohistochemical analysis with a newly generated monoclonal antibody revealed that ESE-3 is a nuclear protein expressed exclusively in differentiated epithelial cells and that it is absent in the epithelial carcinomas tested. In transient transfections, ESE-3 behaves as a repressor of the Ras- or phorbol ester-induced transcriptional activation of a subset of promoters that contain ETS and AP-1 binding sites. ESE-3-mediated repression is sequence- and context-dependent and depends both on the presence of high affinity ESE-3 binding sites in combination with AP-1 cis-elements and the arrangement of these sites within a given promoter. We propose that ESE-3 might be an important determinant in the control of epithelial differentiation, as a modulator of the nuclear response to mitogen-activated protein kinase signaling cascades. PMID- 11259408 TI - Retention of heme in axial ligand mutants of succinate-ubiquinone xxidoreductase (complex II) from Escherichia coli. AB - Succinate-ubiquinone oxidoreductase (SdhCDAB, complex II) from Escherichia coli is a four-subunit membrane-bound respiratory complex that catalyzes ubiquinone reduction by succinate. In the E. coli enzyme, heme b(556) is ligated between SdhC His(84) and SdhD His(71). Contrary to a previous report (Vibat, C. R. T., Cecchini, G., Nakamura, K., Kita, K., and Gennis, R. B. (1998) Biochemistry 37, 4148-4159), we demonstrate the presence of heme in both SdhC H84L and SdhD H71Q mutants of SdhCDAB. EPR spectroscopy reveals the presence of low spin heme in the SdhC H84L (g(z) = 2.92) mutant and high spin heme in the SdhD H71Q mutant (g = 6.0). The presence of low spin heme in the SdhC H84L mutant suggests that the heme b(556) is able to pick up another ligand from the protein. CO binds to the reduced form of the mutants, indicating that it is able to displace one of the ligands to the low spin heme of the SdhC H84L mutant. The g = 2.92 signal of the SdhC H84L mutant titrates with a redox potential at pH 7.0 (E(m)(,7)) of approximately +15 mV, whereas the g = 6.0 signal of the SdhD H71Q mutant titrates with an E(m)(,7) of approximately -100 mV. The quinone site inhibitor pentachlorophenol perturbs the heme optical spectrum of the wild-type and SdhD H71Q mutant enzymes but not the SdhC H84L mutant. This finding suggests that the latter residue also plays an important role in defining the quinone binding site of the enzyme. The SdhC H84L mutation also results in a significant increase in the K(m) and a decrease in the k(cat) for ubiquinone-1, whereas the SdhD H71Q mutant has little effect on these parameters. Overall, these data indicate that SdhC His(84) has an important role in defining the interaction of SdhCDAB with both quinones and heme b(556). PMID- 11259409 TI - Identification of new JNK substrate using ATP pocket mutant JNK and a corresponding ATP analogue. AB - Modification of the ATP pocket on protein kinases allows selective use of an ATP analogue that exhibits high affinity for the altered kinases. Using this approach, we altered the ATP-binding site on JNK and identified N(6)-(2 phenythyl)-ATP, a modified form of ATP that exhibits high specificity and affinity for the modified, but not the wild type form, of JNK. Using modified JNK and its ATP analogue enables the detection of novel JNK substrates. Among substrates identified using this approach is heterogeneous nuclear ribonucleoprotein K, which is involved in transcription and post-transcriptional mRNA metabolism. The newly identified substrate can be phosphorylated by JNK on amino acids 216 and 353, which contribute to heterogeneous nuclear ribonucleoprotein K mediated transcriptional activities. PMID- 11259410 TI - The small ubiquitin-like modifier-1 (SUMO-1) consensus sequence mediates Ubc9 binding and is essential for SUMO-1 modification. AB - SUMO-1 is an ubiquitin-related protein that is covalently conjugated to a diverse assortment of proteins. The consequences of SUMO-1 modification include the regulation of protein-protein interactions, protein-DNA interactions, and protein subcellular localization. At present, very little is understood about the specific mechanisms that govern the recognition of proteins as substrates for SUMO-1 modification. However, many of the proteins that are modified by SUMO-1 interact directly with the SUMO-1 conjugating enzyme, Ubc9. These interactions suggest that Ubc9 binding may play an important role in substrate recognition as well as in substrate modification. The SUMO-1 consensus sequence (SUMO-1-CS) is a motif of conserved residues surrounding the modified lysine residue of most SUMO 1 substrates. This motif conforms to the sequence "PsiKXE," where Psi is a large hydrophobic residue, K is the lysine to which SUMO-1 is conjugated, X is any amino acid, and E is glutamic acid. In this study, we demonstrate that the SUMO-1 CS is a major determinant of Ubc9 binding and SUMO-1 modification. Mutating residues in the SUMO-1-CS abolishes both Ubc9 binding and substrate modification. These findings have important implications for how SUMO-1 substrates are recognized and for how SUMO-1 is ultimately transferred to specific lysine residues on these substrates. PMID- 11259411 TI - Overexpression of TAP/p15 heterodimers bypasses nuclear retention and stimulates nuclear mRNA export. AB - Human TAP and its yeast orthologue Mex67p are members of the multigene family of NXF proteins. A conserved feature of NXFs is a leucine-rich repeat domain (LRR) followed by a region related to the nuclear transport factor 2 (the NTF2-like domain). The NTF2-like domain of metazoan NXFs heterodimerizes with a protein known as p15 or NXT. A C-terminal region related to ubiquitin-associated domains (the UBA-like domain) is present in most, but not all NXF proteins. Saccharomyces cerevisiae Mex67p and Caenorhabditis elegans NXF1 are essential for the export of messenger RNA from the nucleus. Human TAP mediates the export of simian type D retroviral RNAs bearing the constitutive transport element, but the precise role of TAP and p15 in mRNA nuclear export has not yet been established. Here we show that overexpression of TAP/p15 heterodimers bypasses nuclear retention and stimulates the export of mRNAs that are otherwise exported inefficiently. This stimulation of mRNA export is strongly reduced by removing the UBA-like domain of TAP and abolished by deleting the LRR domain or the NTF2-like domain. Similar results are obtained when TAP/p15 heterodimers are directly tethered to the RNA export cargo. Our data indicate that formation of TAP/p15 heterodimers is required for TAP-mediated export of mRNA and show that the LRR domain of TAP plays an essential role in this process. PMID- 11259412 TI - Alternative cyclization in GFP-like proteins family. The formation and structure of the chromophore of a purple chromoprotein from Anemonia sulcata. AB - Anemonia sulcata purple protein (asFP595) belongs to a family of green fluorescent protein (GFP)-like proteins from the Anthozoa species. Similar to GFP, asFP595 apparently forms its chromophore by modifying amino acids within its polypeptide chain. Until now, the GFP-like proteins from Anthozoa were thought to contain chromophores with the same imidazolidinone core as GFP. Mass spectral analysis of a chromophore-containing tryptic pentapeptide from asFP595 demonstrates that chromophore formation in asFP595 is stoichiometrically the same as that in GFP: one H(2)O and two H(+) are released while a Schiff base and dehydrotyrosine are formed. However, structural studies of this asFP595 chromopeptide show that in contrast to GFP, the other peptide bond nitrogen and carbonyl carbon are required for chromophore cyclization, a reaction that yields the six-membered heterocycle 2-(4-hydroxybenzylidene)-6-hydroxy-2,5 dihydropyrazine. Spectrophotometric titration reveals three pH-dependent forms of the asFP595 chromopeptide: yellow (absorption maximum = 430 nm) at pH 3.0; red (absorption maximum = 535 nm) at pH 8.0; and colorless (absorption maximum = 380 nm) at pH 14.0. The pK(a) values for these spectral transitions (6.8 and 10.9) are consistent with the ionization of the phenolic group of dehydrotyrosine and deprotonation of the amidinium cation in the chromophore heterocycle, respectively. The amidinium group in asFP595 accounts for the unique absorption spectrum of the protein, which is substantially red-shifted relative to that of GFP. When the asFP595 chromophore cyclizes, the Cys-Met bond adjacent to the chromophore hydrolyzes, splitting the chromoprotein into 8- and 20-kDa fragments. High performance liquid chromatography analysis of a tryptic digest of denatured asFP595 shows that a pentapeptide with the cleaved Cys-Met bond is the only fragment associated with the red-shifted absorbance. These results imply that fragmentation of asFP595 is a critical step in protein maturation. PMID- 11259413 TI - Biglycan and decorin bind close to the n-terminal region of the collagen VI triple helix. AB - The binding of native biglycan and decorin to pepsin-extracted collagen VI from human placenta was examined by solid phase assay and by measurement of surface plasmon resonance in the BIAcore(TM)2000 system. Both proteoglycans exhibited a strong affinity for collagen VI with dissociation constants (K(D)) of approximately 30 nm. Removal of the glycosaminoglycan chains by chondroitinase ABC digestion did not significantly affect binding. In coprecipitation experiments, biglycan and decorin bound to collagen VI and equally competed with the other, suggesting that biglycan and decorin bind to the same binding site on collagen VI. This was confirmed by electron microscopy after negative staining of complexes between gold-labeled proteoglycans and collagen VI, demonstrating that both biglycan and decorin bound exclusively to a domain close to the interface between the N terminus of the triple helical region and the following globular domain. In solid phase assay using recombinant collagen VI fragments, it was shown that the alpha2(VI) chain probably plays a role in the interaction. PMID- 11259414 TI - Caveolin-3 null mice show a loss of caveolae, changes in the microdomain distribution of the dystrophin-glycoprotein complex, and t-tubule abnormalities. AB - Caveolin-3, a muscle-specific caveolin-related protein, is the principal structural protein of caveolae membrane domains in striated muscle cells. Recently, we identified a novel autosomal dominant form of limb-girdle muscular dystrophy (LGMD-1C) in humans that is due to mutations within the coding sequence of the human caveolin-3 gene (3p25). These LGMD-1C mutations lead to an approximately 95% reduction in caveolin-3 protein expression, i.e. a caveolin-3 deficiency. Here, we created a caveolin-3 null (CAV3 -/-) mouse model, using standard homologous recombination techniques, to mimic a caveolin-3 deficiency. We show that these mice lack caveolin-3 protein expression and sarcolemmal caveolae membranes. In addition, analysis of skeletal muscle tissue from these caveolin-3 null mice reveals: (i) mild myopathic changes; (ii) an exclusion of the dystrophin-glycoprotein complex from lipid raft domains; and (iii) abnormalities in the organization of the T-tubule system, with dilated and longitudinally oriented T-tubules. These results have clear mechanistic implications for understanding the pathogenesis of LGMD-1C at a molecular level. PMID- 11259415 TI - NUB1, a NEDD8-interacting protein, is induced by interferon and down-regulates the NEDD8 expression. AB - NEDD8, a ubiquitin-like protein, covalently conjugates to cullin family members. It appears to control vital biological events through its conjugation to cullins. To study how this conjugation pathway is regulated, we performed yeast two-hybrid screening by using NEDD8 as a bait and isolated a cDNA fragment encoding a potent down-regulator of the NEDD8 expression. Here, we report this novel regulator, NUB1 (NEDD8 Ultimate Buster-1). NUB1 is composed of 601 residues with a calculated 69.1-kDa molecular mass. It is an interferon-inducible protein and predominantly localized in the nucleus. The NUB1 message is specifically expressed in adult human testis, ovary, heart, and skeletal muscle tissues and is developmentally down-regulated in mouse embryos. In biochemical analysis, we found that NUB1 overexpression leads to severe reduction of NEDD8 monomer and NEDD8 conjugates in cells. This reduction is not due to down-regulation of NEDD8 transcription, but due to post-transcriptional mechanism. As expected from this activity, overexpression of NUB1 had a profound growth-inhibitory effect on U2OS cells. Thus, NUB1 is a strong down-regulator of the NEDD8 expression and appears to play critical roles in regulating biological events, including cell growth. PMID- 11259416 TI - Assessment of the role of the inositol 1,4,5-trisphosphate receptor in the activation of transient receptor potential channels and store-operated Ca2+ entry channels. AB - The mechanism for coupling between Ca(2+) stores and store-operated channels (SOCs) is an important but unresolved question. Although SOCs have not been molecularly identified, transient receptor potential (TRP) channels share a number of operational parameters with SOCs. The question of whether activation of SOCs and TRP channels is mediated by the inositol 1,4,5-trisphosphate receptor (InsP(3)R) was examined using the permeant InsP(3)R antagonist, 2 aminoethoxydiphenyl borate (2-APB) in both mammalian and invertebrate systems. In HEK293 cells stably transfected with human TRPC3 channels, the actions of 2-APB to block carbachol-induced InsP(3)R-mediated store release and carbachol-induced Sr(2+) entry through TRPC3 channels were both reversed at high agonist levels, suggesting InsP(3)Rs mediate TRPC3 activation. However, electroretinogram recordings of the light-induced current in Drosophila revealed that the TRP channel-mediated responses in wild-type as well as trp and trpl mutant flies were all inhibited by 2-APB. This action of 2-APB is likely InsP(3)R-independent since InsP(3)Rs are dispensable for the light response. We used triple InsP(3)R knockout DT40 chicken B-cells to further assess the role of InsP(3)Rs in SOC activation. (45)Ca(2+) flux analysis revealed that although DT40 wild-type cells retained normal InsP(3)Rs mediating 2-APB-sensitive Ca(2+) release, the DT40InsP(3)R-k/o cells were devoid of functional InsP(3)Rs. Using intact cells, all parameters of Ca(2+) store function and SOC activation were identical in DT40wt and DT40InsP(3)R-k/o cells. Moreover, in both cell lines SOC activation was completely blocked by 2-APB, and the kinetics of action of 2-APB on SOCs (time dependence and IC(50)) were identical. The results indicate that (a) the action of 2-APB on Ca(2+) entry is not mediated by the InsP(3)R and (b) the effects of 2-APB provide evidence for an important similarity in the function of invertebrate TRP channels, mammalian TRP channels, and mammalian store-operated channels. PMID- 11259417 TI - Transcription factors TFIIF, ELL, and Elongin negatively regulate SII-induced nascent transcript cleavage by non-arrested RNA polymerase II elongation intermediates. AB - TFIIF, ELL, and Elongin belong to a class of RNA polymerase II transcription factors that function similarly to activate the rate of elongation by suppressing transient pausing by polymerase at many sites along DNA templates. SII is a functionally distinct RNA polymerase II elongation factor that promotes elongation by reactivating arrested polymerase. Studies of the mechanism of SII action have shown (i) that arrest of RNA polymerase II results from irreversible displacement of the 3'-end of the nascent transcript from the polymerase catalytic site and (ii) that SII reactivates arrested polymerase by inducing endonucleolytic cleavage of the nascent transcript by the polymerase catalytic site thereby creating a new transcript 3'-end that is properly aligned with the catalytic site and can be extended. SII also induces nascent transcript cleavage by paused but non-arrested RNA polymerase II elongation intermediates, leading to the proposal that pausing may result from reversible displacement of the 3'-end of nascent transcripts from the polymerase catalytic site. On the basis of evidence consistent with the model that TFIIF, ELL, and Elongin suppress pausing by preventing displacement of the 3'-end of the nascent transcript from the polymerase catalytic site, we investigated the possibility of cross-talk between SII and transcription factors TFIIF, ELL, and Elongin. These studies led to the discovery that TFIIF, ELL, and Elongin are all capable of inhibiting SII-induced nascent transcript cleavage by non-arrested RNA polymerase II elongation intermediates. Here we present these findings, which bring to light a novel activity associated with TFIIF, ELL, and Elongin and suggest that these transcription factors may expedite elongation not only by increasing the forward rate of nucleotide addition by RNA polymerase II, but also by inhibiting SII induced nascent transcript cleavage by non-arrested RNA polymerase II elongation intermediates. PMID- 11259418 TI - Control of the orientation of Fos-Jun binding and the transcriptional cooperativity of Fos-Jun-NFAT1 complexes. AB - Heterodimeric transcription regulatory proteins can bind to palindromic recognition elements in two opposite orientations. We have developed a gel-based fluorescence resonance energy transfer assay for quantifying heterodimer orientation preferences. Fos-Jun heterodimers bind in opposite orientations to AP 1 sites with different flanking sequences. The effects of individual amino acid and base pair substitutions on heterodimer binding orientation were quantified. Base pairs at positions +/-6 and +/-10 relative to the center of the AP-1 site were the principal determinants of Fos-Jun binding orientation. Amino acid residues of opposite charge adjacent to the basic regions of Fos and Jun had independent effects on heterodimer orientation. Exchange of these amino acid residues between the basic region-leucine zipper domains of Fos and Jun reversed the binding orientation. Heterodimers formed by full-length Fos and Jun exhibited the same changes in binding orientation in response to amino acid and base pair substitutions. The preferred orientation of heterodimer binding affected the stability of Fos-Jun-NFAT1 complexes at composite regulatory elements. Changes in heterodimer orientation preference altered the transcriptional activity and the promoter selectivity of Fos-Jun-NFAT1 complexes. Consequently, the orientation of Fos-Jun binding can influence transcriptional activity by altering cooperative interactions with other transcription regulatory proteins. PMID- 11259419 TI - The internal repeats in the Na+/Ca2+ exchanger-related Escherichia coli protein YrbG have opposite membrane topologies. AB - We have determined the topology of the Escherichia coli inner membrane protein YrbG, a putative Na(+)/Ca(2+) exchanger with homology to a family of eukaryotic ion exchangers. Our results show that the two homologous halves of YrbG both have five transmembrane segments but opposite membrane orientations. This has implications for our understanding of the function of Na(+)/Ca(2+) exchangers and provides an example of "divergent" evolution of membrane protein topology. PMID- 11259420 TI - Two distinct pathways of formation of 4-hydroxynonenal. Mechanisms of nonenzymatic transformation of the 9- and 13-hydroperoxides of linoleic acid to 4 hydroxyalkenals. AB - The mechanism of formation of 4-hydroxy-2E-nonenal (4-HNE) has been a matter of debate since it was discovered as a major cytotoxic product of lipid peroxidation in 1980. Recent evidence points to 4-hydroperoxy-2E-nonenal (4-HPNE) as the immediate precursor of 4-HNE (Lee, S. H., and Blair, I. A. (2000) Chem. Res. Toxicol. 13, 698-702; Noordermeer, M. A., Feussner, I., Kolbe, A., Veldink, G. A., and Vliegenthart, J. F. G. (2000) Biochem. Biophys. Res. Commun. 277, 112 116), and a pathway via 9-hydroperoxylinoleic acid and 3Z-nonenal is recognized in plant extracts. Using the 9- and 13-hydroperoxides of linoleic acid as starting material, we find that two distinct mechanisms lead to the formation of 4-H(P)NE and the corresponding 4-hydro(pero)xyalkenal that retains the original carboxyl group (9-hydroperoxy-12-oxo-10E-dodecenoic acid). Chiral analysis revealed that 4-HPNE formed from 13S-hydroperoxy-9Z,11E-octadecadienoic acid (13S HPODE) retains >90% S configuration, whereas it is nearly racemic from 9S hydroperoxy-10E,12Z-octadecadienoic acid (9S-HPODE). 9-Hydroperoxy-12-oxo-10E dodecenoic acid is >90% S when derived from 9S-HPODE and almost racemic from 13S HPODE. Through analysis of intermediates and products, we provide evidence that (i) allylic hydrogen abstraction at C-8 of 13S-HPODE leads to a 10,13 dihydroperoxide that undergoes cleavage between C-9 and C-10 to give 4S-HPNE, whereas direct Hock cleavage of the 13S-HPODE gives 12-oxo-9Z-dodecenoic acid, which oxygenates to racemic 9-hydroperoxy-12-oxo-10E-dodecenoic acid; by contrast, (ii) 9S-HPODE cleaves directly to 3Z-nonenal as a precursor of racemic 4-HPNE, whereas allylic hydrogen abstraction at C-14 and oxygenation to a 9,12 dihydroperoxide leads to chiral 9S-hydroperoxy-12-oxo-10E-dodecenoic acid. Our results distinguish two major pathways to the formation of 4-HNE that should apply also to other fatty acid hydroperoxides. Slight ( approximately 10%) differences in the observed chiralities from those predicted in the above mechanisms suggest the existence of additional routes to the 4-hydroxyalkenals. PMID- 11259421 TI - The mouse dystrophin enhancer is regulated by MyoD, E-box-binding factors, and by the serum response factor. AB - In vivo studies in the mouse have revealed that the muscle promoter of the mouse dystrophin gene can target the right ventricle of the heart only, suggesting the need for other regulatory elements to target the skeletal muscle as well as other compartments of the heart. In this study we report the identification of the mouse dystrophin gene enhancer that is located approximately 8.5 kilobases downstream from the mouse dystrophin gene muscle promoter. The enhancer was tested in myogenic G8, H9-C2, and nonmyogenic 3T3 cell lines and is mostly active in G8 myotubes. Sequence analysis of the mouse dystrophin gene enhancer revealed the presence of four E-boxes numbered E1-E4, a putative mef-2 binding site, and a serum response element. Site-directed mutagenesis studies have shown that E-boxes 1, 2, and 3 as well as the serum response element are required for enhancer activity. Gel shift analysis revealed two binding activities at binding sites E1 and E3 which were specific to myotubes, and supershift assays confirmed that myoD binds at both these sites. Our study also shows that werum response factor binds the serum response element but in myoblasts and fibroblasts only, suggesting that serum response factor may repress enhancer function. PMID- 11259422 TI - Agonist-dependent recruitment of phosphoinositide 3-kinase to the membrane by beta-adrenergic receptor kinase 1. A role in receptor sequestration. AB - Agonist-dependent desensitization of the beta-adrenergic receptor requires translocation and activation of the beta-adrenergic receptor kinase1 by liberated Gbetagamma subunits. Subsequent internalization of agonist-occupied receptors occurs as a result of the binding of beta-arrestin to the phosphorylated receptor followed by interaction with the AP2 adaptor and clathrin proteins. Receptor internalization is known to require D-3 phosphoinositides that are generated by the action of phosphoinositide 3-kinase. Phosphoinositide 3-kinases form a family of lipid kinases that couple signals via receptor tyrosine kinases and G-protein coupled receptors. The molecular mechanism by which phosphoinositide 3-kinase acts to promote beta-adrenergic receptor internalization is not well understood. In the present investigation we demonstrate a novel finding that beta-adrenergic receptor kinase 1 and phosphoinositide 3-kinase form a cytosolic complex, which leads to beta-adrenergic receptor kinase 1-mediated translocation of phosphoinositide 3-kinase to the membrane in an agonist-dependent manner. Furthermore, agonist-induced translocation of phosphoinositide 3-kinase results in rapid interaction with the receptor, which is of functional importance, since inhibition of phosphoinositide 3-kinase activity attenuates beta-adrenergic receptor sequestration. Therefore, agonist-dependent recruitment of phosphoinositide 3-kinase to the membrane is an important step in the process of receptor sequestration and links phosphoinositide 3-kinase to G-protein-coupled receptor activation and sequestration. PMID- 11259423 TI - The effect of DNA structure on the catalytic efficiency and fidelity of human DNA polymerase beta on templates with platinum-DNA adducts. AB - DNA adducts formed by platinum-based anticancer drugs interfere with DNA replication. The carrier ligand of the platinum compound is likely to affect the conformation of the Pt-DNA adducts. In addition, the conformation of the adduct can also change upon binding of damaged DNA to the active site of DNA polymerase. From the crystal structures of pol beta ternary complexes it is evident that undamaged gapped and primed single-stranded (non-gapped) DNA templates exist in very different conformations when bound to pol beta. Therefore, one might expect that the constraints imposed on the damaged templates by binding to the polymerase active site should also affect the conformation of the Pt-DNA adducts and their ability to inhibit DNA replication. In support of this hypothesis we have found that the efficiency, carrier ligand specificity, site of discrimination (3'-G versus 5'-G of the Pt-GG adducts), and fidelity of translesion synthesis past Pt-DNA adducts by pol beta are strongly affected by the structure of the DNA template. Previous studies have suggested that the conformation of Pt-DNA adducts may be affected by the sequence context of the adduct. In support of this hypothesis, our data show that sequence context affects the efficiency, fidelity, and pattern of misincorporation by pol beta. PMID- 11259424 TI - Monocyte membrane type 1-matrix metalloproteinase. Prostaglandin-dependent regulation and role in metalloproteinase-2 activation. AB - Membrane type 1-matrix metalloproteinase (MT1-MMP)-mediated activation of MMP-2 is thought to be important in the proteolysis of extracellular matrix in pathological events in which monocytes/macrophages are found. Here we report on the induction and regulation of human monocyte MT1-MMP and its role in MMP-2 activation. Activation of monocytes by lipopolysaccharide resulted in the induction of MT1-MMP mRNA and protein that was suppressed by inhibitors of prostaglandin synthesis (indomethacin), adenylyl cyclase (SQ 22536), and protein kinase A (Rp-cAMPs). Suppression of MT1-MMP by indomethacin and SQ 22536 was reversed by prostaglandin E(2) and dibutyryl cyclic AMP, respectively, demonstrating that induction of monocyte MT1-MMP is regulated through a prostaglandin-cAMP pathway. Functional analysis revealed that pro-MMP-2 in the supernatants from human bone marrow stromal fibroblasts, normal male-derived fibroblasts and melanoma cells (A2058) was converted to active MMP-2 when cultured with activated but not control monocytes. Antibodies against MT1-MMP blocked the activation of MMP-2. Tissue inhibitor of metalloproteinase-2 regulation of MMP-2 activation was shown through the addition of varying amounts of recombinant tissue inhibitor of metalloproteinase-2 with pro-MMP-2 to MT1-MMP expressing monocytes. These findings demonstrate that activated monocytes express functionally active MT1-MMP that may play a significant role in the activation of MMP-2 produced by other cells and as such influence developmental and pathological conditions. PMID- 11259425 TI - Characterization of fibrosurfin, an interfibrillar component of sea urchin catch connective tissues. AB - The Sea URchin Fibrillar (SURF) domain is a four-cysteine module present in the amino-propeptide of the sea urchin 2alpha fibrillar collagen chain. Despite numerous international genome and expressed sequence tag projects, computer searches have so far failed to identify similar domains in other species. Here, we have characterized a new sea urchin protein of 2656 amino acids made up of a series of epidermal growth factor-like and SURF modules. From its striking similarity to the modular organization of fibropellins, we called this new protein fibrosurfin. This protein is acidic with a calculated pI of 4.12. Eleven of the 17 epidermal growth factor-like domains correspond to the consensus sequence of calcium-binding type. By Western blot and immunofluorescence analyses, this protein is not detectable during embryogenesis. In adult tissues, fibrosurfin is co-localized with the amino-propeptide of the 2alpha fibrillar collagen chain in several collagenous ligaments, i.e., test sutures, spine ligaments, peristomial membrane, and to a lesser extent, tube feet. Finally, immunogold labeling indicates that fibrosurfin is an interfibrillar component of collagenous tissues. Taken together, the data suggest that proteins possessing SURF modules are localized in the vicinity of mineralized tissues and could be responsible for the unique properties of sea urchin mutable collagenous tissues. PMID- 11259426 TI - Improved statistical inference from DNA microarray data using analysis of variance and a Bayesian statistical framework. Analysis of global gene expression in Escherichia coli K12. AB - We describe statistical methods based on the t test that can be conveniently used on high density array data to test for statistically significant differences between treatments. These t tests employ either the observed variance among replicates within treatments or a Bayesian estimate of the variance among replicates within treatments based on a prior estimate obtained from a local estimate of the standard deviation. The Bayesian prior allows statistical inference to be made from microarray data even when experiments are only replicated at nominal levels. We apply these new statistical tests to a data set that examined differential gene expression patterns in IHF(+) and IHF(-) Escherichia coli cells (Arfin, S. M., Long, A. D., Ito, E. T., Tolleri, L., Riehle, M. M., Paegle, E. S., and Hatfield, G. W. (2000) J. Biol. Chem. 275, 29672-29684). These analyses identify a more biologically reasonable set of candidate genes than those identified using statistical tests not incorporating a Bayesian prior. We also show that statistical tests based on analysis of variance and a Bayesian prior identify genes that are up- or down-regulated following an experimental manipulation more reliably than approaches based only on a t test or fold change. All the described tests are implemented in a simple-to-use web interface called Cyber-T that is located on the University of California at Irvine genomics web site. PMID- 11259427 TI - Tryptase 4, a new member of the chromosome 17 family of mouse serine proteases. AB - Genomic blot analysis raised the possibility that uncharacterized tryptase genes reside on chromosome 17 at the complex containing the three genes that encode mouse mast cell protease (mMCP) 6, mMCP-7, and transmembrane tryptase (mTMT). Probing of GenBank's expressed sequence tag data base with these three tryptase cDNAs resulted in the identification of an expressed sequence tag that encodes a portion of a novel mouse serine protease (now designated mouse tryptase 4 (mT4) because it is the fourth member of this family). 5'- and 3'-rapid amplification of cDNA ends approaches were carried out to deduce the nucleotide sequence of the full-length mT4 transcript. This information was then used to clone its approximately 5.0-kilobase pair gene. Chromosome mapping analysis of its gene, sequence analysis of its transcript, and comparative protein structure modeling of its translated product revealed that mT4 is a new member of the chromosome 17 family of mouse tryptases. mT4 is 40-44% identical to mMCP-6, mMCP-7, and mTMT, and this new serine protease has all of the structural features of a functional tryptase. Moreover, mT4 is enzymatically active when expressed in insect cells. Due to its 17-mer hydrophobic domain at its C terminus, mT4 is a membrane anchored tryptase more analogous to mTMT than the other members of its family. As assessed by RNA blot, reverse transcriptase-polymerase chain reaction, and/or in situ hybridization analysis, mT4 is expressed in interleukin-5-dependent mouse eosinophils, as well as in ovaries and testes. The observation that recombinant mT4 is preferentially retained in the endoplasmic reticulum of transiently transfected COS-7 cells suggests a convertase-like role for this integral membrane serine protease. PMID- 11259428 TI - PKN regulates phospholipase D1 through direct interaction. AB - The association of phospholipase (PLD)-1 with protein kinase C-related protein kinases, PKNalpha and PKNbeta, was analyzed. PLD1 interacted with PKNalpha and PKNbeta in COS-7 cells transiently transfected with PLD1 and PKNalpha or PKNbeta expression constructs. The interactions between endogenous PLD1 and PKNalpha or PKNbeta were confirmed by co-immunoprecipitation from mammalian cells. In vitro binding studies using the deletion mutants of PLD1 indicated that PKNalpha directly bound to residues 228-598 of PLD1 and that PKNbeta interacted with residues 1-228 and 228-598 of PLD1. PKNalpha stimulated the activity of PLD1 in the presence of phosphatidylinositol 4,5-bisphosphate in vitro, whereas PKNbeta had a modest effect on the stimulation of PLD1 activity. The stimulation of PLD1 activity by PKNalpha was slightly enhanced by the addition of arachidonic acid. These results suggest that the PKN family functions as a novel intracellular player of PLD1 signaling pathway. PMID- 11259429 TI - Tyrosine-phosphorylated low density lipoprotein receptor-related protein 1 (Lrp1) associates with the adaptor protein SHC in SRC-transformed cells. AB - v-Src transforms fibroblasts in vitro and causes tumor formation in the animal by tyrosine phosphorylation of critical cellular substrates. Exactly how v-Src interacts with these substrates remains unknown. One of its substrates, the adaptor protein Shc, is thought to play a crucial role during cellular transformation by v-Src by linking v-Src to Ras. We used Shc proteins with mutations in either the phosphotyrosine binding (PTB) or Src homology 2 domain to determine that phosphorylation of Shc in v-Src-expressing cells depends on the presence of a functional PTB domain. We purified a 100-kDa Shc PTB-binding protein from Src-transformed cells that was identified as the beta chain of the low density lipoprotein receptor-related protein LRP1. LRP1 acts as an import receptor for a variety of proteins and is involved in clearance of the beta amyloid precursor protein. This study shows that LRP1 is tyrosine-phosphorylated in v-Src-transformed cells and that tyrosine-phosphorylated LRP1 binds in vivo and in vitro to Shc. The association between Shc and LRP1 may provide a mechanism for recruitment of Shc to the plasma membrane where it is phosphorylated by v Src. It is at the membrane that Shc is thought to be involved in Ras activation. These observations further suggest that LRP1 could function as a signaling receptor and may provide new avenues to investigate its possible role during embryonal development and the onset of Alzheimer's disease. PMID- 11259430 TI - Cloning and characterization of novel ficolins from the solitary ascidian, Halocynthia roretzi. AB - Ficolins are animal lectins with collagen-like and fibrinogen-like domains. They are involved in the first line of host defense against pathogens. Human ficolin/P35 as well as mannose-binding lectin (MBL) activates the complement lectin pathway in association with MBL-associated serine proteases. To elucidate the origin and evolution of ficolins, we separated approximately 40 kDa (p40) and approximately 50 kDa (p50) N-acetylglucosamine-binding lectins from hemolymph plasma of the solitary ascidian. Binding assays revealed that p40 recognizes N acetyl groups in association with a pyranose ring and that p50 recognizes N acetylglucosamine alone. Based on the amino acid sequences of the proteins, we isolated two clones each of p40 and p50 from the ascidian hepatopancreas cDNA and determined the entire coding sequences of these clones. Because all of the clones contained both collagen-like and fibrinogen-like domains, we concluded that these were homologs of the mammalian ficolin family and designated ascidian ficolins (AsFCNs). The fibrinogen-like domain of the AsFCNs shows 45.4-52.4% amino acid sequence identity with the mammalian ficolin family. A phylogenetic tree of the fibrinogen-like sequences shows that all the fibrinogen-like domains may have evolved from a common ancestor that branched off an authentic fibrinogen. These results suggest that AsFCNs play an important role with respect to ascidian hemolymph lectin activity and the correlation of different functions with binding specificity. PMID- 11259431 TI - Transforming growth factor-beta and ciliary neurotrophic factor synergistically induce vasoactive intestinal peptide gene expression through the cooperation of Smad, STAT, and AP-1 sites. AB - The cytokine ciliary neurotrophic factor (CNTF) and transforming growth factor beta (TGF-beta) both induce transcription of the vasoactive intestinal peptide (VIP) gene through a 180-base pair cytokine response element (CyRE) in the VIP promoter. While CNTF induces STAT and AP-1 proteins to bind to cognate sites in the VIP CyRE, the mechanism through which TGF-beta acts to induce VIP gene transcription is not known. Here we show that Smad3 and Smad4 proteins can bind to two distinct sites within the VIP CyRE. These sites are absolutely required for the induction of VIP CyRE transcription by TGF-beta. TGF-beta induces endogenous Smad-containing complexes to bind to these sites in human neuroblastoma cells. CNTF and TGF-beta synergize to induce VIP mRNA expression and transcription through the VIP CyRE. This synergy is dependent on the Smad, STAT, and AP-1 sites, suggesting that these two independent cytokine pathways synergize through the cooperation of pathway-specific transcription factors binding to distinct sites within the VIP CyRE. PMID- 11259432 TI - IgE receptor type I-dependent tyrosine phosphorylation of phospholipid scramblase. AB - To identify new effectors of IgE receptor (FcepsilonRI) signaling, we purified proteins from FcepsilonRI-stimulated RBL-2H3 rat mast cells on anti phosphotyrosine beads and generated mouse monoclonal antibodies (mAb) against these proteins. Two mAbs bound to a protein that was identified as a new isoform of phospholipid scramblase (PLSCR) after screening an RBL-2H3 cDNA expression library. This isoform differed from PLSCR1 by the absence of an exon 3-encoded sequence and by an insert coding six QGPY(P/A)GP repeats. The PLSCR family of proteins is responsible for a redistribution of phospholipids across the plasma membrane. Although rat PLSCR is a 37-kDa protein, anti-phosphotyrosine immunoblots revealed the presence of 37-49 kDa phosphoproteins in the material immunoprecipitated with either anti-PLSCR mAb but not with unrelated monoclonal or polyclonal antibodies. Depletion of PLSCR resulted in the absence of these phosphoproteins. Additional experiments led to the identification of these phosphoproteins as phospho-PLSCR itself. Stimulation of RBL-2H3 cells upon FcepsilonRI engagement resulted in a dramatic increase in PLSCR tyrosine phosphorylation. A comparison of the relative amounts of phospho-PLSCR and nonphosphorylated PLSCR demonstrated that only a tiny fraction was thus modified, indicating a finely targeted involvement of PLSCR in FcepsilonRI signaling. Thus, this study reports the cloning of a new isoform of PLSCR, as well as the first observation that a member of the PLSCR family is a target for tyrosine kinases and is involved in signaling by an immune receptor. These findings open new perspectives on the role of phospholipid scramblases and to the mechanisms involved in their regulation. PMID- 11259433 TI - Rad23 provides a link between the Png1 deglycosylating enzyme and the 26 S proteasome in yeast. AB - In addition to a role in DNA repair events in yeast, several lines of evidence indicate that the Rad23 protein (Rad23p) may regulate the activity of the 26 S proteasome. We report evidence that a de-N-glycosylating enzyme, Png1p, may be involved in the proteasomal degradation pathway via its binding to Rad23p. Interaction of Rad23p and Png1p was first detected by two-hybrid screening, and this interaction in vivo was confirmed by biochemical analyses. The Png1p-Rad23p complex was shown to be distinct from the well established DNA repair complex, Rad4p-Rad23p. We propose a model in which Rad23p functions as an escort protein to link the 26 S proteasome with proteins such as Rad4p or Png1p to regulate their cellular activities. PMID- 11259434 TI - Oxyanion binding alters conformation and quaternary structure of the c-terminal domain of the transcriptional regulator mode. Implications for molybdate dependent regulation, signaling, storage, and transport. AB - The molybdate-dependent transcriptional regulator ModE of Escherichia coli functions as a sensor of intracellular molybdate concentration and a regulator for the transcription of several operons that control the uptake and utilization of molybdenum. We present two high-resolution crystal structures of the C terminal oxyanion-binding domain in complex with molybdate and tungstate. The ligands bind between subunits at the dimerization interface, and analysis reveals that oxyanion selectivity is determined primarily by size. The relevance of the structures is indicated by fluorescence measurements, which show that the oxyanion binding properties of the C-terminal domain of ModE are similar to those of the full-length protein. Comparisons with the apoprotein structure have identified structural rearrangements that occur on binding oxyanion. This molybdate-dependent conformational switch promotes a change in shape and alterations to the surface of the protein and may provide the signal for recruitment of other proteins to construct the machinery for transcription. Sequence and structure-based comparisons lead to a classification of molybdate binding proteins. PMID- 11259435 TI - The pyrimidine ring-opened derivative of 1,N6-ethenoadenine is excised from DNA by the Escherichia coli Fpg and Nth proteins. AB - It was previously shown that 1,N(6)-ethenoadenine (epsilonA) in DNA rearranges into a pyrimidine ring-opened derivative of 20-fold higher mutagenic potency in Escherichia coli (AB1157 lacDeltaU169) than the parental epsilonA (Basu, A. K., Wood, M. L., Niedernhofer, L. J., Ramos, L. A., and Essigmann, J. M. (1993) Biochemistry 32, 12793-12801). We have found that at pH 7.0, the stability of the N-glycosidic bond in epsilondA is 20-fold lower than in dA. In alkaline conditions, but also at neutrality, epsilondA depurinates or converts into products: epsilondA --> B --> C --> D. Compound B is a product of water molecule addition to the C(2)-N(3) bond, which is in equilibrium with a product of N(1) C(2) bond rupture in epsilondA. Compound C is a deformylated derivative of ring opened compound B, which further depurinates yielding compound D. Ethenoadenine degradation products are not recognized by human N-alkylpurine-DNA glycosylase, which repairs epsilonA. Product B is excised from oligodeoxynucleotides by E. coli formamidopyrimidine-DNA glycosylase (Fpg) and endonuclease III (Nth). Repair by the Fpg protein is as efficient as that of 7,8-dihydro-8-oxoguanine when the excised base is paired with dT and dC but is less favorable when paired with dG and dA. Ethenoadenine rearrangement products are formed in oligodeoxynucleotides also at neutral pH at the rate of about 2-3% per week at 37 degrees C, and therefore they may contribute to epsilonA mutations. PMID- 11259436 TI - Akt stimulates the transactivation potential of the RelA/p65 Subunit of NF-kappa B through utilization of the Ikappa B kinase and activation of the mitogen activated protein kinase p38. AB - The serine/threonine kinase Akt/PKB is a potent regulator of cell survival and has oncogenic transformation potential. Previously, it has been shown that Akt can activate the transcription factor NF-kappaB and that this functions to block apoptosis induced by certain stimuli. The mechanism whereby Akt activates NF kappaB has been controversial, with evidence supporting induction of nuclear translocation of NF-kappaB via activation of IkappaB kinase activity and/or the stimulation of the transcription function of NF-kappaB. Here we demonstrate that Akt targets the transactivation function of NF-kappaB by stimulating the transactivation domain of RelA/p65 in a manner that is dependent on IkappaB kinase beta activity and on the mitogen-activated protein kinase p38 (p38). Activation of RelA/p65 transactivation function requires serines 529 and 536, sites shown previously to be inducibly phosphorylated. Consistent with the requirement of p38 in the activation of NF-kappaB transcriptional function, expression of activated Akt induces p38 activity. Furthermore, the ability of IL 1beta to activate NF-kappaB is known to involve Akt, and we show here that IL 1beta induces p38 activity in manner dependent on Akt and IkappaB kinase activation. Interestingly, activated Akt and the transcriptional co-activators CBP/p300 synergize in the activation of the RelA/p65 transactivation domain, and this synergy is blocked by p38 inhibitors. These studies demonstrate that Akt, functioning through IkappaB kinase and p38, induces the transcription function of NF-kappaB by stimulating the RelA/p65 transactivation subunit of NF-kappaB. PMID- 11259437 TI - Molecular mechanism of tumor necrosis factor-alpha production in 1-->3-beta glucan (zymosan)-activated macrophages. AB - The molecular details of 1-->3-beta-glucans, a fungal cell wall component, induced inflammatory responses are not well understood. In the present study, we conducted a systematic analysis of the molecular events leading to tumor necrosis factor (TNF)-alpha production after glucan stimulation of macrophages. We demonstrated that activation of nuclear factor kappaB (NF-kappaB) is essential in zymosan A (a source of 1-->3-beta-glucans)-induced TNF-alpha production in macrophages (RAW264.7 cells). Zymosan A-induced TNF-alpha protein production was associated with an increase in the TNF-alpha gene promoter activity. Activation of the TNF-alpha gene promoter was dependent on activation of NF-kappaB. Time course studies indicated that DNA binding activity of NF-kappaB preceded TNF alpha promoter activity. Inhibition of NF-kappaB activation led to a dramatic reduction in both TNF-alpha promoter activity and TNF-alpha protein production in the response to zymosan A. Mutation of a major NF-kappaB binding site (kappa3) in the gene promoter resulted in a significant decrease in the induction of the gene promoter by zymosan A, while mutation of Egr or CRE sites failed to inhibit the response to zymosan. Together, these results strongly suggest that NF-kappaB is involved in signal transduction of 1-->3-beta-glucans-induced TNF-alpha expression. PMID- 11259438 TI - Abnormal regulation of photosynthetic electron transport in a chloroplast ycf9 inactivation mutant. AB - The ycf9 (orf62) gene of the plastid genome encodes a 6.6-kDa protein (ORF62) of thylakoid membranes. To elucidate the role of the ORF62 protein, the coding region of the gene was disrupted with an aadA cassette, yielding mutant plants that were nearly (more than 95%) homoplasmic for ycf9 inactivation. The ycf9 mutant had no altered phenotype under standard growth conditions, but its growth rate was severely reduced under suboptimal irradiances. On the other hand, it was less susceptible to photodamage than the wild type. ycf9 inactivation resulted in a clear reduction in protein amounts of CP26, the NAD(P)H dehydrogenase complex, and the plastid terminal oxidase. Furthermore, depletion of ORF62 led to a faster flow of electrons to photosystem I without a change in the maximum electron transfer capacity of photosystem II. Despite the reduction of CP26 in the mutant thylakoids, no differences in PSII oxygen evolution rates were evident even at low light intensities. On the other hand, the ycf9 mutant presented deficiencies in the capacity for PSII-independent electron transport (ferredoxin-dependent cyclic electron transport and NAD(P)H dehydrogenase-mediated plastoquinone reduction). Altogether, it is shown that depletion of ORF62 leads to anomalies in the photosynthetic electron transfer chain and in the regulation of electron partitioning among the different routes of electron transport. PMID- 11259439 TI - Oxidant tone regulates RANTES gene expression in airway epithelial cells infected with respiratory syncytial virus. Role in viral-induced interferon regulatory factor activation. AB - Respiratory syncytial virus (RSV) produces intense pulmonary inflammation, in part, through its ability to induce chemokine synthesis in infected airway epithelial cells. RANTES (regulated upon activation, normal T-cells expressed and secreted) is a CC chemokine which recruits and activates monocytes, lymphocytes, and eosinophils, all cell types present in the lung inflammatory infiltrate induced by RSV infection. In this study we investigated the role of reactive oxygen species in the induction of RANTES gene expression in human type II alveolar epithelial cells (A549), following RSV infection. Our results indicate that RSV infection of airway epithelial cells rapidly induces reactive oxygen species production, prior to RANTES expression, as measured by oxidation of 2',7' dichlorofluorescein. Pretreatment of airway epithelial cells with the antioxidant butylated hydroxyanisol (BHA), as well a panel of chemically unrelated antioxidants, blocks RSV-induced RANTES gene expression and protein secretion. This effect is mediated through the ability of BHA to inhibit RSV-induced interferon regulatory factor binding to the RANTES promoter interferon-stimulated responsive element, that is absolutely required for inducible RANTES promoter activation. BHA inhibits de novo interferon regulator factor (IRF)-1 and -7 gene expression and protein synthesis, and IRF-3 nuclear translocation. Together, these data indicates that a redox-sensitive pathway is involved in RSV-induced IRF activation, an event necessary for RANTES gene expression. PMID- 11259440 TI - Molecular cloning and characterization of Bif-1. A novel Src homology 3 domain containing protein that associates with Bax. AB - Bax is a proapoptotic member of the Bcl-2 protein family that commits the cell to undergo programmed cell death in response to apoptotic stimuli. To gain further insights into Bax mechanisms, we have identified a novel Bax-binding protein, termed Bif-1, by using a yeast two-hybrid cloning technique. Bif-1 is an evolutionarily conserved cytoplasmic protein that contains a predicted Src homology 3 (SH3) domain located near its C terminus but shares no significant homology with members of the Bcl-2 family. A Northern blot analysis indicates that Bif-1 is expressed in most tissues with abundant expression in heart and skeletal muscle. Bif-1 is capable of interacting with Bax as demonstrated by yeast two-hybrid, coimmunoprecipitation, and immunofluorescence studies. Induction of apoptosis in murine pre-B hematopoietic cells FL5.12 by interleukin 3 withdrawal results in increased association of Bax with Bif-1, which is accompanied by a conformational change in the Bax protein. Overexpression of Bif 1 promotes Bax conformational change, caspase activation, and apoptotic cell death in FL5.12 cells following interleukin-3 deprivation. Bif-1 thus represents a new type of regulator of Bax-mediated signaling pathways for apoptosis. PMID- 11259441 TI - Bcl-xL promotes the open configuration of the voltage-dependent anion channel and metabolite passage through the outer mitochondrial membrane. AB - The diffusion of metabolites across the outer mitochondrial membrane is essential for coupled cellular respiration. The outer membrane of mitochondria isolated from growth factor-deprived cells is impaired in its ability to exchange metabolic anions. When added to mitochondria, recombinant Bcl-x(L) restores metabolite exchange across the outer membrane without inducing the loss of cytochrome c from the intermembrane space. Restoration of outer membrane permeability to anionic metabolites does not occur directly through Bcl-x(L) ion channels. Instead, recombinant Bcl-x(L) maintains the outer mitochondrial membrane channel, VDAC, in an open configuration. Consistent with these findings, when ADP-induced oxidative phosphorylation is limited by exogenous beta-NADH, recombinant Bcl-x(L) can sustain outer mitochondrial membrane permeability to ADP. beta-NADH limits respiration by promoting the closed configuration of VDAC. Together these results demonstrate that following an apoptotic signal, Bcl-x(L) can maintain metabolite exchange across the outer mitochondrial membrane by inhibiting VDAC closure. PMID- 11259442 TI - A role for the alpha 113 (GH1) amino acid residue in the polymerization of sickle hemoglobin. Evaluation of its inhibitory strength and interaction linkage with two fiber contact sites (alpha 16/23) located in the AB region of the alpha chain. AB - A cluster of amino acid residues located in the AB-GH region of the alpha-chain are shown in intra-double strand axial interactions of the hemoglobin S (HbS) polymer. However, alphaLeu-113 (GH1) located in the periphery is not implicated in any interactions by either crystal structure or models of the fiber, and its role in HbS polymerization has not been explored by solution experiments. We have constructed HbS Twin Peaks (betaGlu-6-->Val, alphaLeu-113-->His) to ascertain the hitherto unknown role of the alpha113 site in the polymerization process. The structural and functional behavior of HbS Twin Peaks was comparable with HbS. HbS Twin Peaks polymerized with a slower rate compared with HbS, and its polymer solubility (C(sat)) was found to be about 1.8-fold higher than HbS. To further authenticate the participation of the alpha113 site in the polymerization process as well as to evaluate its relative inhibitory strength, we constructed HbS tetramers in which the alpha113 mutation was coupled individually with two established fiber contact sites (alpha16 and alpha23) located in the AB region of the alpha-chain: HbS(alphaLys-16-->Gln, alphaLeu-113-->His), HbS(alphaGlu-23- >Gln, alphaLeu-113-->His). The single mutants at alpha16/alpha23 sites were also engineered as controls. The C(sat) values of the HbS point mutants involving sites alpha16 or alpha23 were higher than HbS but markedly lower as compared with HbS Twin Peaks. In contrast, C(sat) values of both double mutants were comparable with or higher than that of HbS Twin Peaks. The demonstration of the inhibitory effect of alpha113 mutation alone or in combination with other sites, in quantitative terms, unequivocally establishes a role for this site in HbS gelation. These results have implications for development of a more accurate model of the fiber that could serve as a blueprint for therapeutic intervention. PMID- 11259443 TI - Card10 is a novel caspase recruitment domain/membrane-associated guanylate kinase family member that interacts with BCL10 and activates NF-kappa B. AB - BCL10 belongs to the caspase recruitment domain (CARD) family of proteins that regulate apoptosis and NF-kappaB signaling pathways. Analysis of BCL10-deficient mice has revealed that BCL10 mediates NF-kappaB activation by antigen receptors in B and T cells. We recently identified a subclass of CARD proteins (CARD9, CARD11, and CARD14) that may function to connect BCL10 to multiple upstream signaling pathways. We report here that CARD10 is a novel BCL10 interactor that belongs to the membrane-associated guanylate kinase family, a class of proteins that function to organize signaling complexes at plasma membranes. When expressed in cells, CARD10 binds to BCL10 and signals the activation of NF-kappaB through its N-terminal effector CARD domain. We propose that CARD10 functions as a molecular scaffold for the assembly of a BCL10 signaling complex that activates NF-kappaB. PMID- 11259444 TI - The vesiculo-vacuolar organelle (VVO). A new endothelial cell permeability organelle. AB - A newly defined endothelial cell permeability structure, termed the vesiculo vacuolar organelle (VVO), has been identified in the microvasculature that accompanies tumors, in venules associated with allergic inflammation, and in the endothelia of normal venules. This organelle provides the major route of extravasation of macromolecules at sites of increased vascular permeability induced by vascular permeability factor/vascular endothelial growth factor (VPF/VEGF), serotonin, and histamine in animal models. Continuity of these large sessile structures between the vascular lumen and the extracellular space has been demonstrated in kinetic studies with ultrastructural electron-dense tracers, by direct observation of tilted electron micrographs, and by ultrathin serial sections with three-dimensional computer reconstructions. Ultrastructural enzyme affinity cytochemical and immunocytochemical studies have identified histamine and VPF/VEGF bound to VVOs in vivo in animal models in which these mediators of permeability are released from mast cells and tumor cells, respectively. The high affinity receptor for VPF/VEGF, VEGFR-2, was localized to VVOs and their substructural components by pre-embedding ultrastructural immunonanogold and immunoperoxidase techniques. Similar methods were used to localize caveolin and vesicle-associated membrane protein (VAMP) to VVOs and caveolae, indicating a possible commonality of formation and function of VVOs to caveolae. PMID- 11259445 TI - Quantitation of AgNORs by flow versus image cytometry. AB - AgNORs are nucleolar proteins that interact specifically with silver salts. The size of silver precipitates measured by image analysis (ICM) in cycling cells proved to be inversely proportional to the cell cycle time and provided a significant correlation with prognosis for a large spectrum of cancers. Because ICM is time-consuming and poorly reproducible among laboratories using different imaging settings, this article presents a new approach to AgNOR quantitation based on flow cytometry (FCM). We report that silver precipitates caused a great decrease in the forward scattered light and that this effect was correlated with the AgNOR's relative area as measured by ICM. These results were confirmed by measuring cell lines having different cell cycle durations. Moreover, double staining using APase-Fast red fluorescence to reveal the Ki-67/MIB 1 antigen of cycling cells and silver nitrate to stain the AgNORs was successfully analyzed by FCM. The procedure makes it possible, for the first time, to validly and rapidly compare the growth fraction and cycling speed of partially proliferating cell populations, such as tumors. PMID- 11259446 TI - Immunohistochemical distribution of CD9, heparin binding epidermal growth factor like growth factor, and integrin alpha3beta1 in normal human tissues. AB - The tetra-membrane-spanning protein CD9 forms a complex with a membrane-anchored heparin binding epidermal growth factor-like growth factor (HB-EGF) and integrin alpha3beta1 in some human and monkey cell lines. We show here the immunohistochemical distribution of CD9, HB-EGF, and integrin alpha3beta1 in normal human tissues. Distribution of CD9, HB-EGF, and integrin alpha3beta1 was similar in various tissues, including transitional epithelium, squamous epithelium, thyroid follicular epithelium, adrenal cortex, testis, smooth muscle, and stromal fibrous tissue. However, distribution of the three proteins did not coincide in some tissues, such as lung, liver, kidney, gastric and intestinal epithelium, pancreas, salivary gland, and ovary. In striated muscle, including cardiac muscle, CD9 was present not in the muscle cells themselves but in the endomysium and perimysium, whereas HB-EGF was distributed in the muscle cells themselves. CD9 was distributed in the myelin, but HB-EGF was found in the axon of the peripheral and central nervous systems. Coincident distribution of integrin alpha3beta1 with others was not observed in muscles and neural tissues. In conclusion, there is a possibility of complex formation and functional cooperation of CD9 with HB-EGF and/or integrin alpha3beta1 in several tissues. PMID- 11259447 TI - Calmodulin distribution and the actomyosin cytoskeleton in Toxoplasma gondii. AB - The gliding motility of the protozoan parasite Toxoplasma gondii and its invasion of cells are powered by an actin-myosin motor. We have studied the spatial distribution and relationship between these two cytoskeleton proteins and calmodulin (CaM), the Ca(2+)-dependent protein involved in invasion by T. gondii. A 3D reconstruction using labeling and tomographic studies showed that actin was present as a V-like structure in the conoidal part of the parasite. The myosin distribution overlapped that of actin, and CaM was concentrated at the center of the apical pole. We demonstrated that the actomyosin network, CaM, and myosin light-chain kinases are confined to the apical pole of the T. gondii tachyzoite. MLCK could act as an intermediate molecule between CaM and the cytoskeleton proteins. We have developed a model of the organization of the actomyosin-CaM complex and the steps of a signaling pathway for parasite motility. PMID- 11259448 TI - MyoD and myogenin expression patterns in cultures of fetal and adult chicken myoblasts. AB - Isolated chicken myoblasts had previously been utilized in many studies aiming at understanding the emergence and regulation of the adult myogenic precursors (satellite cells). However, in recent years only a small number of chicken satellite cell studies have been published compared to the increasing number of studies with rodent satellite cells. In large part this is due to the lack of markers for tracing avian myogenic cells before they become terminally differentiated and express muscle-specific structural proteins. We previously demonstrated that myoblasts isolated from fetal and adult chicken muscle display distinct schedules of myosin heavy-chain isoform expression in culture. We further showed that myoblasts isolated from newly hatched and young chickens already possess the adult myoblast phenotype. In this article, we report on the use of polyclonal antibodies against the chicken myogenic regulatory factor proteins MyoD and myogenin for monitoring fetal and adult chicken myoblasts as they progress from proliferation to differentiation in culture. Fetal-type myoblasts were isolated from 11-day-old embryos and adult-type myoblasts were isolated from 3-week-old chickens. We conclude that fetal myoblasts express both MyoD and myogenin within the first day in culture and rapidly transit into the differentiated myosin-expressing state. In contrast, adult myoblasts are essentially negative for MyoD and myogenin by culture Day 1 and subsequently express first MyoD and then myogenin before expressing sarcomeric myosin. The delayed MyoD-to-myogenin transition in adult myoblasts is accompanied by a lag in the fusion into myotubes, compared to fetal myoblasts. We also report on the use of a commercial antibody against the myocyte enhancer factor 2A (MEF2A) to detect terminally differentiated chicken myoblasts by their MEF2+ nuclei. Collectively, the results support the hypothesis that fetal and adult myoblasts represent different phenotypic populations. The fetal myoblasts may already be destined for terminal differentiation at the time of their isolation, and the adult myoblasts may represent progenitors that reside in an earlier compartment of the myogenic lineage. PMID- 11259449 TI - Immunolocalization of anion exchanger AE2, Na(+)/H(+) exchangers NHE1 and NHE4, and vacuolar type H(+)-ATPase in rat pancreas. AB - We have studied the expression and localization of several H(+) and HCO(3)(-) transporters, whose presence in the rat pancreas is still unclear. The Cl( )/HCO(3)(-) exchanger AE2, the Na(+)/H(+) exchangers NHE1 and NHE4, and the 31-kD and 70-kD vacuolar H(+)-ATPase (V-ATPase) subunits were detected by immunoblotting and immunocytochemical techniques. Immunoblotting of plasma membranes with transporter-specific antibodies revealed protein bands at approximately 160 kD for AE2, at approximately 90 kD and approximately 103 kD for NHE1 and NHE4, respectively, and at 31 kD and 70 kD for V-ATPase. NHE1 and NHE4 were further identified by amplification of isoform-specific cDNA using RT-PCR. Immunohistochemistry revealed a basolateral location of AE2, NHE1, and NHE4 in acinar cells. In ducts, NHE1 and NHE4 were basolaterally located but no AE2 expression was detected. V-ATPase was detected in zymogen granules (ZGs) by immunogold labeling, and basolaterally in duct cells by immunohistochemistry. The data indicate that NHE1 and NHE4 are co-expressed in rat pancreatic acini and ducts. Basolateral acinar AE2 could contribute to Cl(-) uptake and/or pH regulation. V-ATPase may be involved in ZG fusion/exocytosis and ductal HCO(3)(-) secretion. The molecular identity of the ductal Cl(-)/HCO(3)(-) exchanger remains unclear. PMID- 11259450 TI - Antigen retrieval of basement membrane proteins from archival eye tissues. AB - Antigen retrieval (AR) methods can unmask tissue antigens that have been altered by fixation, processing, storage, or resin interactions. This is particularly important in the study of archival tissues, because primary fixatives and storage times may vary among specimens. We performed an electron microscopic study of basement membrane components of the aqueous humor drainage pathways from archival eye tissue. AR (heated citrate buffer, pH 6.0, LR White resin) increased the amount of label of collagen IV and fibronectin in tissue fixed in four different fixatives, including those containing glutaraldehyde. Labeling density was approximately doubled after AR for most fixatives, with the largest increase for tissues fixed in 4% paraformaldehyde/2% glutaraldehyde. Duration of storage time for archival tissues did not affect AR results. AR did not change the components of the extracellular matrix labeled; no "new" components were labeled after AR. We conclude that AR in citrate buffer can be used on selected extracellular matrix antigens to enhance label that would otherwise be lost due to fixation and storage. PMID- 11259451 TI - Selective processing of chromogranin A in the different islet cells in human pancreas. AB - We studied the immunoreactivity of 12 different region-specific antibodies to the chromogranin A (CgA) molecule in the four major neuroendocrine cell types of the human pancreas by using double immunofluorescence techniques. The antibodies raised to the N-terminal and midportions of CgA showed, on the whole, stronger immunoreactivity than did the C-terminal antibodies, with a few exceptions. Often the immunoreactivity was stronger in glucagon cells. Insulin cells expressed immunoreactivity to all region-specific antibodies, but glucagon cells were nonreactive to two antibodies. Somatostatin cells reacted only with the C terminal antibodies (amino acid sequences CgA 411-424), while PP cells were stained with four CgA region-specific antibodies between amino acid sequences 63 195. The cause of these differences may be that the CgA molecule is cleaved, partly masked, or partly translated from CgA mRNA. Microwave treatment improved only the staining with the CgA 361-372 antibodies, which indicates that masking is not the sole or entire cause. Our findings may indicate that the CgA molecule is cleaved in different ways in the various pancreatic endocrine cell types, giving rise to a variety of biologically functional fragments. PMID- 11259452 TI - Differential expression of a stress-modulating gene, BRE, in the adrenal gland, in adrenal neoplasia, and in abnormal adrenal tissues. AB - Genes that modulate the action of hormones and cytokines play a critical role in stress response, survival, and in growth and differentiation of cells. Many of these biological response modifiers are responsible for various pathological conditions, including inflammation, infection, cachexia, aging, genetic disorders, and cancer. We have previously identified a new gene, BRE, that is responsive to DNA damage and retinoic acid. Using multiple-tissue dot-blotting and Northern blotting, BRE was recently found to be strongly expressed in adrenal cortex and medulla, in testis, and in pancreas, whereas low expression was found in the thyroid, thymus, small intestine and stomach. In situ hybridization and immunohistochemical staining indicated that BRE was strongly expressed in the zona glomerulosa of the adrenal cortex, which synthesizes and secretes the mineralocorticoid hormones. It is also highly expressed in the glial and neuronal cells of the brain and in the round spermatids, Sertoli cells, and Leydig cells of the testis, all of which are associated with steroid hormones and/or TNF synthesis. However, BRE expression was downregulated in human adrenal adenoma and pheochromocytoma, whereas its expression was enhanced in abnormal adrenal tissues of rats chronically treated with nitrate or nitrite. These data, taken together, indicate that the expression of BRE is apparently associated with steroids and/or TNF production and the regulation of endocrine functions. BRE may play an important role in the endocrine and immune system, such as the cytokine-endocrine interaction of the adrenal gland. PMID- 11259453 TI - Differential distribution of sialic acid in alpha2,3 and alpha2,6 linkages in the apical membrane of cultured epithelial cells and tissues. AB - We used lectin cytochemistry and confocal microscopy to examine the distribution of sialic acid in epithelial cells. Maackia amurensis lectin and Sambuccus nigra agglutinin were used to detect alpha2,3 and alpha2,6 sialic acid, respectively. In Caco-2, HT-29 5M12, and MCF-7 cells, which express sialic acid mainly in one type of linkage, the majority of the signal was observed in the apical membrane. In cells that bound both lectins, alpha2,3 sialic acid was distributed apically, whereas alpha2,6 sialic acid showed a broader distribution. In IMIM-PC-1 cultures, alpha2,3 sialic acid was detected mainly in the apical membrane, whereas alpha2,6 sialic acid was more abundant in the basolateral domain of polarized cells. In these cells, treatment with GalNAc-O-benzyl led to reduced alpha2,3 levels and to an increase and redistribution of alpha2,6 to the apical domain. Similarly, sialic acid was predominantly expressed apically in all epithelial tissues examined. In conclusion, (a) sialic acid is mainly distributed to the apical membrane of epithelial cells, (b) there is a hierarchy in the distribution of sialic acids in polarized epithelial cells, i.e., alpha2,3 is preferred to alpha2,6 in the apical membrane, and (c) IMIM-PC-1 cells are a good model in which to study the regulation of the levels and distribution of sialic acids. PMID- 11259454 TI - Aldehyde fixation of thiol-reactive fluorescent cytoplasmic probes for tracking cell migration. AB - Tracking of cell migration plays an important role in the study of morphogenesis, inflammation, and metastasis. The recent development of probes that exist as intracellular peptide-fluorescence dye adducts has offered the possibility of aldehyde fixation of these dyes for detailed anatomic studies of lymphocyte trafficking. To define the conditions for fixation of these cytoplasmic fluorescent probes, we compared fixation conditions containing formaldehyde, glutaraldehyde, paraformaldehyde, zinc formaldehyde, and glyoxylate, as well as fixation by quick-freezing in liquid nitrogen-cooled methylbutane. The efficacy of aldehyde fixation of the cell fluorescence was assessed by quantitative tissue cytometry and flow cytometry. We studied cytoplasmic fluorescent dyes with discrete emissions in the green [5-chloromethylfluorescein diacetate (CMFDA); 492 ex, 516 em] and orange [5-(and-6)-(4-chloromethyl(benzoyl)amino) tetramethylrhodamine (CMTMR); 540 ex, 566 em] spectra. The results demonstrated that aldehyde fixation preserved cell fluorescence for more than 6 months. The primary difference between the aldehyde fixatives was variability in the difference between the yield of the cell fluorescence and the relevant background fluorescence. Formaldehyde and paraformaldehyde were superior to the other fixatives in preserving cell fluorescence while limiting background fluorescence. With these fixatives, both the CMFDA and CMTMR fluorescent dyes permitted sufficient anatomic resolution for reliable localization in long-term cell tracking studies. PMID- 11259455 TI - Identification and purification of functional human beta-cells by a new specific zinc-fluorescent probe. AB - Pancreatic beta-cells contain large amounts of zinc. We took advantage of this to try to localize, quantify, and isolate insulin-producing cells from islet preparations. Our study was designed to identify a non-toxic zinc-sensitive fluorescent probe able to selectively label labile zinc in viable beta-cells and to exhibit excitation and emission wavelengths in the visible spectrum, making this technique exploitable by most instruments. We tested Newport Green, a probe excitable at 485 nm with a dissociation constant in the micromolar range corresponding to a low affinity for zinc. The loading of the lipophilic esterified form of Newport Green was easy, rapid, specific, and non-toxic to cells. Confocal microscopy highlighted an intense fluorescence associated with secretory granules. Regression analyses showed a good relationship between zinc fluorescence and islet number (r = 0.98) and between zinc fluorescence and insulin content (r = 0.81). The determination of Zn fluorescence per DNA enabled us to assess the quality of the different islet preparations intended for islet allografting in terms of both purity and viability. Cell sorting of dissociated Newport Green-labeled cells resulted in a clear separation of beta-cells, as judged by insulin content per DNA and immunocytochemical analysis. This zinc probe, the first able to specifically label living cells in the visible spectrum, appears very promising for beta-cell experimentation, both clinically and for basic research. PMID- 11259456 TI - delta- and gamma-Sarcoglycan localization in the sarcoplasmic reticulum of skeletal muscle. AB - Sarcoglycans are transmembrane proteins that are members of the dystrophin complex. Sarcoglycans cluster together to form a complex, which is localized in the cell membrane of skeletal, cardiac, and smooth muscle fibers. However, it is still unclear whether or not sarcoglycans are restricted to the sarcolemma. To address this issue, we examined alpha-, beta-, delta-, and gamma-sarcoglycan expression in femoral skeletal muscle from control and dystrophin-deficient mice and rats using confocal microscopy and immunoelectron microscopy. Confocal microscopy of the tissues in cross-section showed that all sarcoglycans were detected under the sarcolemma in rats and control mice. delta- and gamma sarcoglycan labeling demonstrated striations in the longitudinal section, suggesting that the proteins were expressed in the sarcoplasmic reticulum (SR) or transverse tubules (T-tubules). Moreover, such striations of both sarcoglycans were recognized in the dystrophin-deficient mouse skeletal muscle. Double labeling with phalloidin or alpha-actinin and delta- or gamma-sarcoglycan showed different labeling patterns, indicating that delta-sarcoglycan localization was distinct from that of gamma-sarcoglycan. Immunoelectron microscopy clarified that delta-sarcoglycan was localized in the terminal cisternae of the SR, while gamma sarcoglycan was found in the terminal cisternae and longitudinal SR over I-bands but not over A-bands. These data demonstrate that delta- and gamma-sarcoglycans are components of the SR in skeletal muscle, suggesting that both sarcoglycans function independent of the dystrophin complex in the SR. PMID- 11259457 TI - Comparative analysis of galectins in primary tumors and tumor metastasis in human pancreatic cancer. AB - Galectins are galactoside-binding proteins that exhibit an important function in tumor progression by promoting cancer cell invasion and metastasis formation. Using Northern blotting and Western blotting analysis, in situ hybridization (ISH), and immunohistochemistry (IHC), we studied galectin-1 and galectin-3 in tissue samples of 33 primary pancreatic cancers and in tumor metastases in comparison to 28 normal pancreases. Furthermore, the molecular findings were correlated with the clinical and histopathological parameters of the patients. Northern blotting and Western blotting analysis showed significantly higher galectin-1 and galectin-3 mRNA and protein levels in pancreatic cancer samples than in normal controls. For galectin-1, no ISH signals and immunoreactivity were observed in acinar or ductal cells in the normal pancreas and in pancreatic cancer cells, whereas fibroblasts and extracellular matrix cells around the cancer mass exhibited strong mRNA signals and immunoreactivity. Galectin-3 mRNA signals and immunoreactivity were strongly present in most pancreatic cancer cells, whereas in the normal controls only faint ISH and IHC signals were seen in some ductal cells. Metastatic pancreatic cancer cells exhibited moderate to strong galectin-3 immunoreactivity but were negative for galectin-1. No relationship between the galectin-1 and galectin-3 mRNA levels and the tumor stage or between the IHC staining score and the tumor stage was found. However, galectin-1 mRNA levels and the IHC staining score were significantly higher in poorly differentiated tumors compared with well/moderately differentiated tumors, whereas for galectin 3 no differences were found. The expression pattern of galectin-1 and galectin-3 in pancreatic cancer tissues indicates that galectin-1 plays a role in the desmoplastic reaction that occurrs around pancreatic cancer cells, whereas galectin-3 appears to be involved in cancer cell proliferation. High levels of galectin-3 in metastatic cancer cells suggest an impact on metastasis formation. PMID- 11259458 TI - Filling the void: urinary markers for bladder cancer risk and diagnosis. PMID- 11259459 TI - Searching for selective cyclin-dependent kinase inhibitors to target the retinoblastoma/p16 cancer gene pathway. PMID- 11259460 TI - Move over, mouse: make way for the woodchucks, ferrets, and zebrafish. PMID- 11259461 TI - Freeman sets goals for center to reduce cancer health disparities. PMID- 11259462 TI - Robotics and more: new technologies emerge for radiotherapy. PMID- 11259463 TI - Radiotherapy technique tackles challenge of moving target. PMID- 11259464 TI - Dramatic changes unlikely for biomedical research with new Congress. PMID- 11259465 TI - Cancer drugs in pipeline span wide spectrum. PMID- 11259468 TI - Biomarker risk assessment and bladder cancer detection in a cohort exposed to benzidine. AB - BACKGROUND: Cancer screening with highly sensitive, specific biomarkers that reflect molecular phenotypic alterations is an attractive strategy for cancer control. We examined whether biomarker profiles could be used for risk assessment and cancer detection in a cohort of Chinese workers occupationally exposed to benzidine and at risk for bladder cancer. METHODS: The cohort consisted of 1788 exposed and 373 nonexposed workers, followed from 1991 through 1997. We assayed urothelial cells from voided urine samples for DNA ploidy (expressed as the 5C exceeding rate [DNA 5CER]), the bladder tumor-associated antigen p300, and a cytoskeletal protein (G-actin). Workers were stratified into different risk groups (high, moderate, and low risk) at each examination based on a predefined biomarker profile. For workers who developed bladder cancer, tumor risk assessment was analyzed from samples collected 6-12 months before the cancer diagnosis. The associations between risk group and subsequent development of bladder cancer were analyzed by Cox proportional hazards regression analysis and logistic analysis, after adjustment. All statistical tests were two-sided. RESULTS: Twenty-eight bladder cancers were diagnosed in exposed workers and two in nonexposed workers. For risk assessment, DNA 5CER had 87.5% sensitivity, 86.5% specificity, an odds ratio (OR) of 46.2 (95% confidence interval [CI] = 8.1 to 867.0), and a risk ratio (RR) of 16.2 (95% CI = 7.1 to 37.0); p300 had 50.0% sensitivity, 97.9% specificity, an OR of 40.0 (95% CI = 9.0 to 177.8), and an RR of 37.9 (95% CI = 16.8 to 85.3). The risk of developing bladder cancer was 19.6 (95% CI = 8.0 to 47.9) times higher in workers positive for either the DNA 5CER or p300 biomarkers than in workers negative for both biomarkers and 81.4 (95% CI = 33.3 to 199.3) times higher in workers positive for both biomarkers. G-actin was a poor marker of individual risk. CONCLUSIONS: Occupationally exposed workers at risk for bladder cancer can be individually stratified, screened, monitored, and diagnosed based on predefined molecular biomarker profiles. PMID- 11259469 TI - Selective in vivo and in vitro effects of a small molecule inhibitor of cyclin dependent kinase 4. AB - BACKGROUND: Cyclin-dependent kinase 4 (Cdk4) represents a prime target for the treatment of cancer because most human cancers are characterized by overexpression of its activating partner cyclin D1, loss of the natural Cdk4 specific inhibitor p16, or mutation(s) in Cdk4's catalytic subunit. All of these can cause deregulated cell growth, resulting in tumor formation. We sought to identify a small molecule that could inhibit the kinase activity of Cdk4 in vitro and to then ascertain the effects of that inhibitor on cell growth and tumor volume in vivo. METHODS: A triaminopyrimidine derivative, CINK4 (a chemical inhibitor of Cdk4), was identified by screening for compounds that could inhibit Cdk4 enzyme activity in vitro. Kinase assays were performed on diverse human Cdks and on other kinases that were expressed in and purified from insect cells to determine the specificity of CINK4. Cell cycle effects of CINK4 on tumor and normal cells were studied by flow cytometry, and changes in phosphorylation of the retinoblastoma protein (pRb), a substrate of Cdk4, were determined by western blotting. The effect of the inhibitor on tumor growth in vivo was studied by use of tumors established through xenografts of HCT116 colon carcinoma cells in mice. Statistical tests were two-sided. RESULTS: CINK4 specifically inhibited Cdk4/cyclin D1 in vitro. It caused growth arrest in tumor cells and in normal cells and prevented pRb phosphorylation. CINK4 treatment resulted in statistically significantly (P: =.031) smaller mean tumor volumes in a mouse xenograft model. CONCLUSIONS: Like p16, the natural inhibitor of Cdk4, CINK4 inhibits Cdk4 activity in vitro and slows tumor growth in vivo. The specificity of CINK4 for Cdk4 raises the possibility that this small molecule or one with a similar structure could have therapeutic value. PMID- 11259470 TI - Cost comparison of mastectomy versus breast-conserving therapy for early-stage breast cancer. AB - BACKGROUND: Choice of treatment for early-stage breast cancer depends on many factors, including the size and stage of the cancer, the woman's age, comorbid conditions, and perhaps the costs of treatment. We compared the costs of all medical care for women with early-stage breast cancer cases treated by breast conserving therapy (BCT) or mastectomy. METHODS: A total of 1675 women 35 years old or older with incident early-stage breast cancer were identified in a large regional nonprofit health maintenance organization in the period 1990 through 1997. The women were treated with mastectomy only (n = 183), mastectomy with adjuvant hormonal therapy or chemotherapy (n = 417), BCT with radiation therapy (n = 405), or BCT with radiation therapy and adjuvant hormonal therapy or chemotherapy (n = 670). The costs of all medical care for the period 1990 through 1998 were computed for each woman, and monthly costs were analyzed by treatment, adjusting for age and cancer stage. All statistical tests were two-sided. RESULTS: At 6 months after diagnosis, the mean total medical care costs for the four groups differed statistically significantly (P:<.001), with BCT being more expensive than mastectomy. The adjusted mean costs were $12 987, $14 309, $14 963, and $15 779 for mastectomy alone, mastectomy with adjuvant therapy, BCT plus radiation therapy, and BCT plus radiation therapy with adjuvant therapy, respectively. At 1 year, the difference in costs was still statistically significant (P:<.001), but costs were influenced more by the use of adjuvant therapy than by type of surgery. The 1-year adjusted mean costs were $16 704, $18 856, $17 344, and $19 081, respectively, for the four groups. By 5 years, BCT was less expensive than mastectomy (P:<.001), with 5-year adjusted mean costs of $41 930, $45 670, $35 787, and $39 926, respectively. Costs also varied by age, with women under 65 years having higher treatment costs than older women. CONCLUSIONS: BCT may have higher short-term costs but lower long-term costs than mastectomy. PMID- 11259471 TI - Scottish adjuvant tamoxifen trial: a randomized study updated to 15 years. AB - BACKGROUND AND METHODS: The Scottish Adjuvant Tamoxifen Trial (main trial) was initiated in April 1978 to assess the effect of tamoxifen given to patients with breast cancer immediately after mastectomy (or mastectomy plus radiation therapy) (adjuvant arm) or only after the patients had had a relapse (control arm); 1323 patients were randomly assigned (667 to the adjuvant arm and 656 to the control arm). Results have been reported for the follow-up period from 2.5 through 8 years. In this article, we report updated results after a median follow-up of 15 years. If agreeable and eligible, patients who were disease free at 5 years in the adjuvant arm of the main trial were entered into a duration trial and randomly assigned either to stop taking tamoxifen (169 patients) or to continue taking it indefinitely until relapse or death (173 patients). For this update, we analyzed information on death, recurrence, survival, and other malignancies for all but 21 of the 560 living patients from the original and duration trials to determine the probabilities of total survival, systemic relapse of disease, and death from breast cancer. All statistical tests are two-sided. RESULTS: The beneficial effect of adjuvant tamoxifen given for 5 years on the probability of total survival (P =.006), systemic relapse (P =.007), and death from breast cancer (P =.002) has been maintained through 15 years. No additional benefit was observed in those randomly assigned to continue taking tamoxifen beyond 5 years. CONCLUSION: Information from this study suggests that, if adjuvant tamoxifen is given to women with operable breast cancer, it need not be for more than 5 years. PMID- 11259472 TI - Activity of a novel bcl-2/bcl-xL-bispecific antisense oligonucleotide against tumors of diverse histologic origins. AB - BACKGROUND: Increased expression of the anti-apoptotic proteins Bcl-2 and Bcl-xL is involved in the development and progression of many tumors. We recently reported that the bcl-2/bcl-xL-bispecific antisense oligonucleotide 4625 induces apoptosis in lung carcinoma cells. To further assess the therapeutic potential of oligonucleotide 4625, we investigated its effect on a series of human tumor cell lines of diverse histologic origins in vitro and in vivo. METHODS: Oligonucleotide 4625-mediated inhibition of bcl-2 and bcl-xL expression in vitro was measured in breast carcinoma cells with the use of reverse transcription polymerase chain reaction (PCR), real-time PCR, and western blotting. Cytotoxicity was assessed in several different cell lines by measurement of tumor cell growth, propidium iodide uptake, and nuclear apoptosis. The in vivo activity of oligonucleotide 4625 was determined by the inhibition of growth of established tumor xenografts in nude mice, immunohistochemical staining of Bcl-2 and Bcl-x proteins in the tumors, and western blotting of tumor lysates. Apoptosis in tumor xenografts was detected with the use of in situ TUNEL (i.e., terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-digoxigenin nick end labeling) staining. All statistical tests are two-sided. RESULTS: In breast carcinoma cells, oligonucleotide 4625 treatment reduced bcl-2 and bcl-xL messenger RNA levels in a dose-dependent manner. At 600 nM:, oligonucleotide 4625 reduced Bcl-2 and Bcl-xL protein levels to 25% (95% confidence interval [CI] = 16% to 34%) and 20% (95% CI = 14% to 26%), respectively, of the levels in untreated cells and it decreased viability in all cell lines mainly by inducing apoptosis. In vivo, oligonucleotide 4625 statistically significantly inhibited the growth of breast and colorectal carcinoma xenografts by 51% (95% CI = 28% to 74%) and 59% (95% CI = 44% to 74%), respectively, relative to those treated with control oligonucleotide 4626; it also reduced Bcl-2 and Bcl-xL protein levels and induced tumor cell apoptosis. CONCLUSION: The bcl-2/bcl-xL-bispecific antisense oligonucleotide 4625 merits further study as a novel compound for cancer therapy. PMID- 11259473 TI - Large nontransplanted hepatocellular carcinoma in woodchucks: treatment with adenovirus-mediated delivery of interleukin 12/B7.1 genes. AB - BACKGROUND: Cytokine-based gene therapy strategies efficiently stimulate immune responses against many established transplanted tumors, leading to rejection of the tumor. In this study, we investigated the therapeutic potential of cancer immunotherapy in a clinically more relevant model, woodchucks with primary hepatocellular carcinomas induced by woodchuck hepatitis virus. METHODS: Large (2 5 cm), established intrahepatic tumors were given an injection once with 1 x 10(9) plaque-forming units of AdIL-12/B7.1, an adenovirus vector carrying genes for murine interleukin 12 and B7.1, or of AdEGFP, the control virus, and regression of the tumors was then monitored. Five animals were used in total. RESULTS: In four tumor-bearing animals, the antitumor response was assessed by autopsy and histologic analysis within 1-2 weeks after treatment. In all animals treated with AdIL-12/B7.1 therapy versus AdEGFP therapy, we observed substantial tumor regression (P =.006; two-sided unpaired Student's t test) accompanied by a massive infiltration of T lymphocytes. These tumors also contained increased levels of CD4(+) and CD8(+) T cells and interferon gamma (IFN gamma). In continuously growing tumor nodules given an injection of the control virus or in nontumoral liver, no such effects (i.e., tumor regression and increased levels of CD4(+) and CD8(+) T cells and IFN gamma) were detected. In the fifth animal, monitored for long-term antitumor efficacy by magnetic resonance imaging (MRI) after intratumoral vector administration by MRI guidance, the tumor was almost completely eliminated (> or = 95%) 7 weeks after treatment. CONCLUSION: Adenovirus vector-based immunotherapy appears to be an effective treatment of large nontransplanted (orthotopic) tumors that acquire malignant characteristics in a stepwise process, reflecting the real-world scenario of hepatocellular carcinoma in humans. PMID- 11259474 TI - More about: improving the cost-effectiveness of colorectal cancer screening. PMID- 11259475 TI - Re: Cost-effectiveness studies fan colonoscopy debate. PMID- 11259476 TI - Cellular defenses against excitotoxic insults. AB - The cellular events mediating necrotic neuron death are now reasonably well understood, and involve excessive extracellular accumulation of glutamate and free cytosolic calcium. When such necrotic neurological insults occur, neurons are not passively buffeted, but instead mobilize a variety of defenses in an attempt to decrease the likelihood of neuron death, or to decrease the harm to neighboring neurons (by decreasing the likelihood of inflammation). This review considers some of these defenses, organizing them along the lines of those which decrease neuronal excitability, decrease extracellular glutamate accumulation, decrease cytosolic calcium mobilization, decrease calcium-dependent degenerative events, enhance neuronal energetics, and bias a neuron towards apoptotic, rather than necrotic, death. Although these are currently perceived as a disparate array of cellular adaptations, some experimental approaches are suggested that may help form a more unified subdiscipline of cellular defenses against neurological insults. Such an advance would help pave the way for the rational design of therapeutic interventions against necrotic insults. PMID- 11259477 TI - The role of presenilins in gamma-secretase activity: catalyst or cofactor? PMID- 11259478 TI - Presenilin and gamma-secretase: structure meets function. PMID- 11259479 TI - The multiple paradoxes of presenilins. PMID- 11259480 TI - Axonal transport of VR1 capsaicin receptor mRNA in primary afferents and its participation in inflammation-induced increase in capsaicin sensitivity. AB - Capsaicin receptors are expressed in primary sensory neurons and excited by heat and protons. We examined the inflammation-induced changes of the level of VR1 capsaicin receptor mRNA in sensory neurons and the sensitivity of primary afferents to capsaicin. Carrageenan treatment induced axonal transport of VR1 mRNA, but not that of preprotachykinin mRNA, from the dorsal root ganglia to central and peripheral axon terminals. The sensitivity of central terminals to capsaicin, which was estimated by measuring the capsaicin-evoked release of glutamate from the dorsal horn, was increased by peripheral inflammation, and such an increase was suppressed by inhibiting the RNA translation in the dorsal horn with cycloheximide and an intrathecal injection of VR1 antisense oligonucleotides. Thus, peripheral inflammation induces the axonal transport of VR1 mRNA, which may be involved in the hypersensitivity of primary afferents to capsaicin and the production of inflammatory hyperalgesia. PMID- 11259481 TI - Vitreal dihydroxyphenylacetic acid (DOPAC) as an index of retinal dopamine release. AB - Dopamine is generally accepted as a major neurotransmitter associated with light adaptive processes in the retina. However, little is known about its precise release pattern in vivo, largely due to the lack of an unambiguous method for the determination of dopamine release. We have found that vitreal levels of dihydroxyphenylacetic acid (DOPAC) reflect the rate of dopamine release in chickens. Blocking re-uptake with nomifensine significantly lowered vitreal DOPAC and retinal dopamine, confirming the retinal origin and reliance of vitreal DOPAC on intact re-uptake mechanisms. Further, inhibition of monoamine oxidase with pargyline reduced vitreal as well as retinal DOPAC levels, confirming that the DOPAC detected is generated by monoamine oxidase. Finally, we found that DOPAC diffused freely into and out of isolated vitreous bodies and we found the vitreous to be metabolically inert with respect to DOPAC, supporting the idea that vitreal levels of DOPAC are consequential to the retinal metabolism of dopamine. Exposure to light, which is known to increase retinal dopamine release, readily increased vitreal DOPAC levels. The accumulation of DOPAC in the vitreous over 6 h light fitted a mathematical model of DOPAC accumulation based on zero order influx (proportional to dopamine release rates) and diffusion driven, first order efflux. PMID- 11259482 TI - Acute and chronic effects of citalopram on postsynaptic 5-hydroxytryptamine(1A) receptor-mediated feedback: a microdialysis study in the amygdala. AB - Microdialysis was used to assess the involvement of postsynaptic 5 hydroxytryptamine(1A) (5-HT(1A)) receptors in the regulation of extracellular 5 HT in the amygdala. Local infusion of the 5-HT(1A) receptor agonist flesinoxan (0.3, 1, 3 microM) for 30 min into the amygdala maximally decreased 5-HT to 50% of basal level. Systemic administration of citalopram (10 micromol/kg) increased 5-HT to 175% of basal level. Local infusion of 1 microM of the 5-HT(1A) receptor antagonist WAY 100.635 into the amygdala augmented the effect of citalopram to more than 500% of basal 5-HT level. 5-HT(1A) receptor responsiveness after chronic citalopram treatment was determined in two ways. First, by local infusion of 1 microM flesinoxan for 30 min into the amygdala, which showed a significant 63% reduction in response (area under the concentration-time curve; AUC) for the citalopram group compared to the saline group. Second, by systemic administration of citalopram (10 micromol/kg), which increased 5-HT to 350% of basal level. The effect was larger than in untreated animals, but more important, local infusion of 1 microM WAY 100.635 into the amygdala now failed to augment the effect of citalopram. Both the flesinoxan and WAY 100.635 data suggest an involvement of postsynaptic 5-HT(1A) receptor-mediated feedback in the amygdala, which diminishes following chronic citalopram treatment. PMID- 11259483 TI - Prion protein with an E200K mutation displays properties similar to those of the cellular isoform PrP(C). AB - Creutzfeldt-Jakob disease (CJD) in Libyan Jews, linked to the E200K mutation in PRNP (E200KCJD), is the most prevalent of the inherited prion diseases. As other prion diseases, E200KCJD is characterized by the brain accumulation of PrP(Sc), a pathologic conformational isoform of a normal glycoprotein denominated PrP(C). To investigate whether the E200K mutation is enough to de novo confer PrP(Sc) properties to mutant PrP, as suggested by experiments in Chinese hamster ovary cells, we examined the biochemical behavior of E200KPrP in brains and fibroblasts from sporadic as well as homozygous and heterozygous E200KCJD patients, asymptomatic transgenic mice carrying the E200K mutation, as well as in normal and scrapie-infected mouse neuroblastoma cells expressing E200KPrP. E200KPrP was examined for protease sensitivity, solubility in detergents, releasibility by phosphoinositol phospholypase-C and localization in cholesterol enriched membrane microdomains (rafts). In all tissues except in brains of CJD patients and ScN2a cells, E200KPrP displayed properties similar to those of PrP(C). Our results indicate that the E200K mutation does not automatically convey the properties of PrP(Sc) to new PrP molecules. A conversion process occurs mainly in the prion disease affected brain, suggesting the presence of a tissue-specific or age dependent factor, in accord with the late onset nature of inherited CJD. PMID- 11259484 TI - Increased expression of NR2A subunit does not alter NMDA-evoked responses in cultured fetal trisomy 16 mouse hippocampal neurons. AB - The trisomy 16 (Ts16) mouse is an animal model for human trisomy 21 (Down's syndrome). The gene encoding the NR2A subunit of the NMDA receptor has been localized to mouse chromosome 16. In the present study, western blot analysis revealed a 2.5-fold increase of NR2A expression in cultured Ts16 embryonic hippocampal neurons. However, this increase did not affect the properties of NMDA evoked currents in response to various modulators. The sensitivity of NMDA receptors to transient applications of NMDA, spermine, and Zn(2+) was investigated in murine Ts16 and control diploid cultured embryonic hippocampal neurons. Peak and steady-state currents evoked by NMDA were potentiated by spermine at concentrations < 1 mM, and inhibited by Zn(2+) in a dose-dependent and voltage-independent manner. No marked difference was observed between Ts16 and control diploid neurons for any of these modulators with regard to IC(50) and EC(50) values or voltage dependency. Additionally, inhibition by the NR2B selective inhibitor, ifenprodil, was similar. These results demonstrate that NMDA evoked currents are not altered in cultured embryonic Ts16 neurons and suggest that Ts16 neurons contain similar functional properties of NMDA receptors as diploid control neurons despite an increased level of NR2A expression. PMID- 11259485 TI - Preferential expression of antioxidant response element mediated gene expression in astrocytes. AB - Transcriptional control of target genes by antioxidant/electrophile response elements has been well described in peripheral tissues. Genes that are regulated by this mechanism include the antioxidant enzymes NAD(P)H:quinone oxidoreductase, gamma-glutamyl cystine synthetase and glutathione-S-transferase. Antioxidant/electrophile response elements within a gene's promoter confer induction by low-molecular-weight electrophilic compounds such as tert butylhydroquinone and dimethyl fumarate. We have now examined the ability of antioxidant/electrophile response elements to elicit gene expression in neurons and astrocytes in both brain slices and primary cultures using transient transfection of promoter reporter constructs. Our results using a heat-stable human placental alkaline phosphatase reporter indicate that antioxidant/electrophile response element mediated gene expression is largely restricted to astrocyte cell populations. Placental alkaline phosphatase expression was significantly elevated in astrocytes treated with the antioxidant/electrophile response element inducer dimethyl fumarate. Mutant constructs lacking a functional antioxidant/electrophile response element abolished all placental alkaline phosphatase expression in astrocytes. We suggest that astrocytic metabolic processes that normally aid and/or protect neurons may be controlled via this inducible system. PMID- 11259486 TI - Characterization of the AT(4) receptor in a human neuroblastoma cell line (SK-N MC). AB - Angiotensin IV (Ang IV), the 3-8 fragment of angiotensin II (Ang II), binds to a distinct receptor designated the AT(4) receptor. The peptide elicits a range of vascular and central actions including facilitation of memory retention and retrieval in several learning paradigms. The aim of this study was to characterize the AT(4) receptor in a human cell line of neural origin. Receptor binding studies indicate that the human neuroblastoma cell line SK-N-MC cells express a high-affinity Ang IV binding site with a pharmacological profile similar to the AT(4) receptor: (125)I]-Ang IV and (125)I]-Nle(1)-Ang IV bind specifically to the SK-N-MC cell membranes (K(d) = 0.6 and 0.1 nM) in a saturable manner (B(max) = 1.2 pmol/mg of protein). AT(4) receptor ligands, Nle(1)-Ang IV, Ang IV and LVV-haemorphin 7 (LVV-H7), compete for the binding of [(125)I]-Ang IV or [(125)I]-Nle(1)-Ang IV to the SK-N-MC cell membranes with rank order potencies of Nle(1)-Ang IV > Ang IV > LVV-H7 with IC(50) values of 1.4, 8.7 and 59 nM ([(125)I]-Ang IV) and 1.8, 20 and 168 nM ([(125)I]-Nle(1)-Ang IV), respectively. The binding of [(125)I]-Ang IV or [(125)I]-Nle(1)-Ang IV to SK-N-MC cell membranes was not affected by the presence of GTP gamma S. Both Ang IV and LVV-H7 stimulated DNA synthesis in this cell line up to 72 and 81% above control levels, respectively. The AT(4) receptor in the SK-N-MC cells is a 180-kDa glycoprotein; under non-reducing conditions a 250-kDa band was also observed. In summary, the human neuroblastoma cell line, SK-N-MC, expresses functional AT(4) receptors that are responsive to Ang IV and LVV-H7, as indicated by an increase in DNA synthesis. This is the first human cell line of neural origin shown to express the AT(4) receptor. PMID- 11259487 TI - Coincident signalling between the Gi/Go-coupled delta-opioid receptor and the Gq coupled m3 muscarinic receptor at the level of intracellular free calcium in SH SY5Y cells. AB - In SH-SY5Y cells, activation of delta-opioid receptors with [D-Pen(2,5)] enkephalin (DPDPE; 1 microM) did not alter the intracellular free Ca(2+) concentration [Ca(2+)](i). However, when DPDPE was applied during concomitant Gq coupled m3 muscarinic receptor stimulation by carbachol or oxotremorine-M, it produced an elevation of [Ca(2+)](i). The DPDPE-evoked increase in [Ca(2+)](i) was abolished when the carbachol-sensitive intracellular Ca(2+) store was emptied. There was a marked difference between the concentration-response relationship for the elevation of [Ca(2+)](i) by carbachol (EC(50) 13 microM, Hill slope 1) and the concentration-response relationship for carbachol's permissive action in revealing the delta-opioid receptor-mediated elevation of [Ca(2+)] (EC(50) 0.7 mM; Hill slope 1.8). Sequestration of free G protein beta gamma dimers by transient transfection of cells with a beta gamma binding protein (residues 495-689 of the C terminal tail of G protein-coupled receptor kinase 2) reduced the ability of delta opioid receptor activation to elevate [Ca(2+)](i). However, DPDPE did not elevate either basal or oxotremorine-M-evoked inositol phosphate production indicating that delta-opioid receptor activation did not stimulate phospholipase C. Furthermore, delta-opioid receptor activation did not result in the reversal of muscarinic receptor desensitization, membrane hyperpolarization or stimulation of sphingosine kinase. There was no coincident signalling between the delta-opioid receptor and the lysophosphatidic acid receptor which couples to elevation of [Ca(2+)](i) in SH-SY5Y cells by a PLC independent mechanism. In SH-SY5Y cells the coincident signalling between the endogenously expressed delta-opioid and m3 muscarinic receptors appears to occur in the receptor activation-Ca(2+) release signalling pathway at a step after the activation of phospholipase C. PMID- 11259488 TI - Reactive oxygen species regulate signaling pathways induced by M1 muscarinic receptors in PC12M1 cells. AB - Activation of the m1 muscarinic receptor subtype in rat pheochromocytoma (PC12) cells stably expressing cloned m1 muscarinic acetylcholine receptors was previously shown to induce morphological changes and growth arrest. However, the signaling pathways which lead to these effects were not identified. In an attempt to characterize the intracellular signaling that might be involved in the muscarinic-induced effects, we investigated the role of reactive oxygen species in the regulation of these processes. Stimulation of the muscarinic receptor in these cells increased the intracellular concentrations of reactive oxygen species. Muscarinic activation induced intracellular signaling pathways that involve activation of Ras, extracellular signal-regulated kinase (ERK), and p38. These pathways were partially blocked when reactive oxygen species (ROS) production was prevented by the antioxidant N-acetylcysteine. Other muscarinic induced signals, such as activation of c-Jun NH(2)-terminal kinase (JNK) or an increase in the binding activity of the transcription factors nuclear factor kappa B and activator protein-1, were inhibited by the antioxidant dicoumarol. N Acetylcysteine also blocked the growth arrest and changes in cell shape induced by stimulation of the muscarinic receptor in PC12M1 cells. These findings suggest that ROS act as second messengers in muscarinic-induced cellular signaling. Moreover, generation of ROS appears to be an early and critical intermediary event, which occurs immediately after stimulation of the muscarinic receptor and affects in a variety of mechanisms the muscarinic-mediated cellular signaling. PMID- 11259489 TI - Nitrogen shuttling between neurons and glial cells during glutamate synthesis. AB - The relationship between neuronal glutamate turnover, the glutamate/glutamine cycle and de novo glutamate synthesis was examined using two different model systems, freshly dissected rat retinas ex vivo and in vivo perfused rat brains. In the ex vivo rat retina, dual kinetic control of de novo glutamate synthesis by pyruvate carboxylation and transamination of alpha-ketoglutarate to glutamate was demonstrated. Rate limitation at the transaminase step is likely imposed by the limited supply of amino acids which provide the alpha-amino group to glutamate. Measurements of synthesis of (14)C-glutamate and of (14)C-glutamine from H(14)CO(3) have shown that (14)C-amino acid synthesis increased 70% by raising medium pyruvate from 0.2 to 5 mM. The specific radioactivity of (14)C-glutamine indicated that approximately 30% of glutamine was derived from (14)CO(2) fixation. Using gabapentin, an inhibitor of the cytosolic branched-chain aminotransferase, synthesis of (14)C-glutamate and (14)C-glutamine from H(14)CO(3)(-) was inhibited by 31%. These results suggest that transamination of alpha-ketoglutarate to glutamate in Muller cells is slow, the supply of branched chain amino acids may limit flux, and that branched-chain amino acids are an obligatory source of the nitrogen required for optimal rates of de novo glutamate synthesis. Kinetic analysis suggests that the glutamate/glutamine cycle accounts for 15% of total neuronal glutamate turnover in the ex vivo retina. To examine the contribution of the glutamate/glutamine cycle to glutamate turnover in the whole brain in vivo, rats were infused intravenously with H(14)CO(3)(-). (14)C metabolites in brain extracts were measured to determine net incorporation of (14)CO(2) and specific radioactivity of glutamate and glutamine. The results indicate that 23% of glutamine in the brain in vivo is derived from (14)CO(2) fixation. Using published values for whole brain neuronal glutamate turnover, we calculated that the glutamate/glutamine cycle accounts for approximately 60% of total neuronal turnover. Finally, differences between glutamine/glutamate cycle rates in these two model systems suggest that the cycle is closely linked to neuronal activity. PMID- 11259490 TI - Synaptotagmin I is a molecular target for lead. AB - Lead poisoning can cause a wide range of symptoms with particularly severe clinical effects on the CNS. Lead can increase spontaneous neurotransmitter release but decrease evoked neurotransmitter release. These effects may be caused by an interaction of lead with specific molecular targets involved in neurotransmitter release. We demonstrate here that the normally calcium-dependent binding characteristics of the synaptic vesicle protein synaptotagmin I are altered by lead. Nanomolar concentrations of lead induce the interaction of synaptotagmin I with phospholipid liposomes. The C2A domain of synaptotagmin I is required for lead-mediated phospholipid binding. Lead protects both recombinant and endogenous rat brain synaptotagmin I from proteolytic cleavage in a manner similar to calcium. However, lead is unable to promote the interaction of either recombinant or endogenous synaptotagmin I and syntaxin. Finally, nanomolar concentrations of lead are able to directly compete with and inhibit the ability of micromolar concentrations of calcium to induce the interaction of synaptotagmin I and syntaxin. Based on these findings, we conclude that synaptotagmin I may be an important, physiologically relevant target of lead. PMID- 11259491 TI - Characterization of the 5' flanking region of the rat D(3) dopamine receptor gene. AB - The D(3) dopamine receptor has a restricted regional distribution in brain and is regulated by dopaminergic agents. Additionally, the D(3) gene is implicated in the pathogenesis of several neuropsychiatric disorders or in their response to pharmacological agents. Elucidating its transcription control mechanisms is therefore of interest in order to explain these biological features of the D(3) gene. In this study, the 5' flanking region of the rat D(3) gene was characterized by isolating the 5' end of its cDNA as well as 4.6 kb of genomic sequence. Analysis of this region revealed the presence of two new exons 196-bp and 120-bp long, separated by an 855-bp intron, located several kilobases upstream of the previously published coding exons. Thus, current evidence indicates that the rat D(3) gene is organized into eight exons. Transcription initiation site was determined by primer extension analysis and repeated rounds of 5' RACE and was found to localize at a pyrimidine-rich consensus 'initiator' sequence, similar to the rat D(2) gene. The D(3) promoter lacks TATA or CAAT boxes but unlike that of other dopamine receptor genes has only 52% GC content. Functional analysis of D(3) promoter deletion mutants fused to a reporter gene in TE671 cells, which endogenously express this gene, revealed strong transcriptional activity localized within 36 nucleotides upstream of transcription start site, and a potent silencer between bases --37 and --537. The D(3) promoter is inactive in C6 and COS7 cells. We conclude that the D(3) gene, similar to the closely related D(2) gene, is transcribed from a tissue specific promoter which is under intense negative control. PMID- 11259492 TI - Caspase 3 inhibition attenuates hydrogen peroxide-induced DNA fragmentation but not cell death in neuronal PC12 cells. AB - Exposure of neurons to H(2)O(2) results in both necrosis and apoptosis. Caspases play a pivotal role in apoptosis, but exactly how they are involved in H(2)O(2) mediated cell death is unknown. We examined H(2)O(2)-induced toxicity in neuronal PC12 cells and the effects of inducible overexpression of the H(2)O(2)-scavenging enzyme catalase on this process. H(2)O(2) caused cell death in a time- and concentration-dependent manner. Cell death induced by H(2)O(2) was found to be mediated in part through an apoptotic pathway as H(2)O(2)-treated cells exhibited cell shrinkage, nuclear condensation and marked DNA fragmentation. H(2)O(2) also triggered activation of caspase 3. Genetic up-regulation of catalase not only significantly reduced cell death but also suppressed caspase 3 activity and DNA fragmentation. While the caspase 3 inhibitor DEVD inhibited both caspase 3 activity and DNA fragmentation induced by H(2)O(2) it did not prevent cell death. Treatment with the general caspase inhibitor ZVAD, however, resulted in complete attenuation of H(2)O(2)-mediated cellular toxicity. These results suggest that DNA fragmentation induced by H(2)O(2) is attributable to caspase 3 activation and that H(2)O(2) may be critical for signaling leading to apoptosis. However, unlike inducibly increased catalase expression and general caspase inhibition both of which protect cells from cytotoxicity, caspase 3 inhibition alone did not improve cell survival suggesting that prevention of DNA fragmentation is insufficient to prevent H(2)O(2)-mediated cell death. PMID- 11259493 TI - Differentiation induces up-regulation of plasma membrane Ca(2+)-ATPase and concomitant increase in Ca(2+) efflux in human neuroblastoma cell line IMR-32. AB - Precise regulation of intracellular Ca(2+) concentration ([Ca(2+)](i)) is achieved by the coordinated function of Ca(2+) channels and Ca(2+) buffers. Neuronal differentiation induces up-regulation of Ca(2+) channels. However, little is known about the effects of differentiation on the expression of the plasma membrane Ca(2+)-ATPase (PMCA), the principal Ca(2+) extrusion mechanism in neurons. In this study, we examined the regulation of PMCA expression during differentiation of the human neuroblastoma cell line IMR-32. [Ca(2+)](i) was monitored in single cells using indo-1 microfluorimetry. When the Ca(2+)-ATPase of the endoplasmic reticulum was blocked by cyclopiazonic acid, [Ca(2+)](i) recovery after small depolarization-induced Ca(2+) loads was governed primarily by PMCAs. [Ca(2+)](i) returned to baseline by a process described by a monoexponential function in undifferentiated cells (tau = 52 +/- 4 s; n = 25). After differentiation for 12-16 days, the [Ca(2+)](i) recovery rate increased by more than threefold (tau = 17 +/- 1 s; n = 31). Western blots showed a pronounced increase in expression of three major PMCA isoforms in IMR-32 cells during differentiation, including PMCA2, PMCA3 and PMCA4. These results demonstrate up regulation of PMCAs on the functional and protein level during neuronal differentiation in vitro. Parallel amplification of Ca(2+) influx and efflux pathways may enable differentiated neurons to precisely localize Ca(2+) signals in time and space. PMID- 11259494 TI - Iron, neuromelanin and ferritin content in the substantia nigra of normal subjects at different ages: consequences for iron storage and neurodegenerative processes. AB - Information on the molecular distribution and ageing trend of brain iron in post mortem material from normal subjects is scarce. Because it is known that neuromelanin and ferritin form stable complexes with iron(III), in this study we measured the concentration of iron, ferritin and neuromelanin in substantia nigra from normal subjects, aged between 1 and 90 years, dissected post mortem. Iron levels in substantia nigra were 20 ng/mg in the first year of life, had increased to 200 ng/mg by the fourth decade and remained stable until 90 years of age. The H-ferritin concentration was also very low (29 ng/mg) during the first year of life but increased rapidly to values of approximately 200 ng/mg at 20 years of age, which then remained constant until the eighth decade of life. L-Ferritin also showed an increasing trend during life although the concentrations were approximately 50% less than that of H-ferritin at each age point. Neuromelanin was not detectable during the first year, increased to approximately 1000 ng/mg in the second decade and then increased continuously to 3500 ng/mg in the 80th year. A Mossbauer study revealed that the high-spin trivalent iron is probably arranged in a ferritin-like iron--oxyhydroxide cluster form in the substantia nigra. Based on this data and on the low H- and L-ferritin content in neurones it is concluded that neuromelanin is the major iron storage in substantia nigra neurones in normal individuals. PMID- 11259495 TI - Monoamine release by neurons of a primitive nervous system: an amperometric study. AB - We measured monoamine release from dissociated neurons of the sea pansy Renilla koellikeri, a representative of the most evolutionarily ancient animals with nervous systems, by real-time monitoring of exocytosis using the amperometric method with carbon-fiber microelectrodes. Depolarization-induced, as well as spontaneously active, neurons exhibited calcium-dependent exocytotic events at both the soma and the terminal bulb of neuritic processes. All spontaneously active neurons exhibited a bursting activity pattern in which amplitudes of exocytotic events appeared to be distributed in a quantal-like fashion. Fast Fourier transform analysis of bursting activity in 20 such neurons revealed burst harmonics with a major frequency of 8 Hz and a dominant rate of 95 Hz for individual exocytotic events within bursts. The results suggest that exocytotic transmitter release is as ancient as neurons and that endogenously bursting neurons in the sea pansy are as complex as those of higher animals. In addition, the observation that both soma and neuritic terminals of the same neuron can release transmitter suggests that local release sites in these cnidarian neurons are not critical for nerve net function. PMID- 11259496 TI - Is Bax a mitochondrial mediator in apoptotic death of dopaminergic neurons in Parkinson's disease? AB - Bax is a proapoptotic member of the Bcl-2 family of proteins. It is believed to exert its action primarily by facilitating the release of cytochrome c from the mitochondrial intermembrane space into the cytosol, leading to caspase activation and cell death. Because alterations in mitochondrial respiratory function, caspase activation and cell death with morphologic features compatible with apoptosis have been observed post mortem in the brain of patients with Parkinson's disease, we tried to clarify the potential role of Bax in this process in an immunohistochemical study on normal and Parkinson's disease post mortem brain and primary mesencephalic cell cultures treated with MPP(+). We found that Bax is expressed ubiquitously by dopaminergic (DA) neurons in post mortem brain of normal and Parkinson's disease subjects as well as in vitro. Using an antibody to Bax inserted into the outer mitochondrial membrane as an index of Bax activation, no significant differences were observed between control and Parkinson's disease subjects, regardless of the mesencephalic subregion analysed. However, in Parkinson's disease subjects, the percentage of Bax positive melanized SNpc neurons containing Lewy bodies, suggestive of DA neuronal suffering, was significantly higher than the overall percentage of Bax-positive neurons among melanized neurons. Furthermore, all melanized SNpc neurons in Parkinson's disease subjects with activated caspase-3 were also immunoreactive for Bax, suggesting that Bax anchored in the outer mitochondrial membrane of melanized SNpc neurons showing signs of neuronal suffering or apoptosis is increased compared with DA neurons that are apparently unaltered. Surprisingly, MPP(+) treatment of tyrosine hydroxylase (TH)-positive neurons in primary mesencephalic cultures did not cause redistribution of Bax, although cytochrome c was released from the mitochondria and nuclear condensation/fragmentation was induced. Taken together, these findings suggest that in the human pathology, Bax may be a cofactor in caspase activation, but our in vitro data fail to indicate a central role for Bax in apoptotic death of DA neurons in an experimental Parkinson's disease paradigm. PMID- 11259497 TI - Evidence for non-oxidative dopamine cytotoxicity: potent activation of NF-kappa B and lack of protection by anti-oxidants. AB - A stable aromatic acid decarboxylase expressing the Chinese hamster ovary cell line was developed to study the cytotoxic properties of intracellular and extracellular dopamine. The relative impermeability of cells to dopamine, but not to L-DOPA, allows the differentiation of extracellular and intracellular dopamine cytotoxicity. In contrast to extracellular dopamine, intracellular dopamine toxicity was resistant to antioxidant protection, and did not require melanin formation for its toxicity. Furthermore, we demonstrated a rapid and potent activation of the stress-inducible transcription factor NF-kappa B by intracellular dopamine, which was also largely insensitive to antioxidant inhibition. A distinctly slower and less potent NF-kappa B activation by extracellular dopamine was blocked by antioxidants and acetylsalicylic acid. Our results indicate the existence of a non-oxidative mechanism of dopamine cytotoxicity. Mitigating intracellular dopamine toxicity could be a novel strategy of slowing the progressive degeneration of dopaminergic neurons in Parkinson's disease. PMID- 11259498 TI - The human delta opioid receptor activates G(i1)alpha more efficiently than G(o1)alpha. AB - To assess the relative capacity of the human delta opioid receptor to activate closely related G proteins, fusion proteins were constructed in which the alpha subunits of either G(i1) or G(o1), containing point mutations to render them insensitive to the actions of pertussis toxin, were linked in-frame with the C terminus of the receptor. Following transient and stable expression in HEK 293 cells, both constructs bound the antagonist [(3)H]naltrindole with high affinity. D-ala(2),D-leu(5) Enkephalin effectively inhibited forskolin-stimulated adenylyl cyclase activity in intact cells in a concentration-dependent, but pertussis toxin-insensitive, manner. The high-affinity GTPase activity of both constructs was also stimulated by D-ala(2),D-leu(5) enkephalin with similar potency. However, enzyme kinetic analysis of agonist stimulation of GTPase activity demonstrated that the GTP turnover number produced in response to D-ala(2),D leu(5) enkephalin was more than three times greater for G(i1)alpha than for G(o1)alpha. As the effect of agonist in both cases was to increase V:(max) without increasing the observed K:(m) for GTP, this is consistent with receptor promoting greater guanine nucleotide exchange, and thus activation, of G(i1)alpha compared with G(o1)alpha. An equivalent fusion protein between the human mu opioid receptor-1 and G(i1)alpha produced a similar D-ala(2),D-leu(5) enkephalin induced GTP turnover number as the delta opioid receptor-G(i1)alpha fusion construct, consistent with agonist occupation of these two opioid receptor subtypes being equally efficiently coupled to activation of G(i1)alpha. PMID- 11259499 TI - Brain-derived neurotrophic factor superinduction parallels anti-epileptic- neuroprotective treatment in the pilocarpine epilepsy model. AB - Antiepileptic drugs provide neuroprotection in several animal models of brain damage, including those induced by status epilepticus (SE). The mechanisms involved in this action are unknown, but neurotrophic factors such as brain derived neurotrophic factor (BDNF) may play a role. In this study we investigated the changes in BDNF levels in rats in which SE had been induced by pilocarpine injection (400 mg/kg i.p.) and continued for several hours (unprotected group). In other animals (protected groups), SE was suppressed after 30 min by intraperitoneal injection of either diazepam (10 mg/kg) + pentobarbital (30 mg/kg) or paraldehyde (0.3 mg/kg). In diazepam + pentobarbital-treated rats the hippocampal damage caused by SE was significantly lower (p < 0.05) than in unprotected animals. In addition, 2 and 24 h after pilocarpine injection, the levels of BDNF mRNA were moderately increased in the unprotected group, but 'superinduced' in protected animals, especially in the neocortex and hippocampus. A time-dependent increase in BDNF immunoreactivity was also found by western blot analysis in rats treated with diazepam + pentobarbital. In contrast, a decrease of BDNF immunoreactivity occurred in the unprotected group. In conclusion, these results show that neuroprotection induced by anti-epileptic drugs in pilocarpine treated rats is accompanied by strong potentiation of BDNF synthesis in brain regions involved in SE. PMID- 11259500 TI - Glycine is taken up through GLYT1 and GLYT2 transporters into mouse spinal cord axon terminals and causes vesicular and carrier-mediated release of its proposed co-transmitter GABA. AB - Glycine and GABA are likely co-transmitters in the spinal cord. Their possible interactions in presynaptic terminals have, however, not been investigated. We studied the effects of glycine on GABA release using superfused mouse spinal cord synaptosomes. Glycine concentration dependently elicited [(3)H]GABA release which was insensitive to strychnine or 5,7-dichlorokynurenic acid, but was Na(+) dependent and sensitive to the glycine uptake blocker glycyldodecylamide. The glycine effect was external Ca(2+) independent, but was reduced when intraterminal Ca(2+) was chelated with 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N' tetracetic acid or depleted with thapsigargin, or when vesicular storage was impaired with bafilomycin. Glycine-induced [(3)H]GABA release was prevented, in part, by blocking GABA transport. The glycine effect was halved by sarcosine, a GLYT1 substrate/inhibitor, or by amoxapine, a GLYT2 blocker, and abolished by a mixture of the two. The sensitivity to sarcosine, used as a transporter inhibitor or substrate, persisted in synaptosomes prelabelled with [(3)H]GABA in the presence of beta-alanine, excluding major gliasome involvement. To conclude, in mice spinal cord, transporters for glycine (both GLYT1 and GLYT2) and for GABA coexist on the same axon terminals. Activation of the glycine transporters elicits GABA release, partly by internal Ca(2+)-dependent exocytosis and partly by transporter reversal. PMID- 11259502 TI - Tamoxifen inhibits nitrotyrosine formation after reversible middle cerebral artery occlusion in the rat. AB - Tamoxifen (TAM), a widely used non-steroidal anti-estrogen, has recently been shown to be neuroprotective in a rat model of reversible middle cerebral artery occlusion (rMCAo). Tamoxifen has several potential mechanisms of action including inhibition of the release of excitatory amino acids (EAA) and nitric oxide synthase (NOS) activity. The question addressed in this study was whether TAM reduces ischemia-induced production of nitrotyrosine, considered as a footprint of the product of nitric oxide and superoxide, peroxynitrite. In rat brain, 2 h rMCAo produced a time-dependent increase in nitrotyrosine content in the cerebral cortex, as measured by Western blot analysis. Compared with vehicle, TAM significantly reduced nitrotyrosine levels in the ischemic cortex at 24 h. The neuronal (n)NOS inhibitor, 7-nitroindazole also tended to reduce nitrotyrosine, but this reduction was not statistically significant. Immunostaining for nitrotyrosine was seen in cortical neurons in the MCA territory and this immunostaining was reduced by TAM. In vitro, TAM and the calmodulin inhibitor trifluoperazine inhibited, with similar EC(50) values, the activity of recombinant nNOS as well as NOS activity in brain homogenates, measured by conversion of [(3)H]arginine to [(3)H]citrulline. There was marginal inhibition of recombinant inducible (i)NOS activity up to 100 microM TAM. These data suggest that TAM is an effective inhibitor of Ca(2+)/calmodulin-dependent NOS and the derived peroxynitrite production in transient focal cerebral ischemia and this may be one mechanism for its neuroprotective effect following rMCAo. PMID- 11259501 TI - Tissue distribution and processing of proSAAS by proprotein convertases. AB - The conversion of inactive precursor proteins into bioactive neuropeptides and peptide hormones involves regulated secretory proteins such as prohormone convertases PC1 and PC2. The neuroendocrine protein 7B2 represents a specific binding protein for PC2, and the protein proSAAS, which interacts with PC1, exhibits certain structural and functional homologies with 7B2. With the intention of better understanding the physiological role of proSAAS and its derived peptides, we investigated its tissue localization using a new radioimmunoassay (RIA) to a C-terminal proSAAS-derived peptide. Immunoreactivity corresponding to this SAAS-derived peptide is mostly localized to the brain and gut. Analysis of the brain distribution of the proSAAS-derived peptides indicates that the hypothalamus and pituitary are the two richest areas, consistent with the previously described high expression of PC1 in these two areas. In order to investigate the cleavage of proSAAS by prohormone convertases, we incubated recombinant His-tagged proSAAS with recombinant mouse proPC2 or furin, separated the cleavage products using high-pressure gel permeation chromatography and analyzed the products by RIA. Our results indicate that either PC2 or furin can accomplish in vitro rapid removal and efficient internal processing of the C terminal peptide, exposing the inhibitory hexapeptide to possible further digestion by carboxypeptidases. Finally, we also studied proSAAS processing in the brains of wild-type and PC2 null mice and found that proSAAS is efficiently processed in vivo. Whereas the C-terminal peptide is mostly internally cleaved in wild-type mouse brain, it is not processed as efficiently in the brain of PC2 null mice, suggesting that PC2 is partially responsible for this cleavage in vivo. PMID- 11259503 TI - Role of Egr-1 in cAMP-dependent protein kinase regulation of the phenylethanolamine N-methyltransferase gene. AB - The molecular mechanism by which cAMP activates the rat phenylethanolamine N methyltransferase (PNMT) gene was examined by transient transfection of the wild type rat PNMT promoter-luciferase reporter gene construct pGL3RP893 into PC12 cells. Forskolin treatment (10 microM) of the transfected cells for 3--6 h maximally induced luciferase threefold. Induction by forskolin was mimicked by the cAMP analog, 8-Br-cAMP, and prevented in PC12 cells pretreated with the protein kinase A (PKA) inhibitor H-89 or co-transfected with an expression construct for PKI, a polypeptide inhibitor of PKA. Furthermore, forskolin did not activate the PNMT promoter when the 893 bp PNMT promoter-reporter gene construct was transfected into the PKA-deficient cell line, A126. Detailed examination of the forskolin responsiveness of PNMT constructs harboring > or = 60 bp and < 893 bp of PNMT promoter demonstrated that the cAMP-responsive element(s) lay between < 392 bp and > or =60 bp. Within this region of the promoter lies a functional binding element for Egr-1, a transcriptional activator of the PNMT gene. Forskolin treatment of PC12 cells also rapidly increased nuclear levels of Egr-1 and the catalytic subunit of PKA (PKA-C), with the rise in PKA-C preceding that of Egr-1. Mutation of the --165 bp Egr-1 site markedly decreased forskolin activation of the PNMT promoter. These findings demonstrate that the rat PNMT gene promoter can be activated via the cAMP-PKA signal transduction pathway, mediated by the immediate early gene transcription factor, Egr-1. PMID- 11259504 TI - Ca(2+) changes induced by different presynaptic nicotinic receptors in separate populations of individual striatal nerve terminals. AB - Presynaptic nicotinic acetylcholine receptors likely play a modulatory role in the nerve terminal. Using laser-scanning confocal microscopy, we have characterized physiological responses obtained on activation of presynaptic nicotinic receptors by measuring calcium changes in individual nerve terminals (synaptosomes) isolated from the rat corpus striatum. Nicotine (500 nM) induced Ca(2+) changes in a subset (10-25%) of synaptosomes. The Ca(2+) responses were dependent on extracellular Ca(2+) and desensitized very slowly (several minutes) on prolonged exposure to agonist. The nicotine-induced Ca(2+) responses were dose dependent and were completely blocked by dihydro-beta-erythroidine (5 microM), differentially affected by mecamylamine (10 microM) and alpha-conotoxin MII (100 nM), and not affected by alpha-bungarotoxin (500 nM). Immunocytochemical studies using well-characterized monoclonal antibodies revealed the presence of the alpha4 and alpha3/alpha5 nicotinic subunits. The nicotine-induced responses were unaffected by prior depolarization or by a mixture of Ca(2+) channel toxins including omega-conotoxin MVIIC (500 nM), omega-conotoxin GVIA (500 nM) and agatoxin TK (200 nM). Our results indicate that nicotinic receptors present on striatal nerve terminals induce Ca(2+) entry largely without involving voltage gated Ca(2+) channels, most likely by direct permeation via the receptor channel itself. In addition, at least two subpopulations of presynaptic nicotinic receptors reside on separate terminals in the striatum, suggesting distinct modulatory roles. PMID- 11259505 TI - The intracellular segment of the sodium channel beta 1 subunit is required for its efficient association with the channel alpha subunit. AB - Sodium channels consist of a pore-forming alpha subunit and auxiliary beta 1 and beta 2 subunits. The subunit beta 1 alters the kinetics and voltage-dependence of sodium channels expressed in Xenopus oocytes or mammalian cells. Functional modulation in oocytes depends on specific regions in the N-terminal extracellular domain of beta 1, but does not require the intracellular C-terminal domain. Functional modulation is qualitatively different in mammalian cells, and thus could involve different molecular mechanisms. As a first step toward testing this hypothesis, we examined modulation of brain Na(V)1.2a sodium channel alpha subunits expressed in Chinese hamster lung cells by a mutant beta1 construct with 34 amino acids deleted from the C-terminus. This deletion mutation did not modulate sodium channel function in this cell system. Co-immunoprecipitation data suggest that this loss of functional modulation was caused by inefficient association of the mutant beta 1 with alpha, despite high levels of expression of the mutant protein. In Xenopus oocytes, injection of approximately 10,000 times more mutant beta 1 RNA was required to achieve the level of functional modulation observed with injection of full-length beta 1. Together, these findings suggest that the C-terminal cytoplasmic domain of beta 1 is an important determinant of beta1 binding to the sodium channel alpha subunit in both mammalian cells and Xenopus oocytes. PMID- 11259506 TI - Steroidogenic factor-1 expression in marmoset and rat hippocampus: co localization with StAR and aromatase. AB - Steroidogenic factor-1 (SF-1), an orphan nuclear receptor, was studied with respect to the expression of steroidogenic enzymes in the hippocampus of rat and marmoset, since SF-1 is a regulator of steroid biosynthesis in the gonads. We used the steroidogenic acute regulatory protein (StAR) as a marker of the first step in the cascade of oestrogen synthesis and aromatase as a marker of the last. StAR transports cholesterol to the inner mitochondrial membrane where it is converted by the cytochrome P-450 enzyme complex. This is the rate-limiting step in steroid biosynthesis. Aromatase metabolizes testosterone to oestrogen. Using an anti-SF-1 antibody we show that SF-1 is highly expressed in neuronal cells of the pyramidal layer (CA1--CA3) and in the dentate gyrus of rat and marmoset hippocampi. Binding of the antibody was seen in more than 60% of all cells in the pyramidal layer and in the fascia dentata. In situ hybridization studies revealed the same expression pattern for StAR and aromatase. StAR and aromatase-positive cells were strictly correlated with SF-1 as shown by computer-assisted confocal microscopy in double labelling experiments (immunohistochemistry and in situ hybridization). This coexpression may imply SF-1 as a possible regulator of steroidogenesis in the hippocampus. However, a few interneurones express solely SF-1 and aromatase but are negative for StAR. Since the expression of StAR represents the first step in steroidogenesis its expression is suggestive for a de novo synthesis of steroids. A small population of interneurones must import precursors for oestrogen synthesis from other sources. Responsive cells, as evidenced by the presence of oestrogen receptor transcripts, were also found in the pyramidal layer and dentate gyrus. In conclusion, (1) SF-1 could play a regulatory role in steroidogenesis in the hippocampus of marmoset and rat and (2) with respect to the capacity of steroidogenesis two populations of hippocampal neurones coexist. PMID- 11259507 TI - Direct binding of beta-arrestins to two distinct intracellular domains of the delta opioid receptor. AB - beta-Arrestins regulate opioid receptor-mediated signal transduction and play an important role in opiate-induced analgesia and tolerance/dependence. This study was carried out to measure the direct interaction between beta-arrestins and opioid receptor. Immunoprecipitation experiments demonstrated that beta-arrestin 1 physically interacts with delta opioid receptor (DOR) co-expressed in human embryonic kidney 293 cells in an agonist-enhanced manner and truncation of the carboxyl terminus of DOR partially impairs the interaction. In vitro data from glutathione-S-transferase pull-down assay showed that the carboxyl terminus (CT) and the third intracellular loop (I3L) of DOR are both capable of and either domain is sufficient for binding to beta-arrestin 1 and 2. Surface plasmon resonance determination further revealed that binding of CT and I3L of DOR to beta-arrestin is additive, suggesting these two domains bind at distinctly different sites on beta-arrestin without considerable spatial hindrance. This study demonstrated for the first time the direct binding of beta-arrestins to the two distinct domains, the carboxyl terminus and the third intracellular loop, of DOR. PMID- 11259508 TI - Neuroprotective effects of non-steroidal anti-inflammatory drugs by direct scavenging of nitric oxide radicals. AB - Recently, it has been reported that inflammatory processes are associated with the pathophysiology of Alzheimer's disease and that treatment of non-steroidal anti-inflammatory drugs reduce the risk for Alzheimer's disease. In the present study, we examined nitric oxide radical quenching activity of non-steroidal anti inflammatory drugs and steroidal drugs using our established direct in vitro nitric oxide radical detecting system by electron spin resonance spectrometry. The non-steroidal anti-inflammatory drugs, aspirin, mefenamic acid, indomethacin and ketoprofen directly and dose-dependently scavenged generated nitric oxide radicals. In experiments of nitric oxide radical donor, NOC18-induced neuronal damage, these four non-steroidal drugs significantly prevented the NOC18-induced reduction of cell viability and apoptotic nuclear changes in neuronal cells without affecting the induction of inducible nitric oxide synthase-like immunoreactivity. However, ibuprofen, naproxen or steroidal drugs, which had less or no scavenging effects in vitro, showed almost no protective effects against NOC18-induced cell toxicity. These results suggest that the protective effects of the former four non-steroidal anti-inflammatory drugs against apoptosis might be mainly due to their direct nitric oxide radical scavenging activities in neuronal cells. These direct NO. quenching activities represent novel effects of non steroidal anti-inflammatory drugs. Our findings identified novel pharmacological mechanisms of these drugs to exert not only their anti-inflammatory, analgesic, antipyretic activities but also neuroprotective activities against neurodegeneration. PMID- 11259509 TI - Effects of glutathione depletion by 2-cyclohexen-1-one on excitatory amino acids induced enhancement of activator protein-1 DNA binding in murine hippocampus. AB - We have investigated the role of glutathione in mechanisms associated with excitatory amino acid signaling to the nuclear transcription factor activator protein-1 (AP1) in the brain using mice depleted of endogenous glutathione by prior treatment with 2-cyclohexen-1-one (CHX). In the hippocampus of animals treated with CHX 2 h before, a significant increase was seen in enhancement of AP1 DNA binding when determined 2 h after the injection of kainic acid (KA) at low doses. The sensitization to KA was not seen in animals injected with CHX 24 h before, in coincidence with the recovery of glutathione contents to the normal levels. By contrast, CHX did not significantly affect the potentiation by NMDA of AP1 binding under any experimental conditions. Prior treatment with CHX resulted in facilitation of behavioral changes induced by KA without affecting those induced by NMDA. These results suggest that endogenous glutathione may be at least in part involved in molecular mechanisms underlying transcriptional control by KA, but not by NMDA, signals of cellular functions. PMID- 11259510 TI - Peroxidative stress selectively down-regulates the neuronal stress response activated under conditions of endoplasmic reticulum dysfunction. AB - Oxidative stress has been implicated in mechanisms leading to neuronal cell injury in various pathological states of the brain. Here, we investigated the effect of peroxide exposure on the expression of genes coding for cytoplasmic and endoplasmic reticulum (ER) stress proteins. Primary neuronal cell cultures were exposed to H(2)O(2) for 6 h and mRNA levels of hsp70, grp78, grp94, gadd153 were evaluated by quantitative PCR. In addition, peroxide-induced changes in protein synthesis and cell viability were investigated. Peroxide treatment of cells triggered an almost 12-fold increase in hsp70 mRNA levels, but a significant decrease in grp78, grp94 and gadd153 mRNA levels. To establish whether peroxide exposure blocks the ER-resident stress response, cells were also exposed to thapsigargin (Tg, a specific inhibitor of ER Ca(2+)-ATPase) which has been shown to elicit the ER stress response. Tg exposure induced 7.2-fold, 3.6-fold and 8.8 fold increase in grp78, grp94 and gadd153 mRNA levels, respectively. However, after peroxide pre-exposure, the Tg-induced effect on grp78, grp94 and gadd153 mRNA levels was completely blocked. The results indicate that oxidative damage causes a selective down-regulation of the neuronal stress response activated under conditions of ER dysfunction. This down-regulation was only observed in cultures exposed to peroxide levels which induced severe suppression of protein synthesis and cell injury, implying a causative link between peroxide-induced down-regulation of ER stress response system and development of neuronal cell injury. These observations could have implications for our understanding of the mechanisms underlying neuronal cell injury in pathological states of the brain associated with oxidative damage, including Alzheimer's disease where the neuronal stress response activated under conditions of ER dysfunction has been shown to be down-regulated. Down-regulation of ER stress response may increase the sensitivity of neurones to an otherwise nonlethal form of stress. PMID- 11259511 TI - Differences in signal transduction pathways by which platelet-derived and fibroblast growth factors activate extracellular signal-regulated kinase in differentiating oligodendrocytes. AB - Treatment of cultured rat oligodendroglial progenitors with either platelet derived growth factor (PDGF) or fibroblast growth factor-2 (FGF-2) activated extracellular signal regulated kinase 2 (ERK2). Activation was transient in response to PDGF, whereas it was greater and more prolonged in response to FGF-2. ERK2 activation by PDGF was preceded by a very rapid, robust and transient tyrosine phosphorylation of the PDGF receptor. Although there was consistently more activation of ERK2 in response to FGF-2 than to PDGF, immunostaining of FGF receptors 1 (FGFR1) and 2 (FGFR2) and their tyrosine phosphorylation in progenitors was very weak, and both receptors were up-regulated during differentiation to oligodendrocytes. Tyrosine phosphorylation of the FGF receptors was maximal from 15 to 60 min of treatment and was sustained for many hours. Binding of radioiodinated FGF-2 to FGFR1 was predominant in progenitors, whereas binding to FGFR2 was predominant in oligodendrocytes. ERK2 activation by PDGF was more sensitive to inhibition of tyrosine kinases, whereas ERK2 activation by FGF-2 was relatively more sensitive to inhibitors of protein kinase C. These differences in signal transduction pathways probably contribute to the different cellular responses of oligodendroglial lineage cells to PDGF and FGF-2, respectively. PMID- 11259512 TI - Hypoxia regulates glutamate metabolism and membrane transport in rat PC12 cells. AB - We investigated the effect of hypoxia on glutamate metabolism and uptake in rat pheochromocytoma (PC12) cells. Various key enzymes relevant to glutamate production, metabolism and transport were coordinately regulated by hypoxia. PC12 cells express two glutamate-metabolizing enzymes, glutamine synthetase (GS) and glutamate decarboxylase (GAD), as well as the glutamate-producing enzyme, phosphate-activated glutaminase (PAG). Exposure to hypoxia (1% O(2)) for 6 h or longer increased expression of GS mRNA and protein and enhanced GS enzymatic activity. In contrast, hypoxia caused a significant decrease in expression of PAG mRNA and protein, and also decreased PAG activity. In addition, hypoxia led to an increase in GAD65 and GAD67 protein levels and GAD enzymatic activity. PC12 cells express three Na(+)-dependent glutamate transporters; EAAC1, GLT-1 and GLAST. Hypoxia increased EAAC1 and GLT-1 protein levels, but had no effect on GLAST. Chronic hypoxia significantly enhanced the Na(+)-dependent component of glutamate transport. Furthermore, chronic hypoxia decreased cellular content of glutamate, but increased that of glutamine. Taken together, the hypoxia-induced changes in enzymes related to glutamate metabolism and transport are consistent with a decrease in the extracellular concentration of glutamate. This may have a role in protecting PC12 cells from the cytotoxic effects of glutamate during chronic hypoxia. PMID- 11259513 TI - Selective blockade of neurokinin (NK)(1) receptors facilitates the activity of adrenergic pathways projecting to frontal cortex and dorsal hippocampus in rats. AB - The selective NK(1) receptor antagonist, GR205,171 (2.5-40.0 mg/kg, i.p.), dose dependently elevated dialysate levels of noradrenaline (NA), but not serotonin (5 HT), in the frontal cortex of freely moving rats. This action was exerted stereospecifically inasmuch as its less active isomer, GR226,206, was ineffective. In the dorsal hippocampus, GR205,171 (but not GR226,206) also significantly increased dialysate levels of NA, whereas levels of 5-HT were unaffected. Further, in anaesthetized rats, GR205,171 dose-dependently (1.0-4.0 mg/kg, i.v.) increased the firing rate of adrenergic perikarya in the locus coeruleus. In contrast, their activity was not modified by GR226,206. These findings indicate that selective blockade of NK(1) receptors enhances the activity of ascending adrenergic pathways in rats. Adrenergic mechanisms may, thus, be involved in the potential antidepressant and other functional properties of NK(1) receptor antagonists. PMID- 11259515 TI - Adolescent peer crowd affiliation: linkages with health-risk behaviors and close friendships. AB - OBJECTIVE: To examine adolescents' peer crowd affiliation and its linkages with health-risk behaviors, their friends' health-risk behaviors, the presence of close friends in the same peer crowd, and adolescents' social acceptance. METHODS: We interviewed 250 high school students and identified six categories: popular, jocks, brains, burnouts, nonconformists, or average/other. Adolescents also reported on their health-risk behaviors (including use of cigarettes, alcohol, marijuana and other drugs; risky sexual behaviors; and other risk-taking behaviors), the health-risk behaviors of their friends, the peer crowd affiliation of their closest friends, and their perceived social acceptance. RESULTS: Burnouts and nonconformists had the highest levels of health-risk behaviors across the areas assessed, the greatest proportions of close friends who engaged in similar behaviors, and relatively low social acceptance from peers. Brains and their friends engaged in extremely low levels of health-risk behaviors. Jocks and populars also showed evidence of selected areas of health risk; these teens also were more socially accepted than others. In general, adolescents' closest friends were highly nested within the same peer crowds. CONCLUSIONS: The findings further our understanding of adolescent behaviors that put them at risk for serious adult onset conditions associated with high rates of morbidity and mortality. We discuss the implications of the findings for developing health promotion efforts for adolescents. PMID- 11259516 TI - Chronic pain and its impact on quality of life in adolescents and their families. AB - OBJECTIVE: To study chronic pain not caused by somatic disease in adolescents and the effect of pain on the quality of life of the adolescents and their families. METHODS: One hundred twenty-eight youngsters (12-18 years) who had reported chronic pain kept a 3-week diary of their pain and completed a questionnaire on quality of life. Their mothers completed a questionnaire on the impact of their youngster's pain on the family. RESULTS: The most prevalent pains were limb pain, headache, abdominal, and back pain. The pain increased during the day, with the highest frequency around dinner time and the highest intensity around bedtime. Girls reported more intense and more frequent pain than boys. The higher the intensity and frequency of the pain, the lower the self-reported quality of life of the female or male adolescent, especially regarding psychological functioning (e.g. feeling less at ease), physical status (a greater incidence of other somatic complaints), and functional status (more impediments to leisure and daily activities). Chronic pain also had a negative impact on family life. The mothers reported restrictions, particularly in social life, and problems dealing with the stress of the adolescent's pain. CONCLUSIONS: Chronic pain, not caused by somatic disease, was present to a higher degree in girls; the pain increased during the day and had a negative impact on quality of life of the adolescents and the family. There is a need for future research aimed at identifying risk factors for chronic pain and pain-associated quality of life in children and adolescents. PMID- 11259517 TI - Childhood illness-related parenting stress: the pediatric inventory for parents. AB - OBJECTIVE: To develop a measure of parenting stress related to caring for a child with an illness and to evaluate its psychometric properties with a group of parents of children with cancer. METHODS: One hundred twenty-six parents (105 mothers, 21 fathers) of children (65 boys and 61 girls, M: age: 12.75 years) being followed by an oncology service were assessed using the 42-item self-report Pediatric Inventory for Parents (PIP). Internal consistency was assessed and construct validity was investigated with standardized, general self-report measures of anxiety and parenting stress. RESULTS: Internal consistency reliability for the PIP was high (Cronbach alpha range:80-.96). PIP scores were significantly correlated with a measure of state anxiety and also with parenting stress, demonstrating construct validity. After we controlled for demographic variables and general parenting stress, PIP scores showed strong independent associations with state anxiety. CONCLUSIONS: Preliminary data indicate that the PIP is a reliable and valid tool to assess parenting stress in pediatric oncology populations. As a measure of illness-related parenting stress, the PIP may be used to provide information about parent well-being that extends beyond that obtained from general measures. PMID- 11259518 TI - Daily coping practice predicts treatment effects in children with sickle cell disease. AB - OBJECTIVE: To examine the 1-month effects of a pain coping skills intervention in children with sickle cell disease (SCD). METHODS: Forty-six African American children (8-17 years old) were randomly assigned to either a coping skills condition or a standard care control condition. Children were asked to practice daily with audiotaped instructions of skills (e.g., relaxation, imagery). RESULTS: Multivariate analyses of summary measures indicated that children in the coping intervention (versus control group) reported a significantly more active approach to managing pain. Multilevel random effects models applied to daily diary data indicated that on pain days when children practiced their strategies, they had fewer health care contacts, fewer school absences, and less interference with household activities than on days when they did not practice. CONCLUSIONS: Brief training in coping skills followed by minimal therapist contact may lead to a range of benefits when children practice with their skills on a consistent basis. PMID- 11259519 TI - Maternal and child reports of behavioral compensation in response to safety equipment usage. AB - OBJECTIVE: To assess maternal and child risk compensation behaviors in response to several commonly used safety measures. METHODS: We administered a previously validated self-report measure of risk tolerance to a total of 151 mothers and their children in grades 3-7. Mothers indicated the level of risk they would permit their child to assume; children were questioned regarding the degree of physical risk they would typically assume while unsupervised by an adult. Participating families were randomly assigned to conditions in which safety equipment either was or was not present during assessments of risk tolerance. RESULTS: Mothers who viewed the stimulus materials depicting the use of safety precautions reported significantly higher levels of tolerance for risky behavior on the part of their children than did mothers who viewed identical materials without the safety precautions. No significant differences in estimated risk taking emerged between children in the two experimental conditions. CONCLUSIONS: These data may reveal a compensatory mechanism by which parents escalate their threshold for acceptable risk behavior in the presence of safety precautions for their children. Such tendencies have the potential to offset some of the protection provided by the use of safety equipment. PMID- 11259520 TI - Pioneers in pediatric psychology: between two professional worlds: personal reflections on a career in a pediatric setting. PMID- 11259521 TI - 2001 ASPET Otto Krayer Award Lecture. Molecular targets for antiviral agents. AB - There are a number of virus-specific processes within the virus replicative cycle or virus-infected cell that have proven to be attractive targets for chemotherapeutic intervention, i.e., virus adsorption and entry into the cells, reverse (RNA --> DNA) transcription, viral DNA polymerization, and cellular enzymatic reactions that are associated with viral DNA and RNA synthesis and viral mRNA maturation (i.e., methylation). A variety of chemotherapeutic agents, both nucleoside (and nucleotide) and non-nucleoside entities, have been identified that specifically interact with these viral targets, that selectively inhibit virus replication, and that are either used or considered for clinical use in the treatment of virus infections in humans. Their indications encompass virtually all major human viral pathogens, including human immunodeficiency virus (HIV), hepatitis B virus (HBV), herpes simplex virus (HSV), varicella-zoster virus (VZV), cytomegalovirus (CMV), human papilloma virus (HPV), orthomyxoviruses (influenza A and B), paramyxoviruses [e.g., respiratory syncytial virus (RSV)] and hemorrhagic fever viruses (such as Ebola virus). PMID- 11259522 TI - Unifying perspectives of the mechanisms underlying the development of tolerance and physical dependence to opioids. AB - The cellular basis of tolerance to, and dependence upon, many types of drugs, including opioids, has long defied identification. Tolerance to opioids cannot be explained solely on the basis of modification of opioid receptors or altered metabolism or disposition of the opioid. The development of tolerance following chronic exposure to opioids presents at least three different types of change in cellular responsiveness, each of which has been suggested to represent some type of adaptive modification in cellular responsiveness. These different forms of tolerance are distinguishable on the basis of their time course and whether or not the tolerance is specific for opioid receptor agonists (homologous) or extends to agonists of other systems (heterologous). The adaptive modulation of responsiveness via regulation of cellular proteins has been proposed to be the basis for both longer-term forms of tolerance. The divergent signaling pathways activated by G-protein-coupled receptors like the mu-opioid receptor provide multiple downstream targets for both short- and long-term regulation of cell function that is associated with the development of tolerance and/or dependence. Since the magnitude of receptor activation is an important determinant of the degree to which various signaling pathways are activated, the expressed characteristics of tolerance and/or dependence may be functionally related to which of these diverse pathways are stimulated to the greatest degree. Thus, the possibility that different signaling events are activated either sequentially or concurrently offers the possibility to explain the interaction between these different forms of tolerance and/or dependence. PMID- 11259523 TI - Bradykinin B(2) receptor endocytosis, recycling, and down-regulation assessed using green fluorescent protein conjugates. AB - Agonist-induced endocytosis and/or down-regulation have been evaluated using green fluorescent protein (GFP) conjugates of the rabbit bradykinin (BK) B2 receptor (B2R). COS-1 cells transiently transfected with vectors coding for either of two rabbit B2R fluorescent variants, B2R-GFP and B2R-GFP DeltaS/T (with previously identified Ser/Thr phosphorylation sites in the C-terminal tail mutated to Ala), exhibited specific and saturable binding (K(D) in the lower nM range). The acute addition of BK (10-100 nM) to HEK 293 cells stably expressing B2R-GFP in the presence of cycloheximide was rapidly followed by translocation of the surface receptors into the cells, with essentially complete recycling of the surface receptors in 1 to 3 h (confocal microscopy, cell fractionation). Adding captopril to inhibit angiotensin I-converting enzyme activity increased the half life of BK in the culture medium (enzyme immunoassay) and, accordingly, promoted B2R-GFP internalization for at least 3 h. However, agonist-induced down regulation was not observed under conditions optimal for endocytosis (microscopy, immunoblot using anti-GFP antibodies). In contrast, B2R-GFP was partially degraded following a short treatment of cells with trypsin. B2R-GFP internalized following agonist treatment was colocalized with fluorescent transferrin, supporting translocation of the receptor to recycling endosomes. B2R-GFP DeltaS/T failed to translocate into the cells following treatment with BK, but exhibited at baseline an altered subcellular distribution relative to B2R-GFP. The agonist BK promotes B(2)R receptor endocytosis followed by recycling to the cell surface, but does not promote receptor down-regulation in the heterologous system that we used here. Digestion initiated by extracellular proteases may be involved in pathological B2R down-regulation, as suggested by the simulation involving trypsin. PMID- 11259524 TI - Effect of acute ethanol on striatal dopamine neurotransmission in ambulatory rats. AB - The effect of ethanol on evoked dopamine release in the caudate putamen has been measured in behaving animals with in vivo electrochemistry. Dopamine was measured with fast-scan cyclic voltammetry in adult male rats to resolve the competing processes of dopamine uptake and release. Ethanol dose dependently decreased dopamine efflux compared with saline-treated animals: to 89% of controls with 0.5 g/kg, 70% with 1 g/kg, 34% with 2.5 g/kg, and 18% with 5 g/kg. This decrease was not due to a change in uptake, as measured by the rate of dopamine disappearance after stimulation, and therefore can be attributed to decreased dopamine release. Additionally, it was not mediated by a decrease in biosynthesis, as measured by L DOPA accumulation after NSD 1015 administration. The selective dopamine uptake inhibitor GBR 12909 compensated for the effects of high doses of ethanol on dopamine release. Moreover, GBR 12909 induced faster restoration of the righting reflex in rats sedated with 2.5 g/kg, but not 5 g/kg, ethanol. In brain slices containing the caudate putamen, ethanol suppressed dopamine release only at the highest dose tested (200 mM). The difference in responses between the slice and the intact animal indicates that ethanol exerts its effects in the cell body regions of dopamine neurons as well as in terminals. These neurochemical results, combined with published accounts of microdialysis measures of extracellular dopamine and electrophysiological recordings of dopamine neurons, demonstrate that ethanol has a profound effect on dopamine neurons whose net result is a suppression of dopamine neurotransmission at high doses. PMID- 11259525 TI - Influence of Short-Term Octreotide Administration on Chronic Tissue Injury, Transforming Growth Factor beta (TGF-beta) Overexpression, and Collagen Accumulation in Irradiated Rat Intestine. AB - The somatostatin analog octreotide was recently found to ameliorate radiation induced tissue injury in rat intestine. The present study addressed whether octreotide reduces chronic intestinal radiation fibrosis, whether enteroprotection is conferred by direct or indirect mechanisms, and whether the effects are dose-dependent. Using a rat model designed for fractionated irradiation, a segment of small intestine was sham-irradiated or exposed to 67.2 Gy X-radiation in 16 daily fractions. Octreotide (0, 2, or 10 microg/kg/h) was administered subcutaneously by osmotic minipumps for 4 weeks, from 2 days before to 10 days after irradiation. Tissue injury was assessed at 2 weeks (early phase) and 26 weeks (chronic phase) by quantitative histopathology and morphometry. Epithelial and smooth muscle cell proliferation was assessed by proliferating cell nuclear antigen staining; connective tissue mast cell hyperplasia by metachromatic staining; and TGF-beta1 and collagen protein and mRNA by quantitative immunohistochemistry, in situ hybridization, and/or real-time fluorogenic probe reverse transcription-polymerase chain reaction. Octreotide conferred dose-dependent protection against early (p = 0.0003) and chronic (p < 0.0001) tissue injury. Octreotide abrogated radiation-induced chronic increases in extracellular matrix-associated TGF-beta (p < 0.0001), collagen I (p = 0.0001), and collagen III (p = 0.0002) immunoreactivity. Octreotide did not affect radiation-induced changes in steady-state TGF-beta1 mRNA levels, mast cell hyperplasia, or smooth muscle cell proliferation. Octreotide reduced crypt epithelial cell proliferation (p = 0.01), but did not otherwise affect unirradiated intestine. Octreotide confers dose-dependent protection against delayed small bowel radiation toxicity and ameliorates radiation fibrosis predominantly by reducing acute mucosal injury. These data strengthen the rationale for using somatostatin analogs as enteroprotective agents in clinical radiation therapy. PMID- 11259526 TI - Cloning and functional characterization of two murine uridine nucleotide receptors reveal a potential target for correcting ion transport deficiency in cystic fibrosis gallbladder. AB - Extracellular nucleotides regulate transepithelial ion secretion via multiple receptors. The P2Y(2) receptor is the predominant transducer of chloride transport responses to nucleotides in the airways, but the P2 receptors that control ion transport in gastrointestinal epithelia have not been identified. UTP and UDP promote chloride secretion in mouse jejuna and gallbladder epithelia, respectively, and these responses were unaffected by P2Y(2) receptor gene disruption. Pharmacological data suggested the involvement of P2Y(4) and P2Y(6) receptors in gastrointestinal responses. To identify the P2Y receptors responsible for the gastrointestinal actions of UTP and UDP, we have cloned the murine P2Y(4) and P2Y(6) receptors and have stably expressed each in a null cell line to examine the nucleotide-promoted inositol phosphate formation and intracellular Ca(2+) mobilization. The (m)P2Y(4) receptor was potently, but not selectively, activated by UTP (UTP > or = ATP >ITP > GTP > CTP), and it was not activated by UDP or ADP. The (m)P2Y(6) receptor was highly selective for UDP (UDP >> ADP = GDP). The nucleotide selectivities observed with the recombinant (m)P2Y(4) and (m)P2Y(6) receptors resemble those for nucleotide-promoted chloride transport in murine P2Y(2)(-/-) jejuna and gallbladder epithelial cells, respectively. Ion transport responses to nucleotide additions were examined in freshly excised tissues from cystic fibrosis transmembrane regulator-deficient mice. Although the effect of UTP on jejunal short-circuit current (I(sc)) was impaired in the CF mouse, UDP-promoted I(sc) changes were not affected in CF gallbladder epithelium, suggesting that the P2Y(6) receptor is a target for treatment of CF gallbladder disease. PMID- 11259527 TI - Stimulatory effect of clofibrate on the action of estradiol in the mammary gland but not in the uterus of rats. AB - Treatment of ovariectomized rats with dietary clofibrate caused a manyfold increase in the liver microsomal esterification of estradiol with fatty acids. The stimulatory effect of clofibrate administration on fatty acid esterification of estradiol by liver microsomes was paralleled in vivo by enhanced estradiol induced increases in the formation of lobules in the mammary gland and by increased incorporation of bromodeoxyuridine into these lobules. In contrast to the stimulatory effect of clofibrate administration on the action of estradiol in the mammary gland, clofibrate administration had no effect on the uterotropic action of estradiol. These results indicate that clofibrate administration has a selective stimulatory effect on the hormonal action of estradiol in the mammary gland but not in the uterus. PMID- 11259528 TI - Matrix metalloproteinase inhibitors cause cell cycle arrest and apoptosis in glomerular mesangial cells. AB - Inflammation is characterized by an excess of cell proliferation often leading to fibrosis and sclerosis with subsequent loss of organ function. We hypothesized that these features may be ameliorated by induction of cell cycle arrest and apoptosis as result of therapy with matrix metalloproteinase (MMP) inhibitors. In our study, mesangial cells and experimental mesangial proliferative glomerulonephritis provided the model of inflammation. First, we investigated the effect of the MMP inhibitor BB-1101 in anti-Thy1.1 nephritis. The numbers of apoptotic glomerular cells in nephritic rats increased about 4 and 6 times as a result of BB-1101 therapy, observed 11 and 14 days after induction of disease, respectively. Subsequently, rat mesangial cells were exposed to an MMP inhibitor in vitro. Fluorescence-activated cell sorter analyses of cells exposed to RO111 3456 demonstrated a dose-dependent cell cycle arrest in the G(0)/G(1) phase associated with increased expression of statin. The cell cycle arrest was followed by apoptosis as investigated by terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP) biotin nick-end labeling (TUNEL) and acridine orange/ethidium bromide stainings, as well as by annexin V binding. The induction of p53, p21, and bax, but not the Fas/FasL pathway appeared to play an important pathogenetic role. In summary, MMP inhibitors induce cell cycle arrest followed by apoptosis in mesangial cells. These features help to explain the anti-inflammatory effects of these compounds, such as reduction of mesangial cell proliferation and attenuation of extracellular matrix accumulation. In conclusion, induction of cell cycle arrest with subsequent apoptosis may offer new perspectives in the therapy of inflammation even beyond kidney diseases. PMID- 11259529 TI - Characterization of opioid receptors modulating neurogenic contractions of circular muscle from porcine ileum and evidence that delta- and kappa-opioid receptors are coexpressed in myenteric neurons. AB - Opioid receptors (ORs) in the myenteric plexus mediate the antimotility actions of opioids in the small intestine. In this study, ORs modulating neurogenic circular muscle contractions in the porcine ileum were characterized by pharmacological and immunohistochemical approaches. Circular muscle-myenteric plexus strips manifested tetrodotoxin- and atropine-sensitive contractions during (ON) and after (OFF) electrical field stimulation. The kappa-OR agonists U 50,488H and U-69,593 inhibited ON contractions (pIC(50) = 7.61 and 8.22, respectively). U-69,593 action was inhibited by the kappa-OR antagonist norbinaltorphimine with an antagonist equilibrium constant (K(e)) of 4.2 nM. Selective delta-OR agonists [D-Ala(2)]-deltorphin II, DSLET, DADLE, SNC80, and DPDPE inhibited OFF contractions (pIC(50) = 9.17, 8.63, 8.50, 8.26, and 7.47, respectively). The selective delta-OR antagonist naltriben reduced the inhibitory actions of SNC80 and DSLET with respective K(e) values of 2.3 and 3.0 nM. In addition, norbinaltorphimine inhibited the actions of these agonists with respective K(e) values of 0.7 and 4.2 nM. The mu-OR agonists DAMGO, loperamide, or morphine exhibited relatively low activities in inhibiting ON and OFF contractions. Using primary antisera directed toward cloned opioid receptors, delta-OR immunoreactivity was observed to be localized alone or in combination with kappa-OR immunoreactivity in myenteric neurons; mu-OR immunoreactivity was absent. The results suggest that myenteric delta- and kappa-opioid receptors mediate the antitransit effects of opioids in the porcine small intestine. These receptors may be functionally coupled in a subpopulation of myenteric neurons. PMID- 11259530 TI - Prostaglandin A(1) protects striatal neurons against excitotoxic injury in rat striatum. AB - Prostaglandin A(1) (PGA1) reportedly inhibits NF-kappaB activation and induces expression of heat shock proteins. Since both these effects could be neuroprotective, the therapeutic potential of PGA1 in neurodegenerative disorders, where excitotoxicity may contribute to pathogenesis, was evaluated in rat striatal neurons exposed to the N-methyl-D-aspartate (NMDA) receptor agonist quinolinic acid (QA). Intrastriatal administration of PGA1 (5-80 nmol) attenuated QA (60 nmol)-induced internucleosomal DNA fragmentation. The inhibitory effects of a single dose of PGA1 (80 nmol) on QA (60 nmol)-induced DNA fragmentation were observed 12 to 48 h after treatment. PGA1 (80 nmol) also attenuated QA-induced DNA fragmentation when administered up to 4 h after QA exposure. PGA1 significantly decreased the loss of D1 dopamine receptors and GAD(67) mRNA in QA injected striatum as measured by quantitative receptor autoradiography and in situ hybridization histochemistry, suggesting that it reduced the neuronal loss induced by QA. Protection of striatal neurons against QA-induced death by PGA1 was further indicated by Nissl staining 10 days after QA administration. PGA1 (5 80 nmol) significantly inhibited QA-induced NF-kappaB activation by blocking inhibitory kappaB-alpha degradation but had no effect on activator protein-1 binding activity. PGA1 (80 nmol) treatment substantially increased 70- and 72-kDa heat shock protein levels in striatum. These results indicate that PGA1 blunts NMDA receptor-mediated neuronal apoptosis by a mechanism possibly involving the up-regulation of neuroprotective heat shock proteins and inhibition of NF-kappaB activation. In view of its potent neuroprotective activity, PGA1 could prove useful in the treatment of certain neurodegenerative disorders related to excitotoxicity. PMID- 11259531 TI - Characterization of the serotonin receptor mediating contraction in the mouse thoracic aorta and signal pathway coupling. AB - The purpose of this study was to characterize pharmacologically the 5-HT receptor(s) mediating contraction in the mouse aorta and the pathways these receptors are coupled with to mediate contraction. We hypothesized that a 5-HT2A receptor, as in the rat, mediates contraction by activating L-type calcium channels, phospholipase C (PLC), and tyrosine kinase(s). Endothelium-denuded aortic strips were placed in a tissue bath for measurement of isometric contractile force. 5-HT, the 5-HT2A receptor agonist alpha-methyl-5-HT, and partial 5-HT2A receptor agonist (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (+/--DOI) caused the most potent and efficacious contraction. The 5 HT(1E/1F) receptor agonist BRL 54443 also induced contraction (-log EC(50) = 6.52); however, the 5-HT2A receptor antagonist ketanserin antagonized this contraction. Our hypothesis was further supported by the finding that antagonists with affinity for the 5-HT2A receptor, ketanserin, 1-(1-naphthyl)piperazine, spiperone, and LY53857, reduced 5-HT-induced contraction. A correlation of 0.927 was found between literature-derived compound binding affinities for the agonists and antagonists at the 5-HT2A receptor of the rat and the data generated in our experiments (-log EC(50) and pK(B) values). The L-type calcium channel blockers nifedipine and nitrendipine, PLC inhibitor 2-nitro-4-carboxyphenyl N,N diphenylcarbamate, and tyrosine kinase inhibitors genistein and PD 098,059 all shifted and/or reduced maximum contraction to 5-HT. We conclude that contraction to 5-HT in the mouse aorta is mediated primarily by a 5-HT2A receptor and is coupled to L-type calcium channels, PLC, and tyrosine kinases. PMID- 11259532 TI - Hydrogen peroxide modifies the kinetics of HERG channel expressed in a mammalian cell line. AB - Reactive oxygen species such as H2O2 were shown to influence both electrical and contractile properties of the heart. H2O2 modulates action potential duration and leads to reperfusion-induced arrhythmias. As these effects could involve the modulation of repolarizing currents, we assessed effects of H2O2 on HERG (which encodes the cardiac potassium channel I(Kr)) expressed in Chinese hamster ovary cells. HERG currents were recorded using the whole-cell patch-clamp technique. HERG activation and deactivation were accelerated when cells were superfused with 30 microM, 100 microM, or 1 mM H2O2. For example, at 1 mM H2O2, tau(act) was decreased from 862 +/- 178 to 633 +/- 151 ms (P < 0.05; n = 6), and fast tau(deact) was reduced from 286 +/- 47 to 151 +/- 18 ms (P < 0.05; n = 6). A negative shift of V1/2 was also observed (from -1.9 to -13.7 mV with 30 microM H2O2; P < 0.05), reflecting the acceleration of the activating current. Effects of H2O2 superfusion were prevented by intracellular application of catalase but superoxide dismutase prevented only H2O2-induced acceleration of activation. This indicates that H2O2 diffuses intracellularly before acting on HERG and that its effects on activation but not deactivation are mediated by the superoxide anion. Moreover, tau(act) decrease preceded fast tau(deact) decrease by about 4 min, suggesting that these effects were not produced by the same intracellular pathway or at the same site on HERG protein. Acceleration of HERG activation kinetics leads to an increase of outward current during the plateau phase of the action potential. This could suggest a reason for H2O2-induced shortening of the action potential. The faster HERG deactivation could be involved in reperfusion-induced arrhythmias by reducing K+ conductance in the early diastole, thus increasing the risks of premature beats. PMID- 11259533 TI - Comparison of the modulatory effects of human and rat liver microsomal metabolism on the estrogenicity of bisphenol A: implications for extrapolation to humans. AB - Bisphenol A [BPA, 2,2-bis(4-hydroxyphenyl)propane], a xenoestrogen, is a monomer for the synthesis of polycarbonate plastics, epoxy resins, and composites. Metabolism of BPA to the monoglucuronide will determine the extent of its estrogenicity in vivo. Investigation of the metabolism of BPA (500 microM) by isolated female rat hepatocytes confirmed the formation of BPA glucuronide as the major metabolite. There was a significant difference (p < 0.05) between the V(max) (mean +/- S.E.M., n = 4) of glucuronidation by pooled male or female human (four livers in each case) and immature female rat liver microsomes (5.9 +/- 0.4, 5.2 +/- 0.3, and 31.6 +/- 8.1 nmol/min/mg of protein, respectively). Estrogenic activity of BPA, assessed in a coupled microsomal metabolism-yeast estrogenicity assay, was decreased 3- and 7-fold following glucuronidation by human female and immature female rat liver microsomes, respectively. Incubations of BPA with pooled human or rat liver microsomes, in the presence of NADPH, resulted in the formation of 5-hydroxybisphenol A [2-(4,5-dihydroxyphenyl)-2-(4 hydroxyphenyl)propane], which was 10-fold less potent than BPA in the yeast estrogenicity assay. However, there was insufficient turnover to achieve a significant effect on the estrogenic activity of BPA. Because human liver microsomes did not glucuronidate BPA as extensively as the rat liver microsomes, estrogen target tissues in humans may be subject to greater exposure to BPA than the tissues of the immature female rats used for assessing estrogenicity of xenobiotics. PMID- 11259534 TI - Discovery of a novel antitumor benzolactone enamide class that selectively inhibits mammalian vacuolar-type (H+)-atpases. AB - A series of naturally occurring compounds reported recently by multiple laboratories defines a new small-molecule class sharing a unique benzolactone enamide core structure and diverse biological actions, including inhibition of growth of tumor cells and oncogene-transformed cell lines. Here we show that representative members of this class, including salicylihalamide A, lobatamides A F, and oximidines I and II inhibit mammalian vacuolar-type (H+)-ATPases (V ATPases) with unprecedented selectivity. Data derived from the NCI 60-cell antitumor screen critically predicted the V-ATPase molecular target, while specific biochemical assays provided confirmation and further illumination. The compounds potently blocked representative V-ATPases from human kidney, liver, and osteoclastic giant-cell tumor of bone but were essentially inactive against V ATPases of Neurospora crassa and Saccharomyces cerevisiae and other membrane ATPases. Essential regulation of pH in cytoplasmic, intraorganellar, and local extracellular spaces is provided by V-ATPases, which are ubiquitously distributed among eukaryotic cells and tissues. The most potent and selective V-ATPase inhibitors heretofore known were the bafilomycins and concanamycins, which do not discriminate between mammalian and nonmammalian V-ATPases. Numerous physiological processes are mediated by V-ATPases, and aberrant V-ATPase functions are implicated in many different human diseases. Previous efforts to develop therapeutic pharmacological modulators of V-ATPases have been frustrated by a lack of synthetically tractable and biologically selective leads. Therefore, availability of the unique benzolactone enamide inhibitor class may enable further elucidation of functional and architectural features of mammalian versus nonmammalian V-ATPase isoforms and provide new opportunities for targeting V ATPase-mediated processes implicated in diverse pathophysiological phenomena, including cancer. PMID- 11259535 TI - Changes in opioid and cannabinoid receptor protein following short-term combination treatment with delta(9)-tetrahydrocannabinol and morphine. AB - Recent studies in our laboratory have shown that in mice, low doses of morphine in combination with Delta(9)-tetrahydrocannabinol (Delta(9)-THC) have a similar antinociceptive effect to high doses of morphine alone. After short-term administration of this combination, there is no behavioral tolerance to the opioid. Previous binding studies and Western analyses following chronic morphine exposure in rodent models indicate significant mu-receptor down-regulation, as well as decreased levels of receptor protein, in both brain and spinal cord regions. We hypothesized that combination-treated animals would show no receptor protein down-regulation. The levels of opioid (mu, delta, kappa) and cannabinoid (CB1) receptor protein were evaluated in mouse models of short-term exposure to Delta(9)-THC, morphine, or both drugs in combination. Western blot analysis revealed that all three types of opioid receptor protein are significantly decreased in morphine-tolerant mouse midbrain. This down-regulation was not seen in combination-treated animals. In the spinal cord, there was an up-regulation of mu-, delta-, and kappa-opioid receptor protein in combination-treated mice when compared with morphine-tolerant mice. There were no apparent changes in levels of CB1 receptor protein in midbrain regions, and there was an up-regulation of CB1 protein in the spinal cord. The data presented here indicate that there is a correlation between morphine tolerance and receptor protein regulation. A combination of Delta(9)-THC and morphine retains high antinociceptive effect without causing changes in receptor protein that may contribute to tolerance. PMID- 11259536 TI - Intravasal peroxynitrite generation causes dysfunction in the isolated perfused rat lung via endothelin. AB - In septic shock excessive nitric oxide and superoxide are produced, thus generating peroxynitrite. This study investigates whether and how intravasal peroxynitrite causes lung dysfunction. To generate peroxynitrite, isolated and ventilated rat lungs were perfused blood-free in a pressure-constant, recirculating mode with hypoxanthine/xanthine oxidase plus sodium nitroprusside. Airway and vascular resistance, and release of thromboxane A2, prostacyclin, and endothelin-1 were assessed over 200 min. Peroxynitrite generation, as demonstrated by oxidation of the marker 2',7'-dichlorodihydrofluorescein diacetate, caused broncho- and vasoconstriction starting after 100 min. Both reactants alone, i.e., NO. or O2, had no effect. The thromboxane A2/prostaglandin H2 receptor antagonist BM13.177 did not affect peroxynitrite-induced broncho- and vasoconstriction. Combined endothelin(A/B) (ET(A/B)) receptor antagonism (BQ123 plus BQ788) prevented broncho- and vasoconstriction more effectively than the ET(A) receptor antagonist BQ123 alone. In tissue from lungs exposed to peroxynitrite, significantly increased amounts of endothelin-1 were detected. This study identifies endothelin-1 rather than prostanoids as a distal mediator induced by the reaction product of superoxide and nitric oxide, i.e., peroxynitrite. It is concluded that 1) endothelin-1 is a causal mediator of peroxynitrite-induced acute rat lung injury, and 2) peroxynitrite-induced broncho and vasoconstriction are mediated by both ET(A) and ET(B) receptors. PMID- 11259537 TI - Dynamic dopamine-antagonist interactions at recombinant human dopamine D(2short) receptor: dopamine-bound versus antagonist-bound receptor states. AB - Antipsychotic drugs comprise a wide range of structurally diverse compounds and are considered to be antagonists at dopamine D2 receptors. High-resolution kinetic analyses of their antagonist properties was performed by monitoring dynamic dopamine (DA)-antagonist interactions at the recombinant human dopamine D(2short) receptor. Time-dependent Ca2+ responses were measured following activation of a chimeric G(alphaq/o) protein in Chinese hamster ovary-K1 cells. DA (10 microM) induced a rapid, high-magnitude Ca2+ response (T(max) = 13.2 +/- 0.7 s) followed by a low-magnitude phase, which continued throughout the recorded time period (15 min). Of a large series of putative DA antagonists, (+)-UH 232 and bromerguride demonstrated positive, DA-like intrinsic activity at the presumably unoccupied, DA-free receptor; the other antagonists being silent. Antagonists differed in terms of their abilities to prevent the high-magnitude Ca2+ phase in the antagonist-bound receptor state, and to reverse the low magnitude Ca2+ phase in the DA-bound state. The benzamide derivatives tropapride and nemonapride fully antagonized both the high- and low-magnitude Ca2+ response. Haloperidol, risperidone, and S 14066 also antagonized both responses but with a maximal effect of only 62 to 79%. Although preventing the high-magnitude response (85-95%), the further putative antagonists (+)-butaclamol (6%), bromerguride (27%), and domperidone (41%) reversed the low-magnitude response only weakly and partially. These Ca2+ data indicate that putative DA antagonists act differently, in particular, at the DA-bound D(2short) receptor. PMID- 11259538 TI - Probenecid-associated alterations in valproate glucuronide hepatobiliary disposition: mechanistic assessment using mathematical modeling. AB - The complexity of processes associated with the hepatobiliary disposition of xenobiotics may require a multiexperimental approach, including pharmacokinetic modeling, to assess mechanisms of drug interactions. The objective of this study was to examine the disposition of valproate glucuronide (VG) in the rat isolated perfused liver (IPL), and to determine the mechanisms of interaction with probenecid (PRB). Livers were isolated and perfused with standard techniques, and valproate (VPA) (20 mg) was administered in the absence and presence of PRB (approximately 75 microg/ml). Concentrations of VPA and VG in perfusate and bile were determined at timed intervals. In the absence of PRB, total recovery of VPA and VG in perfusate and bile was approximately 80%; PRB significantly increased this recovery to approximately 100%, suggesting a decrease in oxidative VPA metabolism. Similarly, pharmacokinetic modeling of the IPL data indicated that PRB competitively inhibited formation of oxidative VPA metabolites. PRB also significantly inhibited formation, biliary excretion, and sinusoidal egress of VG. These observations suggest a competitive interaction between PRB and VG for transport across the canalicular and sinusoidal membranes. Despite PRB-associated impairment of VG formation, mathematical modeling of the data revealed that hepatocyte VG concentrations were increased by PRB, presumably due to simultaneous inhibition of VG biliary excretion and sinusoidal egress by PRB. These results demonstrate the utility of pharmacokinetic modeling in elucidating the mechanisms of alteration in the hepatobiliary disposition of xenobiotics. PMID- 11259539 TI - Dexfenfluramine-associated changes in 5-hydroxytryptamine transporter expression and development of hypoxic pulmonary hypertension in rats. AB - The appetite suppressant dexfenfluramine, which inhibits neuronal 5-HT uptake and elevates plasma 5-HT levels, has been associated with an increase in the relative risk of developing primary pulmonary hypertension. 5-HT is a mitogen for pulmonary artery smooth muscle cells (PA-SMCs), an effect that depends upon activity of the 5-HT transporter (5-HTT). To investigate the relationship between dexfenfluramine and pulmonary hypertension, we examined 1) the effect of dexfenfluramine on 5-HT uptake by PA-SMCs and the mitogenic response of these cells to 5-HT, and 2) 5-HTT mRNA in lung tissue from normoxic and chronically hypoxic rats during and at discontinuation of a 4-week dexfenfluramine treatment (2 mg/kg/day). In cultured PA-SMCs, dexfenfluramine (10(-6) M) markedly reduced [3H]5-HT uptake and [3H]thymidine incorporation in response to 5-HT (10(-6) M). In lungs from rats exposed to 4-week hypoxia (10% O(2)), 5-HTT mRNA levels were higher than in normoxic rats (233.5 +/- 22.5 versus 121.8 +/- 4.8 amol/mg of RNA, P < 0.05), but were not affected by concomitant treatment with dexfenfluramine. One week after discontinuation of dexfenfluramine, 5-HTT mRNA levels increased substantially, this effect being additive with that of hypoxia (364.0 +/- 13.1 in hypoxic versus 164.2 +/- 10 amol/mg of RNA in normoxic rats). When exposure to 2 weeks of hypoxia followed discontinuation of a 4-week treatment, right ventricular hypertrophy was more severe and muscularization of distal pulmonary arteries more marked (P < 0.01) than in rats pretreated with the vehicle. These data show that, in rats, the increased 5-HTT expression that follows dexfenfluramine discontinuation promotes the development of hypoxic pulmonary hypertension. PMID- 11259540 TI - Renal and hepatic toxicity of trichloroethylene and its glutathione-derived metabolites in rats and mice: sex-, species-, and tissue-dependent differences. AB - Acute cytotoxicity (lactate dehydrogenase release) of trichloroethylene (TRI), S (1,2-dichlorovinyl)glutathione (DCVG), and S-(1,2-dichlorovinyl)-L-cysteine (DCVC) in freshly isolated renal cortical cells and hepatocytes from male and female rats was evaluated to test the hypothesis that the assay provides a valid indicator of sex- and tissue-dependent differences in sensitivity to TRI and its metabolites. We then determined mitochondrial toxicity (inhibition of state-3 and/or stimulation of state-4 respiration) in renal cortical and hepatic mitochondria from male and female rats and mice to assess sex-, tissue-, and species-dependent susceptibility. TRI was moderately cytotoxic in renal cells from male rats but was nontoxic in renal cells from female rats or hepatocytes from male or female rats. Acute cytotoxicity of both DCVG and DCVC was greater in renal cells from male rats than in renal cells from female rats. Although DCVC does not target the liver in vivo, it was a very potent hepatotoxicant in vitro. Mitochondrial toxicity in kidney and liver showed similar patterns, with mitochondria from male rats being more sensitive than mitochondria from female rats; order of potency was DCVC > DCVG >> TRI. State-3 respiration in mitochondria from mice was also inhibited, but the patterns and relative sensitivities differed from those in mitochondria from rats. Renal and hepatic mitochondria from mice were less sensitive than corresponding mitochondria from rats and renal mitochondria from female mice were significantly more sensitive than renal mitochondria from male mice. Thus, many of the species-, sex-, and tissue-dependent differences in toxicity observed in vivo are also observed in vitro. PMID- 11259541 TI - Antiasthmatic effect of YM976, a novel PDE4 inhibitor, in guinea pigs. AB - YM976 is a novel and specific phosphodiesterase 4 inhibitor. In our previous report, we indicated that YM976 has less emetogenicity, a major adverse effect of PDE4 inhibitors, than rolipram. In the present study, we examined the antiasthmatic effects of YM976 in guinea pigs. YM976 orally administered exhibited inhibition of antigen-induced bronchoconstriction, airway plasma leakage, airway eosinophil infiltration, and airway hyperreactivity (AHR), with ED(50) values of 7.3, 5.7, 1.0, and 0.52 mg/kg, respectively. Rolipram also dose dependently suppressed these responses. Prednisolone suppressed eosinophil infiltration and AHR, whereas it failed to inhibit bronchoconstriction and plasma leakage. Theophylline moderately suppressed bronchoconstriction and edema, but neither eosinophil infiltration nor AHR. YM976 suppressed the peroxidase activity in the bronchoalveolar lavage fluid, and elevated the intracellular peroxidase activity and cAMP contents of infiltrated cells, suggesting that YM976 inhibited not only the infiltration but also the activation of leukocytes. In vitro studies revealed that YM976 potently suppressed eosinophil activation (EC(30) = 83 nM), and exerted a little relaxation on LTD(4)-precontracted tracheal smooth muscle (EC(50) = 370 nM). Rolipram exhibited a potent tracheal relaxation activity (EC(50) = 50 nM). In vivo studies indicated that the inhibitory effect of YM976 on LTD(4)-induced bronchospasm was marginal even at 30 mg/kg p.o., although rolipram significantly inhibited the bronchospasm at the same dose. These results suggested that YM976, unlike rolipram, showed the inhibition of antigen-induced airway responses due to anti-inflammatory effects, but not to direct tracheal relaxation. In conclusion, YM976 may have potential therapeutic value in the treatment of asthma through its anti-inflammatory activities. PMID- 11259542 TI - Regulation of hepatic cytochrome P450 2C11 via cAMP: implications for down regulation in diabetes, fasting, and inflammation. AB - The effect of glucagon and its second messenger cAMP on cytochrome P450 2C11 (CYP2C11) expression was investigated in primary hepatocytes cultured on Matrigel. Glucagon, epinephrine, forskolin, and the cAMP derivatives dibutyryl cAMP, (S(p))-adenosine 3',5' cyclic monophosphothioate (S(p)-cAMPS), and 8-(4 chlorophenylthio)-cAMP, but not dideoxyforskolin, all down-regulated CYP2C11 mRNA expression to approximately 20% of control levels in a concentration-dependent manner. Using the transcriptional inhibitor 5,6-dichloro-1-beta-D ribofuranosylbenzimidazole, CYP2C11 mRNA was found to have a half-life of 9.8 h. The kinetics of suppression of CYP2C11 mRNA by glucagon and forskolin was similar to that obtained with the transcriptional inhibitor, suggesting that glucagon and forskolin act at the transcriptional level. CYP2C11 expression was more sensitive to suppression by glucagon at low insulin concentrations than at higher concentrations. (R(p))-Adenosine 3',5' cyclic monophosphothioate inhibited the down-regulation of CYP2C11 by S(p)-cAMPS, consistent with a competitive blockade of protein kinase A activation. These results suggest a role for glucagon in the down-regulation of CYP2C11 in diabetic rats, and provide a possible explanation for the known sensitivity of this cytochrome P450 to suppression in various stress and disease models. PMID- 11259543 TI - Natriuretic peptide-induced relaxation of myometrium from the pregnant guinea pig is not mediated by guanylate cyclase activation. AB - We tested both relaxation and cGMP generation by atrial (ANP), brain (BNP), and C type natriuretic peptide (CNP) in oxytocin-stimulated myometrium from near-term pregnant guinea pigs to investigate the ability and mechanism of natriuretic peptides to inhibit myometrial contractility. Myometrial strips were contracted by 10(-8) M oxytocin, and relaxation to the cumulative addition (10(-9)-10(-6) M) of the natriuretic peptides measured. Maximal relaxation to BNP was significantly greater than to ANP (52 versus 32% respectively; p < 0.05), whereas CNP failed to produce relaxation. However, the increase in cGMP produced by BNP (10(-7) M) was significantly less than that produced by ANP (10(-7) M) (4.5 versus 7.0 times basal; p < 0.05); CNP did not increase myometrial cGMP. Anantin, a competitive blocker of the guanylate cyclase A receptor, significantly reduced the increase in cGMP produced by ANP and BNP, but had no effect on relaxation induced by either peptide. Rp-8-Br-cGMP, an inhibitor of the cGMP-dependent protein kinase, did not alter BNP-induced relaxation. The atrial natriuretic peptide-fragment 4 23 amide, a natriuretic peptide clearance receptor agonist, failed to inhibit oxytocin-stimulated myometrial contraction. We conclude that natriuretic peptide induced relaxation of oxytocin-stimulated myometrium from the pregnant guinea pig is not mediated by either guanylate cyclase A or B activation, is independent of the cGMP pathway, and does not involve clearance receptor activation. Our results suggest that natriuretic peptide-induced relaxation of pregnant myometrium is mediated via a novel mechanism. PMID- 11259544 TI - Zebra finch CB1 cannabinoid receptor: pharmacology and in vivo and in vitro effects of activation. AB - Zebra finches (Taeniopygia guttata) learn vocal behavior during sensitive developmental periods, similar to the way in which human language is acquired. As adults, they recite the learned song pattern in a stereotyped manner. Previously, we demonstrated that central nervous system-associated cannabinoid receptors (CB1) are expressed in brain regions known to control both juvenile song learning and adult recitation of song. Here we extend these findings by establishing the zebra finch as a behavioral model to study cannabinoid pharmacology, showing that the cannabinoid agonist WIN55212-2 inhibits both adult song production and locomotor activity, effects that are antagonist-reversed. Through radioligand binding assays we investigated the pharmacology of a number of cannabinoid ligands representing all structural classes and established an affinity profile that can be compared with that of other species. To begin to characterize signal transduction mechanisms we isolated cDNA encoding the receptor protein. The zebra finch CB1 receptor (ZFCB1) is highly expressed in brain with amino acid sequence 92% identical to human CB1 receptor. Establishment of a Chinese hamster ovary cell line stably expressing ZFCB1 allowed demonstration that the cannabinoid agonist WIN55212-2 dose dependently and potently inhibits forskolin-stimulated adenylate cyclase activity (IC(50) = 9.0 nM, maximum inhibition = 49% at 100 nM WIN55212-2, reversed by 1 mM SR141716A). Cyclase inhibition indicates that ZFCB1 mediated signal transduction is consistent with that of mammalian CB1 receptors. Overall, cannabinoid inhibition of adult song production and conserved pharmacology render the zebra finch a promising model to investigate cannabinoid effects on learning by juveniles. PMID- 11259545 TI - Pharmacokinetic/pharmacodynamic modeling of antipyretic and anti-inflammatory effects of naproxen in the rat. AB - Pharmacokinetic/pharmacodynamic modeling was used to characterize the antipyretic and anti-inflammatory effects of naproxen in rats. An indirect response model was used to describe the antipyretic effects of naproxen after short intravenous infusions. The model assumes that basal temperature (T(a)) is maintained by the balance of fever mediators given by a constant (zero order) rate of synthesis (K(syn)), and a first order rate of degradation (K(out)). After an intraperitoneal injection of lipopolysaccharide, the change in T(a) was modeled assuming an increase in fever mediators described as an input rate function [IR(t)] estimated nonparametrically. An inhibitory E(max) model adequately described the inhibition of IR(t) by naproxen. A more complex model was used to describe the anti-inflammatory response of oral naproxen in the carrageenin induced edema model. Before carrageenin injection, physiological conditions are maintained by a balance of inflammation mediators given by K(syn) and K(out) (see above). After carrageenin injection, the additional synthesis of mediators is described by IR(t) (see above). Such mediators induced an inflammatory process, which is governed by a first order rate constant (K(IN)) that can be inhibited by the presence of naproxen in plasma. The sigmoidal E(max) model also well described the inhibition of K(IN) by naproxen. Estimates for IC(50) [concentration of naproxen in plasma eliciting half of maximum inhibition of IR(t) or K(IN)] were 4.24 and 4.13 microg/ml, for the antipyretic and anti inflammatory effects, respectively. PMID- 11259546 TI - Transcriptional and post-translational regulation of CYP1A1 by primaquine. AB - Regulation of the CYP1A1 gene has been shown to involve the aryl hydrocarbon receptor and the CYP1A1 gene expression is induced by AhR ligands. Primaquine is an antimalarial agent that does not exhibit the structural properties of a classical AhR ligand. We have evaluated the mechanisms by which this compound induces CYP1A1 expression using rat hepatoma H4IIE cells and V79 cells stably expressing CYP1A1. In H4IIE cells, primaquine caused a time- and dose-dependent increase of CYP1A1 mRNA and protein expression. The transcriptional activation of the CYP1A1 gene by primaquine was strictly XRE-dependent, as shown by transfection of different CYP1A1 pGL3 reporter constructs in H4IIE cells, and the involvement of the AhR was shown by activation of a Gal4-AhR hybrid protein by primaquine in transfected cells. Furthermore, primaquine caused transformation of the cytosolic AhR to a DNA-binding form, in vitro, suggesting that primaquine directly activates the receptor complex. In addition to its action at the transcriptional level, primaquine caused a dose-dependent inhibition of CYP1A1 degradation with an IC(50) of 3.3 , as seen in mammalian V79 cells. This was not due to the lysosomotropic activity of the drug since other lysosomotropic agents were ineffective. Primaquine formed a type II binding spectrum with CYP1A1 and inhibited the CYP1A1-dependent ethoxyresorufin O-deethylase activity in vitro with a K(i) of 1.3 microM, which is close to the IC(50), suggesting that the drug protects CYP1A1 from degradation by binding at the active site. It is concluded that CYP1A1 is regulated by primaquine both on the transcriptional as well as on a post-translational level. PMID- 11259547 TI - Enadoline discrimination in squirrel monkeys: effects of opioid agonists and antagonists. AB - Squirrel monkeys were trained to discriminate i.m. injections of the kappa-opioid receptor agonist enadoline (0.0017 mg/kg) from saline in a two-lever drug discrimination procedure. Enadoline produced a reliable discriminative stimulus that was reproduced by the kappa-selective agonists PD 117302, U 50,488, GR 89686A, (-)-spiradoline, ICI 204448, and EMD 61753, and by the mixed-action kappa/mu-agonists bremazocine and ethylketocyclazocine. The discriminative stimulus effects of enadoline were not reproduced by the mu-selective agonist morphine, the delta-selective agonist BW373U86, the mixed-action opioids nalbuphine and nalorphine, or by the less active enantiomers of enadoline and spiradoline PD 129829 and (+)-spiradoline, respectively. The selective mu-opioid antagonist beta-funaltrexamine (10.0 mg/kg) did not appreciably alter the dose effect function for enadoline in any subject. However, the nonselective and kappa selective opioid antagonists quadazocine (0.03-3.0 mg/kg) and nor-BNI (3-10 mg/kg), and the mixed-action opioid nalbuphine (0.3-30 mg/kg) served to surmountably antagonize enadoline's discriminative stimulus effects. The antagonist effects of nor-BNI were long-lasting and did not distinguish between drugs purported to act at different kappa-receptor subtypes. The present results bolster the view that common discriminative stimulus effects of enadoline and other opioids are mediated by kappa-agonist actions that are surmountably antagonized by nor-BNI in a long-lasting manner. The enadoline-antagonist effects of nalbuphine support the idea that it acts with low efficacy at kappa-opioid receptors. PMID- 11259548 TI - Effects of diaspirin cross-linked hemoglobin (DCLHb) during and post-CPR in swine. AB - The purpose of the study was to test the hypothesis that diaspirin cross-linked hemoglobin (DCLHb) can produce improved resuscitation during cardiac arrest. DCLHb, a derivative of human hemoglobin, has previously been demonstrated to produce a vasopressor response that is associated with increased blood flow to vital organs. In addition, it is an oxygen carrier. These effects may be beneficial to extreme low flow states, such as that during cardiac arrest and cardiopulmonary resuscitation (CPR). Experimental cardiac arrest and CPR were carried out in 32 anesthetized immature pigs. In each animal, ventricular fibrillation was induced for 5 min, followed by 10 min of standard CPR with a pneumatic device and room air ventilation. High (15 ml/kg) and low (5 ml/kg) doses of DCLHb or equivalent volume of normal saline were infused at the beginning of CPR in a random and blind manner. Cardiac output, organ blood flow, aortic pressure, coronary perfusion pressure, blood gases, and lactate concentrations were obtained before and during CPR. Following the 10-min CPR, the animals were defibrillated and the return of spontaneous circulation (ROSC) determined. DCLHb treatment achieved 75% ROSC compared with 25% in the saline group (p < 0.05). In addition, a better (p < 0.05) myocardial O(2) delivery, venous blood O(2) content, and myocardial and cerebral perfusion pressure were observed in the DCLHb group. DCLHb treatment during cardiac arrest and CPR significantly improves ROSC. This is most likely related to its improvement in coronary perfusion and myocardial oxygen delivery. PMID- 11259549 TI - A-85380 and epibatidine each interact with disparate spinal nicotinic receptor subtypes to achieve analgesia and nociception. AB - Nicotinic agonists, such as epibatidine (EPI) and A-85380, when administered systemically, elicit analgesia. Intrathecal EPI also produces analgesia accompanied by nociceptive and pressor responses. Since spinal administration of drugs offers a well defined pathway connecting the site of administration with behavioral and autonomic responses, we have compared the responses to intrathecal epibatidine and A-85380 to delineate the role of nicotinic acetylcholine receptors in spinal neurotransmission. Following implantation of intrathecal catheters in rats, we monitored cardiovascular, nociceptive, and antinociceptive responses after administration of various nicotinic receptor agonists. Consistent with A-85380 displacement of epibatidine from isolated spinal cord membranes, A 85380 elicited pressor, nociceptive, and antinociceptive responses similar to EPI. Antinociception was preceded by nociception. Both antinociception and nociception were blocked by mecamylamine, methyllycaconitine, and alpha-lobeline, but dihydro-beta-erythroidine only blocked the antinociceptive response. Whereas prior administration of EPI desensitized the nociceptive and antinociceptive responses to EPI, A-85380 pretreatment only desensitized EPI-elicited nociception and not antinociception. 2-Amino-5-phosphopentanoic acid pretreatment blocked the nociceptive response to A-85380, indicating A-85380 stimulated release of glutamate onto N-methyl-D-aspartate receptors to produce the irritant response of nociception. Intrathecal phentolamine virtually abolished A-85380 antinociception, but had no effect on EPI antinociception. Hence, analgesia can be produced by stimulation of distinct spinal preterminal nicotinic receptor subtypes, resulting in the release of neurotransmitters. In the case of A-85380, these sites primarily appear to be localized on adrenergic bulbospinal terminals. Our data suggest that A-85380 and EPI act at separate preterminal spinal sites as well as on distinct nicotinic receptor subtypes to elicit an antinociceptive response at the spinal level. PMID- 11259550 TI - Calcium channel blockade in vascular smooth muscle cells: major hypotensive mechanism of S-petasin, a hypotensive sesquiterpene from Petasites formosanus. AB - In vivo and in vitro studies were carried out to examine the putative hypotensive actions of S-petasin, a sesquiterpene extracted from the medicinal plant Petasites formosanus. Intravenous S-petasin (0.1-1.5 mg/kg) in anesthetized rats produced a dose-dependent hypotensive effect. In isolated aortic ring, isometric contraction elicited by KCl or the L-type Ca2+ channel agonist Bay K 8644 was reduced by S-petasin (0.1-100 microM), an action not affected by the cyclooxygenase inhibitor indomethacin, nitric-oxide synthase inhibitor N(omega) nitro-L-arginine, guanylyl cyclase inhibitor methylene blue, or removal of vascular endothelium. Pretreatment with S-petasin for 10 min shifted the concentration-response curve for KCl (15-90 mM)-induced contraction to the right and reduced the maximal response. In Ca2+-depleted and high K+-depolarized aortic rings preincubation with S-petasin attenuated the Ca2+-induced contraction in a concentration-dependent manner, suggesting that S-petasin reduced Ca2+ influx into vascular smooth muscle cells (VSMCs). Moreover, in cultured VSMCs, whole cell patch-clamp recording indicated that S-petasin (1-50 microM) inhibited the L type voltage-dependent Ca2+ channel (VDCC) activities. Intracellular Ca2+ concentration ([Ca2+[(i)) estimation using the fluorescent probe 1-[2-(5 carboxyoxazol-2-yl)-6-aminobenzofuran-5-oxy]-2-(2'-amino-5'-methylphenoxy)-ethane N,N,N,N-tetraacetic acid pentaacetoxymethyl ester indicated that S-petasin (10, 100 microM) suppressed the KCl-stimulated increase in ([Ca2+[(i)). Taken together, the results suggested that a direct Ca2+ antagonism of L-type VDCC in vascular smooth muscle may account, at least in part, for the hypotensive action of S-petasin. PMID- 11259551 TI - Zolpidem, triazolam, and diazepam decrease distress vocalizations in mouse pups: differential antagonism by flumazenil and beta-Carboline-3-carboxylate-t-butyl ester (beta-CCt). AB - In response to stressful events, neonatal mice emit ultrasonic vocalizations (USVs), which are suppressed by BZ agonists. The present study examined the role of the benzodiazepine/alpha1 (BZ/alpha1) receptor subtype in the suppression engendered by the BZ/alpha1-preferring agonist zolpidem and the nonselective BZ agonists triazolam and diazepam. The role of BZ receptor subtypes was explored further by conducting antagonism studies using the BZ/alpha1-preferring antagonist beta-carboline-3-carboxylate-t-butyl ester (beta-CCt), in comparison with the nonselective BZ antagonist flumazenil. Mouse pups (CFW strain) were separated from their dam and littermates at day 7, and placed for 4 min in a test chamber with reduced ambient temperature (19 +/- 1 degrees C) for recording USVs, motor incoordination (measured as a pup rolling on its back per grid cross), and body temperature. Zolpidem, triazolam, and diazepam suppressed USVs in a dose dependent manner, concomitant with increases in incoordination and augmentation of hypothermia. These effects of the three BZ agonists were blocked by flumazenil in a manner consistent with surmountable antagonism. The ability of zolpidem, but not triazolam or diazepam, to suppress USVs and augment hypothermia was antagonized by beta-CCt, whereas the increase in motor incoordination engendered by zolpidem, triazolam, and diazepam was not sensitive to beta-CCt administration. Collectively, these results suggest that zolpidem suppresses distress USVs in mouse pups by a mechanism distinct from that of typical BZs. Furthermore, suppression of distress USVs by zolpidem may involve BZ/alpha1 receptors and a nonanxiolytic mechanism, such as hypothermia. PMID- 11259552 TI - A novel sodium-hydrogen exchanger isoform-1 inhibitor, zoniporide, reduces ischemic myocardial injury in vitro and in vivo. AB - The cardioprotective efficacy of zoniporide (CP-597,396), a novel, potent, and selective inhibitor of the sodium-hydrogen exchanger isoform 1 (NHE-1), was evaluated both in vitro and in vivo using rabbit models of myocardial ischemia reperfusion injury. In these models, myocardial injury was elicited with 30 min of regional ischemia and 120 min of reperfusion. Zoniporide elicited a concentration-dependent reduction in infarct size (EC(50) of 0.25 nM) in the isolated heart (Langendorff) and reduced infarct size by 83% (50 nM). This compound was 2.5- to 20-fold more potent than either eniporide or cariporide (EC(50) of 0.69 and 5.11 nM, respectively), and reduced infarct size to a greater extent than eniporide (58% reduction in infarct size). In open-chest, anesthetized rabbits, zoniporide also elicited a dose-dependent reduction in infarct size (ED(50) of 0.45 mg/kg/h) and inhibited NHE-1-mediated platelet swelling (maximum inhibition 93%). Furthermore, zoniporide did not cause any in vivo hemodynamic (mean arterial pressure, heart rate, rate pressure product) changes. Zoniporide represents a novel class of potent NHE-1 inhibitors with potential utility for providing clinical cardioprotection. PMID- 11259553 TI - Structure-activity relationships and electrophysiological effects of short-acting amiodarone homologs in guinea pig isolated heart. AB - Antiarrhythmic agents with amiodarone-like electrophysiological actions, but with a more favorable pharmacokinetic profile than amiodarone would be extremely useful for the treatment of many tachyarrhythmias. We designed a series of amiodarone homologs with an alkyl ester group at position 2 of the benzofurane moiety. It was hypothesized that the electrophysiological and pharmacokinetic properties of these compounds are closely related to the size and branching of the ester group. The magnitude and time course of electrophysiological effects caused by methyl (ATI-2001), ethyl (ATI-2010), isopropyl (ATI-2064), sec-butyl (ATI-2042), and neopentyl (ATI-2054) homologs, and their common metabolite (ATI 2000) were investigated in guinea pig isolated heart. In paced hearts (atrial cycle length = 300 ms), each homolog (1 microM) was infused for 90 min followed by a 90-min washout. The stimulus-to-atrium (St-A), atrium-to-His bundle (AH), His bundle-to-ventricle (HV), QRS, and QT intervals, and ventricular monophasic action potential duration at 90% repolarization (MAPD(90)) were measured every 10 min. ATI-2001 and ATI-2064 significantly lengthened the St-A, HV, and QRS intervals, whereas ATI-2042 and ATI-2054 prolonged only the St-A interval. All compounds except the metabolite prolonged the AH interval. The relative rank order for the homologs to lengthen ventricular repolarization (MAPD(90)) was ATI 2042 > or = 2001 = 2010 = 2064 > 2054 > or = 2000. The metabolite was electrophysiologically inactive. Thus, modification of the benzofurane moiety ester group size and branching markedly altered the magnitude and time course of the electrophysiological effects caused by the ATI compounds. The different structure-activity relationships among the amiodarone homologs may have important consequences for further development of amiodarone-like antiarrhythmic agents. PMID- 11259554 TI - Anti-inflammatory and immunomodulatory potential of the novel PDE4 inhibitor roflumilast in vitro. AB - From a series of benzamide derivatives, roflumilast (3-cyclo-propylmethoxy-4 difluoromethoxy-N-[3,5-di-chloropyrid-4-yl]-benzamide) was identified as a potent and selective PDE4 inhibitor. It inhibits PDE4 activity from human neutrophils with an IC(50) of 0.8 nM without affecting PDE1 (bovine brain), PDE2 (rat heart), and PDE3 and PDE5 (human platelets) even at 10,000-fold higher concentrations. Roflumilast is almost equipotent to its major metabolite formed in vivo (roflumilast N-oxide) and piclamilast (RP 73401), however, more than 100-fold more potent than rolipram and Ariflo (cilomilast; SB 207499). The anti inflammatory and immunomodulatory potential of roflumilast and the reference compounds was investigated in various human leukocytes using cell-specific responses: neutrophils [N-formyl-methyl-leucyl-phenylalanine (fMLP)-induced formation of LTB(4) and reactive oxygen species (ROS)], eosinophils (fMLP- and C5a-induced ROS formation), monocytes, monocyte-derived macrophages, and dendritic cells (lipopolysaccharide-induced tumor necrosis factor-alpha synthesis), and CD4+ T cells (anti-CD3/anti-CD28 monoclonal antibody-stimulated proliferation, IL-2, IL-4, IL-5, and interferon-gamma release). Independent of the cell type and the response investigated, the corresponding IC values (for half-maximum inhibition) of roflumilast were within a narrow range (2-21 nM), very similar to roflumilast N-oxide (3-40 nM) and piclamilast (2-13 nM). In contrast, cilomilast (40-3000 nM) and rolipram (10-600 nM) showed greater differences with the highest potency for neutrophils. Compared with neutrophils and eosinophils, representing the terminal inflammatory effector cells, the relative potency of roflumilast and its N-oxide for monocytes, CD4+ T cells, and dendritic cells is substantially higher compared with cilomilast and rolipram, probably reflecting an improved immunomodulatory potential. The efficacy of roflumilast in vitro and in vivo (see accompanying article in this issue) suggests that roflumilast will be useful in the treatment of chronic inflammatory disorders such as asthma and chronic obstructive pulmonary disease. PMID- 11259555 TI - In vivo efficacy in airway disease models of roflumilast, a novel orally active PDE4 inhibitor. AB - We have investigated the bronchodilator and anti-inflammatory properties of roflumilast (3-cyclopropylmethoxy-4-difluoromethoxy-N-[3,5-dichloropyrid-4-yl] benzamide), a novel, highly potent, and selective phosphodiesterase 4 (PDE4) inhibitor. Additionally, we compared the effects of roflumilast and its N-oxide, the primary metabolite in vivo, with those of the PDE4 inhibitors piclamilast, rolipram, and cilomilast. Roflumilast inhibited the ovalbumin-evoked contractions of tracheal chains prepared from sensitized guinea pigs (EC(50) = 2 x 10(-7) M) but showed no relaxant effect on tissues contracted spontaneously. In spasmogen challenged rats and guinea pigs, intravenously administered roflumilast displayed bronchodilatory activity (ED(50) = 4.4 and 7.1 micromol/kg, respectively). Furthermore, roflumilast dose dependently attenuated allergen-induced bronchoconstriction in guinea pigs (ED(50) = 0.1 micromol/kg i.v.). Roflumilast given orally (ED(50) = 1.5 micromol/kg) showed equal potency to its N-oxide (ED(50) = 1.0 micromol/kg) but was superior to piclamilast (ED(50) = 8.3 micromol/kg), rolipram (ED(50) = 32.5 micromol/kg), and cilomilast (ED(50) = 52.2 micromol/kg) in suppressing allergen-induced early airway reactions. To assess the anti-inflammatory potential of orally administered roflumilast, antigen induced cell infiltration, total protein, and TNFalpha concentration in bronchoalveolar lavage fluid of Brown Norway rats were determined. Roflumilast and its N-oxide equally inhibited eosinophilia (ED(50) = 2.7 and 2.5 micromol/kg, respectively), whereas the reference inhibitors displayed lower potency (ED(50) = 17-106 micromol/kg). Besides, orally administered roflumilast abrogated LPS induced circulating TNFalpha in the rat (ED(50) = 0.3 micromol/kg), an effect shared by its N-oxide, with both molecules exhibiting 8-, 25-, and 310-fold superiority to piclamilast, rolipram, and cilomilast, respectively. These results, coupled with the in vitro effects of roflumilast on inflammatory cells, suggest that roflumilast represents a potential new drug for the treatment of asthma and chronic obstructive pulmonary disease. PMID- 11259556 TI - Effect of ovarian hormones and estrous cycle on stimulation of the hypothalamo pituitary-adrenal axis by cocaine. AB - Cocaine is known to exert sexually dimorphic HPA axis effects in rats and to disrupt estrous cyclicity and/or fertility in rats, nonhuman primates, and humans. The present studies investigated the reciprocal interactions between ovarian hormones and HPA axis responses to cocaine. Thirty minutes after injection, cocaine (15 mg/kg i.p.) increased serum ACTH and corticosterone more in cycling than ovariectomized females or male rats. ACTH and corticosterone were highest in proestrus when estradiol was elevated. Cocaine did not alter serum estradiol in females or testosterone in males but did stimulate progesterone secretion in both sexes. Cocaine-stimulated progesterone secretion was significantly greater in females than in males or ovariectomized females, and greater in proestrous than diestrous 1 rats. Cocaine stimulated corticosterone and progesterone secretion in sham-adrenalectomized, but not adrenalectomized rats, indicating that the adrenal gland and not the ovary is the source of cocaine-stimulated progesterone. Estrogen influenced cocaine-stimulated progesterone secretion more than corticosterone, suggesting different release mechanisms for the two steroids in the adrenal. These results suggest that adrenally derived progesterone could contribute to cocaine-induced physiological changes, including inhibited gonadotropin release. PMID- 11259557 TI - Beta(3)-adrenoceptor agonist-induced increases in lipolysis, metabolic rate, facial flushing, and reflex tachycardia in anesthetized rhesus monkeys. AB - The effects of two beta(3)-adrenergic receptor agonists, (R)-4-[4-(3 cyclopentylpropyl)-4,5-dihydro-5-oxo-1H-tetrazol-1-yl]-N-[4-[2-[[2-hydroxy-2-(3 pyridinyl)ethyl]amino]ethyl]phenyl]benzenesulfonamide and (R)-N-[4-[2-[[2-hydroxy 2-(3-pyridinyl)- ethyl]amino]ethyl]phenyl]-1-(4-octylthiazol-2-yl)-5 indolinesulfonamide, on indices of metabolic and cardiovascular function were studied in anesthetized rhesus monkeys. Both compounds are potent and specific agonists at human and rhesus beta(3)-adrenergic receptors. Intravenous administration of either compound produced dose-dependent lipolysis, increase in metabolic rate, peripheral vasodilatation, and tachycardia with no effects on mean arterial pressure. The increase in heart rate in response to either compound was biphasic with an initial rapid component coincident with the evoked peripheral vasodilatation and a second more slowly developing phase contemporaneous with the evoked increase in metabolic rate. Because both compounds exhibited weak binding to and activation of rhesus beta(1)-adrenergic receptors in vitro, it was hypothesized that the increase in heart rate may be reflexogenic in origin and proximally mediated via release of endogenous norepinephrine acting at cardiac beta(1)-adrenergic receptors. This hypothesis was confirmed by determining that beta(3)-adrenergic receptor agonist-evoked tachycardia was attenuated in the presence of propranolol and in ganglion-blocked animals, under which conditions there was no reduction in the evoked vasodilatation, lipolysis, or increase in metabolic rate. It is not certain whether the beta(3)-adrenergic receptor-evoked vasodilatation is a direct effect of compounds at beta(3)-adrenergic receptors in the peripheral vasculature or is secondary to the release or generation of an endogenous vasodilator. Peripheral vasodilatation in response to beta(3)-adrenergic receptor agonist administration was not attenuated in animals administered mepyramine, indomethacin, or calcitonin gene-related peptide(8-37). These findings are consistent with a direct vasodilator effect of beta(3)-adrenergic receptor agonists. PMID- 11259558 TI - Inducible nitric oxide synthase neutralizes carbamoylating potential of 1,3-bis(2 chloroethyl)-1-nitrosourea in c6 glioma cells. AB - Expression of iNOS in glioma and other tumors has been extensively documented but the effects of NO derived from iNOS on tumor-killing mechanisms of chemotherapy drugs remain to be fully defined. We note that increased NO synthesis by cytokine exposure or iNOS overexpression neutralized the cytotoxicity of 1,3-bis(2 chloroethyl)-1-nitrosourea (BCNU) and 1-(2-chloroethyl)-3-cyclohexyl-1 nitrosourea (CCNU), but not cisplatin, in rat C6 glioma cells. Suppression of BCNU cytotoxicity associated with iNOS overexpression could be abolished by pharmacological inhibition of NOS or coexpression of an antisense RNA against iNOS. Both BCNU and CCNU are chloroethylnitrosoureas that kill tumor cells via carbamoylating and alkylating actions. Further studies using compounds that each carry these different activities indicate that iNOS neutralized carbamoylating, but not alkylating, action of chloroethylnitrosoureas. Temozolomide, a novel chemotherapy drug recently available for treating brain tumors, carries only alkylating, but not carbamoylating, action. Overexpression of iNOS in C6 cells failed to neutralize temozolomide cytotoxicity. Results from the present study demonstrate the ability of iNOS-derived NO to confer chemoresistance against the carbamoylating potential of chloroethylnitrosoureas in vitro. Further investigation is needed to test whether iNOS expression, frequently noted in malignant brain tumors, also enhances chemoresistance against chloroethylnitrosoureas in vivo. PMID- 11259559 TI - Serotonergic manipulations both potentiate and reduce brain stimulation reward in rats: involvement of serotonin-1A receptors. AB - A discrete-trial current-threshold self-stimulation procedure was used to assess the effects of increased and decreased serotonergic neurotransmission, and 5 HT(1A) receptor activation on brain stimulation reward. Systemic administration of the 5-HT(1A) receptor agonist 8-OH-DPAT had a biphasic effect on brain reward thresholds, without affecting the latency to respond, a measure of performance. The low dose of 8-OH-DPAT (0.03 mg/kg) lowered reward thresholds, whereas higher doses (0.1 and 0.3 mg/kg) elevated thresholds. The 5-HT(1A) receptor antagonist p MPPI had no effect on brain stimulation behavior, but reversed both the 8-OH-DPAT induced lowering and elevation of thresholds, indicating that both of these effects of 8-OH-DPAT are mediated through the 5-HT(1A) receptor. Injections of 8 OH-DPAT into the median raphe nucleus also lowered brain reward thresholds, without affecting measures of performance, whereas injections of 8-OH-DPAT into the dorsal raphe nucleus had no effect. A high dose of the selective serotonin reuptake inhibitor fluoxetine (10 mg/kg) elevated reward thresholds and responses latencies, whereas lower doses (2.5 and 5.0 mg/kg) increased response latencies without affecting thresholds. Furthermore, the coadministration of a 5-HT(1A) antagonist, p-MPPI, and a previously ineffective dose of fluoxetine, a drug combination that increases serotonin levels, significantly elevated thresholds. Thus, it is suggested here that the reward-potentiating effects of systemically administered low doses of 8-OH-DPAT may be the result of reduced serotonergic neurotransmission, mediated by activation of 5-HT(1A) somatodendritic autoreceptors in the median, but not the dorsal, raphe nucleus. In conclusion, the present data support the hypothesis that serotonin exerts an inhibitory influence on reward processes. PMID- 11259560 TI - Application of the relative activity factor approach in scaling from heterologously expressed cytochromes p450 to human liver microsomes: studies on amitriptyline as a model substrate. AB - The relative activity factor (RAF) approach is being increasingly used in the quantitative phenotyping of multienzyme drug biotransformations. Using lymphoblast-expressed cytochromes P450 (CYPs) and the tricyclic antidepressant amitriptyline as a model substrate, we have tested the hypothesis that the human liver microsomal rates of a biotransformation mediated by multiple CYP isoforms can be mathematically reconstructed from the rates of the biotransformation catalyzed by individual recombinant CYPs using the RAF approach, and that the RAF approach can be used for the in vitro-in vivo scaling of pharmacokinetic clearance from in vitro intrinsic clearance measurements in heterologous expression systems. In addition, we have compared the results of two widely used methods of quantitative reaction phenotyping, namely, chemical inhibition studies and the prediction of relative contributions of individual CYP isoforms using the RAF approach. For the pathways of N-demethylation (mediated by CYPs 1A2, 2B6, 2C8, 2C9, 2C19, 2D6, and 3A4) and E-10 hydroxylation (mediated by CYPs 2B6, 2D6, and 3A4), the model-predicted biotransformation rates in microsomes from a panel of 12 human livers determined from enzyme kinetic parameters of the recombinant CYPs were similar to, and correlated with the observed rates. The model-predicted clearance via N-demethylation was 53% lower than the previously reported in vivo pharmacokinetic estimates. Model-predicted relative contributions of individual CYP isoforms to the net biotransformation rate were similar to, and correlated with the fractional decrement in human liver microsomal reaction rates by chemical inhibitors of the respective CYPs, provided the chemical inhibitors used were specific to their target CYP isoforms. PMID- 11259561 TI - General anesthetic potencies of a series of propofol analogs correlate with potency for potentiation of gamma-aminobutyric acid (GABA) current at the GABA(A) receptor but not with lipid solubility. AB - A series of 27 analogs of the general anesthetic propofol (2,6-diisopropylphenol) were examined for general anesthetic activity in Xenopus laevis tadpoles and for the ability to produce enhancement of submaximal GABA responses and/or direct activation at recombinant GABA(A) receptors. Fourteen of the propofol analogs produced loss of righting reflex in the tadpoles, whereas 13 were inactive as anesthetics. The same pattern of activity was noted with the actions of the compounds at the GABA(A) alpha(1)beta(2)gamma(2s) receptor. The potencies of the analogs as general anesthetics in tadpoles correlated better with potentiation of GABA responses than direct activation at the GABA(A) alpha(1)beta(2)gamma(2s) receptor. The calculated octanol/water partition coefficients for the analogs did not explain the lack of activity exhibited by the 13 nonanesthetic analogs, although this physicochemical parameter did correlate modestly with in vivo anesthetic potency. The actions of one nonanesthetic analog, 2,6-di-tert butylphenol, were examined in detail. 2,6-Di-tert-butylphenol was inactive at GABA(A) receptors, did not function as an anesthetic in the tadpoles, and did not antagonize any of the actions of propofol at GABA(A) receptors or in tadpoles. A key influence on the potency of propofol analogs appears to be the size and shape of the alkyl groups at positions 2 and 6 of the aromatic ring relative to the substituent at position 1. These data suggest steric constraints for the binding site for propofol on the GABA(A) receptor. PMID- 11259562 TI - Neurometer measurement of current stimulus threshold in rats. AB - We examined the current stimulus threshold in rats with the Neurometer, a device used clinically for measuring perception and pain thresholds. Although many studies have indicated the usefulness of this device in the quantification of nerve dysfunction in patients, we have found no published reports on the use of the Neurometer in animals. Transcutaneous nerve stimuli of the three sine-wave pulses produced by the Neurometer (at 2000, 250, and 5 Hz) were applied to plantar surface of rats. The intensity of each stimulation at which rats vocalized or were hardly startled was defined as the current stimulus threshold. With repeated stimulation, the thresholds were almost constant. Repeated topical application to the area around the stimulating electrode of a high concentration of capsaicin, which acts on small-diameter fibers, increased the thresholds at 250 and 5 Hz, but did not affect the 2000-Hz threshold. Intravenous morphine (2-5 mg/kg) increased all three thresholds, whereas intrathecal morphine (20 or 80 microg) increased only the 5-Hz threshold. Intravenous injection of a minor tranquilizer, diazepam, at 1 mg/kg raised the thresholds at 2000 and 250 Hz, but did not affect the 5-Hz threshold. Higher dose of diazepam increased all three thresholds. These results suggest that the Neurometer makes possible selective examination of subsets of nerve fibers that differ in diameter not only in humans but also in animals. The present study in rats, in which we established a method of measurement, may provide helpful suggestions for the interpretation of data in humans. PMID- 11259563 TI - Antagonism of 5-hydroxytryptamine(2a) receptors attenuates the behavioral effects of cocaine in rats. AB - Serotonin (5-hydroxytryptamine; 5-HT) 5-HT(2A) receptors have been shown to modulate dopamine (DA) function and a more thorough appreciation of this modulatory interaction between 5-HT2A receptors and DA systems may yield insight into novel approaches to treatment of cocaine dependence. The present study examined the effects of two ligands with varying selectivity for 5-HT2A receptors on the locomotor stimulant and discriminative stimulus effects of cocaine in male rats. Locomotor activity was measured following intraperitoneal injection of vehicle (1 ml/kg), the selective 5-HT2A receptor antagonist M100907 [R-(+)-(2,3 dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidine-methanol] (0.02-2.0 mg/kg), or the 5-HT(2) receptor antagonist ketanserin (0.04-4 mg/kg) 45 min before administration of saline (1 ml/kg) or cocaine (10 mg/kg); monitoring of activity in photobeam chambers began at once and proceeded for 1 h. Neither M100907 nor ketanserin significantly altered basal locomotor activity, but both drugs attenuated cocaine-induced hyperactivity (p < 0.05). In drug discrimination studies, rats were trained to discriminate cocaine (10 mg/kg) from saline (1 ml/kg) in a two-lever, water-reinforced operant task. M100907 (0.05-1.6 mg/kg) and ketanserin (0.05-4 mg/kg) evoked a dose-related attenuation of the stimulus effects of cocaine (5 mg/kg, p < 0.05). These results suggest that 5-HT2A receptors play an important role in the behavioral effects of cocaine and that 5 HT2A receptors should be considered a viable target for analysis in the search for pharmacotherapies useful in the treatment of cocaine dependence. PMID- 11259564 TI - Antinociceptive and respiratory effects of intrathecal H-Tyr-D-Arg-Phe-Lys-NH2 (DALDA) and [Dmt1] DALDA. AB - DALDA (H-Tyr-D-Arg-Phe-Lys-NH(2)) and [Dmt(1)]DALDA (H-Dmt-D-Arg-Phe-Lys-NH(2)) (Dmt = 2',6'-dimethyltyrosine) are potent and highly selective mu-opioid agonists (K(i)(delta)/K(i)(mu) > 10,000 and K(i)(kappa)/K(i)(mu) > 100). Both peptides carry a 3+ charge at physiological pH. Their antinociceptive and respiratory effects were compared with morphine (MOR) after intrathecal administration in rats. Both DALDA and [Dmt1]DALDA produced dose-dependent and naloxone-reversible antinociceptive effects with relative potencies of 14 and 3000x that of MOR. The antinociceptive potency of [Dmt1]DALDA far exceeded its affinity and potency at the mu-opioid receptor and may be explained by its ability to inhibit norepinephrine (NE) uptake in spinal cord synaptosomes. The antinociceptive response to [Dmt1]DALDA was significantly attenuated by the alpha(2)-adrenergic antagonist yohimbine. Thus, [Dmt1]DALDA may be regarded as a drug with dual actions, and its antinociceptive potency is better described by both its affinity and potency at mu-opioid receptors, and its potency at inhibiting NE uptake. The analgesic duration of an equipotent dose of MOR, DALDA, and [Dmt1]DALDA was 3, 7, and 13 h, respectively, and the long duration may be due to the hydrophilic nature of these peptide analogs. Respiratory effects were determined using whole body plethysmography at 3 and 30x the antinociceptive ED(50). A significant depression in minute ventilation was observed with the higher dose of morphine and both doses of DALDA, but not with either dose of [Dmt1]DALDA. Because of its high antinociceptive potency, long duration of action, and low propensity to induce respiratory depression, [Dmt1]DALDA is of interest as a drug candidate for intrathecal analgesia. PMID- 11259565 TI - Biological characterization of eugeniin as an anti-herpes simplex virus type 1 compound in vitro and in vivo. AB - Eugeniin exhibits antiviral activity against acyclovir and phosphonoacetic acid (PAA)-resistant herpes simplex virus type 1 (HSV-1) as well as the wild-type HSV 1 in vitro. In this study, we characterized the biological activity of eugeniin in cutaneously HSV-1-infected mice and its interaction with HSV-1 DNA polymerase. The oral and intraperitoneal administrations of eugeniin at 0.3 mg/kg showed similar therapeutic efficacy in retarding the development of skin lesions of HSV 1-infected mice. The two routes of administration at 6 or 50 mg/kg significantly prolonged the mean survival times and/or reduced mortality without toxicity. The oral administration of eugeniin at 50 mg/kg reduced virus yields in the skin and brain of infected mice. Thus, the therapeutic efficacy of oral administration at the various doses of eugeniin was similar to that of intraperitoneal administration, suggesting that the oral bioavailability of eugeniin was high with respect to absorption. Furthermore, the anti-HSV-1 activity of eugeniin was characterized by isobolograms analyzing its combined effects with acyclovir or PAA in HSV-1-infected Vero cells. Eugeniin enhanced the anti-HSV-1 activity of acyclovir but was suggested to be antagonistic with PAA. The interaction of eugeniin and PAA on the activity of partially purified HSV-1 DNA polymerase suggested that eugeniin interacted with the polymerase in the vicinity of PAA binding site. Thus, eugeniin showed different anti-HSV-1 action from acyclovir and PAA and therapeutic anti-HSV-1 activity in mice. PMID- 11259566 TI - The nitroderivative of aspirin, NCX 4016, reduces infarct size caused by myocardial ischemia-reperfusion in the anesthetized rat. AB - NCX 4016, a nitro-ester of aspirin endowed with antithrombotic activity, appears to have clinical potential in treating cardiac complications related to coronary insufficiency. This compound has been shown to improve postischemic ventricular dysfunction and to reduce myocardial infarct size in the rabbit. The cardioprotection conferred by NCX 4016 (10, 30, and 100 mg/kg) and aspirin (ASA, 54 mg/kg) was evaluated in anesthetized rats subjected to 30 min of myocardial ischemia followed by 120 min of reperfusion (MI/R). Drugs were given orally for 5 consecutive days. NCX 4016 displayed remarkable cardioprotection in rats subjected to MI/R as was evident in the reduction of ventricular premature beats and in the incidence of ventricular tachycardia and fibrillation; they were reduced dose dependently and correlated with survival of all rats treated with the higher dose of NCX 4016. In these animals, infarct size was restricted proportionally to the dose of NCX 4016 associated with diminution of both plasma creatine phosphokinase and cardiac myeloperoxidase activities. ASA showed only a minor degree of protection against MI/R damage. Rats treated with N(G)-nitro-L arginine methyl ester (L-NAME, 10 mg/kg) demonstrated aggravated myocardial damage in terms of arrhythmias, mortality, and infarct size. Supplementation of nitric oxide (NO) with NCX 4016 (100 mg/kg) greatly reduced the worsening effect caused by L-NAME. The beneficial effects of NCX 4016 appear to derive in large part from the NO moiety, which modulates a number of cellular events leading to inflammation, obstruction of the coronary microcirculation, arrhythmias, and myocardial necrosis. PMID- 11259567 TI - Rebamipide suppresses formyl-methionyl-leucyl-phenylalanine (fMLP)-induced superoxide production by inhibiting fMLP-receptor binding in human neutrophils. AB - The purpose of the present work was to investigate the mechanism underlying the inhibitory action of rebamipide on superoxide anion (O2) production induced by the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP) in human neutrophils. Phosphatidylinositol 3,4,5-trisphosphate (PIP(3)), a product of phosphoinositide 3-OH-kinase (PI 3-kinase) accumulated in response to fMLP and this accumulation was well correlated with O2 production in human neutrophils. Rebamipide inhibited PIP(3) production in parallel with the inhibition of fMLP induced O2 production. PI 3-kinase activity in anti-PI 3-kinase p85 immunoprecipitates was not affected by the presence of rebamipide, therefore rebamipide did not have a direct inhibitory action on PI 3-kinase activity. Since rebamipide had no inhibitory effect on O2 production induced by NaF, a direct activator of G protein, the target of the inhibitory action of rebamipide appears to be a component of the signal transduction pathway upstream of G protein. Scatchard analysis of [3H]fMLP binding to human neutrophil membrane revealed that rebamipide increased the K(D) value of [3H]fMLP without altering the number of [3H]fMLP binding sites, suggesting that rebamipide has a competitive antagonistic action against the fMLP-receptor. The competitive antagonistic action was further confirmed by the finding that rebamipide caused a parallel shift to the right in the dose-response curve of O2 production induced by fMLP. These results provide evidence that the competitive inhibitory action of rebamipide on the fMLP receptor plays a main role in its inhibitory action on fMLP-induced O2 production. PMID- 11259568 TI - A 5-HT(7) receptor-mediated depolarization in the anterodorsal thalamus. I. Pharmacological characterization. AB - Little is currently known regarding the electrophysiological response elicited by 5-hydroxytryptamine-7 (5-HT(7)) receptor stimulation in the brain. Previous anatomical studies have shown that the anterior thalamus expresses a high density of 5-HT7 receptors. Therefore, we used whole-cell recording techniques in the in vitro brain slices to examine the effects of serotonin on neurons of the anterodorsal nucleus of the thalamus (ADn). Bath application of 5-HT induces a large membrane depolarization and inward current in neurons of the ADn. Since these cells expressed 5-HT7 receptor mRNA, as determined by single-cell reverse transcriptase-polymerase chain reaction, we pharmacologically characterized the 5 HT receptor mediating this response. We found that the 5-HT1 and 5-HT7 agonists 5 carboxamidotryptamine (5-CT) and 5-methoxytryptamine mimicked the response to 5 HT, whereas the 5-HT2 agonist 2,5-dimethoxy-4-iodoamphetamine did not. Consistent with the involvement of a 5-HT7 receptor, 5-CT was approximately 18 times more potent than 5-HT. Furthermore, administration of the 5-HT(1A) and 5-HT7 agonist 8 hydroxydipropylaminotetralin mimicked and antagonized the effect of serotonin, suggesting it acted as a partial agonist. To determine if either the 5-HT1 or 5 HT7 receptor mediated the 5-HT-induced inward current, we used antagonists. We found that the 5-HT7 ligands ritanserin, methylsergide, LSD, and mesulergine could inhibit the 5-HT-induced inward current, whereas the 5-HT1 antagonist cyanopindolol had no effect. The pA(2) value determined for mesulergine closely approximated that expected for a 5-HT7 receptor. Finally, we found that bath application of the selective antagonist SB-269770 blocks the 5-HT-induced inward current. These results identify the receptor mediating the serotonin-induced membrane depolarization in the ADn as the 5-HT7 subtype. PMID- 11259569 TI - A 5-HT(7) receptor-mediated depolarization in the anterodorsal thalamus. II. Involvement of the hyperpolarization-activated current I(h). AB - Previous studies have shown that 5-hydroxytryptamine (5-HT) can modulate the hyperpolarization-activated nonselective cation current (I(h)) to elicit a membrane depolarization in neurons. However, the receptor subtype involved in this response remains controversial. In the accompanying study, we have identified a 5-HT7 receptor-mediated depolarization in the anterodorsal nucleus of the thalamus (ADn). In the present study, we have examined the possible role of I(h) in mediating this 5-HT7 receptor-mediated depolarization. We used the blind tight-seal patch clamp technique to examine the ability of 5-HT to modulate I(h) in the ADn. We found that 5-HT induced a shift in the voltage dependence of I(h) to more depolarized potentials. The pharmacology of the receptor mediating this effect was consistent with that of a 5-HT7 receptor. Since the 5-HT7 receptor is coupled positively to adenylate cyclase, we examined the cAMP dependence of the 5-HT-induced modulation of I(h). Intracellular addition of cAMP mimicked and occluded the 5-HT response. Conversely, in the presence of the protein kinase inhibitors H-8 and staurosporine, ADn neurons still expressed a 5 HT-induced shift in the voltage dependence of I(h). These results suggest that 5 HT regulates I(h) in the ADn through a cAMP-dependent but protein kinase A (PKA) independent mechanism. To determine the contribution of I(h) to the 5-HT7 receptor-mediated depolarization, we used the selective I(h) blocker ZD7288. This compound greatly reduced the depolarizing response elicited by activation of 5 HT7 receptors. We conclude that 5-HT7 receptors depolarize ADn neurons primarily by increasing I(h) through a cAMP-dependent, PKA-independent mechanism. PMID- 11259570 TI - Metabolism of levo-alpha-Acetylmethadol (LAAM) by human liver cytochrome P450: involvement of CYP3A4 characterized by atypical kinetics with two binding sites. AB - levo-alpha-Acetylmethadol (LAAM) is a long-acting opioid agonist prodrug used for preventing opiate withdrawal. LAAM undergoes bioactivation via sequential N demethylation to nor-LAAM and dinor-LAAM, which are more potent and longer-acting than LAAM. This study examined LAAM and nor-LAAM metabolism using human liver microsomes, cDNA-expressed CYP, CYP isoform-selective chemical inhibitors, and monoclonal antibody to determine kinetic parameters for predicting in vivo drug interactions, involvement of constitutive CYP isoforms, and mechanistic aspects of sequential N-demethylation. N-Demethylation of LAAM and nor-LAAM by human liver microsomes exhibited biphasic Eadie-Hofstee plots. Using a dual-enzyme Michaelis-Menten model, K(m) values were 19 and 600 microM for nor-LAAM and 4 and 450 microM for dinor-LAAM formation, respectively. LAAM and nor-LAAM metabolism was inhibited by the CYP3A4-selective inhibitors troleandomycin, erythromycin, ketoconazole, and midazolam. Of the cDNA-expressed isoforms examined, CYP2B6 and 3A4 had the highest activity toward LAAM and nor-LAAM at both low (2 microM) and high (250 microM) substrate concentrations. N-Demethylation of LAAM and nor-LAAM by expressed CYP3A4 was unusual, with hyperbolic velocity curves and Eadie Hofstee plots and without evidence of positive cooperativity. Using a two-site model, K(m) values were 6 and 0.2 microM, 1250 and 530 microM, respectively. Monoclonal antibody against CYP2B6 inhibited CYP2B6-catalyzed but not microsomal LAAM or nor-LAAM metabolism, whereas troleandomycin inhibited metabolism in all microsomes studied. The ratio [dinor-LAAM/(nor-LAAM plus dinor-LAAM)] with microsomes and CYP3A4 decreased with increasing LAAM concentration, suggesting most dinor-LAAM is formed from released nor-LAAM that subsequently reassociates with CYP3A4. Based on these results, we conclude that LAAM and nor-LAAM are predominantly metabolized by CYP3A4 in human liver microsomes, and CYP3A4 exhibits unusual multisite kinetics. PMID- 11259571 TI - Futile cycling of estrone sulfate and estrone in the recirculating perfused rat liver preparation. AB - The futile cycling of estrone sulfate (E(1)S) and estrone (E1) was investigated in the recirculating, perfused, rat liver preparation. Although E(1)S was not distributed into bovine erythrocytes, the compound was highly bound to albumin [4% bovine serum albumin (BSA), unbound fraction of 0.03 +/- 0.01]. By contrast, E1 was bound and metabolized to estradiol (E2) by bovine erythrocytes, with metabolic clearances of 0.061 to 0.069 ml/min when normalized to the hematocrit. Due to strong binding of E1 to albumin, BSA (4%) greatly reduced the red cell clearance to a minimum (0.0024 to 0.0031 ml/min/unit of hematocrit). Despite the low unbound fractions of E(1)S (0.027 +/- 0.004) and E1 (0.036 +/- 0.006), clearances of the simultaneously delivered tracers [(3)H]E(1)S and [(14)C]E1 in perfusate (4% BSA and 20% erythrocytes) by the recirculating, perfused rat liver (flow rate of 0.91 +/- 0.1 ml/min/g of liver) were high (0.53 +/- 0.08 and 0.85 +/- 0.2 ml/min/g of liver, respectively). Although low levels of [(3)H]E1 were observed following the tracer [(3)H]E(1)S, both parent and metabolite species displayed similar decay half-lives that were characteristic of compounds undergoing futile cycling. The same decay profile was observed for [(14)C]E(1)S but the half-life of administered [(14)C]E1 was shorter in comparison. A series compartment liver model that incorporated previously noted heterogeneity in estrone sulfation and glucuronidation activities among periportal and perivenous hepatocytes, and homogeneity in sinusoidal transport and desulfation was used to explain the discrepant half-lives. The model described a high partitioning of E1 in the endoplasmic reticulum and the segregation of estrone sulfation activities in the cytosolic space from the desulfation and glucuronidation activities in the endoplasmic reticulum space. PMID- 11259572 TI - Blockade of currents by the antimalarial drug chloroquine in feline ventricular myocytes. AB - The effects of the antimalarial drug chloroquine on cardiac action potential and membrane currents were studied at clinically relevant concentrations. In cat Purkinje fibers, chloroquine at concentrations of 0.3 to 10 microM increased action potential duration, and reduced maximum upstroke velocity. At concentrations of 3 and 10 microM, chloroquine increased automaticity and reduced maximum diastolic potential, and after 60 min of perfusion with a concentration 10 microM, spontaneous activity was abolished. In isolated cat ventricular myocytes, chloroquine also increased action potential duration in a concentration dependent manner, and reduced resting membrane potential at 3 and 10 microM. In voltage-clamped cat ventricular myocytes, chloroquine blocked several inward and outward membrane currents. The order of potency was inward rectifying potassium current (I(K1)) > rapid delayed rectifying potassium current (I(Kr)) > sodium current (I(Na)) > L-type calcium current (I(Ca-L)). Only tonic block of I(Na) and I(Ca-L) was observed at a stimulation frequency of 0.1 Hz and no additional blockade was observed during stimulation trains applied at 1 Hz. The effect of chloroquine on I(K1) was voltage-dependent, with less pronounced blockade at negative test potentials. In addition, unblock was achieved by hyperpolarizing pulses to potentials negative to the current reversal potential. Chloroquine blocked the rapid component of the delayed rectifying outward current, I(Kr,) but not the slow component, I(Ks). These findings provide the cellular mechanisms for the prolonged QT interval, impaired ventricular conduction, and increased automaticity induced by chloroquine, which have been suggested as responsible for the proarrhythmic effects of the drug. PMID- 11259573 TI - Sex-related differences in antinociception and tolerance development following chronic intravenous infusion of morphine in the rat: modulatory role of testosterone via morphine clearance. AB - This study investigated possible sex-related differences in levels of antinociception and the rate of development of tolerance to the antinociceptive effects following prolonged (48 h) intravenous (i.v.) morphine administration in the rat. Groups of adult intact male, castrated male, female, and testosterone pretreated female Sprague-Dawley rats received prolonged (48 h) infusions of i.v. morphine (5 or 10 mg/day) plus intra-arterial (i.a.) saline or i.v. morphine (5 mg/day) plus i.a. chloramphenicol (300 mg/day). Antinociception was quantified using the hotplate test. Serum concentrations of morphine and morphine-3 glucuronide (M3G) were assayed using high performance liquid chromatography with electrochemical detection, whereas the serum testosterone concentrations were quantified using an enzyme-linked immunosorbent assay method. Consistent with our previous findings in intact male rats, prolonged coinfusion of chloramphenicol with morphine produced a marked increase in the extent and duration of morphine antinociception in all experimental groups. Additionally, female and castrated male rats developed tolerance more slowly than either intact male or testosterone pretreated female rats, when coinfused with parenteral morphine plus chloramphenicol. However, mean levels of antinociception were not significantly correlated with either the mean serum morphine or M3G concentrations, but were significantly inversely correlated with the mean values of the M3G/morphine serum molar concentration ratio. Testosterone pretreatment of female rats for 1 week before chronic morphine infusion abolished antinociception and markedly reduced both the serum morphine and M3G concentrations. These latter findings imply that testosterone modulates antinociception evoked by prolonged morphine infusion in rats via a mechanism that appears to involve modulation of morphine metabolism. PMID- 11259574 TI - In vitro and in vivo pharmacological characterization of BIIL 284, a novel and potent leukotriene B(4) receptor antagonist. AB - BIIL 284 is a new LTB(4) receptor antagonist. It is a prodrug and has negligible binding to the LTB(4) receptor. However, ubiquitous esterases metabolize BIIL 284 to the active metabolites BIIL 260 and BIIL 315, the glucuronidated form of BIIL 260. Both metabolites have high affinity to the LTB(4) receptor on isolated human neutrophil cell membranes with K(i) values of 1.7 and 1.9 nM, respectively. On vital human neutrophilic granulocytes K(i) was around 1 nM. BIIL 260 and BIIL 315 interact with the LTB(4) receptor in a saturable, reversible, and competitive manner. BIIL 260 and its glucuronide BIIL 315 also potently inhibited LTB(4) induced intracellular Ca(2+) release in human neutrophils (IC(50) values of 0.82 and 0.75 nM, respectively) as measured with Fura-2. High efficacy of BIIL 284 has been demonstrated in various in vivo models. BIIL 284 inhibited LTB(4)-induced mouse ear inflammation with ED(50) = 0.008 mg/kg p.o., LTB(4)-induced transdermal chemotaxis in guinea pigs with ED(50) = 0.03 mg/kg p.o., LTB(4)-induced neutropenia in various species (monkey: ED(50) = 0.004 mg/kg p.o.), and LTB(4) induced Mac1-expression in monkeys (ED(50) = 0.05 mg/kg p.o. in Tylose). Full blockade of LTB(4) receptors over 24 h was achieved by 0.3 mg/kg BIIL 284 after single oral dose as measured by LTB(4)-induced neutropenia or Mac1-expression in the monkey model. BIIL 284 is an unusually potent and long-acting orally active LTB(4) antagonist, and is therefore under clinical development as a novel anti inflammatory principle. PMID- 11259575 TI - MLL and CREB bind cooperatively to the nuclear coactivator CREB-binding protein. AB - A fragment of the mixed-lineage leukemia (MLL) gene (Mll, HRX, ALL-1) was identified in a yeast genetic screen designed to isolate proteins that interact with the CREB-CREB-binding protein (CBP) complex. When tested for binding to CREB or CBP individually, this MLL fragment interacted directly with CBP, but not with CREB. In vitro binding experiments refined the minimal region of interaction to amino acids 2829 to 2883 of MLL, a potent transcriptional activation domain, and amino acids 581 to 687 of CBP (the CREB-binding or KIX domain). The transactivation activity of MLL was dependent on CBP, as either adenovirus E1A expression, which inhibits CBP activity, or alteration of MLL residues important for CBP interaction proved effective at inhibiting MLL-mediated transactivation. Single amino acid substitutions within the MLL activation domain revealed that five hydrophobic residues, potentially forming a hydrophobic face of an amphipathic helix, were critical for the interaction of MLL with CBP. Using purified components, we found that the MLL activation domain facilitated the binding of CBP to phosphorylated CREB. In contrast with paradigms in which factors compete for limiting quantities of CBP, these results reveal that two distinct transcription factor activation domains can cooperatively target the same motif on CBP. PMID- 11259576 TI - The growth suppressor PML represses transcription by functionally and physically interacting with histone deacetylases. AB - The growth suppressor promyelocytic leukemia protein (PML) is disrupted by the chromosomal translocation t(15;17) in acute promyelocytic leukemia (APL). PML plays a key role in multiple pathways of apoptosis and regulates cell cycle progression. The present study demonstrates that PML represses transcription by functionally and physically interacting with histone deacetylase (HDAC). Transcriptional repression mediated by PML can be inhibited by trichostatin A, a specific inhibitor of HDAC. PML coimmunoprecipitates a significant level of HDAC activity in several cell lines. PML is associated with HDAC in vivo and directly interacts with HDAC in vitro. The fusion protein PML-RARalpha encoded by the t(15;17) breakpoint interacts with HDAC poorly. PML interacts with all three isoforms of HDAC through specific domains, and its expression deacetylates histone H3 in vivo. Together, the results of our study show that PML modulates histone deacetylation and that loss of this function in APL alters chromatin remodeling and gene expression. This event may contribute to the development of leukemia. PMID- 11259577 TI - Association of insulin receptor substrate 1 (IRS-1) y895 with Grb-2 mediates the insulin signaling involved in IRS-1-deficient brown adipocyte mitogenesis. AB - We have recently generated immortalized fetal brown adipocyte cell lines from insulin receptor substrate 1 (IRS-1) knockout mice and demonstrated an impairment in insulin-induced lipid synthesis as compared to wild-type cell lines. In this study, we investigated the consequences of IRS-1 deficiency on mitogenesis in response to insulin. The lack of IRS-1 resulted in the inability of insulin stimulated IRS-1-deficient brown adipocytes to increase DNA synthesis and enter into S/G2/M phases of the cell cycle. These cells showed a severe impairment in activating mitogen-activated protein kinase kinase (MEK1/2) and p42-p44 mitogen activated protein kinase (MAPK) upon insulin stimulation. IRS-1-deficient cells also lacked tyrosine phosphorylation of SHC and showed no SHC-Grb-2 association in response to insulin. The mitogenic response to insulin could be partially restored by enhancing IRS-2 tyrosine phosphorylation and its association with Grb 2 by inhibition of phosphatidylinositol 3-kinase activity through a feedback mechanism. Reconstitution of IRS-1-deficient brown adipocytes with wild-type IRS 1 restored insulin-induced IRS-1 and SHC tyrosine phosphorylation and IRS-1-Grb 2, IRS-1-SHC, and SHC-Grb-2 associations, leading to the activation of MAPK and enhancement of DNA synthesis. Reconstitution of IRS-1-deficient brown adipocytes with the IRS-1 mutant Tyr895Phe, which lacks IRS-1-Grb-2 binding, restored SHC IRS-1 association and SHC-Grb-2 association. However, the lack of IRS-1-Grb-2 association impaired MAPK activation and DNA synthesis in insulin-stimulated mutant cells. These data provide strong evidence for an essential role of IRS-1 and its direct association with Grb-2 in the insulin signaling pathway leading to MAPK activation and mitogenesis in brown adipocytes. PMID- 11259578 TI - Strong functional interactions of TFIIH with XPC and XPG in human DNA nucleotide excision repair, without a preassembled repairosome. AB - In mammalian cells, the core factors involved in the damage recognition and incision steps of DNA nucleotide excision repair are XPA, TFIIH complex, XPC HR23B, replication protein A (RPA), XPG, and ERCC1-XPF. Many interactions between these components have been detected, using different physical methods, in human cells and for the homologous factors in Saccharomyces cerevisiae. Several human nucleotide excision repair (NER) complexes, including a high-molecular-mass repairosome complex, have been proposed. However, there have been no measurements of activity of any mammalian NER protein complex isolated under native conditions. In order to assess relative strengths of interactions between NER factors, we captured TFIIH from cell extracts with an anti-cdk7 antibody, retaining TFIIH in active form attached to magnetic beads. Coimmunoprecipitation of other NER proteins was then monitored functionally in a reconstituted repair system with purified proteins. We found that all detectable TFIIH in gently prepared human cell extracts was present in the intact nine-subunit form. There was no evidence for a repair complex that contained all of the NER components. At low ionic strength TFIIH could associate with functional amounts of each NER factor except RPA. At physiological ionic strength, TFIIH associated with significant amounts of XPC-HR23B and XPG but not other repair factors. The strongest interaction was between TFIIH and XPC-HR23B, indicating a coupled role of these proteins in early steps of repair. A panel of antibodies was used to estimate that there are on the order of 10(5) molecules of each core NER factor per HeLa cell. PMID- 11259579 TI - Role of TATA binding protein (TBP) in yeast ribosomal dna transcription by RNA polymerase I: defects in the dual functions of transcription factor UAF cannot be suppressed by TBP. AB - Initiation of ribosomal DNA (rDNA) transcription by RNA polymerase I (Pol I) in the yeast Saccharomyces cerevisiae involves upstream activation factor (UAF), core factor, the TATA binding protein (TBP), and Rrn3p in addition to Pol I. We found previously that yeast strains carrying deletions in the UAF component RRN9 switch completely to the use of Pol II for rRNA transcription, with no residual Pol I transcription. These polymerase-switched strains initially grow very slowly, but subsequent expansion in the number of rDNA repeats on chromosome XII leads to better growth. Recently, it was reported that TBP overexpression could bypass the requirement of UAF for Pol I transcription in vivo, producing nearly wild-type levels of growth in UAF mutant strains (P. Aprikian, B. Moorefield, and R. H. Reeder, Mol. Cell. Biol. 20:5269-5275, 2000). Here, we demonstrate that deletions in the UAF component RRN5, RRN9, or RRN10 lead to Pol II transcription of rDNA. TBP overexpression does not suppress UAF mutation, and these strains continue to use Pol II for rRNA transcription. We do not find evidence for even low levels of Pol I transcription in UAF mutant strains carrying overexpressed TBP. In diploid strains lacking both copies of the UAF component RRN9, Pol II transcription of rDNA is more strongly repressed than in haploid strains but TBP overexpression still fails to activate Pol I. These results emphasize that UAF plays an essential role in activation of Pol I transcription and silencing of Pol II transcription of rDNA and that TBP functions to recruit the Pol I machinery in a manner completely dependent on UAF. PMID- 11259580 TI - PSF is a novel corepressor that mediates its effect through Sin3A and the DNA binding domain of nuclear hormone receptors. AB - Members of the type II nuclear hormone receptor subfamily (e.g., thyroid hormone receptors [TRs], retinoic acid receptors, retinoid X receptors [RXRs], vitamin D receptor, and the peroxisome proliferator-activated receptors) bind to their response sequences with or without ligand. In the absence of ligand, these DNA bound receptors mediate different degrees of repression or silencing of gene expression which is thought to result from the association of their ligand binding domains (LBDs) with corepressors. Two related corepressors, N-CoR and SMRT, interact to various degrees with the LBDs of these type II receptors in the absence of their cognate ligands. N-CoR and SMRT have been proposed to act by recruiting class I histone deacetylases (HDAC I) through an association with Sin3, although they have also been shown to recruit class II HDACs through a Sin3 independent mechanism. In this study, we used a biochemical approach to identify novel nuclear factors that interact with unliganded full-length TR and RXR. We found that the DNA binding domains (DBDs) of TR and RXR associate with two proteins which we identified as PSF (polypyrimidine tract-binding protein associated splicing factor) and NonO/p54(nrb). Our studies indicate that PSF is a novel repressor which interacts with Sin3A and mediates silencing through the recruitment of HDACs to the receptor DBD. In vivo studies with TR showed that although N-CoR fully dissociates in the presence of ligand, the levels of TR bound PSF and Sin3A appear to remain unchanged, indicating that Sin3A can be recruited to the receptor independent of N-CoR or SMRT. RXR was not detected to bind N-CoR although it bound PSF and Sin3A as effectively as TR, and this association with RXR did not change with ligand. Our studies point to a novel PSF/Sin3-mediated pathway for nuclear hormone receptors, and possibly other transcription factors, which may fine-tune the transcriptional response as well as play an important role in mediating the repressive effects of those type II receptors which only weakly interact with N-CoR and SMRT. PMID- 11259581 TI - Drosophila Mediator complex is broadly utilized by diverse gene-specific transcription factors at different types of core promoters. AB - To decipher the mechanistic roles of Mediator proteins in regulating developmental specific gene expression and compare them to those of TATA-binding protein (TBP)-associated factors (TAFs), we isolated and analyzed a multiprotein complex containing Drosophila Mediator (dMediator) homologs. dMediator interacts with several sequence-specific transcription factors and basal transcription machinery and is critical for activated transcription in response to diverse transcriptional activators. The requirement for dMediator did not depend on a specific core promoter organization. By contrast, TAFs are preferentially utilized by promoters having a specific core element organization. Therefore, Mediator proteins are suggested to act as a pivotal coactivator that integrates promoter-specific activation signals to the basal transcription machinery. PMID- 11259582 TI - Apoptosis suppression by Raf-1 and MEK1 requires MEK- and phosphatidylinositol 3 kinase-dependent signals. AB - Two Ras effector pathways leading to the activation of Raf-1 and phosphatidylinositol 3-kinase (PI3K) have been implicated in the survival signaling by the interleukin 3 (IL-3) receptor. Analysis of apoptosis suppression by Raf-1 demonstrated the requirement for mitochondrial translocation of the kinase in this process. This could be achieved either by overexpression of the antiapoptotic protein Bcl-2 or by targeting Raf-1 to the mitochondria via fusion to the mitochondrial protein Mas p70. Mitochondrially active Raf-1 is unable to activate extracellular signal-related kinase 1 (ERK1) and ERK2 but suppresses cell death by inactivating the proapoptotic Bcl-2 family member BAD. However, genetic and biochemical data also have suggested a role for the Raf-1 effector module MEK-ERK in apoptosis suppression. We thus tested for MEK requirement in cell survival signaling using the interleukin 3 (IL-3)-dependent cell line 32D. MEK is essential for survival and growth in the presence of IL-3. Upon growth factor withdrawal the expression of constitutively active MEK1 mutants significantly delays the onset of apoptosis, whereas the presence of a dominant negative mutant accelerates cell death. Survival signaling by MEK most likely results from the activation of ERKs since expression of a constitutively active form of ERK2 was as effective in protecting NIH 3T3 fibroblasts against doxorubicin-induced cell death as oncogenic MEK. The survival effect of activated MEK in 32D cells is achieved by both MEK- and PI3K-dependent mechanisms and results in the activation of PI3K and in the phosphorylation of AKT. MEK and PI3K dependence is also observed in 32D cells protected from apoptosis by oncogenic Raf-1. Additionally, we also could extend these findings to the IL-3-dependent pro-B-cell line BaF3, suggesting that recruitment of MEK is a common mechanism for survival signaling by activated Raf. Requirement for the PI3K effector AKT in this process is further demonstrated by the inhibitory effect of a dominant negative AKT mutant on Raf-1-induced cell survival. Moreover, a constitutively active form of AKT synergizes with Raf-1 in apoptosis suppression. In summary these data strongly suggest a Raf effector pathway for cell survival that is mediated by MEK and AKT. PMID- 11259583 TI - Protein import channel of the outer mitochondrial membrane: a highly stable Tom40 Tom22 core structure differentially interacts with preproteins, small tom proteins, and import receptors. AB - The preprotein translocase of the yeast mitochondrial outer membrane (TOM) consists of the initial import receptors Tom70 and Tom20 and a approximately 400 kDa (400 K) general import pore (GIP) complex that includes the central receptor Tom22, the channel Tom40, and the three small Tom proteins Tom7, Tom6, and Tom5. We report that the GIP complex is a highly stable complex with an unusual resistance to urea and alkaline pH. Under mild conditions for mitochondrial lysis, the receptor Tom20, but not Tom70, is quantitatively associated with the GIP complex, forming a 500K to 600K TOM complex. A preprotein, stably arrested in the GIP complex, is released by urea but not high salt, indicating that ionic interactions are not essential for keeping the preprotein in the GIP complex. Under more stringent detergent conditions, however, Tom20 and all three small Tom proteins are released, while the preprotein remains in the GIP complex. Moreover, purified outer membrane vesicles devoid of translocase components of the intermembrane space and inner membrane efficiently accumulate the preprotein in the GIP complex. Together, Tom40 and Tom22 thus represent the functional core unit that stably holds accumulated preproteins. The GIP complex isolated from outer membranes exhibits characteristic TOM channel activity with two coupled conductance states, each corresponding to the activity of purified Tom40, suggesting that the complex contains two simultaneously active and coupled channel pores. PMID- 11259584 TI - Novel function of Rad27 (FEN-1) in restricting short-sequence recombination. AB - Saccharomyces cerevisiae mutants lacking the structure-specific nuclease Rad27 display an enhancement in recombination that increases as sequence length decreases, suggesting that Rad27 preferentially restricts recombination between short sequences. Since wild-type alleles of both RAD27 and its human homologue FEN1 complement the elevated short-sequence recombination (SSR) phenotype of a rad27-null mutant, this function may be conserved from yeast to humans. Furthermore, mutant Rad27 and FEN-1 enzymes with partial flap endonuclease activity but without nick-specific exonuclease activity partially complement the SSR phenotype of the rad27-null mutant. This suggests that the endonuclease activity of Rad27 (FEN-1) plays a role in limiting recombination between short sequences in eukaryotic cells. PMID- 11259585 TI - Mitochondrial translation of Saccharomyces cerevisiae COX2 mRNA is controlled by the nucleotide sequence specifying the pre-Cox2p leader peptide. AB - The mitochondrial gene encoding yeast cytochrome oxidase subunit II (Cox2p) specifies a precursor protein with a 15-amino-acid leader peptide. Deletion of the entire leader peptide coding region is known to block Cox2p accumulation posttranscriptionally. Here, we examined in vivo the role of the pre-Cox2p leader peptide and the mRNA sequence that encodes it in the expression of a mitochondrial reporter gene, ARG8m, fused to the 91st codon of COX2. We found within the coding sequence antagonistic elements that control translation: the positive element includes sequences in the first 14 codons specifying the leader peptide, while the negative element appears to be within codons 15 to 91. Partial deletions, point mutations, and local frameshifts within the leader peptide coding region were placed in both the cox2::ARG8m reporter and in COX2 itself. Surprisingly, the mRNA sequence of the first six codons specifying the leader peptide plays an important role in positively controlling translation, while the amino acid sequence of the leader peptide itself is relatively unconstrained. Two mutations that partially block translation can be suppressed by nearby sequence substitutions that weaken a predicted stem structure and by overproduction of either the COX2 mRNA-specific translational activator Pet111p or the large subunit mitochondrial ribosomal protein MrpL36p. We propose that regulatory elements embedded in the translated COX2 mRNA sequence could play a role, together with trans-acting factors, in coupling regulated synthesis of nascent pre-Cox2p to its insertion in the mitochondrial inner membrane. PMID- 11259586 TI - p38 mitogen-activated protein kinase-dependent activation of protein phosphatases 1 and 2A inhibits MEK1 and MEK2 activity and collagenase 1 (MMP-1) gene expression. AB - Degradation of collagenous extracellular matrix by collagenase 1 (also known as matrix metalloproteinase 1 [MMP-1]) plays a role in the pathogenesis of various destructive disorders, such as rheumatoid arthritis, chronic ulcers, and tumor invasion and metastasis. Here, we have investigated the role of distinct mitogen activated protein kinase (MAPK) pathways in the regulation of MMP-1 gene expression. The activation of the extracellular signal-regulated kinase 1 (ERK1)/ERK2 (designated ERK1,2) pathway by oncogenic Ras, constitutively active Raf-1, or phorbol ester resulted in potent stimulation of MMP-1 promoter activity and mRNA expression. In contrast, activation of stress-activated c-Jun N-terminal kinase and p38 pathways by expression of constitutively active mutants of Rac, transforming growth factor beta-activated kinase 1 (TAK1), MAPK kinase 3 (MKK3), or MKK6 or by treatment with arsenite or anisomycin did not alone markedly enhance MMP-1 promoter activity. Constitutively active MKK6 augmented Raf-1 mediated activation of the MMP-1 promoter, whereas active mutants of TAK1 and MKK3b potently inhibited the stimulatory effect of Raf-1. Activation of p38 MAPK by arsenite also potently abrogated stimulation of MMP-1 gene expression by constitutively active Ras and Raf-1 and by phorbol ester. Specific activation of p38alpha by adenovirus-delivered constitutively active MKK3b resulted in potent inhibition of the activity of ERK1,2 and its upstream activator MEK1,2. Furthermore, arsenite prevented phorbol ester-induced phosphorylation of ERK1,2 kinase-MEK1,2, and this effect was dependent on p38-mediated activation of protein phosphatase 1 (PP1) and PP2A. These results provide evidence that activation of signaling cascade MKK3-MKK3b-->p38alpha blocks the ERK1,2 pathway at the level of MEK1,2 via PP1-PP2A and inhibits the activation of MMP-1 gene expression. PMID- 11259587 TI - Establishment and maintenance of DNA methylation patterns in mouse Ndn: implications for maintenance of imprinting in target genes of the imprinting center. AB - Ndn is located on chromosome 7C, an imprinted region of the mouse genome. Imprinting of Ndn and adjacent paternally expressed genes is regulated by a regional imprinting control element known as the imprinting center (IC). An IC also controls imprint resetting of target genes in the region of conserved synteny on human chromosome 15q11-q13, which is deleted or rearranged in the neurodevelopmental disorder Prader-Willi syndrome. Epigenetic modifications such as DNA methylation, which occur in gametes and can be stably propagated, are presumed to establish and maintain the imprint in target genes of the IC. While most DNA becomes substantially demethylated by the blastocyst stage, some imprinted genes have regions that escape global demethylation and may maintain the imprint. We have now analyzed the methylation of 39 CpG dinucleotide sequences in the 5' end of Ndn by sodium bisulfite sequencing in gametes and in preimplantation and adult tissues. While sperm DNA is completely unmethylated across this region, oocyte DNA is partially methylated. A distinctive but unstable maternal methylation pattern persists until the morula stage and is lost in the blastocyst stage, where low levels of methylation are present on most DNA strands of either parental origin. The methylation pattern is then substantially remodeled, and fewer than half of maternally derived DNA strands in adult brain resemble the oocyte pattern. We postulate that for Ndn, DNA methylation may initially preserve a gametic imprint during preimplantation development, but other epigenetic events may maintain the imprint later in embryonic development. PMID- 11259588 TI - Protein tyrosine phosphatase CD148-mediated inhibition of T-cell receptor signal transduction is associated with reduced LAT and phospholipase Cgamma1 phosphorylation. AB - In this study, we investigate the role of the receptor-like protein tyrosine phosphatase CD148 in T-cell activation. Overexpression of CD148 in the Jurkat T cell line inhibited activation of the transcription factor nuclear factor of activated T cells following T-cell receptor (TCR) stimulation but not following stimulation through a heterologously expressed G protein-coupled receptor, the human muscarinic receptor subtype 1. Using a tetracycline-inducible expression system, we show that the TCR-mediated activation of both the Ras and calcium pathways was inhibited by expression of CD148 at levels that approximate those found in activated primary T cells. These effects were dependent on the phosphatase activity of CD148. Analysis of TCR-induced protein tyrosine phosphorylation demonstrated that most phosphoproteins were unaffected by CD148 expression. However, phospholipase Cgamma1 (PLCgamma1) and LAT were strikingly hypophosphorylated in CD148-expressing cells following TCR stimulation, whereas the phosphorylation levels of Slp-76 and Itk were modestly reduced. Based on these results, we propose that CD148 negatively regulates TCR signaling by interfering with the phosphorylation and function of PLCgamma1 and LAT. PMID- 11259589 TI - Molecular distinction between specification and differentiation in the myogenic basic helix-loop-helix transcription factor family. AB - The myogenic basic helix-loop-helix (bHLH) proteins regulate both skeletal muscle specification and differentiation: MyoD and Myf5 establish the muscle lineage, whereas myogenin mediates differentiation. Previously, we demonstrated that MyoD was more efficient than myogenin at initiating the expression of skeletal muscle genes, and in this study we present the molecular basis for this difference. A conserved amphipathic alpha-helix in the carboxy terminus of the myogenic bHLH proteins has distinct activities in MyoD and myogenin: the MyoD helix facilitates the initiation of endogenous gene expression, whereas the myogenin helix functions as a general transcriptional activation domain. Thus, the alternate use of a similar motif for gene initiation and activation provides a molecular basis for the distinction between specification and differentiation within the myogenic bHLH gene family. PMID- 11259590 TI - Site-specific acetylation by p300 or CREB binding protein regulates erythroid Kruppel-like factor transcriptional activity via its interaction with the SWI-SNF complex. AB - Recruitment of modifiers and remodelers to specific DNA sites within chromatin plays a critical role in controlling gene expression. The study of globin gene regulation provides a convergence point within which to address these issues in the context of tissue-specific and developmentally regulated expression. In this regard, erythroid Kruppel-like factor (EKLF) is critical. EKLF is a red cell specific activator whose presence is crucial for establishment of the correct chromatin structure and high-level transcriptional induction of adult beta globin. We now find, by metabolic labeling-immunoprecipitation experiments, that EKLF is acetylated in the erythroid cell. EKLF residues acetylated by CREB binding protein (CBP) in vitro map to Lys-288 in its transactivation domain and Lys-302 in its zinc finger domain. Although site-specific DNA binding by EKLF is unaffected by the acetylation status of either of these lysines, directed mutagenesis of Lys-288 (but not Lys-302) decreases the ability of EKLF to transactivate the beta-globin promoter in vivo and renders it unable to be superactivated by coexpressed p300 or CBP. In addition, the acetyltransferase function of CBP or p300 is required for superactivation of wild-type EKLF. Finally, acetylated EKLF has a higher affinity for the SWI-SNF chromatin remodeling complex and is a more potent transcriptional activator of chromatin assembled templates in vitro. These results demonstrate that the acetylation status of EKLF is critical for its optimal activity and suggest a mechanism by which EKLF acts as an integrator of remodeling and transcriptional components to alter chromatin structure and induce adult beta-globin expression within the beta like globin cluster. PMID- 11259591 TI - S338 phosphorylation of Raf-1 is independent of phosphatidylinositol 3-kinase and Pak3. AB - The Raf-1 serine/threonine protein kinase requires phosphorylation of the serine at position 338 (S338) for activation. Ras is required to recruit Raf-1 to the plasma membrane, which is where S338 phosphorylation occurs. The recent suggestion that Pak3 could stimulate Raf-1 activity by directly phosphorylating S338 through a Ras/phosphatidylinositol 3-kinase (Pl3-K)/-Cdc42-dependent pathway has attracted much attention. Using a phospho-specific antibody to S338, we have reexamined this model. Using LY294002 and wortmannin, inhibitors of Pl3-K, we find that growth factor-mediated S338 phosphorylation still occurs, even when Pl3 K activity is completely blocked. Although high concentrations of LY294002 and wortmannin did suppress S338 phosphorylation, they also suppressed Ras activation. Additionally, we show that Pak3 is not activated under conditions where S338 is phosphorylated, but when Pak3 is strongly activated, by coexpression with V12Cdc42 or by mutations that make it independent of Cdc42, it did stimulate S338 phosphorylation. However, this occurred in the cytosol and did not stimulate Raf-1 kinase activity. The inability of Pak3 to activate Raf-1 was not due to an inability to stimulate phosphorylation of the tyrosine at position 341 but may be due to its inability to recruit Raf-1 to the plasma membrane. Taken together, our data show that growth factor-stimulated Raf-1 activity is independent of Pl3-K activity and argue against Pak3 being a physiological mediator of S338 phosphorylation in growth factor-stimulated cells. PMID- 11259592 TI - Heterozygous disruption of the TATA-binding protein gene in DT40 cells causes reduced cdc25B phosphatase expression and delayed mitosis. AB - TATA-binding protein (TBP) is a key general transcription factor required for transcription by all three nuclear RNA polymerases. Although it has been intensively analyzed in vitro and in Saccharomyces cerevisiae, in vivo studies of vertebrate TBP have been limited. We applied gene-targeting techniques using chicken DT40 cells to generate heterozygous cells with one copy of the TBP gene disrupted. Such TBP-heterozygous (TBP-Het) cells showed unexpected phenotypic abnormalities, resembling those of cells with delayed mitosis: a significantly lower growth rate, larger size, more G2/-M- than G1-phase cells, and a high proportion of sub-G1, presumably apoptotic, cells. Further evidence for delayed mitosis in TBP-Het cells was provided by the differential effects of several cell cycle-arresting drugs. To determine the cause of these defects, we first examined the status of cdc2 kinase, which regulates the G2/M transition, and unexpectedly observed more hyperphosphorylated, inactive cdc2 in TBP-Het cells. Providing an explanation for this, mRNA and protein levels of cdc25B, the trigger cdc2 phosphatase, were significantly and specifically reduced. These properties were all due to decreased TBP levels, as they could be rescued by expression of exogeneous TBP, including, in most but not all cases, a mutant form lacking the species-specific N-terminal domain. Our results indicate that small changes in TBP concentration can have profound effects on cell growth in vertebrate cells. PMID- 11259593 TI - The Cbk1p pathway is important for polarized cell growth and cell separation in Saccharomyces cerevisiae. AB - During the early stages of budding, cell wall remodeling and polarized secretion are concentrated at the bud tip (apical growth). The CBK1 gene, encoding a putative serine/threonine protein kinase, was identified in a screen designed to isolate mutations that affect apical growth. Analysis of cbk1Delta cells reveals that Cbk1p is required for efficient apical growth, proper mating projection morphology, bipolar bud site selection in diploid cells, and cell separation. Epitope-tagged Cbk1p localizes to both sides of the bud neck in late anaphase, just prior to cell separation. CBK1 and another gene, HYM1, were previously identified in a screen for genes involved in transcriptional repression and proposed to function in the same pathway. Deletion of HYM1 causes phenotypes similar to those observed in cbk1Delta cells and disrupts the bud neck localization of Cbk1p. Whole-genome transcriptional analysis of cbk1Delta suggests that the kinase regulates the expression of a number of genes with cell wall-related functions, including two genes required for efficient cell separation: the chitinase-encoding gene CTS1 and the glucanase-encoding gene SCW11. The Ace2p transcription factor is required for expression of CTS1 and has been shown to physically interact with Cbk1p. Analysis of ace2Delta cells reveals that Ace2p is required for cell separation but not for polarized growth. Our results suggest that Cbk1p and Hym1p function to regulate two distinct cell morphogenesis pathways: an ACE2-independent pathway that is required for efficient apical growth and mating projection formation and an ACE2-dependent pathway that is required for efficient cell separation following cytokinesis. Cbk1p is most closely related to the Neurospora crassa Cot-1; Schizosaccharomyces pombe Orb6; Caenorhabditis elegans, Drosophila, and human Ndr; and Drosophila and mammalian WARTS/LATS kinases. Many Cbk1-related kinases have been shown to regulate cellular morphology. PMID- 11259594 TI - Functional analysis of mouse C-terminal kinesin motor KifC2. AB - Proteins of the kinesin superfamily define a class of microtubule-dependent motors that play crucial roles in cell division and intracellular transport. In the mouse, several kinesin motors have been characterized and are suggested to play roles in axonal and/or dendritic transport. One such kinesin is KifC2. Sequence and secondary structure analysis revealed that KifC2 is a member of the C-terminal motor family. Northern and Western blot analyses indicated that KifC2 is specifically expressed in both the central and peripheral nervous systems. The cellular locations of the KifC2 proteins were found to be mainly in neural cell bodies and dendrites but also in axons. To understand the in vivo function of the KifC2 gene, we used homologous recombination in embryonic stem cells to construct knockout mouse strains for the KifC2 gene. Homozygous KifC2 mutants were viable and reproduced normally, and their development was apparently normal. These results suggest that KifC2 is dispensable for normal neural development and behavior in the mouse. PMID- 11259595 TI - Are all DNA binding and transcription regulation by an activator physiologically relevant? AB - Understanding how a regulatory protein occupies its sites in vivo is central to understanding gene regulation. Using the yeast Gal4 protein as a model for such studies, we have found 239 potential Gal4 binding sites in the yeast genome, 186 of which are in open reading frames (ORFs). This raises the questions of whether these sites are occupied by Gal4 and, if so, to what effect. We have analyzed the Saccharomyces cerevisiae ACC1 gene (encoding acetyl-coenzyme A carboxylase), which has three Gal4 binding sites in its ORF. The plasmid titration assay has demonstrated that Gal4 occupies these sites in the context of an active ACC1 gene. We also find that the expression of the ACC1 is reduced fourfold in galactose medium and that this reduction is dependent on the Gal4 binding sites, suggesting that Gal4 bound to the ORF sites affects transcription of ACC1. Interestingly, removal of the Gal4 binding sites has no obvious effect on the growth in galactose under laboratory conditions. In addition, though the sequence of the ACC1 gene is highly conserved among yeast species, these Gal4 binding sites are not present in the Kluyveromyces lactis ACC1 gene. We suggest that the occurrence of these sites may not be related to galactose regulation and a manifestation of the "noise" in the occurrence of Gal4 binding sites. PMID- 11259596 TI - Interleukin-1 (IL-1) receptor-associated kinase leads to activation of TAK1 by inducing TAB2 translocation in the IL-1 signaling pathway. AB - Interleukin-1 (IL-1) is a proinflammatory cytokine that recognizes a surface receptor complex and generates multiple cellular responses. IL-1 stimulation activates the mitogen-activated protein kinase kinase kinase TAK1, which in turn mediates activation of c-Jun N-terminal kinase and NF-kappaB. TAB2 has previously been shown to interact with both TAK1 and TRAF6 and promote their association, thereby triggering subsequent IL-1 signaling events. The serine/threonine kinase IL-1 receptor-associated kinase (IRAK) also plays a role in IL-1 signaling, being recruited to the IL-1 receptor complex early in the signal cascade. In this report, we investigate the role of IRAK in the activation of TAK1. Genetic analysis reveals that IRAK is required for IL-1-induced activation of TAK1. We show that IL-1 stimulation induces the rapid but transient association of IRAK, TRAF6, TAB2, and TAK1. TAB2 is recruited to this complex following translocation from the membrane to the cytosol upon IL-1 stimulation. In IRAK-deficient cells, TAB2 translocation and its association with TRAF6 are abolished. These results suggest that IRAK regulates the redistribution of TAB2 upon IL-1 stimulation and facilitates the formation of a TRAF6-TAB2-TAK1 complex. Formation of this complex is an essential step in the activation of TAK1 in the IL-1 signaling pathway. PMID- 11259597 TI - Critical role of the HMGI(Y) proteins in adipocytic cell growth and differentiation. AB - The high-mobility group I (HMGI) nonhistone chromosomal proteins HMGI(Y) and HMGI C have been implicated in defining chromatin structure and in regulating the transcription of several genes. These proteins have been implicated in adipocyte homeostasis: a severe deficiency of fat tissue is found in mice with targeted disruption of the HMGI-C locus, and lipomagenesis in humans is frequently associated with somatic mutations of HMGI genes. The aim of this study was to examine the role of HMGI(Y) proteins in adipocytic cell growth and differentiation. First, we found that differentiation of the preadipocytic 3T3-L1 cell line caused early induction of HMGI(Y) gene expression. Suppression of HMGI(Y) expression by antisense technology dramatically increased the growth rate and impaired adipocytic differentiation in these cells. The process of adipogenic differentiation involves the interplay of several transcription factors, among which is the CCAAT/enhancer-binding protein (C/EBP) family of proteins. These factors are required for the transcriptional activation of adipocyte-specific genes. We also tested the hypothesis that HMGI(Y) might participate in transcriptional control of adipocyte-specific promoters. We found that HMGI(Y) proteins bind C/EBPbeta in vivo and in vitro. Furthermore, we show that HMGI(Y) strongly potentiates the capacity of C/EBPbeta to transactivate the leptin promoter, an adipose-specific promoter. Taken together, these results indicate that the HMGI(Y) proteins play a critical role in adipocytic cell growth and differentiation. PMID- 11259598 TI - Rfg1, a protein related to the Saccharomyces cerevisiae hypoxic regulator Rox1, controls filamentous growth and virulence in Candida albicans. AB - Candida albicans, the major fungal pathogen in humans, can undergo a reversible transition from ellipsoidal single cells (blastospores) to filaments composed of elongated cells attached end to end. This transition is thought to allow for rapid colonization of host tissues, facilitating the spread of infection. Here, we report the identification of Rfg1, a transcriptional regulator that controls filamentous growth of C. albicans in an environment-dependent manner. Rfg1 is important for virulence of C. albicans in a mouse model and is shown to control a number of genes that have been implicated in this process. The closest relative to Rfg1 in Saccharomyces cerevisiae is Rox1, a key repressor of hypoxic genes. However, Rfg1 does not appear to play a role in the regulation of hypoxic genes in C. albicans. These results demonstrate that a regulatory protein that controls the hypoxic response in S. cerevisiae controls filamentous growth and virulence in C. albicans. The observations described in this paper raise new and intriguing questions about the evolutionary relationship between these processes. PMID- 11259599 TI - F-box protein Grr1 interacts with phosphorylated targets via the cationic surface of its leucine-rich repeat. AB - The flexibility and specificity of ubiquitin-dependent proteolysis are mediated, in part, by the E3 ubiquitin ligases. One class of E3 enzymes, SKp1/cullin/F-box protein (SCF), derives its specificity from F-box proteins, a heterogeneous family of adapters for target protein recognition. Grr1, the F-box component of SCF(Grr1), mediates the interaction with phosphorylated forms of the G(1) cyclins Cln1 and Cln2. We show that binding of Cln2 by SCF(Grr1) was dependent upon its leucine-rich repeat (LRR) domain and its carboxy terminus. Our structural model for the Grr1 LRR predicted a high density of positive charge on the concave surface of the characteristic horseshoe structure. We hypothesized that specific basic residues on the predicted concave surface are important for recognition of phosphorylated Cln2. We show that point mutations that converted the basic residues on the concave surface but not those on the convex surface to neutral or acidic residues interfered with the capacity of Grr1 to bind to Cln2. The same mutations resulted in the stabilization of Cln2 and Gic2 and also in a spectrum of phenotypes characteristic of inactivation of GRR1, including hyperpolarization and enhancement of pseudohyphal growth. It was surprising that the same residues were not important for the role of Grr1 in nutrient-regulated transcription of HXT1 or AGP1. We concluded that the cationic nature of the concave surface of the Grr1 LRR is critical for the recognition of phosphorylated targets of SCF(Grr1) but that other properties of Grr1 are required for its other functions. PMID- 11259600 TI - Essential role of insulin receptor substrate 1 (IRS-1) and IRS-2 in adipocyte differentiation. AB - To investigate the role of insulin receptor substrate 1 (IRS-1) and IRS-2, the two ubiquitously expressed IRS proteins, in adipocyte differentiation, we established embryonic fibroblast cells with four different genotypes, i.e., wild type, IRS-1 deficient (IRS-1(-/-)), IRS-2 deficient (IRS-2(-/-)), and IRS-1 IRS-2 double deficient (IRS-1(-/-) IRS-2(-/-)), from mouse embryos of the corresponding genotypes. The abilities of IRS-1(-/-) cells and IRS-2(-/-) cells to differentiate into adipocytes are approximately 60 and 15%, respectively, lower than that of wild-type cells, at day 8 after induction and, surprisingly, IRS-1( /-) IRS-2(-/-) cells have no ability to differentiate into adipocytes. The expression of CCAAT/enhancer binding protein alpha (C/EBPalpha) and peroxisome proliferator-activated receptor gamma (PPARgamma) is severely decreased in IRS-1( /-) IRS-2(-/-) cells at both the mRNA and the protein level, and the mRNAs of lipoprotein lipase and adipocyte fatty acid binding protein are severely decreased in IRS-1(-/-) IRS-2(-/-) cells. Phosphatidylinositol 3-kinase (PI 3 kinase) activity that increases during adipocyte differentiation is almost completely abolished in IRS-1(-/-) IRS-2(-/-) cells. Treatment of wild-type cells with a PI 3-kinase inhibitor, LY294002, markedly decreases the expression of C/EBPalpha and PPARgamma, a result which is associated with a complete block of adipocyte differentiation. Moreover, histologic analysis of IRS-1(-/-) IRS-2(-/-) double-knockout mice 8 h after birth reveals severe reduction in white adipose tissue mass. Our results suggest that IRS-1 and IRS-2 play a crucial role in the upregulation of the C/EBPalpha and PPARgamma expression and adipocyte differentiation. PMID- 11259601 TI - Activation of hypodermal differentiation in the Caenorhabditis elegans embryo by GATA transcription factors ELT-1 and ELT-3. AB - The Caenorhabditis elegans GATA transcription factor genes elt-1 and elt-3 are expressed in the embryonic hypodermis (also called the epidermis). elt-1 is expressed in precursor cells and is essential for the production of most hypodermal cells (22). elt-3 is expressed in all of the major hypodermal cells except the lateral seam cells, and expression is initiated immediately after the terminal division of precursor lineages (13). Although this expression pattern suggests a role for ELT-3 in hypodermal development, no functional studies have yet been performed. In the present paper, we show that either elt-3 or elt-1 is sufficient, when force expressed in early embryonic blastomeres, to activate a program of hypodermal differentiation even in blastomeres that are not hypodermal precursors in wild-type embryos. We have deleted the elt-3 gene and shown that ELT-3 is not essential for either hypodermal cell differentiation or the viability of the organism. We showed that ELT-3 can activate hypodermal gene expression in the absence of ELT-1 and that, conversely, ELT-1 can activate hypodermal gene expression in the absence of ELT-3. Overall, the combined results of the mutant phenotypes, initial expression times, and our forced-expression experiments suggest that ELT-3 acts downstream of ELT-1 in a redundant pathway controlling hypodermal cell differentiation. PMID- 11259603 TI - The hinge and chromo shadow domain impart distinct targeting of HP1-like proteins. AB - Drosophila heterochromatin-associated protein 1 (HP1) is an abundant component of heterochromatin, a highly condensed compartment of the nucleus that comprises a major fraction of complex genomes. Some organisms have been shown to harbor multiple HP1-like proteins, each exhibiting spatially distinct localization patterns within interphase nuclei. We have characterized the subnuclear localization patterns of two newly discovered Drosophila HP1-like proteins (HP1b and HP1c), comparing them with that of the originally described fly HP1 protein (here designated HP1a). While HP1a targets heterochromatin, HP1b localizes to both heterochromatin and euchromatin and HP1c is restricted exclusively to euchromatin. All HP1-like proteins contain an amino-terminal chromo domain, a connecting hinge, and a carboxyl-terminal chromo shadow domain. We expressed truncated and chimeric HP1 proteins in vivo to determine which of these segments might be responsible for heterochromatin-specific and euchromatin-specific localization. Both the HP1a hinge and chromo shadow domain independently target heterochromatin, while the HP1c chromo shadow domain is implicated solely in euchromatin localization. Comparative sequence analyses of HP1 homologs reveal a conserved sequence block within the hinge that contains an invariant sequence (KRK) and a nuclear localization motif. This block is not conserved in the HP1c hinge, possibly accounting for its failure to function as an independent targeting segment. We conclude that sequence variations within the hinge and shadow account for HP1 targeting distinctions. We propose that these targeting features allow different HP1 complexes to be distinctly sequestered in organisms that harbor multiple HP1-like proteins. PMID- 11259602 TI - NXT1 (p15) is a crucial cellular cofactor in TAP-dependent export of intron containing RNA in mammalian cells. AB - TAP, the human homologue of the yeast protein Mex67p, has been proposed to serve a role in mRNA export in mammalian cells. We have examined the ability of TAP to mediate export of Rev response element (RRE)-containing human immunodeficiency virus (HIV) RNA, a well-characterized export substrate in mammalian cells. To do this, the TAP gene was fused in frame to either RevM10 or RevDelta78-79. These proteins are nonfunctional Rev mutant proteins that can bind to HIV RNA containing the RRE in vivo but are unable to mediate the export of this RNA to the cytoplasm. However, the fusion of TAP to either of these mutant proteins gave rise to chimeric proteins that were able to complement Rev function. Significantly, cotransfection with a vector expressing NXT1 (p15), an NTF2 related cellular factor that binds to TAP, led to dramatic enhancement of the ability of the chimeric proteins to mediate RNA export. Mutant-protein analysis demonstrated that the domain necessary for nuclear export mapped to the C terminal region of TAP and required the domain that interacts with NXT1, as well as the region that has been shown to interact with nucleoporins. RevM10-TAP function was leptomycin B insensitive. In contrast, the function of this protein was inhibited by DeltaCAN, a protein consisting of part of the FG repeat domain of CAN/Nup214. These results show that TAP can complement Rev nuclear export signal function and redirect the export of intron-containing RNA to a CRM1 independent pathway. These experiments support the role of TAP as an RNA export factor in mammalian cells. In addition, they indicate that NXT1 serves as a crucial cellular cofactor in this process. PMID- 11259604 TI - Epidermal growth factor-like repeats mediate lateral and reciprocal interactions of Ep-CAM molecules in homophilic adhesions. AB - Ep-CAM is a new type of cell adhesion molecule (CAM) which does not structurally resemble the members of the four major families (cadherins, integrins, selectins, and CAMs of the immunoglobulin superfamily) and mediates Ca(2+)-independent, homophilic adhesions. The extracellular domain of Ep-CAM consists of a cysteine rich region, containing two type II epidermal growth factor (EGF)-like repeats, followed by a cysteine-poor region. We generated mutated Ep-CAM forms with various deletions in the extracellular domain. These deletion mutants, together with monoclonal antibodies recognizing different epitopes in the extracellular domain, were used to investigate the role of the EGF-like repeats in the formation of intercellular contacts mediated by Ep-CAM molecules. We established that both EGF-like repeats are required for the formation of Ep-CAM-mediated homophilic adhesions, including the accumulation of Ep-CAM molecules at the cell cell boundaries, and the anchorage of the Ep-CAM adhesion complex to F-actin via alpha-actinin. Deletion of either EGF-like repeat was sufficient to inhibit the adhesion properties of the molecule. The first EGF-like repeat of Ep-CAM is required for reciprocal interactions between Ep-CAM molecules on adjacent cells, as was demonstrated with blocking antibodies. The second EGF-like repeat was mainly required for lateral interactions between Ep-CAM molecules. Lateral interactions between Ep-CAM molecules result in the formation of tetramers, which might be the first and necessary step in the formation of Ep-CAM-mediated intercellular contacts. PMID- 11259605 TI - Two immunologically distinct human DNA polymerase alpha-primase subpopulations are involved in cellular DNA replication. AB - Metabolic labeling of primate cells revealed the existence of phosphorylated and hypophosphorylated DNA polymerase alpha-primase (Pol-Prim) populations that are distinguishable by monoclonal antibodies. Cell cycle studies showed that the hypophosphorylated form was found in a complex with PP2A and cyclin E-Cdk2 in G1, whereas the phosphorylated enzyme was associated with a cyclin A kinase in S and G2. Modification of Pol-Prim by PP2A and Cdks regulated the interaction with the simian virus 40 origin-binding protein large T antigen and thus initiation of DNA replication. Confocal microscopy demonstrated nuclear colocalization of hypophosphorylated Pol-Prim with MCM2 in S phase nuclei, but its presence preceded 5-bromo-2'-deoxyuridine (BrdU) incorporation. The phosphorylated replicase exclusively colocalized with the BrdU signal, but not with MCM2. Immunoprecipitation experiments proved that only hypophosphorylated Pol-Prim associated with MCM2. The data indicate that the hypophosphorylated enzyme initiates DNA replication at origins, and the phosphorylated form synthesizes the primers for the lagging strand of the replication fork. PMID- 11259606 TI - The hsp90 chaperone complex regulates intracellular localization of the dioxin receptor. AB - The molecular chaperone complex hsp90-p23 interacts with the dioxin receptor, a ligand-dependent basic helix-loop-helix (bHLH)/Per-Arnt-Sim domain transcription factor. Whereas biochemical and genetic evidence indicates that hsp90 is important for maintenance of a high-affinity ligand binding conformation of the dioxin receptor, the role of hsp90-associated proteins in regulation of the dioxin receptor function remains unclear. Here we demonstrate that the integrity of the hsp90 complex characterized by the presence of the hsp90-associated cochaperone p23 and additional cochaperone proteins is important for regulation of the intracellular localization of the dioxin receptor by two mechanisms. First, in the absence of ligand, the dioxin receptor-hsp90 complex was associated with the immunophilin-like protein XAP2 to mediate cytoplasmic retention of the dioxin receptor. Second, upon exposure to ligand, the p23-associated hsp90 complex mediated interaction of the dioxin receptor with the nuclear import receptor protein pendulin and subsequent nuclear translocation of the receptor. Interestingly, these two modes of regulation target two distinct functional domains of the dioxin receptor. Whereas the nuclear localization signal containing and hsp90-interacting bHLH domain of the receptor regulates ligand dependent nuclear import, the interaction of the p23-hsp90-XAP2 complex with the ligand binding domain of the dioxin receptor was essential to mediate cytoplasmic retention of the ligand-free receptor form. In conclusion, these data suggest a novel role of the hsp90 molecular chaperone complex in regulation of the intracellular localization of the dioxin receptor. PMID- 11259607 TI - A protease-resistant 61-residue prion peptide causes neurodegeneration in transgenic mice. AB - An abridged prion protein (PrP) molecule of 106 amino acids, designated PrP106, is capable of forming infectious miniprions in transgenic mice (S. Supattapone, P. Bosque, T. Muramoto, H. Wille, C. Aagaard, D. Peretz, H.-O. B. Nguyen, C. Heinrich, M. Torchia, J. Safar, F. E. Cohen, S. J. DeArmond, S. B. Prusiner, and M. Scott, Cell 96:869-878, 1999). We removed additional sequences from PrP106 and identified a 61-residue peptide, designated PrP61, that spontaneously adopted a protease-resistant conformation in neuroblastoma cells. Synthetic PrP61 bearing a carboxy-terminal lipid moiety polymerized into protease-resistant, beta-sheet enriched amyloid fibrils at a physiological salt concentration. Transgenic mice expressing low levels of PrP61 died spontaneously with ataxia. Neuropathological examination revealed accumulation of protease-resistant PrP61 within neuronal dendrites and cell bodies, apparently causing apoptosis. PrP61 may be a useful model for deciphering the mechanism by which PrP molecules acquire protease resistance and become neurotoxic. PMID- 11259608 TI - Is cisplatin-induced cell death always produced by apoptosis? AB - It is generally accepted that DNA damage and subsequent induction of apoptosis may be the primary cytotoxic mechanism of cisplatin and other DNA-binding antitumor drugs (Fisher,1994). Because the final step of apoptosis is characterized by morphological changes in the nucleus, the death signals of the execution phase must be transmitted from the cytoplasm to the nucleus. Thus, the recognition and processing of cisplatin-induced DNA damage through"classic" apoptosis, requires that a nuclear signal, generated at the initiation phase, be transmitted to the cytoplasm to be processed through the effector and execution phases. At the end of the execution phase, the apoptotic signal must come back to the nucleus to produce internucleosomal DNA degradation. Therefore, the induction of apoptosis from detection and subsequent processing of cisplatin-induced DNA damage seems to be a long and complex process of cell death. However, because cisplatin is a nonspecific drug and reacts not only with DNA but also with proteins,we cannot rule out the possibility that in some cases of cisplatin induced apoptosis, an easier process of initiation, such as damage to cytoplasmic proteins, may take place (Perez, 1998). Thus, damage to proteins is worth considering as a factor contributing to cisplatin-induced apoptosis. Moreover, it is possible that cisplatin damage to proteins could induce apoptosis at the execution phase level. In fact, initiation of apoptosis at the execution phase (activation of caspases) has been previously reported for the cell killing produced by cytotoxic T lymphocytes (Golstein et al., 1991). Although apoptosis and necrosis are conceptually distinct forms of cell death with very different morphological and biochemical characteristics, these two types of demise may occur simultaneously in tissues or cell cultures exposed to the same insult (Eguchi et al., 1997, Zhan et al., 1999). In fact, both types of cell death have been found in the same population of cisplatin-treated cells (Pestell et al., 2000). Moreover, it has been hypothesized that in a tissue or cell population,apoptosis and necrosis might be two extremes of a continuum of possible types of cell demise. Individual cell death would be decided by factors such as the availability of energy and the metabolic condition of the cell (Leist et al., 1997). Thus, some cells might die as a result of an unfinished apoptotic program. In fact, in L1210 leukemic cells, cisplatin-induced cell death seems to be the result of a defective apoptotic program that lacks some morphological and biochemical characteristics attributed to apoptosis (Segal-Bendirdjian and Jacquemin-Sablon, 1995). In addition, at high doses, cisplatin could damage molecules involved in cellular energy supply (i.e., ATP) and also proteins directly or indirectly involved in the apoptotic process (i.e., p53, Bax, Bcl-2, and caspases), leading to necrotic cell death. In fact, in cisplatin-resistant keratinocytes transformed by H-ras oncogene, a high dose of cisplatin (312 microM) induces characteristic features of necrotic cell death(Perez et al., 1999). Thus, depending on the level of cellular damage induced by cisplatin, necrosis could take place either directly or as a consequence of an unfinished apoptotic program. In summary, a growing body of evidence suggests that cisplatin induced cell death does not always come from "classic"apoptosis. Depending on both cisplatin dose and cellular status, cisplatin may also induced cell death by a defective apoptotic program or even by necrosis. Elucidation of the conditions under which the apoptotic program induced by cisplatin as well as other antitumor drugs is totally or partially executed may have important implications for the outcome of cancer chemotherapy. PMID- 11259609 TI - Maximal aryl hydrocarbon receptor activity depends on an interaction with the retinoblastoma protein. AB - The aryl hydrocarbon receptor (AhR) belongs to the basic helix-loop helix/periodicity/AhR nuclear translocator/simple-minded (Per-Arnt-Sim) family of transcription factors that regulate critical functions during development and tissue homeostasis. Within this family, the AhR is the only member conditionally activated in response to ligand binding, typified by 2,3,7,8-tetrachlorodibenzo-p dioxin (TCDD). We recently demonstrated that the AhR interacts with the retinoblastoma protein (pRb). This report presents evidence that a LXCXE motif in the AhR protein confers pRb binding, which is necessary for maximal TCDD induced G(1) arrest in rat 5L hepatoma cells. The data support a mechanism whereby pRb seems to regulate G(1) cell cycle progression distinct from the direct repression of E2F-mediated transcription. Furthermore, the results indicate that the AhR-pRb interaction regulates TCDD induction of CYP1A1, suggesting that pRb may be a general AhR coactivator. PMID- 11259610 TI - Nootropic drug modulation of neuronal nicotinic acetylcholine receptors in rat cortical neurons. AB - Nefiracetam (DM-9384) is a new pyrrolidone nootropic drug being developed for the treatment of Alzheimer's type and poststroke vascular-type dementia. Because the cholinergic system plays an important role in cognitive functions and Alzheimer's disease dementia, the present study was conducted to elucidate the mechanism of action of nefiracetam and aniracetam on neuronal nicotinic acetylcholine receptors (nnAChRs). Currents were recorded from rat cortical neurons in long term primary culture using the whole-cell, patch-clamp technique. Two types of currents were evoked by acetylcholine (ACh): alpha-bungarotoxin-sensitive, alpha 7-type currents and alpha-bungarotoxin-insensitive, alpha 4 beta 2-type currents. Although nefiracetam and aniracetam inhibited alpha 7-type currents only weakly, these nootropic agents potentiated alpha 4 beta 2-type currents in a very potent and efficacious manner. Nefiracetam at 1 nM and aniracetam at 0.1 nM reversibly potentiated alpha 4 beta 2-type currents to 200 to 300% of control. Nefiracetam at very high concentrations (approximately 10 microM) also potentiated alpha 4 beta 2-type currents but to a lesser extent, indicative of a bell-shaped dose response relationship. Nefiracetam markedly increased the saturating responses induced by high concentrations of ACh. However, human alpha 4 beta 2 subunits expressed in human embryonic kidney cells were inhibited rather than potentiated by nefiracetam. The specific protein kinase A inhibitors (H-89, KT5720, and peptide 5-24) and protein kinase C inhibitors (chelerythrine, calphostin C, and peptide 19--63) did not prevent nefiracetam from potentiating alpha 4 beta 2-type currents, indicating that these protein kinases are not involved in nefiracetam action. The nefiracetam potentiating action was not affected by 24-h pretreatment of neurons with pertussis toxin, but was abolished by cholera toxin. Therefore, G(s) proteins, but not G(i)/G(o) proteins, are involved in nefiracetam potentiation. These results indicate that nnAChRs are an important site of action of nefiracetam and G(s) proteins may be its crucial target. PMID- 11259611 TI - Structural and gating changes of the sodium channel induced by mutation of a residue in the upper third of IVS6, creating an external access path for local anesthetics. AB - Membrane-impermeant quaternary amine local anesthetics QX314 and QX222 can access their binding site on the cytoplasmic side of the selectivity filter from the outside in native cardiac Na(+) channels. Mutation of domain IV S6 Ile-1760 of rat brain IIA Na(+) channel or the equivalent (Ile-1575) in the adult rat skeletal muscle isoform (mu 1) creates an artificial access path for QX. We examined the characteristics of mutation of mu 1-I1575 and the resulting QX path. In addition to allowing external QX222 access, I1575A accelerated decay of Na(+) current and shifted steady-state availability by -27 mV. I1575A had negligible effects on inorganic or organic cation selectivity and block by tetrodotoxin (TTX), saxitoxin (STX), or mu-conotoxin (mu-CTX). It exposed a site within the protein that binds membrane-permeant methanethiosulfonate ethylammonium (MTSEA), but not membrane-impermeant methanethiosulfonate ethyltrimethylammonium (MTSET) and methanethiosulfonate ethylsulfonate (MTSES). MTSEA binding abolished the QX path created by this mutation, without effects on toxin binding. The mu-CTX derivative R13N, which partially occluded the pore, had no effect on QX access. I1575A exposed two Cys residues because a disulfide bond was formed under oxidative conditions, but the exposed Cys residues are not those in domain IV S6, adjacent to Ile-1575. The Cys mutant I1575C was insensitive to external Cd(2+) and MTS compounds (MTSEA, MTSET, MTSES), and substitution of Ile with a negatively charged residue (I1575E) did not affect toxin binding. Ile-1575 seems to be buried in the protein, and its mutation disrupts the protein structure to create the QX path without disturbing the outer vestibule and its selectivity function. PMID- 11259612 TI - Identification of two prokineticin cDNAs: recombinant proteins potently contract gastrointestinal smooth muscle. AB - The motility of gastrointestinal tract is regulated by classical neurotransmitters, neuropeptides, and humoral agents. Two novel human cDNAs have been cloned based on their sequence similarity to a frog skin secretion protein, Bv8, and a nontoxic protein of mamba snake venom. These human cDNAs encode two secreted proteins of 86 and 81 amino acids. Northern blot hybridization has revealed that these cDNAs are expressed in gastrointestinal tract, particularly the stomach. Recombinant proteins with authentic N-terminal sequences have been produced in Escherichia coli and refolded into functional proteins by careful control of protein aggregation. Mass spectrometry has confirmed the formation of five pairs of disulfide bonds. The refolded recombinant proteins potently contract gastrointestinal smooth muscle with EC(50) values in the subnanomolar range. The contractile effects of the recombinant proteins are specific for gastrointestinal smooth muscle, because they have no effect on vascular or respiratory smooth muscle. To reflect their potent and specific effects on gastrointestinal smooth muscle cells, we have named these recombinant proteins prokineticins. Ligand binding studies with iodinated prokineticin revealed the presence of a high-affinity site in ileal smooth muscle. The displacement of specific binding by GTP gamma S suggests that the prokineticin receptor may belong to the family of G protein-coupled receptors. Experiments with verapamil and nifedipine revealed that calcium influx is essential for the contractile activity of prokineticins on gastrointestinal smooth muscle. In summary, we have identified two novel endogenous regulators of gastrointestinal motility. The availability of recombinant prokineticins should provide novel therapeutic agents for disorders involving impaired gastrointestinal motility. PMID- 11259613 TI - Transcriptional regulation of topoisomerase II alpha at confluence and pharmacological modulation of expression by bis-benzimidazole drugs. AB - Topoisomerase II alpha is a critical gene involved in DNA replication and maintenance of genomic stability. Several chemotherapeutic agents target topoisomerase II and levels of expression are an important factor in chemosensitivity. Transcriptional regulation has been demonstrated to regulate topoisomerase II alpha levels under several circumstances, including cellular confluence, heat shock, and expression of oncogenes including ras and myb. Expression of topoisomerase II alpha is regulated by cellular proliferation; transcriptional down-regulation in confluent cells is modulated through sequences within the promoter. In this study, we examined DNA-protein interactions within the topoisomerase II alpha promoter in exponential and confluent phase NIH3T3 cells. Using electrophoretic mobility shift assay and in vitro DNase I footprint experiments, the involvement of NF-Y in transcriptional regulation was established. Incubation of the DNA minor groove-binding agents Hoechst 33342 and Hoechst 33258 with nuclear extracts revealed drug binding to regions surrounding the inverted CCAAT boxes within the topoisomerase II alpha promoter and displacement of proteins binding to these elements. Addition of both Hoechst 33342 and Hoechst 33258 to NIH3T3 cells at confluence resulted in increased expression of topoisomerase II alpha. In addition, MTT cytotoxicity assays in confluent cells showed an additive effect of incubation with Hoechst 33342 and the topoisomerase II alpha poison etoposide. Therefore, DNA binding drugs which block transcription factor activation of the promoter may deregulate topoisomerase II alpha and this strategy may be of value in modifying gene expression and modulating chemosensitivity. PMID- 11259614 TI - Human interferon-inducible 10-kDa protein and human interferon-inducible T cell alpha chemoattractant are allotopic ligands for human CXCR3: differential binding to receptor states. AB - The human CXC chemokines IP-10 (10-kDa interferon-inducible protein), MIG (monokine induced by human interferon-gamma), and I-TAC (interferon-inducible T cell alpha chemoattractant) attract lymphocytes through activation of CXCR3. In the studies presented here, we examined interaction of these chemokines with human CXCR3 expressed in recombinant cells and human peripheral blood lymphocytes (PBL). IP-10, MIG, and I-TAC were agonists in stimulating [(35)S]GTP gamma S binding in recombinant cell and PBL membranes but had no effect in the absence of hCXCR3 expression. (125)I-IP-10 and (125)I-I-TAC bound hCXCR3 with high affinity, although the (125)I-I-TAC B(max) value in saturation bindings was 7- to 13-fold higher than that measured with (125)I-IP-10. Coincubation with unlabeled chemokines decreased (125)I-IP-10 binding with a single discernible affinity. However, with (125)I-I-TAC, competition with IP-10 or MIG was incomplete, and multiple binding affinities were evident. Moreover, in contrast to I-TAC, IP-10 and MIG binding IC(50) values did not increase predictably with increased (125)I I-TAC concentration in competition bindings, suggesting that these chemokines are noncompetitive (i.e., allotopic) ligands. Uncoupling of hCXCR3 eliminated (125)I IP-10 binding but only decreased (125)I-I-TAC binding 30 to 80%, indicating that unlike IP-10, I-TAC binds with high affinity to uncoupled (R) and coupled (R*) hCXCR3. To examine chemokine binding to R*, we tested the effect of anti-hCXCR3 antibody on I-TAC- and IP-10-stimulated [(35)S]GTP gamma S binding. The antibody attenuated [(35)S]GTP gamma S binding in response to IP-10 but not to I-TAC, suggesting that the two chemokines bind differently to R*. Moreover, increased occupancy of R* with a >75-fold increase in (125)I -IP-10 concentration did not increase the I-TAC binding IC(50) value, and I-TAC increased the dissociation rate of (125)I-IP-10. From these data, we conclude that the binding of IP-10 and I-TAC to the R* state of hCXCR3 is allotopic. PMID- 11259615 TI - Cell-specific regulation of human aryl hydrocarbon receptor expression by transforming growth factor-beta(1). AB - Previous studies showed that TGF-beta down-regulates aryl hydrocarbon (AhR) expression in human lung carcinoma cells A549. Here we analyzed the molecular mechanisms by which TGF-beta modulates AhR expression. A 5799-nucleotide 5' flanking region of human AhR gene was isolated. Transient transfection studies of full-length (hAhRP) and deletion promoter constructs indicate the requirement of a cis-regulatory element encompassing -1980 to -1892 for full constitutive activity. Basal hAhRP activity occurs in a cell-specific manner; human hepatoma HepG2 cells possess a 10-fold higher activity compared with A549 cells. TGF-beta exerts cell-specific effects on hAhRP activity. Treatment of cells with 100 pM TGF-beta leads to a 50% inhibition in A549 and a 3-fold induction in HepG2 cells. Deletion mutagenesis identified a TGF-beta-responsive sequence containing a functional conserved Smad-binding element. Transient overexpression of Smad 2, 3, and 4 indicates that these signal transducers modulate hAhRP activity. The down regulation of AhR by TGF-beta is modulated by 5'-TG-3'-interacting factor (TGIF). Transient overexpression of TGIF in MDA-MB231 and HepG2 cells led to inhibition of hAhRP activity and a similar decrease of AhR mRNA expression. Our findings indicate that Smad proteins are involved in the cell-specific regulation of AhR expression by TGF-beta. PMID- 11259616 TI - 2'-C-cyano-2'-deoxy-1-beta-D-arabino-pentofuranosylcytosine: a novel anticancer nucleoside analog that causes both DNA strand breaks and G(2) arrest. AB - The mechanism of 2'-C-cyano-2'-deoxy-1-beta-D-arabino-pentofuranosylcytosine (CNDAC) action was investigated in human lymphoblastoid CEM cells and myeloblastic leukemia ML-1 cells. CNDAC was metabolized to its 5'-triphosphate and incorporated into DNA, which was associated with inhibition of DNA synthesis. After incubation of cells with [(3)H]CNDAC, metabolites were detected in 3'-->5' phosphodiester linkage and at the 3' terminus of cellular DNA. Specific enzymatic hydrolysis of DNA demonstrated that the parent nucleoside and its 2'-epimer 2'-C cyano-2'-deoxy-2-ribo-pentofuranosylcytosine accounted for approximately 65% of the total analogs incorporated into DNA and essentially all of the drug in the 3' ->5' phosphodiester linkage. In contrast, all detectable radioactivity at 3' termini was associated with 2'-C-cyano-2',3'-didehydro-2',3'-dideoxycytidine. This de facto DNA chain-terminating nucleotide arises from an electronic characteristic and cleavage of the 3'-phosphodiester bond subsequent to the addition of a nucleotide to the incorporated CNDAC moiety by beta-elimination, a process that generates a single strand break in DNA. Investigation of the biological consequences of these actions indicated that, after incubation with cytostatic concentrations of CNDAC, cell cycle progression was delayed during S phase, but that cells arrested predominantly in the G(2) phase. This differed from the S phase-arresting actions of ara-C and gemcitabine, other deoxycytidine analogs that inhibit DNA replication but do not cause strand breaks. Thus, once incorporated into DNA, the CNDAC molecule appears to act by a dual mechanism that 1) delays the progress of further DNA replication, but 2) upon addition of a deoxynucleotide results in the conversion of the incorporated analog to a de facto DNA chain terminator at the 3' terminus of a single strand break. It is likely that DNA strand breaks trigger cell cycle arrest in G(2). PMID- 11259617 TI - Modulation of neuronal nicotinic acetylcholine receptors by halothane in rat cortical neurons. AB - Inhalational general anesthetics have recently been shown to inhibit neuronal nicotinic acetylcholine (ACh) receptors (nnAChRs) expressed in Xenopus laevis oocytes and in molluscan neurons. However, drug actions on these systems are not necessarily the same as those seen on native mammalian neurons. Thus, we analyzed the detailed mechanisms of action of halothane on nnAChRs using rat cortical neurons in long-term primary culture. Currents induced by applications of ACh via a U-tube system were recorded by the whole-cell, patch-clamp technique. ACh evoked two types of currents, alpha-bungarotoxin-sensitive, fast desensitizing (alpha 7-type) currents and alpha-bungarotoxin-insensitive, slowly desensitizing (alpha 4 beta 2-type) currents. Halothane suppressed alpha 4 beta 2-type currents more than alpha 7-type currents with IC(50) values of 105 and 552 microM, respectively. Halothane shifted the ACh dose-response curve for the alpha 4 beta 2-type currents in the direction of lower ACh concentrations and slowed its apparent rate of desensitization. The rate of recovery after washout from halothane block was much faster than the rate of recovery from ACh desensitization. Thus, the halothane block was not caused by receptor desensitization. Chlorisondamine, an irreversible open channel blocker for nnAChRs, caused a time-dependent block that was attenuated by halothane. These results could be accounted for by kinetic simulation based on a model in which halothane causes flickering block of open channels, as seen in muscle nAChRs. Halothane block of nnAChRs is deemed to play an important role in anesthesia via a direct action on the receptor and an indirect action to suppress transmitter release. PMID- 11259618 TI - Phthalates rapidly increase production of reactive oxygen species in vivo: role of Kupffer cells. AB - The role of oxidants in the mechanism of tumor promotion by peroxisome proliferators remains controversial. The idea that induction of acyl-coenzyme A oxidase leads to increased production of H(2)O(2), which damages DNA, seems unlikely; still, free radicals might be important in signaling in specialized cell types such as Kupffer cells, which produce mitogens. Because hard evidence for increased oxidant production in vivo after treatment with peroxisome proliferators is lacking, the spin-trapping technique and electron spin resonance spectroscopy were used. Rats were given di(2-ethylhexyl) phthalate (DEHP) acutely. The spin trapping agent alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone was also given and bile samples were collected for 4 h. Under these conditions, the intensity of the six-line radical adduct signal increased to a maximum value of 2.5-fold 2 h after administration of DEHP, before peroxisomal oxidases were induced. Furthermore, DEHP given with [(13)C(2)]dimethyl sulfoxide produced a 12 line electron spin resonance spectrum, providing evidence that DEHP stimulates (*)OH radical formation in vivo. Furthermore, when rats were pretreated with dietary glycine, which inactivates Kupffer cells, DEHP did not increase radical signals. Moreover, similar treatments were performed in knockout mice deficient in NADPH oxidase (p47(phox) subunit). Importantly, DEHP increased oxidant production in wild-type but not in NADPH oxidase-deficient mice. These data provide evidence for the hypothesis that the molecular source of free radicals induced by peroxisome proliferators is NADPH oxidase in Kupffer cells. On the contrary, radical adduct formation was not affected in peroxisome proliferator activated receptor alpha knockout mice. These observations represent the first direct, in vivo evidence that phthalates increase free radicals in liver before peroxisomal oxidases are induced. PMID- 11259619 TI - Activation of the p53 DNA damage response pathway after inhibition of DNA methyltransferase by 5-aza-2'-deoxycytidine. AB - Transcriptional silencing of tumor suppressor genes by DNA methylation occurs in cancer cell lines and in human tumors. This has led to the pursuit of DNA methyltransferase inhibition as a drug target. 5-Aza-2'-deoxycytidine [5-aza-CdR (decitabine)], a potent inhibitor of DNA methyltransferase, is a drug currently in clinical trials for the treatment of solid tumors and leukemia. The efficacy of 5-aza-CdR may be related to the induction of methylation-silenced tumor suppressor genes, genomic hypomethylation, and/or enzyme-DNA adduct formation. Here, we test the hypothesis that 5-aza-CdR treatment is perceived as DNA damage, as assessed by the activation of the tumor suppressor p53. We show that 1) colon tumor cell lines expressing wild-type p53 are more sensitive to 5-aza-CdR mediated growth arrest and cytotoxicity; 2) the response to 5-aza-CdR treatment includes the induction and activation of wild-type but not mutant p53 protein; and 3) the induction of the downstream p53 target gene p21 is partially p53 dependent. The induction of p53 protein after 5-aza-CdR treatment did not correlate with an increase in p53 transcripts, indicating that hypomethylation at the p53 promoter does not account for the p53 response. It is relevant that 5-aza CdR has shown the greatest promise in clinical trials for the treatment of chronic myelogenous leukemia, a malignancy in which functional p53 is often retained. Our data raise the hypothesis that p53 activation may contribute to the clinical efficacy and/or toxicity of 5-aza-CdR. PMID- 11259620 TI - Direct and differential interaction of beta-arrestins with the intracellular domains of different opioid receptors. AB - beta-arrestins have been shown to play important roles in regulation of signaling and desensitization of opioid receptors in many in vivo studies. The current study was carried out to measure the direct interaction of beta-arrestins with two functional intracellular domains, the third intracellular loop (I3L) and the carboxyl terminus (CT), of delta-, mu-, and kappa-opioid receptors (DOR, MOR, and KOR, respectively). Results from the pull-down assay using glutathione S transferase fusion proteins demonstrated that beta-arrestins (1 and 2) were able to bind to the I3L of DOR and to the CT of DOR and KOR. Surface plasmon resonance measurement gave similar results with typical dissociation equilibrium constant (K(D)) values in the micromolar range. The site-directed mutagenesis experiment further revealed that certain specific serine/threonine residues in these receptor domains play a critical role in their interaction with beta-arrestins. Taken together, our data clearly indicated that beta-arrestins interact differentially with the functional domains of different opioid receptors; this may provide a possible molecular basis for differential regulation of opioid receptors by beta-arrestins. PMID- 11259621 TI - Differential effects of rexinoids and thiazolidinediones on metabolic gene expression in diabetic rodents. AB - Both retinoid X receptor (RXR)-selective agonists (rexinoids) and thiazolidinediones (TZDs), PPAR (peroxisome proliferator-activated receptor) gamma-specific ligands, produce insulin sensitization in diabetic rodents. In vitro studies have demonstrated that TZDs mediate their effects via the RXR/PPAR gamma complex. To determine whether rexinoids lower hyperglycemia by activating the RXR/PPAR-gamma heterodimer in vivo, we compared the effects of a rexinoid (LG100268) and a TZD (rosiglitazone) on gene expression in white adipose tissue, skeletal muscle, and liver of Zucker diabetic fatty rats (ZDFs). In adipose tissue, rosiglitazone decreased tumor necrosis factor-alpha (TNF-alpha) mRNA and induced glucose transporter 4 (GLUT4), muscle carnitine palmitoyl-transferase (MCPT), stearoyl CoA desaturase (SCD1), and fatty acid translocase (CD36). In contrast, LG100268 increased TNF-alpha and had no effect or suppressed the expression of GLUT4, MCPT, SCD1, and CD36. In liver, the rexinoid increased MCPT, SCD1, and CD36 mRNAs, whereas rosiglitazone induced only a small increase in CD36. In skeletal muscle, rosiglitazone and LG100268 have similar effects; both increased SCD1 and CD36 mRNAs. The differences in the pattern of genes induced by the rexinoids and the TZDs in diabetic animals found in these studies suggests that these compounds may have independent and tissue-specific effects on metabolic control in vivo. PMID- 11259622 TI - Interaction of co-expressed mu- and delta-opioid receptors in transfected rat pituitary GH(3) cells. AB - mu- and delta-Opioid agonists interact in a synergistic manner to produce analgesia in several animal models. Additionally, receptor binding studies using membranes derived from brain tissue indicate that interactions between mu- and delta-opioid receptors might be responsible for the observation of multiple opioid receptor subtypes. To examine potential interactions between mu- and delta opioid receptors, we examined receptor binding and functional characteristics of mu-, delta-, or both mu- and delta-opioid receptors stably transfected in rat pituitary GH(3) cells (GH(3)MOR, GH(3)DOR, and GH(3)MORDOR, respectively). Saturation and competition binding experiments revealed that coexpression of mu- and delta-opioid receptors resulted in the appearance of multiple affinity states for mu- but not delta-opioid receptors. Additionally, coadministration of selective mu- and delta-opioid agonists in GH(3)MORDOR cells resulted in a synergistic competition with [(3)H][D-Pen(2,5)]enkephalin (DPDPE) for delta opioid receptors. Finally, when equally effective concentrations of [D-Ala(2),N MePhe(4),Gly-ol(5)]enkephalin (DAMGO) and two different delta-opioid agonists (DPDPE or 2-methyl-4a alpha-(3-hydroxyphenyl)-1,2,3,4,4a,5,12,12a alpha octahydroquinolino-[2,3,3-g]-isoquinoline; TAN67) were coadministered in GH(3)MORDOR cells, a synergistic inhibition of adenylyl cyclase activity was observed. These results strongly suggest that cotransfection of mu- and delta opioid receptors alters the binding and functional characteristics of the receptors. Therefore, we propose that the simultaneous exposure of GH(3)MORDOR cells to selective mu- and delta-opioid agonists produces an interaction between receptors resulting in enhanced receptor binding. This effect is translated into an augmented ability of these agonists to inhibit adenylyl cyclase activity. Similar interactions occurring in neurons that express both mu- and delta-opioid receptors could explain observations of multiple opioid receptor subtypes in receptor binding studies and the synergistic interaction of mu- and delta-opioids in analgesic assays. PMID- 11259623 TI - Induction of CDK inhibitors (p21(WAF1) and p27(Kip1)) and Bak in the beta lapachone-induced apoptosis of human prostate cancer cells. AB - beta-Lapachone, a novel anti-neoplastic drug, induces various cancer cells to undergo apoptosis. In a previous report, we showed that beta-lapachone-induced apoptosis of HL-60 cells is mediated by oxidative stress. However, in the present study, we found that beta-lapachone-induced apoptosis of human prostate cancer (HPC) cells may be independent of oxidative stress. In contrast to the 10-fold beta-lapachone-induced increase in H(2)O(2) production seen in HL-60 cells, only a 2- to 4-fold increase was observed in HPC cells. N-acetyl-L-cysteine (NAC), a thiol antioxidant, inhibited the apoptosis in DU145 cells after 12 h exposure to beta-lapachone. Nonetheless, NAC, along with other antioxidants, failed to exert similar effect in HPC cells subjected to beta-lapachone treatment for 24 h. Under this premise, we suggest that the oxidative stress may not play a crucial role in beta-lapachone-mediated HPC cell apoptosis. Here we demonstrate that damage to genomic DNA is the trigger for the apoptosis of HPC cells induced by beta lapachone. According to our results, beta-lapachone stimulates DNA dependent kinase expression and poly(ADP-ribose) polymerase cleavage in advance of significant morphological changes. beta-Lapachone promotes the expression of cyclin-dependent kinase (cdk) inhibitors (p21(WAF1) and p27(Kip1)), induces bak expression, and subsequently stimulates the activation of caspase-7 but not of caspase-3 or caspase-8 during the apoptosis of HPC cells. Taken together, these results suggest that the signaling pathway involving the beta-lapachone-induced apoptosis of HPC cell may be by DNA damage, induction of cdk inhibitors (p21 and p27), and then subsequent stimulation of caspase-7 activation. PMID- 11259624 TI - Sodium salicylate increases CYP2E1 levels and enhances arachidonic acid toxicity in HepG2 cells and cultured rat hepatocytes. AB - Sodium salicylate and acetylsalicylic acid are drugs used as anti-inflammatory agents. Salicylate prevents nuclear factor-kappa B activation and can cause apoptosis. However, salicylate, a substrate of CYP2E1, is also an antioxidant and can scavenge reactive oxygen species. Experiments were carried out to evaluate whether salicylate can modulate CYP2E1-dependent toxicity. Addition of a polyunsaturated fatty acid such as arachidonic acid (AA) to HepG2 cells resulted in loss of cell viability, especially in cells expressing CYP2E1 (E47 cells). Toxicity was enhanced by the addition of 1 to 10 mM salicylate to the E47 cells but not to control HepG2 cells or HepG2 cells expressing CYP3A4. Salicylate alone was not toxic, and the enhanced toxicity by AA in the presence of salicylate was prevented by diallyl sulfide, a CYP2E1 inhibitor, and by the antioxidant (+/-)6 hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid. Salicylate potentiated AA induced lipid peroxidation in the E47 cells, a reaction blocked by diallyl sulfide. CYP2E1 levels were elevated by salicylate at concentrations (<5 mM), which did not increase CYP2E1 mRNA levels. This increase was associated with a decrease of CYP2E1 turnover by salicylate in the presence of cycloheximide. Salicylate also potentiated AA toxicity in hepatocytes isolated from pyrazole treated rats with high levels of CYP2E1 and from saline controls. In view of the potential role of CYP2E1 in contributing to alcohol-induced oxidative stress and liver injury, the potentiation of CYP2E1-dependent toxicity and the elevation of CYP2E1 levels by salicylate may be of clinical significance and merit caution in the use of salicylate and salicylate precursors such as acetylsalicylic acid with certain other drugs. PMID- 11259625 TI - P-glycoprotein limits oral availability, brain, and fetal penetration of saquinavir even with high doses of ritonavir. AB - The low oral bioavailability of the HIV protease inhibitor (HPI) saquinavir is dramatically increased by coadministration of the HPI ritonavir. Because saquinavir and ritonavir are substrates and inhibitors of both the drug transporter P-glycoprotein (P-gp) and of the metabolizing enzyme CYP3A4, we wanted to sort out whether the ritonavir effect is primarily mediated by inhibition of CYP3A4 or P-gp or both. P-gp is known to limit the bioavailability, brain, testis, and fetal penetration of its substrates, so effective inhibition of P-gp by ritonavir in vivo might open up pharmacological sanctuary sites for saquinavir, with the potential of beneficial effects on therapy, but also of increased toxicity. In vitro, P-gp-mediated transport of saquinavir and ritonavir was only moderately inhibited by both HPIs compared with the potent P-gp inhibitor PSC833. When [(14)C]saquinavir was orally coadministered with a maximum tolerated dose of ritonavir to wild-type and P-gp-deficient mice, saquinavir bioavailability was dramatically increased in both strains, but P-gp still limited the oral bioavailability of saquinavir, and its penetration into brain and fetus. These data indicate that in vivo, ritonavir is a relatively poor P-gp inhibitor. The highly increased bioavailability of saquinavir because of ritonavir coadministration most likely results from reduced saquinavir metabolism. Importantly, our data indicate that it is unlikely that ritonavir coadministration will substantially affect the contribution of P-gp to pharmacological sanctuary sites such as brain, testis, and fetus. Thus, if one wanted to effectively open these sites for therapeutic purposes, more efficient P gp inhibitors should be applied. PMID- 11259626 TI - Distinct functional and pharmacological properties of tonic and quantal inhibitory postsynaptic currents mediated by gamma-aminobutyric acid(A) receptors in hippocampal neurons. AB - gamma-Aminobutyric acid (GABA), the principal inhibitory neurotransmitter, activates a persistent low amplitude tonic current in several brain regions in addition to conventional synaptic currents. Here we demonstrate that GABA(A) receptors mediating the tonic current in hippocampal neurons exhibit functional and pharmacological properties different from those of quantal synaptic currents. Patch-clamp techniques were used to characterize miniature inhibitory postsynaptic currents (mIPSCs) and the tonic GABAergic current recorded in CA1 pyramidal neurons in rat hippocampal slices and in dissociated neurons grown in culture. The competitive GABA(A) receptor antagonists, bicuculline and picrotoxin, blocked both the mIPSCs and the tonic current. In contrast, mIPSCs but not the tonic current were inhibited by gabazine (SR-95531). Coapplication experiments and computer simulations revealed that gabazine bound to the receptors responsible for the tonic current but did not prevent channel activation. However, gabazine competitively inhibited bicuculline blockade. The unitary conductance of the GABA(A) receptors underlying the tonic current (approximately 6 pS) was less than the main conductance of channels activated during quantal synaptic transmission (approximately 15--30 pS). Furthermore, compounds that potentiate GABA(A) receptor function including the benzodiazepine, midazolam, and anesthetic, propofol, prolonged the duration of mIPSCs and increased tonic current amplitude in cultured neurons to different extents. Clinically-relevant concentrations of midazolam and propofol caused a greater increase in tonic current compared with mIPSCs, as measured by total charge transfer. In summary, the receptors underlying the tonic current are functionally and pharmacologically distinct from quantally activated synaptic receptors and these receptors represent a novel target for neurodepressive drugs. PMID- 11259628 TI - Modulation of cisplatin cytotoxicity and cisplatin-induced DNA cross-links in HepG2 cells by regulation of glutathione-related mechanisms. AB - Glutathione (GSH), glutathione S-transferase (GST), and glutathione conjugate export pump (GS-X pump) have been shown to participate collectively in the detoxification of many anticancer drugs, including cisplatin. Identification and regulation of the rate-limiting step in the overall system for cisplatin detoxification is of crucial importance for sensitization of human tumor cells to cisplatin. In this study, the GSH content, GST activity, and GS-X pump activity were regulated separately to examine effects of the regulation on cisplatin cytotoxicity and cisplatin-induced DNA interstrand cross-links (ICL) in HepG2 cells. Seventy-percent depletion of GSH by buthionine sulfoximine (BSO) and 50% increase of GSH by monoethyl GSH ester (GSHe) potentiated and decreased cisplatin cytotoxicity, respectively. This was reflected by a significant decrease and increase of their respective IC(50) values by 62 and 107%. Cisplatin-induced ICL was also potentiated by depletion of GSH by BSO and decreased by enrichment of GSH by GSHe, as shown by a 125% increase and a 34% decrease of cross-linked DNA compared with control samples exposed to cisplatin alone (p = 0.008 and 0.03, respectively). On the other hand, inhibition of GST and GS-X pump by ethacrynic acid, quercetin, tannic acid, and indomethacin at concentrations that inhibited activities of GST and GS-X pump by more than 50% had no significant effects on cisplatin cytotoxicity and cisplatin-induced DNA ICL in these cells. The results showed that of the parameters measured, intracellular GSH seems to be the rate limiting factor, and its regulation would provide a more promising strategy for sensitization of human liver tumor cells to cisplatin. PMID- 11259627 TI - Direct effects of candesartan and eprosartan on human cloned potassium channels involved in cardiac repolarization. AB - In the present study, we analyzed the effects of two angiotensin II type 1 receptor antagonists, candesartan (0.1 microM) and eprosartan (1 microM), on hKv1.5, HERG, KvLQT1+minK, and Kv4.3 channels expressed on Ltk(-) or Chinese hamster ovary cells using the patch-clamp technique. Candesartan and eprosartan produced a voltage-dependent block of hKv1.5 channels decreasing the current at +60 mV by 20.9 +/- 2.3% and 14.3 +/- 1.5%, respectively. The blockade was frequency-dependent, suggesting an open-channel interaction. Eprosartan inhibited the tail amplitude of HERG currents elicited on repolarization after pulses to +60 mV from 239 +/- 78 to 179 +/- 72 pA. Candesartan shifted the activation curve of HERG channels in the hyperpolarizing direction, thus increasing the current amplitude elicited by depolarizations to potentials between -50 and 0 mV. Candesartan reduced the KvLQT1+minK currents elicited by 2-s pulses to +60 mV (38.7 +/- 6.3%). In contrast, eprosartan transiently increased (8.8 +/- 2.7%) and thereafter reduced the KvLQT1+minK current amplitude by 17.7 +/- 3.0%. Eprosartan, but not candesartan, blocked Kv4.3 channels in a voltage-dependent manner (22.2 +/- 3.5% at +50 mV) without modifying the voltage-dependence of Kv4.3 channel inactivation. Candesartan slightly prolonged the action potential duration recorded in guinea pig papillary muscles at all driving rates. Eprosartan prolonged the action potential duration in muscles driven at 0.1 to 1 Hz, but it shortened this parameter at faster rates (2--3 Hz). All these results demonstrated that candesartan and eprosartan exert direct effects on Kv1.5, HERG, KvLQT1+minK, and Kv4.3 currents involved in human cardiac repolarization. PMID- 11259629 TI - A single amino-acid in the TM1 domain is an important determinant of the desensitization kinetics of recombinant human and guinea pig alpha-homomeric 5 hydroxytryptamine type 3 receptors. AB - Desensitization of ligand-gated ion channels shapes synaptic responses and provides critical neuroprotection at central synapses, yet the molecular mechanisms underlying the desensitization process are poorly understood. Using the whole-cell voltage-clamp technique, we investigated desensitization kinetics of recombinant human and guinea pig alpha-homomeric 5-hydroxytryptamine type 3 (5 HT(3A)) receptors heterologously expressed in human embryonic kidney 293 cells. Human 5-HT(3A) receptors desensitize 3.5 times faster than does the homologous receptor from guinea pigs. By constructing various chimeras and through site directed mutagenesis, we have identified a single serine in the M1 region of the human 5-HT(3A) receptor sequence (S248) that, when substituted with threonine found in the equivalent guinea pig sequence (T254), conferred guinea pig-like kinetics on the time course of desensitization of the human receptor. Correspondingly, the reverse mutation (guinea pig T254S) resulted in a fast, human-like time constant of desensitization. Thus, the primary structure of the M1 region is an important determinant of desensitization kinetics of recombinant 5-HT(3A) receptors. PMID- 11259630 TI - Regulation of human monoamine oxidase B gene by Sp1 and Sp3. AB - The human monoamine oxidase (MAO) B plays a major role in the degradation of biogenic and dietary amines such as phenylethylamine, benzylamine, dopamine, and tyramine. We previously showed that the -246/-99 MAO B promoter region exhibited the highest activity and contained two clusters of overlapping Sp1 sites, a CACCC element and a TATA box. Here, using a series of 10 deletion constructs of the 2 kilobase pair 5'-flanking sequence, we identified additional potential regulatory elements, including activator proteins 1 and 4, CAAT, GATA, upstream stimulatory factor (USF), estrogen receptor (ER), and sex-determining region Y-box 5 (SOX5). Analysis of nine site-directed mutations of -246/-99 region reveals that both clusters of Sp1 sites contribute positively whereas the CACCC element contributes negatively to the transcriptional activity. Gel shift analysis demonstrates that in addition to Sp1, Sp3 can interact with both clusters of Sp1 sites. Cotransfection experiments show that Sp1 and its closely related family member Sp4 can trans-activate MAO B promoter activity through the proximal cluster of Sp1 sites and its activation can be repressed by the over-expression of Sp3 and a related family member BTEB2. These results suggest that the binding to the overlapping Sp1 sites by various members of Sp family is important for the regulation of the MAO B gene expression. PMID- 11259631 TI - Protein kinase C-alpha coordinately regulates cytosolic phospholipase A(2) activity and the expression of cyclooxygenase-2 through different mechanisms in mouse keratinocytes. AB - Transgenic mice (K5-PKC alpha) in which the keratin 5 promoter directs the expression of protein kinase C-alpha (PKC alpha) to epidermal keratinocytes display a 10-fold increase in PKC alpha protein in their epidermis and alterations in phorbol ester-induced cutaneous inflammation [J Cell Science 1999;112:3497-3506]. In the current study, we have used these K5-PKC alpha mice to examine the role of PKC alpha in keratinocyte phospholipid metabolism/eicosanoid production and cutaneous inflammation. Primary keratinocytes from wild-type and transgenic mice were prelabeled in culture with [(3)H]arachidonic acid (AA) and subsequently treated with TPA. Compared with wild type keratinocytes, K5-PKC alpha keratinocytes displayed a 2-fold increase in AA release. TPA treatment resulted in the phosphorylation of cPLA(2). PKC inhibitors GF-109203X or H7, but not mitogen-activated protein/extracellular signal regulated protein kinase (MEK) inhibitor PD 98059, could inhibit phosphorylation and AA release. Topical 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment of K5-PKC alpha mice resulted in a 5-fold increase in epidermal COX-2 induction and a 2- to 3-fold increase in prostaglandin (PG) E(2) levels above that observed in TPA-treated wild-type mice. PD 98059, GF-109203X, or H7 could block cyclooxygenase-2 (COX-2) induction by TPA. Because C/EBP beta, a basic leucine zipper transcription factor, can be activated via a PKC alpha/mitogen-activated protein kinase pathway and can influence COX-2 expression, we examined whether C/EBP beta is involved in TPA-induced epidermal COX-2 expression. TPA-induced COX 2 expression was similar in C/EBP beta nullizygous and wild-type mice. In summary, our results indicate that epidermal PKC alpha coordinately regulates cPLA(2) activity and COX-2 expression resulting in increased levels of AA and PGE(2). Furthermore, PKC alpha-induced AA release and cPLA(2) phosphorylation are independent of MEK, whereas PKC alpha-induced COX-2 expression and PGE(2) production are MEK-dependent and C/EBP beta-independent events. PMID- 11259632 TI - Inhibition of c-Jun N-terminal kinase 1, but not c-Jun N-terminal kinase 2, suppresses apoptosis induced by ischemia/reoxygenation in rat cardiac myocytes. AB - In the present study, rat cardiac myocytes were used as an in vitro ischemia/reperfusion injury model to delineate the role of c-Jun N-terminal kinase (JNK) 1 and JNK2 isoforms in ischemia/reoxygenation-induced apoptosis using an antisense approach. Exposure of rat cardiac myocytes to ischemia did not induce apoptosis as detected by staining with either acridine orange/ethidium bromide or annexin-V-fluorescein/propidium iodide. In contrast, a time-dependent increase in the number of apoptotic cells was noted after reoxygenation of ischemic myocytes, whereas the level of necrotic cells remained unaltered. Reoxygenation, but not ischemia alone, also caused a time-dependent increase in JNK activation that preceded apoptosis induction. Treatment of cardiac myocytes with antisense (AS) oligonucleotides that specifically targeted either JNK1 or JNK2 significantly reduced both mRNA and protein expression of the target isoform but had no effect on the expression of the alternate isoform. Pretreatment of cardiac myocytes with JNK1 AS, but not JNK2 AS, resulted in almost complete attenuation of reoxygenation-induced apoptosis. Furthermore, control oligonucleotides for JNK1 AS or JNK2 AS had no effect on JNK mRNA or protein expression or reoxygenation-induced apoptosis, indicating a sequence-specific mode of action. Additional studies revealed that apoptosis induced by other JNK activating stimuli, including ceramide, heat shock, and UV irradiation, was partly suppressed after treatment with JNK1 AS but not JNK2 AS. These findings demonstrate that the JNK1 isoform plays a preferential role in apoptosis induced by ischemia/reoxygenation as well as diverse JNK-activating cellular stresses. PMID- 11259633 TI - Activation of guanosine 5'-[gamma-(35)S]thio-triphosphate binding through M(1) muscarinic receptors in transfected Chinese hamster ovary cell membranes; 1. Mathematical analysis of catalytic G protein activation. AB - I analyzed in this work the effect of agonists and unlabeled guanyl nucleotides on [(35)S]GTP gamma S and [(3)H]NMS binding to transfected CHO cells expressing hM(1) muscarinic receptors. I was unable to explain my kinetic results by "traditional" (one-site, two-site, or two-step) bimolecular binding models. I therefore examined the equations that describe catalytic G protein activation. My results were fully consistent with the following interpretation: G protein coupled receptors either interacted with GDP-bound G proteins and facilitated the GDP release or recognized empty G proteins, depending on the incubation conditions. The receptor-coupled empty G proteins (RG) then recognized GTP gamma S, and the occupied G protein (G) dissociated reversibly from the receptor. Agonists accelerated the GDP release from receptor-coupled G proteins and accelerated the G dissociation: both effects accelerated synergically the G protein-GTP gamma S association reaction in the presence of GDP. GTP gamma S bound G proteins, G, competed efficiently with inactive (empty or GDP-bound) G proteins for receptor recognition, and were able, therefore, at low concentrations, to quench the [(35)S]GTP gamma S binding reaction. PMID- 11259634 TI - Activation of guanosine 5'-[gamma-(35)S]thio-triphosphate binding through M(1) muscarinic receptors in transfected Chinese Hamster ovary cell membranes: 2. Testing the "two-states" model of receptor activation. AB - I suggested in the accompanying article [Mol Pharmacol 2001;59:875-885] that muscarinic receptors catalyzed G protein activation. Acetylcholine or carbamylcholine recognition facilitated not only the GDP release from receptor coupled inactive G proteins but also the release of G from the (unstable) HRG complex. The two effects facilitated [(35)S]GTP gamma S binding in the presence of GDP, but could be studied separately by comparing [(35)S]GTP gamma S binding in the absence and presence of GTP. Guanyl nucleotides affected the efficiency of receptor-G protein coupling. The relative efficacies of partial agonists in the absence and presence of GTP should remain nonlinearly correlated if all agonists stabilize (to different extents) the same active receptor conformation. The correlation between M(1) muscarinic agonists' efficacy in accelerating [(35)S]GTP gamma S binding in the absence of other nucleotides and their in vivo efficacy (inositol phosphate accumulation) was in fact very poor. This probably reflected the presence of GTP in intact cells: pertussis toxin pretreatment (which inactivates the G(i/o) proteins) did not affect the agonists' efficacy profile (evaluated in the absence of spare receptors), but the addition of GTP to the [(35)S]GTP gamma S binding medium did. These results did not support the allosteric "two states" model of receptor activation, but suggested that different agonists induced different receptor conformations ("induced fit"). PMID- 11259635 TI - Sensitized increase of period gene expression in the mouse caudate/putamen caused by repeated injection of methamphetamine. AB - Methamphetamine (MAP) causes the sensitization phenomena not only in MAP-induced locomotor activity, dopamine release, and Fos expression, but also in MAP-induced circadian rhythm. Cocaine-induced sensitization is reportedly impaired in Drosophila melanogaster mutant for the Period (Per) gene. Thus, sensitization may be related to induction of the Per gene. A rapid induction of mPer1 and/or mPer2 in the suprachiasmatic nucleus after light exposure is believed to be necessary for light-induced behavioral phase shifting. Although the caudate/putamen (CPu) expresses mPer1 and/or mPer2 mRNA, the function of these genes in this nucleus has not yet been elucidated. Therefore, we examined whether MAP affects the expression of mPer1 and/or mPer2 mRNA in the mouse CPu. Injection of MAP augmented the expression of mPer1 but not mPer2 or mPer3 in the CPu, and this MAP induced increase in mPer1 expression lasted for 2 h. Also, the MAP-induced increase of mPer1 mRNA was strongly antagonized by pretreatment with a dopamine D1 receptor and N-methyl-D-aspartate (NMDA) receptor antagonist, but not by a D2 receptor antagonist. Interestingly, application of either the D1 or the D2 agonist alone did not cause mPer1 expression. The present results demonstrate that activation of both NMDA and D1 receptors is necessary to produce MAP-induced mPer1 expression in the CPu. Repeated injection of MAP caused a sensitization in not only the locomotor activity but also mPer1 expression in the CPu without affecting the level of mPer2, mPer3, or mTim mRNA. Thus, these results suggest that MAP-induced mPer1 gene expression may be related to the mechanism for MAP induced sensitization in the mouse. PMID- 11259636 TI - Cyclooxygenase inhibitors regulate the expression of a TGF-beta superfamily member that has proapoptotic and antitumorigenic activities. AB - The antitumorigenic activity of nonsteroidal anti-inflammatory drugs (NSAIDs), cyclooxygenase (COX) inhibitors, is well established, but responsible molecular mechanisms are not fully understood. NSAIDs stimulate apoptosis by COX dependent and independent mechanisms in colorectal cells in culture. Identification of genes regulated by COX inhibitors could lead to a better understanding of their proapoptotic and anti-neoplastic activities. Using subtractive hybridization, a cDNA which was designated as NSAID activated gene (NAG-1) was identified from NSAID-treated HCT-116, human colorectal cells. NAG-1 has an identical sequence with a novel member of the TGF-beta superfamily that has 5 different names. In the HCT-116 cells, NAG-1 expression is increased and apoptosis is induced by treatment with some NSAIDs in a concentration and time-dependent manner. NAG-1 transfected cells exhibited increased basal apoptosis, increased response to NSAIDs and reduced soft agar cloning efficiency. Furthermore, transplantable tumors derived from NAG-1 transfected HCT-116 cells showed reduced tumorigenicity in athymic nude mice compared with vector-transfected HCT-116 cells. The increased NAG-1 expression by NSAIDs provides a suitable explanation for COX independent apoptotic effects of NSAIDs in cultured cells. These data demonstrate that NAG-1 is an antitumorigenic and proapoptotic protein, and its regulation by COX inhibitors may provide new clues for explaining their proapoptotic and antitumorigenic activities. PMID- 11259637 TI - Competitive CYP2C9 inhibitors: enzyme inhibition studies, protein homology modeling, and three-dimensional quantitative structure-activity relationship analysis. AB - This study describes the generation of a three-dimensional quantitative structure activity relationship (3D-QSAR) model for 29 structurally diverse, competitive CYP2C9 inhibitors defined experimentally from an initial data set of 73 compounds. In parallel, a homology model for CYP2C9 using the rabbit CYP2C5 coordinates was built. For molecules with a known interaction mode with CYP2C9, this homology model, in combination with the docking program GOLD, was used to select conformers to use in the 3D-QSAR analysis. The remaining molecules were docked, and the GRID interaction energies for all conformers proposed by GOLD were calculated. This was followed by a principal component analysis (PCA) of the GRID energies for all conformers of all compounds. Based on the similarity in the PCA plot to the inhibitors with a known interaction mode, the conformer to be used in the 3D-QSAR analysis was selected. The compounds were randomly divided into two groups, the training data set (n = 21) to build the model and the external validation set (n = 8). The PLS (partial least-squares) analysis of the interaction energies against the K(i) values generated a model with r(2) = 0.947 and a cross-validation of q(2) = 0.730. The model was able to predict the entire external data set within 0.5 log units of the experimental K(i) values. The amino acids in the active site showed complementary features to the grid interaction energies in the 3D-QSAR model and were also in agreement with mutagenesis studies. PMID- 11259638 TI - Quinazolone-alkyl-carboxylic acid derivatives inhibit transmembrane Ca(2+) ion flux to (+)-(S)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid. AB - Comparison of the kinetics of the inward Ca(2+) ion flux to (S)-alpha-Amino-3 hydroxy-5-methylisoxazole-4-propionic acid [(S)-AMPA] in cerebrocortical homogenates and that of the previously reported transmembrane Na(+) ion influx mediated by an AMPA receptor in hippocampal homogenates established that the agonist-induced opening of the AMPA receptor channels occurs in two kinetically distinguishable phases. Here we report that the 2-methyl-4-oxo-3H-quinazoline-3 acetic acid (Q1) inhibits the major slow-phase response specifically, whereas the acetyl piperidine derivative (Q5) is a more potent inhibitor of the fast-phase response. Both the quinazolone-3-propionic acid (Q2) and the quinazolone-3-acetic acid methyl ester (Q3) enhanced the slow-phase response to (S)-AMPA. The information provided by docking different Q1, Q2, and Q5 models at the ligand binding core of iGluRs were used to define agonistic and antagonistic modes of interactions. Based on the effects of quinazolone-3-alkyl-carboxylic acid derivatives on specific [(3)H]Glu binding and kinetically distinguishable Ca(2+) ion permeability responses to (S)-AMPA and molecular modeling, the fast- and the slow-phase (S)-AMPA-elicited Ca(2+) ion fluxes were corresponded to different subunit compositions and degrees of S1S2 bridging interaction relative to substitution of kainate thereupon. Substitutions of agonists and antagonists into the iGluR2 S1S2 ligand binding core induced different modes of domain-domain bridging. PMID- 11259639 TI - The role of a conserved inter-transmembrane domain interface in regulating alpha(2a)-adrenergic receptor conformational stability and cell-surface turnover. AB - Functional and structural data from G protein-coupled receptors (GPCR) predict that transmembrane-domain (TM)2 is adjacent to TM7 within the GPCR structure, and that within this interface a conserved aspartate in TM2 and a conserved asparagine in TM7 exist in close proximity. Mutation at this D79(TM2)-N422(TM7) interface in the alpha(2A)-adrenergic receptor (alpha(2A)AR) affects not only receptor activation but also cell-surface residence time and conformational stability. Mutation at TM2(D79N) reduces allosteric modulation by Na(+) and receptor activation more dramatically than affecting cell-surface receptor turnover and conformational stability, whereas mutation at TM7(N422D) creates profound conformational instability and more rapid degradation of receptor from the surface of cells despite receptor activation and allosteric modulation properties that mirror a wild-type receptor. Double mutation of TM2 and 7(D79N/N422D) reveals phenotypes for receptor activation and conformational stability intermediate between the wild-type and singly mutated alpha(2A)AR. Additionally, the structural placement of a negative charge at this TM2/TM7 interface is necessary but not sufficient for receptor structural stability, because mislocalization of the negative charge in either the D79E alpha(2A)AR (which extends the charge out one methylene group) or the D79N/N422D alpha(2A)AR (placing the charge in TM7 instead of TM2) results in conformational lability in detergent solution and more rapid cell-surface receptor clearance. These studies suggest that this interface is important in regulating receptor cell-surface residence time and conformational stability in addition to its previously recognized role in receptor activation. PMID- 11259640 TI - Down-regulation of inducible nitric-oxide synthase (NOS-2) during parasite induced gut inflammation: a path to identify a selective NOS-2 inhibitor. AB - Nitric oxide (NO) possesses potent anti-inflammatory properties; however, an over production of NO will promote inflammation and induce cell and tissue dysfunction. Thus, the ability to precisely regulate NO production could prove beneficial in controlling damage. In this study, advantage was taken of the well characterized inflammatory response caused by an intestinal parasite, Trichinella spiralis, to study the relationship between intestinal inflammation and the regulation of nitric oxide synthase-type 2 (NOS-2) expression. Our study revealed that a specific gut inflammatory reaction results in inhibition of NOS-2 expression. Characteristics of this inhibition are: 1) local jejunal inflammation induced by T. spiralis systemically inhibits NOS-2 gene transcription, protein expression, and enzyme activity; 2) the inhibition blunts endotoxin-stimulated NOS-2 expression; 3) the inhibition does not extend to the expression of other isoforms of NOS, to paxillin, a housekeeper protein, or to cyclooxygenase-2, another protein induced by proinflammatory cytokines; 4) the inhibition is unlikely related to the formation of specific anti-parasite antibodies; and 5) the inhibition may involve substances other than stress-induced corticosteroids. Elucidation of such potent endogenous NOS-2 down-regulatory mechanisms could lead to the development of new strategies for the therapy of inflammatory conditions characterized by the overproduction of NO. PMID- 11259641 TI - Adaptation of the bovine spongiform encephalopathy agent to primates and comparison with Creutzfeldt-- Jakob disease: implications for human health. AB - There is substantial scientific evidence to support the notion that bovine spongiform encephalopathy (BSE) has contaminated human beings, causing variant Creutzfeldt-Jakob disease (vCJD). This disease has raised concerns about the possibility of an iatrogenic secondary transmission to humans, because the biological properties of the primate-adapted BSE agent are unknown. We show that (i) BSE can be transmitted from primate to primate by intravenous route in 25 months, and (ii) an iatrogenic transmission of vCJD to humans could be readily recognized pathologically, whether it occurs by the central or peripheral route. Strain typing in mice demonstrates that the BSE agent adapts to macaques in the same way as it does to humans and confirms that the BSE agent is responsible for vCJD not only in the United Kingdom but also in France. The agent responsible for French iatrogenic growth hormone-linked CJD taken as a control is very different from vCJD but is similar to that found in one case of sporadic CJD and one sheep scrapie isolate. These data will be key in identifying the origin of human cases of prion disease, including accidental vCJD transmission, and could provide bases for vCJD risk assessment. PMID- 11259642 TI - Selenoprotein oxidoreductase with specificity for thioredoxin and glutathione systems. AB - Thioredoxin (Trx) and glutathione (GSH) systems are considered to be two major redox systems in animal cells. They are reduced by NADPH via Trx reductase (TR) or oxidized GSH (GSSG) reductase and further supply electrons for deoxyribonucleotide synthesis, antioxidant defense, and redox regulation of signal transduction, transcription, cell growth, and apoptosis. We cloned and characterized a pyridine nucleotide disulfide oxidoreductase, Trx and GSSG reductase (TGR), that exhibits specificity for both redox systems. This enzyme contains a selenocysteine residue encoded by the TGA codon. TGR can reduce Trx, GSSG, and a GSH-linked disulfide in in vitro assays. This unusual substrate specificity is achieved by an evolutionary conserved fusion of the TR and glutaredoxin domains. These observations, together with the biochemical probing and molecular modeling of the TGR structure, suggest a mechanism whereby the C terminal selenotetrapeptide serves a role of a protein-linked GSSG and shuttles electrons from the disulfide center within the TR domain to either the glutaredoxin domain or Trx. PMID- 11259643 TI - Erythropoietin prevents neuronal apoptosis after cerebral ischemia and metabolic stress. AB - Erythropoietin (EPO) promotes neuronal survival after hypoxia and other metabolic insults by largely unknown mechanisms. Apoptosis and necrosis have been proposed as mechanisms of cellular demise, and either could be the target of actions of EPO. This study evaluates whether antiapoptotic mechanisms can account for the neuroprotective actions of EPO. Systemic administration of EPO (5,000 units/kg of body weight, i.p.) after middle-cerebral artery occlusion in rats dramatically reduces the volume of infarction 24 h later, in concert with an almost complete reduction in the number of terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling of neurons within the ischemic penumbra. In both pure and mixed neuronal cultures, EPO (0.1--10 units/ml) also inhibits apoptosis induced by serum deprivation or kainic acid exposure. Protection requires pretreatment, consistent with the induction of a gene expression program, and is sustained for 3 days without the continued presence of EPO. EPO (0.3 units/ml) also protects hippocampal neurons against hypoxia-induced neuronal death through activation of extracellular signal-regulated kinases and protein kinase Akt-1/protein kinase B. The action of EPO is not limited to directly promoting cell survival, as EPO is trophic but not mitogenic in cultured neuronal cells. These data suggest that inhibition of neuronal apoptosis underlies short latency protective effects of EPO after cerebral ischemia and other brain injuries. The neurotrophic actions suggest there may be longer-latency effects as well. Evaluation of EPO, a compound established as clinically safe, as neuroprotective therapy in acute brain injury is further supported. PMID- 11259644 TI - Solution structure of the A loop of 23S ribosomal RNA. AB - The A loop is an essential RNA component of the ribosome peptidyltransferase center that directly interacts with aminoacyl (A)-site tRNA. The A loop is highly conserved and contains a ubiquitous 2'-O-methyl ribose modification at position U2552. Here, we present the solution structure of a modified and unmodified A loop RNA to define both the A-loop fold and the structural impact of the U2552 modification. Solution data reveal that the A-loop RNA has a compact structure that includes a noncanonical base pair between C2556 and U2552. NMR evidence is presented that the N3 position of C2556 has a shifted pKa and that protonation at C2556-N3 changes the C-U pair geometry. Our data indicate that U2552 methylation modifies the A-loop fold, in particular the dynamics and position of residues C2556 and U2555. We compare our structural data with the structure of the A loop observed in a recent 50S crystal structure [Ban, N., Nissen, P., Hansen, J., Moore, P. B. & Steitz, T. A. (2000) Science 289, 905--920; Nissen, P., Hansen, J., Ban, N., Moore, P. B. & Steitz, T. A. (2000) Science 289, 920--930]. The solution and crystal structures of the A loop are dramatically different, suggesting that a structural rearrangement of the A loop must occur on docking into the peptidyltransferase center. Possible roles of this docking event, the shifted pKa of C2556 and the U2552 2'-O-methylation in the mechanism of translation, are discussed. PMID- 11259645 TI - Nerve growth factor control of neuronal expression of angiogenetic and vasoactive factors. AB - In postnatal tissues, angiogenesis occurs in nontumoral conditions on appropriate stimuli. In the nervous tissue, hypoxia, neural graft, increased neural function, and synaptic activity are associated with neoangiogenesis. We have investigated the occurrence of neoangiogenesis in the superior cervical ganglia (scg) of newborn rats treated for 8--21 days with 6-hydroxy-dopamine (6-OHDA), nerve growth factor (NGF), or 6-OHDA + NGF. The two latter treatments induced a significant increase in scg size. However, the increase after combined treatment far exceeded that of NGF alone. Similarly, histological and histochemical analysis revealed neuronal hypertrophy and endothelial cell hyperplasia associated with stromal hypertrophy (as described by laminin immunostaining) and increased vascular bed (as revealed by platelet/endothelial cell adhesion molecule-1 immunostaining) in 6-OHDA + NGF-treated pups. NGF, either alone or associated with 6-OHDA, also induced a significant up-regulation of NADPH diaphorase, neuronal nitric oxide synthase, and vascular endothelial growth factor expression in scg neurons. The present investigation suggests that the increase of scg size induced by NGF and 6-OHDA + NGF is associated with neoangiogenesis, and that the induction of vasoactive and angiogenic factors in neurons represents a further and previously undisclosed effect of NGF. PMID- 11259646 TI - Microglia at brain stab wounds express connexin 43 and in vitro form functional gap junctions after treatment with interferon-gamma and tumor necrosis factor alpha. AB - Gap junctional communication between microglia was investigated at rat brain stab wounds and in primary cultures of rat and mouse cells. Under resting conditions, rat microglia (FITC-isolectin-B4-reactive cells) were sparsely distributed in the neocortex, and most (95%) were not immunoreactive for Cx43, a gap junction protein subunit. At brain stab wounds, microglia progressively accumulated over several days and formed aggregates that frequently showed Cx43 immunoreactivity at interfaces between cells. In primary culture, microglia showed low levels of Cx43 determined by Western blotting, diffuse intracellular Cx43 immunoreactivity, and a low incidence of dye coupling. Treatment with the immunostimulant bacterial lipopolysaccharide (LPS) or the cytokines interferon-gamma (INF-gamma) or tumor necrosis factor-alpha (TNF-alpha) one at a time did not increase the incidence of dye coupling. However, microglia treated with INF-gamma plus LPS showed a dramatic increase in dye coupling that was prevented by coapplication of an anti TNF-alpha antibody, suggesting the release and autocrine action of TNF-alpha. Treatment with INF-gamma plus TNF-alpha also greatly increased the incidence of dye coupling and the Cx43 levels with translocation of Cx43 to cell-cell contacts. The cytokine-induced dye coupling was reversibly inhibited by 18 alpha glycyrrhetinic acid, a gap junction blocker. Cultured mouse microglia also expressed Cx43 and developed dye coupling upon treatment with cytokines, but microglia from homozygous Cx43-deficient mice did not develop significant dye coupling after treatment with either INF-gamma plus LPS or INF-gamma plus TNF alpha. This report demonstrates that microglia can communicate with each other through gap junctions that are induced by inflammatory cytokines, a process that may be important in the elaboration of the inflammatory response. PMID- 11259647 TI - Complete genome sequence of Caulobacter crescentus. AB - The complete genome sequence of Caulobacter crescentus was determined to be 4,016,942 base pairs in a single circular chromosome encoding 3,767 genes. This organism, which grows in a dilute aquatic environment, coordinates the cell division cycle and multiple cell differentiation events. With the annotated genome sequence, a full description of the genetic network that controls bacterial differentiation, cell growth, and cell cycle progression is within reach. Two-component signal transduction proteins are known to play a significant role in cell cycle progression. Genome analysis revealed that the C. crescentus genome encodes a significantly higher number of these signaling proteins (105) than any bacterial genome sequenced thus far. Another regulatory mechanism involved in cell cycle progression is DNA methylation. The occurrence of the recognition sequence for an essential DNA methylating enzyme that is required for cell cycle regulation is severely limited and shows a bias to intergenic regions. The genome contains multiple clusters of genes encoding proteins essential for survival in a nutrient poor habitat. Included are those involved in chemotaxis, outer membrane channel function, degradation of aromatic ring compounds, and the breakdown of plant-derived carbon sources, in addition to many extracytoplasmic function sigma factors, providing the organism with the ability to respond to a wide range of environmental fluctuations. C. crescentus is, to our knowledge, the first free-living alpha-class proteobacterium to be sequenced and will serve as a foundation for exploring the biology of this group of bacteria, which includes the obligate endosymbiont and human pathogen Rickettsia prowazekii, the plant pathogen Agrobacterium tumefaciens, and the bovine and human pathogen Brucella abortus. PMID- 11259648 TI - 2-arachidonyl glyceryl ether, an endogenous agonist of the cannabinoid CB1 receptor. AB - Two types of endogenous cannabinoid-receptor agonists have been identified thus far. They are the ethanolamides of polyunsaturated fatty acids--arachidonoyl ethanolamide (anandamide) is the best known compound in the amide series--and 2 arachidonoyl glycerol, the only known endocannabinoid in the ester series. We report now an example of a third, ether-type endocannabinoid, 2-arachidonyl glyceryl ether (noladin ether), isolated from porcine brain. The structure of noladin ether was determined by mass spectrometry and nuclear magnetic resonance spectroscopy and was confirmed by comparison with a synthetic sample. It binds to the CB(1) cannabinoid receptor (K(i) = 21.2 +/- 0.5 nM) and causes sedation, hypothermia, intestinal immobility, and mild antinociception in mice. It binds weakly to the CB(2) receptor (K(i) > 3 microM). PMID- 11259649 TI - Suppression of ras-mediated transformation and inhibition of tumor growth and angiogenesis by anthrax lethal factor, a proteolytic inhibitor of multiple MEK pathways. AB - Lethal factor is a protease, one component of Bacillus anthracis exotoxin, which cleaves many of the mitogen-activated protein kinase kinases (MEKs). Given the importance of MEK signaling in tumorigenesis, we assessed the effects of anthrax lethal toxin (LeTx) on tumor cells. LeTx was very effective in inhibiting mitogen activated protein kinase activation in V12 H-ras-transformed NIH 3T3 cells. In vitro, treatment of transformed cells with LeTx caused them to revert to a nontransformed morphology, and inhibited their abilities to form colonies in soft agar and to invade Matrigel without markedly affecting cell proliferation. In vivo, LeTx inhibited growth of ras-transformed cells implanted in athymic nude mice (in some cases causing tumor regression) at concentrations that caused no apparent animal toxicity. Unexpectedly, LeTx also greatly decreased tumor neovascularization. These results demonstrate that LeTx potently inhibits ras mediated tumor growth and is a potential antitumor therapeutic. PMID- 11259650 TI - Ciliary neurotrophic factor activates leptin-like pathways and reduces body fat, without cachexia or rebound weight gain, even in leptin-resistant obesity. AB - Ciliary Neurotrophic Factor (CNTF) was first characterized as a trophic factor for motor neurons in the ciliary ganglion and spinal cord, leading to its evaluation in humans suffering from motor neuron disease. In these trials, CNTF caused unexpected and substantial weight loss, raising concerns that it might produce cachectic-like effects. Countering this possibility was the suggestion that CNTF was working via a leptin-like mechanism to cause weight loss, based on the findings that CNTF acts via receptors that are not only related to leptin receptors, but also similarly distributed within hypothalamic nuclei involved in feeding. However, although CNTF mimics the ability of leptin to cause fat loss in mice that are obese because of genetic deficiency of leptin (ob/ob mice), CNTF is also effective in diet-induced obesity models that are more representative of human obesity, and which are resistant to leptin. This discordance again raised the possibility that CNTF might be acting via nonleptin pathways, perhaps more analogous to those activated by cachectic cytokines. Arguing strongly against this possibility, we now show that CNTF can activate hypothalamic leptin-like pathways in diet-induced obesity models unresponsive to leptin, that CNTF improves prediabetic parameters in these models, and that CNTF acts very differently than the prototypical cachectic cytokine, IL-1. Further analyses of hypothalamic signaling reveals that CNTF can suppress food intake without triggering hunger signals or associated stress responses that are otherwise associated with food deprivation; thus, unlike forced dieting, cessation of CNTF treatment does not result in binge overeating and immediate rebound weight gain. PMID- 11259651 TI - Feeding specialization and host-derived chemical defense in Chrysomeline leaf beetles did not lead to an evolutionary dead end. AB - Combination of molecular phylogenetic analyses of Chrysomelina beetles and chemical data of their defensive secretions indicate that two lineages independently developed, from an ancestral autogenous metabolism, an energetically efficient strategy that made the insect tightly dependent on the chemistry of the host plant. However, a lineage (the interrupta group) escaped this subordination through the development of a yet more derived mixed metabolism potentially compatible with a large number of new host-plant associations. Hence, these analyses on leaf beetles document a mechanism that can explain why high levels of specialization do not necessarily lead to "evolutionary dead ends." PMID- 11259652 TI - Prevention and elimination of upper respiratory colonization of mice by group A streptococci by using a bacteriophage lytic enzyme. AB - Bacteriophage lytic enzymes quickly destroy the cell wall of the host bacterium to release progeny phage. Because such lytic enzymes specifically kill the species in which they were produced, they may represent an effective way to control pathogenic bacteria without disturbing normal microflora. In this report, we studied a murein hydrolase from the streptococcal bacteriophage C(1) termed lysin. This enzyme is specific for groups A, C, and E streptococci, with little or no activity toward several oral streptococci or other commensal organisms tested. Using purified lysin in vitro, we show that 1,000 units (10 ng) of enzyme is sufficient to sterilize a culture of approximately 10(7) group A streptococci within 5 seconds. When a single dose of lysin (250 units) is first added to the oral cavity of mice, followed by 10(7) live group A streptococci, it provides protection from colonization (28.5% infected, n = 21) compared with controls without lysin (70.5% infected, n = 17) (P < 0.03). Furthermore, when lysin (500 units) was given orally to 9 heavily colonized mice, no detectable streptococci were observed 2 h after lysin treatment. In all, these studies show that lysin represents a unique murein hydrolase that has a rapid lethal effect both in vitro and in vivo on group A streptococci, without affecting other indigenous microorganisms analyzed. This general approach may be used to either eliminate or reduce streptococci from the upper respiratory mucosal epithelium of either carriers or infected individuals, thus reducing associated disease. PMID- 11259654 TI - Adenoviral gene transfer of Caenorhabditis elegans n--3 fatty acid desaturase optimizes fatty acid composition in mammalian cells. AB - Omega--3 polyunsaturated fatty acids (PUFAs) are essential components required for normal cellular function and have been shown to exert many preventive and therapeutic actions. The amount of n--3 PUFAs is insufficient in most Western people, whereas the level of n--6 PUFAs is relatively too high, with an n--6/n--3 ratio of >18. These two classes of PUFAs are metabolically and functionally distinct and often have important opposing physiological functions; their balance is important for homeostasis and normal development. Elevating tissue concentrations of n--3 PUFAs in mammals relies on chronic dietary intake of fat rich in n--3 PUFAs, because mammalian cells lack enzymatic activities necessary either to synthesize the precursor of n--3 PUFAs or to convert n--6 to n--3 PUFAs. Here we report that adenovirus-mediated introduction of the Caenorhabditis elegans fat-1 gene encoding an n--3 fatty acid desaturase into mammalian cells can quickly and effectively elevate the cellular n--3 PUFA contents and dramatically balance the ratio of n--6/n--3 PUFAs. Heterologous expression of the fat-1 gene in rat cardiac myocytes rendered cells capable of converting various n -6 PUFAs to the corresponding n--3 PUFAs, and changed the n--6/n--3 ratio from about 15:1 to 1:1. In addition, an eicosanoid derived from n--6 PUFA (i.e., arachidonic acid) was reduced significantly in the transgenic cells. This study demonstrates an effective approach to modifying fatty acid composition of mammalian cells and also provides a basis for potential applications of this gene transfer in experimental and clinical settings. PMID- 11259653 TI - Increased in vivo apoptosis in cells lacking mitochondrial DNA gene expression. AB - We have attempted to determine whether loss of mtDNA and respiratory chain function result in apoptosis in vivo. Apoptosis was studied in embryos with homozygous disruption of the mitochondrial transcription factor A gene (Tfam) and tissue-specific Tfam knockout animals with severe respiratory chain deficiency in the heart. We found massive apoptosis in Tfam knockout embryos at embryonic day (E) 9.5 and increased apoptosis in the heart of the tissue-specific Tfam knockouts. Furthermore, mtDNA-less (rho(0)) cell lines were susceptible to apoptosis induced by different stimuli in vitro. The data presented here provide in vivo evidence that respiratory chain deficiency predisposes cells to apoptosis, contrary to previous assumptions based on in vitro studies of cultured cells. These results suggest that increased apoptosis is a pathogenic event in human mtDNA mutation disorders. The finding that respiratory chain deficiency is associated with increased in vivo apoptosis may have important therapeutic implications for human disease. Respiratory chain deficiency and cell loss and/or apoptosis have been associated with neurodegeneration, heart failure, diabetes mellitus, and aging. Furthermore, chemotherapy and radiation treatment of cancer are intended to induce apoptosis in tumor cells. It would therefore be of interest to determine whether manipulation of respiratory chain function can be used to inhibit or enhance apoptosis in these conditions. PMID- 11259655 TI - Speciation through homoploid hybridization between allotetraploids in peonies (Paeonia). AB - Phylogenies of Adh1 and Adh2 genes suggest that a widespread Mediterranean peony, Paeonia officinalis, is a homoploid hybrid species between two allotetraploid species, Paeonia peregrina and a member of the Paeonia arietina species group. Three phylogenetically distinct types of Adh sequences have been identified from both accessions of P. officinalis, of which two types are most closely related to the two homoeologous Adh loci of the P. arietina group and the remaining type came from one of the two Adh homoeologs of P. peregrina. The other Adh homoeolog of P. peregrina was apparently lost from the hybrid genome, possibly through backcrossing with the P. arietina group. This is a documentation of homoploid hybrid speciation between allotetraploid species in nature. This study suggests that hybrid speciation between allotetraploids can occur without an intermediate stage of genome diploidization or a further doubling of genome size. PMID- 11259656 TI - Ex vivo propagation of infectious sheep scrapie agent in heterologous epithelial cells expressing ovine prion protein. AB - Transmissible spongiform encephalopathies, or prion diseases, are fatal degenerative disorders of the central nervous system that affect humans and animals. Prions are nonconventional infectious agents whose replication depends on the host prion protein (PrP). Transmission of prions to cultured cells has proved to be a particularly difficult task, and with a few exceptions, their experimental propagation relies on inoculation to laboratory animals. Here, we report on the development of a permanent cell line supporting propagation of natural sheep scrapie. This model was obtained by stable expression of a tetracycline-regulatable ovine PrP gene in a rabbit epithelial cell line. After exposure to scrapie agent, cultures were repeatedly found to accumulate high levels of abnormal PrP (PrPres). Cell extracts induced a scrapie-like disease in transgenic mice overexpressing ovine PrP. These cultures remained healthy and stably infected upon subpassaging. Such data show that (i) cultivated cells from a nonneuronal origin can efficiently replicate prions; and (ii) species barrier can be crossed ex vivo through the expression of a relevant PrP gene. This approach led to the ex vivo propagation of a natural transmissible spongiform encephalopathy agent (i.e., without previous experimental adaptation to rodents) and might be applied to human or bovine prions. PMID- 11259658 TI - The nucleotide changes governing cuticular hydrocarbon variation and their evolution in Drosophila melanogaster. AB - The cuticular hydrocarbon (CH) pheromones in Drosophila melanogaster exhibit strong geographic variation. African and Caribbean populations have a high ratio of 5,9 heptacosadiene/7,11 heptacosadiene (the "High" CH type), whereas populations from all other areas have a low ratio ("Low" CH type). Based on previous genetic mapping, DNA markers were developed that localized the genetic basis of this CH polymorphism to within a 13-kb region. We then carried out a hierarchical search for diagnostic nucleotide sites starting with four lines, and increasing to 24 and 43 lines from a worldwide collection. Within the 13-kb region, only one variable site shows a complete concordance with the CH phenotype. This is a 16-bp deletion in the 5' region of a desaturase gene (desat2) that was recently suggested to be responsible for the CH polymorphism on the basis of its expression [Dallerac, R., Labeur, C., Jallon, J.-M., Knipple, D. C., Roelofs, W. L. & Wicker-Thomas, C. (2000) Proc. Natl. Acad. Sci. 97, 9449- 9454]. The cosmopolitan Low type is derived from the ancestral High type, and DNA sequence variations suggest that the former spread worldwide with the aid of positive selection. Whether this CH variation could be a component of the sexual isolation between Zimbabwe and other cosmopolitan populations remains an interesting and unresolved question. PMID- 11259659 TI - CD4(+) T cell recognition of MHC class II-restricted epitopes from NY-ESO-1 presented by a prevalent HLA DP4 allele: association with NY-ESO-1 antibody production. AB - NY-ESO-1 is a tumor-specific shared antigen with distinctive immunogenicity. Both CD8(+) T cells and class-switched Ab responses have been detected from patients with cancer. In this study, a CD4(+) T cell line was generated from peripheral blood mononuclear cells of a melanoma patient and was shown to recognize NY-ESO-1 peptides presented by HLA-DP4, a dominant MHC class II allele expressed in 43- 70% of Caucasians. The ESO p157--170 peptide containing the core region of DP4 restricted T cell epitope was present in a number of tumor cell lines tested and found to be recognized by both CD4(+) T cells as well as HLA-A2-restricted CD8(+) T cells. Thus, the ESO p157--170 epitope represents a potential candidate for cancer vaccines aimed at generating both CD4(+) and CD8(+) T cell responses. More importantly, 16 of 17 melanoma patients who developed Ab against NY-ESO-1 were found to be HLA-DP4-positive. CD4(+) T cells specific for the NY-ESO-1 epitopes were generated from 5 of 6 melanoma patients with NY-ESO-1 Ab. In contrast, no specific DP4-restricted T cells were generated from two patients without detectable NY-ESO-1 Ab. These results suggested that NY-ESO-1-specific DP4 restricted CD4(+) T cells were closely associated with NY-ESO-1 Ab observed in melanoma patients and might play an important role in providing help for activating B cells for NY-ESO-1-specific Ab production. PMID- 11259657 TI - Galanin transgenic mice display cognitive and neurochemical deficits characteristic of Alzheimer's disease. AB - Galanin is a neuropeptide with multiple inhibitory actions on neurotransmission and memory. In Alzheimer's disease (AD), increased galanin-containing fibers hyperinnervate cholinergic neurons within the basal forebrain in association with a decline in cognition. We generated transgenic mice (GAL-tg) that overexpress galanin under the control of the dopamine beta-hydroxylase promoter to study the neurochemical and behavioral sequelae of a mouse model of galanin overexpression in AD. Overexpression of galanin was associated with a reduction in the number of identifiable neurons producing acetylcholine in the horizontal limb of the diagonal band. Behavioral phenotyping indicated that GAL-tgs displayed normal general health and sensory and motor abilities; however, GAL-tg mice showed selective performance deficits on the Morris spatial navigational task and the social transmission of food preference olfactory memory test. These results suggest that elevated expression of galanin contributes to the neurochemical and cognitive impairments characteristic of AD. PMID- 11259660 TI - SQV-7, a protein involved in Caenorhabditis elegans epithelial invagination and early embryogenesis, transports UDP-glucuronic acid, UDP-N- acetylgalactosamine, and UDP-galactose. AB - Caenorhabditis elegans sqv mutants are defective in vulval epithelial invagination and have a severe reduction in hermaphrodite fertility. The gene sqv 7 encodes a multitransmembrane hydrophobic protein resembling nucleotide sugar transporters of the Golgi membrane. A Golgi vesicle enriched fraction of Saccharomyces cerevisiae expressing SQV-7 transported UDP-glucuronic acid, UDP-N acetylgalactosamine, and UDP-galactose (Gal) in a temperature-dependent and saturable manner. These nucleotide sugars are competitive, alternate, noncooperative substrates. The two mutant sqv-7 missense alleles resulted in a severe reduction of these three transport activities. SQV-7 did not transport CMP sialic acid, GDP-fucose, UDP-N-acetylglucosamine, UDP-glucose, or GDP-mannose. SQV-7 is able to transport UDP-Gal in vivo, as shown by its ability to complement the phenotype of Madin-Darby canine kidney ricin resistant cells, a mammalian cell line deficient in UDP-Gal transport into the Golgi. These results demonstrate that unlike most nucleotide sugar transporters, SQV-7 can transport multiple distinct nucleotide sugars. We propose that SQV-7 translocates multiple nucleotide sugars into the Golgi lumen for the biosynthesis of glycoconjugates that play a pivotal role in development. PMID- 11259661 TI - The Sm domain is an ancient RNA-binding motif with oligo(U) specificity. AB - Sm and Sm-like proteins are members of a family of small proteins that is widespread throughout eukaryotic kingdoms. These proteins form heteromers with one another and bind, as heteromeric complexes, to various RNAs, recognizing primarily short U-rich stretches. Interestingly, completion of several genome projects revealed that archaea also contain genes that may encode Sm-like proteins. Herein, we studied the properties of one Sm-like protein derived from the archaebacterium Archaeoglobus fulgidus and overexpressed in Escherichia coli. This single small protein closely reflects the properties of an Sm or Sm-like protein heteromer. It binds to RNA with a high specificity for oligo(U), and assembles onto the RNA to form a complex that exhibits, as judged by electron microscopy, a ring-like structure similar to the ones observed with the Sm core ribonucleoprotein and the like Sm (LSm) protein heteromer. Importantly, multivariate statistical analysis of negative-stain electron-microscopic images revealed a sevenfold symmetry for the observed ring structure, indicating that the proteins form a homoheptamer. These results support the structural model of the Sm proteins derived from crystallographic studies on Sm heterodimers and demonstrate that the Sm protein family evolved from a single ancestor that was present before the eukaryotic and archaeal kingdoms separated. PMID- 11259662 TI - Medial prefrontal cortex and self-referential mental activity: relation to a default mode of brain function. AB - Medial prefrontal cortex (MPFC) is among those brain regions having the highest baseline metabolic activity at rest and one that exhibits decreases from this baseline across a wide variety of goal-directed behaviors in functional imaging studies. This high metabolic rate and this behavior suggest the existence of an organized mode of default brain function, elements of which may be either attenuated or enhanced. Extant data suggest that these MPFC regions may contribute to the neural instantiation of aspects of the multifaceted "self." We explore this important concept by targeting and manipulating elements of MPFC default state activity. In this functional magnetic resonance imaging (fMRI) study, subjects made two judgments, one self-referential, the other not, in response to affectively normed pictures: pleasant vs. unpleasant (an internally cued condition, ICC) and indoors vs. outdoors (an externally cued condition, ECC). The ICC was preferentially associated with activity increases along the dorsal MPFC. These increases were accompanied by decreases in both active task conditions in ventral MPFC. These results support the view that dorsal and ventral MPFC are differentially influenced by attentiondemanding tasks and explicitly self-referential tasks. The presence of self-referential mental activity appears to be associated with increases from the baseline in dorsal MPFC. Reductions in ventral MPFC occurred consistent with the fact that attention demanding tasks attenuate emotional processing. We posit that both self referential mental activity and emotional processing represent elements of the default state as represented by activity in MPFC. We suggest that a useful way to explore the neurobiology of the self is to explore the nature of default state activity. PMID- 11259664 TI - Pectate lyase A, an enzymatic subunit of the Clostridium cellulovorans cellulosome. AB - Clostridium cellulovorans uses not only cellulose but also xylan, mannan, pectin, and several other carbon sources for its growth and produces an extracellular multienzyme complex called the cellulosome, which is involved in plant cell wall degradation. Here we report a gene for a cellulosomal subunit, pectate lyase A (PelA), lying downstream of the engY gene, which codes for cellulosomal enzyme EngY. pelA is composed of an ORF of 2,742 bp and encodes a protein of 914 aa with a molecular weight of 94,458. The amino acid sequence derived from pelA revealed a multidomain structure, i.e., an N-terminal domain partially homologous to the C terminus of PelB of Erwinia chrysanthemi belonging to family 1 of pectate lyases, a putative cellulose-binding domain, a catalytic domain homologous to PelL and PelX of E. chrysanthemi that belongs to family 4 of pectate lyases, and a duplicated sequence (or dockerin) at the C terminus that is highly conserved in enzymatic subunits of the C. cellulovorans cellulosome. The recombinant truncated enzyme cleaved polygalacturonic acid to digalacturonic acid (G2) and trigalacturonic acid (G3) but did not act on G2 and G3. There have been no reports available to date on pectate lyase genes from Clostridia. PMID- 11259663 TI - ClpA mediates directional translocation of substrate proteins into the ClpP protease. AB - The intracellular degradation of many proteins is mediated in an ATP-dependent manner by large assemblies comprising a chaperone ring complex associated coaxially with a proteolytic cylinder, e.g., ClpAP, ClpXP, and HslUV in prokaryotes, and the 26S proteasome in eukaryotes. Recent studies of the chaperone ClpA indicate that it mediates ATP-dependent unfolding of substrate proteins and directs their ATP-dependent translocation into the ClpP protease. Because the axial passageway into the proteolytic chamber is narrow, it seems likely that unfolded substrate proteins are threaded from the chaperone into the protease, suggesting that translocation could be directional. We have investigated directionality in the ClpA/ClpP-mediated reaction by using two substrate proteins bearing the COOH-terminal ssrA recognition element, each labeled near the NH(2) or COOH terminus with fluorescent probes. Time-dependent changes in both fluorescence anisotropy and fluorescence resonance energy transfer between donor fluorophores in the ClpP cavity and the substrate probes as acceptors were measured to monitor translocation of the substrates from ClpA into ClpP. We observed for both substrates that energy transfer occurs 2--4 s sooner with the COOH-terminally labeled molecules than with the NH(2)-terminally labeled ones, indicating that translocation is indeed directional, with the COOH terminus of the substrate protein entering ClpP first. PMID- 11259665 TI - Initiation of clement surface conditions on the earliest Earth. AB - In the beginning the surface of the Earth was extremely hot, because the Earth as we know it is the product of a collision between two planets, a collision that also created the Moon. Most of the heat within the very young Earth was lost quickly to space while the surface was still quite hot. As it cooled, the Earth's surface passed monotonically through every temperature regime between silicate vapor to liquid water and perhaps even to ice, eventually reaching an equilibrium with sunlight. Inevitably the surface passed through a time when the temperature was around 100 degrees C at which modern thermophile organisms live. How long this warm epoch lasted depends on how long a thick greenhouse atmosphere can be maintained by heat flow from the Earth's interior, either directly as a supplement to insolation, or indirectly through its influence on the nascent carbonate cycle. In both cases, the duration of the warm epoch would have been controlled by processes within the Earth's interior where buffering by surface conditions played little part. A potentially evolutionarily significant warm period of between 10(5) and 10(7) years seems likely, which nonetheless was brief compared to the vast expanse of geological time. PMID- 11259666 TI - Peptide hybrids containing alpha - and beta-amino acids: structure of a decapeptide beta-hairpin with two facing beta-phenylalanine residues. AB - A beta-hairpin conformation has been characterized in crystals of the decapeptide t-butoxycarbonyl-Leu-Val-beta Phe-Val-(D)Pro-Gly-Leu-beta Phe-Val-Val-methyl ester [beta Phe; (S)-beta(3) homophenylalanine] by x-ray diffraction. The polypeptide chain reversal is nucleated by the centrally positioned (D)Pro-Gly segment, which adopts a type-I' beta-turn conformation. Four intramolecular cross strand hydrogen bonds stabilize the peptide fold. The beta Phe(3) and beta Phe(8) residues occupy facing positions on the hairpin, with the side chains projecting on opposite faces of the beta-sheet. At the site of insertion of beta-residues, the polarity of the peptide units along each strand reverses, as compared with the alpha-peptide segments. In this analog, a small segment of a polar sheet is observed, where adjacent CO and NH groups line up in opposite directions in each strand. In the crystal, an extended beta-sheet is formed by hydrogen bonding between strands of antiparallel pairs of beta-hairpins. The crystallographic parameters for C(65)H(102)N(10)O(13) x 3H(2)O are: space group P2(1)2(1)2(1); a = 19.059(8) A, b = 19.470(2) A, c = 21.077(2) A; Z = 4; agreement factor R(1) = 9.12% for 3,984 data observed >4 sigma(F) and a resolution of 0.90 A. PMID- 11259668 TI - Visual constraints in foraging bumblebees: flower size and color affect search time and flight behavior. AB - In optimal foraging theory, search time is a key variable defining the value of a prey type. But the sensory-perceptual processes that constrain the search for food have rarely been considered. Here we evaluate the flight behavior of bumblebees (Bombus terrestris) searching for artificial flowers of various sizes and colors. When flowers were large, search times correlated well with the color contrast of the targets with their green foliage-type background, as predicted by a model of color opponent coding using inputs from the bees' UV, blue, and green receptors. Targets that made poor color contrast with their backdrop, such as white, UV-reflecting ones, or red flowers, took longest to detect, even though brightness contrast with the background was pronounced. When searching for small targets, bees changed their strategy in several ways. They flew significantly slower and closer to the ground, so increasing the minimum detectable area subtended by an object on the ground. In addition, they used a different neuronal channel for flower detection. Instead of color contrast, they used only the green receptor signal for detection. We relate these findings to temporal and spatial limitations of different neuronal channels involved in stimulus detection and recognition. Thus, foraging speed may not be limited only by factors such as prey density, flight energetics, and scramble competition. Our results show that understanding the behavioral ecology of foraging can substantially gain from knowledge about mechanisms of visual information processing. PMID- 11259667 TI - The GABAergic reticular nucleus: a preferential target of corticothalamic projections. PMID- 11259669 TI - Integrated control of appetite and fat metabolism by the leptin proopiomelanocortin pathway. AB - Leptin deficiency results in a complex obesity phenotype comprising both hyperphagia and lowered metabolism. The hyperphagia results, at least in part, from the absence of induction by leptin of melanocyte stimulating hormone (MSH) secretion in the hypothalamus; the MSH normally then binds to melanocortin-4 receptor expressing neurons and inhibits food intake. The basis for the reduced metabolic rate has been unknown. Here we show that leptin administered to leptin deficient (ob/ob) mice results in a large increase in peripheral MSH levels; further, peripheral administration of an MSH analogue results in a reversal of their abnormally low metabolic rate, in an acceleration of weight loss during a fast, in partial restoration of thermoregulation in a cold challenge, and in inducing serum free fatty acid levels. These results support an important peripheral role for MSH in the integration of metabolism with appetite in response to perceived fat stores indicated by leptin levels. PMID- 11259671 TI - Role of the arginine-nitric oxide pathway in the regulation of vascular smooth muscle cell proliferation. AB - The objective of this study was to elucidate the mechanisms by which nitric oxide (NO) inhibits rat aortic smooth muscle cell (RASMC) proliferation. Two products of the arginine-NO pathway interfere with cell growth by distinct mechanisms. N(G)-hydroxyarginine and NO appear to interfere with cell proliferation by inhibiting arginase and ornithine decarboxylase (ODC), respectively. S-nitroso-N acetylpenicillamine, (Z)-1-[N-(2-aminoethyl)-N-(2-aminoethyl)-amino]-diazen-1-ium 1,2-diolate, and a nitroaspirin derivative (NCX 4016), each of which is a NO donor agent, inhibited RASMC growth at concentrations of 1-3 microM by cGMP independent mechanisms. The cytostatic action of the NO donor agents as well as alpha-difluoromethylornithine (DFMO), a known ODC inhibitor, was prevented by addition of putrescine but not ornithine. These observations suggested that NO, like DFMO, may directly inhibit ODC. Experiments with purified, recombinant mammalian ODC revealed that NO inhibits ODC possibly by S-nitrosylation of the active site cysteine in ODC. DFMO, as well as the NO donor agents, interfered with cellular polyamine (putrescine, spermidine, spermine) production. Conversely, increasing the expression and catalytic activity of arginase I in RASMC either by transfection of cells with the arginase I gene or by induction of arginase I mRNA with IL-4 resulted in increased urea and polyamine production as well as cell proliferation. Finally, coculture of rat aortic endothelial cells, which had been pretreated with lipopolysaccharide plus a cytokine mixture to induce NO synthase and promote NO production, caused NO-dependent inhibition of target RASMC proliferation. This study confirms the inhibitory role of the arginine-NO pathway in vascular smooth muscle proliferation and indicates that one mechanism of action of NO is cGMP-independent and attributed to its capacity to inhibit ODC. PMID- 11259670 TI - Insulin inhibits transcription of IRS-2 gene in rat liver through an insulin response element (IRE) that resembles IREs of other insulin-repressed genes. AB - Recent data indicate that sustained elevations in plasma insulin suppress the mRNA for IRS-2, a component of the insulin signaling pathway in liver, and that this deficiency contributes to hepatic insulin resistance and inappropriate gluconeogenesis. Here, we use nuclear run-on assays to show that insulin inhibits transcription of the IRS-2 gene in the livers of intact rats. Insulin also inhibited transcription of a reporter gene driven by the human IRS-2 promoter that was transfected into freshly isolated rat hepatocytes. The human promoter contains a heptanucleotide sequence, TGTTTTG, that is identical to the insulin response element (IRE) identified previously in the promoters of insulin repressed genes. Single base pair substitutions in this IRE decreased transcription of the IRS-2-driven reporter in the absence of insulin and abolished insulin-mediated repression. We conclude that insulin represses transcription of the IRS-2 gene by blocking the action of a positive factor that binds to the IRE. Sustained repression of IRS-2, as occurs in chronic hyperinsulinemia, contributes to hepatic insulin resistance and accelerates the development of the diabetic state. PMID- 11259672 TI - The human brm protein is cleaved during apoptosis: the role of cathepsin G. AB - The human brm (hbrm) protein (homologue of the Drosophila melanogaster brahma and Saccharomyces cervisiae SNF-2 proteins) is part of a polypeptide complex believed to regulate chromatin conformation. We have shown that the hbrm protein is cleaved in NB4 leukemic cells after induction of apoptosis by UV-irradiation, DNA damaging agents, or staurosporine. Because hbrm is found only in the nucleus, we have investigated the nature of the proteases that may regulate the degradation of this protein during apoptosis. In an in vitro assay, the hbrm protein could not be cleaved by caspase-3, -7, or -6, the "effector" caspases generally believed to carry out the cleavage of nuclear protein substrates. In contrast, we find that cathepsin G, a granule enzyme found in NB4 cells, cleaves hbrm in a pattern similar to that observed in vivo during apoptosis. In addition, a peptide inhibitor of cathepsin G blocks hbrm cleavage during apoptosis but does not block activation of caspases or cleavage of the nuclear protein polyADP ribose polymerase (PARP). Although localized in granules and in the Golgi complex in untreated cells, cathepsin G becomes diffusely distributed during apoptosis. Cleavage by cathepsin G removes a 20-kDa fragment containing a bromodomain from the carboxyl terminus of hbrm. This cleavage disrupts the association between hbrm and the nuclear matrix; the 160-kDa hbrm cleavage fragment is less tightly associated with the nuclear matrix than full-length hbrm. PMID- 11259673 TI - An impairment in sniffing contributes to the olfactory impairment in Parkinson's disease. AB - Although the presence of an olfactory impairment in Parkinson's disease (PD) has been recognized for 25 years, its cause remains unclear. Here we suggest a contributing factor to this impairment, namely, that PD impairs active sniffing of odorants. We tested 10 men and 10 women with clinically typical PD, and 20 age and gender-matched healthy controls, in four olfactory tasks: (i) the University of Pennsylvania smell identification test; (ii and iii) detection threshold tests for the odorants vanillin and propionic acid; and (iv) a two-alternative forced choice detection paradigm during which sniff parameters (airflow peak rate, mean rate, volume, and duration) were recorded with a pneomatotachograph-coupled spirometer. An additional experiment tested the effect of intentionally increasing sniff vigor on olfactory performance in 20 additional patients. PD patients were significantly impaired in olfactory identification (P < 0.0001) and detection (P < 0.007). As predicted, PD patients were also significantly impaired at sniffing, demonstrating significantly reduced sniff airflow rate (P < 0.01) and volume (P < 0.002). Furthermore, a patient's ability to sniff predicted his or her performance on olfactory tasks, i.e., the more poorly patients sniffed, the worse their performance on olfaction tests (P < 0.009). Finally, increasing sniff vigor improved olfactory performance in those patients whose baseline performance had been poorest (P < 0.05). These findings implicate a sniffing impairment as a component of the olfactory impairment in PD and further depict sniffing as an important component of human olfaction. PMID- 11259674 TI - The contribution of trait-mediated indirect effects to the net effects of a predator. AB - Many prey modify traits in response to predation risk and this modification of traits can influence the prey's resource acquisition rate. A predator thus can have a "nonlethal" impact on prey that can lead to indirect effects on other community members. Such indirect interactions are termed trait-mediated indirect interactions because they arise from a predator's influence on prey traits, rather than prey density. Because such nonlethal predator effects are immediate, can influence the entire prey population, and can occur over the entire prey lifetime, we argue that nonlethal predator effects are likely to contribute strongly to the net indirect effects of predators (i.e., nonlethal effects may be comparable in magnitude to those resulting from killing prey). This prediction was supported by an experiment in which the indirect effects of a larval dragonfly (Anax sp.) predator on large bullfrog tadpoles (Rana catesbeiana), through nonlethal effects on competing small bullfrog tadpoles, were large relative to indirect effects caused by density reduction of the small tadpoles (the lethal effect). Treatments in which lethal and nonlethal effects of Anax were manipulated independently indicated that this result was robust for a large range of different combinations of lethal and nonlethal effects. Because many, if not most, prey modify traits in response to predators, our results suggest that the magnitude of interaction coefficients between two species may often be dynamically related to changes in other community members, and that many indirect effects previously attributed to the lethal effects of predators may instead be due to shifts in traits of surviving prey. PMID- 11259675 TI - Dwarfism and early death in mice lacking C-type natriuretic peptide. AB - Longitudinal bone growth is determined by endochondral ossification that occurs as chondrocytes in the cartilaginous growth plate undergo proliferation, hypertrophy, cell death, and osteoblastic replacement. The natriuretic peptide family consists of three structurally related endogenous ligands, atrial, brain, and C-type natriuretic peptides (ANP, BNP, and CNP), and is thought to be involved in a variety of homeostatic processes. To investigate the physiological significance of CNP in vivo, we generated mice with targeted disruption of CNP (Nppc(-/-) mice). The Nppc(-/-) mice show severe dwarfism as a result of impaired endochondral ossification. They are all viable perinatally, but less than half can survive during postnatal development. The skeletal phenotypes are histologically similar to those seen in patients with achondroplasia, the most common genetic form of human dwarfism. Targeted expression of CNP in the growth plate chondrocytes can rescue the skeletal defect of Nppc(-/-) mice and allow their prolonged survival. This study demonstrates that CNP acts locally as a positive regulator of endochondral ossification in vivo and suggests its pathophysiological and therapeutic implication in some forms of skeletal dysplasia. PMID- 11259676 TI - A signature of the T ---> R transition in human hemoglobin. AB - Allosteric effects in hemoglobin arise from the equilibrium between at least two energetic states of the molecule: a tense state, T, and a relaxed state, R. The two states differ from each other in the number and energy of the interactions between hemoglobin subunits. In the T state, constraints between subunits oppose the structural changes resulting from ligand binding. In the R state, these constraints are released, thus enhancing ligand-binding affinity. In the present work, we report the presence of four sites in hemoglobin that are structurally stabilized in the R relative to the T state. These sites are His alpha 103(G10) and His alpha 122(H5) in each alpha subunit of hemoglobin. They are located at the alpha(1)beta(1) and alpha(2)beta(2) interfaces of the hemoglobin tetramer, where the histidine side chains form hydrogen bonds with specific residues from the beta chains. We have measured the solvent exchange rates of side chain protons of His alpha 103(G10) and His alpha 122(H5) in both deoxygenated and ligated hemoglobin by NMR spectroscopy. The exchange rates were found to be higher in the deoxygenated-T than in ligated-R state. Analysis of exchange rates in terms of the local unfolding model revealed that the structural stabilization free energy at each of these two histidines is larger by approximately 1.5 kcal/(mol tetramer) in the R relative to the T state. The location of these histidines at the intradimeric alpha(1)beta(1) and alpha(2)beta(2) interfaces also suggests a role for these interfaces in the allosteric equilibrium of hemoglobin. PMID- 11259677 TI - The Notch ligand Jagged1 is required for inner ear sensory development. AB - Within the mammalian inner ear there are six separate sensory regions that subserve the functions of hearing and balance, although how these sensory regions become specified remains unknown. Each sensory region is populated by two cell types, the mechanosensory hair cell and the supporting cell, which are arranged in a mosaic in which each hair cell is surrounded by supporting cells. The proposed mechanism for creating the sensory mosaic is lateral inhibition mediated by the Notch signaling pathway. However, one of the Notch ligands, Jagged1 (Jag1), does not show an expression pattern wholly consistent with a role in lateral inhibition, as it marks the sensory patches from very early in their development--presumably long before cells make their final fate decisions. It has been proposed that Jag1 has a role in specifying sensory versus nonsensory epithelium within the ear [Adam, J., Myat, A., Roux, I. L., Eddison, M., Henrique, D., Ish-Horowicz, D. & Lewis, J. (1998) Development (Cambridge, U.K.) 125, 4645--4654]. Here we provide experimental evidence that Notch signaling may be involved in specifying sensory regions by showing that a dominant mouse mutant headturner (Htu) contains a missense mutation in the Jag1 gene and displays missing posterior and sometimes anterior ampullae, structures that house the sensory cristae. Htu/+ mutants also demonstrate a significant reduction in the numbers of outer hair cells in the organ of Corti. Because lateral inhibition mediated by Notch predicts that disruptions in this pathway would lead to an increase in hair cells, we believe these data indicate an earlier role for Notch within the inner ear. PMID- 11259678 TI - Forming classes by stimulus frequency: behavior and theory. AB - Visual classification is the way we relate to different images in our environment as if they were the same, while relating differently to other collections of stimuli (e.g., human vs. animal faces). It is still not clear, however, how the brain forms such classes, especially when introduced with new or changing environments. To isolate a perception-based mechanism underlying class representation, we studied unsupervised classification of an incoming stream of simple images. Classification patterns were clearly affected by stimulus frequency distribution, although subjects were unaware of this distribution. There was a common bias to locate class centers near the most frequent stimuli and their boundaries near the least frequent stimuli. Responses were also faster for more frequent stimuli. Using a minimal, biologically based neural-network model, we demonstrate that a simple, self-organizing representation mechanism based on overlapping tuning curves and slow Hebbian learning suffices to ensure classification. Combined behavioral and theoretical results predict large tuning overlap, implicating posterior infero-temporal cortex as a possible site of classification. PMID- 11259679 TI - A novel site of antibiotic action in the ribosome: interaction of evernimicin with the large ribosomal subunit. AB - Evernimicin (Evn), an oligosaccharide antibiotic, interacts with the large ribosomal subunit and inhibits bacterial protein synthesis. RNA probing demonstrated that the drug protects a specific set of nucleotides in the loops of hairpins 89 and 91 of 23S rRNA in bacterial and archaeal ribosomes. Spontaneous Evn-resistant mutants of Halobacterium halobium contained mutations in hairpins 89 and 91 of 23S rRNA. In the ribosome tertiary structure, rRNA residues involved in interaction with the drug form a tight cluster that delineates the drug binding site. Resistance mutations in the bacterial ribosomal protein L16, which is shown to be homologous to archaeal protein L10e, cluster to the same region as the rRNA mutations. The Evn-binding site overlaps with the binding site of initiation factor 2. Evn inhibits activity of initiation factor 2 in vitro, suggesting that the drug interferes with formation of the 70S initiation complex. The site of Evn binding and its mode of action are distinct from other ribosome targeted antibiotics. This antibiotic target site can potentially be used for the development of new antibacterial drugs. PMID- 11259680 TI - Nicotinic cholinergic signaling in hippocampal astrocytes involves calcium induced calcium release from intracellular stores. AB - In this report we provide evidence that neuronal nicotinic acetylcholine receptors (nAChRs) are present on hippocampal astrocytes and their activation produces rapid currents and calcium transients. Our data indicate that these responses obtained from astrocytes are primarily mediated by an AChR subtype that is functionally blocked by alpha-bungarotoxin (alpha Bgt) and contains the alpha7 subunit (alpha Bgt-AChRs). Furthermore, their action is unusual in that they effectively increase intracellular free calcium concentrations by activating calcium-induced calcium release from intracellular stores, triggered by influx through the receptor channels. These results reveal a mechanism by which alpha Bgt-AChRs on astrocytes can efficiently modulate calcium signaling in the central nervous system in a manner distinct from that observed with these receptors on neurons. PMID- 11259681 TI - Evidence that ultraviolet markings are associated with patterns of molecular gene flow. AB - Recent studies have shown UV vision and markings to be important in vertebrates, particularly birds, where behavioral experiments have demonstrated its potential importance in sexual selection. However, there has been no genetic evidence that UV markings determine patterns of evolution among natural populations. Here we report molecular evidence that UV markings are associated with the pattern of gene flow in the Tenerife lizard (Gallotia galloti). This species has vicariance induced, approximate east--west lineages in Tenerife closely congruent with the primary lineages of the sympatric gecko species. Against expectations, these molecular phylogeographic lineages (representing geological history) and isolation-by-distance do not appear to influence gene flow. Sexually mature males from populations either side of a latitudinal ecotone have different UV markings and gene flow appears to be linked to this difference in UV markings. It may be that these groups with different UV sexual markings mate assortatively, restricting the gene flow between them. This has implications for debate on the relative importance of vicariance and biotopes in influencing biodiversity, with this evidence supporting the latter. PMID- 11259682 TI - Electrostatic surface plasmon resonance: direct electric field-induced hybridization and denaturation in monolayer nucleic acid films and label-free discrimination of base mismatches. AB - We demonstrate that in situ optical surface plasmon resonance spectroscopy can be used to monitor hybridization kinetics for unlabeled DNA in tethered monolayer nucleic acid films on gold in the presence of an applied electrostatic field. The dc field can enhance or retard hybridization and can also denature surface immobilized DNA duplexes. Discrimination between matched and mismatched hybrids is achieved by simple adjustment of the electrode potential. Although the electric field at the interface is extremely large, the tethered single-stranded DNA thiol probes remain bound and can be reused for subsequent hybridization reactions without loss of efficiency. Only capacitive charging currents are drawn; redox reactions are avoided by maintaining the gold electrode potential within the ideally polarizable region. Because of potential-induced changes in the shape of the surface plasmon resonance curve, we account for the full curve rather than simply the shift in the resonance minimum. PMID- 11259685 TI - Our poisoned patients. PMID- 11259684 TI - Role of the cytoplasmic tyrosines of beta 1A integrins in transformation by v src. AB - GD25 cells lacking beta 1 integrins or expressing beta 1A with mutations of conserved cytoplasmic tyrosines (Y783, Y795) to phenylalanine have poor directed migration to platelet-derived growth factor or lysophosphatidic acid when compared with GD25 cells expressing wild-type beta 1A. We studied the effects of v-src on these cells. Transformation with v-src caused tyrosine and serine phosphorylation of wild-type beta1 A but not of Y783/795F doubly mutated beta 1A. v-src-transformed cells had rounded and/or fusiform morphology and poor assembly of fibronectin matrix. Adhesion to fibronectin or laminin and coupling of focal contacts to actin-containing cytoskeleton were preserved in transformed Y783/795F cells but lost on transformation when beta 1A was wild type. Transformed Y783/795F cells also retained ability, albeit limited, to migrate across filters, whereas transformed cells with wild-type beta 1A were unable to transverse filters. Studies of single tyrosine mutants showed that the more important tyrosine for retaining ability to adhere, assemble focal contacts, and migrate is Y783. These results suggest that overactive phosphorylation of cytoplasmic residues of beta 1A, particularly Y783, accounts in part for the phenotype of v src-transformed cells. PMID- 11259686 TI - Growth failure and intestinal inflammation. PMID- 11259687 TI - Age and gender bias in statin trials. AB - Cardiovascular disease is strongly age-related, and is the leading cause of death in older people. Several well-publicized trials have recently reported that statin drugs (HMG CoA reductase inhibitors) are effective in lowering cholesterol and in reducing the risk of myocardial infarction and stroke. In order to determine whether the results of these trials are relevant to our ageing population, we examined the representation of older people and women in randomized controlled trials of statin drugs. A systematic search of the medical literature from 1990 to 1999 was done to identify randomized placebo-controlled trials of statin drugs which evaluated clinical end-points-myocardial infarction, stroke or death. We identified 19 trials: 15 secondary prevention and four primary prevention. The mean age, age range and gender of the participants in these trials were determined. In the secondary prevention trials, the total number of patients randomized was 31683, with a combined mean age of 58.1 years. No trial enrolled people beyond the age of 75 years, and only 23% of the trial population was female. The four primary prevention trials randomized a combined total of 14 557 subjects with a mean age of 56.9 years. Only 10% of study participants were female. Statin drug trials have suffered from age and gender bias, having been mainly conducted in middle-aged male populations. The extrapolation of evidence from these trials to older people and women needs further evaluation. PMID- 11259688 TI - Hypokalaemia and paralysis. AB - It is not uncommon for patients to present to the emergency room with severe weakness and a markedly low plasma potassium concentration. We attempted to identify useful clues to the diagnosis of hypokalaemic periodic paralysis (HPP), because its acute treatment aims are unique. We retrospectively reviewed charts over a 10-year period: HPP was the initial diagnosis in 97 patients. Mean patient age was 29+/-1.1 and the male:female ratio was 77:20. When the final diagnosis was HPP (n=73), the acid-base state was normal, the urine K(+) concentration was low, and the transtubular K(+) concentration gradient (TTKG) was <3. In patients with thyrotoxic periodic paralysis (TPP) (n=39), hypokalaemia was very commonly accompanied by hypophosphataemia (1.9+/-0.1 mg/dl). A clinical diagnosis of sporadic periodic paralysis (SPP) was made if hyperthyroidism and a family history of HPP were both absent (n=29). One subgroup of patients with HPP had a severe degree of hypernatraemia (167+/-5.0 mmol/l, n=3). There were only two patients with familial periodic paralysis (FPP). In 24 patients, the initial diagnosis was HPP, but subsequent studies failed to confirm this diagnosis. Each of these patients had an acid-base disorder, a high rate of renal K(+) excretion in the presence of hypokalaemia, and a TTKG of close to 7. With respect to therapy, much less K(+) was given to patients with HPP, yet 1:3 subsequently had a plasma K(+) concentration that eventually exceeded 5.0 mmol/l. Using plasma acid-base status, phosphate and K(+) excretion parameters allows a presumptive diagnosis of HPP with more confidence in the emergency room. PMID- 11259689 TI - Longitudinal surveillance of antibiotic use in the hospital. AB - We evaluated antimicrobial use in our hospital by department, including indications for use, source of infections, use of the microbiology laboratory, and appropriateness of prescribing, in a prospective, comparative, non interventional study of all patients receiving antimicrobial agents. We excluded departments where antimicrobial use was negligible. The other 19 departments were followed for 3 (n=4) or 4 (n=15) months, including 2 consecutive months in the spring-summer and either 1 or 2 in the autumn-winter. Antimicrobial therapy was followed from initiation, through possible adaptations, and possible change from intravenous to oral therapy, until discontinuation of treatment. Overall, 6376 antibiotics were given to 2306 patients. Of the surveyed hospitalized patients, 62%+/-22% received antibiotics, with a range of 4-100% per department. Antibiotics were prescribed for infections acquired in the community (3037 instances, 47%), in the hospital (2182, 34%), in a nursing home (575, 9%), and for prophylaxis continued post-operatively (582, 9%). The most common indications for antimicrobial use were: respiratory tract infection (1729, 27%), urinary tract infection (955, 15%), sepsis (701, 11%), intra-abdominal infections (663, 10%), prophylaxis 582 (9%), soft-tissue infection (572, 9%), and surgical site infection (319, 5%). Univariate indicators for appropriateness of treatment were: age, department, site of infection, source of infection, antimicrobial drug and serum creatinine (all p<0.001). Forty-nine antimicrobials were prescribed in 279 combinations, 58% as single agent and 42% as drug combinations. Half of all antimicrobial use consisted of four agents: cefuroxime (19.1%), metronidazole (11.3%), gentamicin (10.6%) and ampicillin (10.2%), which together accounted for 20% of expenditure on antibiotics. Although use of as many as 53% of antimicrobials (26/49) surveyed was restricted, use in this category accounted for only 29% of all antimicrobial courses. Of 6376 antibiotic courses, 4101 (64%) were given intravenously and 2275 (36%) orally. Appropriateness of use of restricted drugs was lower (70%) than of unrestricted ones (84%, p<0.001). Of 24571 defined daily doses (DDD) given orally, 4587 (19%) were restricted, compared to 7264 (34%) of 21602 DDDs given intravenously (p<0.001). Antibiotic treatment in our hospital appears to be substantial and increasing, justifying efforts to improve appropriateness of therapy and improve clinical and financial results. PMID- 11259690 TI - Managing chronic hepatitis C acquired through intravenous drug use. AB - We retrospectively reviewed the provision and uptake of hospital services for 253 current and ex-intravenous drug users with hepatitis C virus (HCV). Overall, 237 attended at least one clinic (mean age 32 years, 70% male, 43% on maintenance methadone); 81% had evidence of active viral replication and 137 agreed to a liver biopsy to assess disease severity. Of these 137, 24% had mild chronic hepatitis with a low risk of progression to cirrhosis, but 9% had cirrhosis (mean age 40 years, mean time since initial intravenous drug use 15.8 years). Only 50 of the 100 patients in whom antiviral therapy was indicated, commenced treatment; 18 (36%) have had a sustained virological response. The natural history or response to treatment of chronic HCV in those who acquire it through intravenous drug use is not different to that previously reported for post-transfusion HCV. However, a substantial proportion default from follow-up or decline further intervention. As intravenous drug use is now the main risk factor for acquisition of HCV, these data have implications for future delivery of care aimed at limiting the morbidity of chronic HCV, and limiting the spread of hepatitis C virus infection amongst intravenous drug users. PMID- 11259691 TI - Reduced vitamin B12 binding by transcobalamin II increases the risk of neural tube defects. AB - Periconceptional folic acid supplementation reduces the risk of neural tube defects (NTD). Homocysteine levels are elevated in mothers of NTD children, which may be due to decreased cellular vitamin B12 levels, as vitamin B12 is a cofactor for the methylation of homocysteine. Transcobalamin II (TC II) transports vitamin B12 to the tissues. To examine whether altered plasma transcobalamin levels are a risk factor for NTD, we determined the apo and holo form of TC II and haptocorrin (TCI+TCIII), vitamin B12 and homocysteine concentrations in the plasma of 46 mothers with NTD children, and in 73 female controls. Holo-tc II levels and holo tc II percentages (holo-tc II/total tc II) in the first quartile of the control distribution were related to a three-fold (OR 2.9, 95%CI 0.9-9.2) and five-fold (OR 5.0, 95%CI 1.3-19.3) risk, respectively, for having a child with NTD, when compared with the last quartile. Homocysteine levels were significantly higher among individuals with low holo-tc II, low total vitamin B12 concentrations and low holo-tc II percentages. These low holo-tc II percentages are probably caused by reduced affinity of TC II for vitamin B12, which may be explained by genetic variation in the TC II gene. Vitamin B12 supplementation might therefore be warranted, in addition to folate, in the prevention of NTD. PMID- 11259692 TI - Management of acute physiological parameters after stroke. AB - Considerable effort has been directed towards acute stroke research with numerous drug therapies being tried and tested. As yet there is still no routine treatment that is unequivocally effective in acute stroke. The development of stroke units has been a major breakthrough in reducing disability through co-ordinated rehabilitation, and new interest is being focussed towards limiting acute neurological deterioration through acute stroke units. Monitoring and attempting to stabilize acute physiological parameters within normal limits such as blood pressure, temperature, hydration status, glucose levels and oxygen saturations, has become standard practice for some acute stroke units. Strategies to correct hypertension, hypotension, dehydration, hyperglycaemia, pyrexia and hypoxia may potentially reduce neuronal damage in the acute phase of stroke and subsequently improve functional outcome and survival. Whether we require large prospective randomized controlled trials to test whether these specific interventions are to be used in mainstay practice is unclear. PMID- 11259694 TI - Scenes from the past: CT in the archaeologic study of ancient Greek ceramics. PMID- 11259693 TI - Gallbladder carcinoma: radiologic-pathologic correlation. AB - Primary carcinoma of the gallbladder is an uncommon, aggressive malignancy that affects women more frequently than men. Older age groups are most often affected, and coexisting gallstones are present in the vast majority of cases. The symptoms at presentation are vague and are most often related to adjacent organ invasion. Therefore, despite advances in cross-sectional imaging, early-stage tumors are not often encountered. Imaging studies may reveal a mass replacing the normal gallbladder, diffuse or focal thickening of the gallbladder wall, or a polypoid mass within the gallbladder lumen. Adjacent organ invasion, most commonly involving the liver, is typically present at diagnosis, as is biliary obstruction. Periportal and peripancreatic lymphadenopathy, hematogenous metastases, and peritoneal metastases may also be seen. The vast majority of gallbladder carcinomas are adenocarcinomas. Because most patients present with advanced disease, the prognosis is poor, with a reported 5-year survival rate of less than 5% in most large series. The radiologic differential diagnosis includes the more frequently encountered inflammatory conditions of the gallbladder, xanthogranulomatous cholecystitis, adenomyomatosis, other hepatobiliary malignancies, and metastatic disease. PMID- 11259695 TI - Illuminations: gallbladder carcinoma. PMID- 11259696 TI - Radiologic spectrum of intraductal papillary mucinous tumor of the pancreas. AB - "Intraductal papillary mucinous tumor" is now the preferred term to describe a spectrum of proliferation of the pancreatic ductal epithelium. The tumor produces an excessive amount of mucin and results in progressive dilation of the main pancreatic duct or cystic dilation of the branch ducts, depending on the location of the tumor. This tumor is small and localized in a segment of the main pancreatic duct or in branch ducts, particularly in the branch ducts of the uncinate process, but it may also be diffuse, involving a wide area of the pancreatic ducts. Excessive mucin may impede the pancreatic duct flow and, in turn, produce symptoms of chronic pancreatitis. The following findings are seen on imaging studies: lobulated multicystic dilatation of the branch ducts, diffuse dilatation of the main pancreatic duct, intraductal papillary tumors, elongated or globlike mucous plugs in the dilated ducts, and bulging of the papilla into the duodenal lumen. The diagnosis is suggested at ultrasonography, computed tomography, or magnetic resonance cholangiopancreatography. Endoscopic retrograde cholangiopancreatography is the imaging modality of choice for the diagnosis, because it depicts the communication between the cystically dilated branch ducts and the diffusely dilated main pancreatic duct, as well as intraductal papillary tumor and mucous plugs. PMID- 11259699 TI - Of posters and exhibits and other things. PMID- 11259698 TI - Helical CT in the diagnosis of small bowel obstruction. AB - With recent technologic developments, the role of computed tomography (CT) in the diagnosis of bowel obstruction has expanded. CT is recommended when clinical and initial radiographic findings remain indeterminate or strangulation is suspected. This modality clearly demonstrates pathologic processes involving the bowel wall as well as the mesentery, mesenteric vessels, and peritoneal cavity. CT should be performed with intravenous injection of contrast material, and use of thin sections is recommended to evaluate a particular region of interest. CT is reported to have a sensitivity of 78%-100% for the detection of complete or high grade small bowel obstruction but may not allow accurate diagnosis in cases involving incomplete obstruction. In such cases, the use of adjunct enteroclysis is indicated. Furthermore, multiplanar reformatted imaging may help identify the site, level, and cause of obstruction when axial CT findings are indeterminate. CT can also demonstrate findings that indicate the presence of closed-loop obstruction or strangulation, both of which necessitate emergency exploratory laparotomy. Unfortunately, these pathologic conditions may be missed, and patients with suspected severe obstruction or bowel ischemia in whom CT and clinical findings are widely disparate must also undergo laparotomy. In general, however, CT allows appropriate and timely management of these emergency cases. PMID- 11259700 TI - Three-dimensional gadolinium-enhanced MR angiography: applications for abdominal imaging. AB - Three-dimensional (3D) gadolinium-enhanced magnetic resonance (MR) angiography is a versatile technique that combines speed, superb contrast, and relative simplicity. It has a wide range of applications, particularly in the abdomen and pelvis, where superb images of the abdominal aorta and renal arteries are routinely obtained. Aneurysms, atherosclerotic lesions, and occlusions of the major mesenteric arteries are also well depicted. In addition, 3D gadolinium enhanced MR angiography is ideal for noninvasive evaluation of the systemic and mesenteric veins and can be used to demonstrate parenchymal lesions in the liver, pancreas, kidneys, and other organs. It is also useful in staging genitourinary neoplasms: Parenchymal lesions, venous extension, and adenopathy are all clearly depicted. Three-dimensional gadolinium-enhanced MR angiography can be useful in the preoperative evaluation of potential transplant donors and recipients and in the evaluation of vascular complications following transplantation. Delayed 3D acquisitions of the kidneys, ureters, and bladder can be performed routinely to generate gadolinium-enhanced urograms and demonstrate obstruction, delayed function, filling defects, and masses. A variety of methods for increasing the speed and improving the resolution of 3D acquisition are currently under investigation. These include novel imaging and reformatting techniques and the use of intravascular contrast agents with much longer vascular half-lives. PMID- 11259702 TI - Three-dimensional volume-rendered CT angiography of the renal arteries and veins: normal anatomy, variants, and clinical applications. AB - Three-dimensional volume-rendered computed tomographic (CT) angiography represents an increasingly important clinical tool that, in many institutions, is replacing conventional angiography in the depiction of normal vascular anatomy and the diagnosis of vascular disorders. Evaluation of conditions affecting the renal vasculature constitutes a major focus of volume-rendered CT angiography, which has documented utility for demonstrating both arterial and venous disease. Arterial disorders include renal artery stenosis, renal artery aneurysms, and dissection. Venous disorders include splenorenal shunts, thrombosis, and intravascular tumor extension. In addition, volume-rendered CT angiography accurately displays the normal and variant renal vascular anatomy, which is crucial to detect before surgery, especially partial nephrectomy and laparoscopic nephrectomy. CT angiography is also useful in the evaluation of the renal vasculature following renal transplantation. Familiarity with proper CT protocols and data acquisition techniques are crucial for accurate diagnosis. PMID- 11259703 TI - Systemic arterial supply to the lungs in adults: spiral CT findings. AB - Systemic arterial supply to the lungs can be congenital or due to acquired disease. Congenital diseases encompass bronchopulmonary sequestration and congenital pulmonary venolobar syndrome, in which the involved lung parenchyma is supplied by the aberrant systemic arteries. An anomalous systemic artery can also supply an area of otherwise normal lung parenchyma. In acquired diseases, hypertrophied normal systemic arteries supply the lungs. Hypertrophied systemic arteries include the bronchial arteries, intercostal arteries, internal mammary arteries, inferior phrenic arteries, branches of the thyrocervical trunk, branches of the hepatic arteries, and branches of the abdominal aorta. Hypertrophy of normal systemic arteries is encountered in patients with bronchiectasis, pulmonary tuberculosis, other pulmonary infections, pulmonary thromboembolism, or chronic obstructive pulmonary disease. These systemic arteries are considered to supply the lungs by means of anastomoses between bronchial and pulmonary arteries within the lung parenchyma or transpleural systemic-pulmonary artery anastomoses. In most cases, the correct diagnosis and treatment plan can be determined by identification of the systemic arteries on computed tomographic scans. PMID- 11259704 TI - Atypical pulmonary metastases: spectrum of radiologic findings. AB - Typical radiologic findings of a pulmonary metastasis include multiple round variable-sized nodules and diffuse thickening of interstitium. In daily practice, however, atypical radiologic features of metastases are often encountered that make distinction of metastases from other nonmalignant pulmonary diseases difficult. A detailed knowledge of the atypical radiologic features of a pulmonary metastasis with a good understanding of the histopathologic background is essential for correct diagnosis. Squamous cell carcinoma is regarded as the most common cell type of a cavitating metastasis, but metastatic nodules from adenocarcinomas and sarcomas also cavitate occasionally. Calcification can occur in a metastatic sarcoma or adenocarcinoma, which makes differentiation from a benign granuloma or hamartoma difficult. Peritumoral hemorrhage results in areas of nodular attenuation surrounded by a halo of ground-glass opacity. Pneumothorax commonly occurs in metastases from an osteosarcoma. Air-space consolidation is often seen in cases of metastases from gastrointestinal tract malignancies. Even though tumor emboli in pulmonary arteries can be seen at computed tomography, diagnosis is difficult because they are located in small or medium arteries. A common radiologic appearance of an endobronchial metastasis is an atelectasis. In cases of an endobronchial or a solitary pulmonary metastasis, differentiation between bronchogenic carcinoma and metastasis is difficult. Dilated vascular structures within the mass can be seen in metastatic sarcomas. A sterilized metastasis after chemotherapy is radiologically indistinguishable from a residual viable tumor. Benign tumors such as uterine leiomyomas and giant cell tumors of the bone rarely metastasize to the lung. PMID- 11259705 TI - Unusual lesions of the cerebellopontine angle: a segmental approach. AB - Tumors of the cerebellopontine angle (CPA) are frequent; acoustic neuromas and meningiomas represent the great majority of such tumors. However, a large variety of unusual lesions can also be encountered in the CPA. The site of origin is the main factor in making a preoperative diagnosis for an unusual lesion of the CPA. In addition, it is essential to analyze attenuation at computed tomography (CT), signal intensity at magnetic resonance (MR) imaging, enhancement, shape and margins, extent, mass effect, and adjacent bone reaction. CPA masses can primarily arise from the cerebellopontine cistern and other CPA structures (arachnoid cyst, nonacoustic schwannoma, aneurysm, melanoma, miscellaneous meningeal lesions) or from embryologic remnants (epidermoid cyst, dermoid cyst, lipoma). Tumors can also invade the CPA by extension from the petrous bone or skull base (cholesterol granuloma, paraganglioma, chondromatous tumors, chordoma, endolymphatic sac tumor, pituitary adenoma, apex petrositis). Finally, CPA lesions can be secondary to an exophytic brainstem or ventricular tumor (glioma, choroid plexus papilloma, lymphoma, hemangioblastoma, ependymoma, medulloblastoma, dysembryoplastic neuroepithelial tumor). A close association between CT and MR imaging findings is very helpful in establishing the preoperative diagnosis for unusual lesions of the CPA. PMID- 11259706 TI - Metastatic involvement of the heart and pericardium: CT and MR imaging. AB - Metastases to the heart and pericardium are much more common than primary cardiac tumors and are generally associated with a poor prognosis. Tumors that are most likely to involve the heart and pericardium include cancers of the lung and breast, melanoma, and lymphoma. Tumor may involve the heart and pericardium by one of four pathways: retrograde lymphatic extension, hematogenous spread, direct contiguous extension, or transvenous extension. Metastatic involvement of the heart and pericardium may go unrecognized until autopsy. Impairment of cardiac function occurs in approximately 30% of patients and is usually attributable to pericardial effusion. The clinical presentation includes shortness of breath, which may be out of proportion to radiographic findings in patients with pericardial effusion or may be the result of associated pleural effusion. Patients may also present with cough, anterior thoracic pain, pleuritic chest pain, or peripheral edema. The differential diagnosis of pericardial effusion in a patient with known malignancy includes malignant pericardial effusion, radiation-induced pericarditis, drug-induced pericarditis, and idiopathic pericarditis. Any disease process that causes thickening or nodularity of the pericardium or myocardium or masses within the cardiac chambers can mimic metastatic disease. PMID- 11259709 TI - Stereotactic localization of breast lesions: how it works and methods to improve accuracy. AB - A computer simulation of stereotactic breast biopsy was developed that paralleled the geometric configuration of a currently available breast biopsy system. This model was developed to define and improve the targeting of breast lesions with stereotactic biopsy techniques. Lesions must be clearly identified and accurately targeted on both views for successful localization. Nonvisualization of a lesion may result from overlying tissue or from the geometric configuration of the imaging system. Familiarity with the geometric configuration of the biopsy unit, especially the location of the reference point and center of rotation, facilitates understanding of apparent changes in lesion position (parallax shift). Inaccuracy in lesion targeting on one or both views will manifest predominantly as an error in the calculated z value (depth). The magnitude and direction of this error are largely determined by the direction of the targeting error. Compensatory strategies include use of a long-throw core biopsy gun or directional vacuum-assisted biopsy device and additional sampling along the z axis and should be accompanied by critical evaluation of both pre- and postfire images. Understanding geometric considerations as well as how targeting accuracy affects accuracy in lesion localization should lead to greater success in sampling even challenging breast lesions at stereotactic biopsy. PMID- 11259707 TI - Urachal remnant diseases: spectrum of CT and US findings. AB - Computed tomography (CT) and ultrasonography (US) are ideally suited for demonstrating urachal remnant diseases. A patent urachus is demonstrated at longitudinal US and occasionally at CT as a tubular connection between the anterosuperior aspect of the bladder and the umbilicus. An umbilical-urachal sinus manifests at US as a thickened tubular structure along the midline below the umbilicus. A vesicourachal diverticulum is usually discovered incidentally at axial CT, appearing as a midline cystic lesion just above the anterosuperior aspect of the bladder. At US, it manifests as an extraluminally protruding, fluid filled sac that does not communicate with the umbilicus. Urachal cysts manifest at both modalities as a noncommunicating, fluid-filled cavity in the midline lower abdominal wall located just beneath the umbilicus or above the bladder. Both infected urachal cysts and urachal carcinomas commonly display increased echogenicity at US and thick-walled cystic or mixed attenuation at CT, making it difficult to differentiate between them. Percutaneous needle biopsy or fluid aspiration is usually needed for diagnosis and therapeutic planning. Nevertheless, CT and US can help identify most disease entities originating from the urachal remnant in the anterior abdominal wall. Understanding the anatomy and the imaging features of urachal remnant diseases is essential for correct diagnosis and proper management. PMID- 11259710 TI - Ovarian teratomas: tumor types and imaging characteristics. AB - Ovarian teratomas include mature cystic teratomas (dermoid cysts), immature teratomas, and monodermal teratomas (eg, struma ovarii, carcinoid tumors, neural tumors). Most mature cystic teratomas can be diagnosed at ultrasonography (US) but may have a variety of appearances, characterized by echogenic sebaceous material and calcification. At computed tomography (CT), fat attenuation within a cyst is diagnostic. At magnetic resonance (MR) imaging, the sebaceous component is specifically identified with fat-saturation techniques. The US appearances of immature teratoma are nonspecific, although the tumors are typically heterogeneous, partially solid lesions, usually with scattered calcifications. At CT and MR imaging, immature teratomas characteristically have a large, irregular solid component containing coarse calcifications. Small foci of fat help identify these tumors. The US features of struma ovarii are also nonspecific, but a heterogeneous, predominantly solid mass may be seen. On T1- and T2-weighted images, the cystic spaces demonstrate both high and low signal intensity. Familiarity with the US, CT, and MR imaging features of ovarian teratomas can aid in differentiation and diagnosis. PMID- 11259711 TI - Invasive procedures in the female pelvis: value of transabdominal, endovaginal, and endorectal US guidance. AB - Transabdominal, endovaginal, and endorectal ultrasonographic (US) guidance is indispensable for a multitude of invasive procedures in the female pelvis. Transabdominal uterine US performed with a fluid-filled bladder is appropriate and convenient for guidance of difficult dilation and curettage procedures. Transabdominal intraoperative US can be employed to guide several procedures for which the more expensive intraoperative hysteroscopic procedure is now used. Aspiration of symptomatic ovarian cysts that appear benign at US with an endovaginally guided small-gauge needle is simple and effective. Simple noninfected pelvic fluid collections may be aspirated transvaginally for both diagnosis and therapy by using endovaginal guidance. Endovaginal US demonstrates the anatomic relationships of a pelvic abscess to adjacent structures, allowing safe access for transvaginal drainage. By using an endovaginal transducer with a needle guide, cervical and vaginal cuff masses may be easily sampled. An obstructed uterus may be accessed by puncturing obstructive tissue with a trocar containing needle guided by an endorectal probe. US guidance for placement of a central brachytherapy tandem is performed via the abdominal approach after the bladder has been distended with sterile water. Endorectal US transducers may be effectively used to guide placement of interstitial brachytherapy needles in pelvic soft-tissue masses. PMID- 11259712 TI - Quantification of fluoroscopic imaging system contrast by using video waveform monitoring. AB - A noninvasive method was developed for quantifying the overall contrast of fluoroscopic imaging systems within the clinical setting by using a simple phantom and common video test equipment. In this method, an acrylic phantom with four holes filled with varying amounts of air and aluminum is placed on the entrance exposure side of a patient-equivalent acrylic phantom. The air- and aluminum-filled holes provide a stepped gray-scale pattern that is displayed on the examination room viewing monitor when the phantom is fluoroscopically imaged under automatic brightness control. A video waveform monitor or oscilloscope is then used to quantify those video signal voltage levels as a contrast index value, which is defined as the maximum range of the video signal voltage levels of the gray-scale steps. The method is repeatable and allows quantification of the contrast of the imaging system. It can also be used to optimize video parameters, provide comparative data for quality control monitoring, and characterize overall contrast differences between systems. Experience with this method suggests that there is excellent correlation between the clinical perception of image contrast and the contrast index, with contrast index changes of approximately 15% being seen clinically. PMID- 11259713 TI - Incidental detection of diminished bone marrow metabolic activity with FDG PET. PMID- 11259716 TI - The AAPM/RSNA physics tutorial for residents: digital fluoroscopy. AB - A digital fluoroscopy system is most commonly configured as a conventional fluoroscopy system (tube, table, image intensifier, video system) in which the analog video signal is converted to and stored as digital data. Other methods of acquiring the digital data (eg, digital or charge-coupled device video and flat panel detectors) will become more prevalent in the future. Fundamental concepts related to digital imaging in general include binary numbers, pixels, and gray levels. Digital image data allow the convenient use of several image processing techniques including last image hold, gray-scale processing, temporal frame averaging, and edge enhancement. Real-time subtraction of digital fluoroscopic images after injection of contrast material has led to widespread use of digital subtraction angiography (DSA). Additional image processing techniques used with DSA include road mapping, image fade, mask pixel shift, frame summation, and vessel size measurement. Peripheral angiography performed with an automatic moving table allows imaging of the peripheral vasculature with a single contrast material injection. PMID- 11259717 TI - Image content extraction: application to MR images of the brain. AB - A system for automatically extracting image content features was developed that combines registration to a labeled atlas with natural language processing of free text radiology reports. The system was then tested with T1-weighted, spoiled gradient-echo magnetic resonance (MR) imaging studies of the brain performed in nine patients. The locations of 599 structures were visually assessed by an experienced radiologist and compared with the locations indicated by automated output. The in-plane accuracy of the contours was subjectively evaluated as either good, moderate, or poor. The criterion for classifying a structure as correctly located was that 90% or more of all the images containing the structure had to be correctly identified. For 98% of the structures, the images identified by the automated algorithm agreed with those identified by the radiologist, and in 83% of cases, image contours showed a good in-plane overlap. The results of this validation study demonstrate that this combination of registration and natural language processing is accurate in identifying relevant images from brain MR imaging studies. However, the range of applicability of this technique has yet to be determined by applying the technique to a large number of studies. PMID- 11259719 TI - Paternal and maternal components of the predisposition to preeclampsia. AB - BACKGROUND: There is an inherited maternal predisposition to preeclampsia. Whether there is a paternal component, however, is not known. METHODS: We used records of the Utah Population Database to identify 298 men and 237 women born in Utah between 1947 and 1957 whose mothers had had preeclampsia during their pregnancy. For each man and woman in the study group, we identified two matched, unrelated control subjects who were not the products of pregnancies complicated by preeclampsia. We then identified 947 children of the 298 male study subjects and 830 children of the 237 female study subjects who had been born between 1970 and 1992. These children were matched to offspring of the control subjects (1950 offspring of the male control group and 1658 offspring of the female control group). Factors associated with preeclampsia were identified, and odds ratios were calculated with the use of stepwise logistic-regression analysis. RESULTS: In the group whose mothers had had preeclampsia (the male study group), 2.7 percent of the offspring (26 of 947) were born of pregnancies complicated by preeclampsia, as compared with 1.3 percent of the offspring (26 of 1973) in the male control group. In the female study group, 4.7 percent of the pregnancies (39 of 830) were complicated by preeclampsia, as compared with 1.9 percent (32 of 1658) in the female control group. After adjustment for the offspring's year of birth, maternal parity, and the offspring's gestational age at delivery, the odds ratio for an adult whose mother had had preeclampsia having a child who was the product of a pregnancy complicated by preeclampsia was 2.1 (95 percent confidence interval, 1.0 to 4.3; P=0.04) in the male study group and 3.3 (95 percent confidence interval, 1.5 to 7.5; P=0.004) in the female study group. CONCLUSIONS: Both men and women who were the product of a pregnancy complicated by preeclampsia were significantly more likely than control men and women to have a child who was the product of a pregnancy complicated by preeclampsia. PMID- 11259720 TI - Effects of multisite biventricular pacing in patients with heart failure and intraventricular conduction delay. AB - BACKGROUND: One third of patients with chronic heart failure have electrocardiographic evidence of a major intraventricular conduction delay, which may worsen left ventricular systolic dysfunction through asynchronous ventricular contraction. Uncontrolled studies suggest that multisite biventricular pacing improves hemodynamics and well-being by reducing ventricular asynchrony. We assessed the clinical efficacy and safety of this new therapy. METHODS: Sixty seven patients with severe heart failure (New York Heart Association class III) due to chronic left ventricular systolic dysfunction, with normal sinus rhythm and a duration of the QRS interval of more than 150 msec, received transvenous atriobiventricular pacemakers (with leads in one atrium and each ventricle). This single-blind, randomized, controlled crossover study compared the responses of the patients during two periods: a three-month period of inactive pacing (ventricular inhibited pacing at a basic rate of 40 bpm) and a three-month period of active (atriobiventricular) pacing. The primary end point was the distance walked in six minutes; the secondary end points were the quality of life as measured by questionnaire, peak oxygen consumption, hospitalizations related to heart failure, the patients' treatment preference (active vs. inactive pacing), and the mortality rate. RESULTS: Nine patients were withdrawn from the study before randomization, and 10 failed to complete both study periods. Thus, 48 patients completed both phases of the study. The mean distance walked in six minutes was 22 percent greater with active pacing (399+/-100 m vs. 326+/-134 m, P<0.001), the quality-of-life score improved by 32 percent (P<0.001), peak oxygen uptake increased by 8 percent (P<0.03), hospitalizations were decreased by two thirds (P<0.05), and active pacing was preferred by 85 percent of the patients (P<0.001). CONCLUSIONS: Although it is technically complex, atriobiventricular pacing significantly improves exercise tolerance and quality of life in patients with chronic heart failure and intraventricular conduction delay. PMID- 11259721 TI - Treatment of chronic granulomatous disease with nonmyeloablative conditioning and a T-cell-depleted hematopoietic allograft. AB - BACKGROUND: The treatment of chronic granulomatous disease with conventional allogeneic hematopoietic stem-cell transplantation carries a high risk of serious complications and death. We investigated the feasibility of stem-cell transplantation without ablation of the recipient's bone marrow. METHODS: Ten patients, five children and five adults, with chronic granulomatous disease underwent peripheral-blood stem-cell transplantation from an HLA-identical sibling. We used a nonmyeloablative conditioning regimen consisting of cyclophosphamide, fludarabine, and antithymocyte globulin. The allograft was depleted of T cells to reduce the risk of severe graft-versus-host disease. Donor lymphocytes were administered at intervals of 30 days or more after the transplantation to facilitate engraftment. RESULTS: After a median follow-up of 17 months (range, 8 to 26), the proportion of donor neutrophils in the circulation in 8 of the 10 patients was 33 to 100 percent, a level that can be expected to provide normal host defense; in 6 the proportion was 100 percent. In two patients, graft rejection occurred. Acute graft-versus-host disease (grade II, III, or IV) developed in three of the four adult patients with engraftment, one of whom subsequently had chronic graft-versus-host disease. None of the five children had grade II, III, or IV acute graft-versus-host disease. During the follow-up period, four serious infections occurred among the patients who had engraftment. Three of the 10 recipients died. Preexisting granulomatous lesions resolved in the patients in whom transplantation was successful. CONCLUSIONS: Nonmyeloablative conditioning followed by a T-cell-depleted hematopoietic stem cell allograft is a feasible option for patients with chronic granulomatous disease, recurrent life-threatening infections, and an HLA-identical family donor. PMID- 11259722 TI - The Japanese experience with vaccinating schoolchildren against influenza. AB - BACKGROUND: Influenza epidemics lead to increased mortality, principally among elderly persons and others at high risk, and in most developed countries, influenza-control efforts focus on the vaccination of this group. Japan, however, once based its policy for the control of influenza on the vaccination of schoolchildren. From 1962 to 1987, most Japanese schoolchildren were vaccinated against influenza. For more than a decade, vaccination was mandatory, but the laws were relaxed in 1987 and repealed in 1994; subsequently, vaccination rates dropped to low levels. When most schoolchildren were vaccinated, it is possible that herd immunity against influenza was achieved in Japan. If this was the case, both the incidence of influenza and mortality attributed to influenza should have been reduced among older persons. METHODS: We analyzed the monthly rates of death from all causes and death attributed to pneumonia and influenza, as well as census data and statistics on the rates of vaccination for both Japan and the United States from 1949 through 1998. For each winter, we estimated the number of deaths per month in excess of a base-line level, defined as the average death rate in November. RESULTS: The excess mortality from pneumonia and influenza and that from all causes were highly correlated in each country. In the United States, these rates were nearly constant over time. With the initiation of the vaccination program for schoolchildren in Japan, excess mortality rates dropped from values three to four times those in the United States to values similar to those in the United States. The vaccination of Japanese children prevented about 37,000 to 49,000 deaths per year, or about 1 death for every 420 children vaccinated. As the vaccination of schoolchildren was discontinued, the excess mortality rates in Japan increased. CONCLUSIONS: The effect of influenza on mortality is much greater in Japan than in the United States and can be measured about equally well in terms of deaths from all causes and deaths attributed to pneumonia or influenza. Vaccinating schoolchildren against influenza provides protection and reduces mortality from influenza among older persons. PMID- 11259723 TI - Images in clinical medicine. Erythrophagocytosis. PMID- 11259724 TI - Spontaneous dissection of the carotid and vertebral arteries. PMID- 11259726 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 9-2001. A 64-year-old woman with peripheral neuropathy, paraproteinemia, and lymphadenopathy. PMID- 11259725 TI - Cytokine pathways and joint inflammation in rheumatoid arthritis. PMID- 11259727 TI - Risk factors for preeclampsia. PMID- 11259728 TI - What is the best way to treat inherited disorders? PMID- 11259729 TI - Medicare. PMID- 11259730 TI - Good editorial practice: editors as educators. AB - There may be valuable research going on in the developing and financially less privileged countries, but it usually does not reach international visibility, in spite of a large number of scientific journals in these countries. Such journals are not only invisible but, by perpetuating a vicious circle of inadequacy, may be directly damaging to the local science and research culture. We call for an international action to help journal editors in less privileged countries. International associations of editors may be leaders of these activities by defining, promoting, and perhaps controlling good editorial practice, as a main criterion for international recognition of a journal. However, the editors of small journals have the power and moral obligation to become a stronghold of quality and advancement in their scientific community. Their educational "tools" are editorial integrity and author-friendly policy. Editors can teach the authors study design, statistical analysis, precision, punctuality, research integrity, style and format of writing, and other aspects of scientific communication. The editors of "big", mainstream scientific journals can act as global educators, teaching and providing guidance to editors of small journals. The editors from developed countries as leaders, and editors from less advantageous environments as teachers are the key figures in shaping research communication in less privileged scientific communities. PMID- 11259731 TI - Report of the world association of medical editors: agenda for the future. AB - During a 3-day meeting at Bellagio in January 2001, a group of 20 editors from 12 countries in 5 continents met to map out a strategy for the World Association of Medical Editors (WAME)'s continued development in the service of medical editors over the next several years. The group: 1) Developed a statement of principles on the standards of professionalism and responsibilities of editors (this statement will be posted on the Web site after electronic consultation with and comment by WAME editors); 2) Agreed to assess the extent to which these principles are reflected in practice and to explore barriers to their adoption, using data from a survey and focus groups; 3) Developed and outlined an on-line program for distance learning, targeted at new editors; 4) Planned for formal evaluation of the educational outreach program; and 5) Agreed to support regional initiatives to strengthen local editorial capacity. Underpinning all past and proposed future activities is the WAME Web site. The ambitious plans outlined above will require extensive development of the site, plans for which were made at the Bellagio meeting. PMID- 11259732 TI - Is pain a somatic symptom? AB - The grouping of symptoms into "somatic" or "physical" on the one hand and "mental", "somatoform", or "psychological" on the other are vestiges of an era in medicine when it seemed useful to divide all the phenomena of disease into two groups - one related to the soma and other to the psyche. Today, this division is becoming obsolete and is harmful. Obsolete, because we are discovering changes in the tissues or in biochemical and immunological functions of the body in people with mental disorders, and because psychological complaints are frequent in all physical illnesses. Harmful, because the labels "psychological", "psychogenic", or "somatoform" are so loaded with connotations of being simulations or complaints about nothing that patients are unlikely to recive the help they need. PMID- 11259733 TI - On depleted uranium: gulf war and Balkan syndrome. AB - The complex clinical symptomatology of chronic illnesses, commonly described as Gulf War Syndrome, remains a poorly understood disease entity with diversified theories of its etiology and pathogenesis. Several causative factors have been postulated, with a particular emphasis on low level chemical warfare agents, oil fires, multiple vaccines, desert sand (Al-Eskan disease), botulism, Aspergillus flavus, Mycoplasma, aflatoxins, and others, contributing to the broad scope of clinical manifestations. Among several hundred thousand veterans deployed in the Operation Desert Storm, 15-20% have reported sick and about 25,000 died. Depleted uranium (DU), a low-level radioactive waste product of the enrichment of natural uranium with U-235 for the reactor fuel or nuclear weapons, has been considered a possible causative agent in the genesis of Gulf War Syndrome. It was used in the Gulf and Balkan wars as an armor-penetrating ammunition. In the operation Desert Storm, over 350 metric tons of DU was used, with an estimate of 3-6 million grams released in the atmosphere. Internal contamination with inhaled DU has been demonstrated by the elevated excretion of uranium isotopes in the urine of the exposed veterans 10 years after the Gulf war and causes concern because of its chemical and radiological toxicity and mutagenic and carcinogenic properties. Polarized views of different interest groups maintain an area of sustained controversy more in the environment of the public media than in the scientific community, partly for the reason of being less than sufficiently addressed by a meaningful objective interdisciplinary research. PMID- 11259734 TI - Thrombophilia and adverse pregnancy outcome. AB - Congenital and acquired thrombophilias are the most common predisposing factors for thromboembolism, but they may also contribute to pathophysiological processes involved in recurrent pregnancy loss, fetal death, intrauterine growth restriction, placental abruption, placental infarction, and pre-eclampsia. The most common thrombophilias are deficiencies of antithrombin III, protein C, and protein S, acquired protein C resistance, genetic mutation encoding for factor V Leiden, prothrombin gene, and inherited hyperhomocysteinemia, and antiphospholipid syndrome. Although adverse pregnancy outcomes are more common in women with thrombophilia, the current evidence does not support routine thrombophilia screening of all pregnant women. Selective thrombophilia screening may be justified in certain group of women, particularly those with a history of thromboembolism. More research is required to confirm or refute the causal link between thrombophilia and abnormal placentation, and assess effectiveness and safety of thromboprophylaxis in pregnant women. PMID- 11259735 TI - Serum paraoxonase activities in hemodialyzed uremic patients: cohort study. AB - AIM: To determine whether paraoxonase activity, paraoxonase phenotypes, and lipid status are altered in uremic patients on long-term hemodialysis treatment as compared to healthy population. METHODS: Patients (n = 69) and control subjects (n = 145) were from the area of Slavonski Brod, Croatia. Paraoxon was used as a substrate for measuring basal or sodium chloride-stimulated (NaCl-stimulated) paraoxonase activity, and phenylacetate for measuring arylesterase activity. The double substrate method was used to assign phenotypes. Cholesterol, triglycerides, and high-density lipoprotein cholesterol (HDL-cholesterol) were determined by methods routinely used in medical-biochemical laboratories. Enzyme activities are expressed as international units per liter of serum or per mmol of HDL-cholesterol (HDL-standardized activities). RESULTS: Basal and NaCl-stimulated paraoxonase activity, as well as arylesterase activity expressed per serum volume, were significantly lower in the hemodialyzed uremic patients compared to the controls; 69% (p < 0.001), 73% (p < 0.001) and 49%, (p < 0.001), respectively. However, basal and NaCl-stimulated paraoxonase activity standardized for HDL-cholesterol concentrations were not significantly reduced in the hemodialyzed uremic patients as compared to controls (86%, p = 0.614 and 87%, p = 0.720, respectively), contrary to arylesterase activity, which remained significantly lower (72%, p < 0.001). The distribution of paraoxonase phenotypes in hemodialyzed uremic patients and controls was as follows: AA 45% and 39%, AB 37% and 48%, BB 18%, and 13%, respectively. CONCLUSION: Patients on long-term hemodialysis have decreased paraoxonase/arylesterase activity, which might indicate a greater risk of premature atherogenesis. PMID- 11259737 TI - Children war casualties during the 1991-1995 wars in Croatia and Bosnia and Herzegovina. AB - AIM: To analyze clinical course of war-related injuries in children treated at the Split University Hospital during the wars in Croatia (1991-1995) and Bosnia and Herzegovina (1992-1995). METHODS: Medical records of 94 treated children were analyzed. The severity of wounds was scored according to the Abbreviated Injury Scale (AIS) and Injury Severity Score (ISS) evaluation systems. RESULTS: Most children were wounded during shelling/bombing (n = 28, 10 boys and 18 girls) and by left over explosive devices (n = 26). Children injured by left over explosive devices were predominantly boys (23/26 children), aged 10 to 16 years (19/26 children). Extremities were the most frequently wounded body regions (43% of all wounded regions). The wounds to the head/neck (median AIS = 5.0, range 1-6) and abdomen (median AIS = 4.5, range 3-5) were the most severe. Abdominal wounds required surgical procedures (p < 0.001) and antibiotic treatment (p < 0.05) most frequently, as well as patients with greater AIS and ISS scores (p < 0.05). According to the treatment outcome, more patients wounded to the abdomen and extremities showed improvement than no change or complete recovery (p < .05). Permanent disability remained in 37 (39.4%) children and three (3.3%) children died. CONCLUSION: Boys in upper elementary grades and high school were at greater risk of being wounded by fragments of left over explosive devices than younger boys or girls. The most severe wounds were to the head/neck and the abdomen and inflicted during the shelling or bombing. This should be taken into account in organization of surgical care for the children with war-related injuries. PMID- 11259736 TI - Effect of oral glucose loading on serum gastrin level in pregnant and non pregnant women. AB - AIM: To evaluate the relationship between the changes in gastrin and insulin serum concentrations after oral glucose loading in pregnant and non-pregnant women. METHODS: Thirty women, 12 pregnant and 18 non-pregnant, with normal fasting glucose values were included in the study. Serum concentrations of gastrin, glucose, insulin, and glucagon were analyzed at 0 (t1), 30 (t2) and 60 (t3) minutes after 75 g oral glucose loading. Gastrin, insulin, and glucagon levels were determined by means of radioimmunoassay kits. RESULTS: Serum gastrin concentration in pregnant women increased insignificantly (gastrin median values 57.91, 70.62, and 68.70 for t1, t2, and t3, respectively; Friedman's test, p = 0.264). In non-pregnant women gastrin levels insignificantly increased from t1 to t2, but reduced significantly from t2 to t3 (gastrin median values 62.91, 86.92, and 62.25 for t1, t2 and t3, respectively; Bonferroni adjusted Wilcoxon test, p = 0.002). Unlike in pregnant women, the changes in gastrin release in non-pregnant women were associated with changes in blood glucose concentrations at t2 and t3, which were induced by oral glucose loading. Glucose median values were 7.48 and 6.43 for t2 and t3, respectively. The insulin release due to the oral glucose loading markedly increased at t2 and t3 (Friedman's test, p < 0.001), whereas glucagon release decreased irrespective of pregnancy. CONCLUSION: Changes in blood glucose concentrations induced by oral glucose loading could influence gastrin release, especially in non-pregnant women. Changes in insulin and glucagon levels induced by oral glucose loading, particularly after 60 minutes, could not be associated with changes in gastrin release. PMID- 11259738 TI - Increase of frequency of post-traumatic stress disorder in disabled war veterans during prolonged stay in a rehabilitation hospital. AB - AIM: To explore possible causative factors in the development of post-traumatic stress disorder (PTSD) in disabled Croatian war veterans. METHOD: The sample comprised 42 disabled Croatian war veterans, aged 19 to 44 years, accommodated in the VaraZdinske Toplice Rehabilitation Hospital for the purpose of long-term physical rehabilitation. Manifestation of PTSD symptoms (Mississippi Scale for Combat-Related Post-Traumatic Stress Disorder) and anxiety levels (Spilberger's State Trait Anxiety Inventory) were tested in 1994 and 1999. RESULTS: Patients with PTSD symptoms had significantly higher anxiety levels then patients without PTSD symptoms. The percentage of patients manifesting PTSD increased from 19% in 1994 to 41% in 1999. Over the same period, the anxiety levels decreased in the patients with PTSD. CONCLUSION: Anxiety and PTSD seem to share common etiologic grounds. Nevertheless, staying in the same homogenous group for a substantial period of time, in combination with inadequate social support and deficient psychological care, may contribute to the development of the PTSD symptomatology. PMID- 11259740 TI - Ultrasound screening of the neonatal hip: cost-benefit analysis. AB - AIM: To explore the economic justification for introducing ultrasound screening for developmental dysplasia of the hip in Croatia. METHODS: The analysis was based on the two formulas: that cost-benefit equals benefit/cost, and that net benefit equals benefit minus cost. Screening costs were expressed as a sum of training costs and fee for ultrasound screening of neonates. The neonatologists' working hours and utilization of ultrasound instruments were expressed by multiplying the number of infants born per year in Croatia (N = 47,792) with the standard time needed for one examination and then dividing the product by the number of employed neonatologists (N = 54) and number of ultrasound instruments (N = 58). The benefit was expressed as a late case treatment costs and screening costs ratio. Savings, which would have resulted from the reduction in expected treatment costs of patients with hip problems at later age, represent the indirect benefit. RESULTS: Total hip screening costs would have amounted to US$329,537.80, including the training costs of US$31,035.90. On the average, a neonatologist would spend 71.4 hours screening per year, whereas the instrument utilization would be 64.7 hours. An ultrasound-screening program would save annually US$195,336.50, compared with the existing diagnostic approach. The treatment costs without ultrasound screening were 1.6 times higher than the screening costs. Hospital treatment costs for 165 patients needing endoprosthesis would cover the total screening program in the whole country. CONCLUSION: It is economically justified to introduce ultrasound screening for developmental dysplasia of the hip in neonates in Croatia, a country with transitional and developing economy. PMID- 11259739 TI - Posttraumatic stress disorder and depression in soldiers with combat experiences. AB - AIM: To compare psychological, medical, and trauma-related variables in veterans with combat-related post-traumatic stress disorder (CR-PTSD) comorbid with depression and veterans with CR-PTSD only. METHOD: Out of 402 Croatian veterans recruited during expert evaluation for war-related compensation claims, 346 met the criteria for CR-PTSD: 97 for CR-PTSD only and 249 for PTSD comorbid with other diagnoses (77 comorbid with depression). To reach diagnosis, psychiatrists used clinical interview based on DSM-IV criteria, interview with family and friends, previous medical documentation, and Hamilton Rating Scales for Depression and Anxiety. An independent psychologist used a structured psychological interview, Mississippi CR-PTSD scale, Watson's PTSD criteria, Minnesota Multiphasic Personality Inventory-version 201, and trauma questionnaire based on the Harvard Questionnaire. RESULTS: Out of 402 soldiers, 13.9% did not meet the criteria for PTSD or other psychiatric diagnosis, 61.9% met the criteria for comorbid diagnoses, and 24.2% for PTSD only. The PTSD group with depression did not differ from PTSD-only group in combat experience, number of traumatic events, age, length of employment, sick leave, education, or marital status (chi square test, p = 0.121-0.672). The two groups differed in pre-trauma factors, such as mental disturbances before combat experiences (p = 0.003), positive family history of psychiatric illness (p = 0.008), primary major depression (p = 0.012), and the number of hospital admissions (p = 0.002). CONCLUSION: Different assessment methods in expert examination of combat-experienced soldiers with PTSD for compensation-related purposes are needed to establish the final diagnosis and avoid possibility of factitious disorder or malingering. Combat ability assessment should include assessment of previous psychiatric disturbances of soldiers and their families. PMID- 11259741 TI - Ochratoxin A in corn and wheat: geographical association with endemic nephropathy. AB - AIM: To determine the presence and concentration of ochratoxin A in wheat and corn from Slavonski Brod surroundings, the area of endemic nephropathy allegedly caused by ochratoxin. METHODS: Thin-layer chromatography was used to determine ochratoxin A concentrations in 92 wheat and 51 corn samples from the surroundings of Slavonski Brod, Osijek, Hrvatsko Zagorje, Istria, and Celje (Slovenia). RESULTS: Ochratoxin A was present in 74 of 92 (75.8%) wheat samples and 17 of 51 (33.3%) corn samples, in a concentration range of 0.02-160.00 mg/kg in wheat and 0.02-40.00 mg/kg in corn. Wheat samples from the Slavonski Brod surroundings contained the highest level of ochratoxin A (38.8 +/- 27.2 mg/kg), followed by Osijek (8.7 +/- 8.3 mg/kg). Ochratoxin A levels in the wheat from Hrvatsko Zagorje, Istria, and Celje were considerably lower (2.1 +/- 1.5, 1.3 +/- 2.6 and 0.2 +/- 0.5 mg/kg, respectively). Wheat samples from Slavonski Brod significantly differed from all other sample groups (p < 0.001), and wheat samples from Osijek differed from those from Hrvatsko Zagorje, Istria, and Celje (p < 0.001, p = 0.003, p < 0.001, respectively). Ochratoxin A level was the highest in the corn samples from the Slavonski Brod surroundings (20.0 +/- 14.8 mg/kg) and considerably lower in samples from Osijek, Celje, Hrvatsko Zagorje, and Istria (0.8 +/- 1.4, 0.7 +/- 1.9, 0.4 +/- 0.4, and 0.4 +/- 0.8 mg/kg, respectively). A statistically significant difference was also observed between the Slavonski Brod samples and all other corn samples (p < 0.001). CONCLUSION: Irrespective of the real association between ochratoxin A and endemic nephropathy, our data clearly demonstrate their geographical overlap. PMID- 11259742 TI - Increased incidence of colorectal cancer in the split-dalmatia county: epidemiological study. AB - AIM: To investigate the incidence of colorectal cancer in the Split-Dalmatia County in the 1981-1998 period, and compare it with the incidence in the Republic of Croatia. METHODS: The data were obtained using case records and registries of all hospitals and Public Health Institute in the County and the Croatian Cancer Registry. Age-standardized incidence per 100,000 was calculated from the number of patients with colorectal cancer and the number of inhabitants. RESULTS: There were 2,454 new cases of colorectal cancer (1,383 men and 1,071 women) in the Split-Dalmatia County in 1981-1998. Colon cancer was diagnosed in 55% of the cases. Age-standardized incidence rates for colorectal carcinoma per 100,000 population were 11.4 (men 14.8, women 9.0) in 1981, and 63.5 (men 93.1, women 42.5) in 1998. The total incidence increased from 16.1 (colon cancer 7.9, rectal cancer 8.2) in 1981-1985 period to 52.8 (colon cancer 30.5, rectal cancer 22.3) in 1994-1998 period, or approximately 3.3 times. The colorectal cancer incidence rate in the Split-Dalmatia County increased from 16.2 in 1985 to 46.4 in 1995, and in whole Croatia from 32.4 in 1985 to 37.8 in 1995. CONCLUSION: There was a great increase in the reported incidence of colorectal cancer in the Split Dalmatia County in the 1981-1998 period. The relative increase of incidence in the colorectal cancer was much greater in the Split-Dalmatia County than in Croatia as a whole. These changes call for preventive and screening measures for colorectal carcinoma. PMID- 11259743 TI - Comparison of histopathologists' workloads in two pathology departments in Croatia. AB - AIM: To calculate workloads of Departments of Pathology and staff histopathologists at two Croatian University Hospitals by means of the Suvarna and Kay's Kim Unit (KU) activity index. METHOD: The total number of specimens and KU activity/year were calculated to compare staff pathologists' workload in two Pathology Departments, one at Split and the other at Zagreb University Hospital. The individual specimen types were assigned a difficulty score on a scale of 1-5 units (KUs), depending on the time needed for specimen dissection and macroscopic description, number of sections and stains required, and time spent on microscopy of an "average case". KU activity was calculated for all pathologists individually in terms of histology (1-5 KUs), autopsy (10 KUs), supervision of residents, and outside consultations (2 KUs). RESULTS: According to calculated Kim Unit activity index, pathologists' workload in two investigated pathology departments was not equally spread. In the Split Department of Pathology the distribution of workload was more uniform than in the Zagreb Department of Pathology. The average workload of both institutions was 4,562 KUs. CONCLUSION: KU activity index is a very useful method for assessing average pathologists' workload. It may be also used in hospital administration for predicting changes in service and number of working pathologists. PMID- 11259744 TI - Intradural disc herniation at the T1-T2 level. AB - Intradural disc herniations comprise 0.26-0.30% of all herniated discs. Five percent are found in the thoracic, 3% in the cervical, and 92% in the lumbar region. Although intradural disc herniation may be suspected on preoperatively made CT scans, myelograms, and MRI scans, establishing the diagnosis prior to the surgery is difficult. We present a case of the patient with severe neurological deficits, caused by intradural thoracic disc herniation at T1-T2 interspace, which required surgical treatment. The symptoms were relieved immediately after surgery. This is the first description of an intradural disc herniation at that level. PMID- 11259745 TI - Primary echinococcosis of the sternocleidomastoid muscle. AB - Muscular echinococcosis accounts for 0.5% to 5.4% of all hydatid disease cases, with very little data on the incidence of muscular echinococcosis of the head and neck. We report a unique case of primary echinococcosis of the right sternocleidomastoid muscle in a 56-year-old man. Preoperative assessment by ultrasound and fine needle aspiration did not point to echinococcosis. We suspected the right diagnosis intraoperatively and confirmed it postoperatively by pathohistology and serologic tests. Echinococcosis of the liver and the lungs was also excluded postoperatively. Combination of operative treatment and postoperative albendazole herapy in two 28-day cycles one month apart resulted in complete regression of the disease. Echinococcosis should be considered as differential diagnosis of a multicystic mass in neck, particularly if it is of longstanding duration. Serologic tests for echinococcosis should be included in differential diagnostic procedures for each multicystic formation on the neck, especially in endemic areas. PMID- 11259746 TI - Heart metastasis of extraskeletal myxoid chondrosarcoma. AB - Isolated, remote heart metastasis of myxoid chondrosarcoma is extremely rare. We present an unusual case of isolated remote metastasis of extraosseus myxoid chondrosarcoma from the right ankle region to the right ventricle, its clinical course, and treatment in a 46-year-old woman. Although heart biopsy done before the surgery revealed myxoma, histopathologic diagnosis of the heart tumor was confirmed after its surgical resection from the right ventricle. Nine months after the surgery the patient was doing well but, seven months later, she died in the local hospital because of global heart failure. On the autopsy, the only metastatic lesion found was in the heart. The pericardium and heart muscle were infiltrated with the tumor, whereas all other organs were free from malignant dissemination. PMID- 11259747 TI - Humane warfare. PMID- 11259748 TI - From Gulf War Syndrome to Balkan War Syndrome. PMID- 11259750 TI - Current utilization trends for beta-blockers in cardiovascular disease. AB - Beta blockers have repeatedly demonstrated their therapeutic value in the treatment of a variety of diseases; as a result, multiple treatment guidelines advocate the use of beta blockers. Despite these guidelines, the use of beta blockers is remarkably low. Numerous factors that influence the trends of drug use include pharmaceutical advertisements, physician legal concerns, marketing influences, outdated therapeutic contraindications, and patient and physician demographics. Recent primary evidence from randomized clinical trials has demonstrated a significant benefit to patients with heart failure when beta blocker therapy is added to standard therapy. To ensure proper treatment, continuing efforts must be made to provide patients with appropriate therapy that is proven to reduce the risks of mortality and morbidity. PMID- 11259751 TI - Clinical trials of beta-blockers in heart failure: a class review. AB - Heart failure remains a clinically challenging illness, with increasing incidence and prevalence and a high risk of mortality. The introduction of agents that interfere with the neurohormonal response to chronic left-ventricular dysfunction has resulted in improved patient outcomes. Owing to slowed disease progression and reduced mortality, angiotensin-converting enzyme (ACE) inhibitors are indicated in all patients with heart failure. New data indicate that in appropriate patients, beta-blocker therapy relieves the symptoms associated with heart failure, reduces hospitalizations, and improves survival when added to standard therapy. Questions still remain regarding the ideal use of beta blockers in heart failure, and ongoing trials will attempt to clarify those points. PMID- 11259752 TI - Optimizing the use of beta-blockers in the effective treatment and management of heart failure: a case study approach. AB - Today, heart failure is an increasing concern in the United States. Its prognoses are poor and its treatment is a complicated endeavor, because heart failure is not a single disease state. Rather, it is a syndrome with a cyclic pathophysiology composed of multiple mechanisms. Effective case management of heart failure must address each of the many changes involved in this syndrome, and therapy must be individualized, especially because patients with heart failure often require regimens of five or more drugs. In special populations, such as the elderly and/or patients with concomitant diseases requiring added medication, polypharmacy becomes an important issue. Maintaining consistent compliance with the treatment regimen and patient education regarding symptoms of fluid retention can be critical. Currently, beta-blockers, in addition to standard therapy, are recommended as first-line treatment in mild-to-moderate heart failure. The three cases presented in this article illustrate some common scenarios encountered and clinical decisions made when beta-blockers are used in the management of heart failure. PMID- 11259754 TI - Gender-specific aging effects on the serotonin 1A receptor. AB - The effects of age on serotonergic function have been hypothesized to underlie age-related changes in mood and behaviors such as sleep and eating. Of particular interest is the serotonin type-1A (5-HT1A) receptor, due to its putative role in mediating the therapeutic efficacy of antidepressant treatment. Using positron emission tomography (PET) and [11C--carbonyl] WAY100635, we assessed 5-HT1A receptor binding in 21 healthy subjects (10 men, 11 women) ranging in age from 21 to 80 years. Regional binding potential values were generated using a reference tissue model and corrected for partial volume effects. We observed an inverse relationship between age and binding of [11C--carbonyl] WAY100635 to the 5-HT1A receptor in men, but not women. This finding is in accord with observations reported in the postmortem literature. Gender-specific effects of age on central serotonergic function may relate to differences between men and women in behavior, mood, and susceptibility to neuropsychiatric disease across the adult lifespan. PMID- 11259755 TI - Further characterisation of a thromboembolic model of stroke in the rat. AB - We have used magnetic resonance imaging (MRI) techniques to characterise a rat model of thromboembolic stroke. The consequences of acute perfusion deficit associated with a middle cerebral artery occlusion (MCAo) by a newly formed thrombus was mapped by interrogation of the tissue oxygenation status using gradient echo methods and production of T2* maps. Final infarct size was subsequently assessed at 24-h post-ischaemia by histology with 2,3,5 triphenyltetrazolium chloride (TTC) staining. Animals displayed an infarct volume of 178.7+/-84.2 mm(3) (mean+/-S.D.) with a large coefficient of variation (47%) and range of values (85.6--265.5 mm(3)). This variability provided us with an opportunity to assess the relationships between early imaging observations and eventual infarct size. For a single cerebral slice, at the centre of the MCA territory, a relationship between the area of reduced T2* at 1 and 2 h post MCAo correlated highly with final lesion area (Spearman rank correlation, r=0.98, P<0.01, n=9). Lesion volumes in the thromboembolic MCAo model were compared with a 120-min occlusion, 22-h reperfusion protocol using an intraluminal thread MCAo approach. For the thromboembolic model, the total lesion volume was found to be smaller (178.7+/-84.2 vs. 243.3+/-50.1 mm(3), mean+/-S.D., Student's t-test P=0.046) and showed a greater variability (coefficient of variations: 47% vs. 21%). These data underline the relative variability of this embolic model and provide important preliminary information regarding the value of early changes in T2* in predicting eventual infarct size. PMID- 11259753 TI - Chronic intrathecal morphine administration produces homologous mu receptor/G protein desensitization specifically in spinal cord. AB - Previous studies have shown that chronic i.v. treatment with morphine or heroin decreased mu opioid receptor activation of G-proteins in specific brain regions. The present study examined the effect of intrathecal (i.t.) morphine administration on receptor/G-protein coupling in the spinal cord. In spinal cord membranes, [35S]GTP gamma S binding was stimulated by agonists of several G protein-coupled receptors, including mu opioid (DAMGO), delta opioid (DPDPE), GABA(B) (baclofen), cannabinoid CB(1) (WIN 55,212-2), muscarinic cholinergic (carbachol) and adenosine A(1) (PIA). [35S]GTP gamma S autoradiography revealed that most of this agonist activation of G-proteins was localized to laminae I and II of dorsal horn. To determine the effects of chronic morphine on these receptor activities, rats were treated for 7 days with 0.11 mg/kg/day i.t. morphine, and receptor activation of G-proteins was determined by [35S]GTP gamma S autoradiography of brain and spinal cord. In spinal cord sections, chronic morphine treatment decreased DAMGO-stimulated [35S]GTP gamma S binding in laminae I and II at all levels of spinal cord examined. There were no effects of morphine treatment on [35S]GTP gamma S stimulation in spinal cord by other receptor systems examined (Adenosine A(1) and GABA(B)), and no significant effects of chronic i.t. morphine treatment were observed in brain sections. These data show that homologous desensitization of mu receptor/G-protein coupling occurs specifically in spinal cord following chronic morphine administration. PMID- 11259756 TI - Adrenalectomy causes loss of zinc ions in zinc-enriched (ZEN) terminals and decreases seizure-induced neuronal death. AB - Chelatable zinc ions from synaptic vesicles have been suggested to be involved in neuronal death caused by stroke, epilepsy and head trauma. Elevated glucocorticoid concentration exacerbates such neuron loss, while low levels protect. We have tested the notion that the neuroprotective effect of prior glucocorticoid reduction is mediated by a reduction of zinc ions contained in zinc-enriched (ZEN) synaptic vesicles. The level of vesicular zinc ions was evaluated by toluene sulfonamide quinoline (TSQ) fluorometry and zinc autometallography (ZnS(AMG)) 10 and 30 days, respectively, after adrenalectomy. The hippocampus showed significant vesicular zinc ion depletion following adrenalectomy. After the kainate injection, adrenalectomized rats showed proconvulsive seizure behavior, i.e. shortened latency to seizure onset time and increased seizure score. Additionally they showed decreased hippocampal CA3 neuronal death as compared to control animals. The present data suggest that zinc ions released from damaged ZEN terminals are involved in seizure-induced neuronal death. PMID- 11259757 TI - Cardiorespiratory and metabolic responses to injection of bicuculline into the hypothalamic paraventricular nucleus (PVN) of conscious rats. AB - Stimulation of the PVN increases mean arterial pressure (MAP) and heart rate (HR). However, little is known about its role in modulating ventilation. We tested the hypothesis that the stimulation of the PVN by microinjection of bicuculline methiodide (BMI), a gamma-aminobutyric acid (GABA)(A) receptor antagonist, increases ventilation in conscious rats. Oxygen consumption was also evaluated to determine if the ventilatory responses were associated with increases in metabolic rate. Male Sprague--Dawley rats were instrumented with femoral catheters to measure MAP and HR and cannulae were implanted 1 mm above the PVN. After 5 to 7 days of recovery, metabolic, ventilatory, and cardiovascular responses to artificial cerebrospinal fluid (aCSF) and BMI were evaluated. Rats were given a 50 nl unilateral microinjection of aCSF (the vehicle control) followed by 50 n1 of BMI (1 mM) into the other side. Microinjection of BMI significantly increased MAP compared to aCSF (145+/-4 vs. 124+/-5 mmHg, P<0.02), HR to 460+/-17 from 362+/-22 breaths/min (P<0.01). Ventilation increased by 300% (P=0.01) by stimulating frequency of breathing (176+/-14 compared to 79+/ 12 breaths/min, P<0.005) and increasing tidal volume. Concomitantly, O(2) consumption doubled (P<0.006). These data suggest that in the PVN GABA receptors may be important regulators of cardiopulmonary and metabolic function in conscious rats. PMID- 11259758 TI - Propagation of synchronous epileptiform events from subiculum backward into area CA1 of rat brain slices. AB - The hippocampal trisynaptic pathway is comprised of superficial entorhinal afferents (part of the perforant path) to dentate granule cells, dentate mossy fiber inputs to CA3 pyramidal neurons, and CA3 cell projections to CA1 pyramidal neurons. This CA1 output is among others to the subiculum, and both CA1 and subiculum project to the entorhinal cortex to close the loop. Smaller circuits involving fewer hippocampal and parahippocampal regions have also been described. We present morphological and electrophysiological evidence from rat brain slices for a projection from subiculum back into area CA1. Axons of neurobiotin-labeled subicular pyramidal neurons were visualized in the apical dendritic region of CA1. Spontaneous activity in isolated subiculum--CA1 slices was produced by bathing slices in reduced magnesium media. Events in CA1 always followed events in proximal subiculum. Disruption of this subiculum--CA1 circuit with a radially oriented knife cut in the apical dendritic region between subiculum and CA1 eliminated afterdischarges in subicular and CA1 events, but did not de synchronize the two regions. Full transections between CA1 and subiculum were necessary to functionally isolate the two regions. Only subiculum remained spontaneously active. We conclude that a subiculum--CA1 circuit supports afterdischarges in both regions and synchronizes their activity. This circuit may serve to maintain a level of depolarization in subicular and CA1 pyramidal neurons well beyond the duration of excitatory synaptic potentials resulting from activation of the trisynaptic circuitry. PMID- 11259759 TI - Effects of hypothermia on thrombin-induced brain edema formation. AB - Recent studies have shown that thrombin plays an important role in brain edema formation after intracerebral hemorrhage (ICH). The possible mechanisms of thrombin-induced brain edema formation include blood-brain barrier (BBB) disruption and inflammatory response involving polymorphonuclear (PMN) leukocyte. Animal experiments have revealed that moderate therapeutic hypothermia improves pathological and functional outcome in various models of brain injury. In this study, we examined the effect of hypothermia on thrombin-induced brain edema formation. Effects of hypothermia on BBB permeability and the accumulation of PMN leukocytes were also determined to clarify the protective mechanism of hypothermia in this model. Anesthetized adult rats received an injection of 10 Units of thrombin into the basal ganglia. Animals were separated into the normothermic and hypothermic groups, which were housed in a room maintained at 25 degrees C and in a cold room maintained at 5 degrees C, respectively, for 24 h after the thrombin injection. The brain temperature in rats housed in a cold room reduced temporarily to approximately 30 degrees C and then gradually recovered to 35 degrees C by the end of the observation. Brain water content in the basal ganglia was significantly reduced in rats treated with hypothermia compared to the normothermic rats (84.3+/-0.2 vs. 82.4+/-0.1%; P<0.01). The decrease of brain water content was accompanied with a significant reduction in BBB permeability to Evan's blue dye and in accumulation of PMN leukocytes. This study indicates that hypothermic treatment significantly reduces thrombin-induced brain edema formation in the rat. Inhibition of thrombin-induced BBB breakdown and inflammatory response by hypothermia appear to contribute to brain protection in this model. Hypothermic treatment may provide an approach to potentially reduce ongoing edema after ICH. PMID- 11259760 TI - Lipopolysaccharide-induced ischemic tolerance is associated with increased levels of ceramide in brain and in plasma. AB - Intravenous administration of lipopolysaccharide (LPS) (0.9 mg/kg) has been shown to induce ischemic tolerance in spontaneously hypertensive rats (SHR). TNF-alpha is believed to play a crucial role in preconditioning as its inhibition with TNF alpha-binding protein abolished tolerance. Our recent studies (Liu et al., Am. J. Physiol. 278 C144, 2000) have demonstrated that ceramide, a downstream messenger in TNF-alpha signaling, is a mediator of hypoxia-induced tolerance in neuronal cells. To test the hypothesis that ceramide contributes to LPS-induced tolerance in vivo, SHR were injected intravenously with either LPS or saline and the levels of ceramide in brain and in plasma were determined by reversed phase HPLC. LPS injection resulted in a significant increase of ceramide in plasma with a maximum at 24 h (8.32+/-1.14 pmol/microl (LPS) vs. 2.65+/-0.62 pmol/microl (saline)). LPS also induced ceramide upregulation in brain cortex, which started between 6 and 12 h and remained elevated up to 48 h after LPS injection. Fluorescent NBD-C6 ceramide was able to cross blood-brain barrier and was found in brain vessels, perivascular cells and in brain parenchyma 30 min after intravenous injection. These findings demonstrate that LPS preconditioning leads to elevation of ceramide in brain and plasma and, in conjunction with previous work, suggests that ceramide plays a role in LPS-induced protection against brain ischemic injury in vivo. PMID- 11259761 TI - Oxidative stress induced by ascorbate causes neuronal damage in an in vitro system. AB - Of particular physiological interest, ascorbate, the ionized form of ascorbic acid, possesses strong reducing properties. However, it has been shown to induce oxidative stress and lead to apoptosis under certain experimental conditions. Ascorbate in the brain is released during hypoxia, including stroke, and is subsequently oxidized in plasma. The oxidized product (dehydroascorbate) is transported into neurons via a glucose transporter (GLUT) during a reperfusion period. The dehydroascorbate taken up by cells is reduced to ascorbate by both enzymatic and non-enzymatic processes, and the ascorbate is stored in cells. This reduction process causes an oxidative stress, due to coupling of redox reactions, which can induce cellular damage and trigger apoptosis. Ascorbate treatment decreased cellular glutathione (GSH) content, and increased the rates of lipid peroxide production in rat cortical slices. Wortmannin, a specific inhibitor of phosphatidylinositol (PI)-3-kinase (a key enzyme in GLUT translocation), prevented the ascorbate induced-decrease of GSH content, and suppressed ascorbate induced lipid peroxide production. However, wortmannin was ineffective in reducing hydrogen peroxide (H(2)O(2))-induced oxidative stress. The oxidative stress caused ceramide accumulation, which was proportionally changed with lipid peroxides when the cortical slices were treated with ascorbate. These differential effects support the hypothesis that GLUT efficiently transports the dehydroascorbate into neurons, causing oxidative stress. PMID- 11259762 TI - N-cadherin expression in motoneurons is directly regulated by androgens: a genetic mosaic analysis in rats. AB - We have recently reported that systemic androgens regulate adult N-cadherin (N cad) expression in spinal motoneurons. However, the mechanism through which androgen mediates this effect remains undetermined. Androgen may act directly on motoneurons to regulate N-cad expression, or indirectly, via effects on androgen sensitive afferent or efferent structures. Here, we describe a genetic mosaic investigation of this site-of-action indeterminacy. Following developmental random X chromosome inactivation, androgenized female rats heterozygous for the tfm androgen receptor mutation (X(WT)X(tfm)) are phenotypic mosaics of androgen sensitive wild-type (WT) and androgen-insensitive (tfm) motoneurons. We compared steroid effects on WT and tfm cells in two sexually-dimorphic motoneuron pools, the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN), as well as a less steroid responsive motoneuron pool, the sexually monomorphic retrodorsolateral nucleus (RDLN). Independent of steroid treatment, a greater proportion of wild-type cells were N-cad immunoreactive (IR) in the DLN and RDLN. Following testosterone treatment, increased N-cad expression was observed in both cell types in the DLN, but in the SNB only the androgen competent WT cells increased N-cad expression. Testosterone treatment did not significantly alter N-cad expression in the mosaic RDLN. The results indicate both cell autonomous and cell non-autonomous androgenic regulation of N-cad expression in spinal motoneurons. PMID- 11259763 TI - Alpha-adrenergic agonists inhibit the dipsogenic effect of angiotensin II by their stimulation of atrial natriuretic peptide release. AB - Angiotensin II (ANG-II) and atrial natriuretic peptide (ANP) have opposing actions on water and salt intake and excretion. Within the brain ANP inhibits drinking induced by ANG-II and blocks dehydration-induced drinking known to be caused by release of ANG-II. Alpha-adrenergic agonists are known to release ANP and antagonize ANG II-induced drinking. We examined the hypothesis that alpha agonists block ANG-II-induced drinking by stimulating the release of ANP from ANP secreting neurons (ANPergic neurons) within the brain that inhibit the effector neurons stimulated by ANG-II to induce drinking. Injection of ANG-II (12.5 ng) into the anteroventral region of the third ventricle (AV3V) at the effective dose to increase water intake increased plasma ANP concentrations (P<0.01) within 5 min. As described before, previous injection of phenylephrine (an alpha(1) adrenergic agonist) or clonidine (an alpha(2)-adrenergic agonist) into the AV3V region significantly reduced ANG-II-induced water intake. Their injection also induced a significant increase in plasma ANP concentration and in ANP content in the olfactory bulb (OB), AV3V, medial basal hypothalamus (MBH) and median eminence (ME). These results suggest that the inhibitory effect of both alpha adrenergic agonists on ANG-II-induced water intake can be explained, at least in part, by the increase in ANP content and presumed release from these neural structures. The increased release of ANP from the axons of neurons terminating on the effector neurons of the drinking response by stimulation of ANP receptors would inhibit the stimulatory response evoked by the action of ANG-II on its receptors on these same effector neurons. PMID- 11259764 TI - Carboxyl terminal peptides derived from prepro-orphanin FQ/nociceptin (ppOFQ/N) are produced in the hypothalamus and possess analgesic bioactivities. AB - Orphanin FQ/nociceptin (OFQ/N), the endogenous ligand for the ORL-1/KOR-3 receptor, produces a wide variety of behavioral responses. Its precursor protein, prepro-OFQ/N (ppOFQ/N) contains several series of amino acids bounded by pairs of basic amino acids, raising the possibility that additional functional neuropeptides could be generated by proteolytic posttranslational processing. Several of these processing products have been shown to have pharmacological activity, including the 17 amino acid peptide OFQ/N (mppOFQ/N(140-157)) which is a major product of this precursor in the hypothalamus. Here we have used a newly developed radioimmunoassay and RP-HPLC to detect mppOFQ/N(160-187) in mouse hypothalamic extracts. Murine ppOFQ/N(160-187) has potent analgesic activity supraspinally (3.4 nmol, i.c.v.) and spinally (4.3 nmol, i.t.). This analgesic activity was reversed by the opioid antagonist naloxone (5 mg/kg, s.c.) and kappa(1)-selective opioid antagonist nor-BNI (60 microg, i.c.v.), despite the inability of ppOFQ/N(160-187) to compete binding in mu, delta, kappa(1), kappa(3), or OFQ/N binding assays. These findings suggest that murine ppOFQ/N(160 187) may be a physiologically relevant neuropeptide with a novel mechanism of action. PMID- 11259765 TI - Co-localization of AMPA-type glutamate receptor immunoreactivity in neurons of the rat subthalamic nucleus. AB - In order to determine the precise cellular localization of the alpha-amino-3 hydroxy-5-methyl-4-isoxazole-propionate (AMPA)-type glutamate receptor subunit immunoreactivity in the rat subthalamic nucleus, single and double immunofluorescence was performed. Intense level of GluR1, GluR2, GluR2/3 and GluR4 immunoreactivity was found in almost all neurons of the subthalamic nucleus. By double immunofluorescence, the subthalamic neurons in the same sections that displayed a strong immunoreactivity for GluR1 were found to display a robust GluR2 immunoreactivity and the subthalamic neurons that displayed GluR2 immunoreactivity were also found to express GluR4 immunoreactivity. The present results thus demonstrate that individual neurons of the subthalamic nucleus are likely to co-express GluR1 and GluR2, and GluR2 and GluR4 immunoreactivity. The native AMPA channels in the subthalamic neurons may, therefore, be composed of heteromeric subunits. The present results provide information of the neuroanatomical localization of AMPA receptor subunits in neurons of the subthalamic nucleus. The localization of AMPA receptor subunits may be related to functional characteristics of AMPA channels in the subthalamic neurons. PMID- 11259766 TI - Intracerebroventricular carbachol induces FOS immunoreactivity in lamina terminalis neurons projecting to the supraoptic nucleus. AB - Central application of the non-selective cholinergic receptor agonist, carbachol, induces water intake, vasopressin (VP) release and an acute increase in arterial blood pressure. Forebrain sites, particularly those located along the lamina terminalis (LT) (i.e. the subfornical organ (SFO), organum vasculosum (OV) and the median preoptic nucleus (MePO)) and in the hypothalamus, have been proposed as the major targets for producing the effects induced by intracerebroventricular (i.c.v.) carbachol injections. However, the functional and neuroanatomical relationship among carbachol-activated cells along the LT and hypothalamic areas such as the supraoptic nuclei (SON), is unclear. The present study investigated the i.c.v. carbachol-induced activity of the soma of LT projections which descend from the SFO, OV and MePO and terminate in the region of the SON. Cells along the LT were retrogradely labeled from SON-targeted injections of fluoro-gold, and FOS immunoreactivity (FOS-ir) was used to assess activation. A significant number of cells in the SFO, OV and MePO were double-labeled for both FOS-ir and fluoro gold. The FOS labeling in the cells of the LT-associated structures was significantly reduced by pretreatment with the i.c.v. muscarinic antagonist, atropine. Taken together, the results indicate that neurons located in structures located along the LT and projecting to the region of the SON are activated by i.c.v. carbachol and that these receptors are likely to be muscarinic. PMID- 11259767 TI - In vivo PDGF beta receptor activation in the dorsocaudal brainstem of the rat prevents hypoxia-induced apoptosis via activation of Akt and BAD. AB - Activation of platelet-derived growth factor receptor beta (PDGFR) within the caudal brainstem modulates the hypoxic ventilatory response. Since hypoxia does not induce apoptosis in the caudal brainstem, PDGFR could underlie such protective mechanism via a PI3 kinase-dependent phosphorylation of both Akt and BAD pathways. To further study this issue, caudal brainstem lysates were harvested from Sprague--Dawley rats during hypoxia (10% O(2)) after treatment with either vehicle or CGP 57148B (100 mg/kg), a selective blood-brain barrier permeable PDGFR antagonist. Time-dependent increases in phosphorylated Akt occurred during hypoxia, peaking at 45' and lasting for up to 6 h, without parallel changes in total Akt protein. CGP 57148B attenuated Akt activation at all time points. Similarly, phosphorylation of BAD at serine136 but not at serine 112 occurred in the caudal brainstem as early as 15' of hypoxia, and was completely blocked by CGP 57148B. Furthermore, CGP 57148B treatment elicited significant increases in single-stranded DNA, caspase-like activity, and cleaved caspase 3 after 24 h of hypoxia that were absent in the caudal brainstem of hypoxic vehicle-treated animals. We conclude that PDGFR-dependent in vivo activation of both Akt and BAD during hypoxia prevents induction of apoptosis, and may contribute to the increased hypoxic tolerance of brainstem neurons. PMID- 11259768 TI - Demonstration of a functional motilin receptor in TE671 cells from human cerebellum. AB - BACKGROUND: Our laboratory has described the presence of motilin receptors in the rabbit cerebellum. We discovered its presence in the human TE671 cell line, which is of cerebellar origin. METHODS: Cytosolic Ca(2+) fluxes were monitored on a confocal microscope in cells loaded with Indo-1 and stimulated with motilin under various conditions. Binding studies were performed with 125I-[Nle(13)]porcine motilin. Using primers, PCR for the motilin receptor was performed. RESULTS: Cells responded to motilin after 45+/-20 s. At different concentrations of motilin (10(-8), 10(-7), 10(-6.5), 10(-6) and 10(-5) M) the percentage of responding cells was 0+/-0, 0.6+/-1.5, 4.9+/-4.7, 21.7+/-15 and 35.7+/-12, respectively. The response was blocked by the motilin antagonists [Phe(3), Nle(13)]po-motilin (0.8+/-1.8%) and GM-109 (0.0+/-0.0%) and mimicked by the agonist ABT-229 (23.6+/-15%). After stimulation with motilin, ABT-229 or [Phe(3),Leu(13)]po-motilin, but not with the antagonist GM-109, cells were desensitized. The response to motilin persisted in Ca(2+)-free solution (22.8+/ 14.7%), was not affected by nifedipine (44+/-11%) but was abolished by incubation with thapsigargin (0+/-0%). Neither ryanodine, nor a previous stimulation with caffeine (0+/-0%) in Ca(2+)-free Krebs, nor both could block the response to motilin (28, 32.0+/-5.7, 41.3+/-6.1%, respectively). Binding studies revealed two binding sites for motilin, with a pK(d) of 8.9+/-0.05 and 6.11+/-0.61 (n=4). There were 100 times more low than high affinity receptors per cell. The presence of receptor mRNA was confirmed by PCR. CONCLUSION: Functional motilin receptors are present in TE671 cells. The response requires intracellular IP(3)-sensitive Ca(2+) stores. These cells may serve as a model of the central motilin receptor. PMID- 11259769 TI - Regional distribution of benzodiazepine binding sites in the human newborn and infant hypothalamus. A quantitative autoradiographic study. AB - Using in vitro quantitative autoradiography and [3H]flunitrazepam we examined the rostrocaudal distribution of benzodiazepine binding sites in the human neonate/infant hypothalamus. The autoradiographic analysis shows the presence of a heterogeneous distribution throughout the rostrocaudal extent of this brain structure. High [3H]flunitrazepam binding corresponds primarily to the diagonal band of Broca and the preoptic region. The labelling in the preoptic region showed a rostrocaudal increase, contrasting in that with the other hypothalamic structures. Intermediate densities were present in the septohypothalamic, suprachiasmatic, periventricular and paraventricular nuclei as well as in the mammillary complex. Low binding was observed in the other hypothalamic structures. The benzodiazepine binding sites analyzed belong mostly to type II receptors. In an attempt to unravel possible differences related to age, we compared the autoradiographic distribution in three postnatal age ranges. The topographical distribution of these binding sites was almost identical in each period analyzed. We found, however, that benzodiazepine binding is generally low in the neonatal period and a tendency in increasing densities is observed during development. Taken together, these results provide evidence for a large distribution of benzodiazepine binding sites in neonate/infant hypothalamus, suggesting their implication in the development of this brain structure and the maintenance of its various functions. PMID- 11259770 TI - Retinal projection to the dorsal raphe nucleus in the Chilean degus (Octodon degus). AB - A substantial projection from the retina to the dorsal raphe nucleus (DRN) has been demonstrated in the Chilean degus, a diurnal/crepuscular hystricomorph rodent. Following intraocular injection of cholera toxin subunit B (CTB), immunocytochemically labeled CTB-positive axons and terminals were observed in all major retinorecipient nuclei as well as in the DRN and periaqueductal gray (PAG) of the mesencephalon. Two streams of optic axons to the DRN were observed: one descending from the optic tract at the level of the pretectum and anterior superior colliculus, the other emerging as a small fascicle at the anterior pole of the inferior colliculus and descending bilaterally through the PAG. Contralateral retinal afferents in the DRN appeared to terminate primarily in the dorsomedial and lateral subdivisions of the DRN, and a less extensive ipsilateral component also was observed. Axonal arborizations were characterized by short branches and multiple varicosities, both in the DRN and in the PAG. The extent and density of DRN retinal afferents were not as extensive as previously observed in Mongolian gerbils using identical techniques, but the retinal-DRN projection is considerably larger in degus than in rats. The functional significance of the retinal-DRN pathway remains to be determined, although a variety of evidence indicates that light may directly affect the activity of neurons and serotonin levels in the DRN. PMID- 11259771 TI - Central suppressive effect of octreotide on the hyperglycemic response to 2-deoxy D-glucose injection or cold-swim stress in awake rats: possible mediation role of hypothalamic noradrenergic drive. AB - Somatostatin (SRIH) and its analog have been reported to act within the central nervous system to suppress the hyperglycemic response to a variety of neural stimuli. On the other hand, the hyperglycemic response to 2-deoxy-D-glucose (2 DG) injection or cold-swim stress is well demonstrated to be closely associated with an increase in hypothalamic noradrenergic neuronal activity (NNA). To evaluate whether the suppression of the hypothalamic NNA response could be involved in the central mechanism whereby a SRIH analog inhibits the hyperglycemic response, octreotide, a clinically used long-acting octapeptide SRIH analog, was administered into the third cerebral ventricle of awake rats prior to the intraperitoneal injection of 2-DG or cold-swim stress. Hypothalamic noradrenaline (NA) and its neuronal metabolite, 3,4-dihydroxyphenylethyleneglycol (DHPG), were analyzed, and the ratio of DHPG to NA was used as an index of NNA. Intracerebroventricular (i.c.v.) pretreatment with octreotide suppressed the 2-DG induced increase in hypothalamic NNA, accompanied by the inhibition of the serum glucose, NA and adrenaline responses. This suppressive effect of octreotide was dose-dependent. Similarly, i.c.v. pretreatment with octreotide prevented the hypothalamic NNA response to cold-swim stress, accompanied by a blockade of the increases in serum glucose, NA and adrenaline. A close relationship between hypothalamic NNA and serum glucose emerged from these studies. Intraperitoneal pretreatment with octreotide had no significant effect on the hyperglycemic or hypothalamic NNA response to 2-DG injection. These findings suggest that the inhibitory effect of octreotide on the hypothalamic NNA response to 2-DG injection or cold-swim stress is associated with the simultaneous suppression of the hyperglycemic response. Supporting the concept that hypothalamic NNA contributes to the modulation of blood glucose in stressful conditions, it is suggested that the suppression of the hypothalamic NNA response is, at least in part, involved in the central mechanism by which octreotide inhibits the hyperglycemic response to 2-DG injection or cold-swim stress. PMID- 11259772 TI - Activation of p44/42 mitogen activated protein kinases in thrombin-induced brain tolerance. AB - BACKGROUND: Our recent studies have shown that prior intracerebral injection of a low dose of thrombin attenuates the brain edema formation that results from either an intracerebral hematoma, an intracerebral injection of a large dose of thrombin or cerebral ischemia. The aim of the current study is to investigate whether thrombin-induced tolerance (thrombin preconditioning; TPC) is associated with activation of p44/42 mitogen activated protein (MAP) kinases. METHODS: This study contained three parts. In the first, rats received an intracerebral infusion of either saline or one unit thrombin (the TPC dose) into the right caudate nucleus. After 1, 3 and 7 days, the rats will be killed and brains used to detect p44/42 MAP kinases activation using Western blot analysis and immunohistochemistry. In the second and third parts, rats received intracerebral infusions of either vehicle, one unit thrombin (TPC) or one unit thrombin and 5 nmol PD 098059. These rats were either killed to detect kinases activation after 24 h or received a second intracerebral infusion of five-unit thrombin 7 days later with brain edema being assessed after a further 24 h. RESULTS: Western blot analysis demonstrated that p44/42 MAP kinases were activated in the ipsilateral basal ganglia after the intracerebral infusion of thrombin one unit. Cells immunoreactive for activated p44/42 MAP kinases were found in the ipsilateral basal ganglia and ipsilateral cortex. PD 098059, a MAP kinase kinase inhibitor, abolished thrombin-induced activation of p44/42 MAP kinases. TPC suppressed thrombin-induced brain edema while PD 098059 blocked this protective effect. The water contents in the ipsilateral basal ganglia 24 h after infusion of thrombin five units were 82.6+/-0.8%, 79.2+/-0.4% and 81.8+/-1.9% in the control, TPC alone and TPC plus PD 098059 groups, respectively. CONCLUSION: Thrombin can activate p44/42 MAP kinases within the brain and the protective effects of thrombin preconditioning on brain edema formation are related to this activation. PMID- 11259774 TI - Involvement of the paraventricular nucleus of the hypothalamus in the pressor response to chemoreflex activation in awake rats. AB - Chemoreflex activation with potassium cyanide (KCN, i.v.) produces pressor and bradycardic responses in awake rats in addition to the tachypneic response. In the present study we evaluated the role of the paraventricular nucleus of the hypothalamus (PVN) in the cardiovascular responses to chemoreflex activation in awake rats. Bilateral electrolytic lesion of the PVN was performed 1 day before chemoreflex activation and the results were compared to those obtained with sham lesioned rats. Bilateral electrolytic lesion of the PVN (n=6) produced a significant reduction in both the magnitude (+51+/-5 vs. +22+/-2 mmHg) and duration (+26+/-5 vs. +6+/-2 s) of the pressor response to chemoreflex activation when compared to sham-lesioned rats (n=10). The bradycardic response to chemoreflex activation in rats with bilateral lesion of the PVN was not significantly different from the response of sham-lesioned rats (-229+/-20 vs. 88+/-76 bpm). Unilateral or partial bilateral lesion of the PVN (n=10) produced no significant changes in the pressor response (+51+/-5 vs. +49+/-3 mmHg), in the duration of the response (+26+/-5 vs. +18+/-3 s) or in the bradycardic response ( 229+/-20 vs. -230+/-27 bpm) compared to sham-lesioned rats. The data show that effective bilateral lesion of the PVN produced a significant reduction in the magnitude and duration of the pressor response, indicating that the PVN plays a key role in the processing of the sympathoexcitatory component of the chemoreflex. PMID- 11259773 TI - Characterization of melanin concentrating hormone and preproorexin expression in the murine hypothalamus. AB - Melanin concentrating hormone (MCH) and the orexins (A and B) have been identified as neuropeptides localized to the lateral hypothalamic area (LHA) and are potential regulators of energy homeostasis. Potential factors regulating expression of both MCH and the orexins include fasting and leptin. Previous studies have generated conflicting data and, as there is little leptin receptor expressed in the lateral hypothalamus, it is likely that any observed leptin effects on these peptides are indirect. In this study, we examined MCH and preproorexin expression in mice in physiological states of starvation, with or without leptin administration, in addition to characterizing MCH and preproorexin expression in well-known obesity models, including ob/ob and UCP-DTA mice. Neuropeptide Y (NPY) expression in the arcuate nucleus was used as a positive control. After a 60-h fast, expression of both NPY and MCH mRNA was increased (by 148 and 33%, respectively) while preproorexin expression in the murine LHA did not change. Leptin administration to fasted mice blunted the rise in MCH and NPY expression towards control levels. In contrast, there was a 78% increase in preproorexin expression in fasted mice in response to peripheral leptin administration. MCH expression was increased (by 116%) in ob/ob mice at baseline, as we have previously reported. In addition, leptin treatment of ob/ob mice blunted the increase in MCH expression. In contrast, preproorexin expression did not differ in the leptin-deficient ob/ob mice or in the obese hyperleptinemic brown adipose tissue deficient (UCP-DTA) mice in comparison with controls. In summary, MCH expression is increased in two states of decreased leptin, fasting and ob/ob mice, and leptin replacement blunts MCH expression in both paradigms. Thus, MCH expression appears to be regulated by leptin. In contrast, preproorexin expression does not respond acutely to fasting, although it is acutely increased by leptin treatment during fasting. These preproorexin responses are in contrast to those seen with well-characterized orexigenic neuropeptides, such as NPY and AgRP, suggesting that appetite regulation may not be a significant physiological role of orexins. This conclusion is further supported by the observation that orexin ablated mice have arousal and not feeding deficits. PMID- 11259775 TI - Immunohistochemical study on the distribution of six members of the Kv1 channel subunits in the rat cerebellum. AB - Voltage-gated K(+) (Kv) channels are critical for a wide variety of processes, and play an essential role in neurons. In the present study, we have demonstrated a unique pattern of expression of the six Kv1 channel subunits in the rat cerebellum, for the first time. The greatest concentration of Kv1.2 was found in the basket cell axon plexus and terminal regions around the Purkinje cells. Relatively weak immunoreactivity for Kv1.1 was also found in this area. The somatodendritic Purkinje cell areas were intensely stained with anti-Kv1.5 antibodies. In the cerebellar nuclei, the cell bodies of cerebellar output neurons showed strong Kv1.5 and Kv1.6 immunoreactivities in the nucleus medialis, interpositus and lateralis. Interestingly, Kv1.2 immunoreactivity was found in some neurons with their processes. Our immunohistochemical results may support the notion that the formation of heteromultimeric Kv channels possibly represents an important contribution to the generation of Kv channel diversity in the brain, especially in the cerebellum. PMID- 11259776 TI - Characterization of the glutamatergic system for induction and maintenance of allodynia. AB - Glutamate is the main excitatory neurotransmitter in the central nervous system and has been shown to be involved in spinal nociceptive processing. We previously demonstrated that intrathecal (i.t.) administration of prostaglandin (PG) E(2) and PGF(2 alpha) induced touch-evoked pain (allodynia) through the glutamatergic system by different mechanisms. In the present study, we characterized glutamate receptor subtypes and glutamate transporters involved in induction and maintenance of PGE(2)- and PGF(2 alpha)-evoked allodynia. In addition to PGE(2) and PGF(2 alpha), N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl 4-isoxazolepropionic acid (AMPA), but not kainate, induced allodynia. PGE(2)- and NMDA-induced allodynia were observed in NMDA receptor epsilon 4 (NR2D) subunit knockout (GluR epsilon 4(-/-)) mice, but not in epsilon 1 (NR2A) subunit knockout (GluR epsilon 1(-/-)) mice. Conversely, PGF(2 alpha)- and AMPA-induced allodynia were observed in GluR epsilon 1(-/-) mice, but not in GluR epsilon 4(-/-) mice. The induction of allodynia by PGE(2) and NMDA was abolished by the NMDA receptor epsilon 2 (NR2B) antagonist CP-101,606 and neonatal capsaicin treatment. PGF(2 alpha)- and AMPA-induced allodynia were not affected by CP-101,606 and by neonatal capsaicin treatment. On the other hand, the glutamate transporter blocker DL-threo-beta-benzyloxyaspartate (DL-TBOA) blocked all the allodynia induced by PGE(2), PGF(2 alpha), NMDA, and AMPA. These results demonstrate that there are two pathways for induction of allodynia mediated by the glutamatergic system and suggest that the glutamate transporter is essential for the induction and maintenance of allodynia. PMID- 11259777 TI - Uteroplacental insufficiency lowers the threshold towards hypoxia-induced cerebral apoptosis in growth-retarded fetal rats. AB - Infants suffering uteroplacental insufficiency and hypoxic ischemic injury often demonstrate cerebral apoptosis. Our objective was to determine the global effects of uteroplacental insufficiency upon cerebral gene expression of the apoptosis related proteins Bcl-2 and Bax and their role in increasing vulnerability to hypoxia-induced cerebral apoptosis. We therefore caused uteroplacental insufficiency and growth retardation by performing bilateral uterine artery ligation upon pregnant rats 2 days prior to term delivery and elicited further perinatal fetal hypoxia by placing maternal rats in 14% FiO(2) 3 h prior to delivery. We quantified cerebral levels of Bcl-2 and Bax mRNA, lipid peroxidation, caspase-3 activity, and cAMP in control and growth retarded term rat pups that experienced either normoxia or hypoxia. Uteroplacental insufficiency alone caused a significant decrease in cerebral Bcl-2 mRNA levels without altering cerebral Bax mRNA levels, malondialdehyde levels, or caspase-3 activity. In contrast, uteroplacental insufficiency and subsequent fetal hypoxia significantly increased cerebral Bax mRNA levels, lipid peroxidation and caspase 3 activity; Bcl-2 mRNA levels continued to be decreased. Hypoxia alone increased cerebral cAMP levels, whereas uteroplacental insufficiency and subsequent hypoxia decreased cerebral cAMP levels. We speculate that the decrease in Bcl-2 gene expression increases the vulnerability towards cerebral apoptosis in fetal rats exposed initially to uteroplacental insufficiency and subsequent hypoxic stress. PMID- 11259778 TI - A beta-fiber intensity stimulation of chronically constricted median nerve induces c-fos expression in thalamic projection neurons of the cuneate nucleus in rats with behavioral signs of neuropathic pain. AB - This study was aimed to investigate the possible involvement of neurons in the cuneate nucleus (CN) in the processing of A beta afferent inputs evoked by electrical stimulation of constricted median nerve in rats with behavioral signs of neuropathic pain. Immunohistochemical localization of Fos protein was used to examine the neuronal activation, and the combination of Fos immunohistochemistry with the retrograde labeling of Fluoro-Gold (FG) injected into the ventrobasal complex of the thalamus was used to characterize the activated neurons. Two weeks after unilateral median nerve constriction injury, the rats exhibited behavioral signs of neuropathic pain in the affected forepaws. In rats after nerve injury but without electrical stimulation, some Fos-like immunoreactive (Fos-LI) neurons were detected in the dorsal horn of the seventh cervical segment (C7) but none was found in the CN. Similar features were also noted when the stimulation of the intact median nerve served as an additional control. After A beta-fiber intensity stimulation of the previously constricted median nerve, an increase in number of Fos-LI neurons occurred in the medial half of the ipsilateral C7 dorsal horn as well as in the ipsilateral CN. In the latter, the Fos-LI neurons were located in the median nerve projection territory throughout the nucleus. Most of the Fos-LI neurons were distributed in the middle region of the CN, with about 78% of them emitting FG fluorescence indicating that they were cuneothalamic projection neurons. The results of this study suggest that the dorsal column-medial lemniscal system may contribute to the transmission and modulation of A beta fiber mediated neuropathic pain signals. PMID- 11259779 TI - Sympathetic sprouting in the dorsal root ganglion after spinal nerve ligation: evidence of regenerative collateral sprouting. AB - It is well documented that there is an increase in the number of sympathetic fibers within the dorsal root ganglion (DRG) after a peripheral nerve injury. The present study examined the numbers and distribution of sympathetic fibers in the DRG and their sprouting routes by utilizing various surgical manipulations and retrograde tracing and immunohistochemical staining methods in spinal nerve ligated neuropathic rats. The appearance of many double immunostained fibers with antibodies to tyrosine hydroxylase (TH) and growth associated protein-43 (GAP-43) in the L5 DRG 1 week after L5 spinal nerve ligation, indicated sprouting of sympathetic fibers. The confined location of early sprouting sympathetic fibers in the distal half of the L5 DRG confirmed that sprouting fibers come primarily from the injured spinal nerve. A second cut proximal to the previously ligated L5 spinal nerve -- a process which would transect the regenerating sympathetic fibers extending from the injury site -- did not change the density of sympathetic fibers in the L5 DRG. When retrograde tracers (fast blue and diamidino yellow) were injected into the L5 spinal nerve and DRG, respectively, the number of double-labeled sympathetic postganglionic neurons was greatly increased after spinal nerve ligation, suggesting the increased number of sympathetic neurons projecting to both the spinal nerve and DRG. All these results indicate that many sympathetic fibers in the DRG are regenerating branches that are sprouting from the proximal part of the injured spinal nerve (regenerative collateral sprouting). PMID- 11259780 TI - The organisation of spinal projecting brainstem neurons in an animal model of muscular dystrophy. A retrograde tracing study on mdx mutant mice. AB - Previous studies we performed on the mdx mouse demonstrated marked central nervous system alterations in this model of human Duchenne muscular dystrophy, such as reduction in number and pathological changes of cortico-spinal neurons. Prompted by these findings we extended the survey of the mdx brain to the major brainstem-descending pathways: the rubro-, vestibulo-, reticulo-, and raphe spinal projections. Horseradish peroxidase microinjections were performed in the cervical spinal cord of mdx and control mice. The rubro-spinal neurons were found to be significantly reduced in mutants compared to controls. The vestibulo spinal, reticulo-spinal, and raphe-spinal cell populations, though less numerous in mdx than in control mice, were instead substantially spared. Our data further unveil the selective nature of mdx brain damage indicating a marked and selective involvement of the highest centers for motor control. PMID- 11259781 TI - Effects of pituitary adenylate cyclase activating polypeptide on lumbosacral preganglionic neurons in the neonatal rat spinal cord. AB - The effects of PACAP-38 on phasic and tonic preganglionic neurons (PGN) in L6 and S1 spinal cord slices from neonatal rats (5--11 days old) were studied using the whole-cell patch clamp technique. PGN were identified by retrograde axonal transport of a fluorescent dye (Fast Blue, 5 microl of 4% solution) injected into the intraperitoneal space 3--7 days prior to the study. Bath application of pituitary adenylate cyclase activating polypeptide (PACAP) (20 nM) increased the frequency of spontaneous excitatory postsynaptic potentials (EPSPs) and spontaneous firing in both types of PGN. PACAP markedly increased the number (200 -800%) and frequency of action potentials elicited by depolarizing current pulses in phasic PGN, but had a smaller effect on tonic PGN. PACAP decreased the threshold for action potential generation by approximately 25% in both types of neurons (e.g. -34.0+/-1.5 to -38.4+/-1.7 mV from a holding potential of -50 mV in phasic PGN, P<0.005). PACAP did not affect the duration of the action potential. The amplitude of the spike after hyperpolarization was not changed but the duration was significantly reduced by PACAP from 204.4+/-12.2 to 106.2+/-8.1 ms in tonic but not in phasic PGN. PACAP suppressed a transient outward current that was also suppressed by 4-aminopyridine (0.5 mM). These results coupled with the immunohistochemical identification of a dense collection of PACAP fibers in the region of the PGN, raises the possibility that PACAP may function as an excitatory transmitter in lumbosacral parasympathetic reflex pathways in the neonatal rat. PMID- 11259782 TI - Increased levels of calcium-sensitive adenylyl cyclase subtypes in the limbic system of alcoholics: evidence for a specific role of cAMP signaling in the human addictive brain. AB - We examined the amounts of several adenylyl cyclase (AC) isoforms and of cAMP response element binding protein (CREB) in alcoholic and control brains. Immunoreactivity of type I AC was significantly increased in alcoholic nucleus accumbens and corpus amygdaloideum. Immunoreactivity of type VIII AC was also increased in alcoholic corpus amygdaloideum and hippocampus. CREB immunoreactivities were unchanged. These findings indicate that the brain-region specific increase of Ca(2+)-sensitive AC may contribute to the pathophysiology of alcoholism. PMID- 11259783 TI - Characteristic changes in T(2)-value, apparent diffusion coefficient, and ultrastructure of substantia nigra evolving exofocal postischemic neuronal death in rats. AB - To correlate the magnetic resonance (MR) imaging characteristics of exofocal postischemic neuronal death (EPND) in the substantia nigra (SN) with associated histologic changes, we occluded the left middle cerebral artery of rats for 1, 4, 7, or 12 days. Day 1 (post-occlusion) T(2)-weighted images revealed high signal intensity indicative of infarction in the ipsilateral caudate nucleus, putamen, and cortex but not the SN. Diffusion-weighted images (DWIs) on day 1 similarly failed to reveal any changes in the SN. However, on day 4, DWIs revealed high signal intensity in the ipsilateral SN, in which the apparent diffusion coefficient (ADC) transiently decreased (P<0.05) while the T(2)-value increased (P<0.05). These measures returned to and remained at control levels on days 7 and 12. Histologic examination on day 4 revealed dark-staining neurons, markedly swollen perivascular astrocytic end-feet, many swollen neurons with cytoplasmic microvacuoles that mainly originated in the rough endoplasmic reticulum, and strongly roughed neuropils. Reactive astrocytes and dark neurons most frequently appeared on days 7 and 12. The severity of cellular swelling paralleled the change in the ADC. These results demonstrate that a transient high-intensity signal on DWIs, indicative of a decrease in the ADC, is predictive of EPND in the SN. PMID- 11259784 TI - Dopamine induced protein damage in mitochondrial-synaptosomal fraction of rat brain. AB - Dopamine during in vitro oxidation induced covalent cross-linking of membrane proteins in rat brain crude mitochondrial-synaptosomal fraction. The process is not inhibited by hydroxyl radical scavengers, lipid soluble anti-oxidants, metal chelator or catalase, but reduced glutathione produced a dramatic inhibition of cross-linking. The protein cross-linking mediated by dopamine is not associated with any detectable membrane lipid peroxidation but significant formation of protein bound quinone takes place during incubation. Our results indicate that reactive quinones rather than oxygen free radicals are involved in dopamine induced protein cross-linking in rat brain membrane fraction. PMID- 11259785 TI - Unlike 2-deoxy-D-glucose, 3-O-methyl-D-glucose does not induce long-term potentiation in rat hippocampal slices. AB - Equimolar replacement of 10 mM glucose by 2-deoxy-D-glucose (2-DG) causes substantial depression followed by a sharp and sustained potentiation of CA1 field EPSPs. In the present experiments, similar applications of 3-O-methyl-D glucose, which is also taken up by cells but is not phosphorylated, had only a weak blocking action and elicited no potentiation. Possible explanations for the marked effects of 2-DG include a more rapid block of glycolysis and the production of phosphorylated derivatives of 2-DG. PMID- 11259786 TI - Expression of multidrug resistance protein 1 (MRP1) in the rat cochlea with special reference to the blood-inner ear barrier. AB - Expression of multidrug resistance protein 1 (MRP1) was detected in the rat cochlea by RT-PCR and immunohistochemistry using anti-MRP monoclonal antibody MRPr1. Use of primers specific for rat mrp1 gene resulted in the amplification of an expected 394 bp fragment prepared from brain and cochlear tissues. Immunohistochemically, MRP was found in the choroid plexus, stria vascularis, spiral ligament, spiral prominence and cochlear nerve in the modiolus. From these results, it was suggested that MRP in the rat cochlea might function as an extrusion pump and play an important role in the blood-inner ear barrier. PMID- 11259787 TI - The synaptic vesicle priming protein Munc13-1 is absent from tonically active ribbon synapses of the rat retina. AB - Ribbon synapses, for example of the retina, are specialized synapses that differ from conventional, phasically active synapses in several aspects. Ribbon synapses can tonically and yet very rapidly release neurotransmitter via synaptic vesicle exocytosis. This requires an optimization of the synaptic machinery and is at least partly due to the presence of synaptic ribbons that bind large numbers of synaptic vesicles and which are believed to participate in priming synaptic vesicles for exocytosis. In this paper we analyzed whether ribbon synapses of the retina employ similar priming factors, i.e. Munc13-1, as do conventional, non ribbon containing phasically active synapses. We found that though present in conventional synapses of the retina Munc13-1 was completely absent from ribbon containing synapses of the retina, both in the outer as well as in the inner plexiform layer. This indicates that ribbon synapses of the retina employ other, possibly more potent priming factors than phasically active conventional synapses. PMID- 11259788 TI - A comparative evaluation of the neurotoxic properties of ketamine and nitrous oxide. AB - The general anesthetics, nitrous oxide (N(2)O) and ketamine, are NMDA antagonists which, like other NMDA antagonists such as MK801, induce a neurotoxic reaction in the rat brain. For MK801 neurotoxicity, both age and sex are important variables (adult rats are more sensitive than immature rats and females are more sensitive than males). In this study we found that ketamine has this same age and sex dependency profile, and N(2)O has the same age but not sex dependency. Male and female rats are equally sensitive to N(2)O neurotoxicity. PMID- 11259789 TI - Blockade of lactate transport exacerbates delayed neuronal damage in a rat model of cerebral ischemia. AB - Studies over the past decade have demonstrated that lactate is produced aerobically during brain activation and it has been suggested to be an obligatory aerobic energy substrate postischemia. It has been also hypothesized, based on in vitro studies, that lactate, produced by glia in large amounts during activation and/or ischemia/hypoxia, is transported via specific glial and neuronal monocarboxylate transporters into neurons for aerobic utilization. To test the role of lactate as an aerobic energy substrate postischemia in vivo, we employed the cardiac-arrest-induced transient global cerebral ischemia (TGI) rat model and the monocarboxylate transporter inhibitor alpha-cyano-4-hydroxycinnamate (4-CIN). Once 4-CIN was establish to cross the blood--brain barrier, rats were treated with the inhibitor 60 min prior to a 5-min TGI. These rats exhibited a significantly greater degree of delayed neuronal damage in the hippocampus than control, untreated rats, as measured 7 days post-TGI. We concluded that intra ischemically-accumulated lactate is utilized aerobically as the main energy substrate immediately postischemia. Blockade of lactate transport into neurons prevents its utilization and, consequently, exacerbates delayed ischemic neuronal damage. PMID- 11259790 TI - Effects of rat ventral and dorsal hippocampus temporal inactivation on delayed alternation task. AB - To determine the involvement of the hippocampal regions in a operant-type delayed alternation task of short delay or long delay, microinjections of muscimol into the hippocampus were used for temporal inactivation during the behavioral test in each rat. Dorsal hippocampal inactivation impaired the correct ratio of long delay. Ventral hippocampal inactivation showed no changes in the correct ratio, however, it increased the tendency of perseveration in long delay. These findings suggest hippocampus has regional differentiation in delayed alternation task. PMID- 11259791 TI - Synthetic antigenic decapeptides of human brain acetylcholinesterase cross immunoreact with peptide-specific antibodies against Torpediniformes narcine timlei acetylcholinesterase. AB - Antigenic decapeptides of human brain acetylcholinesterase (AChE) were investigated for immunoreactivity with the rabbit anti-Torpediniformes narcine timlei AChE polyclonal antibody (anti-narcine AChE polyclonal antibody). The decapeptides were synthesized using the multipin combinatorial chemical synthesis technique and biotinylated at N-terminals. Rabbit anti-narcine AChE polyclonal antibodies were purified by Protein A-Sepharose CL 4B column chromatography. Enzyme-linked immunosorbent assay (ELISA) was used for the assay of the reaction between the antigen and the antibody. Seven of 11 antigenic synthetic decapeptides of human brain AChE showed obvious immunoreactivity with the rabbit anti-narcine AChE polyclonal antibodies. The similarity of the AChE sequences of humans and Torpedo species were compared thereby with the epitopes indicated. The results indicate that the epitopes of human brain AChE and Torpedo AChEs have been highly conserved during evolution. In view of this, no N-glycosylation site was found in the antigenic decapeptides tested, they all belong to oligopeptide epitopes. PMID- 11259792 TI - Leptin transport at the blood--cerebrospinal fluid barrier using the perfused sheep choroid plexus model. AB - Leptin is secreted by adipose tissue and thought to regulate appetite at the central level. Several studies have explored the central nervous system (CNS) entry of this peptide across the blood-brain and blood-cerebrospinal fluid (CSF) barriers in parallel, but this is the first to explore the transport kinetics of leptin across the choroid plexus (blood-CSF barrier) in isolation from the blood brain barrier (BBB). This is important as the presence of both barriers can lead to ambiguous results from transport studies. The model used was the isolated Ringer perfused sheep choroid plexus. The steady-state extraction of [(125)I]leptin (7.5 pmol l(-1)) at the blood face of the choroid plexus was 21.1+/-5.7%, which was greater than extraction of the extracellular marker, giving a net cellular uptake for [(125)I]leptin (14.0+/-3.7%). In addition, trichloroacetic acid precipitable [(125)I] was detected in newly formed CSF, indicating intact protein transfer across the blood-CSF barrier. Human plasma concentrations of leptin are reported to be 0.5 nM. Experiments using 0.5 nM leptin in the Ringer produced a concentration of leptin in the CSF of 12 pM (similar to that measured in humans). [(125)I]Leptin uptake at the blood-plexus interface using the single-circulation paired tracer dilution technique (uptake in <60 s) indicated the presence of a saturable transport system, which followed Michaelis-Menten-type kinetics (K(m)=16.3+/-1.8 nM, V(max)=41.2+/-1.4 pmol min( 1) g(-1)), and a non-saturable component (K(d)=0.065+/-0.002 ml min(-1) g(-1)). In addition, secretion of new CSF by the choroid plexuses was significantly decreased with leptin present. This study indicates that leptin transport at the blood-CSF barrier is via saturable and non-saturable mechanisms and that the choroid plexus is involved in the regulation of leptin availability to the brain. PMID- 11259793 TI - Influence of beliefs about the consequences of dizziness on handicap in people with dizziness, and the effect of therapy on beliefs. AB - OBJECTIVE: To determine the longitudinal relationship between beliefs about the consequences of dizziness and handicap levels in dizzy patients, and the effect of therapy on beliefs. METHODS: Symptoms, beliefs, and handicap were assessed at baseline and 6 months follow up in 76 primary care patients complaining of dizziness or vertigo, of whom 33 were assigned to treatment (i.e., vestibular rehabilitation). RESULTS: At baseline most patients believed that dizziness would have negative consequences such as falling, fainting, or losing control. Handicap levels at follow-up were predicted by baseline beliefs that dizziness would have negative consequences. Significant reduction in negative beliefs at follow-up was observed in the patients who received treatment, whereas there was no reduction in negative beliefs in the untreated patients. CONCLUSIONS: Negative beliefs about the consequences of dizziness sustain long-term restriction of activity, and can be modified by therapy. PMID- 11259794 TI - Similar psychological characteristics in mild and severe asthma. AB - OBJECTIVE: Psychological factors have been implicated as potentially contributing to asthma severity. In the present study, we investigated whether patients with mild and severe asthma differ with regard to several psychological characteristics. METHODS: Ninety outpatients with severe asthma (74% female, mean [S.D.] age: 46.5 [13.7] years) and 37 outpatients with mild asthma (73% female, age: 39.4 [13.9] years) were compared with respect to general psychological health, anxiety sensitivity, hyperventilation symptoms, personality, and locus-of control orientation, all measured by well-validated self-report questionnaires. Analysis of (co)variance (ANCOVA) was used to assess between-groups differences. RESULTS: No significant differences in psychological characteristics were found between patients with mild and severe asthma. Only on the subscale for external locus-of-control orientation, severe asthmatic patients differed from those with mild disease (P=.005) in showing less trust in physicians and medication with regard to influencing their asthma. CONCLUSION: The results suggest that mild and severe asthmatic patients cannot be differentiated on the basis of psychopathology or personality. Whether or not the observed lack of confidence in the influence of physicians or medication on asthma course is cause or consequence of disease severity, remains to be established. PMID- 11259795 TI - The contribution of perceptions of stigmatisation to disability in patients with psoriasis. AB - OBJECTIVE: The aim of the present study was to assess the significance of general and psoriasis specific psychological variables in patients with psoriasis and to examine the relative importance of disease status and these psychological variables in predicting psoriasis-related disability. METHOD: A total of 115 patients with psoriasis underwent clinical assessment and completed a number of psychological and psoriasis specific questionnaires. RESULTS: High levels of self reported distress were identified with 43% and 10% of patients scoring as probable cases on the Hospital Anxiety and Depression Scale (HADS) subscales of anxiety (mean 9.3+/-4.9) and depression (mean 4.8+/-3.7), respectively. Multiple regression analysis indicated that clinical severity of psoriasis and anatomical area of involvement had no impact on psychological distress and disability. Perceptions of stigmatisation were significantly related to both psychological distress and degree of disability (P's<.001) and accounted for a significant amount of the variance in disability over and above general psychological distress (F change=11.03; P<.001). CONCLUSION: Psychological factors were much stronger determinants of disability in patients with psoriasis than disease severity, location or duration. This has important implications in relation to the clinical management of psoriasis. PMID- 11259796 TI - Voluntary motor function in patients with chronic fatigue syndrome. AB - INTRODUCTION: The pathogenesis of chronic fatigue syndrome (CFS) remains unknown. In particular, little is known of the involvement of the motor cortex and corticospinal system. METHODS: Transcranial magnetic stimulation (TMS) was used to assess corticospinal function in terms of latency and threshold of motor evoked potentials (MEPs) in thenar muscles. Reaction times and speed of movement were assessed using button presses in response to auditory tones. RESULTS: Patients had higher (P<.05) self-assessed indices of fatigue (7/10) than for pain (5/10), anxiety (4/10) or depression (3/10). Mean (+/-S.E.M.) simple reaction times (SRTs) were longer (P<.05) in the patients (275+/-19 ms) than in the controls (219+/-9 ms); choice reaction times (CRTs) were not significantly longer in the patients. Movement times, once a reaction task had been initiated, were longer (P<.05) in the patients in both SRTs (patients, 248+/-13 ms; controls, 174+/-9 ms) and CRTs (patients, 269+/-13 ms; controls, 206+/-12 ms). There was no difference (P>.05) in threshold or latency of MEPs in hand muscles between the patients (threshold, 54.5+/-2.2% maximum stimulator output [% MSO]; latency 22+/ 0.3 ms) and controls (threshold 54.6+/-3.6% MSO; latency 22.9+/-0.5 ms). Regression analysis showed no correlation (P>.05) of SRTs with either threshold for MEPs or fatigue index. CONCLUSION: Corticospinal conduction times and excitability were within the normal range despite a slower performance time for motor tasks and an increased feeling of fatigue. This suggests that the feeling of fatigue and the slowness of movement seen in CFS are manifest outside the corticospinal system. PMID- 11259798 TI - The relation between work-induced neuroendocrine reactivity and recovery, subjective need for recovery, and health status. AB - OBJECTIVES: The purpose of this cross-sectional study with repeated measurements was to find out to what extent neuroendocrine reactivity during work and neuroendocrine recovery from work, and work characteristics, are related to subjective need for recovery and perceived health status. METHODS: Neuroendocrine reactivity and recovery were studied in 59 subjects by measuring urinary adrenaline and cortisol repeatedly during five consecutive days. Measures of work characteristics, subjective need for recovery, and health status were obtained by self-reports. Two hierarchical multiple linear regression analyses were performed. RESULTS: The work characteristics alone explained 39% and 28% of the variation in subjective need for recovery and health status, respectively, while these figures rose to 49% and 53% in the full models. Significant neuroendocrine contributors were found for cortisol in reactivity during work and recovery immediately after work and recovery during the day off-work, and for adrenaline in baseline level and recovery during the day off-work. Job characteristics contributed significantly as well. CONCLUSION: Both neuroendocrine measures and work characteristics were predictors for the amount of perceived need for recovery after work as well as for health status. The results are consistent with the cognitive activation theory of stress. PMID- 11259797 TI - Psychological job demands as a risk factor for common cold in a Dutch working population. AB - OBJECTIVE: We investigated the effect of Psychological Job Demands (PJD) on the occurrence of the clinical symptoms of common cold. METHODS: Subjects, participating in a large prospective cohort study on psychological determinants of fatigue at work, were asked to fill in a questionnaire on the occurrence of common cold during the previous four months. High PJD were considered as a potential risk factor. Other factors such as age, gender, and having young children were considered as potential confounders. RESULTS: In logistic regression analysis, the adjusted odds ratio (OR) for having a recent cold in subjects reporting high PJD vs. those reporting low PJD was 1.20 (95% confidence interval (CI), 1.08-1.33). A higher risk emerged among those with young children (OR, 1.70; 95% CI, 1.47-1.96), those having a history of asthma (OR, 1.69; 95% CI, 1.28-2.22), or being under the age of 40 (OR, 1.28; 95% CI, 1.14-1.43) and among smokers (OR, 1.23; 95% CI, 1.09-1.38). CONCLUSION: The results support an association between PJD and common cold. In spite of the almost inevitable shortcoming of a large cohort study using questionnaires, this study gave us the opportunity to study the relationship between common cold and work-related factors in a nonexperimental setting with participants observed in a natural environment with all the normal everyday hassles. PMID- 11259799 TI - Mental disorders and medical conditions. A community study in a small island in Spain. AB - OBJECTIVES: This study focuses on the comorbidity between medical and psychiatric illnesses in a sample taken from the general population. METHODS: We design a two stage epidemiological community study in the island of Formentera (Spain), using the Schedules for Clinical Assessment in Neuropsychiatry (SCAN). In the first stage, the sample (N=697) was screened using the General Health Questionnaire (GHQ-28). In the second stage, the SCAN, subjects' clinical histories and General Practitioners' diagnosis were used to evaluate psychiatric disorders and medical illnesses. The final sample comprised 242 subjects. RESULTS: Subjects with diagnosed ICD-10 mental disorders presented higher comorbidity with medical pathologies than the group of respondents without psychiatric diagnosis, although the differences were not statistically significant. Of the psychiatric conditions, mood disorders (in which differences were significant) and neurotic disorders were the ones that most frequently presented comorbidity, followed by sleep disorders. Mean scores on the GHQ-28 used at the screening stage, and mean scores on neurotic, depressive and somatic symptoms on the SCAN, were significantly higher in the comorbid group. Comorbidity was more frequent in females. CONCLUSION: More studies on comorbidity in general population are required, with diagnostic instruments that cover the whole of the psychiatric spectrum and correct assessment systems for medical diagnosis. PMID- 11259800 TI - Salivary cortisol levels and anxiety are not increased in women destined to develop preeclampsia. AB - OBJECTIVE: To compare salivary cortisol levels and maternal anxiety (general and pregnancy-specific) in the early and late second trimester of pregnancy between women who developed preeclampsia (PE) and women who remained normotensive. DESIGN: Nested case-referent study. In a prospectively studied cohort of 250 pregnant women, nine women developed PE in late pregnancy. These nine patients were matched and compared with nine controls. Diurnal cortisol levels were obtained by collecting saliva samples at 17-18 and 27-28 weeks gestation. Salivary cortisol levels were determined by radioimmunoassay. Maternal anxiety was determined by Spielberger's State-Trait Anxiety Inventory (STAI) and a pregnancy-specific stress questionnaire. RESULTS: For both patients and controls, a similar pattern of salivary cortisol excretion was observed. Salivary cortisol levels and anxiety scores (general and pregnancy-specific) did not differ significantly between patients and controls. CONCLUSIONS: Our findings do not lend support to a role for maternal anxiety or second trimester increases in circulating stress hormones in the pathogenesis of PE. PMID- 11259801 TI - A mathematical model of in vitro cancer cell growth and treatment with the antimitotic agent curacin A. AB - A mathematical model of cancer cell growth and response to treatment with the experimental antimitotic agent curacin A is presented. Rate parameters for the untreated growth of MCF-7/LY2 breast cancer and A2780 ovarian cell lines are determined from in vitro growth studies. Subsequent growth studies following treatments with 2.5, 25 and 50 nanomolar (nM), concentrations of curacin A are used to determine effects on the cell cycle and cell viability. The model's system of ordinary differential equations yields an approximate analytical solution which predicts the minimum concentration necessary to prevent growth. The model shows that cell growth is arrested when the apoptotic rate is greater than the mitotic rate and that the S-phase transition rate acts to amplify this effect. Analysis of the data suggests that curacin A is rapidly absorbed into both cell lines causing an increase in the S-phase transition and a decrease in the M-phase transition. The model also indicates that the rate of apoptosis remains virtually constant for MCF-7/LY2 while that of A2780 increases 38% at 2.5 nM and 59% at 50 nM as compared to the untreated apoptotic rate. PMID- 11259802 TI - Simulation of the mechanism of determining the position of the cleavage furrow in cytokinesis of sea urchin eggs. AB - In cytokinesis of sea urchin eggs, the numerical density of astral microtubules extending close to the cell surface has been thought to determine the position of the cleavage furrow. In the present study, a new model was constructed to simulate the relationship between the microtubule density and the furrow formation. In the model, gradients of the microtubule density drive fluid membrane proteins whose accumulation triggers the formation of contractile-ring microfilaments. The model could explain the behavior of the cleavage furrow under various experimental conditions. These simulations revealed two aspects of furrow formation. One is that in some cases, the cleavage furrow appears in a surface region where the microtubule density has neither a minimum nor a maximum. In all furrow regions, however, the second derivative of the microtubule-density function has large positive values. Membrane proteins greatly slow down to accumulate in such a region. The other is that the cleavage furrow is mobile, not fixed in one position, because of the fluidity of membrane proteins. These results strongly suggested that the mitotic apparatus determines the position of the cleavage furrow by redistributing membrane proteins through gradients of the microtubule density at the cell surface. PMID- 11259803 TI - Waiting times to appearance and dominance of advantageous mutants: estimation based on the likelihood. AB - The germinal center reaction (GCR) of vertebrate immunity provides a remarkable example of evolutionary succession, in which an advantageous phenotype arises as a spontaneous mutation from the parental type and eventually displaces the parental type altogether. In the case of the immune response to the hapten (4 hydroxy-3-nitrophenyl)acetyl (NP), as with several other designed immunogens, the process is dominated by a single key mutation, which greatly simplifies the modeling of and analysis of data. We developed a two-stage model of this process in which the primary stage represents the appearance and establishment of the mutant population as a stochastic process while the second stage represents the growth and dominance of the clone as a deterministic process, conditional on its time of establishment from stage one. We applied this model to the analysis of population samples from several germinal center (GC) reactions and used maximum likelihood methods to estimate the waiting times to arrival and to dominance of the mutant clone. We determined the sampling properties of the maximum-likelihood estimates using Monte Carlo methods and compared them to their asymptotic distributions. The methods we present here are well-suited for use in the analysis of other systems, such as tumor growth and the experimental evolution of bacteria. PMID- 11259804 TI - A model of a snorer's upper airway. AB - In snorers, the physiologic decrease of postural muscle tone during sleep results in increased collapsibility of the upper airway. Measurement of nasal pressure changes with prongs is increasingly used to monitor flow kinetics through a collapsing upper airway. This report presents a mathematical model to predict nasal flow profile from three critical components that control upper airway patency during sleep. The model includes the respiratory pump drive, the stiffness of the pharyngeal soft tissues, and the dynamic support of the muscles surrounding the upper airway. Depending on these three components, the proposed model is able to reproduce the characteristic changes in flow profile that are clinically observed in snorers and non-snorers during sleep. PMID- 11259805 TI - Properties of phylogenetic trees generated by Yule-type speciation models. AB - We investigate some discrete structural properties of evolutionary trees generated under simple null models of speciation, such as the Yule model. These models have been used as priors in Bayesian approaches to phylogenetic analysis, and also to test hypotheses concerning the speciation process. In this paper we describe new results for three properties of trees generated under such models. Firstly, for a rooted tree generated by the Yule model we describe the probability distribution on the depth (number of edges from the root) of the most recent common ancestor of a random subset of k species. Next we show that, for trees generated under the Yule model, the approximate position of the root can be estimated from the associated unrooted tree, even for trees with a large number of leaves. Finally, we analyse a biologically motivated extension of the Yule model and describe its distribution on tree shapes when speciation occurs in rapid bursts. PMID- 11259806 TI - Life, quality of life and choice in an ageing society. PMID- 11259807 TI - Early expression of natriuretic peptides and SERCA in mild heart failure: association with severity of the disease. AB - BACKGROUND: We investigated changes in genetic expression of atrial and brain natriuretic peptides (ANP and BNP) and sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) in patients with stable mild to moderate chronic heart failure (CHF), since data on this topic were primarily obtained in end-stage CHF. METHODS: We studied tissue from 25 patients with idiopathic dilated cardiomyopathy (IDC) in New York Heart Association (NYHA) class II (n=12) and III-IV (n=13). Myocardial tissue from normal hearts (n=10) served as controls. Messenger RNA (mRNA) expression of ANP, BNP, and SERCA was isolated, and correlated with severity of CHF, left ventricular function (LVEF), peak oxygen uptake (peak VO(2)), and wedge pressure. RESULTS: A significant trend for gradual changes in mRNA expression according to increasing NYHA class was found for ANP, BNP (P<0.0001) and SERCA (P=0.04), with a marked increase in patients with more advanced CHF (ANP and BNP: P<0.01 vs. controls; SERCA: NS) and less pronounced changes in patients with mild CHF. mRNA of ANP and BNP correlated strongly with LVEF (-0.621 and -0.621, respectively, both P<0.01) and peak VO(2) (-0.625 and -0.555, respectively, both P<0.01) and, to a lesser extent, with wedge pressure (0.440 and 0.488, respectively, both P<0.05). SERCA correlated most strongly with wedge pressure ( 0.623, P<0.01), and weak, non-significant correlations with LVEF and peak VO(2) were found. CONCLUSIONS: Genetic expression of ANP, BNP, and SERCA is progressively altered in proportion to the severity of CHF, although this is more marked for ANP and to a lesser extent BNP, than for SERCA. These changes support the concept that already early in CHF, genetic expression is affected, which has implications for the understanding of the pathophysiology of CHF. PMID- 11259808 TI - The methylenetetrahydrofolate reductase gene polymorphism in Koreans with coronary artery disease. AB - Hyperhomocysteinemia is a known risk factor of cardiovascular diseases. Methylenetetrahydrofolate reductase (MTHFR), involved in folate-dependant metabolism, is associated with homocysteine levels. We studied the associations among MTHFR genotypes, coronary artery disease (CAD), and homocysteine levels in 85 patients with CAD and 152 healthy subjects. The MTHFR genotypes and plasma homocysteine levels were determined. No significant difference in mutation of the MTHFR gene between two groups was observed (P>0.05). While the homozygous mutant genotype (V/V) had the highest homocysteine levels compared to wild (A/A) and heterozygous mutant (A/V) genotypes, there were no significant differences in homocysteine levels among the MTHFR genotype groups. Homocysteine was significantly and inversely related to folate levels, the significant association in V/V genotype (beta coefficient=-1.954, P=0.04). Our data suggested that MTHFR polymorphism was not associated with homocysteine levels, implying no association between gene polymorphism and CAD in Koreans. PMID- 11259809 TI - Endothelial dysfunction in type 2 diabetes mellitus subjects with peripheral artery disease. AB - We strived to characterize the endothelial function status in type 2 diabetic patients with peripheral artery disease which was detected by ankle-brachial index by utilizing high frequency ultrasounds. Predictors of endothelial dysfunction were investigated. We chose 23 type 2 diabetic patients had ankle brachial index <0.97 (0.15-0.95; mean=0.74+/-0.20), 31 diabetic patients had ankle-brachial index >/=1.0 and 28 non-diabetic subjects for study. Older age, a longer duration of diabetes, higher systolic blood pressure, higher prevalence of history of hypertension were observed in patients with peripheral vascular disease. Type 2 diabetic patients showed impaired flow-mediated dilatation than non-diabetic and it showed more impaired in patients with peripheral vascular disease. Nitroglyerin-induced dilatation showed a trend of impairment in patients with peripheral vascular disease but did not reach statistical significance. Age (r=-0.259, P=0.019), baseline brachial artery diameter (r=-0.321, P=0.003), ankle brachial index (r=0.259, P=0.002) and hypertension history (P=0.01) were significantly associated with flow-mediated dilatation. However, after adjusting for age, only baseline diameter and ankle-brachial index were independent predictors of flow-mediated dilatation. In conclusion, we demonstrated flow mediated dilatation was impaired in type 2 diabetic patients and it was further impaired in patients with peripheral vascular disease. Nitroglycerin-induced dilatation showed a trend of impairment but did not reach statistical significance. PMID- 11259810 TI - Altered erythrocyte n-3 fatty acids in Mediterranean patients with coronary artery disease. AB - A low frequency of ischaemic heart diseases in Eskimos has been related to polyunsaturated fatty acids. We therefore studied fatty acid patterns associated with coronary artery disease (CAD) for a possible relationship between fatty acid profile and CAD diagnosis in Mediterranean patients. The gas chromatography method was used to analyze the membranes of patients' erythrocytes. The patients without coronary stenosis were used as controls. Patients with CAD showed increased percentages of saturated fatty acids (35.8 vs. 34.2%, P<0.001) and monounsaturated fatty acids (14.6 vs. 13.6%, P<0.01), as well as reduced percentages of polyunsaturated fatty acids (38.5 vs. 41.3%, P<0.001). The decrease in polyunsaturated fatty acids percentages was due to the series of n-3 fatty acids (9.2 vs. 11.4%, P<0.001), mainly at the expense of docosahexaenoic acid [C22:6 (n-3)] (4.9+/-0.25% vs. 6.4+/-0.23%, P<0.001) and docosapentaenoic acid [C22:5 (n-3)] (3.0+/-0.19% vs. 3.9+/-0.12%, P<0.001). The study shows altered n-3 fatty acids in Mediterranean patients with CAD. Our data suggest that the percentage of docosahexaenoic and docosapentaenoic acids in erythrocytes could be used as indicators of an independent risk factor for coronary artery disease. PMID- 11259812 TI - ST-segment elevation on Q-leads during exercise in patients with ST-segment elevation at rest after myocardial infarction. AB - INTRODUCTION: ST-segment elevation on Q-leads after an acute myocardial infarction is related to a greater infarct size. The meaning of a further exercise-induced ST-segment elevation in these patients has not been analyzed. METHOD: Thirty-six patients with ST-segment elevation on Q-leads were studied after a first acute myocardial infarction. Exercise testing and cardiac catheterization were performed at the first week. Left ventricular volumes (ml/m(2)); the extent of abnormal wall motion (AWM: chords); contractile reserve (AWM improvement with low dose dobutamine) and coronary patency in the culprit artery were analyzed. Cardiac catheterization was repeated at the sixth month in 20 patients; systolic recovery (AWM improvement), left ventricular volumes and coronary patency were again evaluated. RESULTS: Patients with exercise-induced ST segment elevation in two or more Q-leads (n=21) showed lesser contractile reserve (6+/-6 vs. 12+/-7 chords, P=0.01) than patients without exercise-induced ST segment elevation (n=13). AWM (F=8.1) and absence of exercise-induced ST-segment elevation (F=9.5; positive predictive value: 80%; negative predictive value: 68%) were the only independent predictors of contractile reserve. Nevertheless, this electrocardiographic sign was not related to left ventricular volumes, coronary patency or systolic function and it did not predicted late systolic recovery. CONCLUSIONS: In patients with baseline ST-segment elevation on Q-leads an exercise-induced ST-segment elevation is independently related to a lesser contractile reserve but not to the evolution of volumes or regional dysfunction during the first 6 months post-infarction. Therefore, the clinical value of this sign seems to be limited to the non-invasive detection of myocardial viability during the early post-infarction phase. PMID- 11259811 TI - High frequency of arrhythmias after Fontan operation indicates earlier anticoagulant therapy. AB - BACKGROUND: As patients began to survive for longer periods following modified Fontan operations (conventional atrio-pulmonary connection), the late morbidity after this procedure became increasingly apparent. The purpose of the present study was to evaluate late sequelae of modified Fontan operations in long-term survivors (n=14) at our institute. METHODS AND RESULTS: The cohort consisted of patients who underwent a modified Fontan operation between 1981 and 1990. Thus, all patients were examined at least 10 years postoperatively in this study. Early mortality, within 30 days of the operation, was 17.6% (three of 17 patients died from low output syndrome). Excluding these early deaths, the cumulative survival rate at 5 and 10 years was 100 and 79%, respectively. Arrhythmias including atrial fibrillation or flutter were the main late causes of morbidity. The arrhythmia-free rate at 5 and 10 years was 77 and 50%, respectively. Although the quality of life was considered good because all patients (n=11) who survived for 10 years or more were in class I or II according to the New York Heart Association classification, most of them in fact suffered from potentially life threatening arrhythmias. CONCLUSIONS: Meticulous attention to and utilization of established treatment strategies for arrhythmias including anti-arrhythmics, anticoagulants, catheter ablation or re-operation converting the circulation to the total cavopulmonary connection must be considered in long-term survivors following the modified Fontan operation. The fact that no one knows when the thrombogenic arrhythmias occur suggests anticoagulants should be initiated in the early postoperative period. PMID- 11259813 TI - A method for investigating the mechanical properties of intracoronary stents using finite element numerical simulation. AB - The proliferation of stent designs poses difficult problems to clinicians, who have to learn the relative merits of all stents to ensure optimal selection for each lesion, and also to regulatory authorities who have the dilemma of preventing the inappropriate marketing of substandard stents while not denying patients the benefits of advanced technology. Of the major factors influencing long-term results, those of patency and restenosis are being actively studied whereas the mechanical characteristics of devices influencing the technical results of stenting remain under-investigated. Each different stent design has its own particular features. A robust method for the independent objective comparison of the mechanical performance of each design is required. To do this by experimental measurement alone may be prohibitively expensive. A less costly option is to combine computer analysis, employing the standard numerical technique of the finite element method (FEM), with targeted experimental measurements of the specific mechanical behaviour of stents. In this paper the FEM technique is used to investigate the structural behaviour of two different stent geometries: Freedom stent geometry and Palmaz-Schatz (P-S) stent geometry. The effects of altering the stent geometry, the stent wire diameter and contact with (and material properties of) a hard eccentric intravascular lesion (simulating a calcified plaque) on stent mechanical performance were investigated. Increasing the wire diameter and the arterial elastic modulus by 150% results in the need to increase the balloon pressure to expand the stent by 10-fold. Increasing the number of circumferential convolutions increases the pressure required to initiate radial expansion of mounted stents. An incompressible plaque impinging on the mid portion of a stent causes a gross distortion of the Freedom stent and an hour-glass deformity in the P-S stent. These findings are of relevance for future comparative studies of the mechanical performance of stents, in designing newer stents and also in clinical practice. PMID- 11259814 TI - Increased plasma levels of soluble selectins in patients with unstable angina. AB - BACKGROUND: Inflammation plays an important role in the pathogenesis of unstable angina. Adhesion molecules, such as selectins, mediate the interactions between leukocytes, platelets and endothelial cells during inflammation and thrombogenesis. HYPOTHESIS: The purpose of this study was to determine whether soluble E-selectin, P-selectin and L-selectin levels are increased in patients with unstable angina (UA). METHODS: Soluble E-, P- and L-selectin levels were measured by enzyme-linked immunoassay in the peripheral blood of 23 patients with UA, 26 patients with stable angina (SA) and 15 control patients with angiographically normal coronary arteries. RESULTS: Soluble E-selectin levels were significantly higher in patients with UA (45+/-11 ng/ml) than in controls (30+/-8 ng/ml, P<0.001), or patients with SA (34+/-8 ng/ml, P=0.001). Similarly, plasma levels of P- and L-selectin were significantly higher in patients with UA (427+/-144 and 772+/-160 ng/ml, respectively) than in patients with SA (278+/-79 and 643+/-94 ng/ml, respectively, P<0.005 vs. UA for both), or control patients (189+/-43 and 601+/-126 ng/ml, respectively, P=0.001 vs. UA for both). CONCLUSIONS: Plasma levels of soluble selectins were increased in patients with UA compared with patients with SA or patients without angiographically visible coronary artery disease. Measurements of these adhesion molecules may be helpful as non-invasive markers of coronary plaque destabilization in UA. PMID- 11259816 TI - Relationship between mitral regurgitation and myocardial viability after acute myocardial infarction: their impact on prognosis. AB - Mitral regurgitation (MR) after acute myocardial infarction (AMI) is an important prognostic factor. Although its mechanisms are still debated, ventricular remodeling probably plays an important role. Because myocardial viability (MV) in the infarct zone reduces infarct expansion and ventricular remodeling, it is also possible that its presence counteracts the development of mitral regurgitation in infarcted patients. To evaluate this issue 191 patients with uncomplicated AMI, wall motion abnormalities (akinesis) and semiquantitative evaluation of MR were retrospectively selected from those consecutively examined at our echo-laboratory to evaluate MV using low-dose dobutamine echocardiography (DbE). Follow-up evaluation was performed at 30+/-13 months. Seventy-nine patients had no MR; 86 patients had grade 1 MR, 11 patients had grade 2 MR, nine patients had grade 3 MR, and six patients had grade 4 MR. Patients with significant MR (>grade 1) were older (63+/-7 vs. 59+/-10 years, P=0.03), had lower reduction of RWMSI (DeltaRWMSI) during DbE (0.08+/-0.11 vs. 0.22+/-0.28, P=0.01), more stenotic vessels at coronary angiography (2.35+/-0.93 vs. 1.67+/-1.12, P=0.01), and more frequently had anterior-inferior AMI (P<0.0001); they also had a non-significant tendency to higher RWMSI (2.04+/-0.38 vs. 1.92+/-0.28, P=0.06). In a multivariate regression analysis, DeltaRWMSI proved to be significantly related to the grade of MR (P=0.02). Eighteen patients died during follow-up. Death was more frequent in patients with MR (10/165 vs. 8/26, P=0.0003). At multivariate stepwise Cox regression analysis both the extent of ventricular dysfunction and the presence of MR were significantly related to mortality (P<0.0001 and P=0.01, respectively); DeltaRWMSI showed a non-significant tendency to influence mortality (P=0.09). When MR was excluded from the multivariate analysis, DeltaRWMSI remained significantly related to mortality (P=0.05). In conclusion our study suggests that the presence of MV in infarcted patients influences the development of MR. This reduction of MR may be one of the mechanisms by which MV affects mortality after AMI and should be considered in all studies that evaluate MV after myocardial infarction. PMID- 11259815 TI - No long-lasting or intermittent mast cell activation in acute coronary syndromes. AB - BACKGROUND: Unstable coronary syndromes, such as acute myocardial infarction and unstable angina pectoris are mostly due to rupture of an atherosclerotic plaque. Recently mast cells were found to participate actively in the inflammatory process of atherosclerosis by excreting proteolytic and pro-inflammatory substances with the ability to cause plaque instability and rupture. Mast cell activity can be determined by measuring serum levels of tryptase, as has been demonstrated in patients with anaphylaxis and mastcytosis. HYPOTHESIS: Acute coronary events (acute myocardial infarction and unstable angina pectoris) are associated with increased mast cell activity, reflected by elevated serum tryptase levels. METHODS: Serum levels of tryptase were determined in the following three groups of patients: 13 patients with acute myocardial infarction, 10 patients with unstable angina pectoris, and 14 patients without ischaemic cardiovascular disease who were used as controls. Patients with known IgE mediated allergic diseases and/or anti-histaminical drugs were excluded. RESULTS: The groups were comparable for sex, blood pressure, smoking and cholesterol levels. The controls tended to be younger (P=0.05). Levels of tryptase did not differ between patients with acute myocardial infarction (7.9+/-4.6 microg/l), unstable angina pectoris (6.0+/-2.1 microg/l) or controls (6.9+/-4.1 microg/l), nor could a relation with levels of C-reactive protein be demonstrated. CONCLUSION: Serum levels of tryptase are not elevated in patients with acute coronary syndromes. This implies that increased mast cell activity, if any, in unstable coronary syndromes is not reflected systemically. Other, more specific methods will be needed to determine the activity of the mast cell in vivo. PMID- 11259817 TI - Introduction to the special issue--Plenary Sessions of the Ninth Symposium of International Society of Veterinary Epidemiology and Economics (ISVEE). PMID- 11259818 TI - Can epidemiology and economics make a meaningful contribution to national animal disease control? AB - The general role of veterinary epidemiology and economics to national animal disease control throughout the world is considered for the four main groupings of animal diseases: zoonotic, food-borne, endemic and epidemic diseases. This is done by considering how veterinary epidemiology and economics has contributed to priority setting (which diseases come first?), decision-making (for a given disease, which strategy is best?), and disease control implementation (how can optimal delivery and adoption of selected interventions best be achieved?). Within each of these categories, progress made and future opportunities are discussed. In addition, a review is made of how veterinary epidemiology and economics has been institutionalised. We conclude that veterinary epidemiology and economics holds a unique role in the development of national policies and strategies for improved animal health world-wide. However, we consider that we must capitalise more on the unique comparative advantage of the partnership between veterinarians and agricultural economists. We believe that much remains to be done to improve the "institutionalisation" of veterinary epidemiology and economics, and the adoption and impact of the products of our unique partnership, particularly in countries of the developing world. PMID- 11259819 TI - International trade, animal health and veterinary epidemiology: challenges and opportunities. AB - The link between international trade, animal health and epidemiology has been recognized for a long time and has taken an additional importance in the aftermath of the Uruguay Round of Multilateral Trade Negotiations of the General Agreement on Tariffs and Trade (GATT) and of the inception of the World Trade Organization. The Agreement on the Application of Sanitary and Phytosanitary Measures of the World Trade Organization demands that sanitary and phytosanitary measures be scientifically based, placing epidemiology at the center of decisions related animal health and trade. This paper analyses the interactions between international trade of animals (and animal products) and epidemiology with discussion on the inputs of epidemiology in surveillance, risk analysis and regionalization. PMID- 11259820 TI - The role of veterinary epidemiology in the study of free-roaming dogs and cats. AB - Free-roaming dogs or cats are domestic dogs and cats that are not confined to a yard or house. Free-roaming dogs and cats have long caused major public-health problems and animal-welfare concerns in many countries. Free-roaming dogs have been considered to be more of a problem than cats for several reasons, but the literature addressing dogs focuses primarily on their role in rabies spread and control. Free-roaming cats are becoming more of an issue in countries where free roaming dog problems are coming under control. The change in perception of pets, beyond their value as a commodity, has also contributed to the increase in concern and attention focused on free-roaming dogs and cats. Epidemiologists have contributed much to these studies of these populations and have potential to contribute even more. The epidemiologic methods and approaches, the experience of epidemiologists in interdisciplinary teams and the importance of considering the separate sub-populations in study design and analysis all are critical in designing and evaluating interventions for free-roaming dogs and cats. In this paper, I will (1) describe a set of useful definitions regarding free-roaming dogs and cats, (2) summarize past and present topics of study in free-roaming dogs and cats, using selected examples, (3) describe the limitations of existing work and how epidemiologists might strengthen and improve this work, and (4) outline areas needing more attention by epidemiologists and why these are important. PMID- 11259821 TI - The role of epidemiology in the prevention, diagnosis, and control of infectious diseases of fish. AB - Epidemiologic methods are essential to understanding infectious diseases in aquaculture. Unfortunately, many of these methods are poorly understood or not utilized by fish-health scientists and aquaculturists -- often because of the lack of contact with epidemiologists who are willing to investigate fish diseases. In this paper, we describe direct interactions between epidemiologists and fish-health specialists that have resulted in an improved understanding of the causes and management of infectious diseases in aquaculture. We focus on risk factor studies, risk analysis and infectious-disease modeling, evaluation of diagnostic tests and experimental studies. We also describe characteristics of confined fish populations that make them ideal for developing and testing epidemiologic models and the theoretical and practical challenges of designing and conducting epidemiologic studies in fish farms. Throughout our presentation, emphasis is given to characteristics, opportunities and problems associated mainly with conducting epidemiologic studies to intensive aquaculture systems. We conclude that the development of increased cooperation among epidemiologists, fish-health scientists and aquaculturists will be mutually beneficial and, therefore, efforts for such collaboration should be initiated from all parties involved. PMID- 11259822 TI - Methods to investigate spatial and temporal clustering in veterinary epidemiology. AB - Due to their contagious or point-source nature, ill-health and diseases often cluster in time and/or space. Overlooking this characteristic can lead to a delay in the control or eradication of the health problem. In addition to potentially expediting control efforts, cluster identification techniques enable the researcher or health-care official to identify and adjust for confounding factors and to generate new hypotheses regarding disease transmission. This paper examines a variety of temporal and spatial clustering techniques and focuses on those which have been reported recently in the veterinary literature. PMID- 11259823 TI - Behavioural characteristics of two dairy breeds of cows selected (Herens) or not (Brune des Alpes) for fighting and dominance ability. AB - Two breeds of dairy cattle, one selected for intra-specific fighting and dominance ability (Herens, H), the other not selected for this behavioural trait (Brune des Alpes, BA), submitted to the same management techniques, were compared with respect to their social behaviour (dominance, agonistic behaviour, social tolerance, social motivation, social distance), fear reactions, ease of handling and physiological correlates.As expected, cows from the H breed were dominant over the BA cows, they were also less fearful either in response to novel objects or in surprise effect tests and had higher social distances at pasture.On the contrary, H cows were less aggressive in undisturbed groups and more tolerant in a food-competition test than BA cows. There were no differences between the two breeds either in aggressive acts in encounters with unfamiliar animals, or in persistence in conflict situations.Furthermore, H cows were less easy to handle in a standardised test, and tended to be less socially motivated than BA cows. Lastly, H cows had higher plasma testosterone levels, and tended to present a lower increase in plasma cortisol level after a surprise effect than BA cows.Thus, the breeders' selection for fighting and dominance ability in H breed appears to have led to several behavioural and hormonal changes. PMID- 11259824 TI - Vocalisations of the adult female domestic pig during a standard human approach test and their relationships with behavioural and heart rate measures. AB - Vocal communication in the domestic pig is generally not well documented. The aim of this experiment was to categorise and ascribe the function of the vocalisations of 67 Large WhitexLandrace gilts during a standard human approach test. At testing, each group of 3-5 gilts was moved to a handling area where each individual in turn was fitted with a heart rate monitor and introduced individually to a 2.4mx2.4m test arena. After 2min familiarisation, an unfamiliar human entered the pen and stood for 3min against one wall. Behaviour and sound were recorded continuously with sound recordings transferred onto computer for analysis. Three categories of calls were initially identified: single grunts, single squeals and rapidly repeated grunts. Sixty-six gilts performed single grunts, whereas only 28 and 16 gilts performed the other two categories, respectively. Single grunts could be sub-divided into two types based on sound amplitude profile. These types differed significantly in duration. Gilts performed more short and long grunts per minute during the 3min test period than during the familiarisation period. Most short grunts observed in a subset of 15 gilts were performed with the snout close to a pen surface or the human. The rate of short grunts during the test period was negatively correlated with the time taken to make contact with the human and positively correlated with the amount of locomotor behaviour carried out, the total number of interactions with the human and the total time spent within 0.5m of the human. Most long grunts observed in a subset of 15 gilts were performed with the snout away from any surface. The rate of long grunts during the test period positively correlated with amount of locomotor behaviour and heart rate, after the effect of activity had been removed. Squeals could similarly be sub-divided into long and short types on the basis of amplitude profile. Gilts that squealed carried out more locomotor behaviour, interacted with the human more, had higher mean heart rates and lower heart rate rise when touched by the human, suggesting a higher degree of arousal. Rapidly-repeated grunts were associated with close human interaction. The results indicate that the domestic pig performs a number of distinct vocalisations during isolation. Short single grunts appear to be associated with investigatory behaviour. Long single grunts may be a form of contact call, the rate of which is related to physiological and behavioural activity. Squeals may have similar function but result from a higher level of arousal. Short, rapidly-repeated grunts appear to have either a greeting or threat function. With further research, certain pig vocalisations may be identified as providing useful additional information about an individual's welfare. PMID- 11259825 TI - Artificial selection of rams for sexual performance and its effect on the sexual behavior and fecundity of male and female progeny. AB - The objective of this study was to determine the effect of a single generation of artificial selection for sexual performance in rams on the sexual behavior and fecundity of their male and female progeny. Ninety-two ram lambs born to sires selected for either high or low sexual performance were evaluated for their sexual behaviors at approximately 8 months of age when individually exposed to four estrous ewes for 30min in four weekly serving capacity tests. Number of mounts and successful matings (ejaculations) were recorded. Fourteen of the 17 high-performing ram lambs identified were sired by high-performing sires, whereas 22 of 37 low-performing ram lambs were sired by low-performing sires (P<0.01). Sons of high-performing sires exhibited more ejaculations (P<0.04) and more mounts without ejaculation (P<0.02) than sons of low-performing sires. The two groups of ram lambs did not differ in mating efficiency (ratio of ejaculations to total mounts). Daughters of high-performing rams (N=79) exhibited their first behavioral estrus approximately 8 days earlier than daughters (N=61) of low performing sires (P<0.005). Ovulation rates for the two groups of ewe lambs did not differ (P=0.55). It was concluded that there was sufficient genetic variation in the population of sheep studied to obtain a significant response to selection for ram sexual performance in both male and female offspring in a single generation. PMID- 11259826 TI - Domestication effects on foraging strategies in fowl. AB - Birds of two different breeds differing in degree of domestication were studied to reveal any differences in foraging strategies between them. The breeds were wild-type birds (crossing between red jungle fowl (Gallus gallus) and Swedish bantam (Gallus gallus domesticus) and domestic birds (Swedish bantam), breeds representing an increasing level of domestication. Bantam birds have not been selected for any specific characteristics. The birds were allowed to forage in an experimental pen containing two separate food patches, which depleted as a function of being exploited, to see how well the different breeds were able to assess costs and benefits as the distance between patches were changed (short distance between patches compared to long distance between patches). Both breeds behaved in accordance with some general predictions of optimal foraging theory, i.e. moved between patches, left patches before these were empty and stayed for a shorter time in more depleted patches. Wild-type birds responded more than domestic birds to an increase of distance between patches, by spending longer average time in patch when there was a long distance between them compared to when there was a short distance. The wild-type birds adopted what seemed to be a more costly foraging strategy, moving more between patches than the domestic birds without ingesting more feed. During domestication, in the protected environment provided by man, individuals using less costly behavioural strategies may have gained increased fitness over those spending more energy on foraging. Although domestic birds still possessed the ability to respond adaptively to environmental conditions, the differences between the wild-type and the domestic breed might be a result of the reduction of the natural selection pressure which accompanies domestication. PMID- 11259827 TI - Do broiler chicks have a cognitive representation of food quality?. Appetitive, behavioural and ingestive responses to a change in diet quality. AB - It has been observed that when a new diet formulation, or a new batch of the same diet formulation, is presented to poultry, there is a transitory suppression of feeding. It appears that the birds do not recognise the food as being edible, or classify it as being 'unknown'. In order to understand more about food recognition and rejection, the aim of this experiment was to determine whether cognitive processes are involved. Sixteen groups of four broiler chicks were used, and were fed a low quality diet in their home pens. The groups of chicks were trained to run a winding maze to gain access to a high quality diet in a test trough for 15min per day. When training was completed, the feed in the test trough for eight of the groups was changed to that which they received in the home pen, while the other eight groups received no change as a control. Time to traverse the runway did not show an immediate decrease on the day after the change (P>0.05) as would be expected if the birds used a cognitive comparison to determine speed of approach to the test trough. However, the experimental groups were significantly slower compared to the control groups after 4 days (P<0.05). The behaviour observed on the day of the change was indicative of frustration with more scratching and hurried movements shown (P<0.05), and less pecking at feed (P<0.01). Food consumption was lower for experimental groups compared to control groups on all days from the day of change onward (P<0.001). It was concluded that although there was no definitive evidence for the presence of a cognitive representation of food, this may have been due in part to the testing of groups of birds rather than individuals, and the way in which food quality is perceived. The occurrence of behaviours indicative of frustration suggest that a cognitive expectation may have been present. PMID- 11259828 TI - Inappropriate behavior of potential guide dogs for the blind and coping behavior of human raisers. AB - Inappropriate behaviors of potential guide dogs (puppies) for the blind and coping behaviors of their adult female raisers (puppy walkers: PWs) were videotaped in their play situation at home from when the puppies were 2-11 or 12 months of age. The frequency of inappropriate behavior decreased with an increase in the puppies' age, suggesting that human-dog relationships became friendlier. The PWs tended to use moderate coping behaviors to stop the inappropriate behaviors of the puppies. Rejecting interaction with the puppies was effective for stopping the puppies from biting the PWs. Forcible stopping was effective for stopping the puppies from damaging objects. Not responding to the puppies was effective for stopping the puppies from biting the PWs, barking/growling and damaging objects. PMID- 11259830 TI - Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. AB - Experimental and computational approaches to estimate solubility and permeability in discovery and development settings are described. In the discovery setting 'the rule of 5' predicts that poor absorption or permeation is more likely when there are more than 5 H-bond donors, 10 H-bond acceptors, the molecular weight (MWT) is greater than 500 and the calculated Log P (CLogP) is greater than 5 (or MlogP > 4.15). Computational methodology for the rule-based Moriguchi Log P (MLogP) calculation is described. Turbidimetric solubility measurement is described and applied to known drugs. High throughput screening (HTS) leads tend to have higher MWT and Log P and lower turbidimetric solubility than leads in the pre-HTS era. In the development setting, solubility calculations focus on exact value prediction and are difficult because of polymorphism. Recent work on linear free energy relationships and Log P approaches are critically reviewed. Useful predictions are possible in closely related analog series when coupled with experimental thermodynamic solubility measurements. PMID- 11259831 TI - Caco-2 monolayers in experimental and theoretical predictions of drug transport. AB - This review examines the use of Caco-2 monolayers in the prediction of intestinal drug absorption. First, the different routes of drug transport in Caco-2 monolayers are compared with those seen in vivo. Second, the prediction of drug absorption in vivo from transport experiments in cell monolayers is discussed for different classes of drugs. Finally, the use of Caco-2 monolayers as a reference model in physico-chemical and theoretical predictions of drug absorption is discussed. We conclude that Caco-2 monolayers can be used to identify drugs with potential absorption problems, and possibly also to select drugs with optimal passive absorption characteristics from series of pharmacologically active molecules generated in drug discovery programs. PMID- 11259832 TI - Role of dosage regimen in controlling indirect pharmacodynamic responses. AB - The role of drug delivery in controlling indirect pharmacodynamic responses was assessed via computer simulations and literature review. Simulations of responses related to basic indirect response mechanisms were performed for various drug input rates which allowed the importance of drug delivery rate on the overall pharmacodynamic response to be evaluated. Response versus time profiles of integrated or net responses and efficiency were examined. Rate of drug input has the greatest influence on the area under the effect curve when doses are larger and target drug concentrations are above the IC(50)/SC(50). The pharmacodynamics of drugs which elicit indirect pharmacologic responses such as corticosteroids, diuretics, growth hormone, erythropoietin and insulin indicate that sustained drug delivery enhances the therapeutic efficiency and pharmacodynamic availability. PMID- 11259833 TI - The potential use of receptor-mediated endocytosis for oral drug delivery. PMID- 11259834 TI - Influence of physicochemical properties on dissolution of drugs in the gastrointestinal tract. AB - The rate-limiting step to absorption of drugs from the gastrointestinal (GI) tract is often dissolution from the dosage form. Consideration of the Noyes Whitney dissolution model shows that drug diffusivity, solubility in the gastrointestinal contents, the surface area of the solid wetted by the lumenal fluids and the GI hydrodynamics all play a role in determining the in vivo dissolution rate. Solubility in the GI contents is determined by aqueous solubility, crystalline form, drug lipophilicity, solubilization by native surfactants and co-ingested foodstuffs, and pK(a) in relation to the GI pH profile. Compounds with aqueous solubilities lower than 100 microg/ml often present dissolution limitations to absorption. The dose:solubility ratio of the drug provides an estimate of the volume of fluids required to dissolve an individual dose, and when this volume exceeds 1 l, dissolution is often problematic. The surface area of a drug available for dissolution depends on the particle size of the solid and its ability to be wetted by lumenal fluids. Other physiological factors that can play a role in dissolution include the viscosity of the lumenal contents, through its effect on the diffusivity, and mixing and flow patterns within the gut. In order to better predict in vivo dissolution of drugs, dissolution tests which more adequately simulate the physiological conditions are needed. PMID- 11259835 TI - Active secretion and enterocytic drug metabolism barriers to drug absorption. AB - Intestinal phase I metabolism and active extrusion of absorbed drug have only recently been recognized as major determinants of oral drug bioavailability. Both CYP3A4, the major phase I drug metabolizing enzyme in humans, and the multidrug efflux pump, P-glycoprotein (P-gp), are present at high levels in the villus enterocytes of the small intestine, the primary site of absorption for orally administered drugs. Moreover, these proteins are induced by many of the same compounds and demonstrate a broad overlap in substrate and inhibitor specificities, suggesting that they act as a concerted barrier to drug absorption. Clinical studies have demonstrated that inhibition of CYP3A4-mediated intestinal metabolism can significantly improve the oral bioavailability of a wide range of drugs. Intestinal P-gp is a major route of elimination for both orally and intravenously administered anticancer drugs in animal models, and experiments with the Caco-2 cell line have provided strong evidence that inhibition of intestinal P-gp is another means by which oral drug bioavailability could be enhanced. PMID- 11259836 TI - The role of gamma-scintigraphy in oral drug delivery. AB - The gastrointestinal tract is usually the preferred site of absorption for most therapeutic agents, as seen from the standpoints of convenience of administration, patient compliance and cost. In recent years there has been a tendency to employ sophisticated systems that enable controlled or timed release of a drug, thereby providing a better dosing pattern and greater convenience to the patient. Although much about the performance of a system can be learned from in vitro release studies using conventional and modified dissolution methods, evaluation in vivo is essential in product development. The non-invasive technique of gamma-scintigraphy has been used to follow the gastrointestinal transit and release characteristics of a variety of pharmaceutical dosage forms. Such studies provide an insight into the fate of the delivery system and its integrity and enable the relationship between in vivo performance and resultant pharmacokinetics to be examined (pharmacoscintigraphy). PMID- 11259837 TI - Responsive polymeric delivery systems. AB - This paper discusses the state of the art in a relatively new approach in the field of controlled drug delivery-responsive polymeric drug delivery systems. Such systems are capable of adjusting drug release rates in response to a physiological need. The fundamental principles of externally and self-regulated delivery systems are examined. Special attention is paid to specific clinical settings such as diabetes, presenting the advantages and disadvantages of different approaches. PMID- 11259838 TI - Delivery of molecular and cellular medicine to solid tumors. AB - To reach cancer cells in a tumor, a blood-borne therapeutic molecule or cell must make its way into the blood vessels of the tumor and across the vessel wall into the interstitium, and finally migrate through the interstitium. Unfortunately, tumors often develop in ways that hinder each of these steps. Our research goals are to analyze each of these steps experimentally and theoretically, and then integrate the resulting information in a unified theoretical framework. This paradigm of analysis and synthesis has allowed us to obtain a better understanding of physiological barriers in solid tumors, and to develop novel strategies to exploit and/or to overcome these barriers for improved cancer detection and treatment. PMID- 11259839 TI - SMANCS and polymer-conjugated macromolecular drugs: advantages in cancer chemotherapy. AB - This review discusses the development and therapeutic potential of prototype macromolecular drugs for use in cancer chemotherapy, in particular the development and use of SMANCS, a conjugate of neocarzinostatin and poly(styrene comaleic acid). The various topics covered include a brief description of the chemistry and polymer conjugation, the binding of the conjugate to albumin and the biological behaviour in vitro and in vivo after arterial injection in animals, including plasma half-life, and the lipid solubility of SMANCS in medium chain triglycerides and Lipiodol, a lipid contrast medium suitable for use in X ray-computed tomography. The biological response-modifying effects and the tumor targeting mechanism of SMANCS and other macromolecular drugs are also discussed. The latter mechanism is accounted for in terms of a tumor 'enhanced permeability and retention' (or EPR) effect. A principal advantage in the use of SMANCS or other macromolecular drugs is the potential for a reduction or elimination of toxicity. Macromolecular drugs such as a pyran copolymer-NCS conjugate show a marked reduction in bone marrow toxicity normally associated with the use of NCS. This is believed to be due to a hypothetical blood-bone marrow 'barrier' which, relative to NCS, restricts or limits access of the macromolecular drug to the bone marrow. In addition, the clinical possibilities for SMANCS are discussed, including the suggestion that angiotensin II-induced hypertension has clinical potential in improving the selective delivery of macromolecular drugs (i.e. SMANCS) to tumors. Aqueous SMANCS formulations have been tested in pilot studies in patients with solid tumors of the ovary, esophagus, lung, stomach, adrenal gland and in the brain. Formulations based on SMANCS/Lipiodol have been shown to be effective both as a diagnostic tool and for therapeutic use in solid tumors where the formulations are given arterially via a catheter. In a pilot study in primary unresectable hepatoma, an objective reduction in tumor size was observed for about 90% of cases when an adequate amount of the macromolecular drug was administered. A patient receiving such treatment with no active liver cirrhosis and tumor nodules/lesion confined within one liver segment might expect to have a 90% chance of survival after treatment for at least 5 years. PMID- 11259840 TI - Key issues in non-viral gene delivery. AB - The future of non-viral gene therapy depends on a detailed understanding of the barriers to delivery of polynucleotides. These include physicomechanical barriers, which limit the design of delivery devices, physicochemical barriers that influence self-assembly of colloidal particulate formulations, and biological barriers that compromise delivery of the DNA to its target site. It is important that realistic delivery strategies are adopted for early clinical trials in non-viral gene therapy. In the longer term, it should be possible to improve the efficiency of gene delivery by learning from the attributes which viruses have evolved; attributes that enable translocation of viral components across biological membranes. Assembly of stable, organized virus-like particles will require a higher level of control than current practice. Here, we summarize present knowledge of the biodistribution and cellular interactions of gene delivery systems and consider how improvements in gene delivery will be accomplished in the future. PMID- 11259841 TI - Adenovirus-mediated gene transfer to liver. PMID- 11259842 TI - Delivery systems for penetration enhancement of peptide and protein drugs: design considerations. AB - This paper discusses the challenges to be met in designing delivery systems that maximize the absorption of peptide and protein drugs from the gastrointestinal and respiratory tracts. The ideal delivery system for either route of administration is one that will release its contents only at a favorable region of absorption, where the delivery system attaches by virtue of specific interaction with surface determinants unique to that region and where the delivery system travels at a rate independent of the transitory constraints inherent of the route of administration. Such a delivery system, which is as yet unavailable, will benefit not only peptide and protein drugs, but other poorly absorbed drugs. PMID- 11259843 TI - Delivery of peptides and proteins through the blood-brain barrier. AB - Peptide and protein therapeutics are generally excluded from transport from blood to brain, owing to the negligible permeability of these drugs to the brain capillary endothelial wall, which makes up the blood-brain barrier (BBB) in vivo. However, peptides or protein therapeutics may be delivered to the brain with the use of the chimeric peptide strategy for peptide drug delivery. Chimeric peptides are formed when a non-transportable peptide therapeutic is coupled to a BBB drug transport vector. Transport vectors are proteins such as cationized albumin, or the OX26 monoclonal antibody to the transferrin receptor; these proteins undergo absorptive-mediated and receptor-mediated transcytosis through the BBB, respectively. In addition to vector development, another important element of the chimeric peptide strategy is the design of strategies for coupling drugs to the vector that give high efficiency coupling and result in the liberation of biologically active peptides following cleavage of the bond linking the therapeutic and the transport vector. The avidin/biotin system has been recently shown to be advantageous in fulfilling these criteria for successful linker strategies. The use of the OX26 monoclonal antibody, the use of the avidin/biotin system as a linker strategy, and the design of a vasoactive intestinal peptide (VIP) analogue that is suitable for monobiotinylation and retention of biologic activity following cleavage, allowed for the recent demonstration of in vivo pharmacologic effects in brain following the systemic administration of relatively low doses (12 microg/kg) of neuropeptide. PMID- 11259844 TI - The role of electroosmotic flow in transdermal iontophoresis. AB - Iontophoresis enhances transdermal drug delivery by three mechanisms: (a) the ion electric field interaction provides an additional force which drives ions through the skin; (b) flow of electric current increases permeability of skin; and (c) electroosmosis produces bulk motion of the solvent itself that carries ions or neutral species, with the solvent 'stream'. The relative importance of electroosmotic flow is the subject of this review. Experimental observations and theoretical concepts are reviewed to clarify the nature of electroosmotic flow and to define the conditions under which electroosmotic flow is an important effect in transdermal iontophoresis. Electroosmotic flow is bulk fluid flow which occurs when a voltage difference is imposed across a charged membrane. Electroosmotic flow occurs in a wide variety of membranes, is always in the same direction as flow of counterions and may either assist or hinder drug transport. Since both human skin and hairless mouse skin are negatively charged above about pH 4, counterions are positive ions and electroosmotic flow occurs from anode to cathode. Thus, anodic delivery is assisted by electroosmosis, but cathodic delivery is retarded. Water carried by ions as 'hydration water' does not contribute significantly to electroosmotic flow. Rather electroosmotic flow is caused by an electrical volume force acting on the mobile counterions. The simple 'limiting law' theory commonly given in textbooks and some research articles is a very poor approximation for transdermal systems. However, several extensions of the limiting law are compatible with each other and with the available experimental data. One of these theories, the Manning theory, has been incorporated into a theory for the effect of electroosmotic flow on iontophoresis, the latter theory being in good agreement with experiment. Both theory and experimental data indicate that electroosmotic flow increases in importance as the size of the drug ion increases. The 'ionic' or Nernst-Planck effect is the largest contributor to flux enhancement for small ions. Increased skin permeability or the skin 'damage effect', is a significant factor for both large and small ions, particularly for experiments at high current density. For monovalent ions with Stokes radii larger than about 1 nm, electroosmotic flow is the dominant flow mechanism. Because of electroosmotic flow, transdermal delivery of a large anion (or negatively charged protein) from the anode compartment can be more effective than delivery from the cathode compartment. PMID- 11259845 TI - Factors affecting short-term and long-term stabilities of proteins. AB - Proteins are marginally stable and, hence, are readily denatured by various stresses encountered in solution, or in the frozen or dried states. Various additives are known to minimize damage and enhance the stability of proteins. This review discusses the current knowledge of the mechanisms by which these additives stabilize proteins against acute stresses, and also the various factors to be considered for long-term storage of proteins in solution. PMID- 11259846 TI - The effect of retinol on hepatic and renal drug-metabolising enzymes. AB - Retinol pretreatment (75 mg/kg/day, 4 days) potentiated paracetamol-induced hepatotoxicity in BALB/c mice (alanine aminotransferase (ALT) activity; 2510+/ 602 vs 1155+/-282 IU/l; retinol+paracetamol vs corn oil+paracetamol, respectively, P<0.05); however, this potentiation did not occur in the kidney, indicating an organ-specific response. Retinol treatment alone was not toxic in either organ, as indicated by ALT activity, blood urea nitrogen and creatinine. The potentiation effect could be mediated by retinol's induction of CYP450 isoforms relevant to paracetamol metabolism or through depletion of glutathione. Therefore, these parameters were investigated in both organs. Following retinol treatment, renal CYP2E1 and hepatic CYP1A2 and CYP2E1 catalytic activities and polypeptide levels were unchanged. However, retinol significantly decreased both the catalytic activity (0.23+/-0.03 vs 0.53+/-0.06 nmol/mg/min; retinol vs untreated, respectively, P<0.05) and polypeptide levels (58+/-0.6% of control) of hepatic CYP3A. Inhibition of renal CYP3A did not occur as catalytic activity and polypeptide levels were unchanged from control. Following retinol treatment, total reduced glutathione levels, in both organs, were not significantly different from control. Therefore, the potentiation of paracetamol-induced hepatotoxicity is independent of CYP450 and glutathione. PMID- 11259847 TI - An unusual photohaemolytic property of riboflavin. AB - Riboflavin (RF) is a known photoreactive and phototoxic molecule. However, unlike other photosensitizers, it does not induce photohaemolysis of erythrocytes by itself. On the other hand, illuminated RF caused haemolysis but in the presence of serum or plasma. The kinetics of photohaemolysis in the presence of serum/plasma has been studied by monitoring the rate of haemolysis spectrophotometrically and morphological changes at erythrocytes membrane by scanning electron microscopy. We found that the extent of mammalian RBC membrane damage was dependent on the concentration of RF or hematoporphyrin (HP) (0-20 microgram/ml) and dose of sunlight (0-20 min). The RBC membrane-damaging potential of illuminated HP was not affected by the presence of plasma in the reaction system. Furthermore, RF showed a protective role against photohaemolysis caused by photoexcited HP if erythrocytes were preincubated with RF in the absence of serum/plasma. For mechanistic studies, biochemical parameters such as acetylcholinesterase activity (AChE) and formation of TBA-reactive substance (TBA RS) were analysed in RBC and RBC+plasma under a similar set of experimental conditions. We observed about a 25% decrease in AchE activity as a synergistic action of RF or HP (20 microgram/ml) and sunlight (30 min) in both cases (RBC or RBC+plasma). Interestingly, illuminated RF caused about a 125% increase of TBA-RS in a reaction system consisting of RBC+plasma. On the other hand, an increase in the production of TBA-RS by illuminated RF was not observed in the absence of plasma/serum, in the reaction system. These results suggested that photooxidation of RBC membrane lipids by illuminated RF, under the influence of plasma/serum, may be one of the causes of membrane modification leading to RBC lysis. PMID- 11259848 TI - Anti-tumor promoting activity of Asteracantha longifolia against experimental hepatocarcinogenesis in rats. AB - Vegetables, natural products of plant origin and numerous non-nutritive dietary constituents have been shown to play a salutary role in cancer chemoprevention. The present study aims to evaluate the chemopreventive efficacy of the methanol fraction of Asteracantha longifolia seed extract against development of 2 acetylaminofluorene (2-AAF)-selected gamma-glutamyl transpeptidase (gamma-GT) positive foci following diethylnitrosamine (DEN) initiation. Treatment of rats with doses 200 and 400 mg/kg body weight of methanol extract of A. longifolia seeds on alternate days, subsequent to carcinogen treatment, for 6 weeks significantly reduced the incidence and size distribution of gamma-GT-positive foci and tumor formation. Administration of A. longifolia seeds significantly (P<0.001) ameliorated the activities of antioxidant enzymes, glutathione peroxidase (GPx) and catalase (CAT), in a dose-dependent manner. Prophylactic administration of seed extract simultaneous to 2-AAF in the diet, at same doses, significantly suppressed 2-AAF and partial hepatectomy (PH)-induced ornithine decarboxylase (ODC) activity and [(3)H]thymidine incorporation into hepatic DNA, in a dose-dependent manner. Assimilation of the quantitative foci data together with the findings of the modulation of tumor promoting markers give ample evidence to the anti-tumor promoting potential of A. longifolia seeds against chemically-induced hepatocarcinogenesis in Wistar rats. PMID- 11259849 TI - Methods for estimating heterocyclic amine concentrations in cooked meats in the US diet. AB - Heterocyclic amines (HAs) are formed in numerous cooked foods commonly consumed in the diet. A method was developed to estimate dietary HA levels using HA concentrations in experimentally cooked meats reported in the literature and meat consumption data obtained from a national dietary survey. Cooking variables (meat internal temperature and weight loss, surface temperature and time) were used to develop relationships for estimating total HA concentrations in six meat types. Concentrations of five individual HAs were estimated for specific meat type/cooking method combinations based on linear regression of total and individual HA values obtained from the literature. Using these relationships, total and individual HA concentrations were estimated for 21 meat type/cooking method combinations at four meat doneness levels. Reported consumption of the 21 meat type/cooking method combinations was obtained from a national dietary survey and the age-specific daily HA intake calculated using the estimated HA concentrations (ng/g) and reported meat intakes. Estimated mean daily total HA intakes for children (to age 15 years) and adults (30+ years) were 11 and 7.0 ng/kg/day, respectively, with 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) estimated to comprise approximately 65% of each intake. Pan-fried meats were the largest source of HA in the diet and chicken the largest source of HAs among the different meat types. PMID- 11259850 TI - Identification of differentially expressed genes following treatment of monkey kidney cells with the mycotoxin fumonisin B(1). AB - Fumonisin B(1) (FB(1)) is a mycotoxin produced by the phytopathogenic fungus Fusarium moniliforme, which structurally resembles sphingoid bases. FB(1) perturbs sphingolipid synthesis by inhibiting the activity of ceramide synthase. Depending on the host, ingestion of FB(1) causes equine leukoencephalomalacia or porcine pulmonary edema. It is also carcinogenic to rats and may play a role in certain human cancers. Previous studies showed that FB(1) repressed specific isoforms of protein kinase C and cyclin-dependent kinase 2 (CDK2) activity. Conversely, FB(1) induced expression of CDK inhibitors, p21(Waf1/Cip1), p27(Kip1), and p57(Kip2) in monkey kidney cells (CV-1). Consequently, FB(1) treatment of CV-1 cells leads to cell-cycle arrest and apoptosis. The baculovirus IAP gene (inhibitor of apoptosis), which blocks tumor necrosis factor (TNF) induced apoptosis, protects several fibroblast cell types from apoptosis, suggesting the TNF pathway is important for FB(1)-induced apoptosis. To identify genes that are induced by FB(1), we used a PCR-based subtraction approach. Eight genes that showed high similarity (> 90%) to known mammalian genes were identified. These genes included: tumor necrosis factor type 1 receptor associated protein 2 (TRAP2), human leukemia virus receptor (GLVR1), human Scaffold attachment factor A (SAF-A) also called heterogeneous nuclear ribonucleoprotein U (hnRNP-U), human protein kinase C-binding protein (RACK7), human oligosaccharyl transferase STT3 subunit, mouse WW-domain binding protein 2 (WBP2), human fibronectin, and an unknown human clone. The ability of FB(1) to alter gene expression and signal transduction pathways may be necessary for its carcinogenic and toxic effects. PMID- 11259851 TI - Age-related differences in the toxicity of ochratoxin A in female rats. AB - Ochratoxin A (OTA) is a mycotoxin found in food and feedstuffs of plant and animal origin. OTA exposure is related to nephropathy in humans. Age-related differences, especially in nephro- and immunotoxicity of OTA, were investigated in young adult (aged 12 weeks) and old (aged 27-30 months) female SPF Wag rats, treated by gavage with 0, 0.07, 0.34 or 1.68 mg OTA/kg body weight for 4 weeks. In both age groups, survival was significantly decreased in the highest dose group. Clinical condition, body weight, clinical chemistry parameters (ALAT, ASAT, creatinin and urea) and target organs (as identified by weight and pathology - kidney, liver, adrenals, forestomach and brain) were affected by age and dose, but often more severely in old than in young rats. OTA induced primarily nephropathy. Old rats were more sensitive to induction of tubular karyomegaly and vacuolation/necrosis. In young rats, OTA induced a dose-related thickening of the basement membrane and reduction in splenic T-cell fraction. Decreased IgG levels were seen at 0.34 mg/kg OTA (young and old rats) and 1.68 mg/kg OTA (young rats). Vacuolation of the white brain matter (cerebellar medulla and ventral parts of the brain stem) was significantly increased in young rats at 0.34 and 1.68 mg/kg OTA and in old rats at 0.07 and 0.34 mg/kg OTA. It was concluded that: (1) the profiles of OTA toxicity for both age groups are similar, with the kidney and possibly the brain being primary target organs; (2) based on clinical and pathological data old rats are more sensitive to OTA than young rats; and (3) the immune system is probably not the primary target of OTA toxicity. PMID- 11259852 TI - Genotoxic effects after in vivo exposure of vegetable extracts containing heavy metals from the Dhapa area of Calcutta, India. I. Effects of cauliflower, spinach and radish. AB - Several reports have indicated that the sewage-fed vegetables of the Dhapa area, near the city of Calcutta, contain a very high amount of heavy metals. Currently 800 ha of land is being utilised throughout the year to cultivate more than eight types of vegetables, with a production of about 147 tonnes per day. A major population of Calcutta consumes these vegetables grown in the Dhapa area. Recently there has been huge pressure on the State Government to ban vegetables grown in the Dhapa area for human consumption. For this reason, we have studied the genotoxic effects of some of the most commonly used vegetable extracts from the Dhapa area after in vivo acute exposure in mice as measured by chromosomal aberrations (CA) and sister chromatid exchanges (SCE) to find out the minimum threshold dose to induce CA and SCE. Three different concentrations of the three most commonly used vegetable extracts (cauliflower, spinach, radish) were fed by gavage to mice for the study of CA and SCE. A significant increase in CA was observed only at the highest concentration of all the vegetable extract-treated groups when compared with the solvent control. A significant increase in SCE were observed in the middle and high doses of spinach and only the high dose of cauliflower and radish extract-treated series when compared with distilled water control. The lowest dose was equivalent to approximately 1 kg of vegetables consumed by a human (60 kg body weight) in a day. The middle and high doses of each vegetable extract were much higher than the normal amount of vegetables that a human can consume per day. So the minimum dose for inducing SCE and CA was much higher than the amount a human can consume in a day. Therefore this study indicates that these vegetables are safe for human consumption up to a certain limit, and attention should be given to reducing the heavy metal contents in the soil and sewage of the Dhapa area to thus reduce the heavy metal concentrations in the vegetables. PMID- 11259853 TI - Assessment of a two-generation reproductive and fertility study of mercuric chloride in rats. AB - Effects of mercuric chloride (MC) on the reproductive performance of two successive generations of rats was evaluated. F(0) rats were exposed to 0.0:0.0 (males:females), 0.50:0.75 (males:females), 1.00:1.50 (males:females) and 1.50:2.50 (males:females) mg/kg/day MC. Selected parental F(1) males and females were exposed to the same doses received by their parents (F(0)). Significant differences resulting from exposure of the F(0) generation to MC were found in implantation efficiency, fertility, live births and day 4 survival indices, litter size, and the body weight of F(1) pups. However, the continued exposure of the F(1) generation to MC did not affect fertility index or litter size, but did significantly affect implantation efficiency, live births and day 4 survival indices. In F(0) males, body weight and weights of the kidneys, testes, epididymides, prostate and seminal vesicles were significantly different, while in F(1) males, body weight, kidney weight, brain weight, liver weight and the weights of the testes, prostate and seminal vesicles were significantly different. In F(0) females, body weight and the weights of the kidneys, brain and liver were significantly different, while in F(1,) females, body weight, as well as the weights of the kidneys, liver, adrenals, uterus and ovaries were significantly different. These data showed that exposure to MC resulted in more adverse reproductive effects in the first generation and that these effects moderated in the second generation. PMID- 11259854 TI - The stability of cytokeratin 18 in human liver cells during colchicine-induced microtubule disruption. AB - The cytoskeleton plays important roles in cell function and is therefore implicated in the pathogenesis of many human liver diseases, including malignant tumors. The stability of cytokeratin proteins during tumor transformation in human hepatocellular carcinoma has been studied with a molecular approach previously. The results demonstrate that the cytokeratin is modulated in human hepatocellular carcinoma. Besides this, three low molecular weight cytokeratin molecules (named HCC CK) are found. This indicates that these HCC CKs have undergone modulation from the human hepatocyte cytokeratin 18. We also checked the cytokeratin profile of the human hepatoma cell line PLC/PRF/5 with the same methods to ensure the HCC CK molecules are produced by modulation but not protein degradation. The stability of cytokeratin molecules was studied by a different approach. The cytokeratin compositions of human liver cells (Chang cell line) were analysed under the effects of microtubule-disrupting drug (colchicine) by SDS-PAGE, Western blot, immunoprecipitation using a commercially available monoclonal anti-cytokeratin 18 antibody and immunofluorescent staining. Within 1 h of treatment, the microtubule began to collapse and the filamentous structure was shortening. The microtubule had almost collapsed and became fragmented to form a lattice-like network after 24 h of treatment. The cytokeratin was modulated after long-term (24 h) treatment of colchicine, and the molecular weight became 14 kD and the antigenicity was lost. The stability of cytokeratin molecules was related to the intact microtubule network, after disruption of the microtubule the cytokeratin would be modulated. The intact microtubule network was a stabilizing factor of cytokeratin 18 in human liver cells. PMID- 11259855 TI - Storage time and deodorization temperature influence the formation of aniline derived compounds in denatured rapeseed oils. AB - In 1981 an epidemic, named Toxic Oil Syndrome, occurred in Spain as a result of ingestion of rapeseed oil denatured with 2% aniline, which had been imported for industrial use but was fraudulently diverted and processed for human consumption. Two groups of chemical compounds have been identified in the ingested toxic oil: fatty acid anilides and amino-propanediol derivatives. The objective of this work was to assess the effect of several refining process variables on the formation of 3-(N-phenylamino)-1,2-propanediol (PAP) esters. The amount of PAP esters in aniline-denatured oil increased dramatically when oil was heated from 250 degrees C to 300 degrees C. However, the ones formed when 300 degrees C was reached were lost during processing at that temperature. The level maintained during the operation time at 300 degrees C was higher in denatured samples stored for 3 weeks before refining than in denatured samples stored only for 1 week. Anilides were also analyzed. We found that anilides decreased very little with distillation time. In this paper we discuss the influence of storage time prior to refining and of elevated refining temperature, such as temperatures that might occur in close proximity to a deodorizer coil. PMID- 11259856 TI - Motherhood and memory: a review. AB - This paper reviews the literature on intellectual change during and following pregnancy in humans. Whilst much has been written in an impressionistic way, this area has received little serious scientific attention, and at the beginning of the twenty first century considerable gaps in our knowledge remain. Targets for future research are suggested. The second part of the paper reviews the complex hormonal changes that place during pregnancy and birth and examines the impact that these changes may have on cognition, drawing together the findings from a number of scientific fields. Whilst this review may generate more questions than it answers, we hope that it will create interest and stimulate research in this long neglected field. PMID- 11259857 TI - Individual differences in the recovery of the hypothalamic-pituitary-adrenal axis after termination of exposure to a severe stressor in outbred male Sprague-Dawley rats. AB - Individual differences in the speed of recovery of the hypothalamic-pituitary adrenal (HPA) axis response to immobilization in wooden boards (IMO) were studied in two experiments using a normal population of two months old male outbred Sprague-Dawley rats. In a first experiment, rats were subjected to 2 h IMO and were sampled, together with a group of control rats, at various times by the tail nick procedure. Rats were divided into three groups depending on plasma corticosterone levels observed 2 h after termination of exposure to IMO: fast recovery (FR), intermediate recovery (IR) and slow recovery (SR). When the samples obtained at different times were classified in function of the three groups obtained at 2 h post-stress, no differences among groups were observed just after IMO. However, in the morning on the day after stress, all IMO rats showed higher plasma corticosterone levels than controls, but SR rats showed higher levels than FR rats, whereas IR rats were in between. Neither ambulatory activity in the open-field nor behaviour in the plus-maze was related to the HPA responsiveness to IMO. In a second experiment, there were no between-groups differences in ACTH and corticosterone levels obtained just after IMO. However, at 2 h, post-stress ACTH levels followed the same pattern as corticosterone. These data indicate that individual differences in the speed of recovery of the HPA axis after exposure to a severe stressor are not related either to individual differences in fear/anxiety or to the HPA response during exposure to the stressor. These individual differences in the capability to terminate the activation of the HPA axis after stress might be related to individual differences in the predisposition to develop stress-induced pathology. PMID- 11259859 TI - Central dopaminergic function in anorexia and bulimia nervosa: a psychoneuroendocrine approach. AB - Data on central dopamine (DA) function in patients with Anorexia Nervosa (AN) and Bulimia Nervosa (BN) are contradictory. To tentatively clarify the brain secretory state of the amine and its relationship with the nutritional impairments and/or the psychopathological aspects of the two disorders, we measured the responses of growth hormone (GH) to acute stimulation with apomorphine (APO), a selective D-1 and D-2 receptor agonist, in 16 AN patients, 8 restricted (AN-R) and 8 bingeing-purging (AN-BP), in 7 BN patients and in 8 healthy controls (CTR). Interference of impairment of the somatotropic axis in the GH response to APO stimulation was excluded by measuring the GH and insulin like growth factor-1 (IGF-1) basal levels and GH responses to growth hormone releasing hormone (GHRH) stimulation. Psychological aspects of patients and controls were investigated by the rating scales Eating Disorder Inventory (E.D.I.), Bulimic Investigation Test Edinburgh (B.I.T.E)., and Yale-Brown Cornell Eating Disorder Scale (YBC-ED). Basal GH levels were significantly higher and those of IGF-1 lower in AN-R than in AN-BP, BN and CTR subjects. GH responses to GHRH stimulation were significantly higher in AN-R than in AN-BP and BN patients and in CTR. GH responses to APO stimulation were significantly lower in AN-R and AN-BP than in BN and CTR subjects, suggesting that at the hypothalamic level there is a subsensitivity of postsynaptic D-2 receptors and possibly a presynaptic DA hypersecretion. The altered GH responses to APO stimulation did not correlate with the Body Mass Index, while they correlated negatively with E.D.I. scores. PMID- 11259858 TI - Psychological, cardiovascular, and metabolic correlates of individual differences in cortisol stress recovery in young men. AB - The relationship of free salivary cortisol stress recovery and basal cortisol with psychological, cardiovascular and metabolic factors was investigated in 82 healthy young men. Blood pressure, heart rate, cortisol and mood were assessed during a single laboratory session involving mental arithmetic and speech tasks, and lipid profiles were analysed from a fasting blood sample. Participants were divided into high (n=31) and low (n=51) cortisol stress recovery groups on the basis of the magnitude of changes between the peak cortisol responses to tasks and the lowest levels recorded at the end of a 30 min post-stress rest period. The high recovery group showed consistent increases in cortisol following each of the tasks, while the low recovery group showed little change across the session. Cortisol levels in the two groups did not differ at the end of the post-stress recovery period. The groups were indistinguishable in age, body mass index, smoking and alcohol consumption, and did not differ in psychological characteristics including anxiety, depression and perceived social support. However, the high stress recovery group had elevated low density lipoprotein cholesterol and total cholesterol/high density lipoprotein ratios, suggesting raised cardiovascular disease risk. The high stress recovery group also reported greater psychological activation during tasks, and greater recent minor life stress, than did the low recovery group. There was no association between rate of cortisol recovery and cardiovascular responses to tasks. But resting cortisol was related to blood pressure stress reactivity, suggesting that cortisol played a permissive role in augmenting sympathetically-driven cardiovascular responses. The results suggest that the rate of cortisol stress recovery is associated with variations in metabolic risk, and with differences in psychological state but not trait characteristics. PMID- 11259860 TI - Psychosocial factors, respiratory viruses and exacerbation of asthma. AB - The aim of this research was study the role of psychosocial factors in exacerbations of asthma in adults induced by upper respiratory tract infections (URTIs). It involved a longitudinal study (one year) of 92 adults with asthma. The volunteers were 27 men and 65 women 19-46 years of age with a mean duration of wheeze of 19 years. The main outcome measure was symptomatic colds producing asthma exacerbations (infections confirmed by laboratory assays and exacerbation of asthma confirmed by objective changes in peak expiratory flow rate). The results showed that about 20% of the sample did not report an episode. This sub group had a high proportion of males, low negative affectivity scores and consumed more alcohol. When volunteers with at least one episode were considered it was found that those who reported more negative life events and had low levels of social support had more episodes. Smokers were more likely to have to visit their doctor when they developed a cold-induced exacerbation of asthma. Overall, these results show that health-related behaviours, demographic and psychosocial factors influence susceptibility to and severity of exacerbations of asthma by URTIs. PMID- 11259861 TI - Circannual pattern of hypothalamic-pituitary-thyroid (HPT) function and mood during extended antarctic residence. AB - The seasonal variation in thyroid function and mood was examined in 10 men and two women who spent the 1997 or 1998 austral winter at McMurdo Station, Antarctica. Serum samples of TSH, free T3 and free T4 were collected each month over a 10-month period (October-August), along with responses to the Profile of Mood States (POMS) and the Center for Epidemiologic Studies - Depression (CES-D) Scale. Both TSH and mood (a summary score created from the POMS depression, anger, fatigue and confusion subscales) exhibited a circannual pattern with peaks during the months of November and July and a trough during the months of March and April. High levels of tension-anxiety and confusion were preceded by declines in free T3 and T4. However, increases in tension-anxiety and total mood disturbance also preceded a decline in free T3 levels, suggesting a feedback of mood on T3 levels. Levels of free T4 were independently associated with preceding increases in anger scores. These results support the hypothesis that the symptoms characteristic of the winter-over syndrome is a state of relative CNS hypothyroidism. This model of seasonal variation in thyroid function and mood also has implications for an understanding of potential mechanisms underlying the association between latitude and SAD or S-SAD. PMID- 11259862 TI - Short-term sertraline treatment suppresses sympathetic nervous system activity in healthy human subjects. AB - Increased sympathetic nervous system (SNS) activity has been associated with stress, major depression, aging, and several medical conditions. This study assessed the effect of the selective serotonin reuptake inhibitor (SSRI), sertraline, on sympathetic nervous system (SNS) activity in healthy subjects. Twelve healthy volunteers participated in a double-blind, placebo-controlled, norepinephrine (NE) kinetic study, in which the effects of sertraline on SNS activity were ascertained by determining NE plasma concentrations and NE plasma appearance rates and clearance rates in sertraline or placebo conditions. Subjects received 50 mg of sertraline or placebo for two days and then one week later underwent the same protocol with the other drug. By single compartmental analysis, plasma NE appearance rates were significantly lower in the sertraline compared to the placebo condition (0.26+/-0.10 vs 0.40+/-0.23 microg/m(2)/min; P=0.04). Our study found that the net effect of short-term SSRI treatment is an apparent suppression of SNS activity as indicated by a decreased plasma NE appearance rate in the sertraline condition. If this preliminary finding can be extended to long-term treatment of patients, this could have significant therapeutic relevance for treating depression in elderly patients or those with cardiac disease, in which elevated SNS activity may exacerbate underlying medical conditions. PMID- 11259863 TI - Flavonoids as peroxynitrite scavengers: the role of the hydroxyl groups. AB - It has been reported that flavonoids efficiently protect against peroxynitrite toxicity. Two pharmacophores have been identified in flavonoids, namely the catechol group in ring B and the hydroxyl (OH) group at the 3-position. In this study, this structure-activity relationship was further examined. It was found that catechol (1,2-dihydroxybenzene) is a potent peroxynitrite scavenger, whereas phenol (hydroxybenzene) is not. Of the flavonols tested without a catechol group in ring B, kaempferol (OH groups at positions 3,5,7,4') and galangin (OH groups at positions 3,5,7) are also potent scavengers, whereas apigenin (OH groups at positions 5,7,4') and chrysin (OH groups at positions 5,7) are not. This confirms the importance of the OH group at the 3-position. However, the synthetic flavonol TUM 9761 and 3-hydroxyflavone (OH group only at position 3) are poor scavengers. Based on these results, the structure-activity relationship on the peroxynitrite scavenging activity of flavonols was refined. The catechol in ring B remains important. Also the 3-OH group remains important, but the activity of this pharmacophore is influenced by the substituents at position 5 and at position 7. PMID- 11259864 TI - Effects of mercuric chloride exposure on the glutamate uptake by cultured retinal pigment epithelial cells. AB - The cytotoxicity of mercuric chloride and the effects of mercuric chloride on glutamate and calcium uptake and the factors regulating glutamate uptake were studied in retinal pigment epithelium (RPE) cell cultures. RPE cells isolated from pig eyes and human RPE cell line (D407) cells were cultured to confluency and further subcultured according to the test protocol in question. The cytotoxicity caused by 15 min of exposure to mercuric chloride (0.01--1000 microM) was evaluated by WST-1 assay based on the activity of mitochondrial dehydrogenases. [(3)H]Glutamate uptake was measured after the cells were exposed to 0.1--100 microM mercuric chloride and the selected regulators of protein kinase C (PKC) pathway: PKC activator SC10, PKC inhibitor chelerythrine chloride, phospholipase A(2)/C inhibitor manoalide, tyrosine kinase inhibitor lavendustin A, competitive NMDA receptor antagonist AP7 and IP(3) receptor antagonist heparin. Intracellular calcium was monitored with Fluo-3 probe starting immediately after the exposure to 1--1000 microM mercuric chloride. Mercuric chloride showed concentration-dependent effects on cell viability, on glutamate uptake and on intracellular calcium concentration. The results give some support to the concept that glutamate uptake is affected by PKC. The PKC inhibitor chelerythrine chloride decreased glutamate uptake by 25%, but the PKC activator SC10 could partly prevent the inhibitory effect of mercuric chloride. Lavendustin A, manoalide and heparin had smaller, but statistically significant, effects. All these substances act on mediators which can regulate the activity of PKC. However, PKC is not likely to be the only regulator of glutamate uptake. The rise observed in [Ca(2+)](i) may initiate various cellular events during mercury intoxication. PMID- 11259865 TI - Antioxidant and prooxidant roles for beta-carotene, alpha-tocopherol and ascorbic acid in human lung cells. AB - Experiments were conducted to determine the antioxidant and prooxidant effects of beta-carotene, alpha-tocopherol and ascorbic acid on human lung cells at different oxygen (O(2)) tensions. Free radical initiator, 2,2'-azobis (2 amidinopropane) dihydrochloride (AAPH), was used to induce the cellular damage associated with lipid peroxidation, protein oxidation and DNA breaks. Under hypoxic conditions (0 torr O(2) tension) all compounds produced a concentration dependent antioxidant effect. Mixtures of the three compounds exhibited greater protective affects than any individual compound. At 143 torr O(2) tension, all compounds exhibited concentration-dependent protective effects against AAPH induced cellular lipid, protein and DNA damage. At 722 torr O(2) tension, cells exhibited a consistent increase in lipid peroxidation (isoprostane formation), protein oxidation (carbonyl formation) and DNA damage (p53 protein accumulation). beta-Carotene (1.5 microM) produced a prooxidant effect by promoting 12% isoprostane formation. Protein oxidation and DNA damage at 722 torr O(2) tension was not increased by beta-carotene; however, the antioxidant effect of beta carotene was attenuated. The antioxidant effects of alpha-tocopherol, ascorbic acid, and mixtures of the three antioxidant compounds also were reduced by the high O(2) conditions. These results partially substantiate the hypothesis that the antioxidant and prooxidant effects of beta-carotene are dependent on O(2) tension and concentration of beta-carotene. Such findings may partially explain why selected populations, such as smokers, respond adversely when supplemented with beta-carotene. PMID- 11259866 TI - Effects of mycotoxins on human immune functions in vitro. AB - Immunosuppressive and carcinogenic Fusarium mycotoxins may appear in domestic food products. Therefore, the immunological effects of Fusarium mycotoxins were tested on human peripheral blood mononuclear cells from different blood donors. In the present study we investigated deoxynivalenol (DON), 3 acetyldeoxynivalenol, fusarenon-X, T-2 toxin, zearalenone, alpha-zearalenol, beta zearalenol and nivalenol for their effects on T and B cells in a proliferation assay, antibody-dependent cellular cytotoxicity (ADCC) and natural killer (NK) cell activity on human peripheral blood mononuclear cells. The concentrations applied in our experiments were similar to those which can be found in normal human peripheral blood system (0.2--1800 ng/ml). Among the eight mycotoxins tested, T-2 toxin, fusarenon X, nivalenol and deoxynivalenol exerted the highest immunosuppressing effect on human peripheral blood mononuclear cells in vitro. Mycotoxin-induced immunosupression was manifested as depressed T or B lymphocyte activity. Furthermore, by virtue of inhibition of NK cell activity, the protection against tumor development may also be attenuated. PMID- 11259867 TI - Proliferation and differentiation of murine haemopoietic progenitor cells in stroma-free culture in the presence of metabolites of chlorinated pesticides. AB - We have studied the influence of metabolites of chlorinated pesticides (lindane, pentachlorophenol, hexachlorobenzene) on proliferation and differentiation in two stroma-free murine bone marrow culture models, a multipotent progenitor cell line (FDCP-mix) and primary lineage-depleted bone marrow cells. Tetrachlorohydroquinone (Cl(4)pHQ), tetrachloro-p-benzoquinone (Cl(4p)BQ), but not their positional isomers, tetrachlorocatechol (Cl(4)oHQ) and tetrachloro-o benzoquinone (Cl(4)oBQ), nor 2,4,6-trichlorophenol (2,4,6-Cl(3)P), were much more toxic to FDCP-mix cells cultured under conditions which lead to self-renewal than under conditions which lead to granulocyte-macrophage differentiation. Under the latter conditions, Cl(4)pHQ and Cl(4p)BQ even stimulated growth at intermediate concentration levels. In the primary cell cultures, pronounced differences were observed in the sensitivity between individual developmental pathways and between the different compounds. The percent of cells differentiating into the granulocytic lineage was increased at high concentration levels of each test compound. However, stimulatory effects on the macrophage lineage were observed at intermediate concentration levels of Cl(4)pHQ, Cl(4p)BQ and 2,4,6-Cl(3)P, and differentiation into erythrocytes was stimulated at low concentrations of 2,4,6 Cl(3)P. It is concluded that chlorinated monocyclic pesticides, after biotransformation to quinoid metabolites, may interact directly with haemopoietic progenitor cells with differential effects on self-renewal and differentiation. These mechanisms could lead to myeloplastic disorders. PMID- 11259868 TI - Amelioration of cytotoxic effects of aflatoxin by vitamin A: an in vitro study on erythrocytes. AB - We have evaluated the ameliorative role of vitamin A on aflatoxin-induced cytotoxicity in vitro. Aflatoxin (1.95 microM)-induced hemolysis was found to be significantly reduced on addition of vitamin A (125--1250 IU/ml) in incubation medium. The decrease in hemolysis was almost dose dependent. The kinetics of reduction of AFB(1) to B(2) and AFG(1) to G(2) by vitamin A has been investigated in dilute aqueous solution at 37 degrees C. The rate of the reduction was found to be first order with respect to the concentration of vitamin A and aflatoxin concentration. PMID- 11259869 TI - Approaches for the development of cell-based in vitro methods for contact sensitization. AB - Allergic contact dermatitis (ACD) is a cell-mediated immune response to small molecular weight chemicals that contact and penetrate the skin. There are a variety of characteristics that determine whether a chemical can function as a contact sensitizer (or allergen) including the ability to penetrate into the skin, react with protein and be recognized as antigenic by immune cells. The ultimate challenge for developing non-animal test methods for skin sensitization testing will be applying our mechanistic understanding of ACD to the design of predictive in vitro alternative test methods. Specifically, the in vitro approach should be designed so that a chemical's potential to penetrate the skin, react with protein/peptide (biotransformation may be required) and initiate an antigen specific immune response is incorporated in the test methods developed. In this review, we have focused on cellular-based assays that have been developed or proposed for assessing a chemical's skin sensitization potential in vitro. All of the promising leads to date are based on observations made from in vivo studies conducted in animals and humans, and therefore have a strong mechanistic foundation. However, it remains to be demonstrated whether a single in vitro test, or several in vitro tests in combination, which model the critical steps in sensitization, can replace animal experiments for predicting contact allergic reactions in humans. Regardless, the future looks promising with continued development of our understanding of the chemical and biological aspects of allergic contact dermatitis, and most importantly, with the application of genomics/proteomics to this field on the immediate horizon. PMID- 11259870 TI - A prevalidation study on in vitro tests for acute skin irritation. results and evaluation by the Management Team. AB - A prevalidation study on in vitro tests for acute skin irritation was conducted during 1999 and 2000. The overall objective of validation in this area, of which this prevalidation study is an initial stage, is to identify tests capable of discriminating irritants (I) from non-irritants (NI), as defined according to European Union (EU) risk phrases ("R38"; no classification) and the harmonised OECD criteria ("Irritant"; no label). This prevalidation study specifically addressed aspects of: protocol refinement (phase I), protocol transfer (phase II), and protocol performance (phase III), in accordance with the prevalidation scheme defined by the European Centre for the Validation of Alternative Methods (ECVAM). The tests evaluated were: EpiDerm (phases I, II and III), EPISKIN (phases I, II and III), PREDISKIN (phases I and II, and additional protocol refinement), the non-perfused pig ear method (phases I and II, and additional protocol refinement), and the mouse skin integrity function test (SIFT; phases I and II). Modified, standardised test protocols and well-defined prediction models were available for each of the tests at the end of phase I. The results of phase I (intralaboratory reproducibility) were sufficiently promising for all of the tests to progress to phase II. Protocol transfer between the Lead Laboratory and Laboratory 2 was undertaken for all five tests during phase II, and additional refinements were made to the test protocols. For EpiDerm, EPISKIN and the SIFT, the intralaboratory and interlaboratory reproducibilities were acceptable; however, better standardisation of certain aspects of the test protocols was needed prior to commencing phase III. Neither PREDISKIN nor the pig ear test performed sufficiently well in phase II to progress to phase III. The PREDISKIN protocol was overly sensitive, resulting in the prediction of all the NI chemicals as I. The variability in the pig ear test results was too great, indicating that the test would show limited predictive ability. In additional studies (a repeat of phase I), further modification of the PREDISKIN protocol and a change in the prediction model considerably improved the ability of the test to distinguish I from NI chemicals. However, attempts to improve the intralaboratory reproducibility of the pig ear test were unsuccessful. In phase III an initial assessment of the reproducibility and predictive ability, in three independent laboratories per test, was undertaken for the EpiDerm and EPISKIN tests (the SIFT was a late inclusion in the prevalidation study, and is being evaluated in a separate phase III study). A set of 20 coded chemicals (10 I, 10 NI) were tested with the final, refined, test protocols. The intralaboratory reproducibility was acceptable for both EpiDerm and EPISKIN. The interlaboratory reproducibility was considered to be acceptable for EPISKIN; however, for EpiDerm, analysis of variance (ANOVA) indicated that there was a statistically significant laboratory effect on the overall variability, suggesting that the interlaboratory transferability of the test needs to be improved. The EpiDerm test had an overall accuracy of 58%, with an over-prediction rate of 37% and an under-prediction rate of 47%. The EPISKIN test had an overall accuracy of 58%, showing an under prediction rate of 23% and an over-prediction rate of 60%. It is concluded that, as yet, none of the tests evaluated in this prevalidation study are ready for inclusion in a formal validation study on in vitro tests for acute skin irritation. Overall protocol performance of the SIFT is currently being evaluated in a phase III study. Further studies are also in progress to improve the test protocols and prediction models for EpiDerm and EPISKIN. PMID- 11259871 TI - Mediterranean dairy sheep and goat products and their quality. A critical review. AB - Sheep and goat edible products (mainly meat and dairy products) have interesting characteristics in their levels of flavour, taste, aromas and leanness as well as the specific composition of fats, proteins, amino and fatty acids. Their quality is very much linked to historical and cultural uniqueness right through the production, marketing and consumption chains. This refers, at least in the Mediterranean region, to farming systems with dominant extensive grazing situations, specific technologies and conditions for slaughtering as well as for the transformation processes of cheese-making and its maturing; they are also characterised by traditional nutritional habits of the consumers. While the organoleptic properties of the dairy products are very important, the sanitary aspects become more and more influential and tend to modify the accepted definitions of these products and their quality (e.g. non-pasteurised milk, production chains, hygiene, transformation methodologies, etc.). Today there are major research efforts to ameliorate the production aspects but also the quality of the products; better cheese yield with the work undertaken on alpha-s1-casein, flavour and taste of cheeses is improved by the work on lipolysis and the study of the molecules responsible for the taste of the cheeses, studies on the fattening level of animals including genetic variability, and a better understanding of the fat distribution on carcasses. The future evolution of these products is difficult to foresee. Will quality be the criterion which will give the market direction in the developed countries (standardised products and top quality special products)? In the developing regions will we be able to maintain the actual production which is well adapted to the local demand or will this not be handicapped by the fragility and marginalisation of the existing livestock farming systems and untenable rises in costs? PMID- 11259872 TI - Influence of dietary carnitine in growing sheep fed diets containing non-protein nitrogen. AB - The influence of supplemental L-carnitine was investigated in growing sheep fed rations containing non-protein nitrogen (NPN). The experiment was conducted as a randomized block design with a 2x2 factorial arrangement of treatments. Lambs (77.4kg BW, n=24) were fed a total mixed ration (12.1-13.6% CP) with two levels of L-carnitine (0 or 250ppm) and two levels of NPN (urea contributing 0 or 50% of total dietary N) for a 50-day period. Jugular blood samples were collected at 0, 1, and 3h post-feeding, and ruminal fluid samples were collected at 1h post feeding, during days 1, 8, 29, and 50 of the experiment. Average daily gain (121 versus 214g) was lower (P<0.0001) in lambs fed the NPN diets. Lambs consuming diets containing NPN had higher (P<0.0001) ruminal fluid pH (6.6 versus 5.9), ruminal ammonia N (4.8 versus 2.8mmol/l), and plasma ammonia N (177.1 versus 49.5umol/l) than lambs not fed NPN. Additionally, lambs fed the NPN diets had lower plasma urea N (14.5 versus 17.5mmol/l; P<0.003) and thyroxine (T(4)) concentrations (65.8 versus 78.4ng/ml; P<0.02), and lower T(4):triiodothyronine (T(3)) ratio (37.9 versus 43.9; P<0.02). Plasma glucose concentrations were higher (P<0.05) in lambs fed L-carnitine (3.83 versus 3.70mmol/l). Two oral urea load tests (OULT 1 and OULT 2) were conducted during the 50-day trial. Urea solutions (0.835g/kg(0.75) BW) were administered as oral drenches. During the OULT 1 (day 10), plasma ammonia N and glucose concentrations were highest (P<0.0001) in the lambs fed NPN with L-carnitine compared with lambs fed control, L-carnitine, and NPN diets. During the OULT 2 (day 50), plasma ammonia N was highest (P<0.0001) in the NPN and NPN with L-carnitine groups compared with the control and L-carnitine groups. Plasma glucose was lowest (P<0.04) in the NPN with L-carnitine group compared with the NPN and L-carnitine groups, but did not differ (P>0.10) from the control group. Plasma urea N levels in both OULT 1 and OULT 2 were lower (P<0.0001) in the NPN and NPN with L-carnitine groups compared with the control and L-carnitine groups. In the present experiment, production and plasma criteria were affected by NPN incorporation in the diets. Production criteria were not affected by inclusion of L-carnitine in the diet, however, L carnitine reduced experimentally induced hyperammonemia by day 50 of the trial. PMID- 11259873 TI - Effect of dietary propionate on intake patterns and fatty acid composition of adipose tissues in lambs. AB - The aim of the present trial was to study the effect of dietary propionate supplementation on growth performance, intake patterns and on the proportion of odd-numbered and methyl-branched chain fatty acids in internal or in subcutaneous adipose tissues. These fatty acids are responsible, in part, for abnormally soft subcutaneous adipose tissue. Eleven male lambs were fed ad libitum carbohydrate rich diets based mainly on barley, supplemented (P diet) or not supplemented (C diet) with 5.6% DM of sodium propionate. During the trial, the intake patterns were recorded on three non-consecutive days over a 9h-period. The fatty acid compositions of four adipose tissue sites, two subcutaneous (dorsal, caudal) and two internal (omental, perirenal) were analysed. The ME intake (3.32+/-0.44Mcal per day) and average daily gain (316+/-64g per day) were not different (P>0.10) between the C and P treatments. The intake patterns of these lambs given ad libitum access to feed did not show any large meal even after the morning distribution of feed. No differences in intake patterns were observed between the two diets. The dietary propionate supplementation increased (P<0.05) the proportion of odd-numbered fatty acids (Odd FA) in both internal (increase of 0.7wt.% of total fatty acids) and subcutaneous adipose tissues (increase of 1.7% of total fatty acids). The effect of dietary propionate supplementation on branched-chain fatty acids was less conclusive although it tended to increase (P=0.09) the proportion of branched-chain fatty acids other than the iso and anteiso fatty acids (BCFAO) in dorsal adipose tissue. This experiment confirms the role of propionate as an important precursor of odd-numbered and methyl branched chain fatty acids. Moreover, the dorsal adipose tissue seems to be the most sensitive adipose tissue to dietary increment in ruminal propionate content. A positive relationship between the mean ME intake rate measured over 8.5h and the proportions of Odd FA and BCFA in dorsal adipose tissue was shown. However, it does not appear to play a predominant role in the explanation of the individual variability in dorsal fatty acid composition. PMID- 11259874 TI - Economic sustainability of small ruminants production in semi-arid areas of Lebanon. AB - Small ruminant production in the near east region is facing serious constraint of feed availability. A study was conducted in marginal areas of Lebanon, using a cost-benefit analysis technique (CBA), to assess the feasibility of four small ruminant production systems ranging from semi-nomadic to settled. When the owned labor cost was included as an opportunity cost in the economic analysis, CBA revealed negative returns in all four systems. Only the settled system was profitable according to the financial analysis excluding labor cost. Moreover, feed expenses if coupled with grazing costs represented a major constraint to profitability. To remedy to the feed deficit problem, the potential of using agro industrial by-products as feed block supplements was investigated. Simulated feed block diets, using the most available by-products, provided a better nutritive value per unit cost than hand-fed diets used in the systems studied. Preliminary analysis showed that the use of feed block diets could improve the economic sustainability of small ruminant production systems. PMID- 11259875 TI - Citrus pulp and wheat straw silage as an ingredient in lamb diets: effects on growth and carcass and meat quality. AB - Twenty "Merinizzata Italiana" lambs were introduced to two experimental diets. Ten animals (five males and five females, control group) received the traditional diet that is supplied by farmers in southern Italy, which comprised of oat hay ad libitum and commercial concentrate. The second group (the same number of lambs, silage group) received citrus pulp and wheat straw silage ad libitum and 70% of the commercial concentrate supplied to the control group. The animals were slaughtered after 80 days of feeding and carcass and some meat quality parameters were measured. No differences were observed for live weight between treatments, and carcass weights were similar for the two diets, but with obvious differences between sexes. Animals on silage produced carcasses with a better muscular conformation and with a lower fatness score (P<0.05). Subcutaneous fat colour was influenced by sex, being lighter in the female carcasses (P<0.05). Dissection of different anatomical parts showed a higher percentage of lean and a lower proportion of fat in silage samples compared to control ones. Ultimate pH was highly affected by sex being higher in the samples from male lambs (P<0.01), but was unaffected by diet treatment. Diet tended to affect meat shear force value which was lower in the silage samples, although, samples from all the animals were extremely tender. Meat from silage samples had a higher water content (P<0.05). Overall, in our experimental conditions, the use of citrus pulp silage seemed to be economically convenient for producing animals with substantially unmodified carcass and meat quality characteristics. PMID- 11259876 TI - Effect of epidermal growth factor on steroidogenesis by caprine granulosa cells in culture: interaction with FSH. AB - Besides gonadotrophins various peptide growth factors have been implicated in the ovarian folliculogenesis. In this study, the effect of epidermal growth factor (EGF) (0, 0.1, 1.0, 10ng/ml culture medium) on steroidogenesis by caprine granulosa cells at various stages of maturation was investigated using serum free culture medium. Caprine granulosa cells were obtained from ovarian follicles and classified into three classes: small (<3mm), medium (3-6mm) and large (>6mm in diameter). EGF (10ng/m culture medium) alone reduced estradiol secretion in granulosa cell from small, medium and large follicles by 62, 48 and 29%, respectively, as compared with control. This inhibition was 50, 36 and 21%, respectively, when EGF (10ng/ml culture medium) was applied in combination with FSH (100ng/ml culture medium). EGF alone stimulated the secretion of progesterone in granulosa cells from all the three categories of follicles only at highest dose tested (10ng/ml culture medium). FSH acted synergistically with EGF in stimulating progesterone secretion by cultured granulosa cells. EGF in combination with FSH (100ng/ml culture medium) significantly (P<0.05) stimulated progesterone secretion by cultured granulosa cells from all three categories of follicles even at the lowest dose (0.1ng/ml culture medium) tested. In conclusion, EGF significantly influences the steroidogenesis by caprine granulosa cells in vitro and may play important role in the follicular growth and maturation. PMID- 11259877 TI - Nutritional management during fetal and postnatal life, and the influence on testicular stereology and Sertoli cell numbers in Corriedale ram lambs. AB - The aim of the experiment was to determine whether supplementation of the lamb ewe unit during intra-uterine and postnatal life affects testicular stereology, particularly Sertoli cell numbers, in 120 pregnant Corriedale ewes grazed either native pastures (control group) or improved pastures+grain supplement (treated group). Ewes bearing single ram lambs were maintained under the same feeding regime until lambs were castrated (99 days of age). Body weight, testosterone and FSH blood serum levels were recorded at 45, 75 and 99 days of age. Body weight was higher (P<0.01) in the treated group from birth on. Serum testosterone values did not differ between groups. Serum FSH values tended to differ at 45 days of age (P<0.06). Testicular weight and testes histology showed earlier pubertal development and a tendency for higher Sertoli cell numbers in the treated (supplemented) group. This tendency may indicate that extensively reared lambs supplemented during fetal and postnatal life have higher testicular growth and sperm production in later life. PMID- 11259878 TI - Estradiol-17beta and progesterone in the peripheral blood plasma of goats following superovulation with a single dose of pFSH, hMG or eCG. AB - During the breeding season, 42 adult German Improved Fawn nanny goats were superovulated with a single dose of pFSH, hMG or eCG at the end or a single application of pFSH 36h before the end of synchronization treatment using flugestone acetate (FGA). Plasma sampling was performed immediately before and 1h after gonadotrophin treatment, twice daily during pre-estrus and estrus and once daily during post-estrus in order to determine peripheral estradiol-17beta and progesterone levels. During that period, ovarian dynamics was followed by serial ultrasound scans (8h interval during pre-estrus and estrus and once daily during post-estrus). Estradiol-17beta profiles differed between the treatment groups exhibiting significantly positive correlations between the mean estradiol-17beta concentrations and the numbers of developing large and medium-sized follicles during estrus. The early bolus application of FSH 36h before the end of synchronization treatment induced an additional advanced estradiol-17beta peak during gestagen dominance. A sharp decrease of estradiol-17beta at the end of estrus seems to play a major role for normal luteal development. Progesterone profiles during the early luteal phase revealed high premature luteal regression. An early progesterone increase was accompanied by low premature regression rates.Although simple B-mode ultrasonography is suitable to follow follicular growth patterns and to determine the ovulation rate following different superovulation regimen endocrinological supervision is required in order to detect a premature corpus luteum insufficiency. PMID- 11259879 TI - Ultrasonic survey of follicular development following superovulation with a single application of pFSH, eCG or hMG in goats. AB - During the breeding season, 42 adult German Improved Fawn nanny goats were superovulated with a single administration of pFSH, hMG or eCG at the end or a single dose of pFSH 36h before the end of estrus synchronization. Development of follicles and corpora lutea were observed by ultrasonic scanning of the ovaries every 8h from gonadotrophin treatment, until the end of estrus and once daily for the following 6 days. Differences in follicular dynamics could be realized in the four superovulation groups, and the duration of the stimulatory action following the single gonadotrophin challenge was estimated to last for 40-72h in the case of pFSH and to exceed 72h in the case of hMG and eCG treatment groups. Corpora lutea could first be detected 3 days after estrus, whereas an exact count was not possible until day 6. The ovulation rates were satisfactory, suggesting that a single injection of pFSH or hMG provides an adequate stimulus to induce a superovulatory reaction. Premature regression of corpora lutea could not be identified ultrasonographically at these early stages of the luteal phase. However, ultrasonography is a suitable method to follow follicular dynamics after superovulation and to estimate the superovulatory response in goats. PMID- 11259880 TI - Effect of polyethylene glycol on rumen volume and retention time of liquid and particulate matter along the digestive tract in goats fed tannin-rich carob leaves (Ceratonia siliqua). AB - The present work studied the effects of tannins in carob leaves (CL) on rumen volume and kinetics, and on the retention time of fluid and particulate components of the digesta along the gastrointestinal tract (GIT) in goats. The experimental design was a two factor crossover experiment, i.e. in phase 1, two goats were fed CL and 2 CL and polyethylene glycol (PEG) and in phase 2, the treatments were switched. The main effects of tannins were depression of the rumen fluid and particulate content of the rumen, acceleration of the passage of liquid from the abomasum, and delay of the passage of digesta in the intestine. The overall effect was a delay in the passage of fluid and particulate matter throughout the entire GIT. It is hypothesised that these responses are largely the consequence of the interaction of tannins with digestive enzymes and the epithelium lining of the digestive tract. PMID- 11259881 TI - The application of ontologies and problem-solving methods for the development of shareable guidelines. AB - Recently, studies have shown the benefits of using clinical guidelines in the practice of medicine. Computer-based clinical guidelines are increasingly applied in diverse areas such as policy development, utilization management, education, conduct of clinical trials, and workflow facilitation. This paper discusses some of the representations suggested in literature, discusses their weak and strong points, and demonstrates and discusses a new approach that extends earlier developed formalisms by combining primitives, ontologies and the use of problem solving methods (PSMs). The approach is supported by a framework that facilitates the entire guideline authoring process. The paper demonstrates this framework and presents examples of guidelines, PSMs and systems that were developed by means of this approach. The overall goal of this approach is to improve the acceptance of shareable guidelines and decision support systems in daily care by facilitating the guideline acquisition and execution phases. PMID- 11259882 TI - Knowledge-based verification of clinical guidelines by detection of anomalies. AB - As shown in numerous studies, a significant part of published clinical guidelines is tainted with different types of semantical errors that interfere with their practical application. The adaptation of generic guidelines, necessitated by circumstances such as resource limitations within the applying organization or unexpected events arising in the course of patient care, further promotes the introduction of defects. Still, most current approaches for the automation of clinical guidelines are lacking mechanisms, which check the overall correctness of their output. In the domain of software engineering in general and in the domain of knowledge-based systems (KBS) in particular, a common strategy to examine a system for potential defects consists in its verification. The focus of this work is to present an approach, which helps to ensure the semantical correctness of clinical guidelines in a three-step process. We use a particular guideline specification language called Asbru to demonstrate our verification mechanism. A scenario-based evaluation of our method is provided based on a guideline for the artificial ventilation of newborn infants. The described approach is kept sufficiently general in order to allow its application to several other guideline representation formats. PMID- 11259883 TI - ONCODOC: a successful experiment of computer-supported guideline development and implementation in the treatment of breast cancer. AB - Originally published as textual documents, clinical practice guidelines have poorly penetrated medical practice because their editorial properties do not allow the reader to easily solve, at the point of care, a given medical problem. However, despite the proliferation of implemented clinical practice guidelines as decision support systems providing an easy access to patient-centered information, there is still little evidence of high physician compliance to guidelines recommendations. Apart from physicians' psychological reluctance, the incompleteness of guideline knowledge and the impreciseness of the terms used, another reason may be that, although suited to average patients, clinical practice guideline recommendations are not a substitute for the physician controlled clinical judgement that should be applied to each actual individual patient. Therefore, computer-based approaches based on the automation of context free operationalization of guideline knowledge, although providing uniform optimal strategies to problem-focused care delivery, may generate inappropriate inferences for a specific patient that the physician does not follow in practice. Rather than providing automated decision support, ONCODOC allows the clinician to control the operationalization of guideline knowledge through his hypertextual reading of a knowledge base encoded as a decision tree. In this way, he has the opportunity to interpret the information provided in the context of his patient, therefore, controlling his categorization to the closest matching formal patient. Experimented in life-size ONCODOC demonstrated good appropriation of the system by physicians with significantly high scores of compliance. We successfully tested the implemented strategy and the knowledge base in a second medical institution, giving then a noticeable example of reuse and sharing of encoded guideline knowledge across institutions. PMID- 11259884 TI - Flexible guideline-based patient careflow systems. AB - Workflow Management Systems integrate domain and organisational knowledge to support business processes. When applied to the medical environment, they can be termed "Careflow Management Systems", and may be used to manage care delivery by enhancing co-operation among healthcare professionals. This paper focuses on care delivery based on clinical practice guidelines. Healthcare organisations are very different from industrial or commercial companies: their main goal is not profit, but maintaining and improving the health of the public. Therefore, outcomes are difficult to measure. Firstly, physicians, while playing a variety of roles, are quite independent decision-makers; secondly, the object of the process, i.e. the patient, may be involved in choosing treatment options, and may be treated by different institutions. For these reasons, the standard functionality of typical Workflow Management Systems must be strongly enhanced in order to cope with healthcare delivery needs. A major issue is accounting for exceptions. In most non-clinical settings this is not a problem because processes are very well defined and can often be easily controlled by some higher authority. As explained above, this does not happen in healthcare organisations. Responsibilities are widely shared, and health care professionals may be non-compliant with guidelines for a variety of reasons. The paper presents a classification of possible exceptions, and shows how the sequence of tasks described by a guideline may be altered, at the implementation level, in order to meet actual user needs, while maintaining guideline intentions as much as possible. A terminology server is also exploited towards this end. This work illustrates a prototype of a Careflow Management System based on an international guideline for ischemic stroke treatment, developed by the American Heart Association. PMID- 11259885 TI - Welcome to the association of anesthesia clinical directors. PMID- 11259886 TI - Comparison of wire reinforced tubes with warmed standard tubes to facilitate fiberoptic intubation. AB - STUDY OBJECTIVE: To compare the ease of insertion of a warmed standard tracheal tube to that of a wire reinforced tracheal tube when placed over a flexible fiberoptic bronchoscope. DESIGN: Randomized controlled trial. SETTING: Tertiary care hospital. PATIENTS: 50 patients undergoing elective general anesthesia. INTERVENTIONS: Patients' tracheas were intubated with a flexible fiberoptic bronchoscope and had either a standard or wire-reinforced tracheal tube inserted. If resistance was met, the tube was withdrawn, rotated, and readvanced. This was repeated two times. If unsuccessful, the flexible fiberoptic bronchoscope was removed, and intubation was attempted with the other type of tracheal tube. MEASUREMENTS: The ability to advance the tracheal tube was determined. MAIN RESULTS: There were no demographic differences between the two groups. There was a similar ease of advancement of the two tracheal tubes. CONCLUSIONS: When performing elective flexible fiberoptic bronchoscopy for intubation, we recommend using the less expensive warmed standard tracheal tube. PMID- 11259887 TI - Comparison of no airway device, the Guedel-type airway and the Cuffed Oropharyngeal Airway with mask ventilation during manual in-line stabilization. AB - STUDY OBJECTIVE: To compare two different types of oropharyngeal airway: the Guedel-type oral airway and the Cuffed Oropharyngeal Airway (COPA), with respect to the effectiveness of positive-pressure ventilation (PPV) through a face mask in patients with in-line stabilization of the head and neck. DESIGN: Prospective, randomized, crossover study. SETTING: University hospital. PATIENTS: 30 ASA physical status I and II patients undergoing elective surgery. INTERVENTIONS: General anesthesia was induced with propofol and muscle relaxation was produced with vecuronium. In a random sequence, no airway device, the Guedel-type airway, and the COPA were used in each patient while applying a face mask and lifting the jaw forward. MEASUREMENTS AND MAIN RESULTS: Tidal volumes were measured during PPV in each option. The position of the distal tip of each airway was assessed using a fiberscope, and the resulting views were graded and compared. When the Guedel-type airway was used, tidal volumes (V(T)s; means +/- SD) were significantly greater (12.3 +/- 4.5 mL/kg) than those with no airway device (8.5 +/- 4.5 mL/kg) (p < 0.001). When the COPA was used, V(T)s (14.6 +/- 4.4 mL/kg) were significantly greater than those with the Guedel-type airway (p < 0.05). The grade of the fiberscopic view through the distal tip was significantly better with the COPA than with the Guedel-type airway (p < 0.05). CONCLUSIONS: Although clinical differences often appear trivial, the COPA is more effective on mask ventilation than the Guedel-type airway when used in patients with manual in-line stabilization. PMID- 11259888 TI - When should diclofenac be given in ambulatory surgery: preoperatively or postoperatively? AB - STUDY OBJECTIVE: To determine the optimum time of administration of diclofenac in patients undergoing ambulatory knee arthroscopy: either preoperatively or postoperatively. DESIGN: Randomized, double-blind study. SETTING: Ambulatory surgical unit in a tertiary referral hospital. PATIENTS: 127 ASA physical status I and II patients undergoing ambulatory knee arthroscopy. INTERVENTIONS: Patients were randomized into three groups. The Preop group received 50 mg of potassium diclofenac orally 1 hour preoperatively and a placebo 30 minutes postoperatively. The Pre+postop group received 25 mg of potassium diclofenac 1 hour preoperatively and 25 mg diclofenac 30 minutes postoperatively. The Postop group received a placebo 1 hour before surgery and 50 mg of potassium diclofenac 30 minutes postoperatively. MEASUREMENTS AND MAIN RESULTS: The Postop group received a placebo 1 hour preoperatively and 50 mg of potassium diclofenac 30 min postoperatively. Postoperatively, patients used intravenous patient-controlled analgesia (PCA) with fentanyl. Total fentanyl consumption was recorded. During the recovery period, pain was assessed using a visual analog scale (VAS) at 30 minute intervals. Pain was assessed in both legs at rest, on flexion, and extension of the knee. There were no significant differences in pain scores either at rest or on movement of the operative knee among the Preop, Pre+postop, and Postop groups. The consumption of fentanyl via PCA showed no significant differences among the groups. CONCLUSIONS: There is no difference in pain relief whether diclofenac is given preoperatively or postoperatively in patients undergoing unilateral ambulatory knee arthroscopy. Preoperative and postoperative treatment with diclofenac potassium is equally effective. PMID- 11259889 TI - Nicardipine versus nitroprusside for breakthrough hypertension following carotid endarterectomy. AB - STUDY OBJECTIVE: To evaluate the effectiveness of nicardipine and nitroprusside for breakthrough hypertension following carotid endarterectomy. DESIGN: Prospective, randomized, double-blind, controlled effectiveness trial. SETTING: University-based surgical intensive care unit. PATIENTS: 60 ASA physical status I, II, III, and IV patients experiencing breakthrough hypertension at the time of admission to the intensive care unit (ICU). INTERVENTIONS: Patients received either nicardipine (n = 29) and placebo or nitroprusside (n = 31) and placebo for up to 6 hours postoperatively. Loading doses of nicardipine were provided, but placebo was used as a load for patients randomized to nitroprusside. MEASUREMENTS AND MAIN RESULTS: Rapidity and variability of blood pressure (BP) control were assessed. During the first 10 minutes, 83% of nicardipine patients compared to 23% of nitroprusside-treated patients, achieved BP control (p < 0.01). Following initial control, 12 nicardipine- and 24 nitroprusside-treated patients required additional titration of their infusions to maintain blood pressure within the targeted range (p < 0.05). No patient suffered a stroke, myocardial infarction, or was returned to the operating room (OR) for bleeding. CONCLUSIONS: Nicardipine administration produced more rapid BP control, most likely related to the administration of a loading dose. In addition to more rapid control, nicardipine treated patients had less variability in BP and required significantly fewer additional interventions. Although no patient suffered a major event during this study, this study was not powered sufficiently to assess safety. PMID- 11259890 TI - Celite-activated thrombelastography in children. AB - STUDY OBJECTIVE: To quantify global coagulation and establish normal ranges for the celite-activated thrombelastograph (TEG) in healthy pediatric patients. DESIGN: Prospective observational study. SETTING: Operating suite of a university based hospital. PATIENTS: 110 healthy pediatric patients in four age groups and 25 healthy adult patients. INTERVENTIONS: Blood sampling for the celite-activated TEG was carried out after anesthetic induction. MEASUREMENTS: TEG indices: R time (reflecting time to fibrin formation), K time and alpha angle (fibrinogen platelet interaction), maximum amplitude (reflecting maximal clot strength, platelet and fibrinogen function), TEG index (mathematical incorporation of the prior four measurements), and percent fibrinolysis at 30 minutes, were all recorded. MAIN RESULTS: Statistically significant differences between <12-month group in angle (compared to 25-48 month group) and % fibrinolysis (compared to all other pediatric groups). Significant differences in angle between two pediatric groups and adult group, and in the TEG index between three pediatric groups and adult group (all differences p < 0.05). CONCLUSIONS: These data identify changes of small magnitude in three celite-TEG parameters in healthy children compared to adults, without implication of abnormal coagulation between groups. Changes do not seem to be consistently related to age and will be useful for clinicians using the TEG to monitor (ab) normal coagulation in pediatric patients. PMID- 11259891 TI - The relationship of sedation to deliberate self-extubation. AB - STUDY OBJECTIVES: To evaluate the relationship between sedative therapy and self extubation in a large medical-surgical intensive care unit (ICU). DESIGN: Retrospective, case-controlled study. SETTING: Large teaching hospital. PATIENTS: All adult patients who underwent unplanned self-extubation during a 12-month period (n = 50). Each patient was matched to two control patients who did not self-extubate based on age, gender, dates in hospital and diagnosis. INTERVENTIONS: none. MEASUREMENTS: Data collected included time to self extubation, dosages and types of benzodiazepines, opioid analgesics, antipsychotics, and hypnotics. Data on the degree of agitation as assessed by nursing staff also were obtained. MAIN RESULTS: When compared to controls, patients in the self-extubation group were more likely to have received benzodiazepines (59% vs. 35%; p < 0.05), but equally likely to have received opioids and/or paralytic drugs. Patients who self-extubated were twice as likely as controls to be agitated (54% vs. 22%; p < 0.05). Use of benzodiazepines was more common in agitated patients than in nonagitated patients (62% vs. 35%; p < 0.02). Among nonagitated patients who self-extubated, increased use of benzodiazepines (57% vs. 29%; p < 0.05) was noted when compared to nonagitated controls. CONCLUSIONS: In intubated ICU patients, benzodiazepines may not consistently treat agitation effectively or prevent self-extubation. Such an effect may be due to paradoxical excitation, disorientation during long-term administration, or differences in drug administration between ICU and operating room (OR) environments. PMID- 11259892 TI - Pediatric caudal block with mepivacaine, bupivacaine or a mixture of both drugs: requirement for postoperative analgesia and plasma concentration of local anesthetics. AB - STUDY OBJECTIVES: To assess the effects of pediatric caudal block using mepivacaine, bupivacaine, or a mixture of both drugs on postoperative analgesia, and to examine plasma concentrations of the local anesthetics after caudal injection. DESIGN: Prospective, randomized, double-blind study. SETTING: Operating room and pediatric surgical ward. PATIENTS: 60 ASA physical status I children weighing 10 to 20 kg (26 females, 34 males), and scheduled for inguinal herniorrhaphy. INTERVENTIONS: Patients randomly received caudal block with 1 mL/kg of mepivacaine 1% (Group M, n = 20), 1 mL/kg of bupivacaine 0.25% (Group B, n = 20), or a mixture of 0.5 mL/kg of mepivacaine 1% and 0.5 mL/kg of bupivacaine 0.25% (Group MB, n = 20) after induction of anesthesia with sevoflurane in 50% oxygen (O2). Anesthesia was maintained with 66% nitrous oxide in O2 supplemented with sevoflurane at an end-tidal concentration of less than 1%. MEASUREMENTS AND MAIN RESULTS: Postoperative pain scores using a pediatric pain scale and plasma concentration of each local anesthetic were measured. In Group M, four patients required postoperative analgesics within the first 24 hours. However, no patients required postoperative analgesics in Groups B and MB. In Group M, the plasma concentration of mepivacaine of two patients exceeded 5 microg/kg of the level of toxicity. However, these patients did not show any toxic symptoms. Because a mixture of two local anesthetics halves the concentration of each local anesthetic, the plasma concentrations of mepivacaine and bupivacaine in Group MB were significantly lower than those of Groups M and B. CONCLUSIONS: Pediatric caudal block with a mixture of mepivacaine and bupivacaine is effective for intraoperative and postoperative analgesia. PMID- 11259893 TI - Variations on one-lung ventilation. AB - One-lung ventilation is a commonly used technique to facilitate surgical visualization during thoracic surgical procedures. New devices for one-lung ventilation have been introduced into clinical practice over the recent years. One such device is the Arndt Endobronchial Blocker which is a bronchial blocker with a central lumen through which a wire with a looped end has been passed. The loop on the distal end is meant to be placed over a fiberoptic bronchoscope which is then used as a guide to facilitate bronchial placement of the balloon-tipped bronchial blocker. Another advantage of the Arndt device is the airway adaptor that contains ports for the anesthesia circuit, the bronchoscope, the bronchial blocker as well as attachment to the endotracheal tube. The port for the bronchial blocker can be tightened down so as to hold the blocker in place during the procedure. However, patient issues such as size or airway alterations such as the presence of a tracheostomy may make necessary certain alterations in airway management. I describe four cases and provide suggestions for minor alterations in airway management that may be used to provide successful options for one-lung anesthesia. PMID- 11259894 TI - "Apnea-volume" warning during normal ventilation of the lungs: an unusual leak in the Narkomed 4 Anesthesia System. AB - We report a case of an unusual breathing circuit leak in the Narkomed 4 Anesthesia System due to a loose retaining ring at the junction of the expiratory valve assembly and the Spiromed Respiratory Volume Monitor. In the presence of the leak, the monitor panel displayed the messages "apnea volume" and "minute volume low," yet the low airway pressure alarm was not triggered and other parameters and clinical signs pointed to normal ventilation of the lungs. These conflicting data led to some delay in localizing the leak. After conclusion of the case, we found that occult loosening of this ring without causing leaks large enough to fail the FDA generic or manufacturer-recommended leak checks can occur. We recommend checking the tightness of this ring during routine leak checks and after rotating the expiratory valve assembly during re-positioning of the anesthetic system. PMID- 11259895 TI - Negligence in supervision: a case of failed resuscitation. PMID- 11259896 TI - Commentary 1. PMID- 11259897 TI - Commentary 2. PMID- 11259899 TI - Association of Anesthesia Clinical Directors (AACD) abstracts presented at the AACD 13th annual meeting. PMID- 11259898 TI - Regional anesthesia and anticoagulation. AB - The perioperative use of neuraxial techniques in the presence of anticoagulation is a controversial issue. There are significant pharmacokinetic differences between anticoagulants that will affect the timing of neuraxial needle insertion or catheter removal. The pharmacologic profiles of commonly used anticoagulants in the perioperative period are reviewed. Studies examining the use of neuraxial techniques in the presence of various anticoagulants are reviewed and evaluated in the context of the American Society of Regional Anesthesia consensus statements. PMID- 11259900 TI - Erratum. PMID- 11259924 TI - Impaired glucose tolerance is a more advanced stage of alteration in the glucose metabolism than impaired fasting glucose. AB - BACKGROUND: Recent reports have shown a lack of agreement between the impaired glucose tolerance (IGT) and the impaired fasting glucose (IFG) categories, suggesting that correspond to different impaired glucose metabolism stages. OBJECTIVE: To determine the differences of serum insulin levels between subjects with IFG and IGT diagnoses. METHODS: Cross-sectional study of 52 subjects with IFG and 48 with IGT diagnosis, and a euglycemic group of 140 subjects. Serum glucose and insulin were measured in both fasting and 2-h 75-g oral post-load glucose (2-h PG). RESULTS: Subjects with IFG showed the highest fasting and 2-h PG serum insulin levels, whereas subject with IGT the lowest. Serum insulin values showed no significative changes between the fasting and 2-h PG conditions in the subjects with IGT, whereas the subjects with IFG showed significative hyperinsulinemia. The serum glucose 2-h PG showed an increase of 0.2 mmol/l (CI(95%) 0.07-0.33), 0.5 mmol/l (CI(95%) 0.41-0.58) and 3.6 mmol/l (CI(95%) 3.39 3.81) with respect to basal values, whereas the increase of serum insulin 2-h PG was of 54 pmol/l (CI(95%) 53.71-55.29), 918 pmol/l (CI(95%) 917.49-918.51) and 0.5 pmol/l (CI(95%) 0.15-0.84) for the euglycemic, IFG and IGT subjects, respectively. CONCLUSIONS: This study demonstrates that subjects with IFG show hyperinsulinemia whereas those with IGT have low insulin secretion in response to oral load glucose, suggesting that IFG and IGT correspond to different stages of impaired glucose metabolism. PMID- 11259923 TI - Occurrence and predictors of retinopathy and visual acuity in Type 2 diabetic patients and control subjects. 10-year follow-up from the diagnosis. AB - The evolution of visual acuity and retinopathy and their risk factors in patients with newly diagnosed type 2 diabetes and in control subjects. A 10-year prospective study consisting of a representative group of 133 (70 men, 63 women) newly diagnosed type 2 diabetic patients diagnosed at health centers between 1979 and 1981 and 144 (62 men, 82 women) non-diabetic control subjects recruited from the population register. The frequency of retinopathy was determined by grading of 45 degrees fundus photographs at baseline and after 5 and 10 years. By the 10 year follow-up the diabetic patients had lower visual acuity than the control subjects. The impairment of the visual acuity correlated inversely to HbA(1C) value of the 5-year examination. The frequency of retinopathy in type 2 diabetic patients increased sharply after 5 years and at 10-year 55% of diabetic patients had signs of retinopathy. The frequency of retinopathy in the control subjects was low, but detectable. In the diabetic patients poor glycemic control was the most important predictive factor for the development of retinopathy. In the control subjects blood pressure levels were higher and microalbuminuria more common in those with than in those without retinopathy. The visual acuity deteriorated and the frequency of retinopathy increased in newly diagnosed type 2 diabetic patients with duration of disease and poor glycemic control. Interestingly, higher blood pressure levels and microalbuminuria predicted retinopathy in control subjects. PMID- 11259925 TI - Coeliac- and enteropathy-associated autoantibodies in Spanish insulin-dependent diabetes mellitus patients and their relation to HLA antigens. AB - The frequency of reticulin (ARA), endomysium (EmA), and gut epithelial cell (GECA) autoantibodies, and gliadin antibodies (AGA), was investigated in 86 Spanish diabetic patients by indirect immunofluorescence (IFI) and ELISA, along with their HLA phenotype. Four patients (5%) showed ARA-IgG (R1 pattern), eight (9%) showed AGA-IgG, and eight (9%) showed AGA-IgA. No EmA or GECA-positive patients were found. In diabetic patients, HLA-DR7 is increased in ARA-IgG+ vs. ARA-IgG- (though not significantly), and HLA-DR6 and HLA-DQ1 are significantly increased in the AGA-IgG+ group vs. the AGA-IgG- group. Comparison with a non diabetic coeliac group showed that HLA-DR4 and HLA-DQ3 are significantly increased in the AGA-IgA+ group, whereas HLA-DQ2 shows a significant decrease in the AGA-IgG+ and AGA-IgA+ patients. Finally, when compared to the healthy group, HLA-DR7 frequency is decreased in the ARA-IgG- group, while HLA-DQ3 is significantly increased and HLA-DR6 and HLA-DQ1 significantly decreased in the AGA-IgG- group.Altogether, these data suggest that the genetic background leading to the appearance of coeliac-specific autoantibodies in Spanish diabetic patients differ depending on the autoantibody produced and is also different to the genetic background leading to diabetes in Spain. PMID- 11259926 TI - Diabetes mellitus: a hypercoagulable state. AB - Eighty percent of patients with diabetes mellitus die a thrombotic death. Seventy five percent of these deaths is due to cardiovascular complications, and the remainder is due to cerebrovascular events and peripheral vascular complications. Vascular endothelium, the primary defense against thrombosis, is abnormal in diabetes. Endothelial abnormalities undoubtedly play a role in the enhanced activation of platelets and clotting factors seen in diabetes. Coagulation activation markers, such as prothrombin activation fragment 1+2 and thrombin-anti thrombin complexes, are elevated in diabetes. The plasma levels of many clotting factors including fibrinogen, factor VII, factor VIII, factor XI, factor XII, kallikrein, and von Willebrand factor are elevated in diabetes. Conversely, the level of the anticoagulant protein C (PC) is decreased. The fibrinolytic system, the primary means of removing clots, is relatively inhibited in diabetes due to abnormal clot structures that are more resistant to degradation and an increase in plasminogen activator inhibitor type 1 (PAI-1). Increased circulating platelet aggregates, increased platelet aggregation in response to platelet agonists, increased platelet contractile force (PCF), and the presence of higher plasma levels of platelet release products, such as beta-thromboglobulin, platelet factor 4, and thromboxane B(2), demonstrate platelet hyperactivity in diabetes. This constellation of findings supports the clinical observation that diabetes is a hypercoagulable state. This article briefly reviews the published evidence for this conclusion and the putative roles played by hyperglycemia and hyperinsulinemia in its development. PMID- 11259927 TI - Becaplermin and necrobiosis lipoidicum diabeticorum: results of a case control pilot study. AB - Necrobiosis lipoidica diabeticorum (NLD) is a rare skin condition associated with diabetes, which characteristically occurs in the pretibial region of the lower limbs (Boulton et al., 1988). The lesions generally appear as well-circumscribed reddish plaques, which are most often asymptomatic, resulting primarily in cosmetic disability. Currently, there is no reliable form of treatment for NLD, although many regimens have been tried (Shall et al., 1990) PMID- 11259928 TI - Micro- and nanoscale structures for tissue engineering constructs. PMID- 11259929 TI - Comparison of computational analysis with clinical measurement of stresses on below-knee residual limb in a prosthetic socket. AB - Interface pressures and shear stresses between a below-knee residual limb and prosthetic socket predicted using finite element analyses were compared with experimental measurements. A three-dimensional nonlinear finite element model, based on actual residual geometry and incorporating PTB socket rectification and interfacial friction/slip conditions, was developed to predict the stress distribution. A system for measuring pressures and bi-axial shear stresses was used to measure the stresses in the PTB socket of a trans-tibial amputee. The FE predicted results indicated that the peak pressure of 226 kPa occurred at the patellar tendon area and the peak shear stress of 50 kPa at the anterolateral tibia area. Quantitatively, FE-predicted pressures were 11%, on average, lower than those measured by triaxial transducers placed at all the measurement sites. Because friction/slip conditions between the residual limb and socket liner were taken into consideration by using interface elements in the FE model, the directions and magnitudes of shear stresses match well between the FE prediction and clinical measurements. The results suggest that the nonlinear mechanical properties of soft tissues and dynamic effects during gait should be addressed in future work. PMID- 11259930 TI - Intramedullary femoral nails: one or two lag screws? A preliminary study. AB - Failures of proximal femoral nails that treat unstable femoral fractures have been reported. In this communication, a finite element model to include a proximal femoral nail within a fractured femur was used to carry out preliminary investigations into configurations of single or double lag screws. The effects of the different types of fracture were investigated. The results show that in order to share the load evenly between two lag screws, a good configuration seems to be to have a slightly larger screw above the lower screw. This also ameliorates stresses in the nail at the lag screw insertion holes. However, using two screws in this way can lead to large stresses in the cancellous bone in the femoral head, and these stresses may be significant in the initiation of cut-out. PMID- 11259931 TI - Construction and testing of a computer-based intraoral laser scanner for determining tooth positions. AB - An optical set-up for intraoral data acquisition based on the principle of laser triangulation was developed. The system consists of a pig-tailed laser with line generating optics, a stepping motor driven positioning stage, a commercial CCD (charge coupled device) camera system with frame grabber interface, a control personal computer and a mirror system compensating for the fact that there is no possibility of watching an object directly in the mouth under a certain angle except from a facial position during intraoral scanning. Due to the size of the prototype measurements were still restricted to plaster casts. In order to evaluate its accuracy, the measurements were compared with those taken with a commercial laser scanner and a coordinate measurement table. The accuracy of the prototype scanner was determined to be DeltaXYZ=0.04 mm using gauge blocks of given dimensions and proved to range between the commercial laser scanner and the coordinate measurement table (i.e., it was slightly better than that of the commercial scanner). Applications in orthodontics were demonstrated by scanning plaster casts and measuring distances on reconstructed surfaces. The measured distances showed a maximum deviation of about +/-0.2 mm compared with the data of the coordinate measurement table, which served as a reference. In addition, reconstruction of three-dimensional tooth movements was performed on the scan data. The translational and rotational parameters gained from the superimposition of scanned point clouds and describing tooth movement were also in good accordance with the reference. The achieved accuracy proved to be sufficient for further development which should include a reduction in size and the use of more precise device components. PMID- 11259932 TI - The variation in fatigue rate with frequency using kHz frequency alternating current. AB - The most commonly used stimulus for functional electrical stimulation is low frequency, short duration pulsed current. A disadvantage is the associated rapid fatigue. The present study investigated the effect of kHz frequency alternating currents on the rate of fatigue with electrically induced skeletal muscle contractions in normal subjects. Alternating current with frequencies between 1 and 15 kHz, interrupted at 50 Hz and applied in 3-s surges with an effective 1:1 duty cycle, was applied transcutaneously for a 10-min period, during which time electrically induced wrist extensor torque was measured. The decline in torque with time was analysed in terms of "fast" and "slow" fatigue components. A systematic frequency dependence was found in each. "Fast" torque decline is interpreted as reflecting fast-fatigue muscle fibre activity and "slow" torque decline, that of fatigue-resistant fibres. With this interpretation, over the frequency range 1-10 kHz, the proportion of fatigue-resistant fibres contributing to the torque increases. It is argued that this is due to selective dropout of fast-fatigue fibres. The findings suggest the potential usefulness of kHz frequency alternating current for functional electrical stimulation. PMID- 11259934 TI - Evaluation of a sensor for low interface pressure applications. AB - An ultra-thin, small sensor has recently been developed, "FlexiForce" (Tekscan, Boston, MA, USA), which may be effective for the measurement of low interface pressure between the skin, support surfaces and pressure garments. To evaluate the suitability of the sensor for these applications, drift, repeatability, linearity, hysteresis and curvature effects were tested under laboratory conditions. The drift was 1.7-2.5%/logarithmic time, the repeatability was 2.3 6.6% and the linearity was 1.9-9.9% in the range of forces of 10-50 g applied. The hysteresis was 5.4% on average. The output offset of the sensor increased with decreasing radius of curvature for radii less than 32 mm compared with a flat surface when no pressure was applied. The sensitivity to pressure decreased with curvature for radii less than 32 mm. It was found that the sensor had acceptable drift, repeatability, linearity and hysteresis. However, a significant curvature effect was observed indicating that the sensor is suitable for direct measurement on surfaces with the radii greater than 32 mm under static conditions. PMID- 11259933 TI - Modern application of high voltage stimulation for enhanced healing of venous crural ulceration. AB - The objective of this paper was to evaluate the effect of high voltage stimulation (HVS) on the process of healing of crural ulceration. Three comparative groups of patients, A, B and C, were drawn at random from patients with venous crural ulceration. Thirty-three patients in group A were subjected to treatment with HVS. In the course of treatment, doubled-peak monophasic impulses of a total duration of 0.1 ms and frequency of 100 Hz were applied. The voltage was 100 V. Group B, made up of 32 patients treated with topically applied medicine, and group C, made up of 14 patients treated with Unna's boot were used as control. In all patients in all groups (A, B and C) a significant reduction in wound size in relation to baseline was found. The rate of wound area change was highest in group A. The rate of pus cleansing was significantly highest in group A. The degree of granulation after 2 weeks was significantly greater in group A than in the other groups (P<0.003). High voltage stimulation was an efficient method of enhancement of the healing of crural ulceration. PMID- 11259935 TI - Lumbar spine curvature during office chair sitting. AB - Prolonged sitting is generally accepted as a high risk factor in low back pain and it is frequently suggested that a lordotic posture of the lumbar spine should be maintained during sitting. We asked whether the sagittal curvature of the lumbar spine during sitting is affected by the seat tilt, backrest and the direction of the synchronised mechanism of the back and seat tilt (synchro tilt). Two office chairs were tested by multibody analysis interfacing a human model with a chair model. Results indicate that a synchronised mechanism of an office chair representing a posterior tilt of the seat while the backrest is reclined maintains an evenly distributed lumbar lordosis. The segmental angles are between 3.1 and 3.6 degrees at the lumbar vertebrae 1/2-4/5 (L1/2-L4/5). These lumbar spine segmental angles are not sensitive to the backrest height. In contrast, a synchro tilt concept with a reduction of the seat's posterior tilt while the backrest is reclined causes a strong reduction of the lumbar lordosis in backrest recline with a maximum reduction from 11.7 to 2.8 degrees in L4/5. As a consequence of these results, a synchro tilt concept with a posterior tilt of the seat while the backrest is reclined is preferable from the lumbar spine kinematics point of view. PMID- 11259936 TI - Clinical decision-support systems for intensive care units using case-based reasoning. AB - The artificial intelligence approach used in this work focusses on case-based reasoning techniques for the estimation of medical outcomes and resource utilization. The systems were designed with a view to help medical and nursing personnel to assess patient status, assist in making a diagnosis, and facilitate the selection of a course of therapy. The initial prototype provided information on the closest-matching patient cases to the newest patient admission in an adult intensive care unit (ICU). The system was subsequently re-designed for use in a neonatal ICU. The results of a short clinical pilot evaluation performed in both adult and neonatal units are reported and have led to substantial improvement of the prototype. Future work will include longer-term clinical trials for both adult and neonatal ICUs, once all the software changes have been made to both prototypes in response to the comments of the users made during the preliminary evaluations. To date, the results are very encouraging and physician interest in the potential clinical usefulness of these two systems remains high, and particularly so in the new testing environment in Ottawa. PMID- 11259937 TI - High signal intensity of the posterior pituitary gland on T1-weighted MR images. Correlation with plasma vasopressin concentration to water deprivation. AB - PURPOSE: To evaluate the effect of water deprivation on the signal intensity of the posterior pituitary gland on T1-weighted MR images and correlate the signal intensity with the plasma vasopressin concentration. MATERIAL AND METHODS: Fifteen rabbits were studied: Group 1 (n=10) was deprived of water for 9 days and Group 2 (n=5) was replenished water for 7 days after 7-day water deprivation. MR imaging and plasma vasopressin measurement by radioimmunoassay were made before and after water deprivation and replenishment. Sequential changes of the signal intensity ratio of the posterior lobe to the pons and plasma vasopressin concentration were correlated. RESULTS: Before water deprivation, the hyperintense posterior lobe was demonstrated in all rabbits. During water deprivation, the signal intensity ratio decreased and vasopressin concentration increased gradually. On the contrary, the signal intensity ratio increased and vasopressin concentration decreased with water replenishment. The signal intensity ratio correlated well with the plasma vasopressin concentration (p<0.05). CONCLUSION: There was a negative, linear correlation between the signal intensity ratio of the posterior pituitary gland on T1-weighted MR images and plasma vasopressin concentration to water deprivation. The results support that the high signal intensity of the posterior pituitary gland on T1-weighted MR images is attributed to the normal content of vasopressin-neurosecretory granules. PMID- 11259938 TI - Contrast-enhanced ultrasound Doppler examination of the retrobulbar arteries. AB - PURPOSE: To evaluate the diagnostic usefulness of an ultrasound contrast agent in examination of the retrobulbar arteries. MATERIAL AND METHODS: Ten healthy volunteers received a galactose-based echo-contrast medium (Levovist) by i.v. infusion. The ophthalmic, central retinal, the nasal and the temporal posterior ciliary arteries and the short ciliary arteries were studied in 19 eyes by color and spectral Doppler ultrasonography before and after contrast administration. Peak systolic and end diastolic velocities and spectral Doppler indices (pulsatility and resistive) were assessed. The quality of the spectral and color Doppler imaging of the arteries were visually graded on a 5-point scale. RESULTS: There were significant differences in pre- and post-contrast peak systolic velocities in the ophthalmic arteries (p<0.05), but not in the other retrobulbar arteries, or in any of the spectral Doppler indices. After contrast administration the mean spectral Doppler score for vessels poorly visualized before contrast increased from 2.2 (+/-0.4) to 3.1 (+/-0.9). The number of short ciliary arteries with sufficient spectral Doppler quality increased from 7 prior to contrast to 14 following contrast. Prior to the infusion of Levovist, 62 (73%) out of 85 retrobulbar arteries could be evaluated with a sufficient spectral Doppler quality. Following the administration of contrast 66 (78%) arteries had sufficient spectral Doppler quality. However, by combining the results of the pre and post-contrast examinations, sufficient spectral Doppler quality was obtained in 77 (91%) of the 85 retrobulbar arteries. CONCLUSION: Contrast enhancement increased the number of detectable retrobulbar vessels. However, in the case of good quality pre-contrast imaging of the retrobulbar vessels, the use of Levovist did not add any substantial diagnostic information. The optimal spectral Doppler results were obtained when both pre- and post-contrast examinations were performed. PMID- 11259939 TI - Cerebral air emboli from angiography in a patient with stroke. A case report. AB - A 42-year-old woman with subarachnoid and intracerebral hemorrhage was investigated with diagnostic angiography, disclosing an occlusion of the left internal carotid artery and the middle cerebral artery. CT examination immediately after the angiography revealed a 12-h-old infarct of the left middle cerebral artery territory. There was also gas in the arteries supplying the infarcted part of the brain, but not in other vessels. Air had most probably been introduced during the angiography and had consequently been "trapped" in the cortical arteries of the ischemic brain. In the non-ischemic parts of the brain air may have passed through the vessels leaving no trace. PMID- 11259940 TI - Complementary role of MR imaging of ethmomaxillary sinus disease depicted at CT in cystic fibrosis. AB - PURPOSE: To assess whether MR imaging can improve characterization of ethmomaxillary opacification diagnosed at CT in patients with cystic fibrosis (CF) in order to select patients that may benefit from functional endoscopic sinus surgery (FESS). MATERIAL AND METHODS: Sixty-two CF patients (26 females and 36 males) aged 4-50 years (median 20 years) with ethmomaxillary sinus disease at CT underwent MR examination of the paranasal sinuses (coronal T1 and STIR sequences). FESS had been performed in 28 of the patients prior to this study. MR signal intensities were interpreted as mucosal thickening or infectious material, according to a previous study. RESULTS: Three major maxillary sinus MR patterns could be distinguished: Air-filled, oval-shaped pus-filled, and streaky-shaped pus-filled sinus lumen. For air-filled maxillary sinuses with mucosal thickening, CT and MR imaging were diagnostically equivalent. Where CT showed homogeneous opacification of the maxillary sinuses, MR imaging differentiated between thickened mucosa and pus-filled areas. Patients who had undergone FESS most commonly had air-filled or streaky-shaped pus-filled maxillary sinus lumen. In non-operated patients oval-shaped pus-filled sinus lumen was most common and could occur without ethmoid disease. CONCLUSION: MR imaging of the paranasal sinuses can differentiate between infectious material and thickened mucosa and should be used to select CF patients with pus-filled areas that can be eradicated with FESS. PMID- 11259941 TI - CT-guided percutaneous core biopsies of pulmonary lesions. Diagnostic accuracy, complications and therapeutic impact. AB - PURPOSE: To evaluate the diagnostic accuracy, complications, and therapeutic impact of CT-guided percutaneous core biopsies of pulmonary lesions. MATERIAL AND METHODS: Seventy-nine patients underwent diagnostic CT-guided percutaneous core biopsies of pulmonary lesions between July 1995 and March 1999. Evaluation included corresponding clinical data, pathologic results, and therapeutic consequences. RESULTS: There were 29 benign and 50 malignant lesions. Percutaneous core biopsy had an overall diagnostic accuracy of 95%. For malignant lesions, core biopsy was positive in 48 patients (sensitivity 96%), and for benign lesions, in 27 (sensitivity 93%). There were no false-positive findings. Pneumothoraces were observed in 19 patients (24%) and 4 of them required a chest drain (5%). There were no hematothoraces or major bleeding complications; however, postinterventional local hemorrhages were observed in 23 patients (29%). No hemoptysis was noted. CONCLUSION: Percutaneous core biopsies of pulmonary lesions offer excellent diagnostic accuracy for both benign and malignant pulmonary lesions at a low complication rate. PMID- 11259942 TI - Effects of JPEG and wavelet compression of spiral low-dose ct images on detection of small lung cancers. AB - PURPOSE: To compare the effect of compression of spiral low-dose CT images by the Joint Photographic Experts Group (JPEG) and wavelet algorithms on detection of small lung cancers. MATERIAL AND METHODS: Low-dose spiral CT images of 104 individuals (52 with peripheral lung cancers smaller than 20 mm and 52 control subjects) were used. The original images were compressed using JPEG or wavelet algorithms at a ratio of 10:1 or 20:1. Five radiologists interpreted these images and evaluated the image quality on a high-resolution CRT monitor. Observer performance was studied by receiver operating characteristic (ROC) analysis. RESULTS: There was no significant difference in the detection of cancers measuring 6 to 15 mm in uncompressed images and in those compressed by either of the algorithms, although the quality of images compressed at 20:1 with the wavelet algorithm was somewhat inferior. A lower diagnostic accuracy was noted using images compressed by the JPEG or wavelet algorithms at 20:1 in detecting lung cancers measuring 6 to 10 mm and cancers measuring from 6 to 15 mm with ground-glass opacity. CONCLUSION: Compression of low-dose CT images at a ratio of 10:1 using JPEG and wavelet algorithms does not compromise the detection rate of small lung cancers. PMID- 11259943 TI - Abscesses of the breast. US-guided serial percutaneous aspiration and local antibiotic therapy after unsuccessful systemic antibiotic therapy. AB - PURPOSE: The usual therapeutic approach to acute breast abscesses (ABAs) not responsive to systemic antibiotics is surgical incision and drainage. Our purpose was to assess the efficacy of treating ABAs with serial US-guided percutaneous aspiration and local injection of antibiotics. MATERIAL AND METHODS: Twenty-six patients with 28 ABAs, in whom systemic antibiotic therapy had failed, underwent serial US-guided aspiration with local injection a of large-spectrum antibiotic. The treatment was repeated weekly until complete resolution was observed at clinical and US examination. The volume of the ABAs was calculated before each US guided aspiration. Follow-up US examinations were performed at 1, 4, and 12 weeks after clinical and US resolution. A comparison betweeen costs of surgical and US guided treatment of ABAs in our institution was done. RESULTS: In 27 ABAs the treatment was successful: a progressive volume reduction and a complete resolution at clinical and US examination was observed within 1 to 7 weeks. In 1 case only, with a large ABA markedly increased in volume at the second examination, surgical drainage was performed. CONCLUSION: US-guided aspiration with local antibiotic injection is a safe and effective approach to ABAs, cheaper than surgical drainage of these lesions. PMID- 11259944 TI - Palliative treatment of inoperable malignant esophageal strictures with conically shaped covered self-expanding stents. AB - PURPOSE: To investigate the effectiveness of conically shaped covered self expanding (Flamingo) stents in palliative treatment of malignant esophagogastric strictures in terms of patency, improved dysphagia score and survival. MATERIAL AND METHODS: Flamingo stents were placed under fluoroscopic guidance between August 1998 and December 1999 for palliation of malignant dysphagia in 33 cases. There were 21 males and 12 females aged 40--80 years (average 64.2 years). RESULTS: Stent placement was successful in all patients, with good symptomatic control and no procedure-related complications. Spontaneous esophago-respiratory fistula and perforation accompanying malignant esophageal stricture in a total of 4 cases (12.2%) were successfully closed. In 1 case, tumor ingrowth was detected from the distal uncovered segment of the stent. In 2 cases with esophago respiratory fistula, gastrointestinal bleeding occurred. The cause of hemorrhage could not be found by angiography. The mean survival time in 17 patients, later deceased, was 129 days (range 9--360), and the mean observation time in 16 patients still alive is 180 days (range 18--365). CONCLUSION: Flamingo stents provide an effective and safe choice of palliative therapy in inoperable malignant esophagogastric strictures. PMID- 11259945 TI - Frequency of benign hepatic lesions incidentally detected with contrast-enhanced thin-section portal venous phase spiral CT. AB - PURPOSE: To evaluate the frequency of benign focal hepatic lesions incidentally detected at contrast-enhanced thin-section portal venous phase spiral CT. MATERIAL AND METHODS: Between January 1998 and February 1999, contrast-enhanced hepatic spiral CT examinations were performed in 1,892 patients. Out of these, only 100 patients fulfilled the following inclusion criteria: No underlying malignant disease, no liver cirrhosis, no suspected or known focal liver lesions. Standardized spiral CT parameters were applied. All CT studies were reviewed retrospectively by one radiologist. Any focal lesion was recorded and classified. Lesion size and number were noted. RESULTS: A total of 108 hepatic lesions were reported in 33 out of 100 patients (80 cysts; 18 hemangiomas; 3 focal fatty infiltrations; 2 focal non-tumorous perfusion defects; 1 calcification; and 4 non classified lesions). The average lesion size was 9.4 mm (< or =5 mm: n=40; 6-10 mm: n=30; 11-15 mm: n=28; >15 mm: n=10). CONCLUSION: Benign liver lesions are probably a frequent incidental finding at abdominal spiral CT. PMID- 11259946 TI - Self-expandable metal stents in the treatment of antro-pyloric and/or duodenal strictures. AB - PURPOSE: To assess the usefulness of self-expandable metal stents in the recanalization of antro-pyloric and/or duodenal strictures. MATERIAL AND METHODS: We report our experience of 15 patients with inoperable antro-pyloric and/or duodenal strictures treated by implantation of 21 self-expandable metal stents (18 uncovered and 3 covered) inserted perorally under fluoroscopic guidance. The patients were 11 men and 4 women, mean age 65.3 years. Fourteen of 15 patients were affected by a malignant stricture of the antro-pyloric region and/or duodenum either primary or secondary in 10 and 4 cases, respectively. Only in 1 case there was a benign stricture from postoperative scarring. Stricture length and diameter varied from 3 to 9 cm (mean 5.4 cm) and from 0 to 4 mm (mean 1.27 mm), respectively. RESULTS: Twenty-one stents were placed in 15 patients: Technical success was achieved in all cases while clinical improvement was obtained in 14 cases. No short-term complications were observed. A mean 4.3-month follow-up was obtained. Two patients had emesis secondary to peritoneal dissemination of the tumor after 1 and 2 months, respectively. Two other patients showed tumor overgrowth of the oral edge of the prosthesis after 3 and 2 months, respectively, and required another coaxial stent to bridge the new stenosis. The patient treated for a benign stricture had jaundice after 3 months and percutaneous internal-external biliary drainage was necessary. CONCLUSION: Self expandable metal stents are a safe and effective treatment of antro-pyloric and duodenal strictures; therefore, they should be considered an alternative to palliative resection in cases of advanced stage disease or poor general physical condition. PMID- 11259947 TI - MR imaging of upper abdomen following cholecystectomy. Normal and abnormal findings. AB - PURPOSE: To describe the normal MR appearance after cholecystectomy and the findings in patients with postoperative complications using fast pulse sequences in abdominal MR imaging. MATERIAL AND METHODS: In a prospective study of 119 patients, 64 were examined with MR after cholecystectomy. In total, 56 patients with uncomplicated cholecystectomy were examined with MR 1--5 days (mean 1.6 days) after cholecystectomy. Nine patients had an abdominal postoperative complication and 8 of these were examined with MR after the complication commenced 1--12 days after the cholecystectomy. RESULTS: Oedema in the gallbladder fossa was the only finding in 39 patients (61%), all with uneventful recovery. Small fluid collections in an area consistent with the gallbladder fossa were seen in 9/64 (14%) patients, of which 3 had surgical complications: 1 bleeding and 2 bile duct leakage. Twenty-two (34%) patients had small locally situated fluid collections adjacent to the liver, 14 were uneventful and 8 showed postoperative surgical complications. Seven patients had fluid in the rest of the abdomen of which 5 had surgical complications; 4 due to bile duct leakage and 1 acute pancreatitis. One patient had a postoperative bleeding not seen on MR images. CONCLUSION: MR is very sensitive in detecting fluid collections. Early MR findings following cholecystectomy are normally only subtle changes, mainly in the gallbladder fossa. Fluid collections diagnosed elsewhere than in the gallbladder fossa usually indicate a surgical complication and a surgical complication is unlikely if MR fails to show a fluid collection. PMID- 11259948 TI - The prevalence of disc aging and back pain after fusion extending into the lower lumbar spine. A matched MR study twenty-five years after surgery for adolescent idiopathic scoliosis. AB - PURPOSE: To determine the long-term outcome after fusion for adolescent idiopathic scoliosis in terms of degenerative disc findings diagnosed using MR imaging and to elucidate the clinical consequences. MATERIAL AND METHODS: Thirty two patients with adolescent idiopathic scoliosis, who had undergone spinal fusion using Harrington rods to the lower lumbar spine with one or two unfused discs below the fusion, were re-examined 25 years after the fusion. The re examinations included validated questionnaires, clinical examination, full standing frontal and lateral radiographs and MR examination of the lower lumbar region. Curve size and degenerative findings on MR images were evaluated by two unbiased radiologists, blinded to the clinical findings. A matched control group of 32 persons without scoliosis was subjected to the same examinations. RESULTS AND CONCLUSION: There were significantly more degenerative disc changes (p<0.0001), disc height reduction (p=0.0010) and end-plate changes (p<0.0001 for both upper and lower end-plates) in the lowest unfused disc in the patient group compared with the control group. The MR findings in the lowest unfused disc, but not the one above, in the patient group correlated to lumbar pain intensity as well as to the diminished lumbar lordosis. PMID- 11259949 TI - Early response of breast cancer bone metastases to chemotherapy evaluated with MR imaging. AB - PURPOSE: To compare T1-weighted spin-echo and fat-suppressed long echo time inversion recovery turbo spin-echo (long TE IR-TSE) MR images in the evaluation of early response of breast cancer bone metastases to chemotherapy. MATERIAL AND METHODS: Eighteen breast cancer patients with known bone metastases were investigated prospectively by MR, using T1-weighted and long TE IR-TSE sequences on the sternum, spine, pelvis and proximal femora, before and after a median of 6 courses of chemotherapy. Therapeutic response evaluation with MR was based on change in tumor size assessed quantitatively by measuring all focal metastases, and change in pattern and signal intensity (SI) of the metastases, assessed visually. Combined response evaluation based on clinical findings, conventional radiography, and scintigraphy was used as reference. RESULTS: Progressive disease (2 patients) and no change (4 patients) were assessed equally well on both MR sequences. Long TE IR-TSE demonstrated partial response with higher accuracy than T1-weighted images, 58% (7/12 patients) vs. 17% (2/12 patients). In patients without progression there was an SI increase in or around the metastases in 6 patients on T1-weighted images and in 7 patients on long TE IR-TSE images. CONCLUSION: The long TE IR-TSE sequence demonstrated early partial response of breast cancer bone metastases to chemotherapy more accurately than the T1 weighted sequence. PMID- 11259950 TI - Radiographic measurement of femorotibial rotation in weight-bearing. The influence of flexion and extension in the knee on the extensor mechanism and angles of the lower extremity in a healthy population. AB - PURPOSE: A system for examination and measurement of the weight-bearing knee was developed earlier on examination equipment used for QUESTOR Precision Radiography (QPR), adapted to computed radiography and to a picture archiving and communication system. In this study the same system was used to develop a method to define "patellar variables" including measurements of the rotation of the femur and the tibia and the patellar translation, between semiflexion and extension, as well as the Q-angle. The aim of this study was to evaluate the reproducibility of these patellar variables and also to establish "normal" reference values for both the patellar variables and the variables in the original QPR system. MATERIAL AND METHODS: For evaluation of the reproducibility 10 volunteers with healthy knees were examined twice. To obtain the reference values, 80 volunteers with healthy knees, 10 females and 10 males within each decade between 20 and 59 years of age, were examined. RESULTS AND CONCLUSION: The reproducibility of the rotation of the femur and the tibia and of the Q-angle was 2--3 degrees (SD) and of the patellar translation about 3 mm (SD). The result from the healthy volunteers will be a useful tool for evaluation of disorders in the knee joint. PMID- 11259951 TI - Femorotibial rotation and the Q-angle related to the dislocating patella. AB - PURPOSE: To compare the femorotibial rotation, the patellar translation, the hip knee-ankle angle and the Q-angle in patients with a dislocation of the patella with those of healthy volunteers. Further, the clinically measured Q-angle was compared to that measured by radiography. MATERIAL AND METHODS: A system for measurement of patellar variables was previously developed and applied to 80 healthy volunteers. In the present study, 28 patients (20 women, 8 men) with dislocation of the patella were examined bilaterally. Fourteen patients had habitual dislocations (20 affected knees) and 14 patients traumatic dislocations (17 affected knees). In 20 patients the clinical Q-angle was measured bilaterally by an orthopaedic surgeon and in 9 of these patients also by a second independent orthopaedic surgeon. RESULTS: The most striking finding was that dislocating knees in both groups showed a smaller Q-angle than the healthy knees. Further, the habitual group showed greater relative rotation between the tibia and the femur and an increased patellar translation compared to the traumatic group and to the healthy volunteers. There was a poor correlation between clinical and radiographic measurements of the Q-angle and no correlation was found between two independent clinical measurements. CONCLUSION: Surgical operations aiming at decreasing the Q-angle should be challenged. PMID- 11259952 TI - Enhanced visualisation of the normal prostate blood flow in young healthy volunteers using a new ultrasound contrast agent. AB - PURPOSE: To evaluate the sonographic appearance of normal prostate vascularity before and after injection of a new ultrasound contrast agent, Sonazoid (NC100100, Nycomed Amersham). MATERIAL AND METHODS: Five healthy male volunteers were given three injections of Sonazoid each. Transrectal B-mode, colour Doppler and colour Doppler energy (i.e. power Doppler) imaging was performed. The visibility of the vascular pattern and the vascular architecture of the prostate, including dynamics of contrast inflow and blood flow symmetry, were evaluated. RESULTS: The depiction of the vascularity was improved in all subjects after injection of Sonazoid for both Doppler modes. No improvement was seen for B-mode. Contrast dynamics within the prostate vessels were demonstrated with a mean time from injection of the ultrasound contrast agent to enhancement of the Doppler signals in the subcapsular arteries of 14+/-1 s (11--17 s), and the ultrasound contrast agent reached the central periurethral veins 4--7 s later. A symmetric, radial, spoke-like intraprostatic vascular pattern could be identified in all subjects using power Doppler imaging and ultrasound contrast agent. CONCLUSION: Sonazoid improved the detection of normal human prostate vascular anatomy for both colour and power Doppler imaging. Contrast dynamic studies revealed a radial spoke-like intraprostatic vascular pattern. This information might be useful in examination of patients with suspicion of prostate cancer, and needs to be further investigated. PMID- 11259953 TI - Doppler ultrasonography in testicular tumors presenting with acute scrotal pain. AB - PURPOSE: To determine whether Doppler ultrasonography (US) could be useful in the evaluation of testicular neoplasms presenting with acute scrotal pain. MATERIAL AND METHODS: We examined 18 patients with acute scrotal pain using Doppler US. Peak systolic and end-diastolic velocity measurements were performed at first admission and on completion of 1-week proper antibiotic treatment. Data were retrospectively reviewed and compared with findings of a control group of 17 age matched subjects. RESULTS: Of the 18 patients evaluated, 11 were found to have testicular tumors and 7 testicular inflammation. Peak systolic and end-diastolic velocities were significantly increased in patients with testicular tumor (p<0.002) and orchitis (p<0.01) versus normal controls. Follow-up examination on completion of 1-week antibiotic treatment demonstrated persistent high velocity values in patients with tumor and normalization of velocity values in patients with orchitis. CONCLUSION: Our results support the concept that follow-up Doppler US examination is of value in patients with acute scrotal pain submitted to antibiotic treatment since it may contribute to the differentiation between orchitis and testicular tumor. PMID- 11259954 TI - Uterine artery embolization of symptomatic uterine fibroida . Initial success and short-term results. AB - PURPOSE: To evaluate reduction in fibroid volume, the effect on clinical symptoms, adverse events and complications after percutaneous uterine artery embolization (UAE) as primary invasive treatment for symptomatic uterine fibroids. MATERIAL AND METHODS: Sixty-two patients entered the study. Indications for treatment were fibroid-induced menorrhagia, bulk symptoms, pain, and/or large fibroid size. The first 50 patients were evaluated by clinical examination and ultrasonography with measurement of fibroid volume before treament and 1, 6 and 12 months after UAE. The remaining 12 patients were followed 3 and 12 months after treatment. Embolization with microparticles was performed percutaneously in local analgesia by selective catheterization of both uterine arteries. RESULTS: A primary technical success with bilateral UAE was achieved in 60/62 (97%) of the patients. They were treated for postprocedural pain lasting up to 24 h. In 30 of the 62 patients with 6 months follow-up, the mean fibroid volume was reduced 68% 6 months after treatment. Twenty-nine (96%) of the patients experienced reduced bleeding, 21 (70%) reduced pain, and 18 (61%) reduced bulk symptoms at follow-up. CONCLUSION: UAE is a method with a high technical success rate. The treatment has good effect on fibroid volume reduction and clinical symptoms. Severe post procedural pain occurs generally in successful bilateral embolizations, but complications and adverse events are otherwise few and minor. UAE represents a promising new method for treating uterine fibroid-related symptoms. PMID- 11259955 TI - Visualization of tumor vessels in renal tumors. Comparison between power Doppler ultrasonography and angiography. AB - PURPOSE: To compare the ability of power Doppler ultrasonography (PDUS) with that of renal angiography for assessment of renal tumor vessels. MATERIAL AND METHODS: We performed PDUS and angiography in 52 histologically proven renal parenchymal tumors (50 renal cell carcinomas (RCCs) and 2 oncocytomas), and compared vascularity on PDUS and angiography. The vascularity of PDUS was graded as follows: grade 0-- no recognizable tumor vessel; grade 1-- hypovascular to surrounding renal interlobar arteries; grade 2-- hyper- or isovascular to surrounding renal interlobar arteries. RESULTS: With PDUS, 41 tumors were grade 2 and 11 were grade 1. With angiography, 44 lesions had iso/hypervascular pattern, 6 hypovascular pattern, and 2 were judged to be avascular. Among 44 iso/hypervascular tumors, 41 were grade 2, and 3 were grade 1. These latter 3 were located deeper than 7 cm. Six hypovascular tumors and 2 avascular tumors were grade 1. The 2 avascular tumors were small and hypovascular. The kappa-level of agreement was 0.81. CONCLUSION: There was very good agreement between PDUS and angiography in visualizing renal tumor vessels. PDUS appears appropriate for assessing renal tumor vascularity as compared to angiography in small and hypovascular lesions, but deep location reduced the detectability of tumor vessels with PDUS. PMID- 11259956 TI - Image quality in extended arc filtered digital tomosynthesis. AB - PURPOSE: To study image quality in filtered digital tomosynthesis (FDTS) tomograms as a function of their reconstruction arc, using isocentrically acquired, fluoroscopic projection data. MATERIAL AND METHODS: Both digital tomosynthesis (DTS) and cone beam CT (CBCT) reconstruction algorithms are based on backprojection and use cone beam projection data as input. Under limited angle conditions, CBCT is reduced to FDTS, where only a subset of projection data are used for reconstruction. The effect of the reconstruction arc on the spatial resolution, slice thickness, contrast sensitivity, shape distortion and artifacts, was also experimentally studied. The investigation was performed using both simulated and actual fluoroscopic images. RESULTS AND CONCLUSION: Image quality in terms of spatial resolution, slice thickness, shape distortion and artifacts, improved with increasing reconstruction arc and was optimized at 180 degrees, while contrast continued to improve as the arc was increased to 360 degrees. However, DTS was determined to be the technique of choice when reconstruction arcs of less than 40 degrees were used. Consequently, FDTS may be successfully implemented in applications involving extended arc reconstructions, in the range between 40 degrees delimiting the DTS domain and 360 degrees corresponding to CBCT. PMID- 11259957 TI - Self-reported heart health behaviour patterns in a rural context. AB - Does health behaviour change after people receive the results of screening tests? A cohort from rural central Queensland were contacted through telephone interview at 6-month intervals following heart risk screening. Participants self-reported their health behaviour and identified aspects of their lifestyles that supported or inhibited positive health behaviour. Qualitative analysis revealed the types of changes made and the contextual issues that affected health behaviour. The majority of participants did change their behaviour or maintained existing positive behaviours. Some participants were unsuccessful in making health changes. Health enablers included physical ability to undertake activities, appearance and support. The cost and availability of nutritious food in rural areas and the need for more socially supportive environments at a community level were issues raised by participants. Resources need to be made available so that mobile health services can augment local health initiatives, supporting positive health behaviour in rural areas. PMID- 11259959 TI - The impact of coordinated care: Eyre region, South Australia 1997-1999. AB - The SA HealthPlus Coordinated Care Trial in the Eyre Region began in fortuitous circumstances. First, it coincided with the completion of the Eyre Regional Health Service (ERHS) needs assessment in 1996, which highlighted outstanding health service needs and community concerns in relation to health care across the region. Second, although conceived as a formal trial, using standard research techniques, scientific processes and formal control groups to test significant differences between intervention and control groups, the trial did not conform strictly to the rules of social science or pure science and became more an exercise in action research. More significantly still, the Eyre Region became involved in the process, not so much as a way of proving a concept (the SA Health Plus hypotheses around utilisation, funding and health outcomes), but as a way of creating opportunities for change in the regional health system. If nothing else, the region stood to benefit from the implementation of the trial and involvement in the trial process. The present paper outlines the impact of the Eyre Coordinated Care Trial, not in terms of hypotheses and data analysis, but in terms of the impact of the trial processes on systems change and the evolution of an outcome-based health system. Such a system has the potential to deliver improved health outcomes to communities within existing financial resources and make much more effective use of resources by integrating care delivery and encouraging collaboration between health providers. In addition, the success of the change process in Eyre also supports the notion that change is not necessarily predicated upon scientific processes and research outcomes alone, but also upon the human and social structures associated with such endeavours. This perspective also contributes to the debate about the nature and role of science in the advancement of knowledge. PMID- 11259958 TI - Rural practice evaluation: how do rural physicians evaluate their working conditions? AB - Evaluating the quality of rural doctors' working conditions is essential for retaining physicians in rural areas. We carried out a trial to investigate those aspects of working conditions that are important to rural physicians and with which aspects they are satisfied or dissatisfied. Questionnaires were mailed to 204 doctors who were working in rural clinics in Japan. The professional conditions of rural clinic practices were classified into 17 items. The doctors were then asked to evaluate the importance of and degree of satisfaction with each item. Among the 17 items, the clinic's equipment, the municipal government's attitude and the base hospital were evaluated to be more important than the overall average. With regard to satisfaction, the distance to major cities, the municipal government's attitude and locum availability were rated significantly lower than that overall. There were some items where there was a discrepancy between the importance and the degree of satisfaction. Identifying these discrepancies may contribute to creating an environment that will raise the level of rural physicians' satisfaction. PMID- 11259960 TI - Effectiveness of changes in the delivery of diabetes care in a rural community. AB - Diabetes has a significant impact upon health in rural Maori communities. A diabetes club was established to support self-care and improve diabetes management in a rural community in Northland, New Zealand. A structured approach to care and an associated audit were also introduced. Patient involvement and ownership of the condition were considered important issues. Monitoring of care processes increased by 79%. The first year of audit was associated with a reduction in mean fructosamine from 369 +/- 85 micromol L-1 to 321 +/- 65 micromol L-1 and this was sustained for a further 3 years. The number of people using insulin increased from 15 to 22%. The audit process facilitated the implementation of changes in the delivery of care. We conclude that the data indicate that the enthusiastic delivery of care in general practice, with a devolution of power to the patient, linked to an audit service can result in improved management among patients with Type 2 diabetes. PMID- 11259961 TI - Amalgamation and collaboration in rural general practices: early experience with the GP links program in rural South Australia. AB - The GP Links program aims to promote the amalgamation of smaller general practices into larger group practices and is one of several strategies being used to modernise Australian family practice. GP Links provides financial incentives to practices willing to amalgamate. The focus of the program has been on urban practices to date and indeed some of the requirements of the program mean that rural practices are less likely to access the scheme. We report our positive and negative experiences of practice amalgamation through the GP Links program in a regional setting of South Australia. From our experience we suggest that for rural practices, a staged approach of increasing collaboration that may lead to amalgamation, which focuses on rural practices developing a supportive network and alliances with others such as Divisions and University Departments of Rural Health might be a positive way ahead. PMID- 11259962 TI - Work practices of rural and remote physiotherapists. AB - Physiotherapists in rural and remote areas face challenges in their service delivery. The challenge is to provide accessible and comprehensive services to rural and remote Australians. Research was undertaken regarding the work practices of physiotherapists and the availability of other health professionals in rural and remote South Australia and Northern Territory. Rural and remote area physiotherapists in this study are likely to be sole practitioners with a generalist practice. The high number who have practising rights at public hospitals gives rise to service delivery with a wide range of clients and reduced opportunity for specialisation. In addition, the demand for physiotherapists to be multiskilled is increased with the lower number of resident medical specialists and allied health professionals in rural areas. Greater access to other professionals for rural and remote Australians may come from the development of resource networks supported by regional organisation of resources and infrastructure. PMID- 11259963 TI - Improving access for rural Australians to treatment for anxiety and depression: The University of Melbourne Depression and Anxiety Research and Treatment Group Bendigo Health Care Group initiative. AB - Rural Australians have limited access to care for mental health problems. We describe a collaboration between the University of Melbourne Departments of Psychology and Psychiatry and a rural Area Mental Health Service to provide a specialist anxiety and depression treatment service in rural Victoria. The clinical service and the education and training approach are described. PMID- 11259965 TI - Reach for the phone: urgent imaging findings in obstetrics and gynaecology. AB - Some imaging findings indicate that a patient is at risk, and are too urgent or too important for the radiologist to rely on written communication alone. Some examples in obstetrics and gynaecology are demonstrated. PMID- 11259966 TI - An approach to demonstrating cost-effectiveness of diagnostic imaging modalities in Australia illustrated by positron emission tomography. AB - The aim of this study was to develop a framework in which the cost-effectiveness of new imaging technologies could be evaluated using data from other countries, while assessing the impact that any differences between the study populations and Australia may have upon the results. Publications reporting the cost effectiveness or therapeutic impact of positron emission tomography (PET) were re worked using Australian cost structures. PET was assigned a cost of $950. The effects of potential differences between the populations studied and the Australian population were evaluated by applying sensitivity analysis to those publications that describe decision tree methodology. The parameters included in the sensitivity analysis were disease prevalence and specificity of PET. The Australian cost savings per patient examined by PET were $505.50-$912.41 for investigation of solitary pulmonary nodules, $34.65-$360.03 for lung cancer staging, $550.08 for axillary staging of breast cancer, $230.75-$2301.27 for assessment of recurrent colorectal cancer and $300.24-$2069.65 for assessment of myocardial viability. Significant differences in disease prevalence and PET specificity could occur while the cost-effectiveness of PET was preserved. Decision tree sensitivity analysis can demonstrate the cost-effectiveness of diagnostic imaging modalities in Australia and provides indications that PET is cost-effective for a range of clinical indications. PMID- 11259967 TI - The inter-sonographer reliability of carotid duplex ultrasound. AB - Carotid duplex ultrasound (CDUS) is a non-invasive technique used to assess the severity of carotid artery stenosis. It has been shown to have good correlation with digital subtraction angiography (DSA) but has been criticised for its variability. One source of this is the variation in results between studies responsible for re-validating velocity criteria to match the established treatment thresholds of internal carotid artery (ICA) stenosis. The aim of this study was to develop velocity criteria and determine the presence of inter sonographer variation of CDUS when grading ICA stenosis in our department. Five sonographers measured the degree of ICA stenosis with CDUS in 33 patients who also underwent DSA. Receiver operator characteristic curve analysis was used to develop optimal velocity criteria for the 50%, 70% and 90% ICA stenosis thresholds as a group and for each individual sonographer. A peak systolic velocity ratio of > or = 3.25 was shown to have the highest accuracy (91.5%) for predicting a 70% stenosis. A moderate value of kappa (0.53 +/- 0.027) was calculated if the optimum velocity criterion was employed for each sonographer. There was no significant variation between the ability of sonographers to grade ICA stenosis (P > 0.05) and an excellent ICC of 0.911 was calculated. This study provides evidence to suggest that CDUS in our department is not an operator dependant test for the investigation of ICA stenosis. PMID- 11259968 TI - Invasive lobular carcinoma: sonographic features of cancers detected in a BreastScreen Program. AB - Sixty-two screen-detected invasive lobular carcinomas (ILC) were studied for sonographic, mammographic, clinical and histological findings. Ultrasound (US) features were compared with 60 invasive duct cancers (IDC). Size and axillary lymph node status in ILC were compared with all other cancers detected. In 41 ILC examined with US, 36 were found as masses (87.8% sensitivity; 95% CI 77.8-97.8%). Some US features of ILC and IDC differed: ILC were 9.94 times more likely to be hyperechoic (odds ratio, OR, 9.94; 95% CI 3.28-31.74) and 77% less likely to be taller than wide (OR 0.23; 95% CI 0.18-0.62). Thirty-three ILC showed typical malignant features of spiculate margins and acoustic shadowing. invasive lobular carcinomas had a greater mean diameter (20.4 mm; n = 60) than other invasive cancers (14.4 mm; n = 322) (P < 0.001). Ultrasound-guided needle biopsy was the method of diagnosis in 26 of 41 impalpable ILC (63%). Ultrasound has high sensitivity in characterizing screen-detected ILC, which may have atypical sonographic features including hyperechogenicity and a wider than tall shape. Ultrasound was an important contributor to diagnosis. PMID- 11259969 TI - Dual phase 99m-technetium Sestamibi imaging with single photon emission computed tomography in primary hyperparathyroidism: influence on surgery. AB - The purposes of this study were to determine the positive and negative predictive values of 99m Technetium (99mTc) Sestamibi dual phase imaging with single photon emission computed tomography (SPECT) for parathyroid adenomata or hyperplasia and the effect of preoperative localization on duration of surgery. We reviewed 33 adults (14 men, 19 women; mean age 53 years) with newly diagnosed primary hyperparathyroidism who underwent neck exploration. The duration of surgery for this cohort was compared with a group of historical controls (n = 53) who underwent surgery without preoperative SPECT. At surgery, there were 21 adenomata (including one carcinoma), 10 patients with hyperplasia and two with no pathology detected. The positive predictive values (PPV) for adenomata and hyperplasia were 95% and 100%, respectively. The negative predictive values (NPV) for these entities were 67% and 22%, respectively. The mean weight of adenomata detected was 3.4 g (range 0.2-17 g). Mean duration of surgery was 112.6 min as compared with 113 min in the historical controls (P = not significant). We conclude that 99mTc Sestamibi dual phase imaging with SPECT has an excellent PPV for parathyroid adenomata and hyperplasia, but does not contribute to reduced duration of surgery in patients undergoing neck exploration for the first time. The NPV is low, suggesting that a negative result does not exclude an adenoma or hyperplasia. PMID- 11259970 TI - Attitudes and treatment outcome of breast conservation therapy for stage I & II breast cancer using peroperative iridium-192 implant boost to the tumour bed. AB - Breast conservation therapy for early breast cancer is an established but grossly under-utilized treatment option in India for various reasons. Breast conservation therapy was offered to 200 suitable breast cancer patients between June 1993 and June 1998. Fifty-one patients (25%) opted for breast conservation and the remaining preferred mastectomy. In patients agreeing to conservation therapy, surgery was performed first along with peroperative implantation of iridium-192 to deliver a boost. Whole breast irradiation of 45 Gy was delivered 3-4 weeks after the boost. Cosmesis was assessed at the end of 6 months from completion of therapy. The main reason for refusal of breast conservation therapy was fear of recurrence in the remaining breast (60%). There were no locoregional failures in our study at a median follow up of 42 months; one patient experienced a systemic relapse. Cosmesis was good to excellent in 80% of patients. Breast conservation therapy using peroperative iridium-192 implant provides excellent locoregional disease control and cosmesis. The results of our study indicate that patient preference for mastectomy is an important reason for the under-utilization of breast conservation therapy in India. PMID- 11259971 TI - Strontium-89 treatment for prostate cancer bone metastases: does a prostate specific antigen response predict for improved survival? AB - A review of 50 patients treated with strontium-89 for prostate cancer bone metastases from January 1993-1997 at the Wellington Cancer Centre was undertaken to determine if there was any correlation between changes in prostate-specific antigen (PSA) following treatment and subsequent survival. Thirty cases were evaluable for PSA response. Of these, 14 had a fall in PSA following strontium-89 treatment, and their mean survival was 641 days. The remaining 16 patients did not demonstrate a post-treatment fall in PSA and their mean survival was 275 days. A difference between these two groups in the time to development of new bone symptoms following treatment was also observed. No significant correlation between pretreatment PSA and PSA response was observed. In conclusion, a PSA response following strontium-89 treatment appears to predict for improved survival in patients with bone metastases from carcinoma of the prostate. Further prospective studies are indicated. PMID- 11259972 TI - Australian high-dose-rate brachytherapy protocols for gynaecological malignancy. AB - There is no consensus over the optimal dose fractionation schedules for high-dose rate (HDR) brachytherapy used for gynaecological malignancy. In Australian public hospital departments of radiation oncology, HDR brachytherapy for gynaecological cancer is being more commonly used. A survey of public departments that are using this technology, or that plan to introduce this technology, was performed. Their current protocols are presented. In general, protocols are similar biologically; however, the practical aspects such as the number of fractions given do vary and may reflect resource restrictions or, alternatively, differences in interpretations of the literature and of the best protocols by clinicians. PMID- 11259973 TI - Leiomyoma of the oesophagus in an HIV-infected patient. AB - Leiomyomas of the oesophagus are uncommon, and have not been reported in patients infected with the human immunodeficiency virus (HIV). A case of an oesophageal leiomyoma in an adult infected with HIV is presented. PMID- 11259974 TI - Atypical magnetic resonance imaging findings of craniopharyngioma. AB - Three cases of craniopharyngiomas with atypical MRI findings are reported. The first patient had a nasopharyngeal craniopharyngioma. Its unusual location made diagnosis difficult. The second patient had a massive craniopharyngioma with extensive cystic expansion, involving the anterior, middle and posterior cranial fossae, and extending into the foramen magnum. The tumour of the third patient involved the suprasellar region with a large extension into the third ventricle, and demonstrated a predominantly high signal intensity on all T1-weighted, proton weighted and T2-weighted images. These patients further stressed the complexity of MRI findings in craniopharyngiomas. PMID- 11259975 TI - Two-tone pancreas on T1-weighted images: correlation with abnormalities on magnetic resonance pancreatography. AB - Two patients are presented in whom geographical alteration in signal between areas of normal and abnormal pancreatic tissue on T1-weighted magnetic resonance images of the pancreas was observed. This alteration in signal intensity produced a 'two-tone' pattern; magnetic resonance cholangiopancreatography (MRCP) and endoscopic retrograde cholangiopancreatography (ERCP) in both patients revealed altered pancreatic duct drainage. It is believed that the 'two-tone' pancreas effect on T1-weighted images of the abdomen, may indicate aberrant duct drainage and that MRCP is an ideal means of further evaluation. PMID- 11259976 TI - Small ureterocele-like Gartner's duct cyst associated with ipsilateral renal aplasia: a case report. AB - Gartner's duct cyst associated with ipsilateral renal aplasia is a rare anomaly and fewer than 40 cases have been reported in the literature. A case of Gartner's duct cyst presenting like an ureterocele on sonography, intravenous pyelography and CT are described. PMID- 11259977 TI - Emphysematous gastritis: case report and literature review. AB - Emphysematous gastritis is a rare form of gastritis that results from infection of the stomach wall by gas-forming organisms. Diagnosis of this commonly fatal condition rests on radiological demonstration of gas within the stomach wall. This can be observed on plain radiographs or CT scans of the abdomen. Only by prompt diagnosis and treatment can mortality be avoided. A new case of empysematous gastritis, diagnosed on CT scan by the demonstration of both intramural and portal venous gas, is presented and the literature is reviewed. PMID- 11259978 TI - Nodular fasciitis of the breast simulating breast cancer on imaging. AB - Nodular fasciitis is a rare benign soft tissue tumour of the breast that clinically and radiologically can mimic invasive duct carcinoma. The clinical, radiological and pathological findings of nodular fasciitis of the breast in a 38 year-old woman, who presented with a palpable lesion in the upper inner aspect of the left breast, are described. The tumour is characterized histologically by a stellate spindle cell tumour with a focal myxoid background containing scattered inflammatory cells and microhaemorrhages. Pathological assessment of the lesion is essential in making the diagnosis. PMID- 11259979 TI - Hypertrophic osteoarthropathy: an unusual manifestation in nasopharyngeal cancer. AB - A patient with nasopharyngeal carcinoma developed clubbing and hypertrophic osteoarthropathy 6 months before radiological detection of secondary deposits in the lung. Another patient with nasopharyngeal carcinoma developed digital clubbing and hypertrophic osteoarthropathy 6 months after the discovery of lung metastases. Development of a paraneoplastic syndrome in the form of hypertrophic osteoarthropathy and digital clubbing is very rare. This manifestation of nasopharyngeal cancers is presented, with a short review of its biology and pathogenesis. PMID- 11259980 TI - Radiological-pathological correlation: alveolar pattern. AB - This review summarizes important pathological lesions of the lung that typically present radiographically with an 'alveolar pattern'. For each entity, the latest findings as to its pathogenesis, aetiology and pathology are reviewed in the introductory remarks. We then present the typical radiological appearances alongside macroscopic and microscopic pathological photographs. It is hoped that the parallel presentation of radiological image with the pathology will enhance the understanding of the diverse range of diseases the aevolar pattern comprises. PMID- 11259981 TI - Hysterosalpingography. PMID- 11259982 TI - Journal impact factors: a 'bioequivalence' issue? AB - AIMS: Journal impact factors (IMFs) are used increasingly by institutions as performance indicators of the quality of 'individual research output'. Although the need for discretion when using the numbers has been emphasized, there has been little formal analysis of the issues. We therefore investigated citation profiles for three clinical pharmacology journals to assess the validity of using IMF as a measure of 'individual research'. METHODS: We compared the pattern of individual citations for random samples of 120 papers published in Clin Pharmacol Ther (CPT), Br J Clin Pharmacol (BJCP) and Eur J Clin Pharmacol (EJCP) in 1981, 1991, 1995 and 1996. Using an analogy between citation-time profiles of papers and concentration-time profiles of drugs, it was possible to define 'lag-time', Cmax, tmax, t(1/2) and AUC(t), and to investigate 'bioequivalence'. RESULTS: Citation distributions for individual publications were widely variable and skewed (skewness = 1.47, 2.16 and 1.37 for CPT, BJCP and EJCP, respectively). The 90% CI values for the IMF of a publication in each journal (i.e. 90% CI for an observation as opposed to 90% CI for the mean) were 0.24-16.94, 0.08-10.3 and 0.09-5.68. CONCLUSIONS: IMF does not represent the impact of an individual paper. Furthermore, if the comparison of journals is treated as a bioequivalence issue, the citation data should be log transformed prior to calculating IMF such that they represent the likelihood of citation for the median article. After such transformation, absolute differences between the IMF of clinical pharmacology journals become much smaller. PMID- 11259983 TI - Biomarkers for the effects of antipsychotic drugs in healthy volunteers. AB - Studies of novel antipsychotics in healthy volunteers are traditionally concerned with kinetics and tolerability, but useful information may also be obtained from biomarkers of clinical endpoints. A useful biomarker should meet the following requirements: a consistent response across studies and antipsychotics; a clear response of the biomarker to a therapeutic dose; a dose-response relationship; a plausible relationship between biomarker, pharmacology and pathogenesis. In the current review, all individual tests found in studies of neuroleptics in healthy volunteers since 1966 were progressively evaluated for compliance with these requirements. A MedLine search yielded 65 different studies, investigating the effects of 23 different neuroleptics on 101 different (variants of) neuropsychological tests, which could be clustered into seven neuropsychological domains. Subjective and objective measures of alertness, and of visual-visuomotor auditory and motor skills were most sensitive to antipsychotics, although over half of all the studies failed to show statistically significant differences from placebo. The most consistent effects were observed using prolactin response and saccadic eye movements, where 96% and 83% of all studies resp. showed statistically significant effects. The prolactin inducing dose equivalencies relative to haloperidol of 19 different antipsychotic agents correlated with the lowest recommended daily maintenance dose (r(2) = 0.52). This relationship could reflect the clinical practice of aiming for maximum tolerated levels, or it could represent a common basis behind prolactin release and antipsychotic activity (probably D2-receptor antagonism). The number of tests used in human psychopharmacology appears to be excessive. Future studies should look for the most specific and sensitive test within each of the domains that are most susceptible to neuroleptics. PMID- 11259984 TI - Involvement of multiple human cytochromes P450 in the liver microsomal metabolism of astemizole and a comparison with terfenadine. AB - AIMS: The aims of the present study were to investigate the metabolism of astemizole in human liver microsomes, to assess possible pharmacokinetic drug interactions with astemizole and to compare its metabolism with terfenadine, a typical H1 receptor antagonist known to be metabolized predominantly by CYP3A4. METHODS: Astemizole or terfenadine were incubated with human liver microsomes or recombinant cytochromes P450 in the absence or presence of chemical inhibitors and antibodies. RESULTS: Troleandomycin, a CYP3A4 inhibitor, markedly reduced the oxidation of terfenadine (26% of controls) in human liver microsomes, but showed only a marginal inhibition on the oxidation of astemizole (81% of controls). Three metabolites of astemizole were detected in a liver microsomal system, i.e. desmethylastemizole (DES-AST), 6-hydroxyastemizole (6OH-AST) and norastemizole (NOR-AST) at the ratio of 7.4 : 2.8 : 1. Experiments with recombinant P450s and antibodies indicate a negligible role for CYP3A4 on the main metabolic route of astemizole, i.e. formation of DES-AST, although CYP3A4 may mediate the relatively minor metabolic routes to 6OH-AST and NOR-AST. Recombinant CYP2D6 catalysed the formation of 6OH-AST and DES-AST. Studies with human liver microsomes, however, suggest a major role for a mono P450 in DES-AST formation. CONCLUSIONS: In contrast to terfenadine, a minor role for CYP3A4 and involvement of multiple P450 isozymes are suggested in the metabolism of astemizole. These differences in P450 isozymes involved in the metabolism of astemizole and terfenadine may associate with distinct pharmacokinetic influences observed with coadministration of drugs metabolized by CYP3A4. PMID- 11259985 TI - Disposition and metabolism of the flavonoid chrysin in normal volunteers. AB - AIMS: To describe the oral disposition of the dietary flavonoid chrysin in healthy volunteers. METHODS: Oral 400 mg doses of chrysin were administered to seven subjects. Chrysin and metabolites were assayed in plasma, urine and faeces by h.p.l.c. RESULTS: Peak plasma chrysin concentrations were only 3-16 ng ml(-1) with AUCs of 5-193 ng ml(-1) h. Plasma chrysin sulphate concentrations were 30 fold higher (AUC 450-4220 ng ml(-1) h). In urine, chrysin and chrysin glucuronide accounted for 0.2-3.1 mg and 2-26 mg, respectively. Most of the dose appeared in faeces as chrysin. Parallel experiments in rats showed high bile concentrations of chrysin conjugates. CONCLUSIONS: These findings, together with previous data using Caco-2 cells, suggest that chrysin has low oral bioavailability, mainly due to extensive metabolism and efflux of metabolites back into the intestine for hydrolysis and faecal elimination. PMID- 11259986 TI - The effects of lacidipine on the steady/state plasma concentrations of simvastatin in healthy subjects. AB - AIMS: Lacidipine, a long acting 2, 4-dihydropyridine calcium channel antagonist is frequently administered with cholesterol lowering agents, particularly in elderly populations. The effects of lacidipine on the pharmacokinetics of simvastatin were investigated, since they share the CYP3A4 pathway for metabolism. METHODS: The study was an open, randomised, two-way crossover design, with at least 7 days washout. Eighteen healthy subjects received simvastatin, 40 mg once daily, alone and together with lacidipine, 4 mg once daily, for 8 days. The pharmacokinetics of simvastatin were studied on the eighth day. Analysis was made of total simvastatin acid concentrations (naive simvastatin acid plus that derived from alkaline hydrolysis of the lactone). RESULTS: Lacidipine increased the maximum concentration of simvastatin (Cmax) by approximately 70% (P=0.016) and the area under the plasma concentration-time curve AUC(0,24 h) by approximately 35% (P=0.001). The mean Cmax and AUC(0,24 h) of simvastatin (95% confidence interval) when given alone were 8.76 (6.72-11.41) ng ml(-1) and 60.36 (47.15-77.28) ng ml(-1) h. During treatment with lacidipine they were, respectively, 14.89 (10.77-20.58) ng ml(-1) and 80.96 (64.62-101.44) ng ml(-1) h. No significant differences were observed in either time to peak concentration (tmax was 1.0 h for simvastatin alone and 1.5 h for the combination) or in the half-life (t1/2,z was 8.5 h in both cases). The combination was safe and well tolerated. CONCLUSIONS: The observed increased exposure to simvastatin 40 mg following coadministration of lacidipine is unlikely to be of clinical relevance. PMID- 11259988 TI - Nasal retention of budesonide and fluticasone in man: formation of airway mucosal budesonide-esters in vivo. AB - AIMS: The efficacy of topical glucocorticosteroids in rhinitis and asthma is likely to depend on drug retention in the airway mucosa. With fluticasone propionate, retention may be achieved exclusively by lipophilicity, whereas for budesonide an additional possibility may be provided by its ability to form fatty acid esters in the airway mucosa that release the active drug. The aim of the present study was to determine the nasal mucosal retention of budesonide and fluticasone propionate, and the occurrence of budesonide-esters (budesonide oleate, budesonide-palmitate) in the nasal mucosa. METHODS: In the present study, involving 24 healthy subjects, we have examined nasal mucosal drug retention of single doses of topical budesonide (256 microg) and fluticasone propionate (200 microg). Treatments were given consecutively and the administration sequence was randomised. Subjects were randomised into four parallel groups and two nasal biopsies were taken from each subject, i.e. before and at 2 h, at 2 and 6 h, at 6 and 24 h, or before and at 24 h after drug administration, resulting in 12 biopsies/time point. The measurement of unesterified budesonide, budesonide oleate, budesonide-palmitate, and fluticasone propionate was based on microwave extraction procedures combined with liquid-chromatography/tandem mass spectrometry. RESULTS: Neither of the analytes was detected in samples taken before glucocorticosteroid administration. After administration, unesterified budesonide, budesonide-esters, and fluticasone propionate were detected in the tissue from 23, 20, and 19 subjects, respectively. The mean tissue levels of budesonide at 2 and 6 h were 1051 and 176 pmol g(-1); the mean levels of fluticasone propionate at these time points were 237 and 10 pmol g(-1). The dose corrected budesonide/fluticasone propionate tissue concentration ratios were 3.5 (P = 0.07) and 13.7 (P < 0.0002), respectively. At 24 h, budesonide and fluticasone propionate were detected in 8/12 and 3/12 of the biopsies, respectively. CONCLUSIONS: The present study demonstrates the formation of budesonide-esters in the human nasal mucosa in vivo, and that budesonide is retained in the nasal mucosa to a greater extent than fluticasone propionate. It is suggested that the formation of budesonide-esters and their subsequent release of budesonide contributes to an extended retention of budesonide in the airway mucosa. PMID- 11259987 TI - Effect of diuretics on fetal growth: A drug effect or confounding by indication? Pooled Danish and Scottish cohort data. AB - AIMS: The diabetogenic effect of diuretics, as well as the indication for prescribing them, may impact on fetal growth. We analysed whether the purchase of prescription drugs for diuretics during pregnancy was associated with measures of fetal growth. METHODS: During 1991-98 all women who purchased prescription drugs for diuretics during pregnancy were identified in the Northern Jutland Prescription Database (NJDP), Denmark, and in the Medicines Monitoring Unit's Database (MEMO), Scotland. Information on birth weight and gestational age was obtained from the Danish Birth Registry, the Danish Hospital Discharge Registry and the Scottish Tayside Neonatal Database. Information on diabetes, hypertension and prepregnancy weight were obtained by hospital record review in a sample of women in the Danish cohort. Women who did not purchase prescription diuretics during pregnancy were used as a reference group in both cohorts. RESULTS: Danish women who purchased prescription loop diuretics during pregnancy gave birth to infants with higher birth weights than women who did not use diuretics; mean difference 104.7 g (95% CI; 2.6, 206.9). However, the high prevalence of diabetes (10.3%) among Danish women who purchased prescription loop diuretics during pregnancy might explain this result. Both the Danish and the Scottish women who purchased prescription diuretics during their pregnancy were at increased risk of preterm delivery (< 37 completed weeks); ORs: 1.8 (CI; 1.2, 2.7)NJDP, 1.9 (CI; 0.9, 4.3)MEMO. The proportion of hypertension among women who purchased prescription thiazides was 15.8%, and the risk of having an infant with a birth weight (BW) < 2500 g was increased; ORs: 2.6 (CI; 1.4, 5.0)NJDP, 2.4 (CI; 0.8, 7.8)MEMO. CONCLUSIONS: Prescribing diuretics during pregnancy was associated with differences in birth weight and incidence of preterm delivery. Confounding by indication may explain the findings. PMID- 11259989 TI - Pharmacokinetics of sabeluzole and dextromethorphan oxidation capacity in patients with severe hepatic dysfunction and healthy volunteers. AB - AIMS: The primary objective of this study was to determine how the pharmacokinetics of sabeluzole, an investigational drug with specific effects on memory and learning abilities, are affected by chronic liver disease. Since sabeluzole is metabolised by CYP2D6, a secondary objective was to study the correlation between CYP2D6 activity (as assessed by the dextromethorphan dextrorphan metabolic ratio) and hepatic dysfunction. METHODS: The single-dose pharmacokinetics of sabeluzole (10 mg) was compared in 10 healthy Caucasian subjects and 10 patients with severe hepatic dysfunction. The urinary dextromethorphan/dextrorphan (DMP/DRP) metabolic ratio was determined after intake of 20 mg dextromethorphan (NODEX capsules). RESULTS: The terminal half life of sabeluzole was significantly prolonged in subjects with severe hepatic dysfunction vs healthy subjects (respectively 39.3 +/- 11.5 h; 17.5 +/- 10.2 h (mean +/- s.d.)). The areas under the curve (AUC) were significantly higher in subjects with severe hepatic dysfunction than in healthy volunteers (681 +/- 200 ng ml(-1) h vs 331 +/- 282 ng ml(-1) h). There was a significant correlation between the AUC(0,infinity) and the DMP/DRP metabolic ratio in healthy volunteers and subjects with severe hepatic dysfunction. AUC was greater and elimination of sabeluzole slower in poor metabolizers compared with extensive metabolizers. CONCLUSIONS: These results suggest that a) sabeluzole dose should be reduced in patients with severe hepatic dysfunction and b) the AUC of sabeluzole is linked to individual CYP2D6 activity. PMID- 11259990 TI - Influence of gender and oral contraceptives on CYP2D6 and CYP2C19 activity in healthy volunteers. AB - AIMS: The study was carried out in order to assess the effects of gender and the use of oral contraceptives (OCs) on CYP2D6 and CYP2C19 activities in healthy volunteers. METHODS: Six hundred and eleven Caucasian volunteers (330 males and 281 females; age range 18-49 years) were phenotyped with respect to CYP2D6 and CYP2C19 by means of the probe drugs dextromethorphan and mephenytoin, respectively. Extensive metabolisers were selected for this study. RESULTS: The median dextromethorphan/dextrorphan metabolic ratio in non-OC using females was significantly lower than in males (0.067 vs 0.080; P = 0.033) (mean difference in ln dextromethorphan/dextrorphan metabolic ratio 0.023, 95% CI 0.03-0.43). For the mephenytoin S/R ratio, no such difference was observed. However, OC using females had a significantly higher median mephenytoin S/R ratio than non-OC using females (0.230 vs 0.090; P < 0.001) (mean difference in ln mephenytoin S/R ratio 0.082, 95% CI 0.60-1.04). Moreover, females using combined OCs had a significantly higher median ratio than females using OCs with progestins only (median 0.258 vs 0.135; P = 0.008) (mean difference in ln mephenytoin S/R ratio 0.82, 95% CI 0.21 1.34). CONCLUSIONS: Given certain assumptions, the study indicates that females in the fertile age have a slightly higher CYP2D6 activity compared with males. There was no evidence of a gender difference in CYP2C19 activity. The use of combined OCs reduces the activity of CYP2C19, an effect that seems to be related to the ethinyloestradiol component. PMID- 11259991 TI - Acetylsalicylic acid and other salicylates in relation to Stevens-Johnson syndrome and toxic epidermal necrolysis. AB - AIMS: Various nonsteroidal anti-inflammatory drugs are known to increase the risk of Stevens-Johnson syndrome and toxic epidermal necrolysis. The relationship between salicylate treatment and these conditions is not known. METHODS: A case control study was conducted in four countries in Europe from 1989 to 1995. RESULTS: Among 373 cases and 1720 controls, the multivariate relative risk estimate for any salicylate use in the previous week was 1.3 (95% confidence interval, 0.8-2.2); no statistically significant elevations were observed for single ingredient preparations or for salicylate-containing combination products. CONCLUSIONS: Acetylsalicylic acid and other salicylates are not associated with a measurable increase in the risk of these rare but severe reactions. PMID- 11259992 TI - Population pharmacokinetics of antifibroblast activation protein monoclonal antibody F19 in cancer patients. AB - AIMS: The population pharmacokinetics of 131I-mAbF19, a radiolabelled murine monoclonal antibody against fibroblast activation protein and a potential antitumour stroma agent, were investigated during two phase I studies in cancer patients. METHODS: 131I-mAbF19 serum concentration-time data were obtained in 16 patients from two studies involving imaging and dosimetry in colorectal carcinoma and soft tissue sarcoma. Doses of 0.2, 1 and 2 mg antibody were administered as 60 min intravenous infusions. The data were analysed by nonlinear mixed effect modelling. RESULTS: The data were described by a two-compartment model. Population mean values were 109 ml h(-1) for total serum clearance, 3.1 l for the volume of distribution of the central compartment, and 4.9 l for the volume of distribution at steady state. Mean terminal half-life was 38 h. Intersubject variability was high, but no patient covariates could be identified that further explained this variability. In particular, there was no influence of tumour type or mAbF19 dose. CONCLUSIONS: The pharmacokinetics of antistromal mAbF19 were well defined in these two studies with different solid tumour types, and were comparable with those of other murine monoclonal antibodies that do not bind to normal tissue antigens or blood cells. PMID- 11259993 TI - Prescribing patterns in patients using new antidepressants. AB - AIMS: To study possible selective prescribing ('channelling') we compared characteristics of patients using the SSRI sertraline with patients using longer available SSRIs. METHODS: An observational cohort study in 1251 patients being prescribed an SSRI. RESULTS: In contrast to other studies, we found no evidence for channeling of sertraline. Sertraline was mainly prescribed for the labelled indication (depressive disorder), while older SSRIs were more often prescribed also for other indications. Time on the market was inversely associated to the proportion of patients treated for depressive disorder. CONCLUSIONS: We found no evidence for channeling of sertraline compared with prescribing patterns of older SSRIs. PMID- 11259994 TI - Prescribers prefer people: The sources of information used by doctors for prescribing suggest that the medium is more important than the message. AB - AIMS: The sources of prescribing information are legion but there is little knowledge about which are actually used in practice by doctors when prescribing. The aims of this study were to determine the sources of prescribing information considered important by doctors, establish which were used in practice, and investigate if hospital and primary care physicians differed in their use of the sources. METHODS: Two hundred general practitioners (GPs) and 230 hospital doctors were asked to rate information sources in terms of their importance for prescribing 'old' and 'new' drugs, and then to name the source from which information about the last new drug prescribed was actually derived. RESULTS: Among 108 GPs, the Drugs and Therapeutics Bulletin and medical journal articles were most frequently rated as important for information on both old and new drugs while pharmaceutical representatives and hospital/consultant recommendations were more important for information on new drugs, as opposed to old. In practice, information on the last new drug prescribed was derived from pharmaceutical representatives in 42% of cases and hospital/consultant recommendations in 36%, with other sources used infrequently. Among 118 hospital doctors, the British National Formulary (BNF) and senior colleagues were of greatest theoretical importance. In practice, information on the last new drug prescribed was derived from a broad range of sources: colleagues, 29%; pharmaceutical representatives, 18%; hospital clinical meetings, 15%; journal articles, 13%; lectures, 10%. GPs and hospital doctors differed significantly in their use of pharmaceutical representatives (42% vs 18%) and colleagues (7% vs 29%) as sources of prescribing information (P < 0.0001 for both). CONCLUSIONS: The sources most frequently rated important in theory were not those most used in practice, especially among GPs. Both groups under-estimated the importance of pharmaceutical representatives. Most importantly, the sources of greatest practical importance were those involving the transfer of information through the medium of personal contact. PMID- 11259995 TI - The pharmacokinetics of etanercept in patients with heart failure. PMID- 11259996 TI - Sun protection with clothing. PMID- 11259997 TI - Is subacute cutaneous lupus erythematosus a subset of systemic lupus erythematosus or a distinct entity? PMID- 11259998 TI - Carbon dioxide laser treatment promotes repair of the three-dimensional network of elastic fibres in rat skin. AB - BACKGROUND: We have previously reported that ultraviolet (UV) B irradiation induces a loss of linearity in the three-dimensional structure of dermal elastic fibres, which results in the reduction of elastic properties of the skin and leads to wrinkle formation. We further reported that repair of wrinkles by all trans retinoic acid is accompanied by recovery of the linearity of elastic fibres. Carbon dioxide (CO2) lasers are widely used for treating wrinkles in cosmetic surgery. OBJECTIVES: To perform CO2 laser treatment of wrinkles induced in rat skin by UVB irradiation and to evaluate changes in the three-dimensional structure of dermal elastic fibres during wrinkle repair. METHODS: Wrinkles were induced in the hind limb skin of Sprague-Dawley rats by UVB irradiation (130 mJ cm-2 three times weekly for 6 weeks), followed by CO2 laser treatment (11.3 J cm 2). The surface appearance of the skin was evaluated by replica observation 6 and 10 weeks after CO2 laser treatment followed by measurement of mechanical properties using a Cutometer. Subsequently, perfusion fixation and digestion with formic acid were performed and elastic fibres were observed by scanning electron microscopy (SEM). Image analysis of SEM micrographs was carried out to evaluate the linearity in the three-dimensional structure of elastic fibres. RESULTS: Six weeks after CO2 laser treatment, all parameters of skin mechanical properties in the UVB-irradiated group recovered to levels of the control non-irradiated group, accompanied by repair of wrinkles and a significant increase in linearity of the three-dimensional structure of elastic fibres. CONCLUSIONS: These findings indicate that CO2 laser treatment has a therapeutic potential to repair wrinkles to non-irradiated levels through recovery of the three-dimensional structure of elastic fibres. PMID- 11259999 TI - Herpes simplex virus-specific immune responses in subjects with frequent and infrequent orofacial recrudescences. AB - BACKGROUND: Following the primary infection of the orofacial region with herpes simplex virus (HSV) type 1, the virus remains latent in the ganglia for the lifetime of the host but can reactivate at intervals and cause recrudescent lesions. The frequency of these episodes varies considerably from one individual to another. OBJECTIVES: To compare immune responses in two groups of subjects: those with frequent orofacial lesions, defined as 10 or more per year (n = 12), and those with infrequent lesions, defined as three or fewer per year (n = 20). METHODS: Plasma and peripheral blood mononuclear cells were collected from each individual for the series of immunological tests listed in the following results section. RESULTS: Although IgG titres specific for HSV, measured by enzyme-linked immunosorbent assay (ELISA), were not different between the two groups, there was a significantly higher HSV-specific IgE titre in the frequent group. The percentages of CD3-, CD4- and CD8-positive cells in peripheral blood, assessed by flow cytometry, and the in vitro lymphoproliferative response to the non-specific mitogen concanavalin A, did not differ between the two groups. T-cell responses to HSV were assessed by in vitro lymphoproliferation with tritiated thymidine incorporation and subsequent calculation of the stimulation index; cytokine production [interferon (IFN)-gamma and interleukin (IL)-10] into the culture supernatant as a result of the stimulation was measured by ELISAs. The mean +/- SEM stimulation index was 4.1 +/- 0.2 in the subjects with frequent lesions and 11.8 +/- 3.1 in the subjects with infrequent lesions, a difference that was significant. The mean IL-10 concentrations found in the subjects with frequent and infrequent lesions were 154 and 110 pg mL-1, respectively, a difference that did not reach significance. However, the IFN-gamma production was significantly lower in the subjects with frequent lesions compared with those with infrequent lesions: mean 835 and 1679 pg mL-1, respectively. CONCLUSIONS: Thus, from the HSV specific T-cell proliferation, IFN-gamma production and IgE results, patients who experience frequent recrudescences may tend towards the production of T-helper 2 cytokines in response to the virus, which may lead, in turn, to less effective control of viral replication in the periphery following reactivation from latency. PMID- 11260000 TI - Melanoma cell-derived factors stimulate glycosaminoglycan synthesis by fibroblasts cultured as monolayers and within contracted collagen lattices. AB - BACKGROUND: Various tumours exhibit glycosaminoglycan rich, and in particular hyaluronan rich matrices surrounding them that facilitate tumour growth and invasion. In many tumours, this matrix is predominantly synthesized by fibroblasts following stimulation by tumour cell-derived factors. OBJECTIVES: To determine what effect tumour cell-conditioned medium has upon fibroblast glycosaminoglycan synthesis when cells were cultured as monolayers and within contracted collagen lattices. METHODS: Serum-free conditioned medium from melanoma cell lines (C8161, MV3, A375 and Hs294T) was examined for its ability to stimulate the incorporation of 3H-glucosamine and 35SO4 into glycosaminoglycans synthesized by fibroblasts. RESULTS: Conditioned medium from all four melanoma cell lines exhibited potent glycosaminoglycan-stimulating activity. In monolayer culture, C8161-conditioned medium stimulated a 4.2-fold increase in fibroblast hyaluronan, and a 9.9-fold increase in sulphated glycosaminoglycan synthesis, while 35SO4 incorporation was increased only 2.1-fold. In collagen lattice cultures, C8161-conditioned medium stimulated a 4.9-fold increase in hyaluronan synthesis, a 5.4-fold increase in sulphated glycosaminoglycans, and a 1.3-fold increase in 35SO4 incorporation. CONCLUSIONS: Melanoma cells produce factors that are potent stimulators of fibroblast glycosaminoglycan synthesis, in both monolayer culture and within contracted collagen lattices. Synthesis of both hyaluronan and sulphated glycosaminoglycans with a reduced degree of polymer sulphation is stimulated. Such changes are likely to promote tumour cell proliferation and migration. PMID- 11260002 TI - Clinical correlates of Breslow thickness of malignant melanoma. AB - BACKGROUND: Breslow thickness is a major predictor of prognosis in cutaneous malignant melanoma (MM) and patients continue to present with thick lesions, which have a poorer prognosis. OBJECTIVES: To investigate correlations of Breslow thickness with demographic variables, tumour site, clinical features, false negative diagnostic rate and clinic of primary referral. METHODS: Data were obtained from the records of 738 patients with histologically diagnosed cutaneous MM. Tumours included were in situ and invasive MM and lentigo maligna melanoma. In view of the skewed distribution of MM thickness, categories of MM thickness were used for analysis, using the commonly applied cut-offs of 0.75, 1.5 and 3.5 mm. The variables investigated were particularly associated with changes in the proportion of the thickest group, 'thick' MMs. The proportion of this thickness category is proposed as an appropriate and sensitive variable for the investigation of correlations with MM thickness. Thickness >/= 1.5 mm has also been considered in view of its prognostic significance. RESULTS: Results were similar for the two thickness groups, but more significant for the thick group. The previously described correlations of tumour thickness and increasing age (P < 0.00001) and location on head and neck (P = 0.0002), together with the independence of these variables, have been confirmed. The correlation with male gender was also confirmed but this was weak (P = 0.05). Novel findings were correlations of Breslow thickness with all features of the seven-point checklist (P varying from P = 0.01 to P < 0.00001) and tumour elevation (P < 0.00001). In general in the seven-point checklist, the absence of the major features (except variation in size) (P < 0.00001) and presence of minor features (except altered sensation) (P varying from P = 0.004 to P < 0.00001) were strongly associated with thick MMs. These results for tumour thickness are a further argument for retention of the minor features of the seven-point checklist. False negative diagnosis was found to be correlated with tumour thickness (P < 0.02) but this relationship was lost on multivariate analysis with inclusion of the clinical features. MM thickness was highly correlated with primary referral clinic (P < 0.00001). Only approximately 8% of MMs presenting to the Pigmented Lesion Clinic (PLC) were thick, while the proportion presenting to general dermatology was about three times greater and to plastic surgery about five times greater. This was maintained on multivariate analysis, including all other variables and, therefore, there must be other determining factors of referral not examined in the study. Conclusion MM thickness is associated with increasing age, male gender, location on the head and neck, all features of the seven-point checklist, tumour elevation and referral to the PLC. It is important to investigate further the reasons for general practitioner referral of different thickness MM to different types of clinic, as referral to clinics other than the PLC results in delay in the first hospital appointment, and it is now apparent that thicker lesions are disproportionately affected. PMID- 11260001 TI - Timing of excessive ultraviolet radiation and melanoma: epidemiology does not support the existence of a critical period of high susceptibility to solar ultraviolet radiation- induced melanoma. AB - BACKGROUND: The existence of a 'critical period' early in life characterized by a high susceptibility to melanoma initiation due to excessive ultraviolet (UV) radiation has been suggested by various authors based on epidemiological findings from migration studies and some case-control studies. However, the evidence so far is controversial as several epidemiological investigations failed to corroborate these results. Objective To compare the increase in melanoma risk due to excessive UV radiation between different periods in life. METHODS: In a multicentre case-control study we recruited 603 melanoma cases and 627 population controls in seven European countries. We obtained data on the history of sunburns during 'childhood' ( 15 years), respectively, in standardized personal interviews. We employed logistic regression analyses to estimate the impact of the exposure factors under study, while simultaneously controlling for the effect of a variety of confounding variables. RESULTS: We found a very similar upward gradient of melanoma risk in exposure categories related to the frequency of sunburns during both periods in life. More than five sunburns doubled the melanoma risk, irrespective of their timing in life. CONCLUSIONS: Our data do not provide supporting evidence for the existence of a 'critical period'. The hazardous impact of sunburns seems to persist lifelong and thus activities concerned with melanoma prevention should be directed to the entire population rather than being focused only on younger age groups. PMID- 11260003 TI - Ultraviolet protection by summer textiles. Ultraviolet transmission measurements verified by determination of the minimal erythema dose with solar-simulated radiation. AB - BACKGROUND: Apart from sunscreen lotions, clothing provides protection from acute and chronic sun damage. Therefore, it is very important to know the ultraviolet (UV) protection factor (UPF) of textiles, in particular of lightweight summer clothing. Usually, the UPF of a textile is determined by spectrophotometric assessment of the UV transmission (in vitro method). OBJECTIVES: To compare the relationship between in vitro tests and in vivo tests of UPF using solar simulators for determination of the minimal erythema dose (MED), applied to 30 different summer textiles. METHODS: Thirty summer textiles were spectrophotometrically assessed, and UPFs were calculated with respect to the International Commission on Illumination (CIE) erythemal action spectrum.1 Based on the in vitro UPFs 'on skin' and 'off skin', in vivo testing was performed using a solar simulator for the determination of the MEDunprotected and MEDprotected. RESULTS: The UPFs obtained from in vivo 'on skin' testing were significantly (r = 0.95; P < 0.001) lower than the predicted in vitro UPFs. This disparity was also confirmed by chromometric assessment of the MED testing; the erythemal responses measured after textile protection were significantly (P < 0.001) higher than those obtained without protection. However, the in vivo 'off skin' UPFs did not significantly (r = 0.98; P > 0.05) differ from the in vitro UPFs; comparison of the chromometrically assessed erythemal responses was also insignificant (P > 0.05). CONCLUSIONS: The different correlation between in vitro and in vivo measurements of the UPF may be due to the optical-geometrical properties of textiles and the different amount of direct and diffuse radiation passing through the spaces between the yarns. As spectrophotometric measurements of a textile may generally yield lower UPFs than those obtained under average field conditions, the in vitro test method provides 'safe' UPF values representing a 'worst-case scenario'. In contrast to in vitro testing, in vivo methods are much more expensive and time-consuming. Thus, with respect to practicality, spectrophotometric measurements seem to be most suitable for the evaluation of UV protection of textiles. PMID- 11260004 TI - Concentration-dependent phototoxicity in trimethylpsoralen bath psoralen ultraviolet A. AB - BACKGROUND: Long-term use of topical trimethylpsoralen (TMP) psoralen bath plus ultraviolet A (bath PUVA) is considered safe with regard to the risk of skin cancer. However, the potential for severe phototoxicity limits its use. OBJECTIVES: To study the effect of dilution of the TMP bath on the minimal phototoxic dose (MPD). METHODS: Fifteen volunteers participated in the study. The MPD tests were performed for three TMP concentrations: 0.33 mg L-1, 0.1 mg L-1 and 0.033 mg L-1 at 2-week intervals. Geometric UVA dose series increasing by a factor of radical2 were used for the testing on the previously unexposed buttock skin. The MPD72 h was assessed at 72 h from the bath. RESULTS: For the highest TMP concentration of 0.33 mg L-1, the median MPD72 h was 0.14 J cm-2 (95% confidence interval (CI), 0.10-0.14 J cm-2). For the diluted TMP bath concentration of 0.1 mg L-1, the median MPD72 h increased to 0.29 J cm-2 (95% CI, 0.2-0.41 J cm-2) and for 0.033 mg L-1 to 0.81 J cm-2 (95% CI, 0.57-1.15 J cm-2), respectively. Thus, diluting the labelled concentration of 0.33 mg L-1 1 : 10 increased the median MPD72 h 5.6-fold. CONCLUSIONS: With regard to the safety and practicality of the TMP bath PUVA, the lower concentrations of TMP may be of clinical importance, and this needs to be validated in future controlled clinical trials. PMID- 11260005 TI - Calcitriol 3 microg g-1 ointment in combination with ultraviolet B phototherapy for the treatment of plaque psoriasis: results of a comparative study. AB - BACKGROUND: Combinations of topical treatments and ultraviolet (UV) B phototherapy for plaque psoriasis may be more beneficial than either type of treatment used alone. OBJECTIVES: To determine the efficacy of calcitriol 3 microg g-1 ointment in combination with UVB phototherapy in treating plaque psoriasis. METHODS: Calcitriol ointment with UVB was compared with vehicle plus UVB in a randomized, double-blind study in 104 patients. RESULTS: Mean global improvement scores for both groups increased over the 8-week study period; there was a statistically significant difference (P < 0.05) in favour of the calcitriol/UVB combination from week 1. At end-point, 45% of the calcitriol/UVB group showed considerable improvement or clearing of psoriasis, compared with 21% of the control group. The superiority of calcitriol plus UVB was also reflected in the global severity and Psoriasis Area and Severity Index (PASI) scores; at end-point the mean percentage decrease in PASI score was 65% for the calcitriol/UVB group and 43% for vehicle/UVB (P = 0.0014). The incidence of skin related adverse events was low (< 12%) and similar in the two treatment groups. No clinically significant changes in blood chemistry, in particular calcium levels, occurred. The greater efficacy of combined calcitriol and phototherapy allowed a 34% decrease in total UVB exposure. CONCLUSIONS: Calcitriol 3 microg g 1 ointment and UVB phototherapy in combination provides a promising therapy for managing chronic plaque psoriasis. PMID- 11260006 TI - Rationale for use and clinical responsiveness of hexafluoro-1,25-dihydroxyvitamin D3 for the treatment of plaque psoriasis: a pilot study. AB - BACKGROUND: Vitamin D analogues are useful in topical therapy of psoriasis. OBJECTIVES: To evaluate the effect of hexafluoro-1,25-dihydroxyvitamin D3 (F6 1,25(OH)2D3) in treatment of psoriasis. METHODS: Fifteen patients with plaque type psoriasis were enrolled in a single centre double-blind, right/left comparison, placebo-controlled study, and received 0.1 g of petrolatum containing 5 microg of F6-1,25(OH)2D3 or 0.1 g of petrolatum (placebo) for 3 months. After completion of this double-blind study, a subset of these patients (n = 12) applied F6-1,25(OH)2D3 ointment (50 microg g-1 of petrolatum) to all their lesions (total area, 100-5000 cm2, mean area: 3300 m2) for 2 months as a single application at night. RESULTS: The mean severity score in the right/left-sided controlled topical F6-1,25(OH)2D3 (50 microg g-1) therapy group showed a decrease of 85%. In contrast, the mean severity score for the placebo-treated areas showed a decrease of 45% (P < 0.001). In the 12 patients who subsequently applied F6 1,25(OH)2D3 (50 microg g-1) ointment to all of their lesions, 91.6% showed moderate to excellent improvement. The mean Psoriasis Area and Severity Index score decreased by 44.9% (from 33.6 +/- 15 to 18.5 +/- 13). No effect on calcium homeostasis was noted. Adverse events included mild irritation in two patients that resolved during therapy. CONCLUSIONS: Topical F6-1,25(OH)2D3 is a safe and effective once a day treatment for psoriasis. PMID- 11260007 TI - The new topical ascomycin derivative SDZ ASM 981 does not induce skin atrophy when applied to normal skin for 4 weeks: a randomized, double-blind controlled study. AB - BACKGROUND: SDZ ASM 981 is a selective inhibitor of inflammatory cytokines released from T lymphocytes and mast cells, which has been developed for the treatment of inflammatory skin diseases. OBJECTIVES: In the present study, the atrophogenic potential of SDZ ASM 981 1% cream in humans was compared with that of medium and highly potent topical steroids, and vehicle. METHODS: Four different preparations, SDZ ASM 981 1% cream, the corresponding vehicle of SDZ ASM 981 1% cream, betamethasone-17-valerate 0.1% cream and triamcinolone acetonide 0.1% cream, were applied to the volar aspect of the forearms of 16 healthy volunteers, twice daily, 6 days a week, for 4 weeks. Skin thickness was evaluated by ultrasound examination, clinical signs of atrophy by stereomicroscopy, and epidermal thickness was assessed by histology. RESULTS: Both topical corticosteroids induced a significant reduction in skin thickness, as compared with SDZ ASM 981 1% cream and vehicle, which were shown to be equivalent. The difference in skin thickness (measured by ultrasound examination) between patients treated with SDZ ASM 981 1% cream and those receiving either of the two topical steroids was significant from day 8 onwards. Histological analysis performed at day 29 showed significant epidermal thinning with topical steroids compared with SDZ ASM 981 1% cream or the vehicle. Conclusion The lack of atrophogenic properties of SDZ ASM 981 1% cream in this short-term study demonstrates its potential as long-term treatment for inflammatory skin diseases, thus overcoming a major drawback of topical steroids. This may also be important for the treatment of children, and sensitive areas of skin, such as the face and skin-folds. PMID- 11260008 TI - What is the cost of atopic dermatitis in preschool children? AB - BACKGROUND: Atopic dermatitis (AD) is the most common inflammatory skin disorder and an important cause of morbidity in young children in the U.K. Such disability produces significant economic burden reflected in direct medical costs associated with health service utilization, direct family care costs such as transport costs, indirect costs associated with loss of productivity of carers, and intangible costs associated with the psychological effects of the disease. OBJECTIVES: In order to evaluate this economic burden we conducted a cross sectional survey of children aged 1-5 years in the Nottingham area in 1995-96. METHODS: We sent a postal questionnaire to parents of all children aged 1-5 years with questions to identify those with AD; a second questionnaire was sent to parents of identified children regarding costs and utilization of medical services. Intangible costs were not evaluated. RESULTS: The 12-month period prevalence of AD according to a dermatologist's diagnosis in 1761 children was 16.5% (95% confidence interval 14.7-18.2%). Total mean disease costs were estimated to be pound79.59 per child over the 12-month period of the study. The most significant costs were due to costs to the state for National Health Service (NHS) consultations ( pound28.62 mean annual cost) and prescriptions ( pound22.03). Consultations with general practitioners accounted for the significant bulk of consultation costs, with only 6% of children being seen in secondary care (17 of 290). Most prescribing costs (76%) were due to emollients and bath preparations. Family care costs ( pound28.94 mean annual cost) accounted for 36% of total disease costs and were associated with changes to the home environment, purchase of over-the-counter medicines, transport costs, visits to homoeopaths and salary loss. CONCLUSIONS: The results signify that AD is an important cause of economic burden both to the NHS and to the families of affected children. Using population census data and the results in this study, we estimated that the annual U.K. cost of AD in children aged 1-5 years in 1995-96 was pound47 million. PMID- 11260009 TI - Prevalence of atopic dermatitis, asthma, allergic rhinitis, and hand and contact dermatitis in adolescents. The Odense Adolescence Cohort Study on Atopic Diseases and Dermatitis. AB - BACKGROUND: Atopic diseases are common in children and adolescents. However, epidemiological knowledge is sparse for hand eczema and allergic contact dermatitis in this age group. Furthermore, no population-based studies have evaluated the prevalence of atopic diseases and hand and contact dermatitis in the same group of adolescents. OBJECTIVES: To assess prevalence measures of atopic dermatitis (AD), asthma, allergic rhinitis and hand and contact dermatitis in adolescents in Odense municipality, Denmark. METHODS: The study was carried out as a cross-sectional study among 1501 eighth grade school children (age 12-16 years) and included questionnaire, interview, clinical examination and patch testing. RESULTS: The lifetime prevalence of AD was 21.3% (girls 25.7% vs. boys 17.0%, P < 0.001) using predefined questionnaire criteria. The 1-year period prevalence of AD was 6.7% and the point prevalence 3.6% (Hanifin and Rajka criteria). In the interview the lifetime prevalence of inhalant allergy was estimated as 17.7% (6.9% allergic asthma, 15.7% allergic rhinitis). The lifetime prevalence of hand eczema based on the questionnaire was 9.2%, the 1-year period prevalence was 7.3% and the point prevalence 3.2%, with a significant predominance in girls. A significant association was found both between AD and inhalant allergy, and between AD and hand eczema using lifetime prevalence measures. The point prevalence of contact allergy was 15.2% (girls 19.4% vs. boys 10.3%, P < 0.001), and present or past allergic contact dermatitis was found in 7.2% (girls 11.3% vs. boys 2.5%). Contact allergy was most common to nickel (8.6%) and fragrance mix (1.8%). CONCLUSIONS: High prevalence figures were found for atopic diseases, hand eczema and allergic contact dermatitis, and the diseases were closely associated. A considerable number of adolescents still suffers from AD, and a considerable sex difference was noted for hand eczema and allergic contact dermatitis. Nickel allergy and perfume allergy were the major contact allergies. In the future this cohort of eighth grade school children will be followed up with regard to the course and development of atopic diseases, hand eczema and contact dermatitis. PMID- 11260010 TI - HFE mutations and transferrin receptor polymorphism analysis in porphyria cutanea tarda: a prospective study of 36 cases from southern France. AB - BACKGROUND: Porphyria cutanea tarda (PCT) is associated in most cases with iron overload, which may participate in decreased activity of uroporphyrinogen decarboxylase in the liver. The aetiology of this iron overload remains unknown; however, it has been demonstrated that mutations of HFE, the genetic haemochromatosis gene, might be present in a significant proportion of Anglo Saxon and Italian patients. Furthermore, transferrin receptor polymorphism may influence the affinity of this receptor to its ligand with a subsequent increase of cellular iron absorption and storage. OBJECTIVES: To evaluate the incidence and spectrum of HFE mutations and the relative frequency of the two main alleles of transferrin receptor in patients with PCT originating from southern France, and to evaluate the relationship of these genetic data with iron status, and with hepatitis B and C and human immunodeficiency virus (HIV) infections. METHODS: Thirty-six consecutive patients with either sporadic or familial PCT were prospectively included between 1997 and 2000. Search for the presence of the three main mutations of the HFE gene and identification of the transferrin receptor alleles were performed using polymerase chain reaction followed by enzymatic digestion. Iron parameters and viral status for hepatitis B and C viruses and HIV were determined. RESULTS: Seven patients (19%) showed heterozygous C282Y mutation, but no C282Y homozygote was present; five patients (14%) carried homozygous H63D mutation, while eight (22%) were heterozygous for this mutation. One patient was heterozygous for the S65C mutation (3%). Iron parameters demonstrated overload in all patients, without a clear difference between patients with and without deleterious mutations of the HFE gene. Infection by hepatitis C virus was documented in 20 patients (56%), and was significantly less frequent in patients with deleterious HFE mutations. The profile of transferrin receptor alleles in PCT patients did not show significant variation compared with the general population. CONCLUSIONS: This study confirms the high frequency of HFE mutations in patients with PCT and supports the hypothesis that HFE gene abnormalities might play a significant part in the PCT pathomechanism, probably through iron overload; by contrast, transferrin receptor polymorphisms do not appear to play a significant part in iron overload in PCT. PMID- 11260011 TI - Assessment of skin basement membrane zone antibodies in the urine of patients with acquired subepidermal immunobullous diseases. AB - BACKGROUND: In bullous pemphigoid (BP), cicatricial pemphigoid (CP) and linear IgA disease (LAD), autoantibodies to the basement membrane zone (BMZ) are found in skin and mucosa, blood and blister fluid. OBJECTIVES: To assess whether BMZ antibodies might also be detected in urine. METHODS: Urine and serum samples from 62 patients (32 with BP, 17 with CP and 13 with LAD) were analysed for antibody isotypes and subclasses by indirect immunofluorescence, and urine and serum samples from 40 patients (25 with BP, eight with CP and seven with LAD) were screened for target antigens using immunoblotting. RESULTS: Fourteen of 32 patients with BP had detectable levels of IgG BMZ autoantibodies in their urine, and all 32 had positive sera. Of these 14 BP patients, 13 had epidermal-binding serum autoantibodies at a titre > 1 : 160, and one had dermal-binding serum antibodies at a titre of 1 : 40. BMZ autoantibodies were not detected in the urine of the CP or LAD patients, but the corresponding sera were of low titre or negative. IgG subclasses (IgG1-4) were less frequently detected in urine than in serum. IgG4 was the predominant subgroup found (10 urine samples and all 14 sera), followed by IgG1 (two urine samples and 12 sera); IgG2 was detected in a single urine sample and three sera, and IgG3 was not detected. Eight of 25 BP and one of eight CP urine samples were positive on immunoblotting, and bound BP230 and/or BP180 with IgA and/or IgG autoantibodies. IgA autoantibodies were not detected in the urine of the seven LAD patients. The corresponding sera were often more positive, with 21 of 25 BP, five of eight CP and six of seven LAD sera immunoblotting the major BP antigens. CONCLUSIONS: The detection of IgG autoantibodies from urine samples using indirect immunofluorescence correlated with a high titre of IgG autoantibodies in the serum. IgG and IgA autoantibodies in the urine were detected by immunoblotting, although less frequently than in serum. The finding of BMZ antibodies in the urine of many BP patients may have clinical relevance, and may have a restricted application in the diagnosis of immunobullous disease. PMID- 11260012 TI - Resistance to activated protein C due to factor V Leiden mutation: high prevalence in patients with post-thrombotic leg ulcers. AB - BACKGROUND: Activated protein C (APC) resistance is the most frequently diagnosed heritable thrombophilic defect predisposing to thrombosis. OBJECTIVES: To determine the prevalence of APC resistance due to factor V Leiden mutation in patients with leg ulcers. METHODS: Within a 2-year-period 100 consecutive patients with leg ulcers were examined for factor V Leiden mutation. RESULTS: APC resistance due to factor V Leiden mutation was detected in 19 of 53 patients (36%) with post-thrombotic leg ulcers and in three of 47 patients (6%) with ulcers caused by primary varicosis. In a healthy control group APC resistance due to factor V Leiden mutation was found in five of 96 (5%) volunteers. CONCLUSIONS: In view of this high prevalence of APC resistance of 36%, which has never previously been reported, patients with post-thrombotic leg ulcers should be investigated for APC resistance. PMID- 11260013 TI - Seborrhoeic dermatitis and Pityrosporum (Malassezia) folliculitis: characterization of inflammatory cells and mediators in the skin by immunohistochemistry. AB - BACKGROUND: The fact that Pityrosporum ovale plays a part in seborrhoeic dermatitis is well established but the mechanism of this relationship has not been established. OBJECTIVES: To compare the number and type of inflammatory cells and mediators in skin biopsies from normal and lesional skin from the trunk and scalp in patients with seborrhoeic dermatitis, Pityrosporum (Malassezia) folliculitis and in normal skin from healthy controls. METHODS: The skin biopsies were stained using the labelled Streptavidin-biotin METHOD: The following markers were studied: CD4, CD8, CD68, HLA-DR, NK1, CD16, C1q, C3c, IgG, CD54 (ICAM-1), interleukin (IL) -1alpha, IL-1beta, IL-2, IL-4, IL-6, IL-10, IL-12, tumour necrosis factor-alpha and interferon-gamma. RESULTS: HLA-DR+ cells were seen in the highest number, and were higher in lesional skin compared with normal skin from both patients and healthy volunteers. ICAM-1 expression was also increased in lesional skin. C1q and the interleukins showed an increased cellular and intercellular staining in patients compared with healthy controls and the intercellular staining was often more intense in lesions compared with non lesional skin. Staining was often more intense when Malassezia (Pityrosporum ovale) yeast cells were present. CONCLUSIONS: An increase in NK1+ and CD16+ cells in combination with complement activation indicates that an irritant non immunogenic stimulation of the immune system is important. The result with the interleukins showed both an increase in the production of inflammatory interleukins as well as in the regulatory interleukins for both TH1 and TH2 cells. Similarities to the immune response described for Candida albicans infections indicate the role of Malassezia in the skin response in seborrhoeic dermatitis and Pityrosporum folliculitis. PMID- 11260014 TI - Nail pathology in patients with hemiplegia. AB - BACKGROUND: It is well known that nails can be involved in some diseases of the central nervous system; however, no systematic study has been carried out in order to evaluate the incidence and the possible mechanisms of these nail changes in hemiplegia. OBJECTIVES: To study the presence of nail pathology specifically associated with hemiplegia and to evaluate its incidence and its temporal relationship with the onset of the neurological deficit. METHODS: In an open study, fingernails and toenails were examined by a dermatologist; 108 were patients with hemiplegia due to a stroke, consecutively admitted to our Department of Neurology between 1995 and 1998, and 121 were normal controls. RESULTS: Onychodystrophy of fingernails and onychomycosis of toenails were found in both patients with hemiplegia and normal controls. However, three conditions (longitudinal reddish striation, neapolitan nails and unilateral clubbing) were only observed in some patients, always affecting fingernails of the limb affected by hemiplegia. Neapolitan nails were present in three (3%) patients with hemiplegia which had its onset 3-14 months earlier. Hemiplegia had occurred approximately 40 months earlier, on average, in six patients (6%) with longitudinal reddish striation, and 60-120 months prior to unilateral clubbing in another two patients (2%). CONCLUSIONS: In this study we were able to assess the presence of three different fingernail conditions that were characteristically associated with hemiplegia (longitudinal reddish striation, neapolitan nails and unilateral clubbing), to evaluate their incidence and to study the delay with which these changes occur after a stroke. PMID- 11260015 TI - Bone marrow involvement in cutaneous mastocytosis. AB - BACKGROUND: Cutaneous mastocytosis is considered a relatively benign and indolent form of mast cell disease, which either ultimately regresses, remains stable or is only slowly progressive. Previously, it has been purported that no more than 60% of adult patients with cutaneous mastocytosis will have occult bone marrow involvement. OBJECTIVES: To investigate the frequency of bone marrow involvement in patients with mastocytosis but without systemic symptoms. METHODS: Bone marrow aspirate and trephine biopsy were performed in 13 consecutive patients with cutaneous mastocytosis attending our department. RESULTS: All but one of these patients had evidence of bone marrow involvement. Bone marrow cytogenetic abnormalities have been found in patients with cutaneous mastocytosis: all our patients who were analysed showed a normal karyotype. CONCLUSIONS: Bone marrow involvement is common in adults with cutaneous mastocytosis. PMID- 11260016 TI - Bowen's disease, solar keratoses and superficial basal cell carcinomas treated by photodynamic therapy using a large-field incoherent light source. AB - BACKGROUND: Photodynamic therapy (PDT) has not yet been demonstrated to be superior to conventional treatment in the treatment of superficial skin cancers and premalignant skin conditions. A limitation for PDT is the absence to date of a light source suitable for the treatment of larger lesions or 'field changes' where several lesions are present on one anatomical site. OBJECTIVES: To investigate the safety and efficacy of a large field light source, the Waldmann PDT 1200, in the treatment of Bowen's disease (BD), superficial basal cell carcinomas (BCCs) and solar keratoses (SKs). METHODS: After application of 5 aminolaevulinic acid for 4-6 h, each lesion was irradiated with 105 J cm-2 of incoherent red light centred on 640 nm. Eighty-eight patients with 239 lesions were recruited. RESULTS: Within two treatments, 88% of BD lesions, 95% of BCCs and 99% of SKs showed complete clinical clearance. At 12 months the complete response rates were 69% for BD, 82% for BCC and 72% for SK. CONCLUSIONS: This study confirms that PDT is a useful treatment and that selected superficial BCCs and SKs respond well to PDT. The PDT 1200 light source proved capable of treating multiple lesions amounting to a 'field change' and also lesions up to 10 cm in diameter within an acceptable treatment time. Thus far, PDT has failed to become established as a routine treatment for small premalignant and malignant skin lesions as it has not proved superior to simple cheaper conventional therapies such as cryotherapy, curettage and cautery, topical chemotherapy with 5 fluorouracil, or surgery. However, PDT has become established as a treatment for selected cases in some centres. This study suggests a role for PDT in the treatment of large premalignancies, superficial BCCs and field change where existing treatments may be problematic. PMID- 11260017 TI - Photodynamic therapy of acne vulgaris with topical delta-aminolaevulinic acid and incoherent light in Japanese patients. AB - BACKGROUND: Photodynamic therapy (PDT) is useful for treatment of epidermal neoplasia but may also have a role in the treatment of inflammatory dermatoses. OBJECTIVES: To study the effect of PDT in patients with acne. METHODS: Three men and 10 women who suffered from intractable acne vulgaris were treated using PDT with topical delta-aminolaevulinic acid (ALA) and polychromatic visible light. Twenty per cent ALA in an oil-in-water emulsion was applied to the lesions for 4 h with a light-shielding dressing. The lesions were then exposed to polychromatic visible light at 600-700 nm using a halogen light source of energy intensity 17 mW cm-2 and a total energy dose of 13 J cm-2. RESULTS: All patients had apparent improvement of facial appearance and reduction of new acne lesions at 1, 3 and 6 months following PDT treatment. The adverse effects were discomfort, burning and stinging during irradiation, oedematous erythema for 3 days after PDT, epidermal exfoliation from the fourth to the 10th day, irritation and hypersensitivity to physical stimulation for 10 days after PDT, and pigmentation or erythema after epidermal exfoliation; the treated lesions returned to normal skin conditions within 1 month. CONCLUSIONS: PDT was beneficial in the treatment of acne. As a photoactivating light source, polychromatic visible light was thought to be better for use with acne patients than laser light because of its cost effectiveness, uniform illumination and time-efficiency in treating large areas. PMID- 11260018 TI - Acute oedema blisters: a report of 13 cases. AB - BACKGROUND: The development of bullae accompanying acute oedema is a commonly observed clinical sign but has been rarely reported in the literature. OBJECTIVES: To document the clinical features and increase awareness of this entity. METHODS: Retrospective case note review of 13 inpatient consultation cases seen in two dermatology departments. RESULTS: Oedema blisters appear to be related to the speed of development rather than the degree of oedema, and respond rapidly to the reduction of the underlying oedema. CONCLUSIONS: Recognition of oedema blisters is important as they respond quickly and completely to treatment of the underlying oedema. PMID- 11260019 TI - Mycophenolate mofetil as a systemic antipsoriatic agent: positive experience in 11 patients. AB - BACKGROUND: Mycophenolate mofetil (MMF) is a novel immunosuppressive drug. Several case reports have suggested that MMF has a beneficial effect in patients with psoriasis and autoimmune dermatoses. OBJECTIVES: To investigate the efficacy and safety of oral MMF in severe psoriasis. METHODS: Eleven patients with severe stable plaque-type psoriasis and a Psoriasis Area and Severity Index (PASI) between 12 and 53 (mean 30.5) were included in the study. They received oral MMF 1 g twice daily for 3 weeks and then 0.5 g twice daily for 3 weeks. The PASI were determined at baseline (week 0) and after 1, 2, 3 and 6 weeks of treatment. RESULTS: Within 3 weeks of this therapy there was a reduction in PASI of between 40% and 70% in seven of 11 patients, and only one patient achieved a reduction in PASI of < 25% from baseline (mean PASI 15.6). Reducing MMF from 2 g daily to 1 g daily led to further, although only slight, improvement in six of 11 patients during the following 3 weeks. In four of 11 patients, the PASI increased at this lower dosage, and in one patient the drug was withdrawn because of muscle pain, which was possibly drug induced. This side-effect reversed within a few days after stopping the drug. Other side-effects, especially gastrointestinal and haematological toxicity, were not observed in any of the 11 patients treated. Overall, the mean PASI was 16.1 after 6 weeks. CONCLUSIONS: We conclude that the immunosuppressant MMF 2 g daily is effective and safe in the treatment of severe psoriasis. PMID- 11260020 TI - Treatment of psoriatic arthritis with antitumour necrosis factor-alpha antibody clears skin lesions of psoriasis resistant to treatment with methotrexate. AB - BACKGROUND: In inflamed skin, keratinocytes and inflammatory cells both produce large amounts of tumour necrosis factor (TNF) -alpha, a cytokine with broad effects that are relevant to inflammation. Blockade of this proinflammatory cytokine by a monoclonal anti-TNF-alpha antibody might be effectively used in the treatment of inflammatory skin diseases. OBJECTIVES: To gather information about the efficacy of an anti-TNF-alpha antibody (infliximab) in the treatment of skin lesions of psoriatic arthritis. METHODS: Six patients with progressive joint disease and psoriatic skin lesions unresponsive to methotrexate therapy were treated with anti-TNF-alpha antibody. The Psoriasis Area and Severity Index was determined before and 10 weeks after initiation of therapy. RESULTS: Improvement of psoriatic skin lesions was observed in all patients. In addition, a marked improvement of the joint disease was noted. CONCLUSIONS: Therapy with anti-TNF alpha antibody may be an effective treatment regimen for both psoriatic arthritis and psoriatic skin lesions. PMID- 11260021 TI - Cutaneous mucinosis of infancy: is it a real entity or the paediatric form of lichen myxoedematosus (papular mucinosis)? AB - We describe a girl presenting with a childhood dermal mucinosis in which we had the unique opportunity to find all the transitional histological features of lichen myxoedematosus (papular mucinosis), from its early focal mucin deposition in the reticular dermis to its late findings of interstitial mucin deposition, dermal fibrosis and fibroblast proliferation. Her father reported having had similar lesions when he was a child, which completely disappeared during adolescence. This case, and a re-evaluation of the literature, suggests that cases of cutaneous mucinosis of infancy that are not hamartomatous conditions such as mucinous naevi are in fact the infantile presentation of lichen myxoedematosus (papular mucinosis) and, in addition to other cases in the literature, suggests a genetic and familial factor in lichen myxoedematosus (papular mucinosis). PMID- 11260022 TI - Lichen myxoedematosus with associated cardiac abnormalities. AB - We describe a 42-year-old woman who developed lichen myxoedematosus. Twenty years after the onset of the disease she became breathless and hypertensive, and an echocardiogram showed a mass on the mitral valve, which was thought to be a mucin deposit. Her hypertension was resistant to treatment with combination antihypertensives. To our knowledge, this is the first report to link lichen myxoedematosus with a valvular mucinous mass. This case also demonstrates the slow clinical progression of the disease over 20 years. PMID- 11260023 TI - Common variable immunodeficiency treated with a recombinant human IgG, tumour necrosis factor-alpha receptor fusion protein. AB - Common variable immunodeficiency (CVI) is characterized by a failure in B-cell differentiation and impaired immunoglobulin secretion, but with a variable clinical presentation, including the development of sarcoidal granulomas and autoimmune diseases, as well as an increased incidence of malignancies. We present a 21-year-old white man who carried a diagnosis of juvenile rheumatoid arthritis and presented 6 years later with scarring alopecia showing sarcoidal granulomas. Further work confirmed the diagnosis of CVI, and with increasing systemic symptoms, it was elected to treat the patient with a tumour necrosis factor (TNF)-alpha antagonist, a TNF-alpha receptor IgG1 fusion protein. The patient showed improvement in his systemic symptoms and some hair regrowth after 3 months of therapy, and continued improvement in his systemic disease with only mild scalp hair thinning in the areas of prior involvement after almost 1 year of therapy. CVI and sarcoid may have overlapping clinical and immunological findings. Previous therapies for CVI, including intravenous immunoglobulin, have not altered the mortality of the disease. TNF-alpha is a primary cytokine and is elevated in CVI, and specific inhibition of TNF-alpha in this patient was effective in moderating his disease, including his skin disease. PMID- 11260024 TI - Tuberculous gumma following venepuncture. AB - An 88-year-old man with a past medical history of pulmonary tuberculosis presented with a 2-year history of an indolent enlarging ulcerated nodule on his left wrist following a venous blood test. Skin biopsy of the ulcer showed granulomas, Ziehl-Neelsen stain was negative, and cultures grew Mycobacterium tuberculosis. Underlying bone and joint disease was excluded, and the lesion healed completely with 6 months of standard antituberculous treatment. We review the literature on tuberculous gumma following an injury. PMID- 11260025 TI - Arteriovenous haemangioma in chronic liver disease: clinical and histopathological features of four cases. AB - Chronic liver diseases are known to cause several skin manifestations, including cutaneous vascular changes such as spider naevus and palmar erythema. Arteriovenous haemangioma (AVH), a benign acquired cutaneous vascular lesion, has also been reported to be associated with chronic liver disease. We report here four cases of AVH in patients with chronic liver disease: (i) a 59-year-old man who had suffered from chronic active hepatitis associated with hepatitis C virus for 15 years; (ii) a 48-year-old man who had suffered from alcoholic hepatitis for 3 years and was diagnosed with liver cirrhosis 1 year ago; (iii) a 69-year old female who had had chronic active hepatitis associated with hepatitis C virus infection for 20 years and was diagnosed with liver cirrhosis 7 years ago; and (iv) a 48-year-old man who had had chronic active hepatitis associated with hepatitis B virus infection for about 20 years. All patients showed an asymptomatic solitary dome-shaped reddish papule, 5-10 mm in diameter, on the face (first, second and third patients) or chest (fourth patient). Histopathological examination of the tumours revealed features of AVH, namely a well-circumscribed lesion composed of vascular structures of various sizes reminiscent of arteries and veins. In all four cases, elastic-van Gieson stain showed the absence of an internal elastic lamina in the thick-walled vessels as well as the thin-walled vessels. Examination of multiple sections demonstrated glomus cells and an ascending artery feeding the tumour vessels in only one case. Slight inflammatory cell infiltration was seen in the stroma of three patients while Toluidine blue staining revealed an increased number of mast cells in the stroma in all lesions. The present cases suggest that the occurrence of AVH associated with chronic liver disease is not related to any specific liver disease, but may be related to chronic liver dysfunction itself. PMID- 11260026 TI - Paraneoplastic pemphigus without antidesmoglein 3 or antidesmoglein 1 autoantibodies. AB - Paraneoplastic pemphigus (PNP) is an autoimmune mucocutaneous blistering disease characterized by IgG autoantibodies that bind to various epithelia and immunoprecipitate a complex of 250, 230, 210, 190 and 170 kDa proteins. A recent study has suggested that PNP patients have antidesmoglein (Dsg) 3 autoantibody and that the antibody plays a pathogenic role in PNP. We report a 72-year-old woman with PNP associated with thymoma and adenocarcinoma of the lung. Diagnosis of PNP was made by the characteristic clinical, histological and immunopathological findings, as well as immunoprecipitation of characteristic 230, 210 and 190 kDa proteins. Using enzyme-linked immunosorbent assay with baculovirus-expressed recombinant proteins, the patient's serum was negative against both Dsg 3 and Dsg 1. This finding is unusual, and it suggests that the target antigen, which is involved in acantholysis, may be other than Dsg 3 in this case. PMID- 11260027 TI - Hidden scabies: diagnosis by polymerase chain reaction. AB - Diagnosis of scabies infection can be difficult as in many cases only few mites are present on an infected person, and in some cases the skin manifestations can be subtle or atypical. We describe the use of polymerase chain reaction (PCR) to amplify Sarcoptes scabiei DNA in a patient presenting with clinically atypical eczema. Cutaneous scales were PCR positive for S. scabiei DNA before, and negative 2 weeks after, therapy. This method facilitates fast and very sensitive diagnosis of clinically atypical or inapparent scabies infection and therapy control in severely affected patients and may help to identify previously unrecognized scabies cases. PMID- 11260028 TI - Isolated human papillomavirus 18-positive extragenital bowenoid papulosis and idiopathic CD4+ lymphocytopenia. AB - We report a case of isolated extragenital bowenoid papulosis (BP) in a young man with an idiopathic low CD4 count. The lesions occurred on the dorsal aspect of his left middle finger and were not associated with genital involvement. Polymerase chain reaction studies of a biopsy demonstrated human papillomavirus 18. As far as we are aware, this is the first documented case of BP (genital or extragenital) associated with idiopathic CD4 lymphocytopenia. PMID- 11260029 TI - Subacute cutaneous lupus erythematosus and life-threatening hypokalaemic tetraparesis: a rare complication. AB - We describe a patient with lesions of subacute cutaneous lupus erythematosus associated with high titres of anti-Ro and anti-La antibodies who developed a rapidly progressive flaccid tetraparesis due to profound hypokalaemia. After investigation she was found to have distal renal tubular acidosis (dRTA) without pathological evidence of lupus nephritis. It is likely that her dRTA was a manifestation of associated Sjogren's syndrome, which had been otherwise asymptomatic. This is the first report of such a complication in the dermatological literature. PMID- 11260030 TI - Clear cell syringofibroadenoma (of Mascaro) of the nail. AB - Eccrine syringofibroadenoma (ESFA) is a rare disorder. We report the first case of ESFA of the nail apparatus, which presented as a yellow longitudinal onycholytic band of the left fourth finger over an intermittently painful subungual filamentous tumour. Histological examination showed features of ESFA with a digitate pattern of papillomatosis due to the specialized physiological longitudinal arrangement of the ridges in the nail bed. In addition, we describe a new feature of colloidal iron-positive clear cells. In our case, the presence of two types of cells with a central ductal differentiation and a significant amount of mucopolysaccharides in clear cells could suggest differentiation towards both the ductal and the secretory portion of the eccrine gland. PMID- 11260031 TI - Pretreatment of psoriasis with the vitamin D3 derivative tacalcitol increases the responsiveness to 311-nm ultraviolet B: results of a controlled, right/left study. PMID- 11260032 TI - Intralesional injection of enoxaparin in benign symmetrical lipomatosis: an alternative to surgery? PMID- 11260033 TI - What is the cumulative effect of long-term, low-dose isotretinoin on the development of DISH? PMID- 11260034 TI - Intravenous regional anaesthesia for treatment of palmar hyperhidrosis with botulinum toxin type A. PMID- 11260036 TI - Mild, late-onset prolidase deficiency: another Italian case. PMID- 11260035 TI - Hormonal influence on reticular erythematous mucinosis. PMID- 11260037 TI - Compartment syndrome following an insect bite. PMID- 11260038 TI - Successful long-term treatment of severe atopic dermatitis with mycophenolate mofetil. PMID- 11260039 TI - Skin lesions as the only manifestation of the idiopathic hypereosinophilic syndrome. PMID- 11260040 TI - Primary cutaneous nocardiosis mimicking lupus erythematosus. PMID- 11260041 TI - Photosensitivity reaction associated with use of the combined oral contraceptive. PMID- 11260042 TI - Topical cidofovir for bowenoid papulosis in an HIV-infected patient. PMID- 11260043 TI - Porokeratosis of the penis. PMID- 11260044 TI - Pyogenic granuloma arising in port-wine stain during pregnancy. PMID- 11260045 TI - Topical granulocyte-macrophage colony-stimulating factor for radiation dermatitis of the vulva. PMID- 11260046 TI - Stump acne: a new variant of acne mechanica and a cause of immobility. PMID- 11260047 TI - Cutaneous toxicity after intradermal vaccination with Mycobacterium vaccae against lung cancer and malignant mesothelioma. PMID- 11260048 TI - Dermatology dressings: an incendiary potential. PMID- 11260049 TI - Images in haematology: amyloidosis. PMID- 11260050 TI - Images in haematology: non-Hodgkin's lymphoma of the pelvic bone. PMID- 11260051 TI - Clinical governance. PMID- 11260052 TI - Adhesion receptors on haematopoietic progenitor cells. PMID- 11260053 TI - Sir Leonard Parsons and the scientific basis of paediatric haematology. PMID- 11260054 TI - Long-term outcome of individualized prophylactic treatment of children with severe haemophilia. AB - The development of arthropathy is a serious complication of severe haemophilia. With the use of prophylaxis, bleeds can be prevented and arthropathy delayed. We investigated whether an individually tailored prophylactic regimen can prevent arthropathy and whether it had a similar effect on orthopaedic outcome compared with that of a high-dose regimen. Efficacy was determined clinically and by radiographs of six major joints. Prophylaxis was started in 70 patients at a mean age of 4.1 years. Mean follow-up was 15.6 years (range 8-24.5 years). The mean factor VIII consumption was 2319 IU/kg/year. The mean number of joint bleeds was 3.5/year and the mean clinical score (maximum score 90) was 1.0, with a mean Pettersson joint score (maximum score 78) of 3.0 at a mean age of 13.5 years. In conclusion, long-term, early-onset, individualized prophylaxis in haemophilia is feasible and prevents arthropathy. PMID- 11260055 TI - Novel mutations in the Factor VII gene of Taiwanese Factor VII-deficient patients. AB - The genetic defects of four Taiwanese patients with factor VII (FVII) deficiency were studied. FVII activity and antigen levels were < 1 u/dl and 125.7 u/dl (patient I), < 1 u/dl and < 1 u/dl (patient II), 3.4 u/dl and 5.9 u/dl (patient III), and 1.2 u/dl and 30.4 u/dl (patient IV) respectively. The 5' flanking region, and all exons and junctions were amplified using polymerase chain reaction and sequenced. Patient I was homozygous for a 10824C-->A transversion with Pro303-->Thr mutation in exon 8. In patient II, a heterozygous transversion, 9007+1G-->T at the IVS6, a heterozygous decanucleotide insertion polymorphism at 323 (both mutations present in his father) and a heterozygous deletion, del TC (26-27) in exon 1A (originating from his mother) were identified. Patient III had a homozygous 10961T-->G transversion with His348-->Gln mutation in exon 8. Patient IV had a heterozygous 10902T-->G transversion with Cys329-->Gly mutation in exon 8 (transmitted to her second son) and a heterozygous decanucleotide insertion polymorphism at -323 (transmitted to her third son). All but one of the FVII gene mutations detected in the four patients have not been previously reported. In conclusion, four novel mutations of the FVII gene in Taiwanese, including two missense mutations in exon 8, one point mutation at the exon 6 splice site and one deletion in exon 1A, were identified. PMID- 11260056 TI - The association of vitamin K status with warfarin sensitivity at the onset of treatment. AB - We investigated the association of vitamin K status with warfarin sensitivity among 40 orthopaedic patients beginning perioperative algorithm-dosed warfarin. Baseline vitamin K status was assessed using plasma vitamin K-1 and vitamin K-1 2,3 epoxide concentrations, and a questionnaire-based estimation of usual vitamin K intake. Warfarin sensitivity was assessed as the increase in the International Normalized Ratio (INR) after two doses of 5 mg of warfarin and as the 4-d accumulation of under-gamma-carboxylated prothrombin (PIVKA-II), adjusted for warfarin dose requirement. Multivariate models were used to assess vitamin K variables as predictors of warfarin sensitivity. The mean INR increase was 0.53 U and the mean PIVKA-II increase was 771 ng/ml/mg warfarin. Demographic factors were not associated with warfarin response. For each 1 standard deviation (SD) lower value of plasma vitamin K-1, but not the other vitamin K variables, the INR rose 0.24 U (P < or = 0.01). A higher usual vitamin K intake and plasma vitamin K 1, and lower plasma vitamin K-1 2,3 epoxide, were all associated with a lower PIVKA-II increase over 4 d. Respective differences in PIVKA-II accumulation per SD increase of each variable were -165, -218 and 236 ng/ml/mg warfarin (all P < or = 0.05). We concluded that dietary and biochemical measures of vitamin K status were associated with early warfarin sensitivity. PMID- 11260057 TI - Von Willebrand factor collagen binding activity in the diagnosis of von Willebrand disease: an alternative to ristocetin co-factor activity? AB - The capability of von Willebrand factor (VWF) to bind platelet glycoprotein Ib (GPIb) and promote platelet plug formation is currently evaluated in vitro using the ristocetin co-factor activity (VWF:RCo) assay. The replacement of this cumbersome and not always reproducible test with the collagen binding activity of VWF (VWF:CBA) has been attempted with controversial results. To evaluate the capacity of VWF:CBA to identify classic and variant von Willebrand disease (VWD) compared with VWF:RCo, we studied 10 type 2A and 12 type 2B VWD patients, together with 30 type 1 VWD patients with reduced platelet VWF content. In both 2A and 2B VWD, VWF:CBA and VWF:RCo were decreased, but that of VWF:CBA was more consistent. The difference was more evident when values were expressed as a ratio, obtained by normalizing VWF:CBA and VWF:RCo with the VWF antigen value; the ratio for VWF:CBA was always below 0.2, while that for VWF:RCo was greater than 0.4, and in no patient was the VWF:CBA value higher than VWF:RCo. In contrast, in type 1 VWD, the decrease in VWF:CBA was similar to that seen in VWF:RCo with the ratios always within the normal range. To better investigate the relationship between VWF:CBA and VWF:RCo, and the representation of large/intermediate VWF multimers, to which both tests are sensitive, 1-deamino cys-8-D-arginine-vasopressin (DDAVP) was infused in type 2A and 2B VWD patients. The differences between the two tests were even more evident after DDAVP, and in type 2A, even though large multimers were persistently decreased, VWF:RCo was normalized, while VWF:CBA remained defective. These findings clearly indicate that VWF:CBA detects the absence of large and intermediate VWF multimers better than VWF:RCo. Hence, we suggest adding VWF:CBA to the panel of tests employed in the diagnosis of VWD. Moreover, owing to the difficulty in performing VWF:RCo and its low reproducibility, we suggest that, when necessary, VWF:CBA may be substituted for VWF:RCo. PMID- 11260058 TI - Automated CD61 immunoplatelet analysis of thrombocytopenic samples. AB - Revision of the current decision point for prophylactic platelet transfusion in thrombocytopenic patients requires the availability of a method that is able to provide accurate platelet counts to as low as 1 x 109/l. This study is the first to evaluate the immunoplatelet method (CD61-Imm) of the haematological analyser Cell-Dyn 4000 in direct comparison with the flow cytometric procedure. Additionally CD61-Imm results were compared with CD4000 optical (Plto) counts in the ranges 20-547 x 109/l (n = 127) and 1-35 x 109/l (n = 107). The immunoplatelet and Plto results were in good agreement between 20 x 109/l and 547 x 109/l, but for samples of < 25 x 109/l the Plto tended to overestimate the counts. We determined the limits of detection (LD) and quantification (LLQ) for all three methods using standard statistical procedures. The LD for the flow cytometric CD41a method was 0.02 x 109/l compared with 0.009 x 109/l and 1.73 x 109/l for the CD61-Imm and Plto methods respectively. The LLQCV = 15% for the CD41a method was 1.8 x 109/l compared with 1.6 x 109/l and 18.0 x 109/l for the CD61-Imm and Plto procedures. In conclusion, (i) the CD61-Imm method performance is at least equivalent to the reference flow cytometric method, and (ii) in severe thrombocytopenia the CD61-Imm count is superior to the Plto count. PMID- 11260059 TI - Myristoylated alanine-rich C kinase substrate phosphorylation is involved in thrombin-induced serotonin release from platelets. AB - Stimulation of platelets by thrombin induces protein kinase C (PKC) activation, phosphorylation of pleckstrin, aggregation and serotonin release. Here, we demonstrate that, in human platelets, thrombin stimulation also induced phosphorylation of the myristoylated alanine-rich C kinase substrate (MARCKS) and serotonin release in intact and digitonin-permeabilized platelets. MARCKS is known to bind actin and cross-link actin filaments, and this is inhibited by PKC evoked MARCKS phosphorylation. MARCKS phosphorylation and serotonin release in response to increasing concentrations of thrombin have a similar EC50 and time course and, in permeabilized platelets, peptide MPSD, with an amino acid sequence corresponding to the phosphorylation site domain of MARCKS, blocked both responses. However, pleckstrin and myosin light chain phosphorylations were not modified. Ala-MPSD, in which the four serine residues of MPSD were substituted by alanines was ineffective. The results suggest a role for MARCKS in platelet secretion. The fact that pleckstrin phosphorylation has a different time course and was not modified in the presence of MPSD when MARCKS phosphorylation and serotonin release were inhibited would suggest either that pleckstrin phosphorylation is unrelated to secretion or that it might only be involved upstream in the events leading to secretion. PMID- 11260060 TI - Pathogenetic analysis of three cases with a bleeding disorder characterized by defective platelet aggregation induced by Ca2+ ionophores. AB - We report three cases of platelet dysfunction characterized by defective Ca2+ ionophore-induced platelet aggregation without impaired production of thromboxane A2 (TXA2). The patients had mild to moderate bleeding tendencies, and their platelet aggregation and secretion induced by ADP, collagen, arachidonic acid, stable TXA2 (STA2) and Ca2+ ionophore A23187 was defective or much reduced. However, ristocetin- or thrombin-induced platelet aggregation was normal. The analysis of second messenger formation showed that inositol 1,4,5-triphosphate formation or Ca2+ mobilization induced by thrombin, STA2 or A23187 was normal. Furthermore, the phosphorylation of 47 kDa protein (pleckstrin) and 20 kDa protein (myosin light chain, MLC) in response to those agonists was normal. These findings suggest that the defective site in the patients' platelets lies in the process distal to or independent of protein kinase C activation, Ca2+ mobilization and MLC phosphorylation. PMID- 11260061 TI - Platelet activation via the collagen receptor GPVI is not altered in platelets from chronic myeloid leukaemia patients despite the presence of the constitutively phosphorylated adapter protein CrkL. AB - In this study, we show that the adapter proteins CrkL and Cbl undergo increases in tyrosine phosphorylation and form an intracellular complex in platelets stimulated with the snake venom toxin convulxin, a selective agonist at the collagen receptor glycoprotein VI (GPVI). Constitutive tyrosine phosphorylation of CrkL has previously been reported in platelets from chronic myeloid leukaemia (CML) patients. This was confirmed in the present study, and shown to result in a weak constitutive association of CrkL with Cbl and a number of other unidentified tyrosine-phosphorylated proteins. There was no further increase in phosphorylation of CrkL in CML platelets in response to GPVI activation, whereas phosphorylation of Cbl and its association with CrkL were potentiated. In addition, this was accompanied by a small increase in p42/ 44 mapkinase (MAPK) activity in CML platelets. The functional consequence of the presence of constitutively phosphorylated proteins in CML platelets was investigated by measurement of aminophospholipid exposure and alpha-granule secretion. This revealed little alteration in the concentration-response curves for either in CML platelets stimulated via GPVI, although maximal levels of P-selectin were depressed. Despite the minimal effect on platelet activation in CML patients, we cannot exclude a role for CrkL or Cbl in signal transduction pathways stimulated via GPVI. PMID- 11260062 TI - Familial thrombocytosis as a recessive, possibly X-linked trait in an Arab family. AB - Familial thrombocytosis (FT) has previously been described as an autosomal dominant disorder with manifestations similar to those of sporadic essential thrombocythaemia. We studied an Arab family consisting of four brothers, aged 4-8 years, who had either sustained markedly elevated (> 1000 x 109/l) or moderately elevated (> 500 x 109/l) platelet counts, two healthy sisters and their parents who had normal platelet counts. The four brothers with FT had normal plasma thrombopoietin levels and are currently not presenting with any thrombotic or haemorrhagic complications. Mutation analysis at the thrombopoietin gene (THPO) of the affected family members failed to detect the intron 3 G-->C splice mutation that had been described as causing FT. In addition, segregation analysis using a polymorphic CA marker revealed completely discordant THPO alleles among the affected brothers. We postulate the existence of a new locus for FT whereby the disease is transmitted as a recessive, possibly X-linked trait. PMID- 11260063 TI - In vivo platelet activation in atherothrombotic stroke is not determined by polymorphisms of human platelet glycoprotein IIIa or Ib. AB - Platelet membrane glycoprotein polymorphisms are candidate risk factors for thrombosis, but epidemiological data are conflicting. Thus, demonstration of a genotype-dependent alteration in function is desirable to resolve these inconsistencies. We investigated in vivo platelet activation in acute thrombosis and related this to platelet genotype. Frequencies of the 1b and 2b alleles of the HPA 1a/1b and HPA 2a/2b platelet glycoprotein polymorphisms were determined in 150 (52 men/98 women, mean age 58.3 years) patients with atherothrombotic stroke, and the influence of genotype on markers of platelet activation was assessed. Platelet P-selectin (CD62P) expression and fibrinogen binding was measured using whole blood flow cytometry within 24 h of stroke and 3 months later in 77 patients who provided a repeat blood sample. Results were compared with matched controls. Neither the 1b allele [allele frequency 0.11 vs. 0.13, odds ratio (OR) confidence interval (CI) 0.8 (0.5-1.3)] nor the 2b allele [0.09 vs. 0.07, OR (CI) 1.4 (0.8-2.4)] was significantly over-represented in patients. Increased numbers of activated platelets were found following stroke (acute mean P-selectin expression 0.64% vs. control 0.35%, P < 0.001; acute mean fibrinogen binding 1.6% vs. control 0.9%, P < 0.001). Activation persisted in the convalescent phase (P < 0.001 and P = 0.005 vs. controls for P-selectin and fibrinogen respectively). Expression of P-selectin and fibrinogen was not influenced by either the HPA 1a/1b genotype (P > 0.95 for each marker, Scheffe's test) or the 2a/2b genotype (P > 0.95 for each). Although persisting platelet activation is seen in atherothrombotic stroke, it is independent of HPA 1a/1b and 2a/2b genotypes. These data suggest an underlying prothrombotic state, but do not support the polymorphisms studied as risk factors for thrombotic stroke in this population. PMID- 11260064 TI - Platelet glycoprotein IIb polymorphism, traditional risk factors and non-fatal myocardial infarction in young women. AB - Several platelet glycoprotein polymorphisms have been associated with an increased risk of myocardial infarction (MI) in studies that included predominantly men. In a population-based sample of 68 Caucasian women < 45 years old with non-fatal MI and 346 demographically similar control subjects, we found an increased risk of MI among women who possessed at least one copy of the glycoprotein IIb Ser843 allele compared with those lacking the Ser843 allele (odds ratio 1.85; 95% confidence interval = 1.03-3.33). The increased risk was present only in subgroups of women who smoked cigarettes, had hypercholesterolaemia or who had a family history of early onset MI. The Ser843 variant of glycoprotein IIb may be associated with an increased risk of MI in young women with other cardiovascular risk factors. Additional studies involving larger numbers of subjects are needed to confirm this preliminary finding. PMID- 11260065 TI - Age as the major predictive factor of long-term response to splenectomy in immune thrombocytopenic purpura. AB - Sixty-one consecutive patients undergoing splenectomy for chronic immune thrombocytopenia were retrospectively evaluated. Platelet response was considered as complete (CR) when platelet count rose to > 100 x 109/l, partial (PR) when 30 100 x 109/l or absent (NR) if otherwise. Follow-up (mean time 7.6 years) was possible in 54 patients. Forty-eight patients (88%) had an immediate response to splenectomy (39 CR, 9 PR) whereas six (12%) were NR. Thirty-six responders (67%) had sustained remission (31 CR; 5 PR) without further treatment; thrombocytopenia recurred in 12 patients (33%). The probability curve of continued remission showed a constant relapse-rate during the first 36 months; a further step of relapse was observed beginning 70 months after surgery. The only positive predictive factor for the long-term response to splenectomy was age < 40 (P < 0.005). Neither duration of thrombocytopenia nor previous response to medical treatment (steroids and/or intravenous immunoglobulins) were related to splenectomy response. PMID- 11260066 TI - Injecting drug use is a risk factor for deep vein thrombosis in women in Glasgow. AB - Three hundred and twenty-two consecutive women aged 16-70 years who presented with objectively confirmed symptomatic venous thromboembolism (VTE) were studied to determine precipitating factors for thrombosis. One hundred and eighty-seven presented with deep vein thrombosis (DVT), 116 with either definite or possible pulmonary embolism (PE) and 19 with both DVT and PE. Injecting drug use (IDU) via femoral vein puncture was a common risk factor for DVT, associated with 21.4% of all cases of DVT and 52.4% of cases of DVT in women under 40 years. All women with drug-related thrombosis presented with DVT. None presented with symptomatic PE. A number of clinically diagnosed DVT associated with IDU were also documented, suggesting that IDU may be the most common risk factor for DVT in our region. DVT associated with IDU presents significant management challenges. PMID- 11260067 TI - The fleet feet of haematopoietic stem cells: rapid motility, interaction and proteopodia. AB - Haematopoietic stem cells (HSCs) have been extensively characterized regarding in vivo engraftment, surface epitopes and genetic regulation. However, little is known about the homing of these rare cells, and their intrinsic motility and membrane deformation capacity. We used high-speed optical-sectioning microscopy and inverted fluorescent videomicroscopy to study highly purified murine lineage negative, rhodamine-low, Hoechst-low HSCs over time under various in vitro conditions. We discovered extremely rapid motility, directed migration to stromal cells and marked membrane modulation. High resolution images with three dimensional reconstruction showed the general presence of microspikes. Further, pseudopodia (proteopodia) were observed that were induced by stromal-derived factor-1 and steel factor. Proteopodia were directed towards and were quenched by stromal cells, at times bridged HSCs, and could rapidly retract or detach from cells. Proteopodia were also observed in vivo with homed HSCs in frozen sections of murine spleen, lung and heart. This is the first demonstration that HSCs are both fast and highly malleable in phenotype. PMID- 11260069 TI - Decreased prion protein expression in human peripheral blood leucocytes from patients with paroxysmal nocturnal haemoglobinuria. AB - The cellular isoform of the prion protein (PrPC) is a cell surface glycoprotein attached to the outer leaflet of the plasma membrane by a glycosylphosphatidyl inositol (GPI) anchor. PrPC is involved in the pathogenesis of prion diseases and has recently been shown to play a role in haemopoietic cell activation and proliferation. We have used the PrPC-specific monoclonal antibody (mAb) 3F4 in a flow cytometry approach to analyse the constitutive expression of PrPC on human peripheral blood (PB) cell populations from patients with paroxysmal nocturnal haemoglobinuria (PNH), which are characterized by a deficiency of GPI-linked cell surface proteins. Comparable PrPC expression levels (P > 0.05), quantified as mean fluorescent intensity, were measured on lymphocytes isolated from normal donors (n = 10) and patients with PNH (n = 5), whereas PNH PB monocytes and granulocytes exhibited substantially lower PrPC surface immunoreactivity than their normal counterparts (P < 0.05). More detailed histogram analyses of the PNH PB leucocytes revealed that PrPC was absent in PNH granulocytes, but was normally expressed in lymphocytes from four out of five patients. However, in one patient a bimodal distribution of 3F4 mAb staining was observed, indicating the presence of a PrPC-deficient lymphocyte subpopulation. In conclusion, our results show that PNH haemopoietic cells are deficient in cell surface-bound PrPC. PMID- 11260068 TI - The retinoblastoma family member, p107, is active in non-dividing monocyte derived dendritic cells. AB - A member of the retinoblastoma family of cell cycle regulatory proteins, p107, is not normally expressed in non-cycling cells. We demonstrate here that p107 is expressed in monocytes as they differentiate into dendritic cells under the influence of granulocyte-macrophage colony-stimulating factor and interleukin 4, a process shown not to involve cellular proliferation. We show that p107 is localized to the nucleus of these cells and is active, in that it binds an E2F DNA binding site, together with E2F transcription factors. These findings suggest a hitherto unknown role for p107 in non-proliferating dendritic cells that warrants further investigation. PMID- 11260070 TI - Extracellular Tat activates c-fos promoter in low serum-starved CD4+ T cells. AB - The regulatory human immunodeficiency virus-1 (HIV-1) Tat protein shows pleiotropic effects on the survival and growth of both HIV-1-infected and uninfected CD4+ T lymphocytes. In this study, we have demonstrated that low concentrations (10 ng/ml) of extracellular Tat protein induce the expression of both c-fos mRNA and protein in serum-starved Jurkat CD4+ lymphoblastoid T cells. Using deletion mutants, we demonstrates that the SRE, CRE and, to a lesser extent, also the SIE domains (all placed in the first 356 bp of c-fos promoter) play a key role in mediating the response to extracellular Tat. Moreover, the ability of Tat to activate the transcriptional activity of c-fos promoter was consistently decreased by pretreatment with the ERK/MAPK kinase inhibitor PD98058. Activation of c-fos is functional as demonstrated by induction of the AP 1 transcription factor, which is involved in the regulation of critical genes for the activation of T lymphocytes, such as interleukin 2. The Tat-mediated induction of c-fos and AP-1 in uninfected lymphoid T cells may contribute to explain the immune hyperactivation that characterizes the progression to autoimmuno deficiency syndrome and constitutes the optimal environment for HIV-1 replication, occurring predominantly in activated/proliferating CD4+ T cells. PMID- 11260071 TI - Rapid protein-based assays for the diagnosis of T-B+ severe combined immunodeficiency. AB - The severe combined immunodeficiencies (SCID) are a heterogeneous group of conditions arising from a variety of molecular defects. The X-linked form of SCID (X-SCID) is caused by defects in the common gamma chain (gammac), and is characterized by a T-B+NK- immunophenotype. This lymphocyte profile is seen in an autosomal recessive form of SCID caused by mutations in the JAK3 molecule. Thus, X-SCID and JAK3-deficient SCID are clinically and immunologically indistinguishable. Knowledge of the precise molecular defect is essential for antenatal diagnosis, carrier testing and for treatment using somatic gene therapy. To identify the molecular defect in children presenting with a T-B+NK- form of SCID, we have developed rapid assays based on flow cytometric analysis of gammac, immunoblotting for JAK3 and gammac, and detection of interleukin-2 (IL-2) induced tyrosine phosphorylation of JAK3. Sixteen T-B+NK- SCID patients from 15 families were examined. Nine had no detectable gammac, four had abnormal gammac expression and no IL-2-induced JAK3 tyrosine phosphorylation, and one had normal gammac expression but no IL-2-induced JAK3 tyrosine phosphorylation, although JAK3 was present. All these patients had mutations identified in their gammac gene. Two patients exhibited normal gammac expression, but JAK3 was not detected by immunoblotting and these patients were confirmed as having JAK3 gene mutations. Thus, these protein-based assays have led to rapid molecular diagnoses in T-B+ SCID that have subsequently been confirmed by genetic analysis. PMID- 11260072 TI - Bone marrow involvement in Whipple's disease: rarely reported, but really rare? AB - Infection with Tropheryma whippelii, the causative agent of Whipple's disease, involves nearly every organ. Involvement of bone marrow may be an overlooked area of Whipple's disease. We report a case of lymphoma-like Whipple's disease with bone marrow involvement together with a brief review of the literature on this topic. Despite minimal documentation, bone marrow may be commonly involved in Whipple's disease and, although not specific, diastase-resistant periodic acid Schiff (PAS)-positive macrophages in bone marrow may offer an important clue to diagnosis using PAS histology of upper endoscopic biopsies, polymerase chain reaction or electron microscopy. PMID- 11260073 TI - In vitro drug resistance and prognostic impact of p16INK4A/P15INK4B deletions in childhood T-cell acute lymphoblastic leukaemia. AB - p16 gene deletions are present in about 70% of primary paediatric T-cell acute lymphoblastic leukaemia (T-ALL) and 20% of common/precursor B-cell ALL cases. It is not clear what the impact of the frequent p16 deletions is within the subgroup of T-lineage ALL. We studied the relationship between p16/p19ARF deletions, using fluorescence in situ hybridization, and in vitro drug resistance and prognosis in childhood T-ALL at diagnosis. The cellular drug resistance was measured with the methyl thiazol tetrazoliumbromide assay using a panel of drugs and the thymidylate synthase inhibition assay for methotrexate. There was a complete overlap of individual LC50 values of p16 gene homozygously deleted and p16 germ line cases for most of the nine classes of drugs tested. The only difference was for dexamethasone: the p16-deleted group was more sensitive than the germ-line p16 group (P = 0.030). The homozygously deleted p16 T-ALL patients (n = 34) treated with the modern multiagent chemotherapy schemes of the Dutch Childhood Leukaemia Study Group ALL-VII/-VIII or Co-operative ALL-92/-97 protocols have a significantly lower 5-year disease-free survival (DFS) than germ-line p16 T-ALL (n = 25) (65.1 +/- 9.1% vs. 95.5 +/- 4.4%, Plog rank = 0.021). Hence, this study identifies a subpopulation of primary childhood T-ALL that appears to have an extremely high DFS. However, the observed differences in outcome do not seem to be related to intrinsic resistance for the tested drugs. PMID- 11260074 TI - Single site polymorphisms and alternative splicing of the human CD13 gene- different splicing frequencies among patients with acute myeloid leukaemia and healthy individuals. AB - Within the haematopoietic system, CD13/aminopeptidase N (APN), a transmembrane glycoprotein, is expressed on the surface of early committed progenitors of granulocytes and monocytes and by all cells of these lineages as they mature. CD13 is expressed on the majority of leukaemic myeloblasts in acute myeloid leukaemia (AML), and on leukaemic lymphoblasts in a small percentage of acute lymphoid leukaemia cases. Thus, anti-CD13 monoclonal antibodies are used as diagnostic markers in leukaemia typing. By systematically amplifying overlapping reverse transcription polymerase chain reaction (RT-PCR) amplicons throughout the CD13 mRNA, we identified two splice variants in which exon 3 and exon 14 were lost. Fourteen healthy individuals and 34 patients with AML were screened for these splice variants. All healthy individuals, and the majority of AML patients, had both splice variants but they represented less than 10% of the total RT-PCR amplified CD13 product. Increased expression of both truncated CD13 mRNA forms were observed in 6% of AML patients, whereas no detectable exon 3 or exon 14 splice variants could be generated in 26% and 9% of AML patients respectively. The different splicing frequencies may reflect altered processing of pre-mRNA or expansion of certain cell types for some AML patients, even though no correlation existed to blast percentage, FAB classification, surface antigens or cytogenetic characteristics. In addition, we identified an intron of 506 bp between exon 1 and exon 2 as well as two sites of single nucleotide polymorphism with a heterozygosity index of about 0.5, making them useful as genetic markers. PMID- 11260075 TI - Induction of the monocytic differentiation of myeloid leukaemia cells by cotylenin A, a plant growth regulator. AB - Regulators that play an important role in the differentiation and development of plants or invertebrates may also affect the differentiation of human leukaemia cells through a common signal transduction system, and might be clinically useful for treating acute myeloid leukaemia. Cotylenin A has been isolated as a plant growth regulator. We examined the effects of cotylenin A on the differentiation of several myelogenous leukaemia cells, and found that cotylenin A is a potent and novel inducer of the monocytic differentiation of human myeloid leukaemia cells. Cotylenin A induced the functional and morphological differentiation of myeloblastic and promyelocytic leukaemia cells, but did not effectively induce the differentiation of monocytoid leukaemia cells. Cotylenin A-induced differentiation was not affected by several inhibitors of signal transduction, suggesting that this inducer exhibits a unique mode of action. PMID- 11260076 TI - In vivo suppression of Bcl-XL expression facilitates chemotherapy-induced leukaemia cell death in a SCID/NOD-Hu model. AB - Bcl-XL, a member of the Bcl-2-related anti-apoptosis protein family, antagonizes a diverse range of apoptosis-inducing stimuli by preventing mitochondrial permeability transition, release of apoptogenic factors including cytochrome C, and caspase activation. We have tested the hypothesis that the susceptibility of Bcl-XL-expressing leukaemic cells to apoptosis induced by VP16 (etoposide) can be enhanced by pharmacological downregulation of Bcl-XL in vivo. Two subcutaneous xenograft models of B-cell leukaemia-employing SEMK-2 and BV173 cell lines were established in severe combined immunodeficient/non-obese diabetic mice followed by 14 d of continuous subcutaneous administration of Bcl-XL-specific second generation oligonucleotides ISIS 16009 or ISIS 15999. Tumours were disaggregated, enabling investigation of Bcl-XL expression and apoptosis susceptibility at single-cell resolution using cytofluorimetry. Marked sequence-specific reduction of Bcl-XL was associated with sequence-specific enhancement of VP16-induced mitochondrial permeability transition, caspase-3 activation and loss of membrane asymmetry. A negative correlation between Bcl-XL expression and apoptosis susceptibility was observed, together with a positive correlation with respect to a reduced redox state. Bcl-XL downregulation reduces the threshold for VP16 induced apoptosis by potentiating mitochondrial dysfunction and its sequelae, and therefore presents a novel therapeutic strategy for reversing chemoresistance. PMID- 11260077 TI - Apoptosis in refractory anaemia with ringed sideroblasts is initiated at the stem cell level and associated with increased activation of caspases. AB - Treatment with granulocyte colony-stimulating factor plus erythropoietin may improve haemoglobin levels in patients with ringsideroblastic anaemia (RARS) and reduce bone marrow apoptosis. We studied bone marrow from 10 RARS patients, two of whom were also investigated after successful treatment. Mononuclear, erythroid and CD34+ cells were analysed with regard to proliferation, apoptosis, clonogenic capacity and oncoprotein expression, in the presence or absence of Fas-agonist, Fas-blocking antibody 2 and caspase-3 inhibitor. During culture, RARS bone marrow cells showed higher spontaneous apoptosis (P < 0.05) and caspase activity (P < 0.05)) than bone marrow cells from healthy donors. Eight out of nine patients had reduced growth of erythroid colony-forming units (CFU-E) (< 10% of control) and granulocyte-macrophage CFU (CFU-GM) (< 50% of control) from CD34+ cells. Fas ligation increased apoptosis and decreased colony growth equally in RARS and controls, but caused significantly more caspase activation in RARS (P < 0.01). Fas-blocking antibody showed no significant inhibitory effect on spontaneous apoptosis or ineffective haematopoiesis, as measured using phosphatidylserine exposure, the terminal deoxynucleotide transferase-mediated dUTP-biotin nick-end labelling technique, caspase activity or clonogenic growth. Caspase inhibition reduced apoptosis, increased proliferation and enhanced erythroid colony growth from CD34+ cells in RARS, but showed no effect on normal cells. CFU-E improved > 1000% after successful treatment. Thus, erythroid apoptosis in RARS is initiated at the CD34+ level and growth factor treatment may improve stem cell function. Enhanced caspase activation at the stem cell level, albeit not mediated through endogenous activation of the Fas receptor, contributes to the erythroid apoptosis in RARS. PMID- 11260078 TI - Prognostic impact of bone marrow erythropoietic precursor cells and myelofibrosis at diagnosis of Ph1+ chronic myelogenous leukaemia--a multicentre study on 495 patients. AB - A multicentre clinicopathological study was performed on 495 patients with chronic-phase Ph1+ chronic myelogenous leukaemia (CML) to determine bone marrow characteristics that exert a significant impact on survival under standard treatment regimens. Immunohistochemical and morphometric techniques were applied to identify nucleated erythroid precursor cells in the bone marrow and to quantify argyrophilic fibre density. Application of the Sokal index and another recently proposed CML score failed to distinguish three clearly defined risk groups. A borderline increase in fibre content (i.e. doubling of the normal density) and a relevant reduction of medullary erythropoiesis proved to be important predictors for survival, even in low-risk classified patients, according to both clinical scores. With regard to optimal treatment strategies, patients with manifest myelofibrosis showed no significant difference in survival rates under interferon or hydroxyurea treatment. Multivariate analysis confirmed the prognostic value of histological features. A risk model based on three variables (fibre density, erythropoietic precursors and spleen size) was constructed that enabled a distinct discrimination of risk profiles. In conclusion, the presented data provide compelling evidence that bone marrow features at diagnosis exert a significant impact on prognosis in CML. In this context, the generally clinical-based multivariate risk classification can be improved by consideration of morphological variables that are acting independently of treatment modalities. PMID- 11260079 TI - Recognition of chronic myelogenous leukaemia cells by autologous T lymphocytes primed in vitro against the patient's dendritic cells. AB - Defects in immune responses are common in patients with chronic myelogenous leukaemia (CML). However, using dendritic cells (DCs) to promote T-cell immunity in vitro may nonetheless elicit potent specific anti-tumour responses for use in immunotherapy. Here, we show that DCs generated from CML patients had a typical dendritic phenotype and were able to stimulate autologous T cells. Three primed T cell lines were studied in more detail in one patient. They were stimulated by autologous CML cells, but not by normal non-leukaemic cells from the patient's HLA-identical sibling. This was blocked by HLA-DR-specific, but not HLA-DQ- or HLA-DP-specific antibodies. CML-stimulated cytokine secretion, including interferon-gamma and granulocyte macrophage-colony stimulating factor, suggested a Th1-type phenotype for these sensitized anti-leukaemic T cells. This study therefore shows that cells with a functional dendritic phenotype can be generated from the blood of CML patients and are potent inducers of T-cell responses to tumour cells. This approach allows sensitization of patients' T cells by their own particular tumour without the need to identify the exact leukaemia antigens involved, and may find application in immunotherapy of CML. PMID- 11260080 TI - Expression of bcr-abl mRNA in individual chronic myelogenous leukaemia cells as determined by in situ amplification. AB - We present the results of a novel method developed for evaluation of in situ amplification, a molecular genetic method at the cellular level. Reverse transcription polymerase chain reaction (RT-PCR) was used to study bcr-abl transcript levels in individual cells from patients with chronic myelogenous leukaemia (CML). After hybridizing a fluorochrome-labelled probe to the cell bound RT-PCR product, bcr-abl mRNA-positive cells were determined using image analysis. A dilution series of bcr-abl-positive BV173 into normal cells showed a good correlation between expected and actual values. In 25 CML samples, the percentage of in situ PCR-positive cells showed an excellent correlation with cytogenetic results (r = 0.94, P < 0.0001), interphase fluorescence in situ hybridization (FISH) (r = 0.95, P = 0.001) and hypermetaphase FISH (r = 0.81, P < 0.001). The fluorescence intensity was higher in residual CML cells after interferon (IFN) treatment than in newly diagnosed patients (P = 0.004), and was highest in late-stage CML resistant to IFN therapy and lowest in CML blast crisis (P = 0.001). Mean fluorescence values correlated with bcr-abl protein levels, as determined by Western blot analysis (r = 0.62). Laser scanning cytometry allowing automated analysis of large numbers of cells confirmed the results. Thus, fluorescence in situ PCR provides a novel and quantitative approach for monitoring tumour load and bcr-abl transcript levels in CML. PMID- 11260081 TI - Interleukin 4 content in chronic lymphocytic leukaemia (CLL) B cells and blood CD8+ T cells from B-CLL patients: impact on clonal B-cell apoptosis. AB - B-chronic lymphocytic leukaemia (CLL) clonal B cells are characterized by resistance to apoptosis. We evaluated clonal B cells and blood T cells for interleukin 4 (IL-4) content as IL-4 is able to increase CLL cell resistance to apoptosis. The content of IL-4 in CD8+ T cells of CLL patients (n = 9) ranged from 37% to 63% of the total CD8+ T cells (mean level of 49% +/- 3.4) compared with a range of 5-10% for control CD8+ T cells. Clonal B cells positive for cytoplasmic IL-4 ranged from 1% to 97% (mean value 57.8 +/- 6.9%). CD8+ T cells and clonal B cells secreted detectable levels of IL-4, but only clonal CLL B cells (n = 4) secreted IL-4 in association with increasing cell numbers. Fludarabine (F-ara-AMP, 0.1-100 micromol/ml) was able to downregulate the IL-4 content of CD8+ T cells, but not clonal B-cell IL-4. Culture supernatant from CLL CD8+ T cells decreased the spontaneous apoptotic rate of clonal B cells that was reversed with anti-IL-4 and soluble IL-4 receptor. These findings show that IL-4 is present in the microenvironment of B-CLL. In addition, use of agents that can interfere with IL-4 presentation to clonal B cells can be effective in increasing clonal B-cell apoptosis. PMID- 11260082 TI - MDM2 gene amplification and lack of p53 point mutations in Hodgkin and Reed Sternberg cells: results from single-cell polymerase chain reaction and molecular cytogenetic studies. AB - Hodgkin's disease (HD) is the most common haematological malignancy after chronic lymphocytic leukaemia, but very little is known about its pathogenesis or the genetic events that contribute to the malignant phenotype of the tumour cells. p53 is assumed to play an important role in the pathogenesis of HD, based on the observation that p53 protein is frequently accumulated in Hodgkin and Reed Sternberg (H & RS) cells. We investigated single H & RS cells from five different HD patients for point mutations at the genomic level using multiplex polymerase chain reaction amplification and subsequent sequencing. No point mutations were detected in 50 single H & RS cells analysed. Hence, accumulation of p53 protein cannot be explained by mutations within the gene. A genome-wide screening for genomic imbalances using comparative genomic hybridization revealed gain on chromosome 12q14, i.e. the mapping position of the MDM2 gene in several HD cases. Therefore, we assessed the copy number of the MDM2 gene using fluorescence in situ hybridization. In four out of six HD cases analysed, the copy number of the MDM2 gene was found to be increased. As gene amplification is frequently associated with protein overexpression, the observed accumulation of p53 in the nuclei of H & RS cells could be as a result of elevated MDM2 protein levels resulting in stabilization of p53 protein. PMID- 11260083 TI - Cyclin D1 and E2F-1 immunoreactivity in bone marrow biopsy specimens of multiple myeloma: relationship to proliferative activity, cytogenetic abnormalities and DNA ploidy. AB - Cyclin D1, encoded by the CCND1 gene, is immunohistochemically detectable in up to one-third of cases of multiple myeloma (MM). To examine the mechanism of cyclin D1 overexpression, we compared cyclin D1 immunoreactivity with the results of conventional cytogenetics to determine if the t(11;14)(q13;q32) or other abnormalities of 11q11-14 explained cyclin D1 overexpression. Karyotypic abnormalities were found in 45 out of 67 (67%) MM cases; the t(11;14) was present in seven cases (10%). Additional 11q11-14 abnormalities were not identified. The t(11;14) correlated with cyclin D1 upregulation in low to intermediately proliferative MM, but was not present in highly proliferative tumours (assessed using bromodeoxyuridine labelling index). Cyclin D1 indirectly activates the transcription factor E2F-1. In the bone marrow biopsy specimens of MM cases, E2F 1 was concurrently expressed with cyclin D1 (P = 0.001), indicating that cyclin D1 is functional. However, as neither E2F-1 nor cyclin D1 expression correlated with proliferative activity, the speculation that t(11;14) upregulates the CCND1 gene to induce higher proliferation and possibly more aggressive disease is not supported. We conclude that in low to intermediately proliferative MM cases, cyclin D1 is probably upregulated by t(11;14), but an alternative mechanism is more probable in highly proliferative MM. PMID- 11260084 TI - Arsenic trioxide and ascorbic acid: synergy with potential implications for the treatment of acute myeloid leukaemia? AB - Arsenic trioxide (As2O3) induces remission in a high proportion of patients with acute promyelocytic leukaemia (APL) via induction of apoptosis. Preliminary reports suggest that the apoptotic effect of As2O3 is not specific for APL but can also be observed in non-APL acute myeloid leukaemia (AML) cells, although these are less sensitive than APL cells. Ascorbic acid has recently been demonstrated to enhance the apoptotic effect of As2O3. We have therefore evaluated combined As2O3/ascorbic acid treatment in various clinical samples of AML. Our results indicate a significant synergistic effect of As2O3 and ascorbic acid, suggesting a possible future role of As2O3/ascorbic acid combination therapy in patients with AML. PMID- 11260085 TI - Massive immune haemolysis after allogeneic peripheral blood stem cell transplantation with minor ABO incompatibility. AB - Immune haemolysis as a result of minor ABO incompatibility is an underappreciated complication of haematopoietic transplantation. The increased lymphoid content of peripheral blood stem cell (PBSC) transplants may increase the incidence and severity of this event. We observed massive immune haemolysis in 3 out of 10 consecutive patients undergoing HLA-identical, related-donor PBSC transplants with minor ABO incompatibility. Non-ablative conditioning had been given in 9 of these 10 cases, including two with haemolysis. Cyclosporin alone was used as prophylaxis against graft-vs.-host disease (GVHD). Catastrophic haemolysis of 78% of the circulating red cell mass led to anoxic death in the first case seen, but severe consequences were avoided by early, vigorous donor-compatible red cell transfusions in the subsequent two cases. Haemolysis began 7-11 d after PBSC infusion and all patients with haemolysis had a positive direct antiglobulin test (DAT), with eluate reactivity against the relevant recipient antigen. However, neither the intensity of the DAT, the donor isohaemagglutinin titre, nor other factors could reliably be used to predict the occurrence of haemolysis. Our data indicate that haemolysis may be frequent and severe after transplantation of minor ABO-incompatible PBSCs when utilizing cyclosporin alone to prevent GVHD. Meticulous clinical monitoring and early, vigorous donor-compatible red cell transfusions should be practiced in all instances. PMID- 11260086 TI - Stem cell transplantation from HLA-matched related donor for Fanconi's anaemia: a retrospective review of the multicentric Italian experience on behalf of AIEOP GITMO. AB - Twenty-seven consecutive Italian patients with Fanconi's anaemia (FA) underwent stem cell transplantation (SCT) from an HLA-matched related donor in 10 Italian centres of the Associazione Italiana Ematologia ed Oncologia Pediatrica (AIEOP), Gruppo Italiano di Trapianto di Midollo Osseo (GITMO). Twenty-two patients (81.5%) were conditioned with low-dose (median 20 mg/kg) cyclophosphamide (Cy) and thoraco-abdominal or total body irradiation (median dose 500 cGy), five patients (18.5%) with high-dose Cy (median 120 mg/kg). Graft-vs.-host disease (GVHD) prophylaxis was carried out with cyclosporin A in 26 cases; methotrexate (MTX) was added in eight cases. One patient received MTX alone. The median follow up was 36 months. Ninety-two percent of patients (25 out of 27) engrafted, grade II and III acute GVHD occurred in 28% and 8% of patients, respectively, with chronic GVHD in 12.5%. Conditioning-related toxicity was mild: 4% of patients had grade III mucositis, 7.4% had grade II haemorrhagic cystitis, 14.8% had grade III liver toxicity and 11.1% had grade III renal toxicity. Transplant-related mortality at 12 months was 19.2%, survival at 36 months was 81.5%, with a median Karnofsky score of 100%. No late tumours occurred after a mean follow-up of the survivors of 5 years. None of the studied variables significantly affected the survival, including conditioning regimen, acute GVHD and clinical non haematological phenotype. Among the studied variables, only conditioning regimens containing high-dose Cy and the presence of genital abnormalities were significantly (P < 0.05) associated with an increased rate of acute GVHD. Our study demonstrates that the Italian FA patients undergoing SCT from an HLA matched related donor have a very good outcome. These patients, when compared with others of different ethnic origin who underwent allogeneic bone marrow transplantation, showed a less severe non-haematological phenotype, raising the possibility that this milder phenotype may have, at least in part, contributed to the outcome. Our data may provide a useful tool for further studies aiming to correlate genotype with phenotype. PMID- 11260087 TI - Granulocyte colony-stimulating factor-mobilized peripheral blood CD34+ cells from children contain the same levels of long-term culture-initiating cells producing the same numbers of colony-forming cells as those from adults, but display greater in vitro monocyte/macrophage potential. AB - Autologous peripheral blood progenitor cell (PBPC) transplantation is now commonly used in children. The ontogenic differences in haematopoiesis published in recent years suggest differences in the categories of mobilized PBPCs between children and adults. We investigated the frequency and distribution of mature progenitor cells (colony-forming cells, CFCs) and primitive progenitor cells [CD34+ CD38- and CD34+ Thy-1+ cells, long-term culture-initiating cells (LTC ICs)] in children and adults mobilized using granulocyte colony-stimulating factor alone. We found similar proportions of granulocyte colony-forming units (CFU-G) and/or macrophage CFUs (CFU-M), mixed lineage CFUs (CFU-Mix) and megakarocyte CFUs (CFU-Mk), CD34+ CD38- and CD34+ Thy-1+ cells, and LTC-ICs (16.5 +/- 3.5 vs. 10.65 +/- 5 per 104 CD34+ cells), which produced the same number of CFCs (5 +/- 1 vs. 6 +/- 1 CFCs/LTC-ICs) in PB CD34+ cells from children and adults. However, we noted a higher proportion of erythroid blast-forming units (BFU-E) in PB CD34+ cells from adults (x 1.5, P = 0.003). Using cord blood as a third ageing point, we observed an inverse age-related propensity for commitment to the monocyte/macrophage lineage that was still found after normalizing the data per body weight and processed blood mass. This ontogeny-related programming was detected from the LTC-IC level, which produced 1.7 times more CFU-M in children than in adults (P = 0.048). These subtle differences in commitment between children and adults, shown here for the first time, are of interest for the in vitro manipulation of PBPCs and, in particular, for application in adoptive immunotherapy in children. PMID- 11260088 TI - The impact of attaining a minimal disease state after high-dose melphalan and autologous transplantation for multiple myeloma. AB - Initial studies with high-dose therapy (HDT) in myeloma suggest some beneficial effects of attaining a complete response (CR); however, the effect on survival is difficult to assess owing to inconsistencies in the definition of response between studies. We have analysed 96 newly diagnosed patients aged less than 65 years who received HDT and assessed the effect of response on survival using electrophoresis, immunofixation and fluorescent IgH polymerase chain reaction (PCR) to define CR. Patients received induction chemotherapy with C-VAMP (adriamycin, vincristine, methylprednisolone, cyclophosphamide) followed by melphalan 200 mg/m2 and reinfusion of peripheral blood stem cells. There was a high response to C-VAMP [CR = 24%, partial response (PR) = 64%], with all but one patient improving the depth of response after HDT (CR = 69%, PR = 31%). The progression-free survival (PFS) and overall survival (OS) were excellent at a median of 46.4 months and 72+ months. There was a trend towards an improved PFS in patients with an immunofixation-negative CR compared with patients with a PR (49.4 months, 41.14 months; P = 0.26). This was not evident when electrophoresis was used to define CR. The method used to define CR did not impact on the overall survival and fluorescent IgH PCR failed to add any additional prognostic information. This study supports the widespread use of the European Bone Marrow Transplantation group (EBMT) response criteria and suggests that immunofixation should be performed on all patients who become electrophoresis negative. PMID- 11260090 TI - Relationship between transient abnormal myelopoiesis and acute megakaryoblastic leukaemia in Down's syndrome. PMID- 11260089 TI - Treatment of steroid refractory acute and chronic graft-versus-host disease with daclizumab. AB - Competitive inhibition of interleukin 2-dependent lymphocytes by daclizumab demonstrates some beneficial effects in the treatment of graft-versus-host disease (GVHD). Sixteen patients with steroid refractory GVHD received daclizumab (1 mg/kg BW) on d 1, 2 (-5), 7, 14 and 21. Twelve patients suffered from grade III-IV acute GVHD and four patients from extensive chronic GVHD. Responses were observed in nine patients (six acute, three chronic GVHD). Fourteen out of 16 patients acquired infections during daclizumab treatment and three deaths were infection related. Daclizumab demonstrates limited activity and is associated with an increased incidence of infectious complications. PMID- 11260091 TI - Bone marrow morphology in human immunodeficiency virus-infected South Africans with and without tuberculosis. PMID- 11260092 TI - Leucopenia in professional football players. PMID- 11260094 TI - Sentinel lymph node biopsy in the management of thyroid disease. PMID- 11260095 TI - Varicose veins and pregnancy. PMID- 11260096 TI - Laparoscopy in the staging of pancreatic cancer. AB - BACKGROUND: Over the past decade, laparoscopy has emerged as a popular method of detecting extrapancreatic metastatic disease in patients presumed to have localized pancreatic cancer. METHODS AND RESULTS: The English language literature on laparoscopic staging of pancreatic cancer was reviewed. Interpretation of this literature on staging laparoscopy is difficult because (1) there has been inconsistent use of high-quality computed tomography (CT) in prospective studies, (2) many studies have included patients with locally advanced disease, and (3) the R0/R1/R2 resection rates among patients staged by laparoscopy have not been reported, making it impossible to correlate laparoscopic findings with the R0 resection rate. Laparoscopy may prevent unnecessary laparotomy in a proportion of CT-staged patients presumed to have resectable pancreatic cancer. However, routine laparoscopy is performed on patients judged to have resectable disease by high-quality CT, this fraction of patients is between 4 and 13 per cent. CONCLUSION: When state-of-the-art CT is available, the routine use of staging laparoscopy may not be easily justified from the data in the recent literature. Selective use of laparoscopy may be more appropriate and will probably be a more cost-effective staging approach. Criteria are presented for the selective use of laparoscopy in the staging of patients with localized pancreatic cancer. PMID- 11260097 TI - Preoperative (neoadjuvant) chemoradiotherapy in oesophageal cancer. AB - BACKGROUND: Oesophageal cancer carries a poor prognosis. The 5-year survival rate following resection ranges from 10 to 35 per cent. Recent evidence suggests that the addition of non-surgical treatments to surgery may improve resection rates, reduce the risk of recurrence and improve survival. This review examines the role of preoperative chemoradiotherapy (CRT) in oesophageal cancer. METHODS: A Medline based literature review (1980-2000) was performed using the key words 'neoadjuvant or preoperative' and 'chemoradiotherapy or radiochemotherapy'. Additional literature was obtained from original papers and published meeting abstracts. RESULTS: Forty-six non-randomized and six randomized trials of preoperative CRT were found. Resection rates, pathological complete response (pCR), treatment-related mortality rates and relapse patterns are documented. Improved 5-year survival rates approaching 60 per cent may be achieved following pCR. Three of the six randomized trials show a benefit in either overall survival or disease-free survival compared with surgery alone. Treatment-related toxicity can be significant. CONCLUSION: Preoperative CRT may improve survival. Emerging evidence suggests that CRT alone can achieve similar survival rates to surgery alone. New imaging modalities may help to select which patients require surgery. Larger randomized trials of preoperative CRT or chemotherapy are needed to define optimal regimens and produce higher pCR rates with acceptable toxicity. PMID- 11260098 TI - Randomized clinical trial of local bupivacaine perfusion versus parenteral morphine infusion for pain relief after laparotomy. AB - BACKGROUND: Opioids are often used to decrease pain following laparotomy but are associated with unwanted side-effects. The effectiveness of local perfusion of bupivacaine 0.5 per cent following laparotomy was studied. METHODS: A prospective randomized study involving patients undergoing laparotomy for major colorectal surgery using a left iliac fossa skin crease incision was undertaken. Patients were randomized to receive either intermittent intravenous morphine infusion on demand with patient-controlled analgesia (PCA group) or continuous wound perfusion of local bupivacaine 0.5 per cent for 60 h (LA group). RESULTS: Seventy patients were recruited, 35 in each group. Patient demographics, surgical and recovery variables and complications were comparable in the two groups. The wound lengths were similar (median 14 cm in both groups). There was no statistically significant difference in postoperative pain scores at rest and with movement between the two groups, except for pain scores at rest on the first postoperative day (P = 0.03). The median total amount of morphine used was significantly greater in the PCA group (median 38 versus 0 mg in the LA group; P < 0.001). CONCLUSION: Direct continuous local wound perfusion of bupivacaine 0.5 per cent is as effective as PCA for postoperative pain relief after laparotomy. It is a safe and feasible alternative to parenteral opioids. PMID- 11260099 TI - Stoma-related complications are more frequent after transverse colostomy than loop ileostomy: a prospective randomized clinical trial. AB - BACKGROUND: The consequences of leakage from low colorectal or coloanal anastomoses are reduced by the use of a loop stoma to divert the faecal stream. Controversy continues as to whether loop ileostomy (LI) or loop transverse colostomy (LTC) is the optimal method of defunctioning such anastomoses. METHODS: Patients requiring defunctioning following anterior resection and total mesorectal excision were randomized to receive either LI or LTC. Comparison was made between the groups regarding the difficulty of stoma formation and closure, the recovery after stoma closure and stoma-related complications. The minimum follow-up after stoma closure was 6 months (median 36 months). RESULTS: Between October 1995 and August 1999, 70 patients were randomized (LTC 36, LI 34) of whom 63 underwent stoma closure (LTC 31, LI 32). There were no significant differences in the difficulty of formation or closure, or in the postoperative recovery between the groups. However, there were ten complications related directly to the stoma in the LTC group: faecal fistula (one patient), prolapse (two), parastomal hernia (two) and incisional hernia during follow-up (five). None of these complications occurred in the LI group. CONCLUSION: In this randomized study, the frequency of herniation before or after colostomy closure supports the choice of LI as a method of defunctioning a low anastomosis. Both methods appear to provide satisfactory protection for the low anastomosis. PMID- 11260100 TI - Treatment strategy for patients with middle and lower third bile duct cancer. AB - BACKGROUND: The prognosis for patients with middle and lower third bile duct carcinoma remains poor. This study was conducted to identify independent predictors for survival, as well as the patterns of recurrence after curative resection. METHODS: Sixty-seven patients with pathologically verified middle and/or lower third bile duct carcinoma were analysed retrospectively by Cox regression analysis for predictors of survival. RESULTS: The overall 5-year survival rate after resection was 39 per cent, and 0 per cent for patients who did not undergo resection. The 5-year survival rate was 63 per cent in 26 patients without microscopic residual disease (R0), 16 per cent in 25 patients with microscopic residual tumour (R1) and 0 per cent in six patients with macroscopic residual tumour (R2); ten patients did not undergo resection. Radiotherapy improved the 5-year survival rate in eight patients who had R1 resection compared with the rate in 17 patients who underwent resection alone (8 versus 0), but not significantly so (P = 0.137); however, median survival was significantly longer (P = 0.004) in six patients who had R2 resection compared with that in ten inoperable patients (11.4 versus 3.5 months). Multivariate analysis revealed that the primary tumour and tumour node metastasis (TNM) stage were independent predictors of survival; 13 clinicopathological factors were not independent prognostic factors. Of 26 patients having R0 resection, one had a locoregional relapse only, six had distant metastases only, and five had both types of recurrence. The liver was the most frequent site for metastasis, and microscopic venous invasion (MVI) in the primary tumour was a significant predictor of liver metastasis. CONCLUSION: Curative (R0) resection is only one step in curing cancer, and radiotherapy may play a beneficial role in controlling locoregional residual tumour. MVI could be a useful indicator of when systemic adjuvant therapy should be implemented to prevent liver metastasis after R0 resection, although no effective systemic treatment has yet been developed. PMID- 11260101 TI - Impact of laparoscopic surgery on experimental hepatic metastases. AB - BACKGROUND: Metastatic disease to the liver is one of the major factors determining outcome after colonic resection with curative intention. The influence of laparoscopic surgery on metastatic disease in the liver is still largely unknown. METHODS: An intrasplenic tumour cell inoculation was performed in 30 WAG-Rij rats. After 7 days the rats were randomized into three operative groups: laparotomy (n = 10), laparoscopy with 7 mmHg carbon dioxide pneumoperitoneum (n = 10) and gasless laparoscopy (n = 10). A small bowel segmental resection was carried out in all rats. Some 21 days later the rats were evaluated for number and diameter of tumour nodules and cancer index score at eight different abdominal sites. RESULTS: Hepatic tumour growth scored with the cancer index was significantly reduced in the gasless laparoscopy group compared with that in the carbon dioxide laparoscopy group (P = 0.04) and the laparotomy group (P = 0.02). Tumour growth at the port site and total tumour load were significantly reduced in the gasless group compared with the laparotomy group (P < or = 0.04). CONCLUSION: Laparoscopy with carbon dioxide insufflation seems to stimulate the growth of dormant tumour cells into overt liver metastases. Gasless laparoscopy on the other hand may have a protective effect against metastatic disease in the liver. The promoting and inhibiting effects of laparoscopic procedures on growth of liver metastases need further evaluation. PMID- 11260102 TI - Clinicopathological features and treatment of intraductal papillary mucinous tumour of the pancreas. AB - BACKGROUND: The surgical strategy in patients with a pancreatic intraductal papillary mucinous tumour (IPMT) is still controversial. In this study the pathological findings in a series of patients were used to rationalize surgical choice. METHODS: Fifty-one patients with IPMT were observed between 1988 and 1998 and treated by pancreatic resection. Factors evaluated included symptoms, tumour site, type of operation, histological findings and resection margins, tumour stage, follow-up and survival. RESULTS: Pancreaticoduodenectomy was the most frequent surgical treatment (33 patients; 65 per cent), followed by left pancreatectomy (ten), total pancreatectomy (five) and middle pancreatectomy (three). Histological assessment revealed the tumour to be an adenoma in 13 patients (25 per cent), a borderline tumour in ten (20 per cent) and a carcinoma in 28 (55 per cent), 19 of which were invasive. Mild to moderate dysplasia was present at the resection margin in 20 specimens (41 per cent), and carcinoma in one. Local recurrence was observed in four patients (8 per cent), all of whom underwent a second resection. The 3-year actuarial survival rate for benign and malignant disease was 94 and 69 per cent respectively (P = 0.03). CONCLUSION: These results suggest that resection should be the treatment for IPMT. Management of the resection margin could be crucial in avoiding tumour recurrence. PMID- 11260103 TI - Epidemiology of abdominal aortic aneurysms in the Asian community. AB - BACKGROUND: Studies relating to the ethnic origin of patients with an abdominal aortic aneurysm (AAA) are few and are mainly concerned with the differences between black and white Americans. The purpose of this study was to determine whether the incidence of AAA among the Asian population of Bradford is different from that in the Caucasian population. METHODS: A retrospective study of patients with an AAA was carried out between 1990 and 1997 using data collected by the Patient Administrative Service, personal databases of the vascular consultants and theatre records. Information about the ethnic composition of the population of Bradford was obtained from the 1991 national census. Demographic data, including ethnic origin and clinical details, were obtained from patient notes. RESULTS: Two hundred and thirty-three patients with an AAA were identified during the study interval. The Asian population comprised 14.0 per cent of the total population of Bradford. Twenty-eight AAAs would be expected per year. All of the aneurysms identified occurred in the Caucasian population and none in the Asian community. CONCLUSION: These early results suggest that AAA is rare among the Asian population. PMID- 11260104 TI - Preliminary study of D-dimer as a possible marker of acute bowel ischaemia. AB - BACKGROUND: Occlusion of the superior mesenteric artery (SMA) demands prompt recognition and diagnosis. No accurate diagnostic method is available. The aim of this study was to determine whether the fibrinolytic marker D-dimer is a useful early marker of acute bowel ischaemia. METHODS: Fourteen patients suspected of having acute bowel ischaemia were analysed for an increase in plasma D-dimer level. RESULTS: Six patients had embolic or thrombotic occlusion of the SMA and all had significantly higher D-dimer levels than those without thromboembolic occlusion (P < 0.05). Four patients with strangulation of the small bowel due to adhesions and one with a ruptured aortic aneurysm also had raised D-dimer values. CONCLUSION: In patients with suspected thromboembolic occlusive disease of the SMA, a raised level of D-dimer indicated the presence of acute bowel ischaemia, whatever the cause. A more extensive prospective study is needed to evaluate a potential survival benefit using the test as a marker of the need for urgent laparotomy. PMID- 11260105 TI - Value of intraoperative duplex imaging during supervised carotid endarterectomy. AB - BACKGROUND: For overall benefit, carotid endarterectomy requires low perioperative morbidity and mortality rates. Carotid thrombosis is usually secondary to technical error, which may be related to the experience of the operator. In this retrospective study the clinical and technical outcome of carotid endarterectomies performed by one consultant and five trainees were compared. METHODS: Some 149 patients underwent carotid endarterectomy; 89 were operated on by the consultant and 60 by trainees. Intraoperative duplex imaging of the carotid repair was performed before wound closure, and re-exploration was carried out when there was a residual severe stenosis associated with an intimal flap. RESULTS: There was no significant difference in clinical outcome between operations done by consultant or trainees. There was a significant increase in the number of stenoses, kinks and flaps in carotid endarterectomies performed by trainees compared with those of the consultant both before (chi2 = 12.0, 1 d.f., P < 0.001) and after (chi2 = 10.1, 1 d.f., P < 0.001) correction. CONCLUSION: Intraoperative duplex imaging may facilitate training by providing an objective assessment of the quality of the operation. PMID- 11260106 TI - Nitric oxide prevents intestinal mitochondrial dysfunction induced by surgical stress. AB - BACKGROUND: The intestine is highly susceptible to free radical-induced damage and earlier work has shown that surgical stress induces generation of oxygen free radicals in enterocytes, resulting in intestinal damage along with changes in mitochondrial structure and function. Nitric oxide is an important mediator of gastrointestinal function and this study looked at the effect of nitric oxide on surgical stress-induced intestinal mitochondrial alterations. METHODS: Controls and rats pretreated with the nitric oxide donor L-arginine were subjected to surgical stress by opening the abdominal wall and handling the intestine. Enterocytes were isolated, mitochondria prepared and the protection offered by L arginine against damage due to surgical stress was determined. Protection to structural as well as functional aspects of mitochondria was examined. RESULTS: Mild handling of the intestine affected the enterocyte mitochondrial structure as assessed by lipid composition and electron microscopy. Mitochondria were also functionally impaired with altered calcium flux and decreased respiratory control ratio. Pretreatment with the nitric oxide synthase substrate L-arginine prevented these damaging effects of surgical stress. Protection with arginine was abolished by the nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester, indicating the role of nitric oxide. CONCLUSION: Surgical stress in the small intestine can affect enterocyte mitochondrial structure and function. These damaging effects can be prevented by nitric oxide, an important modulator of cellular function. PMID- 11260107 TI - Outcome and late functional results after anastomotic leakage following mesorectal excision for rectal cancer. AB - BACKGROUND: Few studies have evaluated the long-term functional outcome after anastomotic leakage in the treatment of rectal cancer. METHODS: Between 1993 and 1998, 147 patients were admitted with resectable rectal carcinoma, and 92 underwent low anterior resection (LAR). Seventeen patients (18 per cent) developed clinical anastomotic leakage. The functional outcome of 11 of 12 patients, in whom the stoma was subsequently closed and bowel continuity was restored without stricture, was compared with that of 11 matched patients who had undergone LAR without leakage. Anorectal manovolumetry and symptom scoring on visual analogue scales were done 12-48 months after stoma closure. RESULTS: Nine patients made an uneventful recovery after the initial treatment of anastomotic leakage. Eight developed serious septic complications, four of whom had a pelvic abscess, but there was no death. Five patients had chronic complications that precluded closure of the stoma. Patients who had experienced leakage showed reduced neorectal capacity (120 versus 180 ml; P = 0.04), more evacuation problems (P = 0.02), and a trend towards more faecal urgency (P = 0.09) and incontinence (P = 0.06) than control patients. CONCLUSION: Stoma closure was not possible in five of 17 patients who had experienced anastomotic leakage. Patients who had the stoma closed had impaired long-term anorectal function compared with control patients without leakage. PMID- 11260108 TI - Scintigraphic assessment of colonic transit in women with slow-transit constipation arising de novo and following pelvic surgery or childbirth. AB - BACKGROUND: Colonic transit has not been compared between patients with slow transit constipation (STC) arising de novo (idiopathic) and those whose symptoms followed pelvic surgery or childbirth (acquired). METHODS: In 48 women, with either idiopathic (n = 36) or acquired (n = 12) STC, 111In-radiolabelled diethylene-triamine penta-acetic acid colonic scintigraphy was performed to determine patterns of delay (generalized or left sided), the 'severity' of transit disturbance between subgroups, and the association with age or duration of symptoms. Results were compared with those in healthy women. Patterns of colonic transit disturbance were assessed using previously defined criteria. In those with a generalized delay, variables reflecting the overall rate of isotope progression throughout the colon were calculated: gradient of geometric centre of isotope progression and estimated evacuation time of the isotope. RESULTS: The pattern of transit delay was similar between the subgroups, but the 'severity' of the transit abnormality was significantly worse in those with chronic idiopathic symptoms. In the chronic idiopathic STC subgroup only, there was a significant correlation between both age and duration of symptoms and severity of transit disturbance. CONCLUSION: This study demonstrates that differences in colonic transit exist between subgroups of patients with STC. These might be explained by differences in duration of symptoms or differences in aetiology. PMID- 11260110 TI - Clinical significance of mutator phenotype and chromosome 17p and 18q allelic loss in gastric cancer. AB - BACKGROUND: Tumour stage is the only reliable prognostic factor for gastric cancer. The molecular anomalies involved in this process have the potential to serve as additional prognostic markers. METHODS: Forty-four gastric cancers, treated by surgery alone, have been analysed for chromosome 17p and 18q allelic loss and for the presence of microsatellite instability (MSI), using microsatellite markers and DNA from paraffin-embedded tumours. RESULTS: Eight cancers showed a MSI-positive (MSI+) phenotype. Among the 36 MSI-negative cancers, chromosome 17p and 18q allelic losses were found in 22 of 34 and 19 of 33 informative cases respectively. Multivariate survival analysis indicated MSI status to be an independent prognostic factor along with the tumour stage. MSI+ cancers were associated with longer patient survival, whereas MSI-negative cancers had a significantly poorer prognosis (P = 0.007), with a median actuarial survival of 24 months. CONCLUSION: MSI status is an independent prognostic factor among gastric cancers at the same stage. Chromosome 17p and 18q status added no additional prognostic information to that of tumour stage. The combined use of tumour stage and MSI status may help in deciding whether patients with advanced gastric cancer require additional therapy other than surgery alone. PMID- 11260109 TI - Blockade of growth factor receptors in ductal carcinoma in situ inhibits epithelial proliferation. AB - BACKGROUND: Ductal carcinoma in situ (DCIS) expresses c-erbB-2 receptor and epidermal growth factor receptor (EGFR). The aim of this study was to determine whether blocking of c-erbB-2 receptor with a humanized monoclonal antibody, 4D5 (HerceptinTM), or of EGFR with an epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), ZD1839 (IressaTM), would decrease epithelial proliferation in DCIS. METHODS: DCIS tissue from 18 women undergoing surgery was implanted into 16 to 20 athymic nude mice per experiment (eight xenografts per mouse). Treatment commenced 2 weeks after implantation and consisted either of twice-weekly intraperitoneal injections of 4D5 10 mg/kg or of daily gavage with ZD1839 at 100-200 mg/kg for 14 days; appropriate controls were included. Xenografts were removed on days 14, 21 and 28. Proliferation was assessed by counting 1000 epithelial cells after Ki67 immuno- staining. RESULTS: ZD1839 inhibited proliferation compared with that in controls after 14 days (P < 0.01), whereas 4D5 did not. CONCLUSION: Proliferation in DCIS was decreased by EGFR tyrosine kinase inhibition but not by c-erbB-2 receptor blockade. ZD1839, an orally active and selective EGFR-TKI, has potential as adjuvant therapy in DCIS. PMID- 11260111 TI - Lymph node micrometastasis and prognosis in patients with oesophageal squamous cell carcinoma. AB - BACKGROUND: The purpose of this study was to investigate whether the presence of lymph node micrometastasis in pathological lymph node-negative (pN0) oesophageal squamous cell carcinoma had prognostic value. METHODS: Some 1840 lymph nodes were obtained from 50 patients with pN0 oesophageal squamous cell carcinoma who underwent curative resection of the primary tumour with systematic lymphadenectomy. These lymph nodes were examined immunohistochemically with anticytokeratin antibody (AE1/AE3). Lymph node micrometastases newly detected by immunohistochemistry were classified as micrometastasis. Additionally, lymph node micrometastases were classified into three stages: stage 1, one individual AE1/AE3-positive cell; stage 2, multiple individual positive cells; stage 3, one or multiple positive clusters. RESULTS: Micrometastases were detected in 20 patients (40 per cent). A higher stage of micrometastasis was associated with greater pathological tumour (pT) size (P = 0.023). Recurrent tumours developed in nine patients. However, the frequency of recurrence was similar in patients with, or without, micrometastasis (five of 20 and four of 30 patients respectively; P = 0.25). Twenty-three of 30 patients without micrometastasis survived, whereas 15 of 20 patients with micrometastasis were still alive (5-year overall survival 75 and 78 percent respectively, P = 0.91). Twenty-six of 30 patients without micrometastasis had no recurrence, whereas 15 of 20 patients with micrometastasis had no recurrence (5-year relapse-free survival 86 and 73 per cent respectively, P = 0.37). There was no significant difference in prognosis with respect to the stages of micrometastasis. Multivariate analysis also showed that micrometastasis was not an independent prognostic factor (P = 0.73). CONCLUSION: Immunohistochemical detection of lymph node micrometastasis may be an indicator of lymphatic dissemination of tumour cells. However, the presence of micrometastasis had no impact on the prognosis of node-negative patients with oesophageal squamous cell carcinoma. PMID- 11260112 TI - Gastrointestinal complications in lung transplant survivors that require surgical intervention. AB - BACKGROUND: Lung transplantation is widely accepted as a treatment for end-stage lung disease. At present, information regarding the incidence and outcome of acute gastrointestinal complications in lung transplant survivors is limited. METHODS: Since 1990, 127 lung transplantations have been performed in 125 patients: 73 males (58 per cent) and 52 females (42 per cent) of median age 43 (range 9-64) years. Patients received a standard induction and maintenance regimen of immunosuppression. RESULTS: At a median follow-up of 2.6 (range 0-8.6) years the overall survival rate was 68 per cent. An acute abdomen requiring surgical intervention was diagnosed in 12 patients (10 per cent). The median time following lung transplantation was 19 (range 3-68) months. Eight cases of bowel perforation, two of appendicitis, one of colitis, one of cholecystitis, and one pneumoperitoneum were encountered. Four Hartmann procedures, two sigmoid resections, one small bowel resection, two appendicectomies, a subtotal colectomy, a cholecystectomy and an exploratory laparotomy were performed with minimal morbidity and no postoperative death. CONCLUSION: Lung transplant survivors are at increased risk of developing an acute abdomen because of the use of high-dose immunosuppressive agents. Physicians who evaluate lung transplant patients for an acute abdomen should have a low threshold for surgical intervention. PMID- 11260113 TI - Other primary cancers occurring after treatment of superficial oesophageal cancer. AB - BACKGROUND: Recent advances in endoscopic diagnosis and treatment have led to better prognosis for patients with superficial oesophageal cancer. The incidence of subsequent other primary cancer (SOPC) has become a new problem for patients who survive after treatment of superficial oesophageal cancer. METHODS: Between 1966 and 1998, 368 patients with superficial oesophageal cancer, histologically confirmed as squamous cell carcinoma after resection, were reviewed for the presence of SOPC. RESULTS: Among the 368 patients, 43 developed SOPC. The most frequent sites of SOPC were the stomach (11 patients) and hypopharynx (11). Subsequent cancers of the stomach and hypopharynx developed significantly more frequently in heavy smokers. The 5-year cumulative occurrence rate of subsequent cancers within the fields of endoscopy of the upper gastrointestinal tract (stomach, hypopharynx and residual oesophagus) was 15 per cent. CONCLUSION: Gastric and hypopharyngeal cancers were frequently found after resection of superficial oesophageal cancer. A history of heavy smoking at the time of initial resection may be a risk factor. To make an early diagnosis of subsequent cancers, follow-up observation by upper gastrointestinal endoscopy is important after treatment of oesophageal cancer. PMID- 11260114 TI - Evaluation of the necessity for gastrectomy with lymph node dissection for patients with submucosal invasive gastric cancer. AB - BACKGROUND: When cancer cells are found in the submucosal layer of an endoscopically resected specimen, patients are recommended to undergo gastrectomy with lymph node dissection. If it were possible to identify those patients in whom the risk of lymph node metastasis was negligible, it might be possible to avoid surgery. METHODS: Among those who underwent gastrectomy for gastric cancer from 1980 to 1999, 1091 patients with a cancer invading the submucosa were studied. Clinicopathological factors (sex, age, tumour location, macroscopic type, size, ulceration, histological type, lymphatic-vascular involvement and degree of submucosal penetration) were investigated for their possible association with lymph node metastasis. RESULTS: Lymph node metastases were found in 222 patients (20.3 per cent). Univariate analysis showed that larger tumour size (more than 30 mm), undifferentiated histological type, lymphatic-vascular involvement and massive submucosal penetration had a significant association with lymph node metastasis. Tumour size, histological type and lymphatic-vascular involvement were independent risk factors for lymph node metastasis. By combining these three factors with submucosal penetration of less than 500 microm, 117 patients could be selected as having a minimal risk of lymph node metastasis (95 per cent confidence interval 0-3.1 per cent). CONCLUSION: Lymphadenectomy may not be necessary for patients with gastric cancer invading the submucosa who fulfil the above conditions PMID- 11260115 TI - Local heat-shock priming-induced improvement in microvascular perfusion in osteomyocutaneous flaps is mediated by heat-shock protein 32. AB - BACKGROUND: Stress conditioning is thought to improve microvascular free flap survival but the mechanisms of protection are not clear. The aim of this study was to determine whether local induction of heat-shock protein (HSP) 32 improves microvascular perfusion in transferred osteomyocutaneous flaps. METHODS: The hindlimb harvest region of osteomyocutaneous flaps in Wistar rats was subjected to stress conditioning by local heating (30 min, 42.5 degrees C) 24 h before microvascular flap transfer. In a second group of animals, after heat-shock priming, the action of HSP-32 was inhibited by tin protoporphyrin IX. Animals with unconditioned flaps served as controls. After transfer, the microcirculation of the muscle, cutaneous, subcutaneous and periosteal tissue of the flap was analysed quantitatively for 6 h using intravital fluorescence microscopy. RESULTS: Immunohistochemistry revealed that HSP-32 was detectable only after priming and not in unconditioned flaps. Priming did not alter functional capillary density or capillary red blood cell velocity compared with that in unconditioned flaps. However, heat-shock priming induced significant capillary dilatation (P < 0.05) and thus a substantial increase in capillary blood flow volume (P < 0.05) in all tissues of the transferred flaps. Inhibition of HSP-32 by tin protoporphyrin IX completely abolished the priming-induced improvement in capillary perfusion, as indicated by the lack of increased capillary diameters and volumetric blood flow. CONCLUSION: The present study demonstrated that stress conditioning by local heat-shock priming improves nutritive perfusion in osteomyocutaneous flaps by capillary dilatation, probably mediated through the vasoactive action of HSP-32. PMID- 11260116 TI - Extensive surgical cytoreduction and intraoperative hyperthermic intraperitoneal chemotherapy in patients with pseudomyxoma peritonei. AB - BACKGROUND: Pseudomyxoma peritonei remains a fatal disease. However, extensive surgical cytoreduction combined with intraoperative heated intraperitoneal chemotherapy (HIPEC) has recently emerged as a new treatment modality, which might improve survival. METHODS: Patients underwent treatment if the tumour appeared to be technically resectable on preoperative abdominal computed tomography and there were no distant metastases. After aggressive surgical cytoreduction, HIPEC with the administration of mitomycin C was performed for 90 min. Depending on histological grading, patients received adjuvant 5-fluorouracil and leucovorin therapy. RESULTS: Forty-six patients were treated. Optimal surgical cytoreduction was obtained in 40 patients. Postoperative surgical complications occurred in 18 patients. Four patients died as a direct result of the treatment. Bone marrow suppression due to mitomycin C toxicity occurred in 22 patients. There was no other major toxicity related to the HIPEC procedure. After a median follow-up of 12 months, 40 patients are alive, eight of whom have proven recurrence. The actuarial survival rate (Kaplan-Meier) at 3 years was 81 per cent. CONCLUSION: These results confirm that extensive surgery combined with HIPEC is feasible in patients with pseudomyxoma peritonei and that improved long term survival might be achieved. PMID- 11260117 TI - Role of tumour necrosis factor in lung injury caused by intestinal ischaemia reperfusion. AB - BACKGROUND: Despite the well known inflammatory effects of tumour necrosis factor alpha (TNF), the mechanism of TNF-mediated lung injury following ischaemia reperfusion (I/R) is still unclear. In this study, the role of TNF in the development of acute lung injury following intestinal I/R was investigated. METHODS: Male Wistar rats underwent either sham operation (n = 10), 1 h of superior mesenteric artery occlusion and 2 h of reperfusion (I/R, n = 10), or pretreatment with anti-TNF polyclonal antibody 2 mg/kg and I/R (n = 6). Lung injury was evaluated by Evans blue dye concentration, immunohistochemical staining and morphometric analysis. Intestinal injury was assessed by Evans blue dye concentration and histological examination. RESULTS: Intestinal I/R resulted in lung injury characterized by an increase in Evans blue dye concentration, neutrophil sequestration, and obvious staining for expression of pulmonary CD11b and CD18. Pretreatment of animals with anti-TNF antibody led to a reduction in the sequestration of neutrophils, and a decrease in expression of pulmonary intracellular adhesion molecule 1 and CD18. Anti-TNF antibody pretreatment also reduced the intestinal microvascular injury but not histological grade after intestinal I/R. CONCLUSION: Treatment with an anti-TNF antibody resulted in a significant attenuation of lung injury following intestinal I/R. The data indicate that TNF is an important trigger for upregulation of pulmonary endothelial and neutrophil adhesion molecules after intestinal I/R. PMID- 11260118 TI - Causes of re-recurrence after polytetrafluoroethylene patch saphenoplasty for recurrent varicose veins. PMID- 11260121 TI - Cost-effective carotid endarterectomy. PMID- 11260123 TI - Mesh compared with non-mesh methods of open groin hernia repair: systematic review of randomized controlled trials and laparoscopic compared with open methods of groin hernia repair: systematic review of randomized controlled trials. PMID- 11260135 TI - Molecular mechanisms of Plasmodium falciparum placental adhesion. AB - In natural Plasmodium falciparum infections, parasitized erythrocytes (PEs) circulate in the peripheral blood for a period corresponding roughly to the first part of the erythrocytic life cycle (ring stage). Later, in blood-stage development, parasite-encoded adhesion molecules are inserted into the erythrocyte membrane, preventing the circulation of the PEs. The principal molecule mediating PE adhesion is P. falciparum erythrocyte membrane protein 1 (PfEMP1), encoded by the polymorphic var gene family. The population of parasites is subject to clonal antigenic variation through changes in var expression, and a single PfEMP1 variant is expressed at the PE surface in a mutually exclusive manner. In addition to its role in immune evasion, switches in PfEMP1 expression may be associated with fundamental changes in parasite tissue tropism in malaria patients. A switch from CD36 binding to chondroitin sulphate A (CSA) binding may lead to extensive sequestration of PEs in placenta syncytiotrophoblasts. This is probably a key event in malaria pathogenesis during pregnancy. The CSA-binding phenotype of mature PEs is linked to another distinct adhesive phenotype: the recently described CSA-independent cytoadhesion of ring-stage PEs. Thus, a subpopulation of PEs that sequentially displays these two different phenotypes may bind to an individual endothelial cell or syncytiotrophoblast throughout the asexual blood-stage cycle. This suggests that non-circulating (cryptic) parasite subpopulations are present in malaria patients. PMID- 11260136 TI - Attachment of Neisseria gonorrhoeae to the cellular pilus receptor CD46: identification of domains important for bacterial adherence. AB - Pili of Neisseria gonorrhoeae mediate binding of the bacteria to human host cells. Membrane cofactor protein (MCP or CD46), a human cell-surface protein involved in regulation of complement activation, acts as a cellular pilus receptor. In this work, we examined which domains of CD46 mediate bacterial adherence. The CD46 expression was quantified and characterized in human epithelial cell lines. N. gonorrhoeae showed the highest adherence to ME180 cells, which have BC1 as the dominant phenotype. The BC isoforms of CD46 were expressed in all cell lines tested. The adherence was not enhanced by high expression of other isoforms, showing that the BC domain of CD46 is important in adherence of N. gonorrhoeae to human cells. To characterize the pilus-binding site within the CD46 molecule, a set of CD46-BC1 deletion constructs were transfected into COS-7 cells. Piliated N. gonorrhoeae attached well to CD46-BC1 expressing COS-7 cells. We show that the complement control protein repeat 3 (CCP 3) and the serine-threonine-proline (STP)-rich domain of CD46 are important for efficient adherence to host cells. Further, partial deletion of the cytoplasmic tail of CD46 results in low bacterial binding, indicating that the cytoplasmic tail takes part in the process of establishing a stable interaction between N. gonorrhoeae and host cells. PMID- 11260137 TI - Sphingomyelin trafficking in Chlamydia pneumoniae-infected cells. AB - Chlamydia pneumoniae is a bacterial obligate intracellular parasite with a developmental cycle common to all members of the genus Chlamydia. Like other chlamydiae, the developmental cycle of C. pneumoniae occurs entirely within a membrane-bound intracellular vacuole, termed an inclusion, that is non-fusogenic with endosomal or lysosomal compartments. To characterize the vesicular interactions of the C. pneumoniae inclusion, we used a fluorescent analogue of ceramide, (N-[7-(4-nitrobenzo-2-oxa-1,3-diazole)]-6-aminocaproyl-Derythro sphingosine (C6-NBD-Cer), that has previously been used to characterize the endogenous synthesis and transport of sphingolipids from the Golgi apparatus to Chlamydia trachomatis and Chlamydia psittaci inclusions. Sphingolipids are trafficked to C. pneumoniae inclusions in a time-, temperature- and energy dependent manner with properties very similar to the delivery of sphingomyelin to C. trachomatis inclusions. These results indicate that interactions of the inclusion with a subset of sphingomyelin-containing exocytic vesicles is a property common to all species of chlamydiae. PMID- 11260138 TI - Induction of innate immune responses by Escherichia coli and purified lipopolysaccharide correlate with organ- and cell-specific expression of Toll like receptors within the human urinary tract. AB - Mucosal epithelial linings function as physical barriers against microbes. In addition, they participate in the first line of host defence by production of a variety of proinflammatory mediators when exposed to microbes and microbial agents. Here, we use a human urinary tract infection model to demonstrate that organ- and cell-specific innate responses induced by lipopolysaccharides (LPS) present on Gram-negative bacteria correlates with the expression of Toll-like receptor 4 (TLR4). The presence of TLR4 on human bladder epithelial cells enables them to rapidly respond to bacterial infections in vitro and in vivo. In contrast, TLR4 is not expressed on human proximal tubule cells isolated from the renal cortex, which may explain the cortical localization of bacteria in pyelonephritis. TLR4-negative renal epithelial cells, A498, are non-responsive to purified LPS, however, they respond to viable bacteria via a mannose-sensitive attachment-mediated pathway. To identify LPS components recognised by bladder epithelial cells, a bacterial lipid A mutant and LPS of different chemotypes were tested. Full interleukin 8 induction required hexa-acylated lipid A and was decreased by between 50% and 70% in the presence of O-antigen. Taken together, we propose that multiple independent pathways, which are organ- and cell specifically expressed, mediate bacterial recognition and determine the outcome of innate responses to infection. PMID- 11260139 TI - Essential role of the VirB machinery in the maturation of the Brucella abortus containing vacuole. AB - In epithelial cells, the intracellular pathogen Brucella abortus escapes from the endocytic pathway, exploits the autophagic machinery of the host cell and establishes a unique replication niche in the endoplasmic reticulum. The molecular mechanisms underlying these processes are still poorly understood. Recently, a B. abortus type IV-related secretion system encoded by the virB operon has been described as being involved in the intracellular trafficking of the bacteria. In this study, we have analysed the intracellular pathway of B. abortus virB10 mutant strains by confocal microscopy. We demonstrate that a functional virB operon is essential for the biogenesis of the Brucella-containing vacuole. Polar mutation preventing the transcription of virB10 and downstream sequences did not allow Brucella to bypass the endocytic pathway. Consequently, polar mutant-containing vacuoles fused with lysosomes in which bacteria underwent a degradation process. In contrast, virB10 non-polar mutants were capable of avoiding interactions with the endocytic pathway but, diverging to wild-type Brucella, were unable to reach the endoplasmic reticulum to establish their intracellular replication niche and seemed to be recycled to the cell surface. Based on the two particular phenotypes described in this work, a model of maturation of the Brucella-containing vacuole is proposed. PMID- 11260140 TI - Formation of stress fibres in human endothelial cells infected with Bartonella bacilliformis is associated with altered morphology, impaired migration and defects in cell morphogenesis. AB - Bartonella bacilliformis, a Gram-negative, flagellated bacterium, infects human erythrocytes (haematic phase) and endothelial cells (tissue phase), resulting in a biphasic disease. In the tissue phase of disease (verruga peruana), infection leads to infection of endothelial cells and a pronounced proliferation of these cells, resulting in characteristic skin eruptions of papules and nodules. We have studied the properties of endothelial cells infected in vitro. Extensive cytoskeletal remodelling of endothelial cells occurred after infection in vitro with B. bacilliformis. The cells became spindle shaped and contained arrays of actin stress fibres orientated parallel to the long axis of the cell. Cell-cell contacts were disrupted, along with the distribution of the plasma membrane marker protein, PECAM-1, which participates in cell-cell junctions. The prominent stress fibres terminated in an increased number of focal contacts, which were studied using immunofluorescent staining for paxillin, a cytoplasmic protein that localizes in the focal adhesions. These morphological changes are consistent with activation of intracellular Rho by B. bacilliformis. Formation of stress fibres and the increased number of focal adhesions could be prevented by preincubation of the endothelial cells with C3 exoenzyme, which inactivates intracellular Rho by ADP ribosylation. Endothelial cell motility was greatly diminished in infected cells and the cells did not respond effectively to a stimulus that would evoke motility. In addition, infection of endothelial cells interfered with their ability to form networks of capillary tubes when suspended within three dimensional collagen matrices. If the properties of infected endothelial cells in vivo are similar, the infected cells will probably not participate effectively in angiogenesis. PMID- 11260141 TI - Interaction of Bordetella pertussis with mast cells, modulation of cytokine secretion by pertussis toxin. AB - Together with macrophages and dendritic cells, mast cells have recently been shown to interact with certain pathogenic bacteria and present microbial antigens to the immune system. We show here that Bordetella pertussis can adhere to and be phagocytosed by mast cells. In addition, mast cells are able to process and present B. pertussis antigens to T lymphocytes. Furthermore, exposure of mast cells to B. pertussis induced the release of the proinflammatory cytokines tumour necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6). The release of IL-6 was strongly reduced by pertussis toxin expressed by B. pertussis. The production of IL-10, but not that of IL-4, by mast cells was also inhibited by pertussis toxin. Depletion of mast cells in vivo resulted in significant reduction of early TNF-alpha production in bronchoalveolar lavage (BAL) fluids of B. pertussis infected mice. These data suggest that mast cells may play a role in the induction of immune responses against B. pertussis through the release of cytokines, especially TNF-alpha. PMID- 11260142 TI - Basophil histamine release tests in the diagnosis of allergy and asthma. PMID- 11260143 TI - Eosinophil-epithelial cell interactions: a special relationship? PMID- 11260144 TI - Urinary F2-isoprostane metabolite analysis: a step closer to obtaining a reliable measure of oxidative stress? PMID- 11260145 TI - Bonding and transfer: do epithelial conjugates have a role in chemical asthma? PMID- 11260146 TI - Cytokines and cytokine receptors in allergic rhinitis: how do they relate to the Th2 hypothesis in allergy? PMID- 11260147 TI - Differential mediator release from basophils of allergic and non-allergic asthmatic patients after stimulation with anti-IgE and C5a. AB - The differentiation between allergic and non-allergic asthma is a common and important challenge for the clinician. Until now, no in vitro diagnostic characteristics have been described to distinguish between these types. To examine the diagnostic value of a basophil stimulation test, we compared anti-IgE and C5a-induced mediator release from peripheral blood leucocytes in different types of bronchial asthma. Peripheral blood leucocytes (PBL) from 10 aspirin sensitive asthmatics (ASA), 12 non-allergic asthmatics without aspirin intolerance (NAA), seven allergic asthmatics (AA), and nine healthy controls were prepared by dextran sedimentation. After priming with interleukin-3 (IL-3) PBL were stimulated with anti-IgE and C5a, and the release of histamine (HR) and sulfidoleukotrienes (LTR) in the supernatant was compared. Additionally, purified leucocyte fractions were studied to determine the cellular source of mediator release. Upon stimulation with anti-IgE LTR was slightly, but not significantly, lower in ASA and NAA compared to AA and controls. In contrast, C5a-triggered LTR was significantly higher in ASA (14.4 +/- 12.88 pg/105 cells) and NAA (22.9 +/- 22.61 pg/105 cells) than in AA (9.6 +/- 3.29 pg/105 cells) and controls (7.5 +/- 7.19 pg/105 cells) (P < 0.05). This difference between ASA and NAA vs. AA and controls was even more pronounced when determining the quotient C5a-/anti-IgE induced LTR (P < 0.001). At an optimal cut-off point of 1.0, calculated by relative operating characteristics (ROC) analysis, the positive predictive value for a donor to belong to ASA or NAA was 0.94. No significant differences could be found in HR between the asthmatic patient groups and healthy controls in either condition. As cellular source of LTR and HR the basophil could be determined. Determination of anti-IgE- and C5a-induced LTR from basophils allows us to discriminate between allergic and non-allergic asthmatic patients. For diagnostic purposes the quotient C5a-/anti-IgE-induced LTR is more significant than considering a single parameter. ASA cannot be distinguished from NAA. PMID- 11260148 TI - Alpha-1-antitrypsin is present in the specific granules of human eosinophilic granulocytes. AB - Eosinophils may be found at sites of inflammation, for example in asthma, allergy and helminthic infestation, but their role in human inflammatory disease is unclear. In the present study, we investigated the presence of alpha-1 antitrypsin (AAT), a serine proteinase inhibitor, in human eosinophils. When lysates of highly purified eosinophils were subjected to Western blotting, with a chemiluminescent substrate, immunoreactive bands were seen. An ELISA was developed to measure the AAT content, which was found to be about 100 ng/5 x 106 eosinophils, about 50 ng/5 x 106 neutrophils, and about 25 ng/5 x 106 monocytes. Immunoelectron microscopy showed localization of AAT to the specific granules of eosinophils. During prolonged incubation of eosinophils, no significant increase in the total amount of AAT could be detected by ELISA. However, there was an increased level of AAT in the medium, in parallel with a decrease in the intracellular AAT content, suggesting release of preformed AAT. Apparent complex formation between iodinated elastase and AAT in eosinophil lysates provided evidence that the AAT is functionally active. On the basis of these findings it is suggested that by releasing AAT, eosinophils may, in a microenvironment, play a role in counteracting the tissue damage caused by serine proteinases released by neutrophils in inflammatory conditions. PMID- 11260149 TI - Assessment of oxidant stress in allergic asthma by measurement of the major urinary metabolite of F2-isoprostane, 15-F2t-IsoP (8-iso-PGF2alpha). AB - Asthma is a chronic inflammatory disease of the airways which may involve an oxidant injury to the lung. Assessment of oxidant stress is difficult in vivo, but measurement of F2-isoprostanes (F2-IsoPs), free radical-catalysed products of arachidonic acid, appears to offer a reliable approach for quantitative measurement of oxidative stress status in vivo. We have recently developed a mass spectrometric assay for 2,3-dinor-5,6-dihydro-15-F2t-IsoP (15-F2t-IsoP-M), the major urinary metabolite of the F2-IsoP, 15-F2t-IsoP (8-iso-PGF2a). Measurement of the urinary excretion of this metabolite offers a reliable index of oxidative stress status in vivo that has advantages over measuring unmetabolized F2-IsoPs in urine and plasma. To assess the occurrence of oxidative stress in patients with atopic asthma following allergen exposure in vivo by measuring the urinary excretion of 15-F2t-IsoP-M. Analysis of 15-F2t-IsoP-M by GC-NICI-MS in nine mild atopic asthmatics following inhaled allergen provocation and four asthmatic subjects after inhaled challenge with methacholine. Urinary excretion of 15-F2t IsoP-M increased at 2 h after allergen challenge and remained significantly elevated in all urine collections during the subsequent 8-h period of the study compared to the baseline value (ANOVA, and Student-Newman-Keuls multiple comparisons test). No increase in the urinary excretion of 15-F2t-IsoP-M occurred after inhalation of methacholine. Allergen challenge causes an oxidant injury in human atopic asthmatics. 15-F2t-IsoP-M is a valuable marker of oxidant stress in vivo. PMID- 11260150 TI - NAC Manchester Asthma and Allergy Study (NACMAAS): risk factors for asthma and allergic disorders in adults. AB - Asthma and atopic disorders are the most common chronic diseases in the developed countries. Knowledge of the risk factors for these disorders may facilitate the development of preventive strategies aimed at reducing prevalence rates. To investigate the risk factors for asthma and allergic diseases in a large number of adults who are the parents of children in the National Asthma Campaign Manchester Asthma and Allergy Study. All pregnant women and their partners attending "Booking" antenatal clinics were invited to take part in the study. Questionnaire data were collected including the history of asthma and other atopic diseases, pet ownership and smoking habits, and skin prick tests were performed. The prevalence of atopy and the risk factors for asthma and allergic disorders were investigated in all subjects who completed the questionnaire and underwent skin testing. Statistical analysis was carried out using logistic regression. Initially, risk factors were assessed by univariate analysis to see how each potential explanatory variable affected the probability of having allergic disease. Variables were then tested in a forward stepwise multivariate analysis. In 5687 adult subjects there was a very high (48.2%) prevalence of atopy, and 9.7% of subjects had a diagnosis of asthma. In a multivariate regression analysis sensitization to dust mite, cat, dog and mixed grasses were all independently associated with asthma. The odds ratios for current asthma increased with the increasing number of positive skin tests (any two allergens - OR 4.3, 95% CI 3.3-5.5; any three allergens - OR 7.0 95% CI 5.3-9.3; all four allergens - OR 10.4, 95% CI 7.7-14; P < 0.00001). Dog ownership (OR 1.31, 95% CI 1.10-1.57; P = 0.003) and current smoking (OR 1.36, 95% CI 1.15-1.62; P = 0.0004) were significantly and directly associated with "asthma ever". Thirteen per cent of participants reported a history of eczema. In the multivariate analysis the strongest independent associate of eczema was sensitization to dog (OR 1.37, 95% CI 1.14-1.63, P < 0.0001). Apart from dog, the strength of the association between sensitization to common allergens and eczema appeared to be much lower than in the case of asthma. The prevalence of hay fever was high (20.6%), and in the multivariate analysis the association between sensitization to pollen and hay fever was extremely strong (OR 13.6, 95% CI 11.3-16.3; P < 0.0001). The results of the current study emphasize the importance of sensitization to indoor allergens in asthma. However, evidence of a possible direct role of allergen exposure in asthma causation remains unclear. PMID- 11260151 TI - Benefits of high altitude allergen avoidance in atopic adolescents with moderate to severe asthma, over and above treatment with high dose inhaled steroids. AB - Some patients with severe asthma cannot be controlled with high doses of inhaled steroids (ICS), which may be related to ongoing environmental allergen exposure. We investigated whether 10 weeks of high altitude allergen avoidance leads to sustained benefits regarding clinical and inflammatory markers of disease control in adolescents with persistent asthma despite treatment with high dose ICS. Eighteen atopic asthmatic adolescents (12-18 yr, 500-2000 microg ICS daily) with established house dust mite allergy, participated in a parallel-group study. Quality of life (PAQL), lung function, bronchial hyperresponsiveness (BHR) to adenosine and histamine, induced sputum and urine samples were collected repeatedly from 10 patients during a 10-week admission period to the Swiss Alps (alt. 1560 m) and at 6 weeks after return to sea level. Results were compared with those in eight patients, studied in their home environment at sea level for a similar time period. Throughout the study, asthma medication remained unchanged in both groups. During admission to high altitude, PAQL, lung function, BHR to adenosine and histamine, and urinary levels of eosinophil protein X (U-EPX), leukotriene E4 (U-LTE4) and 9alpha11beta prostaglandin F2 (U-9alpha11beta PGF2) improved significantly (P < 0.05), with a similar tendency for sputum eosinophils (P < 0.07). Furthermore, the changes in PAQL and BHR to adenosine and histamine were greater in the altitude than in the control group (P < 0.05). At 6 weeks after renewed allergen exposure at sea level, the improvements in PAQL (P < 0.05), BHR to adenosine (P < 0.07) and histamine (P < 0.05), as well as U-EPX (P < 0.05) and U-LTE4 (P < 0.05) were maintained. A short period of high altitude allergen avoidance, on top of regular treatment with ICS and long-acting beta2 agonists, results in improvement of asthma, as assessed by clinical and inflammatory markers of disease severity. These findings indicate that short term, rigorous allergen avoidance can improve the long-term control of severe asthma over and above what can be achieved even by high doses of inhaled steroids. PMID- 11260152 TI - Exhaled nitric oxide in seasonal allergic rhinitis: influence of pollen season and therapy. AB - Exhaled nitric oxide (eNO) has been proposed as a potential indirect marker of lower airway inflammation in asthma. To investigate the existence of lower airways inflammation in allergic rhinitis eNO measurements were performed in 32 patients with symptomatic and asymptomatic seasonal allergic rhinitis early in and out of pollen seasons and in 80 healthy volunteers. To further define how exhaled NO is modified by therapy, NO levels were detected following 1-month treatment with either inhaled steroids or non-steroids therapy with nedocromil. Exhaled NO (mean +/- SE) was significantly elevated in patients with seasonal allergic rhinitis with and without symptoms (24.2 + 2.5 and 13.9 + 2.9 ppb, respectively) as compared to healthy volunteers (4.5 + 0.3 ppb) both in and out of pollen season (21.2 + 2.1 and 9.0 + 1.4 p.p.b., respectively) with a higher increase during the allergen exposure in season. Higher levels of exhaled NO were detected in patients with symptoms, either from the upper or lower airways, and with bronchial hyperreactivity. The increased exhaled NO in symptomatic patients was reduced only by inhaled steroids and not by nedocromil. These findings possibly suggest the existence of lower airway inflammation in both symptomatic and asymptomatic patients with seasonal allergic rhinitis in and out of pollen season. Thus, exhaled NO may be used as a non-invasive index for early detection of lower airway inflammation and for monitoring the optional treatment in patients with seasonal allergic rhinitis. PMID- 11260153 TI - Relation between exhaled carbon monoxide levels and clinical severity of asthma. AB - Carbon monoxide (CO) can be detected in exhaled air and is increased in asthmatic patients not treated with corticosteroids. However, it is uncertain whether exhaled CO is related to severity of asthma. To study whether exhaled CO is related to severity of asthma in clinical courses, exhaled CO concentrations were measured on a CO monitor by vital capacity manoeuvre in 20 mild asthmatics treated with inhaled beta2-agonists alone, 20 moderate asthmatics treated with inhaled corticosteroids, and 15 stable asthmatics treated with high dose inhaled corticosteroids and oral corticosteroids once a month over 1 years. Exhaled CO concentrations were also measured in 16 unstable severe asthmatics who visited the hospital every 7 or 14 days for treatment with high dose inhaled corticosteroids and oral corticosteroids. The mean values of exhaled CO in severe asthma over 1 year were 6.7 +/- 9.5 p.p.m. (n = 31, mean +/- SD) and significantly higher than those of non-smoking control subjects (1.2 +/- 0.9 p.p.m., n = 20, P < 0.01). Exhaled CO concentrations in unstable severe asthmatics were significantly higher than those in stable severe asthmatics. However, exhaled CO concentrations in mild and moderate asthmatics did not differ significantly from those in non-smoking control subjects (P > 0.20). There was a significant relationship between the exhaled CO concentrations and forced expiratory volume in one second in all asthmatic patients. These findings suggest that exhaled CO concentrations may relate to the severity of asthma and measurements of exhaled CO concentrations may be a useful means of monitoring airway inflammation in asthma. PMID- 11260154 TI - Diagnostic value of skin-prick and patch tests and serum eosinophil cationic protein and cow's milk-specific IgE in infants with cow's milk allergy. AB - The diagnosis of cow's milk allergy is based on a clinical response to an elimination-challenge test with cow's milk. We studied the usefulness of the skin prick and patch tests and measurement of cow's milk-specific IgE and eosinophil cationic protein in serum as diagnostic tools for cow's milk allergy in a cohort of 6209 unselected infants followed from birth for the development of cow's milk allergy. Of the 239 infants challenged with cow's milk, 118 showed a positive and 121 a negative response at a mean age of 6.9 months. A positive reaction to a skin-prick test with cow's milk (> or = 3 mm) was seen in 72 (61%) and 29 (24%) infants with positive and negative challenges, elevated serum cow's milk-specific IgE (> or = 0.7 kU/L) in 52 (45%) and 15 (13%) infants, a positive reaction to patch test with cow's milk protein fractions in 26 (26%) and eight (8%) infants, and elevated serum eosinophil cationic protein (> or = 20 microg/L) in 22 (21%) and seven (13%) infants, respectively. Parallel use of the four tests with the above-mentioned cut-off values correctly classified 73% of the infants with a sensitivity of 0.76 and a specificity of 0.67. An immediate reaction to cow's milk challenge correlated with skin prick test positivity and elevated serum milk specific IgE, and tended to correlate with patch test positivity. No single test or parallel use of the four tests could predict the challenge outcome acceptably in this prospectively followed, unselected cohort of 6209 infants. A positive reaction to one or more tests needs to be confirmed by a challenge test and a negative response to all four tests does not rule out the possibility of cow's milk allergy. PMID- 11260155 TI - Effect of salmeterol on allergen-induced airway inflammation in mild allergic asthma. AB - A previous study suggested that the long-acting beta2-adrenergic agonist salmeterol (SM) had inhibitory effects on bronchial mucosal inflammation 6 hours after allergen exposure. To further evaluate the influence of SM on allergen induced airway inflammation. We studied, in a randomized, double-blind, cross over trial, 16 mild asthmatic patients who had a dual asthmatic response to allergen inhalation. Subjects received 50 microg of SM or placebo (P), twice daily for 1 week each, separated by a 2-week wash-out period. At the end of each treatment period, after withholding SM for 24 h, they had a methacholine inhalation test (medication was resumed after the test), followed 24 h later by an AC with the concentration of allergen that had induced a LAR at baseline. Airway inflammation was assessed 24 h after the AC by bronchoalveolar lavage (BAL) (n = 16) and bronchial biopsies (n = 13). As expected, SM improved baseline FEV1 and PC20 (P < or = 0.009) and decreased the allergen-induced early bronchoconstrictive response. There were no significant differences in BAL cell counts after the two treatments. On bronchial biopsies, numbers (median, mm2) of submucosal CD45 (P: 43 +/- 23; SM: 161 +/- 43, P = 0.031), CD45Ro (P: 37 +/- 19; SM: 126 +/- 41, P = 0.047) and AA1 positive cells (P: 38 +/- 6, SM: 65 +/- 17, P = 0.006) were significantly higher after SM than P treatment. The numbers of CD4 (P: 11 +/- 10; SM: 32 +/- 7, P = 0.085), HLA-DR (P: 65 +/- 30; SM: 116 +/- 36, P = 0.079) and EG2 positive cells (P: 25 +/- 15; SM: 38 +/- 26, P = 0.09) tended to increase with SM treatment. In summary, compared to placebo, 1 week of regular use of SM is associated with an increase in bronchial inflammatory cells 24 h after AC. This is in keeping with the recommendation that salmeterol should only be used with an anti-inflammatory agent, particularly in the context of significant allergen exposure. PMID- 11260156 TI - Lack of allergic cross-reactivity to cephalosporins among patients allergic to penicillins. AB - There are some contradicting data about clinical allergic cross-reactivity to cephalosporins among patients who have had a previous allergic reaction to penicillins. The purpose of this study was to assess the safety of administering cephalosporins to penicillin-allergic patients. The diagnosis of penicillin allergy was made by positive skin tests to penicillin reagents and/or provocation tests with the penicillin suspected of causing the allergic reaction. To assess the clinical tolerance to cephalosporins, 41 well-characterized penicillin allergic patients diagnosed by positive skin tests and/or provocation tests were challenged with three cephalosporins that do not share the same side chain to the penicillin that induced the reactions: cephazoline, cefuroxime and ceftriaxone. Skin prick and intradermal tests with all cephalosporins tested were negative. All penicillin-allergic patients tolerated therapeutic doses of the three cephalosporins tested (cephazoline, cefuroxime and ceftriaxone) without any ill effect. These results indicate that the risk of suffering from an allergic reaction on administering cephalosporins to penicillin-allergic patients seems to be very low, provided that cephalosporins with a different side chain to the penicillin responsible for the allergic reaction are used. PMID- 11260157 TI - Neutrophils enhance eosinophil migration across monolayers of lung epithelial cells. AB - During the late-phase asthmatic response eosinophils and neutrophils infiltrate the lungs and cause severe damage. In this study, we investigated in vitro the migration of eosinophils, in the absence and presence of neutrophils, across a monolayer of lung H292 epithelial cells. The migration of eosinophils towards the complement fragment 5a (C5a) was increased when neutrophils were added to the upper compartment of the Transwells, and decreased when neutrophils were added to the lower compartment. Moreover, neutrophils exclusively stimulated eosinophil migration towards C5a, and not towards other chemoattractants such as RANTES, IL 8 or PAF. Neutrophils and eosinophils differed in that neutrophils, but not eosinophils, rapidly inactivated C5a, suggesting that neutrophils in the upper compartment remove part of the active C5a that has diffused into the upper compartment. Indeed, we found that the addition of other C5a-degrading agents, such as human serum or carboxypeptidase B, also enhanced eosinophil migration when added to the upper compartment and decreased migration when added to the lower compartment. Taken together, these results indicate that the presence of neutrophils influences the migratory behaviour of eosinophils in vitro. The neutrophils presumably maintain a proper C5a chemotactic gradient in the transmigration model, which results in enhanced eosinophil chemotaxis. PMID- 11260158 TI - The development of an antibody to trimellitic anhydride. AB - Acid anhydrides are a group of highly reactive chemicals used widely in the formulation of paints and plastics. Exposure to acid anhydrides causes several occupational lung diseases such as pneumonititis and asthma. Whilst anhydrides, specifically trimellitic anhydride (TMA), have been shown to bind to lung tissue in an animal model, further investigation has been hampered by the lack of a reagent which would enable the identification of primary target proteins for the binding of TMA. Our objective was to develop an antibody to TMA which would enable in vitro studies of TMA interactions with lung epithelial proteins. We developed a monoclonal antibody which binds solely to TMA. We have demonstrated that the antibody can be used to detect TMA bound to different human proteins with little non-specific binding to unconjugated proteins. We then exposed cells of the A549 lung epithelial cell line to TMA in vitro and have shown by western blotting that binding occurs in the 20-35 Kd weight range. We have developed a specific and sensitive reagent to detect TMA bound to proteins. We have used this to show that when TMA is incubated with a lung epithelial cell line, that the TMA binds to proteins with a restricted molecular weight range. These results suggest that the current paradigm for the detection of IgE to small molecular weight reactive chemicals, which presupposes that the chemical binds to serum albumin, may need further investigation. PMID- 11260160 TI - Rye gamma-70 and gamma-35 secalins and barley gamma-3 hordein cross-react with omega-5 gliadin, a major allergen in wheat-dependent, exercise-induced anaphylaxis. AB - Patients with wheat-dependent, exercise-induced anaphylaxis experience severe allergic reactions when exercising after ingestion of wheat. The major wheat allergen associated with these reactions is a omega-5 gliadin, and patients following a gluten-free diet have remained free of symptoms. The aim of this study was to examine whether allergens cross-reacting with wheat omega-5 gliadin are present in rye, barley and oats. Sera from 23 adult patients with wheat dependent, exercise-induced anaphylaxis were examined. Cereal allergens cross reacting with wheat omega-5 gliadin were identified by immunoblot inhibition. The cross-reactive allergens were purified by gel filtration and reversed-phase chromatography and submitted to amino acid sequencing. Cross-reactivity was further studied by IgE ELISA and ELISA inhibition, and in vivo reactivity by skin prick testing. In immunoblotting rabbit anti-omega-5 gliadin antibodies bound to 70 kDa and 32 kDa proteins in rye and a 34-kDa protein in barley, but not to proteins in oats. N-terminal sequencing identified these proteins as rye gamma-70 secalin, rye gamma- 35 secalin and barley gamma-3 hordein, correspondingly. In ELISA 21/23 (91%) patients with wheat-dependent, exercise-induced anaphylaxis showed IgE antibodies to purified gamma-70 secalin, 19/23 (83%) to gamma-35 secalin and 21/23 (91%) to gamma-3 hordein. In ELISA inhibition omega-5 gliadin inhibited over 90% of the IgE binding of pooled patient sera to solid-phase gamma secalins and gamma-3 hordein. Skin prick testing gave positive reactions to gamma 70 secalin in 10/15 (67%) patients, to gamma-35 secalin in 3/15 (20%) patients and to gamma-3 hordein in 7/15 (47%) patients. The results of this study show that gamma-70 and gamma-35 secalins in rye and gamma-3 hordein in barley cross react with omega-5 gliadin, a major allergen in wheat-dependent, exercise-induced anaphylaxis. These findings suggest that also rye and barley may elicit symptoms in patients with wheat-dependent, exercise-induced anaphylaxis. PMID- 11260159 TI - A monoclonal antibody specific for a carbohydrate epitope recognizes an IgE binding determinant shared by taxonomically unrelated allergenic pollens. AB - Carbohydrate epitopes are capable of binding human IgE from allergic subjects and these epitopes play a role in the cross-reactivity between allergens from unrelated sources. A monoclonal antibody (5E6), specific for a carbohydrate epitope detectable on components of Cupressus arizonica pollen extract, has been produced and characterized. To study the relationship between the epitopes recognized by the monoclonal antibody and by IgE from allergic subjects. To investigate the presence of such carbohydrate IgE determinant in extracts from 21 pollen species belonging to 16 taxonomically related and unrelated families, by means of the monoclonal antibody. IgG-depleted fraction from protein G-purified human allergic serum was obtained. The monoclonal antibody and the IgE from the purified fraction were tested on two glycoproteins, polyamine oxidase and ascorbate oxidase, adsorbed on the ELISA plates. The relationship between the monoclonal- and the IgE-recognized epitopes was investigated by ELISA-competition experiments. Analysis of the distribution of this carbohydrate epitope was performed by direct binding of the monoclonal antibody onto the various extracts. The monoclonal antibody and the IgE were able to bind carbohydrate epitopes on the two plant glycoproteins, ascorbate oxidase and polyamine oxidase. Polyamine oxidase shows only one N-glycosilation site whose carbohydrate moiety seems to be composed of a branched chain of seven ordered sugars, i.e. two N-acetyl-D glucosamine-, three mannose-, one fucose- and one xylose-residues. This structure bears the epitope recognized by mAb 5E6. Human IgE from the IgG-depleted fraction were found capable of inhibiting the monoclonal antibody binding. The allergenic epitope identified was shared by a large number of extracts with different levels of reactivity (OD490 ranging from 0.110 to 2.060). Our data support the finding that a monoclonal antibody specific for a carbohydrate epitope of Cupressus arizonica pollen extract detects an epitope which is also recognized by IgE from allergic subjects. This characterized reagent could be a useful tool for studying distribution of cross-reactive carbohydrate determinants in allergenic pollen extracts and their components. PMID- 11260161 TI - Characterization of increased cough sensitivity after antigen challenge in guinea pigs. AB - Increased sensitivity of cough reflex is a fundamental feature of bronchodilator resistant non-productive cough associated with eosinophilic tracheobronchitis. Our hypothesis is that cough sensitivity is increased by airway allergic reaction characterized by airway eosinophilic inflammation. The aim of this study was to elucidate the hypothesis and clarify the characteristics of the increased cough sensitivity. Number of coughs elicited by inhalation of increasing concentrations of capsaicin (10-8, 10-6 and 10-4 M) was counted 24 h after an aerosolized antigen or saline in actively sensitized or non-sensitized (naive) conscious guinea pigs and then bronchoalveolar lavage was performed. The cough response was also measured 1 day before and 1, 2, 3, 5 and 7 days after an aerosolized antigen challenge in sensitized or naive animals. In addition, effect of procaterol (0.1 mg/kg), atropine (1 or 10 mg/kg), phosphoramidon (2.5 mg/kg) given intraperitoneally 30 min before the capsaicin challenge or capsaicin desensitization on the cough response was examined. Furthermore, the thromboxane A2 (TXA2) receptor antagonist S-1452 in a dose of 0.01 or 0.1 mg/kg or vehicle (saline) was given intraperitoneally at 24 and 1 h before the measurement of cough response. Number of coughs caused by capsaicin was extremely increased 24 h after an antigen challenge in sensitized guinea pigs compared with a saline or an antigen challenge in naive animals or a saline challenge in sensitized animals. The increased cough response disappeared at 3-7 days after the antigen challenge. Eosinophils in bronchoalveolar lavage fluid obtained after the measurement of capsaicin-induced coughs, which was performed 24 h after the antigen challenge, were significantly increased in sensitized guinea pigs. The eosinophil count was significantly correlated to the number of capsaicin-induced coughs. Procaterol or atropine did not alter the antigen-induced increase of cough sensitivity, whereas atropine did reduce the cough response in naive animals. Phosphoramidon increased the number of capsaicin-induced coughs in naive guinea pigs but not in sensitized and antigen-challenged animals. Capsaicin desensitization decreased the cough response in both antigen-challenged sensitized guinea pigs and naive animals. S 1452 reduced the antigen-induced increase of cough response in sensitized guinea pigs, but not in naive animals. Airway allergy accompanied with airway eosinophilia induces transient increase in cough sensitivity, which is not mediated by bronchoconstriction. The increased cough sensitivity may result in part from inactivation of neutral endopeptidase and TXA2, one of the inflammatory mediators. PMID- 11260162 TI - Topical glucocorticoids application induced an augmentation in the expression of IL-1alpha while inhibiting the expression of IL-10 in the epidermis in murine contact hypersensitivity. AB - The repeated application of glucocorticoids (GC) on the skin augmented the inflammatory response of both allergic and irritant contact dermatitis in our studies. In order to further clarify the mechanism of such an augmentation of contact hypersensitivity (CHS), we investigated the modulatory effects of cytokines in the epidermis after the administration of GC at challenged sites in CHS. Diflucortolone valerate was applied to BALB/c mice on alternate days for a total of nine times. On day 12, they were contact sensitized with dinitrofluorobenzene (DNFB). Next, on day 17, one day after the last application of GC, they were challenged with DNFB on the ear. The whole challenged ear lobes were removed after a hapten challenge and then were analysed by the RT-PCR method or underwent an immunohistochemical analysis. To clarify the modulatory effects of cytokines in vivo, DNFB sensitized mice pre-treated with GC were injected with rIL-10, IL-1 receptor antagonist (ra) and anti-IL-1alpha monoclonal antibody (mAb) and thereafter were challenged with DNFB. A RT-PCR analysis has demonstrated IL-10 mRNA to be detected in the challenged skin of non-GC pretreated mice but not in that of GC-pre-treated mice after challenge. On the other hand, the expression of IL-1alpha mRNA in the challenged skin of mice pretreated with GC was more strongly detected that that in mice without GC pretreatment. Furthermore, an immuno-histochemical analysis in the challenge showed the expression of IL-10 in the skin showed the expression of IL-10 in the challenged epidermis of the non-GC-pretreated mice but not in the GC-pretreated mice and IL-1alpha was also strongly expressed in the epidermis of the GC pretreated mice. A subcutaneous injection of anti-IL-1alpha mAb or IL-1 ra inhibited the augmented CHS reaction in the GC-pretreated mice. A subcutaneous injection of rIL-10 also inhibited the augmentation of the CHS reaction in the GC pretreated mice; however, no such inhibition was observed in the non-GC pretreated mice. These results indicated that both an up-regulation of IL-1alpha production and the inhibition of the IL-10 production in the epidermis at the challenged skin sites in the GC-pretreated mice appear to play a critical role in the GC-induced augmentation of murine CHS. PMID- 11260163 TI - Infection of mice with the helminth Strongyloides stercoralis suppresses pulmonary allergic responses to ovalbumin. AB - Asthma and helminth infections induce similar immune responses characterized by the presence of peripheral blood eosinophilia and elevated serum IgE levels. Epidemiological surveys have reported either increases or decreases in the development of atopic diseases and asthma based on the prevalence of helminth infections in the population. The aim of this study was to determine if a pre existing helminth infection would increase or decrease subsequent allergic responses to an unrelated allergen in the lungs. BALB/cByJ mice were infected with the nematode parasite Strongyloides stercoralis prior to ovalbumin (OVA) immunization and intratracheal challenge. Bronchoalveolar lavage (BAL) and fluid (BALF) were collected 3 days post-challenge and cellular and humoral immune responses were measured. Intracellular cytokine staining revealed increased IL-4 and IL-5 producing cells in BAL from mice infected with S. stercoralis before OVA sensitization. Increased IL-5 protein levels and decreased IFN-gamma protein levels were also observed in the BALF. There was, however, no increase in airway eosinophil accumulation in mice infectd with parasites before sensitization with OVA as compared to mice exposed to OVA alone. Furthermore, eotaxin levels in the lungs induced by OVA was suppressed in mice infected with the parasite before OVA sensitization. The development of OVA specific IgE responses in BALF was also impaired in mice infected with the parasite before sensitization with OVA. These results suggest that a pre-existing helminth infection may potentiate a systemic Type 2-type response yet simultaneously suppress in the lungs allergen-specific IgE responses and eotaxin levels in response to subsequent exposure to allergens. PMID- 11260164 TI - Image analysis and quantification in lung tissue. AB - On 9-10 September 1999, an international workshop on image analysis and quantification in lung tissue was held at the Leiden University Medical Center, Leiden, The Netherlands. Participants with expertise in pulmonary and/or pathology research discussed the validity and applicability of techniques used for quantitative examination of inflammatory cell patterns and gene expression in bronchial or parenchymal tissue in studies focusing on asthma and chronic obstructive pulmonary disease (COPD). Differences in techniques for tissue sampling and processing, immunohistochemistry, cell counting and densitometry are hampering the comparison of data between various laboratories. The main goals of the workshop were to make an inventory of the techniques that are currently available for each of these aspects, and in particular to address the validity and unresolved problems of using digital image analysis (DIA) as opposed to manual scoring methods for cell counting and assessment of gene and protein expression. Obviously, tissue sampling and handling, fixation and (immunohistochemical) staining, and microscope settings, are having a large impact on any quantitative analysis. In addition, careful choices will have to be made of the commercially available optical and recording systems as well as the application software in order to optimize quantitative DIA. Finally, it appears to be of equal importance to reach consensus on which histological areas are to be analysed. The current proceedings highlight recent advances and state of the art knowledge on digital image analysis for lung tissue, and summarize the established issues and remaining questions raised during the course of the workshop. PMID- 11260167 TI - Liposuction in cosmetic dermatology. AB - Liposuction is one of the most frequently performed cosmetic procedures in the world today. The central role of dermatologists in evolving many of the procedural methods now used is discussed. Tumescent anaesthesia and the associated tumescent technique proper allow liposuction to be performed safely and effectively in an outpatient setting. An overview of the technique and applications is presented. PMID- 11260168 TI - Kawasaki disease: an update. AB - Kawasaki disease is one of the commonest vasculitides seen in children. It presents with prolonged fever and a polymorphic exanthem. It is a major cause of acquired heart disease in western society. Its exact cause is not known, but exposure to a superantigen has been suggested as a possible aetiological factor. Diagnosis of Kawasaki disease still relies on clinical criteria (Table 1) and investigations are done mainly to exclude other diseases and to detect early or established cardiac complications. Coronary complications can be reduced significantly by the use of intravenous immunoglobulin therapy combined with oral aspirin. The serious consequences of Kawasaki disease require a heightened awareness of this condition when dealing with childhood exanthems. PMID- 11260169 TI - Out patient waiting time for common skin conditions - do general practitioners and dermatologists have the same priorities? A questionnaire-based survey. AB - We present the results of a questionnaire-based survey amongst the general practitioners and consultant dermatologists regarding maximum acceptable waiting times for a range of skin conditions. Statistically significant differences were found in the relative prioritization between the general practitioners and the consultants for a selection of the diagnoses. This has implications for planning and implementation of future services in general and developing waiting list prioritization scoring systems in particular. PMID- 11260170 TI - The management of cutaneous leishmaniasis from Belize. AB - We report 20 patients who contracted cutaneous leishmaniasis in Central and South America, 18 of them in Belize. The diagnosis was confirmed by the polymerase chain reaction (PCR) in 79% of those tested; the corresponding figure for histology was 62%, touch smear 46%, and culture 11%. Results of PCR can be falsely positive, so treatment should not be based on PCR alone. Of the 20 cases 18 were healed 6 weeks after intravenous sodium stibogluconate 20 mg/kg per day for 20 days. We present a management protocol. PMID- 11260171 TI - Oral zinc sulphate in the treatment of acute cutaneous leishmaniasis. AB - A clinical trial to evaluate the efficiency of oral zinc sulphate in the treatment of cutaneous leishmaniasis was conducted. One-hundred and four patients with parasitologically proven cutaneous leishmaniasis were included in the trial. Patients were assigned randomly to receive 2.5, 5 or 10 mg/kg of zinc sulphate orally, and a control group of patients did not receive any treatment. All patients were followed up for 45 days. At the end of the follow-up period, lesions were assessed and parasitological proof of cure or otherwise was sought. Results showed that the cure rate for the 2.5 mg/kg group was 83.9%, for the 5 mg/kg treatment group it was 93.1% and for the 10 mg/kg treatment group it was 96.9%. No lesions in the control group showed any sign of healing during the follow-up period. Therefore, oral zinc sulphate can be recommended as a very safe therapy for cutaneous leishmaniasis. PMID- 11260172 TI - Successful treatment of severe recalcitrant psoriasis with combination infliximab and methotrexate. AB - Plaques of psoriasis contain increased levels of cytokines, including tumour necrosis factor-alpha (TNF-alpha), which are thought to be essential to the maintenance of the psoriatic process. We report the successful treatment of severe, recalcitrant psoriasis when infliximab (a monoclonal antibody to TNF alpha) was used in combination with methotrexate. PMID- 11260173 TI - Low-dose ultraviolet-A1 phototherapy for lichen sclerosus et atrophicus. AB - Lichen sclerosus et atrophicus (LSA) is a chronic inflammatory skin disease characterized by white porcelain-like sclerotic skin lesions. It is most commonly seen in adult females and usually affects the genitoanal area. Extragenital LSA appears in 15-20% of cases. We report a 9-year-old Caucasian girl suffering from extragenital LSA that was resistant to conventional treatment. After 40 treatment sessions with low-dose UVA1 phototherapy, all skin lesions were resolved completely. Moreover, the improvement of skin status has been sustained during 6 months of follow-up. Long-wave UVA irradiation has been shown to induce intensively collagenase activity in human dermal fibroblasts. We suggest that UVA1 irradiation could be an effective treatment in patients suffering from extragenital LSA. PMID- 11260174 TI - Treatment of multiple scalp basal cell carcinomas by photodynamic therapy. AB - The use of surface protoporphyrin IX fluorescence detection to delineate multiple superficial basal cell carcinomas on a patient's scalp is described. Photodynamic therapy (PDT) was subsequently performed with clearance of six and partial clearance of the remaining two tumours. The treated lesions have not recurred during 12 months of follow-up. The opportunity to combine diagnostic fluorescence detection with subsequent treatment by PDT offers an effective and practical management option. PDT is a tissue-sparing modality with low morbidity and good cosmesis, leading us to propose 5-aminolaevulinic acid-PDT for multiple superficial basal cell carcinomas of the scalp. PMID- 11260175 TI - Partial re-emergence of a port-wine stain following successful treatment with flashlamp-pumped dye laser. AB - We present a 49-year-old patient with a congenital superficial vascular malformation of port-wine stain (PWS) type which has partially re-emerged in the 2.5 years since it was almost completely obliterated with the flashlamp-pumped short pulse pulsed-dye laser (FPDL). This observation is discussed with respect to the possible pathogenesis of PWS, with particular reference to the underlying autonomic nerve supply. The latter would not be expected to respond to FPDL and may explain re-emergence of the lesion. PMID- 11260177 TI - Regressive effect of intralesional injection of a moderate dose of recombinant interleukin-2 on carcinoma erysipeloides from gastric carcinoma. AB - Cutaneous metastatic diseases remain nearly incurable and a major medical challenge. It has been shown that interleukin-2 (IL-2) has potential as a therapeutic agent for various neoplastic diseases such as melanoma, renal cell carcinoma and myeloid leukaemia. However, IL-2 therapy for metastatic skin lesions has not been established yet. In the present study, we investigated the effect of recombinant IL-2 in a 79-year-old Japanese man with carcinoma erysipeloides, a rare type of cutaneous metastasis from gastric cancer. He was treated with an intralesional injection of rIL-2 (200 000 JRU) daily. Ten days after treatment, an erythematous plaque was eliminated almost completely leaving light brown pigmentation. A skin biopsy from the pigmented area revealed the absence of obvious tumour cells. These findings suggest that this cytokine should be considered for the clinical treatment of several inoperative metastatic cutaneous diseases, including gastric cancer. PMID- 11260176 TI - Aquagenic pruritus responding to intermittent photochemotherapy. AB - Aquagenic pruritus is a rare but distinct entity in which intense itching develops after contact with water, in the absence of cutaneous signs or underlying disorders that could explain the symptoms. The aetiology is currently unknown, and treatment difficult, with a poor response to antihistamines. Psoralen-ultra violet A (PUVA) photochemotherapy can be effective, but frequent maintenance therapy may be required to prevent relapse of symptoms. We present a patient with typical aquagenic pruritus who responded well to PUVA, and who has remained well controlled on infrequent maintenance. PMID- 11260178 TI - Successful treatment of pustulotic arthro-osteitis (Sonozaki syndrome) with systemic cyclosporin. AB - Pustulotic arthro-osteitis, first described by Sonozaki, is a relatively rare disorder, the prevalence of which is probably underestimated in dermatological literature; its early recognition can prevent misdiagnosis, unnecessary surgery, and the use of prolonged and ineffective antibiotic treatment. PMID- 11260179 TI - Fixed drug eruption due to clarithromycin. AB - Clarithromycin is one of the macrolide antibiotics used for cutaneous and respiratory system infections. Only a few cases of adverse cutaneous reactions to this drug have been reported. Here we report a rare case of clarithromycin induced fixed drug eruption which could be reproduced by a peroral provocation test, whereas patch tests on both unaffected and residual pigmented skin yielded negative results. All cutaneous lesions that recurred due to the challenge test developed the same pigmentation after a short course of intravenous corticosteroid. PMID- 11260180 TI - Localized papular cutaneous schistosomiasis: two cases in travellers. AB - Schistosomiasis is endemic in many parts of the tropics and subtropics with an estimated 200 million people, at least, infected worldwide. The symptoms and signs of vesical and gastrointestinal forms are readily recognized but ectopic forms are rare even in endemic areas and present a greater diagnostic challenge, particularly when they are encountered in nontropical climes. We now report two cases of cutaneous schistosomiasis presenting in Edinburgh with subtle, but remarkably similar, skin lesions. PMID- 11260181 TI - Two sporadic cases of idiopathic multiple minute digitate hyperkeratosis. AB - Multiple minute digitate hyperkeratosis (MMDH) is a skin disease of unknown aetiology characterized clinically by multiple minute asymptomatic keratotic lesions with spiky horny projections. The disorder has been classified into early (congenital) and late (acquired) onset forms, the latter occurring as a presenting sign of concomitant inflammatory, metabolic or malignant disease. Here we report two cases of late onset MMDH without any associated pathology. These cases emphasize that some cases of late-onset MMDH may be idiopathic. PMID- 11260182 TI - Linear cutaneous lupus erythematosus in association with ipsilateral submandibular myoepithelial sialadenitis. AB - We present the case of a young Caucasian female with a several years' history of left-sided submandibular gland sialadenitis in association with linear unilateral cutaneous lupus erythematosus apparently following the lines of Blaschko. The affected gland was excised and the cutaneous lupus erythematosus was improved considerably by oral Dapsone. PMID- 11260183 TI - Hair on a gene string: recent advances in understanding the molecular genetics of hair loss. AB - The hair follicle is finally, after remaining a mystery for many years, beginning to yield some of its molecular secrets. The past decade has seen unprecedented and ever quickening advances in understanding the molecular genetics of the many single gene disorders, which have alopecia as a major feature. This article reviews recent novel clinical and experimental observations, which have shed new light on the basic molecular mechanisms underlying hair morphogenesis, differentiation, keratinization and cycling. We consider recent progress in understanding structural hair defects and complex traits and consider where future developments are likely to occur. PMID- 11260184 TI - Leukaemia inhibitory factor and interleukin-8 expression in nonmelanoma skin cancers. AB - Leukaemia inhibitory factor (LIF) and interleukin (IL)-8 possess activities which may contribute to the development of carcinomas. LIF can stimulate proliferation of some tumour cell lines and IL-8 is angiogenic. Using semiquantitative reverse transcription polymerase chain reaction (RT-PCR), we measured the expression of LIF and IL-8 mRNA in cultured normal keratinocytes (NKC) and the malignant carcinoma cells lines A431, SiHa, HeLa, and in biopsies of basal cell carcinoma (BCC), squamous cell carcinoma (SCC) and normal skin. Protein expression for LIF was assessed by immunohistochemistry in the biopsies. LIF mRNA expression was increased significantly (P < 0.01) in all carcinoma lines, except SiHa, compared with NKC but the IL-8 mRNA expression in carcinoma cell lines was similar to that in NKC. Expression of LIF mRNA was elevated in BCC and SCC compared with normal skin, but a significant difference was observed only between SCC and normal skin (P < 0.01). Both BCC and SCC showed significantly greater expression of IL-8 compared with normal skin (P < 0.01). There was no correlation between LIF and IL 8 mRNA expression either in BCCs or in SCCs. Immunoreactivity for LIF was absent throughout BCC and SCC, however, normal epidermis surrounding the tumour stained positive, as in normal skin. These data may suggest a role for LIF and IL-8 in the development of skin carcinomas, but without co-ordinate regulation of these two cytokines in this process. PMID- 11260185 TI - S100A2 coding sequence polymorphism: characterization and lack of association with psoriasis. AB - Psoriasis is a chronic inflammatory skin disease with a strong genetic component. Linkage studies have identified several susceptibility loci for psoriasis including a region on chromosome 1q21 termed the 'epidermal differentiation complex'. At least 20 genes involved in epidermal differentiation and proliferation have been mapped to this region including S100A2, a gene known to be over-expressed in psoriasis lesions. In the course of cloning and sequencing several S100A2 cDNAs, we identified an A/G (Asn62Ser) polymorphism at nucleotide 185 of the S100A2 coding region. To determine whether this polymorphism is associated with psoriasis, we tested DNA from 38 unrelated normal and 40 unrelated psoriatic individuals. The 185G allele was present in 148 of the 156 chromosomes analysed, giving an allele frequency of 94.9%. All of the remaining chromosomes carried 185A. There was no significant difference in the allele distribution between normal and psoriatic individuals (normals 72G, 4A; psoriatics 76G, 4A; P = 1.00 by Fisher's exact test). Our data explain conflicting results in the literature regarding the sequence of S100A2 but provide no support for a direct causal role for S100A2 in psoriasis. PMID- 11260186 TI - Can we safely diagnose pigmented lesions from stored video images? A diagnostic comparison between clinical examination and stored video images of pigmented lesions removed for histology. AB - Rapid expansion of communication technology has permitted the clinician to perform a consultation with a patient located at a different site. Assuming adequate diagnostic accuracy, it could theoretically be possible to use telemedical techniques as a triage tool. Images of pigmented lesions sent by the primary care physician could be viewed by the consultant dermatologist, and those with banal lesions spared from attending clinic. Previous studies assessing diagnostic accuracy of images of lesions have used 'face-to-face' diagnoses as the 'gold standard'. We set out to compare diagnostic accuracy of image examination compared with that of clinical examination, using histological examination as the diagnostic benchmark. We found that pigmented lesions may be diagnosed as accurately by stored video image evaluation as by conventional clinical examination. None of the malignant skin tumours was misdiagnosed as benign in either group. Whilst these results are encouraging in terms of the clinical safety of store-and-forward imaging, the inability to examine the whole patient or to palpate the lesions may limit the acceptability of the technique severely. Further evaluation of the cost : benefit ratio of such a system to the health care provider must be undertaken before considering this technique as a potential adjunct to managing outpatient referrals. PMID- 11260187 TI - Immunohistochemical analysis of human milk fat globulin 1 and cytokeratin expression in median raphe cyst of the penis. AB - Because median raphe cyst of the penis shares histological findings with apocrine cystadenoma, some cases were thought to be erroneously reported as apocrine cystadenoma of the penis. Further, there is some controversy as to whether the entity, apocrine cystadenoma of the penis, exists or not. Nine cases of median raphe cyst which were clinically unequivocal from their location on the ventral aspects of the penis, were analysed immunohistochemically by using an antibody against human milk fat globulin 1 (HMFG) and a panel of monoclonal anti cytokeratin antibodies. HMFG expression was not observed in eight out of nine cases of median raphe cyst of the penis, and the remaining case showed a faint expression of HMFG focally in its luminal surface, while conventional apocrine cystadenoma in extra-genital area expressed HMFG definitively in our previous study. Our results suggest a possibility that apocrine cystadenoma of the penis is very rare or not present. Therefore, we thought that HMFG expression should be examined in the cases in which apocrine cystadenoma on the penis is reported. PMID- 11260188 TI - Dominant dystrophic epidermolysis bullosa presenting as familial nail dystrophy. AB - Nail dystrophy, a well-recognized feature of dystrophic epidermolysis bullosa (EB), is usually accompanied by skin fragility. We present a three-generation family with an autosomal dominant history of dystrophic nails, but without skin fragility or trauma-induced blisters. No specific diagnosis had been made. However, in the fourth generation, an infant presented with nail dystrophy, acral blistering and milia, raising the possibility of dominant dystrophic EB. This was confirmed by mutational analysis of the type VII collagen gene, COL7A1. We identified a glycine substitution mutation, G1776A, in exon 61 of COL7A1, characteristic of dominant dystrophic EB, which segregated with nail dystrophy in this family. A diagnosis of dominant dystrophic EB should be considered in families with autosomal dominant nail dystrophy even when there is no history of blistering. PMID- 11260189 TI - Three cases of de novo dominant dystrophic epidermolysis bullosa associated with the mutation G2043R in COL7A1. AB - In the absence of a positive family history, it is often difficult to determine whether a single case of mild-to-moderately severe dystrophic epidermolysis bullosa (DEB) represents autosomal recessive or de novo dominant disease. Recent molecular analyses of the type VII collagen gene, COL7A1, have established that the vast majority of such cases are recessive in nature. Nevertheless, a small number of de novo dominant patients have been documented. In this report, we describe three further examples of de novo dominant disease. In each case the COL7A1 mutation comprised the same glycine substitution, G2043R. This mutation has previously been reported in both dominant DEB pedigrees and as a de novo phenomenon and is the most common COL7A1 mutation in dominant DEB throughout the world. These cases emphasize the importance of molecular analysis in providing accurate genetic counselling in this genodermatosis. PMID- 11260190 TI - 5% doxepin cream to treat persistent lichenification in a child. PMID- 11260191 TI - Acne of the scalp-why is it so rare? PMID- 11260192 TI - Rosacea in association with the progesterone-releasing intrauterine contraceptive device. PMID- 11260193 TI - Nail dystrophy in association with polydactyly and benign familial hypercalcemia. PMID- 11260194 TI - Case 1. Diagnosis: neo-natal lupus. PMID- 11260195 TI - Case 2. Diagnosis: erythema multiforme as a presentation of autoimmune progesterone dermatitis. PMID- 11260196 TI - Case 3. Superficial granulomatous pyoderma. PMID- 11260197 TI - Case 4. Diagnosis: Tufted angioma (angioblastoma). PMID- 11260198 TI - A vesico-pustular rash and arthralgia. PMID- 11260200 TI - Folliculitis decalvans. PMID- 11260199 TI - A 62-year-old man presenting with widespread nodulo-ulcerative cutaneous lesions. PMID- 11260201 TI - Follow your nose. PMID- 11260203 TI - Advances in the detection of chromosomal aberrations using spectral karyotyping. AB - Spectral karyotyping (SKY) is a powerful 24-color, whole chromosome-painting assay allowing the visualization of each chromosome in one experiment. Subtle karyotype rearrangements can be detected easily so that small translocations lead to a transition from one color to another at the chromosomal breakpoint region. SKY has enabled the elucidation of several examples of hidden or "cryptic" structural aberrations that may otherwise have been left undetected by classical cytogenetic methods. Furthermore, the chromosomal origins of abnormalities once designated "marker chromosomes" can now be determined rather than left unidentified. SKY analysis of cancer cytogenetics samples provides a much more detailed description of the highly abnormal karyotypes that characterize advanced tumors and cancer cell lines. In addition, SKY significantly adds to the power of clinical cytogenetic analysis of constitutional chromosomal aberrations by facilitating the identification of subtle structural rearrangements that may contain aneuploidy with potential pathological consequences. PMID- 11260204 TI - Fetal cells in maternal blood. AB - Fetal lymphocytes, trophoblasts, and nucleated red blood cells have each been separated from maternal blood by methods such as flow cytometry, magnetic cell sorting, and charge flow separation. The frequency of fetal cells among circulating maternal mononuclear cells remains to be ascertained. Current estimates range from about 10-5 to 10-7, but the numbers may be increased in women carrying aneuploid fetuses. Fetal cells separated from maternal blood have been studied by methods such as polymerase chain reaction and fluorescence in situ hybridization. Among fetal conditions so far identified are sex; human leukocyte antigen and Rh blood types; trisomy 13, 18 and 21; triploidy; and sickle cell anemia and thalassemia. Thus, fetal cell separation might one day be used for screening of the common aneuploidies and, ultimately, for prenatal diagnosis. Individual fetal erythroid precursors have been cultured after separation in some laboratories. Culturing and karyotyping of separated fetal cells might enable diagnosis of a spectrum of chromosomal and genetic disorders. Further development will be required, however, before regular clinical application of these methodologies. PMID- 11260208 TI - Dwarfs in art. PMID- 11260209 TI - Identification of a common variant in the lipoprotein lipase gene in a large Utah kindred ascertained for coronary heart disease: the -93G/D9N variant predisposes to low HDL-C/high triglycerides. AB - Defects in the lipoprotein lipase (LPL) gene are associated with dyslipidemia in the general population. Several rare mutations in the gene, as well as two common coding region polymorphisms, D9N and N291S, exhibit deleterious effects on circulating lipid levels. Using a linkage-based approach, we have identified a large Utah kindred segregating the D9N variant in the LPL gene. The kindred was ascertained for premature coronary heart disease and was expanded based on familial dyslipidemia. A genomic scan identified a region of linkage including LPL, and mutation screening identified the segregating variant. In the kindred, the variant shows high penetrance for a hypoalphalipoproteinemia phenotype, but is also associated with hypertriglyceridemia and elevated insulin levels. The strength of linkage was dependent on the combination of phenotype definition and model parameters, favoring the use of a MOD score approach. Most other studies of LPL have proceeded by mutation screening of randomly chosen individuals or selected affected probands; this is the first example identifying a segregating LPL mutation using direct linkage. PMID- 11260210 TI - Retinal detachment and cataract, facial dysmorphism, generalized osteoporosis, immobile spine and platyspondyly in a consanguinous kindred--a possible new syndrome. AB - We report on a consanguineous family with 6 children (out of 7) affected by a spondylo-ocular syndrome. Clinical features include cataract, loss of vision due to retinal detachment, facial dysmorphism, facial hypotonia, normal height with disproportional short trunk, immobile spine with thorakal kyphosis and reduced lumbal lordosis. On ophthalmological examination of the index patient, a dense cataract and complete retinal detachment could be detected on the right eye. On the left eye, an absent lens nucleus was found, but no retinal detachment. On radiological examination, there was generalized moderate osteoporosis; the spine showed marked platyspondyly and the bone age was advanced. On laboratory investigations, a normal excretion of amino acids, mucopolysaccharides and oligosaccharides could be found. The phenotypical spectrum observed in the 6 affected individuals was rather uniform. The karyotype was normal in all affected children. This hitherto undescribed combination of oculo-skeletal symptoms shows most resemblance with connective tissue disorders, suggesting a range of candidate genes for mutation analysis. PMID- 11260211 TI - Identification of two de novo partial trisomies by comparative genomic hybridization. AB - We report the use of comparative genomic hybridization (CGH) to define the extra chromosome region present in two de novo partial trisomies 15q25-qter and Xp21 pter, which could not be clarified by conventional G-banding. Investigation with fluorescence in situ hybridization (FISH) revealed that the partial trisomy corresponded to an unbalanced translocation between Y and 15 chromosomes in 1 patient and an unbalanced X/X reorganization in the other patient. The combination of classical karyotyping, CGH, and FISH is useful for the identification and characterization of partial trisomies in clinical diagnostic laboratories, in order to delineate the chromosome regions implicated in specific clinical disorders. PMID- 11260212 TI - Late-onset ornithine transcarbamylase deficiency in two families with different mutations in the same codon. AB - We report on late-onset ornithine transcarbamylase (OTC) deficiency in two families with mutations in the same codon, but different base substitutions. Onset of symptoms showed great variation, and five hemizygotes finally died. Clinical diagnosis was late and difficult. In family A, 1 patient also developed the signs of Gilbert's disease. In family B, the index case came to attention as OTC deficiency, after the transplantation of his liver when the recipient died of cerebral edema and hyperammonemia. In family A, the hemizygote males died at the ages of 12 and 18 years; in family B, they died at the ages of 20, 26, and 30 years, respectively. Diagnosis was confirmed by reduced OTC activity in liver specimens. The residual activity in autopsy liver of the index patient in family A was less than the activity in the biopsy of the transplanted liver of the index patient in family B. The molecular investigations showed mutations in exon 2 at codon 40 in the OTC gene in both families. However, different bases were substituted. In family A, the single-base mutation was a cytosine-to-thymine transition (Arg 40 Cys); in family B, it was a guanine-to-adenine transition (Arg 40 His). Published data on in vitro expression studies of the recurrent OTC mutation Arg 40 His have shown little effect on the protein structure of the enzyme. These studies would fit well with our observation of higher OTC activity and later age of onset of symptoms in family B. PMID- 11260213 TI - Brachytelephalangic dwarfism due to the loss of ARSE and SHOX genes resulting from an X;Y translocation. AB - Here we report an 8-year-old male patient who had mesomelic shortening of forearms and legs, brachytelephalangia and ichthyotic skin lesions. Chromosomal analysis showed an X;Y translocation involving the short arm of the X chromosome (Xp). Fluorescence in situ hybridization (FISH) and molecular studies localized the breakpoints on Xp22.3 in the immediate vicinity of the KAL gene demonstrating deletions of steroid sulfatase (STS), arylsulfatase E (ARSE), and short stature homeo box (SHOX) genes. It was suspected that the patient was suffering from chondrodysplasia punctata because of a loss of the arylsulfatase E (ARSE) gene. However, no stippled epiphyses were to be seen in the neonatal radiograph. Interestingly, this patient is the first case with a proven loss of the ARSE gene without chondrodysplasia punctata, assuming that chondrodysplasia punctata is not an obligatory sign of ARSE gene loss. Brachytelephalangia was the only result of ARSE gene deletion in this case. The patient's mother also had dwarfism and showed Madelung deformity of the forearms. She was detected as a carrier of the same aberrant X chromosome. The male patient did not show Madelung deformity, demonstrating that Lerri-Weill syndrome phenotype may be still incomplete in children with SHOX gene deletion. The wide clinical spectrum in the male and the Leri-Weill phenotype in his mother are the results of both a deletion involving several sulfatase genes in Xp22.3 and the SHOX gene located in the pseudoautosomal region. Nevertheless, there is no explanation for the absence of chondrodysplasia punctata despite the total loss of the ARSE gene. Further studies are necessary to investigate genotype/phenotype correlation in cases with translocations or microdeletions on Xp22.3, including the ARSE and the SHOX gene loci. PMID- 11260214 TI - The "flap" endonuclease gene FEN1 is excluded as a candidate gene implicated in the CAG repeat expansion underlying Huntington disease. AB - At least 12 disorders including Huntington disease (HD) are associated with expansion of a trinucleotide repeat (TNR). Factors contributing to the risk of expansion of TNRs and the mechanism of expansion have not been elucidated. Data from Saccharomyces cerevisiae suggest that the flap endonuclease FEN1 plays a role in expansion of repetitive DNA tracts. It has been hypothesized that insufficiency of FEN1 or a mutant FEN1 might contribute to the occurrence of expansion events of long repetitive DNA tracts after polymerase slippage events during lagging strand synthesis. The expression pattern of FEN1 was determined, and ubiquitous tissue expression, including germ cells, suggested that FEN1 has the potential to be involved in HD. Fifteen HD parent/child pairs that demonstrated intergenerational increases in CAG length of greater than 10 repeats were examined for possible mutations or polymorphisms within the FEN1 gene that could underlie the saltatory repeat expansions seen in these individuals. No alterations were observed compared to 50 controls, excluding FEN1 as a trans acting factor underlying TNR expansion. The identification of a candidate gene(s) in HD or other CAG-expansion disorders implicated in TNR instability will elucidate the mechanism of expansion for this growing family of neurological disorders. PMID- 11260215 TI - Recombinant Down syndrome: a case report and literature review. AB - We report a case of a child with features of Down syndrome (DS) but with an atypical karyotype. Initial chromosome analysis was 46,XX,dup(21q).ish 21(wcp21+). The father's chromosomes were normal. However, the mother was found to have mosaicism for a pericentric inversion of chromosome 21 (19/30 cells). The revised chromosome result of the child was 46,XX,rec(21)dup(21q)inv(21)(p12q21.1)mat. A literature review of similar cases (hereafter referred to as rec dup(21q)) was conducted to aid counselling about recurrence risks and the prognosis for this child. All previous reports of rec dup(21q) were secondary to a maternal pericentric inversion. Male carriers did not seem to be at risk of having offspring with the rec dup(21q), although the number of male carriers was limited. In those with rec dup(21q), the risk of congenital heart disease was similar to that of trisomy 21. In reported cases, the facial appearance was suggestive of Down syndrome but perhaps less striking. Although the data are limited, there is an indication the developmental disabilities and short stature are milder in those with rec dup(21q) compared to trisomy 21. These observations promote the concept that the region of chromosome 21 proximal to the duplication contains genetic information contributing to the expression of some features of Down syndrome. PMID- 11260216 TI - Analysis of candidate genes for intrinsic neuropathy in a family with chronic idiopathic intestinal pseudo-obstruction. PMID- 11260217 TI - Four half-siblings with infantile myofibromatosis: a case for autosomal-recessive inheritance. PMID- 11260218 TI - Homozygosity for a 4-bp deletion in a patient with Wolfram syndrome suggesting possible phenotype and genotype correlation. PMID- 11260219 TI - Three-way unbalanced translocation in a mildly dysmorphic mentally retarded child. PMID- 11260220 TI - Multicultural education and genetic counseling. AB - The responsibility to provide accessible, useful genetic counseling to individuals from many cultures and ethnicities arises from the increasing ethnocultural diversity of the populations served, coupled with the ethical goal of providing equal access and quality of services for all individuals. The multicultural education, training, and practice of genetic counseling involves three major components: knowledge of relevant ethnocultural groups, ethnocultural self-awareness, and an understanding of institutional and social barriers to services. Despite the diversity of ethnocultural groups served and the critical role of direct experience and training for the genetic counselor, some general guidelines for multicultural genetic counseling can be identified. These include the importance of establishing and maintaining trust, the essential need to respect the counselee's healthcare beliefs and practices, and the necessity of understanding the impact of culture on the process of decision making and on counselee responses to nondirective counseling. PMID- 11260224 TI - The role of genomic imprinting in human developmental disorders: lessons from Prader-Willi syndrome. AB - Normal human development involves a delicate interplay of gene expression in specific tissues at narrow windows of time. Temporally and spatially regulated gene expression is controlled both by gene-specific factors and chromatin specific factors. Genomic imprinting is the expression of specific genes primarily from only one allele at particular times during development, and is one mechanism implicated in the intricate control of gene expression. Two human genetic disorders, Prader-Willi syndrome (PWS, MIM 176270) and Angelman syndrome (AS, MIM 105830), result from rearrangements of chromosome 15q11-q13, an imprinted region of the human genome. Despite their rarity, disorders such as PWS and AS can give focused insight into the role of genomic imprinting and imprinted genes in human development. PMID- 11260225 TI - Shakespeare as a geneticist. AB - There are frequent allusions in Shakespeare's plays to physical and mental characteristics and qualities of individuals that often reflect genetic and related insights, ideas and implications with a bearing on the determination of these phenotypes. Quotations from the plays with comments are presented here to indicate varied perceptions in these regards that Shakespeare brilliantly conveyed some 400 years ago through many of his 'dramatis personae'. PMID- 11260227 TI - Identification of the Arg1086His mutation in the alpha subunit of the voltage dependent calcium channel (CACNA1S) in a North American family with malignant hyperthermia. AB - Individuals from a large North American population were screened for the presence of the mutation in the alpha1 subunit of the voltage-dependent calcium channel (CACNA1S) that has recently been associated with malignant hyperthermia (MH). This Arg1086His mutation was screened for in 154 MH normal (MHN) individuals and 112 MH susceptible (MHS) individuals, who were diagnosed by the North American protocol of the in vitro contracture test. PCR and restriction enzyme analysis was used to test for the mutation. The Arg1086His mutation in the CACNA1S was not found in any of the MHN individuals. In contrast, two related individuals (grandfather and grandson, father and son of the MH proband) among the MHS group exhibited this mutation. However, a third MHS individual in the same family (granddaughter, cousin of the grandson) did not exhibit this mutation. These results indicate that this mutation may be associated with MH in this family. Genetic alterations in the CACNA1S associated with MH are present in approximately 1% of this North American MHS population. PMID- 11260226 TI - Familial aggregation of QT-interval variability in a general population: results from the NHLBI Family Heart Study. AB - QT-interval prolongation is associated with increased risk of cardiac death. Although information on genetics and molecular mechanisms of the congenital long QT syndrome is mounting, limited data are available on the genetics of QT interval in the general population. Heart rate adjusted QT intervals (Bazett's QTc, and QT index (QTI)) were assessed by electrocardiography in 2399 members aged 25-91 years of 468 randomly selected families participating in the NHLBI Family Heart Study. Familial correlation and segregation analyses were performed to evaluate the genetics of the variability of QT interval in this population. The parent-offspring (0.14+/-0.03) and sibling (0.18+/-0.03) correlations for age and sex-adjusted QTc were moderate, while the spouse correlation was close to zero (0.09+/-0.06). This suggests that there are familial/genetic influences on QT-interval variability. Segregation analysis results suggest that there is a major effect in addition to heritable multifactorial effects (h2=0.34), but the major effect did not follow Mendelian inheritance. Further adjustments of QTc for other major cardiovascular risk factors did not significantly change the results. Similar results were found for QTI. The QT-interval variation in the general population is influenced by moderate heritable multifactorial effects in addition to a major effect. A major gene effect is not directly supported. PMID- 11260228 TI - Novel molecular defects in the androgen receptor gene of Mexican patients with androgen insensitivity. AB - The androgen insensitivity syndrome (AIS) is an X-linked form of male pseudohermaphroditism caused by mutations in the androgen receptor (AR) gene. In the present study, we analyzed the AR gene in 8 patients, 4 sporadic and 2 familial cases with the syndrome, using exon-specific polymerase chain reaction, single-stranded conformational polymorphism and sequencing analysis and identified six new single base mutations, including one nonsense mutation at the hinge region of the receptor. These molecular lesions occurred in the steroid binding domain (SBD) and all but one affected the first nucleotide of their respective codons. A nonsense mutation in exon 4, which converts a glutamine into a premature termination signal (Q657stop), a missense mutation changing arginine instead of glycine (G743R) and a conservative substitution of leucine with valine at amino acid 830 (L830V) were detected in patients with CAIS. Three other missense mutations located in exons 4 (L701I), 5 (A765S), and 6 (Q802R) were present in individuals bearing a partial form of AIS. These data allow us to reaffirm the view that nonsense mutations in the AR results almost invariably in a CAIS phenotype and underly the importance of the SBD for the AR functional activity. PMID- 11260229 TI - Mutational screening of the cationic trypsinogen gene in a large cohort of subjects with idiopathic chronic pancreatitis. AB - Several missense mutations, including R122H, N29I, K23R, A16V and D22G, in the cationic trypsinogen gene (PRSS1), have been associated with certain forms of hereditary pancreatitis (HP). Their occurrence in the idiopathic chronic pancreatitis (ICP) and whether novel mutations could be identified in PRSS1 remain to be further evaluated. These were addressed by the mutational screening of the entire coding sequence and the intronic/exonic boundaries of the PRSS1 gene in 221 ICP subjects, using a previously established denaturing gradient gel electrophoresis technique. Among the known PRSS1 mutations, only the R122H was detected in a single subject and the A16V in two subjects in the cohort, strengthening that HP-associated PRSS1 mutations are rare in ICP. Additional missense mutations, including P36R, E79K, G83E, K92N and V123M, were identified once separately. By analogy with the known PRSS1 mutations, predisposition to pancreatitis by some of them, particularly the V123M autolysis cleavage site mutation, is suspected. Functional analysis is expected to clarify their possible medical consequences. PMID- 11260231 TI - Evidence for locus heterogeneity in the Camurati-Engelmann (DPD1) Syndrome. PMID- 11260230 TI - Identification of novel C253Y missense and Y864X nonsense mutations in the insulin receptor gene in type A insulin-resistant patients. AB - Mutations in the insulin receptor gene have been reported in cases of type A insulin resistance. We report herein two cases of type A insulin resistance, which involve some novel mutations. Case 1 is a heterozygote of the C253Y missense mutation and case 2 is a heterozygote of the Y864X nonsense mutation. In the C253Y missense mutation in exon 3, a cysteine residue is replaced with tyrosine in the cysteine-rich domain of the alpha subunit. The Y864X in exon 13 results in a truncated receptor, which is devoid of most of the beta subunit. This mutant receptor could not be expressed on a cell membrane since the transmembrane domain is missing. Other significant mutations were not found for the entire coding regions and splice/donor sites. PMID- 11260232 TI - The use of intragenic polymorphisms in determination of the genomic relevance of whole-exon deletions in MLH1 and MSH2. PMID- 11260233 TI - Cenani-Lenz syndrome with renal hypoplasia is not linked to FORMIN or GREMLIN. PMID- 11260234 TI - Skin occlusion and irritant and allergic contact dermatitis: an overview. AB - Occlusion, widely used to enhance percutaneous absorption of drugs, also increases penetration of other chemicals and antigens, and hence may exacerbate irritant and allergic contact dermatitis. This overview summarizes the adverse effects of occlusion. PMID- 11260235 TI - Identification and allergenic activity of hydroxyaldehydes - a new type of oxidation product from an ethoxylated non-ionic surfactant. AB - Ethoxylated alcohols, which are widely used as surfactants, are susceptible to oxidation on air exposure. A complex mixture of oxidation products is formed, among which alkylated aldehydes, alkylated formates, formaldehyde and peroxides have previously been identified by our group. In the present study, we have identified a new class of oxidation product from the nonionic ethoxylated surfactant C12E5. These oxidation products are highly water-soluble hydroxyaldehydes with the general formula HO(CH2CH2O)nCH2CHO, n=1-4. To facilitate the identification of the hydroxyaldehydes in oxidized C12E5, reference compounds were synthesized. The sensitizing potential of 1 of the identified hydroxyaldehydes, HO(CH2CH2O)3CH2CHO, was studied in guinea pigs and was found to be weak. A significant cross-reactivity between this aldehyde and the next shorter homologue, HO(CH2CH2O)2CH2CHO, was observed. Irritant components, present in the oxidation mixture, facilitate the skin penetration of allergens, which further accentuates the importance of controlling the conditions of storage and handling of ethoxylated surfactants, to reduce the formation of allergenic and irritant oxidation products. PMID- 11260236 TI - Occupational allergic contact dermatitis caused by wood dusts. AB - Exposure to wood dusts may cause various skin and mucosal symptoms. Allergic dermatoses, caused by wood dusts, diagnosed at the Finnish Institute of Occupational Health during 1976-1999 are reported here. 16 had allergic contact dermatitis and, 2 had contact urticaria. 9 men (3 cabinet makers, 3 joiners, 1 carpenter, 1 knifemaker and 1 machinist) were mainly exposed to tropical hardwoods. 1 man had dermatitis caused by western red cedar. 5 patients, 3 men and 2 women, were exposed to Finnish pine or spruce dusts, and 1 man to aspen. 7 also had rhinitis, 4 asthma or dyspnoea and 3 conjunctivitis. On patch testing, 10 men reacted to 9 different wood dusts, including teak (5), palisander (3), jacaranda (2), mahogany (2), walnut (2) and obeche (1). Reactions to wood allergens, including lapachol (2), deoxylapachol (1), (R)-3,4 dimethoxydahlbergione (2), 2,6-dimethoxy-1,4-benzoquinone (1), mansonone A (2) and salicyl alcohol (1), were noted in 4 cases. All but 1 of 5 patients exposed to pine or spruce dusts reacted to the sawdusts, all 5 to colophonium, 3 to abietic acid, 2 to tall oil resin, 3 to wood tar mix and 4 to other wood gum resins. Of the 2 CU patients, 1 was prick and RAST positive to obeche, 1 reacted with urticarial dermatitis to punah wood dust on chamber exposure. Occupational allergic dermatoses are mainly caused by the dusts of hardwoods, mostly due to Type IV allergy, but may also be caused by softwood dusts. Patch tests can be done with wood dusts, but should be confirmed by patch testing with wood allergens if possible. PMID- 11260237 TI - Prevalence of nickel allergy among Finnish university students in 1995. AB - Nickel allergy was studied in a sample of 1st-year university students starting their studies in 1995. A total of 296 subjects (72%) of 413 invited participated in the clinical examination, and 284, 96 male and 188 female, were patch tested (69%). A history of nickel sensitization was enquired for. Prick tests and serum specific IgE levels were determined. Occurrence of atopic dermatitis, hand eczema, and current exposure to metals were recorded. Nickel allergy was encountered in 39% of all female students, in 42% of females with pierced skin, and in 14% of females without pierced skin. The corresponding figures for males were 3%, 7% and 3%. In the multiple regression analysis, the risk factors for nickel allergy were female sex (OR 8.1, p<0.01), current metal exposure at examination (OR 4.1, p<0.01) and skin piercing (OR 3.6, p<0.05). Positive prick tests or elevated IgE levels to common allergens were not significantly associated with nickel allergy. In female students, the prevalence of nickel allergy has increased from 13% in 1986 to 39%. The prevalence among males has remained low at 3%. The results indicate that, in addition to skin piercing, current metal contacts are important risk factors for nickel allergy. This finding gives support to the EU Nickel Directive. PMID- 11260238 TI - A cream containing the chelator DTPA (diethylenetriaminepenta-acetic acid) can prevent contact allergic reactions to metals. AB - Chelating agents in protective barrier creams have often been used in the prevention of allergic contact dermatitis to nickel. In a pilot study, we demonstrated the preventive effect of 10% diethylenetriaminepentaacetic acid (DTPA) in an oil-in-water emulsion in nickel-sensitized patients. Now we reproduced these results in a randomized, double-blind study. Additionally, we investigated the efficacy of the barrier cream in other clinically relevant metal allergies. Individuals sensitized to various metals had a significant decrease in positive patch test reactions after pre-treatment with the DTPA-cream: 2.5% nickel sulfate (24/28 positive without pre-treatment versus 1/28 with pre treatment; p<0.0001), 5% nickel sulfate (30/32 versus 15/32; p=0.0003), 1% cobalt chloride (19/20 versus 6/20; p=0.001) and 5% copper sulfate (13/14 versus 5/14; p=0.02). However, the cream had no protective effect with 1% palladium chloride (17/23 versus 16/23) and with 0.5% potassium dichromate (9/13 versus 7/13). We conclude that the DTPA-cream clearly abrogates positive patch test reactions in nickel-, cobalt- and copper-sensitized subjects and that it may therefore be helpful in the management of allergic contact dermatitis. PMID- 11260239 TI - Effect of different moisturizers on SLS-irritated human skin. AB - Moisturizers are widely used to treat irritant contact dermatitis (ICD). Their use is, however, not well-documented and standardized models for testing skin care products are needed to acquire documentation of their efficacy. The present study was undertaken to evaluate the effect of 6 commonly-used moisturizers on the recovery of irritated human skin. No commercial interests were involved in the study. 36 healthy volunteers had patch tests with SLS 0.5% applied on their forearms/upper arms for 24 h. After irritation of the skin, all volunteers had a moisturizer applied on one forearm/upper arm, respectively, 3 x daily for the following 5 days. The other forearm/upper arm served as an untreated control. Each moisturizer was tested on 12 volunteers and each volunteer tested 2 moisturizers at the same time. Evaluation was done on days 1, 3 and 8 by transepidermal water loss, electrical capacitance, laser Doppler flowmetry, DermaSpectrometry and clinical scoring. All 6 moisturizers were found to accelerate regeneration of the skin barrier function when compared to irritated non-treated skin. The most lipid-rich moisturizers improved barrier restoration more rapidly than the less lipid-rich moisturizers. We suggest this experimental model for further moisturizer efficacy testing. PMID- 11260240 TI - Occupational airborne contact dermatitis caused by thyme dust. AB - The aim of the study was to assess occupational hazards to the farmer's skin associated with processing thyme (Thymus vulgaris L.). 46 farmers were studied during the threshing of dried thyme. They were questioned about work-related skin problems and examined before and after work. In all persons, serum thyme-specific IgE was measured. Skin prick tests, the Ouchterlony test and the migration inhibition test were carried out with allergens of airborne bacteria and fungi present in the working environment. Of the 46 farmers studied, 4 showed skin symptoms after 5-30 min of exposure to thyme dust. Thyme-specific IgE was found in 1 person with work-related symptoms, but also in 2 asymptomatic farmers. Therefore, the importance of IgE seems to be questionable in eczema related to thyme dust. Skin and blood tests with microbial allergens also showed no significant differences between the symptomatic and asymptomatic farmers. To our knowledge, this is the 1st description of occupational airborne contact dermatitis caused by thyme dust. The etiology of thyme-related skin symptoms remains obscure, although an irritant mechanism seems probable. PMID- 11260241 TI - Pathological findings in cumulative irritation induced by SLS and croton oil in hairless mice. AB - It is known that the pathological features of acute irritant contact dermatitis are specific according to the irritant. However, in chronic irritant contact dermatitis, it is not clear whether specific patterns exist. To investigate whether the specific pathology of acute irritant contact dermatitis is sustained in chronic irritant contact dermatitis, sodium lauryl sulfate (SLS) and croton oil were applied 3x a week for 2 weeks on the dorsal skin of hairless mice using Finn Chambers. The pathologic changes induced by irritants at various concentrations were evaluated using H&E and Luna's staining, as well as immunohistochemistry for 5-bromo-2-deoxyuridine (BrdU), keratin 6 and loricrin. Our results showed that epidermal hyperplasia and inflammatory infiltration were relatively marked in the groups treated with higher concentrations of irritants. These features were more prominent in the 1% croton oil treated group than in the 0.25% SLS treated group. However, lower concentrations of irritants resulted in very similar histological changes, characterized by epidermal hyperplasia with minimal inflammatory infiltration, irrespective of the chemical. Our results suggest that the histological responses to irritants vary with concentration in cumulative irritation, as in acute irritation, but repetitive mild irritation may evoke common histological changes, characterized by epidermal hyperplasia with minimal inflammatory infiltration, irrespective of the chemical used. PMID- 11260242 TI - Occupational allergic contact dermatitis from methylchloroisothiazolinone and methylisothiazolinone (MCI/MI) in a silicone-emulsion lock lubricant. PMID- 11260243 TI - False-negative patch test with levobunolol. PMID- 11260244 TI - Changes in the ultrastructure of cytoskeleton and nuclear matrix during HaCaT keratinocyte differentiation. AB - The cellular scaffold that comprises nuclear matrix and cytoskeleton provides mechanical support for the cell and plays a crucial role in motility, cellular signaling, regulation of gene transcription and DNA replication. In this study we examined the structure of cytoskeleton and nuclear matrix in the keratinocyte cell line HaCaT using a recently developed technique, embedment-free electron microscopy. With this method the three-dimensional structure of cellular scaffold is visualized in the cells extracted from soluble proteins and the chromatin. In actively proliferating cells the cytoskeleton appeared to consist of a continuous meshwork of 10--15 nm filaments with a smaller amount of thin (5 nm) and ultrathin (1--2 nm) filaments. In contrast to what could be expected from earlier immunofluorescence and electron microscopy studies, the cytoskeleton in HaCaT keratinocytes did not consist of superposed autonomous networks of different filaments but was a highly integrated, continuous structure filling whole cytoplasmic territory. Moreover, cytoskeletons of adjacent cells were in a direct physical contact. Nuclear matrix consisted of globular ribonucleoprotein aggregates attached to the meshwork of 20--40 nm filaments. Nuclear envelope was firmly fastened to the cytoskeleton. In keratinocytes induced to differentiation by calcium switch both the cytoskeleton and nuclear matrix were drastically rearranged and comprised a monomorphic, dense and regular meshwork of 10--15 nm filaments. Our data underscore the fact that in HaCaT keratinocyte monolayer in vitro, and probably also in the epidermis in vivo, the nuclear matrices and the cytoskeletons of adjacent cells seemed to form a continuous, highly ordered structure which is rapidly rearranged during cell differentiation. This feature may be crucial for the understanding of how the signal initiated by, e.g. mechanical forces generated through the cell--cell and cell--matrix interaction is transmitted to the nucleus producing ultimately changes in the pattern of gene expression. PMID- 11260245 TI - Serum-free cultured keratinocytes fail to organize fibronectin matrix and possess different distribution of beta-1 integrins. AB - The development of serum free medium formulation for culturing keratinocytes was a breakthrough in achieving a high number of epidermal cells for experimental and therapeutic studies, in particular to support the wound healing process. It is not clear, however, if switching the cells to highly proliferative phenotype may reflect change in other cellular functions important for the wound repair as their adhesive interactions with the extracellular matrix components. Remodelling of the extracellular matrix, particularly of fibronectin plays an essential role for guiding the cells during wound healing. The molecular mechanisms for organization of this provisional fibronectin matrix, however, are still not clear. We found that keratinocytes in serum containing medium, although in fewer numbers than fibroblasts, were able to remove adsorbed fluorescent labelled fibronectin from the substratum and reorganize it in a fibrilar pattern along the cell periphery. After 3 days the secreted fibronectin had also been organized as matrix-like fibers and as clusters deposited on the substratum after migrating cells. In contrast, serum free cultured keratinocytes fail to organize pre adsorbed fluorescent labelled fibronectin, as well as the secreted fibronectin, although they grow very well under these conditions. Switching the cells to serum containing medium initiates the removal of fluorescent labelled fibronectin from the substratum, however without reorganization in fibrillar pattern. Most likely, these keratinocytes remove fluorescent labelled fibronectin by the expression of proteolytic activity, rather than with the mechanical function of beta(1) integrins. The latter were diffusely dispersed in serum containing conditions and tend to organize in focal adhesions in serum free cultured cells. We assumed their transient expression and different affinity state might be important for the keratinocyte migration and matrix assembly mechanism. PMID- 11260246 TI - Characterization of the desmosomal cadherin gene family: genomic organization of two desmoglein genes on human chromosome 18q12. AB - The human desmoglein genes, desmogleins 1--3, are members of the desmosomal cadherin superfamily, and encode critical components of the desmosome. These genes are tightly clustered within 150--200 kb of chromosome 18q12.1 and represent excellent candidate genes for genetic disorders of the epidermis linked to this region of the genome. Mutations in desmoglein 1 have already been implicated in the genetic disorder striate palmoplantar keratoderma. Similarly, a mutation in desmoglein 3 underlies the balding mouse phenotype, although no human mutations in desmoglein 3 have been identified to date. In this study, we have characterized the genomic organization of two of the three desmoglein genes mapped to chromosome 18q12. Comparison of their exon-intron structure reveals the high level of evolutionary conservation expected from these related genes. The identification of the genomic structure of the desmoglein genes will facilitate mutation detection in genodermatoses with desmosomal abnormalities resulting from underlying defects in these genes. PMID- 11260247 TI - Structural analysis reflects the evolutionary relationship between the human desmocollin gene family members. AB - Desmocollins, members of the desmosomal cadherin family, are known to play an important role in desmosomal intercellular adhesion. The human desmosomal cadherin cluster is located on chromosome 18q12, and consists of three desmoglein and three desmocollin genes. The cDNAs of all six of these genes have been cloned and sequenced, however, the exon-intron organization was reported for only one human desmocollin gene, DSC2. We elucidated the exon-intron structures of the DSC1 and DSC3 genes using PCR amplification of genomic DNA and direct sequencing of BAC clones. The results suggest a strong evolutionary conservation between the genomic organization of the desmocollin genes. PMID- 11260248 TI - Lectin and proteoglycan histochemistry of Merkel cell carcinomas. AB - Changes in carbohydrate residue expression and in proteoglycan distribution occur during different stages of tumor development and progression. However, few data concerning carbohydrate residue analysis as performed by lectin histochemistry and proteoglycan distribution of Merkel cell carcinoma, a rare malignant tumor of the skin, have been reported. Hence, lectin- and proteoglycan immunohistochemistry was performed on paraffin wax material of 9 cases of Merkel cell carcinomas characterized by cytokeratin and neurofilament immunohistochemistry. The lectin binding pattern of tumor cells varied between lectins with different sugar binding specificities, while within a given nominal sugar specificity intensities were remarkably similar between tumors from different patients. The most intensive reaction was observed using Con A (mannose/glucose-specific) followed by LCA with the same specificity and the N Acetyl glucosamine-specific lectins (WGA, UDA, CMA), while no fucose binding sites were detected (UEA-I). In addition, N-Acetyl galactosamine residues were only occasionally detected. The lectin binding pattern of Merkel cell carcinoma cells indicated that predominantly N-linked glycans and not O-linked glycans, typical for mucins of most epithelia, were present. Hence these tumor cells were relatively undifferentiated and resembled stem cells more closely than differentiated epithelia. The tumor stroma was especially evaluated in this study and showed a lectin reaction, which was intermediate between the tumor cells and extra-tumoral stroma. For example, the reactions of N-Acetyl galactosamine specific lectins were intensive in the extra-tumoral stroma but nearly negative in tumor cells, while the lectin reaction of the intra-tumoral stroma was similar to the cellular reaction. These results indicated an influence of tumor cells on the stromal constituents. Antibodies against chondroitin type glycosaminoglycans reacted with the tumor stroma and the pericellular substance around the tumor cells most intensely in - and around the major tumor septae which, in general, were well vascularized. The most intensive immunoreactivity was detected using the chondroitin-6-sulfate antibody. The cellular and membrane-associated reaction for heparan sulfate was less intensive in comparison to epidermal cells. In conclusion the pattern of lectin-binding sites, the high chondroitin(sulfate) specific reactivity and the relatively low intensity of heparan sulfate immunohistochemistry indicate a low degree of differentiation and high malignity of the tumors, which is consistent with the clinical behavior of Merkel cell carcinomas. PMID- 11260249 TI - Doxepin affects acetylcholine induced cutaneous reactions in atopic eczema. AB - BACKGROUND: Atopic eczema (AE) is a chronic inflammatory skin disease with strong itching as the prominent symptom. The pathology of itch is still in discussion, but acetylcholine (ACH) seems to be a relevant pruritogenic mediator in AE. Since efficient benefit on pruritus and excoriations has been demonstrated with tricyclic agents, we investigated how the topical treatment with doxepin (5%, Boehringer Standard, Mannheim, Germany), a tricyclic compound with anticholinergic properties, may influence ACH induced itch and cutaneous sensations (erythema, wheal, axonreflex flare). METHODS: Eleven patients with AE were included in this double blind study. For 3 days we applied doxepin cream to a defined area on the volar forearm and basic ointment to the other side 4 times daily. On day 4, ACH and sodium chloride were i.c. injected into the pretreated arms. Vasoreactions and cutaneous sensations were measured similar to studies described in previous publications from our group. RESULTS: Doxepin treatment over 3 days reduced ACH provoked flare size more than 53% (P<0.005) and wheal size about 48% (P<0.005) whereas the maximal antipruritic effect was similiar to the basic therapy. The itch intensity, which is expressed as the mean AUC value, was rated at 6.12 arbitrary units after the neutral cream application and 5.9 arbitrary units after doxepin. CONCLUSIONS: The clinical and experimental effectiveness of doxepin as an antipruritic drug has been known for years. However, studies focusing on ACH as a pruritogenic mediator have not been performed. The duration of the doxepin application in our study seems to be appropriate since flare and wheal development were diminished. The reason why doxepin did not develop more antipruritic action compared to the vehicle cream may be due to the fact that the doxepin free cream already possessed an antipruritic action in this experimental study design. This is probably caused by rehydrating and moisturizing effects. PMID- 11260250 TI - Microdialysis of histamine in the skin of patients with mastocytosis. AB - Mastocytosis represents a group of disorders characterized by the proliferation and accumulation of mast cells in tissue. The aim of the present study was to examine whether the interstitial histamine concentration in the skin is increased in mastocytosis patients and whether it correlates with the number of mast cells, the amount of metabolite N-methyl-imidazole acetic acid in the urine and the tryptase in serum. In 7 mastocytosis patients on a standardized diet, the analysis of histamine was performed on microdialysates obtained from catheters positioned intracutaneously in involved and uninvolved skin. N-methyl-imidazole acetic acid in the urine was collected for 24 h. Biopsies for analyses of mast cells were taken from skin adjacent to the microdialysis catheters. The histamine concentrations were 42+/-14, 12+/-3 (P<0.05) and 8+/-2 nmol/l (mean+/-SEM, n=7) in skin eruptions, non-lesional skin and plasma respectively. Mean N-methyl imidazole acetic acid in the urine (9.7+/-3.5 mmol/mol creatinine) and mean tryptase (124+/-54 microg/l) had increased in all patients. In the present study, no linear correlation was found between these parameters and interstitial histamine in lesional skin. This finding corresponds to the fact that the concentration of histamine metabolites and tryptase derives from the entire mast cell population, while interstitial histamine in the dermis represents the local tissue concentration before metabolic transformation. The microdialysis of histamine in the skin of mastocytosis patients could be used as a tool to investigate the effects of dermal mast-cell histamine release in different kinds of treatment regimen. PMID- 11260251 TI - The feasibility of quantitative analysis of androgen metabolism by use of single dermal papillae from human hair follicles. AB - Androgenetic alopecia (AGA) is a dihydrotestosterone-mediated process, characterized by continuous miniaturization of androgen sensitive hair follicles (HF). Although increased 5 alpha-reductase (5aR) activity in affected HF is a key feature in the pathogenesis of AGA, only little is known about the in vivo expression of 5aR within AGA-affected HF. Recent studies have shown that the dermal papilla (DP) is the predominant site of type 2 5aR expression within the human HF, but direct measurements of 5aR activity in intact DP of AGA-affected HF have not been reported so far, mainly because of technical problems. Hence there is a need for a reliable and sensitive method of measuring 5aR activity in fresh tissues. As a novel approach, we used freshly isolated, intact DP and a highly sensitive HPLC-radiomatic flow scintillation system to measure 5aR. In this way we were able to measure 5aR even in small DPs from miniaturized HF. Our results show that DP from the occipital scalp express ex vivo considerable amounts of 5aR activity, but the measurable enzyme activities of individual DP differ considerably. Therefore the use of only one or two DP is at present not a reliable tool to analyze 5aR activity ex vivo. PMID- 11260252 TI - What are the most promising strategies for the therapeutic immunomodulation of allergic diseases? AB - Specific immunotherapy and other immunomodulatory strategies have long been a stronghold in the management of allergic diseases. In particular, "immunodeviation-therapy" or "vaccination for allergies", i.e. the redirection of Th2-type immune responses towards a Th1-response pattern, has become an ever more popular concept. The present feature of CONTROVERSIES complements our previous discussion of atopy (Rocken et al., Exp Dermatol 7: 97--104, 1998), and is dedicated to a critical analysis of the general problems and limitations one faces with the main immunomodulatory strategies traditionally considered in this context. We also explore alternative approaches that appear promising in order to achieve both a more effective and/or a more specific immunotherapy of allergic diseases. Given that the mast cell remains a key protagonist in the pathogenesis of allergic diseases finally, this feature examines how innovative, more selectively mast cell-targeted strategies may be developed for the management of allergic diseases. PMID- 11260253 TI - Histone acetylases--versatile players. AB - Genomic DNA in eukaryotes is tightly packed in the form of a highly ordered chromatin structure. In view of this tight packing, one of the most important questions in biology is how the transcriptional machinery regulates target genes in chromatin. Reversible modification of histones by acetylation is involved in transcriptional activation as well as repression in chromatin contexts. Recent studies with highly purified histone acetylases have provided insights into the mechanisms whereby acetylases contribute to transcriptional control. Furthermore, they suggest the possibility that histone acetylases could play roles in various forms of DNA metabolism as well as in transcription in chromatin contexts. PMID- 11260254 TI - Influenza virus RNA polymerase PA subunit is a novel serine protease with Ser624 at the active site. AB - BACKGROUND: Influenza virus RNA polymerase is a multifunctional enzyme that catalyses both transcription and replication of the RNA genome. The function of the influenza virus RNA polymerase PA subunit in viral replication is poorly understood, although the enzyme is known to be required for cRNA --> vRNA synthesis. The protease related activity of PA has been discussed ever since protease-inducing activity was demonstrated in transfection experiments. RESULTS: PA protein was highly purified from insect cells infected with the recombinant baculovirus carrying PA cDNA, and a novel chymotrypsin-type serine protease activity was identified with the synthetic peptide, Suc-LLVY-MCA, in the PA protein. [3H]DFP was crosslinked with PA and a mutational analysis revealed that serine624 was as an active site for the protease activity. CONCLUSIONS: These results constitute the demonstration of protease activity in PA subunit of the influenza virus RNA polymerase complexes. PMID- 11260255 TI - Sez4 gene encoding an elongation subunit of DNA polymerase zeta is required for normal embryogenesis. AB - BACKGROUND: Sez4 identified as a seizure-activated gene shows a similarity to the yeast REV3 that encodes a catalytic subunit of the nonessential DNA polymerase zeta which is involved in error-prone translesion synthesis. Although yeast REV3 homologues in mouse and human have recently been identified and characterized, their precise roles remain elusive. RESULTS: Here we investigated the role of mouse pol zeta by targeted inactivation of the Sez4 gene. The homozygous Sez4 mutants died around embryonic day (E) 10.5. This lethal effect was the result of developmental defects and apoptotic cell death within the embryo proper at the gastrulation stage, and it was partially rescued at E12.5 by the expression of a Sez4-transgene. In wild-type embryos, Sez4 transcripts were up-regulated within the embryo proper from E7.5, correlating well with the lethal stage of Sez4 inactivation. CONCLUSION: Our findings indicate that Sez4 is essential for epiblast lineage-specific development and suggests a requirement of mammalian DNA polymerase zeta in the survival of certain subcellular populations which are indispensable to normal embryogenesis. PMID- 11260256 TI - Human homologues of the Caenorhabditis elegans cell polarity protein PAR6 as an adaptor that links the small GTPases Rac and Cdc42 to atypical protein kinase C. AB - BACKGROUND: Asymmetric cell division in the Caenorhabditis elegans embryos requires products of par (partitioning defective) genes 1-6 and atypical protein kinase C (aPKC), whereas Cdc42 and Rac, members of the Rho family GTPases, play an essential role in cell polarity establishment in yeast and mammalian cells. However, little is known about a link between PAR proteins and the GTPases in cell polarization. RESULTS: Here we have cloned cDNAs for three human homologues of PAR6, designated PAR6alpha, beta and gamma, comprising 345, 372 and 376 amino acids, respectively. The PAR6 proteins harbour a PDZ domain and a CRIB-like motif, and directly interact with GTP-bound Rac and Cdc42 via this motif and with the aPKC isoforms PKCiota/lambda and PKCzeta via the N-terminal head-to-head association. These interactions are not mutually exclusive, thereby allowing the PAR6 proteins to form a ternary complex with the GTPases and aPKC, both in vitro and in vivo. When PAR6 and aPKC are expressed with a constitutively active form of Rac in HeLa or COS-7 cells, these proteins co-localize to membrane ruffles, which are known to occur at the leading edge of polarized cells during cell movement. CONCLUSION: Human PAR6 homologues most likely play an important role in the cell polarization of mammalian cells, by functioning as an adaptor protein that links activated Rac and Cdc42 to aPKC signalling. PMID- 11260257 TI - Selenocysteine codons decrease polysome association on endogenous selenoprotein mRNAs. AB - BACKGROUND: Selenocysteine incorporation has been reported to be inefficient in all systems studied, including Escherichia coli, baculovirus-insect cell systems, rabbit reticulocyte in vitro translation systems, transiently transfected mammalian cells, and intact animals. Nonetheless, full-length selenoproteins containing up to 17 selenocysteine residues are produced in animals, indicating that the efficiency observed in manipulated systems might not accurately reflect the true efficiency of this process in nature. RESULTS: To begin to address this apparent discrepancy, we have examined the polysome profiles of endogenously expressed selenoprotein mRNAs in a mammalian cell line, and compared them with nonselenoprotein mRNAs. We report that three selenoprotein mRNAs, type 1 deiodinase, glutathione peroxidase and selenoprotein P, are under-loaded with ribosomes, based on their predicted open reading frame sizes. The average numbers of ribosomes per mRNA correspond to the sizes predicted by termination at the UGA selenocysteine codons. Appropriate loading on the type 1 deiodinase mRNA is seen following substitution of a cysteine codon for the selenocysteine codon, indicating that the UGA codon confers a translational penalty on the mRNA. Surprisingly, ribosomal loading is also increased by the expression of eukaryotic release factors eRF1 and eRF3. CONCLUSIONS: These results suggest that the presence of a selenocysteine codon confers a translational penalty on selenoprotein mRNAs, and that increased levels of release factors may alter the kinetics of termination. PMID- 11260258 TI - A serine/threonine kinase p90rsk1 phosphorylates the anti-proliferative protein Tob. AB - BACKGROUND: tob is a member of a gene family with anti-proliferative function. Over-expression of Tob in NIH3T3 cells results in the suppression of cell proliferation. The growth suppression is hampered by the presence of activated ErbB2 kinase. The molecular mechanisms by which Tob suppresses cell growth and by which ErbB2 abrogates Tob function remain to be elucidated. RESULTS: We show that Tob is phosphorylated on serines and threonines, but not tyrosines, by a kinase(s) that associates with Tob in the lysates of various cells, including ErbB2-over-expressed cells. We also show that a 95 kDa kinase associates with Tob in vitro. The autophosphorylation activity of this kinase co-chromatographes with Tob-phosphorylating activity, suggesting that the 95 kDa kinase phosphorylates Tob. Among the known kinases with molecular mass around 95 kDa, p90rsk1 associates with Tob in vitro and in vivo, and phosphorylates Tob at least in vitro. Therefore, it is likely that p90rsk1 represents the 95 kDa kinase and is involved in the regulation of Tob function through phosphorylation. CONCLUSION: p90rsk1 associates with and phosphorylates Tob. Because p90rsk1 is activated downstream of receptor tyrosine kinases, we propose that Tob function is at least in part under the control of growth factor-stimulated tyrosine kinases through its phosphorylation by p90rsk1. PMID- 11260260 TI - Somatic mutations of synaptic cadherin (CNR family) transcripts in the nervous system. AB - BACKGROUND: Cadherin-related neuronal receptor (CNR) family genes have been identified in the nervous system by screening molecules bound to Fyn-tyrosine kinase. The CNR family is comprised of diverse synaptic cadherins. The genomic organization of the CNR genes, composed of variable and constant regions, is similar to that of the immunoglobulin gene cluster. The nervous system is characterized by the acquisition of diverse function. This feature is similar to the immune system. In the immune system, the generation and selection of immunoglobulin gene mutants is the underlying basis for acquired immunity. We therefore examined somatic regulation of the CNR family genes in the nervous system to determine whether a similar mechanism controls nervous system development. RESULTS: We first demonstrated that approximately 10% of the CNR3 transcripts were trans-transcripts between the variable and constant exons at the adult stage. Furthermore, somatic mutations of CNR3 transcripts accumulated during brain development, with a marked bias for A-to-G and U-to-C transitions. Approximately 70% of the CNR3 transcripts exhibited a mutation characterized by the addition of a CU-repeat; these transcripts contained seven continuous CU repeats in the 3' untranslated region at the adult stage, whereas the CNR genomic DNA contained six continuous CU-repeats. Interestingly, a high rate of replacement mutations was detected in the first cadherin domain that functions in acquisition of specificity for cell adhesion in cadherins, while in the other regions the occurrence of silent mutations increased during development. CONCLUSION: The present report is the first description of the generation and possible selection of somatic mutations of synaptic cadherin transcripts. The somatic alterations of CNR family transcripts and their synaptic localization have interesting implications for the molecular basis underlying the establishment of somatic rearrangement of neuronal networks. PMID- 11260259 TI - Identification of Tetrahymena hsp60 as a 14-nm filament protein/citrate synthase binding protein and its possible involvement in the oral apparatus formation. AB - BACKGROUND: Tetrahymena 14-nm filament protein (14FP) is bifunctional, with roles as a citrate synthase in mitochondria and as a cytoskeletal protein in nuclear events during fertilization and in oral morphogenesis. In this study, to further our understanding of the bifunctional property of 14FP, we attempted to screen 14FP-binding proteins using affinity column chromatography. RESULTS: Through the screening of 14FP-binding proteins using 14FP-affinity chromatography, we detected 65 kDa and 70 kDa proteins that bound to 14FP in an ATP dependent manner. From the N-terminal amino acid sequence, these proteins were identified as the Tetrahymena mitochondrial chaperones, hsp60 and mthsp70, respectively. Tetrahymena hsp60 was recognized with a monoclonal antibody raised against human hsp60. Immunofluorescence and immunoelectron microscopy using the monoclonal antibody showed that Tetrahymena hsp60 was localized to mitochondria. Moreover, Tetrahymena hsp60 was also present at extramitochondrial sites including basal bodies of cilia and oral apparatus, and particularly at the developing oral apparatus during cell division. CONCLUSION: These results suggest that Tetrahymena hsp60 is localized in basal bodies and is involved in cortical patterning such as the formation of the oral apparatus as well as having a role in the folding of mitochondrial proteins in mitochondria. PMID- 11260261 TI - TBP-interacting protein TIP120A is a new global transcription activator with bipartite functional domains. AB - BACKGROUND: We previously identified a TBP (TATA-binding protein)-interacting protein 120A (TIP120A) from rat liver nuclear extracts. TIP120A is thought to be a unique global transcription factor that can interact with TBP and can stimulate all classes of eukaryotic transcription. RESULTS: We produced various truncation proteins of TIP120A to delineate its functional domains. TIP120A binds to TBP in the acidic amino acid-rich N-terminal region and in the leucine-rich C-terminal region. These regions exhibited an ability to stimulate basal transcription in vitro. In addition, these two regions overlap with domains that facilitate nonspecific DNA-binding of RNA polymerase II. The sequences of these two regions are significantly conserved among TIP120A homologues of eukaryotes. CONCLUSIONS: TIP120A is a bipartite transcription factor, and both N-terminal and C-terminal regions exhibit TBP-binding activity and stimulate the basal transcription ability. PMID- 11260262 TI - Hes7: a bHLH-type repressor gene regulated by Notch and expressed in the presomitic mesoderm. AB - BACKGROUND: Whereas Notch signalling is essential for somitogenesis, mice deficient for the basic helix-loop-helix (bHLH) genes Hes1 and Hes5, downstream Notch effectors, display normal somite formation, indicating that there may be an as-yet unidentified Hes1-related bHLH gene. RESULTS: We identified a novel bHLH gene, designated Hes7, from mouse embryos. Hes7 has a conserved bHLH domain in the amino-terminal region and the WRPW domain at the carboxy-terminal end, like Hes1. The mouse Hes7 gene is located next to Aloxe3, which is mapped to a position 37.0 cM from the centromere on chromosome 11. In a transfection analysis, Hes7 represses transcription from the N box- and E box-containing promoters. In addition, Hes7 suppresses the E47-induced transcriptional activation. Promoter analysis indicated that Hes7 expression is controlled by Notch signalling. Strikingly, Hes7 is specifically expressed in the presomitic mesoderm in a dynamic manner. We also identified two related bHLH genes from human: one is closely related to mouse Hes7 and therefore designated hHes7 and the other designated hHes4. CONCLUSION: The structure, transcriptional activity and expression pattern in the presomitic mesoderm of Hes7 are very similar to those of Hes1, suggesting that Hes7, together with Hes1, may play a role in somite formation under the control of Notch signalling. PMID- 11260263 TI - Identification and functional analysis of the gene for type I myosin in fission yeast. AB - BACKGROUND: Type I myosin is highly conserved among eukaryotes, and apparently plays important roles in a number of cellular processes. In the budding yeast, two myosin I species have been identified and their role in F-actin assembly has been inferred. RESULTS: We cloned the fission yeast myo1 gene, which apparently encoded a myosin I protein. Disruption of myo1 was not lethal, but it caused growth retardation at high and low temperatures, sensitivity to a high concentration of KCl, and aberrance in cell morphology associated with an abnormal distribution of F-actin patches. An abnormal deposition of cell wall materials was also seen. Homothallic myo1Delta cells could mate, but heterothallic myo1Delta cells were poor in conjugation. Myo1p was necessary for the encapsulation of spores. The tail domain of Myo1p was pivotal for its function. Calmodulin could bind to Myo1p through the IQ domain at the neck. CONCLUSIONS: Myo1p appears to control the redistribution of F-actin patches during the cell cycle. Loss of Myo1p function is likely to slow down the actin assembly/disassembly process, which results in a failure of the actin cycle to catch up with other events in both the mitotic and meiotic cell cycles, including extension of the conjugation tubes. PMID- 11260264 TI - Functional analysis of the C-terminal cytoplasmic region of the M-factor receptor in fission yeast. AB - BACKGROUND: Yeast mating-pheromone receptors facilitate the study of G protein coupled signal transduction. To date, molecular dissection of the budding yeast alpha-factor receptor has been done extensively, but little analysis has been performed with pheromone receptors of fission yeast, another genetically tractable yeast species. RESULTS: We analysed the fission yeast M-factor receptor Map3p. Truncation of the C-terminal 54 amino acids made Map3p dominant-negative over the wild-type. This form, called Map3-dn9p, was competent in the induction of pheromone-dependent gene expression, although it could not direct proper conjugation. Map3-dn9p failed both to provoke the orientated projection of conjugation tubes and to induce adaptation to the pheromone signal associated with endocytosis of the receptor. Deletion and substitution analyses suggested that the integrity of the C-terminal region, rather than a specific subgroup of amino acid residues therein, was vital for the respective Map3p activities. Ubiquitination of the C-terminus was not absolutely essential for Map3p function. CONCLUSIONS: The C-terminal region of Map3p is dispensable for the pheromone signalling per se, but is pivotal for adaptation and pheromone-induced conjugation tube formation, as is true with the budding yeast alpha-factor receptor. However, the mechanisms which induce adaptation appear to differ between fission and budding yeast concerning the necessity of ubiquitination. PMID- 11260265 TI - The Drosophila UBC9 homologue lesswright mediates the disjunction of homologues in meiosis I. AB - BACKGROUND: In Saccharomyces cerevisiae and other organisms, the UBC9 (ubiquitin conjugating 9) protein modifies the function of many different target proteins through covalent attachment of the ubiquitin-like protein SMT-3/SUMO. RESULTS: Using a second-site suppression screen of a mutation in the nod locus with a variable meiotic phenotype, we have identified mutations in the Drosophila melanogaster UBC9 homologue, encoded by the gene lesswright (lwr). lwr mutations dominantly suppress the nondisjunction and cytological defects of female meiotic mutations that affect spindle formation. The lwr lethal phenotype is rescued by a Drosophila UBC9/lwr transgene. CONCLUSIONS: We suggest that LWR mediates the dissociation of heterochromatic regions of homologues at the end of meiotic prophase I. Our model proposes that when there is less LWR protein, homologues remain together longer, allowing for more normal spindle formation in mutant backgrounds and therefore more accurate meiotic chromosome segregation. PMID- 11260266 TI - Coronin forms a stable dimer through its C-terminal coiled coil region: an implicated role in its localization to cell periphery. AB - BACKGROUND: Coronin is an actin-binding protein, which contains WD (Trp-Asp) repeats and a coiled-coil motif, and plays a role in regulating organization of the actin cytoskeletal network. Coronin localizes to the cell periphery, is involved in lamellipodium extension, and has an implicated role in cytokinesis, cell motility and phagocytosis. RESULTS: Our experiments with two different tagged forms of Xenopus coronin (Xcoronin) have shown that Xcoronin forms an oligomer. This oligomer complex is stable, resistant to 2.4 M NaCl, 0.6 M KI or 2 M urea. Physiochemical analysis of endogenous Xcoronin and the protein expressed in COS7 cells or in bacteria has revealed that the oligomer complex is an Xcoronin dimer. A C-terminal coiled-coil motif of Xcoronin is necessary and sufficient for the dimerization. Mutations in the coiled-coil motif generated dimerization deficient mutants of Xcoronin. Moreover, these mutant forms of Xcoronin failed to localize to the cell periphery, suggesting that dimerization is important for the proper subcellular localization of Xcoronin. CONCLUSION: Xcoronin forms a stable dimer via its C-terminal coiled-coil region. We propose that coronin dimerization is necessary for its proper subcellular localization and function. PMID- 11260267 TI - Tumour suppressor activity of human imprinted gene PEG3 in a glioma cell line. AB - BACKGROUND: Mouse imprinted gene Peg3 encodes a large C2H2 type zinc finger protein with unique characteristics. Peg3 knockout mice were found to show an impairment in maternal behaviour of the adult female. Mouse Peg3 is located on the proximal region of chromosome 7 which is syntenic to the long arm of human chromosome 19. It has been reported that a loss of heterozygosity (LOH) of chromosome 19q occurs frequently in several glioma types. RESULTS: We isolated human PEG3 cDNA. Both human and mouse PEG3 were strongly expressed in the adult brain and the Peg3 protein was localized in the nuclei of both neurones and glial cells. A significant decrease in PEG3 expression was more commonly observed in glioma cell lines as compared with that in primary cultures of astrocytes. Transfection of PEG3 cDNA in a glioma cell line resulted in a loss of tumorigenicity in nude mice. CONCLUSIONS: The human PEG3 gene is a paternally expressed imprinted gene. Introduction of PEG3 cDNA into the glioma cells suggests that human PEG3 protein functions as a tumour suppressor. Human PEG3 is located on 19q13.4 and is one of the candidates for tumour suppressor genes that are predicted to be sited in gliomas. PMID- 11260268 TI - Allele-specific detection of nascent transcripts by fluorescence in situ hybridization reveals temporal and culture-induced changes in Igf2 imprinting during pre-implantation mouse development. AB - BACKGROUND: Genomic imprinting causes parental-origin-specific monoallelic transcription of a subset of mammalian genes in the embryo and adult. There is conflicting evidence, however, for the monoallelic transcription of some imprinted genes, such as Igf2, in pre-implantation embryos. RESULTS: We have developed an allele-specific fluorescence in situ hybridization method which involves a pair of oligonucleotide probes designed to detect an intronic polymorphism. The method, called ASO-RNA-FISH, enabled us to distinguish allelic nascent Igf2 transcripts in the cell nuclei of early mouse embryos, avoiding signals from the stored oocyte-specific transcripts. Igf2 transcription was first detectable in two-cell embryos, and biallelic transcription was predominant up to the morula stage. Then, the maternal allele became silenced during the blastocyst stage. When embryos were cultured in vitro, however, a strong bias to maternal transcription was observed up to the morula stage. CONCLUSION: ASO-RNA-FISH revealed that a transition of Igf2 from biallelic to monoallelic transcription occurs in the blastocyst stage. This developmental regulation was modified temporarily by in vitro culture, suggesting a possible link between altered imprinting and abnormalities of the foetuses experienced in vitro culture. ASO RNA-FISH is therefore a powerful technique for the study of allele-specific gene expression. PMID- 11260269 TI - The ERK MAP kinase pathway mediates induction of SGK (serum- and glucocorticoid inducible kinase) by growth factors. AB - BACKGROUND: The ERK MAP kinase pathway plays a pivotal role in growth factor induced gene expression. However, genes whose expression is induced by the ERK pathway are not fully defined. RESULTS: We have identified SGK (serum- and glucocorticoid-inducible kinase) as an ERK-inducible gene by the subtractive screening of Raf-inducible genes. SGK is known to be similar in primary structure to AKT/PKB, PKC and PKA. Treatment of quiescent NIH-3T3 cells with FGF, PDGF or TPA, which induced the sustained activation of ERKs, resulted in the strong induction of SGK, whereas treatment with EGF, which induced the transient activation of ERKs, did not induce a strong expression of SGK. The induction of SGK was blocked by pre-treatment with a specific MEK inhibitor U0126, and expression of constitutively active MEK was able to induce SGK. Treatment with cycloheximide or vanadate prolonged the increased expression of SGK by FGF, concomitant with a more prolonged activation of ERKs. CONCLUSION: Growth factor induced activation of the ERK MAP kinase pathway is necessary and sufficient for the induction of SGK. PMID- 11260270 TI - Restricted feeding entrains liver clock without participation of the suprachiasmatic nucleus. AB - BACKGROUND: There are two main stimuli that entrain the circadian rhythm, the light-dark cycle (LD) and restricted feeding (RF). Light-induced entrainment requires induction of the Per1 and Per2 genes in the suprachiasmatic nucleus (SCN), the locus of a main oscillator. In this experiment, we determined whether RF resets the expression of circadian clock genes in the mouse liver with or without participation of the SCN. RESULTS: Mice were allowed access to food for 4 h during the daytime (7 h advance of feeding time) under LD or constant darkness (DD). The peaks of mPer1, mPer2, D-site-binding protein (Dbp) and cholesterol 7alpha-hydroxylase (Cyp7A) mRNA in the liver were advanced 6-12 h after 6 days of RF, whereas those in SCN were unaffected. The advance of mPer expression in the liver by RF was still observed in SCN-lesioned mice. A 7 h advance in the LD cycle advanced the peaks of clock gene expression in both the liver and SCN, whereas, a shift in the LD did not move the phase of the liver clock when the shift was carried out under a fixed RF schedule during the night-time. CONCLUSIONS: These results suggest that restricted feeding strongly entrained the expression of circadian clock genes in the liver without the participation of an SCN clock function. PMID- 11260271 TI - Pharmacokinetics of recombinant factor IX in relation to age of the patient: implications for dosing in prophylaxis. AB - The aims of this study were to investigate possible age-related changes in the disposition of factor IX procoagulant activity (FIX:C) after administration of recombinant factor IX (rFIX) and to translate the pharmacokinetic findings into suggestions for dosing of rFIX during prophylactic treatment of haemophilia B. Pharmacokinetic data were available from a previous study on 56 patients, aged 4 56 years (one of whom was excluded from analysis). FIX:C curves during prophylactic dosing were computer-simulated from the single-dose data. Clearance and volume of distribution at steady state of FIX:C increased linearly with body weight of the patients, consequently increasing during childhood and adolescence but remaining fairly constant during adulthood. The terminal half-life of FIX:C showed no correlation with age, while in vivo recovery (in U dL(-1) per U kg(-1) given) tended to increase. Computer-predicted trough levels of exogenous FIX:C during repeated doses of rFIX (50 U kg(-1)) and, conversely, doses (in U kg(-1)) needed to maintain a 1-U dL(-1) trough level showed little or no dependence on age. There was considerable interindividual variation in disposition and required doses of rFIX, emphasizing the need for individual dose titration. Dosing of rFIX according to lean body mass instead of body weight did not reduce this variability. During prophylaxis a 1-U dL(-1) trough level can normally be maintained by dosing every 2-3 days, the former schedule resulting in, on average, a 45% lower consumption of rFIX. PMID- 11260272 TI - Recombinant factor IX (BeneFix) by adjusted continuous infusion: a study of stability, sterility and clinical experience. AB - The safety and efficacy of adjusted continuous infusion (CI) of recombinant factor IX (FIX; BeneFix) was assessed in vitro and in a clinical study. BeneFix was reconstituted at 100 IU mL-1 with or without unfractionated heparin (4 U mL 1) and stored at either 4 degrees C or room temperature. Reconstituted BeneFix retained at least 90% activity over 14 days if stored at 4 degrees C but stability was reduced at room temperature. BeneFix reconstituted in a sterile pharmacy was free of bacterial contamination. Six patients with haemophilia B received seven CIs of BeneFix to cover routine surgery and severe bleeding episodes. The CIs lasted between 3 and 10 days. In all cases, haemostasis was excellent and the desired therapeutic FIX level was easily maintained. No thrombotic episodes or inhibitor development occurred but two patients developed thrombophlebitis at the infusion site when heparin was not added to the infusion. BeneFix is not currently licensed for CI and we suggest that studies to enable licensing should be established as soon as possible. PMID- 11260273 TI - Viral safety of a pasteurized, monoclonal antibody-purified factor VIII concentrate in previously untreated haemophilia A patients. AB - The efficacy and viral safety of a pasteurized, immunoaffinity-purified procoagulant factor VIII protein (FVIII:C; Monoclate-P) was studied in two multicentre, prospective, open-label trials in 30 previously untreated patients, 18 with severe (< 1% FVIII:C activity), and 12 with moderate (1% to 5% FVIII:C activity) haemophilia A. Clinical assessments, performed at screening and regularly thereafter for 6 to > 24 months (maximum 34 months), showed that none of 24 assessable patients acquired illnesses consistent with monitored transfusion-transmissible diseases. No patients acquired hepatitis B surface antigen, or antibodies against hepatitis B core antigen, hepatitis C, or human immunodeficiency virus. Likewise, no patients acquired treatment-related hepatitis A antibodies or sustained elevations of alanine aminotransferase levels. The safety profile for Monoclate-P is brought about by a multi-step safety system that incorporates viral inactivation (through a combination of immunoaffinity chromatography and pasteurization) plus donor screening, plasma testing, and quality assurance. The inhibitor development rate (13% low titre, 10% high titre) was similar to that reported in the literature for other FVIII concentrates (24% to 52%). The most frequently reported adverse events were related to typical infant and childhood diseases. Monoclate-P was effective in all patients treated according to protocol, except in two, who developed inhibitors. PMID- 11260274 TI - Immune tolerance treatment in haemophilia patients with inhibitors: the Spanish Registry. AB - We present a retrospective study of immune tolerance treatment (ITT) carried out in 42 Spanish haemophiliac patients. Most of the patients were high responders (39/42), with a median maximum titre of 67 Bethesda units (BU) (range 6-2984). The median inhibitor titre at the start the ITT was 11 BU (range 1-256 BU) and the median age of the patients was 7 years (range 0-57). The mean factor dosage was 140 IU kg bodyweight(-1) day(-1) (range 25-500). In most of the ITTs, plasma derived factor concentrate of intermediate and high purity was used. The inhibitor was eradicated in 26/38 (68%) of the patients who completed the treatment and two patients changed their status from high to low responders. Multivariate logistic regression analysis showed that three significant variables were associated with the highest probability of success: (i) the use of low factor doses for ITT (< or = 100 IU kg(-1) day(-1); P = 0.0106; 95% CI 0.000289 0.342); (ii) a titre of < 10 BU at start of ITT (P = 0.0286; 95% CI 0.00253 0.7189) and (iii) a lower maximum titre (P = 0.0214; 95% CI 0.98-0.9994). PMID- 11260275 TI - Combined prednisolone and intravenous immunoglobulin treatment for acquired factor VIII inhibitors: a 2-year review. AB - Acquired inhibitors to factor VIII (FVIII) are rare, but life-threatening in up to 22% of cases. The optimal therapy for suppression of these inhibitors remains unclear. Prednisolone is the mainstay of therapy, producing responses in approximately 30% of cases. Intravenous immunoglobulin (IVIg) has a similar response rate, but a more rapid effect. We report the results of prednisolone 1 mg kg(-1) combined with IVIg 2 g kg(-1) in divided doses as first-line therapy in seven consecutive patients with acquired FVIII inhibitors. All patients were bleeding at the time of diagnosis with prolonged activated partial thromboplastin time. There were four complete responses, one partial response, one nonresponse and one with an inadequate follow-up for assessment of response, giving an overall response rate of 71%. In all complete responders the inhibitor declined rapidly and was undetectable by day 21 from start of treatment. Therapy was well tolerated and responses have been maintained off treatment for 2-8 months. This is a safe, well-tolerated rapidly acting regimen with good response rates. PMID- 11260277 TI - Utility of the PFA-100 for assessing bleeding disorders and monitoring therapy: a review of analytical variables, benefits and limitations. AB - The PFA-100 (platelet function analyser) is a relatively new tool for the investigation of primary haemostasis. Recent studies have shown its utility in monitoring antiplatelet therapy (including aspirin) and as a screening tool for investigating possible von Willebrand disease (vWD) and various platelet disorders. More recently, the PFA-100 has been shown to be of value in monitoring DDAVP therapy in both vWD and platelet disorders. This paper reviews current findings, details the utility of the PFA-100 for some of these purposes, as well as reviewing analytical variables that may complicate the interpretation of results. The author highlights the benefits, as well as noting the limitations, of its use. Ultimately, the greatest strengths of the PFA-100 are its simplicity in use and excellent sensitivity to particular haemostatic disturbances such as vWD, platelet disorders and platelet-affecting medication. However, because it is thus a 'global' test system, this also creates a significant limitation, as the PFA-100 is not specific for, nor predictive of, any particular disorder. However, utilized appropriately, the PFA-100 can be considered a worthwhile addition to any haemostasis laboratory involved in the diagnosis or therapeutic-monitoring of bleeding disorders including vWD and platelet-dysfunctions. This review should be of value to both haemostasis scientists and clinical specialists. PMID- 11260276 TI - Acquired haemophilia: experiences with a standardized approach. AB - Acquired haemophilia is a rare, life-threatening, acquired bleeding diathesis. No general consensus exists on the best therapeutic approach. We report on the standardized approach at our institution evaluated in ten patients with acquired haemophilia. Factor VIII inhibitors were found in all patients, activities ranging from 1 to 648 Bethesda units (BU). Eight of the ten patients presented with severe bleeding. Two patients died during the acute phase, one from intracranial bleeding and one due to Mycoplasma pneumonia. One patient with mild bleeding was treated with immunosuppression alone. Two patients with factor VIII inhibitor activities below 5 BU were started on factor VIII concentrate therapy. Therapy was successful in one and was changed to recombinant human activated factor VII infusion (rFVIIa) in the other, owing to insufficient factor VIII recovery. Six patients with factor VIII inhibitor activities above 5 BU were started on activated prothrombin complex concentrate (APCC) therapy. APCC treatment was successful initially in all six patients and was changed to rFVIIa infusion in one for rebleeding. One patient did not receive any specific therapy. Immunosuppression with prednisolone (2 mg kg(-1)) was begun in nine patients and was continued with cyclophosphamide (2 mg kg(-1)) in six. A complete remission of the acquired haemophilia was found in seven of the eight patients surviving the acute phase, one had a partial remission. All patients with acquired haemophilia could be managed effectively following our standardized approach. Routine administration of immunosuppression was associated with high inhibitor elimination rates. PMID- 11260278 TI - Laboratory diagnosis of von Willebrand disorder (vWD) and monitoring of DDAVP therapy: efficacy of the PFA-100 and vWF:CBA as combined diagnostic strategies. AB - We have coevaluated a combination of test processes for diagnosing von Willebrand disease (vWD) and monitoring deamino-delta-D-arginine vasopressin (DDAVP) therapy. Using normal controls (n = 23), closure time (CT) ranges measured by PFA 100(R) were (mean +/- 2SD): (i) collagen/ADP cartridge (C/ADP): 67-127 s (ii) collagen/epinephrine (C/Epi): 94-162 s. From a panel of 125 patients undergoing evaluation for clinical haemostatic defects, 29/30 samples from patients with vWD [17/18 type 1, 1/1 type 3, 3/3 type 2A, 7/7 type 2B and 1/1 pseudo-vWD] gave prolonged CTs using C/Epi. The C/ADP was less sensitive, being normal in 7/18 of the type 1 vWD individuals, with higher sensitivity to more severe vWD. Individuals with haemophilia (six factor VIII-deficient, one factor XI-deficient) gave normal CTs, while those with clinical thrombocytopenia (n=13) gave normal or prolonged CTs, somewhat dependent on platelet count. The PFA-100 was also evaluated as a part of the laboratory monitoring procedure in patients with either vWD or haemophilia undergoing a DDAVP trial as a therapeutic management process. For vWD, correction of an initially prolonged CT by DDAVP, accompanied by normalization of von Willebrand factor (vWF) measurable by von Willebrand factor antigen, vWF collagen binding activity and vWF ristocetin cofactor assays (vWF:Ag, vWF:CBA and vWF:RCof), was achieved in type 1 vWD (n=5). In an individual with type 2A vWD, DDAVP normalized vWF:Ag and vWF:RCof, but had no apparent effect on the baseline maximally prolonged CT. In an individual with type 2B vWD, factor VIII/vWF concentrate also normalized vWF:Ag and vWF:RCof, but similarly had no apparent effect on the baseline maximally prolonged CT. vWF:CBA did not normalize for either of these individuals, potentially suggesting that normalization of vWF:CBA might be required for normalization of CT. This concept is supported by correlation analysis undertaken between CT and various vWF parameters. Among these, vWF:CBA held the strongest relationship in our data set, which showed an inverse progressive rise in CT for falling vWF:CBA. Based on these results, we would conclude that the PFA-100 is highly sensitive to the presence of vWD, and may thus provide a valuable screening test for vWD. Furthermore, the combined utility of the PFA-100 and vWF:CBA as markers of DDAVP responsiveness may prove to be simple, quick but powerful, predictors for its clinical efficacy. PMID- 11260279 TI - Validity of health status measurement with the Dutch Arthritis Impact Measurement Scale 2 in individuals with severe haemophilia. AB - The aim of this study was to investigate the validity of the Dutch Arthritis Impact Measurement Scales 2 (D-AIMS2). Hence, D-AIMS2 data of individuals with severe haemophilia were correlated with clinical and perceived health-related quality of life data. Patients with severe haemophilia, who visit the Van Creveldkliniek on a regular basis, were administered the D-AIMS2. In addition, health-related quality of life was measured by the Sickness Impact Profile (SIP). As clinical indices, range of movement (which was converted into Joint Alignment and Motion scores) and muscle strength were recorded during the routine visit. Extensive descriptive and correlational (linear) analyses between corresponding datasets were performed. Thirty-one individuals with severe haemophilia were included. Their scores on the D-AIMS2 demonstrated moderate to very high internal consistency for scales and components (Cronbach's alpha = 0.62-0.92). The physical health components of the D-AIMS2 and the SIP were significantly correlated (Pearson's r = 0.53; P < 0.05). The psychological health and social interaction components of the D-AIMS2 did not correlate significantly with the psychosocial component of the SIP. The physical health component of the D-AIMS2 correlated significantly with the clinical data for the lower extremities (r = 0.52 and r = -0.45; P < 0.05). These data support the reliability and validity of the physical aspects of the D-AIMS2 in patients with severe haemophilia. The next step should be to extend the investigation of psychometric qualities of this health-related questionnaire in a larger population. PMID- 11260280 TI - Home-based factor infusion therapy and hospitalization for bleeding complications among males with haemophilia. AB - Information from the medical records of 2650 US males with haemophilia living in six states was used to examine the influence of infusing factor concentrate at home (home therapy) and other variables on rates of hospitalization for a haemorrhagic bleeding complication (HBC) over a 4-year period. Bleeding complications included actual and suspected haemorrhagic events but excluded elective admissions for procedures necessitated by haemorrhage (e.g. joint synovectomy). Other risk determinants considered in the analyses included age, race, employment status, health insurance type, care received in federally funded haemophilia treatment centres (HTCs), factor deficiency type and severity, amount of factor prescribed, prophylactic treatment, and presence of inhibitors at baseline. Survival analysis methods were used to evaluate relationships between baseline risk factors and subsequent hospitalization rates. During 8708 person years (PYs) of follow-up, 808 subjects (30.5%) had a total of 1847 bleeding related hospitalizations; an overall rate of 21.2 admissions per 100 PYs. Using proportional hazards regression to adjust for all of the studied factors, we found that home therapy use (among residents of four of the states) and care in HTCs were independently associated with a decreased risk for a first HBC. Patients who had government-sponsored health insurance or who had no insurance, those of minority race or ethnicity, those with higher levels of factor use, and those with inhibitors were at increased HBC risk. We conclude that the use of home therapy and receipt of care in HTCs are each associated with a substantially lower risk for HBC among males with haemophilia. PMID- 11260282 TI - High-titre factor VIII inhibitor in two children with mild haemophilia A. AB - A frequently encountered complication of therapy given to patients with severe haemophilia A is the development of antibodies to infused factor VIII. While much less common, inhibitors also occur in patients with mild or moderate severity haemophilia A. Often thought to be of low titre and transient, several cases of high-titre inhibitors have been described in patients with mild or moderate haemophilia A. Generally these occur in adults or adolescents following significant infused factor VIII exposure. A review of reported cases revealed only two cases of high-titre inhibitor formation in mild haemophilia A patients younger than 10 years of age. We wish to report our experience with an additional two children with mild haemophilia A and high titre inhibitors, and offer suggestions for the management of these children. PMID- 11260281 TI - Sustained and therapeutic delivery of factor IX in nude haemophilia B mice by encapsulated C2C12 myoblasts: concurrent tumourigenesis. AB - This study reports the generation of an immunodeficient murine model for haemophilia B, obtained by breeding factor IX-deficient mice with an immunodeficient mouse strain, and use of this mouse model to evaluate the long term efficacy and safety of a gene therapy strategy for treating haemophilia B. Nude haemophilic mice were implanted with biocompatible microcapsules enclosing recombinant myoblasts secreting human factor IX. The activated partial thromboplastin time (APTT) of plasma of mice thus treated was invariably shortened 3 weeks after microcapsule implantation, and remained shortened for at least 77 days. Shortening of the APTT of the haemophilia mice coincided with the appearance of human factor IX in mice plasmas (up to 600 ng mL(-1) on day 77), and normalization of the tail-bleeding time. Thus, the microencapsulated myoblasts reversed the clinical phenotype of haemophilia B. In contrast, plasmas of immunocompetent haemophilic mice similarly implanted with microcapsules only showed a transient shortening of APTT, and coincident transient delivery of human factor IX antigen. Rapid disappearance of human factor IX from plasmas of immunocompetent mice also coincided with production of antibodies to the human transgene. Significantly, 86% of the nude haemophilia mice developed tumours of myoblast origin. Thus, while this study revealed the feasibility of this gene therapy approach to treat severe haemophilia B, it also highlights the importance of using safer cell lines to prevent tumour development. PMID- 11260283 TI - Prophylactic treatment of severe factor X deficiency with prothrombin complex concentrate. AB - Factor X (FX) deficiency is an autosomal recessive trait that occurs in fewer than 1 in 500 000 people. Not surprisingly, reports of prophylactic treatment for FX deficiency are exceedingly rare. We now report our experience of the use of prophylactic therapy in a FX-deficient patient. This 18-year-old African-American male presented at the age of 4(1/2) years with an FX level < 1%. Treatment was on demand with prothrombin complex concentrates (PCCs) given at two times the dose per kilogram of body weight for factor IX. He experienced frequent epistaxis, soft tissue bleeding and joint bleeding. The development of a target joint (right ankle) prompted the initiation of prophylactic treatment in the beginning of 1998 to the present with 30 units kg(-1) Profilnine twice per week via a home infusion programme. If breakthrough bleeding occurred, he was instructed to infuse another dose. He was instructed that Profilnine should not be infused in more than two doses in 24 h or on more than three consecutive days. A trough level drawn 48 h post-infusion showed an FX level of 30%. In the initial 12 months with prophylactic treatment, there was no breakthrough bleeding. Subsequently, with an additional 11 months of follow-up, he has reported one bleed. He rates his quality of life improved since starting prophylactic treatment. There have been no thrombotic events. Prophylaxis with PCC for FX deficiency with adequate education and follow-up can be performed capably in the home setting with a resultant decrease in the frequency of bleeding and attendant complications. PMID- 11260284 TI - The pelvi-femoral incomplete bone bridge in a patient with mild haemophilia. AB - A 15-year-old boy with mild haemophilia who regularly participates in contact sports presented with right hip pain radiating to the groin and buttock areas and difficulty in walking. Conventional radiography disclosed a heterotopic new bone formation in the adductor region. The reformatted and three-dimensional reconstructed images of computerized tomography (CT) scans detailed an incomplete pelvi-femoral bone bridge formation in the quadratus femoris muscle, which was located very close to the sciatic nerve but did not cause any clinical symptoms. Postural exercises and clinical survey were selected as the primary treatment. PMID- 11260285 TI - Heart transplant in a factor VIII-deficient patient with a high-titre inhibitor: perioperative management using high-dose continuous infusion factor VIII and recombinant factor VIIa. AB - Four years prior to transplantation, a 14-year-old boy with severe haemophilia A and a high-responding factor VIII (FVIII) inhibitor developed an anteroseptal myocardial infarct while receiving high doses of an activated prothrombin complex concentrate (PCC). Cardiac transplantation was required for survival because of the ensuing cardiomyopathy. At surgery, the patient's inhibitor titre was 1.8 Bethesda units (BU). High-dose bolus therapy, followed by a continuous infusion of FVIII provided excellent operative and initial postoperative haemostasis without additional blood-product support. Once anamnaesis developed on day 6 postoperatively, recombinant factor VIIa (rFVIIa) therapy was initiated. Haemostasis remained excellent, except for the transient increase in chest-tube bleeding that was noted on day 7. epsilon-Aminocaproic acid was added and haemostasis was re-established. On day 15, rFVIIa was replaced with alternate day infusions of prothrombin complex concentrates (PCCs). On day 21 following the transplant, the patient was discharged, remaining on daily FVIII immune tolerance and thrice-weekly PCC prophylaxis. He remains well 24 months after transplant with an inhibitor titre of 39 BU. This paper describes the second case of cardiac transplantation complicated by haemophilia and an inhibitor, and discusses preoperative planning and operative and postsurgical haemostasis management. PMID- 11260287 TI - Nurses' use of computer databases to identify evidence for practice--a cross sectional questionnaire survey in a UK hospital. AB - The objectives of this study were to determine nurses' use of electronic databases to inform practice. A questionnaire survey of 114 nurses working on five acute wards in a large inner city teaching hospital investigated their general use of computers and the three databases, CINAHL, MEDLINE and the Cochrane Library. Eighty-two qualified nurses responded (response rate 72%). The results show limited confidence and low frequency in using the databases. Thirty four per cent expressed low confidence using CINAHL. Twenty-seven per cent used CINAHL on a regular basis. Twenty-two per cent never used it. Eighteen per cent were unaware that it was available locally. Knowledge and use of MEDLINE was even lower with only 18% using it regularly. Knowledge of the Cochrane Library was extremely limited, with 75% unaware of its existence. Use of a home computer and higher education were associated with higher frequency of use of CINAHL and MEDLINE. If nurses are to make use of electronic resources to contribute to evidence-based practice, effort needs to be put into ensuring that already qualified nurses have basic computer skills and specific knowledge of available resources. More emphasis should be placed on 'evidence-based' resources, such as the Cochrane library, as sources of information for practice. PMID- 11260288 TI - A controlled vocabulary for nursing and allied health in Norway. AB - Nursing and allied health libraries at educational institutions in Norway have generally indexed their book collections with uncontrolled terms. With the reorganization of higher education in 1994, the majority of these libraries joined BIBSYS, which is a joint library system for higher education and research in Norway. This has led to chaos when searching the joint catalogue for literature on nursing and related fields. A term such as 'behaviour problems' may have up to five synonyms. In an attempt to improve the quality of searching the health literature, BIBSYS appointed a working group in the Spring of 1999 to find a suitable controlled vocabulary for this subject area, and to see how this vocabulary could be integrated into BIBSYS. The group presented its recommendations in October 1999. The report has been well received by the BIBSYS Board and by user groups. There are no Norwegian vocabularies that are suitable for use in nursing and allied health, therefore it will be necessary to translate and combine existing thesauri. The group has looked at the Nordic Multilingual Thesaurus on Health Promotion, the Swedish Spriline Thesaurus, MeSH (Medical Subject Headings) and CINAHL Subject Heading List. Other relevant thesauri are AMED/CATS Thesaurus, Bioethics Thesaurus (Bioethicsline) and the RCN thesaurus. The group recommends the development of a Norwegian thesaurus based on a translation of parts of MeSH and CINAHL Subject Heading List. PMID- 11260289 TI - Using metadata to create navigation paths in the HealthInsite Internet gateway. AB - The objective of this study was to evaluate the HealthInsite topic query technique, which uses a dynamic database search to assign resources to a topic. It is an alternative to the explicit classification technique, which relies on the classification of each resource using a predefined classification scheme. We performed a recall-precision analysis on all topics within the broad topic area of Child Health. Recall and precision errors were checked to determine which part of the information retrieval process was at fault. We then compared the topic query technique with the explicit classification technique. The results show errors or problems at every stage of the information retrieval process. This has initiated a review of all the tools used in the process, from indexing guidelines to the search engine. While many errors could be corrected, there were still features of the explicit classification technique that could not be achieved by the topic query technique. In conclusion, the topic query technique has the advantage of flexibility, but close co-operation between the different information retrieval specialists is needed to get the best results. The HealthInsite topic navigation structure should be regarded as an organized set of predefined searches rather than a full classified listing. PMID- 11260290 TI - The focus group: a tool for programme planning, assessment and decision-making- an American view. AB - The objective of this paper is to describe the focus group process used with hospital librarians in the National Network of Libraries of Medicine, Pacific Southwest Region (NN/LM/PSR), in order to illustrate how focus groups can be effectively used in the library setting to plan programmes around identified needs. This paper explores the focus group methodology, a qualitative research technique, by discussing why it is used and the process involved in its use. Aspects of the methodology that are discussed include participant selection, question development, data analysis, and use of results in programme planning. The focus group findings assisted NN/LM/PSR in understanding the needs of hospital librarians related to integrating electronic resources into library services. The focus group data were used to determine the forum, content and speakers for a day-long symposium and subsequent planning meeting. The use of the focus group technique to assess the needs of a specific group on a specific issue resulted in activities and programmes that met these needs successfully. Based on the experiences detailed in this paper, the authors are confident that focus groups are an effective tool for programme planning, needs assessment and decision-making for all types of libraries. PMID- 11260291 TI - Postgraduate training in public health medicine: St George's Hospital Medical School Library public health information service. AB - This article examines the development of the St George's Hospital Medical School Library public health information service. Begun in 1997 as a pilot project to support Public Health Specialist Registrars in South Thames West, it is now an established part of postgraduate training in the region. An outline of the service is described, including the evolution of the post of Public Health Librarian. Issues influencing the development of the service, and the establishment of the Librarian as part of the public health network are discussed. This is a transferable model of public health information provision, which as a centralized resource makes best use of available funding. As a LIS model it is an effective and efficient way of maximizing resources, and delivering a service to a specialist user group that is spread across a wide geographical area. PMID- 11260293 TI - Opportunities on the Web: a role for information professionals, using the development of the BMA Library Online Service as a case study. AB - The paper considers how attributes and skills fundamental to the information profession may be applied to the development and management of Web-based services and resources. It also looks at the need for the acquisition of new skills as part of a continuing professional development programme. The report considers the changing role of the Webmaster and the implications this might have in terms of the role of information professionals. To offer a practical example, from within the healthcare sector, the article goes on to describe the development of BMA Library Online. This is the BMA Library's own suite of Web-based information services and resources for personal members, institutional members and staff. The library's own Web team has been responsible for the development and maintenance of BMA Library Online. To conclude, the paper looks forward to future possibilities for information professionals as Web developers and at how this might be influenced by changes in the Webmaster role. It is crucial that skills and expertise are shared as the roles that go to make up a Webmaster or Webmaster team continue to merge and evolve, and with the possible wider distribution of provision of Web content within organizations. PMID- 11260294 TI - Finding health statistics: making a difficult task easier. PMID- 11260295 TI - Introducing Open University Health Studies students to Internet resources--the ROUTES database. PMID- 11260296 TI - Will health librarians and related information workers ever work together to create an international network? PMID- 11260297 TI - The role of the histopathologist in the management of testicular germ cell tumour in adults. AB - In the last 20--30 years the availability of effective chemotherapy and more accurate clinical staging has greatly improved the prognosis for patients with testicular germ cell tumours. Initially, such treatment appeared to diminish the role of histopathology to the distinction between seminoma and nonseminomatous germ cell tumour (NSGCT) in the primary specimen. However, histopathology has evolved as a prognostic tool indicating the risk of relapse in various defined clinical contexts thereby facilitating therapeutic decisions. The clinical emphasis has been on quality of life and reduction of therapy both in terms of the number of patients treated and the number of chemotherapy courses given to each patient. The treatment of adult testicular germ cell tumours may differ between countries but protocols are established. Therefore it is appropriate to discuss the role of histopathology during this era of relative therapeutic stability. PMID- 11260299 TI - Reproducibility of the WHO/IASLC grading system for pre-invasive squamous lesions of the bronchus: a study of inter-observer and intra-observer variation. AB - AIMS: Although many workers have graded pre-invasive squamous lesions arising in the bronchus, there has been no consensus classification system until the latest edition of the WHO/IASLC histological classification of pulmonary and pleural tumours. Because the value of any such system is dependent on its reproducibility, we have circulated a series of such lesions to a panel of histopathologists to assess interobserver and intra-observer variation when the WHO/IASLC classification was applied. METHODS AND RESULTS: Colour transparencies of 28 pre-invasive squamous lesions were assessed by six histopathologists (two with a special interest in pulmonary pathology, two generalists and two trainees) on three separate occasions over a period of 3 months, using the criteria of the WHO/IASLC (mild, moderate and severe dysplasia, and in-situ carcinoma). An additional category of metaplasia was added for those cases that showed no dysplasia. Weighted kappa coefficents of agreement (K(w)) were used to evaluate paired observations with a standard quadratic weighting being employed, such that kappa coefficients corresponded to intra-class correlation coefficients. Wilcoxon's sign-ranked test was used to measure the statistical significance of group trends, when comparing kappa values for the three grading systems. Various 3-point systems were also assessed, through combination of the above groups. Intra-observer agreement was substantially better than interobserver variation (mean: 0.71 vs. 0.55). Between the various pathologist groups, inter-observer variation was relatively minor, although intra-observer variation was higher within the trainee pathologist group. Using weighted kappa values, there was no significant difference in either inter-observer or intra-observer agreement between the five point grading system and a 3-point system of metaplasia/mild, moderate and severe/in-situ grades. However, there was a significant increase in variation when a 3-point system of metaplasia/mild, moderate/severe and in-situ carcinoma was used. CONCLUSION: This study shows levels of interobserver and intra-observer variation similar to those found in other grading systems in histopathology, with no significant decrease in variability found by abridging the system. The WHO/IASLC system is therefore recommended for future use in both clinical and research fields. PMID- 11260298 TI - Effects of long-term treatment with the anti-androgen bicalutamide on human testis: an ultrastructural and morphometric study. AB - AIMS: To assess the effects of more than 4 years' treatment with the anti androgen bicalutamide on human testis by clinical, ultrastructural and morphometric analysis. METHODS AND RESULTS: Two patients (aged 74 and 69 years) with prostate cancer were treated for more than 4 years with bicalutamide 50 mg daily. Clinical characterization and follow-up included luteinizing hormone (LH), follicle-stimulating hormone (FSH), testosterone and prostate-specific antigen (PSA) measurements and clinical response of the tumours. Due to progression of the disease, patients underwent surgical orchidectomy as a further androgen withdrawal therapy. Testis biopsies were studied by light and electron microscopy and analysed by morphometry. Control samples were obtained from the normal testis of two patients with testicular cancer who underwent orchidectomy. Clinical follow-up showed a good response in the control of tumour growth and serum PSA decreased to < 4 ng/mL; concentrations of serum LH, FSH and testosterone were within the normal range. Testicular morphology of treated patients was unexpectedly well preserved; the organization of seminiferous tubules was normal with all the germ line elements and mature spermatozoa present. In some areas, a net increase of peritubular connective tissue was evident which may be a consequence of the age of the patients. CONCLUSIONS: Long-term bicalutamide (50 mg) treatment appears to have very little impact on testis ultrastructure and sperm maturation, while it is effective in the control of androgen-dependent prostatic tumours. PMID- 11260300 TI - High expression of MDR-1 gene and P-glycoprotein in initial and re-biopsy specimens of relapsed B-cell lymphoma. AB - AIMS: Multidrug resistance (MDR) is a major obstacle in the treatment of lymphoma. The expression of MDR-1 mRNA and P-glycoprotein (MDR-1/P-gp) has been linked to MDR. We aimed to investigate the expression of MDR-1/P-gp in B-cell lymphoma. METHODS AND RESULTS: Samples at diagnosis and relapse from 10 patients with B-cell lymphoma were obtained. We also obtained 14 unselected control cases of B-cell lymphoma at diagnosis. The expression of mRNA and protein were determined semiquantitatively by RT-PCR and immunohistochemistry. High MDR-1 and P-gp expressions were found in seven and seven of 10 samples obtained at diagnosis, eight and eight of 10 samples obtained at relapse, and three and four of 14 control cases at diagnosis, respectively. The results of RT-PCR paralleled those of immunohistochemistry. Concordance of high MDR-1/P-gp expression was noted in 27 of 34 samples (r = 0.73, P = 0.001). There were no significant changes in MDR-1/P-gp expression in all cases at relapse and during the clinical course following chemotherapy. In the 14 control cases, the average survival time was 12.7 months in MDR-1/P-gp positive cases and 29.0 months in the MDR-1/P-gp negative cases (P = 0.20). CONCLUSIONS: Our results showed that at least some B cell lymphomas express MDR-1/P-gp, which could be detected by different methods, and suggested that high MDR-1/P-gp expression in tumour cells may be associated with a high probability of relapse and poor prognosis. PMID- 11260301 TI - Estimation of age of human cadavers by immunohistochemical assessment of advanced glycation end products in the hippocampus. AB - AIMS: Advanced glycation end products (AGEs) are known to accumulate in long lived tissue proteins during normal ageing. In this study, we examined the expression of AGEs in human hippocampus using immunohistochemistry and determined its utility for estimating the age of cadavers of unknown age. METHODS AND RESULTS: Hippocampus tissues were obtained at autopsy from 31 individuals, including 10 fire victims, aged 0--96 years within 3 days postmortem. Immunostaining using anti-AGE antibody demonstrated that the perikarya of pyramidal neurones in the hippocampus was immunoreactive for the anti-AGE antibody, and the immunoreactivity was increased with age. Quantitative analysis of the AGE-immunoreactivity in the pyramidal neurones of the CA4 region revealed a significant correlation between the AGE-immunoreactivity and the age in nonfire death cases with a correlation coefficient of 0.91 (P < 0.01). The significant correlation could be obtained even in fire death cases affected by the unusual environmental condition. CONCLUSIONS: These results suggest that the immunohistochemical analysis of AGEs in human hippocampus may be useful for the age estimation of cadavers with unknown age. PMID- 11260302 TI - Apocrine adenosis of the breast: clonal evidence of neoplasia. AB - AIMS: We report here a case of apocrine adenosis of the breast in a 66-year-old woman. To clarify the nature of this lesion, we examined it by clonal analysis. METHODS AND RESULTS: The lesion, 43 mm at its greatest dimension, was ill circumscribed, lacking a fibrous capsule, and was composed of compact glands that showed typical histological features of apocrine adenosis. Clonality analysis, using the polymerase chain reaction (PCR)-based method for females heterozygous for a BstXI polymorphism of the X-linked phosphoglycerokinase (PGK) gene, revealed the monoclonal nature of the lesion. CONCLUSIONS: This result strongly suggests that some populations of apocrine adenosis are already neoplastic, and this could contribute to the premalignant potential of this lesion. PMID- 11260303 TI - Histological, genomic and clinical heterogeneity of clear cell hepatocellular carcinoma. AB - AIMS: The aim of the study was to determine whether clear cell type hepatocellular carcinoma should still be regarded as a separate uniform diagnostic entity. METHODS AND RESULTS: We retrospectively studied 118 cirrhotic patients with hepatocellular carcinoma treated by orthotopic liver transplantation, and 31 noncirrhotic patients with hepatocellular carcinoma treated by either liver surgical resection or transplantation. The pathology of all liver resections was reviewed. Microsatellite instability was performed on paraffin-embedded samples at loci located on chromosomes 2p, 3p, 5q, 8q, 9p, 13q, 16q and 17p. Among the 118 cirrhotic patients, 10 (8.5%) had a clear cell hepatocellular carcinoma; these had clinical characteristics and prognosis similar to the other cirrhotic patients. No genetic alterations were detected in these tumours. Among the 31 noncirrhotic patients, one (3.2%) had a clear cell hepatocellular tumour. This 170-mm tumour had a lipid density on computed tomography, and its histology resembled chromophobe cell renal carcinoma. Furthermore, this tumour had unusual genomic alterations, with microsatellite instability in 6/8 chromosome loci. CONCLUSIONS: Clear cell hepatocellular carcinoma is a heterogeneous entity in which a chromophobe cell subtype should be identified. PMID- 11260304 TI - Basaloid carcinoma of the colon arising at the splenic flexure. AB - AIMS: Basaloid carcinomas typically arise in the anal canal and there are only three well-documented cases of this neoplasm reported outside the anal canal, none more proximally than the sigmoid colon. The first occurrence of a basaloid colonic carcinoma arising outside the sigmoid colon, at the splenic flexure, is presented. METHODS AND RESULTS: A splenic flexure mass was resected from a 54 year-old man with a 3-week history of abdominal discomfort, diarrhoea and weight loss. This tumour, like typical anal canal basaloid carcinomas, was composed of islands of basaloid cells with peripheral nuclear palisading; within many islands there was central necrosis and focal squamous differentiation. Ultrastructural and immunohistochemical studies confirmed the basaloid nature and focal squamous differentiation within this neoplasm. Basaloid carcinoma of the anal canal has been associated with human papilloma virus. Using in-situ hybridization, HPV DNA was not detected in this case. CONCLUSIONS: Outside the anal canal, it has been postulated that basaloid colonic carcinomas may arise from cloacogenic embryologic rests, squamous metaplastic epithelium, or totipotential basal cells. The location and pathological findings of this tumour suggest that this rare colonic neoplasm arises from a totipotential basal cell. PMID- 11260305 TI - Female adnexal tumours of probable Wolffian origin: an immunohistochemical study comparing tumours, mesonephric remnants and paramesonephric derivatives. AB - AIMS: To establish an immunohistochemical profile of presumed female adnexal mesonephric tumours (FATWO) for diagnostic purposes and to compare the findings with those of mesonephric and paramesonephric derivatives in order to establish supportive evidence for a mesonephric origin. METHODS AND RESULTS: Standard immunohistochemistry was performed on formalin-fixed tissues. Tumours, mesonephric remnants and paramesonephric structures generally show positive staining for vimentin, CAM 5.2 and cytokeratins 7 and 19 but are negative for CK20 and 34 beta E12. EMA is positive in both mesonephric and paramesonephric derivatives but is negative in the tumours. Glutathione S-transferase mu (GST mu) is generally positive in both tumours and mesonephric derivatives but negative in paramesonephric structures. CONCLUSIONS: Immunohistochemistry plays little part in the diagnosis of FATWO. The tumours are generally cytokeratin and vimentin positive and EMA-negative. GST mu, as a marker for the mesonephric duct, is a useful adjunct. Our findings of the study support but do not prove that FATWOs are of mesonephric origin. PMID- 11260306 TI - Apoptotic activity in gestational trophoblastic disease correlates with clinical outcome: assessment by the caspase-related M30 CytoDeath antibody. AB - AIMS: The objective of this study was to assess apoptotic activity in gestational trophoblastic disease (GTD) and its prognostic value in hydatidiform mole (HM). METHODS AND RESULTS: Expression of the specific caspase cleavage site within cytokeratin 18 was assessed immunohistochemically using the monoclonal antibody M30 CytoDeath in 12 spontaneous abortions, 22 partial and 57 complete HM, eight choriocarcinoma (CCA) and 28 normal placentas. The M30 immunoreactivity occurred predominantly in the syncytiotrophoblasts. A significantly higher M30 index in HM and CCA was found when compared with normal placentas and spontaneous abortions (P < 0.001). The M30 index of those HM which spontaneously regressed was significantly higher than those HM which developed persistent disease requiring chemotherapy (P < 0.001). The M30 index correlated with another apoptotic index previously detected by TdT-mediated dUTP nick-end labelling (TUNEL) (P = 0.007) and the proliferation index assessed by the Ki67 antigen (P = 0.034). CONCLUSIONS: We conclude that apoptosis is important in the pathogenesis of GTD. Assessment of apoptotic activity in HM by the M30 index may be considered as an alternative prognostic indicator for predicting the clinical behaviour. PMID- 11260307 TI - Peripheral T/NK-cell lymphoma: a report of the IXth Workshop of the European Association for Haematopathology. AB - AIMS: In April 1998, The European Association for Haematopathology organized the IXth workshop on peripheral T-cell and NK-cell lymphomas and leukaemias. The workshop focused on unusual subtypes of these rare malignancies, allowing evaluation of the recently published WHO classification of neoplastic diseases of the lymphoid tissues. METHODS AND RESULTS: One-hundred and three cases were centrally immunophenotyped and hybridized for EBER1/2 of Epstein--Barr virus. All cases were reviewed by a panel of experienced haematopathologists and classified according to the new WHO classification for lymphoid neoplasms. Three cases were considered as precursor T-cell and 95 cases as peripheral T/NK-cell lymphoma/leukaemia. Although the cases represented a selected series of unusual cases, the following conclusions could be made: (i) Most lymphomas except the hepatosplenic gamma/delta T-cell lymphomas showed a rather broad morphological spectrum, with differences both between and within individual tumours. (ii) This heterogeneity was also reflected by the immunophenotype, for instance a variable expression of CD30 was found in many enteropathy type T-cell lymphomas. (iii) Exceptions in phenotype were regularly found in almost all categories, indicating that phenotype should not be the final determining factor in classification. (iv) The great majority of T-cell lymphomas expressed the alpha/beta T-cell receptor, with the exception of all but one hepatosplenic T-cell lymphomas and a few other extranodal peripheral T cell lymphomas. (v) Malignancies of precursor cells, blastic NK-cell lymphoma/leukaemia, adult T-cell lymphoma/leukaemia and most AIL type T-cell lymphomas did not express cytotoxic molecules such as TIA1 and granzyme-B. In contrast, all five aggressive NK/T-cell lymphomas/leukaemias, a single case of large granular lymphocyte leukaemia and 40 of 47 primary extranodal lymphoma/leukaemias expressed these molecules. In hepatosplenic gamma/delta T-cell lymphoma, five of six cases showed expression of TIA1 but not of granzyme-B. (vi) Seven tumours developed after organ-transplant, four cases being EBV-positive. No distinct phenotype could be attributed to these cases. CONCLUSIONS: Most peripheral T/NK cell lymphomas could be categorized as distinct entities as described in the recently proposed WHO classification for lymphoid neoplasms. PMID- 11260308 TI - Oestrogen and progesterone receptors in breast cancer: past, present and future. PMID- 11260309 TI - Websites review. PMID- 11260311 TI - Primary small cell carcinoma of the breast: report of a case and review of the literature. PMID- 11260312 TI - Spindle cell sarcoma with neuroepithelial [correction of neuroectodermal] differentiation. PMID- 11260313 TI - Apocrine ductal carcinoma in-situ with an unusual morphological presentation. PMID- 11260315 TI - Specificity, restriction and effector mechanisms of immunoregulatory CD8 T cells. PMID- 11260317 TI - Effect of early fetal splenectomy on prenatal B-cell development in sheep. AB - The contribution of early splenic B-cell populations to the colonization of the ileal Peyer's patch was investigated following the surgical removal of the spleen in a series of 56-day-old fetal sheep. The fetuses were killed at 140 days of gestation and the ileal Peyer's patch, the distal jejunal lymph node which drains the Peyer's patch, and a peripheral lymph node, the superficial cervical lymph node, were examined. Enzyme and immunohistochemical evaluation concluded that the distribution of B cells, T cells and stromal cells in the ileal Peyer's patch was similar in splenectomized and normal fetal sheep. Thus, the presence of the fetal spleen was not essential for the colonization of the ileal Peyer's patch and other early sites of B-cell accumulation would appear capable of generating the necessary precursor populations. Investigation of B-cell populations in lymph nodes used a combination of terminal deoxynucleotidyl-transferase-mediated deoxyuridine-triphosphate nick-end-labelling (TUNEL) histochemistry and immunofluorescence to determine the average number of apoptotic B cells in the primary follicles of the outer cortex of splenectomized and normal lambs. A significantly increased number of apoptotic B cells was present in the distal jejunal lymph node but not in the superficial cervical lymph node of splenectomized lambs. This finding suggests that splenectomy affected prenatal B cell development in fetal sheep and raises questions as to the regulation of B cell lymphopoiesis in a species using a post-rearrangement organ of diversification. PMID- 11260316 TI - Kinetics of GATA-3 gene expression in early polarizing and committed human T cells. AB - Different transcription factors have been shown to control the transition of naive T cells into T helper 1 (Th1)/Th2 subsets. The T-cell-specific transcription factor GATA-3 is known to be selectively expressed in murine developing Th2 cells and to exert a positive action on Th2-specific cytokine production. Investigating GATA-3 gene regulation in human T cells we have found that naive T cells highly express GATA-3, and during early T2 or T1 polarization, respectively, they either maintain or quickly down-regulate expression. In developing T2 cells, as well as in committed Th2 cell lines and clones, we found a positive correlation among GATA-3, interleukin (IL)-5 and IL-4 gene expression kinetics, supporting the positive action of GATA-3 on Th2-specific cytokine production. A possible relationship between GATA-3 gene expression and the down regulation of the IL-12 receptor (beta2-chain; IL-12Rbeta2) gene was evident only in the early phases of T2 polarization (within 24 hr), and not demonstrated at later times. During T-cell commitment the presence of IL-4 in the culture was essential to maintain or enhance GATA-3 transcription, while IL-12 was not necessary for full repression of GATA-3. Finally, we showed selective GATA-3 up regulation in human Th2 cell lines and clones and the maintainance of a low basal level of GATA-3 expression in Th1 cells upon activation. PMID- 11260318 TI - Dendritic cells rapidly undergo apoptosis in vitro following culture with activated CD4+ Valpha24 natural killer T cells expressing CD40L. AB - Human Valpha24 natural killer T (Valpha24NKT) cells are activated by alpha glycosylceramide-pulsed dendritic cells (DCs) in a CD1d-dependent and T-cell receptor-mediated manner. There are two major subpopulations of Valpha24NKT cells, CD4- CD8- Valpha24NKT and CD4+ Valpha24NKT cells. We have recently shown that activated CD4- CD8- Valpha24NKT cells have cytotoxic activity against DCs, but knowledge of the molecules responsible for cytotoxicity of Valpha24NKT cells is currently limited. We aimed to investigate whether CD4+ Valpha24NKT cells also have cytotoxic activity against DCs and to determine the mechanisms underlying any observed cytotoxic activity. We demonstrated that activated CD4+ Valpha24NKT cells [CD40 ligand (CD40L) -positive] have cytotoxic activity against DCs (strongly CD40-positive), but not against monocytes (weakly CD40-positive) or phytohaemagglutinin blast T cells (CD40-negative), and that apoptosis of DCs significantly contributes to the observed cytotoxicity. The apoptosis of DCs following culture with activated CD4+ Valpha24NKT cells, but not with resting CD4+ Valpha24NKT cells (CD40L-negative), was partially inhibited by anti-CD40L mAb. Direct ligation of CD40 on the DCs by the anti-CD40 antibody also induced apoptosis of DCs. Our results suggest that CD40-CD40L interaction plays an important role in the induction of apoptosis of DCs following culture with activated CD4+ Valpha24NKT cells. The apoptosis of DCs from normal donors, triggered by the CD40-CD40L interaction, may contribute to the homeostatic regulation of the normal human immune system, preventing the interminable activation of activated CD4+ Valpha24NKT cells by virtue of apoptosis of DCs. PMID- 11260319 TI - The human macrophage cell line U937 as an in vitro model for selective evaluation of mycobacterial antigen-specific cytotoxic T-cell function. AB - Despite strong evidence for CD8+ T-cell function in murine mycobacterial infections, their corresponding role in human tuberculosis has proven more difficult to demonstrate. We have evaluated the human macrophage (Mphi) cell line U937 as an in vitro model for human leucocyte antigen (HLA) class I-restricted presentation of mycobacterial antigens, as HLA class I is constitutively expressed at high levels by U937 cells in the absence of detectable HLA class II or CD1 molecules. U937 cells were evaluated for their ability to phagocytose Mycobacterium tuberculosis and for their ability to present mycobacterial antigens to human HLA class I-matched cytotoxic T lymphocytes (CTLs). Differentiated U937 cells were capable of efficient phagocytosis of M. tuberculosis but did not generate a subsequent respiratory burst response, and were permissive for intracellular growth of both bacillus Calmette-Guerin (BCG) and the virulent M. tuberculosis H37Rv strain. CTL activity was restricted to live mycobacterial organisms and was shown to be mediated by M. tuberculosis specific, HLA class I-matched, purified CD8+ CTL lines and CD8+ T-cell clones. Furthermore, M. tuberculosis-infected U937 targets were more rapidly and strongly lysed by CD8+ CTLs than were infected autologous Mphi. Finally, M. tuberculosis infected U937 cells simultaneously provided a sensitive indicator for detection of mycobacterial-specific, HLA-unrestricted gammadelta+ CTL activity. PMID- 11260320 TI - Primary human alveolar epithelial cells can elicit the transendothelial migration of CD14+ monocytes and CD3+ lymphocytes. AB - The ability of freshly isolated primary human alveolar epithelial cells (type II pneumocytes) to induce leucocyte migration across an endothelial monolayer was investigated. Three-way factorial analysis of variance (ANOVA) demonstrated that resting alveolar endothelial cells (AEC) could produce detectable quantities of monocyte chemoattractant protein 1 (MCP-1), which was upregulated in response to tumour necrosis factor-alpha (TNF-alpha) in a dose- and time-dependent fashion. Interferon-gamma (IFN-gamma) had no significant effect on this process. TNF-alpha and IFN-gamma both induced AEC to provoke migration of CD14+ monocytes and CD3+ lymphocytes across endothelium. IFN-gamma and TNF-alpha synergized in their ability to induce production of T lymphocyte, but not monocyte, chemoattractants from AEC. Leucocyte transendothelial migration was inhibited by anti-MCP-1 neutralizing antibody and by heparin, a polyanionic glycosaminoglycan (GAG). These data suggest that human AEC play a role in the multiple mechanisms that facilitate monocyte and T lymphocyte migration into the alveolar compartment of the lung under homeostasis and inflammatory conditions. One of these mechanisms is mediated via constitutive MCP-1 production by alveolar epithelial cells, which is upregulated by TNF-alpha. PMID- 11260321 TI - Capacity of mouse mast cells to prime T cells and to induce specific antibody responses in vivo. AB - Mouse, human and rat mast cells have been shown to express major histocompatibility complex II molecules and present antigens to specific T-cell hybridomas in vitro. The purpose of our investigation was to determine whether mouse mast cells are able to initiate specific immune responses in vivo. Induction of anti-dinitrophenyl (DNP) immunoglobulin G1 (IgG1) and IgG2a antibodies was performed by transferring ovalbumin (OVA)-DNP-pulsed bone marrow derived mast cells (BMMC), B cells, or macrophages into naive mice which were boosted later with soluble antigen. Cultured spleen cells from immunized mice were tested for their cytokine content. Our data show that mast cells were by far better inducers of anti-DNP IgG1 antibodies than were B cells and macrophages. In contrast, anti-DNP IgG2a response induced by macrophages was much stronger than that obtained with mast cells whereas B cells were completely unable to elicit this response. In addition to a high index of cell proliferation, spleen cells from mast cell-injected mice produced more interferon-gamma than those mice who received macrophages or B cells by two- to fivefold, and almost 10-fold, respectively. Mast cell-deficient Wf/Wf mice were compared with their normal +/+ littermates and with mast cell-reconstituted Wf/Wf mice to develop delayed-type hypersensitivity (DTH) reactions as well as humoral immune responses. Mast cell sufficient mice as well as mast cell-reconstituted Wf/Wf mice developed significantly increased DTH reactions (P = 0.02, and 0.03, respectively) and higher anti-OVA-specific antibody responses as compared with Wf/Wf mice. Our data suggest that mast cells have the potential to up-regulate both humoral and cellular immune responses in vivo. PMID- 11260322 TI - The unusual distribution of the neuronal/lymphoid cell surface CD200 (OX2) glycoprotein is conserved in humans. AB - OX2 (CD200) is a type-1 membrane glycoprotein that contains two immunoglobulin superfamily domains and which is expressed on a variety of lymphoid and non lymphoid cells in the rat. The recent characterization of a receptor for OX2 (OX2R) on myeloid cells, and the phenotype of an OX2-deficient mouse, suggests that OX2 may regulate myeloid cell activity in anatomically diverse locations. Here we report the tissue distribution of the human homologue of the rat OX2 glycoprotein using a new monoclonal antibody (mAb), OX104, raised against recombinant human OX2. Human OX2 was expressed at the cell surface of thymocytes, B cells, T cells, neurons, kidney glomeruli, tonsil follicles, the syncytiotrophoblast and endothelial cells. This broad, but not ubiquitous, distribution pattern is very similar to that observed in rats, suggesting that OX2 may regulate myeloid cell activity in a variety of tissues in humans. PMID- 11260323 TI - Structural and functional analysis of the human CD45 gene (PTPRC) upstream region: evidence for a functional promoter within the first intron of the gene. AB - Expression of the leucocyte common antigen (CD45) in mammals is restricted to the nucleated lineages of haematopoietic cells. It appears in early progenitors in the bone marrow and is expressed at the surface of these cells throughout their differentiation. However, at least in T cells, the pattern of expression switches between different isoforms during the successive stages of differentiation in the thymus and after activation in the periphery. In order to understand the mechanisms controlling the transcription of the human CD45 gene, 2.7 kbp of the 5'-flanking region were sequenced and analysed for their ability to direct expression of a reporter gene. The only region with promoter activity was localized within the first intron of the gene. This promoter shows no tissue specificity but could be enhanced by a heterologous enhancer. Mobility shift assays showed complex but specific protein binding. The sequence in this region lacks similarity with known promoters or initiators but is highly conserved in evolution. No transcription initiation could be detected within or downstream of this region, suggesting that this might be a new type of RNA polymerase II promoter able to drive transcription from an upstream sequence. An additional exon was also found upstream of exon 1. The two exons 1 (1a and 1b) are mutually exclusive and both are spliced to exon 2. This makes the structure of the 5' region of the human CD45 gene identical to its mouse homologue. PMID- 11260324 TI - Efficacy of cytokine gene transfection may differ for autologous and allogeneic tumour cell vaccines. AB - Whole tumour cells are a logical basis for generating immunity against the cancers they comprise or represent. A number of human trials have been initiated using cytokine-transfected whole tumour cells of autologous (patient-derived) or allogeneic [major histocompatibility complex (MHC)-disparate] origin as vaccines. Although precedent exists for the efficacy of autologous-transfected cell vaccines in animal models, little preclinical evidence confirms that these findings will extrapolate to allogeneic-transfected cell vaccines. In order to address this issue a murine melanoma cell line (K1735) was transfected to secrete interleukin (IL)-2, IL-4, IL-7 or granulocyte-macrophage colony-stimulating factor (GM-CSF); cytokines currently in use in trials. The efficacy of these cells as irradiated vaccines was tested head-to-head in syngeneic (C3H) mice and in MHC-disparate (C57BL/6) mice, the former being subsequently challenged with K1735 cells and the latter with naturally cross-reactive B16-F10 melanoma cells. Whilst the GM-CSF-secreting vaccine was the most effective at generating protection in C3H mice, little enhancement in protection above the wild-type vaccine was seen with any of the transfections for the allogeneic vaccines, even though the wild-type vaccine was more effective than the autologous B16-F10 vaccine. Anti-tumour cytotoxic T-lymphocyte (CTL) activity was detected in both models but did not correlate well with protection, whilst in vitro anti-tumour interferon-gamma (IFN-gamma) secretion tended to be higher following the GM-CSF secreting vaccine. Cytokine transfection of vaccines generally increased anti tumour CTL activity and IFN-gamma secretion (T helper type 1 response). Further studies in other model systems are required to confirm this apparent lack of benefit of cytokine transduction over wild-type allogeneic vaccines, and to determine which in vitro assays will correlate best with protection in vivo. PMID- 11260325 TI - Immunological memory in B-cell-deficient mice conveys long-lasting protection against genital tract infection with Chlamydia trachomatis by rapid recruitment of T cells. AB - The role of antibodies and antigen deposition for the development of immunological memory has been incompletely investigated. We addressed whether long-term protection and T-cell memory can be stimulated against a genital tract infection with human Chlamydia trachomatis serovar D in B-cell-deficient (muMT) mice. At 6 months following a primary infection with C. trachomatis, both muMT and wild-type (WT) mice exhibited strong and comparable protection against reinfection. Evidence of long-lasting CD4+ T-cell memory was found in both muMT and WT mice, typified by comparable delayed-type hypersensitivity (DTH) reactions against chlamydial antigens. No bacterial or chlamydial DNA was found in the genital tract of muMT memory mice, suggesting that immunological memory was maintained in the absence of antigen. Whereas few T cells were present in the genital tract of memory mice, rapid recruitment of CD4+, and some CD8+, T cells into the genital tract tissue was observed after challenge with live bacteria. Accumulation of T cells in the genital tract was preceded by a short transient infection of similar magnitude in both muMT and WT memory mice, arguing against a long-term protective role of local antibodies. The rapid recruitment of CD4+ T cells into the genital tract was associated with a transient detection of interferon-gamma (IFN-gamma) mRNA in the genital tract in chlamydia-immune memory mice, which was not found in naive, challenged mice. Thus, long-term protection in the genital tract against C. trachomatis infection is conveyed by IFN-gamma producing CD4+ memory T cells, which appear to be maintained in the absence of antibodies and local antigen deposition. PMID- 11260326 TI - Most parasite-specific CD8+ cells in Trypanosoma cruzi-infected chronic mice are down-regulated for T-cell receptor-alphabeta and CD8 molecules. AB - The present study shows that CD8+ T lymphocytes expressing low levels of T-cell receptor (TCR)alphabeta, CD8 and CD3 accumulate in the spleen, blood, peritoneum and liver, but not in the lymph nodes of mice chronically infected with Trypanosoma cruzi. Analysis of spleen lymphocytes reveals that most CD8LOW TCRLOW T cells have an experienced phenotype (CD44HIGH CD62LLOW and CD45RA,B,CLOW). These cells have small size, lack activation markers such as CD69, CD25 and CD11b (Mac-1), and do not spontaneously secrete cytokines, suggesting they are at the resting state. When stimulated in vitro with T. cruzi-infected macrophages, TCRLOW CD8LOW T cells behave as parasite-specific memory cells, readily responding with interferon-gamma (IFN-gamma) production. Indeed, among parasite activated CD8+ lymphocytes, IFN-gamma production was mostly due to TCRLOW CD8LOW cells. Upon in vitro stimulation with anti-CD3/CD28 monoclonal antibodies, down regulated cells produce IFN-gamma and tumour necrosis factor-alpha, but not interleukin IL-10 or IL-4. Our results indicate that despite parasite persistence, most T. cruzi-specific experienced CD8+ cells are resting. Nevertheless, when encountering infected macrophages these cells differentiate to Tc1 effectors. PMID- 11260327 TI - Impaired protective immunity and T helper 2 responses in alymphoplasia (aly) mutant mice infected with Trichinella spiralis. AB - The alymphoplasia (aly) mutation of mice prevents the development of systemic lymph nodes and Peyer's patches. The mutant homozygotes (aly/aly) are partially deficient in both humoral and cell-mediated immune functions. In the present study, we show that adult worm expulsion was slightly delayed and that T helper 2 (Th2)-type responses were partially defective in aly/aly mice after infection with Trichinella spiralis. Male aly/aly and aly/+ mice (8-weeks old) were infected with 400 muscle larvae. There was no difference in worm recovery between the two groups on day 5. However, worm recovery in aly/aly mice was significantly higher than that in aly/+ mice on day 14. Mucosal mast cells increased in number and peaked 14 days after infection in aly/+ mice. aly/aly mice were deficient in their mucosal mast cell response through out the primary infection. To examine the existence of mast cell precursors, aly/aly mice were treated with recombinant interleukin-3 (rIL-3) before infection. The mast cell response was poorly induced in aly/aly mice treated with rIL-3. An immunoglobulin E (IgE) response was not detected in aly/aly mice during the course of infection. Serum IgG1 levels in aly/aly mice were significantly lower than that of aly/+. The serum IgG2a levels increased in both strains of mice. However, IgG2a production in aly/aly mice on day 14 was half as much as that in aly/+mice. Stimulation of splenic T cells in vitro with anti-CD3 monoclonal antibody (mAb) showed that spleen cells from aly/+ mice on day 5 produced more IL-4 than spleen cells from aly/aly mice. IL-4 production from aly/aly mice on day 14 was half that from aly/+ mice. Interferon gamma (IFN-gamma) was produced in both aly/aly and aly/+ mice on day 14. Proliferation assay showed that T cells of aly/aly mice responded poorly when cultured with antigen-presenting cells. These results suggest that aly gene is needed for the induction of protective immunity and Th2 responses in mice infected with T. spiralis. PMID- 11260328 TI - The ability of heat-killed Mycobacterium vaccae to stimulate a cytotoxic T-cell response to an unrelated protein is associated with a 65 kilodalton heat-shock protein. AB - Exogenous antigens are generally presented by Class II major histocompatibility (MHC) molecules. When administered with an adjuvant, however, they are capable of inducing a CD8+ T-cell response where antigen recognition is associated with Class I MHC. Accordingly, immunization with soluble ovalbumin (OVA) alone does not activate CD8+ cytotoxic T cells (CTL) but when given in complete Freund's adjuvant (CFA), or in formulations of a number of novel adjuvants, an OVA specific CD8+ CTL response can be detected. We show in this report that immunization with soluble OVA mixed with heat-killed Mycobacterium vaccae, but not with other common pathogenic and saprophytic mycobacteria, can activate OVA specific CD8+ CTL. An OVA-specific CTL response is detected when mice are immunized by either the intraperitoneal or intranasal route and their spleen cells are re-stimulated in vitro. Adjuvant activity of heat-killed M. vaccae is present in M. vaccae culture filtrate, in soluble protein components of whole M. vaccae and in the 65 kDa heat-shock protein (hsp) of M. vaccae. Mycobacterium vaccae has previously been shown to have no adverse side-effects in humans. The current results suggest that M. vaccae may be useful as an adjuvant for vaccines and other immunotherapies where CD8+ CTL responses to exogenous proteins are crucial. PMID- 11260329 TI - Induction of in vivo resistance to Mycobacterium avium infection by intramuscular injection with DNA encoding interleukin-18. AB - Interferon-gamma (IFN-gamma) is closely associated with the generation of cell mediated immunity and resistance to intracellular parasites. Interleukin-18 (IL 18) was known to strongly induce IFN-gamma production by T cells and natural killer (NK) cells. In order to determine whether injection with DNA encoding IL 18 can stimulate the resistance to Mycobacterium avium complex (MAC) infection, the mature IL-18 cDNA with kappa leader sequence was cloned under control of the cytomegalovirus (CMV) promoter (TcCMVIL-18) and its effect on MAC infection was investigated in genetically susceptible BALB/c mice. Injection with the TcCMVIL 18 DNA during intranasal infection with MAC resulted in a significant decrease in bacterial load of lung during the entire 8-week observation period, while injection with the TcCMV control DNA did not. Lung cells in mice injected with the TcCMVIL-18 DNA showed persistent production of IFN-gamma throughout the 8 week period. Furthermore, immunization with the TcCMVIL-18 DNA induced and maintained significantly higher levels of cytotoxic activity and nitric oxide production by lung cells than immunization with the TcCMV control vector. This work suggests that IL-18 DNA vaccination may be useful in the immunotherapeutic or immunoprotection approaches of infections by intracellular parasites such as mycobacteria. PMID- 11260330 TI - The failure of Daudi cells to express the cellular prion protein is caused by a lack of glycosyl-phosphatidylinositol anchor formation. AB - The cellular prion protein (PrPc) is a glycosyl-phosphatidylinositol (GPI)-linked cell surface protein, which is expressed at high density on nervous tissues and at lower levels on most other solid-organ tissues. It is also expressed on peripheral blood mononuclear cells (PBMC) of all lineages. In lymphocytes, its level of expression is dependent upon the state of cell activation, and polyclonal anti-PrP antisera partially block lectin-induced T-cell activation, suggesting a functional role of the protein in this process. Using the monoclonal antibody (mAb) 3F4 we examined PrPc surface immunoreactivity on leukaemic cell lines of T- and B-cell origin, and unexpectedly observed a complete lack of PrPc cell-surface expression in Daudi cells, while all other cell lines displayed discernible reactivity. We demonstrated the intracellular presence of PrP specific mRNA and PrP protein. The lack of surface PrPc is unrelated to the well known defect of beta2-microglobulin (beta2m) expression in Daudi cells as other beta2m-deficient cells, such as the melanoma cell line F0-1 and spleen cells from beta2m gene-deleted mice, were not deficient in cell-surface PrPc. Daudi cells failed to bind antibodies directed against all GPI-linked cell surface proteins. In somatic hybridization experiments using murine spleen cells as partners, we observed de novo expression of human PrPc, CD55 and CD59, thus demonstrating in Daudi cells the availability of these gene products for GPI linkage and cell surface expression. PMID- 11260331 TI - Pentoxifylline treatment of mice with chronic pulmonary tuberculosis accelerates the development of destructive pathology. AB - It is well established in animal models that production of the cytokine tumour necrosis factor-alpha (TNF-alpha) is essential to the proper expression of acquired specific resistance following infection with Mycobacterium tuberculosis. This gives rise to an apparent state of chronic disease which over the next 100 200 days is characterized by slowly worsening pathological changes in the lung. To determine whether continued TNF-alpha production was harmful during this phase mice were treated with a TNF-alpha inhibitor, pentoxifylline. It was observed that although this therapy did not alter the numbers of bacteria recovered from the lungs of the infected mice, tissue damage within the lung was accelerated. These data thus demonstrate that production of TNF-alpha, already known to be important during the early expression of resistance to tuberculosis, remains important and beneficial during the chronic stage of the disease. PMID- 11260332 TI - Assessment of renal function with color Doppler ultrasound in autosomal dominant polycystic kidney disease. AB - BACKGROUND: Measurement of renal blood flow by color Doppler ultrasound is useful for assessment of renal function in a variety of renal disorders. In autosomal dominant polycystic kidney disease (ADPKD), however, it might be difficult to visualize interlobar arteries because of deformity of renal structure. To evaluate the usefulness of color Doppler in ADPKD, parameters determined by blood flow examination were compared with the results of ordinal renal function tests. METHODS: Twenty-one patients with ADPKD were examined by color Doppler ultrasound measurement. In each patient, 10 interlobar arteries in both kidneys were investigated. Minimum blood flow velocity (Vmin), maximum blood flow velocity (Vmax), mean blood flow velocity (Vmean), acceleration, resistive index and pulsatility index were measured in relation to the results of creatinine clearance, serum creatinine, blood urea nitrogen and 15 and 120 min values of the phenolsulfonphthalein test. RESULTS: In all patients, interlobar arteries were able to be visualized and blood-flow profile was measured. Although variations of Vmin, Vmax, Vmean and acceleration were relatively large, the resistive index and pulsatility index varied little in each kidney. Mean values of Vmin (P < 0.005), Vmean (P < 0.05), resistive index (P < 0.005) and pulsatility index (P < 0.005) were well correlated to creatinine clearance with statistical significance. CONCLUSIONS: In ADPKD, color Doppler ultrasound measurement is a useful method for assessment of renal function and could be used for monitoring the dynamic state of renal blood flow as a non-invasive technique. PMID- 11260333 TI - Transurethral needle ablation of the prostate: an initial Japanese clinical trial. AB - OBJECTIVES: Transurethral needle ablation of the prostate is a new alternative endoscopic thermal therapy that uses a low-energy radio frequency delivered into the prostatic adenoma. Herein is reported the initial clinical experience by multiple institutes in Japan of transurethral needle ablation of the prostate for the treatment of symptomatic benign prostatic hyperplasia. METHODS: A total of 93 patients were treated with this technique. Transurethral needle ablation of the prostate was generally performed under low-spinal anesthesia. Before and after the procedure, international symptom score (IPSS), quality of life (QOL) score, peak urinary flow rate (Qmax), postvoid residual urine volume and prostate size were evaluated. RESULTS: There was a reduction of IPSS of more than 50% when compared with that of pretreatment, being 51.3% (57/93 patients) and 60.2% (56/93 patients) at 3 months and 6 months after the procedure, respectively. Sixty-seven patients who were available for a 12-month follow-up period demonstrated a markedly decreased mean IPSS when compared with that measured before the treatment for a statistically significant difference (P < 0.01). Fifty-eight patients who were available for uroflowmetric study at 12 months exhibited a notably increased mean Qmax of 11.2 +/- 4.5 mL/s, which was a statistically significant increase when compared with that found before treatment (P < 0.05). Although all patients suffered some degree of gross hematuria after the procedure, none of them required any specific treatment for complications. CONCLUSION: Transurethral needle ablation technique for the treatment of symptomatic benign prostatic hyperplasia is safe and effective. However, a much longer follow-up study is essential for fully evaluating the extended effectiveness of this technique. PMID- 11260334 TI - Are tobacco use and urine pH indicated as risk factors for bladder carcinoma? AB - BACKGROUND: Many case-control and cohort studies have shown a positive relationship between bladder carcinoma and tobacco use. Recently, urine pH has been reported to influence aromatic amine carcinogenesis, which have been implicated as potent carcinogens in bladder carcinoma patients. Herein the correlation between bladder carcinoma, tobacco use and urine pH is reported. METHOD: One hundred and forty-one patients with bladder carcinoma and 128 patients with benign prostatic hyperplasia or urolithiasis as controls were selected. All patients were admitted to Osaka City University Hospital for the purpose of surgical treatment. Urine pH was checked by a test tape. RESULTS: Of the patients with bladder carcinoma, 106 were smokers and 35 were non-smokers. In contrast, the number of smokers in the control group was 76 and that of non smokers was 52. The odds ratio in the bladder carcinoma group calculated for the smoker patients was 2.07, showing a significant correlation between bladder carcinoma and tobacco use. Regarding urine pH, acidic urine was found in 126 patients in the bladder carcinoma group and in 116 patients in the control group. The odds ratio in the bladder carcinoma group for acidic urine was 0.87, showing no significant relationship between bladder carcinoma and urine pH. CONCLUSION: The study found a positive relationship between bladder carcinoma and tobacco use; however, it could not establish a clear relationship between bladder carcinoma and urine pH, even in the smoker group. PMID- 11260335 TI - Direct screening of the IMP-1 metallo-beta-lactamase gene (blaIMP) from urine samples by polymerase chain reaction. AB - BACKGROUND: As Gram-negative bacterial isolates producing plasmid-mediated IMP-1 metallo-beta-lactamase usually demonstrate resistance to various broad-spectrum beta-lactams, including cephamycins and carbapenems, transmission and proliferation of these microorganisms in clinical settings could become a clinical threat in the near future. According to previous studies by the same authors, IMP-1-producing strains are usually isolated from urine samples. Therefore, in this study, a polymerase chain reaction (PCR) was applied for direct screening of the IMP-1 metallo-beta-lactamase gene in urine samples. METHOD: Urine samples were collected from 273 inpatients to whom various broad spectrum beta-lactams, including carbapenems, had been administered in 57 hospitals in 1997. DNA templates for PCR analyses were prepared directly from 19 urine samples from which Serratia marcescens strains demonstrating high-level resistance (minimal inhibitory concentration > 128 microg/mL) to both ceftazidime and cefoperazone-sulbactam were later isolated. RESULTS: The IMP-1 metallo-beta lactamase gene (blaIMP)-specific 578 bp fragments were able to be amplified successfully in eight of the 19 samples. In the seven strains isolated from the eight samples, the presence of blaIMP was also detected by a DNA hybridization analysis. The lower limit of the PCR method was determined as 1 x 10(2) CFU of blaIMP-bearing bacterial cells per 1 mL of urine sample. No false positive result was found. CONCLUSION: The PCR-aided direct screening of blaIMP is applicable to early recognition of IMP-1-producing bacteria in urine samples. This method would help to prevent nosocomial and interhospital transmission of this kind of hazardous bacteria, as well as the advancement of rigorous infection control. PMID- 11260336 TI - Immunohistochemical expression of P-glycoprotein in the rat urinary bladder and the effect of verapamil on intravesical chemotherapy. AB - BACKGROUND: The expression of P-glycoprotein (Pgp) is thought to be common in bladder epithelium and the multidrug resistance mediated by Pgp must be considered to improve the efficacy of chemotherapy for bladder tumors. METHODS: The expression of Pgp in normal and tumor tissue of the rat urinary bladder was first examined immunohistochemically. The effect of verapamil, an expected modulator of Pgp, on intravesical chemotherapy of the rats was then investigated. RESULTS: Pgp was immunohistochemically detected in normal epithelium and in tumor tissue of the rat urinary bladder. In those normal and tumor-bearing bladders, verapamil promoted the uptake of intravesically instilled pirarubicin, but the efflux of intracellular accumulated pirarubicin was observed subsequently in both conditions with and without verapamil. The drug concentration decreased more rapidly in the verapamil group than in the control group. CONCLUSIONS: Verapamil is thought to be useful in promoting uptake of intravesically instilled pirarubicin, but it did not appear to be so efficient at limiting the efflux of intracellular accumulated pirarubicin. PMID- 11260337 TI - Inhibition of calcium oxalate precipitation by bile salts. AB - BACKGROUND: Both urinary and biliary stones can contain calcium. Bile salts (BA), which are known to bind Ca2+, are commonly used to dissolve the latter but not the former. METHODS: The effect of physiologic BA on calcium oxalate (CaOx) precipitation was evaluated by a recently developed method. RESULTS: The Ca2+ binding properties of BA were confirmed by small but significant decreases in pH observed following addition of CaCl2 to bile acids solutions. More importantly, BA inhibited CaOx precipitation with effective concentrations of approximately 10 3 mol/L. The clinical relevance of the latter observation is presently unknown but it is of note that in the same in vitro assay, the activity of BA appeared comparable to that of citric acid, the most common drug for urolithiasis. Although BA do not reach mmol/L levels in urine, they are known to change the physicochemical properties of this fluid, possibly slowing down the crystal growth process. However, the hypothetical therapeutic use of BA in former stone patients would present at least two major problems: (i) hepatotoxicity and (ii) lithogenic activity, due to hyperoxaluria subsequent to increased intestinal absorption of oxalate. CONCLUSION: The ability of BA in effectively binding calcium ions and in inhibiting the precipitation of CaOx appears of interest from both a physiopathologic and a pharmacologic point of view, even if it does not currently seem exploitable for prophylactic or therapeutic purposes. PMID- 11260338 TI - Ureteroarterial fistula in a patient with a single functioning kidney. AB - A rare case is reported of a fistula between the left internal iliac artery and the left ureter in a patient whose left kidney was the only functioning kidney. Internal iliac artery embolization was initially successful in stopping the bleeding, but the fistula recurred when the ureteral stent was removed. Even after embolization, the tissue surrounding a fistula remains very fragile, so the fistula may easily recur as a result of slight injury or inflammation. PMID- 11260339 TI - Primary retroperitoneal ganglioneuroblastoma in an adult. AB - A case of retroperitoneal ganglioneuroblastoma in a 60-year-old man is reported. This retroperitoneal tumor was surgically removed and pathologic diagnosis was ganglioneuroblastoma. Ganglioneuroblastoma usually occurs in children and is extremely rare in adults. The characteristics are described of an unusual tumor based on the published reports. PMID- 11260340 TI - Advanced adenocarcinoma of the urinary bladder successfully treated by the combination of cisplatinum, mitomycin-C, etoposide and tegafur-uracil chemotherapy. AB - Primary adenocarcinoma of the urinary bladder has shown an extremely poor response to radiation or chemotherapy. Therefore, radical surgery is the only therapeutic treatment for it. A case report is presented of a primary advanced adenocarcinoma of the urinary bladder invaded into the uterus with distant metastases which responded completely to systemic combination chemotherapy including tegafur-uracil. The patient was a 53-year-old woman with a history of asymptomatic macrohematuria. She was treated with the combination of cisplatinum, mitomycin-C, etoposide and tegafur-uracil chemotherapy. After four courses of the chemotherapy, computed tomography showed marked regression of the primary tumor of the urinary bladder and the complete disappearance of the distant metastases in the liver, lung and para-aortic lymph node. Subsequently, she underwent radical cystectomy and cutaneous ureterostomy. Pathologically, no viable cancer cells were detected. Three years after the operation, she has no evidence of disease recurrence. Treatment of advanced adenocarcinoma of the urinary bladder by this combination chemotherapy is of benefit. PMID- 11260341 TI - Solitary fibrous tumor of the prostate. AB - An extremely rare case of solitary fibrous tumor of the prostate is presented. The patient underwent a radical retropubic prostatectomy and has remained well with no evidence of recurrence during the last 18 months. This is the fifth reported case of such a lesion arising in the prostate. PMID- 11260342 TI - Primary testicular plasmacytoma with hydrocele of the testis. AB - A case of primary testicular plasmacytoma complicated with hydrocele of the testis is reported. An 86-year-old man presented with hydrocele of the right testis. High inguinal orchiectomy was performed as the preoperative aspiration cytology of the hydrocele fluid showed atypical cells. Immunohistochemical study of the right testis revealed testicular plasmacytoma positive for IgG. He remained well 9 months after the orchiectomy. This is the second reported case where the preoperative diagnosis of testicular plasmacytoma was made based on the hydrocele fluid cytology. PMID- 11260343 TI - Anterior urethral valve as a cause of end-stage renal disease. AB - Although posterior urethral valves are predominant as a cause of obstructive uropathy in children, anterior urethral valves may also appear as the underlying etiologic factor in end-stage renal disease that results from obstruction. Two cases are presented of anterior urethral valve patients that were admitted with end-stage renal disease. The first case was successfully treated with diverticulectomy and urethral reconstruction in preparation for renal transplantation. The second case, however, had been on cystostomy drainage for 6 years and also had a contracted bladder. A more extensive lower urinary tract reconstruction was delayed. Children with poor stream and recurrent infections should be evaluated carefully and anterior urethral valve or diverticula should be considered in differential diagnosis of obstructive lesions. PMID- 11260344 TI - Voiding dysfunction in a patient with adolescent adrenoleukodystrophy. AB - The details are reported of bladder dysfunction in a Japanese boy with adrenoleukodystrophy. He developed gait disturbance at the age of 15 years. Spastic paraparesis progressed from the legs to the hands and brain magnetic resonance imaging showed characteristic degenerative change. Detrusor hyperreflexia was found by a urodynamic study and detrusor-sphincter dyssynergia was also suspected. PMID- 11260345 TI - Acute normovolemic hemodilution for radical prostatectomy: can it replace preoperative autologous blood transfusion? AB - BACKGROUND: Although preoperative autologous blood donation (PAD) is accepted as a standard of care for radical prostatectomy, it is costly, time-consuming and has risks associated with blood storage. Acute normovolemic hemodilution (ANH) is reported to be less expensive and to preserve blood components more effectively than PAD. In the present study, the efficacy and safety of these two autologous blood-collection techniques were compared. METHODS: The study included 16 consecutive patients scheduled for radical prostatectomy. The first eight patients underwent conventional preoperative autologous blood donation of 400 mL 1 week before the operation (PAD group) and the second eight patients underwent acute normovolemic hemodilution followed by immediate operation (ANH group). All blood collected was transfused in the perioperative period. Preoperative and postoperative hematocrit levels in these two groups were compared. RESULTS: There were no differences in preoperative hematocrit, time of operation or operative blood loss between the two groups. In the ANH group, 1080 +/- 160 mL of blood were collected. The postoperative hematocrit level did not differ significantly between the groups. No patient in either group received allogeneic blood transfusion or experienced an adverse event directly related to blood transfusion. CONCLUSION: The two blood-conservation strategies resulted in similar postoperative hematologic outcomes. Given its advantages, which include lower cost, lower risk and higher convenience, ANH is one of the procedures that may replace conventional PAD for use in radical prostatectomy. PMID- 11260346 TI - An anterior urethral stitch improves urinary incontinence following radical prostatectomy. AB - OBJECTIVE: To determine the effects of an anterior urethral stitch, referred to as an endopelvic anterior urethral stitch (EAUS), in reducing recovery time for post-prostatectomy urinary incontinence. METHODS: The urinary continence recovery time for 24 patients, who received a retropubic radical prostatectomy with the EAUS procedure, was compared to that of a historical control series of 22 patients without EAUS. The EAUS is a simple 2-0 polyglactin stitch placed between the bunched dorsal vein complex and the anterior urethra. This procedure was performed at the time of urethro-vesical anastomosis. Continence recovery time was defined as the day after removal of the urethral catheter when the patient no longer required pads for incontinence. RESULTS: A significantly shorter time for continence recovery (median 8.5 days) was obtained in EAUS patients compared with that of the control series (median 72 days) (P < 0.0001). Early recovery of continence was observed in 12/24 patients (50%) within 1 week and 18/24 patients (75%) within 1 month in EAUS patients. No adverse effects or complications were observed in the EAUS patients. CONCLUSION: A surgical procedure, the EAUS, has been developed that reduces urinary incontinence in patients who have undergone a radical prostatectomy. This procedure is simple and quick and improves recovery of continence without any side-effects. PMID- 11260347 TI - Recovery of sexual function after nerve-sparing radical prostatectomy or cystectomy. AB - BACKGROUND: The recovery of sexual function (erectile function and frequency of sexual intercourse) over time after nerve-sparing radical prostatectomy or cystoprostatectomy was evaluated. METHODS: Forty-nine consecutive patients with clinically localized prostate cancer and muscle-invasive bladder cancer were treated with radical prostatectomy and radical cystoprostatectomy with a nerve sparing procedure. Erectile function was evaluated by the circumferential change of the penis during nocturnal penile tumescence (NPT value) with an erectometer before and after surgery. Erectile function and the frequency of sexual intercourse were also evaluated with a self-administered questionnaire before and after surgery. Multivariate analysis by Cox's proportional hazards model was used to evaluate the factor(s) that affected the recovery of erectile function and sexual intercourse. RESULTS: The recovery rates of erectile function were 49% at 3 years and 79% at 5 years. For recovery of sexual intercourse the rates were 36% at 3 years and 57% at 5 years. Multivariate analysis revealed that the preoperative NPT value was the only independent factor which significantly affected the recovery of erectile function. The age at surgery was a significant factor for recovery of sexual intercourse. CONCLUSION: Nerve-sparing operations can often, but not always, provide preservation or recovery of erectile function for patients who receive radical prostatectomy or cystoprostatectomy. Recovery of erectile function depends upon the preoperative NPT value and recovery of sexual intercourse depends upon the age of the patient. PMID- 11260348 TI - One-stage repair of moderately severe hypospadias using a transverse preputial tubularized island flap. AB - BACKGROUND: Transverse preputial tubularized island flap (TPTIF) urethroplasty has been used for the repair of moderately severe hypospadias since Duckett described the procedure in 1980. In spite of the excellent results reported by Duckett, subsequent studies showed high complication rates. A TPTIF procedure modified to reduce the complication rate is presented. METHODS: Between 1996 and 1997, 13 boys with moderately severe hypospadias were repaired with the TPTIF procedure. Patient age ranged from 10 months to 3 years with an average age of 23 months. To prevent urethrocutaneous fistula, the neourethra was constructed with a two-layer closure and the portion of anastomosis was wrapped between the native urethra and the neourethra with the tissue of the corpus spongiosum. RESULTS: The moderately severe hypospadias was repaired without complication in 12 of 13 patients. A urethrocutaneous fistula developed at the midshaft of the penis in one patient. No meatal stenosis, urethral stricture or diverticulum developed. CONCLUSION: Transverse preputial tubularized island flap urethroplasty provided excellent cosmetic and functional results for moderately severe hypospadias, and postoperative complications could be decreased by the two-layer closure of the neourethra and application of the wrapping technique of the proximal anastomosed portion with corpus spongiosum tissue. PMID- 11260349 TI - Urothelial mucosal concentration of levofloxacin administered before transurethral resection: Is the mucosal concentration predictable? AB - BACKGROUND: Although it is an established surgical technique, transurethral resection (TUR) is associated with a certain incidence of postoperative bacteriuria. Assessment was made whether the urothelial mucosal concentration of an antibiotic administered before TUR was high enough to decrease the incidence of urinary tract infection (UTI). Also investigated were factors predicting the organ concentration. METHODS: Forty-nine patients (45 men and four women aged 51 79 years with a median age of 70 years) who underwent TUR between August 1996 and September 1997 were enrolled in the study. Each patient received 200 mg of levofloxacin (LVFX) about two hours before surgery. Blood and bladder urine were collected and urothelial mucosa was harvested at the time of TUR. Then the LVFX concentration in these samples was measured using high-performance liquid chromatography. The association between drug levels, or the ratio to the serum concentration, and factors likely to affect the vascular system that delivers the drug (age, bodyweight, blood pressure, pulse rate, total cholesterol and diabetes mellitus) were investigated. RESULTS: The mean serum drug level was 2.4 microg/mL, and it was 206.4 microg/mL in the urine and 5.7 microg/mL in the urothelial mucosa. The mean ratio of the mucosal to serum concentrations was 2.6. The urinary drug concentration showed no association with any of the factors assessed, while the serum concentration decreased with increasing bodyweight (P = 0.03). As the diastolic blood pressure increased, both the mucosal drug concentration and the mucosa/serum ratio decreased (P < 0.01). When the relationship between the serum and mucosal concentrations was investigated, no correlation was found. However, the mucosa/serum ratio (indicating the transfer of LVFX from the blood) was positively correlated with the mucosal concentration. CONCLUSION: Preoperative administration of LVFX was demonstrated to have potential value for the prophylaxis of UTI after TUR. Both the mucosal concentration and the mucosa/serum ratio were correlated with the diastolic blood pressure. As the diastolic blood pressure seems to be an indicator of the tissue concentration of LVFX, it may be possible to set the optimum dose based on the diastolic pressure. PMID- 11260350 TI - Effect of KMD-3213, an alpha1A-adrenoceptor antagonist, on the prostatic urethral pressure and blood pressure in male decerebrate dogs. AB - BACKGROUND: KMD-3213 is an alpha1A-adrenoceptor-selective antagonist currently being developed for the treatment of urinary outlet obstruction in patients with benign prostatic hyperplasia. In the present study, the uroselectivity of KMD 3213 was evaluated and compared with that of prazosin and tamsulosin in a decerebrate dog model. METHODS: Intercollicular decerebration was carried out in male mongrel dogs under anesthesia. The inhibitory effects of intravenously and intraduodenally administered compounds on the increase in intraurethral pressure (IUP) induced by electrical stimulation of the hypogastric nerve were estimated. Systemic blood pressure was measured simultaneously. RESULTS: The alpha1 antagonists tested produced a dose-dependent inhibition of the induced IUP response and decreased mean blood pressure (MBP). The ID50 of KMD-3213, tamsulosin and prazosin for IUP (dose required to inhibit the increase in IUP by 50%) was 3.15, 1.73 and 11.8 microg/kg i.v., respectively, and the ED20 for the hypotensive effect (dose required to reduce MBP by 20%) was 8.03, 0.59 and 2.46 microg/kg i.v., respectively. The data indicate that uroselectivity (ED20/ID50) of KMD-3213 is 12- and 7.5-fold higher than that of prazosin and tamsulosin, respectively. When the drugs were administered intraduodenally, KMD-3213 was sufficiently absorbed from the digestive tract and continued to demonstrate at least 3.8-fold higher uroselectivity than tamsulosin. CONCLUSION: Based on these findings, KMD-3213 appears to be an effective orally active compound for decreasing urethral resistance during micturition that does not induce any negative cardiovascular effects in patients with benign prostatic hyperplasia. PMID- 11260351 TI - Renal pelvic carcinoma of horseshoe kidney caused systemic metastasis by implantation in prostate. AB - A case is reported of renal pelvic carcinoma of the horseshoe kidney in a 69-year old man, which showed an interesting metastatic pattern by implantation in the prostate. A few months after transurethral resection of the prostate for benign prostate hyperplasia and extracorporal shock wave lithotripsy for renal stones, the patient complained of severe back pain due to multiple metastatic bone tumors. Autopsy revealed transitional cell carcinoma in the pelvis as well as in the prostate with remarkable vessel invasion. The clinical course and autopsy findings suggested that the systemic expansion of cancer cells from the renal pelvis was caused not only by direct metastasis but also by implantation in the prostate. PMID- 11260352 TI - An unusual complication of suprapubic catheterization with Cystofix: catheter knotting within the bladder. AB - Cystofix is commonly used to drain urine temporarily from the bladder. Common complications are hematuria, bladder wall edema and bladder spasm due to irritation by the catheter itself. A case is described where a Cystofix catheter became curled and knotted inside the bladder, probably due to deep introduction or increased detrussor contraction occurring with the irritation of the bladder wall. PMID- 11260353 TI - Primary signet-ring cell carcinoma of the urinary bladder inducing renal failure. AB - A case of primary signet-ring cell carcinoma of the urinary bladder that was found to have induced renal failure is the second such case reported in the world. Primary signet-ring cell carcinoma of the urinary bladder is a rare histologic variant of adenocarcinoma. The patient died of distant metastasis 8 months after undergoing total cystectomy. The neoplasm had a high stage at diagnosis, so the prognosis was very poor. To improve the prognosis, earlier diagnosis and establishing a regimen of chemotherapy is necessary. PMID- 11260354 TI - Candidal infections as a cause of recurrent uretero-ileal anastomotic dehiscence. AB - Fungal infections are common in immunocompromised patients. The presentation is often subtle and therefore treatment is delayed. Uretero-ileal anastomotic dehiscence due to candidal infection has never been reported before. This case represents an uncommon but potentially life-threatening complication in reconstructive surgery; that is, anastomotic dehiscence due to a unique etiology. PMID- 11260355 TI - Appendiceal carcinoma invading the urinary bladder. AB - A case is reported of a 78-year-old woman with appendiceal carcinoma invading the bladder causing irritative symptoms. Although several imaging studies suggested that the secondary bladder tumor was of cecal or appendiceal origin, such as abscess or mucocele, histologic findings on transurethral and transvaginal biopsy were inconclusive. However, following laparotomy, pathologic examination of the frozen sections revealed a mucinous cystadenocarcinoma originating in the appendix and a right hemicolectomy and en bloc partial cystectomy were performed. One year after the operation, the patient was well with no evidence of recurrent cancer. PMID- 11260356 TI - Osteogenic sarcoma of the prostate. AB - A 76-year-old man was treated with bilateral orchiectomy, estramustine phosphate and pelvic irradiation for prostate cancer. Osteogenic sarcoma of the prostate developed 18 months after the treatment. Postmortem examination revealed that the tumor was 8 cm in diameter and had infiltrated into the bladder and rectal walls and had resulted in peritoneal dissemination. There was no distant metastasis. Macroscopically, the tumor was ashen, firm and relatively homogenous and diffusely spread. Histologically, it was composed of spindle and pleomorphic cells, which were making osteoid with calcification. There was no ordinary tubular formation as shown in adenocarcinoma of the prostate. No positive immunostaining for prostate-specific antigen, epithelial membrane antigen and cytokeratin (AE-1, AE-3) were confirmed. Positive immunostaining for nonepithelial marker vimentin was confirmed. The ultimate diagnosis was osteogenic sarcoma of the prostate. PMID- 11260357 TI - Endoscopic management of a traumatic disruption of the bulbous urethra using a thin trocar puncture. AB - A case is reported in which complete disruption of the bulbous urethra resulted in a straddled-type injury, which was managed by endoscopic realignment with a thin trocar needle. The endoscopic urethroplasty consisted of: (i) direct observation of the proximal end of the obliterated urethra by an antegrade flexible cystoscope; (ii) adjustment of the direction of trocar penetration under fluoroscopy and direct vision; (iii) confirmation of the exact trocar position and penetration by antegrade flexible cystoscope; and (iv) placement of the guidewire following the trocar penetration as guidance for urethrotomy. A Foley catheter was left in place for 6 weeks. To date, no further endoscopic revision has been required. Although long-term follow up and more experience are required, this technique is reported because it appears to be safe, reproducible, simple and minimally invasive. PMID- 11260358 TI - The autonomic and sensory innervation of the smooth muscle of the prostate gland: a review of pharmacological and histological studies. AB - 1. We review literature demonstrating (a) the presence and (b) the actions of substances that mediate or modify neuroeffector transmission to the smooth muscle of the prostrate stroma of a number of species including man. 2. In all species studied prostatic stroma, but not secretory acini, receives rich noradrenergic innervation. Stimulation of these nerves causes contractions of prostate smooth muscle that are inhibited by guanethidine and by alpha1-adrenoceptor antagonists that probably act at the alpha1L-adrenoceptor. Such actions underlie the clinical use of alpha1-adrenoceptor antagonists in benign prostatic hyperplasia (BPH). 3. Acetylcholinesterase-positive nerves innervate prostatic stroma as well as epithelium. Atropine reduces nerve-mediated contractions of stromal muscle in the rat, guinea-pig and rabbit. M1, M2 and M3 muscarinic receptors have been implicated in eliciting or facilitating contraction in the prostate from guinea pig, dog and rat, respectively. 4. Adenine nucleotides and nucleosides, nitric oxide (NO), opioids, neuropeptide Y (NPY) and vasoactive intestinal peptide (VIP) may act as co-transmitters or modulators in autonomic effector nerves supplying prostate stroma. Adenosine inhibits neurotransmission to the rat prostate, and NO is inhibitory in prostate from human, rat, rabbit, pig and dog. The activity of peptides present in the relatively sparse sensory innervation of the prostate exhibits species variation, but, when effective, calcitonin gene-related peptide is inhibitory while tachykinins are stimulant. The roles of NPY and VIP in modulating stromal contractility remain unclear. 5. Taken together the current literature indicates that, in addition to noradrenaline, other neurotransmitters and neuromodulators may regulate the tone of prostatic smooth muscle. Whether drugs that mimic or modify their actions might be useful in providing symptomatic relief of the urinary symptoms associated with BPH remains to be established. PMID- 11260359 TI - Potassium channels in gastrointestinal smooth muscle. AB - 1. Electromechanical coupling in smooth muscle serves to coordinate the contractile activity of the syncytium. Electrical activity of smooth muscle of the gut is generated by ionic conductances that regulate and in turn are regulated by the membrane potential of smooth muscle cells. This activity determines the extent of Ca2+ entry into smooth muscle cells, and thus, the timing and intensity of contractions. 2. Potassium channels play an important role in regulating the excitability of the syncytium. The different types of K+ channel are characterized by different sensitivities to membrane potential, to intracellular Ca2+ levels and to modulation by agonists. 3. This review highlights the different types of K+ channels found in gut smooth muscle and describes their possible roles in regulating the electrical activity of the muscle. PMID- 11260360 TI - The relative importance of the time-course of receptor occupancy and response decay on apparent antagonist potency in dynamic assays. AB - 1. The potency of the beta1-adrenoceptor antagonist atenolol was measured as an inhibitor of responses to isoprenaline in guinea-pig left atria. Measurements were made in two ways, firstly, by pre-incubating the atria with a given concentration of atenolol followed by an isoprenaline dose-response curve and, secondly, by measuring the response to isoprenaline followed by addition of atenolol. 2. It was found that the estimation of atenolol potency as an antagonist of beta1-adrenoceptors by these two methods gave divergent results. Specifically, it was found that the isoprenaline-induced increased rate of myocardial relaxation was resistant to receptor blockade. Thus, the rate-limiting step in the relaxation response was dissociated from receptor activation and therefore, could not be used for the measurement of receptor occupancy. 3. In contrast, the positive inotropic response was very responsive to receptor occupancy. However, when atenolol was used to block a steady-state isoprenaline response, there was a complicating depression of basal inotropy after receptor blockade that obfuscated measurement of receptor blockade. 4. In general, these data indicated that the blockade of a steady-state agonist response to measure the potency of an antagonist might in some cases yield erroneous results. These studies indicate some caution in the interpretation of blockade responses in pre contracted or pre-stimulated pharmacological preparations. PMID- 11260361 TI - Inhibition of isolated rat and human detrusor muscle contraction by disopyramide. AB - 1. The aim of this study was to investigate the effect of in vitro and in vivo disopyramide treatment on human and rat isolated detrusor muscle contractile response. Muscle strips were suspended in an organ bath chamber containing Kreb's solution at 37 degrees C aerated with 95% oxygen and 5% carbon dioxide. 2. Disopyramide antagonized significantly the contractile response of isolated human detrusor muscle to carbachol stimulation, shifting the concentration-response curve to the right in a parallel manner. The pA2 for competitive inhibition was estimated to be 6.4 with a slope of 0.64 for disopyramide. 3. Rat detrusor muscle contractile response to electrical field stimulation (EFS) was inhibited by 28% (P < 0.01) after in vitro administration of disopyramide (7.5 x 10(-6) M). Disopyramide had no effect on the atropine-resistant component of the response to EFS or on spontaneous contractions of isolated rat detrusor muscle strips. The concentration-response curve to carbachol was competitively antagonized by disopyramide with a pA2 of 6.3 and a Schild curve a slope of 0.85. 4. Disopyramide (7.5 x 10(-6) M) had no effect on rat detrusor contractile response to low concentrations of KCl but at high concentrations contraction was reduced significantly by 16% (P < 0.01). 5. In vivo treatment of rats with disopyramide for 8 days or with a single dose had a significant inhibitory effect on the contractile response of detrusor muscle strips. 6. In conclusion, disopyramide had a significant anticholinergic effect on isolated human and rat detrusor muscle contraction. PMID- 11260362 TI - Influence of the epithelium on acetylcholine release from parasympathetic nerves of the rat trachea. AB - 1. The present study was undertaken to investigate the influence of the airway epithelium on the release of acetylcholine (ACh) from parasympathetic nerves of the rat trachea. Epithelium-intact and epithelium-denuded preparations of rat trachea were incubated with [3H]-choline to incorporate [3H]-ACh into the cholinergic transmitter stores. Release of radiolabelled transmitter ACh was evoked by electrical field stimulation (60 s trains of 1 ms pulses, 5 Hz, 15 V). 2. Field stimulation both of epithelium-intact and epithelium-denuded radiolabelled tracheal preparations evoked an increase in the efflux of radioactivity; however, the mean stimulation-induced (S-I) efflux from epithelium denuded preparations (2932 +/- 190 d.p.m., n = 9) was approximately 60% of that from epithelium-intact preparations (4802 +/- 820 d.p.m., n = 11). We have shown previously that, in epithelium-intact (but not epithelium-denuded) tracheal preparations, a substantial proportion of the S-I efflux is resistant to tetrodotoxin (1 microM) and to the removal of extracellular Ca2+, indicating that much of the S-I efflux is not caused by exocytotic release of neuronal [3H]-ACh. In epithelium-denuded tracheal preparations, superfused individually, phosphorylcholine (1 and 100 microM) did not alter S-I efflux. In epithelium intact tracheal preparations, both in the absence and in the presence of atropine (1 microM), neither N(G)-nitro-L-arginine (100 microM), superoxide dismutase (100 units ml(-1)), indomethacin (10 microM), capsaicin (30 microM) nor alpha chymotrypsin (1 unit ml(-1)) altered S-I efflux. 3. Experiments were also performed using two tracheal preparations superfused in series. When unlabelled epithelium-intact preparations were present in the upper chamber (superfused first), the S-I efflux from radiolabelled epithelium-denuded preparations in the lower chamber (superfused second) did not differ significantly from radiolabelled epithelium-denuded preparations superfused individually. Moreover, there was no significant difference in the S-I efflux from radiolabelled epithelium-denuded preparations in the lower chamber between experiments in which the upper chamber contained epithelium-intact or epithelium-denuded preparations. 4. Field stimulation of epithelium-intact tracheal preparations in the upper chamber with 90, 120 and 300-s periods (trains of 1 ms pulses, 5 Hz, 15 V) did not significantly alter the S-I efflux from radiolabelled epithelium-denuded tracheal preparations in the lower chamber. 5. When introduced into the upper (unlabelled epithelium-intact) and subsequently allowed to superfuse the lower (radiolabelled epithelium-denuded) tracheal preparations, the stable cholinomimetic carbachol (3 microM) markedly reduced the S-I efflux whereas ACh (0.1 and 1 microM) had no significant effect. However, in the presence of the anti-cholinesterase neostigmine (1 microM), ACh (1 microM) significantly reduced S-I efflux, indicating that ACh is subject to rapid hydrolysis by cholinesterase enzymes. When atropine (10 microM) was only exposed to radiolabelled epithelium-denuded preparations in the lower chamber, the inhibitory effects of ACh (1 microM) and carbachol (3 microM) on S-I efflux were prevented. 6. In conclusion, the findings of the present study do not support the notion that the airway epithelium exerts an inhibitory influence on ACh release from parasympathetic nerves of the rat trachea. Alternatively, if epithelium-dependent modulation of cholinergic transmission does occur in the rat trachea, then the mechanism does not appear to involve phosphorylcholine, nitric oxide, superoxide radicals, cyclo-oxygenase products of arachadonic acid, capsaicin-sensitive neuropeptides or vasoactive intestinal peptide. Moreover, the inhibitory effect of carbachol and ACh on transmitter ACh release in the rat trachea appears to be due solely to activation of prejunctional inhibitory muscarinic cholinoceptors on parasympathetic nerves and does not involve the liberation of a putative epithelium-derived inhibitory factor. PMID- 11260363 TI - Functional identification of beta3-adrenoceptors in the guinea-pig ileum using the non-selective beta-adrenoceptor antagonist (+/-)-bupranolol. AB - 1. To clarify whether there is a species difference or a tissue difference in beta3-adrenoceptors, the beta3-adrenoceptors mediating relaxations to catecholamines ((-)-isoprenaline, (-)-noradrenaline and (-)-adrenaline), a selective beta3-adrenoceptor agonist BRL37344 and a non-conventional partial beta3-adrenoceptor agonist (+/-)-CGP12177A (a potent beta1- and beta2 adrenoceptor antagonist with a partial beta3-adrenoceptor agonist property) were investigated in the guinea-pig ileum. 2. Catecholamines and beta3-adrenoceptor agonists induced concentration-dependent relaxations of pre-contracted strips of the guinea-pig ileum. The rank order for their relaxing potency was (-) isoprenaline (pD2: 7.60) > BRL37344 (7.05) > (-)-noradrenaline (6.38) > (+/-) CGP12177A (6.25) > (-)-adrenaline (6.07). 3. In the presence of the non-selective beta1- and beta2-adrenoceptor antagonist (+/-)-propranolol (1 microM), only small rightward shifts of the concentration-response curves (CRCs) to these agonists were observed and the rank order of potency of agonists was BRL37344 (pD2: 7.00) > (+/-)-CGP12177A (6.17) > (-)-isoprenaline (6.01) > (-)-noradrenaline (5.69) > ( )-adrenaline (5.41). 4. In the presence of (+/-)-propranolol (1 microM), the additional presence of (+/-)-bupranolol (3-30 microM), a non-selective beta1-, beta2- and beta3-adrenoceptor antagonist, caused a concentration-dependent rightward shift of the CRCs to catecholamines and beta3-adrenoceptor agonists. Schild plot analyses of (+/-)-bupranolol against these agonists gave pA2 values of 6.02 ((-)-isoprenaline), 6.03 ((-)-noradrenaline), 6.01 ((-)-adrenaline), 6.56 (BRL37344) and 5.74 ((+/-)-CGP12177A), respectively. All Schild plot slopes were not significantly different from unity. The pA2 values of (+/-)-bupranolol obtained for the guinea-pig beta3-adrenoceptors were about one log unit less than the values obtained for the rat beta3-adrenoceptors and about two log units less than the values obtained for dog beta3-adrenoceptors. 5. These results confirm that functional beta3-adrenoceptors are present in the guinea-pig ileum and that the relaxations of these agonists are mainly mediated via beta3-adrenoceptors in this tissue. The differential antagonistic potency of (+/-)-bupranolol may suggest that there is a species difference between the three species (guinea-pig, dog and rat) in their beta3-adrenoceptors. PMID- 11260365 TI - Wall tension and contraction of the aorta in 6-month-old spontaneously hypertensive rats. AB - 1. The present study aimed at comparing the influence of passive tension on the effect exerted by noradrenaline on the thoracic aorta of 6-month-old Wistar and spontaneously hypertensive rats (SHR). 2. Concentration-response curves to noradrenaline were obtained in aorta rings, at two levels of passive tension: 3 and 0.5 g. 3. The maximal responses (in percentage of the maximal response to noradrenaline obtained in the beginning of the experiment at a tension of 2 g) were significantly larger (P < 0.05) at 3 g than at 0.5 g in both kinds of rats: 171 versus 69%, respectively, for SHR; 139 versus 76%, respectively, for Wistar rats. 4. When expressed as mg of active tension per mg of tissue, the maximal contraction at both 3 and 0.5 g was smaller in SHR than in Wistar rats (at 3 g: 64.6 +/- 6.7, n = 6 versus 122.3 +/- 19.1, n = 8, respectively, P < 0.05; at 0.5 g: 24.0 +/- 1.0, n = 6 versus 49.0 +/- 5.9, n = 8, respectively, P < 0.05). 5. Maximal responses to noradrenaline were markedly decreased by cytochalasin B (50 microM) (to 15.2 +/- 6.0%, n = 6, at 3 g and 2.8 +/- 1.9%, n = 6, at 0.5 g in SHR; to 11.8 +/- 2.3%, n = 4, at 3 g and 4.3 +/- 1.3%, n = 4, at 0.5 g in Wistar rats). Cytochalasin B at a lower concentration (4 microM) produced a less marked decrease in responses to noradrenaline in both strains of rats. The presence of cardiovascular structural changes in SHR was confirmed by the fact that the heart weight (mg):body weight (g) was higher in SHR (3.37 +/- 0.06, n = 10) than in Wistar rats (2.40 +/- 0.12, n = 11) (P < 0.05). 6. It is concluded that in 6 month-old SHR the contractile capacity of the aortic tissue is reduced. However, the differential sensitivity of aortic smooth muscle at the two different levels of tension remains present. This difference may depend on filament interaction. Contractions to noradrenaline in the rat aorta are highly dependent on actin polymerization. PMID- 11260364 TI - Effects of extremely low frequency magnetic fields on pain thresholds in mice: roles of melatonin and opioids. AB - 1. We studied the effects of extremely low frequency (ELF, 60 Hz) magnetic fields (MFs) on pain thresholds using the hot plate test. The implication of opioid and benzodiazepine system in the MFs-induced alteration of pain thresholds was also studied. 2. There was an increase at night time and a decrease at daytime of pain thresholds in normal mice. Exposure of MFs (24 h, 20 gauss (G)) inhibited the increase of pain thresholds at night time and even produced hyperalgesia at daytime. 3. The increase of pain thresholds induced by melatonin at daytime was inhibited by exposure to MFs (24 h, 20 G) or opioid antagonist naloxone. The MFs and naloxone synergically inhibited hypoalgesia produced by melatonin. The hyperalgesia at daytime after MFs exposure was potentiated by the benzodiazepine agonist, diazepam, and inhibited by the benzodiazepine antagonist, flumazenil. There was no significant difference in all rotarod performance we tested. 4. From these results, it is suggested that exposure to MFs inhibits the increase of pain thresholds at night time and produces hyperalgesia at daytime with the involvement of opioid and benzodiazepine systems. PMID- 11260381 TI - Renoprotection: one or many therapies? AB - BACKGROUND: Renal disease that progresses to end-stage renal disease (ESRD) imposes a great burden on the affected individual and on society, which mainly bears the cost of ESRD (currently more than $10 billion to treat about 333,000 patients annually in the U.S.). Thus, there is a great need to identify therapies that arrest the progression mechanisms common to all forms of renal disease. Progress is being made. Perhaps the most visible advance is the randomized controlled trials (RCT) demonstrating the renoprotective effects of angiotensin converting enzyme (ACE) inhibitors. There are also numerous other promising renoprotective therapies. Unfortunately, testing each therapy in RCT is not feasible. Thus the nephrologist has two choices: restrict renoprotective therapy to those shown to be effective in RCT, or expand the use of renoprotective therapies to include those that, although unproven, are plausibly effective and prudent to use. The goal of this work is to provide the documentation needed for the nephrologist to choose between these strategies. METHODS: This work first describes the mechanisms believed to be involved in the progression of renal disease. Based largely on this information, 18 separate interventions that slow the progression are described. Each intervention is assigned a level of recommendation (Level 1 is the highest and Level 3 the lowest) according to the strength of evidence supporting its renoprotective efficacy. RESULTS: The number of interventions at each level of recommendation are: Level 1, N = 4; Level 2, N = 4; Level 3, N = 10. Our own experience with the multiple-risk-factor intervention is that most patients can achieve the majority of the Level 1 and 2 interventions, and many of the Level 3 interventions. We recommend the expanded renoprotection strategy. CONCLUSION: This work advances the hypothesis that, until better information becomes available, a broad-based, multiple-risk-factor intervention intended to slow the progression of renal disease can be justified in those with progressive nephropathies. This work is intended primarily for clinical nephrologists and thus each recommended intervention is described in substantial practical detail. PMID- 11260382 TI - Intrarenal synthesis of complement. AB - During the past decade, research has shown that the kidney has the capacity to synthesize most of the activation pathway components of the complement cascade. As well as implying physiological roles in local clearance of immune complexes and defense against invasive organisms, an increasing amount of evidence indicates that the intrarenal synthesis of complement makes an important contribution in the pathogenesis of renal injury. Here we review this evidence and present a case for more definitive investigation of these functions. PMID- 11260383 TI - Clinical and pathologic features of focal segmental glomerulosclerosis with mitochondrial tRNALeu(UUR) gene mutation. AB - BACKGROUND: Several families have been described in which an A to G transition mutation at position 3243 (A3243G) of the mitochondrial DNA (mtDNA) is associated with focal and segmental glomerulosclerosis (FSGS). However, the prevalence, clinical features, and pathophysiology of FSGS carrying mtDNA mutations are largely undefined. METHODS: Among 11 biopsy-proven primary FSGS patients of unknown etiology, we examined seven FSGS patients to determine whether any of the clinical and pathological features of FSGS were associated with an A3243G mtDNA mutation. In four subjects in whom the A3243G mtDNA mutation was discovered in blood leukocytes, as well as in urine sediments, we retrospectively reviewed the medical records and re-evaluated the renal biopsy specimen using light and electron microscopy. We further screened the patient's family members for the presence and degree of heteroplasmy for this mtDNA mutation and obtained medical histories that were consistent with mitochondrial cytopathy. RESULTS: The four individuals identified with the A3243G mtDNA mutation were female. Proteinuria was diagnosed in these individuals during a routine annual health checkup in their teenage years. None of the patients showed any symptoms related to mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episode, whereas diabetes mellitus in two of the patients and a hearing disturbance in one patient became manifest within a 3- to 13-year follow-up period. Strict maternal transmitted inheritance was confirmed by pedigree studies in all of these patients. Steroid therapy was ineffective in all four patients. In two of these patients, renal function declined slowly to end-stage renal failure. Histologic examination of biopsy specimens revealed that glomeruli were not hypertrophied, while electron microscopic examination identified severely damaged, multinucleated podocytes containing extremely dysmorphic abnormal mitochondria in all patients. CONCLUSIONS: FSGS may belong to the spectrum of renal involvement in A3243G mtDNA mutation in humans. Severely injured podocytic changes containing abnormal mitochondria may explain the pathogenesis of FSGS in association with the A3243G mtDNA mutation. PMID- 11260384 TI - No evidence for AT2R gene derangement in human urinary tract anomalies. AB - BACKGROUND: It has been recently found that mice, especially males, with a disrupted angiotensin type 2 receptor (AT2R) gene, which is located on the X chromosome, often have a range of congenital anomalies of the kidney and urinary tract (CAKUT), including renal hypoplasia, and that Caucasian male patients with ureteropelvic junction stenosis (UPJ) and multicystic dysplastic kidneys frequently have A-G transition in intron 1 of the AT2R gene. We have previously found that renal hypoplasia is remarkably predominant in Japanese boys. METHODS: We investigated sex ratios for the frequency of each CAKUT. The frequency of the A-G transition between the controls and 66 Japanese boys with CAKUT were compared. There was renal hypoplasia in 16, UPJ in 17, vesicoureteral in 20, and other anomalies in 13. We also investigated whether any mutations in AT2R genes were detectable in patients with renal hypoplasia. RESULTS: In contrast to mice with a disruption of the AT2R gene, the male-to-female ratios in human patients proved to be considerably variable: 16 for renal hypoplasia, 2.1 for UPJ, 0.8 for vesicoureteral, and 1.2 for others. The frequency of the A-G transition was not different between the control population and the patients with CAKUT [31 of 102 (30%) vs. 23 of 66 (35%), respectively]. A sequencing study disclosed no mutations in nine boys with renal hypoplasia. CONCLUSIONS: These findings indicate that the AT2R gene may not play a major role in the development of renal hypoplasia and other CAKUT in humans, at least in the Japanese population. PMID- 11260385 TI - Cystinuria type I: identification of eight new mutations in SLC3A1. AB - BACKGROUND: Cystinuria is a heritable disorder of amino acid transport characterized by the defective transport of cystine and the dibasic amino acids through the brush border epithelial cells of the renal tubule and intestine tract. Three types of cystinuria (I, II, and III) have been described based on the urinary excretion of cystine and dibasic amino acids in obligate heterozygotes. The SLC3A1 gene coding for an amino acid transporter named rBAT is responsible for type I cystinuria, whereas the SLC7A9 gene coding for a subunit (b0,+AT) of rBAT is involved in determining non-type I (types II and III) cystinuria. METHODS: The SLC3A1 gene sequence was investigated in a sample of seven type I/type I, three type I/non-type I, six type I/untyped, and four untyped unrelated cystinuric patients by RNA single-strand conformation polymorphism (RNA-SSCP). RESULTS: Eight new point mutations (S168X, 765+1G>T, 766 2A>G, R452Q, Y461X, S547W, L564F, and C673W) and seven previously reported mutations were detected. These new mutations increase the number of mutated alleles so far characterized in SLC3A1 to 62. CONCLUSIONS: We have found SLC3A1 mutations in 0.739 of the type I chromosomes studied. The relatively high proportion of uncharacterized type I chromosomes suggests either that there may be mutations not yet found in SLC3A1 or that many of the assigned type I chromosomes in mixed type I/non-type I patients may have mutations in SLC7A9. If the hypothesis is excluded in the future, we believe that a third gene may be involved in cystinuria. PMID- 11260387 TI - Increased cGMP phosphodiesterase activity mediates renal resistance to ANP in rats with bile duct ligation. AB - BACKGROUND: Liver disease resulting from common bile duct ligation (CBDL) causes abnormal sodium metabolism that is manifested by resistance to the natriuretic action of atrial natriuretic peptide (ANP). This resistance is corrected both in vitro and in vivo by zaprinast, a selective inhibitor of a guanosine cyclic-3'-5' monophosphate (cGMP)-specific phosphodiesterase (PDE5). Several other PDEs with affinity for cGMP are expressed in kidney and could also be involved in this response. METHODS: We measured cGMP hydrolysis in inner medullary collecting duct (IMCD) cell homogenates from kidneys of sham-operated and CBDL rats and quantitated the amount of PDE5 protein by Western blotting and immunoprecipitation studies. We also characterized ANP responsiveness in vivo of kidneys of anesthetized sham and CBDL rats by measuring sodium excretion before and after volume expansion (VE). RESULTS: Kinetic analysis of PDE5 activity in homogenates of IMCD cells isolated from kidneys of sham-operated rats indicated a Vmax of 85.3 +/- 1.7 versus 157 +/- 2.9 pmol/mg/min from CBDL rats (P < 0.01), without a difference in Km. Enzyme activity was inhibited competitively by 1,3 dimethyl-6-(2-propoxy-5-methanesulfonylamidophenyl)pyrazol[3,4d]-pyrimidin-4-(5H) one (DMPPO), a potent and specific inhibitor of PDE5, with an apparent Ki of 4.5 +/- 0.7 and 4.9 +/- 0.7 nmol/L and an IC50 of 6.1 +/- 0.8 and 8.7 +/- 0.7 nmol/L in sham and CBDL rats, respectively (P = NS). DMPPO exhibited very poor inhibitory activity against the calcium-calmodulin-dependent PDE1 in IMCD homogenates from sham rats (Ki 1.3 +/- 0.1 micromol/L and IC50 1.9 +/- 0.2 micromol/L). Western analysis using an antiserum made against bovine lung PDE5 revealed a twofold increase in PDE5 protein in cytosolic extracts from IMCD of CBDL rat kidneys compared with sham-operated controls, and immunoprecipitation studies indicated that the increase in PDE5 protein accounted for the observed increase in cGMP hydrolysis. DMPPO (10 nmol/L) normalized the blunted ANP dependent cGMP accumulation by IMCD cells from CBDL rats in vitro. Intrarenal infusion of DMPPO (0.5 nmol/min) in CBDL rats corrected both the impaired natriuretic response to VE and the blunted VE-related increase in urinary cGMP excretion from the infused, but not the contralateral kidney. CONCLUSION: These results demonstrate that renal resistance to ANP in CBDL rats is accompanied by heightened activity of PDE5, which is due largely to an increase in PDE5 protein. Other PDEs could contribute only a minor part to the enhanced cGMP hydrolysis observed in kidneys of CBDL rats. This PDE5-dependent ANP resistance may represent an important contributor to the sodium retention of liver disease. PMID- 11260386 TI - AT1 receptor antagonist combats oxidative stress and restores nitric oxide signaling in the SHR. AB - The tubuloglomerular feedback (TGF) responses of the spontaneously hypertensive rat (SHR) are under exaggerated regulation by angiotensin II (Ang II) type 1 receptors (AT1-R). Since AT1-Rs enhance oxygen radical (O2-) generation, we tested the hypothesis that the exaggerated TGF was due to a diminished blunting by macula densa (MD)-derived nitric oxide (NO) because of excessive AT1-R dependent generation of O2-. Groups of SHR and control Wistar-Kyoto (WKY) rats received vehicle (Veh), the AT1-R antagonist candesartan (Cand; 3 mg. kg-1. day 1), or nonspecific therapy with hydralazine + hydrochlorothiazide + reserpine (HHR) for two weeks. Compared with WKY rats, the elevated mean arterial pressure of SHR (WKY 125 +/- 2 vs. SHR 163 to 779 mm Hg, P < 0.001) was reduced (P < 0.001) similarly in SHR by Cand and HHR (121 +/- 5 and 116 +/- 5 mm Hg, P = NS). The SHR had an increased maximal TGF response (change in stop flow pressure during luminal perfusion of fluid: SHR 11.2 +/- 0.5 vs. WKY 8.3 +/- 0.4 mm Hg, P < 0.01) and a reduced TGF response to blockade of neuroneal NO synthase (nNOS) in the MD with luminal 7-nitroindazole (7-NI: DeltaTGF in WKY 2.8 +/- 0.4 vs. SHR 1.1 +/- 0.6 mm Hg, P < 0.05). Although the elevated TGF responses of SHR were normalized by both HHR and Cand, only Cand restored a normal TGF response to luminal perfusion of the MD with 7-NI (DeltaTGF with 7-NI in SHR: Veh + 1.8 +/- 0.4 vs. Cand + 3.4 +/- 0.5 mm Hg, P < 0.05). To abrogate the local effects of O2 , tempol (a membrane-permeable superoxide dismutase mimetic) was perfused into the efferent arteriole. During tempol, SHR given vehicle or HHR had a much increased response to blockade of nNOS with 7-NI (DeltaTGF in SHR with 7-NI during tempol: Veh 6.3 +/- 1.0 and HHR 4.5 +/- 0.8 mm Hg, P < 0.01 vs. no tempol for both), implying that the effects of NO had been prevented because of excessive O2-. In contrast, the TGF response to 7-NI in SHR given Cand was unaffected by tempol (DeltaTGF with 7-NI during tempol: 2.9 +/- 0.9, P = NS, compared with no tempol). In conclusion, TGF responses of SHR are exaggerated because of the effects of hypertension and AT1-R. AT1-R blockade specifically diminishes oxidative stress and restores NO signaling in the juxtaglomerular apparatus of the SHR. PMID- 11260388 TI - Increased expression of atrial natriuretic peptide in the kidney of rats with bilateral ureteral obstruction. AB - BACKGROUND: Whether the postobstructive diuresis can be related to an altered regulation of local atrial natriuretic peptide (ANP) in the kidney was investigated. METHODS: Three groups of rats had both of their ureters obstructed for 48 hours. The kidneys were taken without releasing the obstruction in one group [bilateral ureteral obstruction (BUO)]. The obstruction was released in the other two groups and the animals were kept for 4 and 24 hours thereafter to collect urinary data (BUR-4 and BUR-24, respectively). Plasma and urine ANP levels were measured by radioimmunoassay. The mRNA expression of ANP, natriuretic peptide receptor-A (NPR-A), and NPR-C was determined by reverse transcription polymerase chain reaction. ANP receptors were also quantitated by in vitro autoradiography. The activity of guanylyl cyclase was determined by the amount of cGMP generated in response to ANP. RESULTS: Urinary volume and sodium excretion increased in BUR-4, along with the ANP mRNA expression in the kidney and the urinary ANP excretion. The ANP excretion positively correlated with the urinary volume and sodium excretion. The mRNA expression of both NPR-A and NPR-C was decreased by BUO, the latter being far more prominently affected. The maximal binding capacity of radiolabeled ANP was decreased in the glomerulus and papilla in BUO. Not only the urinary parameters but also the mRNA expression of ANP, NPR A, and NPR-C were comparable between BUR-24 and control rats. ANP-stimulated cGMP generation was reduced in the glomerulus and papilla in BUO animals, which was rapidly resumed following the release of the obstruction. CONCLUSIONS: Postobstructive diuresis may be due partially to an increased ANP activity in the kidney. PMID- 11260389 TI - Mesangial cells release untransformed prostaglandin H2 as a major prostanoid. AB - BACKGROUND: Prostaglandin H2 (PGH2) is the precursor of the other prostanoids and exhibits a vasoconstricting activity. Glomerular mesangial cells are an important source of vasoactive prostanoids in kidney. Hence, the present investigation focused on the release of untransformed PGH2 by rat glomerular mesangial cells (RGMCs). METHODS: Synthesis of prostanoid by resting and interleukin-1beta (IL 1beta)-treated (overnight) RGMCs from exogenous or endogenous arachidonic acid (AA) was assessed by high-performance liquid chromtography or enzyme immunoassay, respectively. Cyclo-oxygenase isoforms were determined by Western blotting. Release of untransformed PGH2 from exogenous AA was evaluated in RGMCs and intact glomeruli as the difference of PGF2alpha formed in the incubations performed in the presence and in the absence of SnCl2 or measuring the ability of aspirin treated platelets to form thromboxane B2 (TXB2) in mixed incubations of platelets and RGMCs or glomeruli. RESULTS: The prostanoids formed by RGMCs were PGE2, PGF2alpha, PGI2 and PGD2. SnCl2 totally deviated formation of PGE2 and PGD2 toward PGF2alpha in resting RGMCs, whereas PGE2 was only partially deviated toward PGF2alpha in IL-1beta-treated RGMCs. The PGE2/PGD2 ratio in resting RGMCs was similar to that expected for nonenzymatic isomerization of PGH2, whereas this ratio was higher in IL-1beta-treated RGMCs, suggesting the induction of PGE synthase by IL-1beta. Aspirin-treated platelets formed TXB2 when either RGMCs or intact glomeruli were present in the incubation and formation of TXB2 was approximately fourfold higher with IL-1beta-treated RGMCs or glomeruli. CONCLUSIONS: RGMCs and intact glomeruli released substantial amounts of untransformed PGH2, which was enhanced following exposure to IL-1beta. PMID- 11260390 TI - Role of nitric oxide in renal tubular apoptosis of unilateral ureteral obstruction. AB - BACKGROUND: The obstructed kidney in unilateral ureteral obstruction (UUO) is characterized by renal atrophy and tissue loss, which is mediated by renal tubular apoptosis. We sought to determine whether NO is involved in renal tubular apoptosis in vitro and in vivo. METHODS: Rat renal tubular epithelial cells (NRK 52E) were subjected to mechanical stretch, and apoptosis and cell size were analyzed by flow cytometry. Furthermore, we studied UUO in mice lacking the gene for inducible nitric oxide synthase (iNOS-/-) and their wild-type littermates. Tubular apoptosis and proliferation were detected by immunostaining. NOS activity and NOS expression were assessed by a citrulline assay and Western blot, respectively. RESULTS: Stretching-induced apoptosis in NRK-52E, which was reduced when NO was increased; conversely, stretch-induced apoptosis was increased when a NOS inhibitor was added to the cells. Stretched cells are larger and more apoptotic than unstretched cells. In UUO, the obstructed kidney of iNOS-/- mice exhibited more apoptotic renal tubules than the wild-type mice through 14 days of UUO. The obstructed kidney of iNOS-/- mice at day 3 showed more proliferative tubules compared with wild type. The obstructed kidney of wild-type mice exhibited higher total NOS activity until day 7 after UUO compared with iNOS-/- mice. However, the obstructed kidney of day 14 wild-type mice exhibited significantly lower iNOS activity and protein compared with the day 0 kidney. CONCLUSION: These results suggest that mechanical stretch is related to renal tubular apoptosis and that NO plays a protective role in this system in UUO. PMID- 11260391 TI - Hepatocyte growth factor suppresses interstitial fibrosis in a mouse model of obstructive nephropathy. AB - BACKGROUND: As tubulointerstitial fibrosis (TIF) reflects the prognosis of patients with various chronic renal diseases, the pathogenesis of TIF has to be clarified. Transforming growth factor-beta (TGF-beta) is a key mediator for renal fibrosis. We reported that hepatocyte growth factor (HGF) prevents renal fibrosis in nephrotic mice. However, the function of HGF in chronic renal failure, except for nephrotic syndrome, remains to be determined. METHODS: Using mice subjected to unilateral ureter-ligated obstruction (UUO), we investigated the roles of HGF in TIF, as induced by obstructive nephropathy. Pathophysiological changes in the kidney after UUO treatment were analyzed focusing on expressions of renal HGF and TGF-beta, TIF, tubular proliferation, and apoptosis. Neutralizing antibody against rodent HGF, or recombinant human HGF (rhHGF), was administrated to the UUO mice, and pathophysiological changes after neutralization or supplements of HGF were analyzed. RESULTS: In this UUO model, TIF with tubular apoptosis became evident, and it was accompanied by a decrease in renal HGF expression and an increase in renal TGF-beta expression. Neutralization of endogenous HGF accelerated the progression of TIF, accompanied by increases in TGF-beta expression and tubular apoptosis as well as by decreases in tubular proliferation. In contrast, rhHGF attenuated TIF progression, and there were decreases in TGF-beta expression and tubular apoptosis, and an increase in tubular proliferation. CONCLUSIONS: Endogenous as well as exogenous HGF attenuated the progression of the fibrosis caused by obstructive nephropathy in these mice. Thus, local reduction in HGF levels may account for TIF in chronic renal diseases. PMID- 11260392 TI - Leptin stimulates type I collagen production in db/db mesangial cells: glucose uptake and TGF-beta type II receptor expression. AB - BACKGROUND: Serum leptin levels correlate with fat cell mass and are elevated in patients with massive obesity and type 2 diabetes mellitus, which are strong risk factors for the development of glomerulosclerosis. We have previously shown in cultured glomerular endothelial cells that leptin stimulates cellular proliferation and expression of the prosclerotic cytokine transforming growth factor-beta1 (TGF-beta1). Although the effect of leptin on the hypothalamus to regulate energy homeostasis is well known, the effect of leptin on the kidney, and specifically on the glomerular mesangial cell, is unclear. METHODS: The obese, diabetic db/db mouse, which lacks the functional full-length Ob-Rb leptin receptor, is a suitable model to assess the effects of hyperleptinemia on peripheral tissues that express other receptor isoforms. The effects of leptin on glucose uptake, the TGF-beta system, and type I collagen production were evaluated in db/db mouse mesangial cells in culture. A phosphatidylinositol-3 kinase (PI-3K) inhibitor was used to assess the role of PI-3K in mediating the effects of leptin. RESULTS: A short form of the leptin receptor (Ob-Ra), but not Ob-Rb, was present by reverse transcription-polymerase chain reaction in the kidney and mesangial cells of both nondiabetic db/m and diabetic db/db mice. In db/db mesangial cells, leptin increased 2-deoxy-D-glucose (2DOG) uptake dose dependently and stimulated gene expression of TGF-beta type II receptor (TbetaRII) and alpha1(I) collagen, but not TGF-beta1. Protein production of type I collagen (enzyme-linked immunosorbent assay) was also increased by leptin. Both leptin-stimulated 2DOG uptake and type I collagen production were suppressed by a PI-3K inhibitor, LY294002. Mesangial cells pretreated with leptin exhibited increased responsiveness to exogenous TGF-beta1, as evidenced by a greater production of type I collagen protein in leptin-pretreated cells exposed to low dose TGF-beta1 (0.5 ng/mL). The addition of both TGF-beta1 (2 ng/mL) and leptin (100 ng/mL) increased type I collagen production more than addition of either TGF beta1 or leptin alone. CONCLUSIONS: Leptin increases glucose uptake and type I collagen in db/db mesangial cells through a PI-3K-dependent pathway. We postulate that increased leptin levels may transmit a signal through the short-form leptin receptor to up-regulate TbetaRII and activate the intraglomerular TGF-beta system, which may contribute to the glomerulosclerosis of obesity or type 2 diabetes. PMID- 11260393 TI - PDGF signal transduction inhibition ameliorates experimental mesangial proliferative glomerulonephritis. AB - BACKGROUND: Platelet-derived growth factor (PDGF) has been consistently implicated in the cell proliferation and extracellular matrix accumulation, which characterize progressive glomerular disease. In the present study, the effects of a potent and selective inhibitor of PDGF receptor tyrosine kinase, STI 571, were examined in vitro and in vivo. METHODS: Cultured mesangial cells were incubated with PDGF (50 ng/mL) and fibroblast growth factor-2 (FGF-2; 50 ng/mL) and treated with STI 571 (0.13 to 2.0 micromol/L). Experimental mesangial proliferative glomerulonephritis was induced in male Wistar rats with monoclonal OX-7, anti-rat Thy-1.1 antibody with rats randomized to receive either STI 571 (50 mg/kg intraperitoneally daily) or vehicle. Animals were examined six days later. RESULTS: In vitro, both PDGF and FGF-2 induced a threefold increase in mesangial cell 3H-thymidine incorporation. STI 571 reduced PDGF but not FGF-2-stimulated mesangial cell proliferation in a dose-dependent manner, with complete abolition at 0.4 micromol/L. In animals with Thy-1.1 glomerulonephritis, PDGF receptor tyrosine kinase blockade was associated with significant reductions in mesangial cell proliferation (P < 0.001), the number of activated (alpha-smooth muscle positive) mesangial cells, and glomerular type IV collagen deposition (P < 0.001). CONCLUSION: The amelioration of the pathological findings of experimental mesangial proliferative glomerulonephritis by blockade of PDGF receptor activity suggests the potential clinical utility of this approach as a therapeutic strategy in glomerular disease. PMID- 11260394 TI - Simultaneous blockade of endothelin A and B receptors in ischemic acute renal failure is detrimental to long-term kidney function. AB - BACKGROUND: There is growing evidence of long-term pathological consequences following renal ischemia. Endothelin (ET) receptor antagonists have proved beneficial in the treatment of ischemic acute renal failure (IARF); however, the long-term outcomes have not been assessed in this disease. METHODS: Experimental IARF was induced in uninephrectomized female Sprague-Dawley rats (N = 8) by clamping of the renal pedicle. At 24-hours postischemia, a once-only administration of drug or vehicle was given. One ischemic group received saline only (saline ischemic), and two other ischemic groups received either SB 234551 (ETA receptor antagonist, ETA group) or SB 209670 (ETA and ETB receptor antagonist, ETA/ETB group). A uninephrectomized control group was sham operated to simulate operative conditions without ischemia and was given a once-only saline infusion (sham ischemic). All groups were sacrificed at six-months postischemia. Serum creatinine was assessed daily for one week and then every four weeks. Glomerular filtration rates (GFRs), systolic blood pressure, 24-hour urine collection, and creatinine clearance were performed just prior to sacrifice. Immunohistochemistry for monocytes and macrophages (Mo and Mphi), myofibroblasts (MF, alpha-SMA), collagen IV, and collagen III was also evaluated. Cell kinetics were studied by immunostaining for proliferating cell nuclear antigen (PCNA) and by TUNEL. RESULTS: Urinalysis revealed significant increases in urinary protein and albumin in the ETA/ETB group when compared with all other groups. GFRs and creatinine clearance were also decreased significantly in the ETA/ETB group. Urine albumin, protein, GFR, and creatinine clearance in the ETA group, however, were not different from the sham ischemic and saline ischemic groups. Systolic blood pressure was increased in the saline ischemic group as compared with all other groups. Kidney weights were increased in all ischemic groups, but no differences were observed between the saline ischemic group and ETR antagonist-treated groups. Immunohistochemistry revealed relationships between Mo and Mphi, MF, and tubulointerstitial collagen III, where the saline ischemic and ETA/ETB groups were increased as compared with the sham ischemic and ETA groups. There was no change observed in tubulointerstitial collagen IV accumulation. The largest number of proliferating cells was demonstrated in the ETA/ETB group, whereas apoptotic cells were identified in small amounts in all groups, with the largest number being found in the saline ischemic group. CONCLUSIONS: Renal ischemia appears to have long-term functional and pathological consequences that can be prevented by treatment with ETA receptor antagonists. Blockade of both ETA and ETB receptors, however, appears to be detrimental to long-term kidney function. PMID- 11260395 TI - Selenium-deficient diet induces renal oxidative stress and injury via TGF-beta1 in normal and diabetic rats. AB - BACKGROUND: Oxidative stress has been implicated in the pathogenesis of diabetic nephropathy. Although glucose itself can initiate oxidative stress, deficiency of essential trace elements such as selenium (Se) may exacerbate this oxidative stress in diabetic rats. The mechanism by which Se deficiency causes oxidative stress and renal injury is not completely understood. This study tested the hypothesis that Se deficiency induces renal oxidative stress and renal injury via transforming growth factor-beta1 (TGF-beta1). METHODS: Fifty-four male Wistar rats were used. Diabetes was induced in 27 rats by streptozotocin, and the other 27 rats received buffer only. Ten weeks after induction of diabetes, both normal and diabetic rats were killed, their kidneys removed, and glomeruli were isolated. Glomeruli from normal and diabetic rats were incubated in the presence of TGF-beta1 alone or its neutralizing antibody. Antioxidant enzyme (Cu-Zn) superoxide dismutase (Cu-Zn SOD), catalase, and glutathione peroxidase (GSH-Px) activities; total glutathione; and lipid peroxidation were determined. For Se studies, 15 normal and 15 diabetic rats were divided into groups of five each and fed either a regular, Se-deficient, or Se-supplemented diet one week after induction of diabetes. Ten weeks after feeding these diets, rats were killed and glomeruli were isolated. Oxidative stress was examined by determining the mRNA expressions for antioxidant enzymes and also for TGF-beta1. Plasma glucose and albuminuria were determined. Histology of the kidney and interlobular artery was evaluated by light microscopy. RESULTS: In vitro studies showed that TGF-beta1 significantly reduced glomerular catalase and GSH-Px activities as well as total glutathione levels with an increase in lipid peroxidation in both normal and diabetic rats. Antibody to TGF-beta abrogated these changes. There was no effect of TGF-beta1 on Cu-Zn SOD. Like TGF-beta1, a Se-deficient diet caused a significant decrease in glomerular mRNA expression for Cu-Zn SOD, catalase, and GSH-Px, but a significant increase in TGF-beta1 mRNA expression. Also, a Se deficient diet caused an increase in albuminuria, glomerular sclerosis, and plasma glucose levels in both normal and diabetic rats. The deficient diet caused a decrease in the lumen size of the interlobular artery. Se supplementation to diabetic rats up-regulated mRNA expression for antioxidant enzymes, and significantly reduced but did not normalize that of TGF-beta1. Glomerular sclerosis was normalized and the interlobular artery lumen size was greatly enlarged in diabetic rats by Se supplementation. Also, the tubulointerstitium was preserved by Se supplementation in diabetic rats. CONCLUSIONS: The data show that TGF-beta1 is a pro-oxidant and Se deficiency increases oxidative stress via this growth factor. In addition, Se deficiency may simulate hyperglycemic conditions. Se supplementation to diabetic rats prevents not only oxidative stress but renal structural injury, as well. PMID- 11260396 TI - 12-lipoxygenase is increased in glucose-stimulated mesangial cells and in experimental diabetic nephropathy. AB - BACKGROUND: Arachidonic acid-derived 12-lipoxygenase (12-LO) products have potent growth and chemotactic properties. The present studies examined whether 12-LO and fibronectin are induced in cultured rat mesangial cells (MCs) exposed to high glucose and whether they are expressed in experimental diabetic nephropathy. METHODS: To determine the effect of high glucose on MC 12-LO mRNA and protein expression, rat MCs were incubated with RPMI medium containing 100 (NG) or 450 mg/dL glucose (HG). For animal studies, rats were injected with diluent (control) or streptozotocin. The latter were left untreated (DM) or treated with insulin (DM + I). At sacrifice after four months, GAPDH, 12-LO, and fibronectin mRNA were measured by competitive reverse transcription-polymerase chain reaction (RT-PCR) in microdissected glomeruli (G). Renal sections were semiquantitatively scored (0 to 4+) for diabetic changes and for 12-LO and fibronectin by immunohistochemistry. RESULTS: 12-LO mRNA expression in MC exposed to HG (12.71 +/- 1.17 attm/microL) and DM G (1.78 +/- 0.65 x 10-3 attm/glomerulus) was significantly higher than those of MCs in NG media (6.71 +/- 0.78 attm/microL) and control G (0.34 +/- 0.12 x 10-3 attm/glomerulus, P < 0.005), respectively. Western blot revealed a 1.7- and a 2.8-fold increase in MC and G 12-LO protein expression, respectively (P < 0.05). The immunohistochemistry score for G 12-LO and diabetic nephropathy score was significantly greater in DM and DM + I than controls. MC and G GAPDH mRNA remained unchanged. CONCLUSIONS: In MCs exposed to HG and in diabetic rat glomeruli, increments in 12-LO mRNA and protein are associated with changes modeling diabetic nephropathy. These findings suggest a role for the 12-LO pathway in the pathogenesis of diabetic nephropathy. PMID- 11260397 TI - Gene expression profile in streptozotocin-induced diabetic mice kidneys undergoing glomerulosclerosis. AB - BACKGROUND: To elucidate the molecular mechanism of diabetic nephropathy, a high density DNA filter array was employed to survey the gene expression profile of streptozotocin-induced diabetic CD-1 (ICR) mouse kidneys. METHODS: Ten-week-old CD-1 male mice were divided into four groups: (1) control, (2) unilaterally nephrectomized (UX) mice, (3) streptozotocin (STZ)-induced diabetic (STZ) mice, and (4) STZ mice with unilateral renal ablation (STZ-UX). Pathological changes were examined at 24 weeks after the induction. The gene expression profile was compared between the control and STZ mice by a Gene Discovery Array (GDA). RESULTS: The glomeruli in UX mouse kidney showed prominent glomerular hypertrophy, while the accumulation of mesangial matrix was minimal. Both STZ and STZ + UX mice had significant glomerular hypertrophy and glomerulosclerosis, and the lesions were not enhanced by renal ablation. By comparison between control and STZ mice, 16 clones that increased in expression with the induction of diabetes and 65 clones that decreased in diabetic kidneys were identified. The 37 known genes were related to glucose and lipid metabolism, ion transport, transcription factors, signaling molecules, and extracellular matrix-related molecules. The genes known to be involved in cell differentiation and organogenesis in various tissues (that is, Unc-18 homolog, POU domain transcription factor 2, lunatic fringe gene homolog, fibrous sheath component 1, Sox-17, fibulin 2, and MRJ) were found to be differentially expressed in the early phase of diabetic kidneys. CONCLUSIONS: Hyperglycemia is a major determinant of glomerulosclerosis in STZ-induced diabetic CD-1 mice, and the altered gene expression in the early phase of diabetic kidney may be critical for the development of diabetic nephropathy. PMID- 11260398 TI - Mast cell chymase expression and mast cell phenotypes in human rejected kidneys. AB - BACKGROUND: Mast cells (MCs) are known to participate in various types of chronic disease, but their role in chronic renal rejection is poorly understood. Recently, distinct phenotypes of MCs have been described in humans by the demonstration of one protease, chymase. Hence, we questioned whether chymase in MCs could play a role in the pathogenesis of renal rejection in humans. METHODS: We investigated MC chymase expression and MC phenotypes, using immunohistochemical single- and double-staining techniques, in nephrectomy (N = 13) and biopsy (N = 8) specimens of human rejected kidneys. Tissue chymase levels were determined by enzymatic assay for chymase activity. We also examined the association between MC chymase expression and the degree of interstitial fibrosis in these renal allografts. RESULTS: Based on chymase positivity, rejected kidneys were divided into two groups, a chymase-negative [Chy(-)] group and a chymase positive [Chy(+)] group. Quantitative analysis showed that the number of chymase positive MCs and tissue chymase levels were significantly higher in the Chy(+) group than in the Chy(-) group. Furthermore, the interstitial fibrotic area in the Chy(+) group was significantly larger than that in the Chy(-) group. Immunodouble staining analysis also demonstrated that a new MC phenotype, positive for chymase but negative for tryptase, was present in the human rejected kidney. CONCLUSIONS: These results show that increased expression of chymase in MCs is related to the severity of interstitial fibrosis in human rejected kidneys. PMID- 11260399 TI - Defective expression of B7-2 (CD86) on monocytes of dialysis patients correlates to the uremia-associated immune defect. AB - BACKGROUND: Specific cellular immune reactions in patients with chronic renal failure (CRF) are impaired by a defect of the antigen-presenting cells. To elucidate the molecular background for this defect, we determined the expression of human lymphocyte antigen (HLA)-DR and costimulatory molecules on monocytes of hemodialysis patients. METHODS: The expression of HLA-DR, B7-1 (CD80), and B7-2 (CD86) molecules was determined on CD14+ monocytes of chronic hemodialysis patients prior to a dialysis session. Mononuclear cells of these patients were cultured, and expression of the respective antigens was determined after in vitro activation by various stimuli. Results were correlated with in vitro proliferation of T cells in a phytohemagglutinin (PHA) assay and the clinical response to a hepatitis B vaccination. All data were compared with healthy controls and patients with CRF who were not on dialysis. RESULTS: Monocytes of chronic hemodialysis patients but not CRF patients expressed low levels of costimulatory B7-2, while HLA-DR expression was normal. B7-1 was only expressed on activated monocytes, and the expression reached normal levels in hemodialysis patients. Baseline expression of B7-2 highly correlated with the results of T cell proliferation assays in hemodialysis patients and also with the clinical immune response. CONCLUSIONS: Impaired expression and up-regulation of B7-2 is an important feature of the cellular immune defect in chronic hemodialysis patients. It leads to reduced costimulation and effector activation of T cells and contributes to a molecular explanation for the impaired response of hemodialysis patients to the hepatitis B vaccination. PMID- 11260400 TI - Prolonged transgene expression in glomeruli using an EBV replicon vector system combined with HVJ liposomes. AB - BACKGROUND: Various gene transfer vectors as well as delivery systems have been developed; however, many problems remain to be solved. We already achieved a technique to introduce genes into glomerular mesangial cells by hemagglutinating virus of Japan (HVJ) liposome-mediated gene transfer via renal artery. The main limitation of this method is the transient transgene expression. METHOD: For long term gene expression in glomeruli, Epstein-Barr virus (EBV) replicon-based plasmid was employed, containing the latent viral DNA replication origin (oriP) and EBV nuclear antigen-1 (EBNA-1), which are the minimum EBV component of transgene-nuclear retention. To examine the effect of EBV replicon apparatus on the duration of transgene expression in glomeruli in vivo, the EBV replicon vector pEBActLuc, and the control plasmid vector pActLuc were adopted. These plasmid vectors were transferred into the kidney via renal artery by using artificial viral envelope (AVE)-type HVJ liposome method, and glomerular luciferase activities were analyzed at various time points after transfection. RESULTS: On day 4, pEBActLuc and pActLuc transfer resulted in equal glomerular luciferase activity, and the luciferase gene expression was sustained for at least 56 days in glomeruli transfected with pEBActLuc, whereas it was reduced on seven days in glomeruli transfected with pActLuc. CONCLUSION: The combination of EBV replicon apparatus and HVJ liposomes appears to be a powerful tool for long term gene expression in vivo, and furthermore, it may be a promising new therapeutic method for the progression of renal disease. PMID- 11260401 TI - Viral delivery of superoxide dismutase gene reduces cyclosporine A-induced nephrotoxicity. AB - BACKGROUND: Cyclosporine A (CsA) increases free radical formation in the kidney. Accordingly, this study investigated whether gene delivery of superoxide dismutase (SOD) reduced radical production and nephrotoxicity caused by CsA. METHODS: Rats were given adenovirus (Ad) carrying lacZ or Cu/Zn-SOD genes three days prior to CsA treatment. Histology, glomerular filtration rates (GFRs) and free radical adducts in urine were assessed. RESULTS: SOD activity was increased 2.5-fold three days after viral infection and remained at 2- and 1.6-fold higher 10 and 17 days later. Treatment with CsA for seven days decreased GFR by 70% in rats infected with Ad-lacZ as expected; however, the decrease was diminished significantly in rats receiving Ad-SOD. CsA treatment for two weeks caused a loss of brush border and dilation of proximal tubules, necrosis, and increased leukocyte infiltration into the kidney; these effects were minimized by SOD. Dimethyl sulfoxide (DMSO) was attacked by the hydroxyl radical to produce a methyl radical. Indeed, administration of CsA with 12C-DMSO in rats infected with Ad-lacZ produced a radical adduct with hyperfine coupling constants similar to 4 POBN/methyl radical adduct and another unknown radical adduct. CsA given with 13C DMSO produced a 12-line spectrum, confirming the involvement of hydroxyl radicals. Free radical adducts detected in urine were increased approximately fivefold by CsA, an effect blocked completely by SOD. CONCLUSIONS: CsA increases free radical formation. Gene delivery of SOD blocks formation of free radicals, thereby minimizing nephrotoxicity caused by CsA. PMID- 11260402 TI - Mouse Na+: HCO3- cotransporter isoform NBC-3 (kNBC-3): cloning, expression, and renal distribution. AB - BACKGROUND: Na+:HCO3- cotransporters mediate the transport of HCO3- into or out of the cell. We recently reported the partial cloning and characterization of a new human Na+:HCO3- cotransporter (referred to as NBC-3 or kNBC-3). The purpose of the present studies was to clone the mouse kNBC-3 and to examine its properties and expression in the kidney. METHODS: Using primers from human kNBC-3 cDNA and 5' and 3' rapid amplification cDNA end polymerase chain reaction (RACE PCR), the mouse kNBC-3 full-length cDNA was cloned from inner medullary collecting duct (mIMCD-3) cells. The tissue distribution and functional properties of NBC-3 was determined using established methods. RESULTS: The coding region of the mouse kNBC-3 has 1089 amino acids and shows 73 and 56% identity to human NBC-2 and NBC-1, respectively. The renal distribution of kNBC-3 demonstrated a unique expression pattern: Whereas kNBC-1 is predominantly expressed in the cortex and is absent in the inner medulla, kNBC-3 shows an intense expression level in the inner medulla and is absent in the cortex. Expression studies in oocytes indicated that NBC-3 mediates Na-dependent HCO3- cotransport. Electrophysiological experiments demonstrated that unlike kNBC-1, which is electrogenic, kNBC-3 is electroneutral. CONCLUSIONS: Based on its distribution and electroneutrality, we propose that kNBC-3 mediates the transport of HCO3- into the cells. PMID- 11260403 TI - Effect of acidosis on urine supersaturation and stone formation in genetic hypercalciuric stone-forming rats. AB - BACKGROUND: We have successively inbred over 45 generations a strain of rats to maximize urine calcium excretion. The rats now consistently excrete 8 to 10 times as much calcium as controls and uniformly form poorly crystalline calcium phosphate kidney stones. In humans with calcium nephrolithiasis, consumption of a diet high in acid precursors is often cited as a risk factor for the development of calcium-based kidney stones; however, the effect of this diet on urinary supersaturation with respect to the common solid phases found in kidney stones has not been determined. METHODS: To determine the effect of the addition of an acid precursor on urine ion excretion, supersaturation, and stone formation, we fed these genetic hypercalciuric stone-forming (GHS) rats 13 g/day of a 1.2% calcium diet with 0.0, 0.5, 1.0, or 1.5% NH4Cl in the drinking water for 14 weeks (N = 8 for each). Urine was collected and analyzed every two weeks. RESULTS: As expected, the addition of dietary NH4Cl led to a progressive fall in urine pH and urine citrate, while urine ammonium increased. Urine calcium and phosphorus increased, while urine oxalate fell. Increasing dietary NH4Cl led to a fall in supersaturation with respect to CaHPO4 (brushite) and CaOx and a rise in supersaturation with respect to uric acid. In spite of differences in supersaturation, most rats in each group formed stones that contained calcium phosphate and not calcium oxalate. CONCLUSIONS: Thus, while the provision of additional dietary acids alters urinary ion excretion and lowers supersaturation with respect to CaHPO4 and CaOx, it does not change the character or rate of stone formation in the GHS rats. PMID- 11260404 TI - Dietary zinc deficiency increases uroguanylin accumulation in rat kidney. AB - BACKGROUND: Zinc deficiency in humans produces a secretory diarrhea that is corrected by zinc supplementation. In rats, differential mRNA display analysis has shown that intestinal uroguanylin gene expression is increased in zinc deficiency. An endocrine axis involving intestinal uroguanylin and the kidney may exist. Therefore, we conducted this study to examine whether zinc deficiency would affect uroguanylin expression in the kidney of rats. METHODS: A purified diet, deficient or adequate in zinc content, was fed to rats. Preprouroguanylin mRNA was localized in kidney by in situ hybridization, and prouroguanylin/uroguanylin peptides were localized in the kidney by immunohistochemistry. Abundance was measured by Western blotting and slot blotting analyses. RESULTS: In situ hybridization demonstrated that preprouroguanylin mRNA-expressing cells were localized in the proximal tubules, being primarily limited to the cortical-medullary junction. Zinc deficiency did not alter the abundance or distribution of the mRNA. Immunohistochemistry, using a uroguanylin peptide-specific, affinity-purified antibody, demonstrated that immunoreactive uroguanylin peptide was localized to the same cells but that the staining was stronger in zinc-deficient rats. Western blotting analysis of kidney extracts showed that there was no difference in abundance of prouroguanylin between zinc adequate and deficient rats. However, slot blotting analysis demonstrated that the abundance of a low molecular weight immunoreactive peptide, presumably uroguanylin, was higher in extracts of zinc-deficient rats. CONCLUSION: The results suggest that production of prouroguanylin by the kidney, in contrast to the intestine, is not influenced by dietary zinc intake, but that higher amounts of uroguanylin in kidney extracts may reflect renal processing of the hormone obtained from the systemic circulation. PMID- 11260406 TI - Continuous measurements of renal perfusion in pigs by means of intravascular Doppler. AB - BACKGROUND: Changes in renal blood flow are considered to play a significant role in the induction and maintenance of kidney failure, but are difficult to monitor with currently available techniques. The objective was to validate renal flow measurements with Doppler guidewires and to apply this technique to assess dose and time dependency of the renal vascular effects of norepinephrine (NE). METHODS: In 10 anesthetized pigs, flow velocity in renal arteries (FVart) and veins (FVvein) and volumetric renal blood flow (VBF) were measured before and after intravenous bolus application of incremental doses of NE (2 to 200 microg). RESULTS: FVart curves exactly reflected the changes in VBF. Beat-to-beat analysis revealed a strong linear correlation over a mean VBF range of less than 0.05 to 0.35 L/min (median correlation coefficient with FVart, r = 0.998), and significant but less close relationships were also found between VBF and FVvein. Ten seconds after the administration of 200 microg NE, FVart dropped from 71 to 6 cm/sec and was 90% reversible after 48 seconds. Similarly, the renal vascular resistance temporarily rose from 988 to 13711 mm Hg. min/L. In contrast, NE induced increases in systemic vascular resistance were on average a maximum of 1.5-fold but persisted for more than 60 seconds. CONCLUSIONS: Doppler flow measurements in the renal artery provide an excellent surrogate of volumetric blood flow, which may be useful for continuous monitoring of renal hemodynamics. The renal vasculature is more sensitive when compared with the systemic vasculature, but also appears to evoke more efficient counter-regulatory mechanisms in response to NE. PMID- 11260405 TI - Renal vascular responses to captopril and to candesartan in patients with type 1 diabetes mellitus. AB - BACKGROUND: Enhanced renal vasodilator responses to angiotensin-converting enzyme (ACE) inhibition in diabetes mellitus despite a normal or low plasma renin activity level have suggested intrarenal activation of the renin-angiotensin system in this disease. There is, however, a continuing debate as to the mediators of the renal hemodynamic response to ACE inhibition-reduced angiotensin II formation or pathways involving kinins, prostaglandins, and nitric oxide. METHODS: Twelve patients with type 1 diabetes mellitus of 18 +/- 3.2 (SEM) years of duration (7 females and 5 males, ages 17 to 50, 32 +/- 4.0 years) who were free of sustained microalbuminuria and on a high-salt diet were given the ACE inhibitor captopril (25 mg orally) on one day and the AT1 receptor blocker candesartan (16 mg orally) on another day. Renal plasma flow (RPF) and glomerular filtration rate were measured before and for four hours after administration. RESULTS: Both drugs caused a significant increase in RPF (captopril 574 +/- 26 to 625 +/- 37 mL/min/1.73 m2, P = 0.008; candesartan 577 +/- 26 to 643 +/- 37, P = 0.004). There was a highly significant correlation between the responses to captopril and to candesartan (r = 0.86, P < 0.001). Seven subjects had an RPF response to captopril that was accentuated (90 +/- 13 mL/min/1.73 m2), while five had a response that was normal (-4 +/- 9). There was no significant change in glomerular filtration rate on either drug. CONCLUSION: The remarkable rise in RPF in response to captopril and candesartan despite high-salt balance suggests the intrarenal activation of the renin-angiotensin system in diabetes that is not reflected in plasma renin levels. The high correlation between the renal hemodynamic response to captopril and to candesartan indicates that reduced angiotensin II formation is the main mechanism of action of the ACE inhibitor. PMID- 11260407 TI - Heme oxygenase isoform-specific expression and distribution in the rat kidney. AB - BACKGROUND: The heme oxygenase (HO) genes, HO-1 and HO-2, are the limiting steps in heme degradation and in the regulation of renal heme-dependent enzymes. Previously we reported that selective overexpression of renal HO-1 resulted in a decrease of microsomal heme and the cytochrome P450-dependent arachidonic acid metabolite, 20 HETE, a vasoconstrictor. The present study was undertaken to explore the relative expression and contribution of each of the HO isoforms to HO activity in the rat kidney. METHODS AND RESULTS. Renal HO activity increased above control levels after an injection of the inducers of HO activity, heme or SnCl2. Stannous Mesoporphyrin (SnMP), a nonselective inhibitor of HO, when used alone or in combination with heme or SnCl2, decreased HO activity. Heme alone and combined with SnCl2 decreased the levels of heme content by 13 and 35%, respectively. Western blot analysis showed that both SnCl2 and heme readily induced HO-1 protein, whereas HO-2 was constitutively expressed. Immunohistochemistry showed the distribution of the HO-1 isoform primarily in proximal convoluted tubules. Western blot analysis exhibited relatively higher levels of HO-1 in isolated proximal tubules and relatively higher HO-2 levels in the thick ascending limbs of the loop of Henle and preglomerular arterioles. In vivo administration of HO-1 and HO-2 antisense oligodeoxynucleotides further confirmed that HO-2, but not HO-1, contributed to the basal HO activity; however, following induction of HO with heme, antisense to HO-1, but not to HO-2, inhibited the induced levels of HO activity. CONCLUSION: These results suggest that HO-2 is constitutively expressed in the rat kidney mainly within tubular and arteriolar structures, and its activity may modulate physiological function under basal conditions. On the other hand, the basal levels of expression of HO-1 in the rat kidney are relatively low, and its contribution to HO activity and the regulation of hemoproteins such as cytochrome P450 become apparent only under pathophysiological conditions causing HO induction. PMID- 11260408 TI - Intrarenal angiotensin and bradykinin peptide levels in the remnant kidney model of renal insufficiency. AB - BACKGROUND: The remnant kidney model of renal failure is associated with normal or suppressed plasma renin and angiotensin (Ang) II levels when hypertension is established. However, the hypertension responds to angiotensin-converting enzyme (ACE) inhibition and Ang II receptor antagonism, suggesting a role for Ang II in the hypertensive process. Bradykinin (BK) is a potent vasoactive peptide that may also participate in this model. METHODS: Ang II and BK peptides were measured in the ischemic peri-infarct portion and the intact portion of the remnant kidney at two, five, and seven weeks after surgery. Plasma Ang II, renin, angiotensinogen, and aldosterone levels were also measured. RESULTS: Ang II levels in the peri infarct portion were higher than in the intact portion at all time points and were higher than in sham-operated kidney at two weeks. Ang II levels in the intact portion were similar to the levels in kidneys of sham-operated rats at two and five weeks and were suppressed at seven weeks. BK levels were increased in the peri-infarct portion at all time points and in the intact portion at two and five weeks. Plasma Ang II and aldosterone levels were also elevated at two weeks. CONCLUSIONS: Peri-infarct renal tissue Ang II levels and plasma Ang II and aldosterone levels increase transiently during the evolution of hypertension in the remnant kidney model. Sustained hypertension is associated with an increase in intrarenal BK levels but not with persistent increases in intrarenal or circulating Ang II levels. PMID- 11260411 TI - Incidental renal artery stenosis in peripheral vascular disease: a case for treatment? AB - BACKGROUND: Renal artery stenosis (RAS) is frequently encountered as an incidental finding in peripheral vascular disease. We assessed whether revascularization is indicated to prevent the practical consequences of end-stage renal failure, that is, the need for renal replacement therapy. METHODS: In a retrospective study, a cohort of consecutive patients was followed who had undergone angiography 8 to 10 years previously for peripheral artery disease. Patients with untreated incidental RAS of > or =50% diameter stenosis (68.8 +/- 9.8 years, mean +/- SD) were compared with regard to the prevalence of renal replacement therapy to controls without RAS who were matched for age and gender. RESULTS: RAS was present in 126 of 386 evaluable patients (33%). None of these patients required renal replacement therapy during the 10-year follow-up. Serum creatinine values remained stable during follow-up. CONCLUSIONS: Incidental RAS is frequently seen in patients with peripheral vascular disease. If left untreated, incidental RAS seems not to result in end-stage renal failure or in a need for renal replacement therapy. Revascularization with the aim to prevent end stage renal failure seems less indicated, and further prospective studies are indicated to elucidate this issue. PMID- 11260410 TI - Effect of fluvastatin on endothelium-dependent brachial artery vasodilation in patients after renal transplantation. AB - BACKGROUND: Hypercholesterolemia may affect both endothelial function and arterial distensibility (DC). Renal transplant recipients (NTX) exhibit advanced structural and functional alterations of arterial vessel walls. The aim of this double-blind, randomized trial was to evaluate the effects of fluvastatin (FLU) on brachial artery flow-mediated vasodilation (FMD) and DC in hypercholesterolemic NTX. METHODS: Eighteen NTX received FLU 40 mg/day and 18 NTX placebo (PLA). Before and after six months of treatment, the brachial artery diameter and DC at rest were measured by a Doppler frequency analysis in the M mode, and then changes in diameter during reactive hyperemia (to assess endothelial-dependent FMD) and after 400 microg sublingual nitroglycerin (to assess endothelium-independent vasodilation-NMD). RESULTS: FLU, but not PLA, treatment resulted in significant decreases in total (from 288 +/- 10 to 239 +/- 8 mg/dL, P < 0.05) and low-density lipoprotein cholesterol (from 182 +/- 779 to 138 +/- 8 mg/dL, P < 0.05). Blood pressure did not differ between FLU- and PLA treated patients and was not affected by either treatment. Also, the brachial artery baseline diameter was not different between groups and was not affected by FLU or PLA. Brachial artery flow at rest and during reactive hyperemia as measured by pulsed Doppler did not differ between groups. Brachial artery FMD increased with FLU from 0.23 +/- 0.08 to 0.54 +/- 0.08 mm (P < 0.05), whereas PLA did not alter FMD (0.22 +/- 0.07 vs. 0.14 +/- 0.05 mm at baseline and after six months of PLA treatment, respectively, P = NS). In contrast, NMD did not change significantly with either treatment (0.76 +/- 0.13 vs. 0.83 +/- 0.15 mm at baseline and after 6 months of FLU treatment, respectively, P = NS, and 0.64 +/- 0.09 vs. 0.66 +/- 0.10 mm at baseline and after 6 months of PLA treatment, respectively, P = NS). Also, brachial artery DC was not altered by FLU (6.4 +/- 1.0 vs. 5.8 +/- 0.6 x 10-3/kPa, P = NS) or PLA treatment (5.8 +/- 0.6 vs. 6.8 +/- 0.8 x 10-3/kPa, P = NS). CONCLUSIONS: In hypercholesterolemic NTX, the HMG-CoA reductase inhibitor FLU significantly improves brachial artery FMD as a measure of endothelial function after six months of treatment. In contrast, FLU does not have a beneficial effect on brachial artery DC. PMID- 11260409 TI - Circulating endothelial nitric oxide synthase inhibitory factor in some patients with chronic renal disease. AB - BACKGROUND: Chronic renal disease (CRD) is associated with hypertension and reduced synthesis of nitric oxide (NO). Here, we investigated whether there is a circulating endothelial NO synthase (eNOS) inhibitory factor(s) in some patients with CRD that might directly influence endothelial NOS. METHODS: Human dermal microvascular endothelial cells (HDMECs) were incubated for six hours with 20% plasma from subjects with normal renal function (PCr = 0.8 +/- 0.2 mg%), and patients with moderate renal insufficiency of various causes (PCr = 4.0 +/- 1.5 mg%) and impact on NOS activity, transport of L-arginine, and abundance of eNOS protein were measured. Plasma concentrations of asymmetric and symmetric dimethyl L-arginine (ADMA and SDMA) were also measured. RESULTS: There was no effect of any human plasma on L-arginine transport. The NOS activity was variable in CRD patients and fell into two subgroups: CRD I, individual values similar to control, and CRD II, individual values lower than control mean. The effect of CRD plasma on NOS activity in cultured cells was not related to the primary disease, but was predicted by plasma ADMA levels since plasma ADMA was elevated in CRD II versus both control and CRD I. Blood urea nitrogen and creatinine levels were uniformly elevated in CRD plasma. The abundance of eNOS protein was unaffected by plasma. CONCLUSION: High plasma levels of ADMA in CRD patients are independent of reduced renal clearance, suggesting an alteration in ADMA synthesis and/or degradation. High ADMA is a marker and is partly responsible for the inhibition of eNOS activity in cultured cells and may also result in reduced eNOS activity in vivo, with consequent hypertension. PMID- 11260412 TI - Cyclosporine in patients with steroid-resistant membranous nephropathy: a randomized trial. AB - BACKGROUND: A clinical trial of cyclosporine in patients with steroid-resistant membranous nephropathy (MGN) was conducted. Although MGN remains the most common cause of adult-onset nephrotic syndrome, its management is still controversial. Cyclosporine has been shown to be effective in cases of progressive MGN, but it has not been used in controlled studies at an early stage of the disease. METHODS: We conducted a randomized trial in 51 biopsy-proven idiopathic MGN patients with nephrotic-range proteinuria comparing 26 weeks of cyclosporine treatment plus low-dose prednisone to placebo plus prednisone. All patients were followed for an average of 78 weeks, and the short- and long-term effects on renal function were assessed. RESULTS: Seventy-five percent of the treatment group versus 22% of the control group (P < 0.001) had a partial or complete remission of their proteinuria by 26 weeks. Relapse occurred in 43% (N = 9) of the cyclosporine remission group and 40% (N = 2) of the placebo group by week 52. The fraction of the total population in remission then remained almost unchanged and significant different between the groups until the end of the study (cyclosporine 39%, placebo 13%, P = 0.007). Renal function was unchanged and equal in the two groups over the test medication period. In the subsequent follow up, renal insufficiency, defined as doubling of baseline creatinine, was seen in two patients in each group, but remained equal and stable in all of the other patients. CONCLUSION: This study suggests that cyclosporine is an effective therapeutic agent in the treatment of steroid-resistant cases of MGN. Although a high relapse does occur, 39% of the treated patients remained in remission and were subnephrotic for at least one-year post-treatment, with no adverse effect on filtration function. PMID- 11260413 TI - Upper extremity digital subtraction venography with gadoterate meglumine before fistula creation for hemodialysis. AB - BACKGROUND: The purpose of this study was to evaluate the feasibility, safety, and potential role of gadoterate meglumine (Gd-DOTA) as a contrast agent for upper extremity venography before the creation of an arteriovenous fistula (AVF) for nondialyzed renal insufficiency patients. METHODS: Over a 16-month period, 50 venographies were performed on end-stage renal insufficiency patients, using Gd DOTA as a contrast agent on a high-resolution digital subtraction angiography system. Three sequences were performed on forearm, arm, and chest at 3 mL/sec for a total of 35 mL of Gd-DOTA. Examinations were reviewed by two radiologists for diagnostic and opacification quality. Tolerance was evaluated on the evolution of serum creatinine levels and occurrence of pain during injection. RESULTS: Good interobserver correlation was obtained in evaluating the feasibility of AVF creation by vein segment (0.64 < kappa < 0.88) and in relationship to opacification quality (0.62 to 0.87). No deterioration in renal function (creatinine level before and after) or pain was observed. Twenty-six patients underwent surgical creation of brachiobasilic (N = 8), brachiocephalic (N = 8), radiocephalic (N = 8), and cubitocephalic (N = 1) fistulas or insertion of a polytetrafluoroethylene (PTFE) graft (N = 1). Seventeen were awaiting AVF or were on peritoneal dialysis. Two died before surgery for reasons unconnected with the venography. CONCLUSIONS: Venography with Gd-DOTA is an effective and safe technique in planning AVFs for renal insufficiency patients. PMID- 11260414 TI - Obesity-related glomerulopathy: an emerging epidemic. AB - BACKGROUND: We report the first large renal biopsy-based clinicopathologic study on obesity-related glomerulopathy. METHODS: Obesity was defined as body mass index (BMI)> 30 kg/m2. Obesity-related glomerulopathy (ORG) was defined morphologically as focal segmental glomerulosclerosis and glomerulomegaly (O FSGS; N = 57) or glomerulomegaly alone (O-GM; N = 14). RESULTS: Review of 6818 native renal biopsies received from 1986 to 2000 revealed a progressive increase in biopsy incidence of ORG from 0.2% in 1986-1990 to 2.0% in 1996-2000 (P = 0.0001). Mean BMI in ORG was 41.7 (range 30.9 to 62.7). Indications for renal biopsy included proteinuria (N = 40) or proteinuria and renal insufficiency (N = 31). Seventy-one patients with ORG were compared to 50 patients with idiopathic FSGS (I-FSGS). Patients with ORG were older (mean 42.9 vs. 32.6 years, P < 0.001) and more often Caucasian (75% vs. 52%; P = 0.003). ORG patients had a lower incidence of nephrotic range proteinuria (48% vs. 66%; P = 0.007) and nephrotic syndrome (5.6% vs. 54%; P < 0.001), with higher serum albumin (3.9 vs. 2.9 g/dL; P < 0.001), lower serum cholesterol (229 vs. 335 mg/dL; P < 0.001), and less edema (35% vs. 68%; P = 0.003). On renal biopsy, patients with ORG had fewer lesions of segmental sclerosis (10 vs. 39%; P < 0.001), more glomerulomegaly (100% vs. 10%; P < 0.001), and less extensive foot process effacement (40 vs. 75%; P < 0.001). Glomerular diameter in ORG (mean 226 mu) was significantly larger than age- and sex-matched normal controls (mean 168 mu; P < 0.001). Follow up was available in 56 ORG patients (mean 27 months) and 50 idiopathic FSGS controls (mean 38 months). A total of 75% of ORG patients received angiotensin converting enzyme (ACE) inhibition or A2 blockade while 78% of the I-FSGS patients received immunosuppressive therapy. ORG patients had less frequent doubling of serum creatinine (14.3% vs. 50%; P < 0.001) and progression to ESRD (3.6% vs. 42%; P < 0.001). On multivariate analysis, presenting serum creatinine and severity of proteinuria were the only predictors of poor outcome in ORG. CONCLUSION: ORG is distinct from idiopathic FSGS, with a lower incidence of nephrotic syndrome, more indolent course, consistent presence of glomerulomegaly, and milder foot process fusion. The ten-fold increase in incidence over 15 years suggests a newly emerging epidemic. Heightened physician awareness of this entity is needed to ensure accurate diagnosis and appropriate therapy. PMID- 11260415 TI - Incidence, risk factors, and prognosis of gastrointestinal hemorrhage complicating acute renal failure. AB - BACKGROUND: Few prospective data are currently available on acute gastrointestinal hemorrhage (AGIH) as a complication of acute renal failure (ARF). The aim of the present study was to define incidence, sources, risk factors, and outcome of AGIH in patients with ARF. METHODS: We performed a prospective study on an inception cohort of 514 patients admitted for ARF to a nephrology intermediate care unit. Data on clinical risk factors for bleeding, frequency of occurrence of AGIH, length of hospital stay, and in-hospital mortality were collected. Independent predictors of AGIH were identified. The relative odds of death and the relative increase in length of hospital stay associated with AGIH were calculated after adjusting for baseline comorbidities. RESULTS: Sixty-nine patients out of 514 [13.4% (95% CI, 10.6 to 16.7)] had AGIH as a complication of ARF; 59 were upper AGIH. Forty patients had clinically important bleeding. Erosions and/or ulcers accounted for 71% of cases of upper AGIH. Independent baseline predictors of AGIH were represented by severity of illness [odds ratio 1.45 (95% CI, 1.05 to 2.01) for every 10 point increase in APACHE II score], low platelet count [<50,000 mm3; 3.71 (1.70 to 8.11)], noncirrhotic chronic hepatic disease [2.22 (1.09 to 4.55)], liver cirrhosis [3.38 (1.50 to 7.60)], de novo ARF [2.77 (1.30 to 5.90)], and severe ARF [2.07 (1.10 to 3.88)]. In-hospital mortality was 63.8% in patients with AGIH and 34.2% in the other patients; after adjusting for baseline confounders, AGIH remained significantly associated with an increase in both mortality [2.57 (1.30 to 5.09), P = 0.006] and length of hospital stay [37% (1 to 87%), P = 0.047]. CONCLUSIONS: AGIH and clinically important bleeding are frequent complications of ARF. In this clinical condition, AGIH is more often due to upper gastrointestinal bleeding and is associated with a significantly increased risk of death and length of hospital stay. Both renal and extrarenal risk factors are related to the occurrence of AGIH. PMID- 11260416 TI - Plasminogen activator inhibitor-1 gene polymorphism 4G/4G genotype and lupus nephritis in Chinese patients. AB - BACKGROUND: Abnormal regulation in the coagulation and fibrinolytic system may play an important role in mediating glomerular damage in lupus nephritis. Indeed, glomerular thrombosis occurs frequently in lupus nephritis and predicts the future development of glomerular sclerosis. In the murine model of active lupus nephritis, plasminogen activator inhibitor-1 (PAI-1) gene was overexpressed throughout the kidney, both within the glomeruli and also in tubules and vessels. The level of PAI-1 expression in the tissues appeared to correlate with the progression of lupus nephritis. Recently, a single base pair insertion/deletion 4G/5G polymorphism of the PAI-1 gene has been identified and shown to alter plasma PAI-1 activity. This study was therefore conducted to determine the association of the 4G/5G polymorphism of the PAI-1 gene with the development and severity of lupus nephritis. METHODS: The PAI-1 gene polymorphism of 118 systemic lupus erythematosus (SLE) patients and 103 healthy controls who were gender and age matched was determined using standard polymerase chain reaction. PAI-1 genotype results were studied in relationship to the development and severity of lupus nephritis. RESULTS: Allele frequencies of 4G/5G allele were 0.59/0.41 in lupus patients and 0.59/0.41 in controls (P = 1.000). No significant difference was noted in the genotype distribution between SLE patients with and without nephritis. However, lupus nephritis patients with the 4G4G genotype showed significantly heavier proteinuria (5.0 vs. 3.7 g/day; P = 0.023) when compared with patients with 4G5G and 5G5G genotypes. Also, 73.3% patients with 4G4G had an activity index > or =8 versus 37.3% patients with 4G5G and 5G5G (P = 0.003). Extensive necrotizing lesions were seen in 51.7% patients with 4G4G as compared with 23.5% patients with 4G5G and 5G5G (P = 0.014). The association of the 4G4G gene polymorphism with a higher nephritis activity and more severe necrotizing lesions persisted when only class III and class IV nephritis patients were studied. On the other hand, no significant association was noted between the PAI 1 gene polymorphism and the chronicity of the nephritis. CONCLUSION: These findings suggest that the 4G/5G polymorphism of the PAI-1 gene is associated with the activity but not the chronicity of lupus nephritis. The presence of the 4G4G genotype does not increase the risk of developing SLE or lupus nephritis, but predicts the development of higher nephritis activity and more extensive necrotizing lesions. PMID- 11260417 TI - Bicarbonate/lactate-based peritoneal dialysis solution increases cancer antigen 125 and decreases hyaluronic acid levels. AB - BACKGROUND: In a randomized, controlled trial comparing a pH neutral, bicarbonate/lactate (B/L)-buffered PD solution to conventional acidic, lactate buffered solution (C), the overnight dialysate levels of markers of inflammation/wound healing [hyaluronic acid (HA)], mesothelial cell mass/membrane integrity [cancer antigen 125 (CA125)], and fibrosis [transforming growth factor beta1 (TGF-beta1) and procollagen I peptides (PICP)] were assessed over a six month treatment period. METHODS: One hundred six patients were randomized (2:1) to either the B/L group or C group. Overnight effluents were collected at entry into the study (time = 0 all patients on control solution) and then at three and six months after randomization. Aliquots were filtered, stored frozen, and assayed for HA, CA125, TGF-beta1, and PICP. Differences between groups were assessed by repeated-measures analysis of variance for unbalanced data using the SAS procedure MIXED. RESULTS: In patients treated with B/L, there was a significant (P = 0.03) increase in CA125 after six months compared with time = 0 (19.76 +/- 11.8 vs. 24.4 +/- 13.8 U/mL; mean +/- SD; N = 51). In the same group of patients, HA levels were significantly decreased at both three and six months in the B/L-treated group (time = 0, 336.0 +/- 195.2; time = 3 months, 250.6 +/- 167.6; and time = 6 months, 290.5 +/- 224.6 ng/mL; mean +/- SD; P = 0.006, N = 47 and P = 0.003, N = 48, respectively). No significant changes in CA125 or HA levels were observed in the control group. There were no significant changes observed in the levels of PICP or TGF-beta1 in the B/L or C group over the six month treatment period. CONCLUSIONS: These results suggest that continuous therapy with the B/L solutions modulates the levels of putative markers of peritoneal membrane integrity and inflammation. In the long term, this may positively impact the peritoneal membrane, increasing its life as a dialyzing organ. PMID- 11260418 TI - Exponentially increased risk of infectious death in older renal transplant recipients. AB - BACKGROUND: The benefit of renal transplantation for patients with end-stage renal disease (ESRD) has been well documented. This benefit is seen throughout all age ranges of patients. However, it has been documented that older renal transplant recipients are at increased risk for death because of infectious causes when compared with younger recipients. The present study addresses whether this increased risk merely parallels an age-related increase in infectious mortality or is reflective of a particular vulnerability in older renal transplant recipients. METHODS: Patients wait-listed and transplanted between 1988 and 1997 were analyzed utilizing the United States Renal Data System (USRDS) database. The primary study end point was patient death secondary to infection. Secondary end points included death secondary to cardiovascular cause and malignancy. Cox-proportional hazard models were utilized with all pertinent variables. RESULTS: Death related to infectious cause increased exponentially in transplanted patients with increasing age (slope = 2.90.34x), while it increased linearly (slope = 1.9x + 8.6) with increasing age for those patients on the waiting list. Overall mortality increases with age were equal between the wait listed and transplanted groups. CONCLUSIONS: The overall survival benefit of transplantation is maintained in the older age groups. However, renal transplantation is associated with an increased risk for infectious death beyond the expected age-related increased risk in patients on the renal transplant waiting list. This may have an impact on future immunosuppressive regimens in this population. PMID- 11260420 TI - Graft surveillance: venous pressure, access flow, or the combination? AB - BACKGROUND: Increased venous pressure (VP) and decreased access flow (Qa) are predictors of dialysis access graft thrombosis. VP is easily obtainable. Qa assessment requires a special device and takes more time. The aims of our randomized multicenter studies were to compare outcome in patients with grafts monitored by VP or Qa (study A) or monitored by VP or the combination of VP and Qa (study B). METHODS: We performed VP measurements consisting of weekly VP at a pump flow of 200 mL/min (VP200) and the ratio of VP0/MAP. Qa was measured every eight weeks with the Transonic HD01 hemodialysis monitor. Threshold levels for referral for angiography were VP200> 150 mm Hg or VP0/MAP> 0.5 (both at 3 consecutive dialysis sessions) or Qa <600 mL/min. Subsequent therapy consisted of either percutaneous transluminal angioplasty (PTA) or surgery. RESULTS: Total follow-up was 80.5 patient-years for 125 grafts. The vast majority of a total of 131 positive tests was followed by angiography and corrective intervention. In study A, the rate of thromboses not preceded by a positive test was 0.19 and 0.24 per patient-year (P = NS), and in study B, it was 0.32 versus 0.28 per patient year (P = NS). Survival curves were not significantly different between the subgroups. CONCLUSIONS: These data demonstrate that standardized monitoring of either VP or Qa or the combination of both and subsequent corrective intervention can reduce thrombosis rate in grafts to below the recommended quality of care standard (that is, 0.5 per patient-year, NKF-DOQI). These surveillance strategies are equally effective in reducing thrombosis rates. PMID- 11260419 TI - A novel immunoadsorption device for removing beta2-microglobulin from whole blood. AB - BACKGROUND: High plasma levels of beta2-microglobulin (beta2m) have been implicated in the formation of the severely destructive and potentially fatal amyloid deposits that are characteristic of dialysis-related amyloidosis (DRA). Conventional renal replacement technologies remove insufficient quantities of beta2m to normalize plasma levels. This limitation arises because of nonspecific adsorptive qualities and reliance on size exclusion, which can also remove other middle molecular weight proteins. These nonspecific approaches also make it difficult to evaluate the role and contribution of middle molecular weight molecules to the pathology of DRA and other morbidities of end-stage renal disease. A high-affinity and biologically specific approach could target a protein, prevent a significant loss of other important molecules, and improve the apparent adsorption rate within an extracorporeal device. METHODS: Agarose immobilized murine anti-human beta2m monoclonal antibodies were used in a Vortex Flow Plasmapheretic Reactor (VFPR) to remove donor baseline and controlled amounts of recombinant beta2m from human blood in vitro. The extracorporeal circuit was hemoperfused at 200 mL/min for two hours. RESULTS: The immunoadsorptive media had a binding site density of 30 microg beta2m per mL of settled gel. The VFPR cleared baseline quantities of donor beta2m below detectable limits of the assay. The experiments with higher initial beta2m concentrations reached an equilibrium concentration within 20 minutes, corresponding to a 92% clearance. No deleterious hemocompatibility issues were observed (complete blood count, total protein, and plasma free hemoglobin). CONCLUSIONS: The adsorptive kinetics of the VFPR are optimal for the conditions used and support the use of immunoadsorption for the removal of beta2m. PMID- 11260421 TI - Diagnostic potential of cardiac natriuretic peptides in dialysis patients. AB - BACKGROUND: In the general population, the plasma concentrations of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) are useful to predict left ventricular hypertrophy (LVH) and LV systolic dysfunction. Whether these cardiac hormones have a similar diagnostic potential in dialysis patients is unknown. METHODS: We studied the diagnostic value of ANP and BNP for alterations in LV mass and function in a cohort of 246 dialysis patients without clinical evidence of heart failure. RESULTS: Both ANP and BNP were independently related to left ventricular mass (P < 0.0001) as well as to ejection fraction (P < 0.0001). In an analysis based on a prospectively defined threshold (95th percentile of the normal range), BNP had a significantly higher (P < 0.01) sensitivity (88%) than ANP (51%) for the diagnosis of LVH, but the positive predictive value of the two peptides was very similar (92 and 87%, respectively, P = NS). However, the negative predictive value of BNP for excluding LVH was 22% higher than that of ANP (53 vs. 31%, P = 0.05). Both natriuretic peptides had a high sensitivity for the detection of LV dysfunction (87 and 94%), but their positive predictive value was low (25 and 15%). Importantly, both ANP and BNP proved to be very useful for excluding this alteration (negative predictive value 97 and 96%, respectively). An analysis based on the "best cut-offs" of each peptide as identified on the basis of the ROC curves augmented the positive and negative prediction values of BNP for the diagnosis of LVH to 95 and 61%, respectively. This approach also raised the BNP-positive prediction value for the identification of LV dysfunction to 31% but did not modify the diagnostic potential of ANP (either for LVH or for LV dysfunction). CONCLUSIONS: Measuring the plasma concentration of cardiac natriuretic hormones, particularly BNP, may be useful for the identification of dialysis patients with LVH or for excluding systolic dysfunction. PMID- 11260422 TI - Long-term impact of discontinued or reduced calcineurin inhibitor in patients with chronic allograft nephropathy. AB - BACKGROUND: Chronic allograft nephropathy is the major cause of progressive renal failure in renal transplant recipients. It has no definitive treatment. METHODS: One hundred eighteen renal transplant recipients with declining kidney function and biopsy-proven chronic allograft nephropathy had their cyclosporine or tacrolimus dose reduced or discontinued with either the addition or continuation of mycophenolate mofetil and low-dose steroids at a mean of 853.3 days post transplantation. Their renal function was modeled before and after this intervention by two methods: A least-square regression was used to assess the decay of renal function after the intervention and to compare that with the slope pre-intervention, whereas a hinge regression line method was used to assess the correlation of the intervention with the inflection point and the impact of the intervention on the decay of renal function. Mean follow-up was 651.0 days after the intervention. Serum creatinine at the time of intervention was 2.8 +/- 0.9 mg/dL in the reduced dose cyclosporine (N = 67) and reduced dose tacrolimus (N = 33) groups, and was 2.7 +/- 0.7 mg/dL in the group with discontinued calcineurin inhibitor (N = 18). RESULTS: Using the least-square method, 91.7% of the no calcineurin inhibitor group, 51.6% of the reduced dose cyclosporine group, and 59.3% of the reduced dose tacrolimus group had improved or lack of deterioration in slope after the intervention. Using the hinge regression line method, there was a statistically significant correlation of the inflection point with the intervention (P = 0.001). Moreover, there was a similar relationship with stabilized or improved graft function observed with the hinge regression line method and the least-square method, as 72.2% of the calcineurin inhibitor withdrawal group, 54.4% of reduced-dose cyclosporine group, and 40% of the reduced-dose tacrolimus group had improved the slope of decay of renal function or lack of deterioration after the inflection point. The difference between the calcineurin inhibitor withdrawal group and the reduced-dose cyclosporine/tacrolimus groups on the decay in renal function was significant (P = 0.038) with the least-square method and nearly significant (P = 0.056) using the hinge regression line method. CONCLUSION: This intervention was safe, well tolerated, and associated with a minimal risk of acute rejection. We conclude that the reduction and possible withdrawal of calcineurin inhibitors may be necessary to slow the rate of loss of renal function in patients with chronic allograft nephropathy and deteriorating renal function. PMID- 11260423 TI - Determinants of enhanced thromboxane biosynthesis in renal transplantation. AB - BACKGROUND: Despite great improvement in patient and graft survival, the long term morbidity and mortality in renal transplant recipients (RTRs) are still significant, with a high incidence of cardiovascular disease-related deaths. METHODS: We investigated thromboxane (TXA2) biosynthesis and endothelial and coagulative activation in 65 patients who received a renal transplant. RESULTS: The rate of TXA2 biosynthesis (urinary 11-dehydro-TXB2 excretion largely reflects platelet TXA2 production in vivo) was significantly (P < 0.0001) higher in RTRs than in healthy subjects. Plasma von Willebrand factor (vWF) and thrombin antithrombin (TAT) complexes were significantly higher (P < 0.001) in RTRs compared with controls. Urinary 11-dehydro-TXB2 directly correlated with plasma vWF and cholesterol. We next examined the relative influence of cyclosporine A (CsA) on TXA2 biosynthesis and endothelial activation, comparing a group of RTRs not receiving CsA with an age- and sex-matched group of patients treated with CsA. Urinary excretion of 11-dehydro-TXB2 and plasma levels of vWF were significantly increased in RTRs who received CsA compared with those who did not. After an overall follow-up of 120 months, RTRs who experienced cardiovascular events had a higher frequency of abnormal plasma levels of vWF than patients who remained event free. CONCLUSION: Renal transplantation is associated with in vivo platelet activation highly related to endothelial activation. This is particularly evident in CsA-treated patients. Administration of drugs that are able to reduce or eliminate thromboxane-dependent platelet activation in vivo may be beneficial to reduce the risk of cardiovascular events in RTRs. PMID- 11260424 TI - Substitution of renal function through skin catharsis: evidence from the classical period to the Middle Ages. AB - The skin's cleansing capacity has been known for centuries and has been used therapeutically and extensively for a great number of diseases. We studied the historical evolution of the methods used for catharsis through the skin, particularly for those in renal failure, by reviewing most of the existing ancient Greek and Byzantine codices dealing with the skin's cleansing capacity. From the fragments cited in this article, it is evident that the ancient medical writers were well aware of the mechanism of perspiration, and through this process the excretion of several body toxins, they knew about renal failure as well as the influence of environmental temperature on blood purification via the skin. To validate their views, we reviewed the seasonal variation of the average values for blood urea, creatinine, and electrolytes for 161 regular dialysis treatment (RDT) patients in four dialysis units in southern Greece. The estimations were carried out during the winter/summer 1997, 1998, and 1999 terms and included three winter months and three summer months. We traced an unexpectedly large number of references in the ancient and medieval Greek medical literature concerning detoxification through the skin, mainly regarding patients in renal failure. This seasonal variation hypothesis is supported by the results of our retrospective study: there was a difference of 16 mg/dL in the average blood urea (mean winter urea 182 mg/dL, mean summer urea 166 mg/dL). We suggest that the ancients had a vivid idea about the substitution of renal function by the skin's cleansing ability in renal failure. The previously mentioned phenomenon may be due to the elimination of blood urea through excessive perspiration. Our clinical results seem to verify their notions, and hence, the skin (like the peritoneum) may be considered a natural membrane for dialysis. We were unable to trace a similar report in the literature on the seasonal fluctuation of blood urea in dialysis patients. PMID- 11260425 TI - Endothelin: the yin and yang of ischemic acute renal failure. PMID- 11260426 TI - Platelet-derived growth factor: a new clinical target on the horizon. PMID- 11260427 TI - Expanding the organ donor pool: the Spanish Model. PMID- 11260428 TI - Bone mass evolution after renal transplantation. PMID- 11260429 TI - Selectivity of proteinuria can be estimated reliably from samples of second morning urine. PMID- 11260431 TI - Cited "validation" references for the SphygmoCor device. PMID- 11260433 TI - Psychosocial factors in dialysis patients. PMID- 11260435 TI - Cultural identity and representing culture in medical education. Who does it? PMID- 11260437 TI - Nonsense on stilts. PMID- 11260436 TI - Education for communication: much already known, so much more to understand. PMID- 11260438 TI - The "tropical triangle": a health education partnership in the south-west Pacific. PMID- 11260439 TI - The new NHS and undergraduate medical education. PMID- 11260440 TI - Barriers to breaking bad news among medical and surgical residents. AB - Communicating "bad news" to patients and their families can be difficult for physicians. OBJECTIVE: This qualitative study aimed to examine residents' perceptions of barriers to delivering bad news to patients and their family members. DESIGN: Two focus groups consisting of first- and second-year medical and surgical residents were conducted to explore residents' perceptions of the bad news delivery process. The grounded theory approach was used to identify common themes and concepts, which included: (1) guidelines to delivering bad news, (2) obstacles to delivering bad news and (3) residents' needs. SETTING: McMaster University, Hamilton, Ontario, Canada. SUBJECTS: First- and second-year residents. RESULTS: Residents were able to identify several guidelines important to communicating the bad news to patients and their family members. However, residents also discussed the barriers that prevented these guidelines from being implemented in day-to-day practice. Specifically, lack of emotional support from health professionals, available time as well as their own personal fears about the delivery process prevented them from being effective in their roles. Residents relayed the need for increased focus on communication skills and frequent feedback with specific emphasis on the delivery of bad news. The residents in our study also stressed the importance of processing their own feelings regarding the delivery process with staff. CONCLUSIONS: Although most residents realize important guidelines in the delivery of bad news, their own fears, a general lack of supervisory support and time constraints form barriers to their effective interaction with patients. PMID- 11260441 TI - Training physicians in communication skills with adolescents using teenage actors as simulated patients. AB - Role-play exercises with simulated patients may serve the purpose of training professionals to develop appropriate communication skills with adolescents. Authentic adolescent responses toward the physicians may be achieved by actors who themselves are in their teenage years. We describe our experience in continuing medical education programmes for primary care physicians aimed at improving their skills in communicating with adolescents, using simulation methodology with teenage actors. Eight 16-17-year-old actors from the drama department of a high school for the arts were trained to simulate 20 cases with characteristic adolescent medical problems, as well as confidentiality issues and home and school problems. The actors performed in front of large groups of 20-30 paediatricians, family practitioners, or gynaecologists in continuing medical education. Diagnostic issues as well as therapeutic and management approaches were discussed, while the actors provided feedback to the trainees about their understanding and their feeling regarding the issues raised during the exercises. Normally, smaller learning groups are more suitable for such training purposes; nevertheless the participants could appreciate learning the principles of careful listening, a non-judgmental approach and assuring confidentiality. A collaboration of medical schools and postgraduate programmes with high schools which have drama departments may be fruitful in the teaching of adolescent medicine with special emphasis on communication skills with teenagers. PMID- 11260442 TI - Interactional skills of students from traditional and non-traditional medical schools before and after alcohol education. AB - OBJECTIVE: To compare alcohol-related intervention and general interactional skills performance of medical students from a traditional (Sydney) and a non traditional (Newcastle) medical school, before and after participation in an alcohol education programme about brief intervention. DESIGN: In two controlled trials, students received either a didactic alcohol education programme or didactic input plus skills-based training. Prior to and after training, all students completed videotaped interviews with simulated patients. SETTING: The Faculties of Medicine at the University of Newcastle and the University of Sydney, Australia. SUBJECTS: Fifth-year medical students (n=154). RESULTS: Both alcohol-related intervention and general interactional skills scores of the Newcastle students were significantly higher than those of the Sydney students at pre-test but not after training. Although alcohol-related interactional skills scores improved after training at both universities, they did not reach a satisfactory level. The educational approach used had no effect on post-test scores at either university. CONCLUSIONS: Significant baseline differences in interactional skills scores favouring non-traditional over traditional students were no longer evident after both groups had been involved in an alcohol education programme. Further research is required to develop more effective alcohol intervention training methods. PMID- 11260443 TI - Preliminary evaluation of "interpreter" role plays in teaching communication skills to medical undergraduates. AB - RATIONALE AND OBJECTIVES: Multiculturalism presents linguistic obstacles to health care provision. We explored the early introduction of "interpreter" role play exercises in teaching medical undergraduates communication skills. The interpreter role creates a natural barrier in communication providing an active prompt for recognizing learning needs in this area. METHODS: Bilingual Cantonese first-year medical students (n=160) were randomly allocated to either "Observer" or "Interpreter" role plays at a small-group introductory communication skills workshop using a quasi experimental design, counterbalanced across tutors. Students assessed their own skill competence before and, together with their perceptions of the different role plays' effectiveness, again after the workshop, using an anonymous 16 item Likert-type scale, analysed using ANOVA and MANOVA. RESULTS: Students' assessments of their skills improved significantly following the workshop (F=73.19 [1,156], P=0.0009). Students in the observer group reported greater changes in their scores following the workshop than did students in the interpreter group (F=4.84 [1,156], P=0.029), largely due to improvement in perceived skill (F=4.38 [1,156], P=0.038) rather than perceived programme effectiveness (F=3.13 [1,156], P > 0.05). Subsequent MANOVA indicated no main effect of observer/interpreter conditions, indicating these differences could be attributed to chance alone (F=1.41 [16 141], P > 0.05). CONCLUSION: The workshop positively influenced students' perceived communication skills, but the "Interpreter" role was less effective than the "Observer" role in achieving this. Future studies should examine whether interpreter role plays introduced later in the medical programme are beneficial. PMID- 11260444 TI - Teaching consultation skills: a survey of general practice trainers. AB - BACKGROUND: Consultation skills are vitally important in general practice (GP), and now form part of the summative assessment of GP registrars in the UK. GP trainers need to be skilled in teaching consultation skills, and also need the time and resources to ensure that their registrars are competent in consultation skills. AIMS: To describe the teaching methods used by GP trainers in one deanery, the frequency of teaching of consultation skills, the problems encountered and the training that GP trainers have themselves received both in consultation skills and how to teach them. METHOD: Postal questionnaire survey of all the 164 trainers in the Yorkshire Deanery. RESULTS: Replies were received from 129 trainers (response rate 79%) of which 123 could be analysed. Of these trainers, 45 (37%) trainers taught consultation skills fewer than five times a year, 45 (37%) five to 10 times, and 14 (11%) more than 10 times a year. A total of 24 trainers reported problems with teaching consultation skills, most commonly lack of time, technical difficulties, and unreceptive registrars, and 97 (79%) trainers had had some postgraduate training in consultation skills with 112 (91%) reporting some form of teacher training. CONCLUSION: There is considerable variation in the reported frequency of teaching consultation skills, the models used, and the preparation of trainers for teaching, despite a systematic approach to teacher training in the Yorkshire Deanery. PMID- 11260445 TI - Assessing the development of communication skills in undergraduate medical students. AB - CONTEXT: The teaching of clinical communication skills' teaching has become an important part of medical school curricula. Many undergraduate medical courses include communication skills training at various points in their curriculum. Very few reports have been published on the development of communication skills over the duration of a medical undergraduate training. AIMS: To determine the change in communication skills between early and mid-stages of the students' 5-year curriculum, and to investigate the predictive and theoretical significance of knowledge and understanding of communication skills in relation to observed performance. PARTICIPANTS: Students entering as the first cohort to the new medical curriculum at Liverpool Medical School (n=207). Nine students withdrew leaving 198 students who completed two summative assessments in June 1997 (level 1) and November 1998 (level 2). STATISTICAL ANALYSIS: Repeated measures multivariate ANOVAS were applied to the main study data to detect any change in performance between levels 1 and 2. RESULTS AND CONCLUSIONS: An improvement in communication skills was found in medical students over 17 months of their undergraduate teaching: that is from the level 1 to the level 2 assessment. Knowledge and understanding of communication skills at initial assessment did not show the predicted association with performance at level 2. PMID- 11260446 TI - The development and evaluation of a programme to teach cultural diversity to medical undergraduate students. AB - This paper describes the design (of process and content), implementation and evaluation of a component of the Human Diversity Module developed to teach cultural diversity to undergraduate medical students. The objectives of the teaching were to enable students to gain factual and practical information about other cultures and also for them to examine their own attitudes. METHOD: Students completed a questionnaire, designed in a previous study, at two stages; the first before the component on cultural diversity had been delivered and the second after the sessions on cultural diversity. The time interval between stages 1 and 2 was 1 week. The cultural diversity component was developed using a range of sources. RESULTS: Out of 181 students, 140 (77.3%) completed the questionnaire at both stages. There were a number of statistically significant findings, which indicate that the teaching enabled the session objectives to be successfully met. The findings include statistically significant changes that reflect more "positive" attitudes about cultures coming together and about specific cultures. CONCLUSION: The study indicates that attitudes changed over the period of teaching. There is, however, scope for further development of measures to enable attitudinal shifts to be evaluated. PMID- 11260447 TI - The first sunrise: an experience of cultural immersion and community health needs assessment by undergraduate medical students in New Zealand. AB - CONTEXT: Cultural factors in health and illness, and an awareness of community health needs analysis, are important issues for medical education. Both have received relatively little recognition in the medical education literature. This paper describes the development of an educational attachment to remote predominantly Maori rural communities in New Zealand. The twin purposes of the programme were to encourage students to adopt broad public health approaches in assessing the health needs of defined communities, and to increase their awareness of the importance of cultural issues. METHODS: During a one week attachment, 51 students from the Wellington School of Medicine were hosted in six small communities in the East Cape region of New Zealand. Students gained an insight into the health needs of the communities and were encouraged to challenge their own attitudes, assumptions and thinking regarding the determinants of health and the importance of cultural factors in health and illness. The programme included both health needs assessment and cultural immersion. Students made visits with primary health care professionals and were also introduced to Maori history and cultural protocol, and participated in diverse activities ranging from the preparation of traditional medicines to performing their own songs in concert. CONCLUSIONS: The students evaluated the course extremely highly. Attachments of this sort provide an opportunity for students to appreciate how cultural values have an impact on health care, and how they also make the teaching and learning of topics such as community health needs analysis an enjoyable and dynamic experience. PMID- 11260448 TI - " 'Just think of TB and Asians', that's all I ever hear": medical learners' views about training to work in an ethnically diverse society. AB - The case for being able to respond effectively to cultural and ethnic diversity in health care is attracting increasing debate in medical education. However research exploring the perspectives of learners is lacking. AIMS: We sought medical learners' perceptions and their perceived training needs in relation to cultural and ethnic diversity in health care. METHODS: A series of nine focus group interviews was conducted with 55 medical learners, including undergraduate students in a UK medical school and a group of postgraduate general practitioners in training. Interview data were analysed using qualitative methods. RESULTS: Participants had a broad but superficial awareness of multicultural issues. This focused upon "difference" with students emphasizing their need to acquire knowledge of different beliefs and practices. Current teaching was perceived as inadequate and limited largely to ethnic patterns of disease. Most felt a need for greater training. They regarded development of particular communication skills, such as working with interpreters as helpful. Beyond avoiding stereotyping, learners rarely identified reflecting upon their attitudes or the issue of racism as important. Students anticipated a range of potential problems for further training, but sought learning that was relevant, practically oriented and stimulating. CONCLUSIONS: The study points to learners' experience of inadequate training but suggests a willingness to learn more. The possible predominance of a "difference" perspective might drive a narrow focus upon learning cultural knowledge at the expense of promoting a balance with self reflection upon attitudes and developing generic skills. Educators might heed learners' views about how they should be taught successfully. PMID- 11260449 TI - Cross-cultural communication in medicine: questions for educators. AB - Most research into medical communication has had a western setting. It has been undertaken by western researchers and been influential in shaping communication skills curricula. However we know much less about what communication is effective under other circumstances. This article highlights gaps in our knowledge from research in this field, and poses attendant questions for debate by medical educators. We consider the following key aspects of debate on cross-cultural work. (i) To what extent can our understanding of general principles in other cultures be summarized and presented for teaching in a way which does not descend into caricature? Alternatively, can features of other cultures be presented in ways which do not descend into particularity? (ii) Can such paradigms as "patient centredness" be transferred from culture to culture? Should they be presented across cultures as features of "good" consultations? (iii) What use can be made of the role of interpreters for teaching purposes? What importance does it have to the educator that a doctor may not be a native speaker of the majority language of the culture in which s/he is operating? (iv) Although the language of illness, and particularly metaphors associated with illness, are studied in other cultures, the way in which illness is metaphorized in British English is seldom discussed. What can educators learn and teach from a study of such matters? (v) What are the implications for communication skills teachers of the need to present materials within a culturally diverse environment? PMID- 11260450 TI - Wrestling with ethnic diversity: toward empowering health educators. AB - The importance of training health professionals for work in an ethnically diverse society is increasingly recognized. However, health educators may lack confidence or experience in delivering such teaching, contributing to a self-perpetuating inertia. OBJECTIVES: To identify current experience and challenges perceived by educators of different health professionals, and to facilitate and debate the development of teaching in this field. METHODS: Educators (n=61) from 42 different organizations, participated in facilitated workshops in three different UK settings. They included clinician teachers of medical undergraduates and postgraduates, and educators of nurses, primary care and hospital physicians, physiotherapists, occupational therapists and paramedical staff. Opportunities were provided for educators to discuss experiences; to participate as "learners" in examples of interactive training exercises; to anticipate challenges they might encounter in developing and providing training themselves; and to discuss ways of negotiating them. Qualitative data generated from the workshops were analysed for common themes. RESULTS: Participants had received little relevant training themselves. For many, the workshops provided a first formal opportunity to consider their own responses to ethnic diversity in health care. Current provision of such training in their institutions was limited. Educators lacked specific training to facilitate the learning of others in this field. They wrestled with a wide range of issues: from critical dilemmas about the philosophy of teaching, through to the practicalities and personal challenges of face to face teaching. Strategies to address these were generated that may merit consideration. CONCLUSION: Educators will need help to overcome their uncertainty in approaching this topic and be empowered to develop training. Developing teachers' own awareness and skills, followed by appropriate support, are likely to be prerequisites for successful training. PMID- 11260451 TI - Identifying characteristics that students, interns and residents look for in their role models. AB - OBJECTIVE: To identify characteristics which students, interns and residents look for in their role models. METHODS: A 45-item self-administered questionnaire was sent to a sample (n=96, response rate 80%) consisting of three groups: (1) students in years 3-6 of the medical curriculum (n=66); (2) interns (n=17) and (3) residents (n=13). The questionnaire contained characteristics that participants might use to describe excellent role models, grouped under five general headings: personality, clinical, research and teaching skills, and community service. Other characteristics mentioned by study subjects were qualitatively analysed using content analysis. RESULTS: Personality and teaching and clinical skills were ranked as the top three factors, and research skills and community service as the least important factors by 79 (82%) respondents. Qualitative analysis of characteristics described by respondents for their role models yielded 21 characteristics. These were clustered into three main themes: role models as teacher, physician and person. The most frequently mentioned characteristics were personal characteristics such as positive, respectful attitudes toward patients and their families, and staff and colleagues; honesty; politeness; enthusiasm; competence, and knowledge. Females rated nine personal characteristics significantly higher than males (P < 0.05). Interns and residents valued teaching enthusiasm and competence significantly more than students (P=0.01). Role models had a strong influence on the specialty choice of 53 (55%) respondents. CONCLUSION: Knowing the characteristics of excellent role models should help medical educators to formulate strategies to recruit, retain and develop them. Increasing exposure of a variety of excellent role models to aspiring medical practitioners should be encouraged. PMID- 11260452 TI - Community family medicine teachers' perceptions of their teaching role. AB - OBJECTIVES: Our study explored community preceptors' perceptions of their teaching role, to better understand effective ambulatory and community-based teaching. METHODS: Bandura's social cognitive theory and Schon's notion of reflective practice guided conceptual development of an interview exploring preceptors' views of their role, teaching goals, teaching techniques, student assessment practices, factors affecting teaching and learning, and balance of patient and student needs. Preceptors reflected also on a significant personal teaching experience. A total of 17 highly student-rated preceptors participated. A trained interviewer conducted each interview; all were transcribed and subjected to content analysis. RESULTS: Preceptors (male, 14; female, 3) described learner-centred approaches, setting goals jointly with the student. Demonstration, guided practice, observation and feedback were integral to the experience. Preceptors saw student comfort in the environment as key to effective learning; they attempted to maximize students' learning and breadth of experience. They wanted students to understand content, "know-how" and "being a family physician". Patients remained the primary responsibility, but learners' needs were viewed as compatible with that responsibility. Many preceptors perceived a professional responsibility as "role models". CONCLUSIONS: Preceptors recognized the dynamic environment in which they taught students, and they described strategies which demonstrated how they adapted their teaching to meet the needs of the learner in that environment. These teachers combined learner centred approaches with sound educational practices, broad learning experiences, attention to student learning and concern for development of professional expertise and judgement. These findings may assist faculty development in family medicine, and other disciplines, in providing effective ambulatory care teaching. PMID- 11260453 TI - Refuting patients' obligations to clinical training: a critical analysis of the arguments for an obligation of patients to participate in the clinical education of medical students. AB - CONTEXT: The clinical teaching of medical students is essential to the continuation of medicine, but it has a major impact on the patient's health care and autonomy. Some people believe that there is a moral obligation for patients to participate in this training. Such an obligation, real or perceived, may endanger patients' autonomy. OBJECTIVES: The author makes a critical analysis of the main arguments he encounters supporting such an obligation. These arguments are: (1) the furthering of medical education; (2) compensation when uninsured or unable to pay; (3) an equitable return for the care received in a teaching hospital, and (4) fulfilment of a student's need for (and some say right to) clinical training. METHODS: Related literature is reviewed in search of evidence and/or support for such arguments. CONCLUSIONS: The review reveals that these arguments either cannot be verified or do not necessarily place any obligations on the patient. It is argued that, while a medical student may have a right to clinical education, the obligation to fulfil this right rests with the medical university and not on the patients of its teaching hospitals. SOLUTIONS: Several proposals are made about how to satisfy this need without infringing on the patient's right to refuse participation, explaining the patient's rights and role in clinical teaching, and the use of standardized patients where necessary. PMID- 11260456 TI - Traditional medical education and the new path--they are not mutually exclusive. PMID- 11260454 TI - Impact of a new course on students' potential behaviour on encountering ethical dilemmas. AB - OBJECTIVE: To evaluate the effectiveness of small-group ethics teaching in an integrated medical curriculum. DESIGN: A quasi-experimental, pre- and post-test, non-equivalent control group design. SETTING: University of Glasgow Medical School. SUBJECTS: 111 first-year students from Glasgow University's new learner centred medical curriculum, with a control group of 51 students from the last year of the traditional curriculum. MAIN OUTCOME MEASURE: Student answers consistent with consensus professional judgement on the ethical dilemmas posed by the vignettes of the Ethics and Health Care Survey Instrument. RESULTS: There was a significantly greater increase in the number of post-test consensus answers in the experimental group (P=0.0048): the odds ratio for obtaining the post-test consensus answer in the experimental group compared with the control group was 1.73 (95% confidence interval 1.28-2.33). Significant movement towards consensus occurred in the areas of autonomy, confidentiality and consent. Among controls there was a significant move away from consensus in the area of "whistle blowing" on colleagues (P=0.017). CONCLUSION: Small-group ethics teaching, in an integrated medical curriculum, had a positive impact on the first-year students' potential ethical behaviour. It was more effective than a lecture and a large group seminar-based course in developing students' normative identification with the profession of medicine. PMID- 11260458 TI - Becoming professional. PMID- 11260457 TI - What the GP pre-registration house officer saw--a personal view. PMID- 11260459 TI - USMLE-style exam. PMID- 11260460 TI - Dispensable nature of phosphatidylglycerol in Escherichia coli: dual roles of anionic phospholipids. AB - The major anionic phospholipids of Escherichia coli, phosphatidylglycerol (PG) and cardiolipin (CL), have been considered to be indispensable for essential cellular functions, such as the initiation of DNA replication and translocation of proteins across the cytoplasmic membrane. However, we successfully constructed a null pgsA mutant of E. coli that had undetectable levels of PG and CL if the major outer membrane lipoprotein was deficient, clearly indicating that these anionic phospholipids are not indispensable. In the null mutant, we observed the accumulation of phosphatidic acid, an acidic biosynthetic precursor. This suggests a functionally substitutable nature of these anionic phospholipids and allows us to formulate a dual role model for the physiological roles of the anionic phospholipids in E. coli. The anionic phospholipids may play dual roles in E. coli as (i) substrates for head group-specific enzyme reactions, albeit the viability of null PG mutants indicates that the products of head group-specific reactions are not essential; and (ii) those that are replaceable, partly or entirely, by other phospholipids bearing net negative charges, because of their rather loose head group specificity. These two aspects of the physiological roles of anionic phospholipids are discussed with special reference to the phospholipids of other bacteria and eukaryotic organelles. PMID- 11260461 TI - The phenylacetyl-CoA catabolon: a complex catabolic unit with broad biotechnological applications. AB - The term catabolon was introduced to define a complex functional unit integrated by different catabolic pathways, which are, or could be, co-ordinately regulated, and that catalyses the transformation of structurally related compounds into a common catabolite. The phenylacetyl-CoA catabolon encompasses all the routes involved in the transformation of styrene, 2-phenylethylamine, trans-styrylacetic acid, phenylacetaldehyde, phenylacetic acid, phenylacetyl amides, phenylacetyl esters and n-phenylalkanoic acids containing an even number of carbon atoms, into phenylacetyl-CoA. This common intermediate is subsequently catabolized through a route of convergence, the phenylacetyl-CoA catabolon core, into general metabolites. The genetic organization of this central route, the biochemical significance of the whole functional unit and its broad biotechnological applications are discussed. PMID- 11260462 TI - An essential dimeric membrane protein of trypanosome glycosomes. AB - Kinetoplastid parasites compartmentalize the first seven enzymes of glycolysis in a peroxisome-like microbody, the glycosome. Genes encoding the most abundant protein of the glycosomal membrane, GIM5, have been cloned and the protein characterized. Two genes, GIM5A and GIM5B, encode 26 kDa proteins. Although many microbody membrane proteins are conserved in evolution, the only homologues of GIM5 in the available databases are from the closely related kinetoplastids Trypanosoma cruzi and Leishmania. The N- and C-termini are conserved between the two genes, and between species, and are oriented towards the cytosol. They are separated by a short loop that is located between two transmembrane domains and shows almost no sequence conservation. This suggests that the N- and C-terminal domains are more important for function. GIM5 forms dimers in vivo. Overexpression of GIM5B inhibits growth, whereas depletion of GIM5 to below 10% of wild-type levels is very rapidly lethal. This novel organellar membrane protein is therefore essential for bloodstream trypanosome survival. PMID- 11260463 TI - The multicellular morphotypes of Salmonella typhimurium and Escherichia coli produce cellulose as the second component of the extracellular matrix. AB - Production of cellulose has been thought to be restricted to a few bacterial species such as the model organism Acetobacter xylinus. We show by enzymatic analysis and mass spectrometry that, besides thin aggregative fimbriae, the second component of the extracellular matrix of the multicellular morphotype (rdar) of Salmonella typhimurium and Escherichia coli is cellulose. The bcsA, bcsB, bcsZ and bcsC genes responsible for cellulose biosynthesis are not regulated by AgfD, the positive transcriptional regulator of the rdar morphotype. Transcription of the bcs genes was not co-expressed with the rdar morphotype under any of the environmental conditions examined. However, cellulose biosynthesis was turned on by the sole expression of adrA, a gene encoding a putative transmembrane protein regulated by agfD, indicating a novel pathway for the activation of cellulose synthesis. The co-expression of cellulose and thin aggregative fimbriae leads to the formation of a highly hydrophobic network with tightly packed cells aligned in parallel in a rigid matrix. As the production of cellulose would now appear to be a property widely distributed among bacteria, the function of the cellulose polymer in bacteria will have to be considered in a new light. PMID- 11260464 TI - The Salmonella spvB virulence gene encodes an enzyme that ADP-ribosylates actin and destabilizes the cytoskeleton of eukaryotic cells. AB - ADP-ribosylating enzymes, such as cholera and diphtheria toxins, are key virulence factors for a variety of extracellular bacterial pathogens but have not been implicated previously during intracellular pathogenesis. Salmonella strains are capable of invading epithelial cells and localizing in macrophages during infection. The spvB virulence gene of Salmonella is required for human macrophage cytotoxicity in vitro and for enhancing intracellular bacterial proliferation during infection. Here, we present evidence that spvB encodes an ADP-ribosylating enzyme that uses actin as a substrate and depolymerizes actin filaments when expressed in CHO cells. Furthermore, site-directed mutagenesis demonstrates that the ADP-ribosylating activity of SpvB is essential for Salmonella virulence in mice. As spvB is expressed by Salmonella strains after invasion of epithelial cells or phagocytosis by macrophages, these results suggest that SpvB functions as an intracellular ADP-ribosylating toxin critical for the pathogenesis of Salmonella infections. PMID- 11260465 TI - Chromosomal organization governs the timing of cell type-specific gene expression required for spore formation in Bacillus subtilis. AB - During the early stages of spore formation in Bacillus subtilis, asymmetric division precedes chromosome segregation, such that the forespore transiently contains only about one-third of the genetic material surrounding the origin of replication. Shortly after septum formation, the transcription factor sigmaF initiates forespore-specific gene expression that is essential for the proteolytic activation of pro-sigmaE in the neighbouring mother cell. Moving the sigmaF-dependent spoIIR gene from its original origin-proximal position to an ectopic origin-distal site caused a delay in spoIIR transcription, as well as delays and reductions in the proteolytic activation of pro-sigmaE and sigmaE directed gene expression. These defects correlated with the accumulation of disporic sporangia, thus reducing sporulation efficiency in a manner that depended upon the distance that spoIIR had been moved from the origin-proximal third of the chromosome. A significant proportion of disporic sporangia exhibited sigmaE activity in their central compartment, indicating that delays and reductions in sigmaE activation can lead to the formation of a second septum at the opposite pole. These observations support a model in which chromosomal spoIIR position temporally regulates sigmaE activation, thereby allowing for the rapid establishment of mother cell-specific gene expression that is essential for efficient spore formation. The implications of these findings for cell type specific gene expression during the early stages of spore formation in B. subtilis are discussed. PMID- 11260466 TI - RcoA has pleiotropic effects on Aspergillus nidulans cellular development. AB - Aspergillus nidulans rcoA encodes a member of the WD repeat family of proteins. The RcoA protein shares sequence similarity with other members of this protein family, including the Saccharomyces cerevisiae Tup1p and Neurospora crassa RCO1. Tup1p is involved in negative regulation of an array of functions including carbon catabolite repression. RCO1 functions in regulating pleiotropic developmental processes, but not carbon catabolite repression. In A. nidulans, deletion of rcoA (DeltarcoA), a recessive mutation, resulted in gross defects in vegetative growth, asexual spore production and sterigmatocystin (ST) biosynthesis. Expression of the asexual and ST pathway-specific regulatory genes, brlA and aflR, respectively, but not the signal transduction genes (i.e. flbA, fluG or fadA) regulating brlA and aflR expression was delayed (brlA) or eliminated (aflR) in a DeltarcoA strain. Overexpression of aflR in a DeltarcoA strain could not rescue normal expression of downstream targets of AflR. CreA dependent carbon catabolite repression of starch and ethanol utilization was only weakly affected in a DeltarcoA strain. The strong role of RcoA in development, vegetative growth and ST production, compared with a relatively weak role in carbon catabolite repression, is similar to the role of RCO1 in N. crassa. PMID- 11260467 TI - Sugar transport in Sulfolobus solfataricus is mediated by two families of binding protein-dependent ABC transporters. AB - The extreme thermoacidophilic archaeon Sulfolobus solfataricus grows optimally at 80 degrees C and pH 3 and uses a variety of sugars as sole carbon and energy source. Glucose transport in this organism is mediated by a high-affinity binding protein-dependent ATP-binding cassette (ABC) transporter. Sugar-binding studies revealed the presence of four additional membrane-bound binding proteins for arabinose, cellobiose, maltose and trehalose. These glycosylated binding proteins are subunits of ABC transporters that fall into two distinct groups: (i) monosaccharide transporters that are homologous to the sugar transport family containing a single ATPase and a periplasmic-binding protein that is processed at an unusual site at its amino-terminus; (ii) di- and oligosaccharide transporters, which are homologous to the family of oligo/dipeptide transporters that contain two different ATPases, and a binding protein that is synthesized with a typical bacterial signal sequence. The latter family has not been implicated in sugar transport before. These data indicate that binding protein-dependent transport is the predominant mechanism of transport for sugars in S. solfataricus. PMID- 11260468 TI - The essential role of the promoter-proximal subunit of CAP in pap phase variation: Lrp- and helical phase-dependent activation of papBA transcription by CAP from -215. AB - Catabolite gene activator protein (CAP) is essential for the expression of Pap pili by uropathogenic Escherichia coli. Both in vitro and in vivo analyses indicate that binding of cAMP-CAP centred at 215.5 bp upstream of the papBA promoter is essential for activation of transcription. CAP-dependent activation of papBA requires binding of the leucine-responsive regulatory protein (Lrp) at binding sites that extend from -180 to -149 relative to the start site of papBA. Our data indicate that CAP and Lrp bind independently to their respective pap DNA sites. Activation of papBA transcription was eliminated by mutations in the activating region 1 (AR1) of CAP, but not in the AR2 region, similar to class I CAP-dependent promoters. Also, like class I promoters, the C-terminal domain of the alpha-subunit of RNA polymerase appears to play a role in transcription activation. Moreover, phase variation is strictly dependent upon the helical phase of the CAP DNA binding site with respect to the papBA transcription start site. Using an 'oriented heterodimer' approach with wild-type and AR1 mutant CAPs, it was shown that the AR1 region of the CAP subunit proximal to papBA is required for stimulation of papBA transcription, whereas AR1 of the promoter distal subunit is not. Previously, CAP was hypothesized to activate pap transcription indirectly by disrupting repression mediated by H-NS. The results presented here show that AR1 of the promoter-proximal CAP subunit was required for papBA transcription even in the absence of the histone-like protein H-NS. These results show that the promoter-proximal subunit of CAP, bound 215.5 bp upstream of the papBA transcription start site, plays an active role in stimulating papBA transcription, possibly by interacting with the C-terminal domain of the alpha-subunit of RNA polymerase. PMID- 11260469 TI - Hsf1p and Msn2/4p cooperate in the expression of Saccharomyces cerevisiae genes HSP26 and HSP104 in a gene- and stress type-dependent manner. AB - Saccharomyces cerevisiae possesses several transcription factors involved in the transcriptional activation of stress-induced genes. Among them, the heat shock factor (Hsf1p) and the zinc finger proteins of the general stress response (Msn2p and Msn4p) have been shown to play a major role in stress protection. Some heat shock protein (HSP) genes contain both heat shock elements (HSEs) and stress response elements (STREs), suggesting the involvement of both transcription factors in their regulation. Analysis of the stress-induced expression of two of these genes, HSP26 and HSP104, reveals that the contribution of Hsf1p and Msn2/4p is different depending on the gene and the stress condition. PMID- 11260470 TI - Identification of a non-haem catalase in Salmonella and its regulation by RpoS (sigmaS). AB - We report the identification and functional analysis of katN, a gene encoding a non-haem catalase of Salmonella enterica serotype Typhimurium. katN, which is not present in Escherichia coli, is located between the yciGFE and yciD E. coli homologues in the Salmonella genome. Its predicted protein product has a molecular weight of 31 826 Da and is similar to the Mn-catalases of Lactobacillus plantarum and Thermus spp. Its product, KatN, was visualized as a 37 kDa protein in E. coli maxicells. A KatN recombinant protein, containing six histidine residues at its C-terminus, was purified, and its catalase activity was observed on a non-denaturing polyacrylamide gel. KatN was also visualized by catalase activity gel staining of bacterial cell extracts. Its expression was shown to be regulated by growth phase and rpoS. Northern blotting indicated that kat forms an operon with the upstream yciGFE genes. A putative rpoS-regulated promoter was identified upstream of yciG. Southern blotting revealed that katN is conserved within Salmonella serovars. katN homologues were found in Pseudomonas aeruginosa, Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter cloacae and Serratia marcescens. A katN mutation did not appear to affect the hydrogen peroxide (H2O2) response of Salmonella. However, the expression of katN increased the H2O2 resistance of unadapted cells in the exponential phase and of rpoS mutants in stationary phase. Thus, KatN may contribute to hydrogen peroxide resistance in Salmonella in certain environmental conditions. PMID- 11260471 TI - Bile-induced 'pili' in Campylobacter jejuni are bacteria-independent artifacts of the culture medium. AB - In 1996, it was reported that the enteric pathogen Campylobacter jejuni produces pilus-like appendages in response to bile salts such as deoxycholate (DOC), and that the formation of these appendages requires the putative peptidase PspA. Pili were known to be important virulence determinants in other pathogenic bacteria but had never before been observed for C. jejuni. We report here that these appendages are not pili, but are instead a bacteria-independent morphological artifact of the growth medium. Furthermore, the pspA gene is not required for their formation. Broth cultures containing a threshold concentration of DOC inoculated with no bacteria produced identical abundant, fibrous, pilus-like structures as those cultures that had been inoculated with C. jejuni. These fibres were also found in growth media from DOC-containing pspA:CmR mutant cultures. These results are consistent with the absence of candidate pilin monomers in protein gel analyses as well as the dearth of pilin-like genes and pilus formation gene clusters in the C. jejuni genome. PMID- 11260472 TI - RNase II levels change according to the growth conditions: characterization of gmr, a new Escherichia coli gene involved in the modulation of RNase II. AB - In Escherichia coli, ribonucleases are effectors that rapidly modulate the levels of mRNAs for adaptation to a changing environment. Factors involved in the regulation of these ribonucleases can be relevant for mRNA stability. RNase II is one of the main ribonucleases responsible for exonucleolytic activity in E. coli extracts. We have identified and characterized a new E. coli gene, which was named gmr (gene modulating RNase II). The results demonstrate that a deletion of gmr can be associated with changes in RNase II levels and activity. Western analysis and exoribonuclease activity assays showed a threefold increase in RNase II in the gmr deletion strain. Gmr does not affect RNase II mRNA, but modulates RNase II at the level of protein stability. RNase II protein turnover is slower in the gmr deletion strain. We also show that RNase II levels change in different media, and that this regulation is abolished in a strain lacking gmr. The data presented here show that the regulation of ribonucleolytic activity can depend on growth conditions, and this regulation can be mediated by factors that are not RNases. PMID- 11260473 TI - How does oxygen inhibit central metabolism in the obligate anaerobe Bacteroides thetaiotaomicron. AB - The molecular basis of obligate anaerobiosis is not well established. Bacteroides thetaiotaomicron is an opportunistic pathogen that cannot grow in fully aerobic habitats. Because microbial niches reflect features of energy-producing strategies, we suspected that aeration would interfere with its central metabolism. In anaerobic medium, this bacterium fermented carbohydrates to a mixture of succinate, propionate and acetate. When cultures were exposed to air, the formation of succinate and propionate ceased abruptly. In vitro analysis demonstrated that the fumarase of the succinate-propionate pathway contains an iron-sulphur cluster that is sensitive to superoxide. In vivo, fumarase activity fell to < 5% when cells were aerated; virtually all activity was recovered after extracts were chemically treated to rebuild iron-sulphur clusters. Aeration minimally affected the remainder of this pathway. However, aeration reduced pyruvate:ferredoxin oxidoreductase (PFOR), the first enzyme in the acetate fermentation branch, to 3% of its anaerobic activity. This cluster-containing enzyme was damaged in vitro by molecular oxygen but not by superoxide. Thus, aerobic growth is precluded by the vulnerability of these iron-sulphur cluster enzymes to oxidation. Importantly, both enzymes were maintained in a stable, inactive form for long periods in aerobic cells; they were then rapidly repaired when the bacterium was returned to anaerobic medium. This result explains how this pathogen can easily recover from occasional exposure to oxygen. PMID- 11260474 TI - CspD, a novel DNA replication inhibitor induced during the stationary phase in Escherichia coli. AB - CspD is a stationary phase-induced, stress response protein in the CspA family of Escherichia coli. Here, we demonstrate that overproduction of CspD is lethal, with the cells displaying a morphology typical of cells with impaired DNA replication. CspD consists mainly of beta-strands, and the purified protein exists exclusively as a dimer and binds to single-stranded (ss)DNA and RNA in a dose-dependent manner without apparent sequence specificity. CsdD effectively inhibits both the initiation and the elongation steps of minichromosome replication in vitro. Electron microscopic studies revealed that CspD tightly packs ssDNA, resulting in structures distinctly different from those of SSB coated DNA. We propose that CspD dimers, with two independent beta-sheets interacting with ssDNA, function as a novel inhibitor of DNA replication and play a regulatory role in chromosomal replication in nutrient-depleted cells. PMID- 11260475 TI - An adenosyl-cobalamin (coenzyme-B12)-repressed translational enhancer in the cob mRNA of Salmonella typhimurium. AB - Expression of the cobalamin (Cbl) biosynthetic cob operon in Salmonella typhimurium is repressed by the end-product. This regulation is conferred mainly at the translational level and involves a cobalamin-induced folding of an RNA hairpin that sequesters the ribosomal binding site (RBS) of the cob mRNA and prevents translation initiation. A combined structural and mutational analysis shows that a cis-acting translational enhancer (TE) element, located 83 nucleotides upstream of the Shine-Dalgarno sequence in the 5'-untranslated region (5'-UTR) of the cob mRNA, is required to unfold the inhibitory RBS hairpin in the absence of cobalamin. The TE element, which consists of 5 nucleotides, is proposed to confer its enhancer function in the absence of cobalamin by interacting with nucleotides in the stem of the RBS hairpin. This interaction destabilizes the RNA hairpin and allows ribosome binding. In the presence of cobalamin, the enhancer function is inhibited. As a result, the RBS hairpin forms and prevents translation initiation. Several additional RNA hairpins in the 5' UTR were also identified and are suggested to be important for repression. The above data suggest that normal cobalamin repression of the cob operon requires that the 5'-UTR has a defined secondary and tertiary structure. PMID- 11260476 TI - The novel sigma54- and sigma28-dependent flagellar gene transcription hierarchy of Vibrio cholerae. AB - The human pathogen Vibrio cholerae is a highly motile organism by virtue of a polar flagellum. Flagellar transcriptional regulatory factors have been demonstrated to contribute to V. cholerae virulence, but the role these factors play in the transcription hierarchy controlling flagellar synthesis has been unclear. The flagellar genes revealed by the V. cholerae genome sequence are located in three large clusters, with the exception of the motor genes, which are found in three additional locations. It had previously been demonstrated that the alternative sigma factor sigma54 and the sigma54-dependent activators FlrA and FlrC are necessary for flagellar synthesis. The V. cholerae genome sequence revealed the presence of a fliA gene, which is predicted to encode the alternative flagellar sigma factor sigma28. A V. cholerae DeltafliA mutant strain is non-motile, and synthesizes a truncated flagellum. Vibrio cholerae FliA complements both V. cholerae and Salmonella typhimurium fliA mutants for motility, consistent with its function as an alternative flagellar sigma factor. Analysis of lacZ transcriptional fusions of the V. cholerae flagellar promoters in both V. cholerae and S. typhimurium identified sigma28-, sigma54-, FlrA- and FlrC-dependent promoters, as well as promoters that were independent of all these factors. Our results support a model of V. cholerae flagellar gene transcription as a novel hierarchy composed of four classes of genes. Class I is composed solely of the gene encoding the sigma54-dependent activator FlrA, which along with the sigma54-holoenzyme form of RNA polymerase activates expression of Class II genes. These genes include structural components of the MS ring, switch and export apparatus, as well as the genes encoding both FliA and FlrC. FlrC, along with sigma54-holoenzyme, activates expression of Class III genes, which include basal body, hook and filament genes. Finally, sigma28-holoenzyme activates expression of Class IV genes, which include additional filament genes as well as motor genes. Thus, this novel V. cholerae flagellar hierarchy has incorporated elements from both the sigma54-dependent Caulobacter crescentus polar flagellar hierarchy and the sigma28-dependent S. typhimurium peritrichous flagellar hierarchy. PMID- 11260477 TI - Identification of the teichoic acid phosphorylcholine esterase in Streptococcus pneumoniae. AB - Streptococcus pneumoniae is a major human pathogen and many interactions of this bacterium with its host appear to be mediated, directly or indirectly, by components of the bacterial cell wall, specifically by the phosphorylcholine residues which serve as anchors for surface-located choline-binding proteins and are also recognized by components of the host response, such as the human C reactive protein, a class of myeloma proteins and PAF receptors. In the present study, we describe the identification of the pneumococcal pce gene encoding for a teichoic acid phosphorylcholine esterase (Pce), an enzymatic activity capable of removing phosphorylcholine residues from the cell wall teichoic acid and lipoteichoic acid. Pce carries an N-terminal signal sequence, contains a C terminal choline-binding domain with 10 homologous repeating units similar to those found in other pneumococcal surface proteins, and the catalytic (phosphorylcholine esterase) activity is localized on the N-terminal part of the protein. The mature protein was overexpressed in Escherichia coli and purified in a one-step procedure by choline-affinity chromatography and the enzymatic activity was followed using the chromophoric p-nitrophenyl-phosphorylcholine as a model substrate. The product of the enzymatic digestion of 3H-choline-labelled cell walls was shown to be phosphorylcholine. Inactivation of the pce gene in S. pneumoniae strains by insertion-duplication mutagenesis caused a unique change in colony morphology and a striking increase in virulence in the intraperitoneal mouse model. Pce may be a regulatory element involved with the interaction of S. pneumoniae with its human host. PMID- 11260478 TI - The Caulobacter crescentus flagellar gene, fliX, encodes a novel trans-acting factor that couples flagellar assembly to transcription. AB - The first flagellar assembly checkpoint of Caulobacter crescentus couples assembly of the early class II components of the basal body complex to the expression of class III and IV genes, which encode extracytoplasmic structures of the flagellum. The transcription of class III/IV flagellar genes is activated by the response regulator factor, FlbD. Gain of function mutations in flbD, termed bfa, can bypass the transcriptional requirement for the assembly of class II flagellar structures. Here we show that the class II flagellar gene fliX encodes a trans-acting factor that couples flagellar assembly to FlbD-dependent transcription. We show that the overexpression of fliX can suppress class III/IV gene expression in both wild-type and flbD-bfa cells. Introduction of a bfa allele of flbD into cells possessing a deletion in fliX restores motility indicating that FliX is not a structural component of the flagellum, but rather a trans-acting factor. Furthermore, extragenic motile suppressors which arise in DeltafliX cells map to the flbD locus. These results indicate that FlbD functions downstream of FliX in activating class III/IV transcription. beta-Lactamase fusions to FliX and analysis of cellular fractions demonstrate that FliX is a cytosolic protein that demonstrates some peripheral association with the cytoplasmic membrane. In addition, we have isolated a mutant allele of fliX that exhibits a bfa-like phenotype, restoring flbD-dependent class III/IV transcription in strains that contain mutations in class II flagellar structural genes. Taken together, these results indicated both a positive and negative regulatory function for FliX in coupling the assembly of class II basal body components to gene expression. PMID- 11260479 TI - Mammalian 14-3-3beta associates with the Chlamydia trachomatis inclusion membrane via its interaction with IncG. AB - Chlamydiae replicate intracellularly within a vacuole that is modified early in infection to become fusogenic with a subset of exocytic vesicles. We have recently identified four chlamydial inclusion membrane proteins, IncD-G, whose expression is detected within the first 2 h after internalization. To gain a better understanding of how these Inc proteins function, a yeast two-hybrid screen was employed to identify interacting host proteins. One protein, 14-3 3beta, was identified that interacted specifically with IncG. The interaction between 14-3-3beta and IncG was confirmed in infected HeLa cells by indirect immunofluorescence microscopy and interaction with a GFP-14-3-3beta fusion protein. 14-3-3 proteins are phosphoserine-binding proteins. Immunoprecipitation studies with [32P]-orthophosphate-labelled cells demonstrated that IncG is phosphorylated in both chlamydia-infected HeLa cells and in yeast cells expressing IncG. Site-directed mutagenesis of predicted 14-3-3 phosphorylation sites demonstrated that IncG binds to 14-3-3beta via a conserved 14-3-3-binding motif (RS164RS166F). Finally, indirect immunofluorescence demonstrated that 14-3 3beta interacts with Chlamydia trachomatis inclusions but not C. psittaci or C. pneumoniae inclusions. 14-3-3beta is the first eukaryotic protein found to interact with the chlamydial inclusion; however, its unique role in C. trachomatis pathogenesis remains to be determined. PMID- 11260480 TI - The puzzle of zmpB and extensive chain formation, autolysis defect and non translocation of choline-binding proteins in Streptococcus pneumoniae. AB - Choline-binding proteins (CBPs) from Streptococcus pneumoniae are involved in several important processes. Inactivation of zmpB, a gene that encodes a surface located putative zinc metalloprotease, in a S. pneumoniae serotype 4 strain was recently reported to reveal a composite phenotype, including extensive chain formation, lysis defect and transformation deficiency. This phenotype was associated with the lack of surface expression of several CBPs, including the major autolysin LytA. LytA, normally 36 kDa in size, was reported to form an SDS resistant 80 kDa complex with CinA. ZmpB was therefore proposed to control translocation of CBPs to the surface, possibly through the proteolytic release of CBPs (and RecA) from CinA. Based on the use of 12 independent mariner insertions in the zmpB gene of the well-characterized R6 laboratory strain, we could not confirm several of these observations. Our zmpB mutants: (i) did not form chains; (ii) lysed normally in the presence of deoxycholate, which indicates the presence of a functional autolysin; (iii) transformed at normal frequency; and (iv) contained bona fide CinA and LytA species. Polymorphism of ZmpB between R6 and the serotype 4 isolate could not account for the discrepancy, as inactivation of zmpB (through replacement by transposon-inactivated zmpB R6 alleles) in the latter strain did not affect separation of daughter cells and autolysis. The conflicting observations could be explained by our finding that the reportedly serotype 4 zmpB 'mutant' differed from its S. pneumoniae parent in lacking capsule and in exhibiting characteristic traits of the Streptococcus viridans group, including resistance to optochin. PMID- 11260481 TI - The availability of all technical modalities for pediatric liver transplant programs. PMID- 11260482 TI - Determinants of long-term survival of pediatric kidney grafts reported to the United Network for Organ Sharing kidney transplant registry. AB - Pediatric 1-yr kidney graft survival rates have steadily improved in the US so that, by 1998, over 90% of hospital-discharged young recipients had survived the first year post-transplantation (Tx). However, 25% of the early surviving kidney grafts failed at 5 yr, yielding a projected half-life of 10 yr. Given a median age at transplant of 13 yr (range 0-20 yr), 50% of all current pediatric kidney recipients will need a second graft before the age of 25 years. We examined 8,422 pediatric renal transplants reported to the United Network for Organ Sharing (UNOS) Kidney Transplant Registry and, by using a log-linear multifactorial analysis, determined the relative influence of 26 major transplant factors on long-term graft survival. Results are reported as percentages of assignable variation (totaling 100% for all 26 factors combined) in pediatric outcomes beyond 1 yr and as adjusted graft survival rates. Transplant center, recipient race and age, transplant year, and panel-reactive antibody (PRA) had assignable variation percentages of 25, 24, 16, 12, and 4, respectively. When combined, they accounted for 81% of changes in long-term survival. Besides center effects, Blacks, teenagers, and transplants performed before 1994 exhibited significantly (p <0.0001) lower adjusted 5-yr graft survival rates as did the few sensitized (PRA>40%) pediatric patients (p = 0.02). Patients transplanted with a living donor kidney demonstrated a 5% point advantage at 5 yr post-Tx over cadaver donor kidneys (p = 0.001). Although the survival rate of pediatric kidney transplants has improved steadily, the long-term outcomes for teenagers and for Black recipients lag significantly behind those of younger patients and non-Blacks. PMID- 11260483 TI - From living related to in-situ split liver transplantation: how to reduce waiting list mortality. AB - The insufficient number of suitable cadaveric organs for pediatric recipients is the cause of high pretransplant mortality rates and long waiting times. With the introduction of split and living related liver transplantation, the waiting list mortality rate has dropped from greater than 15% to less than 5% and children are transplanted more rapidly. A 5-year patient and graft survival rate of greater than 80% has been obtained in those centers where split and living related liver transplantations are routinely performed. The data analyzed in this paper show that the only current solution to cadaveric organ shortage is a 'multimodal' approach, where whole liver cadaveric transplantation is associated with split and living related liver transplantation. PMID- 11260484 TI - Extravesical ureteroneocystostomy with and without internalized ureteric stents in pediatric renal transplantation. AB - The use of ureteric double-J stents and the Lich-Gregoir (extravesical) technique of ureteroneocystotomy have both been shown to decrease the rate of urologic complications in adult kidney transplantation (Tx). There are, however, few studies of the systematic use of stents in pediatric renal Tx. Between 1991 and 1997, 32 consecutive pediatric renal transplant recipients routinely received a 6F-12 cm indwelling double-J stent and were studied prospectively. These patients were compared with 32 consecutive pediatric recipients in whom a stent was not used. The latter were transplanted between 1987 and 1991 and formed the control group. All patients had a Lich-Gregoir ureteroneocystotomy. Stents were removed under general-anesthetic cystoscopy 2 3 weeks after Tx. Immunosuppression for stented patients was polyclonal antibody induction, delayed (7-10 days) cyclosporin A, azathioprine, and prednisone. The control group received the same triple drug regimen but with no induction in 29 of the 32 patients. All patients were followed-up with at least one ultrasound evaluation in the first month, and a renal scan and repeat ultrasound were performed if there was any rise in serum creatinine. In the stented group there were two patients with urinary leak and no obstructions. In the non-stented group there were no leaks and one obstruction. There was no graft loss owing to urologic complications in either group. There were three cases of stent expulsion (all in girls) and one case of stent migration in the posterior urethra (a boy). The 1-yr graft survival rate was 90.6% in the stented group and 65.6% in the non-stented group. The prophylactic use of an indwelling ureteral stent in pediatric renal Tx did not reduce the risk of urinary leakage or obstruction. Stent migration is a common phenomenon and, while not a serious complication, is traumatic to children. Furthermore, removal of an internalized double-J stent requires a general anesthetic. We recommend using a stent for selected patients only. PMID- 11260485 TI - Psychosocial responses of adolescent cystic fibrosis patients to lung transplantation. AB - What psychosocial issues do adolescent cystic fibrosis (CF) patients experience after undergoing lung transplantation (Tx)? The aim of this study was to determine, using an ethnographic study design, the common themes and emotional responses in post-lung transplant adolescent CF patients of the Cardiothoracic Transplant Clinic at the Childrens Hospital Los Angeles. Nineteen CF lung transplant recipients were studied (eight males, 11 females: mean age at time of transplant, 15.7 +/- 2.7 yr). The mean time interval from Tx to interview was 25.4 months (range 1-58 months). Sixteen patients had living donor lobar lung Tx while three patients received cadaveric lungs. A series of 25 questions was used to assess the psychosocial impact of Tx, and a semi-structured interview focused on the following five domains: lifestyle, family functioning, social functioning, body image, and psychological functioning. The major themes identified by patients included: a strong desire to set and attain meaningful long-range goals, the need to control as many aspects of their lives as possible while dealing with parental over-protectiveness, and the adjustment to a new lifestyle. Common emotional responses included manageable fear/anxiety of lung rejection and uncertainty of the future, impatience with disruptions of daily routines caused by post-transplant medical management and its effect on the attainment of set goals, and frustration with parental over-protectiveness. In general, patients reported a positive outlook on life, with greater emphasis on sought-after goals as well as inter-personal relationships. This study demonstrates that adolescent CF transplant recipients develop long-term goals and plans for independence. By identifying and anticipating the emotional needs of this population, health care providers can assist patients in improving the quality of their lives from a physiological, as well as a psychological, viewpoint. PMID- 11260486 TI - Tacrolimus: the good, the bad, and the ugly. AB - The aim of this study was to evaluate the efficacy and side-effects of tacrolimus in pediatric transplant patients previously receiving cyclosporin A (CsA). This study was a retrospective chart review strengthened by a concomitant patient interview. Eleven pediatric cardiac or renal transplant patients, who had been converted from CsA to tacrolimus from October 1995 to January 1999 at The Cleveland Clinic Foundation, were included; there were six renal and five cardiac transplant patients. Each chart was reviewed to assess transplanted organ function pre- and post-conversion. For the six renal transplant patients, creatinine levels and biopsy findings were evaluated. For the five cardiac transplant patients, cardiac catheterization and routine biopsy data were analyzed likewise. Epstein Barr virus (EBV) status was also evaluated in each patient. In addition, each parent or patient was interviewed to ascertain dates of transplant, current medications, and side-effects. The patients' ages ranged from 6 to 20 yr (mean age 14.6 yr). All patients had been converted to tacrolimus. Eight patients were converted for treatment of refractory rejection, two were converted because of CsA-associated side-effects, and one patient was converted empirically for a history of multiple previous transplant rejections. Seven out of eight patients who received tacrolimus for rejection therapy improved. One patient had complete resolution of gingival hyperplasia. Another patient who previously developed hemolytic uremic syndrome on CsA had no further evidence of hemolysis. Four patients were weaned off steroid therapy. Despite conversion, two renal transplant patients progressed to chronic rejection. Five patients exhibited no side-effects. Side-effects experienced included transient hyperglycemia in conjunction with steroid use, headaches, and tremors that subsided rapidly. Four of 11 patients developed post-transplant lymphoproliferative disease (PTLD). Fortunately, reducing the dose of tacrolimus and/or surgical resection of the mass (if present), eradicated the disease. In conclusion, conversion therapy successfully provides an alternate treatment for acute rejection. It also enabled some patients to discontinue steroid therapy, maximizing growth potential. PTLD is a severe, potentially life-threatening complication that needs to be recognized and monitored closely. In conclusion, tacrolimus has been shown to be a very effective agent for the treatment of refractory organ rejection, but must be used cautiously. PMID- 11260487 TI - Psychological impact of liver transplantation on children's inner worlds. AB - We carried out an in-depth evaluation of psychosocial status in a sample of 18 children (mean age 6.8 yr, range 4.4-10.8 yr) who had suffered from severe liver disease and undergone orthotopic liver transplantation (OLT). Mean age at OLT was 3.4 yr. The assessment was psychoanalytically oriented and included individual sessions and testing procedures for children--the Children Apperception Test (CAT), the Weschsler Intelligence Scale for Children (WISC-R), the Weschsler Preschool and Primary Scale of Intelligence (WIPPSI), and the Human Figure Test- and a semi-structured interview with a separate questionnaire for parents. Patients were compared with an age- and gender-matched control group. The main findings in patients compared with controls were: IQ 91.6 (range 70-117) vs. 118 (range 94-135) (p<0.0001); immaturity of ego and drives (72.2% vs. 27.7%; p=0.018), fear of death (61.1% vs. 11.1%; p=0.04), anxiety of loss (50%, vs. 27.7%; p=NS), and depressive feelings (61.1% vs. 22.2%; p=0.04); a mild defect of body image (44.4% vs. 33.3%; p=NS) associated with recurrent representations of motionless (72.2% vs. 38.8%; p=NS) and inexpressive (88.8% vs. 16.6%; p<0.0001) human figures. Fantasies about OLT as a 'magic rebirth' or a 'body transformation' were detected in few patients (30%). Although a recurrent set of feelings, conflicts, and fantasies about OLT were expressed by children, individual specific psychological responses to this experience were often detected. In spite of the fact that approximately 50% of the parents mentioned emotional or behavioral disturbances of their child, only three parents were seriously concerned about this problem. The theme of transplantation was most often absent from communication between the child and their parents. Our results suggest that psychic 'working through' of the chronic liver disease and OLT experience is difficult for children. Further studies are necessary to verify whether changes of parental attitude to OLT as a 'family secret' may facilitate integration of the OLT experience in the child's personality development. PMID- 11260488 TI - Pretransplant varicella vaccination is cost-effective in pediatric renal transplantation. AB - Because of the severe complications that may result from varicella zoster virus (VZV) infection following renal transplantation (Tx), transplanted varicella susceptible children exposed to varicella are typically given varicella zoster immunoglobulin (VZIG) as prophylaxis or are admitted and treated with parenteral acyclovir if VZIG prophylaxis fails. As both VZIG and hospitalization are costly, prevention of varicella infection by vaccination could potentially result in significant cost savings in addition to decreasing morbidity and mortality. To test this hypothesis, we developed a decision-analysis model to evaluate the cost effectiveness of vaccinating patients with chronic renal failure (CRF) against varicella prior to renal transplant. Under baseline assumptions, vaccination for varicella pretransplant was a cost-effective strategy, with a cost of $211 per patient vaccinated compared with $1,828 per patient not vaccinated. The magnitude of cost savings from vaccination was sensitive to variations in the cost of varicella vaccine, the percentage of patients hospitalized for treatment with acyclovir, and the percentage of patients exposed to varicella infection. One- and two-way sensitivity analyses confirmed that vaccination was the dominant cost effective strategy under all conditions examined. We conclude that vaccination for varicella pretransplant is cost-effective for patients with CRF, and that the magnitude of cost savings is sensitive to the cost of hospitalization, the percentage of patients exposed to varicella, and the cost of varicella vaccination. Pending results of ongoing studies of the safety and efficacy of VZV vaccine in children with CRF, we recommend that VZV vaccine be given to all children with CRF. PMID- 11260489 TI - Fading renal hyperfiltration in children following liver transplantation. AB - In a prospective longitudinal study, we investigated the renal function (RF) of 23 children before and after orthotopic liver transplantation (OLT). The aim was to assess both the outcome of pretransplant hyperfiltration and the clinical nephrotoxic effects of cyclosporin A (CsA); children with decreased RF prior to OLT were therefore excluded. The RF study of the 13 remaining patients included glomerular filtration rate (GFR) and effective renal plasma flow (RPF) measured by inulin (Cin: mL/min/1.73 m2) and para-amino hippurate (Cpah: mL/min/1.73 m2) clearances, respectively. Hyperfiltration prior to OLT was observed in six children, i.e. Cin>170 [range 172-230] and Cpah>800 [808-1,133]. A significant decrease in RF was noted as soon as 6 months after OLT: Cin (mean+/-SD)=107+/-23 vs. 158+/-46 (p<0.003); Cpah=583+/-119 vs. 791+/-243 (p<0.004). This was due to loss of hyperfiltration in the six children, as there was no significant difference in RF before and 6 months after OLT in the other seven children. With a 36-month follow-up, there was no correlation between CsA trough blood level and RF. In conclusion, following OLT, RF underwent early changes owing to loss of prior hyperfiltration in children without impaired RF before OLT. In addition, no evidence of CsA nephrotoxicity was found and RF remained stable during follow-up. PMID- 11260490 TI - Portal vein phlebolithiasis found post-liver transplantation in the native liver of a child with biliary atresia. AB - Biliary atresia is defined as partial or total obliteration of the extra-hepatic bile ducts. In advanced cases, liver transplantation (LTx) is considered the most appropriate treatment. This report describes a female patient whose biliary atresia and subsequent cirrhosis required LTx at 1 yr of age. Macroscopic inspection of the hilar region of the native liver post-Tx revealed the formation of a pouch in the hepatic duct and a stone in the lumen of the portal vein. X-ray diffraction analysis showed that the stone was composed of cholesteryl cinnamate, gluconic acid phenylhydrazide, Na beta broma-allyl mercaptomethyl penicillinate, and Al2O3 crystals. While the cholesterol component is a known element of gallstones, we attributed the Na beta broma-allyl mercaptomethyl penicillinate to the patient's drug therapy. Our literature search revealed no previous record or crystallographic analysis of portal vein phlebolithiasis. In this report we describe this rare finding. PMID- 11260491 TI - B-cell dysfunction and depletion using mycophenolate mofetil in a pediatric combined liver and kidney graft recipient. AB - The use of mycophenolate mofetil (MMF) in combination with cyclosporin A (CsA) and steroids is well established after kidney transplantation (Tx) in children. A 9-yr-old girl with primary hyperoxaluria type 1 and systemic oxalosis underwent a combined kidney and liver Tx at our institution. The post-operative immunosuppression consisted of CsA, prednisolone, and MMF. Four weeks post transplant the girl suffered from a severe urinary tract infection caused by Pseudomonas aeruginosa, when the serum immunoglobulin G (IgG) concentration was found to be critically low (<1.53 g/L). Additionally, there was an isolated B cell depletion (240/microL) at that time. In the following course, the B-cell count was significantly diminished until the MMF was stopped 13 weeks post transplant. As a result of the very low serum IgG concentration, intravenous immunoglobulin (IVIG) substitution was necessary. There was no significant loss of immunoglobulins in the ascites and urine and no other medication with possible side-effects on B cells was given. We suggest that MMF can lead to suppressed IgG production by B cells and can cause a defective differentiation into mature B cells. In vitro studies demonstrated these effects of MMF on B cells, but no in vivo cases of this phenomenon have been reported. B-cell counts and serum IgG concentrations returned to normal values after discontinuing the MMF. As we can assume that the observed B-cell dysfunction and depletion were MMF related, we suggest that serum IgG concentrations should be monitored when MMF is used after solid-organ Tx. PMID- 11260492 TI - Thoracoabdominal bypass graft with liver retransplantation for the treatment of a pseudoaneurysm of the supraceliac aorta after liver transplantation. AB - Pseudoaneurysm following liver transplantation is a rare but life-threatening complication. Treatment is directed towards control of fatal bleeding. Ligation with or without revascularization of the graft is the treatment of choice. When revascularization is not possible, or the liver does not tolerate the arterial blood deprivation, retransplantation is the only option. We report a case of a 14 month-old girl who developed a pseudoaneurysm at the anastomosis between the recipient supraceliac aorta and the donor graft. The pseudoaneurysm was excised and the aorta was ligated above and below it. An extra-anatomical thoracoabdominal arterial graft was used to provide arterial blood supply to the lower torso and also to arterialize a new orthotopic liver graft. This is the first reported case of the use of thoracoabdominal jump graft to vascularize a transplanted liver. PMID- 11260493 TI - Maize protein kinase CK2: regulation and functionality of three beta regulatory subunits. AB - Biochemical and crystallographic data suggest that, in contrast with other organisms, the active maize protein kinase CK2 might be composed simply of a catalytic polypeptide (CK2alpha), thus lacking CK2beta regulatory subunits. To investigate the existence and functionality of CK2beta regulatory subunits in Zea mays, we have screened a maize cDNA library using different approaches and have isolated three full-length cDNAs encoding CK2beta regulatory subunits (CK2beta-1, CK2beta-2 and CK2beta-3) and a cDNA coding for a novel CK2alpha catalytic subunit, CK2alpha-3. The pattern of expression of all these alpha/beta subunits has been studied in different organs and developmental stages using specific probes for each isoform, and indicates that while CK2alpha subunits are constitutive, CK2beta subunits are expressed differentially during embryo development. The yeast two-hybrid system and pull-down assays have been used to study specific interactions between the different subunits. While CK2alpha subunits are unable to self-associate, preferential interactions between alpha/beta isoforms and beta/beta isoforms can be predicted. Furthermore, we show that maize CK2alpha/beta subunits assemble into a structural tetrameric complex which has very similar properties to those described in other organisms, and that expression of maize CK2beta subunits in yeast allows the rescue of the phenotypic defects associated to the lack of CK2 function, thus demonstrating the functionality of maize CK2beta regulatory subunits. PMID- 11260494 TI - Quinone oxidoreductase message levels are differentially regulated in parasitic and non-parasitic plants exposed to allelopathic quinones. AB - Allelopathic chemicals released by plants into the rhizosphere have effects on neighboring plants ranging from phytoxicity to inducing organogenesis. The allelopathic activity of naturally occurring quinones and phenols is primarily a function of reactive radicals generated during redox cycling between quinone and hydroquinone states. We isolated cDNAs encoding two distinct quinone oxidoreductases from roots of the parasitic plant Triphysaria treated with the allelopathic quinone 2,6-dimethoxybenzoquinone (DMBQ). TvQR1 is a member of the zeta-crystallin quinone oxidoreductase family that catalyzes one-electron quinone reductions, generating free radical semiquinones. TvQR2 belongs to a family of detoxifying quinone oxidoreductases that catalyze bivalent redox reactions which avoid the radical intermediate. TvQR1 and TvQR2 message levels are rapidly upregulated in Triphysaria roots as a primary response to treatment with various allelopathic quinones. Inhibition of quinone oxidoreductase enzymatic activity with dicumarol prior to quinone treatment resulted in increased transcript levels. While TvQR2 homologs were upregulated by DMBQ in roots of all plants examined, TvQR1 homologs were upregulated only in roots of parasitic plants. Phylogenetic trees constructed of TvQR1 and TvQR2 protein homologs in Archea, Eubacteria and Eukaryotes indicated that both gene families are ancient, yet the families have dissimilar evolutionary histories in angiosperms. We hypothesize that TvQR2-like proteins function to detoxify allelopathic quinones in the rhizosphere, while TvQR1 has specific functions associated with haustorium development in parasitic plants. PMID- 11260495 TI - Negative autoregulation of the Arabidopsis homeobox gene ATHB-2. AB - The Arabidopsis homeobox gene ATHB-2 is tightly regulated by light signals, and is thought to direct morphological changes during shade avoidance responses. To understand how ATHB-2 mediates light signals in plant morphogenesis, we investigated its transcriptional network. We constructed a gene encoding a chimeric transcription factor (HD-Zip-2-V-G) that is expected to activate target genes of ATHB-2 in a glucocorticoid-dependent manner. In transgenic Arabidopsis plants expressing HD-Zip-2-V-G, glucocorticoid treatment activates the ATHB-2 gene itself, independent of de novo protein synthesis. An in vitro DNase I footprinting experiment showed that recombinant ATHB-2 protein specifically bound to an ATHB-2 promoter region. These complementary results indicate that ATHB-2 recognizes its own promoter. Consistent with the fact that ATHB-2 itself has been shown to act as a repressor, expression of the endogenous ATHB-2 gene was repressed in transgenic plants overexpressing an ATHB-2 transgene. Moreover, target-gene analysis using the HD-Zip-2-V-G suggested that ATHB-2 recognizes other HD-Zip II subfamily genes. We conclude that ATHB-2 has a negative autoregulatory loop and may be involved in a complicated transcriptional network involving paralogous genes, as is the case with animal homeobox genes. PMID- 11260496 TI - Novel auxin transport inhibitors phenocopy the auxin influx carrier mutation aux1. AB - The hormone auxin is transported in plants through the combined actions of diffusion and specific auxin influx and efflux carriers. In contrast to auxin efflux, for which there are well documented inhibitors, understanding the developmental roles of carrier-mediated auxin influx has been hampered by the absence of specific competitive inhibitors. However, several molecules that inhibit auxin influx in cultured cells have been described recently. The physiological effects of two of these novel influx carrier inhibitors, 1 naphthoxyacetic acid (1-NOA) and 3-chloro-4-hydroxyphenylacetic acid (CHPAA), have been investigated in intact seedlings and tissue segments using classical and new auxin transport bioassays. Both molecules do disrupt root gravitropism, which is a developmental process requiring rapid auxin redistribution. Furthermore, the auxin-insensitive and agravitropic root-growth characteristics of aux1 plants were phenocopied by 1-NOA and CHPAA. Similarly, the agravitropic phenotype of inhibitor-treated seedlings was rescued by the auxin 1 naphthaleneacetic acid, but not by 2,4-dichlorophenoxyacetic acid, again resembling the relative abilities of these two auxins to rescue the phenotype of aux1. Further investigations have shown that none of these compounds block polar auxin transport, and that CHPAA exhibits some auxin-like activity at high concentrations. Whilst results indicate that 1-NOA and CHPAA represent useful tools for physiological studies addressing the role of auxin influx in planta, 1 NOA is likely to prove the more useful of the two compounds. PMID- 11260497 TI - Living under a "dormant" canopy: a molecular acclimation mechanism of the desert plant Retama raetam. AB - Desert plants are exposed to a combination of environmental stress conditions, including low water availability, extreme temperature fluctuations, high irradiance and nutrient deprivation. Studying desert plants within their natural habitat may therefore reveal novel mechanisms and strategies that enable plants to resist stressful conditions. We studied the acclimation of Retama raetam, an evergreen stem-assimilating desert plant, to growth within an arid dune ecosystem. Retama raetam contained two different populations of stems: those of the upper canopy, exposed to direct sunlight, and those of the lower canopy, protected from direct sunlight. During the dry season, stems of the upper canopy contained a very low level of a number of essential proteins, including the large and small subunits of rubisco, ascorbate peroxidase and the D1 subunit of the reaction centre of photosystem II. However, RNA encoding these proteins was present; cytosolic transcripts were associated with polysomes, while chloroplastic transcripts were not. Upon water application, as well as following the first rainfall of the season, these "photosynthetically suppressed" stems recovered and accumulated essential proteins within 6-24 h. In contrast, stems of the lower canopy contained the essential proteins throughout the dry season. We suggest that R. raetam uses an acclimation strategy of "partial plant dormancy" in order to survive the dry season. "Dormancy", as evident by the post transcriptional suppression of gene expression, as well as the suppression of photosynthesis, was induced specifically in stems of the upper canopy which protect the lower canopy by shading. PMID- 11260498 TI - Size constraints for targeting post-transcriptional gene silencing and for RNA directed methylation in Nicotiana benthamiana using a potato virus X vector. AB - Using a recombinant potato virus X (PVX) vector, we investigated the relationship between the length of RNA sequence identity with a transgene and the ability to promote post-transcriptional gene silencing (PTGS) and transgene methylation. The lower size limit required for targeting reporter transgene mRNA de novo using PTGS was 23 nucleotides (nt) of complete identity, a size corresponding to that of small RNAs associated with PTGS in plants and RNA interference (RNAi) in animals. The size and sequence specificity were also explored for PTGS-associated transgene methylation and for the targeting of the vector RNA. The PTGS-competent short sequences resulted in similar patterns of methylation. In all cases, including specific sequences of 33 nt with or without symmetrical cytosine residues, the methylation was distributed throughout the transcribed region of the transgene. In contrast, short sequences lacking symmetrical cytosines were less efficient at promoting PTGS of the transgene mRNA. Short gfp sequences in the PVX vector provided as effective a target for the degradation of viral RNA as was found for PVX carrying the complete gfp cDNA. Short sequences were able to initiate PTGS of an endogenous gene, phyotene desaturase, although this occurred in the absence of DNA methylation. This experimental approach provides important insights into the relationship between short RNA sequences and PTGS. PMID- 11260499 TI - Genetic dissection of blue-light sensing in tomato using mutants deficient in cryptochrome 1 and phytochromes A, B1 and B2. AB - Several novel allelic groups of tomato (Solanum lycopersicum L.) mutants with impaired photomorphogenesis have been identified after gamma-ray mutagenesis of phyA phyB1 double-mutant seed. Recessive mutants in one allelic group are characterized by retarded hook opening, increased hypocotyl elongation and reduced hypocotyl chlorophyll content under white light (WL). These mutants showed a specific impairment in response to blue light (BL) resulting from lesions in the gene encoding the BL receptor cryptochrome 1 (cry1). Phytochrome A and cry1 are identified as the major photoreceptors mediating BL-induced de etiolation in tomato, and act under low and high irradiances, respectively. Phytochromes B1 and B2 also contribute to BL sensing, and the relative contribution of each of these four photoreceptors differs according to the light conditions and the specific process examined. Development of the phyA phyB1 phyB2 cry1 quadruple mutant under WL is severely impaired, and seedlings die before flowering. The quadruple mutant is essentially blind to BL, but experiments employing simultaneous irradiation with BL and red light suggest that an additional non-phytochrome photoreceptor may be active under short daily BL exposures. In addition to effects on de-etiolation, cry1 is active in older, WL grown plants, and influences stem elongation, apical dominance, and the chlorophyll content of leaves and fruit. These results provide the first mutant based characterization of cry1 in a plant species other than Arabidopsis. PMID- 11260500 TI - Analysis of PEAM4, the pea AP1 functional homologue, supports a model for AP1 like genes controlling both floral meristem and floral organ identity in different plant species. AB - APETALA1 (AP1) and its homologue SQUAMOSA (SQUA) are key regulatory genes specifying floral meristem identity in the model plants Arabidopsis and Antirrhinum. Despite many similarities in their sequence, expression and functions, only AP1 appears to have the additional role of specifying sepal and petal identity. No true AP1/SQUA-functional homologues from any other plant species have been functionally studied in detail, therefore the question of how the different functions of AP1-like genes are conserved between species has not been addressed. We have isolated and characterized PEAM4, the AP1/SQUA-functional homologue from pea, a plant with a different floral morphology and inflorescence architecture to that of Arabidopsis or Antirrhinum. PEAM4 encodes for a polypeptide 76% identical to AP1, but lacks the C-terminal prenylation motif, common to AP1 and SQUA, that has been suggested to control the activity of AP1. Nevertheless, constitutive expression of PEAM4 caused early flowering in tobacco and Arabidopsis. In Arabidopsis, PEAM4 also caused inflorescence-to-flower transformations similar to constitutive AP1 expression, and was able to rescue the floral organ defects of the strong ap1-1 mutant. Our results suggest that the control of both floral meristem and floral organ identity by AP1 is not restricted to Arabidopsis, but is extended to species with diverse floral morphologies, such as pea. PMID- 11260501 TI - Expression of MsLEC1- and MsLEC2-antisense genes in alfalfa plant lines causes severe embryogenic, developmental and reproductive abnormalities. AB - Although it has been proposed that plant lectins play a number of roles, the function of these proteins in normal plant growth and development has been unclear. To analyze the functions of putative alfalfa lectin genes, lines of transgenic alfalfa plants expressing approximately half of the open reading frame of MsLEC1 or MsLEC2, in the antisense or sense orientation, were established and analyzed. The antisense plants displayed severe abnormalities in embryogenesis, and both vegetative and reproductive development were perturbed. Some differences were observed between MsLEC1- and MsLEC2-antisense plants, and abnormalities were especially severe during the early stages of development in both the primary and secondary transgenic generations. In contrast, vector-control and sense-transgene plants exhibited normal growth and development. MsLEC1 and MsLEC2 mRNA accumulation levels were reduced in cognate antisense plants, especially during the later stages of embryogenesis, but also tended to be low in MsLEC1 sense transgene plants. However, correlated with the phenotypic abnormalities observed in the MsLEC1-antisense plants was the specific reduction in the accumulation of a candidate MsLEC1 protein. Our results suggest that the MsLEC1 and MsLEC2 gene products, in addition to being important for embryogenesis, are required throughout alfalfa development. PMID- 11260502 TI - Rubisco activase: an enzyme with a temperature-dependent dual function? AB - Heat treatment of intact spinach leaves was found to induce a unique thylakoid membrane association of an approximately 40 kDa stromal protein. This protein was identified as rubisco activase. Most of the rubisco activase was sequestered to the thylakoid membrane, particularly to the stroma-exposed regions, during the first 10 min of heat treatment at 42 degrees C. At lower temperatures (38-40 degrees C) the association of rubisco activase with the thylakoid membrane occurred more slowly. The temperature-dependent association of rubisco activase with the thylakoid membrane was due to a conformational change in the rubisco activase itself, not to heat-induced alterations in the thylakoid membrane. Association of the 41 kDa isoform of rubisco activase occurred first, followed by the binding of the 45 kDa isoform to the thylakoid membrane. Fractionation of thylakoid membranes revealed a specific association of rubisco activase with thylakoid-bound polysomes. Our results suggest a temperature-dependent dual function for rubisco activase. At optimal temperatures it functions in releasing inhibitory sugar phosphates from the active site of Rubisco. During a sudden and unexpected exposure of plants to heat stress, rubisco activase is likely to manifest a second role as a chaperone in association with thylakoid-bound ribosomes, possibly protecting, as a first aid, the thylakoid associated protein synthesis machinery against heat inactivation. PMID- 11260503 TI - Non-conservative substitutions distinguish previously uncharacterized HLA-A molecules. AB - The extent of class I HLA polymorphism is not yet realized, and to provide a glimpse of the HLA-A polymorphism which remains undetected, we have analyzed approximately 3,700 National Marrow Donor Program (NMDP) Donor/Recipient Pair Retrospective Study Samples with HLA-A DNA sequence-based typing (SBT). Seventeen new HLA-A alleles were detected, with a total of 19 nucleotide substitutions distinguishing these new alleles from their closest HLA-A relatives. Nearly all of the new alleles differ by single nucleotide substitutions; a majority of these substitutions can be explained by gene conversion events but 6 alleles likely originated by point mutation. Fifteen of the 19 nucleotide substitutions translate into amino acid differences in the molecule. Structurally, the inferred amino acid alterations were non-conservative in terms of chemical property, and most substitutions were positioned in 1 or more of the specificity pockets which determine peptide binding. Although these new alleles were identified in a primarily Caucasian sample population, 9 of the 17 new HLA-A alleles were found in samples of non-Caucasoid origin. A new allele detection rate of 1 in approximately 200 individuals in our data set would, therefore, be higher in a non-Caucasoid sample population. In summary, the single nucleotide substitutions that distinguish undetected HLA-A alleles translate into functionally distinct HLA-A molecules. Further studies of the role of HLA-A in transplantation, in disease association, and in evolution must therefore accommodate the discovery of new alleles differing by single nucleotides. PMID- 11260504 TI - A MAGE-3 peptide recognized on HLA-B35 and HLA-A1 by cytolytic T lymphocytes. AB - Antigens encoded by MAGE genes are of particular interest for cancer immunotherapy because of their strict tumoral specificity and because they are shared by many tumors. Antigenic peptide EVDPIGHLY encoded by MAGE-3 and known to be presented by HLA-A*0101 is currently being used in therapeutic vaccination trials. We report here that a cytolytic T-lymphocyte (CTL) clone, which is restricted by HLA-B*3501, recognizes the same peptide and, importantly, lyses HLA B*3501 tumor cells expressing MAGE-3. These results infer that the current clinical use of peptide EVDPIGHLY can now be extended to HLA-B*3501 patients. PMID- 11260505 TI - Lack of HLA-class I antigens in human neuroblastoma cells: analysis of its relationship to TAP and tapasin expression. AB - We studied the constitutive and the interferon (IFN)-gamma-induced expression of HLA class I antigen heavy chain, beta2-microglobulin (beta2m), TAP-1, TAP-2 and tapasin in a panel of eleven neuroblastoma cell lines. Surface expression of HLA class I antigens was low in eight out of eight neuroblastoma cell lines bearing MYC-N amplification and/or 1p deletion, while two out of three neuroblastoma cell lines lacking these genetic alterations showed normal expression. IFN-gamma treatment restored HLA class I antigen surface expression in all neuroblastoma cell lines. Eight out of 11 neuroblastoma cell lines did not express TAP-1 mRNA and three of them also lacked TAP-2 mRNA. beta2 m mRNA was barely detectable or absent in five neuroblastoma cell lines, while tapasin mRNA was always expressed. IFN-gamma upregulated the expression of HLA class I heavy chain, beta2 m, TAP-1, TAP-2 and tapasin, as detected at mRNA or protein level. Post-transcriptional events were involved in altered TAP-1 and beta2 m expression in one peculiar neuroblastoma cell line. These data indicate that multiple mechanisms play a role in the HLA class I antigen-deficient phenotype of human neuroblastoma. PMID- 11260506 TI - HLA genes in Macedonians and the sub-Saharan origin of the Greeks. AB - HLA alleles have been determined in individuals from the Republic of Macedonia by DNA typing and sequencing. HLA-A, -B, -DR, -DQ allele frequencies and extended haplotypes have been for the first time determined and the results compared to those of other Mediterraneans, particularly with their neighbouring Greeks. Genetic distances, neighbor-joining dendrograms and correspondence analysis have been performed. The following conclusions have been reached: 1) Macedonians belong to the "older" Mediterranean substratum, like Iberians (including Basques), North Africans, Italians, French, Cretans, Jews, Lebanese, Turks (Anatolians), Armenians and Iranians, 2) Macedonians are not related with geographically close Greeks, who do not belong to the "older" Mediterranenan substratum, 3) Greeks are found to have a substantial relatedness to sub-Saharan (Ethiopian) people, which separate them from other Mediterranean groups. Both Greeks and Ethiopians share quasi-specific DRB1 alleles, such as *0305, *0307, *0411, *0413, *0416, *0417, *0420, *1110, *1112, *1304 and *1310. Genetic distances are closer between Greeks and Ethiopian/sub-Saharan groups than to any other Mediterranean group and finally Greeks cluster with Ethiopians/sub-Saharans in both neighbour joining dendrograms and correspondence analyses. The time period when these relationships might have occurred was ancient but uncertain and might be related to the displacement of Egyptian-Ethiopian people living in pharaonic Egypt. PMID- 11260507 TI - HLA class I in three West African ethnic groups: genetic distances from sub Saharan and Caucasoid populations. AB - Fulani of Burkina Faso (West Africa) are a particularly interesting ethnic group because of their lower susceptibility to Plasmodium falciparum malaria as compared to sympatric populations, Mossi and Rimaibe. Moreover, the occurrence of a Caucasoid component in their genetic make-up has been suggested on the basis of their physical traits and cultural traditions even though this view was not supported by genetic studies. A total of 149 unrelated subjects (53 Mossi, 47 Rimaibe and 49 Fulani) have been typed for 97 HLA class I alleles with the amplification refractory mutation system/polymerase chain reaction (ARMS/PCR) technique. Mossi and Rimaibe data were pooled since none of the 42 statistically testable alleles exhibited a significant heterogeneity. These pooled gene frequencies were found to be very different from those of Fulani: a certain (P<0.001) or a likely (0.001 0.05). The total success rate (success with or without modification) was 100% in both arms. No major adverse effects were observed in either groups. CONCLUSION: CFP is as effective and safe as CFZ for the empirical treatment of febrile episodes in neutropenic patients with solid tumors. PMID- 11260565 TI - Effects of high-dose methotrexate on the hemostatic system in childhood acute lymphoblastic leukemia. AB - BACKGROUND: Thromboembolic and hemorrhagic complications are significant causes of death in patients with malignancy. These are well-known with the use of certain drugs. This study was planned to investigate whether there was any effect of high-dose methotrexate on the hemostatic system in childhood acute lymphoblastic leukemia. PROCEDURE: To evaluate the hemostatic system, we investigated coagulation screening tests (prothrombin time, activated partial thromboplastin time, and fibrinogen), coagulation inhibitors (protein C, protein S, and antithrombin III), and fibrinolytic system (fibrin degradation products and tissue plasminogen activator). These parameters were measured in 35 cycles of high dose-methotrexate (3 g/m(2)) of 20 childhood acute lymphoblastic leukemia cases at baseline and on days 1 and 7 after the therapy. RESULTS: We found that high-dose methotrexate administration adversely affected both the coagulation system (prolonged prothrombin time and activated partial thromboplastin time and decreased fibrinogen levels) and coagulation inhibitors (decreased protein C, protein S, antithrombin III) on day 1 after chemotherapy compared to the baseline values. The hemostatic parameters began to improve on day 7 after chemotherapy, except for fibrin degradation products. Tissue plasminogen activator levels were not changed with the therapy. CONCLUSIONS: Coagulation cascade (prolonged prothrombin time and activated partial thromboplastin time and decreased fibrinogen) and coagulation inhibitors (decreased protein C, protein S, and antithrombin III levels) have been found to be affected by high-dose methotrexate therapy, but these transient changes did not cause clinical thromboembolic or hemorrhagic complications. PMID- 11260567 TI - Cognitive functions of adolescent childhood cancer survivors assessed by event related potentials. AB - BACKGROUND: Neurophysiological methods were applied to examine subtle central nervous system (CNS) adverse effects for adolescent childhood cancer survivors. We analyzed auditory event-related potentials (ERPs)-P300 and MMN/P3a complex-to find out whether there was impaired attention orientation in asymptomatic cancer survivors, and whether these ERP methods could be used as more objective tools in detecting those survivors who might need academic testing. Previous clinical studies of P300 have focused on leukemia survivors. MMN for cancer survivors has not been reported. PROCEDURE: The subjects were survivors of childhood leukemia (n=11) and solid tumors (n=8), as well as healthy controls (n=10). The mean age was 15.5 years for survivors and 15.9 years for controls. Pure sine-wave tones (500 and 553 Hz, 100 ms) were used as stimuli in an oddball paradigm. The ERPs to frequency change were measured. MMN recordings were performed in a passive non attended situation where the subject was watching a voiceless video cartoon. P300 was produced thereafter, but in an active attend situation, by the same auditory oddball paradigm as MMN. RESULTS: A significant difference was detected between the groups for the latency of P300 at electrodes Cz (P = 0.03) and C4 (P = 0.05). The cancer survivors had prolonged P300 latencies as an indication of prolonged short-term memory processing. The area and latency parameters of MMN did not differ significantly between the study groups, but in cancer survivors, the area and the mean amplitude of the subsequent P3a wave were diminished. The results indicate that the discrimination process was not as easy for the survivors as for the controls. However, it seems that in cancer survivors the basic mechanism starting attention shift to novel stimuli is not impaired. CONCLUSIONS: These results indicate that it is important to carefully evaluate the proper methods for the teaching of children who are survivors of malignancies. The auditory information may not always lead to the best possible learning results. PMID- 11260568 TI - Long-acting morphine for pain control in paediatric oncology. AB - BACKGROUND: Guidelines for treatment of paediatric cancer pain recommend the usage of long-acting morphine. However, published paediatric experience with this drug is restricted to 147 children not systematically evaluated, and thus insufficient. We aimed to systematically analyse the age-dependent effects and adverse effects of long-acting morphine in paediatric cancer patients. PROCEDURE: Ninety-five children aged 1 to 19 years were enrolled in a collaborative retrospective study conducted over seven-and-a-half years. Pain was scored according to a numeric rating scale (NRS, range 0 to 5), and the corresponding medication was recorded. RESULTS: In 83 children documentation period started during morphine treatment (71, oral long-acting; 1, rectal; 11, IV). Mean oral/equivalent morphine starting dose was 1.3 mg/kgbw/d (SD 0.9). Mean end dose was 2.8 mg/kgbw/d (SD 2.7). Infants aged < 7 years received the highest average dose (2.6 mg/kgbw/d, SD 2.8), while patients > 12 years received the lowest dose (1.4 mg/kgbw/d, SD 1.1). Median pain intensity decreased from score 1.0 (mean 1.2) NRS at the beginning to 0 (mean 0.6) NRS at the end. The proportion of patients scoring > 2 NRS (severe or most severe pain) under morphine treatment decreased from 26 to 12% (P = 0.08). In children > 12 years pruritus was frequently observed (23% of patients). In all age groups, there were no severe adverse effects during the study period. CONCLUSIONS: In paediatric haematology/oncology, pain control by oral long-acting morphine proved to be safe and effective even in the very young patients. The pharmacological properties of long-acting morphine are ideally suited for paediatric use, combining efficacy and compatibility. PMID- 11260569 TI - Pulmonary abnormalities at long-term follow-up of patients with Langerhans cell histiocytosis. AB - BACKGROUND: In Langerhans cell histiocytosis (LCH) pulmonary involvement, which is often initially asymptomatic, may contribute to significant morbidity and mortality. To determine the long-term prognosis, a cross-sectional study was undertaken. PROCEDURE: Forty-one patients with > or = 5 years follow-up after the diagnosis of LCH were interviewed and underwent physical examination, blood tests, a chest X-ray and a high-resolution CT (HRCT) of the lungs. All patients included had been referred to the Department of Pediatrics at the Karolinska Hospital in Stockholm between July 1962 and February 1990 (median follow-up 16 years). Biopsies from all patients were reviewed and confirmed to be consistent with LCH. Information on previous clinical features including treatment and the results of chest X-rays were also collected for risk factor analysis. RESULTS: Radiographic abnormalities of the lungs (cysts and/or emphysema), found in 10/41 (24%) at follow-up, were classified into five groups according to the extent of the cysts. These patients had more often suffered from multisystem than from single-system disease (P = 0.01), were significantly older at diagnosis (P < 0.001), and had been more heavily treated with chemotherapy and/or radiotherapy. They were also more frequently smokers (P < 0.0001) and 7/10 (70%) had suffered lung involvement at diagnosis. At the time of diagnosis of the pulmonary involvement, 4/10 (40%) patients had respiratory symptoms, but only 2/10 (20%) had symptoms at follow-up. CONCLUSIONS: Ten (24%) of the 41 patients had abnormal findings on radiological examination of the lungs at long-term follow-up and seven are or had been smokers. It is of great importance that patients with LCH be informed about smoking-related pulmonary morbidity. Prolonged monitoring of the lungs for smokers and patients with known pulmonary involvement is recommended. PMID- 11260570 TI - Hepatic dysfunction in children with acute lymphoblastic leukemia in remission: relation to hepatitis infection. AB - BACKGROUND: Viral hepatitis is a cause of hepatic dysfunction in children with ALL in remission during maintenance therapy is debated. The aims of the current study were (1) to explore the incidence of hepatic dysfunction in a group of children (Egyptian and Saudi) with ALL under maintenance therapy, (2) to study the prevalence of hepatitis B (HBV) and/or C (HCV) infection and their contributions to chronic liver disease that might be induced by maintenance therapy. PROCEDURE: The current study included 105 children with ALL (54 Egyptian and 51 Saudi). All eligible patients had been on maintenance therapy for at least 12 months and all had serial assessments of liver function. These included determination of total bilirubin, AST, ALT, and alkaline phosphatase. Markers for HBV and HCV including HBsAg, anti-HBC, and anti-HCV and for some patients HCV RNA by PCR were studied. Percutaneous liver biopsy was performed for a group of children. RESULTS: The prevalence of hepatitis infection (HBV and/or HCV) among Egyptian children was found to be high (43/54-80%). Only five Saudi children had evidence of exposure to HBV (5/51-9.8%), P<0.0001. During the period of study, 22 Egyptian patients vs. four Saudi patients (41 vs. 7.8%, P<0.0001) experienced at least one episode of elevation of liver enzymes, three times the upper limit of normal or more. Twenty-six of the 48 patients (54%) with HBV and/or HCV infection had episodes of elevated liver enzymes, while there was no occurrence among the patients negative for HBV and HCV. In patients with HBV infection, the presence of HBsAg was strongly associated (100%) with elevated liver enzymes. Histopathologic examination of liver biopsies obtained from 35 patients revealed that all five patients negative for HBV and HCV had normal liver biopsies in spite of being under maintenance therapy. CONCLUSION: In children undergoing treatment for ALL, elevations in liver enzymes may be primarily due to hepatitis viruses. However, maintenance therapy using known hepatotoxic drugs, may have additive deleterious effects. Liver enzymes are normalized in affected patients when maintenance therapy is temporarily suspended. PMID- 11260571 TI - Art therapy as support for children with leukemia during painful procedures. AB - BACKGROUND: Children with leukemia undergo painful procedures such as lumbar puncture and bone marrow aspiration. To overcome pain, certain units offer total anesthesia; others offer generic support; others offer no preparation at all. Since September, 1997, we have provided leukemic children with art therapy (AT), a nonverbal and creative modality that develops coping skills. Our goal is to prevent anxiety and fear during painful interventions as well as prolonged emotional distress. PROCEDURE: We treated 32 children aged 2-14 years. The modes of AT before, during, and after the punctures were as follows: clinical dialogue to calm children and help them cope with painful procedures; visual imagination to activate alternative thought processes and decrease the attention towards overwhelming reality and raise the peripheral sensitivity gate; medical play to clarify illness, eliminate doubts, and offer control over threatening reality; structured drawing to contain anxiety by offering a structured, predictable reality (the drawing) that was controllable by children; free drawing to allow children to externalize confusion and fears; and dramatization to help children accept and reconcile themselves to body changes. RESULTS: Children hospitalized before September, 1997, exhibited resistance and anxiety during and after painful procedures. By contrast, children provided with AT from the first hospitalization exhibited collaborative behavior. They or their parents asked for AT when the intervention had to be repeated. Parents declared themselves better able to manage the painful procedures when AT was offered. CONCLUSION: AT was shown to be a useful intervention that can prevent permanent trauma and support children and parents during intrusive interventions. PMID- 11260572 TI - Survival trends of childhood cancer diagnosed during 1970-1994 in Piedmont, Italy: a report from the Childhood Cancer Registry. AB - BACKGROUND: The Childhood Cancer Registry of Piedmont (CCRP) started its activity in 1967. It is population based and covers the Piedmont Region (population 4,500,000; NW Italy). This article reports on time trends in survival after a childhood cancer diagnosed during 1970-1994. PROCEDURE: During 1970-1994, 2,329 incident cases were registered at CCRP on the basis of histological and/or clinical information, excluding 30 cases reported only by death certificate. Histological or hematological diagnosis was available for 2,067 cases. Vital status was assessed through the offices of the town of residence. At the end of follow-up, 1,202 cases were alive, 1,084 dead and 43 were not traceable. Survival was measured for the major diagnostic groups using both univariate and multivariate statistics. RESULTS: The 5-yr survival rate for acute lymphoblastic leukemia (ALL) improved regularly from 24.7% in 1970-1974 to 81.1% in 1990-1994, for acute nonlymphoblastic leukemia (ANLL) from 0% to 38.1%, for non-Hodgkin lymphoma (NHL) from 25.2% to 67.7%, for tumors of the central nervous system (CNS) (all types) from 33.4% to 75.9% and for Ewing tumor from 0% to 90%. Focusing on survival by period of diagnosis, the highest 5-year survival rate was observed for children diagnosed during 1985-1989 for medulloblastoma, neuroblastoma (NB), retinoblastoma, Wilms tumor, osteosarcoma, and rhabdomyosarcoma and for children diagnosed in 1990-1994 for the remaining sites. The trend over time was statistically significant for ALL, ANLL, NHL, CNS tumors, NB, and osteosarcoma as well as for all malignancies together. CONCLUSIONS: Population-based survival studies are useful complements to clinical studies. Survival results in the present study are similar to those presented for other European countries and the United States. For most types of neoplasm (except CNS) survival probability appears to stabilize 5-10 years after diagnosis. PMID- 11260573 TI - Asymmetric salivary gland 123I-meta-iodobenzylguanidine uptake in a patient with cervical neuroblastoma and Horner syndrome. PMID- 11260574 TI - Molecular alterations in a case of bilateral adrenal neuroblastoma. PMID- 11260575 TI - Spontaneous nocturnal growth hormone secretion in children after medulloblastoma therapy. PMID- 11260576 TI - Successful management of neonatal choriocarcinoma. PMID- 11260577 TI - Rapid deterioration of a newborn with congenital spinal cord astrocytoma. PMID- 11260578 TI - Priapism as the first sign of a pelvic tumour in a two-and-a-half-year-old boy. PMID- 11260579 TI - Eyelid leukemia as a relapse sign of B-cell type acute lymphoblastic leukemia. PMID- 11260580 TI - Predicting success using individualized scheduled toileting for memory-impaired elders at home. AB - The purpose of this research was to evaluate the effectiveness of an individualized scheduled toileting (IST) program on incontinent, memory-impaired elders being cared for at home. Using a 2 x 2 mixed design analysis of variance (group by time), 118 patients were randomly assigned to experimental or control groups. Caregivers in the experimental group were taught the IST procedure. Urinary incontinence (UI) was measured at baseline and at 6 months. Weeklong voiding records were kept by caregivers and were used to calculate the percentage of times the incontinence occurred. UI significantly decreased in the experimental group, whereas in the control group it did not. The baseline cognitive ability, mobility, and consistency of implementing IST were entered into a discriminant function equation and significantly predicted patients who would improve with IST. Cognitive ability was the best predictor, with mobility also emerging as a meaningful predictor. Candidates for IST should be selected based on elders' cognitive ability and their ability to cooperate with toileting. Moderately cognitively impaired elders and ones able to cooperate with toileting protocols are prime candidates for IST. PMID- 11260581 TI - Postpartum smoking behaviors and immune response in mothers of term and preterm infants. AB - Infants exposed to secondhand smoke, especially preterm infants with a very low birth weight (VLBW), have an increased risk for developing health problems. Smoking has been associated with numerous health problems in mothers and may reduce immune functioning as well. The purposes of this study were to examine smoking in postpartum mothers of term and preterm infants and to examine the relationship between smoking and immune status. Peripheral blood was drawn on 142 women at four data-collection points and tested for cotinine, immune cell phenotypes, and immune functioning. Overall, 39% of the participants smoked in the postpartum period, but 49% of mothers who delivered preterm infants smoked compared to only 28% of mothers who delivered term infants. There was no difference in cotinine levels between the smokers in both groups of postpartum mothers, nor was smoking related to immune phenotypes or immune function. Given the documented health risks to the mother and infant and the significant number of women who continue to smoke in the postpartum period, it is imperative that health care providers continue to assess smoking status and provide smoking cessation counseling at every encounter. PMID- 11260582 TI - Mediating functions of maternal anxiety and participation in care on young children's posthospital adjustment. AB - The purpose of this study was to determine whether maternal anxiety and mothers' participation in their children's care during hospitalization mediated the effects of a child behavior informational intervention for mothers on their children's posthospital negative behavioral change. Participants were 49 mothers and their young children, ages 24-68 months, who were unexpectedly hospitalized with unplanned medical or surgical conditions. These participants were drawn from a larger study of the separate and combined effects of child behavior information and parent role information on the process and outcomes of maternal and child coping with unplanned hospitalization. Findings indicated that the effects of child behavior information on children's posthospital negative behavioral change were mediated by maternal anxiety and participation in their children's care during hospitalization. Results of this study provide support for targeting mothers with informational interventions in order to enhance outcomes in hospitalized children. PMID- 11260584 TI - Minority women with sexually transmitted diseases: sexual abuse and risk for pelvic inflammatory disease. AB - Mexican American and African American women (N = 617) with a sexually transmitted disease (STD) underwent a targeted physical exam and questioning regarding sexual abuse, current genitourinary symptomatology, and pelvic inflammatory disease (PID) risk behaviors to determine the relationship between sexual abuse and risk for PID. Sexually abused women (n = 194) reported higher PID risk behaviors, including earlier coitus, more sex partners, higher STD recurrence, and a tendency toward delayed health-seeking behavior. They also reported more severe genitourinary symptomatology, confirmed by physical exam, and presumptive diagnoses of PID. These characteristics identify sexually abused women at high risk for PID. Because of its considerable impact on risk for PID, assessment for sexual abuse is essential in clinical management of women with STD and for diagnosis of PID. PMID- 11260583 TI - Cigarette use in adolescents: the Cardiovascular Health in Children and Youth Study. AB - Tobacco is the leading cause of preventable death in the United States, and its use is increasing in adolescents. To determine the interventions needed to prevent the initiation of smoking, it is important to know the factors related to tobacco use by adolescents. In this study the following factors related to cigarette use were examined: age, gender, ethnicity, self-esteem, physical activity, parental smoking, and socioeconomic status. Participants were 1,207 youth completing a written survey for the Cardiovascular Health in Children and Youth Study (CHIC II). Participants ranged in age from 10 to 15 years, with a mean age of 12.2 years; 64.2% were White, 24.0% Black, 5.8% Hispanic, and 6.0% other races. White and Hispanic youth and youth of other races had significantly higher rates of smoking than did Black youth. Significant risk factors for smoking were: higher grade in school, White race, and for girls only, lower self esteem. In White youth those in the lowest socioeconomic status were most likely to be current and experimental smokers. Smoking was as common in girls as in boys at these ages. Multivariate analysis showed that neither physical activity nor parental smoking were significant predictors of smoking behaviors. These results suggest that smoking prevention programs for adolescents should specifically target White and Hispanic youth and those from families with low socioeconomic status. In addition, these interventions should include ways to increase self esteem in girls. PMID- 11260585 TI - Relationship of empathy to appraisal, depression, life satisfaction, and physical health in informal caregivers of older adults. AB - The relationship between empathy and caregiving appraisal and outcomes was examined among 140 informal caregivers of older adults. Caregivers with high cognitive empathy appraised the caregiving situation as less stressful and less threatening, were less depressed, and reported higher life satisfaction than did caregivers with low cognitive empathy. The caregivers' appraisal, along with educational levels and total household income, significantly predicted individual differences in caregiver depression, life satisfaction, and perceived physical health. Emotional empathy was negatively related to life satisfaction. There appeared to be distinct roles for emotional and cognitive empathy in informal caregiving outcomes. The study supported the important role of caregiving appraisal and resources in caregiving outcomes. PMID- 11260586 TI - Hydrotherapy in labor. AB - Maternal anxiety and pain prolong labor and contribute to fetal distress. Hydrotherapy during labor may promote relaxation and decrease pain without the risks caused by other treatments. In this pilot study the psychophysiological effects of hydrotherapy on maternal anxiety and pain during labor were examined. Using a randomized, pretest-posttest control group design with repeated measures, 18 term parturients were assigned to a control or an experimental group. Experimental subjects were placed in a tub of 37 degrees C water for 1 hr during early labor. The Wilcoxon two-sample test revealed statistically significant effects. At 15 min bathers' anxiety and pain scores were decreased compared to nonbathers. At 60 min bathers' pain scores were decreased compared to nonbathers. After 15 min of immersion, bathers had a significantly greater increase in plasma volume than nonbathers. No significant differences were found in urine catecholamines or maternal-fetal complications. The small sample limits conclusions, but the findings offer preliminary support for the therapeutic effects of bathing in labor for acute, short-term anxiety and pain reduction. PMID- 11260587 TI - Psychometric evaluation of the exercise self-efficacy scale among Korean adults with chronic diseases. AB - The purpose of this study was to assess the psychometric properties, appropriateness, and demographic response patterns of an exercise self-efficacy scale for Korean adults with chronic diseases. After assessment of face validity by an expert Korean panel, 249 Korean adults with chronic diseases, ages 18-79 years, were recruited from hospitals or health centers in five Korean cities and surrounding rural areas to complete the questionnaire. In a factor analysis the original 18-item exercise self-efficacy scale converged to one factor without rotation and to three subfactors with rotation: situational/interpersonal factor, competing demands factor, and internal feelings factor. Descriptive analysis showed that Korean adults with chronic diseases perceived they had relatively low exercise self-efficacy, with the situational/interpersonal factor as the lowest factor. Exercise self-efficacy was significantly correlated with gender, education, regular exercise, and frequency of exercise. The exercise self efficacy scale was shown to be a useful measure of exercise beliefs of Korean adults with chronic diseases. PMID- 11260589 TI - Seizing the moment: an opportune time to study the outcomes of interprofessional education and health care delivery. PMID- 11260590 TI - High-density microarrays for gene expression analysis. PMID- 11260591 TI - Hyperspectral imaging: a novel approach for microscopic analysis. AB - BACKGROUND: The usefulness of the light microscope has been dramatically enhanced by recent developments in hardware and software. However, current technologies lack the ability to capture and analyze a high-resolution image representing a broad diversity of spectral signatures in a single-pass view. We show that hyperspectral imaging offers such a technology. METHODS AND RESULTS We developed a prototype hyperspectral imaging microscope capable of collecting the complete emission spectrum from a microscope slide. A standard epifluorescence microscope was optically coupled to an imaging spectrograph, with output recorded by a CCD camera. Software was developed for image acquisition and computer display of resultant X--Y images with spectral information. Individual images were captured representing Y-wavelength planes, with the stage successively moved in the X direction, allowing an image cube to be constructed from the compilation of generated scan files. This prototype instrument was tested with samples relevant to cytogenetic, histologic, cell fusion, microarray scanning, and materials science applications. CONCLUSIONS: Hyperspectral imaging microscopy permits the capture and identification of different spectral signatures present in an optical field during a single-pass evaluation, including molecules with overlapping but distinct emission spectra. This instrument can reduce dependence on custom optical filters and, in future imaging applications, should facilitate the use of new fluorophores or the simultaneous use of similar fluorophores. PMID- 11260592 TI - Fluorescence lifetime imaging: multi-point calibration, minimum resolvable differences, and artifact suppression. AB - BACKGROUND: Frequency-domain fluorescence lifetime imaging microscopy (FLIM) is finding increasing use in the analysis of biological systems. However, the calibration, determination of resolvable lifetime differences, and evaluation of artifacts have not been extensively treated. We describe a multi-point method for calibrating a frequency-domain FLIM system, characterize the minimum detectable heterogeneity and intra- and inter-image lifetime differences, discuss the statistical treatment of FLIM data, and suggest methods for minimizing artifacts. METHODS: A set of solutions exhibiting single-component lifetimes suffice for accurately calibrating a reference material with a single-component lifetime, even in the absence of accurate data on the lifetimes of the individual solutions or the reference material. We used a set of rhodamine 6G solutions quenched with varying concentrations of iodide, leading to lifetimes of 0.5--4.0 ns, to calibrate a 1 microM reference solution of rhodamine 6G in water. RESULTS: We measured a value of 4.11 ns with an estimated absolute error of +/-0.05 ns for the rhodamine 6G reference solution. With 57.7 MHz modulation, the minimum detectable inter-image lifetime difference was 0.1--0.15 ns and the minimum detectable intra-image lifetime difference was 4--5 ps, allowing solutions differing in lifetime by 40 and 70 ps to be easily distinguished. The minimum detectable lifetime heterogeneity was 50--80 ps. Evaluation of replicate measurements of single solutions demonstrated that inter-image instrument errors exceeded those predicted from intra-image statistics by more than an order of magnitude. We also measured lifetimes and heterogeneity in 4 GFP variants (WTGFP, EGFP, S65T, and EYFP) with the technique. CONCLUSION: The multi-point calibration method is applicable to any system consisting of single-component lifetimes. Applying the method in our FLIM microscope allowed us to demonstrate a previously unreported degree of lifetime resolution in a FLIM microscope. Cytometry 43:248 260;2001. PMID- 11260593 TI - Velocity estimation of spots in three-dimensional confocal image sequences of living cells. AB - BACKGROUND: The analysis of three-dimensional (3D) motion is becoming increasingly important in life cell imaging. A simple description of sometimes complex patterns of movement in living cells gives insight in the underlying mechanisms governing these movements. METHODS: We evaluate a velocity estimation method based on intensity derivatives in spatial and temporal domain from 3D confocal images of living cells. Cells of the sample contain intense spots throughout the cell nucleus. In simulations, we model these spots as Gaussian intensity profiles which are constant in intensity and shape. To quantify the quality of the estimated velocity, we introduce a reliability measure. RESULTS: For constant linear velocity, the velocity estimation is unbiased. For accelerated motion paths or when a neighboring spot disturbs the intensity profile, the method results are biased. The influence of the point-spread function on the velocity estimation can be compensated for by introducing anisotropic derivative kernels. The insight gained in the simulations is confirmed by the results of the method applied on an image sequence of a living cell with fluorescently labeled chromatin. CONCLUSIONS: With the velocity estimation method, a tool for estimating 3D velocity fields is described which is successfully applied to a living cell sequence. With the estimated velocity fields, motion patterns can be observed, which are a useful starting point for the analysis of dynamic processes in living cells. PMID- 11260594 TI - Membrane-targeted green fluorescent protein reliably and uniquely marks cells through apoptotic death. AB - BACKGROUND: An understanding of the molecular processes that comprise the program of physiological cell death demands analytical techniques for the assessment of death events on the level of the individual cell, especially among transfectants and within heterogeneous populations. The utility of available transfection markers is limited by the variability of marker retention and discrimination as cells die. For example, soluble green fluorescent protein (GFP) leaks from dying cells and is not useful when fixation is required; conversely, transfected beta galactosidase can be visualized only after fixation and staining. METHODS: We have tested a GFP variant as a marker for the direct identification and visualization of transfected cells. We have explored the utility of this membrane targeted GFP, the genetic fusion of the enhanced GFP and the farnesylation sequence of p21(Ras) (EGFP-F), in a variety of cell death assays. RESULTS: EGFP-F is retained reliably in unfixed dying cells, permitting numerous events of the cell death process to be analyzed in real time in marked cells. Moreover, the cell rounding and shrinkage associated with the loss of adhesion during cell death result in a characteristic condensed EGFP-F signal. CONCLUSIONS: EGFP-F serves to identify transfectants consistently, independent of their ultimate fate. Cellular condensation of EGFP-F provides a specific and quantitative measure of physiological cell death. PMID- 11260596 TI - Evaluation of red blood cell lysing solutions in the study of neutrophil oxidative burst by the DCFH assay. AB - BACKGROUND: Neutrophil subpopulations with enhanced oxidative reactivity have been described in a number of clinical and in vitro settings. In the dichlorofluorescin (DCFH) oxidation assay, it is essential to maintain cellular viability and plasma membrane integrity through all stages of sample preparation. The process of erythrocyte lysing is crucial because a number of commercial lysing reagents can increase leukocyte membrane permeability. METHODS: We assessed viability [propidium iodide (PI) method], DCFH oxidation, and CD11b expression of resting or in vitro-stimulated neutrophils exposed to six different red cell lysing procedures. RESULTS: Formaldehyde-containing reagents (Optilyse B, FACS Lyse, and Erythrolyse) but not hypotonic shock or ammonium chloride (NH(4)Cl) solutions rendered 91.4--99.8% of resting neutrophils PI positive, with concomitant reductions in dichlorofluorescein (DCF) fluorescence, suggesting efflux of the fluorochrome. However, when stimulated with N-formyl-methionyl leucyl-phenylalanine or Yersinia enterocolitica and then treated with FACS Lyse or Erythrolyse, up to 69.9% of neutrophils remained PI negative and exhibited enhanced DCF fluorescence. CD11b expression of PI-positive and -negative neutrophils was comparable, suggesting that they were activated equally. CONCLUSIONS: FACS Lyse and Erythrolyse can modify neutrophil plasma membrane integrity, whereas hypotonic shock and NH(4)Cl solutions retain cellular viability and are lysing methods of choice in evaluation of neutrophil respiratory burst by DCFH oxidation assay. PMID- 11260595 TI - Identification of Echovirus 1 and coxsackievirus A9 receptor molecules via a novel flow cytometric quantification method. AB - BACKGROUND: Virus-receptor binding is an essential step in every virus infectious process. Many viruses employ more than one receptor molecule or even receptor complexes for attachment. In this study, we investigate the binding of Echovirus 1 (Echo1) and Coxsackievirus A9 (CAV-9) on cell surface molecules. CAV-9 has been reported to utilize integrin alpha v beta(3) in binding to cells, whereas Echo1 has been known to utilize integrin alpha 2 beta(1). METHODS AND RESULTS We directly test whether the presence of these molecules alone was sufficient for virus binding. We devised a novel flow cytometric binding assay that enables us to quantify virus particles bound on host cells and to further determine the extent to which viruses utilize specific receptors. CONCLUSIONS: By quantifying virus particles and possible receptor molecules, we found that Echo1 utilizes mainly integrin alpha 2 beta(1). CAV-9 utilizes integrin alpha v beta(3) to a much lesser extent (40%), indicating that CAV-9 also utilizes other receptor(s). PMID- 11260597 TI - Delineation among eight major hematopoietic subsets in murine bone marrow using a two-color flow cytometric technique. AB - BACKGROUND: Many methods have been developed specifically for identifying hematopoietic progenitor cells in murine bone marrow, but few methods allow rapid identification of multiple bone marrow populations. We describe a new, simple method for identifying simultaneously eight populations in murine bone marrow with two-color flow cytometry and phenotypically define these populations. METHODS: Bone marrow was stained with anti-Ly-6C and anti-B220 (CD45R) in one fluorochrome and wheat germ agglutinin (WGA) in another fluorochrome. The eight populations identified in this way were defined further primarily by four-color flow cytometry. RESULTS: Six of the eight populations were characterized phenotypically as containing erythroid, granulocytic, mast, early B, mature B, and stem cell populations. Two additional populations with phenotypic characteristics of partially differentiated precursor cells also were identified. One population was Ly-6C/B220+ and WGA-. It also expressed markers associated with early B, T, and/or dendritic cell differentiation. The second population was Ly-6C(hi)WGA(hi)Mac-1+ and was negative for numerous other lineage-specific and precursor markers. Its morphology suggested monocytic differentiative potential. CONCLUSIONS: A two-color flow cytometric assay profiles six bone marrow populations with identifiable phenotypes and two additional unique, putative hematopoietic precursor populations. PMID- 11260598 TI - Laser scanning cytometry: a novel method for the detection of platelet- endothelial cell adhesion. AB - BACKGROUND: Adherence of platelets to endothelial cells may be a significant event in the development of vascular thrombosis. Existing models, which examine platelet-endothelial cell interactions, compromise endothelial cell integrity or use radioactivity to identify platelets that adhere to endothelial cells. We report a novel method for in vitro detection of platelet-endothelial cell adhesion that allows endothelial cells to remain as an intact monolayer and for visualization of individual platelets. METHODS: Fluorescently labeled platelets were incubated with a confluent monolayer of endothelial cells. Laser scanning cytometry (LSC) identified platelets bound to endothelial cells based on their fluorescent signals. RESULTS: LSC detection of platelets reliably reproduced well described findings of thrombin-induced platelet-endothelial cell adhesion. Results demonstrating reduced adhesion with a glycoprotein IIb-IIIa-specific blocking monoclonal antibody confirmed the specificity of the LSC detection of platelet-endothelial cell adhesion. CONCLUSIONS: LSC is a novel method for detecting platelet--endothelial cell adhesion. Its advantages over other methods are: (a) endothelial cells remain undisturbed and adherent throughout; (b) the ability to detect individual bound platelets and subpopulations; (c) the ability to store images and slides and then relocate, revisualize, and reanalyze individual cells or cell populations of interest; and (d) no radioactivity. PMID- 11260599 TI - A new approach to determine the genetic diversity of viable and active bacteria in aquatic ecosystems. AB - BACKGROUND: Discrimination among viable, active, and inactive cells in aquatic ecosystems is of great importance to understand which species participate in microbial processes. In this study, a new approach combining flow cytometry (FCM), cell sorting, and molecular analyses was developed to compare the diversity of viable cells determined by different methods with the diversity of total cells and active cells. METHODS: Total bacteria were determined by SYBR-II staining. Viable bacteria were determined in water samples from different sites by plate count techniques and by the direct viable count (DVC) method. Substrate responsive cells (i.e., DVC(+) cells) were distinguished from nonresponsive cells (i.e., DVC(-) cells) by FCM and sorted. The genetic diversity of the sorted cell fraction was compared with the diversity of the total microbial community and with that of the culturable cell fraction by denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR)-amplified 16S rDNA fragments. The same approach was applied to a seawater sample enriched with nutrients. In this case, actively respiring cells (CTC+) were also enumerated by FCM, sorted, and analyzed by DGGE. RESULTS: The diversity of viable cells varied depending on the methods (traditional culture or DVC) used for viability assessment. Some phylotypes detected in the fraction of viable cells were not detectable at the community level (from total DNA). Similar results were found for actively respiring cells. Inversely, some phylotypes found at the community level were not found in viable and active cell-sorted fractions. It suggests that diversity determined at the community level includes nonactive and nonviable cells. CONCLUSION: This new approach allows investigation of the genetic diversity of viable and active cells in aquatic ecosystems. The diversity determined from sorted cells provides relevant ecological information and uncultured organisms can also be detected. New investigations in the field of microbial ecology such as the identification of species able to maintain cellular activity under environmental changes or in the presence of toxic compounds are now possible. PMID- 11260600 TI - Knowledge of Down syndrome in pregnant women from different ethnic groups. AB - The uptake of any screening test is influenced by knowledge of the condition being screened for. In the present study, the knowledge and the source of knowledge of women offered antenatal screening for Down syndrome (DS) was assessed by means of a self-administered questionnaire. The questionnaire was administered to 300 consecutive women booking for antenatal care, of the 245 (82%) women who completed and returned the questionnaire, 117 (48%) were Caucasian, 85 (35%) were Asian born outside the UK, 32 (13%) were Asian born in the UK and ten (4%) belonged to other categories. Only 30% of the cohort had a good understanding of the condition. Racial groups other than Caucasian had a poorer understanding of DS. The factors which affected knowledge of DS included quality of spoken English, knowing an affected child, parity and religion. The most significant factor affecting acceptance of screening was the woman's knowledge of DS. The source of information for the condition varied widely: 42% from a general practitioners (GP), 24% from the hospital and 16% from midwives. The proportion with good knowledge was similar in those women whose source of information was the GP (45%) and the midwife (41%). These proportions were, however, higher (though not significantly) when the source of information was from magazines and newspapers (67%) and from friends (53%). Uptake of the screening test was best in those with good knowledge (53%) compared to those with poor knowledge (23%) (p<0.02). Between 28% and 66% (depending on the ethnic group) of women had a screening blood test "allegedly" without knowing why it had been performed. In order to improve uptake of the screening test for DS there is need for better education and counselling of women attending for antenatal care. PMID- 11260601 TI - Prenatal diagnosis on fetal cells from maternal blood: practical comparative evaluation of the first and second trimesters. AB - Objectives- Several attempts have been made to determine the gestational period in which the maximum number of fetal cells can be found in maternal blood and consequently which is the best week in which to perform a reliable non-invasive prenatal diagnosis. Most studies conclude that the number of nucleated red blood cells (NRBC) increases in line with gestation, but the number of cells that are fetal in origin (FNRBC) decreases in the third trimester. The aim of the present study was to make a practical comparative evaluation of the first and second trimesters to ascertain the period in which a greater number of FNRBC can be found of the total number of NRBC identified. Methods- Double density gradient and a posterior positive selection (CD71) by magnetic activated cell sorting (MACS) were employed. In the final fraction, erythroblasts were identified using Kleihauer staining and were studied using the fluorescence in situ hybridization (FISH) interphasic technique. Results- There was a significant difference (p<0.05) between the mean number of FNRBC found in the first and second trimesters. Conclusions- The number of FNRBC increases from the first to the second trimester. It appears that the optimum week in which to perform a reliable non-invasive prenatal diagnosis is around the 15th week. PMID- 11260602 TI - Interphase FISH with chromosome-specific protelomere probes for rapid prenatal diagnosis in a reciprocal translocation carrier. AB - Interphase fluorescence in situ hybridization (FISH) analysis has become an accepted practice for rapid preliminary analysis of chromosome aneuploidy from direct amniocyte preparations. The use of dual-color interphase FISH analysis with chromosome-specific protelomere probes for the rapid exclusion of chromosomally unbalanced segregants in the pregnancy of a reciprocal translocation carrier is reported. Amniocentesis was performed at 16 weeks gestation on the carrier of a t(5;14)(p14.2;p13), who was ascertained after the birth of a son with the der(5) chromosome. Interphase FISH analysis with probes for 5pter, 5qter and 14qter showed two signals for each, consistent with alternate segregation of the maternal translocation. Subsequent metaphase analysis confirmed a 46,XY,t(5;14)(p14.2;p13)mat karyotype in the fetus. This case illustrates the utility of interphase FISH analysis with protelomere probes for rapid prenatal diagnosis in cases of parental reciprocal translocation. PMID- 11260603 TI - The application of fetal magnetocardiography (FMCG) to investigate fetal arrhythmias and congenital heart defects (CHD). AB - OBJECTIVES: Fetal magnetocardiography (FMCG), a new non-invasive diagnostic tool in the analysis of the electrophysiological changes of the heart, was selectively applied in cases of fetal arrhythmias and congenital heart defect (CHD) to demonstrate its value for diagnosis and prenatal management. METHODS: The FMCG was analysed and compared to the postnatal ECG in four cases of fetal arrhythmia [supraventricular tachycardia (two cases), complex tachy-/bradycardia (one case), ventricular extrasystoles (one case)] and a case of right heart hypoplasia diagnosed by established methods prior to investigation. RESULTS: A Wolf Parkinson-White (WPW) syndrome was diagnosed by its characteristic features and the appropriate transplacental therapy chosen. The types of arrhythmia could be characterised in accordance with postnatal ECG findings and irregular conduction was demonstrated in association with a CHD. CONCLUSIONS: The use of the FMCG provides additional information to the common diagnostic tools that influence therapeutic decisions and thus contributes to optimal pre- and postnatal management. PMID- 11260604 TI - Prenatal diagnosis of malignant osteopetrosis in Bedouin families by linkage analysis. AB - Autosomal recessive malignant osteopetrosis (MOP) is a lethal disease, unless bone marrow is successfully transplanted. Yet a donor may not always be available, and even when there is one transplantation results are far from optimal. The difficulty in obtaining conclusive results by sonographic and X-ray evaluation of the fetus makes prenatal molecular diagnosis highly desirable. Subsequent to the chromosomal localization of the MOP gene in Arab-Bedouin families from the Negev region in Israel, linkage analysis was used for the prenatal diagnosis of this disease in Bedouin families at risk. Twelve cases were diagnosed, three fetuses were found to be affected, and one of the pregnancies was terminated. The other two pregnancies continued to term and the diagnosis of osteopetrosis was confirmed by X-ray immediately after birth. This is the first report on prenatal diagnosis of autosomal recessive osteopetrosis by linkage analysis. PMID- 11260605 TI - Optimization of nucleated red blood cell (NRBC) recovery from maternal blood collected using both layers of a double density gradient. AB - The isolation of fetal nucleated red blood cells (NRBC) from maternal blood represents a promising approach to non-invasive prenatal diagnosis. However, the number of fetal NRBC in maternal circulation is quite low and therefore difficult to isolate. An enrichment procedure in which both layers from a double density 1.077/1.107 g/ml gradient are collected was optimized, followed by MACS selection using non-commercial monoclonal antibodies. The influence of the delay in processing maternal blood on the NRBC distribution in both interfaces of the gradient was also studied in cord blood and peripheral maternal blood samples. A significant increase in the number of NRBC isolated from maternal blood was achieved by collecting both layers of the double density gradient compared with the previous protocol in which only the lower layer was recovered. Cord blood samples showed significant differences in the number of NRBC recovered when processed at 24 instead of within 3 h. This effect was also observed in the number of NRBC collected only from the upper layer of peripheral maternal blood samples. Therefore, in order to minimize the target cell losses, it is advisable to process the maternal blood samples as soon as possible. PMID- 11260606 TI - Preliminary data on an association between blood groups and serum markers used for the so-called "triple screening": free oestriol MoM values are decreased in rhesus-negative (Rh-) women. AB - The serum markers human chorionic gonadotrophin (hCG), alpha-fetoprotein (AFP) and unconjugated oestriol (uE3) in 606 rhesus-negative (Rh-) women have been compared with 18 960 controls. There were no significant differences in the distribution of hCG and AFP between these two cohorts. However the uE3 values in Rh- women were significantly lowered (median: 0.85 MoM). PMID- 11260607 TI - Assessment of fetal status in multiple gestation pregnancies using interphase FISH. AB - The use of fluorescence in situ hybridization (FISH) for women with multiple gestation pregnancies has been evaluated. Women were referred for chromosome analysis because of advanced maternal age, abnormal ultrasound findings or a positive family history and/or prior to fetal reduction. FISH was successfully applied to all specimens obtained by amniocentesis or chorionic villus sampling (CVS). Based on FISH results, fetal-fetal contamination of specimens following CVS was 11.5% in twin pregnancies and 16% in triplet or higher multiples. FISH detected trisomy 21 in three cases with no false negatives or positives. Whereas FISH may provide rapid and useful assessment of fetal status in decision-making regarding fetal reduction, the present study also highlighted the obstetrical difficulty of ensuring a sample representative of each fetus following CVS in addition to the possibility of not identifying clinically significant chromosome aberrations using currently available FISH probes. PMID- 11260608 TI - Prenatal diagnosis of oculocutaneous albinism two mutations located at the same allele. AB - A pregnant woman accepted amniocentesis on account of the previous birth of type 1 oculocutaneous albinism (OCA1). PCR revealed that the fetus had two mutations (862delTT, Arg 299His). The father had one missense mutation (Arg 299Ser) and the mother had the same mutations as the fetus. Two mutations of the fetus located at the same allele were suspected. Postpartal follow-up confirmed his carrier status. For recessive disorders, faced with a fetus with two mutations, the importance of performing segregation analysis of mutation on both parents is emphasized. This could exclude two mutations located at the same allele and prevent the unnecessary termination of a fetus with carrier status. PMID- 11260609 TI - Prenatal diagnosis of hereditary spastic paraplegia. AB - Hereditary spastic paraplegia (HSP) is a degenerative neurologic disorder that causes progressive, often severe, spastic weakness in the legs. Autosomal dominant HSP is a highly penetrant, genetically heterogeneous disorder with loci present on chromosomes 2p21-24, 2q24-34, 8q23-24, 10q23.3-24, 12q13, 14q12-23, 15q11-14 and 19q13.1. We identified a large HSP kindred in which the disorder was tightly linked to chromosome 14q12-23. We tested chorionic villus DNA samples of two at-risk fetuses for inheritance of microsatellite polymorphisms flanking and within this locus that segregated with the disease in this family. Whereas samples from the first fetus showed inheritance of a haplotype segregating with the disease allele (indicating high risk of developing HSP), samples from the second fetus showed inheritance of a haplotype segregating with the normal allele (indicating low risk of developing HSP). This is the first report of prenatal testing for HSP. Published in 2001 by John Wiley & Sons, Ltd. PMID- 11260611 TI - Preimplantation genetic diagnosis for sickle-cell anemia and for beta thalassemia. AB - We developed single-cell polymerase chain reaction (PCR) assays for preimplantation genetic diagnosis (PGD) in couples carrying mutations in the beta globin gene. With PGD the genetic status of an embryo obtained after intracytoplasmic sperm injection (ICSI) is determined by PCR analysis in single blastomeres, allowing only healthy embryos to be transferred to the uterus. We carried out nine PGD cycles using fluorescent PCR for two couples in whom the partners carried sickle-cell trait. Both couples achieved pregnancies, one of which was spontaneously aborted. We have developed two beta-thalassemia PGD protocols: one for the analysis of the 25-26delAA and the IVS2+1G>A mutation, and the other for the simultaneous detection of the IVS1+6T>C and the IVS1+110G>A mutations. For the second protocol, both non-labelled PCR and later fluorescent PCR were used. Both protocols were applied in clinical cycles (two non-labelled PCR cycles and one fluorescent PCR cycle) for two couples. The patient with the fluorescent PCR-PGD cycle became pregnant. Overall, the three fluorescent PCR assays were accurate and reliable with amplification efficiencies of minimum 93% and allele dropout (ADO) rates between 0 and 12%. PMID- 11260610 TI - Dehydro-oestriol and dehydropregnanetriol are candidate analytes for prenatal diagnosis of Smith-Lemli-Opitz syndrome. AB - Gas chromatographic/mass spectrometric (GC/MS) analysis of maternal urine and serum steroids from 13 pregnancies at 25% risk for Smith-Lemli-Opitz syndrome (SLOS) was undertaken. All patients were between 12 and 31 weeks' gestational age. From dehydrocholesterol/cholesterol ratios determined in amniotic fluid and chorionic villus cells, five patients were shown to carry SLOS affected fetuses and eight patients were negative for the condition. Because it had previously been shown that dehydro-oestriol and dehydropregnanetriol were novel steroids produced in SLOS, these compounds were measured in the serum and urine samples of the 13 mothers. All five urine samples from SLOS affected pregnancies had high levels of both dehydrosteroid metabolites, which were below the detection limit in the non-affected pregnancies. The ratios of dehydro-oestriol/oestriol (DHE(3)/E(3)) were between 0.073 and 1.42 for the affected patients and less than 0.01 for unaffected patients. Corresponding values for dehydropregnanetriol/pregnanetriol (DHPT/PT) were 0.037-1.02 for affected and less than 0.01 for unaffected. In the positive serum sample available for analysis, the DHE(3)/E(3) ratio was 0.20 [unaffected (n=5), <0.014]. It is proposed that the measurement of DHE(3) and DHPT in maternal urine and serum may allow non-invasive antenatal diagnosis of SLOS. PMID- 11260612 TI - A successful strategy for preimplantation genetic diagnosis of myotonic dystrophy using multiplex fluorescent PCR. AB - The most common form of inherited muscular dystrophy in adults is myotonic dystrophy (DM), an autosomal-dominant disease caused by the expansion of an unstable CTG repeat sequence in the 3' untranslated region of the myotonin protein kinase (DMPK) gene. Expanded (mutant) CTG repeat sequences are refractory to conventional PCR, but alleles with a number of repeats within the normal range can be readily amplified and detected. Preimplantation genetic diagnosis (PGD) of DM has been successfully applied. However, a misdiagnosis using the reported protocol was recently documented. Two new PGD protocols for DM have been developed which utilise multiplex fluorescent PCR. Ideally a linked polymorphic marker, APOC2, is amplified in addition to the normal DMPK alleles, thus providing a back-up diagnostic result. However, the two couples reported in the present study were not fully informative at the APOC2 locus and so an unlinked short tandem repeat (STR) marker, D21S1414, was substituted. The highly polymorphic nature of the D21S1414, DMPK and APOC2 loci means that a very simple genetic fingerprint can be generated by analyses of these loci. This allows most DNA contaminants to be detected. Contamination is a significant problem for PGD and is the primary reason for the inclusion of D21S1414 and APOC2 in this protocol. This paper reports the first clinical experience and pregnancies following PGD for DM using a multiplex fluorescent PCR protocol. PMID- 11260613 TI - Prenatal detection of an inverted X chromosome in a male fetus. PMID- 11260614 TI - Digynic triploidy: possible mechanisms. PMID- 11260616 TI - The total test approach to standardization of immunohistochemistry. PMID- 11260619 TI - Alveolar hemorrhage and renal microangiopathy in systemic lupus erythematosus. AB - CONTEXT: Acute alveolar hemorrhage in systemic lupus erythematosus usually occurs as a pulmonary-renal syndrome. In most cases, the lungs show "bland" alveolar hemorrhage with little or no inflammation. Whether this alveolar injury is similar to the better-defined noninflammatory renal lupus vasculopathy is unresolved. OBJECTIVES: To investigate the relationships and the mechanisms of small vascular injury in the lung and kidney of 2 lupus patients who died of diffuse AH. METHODS: We investigated the relationship of AH to immune complex deposition in the lungs of 6 patients with systemic lupus erythematosus and correlated the findings with glomerular and vascular disease in the kidney. Lung and kidney were studied by light, immunofluorescence, and/or electron microscopy; apoptosis was investigated using in situ nick-end labeling. RESULTS: The clinical course of 2 patients was complicated by alveolar hemorrhage, and the lungs of these patients revealed alveolar wall immune complex deposits and bland alveolar hemorrhage. These 2 patients had World Health Organization class IV lupus nephritis and renal arterioles involved by a noninflammatory lupus vasculopathy. Apoptosis was identified in the lupus microangiopathy and in alveolar walls within areas of alveolar hemorrhage. Alveolar wall immune complex deposits were not found in 4 patients who had a lupus glomerulonephritis but did not have renal lupus vasculopathy. Apoptosis was not seen in renal arterioles or lungs of these 4 cases, except in areas of diffuse alveolar damage or herpesvirus pneumonia. CONCLUSIONS: Our findings indicate that alveolar hemorrhage in systemic lupus erythematosus, characterized by bland alveolar wall changes, is pathogenetically similar to the lupus microangiopathy of the kidney. In both lung and kidney, the pathogenesis of the microvascular injury appears to be related to immune complex deposition and the induction of apoptosis. PMID- 11260620 TI - Accuracy of fine-needle aspiration of thyroid. AB - CONTEXT: Fine-needle aspiration has become an accepted and cost-effective procedure for rapid diagnosis of thyroid lesions. The routine use of fine-needle aspiration has reduced the rate of unnecessary surgery for thyroid nodules. OBJECTIVES: To determine the accuracy of fine-needle aspiration biopsy diagnosis and to discuss the possible pitfalls. Design, Setting, and Participants.-Reports of 6226 fine-needle aspiration biopsies of the thyroid performed during a period of 16 years (1982-1998) were reviewed. Computerized reports of the fine-needle aspiration biopsies were sent to the physicians who performed the procedures, and clinical follow-up information regarding the patients was requested. Twenty-four clinicians participated in the study. Histologic diagnoses were available for 354 cases. The cytopathologic diagnoses were correlated with the histologic findings or clinical outcomes. RESULTS: The cytologic diagnoses were as follows: 210 (3.4%) malignant, 450 (7.2%) suspicious, 3731 (60%) benign, and 1845 (29.5%) unsatisfactory. Most of the cases with negative or unsatisfactory aspirates were followed clinically or by repeat fine-needle aspiration. We identified 11 false negative and 7 false-positive diagnoses. For aspirates considered sufficient for diagnosis, the sensitivity and specificity levels were 93% and 96%, respectively. CONCLUSIONS: Fine-needle aspiration of the thyroid gland is highly accurate and has a low rate of false-negative and false-positive diagnoses. The major diagnostic problems are caused by diagnosis using a marginally adequate specimen, diagnosis of malignancy based on just 1 or 2 atypical cytologic features, or overlapping cytologic features of follicular neoplasm with those of follicular variant of papillary carcinoma. PMID- 11260621 TI - Intraneuronal abeta-amyloid precedes development of amyloid plaques in Down syndrome. AB - CONTEXT: Down syndrome patients who live to middle age invariably develop the neuropathologic features of Alzheimer disease, providing a unique situation in which to study the early and sequential development of these changes. OBJECTIVE: To study the development of amyloid deposits, senile plaques, astrocytic and microglial reactions, and neurofibrillary tangles in the brains of young individuals (<30 years of age) with Down syndrome. METHODS: Histologic and immunocytochemical study of a series of autopsy brains (n = 14, from subjects aged 11 months to 56 years, with 9 subjects <30 years) examined at the Office of the Chief Medical Examiner of the State of Maryland and The Johns Hopkins Hospital. RESULTS: The principal observations included the presence of intraneuronal Abeta immunostaining in the hippocampus and cerebral cortex of very young Down syndrome patients (preceding the extracellular deposition of Abeta) and the formation of senile plaques and neurofibrillary tangles. CONCLUSIONS: We propose the following sequence of events in the development of neuropathologic changes of Alzheimer disease in Down syndrome: (1) intracellular accumulation of Abeta in neurons and astrocytes, (2) deposition of extracellular Abeta and formation of diffuse plaques, and (3) development of neuritic plaques and neurofibrillary tangles with activation of microglial cells. PMID- 11260622 TI - Molecular resistance testing of Helicobacter pylori in gastric biopsies. AB - OBJECTIVE: To evaluate simultaneous diagnosis of infection and molecular resistance testing of Helicobacter pylori. METHODS: Gastric biopsies were obtained from 26 rapid urease-positive and 51 rapid urease-negative test kits used to diagnose H pylori infection. Following glass bead-assisted DNA isolation, amplification of H pylori 16S ribosomal DNA (rDNA), glmM, and 23S rDNA target genes was performed. RESULTS: Helicobacter pylori DNA was successfully amplified from 100% (26/26) of urease-positive and 3.9% (2/51) of urease-negative gastric biopsies. Subsequent restriction enzyme-mediated digestion of 23S rDNA amplification products revealed that 17% (4/24) of urease-positive and H pylori DNA-positive biopsy specimens contained point mutations (A2142G or A2143G) associated with clarithromycin resistance. Helicobacter pylori DNA from gastric biopsies was successfully amplified 8 weeks following rapid urease testing. CONCLUSION: Helicobacter pylori genotyping may be used to detect macrolide resistant H pylori in individuals prior to initiation of therapy or in patients refractory to anti-H pylori therapy. Two urease-negative specimens yielded Helicobacter DNA distinct from that of H pylori and indicated the need for further investigations of Helicobacter species present in the human stomach. PMID- 11260623 TI - Carcinomas of sweat glands: report of 60 cases. AB - CONTEXT: Several aspects of sweat gland carcinomas (incidence, classification, diagnosis, and behavior) have not been definitively clarified and need to be studied further. OBJECTIVE: The clinicopathologic findings of a large series of sweat gland carcinomas, collected during a period of 15 years, are presented. METHODS: Sixty sweat gland carcinomas (41 porocarcinomas, 3 syringomatous carcinomas, 8 ductal carcinomas, 5 adenoid cystic carcinomas, and 3 mucinous carcinomas) were analyzed histologically and immunohistochemically. RESULTS: Porocarcinomas were composed of eosinophilic and clear atypical cells arranged in solid-cystic lobular masses. These tumors were divided into 2 subgroups: horizontal porocarcinomas, showing a prominent intraepidermal component, and nodular porocarcinomas, which demonstrated predominant nodular growth. Syringomatous carcinomas presented keratinizing and nonkeratinizing cysts, dilated tubules (sometimes with a "tadpole" appearance), small neoplastic ducts, solid islands, and cellular cords. Ductal carcinomas were characterized by a prominent formation of tubules, solid islands, and cellular cords. Adenoid cystic carcinomas presented a characteristic pattern, showing basaloid monomorphous cells with moderately atypical nuclei, arranged in cribriform or solid islands and in tubular structures. Mucinous carcinomas were composed of moderately atypical cells with eosinophilic vacuolated cytoplasm, forming solid and cystic islands floating in large mucin pools. Immunohistochemically, cytokeratin was found in neoplastic cells in all cases, carcinoembryonic antigen was detected in 73% of cases, and actin-positive (myoepithelial) cells were not found. CONCLUSIONS: Although numerous studies have been published in recent years, the histologic features, histogenesis, and classification of sweat gland carcinomas still remain controversial and need to be clarified by further studies. PMID- 11260624 TI - Gynecomastia-like changes of the female breast. AB - OBJECTIVES: Gynecomastia-like changes of the female breast are only sparsely reported and are not well defined in the literature to our knowledge. Our objectives were to determine the incidence, clinical presentation, mammographic findings, and the medical background of patients with these changes. DESIGN: Two thousand seven hundred nine female breast surgical cases from 1995 to 1999 were searched by SNOMED. Three observers further reviewed all cases with gynecomastia like changes. Strict criteria were developed and cases that fulfilled the criteria were analyzed further. RESULTS: We found the incidence of female gynecomastia-like changes to be 0.15% (4/2709) of all female breast lesions, which represents an underestimation. Patients were usually young and had an average age of 32 years. The usual clinical presentation was a palpable mass with a size ranging from about 3.5 x 2 x 2 cm to 5 x 4 x 2.5 cm. Mammography showed either negative findings or a nonspecific density. Gross examination of these specimens revealed no distinct lesions. Histologically, the lesions consisted of ductal hyperplasia with periductal stromal fibrosis or edema. They were associated with fibrocystic changes in the adjacent breast. The patients had no significant medical history. CONCLUSION: We propose that the gynecomastia-like change is a specific benign entity within the spectrum of benign fibrocystic changes and that it usually occurs in young patients. PMID- 11260625 TI - Cerebrospinal fluid abeta42, tau, and f2-isoprostane concentrations in patients with Alzheimer disease, other dementias, and in age-matched controls. AB - OBJECTIVE: To test the hypothesis that quantification of cerebrospinal fluid (CSF) F(2)-isoprostanes (F(2)-IsoPs), in vivo biomarkers of free radical damage, along with CSF Abeta(42) and tau levels improves laboratory diagnostic accuracy for Alzheimer disease (AD). PARTICIPANTS: Patients with probable AD (n = 19), dementias other than AD (n = 8), and age-matched controls (n = 10). MAIN OUTCOME MEASURES: Cerebrospinal fluid concentrations of Abeta(42) and tau were determined by a commercially available test (Athena Diagnostics, Worcester, Mass). Cerebrospinal fluid F(2)-IsoP levels were quantified by gas chromatography/mass spectrometry. RESULTS: Individuals were classified as AD or non-AD by a published method using CSF Abeta(42) and tau levels (95% sensitivity, 50% specificity), by CSF F(2)-IsoP levels greater than 25 pg/mL and Abeta(42) concentrations less than 1125 pg/mL (90% sensitivity, 83% specificity), and by combined analysis using CSF F(2)-IsoP, Abeta(42), and tau levels (84% sensitivity, 89% specificity). CONCLUSION: Cerebrospinal fluid F(2)-IsoP quantification may enhance the accuracy of the laboratory diagnosis of AD. PMID- 11260626 TI - Large cell variants of CD5+, CD23- B-cell lymphoma/leukemia. AB - CONTEXT: Mantle cell lymphoma (MCL), and its leukemic phase, constitute a well studied hematologic malignancy with known overall survival, prognostic indicators, morphologic findings at diagnosis and in bone marrow, and known incidence of the bcl-1 immunoglobulin gene rearrangement. Large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype (CD5+, CD23-), including but not limited to blastic MCL, prolymphocytoid MCL, blastic mantle cell leukemia, and prolymphocytic mantle cell leukemia, are not as well characterized. Although blastic MCL is known to be associated with a shorter overall survival than conventional MCL, the large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype have not been described as fully as conventional MCL. OBJECTIVE: The purpose of the present study was to describe the large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype. DESIGN: Nineteen cases of large cell variants of CD5+, CD23- B cell lymphoma/leukemia are reviewed and described in regard to morphology, bone marrow morphological findings, Cyclin D1 immunostaining, and bcl-1 analysis. Clinical data were not available owing to the varied clinical sources of the specimens. SETTING: Tertiary-care academic institution. RESULTS: Lymph node involvement in blastic CD5+, CD23- B-cell lymphoma was diffuse (100%) with a nodular component (33%) or focal mantle zone pattern (10%). Bone marrow involvement in blastic CD5+, CD23- B-cell lymphoma was seen in only 27% of cases and was composed predominantly of small, slightly irregular lymphocytes. Cyclin D1 was demonstrated in 60% of the 15 cases analyzed and more sensitive in B5 fixed tissue. Bcl-1 (performed in 5 cases) was not detected in the 4 cases of blastic CD5+, CD23- B-cell lymphoma analyzed and was detected in the case of the prolymphocytoid MCL. Cyclin D1 was demonstrated in all 4 bcl-1 negative cases and was negative in the bcl-1 positive prolymphocytoid MCL. CONCLUSION: Careful analysis of clinical data, morphology, immunophenotype, Cyclin D1 expression, and molecular analysis are required to differentiate the unusual large cell variants of MCL from other processes. PMID- 11260627 TI - Adult polyglucosan body disease. AB - We describe a case of adult polyglucosan body disease with characteristic clinical symptoms of peripheral neuropathy, upper motor neuron signs, and bowel and bladder dysfunction. Sural nerve biopsy revealed diagnostic intra-axonal polyglucosan bodies. On electron microscopic examination, the inclusions were located mainly within myelinated nerve fibers and consisted of branched filaments that were 6 to 8 nm wide. The diagnosis of adult polyglucosan body disease was confirmed by a skin biopsy from the axilla showing similar inclusions in myoepithelial cells of apocrine glands. This report provides additional evidence that skin biopsy, to date advocated by a single case report only, may be a less invasive and simpler diagnostic alternative to sural nerve or brain biopsies. PMID- 11260628 TI - Transfusion-related acute lung injury secondary to biologically active mediators. AB - Transfusion-related acute lung injury is seen following the transfusion of blood components. The reported incidence is approximately 1 in 2000 transfusions. Clinically, it is similar to adult respiratory distress syndrome. The pathophysiology is unclear but has been attributed to HLA antibodies, granulocyte antibodies, and more recently to biologically active mediators in stored blood components. We report a case with laboratory evidence that supports the role of biologically active mediators in the pathogenesis of transfusion-related acute lung injury. To our knowledge, the case reported here is the first to use lipid extractions of patient samples to determine that lipid-priming activity was present at the time transfusion-related acute lung injury was identified clinically. PMID- 11260629 TI - Intrasellar pituicytoma in a patient with other endocrine neoplasms. AB - Considered a neoplasm of pituicytes, pituicytoma is a rare and distinct type of glioma that arises in the suprasellar space and within the sella turcica. Only 12 previously reported cases of pituicytoma are documented in the literature. We report an intrasellar pituicytoma in a 66-year-old man presenting with symptoms and radiologic appearance indistinguishable from a nonfunctional pituitary adenoma. The patient also had a medical history significant for parathyroid adenomas and follicular carcinoma of the thyroid. The intrasellar tumor had morphologic features of a pituicytoma, with interlacing fascicles and a storiform pattern much like a benign fibrous histiocytoma. Immunoreactivity for S100 was strong, but the tumor lacked intercellular collagen type IV. The differential diagnosis of a low-grade spindle cell lesion of the sellar space is discussed, and the literature is reviewed. A summary of the clinical and pathologic features of this case, as well as the 12 previously reported cases, is presented. PMID- 11260630 TI - Cutaneous angiosarcoma complicating morbid obesity. AB - Herein, we report a case of cutaneous angiosarcoma in a 35-year-old, morbidly obese woman. The tumor arose in the most dependent portion of the lower abdominal panniculus and showed typical changes of chronic lymphedema. The patient underwent a radical resection of her lower abdominal wall panniculus, which showed a multicentric, high-grade angiosarcoma with bilateral superficial inguinal lymph node metastases. Histologically, conventional vasoformative areas were admixed with poorly differentiated sheets of spindle and epithelioid cells. Factor VIII was focally positive (membranous), whereas CD31 showed robust, diffuse positivity (membranous and cytoplasmic). The initial margins of resection were negative, and no follow-up radiation or chemotherapy was given. Following a recurrence at the previous excision site, the patient died 7 months after the surgery. Postmortem examination revealed a widely metastatic tumor that involved multiple organ systems. We believe this is the second report of cutaneous angiosarcoma occurring in a chronically lymphedematous abdominal panniculus due to morbid obesity. PMID- 11260631 TI - A case of fibrillary glomerulonephritis with linear immunoglobulin G staining of the glomerular capillary walls. AB - We report a case of crescentic glomerulonephritis that presented with extensive crescent formation and fibrinoid necrosis in the glomeruli. Immunofluorescence staining was strongly positive for linear and pseudolinear staining of the capillary walls for immunoglobulin G (IgG) in the absence of significant mesangial staining. Histologic examination and immunofluorescence staining suggested a diagnosis of anti-glomerular basement membrane disease. However, electron microscopy showed the presence of numerous fibrillary deposits in the subepithelial areas of the glomerular capillary walls, supporting the diagnosis of fibrillary glomerulonephritis. Test results for circulating anti-glomerular basement membrane antibodies were negative. We report this interesting case to illustrate the point that fibrillary glomerulonephritis should be considered in the differential diagnosis of crescentic glomerulonephritis with linear and pseudolinear IgG deposits within the capillary walls. In such cases, electron microscopy is critical in differentiating the cause of crescentic glomerulonephritis. PMID- 11260633 TI - Combined tall cell carcinoma and Hurthle cell carcinoma (collision tumor) of the thyroid. AB - Tall cell carcinoma and Hurthle cell carcinoma are 2 aggressive forms of differentiated follicular-derived thyroid carcinomas. We present a case where these malignant tumors of thyroid coexisted. The tumor originated in the thyroid as a mixed tumor and metastasized as a tall cell tumor to the lymph nodes and as a Hurthle cell carcinoma to lungs. The coexistence of these tumor types is rare and sheds light on the histogenesis of Hurthle cell tumors and the metastatic behavior of combined tumors. PMID- 11260632 TI - Primary high-grade mucosa-associated lymphoid tissue-type lymphoma of the cervix presenting as a common endocervical polyp. AB - Lymphomas of the uterine cervix are uncommon neoplasms and typically appear as diffuse cervical enlargement. We describe a rare case of primary high-grade lymphoma of mucosa-associated lymphoid tissue of the uterine cervix in a 46-year old white woman. The tumor, incidentally disclosed at gynecological examination, appeared as a single common polyp. Immunohistochemical investigation found the lesion to consist of a monomorphic CD20-positive infiltrate of large blasts and rare intermingling centrocyte-like lymphoid cells. A dense area of monotypic (lambda light-chain restriction) plasma cells was found beneath the endocervical mucosa; only a few scattered lymphoepithelial lesions were present. The neoplastic cells did not stain for CD5, CD10, CD23, CD43, or cyclin D1. A bone marrow biopsy displayed a paratrabecular, centrocyte-like B-cell infiltration, but no lymphadenopathy was detected by instrumental examination (computed tomographic scan, magnetic resonance imaging). The tumor was successfully treated by multiagent chemotherapy followed by total hysterectomy. To our knowledge, this case represents the second reported example of mucosa-associated lymphoid tissue type lymphoma occurring in the uterine cervix. We highlight the very unusual gross appearance of this case and emphasize the difficulty of interpreting lymphoid infiltrates in the lower genital tract by microscopy. PMID- 11260634 TI - Granular cell tumor of the thyroid. AB - A case of granular cell tumor of the thyroid gland occurring in a 23-year-old woman is reported. The patient presented with a painless mass in the neck, but was otherwise in good health and had no associated disorders. Fine-needle aspiration biopsy was performed prior to surgery and yielded thyroid epithelial cells with possible oncocytic-type change. To our knowledge, the English language literature contains only one other report of a granular cell tumor of the thyroid. Our case supports the recent suggestion of a neural crest origin for some types of thyroid neoplasia. PMID- 11260635 TI - Spindle cell epithelioma of the vagina shows immunohistochemical staining supporting its origin from a primitive/progenitor cell population. AB - Spindle cell epitheliomas of the vagina (SCEVs) coexpresses epithelial and mesenchymal markers and were first described as a "mixed tumors of the vagina." However, unlike mixed tumors of other organs, which are believed to originate from myoepithelial cells, SCEVs neither immunohistochemically nor ultrastructurally show features of myoepithelial cells. The present expanded battery of immunohistochemical stains is presented on this rare tumor, including cytokeratin AE1/AE3, CK7, CK20, S100 protein, epithelial membrane antigen, alpha smooth muscle actin, desmin, CD34, CD99, Bcl-2, vimentin, estrogen and progesterone receptors, and Ki-67. There was minimal expression of alpha-smooth muscle actin and negative staining with S100 protein, with coexpression of cytokeratins and vimentin and expression of estrogen and progesterone receptors, as previously reported in SCEVs. In addition, diffuse expression of CD34, CD99, and Bcl-2 immunohistochemical stains was found, which has not previously been reported. The coexpression of CD34, CD99, and Bcl-2 in SCEVs is consistent with its origin from a primitive/progenitor cell population. PMID- 11260636 TI - Primary follicular large cell lymphoma of the testis in a child. AB - Primary follicular lymphoma of the testis in childhood is extremely rare. To our knowledge, only 5 cases have been reported to date. We report a case in a 6-year old boy who presented with painless right scrotal enlargement. Right radical orchiectomy revealed a follicular large cell lymphoma with diffuse areas confined to the testis and epididymis, clinical stage IE. Immunohistochemical stains demonstrated that the neoplastic cells were of B-cell lineage, positive for CD10, CD20, CD79a, and BCL-6. Staining for CD21 accentuated networks of dendritic reticulum cells within the nodules. The cells were negative for BCL-2, p53, and T cell antigens. There was no evidence of the t(14;18) detected by polymerase chain reaction. The data suggest that follicular lymphoma of the testis in children has a different pathogenesis than follicular lymphoma in adults. PMID- 11260637 TI - Urinary beta2-microglobulin masquerading as a Bence Jones protein. AB - Bence Jones proteinuria, a common clinical manifestation of multiple myeloma, can also be seen in patients with other B-cell-derived neoplasms. Measurement of pretreatment levels is a useful adjunct in the diagnosis and staging of multiple myeloma, whereas serial levels reflect response to therapy. Serum concentrations of beta2-microglobulin, a small-molecular-weight protein associated with the major histocompatibility complex class I antigens, are often elevated in hematopoietic neoplasms and are also commonly measured at baseline, before treatment, and serially throughout therapy in patients with multiple myeloma and other lymphoproliferative disorders as a marker of tumor burden. Urinary concentrations, however, are considered an indicator of renal tubular function. High urinary levels are found in tubular proteinuria, a frequent sequela of long standing multiple myeloma. A case is described in which a high urinary concentration of beta2-microglobulin interfered with Bence Jones protein quantification by electrophoresis studies. PMID- 11260638 TI - Colonic adenocarcinoma metastasizing as a germ cell neoplasm: a case report and review of the literature. AB - Mixed tumors of the gastrointestinal tract, including both adenocarcinoma and germ cell neoplasm, have been reported infrequently. In the colon, only 9 cases, to our knowledge, have been described in the English-language literature. This is the case of a 29-year-old man with an unsuspected mixed colonic neoplasm that metastasized as the germ cell component. PMID- 11260639 TI - Three-dimensional microscopic image reconstruction of prostatic adenocarcinoma. AB - CONTEXT: Routine microscopy provides only a 2-dimensional view of the complex 3 dimensional structure that makes up human tissue. Three-dimensional microscopic image reconstruction has not been described previously for prostate cancer. OBJECTIVES: To develop a simple method of computerized 3-dimensional image reconstruction and to demonstrate its applicability to the study of prostatic adenocarcinoma. METHODS: Serial sections were cut from archival paraffin-embedded prostate specimens, immunostained using antikeratin CAM5.2, and digitally imaged. Computer image-rendering software was used to produce 3-dimensional image reconstructions of prostate cancer of varying Gleason grades, normal prostate, and prostatic intraepithelial neoplasia. RESULTS: The rendering system proved easy to use and provided good-quality 3-dimensional images of most specimens. Normal prostate glands formed irregular fusiform structures branching off central tubular ducts. Prostatic intraepithelial neoplasia showed external contours similar to those of normal glands, but with a markedly complex internal arrangement of branching lumens. Gleason grade 3 carcinoma was found to consist of a complex array of interconnecting tubules rather than the apparently separate glands seen in 2 dimensions on routine light microscopy. Gleason grade 4 carcinoma demonstrated a characteristic form of glandular fusion that was readily visualized by optically sectioning and rotating the reconstructed images. CONCLUSIONS: Computerized 3-dimensional microscopic imaging holds great promise as an investigational tool. By revealing the structural relationships of the various Gleason grades of prostate cancer, this method could be used to refine diagnostic and grading criteria for this common tumor. PMID- 11260640 TI - Pathologic quiz case: Recurrent dysplasia in an elderly patient 20 years after treatment for achalasia. PMID- 11260641 TI - Pathologic quiz case: A case of a 35-year-old man with cauda equina syndrome. PMID- 11260642 TI - Pathologic quiz case: A persistent cutaneous eruption in a human immunodeficiency virus-infected man. PMID- 11260643 TI - Pathologic quiz case: An exceedingly rare cause of sudden cardiac death. PMID- 11260644 TI - Pathologic quiz case: Male infant with generalized hypotonia and absence of respirations at birth,. PMID- 11260646 TI - An ovarian cholelithiasis. PMID- 11260645 TI - Pathologic quiz case: Hepatic mass in a patient with renal cell carcinoma. PMID- 11260647 TI - Respiratory distress caused by a hydatid cyst. PMID- 11260653 TI - Improvement of tumor oxygenation by mild hyperthermia. AB - There is now abundant evidence that oxygenation in rodent, canine and human tumors is improved during and for up to 1-2 days after heating at mild temperatures. An increase in tumor blood perfusion along with a decline in the oxygen consumption rate appears to account for the improvement of tumor oxygenation by mild hyperthermia. The magnitude of the increase in tumor pO(2), determined with oxygen-sensitive microelectrodes, caused by mild hyperthermia is less than that caused by carbogen breathing. However, mild hyperthermia is far more effective than carbogen breathing in increasing the radiation response of experimental tumors, probably because mild hyperthermia oxygenates both (diffusion-limited) chronically hypoxic and (perfusion-limited) acutely hypoxic cells, whereas carbogen breathing oxygenates only the chronically hypoxic cells. Mild hyperthermia is also more effective than nicotinamide, which is known to oxygenate acutely hypoxic cells, in enhancing the radiation response of experimental tumors. The combination of mild hyperthermia with carbogen or nicotinamide is highly effective in reducing the hypoxic cell fraction in tumors and increasing the radiation response of experimental tumors. A primary rationale for the use of hyperthermia in combination with radiotherapy has been that hyperthermia is equally cytotoxic toward fully oxygenated and hypoxic cells and that it directly sensitizes both fully oxygenated and hypoxic cells to radiation. Such cytotoxicity and such a radiosensitizing effect may be expected to be significant when the tumor temperature is elevated to at least 42-43 degrees C. Unfortunately, it is often impossible to uniformly raise the temperature of human tumors to this level using the hyperthermia devices currently available. However, it is relatively easy to raise the temperature of human tumors into the range of 39-42 degrees C, which is a temperature that can improve tumor oxygenation for up to 1-2 days. The potential usefulness of mild hyperthermia to enhance the response of human tumors to radiotherapy by improving tumor oxygenation merits continued investigation. PMID- 11260654 TI - Targets for radiation-induced cell death: when DNA damage doesn't kill. AB - Chinese hamster ovary (CHO) K1 and radiosensitive CHO irs-20 cells were synchronized in S phase and labeled for 10 min with 5-[(125)I]-iodo-2' deoxyuridine ((125)IdU). The cells were washed, incubated in fresh medium for 1 h for incorporation of the intracellular radionucleotides into DNA, and then frozen (-80 degrees C) for accumulation of (125)I decays. At intervals after freezing, when the cells had accumulated the desired number of decays, aliquots of the frozen cells were thawed and plated to determine survival. The survival curves for K1 and irs-20 cells were similar from 100% to 30% survival. At higher (125)I doses (more decays/cell), the survival of K1 cells continued to decline exponentially, but the survival of X-ray-sensitive irs-20 cells remained at approximately 30% even after the cells had accumulated 1265 decays/cell. The results contradict the notion that increased DNA damage inevitably causes increased cell death. To account for these findings, we propose a model that postulates the existence of a second radiation target. According to this model, radiation damage to DNA may be necessary to induce cell death, but DNA damage alone is not sufficient to kill cells. We infer from the survival response of irs 20 cells that damage to a second (non-DNA) structure is involved in cell death, and that this structure directly affects the repair of DNA and cell survival. PMID- 11260656 TI - Effector genes altered in MCF-7 human breast cancer cells after exposure to fractionated ionizing radiation. AB - Understanding the molecular mechanisms involved in the response of tumors to fractionated exposures to ionizing radiation is important for improving radiotherapy and/or radiochemotherapy. In the present study, we examined the expression of stress-related genes in an MCF-7 cell population (MCF-IR20) that has been derived through treatment with fractionated irradiation (2 Gy per fraction with a total dose of 40 Gy). MCF-IR20 cells showed a 1.6-fold increase in sensitization with dose at 10% isosurvival in a clonogenic assay, and a reduced growth delay ( approximately 15 h compared to approximately 27 h), compared to the parental MCF-7 cells treated with a single dose of 5 Gy. To determine which effector genes were altered in the MCF-IR20 cells, the expression of stress-related effector genes was measured using a filter with 588 genes (Clontech) that included major elements involved in cell cycle control, DNA repair, and apoptosis. Compared to MCF-7 cells that were not exposed to fractionated radiation, 19 genes were up- regulated (2.2-5.1-fold) and 4 were down-regulated (2.7-3.4- fold) in the MCF-IR20 cells. In agreement with the array results, 6 up-regulated genes tested by RT-PCR showed elevated expression. Also, activities of the stress-related transcription factors NFKB, TP53 and AP1 showed a 1.2-4.5-fold increase after a single dose of 5 Gy in MCF-IR20 cells compared with parental MCF-7 cells. However, when the radioresistant MCF-IR20 cell were cultured for more than 12 passages after fractionated irradiation (MCF-RV), radioresistance was lost, with the radiosensitivity being the same as the parental MCF- 7 cells. Interestingly, expression levels of CCNB1, CD9 and CDKN1A in MCF-RV cells returned to levels expressed by the parental cells, whereas the expression levels of three other genes, MSH2, MSH6 and RPA remained elevated. To determine if any of the changes in gene expression could be responsible for the induced radioresistance, CCNB1 and CDKN1A, both of which were up-regulated in MCF IR20 cells and down-regulated in MCF-RV cells, were studied further by transfection with antisense oligonucleotides. Antisense of CCNB1 significantly reduced the clonogenic survival of MCF- IR20 cells at doses of 5 and 10 Gy, from 42% to 26% and from 5.7% to 1.0%, respectively. Antisense of CDKN1A, however, had no effect on radiation survival of MCF-IR20 cells. In summary, these results suggest that stress-related effector genes are altered in cells after treatment with fractionated irradiation, and that up-regulation of CCNB1 is responsible, at least in part, for radioresistance after fractionated irradiation. PMID- 11260655 TI - Radiosensitivity of mammalian cell lines engineered to overexpress cytosolic glutathione peroxidase. AB - Reactive oxygen species are believed to be involved in radiation lethality. Glutathione peroxidase is an intracellular enzyme with antioxidant functions. To determine whether increasing the cellular antioxidant capacity can confer radiation resistance, the effect of overexpression of glutathione peroxidase on radiosensitivity was determined in two different cell types. An expression construct including the bovine cytosolic glutathione peroxidase cDNA was used to overexpress this enzyme in cells of the human lymphoblast cell line Sup-T1 as well as the Chinese hamster ovary cell line AA8. Supplementation of the culture media with 30 nM sodium selenite was included to obtain optimal glutathione peroxidase activity. Northern blot analysis confirmed the presence of the construct mRNA, and a standard coupled spectrophotometric assay demonstrated significantly increased glutathione peroxidase activity in the transfected cell lines. An approximately 8-fold increase was found in the Sup-T1 cells, and an approximately 30-fold increase was obtained in the Chinese hamster ovary AA8 cells. Clonogenic survival was assayed in the overexpressing cells and compared to that in control cells transfected with vector alone. Despite significantly increased glutathione peroxidase activity, no observable radioprotection was conferred in either of the two cell lines studied, indicating that increased glutathione peroxidase activity is insufficient to confer radioresistance in the two cell types examined. These data are discussed in the context of using antioxidants as adjuncts to clinical radiotherapy. PMID- 11260658 TI - Overnight lysis improves the efficiency of detection of DNA damage in the alkaline comet assay. AB - The ability to detect DNA damage using the alkaline comet assay depends on pH, lysis time and temperature during lysis. However, it is not known whether different lysis conditions identify different types of DNA damage or simply measure the same damage with different efficiencies. Results support the latter interpretation for radiation, but not for the alkylating agent MNNG. For X-ray induced damage, cells showed the same amount of damage, regardless of lysis pH (12.3 compared to >13). However, increasing the duration of lysis at 5 degrees C from 1 h to more than 6 h increased the amount of DNA damage detected by almost twofold. Another twofold increase in apparent damage was observed by conducting lysis at room temperature (22 degrees C) for 6 h, but at the expense of a higher background level of DNA damage. The oxygen enhancement ratio and the rate of rejoining of single-strand breaks after irradiation were similar regardless of pH and lysis time, consistent with more efficient detection of strand breaks rather than detection of damage to the DNA bases. Conversely, after MNNG treatment, DNA damage was dependent on both lysis time and pH. With the higher-pH lysis, there was a reduction in the ratio of oxidative base damage to strand breaks as revealed using treatment with endonuclease III and formamidopyrimidine glycosylase. Therefore, our current results support the hypothesis that the increased sensitivity of longer lysis at higher pH for detecting radiation induced DNA damage is due primarily to an increase in efficiency for detecting strand breaks, probably by allowing more time for DNA unwinding and diffusion before electrophoresis. PMID- 11260657 TI - Permanent growth arrest in irradiated human fibroblasts. AB - Exposure of human fibroblasts to doses of ionizing radiation sufficient to cause a permanent growth arrest repressed the expression of genes induced late during G(0)/G(1)-phase traverse, including both cyclin A and cyclin E. In addition, radiation prevented the cell cycle-dependent activation of cyclin D1-associated kinase activity and the subsequent phosphorylation of the RB tumor suppressor protein. Exposure to radiation did not alter the cellular levels of cyclin D1 protein, nor did it alter the formation of cyclin D1-CDK4 complexes. Surprisingly, the repression of cyclin D1-associated kinase activity in damaged mitogen-stimulated quiescent cells could not be accounted for by a relative increase in the association of CDKN1A (also known as p21(Cip1)) with cyclin D1 complexes, nor was cyclin D1 activity targeted by increased levels of CDKN1A in irradiated, logarithmically growing cultures under conditions where cyclin A activity was acutely repressed. Therefore, a radiation-induced permanent growth arrest is mediated by pathways that are distinct from those that cause cell cycle delay in damaged cells involving repression of cyclin-dependent kinase activity by CDKN1A. PMID- 11260659 TI - The effects of 860 MHz radiofrequency radiation on the induction or promotion of brain tumors and other neoplasms in rats. AB - Sprague-Dawley rats were irradiated with a continuous- wave (CW) or a pulsed-wave (P) radiofrequency (RF) for 6 h/day, 5 days/week from 2 up to 24 months of age. The RFs emanated from dipole antennas (1 W average output) 2.0 +/- 0.5 cm from the tip of each rat's nose. The RFs had an 860 MHz frequency, and the specific absorption rate was 1.0 W/ kg averaged over the brain. Fifteen groups of 60 rats (900 total) were formed from offspring of females injected i.v. with 0 (groups 1, 2, 9, 10, 13), 2.5 (groups 5, 6, 7, 8, 11, 12, 14) or 10 mg/kg (groups 3, 4, 15) ethylnitrosourea (ENU) to induce brain tumors. Groups 1, 3, 5 and 7 received the PRF, and groups 9 and 11 the CWRF; groups 2, 4, 6, 8, 10 and 12 were sham irradiated, and groups 13-15 were cage controls. All rats but 2, totaling 898, were necropsied, and major tissues were studied histopathologically. There was no statistically significant evidence that the PRF or CWRF induced neoplasia in any tissues. Additionally, there was no significant evidence of promotion of cranial or spinal nerve or spinal cord tumors. The PRF or CWRF had no statistically significant effect on the number, volume, location, multiplicity, histological type, malignancy or fatality of brain tumors. There was a trend for the group that received a high dose of ENU and was exposed to the PRF to develop fatal brain tumors at a higher rate than its sham group; however, the result was not significant using the log-rank test (P = 0.14, 2-tailed). No statistically significant differences were related to the PRF or CWRF compared to controls in the low- or zero-dose groups regarding tumors of any kind. PMID- 11260661 TI - A model for optimizing normal tissue complication probability in the spinal cord using a generalized incomplete repair scheme. AB - The purpose of this study was to determine the treatment protocol, in terms of dose fractions and interfraction intervals, which minimizes normal tissue complication probability in the spinal cord for a given total treatment dose and treatment time. We generalize the concept of incomplete repair in the linear quadratic model, allowing for arbitrary dose fractions and interfraction intervals. This is incorporated into a previously presented model of normal tissue complication probability for the spinal cord. Equations are derived for both mono-exponential and bi-exponential repair schemes, regarding each dose fraction and interfraction interval as an independent parameter, subject to the constraints of fixed total treatment dose and treatment time. When the interfraction intervals are fixed and equal, an exact analytical solution is found. The general problem is nonlinear and is solved numerically using simulated annealing. For constant interfraction intervals and varying dose fractions, we find that optimal normal tissue complication probability is obtained by two large and equal doses at the start and conclusion of the treatment, with the rest of the doses equal to one another and smaller than the two dose spikes. A similar result is obtained for bi-exponential repair. For the general case where the interfraction intervals are discrete and also vary, the pattern of two large dose spikes is maintained, while the interfraction intervals oscillate between the smallest two values. As the minimum interfraction interval is reduced, the normal tissue complication probability decreases, indicating that the global minimum is achieved in the continuum limit, where the dose delivered by the "middle" fractions is given continuously at a low dose rate. Furthermore, for bi exponential repair, it is seen that as the slow component of repair becomes increasingly dominant as the magnitude of the dose spikes decreases. Continuous low-dose-rate irradiation with dose spikes at the start and end of treatment yields the lowest normal tissue complication probability in the spinal cord, given a fixed total dose and total treatment time, for both mono-exponential and bi-exponential repair. The magnitudes of the dose spikes can be calculated analytically, and are in close agreement with the numerical results. PMID- 11260660 TI - Effect of immobilization and concurrent exposure to a pulse-modulated microwave field on core body temperature, plasma ACTH and corticosteroid, and brain ornithine decarboxylase, Fos and Jun mRNA. AB - Exposure of humans and rodents to radiofrequency (RF) cell phone fields has been reported to alter a number of stress- related parameters. To study this potential relationship in more detail, tube-restrained immobilized Fischer 344 rats were exposed in the near field in a dose-dependent manner to pulse-modulated (11 packets/s) digital cell phone microwave fields at 1.6 GHz in accordance with the Iridium protocol. Core body temperatures, plasma levels of the stress-induced hormones adrenocorticotrophic hormone (ACTH) and corticosterone, and brain levels of ornithine decarboxylase (Odc), Fos and Jun mRNAs were measured as potential markers of stress responses mediated by RF radiation. We tested the effects of the loose-tube immobilization with and without prior conditioning throughout a 2 h period (required for near-field head exposure to RF fields), on core body temperature, plasma ACTH and corticosteroids. Core body temperature increased transiently (+/-0.3 degrees C) during the initial 30 min of loose-tube restraint in conditioned animals. When conditioned/tube-trained animals were followed as a function of time after immobilization, both the ACTH and corticosterone levels were increased by nearly 10-fold. For example, within 2-3 min, ACTH increased to 83.2 +/- 31.0 pg/dl, compared to 28.1 +/- 7.7 pg/dl for cage controls, reaching a maximum at 15-30 min (254.6 +/- 46.8 pg/dl) before returning to near resting levels by 120 min (31.2 +/- 10.2 pg/dl). However, when non-tube-trained animals were submitted to loose-tube immobilization, these animals demonstrated significantly higher (3-10-fold greater) hormone levels at 120 min than their tube-trained counterparts (313.5 +/- 54.8 compared to 31.2 +/- 10.2 pg/dl; corticosterone, 12.2 +/- 6.2 microg/dl compared to 37.1 +/- 6.4 microg/dl). Hormone levels in exposed animals were also compared to those in swim-stressed animals. Swimming stress also resulted in marked elevation in both ACTH and corticosterone levels, which were 10-20 fold higher (541.8 compared to 27.2-59.1 pg/dl for ACTH) and 2-5 fold higher (45.7 compared to 8.4- 20.0 microg/dl for corticosteroids) than the cage control animals. Three time-averaged brain SAR levels of 0.16, 1.6 and 5 W/ kg were tested in a single 2-h RF-field exposure to the Iridium cell phone field. When RF-exposed and sham-exposed (immobilized) animals were compared, no differences were seen in core body temperature, corticosterone or ACTH that could be attributed to near-field RF radiation. Levels of Odc, Fos and Jun mRNA were also monitored in brains of animals exposed to the RF field for 2 h, and they showed no differences from sham-exposed (loose tube immobilized) animals that were due to RF-field exposure. These data suggest that a significant stress response, indicated by a transient increase in core body temperature, ACTH and corticosterone, occurred in animals placed in even the mild loose-tube immobilization required for near-field RF exposure employed here and in our other studies. Failure to adequately characterize and control this immobilization response with appropriate cage control animals, as described previously, could significantly mask any potential effects mediated by the RF field on these and other stress-related parameters. We conclude that the pulse modulated digital Iridium RF field at SARs up to 5 W/kg is incapable of altering these stress-related responses. This conclusion is further supported by our use of an RF-field exposure apparatus that minimized immobilization stress; the use of conditioned/tube-trained animals and the measurement of hormonal and molecular markers after 2 h RF-field exposure when the stress-mediated effects were complete further support our conclusion. PMID- 11260662 TI - Boron neutron capture therapy of a murine mammary carcinoma using a lipophilic carboranyltetraphenylporphyrin. AB - The first control of a malignant tumor in vivo by porphyrin- mediated boron neutron capture therapy (BNCT) is described. In mice bearing implanted EMT-6 mammary carcinomas, boron uptake using a single injection of either p boronophenylalanine (BPA) or mercaptoundecahydrododecaborane (BSH) was compared with either a single injection or multiple injections of the carboranylporphyrin CuTCPH. The BSH and BPA doses used were comparable to the highest doses of these compounds previously administered in a single injection to rodents. For BNCT, boron concentrations averaged 85 microg (10)B/g in the tumor and 4 microg (10)B/g in blood 2 days after the last of six injections (over 32 h) that delivered a total of 190 microg CuTCPH/g body weight. During a single 15, 20, 25 or 30 MW-min exposure to the thermalized neutron beam of the Brookhaven Medical Research Reactor, a tumor received average absorbed doses of approximately 39, 52, 66 or 79 Gy, respectively. A long-term (>200 days) tumor control rate of 71% was achieved at a dose of 66 Gy with minimal damage to the leg. Equivalent long-term tumor control by a single exposure to 42 Gy X rays was achieved, but with greater damage to the irradiated leg. PMID- 11260663 TI - A pharmacokinetic model for the concentration of 10B in blood after boronophenylalanine-fructose administration in humans. AB - An open two-compartment model has been developed for predicting (10)B concentrations in blood after intravenous infusion of the l-p-boronophenylalanine fructose complex (BPA-F) in humans and derived from studies of pharmacokinetics in 24 patients in the Harvard-MIT Phase I clinical trials of BNCT. The (10)B concentration profile in blood exhibits a characteristic rise during the infusion to a peak of approximately 32 microg/g (for infusion of 350 mg/kg over 90 min) followed by a biphasic exponential clearance profile with half-lives of 0.34 +/- 0.12 and 9.0 +/- 2.7 h, due to redistribution and primarily renal elimination, respectively. The model rate constants k(1), k(2) and k(3) are 0.0227 +/- 0.0064, 0.0099 +/- 0.0027 and 0.0052 +/- 0.0016 min(-1), respectively, and the central compartment volume of distribution, V(1), is 0.235 +/- 0.042 kg/kg. The validity of this model was demonstrated by successfully predicting the average pharmacokinetic response for a cohort of patients who were administered BPA-F using an infusion schedule different from those used to derive the parameters of the model. Furthermore, the mean parameters of the model do not differ for cohorts of patients infused using different schedules. PMID- 11260664 TI - Multifrequency EPR study of carbonate- and sulfate-derived radicals produced by radiation in shells and corallite. AB - Shells of two sea mollusks (Venus sp.), pearl oyster (Meleagrina vulgaris) and corallite (white coral) were exposed to ionizing radiation (gamma and X rays) and then examined by EPR spectroscopy in X, Q and W band. The resulting spectra were analyzed and the g values of the EPR lines in the multicomponent spectra were determined. The increased resolution in Q- and W-band spectra allowed us to assign the observed lines to CO(2)(-) ion radicals (isotropic and orthorhombic), SO(2)(-) isotropic, SO(3)(-) (isotropic and axial), and Mn(2+) species. The assignments were confirmed by simulations of the spectra. Practical implications for the use of Q and/or W band in low-dose quantitative EPR measurements for dating and for accidental dose estimation are discussed. PMID- 11260665 TI - Low-energy (5-40 eV) electron-stimulated desorption of anions from physisorbed DNA bases. AB - We present the results of experiments on anion desorption from the physisorbed DNA bases adenine, thymine, guanine and cytosine induced by the impact of low energy (5-40 eV) electrons. Electron bombardment of DNA base films induces ring fragmentation and desorption of H(-), O(-), OH(-), CN(-), OCN(- ) and CH(2)(-) anions through either single or complex multibond dissociation. We designate the variation of the yield of an anion with electron energy as the yield function. Below 15 eV incident electron energy, bond cleavage is controlled mainly by dissociative electron attachment. Above 15 eV, the portion of a yield function that increases linearly is attributed to nonresonant processes, such as dipolar dissociation. A resonant structure is superimposed on this signal around 20 eV in the anion yield functions. This structure implicates dissociative electron attachment and/or resonant decay of the transient anion into the dipolar dissociation channel, with a minimal contribution from multiple inelastic electron scattering. The yields of all desorbing anions clearly show that electron resonances contribute to the damage of all DNA bases bombarded with 5-40 eV electrons. Comparison of the ion yields indicates that adenine is the least sensitive base to slow electron attack. Electron-irradiated guanine films exhibit the largest yields of desorbed anions. PMID- 11260666 TI - Double lesions are produced in DNA oligomer by ionizing radiation and by metal catalyzed H2O2 reactions. AB - It was demonstrated previously that double lesions are produced in DNA by ionizing radiation. These double lesions consist of adjacent nucleotides each bearing a modified base. The goal of the present investigation was to determine whether Fenton chemistry can generate the same kind of lesions. DNA oligomers were exposed to metal-catalyzed H(2)O(2) reactions, and the products were characterized by chromatography and by mass spectrometry. Double lesions are produced by this treatment in which deoxyguanosine is oxidized to 8-oxo-7,8 dihydrodeoxyguanosine and an adjacent pyrimidine nucleoside is degraded to a formamido remnant. PMID- 11260667 TI - Radiation damage in DNA: possible role of higher triplet states. AB - Triplet states of deoxyribose are expected to dissociate efficiently into radicals, leading to strand breaks. Such states could be excited by slow secondary electrons (A) or result from ion recombination in spurs containing two or more ion pairs (B). Estimates of the efficiencies of these processes are presented and the mechanisms discussed in the light of recent work with electrons, vacuum ultraviolet (VUV) photons, and X rays. Route B could play a significant role in producing double-strand breaks, while route A may be a better approach to characterizing the process experimentally. PMID- 11260668 TI - Japan and its women. PMID- 11260669 TI - Collider's moment of truth. PMID- 11260670 TI - German lab unveils plan to build physicists' next particle collider. PMID- 11260672 TI - More culls planned as Britain wrestles with foot-and-mouth. PMID- 11260673 TI - Space-station cuts leave research in lurch. PMID- 11260675 TI - Conflicting volcano tales erupt in public view. PMID- 11260676 TI - Trials offer way forward for Parkinson's. PMID- 11260677 TI - Bush U-turns on pledge for carbon dioxide emissions. PMID- 11260679 TI - 'One woman is enough...'. PMID- 11260681 TI - Sleepless in Seattle. PMID- 11260682 TI - Action is needed now, or BSE crisis could wipe out endangered birds of prey. PMID- 11260683 TI - We must not be bound by anti-GM extremists. PMID- 11260684 TI - Jewish emigrants and German science. PMID- 11260685 TI - Urban myths of organic farming. PMID- 11260690 TI - Chemical analysis. PMID- 11260691 TI - Protein folds: laws of form revisited. PMID- 11260692 TI - Another face in our family tree. PMID- 11260693 TI - Materials science. A pore view of corrosion. PMID- 11260695 TI - Developmental biology. A twist on embryonic signalling. PMID- 11260696 TI - Mesoscopic physics. Noisy times ahead. PMID- 11260697 TI - Immunology. Telling the brain about pain. PMID- 11260698 TI - Glaciology. Enigmatic Arctic ice sheets. PMID- 11260700 TI - Carbon sink for a century. PMID- 11260701 TI - Asexual reproduction: 'midwives' assist dividing amoebae. PMID- 11260702 TI - Inflammatory response: pathway across the blood-brain barrier. PMID- 11260703 TI - Pollination: flexible style that encourages outcrossing. PMID- 11260704 TI - New hominin genus from eastern Africa shows diverse middle Pliocene lineages. AB - Most interpretations of early hominin phylogeny recognize a single early to middle Pliocene ancestral lineage, best represented by Australopithecus afarensis, which gave rise to a radiation of taxa in the late Pliocene. Here we report on new fossils discovered west of Lake Turkana, Kenya, which differ markedly from those of contemporary A. afarensis, indicating that hominin taxonomic diversity extended back, well into the middle Pliocene. A 3.5 Myr-old cranium, showing a unique combination of derived facial and primitive neurocranial features, is assigned to a new genus of hominin. These findings point to an early diet-driven adaptive radiation, provide new insight on the association of hominin craniodental features, and have implications for our understanding of Plio-Pleistocene hominin phylogeny. PMID- 11260706 TI - Stable ultrahigh-density magneto-optical recordings using introduced linear defects. AB - The stability of data bits in magnetic recording media at ultra-high densities is compromised by the thermal 'flips'--magnetic spin reversals--of nano-sized spin domains, which erase the stored information. Media that are magnetized perpendicular to the plane of the film, such as ultrathin cobalt films or multilayered structures, are more stable against thermal self-erasure than conventional memory devices. In this context, magneto-optical memories seem particularly promising for ultrahigh-density recording on portable disks, and bit densities of approximately 100 Gbit inch(-2) (ref. 7) have been demonstrated using recent advances in the bit writing and reading techniques. But the roughness and mobility of the magnetic domain walls prevents closer packing of the magnetic bits, and therefore presents a challenge to reaching even higher bit densities. Here we report that the strain imposed by a linear defect in a magnetic thin film can smooth rough domain walls over regions hundreds of micrometres in size, and halt their motion. A scaling analysis of this process, based on the generic physics of disorder-controlled elastic lines, points to a simple way by which magnetic media might be prepared that can store data at densities in excess of 1 Tbit inch(-2). PMID- 11260707 TI - Giant lateral electrostriction in ferroelectric liquid-crystalline elastomers. AB - Mechanisms for converting electrical energy into mechanical energy are essential for the design of nanoscale transducers, sensors, actuators, motors, pumps, artificial muscles, and medical microrobots. Nanometre-scale actuation has to date been mainly achieved by using the (linear) piezoelectric effect in certain classes of crystals (for example, quartz), and 'smart' ceramics such as lead zirconate titanate. But the strains achievable in these materials are small--less than 0.1 per cent--so several alternative materials and approaches have been considered. These include grafted polyglutamates (which have a performance comparable to quartz), silicone elastomers (passive material--the constriction results from the Coulomb attraction of the capacitor electrodes between which the material is sandwiched) and carbon nanotubes (which are slow). High and fast strains of up to 4 per cent within an electric field of 150 MV x m(-1) have been achieved by electrostriction (this means that the strain is proportional to the square of the applied electric field) in an electron-irradiated poly(vinylidene fluoride-trifluoroethylene) copolymer. Here we report a material that shows a further increase in electrostriction by two orders of magnitude: ultrathin (less than 100 nanometres) ferroelectric liquid-crystalline elastomer films that exhibit 4 per cent strain at only 1.5 MV x m(-1). This giant electrostriction was obtained by combining the properties of ferroelectric liquid crystals with those of a polymer network. We expect that these results, which can be completely understood on a molecular level, will open new perspectives for applications. PMID- 11260705 TI - Observation of high-energy neutrinos using Cerenkov detectors embedded deep in Antarctic ice. AB - Neutrinos are elementary particles that carry no electric charge and have little mass. As they interact only weakly with other particles, they can penetrate enormous amounts of matter, and therefore have the potential to directly convey astrophysical information from the edge of the Universe and from deep inside the most cataclysmic high-energy regions. The neutrino's great penetrating power, however, also makes this particle difficult to detect. Underground detectors have observed low-energy neutrinos from the Sun and a nearby supernova, as well as neutrinos generated in the Earth's atmosphere. But the very low fluxes of high energy neutrinos from cosmic sources can be observed only by much larger, expandable detectors in, for example, deep water or ice. Here we report the detection of upwardly propagating atmospheric neutrinos by the ice-based Antarctic muon and neutrino detector array (AMANDA). These results establish a technology with which to build a kilometre-scale neutrino observatory necessary for astrophysical observations. PMID- 11260708 TI - Evolution of nanoporosity in dealloying. AB - Dealloying is a common corrosion process during which an alloy is 'parted' by the selective dissolution of the most electrochemically active of its elements. This process results in the formation of a nanoporous sponge composed almost entirely of the more noble alloy constituents. Although considerable attention has been devoted to the morphological aspects of the dealloying process, its underlying physical mechanism has remained unclear. Here we propose a continuum model that is fully consistent with experiments and theoretical simulations of alloy dissolution, and demonstrate that nanoporosity in metals is due to an intrinsic dynamical pattern formation process. That is, pores form because the more noble atoms are chemically driven to aggregate into two-dimensional clusters by a phase separation process (spinodal decomposition) at the solid-electrolyte interface, and the surface area continuously increases owing to etching. Together, these processes evolve porosity with a characteristic length scale predicted by our continuum model. We expect that chemically tailored nanoporous gold made by dealloying Ag-Au should be suitable for sensor applications, particularly in a biomaterials context. PMID- 11260709 TI - Ice shelves in the Pleistocene Arctic Ocean inferred from glaciogenic deep-sea bedforms. AB - It has been proposed that during Pleistocene glaciations, an ice cap of 1 kilometre or greater thickness covered the Arctic Ocean. This notion contrasts with the prevailing view that the Arctic Ocean was covered only by perennial sea ice with scattered icebergs. Detailed mapping of the ocean floor is the best means to resolve this issue. Although sea-floor imagery has been used to reconstruct the glacial history of the Antarctic shelf, little data have been collected in the Arctic Ocean because of operational constraints. The use of a geophysical mapping system during the submarine SCICEX expedition in 1999 provided the opportunity to perform such an investigation over a large portion of the Arctic Ocean. Here we analyse backscatter images and sub-bottom profiler records obtained during this expedition from depths as great as 1 kilometre. These records show multiple bedforms indicative of glacial scouring and moulding of sea floor, combined with large-scale erosion of submarine ridge crests. These distinct glaciogenic features demonstrate that immense, Antarctic-type ice shelves up to 1 kilometre thick and hundreds of kilometres long existed in the Arctic Ocean during Pleistocene glaciations. PMID- 11260710 TI - Geochemical tracing of Pacific-to-Atlantic upper-mantle flow through the Drake passage. AB - The Earth's convecting upper mantle can be viewed as comprising three main reservoirs, beneath the Pacific, Atlantic and Indian oceans. Because of the uneven global distribution and migration of ridges and subduction zones, the surface area of the Pacific reservoir is at present contracting at about 0.6 km2 x y(r-1), while the Atlantic and Indian reservoirs are growing at about 0.45 km2 x yr(-1) and 0.15 km2 x yr(-1), respectively. Garfunkel and others have argued that there must accordingly be net mantle flow from the Pacific to the Atlantic and Indian reservoirs (in order to maintain mass balance), and Alvarez further predicted that this flow should be restricted to the few parts of the Pacific rim (here termed 'gateways') where there are no continental roots or subduction zones that might act as barriers to shallow mantle flow. The main Pacific gateways are, according to Alvarez, the southeast Indian Ocean, the Caribbean Sea and the Drake passage. Here we report geochemical data which confirm that there has been some outflow of Pacific mantle into the Drake passage--but probably in response to regional tectonic constraints, rather than global mass-balance requirements. We also show that a mantle domain boundary, equivalent to the Australian-Antarctic discordance, must lie between the Drake passage and the east Scotia Sea. PMID- 11260711 TI - An exceptionally preserved vermiform mollusc from the Silurian of England. AB - Studies of the origin and radiation of the molluscs have yet to resolve many issues regarding their nearest relatives, phylogeny and ancestral characters. The Polyplacophora (chitons) and the Aplacophora are widely interpreted as the most primitive extant molluscs, but Lower Palaeozoic fossils of the former lack soft parts, and the latter were hitherto unrecognized as fossils. The Herefordshire Lagerstatte is a Silurian (about 425 Myr bp) deposit that preserves a marine biota in remarkable three-dimensional detail. The external surface of even non biomineralized cuticle was preserved by entombment in volcanic ash, subsequent incorporation into concretions, and infilling of the fossils with sparry calcite. Here we describe, from this deposit, a complete vermiform mollusc, which we interpret as a plated aplacophoran. Serial grinding at intervals of tens of micrometres, combined with computer-based reconstruction methods, renders the fossils in the round. PMID- 11260712 TI - Intraspecific competition favours niche width expansion in Drosophila melanogaster. AB - Ecologists have proposed that when interspecific competition is reduced, competition within a species becomes a potent evolutionary force leading to rapid diversification. This view reflects the observation that populations invading species-poor communities frequently evolve broader niches. Niche expansion can be associated with an increase in phenotypic variance (known as character release), with the evolution of polymorphisms, or with divergence into many species using distinct resources (adaptive radiation). The relationship between intraspecific competition and diversification is known from theory, and has been used as the foundation for some models of speciation. However, there has been little empirical proof that niches evolve in response to intraspecific competition. To test this hypothesis, I introduced cadmium-intolerant Drosophila melanogaster populations to environments containing both cadmium-free and cadmium-laced resources. Here I show that populations experiencing high competition adapted to cadmium more rapidly than low competition populations. This provides experimental confirmation that competition in a population can drive niche expansion onto new resources for which competition is less severe. PMID- 11260713 TI - Odour-plume dynamics influence the brain's olfactory code. AB - The neural computations used to represent olfactory information in the brain have long been investigated. Recent studies in the insect antennal lobe suggest that precise temporal and/or spatial patterns of activity underlie the recognition and discrimination of different odours, and that these patterns may be strengthened by associative learning. It remains unknown, however, whether these activity patterns persist when odour intensity varies rapidly and unpredictably, as often occurs in nature. Here we show that with naturally intermittent odour stimulation, spike patterns recorded from moth antennal-lobe output neurons varied predictably with the fine-scale temporal dynamics and intensity of the odour. These data support the hypothesis that olfactory circuits compensate for contextual variations in the stimulus pattern with high temporal precision. The timing of output neuron activity is constantly modulated to reflect ongoing changes in stimulus intensity and dynamics that occur on a millisecond timescale. PMID- 11260714 TI - Interleukin-1beta-mediated induction of Cox-2 in the CNS contributes to inflammatory pain hypersensitivity. AB - Inflammation causes the induction of cyclooxygenase-2 (Cox-2), leading to the release of prostanoids, which sensitize peripheral nociceptor terminals and produce localized pain hypersensitivity. Peripheral inflammation also generates pain hypersensitivity in neighbouring uninjured tissue (secondary hyperalgesia), because of increased neuronal excitability in the spinal cord (central sensitization), and a syndrome comprising diffuse muscle and joint pain, fever, lethargy and anorexia. Here we show that Cox-2 may be involved in these central nervous system (CNS) responses, by finding a widespread induction of Cox-2 expression in spinal cord neurons and in other regions of the CNS, elevating prostaglandin E2 (PGE2) levels in the cerebrospinal fluid. The major inducer of central Cox-2 upregulation is interleukin-1beta in the CNS, and as basal phospholipase A2 activity in the CNS does not change with peripheral inflammation, Cox-2 levels must regulate central prostanoid production. Intraspinal administration of an interleukin-converting enzyme or Cox-2 inhibitor decreases inflammation-induced central PGE2 levels and mechanical hyperalgesia. Thus, preventing central prostanoid production by inhibiting the interleukin 1beta-mediated induction of Cox-2 in neurons or by inhibiting central Cox-2 activity reduces centrally generated inflammatory pain hypersensitivity. PMID- 11260715 TI - Homologues of Twisted gastrulation are extracellular cofactors in antagonism of BMP signalling. AB - Twisted gastrulation (TSG) is involved in specifying the dorsal-most cell fate in Drosophila embryos, but its mechanism of action is poorly understood. TSG has been proposed to modify the action of Short gastrulation (SOG), thereby increasing signalling by the bone morphogenetic protein (BMP) Decapentaplegic. SOG, an inhibitor of BMP signalling, is in turn inactivated by the protease Tolloid. Here we identify Tsg gene products from human, mouse, Xenopus, zebrafish and chick. Expression patterns in mouse and Xenopus embryos are consistent with in vivo interactions between Tsg, BMPs and the vertebrate SOG orthologue, chordin. We show that Tsg binds both the vertebrate Decapentaplegic orthologue BMP4 and chordin, and that these interactions have multiple effects. Tsg increases chordin's binding of BMP4, potentiates chordin's ability to induce secondary axes in Xenopus embryos, and enhances chordin cleavage by vertebrate tolloid-related proteases at a site poorly used in Tsg's absence; also, the presence of Tsg enhances the secondary axis-inducing activity of two products of chordin cleavage. We conclude that Tsg acts as a cofactor in chordin's antagonism of BMP signalling. PMID- 11260716 TI - Twisted gastrulation is a conserved extracellular BMP antagonist. AB - Bone morphogenetic protein (BMP) signalling regulates embryonic dorsal-ventral cell fate decisions in flies, frogs and fish. BMP activity is controlled by several secreted factors including the antagonists chordin and short gastrulation (SOG). Here we show that a second secreted protein, Twisted gastrulation (Tsg), enhances the antagonistic activity of Sog/chordin. In Drosophila, visualization of BMP signalling using anti-phospho-Smad staining shows that the tsg and sog loss-of-function phenotypes are very similar. In S2 cells and imaginal discs, TSG and SOG together make a more effective inhibitor of BMP signalling than either of them alone. Blocking Tsg function in zebrafish with morpholino oligonucleotides causes ventralization similar to that produced by chordin mutants. Co-injection of sub-inhibitory levels of morpholines directed against both Tsg and chordin synergistically enhances the penetrance of the ventralized phenotype. We show that Tsgs from different species are functionally equivalent, and conclude that Tsg is a conserved protein that functions with SOG/chordin to antagonize BMP signalling. PMID- 11260717 TI - Twisted gastrulation can function as a BMP antagonist. AB - Bone morphogenetic proteins (BMPs), including the fly homologue Decapentaplegic (DPP), are important regulators of early vertebrate and invertebrate dorsal ventral development. An evolutionarily conserved BMP regulatory mechanism operates from fly to fish, frog and mouse to control the dorsal-ventral axis determination. Several secreted factors, including the BMP antagonist chordin/Short gastrulation (SOG), modulate the activity of BMPs. In Drosophila, Twisted gastrulation (TSG) is also involved in dorsal-ventral patterning, yet the mechanism of its function is unclear. Here we report the characterization of the vertebrate Tsg homologues. We show that Tsg can block BMP function in Xenopus embryonic explants and inhibits several ventral markers in whole-frog embryos. Tsg binds directly to BMPs and forms a ternary complex with chordin and BMPs. Coexpression of Tsg with chordin leads to a more efficient inhibition of the BMP activity in ectodermal explants. Unlike other known BMP antagonists, however, Tsg also reduces several anterior markers at late developmental stages. Our data suggest that Tsg can function as a BMP inhibitor in Xenopus; furthermore, Tsg may have additional functions during frog embryogenesis. PMID- 11260718 TI - An Arabidopsis circadian clock component interacts with both CRY1 and phyB. AB - Most organisms, from cyanobacteria to mammals, use circadian clocks to coordinate their activities with the natural 24-h light/dark cycle. The clock proteins of Drosophila and mammals exhibit striking homology but do not show similarity with clock proteins found so far from either cyanobacteria or Neurospora. Each of these organisms uses a transcriptionally regulated negative feedback loop in which the messenger RNA levels of the clock components cycle over a 24-h period. Proteins containing PAS domains are invariably found in at least one component of the characterized eukaryotic clocks. Here we describe ADAGIO1 (ADO1), a gene of Arabidopsis thaliana that encodes a protein containing a PAS domain. We found that a loss-of-function ado1 mutant is altered in both gene expression and cotyledon movement in circadian rhythmicity. Under constant white or blue light, the ado1 mutant exhibits a longer period than that of wild-type Arabidopsis seedlings, whereas under red light cotyledon movement and stem elongation are arrhythmic. Both yeast two-hybrid and in vitro binding studies show that there is a physical interaction between ADO1 and the photoreceptors CRY1 and phyB. We propose that ADO1 is an important component of the Arabidopsis circadian system. PMID- 11260719 TI - A metabolic enzyme for S-nitrosothiol conserved from bacteria to humans. AB - Considerable evidence indicates that NO biology involves a family of NO-related molecules and that S-nitrosothiols (SNOs) are central to signal transduction and host defence. It is unknown, however, how cells switch off the signals or protect themselves from the SNOs produced for defence purposes. Here we have purified a single activity from Escherichia coli, Saccharomyces cerevisiae and mouse macrophages that metabolizes S-nitrosoglutathione (GSNO), and show that it is the glutathione-dependent formaldehyde dehydrogenase. Although the enzyme is highly specific for GSNO, it controls intracellular levels of both GSNO and S nitrosylated proteins. Such 'GSNO reductase' activity is widely distributed in mammals. Deleting the reductase gene in yeast and mice abolishes the GSNO consuming activity, and increases the cellular quantity of both GSNO and protein SNO. Furthermore, mutant yeast cells show increased susceptibility to a nitrosative challenge, whereas their resistance to oxidative stress is unimpaired. We conclude that GSNO reductase is evolutionarily conserved from bacteria to humans, is critical for SNO homeostasis, and protects against nitrosative stress. PMID- 11260720 TI - Covalent inhibition revealed by the crystal structure of the caspase-8/p35 complex. AB - Apoptosis is a highly regulated process that is crucial for normal development and homeostasis of multicellular organisms. The p35 protein from baculoviruses effectively prevents apoptosis by its broad-spectrum caspase inhibition. Here we report the crystal structure of p35 in complex with human caspase-8 at 3.0 A resolution, and biochemical and mutagenesis studies based on the structural information. The structure reveals that the caspase is inhibited in the active site through a covalent thioester linkage to p35, which we confirmed by gel electrophoresis, hydroxylamine treatment and mass spectrometry experiments. The p35 protein undergoes dramatic conformational changes on cleavage by the caspase. The repositioning of the amino terminus of p35 into the active site of the caspase eliminates solvent accessibility of the catalytic dyad. This may be crucial for preventing hydrolysis of the thioester intermediate, which is supported by the abrogation of inhibitory activity through mutations at the N terminus of p35. The p35 protein also makes conserved contacts with the caspase outside the active-site region, providing the molecular basis for the broad spectrum inhibitory activity of this protein. We demonstrate a new molecular mechanism of caspase inhibition, as well as protease inhibition in general. PMID- 11260721 TI - The effect of temperature and saturation deficit on mortality in populations of male Glossina m. morsitans (Diptera: Glossinidae) in Zimbabwe and Tanzania. AB - The methods of Bailey and of Jolly and Seber were used to provide maximum likelihood estimates of population parameters for Jackson's classical mark recapture experiments on males of the tsetse fly Glossina m. morsitans Westwood. These were compared with Jolly-Seber (J-S) estimates for the same fly from more recent work on Antelope Island, Lake Kariba, Zimbabwe. The Bailey estimates of birth and death rates and total population size had markedly lower variances than Jackson's originals. Both sets of estimates provided moving averages over 6-week periods, whereas the Jolly-Seber analysis provided independent weekly estimates and their variance is consequently higher. Saturation deficit and maximum temperature (Tmax) accounted for 11 and 16% respectively of the variance in independent 4-week means of the weekly J-S survival probabilities. Analysis of covariance, carried out on a joint data set of smoothed J-S estimates of the survival probability in Tanzania and Zimbabwe, showed a significant effect of Tmax on survival. When this effect was removed, the survival probability in the Tanzania studies was found to be 8% lower than on Antelope Island. The two effects accounted for 50% of the variance in the joint data. When saturation deficit was substituted for Tmax, regression only accounted for 35% of the variance. If saturation deficit is important in determining tsetse survival, it must act on stages other than the post-teneral adult. Given the continuous increase in mortality, even at moderate temperatures, it is hard to envisage a direct effect of Tmax. There may be an indirect effect, however, via the number of hunger-related deaths resulting from the increase in the feeding rate with increasing temperature. PMID- 11260722 TI - Detection of Brugia malayi in laboratory and wild-caught Mansonioides mosquitoes (Diptera: Culicidae) using Hha I PCR assay. AB - An Hha 1 based polymerase chain reaction (PCR) assay developed for the detection of Brugia malayi, the causative agent of Brugian lymphatic filariasis, was evaluated for its sensitivity in the laboratory and for its usefulness in measuring changes in transmission of the disease in the field. Laboratory studies showed that the new assay was highly sensitive in comparison with the standard dissection and microscopy technique. The assay can detect as little as 4 pg of parasite DNA or a single microfilaria in pools of up to 100 mosquitoes. The optimum pool size for convenience was found to be 50 mosquitoes per pool. The efficacy of PCR assay was evaluated in filariasis control programmes in operation in endemic areas of Kerala State, South India. The infection rates obtained by the Hha I PCR assay and the conventional dissection and microscopy technique were 1.2% and 1.7% respectively in operational areas and 8.3% and 4.4% respectively, in check areas, which were not significantly different (P < 0.05). Thus, the Hha I PCR assay was found to be as sensitive as the conventional technique and hence it can be used as a new epidemiological tool for assessing parasite infection in field-collected mosquitoes. PMID- 11260723 TI - Aerial treatment of the Australian plague locust, Chortoicetes terminifera (Orthoptera: Acrididae) with Metarhizium anisopliae (Deuteromycotina: Hyphomycetes). AB - Between October 1999 and April 2000, nearly 4000 ha of nymphal bands and adult swarms of Chortoicetes terminifera (Walker) were aerially treated using a ULV oil formulation of strain FI-985 of Metarhizium anisopliae var. acridum. During the mild weather (maxima 22-30 degrees C) of spring (October), there was little change in nymphal bands during the first week but at all doses between 25-100 g (1-4 x 10(12) conidia) ha(-1), the bands rapidly declined 9-12 days after treatment reaching > 90% mortality by 14 days. Metarhizium persisted for some time as there was 50% mortality of locusts fed vegetation collected from the treated blocks seven days after treatment. Persistence was confirmed by the high mortality of bands that invaded from untreated areas and of nymphs that hatched on the plot five to seven days after treatment, though mortality was then delayed until early in the third week. During summer (January), temperatures were high (maxima 36-42 degrees C), and at all doses between 25 and 125 g (1-5 x 10(12) conidia) ha(-1), there was a rapid decline seven to ten days after treatment. By 12-14 days, there was a > 90% decline in numbers in most blocks which was confirmed by helicopter surveys two weeks after treatment that found very few adults within or near treated areas. Mortality was delayed in the high dose where there were blockages of spray equipment during treatment. The clear demonstration that Metarhizium can suppress small local populations of C. terminifera led to the limited operational use of Metarhizium on an organic farm and in a National Park where nearly 2500 ha of bands and swarms were treated. Continued research is needed to develop a commercially viable product so that Metarhizium can form a significant part of a programme of integrated pest management of locusts in Australia. PMID- 11260724 TI - Description of an invasive new species of Neotropical aleurodicine whitefly (Hemiptera: Aleyrodidae) - a case of complete or partial misidentification? AB - Paraleyrodes pseudonaranjae sp. n. is here described, from material collected from invasive colonies affecting agriculture in Hong Kong, Hawaii and the USA (Florida). This species had previously been identified as P. naranjae Dozier, based on puparial characteristics, but comparison of adult males with male syntypes of P. naranjae has indicated that identification to be mistaken. An adult male has been selected as lectotype of P. naranjae, but the identity of the puparia of the original sample remains uncertain. PMID- 11260725 TI - Emergence trap developed to capture adult large pine weevil Hylobius abietis (Coleoptera: Curculionidae) and its parasite Bracon hylobii (Hymenoptera: Braconidae). AB - A novel type of emergence trap capable of capturing and separating 'live' insect catches is described. The trap was shown to be 96% efficient at capturing newly emergent adult Hylobius abietis Linnaeus on bare ground and at least 82% efficient over stumps on a weedy Sitka spruce clearfell. The trap was more than 80% efficient at capturing Bracon hylobii Ratzeburg, the most commonly found parasite of H. abietis. It was also shown to be effective at capturing adult H. abietis of unknown age (98%), indicating that it could also be used to trap out H. abietis from known areas to estimate on-site overwintering densities. Fifty four percent of newly emergent weevils were captured within 12 h of release on bare mineral soils. Forty-two percent of unknown age weevils and 52% of parasites were captured within 1 h of their release within the trap. The rapid rates of capture mean that when traps are checked frequently they can be used to reflect accurately temporal patterns of emergence. Its potential for use in control programmes and ecological studies is discussed. PMID- 11260726 TI - Synonymy between two spider mite species, Tetranychus kanzawai and T. hydrangeae (Acari: Tetranychidae), shown by ribosomal ITS2 sequences and cross-breeding experiments. AB - Amplification of the second internal transcribed spacer (ITS2) of ribosomal DNA was used to compare seven samples of the Tetranychus kanzawai Kishida-- T. hydrangeae Pritchard & Baker mite complex from five different countries: Australia, the Congo, Indonesia, Japan and the USA. No morphological differences were detected between these mites and their ITS2 sequences displayed strong similarity except for a small nucleotide divergence of 0.2% in specimens from Australia and Indonesia. Reciprocal crosses and backcrosses between mites assumed to be T. kanzawai and T. hydrangeae respectively showed reproductive compatibility. Fertile hybrid females were obtained in all cases, indicating conspecificity of the mites tested. It is concluded that T. hydrangeae is a synonym of T. kanzawai. The evidence suggests that T. kanzawai originated in South-east Asia and probably spread throughout the world on Hydrangea spp. cuttings. PMID- 11260727 TI - Feeding behaviour of the aphid Rhopalosiphum padi (Hemiptera: Aphididae) on nitrogen and water-stressed barley (Hordeum vulgare) seedlings. AB - Electrical penetration graphs (EPGs) were used to examine the probing behaviour of adult apterous Rhopalosiphum padi (Linnaeus) on barley seedlings grown under conditions of nitrogen or water stress. Aphids took significantly longer to reach and ingest from sieve elements of nitrogen-deficient seedlings than from nitrogen sufficient seedlings but there were no such differences between water-stressed or well-watered seedlings. On both nitrogen and water-stressed seedlings the average length of each individual period of salivation into the sieve element was significantly greater compared with their respective unstressed controls. PMID- 11260728 TI - Topical treatment of calves with synthetic pyrethroids: effects on the non-target dung fly Neomyia cornicina (Diptera: Muscidae). AB - Dung from calves treated with synthetic pyrethroids negatively influenced, in varying degrees, survival, reproduction and size of the common dung fly Neomyia cornicina (Fabricius). This was documented in assays where the coprophagous larvae and adults of N. cornicina were exposed to dung collected from calves dosed with topical preparations of deltamethrin, flumethrin, cyfluthrin, and alpha-cypermethrin. Larval mortality was significantly increased in dung collected up to at least seven days after treatment with deltamethrin, alpha cypermethrin and cyfluthrin. Alpha-cypermethrin caused significant mortality of adults allowed to feed on moist dung. Nulliparous flies fed for six days on dung collected three days after treatment of calves with alpha-cypermethrin or deltamethrin showed little or no ovarian development. A tendency for a comparable effect with flumethrin was also observed. A connection between ovarian development and inhibition of feeding was indicated by the observation of significantly lowered excretion rates in flies exposed to residues of deltamethrin, alpha-cypermethrin and flumethrin. Larvae that survived exposure to dung from calves dosed with deltamethrin, alpha-cypermethrin, or cyfluthrin gave rise to smaller flies. The effect on adult fly size decreased when larvae were exposed to dung collected at longer times after treatment of the calves. Adult fly size was significantly reduced in dung collected up to 14 days (alpha cypermethrin) or up to 28 days after treatment (deltamethrin and cyfluthrin). Fluctuating asymmetry of a wing vein character did not reflect the anticipated levels of exposure. The study strongly indicated that the use of synthetic pyrethroids affected the insect dung fauna and that such use may reduce dung decomposition. PMID- 11260729 TI - Microsatellite analysis of the Queensland fruit fly Bactrocera tryoni (Diptera: Tephritidae) indicates spatial structuring: implications for population control. AB - The population structure of a tephritid pest species, the Queensland fruit fly Bactrocera tryoni (Froggatt), has been analysed over a five year period (1994 1998), using six microsatellites. Adult fly samples were collected to cover most regions of eastern and central Australia where the flies are regularly found. Tests for heterogeneity indicated that flies within geographically defined regions were homogeneous. The samples were allocated into five regions, including one very large region, Queensland, which encompasses that portion of the fly's range where breeding can occur year-round. With one exception, the collections from different regions were homogeneous between years, showing a fairly static distribution of the species. However, differences between regions were highly significant. The one case of a change in frequency between years indicated a gradual replacement of flies in a marginal region by flies from the main part of the range. The finding of stability in the distribution of a highly mobile insect is of interest, potentially also for other species which have expanded beyond their native range. It is argued that a contributing reason for this stability may be adaptation to different climatic regimes, and that strategies for control based on this hypothesis afford a reasonable chance of success. PMID- 11260731 TI - Horizon scanning in medical education: 2020 vision. PMID- 11260730 TI - Dependence of Sitona lepidus (Coleoptera: Curculionidae) larvae on abundance of white clover Rhizobium nodules. PMID- 11260732 TI - Rewarding teaching excellence. Can we measure teaching 'excellence'? Who should be the judge? PMID- 11260733 TI - Twelve tips for potential distance learners. AB - Distance learning courses are becoming popular among medical professionals due to their flexibility, allowing minimal disruption to personal and professional commitments. The ability to continue professional duties, allied to the reduced cost of distance learning courses, also makes them attractive to institutions looking to develop the skills of their staff. However the nature of distance learning courses means that they are often of long duration and many students fail to maintain motivation while working in isolation. This is reflected by high non-completion rates. This article outlines issues that all students planning a distance learning course should consider, relating to choice of course, time management, funding and adjusting to the different nature of distance learning. The authors advise developing a support network for distance learning students, either in person or electronically, to increase motivation and completion. PMID- 11260734 TI - Simulation and new learning technologies. AB - Changes in medical practice that limit patient availability and instructors' time have resulted in poor physical diagnosis skills by learners at all levels. Advanced simulation technology, including the use of sophisticated multimedia computer systems, helps to address this problem. For many years 'Harvey', the Cardiology Patient Simulator, and the UMedic Multimedia Computer system have proven to be effective tools to teach and assess bedside cardiovascular skills when they are integrated into the required curriculum of medical school and postgraduate training. In the future, virtual reality technology, based initially on data from the Visible Human Data set, will provide the majority of simulation based training. Models that provide a high level of visual fidelity and use sophisticated haptic devices that simulate the 'touch' and 'feel' of a procedure or examination are now being used in selected medical centers. The presence of these tools is not enough. Evidence-based outcomes must show these systems to be effective instruments for teaching and assessment, and medical educators must be willing to effect change in medical education to ensure the appropriate use of these systems in the next millennium. PMID- 11260735 TI - The development and uses of a computerized evaluation tool for senior house office posts in the West Midlands Region, UK. AB - The senior house officer grade contains the largest group of junior doctors in the United Kingdom. For many years there has been concern over their poor training, long hours of work, poor supervision, little or no feedback (often given inappropriately), and their levels of psychological distress. Evaluation of senior house officer posts has been cumbersome and inconsistent. This paper describes the origins, design, piloting, testing for reliability and subsequent use of a simple 10-point evaluation form for senior house officers in the West Midlands. The process has now been computerized, and has been incorporated into the postgraduate dean's database of all junior doctor posts in the West Midlands. The evaluation of all 1500 senior house officer posts can now be performed every 6 months. Detailed results can be obtained and fed back to clinical tutors, hospitals, and Royal Colleges. The uses of such data in improving the lot of the senior house officer are discussed. PMID- 11260736 TI - Correlation of a written test of skills and a performance based test: a study in two traditional medical schools. AB - Studies in innovative curricula have shown that a written test of skills is potentially able to predict OSCE scores. In this study we verified the potential of a written test of skills as an alternative to an OSCE in two traditional medical schools. A 12 station OSCE was scored using detailed checklists and global rating scales. The written test of skills consisted of 132 true-false questions. Students were assessed immediately before graduation in 1997. The size of the reported correlations confirms that the written test of skills can be used to predict performance-based test scores in traditional medical schools to some extent. For research purposes, particularly for outcome research to compare curricula of medical schools, a written test of skills is a viable alternative to performance-based testing. A written test of skills is unable to replace the OSCE for assessment of individual students. PMID- 11260737 TI - Lessons learned from the development of a distance-learning programme. AB - The development of an effective educational programme, i.e. one that not only delivers education but which also fosters change, requires both educational skills and subject expertise from the producers. This paper looks at the key lessons learned when developing a distance-learning programme. These seven key lessons, which focus heavily on needs assessment as well as team working and the need to consider a multiprofessional approach, offer practical advice to those in the medical and dental professions wishing to produce their own educational programmes. PMID- 11260738 TI - Microburst Teaching and Learning. AB - Amidst changes in the health care field with requirements for increased efficiency and limited time for teaching, there is a need for a teaching-learning model which maximizes the learning process and is exciting, fun, and motivating for both teacher and learner. Microburst Teaching and Learning is one strategy for combining various teaching styles and methods in 'bursts' with different learning styles to enhance the learning process.The model accommodates adult learning theory, adult attention span, learner motivation, the variety of learning styles found in learners, and the need for efficiency. Preliminary reactions to the Microburst Model indicate its appeal and motivating nature as a useful teaching-learning model.The next steps are to more critically evaluate the efficacy of the model for a broader range of clinical preceptors and to examine the variety of specific teaching strategies to determine which methods work best in specific settings. Because there are many potential teaching methods and teaching styles from which medical teachers can choose, a companion article outlining these specific methods and styles is currently in preparation.Weanticipate the article's publication within the next year. PMID- 11260739 TI - Antenatal provision of additional information about the role of students in the labour suite and their subsequent involvement in care: a randomised controlled trial. AB - To determine whether the antenatal provision of additional information to pregnant women about the roles of medical and midwifery students in the labour suite increases their actual and theoretical involvement in care during labour, 624 pregnant women booked for delivery at Ninewells Hospital and Medical School, Dundee were randomised at 34 weeks gestation to receive either a specifically produced information leaflet through the post on the roles of medical and midwifery students in the labour suite, or no additional information. Post delivery and following discharge, women were asked to complete a questionnaire about people who were involved in their care during labour. There were no significant differences in a women's willingness (actual or theoretical) to have medical or midwifery students involved in their care between women who received additional information and those who did not. Of those women who had a student present during labour, midwifery students were significantly more involved in all care activities than medical students (p<0.01 for all activities), with the exception of vaginal examination. Women who did not have a student present because one was not available, but who indicated that they would have allowed a student to participate in their care if one had been available, were significantly more likely to allow a midwifery student to perform vaginal examinations, assist with delivery and attend the baby than medical students (p<0.01 for all activities). They would allow equal participation in other activities. For both groups of students, these women appeared to be willing for significantly more active involvement in care than is happening in practice. PMID- 11260740 TI - Student perceptions about the occurrence of critical incidents in tutorial groups. AB - The study reported here examines student perceptions about the occurrence of critical incidents in tutorial groups across years of medical training. The study investigates the following research questions. (1) Which factors underlie the occurrence of critical incidents in the tutorial group? (2) How do students rate the incidents with respect to whether they occur in the tutorial groups? (3) Are there differences in scores on the factors identified across years of medical training? The subjects consisted of a stratified random sample of 200 students at the Medical School of the University of Maastricht. In general, the results of the confirmatory factor analyses indicate that a six-factor model showed a good fit. The results show that students frequently perceive lack of elaboration, lack of interaction and unequal participation in tutorial groups. For most of the factors (lack of elaboration, difficult personalities, lack of cohesion, and lack of motivation) the average scores differ significantly (p<0.01) across the years. PMID- 11260741 TI - Task-based learning (TBL) in undergraduate medical education. AB - Problem-based learning (PBL) is a proven method to learn medicine during the first years of studies. In the clinical phase the active, self-directive student may experience difficulties in adapting to the life of professionals in health care units, where students usually have to attend and work according to preplanned timetables. Task-based learning (TBL) can serve as an intermediary in the meeting of these two cultures. Here we describe a TBL study module for fourth year medical students and experiences of implementing it at the University of Tampere in Finland. Eighty-five students participated in this study in 1998 and 1999. Our results show that this method works and that it leads to learning. Students evaluate their skills connected with the general practitioner's work in a health centre hospital as better after the study module than at the onset. PMID- 11260742 TI - Developing a questionnaire to assess student awareness of the need to be culturally aware in clinical practice. AB - This study aimed to establish whether students had an awareness of the requirement to consider cultural issues in caring for patients and to identify those issues which are most difficult for students, in order to aid course development. Data was collected using previously developed questionnaires which were modified and added to for the study undertaken at the University of Leicester Medical School in February and June 1998. Thirty fourth year and 75 second year medical students completed the questionnaires. Students accepted that they as doctors have a responsibility to be aware of the different cultures within their practice. Their responses supported the introduction of sessions on cultural and racial awareness as part of the Human Diversity Module. The sessions have been designed to facilitate students to explore their own perception and understanding of culture in a challenging, relevant and enjoyable way. The questionnaire developed in this study will be used to compare student attitudes pre- and post-teaching sessions to assess any change. PMID- 11260743 TI - An assessment of medical students' experiences of learning about the psychosocial enquiry in their introductory clinical course. AB - Medical students' ability to take a meaningful psychosocial history has been shown to decline during clinical training. We postulated that psychosocial histories are given a low priority in busy clinical attachments. The aim of this study was to identify factors that affect how medical students gain skills in psychosocial assessment. A random sample of 37 students filled in a questionnaire before and after their introductory course, and they were asked to keep a logbook of their experiences of teaching about psychosocial history taking. There were 504 teaching experiences recorded of which less than half were positive. Negative experiences often related to poor communication by clinicians. At the end of the course less than half the students felt confident in taking a psychosocial history. To improve doctors' skills in this important area we suggest that teaching in psychosocial history taking should be made explicit, as an integrated part of the overall assessment of a patient. PMID- 11260745 TI - The new medical curriculum at Flinders University, South Australia: from concept to reality. AB - After much discussion and planning, Flinders University in Adelaide, South Australia recently introduced a new Graduate-Entry Medical Program (GEMP) which centres on problem-based learning (PBL). We describe the factors that stimulated the development of this new course, discuss its aims and philosophies and provide a brief outline of its structure. Advice and practical help was freely provided by several institutions who had undertaken similarly radical curricular reform and without this, a difficult task would have been much harder. We hope that our experiences will be of interest and help to others who are considering curricular reform. PMID- 11260744 TI - Ethical issues in the managed care setting: a new curriculum for primary care physicians. AB - Several challenging ethical issues have been associated with the shift to managed healthcare in the United States. Our objective was to develop, implement, and evaluate a curriculum designed to help physicians identify and examine ethical issues encountered in the managed care setting. The curriculum was developed during a year-long workshop at Johns Hopkins Bayview Medical Center. The content of the curriculum was established through literature review, focus group discussions with physicians, and a needs assessment of targeted learners (primary care physicians practicing in managed care settings). Some of the key issues addressed in the curriculum include: changing professional responsibilities of physicians; fair use of resources; and threats to the doctor-patient relationship as a consequence of the new healthcare delivery system. The 7.5-h curriculum was taught over five sessions using varied teaching methods. Evaluations demonstrated that the curriculum was successful in increasing learner awareness of ethical issues confronted in the managed care environment and improved learner knowledge in these areas. The physician-learners reported that this educational experience would change their teaching of medical students and residents. After completing the curriculum, learners felt that they were at least somewhat better able to cope with ethical challenges encountered in the managed care setting. Future research might examine whether such a curriculum could positively affect physician behavior or enhance physician satisfaction with the managed care setting. PMID- 11260746 TI - Intercultural training of medical students. AB - Until recently the Utrecht Medical School had a traditional curriculum with a predominantly biomedical orientation and strong emphasis on curative medicine. In 1997 an experimental 'Multi-cultural Family Attachment Course' started at the Utrecht Medical School with 20 second-year medical students. Each student was attached to a native Dutch and an ethnic minority family with a newborn or chronically ill child. In a period of 1.5 years students had to visit each family at home four times. The students monitored growth and development of the child and discussed several aspects of health and disease with the parents according to a structured schedule. In regular group sessions students reported back their experiences. In this way, the influence of socioeconomic circumstances, culture and environment on health becomes a real-life experience. This paper aims to describe some aspects of this pilot-course and the reactions of the students. PMID- 11260747 TI - Educational benefits of blinding students to information acquired and management plans generated by other physicians. AB - At our university, Internal Medicine clerks are members of a team responsible for the care of patients hospitalized on a teaching ward. Clerks first encounter their patients after the latter have been fully worked up by other physicians who have examined them and initiated investigations and management. Clerks are thus deprived of the opportunity to practice information acquisition, hypothesis generation and problem solving. We therefore undertook a 'blinding' initiative wherein each clerk was required to work up at least one hospitalized patient per week without access to the patient chart and without knowledge of information acquired and hypotheses generated by other physicians. Weekly data collection during the 8-week experiment with 40 clinical clerks revealed that work up of 'blinded' patients was more time-consuming and more difficult than work up of unblinded patients. Clerks were appreciative of the educational value of blinding. Teaching faculty felt clerk 'blinding' to be a practical approach to approximating the true conduct of medical practice and as such was useful for student learning. PMID- 11260748 TI - Supporting medical students to take responsibility for developing their communication skills. AB - We describe a learning structure to support medical students in their own development of communication skills. It is student-centred, patient-independent, cost-effective of staff time, and adaptable to a range of medical school contexts and circumstances. PMID- 11260749 TI - Differentiation and undergraduate medical education. PMID- 11260750 TI - Teaching evidence-based medicine using literature for problem solving. PMID- 11260751 TI - Assessments in evidence-based medicine workshops: loose connection between perception of knowledge and its objective assessment. AB - The outcome of continuing education programs is often based on self-assessment. We evaluated the relationship of self-assessment of knowledge based on rating scales with scores obtained on objective validated tests in evidence-based medicine workshops. In the West Midlands region (1998), 55 participants attended three workshops in critical appraisal of the medical literature.They completed two self-assessment questionnaires: one used a rating scale to subjectively examine the level of knowledge of six different literature appraisal issues; the other objectively assessed participants' literature appraisal knowledge in those issues using validated multiple true-false questions. Comparison of subjective scores reflecting understanding of specific literature appraisal issues with corresponding objective test scores revealed a poor correlation (r(s) ranged from -0.29 to 0.60 for the different knowledge issues assessed). Perception of ones level of knowledge did not always correlate with correctly possessed knowledge. In some instances, those who thought they were knowledgeable actually possessed incorrect knowledge. Therefore, continuing medical education programs should focus on objective, not subjective tests to assess outcome. PMID- 11260752 TI - An intercalated BSc in Primary Health Care - an outline of a new course. AB - This paper describes a BSc in Primary Health Care. The course is in its fourth year, with seventeen graduates. We expect the nine currently enrolled to graduate in summer 2001. The aims of the course are to give students additional skills, that are generic to medicine (research methods etc.) and also specific to primary health care. The course consists of a taught component about the theoretical background to primary care, with input from the social sciences, epidemiology and public health. The self-directed component consists of a research project and, unusually for a BSc, a regular clinical placement. This course may help to change students' expectations that BScs are only for those who are interested in laboratory based medicine. PMID- 11260753 TI - Management development of clinicians in Indonesia. AB - This paper evaluates an open learning and applied learning programme for health professionals delivered by the authors in Indonesia. It also highlights the importance of the active involvement of Indonesian health professionals in the delivery and the need for the programme to be contextualised. The background is a pilot programme for the management development of clinicians in Indonesia. As a country facing huge political, economic and social problems there is an urgent need to improve the health status of the population, for example, by reducing the maternal mortality rate. The objectives of the Indonesian Health Referral and Hospital Programme in the 6th Five Year Development Strategy includes not only this type of health improvement target but also the need for improvement in the management of resources. PMID- 11260754 TI - The problem-based learning tutorial laboratory - a method for training medical teachers. AB - An extensive staff development program was started in 1998 in the Faculty of Medicine at the University of Helsinki. A problem-based learning method was introduced as a new style of teaching in the curriculum reform. This paper describes a teaching method 'Problem-based learning - tutorial laboratory' for training medical teachers to act as tutors and to understand their roles as facilitators of learning and the dynamics of a small group. The method was based on learning cycles: teachers had a possibility to experience tutoring, to get feedback about it from an educational expert and from a peer teacher and also they were able to reflect on their views in the group. The teachers were content with the training. Sessions improved teacher cooperation across the departments and brought new teaching ideas for shared use. It also helped to cope with the resistance related to the curriculum change process. PMID- 11260756 TI - Do you know? PMID- 11260755 TI - Comparison of a general practice and hospital placement on medical students' insights into rural practice. PMID- 11260758 TI - Jottings. PMID- 11260757 TI - What's new in medical education: items of interest to medical educators. PMID- 11260759 TI - Mild traumatic brain injury. PMID- 11260760 TI - Diagnostic criteria and differential diagnosis of mild traumatic brain injury. AB - Brain injury is classified clinically as severe, moderate or mild brain injury characteristics, including admission Glasgow coma score, duration of unconsciousness and post-traumatic amnesia and any focal neurological findings. Most traumatic brain injuries are classified as mild traumatic brain injury (MTBI). Headache, nausea and dizziness are frequent symptoms after MTBI and may continue for weeks to months after the trauma. MTBI may also be complicated by intracranial injuries. Experimental animal models and post-mortem studies have shown axonal damage and dysfunction in MTBI. This damage is mostly localized in the frontal lobes. Serum S-100 and NSE have been reported to be markers for the seventy of brain damage. In the literature, indications for radiodiagnostic evaluation following MTBI have been the subject of debate. Radiographs of the skull are used to exclude skull fractures, but are not useful for an evaluation of brain injury. Computed tomography of the brain seems to be the best way to exclude the development of relevant intracranial lesions. MTBI has a good clinical outcome, although a substantial group of patients develop post concussional complaints (PCC). There is little information on the effectiveness of various methods suggested for reducing the frequency of PCC. PMID- 11260761 TI - To do or not to do? Magnetic resonance imaging in mild traumatic brain injury. AB - Clinical quantification of mild traumatic brain injury (MTBI) patients should be based on Glasgow coma scale (GCS) score, duration of loss of consciousness (LOC) and post-traumatic amnesia (PTA). In addition, a short practicable neuropsychological test might be useful in detecting minor memory and attentional deficits. MRI appears to be the most sensitive imaging method for assessing MTBI so far, but information regarding a visualized lesion is not usually utilized in the classification of MTBI. Magnetic resonance imaging (MRI) should, therefore, play a major role in any MTBI classification scheme. An appropriate MRI protocol has to be chosen using at least T1 weighted, T2 weighted, proton density and gradient-echo (GRE) sequence images, all in at least two planes, in order to detect and classify all lesions precisely. Owing to the fact that acute lesions may be missed, it is advisable to perform MRI in the first 2 weeks following trauma. Further research is necessary to clarify the relationship between chronic symptoms after MTBI and MRI abnormalities. It may, thus, be possible to provide optimal strategies for emergency department management, to define a group of patients with a need for acute and rehabilitative intervention after MTBI, and to predict their outcome. PMID- 11260762 TI - Management of mild traumatic brain injury: lack of consensus in Europe. AB - Mild traumatic brain injury (MTBI) accounts for most traumatic brain injuries and is an important cause of morbidity. Recent studies in various European countries have shown that no consensus exists about management of patients with MTBI. This study describes the management of MTBI patients in various European hospitals. A short questionnaire covering the areas of interest was sent to several EFNS members in European countries. The results of the inquiry show that there is, at present, no consensus about criteria for, or management of MTBI in European hospitals. PMID- 11260763 TI - Factor analysis of the components of 12 standard test batteries, for unilateral spatial neglect, reveals that they contain a number of discrete and important clinical variables. AB - The aim of this study was to investigate a conventional battery of tests capable of assessing the presence of the component and extent of the lesions in patients with unilateral spatial neglect. Ninety-four patients who had unilateral spatial neglect with a stroke in right hemisphere were assessed on 12 traditional neglect batteries 4 weeks after the onset. Computerized tomography was also performed to investigate the possible anatomical relationships with each neglect battery. Factor analysis showed that the tests loaded significantly on five factors. There are not only visual scanning factors but also factors of imaging, visual judgement, visual cognition and effectiveness from left hemisphere in the unilateral spatial neglect. There are high correlations between each neuropsychological test and neglect batteries. Furthermore, lesions in the paraventricular white matter were associated with clock and person drawing tasks. Lesions in the occipital lobe were associated with reading, explaining and visual counting tasks. Lesions in the temporal lobe and the posterior limb of the internal capsule were associated with line bisection tasks. It is suggested that it is possible that there are some different components in unilateral spatial neglect. Failure in some tasks may predict different lesions in terms which include localization. PMID- 11260765 TI - Misconceptions about brain injury among the general public and non-expert health professionals: an exploratory study. AB - The purpose of this pilot study was to investigate the lack of knowledge and misconceptions concerning brain injury, as perceived by those with experience of the condition. Using a qualitative research method, 19 semi-structured interviews were conducted with brain-injured individuals, caregivers and professionals who provide social rehabilitation after brain injury. Interviews were analysed using Interpretative Phenomenological Analysis. According to participants, inaccurate and inadequate knowledge about brain injury is common among the general public and among health professionals without expertise in the field of brain injury. The major themes that emerged from the analysis were: inaccurate beliefs about recovery time and possible extent of recovery from brain injury; lack of awareness of the diversity ofproblems it can cause, particularly the existence of behavioural and cognitive sequelae; misconceptions about the capabilities of brain-injured people depending on the visibility or invisibility of their disability: and misidentification of brain-injured individuals as mentally ill or learning disabled. Results are discussed in terms of a theory of illness cognition. Posibilities for further research are discussed, and it is concluded that the results of this study could help guide future information provision to all who may come into contact with brain injury. PMID- 11260764 TI - Mood and behavioural disorders following traumatic brain injury: clinical evaluation and pharmacological management. AB - In order to investigate phamacotherapeutic responsiveness of major depression and other behavioural disturbances associated with traumatic brain injury (TBI), 20 post-TBI patients were diagnosed as being depressed by two independent neuropsychiatrist observers, out of 37 consecutive TBI subjects sent to psychiatric counselling for poor compliance during rehabilitation programmes or psychiatric/behavioural disturbances after return to society. They were subsequently divided into two subgroups, depending on time elapsed from trauma (A: within 6 months; B: at 24-36 months post-trauma) and were enrolled in an open informed pharmachological study. Rating at baseline included Glasgow Coma Score on hospital admission, length of coma, length of hospitalization, Functional Independence Measure (FIM), Brief Psychiatric Rating Scale (BPRS) and Clinical Global Impression scale (CGI). BPRS and CGI were repeated after 12 weeks of oral administration of citalopram (20 mg a day) and carbamazepine (600 mg a day). At baseline, psychiatric symptoms in group B were worse than in group A (particularly somatic overconcern, anxiety, depressed mood, psychomotor slowness, inappropriate and labile affect). At T1, the global (group A and B combined) CGI and BPRS scores showed a statistically significant improvement when compared with T0, even if group B scores remained higher than group A. The results of this study suggest that: (a) citalopram combined with carbamazepine is effective in reducing depression and behavioural disorders following TBI, and (b) these disturbances should be addressed as soon as possible during the acute rehabilitation period. PMID- 11260766 TI - Methotrimeprazine in the treatment of agitation in acquired brain injury patients. AB - Medical management of the agitation associated with acquired brain injury (ABI) has been proble matic. At least 12 distinct drugs are currently recommended in the medical literature. In recent years, on the ABI in-patient rehabilitation unit, methotrimeprazine (MTZ) has come to be the preferred drug and is used routinely for effective treatment of agitation. The objective of this paper is to describe the use and safety of MTZ in the rehabilitation of ABI patients. A retrospective chart review of all patients discharged from the ABI unit over a course of 2 years was conducted. In addition to demographics such a aetiology of ABI, sex, age, length of stay, Glasgow Coma Scale, length of posttraumatic amnesia and others, a detailed analysis was made of the multidisciplinary progress notes to determine the daily agitation status and the daily use of psychotropic medication. All notes on side effects and adverse reactions were carefully documeneted. 120 first admission recent ABI patients were discharged in the 2 year study period. Of these, 69 (57%) had some level of agitation and 56 (48%) were treated with MTZ, in doses of 2-50 mg up to four times daily. Agitation was controlled in most cases. In only two cases were significant side effects noted. While MTZ has been used as a safe and effective neuroleptic in psychiatry for over 40 years, this is the first report of its use in treating agitation in ABI. PMID- 11260767 TI - Neurobehavioural and cognitive profile of traumatic brain injury patients at risk for depression and suicide. AB - The possibility that patients who have suffered a traumatic brain injury will commit suicide is high, and in many cases clinicians tend to underestimate this possibility. In this study, 39 consecutive patients are studied through a Rorschach technique more than 1.5 years after their hospital discharge. The data show that 48.6% of the patients fulfil the criteria that classifies them as depressive, and, of these, 65% are at clinical risk to commit suicide (33.3% of the total of TBI patients); 25.6% have not met the criteria of depression or suicidal tendencies, and another 25.6% show very low suicide tendency scores. Only 15.6% of the total patients presented only depression without risk of suicide. The neurobehavioural and cognitive profile of the TBI suicide-prone patient shows an emotional person with cognitive difficulties in how they interpret reality, the person tries to understand what is happening around them, but is unable to cope. They show concrete thoughts, although they have difficulties solving problems and have few intellectual resources to cope with their surroundings. They do not know how to distance themselves from the emotional aspects of situations. PMID- 11260768 TI - Guidelines for interpreting the validity of scores on the Recognition Memory Test for use in forensic settings. PMID- 11260769 TI - Traumatic brain injury (TBI) 10-20 years later: a comprehensive outcome study of psychiatric symptomatology, cognitive abilities and psychosocial functioning. AB - The goal of this study was to measure the very long-term mental and psychosocial outcomes of severe traumatic brain injury (TBI). Seventy-six persons with severe TBI were evaluated extensively by means of standardized scales, neuropsychological tests and evaluations by family members, at an average of 14.1 (SD = 5.5) years post-injury. Six mental and functional domains were examined: psychiatric symptomatology, cognitive abilities, vocational status, family integration, social functioning, and independence in daily routines. The findings indicate a long-term differential effect of severe TBI, with seriously affected psychiatric symptomatology, family and social domains, as compared to moderately influenced cognitive, vocational and independent functioning. Relatively high rates of depression, psychomotor slowness, loneliness and family members' sense of burden were found. In addition to their epidemiological importance, the results indicate that persons with TBI and their families may need professional assistance to maintain a reasonable psychosocial quality of life, even more than a decade post-injury. PMID- 11260770 TI - Correlation of atrophy measures on MRI with neuropsychological sequelae in children and adolescents with traumatic brain injury. AB - To examine the relationship between neuropsychological sequelae and atrophy parameters from magnetic resonance imaging (MRI) following paediatric moderate-to severe traumatic brain injury (TBI), 19 head injured children and adolescents were studied at least 6 years after injury. Three-dimensional MRI scans were obtained. A semi-automatic computerized method was used to estimate ventricular volumes and the corpus callosum area. Tests of intellectual, memory, visuospatial, frontal lobe, and motor speed functioning were administered to all patients and to 19 matched normal control subjects. Patients' performance significantly differed from controls in general intellectual function, visual memory, visuospatial and frontal lobe tests. The corpus callosum area correlated strongly with several measures involving processing speed and visuospatial function. Ventricular enlargement was less related to neuropsychological outcome. In conclusion, quantitative measurement of the corpus callosum on MRI reflects neuropsychological outcome better than ventricular dilation in paediatric patients. PMID- 11260771 TI - Caregiver depression following traumatic brain injury (TBI): a consequence of adverse effects on family members? AB - Many studies have demonstrated that the behaviour of individuals with traumatic brain injury (TBI) predicts the emotional adjustment of their caregivers. The primary objective of the present study was to obtain an understanding of potential moderating and mediating variables between carer depression and analogous stressors. Seven sets of predictor variables (demographic variables, concurrent stressful life events, behavioural problems, social role problems, extent of adverse effects on family members, appraisal, and support) and the criterion variable of depression in caregivers were examined. Fifty-eight carers participated in the study at 6 months, 1 year, 2 years, or 3 years following injury. The number of adverse effects on family members (other than the informant) was the only stressor significantly related to carer depression. However, carer appraisal of adverse family effects was found to mediate the relationship between stressor and depression, and carer perception of support effectiveness was found to moderate the effect of adverse family effects on depression. Forty-six per cent of the variance in caregiver depression was accounted for by carers appraisal of adverse family effects and the interaction of adverse family effects and support effectiveness. These findings highlight the importance of supporting families as a whole in the rehabilitation of persons with TBI. PMID- 11260772 TI - Psychological distress in carers of head injured individuals: the provision of written information. AB - Since the early 1970s, researchers have expressed concern about the emotional well-being of family members after traumatic brain injury (TBI), and it is now widely acknowledged that TBI has long-term effects on the patient and relatives alike. Researchers have found a substantial number of relatives caring for head injured patients to show significant levels of anxiety and depression, and have emphasized the need for information for relatives on the prognosis of head injury. There are, however, very few studies that have investigated the usefulness of giving literature to relatives. Using a longitudinal, mixed variable, within- and between-subject design, the present study investigated the effect of an information booklet on levels of distress in a group of 34 carers of individuals with TBI. These results are discussed, and the proposal made that an information booklet such as the one used in the present study should become an integral part of the discharge procedure for relatives of individuals who have sustained a head injury. PMID- 11260773 TI - Personality and neurocognitive correlates of impulsive aggression in long-term survivors of severe traumatic brain injury. AB - This study addresses a common outcome of severe traumatic brain injury (TBI), disinhibited aggressive behaviour. This behaviour has been classified in aggression literature as 'impulsive aggression' (IA). The purpose was to: (1) characterize those TBI patients who are likely to be an aggression risk, and (2) determine if TBI patients with IA demonstrate personality style and neurocognitive performance similar to that seen in other IA groups. Participants were 45 survivors of severe TBI (26 of whom had persisting problems with IA), who were clients of a residential brain injury treatment facility. IA participants had a higher incidence of pre-morbid aggressive behaviour, were younger, had a shorter tenure in the programine, and were more impulsive, irritable, and antisocial than the non-aggressive control participants. Unlike past research, no neurocognitive differences were found. The results are discussed in terms of the conceptualization, identification, and treatment of persisting IA in severe TBI. PMID- 11260774 TI - A psychological assessment of adolescent and young adult inpatients after traumatic brain injury. AB - The purpose of this study was to evaluate the cognitive, behavioural, depressive and self-awareness disorders, and their relationships, after severe traumatic brain injury (TBI) in adolescent and young adult inpatients. Two groups of patients with (n = 83) and without (n = 103) TBI, aged 14-25 years, hospitalized after severe traumatic pathology, were compared using the clinician's report and self-report. A higher frequency of depressive tendencies in TBI patients than in controls was shown in the clinician's therapeutic attitude (i.e. prescription of antidepressant drugs), the clinician's report and the self-report. The same difference between the two groups was observed for behavioural and schooling problems in the clinician's report, but not in the self-report. Discrepancies between self- and clinician's evaluation were in favour of a lack of self awareness of behavioural and cognitive disorders among TBI patients. Correlations of depressive mood with anxiety and cognitive complaints were stronger in TBI than in non-TBI patients. Depression in TBI patients seems compatible with some degree of lack of self-awareness of cognitive and behavioural difficulties. PMID- 11260775 TI - Diagnostic confusion in mild traumatic brain injury (MTBI). Lessons from clinical practice and EFNS--inquiry. European Federation of Neurological Societies. AB - A 1997 inquiry of 130 neurosurgeons throughout Germany, dealing with diagnosis and therapy of patients with mild traumatic brain injury showed a mainly inhomogeneous picture. The European Federation of Neurological Societies inquiry form 'Management of Patients with Mild Head Injury' was sent on behalf of the German Society of Neurological Surgeons to every leading neurosurgeon in Germany, of whom only 74 (57%) answered. The diagnosis 'mild brain injury' is used by 63%, 'commotio cerebri' by 49%, and 'brain concussion' by 4% of the institutions. GCS is used for classification by 60%, PTA 48%, retrograde amnesia by 50%, and LOC by 63% of institutions. Guidelines are used in 78%. Diagnostic x-ray of the skull is used in 77%, cervical spine in 62%, CT in 66%, MRT in 7%; and routine EEG in 35%. Fourteen per cent of the patients are not admitted; home observation is used in 45% of institutions, full bedrest in 19%, working pause in 48%, pain medication in 27%, control in 51%. Seperate guidelines for children in 54% of those departments. PMID- 11260777 TI - The contribution of local distribution substations and associated area distribution system to personal exposure to power frequency magnetic fields. AB - A number of epidemiological studies has shown a significant correlation between wire coding, magnetic fields and childhood cancer, although a more recent study has not [McBride et al. (1999) Am. J. Epidemiol. 149 (9), 831-842]. In the UK there is currently no equivalent to wire-codes and there is some uncertainty about the extent to which the UK medium-voltage electricity distribution systems contribute to personal exposure and how this compares with US overhead supply systems. Studies on four different area types were carried out to measure magnetic field intensities from typical electricity supply utility substations and cabling in the vicinity of domestic housing. Typically at distances of two metres from the substations mean magnetic field intensities were 20 nano teslas (nT) or less, increasing to 0.98 microT or less at the closest public access point. The mean magnetic field exposure level sampled around the four main test areas varied between 0.012 and 0.27 microT increasing to 0.30-0.80 microT at road junctions. PMID- 11260778 TI - Use of norfloxacin in poultry production in the eastern province of Saudi Arabia and its possible impact on public health. AB - Samples of market-ready chicken muscle and liver from 32 local broiler farms were first screened for antibiotic residues by microbiological assay. The antibiotic residue-positive muscles and livers from 22 farms were further analysed for norfloxacin (NFX) residues by high performance liquid chromatography. NFX was detected in 35.0% and 56.7% of raw antibiotic-residue-positive muscles and livers, respectively. The NFX-positive muscles and livers were respectively obtained from 11 (50.0%) and 14 (63.6%) of the 22 antibiotic-residue-positive farms. Since the maximum residue limit (MRL) for NFX has not yet been fixed, the MRL for enrofloxacin was used in the study. All NFX-positive farms had mean raw tissue levels, which were 2.7- to 34.3-fold higher than the MRL. Although cooking markedly reduced NFX tissue concentrations, mean detectable levels remained above MRL in large proportions of NFX-positive samples and farms. Susceptibility patterns of Enterobacteriaceae isolates from chicken and human patients to NFX showed alarmingly high rates of resistance in chicken isolates especially among Escherichia coli (45.9%) and Pseudomonas spp. (70.6%) compared with patients' isolates (10.5% and 18.2%, respectively). The study reveals widespread misuse of NFX in the local poultry industry, which may pose a major risk to public health including possible stimulation of bacterial resistance and hypersensitivity reactions to fluoroquinolones. More prudent use of fluoroquinolones in food producing animals is therefore recommended. Further, there is a need to establish MRL values for NFX. PMID- 11260780 TI - The health promotion implications of the knowledge and attitude of employees in relation to health and safety leaflets. AB - Selected aspects of the efficacy of printed leaflets produced by a government health and safety agency and widely distributed by the enforcement bodies and other organisations to promote workplace health and safety are examined. It is based on a study of 30 small or medium-sized enterprises and examines the views of 120 employers and employees regarding the availability, attractiveness, relevance and usefulness of the leaflets and estimates the reader comprehension and readability of the selected leaflets. The results indicate that the selected leaflets are considered acceptable and comprehensible by the majority of respondents. As these are typical of the leaflets available in the health and safety field this is a positive outcome. The discussion focuses around the ability of the leaflets to engage and to inform and suggestions are made to encourage a wider debate on the criteria which contribute to these two aspects of leaflet use. It is contended that leaflets will continue to be important in the attempts of those involved in workplace health and safety to facilitate learning and to contribute to the overall process of behaviour change. This study raises a number of key issues regarding the future design and use of such leaflets. PMID- 11260779 TI - Socio-environmental determinants of the leptospirosis outbreak of 1996 in western Rio de Janeiro: a geographical approach. AB - The environmental and social context in which a leptospirosis outbreak took place during the summer of 1996 in the Rio de Janeiro Western Region was examined by using spatial analysis of leptospirosis cases merged with population and environmental data in a Geographical Information System (GIS). Important differences were observed between places where residences of leptospirosis cases are concentrated and other places in the region. Water supply coverage, solid waste collection, sewerage system coverage and flood risk area were the main determining variables from an initial list of ten. The influence of these unfavorable social and environmental factors is verified hundreds of meters distant from the leptospirosis case residences, demonstrating a necessity to broaden the area of health surveillance practices. The geocoding indicated that some cases did not report contact with flood water, even though they were geographically adjacent to cases who did report this contact. Cases may only report exposures they believe are related to the disease. Geocoding is a useful tool for evaluating such bias in the exposure recall. PMID- 11260781 TI - Blood lead levels and calcium intake in Mexico City children under five years of age. AB - OBJECTIVE: The relationship between daily calcium intake and blood lead levels was evaluated among children under five years of age living in Mexico City. METHODS: A random sample of 200 children under five years of age, resident in two neighborhoods of Mexico City was selected: Xalostoc, an industrial neighborhood, and Tlalpan, a residential neighborhood (100 from each area). The mothers of these children filled out a questionnaire on predictors of blood lead levels including daily calcium intake. Lead levels were determined from the venous blood samples. Calcium intake was assessed using a short Food Frequency Questionnaire including 11 food items that accounted for 95% of calcium intake in Mexico. RESULTS: The average blood lead level was 9.93 microg dl(-1) (range 1-31 microg dl(-1)). An inverse relationship was observed between blood lead levels and daily calcium intake. This relationship was statistically significant among children aged 13 months-5 years. CONCLUSION: The results suggest that calcium provided a protective effect against lead accumulation in the body among children. Further studies should be undertaken to evaluate this hypothesis through experimental design. PMID- 11260782 TI - Evaluation of oxidative stress in some cases of argimone oil poisoning during a recent outbreak of epidemic dropsy in India. AB - The study was designed to evaluate the oxidative stress and modulation of anti oxidant enzymes in 10 accidental argimone oil poisoning cases admitted in a hospital in Delhi, India during a recent outbreak of epidemic dropsy in 1998. Serum malondialdehyde (MDA) level, oxygen free-radical scavenging enzymes such as superoxide dismutase (SOD) and catalase (CAT), and glutathione (GSH) and related enzymes, e.g. glutathione reductase (GR), glutathione peroxidase (GPx), gamma glutamyl transpeptidase (GGT) and glutathione-S-transferase (GST) in erythrocytes were assayed. The sanguinarine level in serum was measured by high-performance liquid chromatography. The serum MDA level was higher and the GSH level in erythrocytes was lower in argimone oil poisoning cases than those in controls. There was a significant decrease in SOD and GPx activities in erythrocytes of epidemic dropsy cases but no changes were observed in CAT, GR and GST assay. The depletion of GSH in erythrocytes, serum MDA level and clinical severity were dependent on serum sanguinarine level. The results indicate that sanguinarine (argimone oil) poisoning creates an oxidative stress in humans. The oxidative stress and differential modulation of anti-oxidant enzymes by sanguinarine might play a pathogenic role in epidemic dropsy, which suggests the incorporation of anti-oxidant drugs in the treatment protocol of the disease. PMID- 11260783 TI - Bacteriological and virological quality of seawater bathing areas along the Tyrrhenian coast. AB - Monitoring was carried out during summer 1997 along a selected area of the Tyrrhenian coast near the Tiber river mouth. Fifty-eight seawater samples, collected from 19 stations, were examined for coliforms, streptococci, Enteroviruses, Salmonellae, coliphages, Bacteroides fragilis phages, Pseudomonas, alophilic Vibrios, Aeromonas and yeasts. Salmonellae and coliphages were isolated in 3 and 12 out of 58 samples, respectively. Enteroviruses and Bacteroides fragilis phages were not isolated. Reoviruses were isolated only from 2 out of 58 samples. A limited number of samples of the northern stations located near the Tiber and other river mouths exceeded the guide values for bathing water by the EU Directive. All the southern stations, located near canals, were of very good microbiological quality. Pseudomonas, Vibrio, Aeromonas and yeasts were isolated from all stations and their values in 100 ml of seawater were 10-10(6), 10-10(6), 0-10(6) and 1-10(3), respectively. An extensive disinfection practice carried out on domestic wastes, which are discharged in rivers and canals, probably brought pollution levels of most stations to values within the bacterial standards. The spread of Pseudomonas, Aeromonas, etc. showed that all the coastal area studied was characterized by the presence of organic matter coming from land that can support the presence of opportunistic pathogens and other microbial flora. PMID- 11260784 TI - Seasonal and annual variations in air concentrations of Pb, Cd and PAHs in Cairo, Egypt. AB - A study is presented concerning the degree and extent of air pollution by cadmium, lead and polycyclic aromatic hydrocarbons (PAHs) in atmospheric particulates of an urban area (El-Abasya) in Cairo, Egypt. The average yearly and seasonal concentrations are presented. The correlation coefficients between metals, and between metals and suspended PAHs are also reported. Statistical analysis showed seasonal and annual variations. Concentrations of PAHs, Pb and Cd were higher in winter than those measured at other times in the study area. The concentration of lead decreased by 40% from 3.36 microg(-3) in 1994 to 2.4 microg m(-3) in 1997 in consequence of the reduction of the Pb concentration in leaded gasoline fuel. The average cadmium concentrations showed significant decrease in 1997. The sum of the concentrations of PAHs was higher during the winter season 1997 (14.79 ng m(-3)) than in summer (7.53 ng m(-3)) and a highly significant difference was observed between the two seasons. In addition, the ratio of benzo(a)pyrene [B(a)P] to Pb was higher in winter than in summer, suggesting the contribution of a non-vehicular source of lead. The levels of lead observed are higher than those recorded in various other parts of the world. PMID- 11260785 TI - Skin irritation in users of brominated pools. AB - This study investigated adverse skin and eye effects in swimmers using pools with three different disinfection systems (chlorine, chlorine/ozone and bromine/ozone) and monitored water quality parameters that may be related to adverse health effects. A cross-sectional study of 770 children swimming in three school pools was carried out over a 4 week period in November 1994 using a postal questionnaire. Physico-chemical and bacteriological parameters of water quality were monitored on a weekly basis. Responses were obtained for 385 swimmers. Skin rashes with an onset less than 24 h after swimming in the school pool were reported by 4-8% of swimmers. Compared with the bromine/ozone pool, the odds ratio (OR) of having a rash that started less than 24 h after pool use was 1.91 (CI 0.71-5.10) for the chlorine pool and 1.88 (CI 0.61-5.81) for the chlorine/ozone pool. Adjustment for possible confounders made no significant differences to these results. Eye redness, itch or irritation was reported by 23 33% of swimmers and 24% of non-swimmers, and wearing swimming goggles had a protective effect (OR 0.40; CI 0.24, 0.65). Disinfectant levels were more consistently maintained in the pools with automatically controlled systems. The bromine disinfection system was not associated with a greater risk of the development of skin rashes than other disinfection systems, but the numbers were small, and need to be interpreted with caution. PMID- 11260786 TI - The impact of pollution from a mercury processing plant in KwaZulu-Natal, South Africa, on the health of fish-eating communities in the area: an environmental health risk assessment. AB - The objective of this study was to determine the human health risk associated with fish consumption in a contaminated area downstream from a mercury processing plant in KwaZulu-Natal, South Africa. The study population consisted of fish consumers living in close proximity to the u'Mgeni River and the Inanda Dam downstream from the plant. A control group was selected from the area upstream from the mercury plant as far as the Nagle Dam. Total daily mercury consumption per kilogram body weight per day was calculated for each person included in the study. These data were compared with the tolerable daily intake standard published by the World Health Organization, as well as to the United States Environmental Protection Agency's reference dose. Human hair samples obtained from the study population and a control group were analysed for mercury content. The results of the risk estimation indicated that the study population is at risk. Human hair samples, however, indicated that dangerous levels of mercury had not yet been consumed. Humans in this study area could be subject to an excessive health risk from mercury as a result of their fish consumption. Fish mercury levels in the contaminated area should be monitored closely. PMID- 11260787 TI - The role of environmental health officers in the protection of allergic consumers. AB - Those suffering from serious food allergy are advised to avoid contact with the trigger allergens. However, it can prove difficult to avoid such food allergens, especially those hidden as unnamed components of composite foods. Many cases of serious illness and death following the accidental consumption of trigger foods, such as peanuts and other nuts, have been noted in the literature. This study reports the findings of a survey of the role of environmental health officers (EHOs) in Northern Ireland, in relation to the protection of the allergic consumer. This study shows that although EHOs use hazard analysis as a general method of improving food safety, this approach is not currently being applied in the control of the allergenic risks posed to sensitive consumers, consuming nut and peanut-containing foodstuffs. This study has established that the main reasons for not applying HACCP systems in the reduction of this risk are related to concerns in relation to lack of knowledge and appropriate training in this subject. PMID- 11260789 TI - Effects of oil plastic extract on mice. AB - In the plastics industry, various chemical additives are used to improve certain properties of plastics. Some of these chemicals, that might be toxic, have been proved to leach from the plastic containers and mix with their contents such as food oils, beverages, drugs, etc. Locally manufactured polyvinyl chloride (PVC) jerrycans were bought from the market and cut into small chips of 0.5 cm in the larger dimension. Four grams of chips were extracted with 20 ml cottonseed oil in the autoclave for 1 h at 121 degrees C. The extract was prepared daily and given orally to adult MFI mice in a dose of 10 ml kg(-1) body weight. Pure cottonseed oil was prepared under the same conditions and given in a dose of 10 ml kg(-1) body weight to the control group. Treatment of both groups continued for 1 month. Each group comprised 60 animals, regardless of sex. Effects of the oil plastic extract were observed on blood elements, serum transaminases (aspartate aminotransferase, AST, and alanine aminotransferase, ALT), organ-to-body weight of the liver, kidneys and brain, and the nervous system (effects on the neuromuscular junction and analgesia, using the Rota-Rod(R) treadmill 'RRT', and the hot plate, respectively). All the results were subjected to Student's t-test. The results showed that the extract induced significant effects: an increase in the activities of AST (p< 0.001) and ALT (p< 0.02), an increase in the mean corpuscular haemoglobin concentration (MCHC) (p< 0.01), and the monocyte count (p< 0.01). It decreased the white blood cell count (WBC) (p< 0.01), the mean corpuscular volume (MCV) (p< 0.05), and the lymphocyte count (p< 0.05). It also reduced the weight of the liver (P< 0.01), kidneys (P< 0.05), and the endurance time on the RRT (p< 0.001). PMID- 11260788 TI - Heavy metal hazards of Asian traditional remedies. AB - In recent years there has been an increase in the use of traditional Asian medicines. It is estimated that 30% of the US population is currently using some form of homeopathic or alternative therapy at a total cost of over $13 billion annually. Herbal medications are claimed and widely believed to be beneficial; however, there have been reports of acute and chronic intoxications resulting from their use. This study characterizes a random sampling of Asian medicines as to the content of arsenic, mercury, and lead. Traditional herbal remedies were purchased in the USA, Vietnam, and China. The Asian remedies evaluated contained levels of arsenic, lead, and mercury that ranged from toxic (49%) to those exceeding public health guidelines for prevention of illness (74%) when consumed according to the directions given in or on the package. Heavy metals contained in Asian remedies may cause illness of unknown origin and result in the consumption of health care resources that are attributable to other causes. The public health hazards of traditional herbal Asian remedies should be identified and disclosed. PMID- 11260790 TI - Decanting versus sterile pre-filled nutrient containers--the microbiological risks in enteral feeding. AB - A simulated ward study was carried out to compare the microbiological risk of assembling and running four different enteral feeding systems for 24 h. Assembly was carried out, (i) according to manufacturers' instructions with hands either covered by disposable gloves or deliberately contaminated with a test organism, or (ii) touching both the nutrient container top and pump set connector with hands deliberately contaminated with K. aerogenes. Two of the systems were ready to-hang types (pack and bottle), the other two required feed to be decanted from either bottles or cans. When manufacturers' instructions were followed and disposable gloves worn, organisms were only detected in feeds decanted from cans and at levels < or = 20 cfu ml(-1). However, when systems were assembled following manufacturers' instructions, but with contaminated hands, no organisms were found in either of the ready-to-hang systems but average bacterial counts in samples from systems where the feed was decanted from bottles were 1.8 x 10(3) cfu ml(-1) at 24 h and 9.3 x 10(5) cfu ml(-1) for those where feed was decanted from cans. When systems were deliberately touched with contaminated hands, no organisms were detected in any feed samples from the pack system at 24 h, while bacterial counts for the other three systems ranged from 10(1) to 10(5) cfu ml( 1). The results highlight the important role played by system design in reducing both the level and incidence of bacterial contamination of enteral tube feeds and indicate that ready-to-hang feeding systems should be the preferred choice. However, if decanting of feeds cannot be avoided then strict adherence to manufacturers' instructions and the use of disposable gloves is to be advised. PMID- 11260791 TI - Bacteriological profile and holding temperatures of street-vended foods from Addis Ababa. AB - A total of 150 samples of street foods comprising 30 each of 'sambussa', 'macaroni', 'lentil sandwich', 'kitfo' (raw minced meat) and 'egg sandwich' were examined for the relation between their holding temperatures and bacteriological profile. Over 90% of the samples were stored within a temperature range of 15.5 34.5 degrees C. At this holding temperature counts of coliforms and members of the Enterobacteriaceae were below detectable levels in 'sambussa' samples. In 'macaroni', 'lentil sandwich', 'kitfo' and 'egg sandwich' these counts were higher than 10(6) cfu g(-1) in most cases. Over 70% of the street food samples had aerobic mesophilic counts higher than 10(7) cfu g(-1). Of the total of 1552 bacterial strains isolated from the different food samples, Bacillus spp. (29.1%), staphylococci (22.8%) and micrococci (15.4%) were among the dominant groups. Members of the family Enterobacteriaceae (14.5%) were also frequently encountered. PMID- 11260792 TI - Verotoxin-producing Escherichia coli--an environment-induced emerging zoonosis in and around Calcutta. AB - With changes in livestock management practices and food processing industry, along with changes in people's food habits, many diseases have emerged. Infection with verotoxin-producing Escherichia coli (VTEC) is one such illness. In the present study an attempt was made to isolate, identify and characterize VTEC strains with reference to the O157:H7 serotype from animal, human sources and some food products with the aid of the available modern methods. A total of 876 samples (330 animal, 184 human, 362 food samples) were screened for the presence of VTEC by conventional as well as PCR technique. Seventeen VTEC strains (12 animal, one human and four food samples) were isolated. The isolation rate was higher in diarrhoeic animals (6.02%), followed by diarrhoeic handler (3.12%) and raw beef (1.78%) samples. All strains showed the presence of the VT gene by PCR tests and were uniformly sensitive to common antibiotics except tetracycline, cephalexin, dicloxacillin, erythromycin and lincomycin. Since all strains were isolated from various sources of animal and human origin and all strains showed the presence of the VT gene and uniform antibiogram, a zoonotic association is suggested. This study marks the first report of isolation of VTEC strains from animal sources in India. PMID- 11260793 TI - The structural basis of protein folding and its links with human disease. AB - The ability of proteins to fold to their functional states following synthesis in the intracellular environment is one of the most remarkable features of biology. Substantial progress has recently been made towards understanding the fundamental nature of the mechanism of the folding process. This understanding has been achieved through the development and concerted application of a variety of novel experimental and theoretical approaches to this complex problem. The emerging view of folding is that it is a stochastic process, but one biased by the fact that native-like interactions between residues are on average more stable than non-native ones. The sequences of natural proteins have emerged through evolutionary processes such that their unique native states can be found very efficiently even in the complex environment inside a living cell. But under some conditions proteins fail to fold correctly, or to remain correctly folded, in living systems, and this failure can result in a wide range of diseases. One group of diseases, known as amyloidoses, which includes Alzheimer's and the transmissible spongiform encephalopathies, involves deposition of aggregated proteins in a variety of tissues. These diseases are particularly intriguing because evidence is accumulating that the formation of the highly organized amyloid aggregates is a generic property of polypeptides, and not simply a feature of the few proteins associated with recognized pathological conditions. That such aggregates are not normally found in properly functional biological systems is again a testament to evolution, in this case of a variety of mechanisms inhibiting their formation. Understanding the nature of such protective mechanisms is a crucial step in the development of strategies to prevent and treat these debilitating diseases. PMID- 11260794 TI - Quality control in the secretory assembly line. AB - As a rule, only proteins that have reached a native, folded and assembled structure are transported to their target organelles and compartments within the cell. In the secretory pathway of eukaryotic cells, this type of sorting is particularly important. A variety of molecular mechanisms are involved that distinguish between folded and unfolded proteins, modulate their intracellular transport, and induce degradation if they fail to fold. This phenomenon, called quality control, occurs at several levels and involves different types of folding sensors. The quality control system provides a stringent and versatile molecular sorting system that guaranties fidelity of protein expression in the secretory pathway. PMID- 11260795 TI - Folding defects in fibrillar collagens. AB - Fibrillar collagens have a long triple helix in which glycine is in every third position for more than 1000 amino acids. The three chains of these molecules are assembled with specificity into several different molecules that have tissue specific distribution. Mutations that alter folding of either the carboxy terminal globular peptides that direct chain association, or of the regions of the triple helix that are important for nucleation, or of the bulk of the triple helix, all result in identifiable genetic disorders in which the phenotype reflects the region of expression of the genes and their tissue-specific distribution. Mutations that result in changed amino-acid sequences in any of these regions have different effects on folding and may have different phenotypic outcomes. Substitution for glycine residues in the triple helical domains are among the most common effects of mutations, and the nature of the substituting residue and its location in the chain contribute to the effect on folding and also on the phenotype. More complex mutations, such as deletions or insertions of triple helix, also affect folding, probably because of alterations in helical pitch along the triple helix. These mutations all interfere with the ability of these molecules to form the characteristic fibrillar array in the extracellular matrix and many result in intracellular retention of abnormal molecules. PMID- 11260796 TI - Nuclear magnetic resonance characterization of peptide models of collagen-folding diseases. AB - Misfolding of the triple helix has been shown to play a critical role in collagen diseases. The substitution of a single Gly by another amino acid breaks the characteristic repeating (Gly-X-Y)n sequence pattern and results in connective tissue disease such as osteogenesis imperfecta. Nuclear magnetic resonance (NMR) studies of normal and mutated collagen triple-helical peptides offer an opportunity to characterize folding and conformational alterations at the substitution site, as well as at positions upstream and downstream of a Gly mutation. The NMR studies suggest that the local sequences surrounding the substitution site, and the renucleation sequences N-terminal to and adjacent to the substitution site, may be critical in defining the clinical phenotype of osteogenesis imperfecta. These studies may pave the way to understanding the mechanism by which a single Gly substitution in collagen can lead to pathological conditions. PMID- 11260797 TI - Investigating protein conformation-based inheritance and disease in yeast. AB - Our work supports the hypothesis that a protein can serve as an element of genetic inheritance. This protein-only mechanism of inheritance is propagated in much the same way as hypothesized for the transmission of the protein-only infectious agent in the spongiform encephalopathies; hence these protein factors have been called yeast prions. Our work has focused on [PSI(+)], a dominant cytoplasmically inherited factor that alters translational fidelity. This change in translation is produced by a self-perpetuating change in the conformation of the translation-termination factor, Sup35. Most recently, we have determined that new elements of genetic inheritance can be created by deliberate genetic engineering, opening prospects for new methods of manipulating heredity. We have also uncovered evidence that other previously unknown elements of protein-based inheritance are encoded in the yeast genome. Finally, we have begun to use yeast as a model system for studying human protein folding diseases, such as Huntington's disease. Proteins responsible for some of these diseases have properties uncannily similar to those that produce protein-based mechanisms of inheritance. PMID- 11260799 TI - The molecular biology of prion propagation. AB - Prion diseases such as Creutzfeldt-Jakob disease (CJD) in humans and scrapie and bovine spongiform encephalopathy (BSE) in animals are associated with the accumulation in affected brains of a conformational isomer (PrP(Sc)) of host derived prion protein (PrP(C)). According to the protein-only hypothesis, PrP(Sc) is the principal or sole component of transmissible prions. The conformational change known to be central to prion propagation, from a predominantly alpha helical fold to one predominantly comprising beta structure, can now be reproduced in vitro, and the ability of beta-PrP to form fibrillar aggregates provides a plausible molecular mechanism for prion propagation. The existence of multiple prion strains has been difficult to explain in terms of a protein-only infectious agent but recent studies of human prion diseases suggest that strain specific phenotypes can be encoded by different PrP conformations and glycosylation patterns. The experimental confirmation that a novel form of human prion disease, variant CJD, is caused by the same prion strain as cattle BSE, has highlighted the pressing need to understand the molecular basis of prion propagation and the transmission barriers that limit their passage between mammalian species. These and other advances in the fundamental biology of prion propagation are leading to strategies for the development of rational therapeutics. PMID- 11260798 TI - Prions and the lymphoreticular system. AB - Following intracerebral or peripheral inoculation of mice with scrapie prions, infectivity accumulates first in the spleen and only later in the brain. In the spleen of scrapie-infected mice, prions were found in association with T and B lymphocytes and to a somewhat lesser degree with the stroma, which contains the follicular dendritic cells (FDCs) but not with non-B, non-T cells; strikingly, no infectivity was found in lymphocytes from blood of the same mice. Transgenic PrP knockout mice expressing PrP restricted to either B or T lymphocytes show no prion replication in the lymphoreticular system. Therefore, splenic lymphocytes either acquire prions from another source or replicate them in dependency on other PrP-expressing cells. The essential role of FDCs in prion replication in spleen was shown by treating mice with soluble lymphotoxin-beta receptor, which led to disappearance of mature FDCs from the spleen and concomitantly abolished splenic prion accumulation and retarded neuroinvasion following intraperitoneal scrapie inoculation. PMID- 11260800 TI - Prion protein interconversions. AB - The transmissible spongiform encephalopathies (TSEs), or prion diseases, remain mysterious neurodegenerative diseases that involve perturbations in prion protein (PrP) structure. This article summarizes our use of in vitro models to describe how PrP is converted to the disease-associated, protease-resistant form. These models reflect many important biological parameters of TSE diseases and have been used to identify inhibitors of the PrP conversion as lead compounds in the development of anti-TSE drugs. PMID- 11260801 TI - Pathogenesis, diagnosis and treatment of systemic amyloidosis. AB - Amyloidosis is a disorder of protein folding in which normally soluble proteins are deposited as abnormal, insoluble fibrils that disrupt tissue structure and cause disease. Although about 20 different unrelated proteins can form amyloid fibrils in vivo, all such fibrils share a common cross-beta core structure. Some natural wild-type proteins are inherently amyloidogenic, form fibrils and cause amyloidosis in old age or if present for long periods at abnormally high concentration. Other amyloidogenic proteins are acquired or inherited variants, containing amino-acid substitutions that render them unstable so that they populate partly unfolded states under physiological conditions, and these intermediates then aggregate in the stable amyloid fold. In addition to the fibrils, amyloid deposits always contain the non-fibrillar pentraxin plasma protein, serum amyloid P component (SAP), because it undergoes specific calcium dependent binding to amyloid fibrils. SAP contributes to amyloidogenesis, probably by stabilizing amyloid fibrils and retarding their clearance. Radiolabelled SAP is an extremely useful, safe, specific, non-invasive, quantitative tracer for scintigraphic imaging of systemic amyloid deposits. Its use has demonstrated that elimination of the supply of amyloid fibril precursor proteins leads to regression of amyloid deposits with clinical benefit. Current treatment of amyloidosis comprises careful maintenance of impaired organ function, replacement of end-stage organ failure by dialysis or transplantation, and vigorous efforts to control underlying conditions responsible for production of fibril precursors. New approaches under development include drugs for stabilization of the native fold of precursor proteins, inhibition of fibrillogenesis, reversion of the amyloid to the native fold, and dissociation of SAP to accelerate amyloid fibril clearance in vivo. PMID- 11260803 TI - Tobacco/Nicotine Scientific Meetings. PMID- 11260802 TI - From genetics to pathology: tau and alpha-synuclein assemblies in neurodegenerative diseases. AB - The most common degenerative diseases of the human brain are characterized by the presence of abnormal filamentous inclusions in affected nerve cells and glial cells. These diseases can be grouped into two classes, based on the identity of the major proteinaceous components of the filamentous assemblies. The filaments are made of either the microtubule-associated protein tau or the protein alpha synuclein. Importantly, the discovery of mutations in the tau gene in familial forms of frontotemporal dementia and of mutations in the alpha-synuclein gene in familial forms of Parkinson's disease has established that dysfunction of tau protein and alpha-synuclein can cause neurodegeneration. PMID- 11260805 TI - What's new in Nicotine & Tobacco Research? PMID- 11260806 TI - Evaluation of the brief questionnaire of smoking urges (QSU-brief) in laboratory and clinical settings. AB - A brief, 10-item version of the Questionnaire of Smoking Urges (QSU; Tiffany & Drobes, British Journal of Addiction 86:1467-1476, 1991) was administered to 221 active cigarette smokers in a laboratory setting (Study 1) and to 112 smokers enrolled in a comprehensive smoking cessation program (Study 2). In the laboratory setting, craving to smoke was evaluated in response to neutral and smoking-related stimuli. In the clinical setting, craving was assessed prior to cessation and again during treatment. Factor analyses revealed that a two-factor solution best described the item structure of the QSU-Brief across conditions. Factor 1 items reflected a strong desire and intention to smoke, with smoking perceived as rewarding for active smokers. Factor 2 items represented an anticipation of relief from negative affect with an urgent desire to smoke. The findings were consistent with the expressions of craving found in the 32-item version of the QSU (Tiffany & Drobes, 1991). Regression analyses demonstrated stronger baseline mood intensity and self-reported tendency to smoke to achieve pleasurable effects and to experience the desire to smoke when cigarettes are unavailable were predictive of general levels of craving report in active smokers in the laboratory and clinical setting. The findings supported a multidimensional conceptualization of craving to smoke and demonstrated the utility of a brief multidimensional measure of craving. PMID- 11260807 TI - Nicotine enhances latent inhibition and ameliorates ethanol-induced deficits in latent inhibition. AB - Alcohol and nicotine are drugs of abuse that are used frequently together. One possible explanation for this co-administration is that nicotine prevents or lessens alcohol-associated impairments. The present study examined the dose dependent effects of acute administration of nicotine, alcohol, or alcohol plus nicotine on latent inhibition as measured by lick suppression in C57BL/6 mice. Alterations in a lick suppression ratio were measured by assessing the effects of 10 pre-exposures to an auditory conditioned stimulus (CS) on formation of subsequent CS-shock unconditioned stimulus (US) associations. Mice pre-exposed to the CS were expected to develop a weaker CS-US association. Nicotine administered prior to pre-exposure to the CS produced increased suppression ratios, ethanol given prior to pre-exposure to the CS decreased suppression ratios, and nicotine reversed the effects of ethanol when the two drugs were co-administered. These opposing actions of nicotine and ethanol may have relevance to the high incidence of smoking and drinking in humans. PMID- 11260808 TI - Birth and first-year costs for mothers and infants attributable to maternal smoking. AB - Maternal smoking during pregnancy has been linked to high costs. This study estimates the magnitude of excess costs attributable to smoking during pregnancy for mothers and infants. The model estimates smoking-attributable costs for 11 infant and maternal conditions. From a claims database of 7784 mothers and 7901 infants who had deliveries during 1996, we estimated total cost over the infants' first year of life for each mother and infant and identified each complication of interest, based on ICD-9 codes. The average cost for smokers and non-smokers could not be computed directly because smoking status is not available in claims data. Therefore, the population attributable risk percentage (PAR%) due to smoking for each complication was identified from the literature. Multiple linear regression was used to provide estimates of the incremental cost associated with each smoking-related complication. The total cost attributable to smoking was computed as a function of the incremental cost of each complication and the PAR% for each complication. The conditions associated with the largest incremental costs compared to patients without those conditions were abruptio placenta ($23,697) and respiratory distress syndrome ($21,944). Because they were more common, the conditions with the largest smoking-attributable cost were low birth weight ($914) and lower respiratory infection ($428). The sum of the additional costs attributable to smoking for all conditions yielded a total in the first year after birth ranging from $1142 to $1358 per smoking pregnant woman. It was concluded that maternal smoking during pregnancy results in an economic burden to payers and society. These estimates may be useful in formal cost-effectiveness evaluations of individual smoking cessation strategies. PMID- 11260809 TI - Individual differences in responses to the first cigarette following overnight abstinence in regular smokers. AB - We utilized cluster analysis to identify individual differences in response to the initial effects of smoking following overnight abstinence among 183 regular smokers. Participants smoked three cigarettes (1 mg nicotine, spaced 30 min apart) in standardized fashion and completed questionnaires about their subjective responses to each cigarette. Heart rate was monitored throughout the procedure. Participants were grouped into two clusters based on their reported subjective effects and heart rate changes to the first cigarette. Clusters differed in terms of greater increases in heart rate, reports of dizziness, sweating, unpleasantness, nausea, and buzzing sensations in one group compared to the other group. The smokers showing increased responses developed greater acute tolerance to the effects of smoking subsequent cigarettes on subjective negative effects and heart rate, and experienced greater negative affect after quitting. These results are partially consistent with a nicotine sensitivity interpretation or a tolerance model of the effects of initial smoking. PMID- 11260810 TI - Differences among African American light, moderate, and heavy smokers. AB - This study examined differences in demography, behavior, attitude, and physician intervention among African American light, moderate, and heavy smokers. Data were derived from an intervention study designed to assess whether a smoking status stamp would increase screening for smoking status and cessation counseling by physicians. Current analysis included 879 African American smokers categorized into three groups: light (<10 cigarettes/day), moderate (10-19 cigarettes/day), or heavy (> or =20 cigarettes/day) smokers based on number of cigarettes per day smoked. Light smokers constituted 40% of study sample, 33% were moderate smokers, and 27% were heavy smokers. Light smokers were more likely to be female (p<0.001) and have a shorter smoking history (p<0.001). Light smokers were not different in age (p=0.334), or the number of previous quit attempts (p=0.551). Although light smokers were more likely than moderate and heavy smokers to be preparing to quit (p<0.001), they were less likely to be asked their smoking status (p=0.031) or told to arrange follow-up for smoking cessation (p=0.034) by their physicians. Many African American smokers are light smokers. Light smokers are more likely to be female and have a shorter smoking history. Despite their readiness to quit, compared to heavier smokers, African American light smokers are asked about smoking less often by their physicians. Programs are needed to enhance physician intervention in this understudied population of smokers. PMID- 11260811 TI - Characterizing concerns about post-cessation weight gain: results from a national survey of women smokers. AB - Differences among adult women smokers with differing levels of concern about post cessation weight gain were investigated in a national random-digit-dialing survey. To avoid defining weight concerns in terms of possible etiologies or contributory factors, respondents were stratified using a single item querying concern about post-cessation weight gain; 39% described themselves as very concerned (VC), 28% as somewhat concerned (SC), and 33% as not concerned (NC). Significant between-groups differences were detected for measures of weight and body image, eating patterns and weight control practices, and nicotine dependence, but not for depression. Differences, primarily between VC and NC, were also detected for several weight-related smoking variables, including importance of weight as a factor in initiation, smoking as a weight control strategy, increased appetite and weight gain as withdrawal symptoms, willingness to gain weight upon quitting, self-efficacy about relapse in the face of weight gain, and readiness to quit smoking. Most differences persisted even after adjusting for body mass index and nicotine dependence. Although the importance of thinness was rated higher by weight-concerned women, the difference did not reach significance. Rather, what differentiated groups was the importance of overall body image, suggesting a larger pattern of preoccupation with body image that may not be captured by queries about weight concerns alone. We conclude that weight concerned women smokers will be especially unlikely to seek treatment or attempt self-quitting; and that redirecting attention to other aspects of body image is likely to be more helpful than attempting to divert attention away from body image. PMID- 11260812 TI - Determinants of environmental tobacco smoke in a population of Puerto Rican children. AB - This study was designed to determine among various personal, socioeconomic, and environmental factors those which had the greatest influence on exposure to environmental tobacco smoke (ETS) in a population of children residing in a tropical environment and to compare these results with those obtained in the literature of tobacco exposed children in temperate climates. Urine specimens were collected from 606 healthy Puerto Rican children (2-12 years) living in an industrial area and analyzed for cotinine, a quantitative biomarker for exposure to ETS. Parents completed a questionnaire covering smoking habits and socioeconomic information. Seventy per cent of the children were reported to be exposed to ETS, 50% resulting from exposure to smoke from either or both parents. Major determinants to ETS exposure were found to be presence of smoker, number of smokers, identity of smoker, number of cigarettes smoked in the household and child age with the youngest children suffering twice the exposure of older children. Non-determinants were exposure to smoke other than from the parent, sex of the child, season of the year and several socioeconomic factors including civil and employment status of the mother, mother's age and educational background and whether food stamps were being received. Results of a multiple regression analysis showed that our predictors accounted for 40% of cotinine appearing in the urine. Reasons for this relatively low value may be due in part to precision of our analytic method and lower levels of ambient smoke in our population vs. others that reported higher R(2) values. Predictions from questionnaire information for high ETS exposure were not always the same as those indicated by urinary cotinine emphasizing that the bioindicator, which indicates the actual inhalation of ETS, is a better predictor of exposure than responses from a questionnaire. PMID- 11260813 TI - Two behavioral treatments for smoking reduction: a pilot study. AB - This study compared two behavioral treatments on their efficacy and acceptability in reducing smoking, using a crossover design with interposed return to baseline, 2 weeks/condition. 20 US cigarette smokers reporting an interest in reducing, but not quitting, their smoking either increased the inter-cigarette interval or selected easy cigarettes to eliminate, with a goal of 50% reduction. Nicotine gum accompanied both treatments. Measurements taken were self-reported cigarettes per day, carbon monoxide, cotinine, and thiocyanate; self-rated acceptability of the treatments; adverse events. Both treatments decreased self-reported cigarettes per day (-45% and -38%) and carbon monoxide (-20% and -19%), but not cotinine or thiocyanate. Increasing the inter-cigarette interval produced slightly more reduction in cigarettes per day than cigarette selection, but no other differences were found. Both treatments were acceptable and safe. Although our sample size was small and the duration of reduction documented short, both treatments appear to be acceptable and efficacious behavioral treatments for reduction. PMID- 11260814 TI - Subjective and discriminative stimulus effects of two de-nicotinized cigarettes with different tar yields. AB - The role of tar yield in the subjective and discriminative stimulus effects of cigarette smoking was examined. Current smokers (n=18) smoked two non-nicotine cigarettes with FTC yields of 0.06 mg nicotine and 12.4 mg (low tar) or 17.9 mg tar (high tar). Physiological measures and visual analog scales were completed over a 30-min period. Dosing order was determined randomly and counterbalanced. After sampling both cigarettes, volunteers smoked a third, test cigarette. Half of the volunteers received the low-tar cigarette and half the high-tar cigarette. Volunteers identified the test cigarette (i.e., A or B) at 5, 30, 60, 300, and 900 s after the first puff. Eight of the eighteen participants correctly identified the test cigarette on 4/5 of trials. No significant changes in visual analog scale scores were found among the non-discriminators. However, among discriminators, the low-tar cigarette produced significant positive effects including good drug effects and stimulation relative to the high-tar cigarette. Relative to the low-tar cigarette, the high-tar cigarette produced negative effects including harshness, heaviness, and intensity of flavor. Average tar yield of these participants' usual cigarettes was 9.75 mg, lower than that of the low-tar cigarette used here, possibly accounting for their greater liking of the low-tar cigarette. No changes in blood pressure or heart rate were observed and both cigarettes produced similar carbon monoxide increases, indicating similar depth of inhalation when smoking each. Results suggest cigarette tar yields may play a role in cigarette smoking preferences. Further research is needed to verify whether preferences are maintained after associations with nicotine delivery are extinguished. PMID- 11260815 TI - US physicians' treatment of smoking in outpatients with psychiatric diagnoses. AB - A 1996 American Psychiatric Association (APA) guideline recommends the routine treatment of smoking for patients with psychiatric diagnoses. This study evaluates how often US physicians identified and treated smoking among these patients in the ambulatory setting just prior to publication of this guideline, by analysis of 1991-1996 data from the National Ambulatory Medical Care Survey, an annual survey of a random sample of US office-based physicians. Physicians were more likely to identify the smoking status of patients with psychiatric diagnoses compared to patients without these diagnoses (76% vs. 64% of visits, p<0.0001). Smokers with psychiatric diagnoses were more likely to be counseled about smoking than were smokers with non-psychiatric diagnoses (23% vs. 18% of visits, p<0.0001), although the absolute difference was small. Primary care physicians counseled smokers with psychiatric diagnoses more often than did psychiatrists, but both groups of physicians counseled at less than half of smokers' visits. All physicians were more likely to counsel smokers with the diagnosis of anxiety but less likely to counsel smokers with the diagnosis of an affective disorder compared to smokers without these diagnoses. Physicians usually identified the smoking status of patients with psychiatric diagnoses but infrequently acted on this information by counseling smokers to quit. Physicians are missing an important opportunity to prevent tobacco-related morbidity and mortality among this group of patients. PMID- 11260816 TI - From reduced smoking to quitting: improvements in COPD symptoms and lung function: a case report. AB - This case report describes a smoker with chronic obstructive pulmonary disease (COPD) who had given up on trying to stop smoking because of many failed attempts. The patient, a 55-year-old woman, was, however, interested in reducing smoking and harm. During a long reduction phase, aided by nicotine replacement, the patient gained self-confidence, despite a depressive episode, and was finally able to quit smoking completely. Spirometry revealed several major improvements after reduction and abstinence. PMID- 11260818 TI - Physiological significance of anthocyanins during autumnal leaf senescence. AB - The light screen hypothesis states that foliar anthocyanins shade the photosynthetic apparatus from excess light. In this paper we extend the light screen hypothesis, postulating that plant species at risk of photoinhibitory conditions during autumnal leaf senescence often utilize anthocyanins to protect the photosynthetic apparatus during the period of nutrient resorption. When senescence-related photosynthetic instabilities are compounded by other environmental stresses, particularly low temperature, severe photoinhibition may result in reduced resorption of critical foliar nutrients, which can significantly affect plant fitness. There is evidence that environments where low and often freezing temperatures are common in autumn selectively favor the production of anthocyanins in senescing foliage. The stimuli for, and the timing and location of, autumnal anthocyanin production are all consistent with a photoprotective role for these pigments in senescing leaves. Furthermore, differences in nitrogen allocation strategies between early and late successional species appear to affect photosynthetic stability during leaf senescence, resulting in a reduced need for foliar autumnal anthocyanins in many early successional plants. The ecological and physiological evidence presented in this paper suggest that, for many deciduous species, the production of anthocyanins provides effective photoprotection during the critical period of foliar nutrient resorption. PMID- 11260820 TI - Regulation of water flux through tropical forest canopy trees: do universal rules apply? AB - Tropical moist forests are notable for their richness in tree species. The presence of such a diverse tree flora presents potential problems for scaling up estimates of water use from individual trees to entire stands and for drawing generalizations about physiological regulation of water use in tropical trees. We measured sapwood area or sap flow, or both, in 27 co-occurring canopy species in a Panamanian forest to determine the extent to which relationships between tree size, sapwood area and sap flow were species-specific, or whether they were constrained by universal functional relationships between tree size, conducting xylem area, and water use. For the 24 species in which active xylem area was estimated over a range of size classes, diameter at breast height (DBH) accounted for 98% of the variation in sapwood area and 67% of the variation in sapwood depth when data for all species were combined. The DBH alone also accounted for > or = 90% of the variation in both maximum and total daily sap flux density in the outermost 2 cm of sapwood for all species taken together. Maximum sap flux density measured near the base of the tree occurred at about 1,400 h in the largest trees and 1,130 h in the smallest trees studied, and DBH accounted for 93% of the variation in the time of day at which maximum sap flow occurred. The shared relationship between tree size and time of maximum sap flow at the base of the tree suggests that a common relationship between diurnal stem water storage capacity and tree size existed. These results are consistent with a recent hypothesis that allometric scaling of plant vascular systems, and therefore water use, is universal. PMID- 11260819 TI - Internal remobilization of carbohydrates, lipids, nitrogen and phosphorus in the Mediterranean evergreen oak Quercus ilex. AB - Remobilization of internal resources is an important mechanism enabling plants to be partly independent of external nutrient availability. We assessed resource remobilization during the growing period in woody and foliar tissues of leafy branches of mature evergreen Mediterranean oak (Quercus ilex L.) at three field sites. We compared nonstructural carbohydrates, lipids, nitrogen and phosphorus pools in leaves and stems before bud burst (March) and at the end of the growing period (July). We also experimentally defoliated leafy branches to determine the storage function of old leaves. Changes in pools of carbon compounds in leaves and stems during spring and in response to defoliation indicated that foliar and woody tissues could provide carbon to support shoot growth. Independently of stem age, soluble sugar and lipid pools decreased significantly during spring. Changes in leaf pools between March and July involved all compounds measured except starch and were accompanied by a 5% decrease in mean leaf biomass. During the same period, 15% of the nitrogen and 25% of the phosphorus were removed from leaves. In contrast, woody tissues did not remobilize nitrogen or phosphorus. Our results support earlier hypotheses that leaves of evergreen species have a primary role in resource remobilization. PMID- 11260822 TI - Effects of timing of nitrogen fertilization on shoot development in peach (Prunus persica) trees. AB - Shoot development was studied for two consecutive years in peach trees fertilized with N either in the previous fall or in the middle of the growing season. During the first year, two additional treatments were studied: no N supply and nitrate supplied in the irrigation water throughout the growing season. The number of shoots that developed depended on nitrogen availability in the period following bud break. During shoot development, leaf emergence occurred in one, two, or three stages, which ended at about 500 to 600 degree days, 1,000 to 1,200 degree days, and 1,500 to 2,000 degree days after bloom, respectively. The proportion of shoots exhibiting a second or third developmental stage depended on nitrogen availability at the beginning of that stage. Increasing nitrogen availability during a developmental stage prolonged the stage and increased the number of leaves produced. PMID- 11260821 TI - Mechanisms of xylem recovery from winter embolism in Fagus sylvatica. AB - Hydraulic conductivity in the terminal branches of mature beech trees (Fagus sylvatica L.) decreased progressively during winter and recovered in the spring. The objective of this study was to determine the mechanisms involved in recovery. Two periods of recovery were identified. The first recovery of hydraulic conductivity occurred early in the spring, before bud break, and was correlated with the occurrence of positive xylem pressure at the base of the tree trunk. Active refilling of the embolized vessels caused the recovery. The second recovery of hydraulic conductivity occurred after bud break and was correlated with the onset of cambial activity. Formation of new functional vessels, leading to an increase in xylem diameter, was largely responsible for the increase in xylem conductivity. The two mechanisms were complementary: active refilling of embolized vessels occurred mostly in the root and the trunk, whereas formation of new functional vessels occurred mainly in young terminal shoots. PMID- 11260823 TI - Morphological and physiological adjustment to N and P fertilization in nutrient limited Metrosideros polymorpha canopy trees in Hawaii. AB - Leaf-level studies of Metrosideros polymorpha Gaud. (Myrtaceae) canopy trees at both ends of a substrate age gradient in the Hawaiian Islands pointed to differential patterns of adjustment to both nutrient limitation and removal of this limitation by long-term (8-14 years) nitrogen (N), phosphorus (P) and N + P fertilizations. The two study sites were located at the same elevation, had similar annual precipitation, and supported forests dominated by M. polymorpha, but differed in the age of the underlying volcanic substrate, and in soil nutrient availability, with relatively low N at the young site (300 years, Thurston, Hawaii) and relatively low P at the oldest site (4,100,000 years, Kokee, Kauai). Within each site, responses to N and P fertilization were similar, regardless of the difference in soil N and P availability between sites. At the young substrate site, nutrient addition led to a larger mean leaf size (about 7.4 versus 4.8 cm2), resulting in a larger canopy leaf surface area. Differences in foliar N and P content, chlorophyll concentrations and carboxylation capacity between the fertilized and control plots were small. At the old substrate site, nutrient addition led to an increase in photosynthetic rate per unit leaf surface area from 4.5 to 7.6 micromol m(-2) s(-1), without a concomitant change in leaf size. At this site, leaves had substantially greater nutrient concentrations, chlorophyll content and carboxylation capacity in the fertilized plots than in the control plots. These contrasting acclimation responses to fertilization at the young and old sites led to significant increases in total carbon gain of M. polymorpha canopy trees at both sites. At the young substrate site, acclimation to fertilization was morphological, resulting in larger leaves, whereas at the old substrate site, physiological acclimation resulted in higher leaf carboxylation capacity and chlorophyll content. PMID- 11260824 TI - Daily time course of whole-shoot gas exchange rates in two drought-exposed Mediterranean shrubs. AB - Effects of drought on water relations, whole-shoot gas-exchange characteristics, and pigment and zeatin concentrations were investigated in the Mediterranean shrubs rosemary (Rosmarinus officinalis L.) and lavender (Lavandula stoechas L.). Two-year-old, greenhouse-grown plants were placed in a whole-shoot gas-exchange measurement system and subjected to 10 days of drought, resulting in severe water stress, and then re-watered for 5 days in order to study their recovery. Water stress resulted in a significant decline in maximum whole-shoot net CO2 assimilation rates (An) for both species that was associated with reductions in leaf area and stomatal conductance. Because shoot dark respiration rate (Rd) was less sensitive to water stress than An, shoot Rd/An ratio increased from about 15 to 95% during water stress. No major changes in chlorophyll and carotenoid concentrations of rosemary leaves were observed during the experiments, but chlorophyll and carotenoid concentrations fell significantly in water-stressed lavender leaves. Zeatin concentrations were higher in rosemary leaves than in lavender leaves during water stress. After re-watering, whole-shoot An and Rd rapidly recovered to their pre-drought rates. PMID- 11260825 TI - Nitrogen forms in bark, wood and foliage of nitrogen-fertilized Pinus sylvestris. AB - Cycling of soluble non-protein N compounds is thought to be indicative of the N nutritional status of trees. We determined the major N forms in bark, wood and foliage and estimated the dependence of prevalent N forms on N availability in Pinus sylvestris L. trees from northern Sweden. Trees subjected to severe N limitation and trees that had been fertilized with an average 64 kg N ha(-1) year(-1) for 25 years were analyzed. Bark and wood samples were collected by tangentially cryo-sectioning the trunk into 30-microm thick sections, from the bark to the functional xylem. Soluble amino compounds were extracted from the sections for analysis. Sap samples from twigs were obtained by centrifugation, and bark samples from twigs were obtained by tissue extraction. In both needles and bark, arginine dominated the amino-N pool. Because arginine concentrations in needles increased with N fertilization, arginine dominance of the amino-N pool in needles was higher in N-fertilized trees than in control trees. In bark, N fertilization resulted in a large increase in glutamine concentration, so that glutamine accounted for a larger proportion of the amino-N pool in bark in N fertilized trees than in control trees. Glutamine dominated the amino-N pool in wood of control trees. Nitrogen fertilization resulted in an increased proportion of arginine in the wood amino-N pool. We conclude that the composition of the amino-N pools in bark, wood and foliage is highly sensitive to N supply. The composition of the amino-N pools can contribute to the regulation of tree N nutritional status, which is mediated by shoot to root signalling by long distance transport of amino compounds. PMID- 11260826 TI - Effects of shading and removal of plant parts on growth of Trema micrantha seedlings. AB - Effects of artificial shading and removal of plant parts on growth of Trema micrantha (L.) Blume (Ulmaceae) seedlings were studied. Seedlings were grown in pots in a greenhouse in 45, 30, 10.6, 4.8 and 1.8% of full sunlight. Shading for 60 days had no effect on survival, but it influenced all growth parameters measured. Total biomass decreased with decreasing irradiance, reflecting reductions in dry mass of leaves, stems and roots. In response to shading, allocation of biomass to leaves increased, while allocation of biomass to roots decreased. Specific leaf area, leaf area ratio and leaf mass ratio increased with decreasing irradiance. Decreases in relative growth rate were caused by reductions in net assimilation rate rather than leaf area ratio. Photosynthetic efficiency, as determined by the Fv/Fm ratio (Fv = variable fluorescence, Fm = maximal fluorescence), was unaffected by the shading treatments. Partial removal of leaves, stem or roots did not affect seedling survival. Seedlings responded to removal of plant parts by compensatory growth. Topophysis was observed when the apex was removed: the lateral buds developed only as new plagiotropic lateral shoots; consequently, the decapitated plant ceased height growth and was unable to compete with its neighbors for light. PMID- 11260827 TI - Editorial. AB - In this, the 11th Annual Research Review, I have been pleased to work with an outstanding group of contributors. As in past issues of the Annual Research Review the aim is to provide our readers with reviews that update both current knowledge and research findings. Authors are asked to be selective, rather than comprehensive, in their coverage as they identify the issues that they feel are particularly important for future research. I am grateful not only to the authors but to the numerous referees who provided critiques of each paper. In the first paper in this issue David Skuse provides an update on the relevance of behavioural neuroscience to child psychopathology. This paper provides a thoughtful review of the findings of the past decade and outlines possible directions for future research developments; it appears that we are poised for a major explosion of knowledge in this area. In the second paper Robin Chapman provides a very useful review of recent research on language development. This paper provides an update of Dorothy Bishop's earlier review of the topic and illustrates the considerable progress made since the time of that review. In the third paper Eilish Gilvarry summarises recent research on substance abuse in young people. This review covers recent changes in trends and patterns of substance abuse, aspects of risk and comorbidity, and treatment. Brown and colleagues then review recent work on children and adolescents with HIV and AIDS; this global health problem presents unique issues relative both to research and intervention. Danya Glaser then provides an overview of recent work on child abuse and neglect and the brain; the attempt to bring the various perspectives of neuroscience together on this topic is particularly timely and appropriate. Finally, Sparrow and Davis provide an overview of recent advances in the assessment of intelligence. This paper provides a helpful summary of current perspectives on the assessment of intelligence; the review of instruments will be of particular interest to our readers. For the 12th edition of the Annual Research Review we anticipate coverage of the following topics: intersubjectivity, reading disability, longitudinal approaches to developmental data, mental retardation, conduct disorder, and psychopharmacology. PMID- 11260828 TI - Editorial. AB - There has been a consensus that children "in care" show a much increased frequency of behavioural difficulties but the reasons why this should be so are much less well understood. The study by Roy et al. sheds important new light on this issue. They found that children admitted into residential group homes as babies are much more likely than children admitted into foster care at the same age to show hyperactivity and inattention. Although the study sample was small, the groups were closely comparable in coming from a very high-risk family background. The evidence from both questionnaire and observational measures was consistent in indicating that the difference in the pattern of rearing made a substantial difference to the child's behaviour. The findings are sobering in their implication that the pattern of care provided to protect children at high risk seemed to have acted in a detrimental manner. The study clearly provides food for thought in terms of the need to improve provision for this vulnerable group of children. The findings are also provocative in their implication that hyperactivity/inattention, although strongly influenced by the child's biology, can also be affected by the pattern of rearing. The message is that clinicians should not assume that the causes of this hyperactive behaviour necessarily reside entirely within the child but there is also the need to clarify whether the form of hyperactivity/inattention arising from these experiences is in someway atypical. The paper by Reynolds and colleagues is an attempt to evaluate the efficacy of emotional disclosure to offset distress in children experiencing negative life events. There is a developing literature on such interventions in adults but, as these authors suggest, little work has yet been done to test the value of such disclosure in children. Using a randomised controlled trial Reynolds et al. were unable to show a specific benefit from the opportunity provided to write about negative events. Rather, there was a general reduction in symptoms from all groups. Although the results suggest that the efficacy of writing about negative events is less marked in 8-13-year-olds than in adults, they also indicate that it is both feasible and potentially valuable to give children opportunities to engage in discussion about sources of stress and their reaction to them. PMID- 11260829 TI - Editorial. AB - Many children and adolescents do not have the peaceful and carefree youth they are entitled to, because they have been exposed to adverse environmental influences that disturb a normal development. These influences may be caused directly by adults whose task it is to protect these children, but sometimes traumatic experiences, including those caused by natural disasters or war conditions, are beyond the control of parents or other adults. The first three articles of this issue pertain to traumatic events that have a great impact on the child's development. The Practitioner Review in this issue by Perrin, Smith, and Yule provides the most up-to-date summary of the literature on the assessment of Post-traumatic Stress Disorder (PTSD) in young people. Considerable controversy has surrounded PTSD since its inception into the diagnostic nomenclature. The authors make the case that whatever the limits of the diagnostic criteria, children do indeed suffer from post-traumatic stress reactions. The authors draw on their extensive experience working with children exposed to war, natural and man-made disasters, and domestic violence, to give the reader an expert-eye view of the assessment and treatment of childhood PTSD. They make a strong case based on the available literature that cognitive and behavioural approaches hold the most promise as a treatment for childhood PTSD. The reader will find this article useful from both a conceptual and a clinical practical point of view. PMID- 11260830 TI - Editorial. AB - What makes young people act in antisocial ways? This question is discussed from several different angles in our current issue. Delinquency is not, of course, a new phenomenon: for those interested in reading a chilling account of life in London in the early years of the last century, Cyril Burt's The young delinquent, published in 1925, is worth tracking down. Definitive answers seem as far away as ever, but the idea that delinquency might be influenced to a significant extent by genetic mechanisms is still a controversial one. Taylor et al. conducted a twin study, and found the variance in delinquent behaviour among adolescent boys and girls was associated largely with experiences that were unique to individuals. Family influences accounted for half of the remaining variance in risk less than 20% could be attributed to additive genetic factors. Their findings imply, they suggest, scope for prevention and/or intervention. On a similar topic, Hawker and Boulton ask, why is it that some children are bullied? What effect does bullying have on children's emotional adjustment? They review the history of research on this topic, going back over two decades to the pioneering studies of Dan Olweus. Using meta-analytic techniques, they conclude victims become emotionally distressed and, in particular, depressed. Clinicians should realise that children who present with emotional problems may be the victims of bullying; interventions that target either bullying or emotional distress may reduce the severity of both problems. PMID- 11260831 TI - Editorial. AB - It is nearly 30 years since the publication of the edited book by Rutter and Martin (1972), The child with delayed speech (London: Heinemann), which drew attention to the two-way relationship between language and behaviour. In the book the mechanisms whereby psychiatric disorders can have an involvement with language development were identified. Also, Mike Rutter suggested various ways in which delayed language development could impinge upon other aspects of psychological development. The current issue of the JCPP has a number of papers that examine the relationship between language development and aspects of cognition and behaviour. These include a long-term follow-up for a group of individuals with receptive language disorder who were first studied at about the time that the 1972 book was published. PMID- 11260832 TI - Editorial. AB - The readers of the Journal may not be aware that the Joint Editors act independently in making decisions on accepting papers for publication. This means that as an Editor I am just as intrigued as any other reader when I see a new issue of the Journal since it is likely that I will have been responsible for selecting only one third of the papers in that issue. In reading the material in this present issue I was struck by the conceptual and methodological links between a sub-set of papers that were concerned with adverse events and circumstances and their long-term sequelae; moreover, that these papers had between them some important implications for clinical practice. The first of these papers is by Dunn et al. and investigates the transmission within families of qualities of relationships. They found that father-child and mother-child relationships in stepfamilies, single-parent, and non-stepfamilies were found to be related to a number of factors. These included the parents' own earlier life course experiences, current family circumstances, and how a partner and child were getting along. The links with life course experiences meant that children were at risk of a "double dose" of less affectionate relationships in families in which parents had experienced early adversities. They found evidence for both selection effects (similarities in the early experience of both partners) and co parenting effects (effects of one parent's relationship with a child on the other parent) and effects of biological relatedness. PMID- 11260833 TI - Critical Notice. AB - "Goodness of Fit": Clinical Applications from Infancy through Adult Life. By Stella Chess & Alexander Thomas. Brunner/Mazel, Philadelphia, PA, 1999. pp. 229. pound24.95 (hb). Chess and Thomas's pioneering longitudinal studies of temperamental individuality started over 40 years ago (Thomas et al., 1963). Their publications soon became and remain classics. Their concept of "goodness of fit" emerges out of this monumental work but has had a long gestation period. In their new book, the authors distinguish between behaviour disorders that are reactive to the child's life circumstances, including life events, and which are self-correcting or responsive to the relevant changes in their environment, and more serious disorders. PMID- 11260834 TI - [The best in 2000 on heart failure]. AB - The advances in cardiac failure are permanent. In 2000, once again, they concerned all fields of pathology. Complementary information of the epidemiology of the disease in France and Europe has been published. The prevention of sudden death by the implantable defibrillator in hypertrophic cardiomyopathy has been confirmed. The prognostic role of the aetiology of dilated cardiomyopathy has been demonstrated and the distinct clinical entity of acute fulminating viral myocarditis with an excellent long-term prognosis has been identified. New genetic abnormalities have been found in different forms of dilated cardiomyopathy. One of the most important advances concerns the neurohormones and the rise of interest in type B natriuretic peptide, for diagnosis, prognosis, as well as treatment (nesiritide). From the therapeutic point of view, the importance of betablockers has been confirmed, including in severe cardiac failure. The therapeutic value of biventricular stimulation resynchronising the two ventricles seem to be an additional therapeutic opportunity for patients with advanced cardiac failure. The summit of 2000 remains the first cellular transplantation carried out by a Parisian French team. PMID- 11260836 TI - [The best in 2000 on echocardiography]. AB - The advances in Doppler echocardiography, like last year, concern the technology, the indications and the description of the natural history of cardiovascular diseases. The year 2000 will be remembered for the development of portable echocardiographs with two-dimensional and Doppler capabilities but without M mode, pulsed or continuous Doppler. These instruments weigh less than 3 kg and may be used at the bedside; however, problems of training personnel, reimbursement and availability of equipment remain. Myocardial contrast echocardiography has been the subject of many publications which validate continuous intravenous infusion of different contrast agents, which, coupled with techniques of image processing, suggest that quantification of regional myocardial perfusion at rest or during physiological or pharmacological stress may not be far off. The indications of stress echocardiography have increased: in addition to diagnostic and prognostic information in coronary artery disease, the applications of exercise and pharmacological inotropic stress echo extend to asymptomatic valvular regurgitation to detect infraclinical myocardial dysfunction before the usual Doppler echocardiographic parameters show significant changes. The value of Doppler tissue imaging in the assessment of regional myocardial function has been demonstrated: analysis of diastolic function with pulsed Doppler at the mitral annulus, quantification of regional myocardial function in different pathologies such as ischaemic heart disease and cardiomyopathy, validation of indices during stress echocardiography with the hope of a quantitative as well as a qualitative assessment, much improved since the introduction of harmonic imaging. Finally, Doppler haemodynamic evaluation has continued to substitute for cardiac catheterisation, now providing accurate indices of quantification of valvular regurgitation and description of the natural history of cardiac diseases, especially mitral regurgitation irrespective of its cause and aortic stenosis including cases with low output states. The continuing progress of ultrasound technology is therefore confirmed and provides an outlook into the third millennium: development of portable equipment, improved probe technology, use of instruments of quantification and extension of real time three-dimensional reconstruction, systems of image stocking and transmission and better orientation of its usage with the recommendations of scientific societies. PMID- 11260835 TI - [The best in 2000 on arrhythmia]. AB - The two major problems of rhythmology in the year 2000 were mainly therapeutic: on the one hand, the treatment of atrial fibrillation and, on the other hand, the treatment of sudden arrhythmic death. These two subjects should not mask the fermentation of ideas, techniques, and diagnostic and therapeutic suggestions which have made up one year of scientific literature in this field. However, simple analysis of the articles published in the leading clinical review, the New England Journal of Medicine, shows that these two subjects are the main items. In the treatment of atrial fibrillation, Canadian investigators in the CTAF study, showed that amiodarone was more effective than Sotalol or Propafenone for the prevention of recurrences of atrial fibrillation in a population of 403 patients followed up for an average of 16 months. There were 35% of recurrences in the amiodarone group compared with 63% in the Sotalol and Propafenone group. Two articles demonstrated the value of automatic external defibrillation in the prevention of sudden arrhythmic death, especially when used in a relatively confined space such as an aeroplane or a casino. These two American Publications showed the value of this approach for a rapid recovery after ventricular fibrillation, the rapidity being the only guarantee of a high survival rate after hospital discharge, about 40% in this series. These various studies should be placed in the perspective of the general changes in treatment of these two pathologies: the treatment of atrial fibrillation is becoming progressively more hybrid, the same patient being considered at different moments for pharmacological antiarrhythmic therapy, ablation or pacing: arrhythmic sudden death is of increasing concern, either with curative treatment by external automatic defibrillation or by preventive therapy with the implantable defibrillator, the trend being towards "prophylactic" indications in patients in whom the high risk of sudden death has been determined. PMID- 11260837 TI - [The best in 2000 on heart stimulation]. AB - In the last twelve months many important articles have been published in the field of cardiac pacing. The authors analyse 5 of them in this review: the Canadian CTOPP, which compared dual and single chamber pacing in patients with a classical indication for pacing, and concluded that the results were identical in the two modes but not without serious criticism about the validity of this conclusion. A second Canadian trial analysed the effects of atrial stimulation in the prevention of atrial fibrillation but without apparent benefit. In 1999, vaso vagal syncope had already been studied in a randomised trial; a second one was published in 2000 with concordant results in favour of pacing but reserved for a very selected population. Finally, two articles were devoted to "left heart" pacing in cardiac failure. The MUSTIC trial was the first randomised protocol and its results were favourable for this type of pacing. A physiopathological study reported by a Baltimore group, provided fundamental information: the increase of left ventricular contractility (DP/Dt) occurs without any increase in myocardial oxygen consumption. PMID- 11260839 TI - [The best in 2000 on pediatric cardiology]. AB - The year 2000 was rich in events, either spectacular news or confirmed improvement of on-going advances, as far as paediatric cardiology is concerned. The selection presented by the authors includes the first percutaneous implantation in a human being of a biological (bovine) valve which was sewn on a stent, compressed into a catheter and inserted against a stenotic and leaking procine bioprosthesis in a right-ventricle to pulmonary-artery conduit. This may be a new way to further valve replacements as alternatives to surgery. Balloon dilation of late postoperative recoarctations is now also improved with the use of stents able to maintain the result and to avoid traumatic injuries, with new coaxial double balloons making the procedure easier and safer. This is probably one of the main elements in reducing this very particular form of hypertension, the anatomic cause of which is often difficult to understand. As for yesterday's daring innovations now becoming near-routine protocols, two examples are developed. First, the rehabilitation of pulmonary arteries in pulmonary atresia with ventricular septal defect and complex pulmonary blood supply, both by true pulmonary vessels and by collaterals, both being stenotic and/or hypoplastic, anastomosed or not. The anatomic and functional details of such a vascular setting should be accurately understood and treated by early and aggressive surgery and interventional procedures in order to promote antegrade flow, distal angiogenesis, and, finally, active and harmonious vascular growth compatible with complete repair. The second example is Friedreich's ataxia in which, within 3 years of the discovery of the pathogenic mechanism, the deficiency in frataxin and its intra-cellular toxic consequences have been demonstrated, leading to a logical medical therapy which proves to be effective in treating (and maybe in preventing) the severe hypertrophic cardiomyopathy associated to this disease. PMID- 11260838 TI - [The best in 2000 on clinical pharmacology]. AB - The year 2000 provided many new articles in clinical pharmacology and therapeutics in the different fields of cardiology. The authors present some of them in this review. In the field of prevention, the statins were the object of complementary studies showing their value in high cardiovascular risk patients with benefits not only in the reduction of coronary but also cerebrovascular events. These benefits are maintained at long-term. The debate about the value of Vitamin E is still on-going with divergent results (HOPE, SPACE studies...). The absence of secondary coronary prevention by post-menopausal hormone replacement therapy seems to be confirmed. The arrhythmogenic risk of neuroleptic drugs is of increasing concern. New data also suggests the possibility of a venous thromboembolic risk. The NSAIDs are an important factor in the first episode of cardiac failure. The risk of thromboembolism with the 3rd generation of contraceptives has been confirmed. Some data has been published about the safety of drugs used in cardiology: the haemorrhagic risk of LMW heparin in renal failure and of aspirin, even at low doses, drug interactions, aspirin-ACE inhibitors interaction. Future perspectives include the potential value of vasopeptidase inhibitors, of cerebral natriuretic peptide and of therapeutic approaches to induce angiogenesis in ischaemic heart disease (gene therapy, recombining factors). PMID- 11260840 TI - [The best in 2000 on coronary angioplasty or the postintervention period]. AB - The year 2000 I The left main coronary and myocardial infarction have fallen. The last obstacles confronting the interventional cardiologist have been overcome with the help of stenting. The interventional cardiologist is now working in a limited, well defined functional space and anatomic territory, of which stenting is the main element. It is now known that the successful treatment of arterial stenosis is not necessarily victory over the lesion. The time of conquest and pioneers is over. We are now confronted with the problem of justification of the act by validation of the method and strategies. This is an era of training, organisation and management of the practice with the elaboration and respect of scientific recommendations. Finally, there are new challenges: that of restenosis, and the progression of atherothrombosis, which have led interventional cardiologists to develop: on the one hand, diagnostic tools assessing the arterial lesion just beyond the opacification of the arterial lumen and, on the other hand, therapeutic techniques superseding a mechanical effect on the stenosis. Thus, we are beginning to see the first preventive and curative biological treatments (brachytherapy, gene therapy, anti-atheromatous chemotherapy), founded on a multidisciplinary approach to atheromatous disease. PMID- 11260841 TI - [The best in 2000 on thrombosis]. AB - It is nearly impossible to follow and integrate all the new information in each subspeciality of cardiology. In the last months, important data has been published which may change clinical practice. In this domain, over half the cases of suspected coronary chest pain would only require a very short stay in a chest pain unit. The history, an accurate evaluation of symptoms, the application of Bayesian analysis, ECG interpretation and serum troponine measurement, associated with a degree of clinical experience, will allow orientation of the patient to a coronary care unit or hospital discharge (with possible out-patient referral). Patients with true unstable angina will no longer be treated by continuous intravenous injection of non-fractionated heparin because, in theory and in practice, this has been replaced with subcutaneous LMW heparin.... On the other hand, the electric syringe will continue to be required for the integration of the anti-GPIIB-IIIA for the treatment of unstable angina after the recommendations published concomitantly in the United States and Europe. This type of patient, especially with a "positive" troponine, will probably not be kept waiting long before referral to the catheter laboratory for coronary angiography and revascularisation. The long-term results of the FRISC II trial are confirmed by an even earlier invasive approach (Tactics-Timi 18) using anti GPIIb-IIIa. In the first hours, and independently of other older prognostic factors, it will be possible to "predict or compare" the risk of coronary recurrence based on the results of certain biological "markers". In many centres, cases with ST elevation on the ECG could well be included in a phase III "medical protocol", associating a half-dose thrombolytic and an anti-GPIIb-IIIa. Finally, patients who will have been admitted to the chest pain unit with a suspected pulmonary embolism, for example because they had not been treated prophylactically with aspirin before hip surgery, will probably have the choice, after d-dimer measurement, between pulmonary scintigraphy and helicoidal CT scan. If the diagnosis of pulmonary embolism is confirmed, a single subcutaneous injection of LMW heparin could replace the conventional continuous intravenous injection of heparin. The earliest possible oral anticancer ... pardon me I anticoagulant treatment should be prescribed and explained. PMID- 11260842 TI - [The best in 2000 on arterial hypertension]. AB - The authors showed, in a spin-off study of SYST-EUR, that 24% of subjects with isolated systolic hypertension on conventional measurement were not hypertensive during ambulatory blood pressure monitoring. Moreover, in white coat hypertension, treatment had no effect either on the electrical signs of left ventricular hypertrophy or on the incidence of clinical events (cerebrovascular accident and global cardiovascular complications), contrary to what is observed in permanent systolic hypertension. These results raise question as to the diagnosis and treatment of isolated systolic hypertension in the elderly and prompt to a larger usage (if not systematic) of ambulatory blood pressure monitoring in this context. The importance of systolic blood pressure and pulsed pressure For different reasons, diastolic blood pressure was thought to be of greater prognostic significance, as the very large majority of clinical trials recruited on the basis of the value of their diastolic blood pressure alone demonstrate. In recent years, the importance of systolic blood pressure has been underlined in many studies and 3 trials have shown the unquestionable benefits of treatment of isolated hypertension. It would also appear that the pulse pressure, which reflects arterial compliance, has considerable prognostic value. In the absence of established manometric criteria and mostly of therapeutic trials, the practical use of the pulse pressure remains questionable. The interruption of the doxazosin arm of the ALLHAT trial The ALLHAT (Antihypertensive and Lipid Lowering Treatment to Prevent Heart Attack Trial) study showed a doubling of the morbidity from cardiac failure, a 19% excess of cerebrovascular events and 16% of angina pectoris in subjects treated with doxazosin compared with those treated with chlorthalidone. The differences in blood pressure with treatment were minimal and, a priori, unable to explain these results. Beyond the fact that alphablockers cannot be considered as first-line antihypertensive therapy, without doubt, the affirmation that lowering the blood pressure provides the same benefit irrespective of the antihypertensive agent used, probably needs to be reviewed. PMID- 11260843 TI - [The best in 2000 on angiology]. AB - There were no great innovations in angiology in the year 2000 but there was a number of advances, one of which being the publication of an international consensus on peripheral occlusive arterial disease, the TASC consensus. Each year, the number of risk factors for venous thromboembolic disease increases, and 2000 was no exception with the demonstration that high factor IX and XI concentrations were associated with this condition. On the arterial side, oral anticoagulation and aspirin were shown to be equivalent in the prevention of occlusion of lower limb bypass grafts. Finally, some recent data is in favour of a systemic mechanism in the acute complications of atherosclerosis. PMID- 11260844 TI - [The best in 2000 on heart imaging: "the transparent heart"]. AB - Cardiac functional imaging focuses on three variables which complement each other: coronary perfusion reserve (during stress or after maximal vasodilation), myocardial viability (at cellular level) and ventricular contraction (endocardial motion or myocardial thickening). A combination of abnormal findings (location, size and severity) helps to characterise the myocardium in terms of normal, ischaemic, stunned, hibernating or necrosis. This functional information is then used to validate the morphological images of coronary lesions as seen on angiography, to assess the significance of a stenosis, to identify the culprit vessel in cases of multivessel disease, and to delineate the area at risk. "Non invasive" techniques should no longer be considered only as screening methods with more sophisticated investigations in view, but should replace them at every step of patient management: at the time of diagnosis, with a "positive/negative" bayesian approach, before a therapeutic decision and during follow-up, especially when considering the prognosis of patients and risk stratification when the quantitation of functional abnormalities are major determinants. PMID- 11260845 TI - [The best in 2000 on valve diseases]. AB - During the year 2000, publications on valvular heart disease have concerned all aspects of this field of cardiology at a time when old and dated therapeutic procedures are being reassessed. The ageing population of the developed world has led to aortic stenosis playing a large part, and the study of its natural history has provided two keynote publications. Aortic valve replacement, increasingly involving older patients, led to the evaluation of this surgery in this age group in which bioprostheses are often associated with coronary bypass surgery. Conversely, in younger patients, there is a regain in interest in autograft (Ross' procedure) or homograft valve replacement which requires a rigorous infrastructure of supply. In mitral valve disease, the indications of conservative surgery of mitral incompetence, ideal in degenerative forms of the posterior leaflet, have been progressively extended to include bacterial endocarditis in many cases and ischaemic mitral regurgitation according to some authors. Rheumatic lesions are not commonly treated by this technique although some encouraging results have been reported. Percutaneous mitral commissurotomy has attained maturity in the treatment of mitral stenosis, even in the less favourable forms such as restenosis after an initial percutaneous procedure or even after surgical commissurotomy. Valve replacement surgery by prosthetic valves is forty year old and many long-term retrospective and prospective evaluations of the results on large patient population either with one type of prosthesis or comparing different bioprostheses or bioprostheses with mechanical valves have been performed. The ideal age for implanting bioprostheses remains uncertain, between 60 and 70, depending on the authors. Finally, problems of anticoagulation in patients with prosthetic valves were the object of three interesting publications about the use of low molecular weight heparin, aspirin and the risks during pregnancy. 2000 was a year of steady and regular progress in the study of valvular heart disease without any major revolutionary contributions. PMID- 11260846 TI - Progress towards poliomyelitis eradication, Afghanistan, 1999-2000. PMID- 11260847 TI - New study indicates dearth of useful resources for cardiology benchmarking. AB - Resources abound for cardiologists who want clinical benchmarking data, but according to a new study, hospital administrators have to search for accurate, real-time, national and regional benchmarks that integrate cost and clinical quality. PMID- 11260848 TI - ANA survey: nurses say patient care is suffering. AB - The latest American Nurses Association (ANA) staffing survey of its membership shows increasing concern among nurses that patient care is suffering due to declining working conditions. According to ANA president Mary Foley, MS, RN, three-quarters of the 7,300 respondents to the survey feel the quality of nursing care at the facility in which they work has declined. PMID- 11260849 TI - How to take the pain out of hospitalization. AB - The patients at Good Samaritan Hospital in Baltimore had a problem: No one was asking about their pain. When they filled out their patient satisfaction surveys, they let administration know that this was an area where they were less than satisfied with their care. So two years ago, the hospital hired a pain management coordinator to develop a program from scratch. PMID- 11260850 TI - Don't keep women waiting on a diagnosis. AB - In December 1999, the administration at Grant/Riverside Methodist Hospitals in Columbus, OH, launched a benchmark study to determine how long it took for a woman diagnosed with a breast abnormality to find out if she had breast cancer. It was determined that it took an average of 27 days, better than the national average of 42 days, but far below the gold-standard programs around the country that took 24 hours to five working days. To remedy the situation, Patti Dunn, RN, BSN, OCN, was asked to oversee the process and create a system that coordinated care across the continuum to better meet the needs of women with breast health problems. PMID- 11260851 TI - Low-volume hospitals combat low-quality image. AB - Patients with certain conditions have better odds for survival if they go to hospitals that treat a high volume of those conditions. Some researchers question the validity of the volume-outcome studies in general. Even as the debate proceeds, however, health care purchasers are paying closer attention to such studies. One group of high-profile purchasers is even discussing the controversial practice of selective referrals to high-volume facilities. PMID- 11260852 TI - Five ways to deal with information security incidents under HIPAA. PMID- 11260853 TI - Phlebotomist attacked by unruly patient wins $210,000 from hospital. Powell v. Catholic Med. Ctr., No. 97-860 (N.H. Mar. 21, 2000). PMID- 11260855 TI - New technology enhances surveillance, access control in parking lots. PMID- 11260854 TI - When fired worker sues: why hospital did the right thing. Tate v. Baptist Mem'l Hosp., No. W1999-00553-COA-R3-CV (Tenn. Ct. App. 2000). PMID- 11260856 TI - Hospital, community groups sponsor workplace violence conference. PMID- 11260857 TI - Employee theft and fraud: bigger than ever and getting worse. AB - Fraud examiners and investigators say that employee theft is out of control, citing recent U.S. Chamber of Commerce figures that employees steal approximately $400 billion from businesses each year and an Ernst & Young survey showing that nearly 90% of organizations countrywide experienced some type of fraud in the 12 months prior to the survey. An additional undetermined amount is being reported lost because of external fraud committed by customers, vendors, and others having contact with a company or institution. Fraud experts interviewed for this report note that all sectors are targets--including retailers, hospitals and healthcare, the hotel industry, schools, and college campuses. In this report, we'll present the how's and whys of this development and also describe what one medical center is doing to stop thefts of medical equipment and computers. PMID- 11260858 TI - Protecting your hospital from gang activity. PMID- 11260859 TI - Clinical management of malignant adrenal tumors. AB - Malignant primary adrenal tumors are rare forms of cancer with an estimated incidence of two to ten new cases per one million inhabitants per year. The 5 year survival rate for adrenocortical carcinoma is approximately 35%, whereas the 10-year survival rate of malignant pheochromocytoma reaches 40%. Clinical studies support repeated surgery as the mainstay of treatment, either with curative or palliative intention. For adrenocortical carcinoma, adjunctive treatment with oral mitotane leads to well-documented improvement of survival. Rare malignant pheochromocytomas with distant metastases are preferably treated by 131I-MIBG. Chemotherapy is reserved for unresectable tumors without sufficient response to mitotane or 131I-MIBG, respectively. Cisplatin and etoposide as single therapy, or in combination with doxorubicin or etoposide, appear to be effective in adrenocortical carcinoma. Malignant pheochromocytoma may be treated with vincristine, dacarbazine, and cyclophosphamide. Treatment with octreotide is currently being evaluated. Radiotherapy is indicated if unresectable tumor masses cause local symptoms. If symptoms of endocrine activity are not sufficiently controlled by measures aiming at tumor mass reduction, specific inhibitors of hormone synthesis or action are available. Ketoconazole is widely used for adrenocortical carcinoma, and phenoxybenzamine and metyrosine are available for malignant pheochromocytoma. This review provides guidelines for rational disease management based on still scanty clinical evidence. PMID- 11260860 TI - Diagnosis and therapy of sporadic and familial medullary thyroid carcinoma. AB - Medullary thyroid carcinoma (MTC) is a rare thyroid malignancy. About 75% are sporadic (sMTC) while the remaining 25% are hereditary (hMTC). The treatment of choice for both sMTC and hMTC is surgery. An adequate initial operation provides the best chance of cure. Hence, the diagnosis of MTC should be made preoperatively. In sMTC, ultrasound, ultrasound-guided fine-needle aspiration cytology and measurement of calcitonin levels (basal and after injection of calcitonin-stimulating reagents, e.g., pentagastrin) are sensitive diagnostic tools. In hMTC, identification of a germline mutation in the proto-oncogene RET is sufficient for making the diagnosis. Total thyroidectomy is recommended in all patients, sporadic and hereditary. In addition, lymphadenectomy of the cervicocentral and both cervicolateral compartments should be performed. The only indication to perform a less extensive operation may be given in young patients with hMTC. Sufficient treatment of MTC beyond local disease is still non existent. Future research should concentrate on this issue. PMID- 11260861 TI - Transcriptional repression of the human fibronectin gene in laryngeal squamous cell carcinoma cells. AB - PURPOSE: The aim of the experiments was to analyze the mRNA expression pattern and verify the repression of FN gene expression in laryngeal squamous cell carcinoma (SCC) cells in comparison with benign mucosal keratinocytes. METHODS: Messenger RNA from SCC cells and benign keratinocytes was reverse transcribed and subjected to PCR following differential display (DD) analysis of the amplicons. Northern hybridization was carried out to confirm the reduction of the FN-mRNA expression in both laryngeal SCC cells and larynx carcinoma biopsies, in contrast to adjacent normal mucosa. Quantitation of protein synthesis was performed with homogenates of fresh tumor biopsies and their normal phenotypes, as well as of benign keratinocytes and laryngeal SCC cell lines, respectively, using ELISA. In the liposome-mediated transient transfection assay, FN promoter activity was analyzed by linking the FN promoter sequence to the chloramphenicol acetyltransferase (CAT) reporter gene. Transfection efficacy was monitored by co transfection with pGL3 control vector. RESULTS: A 191 bp mRNA fragment revealing a 99% homology with the human FN-mRNA was detected, the expression of which was repressed 20 times as much in SCC cells as compared to benign phenotypes. Northern hybridization confirmed the distinctly reduced expression of FN-mRNA in both laryngeal SCC cells and larynx carcinoma biopsies, in contrast to adjacent normal mucosa. The quantitation experiments showed a correlation between the range of FN synthesis and the expression of FN-mRNA in cell lines and the biopsies which were used. The 1.28 kb FN gene promoter drove expression of the CAT reporter gene, which was similar to the FN-mRNA expression showed by DD and Northern hybridization. CONCLUSIONS: The mechanisms leading to the low level of FN in many tumors have not yet been sufficiently investigated. Our findings suggest that the decrease of FN in laryngeal SCC cells is transcriptionally regulated. PMID- 11260862 TI - Effect of the nonsteroidal anti-inflammatory drug indomethacin on proliferation and apoptosis of colon carcinoma cells. AB - PURPOSE: Nonsteroidal anti-inflammatory drugs lower the incidence of and mortality from colon cancer. In this paper, we present the effect of indomethacin on growth inhibition and alterations in the expression of several genes involved in cell cycle and apoptosis in CaCo-2 colon adenocarcinoma cells. METHODS: We used the MTT test to evaluate the effect of indomethacin on the proliferation rate of colon cancer and normal fibroblast cells in vitro. The expression of c myc oncoprotein and p53 and p27 suppressor proteins was examined using the immunocytochemical method. RESULTS: We have shown that indomethacin reduces the proliferation rate of CaCo-2 colon cancer cells (up to 60% at the concentration of 4 x 10(-4) M), alters their morphology, and induces cell death by apoptosis. The most pronounced inhibitory effect was observed at the concentration of 6 x 10(-4) M where the growth was completely suppressed. However, the growth of normal fibroblasts (Hef 522) was much less inhibited (about 30% of inhibition at the concentration of 6 x 10(-4) M). Indomethacin reduces the proliferation rate and induces apoptosis in CaCo-2 colon cancer cells through enhanced expression of c-myc, p53, and p27 proteins. CONCLUSIONS: This is the first report about p27 increased expression in colon carcinoma cells induced by indomethacin treatment. Increased expression of p27 represents a new mechanism of apoptosis in cells treated with NSAIDs (indomethacin). This effect probably contributes to the anti proliferative effect on colon cancer cells in vitro. PMID- 11260863 TI - Analysis of expressions of components in the plasminogen activator system in high and low-metastatic human lung cancer cells. AB - PURPOSE: To determine the expressive patterns of the components of the plasminogen activator system in human large-cell lung carcinoma strains and to analyze the effects of the patterns on tumor invasion and metastasis. METHODS: The in vitro and in vivo invasive and metastatic potential of two human large cell lung carcinoma strains with high (strain 95D) and low (strain 95C) metastatic potential was further confirmed by the Boyden chamber model and nude mice model. After this, the expressions of the components of the plasminogen activator system--including urokinase-type and tissue-type plasminogen activator (uPA and tPA), urokinase receptor (uPAR), and type-1 and type-2 plasminogen activator inhibitor (PAI-1 and PAI-2) in strain 95D and 95C cells--were determined by RT-PCR and immunohistochemical staining. The effects of monoclonal antibodies of uPA, uPAR, and PAI-1 on the invasive potential of strain 95D cell line were also evaluated. RESULTS: Strain 95D cells were found to have a stronger in vitro and in vivo invasive and metastatic potential than strain 95C cells. In the former, the average number of infiltrating cells in the in vitro model in one field of vision (40055) was 73.75 +/- 7.42, while in the latter, it was 56.33 +/- 6.28 (P < 0.001). Lung metastatic loci were observed in all six nude mice inoculated with 95D cells (6/6), but not in any of the nude mice inoculated with 95C cells (0/6). The high-metastatic strain 95D cells expressed higher uPA and uPAR and lower tPA and PAI-2 than the low-metastatic strain 95C cells. The PAI-1 expressions in both 95D and 95C cells were almost the same. Monoclonal antibodies of uPA and uPAR greatly reduced the invasive potential of strain 95D cells in vitro. CONCLUSIONS: These data suggest that the invasive and metastatic potential of human large-cell lung carcinoma cell lines is associated with differential expressions of the components of the plasminogen activator system and that the determination of these components may be used as a marker for judging clinically the possibility of tumor metastasis as well as the prognoses of patients. PMID- 11260864 TI - Multiple genetic alterations involved in the tumorigenesis of human cholangiocarcinoma: a molecular genetic and clinicopathological study. AB - PURPOSE: Cholangiocarcinoma (CC) is the second most common malignant tumor in the liver and the molecular genetic alterations involved in the tumorigenesis of CC have not been well studied. PATIENTS AND METHODS: The authors analyzed the loss of heterozygosity (LOH) in four tumor suppressor genes, including the adenomatous polyposis coli (APC) gene, the deleted in colon cancer (DCC) gene, the 8 hydroguanine-specific DNA glycosylase (OGG1) gene, and the p53 gene in 22 surgically resected primary CCs by using microdissection-based PCR amplification and direct DNA sequencing. RESULTS: A total of 19 (86.4%) out of 22 CCs exhibited genetic alterations, of which 11 (57.9%) and eight (42.1%) cases showed one and more than one gene alterations, respectively. The frequency of genetic alterations of the four genes studied ranged in order from high to low as APC (68.8%) > DCC (46.2%) > OGG1 (41.7%) > p53 (37.5%). Based on the pattern of altered genes and their correlation with clinical and pathological parameters, the genetic alterations were classified into three groups: group I: no detectable genetic alterations (n = 3, 13.6%); group II: LOH in APC and/or DCC (n = 9, 40.9%); and group III: LOH in OGG1 and/or p53 occurred separately or combined with LOH in APC and/or DCC (n = 10, 45.5%). The > or = 3-year survival rates between group II and group III are 88.9% and 30%, respectively (P < 0.05). No significant differences were found between genetic alterations and tumor size, tumor type, tumor invasion, TNM staging, and tumor differentiation (P > 0.05). CONCLUSION: Accumulation of multiple genetic alterations are involved in the tumorigenesis of CC, of which genetic alterations of APC and DCC occur at a relatively early stage, and of OGG1 and p53 occur at a relatively late stage during development of CC. PMID- 11260865 TI - Clinicopathological significance of fibrotic capsule formation around liver metastasis from colorectal cancer. AB - PURPOSE: The fibrous capsule around hepatocellular carcinoma is well known to be an indicator of a good prognosis. However, the fibrotic stromal response in the liver to a metastatic tumor remains unclear. PATIENTS AND METHODS: In order to clarify the prevalence of fibrotic capsular formation around liver metastases as well as the prognostic and biological significance of the fibrotic capsule, 69 colorectal cancer patients, who underwent radical hepatectomy due to liver metastases, were investigated using immunohistochemical methods. RESULTS: Encapsulated metastases as defined by a thick fibrotic band surrounding the entire surface of a metastasis were detected in 20% of the cases. The rate of initial recurrence in the remnant liver, which is a strong indicator for poor prognosis of colorectal liver metastasis, was significantly lower in the encapsulated metastasis group as compared with the non-encapsulated metastasis group. Proliferating fibroblastic cells in the capsule were myofibroblasts positively stained for alpha-smooth muscle actin (alpha-SMA) and they deposited dense extracellular matrices rich in collagen Type 1 in the layer of the inner half and secreted MMP-1, MMP-2, and TIMP-1 in the layer of the outer half of the capsule. Activation of alpha-SMA positive hepatic stellate cells (HSC) was also observed in the liver parenchyma adjacent to metastases. CONCLUSIONS: The results indicate that fibrotic capsular formation is associated with a lower rate of initial local recurrence in the remnant liver, and that the capsule may serve as a mechanical and chemical barrier to local invasion by metastatic tumor cells. Proliferating stromal cells in the capsule are myofibroblasts, probably derived from HSC activated by colorectal liver metastasis in the liver parenchyma. PMID- 11260866 TI - Identification of a recurrent BRCA1 mutation in German breast-cancer and/or ovarian-cancer families. AB - Specific BRCA1 mutations have been reported to be common within particular populations. We have investigated German breast- and/or ovarian-cancer families and detected a recurrent carboxy-terminal BRCA1 mutation, 5622C > T, using PCR based restriction assay and haplotype analysis. Unrelated families carrying this BRCA1 mutation shared two different disease-associated haplotypes, indicating two independent mutation events. PMID- 11260867 TI - In vitro induction of a bladder cancer-specific T-cell response by mRNA transfected dendritic cells. AB - PURPOSE: To design a tumor-specific immunotherapeutic strategy for treating tumors for which no specific antigens are described (such as bladder urothelial carcinoma), we attempted to activate tumor-specific T-cells by dendritic cells transfected with tumor-derived mRNA. METHODS: Dendritic cells were generated from a patient's peripheral blood and loaded with mRNA derived from the urothelial carcinoma tissue of the same patient. Autologous T-cells were incubated twice on these dendritic cells and tested for their ability to lyse tumor cells. RESULTS: Dendritic cells transfected with tumor-derived mRNA were able to activate T-cells that recognized autologous tumor cells. Cytotoxicity was around 26% for an effector:target ratio of 50:1. Tumor-infiltrating lymphocytes did not kill the autologous tumor cells in vitro, but after a single stimulation with the transfected dendritic cells, they induced tumor cell lysis of 35.7% at an effector:target ratio of 50:1. CONCLUSIONS: These results indicate that dendritic cells transfected with tumor mRNA containing messages for one or more tumor antigens could serve for the ex vivo activation of effector T-cells or directly as vaccines for a wide range of human neoplasias. PMID- 11260868 TI - Two tails of the normal curve. Similarities and differences in the study of mental retardation and giftedness. AB - Professionals in the fields of mental retardation and giftedness have much to teach each other as well as the field of human development in general. Examining the commonalities and differences between the fields in social issues, definitions, developmental differences from the norm, values and policy issues, and educational and long-term implications deepens insights about both normal and deviant development. The authors stress the importance of individual differences in the differential design of educational strategies and the application of approaches developed with specialized populations to normally developing children. Current social inequalities affect both of these fields in particular ways. Finally, numerous research agendas can be enhanced by including representatives of both ends of the normal curve. PMID- 11260869 TI - Reinterpreting individualism and collectivism. Their religious roots and monologic versus dialogic person-other relationship. AB - Examining the religious roots of individualism and collectivism and seeing them as defining alternative conceptions of the person-other relationship reveal a close link between Christianity and the former and between rabbinic Judaism and the latter. Comparisons between these 2 religious formations in the Western world expose a relationship between Christian individualism and an instrumental and monologic understanding of the person-other relationship and a contrasting rabbinic view that offers a formative and dialogic understanding of that relationship. Because the Christian view has been dominant, its understandings have framed the debates on individualism-collectivism and defined the options available for the person-other relationship, providing a somewhat distorted picture of the possibilities for humankind. The dialogic and formative perspective of the rabbinic tradition introduces an alternative portrait of human nature. PMID- 11260871 TI - Gold Medal Award for Life Achievement in the Practice of Psychology. PMID- 11260870 TI - Learning from others. Japan's role in bringing psychology to China. AB - Recent research by Chinese and Japanese historians of psychology and education suggests that it was educational reformers' copying of Japan's education system in the 1st decade of the 20th century that provided the context for developing modern psychology in China. Psychology, although not well understood by those reformers, was thought to be useful in teacher training. In 1902 Japanese psychology teachers came to China and some textbooks were translated. Chinese students studying in Japan also brought back psychological knowledge in translations. However, the Chinese attraction to study in Japan declined after 1906. As the United States opened new universities and provided opportunities for Chinese students to study in U.S. schools, it became a more attractive option for later generations of Chinese, who saw psychology become established as a separate discipline. PMID- 11260872 TI - Guidelines for psychotherapy with lesbian, gay, and bisexual clients. PMID- 11260873 TI - Accredited internship and postdoctoral programs for training in psychology: 2000. PMID- 11260874 TI - Accredited doctoral programs in professional psychology: 2000. PMID- 11260875 TI - [Evaluation of an anti-HIV-1/2 antibodies detection kit based on counting immunoassay]. AB - A novel anti-HIV antibody detection kit, by which anti-HIV-1/2 antibodies were detected based on the counting immunoassay using an auto analyser (PAMIA-50), was evaluated. In an examination using 100 samples each of HIV-1 antibody positive and negative sera, either sensitivity and specificity was 100%. This kit was able to detect all antibodies tested including against HIV-1 subtype A to F, E/F, C/E, B/O, HIV-1 group O, and HIV-2. However, by one of the commercially available kits, one serum containing anti-subtype A antibody was missed. In other comparative studies with the conventional kits using various commercial available panel sera, the same results were obtained. This method was completed within 15 min and was able to examine many samples at one assay. Therefore, this kit is a useful and reliable one if hospitals or institutions had PAMIA-50 in their clinical laboratory. PMID- 11260876 TI - [Clinical and bacteriological studies on hospital outbreak of Salmonella enteritidis food poisoning]. AB - We experienced a hospital outbreak of salmonella food poisoning after ingestion of omelet which was the hospital evening meal on August 8, 1999. Total number of patients was sixty-two (Male 25: female 37) and the mean age was 52.1 years old. Salmonella Enteritidis was isolated from the stool in 59 cases. Twenty-one of them were associated with the immunosuppression (12 with malignancy, 6 with DM, one with nephrotic syndrome, one with chronic nephritis and one with allergic purpura). Clinical symptoms of the patients were composed of watery diarrhea (100%), fever (88.7%), abdominal pain (82.3%), nausea (45.2%) and vomiting (25.8%). The laboratory data revealed leukocytosis (15/47 = 31.9%), increased CRP (44/46 = 95.7%), elevated creatinin (1/37 = 2.7%) and hypokalemia (5/42 = 11.9%). MICs of 20 strains isolated in our laboratory almost coincided with each other indicating that the source of bacteria was probably the same. In vitro, S. Enteritidis were sensitive to OFLX, TFLX, FOM, most of PCs, CEPs, AGs but resistant to MPIPC, CAM, CLDM, VCM. Therefore we administered LVFX to 59 cases (alone in 45cases, combination with FOM in 6 cases), NFLX to two children and FMOX to one pregnant woman. Lactobacillus was administered to 28 cases (45.2%) and antidiarrhetics were given to 6 cases (9.7%). Finally all patients improved within two weeks. We suspect that the salmonella food poisoning was due to infected egg. The partially cooked omelet would permit the growth of a sufficient inoculum to cause disease. To prevent food poisoning, we have to be consistent in cooking the food well (at 75 degrees C, for more than 1 minute) and should not have omelets during the hot summer season. PMID- 11260877 TI - [Serovar-distribution and drug-resistance of Salmonella strains isolated from domestic and imported cases during 1995-1999 in Tokyo]. AB - A total of 2,277 non-typhoidal Salmonella strains consisting of 1,807 domestic strains and 470 imported strains isolated from sporadic cases during 1995-1999 in Tokyo, were examined regarding their serovar-distibution and their drug resistance. The serological typing results showed that the domestic strains were classified into 17 O-groups and 99 serovars, and the imported strains were classified into 12 O-groups and 58 serovars. Among the serovars identified, Salmonella serovar Enteritidis (S. Enteritidis), S. Thompson, S. Hadar, S. Infantis, S. Typhimurium, and S. Litchfield were predominant in the domestic strains, whereas S. Enteritidis, S. Anatum, S. Hadar, and S. Weltevreden were predominant in the imported strains. The drug-resistance test using 9 drugs (CP, TC, SM, KM, ABPC, ST, NA, FOM, and NFLX) showed that 34.0% of the domestic strains and 33.0% of the imported strains were resistant to any of the drugs examined. The serovars of a high resistant rate during this period were S. Blockley (100%), S. Hadar (96.6%), S. Typhimurium (63.6%), and S. Enteritidis (62.2%) in the domestic strains and S. Blockley (100%), S. Hadar (97.1%), S. Rissen (88.9%), S. Emek (83.3%), S. Panama (83.3%), and S. Typhimurium (77.8%) in the imported strains. Drug-resistance patterns of the resistant isolates varied to 60 types. Prevalent patterns recognized were SM, TC.SM, TC, TC.SM.KM.ST, TC.SM.KM, and CP.TC.SM.ABPC in the domestic strains and TC.SM, TC, NA, TC.SM.KM.NA, and TC.SM.NA in the imported strains. PMID- 11260878 TI - [Pathogenic bacteria in the nasal vestivulum of children with acute respiratory tract infection]. AB - The isolation frequency of pathogenic bacteria for acute respiratory infection (ARI) in the pharynx and nasal vestivulum was investigated. Age group-matched children with or without ARI including 109 individuals in each group were examined. Any of the organisms, which are widely regarded as the pathogens causing ARI such as Haemophilus influenzae, Streptococcus pneumoniae, beta haemolytic Streptococcus, Staphylococcus aureus, and Moraxella catarrhalis, were isolated from 91% of the patient group and from 77% of the healthy group. The isolation frequency of S. pneumoniae in the nasal vestivulum of the patient group was outstanding. The healthy carrier rates of S. pneumoniae in the pharynx and nasal vestivulum were 9% and 8%, respectively. Whereas the isolation frequencies from the patient group were 7% and 28%, respectively. alpha-haemolytic Streptococci except S. pneumoniae revealed different tendency from S. pneumoniae. These organisms were almost always isolated from their pharynx but rarely isolated from the nasal vestivulum. The isolation frequency of H. influenzae from the pharynx of the patient group was 41%, which was slightly higher than 34% in the healthy group, but the difference was statistically not significant. H. influenzae was not isolated from the nasal vestivulum of the healthy group, nevertheless it was isolated from 25% of the patient group. The isolation of H. influenzae from the nasal vestivulum may have some important information about ARI. S. aureus was isolated from the pharynx with higher rate than the nasal vestivulum in both groups, and moreover, the isolation frequency of S. aureus in the healthy group was higher than the patient group. It means that the diagnosis of staphylococcal infection should be made very carefully. Considering the results of this study, it could be said that bacteriologic examination of the specimens from nasal vestivulum is valuable to determine S. pneumoniae and H. influenzae as the etiologic agents of ARI. PMID- 11260879 TI - [Bartonella henselae infection in domestic cat and dog fleas]. AB - We studied on the infection of domestic cat and dog fleas with Bartonella henselae by polymerase chain reaction (PCR). A total of 62 fleas (36 Ctenocephalidis felis from cats, 24 C. felis from dogs and 2 Ctenocephalidis canis from dogs), stored in 70% ethanol, were analyzed by PCR for B. henselae specific DNA. Of the 62 fleas, C. felis from cats and dogs were positive for B. henselae specific DNA in 12 of the 36 (33.3%) and in 5 of the 24 (20.8%), respectively, and C. canis from dogs was positive in 2 of the 2 (100%). Our results demonstrated that pet fleas were infected with B. henselae, and suggest that flea transmission of B. henselae between cats or dogs may occur, and direct transmission of B. henselae from pet fleas to human may cause cat scratch disease. PMID- 11260880 TI - [Suppressive effect of lansoprazole on anti-Candida activity of murine macrophages]. AB - This study investigated the influences of lansoprazole (AG1749), a proton pump inhibitor (PPI), and its active derivative (AG2000) on Candida albicans growth and the anti-Candida activity of macrophages. Under concentration of 100 microM, AG1749 and AG2000 had no effect on Candida growth. Murine peritoneal macrophages inhibited the growth of C. albicans in vitro. AG2000 suppressed the anti-Candida activity of macrophages dose-dependently, but AG1749 didn't. The suppressing activity of AG2000 for macrophages was neutralized by adding a SH-compound (L cysteine) in the medium. This suggests that AG2000 may suppress macrophage function in a similar manner with inhibition of proton pump through binding to SH molecules. When macrophages were preincubated with AG2000 for 1 hr and washed, their anti-Candida activity remained to be partially inhibited for 14 hrs. These results were discussed in relation to the pathogenesis of esophageal candidiasis. PMID- 11260881 TI - [Detection of gram-negative bacteria in patients and hospital environment at a room in geriatric wards under the infection control against MRSA]. AB - We prospectively surveyed gram-negative bacteria in patients and hospital environment in a room in the geriatric ward which was specifically under the infection control against MRSA once every two weeks between September and December 1996. We investigated the inpatients in an 8-bed room in the geriatric wards (190 beds) of AINO Memorial Hospital, affiliated with Nagasaki University. During the study period, we performed a total of 431 cultures. The number of specimens cultured was 116 from airways (nose, 42; pharynx, 42; sputum, 32), 24 from decubitus ulcer, 40 from urine, 42 from feces, a total of 125 from skin (head, 42; forearm, 42; inguinal region, 41), and 84 from the hospital environment (floor swab, 42; settled agar plate, 42), respectively. A total of 15 species were isolated from the hospital environment. Some species were the same as those which were recovered from the hospital environment were those observed on each body site. In the hospital environment, the isolation rates of Acinetobacter baumanii and Klebsiella pneumoniae were significantly high in the settled agar plate (A. baumanii, p < 0.01; K. pneumoniae, p < 0.05, respectively). Isolation rates of Pseudomonas aeruginosa, Citrobacter spp., and Enterobacter sakazakii were also high in the settled agar plate (p = 0.078, 0.078, 0.078, respectively). In conclusion, gram-negative bacteria in patients may be associated with the environmental bacteria in the room in the geriatric wards. PMID- 11260882 TI - [A case of loiasis]. AB - Loiasis is quite common in the endemic regions of Central and West Africa. But only three cases were reported in Japan. This is a report of a 28 year old male from Gabon infected with Loa loa with eye symptoms as the chief complaint. For the first time in Japan he was treated with Ivermectin (IVM) which is recently attracting attention as the drug for filariasis world wide. IVM therapy was effective, and decreased the counts of microfilarias in the patient's blood. No adverse effect was seen in this patient. This case suggested that IVM is an useful drug for loiasis, and further study is warranted. PMID- 11260883 TI - [Two cases of cardiac aspergillosis with initial onset of arrhythmia during therapy of acute leukemia]. AB - We have reported two women, aged 86 and 84 years, with cardiac aspergillosis with initial onset of arrhythmia during chemotherapy of acute myeloblastic leukemia and primary plasma cell leukemia, respectively. In leukopenia followed by chemotherapy, they suddenly had arrhythmias with high fever. The former had cardiac infarction with complete atrioventricular block and the latter was also cardiac infarction following to atrial fibrillation. In both cases, cardiac aspergillosis was not diagnosed by echocardigraphy but by autopsy. Since cardiac aspergillosis dose not have characteristic features clinically or examinationally, we need to consider arrhythmias revealed in leukopenia as one symptom of cardiac aspergillosis. PMID- 11260884 TI - [Preparation of Chlamydia pneumoniae in roller bottles with high IFUs]. PMID- 11260885 TI - [Current topics on classification and nomenclature of bacteria. 4. Relations between classification and nomenclature of bacteria]. PMID- 11260886 TI - [Importance of a cleaning in upper airways by using povidone iodine for the prevention of nosocomial pneumonia]. AB - We investigated the efficacy of infection control measures against nosocomial pneumonia in geriatric wards. Cases with nosocomial pneumonia were retrospectively analyzed between January 1991 and March 1995. The study period was divided into four annual periods (periods 1, 2, 3 and 4). Period 1, January to December 1991, was applied as the cotrol. We investigated patients with nosocomial pneumonias in geriatric wards (190 beds) of AINO Memorial Hospital, affiliated with Nagasaki University. During the study period, nosocomial pneumonia significantly diminished. (period 1 vs periods 2, 3 and 4, p < 0.05, p < 0.05, p < 0.05, respectively). Major causative organisms of nosocomial pneumonia were MRSA and Pseudomonas aeruginosa. During the four periods, a significant reduction in cases with MRSA- and P. aeruginosa-induced nosocomial pneumonia was observed (MRSA: period 1 vs periods 2, 3 and 4, p < 0.05, p < 0.05, p < 0.01, respectively; P. aeruginosa: period 1 vs period 3, p < 0.01, period 2 vs periods 3 and 4, p < 0.01, p < 0.05, respectively). On the other hand, the improvement of decubitus ulcers was associated with a significant reduction in nosocomial pneumonia (period 1 vs. periods 2 and 3, p < 0.05 and p < 0.05, respectively). In conclusion, stringent infection control programs, including a cleaning in the upper airways by povidone iodine, are necessary in geriatric wards to reduce and prevent nosocomial pneumonia. PMID- 11260887 TI - [Surgery for cerebral hemisphere astrocytomas]. PMID- 11260888 TI - [Aneurysms of the distal posterior inferior cerebellar artery--analysis of 14 aneurysms in 13 cases]. AB - Thirteen cases of distal posterior inferior cerebellar artery (PICA) aneurysms are reported here. All the aneurysms were found after a subarachnoid hemorrhage. Dissecting aneurysm, incidentally found unruptured aneurysms, and aneurysms associated with arteriovenous malformation have been eliminated from this study. Characteristics for this type of lesion are a high rate of recurrent hemorrhage and rapid death due to direct compression of the brain stem, which clearly indicates the necessity of early surgery. Attention should be paid to the fact that angiography cannot always reveal aneurysms, especially when they are located in the peripheral PICA. One should also pay attention to multiple lesions and rapid growing acute subdural hematoma as initial findings for ruptured distal PICA aneurysm. Prognostic factors for these lesions are, vasospasm, especially when the aneurysm is located proximally in the PICA, and direct compression of the brain stem due to intraventricular hemorrhage when the aneurysm is located distally. It has been suggested that the pathogenesis of this lesion could be hemodynamic stress or embryogenesis. The shape and anomalous arterial structures of the 14 aneurysms presented here tend to agree with this suggestion. Our results suggest that the pathogenesis is hemodynamic stress that had developed due to embryological and/or arteriosclerotic factors. PMID- 11260889 TI - [The relationship between delayed traumatic intracerebral hematoma and coagulopathy in patients diagnosed with a traumatic subarachnoid hemorrhage]. AB - It has long been recognized that a traumatic insult to brain tissue may result in substantive coagulation abnormalities. The present study was carried out in an attempt to find out the association of coagulopathy and the development of delayed traumatic intracerebral hematoma (DTICH) in patients diagnosed with a traumatic subarachnoid hemorrhage (TSAH). Sixty-three patients were diagnosed as having TSAH from the initial CT scans obtained within 2 hours after trauma. On admission, peripheral blood samples for coagulation studies were taken within 6 hours after injury. All patients had subsequent CT scans performed within 24 hours of admission. Thirty (47.6%) of 63 patients exhibited radiological evidence of DTICH on their subsequent CT scans. There was a significant correlation between the increased value of serum fibrinogen degradation product (FDP > 40 micrograms/ml) and the development of DTICH. We observed that the origin of the hematoma might be caused by those radiographically unidentifiable parenchymal lesions often found with TSAH on the initial CT scan. We conclude that a clotting study at the time of admission is of value in predicting the occurrence of DTICH associated with TSAH. PMID- 11260890 TI - [EDAS (encephalo-duro-arterio-synangiosis) for occlusive/stenotic cerebrovascular disease]. AB - The efficacy of encephalo-duro-arterio-synangiosis (EDAS) using superficial temporal artery was evaluated for the treatment of the occlusive/stenotic cerebrovascular disease. Nine patients with the occlusive/stenotic cerebrovascular disease underwent EDAS in our hospital. The mean follow-up period was 6.6 months. Postoperative angiography showed no collateral formation via EDAS in any of the nine patients. We analyzed the following points: 1) operative procedure, 2) follow up period after surgery, 3) preoperative cerebral blood flow, and 4) age of the patients. Results showed that EDAS as a treatment of occlusive/stenotic cerebrovascular disease was not effective. This study failed to reinforce the suggestion that indirect extracranial/intracranial bypass surgery is effective as the treatment of occlusive/stenotic cerebrovascular disease. PMID- 11260891 TI - [A case of convexity meningioma en plaque]. AB - We present a case of convexity meningioma en plaque (MEP). A 51-year-old male occasionally suffering from right parietalgia and numbness of left upper limb. An intracranial abnormal mass was pointed out incidentally by the brain check up. Computed tomographic (CT) scans demonstrated a hyperostosis and an enhanced abnormal mass at the right front-parietal region. Magnetic resonant images (MRI) revealed a carpet like tumor extended along the dura mater. Cerebral angiography disclosed feedings from parietal branches of right middle meningeal artery and superficial temporal artery. The tumor was removed subtotally with adjacent dura mater, leaving the portion of close adhesion to the brain parenchyma. Histologic diagnosis was transitional meningioma. Immunohistological stainings showed a high staining index (6.9%) of MIB-1 (Ki-69 antigen) and high expression of metalloproteinase-9 (MMP-9), especially along the dura mater. Convexity MEP is so rare that we review previous reported cases of convexity MEP, and discuss the clinicopathologic features on that. PMID- 11260892 TI - [Brain abscess and ventriculitis associated with entrapment of the lateral ventricle appearing more like remarkable brain edema than ventricular dilatation- a case report]. AB - We present a case with brain abscess associated with entrapment of the lateral ventricle appearing more like remarkable brain edema in the temporo-occipital lobe than ventricular dilatation. A 72-year-old man suffering from headache and vomiting visited our clinic. CT and MRI showed brain abscess in the right parieto occipital lobe, associated with ventriculitis. Lumbar puncture also revealed purulent meningitis. Both symptoms and CSF findings improved after administration of antibiotics. The improved condition continued for two months after admission, but disturbed consciousness and left hemiparesis than appeared. MRI and CT showed entrapment of the lateral ventricle and brain edema of the right temporo occipital region without ventricular dilatation. Because brain edema was thought to be caused by transudate of the CSF through the ventricular wall, lobectomy of the right temporal lobe and opening of the temporal horn were carried out. Although left hemiparesis and disturbed consciousness and brain edema disappeared after the operation, subdural effusion appeared. Using a subdural-peritoneal shunt, the subdural effusion was prevented and disappeared. In this case, we thought Hounsfield Unit (HU) of the brain edema caused by transudate of CSF through the ventricular wall (12.6) was markedly lower than that of so-called vasogenic edema (25.1) due to active inflammation. Measurement of the HU seemed to be a useful means to differentiate the types of brain edema in this situation from that of vasogenic edema caused by brain abscess, and thus a means for selection of the appropriate treatment. PMID- 11260894 TI - [True posterior communicating artery aneurysm]. AB - We report a case of a true posterior communicating artery aneurysm. A 51-year-old male suffered a subarachnoid hemorrhage with severe headache and vomiting. A true posterior communicating artery aneurysm was recognized after repeated angiography on the seventh day. Right frontotemporal craniotomy was performed and the aneurysm was successfully clipped. The incidence of true posterior communicating artery aneurysms ranges from 0.1-2.8%, and 21 cases including our case have been reported in detail. There are no reported cases in which the aneurysm arises from the branching site of perforating arteries. In almost all cases the dome of the aneurysm projects inferiorly or posteriorly or laterally, so perforating arteries from the posterior communicating artery rarely interfere with dissection of the aneurysm or neck clipping. In a few cases, true posterior communicating artery aneurysms had been diagnosed as IC-PC aneurysms preoperatively, leading to intraoperative aneurysmal rupture or postoperative neurological deficit or death. In the cases of a fusiform aneurysm or an aneurysm of wide-based neck, there may be no other choice than trapping of the aneurysm. It is difficult to predict whether trapping causes postoperative ischemic complications. PMID- 11260893 TI - [Persistent primitive hypoglossal artery aneurysm--case report]. AB - The aneurysm arising from a persistent primitive hypoglossal artery (PHA) is rare, and only 13 such cases have been reported in literature. We present a 62 year-old woman with an aneurysm of PHA at its junction with the basilar artery. The patient consulted our hospital for a transient loss of consciousness and headaches. No neurological deficit was found, but MRI and MRA showed an aneurysm of the vertebrobasilar junction. Cerebral angiogram after admission showed the aneurysm of PHA at its junction with the basilar artery. Perspective 3D-CTA and 3D-T2 weighted MR images were composed to simulate the condition and aneurysmal surgery via the transcondylar approach was carried out. The aneurysm was successfully clipped and the patient was discharged with no neurological deficits. Perspective 3 D-CTA and MRI simulation were very useful for this operation. PMID- 11260895 TI - [Ruptured dissecting aneurysm of the vertebral artery concurrent with contralateral intracerebellar hemorrhage]. AB - A case of a dissecting vertebral aneurysm concurrent with contralateral cerebellar hemorrhage is reported. A 69-year-old man was referred to our hospital for treatment of subarachnoid hemorrhage (SAH). On admission, CT scanning showed SAH and left cerebellar hematoma. Angiography was performed and it revealed a dissecting aneurysm of the right vertebral artery. Proximal clipping of the right vertebral dissecting artery was performed through right suboccipital craniotomy. During the operation, the cerebellar hemisphere gradually became firm, but the operation was finished without any complications. After the operation, the patient's consciousness level decreased from somnolence to semicomatose for a period of 2 hours 30 min. CT scanning showed the left cerebellar hematoma expanding. The cerebellar hematoma was evacuated immediately by midline suboccipital craniectomy, and the patient's consciousness level improved. In such a case, care must be taken to discover the cause of the expansion and to prevent concurrent hematoma during the operation. Through this case, discussion was held concerning the pitfalls of treatment of aneurysmal subarachnoid hemorrhage concurrent with intracerebral hematoma in the remote region. PMID- 11260896 TI - [Increased intracranial pressure caused by obstruction of torcular herophili with hemangiopericytoma: a case report]. AB - A 49-year-old male had experienced diplopia for half a year. The intracranial pressure was markedly elevated (450 mmH2O). Neuroimaging revealed a tumor incompletely occluding the torcular herophili and the bilateral transverse sinuses without cerebral or cerebellar compression by the tumor. Both cortical veins and cervical veins were enlarged, and the Sylvian vein and Rabbe's vein and the tentorial sinus were collateral vessels. Biopsy was performed and histologic examination proved hemangiopericytoma. The patient underwent Gamma-knife treatment and the tumor decreased in size 3 months after the treatment. PMID- 11260897 TI - [Delineation of cerebral aneurysms with fly-through imaging of 3D-MRA using perspective volume rendering]. AB - We used fly-through (FT) imaging of three-dimensional MR Angiography (3D-MRA) with perspective volume rendering to delineate cerebral aneurysms. Four unruptured cerebral aneurysms discovered incidentally on MRA were delineated as FT images and compared with maximum intensity projection (MIP) images. FT imaging was superior to MIP in the realistic representation of cerebral aneurysms from an extraluminal point of view. FT imaging of 3D-MRA may be helpful for the spatial representation of an aneurysm, thereby improving diagnostic accuracy of MRA for cerebrovascular diseases. PMID- 11260899 TI - [Hepatopancreatoduodenectomy (HPD)]. AB - Since the mid-1980s, simultaneous major resection of the liver and the pancreatic head, so-called HPD, has been performed in Japan to improve the survival rate of patients with advanced biliary tract carcinoma. We conducted a study of HPD in dogs in our laboratory from 1986 to 1989 and found that 75% died following 70% hepatectomy with more than 92% pancreatectomy and that only 25% survived. The main cause of death was liver failure. A clinical review of the operative procedures and the outcome of HPD was performed by Mizumoto et al. in our department in 1989. They collected data on 241 patients who underwent HPD for advanced biliary tract carcinoma throughout Japan. The postoperative morbidity and mortality rates after HPD were higher than after other major surgeries. The prognosis after HPD has improved in recent years because of preservation of adequate hepatic or pancreatic parenchyma, improvements in surgical technique, and strict perioperative care. Therefore we plan to review the operative procedures and outcome of HPD again in 2000. PMID- 11260898 TI - [Endovascular treatment for facial arteriovenous malformations using new particles: report of two cases]. AB - Facial arteriovenous malformations (FAVM) are difficult to treat because of their highly vascular networks. Intravascular treatment using liquid material to occlude the FAVM occasionally results in skin necrosis after embolization. The use of particulate materials to obliterate the nidus often fails to obtain a permanent cure due to arterial recanalization. We report two patients with FAVM who were successfully treated with endovascular embolization using a new type of particulate material. One patient was treated with embolization only, the other was treated with embolization followed by surgical resection. Both patients showed clinical and angiographic improvement. Intravascular treatment using particles with a smooth surface and optimal size is safe and effective in the treatment of patients with FAVM. PMID- 11260900 TI - [Indication for and problems of hepatopancreatoduodenectomy for carcinoma of the biliary tract based on the statistical registry in Japan]. AB - Hepatopancreatoduodenectomy (HPD) as radical surgery for advanced carcinoma of the biliary tract was previously eschewed due to the high rate of postoperative complications. However, recently many institutes have performed it due to the improvement of operative procedures, such as hepatectomy and pancreatoenterostomy, and of pre-intra-postoperative management. Four hundred and sixty-five patients undergoing HPD were registered in Japan during the past 10 years, of whom 355 had carcinoma of the gallbladder and 110 carcinoma of the bile duct. The 30-day operative mortality rate was 9.2% (43 patients). The 5-year survival rates according to the Kaplan-Meier method was 18.1% (32 patients). Survival rates of those with ss and se or si gallbladder cancer were 36% and more than 10%, respectively, but that of those with se or si bile duct cancer was less than 6%. Only 3 patients with 16 lymph node metastases survived for more than 5 years. Fewer patients with biliary infiltration survived for more than 5 years compared with those with hepatic infiltration in carcinoma of the gallbladder. For such patients, extended surgery combining so-called total resection of the hepatoduodenal ligament is thought necessary. PMID- 11260901 TI - [Possible inhibitory mechanism of liver regeneration through non-parenchymal cells after combined hepatectomy and pancreatectomy]. AB - BACKGROUND: Recently, simultaneous hepatectomy (Hx) and pancreaticoduodenectomy have been performed in the treatment of biliary tract cancer. Postoperative hepatic failure is a common and potentially fatal complication. The aim of this study was to examine the reduced rate of liver regeneration after 70% Hx alone or in combination with 70% pancreatectomy (HPx). MATERIALS AND METHODS: Male Sprague Dawley rats underwent Hx or combined Hx and Px. The ratio of liver-body weight, labeling index of hepatocytes in vivo, and DNA synthesis of hepatocytes and/or Kupffer cells in primary culture were analyzed. RESULTS: The ratio of liver-body weight in HPx rats was found to be significantly lower than that in Hx rats from 12 hours to 72 hours after surgery. There was no difference in blood glucose or ALT levels between the two groups. An inhibitory effect on DNA synthesis was observed in cocultured hepatocytes and Kupffer cells when portal plasma obtained one hour after surgery was added. We further observed that conditioned medium of Kupffer cells stimulated by portal plasma obtained one hour after HPx inhibited DNA synthesis by hepatocytes. This effect was abolished after incubation at 56 degrees C for 30 min. CONCLUSIONS: These results clearly indicate the existence of a growth inhibitory factor in portal serum after HPx. This heat-labile growth inhibitory factor was released from Kupffer cells stimulated by portal plasma after HPx and appears to act on hepatocytes in a paracrine manner. PMID- 11260902 TI - [Indications for and operative outcome of hepatopancreatoduodenectomy in the treatment of gallbladder carcinoma]. AB - Hepatopacreatoduodenectomy (HPD) was initially performed to resect highly advanced gallbladder carcinoma with direct invasion of the liver and head of the pancreas. High operative morbidity and mortality rates and early recurrence were major problems of this procedure. However, the operative outcome gradually improved with progress in surgical procedures and perioperative management. Recently, HPD has been indicated not only for direct invasion of the liver and pancreas but also for intensive dissection of peripancreatic lymph nodes and resection of occult liver metastasis to subsegments IV and V. Evaluation of all cases in which HPD was performed in our institute suggests that advanced gallbladder carcinoma with lymph node metastasis and without high-grade infiltration of the hepatoduodenal ligament (binf) is the most suitable indication for HPD. PMID- 11260903 TI - [Indication and outcome of extended right hemihepatectomy combined with pancreatoduodenectomy for bile duct carcinoma]. AB - During the past 10 years, we have performed extended right hemihepatectomy combined with pancreatoduodenectomy (rtHPD) in eight patients with bile duct carcinoma. We compared the results in these patients with those in 43 bile duct carcinoma patients who underwent extrahepatic bile duct resection with more extensive hepatectomy than hemihepatectomy. Our indication for rtHPD is bile duct carcinoma of the diffuse type involving the intrapancreatic bile duct. For patients with obstructive jaundice, biliary drainage was performed preferentially in the part of the liver to be preserved. Portal vein embolization was performed before extended right hemihepatectomy or left trisectorectomy. Complete external drainage of pancreatic juice followed by second-stage pancreatojejunostomy was performed in five rtHPD patients. There were no hospital deaths or hepatic failures. There were four 5-year survivors after rtHPD. There was no significant difference between the cumulative 5-year survival rates after rtHPD (71%) and non HPD (42%). Patients with bile duct carcinoma whose prognosis can be improved only by rtHPD exist and should be treated by rtHPD. However, considering the reported high mortality rate after this procedure, rtHPD should not be performed in an institution where its safety cannot be guaranteed. PMID- 11260904 TI - [Post operative complications after hepatopancreatoduodenectomy (HPD)]. AB - The postoperative complications, morbidity and mortality of hepatopancreatoduodenectomy (HPD) are reviewed based on reports by the Japanese Biliary Surgery Association, Japanese Pancreatectomy Association, and leading surgeons. Postoperative hepatic failure, the most important and lethal complication, is significantly correlated with patient age (older than 70 years), resected hapatic volume (hepatic bisegmentectomy or greater), combined resection of the portal vein, or temporal bypass of portal blood flow. In patients who undergo HPD with more than bisegmentectomy, the operative mortality rate is high rate, with reports of 39.7%, 23.0%, 12.5%, and 38.0%. To reduce the morbidity and mortality rates after HPD, it is important to avoid intraoperative hepatic ischemia and to maintain sufficient hepatic blood flow and high oxygen saturation of portal blood postoperatively using a respirator and inotropic agents. PMID- 11260905 TI - [Clinical evaluation of hepatopancreatoduodenectomy]. AB - The clinical outcome of hepatopancreatoduodenectomy (HPD) carried out in 14 patients in our institute was evaluated retrospectively. In principle, HPD is performed in patients with far-advanced biliary carcinoma extending from the level of the liver to pancreas directly or via metastatic lymph nodes around the head of the pancreas. However, the survival periods after surgery did not improve, contrary to expectations. Curative surgery was not achieved in most patients who underwent HPD, even if resection of the portal vein or hepatoduodenal ligament was performed during surgery. Severe lymphatic, vascular, and neural involvement along the hepatoduodenal ligament existed in those patients. In addition, it is possible that they already had micrometastases to other organs based on a study of the pattern of recurrence after surgery. Quality of life did not appear to improve unless the patients survived more than one year. These findings suggest that HPD may be appropriate for patients in earlier disease stages than those in our series to achieve longer survival. However, HPD is associated with high rates of postoperative complications and operative mortality, especially after pancreatoduodenectomy with liver resection involving more than right hepatectomy. We need to establish the appropriate indications for HPD and improve the safety of the procedure. PMID- 11260906 TI - [Portal embolization and second-stage pancreatojejeunostomy improve operative outcomes in patients with advanced biliary carcinoma]. AB - Lethal complications after resection of more than two-thirds of the hepatic parenchyma with pancreatoduodenectomy (PD) are hepatic failure and leakage from pancreatoenterostomy. Twenty-two patients underwent hepatectomy with PD due to advanced biliary carcinoma. Extended hemihepatectomy (EH) with PD was performed in 16 and other types of hepatectomy with PD in 6. The percent volume of the liver to be preserved increased from 31.5% to 43.5% after portal venous branch embolization, which was performed to prevent postoperative hepatic failure. There were no operative and hospital deaths in these 22 patients. Only one patient had a prolonged serum total bilirubin level elevation postoperatively, although the level was less than 5.0 mg/dl. Since leakage from pancreatogastrostomy occurred in 2 of 4 patients who underwent pancreatogastrostomy, second-stage pancreatojejunostomy was chosen to avoid pancreatic fistula in the remaining 18 patients. In 11 patients who underwent EH with PD due to diffuse bile duct carcinoma, the cumulative 1-, 3-, and 5-year survival rates were 90.9%, 64.9%, and 64.9%, respectively. Because EH and PD provides an opportunity for long-term survival for patients with diffuse bile duct carcinoma, prevention of lethal postoperative hepatic failure and leakage from pancreatoenterostomy is important. PMID- 11260907 TI - [Trends in laparoscopic surgery for colorectal cancer: 10-year experience worldwide]. AB - Laparoscopic colorectal surgery was introduced in 1991 and has been performed around the world. We review the English literature on this technique, including reports of case series, case-control studies, and randomized controlled trials. These reports show that laparoscopic surgery for colorectal cancer is safe and useful and that the short-term results of this technique are acceptable and satisfactory. Port-site metastasis is rare in the more recent experience of skilled laparoscopic surgeons, and the survival rates of patients after laparoscopic surgery are comparable to those after conventional open procedures. In the near future, long-term results, including 5-year survival rates, will be clarified in randomized controlled trials comparing laparoscopic and open resections of colorectal cancer. PMID- 11260908 TI - [Role of angiotensin II-forming pathway in ruptured aortic aneurysms]. PMID- 11260911 TI - [Learning from the studies of twitcher mouse]. AB - Twitcher mouse is a naturally occurring mouse model for human Krabbe disease, a lysosomal galactosyl-ceramidase deficiency. Therefore, this mouse provides us an excellent experimental model for studying the pathogenesis and effective therapies for Krabbe disease. First, we succeeded to clone cDNA of human galactosylceramidase from lymphocytes, and determined its sequence. Consequently, many mutations of Krabbe disease have been discovered. For the trial of gene therapy for twitcher mouse, a recombinant retrovirus vector containing human galactosylceramidase cDNA was constructed and targeted to bone marrow cells ex vivo. These cells were transplanted intraperitoneally into twitcher neonates. This protocol resulted in slightly better weight gain and increase in the enzymatic activity in the peripheral nerve in the gene therapy group, but the survival period was prolonged. These results suggest that the efficacy of viral transfection is critical for the gene therapy. In addition, in order to understand the pathogenesis of demyelinating process and to evaluate the gene therapy more precisely, we examined the pathophysiology of oligodendrocytes and their related cells in the nervous system of twitcher mouse. We introduced pi glutathione-S-transferase immunostaining for the specific identification of oligodendrocytes. Using this method, the oligodendrocytes with multiple varicose processes were recognized in the early stages. With the progression of the disease, these cells became shrunk showing the ultrastructural and biochemical characteristics of apoptosis. This may provide a key to the future treatment of Krabbe disease. PMID- 11260912 TI - [Neural stem cell, as a source of graft material for transplantation in neuronal disease]. AB - Self-renewing and multipotent neural stem cells are present in the adult human brain. We successfully harvested neural stem cells from mice and humans using misexpressed EGFP proteins under the control of the nestin second intron enhancer. High-level EGFP expressors derived from mouse embryos included a distinct subpopulation of cells that were self-renewable and multipotent. Further, we obtained that neural progenitor cells from rat fetal spinal cords using a neurosphere technique, and demonstrated their ability to divide and differentiate into neurons in vivo, where they were integrated into the host tissue in the injured rat spinal cord with resultant behavioral improvement of the recipient rat. We also harvested tyrosine hydroxylase-positive neurons from a transgenic mouse expressing GFP under the control of the tyrosine hydroxylase promoter, and successfully transplanted them into the striatum of rats with parkinsonism with marked improvement of the neurological symptoms. Since neural stem cells can adapt well in the host CNS, studies should focus on their application as a vector in gene therapy and on the introduction in vivo or ex vivo of genes to control their proliferation and differentiation. Neural stem cells are a potential, useful source for developing new therapy for CNS disorders. PMID- 11260913 TI - [Surgical treatment of refractory childhood epilepsy with special reference to the role of pediatric neurologists: introductory remarks]. AB - There are many differences between children and adult patients regarding the selection of candidates for epilepsy surgery, decision about the timing of surgery, pre- and intra-surgical evaluation and follow-up. A comprehensive approach by an epilepsy surgery team consisting of pediatric neurologists, neurosurgeons and other medical staffs is absolutely necessary for successful surgical treatment of refractory childhood epilepsy and the improvement of the patient's quality of life. The potential risks and benefits of the surgery must be carefully weighed for each child from various aspects. Pediatric neurologists should make a more active contribution to this whole process. PMID- 11260914 TI - [Surgical treatment of medically refractory epilepsy in childhood]. AB - Twenty five percent of children with epilepsy continue to seize despite best medical management and may be defined as medically refractory. Many children with medically refractory localization-related epilepsy, i.e. seizures which originate in a particular area of brain and secondarily spread to involve other brain regions, may benefit from a variety of surgical treatments including hemispherectomy, corpus callosotomy, focal cortical resection of the temporal lobe, focal cortical resection of extratemporal regions of brain, and multiple subpial resections. A successful outcome from epilepsy surgery is generally defined as a seizure-free state with no imposition of neurologic deficit. In order to achieve these twin goals two criteria must be fulfilled. First, precise localization of the epileptogenic zone in the brain is necessary. The epileptogenic zone may be defined as the region of epileptogenic cerebral cortex whose removal will result in a seizure-free state. Second, one must determine the anatomic localization of eloquent cortex in brain in order to spare these areas during any planned cortical excision of epileptogenic cortex. Several diagnostic measures may be used to achieve a successful surgical outcome. A clinical history to ascertain the earliest symptom in the clinical progression of the seizure (semiology) is imperative as is ictal and interictal scalp EEG, neuropsychological testing, magnetic resonance imaging (MRI), positron emission tomography (PET), single photon emission computerized tomography (SPECT), interictal magnetoencephalography (MEG). In the typical child undergoing evaluation for epilepsy surgery, if the clinical, neuropsychological, EEG, and radiological data are all concordant and point to the same area of epileptogenicity in brain, cortical excision of the suspected epileptogenic zone is undertaken. However, if the data are discordant, and/or the epileptogenic zone resides wholly or in part within eloquent cortex, invasive intracranial monitoring from depth and/or subdural electrodes during a seizure is required to map out the areas of epileptogenicity in brain. The assessment of potential risks and benefits for this type of epilepsy surgery in children involve complex age related issues, including the possible impact of uncontrolled seizures, medication, or surgery, on learning and development. PMID- 11260915 TI - [Pre- and intra-operative evaluation of epileptic children with intractable seizure disorders: the hospital for sick children]. AB - The pediatric epilepsy management team in the Hospital for Sick Children, Toronto, Canada, consists of neurologists, neurophysiologists, neurosurgeons, neuropyschologists, clinical nurse specialist/nurse practitioners, social workers, EEG technologists and psychiatrists. The patients are initially referred to us for the diagnosis of seizure disorders. Epileptic foci and eloquent cortices are identified by neurophysiological studies such as EEG, MEG and SEP. Epileptogenic lesions can be visualized by MRI, the language, motor and sensory cortices by fMRI and the regions of hypoperfusion and hypometabolism in the epileptic foci, by SPECT and PET, respectively. The results of these studies are then discussed by members of the team. For patients with lesional epilepsy, an intraoperative image guided system and intraoperative electrocorticography are used, when lesionectomy, lobectomy and additional multiple subpial transection (MST) are performed. Patients without an identifiable lesion require intracranial invasive video EEG using subdural grids or depth electrodes, which are constructed based on MEG spike sources, seizure semiology and scalp video EEG. After the identification of the epileptogenic and functional zones, maximum cortical excision and MST are performed to control seizures and to minimize functional deficits. Pediatric neurologists should assess the intractability of epilepsy, identify the epileptogenic zone, determine the excisable epileptic region, and minimize postoperative side effects, thereby leading the epilepsy management team. PMID- 11260916 TI - [Surgery for intractable epilepsy in children: pre- and perioperative evaluations with special reference to dipole tracing method estimated from interictal spike]. AB - Methods of preoperative and perioperative evaluation methods for surgical treatment of intractable epilepsy in children are described. Among non-invasive diagnostic methods, EEG-video monitoring is the most fundamental. Amygdalohippocampal volume measurement by MR was useful for the differential diagnosis of mesial temporal lobe epilepsy (TLE) from lateral TLE and generalized epilepsy. The dipole tracing method with a realistic head model was useful for identification of epileptic foci from the interictal spikes of scalp EEG, when an abnormal electric source was estimated as an equivalent current dipole (ECD) in the brain of patients with organic lesion and TLE. ECD concentration ratio ranged from 70 to 90% within 20 mm around the lesion. After lesionectomy seizures disappeared in every patient. The mean distance between the centers of the ECD and epileptic focus (identified by subdural electrode recording) was 14 mm (range: 8 to 18 mm). ECDs of mesial TLE were located in the temporal base rather than mesial temporal lobe, whereas those of lateral TLE in the lateral cortex precisely. In unilateral, intermediate and bilateral TLE, 76%, 52% and 36% of ECDs were localized in the ictal onset zone respectively (p = 0.007). Electrical cortical stimulation with chronically placed intracranial electrodes was used to accurately identify eloquent areas to avoid postsurgical complications. Immediately after operation, 10 to 20% of patients showed better or deteriorated results in neuropsychological examinations, which recovered in all patients after one year. Postoperative seizures were absent in three fourths of patients. Further efforts are needed to obtain better seizure control in future. PMID- 11260917 TI - [Magnetoencephalography analysis of epileptic foci]. AB - Magnetoencephalography (MEG) is very useful for presurgical evaluation for epilepsy surgery because of its high spatio-temporal resolution. Interictal spikes are usually analysed by MEG, whereas ictal studies are performed using EEG or SPECT. We investigated cases in which MEG and other physiological or radiological examinations revealed discordant results. The most important is precise clinical symptomatology. When combined with clinical informations, MEG can be a powerful tool for analyzing epileptic foci. Now pediatric epileptologists should discuss and treat intractable cases with epilepsy surgeons. In Japan, there are only few integrated, epilepsy diagnosis and treatment teams (comprehensive epilepsy programs). They should be widely established in university hospitals and reference centers. PMID- 11260918 TI - [Surgical treatment for pediatric intractable epilepsy--focusing on modified functional hemispherectomy in infants]. AB - Our experience with modified functional hemispherectomy in 14 infants are presented. The etiology of intractable epilepsy was cortical dysgenesis in all of the cases. We applied our new surgical method, transopercular hemispherotomy, in which the frontal operculum was resected en bloc including the upper half of the insula. The corpus callosum was totally sectioned through the lateral ventricle. The resection cavity was communicated to the inferior ventricle, and the medial temporal structures were resected. Finally, the horizontal fibers emerging from the frontal lobe were sectioned along the posterior edge of the ala minor ossis sphenoidalis. There was no mortality or morbidity during and immediately after operation. In 12 cases with more than 1 year follow-up, remarkable seizure reduction was obtained in 75% of the cases and worthwhile improvement in the remaining 25%. No major complications were encountered, except for hydrocephalus in 3 cases and incomplete section of the callosum in 3. In cases showing catch-up of psychomotor development after surgery, swelling of the unaffected hemisphere was observed. PMID- 11260919 TI - [Role of pediatrician in surgical treatment of children with intractable seizures]. AB - Surgery is a useful strategy in the management of intractable and disabling seizures in childhood. Identification of suitable candidates for epilepsy surgery and consideration of the timing of surgery are an important role of pediatrician in surgical treatment of epileptic children. To discuss the timing of surgery, we analyzed clinical courses of 126 patients with TLE and 33 patients with FLE who underwent epilepsy surgery. In many of them seizures continued without remission before surgery. Surgery might have been considered earlier in these cases. Thirty five (27.8%) of 126 patients with TLE and 10 (30.3%) of 33 patients with FLE had seizure remission before surgery. In TLE, seizure remission was rare in the patients who had convulsive seizures at the onset. In conclusion, the timing of surgery should be determined on individual basis in cases with more than one remission. If deletorious effects of seizures on patient's biological and psychosocial state are evident, surgery should be considered earlier. PMID- 11260920 TI - [A case of unilateral moyamoya disease presenting with hemichorea]. AB - We reported a 12-year-old boy with unilateral moyamoya disease whose initial and predominant manifestation was hemichorea. Neurological examinations revealed chorea in his left upper extremity and muscle hypotonia in his left upper and lower extremities. Cranial MRI showed moyamoya vessels only in the right basal ganglia and infarction in the white matter of the right frontal lobe. Right carotid angiography revealed stenosis in the distal part of internal carotid artery, and in the proximal part of anterior and middle cerebral arteries with moyamoya vessels. Left carotid angiography showed normal findings. He was diagnosed as a suspected case of moyamoya disease (unilateral moyamoya disease) according to the diagnostic criteria proposed by the Research Committee on Moyamoya Disease of the Ministry of Health and Welfare of Japan. His chorea responded to haloperidol but encephalo-duro-arterio-synangiosis on the right side improved all symptoms. Chorea occurs in some patients with moyamoya disease. Hypofunction of the striatal indirect pathway is suggested as the cause of chorea. In this case an ischemic lesion in the right striatum may have caused hypofunction of the pathway and developed chorea and hypotonia. PMID- 11260921 TI - [Acute disseminated encephalomyelitis in a 3-month-old infant]. AB - Acute disseminated encephalomyelitis (ADEM) is a demyelinating disease showing multifocal central nervous system lesions due to an autoimmune disorder. We reported a 3-month-old girl with ADEM. One week after having a cold, she presented with somnolence, poor feeding and vomiting. When she was admitted three days after the onset, she could neither fix or follow objects with her eyes nor respond to sound. Her muscle tone was decreased. Cerebrospinal fluid examination revealed pleocytosis, elevated protein concentration and positive myelin basic protein. No oligoclonal band was detected. Diffuse monomorphic slow wave activity was noted on the electroencephalogram. Only wave I was present bilaterally on the auditory brainstem response. T2 weighted images of magnetic resonance imaging revealed multiple areas of high signal in the right posterior limb of the internal capsule, white matter of the cerebellum and brainstem. She was diagnosed as having ADEM, and underwent high dose gamma-globulin therapy. Corticosteroids were not given because of her high blood pressure. The clinical symptoms improved continuously before and after the administration. Two years after the onset, she showed normal growth and development without reoccurrence. The age at onset of childhood ADEM is usually 3 or 4 years. ADEM before one year of age is very rare. The demyelinating lesions of this case corresponded to the regions which normally become myelinated by 3 months. Although ADEM is usually treated with corticosteroids, high dose gamma 1-globulin therapy can be considered if patients are very young or have a high risk for corticosteroid, or respond poorly to corticosteroids. PMID- 11260922 TI - [Efficacy of lidocaine tapes for a school boy with intractable epilepsy]. PMID- 11260923 TI - [Problem in methylphenidate use for the treatment of attention deficit hyperactivity disorder in Japan]. PMID- 11260924 TI - [Two-step tuberculin skin test in general hospital workers--comparison with nursing home workers]. AB - In 1998, the Japanese Society for Tuberculosis recommended two-step tuberculin skin test (TST) for medical workers. As a large majority of the Japanese were BCG vaccinated in their childhood, it is difficult to distinguish true infection from booster effect. In Japan, it is important to record individual baseline tuberculin reactivity by two-step TST. Two-step TST was performed on 126 general hospital workers and 47 nursing home workers, excluding those whose initial TST was strongly positive, according to the recommendation of the Japanese Society for Tuberculosis in 1998 (strongly positive TST in Japan is defined as the reaction with other intensive responses such as double skin erythema, bleb, lymphangitis, etc.). Diameter of erythema of TST among hospital workers v.s. nursing home workers were as follows. In the initial TST: 23.2 +/- 16.7 mm v.s. 14.2 +/- 10.3 mm (p < 0.001), in the second TST: 26.3 +/- 17.1 mm v.s. 16.7 +/- 9.9 mm (p < 0.02), baseline of TST: 32.0 +/- 18.3 mm v.s. 19.4 +/- 10.7 mm (p < 0.001). Booster effect of TST in hospital workers was +9.8 +/- 15.1 mm, while it was +4.8 +/- 7.8 mm (not significant) in nursing home workers. Among those 30 years and over, these differences were no found, except diameter of erythema in the initial TST. In our hospital with no beds for TB, in the past 10 years, tuberculosis has not been broken out among our hospital workers, while several patients with pulmonary tuberculosis have been diagnosed every year (6 patients in 1999). Thus, some hospital workers might be exposed to infection with tuberculosis from these patients. On the other hands, no tuberculosis patients had been diagnosed in the nursing home, and young nursing home workers very rarely exposed to infection with tuberculosis in their life, and they are similar to the general population. This study suggested that hospital workers were more frequently exposed to tuberculosis infection than other workers. PMID- 11260926 TI - [Problems and their transition of long-term hospitalized patients with tuberculosis]. AB - In 1975, the Tuberculosis Research Committee (Ryoken) conducted its first study on long-term hospitalized TB patients who had been staying in a hospital for more than five years. Similar studies were repeated in 1981 (hospitalized for more than three years), 1988 (hospitalized for more than two years), 1993 and 1998 (hospitalized for more than one year). The same patient cohorts in each study were followed up, each after a different time period, i.e., after 68 months for the 1975 study cohort, 36 months for the 1981 study cohort, 26 months for the 1988 study cohort, and 74 months for the 1993 study cohort. Based on the results of these series of studies, changes in the patients' characteristics and factors related to long-term hospitalization during the last 23 years were analyzed. The main findings are summarized below. 1) The proportion of patients who stayed in a hospital for more than one year in the study decreased from 41.6% in 1975 to 9.5% in 1998. 2) The long-term hospitalized patients have become older, are more likely to be previously untreated cases, and initially bacillary cases. 3) The major cause of the treatment failure among the long-term hospitalized cases as reported by the attending doctors was "too chronic disease" in the earlier years, but the causes have changed to adverse reactions to anti-TB drugs, initial drug resistance, and patient's poor compliance or non-adherence to treatment in recent years. 4) The cause of the long-term hospital stay after bacteriological negative conversion was chronic respiratory failure in more than one third of the patients in every study. Among these patients, an increasing proportion of cases has non medical problems, such as poor family acceptance and reluctance to be discharged. 5) The mortality of the long-term hospitalized cases was generally high. Among bacteriologically positive cases in the study, it was 14% to 19% annually, and tuberculosis deaths occupied 60% to 80% of all the deaths. 6) The outcomes of those patients who were eventually discharged has become less favorable. Only 3% of them were returned to light work in the 2000 study, while 15% did in the 1981 study. PMID- 11260925 TI - [Invasion and intracellular growth of Mycobacterium tuberculosis and Mycobacterium avium complex adapted to intramacrophagic environment within macrophages and type II alveolar epithelial cells]. AB - Profiles of the invasion and intracellular growth of M. tuberculosis (MTB) and M. avium complex (MAC), which had been adapted to intramacrophagic environment, within Mono Mac 6 human macrophages (MM6-M phi s) and A-549 human type II alveolar epithelial cells (A-549 cells) were studied. In this study, we used the organisms grown in MM6-M phi s (intracellularly-adapted: I-type) and those passaged in 7H9 liquid medium (extracellularly-adapted: E-type). First, I-type MTB was less efficient than E-type MTB in invading into MM6-M phi s, while I-type MTB invasion into A-549 cells was greater than of E-type MTB. On the other hand, I-type MAC was more efficient than E-type MAC in entering both into MM6-M phi s and A-549 cells. Second, the ability of MTB and MAC to replicate within MM6-M phi s was increased by intracellular passage of these organisms through MM6-M phi s. In contrast, the ability of these organisms to grow within A-549 cells was decreased to some extent by intramacrophagic passage. These findings suggest that growth within M phi s changes the efficiency of MTB and MAC in invading and replicating in M phi s and type II alveolar epithelial cells. PMID- 11260928 TI - [Prevention of development of pulmonary tuberculosis in compromised host]. AB - Recently compromised hosts have increased due to aging of population, advance of medical technology and therapy or changes in the dietary life and social life. Concomitantly the proportion of compromised hosts in the patients with pulmonary tuberculosis has also increased. Taking up diabetes mellitus, hemodialysis, collagen disease and lung cancer as the representatives of compromised hosts, we studied the propriety of chemoprophylaxis to prevent the development of tuberculosis and the standard for the subjects in the case of chemoprophylaxis being given. Diabetics top the patients in the high risk group of developing pulmonary tuberculosis. Therefore, giving chemoprophylaxis is considered necessary to prevent the development of tuberculosis from diabetics. Chemoprophylaxis to diabetics should be given only when healing of tuberculosis has been found despite the history of treatment for tuberculosis being absent. In the patients of hemodialysis, the total morbidity of tuberculosis is high, but the morbidity of pulmonary tuberculosis is not too high, so chemoprophylaxis for the patients on hemodialysis is not always necessary. However, chemoprophylaxis according to the same standard for diabetics is necessary for the patients with diabetic nephropathy. In the patients with collagen disease except rheumatoid arthritis under consideration for administration of corticosteroid preparations, chemoprophylaxis is considered desirable where doses of more than 10 mg in terms of prednisolone are administered over a long period of time. In the patients with lung cancer under consideration for administration of corticosteroid preparations, chemoprophylaxis is considered desirable where doses of more than 10 mg in terms of prednisolone are administered over a long period of time. PMID- 11260927 TI - [A case of tuberculous pericarditis developing constrictive pericarditis]. AB - A case of constrictive pericarditis which developed after the onset of clinical manifestation of tuberculous pericarditis was reported. A 75-year-old male, complaining of anorexia, was admitted to our hospital. Adenosinedeaminase (ADA) level in pericardial effusion was found to be increased, and the culture of pericardial effusion was positive for tubercle bacilli. Diagnosed as having tuberculous pleuritis and pericarditis, he underwent chemotherapy for tuberculosis. However, massive pleural effusion developed later and pleural effusion drainage was carried out. Despite repeated drainage, pleural effusion continued to recur. Chest CT revealed apparent pericardial thickening, in addition, cardiac catheterization revealed elevation of mean right atrial pressure and marked deterioration of cardiac functions including decrease of cardiac output. These findings were compatible with constrictive pericarditis. After these investigations a diagnosis of constrictive pericarditis was established, and the patient underwent a pericardiectomy. Pathological examination of resected specimens revealed tuberculous inflammation. PMID- 11260929 TI - The human immune response to Mycobacterium Tuberculosis infection and disease. PMID- 11260930 TI - Pharmaceutical reimbursement under the Medicare and Medicaid programs. PMID- 11260931 TI - Imaging ventures: a legal primer. AB - As the technology for imaging continues to improve, we expect the number of non radiologists having some involvement in imaging ventures to increase significantly. Further, the variations and numbers of types of ventures will also likely increase. PMID- 11260932 TI - The U.S. pharmaceutical industry: why major growth in times of cost containment? AB - Growth in utilization rather than price, particularly since 1994, has been the primary driver of increased pharmaceutical spending. In this paper I focus on four factors that have increased utilization, even as cost containment efforts have flourished: (1) "the importance of being unimportant"; (2) increased third party prescription drug coverage; (3) the introduction of successful new products; and (4) aggressive technology transfer and marketing efforts by pharmaceutical firms. I also consider the roles that these four factors are likely to play in the future. PMID- 11260933 TI - Prescription drug prices: why do some pay more than others do? AB - The fact that sick elderly people without prescription drug coverage pay far more for drugs than do people with private health insurance has created a call for state and federal governments to take action. Antitrust cases have been launched, state price control legislation has been enacted, and proposals for expansion of Medicare have been offered in response to price and spending levels for prescription drugs. This paper offers an analysis aimed at understanding pricing patterns of brand-name prescription drugs. I focus on the basic economic forces that enable differential pricing of products to exist and show how features of the prescription drug market promote such phenomena. The analysis directs policy attention toward how purchasing practices can be changed to better represent groups that pay the most and are most disadvantaged. PMID- 11260934 TI - The controversy of increased spending for antidepressants. PMID- 11260935 TI - Paying for graduate medical education: the debate goes on. AB - The debate over Medicare payments for graduate medical education has been conducted under the premise that such payments cover the added costs of training. Standard economic theory suggests that residents bear the costs of their training, implying that the additional costs of teaching hospitals are not attributable to training per se but to some combination of a different patient care product, unmeasured case-mix differences, and the costs of clinical research. As a result, payment for the additional patient care costs at teaching hospitals should come from the Medicare trust fund; any subsidies for training should come from general revenues. PMID- 11260936 TI - Economists on academic medicine: elephants in a porcelain shop? PMID- 11260937 TI - Does economic theory justify changing policy that works? PMID- 11260938 TI - Another alternative for financing graduate medical education. PMID- 11260939 TI - Do consumers know how their health plan works? AB - Expanding consumer choice of plans is beneficial only to the extent that consumers make informed choices. Using data from the 1996-97 Community Tracking Study (CTS), this study compares consumers' responses on four key attributes of their health plan with information provided directly by the plan. Plan attributes relate to choice of providers and access to specialists. Although the accuracy of reporting some individual attributes was fairly high, fewer than one-third of consumers accurately reported all four health plan attributes. In general, consumers tended to overreport plan restrictions, especially the need for approval to see specialists. PMID- 11260940 TI - Patients and profits: the relationship between HMO financial performance and quality of care. AB - This paper matches health plans' financial performance with information on quality ratings as measured by 1997 Health Plan Employer Data and Information Set (HEDIS) 3.0 data. We address three policy questions: (1) Is the quality of care delivered by a plan influenced by the plan's financial performance? (2) Do for profit plans behave differently than nonprofits do? (3) What other factors are associated with variation in plan performance? We find, first, that more profitable plans achieve higher quality scores in subsequent years. Profits may enable a plan to pursue higher quality of care and invest in better management systems. Second, there is little systematic evidence that for-profit plans have different HEDIS scores than not-for-profits have. PMID- 11260941 TI - Provider organizations at risk: a profile of major risk-bearing intermediaries, 1999. AB - Provider organizations have evolved to function as intermediaries between managed care plans and individual providers. These organizations assume much financial risk and care management responsibilities. We profile the characteristics of these organizations in markets across the country. The data, taken from a 1999 telephone survey of sixty-four entities in twenty markets and from interviews conducted during site visits to four markets, highlight the youth of many of these organizations, the large financial risk and functional responsibilities they bear, and the mixed views they hold about the health plans they contract with in terms of their willingness to delegate the authority, support, and collaboration that accompany risk. Policymakers need to evaluate what this means for oversight of managed care. PMID- 11260942 TI - Patients' attitudes toward cost control bonuses for managed care physicians. AB - Physicians' cost containment incentives may create conflicts of interest. To understand how patients view these incentives, we interviewed 1,050 patients regarding a 10 percent cost control bonus and a combined cost control/quality bonus. Seventy-three percent said that the cost control bonus was a bad idea; 49 percent viewed the combined bonus more favorably than the cost control bonus; and 91 percent favored disclosure of bonuses. We conclude that patients find bonuses worrisome and favor their disclosure. A quality component reassures some, but not all, patients. Initiating a dialogue with patients about practicing medicine in an era of limited resources may help health plans and physicians to address patients' concerns. PMID- 11260943 TI - Three decades of health care use by the elderly, 1965-1998. AB - Over the past three decades health spending and hospital use increased more for the elderly than for persons under age sixty-five. Medicare spending for the oldest old (age eighty-five and older) increased faster than for persons ages sixty-five to seventy-four, but that increase was due entirely to greater postacute care use. Health care trends are consistent with the idea that Medicare has improved the health of the elderly. Greater spending increases for the elderly may reflect legislative developments such as the passage of Medicare and its continued fee-for-service nature and the failure to pass universal coverage, as well as changes in the health care delivery system such as the rapid growth in managed care enrollment among persons under age sixty-five. PMID- 11260944 TI - Health spending growth up in 1999; faster growth expected in the future. PMID- 11260945 TI - Moving to Medicare: trends in the health insurance status of near-elderly workers, 1987-1996. PMID- 11260946 TI - Trends in mental health services use and spending, 1987-1996. PMID- 11260947 TI - Trends in avoidable hospitalizations, 1980-1998. PMID- 11260948 TI - Hospital finance: signs of 'pushback' amid resurgent cost pressures. PMID- 11260949 TI - Medical safety: from stories to policy. PMID- 11260950 TI - No one needs to know. PMID- 11260951 TI - Casey's legacy. PMID- 11260952 TI - A question of quality. PMID- 11260953 TI - Medicare physician payment changes: impact on physicians and beneficiaries. AB - The Balanced Budget Act (BBA) of 1997 generally reduced Medicare payments for surgical services while increasing them for other services. Concern about implications of these fee reductions prompted the Medicare Payment Advisory Commission to sponsor a national survey of physicians to learn their views on Medicare payment and whether access to care has changed for Medicare beneficiaries. Results suggest that beneficiaries' access to care has not declined. While physicians are concerned about Medicare reimbursement, they are more concerned about reimbursement from managed care plans and Medicaid. Continued monitoring will be important to detect any emerging access problems accompanying upcoming payment reductions. PMID- 11260954 TI - Health plan characteristics and consumers' assessments of quality. AB - Many purchasers and consumers of health care have become concerned about the quality of care being delivered in managed care plans. Little is known, however, about the health plan characteristics that are associated with better performance. We used survey responses from 82,583 Medicare beneficiaries from 182 health plans to study the association of consumers' assessments of care with health plan characteristics. For-profit and nationally affiliated health plans received much worse scores on the outcomes of interest, particularly for overall ratings of the health plan and composite measures of customer service and access to care. Health plans accredited by the National Committee for Quality Assurance did not receive higher scores. PMID- 11260955 TI - Patient safety: grantmakers join the effort to reduce medical errors. PMID- 11260956 TI - The British NHS and U.S. managed care. PMID- 11260957 TI - Americans' views on health policy: a fifty-year historical perspective. AB - A review of data from more than 100 public opinion surveys conducted over a fifty year period finds that the American public has conflicting views about the nation's health policy. They report much dissatisfaction with the health care system and with private health insurance and managed care companies, and they indicate general support of a national health plan. However, most Americans remain satisfied with their current medical arrangements, do not trust the federal government to do what is right, and do not favor a single-payer type of national health plan. The review also finds that confidence in the leaders of medicine has declined but that most Americans maintain trust in the honesty and ethical standards of individual physicians. PMID- 11260958 TI - Trends in out-of-pocket spending by insured American workers, 1990-1997. AB - This paper examines trends in out-of-pocket spending for insured workers from 1990 to 1997. Data are from the Consumer Expenditure Survey conducted by the U.S. Bureau of Labor Statistics. The survey collects detailed quarterly data on all consumer spending from logs kept each year by more than 10,000 households with job-based health insurance. During the study period, total out-of-pocket spending in constant dollars remained unchanged. Spending for medical expenses, drugs, and supplies declined by 23 percent, but this decline was offset by rising employee contributions for health insurance premiums. The shift to managed care, whose benefit structure requires less cost sharing, may have played a role in reducing out-of-pocket spending. PMID- 11260959 TI - Trend data on medical encounters: tracking a moving target. AB - The National Health Care Survey (NHCS), conducted by the National Center for Health Statistics, consists of separate data collection activities that can be used to track the number and content of health care encounters in the United States. Tracking even something as simple as the number of encounters, however, is complicated by the fact that the content of these encounters changes over time. Results from the NHCS indicate that the U.S. population has been receiving more drugs, more cardiac procedures, more ambulatory surgery, more therapies in nursing homes, and more home health care over time. Policymakers and researchers who examine health care trends should be wary about judging whether the number of length of encounters is positive or negative without also examining the content of these encounters. PMID- 11260960 TI - U.S. health care: taking the long view. PMID- 11260961 TI - Growing differences between Medicare beneficiaries with and without drug coverage. AB - Using data from the 1998 Medicare Current Beneficiary Survey (MCBS), we examine changes in beneficiaries' prescription drug coverage from 1997 to 1998 and compare drug use and spending data for beneficiaries with and without drug coverage. The data show that in 1998 the aggregate prescription drug coverage rate of Medicare beneficiaries may have reached a plateau. Also, prescription drug use declined for beneficiaries without drug coverage and increased for those with drug coverage. Covered beneficiaries also paid a larger percentage of their total drug costs out of pocket in 1998 than in 1997. The result was a widening of use and spending differences between beneficiaries with and without coverage. PMID- 11260962 TI - Dynamics in drug coverage of Medicare beneficiaries: finders, losers, switchers. AB - Although the vast majority of elderly Americans have the stability of basic Medicare benefits, Medicare alone offers no protection from the vicissitudes of the market for outpatient prescription drugs. This paper analyzes the sources and stability of prescription coverage maintained by Medicare beneficiaries in 1995 and 1996. The results show that fewer than half of all beneficiaries had continuous drug coverage over this period, while nearly a third gained, lost, or had spells without coverage. PMID- 11260963 TI - The concentration of health care expenditures, revisited. AB - In two previous publications, we described the distribution of health care expenditures among the civilian, noninstitutionalized U.S. population, specifically in terms of the share of aggregate expenditures accounted for by the top spenders in the distribution. Our focus revealed considerably skewed distribution, with a relatively small proportion of the population accounting for a large share of expenditures. In this paper we update our previous tabulations (last computed using data more than a decade old) with new data from the 1996 Medical Expenditure Panel Survey (MEPS). Our findings show that the skewed concentration of health care expenditures has remained very stable; 5 percent of the population accounts for the majority of health expenditures. PMID- 11260964 TI - [Therapeutic prescription of heroin to drug addicts]. AB - Heroin therapeutic prescription to opiate drug addicts through a study of the literature on this topic is valued. Some doubts are put forward with the regard to the scientific quality of the studies, disposable nowdays, on the basis of the experiences of United Kingdom and Switzerland. We are expecting longer and finer studies to answer the question whether this kind of intervention is really useful. PMID- 11260966 TI - [Diagnosis and classification of primary Sjogren syndrome. Comparison of 3 criteria sets in 219 cases]. AB - In spite of all recent years' international meetings, the question of diagnostic criteria of primary Sjogren's syndrome (pSS) is still under debate. The aim of our study is to define sensitivity, specificity and diagnostic accuracy of 3 sets of criteria: those of the European Community Study Group (ECSG), those proposed by Fox, and those proposed by Daniels. We considered 219 subjects complaining of dry mouth and/or dry eyes and/or parotid swelling, evaluated for pSS. The following parameters were considered golden standard for the diagnosis of pSS: focus score > or = 2 foci/mm2, double positivity for SSA and SSB antibodies, and a sialographic grade > or = 2. Our study demonstrates that ECSG criteria show a high sensitivity and a good specificity, resulting in a diagnostic accuracy similar, and sometimes higher, than that obtained with Fox and Daniels' criteria. PMID- 11260965 TI - [Infectious complications in liver transplant in Italy: current status and prospectives]. AB - The impact of infections in orthotopic liver transplantation (OLT) is remarkable. Studies have shown that about 60% of patient may develop at least 1 infectious episode during the first 3 months after transplant. Within the frame of a Finalized Research Project of the Italian Ministry of Health, during the year 2000 a group of investigators belonging to the major Italian Liver Transplant Centers (LTC)--18 out of 20 Centers--met three times in Genoa with the aim of constituting a Research Group aimed at improving our knowledge of infectious complications in liver transplant recipients (PITF = Program of Infections in Liver Transplantation). The group first collected information about anti infective procedure in LTC. The study shows that no Center is supported by a Intensive Care Unit (ICU) exclusively dedicated to the LTC, although 37% of them have a partially dedicated Unit. Surveillance cultures are routinely performed and are frequently used to address the choice of the antibacterial and antifungal regimes. Selective Bowel Decontamination is also very common. The management of CMV infection is usually performed as indicated in international guidelines. PMID- 11260967 TI - [Importation dengue]. AB - We describe two cases of dengue fever (DF) serologicaly confirmed. In both, the clinical features are characterized by: fever, severe headache, myalgias and arthalgias, transient macule-papule rash, leukopenia and thrombocytopenia. The entire illness last few days and terminates abruptly without therapy. A history of travel to dengue-endemic areas and occurrence of other cases in a community are important reminders to include this disease in the differential diagnosis. The hemoagglutination inhibition test for DF at the Laboratory of Virology of the Istituto Superiore di Sanita on two collected sera, during the acute and convalescent phases, has showed a seroconversion. A problem is to advise patients to avoid endemic areas because the second exposure could induce DHF/dengue shock syndrome. PMID- 11260968 TI - [Spontaneous perirenal hemorrhage in patients with polyarteritis nodosa]. AB - Polyarteritis nodosa is a necrotising vasculitis of small and medium-sized arteries. All coats of these vessels are involved easily resulting in aneurysm formations. It is an immunologic disease owing to immune complexes deposits. Kidney affection is over 80%. Polyarteritis nodosa may be complicated by perirenal hematoma, hemobilia, intrahepatic hemorrhage, epidural hematoma. Spontaneous perirenal hematoma can be the initial manifestation of this disease. Abdominal ultrasonography can provide a quick diagnosis of complicating disease and embolization by interventional radiology may be considered a therapeutic alternative to surgery. PMID- 11260969 TI - [Adenocarcinoma of the rectum: role of preoperative radiotherapy for the preservation of the sphincter function]. AB - Surgery is the common upfront treatment for patients who present with clinically resectable rectal cancer. Post-operative radiotherapy and/or chemotherapy are usual for resected rectal cancer in the United States, but in some European countries radiotherapy is preferred as preoperative modality. Preoperative radiotherapy, alone or combined with chemotherapy, increases the chances of tumor downstaging and downsizing and it is less toxic than postoperative combined therapy. More than 10 randomized trials of preoperative radiotherapy for resectable cancers have been recently published: eight trials have reported a significant decrease in local recurrence and two trials have found a significant improvement in survival. If the primary end-point of most clinical trials included survival, a further pivotal end-point included presentation of function of the sphincter in the attempt to improve quality of life. Approximately 70-80 percent of patients irradiated preoperatively are now able to undergo sphincter preserving surgery. Current studies, focused on the interplay between biological properties of rectal cancer and radiation-induced response will clarify the actual role of preoperative radiotherapy and help select patients who may benefit from combined association of chemotherapy with radiotherapy. PMID- 11260970 TI - [Blood pressure self-monitoring]. AB - The blood pressure is measured, usually, in the last years by the patient at home. The method is useful for the screening of the true hypertensive patients, for the follow-up of "white coat" hypertension, for the control of antihypertensive therapy, and for the clinical trials. The home blood pressure is closer to real blood pressure of a subject and it is better correlated than clinical pressure to target organ damage. The normal values, 135/85 mmHg, are been established by meta-analysis studies. Automatic oscillometric devices should be used. The method, very useful in clinical practice, has to improve in accuracy and measures validation. Doctor has to spend more time to train the patient to use the devices correctly. PMID- 11260972 TI - [The physician's duty]. PMID- 11260971 TI - [Mobbing: a problem in occupational health]. AB - Occupational stress may be the cause of psychosomatic and mental disorders. A development of conflicts into a negative direction may trigger a mobbing behaviour at workplace and a mobbing syndrome on the victims. On account of the frequency of mobbing activities at work, it is to assert the need of preventive interventions. PMID- 11260973 TI - [Cryoglobulinemia and hepatitis C virus infection]. AB - The demonstration of the high prevalence of HCV infection (HCV) in patients with MC has changed the clinico-biologic scenario of MC, supporting its subdivision into two groups: MC HCV- and MC HCV+. The former, which is predominantly a polyclonal cryoglobulinemia, should be regarded as an epiphenomenon of the immune system activation in the course of a variety of chronic infections or autoimmune disorders; the latter, which is a oligo- or monoclonal cryoglobulinemia, referred in the past as "essential mixed cryoglobulinemia", might be expression of an indolent B cell proliferation stimulated by HCV in an antigen-driven mechanism. The association of HCV infection with MC may have a pathogenetic an therapeutic significance. There are a number of reports demonstrating the beneficial effects of alpha-interferon (alpha-IFN) in about a half of patients with chronic HCV and MC. However, after the end of alpha-IFN therapy a recurrence of viremia and cryoglobulinemia is frequently observed and less than 25% of treated patients achieve long term remissions. To improve the sustained response rate, prolonged courses of alpha-IFN monotherapy or a combination of alpha-IFN and ribavirin should be considered. New agents with specific antiviral activity against HCV will probably further improve therapeutic options. PMID- 11260974 TI - [Euthanasia. Ten questions for Giovanni Berlinguer]. PMID- 11260975 TI - [Articular involvement in the course of primary hypogammaglobulinemia]. AB - The Authors describe the main features of the most common forms of primary hypogammaglobulinaemia (PH) focusing on the articular involvement. Patients with Bruton's agammaglobulinemia (BA) and common variable immune deficiency (CVID) are predisposed to develop septic arthritis (including arthritis due to atypical microorganisms such as mycoplasma), arthralgia and symmetrical (usually non erosive) polyarthritis. In BA and CVID complicated by recurrent infections, amyloidosis, which may be itself a cause of arthropathy, can occur. In addition, patients with CVID and selective IgA deficiency show an increased prevalence of juvenile rheumatoid arthritis, systemic lupus erythematosus, Sjogren's syndrome and primary biliary cirrhosis, while patients with selective IgA deficiency are prone to developing seronegative spondylarthropathies, including ankylosing spondylitis. The mainstay of treatment for BA and CVID is replacement therapy with human immunoglobulins. Septic arthritis should be promptly treated with antibiotics, whereas other types of arthritis usually respond well to non steroidal antiinflammatory medications. In contrast, the second line agents commonly used to treat rheumatoid arthritis do not appear to be beneficial in patients with PH-associated arthritis. PMID- 11260976 TI - [Replacement therapy in adrenopause. Dehydroepiandrosterone and aging]. AB - Dehydroepiandrosterone (DHEA) and its sulfated ester DHEAS are the most abundant circulating adrenal steroids but their function remains to be elucidated. DHEAS secretion and serum levels decrease with age and this is parallel to the development of a number of the problems of ageing (immunosenescence, increased incidence of osteoporosis, atherosclerosis and cancer, decrease of cognitive functions and/or well-being). For these reasons a growing interest in replacement of DHEA in elderly people has been developed. The findings from recent studies of replacement of DHEA in elderly are reviewed here. Although we have some positive data about benefit results of this therapy on muscle, bone and well-being, at present it is premature to recommend the routine use of DHEA replacement because most of its aspects remains controversial. PMID- 11260977 TI - [Venous thromboembolism: update and prospectives in therapy]. PMID- 11260978 TI - [Strategies for a greater supply of organs for transplantation]. AB - The number of recipients waiting for a solid organ transplantation has increased greatly in the past 5 years. The supply of donor organs during this period has not kept pace, resulting in a large shortage of suitable organs. In an effort to overcome the disparity between supply of donor and demands, various strategies have emerged to expand the existing donor selection criteria. Kidneys from very old donors can be used successfully when a pre-transplant biopsy shows a modest degree of glomerular injury. Kidneys from donors of 50 years or more, or with a history of hypertension and diabetes, or other evidence of renal disease, currently not accepted for single' kidney transplantation, provide excellent function when transplanted together. In contrast, one liver can be enough for two patients. Split-liver transplantation, i.e. sharing one liver between an adult and a pediatric recipient, is becoming routine procedure and has the potential for meeting the need for liver replacement among children without interfering with adult waiting list. Splitting the liver for transplantation in two adults is a further step forward in the more efficient use of hepatic grafts from cadaver donors. Adult-to-adult living donor liver transplantation can further alleviate the pressure in the waiting list, but the risk for the donor must not be underestimated. The decrease in the number of heart-lung transplants in favour of more single and double lung transplants has also made more hearts and more lungs available. It is difficult to quantify the impact of all these procedures on the shortage of organ donors, but the waiting list should be cut by at least one third for kidney and may be more for liver and lung transplants. PMID- 11260979 TI - Fit residents at lower cost. Limited reimbursement for P/OT (physical/occupational therapy) evokes creative answers. PMID- 11260980 TI - When is an ALF (assisted living facility) more than just an ALF? PMID- 11260981 TI - How do you bait the hook? Creative strategies for employee retention. PMID- 11260982 TI - Bringing LTC up to Internet speed. PMID- 11260983 TI - Dot calm: the ease of e-procurement. PMID- 11260984 TI - 10 tips for buying online. Scout out bargains while avoiding the pitfalls. PMID- 11260985 TI - Today's CCRC dilemma. Balancing the cost and quality of care. PMID- 11260986 TI - Alterra COO is now president. PMID- 11260987 TI - Experience with oseltamivir in the control of a nursing home influenza B outbreak. AB - Oseltamivir prophylaxis was very effective in protecting nursing home residents from ILI and in halting this outbreak of influenza B. A portion of the total ILI cases may have been due to influenza A, as this strain was isolated in one resident. The 10% attack rate in this facility, controlled with oseltamivir, compares favourably with another influenza B outbreak in a similar facility in the same region, over the same time frame (ILI onset 27 December to 17 January). Oseltamivir prophylaxis was not used to manage this second outbreak of laboratory confirmed influenza B. Of the 236 residents, 45 developed ILI for an overall attack rate of 19%, nearly double the rate in the oseltamivir-controlled setting (10%). While oseltamivir was effective in controlling influenza B in this outbreak, further experience and evaluation is required before it can be routinely recommended for prophylaxis of influenza in nursing home outbreaks. Although earlier attempts by others using oseltamivir in the control of influenza A outbreaks have also met with success, it is not yet licensed for this purpose. Compared to amantadine, oseltamivir has a relatively high cost for the control of influenza A outbreaks and this may continue to limit its wider acceptance. The cost-effectiveness of oseltamivir in the control of influenza B outbreaks needs to be specifically addressed given the typically milder nature of influenza B strains. However, such a distinction is not clinically reliable and elderly residents of long-term care facilities remain vulnerable to serious complications associated with influenza infection in general. An alternate agent for influenza chemoprophylaxis that is effective against both influenza A and B, is easily administered and has few side effects, could greatly enhance current prevention and control measures and warrants serious assessment. The spread of this outbreak from the geographically separate ward to other areas of the facility in which residents had not received prophylaxis, underscores the likely role of staff as a vehicle for transmission during facility outbreaks. While accurate staff ILI rates could not be determined, their immunization rates were low, and many staff were ill during the outbreak. Isolation of residents with ILI and prophylaxis of non-ill residents on the initial outbreak wards was insufficient to prevent the spread of the outbreak, although it was subsequently halted once prophylaxis was extended to all residents. In view of the uncertainty over this medication's widespread use, in the absence of licensure or previous studies demonstrating its effectiveness in the prophylaxis and control of influenza B outbreaks, initiation of oseltamivir prophylaxis was staggered by ward. In a declared influenza A outbreak, the protocol in a long term care facility is to initiate amantadine prophylaxis on all residents, rather than ward-by-ward. While anti-viral prophylaxis may be an effective secondary control measure in the management of influenza outbreaks, optimal primary prevention would be more effective. This would require increased vaccine coverage of residents and particularly of staff, who play an important role in the importation and transmission of influenza within these facilities. PMID- 11260988 TI - Respiratory virus surveillance FluWatch project update. PMID- 11260989 TI - Me, my torch and my mirror. PMID- 11260990 TI - Teenage pregnancies. 2: Comparative outcomes. PMID- 11260991 TI - The onset of labour. 1: A review of the physiology. PMID- 11260992 TI - Occipito-posterior positioning and some ideas about how to change it! PMID- 11260993 TI - How does it feel to you? Uterine palpation and lochial loss as guides to postnatal 'recovery'. I--The background. PMID- 11260994 TI - The influence of antenatal classes on pain relief in labour. 2: The research. PMID- 11260996 TI - The National Sentinel Caesarean Section Audit. PMID- 11260995 TI - Folic acid fortification. Proposed UK recommendations. PMID- 11260997 TI - Life, love and birth in the Philippines. PMID- 11260998 TI - Video time! PMID- 11260999 TI - Midwives need to make a difference. PMID- 11261000 TI - Pleased to meet you. Georgina Lessing-Turner. PMID- 11261001 TI - Mrs Silver rides again. PMID- 11261003 TI - Research remembered. AB - Another good question to ask yourself and indeed others, is on Jeannette's lips. Of course nursing is not exactly well-endowed with research findings- perioperative practice even less so--but it's the 'change in practice' that must be the key phase for evaluation. So, read on and then reach for the proposal form to the Education and Research Fellowship Fund (ERFF) at NATN, or similar funding body, to join the search for evidence of effective practice. Become one of the 'remembered' in due course. PMID- 11261002 TI - Buy it right, use it right, keep it right! PMID- 11261004 TI - Change and progress.... Life as they see it. The Education Committee of NATN write on issues that concern or interest them. PMID- 11261005 TI - Decontamination and the law. A better understanding. AB - A manufacturer is usually liable in law for a defective product; however, processors are also viewed as manufacturers. Theatre staff who sterilise and label products are also liable; a liability which exists for ten years. Pat Hewitson discusses the implications. PMID- 11261006 TI - A train of events? AB - What sort of relationship do you have with your Sterile Services Department? Do you know what goes on in that department? Do their staff know what goes on in your department? In making a strong case for recognised training in Sterile Services, Olivia and Frank Waller see an exchange of experiences between operating theatres and SSD as a valuable part of the learning process. Too often separate departments only contact each other when things go wrong. The authors quote a damning judgement from an enquiry following the death of five patients: a powerful reminder of the fact that perioperative care gives only one chance to get it right. Just because you work in theatre does not mean you can ignore or take for granted those departments on which you rely for supplies and services. Get involved, find out and understand what they can do for you and what you can do for them. PMID- 11261007 TI - Improving staff communication. The role of the cardiac theatre liaison nurse. PMID- 11261008 TI - Using the Internet to enhance evidence-based practice. AB - Evidence-based healthcare requires reliable and validated research findings to aid improved clinical practice. The Internet, with its growing medical resources, is an invaluable tool for busy healthcare workers. However, as more material becomes available on the web, the quality and reliability of various information sites is a worry for many practitioners. The ability to evaluate web sites is becoming increasingly important and as a novice user, I was interested to discover the extent to which the Internet could assist me researching a specific topic. This article describes the process of conducting an Internet search, on a topic relevant to theatre nursing, and considers how the resources might best be evaluated. PMID- 11261009 TI - Laser safety management. AB - LASER is an acronym for Light Amplification by Stimulated Emission of Radiation. Since the first working laser was demonstrated in 1960 the laser has evolved from being viewed as a weapon, courtesy of the film industry, to its current position as a commonplace medical device within the healthcare industry. As perioperative staff we have become very familiar with the therapeutic use of this device. It is my experience however that, just occasionally, we are guilty of the old adage 'familiarity breeds contempt'. We must remember that the very same features which make lasers so useful in healthcare may also represent major health hazards to patients, staff and others. PMID- 11261010 TI - Controlling the processes. Legislation governing decontamination and sterilisation. AB - Time was, in the not so distant past, that any piece of equipment we wanted to re use in the Theatre Sterile Services Unit (TSSU) was either autoclaved or 'sterilised' in chemicals. The words 'single use' were virtually never used, and I certainly knew very little about the legislation surrounding the whole issue. Indeed, just a little while further back than that, we used to wash and pack and autoclave whole sets of instruments ourselves (it was a good way of learning the sets for the student nurses!). Now we have the whole issue of traceability and responsibility to consider. Martin Williams discusses some of the legislation and issues arising. I wonder who my Authorised Person is? Do you know yours? PMID- 11261011 TI - Survey of nurses' assessment of cancer-related fatigue. AB - Fatigue is now widely recognised as a significant' problem for patients with cancer. While research effort into this subject has grown considerably in recent years, the exact mechanisms underlying fatigue remain unclear. Therefore assessing and managing this symptom can be problematic. This paper describes the findings from a survey evaluating how nurses (n = 84) in a Cancer Centre in Edinburgh currently define and assess fatigue. The results demonstrate that while the problems associated with fatigue are acknowledged, assessment tools are not widely used and the majority of nurses report that they would benefit from further education on the subject to assist in the care of patients. PMID- 11261012 TI - Forum for Applied Cancer Education and Training. Communication with hidden communities. PMID- 11261013 TI - The emphasis of care must be on enabling people to carry on with their lives despite their cancer. PMID- 11261014 TI - Attitudes, beliefs and behaviour regarding the use of sunbeds amongst healthcare workers in Bradford. AB - Although cosmetic tanning and unprotected solar exposure are common, little is known about general attitudes, beliefs and behaviour regarding the use of sunbeds. We sought to determine the frequency of sunbeds use in a select sample and to assess the knowledge and beliefs regarding this behaviour. A self administered anonymous questionnaire was distributed to a sample of 648 employees work for Bradford Hospitals NHS Trust. The questionnaire explored demographic information (including hair and skin type, family history with skin cancer), frequency of sunbeds use, knowledge about the risks of UV exposure and motivations for practising this behaviour. Four hundred and eighteen women and 52 men completed the questionnaire, making a response rate of 73%. Nearly half of respondents (207; 44%) reported using sunbeds to some extent; of those 12% reported frequent use. Appearance ('to look better') was the most popular reason given by respondents for using sunbeds, followed by 'feel healthy'. Frequency of using sunbeds was found to be negatively correlated with the age of respondents and the existence of family experience with skin cancer, and strongly associated with the opinion that it is safer to use a sunbed than subathing outdoors, the female sex and smoking. It is clear from this study that the psychological factors that influence sunbeds use are complex and that so far public education campaigns have had little impact on it. This study highlights some of these psychological factors. PMID- 11261015 TI - Education and training of healthcare staff: the barriers to its success. AB - On-going developments in Britain's healthcare services are placing great demands on its workforce. If high quality care is to be maintained, education and training is needed to provide staff with the necessary knowledge and skills to adapt to their changing roles. A recent research project looking at the education and training needs of non-specialist staff caring for people with cancer identified a number of barriers that prevent education and training from meeting its full potential. These barriers include time, accessibility, financial issues, staff motivation and marketing and advertising and are discussed in this paper together with some possible solutions to overcome them. It is concluded that the planning and development of education and training must address these barriers if it is to be successful in increasing the knowledge and skills of staff, and thereby improve the quality of patient care. PMID- 11261016 TI - Greek parents' reactions, difficulties and resources in childhood leukaemia at the time of diagnosis. AB - Open-ended interviews were used to examine parental psychological reactions, difficulties and resources during the period following the diagnosis of childhood leukaemia. Data were obtained from 71 randomly selected mothers and fathers of children diagnosed with leukaemia at least 3 months prior to the study. The content analysis revealed a wide diversity of parental responses including many of the defensive mechanisms described in the literature such as shock, denial, anxiety and guilt. The most difficult factors for the parents to deal with during the initial period were the psychological upset and the financial burden. Problems associated with relating to others and to the health care system were also identified. Hope, social support and the marital relationship were the most helpful resources in managing the multifaceted problems caused by the diagnosis. Forty-five per cent of the participants felt that the quality of their marital relationship was improved, whereas fewer reported that the diagnosis seriously disturbed their marriage. Spouses were found to adopt symmetrical rather than complementary ways of responding to and coping with the event. Nurses have a key role in assessing the individual parent, the marital unit and the entire family system and planning appropriate interventions. PMID- 11261017 TI - Consumer participation in the development of psychosocial clinical practice guidelines: opinions of women with breast cancer. AB - Clinical practice guidelines are playing an increasingly important role in defining quality care and consumers have a considerable interest in participating in the development of guidelines. The objective of this study was to explore consumer's perceptions of guideline items relating to psychosocial care of women with breast cancer, developed by Australia's National Health and Medical Research Council National Breast Cancer Centre. Women diagnosed with breast cancer in the previous 2 years (n = 313) received a letter about the study via their radiation oncologist. Consenting women were contacted by the researchers to complete a telephone survey. The survey asked women to rate the importance of draft guidelines items, including discussing prognosis, providing information and choice, doctor-patient communication, preparation for surgery, providing emotional support, providing social support, dealing with practical and cultural issues and continuity of care. One hundred and forty women (45%) completed the survey. The results indicated that at least 50% of respondents rated 28 of the 52 items as 'essential' components, with respondents identifying providing information and choice, and doctor-patient communication as the most important aspects of psychosocial care. The findings suggest the guidelines adequately reflect consumer opinions and identify priority areas for clinicians to address in providing psychosocial support to women with breast cancer. PMID- 11261018 TI - Evaluation research. PMID- 11261019 TI - Real time training for mental health staff working in an in-patient setting. AB - Concerns around acute in-patient mental health care including staff training are becoming increasingly prominent (Sainsbury, 1998; Sainsbury 1997). A training solution is proposed in this paper, which aims to deliver content derived from evidence based practice actually within the practitioner's own clinical workplace during their working hours. The benefits of this approach are discussed together with examples of content and methods for evaluating learning outcomes. PMID- 11261020 TI - Psychiatric nursing education: can it make a difference? AB - A review of the literature clearly indicates that psychiatric nursing is not a popular career choice for undergraduate nursing students. While it appears from the literature that exposure to the theory and practice of psychiatric nursing can influence the attitudes of student nurses towards the mentally ill, no clear picture emerges from the research. Furthermore, the design of previous research does not address the relationship between improved attitudes towards the mentally ill and an increase in the popularity of psychiatric nursing. This paper discusses the results of a quasi-experimental study, which compared the attitudes of student nurses towards psychiatric nursing as a career option prior to and following completion of the psychiatric nursing component of the course. The post test results suggest a strong and statistically significant increase in the popularity of psychiatric nursing. The possible impact of problem based learning in influencing these results is discussed. PMID- 11261021 TI - Describing a model of nursing as a focus for psychiatric nursing care. AB - This thesis would not be complete in it's review of Psychiatric nursing and Psychiatric nurse education, without reference to one of the fundamental changes that is occurring in nursing. One of the major changes that had occurred in recent years has been in the focus of nursing management nurses are placing much more emphasis on nursing diagnosis and nursing theories and are detaching from the more traditional medical model when planning nursing care. This movement towards a greater autonomy in planning their nursing care is assisting nurses in becoming a professional and independent group. Nurses are establishing a body of knowledge and skills which may be called "nursing science". Through the use of nursing models for planning patient and health care, nurses are offering a better service to the individual and the community. These changes in the focus of care commenced in the 1950's within the United States with the introduction of nursing models and theories. The Philosophies and concepts of these theories have outlined various methods and ways that the nurse could better plan and organize their professional work, including patient assessment and care. As early as 1953 the term "nursing diagnosis" was introduced by Fray to describe a step in the organization of a nursing care plan. (1) The various nursing models and theories seemed to have coincided with the movement through the 1950's and 1960's within America towards more generic based integrated curricula. PMID- 11261023 TI - Numeracy must become a priority for nurses. PMID- 11261022 TI - Nursing profession suffers from lack of status. PMID- 11261024 TI - Case 31: managing a caseload. Health visitor who failed to manage her caseload appropriately. PMID- 11261025 TI - Bladder washouts in the management of long-term catheters. AB - It has been estimated that 40-50% of patients with long-term catheters can suffer with catheter blockage. This not only causes distress to the patient but also increases the demands on community nurses' time and resources. Infection with bacteria such as Proteus causes the urine to become alkaline. Crystalline deposits can then form inside the catheter lumen which causes blockage. Nurses often manage blocked catheters with the use of bladder 'washouts' or bladder instillations. However, the literature is confused over the terminology of 'bladder washouts', instillations and irrigation and a great deal of controversy surrounds the effectiveness of these procedures. Crisis management of catheters occurs when nurses wait for catheters to become blocked before changing them; this often occurs at inconvenient times and patients frequently have to wait several hours before help is available. To avoid crisis management, nurses should aim to assess individual patients' 'pattern of catheter life' and plan changes accordingly. This would improve patient care and allow nurses to utilize their time more effectively. PMID- 11261026 TI - Antenatal screening for HIV: time to embrace change. AB - In August 1999 the Government set a national objective of reducing the numbers of children acquiring human immunodeficiency virus (HIV) infection from their mothers by 80%. The key policy change towards achieving this objective was that HIV testing was to be recommended as a routine and integral part of antenatal care. The UK has fallen behind other Western industrialized countries in the uptake of antenatal testing and reduction in mother-to-child transmission. This article examines the background to these new recommendations, the substantial benefits of available interventions and the reasons why current testing policies have been failing. PMID- 11261027 TI - Constructive risk in the care of the older adult: a concept analysis. AB - There is much talk within health care at present of the need to manage risk; indeed, it is a cornerstone of the Government's clinical governance policy (Department of Health (DOH), 1998). However, by only concentrating on the dangers associated with risk, the positive effects engendered in the process of taking calculated risk. By analyzing risk as a constructive concept, particularly in the care of the older adult, it is possible to not only identify those attributes that define the phenomenon, but also explore the necessary antecedents and consequences of pursuing this approach to care. Using the work of Walker and Avant (1998), model cases are used to illustrate how the concept may be recognized in practice. PMID- 11261028 TI - Preoperative fluid restrictions: hospital policy and clinical practice. AB - This study examined the practice of preoperative fluid restrictions and the influence of the hospital 'nil by mouth' policy on clinical practice. Structured interviews were used to assess the knowledge of nurses and anaesthetists relating to current hospital policy, their attitudes to fluid fasting and the constituents of clear fluids. The interval between the last intake of fluid and the induction of anaesthesia was measured in 90 adult patients to determine actual periods of fasting. It was found that most patients on the same operating list commenced fasting simultaneously with little or no attempt made to individualize the timing which contributed to prolonged periods without fluids, ranging from 3 hours 30 minutes to 17 hours and 45 minutes. Only 30% of nurses were aware of the hospital policy compared with 75% of anaesthetists. The evidence from this study demonstrated that the hospital policy was not reflected in clinical practice which continued to be based on tradition. PMID- 11261029 TI - Communication is the essence of nursing care. 1: Breaking bad news. AB - This is the first of two articles which support the assumption that effective communication is the foundation upon which nursing should stand. The articles are founded on an examination of an interaction analysis of a role play situation in the classroom where a group of 19 MSc nursing students presented a nursing scenario to their peers and feedback was later generated from the total group. At least seven minority ethnic groups, who were new to the country, were represented. The role play was later examined using theoretical frameworks to guide the analysis. This article provides analysis of the breaking of bad news, giving information through exploring, questioning and reassurance, touch, and, finally, empathy and humanism. The second article will explore and examine ethical issues, such as truth-telling and autonomy, that were also evidenced in the role play. Other themes were personal constraints such as family stress and counselling. This article begins by introducing readers to the role play situation. The number of communication concepts identified in this short 20 minute role play positively support the authors' argument that communication is the essence of nursing care. PMID- 11261030 TI - A clinical evaluation of the Transfoam mattress after 4 years. AB - It is recognized that pressure-reducing foam mattresses can be of benefit in the prevention of pressure sores but it is also recognized that more information is required to demonstrate their long-term efficacy (Cullum et al, 1995). It is therefore necessary to evaluate the pressure-reducing capabilities of this type of mattress some years after purchase to evaluate if in fact it can maintain its initial level of performance. This article presents the results of one such study carried out 4 years after the purchase of Transfoam mattresses. PMID- 11261031 TI - Investigating the use of Aquaform Hydrogel in wound management. AB - This article explores the role of hydrogels in wound management. Methods of debriding necrotic/sloughy wounds are considered and the benefits of using a hydrogel are discussed. The article focuses upon Aquaform Hydrogel, investigating the evidence of its efficacy, and concluding with an evaluation of the product which resulted in a change of practice. PMID- 11261032 TI - Hope: a key concept in the psychology of nursing. PMID- 11261033 TI - Thinking outside the box: strengthening NP certification. PMID- 11261034 TI - Should a sick NP be sent home? PMID- 11261035 TI - Certification for practicing NPs. PMID- 11261036 TI - Asthma in the elderly. A diagnostic and management challenge. PMID- 11261037 TI - Clinical management of the geriatric stroke patient. PMID- 11261038 TI - Amenorrhea in adolescents. Sorting out the clinical picture. PMID- 11261039 TI - AIDS in the elderly. Aging raises unique treatment concerns. PMID- 11261040 TI - Is there really a 'male menopause'? Whatever the term, midlife changes warrant attention. PMID- 11261041 TI - Eye problems in the elderly. PMID- 11261042 TI - Differential diagnosis of lichen simplex chronicus. Meeting the challenge. PMID- 11261043 TI - An ideal combination. PMID- 11261044 TI - Nursing in the developing world: lessons for the future. PMID- 11261045 TI - The Ottawa Charter--from nursing theory to practice: insights from the area of alcohol and other drugs. AB - This article aims to assist nursing services to use the Ottawa Charter as a framework for nursing practice. Incorporation of the Ottawa Charter for Health Promotion into a nursing structure constitutes an innovation in nursing practice that was evaluated as a quality improvement exercise in a health-care organization responsible for providing services in the area of alcohol and other drugs. The evaluation consisted of two stages and sought to identify the degree to which the framework was effective in practice. This involved identifying issues surrounding the implementation of the Ottawa Charter as a framework for nursing practice as well as identifying the means by which quality improvements could occur. The evaluation involved an initial questionnaire to all nursing staff, followed by a series of focus groups. The data collected was both informative and enlightening and revealed a range of pertinent issues such as staff understanding and interpretation of the Ottawa Charter, expansion of the nurse's role and suggestions for organizational change. The Ottawa Charter strategies are discussed in relation to their relevance to the organization under evaluation and also expanded into recommendations to assist those contemplating using the Ottawa Charter as a framework for nursing practice. There was considerable agreement among the respondents that the Ottawa Charter provided a useful framework for nursing practice but was on occasions problematic. PMID- 11261046 TI - Working community as the basis of the quality of care. AB - Nursing takes place in a working community and the working community influences the quality of nursing. The purpose of this study was to find out the effects of nurses' individual characteristics, working conditions, and the ward sisters' management skills on working climate and team spirit. Thirty-seven wards of two Finnish hospitals took part in the study, and 717 nurses answered the questionnaire. Statistical analyses indicate that the ward sister's management skill is the best predictor of the climate and team spirit of the wards as perceived by nurses. The nurses' age, education, and working experience have no direct effect on the working community. Nurses who work by the day, who have plenty of further education and permanent employment, are most likely to be dissatisfied with the climate. According to the results, it is important that the ward sister provides sufficient opportunities for professional development, especially for those who are highly motivated to advance their careers. The ward sister can influence the ward's team spirit also by furthering operational preconditions of the ward. PMID- 11261047 TI - Counselling groups for spouses of elderly demented patients: a qualitative evaluation study. AB - Ten women and eight men who were caring for their demented husbands or wives participated in a closed-group counselling programme, developed and carried out by two district nurses at a local health centre. There were seven or eight participants in each group, which met 13-14 times over a period of 8 months. This study is based on semistructured interviews about the participants' situation just before entering the counselling group, the counselling programme itself, and their situation after the end of the programme. Their situations before the programmes were described as an exhausting, chaotic prison but after the programme they could cope with their situation and plan and manage their daily life. None of the participants needed further organised counselling; engagement in the local dementia association was sufficient for them. The counselling nurses' experience in and about caring for demented patients, their tactful authority, the closed groups and the long duration of the programme were judged to be crucial for the successful outcome of the programme. PMID- 11261048 TI - Reflections on the role of the nursing development facilitator in clinical supervision and reflective practice. AB - The notions of clinical supervision and reflective practice are attracting considerable interest across the spectrum of nursing. This paper focuses upon how the two were linked together and, with the support of an independent survey, were used to evaluate the nursing development facilitator role. It is argued that reflective practice, as a model of clinical supervision, may provide a forum for nurses to explore the value of their actions. Linked to this, the personal knowledge that is revealed may mirror reality and as a result it is argued that there is benefit in evaluating practice through reflection. PMID- 11261049 TI - Feeding outcomes and influences within the neonatal unit. AB - This study examined the feeding intention of mothers (n = 100), and the factors and beliefs, and changes in those factors or beliefs that influenced their choices, in the challenging environment of the neonatal unit. Mothers' experience and the frequency of nurse-assisted feeding activities were examined in mothers intending and not intending to breast-feed on discharge. Eighty-one per cent of mothers were either partially or fully breast-feeding or intending to do so on discharge. The most important factors identified as influencing this feeding choice included personal choice, with other influences being special benefits, more natural and feeling closer to the baby. Experiences such as infants receiving their first sucking feed from either the breast or bottle (inclusive of breast milk) and mothers expressing breast milk more frequently, were found to be significantly different and increased in frequency, in mothers intending to breast-feed. Differences in the mean weighted total daily nurse-assisted feeding score confirmed that these activities varied with gestational age (< or = 32 weeks [2.57], 32 to less than 35 weeks [3.86], and > or = 35 weeks [4.91]; F = 7.04, d.f. 55, P = 0.002), although there was insufficient power to determine differences between breast-feeding and non-breast-feeding mothers. The use of the Feeding Activities Calendar may have contributed to increased activity and high levels of breast-feeding in this preterm group. PMID- 11261050 TI - 'The day of the soft towel?': comparison of the current bed-bathing method with the soft towel bed-bathing method. AB - The impressions of 200 patients (both medical and surgical) and 200 nursing staff (registered, enrolled and trainee enrolled nurses) in relation to two bed-bathing methods were compared by means of questionnaires and semi-structured interviews. Data regarding costs were obtained from appropriate cost centre managers. The results of the study found the soft towel bed-bathing method to be more cost effective and provide more patient and nurse satisfaction than the current bed bathing method. PMID- 11261051 TI - From rituals to reason: creating an environment that allows nurses to nurse. AB - From rituals to reason is a description of the journey that one unit is taking to overturn traditional nursing culture and create an environment that will enable nurses to care for people. In our experience the reality of day-to-day nursing practice does not always correspond to the nursing rhetoric of individualised holistic nursing care for each person. We identified that traditional nursing culture, with a focus on task orientation, rigid hierarchical structures and resultant disempowerment of staff, is an impediment to delivery of patient centered care. We introduced changes to management structures, care delivery models and staff behaviours with the aim of overturning this traditional nursing culture. Changing clinical practice to focus on client outcomes and provide a relaxed and patient-centered environment was easier to achieve than staff empowerment and their full involvement in decision making and innovation. PMID- 11261053 TI - Staff ratios in intensive care: are they adequate? PMID- 11261052 TI - All people with MS have a right to be treated. PMID- 11261054 TI - Case 30: returning to the register. Nurses who seek to regain their professional status. PMID- 11261055 TI - Leg ulcer after care: the role of compression hosiery. AB - The management of venous leg ulcers is an important issue for many healthcare professionals. However, the care and management does not stop when the ulcer has healed. Prevention of recurrence and lifelong compression is an important issue that is often neglected. This article reviews the anatomy and physiology of the venous system, investigates the causes of venous leg ulcers and describes the principles of graduated compression. Compression hosiery and its role in the treatment of varicose veins is also looked at as well as deep vein thrombosis and preventing recurrence of the venous leg ulcer. PMID- 11261056 TI - Quality of life and leg ulcers: will NHS reform address patient need? AB - The present Government has made much of its commitment to gaining patients' views on the health services they receive. Qualitative studies of patients with leg ulcers have highlighted the fact that patients feel healthcare professionals do not always empathize with their plight and sometimes appear to lack the skills to help them. Quality of life in chronic wound care has, thus far, eluded definition but strenuous efforts are being made to quantify the impact that issues such as pain, isolation and frustration can have on leg ulcer patients. If healthcare professionals are able to demonstrate empathy with patients they may be able to maximize the cooperation needed for the management of ulcers. By reflecting on gaps in their knowledge and becoming more assertive in their demands for appropriate training, nurses can improve the outlook for patients' quality of life. PMID- 11261057 TI - Use of psychotropic medication in people with a learning disability. AB - The use of psychotropic medication for people with a learning disability is a controversial issue that has received much attention. This article explores some of the issues for learning disability nurses surrounding the use of psychotropic medication. There are concerns regarding the side-effects that antipsychotic medication can produce. Evidence suggests that some healthcare professionals, including learning disability nurses, need to keep themselves regularly updated on issues surrounding the use of these drugs such as efficacy, side-effects and interactions. Learning disability nurses need a clear understanding of the reasons behind the prescription of such powerful medication especially when it is used in the management of challenging behaviour. There are indications that learning disability nurses would support alternative approaches to medication such as the use of behavioural interventions. More healthcare professionals, direct carers and clients should be encouraged to become part of the multidisciplinary drug-review process. PMID- 11261058 TI - Autism in adulthood: the concepts of identity and difference. AB - It was not until 1980 that autistic spectrum disorders were officially recognized as disorders of development and not of psychoses (Wing, 1996). (In this article the term 'autism' will be used to refer to all individuals diagnosed across the autistic spectrum, including people with Asperger syndrome.) There will be many adults with autism aged over 20 years of age who are not appropriately diagnosed and who are not receiving relevant care (Trevarthen et al, 1998). Many will have been the victims of a medical model of treatment as opposed to receiving the necessary education and support. This article, the second of a two-part series (the first article (Vol 9(12): 779-84) dealt with children with autism) will explore how contemporary knowledge can help healthcare professionals to support adults with autism in the development of positive self-identities. PMID- 11261059 TI - Conservative treatment of erectile dysfunction. 3: Literature review. AB - This is the third of a three-part article addressing whether physiotherapy involving pelvic floor muscle exercises (PFMEs) is efficacious as a first-line treatment for erectile dysfunction (ED). The first part (Vol 9(11): 691-4) highlighted the prevalence of ED, associated risk factors, the anatomy of the penis and the physiology of erection. The second part (Vol 9(12): 755-62) concentrated on the published clinical trials investigating the treatment and prevention of ED. This part will critically analyse the literature. PFMEs using ischiocavernosus muscles (ICMs) and bulbocavernosus muscles (BCMs) seem to have merit as a treatment for ED due to mild or moderate venous leakage. Men suffering from ED due to other causes may also benefit. There is no strong evidence that electrical stimulation or electroacupuncture is effective or ineffective. No studies demonstrating preventive conservative treatments were found. There is evidence that the ICMs and BCMs increase penile rigidity in the tumescent penis, that pelvic floor muscle efficiency is higher in potent men than impotent men and that perineal muscle efficiency reduces with age in impotent men. There is limited evidence that pelvic floor exercises relieve ED due to venous leakage and are a realistic alternative to surgery. Randomized controlled trials are needed to explore the use of PFMEs as a first-line treatment for men with ED. PMID- 11261060 TI - Pharmacological interventions in type 2 diabetes: the role of nurses. AB - Type 2 diabetes is complex and difficult to treat, and patients often present with hyperlipidaemia and hypertension. We now have compelling evidence that adequate treatment of hyperglycaemia, together with tight blood pressure control, can reduce the risk of developing long-term complications of diabetes. This article explores the syndrome of type 2 diabetes, including some of the causal mechanisms. The various forms of treatment are explored: diet, exercise, different oral hypoglycaemic agents and insulin therapy. Finally, the importance of the nurse in assessing the most suitable therapeutic intervention for people with type 2 diabetes is outlined. PMID- 11261061 TI - Reducing the spread of tuberculosis in the homeless population. AB - Tuberculosis (TB) is an old infectious disease that has re-emerged in recent years and is responsible for many deaths throughout the world. Homeless people residing in shelters and hostels within inner city areas of the UK and the USA are at risk from this serious disease. Interventions to control the spread of TB are described in the literature researched; these include the introduction of inducements to encourage participation in screening programmes and the recommendation of directly observed therapy. The literature reflects the partial success of these programmes in the UK and USA. Targeting homeless persons most at risk is challenging as is gaining accurate information on those who are affected by TB. Effective coordination of care by healthcare providers in hospital and the community is imperative. It appears that healthcare professionals are becoming more prescriptive in their approach which is relinquishing the homeless population from taking responsibility for their own health care. PMID- 11261062 TI - The relative influence of personal and workplace descriptors on stress. AB - A survey canvassing self-reported workplace stress levels and personal/workplace demographics was conducted among a sample of Australian nurses. Stress ratings and demographics were cross-sectionally analysed. Quantitative and qualitative analyses of survey responses suggest that excessive workload is the likely dominant predictor of workplace stress. Personal and workplace demographics are shown to be relatively unimportant in comparison with workload-related factors. Regular exercise habit was shown to be correlated with reduced workplace stress ratings. The findings support a previously unreported view that nurses are unlikely to bring their personal stress with them to the workplace--rather the workload characteristics of the workplace itself are the principle stress determinants. PMID- 11261063 TI - Accepting the diversity of overseas nurses. PMID- 11261064 TI - Nursing and gerontology. AB - This article, originally prepared as a think piece for The John A. Hartford Foundation, explores the interrelationship between nursing and gerontology: strengths nursing brings to the area of aging; challenges that must be addressed both at the societal level and within the profession for nursing to achieve its full potential in gerontology; and strategies that might be adopted to maximize strengths and address identified gaps. These strategies include high-lighting the heroic behaviors of the gerontologic nurse, increasing support for gerontologic advanced practice nursing, promoting collaborative gerontologic research, encouraging dissemination of nursing's knowledge base, and collaborating with foundations to promote self-care. It is proposed that nursing's research-practice agenda in the third millennium must be: preventing disease where possible; minimizing morbidity and maximizing quality of life when disease cannot be prevented, and having the wisdom to reconcile the two. PMID- 11261065 TI - Enhancing geriatric nursing scholarship: specialization versus generalization. AB - This article explores the relative merits of encouraging preparation of more nurses with specialization in geriatrics as compared to encouraging geriatric preparation among nurses whose major field of study is outside geriatrics. The article explores two approaches to examining capacity for geriatric nursing scholarship among nurse scholars not involved in geriatrics, and in schools of nursing with strength in research but with little geriatric research. The findings show an ongoing need to strengthen geriatric nursing as an area of specialization. Faculty prepared in geriatric nursing are underrepresented in schools of nursing, and only a small number of doctoral students specialize in geriatric nursing. Academic nursing programs with strength in geriatric nursing need ongoing support to maintain and expand current geriatric programs. Data support that encouraging individual non-geriatric nurse faculty and doctoral candidates to focus their work on areas of concern to geriatric nursing, and strengthening geriatrics in research-intensive schools of nursing that have not heavily invested in geriatric scholarship are viable options for strengthening academic geriatric nursing. Establishing mechanisms to attract nurse scholars working outside the scope of geriatric nursing to address clinical issues of concern to older adults offers promise in rapidly attracting new scholars to geriatric nursing. PMID- 11261066 TI - Improving care for the frail elderly: the challenge for nursing. PMID- 11261067 TI - Individualized care: perceptions of certified nurse's aides. AB - Despite substantial attention devoted to the development of individualized care in nursing homes during recent years, empirical research assessing progress is limited. Further, few studies have explored the experiences of certified nurse's aides (CNAs) in this regard. This survey examines the perceptions and experiences of CNAs in providing individualized care. CNAs (n = 254) were asked to describe a number of current practices and obstacles to implementing individualized care in nursing homes. The majority of respondents reported experiencing: flexibility to change daily schedules; supervisor assistance with challenging residents; active participation in care planning; freedom to test new approaches to care; and supervisors who are open to CNA suggestions. Several barriers to individualized care were also described, including: inadequate staffing; poor team communication; staff attitudes; and a lack of knowledge and training in alternative approaches. These findings provide important insights into the supports and obstacles to implementing individualized care in nursing homes from the perspective of CNAs. PMID- 11261068 TI - Preparing for a meeting of The John A. Hartford Foundation. PMID- 11261069 TI - Ode to my mom. PMID- 11261070 TI - Fall risk assessment. PMID- 11261071 TI - Maintaining health in well older adults: initiatives for schools of nursing and The John A. Hartford Foundation for the 21st century. AB - In the past century, there has been a substantial increase in the number of older Americans and an increase in their life expectancy at birth. These trends will continue over the next century, marked by further increases in active life expectancy and marked changes in the racial and ethnic composition of the older population. While nursing has traditionally focused on the care of frail older individuals, most older adults--well older adults--enjoy a relatively high level of health and function. In this article, well older adults are defined as those with the physical, mental, social, and spiritual function or resources to meet the needs of everyday living. Recommendations for improving care of well older adults are provided, including reconceptualizing ways of thinking, expanding practice initiatives, building the science of nursing research in geriatrics and gerontology, developing education and training opportunities, and rethinking individual safety within the context of autonomy. PMID- 11261072 TI - Women survivors of childhood abuse: the impact of traumatic stress on education and work. AB - A disempowering after-effect of childhood abuse that is not well-researched in nursing is the inability of many women abuse survivors to perform successfully in adulthood tasks such as working, managing money, and parenting. This inability often results from lack of family support, cultural impoverishment, limited formal education, and for some, illiteracy. By default, many women survivors engage in criminal, usually dangerous forms of work. A critical/feminist interview study involved 20 urban low income abuse survivors, who were mostly women of color. Participants were recovering cocaine misusers who had suffered multiple forms of childhood maltreatment. This article reports on a secondary analysis of narratives given by survivors, focused on learning and work difficulties. Findings were grouped into five broad domains: (1) school as problematic, (2) lack of adult life skills, (3) problems with academic and health literacy, (4) legitimate and illicit forms of work, and (5) means of help. In vivo quotes support these themes, and policy and practice implications are discussed. PMID- 11261073 TI - Speaking unavoidable truths: understanding early childhood sexual and physical violence among women in prison. AB - Women in prison have often suffered physical and sexual abuse as children which substantively contributes to their substance abuse, violence, and criminal behaviors. To understand women's offending, and subsequent health issues, it is helpful to understand their own victimization. In this participative action research, women used poetry as both a process and a product to confront the truths of their lives in terms of what was done to them and what they have done to others and themselves. As the women learned to speak their own truths, those same truths became unavoidable to themselves. This rise into consciousness is demonstrated through excerpts from the participants' writings. PMID- 11261074 TI - Intimate partner abuse among women diagnosed with depression. AB - Domestic violence is a pervasive problem for women, and depression is the most prevalent negative mental health consequence of domestic violence. This study investigates the extent to which domestic violence is part of the history of women diagnosed with depression. Eighty two women with a diagnosis of depression were surveyed. A 61.0% lifetime prevalence of domestic violence was found. Lifetime prevalence for forced sex was 29.3%. Demographics of abused and nonabused women were not significantly different. Abused women were found to be less healthy. Prevalence of headaches, chronic pain, rape or marital rape, and sleep problems or nightmares were significantly higher. Severity of abuse was significantly correlated (p < .01) to severity of depression. Implications for mental health practice and training of peer support group leaders for women with depression are described, as well as directions for future research. PMID- 11261075 TI - Sisters of the Yam: African American women's healing and self-recovery from intimate male partner violence. AB - In this womanist ethnographic investigation African American women (N = 21) survivors of intimate male partner violence were interviewed about their resilience-recovering experiences. This article foregrounds the role of therapeutic support groups in African American women's healing experience and addresses how race and ethnicity shape the lives and the recovering process for many African American women. The findings are important to practitioners who strive to provide assistance and interventions for African American women as well as other women of color. PMID- 11261076 TI - Encountering violence and aggression in mental health nursing: a phenomenological study of tacit caring knowledge. AB - Violence is a growing psychosocial problem in the health care working environment. Literature shows that nurses are physically assaulted, threatened, and verbally abused more often than other professionals. However, some nurses are able to relate to clients in a way that produces positive resolution. This study explored the phenomenon of positive encounters with aggressive and violent clients. Guided by a phenomenological method, data were analyzed within a lifeworld perspective. The essential meaning of the phenomenon of caregivers' experiences of encountering violent clients is described as an "embodied moment," which is explicated by seven themes of meaning, "respecting one's fear and respecting the client," "touch," "dialogue," "situated knowledge," "stability," "mutual regard," and "pliability." The authors discuss the meaning of the outcome and propose both theory and praxis-oriented activities toward decreasing aggression and violence in health care. PMID- 11261077 TI - Violent and nonviolent girls: contrasting perceptions of anger experiences, school, and relationships. AB - Arrests of American girls for assault and weapons charges are rapidly increasing, at rates exceeding those for boys. Yet research on girls' violence is scant. We surveyed a national sample of 213 girls (ages 9-19) via personal interview or an Internet questionnaire, regarding anger precipitants and behaviors, interpersonal relationships, and experiences of discipline at home and school. Girls were categorized as violent (n = 54) if they had been suspended or expelled from school for fighting or bringing a weapon, or charged with a violent offense by the juvenile justice system. The remaining girls (n = 159) were categorized as nonviolent. The anger of violent girls tended to be intense and generalized, while the anger of nonviolent girls was precipitated by specific situations of injustice. Correlates of feeling angry enough to hit or hurt someone were loneliness, unfair treatment by adults, not liked by classmates, and somatic anger symptoms. Violent girls were significantly more likely to dislike school and perceive school discipline as unfair. Both groups of girls held negative views of television violence and curfews. Although girls with well-established patterns of aggression need psychotherapy, school-based interventions such as emotional literacy and violence prevention programs may also be helpful. Mental health nurses are well prepared to serve in a consultative role to schools, assisting in the development and delivery of violence prevention programming. PMID- 11261078 TI - Forget the fleecing of America! Tom Brokaw, where are you? PMID- 11261079 TI - From digoxin to angiotensin-converting enzyme inhibitors: issues in pharmacotherapy for congestive heart failure. AB - Congestive heart failure (CHF) is a disease with high mortality rates and increasing prevalence in the United States. As our understanding of the pathophysiologic characteristics of this disease has progressed, so has our pharmacologic approach to treatment. Digoxin was the first documented drug used in the treatment of CHF, and since that time, the efficacy of its use has been the source of great controversy. A more recent treatment option is the use of angiotensin-converting enzyme (ACE) inhibitors, the first drug class shown to increase survival rates in CHF. Although controversy remains, ACE inhibitors appear to be gaining favor over digoxin in the pharmacologic approach to treatment. This article provides a pathophysiologic review of CHF as it is understood today, the rationale behind the use of digoxin and ACE inhibitors, and a challenge for the future in the pharmacotherapy of CHF. PMID- 11261080 TI - Complementary healthcare practices and the implications for nurse practitioners. AB - Complementary, or alternative, healthcare practices are being incorporated into approximately 4 out of 10 Americans' daily health practices. The out-of-pocket expense for such healthcare use was estimated at $21.2 billion in 1997. There are many different forms of complementary health care that nurse practitioners (NPs) must be aware of when evaluating and forming plans of care with their patients. NPs must develop and incorporate interview techniques to obtain this information from their patients to prevent potential interactions. NPs must also be aware of their lack of experience with complementary healthcare practices and the legal liability of incorporating these practices into their practice without appropriate preparation in their use. PMID- 11261081 TI - Stroke: a case review. AB - Stroke is indicated by an abrupt manifestation of neurologic deficits secondary to an ischemic or hemorrhagic insult to a region of the brain. Stroke is ranked as the third leading cause of death in the United States, affects more than 730,000 individuals per year, and accounts for 160,000 deaths annually. Approximately $40 billion is spent annually for health care treating stroke victims. This case report shows that despite the use of antithrombotic and/or antiplatelet aggregating drugs, the key to stroke management is primary prevention. PMID- 11261082 TI - Nurses self-performing and teaching others breast self-examination: implications for advanced practice nurses. AB - Although there is evidence to suggest that breast self-examination (BSE) aids in the early detection of breast cancer, the underuse of this prevention behavior continues to exist. The purpose of this study was to explore the personal behaviors and professional practices of nurses in the use of BSE and to discuss implications for the advanced practice nurse (APN). The sample (N = 300) consisted of nurses and student nurses. While almost all nurses had performed BSE at least once, fewer than half did this monthly. The majority believed it was the nurse's role to teach BSE, but almost three fourths of the sample taught it only occasionally or rarely. Not thinking of it and not knowing when or how to teach were reasons identified for not teaching. The roles of the APN as an educator, leader, consultant, direct care provider, and researcher are examined in light of these findings to promote nurses' performing and teaching BSE. PMID- 11261083 TI - Increasing smoking cessation counseling by advanced practice nurses. AB - The magnitude of individual and societal problems caused by tobacco use mandates that all primary care providers identify and advise smokers to quit. However, this topic has received little attention in the nurse practitioner literature. The purpose of this project is to identify effective methods by which advanced practice nurses can increase the identification and counseling of smokers by reviewing research on this topic. The articles for review were obtained through a computerized literature search and a review of related reference lists. The articles were analyzed and categorized into three groups: office-wide interventions to increase provider identification and counseling of smokers, smoking cessation training programs for providers, and studies using the stages of change theory. Provider smoking cessation programs and office-wide reminders increased the identification and counseling of patients who smoke. The stages of change theory helped explain the steps smokers must progress through to cease smoking. Interventions appropriate for various stages in the cessation process are suggested. PMID- 11261084 TI - Living with hurting and difficulty doing: older women with osteoarthritis. AB - Osteoarthritis (OA) is the most common rheumatologic health problem of older adults. Developing a greater understanding of what it is like to live with this chronic, progressive, and frequently unsuccessfully treated condition is necessary to improve evidence-based nursing care to support independent living. Eighteen women aged 65 to 92 years participated in narrative descriptive research based on naturalistic-framework and qualitative-analysis methods. Data were the transcribed narratives of the participants, field notes of observations and impressions, theoretical memos, coded units of the narratives, and categories noted. Deconstructions and reconstructions of the narratives led to the meaning of "Being With OA," with intermediate categories, "Living With Hurting" and "Living With Difficulty Doing." Recommendations included nursing interventions based on individual problems and strengths and further studies of older adults with chronic health problems. PMID- 11261085 TI - Nurse practitioners: essential providers for the 21st century. PMID- 11261086 TI - Nurse practitioner students in Nicaragua. AB - Despite the growing interest in enhancing the cultural awareness for nurse practitioner (NP) students who work with patients from the developing world, there is a dearth of reports on such experiences. This report describes the clinical experiences of NP students from George Mason University (Fairfax, VA) during an intensive 2 weeks in Nicaragua in their final semester, accompanied by an NP faculty member. The program was planned and implemented in collaboration with the Universidad Politecnica de Nicaragua School of Nursing (Managua, Nicaragua). The students' clinical experiences included working in a Health Post and an impoverished community. Students learned to manage clinical problems using minimal resources and acquired an appreciation for the cultural, political, and economic situations from which many of their patients in the United States originate. Recommendations for establishing this type of experience are included. PMID- 11261087 TI - Nurse practitioners and the Internet. AB - This article describes the Internet as an easily accessible, up-to-date, and valuable information system that nurse practitioners must master to keep up with the needs of their patients and today's healthcare system. It contains a comprehensive review of the multiple uses of the Internet that include education, communication, clinical decision making, and research, and can serve as an update to the information in the healthcare and nursing literature. Strategies are suggested to incorporate the Internet into practice. PMID- 11261088 TI - Profiles of editorial board members: from the academy to the community health center. PMID- 11261089 TI - Non-physician practitioners in radiation oncology: advanced practice nurses and physician assistants. PMID- 11261090 TI - Symptom perception and evaluation in childhood asthma. PMID- 11261091 TI - Training for transformation: reorientating primary health care nurses for the provision of mental health care in South Africa. PMID- 11261092 TI - Cross-cultural study of beliefs about smoking among teenaged females. PMID- 11261093 TI - Needlestick injuries: are you at risk? PMID- 11261094 TI - Astrocytomas: the clinical picture. AB - An astrocytoma is one of the most common brain tumors. Approximately 16,500 people will be diagnosed with brain cancer this year in the United States; of these, an estimated 13,000 will die. Astrocytomas originate from astrocytes in the central nervous system, and the maximum average life expectancy following an astrocytoma diagnosis is 18 months. Intracranial tumors often are devastating because of their growth and spread to vital centers, where they alter neurologic functions that make patients "people." Surgery, radiation, biotherapy, and chemotherapy are among the most common regimens used to treat these life threatening intracranial tumors. Nurses caring for these patients play a vital role in providing pertinent interventions, emotional support, and end-of-life care. PMID- 11261095 TI - Temodar offers promise for treating astrocytomas. AB - Temodar is an exciting new drug that shows promise in the treatment of brain tumors as well as in other types of cancers. Research currently is being conducted with Temodar to determine its efficacy in treating various solid tumors, glioblastoma multiform, and even metastatic melanoma. Temodar provides adult patients with anaplastic astrocytoma with another option of therapy that is convenient, produces minimal side effects, and, above all, instills hope. PMID- 11261096 TI - Hypersensitivity reaction to paclitaxel: nursing interventions. AB - Paclitaxel, a mitotic inhibitor, is used to treat a variety of cancers. A significant incidence of paclitaxel-related hypersensitivity reactions (HSRs) occurs because of the diluent used. Premedication with dexamethasone, diphenhydramine, and H2-histamine antagonists has markedly decreased the incidence of HSRs. Paclitaxel-related HSRs should be managed immediately and appropriately by (a) stopping the infusion, (b) administering oxygen, (c) infusing fluids, (d) continuously monitoring blood pressure, pulse, and oxygenation, and (e) initiating standing orders for i.v. corticosteroids and diphenhydramine or other emergency medications. Oncology nurses are key to the rapid recognition and treatment of paclitaxel-related HSRs. PMID- 11261097 TI - Cutaneous reactions associated with alpha interferon therapy. AB - Although flu-like symptoms are the most common side effect associated with alpha interferon therapy, cutaneous reactions also can occur and present a management challenge for oncology nurses. Cutaneous reactions reported in the literature include generalized effects and localized reactions. Alopecia appears to be the most common generalized cutaneous reaction reported, followed by transient and mild generalized rash-like reactions. Localized manifestations include injection site reactions, induration, or necrosis. Armed with knowledge about interferon associated cutaneous reactions, oncology nurses can identify these reactions early and promptly implement appropriate interventions. PMID- 11261098 TI - Principles of cancer prevention and early detection. AB - Oncology nurses are becoming involved more frequently in answering questions regarding cancer prevention and detection. This article includes a discussion of basic underlying principles of cancer prevention and detection, including types of prevention, frequently used epidemiologic terms, validity assessments of cancer screening tests, and strategies for risk assessment. An understanding of basic principles related to cancer prevention and detection is necessary to provide information and patient education that is accurate and beneficial to patients and their families. With this background information, oncology nurses can help develop programs for cancer prevention and control. PMID- 11261099 TI - Antiandrogen therapy and side effects following radical prostatectomy. PMID- 11261100 TI - Adverse drug reactions and reporting. PMID- 11261101 TI - Assessment of febrile neutropenic patients. PMID- 11261102 TI - Diarrhea. PMID- 11261103 TI - Integrating new information and technology in oncology nursing practice. PMID- 11261105 TI - Focusing on quality cancer care. PMID- 11261104 TI - Methotrexate. PMID- 11261106 TI - "Precarious ordering" on the road. PMID- 11261107 TI - Human papillomavirus, genital warts, Pap smears, and cervical cancer: knowledge and beliefs of adolescent and adult women. AB - The high prevalence of genital warts, human papillomavirus (HPV), and the virus's cancer-causing potential warrant that women be well informed about these conditions and measures to prevent them. The purpose of this descriptive study was to examine women's knowledge and beliefs about genital warts, HPV, cervical cancer, and Pap tests. We interviewed 40 women recruited from health clinics in Chicago (20 adults) and Indianapolis (20 adolescents) about these issues. Audiotapes of the interviews were transcribed and analyzed. Among both the adults and adolescents there was a good deal of misunderstanding about symptoms associated with genital warts, about the purpose of Pap smears, and about the association of genital HPV with abnormal Pap smears and cervical cancer. The gaps in women's understanding about this potentially deadly infection suggest the need for more comprehensive education about preventing genital HPV, the infection's possible sequelae, and the significance of Pap screening for cancer detection and prevention. PMID- 11261108 TI - Attitudes of adolescent/young adult women toward human papillomavirus vaccination and clinical trials. AB - Human papillomavirus (HPV) is the most common sexually transmitted infection. It often inflicts adolescents and young adults shortly after onset of sexual activity. More than 30 types of HPV infect the anogenital area; some HPV types cause cervical cancer in women decades after infection, whereas other types cause genital warts in both men and women within a year after infection. Vaccines are being developed against oncogenic and wart-producing HPV. Knowledge of HPV and attitudes toward HPV vaccination/clinical trial participation among 60 female adolescents and young adults were evaluated. Knowledge of HPV in this group was limited, but almost all participants would be interested in receiving vaccines that prevented cervical cancer and genital warts. Only 30% were likely to participate in an HPV clinical trial that required shots and pelvic examinations. A key motivating factor for clinical trial participation was the potential for a vaccine to help other women. PMID- 11261109 TI - Repatterning care: women's proactive management of family caregiving demands. AB - Caregiving frequently is framed as a family issue when, in practice, women bear the primary responsibility. The goal of this feminist grounded theory study of women's caring was to explain the complex process of family caring by women within the current context of health and social reform. One finding of this study was that women use a process of repatterning, that is, reorganizing caring activities, to reduce or overcome the negative effects of caring demands. These findings make visible both the losses and gains of caring women as they learn and employ the expert strategies of anticipating, setting ground rules, juggling time, and relinquishing and replenishing. These findings draw attention to the complexity of caring and provide direction for health professionals in their work with individuals and families and in lobbying for necessary resources. PMID- 11261110 TI - Gender equity in health care: the case of Swedish diabetes care. AB - To explore the issue of gender equity in diabetes care in Sweden and to develop strategies for monitoring gender equity in health care, population-based studies and statistics published since 1990 were reviewed that contained gender-specific data on health care utilization, glycemic control, patient satisfaction, health related quality of life, and mortality from diabetes. The review shows that diabetic women in Sweden report more frequent outpatient contacts, less patient satisfaction, and a lower health-related quality of life than diabetic men. No gender differences were found in the level of glycemic control. Young and middle aged men with diabetes have a high excess all-cause mortality as compared with nondiabetic men. A trend toward stronger social gradient in mortality among women than men with diabetes was observed in a large nationwide study. PMID- 11261111 TI - A theory of life's lesions: a contribution to solving the mystery of why women get sick more than men. AB - In recent decades there has been an explosion of scholarship dealing with all aspects of women's existence, including gender differences. Whatever one's theoretical position regarding gender differences there is ample empirical evidence to show that rates of morbidity differ between men and women in contemporary society. A paradox exists that although women may live longer they tend to experience non-life-threatening conditions more than men do. What lies behind differential morbidity among women and men remains puzzling. In this paper the author attempts to address this puzzle by proposing the concept of life's lesions. It is proposed that women more than men experience life's lesion because they are caught in situations of unresolved contradictions and moral quandaries more than men. A case history from Mexico illustrates the theoretical proposition. PMID- 11261112 TI - Chronic fatigue syndrome: a woman's dilemma. AB - Chronic Fatigue Syndrome (CFS) is an illness characterized by fatigue with varying levels of disability. According to the Centers for Disease Control (CDC) there are 2 to 5 million people in the United States who suffer from CFS and a disproportionate number are women. There are many theories of etiology of the condition and controversy has surrounded recommendations for diagnosis and treatment. CFS can mimic other diseases and women are doubly affected since many have comorbid conditions. While diagnoses and treatment are critical to the health of women, having the disease and coping with the symptoms may have a greater impact on their well-being and quality of life. The authors report qualitative data describing the experience of having CFS (N = 22) and quantitative responses of 42 CFS sufferers reporting psychosocial factors. The psychosocial factors were measured by the Derogatis Stress Profile (DSP), Spielberger Trait-Anger Scale, Ways of Coping Survey, Profile of Moods States (POMS) Survey, and the Perceived Stress Scale. The findings indicate that CFS changes the lives of women who suffer with the disease and disrupts their relationships, careers, and perceptions of themselves. PMID- 11261113 TI - Follow-up care of cancer survivors. PMID- 11261114 TI - Differential diagnosis of cough: focus on lung malignancy. AB - Evaluating cough in the primary care setting can be very difficult and requires a thorough look through a long list of potential differential diagnoses. The most worrisome diagnosis is that of a lung malignancy. Primary care providers must assess each patient carefully in a logical, precise manner to determine a working diagnosis for acute versus chronic cough in smokers and nonsmokers. Early detection leads to a diagnosis of lung cancer at earlier stages and may offer the only possibility of cure. This article provides primary care providers with an overview of the most common causes of cough, an algorithm to assist with the diagnosis, and a brief overview of the staging, diagnostic workup, treatment, and management of lung cancer. PMID- 11261115 TI - Management of lymphedema. AB - Most providers of health care to persons with a history of cancer will encounter lymphedema requiring an intervention. A basic understanding of the lymphatic system as well as clinical and diagnostic tests will aid the primary care practitioner in determining the correct diagnosis. Knowledge of treatment options and available resources will increase success in limb reduction. Ongoing monitoring, review of lymphedema precautions, and advocacy are the supportive elements needed to help patients cope with this cancer survival issue. PMID- 11261116 TI - Oncologic emergencies. AB - Oncologic emergencies may occur in patients who have no prior diagnosis of malignancy as well as in patients who are known to have cancer. It is important for the primary care practitioner to consider an oncologic cause for symptoms or problems that may bring a patient into the office. These symptoms often are vague and could be indicative of numerous problems frequently associated with individuals who have cancer. A brief overview of the most common oncologic emergencies is given, along with differential diagnostic possibilities and management strategies. PMID- 11261118 TI - Psychosocial needs of patients with cancer in the primary care setting. AB - The diagnosis of cancer exacts an emotional toll on patients and their family members, necessitating attention to psychosocial factors by primary care providers. This article describes the usual course of emotional reactions to cancer, high-risk times for emotional vulnerability, and the role of the primary care provider in assessment, treatment, and referral. Additionally, the unique challenges of cancer survivorship are described, along with resources for patients seeking additional information about medical and psychosocial issues. PMID- 11261117 TI - Chemoprevention research: implications for primary care practice. AB - The field of cancer prevention research is entering a time of growth and opportunity. This important research is identifying agents that are making a substantial difference by reducing cancer incidence in high-risk populations. Primary care providers are natural partners for this research because of their diversity, commitment to disease prevention, and long-term access to their patient population. Several national chemoprevention trials in breast and prostate cancer are open and seeking to affiliate with primary care providers. Information is provided on this research effort, the development of chemoprevention trials, and how to learn more. PMID- 11261119 TI - Young woman presenting with a strong family history of breast cancer. PMID- 11261120 TI - Breast cancer prevention with tamoxifen. PMID- 11261122 TI - Advance directives. PMID- 11261121 TI - Cancer screening and prevention guidelines. PMID- 11261123 TI - Radiation therapy and chemotherapy: what to expect. PMID- 11261124 TI - Cancer resources. PMID- 11261125 TI - Care given to patients should be based on their needs (SOU, 1996). PMID- 11261126 TI - The use of antipyretic medications in the prevention of febrile convulsions in children. AB - Febrile convulsions are a relatively common outcome in paediatric febrile illness, although it is not known why some children suffer these. Antipyretic medications may form the basis for some treatment regimens, although they are not recommended in published guidelines. There is little evidence that the prophylactic use of antipyretics has any effect in reducing the incidence of febrile convulsions. Consequently, educational interventions aimed at reducing parental fear and helping them to care for their children during febrile illnesses may be more efficacious. PMID- 11261127 TI - Communicating with people with stroke and aphasia: understanding through sensation without words. AB - To illuminate the phenomena of 'communicating with people with stroke and aphasia without words', 10 care providers particularly successful at communicating with stroke and aphasia patients who were working at a stroke rehabilitation ward narrated their experiences of communicating with such patients. A phenomenological hermeneutic approach, inspired by Ricoeur's philosophy, was used in the analysis. Two main themes were found: facilitating openness and being in wordless communication. The care providers sensed the feelings of the patients and experienced similar feelings themselves, thus, the communication is guided by the shared feelings between the care provider and the patient, i.e. communion. For this 'communication through sensation' to take place, the following factors were found to be necessary: creative closeness in combination with protective distance; striving for satisfaction and against exhaustion and desperation; meeting the patient halfway to gain understanding; exhibiting attention and accessibility to the patient; and trust and confidence for both care providers and patients. The findings were interpreted and discussed in the light of works by Levinas, Logstrup and Stern. PMID- 11261128 TI - The nursing care of stroke patients in nursing homes. Nurses' descriptions and experiences relating to cognition and mood. AB - Registered nurses working in nursing homes often care for stroke patients with impaired cognition and mood disorders. Understanding the behaviour of these patients often puts great demands on nurses. This study illuminates registered nurses' descriptions and experiences of stroke patients and the nursing care given in nursing homes, with a focus on cognition and mood. Registered nurses responsible for the care of stroke patients in nursing homes were asked to describe the individual patient's state of health and the nursing care given. Patients' cognition and mood have been selected for this article. A qualitative content analysis was used to group the text into categories. Registered nurses' descriptions showed great complexity and variation in patients' disabilities, as well as uncertainty about understanding these patients and the appropriate nursing care. Registered nurses described the need for further education in stroke care, and adequate resources for patient activity training, as well as meeting patients' psychosocial and communicative needs. PMID- 11261129 TI - 'You do know he's had a stroke, don't you?' Preparation for family care-giving- the neglected dimension. AB - Countries throughout the developed world have introduced a policy of community care for older people to reduce costs to the state and maintain quality of life. In reality community care is largely family care and recognition of the need to support family carers is being promoted through the notion of partnership with professional carers. Such a partnership calls for a more complete understanding of how carers' needs change over time and how professional support can be most effective. Support is particularly important at the start of care-giving in order that carers can exercise free choice and be adequately prepared for their role. This paper provides an overview an ongoing longitudinal study and reports specifically on the findings of data from a preliminary study in which a convenience sample of seven experienced carers of stroke survivors who attended a stroke and carers club were interviewed in their own homes. Based on initial data from a longitudinal study of stroke victims, this paper outlines four themes: 'What's it all about', 'Going it alone', 'Up to the job' and 'What about me?' These themes highlight the difficulties carers experience in the immediate aftermath of stroke. PMID- 11261130 TI - Investigating recovery from stroke: a qualitative study. AB - A recent randomized controlled trial evaluated the effects of specialist nurses providing information, advice and support to caregivers and patients at home during the first year after a stroke. Reported here are the results of a complementary study which used qualitative methods to examine the experience of patients and caregivers during the year of recovery after a stroke. We used semi structured interviews with a purposively selected sample of 30 patients and 15 caregivers at the end of a randomized controlled trial (13-16 months post stroke). Patients and caregivers provided vivid descriptions of the recovery process. Recovery was perceived in terms of the degree of congruence patients identified between their lives before, and after, stroke. Patients therefore had individual and personal yardsticks for measuring their recovery. In conclusion, further research and interventions must consider the diverse, complex, dynamic and highly personal character of stroke recovery. Traditional outcome measures are too simplistic to capture patients' and caregivers' experiences. There do not appear to be single or simple solutions to the problems of facilitating psycho social adjustment. PMID- 11261131 TI - Older people and laxative use: literature review and pilot study report. AB - This study explored older adults' perceptions of constipation, and the measures taken if they believed themselves to be afflicted by this condition. The paper provides an overview of the current literature surrounding laxative use, followed by a discussion of the pilot study and its findings. The objectives of the pilot study were to establish older people's definitions of the term 'constipation'; identify prescribed laxatives, over-the-counter laxatives, and home remedies used by older people to manage constipation; produce a detailed account of when these products are used; identify the older person's belief system underpinning their concepts of constipation, and their consequent use of laxative products; and produce information which will inform nursing practice, with a particular focus on nurses in community practice. People who identified themselves as being constipated were interviewed on a one to one basis. Participants shared their stories of loneliness, social isolation and anxiety related to constipation and the need to use laxatives on a daily basis, and described persistent unpleasant and often painful physical symptoms such as bloating, urges, excessive flatus, nausea and cramps, commonly associated with laxative ingestion. Nurses are challenged to work with older people within a 'wellness' framework, helping clients to maintain their bowel function, rather than fall back on short-term options, which provide only brief relief of symptoms, while ignoring the underlying causes. PMID- 11261132 TI - A pilot study to examine the relationships of dyspnoea, physical activity and fatigue in patients with chronic obstructive pulmonary disease. AB - A descriptive-correlational design was used to examine the relationships between dyspnoea, physical activity, and fatigue in patients with chronic obstructive pulmonary disease (COPD). Lazarus and Folkman's theory of stress, appraisal, and coping provided a framework to guide the study. Dyspnoea was measured by a vertical visual analogue scale, fatigue by the Fatigue subscale of the Profile of Mood States, and physical activity by the six-minute walk (6 MW) test and an open ended question. A convenience sample of seven male and 15 female patients with COPD provided data for analysis. The sample was characterized by relatively high forced expiratory volume in one second (FEV1) indicating mild lung impairment and high mean levels of fatigue and dyspnoea. No significant gender difference was found in the ratings of dyspnoea and fatigue and the 6 MW distance. Dyspnoea, physical activities, and fatigue were all significantly inter-related (P < 0.001). Results indicated that the higher the dyspnoea scores, the shorter the 6 MW distance walked, and the higher the fatigue scores. Limitations and suggestions for nursing practice and future research are presented. PMID- 11261133 TI - Men making sense of their chest pain--niggles, doubts and denials. AB - Participant observation was undertaken of the early admission period of 25 men admitted to hospital with acute chest pain, followed by in-depth interviews of 10 of the men after discharge. Grounded theory methods were used in the analysis to develop a model of how the men came to interpret their experiences. An emerging feature of the men's experiences was that, although they had suffered intense pain prior to admission, there had been a series of delays whilst they tried to rationalize their symptoms. We relate our discussion to literature on men and masculinity and the notion of Foucault (1975) of self-surveillance, to offer an insight into the men's self concept and social situation. Our conclusions suggest that men's self concept as 'healthy' may inhibit a speedy response to the signs and symptoms of acute coronary occlusion, increasing the risk of cardiac arrest without nearby life support. PMID- 11261134 TI - Patient participation in bedside reporting on surgical wards. AB - Increasingly nowadays, patients have an opportunity to take part in nurses' reporting sessions via bedside reporting. The aim of this study was to compare nurses' and patients' opinions of the purpose of bedside reports, patient participation in bedside reporting sessions, and factors that promote or prevent their participation. Data were collected by a questionnaire survey of nurses (N = 118) and patients (N = 74). A response rate of 81% was achieved in both groups. Additionally, 76 bedside reporting sessions were observed. According to patients, the main reasons for not participating were tiredness, difficulties in formulating questions, lack of encouragement, difficulties with the language used, nurses concentrating more on their papers than on them, and the reporting sessions were too short. Nurses reported that patients took a more active part in reporting sessions than patients themselves thought. The average time spent on each patient's report was three minutes. PMID- 11261135 TI - Compliance of adolescents with chronic disease. AB - The purpose of this paper is to describe the factors that affect compliance in adolescents with a chronic illness and to compare compliance and factors connected to compliance between adolescents with asthma, epilepsy, rheumatoid arthritis (JRA) and insulin dependent diabetes mellitus (IDDM). The data were collected by questionnaire. Altogether 1200 individuals were selected from the Finnish Social Insurance Institution's register. The response percentage was 88 (n = 1061). One-fifth (23%) of adolescents with chronic disease felt that they had complied fully with health regimens, while 60% placed themselves in the category of satisfactory compliance and the remaining 17% reported poor compliance. In each patient group compliance was promoted by good motivation, a strong sense of normality, a positive attitude towards the disease and treatment, energy and will-power, experience of results, support from the parents, nurses and physicians, and a feeling that the disease was not a threat to social well being. PMID- 11261136 TI - Self-care in adults with asthma: how they cope. AB - The purpose of this study was to find out how well adult asthma patients in Finland cope with self-care in three areas of asthma treatment. The areas of physical, psychological and social asthma treatment were examined. Associations between demographic background data and self-care were also studied. Data (n = 130) for the study were collected using a questionnaire specially developed for this study. A deductive perspective was employed in data analysis. Respondents showed fairly good competence in self-care in all three areas of asthma treatment. However, up to 30% of the asthma patients had pets and 16% were smokers. Extra stress was reduced by exercise and positive thinking. Humour was also important in helping most of the respondents cope mentally. Social support played a significant part in fighting the sense of powerlessness which is caused by asthma. According to the results, women coped better than men in the social area of self-care. PMID- 11261139 TI - Information point: Mann-Whitney test. PMID- 11261138 TI - Patients' views of a new nurse-led continence service. AB - This study used qualitative methods to assess patients' views of a new nurse-led continence service that was being evaluated in a randomized trial as part of the Leicestershire Medical Research Council (MRC) Incontinence Study. The service was provided by a team of five nurses who had received a 3-month training programme on the assessment procedures and the evidence-based practice protocols. In-depth qualitative interviews were carried out by four trained interviewers with 23 respondents, seven male & 16 female (mean age 58 years), and were analysed using NUD*IST software. The main themes to emerge were related to the interpersonal skills and technical skills of the nurse and how these impacted on the effectiveness of treatment. An informal, friendly approach by nurses with good communication skills relieved patients' embarrassment and anxiety, giving them confidence and trust in the nurses, thus facilitating information exchange and effectiveness of care. Good communication skills conveyed the nurses' specialist technical skills and knowledge, encouraging patient compliance with treatments. PMID- 11261140 TI - Information point: Wilcoxon signed rank test. PMID- 11261137 TI - Development, implementation and evaluation of a new nurse-led continence service: a pilot study. AB - The Leicestershire Medical Research Council (MRC) Incontinence Study is a series of interrelated studies exploring the epidemiology of urinary symptoms, including incontinence, and evaluating service provision and treatment options for these symptoms. This paper describes one aspect of the Leicestershire MRC Incontinence Study, namely the development, implementation and evaluation of a new nurse-led continence service. When developing a new service it is important to determine its acceptability and suitability to the target population. The new mode of service delivery was dependent on specially trained Continence Nurse Practitioners (CNP) delivering predefined evidence-based treatment interventions. Objective and subjective outcome measures were used to evaluate the service. The service was shown to be effective in reducing urinary symptoms and led to high levels of patient satisfaction. This service is currently being evaluated in a randomized controlled trial. PMID- 11261141 TI - The effect of occupational socialization on nurses' patient handling practices. AB - Back injury is widely considered an occupational hazard of nursing work. Manual handling of patients has been implicated in the development of back injury in nursing. Legislation has been in place in the UK since 1992 that should have addressed factors implicated in the development of back injury, such as manual handling of patients. Nurses' patient handling practices have been slow to change in line with this legislation. This can be explained in part by a lack of training and resources required for change. However, nurses' attitudes and beliefs about patient handling, and the culture into which new nurses are socialized, may play a significant role in hindering changes in patient handling practice. PMID- 11261142 TI - Discharge planning: an exploratory study. AB - The desire to reduce the length of waiting lists in the modern health service means that strategies for decreasing the length of hospital stay are exercising the minds of service planners. This has led to renewed emphasis on well planned discharge policies and procedures. The aim of this study was to analyse all discharge policies and procedures currently in use in one large integrated NHS trust in Northern Ireland and formulate a Corporate Discharge Policy for general use in NHS trusts. Objectives of the study included examining the current process of discharge, reviewing the interface between ward staff and district nursing services and examining the quality and standard of documentation in use. Findings indicate the need for standardization of the discharge planning process and sufficient notice of discharge, and for clarification and education regarding staff roles, the importance of multidisciplinary working, and the lack of quality communication between acute and community services. PMID- 11261143 TI - Nurses' perceptions of the nature and frequency of aggression in general ward settings and high dependency areas. AB - Studies on aggression in healthcare settings have reported increased frequency of this behaviour in both psychiatric and accident and emergency areas; however, to date, very few studies have addressed this issue in general ward settings in Australia. This descriptive study was conducted to determine nurses' perceptions of the nature and frequency of aggressive behaviours in general wards and high dependency areas and thereby address the gap in the literature. Two hundred and nine nurses in one Australian hospital completed a 23-item questionnaire on aggression in the workplace. The findings revealed that the majority (89.5%) of nurses defined aggressive behaviour as including verbal aggression, physical aggression and intimidation. Ninety-five per cent of respondents had encountered at least several episodes of verbal aggression within the last 12 months. Patients were found to be the main perpetrators of aggressive acts, followed by relatives. After experiencing either verbal or physical aggression nurses most frequently reported feeling angry or emotionally hurt. These findings confirm that acts of aggression are experienced frequently in the general medical and surgical ward areas of the study hospital. Consequently, there is a need to provide staff with education and support in order to deal with this issue and minimize the impact of increasing levels of aggression in the workplace. PMID- 11261144 TI - Accident and emergency nurses' perceptions and experiences of caring for families. AB - This study is an exploration of accident and emergency (A & E) nurses' perceptions and experiences of caring for families of critically ill/injured patients and suddenly bereaved families. In this study a non-probability convenience sample of 54 nurses working in three A & E departments in Glasgow was studied. A descriptive study design was used and data were collected by use of a questionnaire. Closed questions were statistically analysed using SPSS and open questions were content-analysed into themes and items. Results from the study confirm that participants' perceive of their responsibility to include taking care of patients' families. Participants were familiar with study findings on families' needs and felt that it was important to meet their needs. PMID- 11261145 TI - From simplicity to complexity: developing a model of practical skill performance in nursing. AB - The purpose of this article is to present and discuss a new model of practical skill performance in nursing. The model is conceptualized as having five components: substance and sequence; accuracy; fluency; integration; and caring conduct. The model challenges the truism of 'simple' nursing procedures. It is argued that performance of practical skills in nursing is characterized by complexity on many levels. Complexity lies within and between the components of the performance model and in the interaction between the nurse and the clinical context where practical nursing actions are performed. These complexities are described. Examples that illustrate the complex and reciprocal nature of these components are drawn from an empirical study of graduate nurses' development of practical skill in surgical hospital units. Implications of the model for education, practice and research are discussed. PMID- 11261146 TI - Oral health status of psychiatric patients. AB - Many patients suffering from long-term psychiatric illness are on medication for long periods. These medications frequently cause xerostomia leading to an increased risk of caries, gingivitis, periodontitis and stomatitis. Oral hygiene is therefore of the utmost importance for these patients. Nurses interact with patients on a daily basis, and therefore they are the psychiatric caregivers of choice to support these patients. The main aim of this study was to describe the oral health status of patients in short-term and long-term psychiatric care by means of oral assessment. A second aim was to discover whether the assessment guide used could distinguish any differences between these two groups. A modified version of the Oral Assessment Guide (OAG) developed by Eilers et al. (1988) was used. In addition, new items/categories were developed, forming the Oral Assessment Guide for Psychiatric Care (OAG-PC). A total of 57 patients in psychiatric care, short-term (n = 32) and long-term (n = 25), were assessed by the OAG-PC. Patients in long-term psychiatric care had significantly higher scores on the total OAG-PC compared with those in short-term psychiatric care, indicating a worse oral health status. Statistically significant differences were also found in relation to the following OAG-PC categories: odour from the mouth, mucous membranes, gums, teeth or dentures, calculus on teeth and appearance of teeth. Further research should be focused on the difficulties for nurses in approaching their patients in order to perform oral care and on evaluating the effect of teaching and training psychiatric nurses in oral care, preferably with the assistance of the OAG-PC. This assessment guide may thereby also be valuable for nurses' documentation in estimating, planning, implementing and evaluating their psychiatric patients' oral care needs. PMID- 11261148 TI - Evaluating eye donation. PMID- 11261147 TI - Pain and nutrition as experienced by patients with hip fracture. AB - The purpose of this study was to investigate patients' experiences of care in connection with hip fracture. The care process was examined through non participant observation, informal field interviews and healthcare records. The findings showed that many factors in the healthcare services directly or indirectly influence patients' perceptions of the quality of care. Some of these factors may depend upon a varying knowledge and empathy, while others are due to a lack of agreed protocols/procedures. Patients' needs with respect to pain relief and nutrition are discussed. PMID- 11261149 TI - New OSHA standards managers must know. AB - For home care agencies, compliance encompasses far more than HCFA and the Joint Commission specifics. Other organizations also provide strict guidelines regarding daily operations and activities. This article describes the proposed OSHA ergonomics regulations and provides suggestions for managers on how to prepare for the changes. PMID- 11261150 TI - Transition from acute care to home care nursing: how can management help? AB - Supervisors play a pivotal role in helping new staff adjust to home care. This article highlights some of the major areas that should be in your orientation program and approaches as based on the author's recent research. PMID- 11261151 TI - What to do before PPS hits you! For managers. PMID- 11261152 TI - Strategic recruiting. PMID- 11261153 TI - Fall prevention program for community-dwelling older adults and their caregivers. AB - Falls are the leading cause of injury and death among community-dwelling older adults. Many of these falls are a result of environmental and internal risk factors. The authors developed a fall prevention program consisting of a self administered checklist and an audio-visual presentation on ways to reduce or prevent falls for community-dwelling older adults and their caregivers. PMID- 11261154 TI - Boundary setting as a critical supervisory issue. AB - Setting clear boundaries with patients can clarify the function of a home healthcare agency as well the roles of the various visiting staff, especially that of the nurse. When managers model boundary-setting techniques the visiting professional can more easily adopt the same in patient relationships. This article reviews the basics of this concept along with suggested supervisory strategies. PMID- 11261155 TI - Incident reporting: a vital link to organizational performance. AB - It is critical that the reporting, documentation, and evaluation of incident reports be viewed as an educational process rather than as a punitive one. This article outlines constructive ways you can better your organization by constructively using incident reports. PMID- 11261156 TI - Blood Clot: gaming to reinforce learning about disseminated intravascular coagulation. AB - BACKGROUND: The game, Blood Clot, is an educational tool to reinforce learning following a lecture about disseminated intravascular coagulation (DIC). METHOD: The game was implemented as an educational update for thirty Registered Nurses in a hospital setting. RESULTS: The game reinforced learning of lecture material and stimulated group involvement and communication. CONCLUSION: The self-report evaluations indicate Blood Clot enhanced communication within the group and application of lecture material to solve problems. PMID- 11261157 TI - Cranial Nerve Wheel of Competencies. AB - BACKGROUND: The neuroscience intensive care unit (ICU) staff development committee explored ways to test cranial nerve competencies of staff. With the assistance of the nurse manager, clinical nurse specialist, and education specialist, a game called Cranial Nerve Wheel of Competencies was developed. METHOD: The game tested learners' competency knowledge of cranial nerves. Game participants had the opportunity to attend classes on cranial nerve function and to review written materials prior to the day of testing. RESULTS: Staff evaluated the testing methodology as excellent and preferable to a written test. They found the game challenged their knowledge, yet was not intimidating. CONCLUSION: Gaming as a method to test competency knowledge of cranial nerves was an exciting alternative to written testing or return demonstration. PMID- 11261158 TI - Strategy to assess, develop, and evaluate critical thinking. AB - BACKGROUND: To care for patients with complex health problems, nurses need a strong knowledge base and critical thinking skills. Critical thinking enables the nurse to process and analyze information, solve clinical problems, and decide on actions to take. Teaching and evaluation, however, often focus on memorizing facts and details about clinical care rather than on critical thinking. METHOD: Context-dependent test items are designed to evaluate critical thinking and may be used in orientation, in competency testing, and by preceptors and others who work with beginning nurses for formative evaluation and discussions with them. A context-dependent item presents introductory material to analyze and determine a course of action. The introductory material may be a clinical scenario, an issue nurses might face in their practice, patient data, graphs or flow sheets, and other types of material for analysis. Carefully planned questions for assessing critical thinking are then asked. CONCLUSION: The article describes how to develop and use context-dependent items in nursing continuing education. PMID- 11261160 TI - Leadership development course for creating a learning environment. AB - This article describes a leadership development course designed to prepare leadership to promote cultural changes in a large health care system undergoing an initiative of patient care redesign. Entitled "Creating A Learning Environment," the course is based on Peter Senge's work. His five disciplines are presented as central concepts with practice examples. Characteristics of a learning environment and strategies to promote the cultural change necessary for its formation are explained. PMID- 11261159 TI - Recall Rummy: learning can be fun. AB - BACKGROUND: Nurse educators are continuously seeking creative methods to teach nursing skills. Continuing education programs have adapted and used television game show themes as effective teaching strategies. METHOD: The traditional card game of rummy has been modified into a creative learning technique for entertaining and reinforcing skill techniques for nurses practicing in a clinical setting. CONCLUSION: Imagination and creativity are important assets for planning and teaching skills that relate to the practice of nursing. Recall Rummy presents one such approach to teaching nursing skills. PMID- 11261161 TI - Reinvigorating mandatory safety training: a case example. AB - BACKGROUND: Prior to 1998 the New York Department of Veterans Affairs Medical Center in Manhattan conducted traditional classroom lectures for annual mandatory safety training. METHOD: Instead of offering the traditional program, the Nursing Education Department planned and implemented a week-long, theme-based, self paced, interactive training program that included colorful posters, games, models, and opportunities for manipulation of materials. The program titled "Mandatory Cruise 1998" was offered to all medical center employees. RESULTS: Posttests and reaction evaluations showed learning was both successful and fun. CONCLUSION: This innovative method proved to be a cost-effective time-saver because compliance was accelerated; particularly significant in the current climate of reduced employee resources. PMID- 11261162 TI - Using adult learning concepts to assist patients in completing advance directives. AB - BACKGROUND: Despite the fact that advance directives are legal documents that allow people to direct their future health care, few Americans have enacted them. Because lack of knowledge about what they are, and confusion regarding the process needed to complete one are the primary barriers for completion, patient education can be the key element in promoting the use of advance directives. CONCLUSION: This article offers some teaching strategies nurses may use to address patients' ignorance and confusion and help patients understand that, although advance directives do not help people make end-of-life decisions, they serve as a mechanism by which a person's wishes may be carried out. PMID- 11261163 TI - Use of learning contracts in a RN-to-BSN leadership course. AB - Educating RN-to-BSN students can be challenging and rewarding. The diversity of these students can make designing courses quite difficult. In addition, the RN with experience in nursing practice may resent being put in the traditional role of student. Nominal group process and use of learning contracts in a leadership course in one RN completion program are described as two means of meeting the unique learning preferences of working, adult learners. Houle (1984) identified the learning contract as one means of assisting the adult learner seeking continuing professional education. Accordingly, examples of use of these two strategies in continuing nursing education and staff development are described. PMID- 11261164 TI - Creating a game for sexuality and aging: the Sexual Dysfunction Trivia game. AB - BACKGROUND: Older adults often present with signs and symptoms of sexual dysfunction, and nurses must possess the necessary knowledge to address this issue. Therefore, The Sexual Dysfunction Trivia Game was designed to educate staff nurses about sexual dysfunction and the aging process. METHOD: A pilot test was conducted. Five staff nurses played The Sexual Dysfunction Trivia Game. Each nurse completed a pretest before playing the game and a posttest after playing the game. RESULTS: After playing the board game, the nurses were more knowledgeable about what the physical examination includes, what the laboratory tests are, and what the treatment options are for sexual dysfunction. CONCLUSION: Based on the pretest and posttest findings, The Sexual Dysfunction Trivia Game appears to be an effective teaching tool to educate staff nurses about sexual dysfunction in the older adult. However, more studies are needed to measure its effectiveness. PMID- 11261165 TI - Helping patients live and die challenges, rewards oncology nurses. PMID- 11261166 TI - ONS resources help nurses, patients through a difficult time. PMID- 11261167 TI - ONS helps oncology nurses face the challenges of providing quality end-of-life care. PMID- 11261168 TI - Web sites teach patients, nurses about palliative care. PMID- 11261169 TI - Devolving human resource management. PMID- 11261170 TI - Increasing influence. PMID- 11261171 TI - Sarah Mullally interviewed by Tom Keighley. PMID- 11261172 TI - Planning for change. PMID- 11261173 TI - Appointing a new chief executive for the NHS Executive always generates a debate about people and personalities. PMID- 11261174 TI - COPD taskforce. PMID- 11261175 TI - NEdSERV Consortia. A tool to evaluate service quality in nurse education. PMID- 11261176 TI - Conversion: value for money? PMID- 11261177 TI - New money, new nursing, new management, and new solutions. PMID- 11261178 TI - Promoting health. PMID- 11261179 TI - Supreme Court decisions impact ADA. PMID- 11261180 TI - AAOHN. Managing professional risk in occupational and environmental health nursing practice. PMID- 11261181 TI - Hepatitis B immunity in high risk health care workers. Seven years post vaccination. AB - The purpose of this descriptive, quantitative study was to examine hepatitis B immunity in high risk health care workers 7 years post immunization. The study related immunity to age and site of injection, and also described titer results. The study sample was composed of 42 health care workers from areas with frequent blood exposure. The study found that 21% of the health care workers had nonreactive titers 7 or more years post immunization. No significant differences existed in the percentages of the reactive participants according to injection site, although it is recommended the injection be given intramuscularly in the deltoid. No statistically significant relationship was revealed by comparing titer results to age. The low power (.24) displays the need for larger sample sizes which could be obtained by using multicenter data sites. Future research with national collaboration and standard performance measurement systems could provide crucial information related to hepatitis B immunity in health care workers. PMID- 11261184 TI - Getting published online and in print. Understanding the publication process. PMID- 11261183 TI - Unrecognized high blood pressure. A major public health issue for the workplace. AB - Hypertension continues to be prevalent in the general population despite the public's increased awareness of cardiovascular disease. Population-wide detection and prevention of hypertension are high priority goals within preventive health care. According to recent National Heart, Lung, and Blood Institute (NHLBI) guidelines, high normal blood pressure (BP) (systolic 130 to 139 mm Hg or diastolic 85 to 89 mm Hg) is not an innocuous condition (NHLBI, 1997). High normal BP is a detectable, modifiable, antecedent condition to overt hypertension. Little is known about the incidence of high normal BP in the general population and of its relationship to stress. This study examined the prevalence of high normal and hypertensive levels of blood pressure in a convenience sample of 94 volunteer employees from a midsize corporation. Blood pressure and level of reported stress were assessed. Findings revealed rates of 11% and 30% high normal and hypertensive blood pressure levels, respectively. Ninety-six percent of participants assumed their blood pressures were normal. As in other studies, those employees with hypertensive blood pressure reported higher stress levels than normotensive employees. However, the population with high normal BP did not report significantly higher stress levels than normotensive employees. These findings suggest high normal and hypertensive blood pressures are prevalent cardiovascular disease risk factors among employees in the workplace. Most employees are unaware of their elevated BP and the risk of high normal BP. Occupational health nurses are in a strategic position to take a proactive approach to population-wide hypertension prevention by initiating worksite BP screening and education programs. PMID- 11261182 TI - Work related deaths in West Virginia. Targeting research and prevention efforts. PMID- 11261185 TI - The aging employee. Impact on occupational health. AB - 1. The American work force is aging as a result of several trends: the aging of the baby boomer generation; the decline in the birth rate following that generation; the growing demand for workers; and the extended health and interest of older persons in returning to work or continuing to work. As a result of these trends an increasing number of employers now actively seek to employ the older worker. 2. The occupational health nurse must be prepared to recognize and differentiate between normal age related changes in the employee and changes that are pathological. Job placement requires consideration of both. For this reason, the nurse will need to focus the initial assessment on job specific requirements so the older employee can be both safe and effective on the job. 3. To meet the changing profile of the aging work force, the nurse must recognize the need for job modification to accommodate the older worker's age related changes or chronic conditions, and then advocate on the worker's behalf. When the employee can no longer safely or effectively perform the job, a new assessment is needed to enable a smooth transition to a more appropriate assignment. PMID- 11261186 TI - Promoting health and preventing illness. PMID- 11261187 TI - The reality of rural nursing. PMID- 11261188 TI - Bridging the health gaps with Plunket. PMID- 11261189 TI - Taking surgical services to rural communities. PMID- 11261190 TI - Making primary health care accessible. PMID- 11261191 TI - Defining the role of a rural health nurse. PMID- 11261192 TI - A rural hospital adapts and survives. PMID- 11261193 TI - New industrial legislation promises greater equity. PMID- 11261194 TI - Why we need the Employment Relations Act: a case study. PMID- 11261195 TI - Dancing around the medical maypole. PMID- 11261196 TI - Homebirth and independent midwifery. AB - Why do women choose to give birth at home, and midwives to work independently, in a culture that does little to support this option? This article looks at the reasons childbearing women and midwives make these choices and the barriers to achieving them. The safety of the homebirth option is supported in reference to analysis of mortality and morbidity. Homebirth practices and level of success are compared in Australia and New Zealand (NZ), in particular, and The Netherlands, England and America. The success of popularity of homebirths is analysed in terms of socio-economic status. The current situation and challenges of independent midwifery in Darwin are described. PMID- 11261197 TI - ACMI Bachelor of Midwifery Education Project. PMID- 11261198 TI - Profile of a fellow--Pamela Martin. PMID- 11261199 TI - Does evidenced-based practice medicalise midwifery care? Part 2. AB - In this paper evidence-based care is defined. The evidence to support the provision of care by midwives is presented, as is the evidence to support home birth for those women at low obstetric risk. In conclusion midwives are challenged to be political and use this evidence to support changes to improve the quality of care provided to women and their families. PMID- 11261200 TI - Birthing our daughters--a mother midwife perspective. PMID- 11261201 TI - Anticancer effect of Howiinol A and its mechanism of action. AB - Howiinol A (GHM-10) is a kind of phenylethylene pyrone compounds isolated from Goniothalamus howii. By using the techniques of cell growth curve determination, MTT test, soft agar colony assay and experimental therapy of transplantable tumors in mice, it is found that GHM-10 exerts potent inhibitory effect on cancer cells but its influence on normal cells is relatively slight; the sensitivity of a drug-resistant cell line, KB/VCR 2000, to GHM-10 is similar to its parent cell line KB. Remarkable therapeutic effect can be seen in mice bearing H22 hepatoma and Lewis lung cancer and in mice with ascetic sarcoma 180 when GHM-10 is orally or intraperitoneally administered. The IC50s of L1210 cells treated with GHM-10 for 1 and 24 h are 6.85 and 3.32 micrograms.ml-1 respectively. The ratio of IC50 1 h and IC50 24 h is only 2.06, indicating that the action of GHM-10 is conformed to a cell cycle non-specific cytotoxic agent. By using trypan blue exclusive test and morphological examination, it is demonstrated that the main effect of GHM-10 is to inhibit the cell proliferation. Flow cytometery technique is used to analyze the cell cycle of L1210 cells. The results show that to some extent, GHM 10 blocks the cell cycle transition from G1 phase to S phase. By using [3H] labeled precursor incorporation technique, it is shown that GHM-10 significantly suppresses the biosynthesis of DNA, RNA and protein in L1210 cells, and the DNA synthesis is mostly affected. At 1 h after the cells were treated with GHM-10, these inhibitory effects have already been irreversible, suggesting that GHM-10 may cause structural damage on DNA molecules. However, GHM-10 is unable to intercalate into DNA molecules or to destroy its structure directly. By using single cell gel electrophoresis and alkaline elution technology, it is confirmed that GHM-10 causes DNA molecule damage and single strand breakage in L1210 cells. Further studies show that GHM-10 markedly inhibits DNA dehelix induced by DNA topoisomerase II both inside and outside the cells, indicating that GHM-10 is acting as an inhibitor of DNA topoisomerase II. PMID- 11261202 TI - Oligostilbenes from the roots of Vitis amurensis. AB - Two new oligostilbenes, amurensin A (1) and amurensin B (2), were isolated from the roots of Vitis amurensis Rupr. together with five known oligostilbenes. Their structures were elucidated by means of spectroscopic evidence, and amurensin B was the first naturally occurring stilbene trimer with a cis dihydrobenzofuran moiety. PMID- 11261203 TI - Antitumor components from an actinomycete strain 6011W. AB - Three bioactive compounds that inhibited nucleoside transport were isolated from the cultured broth of Streptoverticillium sp. 6011W. The structures of those compounds were characterized as cinnamamide, N-(tetrahydro-2-oxo-3-thienyl) acetamide and benzamide, respectively. They all inhibited radiolabeled thymidine transport into Ehrlich carcinoma cells, with IC50 values of 30.4, 97.2 and 85.4 microM, respectively. When administered i.p., cinnamamide not only inhibited the growth of transplanted tumors but also reduced the number of lung metastases in mice bearing Lewis lung carcinoma. The results suggest that nucleoside transport inhibition assay is a valuable model to search for antitumor agents of natural origin. PMID- 11261204 TI - Biotransformation of 24 alpha-methylcholesterol and 24 beta-methylcholesterol by yeast mutant GL7. AB - Incubation of 24 alpha- and 24 beta-methylcholesterols with yeast mutant GL7 afforded their corresponding C-22-desaturated products under the catalysis of sterol delta 22(23)-desaturase. The metabolites were identified to be 22-dehydro 24 alpha-methylcholesterol (2% yield from 24 alpha-methylcholesterol) and 22 dehydro-24 beta-methylcholesterol (51% yield from 24 beta-methylcholesterol) respectively on the basis of their chromatographic and spectral properties. It was concluded that the sterol delta 22(23)-desaturase prefers the 24 beta-methyl sterols and is highly stereospecific. PMID- 11261205 TI - A new cycloartane saponin from Cimicifuga acerina. AB - A new cycloartane saponin along with two known compounds, cimigenol and cimigenol 3-O-beta-D-xyloside, was isolated from the rhizomes of Cimicifuga acerina (Sieb. et Zucc.) Tanaka (Ranunculaceae). The structure of the new compound was elucidated as 3'-O-acetyl cimigenol 3-O-beta-D-xyloside on the basis of chemical and spectral evidence. PMID- 11261206 TI - Synthesis of tetradecyl (E)-ferulate, a metabolite of Jatropha gossypifolia. AB - Tetradecyl (E)-ferulate (1) has been synthesized starting from vanillin in four steps with an overall yield of 48%. PMID- 11261207 TI - Sphingosine derivatives from the seeds of Allium tuberosum. AB - A new ceramide, named tuber-ceramide (2), along with a known cerebroside (1) were isolated from the seeds of Allium tuberosum. The structure of tuber-ceramide was determined on the basis of spectral data as N-(2',3'-dihydroxy-tetracosenoyl)-2 amino-1,3,4-trihydroxy octadecane (2). This is the first report of sphingosine derivatives isolated from the genus Allium. PMID- 11261208 TI - Three new bicyclic taxane diterpenoids from the needles of Japanese yew, Taxus cuspidata Sieb. et Zucc. AB - Three new bicyclic taxane diterpenes were isolated from the needles of the Japanese yew, Taxus cuspidata Sieb. et Zucc. Their structures were established to be 7,9,10,13-tetraacetoxy-5-cinnamoyloxy-4-hydroxy-methyl-8,12,15,15- tetramethyl bicyclo[9.3.1] pentadeca-3,8,11-trien-2-ol (2,20-dideacetyl taxuspine X), 7,9,10,13,20-pentaacetoxy-5-cinnamoyloxy-8,12,15,15-tetramethyl bicyclo[9.3.1] pentadeca-3,8,11-trien-2-ol (2-deacetyl taxuspine X), and 9,10,13,20-tetraacetoxy 5-cinnamoyloxy-8,12,15,15-tetramethyl-bicyclo[9.3.1] pentadeca-3,8,11-trien-2,7 diol (2,7-dideacetyl taxuspine X) with the aid of spectroscopic techniques and by comparing with taxuspine X. PMID- 11261209 TI - Two new taxane diterpenoids from the seeds of the Chinese yew, Taxus yunnanensis. AB - A new taxoid and a taxinine analogue were isolated from the seeds of the Chinese yew, Taxus yunnanensis. The structures were established as 13 alpha-acetoxy-5 alpha-(3'-dimethylamino-3'-phenyl)-propionyloxy-11(15-->1)-abeotaxa- 4(20),11 diene-9 alpha, 10 beta-diol and 2 alpha-acetoxy-5 alpha-cinnamoyloxy-9 alpha, 10 beta-dihydroxy-taxa- 4(20),11-diene-13-one on the basis of 1D, 2D NMR, and MS spectral analysis. PMID- 11261210 TI - New iridoids from Pedicularis artselaeri. AB - Three new iridoids, named artselaenin I, II and III, were isolated from the whole plants of Pedicularis artselaeri, along with 11 known compounds, 8-epiloganic acid, 7-deoxy-8-epiloganic acid, plantarenaloside, mussaenoside, lariciresinol-4 O-beta-D-glucoside, lariciresinol-4'-O-beta-D-glucoside, alaschaniosideA, cirtusinA, 2-(p-hydroxyphenyl)-ethanol 1-O-beta-D-glucopyranoside, 3-methoxy-4 primeverosylacetophenone and adenine. Their structures were identified mainly by spectral evidence. PMID- 11261211 TI - Glucosylation of salicyl alcohol by cell suspension cultures of Solanum laciniatum. AB - Cell suspension cultures of Solanum laciniatum were able to transform exogenously inoculated salicyl alcohol into salicyl alcohol 7-O-beta-D-glucopyranoside (isosalicin). The highest level of isosalicin (54.6 mg/g dry weight) in the cells was formed within 2 days after inoculation with salicyl alcohol (37.5 mg/flask containing 50 ml of medium). The biotransformation capacity of the cell suspension cultures was about 31.1%. PMID- 11261212 TI - Studies on dammarane-type saponins in the flower-buds of Panax ginseng C.A. Meyer. AB - From the dried flower-buds of Panax ginseng C.A. Meyer, a new minor dammarane type triterpene saponin named ginsenoside III together with nine known saponins was isolated. On the basis of spectral and chemical evidence, the structure of the new saponin was elucidated as 3-O-[beta-D-glucopyranosyl(1-->2)-beta-D glucopyranosyl]-20-O-beta- D-glucopyranosyl-3 beta,12 beta,20(S)-trihydroxy dammar-25-en-24-one. PMID- 11261213 TI - Lignans from Kadsura angustifolia. AB - A new dibenzocyclooctadiene lignan named angustifolin D (1) together with four known lignans: kadsulignan L (2), kadsulignan N (3), schisantherin P (4) and meso dihydroguaiaretic acid (5) were isolated from the stems of Kadsura angustifolia. Their structures and stereochemistries were elucidated by spectral studies. Compounds 2 and 5 showed moderate platelet-activating factor (PAF) antagonistic activities with IC50 values of 2.6 x 10(-5) and 4.1 x 10(-5) M, respectively. PMID- 11261214 TI - A new flavone 2'-glucoside from Andrographis alata. AB - A new flavone glucoside, 5,2',6'-trihydroxy-7-methoxyflavone 2'-O-beta-D glucopyranoside has been isolated from the whole plant of Andrographis alata. The structure was elucidated on the basis of spectral and chemical evidence. PMID- 11261215 TI - A new bio-active flavonol glycoside from the stems of Butea superba Roxb. AB - A new bio-active flavonol glycoside was isolated from the stems of Butea superba Roxb, and its structure was determined by spectral analysis and chemical degradations as 3,5,7,3',4'-pentahydroxy-8-methoxy-flavonol-3-O-beta-D xylopyranosyl(1-->2) -alpha-L-rhamnopyranoside. The compound 1 showed antimicrobial activity against plant pathogenic fungi Trich viride, Asprgillus fumigatus, A. niger, A. terreus, Penicillium expansum, Helmitnhosporium oryzae, Botxitis cinerea, Rhizopus oligosporus, R. chinensis, Kelbsiella pneumoniae, Fusearium moniliforme and gram-positive bacteria Streplococcus pyogenes, Staphylococcus aureus, Bacillus subtilis gram-negative bacteria Escherichia coli, Proteus vulgaris, Klebsiella pneumoniae, Pseudomonas aeruginosa. The maximum inhibitory effect was shown by H. oryzae, A. niger, B. cinera and gram-positive bacteria. PMID- 11261216 TI - A new flavone glycoside: 5,7,4'-trihydroxy-6,3'-dimethoxy flavone 5-O-alpha-L rhamnopyranoside from the leaves of Tridax procumbens Linn. AB - Tridax procumbens Linn. (N.O. Compositae) is commonly known as Tikki Kasa in Hindi. It is distributed throughout in India up to 2400 m above sea level and in all hot countries. The present paper deals with the isolation and identification of a new flavone glycoside, 5,7,4'-trihydroxy-6,3'-dimethoxy-flavone 5-O-alpha-L rhamnopyranoside 1, from the leaves of this plant. PMID- 11261217 TI - Glabcensin Q-U, five new ent-kaurane diterpenoids from Isodon angustifolius var. glabrescens. AB - Examination of the diterpenoid constituents of the dried leaves of Isodon angustifolius var. glabrescens led to the isolation of five new ent-kaurane diterpenoids, named as glabcensin Q-U (2-6). The structures were elucidated on the basis of spectroscopic evidences. PMID- 11261218 TI - Two new diterpenoid alkaloids, beiwusines A and B, from Aconitum kusnezoffii. AB - Two new atisine-type diterpenoid alkaloids, beiwusine A (1) and B (2), have been isolated from the roots of Aconitum kusnezoffii Reichb. Their structures were established on the basis of spectroscopic data. Beiwusines A and B are the first examples of atisine-type diterpenoid alkaloids having a hydroxyl group at C-1. In addition, one known diterpenoid alkaloid spiramine H (3) has been isolated. PMID- 11261219 TI - A new saponin from the leaves and stems of Panax quinquefolium L. collected in Canada. AB - A new dammarane-type saponin named quinquenoside L3 (1) together with vina ginsenoside R3 (2) were isolated from the leaves and stems of Panax quinquefolium L. collected in Canada. On the basis of physicochemical and spectral evidences, 1 was established as 3-O-beta-D-glucopyranosyl-20-O-[beta-D-xylopyranosyl-(1-->6) beta-D -glucopyranosyl] 20(S)-dammar-23-ene-3 beta, 12 beta,20,25-tetryol. PMID- 11261220 TI - Biotransformation of podophyllotoxin to picropodophyllin by microbes. AB - Biotransformation of podophyllotoxin (PT) by several microbial species has been investigated. Among the fungi tested, it was found that Penicillium strains can isomerize PT to picropodophyllin (PPT) in 8% yield and other strains also transform the substrate into the same product but with lower yield. PMID- 11261221 TI - [Helicobacter pylori and immunogenetic factors of the host: relevance of the HLADQA1 *0102 and *0301 alleles in peptic ulcer]. AB - AIM: To investigate the potential contribution of the *0102 and *0301 alleles of the HLADQA1 gene in Helicobacter pylori infection and peptic ulcer disease in a Spanish Caucasian population. PATIENTS AND METHODS: We studied 163 patients with peptic ulcer (117 duodenal ulcers and 46 gastric ulcers; 111 with recent upper gastrointestinal hemorrhage) and 90 controls. The *0102 and *0301 alleles of the HLADQA1 gene were typed by polymerase chain reaction using genomic DNA. H. pylori infection were determined by breath test and/or serology. The cytotoxins CagA and VacA were investigated using serology (Western-blot) in 98 patients and 48 controls with H. pylori infection. RESULTS: H. pylori infection was found in 94.6% of patients with duodenal ulcer, in 84.4% of those with gastric ulcer and in 67.4% of controls (p < 0.001). The distribution of the *0102 allele of the HLADQA1 gene was similar in patients (31.9%) and in controls (36.7%). The *0301 was more frequent in patients with gastric ulcer (32.6%) than in those with duodenal ulcer (16.2%) (p < 0.05) but no differences were found on comparison with the control group (24.4%). No differences were found when the groups were analyzed according to H. pylori infection, CagA- and VacA-positive strains, consumption of non-steroidal antiinflammatory drugs or previous history of ulcer or hemorrhage. CONCLUSION: The *0102 and *0301 alleles of the HLADQA1 gene did not alter susceptibility to H. pylori infection or the evolution of peptic ulcer disease in a Caucasian population in Spain. PMID- 11261222 TI - [Usefulness of magnetic cholangioresonance in the study of hepatobiliary disease in patients adults with cystic fibrosis]. AB - INTRODUCTION: Because alterations in the bile ducts found in cystic fibrosis mimic those found in primary sclerosing cholangitis, magnetic resonance cholangiography (MRC) could be a useful diagnosis technique, especially because it is non-invasive. MATERIAL AND METHODS: We prospectively studied 26 adult patients with cystic fibrosis. Of these, 11 had liver disease previously diagnosed on the basis of symptomatology, physical examination, liver function tests and abdominal ultrasound (group A) and 15 had no apparent liver disease (group B). In all patients liver function tests, abdominal ultrasound and MRC using 1.5 Teslas General Electric and Siemens systems were carried out. The images were interpreted blind by two radiologists with experience in the interpretation of biliary alterations in cystic fibrosis. RESULT: In 6 of the 11 patients in group A, MRC showed signs of liver cirrhosis (nodularity, irregular surface, splenomegaly, varicosity); 4 patients showed rose-colored images in the choledoch and intrahepatic ducts; of the 5 patients with previous non-cirrhotic liver disease, 2 showed rose-colored intrahepatic ducts, 2 showed dilatation of the intrahepatic ducts and 1 showed hepatosplenomegaly with hepatic steatosis. Of the 15 patients in group B, bile duct anomalies were found in 5. Of these, 3 showed rose-colored images of the hepatic ducts and/or choledoch, 1 showed stenosis of the common hepatic duct with rigidity of the intrahepatic ducts and 1 showed irregularities in the caliber of the intrahepatic ducts without dilatation, which were suspicious for intrahepatic lithiasis. CONCLUSIONS: MRC is a useful technique in the study of hepatobiliary disease in cystic fibrosis because it detected anomalies in all our patients previously diagnosed with liver disease and revealed ductal lesions not revealed by other non-invasive techniques. PMID- 11261223 TI - [Hyperinsulinemia in cirrhotic patients infected with hepatitis C virus]. AB - AIMS: a) To prospectively study the frequency of diabetes mellitus in cirrhotic patients with hepatitis C virus (HCV) infection, comparing it with that in cirrhotic patients without HCV infection and b) to investigate basal insulinemia values in both groups, as well as the possible factors causing insulinemia. MATERIAL AND METHODS: Fifty patients with cirrhosis due to HCV infection (group I) and 50 patients with cirrhosis due to other etiologic agents (group II) were studied. In both groups the percentage of diabetic patients, basal insulinemia values and the factors associated with insulin resistance were compared: age, anthropometric indexes, stage of cirrhosis according to Child-Pugh score, plasmatic ferritin concentrations and treatment with drugs inducing insulin resistance. RESULTS: The percentage of diabetics in group I was 36% (18/50) compared with 18% (9/50) in group II (p < 0.05) and basal insulinemia values were 23.5 +/- 9.7 microU/ml compared with 15.7 +/- 9.9 microU/ml respectively (p < 0.05). No differences between the groups were found in the following variables: age (58.7 +/- 16.2 vs. 60.6 +/- 10.0 years), weight (73.2 +/- 10.7 vs 73.9 +/- 11.2 Kg), height (161.9 +/- 8.8 vs. 161.1 +/- 6.8 cm), body mass index (28.2 +/- 3.1 vs. 28.5 +/- 5.2 Kg/height m2) or Child-Pugh stage (A: 40 vs 34, B: 7 vs. 10, C: 3 vs. 6, NS). In contrast, serum ferritin concentrations were much higher in patients in group I than in those in group II [137.7 (12.4-410.2) vs. 87.6 (2.4 420.0) ng/ml p < 0.05]. At the time of inclusion in this study 10 patients in group I were receiving diuretics or non-selective beta adrenergic blockers compared with 24 patients in group II (p < 0.05). CONCLUSIONS: Diabetes mellitus is more prevalent in patients with cirrhosis due to HVC than in those with cirrhosis due to other etiologic agents. Moreover, basal insulinemia values are higher in these patients, which could be explained by an increase in half insulin resistance associated with an increase in iron deposits. PMID- 11261224 TI - [Hepatocellular carcinoma in a patient with hereditary hemochromatosis without cirrhosis]. AB - Hepatocellular carcinoma in patients with hereditary hemochromatosis in the cirrhotic phase is one of the complications causing greatest mortality and may present in spite of removal of excess iron by bloodletting. Hepatocellular carcinoma is usually considered to occur in cirrhotic livers and consequently measures for the early diagnosis of this complication are only recommended in this type of patient. We present the case of a 69-year-old female patient with non-cirrhotic hemochromatosis who, 6 years after undergoing successful treatment, developed hepatocellular carcinoma. This observation should be added to the 12 cases published in the literature. Criteria should be established for the early diagnosis of hepatocellular carcinoma in patients with hereditary hemochromatosis, irrespective of whether they have cirrhosis. PMID- 11261225 TI - [Endoanal and endorectal echography]. PMID- 11261226 TI - [Autoimmune cholangitis or autoimmune hepatitis with cholestatic component]. PMID- 11261227 TI - [Difficulty in the early diagnosis of pseudoachalasia of tumoral origin]. PMID- 11261228 TI - [Informed consent in digestive endoscopy. Who should inform?]. PMID- 11261229 TI - Extensive intraduct component in invasive duct carcinoma of the breast: prevalence and significance in a south Asian setting. AB - Extensive intraduct component (EIC) in invasive duct carcinoma is one of the main factors affecting local cancer recurrence and thereby a major consideration in breast conserving therapy. A retrospective study was undertaken to assess the prevalence of extensive intraductal component in a South Asian setting. 105 cases of invasive duct carcinoma reported at a University Pathology Department during a 75 month period from January 1992, were reviewed. 48.6 percent of all cases assessed had an intraductal component. 13.3 percent had an EIC. Of the seventy two T1 and T2 tumours reviewed 5.6 percent showed EIC. The results of this study indicate that by virtue of its low prevalence. EIC in infiltrating ductal cancer is unlikely to be a major obstacle in the decision of breast conserving therapy in the South Asian region. PMID- 11261230 TI - Systemic bacterial infections in bone marrow transplant patients. AB - The clinical microbiology department at CMC&H, Vellore in conjunction with the haematology department carries out routine surveillance of patients admitted to the hematology department. Since 1994 in a sample population of 55 patients with various underlying clinical conditions who have had bone marrow transplant, sepsis was observed in 16 patients (29%). The predominant Gram negatives associated with sepsis were non-fermenting Gram negative bacilli and all the 5 Gram positives were coagulase negative staphylococci. These organisms were susceptible to most of the routinely used antimicrobial agents. Continued surveillance is needed to determine changing trends with respect to organisms causing systemic infections and their susceptibility to antimicrobials. PMID- 11261231 TI - Production of monoclonal antibodies to a tumor--associated antigen by spontaneous cell fusion. AB - Spontaneous cell fusion induced by the bacterium Haemophilus paragallinarum has been recently reported as an alternative technique to generate hybridomas producing monoclonal antibody (mAb). In order to investigate the advantages of this technique to produce anti-tumor monoclonal antibodies we performed comparative experiments between H. paragallinarum induced spontaneous cell fusion and polyethylene glycol (PEG) mediated fusion. Hybridomas producing monoclonal antibodies to an experimental murine lymphoma antigen, the Dalton's lymphoma associated antigen (DLAA) were generated and their sensitivity and specificity were ascertained. The spontaneous fusion yielded more number of stable and specific hybridomas than PEG mediated fusion. The results suggest the advantage of H. paragalinarum induced cell fusion for the simplified production of specific antitumor monoclonal antibodies. PMID- 11261232 TI - Can 5' nucleotidase estimation be a predictor of liver metastases? AB - The diagnosis of hepatic metastases is important and has prognostic significance in clinical medicine. A variety of biochemical tests have been used to diagnose liver metastases before surgery and during follow up. However, recently, with the introduction of high resolution imaging modalities like CT Scan/Ultrasonography, the value of these biochemical tests declined. Ultrasonography/CT Scan can pick up liver metastases upto 0.5 cm diameter. Many a times bigger lesions and obviously smaller lesions can be missed by these screening methods. Various biochemical markers were proposed to pick up liver metastases. In this study, the estimation of 5'nucleotidase is found to be the most sensitive predictor of liver metastases when compared to conventional markers and imaging modalities. PMID- 11261233 TI - Surgical speech restoration by tracheo-oesophageal puncture--Kidwai experience. AB - This paper addresses our experience with primary (15 patients) and secondary (8 patients) tracheo-oesophageal puncture (TEP) in the laryngectomee. Despite a success rate of 93.3 percent in the primary TEP and 62.5 percent in secondary TEP, in a follow-up period of one month to eight years, prosthesis related problems like maintenance and recurring expenses emerged as significant deterrent factors in adopting prosthetic speech rehabilitation. Successful oesophageal speech training, increased practice of Pearson's near total laryngectomy, prior tracheostomy and advanced disease mandating post-operative radiotherapy in majority of patients are some of the factors in addition to prosthesis after-care maintenance that makes TEP a less practiced option at our center. PMID- 11261234 TI - Primary CNS lymphoma: an audit of cases treated over a nine year period and review of the literature. AB - Primary CNS lymphoma is a rare tumor comprising around one percent of all brain tumors. This report is an audit of eight cases [5 males, 3 females, age range: 17 55 years] which were accrued over nine years. All patients underwent surgical decompression, followed by radical Radiotherapy [RT]. Five out of eight patients received adjuvant chemotherapy in the form of CHOP or PCV. Of the patients who relapsed two received CHOP as a salvage therapy, one received PCV therapy and lomustine with intrathecal methotrexate. At a median follow up of 16 months [range 1 to 39 months] the two year disease free survival and overall survival were 13 percent and 38 percent respectively, which is in accordance with the literature. High dose RT to whole skull with boost therapy is indicated for all the cases. However, the role of chemotherapy and the appropriate regime needs to be defined with certainty. PMID- 11261235 TI - The management of neuropathic pain in cancer: clinical guidelines for the use of adjuvant analgesics. AB - Neuropathic pain is seen in a third of cancer patients and is not always responsive to traditional analgesics. We describe practical guidelines for the use antidepressants and anticonvulsants as adjuvant analgesics in such situations. Newer adjuvant analgesics, interventional procedures and options for the management of pain emergencies, are also briefly outlined. PMID- 11261236 TI - Familial aggregation of cancer in patients with bronchogenic carcinoma. AB - History of cancer among first degree relatives was obtained in 124 patients with bronchogenic carcinoma [probands] and 248 controls, and differences in familial aggregation evaluated by calculation of odds ratio [OR] and their 95 percent confidence intervals [95% CI]. Probands were more likely than controls to have relatives with cancers [OR 1.27, 95% CI 0.49-3.10], both among smokers and nonsmokers [OR 1.31, 95% CI 0.27-6.79 and OR 1.21, 95% CI 0.12-6.16 respectively]. Sisters of probands were particularly at a higher risk [OR 8.72, 95% CI 0.85-430.08]. A genetic component, possibly independent of smoking habits, may be important in the causation of lung cancer. PMID- 11261237 TI - Embryonal rhabdomyosarcoma of prostate in an adult--a diagnostic dilemma. AB - Embryonal rhabdomyosarcoma of the prostate is a rare. Highly malignant tumour. The median age of occurrence is five years, but sporadic cases have been reported in adults' To the best of our knowledge, till date, fewer than ten cases have been reported of which only two are above the age of sixty years. We report a case of embryonal rhabdomyosarcoma of prostate in a patient more than sixty years of age. If one is not aware of this entity, one can make a mistake in the diagnosis as well as treatment. PMID- 11261238 TI - South Dakota court emphasizes supremacy of hospital board over medical staff. Mahan v. Avera St. Luke's. PMID- 11261239 TI - Gainsharing plan given OIG approval. PMID- 11261240 TI - Expiration of contract between hospital and health plan ruled not a termination. Children's Hospital v. Independence Blue Cross. PMID- 11261241 TI - Design and operation of the National Hospital Discharge Survey: 1988 redesign. AB - The National Hospital Discharge Survey (NHDS), a national probability sample survey of discharges from non-Federal hospitals, began in 1965 and has been conducted annually since then. The original design of NHDS was in place through 1987. This report provides information about the survey design, instruments, data collection procedures, and survey methodology used for NHDS since the implementation of its redesign in 1988. PMID- 11261242 TI - [Presentation. The birth of molecular medicine]. PMID- 11261243 TI - [Predictive medicine: its future]. AB - Predictive medicine or individualized preventive medicine, medicine based on the probabilities of carrying genes for susceptibility or resistance to a pathology, should be practiced with caution after a very strict evaluation of the risks benefits. PMID- 11261244 TI - [The human genome project in the year 2000]. AB - The human genome project was officially launched in 1990. This program started with a mapping phase which led to the development of a genetic map, a physical map based on large DNA fragments and more recently, a map of genes. Since 1996, the programme has progressively shifted to massive sequencing. A spectacular acceleration has occurred during the last 12 months and about 90% of the sequence is at present available in a draft format. This will be soon followed by a more complete version and by the progressive completion of each of the 24 chromosomes, a few of which being already in a "finished" state. It is to be hoped that the genome sequence, which can be used to efficiently identify genes involved in Mendelian phenotypes and which will lead to a better understanding of the evolution process, will also allow us to address other questions, such as those involving multifactorial inheritance. PMID- 11261245 TI - [Contribution of human genetics to the understanding of sensory systems]. AB - As in the other animal species, human senses are adapted to their environment. The energy of photonic, mechanical, thermic or chemical, stimulus is perceived by sensory cells, either dispersed or packed in a sensory end organ. Fifteen years ago, our knowledge of the molecular bases of the function of the various modes of sensory perception was rudimentary with the noticeable exception of that of the phototransduction cascade. Since then, several causative genes for human sensory defects have been isolated. We will examine the outcome of this research in fundamental and medical terms, and bring up the vast yet unexplored fields of the sensory perception. Human genetics should contribute to enlighten the molecular aspects of the latter. PMID- 11261246 TI - [Genomics and type I diabetes]. AB - Type I diabetes is the multifactorial disease for which genome-wide studies were conducted for the first time, both in human and in animal models. Today, two susceptibility loci have been identified: one, at the HLA locus, which represents a major genetic determinant to the disease, and the other at the insulin gene. Studies are in progress to try to map and identify other genetic factors involved in disease susceptibility. The task is made difficult due to the large number of susceptibility genes involved, interacting between themselves and with environmental factors, and most of which carrying only minor effects, which may be heterogeneous between populations. The combination of genetic and biological approaches will be essential in order to elucidate the nature and function of these genes. Recent progress on the knowledge of the human genome, as well as the availability of adequate technologies will accelerate these researches. PMID- 11261247 TI - [Genetic predisposition to infective diseases in humans. Mendelian predisposition to mycobacterial infections]. AB - Selective susceptibility to poorly pathogenic mycobacteria, such as bacille Calmette-Guerin (BCG) vaccine and environmental non-tuberculous mycobacteria (NTM), has long been suspected to be a mendelian disorder but its molecular basis has remained elusive. Recently, recessive mutations in the interferon gamma receptor ligand-binding chain (IFNgR1), interferon gamma receptor signalling chain (IFNgR2), interleukin 12 p40 subunit (IL-12p40), and interleukin 12 receptor beta 1 chain (IL-12Rb1) genes have been identified in a number of patients with disseminated BCG or NTM infection. Although genetically distinct, these conditions are immunologically related and highlight the essential role of interferon gamma-mediated immunity in the control of mycobacteria in man. PMID- 11261248 TI - [Gene therapy for immune deficiencies]. AB - Gene therapy offers an attractive option to the most severe forms of primary immunodeficiency diseases. Identification of disease associated genes as well as advances in the technology of gene transfer into hematopoietic progenitor cells have set the basis for the first clinical trials. Settings characterized by the potential for a selective advantage provided to transduced cells are the first diseases to target. The recent example of successful treatment of Severe Combined Immunodeficiency-X1 (gamma c deficiency) illustrates this potential. PMID- 11261249 TI - [Genomics and cardiovascular diseases]. AB - Conversely to the Pasteur's concept of transmissible diseases where an illness have a unique origin, based on the presence of a micro-organism focusing all fighting efforts, researchers and specialists working in the field of cardiovascular diseases had to integrate, as early as in the fifties, the idea of complexity and multifactoriality in their scientific and medical approaches. The characterisation of several determinants of cardiovascular diseases, mainly environmental, initiated with epidemiological studies and cohort follow-up, suggested that the impact of these factors, increasing or decreasing the risk varied among individuals, partly underlying population differences. A major question is to understand what does this susceptibility stand for and what are its physiopathological bases: lifestyles shared by the same group or genetic specificities of individual. The exponential development of genomics knowledge and techniques allows us to address this question in the context of genome complexity and helps to a better understanding of the occurrence, development and evolution of cardiovascular diseases. Beyond knowledge, genomics is modifying our approaches of treatment and prevention of cardiovascular diseases opening the way to new domains as pharmacogenomics and pharmacogenetics. PMID- 11261250 TI - [Cancer genome or the development of molecular portraits of tumors]. AB - The rapid development of cancer genomics is due to important progresses in oncogenesis, human genome sequencing and emergence of new technologies in genome and transcriptome analysis. In this context, the aim of the French program 'Cartes d'Identites des Tumeurs--Molecular Portraits of Tumors' is to build a public data base containing a pan genome assessment of genome and transcriptome alterations in the major types of tumors as well as in relevant normal cells and experimental models. Data mining is done in the context of genome annotations and clinical and biological informations attached to the enrolled samples. The goal of the program is to define new tests useful for diagnostic procedures in clinical laboratories and new targets for biological treatments of tumors. PMID- 11261251 TI - [The genome and neurology. The example of Parkinson's disease]. AB - In the recent years, many genes involved in inherited neurological disorders have been identified, and the achievement of the human genome project should accelerate their discovery. For common disorders which are of multifactorial origin, the identification of genetic susceptibility factors is still difficult. However, the study of rare monogenic forms of these disorders has proven to be fruitful. An example is Parkinson's disease, in which mutations in the alpha synculein gene are responsible for an autosomal dominant form. The study of alpha synculein led to the conclusion that this protein is a major component of Lewy bodies, which constitute the pathological hallmark of the disease. The study of autosomal recessive forms allowed to demonstrate the relative frequency and the large variety of mutations in the Parkin gene. Parkin is probably involved in ubiquitination of proteins before their degradation by the proteasome and the identification of its cellular targets should allow the understanding of the specificity of neurodegenerative process in the human disease. PMID- 11261252 TI - [Endocrine disruption agents: environment, health, public policies, and the precautionary principle]. AB - The already substantial body of evidence and growing web of suspicions as to the scale and severity of the cascade effects of endocrine disrupters (related to persistent organic pollutants or POPs) on the health of ecosystems and humans have sparked such concern that in June 1998, representatives of 94 countries meeting in Montreal under the aegis of UNEP signed a draft international agreement to phase out the most harmful POPs. Related to particular persistent organic pollutants--toxic semi-volatile and persistent chemical compounds now found everywhere in the environment, such as BPCs, organochlorine pesticides, dioxins and furans, that build up in the bodies of organisms that consume other contaminated organisms along the food chain--endocrine disrupters are strongly suspected of affecting the health of animals and adversely impacting the health, fertility and even intellectual faculties of humans. For example, very low-level exposure to some POPs is associated with some hormone-dependent cancers, damage to the central and peripheral nervous systems, impaired immune system function, reproductive disorders and developmental disruptions in newborns and infants, who can be affected in utero or through breast-feeding. Considering the extreme complexity of the scientific and socio-economic effects of POP-related endocrine disrupters, there are those who, advocate a wait-and-see approach, claiming that there is not enough formal scientific proof. There are others who use the available evidence to advance the research, press for bans on incriminated substances and look for global, integrated and viable alternatives. And there are other still who, with careless disregard for the Precautionary Principle, are quite prepared to talk about the perverse effects of POPs in order to justify the increased use of artificial means of reproduction or the replacement of chemical pesticides by pest-resistant genetically modified organisms (GMOs), thereby opening the door to "solutions" that are potentially more biologically and ethically dangerous than the problems they purport to remedy. This paper provides an overview of the current understanding of the main sources and suspected effects of POPs on animal life and human health, explores the complexity of the scientific, economic and political issues involved in any international process to do away with the incriminated products, discusses the risks and perverse consequences of some of the proposed alternatives, and stresses the importance, in the light of these risks and consequences, of placing renewed emphasis on public and environmental health approaches based on the Precautionary Principle. PMID- 11261253 TI - [Function imaging of the hippocampus in Korsakoff's syndrome]. AB - A case of alcoholic Korsakoff syndrome is reported in a 47 year-old-man. Neuropsychological examination revealed an important both anterograd and retrograd amnesia but procedural and short-term memory were not affected nor was intellectual capacity. Cerebral IRM was normal. Pet scan demonstrated a previously unsuspected fact: the diminution of glucose metabolism in the two hippocampus and in the mamillary bodies. PMID- 11261254 TI - [Training of foreign physicians in France. Analysis and suggestions]. AB - The training of foreign physicians in France raises difficult problems unsolved so far but new measures are being studied. The critical analysis of the current situation can help find out solutions which provide a better answer to current needs both for students wishing to follow a complete training in France and for those, already M. D. willing to come here to get a specialisation course. "DIS" have been suppressed but, getting access to the position of "intern" through the competitive examination even tailor-made may not be the best solution. The background in the home country, the opinions of university supervisors must be determining factors. For specialists already graduated requiring upper level further training it is necessary to improve information and circulation of applications. The success of expected reforms implies awarding a specific budget and modifying in-depth the conditions of choice of training positions. PMID- 11261255 TI - ["Idiopathic" acute optic neuropathies in children]. AB - We reported a retrospective study of 27 children (mean age = 10 years), who presented a loss of vision due to an acute optic neuritis, between 1982 and 1997. The symptoms ar more likely bilateral in children, and frequently associated with systemic viral infection or hepatitis B vaccination. Multiple sclerosis occurs approximately in 20 p. cent of the cases, i.e. less frequently than by adults. Magnetic resonance imaging is now systematic: in 20 children hyperintense nodular abnormalities in a various distribution was demonstrated by 9 children. During the follow-up, 23 eyes of 27 have an excellent recovery of vision, but 4 patients have developed multiple sclerosis. Corticotherapy was uses in 23 cases, in the form of methylprednisolone intravenous flashes in 14 cases. PMID- 11261256 TI - [CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy): clinical features and neuroimaging]. AB - Recently identified in a french family, CADASIL (Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy) is a generalised disease of small arteries, largely predominating in the brain. Its clinical manifestations start during mid-adulthood and include recurrent ischaemic subcortical events, attacks of migraine with aura, severe mood disorders, subcortical dementia, and, at magnetic resonance imaging, widespread leuko encephalopathy. There is so far no specific treatment and the mean duration of the disease is 20 years. CADASIL is most frequently a familial disorder with an autosomal dominant mode of transmission. Its responsible gene, Notch 3, is located on Chromosome 19. By the identification of its gene, CADASIL, (which is now known to affect over 400 families worldwide) is a unique variety of cerebro vascular disease, affecting mainly the subcortical white matter. PMID- 11261257 TI - [CADASIL: genetics and physiopathology]. AB - CADASIL, an autosomal dominant adult onset arteriopathy causing stroke and dementia in humans, is underlaid by a non atherosclerotic non amyloid angiopathy involving mainly the media of small cerebral arteries; it is characterized by major lesions of vascular smooth muscle cells. Using a positional cloning approach, we mapped CADASIL locus on chromosome 19 and identified the mutated gene as being Notch3. This gene, previously unknown in humans, encodes for a large transmembrane receptor belonging to the Notch/lin12 gene family which are known to be involved in cell fate specification during development. Genetic analysis of more than 120 CADASIL unrelated families allowed us to show that these mutations are highly stereotyped and affect only the extra cellular domain of the protein. On the basis of these data, a molecular diagnostic test has been set up and is now widely required by clinicians involved in the diagnosis of vascular leukoencephalopathies. Using this test, we recently showed that CADASIL can also occur in patients who do not have any affected relative due to the existence of notch3 de novo mutations. As a first step to investigate the molecular and cellular mechanisms leading from Notch3 mutations to CADASIL phenotype, we analyzed by in-situ hybridization and immunohistochemistry the pattern of expression of this gene. Notch3 expression is highly restricted to the vascular smooth muscle cell in normal human adults. In CADASIL tissues there is a dramatic accumulation of the extracellular domain of the protein which suggests that one of the main mechanisms of CADASIL involves anomalies in the proteolytical cleavage and clearance of this protein. These data provide important clues to the mechanisms of this condition and current work should lead in the next future to a complete understanding of CADASIL and set up the basis of a rational therapeutical approach of this condition. PMID- 11261258 TI - [Medicine and the precaution principle]. PMID- 11261259 TI - Ultrasound in the ED can mean dramatic improvement in care, research shows. AB - The practice of ultrasound in the ED is a quickly growing trend, and experts predict it will soon become the standard of care in the ED. Ultrasound results can provide evidence to specialists that a patient has a life-threatening emergency, such as an abdominal aortic aneurysm. Studies show that ED use of ultrasound can decrease morbidity and mortality, and reduce length of stay. Document gaps in ultrasound service provided by radiology that the ED can fill. PMID- 11261260 TI - Here's how to get buy-in for ultrasound. PMID- 11261261 TI - Improve communication with portable phones. AB - Portable phones can improve communication with staff and colleagues, patient satisfaction, and the care patients receive. Phones allow you to always reach a designated role, such as a triage nurse, at the same number every day. The ED physician can consult with the primary care physician during the patient's exam. There is less noise in the ED because of fewer overhead pages. PMID- 11261262 TI - [Neuraminidase inhibitors in therapy of influenza]. AB - Neuraminidase promotes influenza virus release from infected cells and facilitates virus spread within the respiratory tract. Several specific inhibitors of these enzyme have been developed. Zanamivir and oseltamivir are the nowadays available neuraminidase inhibitors. In contrast to amantadine and rimantadine, which target the M2 protein of influenza A, they inhibit the replication of both influenza A and B. Zanamivir is delivered by inhalation because of its low oral bioavailability. Oseltamivir can be administered orally. Early treatment reduces the severity and duration of illness and associated complications. These drugs are not effective at afebrile, mild courses, or if the influenza symptoms have existed already for more than 2 days. They are not effective against other respiratory viruses. For an optimal usage of the neuraminidase inhibitors a rapid and reliable diagnosis is necessary. The clinical diagnosis is sufficient only in proven epidemics. An increased availability of sensitive and specifically diagnostical tests is necessary for individual therapy decisions. The influenza vaccination is the most effective measure against influenza. PMID- 11261263 TI - The influence of genetic predisposition on the prevalence of atopic diseases in Carinthian school children. AB - The reasons for the origin and increasing rise in incidence of atopic diseases such as bronchial asthma, allergic rhinoconjunctivitis and eczema in many countries are still unknown. Our survey carried out within the frame of the ISAAC protocol comprised three districts in the Austrian province of Carinthia. A complete study of first and second year elementary school children was done in order to uncover a relationship between "presumed exposure" to different possible risk factors with that of an atopic disorder (i.e. asthma, hay fever, eczema). The results show a significant association between the occurrence of atopic disorders and genetic predisposition and the fact that a carpet which had previously been present in the child's bedroom, had been removed due to the presence of an allergic disease in a family member. The logistic regression model explains a very small part of the overall variability (R2 = 7.32%). PMID- 11261264 TI - [Abdominal vascular surgery emergencies: abdominal aortic aneurysm, acute mesenteric ischemia--indications, technique, results]. AB - Ruptured infrarenal aortic aneurysms and mesenteric ischemia are abdominal emergency situations, which should be treated by vascular surgeons. Modern means of patient transport and specialized emergency centers make it possible to bring patients suspect of having a ruptured aneurysm or a mesenteric ischemia to experienced clinics. Indication for surgery in case of symptomatic or ruptured aortic aneurysms is doubtless absolute. If there is suspicion for a ruptured aneurysm, emergency operation is indicated. In patients having undergone emergency surgery for a ruptured aneurysm of the infrarenal aorta, hospital mortality was 41%. The initial indication for surgery for patients with mesenteric ischemia usually is the "acute abdomen". Mostly only the patient's history reveals the suspicion for this disease. There are no valid radiologic examinations for proving or ruling out mesenteric ischemia. Most of our patients had arterial embolism (64%) as a source of mesenteric ischemia, followed by arterial thrombosis (28%), venous thrombosis (3%) and non-occlusive ischemia (5%). Monitoring of levels of serum lactate can be an additional tool for decision making, if a second look operation is discussed. The key for surgical success with these critically ill patients is shortening of the interval between the first symptoms of the patient and the start of surgical therapy. PMID- 11261265 TI - [Development of a radioligand assay for quantifying specific prostaglandin E1 binding in ischemic ulcers]. AB - Ulcers of the lower extremities are of immense socioeconomical importance. Prevention and therapy of these trophic lesion are hence of great interest. In granulation tissue of ischemic ulcers Laser Doppler flow was previously shown to be higher compared to that in ischemic/adjacent skin or in ulcer without granulation tissue. Intravenous infusion of prostaglandin E1 significantly increased Laser Doppler flow. The present study was designed to test the hypothesis whether the increased baseline and prostaglandin E1-stimulated perfusion of granulation tissue is due to an increased number of prostaglandin E1 receptors in granulation tissue. Therefore, a radioligand binding assay was developed. In an initial pilot study the density of the prostaglandin E1 receptors in granulation tissue of ulcers was compared to that in ischemic/adjacent skin in 8 patients suffering from peripheral arterial occlusive disease requiring debridement or amputation of limbs because of ischemic lesions. The amount of specific binding sites detected was not significantly different between granulation tissue and ischemic/adjacent ulcer tissue. However it cannot be excluded that a possible difference might be detectable with more sensitive assays. In the future we hope to establish a more sensitive assay in order to be able to answer the initial question, whether there is a difference concerning the density of prostaglandin E1 receptor sites in granulation tissue and ischemic/adjacent skin. PMID- 11261266 TI - [Serum soluble CD44 isoform variant 5 level in patients with seropositive rheumatoid arthritis treated with cyclosporin A]. AB - CD44 is a widely expressed cell surface glycoprotein which is involved in many cell-cell and cell-matrix interactions. Expression of soluble CD44 splice variants is strictly regulated and is linked to a high rate of cell division. Serum levels of soluble CD44 variant 5 (sCD44v5) were determined in 14 patients with erosive RA. Patients were divided into two groups. In group 1 cyclosporin A treatment (CYA) was initiated after the first visit. In group 2 preliminary CYA was continued. Controls were performed after 6 months. We found a significant decrease of swollen joint count (SJC) and sCD44v5 in group 1. No effect of CYA was found on c-reactive protein, erythrocyte sedimentation rate and IgM rheumatoid factor (IgM-RF). In group 2 a significant decrease of CRP was found. Therefore we conclude that measurement of sCD44v5 might be useful in monitoring RA+ patients with CYA. PMID- 11261267 TI - Unusual cause of leg oedema and plexopathy in a patient with vitiligo. PMID- 11261269 TI - [Spontaneous healing of retroperitonea fibroasis after successful therapy of sigmoid carcinoma]. AB - In a 64 year old man sigma cancer was diagnosed unexpectedly during an operation for retroperitoneal fibrosis (histologically benign fibrosis), that had caused unilateral hydronephrosis. In the following hemicolectomy this tumor of the colon turned out to be a medium high grade adenocarcinoma (tumor staging pT2, pN1, DUKES C). Chemotherapy with 450 mg/m2 5-FU once a week and a concomitant therapy with laevamisol was added for 6 months. Computer-tomography revealed a significant reduction of the retroperitoneal masses already before induction of chemotherapy. One year after termination of chemotherapy retroperitoneal fibrosis was no longer detectable. The course of events makes us assume that the retroperitoneal fibrosis of our patient was paraneoplastic and therefore completely reversible by successful removal of the underlying tumor. PMID- 11261268 TI - [Retroperitoneal fibrosis as the etiology for recent onset of abdominal pain, nausea and vomiting--a rare and easily missed differential diagnosis]. AB - Retroperitoneal fibrosis is a rare disease with unspecific symptomatic signs. Mortality rates are high with a 10-year mortality rate of 10 to 20%. We describe a case of a 55 year old woman with retroperitoneal fibrosis and discuss clinical findings, symptomatic signs, diagnosis, and treatment of this rare disease. PMID- 11261270 TI - [Autologous transplantation of hematopoietic stem cells--therapeutic spectrum and future developments]. AB - Autologous transplantation with haematopoietic blood stem cells (ASTx) is increasingly performed in blood cell disorders, in solid tumours and recently, in severe autoimmune disorders. In acute leukaemia, ASTx is usually offered to patients in complete remission who lack an HLA-identical donor. The chances and risks must be weighed against a transplant from a matched, unrelated donor. ASTx are routinely performed in aggressive lymphoma. Indications for ASTx in intermediate and low grade lymphoma await results of clinical trials. In Hodgkin's disease and myeloma, ASTx are routinely performed in first or subsequent remission, and in early stages, respectively. Among solid cancers, ASTx is an established part of the therapeutic measures in germ cell tumours. Patients with breast cancer may undergo ASTx in an adjuvant setting after complete tumour resection or in a palliative setting after progression. Initial enthusiasm has switched to a more awaiting view of this indication. Severe autoimmune disorders may undergo haemolymphatic ablation from conditioning and a stem cell transplant. The scientific evaluation of this approach should employ three sequential steps: The first step should employ vigorous immunosuppression followed by ASTx; the second step should employ stem cell autografts depleted from autoreactive immune cells; the third step should employ allogeneic stem cells from normal donors. A truly curative approach after complete haemolymphatic reconstitution is conceivable. PMID- 11261271 TI - [Detection of mammaglobin mRNA as a marker for circulating tumor cells in breast carcinoma]. AB - Mammaglobin (hMAM) has been shown to be a marker for the detection of circulating tumor cells in the peripheral blood (pB) of breast cancer (BC) patients via a nested RT-PCR assay. 286 samples from BC patients were classified into four defined clinical subgroups: prior to and after surgery (pre, post), no evidence of disease (NED) and metastatic disease (MD). hMAM mRNA expression was detected in 2/46 pre (4%), 2/24 post (8%), 4/135 NED (3%) and 35/81 MD (43%) patients. 68 BC patients with NED and negative for hMAM mRNA in their pB were repeatedly tested for at least 6 months. Fifteen of these patients relapsed. Eight of them were hMAM-positive, 5 at time of relapse, one patient 13 months before and two patients 10 and 17 months after relapse was diagnosed. 7/15 BC patients relapsed within 24 months, 5 of them were hMAM-positive versus 3 of 8 patients with later relapses. On the basis of these preliminary results we conclude that tumor cells can be detected via hMAM nested RT-PCR in the pB of BC patients and that hMAM could be a marker for early relapse. PMID- 11261272 TI - [Prolactin as pro-inflammatory cytokine--considerations on consolidated immunotherapy after high dosage therapy]. AB - The pituitary hormone prolactin (PRL) is a proinflammatory cytokine, which cooperates with IL-2. Receptors of IL-2 (beta and gamma) and PRL belong to the same "Cytokine Growth Hormone Receptor Superfamily". PRL is necessary for IL-2 mediated activation and proliferation of T-cells and NK-cells and for LAK-cell generation, where optimal effects are found when low IL-2 levels are combined with PRL levels just above the upper normal range. IL-2 is currently investigated for NK and LAK enhancement after autologous haematopoetic stem-cell transplantation (STx) in order to reduce relapse rates. CONCLUSION 1: Drug induced pulsatile PRL elevation (by metoclopramid) enhancing NK-LAK activity could be an easily feasible bridging therapy for the early posttransplant period, when Il-2 production is impaired and high-dose IL-2 therapy is precluded because of side effects. Low-dose cyclosporin A (Cy-A) after autologous STx induces an autologous graft versus host (GvH) and probably graft versus tumor (GvT) effect. On NK- and T-cells PRL interferes with Cy-A in two different, dose-dependent ways (subtherapeutical Cy-A increases PRL-binding to its receptor 4 fold; therapeutical Cy-A competes with PRL for binding in a dose dependent manner). Cy A inhibits PRL-production on the transcriptional level in pituitary cells. In the thymus, which is a prerequisite for the development of autologous GvH, PRL and Cy A are antagonists. CONCLUSION 2: It is not possible to predict the effect of prolactin elevation or reduction on the development of cyclosporin-A-induced autologous GVH as a means of reducing relapse after stem-cell transplantation. PMID- 11261273 TI - [Cytokine combinations for in vivo and ex vivo expansion of hematopoietic progenitor cells]. AB - The critical role of hematopoietic progenitor cells (HPC) for the formation of mature blood cells is well established. Interleukin-3 (IL-3), stem cell factor (SCF), and flt3-ligand (FL) are potential candidates for expansion strategies due to their early acting and lineage-unspecific hematopoietic stimulation. In preclinical and clinical models IL-3, SCF, and FL induced in-vivo expansion of the progenitor cell pool has been shown to be associated with increased HPC mobilization by G-CSF, reduction in the number of leukaphereses required to obtain adequate HPC-numbers and the potentiation of lineage-specific growth factors. Early acting cytokines are also crucial for ex-vivo expansion strategies and are here important partner molecules for IL-1, IL-6 and IL-10. In-vivo as well as ex-vivo expanded HPC are able to reconstitute hematopoiesis in patients following myeloablative chemoradiotherapy. Thus expanded HPC will be increasingly used for clinical purposes and facilitate therapeutic strategies which are currently limited by the problem of insufficient HPC-numbers. PMID- 11261275 TI - [Stem cell transplantation in germ cell tumors]. AB - High dose chemotherapy (HDCT) with autologous stem cell transplantation in patients with relapsed or refractory non-seminomatous germ cell tumors seems to be the only therapeutic option with curative intention. Evaluation of a scored risk profile before HDCT might significantly support the precise estimation of results after HDCT. Patients with a negative or low score could achieve long lasting complete remissions or might be cured, whereas patients with absolutely refractory disease do not benefit from HDCT. Ongoing prospective randomised trials in Europe and USA comparing standard therapy with HDCT will hopefully contribute to the right therapeutic decision in the future. PMID- 11261274 TI - [Autologous stem cell transplantation in lymphomas and Hodgkin's disease]. AB - High-dose therapy and autologous stem-cell transplantation (SCT) have been explored as a therapeutic option for patients with non-Hodgkin's lymphoma (NHL) and Hodgkin's disease (HD) for the past years, in an effort to improve the long term outcome of these patients. After publication of various phase I/II-studies the results of randomized trials in patients with aggressive NHL and HD have been reported. Based on these the conclusion can be drawn that autologous SCT significantly improves the outcome of patients in chemosensitive relapse of aggressive NHL and HD. In patients with aggressive NHL and a high or high intermediate risk according to the International Prognostic Score, high-dose consolidation therapy may improve their prognosis. This could also be the case in patients with primary refractory HD and some low grade lymphoma such as mantle cell lymphoma. However, there is a lack of randomized trials in patients with low grade NHL and, thus, the benefit of autologous SCT in these populations remains to be determined. PMID- 11261276 TI - [High dosage therapy and autologous peripheral stem cell transplantation in breast carcinoma]. AB - 42 breast cancer patients were treated by high-dose chemotherapy (HDC) and autologous peripheral stem-cell transplantation (ASTx) in the Donauspital between 1992 and 1999. 24 patients had stage II/III breast cancer with high risk for relapse. The other 18 patients underwent HDC and ASTx in chemosensitive stage IV. After previous conventional chemotherapy peripheral stem-cells were harvested by one cycle of mobilisation chemotherapy (epirubicin/taxol, FEC 120 or cyclophosphamide) followed by cytokine stimulation. 16 patients were treated by a tandem transplantation (conditioning protocol for 1st ASTx was melphalan 200 mg/m2 and for 2nd transplant it was CTC: cyclophosphamide 6 g/m2; thiotepa 500 mg/m2; carboplatin 800 mg/m2). The other 26 patients received one HDC with CTC as conditioning protocol. The HDC was well tolerated by all patients, there was no transplant-related mortality. The median survival and the progression-free survival (PFS) after HDC and ASTx in stage IV breast cancer patients were 28 and 11 months, respectively. The median survival and PFS were not yet reached in stage II/III patients after 55 months. The actuarial survival and PFS in that patient group were 70% after 55 months. Our data confirm the low risk and good efficacy of HDC and ASTx in breast cancer patients. Nevertheless randomised studies are necessary to evaluate the importance of HDC compared to intensified conventional protocols without ASTx. PMID- 11261277 TI - [High dosage therapy with stem cell transplantation in neuroendocrine carcinoma]. AB - Neuroendocrine carcinoma and small-cell lung cancer (SCLC) are highly responsive to chemo- and radiotherapy. Nevertheless, most patients (pts.) experience relapse. At the 2nd department of medicine in the Donauspital, 4 pts. with neuroendocrine carcinomas of different primary sites underwent high-dose chemotherapy with autologous stem-cell transplantation (ASTx). Pt. 1 suffered from neuroendocrine lung cancer, pt. 2 from a small-cell carcinoma of the pancreas. Pt. 3 had a metastatic small-cell abdominal bulky tumor and pt. 4 presented with neuroendocrine carcinoma of the prostate. After 4-6 cycles induction chemotherapy pts. were consolidated with 1 cycle of HDCht and ASTx. Prior to HDCht pt. 1 and pt. 2 were in complete remission (CR) and pt. 3 and pt. 4 in partial remission. Pt. 3 converted in CR after HDCht. He is still in CR with a disease-free survival of 23 month after ASTx and 30 month after diagnosis. Pt. 1, 2 and 4 died from relapse 10, 16 and 5 month after ASTx and 16, 22 and 9 month after diagnosis. Pts. with neuroendocrine carcinomas might be suitable candidates for HDCht and ASTx. PMID- 11261278 TI - [High dosage chemotherapy with autologous stem cell transplantation in multiple myeloma]. AB - Between 1992 and 1999 15 patients (pts.) suffering from multiple myeloma (MM) were treated with high-dose chemotherapy and consecutive autologous stem-cell transplantation (ASTx). 10/15 pts underwent two courses of ASTx (tandem- or double ASTx). So 25 ASTx were performed in these 15 pts. in total. All pts. were under 60 a. of age. 13/15 pts. received 6 cycles of chemotherapy on an average according to the VAD-protocol (Vincristin, Adriamycin, Dexamethason). Mobilisation of peripheral hematopoietic stem cells was performed with high-dose cyclophosphamide and hematopoietic growth-factors (CSFs). The conditioning protocol consisted of high-dose melphalan (200-225 mg/m2) in 24/25 ASTx. In one single case total body irradiation (TBI) plus melphalan 140 mg/m2 was used. 2/15 pts. died within 30 days from ASTx; one patient from interstitial pneumonia after TBI, and the other, who was in a very advanced stage of his disease with multiple pretreatment courses before ASTx. The overall survival (OS) was in the mean 68 months, the progression-free survival (PFS) after ASTx 21 m respectively. In pts. with MM high-dose melphalan (up to 225 mg/m2) without TBI plus ASTx is a safe and effective procedure when performed in the early course of the disease. PMID- 11261279 TI - Identification and characterisation of minimal residual disease in solid tumors. AB - Malignant tumors of epithelial tissue are the most common form of cancer and are responsible for the majority of cancer-related deaths in Western industrialized countries. As a result of progress in surgical treatment of these tumors lethality is linked increasingly with early metastasis, which is generally occult at the time of primary diagnosis. The decision as to whether systemic adjuvant therapy should be applied for secondary prevention of metastatic relapse following resection of the primary tumor is based solely on the statistical prognosis. For this reason the direct identification of minimal residual cancer is of particular importance. The studies described below demonstrate the utility of immunocytochemical and molecular analysis in the diagnosis and characterization of minimal residual cancer. For the first time these methods give access to this critical stage of tumor progression and also contribute to the development of new approaches to therapy aimed at preventing manifest metastasis. PMID- 11261281 TI - [Medical responsibility: mode of use]. PMID- 11261280 TI - [The cell cycle and its regulation]. PMID- 11261282 TI - [What is a professional error?]. PMID- 11261283 TI - [Implements for improvement of competence and quality in cytology]. PMID- 11261284 TI - [Medical responsibility of the anatomopathologist]. PMID- 11261285 TI - [Civil and penal responsibility of the cytopathologist. Summary from the symposium of the French Society of Clinical Cytology]. PMID- 11261286 TI - [Histologic examination in fetopathology]. PMID- 11261287 TI - [Sentinel lymph node in breast cancer]. PMID- 11261288 TI - [Deregulation of the cell cycle in bronchial cancers]. PMID- 11261289 TI - [Embryonic origin of diffuse endocrine system cells]. PMID- 11261290 TI - [Immunohistochemistry of neuroendocrine tumors]. PMID- 11261291 TI - [New data in the classification of pulmonary neuroendocrine proliferation]. PMID- 11261292 TI - [Thymic carcinoid tumor: realities]. PMID- 11261293 TI - [Cell cycle, proliferation and stomach cancer]. PMID- 11261294 TI - [Cellular proliferation and apoptosis in the development and progression of renal diseases:pharmacological interventions in genetically modified animals]. PMID- 11261295 TI - [Papillary and vesicular carcinoma of the thyroid: diagnostic problems and new data]. PMID- 11261296 TI - [C cell hyperplasia and medullary microcarcinomas of the thyroid]. PMID- 11261297 TI - [Recent data in the diagnosis of pleural mesothelioma, a lesional spectrum which never ceases to amaze us]. PMID- 11261298 TI - [Cryopreserved tumor cell and tissue bank. An essential role for pathologists in cancer research (Recommendations ANAES)]. PMID- 11261300 TI - [Tools of molecular biology. Diagnostic use of DNA arrays]. PMID- 11261299 TI - [Cell cycle and breast cancer]. PMID- 11261301 TI - [What do clinicians look for in urinary cytology?]. PMID- 11261302 TI - [Tumoral urinary cytology: comparison with histologic lesions according to the 1998 ISUP/OMS classification]. PMID- 11261303 TI - [Diagnostic limits and pitfalls in urinary cytology]. PMID- 11261304 TI - [Urinary cytology during kidney transplantation]. PMID- 11261305 TI - [Quality control and thin layer technology in urinary cytopathology. Preliminary results]. PMID- 11261306 TI - [Thin layer technique and urinary tumors]. PMID- 11261307 TI - [Thin layer technique in the detection of exposed persons]. PMID- 11261308 TI - [Cytometry]. PMID- 11261309 TI - ["Immunocyt" technique]. PMID- 11261310 TI - [BTA test (Bard test) and urinary cytology]. PMID- 11261311 TI - [Oncogene her-2/neu and ovarian carcinoma: intraperitoneal cellular immunotherapy]. PMID- 11261313 TI - [Prognostic contribution and therapeutic implications of microsatellite instability in colorectal cancer. Role of the pathologist]. PMID- 11261312 TI - [Factors predictive of chemotherapy resistance in patients with breast cancer]. PMID- 11261314 TI - [Perspectives on anti-angiogenesis treatments and tumor pathology]. PMID- 11261315 TI - Mathematical models and simulations of bacterial growth and chemotaxis in a diffusion gradient chamber. AB - The diffusion gradient chamber (DGC) is a novel device developed to study the response of chemotactic bacteria to combinations of nutrients and attractants [7]. Its purpose is to characterize genetic variants that occur in many biological experiments. In this paper, a mathematical model which describes the spatial distribution of a bacterial population within the DGC is developed. Mathematical analysis of the model concerning positivity and boundedness of the solutions are given. An ADI (Alternating Direction Implicit) method is constructed for finding numerical solutions of the model and carrying out computer simulations. The numerical results of the model successfully reproduced the patterns that were observed in the experiments using the DGC. PMID- 11261316 TI - An explicit representation of the Luria-Delbruck distribution. AB - The probability distribution of the number of mutant cells in a growing single cell population is presented in explicit form. We use a discrete model for mutation and population growth which in the limit of large cell numbers and small mutation rates reduces to certain classical models of the Luria-Delbruck distribution. Our results hold for arbitrarily large values of the mutation rate and for cell populations of arbitrary size. We discuss the influence of cell death on fluctuation experiments and investigate a version of our model that accounts for the possibility that both daughter cells of a non-mutant cell might be mutants. An algorithm is presented for the quick calculation of the distribution. Then, we focus on the derivation of two essentially different limit laws, the first of which applies if the population size tends to infinity while the mutation rate tends to zero such that the product of mutation rate times population size converges. The second limit law emerges after a suitable rescaling of the distribution of non-mutant cells in the population and applies if the product of mutation rate times population size tends to infinity. We discuss the distribution of mutation events for arbitrary values of the mutation rate and cell populations of arbitrary size, and, finally, consider limit laws for this distribution with respect to the behavior of the product of mutation rate times population size. Thus, the present paper substantially extends results due to Lea and Coulson (1949), Bartlett (1955), Stewart et al. (1990), and others. PMID- 11261317 TI - Coevolutionary interactions between a haploid species and a diploid species. AB - We investigate a general model describing coevolutionary interaction between a haploid population and a diploid population, each with two alleles at a single locus. Both species are allowed to evolve, with the fitness of the genotypes of each species assumed to depend linearly on the frequencies of the genotypes of the other species. We explore the resulting outcomes of these interactions, in particular determining the location of equilibria under various conditions. The coevolution here is much more complex than that between two haploid populations and allows for the possibility of two polymorphic equilibria. To allow for further analysis, we construct a semi-symmetric model. The variety of outcomes possible even in this second model provides support for the geographic mosaic theory of coevolution by suggesting the possibility of small local populations coevolving to very different outcomes, leading to a shifting geographic mosaic as neighboring populations interact with each other through migration. PMID- 11261318 TI - Brucellosis, botflies, and brainworms: the impact of edge habitats on pathogen transmission and species extinction. AB - Ecological interactions between species that prefer different habitat types but come into contact in edge regions at the interfaces between habitat types are modeled via reaction-diffusion systems. The primary sort of interaction described by the models is competition mediated by pathogen transmission. The models are somewhat novel because the spatial domains for the variables describing the population densities of the interacting species overlap but do not coincide. Conditions implying coexistence of the two species or the extinction of one species are derived. The conditions involve the principal eigenvalues of elliptic operators arising from linearizations of the model system around equilibria with only one species present. The conditions for persistence or extinction are made explicit in terms of the parameters of the system and the geometry of the underlying spatial domains via estimates of the principal eigenvalues. The implications of the models with respect to conservation and refuge design are discussed. PMID- 11261319 TI - Randomized controlled monocentric comparison of once daily ceftriaxone with tobramycin and cefotaxime three times daily with tobramycin in neutropenic fever. AB - A prospective, randomized, controlled monocentric trial was performed to evaluate the efficacy and safety of once daily ceftriaxone 2 g plus tobramycin 5 mg/kg in comparison to cefotaxime 2 g t.i.d. plus tobramycin 5 mg/kg qd in the treatment of neutropenic fever. In cases of fever > or = 38.5 degrees C and a neutrophil count below 1000/microliter, patients with hematological malignancies were assigned to ceftriaxone or cefotaxime, each with tobramycin. The primary endpoint was defined as defervescence < 37.5 degrees C on day 4-6 followed by at least 7 afebrile days. Secondary endpoints were overall response, defined as defervescence on day 25 and toxicity. There were 160 episodes of 114 patients included. Fever of unknown origin accounted for 79 episodes (51%), microbiologically defined infection for 36 (23%), clinically defined infection for 27 (17%), and both clinically and microbiologically defined infection for 14 episodes (9%). On an intent-to-treat basis 156 episodes could be evaluated for the primary endpoint. Ceftriaxone plus tobramycin and cefotaxime plus tobramycin resulted in a primary response in 46.9% and 45.3%, respectively. Overall response was achieved on study day 25 in 87.7% and 80%, respectively. No significant difference in toxicity was observed. Once-daily ceftriaxone plus tobramycin was not inferior to cefotaxime t.i.d. plus tobramycin qd in the empirical treatment of neutropenic fever. PMID- 11261320 TI - Successful transplantation and engraftment of peripheral blood stem cells after cryopreservation, positive and negative purging procedures, and a second cryopreservation cycle. AB - Transplantation of peripheral blood stem cells (PBSC), positively and/or negatively selected immediately after harvest, has become a widely applied therapeutic option in hematological or oncological patients. The following case of peripheral blood stem cell transplantation represents the first case of successful transplantation of PBSC, cryopreserved twice and purged after cryopreservation. PBSC were harvested in a 44-year-old female patient with a low grade non-Hodgkin's lymphoma stage IV after mobilization with chemotherapy and G CSF. A total number of 15.2 x 10(6) CD34+ cells/kg bodyweight was harvested with a 36.9% contamination of tumor cells coexpressing CD5 and CD20. After subsequent chemotherapy cycles and cyclophosphamide mobilization, only 0.77 x 10(6) CD34+ cells/kg bodyweight, not sufficient for transplantation, were achieved after positive selection. Therefore, 10.8 x 10(6) cryopreserved CD34+ cells/kg bodyweight were thawed and a positive selection was carried out with the BAXTER Isolex 300i machine. Before additional negative selection, the 0.77 x 10(6) positively selected CD34+ cells/kg bodyweight from the second mobilization were added. A total quantity of 4.4 x 10(6) CD34+ cells/kg bodyweight with a purity of 93.1% representing a recovery of 38% was obtained. Cells were again cryopreserved, stored and retransfused after conditioning the patient with TBI and high-dose cyclophosphamide. The patient engrafted with a WBC count > 1000/microliter on day eight and a platelet count > 20,000/microliter without transfusion support on day 12 post-transplantation. This case indicates that purging procedures can successfully be carried out with cryopreserved cell material and that purified CD34+ cells can be cryopreserved a second time before transplantation, without affecting their hematopoietic capacity. PMID- 11261321 TI - Increased susceptibility of a carrier of X-linked chronic granulomatous disease (CGD) to Aspergillus fumigatus infection associated with age-related skewing of lyonization. AB - Chronic granulomatous disease (CGD) is an inherited disorder characterized by the inability of phagocytes to generate normal amounts of superoxide (O2-), leaving patients susceptible to life-threatening infections. It was previously assumed that once carriers of the X-linked form of CGD were found to have 30% or more of functionally normal neutrophils, they would be free of risk for infection because the lyonization ratio was believed to be constant. Our report strongly contradicts this assumption. A 45-year-old X-CGD carrier had approximately 40% of normal neutrophils in her peripheral blood at age 21 years. Recently, she contracted a life-threatening pulmonary infection with Aspergillus fumigatus. After recovery, the ratio of normal-to-nonfunctional neutrophils was re evaluated. She was found to have only 6-8% of normal neutrophils, suggesting that a striking decrease in the number of normal cells over the past 25 years was the reason for an increased susceptibility to Aspergillus infection. We conclude that age-related acquired skewing of the lyonization ratio can result in an increased susceptibility to life-threatening infections in X-CGD carriers. PMID- 11261322 TI - Disseminated scabies evolving in a patient undergoing induction chemotherapy for acute myeloblastic leukaemia. AB - We report on a 34-year-old refugee from the Balkans presenting with a generalized papular rash during induction chemotherapy for acute myeloblastic leukaemia (AML M4eo). This rash appeared on day 5 of the chemotherapy and was diagnosed as disseminated scabies. It was successfully treated with a combination of oral ivermectin and topical lindane. Scabies disappeared completely despite ongoing neutropenia and other severe infectious complications. Disseminated scabies should be included in the differential diagnosis of rash in severely immunosuppressed patients coming from poor housing conditions. PMID- 11261323 TI - Hepatic veno-occlusive disease in two patients with relapsed acute myeloid leukemia treated with anti-CD33 calicheamicin (CMA-676) immunoconjugate. AB - Monoclonal antibodies recognizing hematopoietic antigens are increasingly being used to target therapy directly at leukemic cells, with the aim of achieving sustained remission with little systemic toxicity. Administration of anti-CD33 calicheamicin immunoconjugate is commonly regarded as being safe, with only moderate systemic non-hematological side effects. We report on two cases of hepatic veno-occlusive disease in heavily pretreated patients presenting with relapsed acute myeloid leukemia (AML). Since significant liver toxicity prevented further specific therapy in both patients, we recommend that antibody therapy with anti-CD33 immunoconjugate should be applied with caution in patients presenting with risk factors for the development of hepatic veno-occlusive disease. PMID- 11261324 TI - A case of pulmonary toxicity associated with G-CSF and doxorubicin administration. AB - The cytokine growth factor, G-CSF (granulocyte colony-stimulating factor), is commonly used in oncologic practice and is generally believed to be a safe agent to administer. We describe here a case of pulmonary toxicity associated with the concurrent administration of G-CSF and doxorubicin. We contend that G-CSF contributed to the life-threatening lung injury in our patient, and discuss additional reports in the literature of pulmonary toxicity associated with the use of this agent. PMID- 11261325 TI - Acquired inhibitor to factor VIII in small cell lung cancer: a case report and review of the literature. AB - Acquired hemophilia (antibodies or inhibitors to factor VIII) is the most common acquired disease affecting clotting factors. It has been described in association with autoimmune disease, malignancy, dermatologic disorders, in the postpartum period, and with drug interactions. Factor VIII inhibitors have been previously described with lung cancer, three with squamous cell and one with adenocarcinoma. A 54-year-old woman presented with weight loss and shoulder pain. A chest X-ray revealed a right hilar mass, confirmed by computed tomography (CT) scan and biopsy revealed small cell lung cancer. Coagulation panel prior to bronchoscopy showed an increased partial thromboplastin time (aPTT). The presence of factor VIII inhibitor was demonstrated at 5 Bethesda units. The patient was treated with fresh frozen plasma twice for hemorrhagic episodes, and six cycles of chemotherapy were begun with carboplatin and etoposide 16. Eight months after the diagnosis, her aPTT was normal and the factor VIII inhibitor titer was undetectable. This is the first case report of small cell lung cancer and acquired hemophilia. A causal relationship between the malignancy and the presence of factor VIII inhibitors is suggested by the response to therapy. PMID- 11261326 TI - HLA-DR4-Ala74 beta is associated with risk and poor outcome of severe aplastic anemia. AB - Severe aplastic anemia (SAA) is a heterogeneous hematological disorder with a high mortality. Genetic predisposition has been shown to play a role in a considerable proportion of SAA cases. For instance, the human lymphocyte antigen HLA-DR2 has been repeatedly demonstrated to be over-represented in SAA patients. In this paper, we expand on the evidence for the contribution of HLA polymorphism in the susceptibility to SAA, which was obtained using the "high-resolution" technique of HLA-DRB1 subtyping. The DRB1*1501 allele appeared to be responsible for the predominance of DR2 specificity in SAA patients and was the most significant risk factor for this disease. It was observed in 23/44 (52.3%) patients versus 22/100 (22.0%) donors [odds ratio (OR) = 3.9; 95% confidence interval (CI): 1.8-8.3; P = 0.0005, corrected P (Pc) < 0.05]. In addition, DRB1*04 alleles also displayed non-random distribution in the SAA group. In particular, DRB1*04 variants coding for alanine at position 74 of the DR beta 1 chain (HLA-DR4-Ala74 beta subtype) were detected in all 13 DR4-positive SAA patients but only in 15/24 (62.5%) controls (OR = 16.6; 95% CI: 0.9-312.0; P = 0.015). Multiple comparison analysis confirmed that the HLA-DR4-Ala74 beta subtype confers susceptibility to SAA independently from the DRB1*1501 allele. Finally, examination of the clinical records has shown that the HLA-DR4-Ala74 beta subtype is associated with poor outcome of SAA. PMID- 11261327 TI - Absence of point mutations within the AML-1 gene in patients with MDS/AML and loss of chromosome 5q or 7. AB - There is increasing evidence that the acute myeloid leukemia 1 (AML-1) gene plays a versatile role in hematopoiesis, and its inactivation has been described in various hematopoetic disorders, e.g., leukemia or familial thrombocytopenia. AML 1 can be affected by various mechanisms, such as chromosomal translocations or point mutations. On the other hand, the specific underlying molecular lesions in myelodysplastic syndromes (MDS) or leukemias with aberrations of chromosomes 5q or 7, respectively, are largely unknown. Despite extraordinary scientific effort no specific genes on chromosome 5q or 7, which act as tumor suppressors, have definitely been identified. Therefore, it has recently been speculated that the AML-1 gene, even if distantly located on chromosome 21q22, may be involved in leukemogenesis in patients with aberrations at chromosome 5q or monosomy 7 [2]. Therefore, we sequenced all exons of the AML-1 gene in 15 patients with MDS/AML and deleted chromosome 5q or 7q, respectively. None of the patients analyzed had any AML-1 mutation. PMID- 11261328 TI - The predictive value of vascular risk factors and gender for the development of thrombotic complications in essential thrombocythemia. AB - The impact of the cardiovascular risk factors smoking, hypertension, hypercholesterolemia, and diabetes mellitus on the risk of thrombotic complications was evaluated retrospectively in 132 patients with essential thrombocythemia (ET). The median age at diagnosis was 51 years, and the median follow-up time was 65 months. Sixty-three out of 132 patients (48%) had one or more vascular risk factors, whereas 69 patients (52%) had no risk factors. Thirty two patients were smokers, 27 had hypertension, 21 hypercholesterolemia, and four diabetes mellitus. During the follow-up, 53 patients (40%) had 98 arterial thrombotic events, half of which were disturbances of cerebral circulation. Fifteen patients (11%) experienced 27 venous thrombotic events. The presence of one or more vascular risk factors increased the risk of arterial thrombotic complications. Of the patients, 52% with one or more vascular risk factors and 29% of those without any risk factors had arterial thrombosis (P = 0.01). In multivariate analysis the only independent risk factor was smoking (P = 0.01). Male gender increased the risk of arterial thrombosis significantly. Thirty-six out of 62 men (58%) but only 17 out of 70 women (24%) had an arterial complication (P < 0.001). Smoking had a strong predictive value for the development of arterial complications in women but not in men. Among women 9/15 (60%) of the smokers and 12/82 (15%) of the non-smokers experienced arterial thrombosis (P = 0.002), whereas among men no difference between smokers and non smokers could be found. According to the present findings, the male gender should be regarded as a risk factor when deciding about the indication for treatment. Smoking should be discouraged especially among women with ET. PMID- 11261329 TI - Ineffectiveness of interferon-gamma in the treatment of idiopathic myelofibrosis: a pilot study. AB - It has been proposed that interferon-gamma (IFN) inhibits collagen synthesis in myeloproliferative disorders through an inhibitory effect on PDGF and TGF-beta. We therefore evaluated the role of IFN-gamma on bone marrow fibrosis in idiopathic myelofibrosis (IMF). After a 3-month observation period, nine patients (five female, four male), median age 64 years (range 43-72 years), received 3 x 3 mU IFN-gamma/week over 6 months and were monitored after withdrawal of IFN-gamma for further 3 months. Three out of nine patients have completed the study according to the protocol. Six patients had to be withdrawn from IFN-gamma due to the following reasons: bacterial infection (three patients), splenic infarction or deterioration of splenomegaly (one patient, each) and refusal to continue IFN gamma (one patient). Results from seven patients treated for at least 8 weeks were considered measurable. Leukopenia, initially present in one of the evaluated patients, deteriorated during IFN-gamma treatment. This patient died during the observation period shortly after withdrawal of the therapy as a result of septicemia. Transfusion-dependent anemia, initially observed in two of the evaluated patients, deteriorated during the IFN-gamma treatment. Bone marrow fibrosis increased in three patients, whereas it remained unchanged in another and improved in a further patient. Splenomegaly improved in two patients but deteriorated markedly in one. Taking these observations together, four patients had disease progression during IFN-gamma treatment, two had stable disease and one could be qualified as a partial responder. According to these data IFN-gamma cannot be considered as a treatment option for patients with IMF. PMID- 11261330 TI - Immunomagnetic enrichment of CD138 positive cells from weakly infiltrated myeloma patients samples enables the determination of the tumor clone specific IgH rearrangement. AB - In multiple myeloma, the polymerase chain reaction (PCR) of the Ig heavy chain with allele-specific oligonucleotide (ASO) primers is a common and well-described method of identifying the tumor clone in peripheral blood (PB), bone marrow (BM) or leukapheresis products (LA). A factor which is crucial to the detection of clonal Ig rearrangements lies in the 'purity' of the tumor tissue used for the consensus PCR. We describe the application of a method to enrich CD138 positive myeloma cells derived from weakly infiltrated PB-, BM- and LA-samples. These are subjected to immunomagnetic enrichment with the MACS system, using an CD138 antibody directly conjugated to magnetic beads to obtain an enriched tumor cell population and the subsequent amplification of tumor specific IgH rearrangements. We investigated 29 samples (ten PB, ten BM, nine LA) with a median myeloma cell content of 0.5%. The approach led to a median enrichment factor of 118. Tumor specific rearrangements could be amplified reproducibly from samples containing less than 0.1% myeloma cells. PMID- 11261331 TI - Molecular and clinical follow-up after stem cell transplantation for multiple myeloma. AB - Molecular follow-up has been carried out using immunoglobulin heavy-chain (IgH) gene finger-printing, a polymerase chain reaction (PCR)-based technique with a sensitivity of 0.1-0.01% (10(-3)-10(-4)), in 22 patients affected by multiple myeloma and submitted to stem cell transplantation (SCT). Twelve patients were submitted to either single or double autologous unselected peripheral blood progenitor cell transplantation, eight patients were submitted to autologous CD34+ immunoselected transplantation and two patients were submitted to allogeneic bone marrow (one patient) or peripheral blood CD34+ stem cell (one patient) transplantation. At diagnosis, all patients showed clonal CDIII rearrangement. The molecular analysis performed on leukapheresis products and CD34+ purified fractions proved to be contaminated by myeloma cells. During follow-up after autografting, all but one patient retained clonal rearrangement despite clinical complete remission (CR) in ten of them. These ten patients either relapsed (Rel) or showed progressive disease (PD) after transplantation; four of them died. Only one patient did not retain clonal rearrangement after autologous transplantation; she is currently alive in CR after a follow-up of 100 months. One patient submitted to allogeneic transplantation is currently alive with no evidence of the disease, but still retains clonal rearrangement after a follow-up of 47 months. Another patient died 4 months after transplantation after succumbing to fatal pneumonia showing myeloma progression. PMID- 11261332 TI - High-dose etoposide phosphate and G-CSF mobilizes peripheral blood stem cells in patients that previously failed to mobilize. AB - Ten consecutive patients in our unit who had failed to mobilize a sufficient stem cell yield after either an initial or several mobilization regimens received high dose etoposide phosphate (1500-2000 mg/m2) followed by granulocyte colony stimulating factor (G-CSF; 10 micrograms/kg per day) to stimulate mobilization. Eight of the ten patients were apheresed. A median of 2.1 x 10(6) CD34+/kg (range 0-5.2) was collected. The number of CD34+ cells/microliter peripheral blood (pB) was significantly increased compared to the first-line mobilization [median 13.0 (range 2.68-29) versus median 4.76 (range 1.36-12); P < 0.05]. Besides hematotoxicity and four cases of infection (WHO grade 3), no major side effects were seen. The median duration of neutropenia was short (5 days, range 0-10), which is important in heavily pretreated patients. These results indicate that high-dose etoposide phosphate with G-CSF is safe, well tolerated, and may be effective in peripheral blood stem cell (PBSC) mobilization in patients who had previously failed to mobilize. PMID- 11261333 TI - [Pacing therapies for congestive heart failure considering the results of the PATH-CHF study]. AB - Optimized uni- or biventricular pacing therapy improves left ventricular function in 80% of patients (responders) with wide QRS in surface ECG. Immediate improvement could be shown by an increase of LV dP/dt up to 28% and aortic pulse pressure up to 16%. Chronic improvement was documented by prolongation of the 6 minute-walk-test by 60 meters, an improvement of O2 uptake by 23% at exercise, and improvement of quality of life score and NYHA classification. This controlled study has shown for the first time a significant clinical improvement of congestive heart failure by pacing therapy in a selected group of patients. Conventional right ventricular stimulation is insufficient in this group of patients characterized by LBBB. These results support the hypothesis that optimized ventricular stimulation is an effective chronic therapy of congestive heart failure by improvement of left ventricular hemodynamics. Epicardial placement of the left ventricular electrodes in 50 patients was possible without operative and with low (2%) early mortality. PMID- 11261334 TI - [Indications, technique and initial results of passive cardiomyoplasty]. AB - The cardiac support device (CSD, Acorn Cardiovasc. Inc.), a knitted polyester tissue, is surgically placed over the ventricles to prevent further dilatation of the heart. The aim of this study was to evaluate the feasibility and safety of CSD implantation in patients with advanced heart failure from cardiomyopathy of either idiopathic or ischemic origin. From April 1999, 23 patients received the CSD. In 8 patients the CSD implantation was the only surgical measure; in 12 patients a concomitant mitral valve repair was performed. In three more patients, the CSD implantation was combined with other surgical procedures. The CSD was placed while on bypass with the heart beating, attached to the AV groove and tailored anteriorly to snugly fit the ventricles. There were no intraoperative deaths or complications. Two patients died early postoperatively (4 d, 21 d); 1 late death occurred (44 d postop). The deaths were not considered to be device related. There were no CSD-related adverse events. Six months postoperatively all patients improved by at least one NYHA class. Echocardiography at 6 months revealed an increase in LVEF; the LVEDD decreased accordingly. Mitral valve regurgitation improved in all patients. These findings indicate that the CSD is safe, and improves heart failure symptoms and left ventricular function. Additional studies have to confirm these results. PMID- 11261335 TI - [Epidemiology-etiology of dilated cardiomyopathy]. AB - Among the cardiomyopathies,--dilated cardiomyopathy (dcm), hypertrophic cardiomyopathy, restrictive cardiomyopathy, arrhythmogenic right ventricular cardiomyopathy--, dcm is the most frequent entity. Its prevalence in the United States amounts to 36 cases per 100,000 inhabitants, men being almost 3-fold more involved than women. The etiology of dcm is very heterogenous; 50% of the cases are due to idiopathic dcm whereas the other half comprises a broad spectrum of various etiologies such as myocarditis, ischemic heart disease, peripartal cardiomyopathy, hypertension, HIV infection, toxic cardiomyopathy and others. In 20 to 30% of the cases of idiopathic dcm a genetic transmission of the disease has been found. Another 20 to 30% of idiopathic dcm are associated with inflammatory and immunological phenomena. Infectious myocarditis with enteroviruses, especially with coxsackie-virus type B has been suggested to be an important trigger for an immune-mediated dcm. In both, familiar dcm and infection with coxsackie-virus B, an impairment of constituents of the myocardial cytoskeleton has been shown. This is regarded as a possible pathogenetic mechanism in the development of dcm. PMID- 11261336 TI - [Dynamic cardiomyoplasty: evaluation of an alternative procedure in the treatment of terminal heart failure]. AB - Dynamic cardiomyoplasty (DCMP) was developed as an alternative treatment for patients with end-stage heart failure. The first clinical application of this technique was in 1985 by Carpentier und Chachques. Since then, DCMP has been performed in more than 1000 patients world-wide. During the initial experience with DCMP, survival for NYHA class IV patients was clearly shown to be much worse than that for class III patients. By careful patient selection, operative mortality has decrease from 31% in the past to less than 5% today. The vast majority of patients have demonstrated significant improvement in NYHA class and overall quality of life with only minor effects on systolic cardiac function. Clinical work, as well as recent animal work supports the hypothesis that by a combination of long-term elastic constraint and active dynamic assist, DCMP decreases myocardial wall stress. This process results in a "reverse remodeling" of the insufficient heart with an improvement of the "contractility reserve". To prove the effectiveness of DCMP versus medical therapy alone, the C-SMART study started in 1994, as the first and only randomized trial. Unfortunately the study was stopped in 1998 due to slow patient recruitment after enrolling 103 patients. The study showed that, from a symptoms standpoint, patients with DCMP were improved over those who were medically treated. However, there was no significant difference for survival between the two groups after 12 months. The lack of a clear survival advantage and the relatively poor and inconsistent hemodynamic benefit of DCMP have hindered its acceptance to date as a treatment alternative for patients with end-stage heart failure. The ultimate role of DCMP in the treatment of heart failure will depend on the outcome of future developments to improve the contractility and the long-term durability. PMID- 11261337 TI - [Current aspects of defibrillator therapy in congestive heart failure]. AB - The beneficial effects of implantable cardioverter defibrillator (ICD) therapy in patients (pts) with life-threatening ventricular tachyarrhythmias and impaired left ventricular (LV) function is still unclear. We studied the follow-up of 410 pts (368 males, 42 females, mean age 57 +/- 11 years) after ICD implant. The LV function was assessed by the New York Heart Association functional class of heart failure (NYHA). Fifty pts (12%) were in NYHA I-II, 151 pts (37%) in NYHA II, 117 pts (29%) in NYHA II-III and 92 pts (22%) in NYHA III. Epicardial ICD implantation was performed in 209 pts (51%) and 201 pts (49%) received nonthoracotomy ICDs. Perioperatively (within 30 days after implant), 12 pts (3%) died, significantly more frequent after epicardial (11 of 209 pts, 5%) than after transvenous ICD implant (1 of 201 pts, < 1%) (p < 0.05). During a mean follow-up of 28 +/- 24 months (range < 1 to 114 months), 90 pts (23%) died: 9 pts (2%) died from sudden arrhythmic death and 5 pts (1%) suddenly, but probably not from arrhythmic causes; 55 pts (14%) died from cardiac causes (congestive heart failure, myocardial reinfarction) and 21 pts (5%) from noncardiac causes. The 3 year, 5-year and 7-year survival was 92% to 96% for arrhythmic mortality in NYHA class I, II and III compared to the 3-year survival of 94%, and a 5-year and 7 year survival of 84% in patients with NYHA class II-III. 338 pts (82%) received ICD shocks (mean incidence 21 +/- 43 shocks per pt); pts in NYHA class II (83%), class II-III (84%), class III (90%) received ICD discharges significantly more frequently than in class I-II (64%) (p < 0.05). Our data show that pts with LV dysfunction benefit from ICD therapy and that these pts survive for a considerable time after the first shock. However, survival is clearly influenced by the degree of left ventricular dysfunction and, in addition to ICD therapy, aggressive treatment of heart failure is necessary. PMID- 11261338 TI - Reduction ventriculoplasty. AB - The partial left-sided ventrical resection in dilatative cardiomyopathy is aimed at improving the ejection efficiency of the heart by reducing the ventricular volume. In the meantime, reduction ventriculoplasty has been used worldwide as an alternative to heart transplantation in a multitude of patients with dilatative cardiomyopathy. The intraoperative death rate ranges from 5 to 10%, and the one year survival rate of the patients ranges from 85 to 90%. One year postoperatively, 70-80% of patients are in NYHA class I. PMID- 11261339 TI - [Endoventricular patch plasty in patients with idiopathic dilated cardiomyopathy: an alternative to heart transplantation?]. AB - BACKGROUND: Although the concept of reducing wall tension as a treatment for advanced heart failure is convincing, clinical data from the Batista operation are conflicting. Despite a number of publications, it is not clear whether left ventricular reduction surgery is truly of benefit for patients with idiopathic, dilated cardiomyopathy (DCM). Surgery may reduce wall tension but the reason for dilation and contractile dysfunction remains. Thus, the potential benefit of the operation may be overshadowed by the natural course of the underlying disease. CASES: We report a series of five cases where left ventricular reduction was performed and physiological geometry was restored in patients with DCM by a modification of Dor's endoventricular patch plasty. All patients demonstrated an improvement in cardiac function immediately after the operation. This improvement was sustained in one of the patients at 18 months follow-up. Another patient developed severe heart failure due to therapy resistant ventricular arrhythmia (Lown IVb), and underwent successful transplantation 4 months after ventricular reduction surgery. Left ventricular dilation reoccurred in two patients 9 and 12 months after reduction surgery, and they were listed for transplant. One patient died after 9 weeks due to sepsis and respiratory dysfunction. CONCLUSIONS: Although the endoventricular patch plasty as used in this study is well tolerated by most patients with dilated cardiomyopathy and results in immediate improvement of contractile function, the long-term benefits of this technique for DCM are uncertain. Thus, the technique is currently not an alternative for heart transplantation. However, the procedure may be an option in patients with contraindications for transplantation. PMID- 11261340 TI - Clinical laboratory services' high-tech future. PMID- 11261341 TI - A few cautionary notes about lab services in the millennium. PMID- 11261342 TI - Seven strategies for tomorrow's clinical laboratories. PMID- 11261343 TI - Clinical evaluation of the Shin-Nippon SRW-5000 autorefractor in adults. AB - A clinical evaluation of the Shin-Nippon SRW-5000 (Japan), a newly released commercial autorefractor, was undertaken to assess its repeatability and validity compared to subjective refraction. Measurements of refractive error were performed on 200 eyes of 100 subjects (aged 24.4 +/- 8.0 years) subjectively (non cycloplegic) by one optometrist and objectively with the SRW-5000 autorefractor by a second. Repeatability was assessed by examining the differences between the seven autorefractor readings taken from each eye and by re-measuring the objective prescription of 50 eyes at a subsequent session. Although the SRW-5000 read slightly more plus than subjective refraction (mean spherical equivalent +0.16 +/- 0.44 D), it was found to be highly valid (accurate) compared to subjective refraction and repeatable over the prescription range of +6.50 to 15.00 D examined. The Shin-Nippon SRW-5000 autorefractor is therefore a valuable complement to subjective refraction and as it offers the advantage of a binocular open field-of-view, has a great potential benefit for accommodation research studies. PMID- 11261344 TI - Continuous recording of accommodation and pupil size using the Shin-Nippon SRW 5000 autorefractor. AB - A newly released commercial autorefractor, the Shin-Nippon SRW-5000 (Japan), has been found to be valid compared to subjective refraction and repeatable over a wide prescription range. Its binocular open field-of-view allows the accommodative state to be monitored while a natural environment is viewed. In conventional static mode, the device can take up to 45 readings in 1 min using digital image analysis of the reflected retinal image of a measurement ring. Continuous on-line analysis of the ring provides high (up to 60 Hz) temporal resolution of the refractive state to an accuracy of < 0.001 D. Pupil size can also be analysed to a resolution of < 0.001 mm. The measurement of accommodation and pupil size was relatively unaffected by eccentricity of viewing up to +/- 10 degrees and instrument focusing inaccuracies of +/- 5 mm. The resolution properties of the analysis are shown to be ideal for measurement of dynamic accommodation and pupil responses. PMID- 11261345 TI - Interfacing the Shin-Nippon autorefractor SRW-5000 with a personal computer. PMID- 11261346 TI - Sighting dominance, handedness, and visual acuity preference: three mutually exclusive modalities? AB - It is tempting, even perhaps for the clinician, to assume prima facie that an individual's handedness is indicative of other lateral asymmetries, including ocular (sighting) dominance and preferred monocular acuity. An analysis of new data relating to these three modalities, as collated from counter-balanced groups of normally sighted male and female children and adults examined in optometric practice, confirms the general fallacy of this assumption and considers why it is such a persistent misconception. The degree of association between the three modalities in right-preferent individuals is revealed as statistically no greater than chance. On the basis of this study, estimates of right-sided hand, eye and/or acuity congruency are derived for the information of the clinician in the prescribing environment of the consulting room. PMID- 11261347 TI - A prospective study of contact lens wear in diabetes mellitus. AB - A prospective, controlled, observer-masked study was conducted to investigate the suitability of contact lenses for patients with diabetes mellitus. Forty diabetic patients and 40 non-diabetic control subjects were fitted with soft hydrogel contact lenses to be worn on a daily wear basis for 12 months. The ocular response was assessed using slit lamp biomicroscopy, ultrasonic pachometry, corneal aesthesiometry and visual acuity measures. Compared to non-diabetic subjects, diabetic patients displayed significantly reduced corneal transparency, variable vision and reduced comfort with the contact lenses (p < 0.05). There were no significant differences between the two groups with respect to ocular hyperaemia, corneal staining, corneal thickness, corneal sensitivity or high contrast visual acuity. Contrary to previous reports, the response of the diabetic eye to contact lenses--as observed clinically--does not differ appreciably from that of the non-diabetic eye. These results suggest that current generation daily wear soft contact lenses can be a viable mode of vision correction for diabetic patients. PMID- 11261349 TI - Fatigue reduces tonic accommodation. AB - Ocular accommodation adopts a mean baseline response level of approximately 1.0 D in the absence of blur feedback (open-loop state). This baseline or tonic accommodation (TA) can be elevated following a sustained monocular accommodative response to a dioptric stimulus (lens adaptation) that exceeds the baseline open loop level of TA. The accommodative response to the lens persists in the open loop state (accommodative hysteresis), and eventually decays to a stable end point. Interestingly, if the baseline TA is high, the monocularly adapted accommodative state can decay to an end-point that is below the initial pre adapted baseline level of the TA (counter-adaptive response) (McBrien, N.A. and Millodot, M., (1988). Differences in adaptation of TA with refractive state. Invest. Ophthalmol. Vis. Sci., 29, 460-469). We have investigated the possible contribution of accommodation fatigue to the counter-adaptive change in baseline TA following sustained accommodation. Two fatigue procedures were used while viewing a target at 66 or 33 cm. In a monocular condition, accommodation was stimulated for 3 min with lens values alternating from -1.5 to +1.5 D at a rate of 0.25 Hz. In the binocular condition, convergence was stimulated with alternating prism values from 9 prism diopters (PD) base-out to 9 PD base-in. Both monocular and binocular tasks resulted in a significant reduction of TA. These results suggest that previously reported reductions of baseline TA following sustained monocular accommodation or binocular convergence could have resulted from fatigue of the accommodative system. Accommodative fatigue could be responsible for the lower values of TA observed in counter-adaptive responses to sustained accommodative or convergence effort. PMID- 11261348 TI - How large is the optic disc? Systematic errors in fundus cameras and topographers. AB - PURPOSE: To determine whether or not there are systematic differences in the areas of optic discs as measured by different machines using different measurement algorithms and whether racial or gender differences exist in optic disc area measurements. METHODS: We examined the results of twenty-three published studies on the size of normal optic discs of various patient populations. Studies differed in the type of instrument and method used to measure optic disc area, and the number, age, race and gender of subjects examined. Noticing that different machines exhibited statistically significant systematic differences in optic disc sizes of comparable populations, we computed a "normalization" factor for each machine based on these mean differences. Applying this normalization factor to the results, we then re-examined the differences between racial and gender groups. RESULTS: By comparing the results of mean optic disc areas of different racial groups made with different machines, and normalizing results according to those of the Zeiss fundus camera, we found the normalization factors for the following machines to be, Zeiss fundus camera: 1 (by definition), Rodenstock Optic Disc Analyzer (RODA): 1.51, Topcon fundus camera: 1.04, Heidelberg Retina Tomograph (HRT): 1.15 and TopSS scanning laser ophthalmoscope: 1.29. That is, to bring the results of area measurements made with a RODA machine in line with those made with a Zeiss fundus camera, one should multiply the former by the factor 1.51. Using the normalized results, we confirmed the findings of previous authors that the optic disc areas of black subjects were statistically significantly larger than those of white subjects (n weighted mean effect = 0.556 +/- 0.142 S.E., n = 5). Further, the meta-analysis of various racial populations from five studies shows that males have significantly larger discs than females (n-weighted mean effect = 0.151 +/- .055 S.E., n = 9). CONCLUSION: Different machines and techniques give different results when populations of similar racial composition are measured. We recommend applying the above normalizing factors when comparing studies that employ different instruments. PMID- 11261350 TI - Description of a method for neutralising the Stiles-Crawford effect. AB - The influence of the Stiles-Crawford effect on visual performance can be investigated by filters based on the apodisation model of the Stiles-Crawford effect. We describe the development of practical filters to achieve neutralisation. We present some results of the Stiles-Crawford function showing that the filters work well for expected errors in aligning filters in front of the eye. PMID- 11261351 TI - Clinical evaluation of the Shin-Nippon SRW-5000 autorefractor in children. AB - The Canon Autoref R-1 is an 'open-field' autorefractor which has been widely used for research purposes for the past 20 years, but is no longer manufactured. A new autorefractor, the Shin-Nippon SRW-5000, is now available, and if measures using this instrument are shown to be equally accurate and reliable, is likely to replace the R-1. Here we report on the accuracy and reliability (repeatability and reproducibility) of refraction measures in a paediatric population (from 4 to 8 years of age). Subject numbers were 44 for cycloplegic measures and 53 for non cycloplegic measures. As would be expected, agreement with cycloplegic refraction and reliability were better when SRW-5000 measures were taken using cycloplegia. Repeatability results from the SRW-5000 autorefractor, both with and without cycloplegia were similar to those reported for the Canon R-1. PMID- 11261352 TI - Images of the invisible-prospection methods for the documentation of threatened archaeological sites. AB - To understand the development of prehistoric cultural and economic activities, archaeologists try to obtain as much relevant information as possible. For this purpose, large numbers of similar sites must be identified, usually by non destructive prospection methods such as aerial photography and geophysical prospection. Aerial archaeology is most effective in locating sites and the use of digital photogrammetry provides maps with high accuracy. For geophysical prospection mainly geomagnetic and geoelectrical methods or the ground penetrating radar method are used. Near-surface measurements of the respective contrasts within physical properties of the archaeological structures and the surrounding material allows detailed mapping of the inner structures of the sites investigated. Applying specially developed wheeled instrumentation, high resolution magnetic surveys can be carried out in a standard raster of 0.125 x 0.5 m covering up to 5 ha per day. Measurements of ground resistivity or radar surveys in a raster of 0.5 or 0.5 x 0.05 m, respectively, are used to gain information on archaeological structures and on the main stratigraphic sequence of sites covering up to 0.5 ha per day. Data on intensities of the Earth's magnetic field, apparent resistivities of the ground or amplitudinal information of radar reflections are processed using a digital image processing technique to visualize the otherwise invisible archaeological structures or monuments buried in the ground. Archaeological interpretation, in the sense of detecting, mapping and describing the archaeological structures, is done using GIS technology by combining all relevant prospection data. As most of the Middle European archaeological heritage is under a massive threat of destruction, dramatically accelerated by intensive agriculture or industrial transformation of the landscape, the prospection techniques presented here represent an approach towards an efficient documentation of the disappearing remains of our ancestors. PMID- 11261353 TI - The immunoglobulin-like genetic predetermination of the brain: the protocadherins, blueprint of the neuronal network. AB - The morphogenesis of the brain is governed by synaptogenesis. Synaptogenesis in turn is determined by cell adhesion molecules, which bridge the synaptic cleft and, by homophilic contact, decide which neurons are connected and which are not. Because of their enormous diversification in specificities, protocadherins (pcdh alpha, pcdh beta, pcdh gamma), a new class of cadherins, play a decisive role. Surprisingly, the genetic control of the protocadherins is very similar to that of the immunoglobulins. There are three sets of variable (V) genes followed by a corresponding constant (C) gene. Applying the rules of the immunoglobulin genes to the protocadherin genes leads, despite of this similarity, to quite different results in the central nervous system. The lymphocyte expresses one single receptor molecule specifically directed against an outside stimulus. In contrast, there are three specific recognition sites in each neuron, each expressing a different protocadherin. In this way, 4,950 different neurons arising from one stem cell form a neuronal network, in which homophilic contacts can be formed in 52 layers, permitting an enormous number of different connections and restraints between neurons. This network is one module of the central computer of the brain. Since the V-genes are generated during evolution and V-gene translocation during embryogenesis, outside stimuli have no influence on this network. The network is an inborn property of the protocadherin genes. Every circuit produced, as well as learning and memory, has to be based on this genetically predetermined network. This network is so universal that it can cope with everything, even the unexpected. In this respect the neuronal network resembles the recognition sites of the immunoglobulins. PMID- 11261354 TI - Sulfur nutrition of deciduous trees. AB - Sulfur in its reduced form (-II) is an essential nutrient for growth and development, but is mainly available to plants in its oxidised form as sulfate. Deciduous trees take up sulfate by the roots from the soil solution and reduce sulfate to sulfide via assimilatory sulfate reduction in both roots and leaves. For reduction in the leaves, sulfate is loaded into the xylem and transported to the shoot. The surplus of sulfate not reduced in the chloroplast or stored in the vacuole and the surplus of reduced S not used for protein synthesis in the leaves is loaded into the phloem and transported back to the roots. Along the transport path, sulfate and glutathione (GSH) is unloaded from the phloem for storage in xylem and phloem parenchyma as well as in pit and ray cells. Remobilised S from storage tissues is loaded into the xylem during spring, but a phloem to xylem exchange does not appear to exist later in the season. As a consequence, a cycling pool of S was only found during the change of the seasons. The sulfate:glutathione ratio in the phloem seems to be involved in the regulation of S nutrition. This picture of S nutrition is discussed in relation to the different growth patterns of deciduous trees from the temperate climate zone, i.e. (1) terminated, (2) periodic and (3) indeterminate growth patterns, and in relation to environmental changes. PMID- 11261356 TI - Inverse calefaction. PMID- 11261355 TI - Food caching in orb-web spiders (Araneae: Araneoidea). AB - Caching or storing surplus prey may reduce the risk of starvation during periods of food deprivation. While this behaviour occurs in a variety of birds and mammals, it is infrequent among invertebrates. However, golden orb-web spiders, Nephila edulis, incorporate a prey cache in their relatively permanent web, which they feed on during periods of food shortage. Heavier spiders significantly reduced weight loss if they were able to access a cache, but lost weight if the cache was removed. The presence or absence of stored prey had no effect on the weight loss of lighter spiders. Furthermore, N. edulis always attacked new prey, irrespective of the number of unprocessed prey in the web. In contrast, females of Argiope keyserlingi, who build a new web every day and do not cache prey, attacked fewer new prey items if some had already been caught. Thus, a necessary preadaptation to the evolution of prey caching in orb-web spiders may be a durable or permanent web, such as that constructed by Nephila. PMID- 11261357 TI - ECG of the month. Shades of evil. Inferolateral myocardial infarction. AB - The tracings shown below, 12-lead ECG and rhythm strip (lead II), belong to a 74 year-old man, and were recorded on his eighth hospital day. On admission, he gave a history of a "heart attack" about 11 days earlier. PMID- 11261358 TI - Base of tongue carcinoma. AB - Vigilance must remain high in order to detect the rare occurrence of base of tongue carcinoma. The combination of ubiquitous and nonspecific complaints coupled with difficult physical examination of this area contribute to delays in diagnosis, with patients often presenting at an advanced stage. This advanced presentation creates challenging treatment decisions in which adequate tumor control must be balanced with subsequent patient quality of life. The result is controversy over the primary treatment modality--external beam radiation, with or without brachytherapy, alone or in combination with surgical extirpation. Though the role of neck dissection in patients with neck disease is widely accepted, timing and implementation in patients whose neck disease completely responds to radiation are also contentious issues. Although evolving surgical reconstruction techniques and accelerated radiation protocols are improving treatment regimens, the prognosis for this disease remains poor. PMID- 11261359 TI - Radiology case of the month. All that wheezes is not asthma. Foreign body obstruction FBO of left main stem bronchus with a Luden cough drop. PMID- 11261360 TI - Medical societies of Louisiana in the 19th century. PMID- 11261361 TI - Delayed epidural hematoma: presentation in a pediatric patient. AB - Epidural hematomas remain at the pinnacle of neurosurgical emergencies, representing approximately 3% to 8% of all serious head injuries. Mortality from this entity is usually prevented once the diagnosis is clear. Although readily recognized on non-contrast head CT, the occasional patient may go on to develop a clinically significant hematoma after an initial negative CT. This phenomenon is appropriately termed delayed epidural hematoma. We present a case in which a 3 year-old boy developed a large epidural hematoma, which was not evident until the second CT, several hours after the injury. We feel that maintaining a high clinical suspicion, coupled with a low threshold for CT scanning, is the key to morbidity prevention in this illness. PMID- 11261362 TI - Infant mortality in Louisiana--identifying the risks. AB - The state of Louisiana consistently has one of the five highest infant mortality rates in the nation and the racial disparity within the state is glaring. The purpose of this paper is to analyze the state's linked birth and death infant data set for the birth cohort of 1990-1998, and to identify the "at risk" population. We have analyzed data from the 1990 to 1998 birth and infant death cohort in Louisiana to determine mothers' characteristics that are associated with infant death. These include extremely young and old ages, unmarried marital status, lower levels of education, and prenatal care. A mother's race is also associated with higher future mortality for her infant, with black mothers and their infants being at greater risk than white ones. Because the infant mortality rate is higher for black infants than for white ones, Louisiana's higher infant mortality rate can be seen as a result of the higher proportion of black births. Main concerns in reducing infant mortality rates include reducing incidence of low birth weights specifically among black births primarily by providing adequate prenatal care. PMID- 11261363 TI - Clinical application of botulinum toxin in otolaryngology, head and neck practice (brief review). AB - Botulinum toxin (Botox) is useful in controlling the symptoms of patients with movement disorders. Application of Botox serves to (1) inhibit hypertonicity, (2) enhance the action of the antagonistic muscles, and (3) avoid an impingement in order to reestablish "the balance of forces". In accordance with the principles mentioned above, Botox can be used to treat dystonias of the larynx (adductor laryngeal spasmodic dysphonia, abductor laryngeal spasmodic dysphonia), laryngeal granulomas, laryngeal joint dislocation, cricopharyngeal spasm, and posterior glottic synechiae. In addition, extra-laryngeal disorders such as blepharospasm, hemifacial spasm, oromandibular dystonia, and spasmodic torticollis respond well to Botox. The effects of Botox are reversible and have specific localized activity. Hence, Botox has served as a powerful diagnostic method in exploring the underlying mechanism of various types of dystonias and provides some therapeutic benefits before pursuing surgical options. Here we review the literature and describe our experiences with Botox, including such topics as preparing and storing Botox, identifying the target muscles under EMG-guidance, choosing an appropriate dose, and outlining the applications of Botox in Otolaryngology, Head and Neck Surgery practice. PMID- 11261364 TI - Folic acid: miscarriages, anomalies, thromboses, cancers. AB - Folic acid is an essential micronutrient that merits special attention. In spite of plentiful sources in a balanced diet, recent data indicate that folic acid intake of many persons has long been inadequate. To some degree, this is due to destruction of folic acid by storage and processing of foodstuffs and to dietary practices that vary from recommendations. However, the common hereditary thermolabile variant of methylenetetrahydrofolate reductase (MTHFR) imposes an increased folic acid requirement which, if not met, may result in recurrent early pregnancy loss, anomalous progeny, thrombotic disorders, and cancers. There are recent reports that folic acid metabolism is also altered by other hereditary variations of MTHFR and other enzymes involved in folic acid metabolism. Optimal management requires dietary guidance and, for many patients, folate supplements. Caution is required because mandated folate fortification of grain products may produce levels that are inadequate for some patients and excessive for others. Laboratory tests for red cell folate and serum homocysteine are valuable adjuncts. PMID- 11261365 TI - Mushrooming liability crisis threatens all nursing homes, not just Florida providers. PMID- 11261366 TI - Can HCFA's quality indicators really identity poor care? PMID- 11261367 TI - Births, marriages, divorces, and deaths: provisional data for 1999. PMID- 11261368 TI - Costs rising, managed care gone: bad combo, says Ginsburg. PMID- 11261369 TI - Reentering M+COs forced to reenroll enrolled beneficiaries. PMID- 11261370 TI - HCFA grapples with PPO regulation. PMID- 11261371 TI - Marketplace. Will M+C be Wennberg's window of Medicare opportunity? PMID- 11261372 TI - A 'spigot of waste'. As Medicare-overpayments decline falters, some say more needs to be done. PMID- 11261373 TI - La. payment feud escalates. Alexandria's Rapides system hits Blues for directly reimbursing patients. PMID- 11261374 TI - Single-minded. Nursing homes willing to flout law cracking down on 'single-task workers'. PMID- 11261375 TI - Building new strategies. Construction firms coping with array of fiscal and corporate challenges. PMID- 11261376 TI - Ready to pull the cord. Thanks to decision by Quorum board, CEO packs a hefty golden parachute. PMID- 11261377 TI - Unlikely saviors. HCA or Tenet could rescue not-for-profit hospitals in Florida deal. PMID- 11261378 TI - Can IOM report find a strong perch? PMID- 11261379 TI - The 800-pound gorilla returns. Healthcare spending rising sharply again; new federal cuts foreseen. PMID- 11261380 TI - Shutting down to survive. Rural Georgia hospital closing temporarily to prepare for critical inspection. PMID- 11261381 TI - The heart of the matter. PMID- 11261382 TI - The Web's dark secret. PMID- 11261383 TI - Is it sexual exploitation if victims are 'virtual'? PMID- 11261384 TI - Multiplicity's perils. PMID- 11261385 TI - Vaccine worries get shot down but parents still fret. PMID- 11261386 TI - A prospective test of the dual-pathway model of bulimic pathology: mediating effects of dieting and negative affect. AB - Because there have been few longitudinal investigations of integrative etiological theories of bulimia nervosa, this study prospectively tested the dual pathway model using random regression growth curve models and data from a 3-wave community sample of adolescent girls (N = 231). Initial pressure to be thin and thin-ideal internalization predicted subsequent growth in body dissatisfaction, initial body dissatisfaction predicted growth in dieting and negative affect, and initial dieting and negative affect predicted growth in bulimic symptoms. There was prospective evidence for most of the hypothesized mediational effects. Results are consistent with the assertion that pressure to be thin, thin-ideal internalization, body dissatisfaction, dieting, and negative affect are risk factors for bulimic pathology and provide support for the dual-pathway model. PMID- 11261387 TI - The relation between exposure to violence and social information processing among incarcerated adolescents. AB - Combining evidence from social learning theory with reports of the association between community violence exposure and aggressive behavior development, the authors examined the link between specific characteristics of violence exposure and social information-processing mechanisms (N. R. Crick & K. A. Dodge, 1994; K. A. Dodge, 1980, 1986) in a sample of highly aggressive, incarcerated adolescent boys (N = 110). Results demonstrated that victimization by severe violence was significantly related to approval of aggression as a social response, problems with the interpretation of social cues, and maladaptive social goals. Witnessing severe violence, in contrast, was related to perceived positive outcomes for the use of aggression. These data suggest the importance of examining the severity and modality of exposure to community violence for understanding patterns of social-cognitive functioning among adolescents exposed to violence. PMID- 11261388 TI - Patterns of recovery of autonomic dysfunctions and neurocognitive deficits in schizophrenics after acute psychotic episodes. AB - In this study, the authors aimed to identify patterns of autonomic dysfunction and neurocognitive deficit recovery. The authors performed laboratory assessments on 66 patients with schizophrenia immediately after an acute psychotic episode and 6, 12, and 18 months later. Shortly after the psychotic episode, the patients displayed cardiovascular hyperarousal at rest, cardiovascular and electrodermal hyporeactivity during 2 Continuous Performance Tasks (CPTs) and deficits in 2 behavioral CPT measures (i.e., reaction time and omission error rate) compared with 29 normal controls. In the subsequent postpsychotic course, changes indicative of a process of recovery occurred in all measurement areas, although with regard to autonomic hyporeactivity amelioration was limited to a subgroup of schizophrenics with complete and persistent symptomatic remission. Neurocognitive improvement in CPTs did not appear to depend on unimpaired autonomic reactivity mechanisms. PMID- 11261389 TI - A process-oriented approach for averting confounds resulting from general performance deficiencies in schizophrenia. AB - The most pervasive and least well-addressed problem in cognitive studies of schizophrenia is the propensity of schizophrenia patients to show inferior performance on a variety of cognitive tasks. Consequently, apparent specific cognitive abnormalities may actually reflect the interaction of task discriminating power with generalized deficit. L. J. Chapman and J. P. Chapman (1973a) suggested psychometric approaches for eliminating such artifactual group differences. Unfortunately, their solution neglects important issues of process specification and does not provide a viable strategy for process-oriented investigators. Psychometric remediation of artifactual Group x Task interactions inevitably confounds the processes being measured, resulting in theoretically ambiguous findings. Moreover, evidence that changes in measurement reliability can both increase and decrease group discrimination challenges a basic underlying assumption of the Chapmans' matching solution. This article presents a process oriented approach to solving this problem in schizophrenia research. PMID- 11261390 TI - Directed forgetting of trauma cues in adults reporting repressed or recovered memories of childhood sexual abuse. AB - An item-cuing directed forgetting task was used to investigate whether women reporting repressed (n = 13) or recovered (n = 13) memories of childhood sexual abuse (CSA) exhibit an avoidant encoding style (and resultant impaired memory) for trauma cues relative to women reporting no CSA experience (n = 15). All participants viewed intermixed trauma (e.g., molested), positive (e.g., confident), and categorized neutral (e.g., mailbox) words on a computer screen and were instructed either to remember or to forget each word. The results provided no support for the hypothesis that people reporting either repressed or recovered memories of CSA are especially adept at forgetting words related to trauma. These groups recalled words they were instructed to remember more often than words they were instructed to forget regardless of whether they were trauma related. PMID- 11261391 TI - A comparison of the cognitive deficits in reading disability and attention deficit/hyperactivity disorder. AB - This study used a nonreferred sample of twins to contrast the performance of individuals with reading disability (RD; n = 93), attention-deficit/hyperactivity disorder (ADHD; n = 52), RD and ADHD (n = 48), and neither RD nor ADHD (n = 121) on measures of phoneme awareness (PA) and executive functioning (EF). Exploratory factor analysis of the EF measures yielded underlying factors of working memory, inhibition, and set shifting. Results revealed that ADHD was associated with inhibition deficits, whereas RD was associated with significant deficits on measures of PA and verbal working memory. The RD + ADHD group was most impaired on virtually all measures, providing evidence against the phenocopy hypothesis as an explanation for comorbidity between RD and ADHD. PMID- 11261393 TI - Homophobia and physical aggression toward homosexual and heterosexual individuals. AB - This study examined the relationship between homophobia (defined as self-reported negative affect, avoidance, and aggression toward homosexuals) and homosexual aggression. Self-identified heterosexual college men were assigned to homophobic (n = 26) and nonhomophobic (n = 26) groups on the basis of their scores on the Homophobia Scale (HS; L. W. Wright, H. E. Adams, & J. A. Bernat, 1999). Physical aggression was examined by having participants administer shocks to a fictitious opponent during a competitive reaction time (RT) task under the impression that the study was examining the relationship between sexually explicit material and RT. Participants were exposed to a male homosexual erotic videotape, their affective reactions were assessed, and they then competed in the RT task against either a heterosexual or a homosexual opponent. The homophobic group reported significantly more negative affect, anxiety, and anger-hostility after watching the homosexual erotic videotape than did the nonhomophobic group. Additionally, the homophobic group was significantly more aggressive toward the homosexual opponent, but the groups did not differ in aggression toward the heterosexual opponent. PMID- 11261392 TI - A genetic association with the development of alcohol and other substance use behavior in Asian Americans. AB - Studies of Asian adults have found that alcohol use and alcohol dependence are related to variation in the aldehyde dehydrogenase (ALDH2) gene. To investigate the association of ALDH2 with the development of drug involvement, the authors analyzed retrospective information about the onset and regular use of alcohol and other substances as reported by 180 Asian American college students. Possession of an ALDH2*2 allele was not related to initiation of alcohol use or having ever been intoxicated, but individuals with ALDH2*2 alleles were less likely to be regular drinkers, were less likely to have engaged in a binge-drinking episode, reported a lower number of maximum drinks consumed in a 24-hr period, and were less likely to have used tobacco regularly than those without this genetic variant. These findings suggest that ALDH2 is associated with the development of not only alcohol-related behavior but other substance use behavior as well. PMID- 11261394 TI - Primary and secondary hypohedonia. AB - Having shown taxometrically that there exists a hypohedonic schizotypal taxon in a college population, J. J. Blanchard, S. W. Gangestad, S. A. Brown, and W. P. Horan (2000) suggested that P. E. Meehl erred in revising his 1962 theory by postulating a normal-range individual differences variable of hedonic capacity that potentiates schizotypy into schizophrenia. The aversive drift and secondary anhedonia of Meehl's theory imply that the schizotypal taxon will generate hypohedonic taxonicity in an adult population. Psychometrically measurable hedonic disposition (as distinguished from genetic primary hedonic capacity) is "dragged along" by the schizogene, especially in the social domain. To choose between causal interpretations, it could be ascertained whether the schizotypal anhedonic taxon is composed of individuals who are schizotaxic on the basis of psychophysiological, cognitive, and soft neurologic indicators. PMID- 11261395 TI - Affective reactivity of language symptoms, startle responding, and inhibition in schizophrenia. AB - The speech of some schizophrenia patients becomes markedly more disordered when negative affect is aroused. The authors tested associations between affective reactivity of speech and responsiveness and inhibition on an acoustic startle task in a sample of 27 outpatients. Patients whose language was reactive to negative affect showed significantly higher initial startle amplitudes than those whose language was not reactive. However, they also showed greater habituation to repeated startle stimuli over trials, even after differences in initial amplitudes were controlled statistically. These findings suggest that affective reactivity of speech is associated with higher initial startle responsiveness but also with greater habituation and, conversely, that patients who are relatively nonreactive to excitatory affective and sensory stimuli are also less reactive to inhibitory input. PMID- 11261396 TI - Commentary on two articles concerning generalized and specific cognitive deficits. AB - This is a commentary on essays in this journal issue by M. E. Strauss (2001) and R. A. Knight and S. M. Silverstein (2001) on research methodology for studying cognitive deficits. Concentrating mostly on Knight and Silverstein's article, the authors review the psychometric issues in the matched-task design, analyze Knight and Silverstein's "process-oriented" objections to that design, and scrutinize their methods for studying cognitive deficits, examining 2 of their empirical studies as examples of those methods. PMID- 11261397 TI - Methodology for identifying specific psychological deficits: introduction to the special section. AB - A long-standing problem in psychopathology research is establishing that patterns of differences between groups reflects differences of underlying constructs and not artifacts of research design, measurement, or analysis. This introduction provides a context for 4 articles that consider several controversial issues regarding this problem and presents a precis. Although these articles focus on schizophrenia, it is noted how the issues are of more general relevance to psychopathology researchers. PMID- 11261398 TI - Misunderstanding analysis of covariance. AB - Despite numerous technical treatments in many venues, analysis of covariance (ANCOVA) remains a widely misused approach to dealing with substantive group differences on potential covariates, particularly in psychopathology research. Published articles reach unfounded conclusions, and some statistics texts neglect the issue. The problem with ANCOVA in such cases is reviewed. In many cases, there is no means of achieving the superficially appealing goal of "correcting" or "controlling for" real group differences on a potential covariate. In hopes of curtailing misuse of ANCOVA and promoting appropriate use, a nontechnical discussion is provided, emphasizing a substantive confound rarely articulated in textbooks and other general presentations, to complement the mathematical critiques already available. Some alternatives are discussed for contexts in which ANCOVA is inappropriate or questionable. PMID- 11261399 TI - Reliability of DSM-IV anxiety and mood disorders: implications for the classification of emotional disorders. AB - The reliability of current and lifetime Diagnostic and Statistical Manual of Mental Disorders (4th ed.; DSM-IV; American Psychiatric Association, 1994) anxiety and mood disorders was examined in 362 outpatients who underwent 2 independent administrations of the Anxiety Disorders Interview Schedule for DSM IV: Lifetime version (ADIS-IV-L). Good to excellent reliability was obtained for the majority of DSM-IV categories. For many disorders, a common source of unreliability was disagreements on whether constituent symptoms were sufficient in number, severity, or duration to meet. DSM-IV diagnostic criteria. These analyses also highlighted potential boundary problems for some disorders (e.g., generalized anxiety disorder and major depressive disorder). Analyses of ADIS-IV L clinical ratings (0-8 scales) indicated favorable interrater agreement for the dimensional features of DSM-IV anxiety and mood disorders. The findings are discussed in regard to their implications for the classification of emotional disorders. PMID- 11261400 TI - Is smoking automatic? Demands of smoking behavior on attentional resources. AB - The role of attention in the production of smoking behavior was investigated. Experienced and novice smokers were asked to perform a reaction time (RT) task under 4 conditions: while smoking (smoking), while mimicking all aspects of smoking except inhaling (pseudosmoking), while simply holding a cigarette (holding), and while not smoking (baseline). Experienced smokers' RTs increased during the pseudosmoking and holding conditions compared with baseline but did not differ between the smoking and baseline conditions, suggesting that attentional resources were not required for typical smoking behavior but were required to alter or inhibit smoking behavior. Novice smokers' RTs were slower during both the smoking and pseudosmoking conditions but not the holding condition, suggesting that novice smokers require the use of resources to smoke. Experiment 2 demonstrated that the differences in RT across conditions could not be explained by differences in urges. PMID- 11261401 TI - Demonstrating specific cognitive deficits: a psychometric perspective. AB - Cognitive deficits associated with psychopathology often do not occur in isolation. Consequently, identifying a specific deficit in a disorder requires comparing the magnitude of group differences on theoretically relevant measures with those on control tasks measuring other constructs. L. J. Chapman and J. P. Chapman (1973) noted that common forms of such Group x Task interactions are theoretically ambiguous unless performance measures have comparable discriminating power. The principles of psychometric matching for discriminating power developed in the Chapmans' research program are reviewed, and both criticisms and alternative psychometric approaches are evaluated. Psychometric matching can be mindful of threats to the construct validity of measures and frequently remains methodologically necessary. Otherwise, interactions involving measures that vary in sensitivity to individual differences may be misinterpreted as evidence for specific deficits. PMID- 11261402 TI - Increased semantic and repetition priming in schizophrenic patients. AB - Positive and negative priming (PP and NP) in schizophrenia were studied with a lexical-decision task. Probe words, presented 800 ms after the response to the prime (containing a word and a nonword), were either identical to, semantically related to, or unrelated to the prime target word (PP) or to the prime distractor word (NP). Schizophrenic patients displayed stronger semantic and repetition PP than controls after controlling for their slower responses. Significant NP was observed in both groups for word repetition only. The PP findings contrast with results from studies with similar prime-probe intervals but without prime responses. It is proposed that schizophrenic patients, because of impaired (controlled) processes of response selection, strongly benefit from (or rely on) the automatic retrieval of processing episodes containing response information. Related findings indicating automatic response facilitation in schizophrenia are discussed. PMID- 11261403 TI - The effect of practice on recall of emotional information in individuals with generalized social phobia. AB - Understanding memory processes in social anxiety is important because these individuals often report negative memories of anxiety-provoking situations and because of the recent emphasis on learning and memory in models of anxiety. The authors examined the effect of learning on memory for negative social, positive social, and nonsocial information using the retrieval-induced forgetting paradigm in individuals with generalized social phobia (GSPs) and in nonanxious controls (NACs). Words were presented in 1 of 3 practice conditions: practiced words from a practiced category, unpracticed words from a practiced category, and unpracticed words from an unpracticed category. GSPs and NACs showed the same patterns of memory for practice categories for positive social and nonsocial words. However, for negative social words, GSPs benefited less from practice and were hurt less from the effect of practicing competing negative social information than were NACs. This pattern of processing may hamper GSPs' learning of, and habituation to, negative social information. PMID- 11261404 TI - Mood as input and catastrophic worrying. AB - The authors describe 3 experiments investigating a "mood-as-input" approach to understanding catastrophic worrying. Experiment 1 found that induced negative mood increased the number of steps emitted in both a catastrophizing interview procedure and a positive iteration task. Experiment 2 found that the number of items that worriers emitted in an iterative item generation task was dependent on the stop rules specified by the procedure. Experiment 3 found that manipulating the stop rules for catastrophizing had differential effects on worriers and nonworriers, depending on the nature of the stop rules specified. These results suggest that mood provides information about continuing or terminating the catastrophizing process that is interpreted in the context of the stop rules for the task. It is argued that the mood-as-input hypothesis accounts for the facts of exacerbated catastrophizing in worriers better than explanations couched in terms of either mood congruency effects or worriers possessing a generalized perseverative iterative style. PMID- 11261405 TI - Control-related beliefs and depressive symptoms in clinic-referred children and adolescents: developmental differences and model specificity. AB - The contingency-competence-control (CCC) model links contingency and competence beliefs to perceived control and, in turn, to depression. However, a developmental perspective suggests that noncontingency may be too abstract a concept to be directly tied to depression before adolescence. We tested the CCC model and this developmental notion, using structural equation modeling, with 360 clinic-referred 8- to 17-year-olds. The CCC model fit the data well for the full sample accounting for 46% of the variance in depression. Separate analyses by age group placed perceived contingency in the best-fit model for adolescents (ages 12 17 years) but not for children (8-11 years). This suggests that abstract cause effect concepts may have more direct affective impact after the cognitive changes of adolescence (e.g., formal operations) than before. Finally, the CCC model accounted for much more variance in depression than conduct problems, suggesting diagnostic specificity. PMID- 11261407 TI - Optimizing quality of care and cost effectiveness in the treatment of overactive bladder. PMID- 11261406 TI - OSHA's ergonomics final rule and healthcare: what's the motivation? PMID- 11261408 TI - Overactive bladder: optimizing quality of care. AB - Overactive bladder (OAB), the symptom complex of urinary urgency and frequency with or without urge incontinence, affects the lives of millions of Americans. In recent years, more successful treatment options have emerged as advances have been made in understanding the pathophysiologic processes underlying OAB symptoms. However, because most therapeutic modalities for OAB are aimed at symptom resolution, rather than the treatment of distinct pathologic entities, a basic evaluation is required for all patients to establish whether existing (and treatable) pathologic processes are present. In the absence of these processes, symptom relief is both the objective and the outcome used to judge the efficacy of a specific modality. The type of therapy recommended for OAB may depend on several factors including age, existing behavioral patterns, estrogen status, degree of motivation, environmental surroundings, presence of other coexisting urinary symptoms, family support, and patient expectations. This article focuses on methods of identifying patients with OAB, and the role of developing strategies in treating this common disorder. PMID- 11261410 TI - What this CEO didn't know about his cholesterol almost killed him. PMID- 11261409 TI - Cost effectiveness and quality of life considerations in the treatment of patients with overactive bladder. AB - This article summarizes the quality of life (QOL), cost of illness, and cost effectiveness considerations in the treatment and management of patients with overactive bladder (OAB). Most cost studies have focused primarily on urinary incontinence, which is only one possible symptom of OAB. Prevalence rates of urge and mixed incontinence in the United States ranged from 3% to 8% and 5% to 37%, respectively. The highest prevalence was found in geriatric and psychogeriatric populations, where 40% and 90%, respectively, were classified as incontinent. In patients with OAB, all aspects of QOL can be compromised including physical, social, occupational, domestic, and sexual activities, and associated costs can be substantial. Oxybutynin has been the mainstay of pharmacotherapy for OAB but its more frequent side effects (including dry mouth) may deter patients from full compliance with treatment. Tolterodine, a newer antimuscarinic drug, has proven safe and effective in the treatment of OAB, with fewer side effects and better tolerability than existing agents. Cost effectiveness reports are reviewed. Further research on OAB is needed to characterize the disease process and identify risk factors. PMID- 11261411 TI - Debunking Oxford. PMID- 11261412 TI - Future challenges in pediatric cardiology:the increasing role of medical informatics. PMID- 11261413 TI - When will the MEK inherit the ERK? Acronym alphabet soup. PMID- 11261414 TI - Pharmacogenetics and pharmacogenomics: tailored drug therapy for the 21st century. PMID- 11261415 TI - P2 receptors meet the immune system. PMID- 11261416 TI - Thyroid disorders--an update. PMID- 11261417 TI - Neuromuscular disorders: gene location. PMID- 11261418 TI - Mitochondrial encephalomyopathies: gene mutation. PMID- 11261419 TI - Determination of capsaicin and dihydrocapsaicin in air in a pickle and pepper processing plant. AB - A sampling and analytical method has been developed for measurement of capsaicin and dihydrocapsaicin in air. This method is applicable to measurement of the capsaicinoids in air in pepper processing plants and involves air sampling with a 13-mm glass fiber filter, recovery of the sample with 2 mL of acetonitrile, filtration of the solution, and analysis by high performance liquid chromatography with fluorescence detection. Excitation and emission wavelengths of the detector were 281 and 312 nm, respectively. Average recoveries were 98 to 104% after fortification of glass fiber filters with 0.13- to 2.9-microg quantities of capsaicin. Average recoveries of dihydrocapsaicin were 93 to 99% after fortification of glass fiber filters with 0.11- to 3.0-microg quantities. Detection limits of capsaicin and dihydrocapsaicin were 0.015 and 0.02 microg per sample, respectively. This method was used successfully for determining air concentrations of capsaicin and dihydrocapsaicin in a health hazard evaluation at a large pickle and pepper processing plant. An interesting phenomenon was the fact that the ratio of capsaicin to dihydrocapsaicin in each of the largest air samples was in the range of 0.3:1 to 0.5:1. Generally, capsaicin is the capsaicinoid that occurs in Capsicum fruit in the greatest relative abundance. PMID- 11261420 TI - Detection and sequencing analysis of IL-18 expression in J(6-1_ leukemic cells. PMID- 11261421 TI - Transthyretin-associated neuropathic amyloidosis. Pathogenesis and treatment. AB - Hereditary amyloidoses form a clinically and genetically heterogeneous group of autosomal dominantly inherited diseases characterized by the deposit of insoluble protein fibrils in the extracellular matrix. They typically present with polyneuropathy, carpal tunnel syndrome, autonomic insufficiency, and cardiomyopathy and gastrointestinal features, occasionally accompanied by vitreous opacities and renal insufficiency. Other phenotypes are characterized by nephropathy, gastric ulcers, cranial nerve dysfunction, and corneal lattice dystrophy. Rarely, involvement of the leptomeningeal or cerebral structures dominates the clinical picture. The age at onset is as early as 17 and as late as 78 years. The basic constituents of amyloid fibers are physiologic proteins that have become amyloidogenic through genetically determined conformation changes. Mutated transthyretin (TTR), formerly termed prealbumin, is the most frequent offender in hereditary amyloidosis. Orthotopic liver transplantation (OLT) stops the progression of the disease, which is otherwise invariably fatal, by removing the main production site of amyloidogenic protein. The indications for OLT and its success depend on the grade of cardiovascular and autonomic dysfunction at the time of surgery, age, comorbidity, and type of mutation. Alternative treatment modalities with drugs stabilizing the native tetrameric conformation of TTR and inhibiting fibril formation are currently being studied. PMID- 11261422 TI - Vagus nerve stimulation and drug reduction. AB - The authors prospectively assessed drug reduction and patient satisfaction in 21 patients using vagus nerve stimulation (VNS) for refractory epilepsy and compared results to a case-matched control group with a mean follow-up of 13.2 months. Significant antiepileptic drug (AED) reduction occurred in 9/21 (42.9%) of VNS patients averaging 0.43 AED/patient, with dose reduction in four patients (19.0%). For 12/21 (57.1%) patients not reducing AED, dose reduction occurred in 6/21 (28.6%). Drug and dose reduction of AED is possible in patients using VNS for refractory epilepsy without loss of seizure control and with improved patient satisfaction. PMID- 11261423 TI - Practice parameter: evaluating a first nonfebrile seizure in children: Report of the Quality Standards Subcommittee of the American Academy of Neurology, the Child Neurology Society, and the American Epilepsy Society. PMID- 11261424 TI - Primary care physicians' utilization and perceptions of genetics services. AB - PURPOSE: To document primary care physicians' utilization and perceptions of genetics services. METHODS: A randomized survey of physicians in the Pacific Northwest. RESULTS: The greatest factor prompting a genetics referral was the patient's interest in the evaluation, and the most common reason not to obtain a consultation was the perception that it was of no benefit to the patient. Genetics consultation was rarely sought for a family history of cancer or for deafness, polycystic kidney disease, or congenital heart disease. Even when uncertain about relative risk, physicians usually counseled a patient themselves rather than referring to a specialist. CONCLUSION: Primary care physicians need more education about the genetic component of many diseases to provide directly and to refer appropriately for genetics services. PMID- 11261425 TI - Impact of prenatal screening on the birth status of fetuses with Down syndrome at an urban hospital, 1972-1994. AB - PURPOSE: This hospital-based study has determined the change over time (1972-1974 and 1979-1994) in the methods of prenatal detection of fetuses with Down syndrome and the impact of elective termination on the portion that were liveborn. METHODS: Using a malformations surveillance program, all 265 affected fetuses and infants were identified among 161,560 births and elective terminations during the aforementioned period at Brigham and Women's Hospital in Boston, MA. RESULTS: From 1972 to 1974, Down syndrome was not diagnosed in any affected infants prenatally. In the early 1980s, amniocentesis was the primary method of diagnosis; later, maternal serum screening and ultrasonography were as likely to be the first method of detection. Most couples (86%) elected to terminate pregnancies with affected fetuses. CONCLUSIONS: The effect of prenatal detection and the choice of elective termination produced a significant decrease, between 1972 and 1994, in the portion of fetuses with Down syndrome who were liveborn. PMID- 11261426 TI - Fluorescence in situ hybridization analysis with LIS1 specific probes reveals a high deletion mutation rate in isolated lissencephaly sequence. AB - PURPOSE: Recent revision of the lissencephaly critical region on chromosome 17p13.3 and confirmation of LIS1 as the causative gene for classical lissencephaly has allowed the development and application of fluorescence in situ hybridization (FISH) probes corresponding directly to this gene. METHOD: We have analyzed patients with isolated lissencephaly sequence (ILS) by FISH with probes at D17S379, an anonymous locus distal to LIS1, and with LIS1 specific probes. RESULTS: In 110 patients with ILS, a deletion at D17S379 was detected in 23.6%. Of those patients without a deletion, 32 were available for further study with LIS1 probes. Deletions were found in eight additional individuals. CONCLUSION: The overall deletion mutation rate detectable by FISH with LIS1 probes is approximately 40%. This rate is significantly higher than the deletion rate observed at D17S379. This indicates that FISH studies using probes specific to LIS1 should be undertaken as the initial diagnostic assay for the evaluation of patients with ILS, and the high frequency of deletions raises the possibility of "hotspots" for chromosome breakage in this region. PMID- 11261427 TI - Defective urinary carnitine transport in heterozygotes for primary carnitine deficiency. AB - PURPOSE: Primary carnitine deficiency is an autosomal recessive disorder caused by defective carnitine transport and manifests as nonketotic hypoglycemia or skeletal or heart myopathy. METHODS: To define the mechanisms producing partially reduced plasma carnitine levels in the parents of affected patients, we examined carnitine transport in vivo and in the fibroblasts of a new patient and his heterozygous parents. RESULTS: Kinetic analysis of carnitine transport in fibroblasts revealed an absence of saturable carnitine transport in the proband's cells and a partially impaired carnitine transport in fibroblasts from both parents, whose cells retained normal Km values toward carnitine (6-9 microM) but reduced Vmax. At steady state, normal fibroblasts accumulated carnitine to a concentration that was up to 80 times the extracellular value (0.5 microM). By contrast, cells from the proband had minimal carnitine accumulation, and cells from both parents had intermediate values of carnitine accumulation. Plasma carnitine levels were slightly below normal in both heterozygous, yet clinically normal, parents and in the paternal grandfather and the maternal grandmother. To define the mechanism producing partially decreased carnitine levels, we studied urinary carnitine losses in heterozygous parents compared with controls. Urinary losses increased linearly (P < 0.05) with plasma carnitine levels in normal controls. When urinary carnitine losses were normalized to plasma carnitine levels, a significant difference was observed between controls and heterozygous individuals (P < 0.01). CONCLUSIONS: These results indicate that fibroblasts from heterozygotes for primary carnitine deficiency have a decreased capacity to accumulate carnitine and that heterozygotes have increased urinary losses, which may contribute to their reduced plasma carnitine levels. PMID- 11261428 TI - Clinical applications of comparative genomic hybridization. AB - PURPOSE: Comparative genomic hybridization (CGH) is a powerful DNA-based cytogenetic technique that allows the entire genome to be scanned for chromosomal imbalances without requiring the sample material to be mitotically active. During the past 2 years we received many requests from various medical centers around the country to use CGH to resolve the identity of aberrant chromosomal material. METHODS: We report the use of CGH for the evaluation of 12 clinical postnatal cases in which traditional cytogenetic analysis yielded ambiguous results. This series consisted of five marker chromosomes, five unbalanced translocations, and two intrachromosomal duplications. RESULTS: Identification and characterization of the additional unknown chromosomal material was achieved with use of CGH. All CGH findings were validated by traditional fluorescence in situ hybridization and other specialized staining techniques. CONCLUSLONS: These results demonstrate the effective use of CGH as a focused, single-step method for the identification of chromosomal material of unknown origin. PMID- 11261429 TI - The molecular biology of galactosemia. AB - Classic galactosemia is an autosomal recessive disorder caused by the deficiency of galactose 1-phosphate uridyltransferase (GALT). Although the potentially lethal, neonatal hepatotoxic syndrome is prevented by newborn screening and galactose restriction, long-term outcome for older patients with galactosemia remains problematic. After the cloning and sequencing of the GALT gene, more than 130 mutations in the GALT gene have been associated with GALT deficiency; this review relates them to function and clinical outcome. Two common mutations, Q188R and K285N, account for more than 70% of G alleles in the white population and are associated with classic galactosemia and impaired GALT function. In the black population, S135L accounts for 62% of the alleles causing galactosemia and is associated with good outcomes. A large 5 kb deletion in the GALT gene is found in Ashkenazim Jews. The Duarte galactosemia variant is caused by N314D. Homozygosity for N314D reduces GALT activity to 50%. When either E203K or a 1721C-->T transition (Los Angeles variant) are present in cis with N314D, GALT activity reverts to normal. In this review, we discuss the structural biology of these mutations as they affect both the GALT enzyme and patient outcome. PMID- 11261430 TI - Phenotypic differerencss in African Americans with Prader-Willi syndrome. AB - We report on 10 African Americans with Prader-Willi syndrome whose features differ from those of white patients with this condition. Growth is less affected, hand and foot lengths usually are normal, and the facies are atypical; this may lead to underdiagnosis in this population. We encourage clinicians to recognize these phenotypic differences so that diagnosis is not overlooked. PMID- 11261431 TI - Clinical objectives in medical genetics for undergraduate medical students. Association of Professors of Human Genetics, Clinical Objectives Task Force. PMID- 11261432 TI - Medical genetics: end of the beginning or beginning of the end? PMID- 11261433 TI - Statement on folic acid: fortification and supplementation. PMID- 11261434 TI - The ACMG CYTO2000 subcommittee? PMID- 11261435 TI - From astronomy to biology: could amateurs have a role in research? PMID- 11261436 TI - Uterus didelphys with obstructed hemivagina and ipsilateral renal agenesis. A case report. AB - BACKGROUND: Uterus didelphys with obstructed hemivagina and ipsilateral renal agenesis usually presents after menarche with progressive abdominal pain during menses secondary to hematocolpos. We describe a case with the unique presentation of rectal pain and constipation. CASE: A 13-year-old girl presented to the emergency department complaining of lower abdominal and rectal pain and constipation of two weeks' duration. Pelvic ultrasound, physical examination and laparoscopic findings established a diagnosis of hematometracolpos secondary to uterus didelphys with unilateral imperforate hemivagina. An incision in the vaginal septum allowed drainage of the hematocolpos, providing relief of the patient's symptoms. CONCLUSION: Uterus didelphys with unilateral imperforate hemivagina and ipsilateral renal agenesis may present with apparent gastrointestinal symptoms. With increased awareness of this problem, timely diagnosis may be achieved. PMID- 11261438 TI - A comparison of inhaled fluticasone and oral prednisone for children with severe acute asthma. PMID- 11261439 TI - [Advertising in the Revista de Investigacion Clinica]. PMID- 11261440 TI - [Advertising]. PMID- 11261441 TI - [Comment...]. PMID- 11261442 TI - Magnetic resonance spectroscopy in AD. AB - Proton MR spectroscopy (MRS) studies have found both decreased N-acetylaspartate (NAA) and increased myo-inositol in the occipital, temporal, parietal, and frontal regions of patients with AD, even at the early stages of the disease. This diffuse NAA decline is independent of regional atrophy and probably reflects a decrease in neurocellular viability. Reports of such metabolite changes are now emerging in the mild cognitive impairment prodrome and in investigation of the medial temporal lobe. In vivo quantitation of neural choline in AD has been inconclusive because of poor test-retest repeatability. Less robust evidence using phosphorous MRS has shown significant phosphocreatine decline and increments in the cell membrane phosphomonoesters in the early, and possibly asymptomatic, stages of the disease. These phosphorous metabolite disturbances normalize with disease progression. Phosphodiester concentration has been found to correlate strongly with AD plaque counts. MRS of AD has therefore introduced new pathophysiologic speculations. Studies of automated MRS for AD diagnosis have reported high sensitivity and moderate specificity, but are yet to test prospective samples and should be extended to include at least two MRS regions of interest. MRS has promise for predicting cognitive status and monitoring pharmacologic efficacy, and can assess cortical and subcortical neurochemical change. PMID- 11261443 TI - Bilateral middle cerebellar peduncle infarction caused by traumatic vertebral artery dissection. PMID- 11261444 TI - Flushing and syncopal episode in a 47-year-old female. PMID- 11261445 TI - Congenital rubella and interstitial pneumonitis. PMID- 11261446 TI - Successful thrombolysis of inferior vena cava thrombolysis in a preterm neonate. PMID- 11261447 TI - Thrombolysis in neonates. PMID- 11261448 TI - Severe iron deficiency anemia and asymptomatic nodular gastroduodenitis: an uncommon presentation of Helicobacter pylori infection in a child. PMID- 11261449 TI - gamma-Sarcoglycanopathy in two Palestinian-American siblings. PMID- 11261450 TI - Pediatric methodone poisoning revisted. PMID- 11261451 TI - Predictors of multiple seizures in a cohort of children prospectively followed from th time of their first unprovoked seizure. PMID- 11261452 TI - Behavioral and cognitive status in school-aged children with a history of failure to thrive during early childhood. AB - Twenty-seven school children (aged 8-12 years) earlier diagnosed with nonorganic failure to thrive (FTT) were compared with a normal socioeconomically matched control group (N=17) on current height and weight parameters as well as cognitive, achievement, and behavioral measures from the Child Behavior Checklist (CBCL). The former FTT children were, on average, smaller, less cognitively able, and more behaviorally disturbed than the control children and national normative samples. Sixty percent of former FTT children were below the 20th percentile in height and 48% were below the 20th percentile in weight; 52% had IQs below 80 and 30% had reading standard scores below 80; 48% had clinically adverse attention ratings and 30% had clinically adverse aggression ratings on the CBCL. Within the FTT sample, however, there were no significant associations between current growth measures and cognitive/achievement outcome measures. Mothers' IQs provided the strongest prediction of the FTT children's reading scores. The mothers of the FTT children had not achieved as high levels of education as the mothers of the control children, and more of them were single parents. Early growth problems put children at high risk for multiple adverse sequelae in middle childhood, especially if mothers are poorly educated. Careful ongoing follow-up of such children by pediatricians is encouraged. PMID- 11261453 TI - An examination of the unintended consequences of the rule-out sepsis evaluation: a parental perspective. AB - In order to document unintended consequences of the "rule-out" sepsis (ROS) evaluation, a survey of parents of infants who had undergone such an evaluation at Primary Children's Medical Center in 1997 was conducted. Sixty parents were interviewed. Parental perceptions of the sepsis evaluation and its impact on their families were recorded. Specific data evaluated included parental anxiety, impact on breastfeeding, perceived complications, financial stress, and parental preferences. The majority of parents found the ROS evaluation very stressful. Parental perception of illness increased significantly after being told the infant would require an ROS evaluation, with nearly 30% of parents, after speaking with a physician, believing their infant might die. Breastfeeding problems were reported by 36% of the mothers. Iatrogenic complications were reported by 33%. Although all infants were covered by some form of insurance, 43% of parents reported financial stress. Forty-two percent of parents would have preferred to be treated at home and all parents would prefer an evaluation that could be accomplished in 24 hours. We conclude that unintended consequences of the ROS evaluation included excessive parental anxiety, cessation of breastfeeding, iatrogenic complications, and financial stress. Suggestions to decrease these adverse consequences are given. PMID- 11261454 TI - Effectiveness of inhaled corticosteroids in controlling acute asthma exacerbations in children at home. AB - Many clinicians advise their patients to increase the dose of inhaled corticosteroids during acute asthma exacerbations, without strong clinical evidence supporting this treatment. This study investigates the effectiveness of inhaled corticosteroids in controlling acute asthma exacerbations in children at home. The study population consisted of children with mild intermittent, mild and moderate persistent asthma aged 1 to 14 years who were treated in our outpatient clinic with inhaled budesonide for 1 year. After participating in an asthma education session, the parents were instructed to initiate treatment with inhaled budesonide at the first signs of asthma exacerbation, starting with 200 to 400 microg budesonide, in combination with beta-2 agonists 4 times a day and followed by a decrease in the dose in 4 to 8 days. Asthma status and peak expiratory flow rates were measured in the 3 monthly follow-up visits. Only children who complied with the treatment regimen and came for follow-up visits regularly were included in the final analysis. One hundred fifty children used our treatment protocol with inhaled budesonide to control their asthma attacks. Clinical improvement of asthma symptoms was achieved after a mean of 1.8 +/- 0.7 days from the beginning of treatment. The parents were able to control 94% of the 1,061 episodes of asthma exacerbation occurring during a cumulative follow-up period of 239 years. In the 3-month period before enrollment, 101 children (67%) had used oral corticosteroids to control their asthma attacks and 50 (33%) were hospitalized. During the entire follow-up period, only 11 children (7%) used oral corticosteroids, and none of the children were hospitalized. The present study demonstrates that children with asthma can control their exacerbations at home using inhaled corticosteroids (budesonide). Treatment, starting with relatively high doses followed by a rapid reduction in dose over 4-8 days, resulted in a decrease in the use of oral steroids and in hospitalization. To achieve good results, patient compliance is essential. PMID- 11261455 TI - Medical causes of pneumomediastinum in children. AB - The purpose of this study was to determine the possible causes, clinical findings, and associated complications of pneumomediastinum in children. Medical records from January 1985 to December 1994 were retrospectively reviewed at Children's Hospital Medical Center of Akron using International Classification of Diseases, ninth revision, codes to identify cases of pneumomediastinum. The medical causes, nontraumatic and noniatrogenic, of pneumomediastinum were studied; intubated or trauma patients and patients having undergone procedures were excluded. Neonates were also excluded. Twenty-nine cases of pneumomediastinum were identified. Two patients (7%) had recurrent pneumomediastinum. Only the first episode of pneumomediastinum was included in the data analysis. Twenty males (69%) and nine females (31%) were affected. The most common medical causes of pneumomediastinum were asthma exacerbations (17/59%) and infections (8/28%). Over the 10-year period, the prevalence of pneumomediastinum in children with asthma exacerbations was 0.2% (21/10,472); 1% (1/126) in children with airway foreign bodies and 0.2% (1/351) in children with esophageal foreign bodies. The most common signs and symptoms were subcutaneous emphysema (22/76%), sore throat or neck pain (11/38%), and Hamman's crunch (3/10%). The most common complication was pneumothorax with small pneumothoraces in 2 patients (7%) and a tension pneumothorax in 1 asthmatic with recurrent pneumomediastinum. Patients without sore throat or neck pain and patients admitted to the intensive care unit had greater hospital lengths of stay. Pneumomediastinum appears to be uncommon in children. The most common medical causes were asthma and infections. The most common signs and symptoms were subcutaneous emphysema, sore throat or neck pain, and Hamman's crunch. The most common complication was pneumothorax. The clinical significance of pneumomediastinum is its cause and association with significant complications. PMID- 11261457 TI - Chronic relapsing pancreatitis. PMID- 11261456 TI - Telehealth for children with special health care needs: promoting comprehensive systems of care. AB - This study identified ways that telehealth systems are used to enhance service delivery for children with special health care needs. Various health professionals utilize telehealth to provide physical and behavioral health care, to conduct developmental evaluations, and to address family/social concerns. Telehealth is used most for follow-up visits after in-person specialty care and for child evaluation with treatment recommendations to local care providers. Telehealth systems have the potential to increase access to specialized services and to enhance community-based services for rural and underserved children. However, inadequate reimbursement poses a significant barrier to the development of these services. PMID- 11261458 TI - Some issues relevant to establishing a universal newborn hearing screening program. AB - This article describes some of the factors relevant to the establishment of a universal newborn hearing screening (UNHS) program. First, the difficulty in providing precise estimates of test sensitivity and specificity are reviewed. This section is followed by hypothetical estimates of overall programmatic costs, first for a fixed number of babies to be screened and then as a function of the number of babies to be screened in a year. Included in these estimates are the costs for equipment, disposables, personnel, and follow-up testing. These estimates are provided for three different screening protocols: auditory brainstem response (ABR) alone, otoacoustic emission (OAE) alone, and OAE followed by ABR only for those babies who failed the OAE screening. If follow-up costs are not included, it is less expensive to screen newborns with OAEs compared with the other two protocols. However, once follow-up testing is included as part of the program costs and there are at least 400 births per year, procedures in which OAEs are performed first, followed by an ABR on those infants who do not pass the OAE test, result in the lowest costs. Hospitals with as few as 400 births per year should expect per-baby costs not exceeding $30, regardless of which protocol is used. For all three protocols, the unit costs decrease as the number of babies screened increases. The final section describes data from a local UNHS program in which all infants are screened first with an OAE test, followed by an ABR test on infants not passing the OAE screening. Idiosyncratic features to this program are described, including the fact that all screening tests are performed by audiologists, who are paid on a part-time basis, adding cost to the program. Even under these circumstances, the unit cost is under $30. These data lead us to conclude that all infants can be screened in a cost effective manner. PMID- 11261459 TI - Strategies used in feigning hearing loss. AB - Thirty unsophisticated participants with normal hearing were paid to simulate a hearing loss according to their success in "deceiving" the examiner. The behaviors that these "malingerers" manifested are described. A post-examination interview revealed the strategies used by these participants, which may reflect those strategies used by patients who truly attempt to feign a hearing loss. PMID- 11261460 TI - Technology, expectations, and adjustment to hearing loss: predictors of hearing aid outcome. AB - This study examined the influence of technology, demographic factors, and prefitting expectations, attitudes, and adjustment to hearing loss on hearing aid outcome. Clients obtaining new hearing aids completed questionnaires measuring personal adjustment to hearing loss, expectations of and attitudes toward hearing aids, and hearing aid benefit. Eighty-one percent of the 200 subjects completing the prefitting questionnaires returned questionnaires evaluating hearing aid outcome. Factors affecting hearing aid use, overall satisfaction, and benefit were investigated using regression analyses. Higher use time was associated with higher prefitting expectations and greater acceptance of hearing loss. Greater benefit in easy and difficult listening situations was predicted by higher prefitting expectations. Multiple-memory hearing aids produced higher satisfaction. Benefit was greater for multiple-memory, multiple-channel, and wide dynamic range compression aids. Findings were consistent with previous studies showing positive outcomes for newer technologies but also showed that two subjective factors, prefitting hearing aid expectations and acceptance of hearing loss, significantly influenced hearing aid outcome. PMID- 11261461 TI - Aging and contralateral suppression effects on transient evoked otoacoustic emissions. AB - Transient evoked otoacoustic emissions (TEOAEs) were recorded in 30 normal hearing subjects to nonlinear clicks while continuous contralateral broadband noise (CBBN) was presented at 40, 50, 60, and 70 dB HL. Thirty subjects between 20 and 79 years were divided system-atically into six-decade age groups, five subjects per group. All subjects in each group had hearing thresholds of 20 dB HL or better for the test frequencies from 0.25 to 8.0 kHz and normal acoustic immittance findings. The results provide evidence that contralateral suppression at varying levels of CBBN is interactive with age. Except for subjects in the age ranges between 60 and 69 and 70 and 79 years of age, an increase in CBBN from 40 to 70 dB in 10-dB steps resulted in an average increase in suppression from about 0.5 to 3.5 dB SPL. In addition, the contralateral suppression at 60 and 70 dB HL was significantly greater for subjects between 20 and 59 years of age than for subjects between 60 and 79 years of age. PMID- 11261462 TI - Lexical effects on dichotic word recognition in young and elderly listeners. AB - Dichotic listening was evaluated using monosyllabic word pairs that differed in lexical difficulty as defined by the Neighborhood Activation Model of spoken word recognition. Four combinations of lexically EASY and lexically HARD words were evaluated (same pair: EASY-EASY, HARD-HARD; or mixed pair: EASY-HARD, HARD-EASY) in young adult listeners with normal hearing and older adult listeners with mild to-moderate hearing loss. The same-pair data indicated that for all subjects, EASY words were identified correctly more often than HARD words, and recognition performance on words presented to the right ear was better than performance on words presented to the left ear. Overall performance was lower and the right-ear advantage was larger for the older group. The mixed-pair data for the young group revealed that EASY words were recognized more accurately than HARD words, regardless of the ear to which they were presented. For the older adults, the words presented to the right ear were recognized more accurately than were the words presented to the left ear, regardless of the type of word. The efficiency of the processing of stimuli from the left ear is discussed as an explanation of the results for the mixed-pair conditions. PMID- 11261464 TI - Evidence of normal cerebellar control of the vestibulo-ocular reflex (VOR) in children with high-functioning autism. AB - The effect of "tilt-suppression" on post-rotatory vestibular nystagmus was investigated to assess the function of the caudal cerebellar vermis (lobules IX and X, or nodulus and uvula) in 13 school-age children with high-functioning autism (HFA) and 10 normal controls. Tilt-suppression of the vestibulo-ocular reflex (VOR) refers to the decreasing of the duration of post-rotatory vestibular nystagmus that occurs when the head is moved out of the plane in which it was located during the previous sustained constant-velocity rotation. The participant is rotated in a vestibular chair with the head upright and then the head is tilted forward just after the chair stops rotating. Such tilt-suppression is impaired with lesions of the cerebellar nodulus and portions of the uvula. Results show that children with HFA have normal post-rotatory nystasmus with the head upright and normal attenuation of post-rotatory nystagmus induced by head tilt. These behavioral findings suggest that lobules IX and X of the cerebellum are spared in high-functioning autism. PMID- 11261463 TI - Autism and the cerebellum: evidence from tuberous sclerosis. PMID- 11261465 TI - Are infants with autism socially engaged? A study of recent retrospective parental reports. AB - The purpose of this study was to identify the specific aspects of social engagement that distinguish infants with autism from infants of similar age and developmental level who do not have autism. Ten parents of preschoolers with autism and 10 parents of matched children without autism were given a semistructured interview, the Detection of Autism by Infant Sociability Interview (DAISI), which elicits reports on whether 19 aspects of social engagement characteristic of typically developing infants were present at some time during the child's first 24 months. The reports of infants with autism differed from those of the control group on 16 items. Findings suggest that infants with autism have marked limitation in both person-to-person and person-person-object social engagement, in keeping with the theory that autism involves impairments in primary as well as secondary intersubjectivity (Hobson, 1993a). PMID- 11261467 TI - Teaching on-task and on-schedule behaviors to high-functioning children with autism via picture activity schedules. AB - The purpose of this investigation was to evaluate the effectiveness of a two component teaching package (graduated guidance and visual activity schedules) in teaching young students with autism to increase on-task and on-schedule behavior. Four children enrolled in a resource-based classroom in a public elementary school served as participants. An A-B-A-B withdrawal design was used to evaluate the effectiveness of a picture activity schedule on the percentage of intervals scored as on-task and on-schedule. Generalization measures were taken on the percentage of intervals scored as on-task and on-schedule with novel activities. The results of the investigation indicate that (a) student performance rose to criterion levels upon introduction of the graduated guidance procedure, (b) student performance maintained when the picture activity book was available (Book Only) and dropped when the picture activity book was not available (No Book), and (c) student performance generalized to novel activities. The implications of these findings show the importance for future development and use of visual activity schedules to promote the independent functioning of students with autism spectrum disorders in their least restrictive environments. PMID- 11261466 TI - A comparison of video modeling with in vivo modeling for teaching children with autism. AB - The present study was designed to compare the effectiveness of video modeling with in vivo modeling for teaching developmental skills to children with autism. A multiple baseline design across five children and within child across the two modeling conditions (video and in vivo) and across tasks was used. Each child was presented two similar tasks from his or her curriculum; one task was used for the video condition, while the other was used for the in vivo condition. Video modeling consisted of each child watching a videotape of models performing the target behavior, whereas in vivo modeling consisted of the children observing live models perform the target behavior. After the observations, children were tested for acquisition and generalization of target behaviors. Results suggest that video modeling led to faster acquisition of tasks than in vivo modeling and was effective in promoting generalization. Results are discussed in terms of video modeling's motivating and attention maintaining qualities. PMID- 11261468 TI - Teaching conversational skills to children with autism: effect on the development of a theory of mind. AB - This research examined whether children with autism could be trained to improve their conversational skills and whether this led to changes in standard tests of theory of mind (ToM). Three high-functioning children with autism participated in a multiple baseline across participants design. The children were taught how to initiate a conversation, take turns during conversation, listen attentively, maintain a conversation topic, and change a conversation topic appropriately. The children were tested for ToM using False Belief tasks before and after training sessions. Results indicate that the amount of shared interest exhibited by the children with autism during conversation with their caregivers increased during training sessions. The children also made more responses that were appropriate to the context of the conversation. Performance on the False Belief tasks remained constant throughout the study. Results are discussed with respect to the implications of results of performance in standard ToM tasks. PMID- 11261469 TI - System and cost research issues in treatments for people with autistic disorders. AB - Parents of children with autism and pervasive developmental disorder and educational and clinical practitioners providing services to them regularly confront a wide range of service selection and financial decisions that are not as yet effectively addressed by applied research. Relevant systems issues span a very broad range of concerns: (a) systems delivery models and issues (e.g., costs of services, implementation of intensive intervention, and teacher or therapist training); (b) how best to integrate treatments; (c) providing treatment to those with limited monetary resources; (d) cost and cost/benefit analyses; (e) how to educate adult psychiatrists (as well as other practitioners and personnel) regarding autism; and (f) gaps between research and practice. PMID- 11261470 TI - Benefit-cost analysis and autism services: a response to Jacobson and Mulick. PMID- 11261471 TI - Brief report: reduction of inappropriate vocalizations for a child with autism using a self-management treatment program. AB - Self-management procedures that incorporate elements of self-assessment, self recording, and self-reinforcement have reduced stereotypic (i.e., repetitive) behaviors in children with autism in clinical settings. This study examined the effects of a self-management program used to reduce high rates of inappropriate vocalizations (e.g., humming, tongue clucking, perseverative and echolalic words/phrases) in a 12-year-old girl having autism served in a public school classroom. When self-management was applied to inappropriate vocalizations in a multiple-baseline design during leisure, prevocational, and reading tasks, the occurrence of vocalizations decreased. Implications for teaching these procedures in classroom settings are discussed. PMID- 11261472 TI - Brief report: screening tool for autism in two-year-olds (STAT): development and preliminary data. PMID- 11261473 TI - Brief report: cognitive estimation in individuals with pervasive developmental disorders. PMID- 11261474 TI - Pilot study of the behavioral effects of flumazenil in two children with autism. PMID- 11261475 TI - Einstein: brain and behavior. PMID- 11261476 TI - Ludwig Wittgenstein: autism and philosophy. PMID- 11261477 TI - Recognition of faux pas. PMID- 11261478 TI - Ask the editor. My son is an 8-year-old with high-functioning autism/asperger syndrome. PMID- 11261479 TI - The role of embryonic polarities in preimplantation growth and implantation of mammalian embryos. AB - Strong evidence now exists of polar axes associated with various embryological, morphogenetic and developmental events in mammals from oocyte formation to implantation. These axes strictly regulate morphogenesis throughout the pre- and post-implantation stages. They are expressed by gradients in gene products and organelles, control ooplasmic and cytoplasmic rotations coupled with specific cleavage planes, and the imposition of body axes on embryo and fetus. The expression of polarities in preimplantation and implanting embryos, and their relationship with the implantation process, are discussed in this paper. A deeper understanding of such a fundamental aspect of the genetic regulation of the embryo should help to clarify significant aspects of implantation and the clinical evaluation of human embryos in vitro. PMID- 11261480 TI - Blastocyst stage transfer: the real benefits compared with early embryo transfer. AB - With the development of commercially available sequential media it is now possible to grow human embryos to the blastocyst stage without feeder cells. The transfer of blastocysts offers several advantages, the most important being synchronization of the embryos with the uterine endometrium and selection of the best quality embryos with a high implantation potential. This study was conducted to compare the efficiency of day 2 and day 5 transfer in a prospective randomized trial involving patients for whom embryo selection was possible (i.e. those with more than three embryos on day 2 following insemination). We obtained equivalent clinical pregnancy rates per cycle for day 2 (41.7%) and day 5 (38.8%) transfer, but fewer embryos were transferred on day 5 (2.24 versus 3.03). The implantation rates were 18.9% on day 2 and 24.1% on day 5. Selected patients with a good response to gonadotrophins (at least eight good quality metaphase II oocytes) may therefore benefit from blastocyst transfer by a reduction in the multiple pregnancy rate, provided no more than two (or even one) blastocyst is transferred. PMID- 11261481 TI - Clinical experience employing co-culture of human embryos with autologous human endometrial epithelial cells. AB - Attempts to improve the outcome in IVF and related techniques has drawn our attention to the development of culture systems that can grow embryos up to the blastocyst stage. We have developed a co-culture system with autologous human endometrial epithelial cells (EEC) that retained many features of the endometrium. In this review, we analyse our experience over the last 4 years; in particular, we address the question of whether the system would be safe and useful in cases of IVF and oocyte donation with previous implantation failure, and which factors may contribute to the failure of human embryos to develop in vitro up to the blastocyst stage. In all, 168 IVF cycles were carried out in 127 patients with 3.8+/-0.2 previous implantation failure, and 80 cycles in 57 women having oocyte donation with 3.0+/-0.2 previous implantation failure. In 168 IVF cycles, a 48.9% blastocyst formation rate was recorded; 2.3+/-0.1 blastocysts were transferred per cycle and 30 clinical pregnancies obtained (11.9% implantation and 19.6% pregnancy rates). A total of 20 IVF and 15 oocyte donation patients with three previous implantation failures in whom a day 2 embryo transfer was performed were the controls. In 88% of oocyte donation cycles, a 61.0% blastocyst formation rate was observed; 2.3+/-0.1 blastocysts were transferred per cycle and resulted in 38 clinical pregnancies (32.7% implantation and 54.3% pregnancy rates). In all, 15 cycles were cancelled (9%). In oocyte donation patients with implantation failure undergoing day 2 embryo transfer, implantation and pregnancy rates were significantly lower (4.5 and 13.3%; P < 0.01) than with co-culture; however, in IVF patients with implantation failure with day 2 transfer, results (10.7 and 35%) were similar to those with co culture. A second question addressed was whether chromosomal abnormalities may contribute to the failure of human embryos to develop in vitro. We observed the performance of human embryos from our preimplantation diagnosis programme, which were biopsied and subsequently cultured in EEC before transfer. Out of 68 chromosomally normal embryos, 37 reached the blastocyst stage (54.4%) compared with 35 out of 104 abnormal embryos (33.6%). The present study demonstrates that co-culture of human embryos with EEC and blastocyst transfer is safe, effective, and may be a new approach to improve implantation in patients with implantation failure undergoing oocyte donation, but not in IVF patients. The system shows that abnormal embryos can also grow to the blastocyst stage, although to a lower rate. Prospective randomized studies are needed to confirm the preliminary conclusion that co-culture is an acceptable system to select good quality embryos, and the endometrium is a limiting factor in implantation that needs to be carefully managed. PMID- 11261482 TI - The role of the endometrium during embryo implantation. AB - The endometrium undergoes cyclic growth and development with the sole purpose of successful establishment of pregnancy. As more is known about the gene products of the endometrium, it appears that many of the secreted products of the glandular epithelium function to support the nascent embryo and begin the early communication that continues into pregnancy. Maternal endometrial cells are regulated directly by ovarian steroids and indirectly by various growth factors and cytokines. The time of maximal uterine receptivity is now thought to arise on cycle days 20-24 and is manifest by the expression of many different endometrial products. These proteins can serve as markers of uterine receptivity and have been used to identify women at risk for implantation failure. The use of marker proteins promises to promote our understanding of the mechanism of implantation while providing clues into the causes of some types of infertility. Ultimately, the endometrium provides the opportunity for the embryo while at the same time maintaining constraints on uncontrolled invasion of the 'tumour-like' placenta. Understanding this subject in greater detail will likely improve health care opportunities for the infertile couple and provide new insights into contraception targeting the endometrium and embryo-endometrial interactions. PMID- 11261483 TI - A putative role for oncogenes in trophoblast invasion? AB - Tumour invasion and trophoblastic invasion share the same biochemical mediators: the matrix metalloproteinases (MMP) and their inhibitors (TIMP). MMP are a family of enzymes capable of digesting the extracellular matrices of the host tissues. Human cytotrophoblastic cells are constitutively invasive and produce MMP. That MMP are causally related to trophoblast invasion in the endometrium is shown by the fact that TIMPs inhibit cytotrophoblastic invasion in vitro. In contrast to tumour invasion of a host tissue, trophoblastic invasion during implantation and placentation is stringently controlled both in space and time. The factors responsible for these important regulatory processes are unknown, but in-vitro studies point to autocrine (trophoblastic) and paracrine (endometrial) controls by cytokines and growth factors. These regulators exert their effects directly or indirectly by activating nuclear transcription factors. Transcription factors are proteins or protein complexes (often the products of oncogenes) that activate genes by binding to specific sites of the DNA located in the regulatory (5' flanking) region of genes. This review describes the different DNA binding domains of the MMP-9 gene, summarizes our knowledge about the transcription factors involved and speculates about a potential role of these transcription factors (particularly the oncogenes Jun and Fos) in regulating trophoblast invasion. PMID- 11261484 TI - Angiogenesis and implantation. AB - Mammalian implantation is a complex, but still poorly understood, process in which the developing embryo establishes contact with the endometrial surface epithelium before developing a haemochorial placenta. The growth and development of the placenta and its subsequent invasion into the decidua can be likened to the invasion of healthy tissue by malignancy. The establishment of an angiogenic phenotype is a critical event in the progression of benign disease to malignant. Little is known about the role of angiogenesis in implantation but in view of the importance of vascular development to the receptive endometrium, angiogenesis is likely to be critical for successful implantation. Here, the factors that control the development of blood vessels in the pre-implantation phase are reviewed and the complex interplay between the embryo and decidua that leads to the establishment of a healthy placenta is considered. PMID- 11261485 TI - Relevance of endometrial pinopodes for human blastocyst implantation. AB - Endometrial pinopodes, hormone-dependent protrusions of the endometrial apical plasma membrane, have been suggested as indicators of endometrial receptivity. The lifespan of endometrial pinopodes appears to be tightly regulated. Interestingly, considerable interpatient variations according to the onset of pinopode formation have been observed. In normal fertile women, pinopodes are present on days 6-8 post-ovulation, whereas pinopode formation is observed 1-2 days earlier in patients undergoing controlled ovarian hyperstimulation. In oocyte recipients treated with oestradiol and progesterone in hormone-controlled cycles, pinopode formation seems to be delayed. In-vitro experimental studies on human blastocyst attachment to an artificial endometrium indicate a preference for human blastocysts to attach to pinopode presenting areas on the endometrial surface. Transmission electron microscopy shows that the blastocysts do not attach to the apical plasma membrane of pinopode presenting cells. Trophoblast cells display contact with endometrial epithelial cells at the lateral plasma membranes by sharing apical junctional complexes. The function of endometrial pinopodes during human blastocyst implantation still remains to be elucidated. Ultrastructural studies indicate that the pinopode presenting apical plasma membrane does not participate directly in embryo-endometrial interactions. PMID- 11261486 TI - Embryonic regulation of endometrial epithelial apoptosis during human implantation. AB - Apoptosis is a mechanism of cell death in which cells undergo a genetically determined programme. The implanting blastocyst has to appose and adhere to the endometrial epithelium and subsequently invade it. Locally regulated uterine epithelial apoptosis induced by the embryo is a crucial step in epithelial invasion in rodents. To address the physiological relevance of this process in humans, we have investigated the effect of single human blastocysts on the regulation of apoptosis in cultured human endometrial epithelial cells (EEC) in the apposition and adhesion phases of implantation. We report a co-ordinated embryonic regulation of EEC apoptosis. In the apposition phase, the presence of a blastocyst rescues EEC from the apoptotic pathway. However, when the blastocyst adheres to the EEC monolayer it induces a paracrine apoptotic reaction. PMID- 11261487 TI - Culture and transfer of viable blastocysts: a feasible proposition for human IVF. AB - In spite of the numerous advances in the field of human assisted reproductive technologies (ART) over the past 20 years, a rate-limiting factor in the overall efficiency of the procedure (the implantation rate) has remained at 10-30%. The development of sequential media has led to the ability to culture routinely the human embryo to the viable blastocyst stage. Transfer of such blastocysts has resulted in a significant increase in implantation rates. Increases in implantation rates following blastocyst transfer have been reported for specific groups of patients culminating in the elimination of high order multiple gestations. Of greater significance, however, is that the introduction of blastocyst transfer to all patients entering infertility clinics is associated with an overall increase in implantation and pregnancy rates. Blastocysts derived from the use of sequential media are readily cryopreserved and produce high implantation rates after transfer. Using a model to account for both total embryo utilization per cycle (transferred plus cryopreserved) and implantation rate, it has been calculated that extended embryo culture and blastocyst transfer is approximately 20% more efficient than the transfer of cleavage stage embryos on day 3. Furthermore, as the score of the blastocysts obtained using sequential media is directly related to implantation and pregnancy rates, it is possible to determine which patients should be offered a single blastocyst transfer, thereby addressing the issue of twins conceived through ART. PMID- 11261488 TI - Metschnikowia lochheadii and Metschnikowia drosophilae, two new yeast species isolated from insects associated with flowers. AB - Two new haplontic heterothallic species of Metschnikowia were isolated from floricolous insects and flowers. Metschnikowia lochheadii was recovered from insects found in various flowers on the Hawaiian Islands of Kauai and Maui, and from Conotelus sp. (Coleoptera: Nitidulidae) in northwestern Guanacaste Province, Costa Rica. The morphology, physiology, and sexual cycle are typical of the large spored Metschnikowia species, and the partial ribosomal DNA large subunit (D1D2) sequences suggest that the new species is most closely related to Candida ipomoeae. Metschnikowia lochheadii is nearly indistinguishable from its ascogenous relatives and conjugates freely with Metschnikowia continentalis, forming sterile asci. It also exhibits asymmetric mating with Metschnikowia hawaiiensis. Metschnikowia drosophilae was found in morning glory (Ipomoea sp.) flowers and associated Drosophila bromeliae on Grand Cayman Island. Its nutritional profile is atypical of the genus, being the only species that does not utilize sucrose or maltose as carbon sources, and one of the few that does not utilize melezitose. D1D2 sequences show that Metschnikowia drosophilae is a sister species to Candida torresii, to which it bears considerable similarity in nutritional profile. The type cultures are: Metschnikowia lochheadii, strains UWO(PS)00-133.2 = CBS 8807 (h+, holotype) UWO(PS)99-661.1 = CBS 8808 (h-, isotype); and Metschnikowia drosophilae, strains UWO(PS)83-1135.3 = CBS 8809 (h+, holotype) and UWO(PS)83-1143.1 = CBS 8810 (h-, isotype). PMID- 11261489 TI - Isolation and 16S rRNA sequence analysis of the beneficial bacteria from the rhizosphere of rice. AB - The present study deals with the isolation of plant growth promoting rhizobacteria (PGPR) from rice (variety NIAB IRRI-9) and the beneficial effects of these inoculants on two Basmati rice varieties. Nitrogen-fixing activity (acetylene-reduction activity) was detected in the roots and submerged shoots of field-grown rice variety NIAB IRRI-9. Estimation of the population size of diazotrophic bacteria by ARA-based MPN (acetylene reduction assay-based most probable number) in roots and shoots indicated about 10(5)-10(6) counts/g dry weight at panicle initiation and grain filling stages. Four bacterial isolates from rice roots and shoots were obtained in pure culture which produced phytohormone indoleacetic acid (IAA) in the growth medium. Among these, three isolates S1, S4, and R3 reduced acetylene to ethylene in nitrogen-free semi-solid medium. Morphological and physiological characteristics of the isolates indicated that three nitrogen-fixing isolates S1, S4, and R3 belonged to the genus Enterobacter, while the non-fixing isolate R8 belonged to the genus Aeromonas. 16S rRNA sequence of one isolate from root (R8) and one isolate from shoot (S1) was obtained which confirmed identification of the isolates as Aeromonas veronii and Enterobacter cloacae, respectively. The 1517-nucleotide-long sequence of the isolate R8 showed 99% similarity with Aeromonas veronii (accession No. AF099023) while partial 16S rRNA sequence (two stretches of total 1271 nucleotide length) of S1 showed 97% similarity with the sequence of Enterobacter cloacae (accession No. AJ251469). The seedlings of two rice varieties Basmati 385 and Super Basmati were inoculated with the four bacterial isolates from rice and one Azospirillum brasilense strain Wb3, which was isolated from wheat. In the rice variety Basmati 385, maximum increase in root area and plant biomass was obtained in plants inoculated with Enterobacter S1 and Azospirillum Wb3, whereas in the rice variety Super Basmati, inoculation with Enterobacter R3 resulted in maximum increase of root area and plant biomass. Nitrogen fixation was quantified by using 15N isotopic dilution method. Maximum fixation was observed in Basmati 385 with the inoculants Azospirillum Wb3 and Enterobacter S1 where nearly 46% and 41% of the nitrogen was derived from atmosphere (%Ndfa), respectively. In general, higher nitrogen fixation was observed in variety Basmati 385 than in Super Basmati, and different bacterial strains were found more effective as inoculants for the rice varieties Basmati 385 and Super Basmati. PMID- 11261490 TI - Genetic diversity of Bradyrhizobium strains isolated from Arachis hypogaea. AB - Rhizobia are used exclusively in agricultural systems for enhancing the ability of legumes to fix atmospheric nitrogen. Knowledge about the indigenous population is necessary for the selection and application of inoculant strains. In this study, we have assessed the genetic diversity of Bradyrhizobium strains isolated from the host plant, Arachis hypogaea along the coastline of Tamil Nadu. Different populations collected from varying environmental conditions were analysed for salt and pH tolerance. Genetic diversity among the strains was studied using RAPD markers and PCR-RFLP of 16S rDNA and nifD genes. The approaches used in this study yielded consistent results, which revealed a high degree of heterogeneity among strains and detection of two distinct genetic groups. PMID- 11261491 TI - Improving water stress tolerance of the biocontrol yeast Candida sake grown in molasses-based media by physiological manipulation. AB - The biocontrol agent Candida sake was cultured on either an unmodified molasses based medium (water activity, a(w) 0.996) or on water stressed media produced by the addition of glycerol, glucose, NaCl, sorbitol, or proline to 0.98, and 0.96 a(w) for 24, 48, and 72 h, to study their impact on subsequent cell viability, and on concentrations of endogenous sugars (trehalose and glucose) and polyols (glycerol, erythritol, arabitol, and mannitol). The viability of cells of different ages cultured on these media was evaluated on NYDA medium with freely available water (a(w) 0.995), and on medium modified with polyethylene glycol to a(w) 0.95. Regardless of solute used, viable counts of cells grown on molasses based medium (a(w) 0.98) were equal to or higher than those obtained from the medium with water freely available. The amino acid proline stimulated growth at 10% concentration. In contrast, water stress induced by addition of NaCl, glucose, or sorbitol at a(w) 0.96 caused a significant reduction in viable counts. Older cultures were more resistant to water stress. Glycerol and arabitol were the main solutes accumulated by C. sake cells in response to lowered a(w). Intracellular concentration of these polyols depended more on the solute used to adjust the a(w) than on the a(w) itself. Candida sake was more resistant to water stress with higher intracellular concentration of glycerol and erythritol. PMID- 11261492 TI - Comparison of the PR mutant with the wild-type strain of proteus mirabilis brings insight into peroxide resistance factors and regulation of catalase expression. AB - The peroxide resistant mutant (PR) of Proteus mirabilis was characterized by an increased constitutive catalase activity concomitant with a large production of specific mRNA. Survival toward hydrogen peroxide during exponential phase was increased by H2O2 pretreatment in the wild type but not in the mutant, although the catalase of both strains was not inducible under these conditions. In the mutant, besides catalase, over-produced proteins comprised two different alkyl hydroperoxide reductase subunit C (AhpC) proteins and a protein homologous to the stationary phase transcription factor SspA of Escherichia coli. Conversely, the flagellin A (FlaA) of P. mirabilis was repressed in the PR mutant. Genomic DNA fragments of 2.9 kb carrying the catalase gene (katA) together with the 5' and 3' flanking regions were isolated from both strains and found to be identical. Upstream of katA, a Fur box-like sequence was found, but surprisingly, restricting iron in the culture medium caused a decrease in catalase production. The PR mutant presents similarities with other peroxide resistant mutants, but the regulation of catalase biosynthesis in P. mirabilis seems somewhat different from other close species such as E. coli. PMID- 11261493 TI - Genetic diversity of Sinorhizobium populations recovered from different medicago varieties cultivated in Tunisian soils. AB - A collection of 468 rhizobial isolates was obtained from different ecological areas of Tunisia by trapping them on Medicago sativa cv. Gabes, Medicago scutelleta cv. Kelson, Medicago truncatula, and Medicago ciliaris. A subsample of 134 rhizobia was chosen to determine their plasmid profile, and 89 isolates were subjected to multilocus enzyme electrophoresis (MLEE) and PCR/RFLP analysis using 16S, IGS (inter genic spacer), and nifKD probes. Twenty-five representatives from these isolates were evaluated for their nodulation and nitrogen fixation capacities. MLEE studies revealed two groups with highly heterogeneous host specificity and geographical origin. The discriminatory power was found to be slightly better with the amplified ribosomal intergenic region, than the nifKD genes. Divisions detected by nifKD amplified DNA analysis matched those established by ribosomal PCR- RFLPs. The comparison between different analyses revealed that MLEE illustrated better phenotypic properties of isolates than PCR RFLP or plasmid content analysis. Clear distinction between Sinorhizobium meliloti and Sinorhizobium medicae were observed by analysis of the IGS symbiotic regions between nifD and nifK genes. Were able to distinguish three inoculation groups; isolates trapped from M. sativa cv. Gabes and M. scutelleta cv. Kelson formed one inoculation group which was more closely related to isolates trapped from M. truncatula than those trapped from M. ciliaris. PMID- 11261494 TI - Antibiotic production, accumulation of intracellular carbon reserves, and sporulation in Micromonospora echinospora (ATCC 15837). AB - The physiology of the actinomycete Micromonospora echinospora was examined during growth. Biphasic accumulation of glycogen occurred, initially during the early exponential growth phase, and again following the onset of sporulation at 120 h. Lipid levels increased during growth eventually representing 25% of the cell mass. A significant proportion of the lipid was found to be in the form of triacylglycerols, which were found to accumulate markedly during the sporulation phase. The disaccharide trehalose was also found to accumulate during growth with levels rising to 5% of the dry weight during the mycelial production phase, then remaining constant during sporulation. Antibiotic was produced transiently by the cultures over the period preceding sporulation. PMID- 11261495 TI - ICC/PCR detection of enteroviruses and hepatitis A virus in environmental samples. AB - This study applied the integrated cell culture/polymerase chain reaction methodology (ICC/PCR) for rapid and specific detection of both cytopathogenic and noncytopathogenic viruses. Results of this study showed that the use of direct RT PCR or conventional cell culture alone may yield erroneous results with the analysis of environmental samples. The purpose of this study was to compare cultural, molecular, and combined assays for the most effective method of virus detection in variable environmental samples. Using ICC/PCR, stock enterovirus inocula of > or =10 PFU were PCR positive in at least 4/5 replicate flasks after only 5 h of incubation in cell culture, and in all flasks after > or =10 h. An inoculum of one PFU was detected by PCR after 20 h of cell culture incubation while for concentrations of virus below one PFU, 25 h of incubation was sufficient. Similarly, hepatitis A virus (HAV) inocula of 100 MPN/flask, produced indeterminate CPE in cell culture, but were clearly detected by ICC/PCR following 48 h of incubation. Lower levels of HAV, 1 and 10 MPN, were detected by ICC/PCR after 96 to 72 h of incubation, respectively. Cell culture lysates from 11 environmental sample concentrates of sewage, marine water, and surface drinking water sources, were positive for enteroviruses by ICC/PCR compared to 3 positive by direct RT-PCR alone. Results from ICC/PCR eventually agreed with cell culture but required < or =48 h of incubation, compared to as long as 3 weeks for CPE following incubation with BGM and FRhK cells. PMID- 11261496 TI - Mapping of monoclonal antibodies specific to P64k: a common antigen of several isolates of Neisseria meningitidis. AB - P64k is a minor outer membrane protein from Neisseria meningitidis. This protein has been produced at high levels in Escherichia coli. We generated a group of monoclonal antibodies (mAbs) against recombinant P64k, which recognise four non overlapping epitopes, as shown using competition assays with biotinylated mAbs. The P64k sequences involved in mAbs binding were mapped with synthetic overlapping peptides derived from the P64k protein, and located in the previously determined three-dimensional structure of the protein. These antibodies were also characterised by whole-cell ELISA and bactericidal tests against N. meningitidis. Only two of the recognised epitopes were exposed on the bacterial surface, and none of the mAbs showed bactericidal activity. The relationship between these results and the structural data on the epitopes bound by the mAbs is discussed. PMID- 11261497 TI - Early production of rhizopine in nodules induced by Sinorhizobium meliloti strain L5-30. AB - The rhizopine L-3-O-methyl-scyllo-inosamine (3-O-MSI) is metabolized by approximately 10% of the strains of Rhizobium leguminosarum by. viciae and Sinorhizobium meliloti. Rhizopine strains enjoy a substantial competitive advantage in nodulation, which is manifest before 14 days post-inoculation, implying that rhizopine is produced before this time. We were able to detect this compound in the roots of alfalfa (Medicago sativum L. cv. Hunter River) four days after germination (six days post-infection) with S. meliloti strain L5-30 by gas chromatography-mass spectrometry (GC-MS). At four days, nodules were not visible, and the concentration of rhizopine was extremely low, estimated at 67 pg/gfw (picograms/gram fresh weight). The amount increased gradually but remained low until 16 days, when there was a 50-fold increase from day four, by which time nodules were well established. This pattern of synthesis is consistent with previous studies indicating that rhizopine synthesis is regulated by nifA/ntrA regulatory genes, which are maximally expressed in bacteroids at the onset of nitrogen fixation. However, the low level of rhizopine synthesis must be responsible for the early effects on competition for nodulation. Production of rhizopine at this time most likely results from micro-aerobic induction of mos genes in free-living bacteria, either in the infection threads or in the rhizosphere. PMID- 11261499 TI - The ups and downs of multiple sclerosis therapeutics. PMID- 11261498 TI - Functional integrity and structural stability of freeze-dried ectomycorrhizal fungi established through viability assays. AB - Lyophilized vegetative mycelium of ectomycorrhizal fungi was subjected to various viability tests to confirm functional integrity. Physical integrity of freeze dried cultures was comparable to that of non-lyophilized cultures. Inter- and intraspecific variations in morphology, physiology, and metabolic rate were maintained after lyophilization. Maintenance of total protein content confirmed metabolic stability. According to the assays of viability, a plating assay and determination of total biomass confirmed stable mitotic activity of the freeze dried cultures. PMID- 11261500 TI - The neurochemistry of therapeutics: levodopa pharmacodynamics in Parkinson's disease. PMID- 11261501 TI - How does ACTH work against infantile spasms? Bedside to bench. PMID- 11261502 TI - European/Canadian multicenter, double-blind, randomized, placebo-controlled study of the effects of glatiramer acetate on magnetic resonance imaging--measured disease activity and burden in patients with relapsing multiple sclerosis. European/Canadian Glatiramer Acetate Study Group. AB - Two prior double-blind, placebo-controlled, randomized trials demonstrated that glatiramer acetate (GA) reduces relapse rates in patients with relapsing remitting multiple sclerosis (RRMS). This study was designed to determine the effect, onset, and durability of any effect of GA on disease activity monitored with magnetic resonance imaging (MRI) in patients with RRMS. Two hundred thirty nine eligible patients were randomized to receive either 20 mg GA (n = 119) or placebo (n = 120) by daily subcutaneous injection. Eligibility required one or more relapses in the 2 years before entry and at least one enhancing lesion on a screening MRI. The study was a randomized, double-blind, placebo-controlled phase during which all patients studied underwent monthly MRI scans and clinical assessments over 9 months. The primary outcome measure was the total number of enhancing lesions on T1-weighted images. Secondary outcome measures included the proportion of patients with enhancing lesions, the number of new enhancing lesions and change in their volume; the number of new lesions detected on T2 weighted images and change in their volume, and the change in volume of hypointense lesions seen on unenhanced T1-weighted images. Clinical measures of disease activity were also evaluated. The active treatment and placebo groups were comparable at entry for all demographic, clinical, and MRI variables. Treatment with GA showed a significant reduction in the total number of enhancing lesions compared with placebo (-10.8, 95% confidence interval -18.0 to -3.7; p = 0.003). Consistent differences favoring treatment with GA were seen for almost all secondary end points examined: number of new enhancing lesions (p < 0.003), monthly change in the volume of enhancing lesions (p = 0.01), and change in volume (p = 0.006) and number of new lesions seen on T2-weighted images (p < 0.003). The relapse rate was also significantly reduced by 33% for GA-treated patients (p = 0.012). All effects increased over time. Glatiramer acetate significantly reduced MRI-measured disease activity and burden. PMID- 11261503 TI - Biochemical variations in the synaptic level of dopamine precede motor fluctuations in Parkinson's disease: PET evidence of increased dopamine turnover. AB - Motor fluctuations are a major disabling complication in the treatment of Parkinson's disease. To investigate whether such oscillations in mobility can be attributed to changes in the synaptic levels of dopamine, we studied prospectively patients in the early stages of Parkinson's disease with a follow up after at least 3 years of levodopa treatment. At baseline, 3 positron emission tomography (PET) scans using [11C]raclopride before and after (1 hour and 4 hours) orally administered levodopa were performed on the same day for each patient. Patients who developed "wearing-off" fluctuations during the follow-up period had a different pattern of levodopa-induced changes in [11C]raclopride binding potential (BP) from that observed in patients who were still stable by the end of the follow-up. Thus, 1 hour post-levodopa the estimated increase in the synaptic level of dopamine was 3 times higher in fluctuators than in stable responders. By contrast, only stable responders maintained increased levels of synaptic dopamine in the PET scan performed after 4 hours. These results indicate that fluctuations in the synaptic concentration of dopamine precede clinically apparent "wearing-off" phenomena. The rapid increase in synaptic levels of dopamine observed in fluctuators suggests that increased dopamine turnover might play a relevant role in levodopa-related motor complications. PMID- 11261504 TI - Corticotropin (ACTH) acts directly on amygdala neurons to down-regulate corticotropin-releasing hormone gene expression. AB - The hormone corticotropin (ACTH) is employed as therapy for diverse neurological disorders, but the mechanisms for its efficacy remain unknown. ACTH promotes the release of adrenal steroids (glucocorticoids), and most ACTH effects on the central nervous system (CNS) have been attributed to activation of glucocorticoid receptors. However, in several human disorders, ACTH has therapeutic actions that differ qualitatively or quantitatively from those of steroids. This study tested the hypothesis that ACTH directly influences limbic neurons via the recently characterized melanocortin receptors and focused on the effects of ACTH on the expression of corticotropin-releasing hormone (CRH), a neuropeptide involved in neuroimmune functions and in certain developmental seizures. The results demonstrated that ACTH potently reduced CRH expression in amygdala neurons. This down-regulation was not abolished by experimental elimination of steroids or by blocking their receptors and was reproduced by a centrally administered ACTH fragment that does not promote steroid release. Importantly, selective blocking of melanocortin receptors prevented ACTH-induced down-regulation of CRH expression. Taken together, these data indicate that ACTH activates central melanocortin receptors to modulate CRH gene expression in amygdala, supporting the notion that direct, steroid-independent actions of ACTH may account for some of its established clinical effects on the CNS. PMID- 11261505 TI - Clinical and pathological features of a Parkinsonian syndrome in a family with an Ala53Thr alpha-synuclein mutation. AB - We describe an Australian family of Greek origin with a parkinsonian syndrome and an Ala53Thr alpha-synuclein gene mutation. Five of 9 siblings were affected, the average age of onset was 45 years, and the initial symptoms were variable, including resting tremor, bradykinesia, and gait disturbance, as previously described in families with the same point mutation. Affected family members responded well to levodopa, developed progressive cognitive impairment, and had a disease duration of 5 to 16 years. Pathologic features typical of idiopathic Parkinson's disease were found at autopsy. However, there were several additional features not previously reported in families with this gene mutation. These features included severe central hypoventilation, orthostatic hypotension, prominent myoclonus, and urinary incontinence. An abundance of alpha-synuclein immunoreactive Lewy neurites were found in the brainstem pigmented nuclei, hippocampus, and temporal neocortex. The Lewy neurites were associated with temporal lobe vacuolation. Subcortical basal ganglia cell loss and gliosis were seen. These additional clinical and pathological features suggest that the Ala53Thr alpha-synuclein mutation can produce a more widespread disorder than found in typical idiopathic Parkinson's disease. PMID- 11261506 TI - Differences in sensory and motor cortical organization following brain injury early in life. AB - There have been a number of physiological studies of motor recovery in hemiplegic cerebral palsy which have identified the presence of novel ipsilateral projections from the undamaged hemisphere to the affected hand. However, little is known regarding the afferent projection to sensory cortex and its relationship to the reorganized cortical motor output. We used transcranial magnetic stimulation (TMS) to investigate the corticomotor projection to the affected and unaffected hands in a group of subjects with hemiplegic cerebral palsy, and also performed functional magnetic resonance imaging (fMRI) studies of the patterns of activation in cortical motor and sensory areas following active and passive movement of the hands. Both TMS and fMRI demonstrated a normal contralateral motor and sensory projection between the unaffected hand and the cerebral hemisphere. However, in the case of the affected hand, the TMS results indicated either a purely ipsilateral projection or a bilateral projection in which the ipsilateral pathway had the lower motor threshold, whereas passive movement resulted in fMRI activation in the contralateral hemisphere. These results demonstrate that there is a significant fast-conducting corticomotor projection to the affected hand from the ipsilateral hemisphere in this group of subjects, but that the predominant afferent projection from the hand is still directed to the affected contralateral hemisphere, resulting in an interhemispheric dissociation between afferent kinesthetic inputs and efferent corticomotor output. The findings indicate that there can be differences in the organization of sensory and motor pathways in cerebral palsy, and suggest that some of the residual motor dysfunction experienced by these subjects could be due to an impairment of sensorimotor integration at cortical level as a result of reorganization in the motor system. PMID- 11261507 TI - Genome scan of idiopathic generalized epilepsy: evidence for major susceptibility gene and modifying genes influencing the seizure type. AB - Idiopathic generalized epilepsy (IGE) is a common, complex disease with an almost exclusively genetic etiology but with variable phenotypes. Clinically, IGE can be divided into different syndromes. Varying lines of evidence point to the involvement of several interacting genes in the etiology of IGE. We performed a genome scan in 91 families ascertained through a proband with adolescent-onset IGE. The IGEs included juvenile myoclonic epilepsy (JME), juvenile absence epilepsy (JAE), and epilepsy with generalized tonic clonic seizures (EGTCS). Our linkage results support an oligogenic model for IGE, with strong evidence for a locus common to most IGEs on chromosome 18 (lod score 4.4/5.2 multipoint/two point) and other loci that may influence specific seizure phenotypes for different IGEs: a previously identified locus on chromosome 6 for JME (lod score 2.5/4.2), a locus on chromosome 8 influencing non-JME forms of IGE (lod score 3.8/2.5), and, more tentatively, two newly discovered loci for absence seizures on chromosome 5 (lod scores 3.8/2.8 and 3.4/1.9). Our data also suggest that the genetic classification of different forms of IGE is likely to cut across the clinical classification of these subforms of IGE. We hypothesize that interactions of different combinations of these loci produce the related heterogeneous phenotypes seen in IGE families. PMID- 11261508 TI - Mortality in epilepsy in the first 11 to 14 years after diagnosis: multivariate analysis of a long-term, prospective, population-based cohort. AB - The United Kingdom National General Practice Study of Epilepsy is a prospective, population-based study of newly diagnosed epilepsy. A cohort of 792 patients has now been followed for up to 14 years (median follow-up [25th, 75th percentiles] 11.8 years, range 10.6-11.7 years), a total of 11,400 person-years. These data are sufficient for a detailed analysis of mortality in this early phase of epilepsy. Over 70% of patients in this cohort have developed lasting remission from seizures, although the mortality rate in the long term was still twice that of the general population. The standardized mortality ratio (SMR), the number of observed deaths per number of expected deaths, was 2.1 (95% confidence interval [CI] = 1.8, 2.4). Patients with acute symptomatic epilepsy (SMR 3.0; 95% CI = 2.0, 4.3), remote symptomatic epilepsy (SMR 3.7; 95% CI = 2.9, 4.6), and epilepsy due to congenital neurological deficits (SMR 25; 95% CI = 5.1, 73.1) had significantly increased long-term mortality rates, whereas patients with idiopathic epilepsy did not (SMR 1.3; 95% CI = 0.9, 1.9). This increase in mortality rate was noted particularly in the first few years after diagnosis. Multivariate Cox regression and time-dependent co-variate analyses were utilized for the first time in a prospective study of mortality in epilepsy. The former showed that patients with generalized tonic-clonic seizures had an increased risk of mortality. The hazard ratio (HR), or risk of mortality in a particular group with a particular risk factor compared to another group without that particular risk factor, was 6.2 (95% CI = 1.4, 27.7; p = 0.049). Cerebrovascular disease (HR 2.4; 95% CI = 1.7, 3.4; p < 0.0001), central nervous system tumor (HR 12.0; 95% CI = 7.9, 18.2; p < 0.0001), alcohol (HR 2.9; 95% CI = 1.5, 5.7; p = 0.004), and congenital neurological deficits (HR 10.9; 95% CI = 3.2, 36.1; p = 0.003) as causes for epilepsy and older age at index seizure (HR 1.9; 95% CI = 1.7,2.0; p < 0.0001) were also associated with significantly increased mortality rates. These hazard ratios suggest that epilepsy due to congenital neurological deficits may carry almost the same risk of mortality as epilepsy due to central nervous system tumors and that epileptic seizures subsequent to alcohol abuse may carry almost the same risk of mortality as epilepsy due to cerebrovascular disease. The occurrence of one or more seizures before the index seizure (the seizure that led to the diagnosis of epilepsy and enrolment in the study) was associated with a significantly reduced mortality rate (HR 0.57; 95% CI = 0.42, 0.76; p = 0.00001). Time-dependent co-variate analysis was used to examine the influence of ongoing factors, such as seizure recurrence, remission, and antiepileptic drug use, on mortality rates in the cohort. Seizure recurrence (HR 1.30; 95% CI = 0.84, 2.01) and antiepileptic drug treatment (HR 0.97; 95% CI = 0.67, 1.38) did not influence mortality rate. There were only 5 epilepsy-related deaths (1 each of sudden unexpected death in epilepsy, status epilepticus, burns, drowning, and cervical fracture), suggesting that death directly due to epileptic seizures is uncommon in a population-based cohort with epilepsy. PMID- 11261509 TI - Sequence-selective DNA binding drugs mithramycin A and chromomycin A3 are potent inhibitors of neuronal apoptosis induced by oxidative stress and DNA damage in cortical neurons. AB - Global inhibitors of RNA or protein synthesis such as actinomycin D or cycloheximide abrogate neuronal apoptosis induced by numerous pathological stimuli in vitro and in vivo. The clinical application of actinomycin D or cycloheximide to human neurological disease has been limited by the toxicities of these agents. To overcome these toxicities, strategies must be developed to inhibit selectively the expression of deleterious proapoptotic proteins, while leaving the expression of antiapoptotic, proregeneration, and other critical homeostatic proteins unperturbed. Mithramycin A (trade name Plicamycin) is an aureolic acid antibiotic that has been used in humans to treat hypercalcemia and several types of cancers. This class of agents is believed to act, in part, by selectively inhibiting gene expression by displacing transcriptional activators that bind to G-C-rich regions of promoters. Here we demonstrate that mithramycin A and its structural analog chromomycin A3 are potent inhibitors of neuronal apoptosis induced by glutathione depletion-induced oxidative stress or the DNA damaging agent camptothecin. We correlate the protective effects of mithramycin A with its ability to inhibit enhanced DNA binding of the transcription factors Sp1 and Sp3 to their cognate "G-C" box induced by oxidative stress or DNA damage. The protective effects of mithramycin A cannot be attributed to global inhibition of protein synthesis. Together, these results suggest that mithramycin A and its structural analogs may be effective agents for the treatment of neurological diseases associated with aberrant activation of apoptosis and highlight the potential use of sequence-selective DNA-binding drugs as neurological therapeutics. PMID- 11261510 TI - Orbitofrontal and anterior cingulate cortex neurofibrillary tangle burden is associated with agitation in Alzheimer disease. AB - Few studies evaluate neuropathological correlates of behavioral changes in Alzheimer disease (AD). We identified 31 autopsy patients with a diagnosis of definite AD. Behavioral changes were assessed with the Neuropsychiatric Inventory. Brain sections were collected from bilateral orbitofrontal and left anterior cingulate, superior temporal, inferior parietal, occipital, and hippocampal cortices for quantification of neurofibrillary tangles (NFTs) and diffuse and neuritic plaques. Sections from frontal, cingulate, and hippocampal cortices were reviewed for the presence of Lewy bodies (LBs). Hypothesis-driven correlational analyses were performed by the bootstrap method. Subgroup analyses contrasted a group with high scores of one specific behavior to a group with low scores after equating groups for other behaviors. NFT burden in the left orbitofrontal cortex across all 31 patients significantly correlated with agitation scores (r = 0.41, p < 0.015) and NFTs correlated significantly (r = 0.66, p = 0.004) with higher agitation scores in the subgroup analysis. Left anterior cingulate NFTs, although not within our hypotheses, also showed a significant relationship to agitation within the subgroups (r = 0.76, p = 0.0003; Bonferroni p = 0.02). Seven patients, including three in the agitation subgroup, had cortical LBs. Aberrant motor behavior and NFT density in the left orbitofrontal cortex showed a significant relationship for the entire group (r = 0.38, p < 0.03) and for subgroups (r = 0.49, p = 0.04), whereas apathy and left anterior cingulate NFTs showed a significant relationship only for the entire group (r = 0.25, p < or = 0.01). These observations suggest that agitation and aberrant motor behavior are correlates of greater NFT pathology in the orbitofrontal cortex in AD, whereas increasing apathy may relate to greater NFT burden in the anterior cingulate. PMID- 11261511 TI - DYT13, a novel primary torsion dystonia locus, maps to chromosome 1p36.13--36.32 in an Italian family with cranial-cervical or upper limb onset. AB - Primary torsion dystonia (PTD) is a clinically and genetically heterogeneous group of movement disorders, usually inherited in an autosomal dominant fashion with reduced penetrance. The DYT1 gene on chromosome 9q34 is responsible for most cases of early limb-onset PTD. Two other PTD loci have been mapped to date. The DYT6 locus on chromosome 8 is associated with a mixed phenotype, whereas the DYT7 locus on chromosome 18p is associated with adult onset focal cervical dystonia Several families have been described in which linkage to the known PTD loci have been excluded. We identified a large Italian PTD family with 11 definitely affected members. Phenotype was characterized by prominent cranial-cervical and upper limb involvement and mild severity. A genome-wide search was performed in the family. Linkage analysis and haplotype construction allowed us to identify a novel PTD locus (DYT13) within a 22 cM interval on the short arm of chromosome 1, with a maximum lod score of 3.44 between the disease and marker D1S2667. PMID- 11261512 TI - Positron emission tomographic analysis of the nigrostriatal dopaminergic system in familial parkinsonism associated with mutations in the parkin gene. AB - A kindred from South Tyrol (northern Italy) with familial, adult-onset parkinsonism of pseudo-dominant inheritance and mutations in the parkin gene was recently described. To gain insight into basal ganglia dysfunction in this form of hereditary parkinsonism, positron emission tomography (PET) with 18-fluorodopa (FDOPA) and 11C-raclopride (RAC) was performed in 5 affected family members and 5 asymptomatic relatives with proven compound heterozygous or heterozygous parkin mutations. Results were compared to findings in healthy control subjects and patients with typical sporadic, idiopathic Parkinson's disease. Similar to findings in the sporadic Parkinson's disease group, presynaptic striatal FDOPA storage was decreased in patients with compound heterozygous parkin mutations, with the most prominent reduction in the posterior part of the putamen. Along with the presynaptic lowered FDOPA uptake, we found a uniform reduction of the striatal 11C-raclopride binding index in all affected family members as compared to asymptomatic family members carrying a heterozygous parkin mutation, sporadic Parkinson's disease, and control subjects. Our PET data provide evidence that parkinsonism in this family is associated with presynaptic dopaminergic dysfunction similar to idiopathic Parkinson's disease pathophysiology, along with alterations at the postsynaptic D2 receptor level. In asymptomatic carriers of a single parkin mutation with an apparently normal allele, we found a mild but statistically significant decrease of mean FDOPA uptake compared to control subjects in all striatal regions. These data indicate a preclinical disease process in these subjects. PMID- 11261513 TI - The incidence of mitochondrial encephalomyopathies in childhood: clinical features and morphological, biochemical, and DNA abnormalities. AB - In this study we present incidence, point prevalence, and mortality figures of mitochondrial encephalomyopathies in a population-based study of children from western Sweden. Through the screening of registers and review of medical records, we identified 32 patients under 16 years of age from the study population who were diagnosed between January 1, 1984, and December 31, 1998. The incidence of mitochondrial encephalomyopathies in preschool children (<6 years of age) was 1 out of 11,000. The preschool incidence of Leigh's syndrome was 1 out of 32,000, and the preschool incidences of both Alper's syndrome and infantile mitochondrial myopathy with cytochrome C oxidase deficiency were 1 out of 51,000. The point prevalence January 1, 1999) of mitochondrial encephalomyopathies in children under 16 years of age was 1 out of 21,000. The median survival for patients with infantile onset was until 12 years of age. We identified 4 cases with mitochondrial DNA point mutations, 2 cases with mitochondrial DNA deletions, and 2 cases with nuclear mutations in the SURF1 gene. We conclude that mitochondrial encephalomyopathies are relatively common neurometabolic disorders in childhood. PMID- 11261514 TI - McLeod syndrome: a novel mutation, predominant psychiatric manifestations, and distinct striatal imaging findings. AB - The McLeod syndrome is an X-linked disorder caused by mutations of the XK gene encoding the XK protein. The syndrome is characterized by absent Kx erythrocyte antigen, weak expression of Kell blood group system antigens, and acanthocytosis. In some allelic variants, elevated creatine kinase, myopathy, neurogenic muscle atrophy, and progressive chorea are found. We describe a family with a novel point mutation in the XK gene consisting of a C to T base transition at nucleotide position 977, introducing a stop codon. Among seven affected males, five manifested with psychiatric disorders such as depression, bipolar disorder, or personality disorder, but only two presented with chorea Positron emission tomography and magnetic resonance volumetry revealed reduced striatal 2-fluoro-2 deoxy-glucose (FDG) uptake and diminished volumes of the caudate nucleus and putamen that correlated with disease duration. In contrast, none of 12 female mutation carriers showed psychiatric or movement disorders. However, a semidominant effect of the mutation was suggested by erythrocyte and blood group mosaicism and reduced striatal FDG uptake without structural abnormalities. Therefore, patients with psychiatric signs or symptoms segregating in an X-linked trait should be examined for acanthocytosis and Kell/Kx blood group serology. PMID- 11261515 TI - Children with comorbid attention-deficit-hyperactivity disorder and tic disorder: evidence for additive inhibitory deficits within the motor system. AB - For children with attention-deficit-hyperactivity disorder (ADHD) or tic disorder (TD), we recently reported deficient inhibitory mechanisms within the motor system by using transcranial magnetic stimulation. These deficits--stated as reduced intracortical inhibition in ADHD and shortened cortical silent period in TD--could be seen as neurophysiological correlates of motor hyperactivity and tics, respectively. To investigate neurophysiological aspects of comorbidity, we measured motor system excitability for the first time also in children with combined ADHD and TD. The findings of a reduced intracortical inhibition as well as a shortened cortical silent period in these comorbid children provide evidence for additive effects at the level of motor system excitability. PMID- 11261516 TI - Neurological and neuropathologic heterogeneity in two brothers with cobalamin C deficiency. AB - Two adult brothers, one documented to have methylmalonic acidemia with homocystinuria, or cobalamin C deficiency, after autopsy, displayed severe but divergent neurological presentations. One exhibited a myelopathy and the other chronic endocrine problems (Schmidt's syndrome) followed by a neuropsychiatric and dementing disorder owing to cerebral perivascular demyelination. The recognition of cobalamin C deficiency has practical implications because it is one of the few inherited diseases of central white matter that is treatable. PMID- 11261517 TI - Irreversible brain creatine deficiency with elevated serum and urine creatine: a creatine transporter defect? AB - Recent reports highlight the utility of in vivo magnetic resonance spectroscopy (MRS) techniques to recognize creatine deficiency syndromes affecting the central nervous system (CNS). Reported cases demonstrate partial reversibility of neurologic symptoms upon restoration of CNS creatine levels with the administration of oral creatine. We describe a patient with a brain creatine deficiency syndrome detected by proton MRS that differs from published reports. Metabolic screening revealed elevated creatine in the serum and urine, with normal levels of guanidino acetic acid. Unlike the case with other reported creatine deficiency syndromes, treatment with oral creatine monohydrate demonstrated no observable increase in brain creatine with proton MRS and no improvement in clinical symptoms. In this study, we report a novel brain creatine deficiency syndrome most likely representing a creatine transporter defect. PMID- 11261519 TI - Anticardiolipin antibodies are not a useful screening tool in a nonselected large group of patients with multiple sclerosis. AB - Recent works claiming that primary antiphospholipid syndrome (PAPS) cannot be clinically distinguished from multiple sclerosis (MS) recommend that MS patients be screened for anticardiolipin antibodies (ACA). In this study 296 randomly selected patients with MS and clinically isolated syndromes and 51 healthy controls were analyzed; ACA, anti-beta2-glycoprotein I, or antiprothrombin was found in 6 patients. No predominance of any kind of clinical manifestations and no cardinal manifestations of PAPS were found in these patients. ACA tests should be performed only when a suspicion of PAPS is raised and atypical clinical presentation for MS is found. PMID- 11261518 TI - Reduction of Menkes mRNA and copper in leukocytes of patients with primary adult onset dystonia. AB - Studies on postmortem tissue of patients with primary adult-onset dystonia revealed a significant increase in copper levels and a reduction of copper transporting Menkes protein of the lentiform nuclei. Here we demonstrate that patients with idiopathic adult-onset cervical dystonia (n = 14) have reduced Menkes mRNA copies and lower copper levels in leukocytes compared to controls (n = 17; U test, p < 0.05). Changes were less distinct in patients with blepharospasm. Therefore, disturbances of copper metabolism in focal dystonia may not be restricted to the basal ganglia. PMID- 11261520 TI - Low cerebral blood flow velocity and risk of white matter hyperintensities. AB - Cerebral blood flow velocity (CBF-V) measured by transcranial doppler was assessed in 628 elderly individuals who had cerebral magnetic resonance imaging performed as part of a population-based study on vascular aging. Cerebral white matter hyperintensities (WMHs) were associated with low CBF-V, such as the adjusted odds ratios of severe WMHs from highest (referent) to lowest quartile of mean CBF-V were 1.0, 1.7, 3.7, and 4.3 (p = 0.001). Further, CBF-V was found to be a stronger risk factor for WMHs than high blood pressure. These findings suggest that the assessment of CBF-V might be a powerful tool in future studies on WMHs. PMID- 11261522 TI - Central acetylcholinesterase inhibition in Alzhemier patients. PMID- 11261521 TI - Effects of seasons on magnetic resonance imaging--measured disease activity in patients with multiple sclerosis. PMID- 11261523 TI - Converting patients from brand-name clozapine to generic clozapine. AB - OBJECTIVE: To evaluate safety and dosage requirements when patients taking brand name clozapine (Clozaril, Novartis Pharmaceuticals) are converted to generic clozapine (Zenith Goldline). METHODS: In November 1999, patients at Colorado Mental Health Institute at Pueblo taking Clozaril were changed to generic clozapine. Seventeen patients had been prescribed Clozaril for three years and were included in the study. Drug dosage, white blood cell (WBC) count values, and adverse drug reaction reports were compared. Data regarding patients on the brand name product were evaluated retrospectively for the months of November, December, January, and February during the years 1996/1997, 1997/1998, and 1998/1999. These data were compared with those from the same patients after switching to generic clozapine for the same months in 1999/2000. A one-year comparison of brand-name (1998/1999) with generic drug (1999/2000) was also performed. Statistical analysis included a standard test comparing WBC values and a Brown-Forsythe test for comparing dosages. RESULTS: There were no differences between the values obtained for the brand-name and generic products. WBC counts for the three-year data resulted in a p value of 0.9992. There was no difference when comparing the samples one year prior to switching and after the switch (p = 0.9991). There was no difference in dosages at three years or one year (p = 0.9999 and p = 0.9993, respectively). No adverse events were noted with the generic product. CONCLUSIONS: No differences were found between the brand-name and generic clozapine groups with regard to WBC count, dosage, and adverse events. The conversion to the generic product is projected to save the pharmacy $90,000 annually. PMID- 11261524 TI - Comparison of FDA reports of patient deaths associated with sildenafil and with injectable alprostadil. AB - BACKGROUND: Sildenafil (Viagra) has been linked to 240 deaths (128 verified, 112 unverified) reported to the Food and Drug Administration (FDA) during 7.5 months of availability, and to 522 reported deaths after 13 months of availability. To date, no updated information about FDA-reported deaths has emerged, and no comparative analyses have been published. OBJECTIVE: To compare the mortality rates between sildenafil and injectable alprostadil, both of which are used exclusively for treating erectile dysfunction. METHODS: A comparison of the number of deaths per filled prescriptions reported to the FDA involving sildenafil and injectable alprostadil was undertaken to perhaps provide further insight into this issue. Materials included FDA statements on sildenafil adverse event reports to the FDA involving sildenafil and injectable alprostadil, and data on prescriptions filled for sildenafil and injectable alprostadil. RESULTS: The number of deaths per prescriptions filled reported in association with sildenafil was significantly greater (5.15-6.28 times) than in association with injectable alprostadil. DISCUSSION: Previous explanations for sildenafil associated deaths have been based on the expected attrition within the population of men with erectile dysfunction and its commonly associated disorders, the physiologic stress of renewed sexual activity, and a pharmacologic effect of sildenafil. The results of this analysis may indicate that a pharmacologic effect of sidenafil is responsible for these deaths. However, other factors may also explain these findings: a greater frequency of reporting of sildenafil-associated events by physicians, a difference in the populations using these two drugs, or the number of prescriptions filled may not accurately reflect actual exposure. CONCLUSIONS: Further study should be undertaken to clarify the issues associated with sildenafil-related deaths. In the meantime, reasonable precautions might be considered in prescribing sildenafil, such as initiating treatment with a low test dose of sildenafil. PMID- 11261525 TI - Pharmacists' preferences for continuing education and certificate programs. AB - OBJECTIVE: To conduct an assessment of the needs and interests of West Virginia pharmacists with respect to continuing education (CE) and certificate programs (CP) versus a nontraditional PharmD program (NTP). METHODS: A cross-sectional study was conducted. The survey was mailed to 2800 West Virginia University School of Pharmacy alumni and West Virginia licensed pharmacists. The survey collected data pertaining to pharmacists' perceptions for the needs of CE, CP, and the NTP program; the optimal structuring of these programs; and the demographics of participants. RESULTS: A 24% (674) usable response rate was achieved from two mailings. Respondents were asked to address all areas of interest: approximately 75% showed interest in enrolling in CE, 45% in CP, and 40% in the NTP program. Interest levels varied across demographic and practice characteristics. Seven methods of instruction were evaluated by pharmacists, with live lectures being the most preferred for both CE and CP. Interest for specific content areas and topics for CE workshops and CP were identified. The type and amount of employer support and willingness to pay for enrollment in the two types of programs were obtained. Markets' needs for CE and CP were identified for five typical pharmacists' profiles (e.g., staff pharmacists in community practice). CONCLUSIONS: Results can be used by CE providers to develop CE and CP programs of most interest to pharmacists and target them to appropriate demographic segments in order to be cost-effective. PMID- 11261526 TI - Olanzapine-lnduced hyperglycemic nonketonic coma. AB - OBJECTIVE: To report a case of olanzapine-induced hyperglycemia leading to a hyperosmolar, hyperglycemic, nonketonic coma. CASE SUMMARY: A 51-year-old, 85.5 kg (ideal body weight 79.9 kg), white man presented to a Veterans Affairs hospital with a serum glucose concentration of 1596 mg/dL. Soon thereafter, he went into a hyperosmolar, hyperglycemic, nonketonic coma. Olanzapine therapy had been instituted less than six months prior to this event; approximately two months before this event, his blood glucose was 108 mg/dL. Eight days after stopping olanzapine, the glucose concentration returned to normal, and the patient no longer required insulin nor any other glucose-lowering agents. DISCUSSION: The insulin resistance caused by olanzapine is normally attributed to the weight gain associated with the drug. In this patient, it appears that olanzapine caused hyperglycemia by a mechanism other than weight gain. CONCLUSIONS: This case report and others from the literature suggest that olanzapine therapy may induce hyperglycemia in some patients. PMID- 11261528 TI - Amphotericin B--not so terrible. AB - OBJECTIVE: To describe a patient who developed adverse reactions to two different lipid formulations of amphotericin B: liposomal amphotericin B (AmBisome) and amphotericin B colloidal dispersion (ABCD, Amphocil), yet tolerated amphotericin B deoxycholate (Fungizone) despite renal toxicity. CASE SUMMARY: A 72-year-old woman with acute myelomonocytic leukemia was treated with amphotericin B deoxycholate for suspected pulmonary aspergillosis; the drug was well tolerated but resulted in renal failure. Antifungal therapy was then changed to liposomal amphotericin B. Within 10 minutes of liposomal amphotericin B infusion, the patient developed severe dyspnea, chest pain, and a feeling of imminent death. On the following day, liposomal amphotericin B was switched to amphotericin B colloidal dispersion. Again, within 10 minutes of this infusion, the patient developed fever, chills, hypotension, severe chest pain, dsypnea, and a feeling of imminent death. The patient refused any further treatment with these drugs and insisted on switching back to amphotericin B deoxycholate, which was then administered for 10 days and was well tolerated. DISCUSSION: Severe adverse reactions, such as anaphylaxis, cardiac toxicity, and respiratory failure, following administration of all three lipid formulations of amphotericin B have been reported. In most reported cases, switching to a different lipid formulation of amphotericin B was well tolerated. This is in contrast to our case, where a severe reaction was repeated when another lipid preparation was given, necessitating switching back to amphotericin B deoxycholate despite its nephrotoxicity. CONCLUSIONS: In some patients, paradoxically, lipid formulations of amphotericin B may be less tolerable than conventional amphotericin B. PMID- 11261527 TI - Intravenous theophylline--an alternative to temporary pacing in the management of bradycardia secondary to AV nodal block. AB - OBJECTIVE: To report a case of bradycardia secondary to atrioventricular nodal block (AVNB) successfully treated with intravenous theophylline. Intravenous theophylline was used as an alternative to temporary pacing in a patient with sepsis secondary to thermal injury. CASE SUMMARY: A 79-year-old white woman with significant cardiac history was admitted with 14.5% total body surface area burns after a house fire. Cardiac events included intermittent episodes of sinus bradycardia complicated by the development of second-degree AVNB and periods of sinus arrest. Intravenous theophylline initiation maintained normal sinus rhythm without further episodes of sinus bradycardia or heart block, thus preventing the need for cardiac pacemaker placement. DISCUSSION: This is the first case published in the English-language literature describing the use of intravenous theophylline as an alternative therapy to temporary pacing in a patient with sepsis secondary to thermal injury. Bradyarrhythmic events in sepsis patients have been associated with catecholamine production increasing adenosine formation. High concentrations of adenosine in the areas of the sinoatrial or atrioventricular nodal regions may induce sinus bradycardia or AVNB. Theophylline, an adenosine antagonist, has been identified as a treatment option for such bradyarrhythmic events. CONCLUSIONS: Theophylline, a methylxanthine derivative, may represent an alternative to other pharmacologic therapies and temporary pacing in the treatment of bradycardia secondary to AVNB. These agents may represent a pharmacologic alternative in patients in whom other pharmacologic strategies or cardiac pacemaker insertion may be contraindicated. PMID- 11261529 TI - Seizure induced by ropivacaine. AB - OBJECTIVE: To report development of a seizure after administration of ropivacaine. CASE SUMMARY: A 26-year-old woman was scheduled for a cesarean section because of a stagnation of the uterine neck dilatation after 4.5 hours. After peridural administration of 279 mg of ropivacaine (total dose) over five hours, she presented with oculogyric movements and slurred speech that preceded convulsions of the face and of the upper limbs. DISCUSSION: Convulsions are well known complications of local anesthetics. Ropivacaine, a relatively new agent, is considered safer for the central nervous system. Currently, there are only four published reports that implicate ropivacaine as being associated with convulsions. The likelihood that ropivacaine caused the seizure in our patient was possible based on the Naranjo probability scale. CONCLUSIONS: Clinicians should be aware of the possibility of seizures as an adverse effect of ropivacaine. PMID- 11261530 TI - Orlistat--a novel weight loss therapy. AB - OBJECTIVE: To review the pharmacology, pharmacokinetics, clinical safety and efficacy, drug interactions, and therapeutic issues related to the use of orlistat for treatment of obesity. DATA SOURCES: English-language articles were identified from MEDLINE (1966-July 2000), Roche Laboratories, organizational guidelines, National Institutes of Health and Food and Drug Administration Web sites, and Doctor's Guide online. Key words included obesity, orlistat, and lipase inhibitors. References were also identified from reference sections of published articles. STUDY SELECTION AND DATA EXTRACTION: Prospective, randomized, double-blind, placebo-controlled, human trials were selected for review and discussion. DATA SYNTHESIS: Orlistat is the first agent in the lipase inhibitor class of antiobesity drugs. Orlistat is minimally absorbed and has been shown to reduce body weight by inhibiting absorption (by approximately 30%) of ingested dietary fat. Safety and efficacy have been established in one- and two-year double-blind, placebo-controlled trials; adverse effects were primarily, and almost exclusively, gastrointestinal. Due to its ability to block fat absorption, orlistat also has the capability to inhibit absorption of fat-soluble vitamins. Therefore, a daily multiple vitamin is recommended while taking orlistat. CONCLUSIONS: By inhibiting fat absorption, orlistat offers a new treatment modality for weight loss and maintenance. Preliminary data from clinical trials suggest that orlistat may be beneficial in patients with comorbid conditions related to obesity, such as diabetes and hyperlipidemia. However, further studies during postmarketing surveillance are needed to fully establish orlistats long term benefits and safety. Orlistat should be considered a useful adjunctive therapy for weight loss and maintenance in obese patients (i.e., body mass index > or = 30 kg/m2 or > or = 27 kg/m2 if other risk factors are present) committed to lifestyle changes including diet, exercise, and behavioral modification. PMID- 11261531 TI - Implanon: a critical review. AB - OBJECTIVE: To evaluate clinical information on Implanon as a long-term method of contraception and, specifically, to review the efficacy and adverse effect profile of Implanon. DATA SOURCES: A MEDLINE search (from 1966 through June 1999) was performed to retrieve primary and review articles. The search was limited to data on human subjects. Some references were identified through secondary sources. STUDY SELECTION AND DATA EXTRACTION: Pharmacokinetic and pharmacodynamic studies, and clinical trials assessing the efficacy and safety of desogestrel implants were reviewed. Relevancy and consistency of information was assessed for each trial. DATA SYNTHESIS: Implanon is the newest contraceptive implant system that has completed worldwide Phase III clinical trials. It is a single rod that contains a core of 68 mg of 3-keto-desogestrel with a membrane of ethylene vinyl acetate. Inhibition of ovulation occurs within one day of implantation, and effective contraception lasts for three years. Fertility returns within one month after implant removal. Insertion and removal by trained medical professionals is simple, and only minor complications have been documented. Adverse effects are mild and primarily consist of abnormal bleeding, weight gain, acne, breast pain, and headache. CONCLUSIONS: The data on Implanon indicate that it provides effective long-term contraception with limited adverse effects. It appears to be a good addition to the currently available contraceptives. PMID- 11261532 TI - Is the sildenafil product information adequate to facilitate informed therapeutic decisions? AB - BACKGROUND: Optimal therapeutics and the prevention of adverse drug effects begin with complete information. When new drugs are released, the manufacturer's product information is the main and often only readily available source of drug information and, therefore, greatly influences treatment strategies. Thus, it is vital that the information in the package insert is not only complete, but also as relevant as possible for the great diversity of patients that physicians encounter. OBJECTIVE: To review the product information for sildenafil for comprehensiveness and accuracy, with respect to whether the information is sufficient to facilitate optimal therapeutics and to prevent avoidable adverse events in the wide diversity of patients with erectile dysfunction seen in clinical practice. DATA SOURCES: Sildenafil package inserts, unpublished information provided by the manufacturer, Food and Drug Administration reports, and articles retrieved through MEDLINE through March 2000. DATA SYNTHESIS: Deficiencies or inaccuracies persist in the sildenafil product information regarding sildenafil's effects on blood pressure; potential drug interactions with cimetidine, protease inhibitors, some antihypertensive drugs, alcohol, and drugs that may competitively inhibit cytochrome P450 pathways; and recommended sildenafil doses for older patients. CONCLUSIONS: For physicians to practice optimal therapeutics, adequate, clinically relevant drug information is required. Several brief changes and additions in the sildenafil product information would assist physicians in making therapeutic decisions regarding the use of sildenafil in a very diverse patient population and in avoiding preventable adverse events. PMID- 11261533 TI - Glucose disorders associated with HIV and its drug therapy. AB - OBJECTIVE: To review the impact that factors such as HIV infection, antiretrovirals, and other commonly used drug therapies have on glucose metabolism in HIV-infected patients. DATA SOURCES: Pertinent literature was identified via a MEDLINE search from 1980 to April 2000 and through secondary sources (abstracts presented at recent scientific meetings, manufacturers' package inserts). The key words used were antiretroviral therapy, HIV infection, insulin resistance, and metabolic abnormalities. All information deemed relevant to evaluate the impact that HIV infection and drug therapy have on glucose metabolism in HIV-infected patients was included. DATA SYNTHESIS: The viral burden and stress that are present in HIV-infected patients elicit a complex hormonal and immunologic response that may alter various biochemical pathways, including glucose metabolism. Although rare before the era of potent antiretroviral therapy, insulin resistance has now been described as an important component of the lipodystrophy syndrome. The complex and multifactorial nature of glucose metabolism dysregulation makes management of hyperglycemia or diabetes mellitus challenging in HIV-infected patients. In such a context, a set of recommendations was developed to guide practitioners in assessing, treating, and monitoring hyperglycemia or diabetes mellitus in HIV-infected patients. CONCLUSIONS: Alterations of glucose metabolism observed in HIV-infected patients are more frequent since the introduction of potent antiretroviral therapy. Although the etiology of such abnormalities remains unknown, protease inhibitors and, to a lesser extent, nucleoside reverse transcriptase inhibitors are believed to participate in their pathogenic mechanisms. Because of similarities to the pathogenesis of diabetes mellitus, management of antiretroviral-induced hyperglycemia could follow that the recommendations of the American Diabetes Association, with special considerations for monitoring patients with HIV infection. Future studies of altered glucose metabolism in HIV-infected patients should focus on understanding the precise mechanism or causes of this complication so that preventive and therapeutic guidelines can be further evaluated. PMID- 11261534 TI - Religion and coping with serious medical illness. AB - OBJECTIVE: To review and discuss some of the research published in the last several decades that has addressed the role that religion plays in helping patients cope with serious medical illness. DATA SOURCES: Although this is not a systematic review of the literature, it provides a sampling of the studies that have examined the relationship between religious involvement, coping with illness, and health outcomes. This sampling of studies reflects the findings of a much larger systematic review of research (MEDLINE, Current Contents, Psychlit, Soclit, HealthStar, Cancerlit, CINAHL, and others) during the past century that was recently completed by the authors. DATA EXTRACTION: Epidemiologic studies published in the English-language literature were reviewed and discussed. DATA SYNTHESIS: A number of well-designed cross-sectional and prospective studies have examined the relationship between religious beliefs and activities and adaptation to physical illness in patients with general medical conditions, neurologic disorders, heart disease, renal failure, AIDS, and a host of other physical disorders. This review demonstrates the widespread use of religion in coping with medical illness and provides circumstantial evidence for the possible benefits of this lifestyle factor. CONCLUSIONS: When people become physically ill, many rely heavily on religious beliefs and practices to relieve stress, retain a sense of control, and maintain hope and their sense of meaning and purpose in life. Religious involvement appears to enable the sick, particularly those with serious and disabling medical illness, to cope better and experience psychological growth from their negative health experiences, rather than be defeated or overcome by them. PMID- 11261535 TI - Use of sucralfate in renal failure. AB - OBJECTIVE: To assess the potential for the development of aluminum toxicity in patients with renal insufficiency or chronic renal failure who are taking sucralfate. DATA SOURCES: Clinical literature accessed through MEDLINE (1966 December 1999) and International Pharmaceutical Abstracts (1970-December 1999). Key search terms included sucralfate, renal failure, renal insufficiency, and end stage renal disease. DATA SYNTHESIS: Urinary excretion is an important route of elimination for systemically absorbed aluminum. Accumulation of aluminum in patients with impaired renal function may lead to significant toxicity. A potential source of aluminum is the antiulcer medication sucralfate. Studies and case reports evaluating the use and toxicity of sucralfate in patients with normal renal function, as well as those with renal failure or renal insufficiency, were reviewed. CONCLUSIONS: Aluminum accumulation and toxicity have been reported with the use of sucralfate in patients with compromised renal function. The risk of toxicity most likely represents a long-term complication of sucralfate use in this patient population. Toxicity may be enhanced by concurrent use of other aluminum-containing medications, such as phosphate binders or antidiarrheal preparations. These medications, in addition to sucralfate, should be avoided if possible in patients with end-stage renal disease. Patients with renal failure or renal insufficiency who are undergoing prolonged sucralfate therapy should be monitored for potential signs of aluminum toxicity. PMID- 11261536 TI - Corticotropin for acute management of gout. AB - OBJECTIVE: To evaluate safety and efficacy of corticotropin for acute gout. DATA SOURCES: Clinical literature was accessed through MEDLINE (1966-August 2000). Key search terms included gout, ACTH, adrenocorticotropic hormone, and corticotropin. DATA SYNTHESIS: Joint pain and local signs of inflammation characterize gout. Acutely, colchicine and nonsteroidal antiinflammatory drugs (NSAIDs) are first line therapy. Adverse effects or concomitant diseases limit therapy and necessitate alternative options. An evaluation of studies involving corticotropin was conducted. CONCLUSIONS: Corticotropin alone or in combination with colchicine was more rapidly effective and associated with fewer adverse effects than indomethacin. This regimen may be considered an alternative, especially for patients with medical problems in which other regimens are contraindicated. PMID- 11261538 TI - Mifepristone--controversy, beliefs, and politics--issues for everyone. PMID- 11261537 TI - Fluconazole dose recommendation in urinary tract infection. AB - OBJECTIVE: To identify the most appropriate dose of fluconazole for the treatment of symptomatic fungal urinary tract infection (UTI). DATA SOURCES: Primary literature identified through MEDLINE (1990-June 2000). Key search terms included fluconazole and urinary tract infection. DATA SYNTHESIS: Fluconazole is approved for the treatment of candidal UTIs, but dosage recommendations are not consistent. An evaluation of clinical studies of fluconazole for the treatment of candidal UTI was performed. CONCLUSIONS: Questions remain about the optimal dosing of fluconazole, including the most appropriate dose in non-albicans species of candida as well as the optimal duration of therapy. Until further studies are performed, a fluconazole 200-mg loading dose followed by 100 mg/d for at least four days appears to be the most appropriate dose for the treatment of symptomatic candidal UTI in patients without systemic fungal infection or severe renal failure. PMID- 11261539 TI - Mifepristone--a boom or a bust? PMID- 11261540 TI - Mifepristone: less obvious adverse effects. PMID- 11261541 TI - Increase in international normalized ratio associated with smoking cessation. PMID- 11261542 TI - Bromide intoxication and pseudohyperchloremia. PMID- 11261543 TI - Immunohistochemical localisation of ET-1 and eNOS in lymphatic stomata of the porcine broad ligament of the uterus. AB - Immunohistochemical localization and distribution of endothelin (ET-1) and nitric oxide synthase (eNOS) were investigated in lymphatic stomata in areas of their special accumulation in the porcine broad ligament during the estrous cycle. The study was performed using polyclonal antibody for ET-1 and monoclonal antibody for eNOS. ET-1 and eNOS immunoreactivities were demonstrated in some thin endothelial lymphatic lacuna walls throughout the estrous cycle. In the mesothelial cell layer, ET-1 and eNOS were detected only in stomata-related cuboidal mesothelial cells, however, the intensity of the immunostaining and distribution of the positive cells varied during the cycle. These results suggest that ET-1 and eNOS can play a role in mechanisms regulating the tone of lymphatic stomata during the absorption and passage of fluids, particles and cells from the peritoneal cavity to lymphatic vessels in the porcine broad ligament. PMID- 11261544 TI - Correlation of myofibrillar ATPase activity and myosin heavy chain content in ventricular and atrial myocardium of fish heart. AB - In the fish heart, ventricular and atrial muscles contain different isoforms of native myosin and myosin heavy chain (MyHC) but the significance of this diversity is still not known. We have analysed ventricular and atrial myocardium of six freshwater fish species (goldfish, roach, bream, rudd, perch and pike perch) using histochemical staining for myofibrillar ATPase activity as well as non-denaturing and SDS gel electrophoreses for native myosin and MyHC content. In the range of fish species studied, the intensity of ATPase reaction was higher in the atrial myocardium than in the ventricular myocardium and the composition of native myosin isoforms differed between these two muscles. The MyHC content in the cardiac muscle showed some species-related differences. In the goldfish, both atrial and ventricular cardiac muscle contained electrophoretically similar MyHC. In the other fish species, however, the ventricular myocardium showed electrophoretically faster MyHC than that present in the atrial myocardium. These results indicate that there are consistent and characteristic species-related differences between the ventricular and atrial muscles at the level of ATPase staining and the type of MyHC expressed. The findings suggest that fish ventricular and atrial muscles may differ in their contractile properties. PMID- 11261545 TI - Influence of culture medium pH on cytotoxicity of CD95L in vitro. AB - CD95L belongs to the tumor necrosis factor-alpha (TNF-alpha) family, the members of which induce apoptosis by activation of their specific receptors. However, there are a few publications suggesting that two of these factors, TNF-alpha and TNF-beta, are able to reveal cytotoxic effect in pH-dependent manner. Therefore we investigated, whether CD95L may also reveal pH-dependent cytotoxicity. We analyzed influence of CD95L on U937 and K562 human cell lines at pH 5.1 and pH 7.4 using radioactive chromium release and tetrazolium salt (MTT) reduction assays. Expression of CD95 in both cell lines was estimated using RNase Protection Assay and FACS analysis. It has been found that short incubation of cells at pH 5.1 did not visibly affect their viability, as measured after 16 or 20 h. Incubation of U937 with CD95L at pH 7.4 resulted in a dose-dependent cell cytotoxicity. The effect was significantly augmented by incubation of cells with CD95L at pH 5.1. K562 cell line was resistant to CD95L at pH 7.4. This result correlated with the lack of CD95 expression in K562 cells. However, incubation at pH 5.1 resulted in a sensitization of K562 cells to CD95L. Our results suggest that CD95L, similarly to TNF-alpha, is able to reveal its cytotoxic activity in a receptor-independent manner and this activity strongly depends on pH of the environment. PMID- 11261546 TI - Autophagy-related vacuoles in mouse gallbladder epithelium. AB - The mouse gallbladder epithelial cells contain very heterogeneous vacuolar population. In an attempt to classify these vacuoles we identified NADPase and TPPase activity as well as the location of HRP which is used as the endocytotic marker. The results of the present study show that the vacuoles can be classified into three categories: (1) the vacuoles predominantly containing loose membrane coils related to the nascent autophagic vacuoles, (2) vacuoles containing densely packed membranes and exhibiting a positive HRP reaction, indicating the convergence of endocytotic and autophagic pathway, and (3) vacuoles composed of degraded membrane structures and containing the reaction product of NADPase activity, showing that the fusion of the lysosomes with the autophagosome endosome took place. The highly developed cis, medial and trans Golgi compartments reflect the biosynthetic and endocytotic activity of the gallbladder epithelium. PMID- 11261547 TI - Idiopathic infertility in married couples in the light of cytogenetic analysis and sperm penetration assay. AB - We have selected 47 couples with unexplained infertility in order to analyse a possible link between sperm dysfunction studied in males in in vitro conditions and karyotype analysis of somatic cells. In order to identify so called "idiopathically infertile" couples we had to exclude any change in reproductive organs in both partners or in spermiogram which would qualify any of spouses into known category of infertility. We have revealed chromosome aberrations (translocations and marker chromosomes) in 19% of infertile males and in 6% of infertile females. Idiopathically infertile males had an overall decreased ability of sperm function (measured by proportion of penetrated hamster oocytes by human sperm) in comparison to fertile controls, however, still well placed within physiological range of values. Only sperm from a patient with identified translocation was clearly below the normal level of penetration (20% of penetrated oocytes), however, also the patients with revealed chromosome variant polymorphisms presented statistically lower values of penetration in comparison to fertile controls (39% vs 57%, p<0.05). On the contrary, patients with marker chromosomes did not exhibit affected sperm function. It can be speculated that only particular chromosome aberration in group of idiopathically infertile males may affect sperm functional capability (measured in vitro), however, the intragonadal genetic analysis has to be recommended in order to confirm such a causative link. PMID- 11261548 TI - Studies on the influence of aneuploidy of spermatozoa obtained from patients with oligozoospermia on their structure. AB - The presence of aneuploidy in spermatozoa influences their biological characteristics, especially their ability to fertilise the ovum. The aim of the present study was to investigate if aneuploidy is accompanied by any changes in the morphology of spermatozoa in oligozoospermic patients. For this purpose, the percentage of aneuploid cells in sperm and the correlation between the specific morphological forms of spermatozoa and aneuploidy were evaluated. The study proved a negative correlation between DNA content of aneuploid and normal spermatozoa. A weak positive correlation was demonstrated between the presence of aneuploid spermatozoa and DNA content of spermatozoa with large heads. No such correlations could be detected for DNA content of the remaining morphological forms of spermatozoa. Thus, men with a lowered number of spermatozoa and/or with abnormal spermatozoal morphology should have their spermatozoal DNA content tested in order to evaluate the degree of aneuploidy, especially in cases where in vitro fertilisation is intended. PMID- 11261549 TI - High molecular weight dextran sulphate enhances the activity of NF-kappaB regulated promoter in monocytic cell line. AB - It has been known that lipopolysaccharides (LPS) as well as superantigens increase activity of transcription factor NF-kappaB in monocyte-derived macrophages and monocytic cell lines. To examine the mitogenic effect of high molecular weight dextran sulphate (HMDS), the monocytic cell line THP-1 was transfected with a plasmid vector containing chloramphenicol acetyl transferase (CAT) reporter gene linked to the HIV-1 long terminal repeat (LTR) promoter, that is regulated by the transcription factor NF-kappaB. We have demonstrated that HMDS, similar to bacterial LPS, increases the expression of CAT reporter gene, indicating increased activity of NF-kappaB. PMID- 11261550 TI - The expression of androgen receptor, cytochrome P450 aromatase and FSH receptor mRNA in the porcine ovary. AB - The aim of this study was to visualize the expression of androgen receptor, cytochrome P450 aromatase and FSH receptor mRNAs in various structures of porcine ovary. Porcine ovaries were frozen in liquid nitrogen, and 8 microm sections were prepared for in situ hybridization. In the small, medium and large antral follicles as well as in early, midluteal and regressing corpora lutea, mRNAs for androgen receptor, P450 aromatase and FSH receptor were detected. In small antral follicles high levels of mRNAs for androgen and FSH receptors were observed, mainly in the granulosa layer, while mRNA expression for P450 aromatase was negligible. As follicles grew, amount of mRNAs for androgen receptor and FSH receptor decreased, and that for P450 aromatase increased. Small amounts of androgen receptor mRNA were also present in corpora lutea at all examined stages. P450 aromatase mRNA was not detected in early and midluteal corpora lutea. However, regressing corpus luteum showed a weak expression of aromatase mRNA. PMID- 11261551 TI - Description of a growth simulation model for predicting the effect of diet on broiler composition and growth. AB - The growth simulation program, BPHL (Bromley Park Hatcheries Limited), is a computerized, mechanistic, deterministic and dynamic approach to the evaluation of the effects of diet on broiler carcass composition and growth. Daily growth is simulated with information on the initial age and live weight of the bird, number of days over which the diet is to be fed, protein and amino acid densities of the diet, dietary metabolizable energy, and whether feed intake is to be simulated or data provided. Output provides information on a daily basis with respect to daily and accumulated deposition and current bird status for protein, fat, water, and ash body content. Carcass weight, feather weight, live weight, feed eaten, feed deprivation heat loss, limiting amino acids, food conversion ratio, and percentage carcass fat are also provided. The approach employs empirically derived first-limiting amino acid coefficients relating to accretion efficiency and dietary concentration to define limits of protein retention, uses mathematical expressions describing feed intake and heat loss trajectories as datum patterns prescriptive of the strain, introduces calibration as a device for improving correspondence between simulated and field performance, and relies on an assumption that deviations to the datum patterns of food intake and heat output caused by strain and environmental factors can be duplicated by simple multiplers acting on the expressions. The program may be used as a tool for exploring the predicted effect of specific dietary characteristics and strain parameters on growth, body composition, and performance. PMID- 11261552 TI - Minimal number of chicken daily growth velocities for artificial neural network detection of pulmonary hypertension syndrome (PHS). AB - Previously, evaluation of the first 2 wk of daily growth velocity with an artificial neural network (ANN) provided an effective noninvasive approach for predicting the susceptibility of broilers to pulmonary hypertension syndrome (PHS). This study was conducted to define the minimum number of days of growth data and the type of ANN required for the best prediction of PHS susceptibility. Four experiments were conducted in which broilers were weighed daily at 0800 h. In Experiment 1, Hubbard male broilers were reared to 50 d of age, with 13 developing PHS and 33 remaining normal (N), for a PHS:N ratio of 13:33. In Experiment 2, ANAK broilers were exposed to cool temperatures (16 to 17 C) from 17 to 42 d of age, resulting in a PHS:N ratio of 16:46 for males. In Experiments 3 and 4, Hubbard male and female chicks from a base population and a PHS resistant line were exposed to cool temperatures from 17 to 42 d (Experiment 3) or 49 d of age (Experiment 4). The PHS:N ratios were 40:68 for males and 6:96 for females in Experiment 3 and 26:91 for males and 10:58 for females in Experiment 4. Four ANN, back propagation (BP3), Ward back propagation (WardBP), probabilistic (PNN), and general regression (GRNN), were evaluated for their ability to predict PHS in the shortest number of days based on daily growth velocities (BWd+1-BWd). A 100% prediction of PHS and N birds was considered the criterion of success. Starting with 14 d of data, each ANN was trained on daily growth velocity, and the number of predictive days was reduced with each run of the ANN. The best ANN was a GRNN, which correctly diagnosed PHS and N male broilers on 4 and 6 d of growth velocity data for Experiments 1 and 2, respectively. The results were poorer with the BP3, WardBP, and PNN. The diagnostic ability of the neural network was not consistent over all four experiments. In Experiment 2, a minimum of 6 d was required for 100% PHS detection for males. In Experiment 3, the best diagnostic value for males was 93% PHS detection and 100% N detection at 15 d. For females, the 100% PHS detection occurred at a minimum of 8 d. In Experiment 4, males had 100% PHS and N detection at a minimum of 11 d. Females had a 100% PHS and N detection at a minimum of 10 d. An attempt to build a single neural network that would detect PHS susceptibility in Hubbard (Experiment 1) and ANAK (Experiment 2) broilers was unsuccessful. The application (validation) of neural networks between experiments also was not successful (data not presented). However, these studies demonstrate that within a breed or line reared under similar selection pressures for ascites, a GRNN based on the first 14 d of growth velocity can detect, with at least 93% accuracy, broilers susceptible to PHS. PMID- 11261554 TI - Plumage condition and health of aviary-kept hens fed mash or crumbled pellets. AB - In the present experiment, we evaluated the effects on plumage condition and health of feeding a mash or a crumbled diet to two hybrids of laying hens in an aviary system. The two diets had the same composition and calculated nutrient content. A total of 3,204 birds was studied from 20 to 80 wk of age. Two hybrids, Lohmann Selected Leghorn and SLU-1329 (two line crosses of Leghorn and Rhode Island Red), were housed in six pens each of an aviary system with groups of 269 and 265 birds, respectively. There were three replicates per treatment (diet x hybrid). Diet generally had little effect on plumage condition, health, and tonic immobility. However, birds fed the crumbled diet had significantly fewer problems with bumble foot than those fed the mash diet. Hybrids reacted differently in most traits studied; SLU-1329 had better health scores but more problems with cannibalism and salpingitis than Lohmann Selected Leghorns, whereas the reverse was found in the proportion of cases with coccidiosis. The hybrid differences found underline the importance of genotype. PMID- 11261553 TI - Effects of the ionophore anticoccidial semduramicin on broiler breeders. AB - Three experiments were conducted to assess the effects on broiler breeders of contamination of feed with the ionophore anticoccidial semduramicin. In Experiment 1, individually caged females received 0, 12.5, or 25 mg/kg diet for 3 wk from 48 to 50 wk of age. In Experiment 2, males and females in floor pens received 0, 12.5, or 25 mg/kg diet for 3 wk from 63 to 65 wk of age. In Experiment 3, individually caged males and females received 0, 3, 6, or 25 mg/kg diet for 1 wk at 31 wk of age and were mated by artificial insemination. There was a dose-related decrease in cumulative egg production and percentage shell in Experiment 1 after more than 1 wk exposure, but these effects were not observed in the other experiments. There was a decrease in cumulative fertile hatchability and a dose-related decrease after 3 wk exposure due to an increase in early embryonic mortality in Experiment 2, but these changes were not observed during the 1-wk exposure in Experiment 3. The data show that adverse effects of semduramicin require greater than 1 wk of exposure to be evident. PMID- 11261555 TI - Breeder age influences embryogenesis in broiler hatching eggs. AB - The effects of dietary fat and broiler breeder age on egg and embryo characteristics during incubation were investigated. Breeders were fed diets containing no added fat or 3.0% added poultry fat (PF) for peak energy intakes of 430 and 467 kcal/hen day (pC/d), or 1.5% PF or 3.0% corn oil at 449 pC/d. Feeding of diets was initiated at 22 wk, and eggs were collected for incubation at 27 and 36 wk of age. Percentage incubational egg weight loss was determined between day of set and Days 6, 12, and 18. Percentage wet and dry embryo weights, embryo moisture content, and eggshell weights were determined at 6, 12, and 18 d of incubation. Percentage yolk sac weight and wet and dry liver weights and moisture content were determined on Days 12 and 18. Percentage gall bladder weight was determined on Day 18. There were no observed effects due to breeder diet. However, eggshell weight at Days 6, 12, and 18 was higher in 27-wk-old hens compared with 36-wk-old hens. Conversely, egg weight loss between Day 0 and Days 6, 12, and 18 and yolk sac weight across Days 12 and 18 of incubation were lower in eggs at 27 wk of age compared with 36 wk. At Day 18, dry embryo weight was higher and wet liver weight was lower at 27 wk compared with 36 wk. A slower rate of DM accumulation in embryos at Week 36 compared to Week 27 was associated with increased incubational water loss and decreased embryo moisture content, eggshell percentage, and yolk sac absorption rate. These data demonstrate that changes in eggshell characteristics with broiler breeder age can alone impact yolk uptake, growth, and body composition in subsequent embryos. PMID- 11261556 TI - Effect of lactic acid administration in the drinking water during preslaughter feed withdrawal on Salmonella and Campylobacter contamination of broilers. AB - The crop is a known source of Salmonella and Campylobacter contamination. We evaluated the use of selected organic acids (0.5% acetic, lactic, or formic) in drinking water during a simulated 8-h pretransport feed withdrawal (FW). Salmonella typhimurium was recovered from 53/100 control crops and from 45/100 of crops from acetic acid-treated broilers. However, treatment with lactic acid (31/100) or formic acid (28/76) caused significant (P < 0.05) reduction in incidence. Reductions of recovered incidence were also associated with reduced numbers of S. typhimurium recovered (e.g., control, log 1.45 cfu/crop; lactic acid, 0.79 cfu/crop). In an additional commercial farm study, broilers were provided 0.44% lactic acid during a 10-h FW (4 h on the farm and 6 h transport) and pre-FW crop, post-FW crop, and pre-chill carcass wash samples were collected for Campylobacter and Salmonella detection. Crop contamination with Salmonella was significantly reduced by lactic acid treatment (6/175) as compared with controls (29/175). Importantly, Salmonella isolation incidence in prechill carcass rinses was significantly reduced by 52.4% with the use of lactic acid (26/175 vs. 55/176). Crop contamination with Campylobacter was significantly reduced by lactic acid treatment (62.3%) as compared with the controls (85.1%). Lactic acid also reduced the incidence of Campylobacter found on pre-chill carcass rinses by 14.7% compared with the controls. These studies suggest that incorporation of lactic acid in the drinking water during pretransport FW may reduce Salmonella and Campylobacter contamination of crops and broiler carcasses at processing. PMID- 11261557 TI - Genetic variation among chicken lines and mammalian species in specific genes. AB - Thirteen gene-specific primer sets provided by the U.S. Poultry Genome Coordinators were used to investigate DNA polymorphisms between two highly inbred chicken lines of Leghorn and Fayoumi origin. Nucleotide and predicted amino acid sequences were then compared among these chicken lines and the Genbank sequences of chicken, mouse, and human. The following genes were selected as candidates for immune response or transcription activation: B2M, DAD1, IAP1, IL2, IREB1, LAP18, MAFL, POU1F1, RREB1, TAD, TBP1, TCRG, and ZOV3. Total cDNA was obtained from the spleens of Leghorn and Fayoumi lines by reverse transcriptase-polymerase chain reaction (PCR) and was used as a template to PCR-amplify gene-specific products. All primers except POU1F1 and TCRG generated single PCR products of the predicted 325- to 667-bp size, confirming the efficacy of these gene-specific primers in the chicken. Three and seven of the 11 amplified gene fragments yielded line specific nucleotide polymorphisms between the Leghorn and Fayoumi sequences and between the Leghorn and Genbank chicken sequences respectively. Similarities between inbred Leghorn and mammalian species were 36 to 86% for nucleotides and 25 to 96% for predicted amino acid sequence. The polymorphisms of some gene fragments between the Leghorn and Fayoumi lines will allow for investigation of associations of these genes with immune response and other biological traits. PMID- 11261558 TI - Sodium and chloride requirements of growing broiler chickens (twenty-one to forty two days of age) fed corn-soybean diets. AB - Two trials were conducted to determine Na+ and Cl- nutritional requirements and dietary electrolyte balance (DEB) and its effects on acid-base balance, litter moisture, and incidence of tibial dyschondroplasia (TD) in broiler chickens during the growing period. Cobb broilers were distributed in a completely randomized design (30 pens) with six treatments, five replicates, and 50 birds per experimental unit at 21 d of age. Treatments used in both trials were a basal diet with 0.10% Na+ (Trial 1) or Cl- (Trial 2) supplemented to result in diets with Na+ or Cl- levels of 0.10, 0.15, 0.20, 0.25, 0.30, and 0.35%. In the first trial, the results indicated an optimum Na+ requirement of 0.15%. The Na+ levels, obtained with supplemental NaHCO3, did not affect blood gas parameters and TD incidence. Litter moisture increased linearly with Na+ levels. In the second trial, the Cl- requirement was estimated at 0.23%. Increasing Cl- levels, provided by NaCl with NaHCO3 to balance Na+, caused a linear effect (P < or = 0.01) on blood gas parameters, with an estimated equilibrium at 0.19% dietary Cl . No effect (P > or = 0.05) of Cl- levels on litter moisture was observed. The hypertrophic area of growth plate in the proximal tibiotarsus increased with Cl- levels (P < or = 0.001). A nonlinear model describes this response. The best dietary electrolyte balance (DEB) was between 250 to 261 mEq/kg in Trial 1 and 249 to 257 mEq/kg in Trial 2. We concluded that the Na+ requirement was 0.15%, and the Cl- requirement was 0.23% for maximum performance of growing chickens between 21 and 42 d of age, and the best DEB was between 249 and 261 mEq/kg. PMID- 11261560 TI - In vitro evaluation of nonstarch polysaccharide digestibility of feed ingredients by enzymes. AB - Some of the commonly used feed ingredients for poultry (corn, sorghum, finger millet, deoiled ricebran, soybean meal, peanut meal, sunflower meal, and rapeseed meal) were screened for pentosans, cellulose, pectin, and total nonstarch polysaccharides. The ingredient in vitro digestibilities by enzymes were evaluated. Cereal samples screened contained mainly pentosans. Pectin content was rich in oilseed meals. Sunflower meal, soybean meal, deoiled rice bran, and a broiler starter diet were subjected to a two-stage in vitro digestion assay with three different enzyme mixtures viz., Enzyme-I (xylanase + cellulase from Trichoderma viridae), Enzyme-II (xylanase + cellulase + beta-glucanase from Huminicola insolens), and Enzyme-III (xylanase + cellulase + pectinase + beta glucanase from Aspergillus aculeatus) by incubating 0.1 g of the sample with 3 mL of a pepsin-HCl mixture (2,000 U pepsin/mL of 0.1N HCl) for 45 min to simulate the peptic phase of bird digestion. A pancreatin-NaHCO3 mixture (2 mg pancreatin/mL of 1 M NaHCO3) was used for 2 h at 40 C to simulate the pancreatic phase. Digestibility was assessed by measuring the relative viscosity of the digesta supernatent and the total sugars released. Enzyme-I produced the least relative viscosity and highest total sugars in sunflower meal, deoiled rice bran, and broiler starter diet, whereas Enzyme-III was very effective in soybean meal subjected to in vitro digestion. The assay was a convenient and rapid method of screening for effective and stable enzymes. PMID- 11261559 TI - Oral treatment of mule ducks with arsenicals for inducing fatty liver. AB - The aim of this study was to determine the dosage and the compounds of arsenic that induce fatty liver in mule ducks and also to investigate their effects on tissue residues. One hundred four ducks, 8 wk old, were randomly selected for one of six dietary treatments in Trial 1 or one of seven dietary treatments in Trial 2. Different levels of roxarsone were administrated: 0, 10, 20, 30, 40, or 50 mg/d, respectively, in Trial 1. In Trial 2, the experimental treatments were of the same level (11.36 mg/d) with different sources of arsenic that included the control without As, roxarsone (3-nitro-4-hydroxyphenylarsonic acid), arsanilic acid, phenylarsonic acid, O-nitro-phenylarsonic acid, As2O3, or As2O5. Both trials lasted 3 wk, with 1 wk on the treatment followed by 2 wk of withdrawal. Results in Trial 1 showed that a dose of 40 mg roxarsone/d increased liver weight and caused fatty liver, whereas administration of 50 mg/d was lethal. In Trial 2, administration of arsenic (11.36 mg/d) for 1 wk significantly depressed feed intake in the roxarsone, As2O3, and As2O5 groups (P < 0.05), whereas the treatment significantly decreased only live weight gain in the roxarsone group (P < 0.05). Administration of roxarsone alone increased (P < 0.05) serum cholesterol (CHOL), albumin (ALB), and total protein (TP), whereas only As2O3 among treatments significantly decreased serum triacylglycerol (TG) concentration (P < 0.05). In the roxarsone, arsanilic acid, and phenylarsonic acid groups, serum high density lipoprotein (HDL) decreased to a greater extent (P < 0.05), and arsanilic acid treatment significantly increased the very low density lipoprotein (VLDL) (P < 0.05). After 2 wk of withdrawal, liver weights and relative liver weights were heavier in the treatment groups of roxarsone, As2O3, and As2O5 as compared to the control (P < 0.05). Levels of CHOL, TG, TP, and ALB were significantly higher in the groups treated with As2O3 or As2O5 as compared to the control (P < 0.05). The roxarsone and arsanilic acid treatments significantly decreased HDL and increased VLDL in plasma (P < 0.05). The creatine kinase (CK) level in the roxarsone, As2O3, and As2O5 groups was significantly higher compared to the control group (P < 0.05). Among the As sources, roxarsone, As2O3, and As2O5 caused fatty liver in mule ducks. PMID- 11261561 TI - Studies on feeding peanut meal as a protein source for broiler chickens. AB - Four experiments were conducted to compare the performance of broilers fed soybean meal (SBM) versus peanut meal (PNM) as protein sources. Ross x Ross 208 broiler chickens were placed in battery brooders (Experiments 1 to 3, four replicates of 8 chicks per treatment) and floor pens (Experiment 4, four replicates of 34 chicks per treatment). In Experiment 1, addition of 0, 0.1, 0.2, and 0.3% Thr to a corn-PNM-based diet increased 0 to 18 d BW gain (BWG; 0.374c vs. 0.495b vs. 0.508b vs. 0.508b kg, respectively) and decreased feed conversion ratio (FCR; 2.09c vs. 1.63b vs. vs. 1.54b vs. 1.54b g/g, respectively) compared to the corn-SBM-based control diet (BWG = 0.593a and FCR = 1.36a). In Experiment 2, diets were formulated with the same amino acid minimums, and as the percentage of PNM increased in the diets (0, 10, 20, and 32%), BWG decreased (0.560a vs. 0.532a vs. 0.521a vs. 0.458b kg, respectively) and FCR increased (1.72b vs. 1.71b vs. 1.79bc vs. 1.86c g/g, respectively). In Experiment 3, addition of Thr to a corn-PNM-based diet increased BWG (-Thr = 0.284c vs. +Thr = 0.397b kg) and decreased FCR (-Thr = 1.60b vs. +Thr = 1.54b g/g). The BWG and FCR were best for the corn-SBM-based control diet (0.499a kg and 1.38a g/g, respectively). In Experiment 4, during the growing period (18 to 42 d), significant interactions occurred between protein source (PNM vs. SBM) and protein level (16 and 20% vs. 24%) for BW and FCR but not for carcass, breast, or leg quarter yield or fat pad weights (P < 0.05) at 42 d of age. Technical (not economic) performance of birds fed PNM was similar to SBM at the highest protein levels fed. PNM could be used as a protein source for broilers under appropriate economic conditions. PMID- 11261562 TI - The use of near-infrared reflectance spectroscopy to predict the moisture, nitrogen, calcium, total phosphorus, gross energy, and phytate phosphorus contents of broiler excreta. AB - One hundred forty-three broiler chick excreta samples were obtained from previous experiments dealing with phytate phosphorus utilization. The air-dried samples were ground in a Cyclotech 1093 sample mill and analyzed for the following: moisture, N, Ca, energy, total P, and phytate P. By chemical assay, the sample compositions were moisture: mean = 9.62, SD = 1.27% (range = 7.37-13.59); N: mean = 5.31, SD = 0.37% (range = 4.28 to 6.48); Ca: mean = 1.66, SD = 0.32% (range = 0.85 to 2.6); total P: mean = 1.13, SD = 0.28% (range = 0.66 to 1.75); gross energy: mean = 3,560, SD = 120 kcal/kg (range = 3,309 to 3,882); phytate P: mean = 0.63, SD = 0.17% (range = 0.32 to 0.97). The samples were scanned in a Feed & Forage Analyzer Model 5000 with near-infrared reflectance spectroscopy (NIRS)-2 Software. One hundred twenty-three samples were used to create the calibration curves (20 randomly selected samples were set aside for validating the calibration). The combination of math treatments and scatter corrections that provided the best standard error of cross validation (and its correlation coefficient) was chosen for the standard curves. The coefficients of determination (R2) were moisture, 0.96; N, 0.88; Ca, 0.84; total P, 0.91; gross energy, 0.86; and phytate P, 0.86. The standard errors of prediction were moisture, 0.342%; N, 0.193%; Ca, 0.143%; total P, 0.134%; gross energy, 74.66 kcal; and phytate P, 0.91%. We concluded that it is possible to predict the moisture, N, Ca, gross energy, total P, and phytate P in broiler excreta by using NIRS. PMID- 11261563 TI - Influence of source and ratio of xanthophyll pigments on broiler chicken pigmentation and performance. AB - One experiment was conducted using 960 1-d-old, sexed broilers of Ross 308 strain from 1 to 43 d to evaluate if one type of chemically isomerized marigold with 25% of xanthophylls as zeaxanthin (SME-25) could produce pigmentation equivalent to the current addition of conventional marigold with 10% of xanthophylls as zeaxanthin (SME-10) plus canthaxanthin (CTX) in practical broiler diets (maize wheat-soybean). Birds were allocated in 32 pens, in a randomized complete block design (four blocks x four treatments). The treatments consisted of a nonpigmented control (T1), a combination of 35 ppm of yellow xanthophylls (YX) from SME-10 + 5 ppm of CTX (T2), a combination of 32 ppm of YX from SME-10 + 2 ppm of CTX (T4), and one treatment with 40 ppm of YX from a new SME-25 (T3). There were no significant treatment effects on chicken performance. All color parameters (Minolta coordinates, Roche color fan scores, Rank test) presented significant differences (P < 0.0001) because of dietary pigments on shanks and breast skin. Birds fed the SME-25 diet had less pigmentation than those fed equivalent quantities of a combination of SME-10 + CTX. The Minolta coordinate "b" measured in breast skin was a good indicator of YX content in feed, whereas the "a" coordinate measured on the shank showed a linear relationship with the dietary CTX level (r = 0.61, P < 0.0001). The same visual color classification of chickens was achieved irrespective of the rank test performed (by shank or carcass color). Lutein and zeaxanthin from the SME-25 product had lower deposition rates in skin and fat tissues than those from the SME-10 product. This finding seems to be related to the ratio of zeaxanthin stereoisomer RR (optically active) vs. RS that was found in tissues from the SME-10 product (97.8%:2.2%), whereas with SME-25 this ratio was 16.0:84.0%. These results suggest that inclusion of only the SME-25 product could not replace the current addition of SME-10 and CTX combinations. PMID- 11261564 TI - Lipid oxidation in fresh and spray-dried eggs enriched with omega3 and omega6 polyunsaturated fatty acids during storage as affected by dietary vitamin E and canthaxanthin supplementation. AB - A 2 x 2 x 2 factorial experiment was planned to study the influence of dietary fat source (linseed oil or sunflower oil) and dietary doses of alpha-tocopheryl acetate (alpha-TA) (0 or 200 mg/kg of feed) and canthaxanthin (CX) (0 or 5 mg/kg of feed) on fatty acid (FA) composition and lipid oxidation of fresh and spray dried eggs. Dietary supplementation with alpha-TA and CX modified the levels of certain long-chain polyunsaturated FA (PUFA). Lipid oxidation in fresh eggs and spray-dried eggs at 0, 6, and 12 mo of storage was measured by the lipid hydroperoxide (LHP) and TBA values. The LHP and TBA values were up to 10 times higher in spray-dried eggs than in fresh eggs. The evolution of LHP and TBA values in spray-dried eggs showed that omega3 FA-enriched eggs were more susceptible to lipid oxidation than those enriched with omega6 FA. The omega-TA supplementation increased the lipid stability of enriched eggs and was very effective throughout the storage of spray-dried eggs. On the other hand, CX supplementation did not prevent lipid oxidation in PUFA-enriched eggs. Moreover, no synergistic effect between both compounds was detected. PMID- 11261565 TI - Microbial phytase improves performance, apparent metabolizable energy, and ileal amino acid digestibility of broilers fed a lysine-deficient diet. AB - An experiment was conducted to examine the effects of adding microbial phytase (Natuphos) on the performance in broilers fed a phosphorus-adequate, lysine deficient diet. A wheat-soybean meal-sorghum-based diet, containing 1.00% lysine and 0.45% nonphytate phosphorus, was supplemented with L-lysine monochloride to provide 1.06, 1.12, or 1.18% lysine or with 125, 250, 375, 500, 750, or 1,000 phytase units (FTU)/kg diet. Each diet was fed to six pens of 10 chicks each from Day 7 to 28 posthatching. Addition of lysine to the lysine-deficient diet linearly increased (P < 0.001) weight gain and gain per feed of broilers. The response in weight gain to added phytase reached a plateau at 500 FTU/kg diet (quadratic effect, P < 0.001). Phytase had no effect on gain per feed to 250 FTU/kg diet and then increased (quadratic effect, P < 0.05) with further additions. Assuming that the observed responses in weight gain and gain per feed to added phytase were due to the release of lysine alone and by solving linear or nonlinear response equations of lysine and phytase levels, the lysine equivalency value was calculated to be 500 FTU phytase/kg diet = 0.074% lysine. Addition of increasing levels of supplemental phytase to the lysine-deficient diet improved (P < 0.001) the digestibilities of nitrogen and all amino acids. Phytase also increased the AME, and the response reached a plateau at 750 FTU/kg diet (quadratic effect, P < 0.001). These results showed that amino acid and energy responses are responsible for the performance improvements observed when phytase was added to a wheat-soybean meal-sorghum-based diet. PMID- 11261566 TI - Phase-feeding supports maximum growth performance of broiler chicks from forty three to seventy-one days of age. AB - Phase-feeding (PF) was tested to evaluate its efficacy compared with NRC recommendations for broilers. Two modified PF regimens were also tested that involved lowering amino acid requirements predicted by PF linear regression equations by 10% (PF10) and increasing the slope of the linear regression equations by 15% (PF15). Experimental diets were fed from 43 to 71 d. Broilers fed the NRC regimen were given a single diet throughout the experiment, whereas PF, PF10, and PF15 were tested with a series of four diets (43 to 50 d, 50 to 57 d, 57 to 64 d, and 64 to 71 d). At 71 d, no differences (P < 0.05) in weight gain, feed intake, or feed efficiency were observed among treatments. Intake of crude protein, digestible lysine, sulfur amino acids (SAA), and threonine were decreased (P < 0.05) by PF, PF10, and PF15 relative to that of broilers fed the NRC diet. Gain per unit digestible lysine and threonine intake were increased (P < 0.05) by PF, PF10, and PF15, and gain per unit digestible SAA intake was increased (P < 0.05) by PF10 and PF15 relative to broilers fed the NRC diet. No differences (P > 0.05) were observed in carcass, breast, wing, or leg yield, but abdominal fat was increased (P < 0.05) by the PF10 regimen relative to that of broilers fed the NRC diet. Economic analysis indicated that PF and PF10 may lower the cost of feed consumed and the cost per unit weight gain or breast yield relative to broilers fed the NRC diet. PMID- 11261567 TI - Early postmolt performance of laying hens fed a low-protein corn molt diet supplemented with spent hen meal. AB - We used a total of 504 commercial Single Comb White Leghorn hens (69 and 65 wk of age) in each of two experiments, and hens were induced to molt by feed withdrawal only. Feed withdrawal lasted for 12 and 11 d, and hens lost 26 and 25%, body weight in Experiments 1 and 2, respectively. All hens were then weighed, and seven replicate groups of 12 hens each were assigned to molt diet treatments. In Experiment 1, diets consisted of a corn basal diet (7.9% CP) or corn basal diet supplemented with 7.5 or 10% spent hen meal (SHM) each from two different sources. In Experiment 2, the corn basal diet or this diet supplemented with 5 or 10% SHM alone or 5% SHM plus Met, Lys, and Trp was evaluated. A molt diet of 16% CP corn-soybean meal was used as a positive control in both experiments. Molt diets were fed for 15 d in both experiments, at which time all hens were fed a 16% CP layer diet. Performance was measured for 8 wk following the beginning of feeding the layer diet. Feeding the low-protein corn molt diet supplemented with 5 to 10% SHM improved early postmolt egg production performance and body weight gain compared with hens fed the corn basal diet alone. The 7.5 and 10% SHM diets yielded early postmolt performance that was not significantly different (P > 0.05) from that of hens fed the high-protein (16% CP) diet. Supplementing the 5% SHM diet with amino acids generally did not significantly improve performance. The present study thus indicates that improved early postmolt performance may be achieved by supplementation of a low-protein corn molt diet with 5 to 10% SHM. PMID- 11261569 TI - The oxidation of cholesterol in the yolk of selective traditional Chinese egg products. AB - The yolks of traditional chicken egg products (Tiedan, Ludan, and Chayedan) and duck egg products (raw and cooked Xiandan, immersed and coated Pidan) were subjected to moisture, lipid, and thiobarbituric acid (TBA) determinations as well as cholesterol and cholesterol oxidation products (COP) analysis. The main COP detected for these egg products included 20-hydroxycholesterol and 7beta hydroxycholesterol, other types of COP were not detected. The contents of COP formed in traditional egg products varied, depending upon the types of egg products. The cholesterol oxidation ratio for traditional Chinese chicken egg products ranged from 1.14 to 1.75%, whereas that for traditional Chinese duck egg products ranged from 1.18 to 1.90%. Those traditional egg products that required pickling in salt or alkaline, cooking, hot air drying, and exposure to oxygen and heat all produced COP. PMID- 11261568 TI - Plasma levels of arginine, ornithine, and urea and growth performance of broilers fed supplemental L-arginine during cool temperature exposure. AB - Two experiments (Experiment 1 and 2) were conducted to evaluate growth performance, ascites mortality, and concentrations of plasma Arg, urea, and ornithine in male broilers raised in floor pens (2 x 4 factorial experiment, six pens for treatment) and exposed to cool temperatures averaging 16 C after 21 d of age. Broilers were fed low- or high-CP diets in both Experiments. In Experiment 1, Arg treatments consisted of control (no supplemental Arg); 0.15 or 0.3% supplemental Arg in the diet (low- and medium-Arg feed, respectively); and 0.3% supplemental Arg in the drinking water (Arg-water). Arginine levels were increased in Experiment 2 and consisted of the following: control (no supplemental Arg); 0.3 or 0.85% supplemental Arg in the diet (medium- and high Arg feed, respectively); and 0.6% supplemental Arg in the drinking water (Arg water). The water treatment followed a 3-d cyclic regimen, with supplemental Arg being provided for 24 h, followed by tap water for 48 h. When the broilers reached 37 d of age and all groups had consumed tap water for the previous 48 h, blood samples were collected from one bird per pen (Time 0, 0700 h); then supplemental Arg was provided in the Arg-water group, and additional blood samples were collected from the control and Arg-water groups at 3, 6, 12, and 36 h after Time 0. Plasma amino acids were analyzed using HPLC. Birds fed the high CP diet were heavier at 49 d than birds fed the low-CP diet in Experiment 1, but not in Experiment 2. No differences were found in feed conversion or ascites mortality due to CP or Arg treatments in either experiment. In both experiments, plasma Arg was similar for all groups at Time 0, but increased in the Arg-water group at 3, 6, and 12 h after Arg was provided in the water. Within 12 h after returning to tap water, plasma Arg levels of the Arg-water group did not differ from the control group. Plasma urea and ornithine were parallel to plasma Arg concentrations, and the high-CP diets resulted in higher plasma levels of urea and ornithine compared with low-CP diets. These results indicate that kidney arginase was readily activated by Arg provided in the water, resulting in an immediate increase in plasma urea and ornithine. Plasma Arg was increased significantly, but no effects were observed in ascites mortality. PMID- 11261570 TI - Effects of age, sex, and duration of postmortem aging on percentage yield of parts from broiler chicken carcasses. AB - The objective of this study was to evaluate effects of age, sex, and postmortem carcass aging duration on parts yield from broiler chickens. Two hundred twenty four mixed-sex broilers were reared under commercial-like conditions for various periods between 37 and 51 d, slaughtered, packed in ice, and then aged for 0, 2, 4, or 6 h. Mean percentage yield of thighs, drumsticks, forequarters, wings, breasts, and filets were evaluated for each rearing period, sex, and postmortem aging duration. Yield of meatier parts such as thighs, forequarters, breasts, and filets increased with birds' ages. Female carcasses produced higher percentage yields of forequarters, breasts, and filets but lower yields of drumsticks. Carcasses aged 2 h or more postmortem tended to have lower yields of forequarters, breasts, and drumsticks than did carcasses aged for shorter durations. No statistically significant interactions among age, sex, or postmortem aging duration that affected yield of parts were detected. This information is useful to integrated poultry firms wishing to optimize yield of the most commercially valuable parts. PMID- 11261571 TI - Who benefits from biopiracy? PMID- 11261573 TI - Biotransformation of cedrol by Curvularia lunata ATCC 12017. AB - Bioconversion of cedrol (1) with the Curvularia lunata was investigated in two different growth media. Five products were obtained in potato dextrose broth, whereas nine compounds were produced in a medium containing beef extract. Only three of the metabolites: 3beta-hydroxycedrol (2), 3alpha-hydroxycedrol (3) and 12-hydroxycedrol (4) were common to both media. They were also obtained as the major products in each case. Three new derivatives have also been identified. PMID- 11261572 TI - Cytosolic aldolase is a ripening related enzyme in strawberry fruits (Fragaria x ananassa). AB - Two aldolase isoenzymes have been isolated from ripe strawberry fruits (Fragaria x ananassa cv. Camarosa and Elsanta) and partially purified by DEAE anion exchange and Sephacryl size exclusion chromatography. The isoenzymes were identified as class I cytosol and plastid aldolase on the basis of their chromatographic behavior on DEAE-cellulose columns, native molecular weight, pH optimum pattern, Km value for D-fructose-1,6-bisphosphate, tendency to be inactivated by lower pH values and SDS-PAGE subunit determination of 40 and 38 kDa, respectively. Total aldolase activity and distribution of both aldolase isoenzymes was also investigated at different stages of strawberry fruit ripening. Strawberries in the green and white ripening stage showed the same ratio of the two isoenzymes as green leaves with 15 and 8% cytosol aldolase activity, respectively. During strawberry fruit development the overall total aldolase activity decreased until the pink ripening stage and then increased due to a rise of cytosol aldolase yielding up to 75% in red strawberries. A cDNA putatively encoding the cytosolic form of aldolase in strawberry was cloned during the course of this study. Both microarray and RNA gel blot analyses showed that the cytosolic aldolase gene expression is induced during ripening as detected for the cytosolic aldolase enzyme. We suggest that induction of the cytosolic aldolase both at the levels of transcription and translation might be part of a ripening related stress response in the receptacle tissue. PMID- 11261574 TI - Ripening-related changes in raspberry cell wall composition and structure. AB - Cell walls were prepared from the fruit of two cultivars of raspberry at three stages of ripening; green, white and red (ripe). The cultivars. Glen Clova and Glen Prosen, are subjectively classified, at harvest by growers, as soft and firm fruit, respectively. The cell walls were analysed for neutral sugar composition, uronic acid content, degree of methyl esterification, lignin and ferulic acid derived dehydrodimers. Solid-state 31C NMR and diffuse reflectance infrared (DRIFT) spectra were acquired for the cell wall residues. For both cultivars the progression from green to white produced minimal changes, save for a reduction in pectin. NMR analyses indicated that the solubilized pectin was acetylated. Progression to the red (ripe) stage, in both cultivars, was accompanied by a reduction in the ordered cellulose and a dramatic reduction in pectin content and the degree of methyl-esterification. Significantly, the softer fruit (Glen Clova) exhibited greater reductions in both parameters, implicating increased pectin hydrolysis, as one of the main factors contributing to the difference in firmness between the cultivars. A relative increase in cell wall-associated protein was seen at the red stage. The nature and function of the protein(s) are, as yet unknown. PMID- 11261575 TI - Formation and occurrence of dopamine-derived betacyanins. AB - In light of the fact that the main betaxanthin (miraxanthin V) and the major betacyanin (2-descarboxy-betanidin) in hairy root cultures of yellow beet (Beta vulgaris L.) are both dopamine-derived, the occurrence of similar structures for the minor betacyanins was also suggested. By HPLC comparison with the betacyanins obtained by dopamine administration to beet seedlings, enzymatic hydrolysis, LCMS and 1H NMR analyses, the minor betacyanins from hairy roots were identified as 2 descarboxy-betanin and its 6'-O-malonyl derivative. A short-term dopamine administration experiment with fodder beet seedlings revealed that the condensation step between 2-descarboxy-cyclo-Dopa and betalamic acid is the decisive reaction, followed by glucosylation and acylation. From these data a pathway for the biosynthesis of dopamine-derived betalains is proposed. Furthermore, the occurrence of these compounds in various cell and hairy root cultures as well as beet plants (Fodder and Garden Beet Group) is shown. PMID- 11261576 TI - Seasonal variation in hypericin content of Hypericum perforatum L. (St. John's Wort). AB - Hypericin and pseudohypericin, bioactive constituents in St. John's Wort (Hypericum perforatum), have been determined in the soft tops of the plant that are most likely to be browsed by foraging livestock. In two consecutive seasons, the hypericin/pseudohypericin concentration in a broad leaf biotype varied from a winter minimum of less than 100 ppm to a summer maximum approaching 3000 ppm. In contrast the narrow leaf biotype increased from similar winter values to summer maxima approaching 5000 ppm. The latter biotype was slower in returning to low levels of hypericin/pseudohypericin. PMID- 11261577 TI - The ecological and taxonomic importance of flower volatiles of Clusia species (Guttiferae). AB - The chemical composition of floral volatiles of sixteen species of Clusia (Guttiferae) belonging to four different taxonomic sections of the genus was investigated. The volatiles were extracted from fresh petals by microhydrodistillation and analysed by GC/MS. The composition of the volatiles was in part, but not always, related to the taxonomic position of the species, and to a minor extent to the type of pollinator observed on the flowers as revealed by clustering analysis. The composition of the volatile components of female and male flowers belonging to the same species (C. grandiflora, C. lanceolata, C. paralicola, C. parviflora and C. spiritu-sanctensis) was found to be almost identical. Field bioassays showed the petal extracts to be attractive to pollinating bees. PMID- 11261578 TI - Flavonoid characters contributing to the taxonomic revision of the Hebe parviflora complex. AB - Comparative flavonoid chemistry is a key element of a multidisciplinary study aimed at a revision of the genus Hebe, New Zealand's largest genus of flowering plants. One aspect of this study has been an investigation of the Hebe parviflora complex. A recent botanical paper on this topic marshalls generalised flavonoid data and morphological characters to support the recognition of two species in this complex, Hebe stenophylla and Hebe parviflora, which are clearly distinguishable from each other and from the related Hebe traversii and Hebe strictissima. A detailed study of the flavonoid chemistry and the distributional data used to support these conclusions are presented here. Six new compounds have been isolated in this study, including 6-hydroxyapigenin-7-O-beta-[2-O-beta xyloxyloside] and-7-O-beta-[2-O-beta-xyloglucoside], 6-hydroxyluteolin-7-O-beta [2-O-beta-xyloxyloside] and, luteolin-, 6-hydroxyluteolin- and 4'-O methylluteolin-7-O-beta-[6-O-beta-xyloglucoside]. Other flavonoids include apigenin and luteolin 7- and 4'- mono-, di- and possibly tri-O-glycosides, 8 hydroxyluteolin 7- and 8-O-glucosides, and kaempferol and quercetin 3-O-mono- and di-glycosides. New structure assignments are supported with detailed 1H and 13C NMR data, including HMQC and HMBC measurements. PMID- 11261579 TI - Ambuic acid, a highly functionalized cyclohexenone with antifungal activity from Pestalotiopsis spp. and Monochaetia sp. AB - Ambuic acid, a highly functionalized cyclohexenone, was isolated and characterized from Pestalotiopsis spp. and Monochaetia sp. these being biologically related endophytic fungi associated with many tropical plant species. This compound was found in representative isolates of these fungal species obtained from rainforest plants located on several continents. The relevance of ambuic acid to the biology of the association of these fungi to their host plants is also discussed. PMID- 11261580 TI - Lactone diterpenes from the aquatic plant Potamogeton natans. AB - Four lactone diterpenes and two related glucosides with a labdane skeleton have been isolated from the aquatic plant Potamogeton natans. The structures of three new compounds were determined as 19-acetoxy-20-oxo-8(17),13-ent-labdadien-15-->16 lactone, 8(17), 13-ent-labdadien-15-->16,19-->20 dilactone and 6'-acetyl-19 glucopyranosyloxy-8(17),13-ent-labdadien-15-->16 lactone, respectively, by means of spectral analysis. Antialgal assays showed inhibitory activity for some compounds. PMID- 11261581 TI - Germacranolides from Mikania guaco. AB - Fourteen novel sesquiterpene lactones of the germacranolide type have been isolated from the aerial parts of Mikania guaco: six costunolide, two melampolide and six germacra-4-trans,10(14),11(13)-trien-12.6alpha-olide derivatives. Except for one compound all the others possess a carbonyl function at C-9. Eight were obtained in the form of four isomer pairs which were difficult to separate. Structure elucidation was based on mass and ID and 2D NMR measurements. Low energy conformations were obtained by quantum mechanical calculations. Pyrrolizidine alkaloids could not be detected. PMID- 11261582 TI - Five saponins from the root bark of Aralia elata. AB - Five saponins, 3-O-[beta-D-glucopyranosyl (1-->2)-[beta-D-glucopyranosyl (1-->3)] beta-D-glucopyranosyl]-oleanolic acid 28-O-beta-D-glucopyranosyl ester (aralia saponin V), 3-O-[beta-D-glucopyranosyl (1-->2)-[beta-D-glucopyranosyl (1-->3)] beta-D-glucopyranosyl]-echinocystic acid 28-O-beta-D-glucopyranosyl ester (aralia saponin VI), 3-O-beta-D-glucopyranosyl (1-->2)-[beta-D-glucopyranosyl (1-->3)] beta-D-glucopyranosyl]-hederagenin 28-O-beta-D-glucopyranosyl ester (aralia saponin VII), 3-O-[beta-D-glucopyranosyl-(1-->3)-beta-D-glucopyranosyl-(1-->3) [beta-D-glucopyranosyl-(1-->2)]-beta-D-glucopyranosyl]-caulophyllogenin 28-O-beta D-glucopyranosyl ester (aralia-saponin VIII), 3-O-[beta-D-glucopyranosyl (1-->2) [beta-D-glucopyranosyl(1-->3)]-alpha-L-arabinopyranosyl]-hederagenin 28-O-beta-D glucopyranosyl ester (aralia-saponin IX), were isolated from the root bark of Aralia elata (Miq.) Seem., together with four known compounds. Their structures were determined on the basis of chemical and spectroscopic methods. PMID- 11261583 TI - Lepidimoic acid increases fructose 2,6-bisphosphate in Amaranthus seedlings. AB - This study examines the influence of the growth promoter, lepidimoic acid, on the level of an important cytosolic signal metabolite, fructose 2,6-bisphosphate (Fru 2,6-P2), which can activate pyrophosphatedependent:phosphofructokinase (PFP, EC 2.7.1.90), and on glycolytic metabolism in Amaranthus caudatus seedlings. Fru-2,6 P2 concentrations were respectively increased by approximately 2-, 3- and 4-fold when the seedlings were treated with 0.3, 3 and 30 mM lepidimoic acid. Exogenous lepidimoic acid also affected levels of glycolytic intermediates in the seedlings. The increase in fructose 1,6-bisphosphate and decreases in fructose 6 phosphate and glucose 6-phosphate were found in response to the elevated concentration of lepidimoic acid. These results suggest that lepidimoic acid may affect glycolytic metabolism in the Amaranthus seedlings by increasing the activity of PFP due to increasing level of Fru-2,6-P2. PMID- 11261584 TI - Monooxygenases involved in GA12 and GA14 synthesis in Gibberella fujikuroi. AB - A microsomal preparation from mycelia of the gibberellin (GA)-producing fungus Gibberella fujikuroi catalyzed the first two steps in the conversion of the biosynthetic intermediate GA12-aldehyde to gibberellic acid (GA3). [14C]GA12 Aldehyde was converted to radiolabelled GA14, the major product, together with smaller amounts of non-hydroxylated GA12. The microsomal activities required reduced pyridine nucleotides and molecular oxygen. However, GA12 and GA14 synthesis differed markedly in the preferred electron source. Formation of GA12 required NADH or NADPH, while GA14 synthesis from GA12-aldehyde occurred only with NADPH. Marked differences were also found in the activating effect of FAD. When NADPH was the reductant, the rate of GA14 synthesis was enhanced 3.5 times by 5 microM FAD while this flavin nucleotide did not alter the synthesis of GA12. In contrast, GA12 synthesis was activated 3.8 times by 50 microM FAD in the presence of NADH. Both activities were inhibited by carbon monoxide and cytochrome c. These properties suggest that the 3beta-hydroxylation of GA12 aldehyde and further oxidation of carbon 7 are catalyzed by cytochrome P-450 monooxygenases in Gibberella fujikuroi. PMID- 11261585 TI - Ferulic acid is esterified to glucuronoarabinoxylans in pineapple cell walls. AB - The ester-linkage of ferulic acid (mainly E) to polysaccharides in primary cell walls of pineapple fruit (Ananas comosus) (Bromeliaceae) was investigated by treating a cell-wall preparation with 'Driselase' which contains a mixture of endo- and exo-glycanases, but no hydroxycinnamoyl esterase activity. The most abundant feruloyl oligosaccharide released was O-[5-O-(E-feruloyl)-alpha-L arabinofuranosyl](1-->3)-O-beta-D-xylopyranosyl-(1-->4)-D-xylopyranose (FAXX). This indicated that the ferulic acid is ester-linked to glucuronoarabinoxylans in the same way as in the primary walls of grasses and cereals (Poaceae). Glucuronoarabinoxylans are the major non-cellulosic polysaccharides in the pineapple cell walls. PMID- 11261586 TI - Lipophilization of somatostatin analog RC-160 with long chain fatty acid improves its anti-proliferative activity on human oral carcinoma cells in vitro. AB - Oral cancer which comprises about 40% of total cancers in India, has one of the lowest relative survival rates of all cancers. Epidermal growth factor (EGF) has been known to play a role in the proliferation/malignant transformation of oral neoplasms. Since, the somatostatin analog RC-160 is reported to be a potent inhibitor of EGF stimulated cell proliferation, its anti-proliferative activity in the human oral carcinoma cell line KB was investigated, in this study. RC-160 was found to potently inhibit EGF-induced proliferation in KB cells in vitro, suggesting a therapeutic potential of the same in oral carcinoma. However, the therapeutic potential of RC-160 is limited by its short serum half life. To overcome this limitation, fatty acids namely butanoic acid and myristic acid individually were coupled to RC-160. The lipophilized derivatives of RC-160 were synthesized, purified and characterized. The anti-proliferative activity of lipophilized derivatives of RC-160 on KB cells was evaluated in vitro. Myristoyl RC-160 (0.75 nM) inhibited the growth of KB cells at a 10-fold lower concentration relative to RC-160 (8.8 nM) and at a 100-fold lower concentration relative to butanoyl-RC-160 (0.83 microM) (p<0.001). The affinity of RC-160 towards somatostatin receptors remains unaltered by lipophilization. The signaling pathways underlying the antineoplastic activity of these lipopeptides are similar to RC-160, and do not involve the stimulation of a protein tyrosine phosphatase or a serine threonine phosphatase 1A and 2A. The anti-proliferative activity of the lipopeptides was found to be mediated by somatostatin receptors and correlates with the inhibition of protein tyrosine kinase activity and decrease in intracellular cAMP levels. Myristoyl-RC-160 displayed significantly greater resistance towards trypsin and serum degradation than RC-160 (p<0.01). These findings demonstrate that RC-160 can inhibit the growth of oral cancer cells in vitro. Lipophilization of RC-160 with long chain fatty acids like myristic acid improves its stability and anti-proliferative activity, in human oral carcinoma cells in vitro, thereby enhancing the scope of improving its therapeutic index. PMID- 11261587 TI - Effects of Welsh onion extracts on human platelet function in vitro. AB - Welsh onion has been consumed for prevention of cardiovascular disorders. However, its underlying mechanisms are still unclear. This study investigated whether Welsh onion extracts can alter human platelet function (ie, platelet adhesion, aggregation, and thromboxane release). To clarify the underlying mechanisms, we also measured the intracellular calcium ([Ca2+]i) and cyclic nucleotide levels in platelets. Our results showed that 1) boiled extracts directly induced platelet aggregation in a dose-dependent manner; 2) raw extracts inhibited platelet adhesion and ADP-evoked platelet aggregation, while boiled extracts enhanced them; 3) raw green extract suppressed ADP-stimulated platelet [Ca2+]i elevation and thromboxane production, whereas boiled green extract enhanced them; 4) raw green extract elevated platelet cAMP level, whereas boiled green extract had no effect on cAMP level. Furthermore, the boiled green extract, but not the raw extract, induced pronounced platelet morphological changes. In conclusion, raw extracts of Welsh onion inhibit platelet function in vitro while boiled extracts activate platelets. PMID- 11261588 TI - Pharmacological characterization, molecular subtyping, and autoradiographic localization of putative melatonin receptors in uterine endometrium of estrous rats. AB - The objective of this study was to determine the biochemical characteristics, subtypes, and localization of melatonin receptors in the rat uterus in estrous stage. Autoradiography with the melatonin ligand, 2-[125I]iodomelatonin, showed that melatonin receptors were localized in the rat uterine endometrium. Binding of 2-[125I]iodomelatonin in crude membrane preparations of rat uterine endometrium in estrous stage was stable, saturable, reversible and of high affinity. Rosenthal analysis yielded an equilibrium dissociation constant (Kd) of 28.9 +/- 3.59 pmol/l (n = 8) and a maximum number of binding sites (Bmax) of 1.6 +/- 0.15 fmol/mg protein (n = 8). The Kd value determined from kinetic analysis was 16.5 +/- 3.02 pmol/l (n = 3). Competition studies using various indoles and neurotransmitters demonstrated that 2-iodomelatonin, melatonin, 6 chloromelatonin, 6-hydroxymelatonin and N-acetylserotonin showed significant inhibition of the 2-[125I]iodomelatonin binding, while the other indole compounds tested had no significant inhibition. The expression of rat uterine endometrial melatonin receptor subtypes was studied by reverse transcription-polymerase chain reaction (RT-PCR) using mt1 and MT2 receptor gene-specific primers. mt1 receptor cDNA was amplified and confirmed by nucleotide sequencing. These findings indicate that mt1 receptors were present in the rat uterine endometrium, and suggest that melatonin plays an integral part in uterine physiology. PMID- 11261589 TI - Trophoblastic cells expressing human chorionic gonadotropin genes in peripheral blood of patients with trophoblastic disease. AB - We attempted to identify the cells expressing alpha and beta subunits of human chorionic gonadotropin (hCG) in the peripheral blood of patients with trophoblastic disease and normal pregnant women by using reverse transcriptase polymerase chain reaction (RT-PCR) and Southern blot. By this method, the mRNAs of hCG alpha and hCG beta were detected in the peripheral blood mononulear cells (PBMNC) from 3 of 7 hydatidiform mole (mole) and 1 of 4 choriocarcinoma patients as well as from normal pregnant women during the first trimester. None of the mRNAs of hCG subunits was detected in the PBMNC from healthy male and nonpregnant healthy women examined. The expression of hCG alpha and hCG beta in patients with trophoblastic disease and normal pregnant women almost correlated with their plasma levels of intact hCG. The present study indicates that the cells expressing hCG alpha and hCG beta, which virtually represent trophoblasts, are circulating in the peripheral blood of patients with trophoblastic disease as well as of normal pregnant women. PMID- 11261590 TI - Activin receptors are expressed on human lung fibroblast and activin A facilitates fibroblast-mediated collagen gel contraction. AB - Activin A is a member of the transforming growth factor-beta superfamily that exerts its diverse biological effects through bindings to activin specific transmembrane serine/threonine kinase receptors. The fibroblast-mediated contraction of a collagen gel is thought to be a model of part of the wound repair response and tissue contraction. In this study, we found the expression of activin type I receptors (ActR-I and ActR-IB) and type II receptor (ActR-II) on human fetal lung fibroblasts (HFL-1) by RT-PCR and immunocytochemistry. We also examined the effects of activin A on the HFL-1-mediated collagen gel contraction. Activin A stimulated collagen gel contraction in a dose dependent manner and its effect was abolished by an activin-binding protein, follistatin, that specifically suppresses activin A activities. This study demonstrated that ActR I, ActR-1B and ActR-II are expressed on human fetal lung fibroblast and that activin A regulates fibroblast-mediated collagen gel contraction, suggesting that activin A might contribute to human lung fibroblast activities and structural remodeling observed in pulmonary fibrosis. PMID- 11261591 TI - Postnatal changes of vascular endothelial growth factor (VEGF) expression in the retinae of normal and hypertensive rats. AB - Vascular endothelial growth factor (VEGF) has been shown to have potent mitotic activity specific to vascular endothelial cells and has been related to vascular permeability, angiogenesis and cell proliferation in both normal and pathological situations. The present study aimed at elucidating the spatio-temporal changes in the postnatal expression pattern of VEGF in the retinae of both normal and hypertensive rats. In situ hybridization with a riboprobe showed that in the pre hypertensive stage (2 weeks postnatal, prior to the increase of the blood pressure of the hypertensive rat), VEGF expressed strongly in the retinal pigment epithelium (RPE) and inner nuclear layer (INL) but weakly in the ganglion cell layer and nerve fiber layer in both the normal and hypertensive rats. During the early hypertensive stage (6 weeks postnatal, initial increase of the blood pressure of the hypertensive rat), similar expression pattern was maintained but the INL of the hypertensive rat was found to have more positive cells in clusters than that of the normal rat. When a sustained high blood pressure was developed (12 weeks postnatal, sustained hypertensive stage) in the hypertensive rat, the VEGF expression was much reduced in all layers of the retina although weak expression was still observed in the RPE of the normal rat and RPE and INL of the hypertensive rat. Western blot analysis however showed that VEGF protein expression in the retina was much stronger in the hypertensive rat than in the normal rat at 2 and 6 weeks postnatal. At 12 weeks, the VEGF protein returned to a level comparable to that found in the normal rat. It is speculated that the change of the VEGF protein expression pattern during the early phase of the development of hypertension may be related to the subsequent changes in the retinal vasculature of the hypertensive rat. PMID- 11261592 TI - The antinociceptive effect of tramadol on a model of neuropathic pain in rats. AB - The antinociceptive activity of tramadol was investigated on the vocalization threshold to paw pressure in a rat model of unilateral mononeuropathy produced by loose ligatures around the common sciatic nerve. Despite the analgesic activity of tramadol was clearly established in motor and sensory responses of the nociceptive system in rats, the effect of this atypical opioid on experimental neuropathic pain models is not investigated. The intraperitoneally injected tramadol (2.5, 5 and 10 mg/kg) produced a potent and dose-dependent antinociceptive effect on both lesioned and non-lesioned hind paws. However, the analgesic effect on the lesioned paw was significantly more potent than the non lesioned paw. This effect was partially antagonized by intraperitoneally administered naloxone (0.1 mg/kg) suggesting an additional non-opioid mechanism. Our results suggest that tramadol may be useful for the alleviation of some symptoms in peripheral neuropathic conditions PMID- 11261593 TI - Effects of salmeterol on secretion of phosphatidylcholine by alveolar type II cells. AB - Beta-adrenergic agents enhance secretion of phosphatidylcholine (PC) by adult and fetal type II cells. We have previously shown that terbutaline stimulates secretion of PC by fetal type II cells, but the response wanes after 30 minutes. We studied the effects of salmeterol, a highly selective, long-acting beta2 adrenergic agonist that does not cause receptor desensitization, on PC secretion by adult type II alveolar cells in primary culture. Release of lactate dehydrogenase was < 4% and did not vary with the concentration of salmeterol. Salmeterol stimulated PC secretion in a concentration-dependent manner. The maximum effective-concentration tested was 50 nM and the EC50 was 11.40 +/- 1.14 nM. Propranolol inhibited the effect of salmeterol on release of PC, confirming that the effects of salmeterol are mediated by beta-receptors. OT50, the time for onset of action, was 32.0 +/- 1.6 minutes. RT50, the time to achieve 50% recovery from maximal stimulation was, 393.0 +/- 20.2 minutes. We conclude that salmeterol stimulates PC secretion by type II cells through activation of beta-adrenergic receptors and has a longer duration of action (>6 hours) compared to other beta2 agonists. Salmeterol may be a useful drug with which to study the role of receptor desensitization in the developmental changes in PC secretion. PMID- 11261595 TI - Effects of brain natriuretic peptide on pituitary-adrenal activation in rats. AB - The aim of this study was to characterize the effects of brain natriuretic peptide (BNP) on the hypothalamo-pituitary-adrenal (HPA) responses to different stress paradigms (ether stress, electric shock and restraint). Rats were subjected to the stressful stimuli after intracerebroventricular administration of BNP (32.5 ng-6.5 microg) and plasma corticosterone was used as an indicator of the HPA activation. BNP did not modify the basal secretion, but inhibited the stress-induced rise in plasma corticosterone in a dose-dependent manner. BNP proved most effective in decreasing the corticosterone response to ether stress and attenuated the electric shock and restraint-induced HPA activation to a lesser extent. These results confirm the view that BNP takes part in the regulation of the HPA system. PMID- 11261594 TI - Fatty acid composition of phospholipids, triglycerides and cholesterol in serum of castrated and estradiol treated rats. AB - We have studied the levels of phospholipids, triglycerides, cholesterol esters, and their fatty acid composition in serum for normal, castrated and estradiol treated rats. The sex hormones did not greatly affect the levels of the various lipid fractions which did not undergo great significant variations, under the different treatments. More evident variations occurred in the percent composition of fatty acid and in the content of the various saturated (SAT), unsaturated (UNSAT), essential (EFA) and non essential fatty acids (NEFA). We studied the most important ratios: EFA/NEFA; UNS/SAT; 16:0/16:1; 18:0/18:1, 18:2/18:3; 18:2/20:4. 16:0/16:1; 18:0/18:1 represent the delta9 desaturase, one specific for palmitic, the other for stearic acid. 18:2/18:3 ratio is an index of the delta6 desaturase activity: 18:2/20:4 ratio of delta5 desaturase-elongase. Most changes were evident in triglycerides. We observed a different behaviour of the UNS/SAT and EFA/NEFA ratios in phospholipids and cholesterol esters, which may reflect either an effect of the sex hormones on the exchange of fatty acids between the same lipid fractions, or a redistribution of lipids among different tissues. Great variations were observed of the ratios 16:0/16:1; 18:0/18:1; 18:2/18:3; 18:2/20:4, which are ascribed a different effect of the sex hormones of delta9, delta6, delta5 desaturases. PMID- 11261596 TI - Disopyramide block of K(ATP) channels is mediated by the pore-forming subunit. AB - The class Ia antiarrhythmic agent disopyramide blocks native ATP-sensitive K+ (K(ATP)) channels at micromolar concentrations. The K(ATP) channel is a complex of a pore-forming inwardly rectifying K+ channel (Kir6.2) and a sulfonylurea receptor (SUR). The aim of the present study was to further localize the site of action of disopyramide. We have used a C-terminal truncated form of Kir6.2 (Kir6.2delta26), which--in contrast to Kir6.2--expresses independently of SUR. Kir6.2delta26 channels were expressed in African green monkey kidney COS-7 cells, and enhanced green fluorescent protein (EGFP) cDNA was used as a reporter gene. EGFP fluorescence was visualized by a laser scanning confocal microscope. Disopyramide applied to the cytoplasmic membrane surface of inside-out patches inhibited Kir6.2delta26 channels half-maximally at 7.1 microM (at pH 7.15). Lowering the intracellular pH to 6.5 potentiated the inhibition of Kir6.2delta26 channels by disopyramide. These observations suggest that disopyramide directly blocks the pore-forming Kir6.2 subunit, in particular at reduced intracellular pH values that occur under cardiac ischaemia. PMID- 11261597 TI - Effect of GLG-V-13, a class III antiarrhythmic agent, on potassium currents in rabbit ventricular myocytes. AB - The effects of a new Class III antiarrhythmic drug, GLG-V-13, on the 4 aminopyridine sensitive transient outward current, on the inward rectifier potassium current, on the ATP sensitive potassium current and on the rapid and slow components of the delayed rectifier potassium current were studied in single rabbit ventricular myocytes using the whole-cell voltage-clamp technique. GLG-V 13 blocked the rapid component of the delayed rectifier potassium current in a dose-dependent manner, with an estimated EC50 value of 0.36 microM. At high concentration, the slow component of the delayed rectifier potassium current was also depressed by the drug (40% effect at 10 microM concentration). The transient outward current, the inward rectifier potassium current and the ATP sensitive potassium current were not influenced by GLG-V-13, even at 10 microM concentration. Thus, GLG-V-13 blocks predominantly the rapid component of the delayed rectifier potassium current which may play a significant role in the prolongation of repolarization by the drug in ventricular tissue. PMID- 11261598 TI - 17Beta-estradiol inhibits ovariectomy-induced expression of inducible nitric oxide synthase in rat aorta in vivo. AB - The objective of this study was to elucidate a role of ovarian steroid hormones in the production of immunologic nitric oxide (NO) synthases in the female rat aorta in vivo. Aortic homogenates were analyzed by using western blot with isoform-specific antibodies against endothelial NOS (eNOS) and inducible NOS (iNOS). Two weeks after ovariectomy (OVX), rats (10-week-old) were treated with 17beta-estradiol (E2) and/or progesterone (P4) for 5 days, and aortae were obtained from these rats on the following day. OVX markedly increased the levels of iNOS protein in abdominal aorta, whereas treatment with E2 or a combination of E2 and P4 inhibited the induction of iNOS in the aorta. The present findings indicate that endogeneous estrogen negatively regulates the expression of iNOS in abdominal aorta, and suggest that changes in the levels of circulating estrogen may affect vascular function. PMID- 11261599 TI - The influence of photodynamic therapy on the wound healing process in rats. AB - In photodynamic therapy (PDT), photosensitisers (PS) are used along with lasers for the treatment of tumors. The combined effect of photosensitisers and lasers on the wound healing process is studied using delta-aminolevulinic acid (ALA) (5 mg/kg) and hematoporphyrin derivative (HPD) (5 mg/kg) as photosensitisers in the open excision wounds of rats. The lasers used were He-Ne laser (3 J/cm2) and Nd:YAG laser (30 J/cm2). This study is important for understanding the healing process involved after PDT. Open excision wounds treated with He-Ne lasers in animals that received ALA as photosensitiser showed complete wound closure at the earliest by 13 +/- 1 days, and with results obtained for HPD and the combination of lasers with complete closing by 14 +/- 1 days. However, the control group of animals that received ALS or HPD with no laser treatment showed wound healing on the twentieth and eighteenth days with a deviation of one day and two days, respectively. ALA with the combination of Nd:YAG and He-Ne lasers and HPD with He Ne laser alone does not show quicker wound healing effects. Histopathological results also gave similar results. Tensile strength measurements do not vary significantly from control group to the test group. ALA along with He-Ne laser of HPD along with the combination of He-Ne and low power Nd-YAG lasers are found to be ideal methods for quickening the wound healing process in rat. PMID- 11261600 TI - Ageing human bone: factors affecting its biomechanical properties and the role of collagen. AB - The incidence of fractures increases with age. This is partly due to extraosseous factors and partly to the increased fragility of the bone material itself. Ageing adversely affects the "quality" of human bone material, its elastic and ultimate properties. The hypothesis here is that these effects are caused by factors such as architectural changes, compositional changes, physicochemical changes, changes at the micromechanical level, and the degree of prior in vivo microdamage. Examination of the extent of the secondary osteonal area, the porosity level, the calcium content, the mineral/wet weight fraction, the dry density, the condition of the collagen and its content in mature x-links, the elasticity of osteonal and interstitial lamellae at the microscopic level and the numerical- and surface density of the in vivo fatigue microcracks has been undertaken. The findings show that some factors simply affect the stiffness and the strength of bone, while others soley affect its toughness. We discuss the implications of these findings in the context of the composite nature of the ageing bone material matrix. PMID- 11261601 TI - Ultrastructure of exogenous surfactants using cryogenic scanning electron microscopy. AB - Therapy with specialised biomaterials, exogenous surfactants, is known to significantly decrease the mortality rates in Respiratory Distress Syndrome (RDS). Surfactants available commercially vary widely in composition and biophysical properties. The present paper studies the ultrastructure of three exogenous surfactants used for the treatment of Respiratory Distress Syndrome, namely, Survanta, ALEC and Exosurf Neonatal with respect to their ability to form liposomes using cryogenic scanning electron microscopy. Liposomal organisation is more obvious in Exosurf than in Survanta and is most pronounced in ALEC. ALEC forms closed regular liposomes with an onion-ring-like internal bilayer arrangement. Survanta forms open membranous structures with wavy ribbon-like membranes. The complex membrane-like structures seen with Survanta may be due to the interaction of lipids with surfactant-specific proteins present in this surfactant which is derived from natural lung extracts and might indicate superior spreading at the lipid-water interface. Artificial protein-free surfactants (ALEC and Exosurf) did not appear to form these open membranous structures. Further study of the ultrastructure of possible biomaterials as surfactants could help in the development of new, improved artificial protein free surfactants with open membranous structures that might facilitate spreading at the air-liquid interface of lungs. PMID- 11261602 TI - The mechanical behavior of vascular grafts: a review. AB - The development of intimal hyperplasia (IH) near the anastomosis of a vascular graft to artery is directly related to changes in the wall shear rate distribution. Mismatch in compliance and diameter at the end-to-end anastomosis of a compliant artery and rigid graft cause shear rate disturbances that may induce intimal hyperplasia and ultimately graft failure. The principal strategy being developed to prevent IH is based on the design and fabrication of compliant synthetic or innovative tissue-engineered grafts with viscoelastic properties that mirror those of the human artery. The goal of this review is to discuss how mechanical properties including compliance mismatch, diameter mismatch, Young's modulus and impedance phase angle affect graft failure due to intimal hyperplasia. PMID- 11261603 TI - Quantitative radiographic analysis of fiber reinforced polymer composites. AB - X-ray radiographic examination of the bone fracture healing process is a widely used method in the treatment and management of patients. Medical devices made of metallic alloys reportedly produce considerable artifacts that make the interpretation of radiographs difficult. Fiber reinforced polymer composite materials have been proposed to replace metallic alloys in certain medical devices because of their radiolucency, light weight, and tailorable mechanical properties. The primary objective of this paper is to provide a comparable radiographic analysis of different fiber reinforced polymer composites that are considered suitable for biomedical applications. Composite materials investigated consist of glass, aramid (Kevlar-29), and carbon reinforcement fibers, and epoxy and polyether-ether-ketone (PEEK) matrices. The total mass attenuation coefficient of each material was measured using clinical X-rays (50 kev). The carbon fiber reinforced composites were found to be more radiolucent than the glass and kevlar fiber reinforced composites. PMID- 11261604 TI - Regulation of human papillomavirus late gene expression. AB - Human papillomaviruses (HPVs) comprise a large group of small DNA tumor viruses with tropism for squamous epithelial cells. The papillomavirus life cycle is strictly linked to the epithelial differentiation program and production of virus particles is dependent on terminal cell differentiation. The virus structural proteins L1 and L2 are therefore detected only in the upper layers with differentiated cells in the infected epithelium. This property is common to all known HPVs. However, the abundance of virus particles detected in vivo may vary between the various HPV types and one can divide the HPV types according to the amount of virus they produce. It is reasonable to speculate that the differentiation dependent production of L1 and L2 and the differences in L1 and L2 protein levels among various HPV types reflect an adaptation of the viruses to the environment of the host that results in escape from the immune surveillance for various periods of time and efficient transmission of virus in the human population. Our research group is interested in the regulation of expression of the HPV structural proteins L1 and L2. The results of our research are summarised in this article. PMID- 11261606 TI - Neuropeptide levels in murine liver and biliary pathways. AB - Neuropeptide concentrations in the liver, gallbladder and bile duct of 30 male mice were determined by radioimmunoassay of tissue extracts. Vasoactive intestinal peptide (VIP), substance P, somatostatin, neurotensin, neuropeptide Y (NPY), galanin, gastrin-releasing peptide (GRP) and enkephalin were found in extracts of liver, gallbladder and bile duct. The level of enkephalin was low in all tissues investigated. The most predominant neuropeptide in liver and bile duct was VIP and in the gallbladder, NPY. The function of these neuropeptides in the liver and biliary pathways is unclear. One can speculate, however, that in the liver they regulate blood flow and probably secretion. In the gallbladder and bile duct, they may regulate motility and secretion. PMID- 11261605 TI - Human islets transplanted to nude mice: in vitro insulin release from retrieved grafts. AB - Limitations in success of clinical islet transplantation may be coupled to a long term decline in the secretory capacity of the grafted human islet tissue. To address this issue human or mouse islets were transplanted to the subcapsular space of the kidney of nude mice. After 4 or 12 weeks, the grafts were removed and tested for insulin secretory dynamics in a perifusion system. Insulin secretion of non-transplanted human islets was examined as well. Insulin extracted from 12-week human islet grafts was significantly lower than that from 4-week grafts. Quite in contrast, 12-week mouse islet grafts contained as much insulin as the 4-week grafts. When stimulated with high glucose, insulin secretion was increased by about 6-fold in non-transplanted human islets and 3 fold in 4-week-grafts. The 12-week-grafts were just marginally stimulated by the high glucose stimulation. The mouse islets maintained a 2- to 3-fold insulin response at both time points when challenged high glucose. In non-transplanted human islets glucose-induced insulin secretion was inhibited by noradrenaline, while there was no such effect in the human islet grafts. Addition of acetylcholine potentiated glucose-induced insulin secretion 1-4 fold in both non transplanted and grafted human islets. When human islet grafts were stimulated by both glucose and caffeine or arginine, insulin secretion was increased severalfold in comparison to glucose stimulation alone. The present results indicate that human islets, in contrast to mouse islets, progressively diminish their insulin content, as well as the capacity to secrete insulin in response to glucose after transplantation into nude mice. Moreover, grafted human islets also lose their responsiveness to the neurotransmittor noradrenaline. These findings may partly explain why clinical islet transplantation so far has met with limited success. PMID- 11261607 TI - Cigarette smoke and hypoxia induce acute changes in the testicular and cerebral microcirculation. AB - The acute effects of cigarette smoking and hypoxia on the cerebral and testicular microcirculation were studied in anestethised adult rats. Smoking for 2 min did not influence arterial pO2, pCO2 or pH but it induced an increase in cerebral blood flow by 34% and inhibited vasomotion in the testis for about 1 h. One hour after smoke exposure apnea induced a slight increase in arterial pCO2, a significant decrease in pO2, and an increase in cerebral blood flow (CBF) by 54%. In animals not previously exposed to cigarette smoke apnea increased CBF by 121%, demonstrating that a short-term exposure to tobacco smoke influences the cerebrovascular reactivity for more than one hour. In the testis, apnea resulted in a decreased blood flow by 39% and a complete depression of vasomotion. Breathing 10% O2/90% N2 resulted in moderate hypoxia, a total disappearance of the vasomotion in the testis, a 24% decrease in testicular blood flow, but a 23% increase in CBF. PMID- 11261608 TI - Clinicopathologic prognostic factors in patients with Borrmann type 4 gastric cancer: univariate and multivariate analyses. AB - BACKGROUND: Advanced gastric cancer is classified into four Borrmann types, types 1 to 4. Type 4 is a relatively undifferentiated carcinoma with little or no gland forming capability. Despite recent advances in the diagnosis and surgical management of gastric cancer, most tumors of Borrmann type 4 are not detected at an early stage and the prognosis remains poor; the five-year survival rate after gastric resection ranges from 10 to 20 percent. We evaluated the affects of several clinicopathologic variables on the 5-year survival rate after resection of Borrmann type 4 gastric cancer. METHODS: Data on clinical characteristics were obtained from the records of patients who underwent gastric resection between 1985 and 1995 at the Department of Surgery, Sendai National Hospital, and follow up data were obtained from our tumor registry. Pathologic characteristics were determined from a detailed review of all available histopathologic slides. The relationship between clinicopathologic variables and 5-year survival rate was estimated by the Kaplan-Meier survival curve and the logrank test. Multivariate Cox's proportional hazards regression analysis was then performed to determine which variables were independent prognostic factors. RESULTS: Eighty-seven patients with Borrmann type 4 gastric cancer underwent a resection during the study period at our hospital. The overall 5-year survival rate was 14.8%. The relationship between clinicopathologic variables and 5-year survival rate was determined by constructing a Kaplan-Meier survival curve. Tumor location (upper, middle and distal vs whole stomach, p=0.0214), lymph node metastasis, capillary microinvasion, and peritonitis carcinomatosa (absent vs present, p<0.05) significantly influenced survival. When multivariate analysis using Cox's proportional hazards regression of 5-year survival was performed, capillary microinvasion, peritonitis carcinomatosa (absent vs present) and tumor location (distal vs whole stomach) emerged as the statistically significant independent prognostic factors associated with long-term survival. CONCLUSION: Capillary microinvasion and the presence or absence of peritonitis carcinomatosa are more powerful predictors of 5-year survival than is lymph node metastasis. Patients with gastric cancer of the whole stomach have a poorer prognosis than do those with carcinoma in the antrum of the stomach. PMID- 11261609 TI - Spread of epidural blockade in the anaesthetised pig using technetium99 and sensory-evoked potentials. AB - Our objective was to determine the extension and duration of a stable segmental thoracic or lumbar epidural blockade by lidocaine in the 20 to 25-kg pig. Fourteen anaesthetised pigs were investigated. The radioactive isotope technetium99 (Tc99) was added to lidocaine to evaluate the spread in the epidural space. One ml of this solution was administered via an epidural catheter either at the Th(6-7) or L6-S1 level. Sensory-evoked potentials (SEPs) were measured following peripheral nerve stimulation, to determine duration and sensory extension of the epidural blockade. The thoracic anatomical mean spread was 14.2 vertebrae. The end points of the spread were C3-C7 and Th8-Th13 and the lumbar anatomical mean spread was 10.2 vertebrae. The end points were Th10-Th12 and L6 S1. It was possible to abolish the SEPs with 2 and 4% lidocaine. A smaller volume of lidocaine was needed if a second dose was administered within 20 min. With 4% lidocaine it was possible to abolish the SEPs for more than 180 min. The use of 1 ml of 4% lidocaine repeated after 20 and 60 min at the Th(6-7) or L6-S1 level achieved a stable thoracic or lumbar epidural blockade. PMID- 11261611 TI - Prevalence of sexually transmitted diseases (STD) among women in a suburban Sudanese community. AB - 338 women with age ranging from 15 to 69 years in a suburban Sudanese community were randomly selected and studied. Urine sample, high vaginal swabs and blood samples were investigated for bacterial vaginosis, candidiasis, trichomoniasis, gonorrhoea, HIV and syphilis. The sensitivity and specificity of some laboratory tests were evaluated. Bacterial vaginosis was found in 17.2% of the subjects, candidiasis in 10.1%, trichomoniasis in 7.7%, gonorrhoea in 1.2%, HIV in 1.2% and syphilis in 0.9% of the subjects. The sensitivity and specificity of amine test as a criterion for diagnosing bacterial vaginosis was 58.6% and 73.2%, respectively. The respective values of clue cells in wet preparation were 43.1% and 99.6%. The vaginal discharge in women with bacterial vaginosis lacked pus cells unless associated with concurrent infection. PMID- 11261610 TI - Anastomotic rupture at the site of polytetrafluoroethylene (PTFE) and distal vein cuff of femoropopliteal bypass. Two case reports. AB - Two female patients, 63 and 78 years of age, underwent femoropopliteal bypass with polytetrafluoroethylene (PTFE) graft and distal vein cuff. They developed graft occlusion due to false aneurysm at the site of vein cuff during one and eight weeks after surgery, respectively. Improper suture technique or weak vein wall might lead to suture disruption leading to false aneurysm as presented in this article. PMID- 11261612 TI - Nutrition borne myolysis; an uncommon cause of myolysis in Europe. PMID- 11261613 TI - CT imaging of trunk muscles in chronic low back pain patients and healthy control subjects. AB - Increasing documentation on the size and appearance of muscles in the lumbar spine of low back pain (LBP) patients is available in the literature. However, a comparative study between unoperated chronic low back pain (CLBP) patients and matched (age, gender, physical activity, height and weight) healthy controls with regard to muscle cross-sectional area (CSA) and the amount of fat deposits at different levels has never been undertaken. Moreover, since a recent focus in the physiotherapy management of patients with LBP has been the specific training of the stabilizing muscles, there is a need for quantifying and qualifying the multifidus. A comparative study between unoperated CLBP patients and matched control subjects was conducted. Twenty-three healthy volunteers and 32 patients were studied. The muscle and fat CSAs were derived from standard computed tomography (CT) images at three different levels, using computerized image analysis techniques. The muscles studied were: the total paraspinal muscle mass, the isolated multifidus and the psoas. The results showed that only the CSA of the multifidus and only at the lowest level (lower end-plate of L4) was found to be statistically smaller in LBP patients. As regards amount of fat, in none of the three studied muscles was a significant difference found between the two groups. An aetiological relationship between atrophy of the multifidus and the occurrence of LBP can not be ruled out as a possible explanation. Alternatively, atrophy may be the consequence of LBP: after the onset of pain and possible long loop inhibition of the multifidus a combination of reflex inhibition and substitution patterns of the trunk muscles may work together and could cause a selective atrophy of the multifidus. Since this muscle is considered important for lumbar segmental stability, the phenomenon of atrophy may be a reason for the high recurrence rate of LBP. PMID- 11261614 TI - Influence of age and duration of symptoms on fibre type distribution and size of the back muscles in chronic low back pain patients. AB - Many studies have documented an association between chronic low back pain (LBP) and deficits in back muscle strength and endurance. The sub-optimal performance is believed to be the result of alterations in the size and structure of the muscle, although the long-standing issue of whether the observed changes precede or are a consequence of the pain remains unresolved. If consequent to the problem, and predominantly related to disuse of the muscles, then it may be expected that a relationship between muscle structure and symptom duration would exist. Lumbar paraspinal muscle samples were obtained from 59 chronic LBP patients using the percutaneous biopsy technique. The samples were subject to routine histochemical analysis for the examination of muscle fibre type characteristics and cytochemical architectural changes. In 55 of the patients, the gross cross-sectional areas of magnetic resonance images of the trunk muscles were also measured. Multivariate analysis showed that symptom duration was the strongest predictor of the individual proportions of the fast-fatigable type IIX fibres; with age and gender included in the model, nearly 30% of the variance in fibre type distribution could be accounted for. Duration of pain had no influence on fibre size. Gross muscle cross-sectional area correlated directly with lean body mass and inversely with age, but showed no relationship with symptom duration. Pathological changes in the internal fibre structure were more frequently encountered in older patients, and were independent of symptom duration. The results suggest that, over the long term, fibre type transformations rather than alterations in fibre size are the predominant changes to be found in the muscles of chronic LBP patients. The direction of change supports the results of many previous studies that have demonstrated corresponding differences in the fatigability of the muscles. There is a strong case for the early implementation of active measures to attempt to offset the development of these changes in back pain patients. PMID- 11261616 TI - Nuclear factor kappaB activation in acute appendicitis: a molecular marker for extent of disease? AB - Nuclear factor-kappaB (NF-kappaB) has been demonstrated to regulate the transcription of target genes and stimulate inflammatory cytokine responses in a variety of inflammatory diseases. Preliminary studies have demonstrated that NF kappaB is activated early in acute inflammation and sepsis and may serve as an indicator of clinical severity. The present study was designed to evaluate the degree of activation of NF-kappaB in patients with acute appendicitis and correlate activation with clinical extent of disease. Ten patients with acute appendicitis and five control patients (elective inguinal hernia repair) were evaluated by assaying NF-kappaB activity preoperatively and 12 to 18 hours postoperatively. Assaying of NF-kappaB was determined by binding activity for consensus probes in nuclear extracts from peripheral mixed white blood cells obtained by venous puncture. The bands of NF-kappaB activity from gel electrophoresis were quantified with a phosphor imager and reported as units of integrated intensity. The preoperative NF-kappaB activity was increased in all patients with appendicitis versus the controls [mean 151 (range 97-189) vs mean 50.3 (range 13.7-77); P < 0.0001]. The increased NF-kappaB activity also correlated with length of time of symptoms before operation. The patients who were symptomatic for less than 24 hours had an average NF-kappaB value of 103 (range 97-105) versus 171.4 (range 152-189) (P < 0.0001) in those who were symptomatic 24 or more hours. The NF-kappaB activity did not correlate with the white blood cell count. Postoperative NF-kappaB binding activity in the appendicitis patients dropped to minimal levels (mean 50.3), even lower than the control patients' baseline values (mean 55.6). Control patients demonstrated low baseline values preoperatively and a slight rise postoperatively [mean 50.3 (range 13.7-77) vs mean 100 (range 45-186)]. We conclude the following: (1) NF kappaB binding activity is elevated in patients with acute appendicitis and correlates with symptoms longer than 24 hours. (2) This increased activity returns to baseline values within 18 hours after appendectomy. (3) Molecular indicators of inflammation may have a role in both staging surgical inflammatory conditions and predicting ultimate outcome. PMID- 11261615 TI - Acute lung injury after hemorrhagic shock is dependent on gut injury and sex. AB - Recent studies have established gut-derived lymph rather than portal blood as the major source of toxic mediators after hemorrhagic shock that causes distant organ injury. Similarly, emerging data have identified sex as a major modifier of the response to injury and illness. Thus we tested the hypothesis that female rats would be more resistant to shock-induced lung injury than male rats because females are more resistant to shock-induced gut injury and produce mesenteric lymph that is less toxic to endothelial cells. Male and female rats were subjected to sham or hemorrhagic shock and lung permeability was quantitated by Evans blue dye and protein extravasation into the alveolar space. Next, mesenteric lymph collected from shocked and sham-shocked rats of both sexes was incubated with human umbilical vein endothelial cells (HUVECs) and assayed for toxicity. Trypan blue dye exclusion and the release of lactate dehydrogenase assessed HUVEC viability and injury respectively. Lastly, sections of the terminal ileum were histologically examined for evidence of shock-induced mucosal injury. Male rats but not female rats subjected to hemorrhagic shock had evidence of increased lung permeability and produced mesenteric lymph that was cytotoxic to HUVECs. Shock caused gut injury in the male rats whereas histological evidence of gut injury was not observed in the female rats. Hemorrhagic shock-induced lung injury depends on gut injury and mesenteric lymph appears to be the route by which gut-derived toxic factors exit the gut to cause lung injury. The resistance of female rats to shock-induced lung injury appears to be secondary to their resistance to shock-induced gut injury. PMID- 11261618 TI - Six-year experience with management of subclavian artery injuries. AB - Penetrating injuries of the subclavian artery are rare; however, the associated morbidity and mortality may be high. Retrospective data on 25 patients who sustained penetrating subclavian artery injuries are reported. Diagnosis of subclavian artery injuries was made clinically and was followed by expedient surgical exploration in 65.4 per cent of patients. Patients who were hemodynamically unstable at presentation (26.9%) underwent immediate operation. The remaining hemodynamically stable group of patients with hard signs indicative of vascular injury were also expediently taken to the operating room after initial evaluation and resuscitation. Angiographic evaluation was performed in 34.6 per cent of patients who were stable hemodynamically. Preoperative angiography localized the injury and helped in planning the optimal incision and approach to obtain vascular control. Vascular flow was reestablished in all patients operated except for three who underwent ligation of subclavian artery. Limb salvage rate was 100 per cent, and operative mortality was less than 5 per cent. Morbidity was related to hemodynamic stability at presentation and associated injuries. A low morbidity and mortality rate was achieved by aggressive initial resuscitation and early surgical intervention coupled with selective use of preoperative angiography in hemodynamically stable patients. PMID- 11261617 TI - A successful multimodality strategy for management of liver injuries. AB - The treatment of liver injuries involves many strategies ranging from observation to operative intervention and includes numerous options such as angiography, packing, and damage-control procedures. In July 1994 we instituted a protocol for the management of traumatic liver injuries. The main objective of this study was to evaluate the management of liver injuries occurring since the institution of the protocol. Two hundred three consecutive adult patients with liver injuries were evaluated at our Level I trauma center between July 1994 and May 1999. Eighty-eight per cent of the injuries were blunt with a mean Injury Severity Score (ISS) of 24.3 +/- 0.8 and a survival probability (Ps) of 90.0 +/- 1.5 per cent. The overall mortality was 6.4 per cent. A comparison between patients with minor liver injuries and patients with more severe injuries [Abbreviated Injury Score (AIS) <3 vs >3] demonstrated no difference in mortality between the two groups despite a Ps of 93.8 +/- 1.3 per cent in patients with an AIS <3 versus 84.1 +/- 3.3 per cent in patients with an AIS >3. The most common complication in our patient population was posthemorrhagic anemia, which was seen in 10.8 per cent of cases. Severity of injury did not result in a significant difference in the complication rate. Patients who underwent laparotomy had a statistically higher ISS, a lower Ps, and a mortality rate of 11.5 per cent compared with 3.7 per cent (P = 0.03) in patients managed nonoperatively. However, a comparison of patients undergoing laparotomy with those who did not and who had equivalent ISS demonstrated no difference in mortality. Our results demonstrated that a preplanned management strategy was a successful way in which to treat patients with traumatic liver injuries. Although nonoperative management of liver injuries has been common practice a management plan that involves a multimodal surgical strategy is essential. PMID- 11261619 TI - Laparoscopic adrenalectomy is superior to an open approach to treat primary hyperaldosteronism. AB - We reviewed our institutional experience with primary hyperaldosteronism to compare clinical outcomes after laparoscopic versus open adrenalectomy. All patients surgically treated for primary hyperaldosteronism from 1988 through 1999 are included in this study. Patients were assigned to either the LA (laparoscopic) or OA (open) group depending on the initial surgical approach selected for treatment. Records were reviewed to determine demographics, operative results, and complications. Twenty-four patients were surgically treated for primary hyperaldosteronism. There were no significant differences between groups with respect to age, weight, number of preoperative antihypertensive medications, or preoperative potassium level. The results of adrenalectomy with respect to number of postoperative antihypertensive medications or serum potassium level were also similar. Operative times were not significantly different (191 +/- 53 minutes for OA and 205 +/- 88 minutes for LA) between groups, but four LA patients were converted to OA. Estimated blood loss was 401 +/- 513 cm3 for OA and 127 +/- 131 cm3 for LA (P = 0.07). Hospital length of stay was 6.7 +/- 3.7 days for OA and 3.3 +/- 2.7 days for LA (P = 0.02). Complications were nine for OA and three for LA (P = 0.001 by Pearson's Chi square). LA is similar to OA in the treatment of primary hyperaldosteronism. The significantly fewer complications and shorter length of hospital stay associated with LA makes the laparoscopic approach the preferred method for treating primary hyperaldosteronism. PMID- 11261620 TI - Ultrasound-guided thrombin injection of iatrogenic femoral pseudoaneurysms: a prospective analysis. AB - An adverse consequence of the use of the femoral artery for the endovascular evaluation and treatment of arterial disease is the increased incidence of iatrogenic femoral pseudoaneurysms. Although surgical repair has traditionally been used to treat such aneurysms, less invasive modalities have emerged. The purpose of this study is to prospectively evaluate ultrasound-guided thrombin injection (UGTI) for the treatment of iatrogenic femoral pseudoaneurysms. A treatment protocol was approved and 30 stable patients (21 female; age range 43 85 years; mean 67 years) were prospectively enrolled from December 1997 through June 1999 to undergo UGTI on 30 iatrogenic femoral pseudoaneurysms. Pseudoaneurysms occurred after cardiac intervention (n = 22, 73%), peripheral intervention (n = 7, 23%), and after a femoral line placement (n = 1, 3%). They ranged in size from one to 5 cm with a time interval from intervention until UGTI of one to 132 days (median 3 days). Eleven patients (37%) were systematically anticoagulated at the time of UGTI. All pseudoaneurysms were treated using sterile technique and local anesthesia with ultrasound-guided injection via a 20 gauge spinal needle of 0.1 to 2 cm3 (median 0.6 cm3) of 1000 units/cm3 topical thrombin solution administered by one of six physicians. A period of bedrest for 4 to 6 hours after injection was followed by repeat groin duplex scan at 24 hours and a clinical follow-up at 30 days. There were no procedural deaths or nonvascular complications. Twenty-seven (90%) UGTIs resulted in successful pseudoaneurysm ablation with no recurrences at 24 hours or 30 days. Two (7%) UGTIs failed and one (3%) femoral artery embolic complication occurred; all were successfully treated with surgery. Success appeared to be independent of anticoagulation status, pseudoaneurysm age, size, or operator experience. We conclude that UGTI is a safe, easy, well-tolerated and effective noninvasive method for treatment of iatrogenic femoral pseudoaneurysms and should be considered in all stable patients before operative repair. PMID- 11261621 TI - Sentinel lymph node mapping for carcinoma of the colon: a pilot study. AB - Sentinel lymph node (SLN) mapping has evolved into the standard of care for melanoma and may replace routine node dissection in the treatment of breast cancer. There are few data evaluating sentinel node mapping in patients with cancer of the colon. This trial represents our initial experience with SLN mapping for carcinoma of the colon. SLN mapping was performed in 22 patients most of whom had biopsy-proven adenocarcinoma of the colon. One milliliter of isosulfan blue was injected with a 25-gauge needle into the subserosa at four sites around the edge of the palpable tumor. The SLN was identified visually and excised. A standard lymphadenectomy was then performed. The SLN was analyzed with standard hematoxylin and eosin evaluation. Immunohistochemical techniques for carcinoembryonic antigen and cytokeratin (Imm) were performed if the H&E was negative. The mapping added approximately 5 minutes to the total operative time and no adverse reactions to the dye occurred. A SLN was identified in 20 of 22 cases. In cases with negative lymph nodes the SLN was predictive of all the regional nodes by both H&E and Imm (14 of 14). In patients with positive lymph nodes the SLN was predictive in all cases (six of six). In one case the only node with disease was the SLN, and in this case the diease was identified by only Imm; thus this patient was upstaged. SLN mapping is feasible and safe and can readily be performed in patients with colonic cancer. In conjunction with SLN mapping, Imm techniques may upstage a subset of patients likely to be at increased risk for metastatic disease. Consequently SLN mapping of colon cancer should be evaluated in large prospective trials. PMID- 11261622 TI - Isoproterenol inhibits bacterial lipopolysaccharide-stimulated release of tumor necrosis factor-alpha from human heart tissue. AB - Recent evidence suggests that inflammatory cytokines, particularly tumor necrosis factor alpha (TNF-alpha), may play a role in heart disease. Elevated plasma levels of the cytokine have been reported in congestive heart failure and severe angina and after myocardial infarction. The exact role of TNF-alpha in heart disease and how production is stimulated and regulated in the heart are current areas of investigation. Regarding regulation of production, isoproterenol elevates cyclic AMP and inhibits TNF-alpha release in macrophages. Therefore we hypothesized that stimulation of beta-adrenergic receptors of the sympathetic nervous system would inhibit release of the cytokine from heart tissue. With Institutional Review Board approval and patient consent atrial tissue was obtained during preparation for cardiac bypass. The tissue was divided into segments, placed in culture medium, and incubated for various times in the presence or absence of lipopolysaccharide (LPS) (20 microg/mL) and/or isoproterenol (1 microM). The medium was removed and analyzed for biologically active TNF-alpha by the L929 cell cytotoxicity assay. Tissue samples were weighed and TNF-alpha release was expressed as pg TNF-alpha/mg tissue. Initially, to determine the time course of release, measurements were made at 2, 5, 10, 15, 30, 60, 120, 180, and 360 minutes after the addition of LPS. Elevated TNF-alpha levels in the culture medium were reliably detected at 360 minutes after exposure to LPS. In atrial tissue obtained from seven patients TNF-alpha released into the culture medium at 360 minutes was 6 +/- 3 pg/mg tissue. In the presence of LPS, levels of the cytokine in the culture medium increased to 604 +/- 233 pg/mg tissue (P < 0.05 vs LPS alone). When isoproterenol and LPS were simultaneously added to the culture medium release of TNF-alpha was reduced by 87 per cent to 82 +/- 40 pg/mg tissue (P < 0.05 vs LPS alone). Our results show that activation of the beta-adrenergic receptor inhibits myocardial production of TNF-alpha. This finding suggests that the sympathetic nervous system inhibits production of the cytokine and that impaired sympathetic function in heart failure may play a role in the elevated levels of TNF-alpha. PMID- 11261623 TI - Splenectomy for idiopathic thrombocytopenic purpura: a five-year retrospective review. AB - Idiopathic thrombocytopenic purpura is a condition that is characterized by persistently low platelet counts. Idiopathic thrombocytopenic purpura results from splenic sequestration and accelerated platelet destruction mediated by antiplatelet antibody. Most cases arise in previously healthy patients, mostly women ages 20 to 40. Clinical symptoms consist of bruising, petechiae, mucosal bleeding, menorrhagia, and intracranial bleeding. Platelet-associated immunoglobulin G can be detected in 90 per cent of patients. Therapy for adults and children is somewhat different. Splenectomy in adults should be considered in patients who fail to respond to steroids, develop thrombocytopenia after taper, or develop steroid toxicity. Ninety per cent of children will maintain normal platelet counts in 9 to 12 months. Some will recover spontaneously without medical therapy. Splenectomy in children is recommended if idiopathic thrombocytopenic purpura persists for more than one year or fails to respond to steroids. Our purpose was to determine whether management of idiopathic thrombocytopenic purpura in patients who undergo splenectomy at our institutions is appropriate and effective. We undertook a 5-year retrospective review of 27 patients with idiopathic thrombocytopenic purpura which have undergone splenectomy. All of the 27 patients were referred to surgeons after initial medical management. The patients were divided into two groups on the basis of length of therapy: longer than 6 months and less than 6 months. The longer than 6 months group contained 15 patients. This group had a postoperative complication rate of 40 per cent. Those in the group with <6 months therapy had a complication rate of 7 per cent. Average follow-up for all patients was 20 months. Eighty eight per cent of the patients had complete response. Three per cent had a partial response with platelet counts >50,000. The partial response group did not respond well to preoperative steroid boluses with a great rise in platelet counts. Eighteen per cent of patients received platelet transfusions. Sixty per cent of the transfusions were given for inappropriate reasons. A large percentage of our patients had prolonged medical therapy before splenectomy. The inappropriate use of platelets was a common error in management. Patients treated for more than 6 months had more postoperative complications. An initial increase in platelets after steroid bolus is a good indicator for favorable response to splenectomy. We conclude that splenectomy is a safe and effective method of treatment for idiopathic thrombocytopenic purpura with no deaths or postsplenectomy sepsis to date. PMID- 11261624 TI - Self-expanding esophageal metallic stents in the treatment of esophageal obstruction. AB - Esophageal obstruction from any cause is debilitating. In patients with malignant obstruction palliation to relieve pain and dysphagia is the primary goal. Conventional endoluminal prostheses allow variable palliation. Covered expandable metallic stents with an 18-mm lumen allow improved deglutition. From December 1994 through December 1998, 59 patients underwent placement of self-expanding silicone-covered esophageal stents for esophageal obstruction. There were 36 men and 23 women ranging in age from 41 to 94. All patients underwent esophageal dilation using a flexible gastroscope and Savary bougies. After dilation placement of the stent was performed under fluoroscopic control. Follow-up was complete in all patients. Technical success was achieved in all patients. There was one postoperative death (bronchopulmonary fistula), one migration of the stent requiring removal, and one recurrent obstruction. The remaining stents were well tolerated even in the cervical region (four patients). All patients returned to a diet of solid foods. We conclude that covered self-expanding esophageal metallic stents are technically simple and safe to insert and appear to provide durable excellent palliation of esophageal obstruction due to either benign or malignant conditions. PMID- 11261625 TI - A simple approach to nipple discharge. AB - Evaluation and management of patients with nipple discharge (ND) aims to identify carcinoma when present, and in benign cases, stop the discharge when bothersome. We reviewed our recent experience with ND to develop a simple and effective algorithm to manage these patients. Records of all patients with ND evaluated from December 1996 through June 1999 were reviewed. Patients were liberally offered duct excision for a clinical suspicion of malignancy (persistent clear or bloody fluid) or to stop bothersome discharge. Patients with breast imaging abnormalities (mammography or ultrasound) related to their ND underwent biopsy and were considered separately. Of 104 patients with ND, 11 underwent biopsy as a result of mammographic findings; three of these cases proved malignant. The remaining 93 patients were evaluated with 55 tests that did not demonstrate malignancy, including ductography, discharge fluid cytology, serum prolactin and thyroid-stimulating hormone levels, and image-guided breast or nipple biopsy. Thirty-nine patients underwent duct excision with only a single patient demonstrating malignancy. Clinical follow-up has not identified malignancy in any patient managed nonoperatively. When diagnostic breast imaging is negative, malignancy related to ND is uncommon. Patients with ND should have diagnostic breast imaging and, if it is negative, should be offered duct excision. There is little role for ductography, cytology, or laboratory studies in evaluating these patients. PMID- 11261626 TI - Necrotizing soft tissue infections: a surgical disease. AB - Despite advances in antibiotics and infection control practices necrotizing fasciitis is still a potentially lethal disease. We reviewed 37 patients with necrotizing fasciitis to identify prognostic factors indicating outcome. Overall mortality was 24 per cent. Mortality was significantly increased for elderly patients. Solid-organ transplant recipients also represented a subset of patients with increased mortality. Most infections were polymicrobial. There was no Clostridium perfringens cultured. Rapid diagnosis and treatment with surgical debridement remains the cornerstone of therapy. PMID- 11261627 TI - An unusual case of corneal perforation secondary to Pseudomonas keratitis complicating a patient's surgical/trauma intensive care unit stay. AB - We report a case of corneal perforation secondary to bacterial keratitis caused by Pseudomonas aeruginosa in a trauma patient in our intensive care unit. A 43 year-old man was involved in a motorcycle crash and suffered multiple injuries necessitating a prolonged intensive care unit (ICU) stay. Subsequently P. aeruginosa was cultured from his sputum, blood, and open abdomen. He developed a bacterial keratitis in his right eye, which also grew P. aeruginosa. This infection rapidly progressed to corneal perforation requiring a Gunderson conjunctival flap and lateral tarsorrhaphy in addition to aggressive antibiotic treatment. At the time of discharge from the hospital the patient had the return of vision to light only in his right eye. Corneal perforation is an unusual event in the ICU. Prevention or early detection of bacterial keratitis with aggressive antibiotic treatment is needed to prevent such complications. Pseudomonas is one of the more virulent organisms that can infect the cornea and early identification is paramount for a good outcome. Management of this complicated case is discussed and the limited amount of literature on nosocomial bacterial keratitis in the ICU is reviewed. PMID- 11261628 TI - Ultrasound-guided percutaneous thrombin injection for femoral artery pseudoaneurysms. AB - We reviewed 13 cases of ultrasound-guided thrombin injection of femoral pseudoaneurysms. All cases occurred within a 17-month period from January 1998 through May 1999 and were complications of femoral artery puncture. Immediate total thrombosis occurred in nine of 13 patients. Twenty-four-hour follow-up ultrasound in seven patients revealed no recurrence of pseudoaneurysm. Two of 13 patients required operative repair. One pseudoaneurysm thrombosed with 15 minutes of compression after injection and one case required a second injection. No cases of arterial thrombosis were noted. Ultrasound-guided thrombin injection for femoral artery pseudoaneurysm represents a safe and effective alternative to operative repair. PMID- 11261629 TI - Age-adjusted outcomes in traumatic flail chest injuries in the elderly. AB - Severe chest trauma does not independently predict poor outcome in elderly patients. We chose a specific injury, flail chest, to determine whether age factored into outcome of these patients. A retrospective chart review of all trauma admissions to our Level I trauma center between January 1994 and January 1998 sustaining flail chest was undertaken. Sixty-eight patients were identified, but ten patients were excluded because of death on arrival. Fifty-eight patients were included in the study and separated into groups. The first group comprised those under the age of 55 (n = 32) and the second comprised those over age 55 (n = 26). Parameters evaluated were age, Injury Severity Score (ISS), neurologic injury, the need for mechanical ventilation, need for tracheostomy, length of stay, and death. Statistical analysis was performed with Wilcoxon t test, chi2, and logistic regression where appropriate. A 95 per cent confidence interval was sought as determinant of significance. Of the 58 surviving patients analyzed there was no significant difference between the groups regarding ISS, length of stay, days on the ventilator, head injury, tracheostomy, or development of pneumonia or adult respiratory distress syndrome. The likelihood of death was shown to increase by 132 per cent for every 10 years starting at the second decade and continuing to the eighth decade of life. The likelihood of death also increased by 30 per cent for each unit increase in ISS. The likelihood of death decreased by 23 per cent for every day survived in the hospital. Blunt chest trauma directly impacts respiratory mechanics. Elderly patients are more likely to have comorbid conditions and less likely to tolerate traumatic respiratory compromise. Age (and its effects on the body) is the strongest predictor of outcome with flail chest and is associated with an increased mortality (P < or = 0.05). PMID- 11261630 TI - Effect of fibrin glue on lymphatic drainage after modified radical mastectomy: a prospective randomized trial. AB - Fibrin as a tissue sealant has been used since the turn of the century for hemostasis. The development of cryoprecipitate and the resultant availability of higher concentrations of fibrinogen have led to a resurgence of interest in this material. Fibrin glue has since been shown to be effective for numerous applications throughout the field of surgery. Animal studies have shown fibrin glue to be effective at reducing drain output after mastectomy. Human studies, however, have been equivocal. Our objectives were to determine whether the use of fibrin glue would decrease lymphatic drainage after modified radical mastectomy and subsequently reduce time to drain removal. A prospective randomized trial was conducted consisting of 27 women. All women received modified radical mastectomy. At the completion of the mastectomy they were randomized to receive either standard closure or the application of fibrin glue before standard closure. Patients were then monitored for daily drain output, time to drain removal, and wound complications. A total of 14 women received fibrin glue and 13 received no glue. Those patients receiving fibrin glue had a significantly higher average drain output than patients who did not receive glue (1308 vs 754 cm3; P = 0.012). Time to drain removal was also increased by 4 days, although this did not reach statistical significance. The overall complication rate was higher for the fibrin glue group, although again, this did not reach significance. The application of fibrin glue significantly increased drain total drain output after modified radical mastectomy. Time to drain removal was increased as was the complication rate. On the basis of these data fibrin glue cannot be recommended for routine use in modified radical mastectomy. PMID- 11261631 TI - Duplex examination of the inferior vena cava. AB - Duplex examination of the inferior vena cava (IVC) was performed in 270 patients from 1/1/96 to 1/1/00 to define suitability of the IVC for caval interruption using noninvasive means. The IVC was interrogated using a 3-mHz curved linear array probe and an ATL Ultramark 9 ultrasound machine (Bothell, WA). Duplex measured IVC dimensions and defined presence of thrombus or anomalies. Of the 270 IVC duplex examinations 10.7 per cent (n = 29) could not be completed because of overlying bowel gas or for other technical reasons. Of the 241 completed studies 4.1 per cent (n = 10) revealed acute or chronic thrombosis of the IVC. The lateral diameter of the IVC was 20.3 +/- 4.4 mm (95% confidence interval 19.8 20.9 mm), whereas the anteroposterior diameter was 12.6 +/- 4.0 mm (95% confidence interval 12.1-13.1 mm). Excluding those with vena cava thrombosis maximum vena cava diameters exceeded 28 mm in only 2.2 per cent (n = 5) of those with technically adequate studies. Apart from the latter megacavas there were no major IVC anomalies detected. For those with incomplete studies body weight was 192 +/- 59 lb versus 169 +/- 38 lb for those with technically adequate studies (P = 0.008). Technically adequate vena cava duplex examinations can be performed in 89 per cent of patients. On the basis of this and one prior study done at this center IVC duplex can define vena cava dimensions and presence of thrombus. Using the standard criteria for IVC filter insertion that require presence of a maximum cava diameter < or = 28 mm and absence of caval thrombus or anomalies, 94 per cent (226 of 241) of those with complete duplex examinations would have been anatomically suitable for standard Greenfield filter insertion based on noninvasive testing. PMID- 11261632 TI - Eosinophilic gastroenteritis mimicking acute appendicitis. AB - Eosinophilic gastroenteritis is a rare entity that can be treated successfully with glucocorticoid therapy if the appropriate diagnosis is made. However, it may present with symptomatology mimicking acute surgical conditions. We present the case of a 26-year-old man who presented with diffuse epigastric pain, nausea, vomiting, and diarrhea. Extensive workup including upper endoscopy and imaging study revealed gastritis with ulcer and ascites. The patient developed right lower quadrant pain with localized peritonitis and leukocytosis. He underwent appendectomy and small bowel biopsy. Pathology revealed eosinophilic cellular infiltrate of both the appendiceal and small intestinal wall. The unique features of this condition are reviewed and surgical approaches are discussed. PMID- 11261633 TI - Models of community treatments in schizophrenia: do they travel? AB - OBJECTIVE: To explore the stability of conclusions from mental health services research across differing care systems. Contradictory results in different countries for similar studies of programmes for patients with schizophrenia have usually been attributed to poor replication. This paper explores whether these differing results can illuminate aspects of schizophrenia by examining the interaction of the disorder with the care context as an alternative explanation. METHOD: The findings of a large UK random controlled trial of intensive case management with such patients is compared to previous UK and US studies. RESULTS: Reduction of case-load size of psychotic patients did not significantly reduce their need for hospitalization in the context of locally available co-ordinated care. CONCLUSION: There is more to be gained in understanding complex disorders such as schizophrenia by interpreting the impact of context on treatment study outcomes than by simply dismissing contradictory findings as failures of implementation of either research or clinical practice. PMID- 11261634 TI - Schizophrenia costs and treatment cost-effectiveness. AB - OBJECTIVE: The paper sets out to summarize evidence on the costs of schizophrenia and on the cost-effectiveness of three broad treatment areas. METHOD: Evidence from a number of countries was examined, both published and unpublished, and systematic reviews and meta-analyses were consulted. RESULTS: The costs of schizophrenia are high and wide-ranging. They fall not only to health-care agencies but also to other parts of the public sector, to families, to sufferers themselves and to the wider society. However, there are interventions--a counselling intervention to address non-compliance with medication, family interventions to reduce levels of expressed emotion, and atypical antipsychotic drugs--that have been found to be not only effective (improving patient outcomes) but also appear to be cost-effective. CONCLUSION: Resource constraints and policy pressures make it increasingly common for economic as well as clinical questions to be asked about new modes of treatment. This is the new reality of mental health practice. Reliable evidence is now available to address these economic questions and can be factored into decision-making processes. PMID- 11261635 TI - The interactive developmental model revisited. AB - OBJECTIVE: To provide a more adequate method for integrating our understanding of biological, psychological and social factors in schizophrenia, new concepts and a broader view of scientific method are required. This report aims to describe steps towards these concepts and methods. METHOD: Using experiences from a range of research, clinical, and general life encounters, the author reviews assumption about objectivity and science in the mental health field where human experience is a crucial source of data and concepts. RESULTS: These experiences indicate that the more complex concepts of intentionality, emotional experience and expression, and the concept of experience have been largely ignored in much schizophrenia research and theory. CONCLUSION: Some approaches to overcoming this lack are suggested. PMID- 11261636 TI - Public acceptance of restrictions on mentally ill people. AB - OBJECTIVE: To assess the influence of demographic, psychological, sociological and cultural variables on the public acceptance of restrictions on mentally ill people. METHOD: Multiple logistic regression analysis of the results of an opinion survey conducted on a representative sample in the German, French and Italian-speaking part of Switzerland. RESULTS: Public acceptance of restrictions depends on age, education and gender. The influence of demographic variables is reduced when including rigidity, anomie, social distance and contact with mentally ill people in the analysis. The cultural influence of the language region is significant. All these factors predict acceptance. Stereotypes and emotional reactions have no influence. CONCLUSION: Attitudes to mentally ill people are shaped on three distinct levels, the psychological, social and cultural, respectively. Demographic variables partly play the role of placeholders covering the underlying relationships. PMID- 11261637 TI - Schizophrenia and quality of life. AB - OBJECTIVE: The purpose of this paper is to examine critically results of quality of-life research in schizophrenic patients living in the community. METHOD: Based on the relevant literature results of specific studies are discussed in the light of the methodological problems of assessing quality of life in these people. RESULTS: Subjectively assessed quality of life was found to be higher in the less educated and in female patients, and when a sense of control is experienced. If negative or extrapyramidal symptoms are experienced and stigmatization is perceived, subjective quality of life is reported as being poorer. These results are discussed in view of the additional needs and fewer resources of this population, their stigma-dilemma and their occasional difficulties in adequately assessing quality of life. CONCLUSION: It is argued that subjective and quantitative measures of quality of life in schizophrenic patients should be supplemented by external assessment and qualitative methods. PMID- 11261639 TI - Onset and early course as determinants of the further course of schizophrenia. AB - OBJECTIVE: First treatment contact is preceded by a lengthy prodromal and psychotic prephase. We analysed the occurrence of symptoms and disabilities prior to first contact and their consequences for medium-term illness course. METHOD: A population-based sample of 232 first episodes of schizophrenia was studied retrospectively using the IRAOS and compared with matched peers from the population of origin. Further illness course was studied prospectively at six cross-sections over 5 years from first admission onwards. RESULTS: Three quarters showed a prodromal phase of 5 years (mean) and a 1-year accumulation of psychotic symptoms. First to appear were depressive and negative symptoms and early signs of cognitive and social impairment. Social disability emerged 4 to 2 years before first contact. Further illness course was determined by stage of social development at psychosis onset with the consequence of a significantly poorer course for men than women because of men's earlier illness onset. Symptomatology, type of onset, age and gender influenced social course via stage of social development and, additionally, via young men's socially adverse illness behaviour. CONCLUSION: Social course is determined by individual stage of social development at illness onset and by early illness course. Therefore, early detection and intervention are needed. PMID- 11261638 TI - Ethnic disadvantage and schizophrenia. AB - OBJECTIVE: To review and interpret epidemiologic research on ethnic disadvantage and schizophrenia. METHOD: A search of the research literature was conducted. RESULTS: Seventeen population-based studies were reported in the United Kingdom and the Netherlands from 1967 to 1997. The studies report high incidence rates for immigrants whose position in society is disadvantaged, than majority-group native-born, with a range of relative incidence from 1.7 to 13.2. It is proposed that the developmental task for formulating the life plan challenges the young adult's executive function abilities, which may be weaker in individuals vulnerable to schizophrenia. Formulating the life plan may be made more difficult by the position in society of disadvantaged ethnic minorities, raising the risk for schizophrenia. CONCLUSION: Further research on executive function, and the developmental challenge of formulating the life plan, might provide insights into the etiology of schizophrenia, as well as suggest avenues for prevention. PMID- 11261640 TI - Long-term course in schizophrenia: concepts, methods and research strategies. AB - OBJECTIVE: This paper summarizes current knowledge about course and outcome in schizophrenia by selecting particular information and by drawing conclusions in the light of theoretical and methodological considerations, in order to illustrate future research strategies, as well as to consider novel targets of treatment. METHOD: Based on examples from the literature the concepts of course and outcome as well as intervening factors of course and outcome are discussed by considering study results within their methodological and theoretical framework restrictions. RESULTS: The heterogeneity of the disorder, manifest in its variable phenomenology, illness course, treatment response and outcome, complicates research but at the same time offers clues to elucidate the heterogeneity of aetiology and pathogenesis. The search for intervening factors and predictors is of particular interest in order to explore the illness mechanisms. CONCLUSION: Future research should focus on the biopsychosocial determinants of course and outcome in methodologically improved long-term studies. PMID- 11261641 TI - Long-term course of schizophrenic, affective and schizoaffective psychosis: focus on negative symptoms and their impact on global indicators of outcome. AB - OBJECTIVE: The aim of this study was to compare indicators of outcome between different types of psychosis and to verify whether or not negative symptoms (NS) have a special relevance for schizophrenia. METHOD: This is a follow-up study on functional psychosis according to ICD-9. Patients were assessed standardized at the time of first hospitalization and about 12.5 years later. RESULTS: Comparison of global outcome parameters and NS revealed that schizophrenia had the poorest outcome of all types of psychosis. NS had the highest impact on global functioning and the severity of illness in schizophrenia. NS assessed at the first hospitalization were associated with the different outcome parameters only in schizophrenia at follow-up. CONCLUSION: The course of schizophrenia is a more deteriorating one than that of affective or schizoaffective psychosis. The findings point to the special relevance of NS for the outcome and their relative specificity for schizophrenia. PMID- 11261642 TI - Gender aspects in schizophrenia: bridging the border between social and biological psychiatry. AB - OBJECTIVE: This paper tries to show that gender differences in mental diseases are a valuable paradigm for research into the interplay between biological and psychosocial factors--not only regarding pathogenetic mechanisms, but also concerning therapeutic approaches. METHOD: Based on relevant literature, this topic is highlighted using schizophrenia as an example. RESULTS: Schizophrenic disorders show a later age of onset in women and a slightly better course, especially in young women. As to pathogenesis, there is some evidence that the age difference might be due at least partly to the female sex hormone oestradiol being a protective factor. Differences in course might also have to do with this biological factor, but at the same time with the psychosocial advantages of a higher age of onset and other psychosocial factors. Concerning therapy, these gender differences have important implications for pharmacotherapy, but also psychotherapy and social measures. CONCLUSION: A gender-sensitive approach in psychiatry improves our understanding of mental illness and our therapeutic strategies and at the same time illustrates that comprehensive psychiatry cannot be practised in artificially separated 'drawers' called 'biological psychiatry', on one hand, and 'social psychiatry' on the other. PMID- 11261643 TI - Psychosocial rehabilitation and psychiatry in the treatment of schizophrenia- what are the boundaries? AB - OBJECTIVE: To examine the relationship between psychosocial rehabilitation and psychiatry with particular emphasis upon treatment of people who suffer from schizophrenia. METHOD: Current literature is examined in an effort to identify prevailing premises and myths held by each discipline as it relates to the other. RESULTS: A working definition of psychosocial rehabilitation is proposed, and nine interrelated concepts that constitute the belief system of that discipline are identified. CONCLUSION: Rehabilitation's biopsychosocial perspective provides a strong basis for joint enterprise with psychiatry in the care of people with schizophrenia. However, members of both disciplines must pursue active measures in order to effect co-operative treatment interventions. PMID- 11261644 TI - Schizophrenia and violence. AB - OBJECTIVE: The relationship between schizophrenia and violence is studied from a psychiatric and a public health perspective. METHOD: All epidemiological studies which have been published since 1990 are reviewed. RESULTS: Despite differences in the methodological approaches chosen the studies reviewed concur in supporting the assumption that there is a moderate but significant association between schizophrenia (or more generally psychotic disorders) and violence. However, compared with the magnitude of risk associated with substance abuse and personality disorders, that associated with schizophrenia or other major mental disorders is small. In addition, the elevated risk to behave violently appears to be limited to particular symptom constellations. The evidence available so far suggests that the proportion of violent crimes committed by people suffering from a severe mental disorder is small. There is no unambiguous evidence of an increase of violent acts committed by severely mentally ill people in general and people suffering from schizophrenia in particular during recent years. Strangers appear to be at an even lower risk of being violently attacked by someone suffering from severe mental disorder than by someone who is mentally healthy. CONCLUSION: While the assessment of relative risk is of great interest for psychiatric researchers who are trying to identify factors which may increase or decrease the risk of violent behaviour among the mentally ill, which in turn may provide some clues as to how to intervene best in order to reduce the risk, the attributable risk is of special interest for the public since it informs about the risk of becoming victim of a violent act committed by someone who is suffering from a mental disorder. PMID- 11261645 TI - Migration and schizophrenia. AB - OBJECTIVE: Since the beginning of the human race, individuals have migrated alone or in groups. This process of migration has often been considered to be an aetiological factor in the genesis of many mental disorders. METHOD: Two studies collecting rates of first onset schizophrenia in Trinidad and in London using the same assessment instruments. RESULTS: The sending countries have low rates of schizophrenia. The impact of migration itself produces high stress but rates of schizophrenia are even higher in the second generation, suggesting that that other social factors may be responsible for the increase if genetic vulnerability is excluded. CONCLUSION: Individual social factors, such as cultural identity and the impact of racism, are more likely to play a key role in the genesis of schizophrenia. PMID- 11261646 TI - Psychological therapy in schizophrenia: what is the evidence? AB - OBJECTIVE: The present contribution provides a critical outline of the current position of psychological therapies in schizophrenia. METHOD: Therefore, empirical research into the efficacy of psychological interventions in the treatment of schizophrenic disorders has been reviewed. RESULTS: Four cognitive behavioural approaches have emerged as preeminently effective, or at least especially promising, as adjuncts to pharmacotherapy, i.e. the training of social skills, cognitive training programs for the remediation of neurocognitive deficits, psychoeducative, coping-orientated interventions with patients and their families, and cognitive-behavioural therapy of residual symptoms. These approaches are discussed with regard to their efficacy in reducing relapse rates, psychopathology as well as cognitive and social disability. CONCLUSION: Open questions and possibilities for the further development of these approaches are considered and prognostications are made concerning the future of psychotherapy research in schizophrenia, notably in the light of changing conditions in public health care systems. PMID- 11261647 TI - Family work for schizophrenia: practical application. AB - OBJECTIVE: To review the evidence for the efficacy, efficiency and effectiveness of family work for schizophrenia. METHOD: The review is based on the relevant literature but is not intended to be exhaustive, except in the area of practical application. RESULTS: The effectiveness of family work has been established by a series of randomized controlled trials. Relatives groups are efficient in terms of staff time, and multiple family groups may be more efficacious than sessions with single families. However, a substantial proportion of relatives refuse to attend a group and need sessions in the home. Family work skills can be acquired by clinicians working in ordinary settings, although few studies have addressed this question. Problems have been encountered regularly by trained community workers trying to practise their newly acquired skills. CONCLUSION: Difficulties in implementation may be remedied by adopting a systemic approach and including the managers of the service in the initial training sessions. PMID- 11261648 TI - Compliance with antipsychotic treatment. AB - OBJECTIVE: Despite the demonstrated efficacy of antipsychotics the relapse rate among patients with schizophrenia remains high. One major reason for this is non compliance. In this article we review different factors influencing compliance and discuss possibilities to enhance compliance among schizophrenic patients. METHOD: This review is based on a systematic literature search in Medline. RESULTS: We summarize the four main factors (patient-, environment-, physician- and treatment-related) that influence compliance and discuss possible measures to enhance compliance. Next to many other variables discussed in more detail, it is crucial to ensure a positive doctor-patient relationship and to provide sufficient information about the benefit/risk ratio of the medication as well as about the illness itself to build up and sustain compliance. Significant others should be included into the therapeutic alliance whenever possible. CONCLUSION: Despite many published reports on compliance, it remains to be a problem of eminent clinical relevance. Clinicians must not underestimate it in order to optimize the treatment of patients with schizophrenia. PMID- 11261649 TI - Integrating pharmacological and psychosocial treatments for schizophrenia. AB - OBJECTIVE: The objective of this study was to develop principles of combining pharmacological and psychosocial treatment that can be useful for clinicians who are treating patients with schizophrenia. METHOD: Research studies in schizophrenia that controlled both pharmacological and psychosocial treatments were reviewed. These included studies using conventional and newer antipsychotics as well as a number of psychosocial methods. RESULTS: The interactions between the forms of treatment appear to be more than merely additive, since each can enhance the effects of the other. Drug and psychosocial treatments may affect different outcome domains, with the former affecting symptoms and the later affecting social outcomes. Recent studies using newer antipsychotics suggest that these agents improve the participation of patients in psychosocial treatments. CONCLUSION: Understanding the interactions between psychosocial and pharmacological treatments can be useful for clinicians who are developing treatment strategies for patients with schizophrenia. Newer agents with different side-effect profiles and broader effectiveness appear to have improved the outcomes of psychosocial treatments. PMID- 11261650 TI - Actigraphy to measure day structure as a therapeutic variable in the treatment of schizophrenic patients. AB - OBJECTIVE: A component of social skills is the ability to adapt to the social rhythms of the environment. Patients with schizophrenia are often disabled in this adaptation. Thus, structuring activities throughout the day has long been known as part of psychosocial treatments. Actigraphy as a tool to measure acitvity and circadian rhythms may even serve as an indicator for the day structuring of schizophrenic patients. METHOD: Actigraphy was used in a patient with affective disorder and one with chronic schizophrenia for more than 2 weeks. RESULTS: In comparison to a regular 24-hour rest-activity cycle in a depressed patient, the actigraph of the patient with schizophrenia presents active phases at night, irregular activity levels at day and signs of a delayed-sleep-phase syndrome. CONCLUSION: Actigraphy could serve as a tool to investigate activity levels and circadian rest-activity phases, even in schizophrenia. There may be some further benefit of actigraphy as a tool in psychosocial treatments. PMID- 11261651 TI - The schizophrenic patient in the somatic hospital. AB - OBJECTIVE: There is an increased risk in schizophrenia for premature death from illnesses in almost all organic systems. The present study analyses the Rate Ratio (RR) for schizophrenic patients' admissions to somatic departments in Denmark. METHOD: 20000 schizophrenic patients were identified in the Danish Psychiatric Central Register and 200000 sex- and age-matched controls were identified in the Danish Central Person Register. Both groups were searched for in the Danish National Patient Register in which admissions to all somatic departments are registered. Pulmonary and cardiovascular diseases are used as examples. RESULTS: RR is increased for several diseases, especially infectious, up to a maximum of RR = 4.15 for severe heart failures and decreased to as low as RR = 0.35 for atherosclerotic diseases of the brain vessels. CONCLUSION: It seems that individuals with schizophrenia are rarely treated for their physical illness in its early, less severe phases, but more likely in its acute phases when the disease is severe, life-threatening or painful. PMID- 11261652 TI - Combinatorial chemistry toward understanding the function(s) of carbohydrates and carbohydrate conjugates. AB - Combinatorial chemistry has contributed significantly to understanding the structure-function relationships of biologically important molecules such as proteins and nucleic acids. However, carbohydrates and carbohydrate conjugates, which have been identified as key modulators of several biological functions have not enjoyed the same measure of success. The complexity and synthetic challenges of carbohydrate conjugates have resulted in a number of conceptual approaches to rapidly access sufficient quantities of these biomolecules. This article summarizes these combinatorial approaches and also highlights fully automated library synthesis of artificial glycopeptides with the goals of understanding their biological roles. PMID- 11261653 TI - Regio- and stereoselective synthesis of Nor-nonactinic acid derivatives--kinetic reaction control in the Lewis acid mediated domino reaction of 1,3-dicarbonyl dianions with 1-bromo-2,3-epoxypropanes. AB - Reaction of 1,3-dicarbonyl dianions with epibromohydrin derivatives results in formation of functionalized 2-alkylidene-5-hydroxymethyltetrahydrofurans. These reactions proceed by chemoselective attack of the dianion onto the carbon attached to the bromine atom and subsequent nucleophilic attack of the resultant monoanion onto the epoxide. The cyclization products, which were formed with very good regio- and stereoselectivities, are of pharmacological relevance and represent versatile building blocks for the synthesis of natural products. PMID- 11261654 TI - Direct transformation of silyl enol ethers into functionalized allenes. AB - The first elimination reactions of silyl enol ethers to lithiated allenes are reported. These reactions allow a direct transformation of readily available silyl enol ethers into functionalized allenes. The action of three to four equivalents of lithium diisopropylamide (LDA) on silyl enol ethers results in the formation of lithiated allenes by initial allylic lithiation, subsequent elimination of a lithium silanolate, and finally, lithiation of the allene thus formed. Starting with amide-derived silyl imino ethers, lithiated ketenimines are obtained. A variety of reactions of the lithiated allenes with electrophiles (chlorosilanes, trimethylchlorostannane, dimethyl sulfate and ethanol) were carried out. Elimination of silanolate is observed only for substrates that contain the hindered SiMe2tBu or Si(iPr)3 moiety, but not for the SiMe3 group. The reaction of 1,1-dilithio-3,3-diphenylallene with ketones provides a convenient access to novel 1,1-di(hydroxymethyl)allenes which undergo a domino Nazarov-Friedel-Crafts reaction upon treatment with p-toluenesulfonic acid. PMID- 11261656 TI - Probing the reactivity of oxomanganese-salen complexes: an electrospray tandem mass spectrometric study of highly reactive intermediates. AB - Electrospray ionization in combination with tandem mass spectrometric techniques has been employed to study the formation of oxomanganese-salen complexes upon oxidation of [Mn(III)(salen)]+ cations as well as the properties and reactions of the oxidized species in the gas phase. Two species could be characterized as the principal oxidation products: the oxomanganese(v) complex, [Mn=O(salen)]+, which is the actual oxygen-transfer agent in epoxidation reactions, and the dinuclear, mu-oxo bridged [L(salen)Mn-O-Mn(salen)L]2+ with two terminal ligands L; the latter acts as a reservoir species. The effects of various substituents in the 5- and 5'-positions, respectively, of the salen ligand on the reactivity of the epoxidation catalyst were determined quantitatively from CID (collision-induced dissociation) experiments and B3LYP density functional calculations. Accordingly, the effect of axial donor ligands on the reactivity of the epoxidation catalyst was studied. Electron-withdrawing substitutents on the salen ligand and additional axial ligands decrease the stability and thus enhance the reactivity of the Mn=O moiety, while electron-donating salen substituents have a strong stabilizing effect. PMID- 11261655 TI - Facile solid-phase synthesis of an antifreeze glycoprotein. AB - The antifreeze glycoproteins (AFGPs) 1 are composed of a repeating tripeptide unit (Ala-Thr-Ala) in which the threonine residue is glycosylated with the disaccharide beta-D-Gal-(1-->3)-alpha-D-GalNAc. A new procedure for synthesizing AFGPs using Fmoc-(Ac4-beta-D-Gal-(1-->3)-benzylidene- alpha-D-GalNAc)Thr-OH (10) as a building block has been developed. Total synthesis of the AFGPs (n = 4, 8) in overall yields of 61% and 33 %, respectively, has demonstrated the usefulness of the method. The synthetic AFGPs 1 (n = 4, 8) showed a similar conformation to the native AFGPs in their circular dichroism spectra. PMID- 11261657 TI - Molecular dynamics simulations of MRI-relevant GdIII chelates: direct access to outer-sphere relaxivity. AB - The structure and dynamics of the surrounding water were studied through molecular dynamics (MD) simulations for several GdIII polyaminocarboxylate and polyaminophosphonate complexes in aqueous solution. The radial distribution functions (rdf) show that a few water molecules are bonded to the ligand through hydrogen bonds to hydrophilic groups such as carboxylates and phosphonates. Residence times are of the order of 20-25 ps for the polyaminocarboxylate and 56ps for the polyaminophosphonate chelates. No preferred orientation or bonding of water molecules is observed in the hydrophobic region of the anisotropic macrocyclic complexes. Our rdf allow calculation of the outer-sphere contribution to the nuclear magnetic resonance dispersion (NMRD) profiles using Freed's finite differences method, including electronic relaxation. The results show that the commonly used analytical force-free model is only an empirical relationship. When experimental outer-sphere NMRD profiles are available ([Gd(teta)]- and [Gd(dotp)]5-(teta=N,N',N",N"'-tetracarboxymethyl-1,4,8,11- tetraazacyclotetradecane; dotp = N,N',N",N"'-tetraphosphonatomethyl-1,4,7,10 tetraazacyclododecane) the calculated curves are in good agreement. In the case of [Gd(teta)]-, the comparison with the experimental NMRD profile has led us to predict a very fast electronic relaxation, which has been confirmed by the EPR spectrum. PMID- 11261658 TI - Planar chirality: cycloaddition and transannular reactions of optically active azoninones that contain (E)-olefins. AB - Unsaturated nine-membered ring lactams that contain (E)-olefins within the ring are characterized by planar chiral properties. Thus, selective conversions of the double bond allowed a complete transfer of the planar chiral information into new stereogenic centers The basis of the transformations was the high activation barrier that prevented efficient flipping of the double bond at room temperature (epimerization pR <=> pS) with respect to the ring. Cycloadditions led diastereoselectively to cyclopropano, epimino, epoxy, and dihydroxy azonanones under mild conditions with moderately high yields. The epoxy azonanones were subjected to regio- and diastereoselective transannular epoxide opening/ring contraction sequences to give hydroxy indolizidinones. The regiochemical and stereochemical outcome strongly depends on the configuration of the oxirane and the chiral information of the lactam unit. The so-formed optically active bicycles with defined substitution patterns should serve as versatile building blocks in alkaloid synthesis. PMID- 11261659 TI - Complete defluorination of 1,2,3,4-tetramethyl-5-(trifluoromethyl)cyclopentadiene by titanium tetrakis(dimethylamide)--selective formation of a cyclic hexanuclear titanium fluoroamide and 6,6-dimethylaminotetramethylfulvene. AB - 1,2,3,4-Tetramethyl-5-(trifluoromethyl)cyclopentadiene (Cp*CF3-H, 1) reacts with [Ti(NMe2)4] (2) under mild conditions to give [Ti(mu-NMe2)(NMe2)(mu-F)(F)]6 (3) in nearly quantitative yield. The molecular structure of 3 consists of a ring of six [TiF2(NMe2)2] edge-bridged octahedra. Titanium complexes containing the Cp*CF3 ligand, which was the primary intention of these investigations, were not observed. C5Me4=C(NMe2)2 (4) was isolated as a by-product. The complete defluorination of an aliphatic CF3 group occurs during the reaction. The reaction mechanism involves the primary formation of a difluorofulvene intermediate C5Me4=CF2 (5), which was monitored by NMR studies. Density functional theory calculations predict a highly charged C6 atom (+0.87) in 5, which is discussed as the driving force of the reaction. PMID- 11261660 TI - Exchange coupling in carboxylato-bridged dinuclear copper(II) compounds: a density functional study. AB - A computational study of the exchange coupling is presented for a selected sample of carboxylato-bridged dinuclear copper(II) compounds. Model calculations have been used to examine the influence of several factors on the coupling constants: a) the electron-withdrawing power of the bridging ligands; b) the nature of the axial ligands; c) the number of bridging carboxylato groups; d) some structural distortions frequently found in this family of compounds; and e) the coordination mode of the carboxylato bridge. Coupling constants calculated for some complete structures, as determined by X-ray diffraction, are in excellent agreement with experimental data, confirming the ability of the computational strategy used in this work to predict the coupling constant for compounds for which experimental data are not yet available. PMID- 11261661 TI - The first acetimine gold(I) and gold(III) complexes and the first acetonine complexes. AB - Ketimino(phosphino)gold(I) complexes of the type [Au[NR=C(Me)R']L]X (X = ClO4, R = H, L = PPh3, R'=Me (la), Et (2a); L=PAr3 (Ar=C6H4OMe-4), R'=Me (1b), Et (2b); L=PPh3, R=R'=Me (3); X= CF3SO3 (OTf), L=PPh3, R=R'=Me (3'); R=Ar, R'=Me (4)) have been prepared from [Au(acac)L] (acac = acetyl acetonate) and ammonium salts [RNH3]X dissolved in the appropriate ketone MeC(O)R'. Complexes [Au(NH=CMe2)2]X (X = C1O4 (6), OTf (6')) were obtained from solutions of [Au(NH3)2]X in acetone. The reaction of 6 with PPN[AuCl2] or with PhICl2 gave [AuCl(NH=CMe2)] (7) or [AuCI2(NH=CMe2)2]ClO4 (8), respectively. Complex 7 was oxidized with PhICl2 to give [AuCl3(NH=CMe2)] (9). The reaction of [AuCl(tht)] (tht = tetrahydrothiophene), NaClO4, and ammonia in acetone gave [Au(acetonine)2]ClO4 (10) (acetonine = 2,2,4,4,6-pentamethyl-2,3,4,5-tetrahydropyrimidine) which reacted with PPh3 or with PPN[AuCl2] to give [Au(PPh3)(acetonine)]ClO4 (11) or [AuCl(acetonine)] (12), respectively. Complex 11 reacts with [Au(PPh3)(Me2CO)]ClO4 to give [(AuPPh3)2(mu-acetonine)](ClO4)2 (13). The reaction of AgClO4 with acetonine gave [Ag(acetonine)(OClO3)] (14). The crystal structures of [Au(NH2Ar)(PPh3)]OTf (5), 6' and 10 have been determined. PMID- 11261662 TI - Novel cationic selenium-cluster nitride species [SenN]+(n = 1-11) formed by laser ablation of a Se target in the presence of N2. AB - Nitride cations of selenium clusters [SenN]+ (n = 1-11) were readily produced by laser ablation of a selenium disk that was surrounded by a trace amount of nitrogen seeded in helium and followed by supersonic expansion into a high vacuum. Even at high nitrogen partial pressures, the cluster mononitride cations were found to be essentially the only nitride products in the whole size range we studied. The exception was [Se3N2]+, which is known to be a stable five-membered ring with seven pi electrons. We propose that, in the laser-ablation plasma, the selenium clusters with n > 2 take on a chain conformation, and that the N species links the two ends of the selenium chains, thus forming stable mononitride cations of the cyclic selenium clusters. Their stability is supported by the results of ab initio calculations (at both B3LYP/ 6-31 + G* and MP2/6-31 + G* levels) and of mass-selected cluster-ion photodissociation experiments. PMID- 11261663 TI - Zirconium(IV) complexes of oxydiacetic acid in aqueous solution and in the solid state as studied by multinuclear NMR and X-ray crystallography. AB - The structures of complexes of Zr(IV) and oxydiacetate (ODA2-) in aqueous solutions of pH 0-7 were investigated with the use of 1H, 13C, and 17O NMR spectroscopy. Equilibria of mononuclear [Zr(oda)]2+, [Zr(oda)2], and [Zr(oda)3]2- complexes have been observed. In all complexes ODA2- is bound in a tridentate fashion through the two carboxylate groups and the ether oxygen. No di- or oligonuclear species containing ODA2- were observed. An excess of free Zr(IV) remains in solution, probably as a result of weak electrostatic interactions between negatively charged Zr-ODA complexes or free ODA2- and a positively charged cyclic tetranuclear hydroxy zirconium complex. CP-MAS 13C NMR spectra of solid compounds isolated from the samples indicated that the structures of the [Zr(oda)2] and [Zr(oda)3]2- complexes in solution are similar to those in the solid state. This is corroborated by the single-crystal X-ray structure of Na2[Zr(oda)3] x 5.5 H2O, which was obtained from a solution containing exclusively the [Zr(oda)3]2- complex. In this structure Zr(IV) is nine-coordinate with the three ODA2- ligands bound in a tricapped trigonal prismatic geometry. The negative charge of this [Zr(oda)3]2complex is balanced by two Na+ ions, one of which is on a center of symmetry between delta and lambda enantiomers of [Zr(oda)3]2-. This Na+ is octahedrally coordinated to six (non Zr(IV)-bound) carboxylate oxygen atoms of six different [Zr(oda)3]2- units. PMID- 11261664 TI - Molecular recognition of carbohydrates by artificial receptors: systematic studies towards recognition motifs for carbohydrates. AB - The synthesis and binding properties for carbohydrates of several artificial, acyclic receptors containing two or three heterocyclic recognition units covalently attached to a phenyl spacer is described. These host molecules having uncharged hydrogen-bonding sites were used in a systematic study towards the evaluation of recognition motifs for carbohydrates. A novel effective, acyclic hydrogen-bonding receptor possessing naphthyridine-amide moieties as heterocyclic recognition units has been developed. PMID- 11261665 TI - Chiral bicyclic phosphoramidites--a new class of ligands for asymmetric catalysis. AB - The development of new ligands for catalytic asymmetric C-C bond formation is of great interest to organic synthesis. We describe here a new class of chiral phosphoramidites that embody one or two binaphthol units attached to an achiral azabicyclic [3.3.1] or [3.3.0] framework. These ligands were easily accessible from (R)1,1'-binaphthyl-2,2'-dioxaphosphorchloridite (4) and the corresponding heterobicyclic core 1, 2, or 3. They were employed in enantioselective Cu catalyzed additions of different dialkylzinc reagents to cyclic and acyclic enones. The chiral ketones were obtained with an enantiomeric ratio up to 91:9. The choice of the best ligand proved to be strongly dependent on each substrate. In addition, ligand 6 was found to be the most suitable for Rh-catalyzed hydrogenations of a,beta-unsaturated esters, giving rise to dimethyl 2 methylsuccinate and methyl N-acetylalaninate with enantiomer ratios up to 95:5. PMID- 11261666 TI - Synthesis and spectroscopic and electrochemical characterisation of a conducting polythiophene bearing a chiral beta-substituent: polymerisation of (+)-4,4 bi. AB - A regioregular head-to-head/ tail-to-tail poly(beta,beta'-disubstituted bithiophene) P1 was synthesised by chemical and electrochemical polymerisation of 2,2'-bithiophene that bears (S)-2-methylbutylsulfanyl chains in the beta and beta'-positions. The polymer was characterised by GPC, NMR and UV/Vis spectroscopy, CD, AFM and by electrochemical and conductivity measurements. The CD spectra of P1 in solutions in which poor solvents are present show interesting features and allow the presence of different optically active species to be distinguished. Upon varying the casting conditions of P1, different relative amounts of grainy and homogeneous aggregated phases were observed in AFM micrographies of films and corresponding negative or positive first Cotton effects were found in the CD spectra. AFM, CD and UV/Vis characterisations were also performed on an electrogenerated optically active polymer PE1, in order to make a comparison with the chemically synthesised one. The interesting, small band gap of P1 allows for easy p- and n-electrochemical doping. PMID- 11261667 TI - Beta-alanine-based dendritic beta-peptides: dendrimers possessing unusually strong binding ability towards protic solvents and their self-assembly into nanoscale aggregates through hydrogen-bond interactions. AB - A series of poly(beta-alanine) dendrimers 1-4 with Boc-carbamate as the surface functionality, beta-alanine as the dendritic branch, 3,5-diaminobenzoic acid as the branching agent, and 1,2diaminoethane as the interior core has been synthesized by a solution-phase peptide-coupling method. The structural identities and purities of the products have been fully characterized by spectroscopic and chromatographic methods. 1H NMR studies on the dendrimers indicated that the Boc-carbamate surface groups exist as a mixture of syn and anti rotamers in solution, and that the dendrimers adopt an open structure in polar solvents; this allows the free interaction of the interior core functionality with solvent molecules. Due to the cooperative effect of a large number of carbamate and amide groups, the dendrimers exhibit an unusually strong binding ability towards protic solvents and behave as H-bond sponges. As a result, the H/D exchange rates of the N-H protons are significantly enhanced in such dendritic structures, as compared to those of nondendritic carbamates and amides. These dendritic peptide dendrimers also exhibit a strong tendency to form nanoscopic aggregates in nonpolar or polar aprotic solvents through intermolecular H-bond interactions. PMID- 11261668 TI - "Dimers" and "trimers" of tetrahydroindenes and hexahydroazulenes, respectively generated from [2-(1-cycloalkenyl)ethynyl]carbene complexes (M = W, Cr) by cascade cyclization/cycloaddition reactions. AB - A cascade of cyclization/cycloaddition reactions was triggered by addition of protic oxygen nucleophiles ROH 2 (RO = CH3CO2, PhCO2, PhO) to [2-(1 cyclohexenyl)ethynyl]carbene complexes 1b and 1c (M=W, Cr, respectively), affording highly strained "dimers" 11/11' and "trimers" 12 of the carbene ligand. The first reaction step involved the formation of 1-metalla1,3,5-hexatrienes 7, which readily gave tetrahydroindenes 8 by pi cyclization and extrusion of the metal unit. "Dimers" 11/11' were generated from tetrahydroindenes 8 by a highly exo selective [4+2] cycloaddition of compounds 1b and 1c to afford 1-metalla 1,3,5-hexatriene intermediates 9, and a spontaneous pi cyclization of the latter compounds involving the disengagement of the metal unit. Propenylidene cyclohexenes 13/13' were formed in "ene"-type side reactions to the pi cyclization of 1-metalla-1,3,5-hexatrienes 7, by loss of the metal unit. "Dimers" 11 were transformed into "trimers" 12 by a [4+2] cycloaddition and subsequent pi cyclization of the resulting 1-metalla-1,3,5-hexatriene system. The course of the reaction was elucidated by means of model reactions with (2-phenylethynyl)carbene complex 14, in which 1-metalla-1,3,5-hexatriene intermediates 16 and 17 were isolated and characterized. Alkynyl benzene derivatives 19 were obtained by an unprecedented ring-expansion of a cyclopentadiene unit of "dimers" 11a and 11c, involving the insertion of a carbene carbon atom of compound 14 into a C=C bond. A reaction cascade leading to "dimers" 24/24' could also be triggered by treatment of compounds 2 with [2-(1-cycloheptenyl)ethynyl]carbene tungsten complex 1d. PMID- 11261669 TI - Enaminone substitutents attached to cyclopentadienes: 3E/3Z stereochemistry of 1 metalla-1,3,5-hexatriene intermediates (M = Cr, W) as a functional criterion for the formation of cyclopentadienes and six-membered heterocycles, respectively. AB - Reactions of NH-enaminones 2 with [2-(1-cycloalkenyl)ethynyl]carbene complexes 7 (M=W, Cr) gave tetrahydropentalenes, tetrahydroindenes, and hexahydroazulenes 8a i, in which the NH-enaminone moiety is attached to the cyclopentadiene unit. The reaction involved formation of (3E)-1-metalla-1,3,5-hexatriene intermediates, which underwent pi cyclization faster than 3E/3Z isomerization. Tungsten complexes 12 and 13 were characterized as reaction intermediates. Compounds 8 are potentially bidentate ligands with respect to coordination both of the cyclopentadienyl and the enaminone moieties. PMID- 11261670 TI - Self-assembled helices from 2,2'-biimidazoles. AB - 2,2'-Biimidazoles were synthesized by palladium(0)-catalyzed coupling of 2 iodoimidazoles bearing an alkyl and an ester group at the 4- and 5-positions, respectively. The products were found to be fluorescent and moderately soluble in organic solvents. Three biimidazoles were subjected to single crystal X-ray diffraction analysis. In all three instances, adjacent molecules were found to be bound together in the solid state by pairs of N-H...N hydrogen bonds, forming twisted ribbon-like columns which resemble double helices. The amount of helical twist observed between neighboring biimidazole subunits in these helices varies with the identity of the alkyl and ester groups; in two cases it is approximately 60 degrees, whereas in the third it is about 90 degrees. Mass spectra of six different biimidazoles display ions with masses corresponding to dimers; this indicates that these compounds retain some affinity for each other in the gas phase. The three most soluble biimidazoles also show mass spectrometric peaks ascribable to trimers and tetramers. The solution-phase aggregation tendencies of these latter three compounds were studied by vapor pressure osmometry. In each case, the apparent molecular weight in 1,2-dichloroethane solution is higher than would be expected for free monomers. PMID- 11261671 TI - Asymmetric activation of chiral alkoxyzinc catalysts by chiral nitrogen activators for dialkylzinc addition to aldehydes: super high-throughput screening of combinatorial libraries of chiral ligands and activators by HPLC-CD/UV and HPLC-OR/RIU systems. AB - Asymmetric catalysts, prepared by chiral ligand exchange or chiral modification, can evolve further into highly activated catalysts through engineering with chiral activators. Two new methodologies for "super high-throughput screening" (SHTS) of chiral ligands and activators have been developed as a combination of HPLC-CD/UV (CD/ UV = circular dichroism/ultraviolet spectroscopy) or -OR/RIU (OR/RIU = optical rotation/refractive index unit) with a combinatorial chemistry (CC) factory. With these techniques, the % ee of the product is determined within minutes without separation of the enantiomeric products by using a nonchiral stationary phase. Therefore, those SHTS techniques combined with our 'asymmetric activation' concept can provide a powerful strategy for finding the best activated chiral catalyst. PMID- 11261672 TI - Chiral metal-dithiolene/viologen ion pairs: synthesis and electrical conductivity. AB - Enantiomerically pure dithiolene complexes NBu4[Ni[(R,R)-diotte)2] and NBu4[Ni((S,S)-diotte]2] (diotte2- = a 1,3-dioxolane-tetrathiaethylene), were prepared from the corresponding enantiomers of a diotte2- precursor. The structure of the precursor was solved by single-crystal X-ray analysis; desulfurization afforded a novel tetrathiafulvalene derivative. Combination of the complex monoanion with the enantiomers of the viologen derivative bis(2 methyl-3-hydroxypropyl)-4,4'-dipyridinium (HiBV2+) afforded enantiomeric and diastereomeric ion-pair complexes of the type HiBV[Ni(diotte)2]2. For comparison, the analogous compounds A[Ni(diotte)2]2, (A2+ = methyl (MV2+), octyl (OV2-), stearyl (StV2+) viologen or two 2,2'-bipyridinium acceptors), HiBV-[Ni(diotte)L] [L = mnt2- (maleonitrile-1,2-dithiolate), dmit2- (2-thioxo-1,3-dithiol-4,5 dithiolate)], MV[Ni(dmit)2)]2, [Ni(diotte)2], and [Ni(diotte)(dmit)] were synthesized. An X-ray powder diffraction structural analysis of MV-[Ni(dmit)2)]2 revealed the presence of mixed stacks that contain the sequence anion-anion cation. While no short contacts are observable within a stack, these are observed between the stacks for the dication-anion interaction by short S...H distances in the range of 2.77 to 2.86 A, and for the anion-anion interaction short S...S distances of 3.55 to 3.65 A. In agreement with the absence of intrastack interactions, no ion-pair charge-transfer band can be detected in this and the other complexes. ESR and UV/Vis data suggest that in [Ni(diotte)2]- electron delocalization is less pronounced than in the corresponding mnt2- and dmit2- complexes. The specific electrical conductivity (sigma) of pressed powder pellets ranges from 10(-2) to 10(-12) ohm(-1) cm(-1) and in all cases increases with increasing temperature (293 - 393 K) according to an Arrhenius law. Corresponding activation energies vary from 0.14 to 0.93 eV and increase linearly with log a for structurally similar ion pairs. Charge generation is postulated to occur by disproportionation of the monoanion as suggested by the almost linear increase of log(sigma) with decreasing disproportionation energy. The conductivity of diastereomers of ions with two unlike configurations like [(S,S)-HiBV]-[Ni[(R,R) diotte]2]2 (1.1 x 10(-1) ohm(-1) cm(-1)) is one to two orders of magnitude higher as compared to the diastereomers with two like-configured ions. PMID- 11261673 TI - Thoracocentesis helps diagnose diaphragmatic defects in peritoneal dialysis patients. AB - Hydrothorax is an infrequent complication of peritoneal dialysis. Hydrothorax is often secondary to diaphragmatic defects. Available diagnostic tools are usually insensitive and false-negative results are common. We describe a modification to the standard isotopic peritoneogram that proved effective when other tests yielded false-negative results. We conclude that thoracocentesis prior to the standard isotopic peritoneogram facilitates the diagnosis of diaphragmatic defects. PMID- 11261674 TI - Platelet adenylyl cyclase signaling remains unaltered in children undergoing hemodialysis treatment. AB - Patients with chronic renal failure exhibit multiple endocrine, gastrointestinal and cardiovascular abnormalities, many of which may be explained by alterations of adenylyl cyclase (AC) responsiveness and/or G-protein expression. Since such alterations were previously reported, e.g., for platelets of adult chronic renal failure patients undergoing hemodialysis treatment (HD), we have investigated whether children with chronic renal failure undergoing HD exhibit similar alterations. Eleven uremic children undergoing HD were compared with 11 age matched healthy controls. Platelet AC activity was determined in the absence (basal) and presence of a receptor agonist, direct G-protein activators and direct AC stimulators. G-protein alpha-subunits were measured by quantitative immunoblotting. Basal and stimulated platelet AC and immunoreactivity for platelet G-protein alpha-subunits did not significantly differ between HD and control children. We conclude that HD in children is associated with much smaller, if any, abnormalities of blood cell signal transduction than in adult patients. We speculate that quality of dialysis, age, and underlying disease might differentially influence blood cell signal transduction cascades. PMID- 11261675 TI - A new approach to mRNA in proximal tubule cells of patients with CLCN5 channelopathy. AB - ClC-5 is a chloride channel whose gene mutations have been reported to be associated with X-linked nephrolithiasis (XRN), X-linked recessive hypophosphatemic rickets (XLRH), Dent disease, and idiopathic low-molecular weight proteinuria (ILMWP) in Japanese children. To establish more efficient screening for CLCN5 abnormalities, we developed a new diagnostic method using reverse transcription and polymerase chain reaction (RT-PCR) of cultured renal tubular cells from the urine of patients. Using this new method, we successfully detected microdeletion of ClC-5 mRNA in a patient and splicing abnormality of the CLCN5 Cl channel. PMID- 11261676 TI - Attitude of Belgian pediatricians toward strategy in acute pyelonephritis. AB - To investigate the attitude of Belgian pediatricians toward the management and treatment of children with suspected acute pyelonephritis, a short letter was sent to all Belgian pediatricians (1,200). It contained a brief description of a clinical case strongly suggestive of acute pyelonephritis followed by a series of questions centered on complementary examinations to be performed, need of hospitalization and treatment. A total of 583 responses were received (49%). In the acute phase, 99% of pediatricians perform urine cultures, 87% blood examinations, and 76% renal ultrasound. Dimercaptosuccinate (DMSA) scintigraphy is performed during the acute phase by 37% and during follow-up by 32% of all pediatricians. A voiding cystogram is requested by 71%. Ambulatory treatment is considered by 30% of responders. Amoxicillin/clavulanic acid (44%) and trimethoprim/sulfonamide (22%) are the most frequently used oral antibiotics. Private pediatricians perform fewer examinations and more frequently consider ambulatory treatment of acute pyelonephritis, compared to pediatricians working in hospitals. Among Belgian pediatricians, attitudes toward the diagnosis and treatment of acute pyelonephritis are heterogeneous. This survey underlines the need for properly documented prospective studies for the evaluation of different treatment modalities in childhood acute pyelonephritis. PMID- 11261677 TI - Plasma prorenin levels may predict persistent microalbuminuria in children with diabetes. AB - Diabetic microangiopathy is characterized by increased prorenin concentrations. In the present study, we evaluated plasma prorenin concentrations in a large group of adolescents with onset of diabetes during childhood to determine whether increasing prorenin levels may predict the development of persistent microalbuminuria. Ninety-seven young diabetic patients were studied; they were divided according to the presence of persistent microalbuminuria, at the end of follow-up, into group A and group B (patients who did not develop and who developed persistent microalbuminuria, respectively). One hundred and two healthy subjects, matched for age and sex, were also selected. Patients were followed up for at least 10 years. At the beginning of the study there were no significant differences in prorenin levels between either the two diabetic groups or the healthy controls. During follow-up, an increase in plasma prorenin started at 4 years and became statistically significant (P<0.01) 3 years before the onset of persistent microalbuminuria. No correlation was found between plasma prorenin levels and HbAlc percentages. In conclusion, an increased concentration of prorenin in plasma precedes the elevation of albumin excretion rate (AER) and, therefore, can be useful for identifying patients with onset of diabetes during childhood at risk of developing incipient nephropathy later in life. PMID- 11261678 TI - Idiopathic carpotarsal osteolysis with nephropathy. AB - Idiopathic carpotarsal osteolysis is characterized by gradual lysis and resorption of bones, occurring mainly on hands and feet. It may be sporadic or hereditary and either form can manifest renal involvement. Nephropathy is seen more frequently and is more severe in the sporadic form. We present herein two new cases of unrelated boys with the sporadic form associated with nephropathy. One of the patients had focal segmental glomerulosclerosis. The other patient revealed isolated proteinuria. PMID- 11261679 TI - Folate, vitamin B12, and sulfur amino acid levels in patients with renal failure. AB - We examined the plasma profile of sulfur amino acids (SAA) in patients with chronic renal failure (CRF) and looked for any correlation with serum folate (FA) and/or vitamin B12. Group 1 comprised 9 patients with CRF and glomerular filtration rate (GFR) >20 ml/min per 1.73 m2, 9 patients with GFR<20 ml/min per 1.73 m2 comprised group 2, and 14 patients on hemodialysis group 3. The control group comprised 16 healthy children. Homocysteine (Hcy), methionine (Met), cysteine (Cys), and serine (Ser) were measured with gas chromatography. FA and vitamin B12 were measured using enzymatic immunoassay. Median SAA concentrations were significantly lower in controls than in the three groups of patients. Hcy concentrations were 0.8 micromol/l in controls versus 5 micromol/(group 1), 9 micromol/l (group 2), and 20 micromol/l (group 3). Met concentrations were 26 micromol/l in controls versus 26 micromol/l (group 1), 66 micromol/l (group 2), and 281 micromol/l (group 3). Cys concentrations were 10 micromol/ in controls versus 98 micromol/l (group 1), 54 micromol/l (group 2), and 122 micromol/l (group 3). Ser concentrations were 88 micromol/ in controls versus 153 micromol/l (group 1), 239 micromol/l (group 2), and 240 micromol/l (group 3). The median concentrations of FA were lower in controls than in groups 2 and 3: 5.5 ng/ml versus 8 ng/ml and 15 ng/ml, respectively. Vitamin B12 concentrations did not differ between groups. Vitamin levels did not correlate with SAA. The only difference between patients with Hcy levels in the lower and upper quartile was in Met concentration (38 vs. 263 micromol/l, P<0.02) and GFR (P<0.01). In conclusion, patients with CRF had higher SAA concentrations than healthy children. FA concentrations are higher in CRF patients than in healthy children but did not correlate with concentrations of SAA. PMID- 11261681 TI - Chronic renal failure in Iranian children. AB - We investigated chronic renal failure (CRF) in 166 Iranian children (95 boys and 71 girls) from July 1991 to June 1999. The mean age at onset of CRF was 7.9+/-4.5 years. The most common cause of CRF was congenital urological malformations (78 cases). The second most common cause of CRF was hereditary nephropathy (21%). Glomerular diseases accounted for only 10% of children who later went on to develop renal failure. High rates of cystinosis and primary hyperoxaluria were seen, and these elevated rates could be due to a high prevalence of parental consanguinity. Eighty-six patients required renal replacement therapy, of whom the majority underwent hemodialysis. The prevalence of primary reflux as a cause of CRF was high compared with reports from western countries. Earlier diagnosis and management of urinary tract infections in this group could reduce the prevalence of reflux as a cause of CRF in this population. PMID- 11261680 TI - Normal urinary calcium/creatinine ratios in African-American and Caucasian children. AB - A random urine calcium/creatinine ratio (UCa/Cr) is of practical use in screening for hypercalciuria. However, due to worldwide variations, reference values for the pediatric population are not yet well established. Furthermore, no study has been conducted to establish normal UCa/Cr values in young African-American (AA) children. It has also been previously reported that an elevated UCa/Cr is related to a high urine Na/K ratio (UNa/K). The objectives of the present study were: (1) to set normal values of random UCa/Cr by age and race in the pediatric population of Metropolitan Kansas City, (2) to identify potential racial differences in UCa/Cr between Caucasian (CS) and AA children, and (3) to determine the relationship between UCa/Cr and UNa/K in healthy children. A total of 368 healthy children of both genders were enrolled in the study. They were divided into four age groups as follows: (1) <7 months, (2) 8-18 months, (3) 19 months to 6 years, and (4) 7-16 years. Each group was subdivided into AA and CS. A non-fasting random urine specimen from each subject was analyzed for Ca, Na, K and creatinine. The median UCa/Cr values for AA were: (1) 0.13, (2) 0.09, (3) 0.06, and (4) 0.04 and for CS they were (1) 0.26, (2) 0.11, (3) 0.10, and (4) 0.09. The data showed a strong inverse relationship between UCa/Cr and age, the youngest children demonstrating the highest UCa/Cr. In each age group, UCa/Cr in CS exceeded the corresponding value in AA. The age-dependent 95th percentiles of UCa/Cr values for CS were (1) 0.70, (2) 0.50, (3) 0.28, and (4) 0.20 and for AA they were (1) 0.38 and (3) 0.24. Due to outliers, the 95th percentile could not be established for the other two AA subgroups. The relationship between UCa/Cr and UNa/K was found to be extremely weak in both AA (r2=0.00005) and CS (r2=0.02). On the other hand, a strong linear correlation was observed between UNa/K and age (CS r2=0.23, P<0.001, AA r2=0.19, P<0.001), explaining in part the lack of correlation between UNa/K and UCa/Cr. We conclude that the child's age, ethnicity and geographic location should be taken into consideration when assessing UCa/Cr ratio. Contrary to what has previously been reported in hypercalciuric children, no significant relationship was found between UCa/Cr and UNa/K in healthy children. PMID- 11261682 TI - Urea resolves gross hematuria in a 15 year old with hemoglobin C trait. AB - Hematuria is a rare complication seen in patients with hemoglobin C trait. We report a 15-year-old African-American female with hemoglobin C trait, who presented with persistent hematuria. None of the urological, serological or histological workups revealed any other pathology. Hematuria failed to respond to all conventional modalities used in the treatment of the same condition seen in sickling hemoglobinopathies. This case is the first known case of persistent hematuria in a pediatric patient with hemoglobin C trait, which resolved with intravenous urea administration. PMID- 11261683 TI - Severe renal impairment in the case of classic polyarteritis nodosa. AB - A 14-year-old boy with classic polyarteritis nodosa (cPAN) and a clinical picture resembling rapidly progressive glomerulonephritis (RPGN) is described. He had severe hypertension, malaise, weight loss, fever, myalgia, and rapid deterioration of renal function. Renal biopsy revealed acute necrotizing vasculitis. Angiography showed small saccular aneurysmatic dilatations in the intrarenal branches of the right renal artery and the intrahepatic branches of the hepatic artery. cPAN was diagnosed and pulse methylprednisolone (MP), pulse cyclophosphamide (CYC) and subsequently oral prednisolone were given. Clinical and laboratory findings improved dramatically and remission was attained rapidly. The patient has remained in remission for the last 11 months. cPAN should be considered in patients who present with severe systemic symptoms and hypertension. Progressive renal insufficiency can occur during the acute course of cPAN due to renal vascular involvement without glomerulonephritis. Prompt and aggressive corticosteroid and cytotoxic therapy is essential to suppress disease activity and to maintain remission. PMID- 11261684 TI - End-stage renal disease in a patient with congenital lymphangiectasia and lymphedema. AB - Congenital lymphangiectasia with lymphedema is a disorder constituting the main defect in many different genetic syndromes. Herein we describe a 23-year-old male patient with congenital lymphangiectasia and severe lymphedema of the right leg, scrotum, and abdominal wall, who presented with end-stage renal disease, presumably due to cystic renal lymphangiectasia, and is undergoing chronic hemodialysis treatment. PMID- 11261685 TI - A pharmacokinetic study of Neoral in childhood steroid-dependent nephrotic syndrome. AB - Seven children with steroid-dependent nephrotic syndrome who were on stable remission under Sandimmun therapy were switched to Neoral at the same dosage. During the 4-month follow-up period, two patients relapsed, due to poor compliance in one of them. Serum creatinine remained stable in all patients. Pharmacokinetic profiles were performed at day 0 while on Sandimmun and 4 weeks after conversion to Neoral. Following conversion to Neoral, the peak concentration occurred earlier (2+/-1.4 h vs 3.9+/-2.4 h), and the maximum concentration (677+/-386 ng/ml vs 488+/-265 ng/ml) and the area under the curve (3,082+/-1,536 ng/ml/h vs 2,201+/889 ng/ml/h) were higher. We conclude that Neoral results in an increased bioavailability of cyclosporine (CsA) as compared to Sandimmun in patients with steroid-dependent nephrotic syndrome in remission. PMID- 11261686 TI - Evidence-based assessment of treatment options for children with IgA nephropathies. AB - We present an evidence-based evaluation of published data on therapy for children with various presentations of the IgA nephropathies--idiopathic IgA nephropathy (IgAN) and Henoch-Schonlein purpura nephritis (HSPN). Particular attention has been paid to the outcome markers used in the studies reviewed, with the best evidence provided by markers highly associated with progressive renal failure. No treatment modality for either IgAN or HSPN in pediatric patients has been shown to be effective by a properly designed and administered randomized controlled trial (i.e., the highest level of evidence--level 1). Lower levels of evidence support the use of a variety of corticosteroid regimens, often in combination with other agents, although there are some conflicting studies in this area. No convincing evidence has been published to date to support the use of fish oil, angiotensin-converting enzyme inhibitors or tonsillectomy for the treatment of children with IgAN or HSPN. Well designed randomized controlled trials in children with the IgA nephropathies need to be undertaken. PMID- 11261687 TI - New insights: nephronophthisis-medullary cystic kidney disease. AB - Nephronophthisis (NPH) and medullary cystic kidney disease (MCKD) constitute a group of renal cystic diseases, which share a common characteristic renal histologic triad of tubular basement membrane disintegration, tubular atrophy with cyst development, and interstitial cell infiltration with fibrosis. The different disease variants lead to chronic renal failure with onset at characteristic age ranges for recessive NPH and dominant MCKD. There is extensive gene locus heterogeneity with at least three different loci for nephronophthisis (NPHP1, NPHP2, and NPHP3) and two different loci for MCKD (MCKD1 and MCKD2). Juvenile nephronophthisis, in addition, can be associated with extrarenal organ involvement. We have identified by positional cloning the gene (NPHP1) for juvenile nephronophthisis (NPH1), as a first step towards understanding the pathogenesis of this disease group. Its gene product, nephrocystin, is a novel protein, which contains a src-homology 3 (SH3) domain. We put forward a hypothesis that the pathogenesis of NPH might be related to signaling processes at focal adhesions (the contact points between cells and extracellular matrix) and/or adherens junctions (the contact points between cells). PMID- 11261688 TI - Angiopoietin growth factors and Tie receptor tyrosine kinases in renal vascular development. AB - Angiopoietin-1 (Ang-1) is a secreted growth factor which binds to and activates the Tie-2 receptor tyrosine kinase. The factor enhances endothelial cell survival and capillary morphogenesis, and also limits capillary permeability. Ang-2 binds the same receptor but fails to activate it: hence, it is a natural inhibitor of Ang-1. Ang-2 destabilises capillary integrity, facilitating sprouting when ambient vascular endothelial growth factor (VEGF) levels are high, but causing vessel regression when VEGF levels are low. Tie-1 is a Tie-2 homologue but its ligands are unknown. Angiopoietin and Tie genes are expressed in the mammalian metanephros, the precursor of the adult kidney, where they may play a role in endothelial precursor growth. Tie-1-expressing cells can be detected in the metanephros when it first forms and, based on transplantation experiments, these precursors contribute to the generation of glomerular capillaries. During glomerular maturation, podocyte-derived Ang-1 and mesangial-cell-derived Ang-2 may affect growth of nascent capillaries. After birth, vasa rectae acquire their mature configuration and Ang-2 expressed by descending limbs of loops of Henle would be well placed to affect the growth of this medullary microcirculation. Finally, preliminary data implicate angiopoietins in deregulated vessel growth in Wilms' kidney tumours and in vascular remodelling after nephrotoxicity. PMID- 11261689 TI - Congenital nephrotic syndrome. AB - A female infant born at 34 weeks' gestation after several days of ruptured membranes had a Potter-like face and compression-induced limb posture consistent with oligohydramnios. Oedema developed on day 2; initial investigations showed massive proteinuria, hypoalbuminaemia, hyponatraemia, acidosis and marked renal insufficiency. The infant was intubated and despite albumin infusion and intravenous antibiotics she became oligoanuric by day 8 and required haemofiltration. Renal biopsy at this stage showed cystic dilatation of tubules in the cortex and glomerular lesions consisting of shrunken tufts with sclerotic centres and a corona of epithelial cells at the periphery. Due to a very poor prognosis treatment was withdrawn. Postmortem examination of the kidneys confirmed the histological diagnosis of diffuse mesangial sclerosis. Genetic studies found no mutations in WT1 and NPHS1 genes although the entire genes could not be screened for mutations due to lack of DNA. PMID- 11261690 TI - Congenital nephrotic syndrome: commentary. PMID- 11261691 TI - Recommendations for the training of European Pediatric Nephrologists by the European Society for Paediatric Nephrology. An outline of the minimal requirements for accreditation in the European Economic Community. PMID- 11261692 TI - Intravenous cyclosporine A for patients with nephrotic syndrome. PMID- 11261693 TI - Childhood crush syndrome sustained in the 1999 Marmara earthquakes. PMID- 11261694 TI - Neisseria meningitidis infection in nephrotic children. PMID- 11261695 TI - Epidemiology of hepatitis C virus infection. PMID- 11261696 TI - Diagnostic tests for hepatitis C virus infection. PMID- 11261697 TI - The natural history of hepatitis C virus infection. PMID- 11261698 TI - Hepatitis C. PMID- 11261699 TI - Extrahepatic symptoms of hepatitis C virus infection: relation to autoimmune response. PMID- 11261700 TI - Glomerular disease associated with hepatitis C virus infection in native kidneys. PMID- 11261701 TI - Treatment of glomerulonephritides associated with hepatitis C virus infection. PMID- 11261702 TI - Epidemiology and mechanisms of transmission of the hepatitis C virus in haemodialysis. PMID- 11261703 TI - Hepatitis C virus-induced liver disease in dialysis patients. PMID- 11261704 TI - Treatment of hepatitis C virus infection in dialysis patients. PMID- 11261705 TI - Hepatitis C and management of end-stage renal disease. PMID- 11261706 TI - Epidemiology and mode of transmission of hepatitis C virus infection after renal transplantation. PMID- 11261707 TI - Systematic evaluation of liver disease in hepatitis C virus-infected renal transplant recipients: clinical and pathological study. PMID- 11261708 TI - Hepatitis C virus infection in the renal transplant recipient. PMID- 11261709 TI - Glomerulonephritis associated with hepatitis C virus infection after renal transplantation. PMID- 11261710 TI - Hepatitis C virus-positive patients on the waiting list for transplantation: study, evaluation and treatment. PMID- 11261711 TI - Policies concerning the use of kidneys from donors infected with hepatitis C virus. PMID- 11261712 TI - Treatment of hepatitis C virus infection after renal transplantation: new insights. PMID- 11261714 TI - Positional effect of solute functional group among positional isomers in hydroxyl group-solvent and carbonyl group-solvent specific interactions in menthanol- water mixed solvents monitored by high-performance liquid chromatography. AB - We have evaluated the hydroxyl group-solvent and carbonyl group-solvent specific interactions by using mostly an Alltima C18 stationary phase and subsidiarily squalane-adsorbed C18 phase, and by measuring the retention data of carefully selected solutes in 60:40, 70:30, 80:20 and 90:10 (%, v/v) methanol-water eluents at 25, 30, 35, 40, 45 and 50 degrees C. The selected solutes are four positional isomers of phenylbutanol, 5-phenyl-1-pentanol, three positional isomers of alkylarylketone derived from butylbenzene, and 1-phenyl-2-hexanone. The magnitudes of carbonyl group-solvent specific interaction enthalpies are larger than those of hydroxyl group-solvent specific interaction enthalpies in general. We observed clear discrepancies in functional group-solvent specific interactions among positional isomers. The spatial accessibility of the functional group by the solvent molecules seems to govern the strength of interaction. The relationships between molecular structures and functional group accessibilities have been discussed. The specific functional group-mobile phase interactions obtained by the Alltima C18 stationary phase were systematically different from those obtained by the squalane-impregnated C18 stationary phase, which may be due to structural differences between the two phases. PMID- 11261713 TI - Retention of ionizable compounds on high-performance liquid chromatography. VII. Characterization of the retention of ionic solutes in a C18 column by mass spectrometry with electrospray ionization. AB - The elution of ions from a C18 column with mobile phases containing methanol (60%, v/v) and aqueous buffers is studied by mass spectrometry. It is demonstrated that the anions are excluded from the stationary phase by the ionized silanols. However, the ionized silanols interact strongly with cations, which are retained in the column. These cations are later eluted from the column by ion exchange with the cations present in the pH buffered mobile phase. The size of the ions, the mobile phase cation concentration and the mobile phase pH are the main parameters that affect elution of the retained cations. It is also demonstrated that there are at least two different types of ionizable silanols, with different acidities, that contribute to the retention of cations. An estimate of the pKa values of these two groups of silanols in 60% methanol is given. PMID- 11261715 TI - Liquid exclusion-adsorption chromatography: new technique for isocratic separation of nonionic surfactants. I. Retention behaviour of fatty alcohol ethoxylates. AB - A new technique of liquid chromatography is described, which allows a baseline separation of fatty alcohol ethoxylates with 15-20 ethylene oxide units under isocratic conditions. The new method is based on a combination of two different chromatographic modes for the individual structural units: size exclusion for the poly(oxyethylene) chain and adsorption interaction for the hydrophobic end fragments. A theory is provided for this mixed exclusion-adsorption mode of liquid chromatography. Chromatographic data are found to be in a good agreement with this theory. PMID- 11261716 TI - Multi-beam polarized photometric detector for high-performance liquid chromatography. AB - A novel multi-beam polarized photometric detector (PPD) for high-performance liquid chromatography (HPLC) is described. By pairing a polarizing prism with a thin quartz plate as a retarder, many linear polarized beams are produced at every 1/2 wavelength of the plate, and the polarizing axes of the adjacent beams intersect each other. The addition of another prism inclining its polarizing axis by pi/4 against the first one enables the simultaneous measurement of optical rotations based on the PPD at many wavelengths. The combination of these optics with a photo-diode array detector can be used to construct a modulated type polarimeter. This detector is designed to measure the optical rotation of an analyte at its absorption band. The spline function connecting the points at 1/4 wavelengths of the plate was used as a baseline to extract the PPD waves. The use of the similarity factor as a noise filter gave high sensitivity. Application of the proposed technique to an analyte carrying the Cotton absorption band provided good results. PMID- 11261717 TI - Impact of the post-treatment conditions of parent silica on the silanization of n octadecyl bonded silica packings in reversed-phase high-performance liquid chromatography. AB - Native mesoporous silica beads were subjected to a sequence of post-treatment procedure including hydrochloric acid treatment, calcination and subsequent rehydroxylation. The post-treated silica beads were converted into RP-18 silica by silanization with monochloro- and dimethoxy-n-octadecylsilanes, respectively. The influence of post-treatments and silanization conditions on the physico chemical characteristics and on the chromatographic behaviour of the RP-silicas was studied. Also the changes of the pore structural parameters and the silanol group densities during the post-treatment and silanization were assessed. PMID- 11261718 TI - Spectroscopic characterisation and identification of ecdysteroids using high performance liquid chromatography combined with on-line UV--diode array, FT infrared and 1H-nuclear magnetic resonance spectroscopy and time of flight mass spectrometry. AB - A prototype multiply hyphenated reversed-phase HPLC system has been applied to the analysis of a mixture of pure ecdysteroids and an ecdysteroid-containing plant extract. Characterisation was achieved via a combination of diode array UV, 1H NMR, FT-IR spectroscopy and time of flight (TOF) mass spectrometry. This combination of spectrometers allowed the collection of UV, 1H NMR, IR and mass spectra for a mixture of pure standards enabling almost complete structural characterisation to be performed. The technique was then applied to a partially purified plant extract in which 20-hydroxyecdysone and polypodine B were identified despite incomplete chromatographic resolution and the presence of co chromatographing interferents. The experimental difficulties in the use of such a systems for these analytes are described. PMID- 11261719 TI - Liquid chromatographic enantioseparation of beta-blocking agents with (1R,2R)-1,3 diacetoxy-1-(4-nitrophenyl)-2-propyl isothiocyanate as chiral derivatizing agent. AB - The applicability of (1R,2R)-1,3-diacetoxy-1-(4-nitrophenyl)-2-propyl isothiocyanate [(R,R)-DANI] as a recently developed chiral derivatizing agent for the enantioseparation of a series of beta-blockers is described. The thiourea diastereomers formed were analyzed by reversed-phase high-performance liquid chromatography, mixtures of water and methanol or acetonitrile being used for elution. Conditions of derivatizations (temperature, reagent excess and reaction time) were optimized, and the effects of organic modifiers on the retention and separation were investigated; the diastereomers could readily be baseline separated with methanol-containing mobile phases with resolutions between 1.58 and 2.72. PMID- 11261720 TI - Determination of anthocyanins in wine based on flow-injection, liquid--solid extraction, continuous evaporation and high-performance liquid chromatography- photometric detection. AB - A continuous method, easy to automate, for the determination of anthocyanins in wine based on the coupling of continuous liquid-solid extraction, evaporation, HPLC individual separation and photometric detection is proposed. The target analytes are removed from the wine in a continuous fashion using a C18 minicolumn and eluted with an aqueous solution (pH 2) with 16% acetonitrile. The eluted fraction is concentrated by solvent evaporation assisted by heat and dragging off the vapour using a flow of N2. For in-line preconcentration, a continuous evaporation module was designed and located in the manifold between the solid phase minicolumn and the injection valve of the chromatograph. In this way, injection of the sample into the dynamic system leads the plug through it for liquid-solid extraction of the anthocyanins, partial evaporation of the eluent (with a preconcentration factor as required) and transport to the high-pressure injection valve of the chromatograph, where individual separation and subsequent photometric detection take place. The method thus developed for the determination of malvidin-3-glucoside, cyanidin-3-glucoside and peonidin-3-glucoside anthocyanins in Spanish red wines is more sensitive than the batch manual method based on the same steps, has better linearity of the calibrations curves with lower detection limits and much wider determination range for the most abundant anthocyanins in wine. In addition, the method can be fully automated with low acquisition and maintenance costs. PMID- 11261721 TI - Determination of phenolic acids and flavonoids of apple and pear by high performance liquid chromatography. AB - A new HPLC stationary phase has been applied to the analysis of phenolic acids and flavonoids with diode array and mass spectrometric detection. The separation of 26 standard compounds was achieved within 1 h. The stationary phase displayed excellent resolution especially of flavonol glycosides. The analytical system has been used for the determination of phenolic compounds in apple pomace and apple juice, and in extracts of pear fruits of different cultivars. Apple pomace was found to be a promising source of phenolics. However, yields are affected by the drying conditions applied. Furthermore, the applicability of the analytical system for the authenticity control of apple and pear juice was demonstrated by determination of characteristic quercetin and isorhamnetin glycosides, and dihydrochalcones, respectively. Since isorhamnetin-3-glucoside was present in all pear cultivars investigated, the usefulness of arbutin as a specific marker of pear products appears to be doubtful. PMID- 11261722 TI - Genetically engineered peptide fusions for improved protein partitioning in aqueous two-phase systems. Effect of fusion localization on endoglucanase I of Trichoderma reesei. AB - Genetic engineering has been used for fusion of the peptide tag, Trp-Pro-Trp-Pro, on a protein to study the effect on partitioning in aqueous two-phase systems. As target protein for the fusions the cellulase, endoglucanase I (endo-1,4-beta Dglucan-4-glucanohydrolase, EC 3.2.1.4, EGI, Cel7B) of Trichoderma reesei was used. For the first time a glycosylated two-domain enzyme has been utilized for addition of peptide tags to change partitioning in aqueous two-phase systems. The aim was to find an optimal fusion localization for EGI. The peptide was (1) attached to the C-terminus end of the cellulose binding domain (CBD), (2) inserted in the glycosylated linker region, (3) added after a truncated form of EGI lacking the CBD and a small part of the linker. The different constructs were expressed in the filamentous fungus T. reesei under the gpdA promoter from Aspergillus nidulans. The expression levels were between 60 and 100 mg/l. The partitioning behavior of the fusion proteins was studied in an aqueous two-phase model system composed of the thermoseparating ethylene oxide (EO)-propylene oxide (PO) random copolymer EO-PO (50:50) (EO50PO50) and dextran. The Trp-Pro-Trp-Pro tag was found to direct the fusion protein to the top EO50PO50 phase. The partition coefficient of a fusion protein can be predicted with an empirical correlation based on independent contributions from partitioning of unmodified protein and peptide tag in this model system. The fusion position at the end of the CBD, with the spacer Pro-Gly, was shown to be optimal with respect to partitioning and tag efficiency factor (TEF) was 0.87, where a fully exposed tag would have a TEF of 1.0. Hence, this position can further be utilized for fusion with longer tags. For the other constructs the TEF was only 0.43 and 0.10, for the tag fused to the truncated EGI and in the linker region of the full length EGI, respectively. PMID- 11261723 TI - Determination of macrolide antibiotics by liquid chromatography. AB - The liquid chromatographic separation of seven macrolides used in food producing animals in the European Union has been studied. Separation was performed by using an end-capped high-purity silica-based C18 column and mobile phases consisting of phosphate buffer (pH 2.5)-acetonitrile mixtures. The effect of pH and acetonitrile percentage on the separation was examined. Two UV-based detection systems, wavelength programming and diode array, were assayed. Detection limits were in the range 6-33 microg l(-1) for spiramycin, tilmicosin, tylosin, kitasamicin and josamicin and about 400 microg l(-1) for erythromycin and oleandomycin. The suitability of the method for multiresidue determination of the five macrolides is demonstrated by the analysis of spiked samples of chicken muscle. PMID- 11261724 TI - Mass spectrometric confirmation criterion for product-ion spectra generated in flow-injection analysis. Environmental application. AB - The suitability of a confirmation criterion recently recommended in the Netherlands for gas chromatography with mass spectrometric detection (GC-MS), was evaluated for flow-injection analysis (FIA) with atmospheric pressure chemical ionisation MS-MS detection. The main feature of the criterion is that the relative ion abundances of the four diagnostic ions are taken into account. That is, for lower-intensity peaks, relative standard deviations may be higher; this is an advantage with chemical ionisation MS procedures. A series of triazines and their degradation products were used as test compounds. Tap and surface water samples spiked at 0.33 microg/l were analysed by means of a selected reaction monitoring MS-MS procedure. For all analytes but hydroxysimazine (3 transitions), 4-9 transitions could be selected which invariably met the demands of the criterion. Some of the transitions used originate from the 37Cl isotopic mass of the parent compounds which provides additional structural information. Data for twenty surface water samples analysed by means of FIA-MS-MS as well as GC-MS and liquid chromatography with diode array UV and MS-MS detection gave essentially the same results over the 0.1-1.0 microg/l range. In two samples desethylatrazine was reported by FIA-MS-MS whereas this compound was not detected by GC-MS. For a first test, this is a promising result. PMID- 11261725 TI - Sensitive and selective ion chromatographic method for the determination of trace beryllium in water samples. AB - A selective and sensitive ion chromatographic method has been developed for the determination of beryllium in a number of water samples at low-microg/l concentrations. The separation was performed on a 250x4.0 mm I.D. iminodiacetic acid functionalised silica gel column. Chromatographed Be(II) was detected using visible detection at 590 nm following post-column reaction with chrome azurol S (CAS). The optimum separation and derivatisation conditions were studied in detail. The optimum eluent conditions were found to be 0.4 M KNO3, adjusted to pH 2.5 using HNO3, with optimum post-column detection being achieved using a solution containing 0.26 mM CAS, 2% Triton X-100, 50 mM 2-(N morpholino)ethanesulfonic acid, pH 6.0. Under the above conditions, the concentration detection limit for Be(II) was found to be 3 microg/l in a standard solution and 4 microg/l in a typical tap water sample, using a 250 microl injection. The method was linear over the investigated range of 10 microg/l to 10 mg/l and highly reproducible. The method was successfully applied to a number of water samples of varying matrix complexity, including simulated seawater, and also to a natural freshwater certified reference material NIST 1640. PMID- 11261726 TI - Study of a novel cationic calix[4]arene used a selectivity modifier in capillary electrophoresis with electrochemical detection. AB - The use of a novel cationic calixarene, p-(quaternary ammonium) calix[4]arene, as selectivity modifier in capillary electrophoresis coupled with electrochemical detection was reported. The calixarene displayed good selectivity for the positional isomers of benzenediol and aminophenol and their successful separation was obtained under optimum conditions. The interaction mechanism between p (quaternary ammonium) calix[4]arene and the solutes is discussed using the molecular modeling method. The detection limits by electrochemical detection for the most solutes studied here were below picogram level, which was approximately 2 orders of magnitude lower than those reported in the literature using UV detection. The results showed that electrochemical detection is especially suitable for an electrophoresis system where calixarenes are used as modifier. PMID- 11261727 TI - Analysis of sialo-N-glycans in glycoproteins as 1-phenyl-3-methyl-5-pyrazolone derivatives by capillary electrophoresis. AB - A method for the analysis of the sialo-N-glycans in glycoproteins was established by the electrokinetic chromatography mode of capillary electrophoresis (CE) in sodium dodecyl sulfate (SDS) micelles as 1-phenyl-3-methyl-5-pyrazolone (PMP) derivatives, using sialo-N-glycans in fetuin as a model. Six major and some minor peaks were observed for the N-glycans in fetuin, which were well separated from each other using 50 mM phosphate buffer, pH 6.0, containing SDS to a concentration of 30 mM in an uncoated fused-silica capillary, and these peaks were assigned to sialo-N-glycans having either of the biantennary or beta1 3/beta1-4 linked galactose-containing complex type triantennary N-glycans as the basic structures, by an indirect method based on the assignment of the peaks in high-performance liquid chromatography separated in parallel with CE and peak collation between these two separation methods. The attaching position of the sialic acid residue was determined using the linkage preference of neuraminidase isozymes. The established system is considered to be useful for routine analysis of microheterogeneity of the carbohydrate moiety of this model glycoprotein from the following reasons: (1) the derivatization with PMP proceeds quantitatively under mild conditions without causing release of the sialic acid residue, (2) the derivatives can be sensitively detected by UV absorption, (3) the procedure is simple, rapid and reproducible. Preliminary results of N-glycan analysis for several other glycoproteins under these conditions are also presented. PMID- 11261728 TI - Determination of low-molecular-mass carboxylic acids in atmospheric aerosol and vehicle emission samples by capillary electrophoresis. AB - A method is developed for the determination of a large number of airborne and vehicle-emitted low-molecular-mass mono- and dicarboxylic acids using capillary electrophoresis with indirect UV detection. A background electrolyte (BGE) consisting of 2,6-naphthalenedicarboxylic acid and tetradecylmethylammonium bromide, adjusted to pH 6.2 with 2,2-bis(hydroxymethyl)-2,2',2" nitrilotriethanol, is employed. Separations are robust using the buffered BGE, proper rinse steps, and constant current mode with migration time variations less than 3% RSD on a day-to-day basis, using different capillaries and performed by different analysts. Detection limits are at the tens of microg/l level using pressure injection. A comparison of the CE method with ion chromatography is also made. PMID- 11261729 TI - Dual effect of high electric field in capillary electrophoresis study of the conformational stability of Bungarus fasciatus acetylcholinesterase. AB - The effect of high electric field in capillary zone electrophoresis (CZE) was evaluated for the study of the thermally induced unfolding of Bungarus fasciatus acetylcholinesterase. This monomer enzyme is characterised by two interdependent uncommon structural features, the asymmetrical distribution of charged residues and a relatively low thermal denaturation temperature. Both traits were presumed to interfere in the thermal unfolding of this enzyme as investigated by CZE. This paper analyses the effect of high electric field on the behaviour of the enzyme native state. It is shown that increasing the applied field causes denaturation like transition of the enzyme at a current power which does not induce excessive Joule heating in the capillary. The susceptibility to electric field of proteins like cholinesterases, with charge distribution anisotropy, large permanent dipole moment and notable molecular flexibility associated with moderate thermal stability, was subsequently discussed. PMID- 11261730 TI - New chiral crown ether stationary phase for the liquid chromatographic resolution of alpha-amino acid enantiomers. AB - A new chiral stationary phase (CSP) for the liquid chromatographic separation of enantiomers was prepared by bonding a novel enantiopure (diphenyl-substituted 1,1'-binaphthyl) crown ether to 5 microm silica gel. The resulting CSP was applied to the separation of the enantiomers of various natural and unnatural alpha-amino acids. All alpha-amino acids tested were resolved very well on the new CSP, with the exception of proline, which does not contain a primary amino group. The resolution of alpha-amino acid enantiomers on this new CSP was found to be dependent on the type and amounts of organic and acidic modifiers, and on column temperature. PMID- 11261731 TI - New method for high-performance liquid chromatographic separation and fluorescence detection of ginsenosides. AB - A novel pre-column derivatization method for the quantitative determination of ginsenosides by HPLC with fluorescence detection was established. The double bond at the C24-C25 position of ginsenoside was converted into an aldehyde group by means of ozonolysis. Then the aldehyde group reacts with FMOC-hydrazine forming the ginsenoside FMOC-hydrazone. The derivatized products were separated by RP HPLC with gradient elution. The detection limits of ginsenosides Rg1 and Rb1 were 2.0 ng (about 2.5 pmol) and 1.0 ng (about 0.9 pmol), respectively. This method can be used for all ginsenosides having the C24-C25 double bond. PMID- 11261732 TI - Fluorimetric determination of magnesium and aluminum via complexation with oxine in high-performance liquid chromatography. AB - Aluminum and magnesium were determined by fluorimetric detection via pre-column and/or in-column derivatization with 8-hydroxyquinoline (oxine) in high performance liquid chromatography. The oxine complex of aluminum was selectively detected when the eluent contained no oxine, whereas the aluminum and magnesium complexes could be simultaneously detected when the eluent contained oxine. The sensitivity was improved by using eluents containing oxine by a factor of 7.1 for aluminum, and the detection limits at S/N=3 were 18 and 16 ng/ml for magnesium and aluminum, respectively. The present system was applied to the determination of magnesium and aluminum in various water samples. PMID- 11261733 TI - Method for determination of methyl tert-butyl ether in gasoline by gas chromatography. AB - A simple method for the determination of methyl tert-butyl ether (MTBE) in gasoline has been developed. The separation of MTBE from other analytes was controlled by the use of gas chromatography-mass spectrometry in the full scan mode using the characteristic primary, secondary and tertiary ions m/z 73, 57 and 43. The sample mass spectrum did not show any superimposition of other analytes. The separation from the common gasoline component 2-methylpentane was sufficient for reliable quantitation. An application of the developed conditions using gas chromatography with flame ionization detection was performed by the analysis of regular, euro super, super premium unleaded and 'Optimax' gasoline from petrol stations in the area of Frankfurt/Main, Germany. Regular unleaded gasoline shows an average MTBE content of 0.4% (w/w), whereas the MTBE content in euro super gasoline varies between 0.4 and 4.2% (w/w). The blending of MTBE to super premium has increased from 8.2% (w/w) in 1998 to 9.8% (w/w) on average in 1999. The recently introduced gasoline 'Optimax' shows an average MTBE content of 11.9% (w/w). The presented method might also be used for the analysis of other ethers, such as ethyl tert-butyl ether, which requires the use of another internal standard. PMID- 11261734 TI - Risk factors and pathogenesis of cholesterol gallstones: state of the art. AB - The aim of this article is to present an update of selected aspects of the pathogenesis and risk factors of cholesterol gallstones, a highly prevalent Western disease. The etiology of cholesterol cholelithiasis is considered to be multifactorial, with interaction of genetic and environmental factors. Mechanisms of cholesterol lithogenesis include biliary cholesterol hypersecretion, supersaturation and crystallization, stone formation and growth, and bile stasis within the gallbladder. Each of these various steps could be under genetic control and/or be influenced through intermediate pathogenic steps linked to a variety of environmental factors. PMID- 11261735 TI - State of the art and new perspectives on dry powder inhalers. AB - Modern local therapy for lung diseases is now largely based on pressurized metered-dose inhalers (MDIs). The research of alternatives to MDIs has recently accelerated, primarily due to environmental concerns related to the use of chlorofluorocarbon (CFC) propellants. The most recent and attractive solution to this problem is represented by the development of dry powder inhalers (DPIs), particularly designed to avoid the use of propellants. DPIs have been developed for specific products, therefore they possess a reduced versatility in term of application of the same device to different drugs. However, they did introduce new concepts in pulmonary drug delivery, solving some disadvantages of the pressurized devices. They are in their infancy and the efforts of researchers are now impressive. The future will certainly see many other devices containing additional innovative features for the effective respiratory delivery of drug. The goals still remain the delivery of precise and uniform drug doses and increasing the respirable fraction in relation to the dose emitted from the device. PMID- 11261736 TI - Tolerability of doxofylline in the maintenance therapy of pediatric patients with bronchial asthma. AB - A retrospective open study was performed to ascertain the tolerability of doxofylline in pediatric patients with bronchial asthma or airway obstruction complicating acute bronchitis. The study population included 806 patients aged between 3 and 16 years. Doxofylline (200 mg sachets) was administered at daily doses ranging from 100 to 400 mg. The percentage of patients reporting side effects was 11%. The percent of patients reporting moderate side effect was 5%, the others being mild. The percent of patients reporting adverse event very likely due to doxofylline was 6%. The percent of patient drop-outs related to side effects was 5%. PMID- 11261737 TI - A multidisciplinary approach to the treatment of small-cell lung cancer: the role played by surgery. AB - The authors report their data on 344 cases of small-cell lung cancer treated according to indications with combined chemoradiotherapy and in selected cases with surgical intervention. In patients with limited disease, the results of pharmacologic therapy significantly improve the prognosis only in association with surgery. The role of surgery has been reappraised in the treatment of small cell lung cancer which appears, nowadays, multidisciplinary. PMID- 11261738 TI - Hyperthyroidism and concurrent thyroid carcinoma. AB - In the last twenty years, medical studies have reported a significant increase in thyroid neoplasms among patients with hyperthyroidism. Aim of the present work is to reconsider the real incidence of this not uncommon association and to establish a model for surgical treatment of hyperthyroidism for a possible concurrence with thyroid carcinoma. At the Department of Surgical Sciences and Applied Medical Technologies "La Sapienza" Rome's University, during the period 1994 to 1999, an homogeneous group of 82 patients was surgically treated for hyperthyroidism. Of our patients, fifty-four (66%) had a "multinodular toxic goiter" (MTG), twenty (24%) a "functional autonomous nodule" (FAN) while the remaining patients were affected by Graves' disease. The surgical procedures adopted were: 1) total extracapsular ipsilateral lobectomies and isthmectomies in sixteen patients with FAN; 2) total extracapsular thyroidectomy in all patients with MTG and with Graves' disease and in the remaining four patients with FAN after a long time treatment with thyrostatic drugs. On six (7%) of our patients we found out a thyroid carcinoma: five with MTG and one with Graves' disease. However, no association with thyroid carcinoma was observed in anyone with FAN. The correct treatment of thyroid surgical diseases is a single definitive operative approach. The procedure must be a total thyroidectomy in MTG and Graves' disease. However, in patients with FAN it's possible, after careful evaluation, to carry out a total extracapsular ipsilateral lobectomy with isthmectomy, justified by the normal morphology of the remaining thyroid tissue. It is always possible, in these cases, a subsequent complete exeresis if a carcinoma is present in the removed lobe. PMID- 11261740 TI - Evidence of melatonin involvement in pindolol-induced suppression of REM sleep. AB - The effects of pindolol, melatonin, and the melatonin receptor agonist agomelatine were studied in rats implanted for chronic sleep procedures. Administration of pindolol (1.0-4.0 mg/kg) during the light phase induced a significant reduction of rapid-eye-movement sleep (REMS) and an increase of waking (W). In the rats recorded after receiving 1.0-6.0 mg/kg melatonin no significant differences were found in sleep or W compared with controls. Agomelatine (1.0-6.0 mg/kg) induced a significant increase of light sleep during the first 3 h of the recording period. Pretreatment with melatonin partly prevented the pindolol-induced suppression of REMS. However, agomelatine was ineffective in this respect. Overall, these data suggest that the decreased production of melatonin could play a role in REMS suppression related to pindolol administration. PMID- 11261739 TI - Management of antihypertensive treatment with Lisinopril: a chronotherapeutic approach. AB - Risk for cardiovascular events seems to be higher in the early morning, also as consequence of a rise in blood pressure (BP) values due to the characteristic circadian pattern of BP variability. Therefore, a suitable therapeutic BP control should be tightest during the early morning. On the basis of the ambulatory blood pressure monitoring (ABPM) studies, it has been previously demonstrated that the antihypertensive effect of once daily drug, generally administrated in the morning, decreases at the end of the dosing period. A chronotherapeutic approach to the management of hypertension (this field has been pourly investigated so far) would allow the assessment of the optimum timing of drug dosing, according to the circadian BP rhythm and to the chronorisk maps, in hypertensive patients affected by associated vascular pathologies. This would increase the therapeutic effects. The aim of this study was to assess BP changes due to ACE-inhibitor (Lisinopril 20 mg/die) once daily administration at three different times (8.00 AM, 4.00 PM, 10.00 PM), in order to optimise the dosing time. 40 subjects (mean age +/- SD: 45 +/- 10) affected by primary mild to moderate hypertension were submitted to ABPM for 24 hours, by means of Spacelabs 90207, before and after pharmacological treatment. Patients were randomised to take the drug at 8.00 AM, 4.00 PM or 10.00 PM, and they repeated ABPM every two months, by changing the dosing time. The chronobiological analysis showed: 1) a sensible decrease both in Systolic (S)BP and Diastolic (D)BP without affecting the circadian rhythm, in all evaluations; 2) a greater reduction of SBP and DBP from 6.00 AM to 11.00 AM, period in which cardiovascular risk is higher, after 10.00 PM dosing; 3) no other sensible reduction in SBP and DBP occurred after night administration as compared to that caused by other dosing times. Lisinopril administration at 10.00 PM. has been shown to be much more useful since, although BP circadian rhythm was unmodified, it protects hypertensive patients from both vascular chronorisk and Cruickshank effect (J-curve). Therefore, a chronobiologist antihypertensive treatment in order to increase the therapeutic effect already obtained with the traditional statistic methods. PMID- 11261741 TI - Postnatal maturation of prefrontal pyramidal neurones is sensitive to a single early dose of methamphetamine in gerbils (Meriones unguiculatus). AB - The effect of a single methamphetamine application on postnatal maturation of the prefrontal cortex was studied using pyramidal cell morphology and spine density as parameters of systemic plasticity. Male gerbils were injected a single dose of methamphetamine (METH, 50mg/kg, i.p.) on postnatal day 14. On postnatal day 90, prefrontal cortices of METH-treated animals and saline-treated controls were processed for Golgi-staining. Dendritic arbours of layer III and V pyramidal neurones were measured to describe pyramidal cell morphology, and segmental spine counts were carried out. The results showed that a single postnatal METH challenge significantly alters morphological differentiation of pyramidal cells towards adulthood. Cells from METH-treated animals showed a higher total dendritic length based on longer segments between subsequent dendritic branchings, with only the apical stem dendrite being shorter in METH-treated than in control subjects. The branching rate was slightly but not significantly increased in METH-treated animals. Nevertheless, spine density was significantly increased on all types of dendrites, with apical dendrites of both layers III and V showing the highest drug-induced progression of about 50% compared to control values. The present results are discussed with regard to probable clues they may provide for investigating neurobiological principles of psychotic behaviour in an animal model. PMID- 11261742 TI - Enantio-specific induction of apoptosis by an endogenous neurotoxin, N methyl(R)salsolinol, in dopaminergic SH-SY5Y cells: suppression of apoptosis by N (2-heptyl)-N-methylpropargylamine. AB - Endogenous N-methyl(R)salsolinol, which caused parkinsonism in rats by injection in the striatum, was found to induce apoptosis in dopaminergic neuroblastoma SH SY5Y cells. After 12-h incubation with 500[microM N-methyl(R)salsolinol, almost all the cells died with apoptosis and necrotic cell death was negligible. N Methyl(R)salsolinol was much more potent to induce apoptosis than the (S) enantiomer. The mechanism of apoptosis was studied in relation to changes in mitochondrial membrane potential, deltapsi(m), using a fluorescent indicator, JC 1. Red fluorescence of J-aggregates representing hyperpolarized deltapsi(m) was found to decrease significantly within 60 min after incubation with N methyl(R)salsolinol, but not by the (S)-enantiomer at the same concentration. It suggests that mitochondria may recognize the stereo-chemical structure of N methyl(R) salsolinol. Aliphatic propargylamines, (R)-N-(2-heptyl)-N methylpropargylamine and (R)-N-(2-heptyl)propargylamine, were found to prevent deltapsim loss and subsequent apoptosis induced by N-methyl(R)salsolinol. These results suggest that mitochondria play a key role in the induction of apoptosis by the neurotoxin and the prevention by aliphatic propargylamines. PMID- 11261743 TI - No association between dopamine D2 receptor gene (DRD2) and human intelligence. AB - Significantly diminished intellectual functioning, as indicated by appropriately administered IQ tests with scores below 70, is a frequent mental handicap leading to severe social disadvantages and serves as a paradigm for molecular genetic research of complex disorders and traits due to its multitude of known and unknown, genetic as well as environmental causes. Since the number of confounding variables is expected to be considerably reduced in the normal population at the opposite ends of the IQ distribution, we employed a contrast of extremes approach by comparing adults of high (N = 71) and average IQ (N = 78) in association studies to search for genes involved in the multigenic forms of familial mental retardation. The dopamine D2 receptor gene (DRD2) was chosen as a candidate gene for general cognitive ability (g) since it has been found to be associated with visuospatial ability which in turn is highly correlated with g. Confirming two similar studies in children, however, no significant differences were obtained. Given three negative studies, the DRD2 gene is unlikely to pay a major role in g. PMID- 11261744 TI - Anti-apoptotic and apoptotic action of (-)-deprenyl and its metabolites. AB - The mode of cytoprotective action of the monoamine oxydase B inhibitor (-) deprenyl was studied using A-2058 human melanoma cells in culture. Serum deprivation caused apoptosis of the cultured cells, which could be decreased by administration of 10(-9) - 10(-13)M (-)-deprenyl. The known metabolites of (-) deprenyl, (-)-desmethyl-deprenyl, (-)- and (+)-methylamphetamine failed to exert the same effect. The anti-apoptotic activity of (-)-deprenyl was prevented by the simultaneous application of the microsomal drug-metabolizing enzyme inhibitor SKF 525A. These results show that (-)-deprenyl needs metabolic conversion in order to be anti-apoptotic, but the effective metabolite is still unknown. On the other hand, higher dose (10(-13)M) of (-)-deprenyl, (-)-desmethyl-deprenyl, (-)- and (+)-methylamphetamine induced apoptosis in the non-serum-deprived A-2058 cell culture. SKF-525A did not prevent the apoptosis-inducing effect of (-)-deprenyl, which means that no metabolic changes are needed for this activity. High dose (10(-3)M) of (-)-deprenyl induced very high Caspase 3 activity in non-serum deprived A-2058 cell culture, low doses (10(-9) - 10(-3) M) of (-)-deprenyl maintained Caspase 3 activity on control level in case of serum-deprivation. PMID- 11261745 TI - Deconvolution of transcranial magnetic stimulation (TMS) maps. AB - Transcranial magnetic stimulation (TMS) is a noninvasive method for local stimulation of cerebral cortex using a small coil's pulsed magnetic field. TMS response maps consist of measured responses to stimulations at points on a scalp referenced grid and are used to study the topography of the brain's inhibitory and excitatory response. Because the magnetic field distributions of stimulation coils are 1-2 centimeters wide and 2-3 centimeters long, and the induced electric fields are even broader, the resolution of TMS maps is limited and the actual region of cortical stimulation is poorly defined. To better characterize the activation pattern, a practical mathematical procedure was developed for deconvolving a spherical model approximation of the coil's induced electric field distribution (here measured in a phantom) from the TMS response maps. This procedure offers an integrated, internally consistent method for processing TMS response maps to estimate the spatial distribution of motor cortex activations and inhibitions. PMID- 11261746 TI - Freezing of gait in patients with advanced Parkinson's disease. AB - BACKGROUND: Freezing of Gait (FOG) is one of the most disturbing and least understood symptom in advanced stage of Parkinson's disease (PD). The contribution of the underlying pathological process and the antiparkinsonian treatment to the development of FOG are controversial. OBJECTIVE: To study the relationships between clinical features of PD and therapeutic modalities in patients with advanced PD and FOG. METHODS: Consecutive patients with 5 years or more of PD symptoms (n = 172) (99 men) with mean age at symptoms onset of 58.3 +/ 13.2 years and mean symptoms duration of 11.8 +/- 5.6 years were studied. Clinical data were collected during the last office visit through physical examination, detailed history, review of patients' charts, and other documents. A patient was considered as "freezer" if he/she reported recent experience that the legs got stuck to the ground while trying to walk. The presence of dyskinesia, early morning dystonia or significant postural reflex abnormalities were assessed through history and neurological examination. Duration of treatment with antiparkinsonian drugs was calculated from history charts. Chi square and t test were used to compare the patients with and without FOG. Logistic regression was used for the comparison of association between the presence of FOG (dependent variable) disease duration and disease stage (explanatory variables) and duration of treatment with anti-parkinsonian drugs. RESULTS: The study population consisted of 45 patients at Hoehn and Yahr (H&Y) stage 2.5 (26%), 104 patients at stage 3 (60.5%), and 23 patients at H&Y stages 4-5 (13.5%). Ninety one patients (53%) reported FOG at the time of the study. Severity of the disease expressed by H&Y stage at "off" was a significant contributing factor for FOG with a significant trend (z = 4.38, p < 0.0001), as was longer duration of levodopa treatment, and confirmed by FOG using the multivariate logistic regression (p = 0.01 and p = 0.004, respectively). Using a univariate model, longer duration of treatment with dopamine agonists contribute to the appearance of FOG (p = 0.07) while longer duration of amantadine treatment decreased the appearance of FOG (p = 0.09). There was a significant association between FOG and the presence of dyskinesia (p < 0.002), early morning foot dystonia (p < 0.003) and significant postural instability (p < 0.0005). CONCLUSION: FOG is a common symptom in advanced PD. It is mainly related to disease progression and levodopa treatment. PMID- 11261747 TI - Switching from pergolide to pramipexole in patients with Parkinson's disease. AB - OBJECTIVE/BACKGROUND: To compare the safety and efficacy of pramipexole and pergolide in the treatment of mild to moderate Parkinson's disease (PD). In contrast to pergolide, a D1 and D2 dopamine agonist, pramipexole is a nonergoline dopamine agonist with D2 and preferential D3 dopamine receptor activity. This selective activity may result in clinically different effects. No prospective head-to-head comparison studies of pergolide and pramipexole have been reported. METHODS: Patients with PD who were maintained on an optimal dose of pergolide were converted to pramipexole, typically over a one-month period. Clinical assessments were performed just prior to conversion and after an optimal dose of pramipexole was achieved. RESULTS: Twenty-five patients were converted from pergolide to pramipexole during the period of July, 1997 to January, 1999. Three patients were lost to follow-up, and one patient died. Of the remaining 21 patients there were 11 men and 10 women, mean age was 67.3 years +/- 10.0 (range 51-84). Mean duration of symptoms prior to conversion was 12.5 years +/- 3.4 (range 5-19). All patients (except one) were on concomitant carbidopa/ levodopa and experienced motor fluctuations. After a mean follow-up of 5.9 +/- 2.9 months on pramipexole, the mean levodopa daily dose was reduced from 618.7mg to 581.2mg (16.5% reduction, p = 0.61). The mean daily doses of pergolide and pramipexole (in milligrams per day) were 2.1 +/- 1.5 (0.15-6) and 3.2 +/- 1.1 (0.75-6) respectively. Thirteen patients (62%) reported overall improvement (subjective global response) on pramipexole as compared to pergolide, 5 (24%) were unchanged and 3 (14%) reported worsening. Eighteen of the 21 patients (86%) remained on pramipexole after the study period. Although there was a slight trend toward improved scores on pramipexole, the difference was not statistically significant. CONCLUSION: This open label study failed to provide evidence of superior efficacy of either dopamine agonist. It is possible, however, that while somepatients may benefit more from either pergolide or pramipexole, other patients may obtain additional benefit from other DA agonists or combination therapy. Future randomized, controlled, double-blinded therapeutic trials are needed to determine which, if any, dopamine agonist is superior in the treatment of PD. PMID- 11261748 TI - A study of excessive daytime sleepiness and its clinical significance in three groups of Parkinson's disease patients taking pramipexole, cabergoline and levodopa mono and combination therapy. AB - OBJECTIVE: To determine if therapy with an ergot and a non-ergot dopamine agonist and levodopa confers an increased risk of excessive daytime sleepiness and secondary "sleep attacks" in Parkinson's disease (PD). METHODS: Comparative study of three clinical groups taking, pramipexole (Group 1, n = 19, 8 monotherapy), cabergoline (Group 2, n = 22, 10 monotherapy), and levodopa monotherapy (Group 3, n = 14). Clinical and demographic characteristics, occurrence of "sleep attacks", and assessment of daytime sleepiness [using the Epworth Sleepiness Scale (ESS)], recorded. RESULTS: No patients reported "sleep attacks". Mean ESS scores: Group 1 (pramipexole) 8.0 +/- 4.5 (range 0-16), Group 2 (cabergoline) 8.1 +/- 3.9 (range 0-19), Group 3 (levodopa), 8.1 +/- 5.5 (range 1-18). There was no significant difference between groups (p = 0.897). Scores of > or = 16 indicating excessive daytime sleepiness (EDS) were evenly distributed throughout treatment groups, particularly in older patients with more advanced disease. CONCLUSIONS: a) EDS is not unique to pramipexole therapy and occurs with both cabergoline and levodopa. b) Increasing age, advanced disease, and higher treatment dose appear important predictors for EDS. c) Driving regulations should be reviewed accordingly. PMID- 11261749 TI - Comparative toxicological study on the hepatic safety of entacapone and tolcapone in the rat. AB - Entacapone and tolcapone are novel COMT (catechol-O-methyltransferase) inhibitors indicated for the adjunctive treatment of Parkinson's disease (PD) in combination with levodopa. The marketing authorisation of tolcapone was suspended in the European Union (EU) in 1998 mainly due to severe abnormal hepatic reactions. This fact raised concern about the safety of COMT inhibitors in the treatment of parkinsonian patients. In order to investigate whether these COMT inhibitors exhibit different effects on the liver comparative toxicological studies were performed in the rat. Short term toxicological studies in rats at high oral doses of entacapone and tolcapone (200, 400 or 600mg/kg daily) were carried out. Tolcapone (400 mg/kg/day or 600 mg/kg/day) increased mortality after only one week treatment and induced signs of toxicity such as a rise in body temperature, stimulation of respiration and rapid onset of rigor mortis after death. Entacapone did not show any adverse effects at the tested dose levels. In the histopathological examination liver cell necrosis was observed in the tolcapone (400 and 600mg/kg/day) treated rats, but it revealed no treatment related signs of toxicity in entacapone-treated rats. We conclude that the toxicological profile of the two COMT inhibitors, entacapone and tolcapone, differ from each other, tolcapone--unlike entacapone--showed hepatotoxicity. PMID- 11261750 TI - CSF-folate levels are decreased in late-onset AD patients. AB - Folates are involved in the cerebral metabolism of cobalamine, methionine, L tyrosine and acetylcholine. Remarkably CSF-folate levels are 3 to 4 times higher than blood-folate levels. To reach the brain, folates are actively transported by choroid plexus (CP) as well as vitamins B6, B12, C and E. Epithelial atrophy having been reported in aging and in Alzheimer's disease (AD), we measured the CSF folate-levels of 126 patients, including 30 AD consecutive patients to evaluate whether CP functions of folate-transport were impaired. CSF-folate concentrations did not vary with age (10.47 +/- 1.93ng/ml between 20 and 60 years; 9.96 +/- 2.01 ng/ml in elderly control patients older than 60 years of age, p > 0.05) while late-onset AD patients had significantly lower CSF-folate levels (8.26 +/- 1.82 ng/ml, p < 0.001). These data support a specific alteration of CP transport function in AD patients. PMID- 11261751 TI - Rapid ribosequencing--an effective diagnostic tool for detecting microbial infection. AB - BACKGROUND: Rapid and reliable identification of microorganisms is a prerequisite for the diagnosis and subsequent treatment of infectious diseases. The identification of pathogenic bacteria is traditionally based on their isolation from clinical samples and propagation on culture medium in the routine laboratory. However, despite clinical signs of infection, culture of the pathogenic agent often fails. This may be due to a low number of microorganisms, prior antibiotic treatment, nonculturable microorganisms or specific culture requirements for presently unknown pathogens. Amplification and sequencing of the entire prokaryotic 16S-rRNA is time consuming, labor intensive and expensive. MATERIALS AND METHODS: We describe here a procedure for the identification of a wide range of known and unknown clinically relevant microorganisms by sequencing a small, but highly informative region of the prokaryotic 16S-rRNA gene. This rapid ribosequencing method was evaluated with various reference strains and with clinical samples including eye anterior chamber fluid, cerebrospinal fluid (CSF) and blood cultures. RESULTS: All sequences obtained from the reference strains corresponded to the sequences in databases. We correlated severe eye infection with the isolation of Pseudomonas putida, neurological disorder with Tropheryma whippelii and disseminated visceral abscesses in a child with Blastobacter denitrificans. CONCLUSION: We consider the rapid ribosequencing method as a promising new tool for the analysis of infectious agents in primarily sterile body fluids where conventional culturing of microorganisms fails. PMID- 11261752 TI - Antibodies to the E2 protein of GB virus C/hepatitis G virus: prevalence and risk factors in different populations in Italy. AB - BACKGROUND: Many aspects of the epidemiology of GB virus C/hepatitis G virus (GBV C) infection have not been fully elucidated. The purpose of this study was to assess the prevalence of GBV-C antibodies and risk factors in different subjects living in Italy. MATERIALS AND METHODS: A total of 1,005 sera were tested for the presence of antibodies to the of GBV-C E2 protein using a recently developed enzyme immunoassay. RESULTS: A high prevalence of GBV-C antibodies was found in healthy blood donors (12.6%). Hemodialysis patients and drug users showed higher rates of GBV-C seropositivity (22% and 39%, respectively). Immigrants from sub Saharan Africa and South Asia had anti-GBV-C prevalence comparable to that found in Italian blood donors, whereas higher and lower rates were detected in immigrants from Latin America and the Caribbean (19.5%) and from Mediterranean Africa (5.6%). CONCLUSION: GBV-C infections are widespread in the general population in Italy and particularly common in risk groups. The different prevalence of GBV-C antibodies detected in third world immigrants is likely to reflect the prevalence in the countries of origin. However, the observation that the length of residency in Italy is a significant risk factor may suggest that at least some GBV-C infections are contracted in Italy. PMID- 11261754 TI - Collagen patches impregnated with antimicrobial agents have high local antimicrobial efficacy and achieve effective tissue gluing. AB - BACKGROUND: Local antimicrobial systems have gained importance, as illustrated by current research on drug delivery systems (DDS). We aimed to develop materials that combine hemostatic and antimicrobial efficacy as well as adhesiveness for use in surgical tissue management. MATERIALS AND METHODS: Materials were evaluated by in vitro studies employing microbiological and technological methods. RESULTS: Antimicrobial impregnation of a collagen fleece, which is a pre coated fibrinogen-based adhesive and therefore ready-to-use (TachoComb), is significantly more efficient--both in terms of the antimicrobial efficacy (p < 0.001) as well as the adhesive strength (p = 0.03) -than coating an antibiotic containing collagen fleece "on-site" with fibrin glue. CONCLUSION: Due to ease of practical handling and favorable pharmacoeconomics, this DDS is recommended for both open and minimally invasive surgery. PMID- 11261753 TI - Hepatitis C virus genotyping versus serotyping in Egyptian patients. AB - BACKGROUND: The RNA genome of hepatitis C virus (HCV) displays extensive sequence variation. In this study, serotyping and genotyping techniques were applied to assess this variability by comparing the performance of the serotyping assay with a panel of well-characterized HCV strains isolated from chronic active hepatitis (CAH) patients. PATIENTS AND METHODS: 60 serum samples from CAH patients were analyzed. All isolates were genotyped by a line probe assay and the results of genotyping and serotyping were evaluated. RESULTS: The overall sensitivity of the serotyping and genotyping techniques was 81.16% with a concordance of 73.3%. Type 4 was detected in 73.3% of cases and it was highly heterogeneous. CONCLUSION: Type 4 HCV is the most prevalent type in Egyptian CAH patients and there is a high concordance between the results of serotyping and genotyping techniques. PMID- 11261755 TI - Hospital admissions for pneumonia in Spain. AB - BACKGROUND: Hospital-based surveillance study to estimate the burden of hospitalizations due to pneumonia in Spain. PATIENTS AND METHODS: The national surveillance system for hospital data (Conjunto Minimo de Datos, CMBD) maintained by the Ministry of Health was used to obtain information concerning hospital discharges for pneumonia for the period January 1, 1995 to December 31, 1996. RESULTS: There were 109,644 hospitalizations for pneumonia during this period. For the years 1995 and 1996 the incidence of hospitalizations for pneumonia was 162 and 189 cases per 100,000 population, respectively. Persons > or = 65 years accounted for 49.5% of cases. The average length of stay in hospital was 11.2 days and the case-fatality rate was 7.4%. CONCLUSION: Each year more than 50,000 persons were hospitalized for pneumonia and approximately 4,000 died. PMID- 11261756 TI - Th1/Th2 shift in HIV lymph nodes: no contribution of CD6o cells. AB - BACKGROUND: An expansion of CD8+60+ cells with Th2 type helper function has been observed in HIV-infected individuals. A Th1/Th2 shift in late HIV infection might be related to the functional activity of this subset. Our objective was to test the ability of lymph node (LN) lymphocytes to produce cytokines of the Th1 and Th2 type. PATIENTS AND METHODS: LN cells were stimulated with PMA in the presence of ionomycin and brefeldin A. After surface staining for CD4, CD8 and CD60 and intracellular staining for interferon gamma (IFNgamma), interleukin 2 (IL-2) or IL-4 and IL-10, the percentage of cytokine producing lymphocytes (CPL) was determined by flow cytometry. LN of nine individuals in the early stage of HIV infection were compared to late stage patients. RESULTS: CD4+ subset: in early stages of HIV infection the percentage of Th1 CPL was 1.9 times higher than that of Th2 CPL. In late stages of infection the Th1 responding cells were not found more frequently than Th2 ceLLs. CD8+ subset: no Th1/Th2 shift was detected during early or late stages of HIV infection. CD60+ subset: a maximum of 32.1 +/- 7.8% of double positive cells of the CD8+60+ type produced Th2 type cytokines. A small percentage (< 8%) of CD60+ cells also produced Th1 cytokines. No shift in the cytokine production was seen in early or late stages of infection. CONCLUSION: At single cell level a shift in cytokine production in LN cells can only be detected in the CD4+ subset. Thus, the CD60+ subset does not seem to contribute to the putative Th1/Th2 shift attributed to the immunopathogenesis of HIV-induced destruction of the immune system. PMID- 11261757 TI - Varicella zoster virus infection associated with erythema multiforme in children. AB - BACKGROUND: Erythema multiforme (EM) is a vesiculobullous disorder with variable manifestations which predominantly affects the skin. It is regarded as a hypersensitivity disorder which is triggered by multiple factors such as infection, drugs and food. Varicella zoster virus (VZV) has rarely been reported as an etiological agent, despite its high incidence as a pathogen in childhood. PATIENTS: We describe two children in whom EM preceded VZV infection. In the first, a 5-year-old boy, EM was followed 3 days later by a classical disseminated varicella eruption. The diagnosis was reached by clinical, epidemiological and serological means. The second patient was a 13-year-old boy with EM which was followed 2 weeks later by Ramsay-Hunt syndrome. The diagnosis was confirmed by skin biopsy, positive serology and viral culture. CONCLUSION: The association of EM and VZV infection is probably more common than reported. In clinical cases of EM, VZV should be included in the list of possible causative agents. PMID- 11261758 TI - Acute posttransfusion hepatitis C: identification of a common hepatitis C virus strain in donor and recipient using polymorphism analysis. AB - An 11-year-old Thai boy who had received multiple blood transfusions from 12 different donors for treatment of Dengue shock syndrome presented with symptoms of acute hepatitis 5 weeks thereafter. He was found positive for antibodies to hepatitis C virus (HCV) and HCV-RNA was detected by reverse transcription PCR (RT PCR). When his alanine aminotransferase (ALT) level peaked at 1,879 U/l in the 8th week, interferon therapy (3 million units, thrice weekly for 6 months ) was initiated. After initially decreasing to tenfold the normal level, the ALT dropped to fivefold the normal level at 6 months. HCV RNA is still detectable in his serum 6 months later. Using RT-PCR and subsequent restriction fragment length polymorphism (RFLP) analysis we identified one of the donors as harboring HCV genotype 3a, identical to that found in the patient. Moreover, polymorphism analysis on the hypervariable region employing five distinct restriction endonucleases suggested this donor as the source of infection. We hence recommend thorough screening of all blood donors as the only means of prevention presently feasible. PMID- 11261759 TI - A case of aortic valve disease associated with Tropheryma whippelii infection in the absence of other signs of Whipple's disease. AB - A case of endocarditis caused by Tropheryma whippelii is reported. The 69-year old patient was diagnosed as suffering from severe aortic regurgitation requiring aortic valve replacement, but showed no other symptoms of Whipple's disease. T. whippelii was detected in the explanted aortic valve by broad-range PCR amplification of the 16S rDNA and subsequent sequence analysis of the product. The etiologic agent was classified as a type 2A sequence variant based on the 16S 23S intergenic spacer and the 23S rDNA (domain III) sequences. The histological examination of the aortic valve was compatible with Whipple's disease. A duodenal biopsy revealed an infection with Giardia lamblia, but T. whippelii and histological signs of Whipple's disease were not detectable. PMID- 11261760 TI - Mycobacterium kansasii pericarditis as a presentation of AIDS. AB - Mycobacterium kansasii infection is a recognized complication of AIDS and a broad spectrum of extrapulmonary manifestations has been reported. However, AIDS related M. kansasii pericarditis is an extremely rare disease. We report the first European case of this infection, that presented some different clinical findings to those previously described in HIV-infected individuals. M. kansasii pericarditis was the first AIDS-defining illness presented by the patient. The stained smears of pericardial fluid were negative for acid-fast bacilli and an increased level of adenosine deaminase was observed in pericardial fluid. A short course of prednisone therapy was added to antituberculous treatment, with a good clinical response. PMID- 11261761 TI - Pericarditis after allogeneic peripheral blood stem cell transplantation caused by Legionella pneumophila (non-serogroup 1). AB - A case of Legionella pericarditis caused by a Legionella pneumophila isolate other than serogroup 1 is reported in a 59-year-old man after allogeneic peripheral blood stem cell transplantation. On admission a 5 mm pericardial effusion was detected on echocardiography. Antibodies were detected against L. pneumophila serogroups 7 to 14 using the antigen pool and against serogroup 12 alone. Antibodies were not detected against the serogroup 1 to 6 antigen pool. The patient's clinical condition improved dramatically after treatment with clarithromycin and an echocardiography revealed the total disappearance of the pericardial effusion. PMID- 11261762 TI - Scrub typhus pneumonitis acquired through the respiratory tract in a laboratory worker. AB - We report a case of scrub typhus pneumonitis in a laboratory worker who apparently acquired it through the respiratory tract. The patient was suffering from fever, cough and dyspnea. He had both cervical and axillary lymphadenopathy, and hepatomegaly. A chest X-ray showed interstitial infiltrates. A diagnosis of scrub typhus was established upon isolation of Orientia tsutsugamushi. 12 days before the patient showed symptoms, he had purified O. tsutsugamushi proteins from infected cells using an ultrasonication method which could generate aerosols containing O. tsutsugamushi. PMID- 11261763 TI - Buccal adherence of Pseudomonas aeruginosa in patients with cystic fibrosis under long-term therapy with azithromycin. AB - BACKGROUND: The oropharyngeal barrier is an innate host defence mechanism to prevent bacterial Lung infection. A compromised barrier function is observed in patients with cystic fibrosis (CF) who are chronically infected with Pseudomonas aeruginosa. Macrolides are assumed to modify host defence. We investigated the oropharyngeaL barrier function in CF patients treated with azithromycin (AZM). PATIENTS AND METHODS: In a prospective study, eleven chronically infected children with CF were treated with longterm low-dose AZM. The oropharyngeal barrier function was assessed by adherence of P. aeruginosa (strain PACF 12-1) to buccal epithelial cells of the patients before and after therapy. RESULTS: The mean (standard deviation, SD) buccaL adherence before therapy was markedly high with 8.0 (4.8) bacteria/cell. Following therapy with AZM adherence decreased in all patients by 70% or 5.6 to 2.4 (1.1) bacteria/cell (p = 0.007), representing close to normal LeveLs (1.2 +/- 0.6). CONCLUSION: Long-term low-dose AZM therapy may improve the compromised oropharyngeaL barrier function in patients with CF, opening new perspectives for early treatment of P. aeruginosa infection in CF. PMID- 11261764 TI - Anticonvulsion effect of acupuncture might be related to the decrease of neuronal and inducible nitric oxide synthases. AB - To measure the levels of hippocampal nitric oxide synthase isoforms in penicillin induced epilepsy and to test the effect of electroacupuncture (EA) on changes of theses levels during epilepsy, we injected penicillin into rat hippocampus to make an epilepsy model and performed electroacupuncture treatment on "Feng Fu" (DU 16) and "Jin Suo" (DU 8) points in Wistar rats. Nitric Oxide synthase (NOS) mRNA levels of rat hippocampus were determined by reverse transcription polymerase chain reaction (RT-PCR). The neuronal nitric oxide synthase (nNOS) mRNA markedly increased (P<0.01) and inducible nitric oxide synthase (iNOS) mRNA significantly emerged during epilepsy, whereas no significant change in epithelial nitric oxide synthase (eNOS) mRNA was observed. EA inhibited the epilepsy and decreased nNOS (P<0.01) and iNOS (P<0.01) correspondingly but had no effect on the amount of eNOS mRNA. The data suggest that penicillin-induced epilepsy caused an increase in nNOS and iNOS, and the EA anticonvulsant effect might be related to the decrease of these nitric oxide synthases. PMID- 11261765 TI - The central mechanism of the depressor-bradycardia effect of "Tinggong. AB - Roles of central adrenergic receptors and opioid receptors in the depressor bradycardia effect of 3V, 2Hz "Tinggong-Quchi" electroacupuncture (the EA-DpB, i.e.the depressor bradycardia induced by electroacupuncture) were studied by intracerebroventricula (icv) injection of prazosin, yohimbine or propanol, naloxone or by intra-arachnoid (ith) injection of naloxone. Voltage-dependent depressor effects were induced by 2Hz "Tinggong-Quchi" acupuncture. The depressor effect of 3V, 2Hz " Tinggong-Quchi " acupuncture was attenuated by icv injection of a beta-receptor antagonist-propranolol, but was not blocked by the icv injection of an alpha1 -or (alpha2 -receptor antagonist prazosin or yohimbine. Icv injected naloxone but not ith injected naloxone blocked or reversed the EA DpB. Results suggest that central P-receptors or opioid receptors in the brain are selectively involved in the EA-DpB. PMID- 11261766 TI - Tolosa Hunt Syndrome--intractable pain treatment with acupuncture? AB - PURPOSE: The Tolosa Hunt Syndrome (THS) is a painful granular inflammation of the cerebral vessels followed by pain and disorders of the extrabulbar muscles. The therapy consists of corticosteroids and analgetics. There was a 70 year old woman who suffered from painful paresis of the abducent and oculomotor nerves following an infection with Borrelia Burgdorferi--but without ocular symptoms. The treatment with corticosteroids reduced the palsy but she complained of excessively painful attacks in the region of the first branch of the trigeminal nerve. Opiold analgetic therapy did not bring about any relief. Acupuncture is an irritative method with a physical effect on the nervous system: its pain-reducing effect is caused by the activation of transmitters like endorphins in thalamus and brain stem. Knowing this effect, the THS patient, after informed consent, was treated with acupuncture. To measure the extent of pain, a visual analog scale (0: no pain - 10: maximum pain) was used. Acupuncture was performed according to the empirical rules of the Traditional Chinese Medicine (TCM), during a period of 10 weeks and 12 weeks. There was a significant pain relief after acupuncture from VAS 10 to VAS 5. The effect vanished during the next four months. After a second series of 12 sessions pain reduction was reported from VAS 10 to 4. One year after the last Tolosa Hunt Syndrome - intractable pain pain strength ranged between VAS 4 - 6. Therefore acupuncture seems to be a good additional method for reduction of intractable pain. PMID- 11261767 TI - Preliminary results of a new method for locating auricular acupuncture points. AB - Auricular acupuncture is widely used for the treatment of cocaine addiction, and there is an urgent need to conduct controlled clinical research of this intervention. One impediment to this endeavor is the lack of an objective and reliable method for identifying the hypothesized active and control points. In order to address this issue, we conducted two studies employing a constant current electrical device and a novel probing technique. In the first study, we assessed the reliability of our technique for measuring electrical skin resistance points (acupuncture or non-acupuncture) on the body and auricles. In the second study, we analyzed and compared the measurements of skin resistance of auricular acupuncture and control zones in a group of cocaine abusing patients. Findings suggest that our measurement method produced reliable measurements, and that active acupuncture zones revealed a significantly different pattern of electrical skin resistance readings compared to control zones. This method may be useful for locating active and control points in controlled clinical trials of auricular acupuncture. PMID- 11261768 TI - Consolidation and the medial temporal lobe revisited: methodological considerations. AB - It is widely believed that new memories are stored in the medial temporal lobe structures in the short term, but then are reorganized over time as the neocortex gradually comes to support stable long-term storage. On this view, the medial temporal lobe structures play a time-limited role in information storage. This putative process of reorganization, known as consolidation, is supported by some clinical findings in humans and by some data from nonhuman animals. Here we review prospective studies of retrograde memory in nonhuman animals, with particular emphasis on experimental design. In considering the evidence for a time-limited role for the medial temporal lobe in information storage, we note that there are alternative interpretations for at least some of the findings typically cited in support of the consolidation process. In addition, we suggest that some studies arguing against the consolidation view should probably be given more weight than they have so far received. Finally, we observe that different structures in the medial temporal lobe are unlikely to operate together as a single functional unit mediating a single consolidation process. Although evidence for a time-limited role for medial temporal lobe structures in memory is at present equivocal, future studies that consider some of the alternative accounts we and others have identified will provide a clearer picture of the mechanisms underlying information storage and retrieval in the brain. PMID- 11261769 TI - Anterograde and retrograde amnesia in rats with large hippocampal lesions. AB - A test of socially acquired food preferences was used to study the effects of large lesions to the hippocampal formation (HPC) on anterograde and retrograde memory in rats. In the anterograde test, rats with HPC lesions normally acquired the food preference but showed a faster rate of forgetting than control groups. When the food preference was acquired preoperatively, HPC groups exhibited a temporally graded retrograde amnesia in which memory was impaired when the preference was acquired within 2 days of surgery but not at longer delays. The results support the traditional theory that the HPC contributes to the consolidation of newly acquired information into a durable memory trace that is represented in other brain areas. Consistent with this view, the results indicate that, once a memory trace is consolidated, the HPC does not participate in its storage or retrieval. The possibility is considered that extrahippocampal areas in the medial temporal lobe are needed to maintain a memory trace throughout its existence. PMID- 11261771 TI - Retrograde amnesia and consolidation: anatomical and lesion considerations. AB - The four papers in this issue of Hippocampus dealing with retrograde amnesia, together with relevant animal studies in the literature, are reviewed from the perspective of the anatomical location of the lesion and extent of damage to the brain. In order to evaluate the underlying damage in these and related prospective experimental studies, it is necessary to consider both the lesion techniques that were used as well as the care with which the resulting damage was determined. Both temporally graded and flat, ungraded retrograde amnesia have been reported, as well a lack of effects, following damage to structures in the medial temporal area. Most research has centered around damage to the hippocampus, but differences in selectivity of the lesions and behavioral testing procedures preclude any definite conclusions regarding the precise nature of the involvement of this structure. With a greater appreciation for the importance of the locus and extent of the damage, together with the kind of information being processed, it should be possible to obtain a better understanding of the neural substrates underlying retrograde amnesia. PMID- 11261770 TI - Retrograde amnesia after hippocampal damage: recent vs. remote memories in two tasks. AB - We review evidence from experiments conducted in our laboratory on retrograde amnesia in rats with damage to the hippocampal formation. In a new experiment reported here, we show that N-methyl-D-aspartate (NMDA)-induced hippocampal damage produced retrograde amnesia for both hidden platform and two-choice visible platform discriminations in the Morris water task. For both problems there was a significant trend for longer training-surgery intervals to be associated with worse retention performance. Little support is offered by our work for the concept that there is a process involving hippocampal-dependent consolidation of memories in extrahippocampal permanent storage sites. Long-term memory consolidation may take place within the hippocampus. The hippocampus may be involved permanently in storage and/or retrieval of a variety of relational and nonrelational memories if it was intact at the time of learning, even involving information which is definitely not affected in anterograde amnesia after hippocampal damage. PMID- 11261772 TI - Retrograde amnesia. AB - In humans, the phenomenon of temporally graded retrograde amnesia has been described in the clinic and the laboratory for more than 100 years. In the 1990s, retrograde amnesia began to be studied prospectively in experimental animals. We identified 13 published studies in which animals were given equivalent training at two or more separate times before damage to the fornix or hippocampal formation. Eleven of these studies found temporally graded retrograde amnesia, with the extent of amnesia ranging from several days to a month or two. We consider these studies and also suggest why temporally graded retrograde amnesia has sometimes not been observed. Although the evidence in favor of temporally graded retrograde amnesia is substantial, the inference from this work, that memory is reorganized as time passes, is rather vague and depends on mechanisms yet to be identified. It is therefore encouraging that many opportunities exist for moving beyond purely descriptive studies to studies that involve treatments or manipulations directed toward yielding information about mechanisms. PMID- 11261773 TI - Consolidation of memory. AB - Animal studies have proven useful in addressing aspects of memory formation and consolidation that cannot be readily answered in research with humans. In particular, they offer the possibility of controlling both the extent and locus of brain lesions, and the exact nature of the experiences to be remembered. Taking advantage of these possibilities, recent studies indicated that the graded retrograde amnesia often seen after lesions to the hippocampal system is not uniform across lesion site and task, nor is it an indication that all of the remembered information available in intact subjects becomes available after hippocampal system lesions made a long time after learning. Rather, these studies support the notion that information is stored in both hippocampal and extrahipocampal sites, and that retrieval from different sites involves access to different kinds of information. The strongest evidence in support of this view is the set of findings indicating that when remote memories are retrieved, in either human or animal subjects that have suffered hippocampal system damage, these memories are not qualitatively the same as remote memories retrieved in intact subjects. In sum, memory appears to be rather more dynamic than most current conceptions allow, such that retrieval events trigger new encodings, and these new encodings engage the hippocampal system once again. As a result, older, reactivated memories become more resistant to disruption, and this mechanism helps to explain why graded retrograde amnesia is sometimes seen after brain damage. The use of new neuroimaging techniques, coupled with more sensitive neuropsychological tests in lesioned subjects, should further illuminate the complex nature of memory in coming years. It is likely that animal studies will continue to prove important in these developments. PMID- 11261774 TI - Opposite relationship of hippocampal and rhinal cortex damage to delayed nonmatching-to-sample deficits in monkeys. AB - Three recent studies in macaque monkeys that examined the effects on memory of restricted hippocampal lesions (Murray and Mishkin, J Neurosci 1998;18:6568-6582; Beason-Held et al., Hippocampus 1999;9:562-574; Zola et al., J Neurosci 2000;20:451-463) differed in their conclusions about the involvement of the hippocampus in recognition memory. Because these experiments used a common behavioral procedure, trial-unique visual delayed nonmatching-to-sample (DNMS), a quantitative synthesis ("meta-analysis") was performed to determine whether hippocampal lesions produced a reliable net impairment in DNMS performance, and whether this impairment was related to the magnitude of hippocampal damage. A similar analysis was performed on data from monkeys with perirhinal or rhinal cortex damage (Meunier et al., J Neurosci 1993;13:5418-5432; Buffalo et al., Learn Mem 1999;6:572-599). DNMS performance scores were transformed to d' values to permit comparisons across studies, and a loss in d' score, a measure of the magnitude of the recognition deficit relative to the control group, was calculated for each operated monkey. Two main findings emerged. First, the loss in d' following hippocampal damage was reliably larger than zero, but was smaller than that found after lesions limited to the perirhinal cortex. Second, the correlation of loss in d' with extent of hippocampal damage was large and negative, indicating that greater impairments were associated with smaller hippocampal lesions. This relationship was opposite to that between loss in d' and rhinal cortex damage, for which larger lesions were associated with greater impairment. These findings indicate that damage to the hippocampus and to the rhinal cortex affects recognition memory in different ways. Furthermore, they provide a framework for understanding the seemingly disparate effects of hippocampal damage on recognition memory in monkeys, and by extension, for interpreting the conflicting reports on the effects of such damage on recognition memory abilities in amnesic humans. PMID- 11261775 TI - Hippocampus and contextual fear conditioning: recent controversies and advances. AB - Dorsal hippocampal (DH) lesions produce a severe deficit in recently, but not remotely, acquired contextual fear without impairing memory of discrete training stimuli, i.e., DH lesions produce an anterograde and time-limited retrograde amnesia specific to contextual memory. These data are consistent with the standard model which posits temporary involvement of the hippocampus in recent memory maintenance. However, three recent controversies apparently weaken the case for a selective mnemonic role for the hippocampus in contextual fear. First, although retrograde amnesia (from posttraining lesions) is severe, anterograde amnesia (from pretraining lesions) may be mild or nonexistent. Second, a performance, rather than mnemonic, account of contextual freezing deficits in hippocampal-lesioned animals has been offered. Third, damage to the entire hippocampus, including the ventral hippocampus, can produce a dramatic and temporally stable disruption of context and tone fear. These data are reviewed and explanations are offered as to why they do not necessarily challenge the standard model of hippocampal memory function in contextual fear. Finally, a more complete description of the hippocampus' proposed role in contextual fear is offered, along with new data supporting this view. In summary, the data support a specific mnemonic role for the DH in the acquisition and consolidation of contextual representations. PMID- 11261776 TI - Regulation of hematopoiesis through adhesion receptors. AB - Normal steady-state hematopoiesis takes place in the bone marrow microenvironment. Soluble factors as well as contact interactions between the hematopoietic cells and the marrow microenvironment dictate the fate of hematopoietic stem cells and progenitors. Over the last decade it has become clear that cell-cell and cell-extracellular matrix interactions through adhesion receptors play a major role in the hematopoietic process. They are required for the residence of stem cells and progenitors in the marrow, as well as for homing of stem and progenitor cells to the marrow in the setting of stem cell transplantation. Furthermore, adhesion receptors play an important role in regulation of cell behavior, either through direct activation of signal pathways important for cell survival, cell growth, and cell fate decision-making processes, or by modulating responses to growth factors. Insights in the abnormalities seen in these interactions in diseases of the hematopoietic system will help to develop better therapeutic strategies based on the pathogenesis of these diseases. PMID- 11261777 TI - Molecular motors involved in T cell receptor clusterings. AB - Engagement of antigen receptors on T and B cells triggers reorganization of the cytoskeleton and ordered clustering of cell surface receptors. These receptor clusters constitute spatially organized signaling machines and form the immune synapse with antigen-presenting cells. Formation of supramolecular activation clusters appear to be essential to induce functional lymphocyte responses and have been implicated as molecular mechanisms of costimulation. The Vav1-Rho GTPase-WASP pathway constitutes a molecular motor that relays antigen receptor stimulation to changes in the cytoskeleton and receptor clustering. PMID- 11261778 TI - Lymphoid neogenesis: de novo formation of lymphoid tissue in chronic inflammation through expression of homing chemokines. AB - Chronic inflammation is a complex pathophysiological process with accumulation of mononuclear cells seen in response to invading pathogens, neoplastic transformation, or autoimmune recognition of self-antigens. The inflammatory process has evolved to facilitate effective elimination of pathogens and tumors and it is normally transient and turned off when the causative stimulus has been eliminated. Occasionally, however, the process is sustained for a long time and can lead to severe tissue damage. This is seen in organ-specific autoimmune diseases such as rheumatoid arthritis, Sjogren's syndrome, and Hashimoto's thyroiditis, but also in infectious diseases such as Helicobacter pylori-induced gastritis. Disturbingly, many of these chronic inflammatory diseases are associated with an increased risk for neoplastic transformation and development of lymphomas. This review summarizes experimental evidence suggesting that chronic inflammation involves ectopic de novo formation of organized lymphoid tissue and that this lymphoid neogenesis is regulated by expression of homing chemokines. PMID- 11261779 TI - DNA damage and apoptosis in mononuclear cells from glucose-6-phosphate dehydrogenase-deficient patients (G6PD Aachen variant) after UV irradiation. AB - Patients affected with X chromosome-linked, hereditary glucose-6-phosphate dehydrogenase (G6PD) deficiency suffer from life-threatening hemolytic crises after intake of certain drugs or foods. G6PD deficiency is associated with low levels of reduced glutathione. We analyzed mononuclear white blood cells (MNC) of three males suffering from the German G6PD Aachen variant, four heterozygote females of this family, one G6PD-deficient male from another family coming from Iran, and six healthy male volunteers with respect to their DNA damage in two different genes (G6PD and T-cell receptor-delta) and their propensity to enter apoptosis after UV illumination (0.08-5.28 J/cm2). As determined by PCR stop assays, there was more UV-induced DNA damage in MNC of G6PD-deficient male patients than in those of healthy subjects. MNC of G6PD-deficient patients showed a higher rate of apoptosis after UV irradiation than MNC of healthy donors. MNC of heterozygote females showed intermediate rates of DNA damage and apoptosis. It is concluded that increased DNA damage may be a result of deficient detoxification of reactive oxygen species by glutathione and may ultimately account for the higher rate of apoptosis in G6PD-deficient MNC. PMID- 11261780 TI - Importance of CD44v7 isoforms for homing and seeding of hematopoietic progenitor cells. AB - The adhesion molecule CD44 consists of many isoforms of which particularly CD44v7 is of major importance in hematopoietic progenitor cell homing. An increase of progenitor cells in the periphery was observed after treating mice with a CD44v7 specific antibody, concomitant with a substantially augmented marrow-repopulating ability (MRA). Because CD44v7 is expressed on a fraction of bone marrow cells (BMC) as well as on long-term bone marrow culture-derived stromal cells, we aimed to differentiate between the functional relevance of CD44v7 on either cell type by transferring CD44v7+/+ BMC into CD44v7-/- mice and vice versa. CD44v7+/+ BMC homed poorly in the bone marrow of CD44v7-/- mice and their MRA was severely impaired. CD44v7-/- BMC, instead, exhibited an improved MRA when transferred into the CD44v7+/+ host, although their MRA remained below that of CD44v7+/+ BMC. Thus, it is predominantly, but not exclusively, expression of CD44v7 on stromal cells which supports progenitor cell homing. PMID- 11261781 TI - Eosinophil recruitment into sites of delayed-type hypersensitivity reactions in mice. AB - The selective accumulation of eosinophils in tissue is a characteristic feature of allergic diseases where there is a predominance of lymphocytes expressing a Th2 phenotype. In an attempt to define factors determining specific eosinophil accumulation in vivo, we have used a radiolabeled technique to assess the occurrence and the mechanisms underlying (111)In-eosinophil recruitment into Th1- and Th2-predominant, delayed-type hypersensitivity (DTH) reactions. Eosinophils were purified from the blood of IL-5 transgenic mice, labeled with (111)In and injected into nontransgenic CBA/Ca mice. Th1- and Th2-predominant, DTH reactions were induced in mice by immunization with methylated bovine serum albumin (MBSA) in Freund's complete adjuvant or with Schistosoma mansoni eggs, respectively. In these animals, (111)In-eosinophils were recruited in skin sites in an antigen-, time-, and concentration-dependent manner. Depletion of CD4+ lymphocytes abrogated (111)In-eosinophil recruitment in both reactions. Pretreatment of animals with anti-IFN-gamma mAb abrogated (111)In-eosinophil recruitment in MBSA immunized and -challenged animals, whereas anti-IL-4 inhibited (111)In-eosinophil recruitment in both models. Local pretreatment with an anti-eotaxin polyclonal antibody inhibited the MBSA and SEA reactions by 51% and 39%, respectively. These results demonstrate that, although eosinophilia is not a feature of Th1 predominant, DTH reactions, these reactions produce the necessary chemoattractants and express the necessary cell adhesion molecules for eosinophil migration. The control of the circulating levels of eosinophils appears to be a most important strategy in determining tissue eosinophilia. PMID- 11261782 TI - Retinoic acid up-regulates myeloid ICAM-3 expression and function in a cell specific fashion--evidence for retinoid signaling pathways in the mast cell lineage. AB - Investigation of mast cell responsiveness toward retinoic acid (RA) revealed selective promotion of ICAM-3 expression in the human mast cell line HMC-1. This process was dose- and time-dependent and detectable by flow cytometry, Western blot analysis, ELISA, and Northern blot analysis. ICAM-3 modulation was found to be cell-type dependent, detectable also for HL-60 cells and monocytes but not U 937 and only weakly for KU812 cells. Terminally differentiated skin mast cells also failed to up-modulate their ICAM-3, suggesting the requirement for some degree of immaturity for the process. RA-mediated effects on ICAM-1 expression, studied in parallel, were clearly distinct from those on ICAM-3. Investigation of retinoid receptor expression, known to mediate intracellular RA signaling, revealed presence of RAR alpha, RAR gamma, RXR beta, and RXR gamma transcripts in all cell lines studied, and HMC-1 cells were the only line lacking RXR alpha. RAR beta, not expressed at baseline, was induced by RA in a fashion obviously correlating with ICAM-3 up-regulation. Increased ICAM-3 expression was of functional significance, such that processes stimulated or co-stimulated via ICAM 3 (homotypic aggregation, IL-8 secretion) were clearly enhanced upon RA pretreatment, suggesting that RA may contribute via hitherto unrecognized pathways to immune function and host defense. PMID- 11261783 TI - Homotypic cluster formation of dendritic cells, a close correlate of their state of maturation. Defects in the biobreeding diabetes-prone rat. AB - Aggregation of dendritic cells (DCs) in homotypic clusters has been described in vivo in lymph and skin, and here we report studies on homotypic clustering of rat splenic (s) DCs in vitro. Wistar rat sDCs readily formed homotypic clusters in culture, which increased in number and size over time (with a peak at t = 3 h). Keeping the cells at higher densities or treatment with anti-CD43 induced more and larger homotypic clusters. After such enhanced clustering the DCs had increased their T cell stimulating capabilities in syngeneic mixed lymphocyte reaction, and had a higher expression of CD80 and CD86 (signs of maturation). Ag transfer from bovine serum albumin-fluorescein isothiocyanate-pulsed to unpulsed DCs was observed during clustering. Here we also show that sDCs of the biobreeding diabetes-prone (BB-DP) rat, a model of autoimmune diabetes/thyroiditis, formed fewer and smaller clusters than Wistar sDCs, and that DC-DC clustering resulted in only a modest maturation of the cells (as determined in syn MLR and by phenotyping). Anti-CD43 completely restored the clustering defect BB-DP DCs in vitro, yet T cell-stimulating capability was only restored to a limited extent. Ag transfer in BB-DP DC clusters was similar. PMID- 11261784 TI - MHCII, Tlr4 and Nramp1 genes control host pulmonary resistance against the opportunistic bacterium Pasteurella pneumotropica. AB - MHCII, Tlr4, and Nramp1 genes are each independently important in pulmonary immunity. To determine the effect of these genes on host resistance, mice carrying various combinations of functional alleles for these three genes were experimentally challenged with the opportunistic bacterium, Pasteurella pneumotropica. MHCII-/-, Tlr4d/d, and Nramp1s/s mice were significantly more susceptible to experimental infections by P. pneumotropica after intranasal challenge compared to mice carrying functional alleles at only one of those genes. P. pneumotropica were cultured from the lungs of challenged mice, and the severity of the pneumonia strongly correlated with the number of isolated bacteria. Mice with the genotype MHCII-/- Tlr4n/n genotype were less susceptible to pneumonia than MHCII+/+, Tlr4d/d mice. It is interesting that the Nramp1 gene contribution to host resistance was apparent only in the absence of functional MHCII or Tlr4 genes. These data suggest that MHCII, Tlr4, and Nramp1 genes are important to pulmonary bacterial resistance. PMID- 11261785 TI - Alternative versus classical macrophage activation during experimental African trypanosomosis. AB - The type I/type II cytokine balance may influence the development of different subsets of suppressive macrophages, i.e., classically activated macrophages (caMphi, type I) versus alternatively activated macrophages (aaMphi, type II). Recently, we showed that although mice infected with phospholipase C-deficient (PLC-/-) Trypanosoma brucei brucei exhibit a clear shift from type I to the type II cytokine production, wild type (WT)-infected mice remain locked in a type I cytokine response. In the present study, phenotype and accessory cell function of macrophages elicited during WT and PLC-/- T. b. brucei infection were compared. Results indicate that caMphi develop in a type I cytokine environment in the early phase of WT and PLC-/- trypanosome infection, correlating with inhibition of T cell activation triggered by a mitogen, a superantigen, or an antigen. In the late stage of infection, only PLC(-/-)-infected mice resisting the infection develop type II cytokine-associated aaMphi correlating with impaired antigen- but not mitogen- or superantigen-induced T cell activation. PMID- 11261786 TI - Phagocytosis and killing of Mycobacterium avium complex by human neutrophils. AB - Organisms belonging to the Mycobacterium avium complex (MAC) cause life threatening bacteremia in immunocompromised patients. Monocytes and macrophages are thought to be responsible for ingestion and killing of MAC. However, it has been suggested that neutrophils may play a role in the early immune response to MAC infection. Here, neutrophils in autologous plasma were incubated (at 0 and 37 degrees C) with M. avium labeled with Auramine O, a potent fluorochrome. Neutrophil phagocytosis was measured by flow cytometry. Neutrophils incubated at 37 degrees C showed an increase in fluorescence over time with a maximum at 15 min, whereas neutrophils on ice showed no time-dependent increase in FL1. At 15 min Fl 1 at 37 degrees C was twice as high as FL1 at 0 degrees C. Examination under the fluorescent microscope showed multiple intracellular fluorescent mycobacteria. Results in nine independent experiments showed time-dependent decrease of colony-forming units in neutrophil-associated live M. avium. Significant killing was observed within 30 min and was complete by 120 min. Observation by electron microscopy clearly confirmed the presence of intraphagosomal MAC, both intact and with evidence of degradation. These data demonstrate that MAC is rapidly phagocytized and killed by human neutrophils. The newly established flow cytometry method should be useful in further studies of neutrophil function and of the role of G-CSF and other cytokines in MAC disease. PMID- 11261787 TI - CD86 expression correlates with amounts of HIV produced by macrophages in vitro. AB - Primary macrophages from different donors produce variable levels of HIV; however, the mechanisms are unclear. We tested whether variations in cell-surface or cell-cycle characteristics influenced HIV production. We found that greater basal proliferation of the macrophages prior to infection resulted in more arrested in G2M 3 days post-infection (r2=0.7, P<0.04). Likewise, the number of G2M-arrested macrophages correlated with p24 production (r2=0.78, P<0.02) and apoptosis (r2=0.67, P<0.05) later in the infection. Serum-starvation or reduction, which limit HIV spread, reduced G2M arrest and HIV amounts. Surprisingly, the amount of HIV produced correlated with expression levels of the costimulating ligand, CD86, but not with other important molecules, including class II, CD40, or CD54 (r2=0.96, P<0.0005). These data establish donor characteristics related to variable HIV production in vitro and suggest that altered expression of costimulatory ligands may influence HIV production in vivo. PMID- 11261788 TI - Leptin: a potential regulator of polymorphonuclear neutrophil bactericidal action? AB - It is well known that leptin, the ob gene product, is involved in the regulation of food intake and thermogenesis. Recent studies also demonstrate that leptin may be able to modulate functions of cells involved in nonspecific immune response such as phagocytosis and secretion of cytokines by macrophages. This and the prominent implication of polymorphonuclear neutrophils (PMNs) in infectious response suggested a possible role of leptin as a modulator of PMN functions. We detected a leptin receptor on the PMN membrane by immunocytochemistry with an anti-leptin receptor. Using chemiluminescence we then demonstrated that leptin enhances oxidative species production by stimulated PMNs. These results show for the first time that a functional leptin receptor is present on PMNs and that leptin may be able to influence their oxidative capacity. PMID- 11261789 TI - A p74 common gamma receptor chain isoform facilitates IL-2 and IL-15 responses by the myelomonocytic cell line Tf-1beta2. AB - Functional forms of the IL-2, IL-4, IL-7, IL-9, and IL-15 receptors require the gamma c receptor component. We have described previously a myeloid cell line called Tf-1beta, which binds IL-2 with intermediate-affinity and proliferates in response to IL-2. In this study, we characterize gamma c expression on Tf-1beta2 cells, a derivative of Tf-1beta cells stimulated exclusively with IL-2. Although Tf-1beta2 cells bind IL-2 with intermediate-affinity and proliferate in response to IL-2, this cell line does not express the p64 gamma c chain at the protein level. This result was surprising because prior studies suggest these cells should not be expected to proliferate in response to IL-2 or IL-15 in the absence of the p64 gamma c chain. A p74 protein was detected by western blot following immunoprecipitation with an anti-gamma c polyclonal antibody, and a p74 protein was identified consistently in complex with IL-2 and IL-15 on these cells. However, the gamma c gene in these Tf-1beta2 cells shows no evidence of mutation by sequence analysis. Furthermore, inhibition of glycosylation of these Tf-1beta2 cells by tunicamycin treatment yields a standard 39-kDa molecule recognized on western blot with anti-gamma c antibody, as seen for the standard 64-kDa isoform of gamma c. These results demonstrate that a 74-kDa gamma c receptor isoform was involved in the response of the Tf-1beta2 cells to cytokines which normally interact with the 64-kDa gamma c chain. PMID- 11261790 TI - Inhibition of glucocorticoid-mediated, caspase-independent dendritic cell death by CD40 activation. AB - Glucocorticoids (GC) are potent anti-inflammatory and immunosuppressive agents that act on a variety of immune cells, including T cells, monocytes/macrophages, osteoclasts, and dendritic cells (DC). However, the mechanism(s) by which GC exert anti-inflammatory effects is still largely unknown. It is already well known that GC treatment inhibits DC maturation and interleukin (IL)-12 production by DC. In this study, we investigated the apoptosis induction of DC by a synthetic GC, dexamethasone (Dex). The stimulation with Dex resulted in DC apoptosis in a dose- and time-dependent manner as it was measured by determining annexin V-positive cells and mitochondrial potential. In contrast, monocytes that are precursor cells of DC are resistant to Dex-mediated apoptosis. The Dex induced apoptosis of DC was independent of caspase activation because it was not inhibited by the broad caspase inhibitor, Z-VAD-fmk. It is interesting that agonistic CD40 antibody completely inhibited Dex-induced cell death, whereas other inflammatory stimuli did not show the same effect, suggesting that CD40 signaling may selectively modulate GC-mediated DC apoptosis. Taken together, our findings revealed an important role of GC and CD40 signaling in the regulation of immune responses in which DC play a key role in the inflammatory process of various immunomediated diseases. PMID- 11261791 TI - Lymphocytes induce monocyte chemoattractant protein-1 production by renal cells after Fc gamma receptor cross-linking: role of IL-1beta. AB - Leukocyte recruitment to the kidney in immune complex disease like systemic lupus erythematosus (SLE) is mediated in part by local expression of chemokines such as monocyte chemoattractant protein-1 (MCP-1). Recent studies from this laboratory demonstrated that cross-linking Fc gammaR on lymphocytes causes release of a soluble factor that induces monocyte chemokine production. To explain the induction of renal chemokine expression in immune complex disease, we postulated that this lymphocyte factor stimulates renal parenchymal cell MCP-1 expression. To test this hypothesis, human peripheral blood lymphocytes were incubated on immobilized IgG, a model for immune complex Fc gammaR cross-linking. Supernatants from these lymphocyte cultures significantly increased MCP-1 production by human mesangial, glomerular capillary endothelial, and proximal tubular epithelial cells. Mesangial cells incubated on immobilized IgG or with soluble, preformed immune complexes did not secrete MCP-1 above control levels. Lymphocyte supernatant-induced MCP-1 production appeared to be dependent on the presence of interleukin (IL)-1beta in the supernatant. Removing IL-1beta from the supernatants, antagonizing its activity, or preventing conversion to mature IL 1beta abrogated renal cell MCP-1 expression by the lymphocyte supernatants. These data demonstrate that in response to cross-linking Fc gammaR, lymphocytes induce renal cell MCP-1 expression by secreting IL-1beta. Renal chemokine expression in immune complex disease may thus be triggered as lymphocytes traffic through the kidney and encounter deposited immune complexes. PMID- 11261792 TI - SJL and NOD macrophages are uniquely characterized by genetically programmed, elevated expression of the IL-12(p40) gene, suggesting a conserved pathway for the induction of organ-specific autoimmunity. AB - Genetic susceptibility of the SJL mouse to experimental autoimmune encephalomyelitis (EAE) appears, in part, to be a result of genes that promote abnormal development of the pathogenic Type 1 (Th1) phenotype of neuroantigen specific T-cells. Because antigen-presenting/accessory cells (APCs) produce cytokines that can modulate the development of Th1 and Th2 phenotypes, we addressed whether APCs from SJL mice were genetically programmed for elevated expression of the Th1-promoting cytokine, IL-12. Activated peritoneal macrophages (Mphi; i.e., APC) from naive SJL mice produced levels of TNF-alpha, IL-1, IL-6, IL-10, and TGF-beta within the range of six normal strains. In contrast, SJL IL 12p40 (in addition to IL-12p70) production was consistently five- to 20-fold greater than that of any normal strain tested, which arose from elevated expression of the IL-12p40 but not the IL-12p35 gene, because p40 mRNA levels were eight- to 15-fold greater than those of normal strains. This aberrancy in IL 12p40 expression appears identical to that observed in the NOD mouse, another strain prone to organ-specific autoimmunity. A genetically programmed bias toward elevated expression of IL-12 in Mphi from the SJL and NOD strains of autoimmunity provides a conserved mechanism for the dominant Th1 development of naive, autoantigen-specific T-cells in these strains. This study is the first demonstration of a genetically programmed aberrant phenotype that is intrinsically expressed within a cell type in the SJL mouse and provides insight into its predisposition for EAE. PMID- 11261793 TI - Dietary fatty acids influence the production of Th1- but not Th2-type cytokines. AB - C57B16 mice were fed for 6 weeks on a low-fat diet or on high-fat diets containing coconut oil (rich in saturated fatty acids), safflower oil [rich in n 6 polyunsaturated fatty acids (PUFAs)], or fish oil (rich in n-3 PUFAs) as the main fat sources. The fatty acid composition of the spleen lymphocytes was influenced by that of the diet fed. Thymidine incorporation into concanavalin A stimulated spleen lymphocytes and interleukin (IL)-2 production were highest after feeding the coconut oil diet. Interferon (IFN)-gamma production was decreased by safflower oil or fish oil feeding. IL-4 production was not significantly affected by diet, although production was lowest by lymphocytes from fish oil-fed mice. The ratio of production of Th1- to Th2-type cytokines (determined as the IFN-gamma/IL-4 ratio) was lower for lymphocytes from mice fed the safflower oil or fish oil diets. After 4 h of culture, IL-2 mRNA levels were higher in cells from mice fed coconut oil, and IFN-gamma mRNA levels were higher in cells from mice fed coconut oil or safflower oil. After 8 h of culture, IL-2, IFN-gamma, and IL-4 mRNA levels were lowest in cells from mice fed fish oil. The ratio of the relative levels of IFN-gamma mRNA to IL-4 mRNA was highest in cells from mice fed coconut oil and was lowest in cells of mice fed fish oil. The influence of individual fatty acids on IL-2 production by murine spleen lymphocytes was examined in vitro. Although all fatty acids decreased IL-2 production in a concentration-dependent manner, saturated fatty acids were the least potent and n-3 PUFAs the most potent inhibitors, with n-6 PUFAs falling in between in terms of potency. It is concluded that saturated fatty acids have minimal effects on cytokine production. In contrast, PUFAs act to inhibit production of Th1-type cytokines with little effect on Th2-type cytokines; n-3 PUFAs are particularly potent. The effects of fatty acids on cytokine production appear to be exerted at the level of gene expression. PMID- 11261794 TI - Chemokines stimulate human T lymphocyte transendothelial migration to utilize VLA 4 in addition to LFA-1. AB - Lymphocyte infiltration in inflammation is induced by the dual actions of chemokines and cell adhesion molecules. The role of LFA-1 and VLA-4 in chemokine induced T cell transendothelial migration (TEM) across cytokine-activated endothelium has not been examined. LFA-1, but not VLA-4, mediated blood T cell TEM to RANTES, macrophage inflammatory protein-1alpha (MIP-1alpha), and stromal cell-derived factor-1 (SDF-1), and across tumor necrosis factor alpha (TNF-alpha) or interferon-gamma (IFN-gamma) -stimulated endothelial cells (EC). Chemokine stimulation in combination with TNF-alpha activation of EC induced TEM, which was partially mediated by VLA-4. SDF-1 increased a beta1-integrin activation epitope on T cells and enhanced VLA-4-mediated adhesion. Thus, LFA-1 mediates TEM under most conditions, but VLA-4 can also mediate TEM, although, in contrast to LFA-1, this requires exogenous chemokines and EC activation. In addition, an LFA-1- and VLA-4-independent pathway of lymphocyte TEM can also be induced by SDF-1. PMID- 11261795 TI - Divergent effects of tumor necrosis factor alpha on apoptosis of human neutrophils. AB - Apoptosis of neutrophils is a key mechanism to control the intensity of the acute inflammatory response. Previously, the cytokine tumor necrosis factor alpha (TNF alpha) was reported by some to have pro-apoptotic and by others to have antiapoptotic effects on neutrophils. The aim of this study was to explain these contradictory results. We found that TNF-alpha at low concentrations strongly decreased apoptosis of neutrophils. However, at higher concentrations, TNF-alpha lost its protective effects, and also reversed the protective effects of interferon-gamma (IFN-gamma) and granulocyte-macrophage colony-stimulating factor (GM-CSF). This pro-apoptotic effect of TNF-alpha was blocked by anti-CD11b and was absent in neutrophils from patients with chronic granulomatous disease, which cannot produce toxic oxygen metabolites. Under these circumstances, we found that TNF-alpha retained its anti-apoptotic effects even at high concentrations. In conclusion, the protective effects against apoptosis of IFN-gamma, GM-CSF, and TNF-alpha itself are overruled when the concentration of TNF-alpha is high enough to produce a respiratory burst. These dual, concentration-dependent effects of TNF-alpha provide an explanation for previous controversial reports and support a dominant role for TNF-alpha in neutrophil apoptosis. PMID- 11261796 TI - Differential expression of Toll-like receptor 2 in human cells. AB - Human Toll-like receptor 2 (TLR2) is a receptor for a variety of microbial products and mediates activation signals in cells of the innate immune system. We have investigated expression and regulation of the TLR2 protein in human blood cells and tissues by using two anti-TLR2 mAbs. Only myelomonocytic cell lines expressed surface TLR2. In tonsils, lymph nodes, and appendices, activated B cells in germinal centers expressed TLR2. In human blood, CD14+ monocytes expressed the highest level of TLR2 followed by CD15+ granulocytes, and CD19+ B cells, CD3+ T-cells, and CD56+ NK cells did not express TLR2. The level of TLR2 on monocytes was after 20 h up-regulated by LPS, GM-CSF, IL-1, and IL-10 and down regulated by IL-4, IFN-gamma, and TNF. On purified granulocytes, LPS, GM-CSF, and TNF down-regulated, and IL-10 modestly increased TLR2 expression after 2 h. These data suggest that TLR2 protein expression in innate immune cells is differentially regulated by inflammatory mediators. PMID- 11261797 TI - Inhibition of fasL sustains phagocytic cells and delays myogenesis in regenerating muscle fibers. AB - Macrophage-muscle cell interactions are complex, and the majority is unknown. The persistence of inflammatory cells in skeletal muscle could be critical for myofiber viability. In the present paper, we show that FasL plays a role in the resolution of muscle inflammation. We analyzed inflamed muscles of normal mice treated from day 3 to day 8 with a FasL inhibitor (Fas-Ig) or with control Ig. Treated muscles were collected at 3, 5, and 10 days. The treatment with recombinant Fas-Ig protein induced a severe persistence of inflammatory cells at 5 days (115,000+/-27,838 vs. 41,661+/-6848, p<0.01) and 10 days from injury (145,500+/-40,850 vs. 5000+/-1000, p<0.001). Myofiber regeneration was highly impaired (37+/-14 vs. 252+/-28, p<0.01). Apoptosis of phagocytic cells was absent during Fas-Ig treatment (0.9+/-0.6 vs. 1300+/-150, p<0.0001), but apoptotic, mononucleated cells appeared at day 10, 2 days after the suspension of Fas-Ig administration. The time course of FasL expression during muscle inflammation, at mRNA and protein level, reveals a peak during myoblast proliferation. The peak of FasL expression coincides with the peak of apoptosis of phagocytic cells. In situ hybridization shows the co-expression of FasL and MyoD mRNA in mononucleated cells, i.e., myoblasts. Experiments on the myoblast cell culture confirmed the expression of FasL in myoblasts. The findings shown here indicate one of the pathways to control myoblast-macrophage interaction and might be relevant for the control of inflammatory cells in muscle tissue. Perhaps altering FasL expression with recombinant proteins could ameliorate inflammation in degenerative myopathies and up-regulate muscle regeneration. PMID- 11261798 TI - Caspase-mediated cleavage of TRAF3 in FasL-stimulated Jurkat-T cells. AB - The tumor necrosis factor receptor (TNFR)-associated factor (TRAF) proteins play a central role in the early steps of signal transduction by TNFR superfamily proteins, which induce various cellular responses, including apoptosis. Influences of TRAF proteins on the regulation of cell death and physical interactions between TRAFs and caspases have been reported. In this study, we demonstrate that TRAF3 is proteolyzed during cell death in a caspase-dependent manner. TRAF3 was found to be cleaved by incubation with caspase3 in vitro and by Fas- or CD3-triggering in Jurkat-T cells. The Fas- or CD3-induced cleavage of TRAF3 was blocked by caspase inhibitors and by introduction of alanine substitutions for D347 and D367 residues. Furthermore, the amino-terminal fragment of TRAF3 showed a different intracellular localization from the full length TRAF3 with preferential distribution to particulate fractions and the nucleus. These findings suggest that TRAF3 may be regulated by caspases during apoptosis of T cells. PMID- 11261799 TI - Leukocyte-specific gene 1 protein (LSP1) is involved in chemokine KC-activated cytoskeletal reorganization in murine neutrophils in vitro. AB - Leukocyte-specific gene 1 protein (LSP1) is a cytoskeletal-associated protein of leukocytes that in vitro cross-links F-actin into extensively branched bundles of mixed polarity. In this study, we examined chemotaxis and superoxide production in neutrophils prepared from wild-type (WT) and Lsp1 knockout mice. Compared to WT neutrophils, Lsp1-/- neutrophils showed impairment in both migration speed and chemotaxis direction during chemokine KC-directed chemotaxis. When examined by confocal microscopy, chemotaxing Lsp1-/- neutrophils showed abnormal morphologies. They had discontinuous primary actin-rich cortexes and large membrane protrusions. When stimulated by phorbol 12-myristate 13-acetate (PMA), Lsp1-/- peritoneal neutrophils produce more superoxide than WT. The data presented suggest that LSP1 plays important roles in the regulation of neutrophil morphology, motility, and superoxide production. PMID- 11261800 TI - A pharmacogenomic approach to Alzheimer's disease. AB - Single nucleotide polymorphisms (susceptibility genetics) and genomic point mutations (mendelian genetics) can be used in Alzheimer's disease (AD) for diagnostic, predictive and therapeutic purposes. Using a matrix genetic model, including APOE, PS1 and PS2 allelic variants, we have studied the distribution of 36 different genotypes in the AD population (N= 479) and the genotype-related cognitive response to a multifactorial therapy in AD patients with mild-to moderate dementia. The 10 most frequent AD genotypes are the following: 1) E33P112P2 + (17.75%), 2) E33P112P2- (15.55%), 3) E33P111P2+ (10.85%), 4) E34P112P2+ (9.60%), 5) E34P112P2- (7.56%), 6) E33P111P2- (7.10%), 7) E34P111P2+ (4.80%), 8) E33P122P2+ (4.38%), 9) E34P111P2- (4.18%), and 10) E34P122P2+ (3.55%). APOE-4/4-related genotypes represent less than 3% in the following order: E44P112P2 + > E44P111P2+ = E44P111P2- > E44P112P2+ > E44P122P2+ = E44P122P2. Multifactorial therapy with CDP-choline (1,000 mg/day) + piracetam (2,400 mg/day) + anapsos (360 mg/day) did improve mental performance during the first 6-15 months in a genotype-specific fashion. The best responders in the APOE series were patients with APOE-3/4 genotype (r= +0.013), while the worst responders were APOE-4/4 patients (r= -0.93). PS1-related genotypes responded in a similar manner; and patients with a defective PS2 gene exon 5 (PS2+) always showed a poorer therapeutic response than PS2- patients. All these data suggest that the therapeutic outcome in AD exhibits a genotype-specific pattern, and that a pharmacogenomic approach to AD might be a valuable strategy for drug development and monitoring. PMID- 11261801 TI - Magnetic resonance spectroscopy in Alzheimer's disease: focus on N acetylaspartate. AB - This paper reviews published post-mortem brain and in-vivo proton magnetic resonance spectroscopy (1H-MRS) studies in Alzheimer's disease (AD) and focuses on the emerging role of N-acetylaspartate (NAA) as a prognostic marker of neuronal function. Post-mortem brain studies have reported significantly lower NAA levels in AD brains than in control brains, and some have correlated the low levels with neuropathological findings (i.e. amyloid plaques and neurofibrillary tangles). Similarly, almost all published in-vivo studies have reported lower NAA levels in AD patients compared to elderly controls. While some studies have found changes in metabolite levels that were considered useful for the diagnosis of AD, most have found that 1H-MRS provided little or no advantages over other, more common diagnostic tools. Instead, recent studies in AD and other neuropsychiatric disorders suggest that NAA may be more useful as a prognostic marker for monitoring neurodegeneration, stabilization, or improvement, and for evaluating therapeutic response to novel drugs. PMID- 11261802 TI - Ligands for in vivo imaging of nicotinic receptor subtypes in Alzheimer brain. AB - The neuronal nicotinic acetylcholine receptors (nAChR) are involved in functional processes in brain including cognitive function and memory. A severe loss of the nAChRs has been detected in brain of patients with Alzheimer's disease (AD). There is a great interest to image nAChRs noninvasive for detection of receptor impairments even at a presymptomatic stage of AD as well for monitoring outcome of drug treatment. (S) [11C]Nicotine, has so far been the only nAChR ligand used in positron emission tomography (PET) studies for visualizing nAChRs in human brain. In order to develop PET/SPECT nAChRs ligands for detection of subtypes of nAChRs nicotine analogues, epibatidine and A-85380 compounds have been characterized in vitro and investigated in vivo. Epibatidine and A-85380 have been found to have higher specific signals and more favorable kinetic parameters than nicotine and its analogues. The epibatidine and A-85380 compounds can also be radiolabeled with high specific radioactivity, show affinities for the nAChRs in the pM range and readily cross the blood-brain barrier. In addition they reversibly bind to the nAChRs and show low non-specific binding and moderately fast metabolism. Due to a probably high alpha4beta2 nAChR selectivity combined with low toxicity, the A-85380 analogs presently seem to be the most promising nAChR ligand imaging of subtypes of nAChRs in human brain. PMID- 11261804 TI - Classical Alzheimer features and cholinergic dysfunction: towards a unifying hypothesis? AB - OBJECTIVE: Our autopsy studies show possible links between classical Alzheimer pathology and decreased expression of nicotinic acetylcholine receptors. For further elucidation we are now using in vitro models. We report preliminary evidence for the impact of beta-amyloid on nicotinic receptor expression in hippocampal dissociation culture. METHODS: Cultures (E18 rats) were grown in a serum-free medium and incubated at 8 days in vitro for 3 days with 1 microM Abeta1-42. Expression of alpha4, alpha7, and beta2 nicotinic receptor subunit protein was assessed immunohistochemically and rated semiquantitatively. RESULTS: Abeta1-42 incubation resulted in a massive reduction of alpha4 protein-expressing neurons, this effect was less pronounced for the alpha7 and beta2 subunit protein. CONCLUSION: These findings provide first evidence for a direct impact of classical Alzheimer pathology features on nicotinic receptor expression in vitro. Our model will be useful for testing the potential of drugs to stop or reverse these effects. PMID- 11261803 TI - Nicotinic receptors in dementia of Alzheimer, Lewy body and vascular types. AB - OBJECTIVES: Comparisons were made of nicotinic receptors in 3 major forms of dementia in old age. Although it is well established the involvement of nicotinic receptors in Alzheimer's disease (AD), their status in the other two main causes of dementia in old age-dementia with Lewy bodies (DLB) and vascular dementia (VaD) is not widely reported. METHODS: Temporal cortex was examined for epibatidine and alpha-bungarotoxin binding, and immunoreactivity of alpha4 and alpha7 nAChR subunits. RESULTS: There were selective abnormalities in nicotinic receptor subtypes in the disorders examined. In AD there is a loss of high affinity receptor binding, reflecting a selective loss of alpha4 subunit, but no change in alpha7 subunits. Similar abnormalities in ligand binding are also apparent in DLB. In the VaD series, there was no overall loss of epibatidine binding or immunoreactivity for alpha4 or alpha7 subunits. CONCLUSIONS: Loss of cortical receptor alpha4 subunit appears to be a characteristic feature of neurodegenerative dementia but not dementia of vascular origin. Since nicotinic receptors control cerebral vasodilation, the relative integrity of the receptors in VaD may auger well for nicotinic therapy in this disorder in which there is a cholinergic abnormality, to judge by the loss of the presynaptic enzyme. PMID- 11261805 TI - An unusually glycosylated form of acetylcholinesterase is a CSF biomarker for Alzheimer's disease. AB - The identification of a biochemical marker of Alzheimer's disease (AD) is a major research aim of many groups. Abnormal levels of tau and Abeta have been identified in the cerebrospinal fluid (CSF) of AD patients, although the sensitivity and specificity of the changes in these two biomarkers alone is not sufficient to be of diagnostic value. Recently, our group has identified an abnormality in the glycosylation of acetylcholinesterase (AChE). The increase in this glycoform of AChE is very specific for Alzheimer's disease and is not seen in many other neurological diseases including other dementias. PMID- 11261806 TI - Acetylcholinesterase-amyloid-beta-peptide interaction and Wnt signaling involvement in Abeta neurotoxicity. AB - Previous studies have indicated that acetylcholinesterase (AChE) promotes amyloid beta-peptide (Abeta) fibril formation and AChE-Abeta complexes increase Abeta dependent neurotoxicity. Here we present evidence for the: i) identification of the AChE motif that promotes amyloid formation, ii) in vivo effect of AChE on brain plaque formation, and iii) connection between AChE-Abeta neurotoxicity and the Wnt signal transduction pathway. Computer modeling, stereotaxic infusions and cell biological techniques were used to study the above problems. Results indicated that a 3.4 kDa AChE peptide promotes Abeta fibril formation. AChE infusion into rat hippocampus determines the appearance of anti-Abeta and thioflavine-S positive plaques, and AChE-Abeta toxicity on hippocampal cultures was blocked by lithium, an activator of the Wnt cascade. We suggest that AChE Abeta/Abeta dependent neurotoxicity may result in loss of function of Wnt signaling components, and open the possibility that lithium may be considered as a candidate for therapeutic intervention in Alzheimer's disease pathology. PMID- 11261807 TI - Mutation of conserved aspartates affect maturation of presenilin 1 and presenilin 2 complexes. AB - Presenilin (PS1 and PS2) holoproteins are transiently incorporated into low molecular weight (MW) complexes. During subsequent incorporation into a higher MW complex, they undergo endoproteolysis to generate stable N- and C-terminal fragments (NTF/CTF). Mutation of either of two conserved aspartate residues in transmembrane domains inhibits both presenilin-endoproteolysis and the proteolytic processing of APP and Notch. We show that aspartate-mutant holoprotein presenilins are not incorporated into the high molecular weight, NTF/CTF-containing complexes. Aspartate-mutant presenilin holoproteins also preclude entry of endogenous wild-type PS1/PS2 into the high molecular weight complexes, but do not affect the incorporation of wild-type holoproteins into lower molecular weight holoprotein complexes. These data suggest that the loss-of function aspartate-mutants cause altered PS complex maturation, and argue that the functional presenilin moieties are contained in the high molecular weight presenilin NTF/CTF-containing complexes. PMID- 11261808 TI - Galantamine is an allosterically potentiating ligand of the human alpha4/beta2 nAChR. AB - Galantamine (Reminyl) is a novel drug treatment for mild to moderate Alzheimer's disease (AD). Originally established as a reversible inhibitor of the acetylcholine-degrading enzyme acetylcholinesterase (AChE), galantamine also acts as an allosterically potentiating ligand (APL) on nicotinic acetylcholine receptors (nAChR). Having previously established this second mode of action on nAChRs from murine brain, we demonstrate here the same action of galantamine on the most abundant nAChR in the human brain, the alpha4/beta2 subtype. This nAChR sensitizing action is not a common property of all, or most, AChE inhibitors, as is shown by the absence of this effect for other therapeutically applied AChE inhibitors including tacrine, metrifonate, rivastigmine and donepezil. The possible benefits for therapy of AD of an APL action on nicotinic receptors is discussed. PMID- 11261809 TI - The experimental Alzheimer drug phenserine: preclinical pharmacokinetics and pharmacodynamics. AB - Phenserine, a phenylcarbamate of physostigmine, is a new potent and highly selective acetylcholinesterase (AChE) inhibitor, with a > 50-fold activity versus butyrylcholinesterase (BChE), in clinical trials for the treatment of Alzheimer's disease (AD). Compared to physostigmine and tacrine, it is less toxic and robustly enhances cognition in animal models. To determine the time-dependent effects of phenserine on cholinergic function, AChE activity, brain and plasma drug levels and brain extracellular acetylcholine (ACh) concentrations were measured in rats before and after phenserine administration. Additionally, its maximum tolerated dose, compared to physostigmine and tacrine, was determined. Following i.v. dosing, brain drug levels were 10-fold higher than those achieved in plasma, peaked within 5 min and rapidly declined with half-lives of 8.5 and 12.6 min, respectively. In contrast, a high (> 70%) and long-lasting inhibition of AChE was achieved (half-life > 8.25 h). A comparison between the time dependent plasma AChE inhibition achieved after similar oral and i.v. doses provided an estimate of oral bioavailability of 100%. Striatal, in vivo microdialysis in conscious, freely-moving phenserine-treated rats demonstrated > 3-fold rise in brain ACh levels. Phenserine thus is rapidly absorbed and cleared from the body, but produces a long-lasting stimulation of brain cholinergic function at well tolerated doses and hence has superior properties as a drug candidate for AD. It selectively inhibits AChE, minimizing potential BChE side effects. Its long duration of action, coupled with its short pharmacokinetic half life, reduces dosing frequency, decreases body drug exposure and minimizes the dependence of drug action on the individual variations of drug metabolism commonly found in the elderly. PMID- 11261810 TI - Anti-inflammatory therapy for Alzheimer's disease: implications of the prednisone trial. AB - The inflammatory hypothesis of Alzheimer's disease (AD), which is supported both by basic laboratory evidence and epidemiological studies, suggests that treatment with anti-inflammatory drugs may reduce the risk or slow the progression of AD. In the first large-scale test of this hypothesis, the Alzheimer's Disease Cooperative Study (ADCS) conducted a randomized placebo-controlled trial of low dose prednisone treatment in subjects with probable AD. There was no difference in cognitive decline between the prednisone and placebo treatment groups; subjects treated with prednisone showed behavioral decline compared to those in the placebo group. While this study indicates that a low-dose regimen of prednisone is not useful in the treatment of AD, it does not refute the inflammatory hypothesis; recent evidence supports testing of a number of alternative anti-inflammatory regimens, for prevention and/or treatment of AD. The ADCS has initiated a trial to determine whether treatment with a non selective non-steroidal anti-inflammatory drug or a selective cyclooxygenase-2 inhibitor is effective in slowing the rate of cognitive decline in AD. PMID- 11261811 TI - Inhibiting the conversion of soluble amyloid-beta peptide into abnormally folded amyloidogenic intermediates: relevance for Alzheimer's disease therapy. AB - Alzheimer's disease is a degenerative disorder of the brain for which there is no cure or effective treatment. Recent studies suggest that cerebral amyloid plaques are central to the disease process. However, it is not clear which of the species going from the normal soluble amyloid-beta peptide to the mature amyloid plaque is the toxic agent in the brain. Therefore, an attractive therapeutic strategy for Alzheimer's disease is to block the early steps involving the pathological conversion of the soluble peptide into the abnormally folded oligomeric intermediate precursor of the amyloid fibrils. We have engineered synthetic beta sheet breaker peptides to bind amyloid-beta peptide, stabilize the normal conformation and destabilize the beta-sheet rich structure of the potentially toxic intermediates and hence the formation of amyloid plaques. Results in vitro, in cell culture and in vivo suggest that beta-sheet breaker peptide may be useful for blocking the pathway that lead to the formation of cerebral amyloid deposits. It remains to be proved that inhibition of the defective folding of amyloid-beta peptide and/or amyloid plaque deposition could be beneficial for the therapeutic treatment of Alzheimer's disease. PMID- 11261812 TI - Immediate causes of death of demented and non-demented elderly. AB - OBJECTIVE: To investigate the immediate causes of death, in autopsied demented and non-demented elderly. DESIGN: Retrospective clinicopathologic correlations. SETTING: Acute and intermediate care geriatric hospital. PARTICIPANTS: 342 hospitalized demented and non-demented elderly (mean age 84.94 +/- 6.9 years) who underwent consecutive postmortem examinations: 120 demented patients with either vascular dementia (VaD, n = 34), mixed dementia (MD, n = 65) or Alzheimer's disease (AD, n=21) neuropathologically confirmed and 222 nondemented elderly. RESULTS: Primary causes of death were similar in both demented and non-demented patients; the commonest were cardiovascular disease and bronchopneumonia. Cardiac causes of death and especially cardiac failure were more frequent in VaD than in AD or MD (respectively P = 0.027 and 0.005). Dementia was an underlying but never a primary cause of death. CONCLUSIONS: Immediate causes of death are similar in elderly demented and non-demented patients. PMID- 11261814 TI - Connexin 43 expression in rat aortic smooth muscle after ovariectomy and hormonal replacement. AB - The purpose of this study was to determine the effects of ovariectomy and long term combined sexual hormone replacement on the gap junctional protein, connexin 43 (Cx43) of aortic medial smooth muscle cells in rats. Twenty non-pregnant mature Wistar female rats were divided into five groups (four animals in each group). Group A underwent ovariectomy, Group B underwent ovariectomy and received estradiol propionate, Group C underwent ovariectomy and received medroxyprogesterone acetate and Group D underwent ovariectomy and received both hormones. Group E was sham-operated and used as control. After 15 weeks of treatment, thoracic aortas were removed and immunohistochemistry was carried out using a specific fluorescent antibody against Cx43. Tissue sections were examined by confocal laser scanning microscopy and analysed by the Scion Image program. All five different groups had the same distribution and extent of Cx43 in the aorta. Neither the ovariectomy nor the hormone replacement had any effect on the Cx43 expression of aortic smooth muscle cells in rats as compared to control animals. These results indicate that sexual steroids do not influence the gap junctional protein Cx43 of the medial layer of aorta in rats. They may suggest that the beneficial effects of estrogen are not mediated via gap junctions in the human aorta either. PMID- 11261813 TI - Expression of tyrosine kinase receptors Tie-1 and Tie-2 in giant cell tumor of the tendon sheath: a possible role in synovial proliferation. AB - We have recently demonstrated that Tie-1 and Tie-2 are expressed in synovial cells from rheumatoid arthritis (RA). To elucidate the possible involvement of Tie receptors in synovial proliferation, we analyzed their expression by immunostaining in five cases of giant cell tumor of tendon sheath (GCTTS), which represents a proliferating lesion of synovial cells. Strong immunoreactivity for both Tie-1 and Tie-2, regardless of the individual patient's profile, was observed in all cases of GCTTS. Six sets of double immunohistochemical stainings for Tie-1/Tie-2 and fibronectin, CD68, or CD34 were carried out to determine the phenotype of Tie-1 and Tie-2-positive tumor components. In these studies, both Tie-1 and Tie-2 immunoreactivity were widely observed in the fibronectin-positive fibroblastic and the CD68-positive histiocytic mononuclear cells, as well as in the osteoclast-like giant cells. In tumor vasculature, Tie receptors were expressed in the CD34-positive endothelial cells possessing proliferating cell nuclear antigen (PCNA) immunoreactivity. We also evaluated the correlation of Tie 1/Tie-2 expression and proliferating cells in GCTTS by using double staining of Tie-1/Tie-2 together with PCNA. Overexpression of PCNA immunoreactivity was frequently found in Tie receptors-positive cells with no obvious differences in the expression pattern of Tie-1 and Tie-2. These findings suggest the possible involvement of Tie receptors in the pathogenesis of GCTTS other than solely via their involvement in angiogenesis and subsequent vascularization. It was demonstrated that Tie-2 immunoreactivity was restricted to the fibroblastic, but not histiocytic, phenotype in RA synovium, suggesting different regulatory control of Tie-2 expression in GCTTS and RA synovium. Overexpression of Tie receptors in GCTTS may imply a biological role for these receptors in synovial proliferation. PMID- 11261815 TI - Primary pulmonary primitive neuroectodermal tumor (PNET). A case report. AB - We describe a rare case of a primary primitive neuroectodermal tumor (PNET) in the lung of a 17-year-old girl. Grossly, the tumor, located in the right lower lobe, was relatively well-circumscribed and whitish to yellowish in color with scattered hemorrhagic necrosis. Microscopically, the tumor was composed of ovoid to polygonal cells with a high nuclear to cytoplasmic ratio and relatively scant cytoplasm, arranged in solid sheets with intervening fine fibrovascular stroma. Immunohistochemically, the tumor was positive for the MIC2 gene product, whereas AE1/AE3, CAM5.2, and a variety of neuroendocrine markers such as chromogranin A, synaptophysin, and ProGRP, were negative. Three months after the lobectomy, recurrent tumors were noted in the mediastinum and right thoracic wall, and she died despite combined chemotherapy and radiation therapy. In this case cytogenetic analysis showed a hypertriploid karyotype with multiple numerical and structural chromosomal aberrations, but failed to disclose distinct evidence of translocation between chromosome 11 and 22. However, the reverse transcriptase polymerase chain reaction (RT-PCR) demonstrated EWS/FLI-1 fusion transcripts, confirming the histopathologic diagnosis of PNET. This case indicates that the primary pulmonary PNET is a highly aggressive neoplasm occurring at a young age, and should prompt combined systemic chemotherapy, even though it is organ confined. PMID- 11261816 TI - Molecular diagnosis of a Mycobacterium chelonae infection. AB - A 23-year-old female presented with enlarged cervical lymph nodes, and a diagnosis of nonspecific lymphadenitis with formation of pyogranulomas was rendered. Despite an initial oral antibiosis and subsequent long-term intravenous and oral antibiosis under hospitalized conditions, the symptoms progressed. The lymph nodes became larger and then affected the cervical region bilaterally. Her general condition worsened, and an exanthema of the extremities accompanied by a reactive arthritis occurred. Serological assays of various viral and bacterial markers and blood cultures were negative. Application of a polymerase chain reaction (PCR) protocol allowing specific amplification of mycobacterial DNA revealed DNA of Mycobacterium chelonea in formalin-fixed, paraffin-embedded lymph node tissue. Sequencing of the PCR product showed a 97% homology with the known Mycobacterium chelonae sequence. Modification of the antibiotic therapy with clarithromycin, imipenem and amikacin resulted in a rapid regression of the symptoms. The clinical course, in combination with the difficulties in detecting the infectious agent, supports the usefulness of molecular pathological analyses specific for nontuberculous mycobacteria (NTM). PMID- 11261817 TI - Small cell neuroendocrine carcinoma of the thymus complicated by Cushing's syndrome. Report of a 58-year-old woman with a 3-year history of hypertension. AB - A 58-year-old woman with a history of Cushing's syndrome for three years presented with a mediastinal mass and received the diagnosis of small cell neuroendocrine carcinoma of the thymus invading the pericardium. On immunohistochemical study, the neoplastic cells reacted with antibodies against cytokeratin, epithelial membrane antigen, neuron-specific enolase, chromogranin, synaptophysin, and ACTH. Clinicopathologic findings of this rare case of ectopic adrenocorticotropic hormone (ACTH) syndrome are discussed with a literature review. PMID- 11261818 TI - Uterine leiomyoma with massive lymphocytic infiltration simulating malignant lymphoma. A case report with immunohistochemical study showing that the infiltrating lymphocytes are cytotoxic T cells. AB - Uterine leiomyoma with massive lymphoid infiltration is very rare and may simulate malignant lymphoma. To the best of our knowledge, this is the first description of such a lesion occurring in an Oriental, and the ninth case in the English literature. A 50-year-old Taiwanese woman had urinary frequency and nocturia because of a uterine myoma. The myomectomy specimen was identified as a well-defined tumor, 6.5-cm in diameter, the cut surface of which was pale, white and whorled. A massive lymphocytic infiltration accompanied by plasma cells and histiocytes was noted in the leiomyoma but not in the surrounding non-neoplastic myometrial fibers. Most infiltrating lymphocytes were positive for CD3 and T cell intracellular antigen-1, a cytotoxic marker. The postoperative course was uneventful, and the urinary symptoms improved within a 6-month follow-up period. PMID- 11261819 TI - Aggressive angiomyxoma of the spermatic cord. Two unusual cases occurring in childhood. AB - We report on two cases of aggressive angiomyxoma (AAM) of the spermatic cord occurring in two 13-year-old children. Clinically, the tumor simulated a mass of the spermatic cord. Histologically, it represented a poorly circumscribed, benign myxoid tumor, with a sparse population of stromal cells immunoreactive for vimentin and, focally, for smooth muscle actin. No immunostaining for desmin, S 100, p53, p21waf-1, c-Erb-B2 and estrogen-progesterone receptors was found. High proliferating cell nuclear antigen (PCNA) immunoexpression found in most of the tumor cells may explain the high risk of recurrence. AAM should be considered in the differential diagnosis of a spermatic cord mass occurring during infancy. PMID- 11261820 TI - Pathology and pathogenesis of idiopathic portal hypertension with an emphasis on the liver. AB - Idiopathic portal hypertension (IPH) is characterized by a long-standing presinusoidal portal hypertension of unknown etiology in adults. Some unidentified agent(s) affect(s) the intrahepatic small portal veins or portal tracts. Immunological disturbance, thromboembolism, infectious etiology and/or increased fibrogenesis in portal tracts are suspected as being candidates for the primary agent(s). During the long clinical course of IPH, several pathological changes may occur, including subcapsular parenchymal atrophy, atrophy of the liver, portal and parenchymal fibrosis, and portal venous phlebosclerosis and thrombosis. The last-named of these lesions is mostly found in patients with a history of splenectomy. Subcapsular parenchymal and hepatic atrophy may result from a hepatocellular dropout via apoptosis or necrosis because of intrahepatic hemodynamic disturbances, particularly chronic portal venous blood insufficiency. Pericellular fibrosis and thin fibrous septa are also frequently found and associated with activated perisinusoidal cells positive for smooth muscle actin. At the same time, vague nodular hyperplasia of hepatocytes not surrounded by fibrous septa is not infrequently seen. It may resemble nodular regenerative hyperplasia, partial nodular transformation, or focal nodular hyperplasia. However, liver cirrhosis does not occur even at the terminal stage. Taking these findings into consideration, a new staging of IPH with a combination of hepatic parenchymal atrophy and portal venous thrombosis was proposed: non-atrophic liver without subcapsular parenchymal atrophy (stage I), non-atrophic liver with subcapsular parenchymal atrophy (stage II), atrophic liver with subcapsular parenchymal atrophy (stage III), and portal venous occlusive thrombosis (stage IV). IPH livers are likely to progress from stage I to stage III. Stage IV, which occurs relatively late, has a poor prognosis. This staging is applicable to clinical and autopsy cases without any histological data. PMID- 11261821 TI - Polycythemia rubra vera versus secondary polycythemias. A clinicopathological evaluation of distinctive features in 199 patients. AB - To determine parameters of distinctive value in polycythemia rubra vera (PV) versus secondary polycythemias (SP), a clinicopathological study was performed on 199 patients. These presented with a borderline to marked elevation of the hemoglobin level (> 18 g/dl in men and > 16 g/dl in women). Evaluations of clinical features and bone marrow histopathology were carried out independently. According to the results derived from laboratory data and representative pretreatment trephine biopsies, three groups of patients emerged: group I presenting with the concordant clinical and morphological findings of early to manifest PV (136 patients), group II consisting of 55 patients with the congruent signs and symptoms of SP mostly caused by various chronic bronchopulmonal disorders, and finally eight patients (group III) with divergent findings. Between group I and II patients (PV versus SP), a number of clinical parameters proved to be significantly different. With the exception, of the red cell mass, platelet count, leukocyte alkaline phosphatase, LDH, spleen size, and the erythropoietin level had a significantly discriminating impact. Morphological features of distinctive value consisted of a set of specific lesions. Contrasting SP with an only borderline to slight increase in cellularity associated with a moderate enlargement of the erythroblastic islets, PV was always characterized by a significant increase in hematopoiesis, revealing a trilinear proliferation (panmyelosis). Megakaryopoiesis was strikingly different in PV as compared to SP by displaying clustering and a pleomorphous appearance. i.e., very small and giant megakaryocytes with staghorn-like nuclei were neighboring each other. Moreover, conspicuous alterations of the interstitial compartment were recognizable in SP. These consisted of deposits of cell debris in histiocytic reticular cells, iron-laden macrophages, and a plasmacytosis, implying an inflammatory reaction. These changes were only very rarely observed in PV, as opposed to a minimal to slight increase in reticulin fibers in about 12% of patients. In conclusion, a more elaborate evaluation of bone marrow features resulted in a set of diagnostic criteria with discriminating capacity that should be considered in prospective clinical trials. PMID- 11261822 TI - Perinuclear and cytoplasmic distribution of desmoglein in esophageal squamous cell carcinomas. AB - The desmosomal glycoproteins desmoglein (Dsg) and desmocollin (Dsc) are members of the cadherin family of cell adhesion molecules. They play an important role in epithelial adhesion. To observe the distribution pattern of Dsg in esophageal squamous cell carcinomas (SCC), immunohistochemical and immunoelectron microscopic analyses were performed. Immunohistochemically, normal esophageal squamous cells strongly expressed Dsg at the cell-cell boundaries, while moderately differentiated esophageal SCC cells showed a perinuclear distribution in addition to the cell boundary staining. At the ultrastructural level, the reaction product was concentrated at the desmosomes in the cell membrane region of normal epithelial cells, but was reduced at the membrane and found throughout the cytoplasm as well as in the surrounding outer nuclear envelope in SCC cells. These results demonstrate an aberrant distribution of Dsg in SCC cells. This may have important consequences for invasion and metastasis, as it may indicate loosened intercellular adhesion. PMID- 11261823 TI - Early gastric stump carcinoma with rhabdoid features. PMID- 11261824 TI - Expression and clinical significance of the erbB family in intrahepatic cholangiocellular carcinoma. AB - The type I family of growth factor receptors is known to play a role in the development of several carcinomas, but its role in hepatic malignancies is not clearly understood. In this study we investigated the expression of this family of EGF-R, c-erbB-2, c-erbB-3 and c-erbB-4 in 38 intrahepatic cholangiocellular carcinomas (CCC) by means of immunohistochemistry. EGF-R expression was related to lymph node metastasis, aberrant p53 expression, proliferating activity, and carcinoma differentiation. c-erbB-2 expression was observed in more than 50% of the cases, but was not related to any clinicopathological features, c-erbB-3 expression was linked to lymph node metastasis, and c-erbB-4 expression was directly related to proliferating activity and lymph node metastasis. These results indicate that: 1) EGF-R contributes greatly to CCC progression, and c erbB-3 and c-erbB-4 have roles similar to but less than that of EGFR, and 2) c erbB-2 is expressed in CCC in high incidence, but its clinical role in CCC remains unclear. PMID- 11261825 TI - Malignant MCF10CA1 cell lines derived from premalignant human breast epithelial MCF10AT cells. AB - The MCF10 series of cell lines was derived from benign breast tissue from a woman with fibrocystic disease. The MCF10 human breast epithelial model system consists of mortal MCF10M and MCF10MS (mortal cells grown in serum-free and serum containing media, respectively), immortalized but otherwise normal MCF10F and MCF10A lines (free-floating versus growth as attached cells), transformed MCF10AneoT cells transfected with T24 Ha-ras, and premalignant MCF10AT cells with potential for neoplastic progression. The MCF10AT, derived from xenograft passaged MCF10-AneoT cells, generates carcinomas in approximately 25% of xenografts. We now report the derivation of fully malignant MCF10CA1 lines that complete the spectrum of progression from relatively normal breast epithelial cells to breast cancer cells capable of metastasis. MCF10CA1 lines display histologic variations ranging from undifferentiated carcinomas, sometimes with focal squamous differentiation, to well-differentiated adenocarcinomas. At least two metastasize to the lung following injection of cells into the tail vein; one line grows very rapidly in the lung, with animals moribund within 4 weeks, whereas the other requires 15 weeks to reach the same endpoint. In addition to variations in efficiency of tumor production, the MCF10CA1 lines show differences in morphology in culture, anchorage-independent growth, karyotype, and immunocytochemistry profiles. The MCF10 model provides a unique tool for the investigation of molecular changes during progression of human breast neoplasia and the generation of tumor heterogeneity on a common genetic background. PMID- 11261826 TI - P53 protein accumulation in non-invasive lesions surrounding p53 mutation positive invasive breast cancers. AB - p53 mutation is a common event in sporadic breast cancer being found in 15-50% of invasive carcinomas. The purpose of this study was to determine the earliest histologic stage at which p53 mutation could be detected with a widely used anti p53 antibody (DO7, Novocastra) which recognizes both wild type and mutant forms. p53 expression was assessed immunohistochemically in 12 primary breast carcinomas with known p53 mutations and in all pre-malignant epithelial lesions surrounding these invasive cancers. Strong p53 nuclear staining was found in all of the tumors known to have missense mutations and none of the tumors with truncation mutations. In cases with intense staining in the invasive carcinoma, a similar quality of staining was also seen in all areas of DCIS (ductal carcinoma in situ) and was representative of missense p53 mutations. Lighter nuclear staining intensity was observed in up to 40% of cells in areas of hyperplasia and in up to 30% of normal breast lobules irrespective of the type of mutation found in the invasive carcinoma. This weak staining was not specific to mutated p53 and may indicate increased amounts of normal p53 protein. We conclude that p53 inactivation occurs prior to invasion in breast carcinogenesis, with mutations being uniformly identified in DCIS associated with p53-mutated invasive carcinomas. In contrast, there is no evidence that epithelial hyperplasia or epithelial cells of the terminal duct lobular unit harbor the same mutations as their associated invasive carcinoma. PMID- 11261827 TI - Phase III randomized trial of toremifene vs tamoxifen in hormonodependant advanced breast cancer. AB - PURPOSE: Efficacy and safety of toremifene (TOR) 60 mgs/dayly/o.r. was compared with tamoxifen (TAM) 40 mgs/dayly/o.r. in a group of postmenopausal women with advanced breast cancer, without previous systemic therapy for advanced breast cancer. MATERIAL AND METHODS: The study was a prospective double-blind randomized trial. All treated patients presented with positive estrogen receptors. Main end points were response rates, toxicity profile analysis, time to progression and survival. WHO and ECOG criteria were employed for response evaluation while toxicity was assesed according to WHO guidelines. Curves were constructed by means of Kaplan-Meier methodology and were compared by means of log-rank test. RESULTS: From January 1996 to January 1999 a total of 217 patients were included in the study (106 in the TOR branch and 111 in the TAM arm). Both groups of patients were homogeneous regarding the main prognostic factors. A response rate of 64% (68/106) was observed in the TOR group as compared with a 52% (58/111) in the TAM group. Median times to progression and overall survival were not significantly different. A lower incidence of undesirable effects was apreciated in the TOR arm. CONCLUSIONS: Our data suggest that TOR is an efficient and well tolerated agent for the therapy of postmenopausal women with hormonal positive receptors advanced breast cancer, and must be considered an alternative to TAM as first line therapy for ER+ advanced breast cancer patients and as well as an adjuvant treatment. PMID- 11261828 TI - Continued breast cancer risk reduction in postmenopausal women treated with raloxifene: 4-year results from the MORE trial. Multiple outcomes of raloxifene evaluation. AB - Raloxifene, a selective estrogen receptor modulator approved for the prevention and treatment of postmenopausal osteoporosis, has shown a significant reduction in breast cancer incidence after 3 years in this placebo-controlled, randomized clinical trial in postmenopausal women with osteoporosis. This article includes results from an additional annual mammogram at 4 years and represents 3,004 additional patient-years of follow-up in this trial. Breast cancers were ascertained through annual screening mammograms and adjudicated by an independent oncology review board. A total of 7,705 women were enrolled in the 4-year trial; 2,576 received placebo, 2,557 raloxifene 60 mg/day, and 2,572 raloxifene 120 mg/day. Women were a mean of 66.5-years old at trial entry, 19 years postmenopause, and osteoporotic (low bone mineral density and/or prevalent vertebral fractures). As of 1 November 1999, 61 invasive breast cancers had been reported and were confirmed by the adjudication board, resulting in a 72% risk reduction with raloxifene (relative risk (RR) 0.28, 95% confidence interval (CI) 0.17, 0.46). These data indicate that 93 osteoporotic women would need to be treated with raloxifene for 4 years to prevent one case of invasive breast cancer. Raloxifene reduced the risk of estrogen receptor-positive invasive breast cancer by 84% (RR 0.16, 95% CI 0.09, 0.30). Raloxifene was generally safe and well-tolerated, however, thromboembolic disease occurred more frequently with raloxifene compared with placebo (p=0.003). We conclude that raloxifene continues to reduce the risk of breast cancer in women with osteoporosis after 4 years of treatment, through prevention of new cancers or suppression of subclinical tumors, or both. Additional randomized clinical trials continue to evaluate this effect in postmenopausal women with osteoporosis, at risk for cardiovascular disease, and at high risk for breast cancer. PMID- 11261829 TI - Change in expression of ER, bcl-2 and MIB1 on primary tamoxifen and relation to response in ER positive breast cancer. AB - Pre-treatment oestrogen receptor (ER) expression in breast cancer predicts for rate of response to endocrine therapy but not for the quality or duration of response (DofR). ER is known to be down-regulated by anti-oestrogens. This study has tested the hypothesis that the degree of down-regulation of ER and the ER regulated marker bcl-2 are associated with the quality and duration of tamoxifen response. 80 patients with ER+ve breast cancer (H-score > 10) receiving primary tamoxifen (n = 51 Stage I-II elderly; n = 29 Stage III) underwent sequential tumour biopsies for immunocytochemical assessment of ER, bcl-2 and the proliferation marker MIB1. Median follow-up is 45 months. By 6-months on therapy three patients had attained complete response (CR), 27 partial response (PR); 44 static disease (SD) and six progression (PD) by UICC criteria. Greater decrease in ER and bcl-2 H-score from pre-treatment to 6 weeks (p = 0.035, p = 0.037) and ER and bcl-2 H-score from pre-treatment to 6 months (p = 0.058, p = 0.036) were significantly associated with better quality of response (CR/PR vs SD/PD). Greater 6-week and 6-month reduction in bcl-2 H-score (p = 0.041, p = 0.036) and 6-week reduction in MIB1 (p = 0.013) were significantly correlated with longer DofR. This study demonstrates that greater down-regulation of ER and the ER regulated protein bcl-2 on primary tamoxifen are significantly associated with a better quality of response and bcl-2 and the proliferation marker MIB1 a longer duration of response in ER+ve breast cancer. PMID- 11261830 TI - Fractionated radiosurgery for brain metastases in 43 patients with breast carcinoma. AB - About 15% of metastatic breast carcinoma patients are diagnosed with brain metastases. Historically, the majority are treated with palliative external whole brain radiation with a median survival of 4 months. We examined stereotactic radiosurgery's effect on treatment outcome in such patients. Four hundred and fifty four consecutive patients with brain metastases were treated with stereotactic radiosurgery at Staten Island University Hospital, NY, between 1991 and 1999. The medical records of 60 women with histologically confirmed breast cancer were retrospectively reviewed. Forty-three patients (71%) received fractionated radiosurgery (4 x 600 cGy) and form the core of this report. Sixty five percentage had been previously unsuccessfully treated by whole-brain radiation or had recurrence after craniotomy. Survival was calculated by the Kaplan-Meier method. The median age at diagnosis of brain metastases was 52 years, with median interval of 49 months following the diagnosis of tumor primary. Median survival from brain diagnosis reached 13.6 months. Overall median survival from radiosurgery treatment was 7.5 months. Fifteen patients with one or two brain lesions survived a median of 11.5 months. For the fractionated cohort of patients 1- and 2-year actuarial survival was 28.2% and 12.8%, respectively. Three patients are alive at 32, 34 and 64 months, respectively. We conclude that fractionated radiosurgery improves survival of patients with brain metastases from breast cancer, especially those with small lesions, good functional status and no other metastatic disease. These patients should be encouraged to consider radiosurgery, possibly before WBRT. Considering our 7.5 months overall survival including patients with multiple metastases, and patients with progressive brain metastases despite extensive standard therapy and often systemic disease, these results suggest that radiosurgery could benefit breast cancer patients with brain metastases and extend life. PMID- 11261831 TI - Selection of primary breast cancer patients for adjuvant endocrine therapy--is oestrogen receptor alone adequate? AB - Among 834 patients who had primary breast cancer treated by surgery without adjuvant systemic therapy, 363 had relapse treated by endocrine therapy alone. Patients with oestrogen receptor positive tumours (median: 70 vs. 45 months, p < 0.0001) or with non-progression at 6 months of therapy (median: 111 vs. 37 months, p < 0.0001) survived longer than those with oestrogen receptor negative tumours or with disease progression respectively, presumably due to the effect of therapy. On the other hand, the median disease-free interval, uninfluenced by therapy, showed a similar difference: oestrogen receptor positive versus negative = 29 versus 21 months, p < 0.005; non-progression versus progression = 40 versus 19 months, p < 0.0001. Patients with oestrogen receptor-positive tumours and non progression at 6 months had the longest disease-free interval. The present study has established that there are factors, other than the oestrogen receptor, inherent in the primary tumour as reflected by the disease-free interval, which affect hormone sensitivity. Selection of adjuvant endocrine therapy based on the oestrogen receptor alone would deem inadequate. Further studies to elucidate other possible factors are warranted to refine the use of endocrine therapy, especially in the adjuvant setting when no indication of response is available. PMID- 11261832 TI - Effects of oral contraceptives on breast epithelial proliferation. AB - The association between oral contraceptive (OC) use and breast cancer is not fully understood. Estrogen is a known mitogen to breast epithelial cells, but there is still a controversy about the effect of added progestogens. Fine needle aspiration (FNA) biopsies were used to assess epithelial proliferation in normal breast tissue from 106 healthy premenopausal women with and without oral contraceptives. In 26 women biopsies were performed before and after 2 months of OC use. Proliferation, expressed as percentage of Ki-67/MIB-1 positive cells, was correlated to endogenous progesterone, androgenic/anabolic compounds and exogenous progestogen. We found a higher proliferation (p = 0.03) in OC users compared to non users, with mean values of 4.8% and 2.2%, respectively. There was a positive correlation between proliferation and progesterone levels in non-users and with serum levonorgestrel concentrations in women using OCs containing this progestogen (rs = 0.43, p = 0.02). Women using OCs had significantly lower serum androgen levels compared to naturally cycling women and free testosterone levels displayed an inverse relation to breast epithelial proliferation. There was a marked variation in the response to exogenous sex steroids. In certain women after 2 months of OC use, the percentage of MIB-1 positive cells was as high as 40-50%. The results add to the growing evidence that progestogens may be mitogenic in breast tissue. Increased proliferation during hormonal contraception should be regarded as an unwanted and potentially hazardous side effect. Efforts should be made to define hormonal contraceptive regimens which minimize breast epithelial proliferation and to identify those women with the most pronounced proliferative response. PMID- 11261833 TI - Microsatellite instability is associated with the loss of apoptosis in ductal breast carcinomas. AB - Metastatic progression in ductal breast carcinomas are related to apoptosis in primary tumors. Frameshift mutations in a single-repeat sequence within the coding region (G)8 of the pro-apoptotic Bax gene have been related to microsatellite instability (MSI) and progression of some carcinomas and lymphomas. The aim of this study was to explore whether the extended lifespan of breast cancer cells can also be triggered by Bax mutation in ductal-breast carcinomas, and whether breast cancer cell MSI is related to the loss of apoptosis. For this purpose we studied frameshift mutations of a microsatellite (G)8 in the third exon of the Bax gene in a series of 105 ductal breast carcinomas, at T1 and T2-3 stages, 45 of which had lymph node metastasis. We analyzed MSI in five sequences of DNA isolated from normal and tumor tissue samples taken from 86 patients, and we explored the relationship between MSI and tumor apoptosis status. Bax mutation was not present in ductal breast carcinomas. MSI (two or more markers altered) was detected in 11.6% of tumors. Loss of apoptosis occurred in 80% (8/10) tumors with MSI, versus 17.8% of tumors without MSI (chi2 test, p = 0.0004), independently of Bax protein expression. We conclude that frameshift mutations of a microsatellite (G)8 of the Bax gene are not critical for the loss of apoptosis in breast cancer, and that loss of apoptosis may be a consequence of overexpression of anti-apoptotic protein Bcl-2 or Bcl-xL. Moreover, MSI in breast carcinomas might be the cause of loss of an apoptotic pathway that is not induced by frameshift mutations of a microsatellite (G)8 of the Bax gene. PMID- 11261834 TI - An in vitro model of human breast carcinogenesis: epigenetic aspects. AB - A review is given of 12 years of research on a human breast epithelial cell line, HMT-3522, which has undergone malignant transformation in vitro without being exposed to known carcinogenic agents. Epigenetic aspects of the malignant transformation have been considered and the results have been viewed in a clinical context. It has been concluded that the history and characteristics of the cell line resembles the comedocarcinoma of the human breast. It is hypothesized that progression from benign lesion to comedo in situ carcinoma and invasive carcinoma occurs if low levels of epidermal growth factor are prevailing in the microenvironment. Our data also suggest that breast cancer developed under high epidermal growth factor receptor activity is estrogen receptor negative, while suppression of epidermal growth factor receptor activity promotes estrogen responsive breast cancer. PMID- 11261835 TI - Chronic in vitro exposure to 3'-azido-2', 3'-dideoxythymidine induces senescence and apoptosis and reduces tumorigenicity of metastatic mouse mammary tumor cells. AB - Normal cells in culture divide a certain amount of times and undergo a process termed replicative senescence. Telomere loss is thought to control entry into senescence. Activation of telomerase in tumors bypasses cellular senescence and is thus a requirement for tumor progression. We reported previously the preferential incorporation of 3'-azido-2', 3'-dideoxythymidine (AZT) in telomeric sequences of immortalized cells in culture. In this work, we have investigated the effects of chronic in vitro AZT exposure on F3II mouse mammary carcinoma cells. We demonstrate, for the first time, that AZT-treated tumor cells have a reduced tumorigenicity in syngeneic BALB/c mice. Tumor incidence was reduced and survival was prolonged in animals inoculated with AZT-treated cells when comparing with control counterparts. The number and size of spontaneous metastases were also decreased in animals inoculated with AZT-treated cells. In addition, we present evidence of morphological and biochemical signs of senescence, as shown by the staining for senescence associated beta-galactosidase activity, and induction of programmed cell death, as demonstrated by an increase of caspase-3 activity, in tumor cells exposed to AZT. These data indicate that chronic exposure of mammary carcinoma cells to AZT may be sufficient to induce a senescent phenotype and to reduce tumorigenicity. PMID- 11261836 TI - Synthesis and reactivity of the monosaccharide esters of amino acids as models of teichoic acid fragment. AB - The increasing prevalence of sepsis from gram-positive bacterial pathogens necessitates further evaluation of the basic assumptions about the molecular pathogenesis of septic shock. Since diverse physiological functions of gram positive bacteria are controlled by the degree of esterification of teichoic acids with D-alanine, we examined the reactivity of monosaccharide esters in which anomerically free or protected D-glucose is linked through its C-6 hydroxy group to either phenylalanyl or tyrosyl residues as models for teichoic acid fragment. We show that the attached sugar moiety induces activation of the amino acid residue. Due to the enhanced reactivity of the NH2 group in the monosaccharide esters studied, the formation of products generated by intramolecular and intermolecular glycation reactions is accelerated resulting in heterogeneous mixture of compounds. These findings suggest that, if similar adducts are formed by glycation of D-alanine in teichoic acid of gram-positive bacteria, they should be examined as potential bioactive ligands or chemical message for infection. PMID- 11261837 TI - Modeling ganglioside headgroups by conformational analysis and molecular dynamics. AB - The conformations and dynamics of gangliosides GM1, GM2, 6'-GM2 and GM4 have been studied by computational means, and the results compared to NMR data. Unconstrained conformational searches were run using the AMBER* force field augmented by MNDO derived parameters for the Neu5Ac anomeric torsion, the GB/SA water solvation model, and the MC/EM alogorithm; extended (10-12 ns) dynamic simulations in GB/SA water were performed with the MC/SD protocol, and the stored structures were minimized. The overall mobility of the Neu5Ac alpha2,3Gal linkage and the position of its minimum energy conformation have been shown to depend mainly on the presence or the absence of a GalNAc residue at the adjacent position. The best quantitative agreement with the available NOE data was achieved after minimization of the structures stored during the MC/SD dynamic runs. The latter protocol appears to reproduce satisfactorily the available experimental data, and can be used with confidence to build three-dimensional models of ganglioside headgroups. PMID- 11261839 TI - Differentiation-dependent glycosylation of gp190, an oncofetal crypt cell antigen expressed by Caco-2 cells. AB - gp190 is a glycoprotein expressed on the cell surface of several human colon carcinoma cells in culture, on epithelial cells of fetal colon, but not on the normal mucosa of adult colon; thus it is referred to as an oncofetal crypt cell antigen. We report the characterisation of O-linked glycans carried by gp190 synthesised by [3H]glucosamine-labelled Caco-2 cells at the confluence (undifferentiated cells) and at three weeks of postconfluence (differentiated cells). By using a specific monoclonal antibody, gp190 was isolated and analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The mobility of gp190 from differentiated cells was found to be lower than that from undifferentiated cells, suggesting a more extensive glycosylation process in the former glycoprotein. The major results of the glycan characterisation have been as follows: (i) gp190 carries mainly, if not exclusively, O-linked glycans with the core-2 structure; (ii) the elongation with N-acetyllactosamine units of the Gal beta1,4GlcNAc beta1,6(Gal beta1,3)GalNAc tetrasaccharide predominates in gp190 synthesised by differentiated cells, whereas the direct alpha2,3sialylation of the tetrasaccharide is prevalent in gp190 synthesised by undifferentiated cells. The increment in the core-2 beta1,6GlcNAc-transferase activity under the Caco-2 differentiation process may be relevant in producing the larger occurrence of polylactosaminoglycans in gp190 from differentiated cells. Since no change in the activity of the alpha2,3sialyltransferases upon cell differentiation was observed, we suggest that the lower alpha2,3sialylation in gp190 synthesised by polarised cells might be due to a changed transit-rate through the distal Golgi apparatus. PMID- 11261838 TI - Decrease in cell surface sialic acid in etoposide-treated Jurkat cells and the role of cell surface sialidase. AB - The present study investigated the mechanism underlying alterations of cell surface sugar chains of Jurkat cells by inducing apoptosis with etoposide, an inhibitor of topoisomerase II. Within 3 h of etoposide treatment, flowcytometric analysis revealed a decrease in Maackia amurensis agglutinin recognized alpha2,3 linked sialic acid moieties and an increase in Ricinus communis agglutinin recognized galactose. The results suggested that asialo-sugar chains on glycoconjugates were rapidly induced on the etoposide-treated cell surface. To clarify the desialylation mechanism, we studied alpha2,3-sialyltransferase mRNA expression and the activity of sialidase on the cell surface during etoposide induced apoptosis. The expression of hST3Gal III and hST3Gal IV mRNAs were down regulated and sialidase activity on the cell surface increased threefold within 2 h of etoposide treatment. Moreover, the decrease in alpha2,3-linked sialic acid levels was significantly suppressed in the presence of 2,3-dehydro-2-deoxy-N acetylneuraminic acid, an inhibitor of sialidase. These results suggested that activation or exposure of sialidase on the cell surface was induced by etoposide treatment and was the main cause of the decrease in sialic acids. PMID- 11261840 TI - Modulation of the basal activity of phosphatidylinositol-3-kinase/protein kinase B signaling pathway in human hepatocarcinoma cells. AB - The modulation of GnT-V activity by signaling molecules in PI-3-K/PKB pathway in human hepatocarcinoma cell line 7721 was studied. GnT-V activity was determined after the transfection of sense or antisense cDNA of PKB into the cells, as well as the addition of activators, specific inhibitors, and the antibodies to the enzyme assay system or culture medium. It was found that the basal activity of GnT-V was up regulated by the sense and down regulated by the antisense cDNA of PKB transfected into 7721 cells. GnT-V was activated by PIP2, PIP3 or GTPgamma[S] added to the assay system, and the activation of PIP2 or GTPgamma[S] was abolished by LY2940002, a specific inhibitor of PI-3-K, but the activation of PIP3 was not attenuated by LY2940002. In addition, GnT-V activity in cultured parental or H-ras transfected cells was inhibited by the antibody against PKB or PI-3-K. These findings demonstrated the involvement of PI-3-K/PKB signaling pathway in the regulation of GnT-V. Moreover, ET18-OCH3, an inhibitor of Raf translocation and PI-PLC enzyme, which produces the activator of PKC, as well as the antibodies against Raf-1 or MEK also inhibited GnT-V activity in the parental and H-ras transfected cells. The inhibitory rates, however, were less in the transfected cells than those in the parental cells. These results reveal that in parental and H-ras transfected 7721 cells, the basal activity of GnT-V is also regulated by the Ras/Raf-1/MEK/MAPK cascade in addition to PI-3-K/PKB signaling pathway. The significance of these two pathways in the regulation of GnT-V and their relations to the activation of PKC previously reported by our laboratory (Ju TZ et al., 1995 Glyconjugate J 12, 767-772) was discussed. PMID- 11261841 TI - Analysis of agalacto-IgG in rheumatoid arthritis using surface plasmon resonance. AB - It is well established that IgG from rheumatoid arthritis (RA) patients are less galactosylated than IgG from normal individuals. Determination of agalacto-IgG may therefore aid in diagnosis and treatment of RA. The decrease in galactosylation of IgG leads to an increase in terminal N-acetylglucosamine residues, which can be detected using a specific lectin from Psathyrella velutina. In the present study IgG from RA and control serum was purified using affinity chromatography. The samples were then, after reduction, analyzed on a BIOCORE 2000 system with immobilized Psathyrella velutina lectin. Using this technique it was possible to discriminate between IgG from RA patients and IgG from control individuals with respect to its content of IgG with terminal N acetylglucosamine. The affinity biosensor technique makes it possible to detect binding without labeling or using secondary antibodies. PMID- 11261843 TI - Identification of lectin isoforms in juvenile freshwater prawns Macrobrachium rosenbergii (DeMan, 1879). AB - From the serum of juvenile freshwater prawns, we isolated by affinity chromatography on glutaraldehyde-fixed rat erythrocytes stroma, immobilized in Sephadex G-25, a sialic acid specific lectin of 9.6 kDa per subunit. Comparative analysis against adult organisms purified lectin, by chromatofocusing, showed that the lectin from juvenile specimens is composed by four main isoforms with a pI of 4.2, 4.6, 5.1, and 5.6, whereas the lectin from adults is eluted at pH 4.2. The amino acid composition of the lectin obtained from adult and juvenile stages suggest identity, but the compositions are not identical since a higher content of carbohydrates was found in the lectin from younger organisms. The freshwater prawn lectin showed specificity toward N-acetylated amino sugar residues such as GlcNAc, GalNAc, Neu5Ac and Neu5,9Ac; but in juvenile organisms the lectin showed three times less hemagglutinating activity than the lectin from adults. Both lectins agglutinated rat, rabbit and chicken erythrocytes, indicating that Neu5,9Ac in specific O-glycosydically linked glycans seems to be relevant for the interaction of M. rosenbergii lectins with their specific cellular receptor. Our results suggest that the physicochemical characteristics of the lectin from the freshwater prawn are regulated through maturation. PMID- 11261844 TI - Genomic DNA hybridizes with the same rate constant on the QCM biosensor as in homogeneous solution. AB - Hybridization rates of sheared, genomic E. coli DNA in 0.14 M, pH 6.7 phosphate buffer at 65 degrees C were determined by: (1) observing the rate of absorbance decrease at 260 nm due to self-hybridization in solution; and (2) measurement of the rate of mass increase caused by hybridization between DNA in solution and DNA photografted to polystyrene. The latter measurement was done using a quartz crystal microbalance (QCM). In both the spectrophotometric and QCM experiments the probe was identical to the target, as both were taken from the same sample of sheared E. coli DNA. In the QCM measurements, viscoelastic effects were made negligible by drying the biopolymer layer on the QCM's surface before taking the frequency readings. Our purpose was to explore the effect of immobilizing DNA on its hybridization rate constant. A second-order constant of 2.32 +/- 0.09 x 10( 6) ml microg(-1) s(-1), n = 14, for hybridization in solution was obtained spectrophotometrically, while the QCM experiment gave a constant of 2.2 +/- 0.3 x 10(-6) ml microg(-1) s(-1), n = 6. These values are not statistically different. The reaction half-lives for the spectrophotometric and QCM experiments were 6.5 h and 13 min, respectively. The shorter half-life on the QCM can be explained solely by the much greater reactant concentration in the QCM experiment. About 25% of the DNA was inactivated by the attachment reaction. After correcting for this, the surface-attached DNA hybridized with the same rate constant as DNA free in solution. Therefore, it is concluded that, in these specific experiments with genomic DNA, the immobilized regions must have been short compared to the length of the molecules. The data demonstrate the high hybridization rate obtainable when nucleic acids are hybridized in a thin-film, micro-volume reaction on a non porous surface. PMID- 11261842 TI - Expression of H type 1 antigen of ABO histo-blood group in normal colon and aberrant expressions of H type 2 and H type 3/4 antigens in colon cancer. AB - We have immunohistochemically examined the distribution of the H antigens of type 1, type 2 and type 3/4 chains of the ABO(H) histo-blood group system in human normal colon and in colon cancer using three monoclonal antibodies specific for each of the H type 1/2, H type 2, and the H type 3/4 chain. We unexpectedly found that mucosa of the normal colon from secretors but not that from nonsecretors expressed only H type 1 and did not express H type 2 or H type 3/4. The H type 1 was expressed in goblet cells. Positive goblet cells expressing H type 1 were decreased in number progressively from the proximal colon to the rectum. In tumors, 4 (57%) of 7 cancer tissues of the proximal colon from secretors expressed no H type 1, whereas all 8 cancer tissues of the distal colon from secretors expressed H type 1. The aberrant expressions of H type 2 and H type 3/4 (47 and 67%, respectively) were found in cancer tissues from both the proximal and the distal colon. Tumors from nonsecretors did not express any H antigens. Our results suggested that the expression of H type 1 in the normal colon and the aberrant expressions of H type 2 and H type 3/4 in colon cancer tissues were regulated by FUT2-encoded Se type alpha(1,2)fucosyltransferase. However, UEA-I positive substance(s) rather than H type 2 were uniquely expressed throughout the normal colon and in colon cancers from both secretors and nonsecretors. PMID- 11261845 TI - A fiber-optic lactate sensor based on bacterial cytoplasmic membranes. AB - A new type of fiber-optic biosensor based on bacterial cytoplasmic membranes (CPM) as the biological recognition element and an oxygen sensitive dye layer as the transducer is described for the detection of lactate. CPMs from bacteria with an induced lactate oxidase system are adsorbed onto a cellulose disk. The disk is fixed mechanically over an oxygen sensitive siloxane layer on the distal end of an optical fiber. This system detects lactate with no interference from glucose, fructose or glutamic acid. PMID- 11261846 TI - Characterization for didodecyldimethylammonium bromide liquid crystal film entrapping catalase with enhanced direct electron transfer rate. AB - Direct electrochemistry for catalase (CAT) embedded in the liquid crystal film of didodecyldimethylammonium bromide (DDAB) was investigated at pyrolytic graphite (PG) electrode by voltammetric methods. The reduction reaction involved the redox couple in CAT, i.e. FeIII/FeII couple. The electron transfer between the incorporated CAT and PG electrode was found to be greatly enhanced by DDAB. The heterogeneous electron transfer rate constant k(s) was fitted as 3.0 +/- 0.4 s( 1) using the nonlinear regression analysis of the square wave voltammograms at a series of pulse heights. The pH dependence of the formal potential for CAT in DDAB film was 57 mV/pH, which suggested one-proton-transfer together with a one electron reaction. Visible absorption and reflectance-absorbance infrared (RAIR) spectra inferred the similar heme environment of CAT in DDAB film to its native status. Circular dichroism (CD) results indicated DDAB affected slightly on the second structure of CAT. PMID- 11261847 TI - Electrochemical biosensors: recommended definitions and classification. AB - Two Divisions of the International Union of Pure and Applied Chemistry (IUPAC), namely Physical Chemistry (Commission 1.7 on Biophysical Chemistry formerly Steering Committee on Biophysical Chemistry) and Analytical Chemistry (Commission V.5 on Electroanalytical Chemistry) have prepared recommendations on the definition, classification and nomenclature related to electrochemical biosensors: these recommendations could, in the future, be extended to other types of biosensors. An electrochemical biosensor is a self-contained integrated device, which is capable of providing specific quantitative or semi-quantitative analytical information using a biological recognition element (biochemical receptor) which is retained in direct spatial contact with an electrochemical transduction element. Because of their ability to be repeatedly calibrated, we recommend that a biosensor should be clearly distinguished from a bioanalytical system, which requires additional processing steps, such as reagent addition. A device that is both disposable after one measurement, i.e. single use, and unable to monitor the analyte concentration continuously or after rapid and reproducible regeneration, should be designated a single use biosensor. Biosensors may be classified according to the biological specificity-conferring mechanism or, alternatively, to the mode of physico-chemical signal transduction. The biological recognition element may be based on a chemical reaction catalysed by, or on an equilibrium reaction with macromolecules that have been isolated, engineered or present in their original biological environment. In the latter cases. equilibrium is generally reached and there is no further, if any, net consumption of analyte(s) by the immobilized biocomplexing agent incorporated into the sensor. Biosensors may be further classified according to the analytes or reactions that they monitor: direct monitoring of analyte concentration or of reactions producing or consuming such analytes; alternatively, an indirect monitoring of inhibitor or activator of the biological recognition element (biochemical receptor) may be achieved. A rapid proliferation of biosensors and their diversity has led to a lack of rigour in defining their performance criteria. Although each biosensor can only truly be evaluated for a particular application, it is still useful to examine how standard protocols for performance criteria may be defined in accordance with standard IUPAC protocols or definitions. These criteria are recommended for authors. referees and educators and include calibration characteristics (sensitivity, operational and linear concentration range, detection and quantitative determination limits), selectivity, steady-state and transient response times, sample throughput, reproducibility, stability and lifetime. PMID- 11261848 TI - The effect of UV irradiation on uracil thin layer measured by optical waveguide lightmode spectroscopy. AB - The polycrystalline uracil thin-layer dosimeter is a well-established method to monitor the biological effects of the environmental ultraviolet (UV) radiation. It is based on the optical density (OD) decrease of the uracil layer in the UV absorption band due to photodimerization of the crystal caused by UV irradiation. In the present study, we report measurements made with optical waveguide lightmode spectroscopy (OWLS) to characterize the changes in the optogeometrical parameters of the uracil layer caused by an artificial UV source. It is shown that UV irradiation causes a decrease in the refractive index and an increase of the optical anisotropy. The determined kinetic parameters of the UV dose-sensor response curves correlate well with results of OD measurements, but the sensitivity of OWLS is about ten times higher. The results show that OWLS is capable of analyzing the UV response of the uracil layer and opens the way for dosimetrical applications. PMID- 11261849 TI - Characterisation and optimisation of AC conductimetric biosensors. AB - Urease, immobilised on interdigitated gold electrodes, is employed as a model enzyme for characterisation and optimisation of a.c. conductimetric sensors. The sensors' response is measured over a frequency range of 20 Hz to 300 kHz and an optimum operating frequency established. The activity of the urease, both in solution and immobilised states, is investigated and Km values obtained. The effect of method of immobilisation and enzyme loading on the sensors' performance are studied and urease electrodes are characterised as a function of temperature, pH and electrolyte concentration. An important finding, particularly for conductimetric sensors designed for clinical use, is that proper consideration of the effects of electrode polarisation must be taken into account in order to maintain high sensor sensitivity at physiological electrolyte concentrations. Measurements of urea concentration in untreated serum are described. PMID- 11261850 TI - The electric charge of pigment granules in pigment cells. AB - Black pigment cells called melanophores change colour in response to environmental changes and have lately been studied as promising biosensors. To further elucidate the intracellular processes involved in the colour changes of these cells, and to find optimal biosensing principles, the electric charge of intracellular pigment granules, melanosomes, has been determined in vitro by electrophoresis. Melanosomes from the two extreme states in the cell colour change (aggregated and dispersed melanosomes) were measured. The charge was found to be -1.5 x 10(-16) and -1.7 x 10(-16) C, aggregated and dispersed melanosomes, respectively, without significant difference between the two conditions. This charge is of the same order of magnitude as the one of 1000 electrons. The origin of the melanosome charge, and the use of these findings in new biosensor principles, is discussed. PMID- 11261851 TI - Compact integrated optical sensor system. AB - A compact integrated optical sensor system for a large variety of different (bio ) chemical applications using replicated sensor chips is described. Features of the refractometric system to be emphasized for practical applications include a high-resolution window that can be positioned within a wide measuring range, an in situ chip testing and characterization procedure, and on-chip referencing. As an application example, experimental results on refractometric measurements as well as on the suppression of non-specific binding are given. PMID- 11261853 TI - Dynamic model of amperometric biosensors. Characterisation of glucose biosensor output. AB - An integrated model for the characterisation of the output signal course of oxidase-bound amperometric biosensors is presented and evaluated in the case of glucose biosensors. This model integrates two earlier proposed models, the model of oxygen transducer-based biosensors, allowing the prediction of steady state parameters from the transient response and the dynamic signal lag model, characterising the electrochemical diffusion-limited sensors. The integrated model allows the characterisation of the whole biosensor signal output, originating from the output curve itself with errors less than 3% and no need to determine the system's geometrical, diffusion and partition parameters. PMID- 11261852 TI - A facile approach to immobilize protein for biosensor: self-assembled supported bilayer lipid membranes on glassy carbon electrode. AB - A kind of solid substrate, glassy carbon (GC) electrode, was selected to support self-assembled lipid layer membranes. On the surface of GC electrode, we made layers of dimyristoylphosphatidylcholine (DMPG, a kind of lipid). From electrochemical impedance experiments, we demonstrated that the lipid layers on the GC electrode were bilayer lipid membranes. We immobilized horseradish peroxidase (HRP) into the supported bilayer lipid membranes (s-BLM) to develop a kind of mediator-free biosensor for H2O2. The biosensor exhibited fine electrochemical response, stability and reproducibility due to the presence of the s-BLM. As a model of biological membrane, s-BLM could supply a biological environment for enzyme and maintain its activity. So s-BLM is an ideal choice to immobilize enzyme for constructing the mediator-free biosensor based on GC electrode. PMID- 11261854 TI - Amperometric flow-injection determination of sucrose with a mediated tri-enzyme electrode based on sucrose phosphorylase and electrocatalytic oxidation of NADH. AB - A new sucrose electrode is described for the determination of sucrose without interference from glucose or fructose. The sucrose electrode is based on the tri enzymatic system of sucrose phosphorylase, phosphoglucomutase and glucose-6 phosphate 1-dehydrogenase, where NAD(P)H is produced from the last enzymatic reaction and recycled into NAD(P)+ through its electrocatalytic oxidation by Os(4,4'-dimethyl-2,2-bypyridine)2(1,10-phenanthroline-5,6-dione). The electrodes were optimised with respect to the various construction parameters and carrier composition in a FIA system and their response as a function of the pH and flow rate was examined. The electrodes were suitable for operation in a FIA system and the analysis of real samples showed good agreement with the reference method. Typical optimised electrodes showed detection limits of 1 mM sucrose, response time of 5 min, sensitivity 1.010 nA mM(-1), and current density of 8.38 microA cm(-2), using 200 mM PIPES pH 7.25 with 10 mM phosphate and 5 mM MgCl2 as carrier. PMID- 11261855 TI - The influence of external environment fluctuations on the signal formation of microbiosensors. AB - This part of theoretical analysis describes the fluctuations of output signal of microbiosensors when the number of accessible molecular recognition elements (enzymes, receptors, antibodies, etc.) fluctuated under external environmental influences. The mean electric current, dispersion correlating function, as well as spectral density of output current fluctuation are analyzed, and it is shown that a comparison of theoretically calculated mean current and correlation function with experimental data allow a determination of the kinetic parameters of substrate binding reaction with the molecular recognition element of biosensor. PMID- 11261856 TI - On applicability of laccase as label in the mediated and mediatorless electroimmunoassay: effect of distance on the direct electron transfer between laccase and electrode. AB - Applicability of laccase as enzyme-label has been investigated. It was shown that the property of laccase to catalyze the oxygen electroreduction at an electrode allows to develop a mediatorless and pseudoreagentless electro-enzyme-immunoassay (EEIA). In this case the electrode acts as an electron-donor substrate. When the bioelectrocatalytic reaction takes place, some electric charge is collected on the electrode. A method of determination of the electrode charge as well as the concentration of oxidized form of the mediator at the electrode surface has been elaborated. For this aim a technique of the measurement of current-surge was employed. Human immunoglobulin G and insulin were taken as model in this investigation. A back titration schemes without any mediator and in the presence of o-carboxybenzoylferrocene as a mediator was applied. The antibody carbon-black and the antigen glassy-carbon electrodes were used. The limits of detection were found to be 0.3 and 1.6 nM, respectively. The advantage of the mediatorless assay is that the charge leakage is imperceptible by open circuit for a long time and the accumulation of the charge occurs linearly with time. The charge accumulation for a long time allows to diminish the limit of detection. However, there is a limitation of the method. The direct electron transfer slows down with increasing the distance between the enzyme molecule and the electrode surface. This effect reduces the sensitivity of the method. The decrease of the electron transfer rate with distance has been estimated. Monolayer of hemoglobin dividing the laccase molecule from the electrode surface decreases the rate by four times. The electron transfer rate for the antibody electrode with associated antigen-laccase conjugate is less than that for the analogous electrode, covered with monolayer of covalently attached laccase, by 210 times. The current-surge peak was expected to decrease with distance by an equation of the form I = I0 exp[-r/r0]. The parameter r0 is equal to 2.2 +/- 0.8 nm. The possibility of the sensitivity increase in the mediatorless mode by 'wiring' through the multilayer film of immunoproteins immobilized on the electrode is discussed. PMID- 11261857 TI - Microgravimetric DNA sensor based on quartz crystal microbalance: comparison of oligonucleotide immobilization methods and the application in genetic diagnosis. AB - We report on the study of immobilization DNA probes onto quartz crystal oscillators by self-assembly technique to form variety types of mono- and multi layered sensing films towards the realization of DNA diagnostic devices. A 18-mer DNA probe complementary to the site of genetic beta-thalassaemia mutations was immobilized on the electrodes of QCM by covalent bonding or electrostatic adsorption on polyelectrolyte films to form mono- or multi-layered sensing films by self-assembled process. Hybridization was induced by exposure of the QCMs immobilized with DNA probe to a test solution containing the target nucleic acid sequences. The kinetics of DNA probe immobilization and hybridization with the fabricated DNA sensors were studied via in-situ frequency changes. The characteristics of QCM sensors containing mono- or multi-layered DNA probe constructed by direct chemical bonding, avidin-biotin interaction or electrostatic adsorption on polyelectrolyte films were compared. Results indicated that the DNA sensing films fabricated by immobilization of biotinylated DNA probe to avidin provide fast sensor response and high hybridization efficiencies. The effects of ionic strength of the buffer solution and the concentration of target nucleic acid used in hybridization were also studied. The fabricated DNA biosensor was used to detect a set of real samples. We conclude that the microgravimetric DNA sensor with its direct detection of amplified products provide a rapid, low cost and convenient diagnostic method for genetic disease. PMID- 11261858 TI - A methylene blue-mediated enzyme electrode for the determination of trace mercury(II), mercury(I), methylmercury, and mercury-glutathione complex. AB - A methylene blue-mediated enzyme biosensor has been developed for the detection of inhibitors including mercury(II), mercury(I), methylmercury, and mercury glutathione complex. The inhibition to horseradish peroxidase was apparently reversible and noncompetitive in the presence of HgCl2 in less than 8 s and irreversibly inactivated when incubated with different concentrations of HgCl2 for 1-8 min. The binding site of horseradish peroxidase with HgCl2 probably was a cysteine residue SH. Mercury compounds can be assayed amperometrically with the detection limits 0.1 ng ml(-1) Hg for HgCl2 and methylmercury, 0.2 ng ml(-1) Hg for Hg2(NO3)2 and 1.7 ng ml(-1) Hg for mercury glutathione complex. Inactivation of the immobilized horseradish peroxidase was displayed in the AFM images of the enzyme membranes. PMID- 11261859 TI - A surface plasmon resonance array biosensor based on spectroscopic imaging. AB - We have developed a multi-element transduction system which combines conventional SPR spectroscopy with one-dimensional SPR microscopy to create an effective platform for monitoring binding events on macro- or micro-patterned receptor arrays created on disposable sensor chips. This creates an effective platform for monitoring simultaneous binding events on each of the regions patterned with the receptors. This system has been specifically designed with commercially available components to allow relatively easy duplication. Furthermore, this system can use a proven, simple method to compensate for changes in the bulk index of refraction of the solution containing the analytes due to changes in temperature or solute concentration with simple modifications to the sensor chips alone. Preliminary results demonstrate how this system can be used to monitor several independent biospecific binding events simultaneously. PMID- 11261860 TI - Ergogenic aids: powders, pills and potions to enhance performance. PMID- 11261861 TI - Evaluating children for possible sexual abuse. PMID- 11261862 TI - Toxic cascades: a comprehensive way to think about medical errors. PMID- 11261863 TI - Pharmacotherapy for nonmalignant pain. PMID- 11261864 TI - Nutritional supplements and treatment of osteoarthritis. PMID- 11261865 TI - Evaluating the child for sexual abuse. AB - Child victims of sexual abuse may present with physical findings that can include anogenital problems, enuresis or encopresis. Behavioral changes may involve sexual acting out, aggression, depression, eating disturbances and regression. Because the examination findings of most child victims of sexual abuse are within normal limits or are nonspecific, the child's statements are extremely important. The child's history as obtained by the physician may be admitted as evidence in court trials; therefore, complete documentation of questions and answers is critical. A careful history should be obtained and a thorough physical examination should be performed with documentation of all findings. When examining the child's genitalia, it is important that the physician be familiar with normal variants, non-specific changes and diagnostic signs of sexual abuse. Judicious use of laboratory tests, along with appropriate therapy, should be individually tailored. Forensic evidence collection is indicated in certain cases. Referral for psychologic services is important because victims of abuse are more likely to have depression, anxiety disorders, behavioral problems and post-traumatic stress disorder. PMID- 11261866 TI - Osteoporosis: part I. Evaluation and assessment. AB - Osteoporosis afflicts 75 million persons in the United States, Europe and Japan and results in more than 1.3 million fractures annually in the United States. Because osteoporosis is usually asymptomatic until a fracture occurs, family physicians must identify the appropriate timing and methods for screening those at risk. Prevention is the most important step, and women of all ages should be encouraged to take 1,000 to 1,500 mg of supplemental calcium daily, participate in regular weight-bearing exercise, avoid medications known to compromise bone density, institute hormone replacement therapy at menopause unless contraindicated and avoid tobacco and excessive alcohol intake. All postmenopausal women who present with fractures as well as younger women who have risk factors should be evaluated for the disease. Physicians should recommend bone mineral density testing to younger women at risk and postmenopausal women younger than 65 years who have risk factors for osteoporosis other than being postmenopausal. Bone mineral density testing should be recommended to all women 65 years and older regardless of additional risk factors. Bone mineral density screening should be used as an adjunct to clinical judgment only if the results would influence the choice of therapy or convince the patient to take appropriate preventive measures. PMID- 11261867 TI - Ergogenic aids: counseling the athlete. AB - Numerous ergogenic aids that claim to enhance sports performance are used by amateur and professional athletes. Approximately 50 percent of the general population have reported taking some form of dietary supplements, while 76 to 100 percent of athletes in some sports are reported to use them. Physicians can evaluate these products by examining four factors (method of action, available research, adverse effects, legality) that will help them counsel patients. Common ergogenic aids include anabolic steroids, which increase muscle mass. These illegal supplements are associated with a number of serious adverse effects, some irreversible. Creatine modestly improves athletic performance and appears to be relatively safe. Dehydroepiandrosterone and androstenedione do not improve athletic performance but apparently have similar adverse effects as testosterone and are also banned by some sports organizations. Caffeine has mild benefits and side effects and is banned above certain levels. Products that combine caffeine with other stimulants (e.g., ephedrine) have been linked to fatal events. Protein and carbohydrate supplementation provides modest benefits with no major adverse effects. PMID- 11261868 TI - Otitis externa: a practical guide to treatment and prevention. AB - Otitis externa is most commonly caused by infection (usually bacterial, although occasionally fungal), but it may also be associated with a variety of noninfectious systemic or local dermatologic processes. The most characteristic symptom is discomfort that is limited to the external auditory canal, while the most characteristic signs are erythema and swelling of the canal with variable discharge. Excessive moisture and trauma, both of which impair the canal's natural defenses, are the two most common precipitants of otitis externa, and avoidance of these precipitants is the cornerstone of prevention. Thorough cleansing of the canal is essential for diagnosis and treatment, but flushing should be avoided. Acidification with a topical solution of 2 percent acetic acid combined with hydrocortisone for inflammation is effective treatment in most cases and, when used after exposure to moisture, is an excellent prophylactic. Other prophylactic measures such as drying the ears with a hair dryer and avoiding manipulation of the external auditory canal may help prevent recurrence. PMID- 11261869 TI - Photo quiz. Suspicious ulcers in the colon. PMID- 11261870 TI - ATS adopts diagnostic standards for tuberculosis. American Thoracic Society. PMID- 11261871 TI - A major medical error. PMID- 11261873 TI - Poly(ethylene glycol): protein-repulsive or albumin-compatible? AB - In the literature, many papers deal with the behavior of proteins in aqueous media in the presence of poly(ethylene glycol) (PEG) molecules or poly(ethylene oxide) (PEO) segments, physically adsorbed onto, or covalently attached to, macromolecules or to solid surfaces. In particular, it is well known that PEO segments make foreign materials stealthy, i.e. they are much less detected by the immune system either through humoral reactions or, at the cell level, through opsonins. Revisiting the literature led us to challenge the largely accepted opinion that the decreased recognition of PEO segment-bearing foreign macromolecules and particles by the mononuclear phagocyte system is primarily the consequence of the repulsion of all blood proteins by PEG segments through the excluded volume effect. This challenge is based on the finding that albumin and PEG are compatible in phosphate-buffered saline at room temperature and at concentrations comparable to those measured by others on the surface of PEO segment-bearing species, whereas fibrinogen and PEG phase-separated and were incompatible despite the much lower concentration of the latter protein. According to literature and to these observations, it is proposed that the stealth effect induced by PEO segments is primarily due to the compatibility between PEO segments of intermediate molar mass and albumin, thus rendering PEO bearing macromolecules or surfaces to look like native albumin. Under such conditions, the hospitality offered by PEG macromolecules or PEO segments to albumin, the dominant plasma protein, results in a 'chameleon' effect that prevents the activation of other PEG-compatible or -incompatible plasma proteins or cells involved in foreign body recognition and elimination. PEG with molar masses > or = 8000 did not accommodate albumin in agreement with the excluded volume phenomenon. PMID- 11261872 TI - Professor Bamford's research in the field of biomaterials. AB - Professor Bamford was regarded by many as the greatest British polymer chemist of the twentieth century and when Bam passed away in November 1999 tribute was quite rightly made to his considerable achievements in the field of polymer science. The aim of this paper is to highlight Bam's contribution to biomaterials research that occupied his attention for over 15 years after his official retirement. In particular a review of the synthetic methods employed by Bam for the modification of polymers to improve haemocompatibility and to function as affinity separation membranes for protein purification is presented. PMID- 11261874 TI - Insoluble ionic complexes of polyacrylic acid with a cationic drug for use as a mucoadhesive, ophthalmic drug delivery system. AB - We have developed a new mucoadhesive drug delivery formulation based on an ionic complex of partially neutralized poly(acrylic acid) (PAA) and a highly potent beta blocker drug, levobetaxolol x hydrochloride (LB x HCl), for use in the treatment of glaucoma. PAA was neutralized with sodium hydroxide to varying degrees of neutralization. Aqueous solutions containing concentrations of LB x HCl equivalent to the degree of PAA neutralization were added to the PAA solutions and formed insoluble complexes, which were isolated. The complex formation was followed by turbidimetric titration, and the complexes were characterized by IR and 1H NMR spectroscopy. Complexes were prepared with varying degrees of drug loading, such that the same PAA chain would have free -COOH groups for mucoadhesion along with ionic complexes of LB x H+ with COO- groups. Thin films of the complexes dissociated to release the drug by ion exchange with synthetic tear fluid. The films shrunk continuously during release of the drug and dissolved completely in 1 h. Solid inserts of these films could be useful as a mucoadhesive ophthalmic drug delivery system. PMID- 11261875 TI - Preparation and characterization of poly(ethylene glycol) hydrogels cross-linked by hydrolyzable polyrotaxane. AB - PEG hydrogels cross-linked by a hydrolyzable polyrotaxane were prepared and their hydrolytic erosion characterized in terms of supramolecular dissociation of the polyrotaxane. The hydrolyzable polyrotaxane, in which many alpha-cyclodextrins (alpha-CDs) are threaded onto a poly(ethylene glycol) (PEG) chain capped with L phenylalanine via ester linkages, was used as a multifunctional cross-linker: the PEG network was covalently bound to hydroxyl groups of alpha-CDs in the polyrotaxane. The contact angle and water content of the hydrogels were varied with the polyrotaxane content in the feed. In vitro hydrolysis study revealed that the time to reach complete gel erosion was shortened by increasing the polyrotaxane content in the feed in relation to the decreased number of chemical cross-links between PEG and alpha-CDs in the polyrotaxane. The hydrogel degradation in a physiological condition was found to be followed by bulk mechanism. These findings suggest that changing the preparative conditions such as polyrotaxane content will make it possible to control programmed gel erosion for tissue engineering. PMID- 11261876 TI - Poly(rac-lactide) nanocapsules containing diclofenac: protection against muscular damage in rats. AB - The aim of this work was to determine whether encapsulation of a non steroidal antiinflammatory agent within nanocapsules could reduce local toxicity after intramuscular injection. Diclofenac-loaded nanocapsules were prepared by deposition of poly(rac-lactic acid) polymer, and administered intramuscularly to male Wistar rats. Plasma creatine phosphokinase (CPK) activity and histological examination were used to assess local tissue damage. Following a single intramuscular injection of diclofenac (0.8 mg), CPK activity was shown to depend on both the type of dosage form and, in the case of nanocapsules, on the chemical nature of the central oily core. Lower CPK activity was observed with nanocapsules prepared from Miglyol 810, a caprylic/capric triglyceride, while nanocapsules prepared from benzyl benzoate, either empty or containing diclofenac, exhibited the same CPK activity as the drug solution. Histopathological examination performed three days after administration of free diclofenac or nanocapsules containing diclofenac prepared from Miglyol 810 revealed that a much more intense inflammation was obtained with the solution than with nanocapsules. In conclusion, when appropriately formulated, nanocapsules can considerably reduce the muscular damage caused by diclofenac. PMID- 11261877 TI - Synergistic effect of gelatin microspheres incorporating TGF-beta1 and a physical barrier for fibrous tissue infiltration on skull bone formation. AB - The objective of this study is to examine whether or not bone formation at a skull bone defect induced by gelatin microspheres incorporating transforming growth factor (TGF)-beta1 is promoted by prevention of fibrous tissues into the defect. The 6-mm diameter bone defect of rabbit skulls was applied with gelatin microspheres incorporating TGF-beta1 or free TGF-beta1 and physically covered by a barrier membrane. When the bone formation at the defect was assessed 6 weeks postoperatively, combinational application of gelatin microspheres incorporating 0.1 microg of TGF-beta1 with the barrier membrane induced bone formation at the skull defect, in marked contrast to that of 0.1 microg of free TGF-beta1 and empty gelatin microspheres. Complete defect closure was histologically observed by the newly formed bone tissue. Without the barrier membrane, gelatin microspheres incorporating TGF-beta1 were less effective in inducing bone formation, whereas free TGF-beta1 and empty gelatin microspheres were ineffective. The skull defect was occupied by fibrous tissue infiltrated in place of bone tissue. The bone mineral density at the skull defect applied with gelatin microspheres incorporating TGF-beta1 plus the membrane was significantly higher than that of gelatin microspheres incorporating TGF-beta1 alone. The present data indicated that physical protection from the soft tissue infiltration enabled gelatin microspheres incorporating TGF-beta1 to synergistically enhance the osteoinductive ability at the skull defect. PMID- 11261878 TI - pH-sensitivity of fast responsive superporous hydrogels. AB - Stimuli-sensitive hydrogels (or smart hydrogels) are hydrogels that swell or shrink in response to small changes in environmental conditions in which they are placed. While the extent of swelling or shrinking may be large, the kinetics of such changes is slow, since the diffusion of water into and out of the hydrogel is a slow process. To obtain fast responses, we have prepared superporous hydrogels (SPHs) that can swell or shrink extremely fast regardless of their dimensions. The swelling and shrinking are orders of magnitude faster than expected for a nonporous hydrogel of the same dimensions. Water molecules are taken up into the SPHs by capillary forces, and this makes water uptake much faster than diffusion. The swelling ratio of the poly(acrylamide-co-acrylic acid) (p(AM-co-AA)) SPHs was dependent on the pH and ionic strength of the medium. The effect of pH was most pronounced and the effect of ionic strength was observed at all pH values. SPHs made at pH around 5 showed transient maximum swelling when exposed to pH 1.2 medium due to the transient low hydrogen ion concentration inside the swelling SPHs. The p(AM-co-AA) SPHs showed repeated swelling and shrinking by alternating the medium pH between 1.2 and 7.5, and the changes in swelling ratio was quite fast occurring in a matter of a minute. This fast sensitivity may make the stimuli sensitive hydrogels useful in many applications not previously possible. These materials can be used for applications where a single-piece hydrogel is more advantageous than hydrogel microparticulates. PMID- 11261879 TI - The use of immunoliposomes for specific delivery of antimicrobial agents to oral bacteria immobilized on polystyrene. AB - Antibacterial immunoliposomes have been prepared using covalently bound antibody, raised to the cell surface of the bacterium Streptococcus oralis (S. oralis), and incorporating the bactericides chlorhexidine and Triclosan. A regrowth assay, in which the ability of a bacterial biofilm immobilised on polystyrene to grow after exposure to a test solution, was undertaken to study the action of the antibacterial immunoliposomes. The antibacterial anti-oralis immunoliposomes show enhanced growth inhibition of S. oralis, compared to free bactericide, using low bactericide concentrations. For short exposure times to the biofilms, antibacterial anti-oralis immunoliposomes can show several times enhanced growth inhibition of S. oralis compared to free bactericide. Antibacterial anti-oralis immunoliposomes inhibit the growth of S. oralis more than that of other oral bacteria. The extent of growth inhibition by antibacterial anti-oralis immunoliposomes is linearly related to the number of immunoliposomes targeted to the biofilm surface. PMID- 11261880 TI - Surface modification of polystyrene nanoparticles using dextrans and dextran-POE copolymers: polymer adsorption and colloidal characterization. AB - Hydrophobically-modified dextran (dextran-phenoxy, DexP) and dextran-phenoxy poly(oxyethylene) (DexP-POE) copolymers have been used to modify the surface properties and the stability of polystyrene nanoparticles. We examined the effect of phenoxy group and POE chain concentrations on their adsorption behaviour. The adsorbed amount was determined by the standard depletion method and the layer thickness of the adsorbed layer by photon correlation spectroscopy and electrokinetic measurements. The results show that the hydrophobic interaction is the driving force during the adsorption while the layer thickness correlates with the interfacial concentration of grafted POE chains. The effects of adsorbed layers on the properties of latex dispersions have been characterized in terms of the stability of the dispersions toward added electrolyte and temperature. The conformation of the adsorbed copolymers is discussed in relation to layer thickness and colloidal stability of suspensions. PMID- 11261881 TI - Strength retention of self-reinforced drawn poly-L/DL-lactide 70/30 (SR-PLA70) rods and fixation properties of distal femoral osteotomies with these rods. An experimental study on rats. AB - Self-reinforced polylevo-dextro-lactic acid (SR-PLA) 70 composite rods, (2 mm x 26 mm) were implanted in the dorsal subcutaneus tissue of sixteen rats. Osteotomies of the distal femur were fixed with SR-PLA70 composite rods (2 mm x 15 mm) in 39 rats. The follow-up times varied from 1 week to 1 year. After sacrifice three-point bending and shear tests were performed for subcutaneously placed rods, and radiological, histological, histomorphometrical, microradiographic, and oxytetracycline-fluorescence studies of osteotomized and intact control femora were performed. At 52 weeks the shear strength and flexural modulus of the rods were 41% of the initial value, and the flexural strength was 43% of the initial value. In the osteotomies seven specimens had to be excluded due to postoperative infection or dislocation of the fragment. Six of the thirty two evaluated osteotomies showed signs of postoperative infection. Twenty-six osteotomies healed uneventfully. No signs of inflammatory or foreign-body reaction were observed. The present investigation demonstrated that the mechanical strength and fixation properties of the SR-PLA70 rods are suitable for fixation of cancellous bone osteotomies in rats. The present article is the first report on successful application of SR-PLA70 rods for fixation of cancellous bone osteotomies studied. PMID- 11261882 TI - HEMA-based methacrylic carriers incorporating ketoprofen: chain flexibility and swelling behaviour. AB - The chain flexibility, swelling and release behaviour of several copolymer systems of pharmacological interest have been studied. The systems are copolymers of 2-hydroxyethyl methacrylate with two support comonomers, i.e. N-(4 hydroxyphenyl) methacrylamide and N-[4-(2-hydroxy)ethoxyphenyl] methacrylamide and copolymers of 2-hydroxyethyl methacrylate with these carrier structures bearing ketoprofen (N-(4-[2-(3-benzoylphenyl)propionyloxy]phenyl methacrylamide and N-(4-[2-(3-benzoylphenyl)propionyloxy]2-ethoxyphenyl methacrylamide). In these copolymers ketoprofen was attached via different spacer groups, i.e. 4 aminophenoxy and 4-aminoethoxyphenyl. The chain structures are discussed on the basis of reactivity ratios. The properties of the polymeric drugs and of the parent carriers were studied comparatively by DSC and through the swelling of films from DMF solutions in pH 7.4 buffers. Data are discussed in terms of chain flexibility and swelling. PMID- 11261883 TI - Doxorubicin-peptide conjugates overcome multidrug resistance. AB - A well-known mechanism leading to the emergence of multidrug-resistant tumor cells is the overexpression of P-glycoprotein (P-gp), which is capable of lowering intracellular drug concentrations. To overcome this problem, we tested the capability of two peptide vectors that are able to cross cellular membranes to deliver doxorubicin in P-gp-expressing cells. The antitumor effect of peptide conjugated doxorubicin was tested in human erythroleukemic (K562/ ADR) resistant cells. The conjugate showed potent dose-dependent inhibition of cell growth against K562/ADR cells as compared with doxorubicin alone. Doxorubicin exhibited IC50 concentrations of 65 microM in the resistant cells, whereas vectorized doxorubicin was more effective with IC50 concentrations of 3 microM. After treatment of the resistant cells with verapamil, the intracellular levels of doxorubicin were markedly increased and consequent cytotoxicity was improved. In contrast, treatment of resistant cells with verapamil did not cause any further enhancement in the cell uptake nor in the cytotoxic effect of the conjugated doxorubicin, indicating that the conjugate bypasses the P-gp. Finally, we show by the in situ brain perfusion method in P-gp-deficient and competent mice that vectorized doxorubicin bypasses the P-gp present at the luminal site of the blood brain barrier. These results indicate that vectorization of doxorubicin with peptide vectors is effective in overcoming multidrug resistance. PMID- 11261884 TI - Hydrolyzable hydrophobic taxanes: synthesis and anti-cancer activities. AB - A series of taxane prodrugs with 2-bromoacyl chains attached at the 2'-position of the paclitaxel side chain, varying from six, eight, 12, 14 to 16 carbons in length, were synthesized, characterized and evaluated against human breast MCF-7 cancer cell line for their growth inhibitory (GI50) activities. The GI50 is the drug concentration required to inhibit cell growth by 50%. For comparison, hydrophobic taxanes varying in acyl chain lengths from six to 16 carbons were also synthesized and compared for their G050s with taxanes having equivalent bromoacyl chain lengths. The bromoacyl taxanes bearing six, eight and 12 carbon acyl chain lengths had GI50 values very similar to parent paclitaxel. The GI50 was 3 nM for three taxanes versus 1 nM for paclitaxel on the MCF-7 cell line. Increasing the acyl chain length to 14 or 16 carbons resulted in a significant decrease in cytotoxicity and an increase in the GI50 to 20 or 70 nM, respectively. In general, the GI50 values were directly related to the bromoacyl chain lengths in cultured tumor cells. Unlike bromoacyl taxanes, the taxanes lacking bromine in their acyl chain composition were 50- to 250-fold less active, suggesting that the heteroatom facilitated the hydrolysis of acyl chains to yield free paclitaxel. These differences in growth inhibitory activities may indirectly reflect differences in the susceptibility of the acyl chain to bromine-induced hydrolysis after association of the derivative with cell membranes. Liposome formulations of 2-bromoacyl taxanes bearing six, eight, 12 and 16 carbons were prepared and tested in SCID mice against a xenografted human ovcar-3 ovarian tumor. In vivo results showed that bromoacyl taxanes with a longer chain were therapeutically more efficacious than those with a short chain, presumably due to slow hydrolysis of the prodrug followed by sustained delivery of paclitaxel to the tumor. PMID- 11261886 TI - Neuropeptide receptor status in human tumor cell lines. AB - Tumor types expressing a neuroendocrine phenotype secrete neuropeptides with paracrine or autocrine growth factor activity. The efficacy of these paracrine or autocrine loops depends on the expression of specific receptors on tumor cells. Once specific receptors are identified, specific neuropeptide antagonists disrupting paracrine and autocrine loops could be potential treatments in neuropeptide-secreting tumors. In the present study, 11 human tumor cell lines representing astrocytoma, lymphoma, and pancreatic, prostate, lung and colon carcinomas were examined for expression of five different neuropeptide receptors (cholecystokinin, neurotensin, vasopressin, tachykinine substance P and cannabinoid) using RT-PCR and radioligand binding. The presence of various neuropeptide receptors in different human cancer cell lines supports development of new antitumor treatments based on disruption of neuropeptide autocrine growth pathways. PMID- 11261885 TI - Overexpression of aldose reductase in liver cancers may contribute to drug resistance. AB - We previously found that about 29% of human liver cancers overexpressed aldose reductase (AR) and about 54% of them overexpressed an AR-like gene called ARL-1 that has similar enzymatic activities to AR. Since these aldo-keto reductases can reduce a broad spectrum of substrates including cytotoxic aldehydes, we were interested to find out if these enzymes can contribute to the resistance of liver cancer chemotherapy by inactivating some of the anticancer drugs. HepG2 cells, a stable line of liver cells, were induced to overexpress AR by hypertonicity. Cells that were cultured in hypertonic medium became more resistant to daunorubicin, suggesting that overexpression of AR made the cells more resistant to this drug. This is confirmed by the fact that addition of AR inhibitor sensitizes the cells to this drug again. This information may be important for designing new drugs to treat this deadly disease. PMID- 11261887 TI - Antitumor activity of L-asparaginase from Thermus thermophilus. AB - L-asparaginase (EC 3.5.1.1) was purified to homogeneity from Thermus thermophilus. The apparent molecular mass of L-asparaginase was found to be 33 kDa by SDS-PAGE, whereas by Sephacryl S-300 superfine column it was found to be 200 kDa, indicating that the enzyme in the native stage acts as hexamer. It is a thermostable enzyme and keeps all of its activity at 80 degrees C for 10 min. The antiproliferative activity of the purified L-asparaginase from T. thermiphilus was tested against the following human cell lines: K-562 (chronic myelogenous leukemia), Raji (Burkitt's lymphoma), SK-N-MC (primitive neuroectodermal tumor), HeLa (cervical cancer), BT20 and MCF7 (breast cancers), HT-29 (human colon cancer), and OAW-42 (ovarian cancer). The antiproliferative activity of T. thermophilus enzyme was compared with Erwinase, the commercially available L asparaginase from Erwinia corotovora. The potency difference between the two L asparaginases was greater in HeLa and SK-N-MC than in other cell lines. The fact that L-asparaginase from T. thermophilus does not hydrolyse L-glutamine makes it advantageous for future clinical trials. PMID- 11261888 TI - The antioxidant caffeic acid phenethyl ester induces apoptosis associated with selective scavenging of hydrogen peroxide in human leukemic HL-60 cells. AB - Caffeic acid phenethyl ester (CAPE), an active component of propolis, has many biological and pharmacological activities including antioxidation and tumor cell cytotoxicity. We examined the type of cell death in human leukemic HL-60 cells after CAPE treatment in order to elucidate the relationship between CAPE-induced alterations of the redox state and apoptosis. CAPE treatment (6 microg/ml) resulted in marked growth inhibition up to 70.3+/-4.0% at day 2. This inhibition was partially blocked by pretreatment with N-acetyl-L-cycteine (NAC). Agarose gel electrophoresis showed evident DNA fragmentation after CAPE treatment. CAPE induced a significant decrease in mitochondrial transmembrane potential to about half of the untreated level after 6 h and a rapid depletion of intracellular glutathione (GSH) down to 41.7+/-6.0% after 1 h. Pretreatment of HL-60 cells with NAC reversed the GSH depletion and partially rescued cells from CAPE-induced apoptosis. With regard to intracellular reactive oxygen species, CAPE caused a fast and profound scavenging of H202 (19% of untreated cells after a 2-h treatment) but not of superoxide anion. These results suggest that apoptosis induced by CAPE is associated with mitochondrial dysfunction, GSH depletion and selective scavenging of H2O2 in human leukemic HL-60 cells. PMID- 11261889 TI - Simulation tool for schedule-dependent etoposide exposure based on pharmacokinetic findings published in the literature. AB - It is the aim of this study to establish a simulation tool for etoposide (Eto) which can be used to interpret drug monitoring data in clinical practice and to design new schedules for future protocols. As schedule dependency was observed for Eto, knowledge of concentration-time profiles is important. Pharmacokinetic data from children after low-dose i.v. administration of Eto together with data reported in the literature were used to construct the simulation tool. Validation was performed by independently reproducing various published data. Dose linearity of AUC was shown over the whole dose range of 20-2000 mg/m2 reported in the literature and fits the predictions by the simulation tool. There was no difference in clearance between children and adults. Close agreement was found between predicted and reported concentration-time profiles after various administration schedules. However, subgroups with significantly altered pharmacokinetics of Eto, such as patients with renal impairment or concurrent cisplatin chemotherapy, were excluded from the comparisons. In these patients values predicted for a 'regular' patient might be used as a base for possible dose modifications. In summary, a pharmacokinetic model of high predictive value is presented which allows simulations of Eto concentration-time profiles for low- as well as high-dose conditions and various infusion times. PMID- 11261890 TI - D-TRP-6-LHRH (Triptorelin) is not effective in ovarian carcinoma: an EORTC Gynaecological Cancer Co-operative Group Study. AB - Between March and September 1988, 74 patients with progressive ovarian cancer after prior platinum-based therapy were treated with the luteinizing hormone releasing hormone (LHRH) agonist Triptorelin (Decapeptyl degrees). Treatment consisted of i.m. injection of 3.75 mg of microencapsulated Triptorelin on days 1, 8 and 28 followed by 4-weekly injections until tumor progression. No objective responses were observed. Eleven out of 68 evaluable patients (16%) had stable disease. The median progression-free survival was 5 months in patients with disease stabilization and 2 months for all evaluable patients. The median survival for patients with disease stabilization was 17 months, whereas for all patients it was 4 months. The treatment was well tolerated; the only reported adverse events were incidental hot flushes. This study showed that the LHRH agonist Triptorelin has only modest efficacy in patients pretreated with platinum containing chemotherapy. PMID- 11261891 TI - High-dose chemotherapy in breast cancer--interpretation of the randomized trials. AB - High-dose chemotherapy was widely viewed as an effective treatment modality in breast cancer until the preliminary results of randomized trials proved disappointing. When it transpired that the author of the two unequivocally positive studies had fabricated data, many decided that high-dose chemotherapy in breast cancer was no longer worth studying. In fact, however, the reported results from randomized studies are consistent with a modest progression-free survival advantage of high-dose chemotherapy over conventional dose. Several more years of data maturation and the results of additional randomized trials must be awaited. PMID- 11261892 TI - Modulation of camptothecin analogs in the treatment of cancer: a review. AB - The topoisomerase I inhibitors reviewed in this paper are all semisynthetic analogs of camptothecin (CPT). Modulation of this intranuclear enzyme translates clinically in to antitumor activity against a broad spectrum of tumors and is therefore the subject of numerous investigations. We present preclinical and clinical data on CPT analogs that are already being used in clinical practice [i.e. topotecan and irinotecan (CPT-11)] or are currently in clinical development (e.g. 9-aminocamptothecin, 9-nitrocamptotecin, lurtotecan, DX 8951f and BN 80915), as well as drugs that are still only developed in a preclinical setting (silatecans, polymer-bound derivates). A variety of different strategies is being used to modulate the systemic delivery of this class of agents, frequently in order to increase antitumor activity and/or reduce experienced side effects. Three principal approaches are discussed, including: (i) pharmaceutical modulation of formulation vehicles, structural alterations and the search for more water-soluble prodrugs, (ii) modulation of routes of administration and considerations on infusion duration, and (iii) both pharmacodynamic and pharmacokinetic biomodulation. PMID- 11261893 TI - Polycyclic aromatic hydrocarbon exposure and burden of outdoor workers in Budapest. AB - Polycyclic aromatic hydrocarbons exposure (PAHs: (benz[a]anthracene, benzo[a]pyrene, dibenz[a,h]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, indeno[1,2,3-cd]-pyrene, fluoranthene, chrysene, pyrene) of policemen on street duty in downtown Budapest and workers repairing the road (asphalting) at a traffic junction and their excretion of PAH metabolites (1 hydroxypyrene, 3-hydroxybenz[a]anthracene, and 3-hydroxybenzo[a]pyrene) were determined. As controls, health-care workers were investigated. In addition PAH pollution of the air of a factory processing asphalt was also measured. The measurements were performed on air samples gained using personal samplers and from urine of end-shift samples using a high-performance liquid chromatography method. It was found that PAH pollution of the most crowded and busy center of Budapest was similar to that of several other cities in the world. PAH exposure of road builders was actually not higher than that of policemen; the slight difference resulted from diverging life-styles. PAH metabolite excretion of smoking health-care workers, road builders, or policemen significantly exceeded that of the nonsmokers. The PAH metabolite values of the three groups engaged in various activities did not show any difference. It was concluded that cancer related risk due to PAH compounds in the case of policemen on street duty and road builders (asphalting) does not exceed significantly that of workers not exposed occupationally to PAHs in the ambient air, but that smoking is a decisive factor. PMID- 11261894 TI - Follow-up genotoxicological monitoring of nurses handling antineoplastic drugs. AB - Most of the antineoplastic drugs used in the treatment of tumors are carcinogenic to humans. Hospital nurses are often subject to possible occupational carcinogen exposure. Exposure may occur during handling and administration of infusion solutions containing cytostatics. A genotoxicological monitoring system to detect genotoxic changes was developed in our laboratory, helping to improve working conditions and subserving primary prevention. Multiple-endpoint follow-up genotoxicological monitoring was performed in peripheral blood lymphocytes (PBLs) among 4 groups of 95 nurses (152 investigations) occupationally exposed to cytostatics. The results were compared to those of historical and industrial controls. The genotoxicological endpoints were the determination of the frequency of sister chromatid exchanges (SCEs) and the cells with high-frequency SCEs (HFC), the frequency of structural and numerical chromosome aberrations. and the measurement of ultraviolet-light-induced unscheduled DNA-repair synthesis (UDS). In Hospital 1, where nurses worked without a safety cabinet, the percentage of cells with chromosome aberrations (AC) was significantly higher than that of the controls. In Hospital 2, where nurses used inadequate safety cabinets (with horizontal airflow), significantly elevated levels of AC, SCE, HFC, and UDS were detected. During follow-up, in Hospital 2 at the time of the second investigation AC was still significantly higher, although safety conditions had been improved. The results indicate the presence of genotoxic damage in hospital nurses working with no or inadequate safety equipment. In Hospitals 3 and 4 where nurses using biological safety cabinets, the results were lower than those in the previous two groups. In Hospital 3 in the first year of the study AC was as at the level of industrial controls. During follow-up in the course of the repeated investigations a fluctuation in AC above the control level and an increase in HFC in yr 4 and 6 of the study were observed. In this group, the fluctuation in AC and HFC during the study points to the possibility of genotoxic exposure with cytostatics despite of the use of suitable safety cabinets, drawing attention to other possible routes of exposure. In Hospital 4, both AC and HFC were elevated. These data corroborate the need to maintain safety measures to avoid exposure, and the necessity of intervention in the case of exposure when using and handling hazardous carcinogenic agents. The results also indicate a certain expression time for genotoxic changes, which can lead to late somatic mutations as well as to a possible higher risk of cancer. PMID- 11261895 TI - Working condition-related improvement in genotoxicological parameters of Hungarian road pavers. AB - Multiple-endpoint follow-up genotoxicology monitoring was performed in a group of 22 Hungarian road pavers between 1996 and 1999. The studied endpoints were the determination of structural and numeric chromosome aberration (CA), sister chromatid exchange (SCE), high-frequency SCE and HPRT mutation frequencies, and ultraviolet (UV)-light-induced unscheduled DNA synthesis in peripheral blood lymphocytes (PBLs). The workers (8 hand pavers and 14 finishers, mean age 37 yr) used tar-free asphalt. The results were compared with those of 6 work-site controls (35 yr), 101 historical controls (38 yr), and 87 industrial controls (38 yr). The most marked changes were found in the CA frequencies. In the control, the mean CA frequency was 1.6%. In the first study, increased CA frequencies were found in the donors that either had been exposed to hot asphalt fumes or had cleaned the equipment with crude oil. The mean CA frequency of the 14 finishers working in closed cabins was 3.67% in 1996. The increased CA frequency was attributed to the high level of hot asphalt fumes due to insufficient ventilation. By 1999 the mean CA frequency decreased to 1.23%. For the 8 hand pavers working in open air the mean CA frequency was 3.6% in 1996. The obtained data suggested that the increase in CA frequencies was due to the use of petroleum and crude oil; therefore, these substances were replaced with harmless detergents. By 1999 the mean CA frequency decreased to 1%. In finishers the mean CA frequency returned to the control level 1 yr later (1999) than in the case of hand pavers. The chromosome-type aberrations remained predominant during the follow-up. The individual variations observed were attributed to smoking and inadequate personal protection. The obtained results suggest that the use of tar free asphalt and harmless detergents with adequate personal protection does not increase the frequencies of the genotoxicological parameters compared to controls. Consequently, an improvement in working conditions can prevent further exposures and thus decrease the cancer risk of road pavers. PMID- 11261896 TI - Alterations of K-ras and p53 mutations in colorectal cancer patients in Central Europe. AB - Many molecular investigations of colorectal cancer (CRC) have suggested that the accumulation of specific mutations in proto-oncogenes and tumor suppressor genes regulating cell growth via signal transduction trigger the stagewise progression to malignancy. In this study, the frequency, location, and type of mutations of the K-ras proto-oncogene exon I and p53 tumor suppressor gene exons 5-8 were analyzed in colorectal carcinomas of 65 patients from Central Europe, using polymerase chain reaction (PCR)-cold single-strand conformation polymorphism (SSCP) screening and direct sequencing. The incidence of K-ras activating mutations in these Central European samples was lower (25%) compared to that obtained in American and western European populations (40-50% at least), while the incidence of p53 inactivating mutations was similar (58%). These results suggest that some other genetically linked mechanisms may play a role in CRC development and progression, and hence K-ras and p53 mutations cannot be considered to be universal genetic markers for CRC. PMID- 11261897 TI - Specific amino acids moderate the effects on Ni2+ on the testosterone production of mouse leydig cells in vitro. AB - The purpose of this investigation was to study the effectiveness of two nickel binding amino acids, histidine (His) and cysteine (Cys), to prevent the inhibitory action of Ni2+ on testosterone (T) production by mouse primary Leydig cell culture. The maximal human chorionic gonadotropin (hCG)-stimulated T response was measured by radioimmunoassay (RIA) in the culture media. Three types of experiments were performed. In a concentration-response study, Ni2+ (62.5 to 1,000 microM) was added to the cells simultaneously with equimolar or twice the equimolar concentrations of His or Cys and the cultures were maintained for 48 h. Nickel-induced reduction in T production was completely prevented by equimolar concentrations of His at Ni2+ concentrations of 125, 250, and 500 microM; equimolar or twice the equimolar concentrations of His were only partially effective at 1,000 microM Ni2+. Protective action of Cys was complete only at the lowest concentration of Ni2+ (125 microM). In a second series, the cells were incubated for various times (0.5 to 48 h) with 1,000 microM Ni2+ in the presence of 2,000 microM His or Cys. Increasing the time of incubation, the protective effect of both amino acids against Ni2+ was reduced. In a third series, attempts were made to reverse the action of 1,000 microM Ni2+ after incubation with cells for various times (0.5 to 24 h), followed by exposure to 2,000 microM His or Cys. Cell cultures were maintained for 48 h. A partial recovery of hCG-stimulated T production could be observed only if the amino acid was added not later than 4 h after the metal. This time was also required to elicit the T depression produced by Ni2+. Administration of either His or Cys at later times had no effect. Our results show that both His and Cys are able to moderate the effects of Ni2+ on Leydig cell T production, depending on the concentration of this metal ion, as well as on amino acid. However, at higher Ni2+ concentrations the complete protection by His or Cys is only temporary. Administration of these amino acids after the Ni2+-produced decrease in T production was not able to reverse the process. PMID- 11261898 TI - Lead accumulation in human ovarian follicular fluid, and in vitro effect of lead on progesterone production by cultured human ovarian granulosa cells. AB - Lead content of ovarian follicular fluid obtained from 23 women was determined by atomic absorption spectrophotometry. In an in vitro experiment the direct effect of lead on the morphology and on progesterone (P) production by cultured granulosa cells of six women was investigated. Follicular fluid and granulosa cells were obtained from follicular aspirates of women undergoing in vitro fertilization (IVF) and embryo transfer (ET). Granulosa cells were cultured for 48 h to form monolayers in the presence or absence of lead acetate (100-1,600 microM). The effect of the metal proved to be concentration dependent. While 100 400 microM lead had no effect on the integrity of the monolayer, concentrations as high as 800 microM or higher inhibited cell adhesion and induced detachment of cells. The lead levels found in follicular fluid were 11.29 +/- 1.38 microg/L (0.056 +/- 0.007 microM). With lead in vitro at 1,600 microM (331.5 mg/L) there resulted a significant decrease in P production by granulosa cells. This concentration is very much higher than that measured in follicular fluid of IVF/ET patients, specifically nonexposed to lead, and even higher than mean blood levels reported by others in high exposure groups. In conclusion, lead seems not to exert a specific effect on the steroidogenesis by cultured human granulosa cells. Therefore, the lead levels measured in the ovarian follicular fluid seem not to pose a hazard with respect to progesterone secretion by the ovary. PMID- 11261899 TI - Effects of cobalt sulfate on prenatal development of mice, rats, and rabbits, and on early postnatal development of rats. AB - The effects of cobalt sulfate administered to pregnant C57BI mice, OFA-SD rats, and New Zealand rabbits was studied on fetal and postnatal offspring. Cobalt concentration in the maternal blood was increased in proportion to the administered doses. Cobalt crossed the placenta and appeared in the fetal blood and amniotic fluid. Regardless of the administered dose of cobalt sulfate, cobalt concentration in the blood peaked 2 h after administration. Cobalt produced dose dependent maternal toxicity and was found to be embryotoxic in all three species, as evidenced by elevated frequency of fetuses with body weight or skeletal retardation and embryolethality. Cobalt increased the frequency of major anomalies significantly in mice and rats, with anomalies of the eyes, kidneys, skull, spine, and sternum in mice, and anomalies of the urogenital system in rats. Cobalt sulfate was not teratogenic in rabbits. Intra-amnial administration of cobalt sulfate produced a dose-dependent increase of the frequency of dead fetuses, and weight retardation of the live fetuses. The direct cytotoxic effect probably plays a role in the embryotoxic and teratogenic effects of cobalt. The postnatal examinations revealed a decrease of the perinatal index in the treated group. The body weight of the pups in the treated group was lower during wk 1 of life, but no difference was found between the control and treated by the end of wk 2. Eye opening was completed in the usual time period in both groups, while time of appearance of the teeth, descending of the testes, shaping of ears, and development of hearing was delayed in the treated group. The development of muscle strength and of the locomotor system was delayed. All the functions studied (forward movement, swimming, righting reflex) normalized by postnatal d 21, with the exception of muscle strength. It was concluded that cobalt sulfate exposure decreases the perinatal viability of the fetuses, but the functions of the surviving fetuses with perinatal retardation become compensated by postnatal wk 2-3. The development of fetuses is undisturbed thereafter. PMID- 11261900 TI - The effect of prenatal indium chloride exposure on chondrogenic ossification. AB - Daily indium chloride doses of control (0) or 400 mg/kg were administered orally to pregnant Sprague-Dawley (SD) rats by gavage, on d 20 of gestation. Indium concentration was determined in the maternal and fetal blood, livers, kidneys, skulls, and femurs by atomic absorption spectrometry. Further groups of pregnant rats were treated with control (0) or 400 mg/kg indium chloride orally, during the whole gestation period. The fetuses were examined on d 21 of gestation, using histological and histochemical methods. Four hours after the administration indium concentration was found to be significant in the blood, liver, and kidneys of the dams. Twenty-four hours later it increased in the blood but not in the liver and kidney. Fetal indium concentrations were 40-50% of the maternal levels due to a barrier of the placenta. In the skull and the femur, indium was already detectable at 4 h after the administration, and by the end of 24 h, metal concentration was several times higher than that at 4 h, indicating accumulation. Furthermore, it was found that the birefringency of collagen detectable by picrosirius red staining in polarized light around the chondrocytes disappeared and became irregular. In the matrix of the epiphyseal cartilage, the regular, birefringent network demonstrable by Rivanol reaction became irregular and hardly recognizable. In the cytoplasm of the chondrocytes, the diffuse, evenly distributed positive Ricinus communis agglutinin reaction became irregular or disappeared. Similar but much weaker changes were observed with concanavalin A and wheat germ agglutinin stainings. It was concluded that the missing femur and micromelia diagnosed by alizarin staining is the consequence of a specific toxic effect of indium that inhibits chondrogenic ossification. No similar histochemical changes were observed in the bones of the skull developing by desmogenic ossification, despite the presence of indium. Data indicate that the mechanisms of the effects of indium causing retardation and/or malformation differ in the bones developing through desmogenic or chondrogenic ossification. PMID- 11261901 TI - Hemodynamic effect of indium chloride in pregnant rats. AB - Daily indium chloride doses of control (0) or 200 mg/kg were administered orally to pregnant Sprague-Dawley (SD) rats by gavage, on d 6-15 of gestation. On d 16 of gestation hemodynamic tests were performed; Arterial blood pressure, cardiac output (CO), and volume organ blood flow were determined with radioactive microspheres using the reference sample method (McDevitt & Nies, 1976). Indium chloride increased the cardiac index (CI), but did not change arterial blood pressure and total peripheral resistance (TPR). Indium decreased the organ fractions of the cardiac output to kidneys, ovaries, uterus, and placenta, while those to brain, lungs, and liver were not affected. In the placenta the blood flow was reduced significantly while the vascular resistance increased. The blood flow and vascular resistance did not change in the rest of the organs studied. The changes in arterial blood pressure, CO, Cl, TPR, organ fraction of cardiac output, blood flow, and vascular resistance in most of the organs displayed normal responsiveness to noradrenaline (NA) infusion. The reduction of uterine and placenta fractions and placental blood flow, produced by NA infusion were significantly greater in control than in the indium-treated group. Data indicate that the hemodynamic changes induced by indium are detrimental to the fetus. Indium chloride exposure modifies the maternal effect of noradrenaline such that there is maternal survival at the expense of fetal mortality. PMID- 11261902 TI - Study of inflammatory responses to crocidolite and basalt wool in the rat lung. AB - The subacute effects of crocidolite and basalt wool dusts were studied by nmeans of biochemical, morphological. and histological methods 1 and .3 mo after intrabronchial instillation. The cell count, protein and phospholipid contents, and lactate dehydrogenase (LDH) activity were determined in the bronchoalveolar lavage (BAL). Both types of fibers induced a prolonged inflammatory reaction in the lung. All the parameters studied in the experimental groups were more markedly elevated after 3 mo. Relative to the control, the protein and LDH values were increased three- to fivefold, the phospholipid content twofold, and the number of free cells in the BAL exceeded the control level up to ninefold. The inflammatory responses to crocidolite and basalt wool in the lung did not differ significantly. In spite of this, basalt wool is recoinmended as an asbestos substitute, as the use of this man-nade fiber may result in a significantly lower release of dust than that from crocidolite. PMID- 11261903 TI - Anticoagulation and cataract surgery: a review of the current literature. AB - The anticoagulated patient presenting for cataract surgery presents many dilemmas for anaesthetist and surgeon alike. Current evidence suggests that warfarin therapy significantly improves prognosis in patients with atrial fibrillation with coexisting cerebrovascular disease, and those with non-tissue prosthetic heart valves. Inadequate anticoagulation in these groups exposes them to higher risk of systemic embolic complications, which are frequently devastating. Warfarin is an extremely complex drug. Attempted cessation and recommencement of warfarin therapy may not only reverse anticoagulation for unpredictable periods of time but may also expose patients to a transient yet dangerous hypercoagulable state. In most instances this state represents an additive risk to the untreated disease for which warfarin is being prescribed. It is difficult to accurately measure risks of local anaesthetic blockade in anticoagulated patients as techniques are not standardized. Smaller needles and single injections appear safer with deep eye blocks, while sub-Tenon's block and topical techniques appear safer still, and acceptable provided patients and surgeons are happy with the conditions so created. Retrobullbar haemorrhage appears to occur more frequently in anticoagulated patients who have their anticoagulation continued up to the time of cataract surgery. Retrobulbar haemorrhage is also more frequent in this same group even when anticoagulation is ceased prior to surgery when compared to non-anticoagulated patients. Prognosis for visual acuity with retrobulbar haemorrhage is generally good, given an experienced surgeon is present to rapidly decompress the eye. PMID- 11261904 TI - Non-invasive cardiac output determination: comparison of a new partial rebreathing technique with thermodilution. AB - This study compares a derivative Fick technique using carbon dioxide (CO2) with the thermodilution pulmonary artery catheter (PAC), for determination of cardiac output (CO). Subjects were sedated, mechanically ventilated adults following elective cardiac surgery Microprocessor controlled deadspace activation and side stream capnography in a ventilator circuit enabled calculation of CO (CO(CO2)) every four minutes. Thermodilution CO (CO(TD)) was performed as clinically indicated and at 20-minute intervals. Simultaneous CO(TD)/CO(CO2) pairs were recorded from time of admission to ICU for a minimum period of two hours for each patient. There were 358 CO(TD)/CO(CO2) pairs recorded from 41 patients. Cardiac output measurements ranged from 2. 7 to 10.6 l/min. The bias (Bland-Altman) was 0.050 l/min (95% CI -0.024 to 0.125 l/min). The 95% limits of agreement were 1.354 to 1.455 l/min. This simple, non-invasive partial-rebreathing technique is a valid alternative to thermodilution for cardiac output determination in sedated, mechanically ventilated patients. There are significant implications for improved safety, reduced complexity and reduced cost in anaesthesia and intensive care. PMID- 11261905 TI - Anaesthesia for amiodarone-induced thyrotoxicosis: a case review. AB - Amiodarone-induced thyrotoxicosis is a life-threatening problem that is very effectively managed by total thyroidectomy, although often many of these patients are considered "too unfit" for anaesthesia. The aim of this study was to review the safety of anaesthesia for total thyroidectomy in the acute management of amiodarone-induced thyrotoxicosis. Information was obtained retrospectively from a prospective endocrine surgical database and from hospital records. Data, including outcomes and morbidity, are presented from 12 patients who underwent anaesthesia for total thyroidectomy as an urgent procedure for amiodarone-induced thyrotoxicosis. Despite the fact that these patients had uncontrolled thyrotoxicosis and marked cardiac failure at the time of anaesthesia, no anaesthetic or surgical mortality was seen. We conclude that a challenging patient with amiodarone-induced thyrotoxicosis that has not responded to conservative measures may be considered for total thyroidectomy early in their management. Total thyroidectomy can be performed under general or local anaesthesia with apparent relative safety. PMID- 11261906 TI - Early intravenous anaesthesia. PMID- 11261907 TI - A randomized comparison of low-dose ketamine and lignocaine infiltration with ketamine-diazepam anaesthesia for post partum tubal ligation in Vanuatu. AB - Ketamine remains one of the most commonly used anaesthetic agents around the world. Despite it being the anaesthetic agent of choice in many developing nations, there is a paucity of literature describing ketamine in the developing world. In what we believe is the first randomized controlled trial to be performed in Vanuatu (formerly the New Hebrides) we compared the use of ketamine 0.9 mg/kg and diazepam 0.07 mg/kg with ketamine 0.3 mg/kg and 2% lignocaine infiltration in 50 Melanesian women undergoing post partum tubal ligation. All women received 0.5 mg/kg intramuscular pethidine. Visual analog pain scores and verbal numeric satisfaction scores were similar between the groups. However the time to obeyed command was significantly faster in the 0.3 mg/kg ketamine group (7.0+/-4.9 vs 13.0+/-9.2 min). The incidence of dreaming was similar and the content rated as pleasant by both groups. In institutions where post-anaesthesia care resources are limited, 0.3 mg/kg ketamine with local anaesthesia provides for earlier self-care of patients after tubal ligation, without compromise of analgesia, emergence or satisfaction. The implications of these findings extend to other procedures that require short general anaesthesia, which can be adequately performed with low-dose ketamine and local anaesthesia. The latter technique allows more rapid awakening. PMID- 11261908 TI - Anaesthesia and myotonia--an Australian experience. AB - The purpose of this audit was to retrospectively examine the anaesthetic technique and perioperative complications among adult patients with confirmed myotonic dystrophy who presented for surgery at Royal Perth Hospital. A total of 18 general anaesthetics, two spinal anaesthetics, one conscious sedation and six eye blocks were performed. No deaths and no myotonic episodes were described despite the use of a variety of techniques, including the administration of succinylcholine to three patients. The only patient to experience postoperative complications was severely unwell preoperatively and underwent upper abdominal surgery. The audit revealed an overall complication rate of 5.5% of general anaesthetics or 3.8% of all anaesthetics in this patient population. PMID- 11261909 TI - The use of thiopentone/propofol admixture for laryngeal mask airway insertion. AB - An admixture of thiopentone and propofol was evaluated against propofol for laryngeal mask airway (LMA) insertion. Eighty-one ASA 1 and 2 18- to 65-year-old patients, premedicated with 7.5 mg midazolam orally were assigned randomly to receive either propofol 1% or an admixture of thiopentone and propofol (1.25% and 0.5% respectively), both at a dose of 0.25 ml x kg(-1). Satisfactory conditions for insertion were achieved with the admixture, which was comparable to propofol (73% vs 85%, P>0.05). There was no statistical difference in the incidence or severity of gagging, coughing, inadequate jaw relaxation and laryngospasm. The incidence of hypotension was lower in the admixture group (51% vs 78%, P=0.02). The duration of apnoea was not different between the admixture and propofol group (mean 103s vs 109s respectively, P>0.05). We conclude that thiopentone/propofol admixture can be a suitable alternative to propofol for LMA insertion, producing less hypotension while allowing cost savings of up to 45%. An admixture of thiopentone and propofol (1.25% and 0.5% respectively) can produce suitable conditions compared to propofol 1%, for laryngeal mask insertion. In addition to cost containment, the admixture also produces less hypotension. PMID- 11261910 TI - The prevalence of latex allergy in operating theatre staff. AB - Latex allergy has become increasingly common amongst health care workers. The prevalence of latex allergy in 102 theatre personnel at Princess Alexandra Hospital was determined by the results of a standardized questionnaire and a latex specific IgE radioallergosorbent test (RAST). Volunteers had their forced expiratory volume in 1 second/forced vital capacity (FEV1/FVC) measured at the beginning and end of their working shifts. Only one of 102 volunteers had a positive latex specific IgE RAST and he also experienced both local and systemic symptoms with latex exposure and a deterioration in daily FEV1/FVC. The 14 volunteers using asthma medications had significantly higher incidence of operating theatre symptoms (10/14), local reactions (8/14), and atopy (14/14). There was no clinically significant daily change in FEV1 in the total population or any specific group. The 1% (1/102) prevalence of latex allergy and sensitization in theatre staff demonstrates a much lower incidence than previously reported (12.5%, 15.8%). PMID- 11261911 TI - A standardized, uniform and universal dental chart for documenting state of dentition before anaesthesia. AB - It is vital to have adequate and precise documentation of the condition of a patient's dentition before commencing an anaesthetic. The incidence of dental damage during anaesthesia is not low. To the authors' knowledge, there is no standardized method used by anaesthetists to document the state of a patient's dentition. We propose the introduction of a standardized uniform dental chart to enable anaesthetists to accurately document the condition of their patients' teeth. This vital information can be easily obtained during the preanaesthetic assessment. With the increase in medical litigation and demands for adequate documentation, we believe this chart can become an invaluable part of every hospital's preanaesthetic assessment form. The dental chart is to be offered as a service to anaesthetists in the form of a copyright-free "Freeware" computer diskette or adhesive sticker and will be downloadable from the internet. PMID- 11261912 TI - Bilateral frontal haemorrhages associated with continuous spinal analgesia. AB - We report a case of intracerebral haemorrhages associated with continuous spinal analgesia. Continuous spinal analgesia is frequently employed for postoperative analgesia in high-risk patients in our institution. The analgesia is administered via a 20 gauge catheter passed through an 18 gauge Tuohy needle (Portex). A 71 year-old man with severe respiratory impairment had an intrathecal catheter placed for postoperative analgesia. He had a difficult postoperative course, including wound dehiscence, and died from respiratory failure some five weeks postoperatively. On day nine postoperatively he had two tonic-clonic seizures and was subsequently found to have developed bilateral frontal intracerebral haemorrhages. There was no previous history of seizures. Although several confounding variables exist, the most likely explanation for the intracerebral event appears to be an association with the dural puncture and intrathecal catheter Possible mechanisms and risk factors are discussed. PMID- 11261913 TI - Acute haemorrhagic leucoencephalitis complicating sepsis. AB - A case of acute haemorrhagic leucoencephalitis presenting as fatal septic encephalopathy is reported. The clinical features of this condition are reviewed and the potential for earlier diagnosis is considered. PMID- 11261914 TI - Patient-controlled spinal analgesia for labour and caesarean delivery. AB - Continuous spinal anaesthesia has not been widely used in Australia. Epidural anaesthesia is often inadequate in patients with previous spinal surgery, as distribution of local anaesthetic in the epidural space is unpredictable. Two cases are presented where continuous spinal anaesthesia enabled satisfactory analgesia and anaesthesia to be obtained for labour and caesarean delivery respectively. PMID- 11261915 TI - Anterior spinal artery syndrome following total hip arthroplasty under epidural anaesthesia. AB - We present a case of anterior spinal artery syndrome in a 57-year-old man having a total hip arthroplasty under epidural anaesthesia. Epidural insertion and surgery were uneventful. Postoperatively bilateral lower limb motor weakness was attributed to the initial dose of local anaesthetic. There was no change in neurological status 24 hours later. Magnetic resonance imaging demonstrated spinal cord infarction. The diagnosis of anterior spinal artery syndrome was made based on the patient's neurological condition and MRI findings. PMID- 11261916 TI - Anaesthetic management of a caesarean section in a patient with Marfan's syndrome and aortic dissection. AB - This report describes a case of a Stanford Type B aortic dissection (originating distal to the left subclavian artery and extending to the aortic bifurcation and proximal left iliac artery) in a 31-year-old primigravid woman who was at 39 weeks gestation and had Marfan's syndrome. The dissection was managed conservatively. Caesarean section was performed under epidural anaesthesia with aggressive control of hypertension. Postoperatively, there was no extension of the dissection and no aneurysm formation. She was discharged from hospital two weeks after delivery and remained asymptomatic at six months. There are no plans for surgical intervention. PMID- 11261917 TI - Anaesthesia in a child with autosomal recessive omodysplasia. AB - A child with a recently described form of dwarfism, autosomal recessive omodysplasia, presented for general anaesthesia for dental conservation. Despite the patient's dysmorphic features and craniofacial and mandibular deformities, no significant anaesthetic problems were encountered. Anaesthetic implications of the condition are reviewed. PMID- 11261918 TI - Yet another use for the laryngeal mask airway--ventilation of a patient with a tracheostomy stoma. PMID- 11261919 TI - Non-pharmacologic cause of anti-emetic resistant postoperative nausea and vomiting. PMID- 11261920 TI - Tracheal foreign body following tube change during percutaneous dilational tracheostomy: comment. PMID- 11261921 TI - Postobstructive pulmonary oedema. PMID- 11261922 TI - Early vs late LMA removal; risks to patients and damage to equipment. PMID- 11261923 TI - Non-invasive cardiac output (NICO) monitor. PMID- 11261924 TI - The endogenous retroviral insertion in the human complement C4 gene modulates the expression of homologous genes by antisense inhibition. AB - Intron 9 contains the complete endogenous retrovirus HERV-K(C4) as a 6.4-kb insertion in 60% of human C4 genes. The retroviral insertion is in reverse orientation to the C4 coding sequence. Therefore, expression of C4 could lead to the transcription of an antisense RNA, which might protect against exogenous retroviral infections. To test this hypothesis, open reading frames from the HERV sequence were subcloned in sense orientiation into a vector allowing expression of a beta-galactosidase fusion protein. Mouse L cells which had been stably transfected with either the human C4A or C4B gene both carrying the HERV insertion (LC4 cells), and L(Tk-) cells without the C4 gene were transiently transfected either with a retroviral construct or with the wild-type vector. Expression was monitored using an enzymatic assay. We demonstrated that (1) HERV K(C4) antisense mRNA transcripts are present in cells constitutively expressing C4, (2) expression of retroviral-like constructs is significantly downregulated in cells expressing C4, and (3) this downregulation is further modulated in a dose-dependent fashion following interferon-gamma stimulation of C4 expression. These results support the hypothesis of a genomic antisense strategy mediated by the HERV-K(C4) insertion as a possible defense mechanism against exogenous retroviral infections. PMID- 11261925 TI - Sequence and peptide-binding motif for a variant of HLA-A*0214 (A*02142) in an HIV-1-resistant individual from the Nairobi Sex Worker cohort. AB - As part of the ongoing study of natural HIV-1 resistance in the women of the Nairobi Sex Workers' study, we have examined a resistance-associated HLA class I allele at the molecular level. Typing by polymerase chain reaction using sequence specific primers determined that this molecule is closely related to HLA-A*0214, one of a family of HLA-A2 supertype alleles which correlate with HIV-1 resistance in this population. Direct nucleotide sequencing shows that this molecule differs from A*0214, having a silent nucleotide substitution. We therefore propose to designate it HLA-A*02142. We have determined the peptide-binding motif of HLA A*0214/02142 by peptide elution and bulk Edman degradative sequencing. The resulting motif, X-[Q,V]-X-X-X-K-X-X-[V,L], includes lysine as an anchor at position 6. The data complement available information on the peptide-binding characteristics of this molecule, and will be of use in identifying antigenic peptides from HIV-1 and other pathogens. PMID- 11261926 TI - A major determinant quantitative-trait locus responsible for atopic dermatitis like skin lesions in NC/Nga mice is located on Chromosome 9. AB - NC/Nga (NC) is a newly discovered model mouse for human atopic dermatitis, NC mice showing specific symptoms such as dermatitis and overproduction of IgE. To detect the loci responsible for the onset of dermatitis in the mice, backcross (N2) progeny between (NCxMSM/MS)F1 and NC were generated, where MSM/MS is an inbred strain from Japanese wild mice, Mus musculus molossinus. Linkage disequilibrium between dermatitis and various chromosome-specific microsatellite markers was then examined in the N2 segregants with severe dermatitis. The analysis revealed that the locus of the major determinant (designated here as derm1) was tightly linked to D9Mit163, D9Mit72, D9Mit143, D9Mit103, D9Mit207, and D9Mit209, because these markers showed the highest and most significant chi2 values. Since no recombination was observed among the markers in our linkage map, a radiation hybrid (RH) panel was applied to locate the derm1 locus more precisely. The markers were separated on the RH map, and their order was D9Mit163 D9Mit72-D9Mit143-D9Mit103-D9Mit207-D9Mit209 from the centromere. Several functional candidate genes are located near the locus derm1. These candidates are Thy1, Cd3d, Cd3e, Cd3g, Il10ra, 1118, and Csk, all of which could be involved in allergic responses through effects on T-cell function. Of these candidates, Csk is the strongest for NC dermatitis, since its map position was most tightly linked to the derm1 locus. PMID- 11261927 TI - Distribution of human kappa locus IGKV2-29 and IGKV2D-29 alleles in Swedish Caucasians and Hong Kong Chinese. AB - Polymorphism in the IGKV2-29 gene was shown to decrease the recombination frequency in B cells and to be important for immune responses to Haemophilus influenzae type b polysaccharide. By using the combination of PCR and restriction enzyme mapping, the distribution of IGKV2D-29 and IGKV2-29 gene alleles was estimated in two geographically and ethnically different groups. We found that V2D-29*01 homozygous individuals were most common in Swedish Caucasians (82%), but less common in the Chinese population of Hong Kong (28%). The homozygous V2D 29*02 genotype was found in 19% Chinese, but only in one Caucasian (1%). The frequency of the heterozygous V2D-29*01/V2D-29*02 genotype was also higher in the Chinese population (46%) compared with the Caucasians (7%). V2-29*01 homozygosity was more frequent among Caucasians (85%) than among Chinese (19%). In contrast, homozygous V2-29*02 individuals were over-represented in the Chinese population (18%), whereas only one was found among Caucasians (1%). Heterozygous V2-29*01/V2 29*02 individuals were also more common in the Chinese (63%) than the Caucasian (15%) population. Most Caucasians had the combination of V2D-29*01/V2D-29*01+V2 29*01/V2-29*01 (74%), while the most common genotype for Chinese was V2D 29*01/V2D-29*02+ V2-29*01/V2-29*02 (41%). Analysis of the association of V2D 29*02 and V2-29*02 alleles demonstrated a high degree of linkage, as for V2D 29*01 with V2-29*01. These data show a significant difference in the distribution of IGKV2D-29 and IGKV2-29 alleles among Swedish Caucasians and Hong Kong Chinese. This may help to explain differences in the occurrence of H. influenzae type b infection in the two populations. Evaluated methods for IGKV2D-29 and IGKV2-29 allele detection can be used for the screening allele polymorphisms in other particular patient groups. PMID- 11261928 TI - Characterization of the Japanese pufferfish (Takifugu rubripes) T-cell receptor alpha locus reveals a unique genomic organization. AB - Polymerase chain reactions with degenerate V gene segment primers were used to isolate the putative T-cell receptor alpha-chain gene (TCRA) from Japanese pufferfish (Takifugu rubripes). The putative TCRA chain cDNA is composed of an N terminus leader peptide followed by the variable region and the constant region. The variable portion of the TCRA gene is encoded by V and J gene segments separated in the germline. As in mammals, the V-J junction sequences are GC rich and highly diversified. Amino acid residues that are required to maintain the function and structural integrity of the TCRA polypeptide, including the conserved Trp-Tyr-Lys and Tyr-Tyr-Cys motifs in the V gene segments, the Lys-Leu X-Phe-Gly-X-Gly-Thr-X-Leu motif in the J gene segment, the three cysteine residues in the constant region and the charged residues in the transmembrane region are all preserved in the pufferfish. These conserved features suggest that the TCRA gene families in fish and mammals have evolved from a common ancestor. PMID- 11261929 TI - HLA-B*4034 identified in a UK Caucasoid potential bone marrow donor. PMID- 11261930 TI - Tumor necrosis factor-alpha promoter polymorphism TNF2 is associated with a stronger delayed-type hypersensitivity reaction in the skin of borderline tuberculoid leprosy patients. PMID- 11261931 TI - Characterization of a new HLA-G allele encoding a nonconservative amino acid substitution in the alpha3 domain (exon 4) and its relevance to certain complications in pregnancy. PMID- 11261933 TI - Genomic structure, expression, and chromosome mapping of the mouse homologue for the WSL-1 (DR3, Apo3, TRAMP, LARD, TR3, TNFRSF12) gene. PMID- 11261932 TI - Identity of IGHV-7183.1 (V81x) coding and recombination signal sequences among wild-derived mice. PMID- 11261934 TI - G7c in the lung tumor susceptibility (Lts) region of the Mhc class III region encodes a von Willebrand factor type A domain protein. PMID- 11261935 TI - Selective expression of NKG2-A and NKG2-C mRNAs and novel alternative splicing of 5' exons in rhesus monkey decidua. PMID- 11261936 TI - Identification of new cattle BoLA-DRB3 alleles by sequence-based typing. PMID- 11261937 TI - A practical approach to the older patient with cancer. PMID- 11261938 TI - Cell numbers: can they be controlled without programmed cell death? PMID- 11261939 TI - Serine proteases and brain damage - contribution of the urokinase- plasminogen activator system. PMID- 11261940 TI - Membrane bioenergetics and virulence: problems and prospects. PMID- 11261941 TI - The International Society for Heart and Lung Transplantation 21st Annual Meeting and Scientific Sessions. April 25-28, 2001. Vancouver, Canada. Abstracts. PMID- 11261942 TI - Synthesis and characterization of NaGaTe2O6.2.4h2O: a new open-framework tellurite related to zemannite. PMID- 11261943 TI - Semiconductor properties in an iodine-doped platinum(II) dinuclear complex. PMID- 11261944 TI - Pi* level tuning in a series of diimine ligands based on density functional theory: application to photonic devices. AB - Energy- and electron-transfer processes are very important for artificial photosynthesis and a variety of other applications. [(bpy)2Ru(PAP)Os(bpy)2]4+ and its oxidized form [(bpy)2Ru(PAP)Os(bpy)2]5+ perform efficient photoinduced energy and electron-transfer processes, respectively (k(en) = 5.2 x 10(7) s(-1), k(el) = 7.2 x 10(6) s(-1)). The introduction of appropriate donor and acceptor units on the Ru2+ center can improve the lifetime of the excited state, resulting in a much longer and efficient storage of energy. Nonempirical (density functional) calculations and experimental data are used to predict the best donor and acceptor ligands for improving electron- and energy-transfer processes. Such a result can be extended to all polynuclear complexes where electronic coupling between the metal centers is very weak. PMID- 11261945 TI - Synthesis and electrochemical studies of diiron complexes of 1,8-naphthyridine based dinucleating ligands to model features of the active sites of non-heme diiron enzymes. AB - A bis(mu-carboxylato)(mu-1,8-naphthyridine)diiron(II) complex, [Fe2(BPMAN)(mu O2CPhCy)2](OTf)2 (1), was prepared by using the 1,8-naphthyridine-based dinucleating ligand BPMAN, where BPMAN = 2,7-bis[bis(2-pyridylmethyl)aminomethyl] 1,8-naphthyridine. The cyclic voltammogram (CV) of this complex in CH2Cl2 exhibited two reversible one-electron redox waves at +296 mV (DeltaE(p) = 80 mV) and +781 mV (DeltaE(p) = 74 mV) vs Cp2Fe+/Cp2Fe, corresponding to the FeIIIFeII/FeIIFeII and FeIIIFeIII/FeIIIFeII couples, respectively. This result is unprecedented for diiron complexes having no single atom bridge. Dinuclear complexes [Fe2(BPMAN)(mu-OH)(mu-O2CPhCy)](OTf)2 (2) and [Mn2(BPMAN)(mu O2CPhCy)2](OTf)2 (3) were also synthesized and structurally characterized. The cyclic voltammogram of 2 in CH2Cl2 exhibited one reversible redox wave at -22 mV only when the potential was kept below +400 mV. The CV of 3 showed irreversible oxidation at potentials above +900 mV. Diiron(II) complexes [Fe2(BEAN)(mu O2CPhCy)3](OTf) (4) and [Fe2(BBBAN)(mu-OAc)2(OTf)](OTf) (6) were also prepared and characterized, where BEAN = 2,7-bis(N,N-diethylaminomethyl)-1,8-naphthyridine and BBBAN = 2,7-bis[2-[2-(1-methyl)benzimidazolylethyl]-N-benzylaminomethyl]-1,8 naphthyridine. The cyclic voltammograms of these complexes were recorded. The Mossbauer properties of the diiron compounds were studied. PMID- 11261946 TI - Redox properties of C6S8(n-) and C3S5(n-) (n = 0, 1, 2): stable radicals and unusual structural properties for C-S-S-C bonds. AB - The new anionic carbon sulfides C6S10(2-) and C12S16(2-) are described and crystallographically characterized. The C12S16(2-) anion consists of two C6S8 units connected by an exceptionally long (2.157(12) A) S-S bond. In solution, C12S16(2-) exists in equilibrium with the radical C6S8(-*). The equilibrium constant for radical formation (293 K, THF) is 1.2 x 10(-4) M, as determined by optical spectroscopy at varying concentrations. Radical formation occurs through scission of the S-S bond. On the basis of variable temperature EPR spectra, the thermodynamic parameters of this process are DeltaH = +51.5 +/- 0.5 kJ x mol(-1) and DeltaS = +110 +/- 3 J x mol(-1) x K(-1). C6S10(2-) is an oxidation product of C3S5(2-) and consists of two C3S5 units connected by an S-S bond. The S-S bond length (2.135(4) A) is long, and the CS-SC torsion angle is unusually acute (52.1 degrees ), which is attributed to an attractive interaction between C3S2 rings. The oxidation of (Me4N)2C3S5 occurs at -0.90 V vs Fc+/Fc in MeCN, being further oxidized at -0.22 V. The similarity of the cyclic voltammogram of (Me4N)2C6S10 to that of (Me4N)2C3S5 indicates that C6S10(2-) is the initial oxidation product of C3S5(2-). PMID- 11261947 TI - Thermodynamic and kinetic studies on the reaction between the vitamin B12 derivative beta-(N-methylimidazolyl)cobalamin and N-methylimidazole: ligand displacement at the alpha axial site of cobalamins. AB - The equilibria and kinetics of substitution of the 5,6-dimethylbenzimidazole at the alpha site of beta-(N-methylimidazolyl)cobalamin by N-methylimidazole have been investigated, and the product, bis(N-methylimidazolyl)cobalamin, has been characterized by visible and 1H NMR spectroscopies. The equilibrium constant for (N-MeIm)Cbl+ + N-MeIm right harpoon over left harpoon (N-MeIm)2Cbl+ was determined by 1H NMR spectroscopy (9.6 +/- 0.1 M(-1), 25.0 degrees C, I = 1.5 M (NaClO4)). The observed rate constant for this reaction exhibits an unusual inverse dependence on N-methylimidazole concentration, and it is proposed that substitution occurs via a base-off solvent-bound intermediate. Activation parameters typical for a dissociative ligand substitution mechanism are reported at two different N-MeImT concentrations, 5.00 x 10(-3) M (DeltaH++ = 99 +/- 2 kJ x mol(-1), DeltaS++ = 39 +/- 5 J x mol(-1) x K(-1), DeltaV++ = 15.0 +/- 0.7 cm3 x mol(-1), and 1.00 M (DeltaH++ = 109.4 +/- 0.8 kJ x mol(-1), DeltaS++ = 70 +/- 3 J x mol(-1) x K(-1), DeltaV++ = 16.8 +/- 1.1 cm3 x mol(-1)). According to the proposed mechanism, these parameters correspond to the equation of (N-MeIm)2Cbl+ and the ring-opening reaction of the alpha-DMBI of (N-MeIm)Cbl+ to give the solvent-bound intermediate in both cases, respectively. PMID- 11261948 TI - [(Zr6B)Cl11-xI2+x] (0< or = x < or = 6): a new mixed-halide structure with zigzag cains of clusters in multiply twinned crystals. AB - The new [(Zr6B)Cl11-xI2+x] phase (with 0 < or = x < or = 6) is obtained from reactions of ZrI4, ZrCl4, and elemental Zr and B for 2-4 weeks in sealed Ta tubing at 800-850 degrees C. Single crystals of [(Zr6B)Cl6.44(7)I6.56] have been characterized by X-ray diffraction at room temperature (orthorhombic Pbcn, Z = 4, a = 12.365(2) A, b = 15.485(3) A, c = 13.405(2) A). This structure contains zigzag chains of boron-centered (Zr6B) octahedra that are interconnected by Cl(i i) halides. Further three-dimensional connectivity is achieved by I(a-a-a) bridges. The noncluster interconnecting two-bonded X(i) sites are occupied statistically by a mixture of Cl and I. For each site both positions were resolved. This structure forms within a phase width of 0 < or = x < or = 6 at temperatures between 800 and 850 degrees C. Crystals of this phase appear to be always multiply twinned. PMID- 11261949 TI - N-monofunctionalized 1,4,7-triazacyclononane macrocycles as building blocks in inorganic crystal engineering. AB - The syntheses of N-(3-prop-1-ene)-1,4,7-triazacyclononane molybdenum tricarbonyl (2), N-(4-but-1-ene)-1,4,7-triazacyclononane molybdenum tricarbonyl (3), N-(3 prop-1-ene)-1,4,7-triazacyclononane molybdenum trioxide (5), N-(4-but-1-ene) 1,4,7-triazacyclononane molybdenum trioxide (6), N-(hydroxyethyl)-1,4,7 triazacyclononane molybdenum trioxide (7), and N-(2-methylpyridyl)-1,4,7 triazacyclononane molybdenum trioxide (8) have been achieved. The objective of this work is to systematically vary the functionality of the pendant group in order to create different crystal packing in the solid state. This is evidenced in comparing the structures of 1,4,7-triazacyclononane molybdenum trioxide (4) and 5-8, which were determined using X-ray crystallography. The synthesis and characterization of the new ligand N-(2-methylpyridyl)-1,4,7-triazacyclononane (L5) is reported. PMID- 11261950 TI - A mixed oxidation state, binuclear iron complex containing an unsymmetrically coordinating ligand. A ligand-induced switch in redox behavior. AB - Binuclear [FeIIFeIII(BMDP)(O2CPh)3](BF4) (1) was obtained by treating an acetonitrile solution of the fully reduced [FeIIFeII(BMDP)(O2CPh)(MeOH)1.5(H2O)0.5](BF4)2 with 5 equiv of benzoate and then exposing the mixture to oxygen. Examination of [FeIIFeIII(BMDP)(O2CPh)3](BF4) by X-ray crystallography reveals the localized, mixed oxidation state nature of the cation in the solid state. 1H NMR and magnetic susceptibility data for the new complex are also reported. In the absence of dioxygen and other oxidants, treatment of FeIIFeII(BMDP)(O2CPh)(MeOH)1.5(H2O)0.5](BF4)2 with excess benzoate results in the formation of [FeIIFeII(BMDP)(O2CPh)2](BF4)2, which has also been characterized by X-ray diffraction. PMID- 11261951 TI - Syntheses of Ru-S clusters with kinetically labile ligands via the photolysis of [(cymene)3RuS2](PF6)2. AB - Three ruthenium sulfide clusters with labile CH3CN ligands have been photochemically synthesized. Irradiation of [(cymene)3Ru3S2](PF6)2 ([1](PF6)2) in CH3CN gives [(cymene)2(CH3CN)3Ru3S2](PF6)2 ([2](PF6)2), which has been characterized by 1H NMR spectroscopy, ESI mass spectrometry, and chemical reactivity. Treatment of [2](PF6)2 with PPh3 gives [(cymene)2(CH3CN)2(PPh3)Ru3S2](PF6)2 ([3](PF6)2) and [(cymene)2(CH3CN)(PPh3)2Ru3S2](PF6)2 ([4](PF6)2), while treatment with 1,4,7 trithiacyclononane (9S3) gives [(cymene)2(9S3)Ru3S2](PF6)2 ([5](PF6)2). A crystallographic study demonstrated that the Ru3 core in [3](PF6)2, [4](PF6)2, and [5](PF6)2 is distorted with a pair of elongated Ru-Ru bonds. Cyclic voltammetry shows that [3](PF6)2 and [4](PF6)2 undergo two closely spaced reversible one-electron reductions whereas [5](PF6)2 undergoes one irreversible one-electron reduction and one reversible one-electron reduction. Prolonged irradiation of [1](PF6)2 in CH3CN causes decomposition, resulting in the pentanuclear cluster [(cymene)4Ru5S4](PF6)2 ([6](PF6)2). PMID- 11261952 TI - Time-resolved optical and infrared spectral studies of intermediates generated by photolysis of trans-RhCl(CO)(PR3)2. Roles Played in the Photocatalytic Activation of Hydrocarbons. AB - Described are picosecond and nanosecond time-resolved optical (TRO) spectral and nanosecond time-resolved infrared (TRIR) spectral studies of intermediates generated when the rhodium(I) complexes trans-RhCl(CO)L2 (L = PPh3 (I), P(p tolyl)3 (II), or PMe3 (III)) are subjected to photoexcitation. Each of these species, which are precursors in the photocatalytic activation of hydrocarbons, undergoes CO labilization to form an intermediate concluded to be the solvated complex RhCl(Sol)L2 (A(i)). The picosecond studies demonstrate that an initial transient is formed promptly (<30 ps), which decays to A(i) with lifetimes ranging from 40 to 560 ps depending upon L and the medium. This is proposed on the basis of ab initio calculations to be a metal-to-ligand charge transfer (MLCT) excited state. Second-order rate constants (kCO) for reaction of the A(i) with CO were determined, and these depend on the nature of L and the solvent, the slowest rate being for A(I) in tetrahydrofuran (kCO = 7.1 x 10(6) M(-1) x s(-1)), the fastest being for A(III) in dichloromethane (1.3 x 10(9) M(-1) x s(-1)). Each A(i) also undergoes competitive unimolecular reaction with solvent to form long lived transients with TRIR properties suggesting these to be Rh(III) products of oxidative addition. Although this was mostly suppressed by the presence of higher concentrations of CO (which trapped A(i) to re-form the starting complexes in each case), both TRO and TRIR experiments indicate that a fraction of the oxidative addition could not be quenched. Thus, the short-lived MLCT state or a vibrationally hot species formed during the decay of this excited state appears to participate directly in C-H activation. PMID- 11261953 TI - Synthesis and structural characterization of Zn(O3PCH2OH), a new microporous zinc phosphonate. AB - Zn(O3PCH2OH) (1) has been formed by reaction of zinc acetate with diethyl hydroxymethylphosphonate. The acidity of the zinc solution effects hydrolysis of the phosphonate to produce phosphonic acid in situ. 1 crystallizes in the trigonal spacegroup R3, with a = 15.9701(2) A, c = 7.783(2) A, and Z = 18. The compound has channels in the [001] direction, formed by phosphonate groups bridging the octahedral coordinated zinc atoms. The zinc atoms are coordinated by the three oxygens of the phosphonate group and the oxygen of the hydroxy group. PMID- 11261954 TI - Optical dimer excitations and exchange parameters in (Et4N)3Cr2F9: first observation of the 4A2 --> 2A1 transition. AB - The synthesis, crystal growth, and polarized optical absorption spectra in the visible and near-UV of (Et4N)3Cr2F9 are reported. In the energy range 25800-27700 cm(-1) the 4A2 --> 2A1 (O notation) ligand field transition can be resolved in detail for the first time in any Cr3+ compound. This allows the determination of the antiferromagnetic ground-state exchange splitting with great accuracy: J = 25.9 cm(-1) and j = 0.27 cm(-1) using the Hamiltonian H = J(S(A).S(B)) - j(S(A).S(B))2, where j leads to deviations from the regular Lande pattern. The temperature dependence of the magnetic susceptibility is nicely reproduced by these parameters. A comparison with Cs3Cr2Cl9 and Cs3Cr2Br9 reveals an exponential dependence of the ground-state splitting upon the Cr-Cr distance in the [Cr2X9]3- dimers. This is the result of a dominant sigma-type orbital exchange pathway along the Cr-Cr axis. PMID- 11261955 TI - Synthesis and reactivity of perhalogenated acyclic and metallacyclic tantalum(V) phosphoraniminato complexes: discovery of an unexpected ligand coupling reaction to form the novel phosphazenium salt. AB - The reaction of TaCl5 with a single equivalent of Cl3P=NSiMe3 resulted in the isolation of the perhalogenated (phosphoraniminato) tantalum(V) complex TaCl4(N=PCl3) (1). Reaction of 1 with an excess of THF and subsequent cooling produced crystals of TaCl4(N=PCl3)(THF) (1.THF), which possesses a distorted octahedral Ta center with a THF molecule coordinated trans to the phosphoraniminato ligand. The reaction of 1 with the aminophosphoranimine, (Me3Si)2NPCl2=NSiMe3, resulted in a [3 + 1] cyclocondensation reaction to form the metallacyclic complex, TaCl3(N=PCl3)[N(SiMe3)PCl2N(SiMe3)] (2), which contains a TaNPN four-membered ring and a phosphoraniminato ligand (N=PCl3). The analogous [3 + 1] cyclocondensation reaction between (Me3Si)2NPCl2=NSiMe3 and TaCl5 led to the isolation of TaCl4[N(SiMe3)PCl2N(SiMe3)] (3). An attempt to cleave the NPN ligand from the Ta center in 2 via protonolysis with HCl led to an unusual phosphoraniminato ligand coupling reaction to yield the novel phosphazenium salt [N(PCl2NH2)2][TaCl6] (4). All new compounds (1.THF and complexes 1-4) were characterized by single-crystal X-ray diffraction. PMID- 11261956 TI - Platinum(IV) complexes with dipeptide. X-ray crystal structure, 195Pt NMR spectra, and their inhibitory glucose metabolism activity in Candida albicans. AB - Three dipeptide complexes of the form K[Pt(IV)(dipep)Cl3] and two complexes of the form K[Pt(IV)(Hdipep)Cl4] were newly prepared and isolated. The platinum(IV) complexes containing the dipeptide were obtained directly by adding KI to H2[PtCl6] solution. The reaction using KI was rapidly completed and provided analytically pure yellow products in the form of K[Pt(dipeptide)Cl3] for H2digly, H2gly(alpha)-ala, H2alpha-alagly and H2di(alpha)-ala. The K[Pt(IV)(digly)Cl3] complex crystallizes in the monoclinic space group P2(1)/c with unit cell dimensions a = 10.540(3) A, b = 13.835(3) A, c = 8.123(3) A, beta = 97.01(2) degrees, Z = 4. The crystal data represented the first report of a Pt(IV) complex with a deprotonated peptide, and this complex has the rare iminol type diglycine(2-) coordinating to Pt(IV) with the bond lengths of the C2-N1 (amide) bond (1.285(13) A). The 195Pt NMR peaks of the K[Pt(IV)(dipep)Cl3] and the K[Pt(IV)(Hdipep)Cl4] complexes appeared at about 270 ppm and at about -130 ppm, respectively, and were predicted for a given set of ligand atoms. While the K[Pt(IV)(x-gly)Cl3] complexes, where x denotes the glycine or alpha-alanine moieties, were easily reduced to the corresponding platinum(II) complexes, the K[Pt(IV)(x-alpha-ala)Cl3] complexes were not reduced, but the Cl- ion was substituted for OH- ion in the reaction solution. The K[Pt(digly)Cl3] and K[Pt(gly-L-alpha-ala)Cl3] complexes inhibited the growth of Candida albicans, and the antifungal activities were 3- to 4-fold higher than those of cisplatin. The metabolism of glucose in C. albicans was strongly inhibited by K[Pt(digly)Cl3] and K[Pt(gly-L-alpha-ala)Cl3] but not by the antifungal agent fluconazole. PMID- 11261957 TI - Synthesis and Structure of [DB24C8)Na][Cd(SCN)3. Formation of a novel linear Cd...Cd...Cd chain with a mer-CdN3S3 coordination confirmation and new coiled [(DB24C8)Na]+ cation. AB - We report herein a novel coordination solid, [(DB24C8)Na][Cd(SCN)3] (6) (DB24C8 denotes dibenzo-24-crown-8), which exhibits a new type of [Cd(SCN)3-]infinity chain with two unusual stereochemical characteristics: (1) a mer-CdN3S3 coordination and (2) a linear Cd chain with a Cd...Cd...Cd angle of 180 degrees. In addition, the [(DB24C8)Na]+ monocation adopts a new structural type-a coiled structure-for the combination of crown ether DB24C8 and alkali metal Na+. The title compound crystallizes in a monoclinic unit cell of C2/c space group symmetry with lattice parameters a = 16.110(8) A, b = 20.380(5) A, c = 11.01(1) A, beta = 119.87(3) degrees, and Z = 4. The arrangement of the [Cd(SCN)3 ](infinity) chains in the crystal lattice in the title compound is approximately hexagonal, creating triangular channels which are filled with [(DB24C8)Na]+ monocations. It was previously reasoned by us that the coiled [(DB24C8)Na]+ monocation, which lacks inversion or mirror symmetries, should enhance the tendency for the formation of the noncentrosymmetric space group of the title crystal, making it a potential second-order nonlinear optical crystal. Interestingly, however, the title compound crystallizes in a centrosymmetric space group (C2/c) and gives rise to no second harmonic generation (SHG). Previously known [Cd(SCN)3-](infinity) chains adopt fac-CdN3S3 coordination and a zigzag Cd chain configuration with a Cd...Cd...Cd angle of 165 degrees. The zigzag chains can align in either parallel or antiparallel fashion, resulting in efficient or no SHG effects, respectively. The linear Cd.Cd.Cd chain configuration observed in the title compound, on the other hand, makes it indistinguishable between parallel and antiparallel alignments. It is concluded that, to ensure the formation of noncentrosymmetric space groups, it is necessary to employ optically pure chiral cations as spacers and/or controllers. Furthermore, to enhance the nonlinear optical responses, [Cd(SCN)3-]infinity chains with fac-CdN3S3 coordination and parallel alignments of the zigzag Cd chains should be used. PMID- 11261958 TI - A synthetic, structural, magnetic, and spectral study of several [Fe[tris(pyrazolyl)methane]2](BF4)2 complexes: observation of an unusual spin state crossover. AB - The complexes [Fe[HC(3,5-Me2pz)3]2](BF4)2 (1), [Fe[HC(pz)3]2](BF4)2 (2), and [Fe[PhC(pz)2(py)]2](BF4)2 (3) (pz = 1-pyrazolyl ring, py = pyridyl ring) have been synthesized by the reaction of the appropriate ligand with Fe(BF4)2.6H2O. Complex 1 is high-spin in the solid state and in solution at 298 K. In the solid phase, it undergoes a decrease in magnetic moment at lower temperatures, changing at ca. 206 K to a mixture of high-spin and low-spin forms, a spin-state mixture that does not change upon subsequent cooling to 5 K. Crystallographically, there is only one iron(II) site in the ambient-temperature solid-state structure, a structure that clearly shows the complex is high-spin. Mossbauer spectral studies show conclusively that the magnetic moment change observed at lower temperatures arises from the complex changing from a high-spin state at higher temperatures to a 50:50 mixture of high-spin and low-spin states at lower temperatures. Complexes 2 and 3 are low-spin in the solid phase at room temperature. Complex 2 in the solid phase gradually changes over to the high-spin state upon heating above 295 K and is completely high-spin at ca. 470 K. In solution, variable-temperature 1H NMR spectra of 2 show both high-spin and low-spin forms are present, with the percentage of the paramagnetic form increasing as the temperature increases. Complex 3 is low-spin at all temperatures studied in both the solid phase and solution. An X-ray absorption spectral study has been undertaken to investigate the electronic spin states of [Fe[HC(3,5-Me2pz)3]2](BF4)2 and [Fe[HC(pz)3]2](BF4)2. Crystallographic information: 2 is monoclinic, P2(1)/n, a = 10.1891(2) A, b = 7.6223(2) A, c = 17.2411(4) A, beta = 100.7733(12) degrees, Z = 2; 3 is triclinic, P1, a = 12.4769(2) A, b = 12.7449(2) A, c = 13.0215(2) A, alpha = 83.0105(8) degrees, beta = 84.5554(7) degrees, gamma = 62.5797(2) degrees, Z = 2. PMID- 11261959 TI - Syntheses of hydrated molybdenum bronzes by reduction of MoO3 with NaBH4. AB - Hydrated molybdenum bronzes have been prepared by reduction reaction of MoO3 with NaBH4 in ethanol and DMSO. The reduction reactions in both solvents occur smoothly; thus, the layered structure of MoO3 is maintained in the product. Divalent cation Ca2+ has been intercalated between the MoO3 layers, which leads to highly reduced molybdenum bronze (Mo5.26+). Solvated molybdenum bronze catalyzes the reduction reaction of DMSO by NaBH4, producing CH3SCH3. The structure model of hydrated sodium molybdenum bronze has also been reinvestigated by using the Rietveld analysis. The hydrated molybdenum bronze crystallizes in an orthorhombic structure, in which the structure of Mo octahedron layers is closely related to that in MoO3. However, the structure refinement reveals that the Mo octahedron in the MoO3 layers is axially distorted, which is different from that in MoO3 but similar to an isoelectron compound H0.33MoO3. PMID- 11261960 TI - Effects of sequential replacement of -NH2 by -OH in the tripodal tetraamine tren on its acidity and metal ion coordinating properties. AB - The preparation is described of two modified derivatives of the tripodal tetraamine tren, 2-hydroxy-N,N-bis(2-aminoethyl)ethylamine, NN(2)O222, and 2 amino-N,N-bis(2-hydroxyethyl)ethylamine, NNO(2)222, in which one and two primary amines, respectively, have been replaced with hydroxyl groups. The aqueous acid base and metal ion (Ni2+, Cu2+, Zn2+) coordination properties of these two compounds were studied by potentiometric, spectrophotometric, and NMR titrations. Two and three acidity constants, respectively, were determined for NNO(2)222 and NN(2)O222 by potentiometry. NMR titrations proved that deprotonation of the two OH residues in NNO(2)222, and of the one in NN(2)O222, corresponded to pK(a) > 14. Acidity constants related to deprotonation of the terminal primary amine functions were similar in both NNO(2)222 and NN(2)O222 (and to those in the parent compound tren), whereas deprotonation of the tertiary ammonium N atom had a very different acidity constant in each of these three compounds. Charge repulsion, polar effects, and intramolecular hydrogen bond formation are responsible for the discrepancy. Chelated diamine metal complexes for each ligand studied depended only on the basicity of the corresponding two amines, suggesting that the hydroxyl group interacted with the metal ion very weakly in acidic or neutral solutions. The ML2+ species further deprotonated to form M(L - H)+ and M(L - 2H) complexes, in which the protons are released from the coordinated OH group. A pM vs pH correlation showed that replacing an NH2 group with a OH group in tren or NN(2)O222 makes the resulting metal complex less stable. Electronic spectra showed that the Cu(II) complexes of both NNO(2)222 and NN(2)O222 adopted a square pyramidal geometry rather than a trigonal bipyramidal geometry. The X ray crystal structure analysis of the zinc complex [Zn(OH)(mu-NNO(2)222 - H)Zn(NNO(2)222)]2+, as its [BF4]- salt, shows a dinuclear molecule containing two zinc ions, each coordinated in a distorted trigonal bipyramid. The coordination environment at one zinc atom is composed of the four donor groups of a mono-O deprotonated ligand NNO(2)222 and a hydroxyl ion with the central nitrogen atom of the ligand and the hydroxyl ion in equatorial positions. The oxygen atom of the deprotonated alkoxo group bridges to the second zinc atom, which is coordinated by this atom and one undeprotonated ligand NNO(2)222. PMID- 11261961 TI - Syntheses, crystal structures, magnetic properties, and EPR spectra of tetranuclear copper(II) complexes featuring pairs of "roof-shaped" Cu2X2 dimers with hydroxide, methoxide, and azide bridges. AB - Hydroxo- and methoxo-bridged tetranuclear copper(II) complexes of the tetramacrocyclic ligand 1,2,4,5-tetrakis(1,4,7-triazacyclonon-1-ylmethyl)benzene (Ldur), have been prepared from [Cu4Ldur(H2O)8](ClO4)8.9H2O (1). Addition of base to an aqueous solution of 1 gave [Cu4Ldur(mu2-OH)4](ClO4)4 (2). Diffusion of MeOH into a DMF solution of 2 produces [Cu4Ldur(mu2-OMe)4](ClO4)4.HClO4.2/3MeOH (3), a complex which hydrolyzes on exposure to moisture regenerating 2. The structurally related azido-bridged complex, [Cu4Ldur(mu2-N3)4](PF6)4.4H2O.6CH3CN (4), was produced by reaction of Ldur with 4 molar equiv of Cu(OAc)2.H2O and NaN3 in the presence of excess KPF6. Compounds 2-4 crystallize in the triclinic space group P1 (No. 2) with a = 10.248(1) A, b = 12.130(2) A, c = 14.353(2) A, alpha = 82.23(1) degrees, beta = 80.79(1) degrees, gamma = 65.71(1) degrees, and Z = 1 for 2, a = 10.2985(4) A, b = 12.1182(4) A, c = 13.9705(3) A, alpha = 89.978(2) degrees, beta = 82.038(2) degrees, gamma = 65.095(2) degrees, and Z = 1 for 3, and a = 12.059(2) A, b = 12.554(2) A, c = 14.051(2) A, alpha = 91.85(1) degrees, beta = 98.22(1) degrees, gamma = 105.62(1) degrees, and Z = 1 for 4. The complexes feature pairs of isolated dibridged copper(II) dimers with "roof shaped" Cu2(mu2-X)2 cores (X = OH-, OMe-, N3-), as indicated by the dihedral angle between the two CuX2 planes (159 degrees for 2, 161 degrees for 3, and 153 degrees for 4). This leads to Cu.Cu distances of 2.940(4) A for 2, 2.962(1) A for 3, and 3.006(5) A for 4. Variable-temperature magnetic susceptibility measurements indicate weak antiferromagnetic coupling (J = -27 cm(-1)) for the hydroxo-bridged copper(II) centers in 2 and very strong antiferromagnetic coupling (J = -269 cm(-1)) for the methoxo-bridged copper(II) centers in 3. Pairs of copper(II) centers in 4 display the strongest ferromagnetic interaction (J = 94 cm(-1)) reported thus far for bis(mu2-1,1-azido)-bridged dicopper units. Spectral measurements on a neat powdered sample of 4 at 33.9 GHz or 90 Ghz confirm the spin-triplet ground state for the azido-bridged copper(II) pairs. PMID- 11261962 TI - Density functional study of tetraphenolate and calix[4]arene complexes of early transition metals. AB - Density functional calculations have been performed on some calix[4]arenes complexes of early transition metals. Particular emphasis has been placed on the comparison of the main properties of these metal complexes with the analogous metal complexes based on four monodentate phenolate ligands to study the effect of the geometrical constraints imposed by the calixarenes framework on the electronic structure. The results show that the most stable geometry of titanium and molybdenum tetraphenolates is pseudotetrahedral (slightly flattened for molybdenum) and that the distortion to a square planar coordination requires, respectively, 52.0 and 21.5 kcal x mol(-1). However, a significant energy decrease is found when the four phenolate groups are bent in the same hemisphere, reproducing the calix[4]arene geometry. Such a coordination determines the energy decrease of the unoccupied metal d orbitals of sigma and pi symmetry, which leads to an increase of the electron-accepting properties of these metal fragments. PMID- 11261963 TI - Photochemical and chemical oxidation of alpha-dimine-dithiolene metal complexes: insight into the role of the metal atom. AB - [Pd(bpy)(bdt)], 2 (bpy = 2,2'-bipyridine, bdt = 1,2-benzenedithiolate), was prepared in good yield by the reaction of bdtNa2 with [(bpy)PdCl2] in DMSO. The analogous nickel complex, 1, was prepared in a similar reaction using MeOH/CH2Cl2 and [(bpy)NiCl2.dmf]2. Both 1 (a = 7.9920(1) A, b = 11.4385(1) A, c = 16.1415(1) A, beta = 103.327(1) degrees, V = 1435.86(2) A3, Z = 4) and 2 (a = 8.1631(5) A, b = 11.4379(7) A, c = 16.2475(10) A, beta = 103.7010(10) degrees, V = 1473.84(12) A3, Z = 4) crystallize in the monoclinic space group P2(1)/c and are isostructural with their previously reported platinum analogue. In accord with the results observed for platinum but not nickel, photochemical oxidation of 2 in DMF provides the monosulfinate complex [Pd(bpy)(bdtO2)], 4, along with a minor amount of the corresponding disulfinate [Pd(bpy)(bdtO4)], 5, while chemical oxidation yields only the latter. 4 cocrystallizes with 5 in the monoclinic space group P2(1)/c (a = 8.026(3) A, b = 14.600(6) A, c = 13.371(3) A, beta = 101.80(3) degrees, V = 1533.8(9) A3, Z = 4) as does pure 5 (a = 8.5611(9) A, b = 14.4586(15) A, c = 13.3677(14) A, beta = 108.122(2) degrees, V = 1572.6(3) A3, Z = 4). Comparison of spectroscopic and electrochemical properties of the three complexes, [M(bpy)(bdt)], yields the following ordering for the energy of the HOMO: Pd < Ni < Pt. The observed reactivity patterns and the electronic data suggest that the "anomalous" reactivity of 1 be attributed to the greater relative flexibility of the coordination geometry for nickel(II) complexes rather than electronic differences such as the energies of the frontier orbitals. PMID- 11261964 TI - Conformational/configurational analysis of all the binding geometries of cobalt(III) bleomycin. AB - No crystal structure of metallobleomycin (BLM) exists, and the exact coordination mode of the ligand is unknown. To date, spectroscopic investigations of BLM complexes and crystal structures of BLM models have been used to propose its metal coordination sites. This has led to contradictory interpretations of the metal coordination sphere in BLM. Inorganic molecular mechanics and configurational/conformational searches were used to analyze HOO-CoBLM A2, H2O CoBLM A2, and HOO-CoPEP with commonly proposed binding geometries. The lowest energy binding geometry found was one with the mannose carbamoyl bound to the cobalt ion. The Monte Carlo dihedral and translational variational searches were able to find most of the configurations available to cobalt(III) bleomycin in the three binding geometries examined. PMID- 11261965 TI - Preparation of site-differentiated mixed ligand and mixed ligand/mixed metal metallacrowns. AB - Assembly reactions that can prepare reliably regioselective metallamacrocyclic complexes have been a target in the development of metallacrowns. To this end, a series of mixed ligand and mixed ligand/mixed metal metallacrowns have been synthesized in high yield and structurally characterized. Two distinct connectivities have been observed in these types of metallacrowns. The monomeric, vacant metallacrown with mixed ligand composition [12-MC(Ni(II)N(Hshi)2(pko)2-4)] (1a) shows the connectivity pattern [-O-Ni-O-N-Ni-N-]2 while the other Ni metallacrowns, [12-MC(Ni(II)N(shi)2(pko)2-4)] (2a) and the coupled [12 MC(Ni(II)N(shi))2(pko)2-4)][12-MC(Ni(II)N(shi))3(pko)-4)] (3a) fused metallacrowns as well as the mixed metal Mn-Ni metallacrown [12 MC(Ni(II)Mn(III)N(shi)2(pko)2-4)] (4a), follow the pattern [-Ni-O-N-]4. Also, three distinct arrangements of the chelate rings around the metal ions have been observed. The syntheses are completely general, allowing for the substitution of different ligands into the metallacrown core. Compounds 1 and 4 show the 6-5-6-5 6-5-6-5 arrangement, compounds 2 and 3(1) the 6-6-5-5-6-6-5-5, and the 3(2) component the 6-6-5-5-6-5-6-5. The obtained structures can be rationalized by balancing the charge at each metal site in the metallacrown. Variable temperature magnetic susceptibility measurements show that exchange interactions for all the compounds are weak and dominantly antiferromagnetic (e.g., 2a gives an exchange coupling of J = -1.2 cm(-1) with g = 2.2). In solution, the metallacrowns are shown to be stable both to decomposition and ligand exchange. PMID- 11261966 TI - Assembly of the inorganic double helix Mg6(Et2NCO2)12 by fixation of carbon dioxide: crystal and solution structure. AB - The homoleptic magnesium carbamato complex Mg6(Et2NCO2)12, 1 (Et2NCO2- = diethylcarbamato anion), was prepared by the reaction of dibutylmagnesium with diethylamine, followed by carboxylation using gaseous carbon dioxide. Crystallographic characterization demonstrated that 1 has the standard M6(R2NCO2)12 structure and is a double helix of MgO(x)(x = 5, 6) coordination polyhedra with Delta or Lambda stereochemistry arising from the configuration around the six-coordinate Mg2+ cations. It crystallized in the orthorhombic space group Ccca with two molecules of Delta1 and two of Lambda1 per unit cell (a = 21.548 A, b = 25.094 A, c = 15.4485(11) A, alpha = beta = gamma = 90 degrees ). Extensive solution characterization of 1 by 1-dimensional proton and 13C NMR spectroscopy and by two-dimensional 1H-[13C] NMR correlation techniques verified that the helical structure is maintained in solution. Moreover, these measurements indicated that the intramolecular dynamics of 1 relating to motions of the ethyl groups was substantially hindered in solution. Correlation of the crystallographic and NMR structural studies indicated that this arises from a combination of hindered rotation about the carbamato C-N bond and efficient packing of the ethyl groups around the Mg6O24 core. The result is an inverted micelle-like structure for 1 in which the hydrophobic ethyl groups form a sheath largely restricting access to the hydrophilic Mg6O24 core. PMID- 11261967 TI - Dimeric W3SO3 cluster complexes: synthesis, characterization, and potential applications as X-ray contrast agents. AB - Our continued research on the use of heavy metal cluster complexes as a new class of X-ray contrast agents in medical diagnostic imaging is described. A series of 2:3 cluster-ligand complexes, [(W(IV)3SO3)2L3]4- (L = linear polyaminopolycarboxylate ligands), were isolated from the reaction of aqua ion [W(IV)3SO3(H2O)9]4- (prepared in large quantities through an improved literature process) with respective ligands in refluxing DMF. The salts of [(W(IV)3SO3)2L3]4 complex anions were fully characterized using routine techniques such as elemental analysis, MS, HPLC, UV-vis, IR, and NMR. The solid structures of two complex anions, [(W(IV)3SO3)2(PDTA)3]4- and [(W(IV)3SO3)2(HO-PDTA)3]4-, were determined by X-ray crystallography. They are the first examples wherein two W(IV)3SO3 clusters are complexed and linked by three ligands that contain two terminal iminodiacetate (bis-IDA) groups. Complexation of the unstable aqua ion [W(IV)3SO3(H2O)9]4- with ligands has imparted desired biological compatibility to the tungsten metal cluster. These complexes are stable and highly soluble in H2O. The potential utility of such tungsten cluster complexes as X-ray contrast agents was evaluated in both in vitro and in vivo animal studies. In addition, the syntheses of several new linear polyaminopolycarboxylate ligands used in this study are reported. PMID- 11261968 TI - Charge distribution in bis-dioxolene radical metal complexes. synthesis and DFT characterization of dinuclear Co(III) and Cr(III) complexes with a mixed-valent, S = 1/2 semiquinone-catecholate ligand. AB - Bis-dioxolene bridged dinuclear metal complexes of general formula M2(CTH)2(diox diox)(PF6)n (n = 2, 3; M = Co(III), Cr(III); CTH = tetraazamacrocycle) have been synthesized using the bis-bidentate ligand 5,5'-di-tert-butyl-3,3',4,4' tetrahydroxybiphenyl. These complexes were characterized by means of ESR, UV-vis, temperature dependent magnetic susceptibility, and cyclic voltammetry. Our results unambiguously suggest that the tripositive dimetal cations can be described as containing a fully delocalized bis-dioxolene trinegative radical ligand (Cat-Sq) bridging two tripositive metal cations. In this frame the sextet electronic ground state characterizes the Cr2(CTH)2(Cat-SQ)3+ as a result of the antiferromagnetic coupling of the radical bridging ligand with the two equivalent paramagnetic metal centers. The electronic and geometrical structure and the magnetic properties of Cat-Sq and of its complexes have been studied with density functional theory. PMID- 11261969 TI - Chloroform-soluble schiff-base Zn(II) or Cd(II) complexes from a dynamic combinatorial library. AB - A dynamic combinatorial library of metal ion Schiff-base complexes have been studied for the extraction of Zn(II) or Cd(II) from aqueous solution into chloroform. Library components consist of different aminophenols and 2 pyridinecarboxaldehyde. Extraction of both Zn(II) and Cd(II) into chloroform was observed from aqueous solutions containing 0.0500 mM M(NO3)2, 0.100 M aminophenol, 0.100 M 2-pyridinecarboxaldehyde, 0.100 M NaCl, and 5.00 mM buffer at pH 8.5. Extraction was dependent on pH but not on counterions including Cl-, Br-, or NO3-. Studies showed that equilibrium was attained between the Schiff base complexes across the two-phase chloroform-water system after 24 h of stirring. Analysis of the extracted species by use of 1H NMR spectroscopy and mass spectrometry as well as solubility studies on characterized complexes suggested that the major extracted species is the neutral bis-Schiff-base metal ion complex. In libraries containing mixtures of two different aminophenols and 2 pyridinecarboxaldehyde, an enhanced extent of extraction of Zn(II) into chloroform is observed. Studies suggest that a Zn(II) complex, which is likely the mixed Schiff-base complex, has superior extraction properties compared to simple libraries with a single aminophenol component. The structures of two bis Schiff-base complexes of Zn(II) and one of Cd(II) have been determined by X-ray crystallography. The geometries of the two Zn(II) complexes, which differ only by a methyl substituent on the Schiff-base ligand, are markedly different, supporting the use of combinatorial methods in coordination chemistry. Zn(SB14)2 crystallized as the sesquihydrate (C24H18N4O2Zn.1.5 H2O) in the space group C2/c, with cell dimensions a = 23.219(15) A, b = 11.299(7) A, c = 16.822(11) A, beta = 102.91(5) degrees, V = 4302(5) A3, and Z = 8. Zn(SB15)2 crystallized as a 1:1 methanol solvate (C26H22N4O2Zn.CH3OH) in the space group P2(1)/c with cell dimensions a = 13.981(5) A, b = 7.978(3) A, c = 22.568(8) A, beta = 104.53(3) degrees, V = 2436.8(15) A3, and Z = 4. Cd(SB14)2 crystallized as a 1:1 ethanol solvate (C24H18N4O2Cd.CH3CH2OH) in the space group R3 with unit cell dimensions of a = 36.423(2) A, c = 9.2930(10) A, V = 10678(2) A3, and Z = 18. PMID- 11261970 TI - Synthesis of beta-P,N aminophosphines and coordination chemistry to Pd(II). X-ray structures of [PdCl2(Ph2PCH(Ph)NHPh-kappaP,kappaN)] and PdCl(eta3 C3H5)(Ph2PCH2CH(Ph)NHPh-kappaP)]. AB - The reaction of the C=N bond in PhCH=NPh with the carbanionic species Ph2PCH2-, leading to the N-phenyl beta-aminophosphine Ph2PCH2CH(Ph)NHPh, L1, is described. This molecule reacts with different organic electrophiles to afford related compounds Ph2PCH2CH(Ph)NPhX (X = SiMe3, L2; COPh, L4), [Ph2MePCH2CH(Ph)NHPh]+(I ), L3, and [Ph2PCH2CH(Ph)N(Ph)CO]2, L5, containing two amido and two phosphino functions. The coordination properties of L1, L2, and L4 have been studied in palladium chemistry. The X-ray structure of [PdCl2(Ph2PCH2CH(Ph)NHPh kappaP,kappaN)] shows the bidentate coordination mode for the L1 ligand with equatorial C(Ph)-N(Ph) phenyl groups. [PdCl2(Ph2PCH2CH(Ph)NHPh-kappaP,kappaN)] crystallizes at 298 K in the space group P2(1)/n with cell parameters a = 10.689(2) A, b = 21.345(3) A, c = 12.282(2) A, beta = 90.294(12) degrees, Z = 4, D(calcd) = 1.526. The reaction between 2 equiv of L1 and [PdCl(eta3-C3H5)]2 affords the [PdCl(eta3-C3H5)(Ph2PCH2CH(Ph)NHPh-kappaP)] complex in which an unexpected N-H.Cl intramolecular interaction has been observed by an X-ray diffraction analysis. [PdCl(eta3-C3H5)(Ph2PCH2CH(Ph)NHPh-kappaP)] crystallizes at 298 K in the monoclinic space group Cc with cell parameters a = 10.912(1) A, b = 17.194(2) A, c = 14.169(2) A, beta = 100.651(9) degrees, Z = 4, D(calcd) = 1.435. Neutral and cationic alkyl or allyl palladium chloride complexes containing L1 are also reported as well as a neutral allyl palladium chloride complex containing L4. Variable-temperature 31P[1H] NMR studies on the allyl complexes show that the eta3/eta1 allyl interconversion is enhanced by a positive charge and also by a N-H.Cl intramolecular interaction. PMID- 11261971 TI - Structural and spectroscopic studies of the versatile coordination chemistry of the chiral ligand N,N-bis(1-propan-2-onyl oxime)-L-methionine N'-methylamide with Ni(II) and Zn(II). AB - The potentially pentadentate, chiral ligand N,N-bis(1-propan-2-onyl oxime)-L methionine N'-methylamide (L-MABO) shows remarkable versatility in its coordination chemistry with Ni(II) and Zn(II). In the crystal structure of the ZnCl2 complex of L-MABO, the ligand coordinates to the metal only through its three nitrogen donor groups (one amine and two oximes), with two chloride anions completing the distorted trigonal bipyramidal coordination sphere. In the NiCl2 complex, the three nitrogen donors and the thioether sulfur coordinate, along with two chlorides. The crystal structure of the Ni(NO3)2 complex contains two independent molecules, one of which coordinates the three nitrogens, the thioether sulfur, and the amide oxygen of L-MABO in addition to one nitrate anion. The second molecule coordinates the three nitrogen donors, the amide oxygen, one nitrate anion, and a methanol molecule. Thus, in only three crystal structures, L-MABO demonstrates its ability to provide N3, N3S, N3O, and N3OS donor sets. The thioether-bound complexes are unusual in that they have a predominantly nitrogen environment with a nickel-thioether bond that is not constrained by surrounding donor groups in a macrocyclic or linear polydentate motif. Comparison of the thioether-coordinated and methanol-coordinated molecules in the Ni(NO3) salt of L-MABO demonstrate the effect of the thioether bond on the relative "hardness" of the nickel. The electronic absorption and circular dichroism spectra of the aqueous solutions of the nickel complexes are interpreted in terms of a "descent in symmetry" model based on successive C3v and Cs distortions from octahedral geometry. These ligand field spectra indicate that in aqueous solution all ligand groups except for the three nitrogens of L-MABO are displaced by water. In acetonitrile, the non-nitrogen donors in the nitrate salt may also be displaced, while the chlorides remain coordinated. PMID- 11261972 TI - Electron/atom transfer in halo-bridged homobimetallic complexes. structure and donor-acceptor properties of face-to-face dicopper complexes with teraazamacrocyclic ligands. AB - The syntheses and donor-acceptor properties of some novel, halo-bridged dicopper(II) complexes of alpha,alpha'-bis(5,7-dimethyl-1,4,8,11 tetraazacyclotetradecane-6-yl)-o-xylene are reported. These complexes were characterized by their magnetic and electrochemical behavior, X-ray structure analysis, FAB mass spectroscopy, and electronic spectra. The bromo-bridged complex crystallized in the tetragonal system, space group P4(3)2(1)2, with a = 12.6584(5) A, c = 28.6483 (14) A, Z = 4, R = 0.071, and Rw = 0.147. The chloro bridged complex crystallized in the monoclinic system, space group C2/c, with a = 32.749(2) A, b = 18.8915(9) A, c = 26.022(2) A, beta =114.831 degrees, Z = 12, R = 0.080, and Rw = 0.132. Both molecules have C2 symmetry. The two copper(II) ions are axially bridged by a bromine or a chlorine, and the two macrocycles are bridged by an o-xylene group. Each complex displays a cofacial ring arrangement. The Cu-X distance (where X = Cl, Br) is shorter than the sum of van der Waals radii of Cu and X. The phenyl ring is approximately orthogonal to the Cu-X-Cu axis. The nonhalo-bridged complex has a significant affinity for halides (Kf approximately 10(4) M(-1)). The chloride-bridged complex had barely resolved differential pulse polarographic waves (DeltaE1/2 approximately 28 mV), while the bromide-bridged complex exhibited two CV waves in the 1.0-1.5 V range (DeltaE1/2 = 0.24 V). All the Cu(II)/Cu(I) couples were irreversible with a cathodic peak at about - 0.9 V. The magnetic susceptibility results below 20 K follow Curie-Weiss behavior, indicating that the magnetic interaction between the two Cu centers is weakly antiferromagnetic with J < or = -1 cm(-1) for all three complexes. A bridging-ligand-mediated superexchange model is used to treat the magnetic and electron-transfer coupling in the Cu(II)(X-)Cu(II) complexes. A single set of perturbation theory parameters is consistent with the magnetic and electrochemical observations on the chloride-bridged complex and the magnetic properties of the bromide-bridged complex. The electrochemical behavior of the latter suggests a relatively low-energy, high-spin configuration for the Cu(III)(Br-)Cu(II) complex. The analysis attributes the weak Cu(II)/Cu(II) coupling to the orthogonality of the donor and acceptor orbitals to the bridging axis. It is inferred that bridging halide-mediated metal-metal dsigma/psigma coupling significantly alters the chemical properties of the bimetallic complexes only when the donor and acceptor orbitals are coaxial with the bridging ligand. In such a limit, the coupling takes the form of a three-center bonding contribution. PMID- 11261973 TI - Nickel(II) and zinc(II) dibenzoylmethanates: molecular and crystal structure, polymorphism, and guest- or temperature-induced oligomerization. AB - Four forms of nickel(II) and two of zinc(II) dibenzoylmethanates have been isolated and characterized with powder and single-crystal X-ray diffraction analyses, differential scanning calorimetry, magnetic susceptibility measurements, and solid-state 13C cross-polarization/magic angle spinning NMR. Nickel dibenzoylmethanate, Ni(DBM)2 (DBM = PhCOCHCOPh-), forms three polymorphic forms (light-green, brown, and green) and a fourth clathrate form with guest benzene included. The light-green polymorph is metastable. Substituted benzenes induce recrystallization of the polymorph into a stable brown form (C30H22NiO4; a = 26.502(3) A, b = 5.774(1) A, c = 16.456(2) A, beta = 116.03(1) degrees; monoclinic, C2/c; Z = 4). Unlike the other forms, the brown form is diamagnetic and is comprised of monomers of the low-spin [Ni(DBM)2] complex. The Ni(II) is chelated by two DBM ligands in a square planar environment by four donor oxygen atoms. When heated, the brown form transforms to a green form which is stable above 202 degrees C (C90H66Ni3O12; a = 13.819(2) A, b = 16.252(2) A, c = 17.358(2) A, beta = 108.28(1) degrees; monoclinic, P2(1)/n; Z = 2). This polymorph is formed by van der Waals packing of trimers [Ni3(DBM)6] containing linear Ni3 clusters with an Ni-Ni distance of 2.81 A. The cluster is surrounded by six DBM ligands, providing a distorted octahedral environment about each Ni by six oxygen atoms. Benzene stabilizes the trimeric structure at room temperature, forming a [Ni3(DBM)6].2(benzene) inclusion compound (Ni-Ni distance of 2.83 A) with guest benzene molecules located in channels (C90H66Ni3O12 + 2(C6H6); a = 17.670(2) A, b = 20.945(3) A, c=11.209(2) A, beta = 102.57(1) degrees; monoclinic, P2(1)/c; Z = 2). Zinc dibenzoylmethanate has been prepared in two polymorphic forms. The monomeric form contains [Zn(DBM)2] molecules with the zinc center in a distorted tetrahedral environment of four oxygens from the two chelated DBMs (C30H22O4Zn; a = 10.288(2) A, b = 10.716(2) A, c = 12.243(2) A, alpha = 89.19(1) degrees, beta = 75.39(1) degrees, gamma = 64.18(1) degrees; triclinic, P1; Z = 2). Another, dimeric form contains [Zn2(DBM)4] species, with two zinc atoms separated by a distance of 3.14 A and each zinc coordinated by five oxygen atoms (C60H44O8Zn2; a = 25.792(3) A, b = 7.274(1) A, c = 24.307(2) A, beta = 90.58(1) degrees; monoclinic, C2/c; Z = 4). The polymorphic variety of the title complexes and the peculiarities of the Ni(II) and Zn(II) coordination environments are discussed in the context of using the complexes as precursors for new metal complex hosts. PMID- 11261974 TI - Interaction of Rh(I) with meso-arylsapphyrins and -rubyrins: first structural characterization of bimetallic hetero-rubyrin complex. AB - The ligational behavior of meso-arylsapphyrins and rubyrins toward Rh(I) is investigated. Sapphyrins form monometallic complexes with coordination of one imine and amine type nitrogens of the bipyrrole unit in an eta2 fashion. The Rh(I) coordination is completed by the presence of two ancillary carbon monoxide ligands. Rubyrins form both monometallic and bimetallic complexes. Two types of bimetallic complexes have been isolated. In the first type, both rhodium atoms are projected above the mean rubyrin plane, while in the second type, one rhodium atom is projected above and the other below the mean plane. Detailed 1H and 2D NMR spectral analyses along with IR and UV-visible spectra of the complexes confirm the proposed binding modes for the rhodium complexes. Furthermore, the single-crystal X-ray analysis of one of the bimetallic complexes of rubyrin shows a bowl-shaped symmetric structure where both Rh(I) atoms are projected above the mean rubyrin plane at an angle of 71.73 degrees. The geometry around each rhodium center is approximately square planar [N1-Rh1-N2, 80.38(9) degrees; C15-Rh1-C16, 86.95(14) degrees; N1-Rh1-C15, 97.13(12) degrees; and N2-RH1-C16, 94.97(12) degrees ]. The omicronbserved distance of 4.313 A between the two rhodium centers reveals very little interaction between the two rhodium atoms. This type of metal binding is accompanied by a 180 degrees ring flip of the heterocyclic ring connecting the two bipyrrole units. In dioxarubyrin, where one of the pyrrole rings of the bipyrrole unit is inverted, Rh(I) binds at the periphery to the pyrrole nitrogen, leaving the rubyrin cavity empty. The absence of one amino and one imino nitrogen on the dipyrromethene subunits in the sapphyrins and rubyrins described here forces Rh(I) to bind to bipyrrole nitrogens. PMID- 11261976 TI - Ion pairing between Cl- or ClO4- and alkali metal complexes of ionophore antibiotics in organic solvents: a multinuclear NMR and FT-IR study. AB - The extent of ion pairing in chloride and perchlorate salts was studied by measurement of the Cl- and ClO4- resonances and the observation of the perchlorate stretching frequency by use of nuclear magnetic resonance (NMR) spectroscopy and Fourier transform infrared spectroscopy (FT-IR), respectively, for a variety of ionophores in various solutions and in large unilaminar vesicles (LUVs). The NMR line widths of chloride and perchlorate were larger in solutions containing the neutral ionophores valinomycin (Val) and nonactin (Non) than in solutions containing the negatively charged ionophores nigericin (Nig), lasalocid (Las), and monensin (Mon). The viscosity-corrected perchlorate NMR line widths in solutions containing Val and Las were significantly negatively correlated (r2 > or = 0.99) with the dielectric constant of the solvent. Solvents with low dielectric constants favored ion pair formation. From methanolic solutions containing the Li+, Na+, K+, and Cs+ salts of Cl- and ClO4-, it was determined that the cation with the highest selectivity for the ionophore affords the most ion pairing. A decrease in pH from 7 to 3 had no significant effect on the NMR line widths of chloride and perchlorate in methanolic solutions containing Val, whereas a similar decrease in pH in a methanolic solution containing Mon caused a 2-fold increase in the line widths. The FT-IR difference spectrum of KClO4 in a methanolic solution containing Val showed splitting at the perchlorate stretching frequency. No band splitting was observed in the FT-IR difference spectrum of KClO(4) in methanolic solutions containing Las. The efflux of 35Cl in LUVs containing the neutral ionophore Val followed first-order kinetics with an efflux constant of 1.70 x 10(-3) x min(-1), as determined by 35Cl NMR spectroscopy. The induction of increased membrane permeability in LUVs by the ionophore was determined to be negligible for Val and Nig by fluorescence spectroscopy. PMID- 11261977 TI - Metal-metal bonded diruthenium(II, III) assemblies with the polycyano anionic linkers N(CN)2-, C(CN)3-, and 1,4-dicyanamido-2,5-dimethylbenzene (DM-dicyd2-): syntheses, structures, and magnetic properties. AB - The new metal-metal bonded diruthenium(II,III) compounds [Ru2(O2CCH3)4(mu L)]infinity (L = N(CN)2-, 1; C(CN)3-, 2) and [[Ru2(O2CCH3)2(mhp)2]2(mu-DM-Dicyd)] (3) (mhp = 2-oxy-6-methylpyridinate, DM-Dicyd = 1,4-dicyanamido-2,5 dimethylbenzene dianion) have been synthesized and fully characterized. Compounds 1 and 2 were synthesized by the reaction of [Ru2(O2CCH3)4(NCCH3)2](BF4) with NaN(CN)2 and KC(CN)3, respectively. The "dimer-of-dimers", 3, was synthesized by a 2:1 reaction of [Ru2(O2CCH3)2(mhp)2(MeOH)](BF4) with [As(Ph)4]2[DM-Dicyd]. Compound 1 crystallizes in the monoclinic space group C2/m with a = 10.174(2) A, b = 13.016(3) A, c = 7.0750(14) A, beta = 101.83(3) degrees, and Z = 2. Compound 2 crystallizes in the orthorhombic space group Fdd2 with a = 29.679(6) A, b = 31.409(6) A, c = 7.3660(15) A, V = 6866(2) A3, and Z = 16. In compound 1, dicyanamide anions (N(CN)2-) bridge the [Ru2(O2CCH3)4]+ units in an end-to-end bridging mode, thereby forming an alternating one-dimensional chain. In compound 2, two cyano groups of tricyanomethanide anion (C(CN)3-) are coordinated to independent [Ru2(O2CCH3)4]+ units to give a chain similar to that found in 1. The Ru-Ru bond distances in 1 and 2 are 2.2788(14) and 2.2756(5) A, respectively, which are typical values for Ru2(O2CR)4Cl and [Ru2(O2CR)4]+ compounds. The Ru-N distances are 2.257(8) A in 1 and 2.259(4) and 2.283(4) A in 2. The temperature dependence of the magnetic susceptibilities of compounds 1-3 reveals a weak antiferromagnetic interaction between Ru2 units (S = 3/2) through each polycyano anionic linker: g = 2.16, zJ = -0.33 cm(-1), D = 63.3 cm(-1) for 1; g = 2.15, zJ = -0.22 cm(-1), D = 58.0 cm(-1) for 2; and g = 2.10, zJ = -0.90 cm(-1), D = 75.0 cm(-1) for 3. PMID- 11261975 TI - Probing the influence of local coordination environment on the properties of Fe type nitrile hydratase model complexes. AB - A series of four structurally related cis-dithiolate-ligated Fe(III) complexes, [Fe(III)(DITpy)2]Cl (1), [Fe(III)(DITIm)2]Cl (2), [Fe(III)(ADIT)2]Cl (3), and [Fe(III)(AMIT)2]Cl (4), are described. The structural characterization of 3 as well as the spectroscopic properties of 3 and 4 has been previously reported. Crystal data for 1, 2, and 4 are as follows: 1.3H2O crystallizes in the orthorhombic space group Pca2(1) with a = 19.800(4) A, b = 18.450(4) A, c = 14.800(3) A, and Z = 8. 2.(1/2)EtOH.1/2H2O crystallizes in the monoclinic space group Cc with a = 24.792(4) A, b = 14.364(3) A, c = 17.527(3) A, beta = 124.91(2) degrees, and Z = 8. 4 crystallizes in the triclinic space group P1 with a = 8.0152(6) A, b = 10.0221(8) A, c = 11.8384(10) A, alpha = 73.460(3) degrees, beta = 71.451(5) degrees, gamma = 72.856(4) degrees, and Z = 2. Complexes 1-4 share a common S2N4 coordination environment that consists of two cis-thiolates, two trans-imines, and two cis-terminal nitrogen donors: Nterm = pyridine (1), imidazole (2), and primary amine (3 and 4). The crystallographically determined mean Fe-S bond distances in 1-4 range from 2.196 to 2.232 A and are characteristic of low-spin Fe(III)-thiolate complexes. The low-spin S = 1/2 ground state was confirmed by both EPR and magnetic susceptibility measurements. The electronic spectra of these complexes are characterized by broad absorption bands centered near approximately 700 nm that are consistent with ligand-to-metal charge-transfer (CT) bands. The complexes were further characterized by cyclic voltammetry measurements, and all possess highly negative Fe(III)/Fe(II) redox couples ( approximately -1 V vs SCE, saturated calomel electrode) indicating that alkyl thiolate donors are effective at stabilizing Fe(III) centers. Both the redox couple and the 700 nm band in the visible spectra show solvent-dependent shifts that are dependent upon the H-bonding ability of the solvent. The implications of these results with respect to the active site of the iron containing nitrile hydratases are also discussed. PMID- 11261978 TI - Gas-phase structures of acetyl peroxynitrate and trifluoroacetyl peroxynitrate. AB - The molecular structures and conformational properties of acetyl peroxynitrate (PAN, CH3C(O)OONO2) and trifluoroacetyl peroxynitrate (FPAN, CF3C(O)OONO2) were investigated in the gas phase by electron diffraction (GED), microwave spectroscopy (MW), and quantum chemical methods (HF/3-21G, HF/6-31G*, MP2/6-31G*, B3PW91/6-31G*, and B3PW91/6-311+G*). All experimental and theoretical methods show the syn conformer (C=O bond of acetyl group syn to O-O bond) to be strongly predominant relative to the anti conformer. The O-NO2 bonds are extremely long, 1.492(7) A in PAN and 1.526(10) A in FPAN, which correlates with their low bond energy and the easy formation of CX3C(O)OO* and *NO2 radicals in the atmosphere. The O-O bonds (1.418(12) A in PAN and 1.408(8) A in FPAN) are shorter than that in hydrogen peroxide (1.464 A). In both compounds the C-O-O-N dihedral angle is close to 85 degrees. PMID- 11261979 TI - Preparation of mononuclear tungsten tris(sulfido) and molybdenum sulfido tetrasulfido complexes with hydridotris(pyrazolyl)borate coligand and conversion of the former into sulfido-bridged bimetallic complex having Pt(mu-S)2WS core. AB - Treatment of [Et4N][(Me2Tp)W(CO)3] (Me2Tp = HB(3,5-dimethylpyrazol-1-yl)3) with S8 in DMF at room temperature afforded a tris(sulfido) complex [Et4N][(Me2Tp)WS3] (1a), while that of [Et4N][TpW(CO)3] (Tp = HB(pyrazol-1-yl)3) in MeCN resulted in the formation of [Et4N][TpWS3] (1b) along with [Et4N]2[[WO(S2)2]2(mu-S)] (6) as a byproduct. Under similar conditions, [Et4N][(Me2Tp)Mo(CO)3] gave a mixture of a sulfido-tetrasulfido complex [Et4N][(Me2Tp)MoS(S4)] (2a) and its monooxo analogue [Et4N][(Me2Tp)MoO(S4)], although a sulfido-tetrasulfido complex [Et4N][TpMoS(S4)] (2b) was exclusively obtained from [Et4N][TpMo(CO)3]. The reaction of 1a with [PtCl2(cod)] (cod = 1,5-cyclooctadiene) in MeCN at room temperature led to the formation of a sulfido-bridged mixed-metal complex [Et4N][(Me2Tp)WS(mu-S)2PtCl2] (10). The structures of new complexes have been determined in detail by the X-ray analyses for 1a.MeCN, 1b, 2a, 2b, 6, and 10. PMID- 11261980 TI - Reduction of (imine)Pt(IV) to (imine)Pt(II) complexes with carbonyl-stabilized phosphorus ylides. AB - A novel method is reported for generation of the difficult-to-obtain (imine)Pt(II) compounds that involves reduction of the corresponding readily available Pt(IV)-based imines by carbonyl-stabilized phosphorus ylides, Ph3P=CHCO2R, in nonaqueous media. The reaction between neutral (imino)Pt(IV) compounds [PtCl4[NH=C(Me)ON=CR1R2]2] [R1R2 = Me2, (CH2)4, (CH2)5, (Me)C(Me)=NOH], [PtCl4[NH=C(Me)ONR2]2] (R = Me, Et, CH2Ph), (R1 = H; R2 = Ph or C6H4Me; R3 = Me) as well as anionic-type platinum(IV) complexes (Ph3PCH2Ph)[PtCl5[NH=C(Me)ON=CR2]] [R2 = Me2, (CH2)4, (CH2)5] and 1 equiv of Ph3P=CHCO2R (R = Me, Et) proceeds under mild conditions (ca. 4 h, room temperature) to give selectively the platinum(II) products (in good to excellent isolated yields) without further reduction of the platinum center. All thus prepared compounds (excluding previously described Delta4-1,2,4-oxadiazoline complexes) were characterized by elemental analyses, FAB mass spectrometry, IR and 1H, 13C[1H], 31P[1H] and 195Pt NMR spectroscopies, and X-ray single-crystal diffractometry, the latter for [PtCl2[NH=C(Me)ON=CMe2]2] [crystal system tetragonal, space group P4(2)/n (No. 86), a = b = 10.5050(10) A, c = 15.916(3) A] and (Ph3PCH2CO2Me)[PtCl3(NCMe)] [crystal system orthorhombic, space group Pna2(1) (No. 33), a = 19.661(7) A, b = 12.486(4) A, c = 10.149(3) A]. The reaction is also extended to a variety of other Pt(II)/Pt(IV) couples, and the ylides Ph3P=CHCO2R are introduced as mild and selective reducing agents of wide applicability for the conversion of Pt(IV) to Pt(II) species in nonaqueous media, a route that is especially useful in the case of compounds that cannot be prepared directly from Pt(II) precursors, and for the generation of systematic series of Pt(II)/Pt(IV) complexes for biological studies. PMID- 11261981 TI - Oxidation of 7-deazaguanine by one-electron and oxo-transfer oxidants: mismatch dependent electrochemistry and selective strand scission. AB - Addition of oligonucleotides containing 7-deazaguanine (Z) to solutions containing Ru(dmb)3(2+) (dmb = 4,4'-dimethyl-2,2'-bipyridine) produces an enhancement in the oxidative current in the cyclic voltammogram of the metal complex that can be used, through digital simulation, to determine the rate of oxidation of 7-deazaguanine by Ru(dmb)3(3+). The measured rate constants are about 10 times higher than those for oxidation of guanine by Ru(bpy)3(3+), even though the redox potential of Ru(dmb)3(3+/2+) is 200 mV lower. A potential of 0.75 V (vs Ag/AgCl) can therefore be estimated for the oxidation of 7 deazaguanine, which can be selectively oxidized over guanine when Ru(dmb)3(3+) is the oxidant. The rate of oxidation was much faster in single-stranded DNA, and the difference between rates of single-stranded and duplex DNA was higher than for guanine. The oxidation rate was also sensitive to the presence of a single base mismatch at the 7-deazaguanine in the order Z.C < Z.T < Z.G approximately Z.A < single-stranded. The Z.T mismatch was much more readily distinguished than the G.T mismatch, consistent with the overall greater sensitivity to secondary structure for Z. The oxidation reaction was also probed by monitoring piperidine labile cleavage at the Z nucleotide, which could be generated by treatment with either photogenerated Ru(bpy)3(3+) or the thermal oxidant Ru(tpy)(bpy)O2+ (tpy = 2,2',2' '-terpyridine). These oxidants gave qualitatively similar selectivities to the electron-transfer rates from cyclic voltammetry, although the magnitudes of the selectivities were considerably lower on the sequencing gels. PMID- 11261982 TI - High-frequency EPR study of a new mononuclear manganese(III) complex:. [(terpy)Mn(N3)3] (terpy = 2,2':6',2''-terpyridine). AB - The isolation and structural characterization of [(terpy)Mn(III)(N3)3], complex 1, is reported (terpy = 2,2':6',2' '-terpyridine). Complex 1, a product of the reaction between the mixed-valence dimer [(terpy)(H2O)Mn(III)(O)2Mn(IV)(OH2)(terpy)](NO3)3 and NaN3, crystallizes in a triclinic system, space group P1, a = 8.480(1) A, b = 8.9007(2) A, c = 12.109(2) A, alpha = 93.79(1) degrees, beta = 103.17(1) degrees, gamma = 103.11(1) degrees, and Z = 2. Complex 1 exhibits a Jahn-Teller distortion of the octahedron characteristic of a six-coordinated high-spin Mn(III). A vibrational spectroscopic study was performed. The nu(asym)(N3) mode of complex 1 appears in the IR as a strong band at 2035 cm(-1) with a less intense feature at 2072 cm( 1), and in the FT-Raman as a strong band at 2071 cm(-1) with a weaker broad band at 2046 cm(-1). The electronic properties of complex 1 were investigated using a high-field and high-frequency EPR study (190-475 GHz). The different spin Hamiltonian parameters have been determined (D = -3.29 (+/-0.01) cm(-1), E = 0.48 (+/-0.01) cm(-1), E '= 0.53 (+/-0.01) cm(-1), g(x) = 2.00 (+/-0.005), g(y) = 1.98 (+/-0.005), g(z) = 2.01 (+/-0.005)). These parameters are in agreement with the geometry of complex 1 observed in the crystal structure, a D < 0 related to the elongated distortion, and a value of E/D close to 0.2 as expected from the highly distorted octahedron. The two values of the E-parameter are explained by the presence of two slightly different structural forms of complex 1 in the crystal lattice. A second hypothesis was explored to explain the experimental data. The calculation for the simulation was done taking into account that the g and D tensors are not collinear due to the low symmetry of complex 1. In that case, the spin Hamiltonian parameters found are D = -3.29 (+/-0.01) cm(-1), E = 0.51 (+/ 0.01) cm(-1), g(x) = 2.00 (+/-0.005), g(y) = 1.98 (+/-0.005), and g(z) = 2.01 (+/ 0.005). PMID- 11261983 TI - Synthesis and characterization of phosphorescent cyclometalated iridium complexes. AB - The preparation, photophysics, and solid state structures of octahedral organometallic Ir complexes with several different cyclometalated ligands are reported. IrCl3.nH2O cleanly cyclometalates a number of different compounds (i.e., 2-phenylpyridine, 2-(p-tolyl)pyridine, benzoquinoline, 2 phenylbenzothiazole, 2-(1-naphthyl)benzothiazole, and 2-phenylquinoline), forming the corresponding chloride-bridged dimers, CwedgeN2Ir(mu-Cl)2IrCwedgeN2 (CwedgeNis a cyclometalated ligand) in good yield. These chloride-bridged dimers react with acetyl acetone (acacH) and other bidentate, monoanionic ligands such as picolinic acid (picH) and N-methylsalicylimine (salH), to give monomeric CwedgeN2Ir(LX) complexes (LX = acac, pic, sal). The emission spectra of these complexes are largely governed by the nature of the cyclometalating ligand, leading to lambda(max) values from 510 to 606 nm for the complexes reported here. The strong spin-orbit coupling of iridium mixes the formally forbidden 3MLCT and 3pi-pi* transitions with the allowed 1MLCT, leading to a strong phosphorescence with good quantum efficiencies (0.1-0.4) and room temperature lifetimes in the microsecond regime. The emission spectra of the CwedgeN2Ir(LX) complexes are surprisingly similar to the fac-IrCwedgeN3 complex of the same ligand, even though the structures of the two complexes are markedly different. The crystal structures of two of the CwedgeN2Ir(acac) complexes (i.e., CwedgeN = ppy and tpy) have been determined. Both complexes show cis-C,C', trans-N,N' disposition of the two cyclometalated ligands, similar to the structures reported for other complexes with a "CwedgeN2Ir" fragment. NMR data (1H and 13C) support a similar structure for all of the CwedgeN2Ir(LX) complexes. Close intermolecular contacts in both (ppy)2Ir(acac) and (tpy)2Ir(acac) lead to significantly red shifted emission spectra for crystalline samples of the ppy and tpy complexes relative to their solution spectra. PMID- 11261984 TI - Molecular ladders with macrocyclic platforms. PMID- 11261985 TI - Antimony in the Sr4PtO6 structure: a neutron diffraction study of Sr3NaSbO6. PMID- 11261986 TI - Cation templation of anionic metal dicyanamide networks. PMID- 11261988 TI - Determination of arylglycerol-beta-aryl ethers and other linkages in lignins using DFRC/(31)P NMR. AB - An analytical method for lignins has been developed that involves derivatization followed by reductive cleavage (DFRC), depolymerization, and quantitative (31)P NMR spectroscopy. This technique detects and quantifies the various ether linkages present in softwood residual kraft lignins (RKL) and milled wood lignins (MWL). In addition, the technique supplies new quantitative information about beta-aryl ethers linked to condensed and noncondensed aromatic moieties, including dibenzodioxocins. Within RKL, beta-aryl ether bonds connected to condensed phenolic moieties predominated over those connected to noncondensed phenolic moieties. In addition, the amount of DFRC monomers determined by gas chromatography was minute in the RKL but large in the MWL. This indicates that almost all noncondensed beta-aryl ether linkages were cleaved during kraft pulping. The method offers new avenues for the detailed investigation of the bonding patterns of native and technical lignins. PMID- 11261989 TI - Immunoaffinity sample purification and MALDI-TOF MS analysis of alpha-Solanine and alpha-chaconine in serum. AB - A sample purification technique was developed for the detection of potato glycoalkaloids (GAs) in blood serum by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). GAs were extracted from spiked serum (5 mL) using a C(18) solid-phase extraction cartridge. The GAs were then selectively captured on antibody-coated agarose beads. The agarose beads were washed with water and the GAs eluted with 25 microL of methanol. MALDI-TOF MS was used to detect the GAs in the methanol eluent. Immunoaffinity sample purification of the GAs effectively reduced the signal suppression observed during the analysis of unpurified samples. alpha-Chaconine and alpha-solanine were detected in serum spiked with 1 ng/mL of each GA. PMID- 11261990 TI - Quantitative determination of L-arginine by enzymatic end-point analysis. AB - An enzymatic end-point method for the quantitative determination of L-arginine was evaluated with samples of synthetic wine and natural grape juice. The enzymes arginase, urease, and glutamate dehydrogenase were used in this simple assay, similar to those described for many metabolites by Boehringer-Mannheim. In synthetic wine, recovery of L-arginine ranged between 98.3 and 104.4% and the precision as coefficient of variation was between 0.4 and 1.47% in the concentration range of the method, 0-100 mg/L L-arginine. The recovery of L arginine in a grape juice with added L-arginine after clarification with polyvinylpolypyrrolidone ranged between 100 and 101.3%, and the coefficient of variation was 0.6%. The method has low material costs of approximately 0.43 U.S.$ per assay, and the time course of the reaction facilitates measurement of several samples concurrently. The results of this evaluation indicate that the enzymatic assay is a preferred method over colorimetric methods for the manual determination of L-arginine. PMID- 11261991 TI - On-line supercritical fluid extraction/enzymatic hydrolysis of vitamin a esters: a new simplified approach for the determination of vitamins a and e in food. AB - An on-line supercritical fluid extraction (SFE)/enzymatic hydrolysis procedure using immobilized lipase has been developed for the determination of vitamin A in dairy and meat products. Several lipases were tried, of which Novozyme 435 (Candida antarctica type B) showed the highest activity toward retinyl palmitate. There was no observed activity with alpha-tocopheryl acetate. When pressure, temperature, modifiers, flow rate, extraction time, and water content were varied, high vitamin A recovery was obtained in milk powder. Collected extracts were analyzed by reversed-phase high-performance liquid chromatography with ultraviolet and fluorescence detection without additional sample cleanup. The procedure gave reliable values of vitamin A as well as of vitamin E in other food items such as infant formula, minced pork and beef meat, and low- and high-fat liver paste. The described method is faster and more automated than conventional methods based on liquid-liquid extraction, or SFE using off-line saponification, for vitamin A and E determination. Results obtained with the new method did not differ significantly from those obtained with the other two methods mentioned above. PMID- 11261992 TI - A novel method to analyze betaine in chicken liver: effect of dietary betaine and choline supplementation on the hepatic betaine concentration in broiler chicks. AB - Betaine was measured from liver tissue by a high-performance liquid chromatographic (HPLC) method developed for this study. The method involves homogenization of liver in acetate buffer at pH 6 and precipitation of protein with trichloroacetic acid, which was removed by diethyl ether extraction. Betaine was separated using a cation exchange column in Ca(2+) form and detected with a refractive index detector. This method also allows the determination of S adenosylmethionine (S-AM) from the same liver extract but with different HPLC conditions. Broiler chicks were fed with experimental diets supplemented with four different doses of betaine or choline ranging from 0 to 5 mol equiv. After a 3 week feeding period, the livers were analyzed for betaine and S-AM. Dietary betaine was twice as efficient in increasing the hepatic betaine concentration as dietary choline. The hepatic S-AM concentrations were similar in all dietary treatments. PMID- 11261993 TI - Problems associated with measuring phytate in infant cereals. AB - The inositol hexaphosphate (IP6) content of commercially available dried infant cereals was measured by ion pair high-pressure liquid chromatography (ion pair HPLC) and ion exchange high-pressure liquid chromatography (ion exchange HPLC). Large differences between methods were apparent: ion pair HPLC gave values 14 to 190-fold lower than the values from ion exchange HPLC. Poor recoveries of added IP6 (25 to 60%) by ion pair HPLC suggested that some component of the infant cereal was responsible for the difference. Further experimentation suggested that an excess of minerals (approximately 11 mg/g calcium and 0.3 mg/g iron) in these samples sequestered the endogenously low phytate content. This problem may be unique to samples with low IP6 and high mineral content as wheat bran was not problematic. These results suggest that ion exchange HPLC is the method of choice for measuring inositol phosphates in infant cereals. PMID- 11261994 TI - Development of a gas chromatographic method for fungicide cymoxanil analysis in dried hops. AB - An analytical method for detecting cymoxanil [2-cyano-N-[(ethylamino)carbonyl]-2 (methoxyimino)acetamide] residues in dried hops was developed utilizing liquid liquid partitioning, automated gel permeation chromatography (GPC), solid phase extraction (SPE) cleanup, and gas chromatography (GC). Method validation recoveries from dried hops were 96 +/- 12, 108 +/- 11, and 136 +/- 8% over three levels of fortification (0.05, 0.5, and 1.0 ppm, respectively). The hop samples from three field sites, which were treated with cymoxanil, had residue levels ranging from 0.146 to 0.646 ppm. The detection limit and the quantitation limit of the method developed in the present study were 0.022 and 0.050 ppm, respectively. PMID- 11261995 TI - Rapid detection and identification of bacterial strains by Fourier transform near infrared spectroscopy. AB - The use of Fourier transform near-infrared (FT-NIR) spectroscopy and multivariate pattern recognition techniques for the rapid detection and identification of bacterial contamination in liquids was evaluated. The complex biochemical composition of bacteria yields FT-NIR vibrational transitions (overtone and combination bands) that can be used for classification and identification. Bacterial suspensions (Escherichia coli HB101, E. coli ATCC 43888, E. coli 1224, Bacillus amyloliquifaciens, Pseudomonas aeruginosa, Bacillus cereus, and Listeria innocua) were filtered to harvest the cells and eliminate the matrix, which has a strong NIR signal. FT-NIR measurements were done using a diffuse reflection integrating sphere. Principal component analysis showed tight clustering of the bacterial strains at the information-rich spectral region of 6000-4000 cm(-1). The method reproducibly distinguished between different E. coli isolates and conclusively identified the relationship between a new isolate and one of the test species. This methodology may allow for the rapid assessment of potential bacterial contamination in liquids with minimal sample preparation. PMID- 11261996 TI - Discrimination between orange juice and pulp wash by (1)H Nuclear Magnetic Resonance spectroscopy: identification of marker compounds. AB - The potential of NMR spectroscopy and multivariate analysis methods to detect the adulteration of orange juice with pulp wash is demonstrated. Principal component analysis has been applied to (1)H NMR spectra of >300 orange and pulp wash juices, and stepwise linear discriminant analysis was used to classify the samples. A model with six principal components gave a high success rate of classification (94%) for both training and validation sets. An important principal component loading showed that dimethylproline played a key role in the discrimination between the two types of juice, with higher levels in pulp wash. Dimethylproline was not previously known as a marker compound for orange juice adulteration. An ANOVA test revealed at least 21 other NMR signals that differed significantly between the authentic and pulp wash groups. The compounds they represent could be seen as potential marker compounds in addition to dimethylproline. This makes NMR with chemometrics an attractive screening tool with advantages in terms of rapidity, simplicity, and diversity of information provided. PMID- 11261997 TI - Monitoring ester formation in grape juice fermentations using solid phase microextraction coupled with gas chromatography-mass spectrometry. AB - Volatile esters contribute important floral and fruity sensory properties to wine. Numerous factors influence the biosynthesis and hydrolysis of esters throughout yeast fermentation; however, methods to monitor the dynamic changes in ester production that occur during winemaking processes are limited. In this study, we showed that solid phase microextraction (SPME), a rapid, solventless sampling procedure, combined with GC/MS analysis is a useful method for the nearly continuous analysis of volatile compounds such as esters that are produced during fermentation. Accuracy, precision, and limits of quantification were comparable to those of other sample preparation methods such as liquid-liquid extraction. Using GC/MS-SPME to monitor fatty acid ethyl esters and acetate esters, we obtained detailed information on the production patterns of ester formation during fermentation. This method now enables the monitoring of volatiles during fermentation and can provide greater insight into yeast metabolism and flavor formation. PMID- 11261998 TI - Fluorescence polarization as a means for determination of fumonisins in maize. AB - Fumonisins, mycotoxins produced by certain species of Fusaria, are commonly found worldwide as contaminants in maize. This paper reports the development of a rapid, portable fluorescence polarization-based assay for fumonisins in maize. The assay was based on the competition of unlabeled fumonisin, from a sample, with a fluorescently tagged fumonisin (FB(1)-FL) for a fumonisin-specific monoclonal antibody in solution. The fluorescence polarization (FP) of the tagged fumonisin was increased upon binding with the antibody. In the presence of free toxin, less of the FB(1)-FL was bound and the polarization signal was decreased. The assays were very simple to perform, requiring only mixing of an aqueous extract of maize with the tagged fumonisin and antibody, and required <2 min per sample, excluding extraction time. Two permutations of the assay were tested, one with each sample matrix serving as its own blank, and the other with all of the samples compared relative to a PBS blank with normalization of the data similar to an ELISA. The limit of detection, defined as the toxin content associated with a fluorescence polarization signal 5 standard deviations from that of a fumonisin free control, was 0.5 microg of FB(1)/g in spiked maize. Recoveries from spiked maize over the range of 0.5-20 ppm averaged 94.3 +/- 13.8%. Forty-eight samples of field-contaminated maize were tested by the FP and an established HPLC method, with a good correlation between the two (r(2) = 0.85-0.88). For these samples, the two variations of the FP assay also compared well to one another (r(2) = 0.97), suggesting the assay principle is very robust. The results, combined with the speed and ease of use for the assay, suggest that this technology has substantial potential as a screening tool for mycotoxins in foods. PMID- 11261999 TI - Comparison of spectroscopic techniques for the determination of Kjeldahl and ammoniacal nitrogen content of farmyard manure. AB - The feasibility of determining the nitrogen content of farmyard manure using infrared spectroscopy was investigated. Fifteen samples each of cattle, pig, and turkey manure were analyzed by three infrared techniques: Fourier transform mid infrared (MIR), using attenuated total reflection (ATR); near-infrared reflectance (NIR-R); and near-infrared optothermal photoacoustic (NIR-OT). The near-infrared measurements were made at wavelengths determined respectively by four (NIR-OT) and five (NIR-R) band-pass filters. The total nitrogen (using the Kjeldahl method) and volatile (ammoniacal) nitrogen contents of all samples were measured by wet chemistry. Internally cross-validated (ICV) partial least-squares (PLS) regression was then used to obtain calibrations for the nitrogen content. The data sets obtained by each technique were treated separately. Within these sets, data from each manure type were treated both separately and combined: the best predictive ability was obtained by combining data from all three manure types. From the combined data set, the residual standard deviations and correlation coefficients for the ICV-predicted versus actual Kjeldahl nitrogen content were, respectively, 6772 mg/kg dry wt, 0.862 (MIR); 9434 mg/kg dry wt, 0.771 (NIR-OT); and 8943 mg/kg dry wt, 0.865 (NIR-R). For the ammoniacal nitrogen content, the residual standard deviations and correlation coefficients were 3869 mg/kg dry wt, 0.899 (MIR); 6079 mg/kg dry wt, 0.820 (NIR-OT); and 3498 mg/kg dry wt, 0.961 (NIR-R). PMID- 11262000 TI - Determination of flubendazole and its metabolites in eggs and poultry muscle with liquid chromatography-tandem mass spectrometry. AB - The optimization of a quantitative and sensitive LC-MS/MS method to determine flubendazole and its hydrolyzed and reduced metabolites in eggs and poultry muscle is described. The benzimidazole components were extracted from the two matrices with ethyl acetate after the sample mixtures had been made alkaline. The HPLC separation was performed on an RP C-18 column with gradient elution, using ammonium acetate and acetonitrile as mobile phase. The analytes were detected after atmospheric pressure electrospray ionization on a tandem quadrupole mass spectrometer in MS/MS mode. The components were measured by the MS/MS transition of the molecular ion to the most abundant daughter ion. The overall extraction recovery values for flubendazole, the hydrolyzed metabolite, and the reduced metabolite in eggs (fortification levels of 200, 400, and 800 microg kg(-1)) and muscle (fortification levels of 25, 50, and 100 microg kg(-1)) were, respectively, 77, 78, and 80% and 92, 95, and 90%. The trueness (fortification levels of 400 and 50 microg kg(-1), respectively, for eggs and muscle), expressed as a percentage of the added values for these analytes, was, respectively, 89, 100, and 86 and 110, 110, and 98%. The proposed MS detection method operating in the MS/MS mode is very selective and very sensitive. The limits of detection for flubendazole and its hydrolyzed and reduced metabolites in egg and muscle were, respectively, 0.19, 0.29, and 1.14 microg kg(-1) and 0.14, 0.75, and 0.31 microg kg(-1). The limits of quantification were, respectively, 1, 1, and 2 microg kg( 1) and 1, 1, and 1 microg kg(-1). The discussed method was applied to a pharmacokinetic study with turkeys. Residue concentrations in breast and thigh muscle of turkeys orally treated with flubendazole were quantified. Medicated feed containing 19.9 and 29.6 mg kg(-1) flubendazole was provided to the turkeys for seven consecutive days. For the trial with the recommended dose of 19.9 mg kg(-1), one day after the end of the treatment, the mean sum of the flubendazole plus hydrolyzed metabolite residue values in thigh and breast muscle declined to below the maximum residue limit (50 microg kg(-1)) and were, respectively, 36.6 and 54.1 microg kg(-1). The corresponding values with the higher dose of 29.6 mg kg(-1) were, respectively, 101.7 and 119.7 microg kg(-1). PMID- 11262001 TI - Rapid high-performance liquid chromatography analysis for the quantitative determination of oleuropein in Olea europaea leaves. AB - Olea europaea (Oleaceae) leaves of 14 different cultivars have been studied by a new isocratic HPLC method. Qualitative and quantitative determinations of principal compounds were established for each cultivar. Oleuropein concentration was determined for each sampled tree, using coumarin as internal standard. Bid el Haman, Chemlali, Meski, Cailletier, Tanche, a Verdale-Picholine hybrid, and Lucques, in particular, had high oleuropein concentrations and could be useful sources for industrial extractions. PMID- 11262002 TI - Analysis of molecular species of glycolipids in fruit pastes of red bell pepper (Capsicum annuum L.) by high-performance liquid chromatography-mass spectrometry. AB - Five major glycolipid classes (acylated steryl glucoside, steryl glucoside, monogalactosyldiacylglycerol, digalactosyldiacylglycerol, and glucocerebroside) from fruit pastes of red bell pepper were separated by silica gel column chromatography. The molecular species of each glycolipid were separated and characterized by reversed-phase high-performance liquid chromatography coupled with on-line mass spectrometry using atmospheric pressure chemical ionization. The molecular species of steryl glucoside were beta-sitosteryl and campesteryl glucosides, and those of the acylated steryl glucoside were their fatty acid esters. The dilinolenoyl species was predominant in monogalactosyldiacylglycerol in addition to small amounts of another five molecular species, whereas digalactosyldiacylglycerol consisted of seven molecular species varying in their degree of unsaturation. The glucocerebroside class contained at least seven molecular species, which were characterized by proton nuclear magnetic resonance spectroscopy. PMID- 11262003 TI - Supercritical fluid extraction of 5-hydroxymethyl-2-furaldehyde from raisins. AB - An extraction method based on supercritical CO(2) has been developed for the analysis of 5-hydroxymethyl-2-furaldehyde in raisins. To optimize extraction variables, a fractional factorial experimental design was applied. Six extraction variables were optimized. The organic modifier used for increasing the extraction fluid solvating power was the most important factor. Methanol as organic modifier produced 10-fold higher recoveries of 5-hydroxymethyl-2-furaldehyde than ethyl acetate. The efficiency of the organic modifier in the static extraction phase was compared with using it in the dynamic extraction phase. Repeatability of the analysis method was evaluated, which resulted in an RSD of <5%. 5-Hydroxymethyl-2 furaldehyde was quantified in raisins, and the concentration was found to be 0.128 mg/g of raisin. PMID- 11262004 TI - Determination of estrogenic activity in beer by biological and chemical means. AB - It has been suspected that beer drinking may change the hormonal status of men caused by phytoestrogens. Five different Austrian lager beers have been investigated for estrogenic activity by a yeast two-plasmid system harboring the human estrogen receptor alpha, after concentration by solid phase extraction. The beer concentrate was further fractionated by reversed phase HPLC, and then the fractions were characterized by the biological assay and GC-MS. The most potent fraction did not contain a known phytoestrogen. The total activity corresponded to an average of 43 ng of 17beta-estradiol/L of beer. It was concluded that the human health hazard of beer drinking originating from compounds activity on the estrogen receptor alpha is negligible. PMID- 11262005 TI - High-level production of recombinant chicken cystatin by Pichia pastoris and its application in mackerel surimi. AB - A high level of the secreted form of recombinant chicken cystatin was expressed in Pichia pastoris X-33 by chromosomal integration of multiple copies of an expression cassette containing chicken cystatin under the control of glyceraldehyde-3-phosphate dehydrogenase promoter. The inhibition ability of the recombinant for papain-like proteinase was found to correspond to those of natural chicken cystatin. The recombinant cystatin substantially inhibited the proteolysis of myosin and gel softening, which consequently improved the gel properties of mackerel surimi. PMID- 11262006 TI - High-yield preparation of wax esters via lipase-catalyzed esterification using fatty acids and alcohols from crambe and camelina oils. AB - Fatty acids obtained from seed oils of crambe (Crambe abyssinica) and camelina (Camelina sativa) via alkaline saponification or steam splitting were esterified using lipases as biocatalysts with oleyl alcohol and the alcohols derived from crambe and camelina oils via hydrogenolysis of their methyl esters. Long-chain wax esters were thus obtained in high yields when Novozym 435 (immobilized lipase B from Candida antarctica) and papaya (Carica papaya) latex lipase were used as biocatalysts and vacuum was applied to remove the water formed. The highest conversions to wax esters were obtained with Novozym 435 (> or =95%) after 4-6 h of reaction, whereas with papaya latex lipase such a high degree of conversion was attained after 24 h. Products obtained from stoichiometric amounts of substrates were almost exclusively (>95%) composed of wax esters having compositions approaching that of jojoba (Simmondsia chinensis) oil, especially when crambe fatty acids in combination with camelina alcohols or camelina fatty acids in combination with crambe alcohols were used as substrates. PMID- 11262007 TI - Control of enzymatic browning in potato (Solanum tuberosum L.) by sense and antisense RNA from tomato polyphenol oxidase. AB - Polyphenol oxidase (PPO) activity of Russet Burbank potato was inhibited by sense and antisense PPO RNAs expressed from a tomato PPO cDNA under the control of the 35S promoter from the cauliflower mosaic virus. Transgenic Russet Burbank potato plants from 37 different lines were grown in the field. PPO activity and the level of enzymatic browning were measured in the tubers harvested from the field. Of the tubers from 28 transgenic lines that were sampled, tubers from 5 lines exhibited reduced browning. The level of PPO activity correlated with the reduction in enzymatic browning in these lines. These results indicate that expression of tomato PPO RNA in sense or antisense orientation inhibits PPO activity and enzymatic browning in the major commercial potato cultivar. Expression of tomato PPO RNA in sense orientation led to the greatest decrease in PPO activity and enzymatic browning, possibly due to cosuppression. These results suggest that expression of closely related heterologous genes can be used to prevent enzymatic browning in a wide variety of food crops without the application of various food additives. PMID- 11262008 TI - Effect of carbohydrate substrate on fermentation by kefir yeast supported on delignified cellulosic materials. AB - The suitability of delignified cellulosic (DC) material supported kefir yeast to ferment raw materials that contain various single carbohydrates, for the production of potable alcohol and alcoholic drinks, is examined in this investigation. Results are reported of fermentations carried out with sucrose, fructose, and glucose in synthetic media. Repeated batch fermentations at various initial sugar concentrations of sucrose, fructose, and glucose were performed at 30 degrees C in the presence of the aforementioned biocatalyst. The results clearly show feasible yields in the range of 0.38-0.41 g/g, alcohol concentrations of 7.6-8.2% v/v, fermentation time of 90-115 h, and conversion of 92-96%. DC material supported kefir fermented 11-fold more rapidly than free cells and 9-fold more rapidly in comparison to kissiris supported kefir. The main volatile byproducts such as amyl alcohols (mixture of 2-methyl-1-butanol and 3 methyl-1-butanol), ethanal, and ethyl acetate were formed in all sugar fermentation products. The formation of 65-110 ppm of ethyl acetate is as high and even higher than that obtained with traditional wine yeasts. The increase of the initial concentration of sugar in the fermentation media resulted in an increase in contents of volatiles. The fine aroma that was obtained in the product of fructose could be attributed to the high percentage of ethyl acetate on total volatiles. The efficiency of DC material supported kefir was the same for the fermentations of individual sugars or a mixture of fructose, sucrose, and glucose. When whey with raisin extracts was fermented, lower yields were obtained but the aroma of the product was even better. PMID- 11262009 TI - Polygalacturonase, pectinesterase, and lipoxygenase activities in high-pressure processed diced tomatoes. AB - High-pressure processing (HPP) can inactivate pathogenic microorganisms and degradative enzymes without the use of heat, thereby minimizing the destruction of flavors, nutrients, and other quality attributes. Lipoxygenase plays a role in the off-flavor production of tomatoes, whereas pectinesterase and polygalacturonase impact tomato texture. The purpose of this study was to determine HPP's ability to inactivate lipoxygenase, pectinesterase, and polygalacturonase in diced tomatoes. Processing conditions used were 400, 600, and 800 MPa for 1, 3, and 5 min at 25 and 45 degrees C. The magnitude of applied pressure had a significant effect on inactivating lipoxygenase and polygalacturonase (p < 0.05), with complete loss of activity occurring at 800 MPa. Pectinesterase was very resistant to pressure treatment. Percent soluble solids, pH, titratable acidity, and color a/b values did not differ significantly among the high-pressure-processed samples as compared to the control, but color L values increased. This change in L values was not considered of practical importance. Apparent protein content decreased in the pressure-processed samples, due possibly to protein denaturation, loss of solubility, and/or a decrease in dye binding sites to assay protein content. PMID- 11262010 TI - Flavor, color, and vitamin C retention of pulsed electric field processed orange juice in different packaging materials. AB - Effects of packaging materials, storage temperature, and time on the stability of pulsed electric field (PEF) processed orange juice were investigated. Single strength orange juice was treated with PEF at an electric field strength of 35 kV/cm for 59 micros using an integrated pilot plant scale PEF processing and glovebox packaging system. The retention of eight orange juice aroma compounds, color, and vitamin C in glass, polyethylene terephthalate (PET), high-density polyethylene, and low-density polyethylene were evaluated at 4 and 22 degrees C for 112 days. Packaging material had a significant effect (p < or = 0.05) on the retention of orange juice aroma compounds, color, and vitamin C. PEF-treated orange juice had a shelf life of >16 weeks in glass and PET at 4 degrees C. PMID- 11262011 TI - Photodegradation of cassava and corn starches. AB - The baking expansion properties of sour cassava starch (Polvilho azedo) are attributable to photochemical starch degradation induced by heterolactic fermentation after sun-drying. This study investigated the effects of UV irradiation on the different structural levels of cassava starch as compared to those of corn starch and dextrins. Photosensitive compounds excited at 360 and 290 nm in cassava starch were photodegraded when starch was exposed to sunlight or 360 nm irradiation. UV irradiation depolymerized cassava and corn starches, inducing modifications due, at least in part, to a mechanism involving free radicals. Lactic acid was also photodegraded. Photodegradation induced by UV absorption could have been due to fluorescent chromophores found in starches and nonfluorescent chromophores present in glucosidic units. PMID- 11262012 TI - Direct electron paramagnetic resonance study of tobacco. 1. Manganese(ii) as a marker. AB - Three categories of tobacco products were studied using electron paramagnetic resonance (EPR) spectroscopy: Cuban cigar brand name Montecristo, four international trademark cigarettes, and three types of Middle Eastern tobacco blends called Al-Moassal or Jurak. The Montecristo Cuban cigar is used as standard of high-quality tobacco. Mainly two EPR signals from all of the studied samples are observed: a very weak sharp EPR signal superimposed on a broad signal. The broad EPR signal is attributed to a manganese(II) complex. The intensity of the manganese(II) EPR signal is found to be related to the quality of the tobacco content. The sharp signal, which is characteristic of semiquinone radicals, is observed at room temperature, and its intensity increases drastically with temperature. PMID- 11262013 TI - Chemical investigation of gamma-irradiated saffron (Crocus sativus L.). AB - Changes in aroma and coloring properties of saffron (Crocus sativus) after gamma irradiation at doses of 2.5 and 5 kGy (necessary for microbial decontamination) were investigated. The volatile essential oil constituents responsible for aroma of the spice were isolated by steam distillation and then subsequently analyzed by gas chromatography/mass spectrometry (GC/MS). No significant qualitative changes were observed in these constituents upon irradiation, although a trained sensory panel could detect slight quality deterioration at a dose of 5 kGy. Carotene glucosides that impart color to the spice were isolated by solvent extraction and then subjected to thin-layer chromatography and high-performance liquid chromatography (HPLC). Fractionation of the above pigments into aglycon and glucosides was achieved by using ethyl acetate and n-butanol, respectively. Analysis of these fractions by HPLC revealed a decrease in glucosides and an increase in aglycon content in irradiated samples. The possibility of degradation of pigments during gamma irradiation is discussed. PMID- 11262015 TI - Effects of olive fruit quality and oil storage practices on the diacylglycerol content of virgin olive oils. AB - The evolution of 1,3- and 1,2-isomers of diacylglycerols (DGs) in olive oils obtained from healthy olives and the influence of the olive quality was studied. Healthy olive fruits yielded oils containing almost exclusively 1,2-isomers whereas altered olives produced oils with significant amounts of 1,3-isomers. Virgin olive oils obtained from various olive cultivars and stored at different temperatures showed triacylglycerol hydrolysis and diacylglycerol isomerization depending on the acidity and temperature. The results indicated that the relationship between acidity and total diacylglycerol content has scarce utility for detecting mild refined oil in virgin olive oil. On the other hand, the 1,3 /1,2-DG isomers ratio is useful for assessing the genuineness of virgin olive oils with low acidities during the early stages of storage. PMID- 11262014 TI - Assessment of home-based processing methods to reduce the phytate content and phytate/zinc molar ratio of white maize (Zea mays). AB - Various methods of processing maize suitable for household use in rural Malawi, Central Africa, were investigated for their ability to reduce its phytate content and phytate/zinc molar ratio. These methods included fermentation, germination, and soaking. Penta- and hexainositol phosphates were measured by HPLC, and zinc was measured by atomic absorption spectrophotometry. Natural lactic fermentation of maize flour slurries resulted in 88% phytate retention compared to unprocessed, unrefined maize flour porridges, whereas lower phytate retention was observed when a starter culture (61%) or germinated flour (71%) was also used. Fermentation of cooked maize flour porridges with germinated flour added resulted in 54-85% retention of phytate compared to controls. Soaking maize flour or pounded maize and decanting excess water resulted in 43 and 49% retention of phytate, respectively. The latter soaking procedures were simple and effective and were suitable for household use in rural Malawian communities. PMID- 11262016 TI - Natural levels of dimethyl sulfide in rough rice and its products. AB - Natural levels of dimethyl sulfide (DMS) in rough rice and its products (polished rice, brown rice, and broken rice) were determined by a gas chromatograph equipped with a flame photometric detector and sulfur mode, after extraction with 25% KBr solution in a sealed system. DMS was found to occur naturally in nine newly harvested and stored Australian varieties of rough rice and its products and decreased during storage after harvesting. Natural levels of DMS in rough rice and its products varied with variety, fraction, and period of storage. The order of levels of DMS was rough rice = brown rice > polished rice = broken rice. The range of values was 0.002-30 mg kg(-1) (ppm, w/w). PMID- 11262017 TI - Residue levels of chlorpropham in individual tubers and composite samples of postharvest-treated potatoes. AB - Chlorpropham, a herbicide and sprout suppressant, is used on stored potatoes to prolong the storage period without deterioration of produce quality. Data for residue concentrations on an individual tuber basis are required by WHO for the estimation of the variability factor. In this study, the levels of chlorpropham in individual tubers and in composite samples were determined. The distribution of chlorpropham between the peel and the tuber flesh was examined, and the fate during the cooking process (washing, boiling, frying) was studied. The concentrations in individual tubers ranged from 1.8 to 7.6 mg/kg 10 days postapplication (mean 3.8 mg/kg, RSD 39%), from 0.7 to 4.0 mg/kg 28 days postapplication (mean 2.9 mg/kg, RSD 28%), and from 0.8 to 3.8 mg/kg 65 days postapplication (mean 2.2 mg/kg, RSD 48%). The calculated residues in composite samples 10 days postapplication ranged from 4.3 to 6.1 mg/kg (mean 4.9 mg/kg, RSD 20%). Those in samples taken 28 days postapplication ranged from 3.1 to 4.2 mg/kg (mean 3.8 mg/kg, RSD 15%). The concentrations determined in composite samples of whole tubers 65 days postapplication ranged between 2.6 and 3.2 (mean 2.9 mg/kg, RSD 11%). Peeling removed 91-98% of the total residue; washing reduced residues by 33-47%. Detectable residues were found in boiled potatoes and the boiling water, and in french fries and the frying oil. Monitoring data on commercial prefried frozen french fries are reported. PMID- 11262018 TI - Acute, sublethal, antifeedant, and synergistic effects of monoterpenoid essential oil compounds on the tobacco cutworm, Spodoptera litura (Lep., Noctuidae). AB - Monoterpenoids (terpenes and biogenically related phenols) commonly found in plant essential oils were tested for acute toxicity via topical application to tobacco cutworms (Spodoptera litura Fab.). The most toxic among 10 such compounds was thymol (LD(50) = 25.4 microg/larva) from garden thyme, Thymus vulgaris. The compounds were then tested for sublethal effects, specifically inhibition of larval growth after topical application of low doses. Among 6 compounds tested, an LD(10) dose reduced growth by 20% on average 3 days after administration. Feeding deterrence was determined using a cabbage leaf disk choice test. The most deterrent compound was thymol, with a DC(50) of 85.6 microg/cm(2) leaf disk area. Because minor constituents in complex essential oils have been suggested to act as synergists, binary mixtures of the compounds were tested for synergy vis a vis acute toxicity and feeding deterrence. trans-Anethole acted synergistically with thymol, citronellal, and alpha-terpineol, in terms of both acute toxicity and feeding deterrence. On the basis of these findings, several complex mixtures were developed and tested as leads for effective control agents. Candidate mixtures demonstrated good synergistic effects. The observed LD(50) of mixture 3 was 40.6 microg/larvae compared to an expected value of 74.6 microg/larvae. The result of this research is a proprietary product suitable for commercial production. PMID- 11262019 TI - Tissue-specific patterns of lignification are disturbed in the brown midrib2 mutant of maize (Zea mays L.). AB - Despite recent progress, several aspects of lignin biosynthesis, including variation in lignin composition between species and between tissues within a given species, are still poorly understood. The analysis of mutants affected in cell wall biosynthesis may help increase the understanding of these processes. We have analyzed the maize brown midrib2 (bm2) mutant, one of the four bm mutants of maize, using pyrolysis-mass spectrometry (Py-MS) and pyrolysis-gas chromatography mass spectrometry (Py-GC-MS). Vascular tissues from the leaf blade and leaf sheath from different parts of the plant were investigated and compared to the corresponding samples from a wild-type plant of the same genetic background (inbred line A619). Multivariate analysis revealed that the bm2 mutant had reduced amounts of di- and trimeric lignin derivatives, notably species with m/z 272 and m/z 330, and that the ratio of guaiacyl residues to polysaccharides was reduced in the bm2 mutant. In addition, differences in cell wall composition between different parts of the plant (blade versus sheath, young versus old tissue) were much less pronounced in the bm2 mutant. These changes suggest that the functional Bm2 gene is important for the establishment of tissue-specific cell wall composition. PMID- 11262020 TI - beta-Conglycinin and glycinin in high-protein soybean seeds. AB - The agronomic performance and storage proteins of high seed protein lines of soybeans [Glycine max L. (Merr.)] were investigated to determine if the two major storage proteins, beta-conglycinin and glycinin, contribute to the increased protein content of high seed protein lines. Subunits of these two major storage proteins were estimated by scanning SDS-PAGE gels by scanning densitometry. The relative rankings of the lines with respect to seed size and protein content were not different between years in one environment over 5 years, but oil and total protein and oil contents and the ratio of protein to oil differed. The alpha', alpha, and beta subunits of beta-conglycinin were significantly higher in the high-protein lines except CX797-115, CX804-108, CX804-3, D81-8498, and NC-2-62. The acidic A(3) polypeptide of glycinin was significantly higher in high-protein lines except 76-48773, CX804-108, CX804-3, D81-8498, and NC-2-62, whereas the acidic polypeptides A(1,2,4) of glycinin were significantly higher in all of the high-protein lines. The basic polypeptides of glycinin were significantly higher in all high seed protein lines except D81-8259. In conclusion, high-protein lines appear to contain more beta-conglycinin and glycinin than normal-protein soybean lines, and the amounts of subunits and polypeptides differ among lines. PMID- 11262021 TI - Advanced technique for three-dimensional visualization of compound distributions in a rice kernel. AB - A three-dimensional (3D) visualization technique for the compound distribution in a rice kernel was developed. This technique is a combination of sectioning, staining, and digital image postprocessing. By using a special microtome system with adhesive tapes, a set of sequential sections of a rice kernel, which can be preserved with their own set of relative position data, was obtained. A single set of sequential sections was stained by various chemical techniques for the visualization of protein, starch, or lipid content. Each stained section was digitally captured using a CCD imaging device. As the stained areas represent areas containing dye-target complex, the distribution of each compound in the section was visualized in two dimensions. The digitally captured images of a single set of sequential sections were reconstructed to produce a 3D plotting image. As a result, the distributions of various compounds in a rice kernel could be visualized in a new 3D model. PMID- 11262022 TI - Studies on the constituents of Chenopodium quinoa seeds: isolation and characterization of new triterpene saponins. AB - Six triterpenoid saponins were isolated from the edible grain quinoa, which is seeds of Chenopodium quinoa (Chenopodiaceae). Following are their structures: phytolaccagenic acid 3-O-[alpha-L-arabinopyranosyl-(1' '-->3')-beta-D glucuronopyranosyl]-28-O-beta-D-glucopyranoside (1); phytolaccagenic acid 3-O [beta-D-glucopyranosyl-(1' '-->3')-alpha-L-arabinopyranosyl]-28-O-beta-D glucopyranoside (2); phytolaccagenic acid 3-O-[beta-D-glucopyranosyl-(1' "-->3' ')-beta-D-xylopyranosyl-(1' '-->2')-beta-D-glucopyranosyl]-28-O-beta-D glucopyranoside (3); phytolaccagenic acid 3-O-[beta-D-glucopyranosyl-(1' "-->2' ')-beta-D-glucopyranosyl-(1' '-->3')-alpha-L-arabinopyranosyl]-28-O-beta-D glucopyranoside (4); oleanolic acid 3-O-[alpha-L-arabinopyranosyl-(1' '-->3') beta-D-glucuronopyranosyl]-28-O-beta-D-glucopyranoside (5); and oleanolic acid 3 O-[beta-D-glucopyranosyl-(1' '-->3')-alpha-L-arabinopyranosyl]-28-O-beta-D glucopyranoside (6). The oleanane-type saponins (5, 6) were isolated for the first time in this plant, two of the phytolaccagenane (1, 3) were new compounds and two (2, 4) were previously found in quinoa. The structures were characterized on the basis of hydrolysis and spectral evidence, including 1D- and 2-D NMR (HMQC and HMBC) and ESI-MS analyses. PMID- 11262023 TI - Resveratrol and other phenolics from the bark of Yucca schidigera roezl. AB - Five phenolic constituents have been identified in Yucca schidigera bark, and their structures were established by spectral (FABMS and NMR) experiments. These included two known stilbenes, trans-3,4',5-trihydroxystilbene (resveratrol) and trans-3,3',5,5'-tetrahydroxy-4'-methoxystilbene, as well as three novel compounds, yuccaols A, B, and C, with spiro-structures rarely occurring in the plant kingdom. It is suggested that yuccaols A-C are biosynthethized via attachment of a stilbenic derivative to the carbocationic intermediate of the oxidative flavanone-flavonol conversion. PMID- 11262024 TI - Alfalfa (Medicago sativa L.) flavonoids. 1. Apigenin and luteolin glycosides from aerial parts. AB - Nine flavones and adenosine have been identified in aerial parts of alfalfa, and their structures were established by spectral (FABMS and NMR) techniques. Five of the identified compounds, including apigenin 7-O-[beta-D-glucuronopyranosyl(1- >2)-O-beta-D-glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, apigenin 7-O-[2 O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranosyl]-4'-O beta-D-glucuronopyranoside, apigenin 7-O-[2-O-feruloyl-[beta-D glucuronopyranosyl(1-->3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D glucuronopyranoside], apigenin 7-O-[2-O-p-coumaroyl-[beta-D-glucuronopyranosyl(1- >3)]-O-beta-D-glucuronopyranosyl(1-->2)-O-beta-D-glucuronopyranoside], and luteolin 7-O-[2-O-feruloyl-beta-D-glucuronopyranosyl(1-->2)-O-beta-D glucuronopyranosyl]-4'-O-beta-D-glucuronopyranoside, have not been reported before in the plant kingdom. Additionally, five known compounds, including apigenin 7-O-beta-D-glucuronopyranoside, apigenin 4'-O-beta-D glucuronopyranoside, apigenin 7-O-[beta-D- glucuronopyranosyl(1-->2)-O-beta-D glucuronopyranoside], luteolin 7-O-beta-D-glucuronopyranoside, and adenosine, were identified. PMID- 11262025 TI - Structure of a persistent heptachlorobornane in toxaphene (b7-1000) agrees with molecular model predictions. AB - A Cl(7) component of technical toxaphene (CTT), previously detected in marine mammals and fish and referred to as "7-1", was isolated from contaminated estuarine sediment using preparative solid-liquid chromatography followed by reversed-phase HPLC. The structure of this compound, elucidated by GC/MS and (1)H NMR, was 2-endo,3-exo,5-endo,6-exo,8,8,10-heptachlorobornane (hereafter referred to as B7-1000). This newly identified CTT eluted in the nonpolar fraction from silica and shares the alternating endo-exo chlorine substitution pattern with other relatively nonpolar, persistent congeners (e.g., B8-1413 and B9-1679). Based on ECNI-MS response, levels of B7-1000 in tissue samples of various higher organisms including humans were as high as 16% of B8-1413. Enantioselective determination of B7-1000 using a modified cyclodextrin chiral stationary phase (beta-BSCD) resulted in enantiomer ratios that were depleted in adipose tissue of a marine bird (skua) and Weddell seal blubber (0.3 and 0.5, respectively), but not in elephant seal blubber (1.1). Elucidation of the structure of B7-1000 thus validates previous predictions of persistence based on structure-activity relationships, chromatographic properties, and molecular modeling. PMID- 11262026 TI - Mid-infrared diffuse reflectance spectroscopy for the quantitative analysis of agricultural soils. AB - Soil samples were analyzed conventionally and by mid-infrared diffuse reflectance spectroscopy for total C, total N, pH, and measures of biological activity. Ground, non KBr diluted, samples (n = 180) from experimental plots (two locations, three replicate plots, under plow and no-till practices, three rates of N fertilizer, and from five depths) were scanned from 4000 to 400 cm(-1) (4 cm(-1) resolution, 64 co-added scans) on a DigiLab FTS-60 Fourier transform spectrometer using a custom-made linear sample transport (50 by 2 mm sample area scanned). Results using partial least-squares regression showed that accurate calibrations can be developed for the determination of a number of compositional parameters: total C, total N, pH, and many measures of biological activity. In general, the results achieved using mid-infrared spectra were at least as accurate as those found previously using near-infrared spectra and were sometimes significantly better, that is, pH. PMID- 11262027 TI - Quantification of the rice aroma compound, 2-acetyl-1-pyrroline, in uncooked Khao Dawk Mali 105 brown rice. AB - Volatile components of uncooked Khao Dawk Mali 105 brown rice were extracted using indirect steam distillation under reduced pressure and controlled temperature in order to prevent cooking. Analysis of the fresh extract by capillary gas chromatography-mass spectrometry revealed that there were >140 volatile constituents. Among these, 70 volatiles were identified, including 2 acetyl-1-pyrroline (2AP), a key aroma compound of cooked rice. Further study concentrated on an improved method for the quantification of 2AP in uncooked brown rice. The method was simplified by utilizing a solvent extraction procedure. Quantitative analysis was performed using a capillary gas chromatographic system employing a flame ionization detector with the aid of a more selective column, CP-Wax 51, for amines. This improved chromatographic system had remarkable detection sensitivity for 2AP in the rice extracts so that 2AP in an extract of only 0.5 g of uncooked Khao Dawk Mali 105 brown rice could be detected. PMID- 11262028 TI - Stability of monoterpenes encapsulated in gum Arabic by spray-drying. AB - Microencapsulation using spray-drying was tested with gum arabic and monoterpenes as wall and core materials, respectively. Citral, linalool, beta-myrcene, limonene, and beta-pinene were used at concentrations of 10, 20, and 30% with respect to the wall material. The greatest percentages of retention occurred at a concentration of 10%. Linalool and citral presented the greatest losses with increase in concentration. The hydrocarbons used were the most retained. Of the hydrocarbons, beta-pinene was better retained in the capsules than limonene, and beta-myrcene was the least retained of all. The capsules presented similar external morphologies, with no apparent cracks or porosity and an average size varying between 15.7 and 23.2 microm. The stability of the capsules to temperature was monitored for 33 days. The degradation products of the monoterpenes were evaluated. The results indicated a greater stability of the capsules containing beta-pinene and citral than of those containing linalool and beta-myrcene presenting the lowest retentions. PMID- 11262029 TI - Modifying the temporal profile of the high-potency sweetener neotame. AB - It is possible, using hydrophobic organic acids (such as cinnamate) or hydroxyamino acids (such as serine and tyrosine), to modify the temporal profile of the high-potency sweetener neotame. On the basis of Monte Carlo simulations, it was concluded that it is unlikely that this effect is due to direct interaction between the neotame molecule and the taste modifier. It is shown, using conformational analysis and molecular modeling, that the taste modifiers can adopt low-energy conformers which mimic the proposed active conformation of neotame, which suggests that the modifiers may compete for binding at the receptor site. PMID- 11262030 TI - Aroma extract dilution analysis of a beeflike process flavor from extruded enzyme hydrolyzed soybean protein. AB - Aroma-active compounds from a beeflike process flavor, produced by extrusion of enzyme-hydrolyzed vegetable protein (E-HVP), were analyzed using aroma extract dilution analysis. The number of aroma-active compounds and the aroma intensity were increased by the addition of aroma precursors prior to extrusion. The most intense compound was 2-methyl-3-furanthiol having a cooked rice/vitamin like/meaty aroma note. Several sulfur-containing furans, such as 2-methyl-3 (methylthio)furan, 2-methyl-3-(methyldithio)furan, and bis(2 methylfuryl)disulfide, were detected with high flavor dilution (FD) factors. Some pyrazines, such as 2-ethyl-3,5-dimethylpyrazine, 2,6-diethylpyrazine, and 3,5 diethyl-2-methylpyrazine, also had high FD factors. It is hypothesized that sulfur-containing amino acids and thiamin were important precursors in aroma formation in process flavor from E-HVP. PMID- 11262031 TI - Acetyl-coa: alcohol acetyltransferase activity and aroma formation in ripening melon fruits. AB - Melon varieties (Cucumis melo L.) differ in a range of physical and chemical attributes. Sweetness and aroma are two of the most important factors in fruit quality and consumer preference. Volatile acetates are major components of the headspace of ripening cv. Arava fruits, a commercially important climacteric melon. In contrast, volatile aldehydes and alcohols are most abundant in cv. Rochet fruits, a nonclimacteric melon. The formation of volatile acetates is catalyzed by alcohol acetyltransferases (AAT), which utilize acetyl-CoA to acetylate several alcohols. Cell-free extract derived from Arava ripe melons exhibited substantial levels of AAT activity with a variety of alcohol substrates, whereas similar extracts derived from Rochet ripe melons had negligible activity. The levels of AAT activity in unripe Arava melons were also low but steadily increased during ripening. In contrast, similar extracts from Rochet fruits displayed low AAT activity during all stages of maturation. In addition, the benzyl- and 2-phenylethyl-dependent AAT activity levels seem well correlated with the total soluble solid content in Arava fruits. PMID- 11262032 TI - Carbohydrate and amino acid degradation pathways in L-methionine/D-[13C] glucose model systems. AB - Maillard model systems consisting of labeled D-[(13)C]glucoses, L [(15)N]methionine, and L-[methyl-(13)C]methionine, have been utilized to identify the amino acid and carbohydrate fragmentation pathways occurring in the model system through Py-GC/MS analysis. The label incorporation analyses have indicated that the carbohydrate moiety produces 1-deoxy- and 3-deoxyglucosones and undergoes C(2)/C(4) and C(3)/C(3) cleavages to produce glycolaldehyde, tetrose, and C(3)-reactive sugar derivatives such as acetol, glyceraldehyde, and pyruvaldehyde. Glycolaldehyde was found to incorporate C-1, C-2 (70%) and C-5, C 6 (30%) glucose carbon fragments, whereas the tetrose moiety incorporates only C 3, C-4, C-5, C-6 glucose carbon atoms. In addition, the major source of reactive C(3) fragments was found to contain C-4, C-5, C-6 sugar moiety. On the other hand, methionine alone also generated Strecker aldehyde as detected by its condensation product with 3-(methylthio)propylamine. Plausible mechanisms were proposed for the formation of the interaction products between sugar and amino acid degradation products on the basis of the label incorporation patterns. PMID- 11262033 TI - Volatile release from an emulsion: headspace and in-mouth studies. AB - The headspace concentrations of three esters above solutions containing emulsified lipids were more resistant to dilution by a stream of gas than those above water alone. The effect was greatest for ethyl octanoate, and least for ethyl butyrate, with ethyl hexanoate showing intermediate behavior. This correlated with their solubility in the lipid fraction of the emulsion. Headspace analysis (comparing the emulsion with water) underestimated the release of the esters during consumption. The ratios observed between water and emulsion systems were different for the maximum breath concentration compared with headspace analysis. The emulsion appears to have acted as a reservoir for volatile release, counteracting the effects of sample dilution by saliva. PMID- 11262035 TI - Heterocyclic volatiles formed by heating cysteine or hydrogen sulfide with 4 hydroxy-5-methyl-3(2H)-furanone at pH 6.5. AB - The reaction of 4-hydroxy-5-methyl-3(2H)-furanone (HMF) with cysteine or hydrogen sulfide at pH 6.5 for 60 min at 140 degrees C produced complex mixtures of volatile compounds, the majority of these containing either sulfur or nitrogen. Of the 68 compounds detected, 63 were identified, some tentatively, by GC-MS. Among the identified compounds were thiophenes (10), thiophenones (6), thienothiophenes (5), thiazoles (5), trithiolanes (4), pyrazines (6), and oxazoles (4). More compounds were produced in the reaction of HMF with cysteine (63) than were formed in the reaction with hydrogen sulfide (33). In both systems, thiophenones were major reaction products, accounting for 25-36% of the total volatiles formed. Possible reasons for the differences in the composition of the two systems are discussed. The contributions of these reactions, and their products, to the flavor of heated foods are considered. PMID- 11262034 TI - Composition and antimicrobial activity of the essential oils of five taxa of Sideritis from Greece. AB - The chemical compositions of the essential oils obtained from the aerial parts of five taxa of Sideritis were analyzed using various GC-MS techniques. A total of 99 different compounds was identified, and significant differences (qualitative and quantitative) were observed between the samples. The in vitro antimicrobial activity of the essential oils against six bacteria and three fungi is also reported. PMID- 11262036 TI - Reduced immunogenicity of beta-lactoglobulin by conjugation with carboxymethyl dextran differing in molecular weight. AB - To reduce the immunogenicity of beta-lactoglobulin (beta-LG), two beta-LG carboxymethyl dextran (CMD) conjugates (Conj. 40 and Conj. 162) were prepared by using water-soluble carbodiimide (EDC). The molar ratios of beta-LG to CMD in Conj. 40 and Conj. 162 were 8:1 and 7:1, respectively. Each conjugate maintained approximately 50% of the retinol binding activity of beta-LG. Structural analyses by intrinsic fluorescence, CD spectra, and ELISA with monoclonal antibodies indicated that the surface of beta-LG in each conjugate was covered by CMD without great disruption of native conformation. By conjugation with CMD, the antibody response to beta-LG was reduced in BALB/c, C3H/He, and C57BL/6 mice, which was eminent in Conj. 162. The results of B cell epitope scanning using overlapping synthesized peptides showed that the linear epitope profiles of the conjugates were similar to those of beta-LG, whereas the antibody response to each epitope was reduced, which was eminent in Conj. 162. It was concluded that conjugation with CMD of higher molecular weight is effective in reducing the immunogenicity of beta-LG and that masking of epitopes by CMD is responsible for the reduced immunogenicity. PMID- 11262037 TI - Changes in chloroplast pigments of olive varieties during fruit ripening. AB - Changes in chlorophyll and carotenoid pigments of five olive (Olea europaea L.) varieties destined for milling were investigated at six consecutive ripening stages. There was a manifest dependence between olive variety, moment of picking, and chloroplast pigment composition of the fruits. Although the content of chlorophylls and carotenoids differed with fruit variety, ripening always involved their gradual loss, which becames more pronounced with increased presence of anthocyanin compounds. The relative rates of disappearance of chlorophylls and carotenoids were markedly different between varieties, implying that the catabolism of these pigments takes place at a relative rate inherent to each variety. The varieties less rich in pigments showed the most extreme behavior. The highest relative rate of disappearance was observed in fruits of the Blanqueta variety, and the lowest was observed in those of Arbequina. The chlorophyll a/chlorophyll b ratio remained practically constant during ripening, with a value very similar for Hojiblanca, Picual, Cornicabra, and Blanqueta, but much higher for Arbequina, implying that the structure of the photosynthetic apparatus is different in the latter variety. In the five varieties studied, lutein was the slowest carotenoid to be degraded, so that its percentage in the fruits increased with ripening, whereas beta-carotene was the fastest to disappear. In ripe fruits covered with anthocyanins, chloroplast pigments were retained in both skin and pulp, with the rate of disappearance being much higher in the latter. PMID- 11262038 TI - Soluble and total myrosinase activity in defatted Crambe abyssinica meal. AB - Crambe defatted meal contains 4-6% w/w of glucosinolates, with epiprogoitrin accounting for >90% of the total. This feature limits the use of the meal as feed due to the antinutritional properties of myrosinase-glucosinolate breakdown products. In this context, myrosinase activity assumes particular importance. In this study the total and soluble myrosinase activities have been evaluated directly on defatted meals of eight Crambe abyssinica varieties. The pH-stat method, which is the most suitable for assays in heterogeneous solid-water systems, was used. The total myrosinase activity in C. abyssinica varieties, determined using epiprogoitrin as substrate, ranged from 288 to 653 units g(-1). These activity values were up to 26 times higher than those obtained using other substrates, namely, sinigrin, glucosinalbin, glucotropaeolin, progoitrin, and glucoraphenin. Crambe myrosinase is unusual in that, unlike other Brassicaceae containing a typical main glucosinolate, it does not show the same specificity toward its natural substrates. PMID- 11262039 TI - Capillary electrophoresis analysis of orange juice pectinesterases. AB - Pectinesterase (PE) was extracted from orange juice and pulp with 1 M NaCl, desalted, and separated using capillary electrophoresis (CE) gel procedures (CE SDS-CGE) and isoelectric focusing (CE-IEF). PE resolved as a single peak using noncoated fused silica columns with CE-SDS-CGE. CE-IEF separation of PE required acryloylaminoethoxyethanol-coated columns, which had limited stability. Thermal stability of PE extracts before and after heating at 75 degrees C for 30 min and at 95 degrees C for 5 min established heat labile and heat stabile fractions with identical PE migration times by CE-SDS-CGE or CE-IEF. Peak magnitude decreased to a constant value as heating time increased at 75 degrees C. Regression analysis of CE-SDS-CGE peak migration times of molecular weight (MW) standards estimated both heat labile and heat stable PE at MW approximately 36 900. Traditional SDS PAGE gel separation of MW standards and active PE isolated by IEF allowed estimation of MW approximately 36 000. CE-SDS-CGE allowed presumptive, but not quantitative, detection of active PE in fresh juice. PMID- 11262040 TI - Quantitative structure-activity relationship modeling of peptide and protein behavior as a function of amino acid composition. AB - A quantitative structure-activity relationship (QSAR) modeling approach based on the location of each amino acid along three axes obtained by principal component analysis (called z scores) was extended to physical and functional properties of proteins, where the proportion of particular amino acids rather than a precise sequence is the determining factor. Coomassie Brilliant Blue spectral responses to amino acid homopolymers (R = 0.926) and proteins, either as a function of their contents of six basic and aromatic amino acids (R = 0.976) or as a function of the contributions of these amino acids to the three z scores (R = 0.935), were modeled. The ultraviolet absorbance of proteins was modeled in terms of the z score contributions of tyrosine, tryptophan, and cysteine (R = 0.995). Modeling many protein functional properties in this manner appears to be possible. An approach to modeling peptide behaviors that depend on short sequences of amino acids was also considered. PMID- 11262041 TI - Adsorbed protein secondary and tertiary structures by circular dichroism and infrared spectroscopy with refractive index matched emulsions. AB - The secondary structure of protein adsorbed at the emulsion interface has been studied in refractive index matched emulsions using the techniques of circular dichroism (CD) and Fourier transform infrared spectroscopy. Bovine serum albumin (BSA) and bovine beta-lactoglobulin (betalg) stabilized emulsions were studied, and the refractive index was altered by the addition of glycerol or polyethylene glycol. The effect of additive on the solution and adsorbed protein structure in addition to the effect of adsorption time was considered. Both adsorption and glycerol addition alter protein secondary structure; however, the majority of secondary structure remains. Small changes are observed in the secondary structure of adsorbed protein with time. Near-ultraviolet CD studies showed the effect of glycerol and adsorption on the aromatic groups. BSA showed small changes both upon the addition of glycerol to protein in solution and upon adsorption. betalg showed slightly larger changes upon the addition of glycerol to protein in solution and a larger change upon adsorption. PMID- 11262042 TI - Evolution of wheat gliadins conformation during film formation: a fourier transform infrared study. AB - The secondary structures of wheat gliadins (a major storage protein fraction from gluten) in film-forming solutions and their evolution during film formation were investigated by Fourier transform infrared spectroscopy. In the film-forming solution, wheat gliadins presented a mixture of different secondary structures, with an important contribution of beta-turns induced by proline residues. The presence of plasticizer did not have any influence on protein secondary structure in the film-forming solution. The evolution of protein conformation was followed during drying; the major feature of this evolution was a clear growing of the infrared band at 1622 cm(-1), characteristic of intermolecular hydrogen-bonded beta-sheets. This revealed the formation of protein aggregates during film drying. The influence of the drying temperature on film properties and gliadin secondary structures was also investigated. Higher drying temperatures induced an increase of both the tensile strength of the films and the amount of beta-sheets aggregates. Although the appearance of heat-induced disulfide bridge cross-links has already been described, there is clear evidence that hydrogen-bonded beta sheets aggregates are also induced by thermal treatment. It was not possible, however, to determine whether there is a direct relationship between the occurrence of these aggregates and the increase of the tensile strength of the films. PMID- 11262043 TI - Pressure-temperature phase transition diagram for wheat starch. AB - Wheat starch suspensions in water (5% dry matter) were subjected to various pressures (0.1-600 MPa) and temperatures (-20 to 96 degrees C) for 15 min. The gelatinization rate was measured after treatment by using microscopic measurements of the loss of birefringence of the granules. This method was previously calibrated by differential scanning calorimetry. Curves of isogelatinization were found to be quite similar to a pressure-temperature (P-T) diagram of unfolding proteins. Results were first analyzed by considering the thermodynamic aspects related to the dT/dP curve shifts. On the basis of equations already shown for proteins, the P-T gelatinization diagram of wheat starch would show different kinds of thermal contributions, suggesting endothermic, athermic, or exothermic melting reactions. Second, as a practical consequence, these previous P-T areas corresponded to specific gelatinization conditions as confirmed by hydration evaluation measured by starch swelling index. Depending on the pressure-temperature conditions, gelatinization would involve hydration. Lowering the pressure and temperature resulted in a complete gelatinization with less hydration in comparison with a thermal treatment at atmospheric pressure. A hydration model based on an energetic approach was proposed. PMID- 11262044 TI - Seasonal changes of carotenoid pigments and color in Hamlin, Earlygold, and budd blood orange juices. AB - The developmental patterns of carotenoids in Hamlin, Earlygold (an early-maturing selection), and Budd Blood sweet orange juices were studied during the September to mid-January period of the 1996-97 and 1997-98 seasons. The carotenoid concentration of Earlygold increased by as much as 4.9 times during the color development compared to 3.9 times in Hamlin and 4.5 times in Budd Blood juice in the same period. For the profiles of carotenoid pigment, dramatic changes occurred among the pigments that were present in high concentrations at the beginning of the season, with lutein and violaxanthin noted as the predominant pigments in Hamlin fruit. A marked increase in the percentage of beta cryptoxanthin allowed it to become a major pigment in the late stage of maturation. The color development in the new cultivar Earlygold was especially notable, reaching the 36 color number, which is grade A, by late October to mid November whereas Hamlin juice did not reach this grade A color number until January. Budd Blood juice was similar in carotenoid pigment content and seasonal changes to Hamlin juice, but also, the development of red anthocyanin pigment in January significantly increased juice color. PMID- 11262045 TI - Growth of Trichoderma viride on crude cell wall preparations from barley. AB - Trichoderma viride can utilize crude cell wall preparations from barley starchy endosperm as sole source of carbon and energy. In the process beta-(1-->3)(1-->4) glucan and arabinoxylan are released. The onset of release of the latter preceded that of glucan, consistent with arabinoxylan being encountered and utilized first. The release of several enzymes was observed during growth of Trichoderma on this substrate: endo-beta-(1-->3)(1-->4)-glucanase, endo-beta-(1-->4) glucanase, endo-xylanase, arabinofuranosidase, esterase, carboxypeptidase, and "beta-glucan solubilase". It is inferred that each of these activities is necessary for the digestion of this substrate. PMID- 11262046 TI - Gelation of edible blue-green algae protein isolate (Spirulina platensis Strain Pacifica): thermal transitions, rheological properties, and molecular forces involved. AB - Proteins isolated from blue-green algae Spirulina platensis strain Pacifica were characterized by visible absorption, differential scanning calorimetry (DSC), viscometry, and dynamic oscillatory rheological measurements. Unique thermal unfolding, denaturation, aggregation, and gelation of the algal protein isolate are presented. DSC analysis showed that thermal transitions occur at about 67 and 109 degrees C at neutral pH. Calcium chloride stabilized the quaternary structure against denaturation and shifted the transitions at higher temperatures. Viscometric studies of Spirulina protein isolate as a function of temperature showed that the onset of the viscosity increase is closely related to the dissociation-denaturation process. Lower viscosities were observed for the protein solutions dissolved at pH 9 due to an increased protein solubility. Solutions of Spirulina protein isolate form elastic gels during heating to 90 degrees C. Subsequent cooling at ambient temperatures caused a further pronounced increase in the elastic moduli and network elasticity. Spirulina protein isolate has good gelling properties with fairly low minimum critical gelling concentrations of about 1.5 and 2.5 wt % in 0.1 M Tris buffer, pH 7, and with 0.02 M CaCl(2) in the same buffer, respectively. It is suggested that mainly the interactions of exposed hydrophobic regions generate the molecular association, initial aggregation, and gelation of the protein isolate during the thermal treatment. Hydrogen bonds reinforce the network rigidity of the protein on cooling and further stabilize the structure of Spirulina protein gels but alone are not sufficient to form a network structure. Intermolecular sulfhydryl and disulfide bonds were found to play a minor role for the network strength of Spirulina protein gels but affect the elasticity of the structures formed. Both time and temperature at isothermal heat-induced gelation within 40-80 degrees C affect substantially the network formation and the development of elastic modulus of Spirulina protein gels. This is also attributed to the strong temperature dependence of hydrophobic interactions. The aggregation, denaturation, and gelation properties of Spirulina algal protein isolate are likely to be controlled from protein-protein complexes rather than individual protein molecules. PMID- 11262047 TI - Effect of oleic and linoleic acids on the production of deep-fried odor in heated triolein and trilinolein. AB - To determine sources of desirable deep-fried flavor in frying oils, degradation products from heated triolein and trilinolein with 5-31% polar compounds representing low to high deterioration were evaluated by purge-trap gas chromatography-mass spectrometry-olfactometry. (E,E)-2,4-Decadienal, 2-heptenal, 2-octenal, 2,4-nonadienal, and 2,4-octadienal produced deep-fried odor at moderate-strong intensities in heated trilinolein. However, unexpected aldehydes 2,4-decadienal, 2,4-undecadienal, 2,4-nonadienal, and 2-octenal (all <15 ppm) were produced in triolein heated for 6 h. These dienals possibly were produced by hydroperoxidation and/or hydroxylation followed by dehydration of 2-alkenals. The 2-alkenals were produced from thermal decomposition of hydroperoxides, epoxides, and keto and dimeric compounds produced during the heating of triolein. These aldehydes produced low intensities of deep-fried odor in triolein. This information helps to explain sources of the deep-fried flavor that is characteristic of high linoleic frying oils but which is only at low intensity levels in high oleic frying oils. PMID- 11262048 TI - Denaturation and aggregation of myosin from two bovine muscle types. AB - The thermal behaviors of myosin from bovine vastus intermedius (VI, predominantly red muscle) and semimembranosus (SM, predominantly white muscle) at pH 6.05 (ultimate pH of VI muscle) and 5.50 (ultimate pH of SM muscle) were compared. Differential scanning microcalorimetry and turbidity measurements were used to monitor changes in myosin during heating from 25 to 80 degrees C at 1 degrees C/min. VI and SM myosin heavy chain isoforms were identified on gradient SDS PAGE. Endotherms of VI myosin at pH 6.05 had three transition temperatures (T(m)) of 45, 53, and 57 degrees C, whereas at pH 5.50 two transitions were observed at 42 and 59 degrees C. SM myosin had two T(m) values of 46 and 58 degrees C at pH 6.05 and T(m) values of 43 and 62 degrees C at pH 5.5. SM myosin at its ultimate pH was less heat stable than VI myosin at its ultimate pH; however, when SM and VI myosin were compared at the same pH, VI myosin was less stable. PMID- 11262049 TI - Effect of freezing and frozen storage of doughs on bread quality. AB - The effects of freezing and storage in frozen conditions on bread quality, crumb properties, and aggregative behavior of glutenins were analyzed. The effect of different additives on bread quality was also studied. The results obtained showed that freezing and storage at -18 degrees C decreased the bread quality. Samples stored in frozen conditions supplemented with diacetyl-tartaric acid ester of monoglycerides, gluten, and guar gum produced breads of greater volume and more open crumb structure than those prepared with the base formulation (without additives). All additives analyzed increased the proof time. Crumb firmness increased with dough frozen storage and bread aging time at 4 degrees C. A decrease in the amount of glutenin subunits of high molecular mass was observed by electrophoresis analysis of the SDS-soluble proteins aggregates extracted from the frozen dough. This result suggested that the protein matrix of bread underwent depolymerization during storage in frozen conditions. PMID- 11262050 TI - Combined effect of ascorbic acid and gamma irradiation on microbial and sensorial characteristics of beef patties during refrigerated storage. AB - The present study was undertaken to evaluate the effect of ascorbic acid concentrations (0.03 to 0.5%) and irradiation doses (0.5 to 4 kGy) on microbial growth, color coordinates (L, a, and b), and sensory characteristics (taste and odor) of beef patties during storage at 4 +/- 1 degrees C. Ascorbic acid was also compared to citric acid at a similar pH value in order to differentiate the effects of ascorbic acid from those of pH reduction. Results showed significant reduction (p< or = 0.05) of aerobic plate counts (APCs) and total coliforms, and a significant interaction (p< or = 0.05) between ascorbic acid and irradiation dose was observed. The irradiation treatment had detrimental effects on redness, yellowness, and hue angle values of meat. However, incorporation of ascorbic acid into the meat before irradiation resulted in significant (p< or = 0.05) stabilization of color parameters. The color improvement obtained with ascorbic acid was not related to the pH reduction. Also, no significant detrimental effect on taste or odor was found in irradiated samples containing ascorbic acid. PMID- 11262051 TI - Nutrient distribution and phenolic antioxidants in air-classified fractions of beach pea (Lathyrus maritimus L.). AB - Beach pea (Lathyrus maritimus L.) cotyledons and hulls were air-classified into different fractions. The crude protein content (%N x 6.25) of samples ranged from 32.8 to 35.3% in cotyledons and 14.7 to 16.8% in hulls. Crude fiber content was higher in hulls fraction 1 (37.13%) and fraction 2 (36.85%) than in cotyledons (2.83, 2.99, and 3.08% in fractions 1, 2, and 3, respectively). Condensed tannins of cotyledons ranged from 5.76 to 6.90% and of hulls ranged from 52.49 to 57.24%, expressed as catechin equivalents. Minerals, namely P, K, and Zn, were higher in cotyledons, but Ca and Mn were more prevalent in hulls. Nonprotein nitrogen was concentrated in hulls, whereas phytic acid was more abundant in the cotyledons. The UV absorption pattern showed that flavonoids were present in fractions (I III) from hulls separated on Sephadex LH-20. Fraction III from hulls had the highest content of total phenolics and condensed tannins, but no condensed tannins were detected in fractions I and II from hulls. The antioxidant activity of fractions separated on Sephadex LH-20 from hulls and crude extracts in a beta carotene-linoleate model system was in the order of fraction III > crude extract > fraction II > fraction I. Spots on silica gel TLC plates, sprayed with a solution of beta-carotene and linoleic acid, indicated that many of the individual compounds were antioxidative in nature. Further, separation of fraction III from hulls on a semipreparative HPLC showed the presence of (+) catechin and (-) epicatechin as the main low-molecular-weight phenolic compounds present. PMID- 11262052 TI - Ascorbic acid-induced browning of (+)-catechin in a model wine system. AB - The ability of ascorbic acid to induce browning of (+)-catechin in a model wine system has been studied. A significant increase in absorbance at 440 nm was observed over 14 days when ascorbic acid was incubated at 45 degrees C with (+) catechin in a model wine base. The onset of browning was delayed for about 2 days, although the length of the lag period was dependent on the amount of molecular oxygen in the headspace of the reaction system. The lag period was not observed when a preoxidized solution of ascorbic acid was used, suggesting that a product of ascorbic acid oxidation is responsible for the onset of browning. Hydrogen peroxide, when added directly to (+)-catechin in the model system, was not capable of producing the same degree of browning as that generated by ascorbic acid. Liquid chromotography evidence is presented to show that different reaction products are produced by ascorbic acid and hydrogen peroxide. PMID- 11262053 TI - Structural features of procyanidin interactions with salivary proteins. AB - Procyanidin dimers and trimer C1 were synthesized, whereas (-)-epicatechin O gallate and B2-3"-O-gallate were isolated from grape seeds. Human saliva was separated into two fractions. One of these was mainly alpha-amylase and the other mainly proline-rich proteins (PRPs). The procyanidin compounds were combined with each of the saliva protein fractions and with bovine serum albumin. The protein polyphenol interactions were observed using nephelometry. (+)-Catechin had a higher tannin specific activity (TSA) for PRPs than (-)-epicatechin (1.45 versus 0.65 nephelos turbidity units/mg of polyphenol). This indicated the effect of the stereochemistry of flavan-3-ols on their interaction with proteins. Procyanidin dimers linked through a C(4)-C(8) interflavanoid bond had consistently greater TSA than their counterparts with a C(4)-C(6) linkage. Esterification of a galloyl group to the C(3) hydroxyl function of (-)-epicatechin or to the epicatechin moiety of procyanidin dimer B2 increased TSA. This was not as strong an effect for the dimer, probably as a result of the expected "closed" structure of B2-3"-O gallate. PMID- 11262054 TI - Polyphenol oxidase activity, color changes, and dehydration in table grape rachis during development and storage as affected by n-(2-chloro-4-pyridyl)-n phenylurea. AB - Flame Seedless grapes were sprayed with N-(2-chloro-4-pyridyl)-N-phenylurea (CPPU) at 0, 2.5, and 5.0 ppm to develop rachis resistant to browning and dehydration. Rachis polyphenol oxidase (PPO) activity was determined during cluster development. Cluster components were weighed at commercial (CM), and physiological maturity (PM). PPO activity, rachis color changes (L and a), and cluster weight loss were evaluated at 0 degrees C for 8, 16, 32, and 56 days. CPPU-treated rachis had a decrease of 36% in PPO activity and a week delay in peak activity. At PM, dry weight of CPPU-treated rachis increased by 3 g. Postharvest rachis PPO activity declined with CPPU application, and color changes followed the same pattern for CM and PM. After 32 days of storage, L and a in lateral branches were significantly superior in CPPU treatments. Weight losses below 2.1% were significantly lowest in CPPU-treated clusters for 16 days of storage regardless of cluster maturity. PMID- 11262055 TI - Comparative contents of dietary fiber, total phenolics, and minerals in persimmons and apples. AB - Dietary fibers, major phenolics, main minerals, and trace elements in persimmons and apples were analyzed and compared in order to choose a preferable fruit for an antiatherosclerotic diet. Fluorometry and atomic absorption spectrometry following microwave digestion were optimized for the determination of major phenolics and minerals. Total, soluble, and insoluble dietary fibers, total phenols, epicatechin, gallic and p-coumaric acids, and concentrations of Na, K, Mg, Ca, Fe, and Mn in whole persimmons, their pulps, and peels were significantly higher than in whole apples, pulps, and peels (P < 0.01-0.0025). Conversely, the contents of Cu and Zn were higher in apples than in persimmons. In persimmons and apples all of the above components were higher in their peels than in whole fruits and pulps. The relatively high contents of dietary fibers, total and major phenolics, main minerals, and trace elements make persimmon preferable for an antiatherosclerotic diet. PMID- 11262056 TI - The anthocyanidins cyanidin and delphinidin are potent inhibitors of the epidermal growth-factor receptor. AB - The aglycons of the most abundant anthocyanins in food, cyanidin (cy) and delphinidin (del), were found to inhibit the growth of human tumor cells in vitro in the micromolar range, whereas malvidin (mv), a typical anthocyanidin in grapes, was less active. The aglycons preferentially inhibited the growth of the human vulva carcinoma cell line A431, overexpressing the epidermal growth-factor receptor (EGFR). The glycosides cyanidin-3-beta-D-galactoside (cy-3-gal, idaein) and malvidin-3-beta-D-glucoside (mv-3-glc, oenin) did not affect tumor cell growth up to 100 microM. The tyrosine kinase activity of the EGFR, isolated from A431 cells, was potently inhibited by cy and del. Mv and the glycosides cy-3-gal and mv-3-glc were inactive up to 100 microM. In intact cells the influence of anthocyanin treatment on downstream signaling cascades was investigated by measuring the phosphorylation of the transcription factor Elk-1. A431 cells were transiently transfected with a luciferase reporter gene construct whose expression is controlled by MAP kinase pathway dependent phosphorylation of a GAL4-Elk-1 fusion protein. We found that cy and del inhibited the activation of the GAL4-Elk-1 fusion protein in the concentration range where growth inhibition was observed. Thus, the anthocyanidins cy and del are potent inhibitors of the EGFR, shutting off downstream signaling cascades. These effects might contribute substantially to the growth-inhibitory properties of these natural food constituents. PMID- 11262057 TI - Studies on the antioxidative activities of Hsian-tsao (Mesona procumbens Hemsl) leaf gum. AB - This study aimed at evaluating the antioxidative activity of crude hsian-tsao leaf gum extracted by sodium bicarbonate solutions and precipitated by 70% ethanol. The antioxidative activities, including the radical-scavenging effects, Fe(2+)-chelating ability, and reducing power as well as the inhibition of FeSO(4) H(2)O(2)-induced malondialdehyde formation in rat tissue homogenate were studied in vitro. It was found that the antioxidative effect provided by hsian-tsao leaf gum was strongly concentration dependent. In general, the antioxidative activity increased with increasing gum concentration, to a certain extent, and then leveled off with further increase in gum concentration. A concentrtaion-dependent kinetics for the rate of change in antioxidative activity was proposed. The antioxidative activity constant (k) and the half-inhibition concentration (IC(50)) for each antioxidative reaction studied were calculated. From a comparison of the IC(50) values for different antioxidative reactions, it seemed that hsian-tsao leaf gum was more effective in scavenging superoxide radicals than chelating Fe(2+) or scavenging alpha,alpha-diphenyl-beta-picrylhydrazyl (DPPH) radicals. As compared to the commercial antioxidants, hsian-tsao leaf gum showed less scavenging effect on the DPPH radical and reducing power but better superoxide radical-scavenging effect and Fe(2+)-chelating ability than alpha tocopherol and BHT. PMID- 11262058 TI - Antioxidant capacity in cranberry is influenced by cultivar and storage temperature. AB - Ten cranberry (Vaccinium macrocarpon Aiton) cultivars were evaluated for oxygen radical absorbance capacity (ORAC), anthocyanins, and total phenolics contents after three months of storage at 0, 5, 10, 15, and 20 degrees C. The antioxidant capacity of cranberry was affected by cultivars and storage temperatures. Among the 10 cranberry cultivars used in this study, Early Black, Crowley, and Franklin had higher antioxidant capacities than the other cultivars. ORAC values, anthocyanins, and total phenolics contents increased during storage. The highest increases in antioxidant activity, anthocyanin, and phenolics contents occurred at 15 degrees C storage. Fruit stored at 20 degrees C had lower ORAC values than those stored at 15 degrees C. A positive relationship existed between ORAC values and anthocyanin or phenolic content in all 10 cranberry cultivars at different storage temperatures. PMID- 11262059 TI - Degradation of starchy endosperm cell walls in nongerminating sterilized barley by fungi. AB - Strains of fungi from different origins, including isolates of the natural microflora of barley, were screened for their ability to modify barley starchy endosperm cell walls in situ. In an initial step, fungi were selected that degrade the major component of the cell walls, that is, (1-->3),(1-->4)-beta-D glucan, in vitro on artificial media. Nongerminating, sterilized barley, obtained by gamma-irradiation, was inoculated with such fungi and subjected to solid state fermentation under conditions resembling those of a traditional malting process. For some strains of fungi, a clear correlation between the production of endo beta-glucanase and the friability of the treated kernels was found. Image analysis of Calcofluor stained longitudinal sections of barley kernels fermented with the endo-beta-glucanase producing strains showed that starchy endosperm cell walls were modified. As malt quality is inversely related to its (1-->3),(1-->4) beta-D-glucan content, the selected strains have high potential to be used as starter cultures during malt production, contributing to the processing quality of the final product. PMID- 11262060 TI - Effect of storage on fructan and fructooligosaccharide of onion (Allium cepa L.). AB - The purpose of this study was a comparative examination of the fructan and fructooligosaccharide (FOS) content of different varieties of onions (Allium cepa L. cv. Sturon, Hysam, Durco, Grano de Oro, and Caribo) and the changes produced during their commercial storage. In fresh onions, the Grano de Oro variety presented a remarkably different behavior, showing low contents of total fructans and FOS and high levels of reducing sugars. In the other varieties, Sturon, Hysam, Durco, and Caribo, fructans were the main carbohydrates, the lowest polymerized FOS being the major oligomer. Storage period caused in these varieties important increased levels of free fructose attributed to fructan hydrolysis. Maleic hydrazide treatment had no significant effect in avoiding the hydrolysis of fructans during storage conditions for the Sturon variety. Varieties with >16% dry matter or 15% soluble solids contents could be stored for 6 months at 0 degrees C and 60-65% relative humidity. PMID- 11262061 TI - Aging of whey protein films and the effect on mechanical and barrier properties. AB - This work focuses on the aging of whey protein isolate (WPI) films plasticized with glycerol (G) and sorbitol (S). The films were cast from heated aqueous solutions at pH 7 and dried at 23 degrees C and 50% relative humidity (RH) for 16 h. They were stored in a climate room (23 degrees C, 50% RH) for 120 days, and the film properties were measured at regular intervals. The moisture content (MC) of the WPI/G films decreased from 22% (2 days) to 15% (45 days) and was thereafter constant at 15% (up to 120 days). This affected the mechanical properties and caused an increased stress at break (from 2.7 to 8.3 MPa), a decreased strain at break (from 33 to 4%), and an increased glass transition temperature (T(g)) (from -56 to -45 degrees C). The barrier properties were, however, unaffected, with constant water vapor permeability and a uniform film thickness. The MC of the WPI/S films was constant at approximately 9%, which gave no change in film properties. PMID- 11262062 TI - Lipid particle size effect on water vapor permeability and mechanical properties of whey protein/beeswax emulsion films. AB - Lipid particle size effects on water vapor permeability (WVP) and mechanical properties of whey protein isolate (WPI)/beeswax (BW) emulsion films were investigated. Emulsion films containing 20 and 60% BW (dry basis) and mean lipid particle sizes ranging from 0.5 to 2.0 microm were prepared. BW particle size effects on WVP and mechanical properties were observed only in films containing 60% BW. WVP of these films decreased as lipid particle size decreased. As drying temperature increased, film WVPs decreased significantly. Meanwhile, tensile strength and elongation increased as BW particle size decreased. However, for 20% BW emulsion films, properties were not affected by lipid particle size. Results suggest that increased protein-lipid interactions at the BW particle interfaces, as particle size decreased and resulting interfacial area increased, result in stronger films with lower WVPs. Observing this effect depends on a large lipid content within the protein matrix. At low lipid content, the effect of interactions at the protein-lipid interfaces is not observed, due to the presence of large protein-matrix regions of the film without lipid, which are not influenced by protein-lipid interactions. PMID- 11262063 TI - Proteolytic activation of latent Paraguaya peach PPO. Characterization of monophenolase activity. AB - The kinetics of the activation process of latent peach PPO by trypsin was studied. By coupling this activation process to the oxidation of 4-tert butylcatechol (TBC) to its corresponding quinone, it was possible to evaluate the specific rate constant of active PPO formation, k(3), which showed a value of 0.04 s(-1). This proteolytic activation of latent peach PPO permitted us to characterize the monophenolase activity of peach PPO for the first time using p cresol as substrate, and it showed the characteristic lag period of the kinetic mechanism of monophenols hydroxylation, which depended on the enzyme and substrate concentration, the pH and the presence of catalytic amounts of o diphenol (4-methylcatechol). The enzyme activation constant, k(act), was 2 microM. PMID- 11262064 TI - Lipid oxidation in fish oil enriched mayonnaise: calcium disodium ethylenediaminetetraacetate, but not gallic acid, strongly inhibited oxidative deterioration. AB - The antioxidative effects of gallic acid, EDTA, and extra emulsifier Panodan DATEM TR in mayonnaise enriched with 16% fish oil were investigated. EDTA reduced the formation of free radicals, lipid hydroperoxides, volatiles, and fishy and rancid off-flavors. The antioxidative effect of EDTA was attributed to its ability to chelate free metal ions and iron from egg yolk located at the oil water interface. Gallic acid reduced the levels of both free radicals and lipid hydroperoxides but promoted slightly the oxidative flavor deterioration in mayonnaise and influenced the profile of volatiles. Gallic acid may therefore promote the decomposition of lipid hydroperoxides to volatile oxidation products. Addition of extra emulsifier reduced the lipid hydroperoxide levels but did not influence the level of free radicals or the oxidative flavor deterioration in mayonnaisse; however, it appeared to alter the profile of volatiles. The effect of the emulsifier on the physical structure and rheological properties depended on the presence of antioxidants. PMID- 11262065 TI - Partial isolation and characterization of a cysteine proteinase inhibitor from Lima bean (Phaseolus lunatus). AB - Lima beans (Phaseolus lunatus) have been shown to contain cysteine proteinase inhibitor (CPI) activity, but the CPI has not been isolated or characterized. Accordingly, our objective was to isolate and partially characterize a CPI from lima bean. The isolation scheme included water extraction of lima bean flour followed by a chromatography series using DEAE Sepharose, Phenyl Sepharose, hydroxyapatite, and reversed-phase high performance liquid chromatography. This scheme resulted in the partial purification of a approximately 20 000-dalton protein with high inhibitory activity against papain. This isolated lima bean CPI had an N-terminal sequence homologous with other members of the cystatin class of CPIs. The protein was relatively heat labile; suggesting it could be inactivated with normal cooking, which is favorable for its use in transforming plants to create insect resistance. PMID- 11262066 TI - In vitro study of possible role of dietary fiber in lowering postprandial serum glucose. AB - There have been many reports concerning the role of dietary fiber in lowering postprandial serum glucose, and the main mechanism was regarded as the viscosity of different dietary fibers in hampering diffusion of glucose and postponing absorption and digestion of carbohydrates. In this paper, two kinds of water insoluble dietary fibers, water-insoluble dietary fiber of wheat bran and enzyme resistant starch of maize amylose, and four kinds of water-soluble dietary fibers, water-soluble dietary fiber of wheat bran, carboxymethyl cellulose, guar gum, and xanthan gum, were used to investigate their postprandial serum glucose lowering mechanism in vitro. The results showed that these dietary fibers lowered postprandial serum glucose levels at least by three mechanisms. First, dietary fibers increase the viscosity of small intestine juice and hinder diffusion of glucose; second, they bind glucose and decrease the concentration of available glucose in the small intestine; and, third, they retard alpha-amylase action through capsuling starch and the enzyme and might directly inhibit the enzyme. All of these decreased the absorption rate of glucose and the concentration of postprandial serum glucose. PMID- 11262068 TI - In vitro availability of flavonoids and other phenolics in orange juice. AB - Hand-squeezed navel orange juice contains 839 mg/L phenolics, including flavanones, flavones, and hydroxycinnamic acid derivatives. The flavanones are the main phenolics in the soluble fraction (648.6 mg/L) and are also present in the cloud fraction (104.8 mg/L). During refrigerated storage of fresh juice (4 degrees C), 50% of the soluble flavanones precipitate and integrate into the cloud fraction. Commercial orange juices contain only 81-200 mg/L soluble flavanones (15-33%) and the content in the cloud is higher (206-644 mg/L) (62 85%), showing that during industrial processing and storage the soluble flavanones precipitate and are included in the cloud. An in vitro simulation of orange juice digestion shows that a serving of fresh orange juice (240 mL) provides 9.7 mg of soluble hesperidin (4'-methoxy-3',5,7-trihydroxyflavanone-7 rutinoside) and 4.7 mg of the C-glycosylflavone vicenin 2 (apigenin, 6,8-di-C glucoside) for freshly squeezed orange juice, whereas pasteurized commercial juices provide 3.7 mg of soluble hesperidin and a higher amount of vicenin 2 (6.3 mg). This means that although orange juice is a very rich source of flavanones, only a limited quantity is soluble, and this might affect availability for absorption (11-36% of the soluble flavanones, depending on the juice). The flavanones precipitated in the cloud are not available for absorption and are partly transformed to the corresponding chalcones during the pancreatin-bile digestion. PMID- 11262067 TI - Characterization of Aloeride, a new high-molecular-weight polysaccharide from Aloe vera with potent immunostimulatory activity. AB - We have characterized a new immunostimulatory polysaccharide called Aloeride from commercial aloe vera (Aloe barbadensis) juice. Aloeride is between 4 and 7 million Da, and its glycosyl components include glucose (37.2%), galactose (23.9%), mannose (19.5%), and arabinose (10.3%). At 0.5 microg/mL Aloeride increased NF-kappa B directed luciferase expression in THP-1 human monocytic cells to levels 50% of those achieved by maximal concentrations (10 microg/mL) of LPS. Aloeride induced the expression of the mRNAs encoding IL-1beta and TNF-alpha to levels equal to those observed in cells maximally activated by LPS. Acemannan, the major carbohydrate component from aloe, used at 200 microg/mL in the macrophage assay resulted in negligible NF-kappa B activation. Analysis of acemannan and Aloeride using size-exclusion chromatography suggests that the low activity of acemannan is due to trace amounts of Aloeride. Although Aloeride comprises only 0.015% of the aloe juice dry weight, its potency for macrophage activation accounts fully for the activity of the crude juice. PMID- 11262069 TI - Synthesis of (-)-[4-3H]epigallocatechin gallate and its metabolic fate in rats after intravenous administration. AB - Because a great deal of attention has been focused on the metabolism of (-) epigallocatechin gallate (EGCg), quantitative analysis of this compound is required. For this purpose we developed a method of chemical synthesis of [4 (3)H]EGCg. Synthesized [4-(3)H]EGCg showed 99.5% radiochemical purity and a specific activity of 13 Ci/mmol. To clarify the excretion route of EGCg, the radioactivity levels of bile and urine were quantified after intravenous administration of [4-(3)H]EGCg to bile-duct-cannulated rats. Results showed that the radioactivity of the bile sample excreted within 48 h accounted for 77.0% of the dose, whereas only 2.0% of the dose was recovered in the urine. The excretion ratio of bile to urine was calculated to be about 97:3. These results clearly showed that bile was the major excretion route of EGCg. Time-course analysis of the radioactivity in blood was also performed to estimate the pharmacokinetic parameters following intravenous administration of [4-(3)H]EGCg. In addition, EGCg metabolites excreted in the bile within 4 h after the intravenous dose of [4 (3)H]EGCg were analyzed by HPLC. The results showed that 4',4"-di-O-methyl-EGCg was present in the conjugated form and made up about 14.7% of the administered radioactivity. PMID- 11262070 TI - Effects of heat treatment of casein in the presence of reducing sugars on calcium bioavailability: in vitro and in vivo assays. AB - Casein-glucose-fructose mixtures unheated (C) or after heating (HC) were added to a solution of ionic calcium to study calcium speciation and included in diets (C D, HC-D) for rats. Samples and diets were digested in vitro. Supernatants of digested samples were used for transport experiments with Caco-2 cells. Total soluble and ionic calcium levels were lower and precipitated calcium levels higher with HC compared to C. Dialyzed calcium from the diets was highly ionic and lower in HC-D compared to C-D. Nondialyzed soluble calcium was also lower, whereas precipitated calcium was higher, in HC-D. HC increased calcium transport in Caco-2 cells compared to C, but transport efficiency decreased due to lower calcium solubililty after digestion. Urinary calcium increased with HC-D consumption without changes in calcium absorbed and retained. Maillard reaction products in HC decrease calcium solubility, but enterocyte metabolism and calcium absorption and retention seem to be unaffected. Nevertheless, urinary calcium losses increase. PMID- 11262071 TI - Molecular dynamics simulations on the mycotoxin fumonisin B1. AB - The solution conformational properties of the mycotoxin fumonisin B(1) have been studied using molecular dynamics methodology. Fumonisins have been shown to inhibit sphinganine (sphingosine) N-acyltransferase (ceramide synthase) and show a wide range of toxic effects in many animals. This study of the solution properties of fumonisin B(1) attempts to add to the structural models necessary for the understanding of the binding and activity properties. The computational method uses a box with periodic boundaries, filled with explicit TIP3P water molecules, the substrate fumonisin B(1), and selected counterions for charge neutrality. The starting structure of fumonisin B(1) is added to the box by excluding water molecules. The explicit image method using 12-A cutoffs is applied to the system and molecular dynamics are carried out on different starting conformations at 300 K in 100-picosecond (ps) steps. Examination of the resulting equilibrated conformations suggests that the structure is relatively extended and that previous computational studies in vacuo, showing a compact folded structure, may not be consistent with the solution structure. PMID- 11262072 TI - Areas with high concentrations of selenium in the soil and forage produce beef with enhanced concentrations of selenium. AB - Beef provides a significant portion of human dietary selenium (Se), and it is possible that modest portions of beef produced in areas with high-Se soil and forage could provide the entire Recommended Dietary Allowance (RDA) for Se. The present study has addressed the environmental conditions that resulted in the production of high-Se beef. One hundred and thirty-eight cull cows were obtained from 21 ranches in five distinct geographic regions that, on the basis of soil parent material, reports of Se deficiency, and previous soil and forage Se surveys, were likely to have high or low Se concentrations in the soil. Grass and soil samples were taken from ranch sites, and hair, whole blood, skeletal muscle, diaphragm muscle, and liver samples were obtained from the animals. Hair and whole blood samples were taken 1 day prior to shipping. Selenium concentrations of all samples were determined by hydride generation atomic absorption spectroscopy. Geographic origin affected Se content of all samples (p < 0.05). Selenium concentrations in soil (r = 0.53; p < 0.01) and grass (r = 0.63; p < 0.01) were correlated to Se content of skeletal muscle. Selenium concentrations in whole blood, diaphragm, hair, and liver also were significantly correlated to Se content of skeletal muscle (p < 0.01). Cows that received Se in mineral supplements did not have significantly higher concentrations of Se in sampled tissues (p > 0.05). Results of this study suggest that the greatest source of variation in Se content of bovine skeletal muscle was the geographic region from which the beef originated and not production or management practices. Results also demonstrated that a 100 g serving of high-Se beef could provide 100% of the RDA for Se. PMID- 11262075 TI - Perspectives in animal health: old targets and new opportunities. PMID- 11262077 TI - Anti-AIDS agents. 42. Synthesis and anti-HIV activity of disubstituted (3'R,4'R) 3',4'-di-O-(S)-camphanoyl-(+)-cis-khellactone analogues. AB - A series of disubstituted 3',4'-di-O-(S)-camphanoyl-(+)-cis-khellactone (DCK) analogues (1-10) were synthesized and evaluated for inhibition of HIV-1 replication in H9 lymphocytes. 5-Methoxy-4-methyl DCK (8) was the most promising compound with an EC(50) value of 7.21 x 10(-6) microM and a therapeutic index of >2.08 x 10,(7) which were much better than those of lead compound DCK in the same assay. Another six disubstituted DCK analogues (1-5 and 7) were more potent than AZT but less active than DCK. Conformational analysis suggested that resonance of the coumarin system is an essential structural feature for potent anti-HIV activity. Steric compression of C(4) and C(5) substituents of the coumarin moiety can reduce the overall planarity and thus resonance of the coumarin nucleus, resulting in a decrease or lack of anti-HIV activity. PMID- 11262076 TI - X-Ray structure of citrate bound to Src SH2 leads to a high-affinity, bone targeted Src SH2 inhibitor. PMID- 11262078 TI - A new class of nonsteroidal aromatase inhibitors: design and synthesis of chromone and xanthone derivatives and inhibition of the P450 enzymes aromatase and 17 alpha-hydroxylase/C17,20-lyase. AB - Aromatase (P450arom) is a target of pharmacological interest for the treatment of breast cancer. In this paper, we report the design, synthesis, and in vitro biological evaluation of a series of new (di)benzopyranone-based inhibitors of this enzyme. The design of the new compounds was guided by a CoMFA model previously developed for a series of nonsteroidal aromatase inhibitors. Both the chromone and the xanthone nuclei were taken as molecular skeletons, and the functions supposed to be critical for binding to the aromatase active site - a heterocyclic ring (imidazole or 1,3,4-triazole) linked to the aromatic moiety by a methylene unit and an H-bond accepting function (CN, NO(2), Br) located on the aromatic ring at a suitable distance from the heterocyclic nitrogen carrying the lone pair--were attached to them. The chromone, xanthone, and flavone derivatives were prepared by conventional synthetic methods from the appropriate methyl analogues. Aromatase inhibitory activities were determined by the method of Thompson and Siiteri, using human placental microsomes and [1 beta,2 beta (3)H]testosterone as the labeled substrate. All the compounds were also tested on 17 alpha-hydroxylase/C17,20-lyase (P450 17), an enzyme of therapeutic interest for the treatment of prostatic diseases. The goal to find new potent inhibitors of aromatase was reached with the xanthone derivatives 22d,e (IC(50) values 43 and 40 nM, respectively), which exceeded the potency of the known reference drug fadrozole and also showed high selectivity with respect to P450 17. Moreover, compounds 22g-i based on the same xanthonic nucleus showed fairly high potency as P450 17 inhibitors (IC(50) values 220, 130, and 42 nM, respectively). Thus, they might be new leads for the development of drug candidates for androgen-dependent diseases. PMID- 11262079 TI - Discovery of 4-[3-(trans-3-dimethylaminocyclobutyl)-1H-indol-5-ylmethyl]-(4S) oxazolidin-2-one (4991W93), a 5HT(1B/1D) receptor partial agonist and a potent inhibitor of electrically induced plasma extravasation. AB - Utilizing a pharmacophoric model of binding of 3-(2-aminoethyl)indoles to 5HT(1B/1D) receptors, we identified the 3-aminocyclobutyl group as a potential ethylamine isostere. A novel multidimensional chemometric approach was used to predict the intrinsic activity (degree of agonism) at the receptor. A qualitative model for pharmacokinetic properties was then used to guide the synthesis toward molecules likely to have oral bioavailability in humans. A novel synthetic route to 3-(3-dimethylaminocyclobutyl)indoles was developed. Analogues showed generally lower intrinsic activity at 5HT(1B/1D) receptors than their ethylamine counterparts. 4-[3-(trans-3-Dimethylaminocyclobutyl)-1H-indol-5-ylmethyl]-(4S) oxazolidin-2-one (4991W93, 1) was identified as a partial agonist against 5HT(1B/1D) receptors, with low intrinsic activity. This molecule also has significant activity against 5HT(1F) receptors but is selective over other 5HT receptors. In addition this compound was found to be an exceptionally potent inhibitor of electrically induced plasma extravasation. Compound 1 may have utility in the treatment and prophylaxis of migraine. PMID- 11262080 TI - Antimitotic antitumor agents: synthesis, structure-activity relationships, and biological characterization of N-aryl-N'-(2-chloroethyl)ureas as new selective alkylating agents. AB - A series of N-aryl-N'-(2-chloroethyl)ureas (CEUs) and derivatives were synthesized and evaluated for antiproliferative activity against a wide panel of tumor cell lines. Systematic structure--activity relationship (SAR) studies indicated that: (i) a branched alkyl chain or a halogen at the 4-position of the phenyl ring or a fluorenyl/indanyl group, (ii) an exocyclic urea function, and (iii) a N'-2-chloroethyl moiety were required to ensure significant cytotoxicity. Biological experiments, such as immunofluorescence microscopy, confirmed that these promising compounds alter the cytoskeleton by inducing microtubule depolymerization via selective alkylation of beta-tubulin. Subsequent evaluations demonstrated that potent CEUs were weak alkylators, were non-DNA-damaging agents, and did not interact with the thiol function of either glutathione or glutathione reductase. Therefore, CEUs are part of a new class of antimitotic agents. Finally, among the series of CEUs evaluated, compounds 12, 15, 16, and 27 were selected for further in vivo trials. PMID- 11262081 TI - Correlation of anti-HIV activity with anion spacing in a series of cosalane analogues with extended polycarboxylate pharmacophores. AB - Cosalane and its synthetic derivatives inhibit the binding of gp120 to CD4 as well as the fusion of the viral envelope with the cell membrane. The binding of the cosalanes to CD4 is proposed to involve ionic interactions of the negatively charged carboxylates of the ligands with positively charged arginine and lysine amino acid side chains of the protein. To investigate the effect of anion spacing on anti-HIV activity in the cosalane system, a series of cosalane tetracarboxylates was synthesized in which the two proximal and two distal carboxylates are separated by 6--12 atoms. Maximum activity was observed when the proximal and distal carboxylates are separated by 8 atoms. In a series of cosalane amino acid derivatives containing glutamic acid, glycine, aspartic acid, beta-alanine, leucine, and phenylalanine residues, maximum activity was displayed by the di(glutamic acid) analogue. A hypothetical model has been devised for the binding of the cosalane di(glutamic acid) conjugate to CD4. In general, the compounds in this series are more potent against HIV-1(RF) in CEM-SS cells than they are vs HIV-1(IIIB) in MT-4 cells, and they are least potent vs HIV-2(ROD) in MT-4 cells. PMID- 11262082 TI - Heterobifunctional ligands: practical chemoenzymatic synthesis of a cell adhesive glycopeptide that interacts with both selectins and integrins. AB - An efficient and practical synthesis of cell adhesive glycopeptides exhibiting unique properties as a novel type of modulator of cellular recognition is described. A nonnatural glycopeptide 1 composed of sialyl Lewis x and Lys-Gly-Arg Gly-Asp-Ser that interacts with both selectins and integrins has been systematically synthesized by combined chemical and enzymatic strategy. It is suggested that glycopeptide 1 showed much higher affinity with P-selectin (K(a) = 6.6 x 10(7) M(-1)) and E-selectin (K(a) = 4.5 x 10(5) M(-1)) than sialyl Lewis x. This compound also inhibited a specific interaction between human integrin beta(1) and its monoclonal antibody more effectively than the tetrapeptide Arg Gly-Asp-Ser. Interestingly, it was demonstrated by surface plasmon resonance analysis that this heterobifunctional glycopeptide exhibited a capacity to form stable complexes with P-selectin and integrin beta(1) concurrently. It is also suggested that this activity can be used for the inhibition of integrin-mediated adhesion of activated helper T cells onto collagen-coated plates as a cell migration model. These results indicate that the chemoenzymatic hybridization strategy of different biological functions of carbohydrates and peptides is a new concept for designing potent glycoconjugates as antiinflammatory and anticancer metastasis reagents. PMID- 11262083 TI - Cyclic ketone inhibitors of the cysteine protease cathepsin K. AB - Cathepsin K (EC 3.4.22.38), a cysteine protease of the papain superfamily, is predominantly expressed in osteoclasts and has been postulated as a target for the treatment of osteoporosis. Crystallographic and structure--activity studies on a series of acyclic ketone-based inhibitors of cathepsin K have led to the design and identification of two series of cyclic ketone inhibitors. The mode of binding for four of these cyclic and acyclic inhibitors to cathepsin K is discussed and compared. All of the structures are consistent with addition of the active site thiol to the ketone of the inhibitors with the formation of a hemithioketal. Cocrystallization of the C-3 diastereomeric 3-amidotetrahydrofuran 4-one analogue 16 with cathepsin K showed the inhibitor to occupy the unprimed side of the active site with the 3S diastereomer preferred. This C-3 stereochemical preference is in contrast to the X-ray cocrystal structures of the 3-amidopyrrolidin-4-one inhibitors 29 and 33 which show these inhibitors to prefer binding of the 3R diastereomer. The 3-amidopyrrolidin-4-one inhibitors were bound in the active site of the enzyme in two alternate directions. Epimerization issues associated with the labile alpha-amino ketone diastereomeric center contained within these inhibitor classes has proven to limit their utility despite promising pharmacokinetics displayed in both series of compounds. PMID- 11262084 TI - Design, synthesis, and evaluation of a novel pyrrolobenzodiazepine DNA interactive agent with highly efficient cross-linking ability and potent cytotoxicity. AB - A novel sequence-selective pyrrolobenzodiazepine (PBD) dimer 5 (SJG-136) has been developed that comprises two C2-exo-methylene-substituted DC-81 (3) subunits tethered through their C8 positions via an inert propanedioxy linker. This symmetric molecule is a highly efficient minor groove interstrand DNA cross linking agent (XL(50) = 0.045 microM) that is 440-fold more potent than melphalan. Thermal denaturation studies show that, after 18 h incubation with calf thymus DNA at a 5:1 DNA/ligand ratio, it increases the T(m) value by 33.6 degrees C, the highest value so far recorded in this assay. The analogous dimer 4 (DSB-120) that lacks substitution/unsaturation at the C2 position elevates melting by only 15.1 degrees C under the same conditions, illustrating the effect of introducing C2-exo-unsaturation which serves to flatten the C-rings and achieve a superior isohelical fit within the DNA minor groove. This behavior is supported by molecular modeling studies which indicate that (i) the PBD units are covalently bonded to guanines on opposite strands to form a cross-link, (ii) 5 has a greater binding energy compared to 4, and (iii) 4 and 5 have equivalent binding sites that span six base pairs. Dimer 5 is significantly more cytotoxic than 4 in a number of human ovarian cancer cell lines (e.g., IC(50) values of 0.0225 nM vs 7.2 nM, respectively, in A2780 cells). Furthermore, it retains full potency in the cisplatin-resistant cell line A2780cisR (0.024 nM), whereas 4 loses activity (0.21 microM) with a resistance factor of 29.2. This may be due to a lower level of inactivation of 5 by intracellular thiol-containing molecules. A dilactam analogue (21) of 5 that lacks the electrophilic N10-C11/N10'-C11' imine moieties has also been synthesized and evaluated. Although unable to interact covalently with DNA, 21 still stabilizes the helix (Delta T(m) = 0.78 degrees C) and has significant cytotoxicity in some cell lines (i.e., IC(50) = 0.57 microM in CH1 cells), presumably exerting its effect through noncovalent interaction with DNA. PMID- 11262085 TI - Thiazole and thiadiazole analogues as a novel class of adenosine receptor antagonists. AB - Novel classes of heterocyclic compounds as adenosine antagonists were developed based on a template approach. Structure-affinity relationships revealed insights for extended knowledge of the receptor-ligand interaction. We replaced the bicyclic heterocyclic ring system of earlier described isoquinoline and quinazoline adenosine A(3) receptor ligands by several monocyclic rings and investigated the influence thereof on adenosine receptor affinity. The thiazole or thiadiazole derivatives seemed most promising, so we continued our investigations with these two classes of compounds. The large difference between a pyridine and isoquinoline ring in binding adenosine A(1) and A(3) receptors showed the importance of the second ring of the isoquinoline ligands. We prepared several N-[4-(2-pyridyl)thiazol-2-yl]benzamides, and these compounds showed adenosine affinities in the micromolar range. Most surprising in the series of the N-[4-(2-pyridyl)thiazol-2-yl]amides were the retained adenosine affinities by introduction of a cylopentanamide instead of the benzamide. A second series of compounds, the thiadiazolobenzamide series of compounds, revealed potent and selective adenosine receptor antagonists, especially N-(3-phenyl-1,2,4-thiadiazol 5-yl)-4-hydroxybenzamide (LUF5437, 8h) showing a K(i) value of 7 nM at the adenosine A(1) receptor and N-(3-phenyl-1,2,4-thiadiazol-5-yl)-4-methoxybenzamide (LUF5417, 8e) with a K(i) value of 82 nM at the adenosine A(3) receptor. 4 Hydroxybenzamide 8h is the most potent adenosine A(1) receptor antagonist of this new class of compounds. Structure--affinity relationships showed the existence of a steric restriction at the para-position of the benzamide ring for binding adenosine A(1) and A(3) receptors. The electronic nature of the 4-substituents played an important role in binding the adenosine A(3) receptor. Cis- and trans-4 substituted cyclohexyl derivatives were made next to the 4-substituted benzamide analogues. We used them to study the proposed specific interaction between the adenosine A(1) receptor and the 4-hydroxy group of this class of thiadiazolo compounds, as well as a suggested special role for the 4-methoxy group in binding the A(3) receptor. Both the adenosine A(1) and A(3) receptor slightly preferred the trans-analogues over the cis-analogues, while all compounds showed low affinities at the adenosine A(2A) receptor. Our investigations provided the potent and highly selective adenosine A(1) antagonist N-(3-phenyl-1,2,4 thiadiazol-5-yl)-trans-4-hydroxycyclohexanamide (VUF5472, 8m) showing a K(i) value of 20 nM. A third series of compounds was formed by urea analogues, N substituted with thiazolo and thiadiazolo heterocycles. The SAR of this class of compounds was not commensurate with the SAR of the previously described quinazoline urea. On the basis of these findings we suggest the existence of a special interaction between adenosine receptors and a region of high electron density positioned between the thia(dia)zole ring and phenyl(pyridyl) ring. Molecular electrostatic potential contour plots showed that for this reason the ligands need either a thiadiazole ring instead of a thiazole or a 2-pyridyl group instead of a phenyl. The derived novel classes of antagonists will be useful for a better understanding of the molecular recognition at the adenosine receptors. PMID- 11262086 TI - C-Isoprenylation of flavonoids enhances binding affinity toward P-glycoprotein and modulation of cancer cell chemoresistance. AB - Previous studies have shown that flavones bind to P-glycoprotein (Pgp) with higher affinity than isoflavones, flavanones, and glycosylated derivatives. In the present work, a series of C- or O-substituted hydrophobic derivatives of chrysin were synthesized to further investigate structural requirements of the A ring toward Pgp modulation. Increasing hydrophobicity at either position 6, 8, or 7 increased the affinity of in vitro binding to a purified cytosolic domain of Pgp, but only benzyl and 3,3-dimethylallyl C-substitution produced a high maximal quenching of the protein intrinsic fluorescence. Inhibition of membrane Pgp within leukemic cells, characterized by intracellular drug accumulation, was specifically produced by isoprenylated derivatives, with 8-(3,3 dimethylallyl)chrysin being even more efficient than the commonly used cyclosporin A. PMID- 11262087 TI - Electrooxidation potential as a tool in the early screening for new safer clozapine-like analogues. AB - The chemical modification of clozapine (1) has permitted the finding of new analogues, e.g., olanzapine (2), quetiapine (3), 5-(4-methylpiperazin-1-yl)-8 chloropyrido[2,3-b][1,5]benzoxazepine fumarate (9), with a clinical or psychopharmacological profile similar to that of clozapine. However, when developing new derivatives, the designers are discouraged by the development of clozapine-induced agranulocytosis. Different researchers have raised the role played by the oxidizability of the molecule in such a deleterious effect. In the present paper, we examined the oxidation profile (direct scavenging abilities, efficacy in inhibiting lipid peroxidation, and electrooxidation potential) of newly developed methoxy and trifluoromethylsulfonyloxy analogues related to clozapine, some of them being described as putative antipsychotic. The oxazepine derivative 7, unlike the other diazepine derivatives (6, 10--12), was not readily oxidized. Using a statistical predictive model for hematotoxicity previously described, 7 was found in the cluster of potentially nontoxic compounds while diazepine derivatives 6 and 10-12 were classified as potentially toxic compounds. Among these original compounds, 7, which presents a preclinical clozapine-like profile and a low sensitivity to oxidation, could be a promising antipsychotic candidate with low side effects. Considering the tricyclic derivatives examined so far, some elements of structure-oxidation relationship (SOR) might be pointed out. Regarding the nature of the tricyclic ring substituent, from the most to the least sensitive to oxidation, the sequence was as follows: HO > Cl > CH(3)O > CF(3)SO(2)O. The nature of the tricyclic ring influenced also the sensitivity to oxidation; the diazepine moiety appeared to be the most reactive ring compared to oxa- and thiazepine congeners. These parameters could be advantageously integrated in the early design of new safer clozapine-like analogues. PMID- 11262088 TI - New 3'-azido-3'-deoxythymidin-5'-yl O-(omega-hydroxyalkyl) carbonate prodrugs: synthesis and anti-HIV evaluation. AB - Prodrugs of zidovudine (AZT) have been synthesized in an effort to enhance its uptake by HIV-1 infected cells and its anti-HIV activity. The 5'-OH function of AZT was functionalized with various enzymatically labile alkyl groups using specific procedures. The prodrug moieties included 5'-O-carbonate, 5'-O carbamate, and 5'-O-ester. Analogues of the 3'-azido-3'-deoxythymidin-5'-yl O (omega-hydroxyalkyl) carbonate series were particularly interesting since they were rearranged through an intramolecular cyclic process during their enzymatic hydrolysis. Evidence of this prodrug rearrangement was confirmed by comparison of the serum half-lives of 5'-O-carbonate prodrugs with their corresponding 5'-O ester- and 5'-O-carbamate-AZT prodrugs. Interestingly, the anti-HIV-1 activities (EC(50)) of 3'-azido-3'-deoxythymidin-5'-yl O-(4-hydroxybutyl) carbonate 10 in acutely infected MT-4 cells and in peripheral blood mononuclear cells (PBMCs) were 0.5 nM and 0.78 nM, respectively. Compound 10 was 30 to 50 times more potent than its parent drug AZT. Our results suggest that the specific intramolecular rearrangement associated with the 3'-azido-3'-deoxythymidin-5'-yl O-(omega hydroxyalkyl) carbonate prodrugs could explain the remarkable anti-HIV-1 activity of this series of AZT prodrugs. Prodrug 10 may therefore have better clinical potential than AZT for the treatment of AIDS. PMID- 11262089 TI - Potential antidepressants displayed combined alpha(2)-adrenoceptor antagonist and monoamine uptake inhibitor properties. AB - Classical antidepressants are thought to act by raising monoamine (serotonin and noradrenaline) levels in the brain. This action is generally accomplished either by inhibition of monoamine metabolism (MAO inhibitors) or by blockade of monoamine uptake (tricyclic antidepressants and selective serotonin or noradrenaline reuptake inhibitors). However, all such agents suffer from a time lag (3--6 weeks) before robust clinical efficacy can be demonstrated. This delay may reflect inhibitory actions of noradrenaline at presynaptic alpha(2A) adrenergic auto- or heteroreceptors which gradually down-regulate upon prolonged exposure. Blockade of presynaptic alpha(2A)-adrenoceptors by an antagonist endowed with monoamine uptake inhibition properties could lead to new antidepressants with greater efficacy and a shorter time lag. In the literature, only two molecules have been described with such a pharmacological profile. Of these, napamezole (2) was chosen as a point of departure for the design of 4(5) [(3,4-dihydro-2-naphthalenyl)methyl]-4,5-dihydroimidazole (4a), which displayed the desired profile: alpha(2A)-adrenoceptor antagonist properties and serotonin/noradrenaline uptake inhibition. From this original molecule, a series of derivatives was designed and synthesized, encompassing substituted as well as rigid analogues. Structure-activity relationships permitted the selection of 14c (4(5)-[(5-fluoroindan-2-yl)methyl]-4,5-dihydroimidazole) as a development candidate. PMID- 11262090 TI - Syntheses and structure--activity relationships of novel apio and thioapio dideoxydidehydronucleosides as anti-HCMV agents. AB - On the basis of the fact that apio dideoxynucleosides, in which the furanose oxygen and the C2 of the 2,3-dideoxyribose are transposed, exhibited potent anti HIV activity and 2',3'-dideoxy-2',3'-didehydronucleosides also showed potent anti HIV activity, we synthesized apio dideoxydidehydronucleosides in which the oxygen atom and the double bond of the 2,3-dideoxy-2,3-didehydroribose are exchanged. The thioapio dideoxydidehydronucleosides were also synthesized since sulfur serves as a bioisostere of oxygen. Apio dideoxydidehydronucleosides 13a--f were synthesized starting from 1,3-dihydroxyacetone, utilizing phenylselenenyl chemistry as a key step. The ratio of the anomeric mixture was variable from 1:1 to 5:1 during the condensation of nucleosidic bases with the phenylselenyl acetate 11 in the presence of a Lewis acid. This is in contrast with other glycosyl donors such as 5-O-(tert-butyldiphenylsilyl)-2-phenylselenenyl-2,3 dideoxyribosyl acetate which shows excellent neighboring group effect (alpha:beta = 1:99). Thioapio dideoxydidehydronucleosides 22a,b were synthesized from the lactone 9 via thiolactone 17 as a key intermediate which was synthesized from dicyclohexylcarbodiimide coupling of the mercapto acid produced from the basic hydrolysis of thioacetate 16. The majority of apio analogues synthesized in this study exhibited moderate to potent anti-HCMV activity, among which the 5 fluorouracil derivative 13c was found to be the most potent against HCMV, while thioapio analogues showed no activity against HCMV. However, all synthesized compounds did not exhibit any significant activities against HIV-1, HSV-1, and HSV-2. The fact that apio dideoxydidehydronucleosides were active against HCMV suggests that the apio dideoxydidehydro sugar moiety can serve as a novel template for the development of new antiviral agents. PMID- 11262091 TI - Antipsoriatic anthrones with modulated redox properties. 5. Potent inhibition of human keratinocyte growth, induction of keratinocyte differentiation, and reduced membrane damage by novel 10-arylacetyl-1,8-dihydroxy-9(10H)-anthracenones. AB - The synthesis and structure-activity relationships (SARs) of a series of novel 10 arylacetyl-1,8-dihydroxy-9(10H)-anthracenones are described. Acylation of anthralin with either the appropriate arylacetyl chlorides or arylacetic acids in the presence of pyridine or via the coupling agent dicyclohexylcarbodiimide (DCC), respectively, furnished this structural class of antipsoriatic agents. Potential antipsoriatic activity was evaluated in complementary assays specifically addressed to three important aspects of psoriasis. First, several compounds were identified which are equally potent as inhibitors of human keratinocyte growth as the antipsoriatic agent anthralin. Furthermore, improved ratio of antiproliferative activity to cytotoxicity is demonstrated by the reduced potential of the novel analogues to induce membrane damage, which is a benefit of their reduced ability to generate oxygen radicals as documented by deoxyribose degradation. Second, analogue 3o bearing a hydroxamate functional group was also a highly potent inhibitor of LTB(4) biosynthesis in addition to its excellent antiproliferative activity. SARs of these inhibitors of both keratinocyte growth and LTB(4) biosynthesis with respect to the nature of the para-substitution in the 10-phenylacetyl side chain are discussed. Third, the compounds were also evaluated for their ability to induce the formation of cornified envelope protein in keratinocytes. Cross-linking of cellular protein as a marker of terminal differentiation of keratinocytes was observed for many 10 arylacetyl analogues at concentrations required to arrest cell growth. This newly uncovered activity of the novel anthracenones suggests antipsoriatic potential with respect to disturbance of keratinocyte differentiation, in addition to hyperproliferative and inflammatory aspects of psoriasis. PMID- 11262092 TI - Synthesis and Src kinase inhibitory activity of a series of 4-phenylamino-3 quinolinecarbonitriles. AB - Screening of a directed compound library in a yeast-based assay identified 4 [(2,4-dichlorophenyl)amino]-6,7-dimethoxy-3-quinolinecarbonitrile (2a) as a Src inhibitor. An enzymatic assay established that 2a was an ATP-competitive inhibitor of the kinase activity of Src. We present here SAR data for 2a which shows that the aniline group at C-4, the carbonitrile group at C-3, and the alkoxy groups at C-6 and C-7 of the quinoline are crucial for optimal activity. Increasing the size of the C-2 substituent of the aniline at C-4 of 2a from chloro to bromo to iodo resulted in a corresponding increase in Src inhibition. Furthermore, replacement of the 7-methoxy group of 2a with various 3 heteroalkylaminopropoxy groups provided increased inhibition of both Src enzymatic and cellular activity. Compound 25, which contains a 3 morpholinopropoxy group, had an IC(50) of 3.8 nM in the Src enzymatic assay and an IC(50) of 940 nM for the inhibition of Src-dependent cell proliferation. PMID- 11262093 TI - Syntheses and biological activities of a novel group of steroidal derived inhibitors for human Cdc25A protein phosphatase. AB - Silica gel supported pyrolysis of an azido-homo-oxa steroid led to rearrangement, presumably by a mechanism similar to that of solution phase Schmidt fragmentation, to produce a group of novel inhibitors for the oncogenic cell cycle regulator Cdc25A phosphatase. Cyano-containing acid 17, one of the best inhibitors in this group, inhibited the activity of Cdc25A protein phosphatase reversibly and noncompetitively with an IC(50) value of 2.2 microM. Structure activity relationships revealed that a phosphate surrogate such as a carboxyl or a xanthate group is required for inhibitory activity, and a hydrophobic alkyl chain, such as the cholesteryl side chain, contributes greatly to the potency. Without the cyano group, acid 26 and xanthate 27 were found to be more selective over Cdc25A (IC(50) = 5.1 microM and 1.1 microM, respectively) than toward CD45 (IC(50) > 100 microM, in each case), a receptor protein tyrosine phosphatase. Several of these inhibitors showed antiproliferative activities in the NCI 60 human tumor cell line screen. These steroidal derived Cdc25 inhibitors provide unique leads for the development of dual-specificity protein phosphatase inhibitors. PMID- 11262094 TI - Synthesis and photochromism of spirobenzopyran derivatives bearing an oxymethylcrown ether moiety: metal ion-induced switching between positive and negative photochromisms. AB - Spirobenzopyran derivatives carrying an oxymethylcrown ether moiety were synthesized, and their photochromism was studied in the presence of various metal ions in acetonitrile. The metal ion complexing ability of the crown ether moiety in crowned spirobenzopyrans affects both thermal isomerization and photoisomerization of their spirobenzopyran moiety to a great extent. When the interaction of the crown ether moiety with a metal ion was strong enough to cause thermal isomerization of the spirobenzopyran moiety to its corresponding merocyanine form and to suppress UV-induced isomerization to the merocyanine form, a negative photochromism appears. On the other hand, a relatively weak interaction of the crown ether moiety with a metal ion affords a positive photochromism. This phenomenon enables us to switch the photochromic behavior between positive and negative photochromisms. PMID- 11262096 TI - Effect of cyclodextrin on the intramolecular catalysis of amide hydrolysis. AB - The hydrolysis reaction of phthalamic acids (HOOCArCONHR, R = p-NO(2)Ph 1a, Ph 1b, adamantyl 1c) and N-phenyl maleamic acid 2b was studied in the presence of hydroxypropyl-beta-cyclodextrin (HPCD) in acid solution. The reactions of 1a and 1b were studied also in the presence of beta-cyclodextrin (beta-CD). All the compounds formed inclusion complexes with HPCD, and the association constant was determined from the change in absorption of the substrate when the host is added in the case of 1a (90 M(-)(1)) and 2b (49 M(-)(1)). For 1c ( 4 x 10(4) M(-)(1)) a competition method was used, and for 1b the association equilibrium constant was obtained from the kinetic data (37 M(-)(1)) because it is too reactive for the spectrophotometric method. Both cyclodextrins strongly inhibited the reactions, and analysis of the kinetic data for HPCD indicated that the reactions of complexed 1a, 1b, and 2b are at least 10-30 times slower than in the bulk solution whereas 1c reacts only 4.6 times slower when it is complexed. The inhibition is attributed to changes in the geometry of the substrate due to interaction of the carboxylic group and/or the amide with the OH at the rim of the cyclodextrin. The differences in the relative effect observed for 1c are attributed to the formation of a tighter complex with this substrate. PMID- 11262095 TI - Synthesis, aggregation, and binding behavior of synthetic amphiphilic receptors. AB - Amphiphilic bowl-shaped receptor molecules have been synthesized starting from diphenylglycoluril. Upon dispersion in water, these molecules self-assemble to form vesicles that bind neutral guests and alkali metal ions. In the case of bis(alkylester)-modified receptor compound 4, electron microscopy reveals that an increase in the size of the alkali metal ion (from Na(+) or K(+) to Rb(+) and to Cs(+)) leads to a change in the shape of the aggregates, viz. from vesicles to tubules. Monolayer experiments suggest that this behavior is due to a change in the conformation of this amphiphilic receptor. In water, molecules of 4 have an elongated conformation that changes to a sandwich-like one upon binding of alkali metal ions. Binding studies with vesicles from the bis-ammonium receptors 6 and 9 and the guest 4-(4-nitrophenylazo)resorcinol (Magneson) reveal that below the critical aggregation concentration (CAC) of the amphiphile 1:1 host-guest complexes are formed with high host-guest association constants. Above the CAC, a host-guest ratio of 2:1 was observed that indicates that only the cavities on the outside of the vesicle can be occupied. In the case of the naphthalene walled compound 8 changes in the vesicle structure are induced by the organic guest Magneson. PMID- 11262097 TI - Tetrapyrrole derivatives substituted with ferrocenylethynyl moieties. synthesis and electrochemical studies. AB - Up to eight redox-active ferrocenyl units have been incorporated, through the unsaturated ethynyl linkers, on the periphery of a series of cyclic tetrapyrrole derivatives including zinc(II) phthalocyanine and 2,3-naphthalocyanine, and nickel(II) meso-diphenylporphyrin. The synthesis of the former two macrocycles 4 and 7 involves the Sonogashira coupling reaction of ferrocenylethyne with 4,5 dichlorophthalonitrile (1) or 6,7-dibromonaphthalonitrile (5), respectively, followed by a base-promoted cyclization. The meso-bis(ferrocenylethynyl)porphyrin 11 has been prepared from the dibromo analogue 10 also by a palladium-catalyzed coupling reaction. These novel macrocyclic compounds have been spectroscopically and electrochemically characterized. As revealed by cyclic voltammetry, the ferrocenyl moieties appear to be electrochemically independent in these complexes and there is no significant electronic coupling among the iron(II) centers. PMID- 11262098 TI - New ferrocenyl chiral auxiliary substituents for amines: applications to syntheses of mossambine and vinblastine. AB - (+)-(R)-1,2-(alpha-(R)-Mesyloxy-beta-dimethyltetramethylene)-ferrocene was synthesized and used as a chiral auxiliary for N-alkylation of methyl 1,2,3,4,5,6 hexahydroazepino[4,5-b] indole-5-xi-carboxylates. Condensation with aldehydes then provided tetracyclic products in a diastereomeric ratio of at least 97:3. Gentle cleavage in acetic acid removed the chiral auxiliary to give the corresponding secondary amines in >99% ee. Thus, key intermediates leading to mossambine and vinblastine could be synthesized with high enantioselectivity. The enantioselectivity greatly exceeds that found with other chiral N-auxiliaries developed in our studies. PMID- 11262099 TI - Stereoisomerism of molecular multipropellers. 1. Static stereochemistry of bis- and tris-triaryl systems. AB - The static stereoisomerism of bis- and tris-triaryl systems has been analyzed by a systematic stereochemical analysis, and the resulting theoretical predictions have been experimentally confirmed by using reversed-phase HPLC and ESR and (1)H NMR spectroscopies with a family of seven distinct polychlorinated aromatic multipropellers. To analyze the static stereochemistry of these molecules, we have developed a specific procedure that uses a symmetry-adapted symbolic notation, allowing the theoretical prediction of both the number and symmetry of the isomers of the investigated molecules. Due to the steric hindrance introduced by the presence of bulky chlorine substituents, (all) conformational isomers can be characterized experimentally by several independent techniques confirming the theoretical stereochemical predictions. The different propeller moieties that constitute the molecule appear to be nearly independent of each other. Consequently, most of the observed isomers show comparable populations in solution at room temperature. PMID- 11262100 TI - Stereoisomerism of molecular multipropellers. 2. Dynamic stereochemistry of bis- and tris-triaryl systems. AB - The dynamic stereochemistry of bis- and tris-triaryl systems, the most simple "molecular multipropellers", is discussed on the basis of an extension of a systematic stereochemical analysis based on a symmetry-adapted symbolic notation developed specifically for these molecules. A suitable theoretical basis for our study is provided by the classical hypotheses concerning the dynamics of simple triaryl systems as formulated by Mislow and co-workers (J. Am. Chem. Soc. 1973, 95, 1535-1547), which, once applied to molecular multipropellers, show the existence of two modes of rearrangement for each propeller. Interconversion graphs for all molecules under study, covering a wide span of structural complexity, are presented. A complete NMR study of a two- and a three-propeller molecule indicates that all experimentally observable exchange pathways are indeed predicted by theoretical analysis. Moreover, quantitative analysis of 2D EXSY experiments affords the activation energy of the subset of pathways that give rise to observable interconversions on the NMR time scale. Assuming that two ring flips are the threshold mechanism for individual propeller interconversion, the experimental evidence indicates a preference for the flip of the central ring and one of the outer rings over the flip of two outer rings. PMID- 11262101 TI - NAD/NADH models with axial/central chiralities: superiority of the quinoline ring system. AB - Precursors of NAD model compounds 1c and 3a,b were successfully resolved into their atropisomers with respect to carbamoyl rotation. Atropisomers of quinoline derivatives are much more stable than pyridine derivatives as determined by cyclic voltammetry and X-ray crystallography. The 1,4-reduction of NAD model compound 4 was successfully achieved, affording novel NADH model compound 5. The rotational properties of the side chain of 5 were investigated by means of dynamic NMR. The rotational rate and syn/anti ratio, which indicate the orientation between carbonyl oxygen and hydrogen at the 4-position, are significantly affected by addition of magnesium ion. In the rotational transition state, the double-bond character of the C(carbonyl)-N(amide) bond is disrupted judging from the activation parameters. The oxidation of chiral 5 with p benzoquinone in the presence of magnesium ion catalyst gave predominantly one enantiomer of 4. On the other hand, oxidation of 5 with p-chloranil (tetrachloro p-benzoquinone) in the absence of magnesium ions affords the opposite enantiomer of 4 as the major product. The product enantiomer ratio is parallel to the syn/anti ratio in the starting material, indicating the importance of ground state conformation to stereochemistry of the reaction. PMID- 11262102 TI - An experimental and computational study of 1,2-hydrogen migrations in 2 hydroxycyclopentylidene and its conjugate base. AB - Thermal decomposition of alpha-hydroxydiazirine 2 gives primarily cyclopentanone and some allylic alcohol, in similar amounts as the known cyclohexyl analogue 1. Calculations (B3LYP/6-31+G) also show cyclopentanone to be the major product of this carbene rearrangement. Diazirine 2 and the lithium salt of the corresponding conjugate base 3 were decomposed by photolysis. The proportion of ketone formed increases with deprotonation, a trend also found computationally. In comparison, the base-induced isomerization of cyclopentene oxide, which proceeds via alpha elimination to a carbenoid intermediate similar to that obtained from 3, yields primarily allylic alcohol rather than ketone; neither ring size nor charge thus accounts for the unusual product distribution observed. Interestingly, the calculations reveal that in the gas phase with no counterion, the singlet, oxyanionic carbene, and the alpha-deprotonated epoxide are the same, rather than discrete structures. This intramolecular complexation stablilizes the oxyanionic carbene by 20-25 kcal/mol. PMID- 11262103 TI - New chiral molecular tweezers with a bis-Troger's base skeleton. AB - A convenient synthesis of 5alpha,8alpha,14alpha,17alpha-5,17:8,14-dimethano 5,8,14,17-tetraaza-5,6,7,8,13,14,17,18-octahydrodibenzo[e,e']benzo[1,2-a:3,4 a']dicyclooctene derivatives is described, and the compounds have been fully characterized by NMR; in some cases, the molecular structure has been determined by X-ray crystallography. These compounds represent the first examples of a new class of molecular tweezers. PMID- 11262104 TI - Straightforward asymmetric entry to highly functionalized medium-sized rings fused to beta-lactams via chemo- and stereocontrolled divergent radical cyclization of Baylis-Hillman adducts derived from 4-oxoazetidine-2 carbaldehydes. AB - DABCO promoted reactions of various activated vinyl systems with optically pure 4 oxoazetidine-2-carbaldehydes 1 gave rise to Baylis-Hillman adducts 3 with excellent syn stereoselectivities, without detectable racemization. Products 3 are used for the asymmetric preparation of unusual 2-azetidinones fused to medium sized rings via chemo- and stereocontrolled divergent radical cyclization. The formation of bicyclic beta-lactams 4-6 could be rationalized through a tandem radical Michael addition/endo cyclization or a tandem radical addition/Michael addition, depending on the electronic nature of the radical promoter. PMID- 11262105 TI - Crystal engineering of primary cubanecarboxamides: Repetitive formation of an unexpected N-H...O hydrogen-bonded network. AB - The unusual N-H.O hydrogen bond pattern in a family of primary cubanecarboxamides is described. 4-Chloro-1-cubanecarboxamide, 1, and the corresponding bromo and iodo derivatives, 2 and 3, form the "shallow-glide" hydrogen-bonded motif instead of the usual 5.1 A translated ribbon pattern, more characteristic of primary amides. This behavior is also seen, somewhat unexpectedly, for cubanecarboxamide, 4, but more or less unsurprisingly for 1,4-cubanedicarboxamide, 5. This repetitive occurrence of the same hydrogen bond pattern is of significance in crystal engineering wherein synthon robustness is measured in terms of such repetitivity. The cubyl group is directly responsible for the appearance of the shallow-glide motif in this family in preference to the 5.1 A translation pattern for two reasons: (1) steric--it is too large to fit in a 5.1 A translated structure and (2) electronic--its carbon acidity is sufficient to result in the appearance of C-H.O hydrogen bonds to the C=O group, disrupting any putative 5.1 A translated structure. Such a molecule --> supermolecule relationship is of value in crystal engineering strategies. PMID- 11262106 TI - Enzymatic coupling of specific peptides at nonspecific ligation sites: effect of Asp189Glu mutation in trypsin on substrate mimetic-mediated reactions. AB - Two main drawbacks seriously restrict the synthetic value of proteases as reagents in peptide fragment coupling: (i) native proteolytic activity and, thus, risk of undesired peptide cleavage; (ii) limited enzyme specificities restricting the amino acid residues between which a peptide bond can be formed. While the latter can be overcome by the use of substrate mimetics achieving peptide bond formation at nonspecific ligation sites, the risk of proteolytic cleavage still remains and hinders the wide acceptance of this powerful strategy for peptide coupling. This paper reports on the effect of the trypsin point mutant Asp189Glu on substrate mimetic-mediated reactions. The effect of this mutation on the steady-state hydrolysis of substrate mimetics of the 4-guanidinophenyl ester type and on trypsin-specific Lys- and Arg-containing peptides was investigated. The results were confirmed by enzymatic coupling reactions using substrate mimetics as the acyl donor and specific amino acid-containing peptides as the acyl acceptor. The competition assay verifies the predicted shift in substrate preference from Lys and Arg to the substrate mimetics and, thus, from cleavage to synthesis of peptide bonds. The combination of results obtained qualifies the trypsin mutant D189E as the first substrate mimetic-specific peptide ligase. PMID- 11262107 TI - A new tetratertiary phosphine ligand and its use in Pd-catalyzed allylic substitution. AB - A new tetraphosphine, the cis-cis-cis-1,2,3,4 tetrakis(diphenylphosphinomethyl)cyclopentane (Tedicyp) 1 has been synthesized, characterized, and used in Pd-catalyzed allylic substitutions. The Tedicyp was easily prepared in seven steps from the commercially available himic anhydride. The structure of the complex Tedicyp-borane was determined by X-ray analysis. The tetraphosphine in combination with [Pd(eta(3)-C(3)H(5))Cl](2) affords a very efficient catalyst for allylic substitution of several allylic acetates. Under mild conditions, very high turnover numbers and turnover frequencies have been obtained. PMID- 11262108 TI - Tetrahydroquinolizinium ylides: preparation and 1,3-dipolar cycloaddition. AB - By the Cu(II)-catalyzed reaction of 2-(4-diazo-3-oxoalkyl)pyridines (2), 4 alkoxycarbonyl (or 4-acyl)-3-oxo-1,2,3,4-tetrahydroquinolizinium ylides (3) were obtained in high yields. From the cycloaddition reaction of 3 with acetylenic esters (propynoates or acetylenedicarboxylates) the labile [2 + 3] cycloadducts, 3-oxo-3H-2a,4,5,8a-tetrahydropyrrolo[2,1,5-de]quinolizine-2a-carboxylates (8 or 12), were identified, which further reacted with DMAD (dimethyl acetylenedicarboxylate) to afford azocine derivatives (15 or 16) and produced pyrrolodihydroquinolizines (9 or 20) by dealkoxycarbonylation. PMID- 11262109 TI - Selective liquid-phase semihydrogenation of functionalized acetylenes and propargylic alcohols with silica-supported bimetallic palladium-copper catalysts. AB - Silica-supported, bimetallic palladium-copper catalysts were prepared in solution under mild conditions by reacting lithium di(4-tolyl)cuprate with palladium acetate in the presence of silica particles. Small bimetallic palladium-copper particles were deposited on the silica surface as confirmed with TEM-EDAX and EXAFS. The new material has been applied as catalyst in the liquid-phase semihydrogenation of mono- and disubstituted alkynes and showed high selectivity toward the cis-alkenes. The influence of addition of quinoline or potassium hydroxide to the semihydrogenation reaction mixture and the effects of exposure of the catalyst to air before use have been investigated. PMID- 11262110 TI - Cyclopropane-derived peptidomimetics. design, synthesis, and evaluation of novel Ras farnesyltransferase inhibitors. AB - Trisubstituted cyclopropanes have previously been established as rigid replacements of dipeptide arrays in several biological systems. Toward further evaluating the utility of these dipeptide mimics in the design of novel CA(1)A(2)X-based inhibitors of Ras farnesyltransferase (FTase), the conformationally constrained, diastereomeric pseudopeptides CAbuPsi[COcpCO]FM 7 9, the flexible analogue CAbuPsi[CHOHCH(2)]FM (10), and the tetrapeptide CAbuFM (6) were prepared. The orientations of the two peptide backbone substituents and the phenyl group on the cyclopropane rings in 7-9were specifically designed to probe selected topological features of the hydrophobic binding pocket of the A(2) subsite of FTase. The syntheses of the requisite trisubstituted cyclopropane carboxylic acid 22 and the diastereomeric cyclopropyl lactones 32a,b featured diastereoselective intramolecular cyclopropanations of chiral allylic diazoacetates and a new method for introducing side chains onto the C-terminal amino acid of cyclopropane-derived dipeptide replacements via the opening of an N Boc-aziridine with an organocuprate. These cyclopropane intermediates were then converted into the targeted FTase inhibitors 7-9 by standard peptide coupling techniques. The pseudopeptides 7-9 were found to be competitive inhibitors of Ras FTase with IC(50)s of 1055 nM for 7, 760 nM for 8, and 7200 nM for 9. The flexible analogue 10 of these constrained inhibitors exhibited a IC(50) of 320 nM and hence was slightly more potent than 7 and 8. All of these pseudopeptides were less potent than the tetrapeptide parent CAbuFM (6), which had an IC(50) of 38 nM. Because 7 and 8 are approximately equipotent, it appears that the orientation of the peptide backbone substituents on the cyclopropane rings in 7 and 8 do not have any significant effect on binding affinity and that multiple binding modes are possible without significant changes in affinity. On the other hand, this flexibility does not extend to the orientation of the side chain of the A(2) residue as 7 and 8 were both nearly 1 order of magnitude more potent than 9. Comparison of the relative potencies of 6 and 10 suggests that the amide linkage between the A(1) and the A(2) residues of CA(1)A(2)X-derived FTase inhibitors is important. PMID- 11262111 TI - A direct reduction of aliphatic aldehyde, acyl chloride, ester, and carboxylic functions into a methyl group. AB - The aliphatic carboxylic group was efficiently reduced to the methyl group by HSiEt(3) in the presence of catalytic amounts of B(C(6)F(5))(3). To the best of our knowledge, this is the first example of a direct exhaustive reduction of aliphatic carboxylic function. Aliphatic aldehydes, acyl chlorides, anhydrides, and esters also underwent complete reduction under similar reaction conditions. Aromatic carboxylic acids, as well as other carbonyl functional equivalents, underwent smooth partial reduction to the corresponding TES-protected benzylic alcohols. It was shown that, unlike the reduction of aliphatic substrates, the exhaustive reduction of aromatic substrates was not straightforward: a concurrent Friedel-Crafts-like alkylation process competed with the reduction yielding trace to notable amounts of dimeric products, thus decreasing the overall selectivity of the reduction process. PMID- 11262112 TI - A facile and highly diastereoselective aziridination of chiral camphor n enoylpyrazolidinones with n-aminophthalimide. AB - Reaction of various chiral camphor N-enoylpyrazolidinones 2a-g with N aminophthalimide in the presence of lead tetraacetate in CH(2)Cl(2) proceed smoothly afford the corresponding N-phthalimidoaziridines (3a-e, 4f-g) with excellent material yields (86-95%) at room temperature in 5 min. High levels of diastereoselectivities (up to >95:5 dr) were obtained. The solvent effect was investigated, and the auxiliary can be easily recovered in high yields under mild reaction conditions. PMID- 11262113 TI - Electrochemical and ferromagnetic couplings in 4,4',4' '-(1,3,5 benzenetriyl)tris(phenoxyl) radical formation. AB - 4,4',4' '-(1,3,5-Benzenetriyl)tris(2,6-di-tert-butylphenol) was prepared by the cross-coupling of 1,3,5-tribromobenzene and [4-(trimethylsiloxy)phenyl]magnesium bromide. X-ray analysis of the single crystal showed a propeller-like structure with a mean dihedral angle of 39 degrees between the hydroxyphenyl and the core benzene. The phenoxyl mono-, di-, and triradicals were generated by the electrochemical oxidation of the trianion. A stepwise radical formation was revealed by a differential pulse voltammogram, electrolytic ESR spectroscopy, and a comproportionation reaction between the radicals, which was discussed as an effect of the pi-conjugated but non-Kekule-type coupler. The quartet and triplet ground state for the tri- and diradical, respectively, were confirmed by a SQUID measurement. PMID- 11262114 TI - Control of the regioselectivity in catalytic transformations involving amphiphilic bis-allylpalladium intermediates: mechanism and synthetic applications. AB - Various dialkyl-substituted allyl chloride derivatives (2d-i) undergo regioselective palladium-catalyzed coupling reactions with allylstannanes (1a,b) and benzylidenemalonitrile (4), providing functionalized 1,7-octadienes in good yield. The catalytic reaction proceeds through an unsymmetrical amphiphilic bis allylpalladium intermediate. An introductory electrophilic attack on the terminal position of the unsubstituted allyl moiety is followed by a nucleophilic attack on the alkyl-substituted allyl ligand. A theoretical analysis was performed by applying density functional theory at the B3PW91/DZ+P level to study the substituent effects on the electrophilic attack. According to the theoretical results, the high regioselectivity can be ascribed to the electronic effects of the alkyl substituents: The terminal alkyl groups destabilize the eta(1),eta(3) bis-allylpalladium intermediate of the reaction; in addition, the alkyl substitution increases the activation barrier for the electrophilic attack. PMID- 11262115 TI - An examination of hyperconjugative and electrostatic effects in the hydride reductions of 2-substituted-4-tert-butylcyclohexanones. AB - To better understand electronic effects on the diastereoselectivity of nucleophilic additions to the carbonyl group, a series of 2-X-4-tert butylcyclohexanones (X = H, CH(3), OCH(3), F, Cl, Br) were reacted with LiAlH(4). Reduction of ketones with equatorial substituents yields increasing amounts of axial alcohol in the series for X [H < CH(3) < Br < Cl < F << OCH(3)]. These data cannot be explained by steric or chelation effects or by the theories of Felkin Anh or Cieplak. Instead, an electrostatic argument is introduced: due to repulsion between the nucleophile and the X group, axial approach becomes energetically less favorable with an increase in the component of the dipole moment anti to the hydride approach trajectory. The ab initio calculated diastereoselectivities were close to the experimental values but did not reproduce the relative selectivity ordering among substituents. For reduction of ketones with axial substituents, increasing amounts of axial alcohol are seen in the series for X [Cl < Br < CH(3) < OCH(3) < H < F]. After some minor adjustments are made, this ordering is consistent with both the electrostatic model and Felkin-Anh theory. Cieplak theory cannot account for these data regardless of adjustments. Ab initio calculated diastereoselectivities were reasonably accurate for the nonpolar substituents but were poor for the polar substituents. PMID- 11262116 TI - A consecutive double-Criegee rearrangement using TFPAA: stepwise conversion of homoadamantane to oxahomoadamantanes. AB - Rearrangement of 4-methylhomoadamantan-4-ol (1) with trifluoroperacetic acid (TFPAA) in trifluoroacetic acid (TFAA) proceeds with the formation of 4 oxahomoadamantane 6 and its derivatives (4 and 5). 2-exo-Hydroxy-4 oxahomoadamantane (5) and 6 were identified as a result of consecutive O insertion Criegee rearrangement processes. The absence of methyl trifluoroacetate and methyl trifluoroperacetate among the reaction products, as well as the presence of acetyltrifluoroacetyl peroxide, is consistent with a double rather that a triple oxygen insertion during the course of the Criegee reaction. A mechanism involving initial Criegee rearrangement followed by a Baeyer-Villiger reaction is also excluded by kinetic considerations. The parallel formation of 4 ethyl-3-oxahomoadamantan-2-one (4) was determined to be the result of 4 methylhomoadmantan-4-ol (3) dehydration, with subsequent epoxidation of 4 methylhomoadamant-4-ene (32) to 4,5-epoxy-4-methylhomoadamantane (33), acid catalyzed isomerization of 33 to 3-methylhomoadamantan-2-one (34), and Baeyer Villiger oxidation to 3-methyl-5-oxabishomoadamantan-6-one (35). This sequence of reactions was followed by the acid-catalyzed isomerization to the final product 4. The proposed mechanisms for these transformations are discussed on the basis of model experiments and supporting density functional theory (DFT) calculations. PMID- 11262117 TI - The first total synthesis of concanamycin f (concanolide a). AB - A highly stereoselective total synthesis of the macrolide antibiotic concanamycin F (1), a specific and potent inhibitor of vacuolar H(+)-ATPase, has been achieved by a convergent route involving the synthesis and coupling of its 18-membered tetraenic lactone and beta-hydroxyl hemiacetal side chain subunits. The C1-C19 18 membered lactone aldehyde 4 was synthesized through the intermolecular Stille coupling of the C5-C13 vinyl iodide 24 and the C14-C19 vinyl stannane 25, followed by construction of the C1-C4 diene and macrolactonization. Synthesis of 4 via a second convergent route including the esterification of the C1-C13 vinyl iodide 45 and the C14-C19 vinyl stannane 47 followed by the intramolecular Stille coupling was also realized. The highly stereoselective aldol coupling of 4 and the C20-C28 ethyl ketone 5 followed by desilylation provided 1 which was identical with natural concanamycin F. PMID- 11262118 TI - Formal total synthesis of (+/-)-gamma-lycorane and (+/-)-1-deoxylycorine using the [4+2]-cycloaddition/rearrangement cascade of furanyl carbamates. AB - The total syntheses of gamma-lycorane and (+/-)-1-deoxylycorine were accomplished using an intramolecular Diels-Alder cycloaddition of a furanyl carbamate as the key step. The initially formed [4+2]-cycloadduct undergoes nitrogen-assisted ring opening followed by deprotonation/reprotonation of the resulting zwitterion to give a rearranged hexahydroindolinone. The stereochemical outcome of the IMDAF cycloaddition has the side arm of the tethered alkenyl group oriented syn with respect to the oxygen bridge. The key intermediate used in both syntheses corresponds to hexahydroindolinone 20. Removal of the t-Boc group in 20 followed by reaction with 6-iodobenzo[1,3]dioxole-5-carbonyl chloride afforded enamide 22. Treatment of this compound with Pd(OAc)(2) employing the Jeffrey modification of the Heck reaction provided the galanthan tetracycle 24 in good yield. Compound 24 was subsequently converted into (+/-)-gamma-lycorane using a four-step procedure to establish the cis-B,C-ring junction. A radical-based cyclization of the related enamide 33 was used for the synthesis of 1-deoxylycorine. Heating a benzene solution of 33 with AIBN and n-Bu(3)SnH at reflux gave the tetracyclic compound 38 possessing the requisite trans fusion between rings B and C in good yield. After hydrolysis and oxidation of 38 to 40, an oxidative decarboxylation reaction was used to provide the C(2)(-)C(3)(-)C(12) allylic alcohol unit characteristic of the lycorine alkaloids. The resulting enone was eventually transformed into (+/-)-1-deoxylycorine via known synthetic intermediates. PMID- 11262119 TI - Preparation of stilbene-tethered nonnatural nucleosides for use with blue fluorescent antibodies. AB - The synthesis of the first examples of stilbene-tethered hydrophobic C nucleosides is described. Compounds of this type are targeted for use with our recently reported "blue-fluorescent antibodies" with the aim of probing native and nonnatural DNA. The nucleophilic addition of aryl Grignard reagents to either a protected 2'-deoxy-1'-chloro-ribofuranose or a protected 2'-deoxy-ribonolactone was the key synthetic step and afforded C-nucleosides in good yields. Both routes resulted in a final product that was >/=90% of the beta-anomer. Amide- and ether based linkers for attachment of trans-stilbene to the nucleobase were assessed for utility during synthesis and in binding of the ligands to a blue-fluorescent monoclonal antibody. X-ray structures of each complex were obtained and serve as a guideline for second-generation stilbene-tethered C-nucleosides. The development of these hydrophobic nucleosides will be useful in current native and nonnatural DNA studies and invaluable for investigations regarding novel, nonnatural genomes in the future. PMID- 11262121 TI - Carbonyl- and carboxyl-substituted enediynes: synthesis, computations, and thermal reactivity. AB - The influence of electron-withdrawing groups (carbonyl and carboxyl) at the alkyne termini on the reactivity of enediynes was investigated by a combination of experimental and computational techniques. While the general chemical reactivity of such enediynes, especially if non-benzannelated, is increased markedly, the thermal cyclization, giving rise to Bergman cyclization products, is changed little relative to the parent enediyne system. This is evident from kinetic measurements and from density functional theory (DFT, BLYP/6-31G + thermal corrections) computations of the experimental systems which show that the Bergman cyclization barriers slightly (3-4 kcal/mol) increase, in contrast to earlier theoretical predictions. The effect on the endothermicities is large (DeltaDeltaH(r) = 7-12 kcal/mol). Hence, the increased reactivity of the substituted enediynes is entirely due to nucleophiles or radicals present in solution. This was demonstrated by quantitative experiments with diethylamine and tetramethyl piperidyl oxide (TEMPO) which both give fulvenes through 5-exo-dig cyclizations. PMID- 11262120 TI - Bengamides revisited: new structures and antitumor studies. AB - The structural chemistry and biological activity of the bengamide class of compounds have been further characterized. Extracts prepared from recollected Jaspis cf. coriacea from five sites in Fiji were pooled. Six new bengamides, M (7b), N (8a), O (8b), P (9a), Q (9b), and R (10), were identified, accompanied by the known bengamides A (1a), B (1b), E (3a), F (3b), Y (5), Z (6), L (7a), G (11a), H (11b), and I (12). The structures of the new compounds were determined from spectroscopic data, and some were additionally confirmed by semisynthesis. Cytotoxicity screening data were obtained from the NCI-DTP 60 cell screen for bengamides A, B, and P. Bengamides A and B were more potent than bengamide P, with average IC(50) values of 0.046, 0.011, and 2.70 FM, respectively. The in vitro antitumor activity against MDA-MB-435 human mammary carcinoma was also determined for natural bengamides A, B, E, F, P, M, O, and Z and for synthetic samples of B and O. The best activity was observed for the natural bengamides A (IC(50) = 1 nM) and O (IC(50) = 0.3 nM). PMID- 11262122 TI - Highly functionalized five-membered carbocycles from (3-dialkylamino-1 ethoxyalkenylidene)pentacarbonylchromium complexes and alkynes: the effects of substituents, solvents, ligand additives, and reagent concentrations on the product distribution. AB - The cocyclization reaction of pentacarbonyl(beta-amino-1 ethoxyalkenylidene)chromium complexes 1 with alkynes has been studied with respect to the effects of substituents, solvents, ligand additives, and reagent concentrations upon the product distribution. This reaction proceeds either as a formal [2 + 2 + 1] cycloaddition to give 5-(1'-dialkylaminoalkylidene)-4 ethoxycyclopent-2-enones 8 or a formal [3 + 2] cycloaddition to give 5 dialkylamino-3-ethoxy-1,3-cyclopentadienes 9. A working hypothesis for the mechanism of this reaction is proposed on the basis of that previously determined for the Dotz reaction. The effects of the aforementioned parameters upon the product distribution of this current reaction are explained in terms of this model. A pronounced ligand-induced allochemical effect has been observed. Conditions for the selective preparation of both classes of cycloadducts 8 and 9 have been determined. PMID- 11262123 TI - Synthesis of carbobicyclic compounds via palladium-catalyzed cyclization/hydrosilylation: evidence for reversible silylpalladation. AB - Cyclization/hydrosilylation of substituted 1-vinyl-1-(3-butenyl)cycloalkanes catalyzed by a 1:1 mixture of (phen)Pd(Me)Cl (1) and NaBAr(4) [phen = 1,10 phenanthroline; Ar = 3,5-C(6)H(3)(CF(3))(2)] formed silylated spirocycles in high yield with excellent regio and diastereoselectivity. Cyclization/hydrosilylation of substituted 3-(3-butenyl)cycloalkenes or 2,3-diallyl-5,6-dimethyl-1,4 hydroquinone diacetate (16) formed silylated fused bicyclic complexes in good yield. Reaction of substituted 1,6,11-nonatrienes with silane catalyzed by 1/NaBAr(4) led to cascade cyclization with hydrosilylation. This latter procedure was employed in the synthesis of silylated bicyclopentanes and a linear triquinane. PMID- 11262125 TI - Stereospecific synthesis of alpha- and beta-C-glycosides from glycosyl sulfoxides: scope and limitations. AB - C-Glycosides derived from alpha-L-fuco-, alpha-D-gluco-, beta-D-gluco-, and alpha D-mannopyranose have been synthesized from the corresponding glycosyl phenyl sulfoxide through phenylsulfinyl-lithium exchange, to generate an anomeric carbanion, and subsequent reaction with a carbon electrophile. The reactions were stereospecific and proceeded with retention of the configuration at the anomeric center. Improved yields of C-glycosides were obtained by an inverse addition protocol. Trapping of the anomeric carbanion with aldehydes gave best results. Reaction with ketones, chloroformates, nitriles, and alkyl halides was also explored. Mechanistic aspects of the reaction are discussed. PMID- 11262126 TI - Photo-Fries rearrangements of phenyl phenylacylates in polyethylene films: comparison of reactivity and selectivity with 1-naphthyl phenylacylates. AB - The fates and kinetics of recombination of singlet radical pairs generated by photolyses of three phenyl phenylacylates have been examined in unstretched and stretched polyethylene films. Comparisons with results from photolyses of analogous 1-naphthyl phenylacylates in the same media lead to the conclusions that (1) phenoxy is less reactive overall than 1-naphthoxy toward a common phenylacyl radical but (2) the constrained cages in which the radical pairs reside exert greater control over the movements of the 1-naphthoxy/phenylacyl pairs. The reasons for these observations are discussed in the context of the shapes and van der Waals volumes of the radical pairs, the void volumes of sites in native polyethylene films, and the electronic properties of the aryloxy radicals. PMID- 11262124 TI - Allylated monosaccharides as precursors in triple reductive amination strategies: synthesis of castanospermine and swainsonine. AB - The feasibility of the triple-reductive amination reaction for the synthesis of complex indolizidine frameworks is illustrated by application to the potent glycosidase inhibitors castanospermine and swainsonine. The target compounds were obtained from known carbohydrate precursors in yields of 23 and 14%, over nine and 13 steps, respectively. The iodoetherification reaction of allylated monosaccharides was shown to be a practical reaction for the synthesis of the tricarbonyl precursors for the key triple reductive amination reactions. PMID- 11262127 TI - Stereocontrolled synthesis of acyclic 1,3-diols via condensation of tungsten-syn pi-pentadienyl complexes with aldehydes. A new Prins reaction via s-trans-diene cationic intermediates. AB - In the presence of BF(3).Et(2)O, condensation of CpW(CO)(2)(syn-pi-2 methoxycarbonylpentadienyl) with aldehydes generated tungsten-eta(4)-trans-diene cation in cold toluene, and hydrolysis of this salt afforded tungsten-pi-allyl anti-1,3-diols in good yields. This new synthesis of anti-1,3-diols represents an atypical Prins reaction that is applicable to normal aldehydes. The anti/syn ratios of 1,3-diols increased with an increase in the size of the aldehydes. These anti-1,3-diols were transformed into various complex oxygen heterocycles based on two demetalations: (1) conversion to an allyl cation followed by nucleophilic attack and (2) condensation with aldehydes via its CpW(NO)Cl derivative, to give functionalized alpha-methylene butyrolactones. A semi emperical calculation was performed to deduce the transition-state structure to rationalize the anti-stereoselectivity. PMID- 11262128 TI - Synthesis and reactivity of allenes substituted by selenenyl groups at 1- and 3 positions. AB - 1,3-Bis(methylseleno)- and 1,3-bis(benzylseleno)-1,3-diphenylpropadienes were synthesized by reaction of Ph(2)C(3) dianion, prepared from 1,3-diphenylpropyne and n-butyllithium, with dimethyl diselenide or benzylselenocyanate in the presence of TMEDA, and reaction of the dianion with a mixture of dimethyl diselenide and benzylselenocyanate yielded 1-benzylseleno-3-methylselenoallene along with the symmetric allenes. Diselenocyclic allenes and tetraselenocyclic bisallenes were also obtained by reacting the dianion with corresponding alkane diselenocyanates. The thermal reaction of the 1,3-bis(alkylseleno)allenes mainly afforded enediynes through radical pathway, and the nine-membered cyclic allene provided intramolecular cyclization product via an intramolecular rearrangement. Heating of the cyclic bisallenes gave compounds derived from intramolecular cyclization products together with a small amount of the enediynes. Irradiation of allenes caused rearrangement of the selenenyl group to give alkynes, and the alkynes also reacted photochemically to yield the enediynes. PMID- 11262129 TI - Palladium-catalyzed allylic alkylation using chiral hydrazones as ligands. AB - Palladium-catalyzed asymmetric allylic alkylation of 1,3-diphenyl-2-propenyl acetate (4) with a dimethyl malonate-BSA-LiOAc system and its derivatives has been successfully carried out in the presence of a new chiral hydrazone ligands such as 2-(diphenylphosphino)benzaldehyde SAMP hydrazone (DPPBA-SAMP) (3a) in high yields with high enantioselectives. PMID- 11262130 TI - Phosphoramidite coupling to oligonucleotides bearing unprotected internucleosidic phosphate moieties. AB - The coupling of 2-cyanoethyl thymidine phosphoramidite to solid-support-bound, phosphate-unprotected oligothymidylates and their phosphorothioate analogues was studied. The yield of the coupling reaction depended on the pK(BH)()+ values of protonated nitrogen bases that served as counterions to the phosphodiester functions of oligonucleotides. To maximize the coupling efficiency, the oligonucleotides were detritylated and washed with a mixture of 0.1 M DMAP and 0.1 M 1H-tetrazole, which resulted in a 98+% coupling efficiency. The utility of the results was demonstrated in the preparation of oligonucleotides with a mixed backbone that required the successive use of H-phosphonate and phosphoramidite methods of synthesis. Using this approach, 20-mer antisense oligonucleotides containing 2'-O-(2-methoxyethyl) ribonucleoside residues and phosphorothioate and phosphoramidate internucleosidic linkages were synthesized in high yield. PMID- 11262132 TI - Synthesis of novel 2-azabicyclo[2.2.0]- and [2.1.1]hexanols. AB - Methyl- and phenyl-substituted N-(ethoxycarbonyl)-2-azabicyclo[2.2.0]hex-5-enes 6 were reacted with NBS in wet DMSO to afford bromohydrins. Mixtures of unrearranged 6-exo-bromo-5-endo-hydroxy-2-azabicyclo[2.2.0]hexanes 7a,b and rearranged 5-anti-bromo-6-anti-hydroxy-2-azabicyclo[2.1.1]hexanes 8a,b were formed stereoselectively from the parent alkene 6a and 4-methyl alkene 6b. The 5 methyl alkene 6c affords only unrearranged bromohydrin 7c and dibromohydrin 9. By contrast, solely rearranged 3-endo-substituted-2-azabicyclo[2.1.1]hexane bromohydrins 8d-f result from additions to 3-endo-methyl alkene 6d, 3-endo-4 dimethyl alkene 6e, and 3-endo-phenyl alkene 6f. As an alternative route to bromohydrins, the parent 5,6-exo-epoxide 10a and 5-endo-methyl-5,6-exo-epoxide 10b were ring opened with bromine/triphenylphosphine to afford unrearranged 5 endo-bromo-6-exo-hydroxy-2-azabicyclo[2.2.0]hexanes 11a,b, while the 3-endo methyl epoxide 10c afforded solely the rearranged 5-anti-bromo-6-anti-hydroxy-3 exo-methyl-2-azabicyclo[2.1.1]hexane isomer 8g. Tributyltin hydride reduction of bromohydrins 7a,b and 11a afforded novel 2-azabicyclo[2.2.0]hexan-5-ols 13a,b and -6-ol 14, and bromohydrins 8a,b, 8d-g afforded new 2-azabicyclo[2.1.1]-hexan-5 ols 15a,b and 15d-g. PMID- 11262131 TI - 2-Azabicyclo[2.1.1]hexanes. 2. Substitutent effects on the bromine-mediated rearrangement of 2-azabicyclo[2.2.0]hex-5-enes. AB - Methyl- and phenyl-substituted N-(ethoxycarbonyl)-2-azabicyclo[2.2.0]hex-5-enes 6 have been prepared by photoirradiation of appropriately substituted 1,2 dihydropyridines. Torquoselectivity is observed in the synthesis of the 3-endo methyl- and 3-endo-phenyl-2-azabicyclo[2.2.0]hexenes 6c-e from 2-methyl- and 2 phenyl-1,2-dihydropyridines 5c-e. Products formed upon addition of bromine to 3 endo-, 4-, and 5-methyl- and 3-endo-phenyl-substituted N-(ethoxycarbonyl)-2 azabicyclo[2.2.0]hex-5-enes 6a-f were substituent dependent. For 6a,b, which lack substituents at C(3) or C(5), mixtures of unrearranged dibromides 8a,b and rearranged dibromides 9a,b were obtained. With the 3-endo-substituents in 6c-e, only rearranged dibromides 9c-e were formed; 5-methyl substitution afforded mainly unrearranged dibromide 8f and some allylic bromide 10. Both unrearranged 5 endo,6-exo-dibromo-2-azabicyclo[2.2.0]hexanes 8 and rearranged 5-anti-6-anti dibromo-2-azabicyclo[2.1.1]hexanes 9 are formed stereoselectively. The dibromoazabicyclo[2.1.1]hexanes 9 have been reductively debrominated to afford the first reported 2-azabicyclo[2.1.1]hexanes 11 with alkyl or aryl substituents at C-3. PMID- 11262133 TI - Enantioselective synthesis and stereoselective rearrangements of enol ester epoxides. AB - Enol esters can be epoxidized with high enantioselectivities using the fructose derived chiral ketone 1 as catalyst and Oxone as oxidant. A detailed study of enantiomerically enriched enol ester epoxides has revealed that the acid catalyzed rearrangement can proceed through two distinct pathways, one with retention of configuration and the other with inversion. The competition between the two pathways is highly dependent upon the nature of the acid catalyst. A strong acid favors retention of configuration and a weak acid favors inversion of configuration. Under thermal conditions, these epoxides rearrange highly stereoselectively with inversion of configuration. Either enantiomer of an alpha acyloxy ketone can be formed from one enantiomer of an enol ester epoxide by judicious choice of reaction conditions. PMID- 11262134 TI - Iminopropadienethiones, Ar=N=C=C=C=S. AB - Aryliminopropadienethiones 9 have been generated by flash vacuum thermolysis of isoxazolones of the type 5 and characterized by mass spectrometry and matrix isolation IR spectroscopy in conjunction with DFT calculations and chemical trapping. PMID- 11262135 TI - Synthesis and metabolism of the first thia-bilirubin. AB - A symmetrical C(10)-thiabilirubin analogue, 8,12-bis(2-carboxyethyl)-2,3,17,18 tetraethyl-7,13-dimethyl-10-thia-(21H,23H,24H)-bilin-1,19-dione (1), was synthesized from 8-(2-carboxyethyl)-2,3-diethyl-7-methyl-10H-dipyrrin-1-one in one step by reaction with sulfur dichloride. The thia-rubin exhibited the expected IR, UV-vis, and NMR spectroscopic properties, which are rather similar to those of mesobilirubin-XIIIalpha. Like bilirubin and mesobilirubin, 1 adopts an intramolecularly hydrogen-bonded conformation, shaped like a ridge-tile but with a steeper pitch. The longer C-S bond lengths and smaller bond angles at C-S C, as compared to C-CH(2)-C, lead to an interplanar angle between the two dipyrrinones of only 74 degrees -or considerably less than that of bilirubin (approximately 100 degrees). On normal- and reversed-phase chromatography, 1 is substantially less polar than bilirubin. Despite this conformational distortion, 1 is metabolized in normal rats to acyl glucuronides, which are secreted into bile. In mutant (Gunn) rats lacking bilirubin glucuronosyl transferase, 1 (like bilirubin) was not excreted in bile. PMID- 11262136 TI - Excited- and ground-state versions of the tri-pi-methane rearrangement: mechanistic and exploratory organic photochemistry. AB - The di-pi-methane rearrangement with two pi-groups bonded to a single carbon leading to pi-substituted cyclopropanes is now well established. The present research had as its goal the exploration of molecular systems having three pi moieties attached to an sp(3)-hybridized atom in a search for a tri-pi-methane rearrangement. Indeed, it was found that such systems do rearrange photochemically to afford cyclopentenes. However, it was also established that vinylcyclopropanes ring-expand to cyclopentenes on direct irradiation. Since both three-ring and five-ring photoproducts often are found to be produced, it was important to establish that the observed photochemistry was really the result of a true single-step tri-pi-methane rearrangement and not the consequence of two sequential rearrangements, first to form a vinyl cyclopropane which subsequently ring expanded to the cyclopentene. The general situation has three species-A, B, and C-corresponding to tri-pi-methane reactant A, vinylcyclopropane photoproduct B, and cyclopentene photoproduct C. Three rate constants are involved, k(1) for A --> B, k(2) for A --> C, and k(3) for B --> C. The kinetics were applied to two examples with provision to avoid differential light absorption; this utilized singlet sensitization. It was determined that direct formation of the cyclopentene photoproduct proceeds more rapidly than the ring-expansion route. In contrast to the di-pi-methane rearrangement, the tri-pi-methane reaction was found to be preferred by the singlet, while in these sterically congested systems, the triplet led to di-pi-methane reactivity. Finally, a ground-state counterpart of the reaction was obtained. PMID- 11262137 TI - Intramolecular Pd-catalyzed carbocyclization, Heck reactions, and aryl-radical cyclizations with planar chiral arene tricarbonyl chromium complexes. AB - (o-butenylhalobenzene)Cr(CO)(3) complexes were synthesized by diastereoselectve allylmetal additions to o-halo benzaldehyde complexes. The addition of allylZnBr proved particularly convenient and clean. The complexes undergo intramolecular Pd catalyzed cyclizations (Heck reactions) without decomplexation and/or alkene isomerization. In complexes with a benzylic stereogenic center, the diastereoselectivity of the alkene carbopalladation is governed by the planar chirality of the complex rather than by the benzylic stereogenic center in the side chain. This reaction outcome can be rationalized by the geometry of the arene plane vs that of the Pd coordination plane in the transition step of the alkene carbopalladation step. An alternative cyclization procedure involves the generation of a Cr(CO)(3)-coordinated arene radical from the bromo and iodo complexes. Intramolecular aryl-radical cyclization affords indan complexes. The transition metal arene pi-bond remains intact during this process. PMID- 11262138 TI - Fragmentation of carbohydrate anomeric alkoxy radicals. synthesis of polyhydroxy piperidines and pyrrolidines related to carbohydrates. AB - Imino sugars of the piperidine and pyrrolidine types can be specifically obtained when protected 5-amino-5-deoxyfuranopentoses, 5-amino-5-deoxyfuranohexoses, 6 amino-6-deoxyfuranohexoses, and 6-amino-6-deoxypyranohexoses undergo a tandem alkoxy radical beta-fragmentation-intramolecular cyclization reaction. The reaction is promoted by the system: iodosylbenzene-iodine under mild conditions. The tert-butoxycarbonyl, benzyloxycarbonyl, and diphenylphosphinoyl radicals have been studied as amino-protecting groups. Using this methodology, polyhydroxylated pyrrolidines of the D-erythrofuranoses 34 and 35, D-threofuranose 36, L xylofuranose 42, and D-arabinofuranose 43 series and polyhydroxylated piperidines of the D-arabinopyranose type 37 and 38 were obtained. PMID- 11262139 TI - 2-ethynylaziridines as chiral carbon nucleophiles: stereoselective synthesis of 1,3-amino alcohols with three stereocenters via allenylindium reagents bearing a protected amino group. AB - Allenylindium reagents bearing a protected amino group were effectively prepared from N-protected 3-alkyl-2-ethynylaziridines by treatment with InI in the presence of Pd(PPh(3))(4) and H(2)O. Stereoselective addition of the allenylindium to aliphatic or aromatic aldehydes affords 1,3-amino alcohols bearing three contiguous chiral centers: while 2,3-trans-2-ethynylaziridines yield syn,syn-2-ethynyl-1,3-amino alcohols predominantly, 2,3-cis-aziridines give anti,syn isomers selectively. Synthesis of highly substituted ethynylazetidines is also described. PMID- 11262140 TI - Total synthesis of the four enantiomerically pure diastereomers of 8-F(2t) isoprostane. AB - Syntheses of the four enantiomerically pure diastereomers of 8-F(2t)-isoprostane (5-8) are described. The key to this approach was to prepare the racemic alcohol 9 in high diastereomeric purity and then resolve 9 by lipase-mediated acetylation to yield the enantiomerically pure alcohols 30 and 32. PMID- 11262142 TI - Photoinduced 1,3-proton shift in methyldithiepines as a potential way of modulating hyperpolarizabilities. PMID- 11262141 TI - The first total synthesis of +-ratjadone. AB - The first total synthesis of ratjadone was achieved using a highly convergent approach joining three subunits together with a Wittig olefination and a selective Heck reaction as the pivotal steps. Besides establishing a robust and reliable route for the synthesis of this orphan ligand, the configuration of unknown stereocenters could also be determined. This synthesis not only provides an additional access to a biologically important compound but also enables the synthesis of structural analogues for biological target identification. PMID- 11262143 TI - Counteranion effect on complexation of quats by a neutral calix[5]arene receptor. PMID- 11262145 TI - A highly chemo- and stereoselective synthesis of beta-keto esters via a polymer supported lipase catalyzed transesterfication. PMID- 11262144 TI - An improved synthesis of chiral alpha-(4-bromobenzyl)alanine ethyl ester and its application to the synthesis of LFA-1 antagonist BIRT-377. PMID- 11262146 TI - A highly selective synthesis of diarylethynes and their oligomers by a palladium catalyzed Sonogashira coupling reaction under phase transfer conditions. PMID- 11262147 TI - Improved synthesis of indolyl fulgides. PMID- 11262148 TI - A novel approach to the synthesis of chiral terminal 1,2-diamines. PMID- 11262149 TI - Nad(p)(+)-nad(p)h models. 90. stereoselection controlled by electronic effect of a carbonyl group in oxidation of nad(p)h analog. PMID- 11262150 TI - Analytical characterization of the conformational transitions of polynucleotides by means of different molecular spectroscopies and multivariate curve resolution. AB - A general procedure for the study of conformational transitions of polynucleotides is described. The equilibria between different conformations induced by salt, ethidium bromide, and temperature of poly(dG-dC). poly(dG-dC) and induced by salt and temperature of poly(A). poly(U) are investigated using molecular absorption, circular dichroism, and fluorescence spectroscopies. Spectral data obtained from experiments are analyzed by means of a factor analysis method, namely, multivariate curve resolution, which allows possible intermediate states to be detected and the pure spectra and the concentration profiles of all species present in the system to be estimated. This work shows the application of this procedure for the analysis of data matrices obtained in individual experiments but also for the analysis of several data matrices simultaneously. PMID- 11262151 TI - A high-performance liquid chromatography method for the detection of diaminopimelic acid in urine. AB - We describe a quantitative assay for diaminopimelic acid (DAP) in urine. It involves (i) hydrolysis of urine samples, (ii) purification by several different liquid chromatography steps, and (iii) analysis by high-performance liquid chromatography on a reversed-phase C18 column. Tritiated-DAP, the internal standard, allows one to precisely follow the DAP-containing fractions and to determine the yield during purification. Sensitive and relatively accurate quantification of DAP, with a threshold of 50 fmol, is based on ion-pairing properties of eluants and ortho-phthaldialdehyde derivatization. The presence of DAP in relevant fractions was confirmed by combined gas chromatography and mass spectrometry. The DAP concentration in adult human urine pooled over 24 h ranges from 0.69 to 2.01 microM, a result in fair agreement with previously published values obtained by ninhydrin derivatization or gas chromatography. PMID- 11262152 TI - Investigation of the metabolite variation in control rat urine using (1)H NMR spectroscopy. AB - An exploratory statistical analysis has been undertaken of 640 (1)H NMR spectra of rat urine, obtained from predose and control animals during the course of eight separate toxicology studies. The aim was to determine the degree and type of variation between (1)H NMR spectra from such control animals and to investigate the variations in the spectral descriptors based on averaged peak intensities. The results showed that many of the spectral descriptors had skew and/or multimodal distributions, and that it was possible to distinguish between samples of urine collected at different times of day with a success rate of (89%) and to classify 90% of the predose spectra into their correct study group using principal component and linear discriminant analyses. The results show that successful classification can be achieved of NMR spectra of control rat urine, which exhibit more subtle changes than those previously reported when treated and control animals were compared. The results presented here suggest that it will be possible to identify very subtle toxicological changes if care is taken to standardize the experimental conditions used during toxicity screens. PMID- 11262153 TI - Incorporation of fluorescent enzyme substrates in agarose gel for in situ zymography. AB - The currently available methods for the detection of proteases in tissue sections are characterized by limited substrate specificity and low sensitivity and are also cumbersome. We have developed a novel in situ zymography method that uses a synthetic substrate conjugated to a fluorescent tag for detection of proteases in tissue sections. In the presence of active enzyme, the fluorescent tag is cleaved off from the substrate peptide chain resulting in an approximately 100-fold increase in the fluorescent signal. In order to minimize the diffusion of the fluorescent tag, the substrate is incorporated into 1% agarose prior to overlaying onto the tissue section. This method retains the morphological details of the tissue section, is highly sensitive and specific for the designated peptide sequence, and provides information regarding the functional status of the enzyme. Thus, this method could be used for detection and monitoring of enzymatic activity in tissue sections for a variety of applications. PMID- 11262154 TI - A kinetic study of analyte-receptor binding and dissociation, and dissociation alone, for biosensor applications: a fractal analysis. AB - A fractal analysis is presented for (a) analyte-receptor binding and dissociation kinetics and (b) dissociation kinetics alone for biosensor applications. Emphasis is placed on dissociation kinetics. Data taken from the literature may be modeled, in the case of binding, using a single-fractal analysis or a dual fractal analysis. The dual-fractal analysis represents a change in the binding mechanism as the reaction progresses on the surface. A single-fractal analysis is adequate to model the dissociation kinetics in the examples presented. Predictive relationships developed for the dissociation rate coefficient(s) as a function of the analyte concentration are of particular value since they provide a means by which the dissociation rate coefficients may be manipulated. Relationships are also presented for the binding and dissociation rate coefficients as a function of their corresponding fractal dimension, D(f), or the degree of heterogeneity that exists on the surface. When analyte-receptor binding or dissociation is involved, an increase in the heterogeneity on the surface (increase in D(f)) leads to an increase in the binding and in the dissociation rate coefficient. PMID- 11262155 TI - Mass spectrometry as a novel approach to probe cooperativity in multimeric enzymatic systems. AB - Investigating cooperativity in multimeric enzymes is of utmost interest to improve our understanding of the mechanism of enzymatic regulation. In the present article, we propose a novel approach based on mass spectrometry to probe cooperativity in the binding of a ligand to a multisubunit enzyme. This approach presents the selective advantage of giving a direct insight into all the subsequent ligation states that are formed in solution as the ligand is added to the enzyme. A quantitative interpretation of the electrospray ionization (ESI) mass spectra gives the relative abundance of all the distinct enzymatic species, which allows one to directly deduce the cooperativity of the system. The overall method is described for the addition of the oxidized cofactor nicotinamide adenine dinucleotide (NAD(+)) to a dimeric mutant of Bacillus stearothermophilus glyceraldehyde-3-phosphate dehydrogenase (GPDH). It is then applied to four tetrameric enzymes: sturgeon muscle GPDH, wild type and S48G mutant of GPDH from B. stearothermophilus, and alcohol dehydrogenase (ADH) from Bakers yeast. The results illustrate the possibilities offered by this new technique. First, mass spectrometry allows a control of the enzymes before the addition of NAD(+). Second, the cooperative behavior can be drawn from one single ESI mass spectrum, which makes the method highly attractive in terms of the amount of biological material required. Above all, the major benefit lies in the direct visualization of all the enzymatic species that are in equilibrium in solution. The direct measurement of cooperativity readily resolve the inconvenience of the classical approaches employed in this field, which all need to model the experimental data in order to get the cooperative behavior of the system. PMID- 11262156 TI - Determination of protection from serum nuclease activity by DNA-polyelectrolyte complexes using an electrophoretic method. AB - Polyelectrolyte complexes between cationic polymers and DNA have emerged as potential nonviral vectors for DNA delivery. For successful in vivo delivery, methods for analyzing their ability to prevent digestion of the DNA payload by serum nucleases are essential. We report here a simple assay to determine degradation of DNA in these complexes using standard electrophoretic techniques. The assay is based on a high pH buffer which can dissociate the complexes under standard electrophoretic conditions. This assay can be used qualitatively to determine the time taken for degradation to occur. Alternatively, with a standard gel analysis program it can be used quantitatively to investigate rates of DNA degradation from complexes in the presence of serum nucleases. We have shown that it can distinguish between different formulations with the same polymer, and also to distinguish between the time taken to degradation and the rates of degradation of DNA in complexes formed with two structurally related, linear polyamidoamine polymers. The assay could also distinguish between the time to degradation using poly-l-lysine complexes, although these were less well dissociated by the electrophoresis buffer, and could not be analyzed quantitatively. This assay will be of value in investigating and developing polyelectrolyte formulations for parenteral administration. PMID- 11262157 TI - The solubility of calcium oxalate in tissue culture media. AB - The equilibrium parameters for calcium oxalate solubility in tissue culture media were investigated because of the current interest in oxalate toxicity. The calcium selective ion electrode methodology was evaluated and calcium concentrations from potentiometric calculations were verified by d-c argon plasma emission spectroscopy. The experimental K(sp)'s at 25 degrees C for Dulbecco's modified Eagle media and McCoys 5A media are equivalent to the literature K(sp) of 2.3 x 10(-9) for low ionic strength. The equilibrium concentration products, [Ca2+] [C2O2-(4)], are ten times higher than the K(sp)'s due to the high ionic strengths of tissue culture media. At 37 degrees C, addition of soluble oxalate at the 10(-3) to 10(-4) M level causes >50% precipitation of the oxalate resulting in equilibrium oxalate concentrations of less than 6 x 10(-5) M. This relatively inexpensive selective ion technique allows the determination of oxalate concentrations in equilibrium-saturated media which are substantially less than those calculated by the amount of soluble oxalate added to the media. PMID- 11262159 TI - Assays for angiotensin converting enzyme inhibitory activity. AB - A colorimetric method and a capillary electrophoresis procedure were developed for quantifying histidyl-leucine and hippurate, respectively. The colorimetric method is sensitive (extinction coefficient = 7.5 mM(-1) cm(-1)) and reproducible (CV = 1.7%, n = 5), which is based on a selective chromogenic reaction for histidyl-leucine (lambda(max) = 390 nm) using o-phthalaldehyde. For samples containing unusually high levels of histidine and/or histidyl peptides, the separation-based approach is preferable. The capillary electrophoresis method makes use of an in-capillary microextraction technique; complicated samples can be measured in less than 4 min without pretreatment. Protocols using both methods to measure angiotensin converting enzyme inhibitory activity were proposed. PMID- 11262158 TI - Adenosine triphosphate-dependent degradation of a fluorescent lambda N substrate mimic by Lon protease. AB - Escherichia coli Lon exhibits a varying degree of energy requirement toward hydrolysis of different substrates. Efficient degradation of protein substrates requires the binding and hydrolysis of ATP such that the intrinsic ATPase of Lon is enhanced during protein degradation. Degradation of synthetic tetrapeptides, by contrast, is achieved solely by ATP binding with concomitant inhibition of the ATPase activity. In this study, a synthetic peptide (FRETN 89-98), containing residues 89-98 of lambda N protein and a fluorescence donor (anthranilamide) and quencher (3-nitrotyrosine), has been examined for ATP-dependent degradation by E. coli and human Lon proteases. The cleavage profile of FRETN 89-98 by E. coli Lon resembles that of lambda N degradation. Both the peptide and protein substrates are specifically cleaved between Cys93 and Ser94 with concomitant stimulation of Lon's ATPase activity. Furthermore, the degradation of FRETN 89-98 is supported by ATP and AMPPNP but not ATPgammaS nor AMPPCP. FRETN 89-98 hydrolysis is eight times more efficient in the presence of 0.5 mM ATP compared to 0.5 mM AMPPNP at 86 microM peptide. The ATP-dependent hydrolysis of FRETN 89-98 displays sigmodial kinetics. The k(cat), [S](0.5), and the Hill coefficient of FRETN 89-98 degradation are 3.2 +/- 0.3 s(-1), 106 +/- 21 microM, and 1.6 respectively. PMID- 11262160 TI - Optical determination of glutamine using a genetically engineered protein. AB - We have developed a reagentless optical biosensor for glutamine based on the Escherichia coli glutamine binding protein (GlnBP). Site-directed mutagenesis was performed to engineer single cysteine mutants which were covalently modified with environmentally sensitive sulfhydryl-reactive probes. The fluorescence emission of acrylodan and 2-(4'-(iodoacetamido)anilino)naphthalene-6-sulfonic acid (IAANS) attached to GlnBP mutant S179C was shown to decrease 65 and 35%, respectively, upon titration with increasing amounts of glutamine (0 to 6.4 microM; K(Dapp) 160 nM). No significant changes in the fluorescence intensity were observed for the structurally similar amino acids glutamate, asparagine, and arginine. Time resolved intensity decays showed a 2.4-fold decrease in mean lifetime for GlnBP S179C-acrylodan upon the addition of glutamine, indicating the possibility of a lifetime-based assay. Anisotropy decay measurements for GlnBPS179C-acrylodan showed a 13-ns rotational correlation time in the ligand-free state, whereas multiple correlation times were assigned in the glutamine-bound conformation. The decrease in fluorescence intensity of S179C-acrylodan was adapted to polarization sensing of glutamine. The engineered GlnBP is a first step toward the development of a nonenzymatic biosensor capable of determining glutamine concentrations in cell cultures. PMID- 11262161 TI - Application of the primer in situ DNA synthesis (PRINS) technique to titer recombinant virus and evaluation of the efficiency of viral transduction. AB - Titration is an important and critical step in dosing recombinant virus for gene therapy. We present a relatively fast, convenient, and sensitive method that allows for precise quantification of recombinant retrovirus. The method is based on PCR amplification of a foreign gene by the PRINS (primer in situ DNA synthesis) technique. The PRINS technique is based on the sequence-specific annealing of unlabeled oligonucleotide DNA in situ. This oligonucleotide operates as a primer for in situ chain elongation catalyzed by the Taq I polymerase. Using digoxygenin-labeled nucleotides as a substrate for chain elongation, the neo synthetic DNA is labeled by an FITC-conjugated anti-digoxygenin antibody. To avoid the possibility of false positives, we amplified the puromycin-resistance gene, which is associated with the transgene in the same viral vector and is not normally present in mammalian cells. The retroviral titer was evaluated by counting fluorescein isothiocyanate-positive cells after PRINS labeling, while knowing the number of plated cells that were transduced with different amounts of viral supernatant. A comparable viral concentration of 1 x 10(7) infectious units/mL was found among the retroviruses. PMID- 11262162 TI - Prediction of affinity and kinetics in biomolecular interactions by affinity chromatography. AB - Computer simulation of affinity chromatography is a valuable tool for accurate prediction of column performance. In our study affinity pairs based on lectin and antibody interactions with carbohydrates have been used as model systems. In this well-characterized system we have demonstrated the usefulness of the simulation approach for determination of affinity and kinetics. These properties are typically difficult to obtain for many weakly interacting molecular species (i.e., when dissociation constants (K(D)) are greater than 10(-5) M). The influence of affinity and kinetics on peak broadening in affinity chromatography has also been investigated. PMID- 11262163 TI - Fluorescent BODIPY-GTP analogs: real-time measurement of nucleotide binding to G proteins. AB - Three BODIPY GTPgammaS analogs (FL, 515, and TR), BODIPY FL GppNHp and BODIPY FL GTP molecules were synthesized as possible fluorescent probes to study guanine nucleotide binding spectroscopically. Binding to G(alphao) increases baseline analog fluorescence by 6-, 8.5-, 2.8-, 3.5-, and 3.0-fold, respectively. Binding of GTPgammaS and GppNHp analogs to G(alphao) is of high affinity (K(D) 11, 17, 55, and 110 nM, respectively) and reaches a stable plateau while fluorescence of BODIPY FL GTP shows a transient increase which returns to baseline. Furthermore, BODIPY FL GTPgammaS shows varying affinities for alpha(o), alpha(s), alpha(i1), and alpha(i2) (6, 58, 150, and 300 nM). The affinities of BODIPY FL GppNHp for all four G(alpha) subunits are 10-fold lower than for BODIPY FL GTPgammaS. Half times for the fluorescence increase are consistent with known GDP release rates for those proteins. Enhancement of fluorescence upon binding the G(alpha) subunit is most likely due to a rotation around the gamma-thiol (GTPgammaS) or the 3' ribose-hydroxyl (GppNHp) bond to relieve the quenching of BODIPY fluorescence by the guanine base. Binding to G(alpha) exposes the BODIPY moiety to the external environment, as seen by an increase in sodium iodide quenching. The visible excitation and emission spectra and high fluorescence levels of these probes permit robust real-time detection of nucleotide binding. PMID- 11262165 TI - Polymerase chain reaction in polymeric microchips: DNA amplification in less than 240 seconds. AB - There is much interest in developing methods amenable to amplifying nucleic acids by the polymerase chain reaction (PCR) in small volumes in microfabricated devices. The use of infrared-mediated temperature control to accurately thermocycle microliter volumes in microchips fabricated from polyimide is demonstrated. Amplification of a 500-base-pair fragment of lambda-phage DNA was achieved in a 1.7-microl chamber containing a thermocouple that allowed for accurate control of temperature. While previous work showed that Taq polymerase was inactivated when in direct contact with the thermocouple, this was circumvented with the polyimide chip by the addition of polyethylene glycol as a buffer additive. This, consequently, allowed for adequate amounts of PCR product to be observed after only 15 cycles, with a total time for amplification of 240 s. PMID- 11262164 TI - Measurement of hemoglobin synthesis rate in vivo using a stable isotope method. AB - We developed a method to measure hemoglobin synthesis rate (SynHb) in humans, assuming that free glycine in the red blood cell (RBC) represents free glycine in bone marrow for hemoglobin synthesis. The present rat study examines this assumption of the method and quantifies SynHb in rats. Sprague-Dawley rats (n = 9) were studied, [2-(13)C]glycine was intravenously infused over 24 h (2.5 mg kg( 1) h(-1)), blood was drawn for glycine and heme isolation, and bone marrow was harvested for glycine isolation. Isotopic enrichments of glycine and heme were measured, fractional hemoglobin synthesis rate (fSynHb% day(-1)) was calculated, and from this a value for SynHb (mg g(-1) day(-1)) was derived. Mean body weight was 446 +/- 10 g (mean +/- SE) and hemoglobin concentration was 14 +/- 0.5 g dl( 1). At 24 h, the mean isotopic enrichment, atom percentage excess (APE), of the RBC free glycine (1.56 +/- 0.18 APE) was similar to the bone marrow (1.68 +/- 0.15 APE). The rate of incorporation of (13)C into heme increased over time from 0.0004 APE/h between 6 and 12 h, to 0.0014 APE/h between 12 and 18 h, and 0.0024 APE/h between 18 and 24 h. Consequently, fSynHb (1.19 +/- 0.32, 2.92 +/- 0.66, and 4.22 +/- 0.56% day(-1), respectively) and SynHb (0.11 +/- 0.03, 0.28 +/- 0.05, and 0.42 +/- 0.05 mg g(-1) day(-1), respectively) showed similar patterns over the 24-h study period. We conclude that (1) enrichment of free glycine in the circulating RBC approximates enrichment of bone marrow free glycine for heme formation and (2) this pattern of hemoglobin synthesis rate is reflecting the characteristic release and gradual maturation of reticulocytes in the circulation. PMID- 11262166 TI - Detection of multivalent interactions through two-tiered energy transfer. AB - A method based on two-tiered fluorescence resonant energy transfer (FRET) has been developed for selective and sensitive detection of species involved in a multivalent interaction. Pentavalent binding between cholera toxin and ganglioside GM1 is used as a model system to demonstrate the advantage of the two tiered FRET over one-stage FRET in both conventional fluorimeter and flow cytometer. In the system, three fluorescent probes (namely, fluorescence donor, acceptor, and intermediate) are covalently tagged to receptors, and the intermediate is used to bridge the energy transfer between the donor and acceptor even though the donor's fluorescence spectrum does not overlap with absorption spectrum of the acceptor. One of the most significant improvements of the scheme over one-stage FRET is a dramatic decrease in the background fluorescence of the acceptor fluorescence, which, theoretically and practically, increases the detection sensitivity. PMID- 11262167 TI - Development of two bacterial artificial chromosome shuttle vectors for a recombination-based cloning and regulated expression of large genes in mammalian cells. AB - Most conditional expression vectors designed for mammalian cells have been valuable systems for studying genes of interest by regulating their expressions. The available vectors, however, are reliable for the short-length cDNA clones and not optimal for relatively long fragments of genomic DNA or long cDNAs. Here, we report the construction of two bacterial artificial chromosome (BAC) vectors, capable of harboring large inserts and shuttling among Escherichia coli, yeast, and mammalian cells. These two vectors, pEYMT and pEYMI, contain conditional expression systems which are designed to be regulated by tetracycline and mouse interferons, respectively. To test the properties of the vectors, we cloned in both vectors the green fluorescence protein (GFP) through an in vitro ligation reaction and the 17.8-kb-long X-inactive-specific transcript (Xist) cDNA through homologous recombination in yeast. Subsequently, we characterized their regulated expression properties using real-time quantitative RT-PCR (TaqMan) and RNA fluorescent in situ hybridization (FISH). We demonstrate that these two BAC vectors are good systems for recombination-based cloning and regulated expression of large genes in mammalian cells. PMID- 11262168 TI - Differentiation of bacterial autolysins by zymogram analysis. AB - The use of zymograms in which the bacterial cell wall heteropolymer peptidoglycan is incorporated into the resolving gel of SDS-PAGE has led to the identification of various SDS stable peptidoglycan hydrolases (autolysins). To examine the specificity of autolysins with respect to O-acetylated peptidoglycan, a discontinuous SDS-PAGE system has been developed that operates under neutral conditions. [Bis(2-hydroxyethyl)imino]tris(hydroxymethyl)methane (Bis-Tris) buffers are employed with pH 6.8 and 6.3 for the separating and stacking gels, respectively, while the anode buffer N-2-acetamido-2-hydroxyethanesulfonic acid (Aces)-HCl and the Bis-Tris cathode buffer both had a pH of 6.8. These conditions resulted in a relative trailing ion mobility of 0.349 and 0.137 in the resolving and staking gel, respectively, under room temperature conditions. Peptides and proteins were resolved in the 3-100 kDa range with a 10% acrylamide resolving gel. Comparison of zymograms that incorporated unacetylated or chemically O acetylated peptidoglycan revealed the specificity of hen egg-white lysozyme for the unacetylated material. A preliminary analysis of the autolysins produced by the urinary tract pathogen Proteus mirabilis indicated that some enzymes were specific for either O-acetylated or non-O-acetylated peptidoglycan while others displayed no clear preference toward either of the two substrates. PMID- 11262169 TI - Detection of insulin receptor tyrosine kinase activity using time-resolved fluorescence energy transfer technology. PMID- 11262170 TI - Relationship between turbidity of lipid vesicle suspensions and particle size. PMID- 11262171 TI - Concurrent measurement of promoter activity and transfection efficiency using a new reporter vector containing both Photinus pyralis and Renilla reniformis luciferase genes. PMID- 11262172 TI - Immunoprecipitation of human telomerase reverse transcriptase with telomerase activity. PMID- 11262176 TI - Role of ERK activation in growth and erythroid differentiation of K562 cells. AB - Inhibition of signaling through Ras in BCR-ABL-positive pluripotent K562 cells leads to apoptosis and spontaneous differentiation. However, Ras-induced activation of the mitogen-activated protein kinase ERK has been suggested to play a critical role in either growth or differentiation in different model systems. We studied the role of ERK activation in the growth-promoting and anti-apoptotic effect of Ras and its involvement in hemin-induced nonterminal erythroid differentiation using the BCR-ABL-positive K562 cell line as a model. K562 cells were stably transfected with ERK1 or the dominant inhibitory mutant of ERK1 (ERK1 KR). Overexpression of ERK1-KR inhibited cell growth with an approximately fourfold increase in doubling time and induced apoptosis in K562 cells. Incubation with the MEK1 inhibitor UO126 inhibited cell growth and induced apoptosis in K562 cells in a dose-dependent manner as well. In the presence of exogenously added hemin, K562 cells differentiate into erythroblasts, as indicated by the production of large amounts of fetal hemoglobin. We examined the activation of MAP kinases during hemin-induced differentiation. The ERK1 and 2 activity increased within 2 h post hemin treatment and remained elevated for 24 48 h. During this time, fetal hemoglobin synthesis also increases from 0.8 to 10 pg/cell. There was no activation of JNK or p38 protein kinases. The hemin-induced accumulation of hemoglobin was inhibited in ERK1-KR overexpressing cells and was enhanced in the wild-type ERK1 transfectants. Our results suggest that ERK activation is involved in both growth and hemin-induced erythroid differentiation in the BCR-ABL-positive K562 cell line. PMID- 11262177 TI - The nucleoskeleton: a permanent structure of cell nuclei regardless of their transcriptional activity. AB - Nuclear matrix or nucleoskeleton is thought to provide structural basis for intranuclear order. However, the nature of this structure is still uncertain because of numerous technical difficulties in its visualization. To reveal the "real" morphology of the nucleoskeleton, and to identify possible sources of structural artifacts, three methods of nucleoskeleton preparations were compared. The nucleoskeleton visualized by all these techniques consists of identical elements: nuclear lamina and an inner network comprising core filaments and the "diffuse" nucleoskeleton. We then tested if the nucleoskeleton is a stable structure or a transient transcription-dependent structure. Incubation with transcription inhibitors (alpha-amanitin, actinomycin D, and DRB) for various periods of time had no obvious effect on the morphology of the nucleoskeleton. A typical nucleoskeleton structure was observed also in a physiological model-in transcriptionally inactive mouse 2-cell embryos and in active 8- to 16-cell embryos. Our data suggest that the nucleoskeleton is a permanent structure of the cell nucleus regardless of the nuclear transcriptional state, and the principal architecture of the nucleoskeleton is identical throughout the interphase. PMID- 11262178 TI - Trypanosoma cruzi: phosphatidylinositol 3-kinase and protein kinase B activation is associated with parasite invasion. AB - Multiple signal transduction events are triggered in the host cell during invasion by the protozoan parasite Trypanosoma cruzi. Here, we report the regulation of host cell phosphatydilinositol 3-kinase (PI3K) and protein kinase B (PKB/Akt) activities by T. cruzi during parasite-host cell interaction. Treatment of nonphagocytic cells (Vero, L(6)E(9), and NIH 3T3) and phagocytic cells (human and J774 murine macrophages) with the selective PI3K inhibitors Wortmannin and LY294002 significantly impaired parasite invasion in a dose-dependent fashion. A strong activation of PI3K and PKB/Akt activities in Vero cells was detected when these cells were incubated with trypomastigotes or their isolated membranes. Consistently, we were unable to detect activation of PI3K or PKB/Akt activities in host cells during epimastigote (noninfective) membrane-host cell interaction. Infection of transiently transfected cells containing an inactive mutant PKB showed a significant inhibition of invasion compared with the active mutant transfected cells. T. cruzi PI3K-like activity was also required in host cell invasion since treatment of trypomastigotes with PI3K inhibitors prior to infection reduced parasite entry. Taken together, these results indicate that PI3K and PKB/Akt activation in parasites, as in host cells induced by T. cruzi, is an early invasion signal required for successful trypomastigote internalization. PMID- 11262179 TI - Dynamic nuclear localization of the baculovirus proteins IE2 and PE38 during the infection cycle: the promyelocytic leukemia protein colocalizes with IE2. AB - The early gene products IE2 and PE38 of Autographa californica multicapsid nuclear polyhedrosis virus localize to distinct nuclear domains after transient expression. Here, the nuclear localization pattern and the putative association with cellular proteins have been determined during virus infection to shed light on the functional significance of the nuclear domains. IE2 was always localized to distinct nuclear structures while PE38 was partly present in nuclear dots. Confocal imaging indicated colocalization of PE38 and IE2 to common domains, prominently at 2 h p.i. The nuclear dot localization of PE38 in infected cells was different from that in transfected cells. Hence, we have performed cotransfection experiments that suggested that a viral factor influences the nuclear distribution. Since the promyelocytic leukemia protein (PML) that localizes to distinct nuclear multiprotein complexes termed ND10/PODs in mammalian cells functions as a target for some immediate early viral proteins, we have investigated whether baculovirus proteins act similarly. Transiently expressed IE2 and PE38 were found to be associated with endogenous PML in the mammalian cell line BHK21. Infection with a recombinant virus that expresses the human pml gene in insect cells reveals IE2 and PML to be colocalized during the early phase of infection followed by a redistribution of both proteins. Taken together our results provide first evidence that the early baculovirus protein IE2 associates at least with one component of mammalian PODs during virus infection, suggesting that POD-like structures can be formed in insect cells. PMID- 11262180 TI - The role of B-cell-specific activator protein in the response of malignant B-1 cells to LPS. AB - Chronic lymphocytic leukemia (CLL) results from the uncontrolled proliferation and accumulation of B-1 cells, many of which demonstrate self-reactivity. The response of B-1 cells to mitogen after undergoing malignant transformation is still unclear. Using our established malignant B-1 cell lines derived from the NZB murine model of human CLL, we investigated the response of malignant B-1 cells to the mitogen LPS. Interestingly, these malignant B-1 cells proliferated initially, but the proliferation rate decreased after a 48-h transition. Prolonged LPS treatment induced apoptosis and pathological differentiation. We studied possible underlying molecular mechanisms and found that the level of the DNA binding protein BSAP (B-cell-specific activator protein) was upregulated by LPS at the initial activation stage, followed by an increase in the apoptotic factor caspase-3 (CPP32) at 48 h and a subsequent decrease of BSAP at 72 h. The pathological differentiation induced by LPS was partially prevented by treatment with antisense BSAP. This study indicates that malignant B-1 cells could be driven to apoptosis and pathological differentiation when activated by the mitogen LPS, and BSAP may be an important factor in regulating these responses. PMID- 11262181 TI - Rho GTPases are involved in the regulation of NF-kappaB by genotoxic stress. AB - A common cellular response to genotoxic agents and inflammatory cytokines is the activation of NF-kappaB. Here, we addressed the question of whether small GTPases of the Rho family are involved in the stimulation of NF-kappaB signaling by genotoxic agents or TNFalpha in HeLa cells. Inhibition of isoprenylation of Rho proteins by use of the HMG-CoA reductase inhibitor lovastatin attenuated UV-, doxorubicin-, and TNFalpha-induced degradation of IkappaBalpha as well as drug stimulated DNA binding activity of NF-kappaB. Furthermore, NF-kappaB-regulated gene expression stimulated by either UV irradiation or treatment with TNFalpha was abrogated by lovastatin pretreatment. This indicates that isoprenylated regulatory proteins participate in the regulation of NF-kappaB by DNA-damaging agents as well as by TNFalpha. Specific blockage of Rho signaling by Clostridium difficile toxin B attenuated UV- and doxorubicin-induced activation of NF-kappaB, but did not affect stimulation of NF-kappaB by TNFalpha. Obviously, signaling to NF-kappaB by genotoxic and nongenotoxic stimuli occurs via different molecular mechanisms, either involving Rho GTPases or not. Based on the data, we suggest Rho GTPases to be essentially required for genotoxic stress-induced signaling to NF-kappaB. PMID- 11262182 TI - Type I protein kinase a is localized to interphase microtubules and strongly associated with the mitotic spindle. AB - We show here that type I protein kinase A is localized to microtubules during the entire cell cycle in epithelial (hepatoma, cervical carcinoma) and nonepithelial (myoblast) cell lines. The association of the type Ialpha regulatory subunit is very strong in all phases of mitosis, from prophase to cytokinesis. In interphase, the association appears weaker, reflecting perhaps a more dynamic molecular interaction. This regulatory subunit appears to recruit catalytic subunits as the latter are also associated with microtubules. BW1J hepatoma cells, stably transfected with either wild-type or mutant Ialpha regulatory subunit, are enriched in aberrant mitoses with multipolar spindles and in mono- or multinucleated giant cells. This suggests that type I protein kinase A could have a role in centrosome duplication and/or segregation, sister chromatid separation, or cytokinesis. PMID- 11262183 TI - A Cdk5-p35 stable complex is involved in the beta-amyloid-induced deregulation of Cdk5 activity in hippocampal neurons. AB - The cdk5 and its activator p35 constitute one of the main tau-phosphorylating systems in neuronal cells. Under normal conditions for neurons, its activity is required for modulating tau involvement in neuronal polarity and in development of the mammalian central nervous system. Recently, we reported that the treatment of rat hippocampal cells in culture with fibrillary beta-amyloid (Abeta) results in deregulation of the protein kinase cdk5. The neurotoxic effects of Abeta fibrils were prevented by inhibition of cdk5 activity by butyrolactone I or by using antisense oligonucleotides that control the expression of this kinase. Here, we show that the Abeta-promoted increase of cdk5 activity is associated with changes in tau phosphorylation patterns and in the intraneuronal distribution of tau. In addition to hippocampal cells, deregulation of cdk5 was observed in other cell types. However, butyrolactone I prevented Abeta-induced cell death only in neuronal cells in which cdk5 activation was sensitive to Abeta fibrils. This lost of cdk5 regulation in hippocampal cells exposed to Abeta fibrils appears to be associated with an increase in the cdk5-p35 complex stability. Complex stabilization was sensitive to phosphorylation of cdk5. However, no changes in cdk5 and p35 mRNAs were observed, suggesting that the main effects on cdk5 occur at the posttranslational level. These studies indicate that cdk5 phosphorylation and the formation of an abnormally active cdk5-p35 complex are directly involved in the molecular paths leading to the neurodegenerative process of rat hippocampal neurons triggered by Abeta fibrils. PMID- 11262184 TI - The role of mitogen-activated protein kinase activation within focal adhesions in chemotaxis toward FGF-2 by murine brain capillary endothelial cells. AB - Fibroblast growth factors (FGFs) regulate a number of angiogenic cellular responses such as migration of endothelial cells. To examine the role of mitogen activated protein kinase (MAPK) in endothelial cell migration, chemotaxis toward FGF-2 was determined in murine brain capillary endothelial cells, denoted IBE cells. PD98059, a specific inhibitor for MAPK/Erk kinase, inhibited FGF-2-induced chemotaxis of IBE cells. It has been reported that c-Src tyrosine kinase phosphorylates focal adhesion kinase at tyrosine 925 within focal adhesions, which in turn creates the binding site for Grb2, leading to MAPK activation. The Src family tyrosine kinase inhibitor, PP1, as well as overexpression of kinase inactive c-Src, attenuated chemotaxis toward FGF-2. To investigate the signaling events involved in FGF-2-induced chemotaxis, MAPK activation was monitored in IBE cells by indirect immunofluorescence staining. Activated MAPK was initially observed in the cytoplasm and gradually moved into nuclei. A fraction of MAPK was activated by FGF-2 within focal adhesions, where FGF receptor-1 and Src family kinases were also colocalized. MAPK activation within focal adhesions was remarkably decreased in kinase-inactive c-Src-expressing IBE cells. Our data suggest that activation of MAPK by FGF-2 within focal adhesions may depend on c Src activity and is crucial for FGF-2-induced migration of IBE cells. PMID- 11262185 TI - Molecular characterization of Ct-hrp65: identification of two novel isoforms originated by alternative splicing. AB - Hrp65, a protein with two conserved RNA-binding domains, has been identified in Chironomus tentans as a component of nuclear fibers associated with ribonucleoprotein particles in transit from the gene to the nuclear pore. We have cloned two novel hrp65 isoforms and characterized the structure of the hrp65 gene. Comparison of the hrp65 gene to the hrp65 cDNAs revealed that the multiple hrp65 isoforms, hrp65-1, hrp65-2 and hrp65-3, are generated by alternative splicing of a single pre-mRNA. The hrp65-3 mRNA is only detected in C. tentans tissue culture cells of embryonic origin, whereas hrp65-1 and hrp65-2 mRNAs appear to be constitutively expressed. The hrp65 mRNAs are generated by differential 3' splice site selection at the last exon of the gene. Thus, the three hrp65 transcripts contain different 3' UTRs and encode proteins that vary in their C-terminal ends. Interestingly, the variant C-terminal region determines the subcellular localization of the hrp65 proteins. In transient transfection assays, hrp65-1 is efficiently targetted to the nucleus, whereas hrp65-2 and hrp65-3 localize mainly to the cytoplasm. Moreover, hrp65-3 is associated with cytoplasmic actin fibers. All together, our findings suggest that the different hrp65 isoforms serve specialized roles related to mRNA localization/transport in the different cell compartments. PMID- 11262186 TI - PKC-dependent activation of FAK and src induces tyrosine phosphorylation of Cas and formation of Cas-Crk complexes. AB - SH-SY5Y neuroblastoma cells are a well-characterized model for studying the induction of neuronal differentiation. TPA treatment of these cells induces cytoskeletal rearrangements that ultimately result in neurite extension. However, the signaling pathways that precede these changes are poorly understood. Other investigators have shown that TPA treatment of SH-SY5Y cells results in increased tyrosine phosphorylation of cytoskeletal-associated proteins, including the adapter protein Cas. In this report, we examine the events upstream and downstream of Cas phosphorylation. We show that TPA treatment induces the PKC dependent association of tyrosine-phosphorylated Cas with Crk. The activity of two protein tyrosine kinases, Src and FAK, was shown to be necessary and sufficient for TPA-induced Cas phosphorylation. We propose that the PKC-dependent phosphorylation of Cas by Src and FAK promotes the establishment of Cas-Crk complexes and that these interactions may play an important role in regulating the actin cytoskeleton during neuronal differentiation. PMID- 11262187 TI - Testosterone-induced growth of S115 mouse mammary tumor cells is dependent on heparan sulfate. AB - The androgen-induced proliferation of S115 mouse mammary tumor cells has been suggested to involve autocrinic fibroblast growth factor signaling. Heparan sulfate proteoglycans are required for fibroblast growth factor signaling, presumably due to their ability to alter binding of fibroblast growth factors to their receptors. We have investigated the role of heparan sulfate proteoglycans in the testosterone-induced proliferation of S115 cells. We demonstrate that when the cells are treated with sodium chlorate, which inhibits the sulfation of endogenous heparan sulfate proteoglycans, cell growth becomes dependent on exogenous heparin. The shortest heparin oligosaccharides supporting cell growth were octasaccharides, whereas dodecasaccharides were almost as effective as native heparin. The N-, 2-O-, and 6-O-sulfate groups of heparin were all required for full testosterone response. Treatment of S115 cells with chlorate or testosterone did not alter the expression of fibroblast growth factor receptors 1 or 3, whereas the expression of fibroblast growth factor receptor 2 was down regulated. We have previously shown that overexpression of syndecan-1 heparan sulfate proteoglycan renders S115 cells insensitive to testosterone and now demonstrate that this effect can be overcome by sodium chlorate treatment in combination with exogenous heparin. Our results suggest that heparin-like molecules are intimately involved in the androgen-mediated proliferation of S115 cells. PMID- 11262188 TI - Comprehensive studies on subcellular localizations and cell death-inducing activities of eight GFP-tagged apoptosis-related caspases. AB - By using green fluorescent protein fusion, we investigated the subcellular localization of all the caspases that have been cloned from humans and implicated in the execution of apoptosis. We divided these caspases into three groups according to subcellular localization. The first group includes caspase-1, -3, 6, -7, and -9, which are expressed mainly in the cytoplasm with various levels of nuclear localization depending on the cell type. The second group has a single member, caspase-2, which is primarily localized in the nucleus. The nuclear localization was demonstrated to be mediated by a nuclear localization signal near the NH(2)-terminus of the prodomain. The third group includes caspase-8 and 10, which have a cytoplasmic distribution. These two members have potent, rapid cell death-inducing activity and are prone to make aggregates when overexpressed. Their prodomains formed marked fibrous structures in the cytoplasm whose localization seemed distinct from organelles or cytoskeletons. None of the GFP caspases examined in this study showed a predominant mitochondrial localization as has been reported for some caspases. PMID- 11262189 TI - uPA/plasmin system-mediated MMP-9 activation is implicated in bronchial epithelial cell migration. AB - To examine the effects of the uPA/plasmin system on cell migration in relation to the activation of MMP-9, we used ex vivo and in vitro wound-repair models of human bronchial epithelial cells and videomicroscopy techniques that make possible cell tracking and quantification of cell migration speeds. We observed that uPA was only detected in migrating cells at the wound edges and located at crucial sites for cell/extracellular matrix interactions. The implication of uPA in human bronchial epithelial cell migration was studied by incubating cultures with a monoclonal antibody raised against uPA and these experiments led to a 70% reduction in cell velocity. To examine the effects of the plasmin system on cell migration, we incubated cultures with increasing concentrations of plasmin or activated MMP-9. We observed a significant dose-dependent increase in cell migration velocity with plasmin (P < 0.001) and MMP-9 (P < 0.001). Moreover, addition of exogenous plasmin led to a twofold increase of activated MMP-9 in migrating cells. We also demonstrated that the addition of anti-uPA IgG led to an inhibition of 43% of activated MMP-9. In conclusion, these results show that uPA is involved in human bronchial epithelial cells migration. This action is mediated by the generation of plasmin, which in turn activates MMP-9, thus making possible cell migration. PMID- 11262190 TI - Trefoil peptides promote restitution of wounded corneal epithelial cells. AB - The ocular surface shares many characteristics with mucosal surfaces. In both, healing is regulated by peptide growth factors, cytokines, and extracellular matrix proteins. However, these factors are not sufficient to ensure most rapid healing. Trefoil peptides are abundantly expressed epithelial cell products which exert protective effects and are key regulators of gastrointestinal epithelial restitution, the critical early phase of cell migration after mucosal injury. To assess the role of trefoil peptides in corneal epithelial wound healing, the effects of intestinal trefoil factor (ITF/TFF3) and spasmolytic polypeptide (SP/TFF2) on migration and proliferation of corneal epithelial cells were analyzed. Both ITF and SP enhanced restitution of primary rabbit corneal epithelial cells in vitro. While the restitution-enhancing effects of TGF-alpha and TGF-beta were both inhibited by neutralizing anti-TGF-beta-antibodies, trefoil peptide stimulation of restitution was not. Neither trefoil peptide significantly affected proliferation of primary corneal epithelial cells. ITF but not SP or pS2 mRNA was present in rabbit corneal and conjunctival tissues. In summary, the data indicate an unanticipated role of trefoil peptides in healing of ocular surface and demand rating their functional actions beyond the gastrointestinal tract. PMID- 11262191 TI - Respiratory chain-generated oxidative stress following treatment of leukemic blasts with DNA-damaging agents. AB - Oxidative stress occurs in diverse life forms during programmed cell death and appears to be a significant mediator since a wide range of manipulations that enhance cellular antioxidant systems are protective. Using a recently developed flow cytometry technique to assess respiratory chain function, we have investigated the mechanism of reactive oxygen generation in OCI/AML-2 leukemic blasts following treatment with cytosine arabinoside, etoposide, and gamma irradiation. Increases in mitochondrially generated reactive oxygen were seen using all three agents, in association with hyperpolarization of the mitochondrial inner membrane. Increased reactive oxygen occurred when mitochondria were energized using substrates for either complex I or complex II, indicating that the likely source is complex III (cytochrome c reductase). These findings are consistent with impaired adenine nucleotide exchange across the mitochondrial membrane, recently proposed to be an important event during the early stages of apoptosis induction (M. G. Vander Heiden et al., 1999, Mol. Cell 3, 159-167). Elevations of the antioxidants glutathione and thioredoxin occurred in association with this oxidative stress, likely the result of feedback mechanisms based on redox-sensitive transcription factors. Since glutathione and thioredoxin can protect from drug-induced apoptosis, their upregulation in response to respiratory chain-generated reactive oxygen might represent a cellular adaptation to DNA damage that promotes cell survival. PMID- 11262192 TI - EGF regulates a complex pattern of gene expression and represses smooth muscle differentiation during the neurotypic conversion of the neural-crest-derived TC 1S cell line. AB - EGF, known to sustain CNS neuronal progenitors, also promotes a neurotypic response in the thymic neural-crest-derived TC-1S cell line. We report here the use of TC-1S cells as a model to identify the genetic programs regulated during the neurotypic response induced by EGF and to isolate 23 EGF-responsive genes. Among them 5 represent novel cDNAs, while 18 are known genes, whose regulation by EGF is associated with the mitogenic or differentiating effects of the growth factor. The repression of smooth muscle alpha-actin and SM22alpha genes by EGF and their increase by TGFbeta suggest that the TC-1S line includes neural crest multipotent cells whose smooth muscle differentiation is repressed upon EGF treatment and stimulated by TGFbeta. Therefore, we identified a complex pattern of EGF-target genes and propose EGF as a novel signal able to recruit postmigratory neural-crest-derived cells along proliferation and cell lineage choice pathways. PMID- 11262193 TI - Subcellular localization of procollagen I and prolyl 4-hydroxylase in corneal endothelial cells. AB - To investigate the molecular mechanism of intracellular degradation of type I collagen in normal corneal endothelial cells (CEC), we studied the role of prolyl 4-hydroxylase (P4-H) and protein disulfide-isomerase (PDI; the beta subunit of P4 H) during procollagen I biosynthesis. When the subcellular localization of P4-H and PDI was determined, P4-H demonstrated a characteristic diffuse endoplasmic reticulum (ER) pattern, whereas PDI showed a slightly more restricted distribution within the ER. When colocalization of procollagen I with the enzymes was examined, procollagen I and PDI showed a large degree of colocalization. P4-H and procollagen I were predominantly colocalized at the perinuclear site. When colocalization of type IV collagen with PDI and P4-H was examined, type IV collagen was largely colocalized with PDI, which showed a wider distribution than type IV collagen. Type IV collagen is similarly colocalized with P4-H, except in some perinuclear sites. The colocalization profiles of procollagen I with both PDI and P4-H were not altered in cells treated with alpha,alpha'-dipyridyl compared to those of the untreated cells. The underhydroxylated type IV collagen demonstrated a colocalization profile with PDI similar to that observed with procollagen I, while the underhydroxylated type IV collagen was predominantly colocalized with P4-H at the perinuclear sites. Immunoblot analysis showed no real differences in the amounts of the beta subunit/PDI and the catalytic alpha subunit of P4-H in CEC compared to those of corneal stromal fibroblasts (CSF). When protein-protein association was determined, procollagen I was associated with PDI much more in CEC than it was in CSF, whereas type IV collagen showed no differential association specificity to PDI in both cells. Limited proteolysis of the newly synthesized intracellular procollagen I with pepsin showed that procollagen I in CEC was degraded by pepsin, whereas CSF contained type I collagen composed of alpha1(I) and alpha2(I). These findings suggest that procollagen I synthesized in CEC is not in triple helical conformation and that the improperly folded procollagen I may be preferentially associated with PDI before targeting to the intracellular degradation. PMID- 11262194 TI - The Saccharomyces cerevisiae phosphotyrosyl phosphatase activator proteins are required for a subset of the functions disrupted by protein phosphatase 2A mutations. AB - In Saccharomyces cerevisiae, PTPA is encoded by two genes, YPA1 and YPA2. In order to examine the biological role of PTPA as potential regulator of protein phosphatase 2A (PP2A), we compared the phenotypes of the ypaDelta mutants with these of PP2A-deficient strains. While deletion of both YPA genes is lethal, deletion of YPA1 alone results in a phenotype resembling that of PP2A-deficient strains in specific aspects such as aberrant bud morphology, abnormal actin distribution, and similar growth defects under various growth conditions. These phenotypes were even more pronounced when YPA1 was deleted in a pph21Delta genetic background. Moreover, ypaDelta mutants are hypersensitive to nocodazole and show inappropriate mitotic spindle formation as previously described for mutants in the catalytic subunit of PP2A, suggesting that Ypa, like PP2A, has a function in mitotic spindle formation. These results are consistent with an in vivo role of Ypa as a regulator of PP2A. However, unlike a PP2A-deficient strain, ypaDelta mutants do not show a G2 arrest. Therefore, Ypa does not seem to play a role in the regulation of PP2A at this stage of the cell cycle. These results imply that Ypa regulates a specific subset of PP2A functions, possibly by controlling the subunit composition of PP2A. PMID- 11262195 TI - Loss of insulin-like growth factor II receptor expression promotes growth in cancer by increasing intracellular signaling from both IGF-I and insulin receptors. AB - The insulin-like growth factor-II receptor (IGF-IIR) is frequently mutated or deleted in some malignant human tumors, suggesting that the IGF-IIR is a tumor suppressor. However, the exact mechanism by which IGF-IIR suppresses growth in tumors has not been definitively established. We demonstrate that IGF-IIR deficient murine L cells (D9) have higher growth rates than IGF-IIR-positive L cells (Cc2) in response to IGF-II. IGF-II levels are higher in growth-conditioned medium from D9 versus Cc2 cells. Receptor neutralization studies and measurements of insulin receptor substrate 1 phosphorylation confirm that the enhanced growth of D9 cells is due to increased stimulation of the IGF-I and insulin receptors by IGF-II. In contrast, the levels of secreted latent and active transforming growth factor beta (TGF-beta) are similar for both D9 and Cc2 cells, indicating that the slower growth of Cc2 cells is not due to activation of latent TGF-beta by IGF-IIR and growth inhibition. The results directly demonstrate that down regulation of the IGF-IIR promotes the growth of transformed D9 cells by sustaining IGF-II, which binds to and activates IGF-IR and insulin receptor to increase intracellular growth signals. PMID- 11262196 TI - Enhancement of connexin 43 expression increases proliferation and differentiation of an osteoblast-like cell line. AB - Bone cells form a functional syncytium as they are coupled by gap junctions composed mainly of connexin 43 (Cx43). To further understand the role of Cx43 in bone cell growth and differentiation, we stably transfected Cx45-expressing UMR 106-01 cells with Cx43 using an expression vector containing rat Cx43 cDNA. Three stably transfected clones were analyzed, all of which showed altered expression of Cx43 and/or Cx45 as was obvious from immunocytochemistry and Northern blotting. Double whole-cell patch clamping revealed single-channel conductances of 20 (Cx45) and 60 pS (Cx43). The overexpression of Cx43 led to an increase in dye coupling concomitant with elevated gap-junctional conductance. The phenotype of the transfected clones was characterized by an increased proliferation (4- to 7-fold) compared to controls. Moreover, a transfectant clone with 10- to 12-fold enhanced Cx43 expression showed a significantly increased calcium content of the extracellular matrix and enlarged mineralization nodules, while alkaline phosphatase was moderately increased. We conclude that enhanced gap-junctional coupling via Cx43 significantly promotes proliferation and differentiation of UMR cells. PMID- 11262197 TI - Transcriptional activation of prion protein gene in growth-arrested and differentiated mouse erythroleukemia and human neoplastic cells. AB - The prion protein (PrP) is a GPI-anchored sialoglycoprotein that has attracted worldwide attention over the years due to its involvement in the pathogenesis of transmissible spongiform encephalopathies in sheep (scrapie), cattle (BSE), and humans (CJD). To understand the precise role of the Prn-p gene in cell growth and differentiation we investigated the expression pattern of the Prn-p gene in proliferating cells and in cells arrested in growth either by confluency or by induction of terminal differentiation. Viral-transformed mouse spleen hematopoietic cells named murine erythroleukemia (MEL) and other types of inducible cells (human neuroectodermal RD/TE-671, myoid RD cells) were employed. Cells grown exponentially, at confluency, or irreversibly arrested in growth at terminal differentiation state were analyzed by fluorescence cell sorting and Northern blot hybridization to estimate the steady-state level of PrP mRNA at different phases of the cell cycle. MEL cells that failed to differentiate from treatment with N(6)-methyladenosine (N(6)mAdo), an inhibitor of differentiation, were also analyzed for PrP mRNA level. Our results indicate the following: (a) growth arrest of cells at G(1) phase by confluency or by induction of terminal differentiation led to increased accumulation of PrP mRNA transcripts, an event observed also in differentiated MEL, RD/TE-671, and RD cells independent of the inducer used; (b) treatment of MEL cells with N(6)mAdo prevented early activation of the Prn-p gene in cells treated with the inducer; and (c) cell-free nuclear run-off studies showed enhanced expression of the Prn-p gene due to transcriptional activation. These findings indicate, for the first time, that the Prn-p gene, which is thought to be a housekeeping gene, is transcriptionally activated in G(1) phase in confluent and terminally differentiated cells. This information may be valuable in understanding the overaccumulation of PrP in some differentiated tissues and may let us repress Prn-p gene activation by novel agents. PMID- 11262198 TI - A canarypox vector-expressing cytomegalovirus (CMV) phosphoprotein 65 induces long-lasting cytotoxic T cell responses in human CMV-seronegative subjects. AB - The major matrix phosphoprotein 65 (pp65) of cytomegalovirus (CMV) is an important target of HLA-restricted cytotoxic T cells (CTL) after natural infection. A canarypox-CMV pp65 recombinant was studied for its ability to induce CMV pp65-specific CTL, helper T lymphocytes, and antibodies in a phase I clinical trial. Twenty-one CMV-seronegative adult volunteers were randomized to receive immunizations at months 0, 1, 3, and 6 with either canarypox-CMV pp65 or placebo. In canarypox-CMV pp65-immunized subjects, pp65-specific CTL were elicited after only 2 vaccinations and were present at months 12 and 26 in all subjects tested. Cell-depletion studies indicated that the CTL were phenotype CD8(+). Peripheral blood mononuclear cells proliferated in response to stimulation with purified pp65, and antibodies specific for pp65 also were detected. Canarypox-CMV pp65 is the first recombinant vaccine to elicit CMV-specific CTL responses, which suggests the potential usefulness of this approach in preventing disease caused by CMV. PMID- 11262199 TI - Expression of Th2 cytokines decreases the development of and improves Behcet's disease-like symptoms induced by herpes simplex virus in mice. AB - In the etiology of Behcet's disease (BD), viral infection has long been postulated as a contributing factor, and viral involvement has been demonstrated. However, viral infection alone is not sufficient to explain the pathogenesis of BD, and some evidence suggests that immunologic abnormalities are also important. To study the possible role of immune regulation in the development of BD-like symptoms induced by herpes simplex virus inoculation in ICR mice, macrophages were deleted by use of liposome-encapsulated clodronate (lip-Cl(2)MDP). Treatment with lip-Cl(2)MDP suppressed the development of BD-like symptoms, and this suppression was correlated with the induction of interleukin-4 expression in mouse spleens. When the Th2 adjuvant ovalbumin (OVA)-alum was injected into mice with BD-like symptoms, their cutaneous symptoms improved. Adoptive transfer with splenocytes from OVA-alum-injected mice also resulted in improvement. These findings suggest that up-regulated Th2 cytokine expression can attenuate the development of and improve some BD-like symptoms. PMID- 11262200 TI - Hepatitis C virus-specific CD4+ T cell response after liver transplantation occurs early, is multispecific, compartmentalizes to the liver, and does not correlate with recurrent disease. AB - The role of hepatitis C virus (HCV)-specific CD4+ T cells in recurrent HCV infection after orthotopic liver transplantation (OLTx) is unclear. In parallel, 73 intrahepatic and 73 blood-derived T cell lines were established from 34 patients. At a single cell level, virus-specific interferon (IFN)-gamma production to various HCV proteins was determined by ELISPOT assay: 45 (62%) of 73 liver- or blood-derived T cell lines produced IFN-gamma in response to one of the HCV antigens. HCV specificity was detected mainly in the liver (47% vs. 23% in the blood; P<.05, chi(2) test) and was detectable earlier (< or =6 months) significantly more often than later (>6 months) after OLTx (78% vs 49%; P<.05, chi(2) test). Histology, histologic activity index, liver enzymes, and virus load did not correlate with the occurrence of HCV-specific CD4+ T cells. Despite strong immunosuppressive treatment, OLTx recipients can develop an early, multispecific, preferentially intrahepatic CD4+ T cell response that decreases over time, making it a potential candidate target for novel therapeutic approaches in the transplant setting. PMID- 11262202 TI - Cervical and prostate primary epithelial cells are not productively infected but sequester human immunodeficiency virus type 1. AB - Primary prostate and cervical epithelial cells and epithelial cell lines were examined for human immunodeficiency virus type 1 (HIV-1) infection or transmission to peripheral blood mononuclear cells (PBMC). Neither cell-free nor cell-associated HIV-1 infected primary epithelial cells or cell lines. Pretreatment of HIV-1 to enhance CD4-independent entry did not augment infection. Cell surface expression was detected for galactosyl ceramide but not for CC chemokine receptor 5, CXC-chemokine receptor 4, or CD4. The ability to transfer HIV-1 to resting or activated PBMC was tested by culturing with rinsed or trypsinized and replated HIV-1-exposed epithelial cells. Virus was not recovered from the rinsed or replated cocultures with resting PBMC; however, activated PBMC recovered HIV-1 from rinsed epithelial cells and rarely from replated epithelial cells. Although urogenital epithelial cells are not infected, these data suggest that they can transfer virus to activated immune cells and have implications for sexual transmission of HIV-1. PMID- 11262201 TI - Number and position of mutations in the interferon (IFN) sensitivity-determining region of the gene for nonstructural protein 5A correlate with IFN efficacy in hepatitis C virus genotype 1b infection. AB - To explore the relationship between responses to interferon (IFN) and the mutation patterns in the IFN sensitivity-determining region (ISDR; amino acid positions 2209-2248) in the NS5A gene of hepatitis C virus genotype 1b, a cohort of 334 patients was analyzed. The number of mutations in the ISDR was higher in patients with sustained response (SR) than in patients with transient or no response (P<.001). Patients with viruses mutated at positions 2209 (P=.02), 2216 (P=.01), or 2227 (P=.02) more frequently experienced SR than did those without these mutations. Mutation occurred most frequently at position 2218, where the presence of cysteine was significantly associated with SR. Thus, the mutation pattern in the ISDR affects the virologic response to IFN and reflects different influences on the function of the NS5A protein. ISDR sequence analysis would allow the prediction of clinical IFN efficacy in individual patients. PMID- 11262203 TI - Two distinct receptors mediate nonopsonic phagocytosis of different strains of Pseudomonas aeruginosa. AB - Complement receptor 3 (CR3) mediates both opsonic and nonopsonic phagocytosis of bacteria. Leukocyte adhesion deficiency (LAD) allows for the study of CR3 dependent phagocyte-bacterial ingestion, since LAD phagocytes do not express this receptor. Phagocytes from an infant with LAD were unable to ingest 50% of the Pseudomonas aeruginosa strains studied, which indicates a requirement for CR3. However, the remaining strains were phagocytosed in the absence of CR3, and ingestion was blocked by monoclonal antibodies directed at CD14. This CR3/CD14 receptor bias was further confirmed by using thioglycollate-elicited murine peritoneal macrophages, which have nonfunctional CR3 before activation. Results indicate that either CR3 or CD14 is involved independently in nonopsonic phagocytosis of different P. aeruginosa strains. Clearance of P. aeruginosa from the endobronchial space may be facilitated by nonopsonic phagocytosis, since low levels of opsonins are present. The impact of lung infection with P. aeruginosa may be determined, in part, by the phagocytic receptor that mediates ingestion. PMID- 11262204 TI - Borrelia burgdorferi stimulates in vitro a selective expansion of CD3-CD4-CD8- (triple negative) autoreactive cells in naive rhesus monkey lymphocytes. AB - Autoreactive cell lines were generated from cell suspensions of freshly isolated naive monkey lymph node (LN) cells and peripheral blood mononuclear cells by cocultivation with freeze-thawed Borrelia burgdorferi spirochetes (Bb/FT). These cells produced interleukin (IL)-6 when cocultured with autologous antigen presenting cells (APCs) alone without Bb/FT. IL-6 production was not observed when control cell lines were stimulated in the same fashion. CD4+-enriched T cell populations obtained from the LN autoreactive cell line also produced IL-6 when cultivated with APCs alone. When these cells were cultivated further in the presence of APCs, a population of cells whose phenotype was CD56+/-CD4-CD8-CD3- was predominantly selected. These cells both proliferated and produced IL-6 when cocultured with APCs alone. The possible relevance of these cells to Lyme disease pathogenesis remains to be determined. PMID- 11262205 TI - Localization of Tropheryma whippelii rRNA in tissues from patients with Whipple's disease. AB - Whipple's disease is caused by a cultivation-resistant bacterium, Tropheryma whippelii. Ultrastructural studies of intestinal biopsy specimens from patients with Whipple's disease have shown that intracellular and extracellular bacteria are present, but the preferred site of growth is unknown. Tissue sections from 8 patients with Whipple's disease and from 19 healthy control subjects were analyzed by use of fluorescence in situ hybridization and laser scanning confocal microscopy, to determine the location of rRNA that would indicate the presence of metabolically active bacteria. T. whippelii rRNA was most prevalent near the tips of intestinal villi, in the lamina propria, just basal to epithelial cells. Most of the bacterial rRNA signal appeared to be located between cells and did not colocalize with the human intracellular protein vimentin. The location of bacterial rRNA in tissues from patients with Whipple's disease provides evidence that bacteria are growing outside cells and suggests that T. whippelii is not an obligate intracellular pathogen. PMID- 11262206 TI - Antibodies to glycans dominate the host response to schistosome larvae and eggs: is their role protective or subversive? AB - Multiple exposures of chimpanzees to the radiation-attenuated schistosome vaccine provoked a strong parasite-specific cellular and humoral immune response. Specific IgM and IgG were directed mainly against glycans on antigens released by cercariae; these were also cross-reactive with soluble antigens from larvae, adult worms, and eggs. Egg deposition was the major antigenic stimulus after challenge of vaccinated and control chimpanzees with normal parasites, eliciting strong antiglycan responses to egg secretions. Glycan epitopes recognized included LacdiNAc, fucosylated LacdiNAc, Lewis(X) (weakly), and those on keyhole limpet hemocyanin. Antibodies to peptide epitopes became prominent only during the chronic phase of infection, as glycan-specific IgM and IgG decreased. Because of their intensity and cross-reactivity, the antiglycan responses resulting from infection could be a smoke screen to subvert the immune system away from more vulnerable larval peptide epitopes. Their occurrence in humans might explain the long time required for antischistosome immunity to build up after infection. PMID- 11262207 TI - Effect of testing for IgG avidity in the diagnosis of Toxoplasma gondii infection in pregnant women: experience in a US reference laboratory. AB - The usefulness of testing for IgG avidity in association with Toxoplasma gondii was evaluated in a US reference laboratory. European investigators have reported that high-avidity IgG toxoplasma antibodies exclude acute infection in the preceding 3 months. In this US study, 125 serum samples taken from 125 pregnant women in the first trimester were chosen retrospectively, because either the IgM or differential agglutination (AC/HS) test in the Toxoplasma serologic profile suggested or was equivocal for a recently acquired infection. Of 93 (74.4%) serum samples with either positive or equivocal results in the IgM ELISA, 52 (55.9%) had high-avidity antibodies, which suggests that the infection probably was acquired before gestation. Of 87 (69.6%) serum samples with an acute or equivocal result in the AC/HS test, 35 (40.2%) had high-avidity antibodies. Forty women were given spiramycin, to prevent congenital transmission, and 7 (17.5%) had high avidity antibodies. These findings highlight the value of testing a single serum sample obtained in the first trimester of pregnancy for IgG avidity. PMID- 11262208 TI - Artesunate reduces but does not prevent posttreatment transmission of Plasmodium falciparum to Anopheles gambiae. AB - Combination therapy that includes artemisinin derivatives cures most falciparum malaria infections. Lowering transmission by reducing gametocyte infectivity would be an additional benefit. To examine the effect of such therapy on transmission, Gambian children with Plasmodium falciparum malaria were treated with standard regimens of chloroquine or pyrimethamine-sulfadoxine alone or in combination with 1 or 3 doses of artesunate. The infectivity to mosquitoes of gametocytes in peripheral blood was determined 4 or 7 days after treatment. Infection of mosquitoes was observed in all treatment groups and was positively associated with gametocyte density. The probability of transmission was lowest in those who received pyrimethamine-sulfadoxine and 3 doses of artesunate, and it was 8-fold higher in the group that received pyrimethamine-sulfadoxine alone. Artesunate reduced posttreatment infectivity dramatically but did not abolish it completely. The study raises questions about any policy to use pyrimethamine sulfadoxine alone as the first-line treatment for malaria. PMID- 11262209 TI - Malaria infection induces virus expression in human immunodeficiency virus transgenic mice by CD4 T cell-dependent immune activation. AB - To test the capacity of malaria parasites to trigger virus expression in vivo, human immunodeficiency virus (HIV) transgenic mice were infected with Plasmodium chabaudi chabaudi clone AS. Splenocytes recovered during peak parasitemia showed a dramatic elevation in viral p24 production that returned to baseline by day 15 and failed to rebound at recrudescence or after reinfection. The major sources of virus expression were antigen-presenting cells (APCs) rather than T lymphocytes. Nevertheless, T cells from infected mice stimulated with plasmodial antigen triggered 5-10-fold increases in p24 production from dendritic cells in vitro, which suggests that viral induction stems from interaction of malaria-specific T lymphocytes with HIV-expressing APCs. Indeed, depletion of CD4 T cells resulted in a 70% reduction in the p24 response stimulated by malaria in vivo. These findings demonstrate the ability of Plasmodium species to immunologically activate latently integrated HIV in vivo but suggest that this process may be restricted to acute infection. PMID- 11262210 TI - Detection by reverse transcription-polymerase chain reaction of influenza C in nasopharyngeal secretions of adults with a common cold. AB - The lack of practical methods for a laboratory diagnosis of influenza C virus infections and the seemingly benign nature of the virus contribute to the fact that 50 years after its first isolation, relatively little is known about the epidemiology and the clinical impact of this virus. Reverse transcription polymerase chain reaction (RT-PCR) was used to amplify influenza C RNA fragments from clinical specimens. Two hundred otherwise healthy adults with recent onset of a common cold were studied. Nasopharyngeal aspirates were collected at entry to the study and 1 week later. Serum samples for antibody determinations were obtained at the first visit and after 3 weeks. Influenza C was detected in 7 of the 200 patients by 2 different RT-PCR formats. All 7 patients had a significant increase in antibody titers between serum samples collected during the acute and convalescent phases of the illness. Influenza C appears to be one of the many viruses that cause acute upper respiratory tract infections in adults. PMID- 11262211 TI - An outbreak of hepatitis A associated with green onions. AB - Forty-three cases of serologically confirmed hepatitis A occurred among individuals who ate at restaurant A in Ohio in 1998. Serum samples from all restaurant A employees who worked during the exposure period were negative for IgM antibodies to hepatitis A virus (HAV). A matched case-control study determined that foods containing green onions, which were eaten by 38 (95%) of 40 case patients compared with 30 (50%) of 60 control subjects, were associated with illness (matched odds ratio, 12.7; 95% confidence interval, 2.6-60.8). Genetic sequences of viral isolates from 14 case patients were identical to each other and to those of viral isolates from 3 patients with cases of hepatitis A acquired in Mexico. Although the implicated green onions, which could have come from one of 2 Mexican farms or from a Californian farm, were widely distributed, no additional green onion-associated cases were detected. More sensitive methods are needed to detect foodborne hepatitis A. A better understanding of how HAV might contaminate raw produce would aid in developing prevention strategies. PMID- 11262212 TI - Quantitative hepatitis B virus DNA testing for the early prediction of the maintenance of response during lamivudine therapy in patients with chronic hepatitis B. AB - To determine whether a dramatic decrease in hepatitis B virus (HBV) DNA levels within the first months of lamivudine therapy can predict the emergence of YMDD variants in patients with chronic hepatitis B, quantitative testing was done every 3 months on serum samples from 35 patients who were treated with lamivudine for >1 year. The decline in HBV DNA levels from baseline to month 3 was higher in 22 responders than in 13 nonresponders (mean+/-SD, 4.16+/-1.06 vs. 2.88+/-1.77 log(10) copies; P=.002), whereas no differences were observed in patients with and without YMDD variants at 1 year of therapy. At 3 months, HBV DNA was undetectable in 77% of the responders, whereas, after 1 year, it was undetectable in 23% of nonresponders, 40% of patients with YMDD variants, and 74% of those without variants. Therefore, quantitative HBV DNA testing is very useful in deciding whether to continue therapy, because of the low likelihood of response in patients who remain HBV DNA positive at month 3 of treatment. PMID- 11262213 TI - Half-life of human parainfluenza virus type 3 (hPIV3) maternal antibody and cumulative proportion of hPIV3 infection in young infants. AB - During a phase 2 trial of parainfluenza virus type 3 (PIV3) vaccine, sequential serum samples were obtained from infants at 2, 6, 7, 12-15, and 13-16 months of age. Paired serum samples obtained at 2 and 6 months of age were used to estimate the biologic half-life of human PIV3 (hPIV3) maternal antibody in young infants. On the basis of the assumption that hPIV3 maternal antibody decays exponentially and constantly, the biologic half-life was estimated without adjusting for body weight increases. Cumulative proportions of hPIV3 infection in young infants were further estimated after adjusting for maternal antibody decline. A hemagglutination inhibition assay was used to quantify hPIV3 antibody. The mean (95% confidence interval) biologic half-life was estimated to be 51 (42-60) days, on the basis of which cumulative proportions of hPIV3 infection were estimated to be 11% at 6 months of age, 47% at 12-15 months of age, and 50% at 13-16 months of age. PMID- 11262215 TI - Influence of age on CD4 cell recovery in human immunodeficiency virus-infected patients receiving highly active antiretroviral therapy: evidence from the EuroSIDA study. AB - Influence of age on the CD4 cell response to highly active antiretroviral therapy (HAART) was examined in 1956 patients (median age, 37.2 years) in the EuroSIDA study. Median initial CD4 cell count was 192x106 cells/L, follow-up was 31 months, and time to maximum CD4 cell response was 20 months. Age groups were not different for baseline CD4 cell count, baseline human immunodeficiency virus RNA load, or treatment history. CD4 cell increase, stratified by age quartiles, differed during months 3-36 of HAART (P=.023). Maximum CD4 cell increase from start of HAART differed by age group (P=.0003), as did maximum CD4 cell count (P<10-4). Multivariate analysis confirmed the inverse relationship between age and maximum CD4 cell response (P=.023). Time to a CD4 increase of >200x106 cells/L was shorter for patients in the younger age groups (P=.0026), as confirmed by multivariate analysis (P<10-4). Younger age may favor CD4 cell restoration because of preserved thymic function. PMID- 11262214 TI - Loss of cytomegalovirus-specific CD4+ T cell responses in human immunodeficiency virus type 1-infected patients with high CD4+ T cell counts and recurrent retinitis. AB - Clinical histories are reported for 2 patients treated with highly active antiretroviral therapy (HAART) who experienced multiple relapses of cytomegalovirus (CMV) retinitis, despite suppression of human immunodeficiency virus type 1 (HIV-1) viremia and improvement in CD4+ T cell counts (to >400 cells/microL). CMV-specific CD4+ T cell immune reconstitution was measured directly, using cytokine flow cytometry, which revealed persistent deficits in CMV-specific CD4+ T cell responses in both patients. CMV-specific T cells constituted 0.14% and 0.05% of the total CD4+ T cell count in these patients, which is significantly lower than the percentages for 34 control subjects (0.6% 46%; CD4+ T cell count range, 7-1039 cells/microL; P=.019). Deficits in pathogen specific immune responses may persist in some individuals, despite suppression of HIV-1 replication and substantial increases in circulating CD4+ T cells after HAART, and such deficits may be associated with significant morbidity from opportunistic infections. PMID- 11262216 TI - Serotype distribution of Salmonella isolates from food animals after slaughter differs from that of isolates found in humans. AB - If raw meat and poultry are the primary point of entry for Salmonella species into human populations, a correlation might be expected between the serotype distribution of Salmonella species isolated from animals at the time of slaughter and that of isolates found in humans. For 1990-1996, sufficient national data were available to permit such a comparison. A mathematical model was developed to predict serotype distributions of Salmonella isolates among humans on the basis of animal data. There was a significant mismatch between the serotype distributions among humans predicted by the model and those actually observed. This mismatch raises questions about the validity of the "standard" assumptions about Salmonella transmission on which the model was based-namely, that raw animal products are the primary source for human salmonellosis, that the risk of transmission to humans is equal for all food product categories, and that all Salmonella serotypes have an equal ability to cause human illness. PMID- 11262217 TI - A whole blood bactericidal assay for tuberculosis. AB - The bactericidal activity of orally administered antituberculosis (anti-TB) drugs was determined in a whole blood culture model of intracellular infection in which microbial killing reflects the combined effects of drug and immune mechanisms. Rifampin (Rif) was the most active compound studied and reduced the number of viable bacilli by >4 logs. Isoniazid (INH), 2 quinolones, and pyrazinamide (PZA) showed intermediate levels of activity. Ethambutol exerted only a bacteristatic effect; amoxicillin/clavulanate was inactive. The combination of INH-Rif-PZA showed strong activity against 11 drug-sensitive isolates (mean, -3.8 log) but no activity against 12 multidrug-resistant (MDR) strains. The combination of levofloxacin-PZA-ethambutol had intermediate bactericidal activity against MDR isolates (mean, -1.2 log) but failed to equal that of INH-Rif-PZA against sensitive isolates (P<.001). The whole blood BACTEC method (Becton Dickinson) may be useful for the early clinical evaluation of new anti-TB drugs and in the management of individual patients. PMID- 11262218 TI - Effect of order of infection with human immunodeficiency virus and human herpesvirus 8 on the incidence of Kaposi's sarcoma. PMID- 11262220 TI - Viral diversity in some immunodominant epitopes: possible implications for retroviral immunopathogenesis. PMID- 11262221 TI - Misleading negative findings in a field trial of killed, oral cholera vaccine in Peru. PMID- 11262224 TI - Direct regulation of the muscle-identity gene apterous by a Hox protein in the somatic mesoderm. AB - Hox genes control segment identity in the mesoderm as well as in other tissues. Most evidence indicates that Hox genes act cell-autonomously in muscle development, although this remains a controversial issue. We show that apterous expression in the somatic mesoderm is under direct Hox control. We have identified a small enhancer element of apterous (apME680) that regulates reporter gene expression in the LT1-4 muscle progenitors. We show that the product of the Hox gene Antennapedia is present in the somatic mesoderm of the second and third thoracic segments. Through complementary alterations in the Antennapedia protein and in its binding sites on apME680, we show that Antennapedia positively regulates apterous in a direct manner, demonstrating unambiguously its cell autonomous role in muscle development. Finally, we determine that LT1-4 muscles contain more nuclei in the thorax than in the abdomen and we propose that one of the segmental differences under Hox control is the number of myoblasts allocated to the formation of specific muscles in different segments. PMID- 11262225 TI - The AP-2 transcription factor is required for joint formation and cell survival in Drosophila leg development. AB - Flies mutant for the Drosophila homologue of the mammalian transcription factor AP-2 show a severe reduction in leg length and fail to develop joint structures. Presumptive joint cells express dAP-2 in response to Notch signaling. dAP-2 is required for joint cell differentiation and can induce formation of supernumerary joints when misexpressed. Although dAP-2 is expressed only in presumptive joint cells, its activity is required to support cell survival in the entire leg segment. Taken together, our data indicate that dAP-2 is an important mediator of Notch activity in leg development. PMID- 11262226 TI - Drosophila transcription factor AP-2 in proboscis, leg and brain central complex development. AB - We report loss- and gain-of-function analyses that identify essential roles in development for Drosophila transcription factor AP-2. A mutagenesis screen yielded 16 lethal point mutant alleles of dAP-2. Null mutants die as adults or late pupae with a reduced proboscis, severely shortened legs (approximately 30% of normal length) lacking tarsal joints, and disruptions in the protocerebral central complex, a brain region critical for locomotion. Seven hypomorphic alleles constitute a phenotypic series yielding hemizygous adults with legs ranging from 40-95% of normal length. Hypomorphic alleles show additive effects with respect to leg length and viability; and several heteroallelic lines were established. Heteroallelic adults have moderately penetrant defects that include necrotic leg joints and ectopic growths (sometimes supernumerary antennae) invading medial eye territory. Several dAP-2 alleles with DNA binding domain missense mutations are null in hemizygotes but have dominant negative effects when paired with hypomorphic alleles. In wild-type leg primordia, dAP-2 is restricted to presumptive joints. Ectopic dAP-2 in leg discs can inhibit but not enhance leg elongation indicating that functions of dAP-2 in leg outgrowth are region restricted. In wing discs, ectopic dAP-2 cell autonomously transforms presumptive wing vein epithelium to ectopic sensory bristles, consistent with an instructive role in sensory organ development. These findings reveal multiple functions for dAP-2 during morphogenesis of feeding and locomotor appendages and their neural circuitry, and provide a new paradigm for understanding AP-2 family transcription factors. PMID- 11262227 TI - Inactivation of the beta-catenin gene by Wnt1-Cre-mediated deletion results in dramatic brain malformation and failure of craniofacial development. AB - beta-Catenin is a central component of both the cadherin-catenin cell adhesion complex and the Wnt signaling pathway. We have investigated the role of beta catenin during brain morphogenesis, by specifically inactivating the beta-catenin gene in the region of Wnt1 expression. To achieve this, mice with a conditional ('floxed') allele of beta-catenin with required exons flanked by loxP recombination sequences were intercrossed with transgenic mice that expressed Cre recombinase under control of Wnt1 regulatory sequences. beta-Catenin gene deletion resulted in dramatic brain malformation and failure of craniofacial development. Absence of part of the midbrain and all of the cerebellum is reminiscent of the conventional Wnt1 knockout (Wnt1(-/-)), suggesting that Wnt1 acts through beta-catenin in controlling midbrain-hindbrain development. The craniofacial phenotype, not observed in embryos that lack Wnt1, indicates a role for beta-catenin in the fate of neural crest cells. Analysis of neural tube explants shows that (beta-catenin is efficiently deleted in migrating neural crest cell precursors. This, together with an increased apoptosis in cells migrating to the cranial ganglia and in areas of prechondrogenic condensations, suggests that removal of beta-catenin affects neural crest cell survival and/or differentiation. Our results demonstrate the pivotal role of beta-catenin in morphogenetic processes during brain and craniofacial development. PMID- 11262228 TI - Reduced fertility and spermatogenesis defects in mice lacking chromosomal protein Hmgb2. AB - High mobility group 2 protein (Hmgb2) is a member of the HMGB protein family, which includes the ubiquitous Hmgb1 and the embryo-specific Hmgb3. The three proteins are more than 80% identical at the amino acid level and their biochemical properties are indistinguishable. Hmgb1 is an abundant component of all mammalian nuclei and acts as an architectural factor that bends DNA and promotes protein assembly on specific DNA targets. Cells that lack Hmgb1 can survive, although mutant mice die shortly after birth. As Hmgb2 is present in all cultured cells and is abundant in thymus, the preferred source for HMGB proteins, it was considered a ubiquitous variant of Hmgb1. We show that in adult mice Hmgb2 is restricted mainly to lymphoid organs and testes, although it is widely expressed during embryogenesis. Mice that lack Hmgb2 are viable. However, male Hmgb2(-/-) mice have reduced fertility, that correlates with Sertoli and germ cell degeneration in seminiferous tubules and immotile spermatozoa. Significantly, Hmgb2 is expressed at very high levels in primary spermatocytes, while it is barely detectable in spermatogonia and elongated spermatids. This peculiar pattern of expression and the phenotype of mutants indicate that Hmgb2 has a specialised role in germ cell differentiation. PMID- 11262229 TI - Regulation of imprinted X-chromosome inactivation in mice by Tsix. AB - In mammals, X-chromosome inactivation is imprinted in the extra-embryonic lineages with paternal X chromosome being preferentially inactivated. In this study, we investigate the role of Tsix, the antisense transcript from the Xist locus, in regulation of Xist expression and X-inactivation. We show that Tsix is transcribed from two putative promoters and its transcripts are processed. Expression of Tsix is first detected in blastocysts and is imprinted with only the maternal allele transcribed. The imprinted expression of Tsix persists in the extra-embryonic tissues after implantation, but is erased in embryonic tissues. To investigate the function of Tsix in X-inactivation, we disrupted Tsix by insertion of an IRESbetageo cassette in the second exon, which blocked transcripts from both promoters. While disruption of the paternal Tsix allele has no adverse effects on embryonic development, inheritance of a disrupted maternal allele results in ectopic Xist expression and early embryonic lethality, owing to inactivation of both X chromosomes in females and single X chromosome in males. Further, early developmental defects of female embryos with maternal transmission of Tsix mutation can be rescued by paternal inheritance of the Xist deletion. These results provide genetic evidence that Tsix plays a crucial role in maintaining Xist silencing in cis and in regulation of imprinted X-inactivation in the extra-embryonic tissues. PMID- 11262230 TI - Analysis of RNA associated with P granules in germ cells of C. elegans adults. AB - P granules are cytoplasmic structures of unknown function that are associated with germ nuclei in the C. elegans gonad, and are localized exclusively to germ cells, or germ cell precursors, throughout the life cycle. All the known protein components of P granules contain putative RNA-binding motifs, suggesting that RNA is involved in either the structure or function of the granules. However, no specific mRNAs have been identified within P granules in the gonad. We show here that P granules normally contain a low level of RNA, and describe conditions that increase this level. We present evidence that several, diverse mRNAs, including pos-1, mex-1, par-3, skn-1, nos-2 and gld-1 mRNA, are present at least transiently within P granules. In contrast, actin and tubulin mRNA and rRNA are either not present in P granules, or are present at relatively low levels. We show that pgl-1 and the glh (Vasa-related) gene family, which encode protein components of P granules, do not appear essential for RNA to concentrate in P granules; these proteins may instead function in events that are a prerequisite for RNAs to be transported efficiently from the nuclear surface. PMID- 11262231 TI - Meis3 synergizes with Pbx4 and Hoxb1b in promoting hindbrain fates in the zebrafish. AB - Many Hox proteins are thought to require Pbx and Meis co-factors to specify cell identity during embryogenesis. Here we demonstrate that Meis3 synergizes with Pbx4 and Hoxb1b in promoting hindbrain fates in the zebrafish. We find that Hoxb1b and Pbx4 act together to induce ectopic hoxb1a expression in rhombomere 2 of the hindbrain. In contrast, Hoxb1b and Pbx4 acting together with Meis3 induce hoxb1a, hoxb2, krox20 and valentino expression rostrally and cause extensive transformation of forebrain and midbrain fates to hindbrain fates, including differentiation of excess rhombomere 4-specific Mauthner neurons. This synergistic effect requires that Hoxb1b and Meis3 have intact Pbx-interaction domains, suggesting that their in vivo activity is dependent on binding to Pbx4. In the case of Meis3, binding to Pbx4 is also required for nuclear access. Our results are consistent with Hoxb1b and Meis3 interacting with Pbx4 to form complexes that regulate hindbrain development during zebrafish embryogenesis. PMID- 11262232 TI - Roles of homeobox and bHLH genes in specification of a retinal cell type. AB - Previous analysis of mutant mice has revealed that the bHLH genes Mash1 and Math3, and the homeobox gene Chx10 are essential for generation of bipolar cells, the interneurons present in the inner nuclear layer of the retina. Thus, a combination of the bHLH and homeobox genes should be important for bipolar cell genesis, but the exact functions of each gene remain largely unknown. We have found that in Mash1-Math3 double-mutant retina, which exhibits a complete loss of bipolar cells, Chx10 expression did not disappear but remained in Muller glial cells, suggesting that Chx10 expression per se is compatible with gliogenesis. In agreement with this, misexpression of Chx10 alone with retrovirus in the retinal explant cultures induced generation of the inner nuclear layer cells, including Muller glia, but few of them were mature bipolar cells. Misexpression of Mash1 or Math3 alone did not promote bipolar cell genesis either, but inhibited Muller gliogenesis. In contrast, misexpression of Mash1 or Math3 together with Chx10 increased the population of mature bipolar cells and decreased that of Muller glia. Thus, the homeobox gene provides the inner nuclear layer-specific identity while the bHLH genes regulate the neuronal versus glial fate determination, and these two classes of genes together specify the bipolar cell fate. Moreover, Mash1 and Math3 promoted the bipolar cell fate, but not the other inner nuclear layer-specific neuronal subtypes in the presence of Chx10, raising the possibility that the bHLH genes may be involved in neuronal subtype specification, in addition to simply making the neuronal versus glial fate choice. PMID- 11262233 TI - The ULTRAPETALA gene controls shoot and floral meristem size in Arabidopsis. AB - The regulation of proper shoot and floral meristem size during plant development is mediated by a complex interaction of stem cell promoting and restricting factors. The phenotypic effects of mutations in the ULTRAPETALA gene, which is required to control shoot and floral meristem cell accumulation in Arabidopsis thaliana, are described. ultrapetala flowers contain more floral organs and whorls than wild-type plants, phenotypes that correlate with an increase in floral meristem size preceding organ initiation. ultrapetala plants also produce more floral meristems than wild-type plants, correlating with an increase in inflorescence meristem size without visible fasciation. Expression analysis indicates that ULTRAPETALA controls meristem cell accumulation partly by limiting the domain of CLAVATA1 expression. Genetic studies show that ULTRAPETALA acts independently of ERA1, but has overlapping functions with PERIANTHIA and the CLAVATA signal transduction pathway in controlling shoot and floral meristem size and meristem determinacy. Thus ULTRAPETALA defines a novel locus that restricts meristem cell accumulation in Arabidopsis shoot and floral meristems. PMID- 11262234 TI - Identification of NKL, a novel Gli-Kruppel zinc-finger protein that promotes neuronal differentiation. AB - The proneural basic helix-loop-helix proteins play a crucial role in promoting the differentiation of postmitotic neurons from neural precursors. However, recent evidence from flies and frogs indicates that additional factors act together with the proneural bHLH proteins to promote neurogenesis. We have identified a novel zinc finger protein, neuronal Kruppel-like protein (NKL), that positively regulates neurogenesis in vertebrates. NKL is expressed in Xenopus primary neurons and in differentiating neuronal precursors in the intermediate zone of the mouse and chick neural tube. In frog embryos, NKL is induced by overexpression of Neurogenin (Ngn), arguing that NKL is downstream of the proneural determination genes. Our results show that NKL and a NKL/VP16 fusion protein promote differentiation of neuronal precursors in the embryonic chick spinal cord. Following in ovo misexpression of NKL, neuroepithelial cells exit the cell cycle and differentiate into neurons. Similarly, NKL/VP16 induces extra primary neurons in frogs and upregulates expression of the neural differentiation factors, Xath3 and MyT1, as well as the neuronal markers, N-tubulin and elrC. Our findings establish NKL as a novel positive regulator of neuronal differentiation and provide further evidence that non-bHLH transcription factors function in the neuronal differentiation pathway activated by the vertebrate neuronal determination genes. PMID- 11262236 TI - Regulated nuclear export of the homeodomain transcription factor Prospero. AB - Subcellular distribution of the Prospero protein is dynamically regulated during Drosophila embryonic nervous system development. Prospero is first detected in neuroblasts where it becomes cortically localized and tethered by the adapter protein, Miranda. After division, Prospero enters the nucleus of daughter ganglion mother cells where it functions as a transcription factor. We have isolated a mutation that removes the C-terminal 30 amino acids from the highly conserved 100 amino acid Prospero domain. Molecular dissection of the homeo- and Prospero domains, and expression of chimeric Prospero proteins in mammalian and insect cultured cells indicates that Prospero contains a nuclear export signal that is masked by the Prospero domain. Nuclear export of Prospero, which is sensitive to the drug leptomycin B, is mediated by Exportin. Mutation of the nuclear export signal-mask in Drosophila embryos prevents Prospero nuclear localization in ganglion mother cells. We propose that a combination of cortical tethering and regulated nuclear export controls Prospero subcellular distribution and function in all higher eukaryotes. PMID- 11262235 TI - eFGF and its mode of action in the community effect during Xenopus myogenesis. AB - The community effect is an interaction among a group of many nearby precursor cells, necessary for them to maintain tissue-specific gene expression and differentiate co-ordinately. During Xenopus myogenesis, the muscle precursor cells must be in group contact throughout gastrulation in order to develop into terminally differentiated muscle. The molecular basis of this community interaction has not to date been elucidated. We have developed an assay for testing potential community factors, in which isolated muscle precursor cells are treated with a candidate protein and cultured in dispersion. We have tested a number of candidate factors and we find that only eFGF protein is able to mediate a community effect, stimulating stable muscle-specific gene expression in demonstrably single muscle precursor cells. In contrast, Xwnt8, bFGF, BMP4 and TGF(&bgr;)2 do not show this capacity. We show that eFGF is expressed in the muscle precursor cells at the right time to mediate the community effect. Moreover, the time when the muscle precursor cells are sensitive to eFGF corresponds to the period of the endogenous community effect. Finally, we demonstrate that FGF signalling is essential for endogenous community interactions. We conclude that eFGF is likely to mediate the community effect in Xenopus myogenesis. PMID- 11262237 TI - Distinct sites of origin of oligodendrocytes and somatic motoneurons in the chick spinal cord: oligodendrocytes arise from Nkx2.2-expressing progenitors by a Shh dependent mechanism. AB - In the vertebrate spinal cord, oligodendrocytes arise from the ventral part of the neuroepithelium, a region also known to generate somatic motoneurons. The emergence of oligodendrocytes, like that of motoneurons, depends on an inductive signal mediated by Sonic hedgehog. We have defined the precise timing of oligodendrocyte progenitor specification in the cervico-brachial spinal cord of the chick embryo. We show that ventral neuroepithelial explants, isolated at various development stages, are unable to generate oligodendrocytes in culture until E5 but become able to do so in an autonomous way from E5.5. This indicates that the induction of oligodendrocyte precursors is a late event that occurs between E5 and E5.5, precisely at the time when the ventral neuroepithelium stops producing somatic motoneurons. Analysis of the spatial restriction of oligodendrocyte progenitors, evidenced by their expression of O4 or PDGFR(&agr;), indicate that they always lie within the most ventral Nkx2.2-expressing domain of the neuroepithelium, and not in the adjacent domain characterized by Pax6 expression from which somatic motoneurons emerge. We then confirm that Shh is necessary between E5 and E5.5 to specify oligodendrocyte precursors but is no longer required beyond this stage to maintain ongoing oligodendrocyte production. Furthermore, Shh is sufficient to induce oligodendrocyte formation from ventral neuroepithelial explants dissected at E5. Newly induced oligodendrocytes expressed Nkx2.2 but not Pax6, correlating with the in vivo observation. Altogether, our results show that, in the chick spinal cord, oligodendrocytes originate from Nkx2.2-expressing progenitors. PMID- 11262238 TI - Notch signaling represses the glial fate in fly PNS. AB - By using gain-of-function mutations it has been proposed that vertebrate Notch promotes the glial fate. We show in vivo that glial cells are produced at the expense of neurons in the peripheral nervous system of flies lacking Notch and that constitutively activated Notch produces the opposite phenotype. Notch acts as a genetic switch between neuronal and glial fates by negatively regulating glial cell deficient/glial cells missing, the gene required in the glial precursor to induce gliogenesis. Moreover, Notch represses neurogenesis or gliogenesis, depending on the sensory organ type. Numb, which is asymmetrically localized in the multipotent cell that produces the glial precursor, induces glial cells at the expense of neurons. Thus, a cell-autonomous mechanism inhibits Notch signaling. PMID- 11262239 TI - The role of presenilin 1 during somite segmentation. AB - The Notch signalling pathway plays essential roles during the specification of the rostral and caudal somite halves and subsequent segmentation of the paraxial mesoderm. We have re-investigated the role of presenilin 1 (Ps1; encoded by Psen1) during segmentation using newly generated alleles of the Psen1 mutation. In Psen1-deficient mice, proteolytic activation of Notch1 was significantly affected and the expression of several genes involved in the Notch signalling pathway was altered, including Delta-like3, Hes5, lunatic fringe (Lfng) and Mesp2. Thus, Ps1-dependent activation of the Notch pathway is essential for caudal half somite development. We observed defects in Notch signalling in both the caudal and rostral region of the presomitic mesoderm. In the caudal presomitic mesoderm, Ps1 was involved in maintaining the amplitude of cyclic activation of the Notch pathway, as represented by significant reduction of Lfng expression in Psen1-deficient mice. In the rostral presomitic mesoderm, rapid downregulation of the Mesp2 expression in the presumptive caudal half somite depends on Ps1 and is a prerequisite for caudal somite half specification. Chimaera analysis between Psen1-deficient and wild-type cells revealed that condensation of the wild-type cells in the caudal half somite was concordant with the formation of segment boundaries, while mutant and wild-type cells intermingled in the presomitic mesoderm. This implies that periodic activation of the Notch pathway in the presomitic mesoderm is still latent to segregate the presumptive rostral and caudal somite. A transient episode of Mesp2 expression might be needed for Notch activation by Ps1 to confer rostral or caudal properties. In summary, we propose that Ps1 is involved in the functional manifestation of the segmentation clock in the presomitic mesoderm. PMID- 11262240 TI - Nuclear import of activated D-ERK by DIM-7, an importin family member encoded by the gene moleskin. AB - The initiation of gene expression in response to Drosophila receptor tyrosine kinase signaling requires the nuclear import of the MAP kinase, D-ERK. However, the molecular details of D-ERK translocation are largely unknown. In this regard, we have identified D-Importin-7 (DIM-7), the Drosophila homolog of vertebrate importin 7, and its gene moleskin. DIM-7 exhibits a dynamic nuclear localization pattern that overlaps the spatial and temporal profile of nuclear, activated D ERK. Co-immunoprecipitation experiments show that DIM-7 associates with phosphorylated D-ERK in Drosophila S2 cells. Furthermore, moleskin mutations enhance hypomorphic and suppress hypermorphic D-ERK mutant phenotypes. Deletion or mutation of moleskin dramatically reduces the nuclear localization of activated D-ERK. Directly linking DIM-7 to its nuclear import, this defect can be rescued by the expression of wild-type DIM-7. Mutations in the Drosophila Importin beta homolog Ketel, also reduce the nuclear localization of activated D ERK. Together, these data indicate that DIM-7 and Ketel are components of the nuclear import machinery for activated D-ERK. PMID- 11262241 TI - 5-HT causes an increase in cAMP that stimulates, rather than inhibits, oocyte maturation in marine nemertean worms. AB - In the nemertean worms Cerebratulus lacteus and Micrura alaskensis, 5-HT (=5 hydroxytryptamine, or serotonin) causes prophase-arrested oocytes to mature and complete germinal vesicle breakdown (GVBD). To identify the intracellular pathway that mediates 5-HT stimulation, follicle-free oocytes of nemerteans were assessed for GVBD rates in the presence or absence of 5-HT after being treated with various modulators of cAMP, a well known transducer of 5-HT signaling and an important regulator of hormone-induced maturation in general. Unlike in many animals where high levels of intra-oocytic cAMP block maturation, treatment of follicle-free nemertean oocytes with agents that elevate cAMP (8-bromo-cAMP, forskolin or inhibitors of phosphodiesterases) triggered GVBD in the absence of added 5-HT. Similarly, 5-HT caused a substantial cAMP increase prior to GVBD in nemertean oocytes that had been pre-injected with a cAMP fluorosensor. Such a rise in cAMP seemed to involve G-protein-mediated signaling and protein kinase A (PKA) stimulation, based on the inhibition of 5-HT-induced GVBD by specific antagonists of these transduction steps. Although the downstream targets of activated PKA remain unknown, neither the synthesis of new proteins nor the activation of MAPKs (mitogen-activated protein kinases) appeared to be required for GVBD after 5-HT stimulation. Alternatively, pre-incubation in roscovitine, an inhibitor of maturation-promoting factor (MPF), prevented GVBD, indicating that maturing oocytes eventually need to elevate their MPF levels, as has been documented for other animals. Collectively, this study demonstrates for the first time that 5-HT can cause immature oocytes to undergo an increase in cAMP that stimulates, rather than inhibits, meiotic maturation. The possible relationship between such a form of oocyte maturation and that observed in other animals is discussed. PMID- 11262242 TI - The domino gene of Drosophila encodes novel members of the SWI2/SNF2 family of DNA-dependent ATPases, which contribute to the silencing of homeotic genes. AB - The Drosophila domino gene has been isolated in a screen for mutations that cause hematopoietic disorders. Generation and analysis of loss-of-function domino alleles show that the phenotypes are typical for proliferation gene mutations. Clonal analysis demonstrates that domino is necessary for cell viability and proliferation, as well as for oogenesis. domino encodes two protein isoforms of 3202 and 2498 amino acids, which contain a common N-terminal region but divergent C termini. The common region includes a 500 amino acid DNA-dependent ATPase domain of the SWI2/SNF2 family of proteins, which function via interaction with chromatin. We show that, although domino alleles do not exhibit homeotic phenotypes by themselves, domino mutations enhance Polycomb group mutations and counteract Trithorax group effects. The Domino proteins are present in large complexes in embryo extracts, and one isoform binds to a number of discrete sites on larval polytene chromosomes. Altogether, the data lead us to propose that domino acts as a repressor by interfering with chromatin structure. This activity is likely to be performed as a subunit of a chromatin-remodeling complex. PMID- 11262243 TI - Steroid regulation of autophagic programmed cell death during development. AB - Apoptosis and autophagy are morphologically distinct forms of programmed cell death. While autophagy occurs during the development of diverse organisms and has been implicated in tumorigenesis, little is known about the molecular mechanisms that regulate this type of cell death. Here we show that steroid-activated programmed cell death of Drosophila salivary glands occurs by autophagy. Expression of p35 prevents DNA fragmentation and partially inhibits changes in the cytosol and plasma membranes of dying salivary glands, suggesting that caspases are involved in autophagy. The steroid-regulated BR-C, E74A and E93 genes are required for salivary gland cell death. BR-C and E74A mutant salivary glands exhibit vacuole and plasma membrane breakdown, but E93 mutant salivary glands fail to exhibit these changes, indicating that E93 regulates early autophagic events. Expression of E93 in embryos is sufficient to induce cell death with many characteristics of apoptosis, but requires the H99 genetic interval that contains the rpr, hid and grim proapoptotic genes to induce nuclear changes diagnostic of apoptosis. In contrast, E93 expression is sufficient to induce the removal of cells by phagocytes in the absence of the H99 genes. These studies indicate that apoptosis and autophagy utilize some common regulatory mechanisms. PMID- 11262244 TI - A requirement for Notch in the genesis of a subset of glial cells in the Drosophila embryonic central nervous system which arise through asymmetric divisions. AB - In the Drosophila central nervous system (CNS) glial cells are known to be generated from glioblasts, which produce exclusively glia or neuroglioblasts that bifurcate to produce both neuronal and glial sublineages. We show that the genesis of a subset of glial cells, the subperineurial glia (SPGs), involves a new mechanism and requires Notch. We demonstrate that the SPGs share direct sibling relationships with neurones and are the products of asymmetric divisions. This mechanism of specifying glial cell fates within the CNS is novel and provides further insight into regulatory interactions leading to glial cell fate determination. Furthermore, we show that Notch signalling positively regulates glial cells missing (gcm) expression in the context of SPG development. PMID- 11262245 TI - The winged-helix transcription factor FoxD3 is important for establishing the neural crest lineage and repressing melanogenesis in avian embryos. AB - The winged-helix or forkhead class of transcription factors has been shown to play important roles in cell specification and lineage segregation. We have cloned the chicken homolog of FoxD3, a member of the winged-helix class of transcription factors, and analyzed its expression. Based on its expression in the dorsal neural tube and in all neural crest lineages except the late emigrating melanoblasts, we predicted that FoxD3 might be important in the segregation of the neural crest lineage from the neural epithelium, and for repressing melanogenesis in early-migrating neural crest cells. Misexpression of FoxD3 by electroporation in the lateral neural epithelium early in neural crest development produced an expansion of HNK1 immunoreactivity throughout the neural epithelium, although these cells did not undergo an epithelial/mesenchymal transformation. To test whether FoxD3 represses melanogenesis in early migrating neural crest cells, we knocked down expression in cultured neural crest with antisense oligonucleotides and in vivo by treatment with morpholino antisense oligonucleotides. Both experimental approaches resulted in an expansion of the melanoblast lineage, probably at the expense of neuronal and glial lineages. Conversely, persistent expression of FoxD3 in late-migrating neural crest cells using RCAS viruses resulted in the failure of melanoblasts to develop. We suggest that FoxD3 plays two important roles in neural crest development. First, it is involved in the segregation of the neural crest lineage from the neuroepithelium. Second, it represses melanogenesis, thereby allowing other neural crest derivatives to differentiate during the early stages of neural crest patterning. PMID- 11262246 TI - [Transport of critically ill pediatric patients: beginning of a new era]. PMID- 11262247 TI - [Instruments for measuring health-related quality of life in children and adolescents with asthma]. AB - OBJECTIVES: Measures of health-related quality of life (HRQOL) are proving to be useful in providing a comprehensive evaluation of illness and its effects on patients' daily lives. The aim of this review is to describe HRQOL instruments that are currently available to measure the HRQOL in children and adolescents with asthma. METHODS: The MEDLINE database from 1980to 2000was reviewed, as well as the website of the American Thoracic Society and the Quality of Life Research journal. Studies that included instruments measuring HRQOL in children with asthma were included as long as the instruments included met the selection criteria of multidimensionality, scoring was through standardized ordinal scales and psychometric properties were evaluated. RESULTS: Of the 21instruments initially identified 7were excluded because they did not meet one or more of the selection criteria. Fourteen instruments were included in the final review, 6were generic instruments for children (CHQ, KINDL, PedsQL, FS-IIR, RAND and CHIP-AE) and eight were specific to children with asthma (SSES, ASDQ, AMA, CAQ, LAQCA, PACQLQ, PAQLQ and APBC). The generic instruments measured the four basic aspects of HRQOL (symptoms, physical, mental and social functioning), whilst the majority of the specific instruments focused more closely on symptom measurement and physical functioning. Reliability (Cronbach's alpha) and construct validity were the most widely tested psychometric properties. In general, sensitivity to change was the least widely tested property, and only three disease-specific instruments were sensitive to change (LAQCQ, PACQLQ and PAQLQ). All the instruments could be self-administered. Only two of the generic instruments (FS-IIR and RAND) had been validated for use in Spain. At present two more generic instruments (PedsQL and CHIP-AE) and two specific instruments (PACQLQ and PAQLQ) are currently being validated. CONCLUSION: The availability of instruments to measure the HRQOL of children with asthma in Spain is currently limited. Validated versions of the PedsQL (generic) and PAQLQ (specific) instruments, both of which have been demonstrated to be useful in other countries, should shortly be available to measure the HRQOL of children with asthma in Spain. PMID- 11262248 TI - [Clinical findings in thoracic neuroblastomas]. AB - OBJECTIVES: To analyze clinical onset and other epidemiological findings in a group of children diagnosed with thoracic neuroblastoma. PATIENTS AND METHODS: We reviewed 46 non-metastatic thoracic neuroblastomas diagnosed and treated from January 1984 to December 1998 at the Pediatric Oncology Unit. Abdominal neuroblastomas were excluded. RESULTS: Mean age was 2 years and 5 months (range, 0-12 years). Eighty-five percent of the patients were infants (aged less than 1 year). Neuroblastoma was more frequent in males than in females. Fifty-seven percent of the patients were symptomatic and 43% were asymptomatic. The most frequent clinical findings were cough (18%) and palpable tumor or adenopathy (9%). Mean lag-time to diagnosis was 10 weeks in symptomatic patients. Thoracic x ray was the most useful diagnostic method (89%). Previous family history of neoplasias was present in 26%. Four patients (9%) had personal antecedents of other diseases. At the time of the study the event-free survival for the 46patients was 90% at 5 years (1-14 years). CONCLUSIONS: Thoracic neuroblastoma was more prevalent in male infants aged less than 6months. Most symptomatic patients were diagnosed in the 2first months after onset, and asymptomatic cases were common. The most frequent symptom was cough and most of the patients were diagnosed by thoracic x-ray. PMID- 11262249 TI - [Pulmonary function after liver transplantation in cystic fibrosis. Report of four cases]. AB - OBJECTIVES: The aim of this study was to evaluate pulmonary function in four patients with cystic fibrosis (CF) after liver transplantation. PATIENTS AND METHODS: From 1993 to 1997 three males and one female, aged 12 to 15 years, required liver transplantation for CF with cirrhosis and portal hypertension. Three had a history of esophageal variceal bleeding. In three patients, forced vital capacity (FVC) and forced expiratory volume in one second (FEV1) before liver transplantation were over 80 and 75% of predicted values, respectively; in the fourth patient FVC was 37% and FEV1was 26%. Two patients presented allergic bronchopulmonary aspergillosis before transplantation. Only one patient was chronically infected in sputum with multiresistant Pseudomonas aeruginosa and none had Burkholderia cepacea. RESULTS: After liver transplantation, only the patient with P. aeruginosa in sputum culture and the worst pulmonary function presented a complicated course requiring mechanical ventilation for 43 days followed by non-invasive nasal ventilation for 8 months. This patient died 19 months after transplantation. The remaining three patients, with better pulmonary function before transplantation, presented an uncomplicated course and currently lead normal lives. CONCLUSIONS: We conclude that liver transplantation can improve pulmonary function and is well tolerated in children with CF and mild or moderate pulmonary involvement. When pulmonary involvement is severe, combined lung and liver transplantation should be considered. PMID- 11262251 TI - [Causes of adolescent visits to the emergency department]. AB - OBJECTIVE: To determine the complaints leading adolescents to a third level emergency department, as well as the distinguishing features between the different groups of complaints. MATERIAL AND METHODS: Retrospective study of the emergency department reports of 170 adolescents selected from the 1,172 adolescents who visited the Hospital Sant Joan de Deu pediatric emergency department in May 1997. We excluded 517 patients with trauma. Statistical analysis was carried out with the chi-square test for qualitative variables and analysis of variance for quantitative variables; differences were accepted as significant at p,0.05. RESULTS: Eighty-nine patients were female and 81 were male. Median age was 13.8 years. The most frequent discharge diagnoses were abdomen pain in 26 patients, viral illness in 15 and depression-anxiety in 14. Complementary investigations were carried out in 60 patients. One hundred forty one patients were discharged. Of the 26 patients with psychiatric disorders, 18 were girls and 8 were boys. The proportion of girls was also higher among patients with abdominal and neurological symptoms, although not among patients with organic disorders (p50.03). Most of the patients with psychiatric antecedents were diagnosed with a psychiatric disorder. Ten (11.8%) of the 85 patients with organic complaints were admitted to hospital compared with 13(50%) of the 26 patients with psychiatric disorders (p,0.001). CONCLUSIONS: Adolescents visit emergency departments for a variety of complains. The proportion of psychiatric disorders and non-specific symptoms is high. PMID- 11262250 TI - [Triage criteria in a emergency department]. AB - OBJECTIVE: Accurate patient triage for the early identification of potentially seriously ill or high-risk children is needed due to the increasing demands made on paediatric emergency departments. MATERIAL AND METHODS: A set of triage criteria for patient selection was established. Between October 1998 and April 1999 we selected 20 days with maximum patient flow in our emergency department (average 225 children/day). Patients who satisfied the triage criteria were selected. The following variables were studied: age, sex, triage criteria, waiting time, visit time and final diagnosis. RESULTS: Five hundred and thirteen patients satisfied our triage criteria (11% of patients admitted to the emergency department). Median age was 3.4 years. The most frequent complaints (criteria) were dyspnea (36.4%), referral to the emergency department by another physician (29%), fever and rash (11.7%) and age under 1 month (5.8%). The most common causes of patient referral to hospital were acute abdominal pain and fever (31% each). Waiting and visit times were 29 and 55 minutes, respectively. CONCLUSION: Definition of selection criteria seems effective and equivalent to other triage systems and allows optimization of human resources. PMID- 11262252 TI - [Sexual abuse. Experience in a child sexual abuse unit]. AB - OBJECTIVE: To describe the clinical findings in children treated in a child sexual abuse unit. PATIENTS AND METHODS: We carried out a retrospective study of the clinical histories of children under suspicion of sexual abuse who visited the hospital from January 1992 to April 2000. Data on age, sex, need of urgent medical care, means of arrival, mechanism of discovery of abuse, parental separation, anamnesis, physical findings and complementary investigations were collected. The patients were then classified in four groups: normal, compatible, highly probable or certain sexual contact. In cases with a high probability of abuse, data of the aggressor's identity, place, duration and type of abuse were also collected. RESULTS: We studied 704 patients. Seventy-five percent were girls. The child's account of events was the most frequent means of discovering abuse (51%). Anamnesis was positive in 45% of the patients, genital examination was normal in 74% and anal examination was normal in 79%. According to our classification, 40% of the patients were normal, 11% were compatible, 41% were highly probable and 4% were of certain sexual contact. Ninety-two percent of aggressors were male. Molestation was the most frequent form of abuse and in 25% of cases abuse took place for more than 1 year. CONCLUSION: Diagnosis of sexual abuse is difficult and is almost always based on the child's account of events. The diagnostic yield of physical examination and complementary investigations is very low. We propose a diagnostic classification of four levels: normal, compatible, highly probable abuse and certain sexual contact. PMID- 11262253 TI - [Primary torsion of the greater omentum]. AB - OBJECTIVE: Primary torsion of the greater omentum is an infrequent cause of acute abdomen in children. A retrospective review was conducted to establish the prevalence and clinical features of omental torsion as a cause of acute abdominal pain in childhood. PATIENTS AND METHODS: We reviewed the clinical histories of the children given surgical treatment for torsion of the greater omentum in our hospital in the last 25 years. The following data were studied: age at presentation, sex, predisposing factors, symptomatology, complementary investigations, treatment and evolution. RESULTS: The male:female ratio among the 15 patients who underwent surgery was 2:1. Symptomology was similar to that of acute appendicitis with certain peculiarities such as a longer period of evolution at the moment of diagnosis, lower temperature and leucocytosis lower than would be expected in appendicitis at the same time of evolution and, in 12 patients, absence of vomiting. After surgical treatment evolution was satisfactory. Torsion was primary in 13 patients, secondary to inguinal hernia in 1 and secondary to cystic lymphangioma of the omentum in 1. The etiology and pathogenesis, as well as the diagnostic and therapeutic problems of this process, are discussed. CONCLUSIONS: In all the patients with primary torsion the clinical diagnosis was of acute appendicitis. Although primary torsion is classically associated with obesity, only 1 of the 13 patients weighed significantly more than the average for the same age and sex in our region and none of the patients showed a clearly associated anatomic malformation. PMID- 11262254 TI - [Uveitis and juvenile idiopathic arthritis]. AB - OBJECTIVE: Anterior uveitis is one of the most important extra-articular manifestations of juvenile idiopathic arthritis (JIA). The objective was to analyze the frequency of uveitis in patients with JIA and to describe its clinical and evolutive characteristics. PATIENTS AND METHOD: Among the 234 children diagnosed with JIA in our hospital, those presenting uveitis were studied. RESULTS: Seventeen children, 16 girls and 1 boy, presented uveitis in 28 eyes, representing a prevalence of 7.3%. Among patients with pauci- or oligo articular forms of the disease, the percentage increased to 13.3%; polyarticular forms accounted for 10%. Only one of the 12 patients with psoriatic arthritis developed uveitis. Mean age at diagnosis of the ocular condition was 4.5 years and the interval between diagnosis of arthritis to detection of uveitis was 661.5 months. In two patients uveitis was diagnosed before arthritis. Thirty-seven episodes of uveitic activity were identified, of which 27 were asymptomatic. Fifty-three percent of the affected eyes developed complications (posterior synechias in 43%, cataracts in 25%, in-band keratopathy in 18% and glaucoma in 7%). Surgery was required in six eyes. A marked loss of vision occurred in four eyes, despite ophthalmologic treatment. Conclusions Anterior uveitis is a cause of morbidity in JIA. Periodic ophthalmologic explorations are essential for early diagnosis and treatment. PMID- 11262255 TI - [Pediatric perspectives on medical transport]. AB - OBJECTIVE: To highlight the need for the organization and development of pediatric medical transport systems in Spain. METHODS: Analysis of the different types of pediatric medical transport, with a brief historical introduction and theoretical-practical discussion of the different types of transport team, their human and material resources and systematic organization. RESULTS AND CONCLUSIONS: The retrospective experience of a state medical emergency team is reported. The need for multidisciplinary coordination among pediatricians, critical care pediatricians, neonatologists, emergency physicians and nurses is identified as crucial in order to optimize resources and achieve the proposed objectives. PMID- 11262256 TI - [Sexual abuse in children: prevention of sexually transmitted diseases]. AB - When a child suffers from sexual abuse clinical guidelines must be established. There is a risk of infection from the following agents responsible for sexually transmitted diseases: the hepatitis B, hepatitis C and human immunodeficiency viruses, Neisseria gonorrhoeae, Chlamydia trachomatis, syphilis, herpes simplex virus, bacterial vaginosis, papillomavirus, Trichomonas vaginalis and Pediculus pubis. Therefore, a follow-up with periodic serological monitoring for 1year and immunoprophylaxis or chemoprophylaxis for some of these diseases should be started. Postpuberal girls should receive emergency contraception. PMID- 11262257 TI - [Management of parapneumonic pleural effusions]. AB - Pleural effusion in children is most often due to bacterial pneumonia. Between 0.6 and 2% of pneumonias are complicated by empyema and approximately 40% of children hospitalized with pneumonia have a pleural effusion. In recent years Streptococcus pneumoniae is the most prevalent organism. Treatment is based on the early and judicious use of antibiotics, imaging techniques, thoracocentesis, pleural drainage, fibrinolytics, thoracoscopy and thoracotomy. Indications for early pleural drainage are gross pus, positive Gram stain in pleural fluid, pleural glucose less than 50mg/dL, pleural fluid pH of less than 7 and sonographic evidence of loculations. Local fibrinolytics may decrease the need for surgical treatment, with a success rate between 38 and 100%, according to the effusion stage. Thoracoscopic debridement is useful in the fibrinopurulent stage with loculations, with favorable results in 30-100% of patients, also depending on the effusion stage. PMID- 11262258 TI - [Neonatal hearing loss screening using otoacoustic emission with Echocheck]. AB - OBJECTIVES: Few studies have been published on neonatal hearing loss screening using the Echocheck system (a new and easy method for detecting otoacoustic emissions). The aim of this study was to demonstrate how, in combination with auditory evoked potentials, this system can be used with satisfactory results for universal screening of all live births. METHODS: Otoacoustic emissions in the 1,000 infants born in our hospital during a 10-months period were checked by Echocheck. According to a preestablished protocol, neonates who failed the first screening test were sent for diagnosis by auditory brainstem responses. RESULTS: Coverage of the test was 99.3%. In 25% of the patients the test was repeated. Deafness was found in eight infants (only in two infants at high risk). The results obtained with this new method were compared with published results for the ILO systems. CONCLUSIONS: Echocheck provides more data than the standard recommended for neonatal hearing loss screening which, together with the results obtained in previous studies, suggests that Echocheck can be used like other systems for neonatal screening programmes. Since it is easy to use and cheaper than screening programmes, this system facilitates universal screening. PMID- 11262259 TI - [Concentrations of calcium and bone remodeling biomarkers in umbilical cord blood and urine of newborn infants during delivery]. AB - OBJECTIVE: To evaluate calcium, phosphorus, parathormone (PTH), osteocalcin, and alkaline phosphatase concentrations in cord blood, pyridinoline concentrations in urine, and the pyridinoline/creatinine ratio in order to indirectly assess late bone remodeling in the newborn by comparing concentrations of these substances with those in the mother during the third trimester and delivery. PATIENTS AND METHODS: Calcium, phosphorus, PTH, osteocalcin, and alkaline phosphatase levels in cord blood, pyridinolines in urine and the pyridinoline/creatinine ratio were evaluated in 38 healthy pregnant women during the last two months of pregnancy and during delivery. To determine concentrations of these substances, samples from vein umbilical cord blood were obtained immediately after birth and urine samples were obtained from the neonates within 24 hours of delivery. RESULTS: The mean calcium concentration in cord blood was higher than maternal values during delivery. Concentrations of PTH in cord blood were much lower than those in maternal blood during pregnancy, alkaline phosphatase levels were similar, and osteocalcin levels were higher. Mean urine pyridinoline concentrations in the neonates were much higher than those in the mother during the third trimester and delivery. CONCLUSION: During delivery newborn infants are hypercalcemic compared with their mothers, presenting a state of relative hypoparathyroidism. Concentrations of remodeling biomarkers are higher in the newborn than in the mother, revealing a higher rate of bone turnover in the fetus. PMID- 11262260 TI - [Breast-feeding in southern Catalonia. Epidemiological analysis of sociocultural and health factors influencing choice and duration]. AB - OBJECTIVE: To study the incidence and prevalence of breast-feeding and to determine the factors that influence the mother's decision to breast-feed or to use adapted milk. MATERIAL AND METHODS: Two hundred families were included in a survey in the hospital's maternity department. Those who breast-fed were followed up by means of a telephone call on days 15, 30, 90, and 180. RESULTS: On leaving hospital 78% of the neonates were receiving breast milk only. After 15 days, 89.7% of the neonates continued to receive breast milk and at 6 months this figure was 39%. Breast-feeding was discontinued after a mean of 2.5 months. The mean age of mothers who breast-fed was 30.2 years and that of mothers using adapted milk was 27.9 years (p,0.05). Mothers decided on the type of feeding before pregnancy (52.5%). This decision was unchanged by prenatal information except in the case of information provided by the family, especially if both parents were breast-fed (p,0.05). Doctors provided little information. The mother's level of education did not influence the decision to breast-feed although the higher the mother's education, the greater the tendency to breast feed (74.7% with primary education vs 81.5% with higher education). Being in paid employment did not influence the decision to breast-feed (76% of mothers worked vs 79% of mothers who did not). The main reasons for discontinuance were hypogalactia, "feeling hungry", and work. In general, giving up breast-feeding was the mother's decision. CONCLUSIONS: The information pregnant women receive on breast-feeding should be based on unified criteria. The implementation of joint protocols between primary and hospital care as well as breast-feeding support groups help mothers to begin and continue breast-feeding. PMID- 11262261 TI - [Vulvar and perianal erythema]. PMID- 11262262 TI - [Evolution of a case of tyrosinemia type I treated with NTBC]. AB - Tyrosinemia type I is an autosomal recessive inherited disorder caused by deficient fumarylacetoacetase activity. Treatment with 2-(2-nitro-4-trifluoro methylbenzoyl)-1,3-cyclohexanedione (NTBC), an inhibitor of 4 hydroxyphenylpyruvate dioxygenase, has successfully been applied for the last few years. Our aim was to evaluate the clinical and biochemical response to treatment with NTBC of a 18-year-old patient with a chronic form of tyrosinemia type I, whose main clinical feature was vitamin D-resistant rickets leading to severe osteoporosis with multiple bone fractures and skeletal deformities. After treatment, toxic metabolites became undetectable and porphobilinogen synthase activity returned to normal. Renal function improved, blood hemoglobin returned to normal and alfa-fetoprotein decreased. The patient's general condition greatly improved. However, the alfa-fetoprotein concentration slowly increased during the second year of NTBC treatment and hepatocellular carcinoma developed. NTBC treatment should be considered even in advanced cases of tyrosinemia type I, although only as a palliative therapy. PMID- 11262263 TI - [Gaucher's disease with D409H/D409H genotype. evolution with enzyme replacement therapy]. AB - Gaucher's disease is caused by mutations in the gene encoding glucocerebrosidase. The D409H mutation is the third most frequent mutation in Spain and has been associated with a particular phenotype, including oculomotor apraxia and cardiac valvular calcifications in late childhood. We report a 4-year-old patient, homozygous for the D409H mutation, who was diagnosed with Gaucher's disease at the age of 45days. Enzyme replacement therapy was started at the age of 2months. We report the patient's evolution after 4years of treatment. PMID- 11262265 TI - [Erythromycin and hypertrophic stenosis of the pylorus]. PMID- 11262264 TI - [Microangiopathic hemolytic anemia and thrombocytopenia. Thrombotic thrombocytopenic purpura]. AB - OBJECTIVE: Thrombotic thrombocytopenic purpura (TTP) or Moschovitz' syndrome is rare and is even rarer in childhood. Clinically, it is characterized by microangiopathic hemolytic anemia, thrombocytopenia, neurologic abnormalities, fever and renal dysfunction. The etiology is still unknown, although different factors such as large von Willebrand factor multimers and prostacyclin have been implicated. The acute form is more frequent, and in most cases the course is fulminant if treatment is not initiated. Laboratory data typically reveal hemolytic anemia, with schistocytes on the peripheral smear, diminished serum haptoglobin, and thrombocytopenia. MATERIAL AND METHODS: We present the clinical cases of two children, aged 4 and 7 respectively, with TTP, but with different evolution and treatment. Evolution was favorable in both patients. The first child recovered spontaneously. In the second plasmapheresis was required and produced remission of all the symptomatology. Normality has been maintained for 36 and 24 months respectively, and the children have presented no clinico biological alterations. PMID- 11262266 TI - [Should drinking water be boiled for 10 minutes to prepare feeding bottles?]. PMID- 11262278 TI - [Prescription patterns of older and newer antidepressants for geriatric depressive out-patients]. AB - OBJECTIVE: To study the prescription patterns of antidepressants for an elderly population. METHODS: 140 out-patients suffering an index depressive episode were studied and followed for at least one year. RESULTS: Demografics: 75% women. Mean age: 72.4%. Mayor depression: 71%. Late-onset: 41%. The most frequently used types the frist choice were SSRIs (48.6%) and Tricyclics (45%), the molecules initially most prescribed: Fluoxetine (26%), Mianserin (13%) and Paroxetin (10%). Initial choice of treatment achieves satisfactory remission in 32.9% of the cases. 57% of patients needed a frist malecule switch, 23% received only two molecules, 16% three, 5% four and 11% five molecules or more. After the first switch, the initial SSRI was substituted by another SSRI in 48% of the cases and by a Tricyclic in 30%. Initial Tricyclic chaged to a SSRI in 33% and another Tricyclic in 45%. The patterns of Tricyclics and SSRIs use were not essentially different and both were used globally in the same proportion, although the most prescribed molecules were Fluoxetine, Fluvoxamine and Paroxetine. Both groups achieved statistically similar number of remissions and suppressions by intolerance or ineficcacy but tricyclics were increasingly used from the frist switch onwards. PMID- 11262279 TI - [Open study of fluoxetine in children with autism]. AB - INTRODUCTION: Serotonergic dysfunction have been implicated in the pathophysiology of the autism. SRIs have shown efficacy in improving some symptoms in children with Autism. OBJECTIVES: To evaluate, in a pilot study, the efficacy and safety of fluoxetine in very young children with autism. METHOD: 1 year open-label trial was made with fluoxetine on 12 patients (3 to 13 years old) with Pervasive Developmental Disorder. CGI-severity was used to assess the severity and the improvement of symptoms. Individual symptoms improvement was assessed by both parents and therapists. Fluoxetine dose was titrated in all patients from 1.2 ml/day to reach a final doses of 3,6 or 5 ml/day in four weeks. Tolerance was assessed by collecting spontaneous adverse events. RESULTS: 11 children completed the study. Children experienced a moderate or marked improvement (final score in CGI 3 to 5). Communication and attention skills were improved. There were also a decrease of rituals, stereotypies and repetitive behaviours. Most common adverse events were increase of impulsivity and restlessness, other events were sleep disturbances and lost of appetite. 6 children needed concomitant medication with carbamacepine and 1 with levopromacine along the study. CONCLUSIONS: These results support that Fluoxetine at doses of 5 ml/day (20 mg/day) could help to improve some symptoms in Pervasive Developmental Disordersand allow to increase the effectiveness of therapies in these patients. More studies are needed to confirm these results. PMID- 11262280 TI - [Attention-deficit hyperactivity disorder and vulnerability to the development of alcoholism: use of the Wender-Utah Rating Scale for retrospective diagnostic of ADHD in the childhood of alcoholic patients]. AB - In the last years, it has been accumulated data about an important association between addictions and attention-deficit hyperactivity disorder (ADHD). Both disorders share clinic aspects and relevant biological markers, and for both it has been postulated alterations in the same cerebral systems. OBJECTIVE: To evaluate the rate of possible ADHD in the early ages of adult alcoholic patients, in contrast with controls. METHOD: It was realized an adaptation of Wender-Utah Rating Scale (WURS) and it was analyzed its psychometric characteristics. It was administered to 117 alcoholic patients and to 52 controls. RESULTS: The mean score of WURS is significatively higher in alcoholic group than in the control one (32.26 vs. 16.55, p< 0.0001). The percentage of alcoholic patients who has a score upper the different cut-off points (36 and 46) is also higher in alcoholic group than in the control one (36.75% vs 7.69%; p< 0.0005; 18.8% vs. 1.92%; p< 0.01, respectively). The mean score is higher in alcoholics with other comorbid addiction than in alcoholics without it (37.61 vs. 29.17; p< 0.018), and is higher in alcoholic patients who usually have intoxicated states in an high moderate grade than those who have it in a low-nule one. CONCLUSIONS: Among alcoholic patients exists an important group with high scores in the WURS, it could indicate high rates of ADHD in early ages. It was discused the clinic and etiopathogenetics implications, and the convenience of advancing in the developemnt of diagnostic tools. PMID- 11262281 TI - [Risperidone: a real alternative for patients treated with depot neuroleptics]. AB - OBJECTIVE: to determine the efficacy and tolerability of risperidone in schizophrenic outpatients. MATERIAL AND METHOD: DESIGN: multicentre, observational, 6 month follow-up study. PATIENTS: 421 schizophrenic (ICD-10 criteria) outpatients previously treated with depot neuroleptics and who were switched from depot medication to risperidone due to inefficacy and/or low tolerability. ASSESSMENT: BPRS, CGI, GAF, UKU, and DAI were administered at baseline and months 1, 3, and 6. STATISTICAL ANALYSIS: the Wilcoxon's T, Friedman's test, and Cochran's Q have been employed. RESULTS: Mean total dose of risperidone: 5.5 mg/d (SD 2.4). Scores on BPRS, CGI, GAF, UKU, DAI showed statistically significant improvements (p<0.0001). Significant decrease in the proportion of patients using anticholinergic drugs to control extrapyramidal symptoms and in the rate of hospitalizations. CONCLUSIONS: patients benefited from the change to risperidone in terms of efficacy, tolerance, compliance and admissions. Depot neuroleptic treated schizophrenic patients can benefit from treatment change to an oral atypic antipsychotic like risperidone to ease compliance, illness outcome and patient community integration. PMID- 11262282 TI - [Relation between patient's subjective complaints and family reports in patients with Alzheimer's type dementia]. AB - INTRODUCTION: Reduced ability to remember facts and events of everyday life is a common complaint in the elderly and is also the first sign of Alzheimer's disease (AD). The present research was designed to study the interrelationship between severity of memory complaints (MC), informant report, and performance in memory tests. MATERIAL AND METHODS: Seventy three (73) patients (41 age associated memory impairment (AAMI), and 32 Alzheimer's disease) and 30 normal controls were studied using the Subjective Memory Questionnaire (modified version), an objective memory battery and the Hamilton depression scale. RESULTS: Age Associated Memory Impairment subjects reported more severe MC (p< 0.001). No relationship was found between severity of MC and age, sex or educational level. Patient's MC didn't correlate with objective memory battery. A strong correlation was found with Hamilton depression score. Caregiver memory reports correlated with objective memory performances. CONCLUSION: Depressive features in AAMI and anosognosia in dementia patients would explain these results. These data suggest that informant report is the best predictor of patient's memory performance. PMID- 11262284 TI - [Simple schizophrenia: personality development or a condition?]. AB - The term simple schizophrenia was firstly used in the early twentieth century to designate a formal variety of schizophrenia. Without a precise definition the concept progressively looses its clinical significance as a subtype of schizophrenia and becomes paradoxically the representation of the pure fundamental disorder. The intersection of three clinical psychopathology categories psychoses, endogenity and process has historically determined the meaning of simple schizophrenia. The recent exclusion of the term simple schizophrenia from the diagnostic manuals and its taxonomical realignment as a personality disorder are the result of the nosological approach. The review of the concept of simple schizophrenia allow us to question the viability of a unique definition of clinical objects and to point out the configurating effect of theory on psychiatric taxonomy. PMID- 11262283 TI - [Biological basis of posttraumatic stress disorder]. AB - The postraumatic psychobiological stress disorder (PTSD) remains unclear although it is suggested that many systems are implicated. In this article, the PTSD biological aspects are reviewed, providing an exposition of main changes which are taking place in both the hypothalamic pituitary adrenal (HPA) and sympathetic nervous system (SNS). There are also some other mechanisms which are involved into human stress response such as thyroid function, neuropeptide Y, substance P and the opiod and glutamatergic system. Finally, we remark some changes which were found in neuroimaging techniques. PMID- 11262285 TI - [Training guidelines for linkage psychiatry]. AB - This review is aimed to provide some basic criteria intended to be useful as a C L training guideline. Teaching foundations for C-L Psychiatry are reviewed and the main features of the training schemes in C-L Psychiatry Services or Units are outlined taking in to considerations the four main perspectives such as clinical, teaching, research and management activities. Furthermore, a core curriculum for a teaching is recommended. Teaching principles for C-L Psychiatry should be meditated; due to the absence of national publications in this specific area, this article might give the opportunity to get started in an open debate. In spite of a worldwide promising future for C-L Psychiatry, in Spain there are still few Services or units developed and fewer of those providing a formal teaching program, so an opportunity for a major shift is served. PMID- 11262286 TI - [Obsessive psychotic disorder in adolescence]. AB - Relationship between obsessive-compulsive disorder (OCD) and psychotic pathology is a controversial one. Case reports indicate that OCD with psychotic features is generally more severe and antidepressant-refractory than neurotic OCD. Behaviour therapy is shown to be ineffective in OCD patients with psychotic features. One of the subtypes includes reactive paranoid forms, with fluctuating clinical course, in obsessive patients with delusional thoughts that improve with neuroleptic augmentation of the serotonin reuptake inhibitor (SRI) treatment for OCD. We present the case of a 17-year-old woman with family history of OCD, starting with panic symptoms after cannabis use, but suddenly developing OCD with avoidant behavior and delusional ideas of self-reference and persecution. Response to behavior therapy and SRI and neuroleptics is analyzed. PMID- 11262300 TI - [Evolutionary theories on schizophrenia]. AB - The universal distribution of schizophrenia associated with the low fertility of these patients outlines the paradox: why doesn't diminish the prevalence of this dysfunction? A critical revision is made of the main hypotheses that have thought about to solve this question. Some theories have postulated a physiologic or social advantage. Others have analysed this illness from the mark of the Evolutionary Psychology. We stand out the theories that have looked for an association of a Darwinian approach with genetic or neurodevelopmental data. Finally a possibility of integration of the evolutionary and neuropsychological models is proposed. PMID- 11262299 TI - [Neuroimaging and basic symptoms of schizophrenia]. AB - The complexity of the clinical picture of schizophrenia is an essential feature of the disease that has led to attempts at organizing the symptoms and clinical course into different subtypes or grouping them into symptom constellations. Variations in response to treatment, particularly biological treatment, justifies the effort of differentiation. Various authors have tried to define primary or fundamental symptoms, that are directly related with the underlying abnormality, and secondary symptoms, which involve adaptive or reactive mechanism. Similarly, first-order symptoms, pathognomic symptoms, and second-order symptoms have been described. A constant latent problem in psychopathology is to determine the degree in which first-order symptoms are primary. PMID- 11262301 TI - [Fronto-temporal dementia]. AB - Defined barely one decade ago, Frontotemporal Dementia emerges as a new clinical entity that reestablishes the classical Pick's disease as part of a extensive syndrome with important and unsuspected prevalence. Frontotemporal Dementia includes all primary degenerative processes starting in the anterior portions of the brain. This type of dementia is clinically characterized by behavior and personality disorders, more than cognitive alterations. In the present research we performed a bibliographic review of the subject including clinical characteristics, laboratory tests, and neuropathology. Furthermore we assessed areas that require further development. PMID- 11262303 TI - [Naltrexone in the treatment of alcohol addiction]. AB - This paper reviews preclinical research which demonstrates the involvement of the opioid system in the reinforcing effects of alcohol, and the effects of naltrexone, a long-acting, nonselective opioid receptor antagonist to reduce alcohol intake. Naltrexone (50 mg/day) may prevent the return to drinking by blocking the pleasurable effects or "high" associated with alcohol drinking, and relapse rates were reduced. The most common adverse effects reported include nausea and vomiting and it does not appear to be hepatotoxic in dosages recommend. Thus, naltrexone appears to offer significant therapeutic benefits, when used with behavioural treatment for alcohol dependent patients. PMID- 11262302 TI - [Role of personality in eating behavior disorders]. AB - Personality disorders are common in eating disorders and preliminary reports indicate that this type of disorders implicate a poor prognosis in anorexia and bulimia nervosa. Cluster C personality disorders, particularly avoidant and dependent personality disorder are the most frequent in anorexia nervosa. In bulimia nervosa, however, cluster B personality disorders, including borderline and histrionic personality disorders, are more frequent. Furthermore, temperament could differentiate anorectic patients from bulimic patients. Eating disorders seem to be associated with high scores in neuroticism and harm avoidance. However, while anorexia nervosa patients might have higher persistence, bulimia nervosa patients seem to have an impulsive temperament. According to the character model of Cloninger, both anorexia and bulimia present lower scores in the dimension self-directedness. PMID- 11262304 TI - [Management and treatment of severe mental disorders in pregnancy]. AB - The pharmacological treatment of serious mental disorders in the pregnancy, supposes a clinical dare by the possible repercussions on the fetus and the pregnancy: theratogenesis, perinatal syndrome or postnatal sequels in the development. The electroconvulsive therapy (ECT) as much takes implicit a minimum risk for the mother as for the fetus and therefore, it must be located in the highest positions of the therapeutic decision trees. In the present article, are reviewed the consequences of the pharmacological treatment and the ECT in the serious mental disorders during the pregnancy. Is referenced to all of the pharmacological groups and with respect to the ECT: their indications, counterindications, complications and technical procedures advisables. Finally is reviewed the guide line for each syndromical group of psychiatric diseases. PMID- 11262305 TI - [Neuropsychological changes in schizophrenia and its modification]. AB - The main experimental works about neuropsychological impairments of schizophrenia are reviewed. The underlying mechanisms of the cognitive deficits are set in a framework of the limited capacity model. In second point, the current status of the modificability of the cognitive deficits and the clinical and psychosocial consequences of this deficits are presented. At least, neuropsychological rehabilitation programs are reviewed from a clinical point of view. PMID- 11262318 TI - [Evaluation of residents]. PMID- 11262319 TI - [Mortality attributable to smoking in Castilla and Leon]. AB - OBJECTIVE: To calculate the contribution of tobacco consumption to mortality in Castilla y Leon, and its effect on premature mortality in this community. DESIGN: Cross-sectional study. SETTING: Community. PARTICIPANTS: Population of Castilla y Leon. MEASUREMENTS AND MAIN RESULTS: The source of information used were the life statistics from the National Institute of Statistics for 1995. The proportion and number of deaths attributed to tobacco were calculated by working out aetiological population fractions. 9.37% of the total deaths occurring were attributed to tobacco consumption. Mortality was higher in men (7.22%) than in women (2.27%). The diagnostic categories contributing most to these figures were tracheal-bronchial-lung cancer (36.9%) and ischaemic heart disease (22%). CONCLUSIONS: The study shows the importance of tobacco dependency as a public health problem in Castilla y Leon, and its major contribution to premature death. PMID- 11262320 TI - [Alcohol consumption among students of a health area. Habits and beliefs]. AB - OBJECTIVES: 1. To describe the habits and conduct of school students (13-17 years old) around alcohol in the Cartagena Health Area. 2. To discover the factors underlying consumption. 3. To analyse beliefs and behaviour about consumption. DESIGN: Cross-sectional descriptive study. SETTING: Health area. PARTICIPANTS: 16,657 school students. Sample population (n = 1004). Large-group sampling (sampling unit: class-room), accuracy = 3%, CI = 95%, p = q = 50%, stratified by school and geographical area. MEASUREMENTS AND MAIN RESULTS: Anonymous, self filled questionnaire (33 open and closed questions). SELECTION CRITERIA: attendance at class on the day of the questionnaire. VARIABLES: social, economic and cultural data on parents and students, course, type of family, alcohol consumption (starting age, access, individual habits, friends and family), beliefs about consumption, study profile of consumer. STATISTICAL ANALYSIS: description of variables, chi-squared test, t test, logistical regression. 99.2% filled in the questionnaire. Students' average age was 15.84 (SD, 1.21). 89.96% lived with their parents. 83.4% had consumed alcohol on some occasion, with no relationship to current consumption (p > 0.05). Starting age was 13.77 (SD, 1.78), 48.5% in discos/bars and 19% at home (p > 0.05). 47.5% had got drunk on some occasion (p > 0.05). 88.2% customarily drank alcohol mixed with other drinks (p > 0.05). 51.8% drank at week-ends (2.4% every day) (p > 0.05). 63.3% thought it easy to acquire drinks: 48.9% in supermarkets, 26.4% at petrol stations (p = 0.038). 71% of consumers agreed that "alcohol is a drug" (p = 0.005). 13% believed "it does not cause dependency" (p = 0.00001). 23.3% thought that it is not true to say "the person who does sport is healthier and does not take alcohol". CONCLUSIONS: The pattern of consumption is similar to that found in other autonomous communities, although in greater quantity and without differences in kind. Mean age of consumer: 15 years old. They usually drink alcohol mixed with other soft drinks, normally at week-ends. They are very ignorant of the effects of consumption. PMID- 11262321 TI - [Factors associated with dental enamel defects in the first molar in a population of children]. AB - OBJECTIVES: To assess factors related to defects in the tooth enamel of the first permanent molar. Factors studied include, pediatric assistance, hospital admittance, high and low respiratory illness, varicella, gastroenteritis, ear infections, urinary tract infections, and different pharmacological treatments. DESIGN: A retrospective case control study. PARTICIPANTS: Cases and controls were selected randomly from a pool of 1382 scholars born in years 1980, 1981 and 1982. MEASUREMENTS AND MAIN RESULTS: The study was conducted in march and may of year 2000. Finally 48 cases and 148 controls were selected. A dental examination was conducted at 8 years of age using the WHO criteria. Defects in the tooth enamel were defined according to the FDI criteria. We've measured and compared the study variables along the first five years of live. The epidemiological association was quantified by means of the odds ratio (OR) an its 95% confidence interval (CI). In the first year of live we estimated for pediatric assistance an OR of 2.26 (95% CI 1.05-4.92); in the second year for Urinary Tract Infections (UTI) we obtained an OR of 25.27 (95% CI 2.98-562.2); in the third year for UTI an OR of 6.68 (95% CI 1.01-54.52); in the fourth year the OR for pneumonia was 13.45 (95% CI 1.36-324.5) and finally in the fifth year the significant OR were: 2.56 (95% CI 1.23-5.34) for ear infections, 2.28 (95% CI 1.03-5.03) for macrolides and OR of 2.20 (95% CI 1.08-4.50) for anticongestive medications. CONCLUSIONS: UTI and pneumonia show a high association with the presence of defects in the tooth enamel. Other variables such as high frequency of pediatric assistance, hospital admittance, ear infections, varicella, and pharmacological treatments with macrolides, cefolosporines, anticongestive medications and lungs medications showed a weak association. PMID- 11262322 TI - [Professionals and managers in face of training]. AB - OBJECTIVES: To analyse ongoing education within primary care teams from managers' perspective. DESIGN: Cross-sectional descriptive study. SETTING: The Mother Infant Health Programme of Andalusia in primary care. INTERVENTION: The views of health centre professionals were obtained through 8 groups. The type of health district and the number of years in operation were used as criteria for division. The views of the area managers were collected through a self-filled questionnaire from all the managers in the autonomous community. MEASUREMENTS AND MAIN RESULTS: Professionals believed it necessary to keep up to date and train in new technology. They thought that training activities should focus on questions such as evaluation of programmes, focus of risk, counselling and communication, and certain techniques such as IUD insertion. Both managers and professionals coincided in recognising the effect of training on the development of mother child services. There was consensus among professionals in affirming that training increases the quality of care delivered. Access to training was the main incentive element used by most area managers and one of those most highly valued by professionals. The professionals affirmed that in recent years there had been obstacles to their education by attending scientific events outside their areas. Many professionals stated that the economic assistance of the pharmaceutical industry for courses caused inequalities between doctors and nurses in access to training. Professionals were very critical of training combined with specialist training. CONCLUSIONS: Professionals think they have sufficient training for mother-child health care. It is generally recognised that family doctors are better trained for working on health programmes than other professionals at the centres. Ongoing training was rated very highly by both professionals and directors, as it was seen as a motivating element and thought to be an activity almost solely carried out at health centres. PMID- 11262323 TI - [ADA-97 criteria, prevalence of diabetes mellitus and the most southern counties of Catalonia]. AB - OBJECTIVES: To find the prevalence of diabetes mellitus (DM) and disturbed basal glucaemia in the population over 24 years old and the relationship of this to factors of risk of becoming diabetic. DESIGN: Descriptive, cross-sectional population study. SETTING: Community, county (Baix Ebre-Montsia-Terra Alta) and primary care (9 health districts) context. PATIENTS: Inhabitants of the three most southerly counties of Catalonia over 24 years old (106,551 out of 132,938). INTERVENTIONS: We randomised from the data base of the computer service of the Catalan Institute of Health (100% coverage), for an estimated prevalence of 15%, losses of 20%, 95% CI and +/- 5% accuracy, a sample of 245 people (we studied 198). We informed each doctor of his/her patients in the study. If the patient was diabetic, his/her doctor filled out a questionnaire; if not, the doctor also requested from the laboratory two glucaemia analyses taken after fasting. MEASUREMENTS AND MAIN RESULTS: Using the diagnostic criteria and screening methods of the ADA-1997, we obtained the following results: 1. 14.1% prevalence of DM (1.5% new diagnoses); 4% prevalence of disturbed basal glucaemia. 2. Likelihood of diabetes: age > 45 (4.7 times greater); triglyceridaemia > 250 mg/dl (4.5 times greater); BMI > 27 (2.9 times). CONCLUSIONS: High prevalence of DM, with high proportion known through primary care. We know the prevalence of disturbed basal glucaemia. DM-related risk factors in our population were: age > 45, BMI > 27 and hyper-triglyceridaemia. Our primary care focus can better manage the resources dedicated to DM. PMID- 11262324 TI - [Should we ask for vitamin B12 concentrations in the initial study of dementia?]. PMID- 11262325 TI - [Costs of economic evaluation of health technologies]. PMID- 11262326 TI - [Thyroid disease in pregnancy]. PMID- 11262327 TI - [The COMBO project. Criteria and guidelines for the combined treatment of type 2 diabetes. Consensus document (I)]. PMID- 11262328 TI - [Health center pediatricians as consultants for family doctors]. PMID- 11262329 TI - [Pre-transfer moment. Something more than a transfer of powers in health]. PMID- 11262330 TI - [Is the Papanicolaou smear useful as aid for diagnosing some sexually transmitted infections?]. AB - OBJECTIVE: We undertook this study to assess the validity of cytologic diagnosis of sexually transmitted infections like: bacterial vaginosis (BV), tricomoniasis and candidiasis using the Papanicolaou (Pap) smear. DESIGN: Prospective, descriptive transverse study. SETTING: The present study was carried out in the Health Center Dr. Jose Castro Villagrana in Tlalpan, Mexico, D.F. from January 1997, to February 2000. PARTICIPANTS: Routine Pap smears and vaginal secretion smears were collected from two hundred and seventy one patients ranged from age 16-66 years, with cervicovaginitis diagnosis. MEASUREMENTS AND MAIN RESULTS: Of the 271 patients, 92 (33.9%) had bacterial vaginosis diagnosed by Amsel criteria, 47 (17.3%) had candidiasis by culture and 5 (1.8%) had tricomoniasis by wet smear. The Bethesda system for diagnosing BV on Pap smear had 66% sensitivity and a specificity of 86%. The respective positive predictive and negative predictive value were 79% and 84%. Therefore, compared to the Candida culture, cervical cytologic test results had a sensitivity of 21% and specificity of 99%. The predictive positive predictive and negative predictive values were 90% and 85%. CONCLUSIONS: Specificity tended to be higher than sensitivity, in other words cytology tended to be more efficient in identifying women without sexually transmitted infection than in identifying those with infection. In summary, the Pap smear should not be used in lieu of more effective diagnostic test for sexually transmitted disease, and treatment should not be based on cytologic findings alone. PMID- 11262331 TI - [Can we improve the therapeutic management of allergic rhinitis in primary care?]. AB - OBJECTIVE: To evaluate improvement in therapeutic management of allergic rhinitis. DESIGN: Study of level of quality (longitudinal, prospective, intervention). SETTING: Primary care. PATIENTS AND OTHERS PARTICIPANTS: First evaluation (second quarter of 1995): 73 out of 305 patients were sampled (confidence 5%, accuracy 10%). Second evaluation (second quarter of 1996). Sample of 51 patients from a total of 210. INTERVENTIONS: Explicit criteria and standard procedure, based on consensus, for rhinitis treatment and an overall indicator of the general quality of rhinitis management were analysed. Criterion 1 (C1): correctly scaled treatment; criterion 2 (C2): initial treatment of choice with inhaled corticoids; criterion 3 (C3): correct use of oral corticoids or immunotherapy; criterion 4 (C4): coadjutant therapy according to predominant symptoms. Corrective measures: ongoing training and routine use of guide to practice. STATISTICS: index of compliance with criteria, Chi squared and Fisher's Z tests of a tail to compare both evaluations. MEASUREMENTS AND MAIN RESULTS: First evaluation: index of compliance with C1 = 59% (CI +/- 11), C2 = 41% (CI +/- 11), C3 = 90% (CI +/- 6) and C4 = 83% (CI +/- 8). Criteria and summary indicator obtained better results in patients attended by allergists. Second evaluation with overall improvement: C1 = 68.6% (CI +/- 13), C2 = 57% (CI +/- 13), C3 = 94% (CI +/- 6), C4 = 98% (CI +/- 3). Significant differences for C4 and C2 (p < 0.05). Overall quality and quality of criteria improved for patients attended in our environment. The summary indicator went up from 35.6% to 57% (p = 0.019), with the quality levels (C1-C4) becoming the same as those of the patients with allergy attended and with significant differences in the first evaluation disappearing. CONCLUSIONS: Ongoing training and routine use of guides to practice enables the therapeutic management of allergic rhinitis in primary care to be improved. PMID- 11262332 TI - [Agreement in the measurement of blood pressure among different health professionals. Are mercury sphygmomanometers reliable?]. AB - OBJECTIVES: To assess reliability in terms of inter-observer agreement of blood pressure (BP) readings. Various health professionals and measuring systems. Influence of observer's experience. DESIGN: Observational, descriptive, cross sectional study. SETTING: Urban health centre, Cordoba. PARTICIPANTS: 131 hypertensive, randomised patients, belonging to a functional care unit. 11 were excluded. MEASUREMENTS: To reduce variability: course on the right way to take blood pressure, otoscope and verification of visual sharpness of observers, calibration and validation of measuring devices, limited time and blinding of measurements. 4 BP measurements per patient: 3 with mercury sphygmomanometer (2 simultaneously, one individual) and one with an automatic device. Descriptive, clinical and somatometric variables were gathered. Inter-observer agreement was evaluated through the intraclass correlation coefficient (ICC), the mean of differences method (MDM) and the simple concordance index (CI). An ICC > 0.75 was thought acceptable. A difference > 5 mmHg was thought clinically relevant (MDM and CI). MAIN RESULTS: Acceptable consistency for MDM: alone, systolic and diastolic pressure of OBS 1/ OBS 2, bi-auricular, -6.1/+8.9 mmHg and -6.8/+5.8 mmHg. Less favourable results: for systolic and diastolic pressure: OBS 1/AUTO 20.9/25.0 and -16.4/15.1; OBS 2/AUTO -22.8/24.4 and -16.6/15.2. Remaining intervals always > 10 mmHg; CI > 0.75 in all comparisons except diastolic pressure OBS 1/AUTO and diastolic pressure OBS 2/AUTO (0.69 in both cases). 41% of comparisons were > 5 mmHg. No differences in less expert professionals were found. CONCLUSIONS: Inaccuracy of the standard BP measurement method (mercury sphygmomanometer) for MDM and CI. Contradictory conclusions according to method of measurement. Differences not clinically acceptable. PMID- 11262333 TI - [Delivery of care pressure, frequency of attendance and pediatric morbidity at a health center. Age and seasonal variations]. AB - OBJECTIVE: To inform on the demand for primary care paediatric services, with a view to improving health care delivery. DESIGN: Observational retrospective study. SETTING: Estella Health Centre (Navarra). PATIENTS: All patients attended during 1999 at one of the two paediatric clinics at the health centre. They were divided by ages into breast-feeders (0-12 months), pre-school (1-5 years old), primary school (6-9) and adolescents (10-14). MEASUREMENTS: The date, sex, age, type of consultation (on demand/scheduled) and health problem (CIPSAP-2) of the 6611 consultations were recorded. Frequency of visits, care pressure and seasonal distribution, related to age and type of consultation, were calculated. RESULTS: There were 4600 on-demand consultations (69.6%) and 2011 scheduled ones (30.4%). Total frequency of attendance was 5.46, which was greater in on-demand pre-school children (6.4) and scheduled breast-feeders (13.3). Overall patient pressure was 28.2, though less (p < 0.05) in the summer months. The most common health problems in on-demand consultations were respiratory illnesses (52.8%), infectious diseases (7.5%), neuro-sensory problems (6.8%), accidents (6.0%) and digestive problems (4.7%). There was negative correlation (p < 0.05) between age and the prevalence of respiratory diseases, whereas age was directly proportional (p < 0.05) to the prevalence of accidents and locomotive illnesses. Respiratory and infectious diseases were more common (p < 0.05) in the autumn and winter months. CONCLUSIONS: Paediatrics at a health centre suffers patient overload, with acute illnesses of the respiratory apparatus, and to a lesser extent infectious diseases, the main causes of consultation. Health check-ups are becoming steadily more important. There needs to be better coordination between PC teams in order to unify diagnostic and therapeutic criteria, for the more thorough non-hospital paediatrics becomes, the greater its effect on improving child health quality. PMID- 11262334 TI - [Continuing medical education about the use of antilipemic agents in elderly patients aged 65-75 years]. AB - OBJECTIVE: The objective is evaluating the efficacy of the educative intervention to primary care physicians, about the accurate dyslipidaemia management in population between 65 and 75 years old with hypercholesterolemia. DESIGN: Simple blind random clinical trial. SETTING: Area 10 primary care (National Institute of Health of Spain). STUDY SUBJECTS: . Thirty eight primary care physicians of Area 10. Seven hundred and five patients between 65 and 75 years old with dyslipidaemia. INTERVENTIONS: Clinical session to physicians about the dyslipidaemia management, reinforced with the shipment of the accurate management criteria and bibliographic information. Physicians were followed up for one year. RESULTS: The therapeutic management varied (p = 0.03) in the experimental group after educative intervention. The dietetic therapeutic increased 6.56 percent (p = 0.21), the therapeutic with HMG-CoA-reductase inhibitors increased 4.16 percent (p = 0.36), and the therapeutic with fibric-acid derivates decreased 4.22 percent (p = 0.24). The criteria fulfillment rate of accurate dyslipidaemia management did not vary (p = 1.0) in the control group (44.3 percent) and there was hardly any variation (from 49.4 percent to 49.1) in the experimental group (p = 0.96). The fulfillment rate decreased 7,56 percent (p = 0.25) when dyslipidaemia managed with only diet. The fulfillment improved 17,17 percent (p = 0.14) if dyslipidaemia managed with fibric-acid derivates, and it improved 17,58 percent (p = 0.06) if was managed with HMG-CoA-reductase inhibitors. CONCLUSIONS: The educative session to primary care physicians reinforced with the shipment of the received information, is not likely to relieve the criteria fulfillment rate of accurate management of population between 65 and 75 years old with hypercholesterolemia. PMID- 11262335 TI - [Prevalence of goitre and iodine deficiency in a school population from a traditionally endemic health area]. AB - OBJECTIVES: To determine the current prevalence of simple goitre in the school population of a health district where goitre is traditionally endemic. Calculation of the deficiency or otherwise of iodine through the determination of mean urinary excretion of iodine in the population under study. DESIGN: Cross sectional descriptive study. SETTING: Olvera Health District (Cadiz). PARTICIPANTS: School students in the health district between 6 and 14 years old out of a total of 1969. Sample size of 92 school students was chosen at random, for a 95% confidence interval. MEASUREMENTS AND MAIN RESULTS: Dependent variables were the existence of goitre found in a physical examination, urinary excretion of iodine measured in microg/dl in the first urine of the morning, origin of water consumed and habitual consumption of iodised salt in their diet. 87% of the population under study habitually drank water from the normal supply, 4% from wells or springs, and 9% mineral water. 57% of parents did not know whether the salt in their normal diet was iodised or not. 29.3% of school students included in the study had some degree of goitre. The mean excretion of iodine in urine was 13.78 microg/dl (95% CI, 12.30-15.26). Ioduria was below 9.9 microg/dl in 28.2%, within the endemic figures. CONCLUSIONS: The mean amount of iodine in urinary excretion in the sample means that the risk of developing goitre is low, although the prevalence of goitre continues at endemic figures. PMID- 11262337 TI - [Anorexia and bulimia: need to distinguish primary prevention from early detection in the context of health education in teaching facilities]. PMID- 11262336 TI - [Nutrition among Moroccan immigrants in the community of Madrid: factors affecting the choice of food]. AB - OBJECTIVES: To describe the main characteristics of the diet of Marrocan immigrants in the Autonomous Community of Madrid, and to explore the factors that may influence their selection of foods. DESIGN: Cross-sectional study. Two-stage cluster sampling of 179 immigrants. SETTING: Health Area number 6 of the Autonomous Community of Madrid. PARTICIPANTS: Marrocan immigrants, older than 14 years of age who had lived in Spain more than 3 months. INTERVENTIONS: Personal interview including a 24-hours recall of food consumption. Descriptive analysis and logistic regression using SPSS 8.0 for Windows. RESULTS: The factors that have a larger influence in the food consumed are the age of the person, whether the person lives in a couple or not, and whether the person is able to read and write in Spanish. Gender and number of years living in Spain had very limited or no influence in the food items referred to as consumed by the person. Conclusions. The results of our study contrast with findings from studies conducted in other countries. The diet of the immigrants in our study would seem to become more varied and balanced, and therefore improve, with the progression of the acculturation process, with age and in people who live with their partners. People who have migrated recently have a less varied diet and may be proned to suffer nutritional deficiences. PMID- 11262338 TI - [Cost effectiveness analysis in health]. PMID- 11262339 TI - [Cardiovascular risk on the Internet]. PMID- 11262340 TI - ["Fortunately, there's nothing wrong with you." PSA and prostate cancer]. PMID- 11262342 TI - [Modification of the prescription of anticholesteremic agents]. PMID- 11262344 TI - [Do we have to rush to diagnose fibromyalgia?]. PMID- 11262348 TI - [Manolo Gomis] PMID- 11262349 TI - [Microorganisms isolated from patients with pertussis-like syndrome] PMID- 11262351 TI - [Value of MR myelography in the diagnosis of the spine disorders]. AB - To evaluate the utility of myelography obtained with MR imaging (MR-myelography) as a complementary tool in patients studied with a conventional MR examination of the spine. 275 consecutive patients were included. All of them were studied with MR-myelography in 2 planes, coronal and sagittal, with a turbo spin-echo single shot technique, as a complement to a conventional MR study of the spine; 130 were males and 145 women, with ages ranging from 20 to 71 years (mean, 45 years). The analyzed variables were age, sex, vertebral segment studied, alteration of the dural sac, intradural nerve roots, emergent roots, and presence of intradural lesions, meningeal cysts, and spinal stenosis. The added value of MR-myelography regarding conventional MR was categorized. MR-myelography obtained new information in 88 cases (32%), being considered irrelevant information in 42 cases and relevant in 46 cases (16.7% of all cases) (amputations of the emergent roots and alterations of the intradural roots). MR-myelography did not contribute to any type of additional information to the conventional MR study in 187 cases (68% of all studies). MR-myelography is a rapid acquisition technique that supplements the conventional MR study of the spine, contributing with relevant new information in the analysis of the spine diseases 16.7% of cases. PMID- 11262350 TI - [Mediterranean diet improves low density lipoprotein susceptibility to oxidative modifications]. AB - Most experts, specially from Anglo-Saxon countries, recommend a low fat diet in order to prevent cardiovascular diseases. However, mortality rate by ischemic cardiopathy is low in Mediterranean countries, probably because of the consumption of a Mediterranean diet, with a high level of monounsaturated fats provided by the olive oil. We have conducted this study in order to investigate the possible influence of this kind of diet on the oxidation of LDL in vitro, the key element for the development of atherosclerosis. PATIENTS AND METHODS: 41 healthy male subjects were submitted to three consecutive 4-week diets. The first was a saturated fat-rich diet (SAT diet, 38% fat, 20% saturated). This was followed by a low fat diet (NCEP-I, 28% fat, 10% saturated) and after that a Mediterranean diet (38% fat, 22% monounsaturated fat). Plasma levels of total cholesterol, LDL-c, HDL-c, triglycerides, apolipoproteins A-I and B, -tocopherol, and the in vitro susceptibility to oxidation of LDL particles. Both hypolipidemic diets produced a significant decrease in total cholesterol, LDL-c, and apo-B plasma levels. However, it was only the NCEP-I diet that revealed a decrease in the HDL-c. The shift from a saturated fat-rich diet, or a diet rich in carbohydrates, to a Mediterranean diet increased the resistance of LDL particles to oxidation increasing the lag time period (p < 0.038), and decreasing (p < 0.001) the progression rate of the curve of oxidation of LDL. Our results point out two positive consequences of the consumption of a Mediterranean diet by healthy young males, compared with the low fat diet recommended by most Anglo Saxon experts. On the one hand, the Mediterranean diet increases HDL-c plasma levels, and on the other hand, it decreases the susceptibility of LDL to oxidation. This is why the Mediterranean diet must be recommended in order to prevent cardiovascular diseases. PMID- 11262353 TI - [Seroprevalence of toxoplasmosis in women of childbearing age, 1992-1999]. AB - To determine the need of prenatal screening for toxoplasmosis in our hospital from a seroepidemiological point of view. PATIENTS AND METHODS: The prevalence of IgG anti-T. gondii was retrospectively analyzed in 7.090 women of childbearing age attended in the Hospital Clinic of Barcelona from February 1992 to April 1999. The association among the seroprevalence and the variables year, age, birthplace (province of Barcelona/other provinces) and place of residence (urban/rural) was analyzed. A decreasing trend was observed in the prevalence (p < 0.001), currently being < 40% in the average women between 15 and 45 years. Infection was also directly related to age of women (p < 0.001) and birthplace out of the province of Barcelona (p = 0.001). Habitat (rural or urban) was not associated with seroprevalence. Prenatal screening for toxoplasmosis is necessary due to the high rate of seronegative women exposed to infection and the evidence of a high number of primoinfections in the childbearing period. PMID- 11262352 TI - [Sib-sib correlations of cardiovascular risk factors: the Cuenca study]. AB - To analyze the sib-sib correlation of the cardiovascular risk factors. SUBJECTS AND METHOD: A cross-sectional study was made in 115 sibling pairs. Sociodemographic variables, weight, height, body mass index, systolic blood pressure, diastolic blood pressure and fasting plasma total cholesterol, LDL-C, HDL-C, apo A-I, apo B and Lp(a) concentrations. Partial correlation coefficients adjusted for age and body mass index for plasmatic lipid and lipoprotein levels in sibling pairs showed positive values range from 0.27 for triglyceride to 0.53 for apo B/apo A-I ratio (p < 0.005). Likewise, the intraclass correlation coefficients ranged from 0.20 to 0.48. Both coefficient were higher in brother brother pairs than in sister-sister pairs. The Quetelet index coefficients were slightly lower than lipid coefficients, and showed similar values among the different kinds of sibling pairs. Blood pressure coefficients were low (p > 0.05). CONCLUSIONS: An association for lipid and lipoprotein plasma concentrations has been evidenced in sibling pairs. This association is stronger in brother-brother pairs than in sister-sister pairs. This fact could indicate a sex linked heredability of lipid and lipoprotein levels. A weaker association for body mass index has been observed. Sib-sib aggregation of blood pressure levels is not significant. PMID- 11262354 TI - [Inappropiate visits to emergency department of a general hospital]. AB - The proportion of inadequate attendances at emergency department (ED) is 20-80%. The suitability of attendances at ED was evaluated using an Hospital Emergency Suitability Protocol, which was validated. 37.9% of attendances were inappropriate and they are more frequent in children. Patients who were referred by a doctor, with trauma or surgical consulted more adequately. The suitability of attendances at ED are related with illness. PMID- 11262356 TI - [Health Sciences University: an innovatory option?]. PMID- 11262355 TI - [Susceptibility of low density lipoproteins to oxidation and the Mediterranean diet]. PMID- 11262357 TI - [Parapneumonic effusions and empyema]. PMID- 11262358 TI - [PPARgamma and thiazolidinediones, something more than a treatment for diabetes]. PMID- 11262360 TI - [Immigration in the Canary Islands: a change in health problems]. PMID- 11262359 TI - [Tetany as the first manifestation of common variable immunodeficiency]. PMID- 11262361 TI - [Diffuse infiltrative CD8 lymphocytosis syndrome in a patient with HIV-1 infection]. PMID- 11262365 TI - [Surgery for active valvular endocarditis]. PMID- 11262364 TI - [Diabetes mellitus and coronary revascularization. The controversy continues]. PMID- 11262366 TI - [Impact of diabetes mellitus on the late clinical outcome of coronary revascularization with stents]. AB - INTRODUCTION: The Influence of diabetes mellitus in the late outcome of coronary stenting remains controversial. AIM: The aim of this study was to determine the late clinical outcome of diabetics in comparison with non diabetics and to establish whether there are subgroups of diabetic patients with a greater need for target lesion revascularization. METHODS: Two hundred sixteen consecutive patients (74 diabetics; 95 stents in 90 lesions and 142 non diabetics) who had successfully undergone coronary stenting were included in the study and followed over 17.6 +/- 10 months. The clinical events evaluated were target lesion revascularization, death and acute myocardial infarction. Independent predictive variables of target lesion revascularization were studied in both groups of patients. RESULTS: The diabetic patients presented greater cardiovascular mortality (6.7% vs 1.4%; p=0.02) but the incidence of infarction was similar in the two groups (2.7% vs. 3.5%; p=0.6). The accumulated rate of target lesion revascularization at two years was 18.2% in diabetics vs 13.3% in non diabetics (p=0.09), respectively. The presence of three vessel disease (p=0.014), history of arterial hypertension ([=0.011) and residual stenosis > 0% (p=0.005) were specific predictive factors of target lesion revascularization for diabetic patients and together with vessel diameter < 3mm (p<0.001) subgroups of diabetics were independently selected with a significantly greater incidence of target lesion revascularization than the non diabetic patients. CONCLUSIONS: Following coronary stenting, diabetic patients show a greater cardiovascular mortality than non diabetics, but only some subgroups of diabetics (small vessels extensive coronary disease, associated arterial hypertension, residual stenosis) show a significantly greater risk of target lesion revascularization. PMID- 11262367 TI - [Adult congenital anomalies of the coronary arteries described over 31 years of angiographic studies in the Asturias Principality: main angiographic and clinical characteristics]. AB - OBJECTIVE: The aim of this study was to determine the incidence of adult congenital anomalies of the coronary arteries over 31 years of angiographic studies, describing their angiographic and clinical characteristics. The results have been compared with the main series published. METHODS: The diagnostic angiographic reports done in the Principado de Asturias from 1968 to 1999 are reviewed. In those in which a congenital anomaly was diagnosed, the clinical report and the angiography were studied. The initial course of the anomaly was defined following angiographic criteria. RESULTS: Thirteen thousand five hundred reports were reviewed describing 75 patients with 75 anomalies (0.5%) including: anomalous origin of the left circumflex coronary artery (n = 24), coronary artery fistulae (n = 21), both coronary arteries arising from the left coronary sinus (n = 15), single coronary arteries (n = 6), both coronary arteries arising from the right coronary sinus (n = 2), separated origin of anterior descending and left circumflex coronary arteries (n = 3), anterior descending artery arising from the right coronary sinus (n = 2), and others (n = 1). Angiographic studies were done because of: angina (59%), dysnea (25%), atypical chest pain (7%), syncope (3%), dizziness (3%) and palpitations (3%). The initial course was retroaortic in all the circumflex arteries, interarterial in the right coronaries, anterior in the anterior descending arteries and retroaortic, septal and combined, in the left coronaries. CONCLUSIONS: Adult congenital anomalies of the coronary arteries are not very common and are usually casual findings of diagnostic angiographic studies. Left circumflex coronary artery anomalies are the most frequently diagnosed. PMID- 11262368 TI - [Creatine kinase increases after coronary interventions]. AB - INTRODUCTION AND OBJECTIVES: Percutaneous revascularization has led to an important change in the treatment of patients with symptomatic ischemic heart disease in recent years. There is controversy concerning the incidence and prognostic significance of postprocedural increases in creatine kinase. The aim of this study was to assess the incidence of these elevations and the related factors and to observe the prognosis of patients with and without creatin kinase elevations. METHODS: We reviewed 447 patients in whom an angioplasty was done in our department from January 1997 to June 1998, excluding 138 patients with myocardial infarction in the previous four days or unsuccessful angioplasty. Creatine kinase was measured in all patients at 0, 4, 8 and 24 hours after angioplasty. We analyzed the incidence of elevated levels of creatine kinase following coronary surgery and the characteristics of the patients in comparison with a control group made up of patients who, at a similar time had undergone a similar angioplasty procedure including, a similar vessel and type of lesion, and equivalent left ventricular function but without elevated serum levels of creatine kinase. Major adverse coronary events were defined as: cardiac death, nonfatal myocardial infarction, new revascularization and unstable angina in which hospitalization was required. RESULTS: Out of 309 patients studied, an elevation in creatine kinase was observed in 24 patients (7.7%). Complications related to the procedure were found in 50% of these elevations, most of which involved side branch occlusion. There were no differences with respect to the demographical or anatomical characteristics of the lesions in the groups studied. During the follow-up of 9.5 months, complications were observed in 37.5% of the group of patients with elevated creatine kinase levels and in 20% of the control group, but this difference did not achieve statistical significance. CONCLUSIONS: Creatine kinase elevations are produced in 7.7% of the patients after coronary angioplasty. Complications related to the procedure were observed in 50% of the cases, most being side branch occlusion and no complications were seen in the remaining patients. Continuous measurement of creatine kinase after angioplasty shows a low sensitivity for detecting complications during follow-up. New, more sensitive and specific cardiac markers, such as troponin, could define this group of patients. PMID- 11262370 TI - [Treatment with intrapleural streptokinase for coagulated hemothorax after cardiac surgery with cardiopulmonary bypass]. AB - INTRODUCTION AND OBJECTIVES: Coagulated hemothorax is a complication of cardiac surgery with cardiopulmonary bypass. The objective of this study was to present the authors' experience in the intrapleural infusion of streptokinase for the treatment of this complication. METHODS: From January 1996 to June 1999, nine patients (6 males, 3 females, age range: 1-75 years) were clinically and radiographically diagnosed with coagulated hemothorax after cardiac surgery. All patients were treated with intrapleural infusion of streptokinase at a standard dose of 250,000 units in adult patients and 12,000 U/kg in pediatric cases. In cases of occluded chest drainage, the position of the patient was changed and drainage was opened. RESULTS: In all the cases clinical and radiological improvement was observed and 100 to 200 ml of hemothorax was obtained on drainage. One patient died of multiorgan failure due to the underlying disease not related to the procedure. No alteration were observed in hematological tests including coagulation. The other 8 patients were discharged from hospital and remain without pulmonary compromise to date. CONCLUSION: Treatment of coagulated hemothorax with intrapleural infusion of streptokinase is a useful procedure and avoid the need for surgical drainage of hemothorax. PMID- 11262369 TI - [Surgical reconstruction of intervalvular fibrous body in active infective endocarditis]. AB - INTRODUCTION AND OBJECTIVES: Surgery for infective endocarditis with paravalvular abscesses and fibrous body destruction has the highest mortality and morbidity rates in this disease with high surgical risk. We report a new approach of radical resection of the abscess and affected tissues and reconstruction of the heart with pericardium as an alternative to conventional surgery. METHODS: In the last two years six patients with infective endocarditis, paravalvular abscesses and fibrous body destruction underwent surgery (five prostheses with infective endocarditis). The main indication for surgery was persistent sepsis despite adequate antibiotic treatment in five patients and congestive heart failure in one. After wide resection of the abscesses and fibrous body the heart was reconstructed with glutaraldehyde-fixed bovine pericardium. RESULTS: There was no hospital mortality. The median bypass and clamp times were 198 and 174 minutes, respectively. One patient presented complete AV block and a permanent transvenous pacemaker was implanted. Doppler echocardiographic studies performed in all the patients prior to discharge indicated that no patient had patch dehiscence or paravalvular leaks. Patients were followed a mean of 15 months with no deaths or other complications being reported. CONCLUSIONS: Resection of the abscesses and fibrous body, and reconstruction of the heart with glutaraldehyde-fixed bovine pericardial patch is a radical, feasible technique with all infected tissues being resected to thereby prevent reinfection or paravalvular leaks. PMID- 11262371 TI - [Influential factors in mortality rate from congenital heart disease. Study of 1,216 children in the Autonomous Community of Murcia (1978-1990)]. AB - INTRODUCTION: In the last few years, important progress has taken place in management of congenital heart disease. These changes have had an influence on diagnosis, preoperative management, surgery treatment and postoperative care, giving rise to better results in the treatment of children suffering from congenital heart disease. AIM: To assess the results of congenital heart diseases in a reference hospital by comparing two periods with reference to both diagnosis and therapeutical management. We also intend to investigate the influence that factors such as the existence of extracardiac congenital malformations and heart surgery have on mortality. PATIENTS AND METHODS: Our sample group was made up of 1,216 children suffering from congenital heart disease. Their ages ranged from 1 day to 7 years old. These children were born over a period of thirteen years and studied at the paediatric cardiology unit in a reference hospital in the Autonomous Community of Murcia, a region of Spain. We retrospectively analysed their development by individual heart diseases (and their associated factors), and the global results. Our research was divided into two periods: between 1978 and 1983, and between 1984 and 1990. Differences were found regarding diagnosis and treatment. RESULTS: a) Mortality rate from congenital heart disease decreased in the period between 1984 and 1990 in comparison to the period between 1978 and 1983, from 28 to 21,7% (p < 0.05); b) individually, the mortality rate decreased with statistical significance in two diseases: interventricular communication and patent ductus arteriosus, and c) there is a higher mortality rate of patients with no surgery treatment and/or extracardiac malformations. CONCLUSION: Progress in the management of congenital heart disease has led to a more favourable outcome in the last years. PMID- 11262373 TI - [The heart myth]. PMID- 11262372 TI - [Practice guidelines of the Spanish Society of Cardiology on cardiac arrythmias]. PMID- 11262375 TI - [Infection and atherosclerosis]. PMID- 11262374 TI - [New concepts on the mechanisms of ventricular fibrillation]. PMID- 11262377 TI - [Aortic pseudoaneurysm]. PMID- 11262376 TI - [Hemopericardium pericardiocentesis guided with echo-contrast]. PMID- 11262378 TI - [Shock secondary to extrinsic compression of the right atrium by postoperative mediastinal hematoma. Pseudotumor echocardiography image in right atrium]. AB - We present the case of a 64 year-old patient in whom an aortic Saint Jude prosthesis, a Cosgrove's mitral annulus and triple coronary artery by-pass graft were implanted, and who presented with shock related to extrinsic compression of the right atrium by a mediastinal hematoma within the first postoperative month. Transthoracic echocardiogram showed a right atrial <>, hampering right atrium drainage. The extrapericardial location of the hematoma is of note and was diagnosed with the aid of thoracic computerized tomography. We present the case of a 64 year-old patient in whom an aortic Saint Jude prosthesis, a Cosgrove's mitral annulus and triple coronary artery by-pass graft were implanted, and who presented with shock related to extrinsic compression of the right atrium by a mediastinal hematoma within the first postoperative month. Transthoracic echocardiogram showed a right atrial <>, hampering right atrium drainage. The extrapericardial location of the hematoma is of note and was diagnosed with the aid of thoracic computerized tomography. PMID- 11262379 TI - [Large myxoma of the right atrium]. AB - We describe the case of a patient in whom two-dimensional echocardiography, performed due to dissociated cholestasis and jugular ingurgitation, demonstrated a huge mass in the right atrium which prolapsed in the right ventricle. Intraoperative transesophageal echocardiography was performed to further assess the dimension and characteristics of the mass and to discard the involvement of associated structures. The patient underwent a cardiopulmonary bypass surgery and the mass (12 * 5 cm) was removed without complications. Histologic examination confirmed the diagnosis of myxoma. This case is of interest because of the size of the mass, and is centered in the diagnosis following clinical suspicion due to the pattern of dissociated cholestasis and jugular ingurgitation leading to surgery to prevent the potential embolic complications. PMID- 11262380 TI - [Isolated pulmonary endocarditis on native valve in elderly patient without predisposing factors]. AB - Isolated infective endocarditis in the native pulmonary valve is an unusual clinical entity in patients without predisposing factors and in non-intravenous drugs users. We present the case of a 75-year-old patient, with a subacute clinical picture of fever and pulmonary cavity nodules, admitted to our hospital with an initial suspected diagnosis of pulmonary tuberculosis. The presence of Enterococcal bacteremia in hemocultive and the documentation of a large vegetation in pulmonary valve by transtoracic and transesophageal echocardiography were key factors for final diagnosis. PMID- 11262381 TI - [Radiofrequency ablation of recurrent monomorphic ventricular tachycardia in a patient with severe systemic scleroderma]. AB - A case of progressive systemic scleroderma in a 33 year-old woman who was referred to our Arrhythmia Unit due to daily palpitations and dizziness is presented. The 24-hour Holter recording showed monomorphic ventricular tachycardia which lasted several minutes. Hemodynamic study showed dilated right chambers and right ventricular dysfunction, without pulmonary hypertension. Left ventricular angiography and coronary arteries were normal. During programmed electrical stimulation, two different ventricular tachycardia were induced and ablated with radiofrequency on the right ventricle. The patient remains free of recurrence of tachycardia after (10 months of follow up). Patients with progressive systemic scleroderma may present several different cardiac arrhythmias. Involvement of the right ventricle is particularly frequent as is the origin of ventricular tachycardia in this ventricle. Radiofrequency catheter ablation is safe and effective in the management of these patients. PMID- 11262386 TI - Tip60 and HDAC7 interact with the endothelin receptor a and may be involved in downstream signaling. AB - Endothelins exert their biological effects through G protein-coupled receptors. However, the precise mechanism of downstream signaling and trafficking of the receptors is largely unknown. Here we report that the histone acetyltransferase Tip60 and the histone deacetylase HDAC7 interact with one of the ET receptors, ETA, as determined by yeast two-hybrid analysis, glutathione S-transferase pull down assays, and co-immunoprecipitation from transfected COS-7 cells. In the absence of ET-1, Tip60 and HDAC7 were localized mainly in the cell nucleus while ETA was predominantly confined to the plasma membrane. Stimulation with ET-1 resulted in the internalization of ETA to the perinuclear compartment and simultaneously in the efflux of Tip60 and HDAC7 from the nucleus to the same perinuclear compartment where each protein co-localized with the receptor. Upon co-transfection with ETA into COS-7 cells, Tip60 strongly increased ET-1-induced ERK1/2 phosphorylation, whereas HDAC7 had no significant effect. We thus suggest that protein acetylase and deacetylase interact with ETA in a ligand-dependent fashion and may participate in ET signal transduction. PMID- 11262387 TI - A catalytically inactive mutant of type I cGMP-dependent protein kinase prevents enhancement of large conductance, calcium-sensitive K+ channels by sodium nitroprusside and cGMP. AB - The activation of large conductance, calcium-sensitive K(+) (BK(Ca)) channels by the nitric oxide (NO)/cyclic GMP (cGMP) signaling pathway appears to be an important cellular mechanism contributing to the relaxation of smooth muscle. In HEK 293 cells transiently transfected with BK(Ca) channels, we observed that the NO donor sodium nitroprusside and the membrane-permeable analog of cGMP, dibutyryl cGMP, were both able to enhance BK(Ca) channel activity 4-5-fold in cell-attached membrane patches. This enhancement correlated with an endogenous cGMP-dependent protein kinase activity and the presence of the alpha isoform of type I cGMP-dependent protein kinase (cGKI). We observed that co-transfection of cells with BK(Ca) channels and a catalytically inactive ("dead") mutant of human cGKIalpha prevented enhancement of BK(Ca) channel in response to either sodium nitroprusside or dibutyryl cGMP in a dominant negative fashion. In contrast, expression of wild-type cGKIalpha supported enhancement of channel activity by these two agents. Importantly, both endogenous and expressed forms of cGKIalpha were found to associate with BK(Ca) channel protein, as demonstrated by a reciprocal co-immunoprecipitation strategy. In vitro, cGKIalpha was able to directly phosphorylate immunoprecipitated BK(Ca) channels, suggesting that cGKIalpha-dependent phosphorylation of BK(Ca) channels in situ may be responsible for the observed enhancement of channel activity. In summary, our data demonstrate that cGKIalpha alone is sufficient to promote the enhancement of BK(Ca) channels in situ after activation of the NO/cGMP signaling pathway. PMID- 11262388 TI - Modulation of contractile activation in skeletal muscle by a calcium-insensitive troponin C mutant. AB - Calcium controls the level of muscle activation via interactions with the troponin complex. Replacement of the native, skeletal calcium-binding subunit of troponin, troponin C, with mixtures of functional cardiac and mutant cardiac troponin C insensitive to calcium and permanently inactive provides a novel method to alter the number of myosin cross-bridges capable of binding to the actin filament. Extraction of skeletal troponin C and replacement with functional and mutant cardiac troponin C were used to evaluate the relationship between the extent of thin filament activation (fractional calcium binding), isometric force, and the rate of force generation in muscle fibers independent of the calcium concentration. The experiments showed a direct, linear relationship between force and the number of cross-bridges attaching to the thin filament. Further, above 35% maximal isometric activation, following partial replacement with mixtures of cardiac and mutant troponin C, the rate of force generation was independent of the number of actin sites available for cross-bridge interaction at saturating calcium concentrations. This contrasts with the marked decrease in the rate of force generation when force was reduced by decreasing the calcium concentration. The results are consistent with hypotheses proposing that calcium controls the transition between weakly and strongly bound cross-bridge states. PMID- 11262389 TI - Epidermal growth factor activates hyaluronan synthase 2 in epidermal keratinocytes and increases pericellular and intracellular hyaluronan. AB - Hyaluronan is an abundant and rapidly turned over matrix molecule between the vital cell layers of the epidermis. In this study, epidermal growth factor (EGF) induced a coat of hyaluronan and a 3-5-fold increase in its rate of synthesis in a rat epidermal keratinocyte cell line that has retained its ability for differentiation. EGF also increased hyaluronan in perinuclear vesicles, suggesting concurrent enhancement in its endocytosis. Cell-associated hyaluronan was most abundant in elongated cells that were stimulated to migrate by EGF, as determined in vitro in a wound healing assay. Large fluctuations in the pool size of UDP-N-acetylglucosamine, the metabolic precursor of hyaluronan, correlated with medium glucose concentrations but not with EGF. Reverse transcriptase polymerase chain reaction (RT-PCR) showed no increase in hyaluronan synthases 1 and 3 (Has1 and Has3), whereas Has2 mRNA increased 2-3-fold in less than 2 h following the introduction of EGF, as estimated by quantitative RT-PCR with a truncated Has2 mRNA internal standard. The average level of Has2 mRNA increased from approximately 6 copies/cell in cultures before change of fresh medium, up to approximately 54 copies/cell after 6 h in EGF-containing medium. A control medium with 10% serum caused a maximum level of approximately 21 copies/cell at 6 h. The change in the Has2 mRNA levels and the stimulation of hyaluronan synthesis followed a similar temporal pattern, reaching a maximum level at 6 h and declining toward 24 h, a finding in line with a predominantly Has2-dependent hyaluronan synthesis and its transcriptional regulation. PMID- 11262390 TI - Glucagon-like peptide (GLP)-2 action in the murine central nervous system is enhanced by elimination of GLP-1 receptor signaling. AB - Glucagon-like peptide-2 (GLP-2) regulates energy homeostasis via effects on nutrient absorption and maintenance of gut mucosal epithelial integrity. The biological actions of GLP-2 in the central nervous system (CNS) remain poorly understood. We studied the sites of endogenous GLP-2 receptor (GLP-2R) expression, the localization of transgenic LacZ expression under the control of the mouse GLP-2R promoter, and the actions of GLP-2 in the murine CNS. GLP-2R expression was detected in multiple extrahypothalamic regions of the mouse and rat CNS, including cell groups in the cerebellum, medulla, amygdala, hippocampus, dentate gyrus, pons, cerebral cortex, and pituitary. A 1.5-kilobase fragment of the mouse GLP-2R promoter directed LacZ expression to the gastrointestinal tract and CNS regions in the mouse that exhibited endogenous GLP-2R expression, including the cerebellum, amygdala, hippocampus, and dentate gyrus. Intracerebroventricular injection of GLP-2 significantly inhibited food intake during dark-phase feeding in wild-type mice. Disruption of glucagon-like peptide 1 receptor (GLP-1R) signaling with the antagonist exendin-(9-39) in wild-type mice or genetically in GLP-1R(-)/- mice significantly potentiated the anorectic actions of GLP-2. These findings illustrate that CNS GLP-2R expression is not restricted to hypothalamic nuclei and demonstrate that the anorectic effects of GLP-2 are transient and modulated by the presence or absence of GLP-1R signaling in vivo. PMID- 11262391 TI - Bcl10 and MALT1, independent targets of chromosomal translocation in malt lymphoma, cooperate in a novel NF-kappa B signaling pathway. AB - At least two distinct recurrent chromosomal translocations have been implicated in the pathogenesis of MALT lymphoma. The first, t(1;14), results in the transfer of the entire Bcl10 gene to chromosome 14 wherein Bcl10 expression is inappropriately stimulated by the neighboring Ig enhancer. The second, t(11;18), results in the synthesis of a novel fusion protein, API2-MALT1. Until now, no common mechanism of action has been proposed to explain how the products of these seemingly unrelated translocations may contribute to the same malignant process. We show here that Bcl10 and MALT1 form a strong and specific complex within the cell, and that these proteins synergize in the activation of NF-kappaB. The data support a mechanism of action whereby Bcl10 mediates the oligomerization and activation of the MALT1 caspase-like domain. This subsequently activates the IKK complex through an unknown mechanism, setting in motion a cascade of events leading to NF-kappaB induction. Furthermore, the API2-MALT1 fusion protein also strongly activates NF-kappaB and shows dependence upon the same downstream signaling factors. We propose a model whereby both the Bcl10.MALT1 complex and the API2-MALT1 fusion protein activate a common downstream signaling pathway that originates with the oligomerization-dependent activation of the MALT1 caspase like domain. PMID- 11262392 TI - The rational of catalytic activity of herpes simplex virus thymidine kinase. a combined biochemical and quantum chemical study. AB - Most antiherpes therapies exploit the large substrate acceptance of herpes simplex virus type 1 thymidine kinase (TK(HSV1)) relative to the human isoenzyme. The enzyme selectively phosphorylates nucleoside analogs that can either inhibit viral DNA polymerase or cause toxic effects when incorporated into viral DNA. To relate structural properties of TK(HSV1) ligands to their chemical reactivity we have carried out ab initio quantum chemistry calculations within the density functional theory framework in combination with biochemical studies. Calculations have focused on a set of ligands carrying a representative set of the large spectrum of sugar-mimicking moieties and for which structural information of the TK(HSV1)-ligand complex is available. The k(cat) values of these ligands have been measured under the same experimental conditions using an UV spectrophotometric assay. The calculations point to the crucial role of electric dipole moment of ligands and its interaction with the negatively charged residue Glu(225). A striking correlation is found between the energetics associated with this interaction and the k(cat) values measured under homogeneous conditions. This finding uncovers a fundamental aspect of the mechanism governing substrate diversity and catalytic turnover and thus represents a significant step toward the rational design of novel and powerful prodrugs for antiviral and TK(HSV1) linked suicide gene therapies. PMID- 11262393 TI - Calcium-dependent activation of nuclear factor regulated by interleukin 3/adenovirus E4 promoter-binding protein gene expression by calcineurin/nuclear factor of activated T cells and calcium/calmodulin-dependent protein kinase signaling. AB - An increase in the intracellular Ca(2+) concentration controls a diverse range of cell functions, including gene expression, apoptosis, adhesion, motility, and proliferation. We have investigated Ca(2+) regulation of gene expression in rat aortic smooth muscle cells. We found that the expression of nuclear factor regulated by interleukin 3 (NFIL3)/adenovirus E4 promoter-binding protein (E4BP4)/basic region/leucine zipper (bZIP) type of a transcription factor that has a very important function in cell survival, was activated by thapsigargin (TG). This activation was inhibited by chelation of extra- or intracellular Ca(2+), suggesting that the induction by TG was dependent on the elevation of [Ca(2+)](i). Specific inhibition of calcineurin or calcium/calmodulin-dependent protein kinase (CaM kinase) by chemical means impaired the TG-induced NFIL3/E4BP4 expression. Expression of dominant negative forms of calcineurin or nuclear factor of activated T cells (NFAT) inhibited the induction of NFIL3/E4BP4 mRNA by TG. These results suggest that intracellular Ca(2+) plays a critical role in regulating gene expression of NFIL3/E4BP4 by calcineurin/NFAT and CaM kinase signaling in vascular smooth muscle cells. PMID- 11262394 TI - G Protein beta subunit types differentially interact with a muscarinic receptor but not adenylyl cyclase type II or phospholipase C-beta 2/3. AB - In comparison with the alpha subunit of G proteins, the role of the beta subunit in signaling is less well understood. During the regulation of effectors by the betagamma complex, it is known that the beta subunit contacts effectors directly, whereas the role of the beta subunit is undefined in receptor-G protein interaction. Among the five G protein beta subunits known, the beta(4) subunit type is the least studied. We compared the ability of betagamma complexes containing beta(4) and the well characterized beta(1) to stimulate three different effectors: phospholipase C-beta2, phospholipase C-beta3, and adenylyl cyclase type II. beta(4)gamma(2) and beta(1)gamma(2) activated all three of these effectors with equal efficacy. However, nucleotide exchange in a G protein constituting alpha(o)beta(4)gamma(2) was stimulated significantly more by the M2 muscarinic receptor compared with alpha(o)beta(1)gamma(2). Because alpha(o) forms heterotrimers with beta(4)gamma(2) and beta(1)gamma(2) equally well, these results show that the beta subunit type plays a direct role in the receptor activation of a G protein. PMID- 11262395 TI - Bcl-rambo, a novel Bcl-2 homologue that induces apoptosis via its unique C terminal extension. AB - The Bcl-2 family of proteins plays a central regulatory role in apoptosis. We have identified a novel, widely expressed Bcl-2 member which we have named Bcl rambo. Bcl-rambo shows overall structural homology to the anti-apoptotic Bcl-2 members containing conserved Bcl-2 homology (BH) motifs 1, 2, 3, and 4. Unlike Bcl-2, however, the C-terminal membrane anchor region is preceded by a unique 250 amino acid insertion containing two tandem repeats. No interaction of Bcl-rambo with either anti-apoptotic (Bcl-2, Bcl-x(L), Bcl-w, A1, MCL-1, E1B-19K, and BHRF1) or pro-apoptotic (Bax, Bak, Bik, Bid, Bim, and Bad) members of the Bcl-2 family was observed. In mammalian cells, Bcl-rambo was localized to mitochondria, and its overexpression induces apoptosis that is specifically blocked by the caspase inhibitors, IAPs, whereas inhibitors controlling upstream events of either the 'death receptor' (FLIP, FADD-DN) or the 'mitochondrial' pro-apoptotic pathway (Bcl-x(L)) had no effect. Surprisingly, the Bcl-rambo cell death activity was induced by its membrane-anchored C-terminal domain and not by the Bcl-2 homology region. Thus, Bcl-rambo constitutes a novel type of pro-apoptotic Bcl-2 member that triggers cell death independently of its BH motifs. PMID- 11262396 TI - Vav2 activates c-fos serum response element and CD69 expression but negatively regulates nuclear factor of activated T cells and interleukin-2 gene activation in T lymphocyte. AB - Vav1 and Vav2 are members of the Dbl family of guanine nucleotide exchange factors for the Rho family of small GTPases. Although the role of Vav1 during lymphocyte development and activation is well characterized, the function of Vav2 is still unclear. In this study, we compared the signaling pathways regulated by Vav1 and Vav2 following engagement of the T cell receptor (TCR). We show that Vav2 is tyrosine-phosphorylated upon TCR stimulation and by co-expressed Src and Syk family kinases. Using glutathione S-transferase fusion proteins, we observed that the Src homology 2 domain of Vav2 binds tyrosine-phosphorylated proteins from TCR-stimulated Jurkat T cell lysates, including c-Cbl and SLP-76. Like Vav1, Vav2 cooperated with TCR stimulation to increase extracellular signal-regulated kinase activation and to promote c-fos serum response element transcriptional activity. Moreover, both proteins displayed a similar action in increasing the expression of the early activation marker CD69 in Jurkat T cells. However, in contrast to Vav1, Vav2 dramatically suppressed TCR signals leading to nuclear factor of activated T cells (NF-AT)-dependent transcription and induction of the interleukin-2 promoter. Vav2 appears to act upstream of the phosphatase calcineurin because a constitutively active form of calcineurin rescued the effect of Vav2 by restoring TCR-induced NF-AT activation. Interestingly, the Dbl homology and Src homology 2 domains of Vav2 were necessary for its inhibitory effect on NF-AT activation and for induction of serum response element transcriptional activity. Taken together, our results indicate that Vav1 and Vav2 exert overlapping but nonidentical functions in T cells. The negative regulatory pathway elicited by Vav2 might play an important role in regulating lymphocyte activation processes. PMID- 11262397 TI - The signal transduction pathway underlying ion channel gene regulation by SP1-C Jun interactions. AB - During neuronal differentiation, an exquisitely controlled program of signal transduction events takes place, leading to the temporally and spatially regulated expression of genes associated with the differentiated phenotype. A critical class of genes involved in this phenomenon is that made up of genes encoding neurotransmitter-gated ion channels that play a central role in signal generation and propagation within the nervous system. We used the well established PC12 cell line to investigate the molecular details underlying the expression of the neuronal nicotinic acetylcholine receptor class of ion channels. Neuronal differentiation of PC12 cells can be induced by nerve growth factor, leading to an increase in neuronal nicotinic acetylcholine receptor gene expression. Nerve growth factor initiates several signal transduction cascades. Here, we show that the Ras-dependent mitogen-activated protein kinase and phosphoinositide 3-kinase pathways are critical for the nerve growth factor mediated increase in the transcriptional activity of a neuronal nicotinic acetylcholine receptor gene promoter. In addition, we show that a component of the Ras-dependent mitogen-activated protein kinase pathway, nerve growth factor inducible c-Jun, exerts its effects on receptor gene promoter activity most likely through protein-protein interactions with Sp1. Finally, we demonstrate that the target for nerve growth factor signaling is an Sp1-binding site within the neuronal nicotinic acetylcholine receptor gene promoter. PMID- 11262398 TI - MACROH2A2, a new member of the MARCOH2A core histone family. AB - MACROH2As are core histones that have a unique hybrid structure consisting of an amino-terminal domain that closely resembles a full-length histone H2A followed by a large nonhistone region. The human MACROH2A1 gene, on chromosome 5, encodes two MACROH2A subtypes, MACROH2A1.1 and MACROH2A1.2, produced by alternate splicing. Here we report the identification of MACROH2A2, a new MACROH2A subtype encoded by a separate gene on human chromosome 10, MACROH2A2. The amino acid sequence of human MACROH2A2 is 68% identical to human MACROH2A1.2. We show by immunofluorescence on mouse tissue sections that MACROH2A2, like MACROH2A1.2, is concentrated in the inactive X chromosome. However, MACROH2A2 has a very different pattern of expression in the cell types present in the liver and kidney. When MACROH2A2 and MACROH2A1.2 are present in the same nucleus, they have a similar, though nonidentical, pattern of localization, with both subtypes present in the inactive X chromosome. Our results suggest a developmental role for MACROH2A subtypes. PMID- 11262399 TI - An "elongated" translation elongation factor Tu for truncated tRNAs in nematode mitochondria. AB - We have found the gene for a translation elongation factor Tu (EF-Tu) homologue in the genome of the nematode Caenorhabditis elegans. Because the corresponding protein was detected immunologically in a nematode mitochondrial (mt) extract, it could be regarded as a nematode mt EF-Tu. The protein possesses an extension of about 57 amino acids (we call this domain 3') at the C terminus, which is not found in any other known EF-Tu. Because most nematode mt tRNAs lack a T stem, domain 3' may be related to this feature. The nematode EF-Tu bound to nematode T stem-lacking tRNA, but bacterial EF-Tu was unable to do so. A series of domain exchange experiments strongly suggested that domains 3 and 3' are essential for binding to T stem-lacking tRNAs. This finding may constitute a novel example of the co-evolution of a structurally simplified RNA and the cognate RNA-binding protein, the latter having apparently acquired an additional domain to compensate for the lack of a binding site(s) on the RNA. PMID- 11262400 TI - Pax3 is essential for skeletal myogenesis and the expression of Six1 and Eya2. AB - Pax3 is a paired box transcription factor expressed during somitogenesis that has been implicated in initiating the expression of the myogenic regulatory factors during myogenesis. We find that Pax3 is necessary and sufficient to induce myogenesis in pluripotent stem cells. Pax3 induced the expression of the transcription factor Six1, its cofactor Eya2, and the transcription factor Mox1 prior to inducing the expression of MyoD and myogenin. Overexpression of a dominant negative Pax3, engineered by fusing the active transcriptional repression domain of mouse EN-2 in place of the Pax3 transcriptional activation domain, completely abolished skeletal myogenesis without inhibiting cardiogenesis. Expression of the dominant negative Pax3 resulted in a loss of expression of Six1, Eya2, and endogenous Pax3 as well as a down-regulation in the expression of Mox1. No effect was found on the expression of Gli2. These results indicate that Pax3 activity is essential for skeletal muscle development, the expression of Six1 and Eya2, and is involved in regulating its own expression. In summary, the combined approach of expressing both a wild type and dominant negative transcription factor in stem cells has identified a cascade of transcriptional events controlled by Pax3 that are necessary and sufficient for skeletal myogenesis. PMID- 11262402 TI - The two phosphofructokinase gene products of Entamoeba histolytica. AB - Two phosphofructokinase genes have been described previously in Entamoeba histolytica. The product of the larger of the two genes codes for a 60-kDa protein that has been described previously as a pyrophosphate (PP(i))-dependent enzyme, and the product of the second, coding for a 48-kDa protein, has been previously reported to be a PP(i)-dependent enzyme with extremely low specific activity. Here it is found that the 48-kDa protein is not a PP(i)-dependent enzyme but a highly active ATP-requiring enzyme (k(cat) = 250 s(-)1) that binds the cosubstrate fructose 6-phosphate (Fru-6-P) with relatively low affinity. This enzyme exists in concentration- and ATP-dependent tetrameric active and dimeric inactive states. Activation is achieved in the presence of nucleoside triphosphates, ADP, and PP(i), but not by AMP, P(i), or the second substrate Fru 6-P. Activation by ATP is facilitated by conditions of molecular crowding. Divalent cations are not required, and no phosphoryl transfer occurs during activation. Kinetics of the activated enzyme show cooperativity with Fru-6-P (Fru 6-P(0.5) = 3.8 mm) and inhibition by high ATP and phosphoenolpyruvate. The enzyme is active without prior activation in extracts of E. histolytica. The level of mRNA, the amount of enzyme protein, and the enzyme activity of the 48-kDa enzyme are about one-tenth that of the 60-kDa enzyme in extracts of E. histolytica trophozoites. PMID- 11262401 TI - Effects of stimulation of AMP-activated protein kinase on insulin-like growth factor 1- and epidermal growth factor-dependent extracellular signal-regulated kinase pathway. AB - AMP-activated protein kinase (AMPK) is tightly regulated by the cellular AMP:ATP ratio and plays a central role in the regulation of energy homeostasis. Previously, AMPK was reported to phosphorylate serine 621 of Raf-1 in vitro. In the present study, we investigated a possible role of AMPK in extracellular signal-regulated kinase (Erk) cascades, using 5-aminoimidazole-4-carboxamide-1 beta-d-ribofuranoside (AICAR), a cell-permeable activator of AMPK and antisense RNA experiments. Activation of AMPK by AICAR in NIH-3T3 cells resulted in drastic inhibitions of Ras, Raf-1, and Erk activation induced by insulin-like growth factor 1 (IGF-1). Expression of an antisense RNA for the AMPK catalytic subunit decreased the AMPK activity and significantly diminished the AICAR effect on IGF 1-induced Ras activation and the subsequent Erk activation, indicating that its effect is indeed mediated by AMPK. Phosphorylation of Raf-1 serine 621, however, was not involved in AMPK-mediated inhibition of Erk cascades. In contrast to IGF 1, AICAR did not block epidermal growth factor (EGF)-dependent Raf-1 and Erk activation, but our results demonstrated that multiple Raf-1 upstream pathways induced by EGF were differentially affected by AICAR: inhibition of Ras activation and simultaneous induction of Ras-independent Raf activation. The activities of IGF-1 and EGF receptor were not affected by AICAR. Taken together, our results suggest that AMPK differentially regulate Erk cascades by inhibiting Ras activation or stimulating the Ras-independent pathway in response to the varying energy status of the cell. PMID- 11262403 TI - VP4 differentially regulates TRAF2 signaling, disengaging JNK activation while directing NF-kappa B to effect rotavirus-specific cellular responses. AB - Rotaviruses rapidly activate NF-kappaB and induce the secretion of selected chemokines after infection. The ability of rotavirus particles lacking genomic RNA to activate NF-kappaB suggested that rotavirus proteins direct cell signaling responses. We identified conserved TNFR-associated factor (TRAF) binding motifs within the rotavirus capsid protein VP4 and its N-terminal VP8* cleavage product. TRAFs (-1, -2, and -3) are bound by the rhesus rotavirus VP8* protein through three discrete TRAF binding domains. Expression of VP4 or VP8* from rhesus or human rotaviruses induced a 5-7-fold increase in NF-kappaB activity and synergistically enhanced TRAF2-mediated NF-kappaB activation. Mutagenesis of VP8* TRAF binding motifs abolished VP8* binding to TRAFs and the ability of the protein to activate NF-kappaB. Expression of pathway-specific dominant negative (DN) inhibitors DN-TRAF2 or DN-NF-kappaB-inducing kinase also abolished VP8*-, VP4-, or rotavirus-mediated NF-kappaB activation. These findings demonstrate that rotavirus primarily activates NF-kappaB through a TRAF2-NF-kappaB-inducing kinase signaling pathway and that VP4 and VP8* proteins direct pathway activation through interactions with cellular TRAFs. In contrast, transcriptional responses from AP-1 reporters were inhibited 5-fold by VP8* and were not activated by rotavirus infection, suggesting the differential regulation of TRAF2 signaling responses by VP8*. VP8* blocked JNK activation directed by TRAF2 or TRAF5 but had no effect on JNK activation directed by TRAF6 or MEKK1. This establishes that fully cytoplasmic rotaviruses selectively engage signaling pathways, which regulate cellular transcriptional responses. These findings also demonstrate that TRAF2 interactions can disengage JNK signaling from NF-kappaB activation and thereby provide a new means for TRAF2 interactions to determine pathway-specific responses. PMID- 11262404 TI - Increased sensitivity of transforming growth factor (TGF) beta 1 null cells to alkylating agents reveals a novel link between TGFbeta signaling and O(6) methylguanine methyltransferase promoter hypermethylation. AB - Inactivation of the transforming growth factor beta (TGFbeta)-signaling pathway and gene silencing through hypermethylation of promoter CpG islands are two frequent alterations in human and experimental cancers. Here we report that nonneoplastic TGFbeta1-/- keratinocyte cell lines exhibit increased sensitivity to cell killing by alkylating agents, and this is due to lack of expression of the DNA repair enzyme O(6)-methylguanine DNA methyltransferase (MGMT). In TGFbeta1-/- but not TGFbeta1+/- cell lines, the CpG dinucleotides in the MGMT promoter are hypermethylated, as measured by restriction enzyme analysis and methylation specific polymerase chain reaction. In one unstable TGFbeta1+/- cell line, loss of the wild type TGFbeta1 allele correlates with the appearance of methylation in the MGMT promoter. Bisulfite sequencing shows that in the KO3 TGFbeta1-/- cell line nearly all of the 28 CpG sites in the MGMT promoter 475 base pairs upstream of the start site of transcription are methylated, whereas most are unmethylated in the H1 TGFbeta1+/- line. Treatment of the TGFbeta1-/- cell lines with 5-azacytidine causes reexpression of MGMT mRNA and demethylation of CpG islands in the promoter. Analysis of the time course of methylation using methylation-specific polymerase chain reaction shows a lack of methylation in primary TGFbeta1-/- keratinocytes and increasing methylation with passage number of immortalized clones. Subcloning of early passage clones reveals a remarkable heterogeneity and instability of the methylation state in the TGFbeta1-/- keratinocytes. Thus, the TGFbeta1-/- genotype does not directly regulate MGMT methylation but predisposes cells to immortalization-associated MGMT hypermethylation. PMID- 11262405 TI - Mass spectral analysis of protein-based radicals using DBNBS. Nonradical adduct formation versus spin trapping. AB - Protein-based radicals generated in the reaction of ferricytochrome c (cyt c) with H(2)O(2) were investigated by electrospray mass spectrometry (ESI-MS) using 3,5-dibromo-4-nitrosobenzenesulfonate (DBNBS). Up to four DBNBS-cyt c adducts were observed in the mass spectra. However, by varying the reaction conditions (0 5 molar equivalents of H(2)O(2) and substituting cyt c with its cyanide adduct which is resistant to peroxidation), noncovalent DBNBS adduct formation was inferred. Nonetheless, optical difference spectra revealed the presence of a small fraction of covalently trapped DBNBS. To probe the nature of the noncovalent DBNBS adducts, the less basic proteins, metmyoglobin (Mb) and alpha lactalbumin, were substituted for cyt c in the cyt c/H(2)O(2)/DBNBS reaction. A maximum of two DBNBS adducts were observed in the mass spectra of the products of the Mb/H(2)O(2)/DBNBS reactions, whereas no adducts were detected following alpha lactalbumin/H(2)O(2)/DBNBS incubation, which is consistent with adduct formation via spin trapping only. Titration with DBNBS at pH 2.0 yielded noncovalent DBNBS cyt c adducts and induced folding of acid-denatured cyt c, as monitored by ESI-MS and optical spectroscopy, respectively. Thus, the noncovalent DBNBS-cyt c mass adducts observed are assigned to ion pair formation occurring between the negatively charged sulfonate group on DBNBS and positively charged surface residues on cyt c. The results reveal the pitfalls inherent in using mass spectral data with negatively charged spin traps such as DBNBS to identify sites of radical formation on basic proteins such as cyt c. PMID- 11262406 TI - gp130 plays a critical role in pressure overload-induced cardiac hypertrophy. AB - gp130, a common receptor for the interleukin 6 family, plays pivotal roles in growth and survival of cardiac myocytes. In the present study, we examined the role of gp130 in pressure overload-induced cardiac hypertrophy using transgenic (TG) mice, which express a dominant negative mutant of gp130 in the heart under the control of alpha myosin heavy chain promoter. TG mice were apparently healthy and fertile. There were no differences in body weight and heart weight between TG mice and littermate wild type (WT) mice. Pressure overload-induced increases in the heart weight/body weight ratio, ventricular wall thickness, and cross sectional areas of cardiac myocytes were significantly smaller in TG mice than in WT mice. Northern blot analysis revealed that pressure overload-induced up regulation of brain natriuretic factor gene and down-regulation of sarcoplasmic reticulum Ca(2+) ATPase 2 gene were attenuated in TG mice. Pressure overload activated ERKs and STAT3 in the heart of WT mice, whereas pressure overload induced activation of STAT3, but not of ERKs, was suppressed in TG mice. These results suggest that gp130 plays a critical role in pressure overload-induced cardiac hypertrophy possibly through the STAT3 pathway. PMID- 11262407 TI - The active N-terminal region of p67phox. Structure at 1.8 A resolution and biochemical characterizations of the A128V mutant implicated in chronic granulomatous disease. AB - Upon activation, the NADPH oxidase from neutrophils produces superoxide anions in response to microbial infection. This enzymatic complex is activated by association of its cytosolic factors p67(phox), p47(phox), and the small G protein Rac with a membrane-associated flavocytochrome b(558). Here we report the crystal structure of the active N-terminal fragment of p67(phox) at 1.8 A resolution, as well as functional studies of p67(phox) mutants. This N-terminal region (residues 1-213) consists mainly of four TPR (tetratricopeptide repeat) motifs in which the C terminus folds back into a hydrophobic groove formed by the TPR domain. The structure is very similar to that of the inactive truncated form of p67(phox) bound to the small G protein Rac previously reported, but differs by the presence of a short C-terminal helix (residues 187-193) that might be part of the activation domain. All p67(phox) mutants responsible for Chronic Granulomatous Disease (CGD), a severe defect of NADPH oxidase function, are localized in the N-terminal region. We investigated two CGD mutations, G78E and A128V. Surprisingly, the A128V CGD mutant is able to fully activate the NADPH oxidase in vitro at 25 degrees C. However, this point mutation represents a temperature-sensitive defect in p67(phox) that explains its phenotype at physiological temperature. PMID- 11262408 TI - Roles of cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1. AB - Gab-1 is a multiple docking protein that is tyrosine phosphorylated by receptor tyrosine kinases such as c-Met, hepatocyte growth factor/scatter factor receptor, and epidermal growth factor receptor. We have now demonstrated that cell-cell adhesion also induces marked tyrosine phosphorylation of Gab-1 and that disruption of cell-cell adhesion results in its dephosphorylation. An anti-E cadherin antibody decreased cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas the expression of E-cadherin specifically induced tyrosine phosphorylation of Gab-1. A relatively selective inhibitor of Src family kinases reduced cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas expression of a dominant-negative mutant of Csk increased it. Disruption of cell cell adhesion, which reduced tyrosine phosphorylation of Gab-1, also reduced the activation of mitogen-activated protein kinase and Akt in response to cell-cell adhesion. These results indicate that E-cadherin-mediated cell-cell adhesion induces tyrosine phosphorylation by a Src family kinase of Gab-1, thereby regulating the activation of Ras/MAP kinase and phosphatidylinositol 3-kinase/Akt cascades. PMID- 11262409 TI - Effects of a cardiomyopathy-causing troponin t mutation on thin filament function and structure. AB - Familial hypertrophic cardiomyopathy (FHC) is caused by missense or premature truncation mutations in proteins of the cardiac contractile apparatus. Mutant proteins are incorporated into the thin filament or thick filament and eventually produce cardiomyopathy. However, it has been unclear how the several, genetically identified defects in protein structure translate into impaired protein and muscle function. We have studied the basis of FHC caused by premature truncation of the most frequently implicated thin filament target, troponin T. Electron microscope observations showed that the thin filament undergoes normal structural changes in response to Ca(2+) binding. On the other hand, solution studies showed that the mutation alters and destabilizes troponin binding to the thin filament to different extents in different regulatory states, thereby affecting the transitions among states that regulate myosin binding and muscle contraction. Development of hypertrophic cardiomyopathy can thus be traced to a defect in the primary mechanism controlling cardiac contraction, switching between different conformations of the thin filament. PMID- 11262410 TI - Repression of dpp targets by binding of brinker to mad sites. AB - Signaling by decapentaplegic (Dpp), a Drosophila member of the transforming growth factor (TGF) beta superfamily of growth factors, has recently been shown to activate targets such as vestigial (vg) indirectly through negative regulation of brinker (brk). Here we show that the Brk protein functions as a repressor by binding to Dpp response elements. The Brk DNA binding activity was localized to an amino-terminal region containing a putative homeodomain. Brk bound to a Dpp response element of the Ultrabithorax (Ubx) midgut enhancer at a sequence that overlaps a binding site for the Smad protein, Mothers Against Dpp (Mad). Furthermore, Brk was able to compete with Mad for occupancy of this binding site. This recognition of overlapping binding sites provides a potential explanation for why the G/C-rich Mad binding site consensus differs the Smad3/Smad4 binding site consensus. We also found that the Dpp response element from Ubx was more sensitive than the vg quadrant enhancer to repression by Brk. This difference correlates with short-range activation of Ubx by Dpp in the visceral mesoderm, whereas vg exhibits a long-range response to Dpp in the wing imaginal disc, indicating that Brk binding sites may play a critical role in limiting thresholds for activation by Dpp. Finally, we provide evidence that Brk is capable of functioning as an active repressor. Thus, whereas Brk and Mad compete for regulation of Ubx and vg, Brk may regulate other Dpp targets without direct involvement of Mad. PMID- 11262411 TI - Phytotoxic protein PcF, purification, characterization, and cDNA sequencing of a novel hydroxyproline-containing factor secreted by the strawberry pathogen Phytophthora cactorum. AB - A novel protein factor, named PcF, has been isolated from the culture filtrate of Phytophthora cactorum strain P381 using a highly sensitive leaf necrosis bioassay with tomato seedlings. Isolated PcF protein alone induced leaf necrosis on its host strawberry plant. The primary structure and cDNA sequence of this novel phytotoxic protein was determined, and BLAST searches of Swiss-Prot, EMBL, and GenBank(TM)/EBI data banks showed that PcF shared no significant homology with other known sequences. The 52-residue PcF protein, which contains a 4 hydroxyproline residue along with three S-S bridges, exhibits a high content of acidic sidechains, accounting for its isoelectric point of 4.4. The molecular mass of isolated PcF is 5,622 +/- 0.5 Da as determined by mass spectrometry and matches that calculated from the deduced amino acid sequence with cDNA sequencing. The cDNA sequence indicates that PcF is first produced as a larger precursor, comprising an additional N-terminal, 21-residue secretory signal peptide. Maturation of this protein involves the hydroxylation of proline 49, a feature that is unique among other known secreted fungal phytopathogenic proteins. PMID- 11262412 TI - Ubiquinol:cytochrome c oxidoreductase (complex III). Effect of inhibitors on cytochrome b reduction in submitochondrial particles and the role of ubiquinone in complex III. AB - Two sets of studies have been reported on the electron transfer pathway of complex III in bovine heart submitochondrial particles (SMP). 1) In the presence of myxothiazol, MOA-stilbene, stigmatellin, or of antimycin added to SMP pretreated with ascorbate and KCN to reduce the high potential components (iron sulfur protein (ISP) and cytochrome c(1)) of complex III, addition of succinate reduced heme b(H) followed by a slow and partial reduction of heme b(L). Similar results were obtained when SMP were treated only with KCN or NaN(3), reagents that inhibit cytochrome oxidase, not complex III. The average initial rate of b(H) reduction under these conditions was about 25-30% of the rate of b reduction by succinate in antimycin-treated SMP, where both b(H) and b(L) were concomitantly reduced. These results have been discussed in relation to the Q cycle hypothesis and the effect of the redox state of ISP/c(1) on cytochrome b reduction by succinate. 2) Reverse electron transfer from ISP reduced with ascorbate plus phenazine methosulfate to cytochrome b was studied in SMP, ubiquinone (Q)-depleted SMP containing 3sigma(I); R = 0.0095, wR = 0.0065, Deltarho(min) = -0.91 e A(-3), Deltarho(max) = 0.65 e A(-3)]. Defects in the O and K atomic positions were found. (1) is a semiconductor in the temperature range 4-300 K, whereas the well studied and closely related colourless transparent crystals KTa(+5)O(3) (2) are dielectric. Differences in the properties of (1) and (2) are assumed to be connected with the existence of Ta dumb-bells statistically distributed into the KTaO(3) matrix. PMID- 11262431 TI - Systematic ab initio study of the compressibility of silicate garnets. AB - The structural properties of the silicate garnets andradite, Ca(3)Fe(2)Si(3)O(12), uvarovite, Ca(3)Cr(2)Si(3)O(12), knorringite, Mg(3)Cr(2)Si(3)O(12), goldmanite, Ca(3)V(2)Si(3)O(12), blythite, Mn(2+)(3)Mn(3+)(2)Si(3)O(12), skiagite, Fe(2+)(3)Fe(3+)(2)Si(3)O(12), calderite, Mn(2+)(3)Fe(3+)(2)Si(3)O(12), and khoharite, Mg(3)Fe(3+)(2)Si(3)O(12), have been investigated with a quantum-mechanical model as a function of applied pressure. The study has been performed with the density functional theory code CASTEP, which uses pseudopotentials and a plane-wave basis set. All structural parameters have been optimized. The calculated static geometries (cell parameters, internal coordinates of atoms and bond lengths), bulk moduli and their pressure derivatives are in good agreement with the experimental data available. Predictions are made for those cases where no experimental data have been reported. The data clearly indicate that the elastic properties of all silicate garnets are dominated by the compressibility of the dodecahedral site. The compression mechanism is found to be based on a bending of the angle between the centers of the SiO(4) tetrahedra and the adjacent octahedra, as in the aluminosilicate garnets. An analysis of the relationship between ionic radii of the cations and the compressibility of silicate garnets is presented. PMID- 11262432 TI - Structure determination of two intercalated compounds VOPO4.(CH2)4O and VOPO4.OH (CH2)2-O-(CH2)2-OH; synchrotron powder diffraction and molecular modelling. AB - The crystal structures of two intercalated compounds have been determined using a combination of synchrotron powder diffraction and molecular mechanics simulations: (1) vanadyl phosphate intercalated with tetrahydrofuran, VOPO(4).(CH(2))(4)O, and (2) vanadyl phosphate intercalated with diethylene glycol, VOPO(4).HO(CH(2))(2)O(CH(2))(2)OH. Both intercalates preserve the tetragonal space group P4/n, as found in the host structure VOPO(4).2H(2)O. (1): a = 6.208, c = 8.930 A, Z = 2, D(x) = 2.51 g cm(-3); (2): a = 6.223, c = 11.417 A, Z = 2, D(x) = 2.66 g cm(-3). Both intercalates exhibit the same type of orientational disorder in the arrangement of guest molecules, as observed in the same host compound intercalated with water. These two intercalates also exhibit, rather surprisingly, perfect ordering in layer stacking without the displacement disorder, characteristic of many intercalated layered structures. Thanks to this regularity in the arrangement of guests and layers, synchrotron powder diffraction could be used in the present structure determination. The present results also enabled the analysis of the effect of geometrical parameters characterizing the mutual host-guest complementarity and the effect of host-guest and guest-guest interaction on the crystal packing of intercalates. PMID- 11262433 TI - Ab initio structure determination of monoclinic 2,2-dihydroxymethylbutanoic acid from synchrotron radiation powder diffraction data: combined use of direct methods and the Monte Carlo method. AB - The crystal structure of 2,2-dihydroxymethylbutanoic acid (C(6)H(12)O(4)) in monoclinic form has been determined ab initio from synchrotron radiation powder diffraction data. Two O and five C atoms were first derived by direct methods. Two missing O atoms and one C atom were found by the Monte Carlo method without applying constraint to their relative positions. Positional and isotropic displacement parameters of these non-H atoms were refined by the Rietveld method. Molecules are linked by hydrogen bonds and they make sheet-like networks running parallel to the (010) plane. The Monte Carlo method is demonstrated to be a powerful tool for finding missing atoms in partially solved structure. PMID- 11262434 TI - Patterns of soft C-H...O hydrogen bonding in diaryl sulfones. AB - In bis(4-tolyl) sulfone, C(14)H(14)O(2)S (1), 2,5,4'-trimethyldiphenyl sulfone, C(15)H(16)O(2)S (2), and 4-chlorodiphenyl sulfone, C(12)H(9)ClO(2)S (3), the molecules are linked by soft C-H...O hydrogen bonds into three different types of one-dimensional aggregate: simple chains in (1), molecular ladders in (2) and chains of fused rings in (3). In each of 3,4-dimethyl-4'-chlorodiphenyl sulfone, C(14)H(13)ClO(2)S (4), and 2,5-dimethyldiphenyl sulfone, C(14)H(14)O(2)S (5), the C-H...O hydrogen bonds link the molecules into two different types of two dimensional sheet, based on a (4,4) net in (4) and a (3,6) net in (5). The patterns of soft C-H...O hydrogen bonds in (1)-(5) are compared with those in other diaryl sulfones, mainly retrieved from the Cambridge Structural Database, whose substitution patterns preclude the formation of hard hydrogen bonds. Observed aggregation modes range from the formation of no C-H...O hydrogen bonds at all, via finite (zero-dimensional) arrays through one-, two- and three dimensional systems. PMID- 11262435 TI - Salts of 3,5-dinitrobenzoic acid with organic diamines: hydrogen-bonded supramolecular structures in one, two and three dimensions. AB - The trigonally trisubstituted carboxylic acid 3,5-dinitrobenzoic acid, (O(2)N)(2)C(6)H(3)COOH, forms 2:1 salts with a range of organic diamines L, with the general composition [LH(2)](2+) x [[(O(2)N)(2)C(6)H(3)COO](-)](2). When L is a bis-tertiary amine the hard N-H...O hydrogen bonds generate finite three component aggregates, anion...cation...anion, and these aggregates are further linked by soft C-H...O hydrogen bonds to form one-dimensional molecular ladders when L is N,N,N',N"-tetramethyl-1,2-diaminoethane and chains of rings when L is 4,4'-dipyridylethane or 4,4'-dipyridylethene; two-dimensional sheets are formed when L is 1,4-diazabicyclo[2.2.2]octane and a three-dimensional framework is formed when L is N,N'-dimethylpiperazine. When L is the bis-secondary amine piperazine, the hard N-H...O and soft C-H...O hydrogen bonds each generate continuous motifs in the form of distinct chains of rings, the combination of which generates sheets, while when L is the bis-primary amine 1,2-diaminoethane the hard N-H...O hydrogen bonds alone generate a three-dimensional framework. PMID- 11262436 TI - Structures of furanosides: geometrical analysis of low-temperature X-ray and neutron crystal structures of five crystalline methyl pentofuranosides. AB - Crystal structures of all five crystalline methyl D-pentofuranosides, methyl alpha-D-arabinofuranoside (1), methyl beta-D-arabinofuranoside (2), methyl alpha D-lyxofuranoside (3), methyl beta-D-ribofuranoside (4) and methyl alpha-D xylofuranoside (5) have been determined by means of cryogenic X-ray and neutron crystallography. The neutron diffraction experiments provide accurate, unbiased H atom positions which are especially important because of the critical role of hydrogen bonding in these systems. This paper summarizes the geometrical and conformational parameters of the structures of all five crystalline methyl pentofuranosides, several of them reported here for the first time. The methyl pentofuranoside structures are compared with the structures of the five crystalline methyl hexopyranosides for which accurate X-ray and neutron structures have been determined. Unlike the methyl hexopyranosides, which crystallize exclusively in the C(1) chair conformation, the five crystalline methyl pentofuranosides represent a very wide range of ring conformations. PMID- 11262437 TI - Comparison of nebulized particle size distribution with Malvern laser diffraction analyzer versus Andersen cascade impactor and low-flow Marple personal cascade impactor. AB - Particle size of nebulized aerosols can be measured directly using laser diffraction or by evaluating aerodynamic properties by cascade impaction. As of today, there are no generally accepted standards for measuring particle size distribution from nebulizers. Laser diffraction has been questioned because of potential evaporative losses of the small particles at the edge of the plume, causing an apparent shift in the particle size distribution and thus a larger mass median diameter (MMD). When particle-sizing wet aerosols, cascade impaction may give rise to an apparent shift in the distribution, resulting in a smaller mass median aerodynamic diameter (MMAD) due to evaporative losses of aerosol droplets as they enter the impactor at ambient temperature. The modified low-flow Marple 296 Personal Cascade Impactor (MPCI) is currently being proposed as the European standard for wet aerosol analysis to minimize evaporative losses during sampling. The present study compared the particle size distribution of salbutamol and sodium cromoglycate aerosols nebulized by the Pari LC Star, using laser diffraction (Malvern Mastersizer X; MMX) and cascade impaction (Andersen Cascade Impactor [ACI] and the commercially available MPCI), which was either at ambient temperature or cooled to the nebulized aerosol temperature (10 degrees C). MMDs obtained with the MMX were virtually identical to the MMADs measured with both impactors when cooled with no significant differences in geometric standard deviation (sigma(g)). When the impactors were operated at ambient temperature, MMADs were smaller (18 to 30%) with a significantly larger sigma(g) (p < 0.05) compared to the MMX. These findings suggest that droplet distribution data for wet aerosol where evaporation process has not been minimized must be viewed with caution. There was no evidence suggesting a significant evaporative loss of small droplets from the edge of the plume during laser particle sizing. The MPCI does not minimize evaporative losses of aerosol particles during sampling. PMID- 11262438 TI - The distribution of inhaled particles in aerosol measurements of pulmonary airspace size. AB - When the deposition of aerosol boluses is used to estimate mean pulmonary airspace size, an implicit assumption is made that the inhaled particles are distributed uniformly among normal and diseased lung regions. This assumption was examined in a series of dogs in which emphysema was experimentally induced by exposure to papain. After the experimental disease had developed for several weeks, boluses of fluorescent particles were inhaled, using a breathing pattern similar to that used for aerosol measurements of airspace size. The lungs were then excised and 18-20 tissue blocks were obtained from each lung. A section from each tissue block was analyzed to determine the mean liner intercept (Lm), which was considered as an index of lung injury. In the same sections, the density of particles was determined by counting particles in a number of microscopic fields and dividing the particle count by the number of fields sampled. Correlation analysis generally revealed a negative correlation of particle density with Lm, indicating fewer particles being delivered to diseased regions. One lung, however, showed a positive correlation between particle density and Lm. Correction for the fractional deposition of aerosol in the lung regions weakened but did not reverse the relationship between particle density and Lm. A model calculation of the effect of the observed nonuniform distribution of aerosol on the determination of airspace size found a negligible effect of uneven ventilation on mean airspace size determination in this experimental preparation. PMID- 11262439 TI - Jet and ultrasonic nebulization of single chain urokinase plasminogen activator (scu-PA). AB - Recent studies have indicated that the deposition of intra-alveolar fibrin may play a central role in the pathogenesis of acute respiratory distress syndrome (ARDS). Our aim was to study whether the indigenous fibrinolytic agent (urokinase) normally present in the alveoli can be administered locally by nebulization in a recombinant zymogen form as single chain urokinase plasminogen activator (scu-PA). We aimed to characterize the particle size distribution, drug output, and enzymatic activity of scu-PA after nebulization with a Ventstream jet nebulizer (Medic-Aid, Bognor Regis, UK) and a Syst'AM DP-100 ultrasonic nebulizer (Pulmolink, Kent, UK). The particle size distribution was measured with a laser diffraction method and the drug output was determined by collection on filters. The amount of protein on the filters was determined with the Lowry method, and the enzymatic activity after nebulization was measured with a microtiter fibrin plate assay. The mass median diameter (MMD) of the scu-PA aerosol generated with the ultrasonic nebulizer was 3.69 (3.53-3.83) microm and with the jet nebulizer 2.96 (2.91-3.03) microm (p < 0.001). The drug output from the two nebulizers did not differ between nebulizers (p = 0.054). Fibrinolytically active scu-PA was generated with both nebulizers, but in contrast to jet nebulization, ultrasonic nebulization caused partial inactivation of scu-PA (p < 0.001). In conclusion, nebulization of scu-PA with the jet nebulizer is superior to ultrasonic nebulization in terms of particle size distribution and preservation of fibrinolytic activity. PMID- 11262440 TI - What do epidemiologic findings tell us about health effects of environmental aerosols? AB - In the last 10 years there has been an abundance of new epidemiological studies on health effects of particulate air pollution. The overall evidence suggests that fine particulate pollution can be an important risk factor for cardiopulmonary disease. Long-term, repeated exposure to fine particulate air pollution may increase the risk of chronic respiratory disease and the risk of cardiopulmonary mortality. Short-term exposures exacerbate existing cardiovascular and pulmonary disease and increase the risk of becoming symptomatic, requiring medical attention, or even dying. This paper outlines the results of the basic epidemiologic studies and briefly reviews and discusses recent studies that have looked at specific physiologic health endpoints in addition to lung function. A few recent, mostly exploratory pilot studies, have observed particulate pollution associations with blood plasma viscosity, heart rate, heart rate variability, and indicators of bone marrow stimulation. A systemic response to particulate-related pulmonary inflammation remains somewhat speculative. The epidemiologic evidence, nevertheless, seems consistent with the hypothesis that particle-induced pulmonary inflammation, cytokine release, and altered cardiac autonomic function may be part of the pathophysiological mechanisms or pathways linking particulate pollution with cardiopulmonary disease. PMID- 11262441 TI - Acute health effects of ambient air pollution: the ultrafine particle hypothesis. AB - A strong and consistent association has been observed between adjusted mortality rates and ambient particle concentration. The strongest associations are seen for respiratory and cardiac deaths, particularly among the elderly. Particulate air pollution is also associated with asthma exacerbations, increased respiratory symptoms, decreased lung function, increased medication use, and increased hospital admissions. The U.S. Environmental Protection Agency (EPA) has recently promulgated a new national ambient air quality standard for fine particles, and yet the mechanisms for health effects at such low particle mass concentrations remain unclear. Hypotheses to identify the responsible particles have focused on particle acidity, particle content of transition metals, bioaerosols, and ultrafine particles. Because ultrafine particles are efficiently deposited in the respiratory tract and may be important in initiating airway inflammation, we have initiated clinical studies with ultrafine carbon particles in healthy subjects. These studies examine the role of ultrafines in: (1) the induction of airway inflammation; (2) expression of leukocyte and endothelial adhesion molecules in blood; (3) the alteration of blood coagulability; and (4) alteration in cardiac electrical activity. These events could lead to exacerbation of underlying cardiorespiratory disease. For example, airway inflammation may activate endothelium and circulating leukocytes, and induce a systemic acute phase response with transient hypercoagulability; this could explain the epidemiologic linkages between pollutant exposures and cardiovascular events. These approaches should be useful in identifying mechanisms for pollutant-induced respiratory and systemic effects, and in providing data for determining appropriate air quality standards. PMID- 11262442 TI - Characterization of dendritic cell populations in the respiratory tract. AB - Research in a variety of experimental animal species and in humans has identified dendritic cells (DCs) as the principal resident antigen presenting cells in respiratory tract tissues. The two major populations of respiratory tract DC (RTDC) comprise those found in the parenchymal tissues of the peripheral lung and in the epithelium of the conducting airways, in which they are distributed as a contiguous network comparable to the langerhans cells (LC) of the epidermis. Under steady state conditions, the airway DC population turns over every 36-48 h, whereas those in the lung parenchyma display a half-life of approximately 7 days. However, under conditions of local stress (e.g., inflammatory challenge), the turnover of these RTDC populations further accelerates, reflecting their important role in local antigen surveillance. In the resting state, they are specialized for the efficient endocytosis and processing of internalized antigens, but lack the capacity to efficiently present antigen to T-cells until they receive appropriate cytokine signals (especially GM-CSF); responsiveness to the latter is inhibited by nitric oxide, in particular from adjacent lung tissue macrophages. Our most recent findings indicate that the "default" function of resting RTDC involves selective priming for Th2 responses, and induction of optimal Th1 responses requires exposure to GM-CSF together with TNFalpha or CD40L. PMID- 11262444 TI - Validating deposition models in disease: what is needed? AB - To develop theoretical deposition models, assumptions are introduced to make the models computationally affordable. For this reason, experimental (in vivo) validation of such models is needed to give confidence to the assumptions being made. However, for an in vivo deposition experiment to be considered useful for validation of a model, a number of parameters must be measured in the experiment for input to the model. Ideally, these parameters would include time-dependent breathing flow rates during aerosol exposure, properties of the inhaled aerosol as a function of time during the breath (including particle size distribution, aerosol mass fraction, as well as hygroscopic properties, inhaled temperature and humidity if hygroscopicity is important), in addition to anatomical regional deposition data and detailed lung geometry measurements. Furthermore, because of the dependence of extrathoracic filtering on the inlet conditions at the mouth and the complexity of modeling deposition in this region, experimental data on the filtering properties of the mouth-throat are needed. Although some of the above parameters are impractical to measure with current experimental techniques, it would greatly aid the development of deposition models if as many of these parameters as possible were measured in future in vivo deposition experiments. Data exemplifying the importance of measuring the above parameters is discussed. PMID- 11262443 TI - Interaction of alveolar macrophages with inhaled mineral particulates. AB - Pulmonary disorders triggered by inhalation of occupational and environmental mineral particulates can be endpoints of a chronic inflammatory process in which alveolar macrophages (AMs), as a first line of defense, play a crucial role. The biological processes involved in particulate-induced activation of AMs include indirect or direct interactions of particulates with the cell membrane, subsequent stimulation of signal transduction pathways, and activation of gene transcription. Depending on the nature of particulate involved, particulate induced activation of AMs has been shown to result in the release of potent mediators, such as reactive oxygen and nitrogen species, cytokines, eicosanoids, and growth factors. The prolonged and enhanced production of such effector molecules may result in a complex cascade of events that can contribute to the development of pulmonary disorders. This paper will give a short review of the present knowledge of AM interaction with inhaled mineral particulates and of the possible implications these interactions may have in the development of pulmonary disorders. PMID- 11262445 TI - Physiological and pathological considerations for aerosol deposition: expiration and models of deposition. AB - Theoretical models are often used to predict fractional and regional deposition of inhaled particles in the respiratory tract. The distribution of particle diameters in the aerosol, airway geometry, breathing pattern, and local flow profiles are major determinants of deposition in the lung. However, most models predicting deposition consider airway geometry to be fixed and concentrate on inspiratory events in their calculations. When particle losses during expiration are estimated, inspiratory and expiratory flow patterns and airspace geometry are usually considered to be similar with similar effects on deposition. The theme of this presentation will be the analysis of events during expiration that influence particle deposition. In the normal lung, during quiet breathing, experiments performed on excised lungs have suggested that convective forces may be different between inspiration and expiration that significantly affect deposition. Bennett and Smaldone, in excised dog lungs, by regulating the duty cycle of tidal breathing found that more particles deposited during inspiration than expiration and that the effects were density dependent. In human subjects with obstructive lung disease, the situation is reversed. Major differences in large airway geometry between inspiration and expiration can occur with each tidal breath. Once the FEV(1) decreases to about 60% of the FVC, flow-limiting segments (FLS) are known to form in central airways. Large pressure drops can occur over short lengths of airway indicating disturbed regions of convective streamlines that are not present during inspiration. Using radiolabeled monodisperse particles, Smaldone and Messina have determined that FLS can be a major determinant of deposition in central airways. Theoretical predictive models of particle deposition and clearance should consider inspiratory and expiratory differences in airway physiology in health and disease. PMID- 11262446 TI - Rationale for the choice of an aerosol delivery system. AB - The choice of an aerosol delivery system depends on numerous factors such as the drug itself, the characteristics of the aerosol generator, the patient and his or her disease, the physician, and the clinical setting, notably an emergency situation or not. Some rules always apply: an ultrasonic nebulizer should not be used to aerosolize a drug suspension; whenever possible, the same type of aerosol generator should be used for all inhaled medications received by a given patient; for outpatients, education is a major factor to ensure treatment efficacy. When the deposition of the aerosolized drug is aimed at the terminal respiratory units, nebulizers that generate micronic aerosols should be chosen. When the deposition of the aerosolized drug is aimed at the conducting airways, the metered dose inhaler (MDI) is the first choice. However, the MDI is often ill used, notably in children and elderly people. Therefore, other inhalation devices have been developed: spacers, dry-powder inhalers, breath-actuated MDIs and, more recently, piezo-electric devices. They have been shown to increase lung deposition of drugs in poor coordinators but they all have limitations, which may affect their clinical efficacy. These limitations include the cumbersome dimensions of spacers, the dependency of lung deposition of dry powders on the inspiratory flow rate, the need for reformulation of breath-actuated or not MDIs with CFC-free gases. Nebulization of drugs should be considered only when no portable device is available for the considered drug, or in case of failure of other forms of aerosol administration. PMID- 11262448 TI - Patterns of ambulatory care use for gynecologic conditions: A national study. AB - OBJECTIVE: This study was undertaken to describe the site of ambulatory care visits for gynecologic conditions in the United States and to identify patient factors associated with the site of care for these conditions. STUDY DESIGN: We conducted a national cross-sectional study using data from the 1995-1996 National Ambulatory Medical Care and National Hospital Ambulatory Medical Care Surveys. Visits to private physician offices, hospital outpatient departments, and emergency departments were selected if the principal diagnoses were consistent with 1 of 9 gynecologic categories. Multiple logistic regression was used for all diagnoses to identify factors associated with visits to emergency departments or hospital-based outpatient departments compared with factors associated with visits to private physician offices. Separate regression models were developed for individual diagnoses to test the hypothesis that the factors associated with the site of care would vary across different gynecologic conditions. RESULTS: There were 23,194,000 visits for gynecologic conditions during the 2-year study period. Genital dysplasia, ovarian disorders, and uterine disorders were associated with greater use of hospital outpatient departments and emergency departments compared with physician offices. There was a 30% to 50% reduction in emergency room use for visits by women aged 45 years and older compared with visits by women aged 18 to 29 years. Emergency department use for several gynecologic conditions was 5 to 8 times greater for visits by women with household income <$29,000 than for visits by women with household income > or =$40,000. CONCLUSION: Specific gynecologic diagnoses and patient factors are associated with greater use of emergency departments or hospital outpatient departments compared with physician offices. The association of these factors with the site of care varies across different gynecologic conditions. PMID- 11262449 TI - Effectiveness of emergency contraceptive pills between 72 and 120 hours after unprotected sexual intercourse. AB - OBJECTIVE: This article reopens the issue of a medical practice that has been well established for more than three decades by proposing an extension of the period during which treatment with the "morning-after" pill is currently prescribed. The objective is to determine the effectiveness of the regimen of Yuzpe and Lancee when it is administered between 72 and 120 hours after sexual intercourse. STUDY DESIGN: We conducted an observational study comparing 2 groups of women for whom the regimen of Yuzpe and Lancee was administered after unprotected sexual intercourse. One group (usual time frame treatment group) sought consultation within 72 hours (n = 131), and the other (extended time frame treatment group) after 72 to 120 hours (n = 169). RESULTS: The pregnancy rate was 0.8% for the <72-hour group and 1.8% for the 72- to 120-hour group. The effectiveness rate varied from 87% to 90% for the <72-hour group and from 72% to 87% for the 72- to 120-hour group. In both groups the chi(2) tests showed that emergency contraceptive pills significantly reduced the risk of pregnancy. CONCLUSIONS: Women should be encouraged to seek consultation as quickly as possible after unprotected sexual intercourse. However, if the usual time limit (<72 hours) has expired, the so-called "morning-after" pill should be recommended if an intrauterine contraceptive device is not available. Emergency contraceptive pills have a favorable success rate after 72 hours, with a pregnancy rate that is significantly lower than would be expected if no contraceptive were administered. PMID- 11262450 TI - Effect of excisional therapy and highly active antiretroviral therapy on cervical intraepithelial neoplasia in women infected with human immunodeficiency virus. AB - OBJECTIVE: Our purpose was to determine the rates of recurrence, persistence, and progression of cervical intraepithelial neoplasia in women who were seropositive for human immunodeficiency virus after excisional therapy with and without highly active antiretroviral therapy. STUDY DESIGN: The records of 118 women with cervical intraepithelial neoplasia, 56 of whom were infected with human immunodeficiency virus and 62 of whom were not infected, were examined to compare outcomes. Demographic, behavioral, and clinical indices were analyzed. RESULTS: Of 54 women infected with human immunodeficiency virus, 31 (57.4%) had persistent or recurrent cervical intraepithelial neoplasia, in comparison with 10 (16.7%) of 60 noninfected women (P <.01). Progression occurred in 4 (16.7%) of 54 in the infected group and in 3 (5.0%) of 60 in the noninfected group (P <.05). In 21 (60.0%) of 35 infected women, in comparison with 8 (32%) of 25 noninfected women, disease persisted 6 months after diagnosis if treatment was not given (P <.05). Of 19 infected women, 10 (52.6%) had recurrent disease after treatment, compared with 2 (5.7%) of 35 noninfected women (P <.01). Risk factors for recurrence in women who were seropositive for human immunodeficiency virus included margin involvement of specimens obtained by loop electrosurgical excision (87.5% vs 20.0%l; P <.05). Exposure to highly active antiretroviral therapy, including therapy with protease inhibitors, was associated with a lower recurrence or persistence rate (17.6% vs. 70.3%; P <.05) and a lower progression rate (0% vs. 24%; P <.05). CONCLUSION: Women infected with human immunodeficiency virus had high rates of recurrent and persistent cervical intraepithelial neoplasia despite standard therapy. Low CD4(+) levels and margin involvement of specimens obtained by loop electrosurgical excision are risk factors for recurrence. The use of highly active antiretroviral therapy is associated with a lower risk of recurrence, persistence, and progression of cervical intraepithelial neoplasia. PMID- 11262451 TI - Hemodynamic response to tibolone in reproductive and nonreproductive tissues in the sheep. AB - OBJECTIVE: We sought to determine the hemodynamic effects of tibolone in reproductive and nonreproductive tissues in the nonpregnant ovariectomized sheep. STUDY DESIGN: Six ewes were chronically instrumented for measurement of mean arterial pressure, heart rate, cardiac output, and coronary and uterine blood flow. A dose response curve was generated for intravenous tibolone (1.25, 2.5, and 5 mg) and compared with intravenous 17beta-estradiol (1 microg/kg body weight). To determine whether tibolone-related cardiovascular responses were estrogen receptor mediated and produced by nitric oxide, animals were treated on separate days with either estrogen receptor antagonist ICI 182,780 or the nitric oxide synthase inhibitor, L -nitroarginine methyl ester. RESULTS: Tibolone significantly increased coronary blood flow in a dose-related fashion by 5% +/- 3%, 9% +/- 2%, and 11% +/- 2% for the 1.25, 2.5, and 5 mg doses, respectively. Uterine blood flow was also increased significantly in a dose-dependent manner by 98 +/- 15, 216 +/- 59, and 303 +/- 56 mL/min, for the 1.25, 2.5, and 5 mg doses, respectively. L -Nitroarginine methyl ester attenuated tibolone-induced increases in uterine blood flow by 84% +/- 4% and abolished the increase in coronary blood flow. ICI 182,780 inhibited all tibolone-induced cardiovascular responses. CONCLUSION: Tibolone significantly increases coronary and uterine blood flow in ovariectomized ewes. The coronary and uterine vascular responses are mediated via an estrogen receptor-dependent mechanism and are produced mainly by nitric oxide. PMID- 11262452 TI - A new instrument to measure sexual function in women with urinary incontinence or pelvic organ prolapse. AB - OBJECTIVE: Our aim was to develop a condition-specific, reliable, validated, and self-administered instrument to evaluate sexual function in women with pelvic organ prolapse or urinary incontinence. STUDY DESIGN: The questionnaire was designed after review of the literature and of nonspecific validated instruments. The study was completed in 2 phases. In phase 1 a total of 83 women completed both our questionnaire and the Incontinence Impact Questionnaire-7, with 20 women undergoing test-retest reliability analyses. Item analysis was based on the internal consistency, the correlations with the Incontinence Impact Questionnaire 7, the patient's age and self-rating of satisfaction, and the results of reliability testing. For final validation the questionnaire was administered in phase 2 to 99 women. Factor and item analyses were repeated, results were correlated with the Sexual History Form-12, and comparison was made between patients with high depression scores and those with low depression scores on the Symptom Questionnaire. RESULTS: Factor analysis identified 3 domains, labeled Behavioral/Emotive, Physical, and Partner-Related. Sexual function scores were highly correlated with scores on the Sexual History Form-12 for the questionnaire (r = -0.74; P <.001) and for both the Behavioral/Emotive and the Partner-Related domains (r = -0.79 and -0.5, respectively; P <.001). The Physical domain was correlated with scores on the Incontinence Impact Questionnaire-7 (r = -0.63; P <.001). Women with high depression scores on the Symptom Questionnaire had significantly lower scores on the final questionnaire, in comparison with women without depression (P <.001). CONCLUSION: We developed a condition-specific, validated, and reliable instrument, containing 31 items divided into 3 domains, to evaluate sexual functioning in women with urinary incontinence or pelvic organ prolapse. PMID- 11262454 TI - Expressions of proliferation markers (Ki-67, proliferating cell nuclear antigen, and silver-staining nucleolar organizer regions) and of p53 tumor protein in gestational trophoblastic disease. AB - OBJECTIVE: This study was undertaken to determine whether the expressions of 3 proliferation markers (Ki-67, proliferating cell nuclear antigen, and silver staining nucleolar organizer regions) and of p53 tumor protein could differentiate spontaneous abortions from gestational trophoblastic diseases and also discriminate among gestational trophoblastic disease subgroups. STUDY DESIGN: Twenty-two partial hydatidiform moles, 17 complete hydatidiform moles, 6 invasive hydatidiform moles, and 20 nonhydropic spontaneous abortions (control group) were evaluated by means of immunohistochemical techniques with antibodies to Ki-67, proliferating cell nuclear antigen, and p53. One-step silver staining was used to detect silver-staining nucleolar organizer regions. RESULTS: The expressions of Ki-67, proliferating cell nuclear antigen, silver-staining nucleolar organizer regions, and p53 were significantly higher in the gestational trophoblastic disease group than in the control group. The results of linear discriminant analysis showed that silver-staining nucleolar organizer region count had the highest sensitivity and specificity (93.3% and 100%, respectively) for distinguishing gestational trophoblastic disease from spontaneous abortion. Sensitivity and specificity for discriminating gestational trophoblastic disease from spontaneous abortion increased to 100% when all four markers were used together. Proliferating cell nuclear antigen was found to be the best discriminating variable for differentiating among gestational trophoblastic disease subgroups. CONCLUSION: Our findings suggest that expressions of Ki-67, proliferating cell nuclear antigen, silver-staining nucleolar organizer regions, and p53 may aid in the diagnosis of gestational trophoblastic diseases. These fairly rapid, simple, and economic techniques could serve as a useful adjunct to conventional methods in the diagnosis of gestational trophoblastic diseases. PMID- 11262453 TI - Interleukin 8 production and interleukin 8 receptor expression in human myometrium and leiomyoma. AB - OBJECTIVE: Interleukin 8 is a potent chemoattractant cytokine that is expressed in a variety of human tumors and is known to induce mitogenesis. We aimed to investigate the production of interleukin 8 and the expression of its receptor in myometrium and leiomyoma, in which we hypothesized that interleukin 8 may contribute to cellular proliferation. STUDY DESIGN: Myometrial and leiomyoma tissue pairs (n = 14) were obtained from human uteri after hysterectomy conducted for leiomyomatous uterus. Expression of interleukin 8 and interleukin 8 receptor type A was identified in the leiomyomatous myometrium by means of specific antibodies directed against interleukin 8 and interleukin 8 receptor type A for immunohistochemical detection. Interleukin 8 production by cultured cells was measured by enzyme-linked immunosorbent assay. The regulation of interleukin 8 messenger ribonucleic acid expression was assessed by means of the Northern blot analysis after treatment of myometrial cells with interleukin 1alpha and tumor necrosis factor alpha. Myometrial cell proliferation was determined by means of colorimetric assay after cells were treated with interleukin 8 and antihuman interleukin 8 neutralizing antibody. RESULTS: Immunostaining for both interleukin 8 and interleukin 8 receptor type A was stronger in the myometrium adjacent to leiomyoma compared with leiomyoma itself (2-fold, P <.05). Compared with samples from nonusers, samples from patients who had used gonadotropin-releasing hormone agonists revealed a trend for decreased staining for both interleukin 8 and interleukin 8 receptor type A. Interleukin 1alpha and tumor necrosis factor alpha caused a time- and dose-dependent increase in interleukin 8 production by myometrial cells (P <.001). There was a dose-dependent inhibition of cell proliferation with antihuman interleukin 8 antibody to 55% of the control (P <.001). CONCLUSION: Our demonstration of high levels of interleukin 8 and its receptor in myometrium immediately surrounding leiomyoma and the inhibition of cell proliferation when interleukin 8 is blocked by a neutralizing antibody suggest a potential role for interleukin 8 in the growth of myometrial tissue surrounding leiomyomatous tissue. This study could lead to a better understanding of potential involvement of cytokines in leiomyoma growth and in gonadatropin releasing hormone agonist-induced regression. PMID- 11262455 TI - Troglitazone is a competitive inhibitor of 3beta-hydroxysteroid dehydrogenase enzyme in the ovary. AB - OBJECTIVE: Troglitazone is a potent inhibitor of progesterone release from porcine granulosa cells. This is associated with a marked increase in pregnenolone secretion, implicating inhibition of the 3beta-hydroxysteroid dehydrogenase enzyme. This study determined whether troglitazone is a direct inhibitor of 3beta-hydroxysteroid dehydrogenase activity. STUDY DESIGN: Homogenates of porcine granulosa cells underwent classic enzyme kinetic analysis through Lineweaver-Burke and Dixon plotting. Human ovarian homogenates were also assayed for the effects of troglitazone on 3beta-hydroxysteroid dehydrogenase enzyme activity. Enzyme kinetics data were analyzed by the HyperKinetics software program. Analysis of variance was used to determine statistical significance for human ovarian homogenate experiments. RESULTS: In porcine granulosa cells Lineweaver-Burke analysis found that troglitazone inhibition of 3beta hydroxysteroid dehydrogenase enzyme activity was competitive in nature, with 5 microg/mL troglitazone increasing the apparent Michaelis constant from 1.3 to 4.3 micromol/L (no change in maximum velocity). Dixon plot analysis demonstrated that the inhibition constant for troglitazone of 3beta-hydroxysteroid dehydrogenase is approximately 6.5 microg/mL, which is in the same order of magnitude as its therapeutic concentration in blood. Troglitazone also significantly decreased the activity of 3beta-hydroxysteroid dehydrogenase in homogenates of human ovarian tissue. CONCLUSION: We conclude that troglitazone can inhibit steroidogenesis in the ovary by direct competitive inhibition of 3beta-hydroxysteroid dehydrogenase. PMID- 11262456 TI - Hormone replacement therapy for bone protection in multiparous women: when to initiate it. AB - OBJECTIVE: Hormone replacement therapy is used in postmenopausal women to improve symptoms of menopause and to protect bone and the cardiovascular system. We have evaluated the effects of parity in terms of number of deliveries on bone density and fracture risk at different ages. STUDY DESIGN: We evaluated 1875 Hispanic women > or =50 years old (61.3 +/- 8.3 years), 425 with a history of nonselective fractures and 1450 without previous fractures. Body mass index was 27.3 +/- 4.3 kg/m(2). Bone mineral densities were determined for the total body in 1468 cases, the femur in 221 cases, and the lumbar spine in 189 cases. Women were classified according to lifetime number of deliveries (from 0 to > or =5), and bone mineral densities and odds ratios for fracture risk were calculated relative to the number of deliveries. RESULTS: Bone mineral densities in total body, pelvis, and legs and total calcium and total mineral contents increased (P <.001) with > or =2 deliveries among women 50 to 59 years old but not among those > or =70 years old. The prevalence of fractures was higher in nulliparous than in multiparous women at all ages. Fracture risk was lower in multiparous women at all age groups, including those > or =70 years old (odds ratio, 0.47; 95% confidence interval, 0.26-0.84; P <.006). CONCLUSION: Bone mineral density increases with the number of deliveries until the age of 69 years. Fracture prevalence and fracture risk are lower among multiparous women even at older ages. These findings suggest that hormone replacement therapy can be delayed until 65 years of age for multiparous women but should be initiated at the beginning of menopause for nulliparous women. PMID- 11262457 TI - Human papillomavirus-associated cervical cytologic abnormalities among women with or at risk of infection with human immunodeficiency virus. AB - OBJECTIVE: Correlates of abnormal human immunodeficiency virus cervical cytologic findings were examined among women infected with human immunodeficiency virus and uninfected women. STUDY DESIGN: We performed a cross-sectional analysis of baseline data on demographically similar women with infection or risk factors for it. RESULTS: Among 1050 women without hysterectomy, squamous intraepithelial lesions were more common among women infected with human immunodeficiency virus than among uninfected women (18.8% vs 5.3%; P <.001). In multivariate analysis the association of squamous intraepithelial lesions with human papillomavirus infection was strong; adjusted prevalence ratios were 27 for high-risk, 25 for intermediate-risk, and 10 for low-risk types (95% confidence intervals, 12-58, 12 54, and 4-25, respectively). Much lower adjusted prevalence ratios were seen for the only other factor significantly associated with squamous intraepithelial lesions, namely, infection with human immunodeficiency virus in conjunction with a reduced CD4(+) cell count. Adjusted prevalence ratios were 1.9 for CD4(+) cell counts <200 and 1.6 for CD4(+) cell counts between 200 and 500 (95% confidence intervals, 1.2-3.0 and 1.0-2.5, respectively). Adjusted attributable fractions calculated for this study population indicated that if both human immunodeficiency virus and human papillomavirus were removed, 47.6% of the observed lesions with atypical squamous cells of uncertain significance and 93.4% of the observed squamous intraepithelial lesions would be prevented. CONCLUSION: Squamous intraepithelial lesions are more common among human immunodeficiency virus-infected women and are associated most commonly with high- and intermediate risk human papillomavirus types and secondarily with human immunodeficiency virus associated immune compromise. PMID- 11262458 TI - Access to services at assisted reproductive technology clinics: a survey of policies and practices. AB - OBJECTIVE: Our goal was to investigate policy on patient access to services at assisted reproductive technology clinics in the United States. STUDY DESIGN: Surveys asked about a variety of ethically and socially challenging cases and were mailed to directors of all Society for Assisted Reproductive Technology associated assisted reproductive technology clinics. RESULTS: Written policies on access to services are present at 40% of assisted reproductive technology clinics. Universal agreement was not found on any issue; 79% of clinics treat single women, 27% treat patients with a history of schizophrenia, 10% treat patients who use alcohol excessively, 7% treat human immunodeficiency virus positive women, and 2% would treat patients previously convicted of child abuse. A breakdown of the responses indicated that some clinics are more permissive in terms of access to services than others, whereas some are more restrictive. CONCLUSIONS: The data demonstrate considerable variability in policy among clinics on most access-to-services questions. The results highlight the importance of ongoing discussion of the ethical and legal issues related to access and the need to develop consistent methods to deal with complex cases. PMID- 11262459 TI - Efficacy of maintenance therapy with topical boric acid in comparison with oral itraconazole in the treatment of recurrent vulvovaginal candidiasis. AB - OBJECTIVE: Our purpose was to examine the efficacy of a topical long-term treatment with boric acid versus an oral long-term treatment (itraconazole) in the cure and prevention of recurrent vulvovaginal candidiasis. STUDY DESIGN: A prospective, nonrandomized study of patients affected by recurrent vulvovaginal candidiasis was undertaken. In 3 years we recruited 22 consecutive patients who underwent therapy with itraconazole (group 1) or boric acid (group 2). Women were followed up for 1 year, with clinic and microbiologic controls after 1, 3, 6, and 12 months after the first visit. RESULTS: During the treatment, the positive culture results (15.1% vs 12.1%) and the signs and symptoms (33.3% vs. 24.2%) were similar within the 2 groups, with no significant statistical difference. With the withdrawal, after 6 months relapses were common in the 2 groups (54.5%). CONCLUSIONS: Boric acid seems to be a valid and promising therapy both in the cure of the vaginal infection and in the prevention of relapses of recurrent vulvovaginal candidiasis, but its efficacy ends with the suspension of the therapy. PMID- 11262460 TI - Introduction of the new Centers for Disease Control and Prevention group B streptococcal prevention guideline at a large West Coast health maintenance organization. AB - OBJECTIVE: Our purpose was to assess the impact of new consensus guidelines issued by the Centers for Disease Control and Prevention, The American College of Obstetricians and Gynecologists, and the American Academy of Pediatrics to prevent perinatal group B streptococcal disease. STUDY DESIGN: We performed a descriptive analysis and a before-and-after analysis of implementation of the group B streptococcal disease prevention guidelines among singleton-birth pregnancies in 2 Group Health Cooperative hospitals from October 1, 1995, through December 31, 1997. We studied the speed and completeness of implementation and the effect on pregnancy care practices including intrapartum antibiotic use, test ordering, and maternal and neonatal health. RESULTS: Guideline implementation occurred rapidly. The proportion of term pregnancies screened according to the guideline increased markedly, and overall intrapartum antibiotic use more than doubled. Among group B streptococci-positive women, intrapartum antibiotic prophylaxis increased from 24% before to 74% after guideline implementation. Median duration of treatment before delivery increased from 1.8 to 4.3 hours. The rate of rash did not increase, and there were no cases of anaphylaxis or pseudomembranous colitis. The proportion of infants undergoing evaluation decreased after implementation of the neonatal guidelines; among infants of group B streptococci-negative women, test ordering dropped by almost 40%. CONCLUSIONS: Implementation of the new guidelines is feasible and can be accomplished rapidly. The guidelines were associated with increased maternal intrapartum antibiotic use, particularly among women at highest risk, and with a decrease in laboratory use for infants. PMID- 11262461 TI - Temporal changes in rates and reasons for medical induction of term labor, 1980 1996. AB - OBJECTIVE: This study was undertaken to assess temporal changes in rates and reasons for medical induction of term labor. STUDY DESIGN: A retrospective medical record review was conducted on a population-based cohort of 1293 women with term deliveries. RESULTS: The rate of medical labor induction increased from 12.9% in 1980 to 25.8% in 1995. Stated indications also changed, with a 2-fold increase in induction for postdate gestation, a 23-fold increase in induction for macrosomia, a 15-fold increase in elective induction, and a 22-fold decline in induction for premature rupture of membranes. The average gestational age at delivery of postdate pregnancies declined from 41.9 weeks in 1980 to 41.0 weeks in 1995. By 1995, the average maternal length of stay and the percentage of cesarean deliveries were higher among women with induced labor at term than among those with spontaneous labor at term. CONCLUSION: Induction of term labor has almost doubled in prevalence during the past 15 years. The most common indications are elective induction and postdate pregnancy, often applied to gestations of 40 to 41 weeks' duration. PMID- 11262462 TI - Dystocia among women with symptomatic uterine rupture. AB - OBJECTIVE: The purpose of this study was to analyze cervical dilatation patterns among women with uterine rupture by means of a mathematic model and to use the results to determine optimal intervention criteria. STUDY DESIGN: This was a case control review that compared a case patient group of 19 women with uterine rupture during labor with control groups with either no previous cesarean deliveries, vaginal birth after cesarean delivery, or failure of attempted vaginal birth after cesarean delivery. The mathematic model quantified dilatation and adjusted for conditions specific to each patient. Case patients were compared with matched control subjects by means of paired t tests, analysis of variance, odds ratios, and conditional logistic regression. RESULTS: Dystocia was present in 31.6% to 47.4% of patients with uterine rupture, versus 2.6% to 13.2% of the control group with no previous cesarean deliveries (P< or =.001). The incidence of an arrest disorder among patients with uterine rupture was similar to that seen in the control group with failure of attempted vaginal birth after cesarean delivery. However, the interval from diagnosis to rupture or cesarean delivery was 5.5 +/- 3.3 hours among case patients with uterine rupture and 1.5 +/- 1.3 hours in the control group with failure of attempted vaginal birth after cesarean delivery. CONCLUSION: When cervical dilatation was lower than the 10th percentile and was arrested for > or =2 hours, cesarean delivery would have prevented 42.1% of the cases of uterine rupture and resulted in excess 2.6% and 7.9% cesarean delivery rates among women with no previous cesarean deliveries and women with vaginal birth after cesarean delivery, respectively. PMID- 11262463 TI - Effect of parathyroid hormone-related peptide on human and rat myometrial contractility in vitro. AB - OBJECTIVES: The aims of this study were primarily to investigate the effects of parathyroid hormone-related peptide (human fragment 1-34) on human nonpregnant and pregnant (nonlabor and labor) myometrial contractility in vitro and secondarily to compare these effects with those of parathyroid hormone-related peptide on rat myometrial contractility. STUDY DESIGN: Isometric tension recording was performed under physiologic conditions in isolated myometrial strips obtained at hysterectomy and cesarean delivery and from Sprague-Dawley rats. The effect of cumulative additions of parathyroid hormone-related peptide (1, 10, and 100 nmol/L) on myometrial contractility was measured and the significance of results was assessed by 2-way analysis of variance. RESULTS: Parathyroid hormone-related peptide exerted a statistically significant net relaxant effect on myometrial contractility in human nonpregnant myometrium (34.71%; P<.01), in human pregnant myometrium obtained before (18.27%; P <.05) but not after (10.32%; P>.05) the onset of labor, and in rat tissue (31.60%; P<.01). CONCLUSIONS: Parathyroid hormone-related peptide exerts a relaxant effect on human and rat myometrial tissue. In human myometrium, sensitivity to parathyroid hormone-related peptide is reduced in pregnancy and abolished by the onset of labor. PMID- 11262464 TI - Correlation between prenatal velocity waveforms in the aortic isthmus and neurodevelopmental outcome between the ages of 2 and 4 years. AB - OBJECTIVE: Experimental studies on fetal lambs have shown that during an increase in the resistance to placental flow the delivery of oxygen to the brain is preserved as long as net flow through the aortic isthmus is antegrade. Our purpose was to determine whether the same changes in aortic isthmus flow in human subjects have any impact on neurodevelopmental outcome. STUDY DESIGN: Forty-four fetuses were retrospectively included in this study on the basis of an abnormal Doppler velocity in the umbilical artery. Mean gestational age at delivery was 33.0 +/- 2.0 weeks and mean birth weight 1386 +/- 435 g. The neurodevelopmental condition was assessed between the ages of 2 and 4 years. The developmental score was analyzed in relation to the flow patterns in the fetal aortic isthmus, which were classified as follows: group A, net isthmic flow antegrade (defined as the ratio of the systolic antegrade to the diastolic retrograde velocity integrals) (n = 39); group B, net isthmic flow retrograde (n = 5). RESULTS: Nonoptimal neurodevelopment was observed in 19 (49%) of 39 fetuses in group A and in all 5 fetuses (100%) in group B. This difference is significant and leads to a relative risk of 2.05 (95% confidence interval, 1.49-2.83) for neurodevelopmental deficit when predominantly retrograde flow is observed in the fetal aortic isthmus before birth. CONCLUSION: Measuring the ratio of antegrade to retrograde velocity integrals in the aortic isthmus could help in the indirect assessment of cerebral oxygenation during placental circulatory insufficiency. PMID- 11262465 TI - When stress happens matters: effects of earthquake timing on stress responsivity in pregnancy. AB - OBJECTIVE: The purpose of the study was to assess the effects of the timing of stress during pregnancy on emotional responses and birth outcome. We hypothesized that as pregnancy advanced women would become increasingly resistant to the adverse effects of stress, and so early stress would have more profound effects than later stress. STUDY DESIGN: Forty pregnant women who had experienced an earthquake during pregnancy or shortly afterward were identified. Using regression analyses we determined whether the timing of the earthquake was related to an affective response to this event and to length of gestation. RESULTS: The earthquake was rated as more stressful when it occurred early in pregnancy compared with late in pregnancy, and postpartum ratings were similar to first-trimester ratings (r (quad) =.39; P <.05). Stress experienced early in pregnancy was associated with shorter gestational length (r =.35; P <.05). CONCLUSIONS: As pregnancy advances, women become decreasingly sensitive to the effects of stress. This decrease in vulnerability may reflect increasing protection of the mother and fetus from adverse influences during pregnancy. PMID- 11262466 TI - Nutrient intake and hypertensive disorders of pregnancy: Evidence from a large prospective cohort. AB - OBJECTIVE: The objective of this analysis was to prospectively determine the effects of nutrient intakes on the incidences of preeclampsia and pregnancy associated hypertension among women enrolled in the Calcium for Preeclampsia Prevention study. STUDY DESIGN: This was a prospective observational cohort study of women in a randomized clinical trial that included women seeking prenatal care at university medical centers and affiliated clinics and hospitals in 5 US communities. A total of 4589 nulliparous women were recruited between 13 and 21 weeks' gestation. Preeclampsia and pregnancy-associated hypertension were the main outcome measures. RESULTS: Preeclampsia was noted in 326 (7.6%) of the 4314 women with known pregnancy outcomes followed up until > or =20 weeks' gestation, and pregnancy-associated hypertension was noted in 747 (17.3%). As previously reported, there was no significant difference in these outcomes between cohorts randomly assigned to supplementation with calcium or placebo. By means of logistic regression a baseline risk model was constructed for preeclampsia and pregnancy-associated hypertension. After adjustment for treatment and clinical site, body mass index >26 kg/m(2) and race were significantly associated with an increased risk of preeclampsia. Body mass index > or =35 kg/m(2), race, and never smoking were significantly associated with an increased risk of pregnancy associated hypertension. After adjustment for baseline risks, none of the 28 nutritional factors analyzed were significantly related to either preeclampsia or pregnancy-associated hypertension. CONCLUSION: We found no evidence in this study for a significant association of hypertensive disorders of pregnancy with any of the 23 nutrients measured. PMID- 11262467 TI - The preterm prediction study: can low-risk women destined for spontaneous preterm birth be identified? AB - OBJECTIVE: Half of all preterm births occur in women without clinical risk factors. Our goal was to assess fetal fibronectin assay, Bishop score, and cervical ultrasonography as screening tests to predict which low-risk pregnancies will end in preterm birth. STUDY DESIGN: We performed a secondary analysis of data collected at 22 to 24 weeks' gestation from low-risk subjects enrolled in the Preterm Prediction Study, an observational study of risk factors for preterm birth conducted by the National Institute of Child Health and Human Development Maternal-Fetal Medicine Units Network. Analysis was limited to primigravid women and to women who did not have a history of preterm birth or spontaneous pregnancy loss at <20 weeks' gestation. Bishop score (> or =4), fetal fibronectin level (> or =50 ng/mL), and cervical length (< or =25 mm) at 24 weeks' gestation were evaluated alone and in sequence as tests to predict spontaneous delivery before 35 weeks' gestation. RESULTS: Of the 2929 subjects enrolled in the original study, 2197 (1207 primigravid women and 900 low-risk multiparous women) met criteria for this analysis. There were 64 spontaneous births before 35 weeks' gestation (3.04%). All three tests were significantly related to birth before 35 weeks' gestation (high Bishop score: relative risk, 3.6; 95% confidence interval, 2.1-6.3; fetal fibronectin detection: relative risk, 8.2; 95% confidence interval, 4.8-13.9; short cervical length: relative risk, 6.9; 95% confidence interval, 4.3-11.1). However, the sensitivities of the tests alone were low (23.4% for high Bishop score, 23.4% for fetal fibronectin detection, and 39.1% for short cervix), as were the sensitivities for Bishop score followed by cervical ultrasonography (14.1%) and fetal fibronectin assay followed by cervical scan (15.6%). CONCLUSION: In the setting of low-risk pregnancy, fetal fibronectin assay and cervical ultrasonography have low sensitivity for preterm birth before 35 weeks' gestation. Sequential screening with Bishop score or fetal fibronectin assay followed by cervical ultrasonography further decreased sensitivity to only 15% among low-risk women. PMID- 11262468 TI - Prophylactic use of antibiotics for nonlaboring patients undergoing cesarean delivery with intact membranes: a meta-analysis. AB - OBJECTIVE: We performed a meta-analysis to examine the role of antibiotic prophylaxis in preventing postoperative infections among nonlaboring women undergoing cesarean delivery with intact membranes. STUDY DESIGN: A computerized literature search was performed with MEDLINE. Studies were included if they contained data on patients undergoing cesarean delivery in the absence of labor and ruptured membranes. Only randomized trials with a placebo control group were included. RESULTS: Seven studies were found. Use of antibiotics decreased the risk of all infectious outcomes reported. When the results of 4 studies were pooled, prophylactic antibiotic use was associated with a significant reduction in postoperative fever (relative risk, 0.25; 95% confidence interval, 0.14-0.44). A similar reduction was noted for endometritis in 4 studies (relative risk, 0.05; 95% confidence interval, 0.01-0.38). Two studies reported on wound infection and showed a trend toward a protective effect (relative risk, 0.59; 95% confidence interval, 0.24-1.45). CONCLUSION: The prophylactic use of antibiotics reduces the risk of postoperative infectious complications after cesarean delivery even in the population at lowest risk. PMID- 11262469 TI - Efficacy and safety of intravenously administered iron sucrose with and without adjuvant recombinant human erythropoietin for the treatment of resistant iron deficiency anemia during pregnancy. AB - OBJECTIVE: This study was undertaken to determine the efficacy and safety of intravenously administered iron sucrose with versus without adjuvant recombinant human erythropoietin in the treatment of gestational iron-deficiency anemia resistant to therapy with orally administered iron alone. STUDY DESIGN: Forty patients with gestational iron-deficiency anemia were randomly assigned to receive intravenously iron sucrose plus recombinant human erythropoietin or iron sucrose alone twice weekly. Target hemoglobin value was 11.0 g/dL. Efficacy measures were reticulocyte count, increase in hematocrit, and time to target hemoglobin level (treatment duration in weeks and need for continued therapy after 4 weeks). RESULTS: Both regimens were effective, but with adjuvant recombinant human erythropoietin the reticulocyte counts were higher from day 4 (P<.01), increases in hematocrit were greater from day 11 (P <.01), and the median duration of therapy was shorter (18 vs 25 days), with more patients reaching the target hemoglobin level by 4 weeks of treatment (n = 19 vs. n = 15). The groups did not differ with respect to maternal-fetal safety parameters. CONCLUSION: Adjuvant recombinant human erythropoietin safely enhanced the efficacy of iron sucrose in the treatment of gestational iron-deficiency anemia resistant to orally administered iron alone. PMID- 11262470 TI - Relationship of race and severity of neonatal illness. AB - OBJECTIVE: Our goal was to determine whether there are racial differences in the severity of illness on admission for premature newborn infants independent of gestational age. STUDY DESIGN: The study population consisted of all African American and Caucasian singleton infants with gestational ages <34 weeks who were admitted to the neonatal intensive care unit at Brigham and Women's Hospital between December 1994 and November 1995. Illness severity was measured with a neonatal severity of illness score, the SNAP score (Score for Neonatal Acute Physiology). The SNAP score is a physiologic scoring system that ranks the worst physiologic derangements in each organ system in the first 12 hours of life. It is an objective measure of neonatal illness severity with scores ranging from 0 (healthy) to 42 (most severely ill). Student t tests, chi(2) analysis, and Fisher exact tests were used to assess statistical significance. Linear and logistic regression analyses were used to examine associations while confounding factors were controlled for. RESULTS: There were 129 (79%) Caucasian and 36 (22%) African American newborns included in the analysis. Caucasian newborns had significantly higher mean SNAP scores than African American newborns (8.8 vs. 6.3; P <.05). Compared with African American newborns, Caucasian newborns were more than twice as likely to have a SNAP score >10 (33% vs. 14%; P <.05). In a linear regression analysis in which we controlled for gestational age, birth weight, preterm premature rupture of membranes, preterm labor, preeclampsia, intrapartum fever > or =100.4 degrees F, route of delivery, and other maternal and fetal factors, African American newborns were predicted to have a SNAP score that was on average 3.0 points lower than that of Caucasian newborns (P =.005). In a logistic regression in which we controlled for the above-mentioned confounders, African American newborns were only 14% as likely to have a SNAP score >10 when compared with Caucasian newborns (odds ratio, 0.14; 95% confidence interval, 0.04-0.51). CONCLUSIONS: Over a broad range of prematurity, Caucasian newborns were more ill than African American newborns on admission to the neonatal intensive care unit. PMID- 11262471 TI - The effect of vascular coiling on venous perfusion during experimental umbilical cord encirclement. AB - OBJECTIVE: The aim of these studies was to compare venous perfusion in umbilical cords subjected to a standardized tight encirclement force. Comparisons were made between cords from normal pregnancies and those complicated by gestational diabetes mellitus and intrauterine growth restriction. STUDY DESIGN: The cannulated cord segment was wrapped around a plastic container, which in turn was attached with nylon string to a hanging graduated measuring cylinder in which known volumes of water could be applied for weight. The cord was perfused with Krebs solution to a constant venous perfusion pressure of 40 mm Hg. Weights of 100-g increments were applied until total cessation of venous perfusion was observed. The weight, length, number of vascular coils, and degree of hydration were recorded for each cord. The coiling index was defined as the number of vascular coils per 10 cm of cord. RESULTS: Regression analysis of 34 cords (normal, n = 16; gestational diabetes mellitus, n = 12; intrauterine growth restriction, n = 6) identified a significant inverse correlation (P =.0003, Spearman rank correlation) between coiling index and the minimum weight required to occlude venous perfusion. Cords from pregnancies complicated by intrauterine growth restriction displayed a higher frequency of vascular coiling and were more easily occluded (median weight, 350 g) than were cords from pregnancies complicated by gestational diabetes mellitus, which were less coiled and tended to resist occlusion (median weight, 1100 g). CONCLUSION: During experimental cord encirclement there was a significant inverse relationship between vascular coiling and susceptibility to cord venous occlusion when traction was applied to the encirclement. PMID- 11262472 TI - Proteoglycans and hyaluronan in human fetal membranes. AB - OBJECTIVE: The aim of this study was to describe the distributions of major extracellular matrix components, such as proteoglycans, collagen and hyaluronan, in the fetal membranes at term. STUDY DESIGN: Fetal membranes were obtained from elective cesarean deliveries at term. Guanidinium extracts were analyzed for proteoglycans with alcian blue precipitation, sodium dodecyl sulfate- polyacrylamide gel electrophoresis, and Western blotting and for hyaluronan with a radioimmunoassay. Collagen was measured by estimating hydroxyproline content. Tissue sections were immunostained for decorin and biglycan and stained for hyaluronan with a biotin-labeled hyaluronan-binding protein. RESULTS: The fetal membranes contained predominantly smaller proteoglycans, such as biglycan and decorin. The amnion consisted of typical fibrous connective tissue with a high concentration of collagen. The amnion was dominated by decorin located in close connection with the collagen fibrils. The chorion was composed of a fibroblastic part containing collagen and decorin and a trophoblastic part mainly containing biglycan. In addition, large amounts of hyaluronan were found, especially in the amnion and in the decidual cell layers. CONCLUSION: The distributions of proteoglycans, collagen, and hyaluronan in human fetal membranes may explain the biomechanical properties of this tissue. We suggest that changes in the relative proportions of these extracellular molecules are crucial for the proposed maturation process in the fetal membranes during the last weeks of pregnancy. PMID- 11262473 TI - Marked variation in responses to long-term nitric oxide inhibition during pregnancy in outbred rats from two different colonies. AB - OBJECTIVE: Some but not all studies have shown that long-term nitric oxide synthase inhibition during pregnancy induces symptoms similar to those of preeclampsia that include hypertension, proteinuria, and intrauterine growth restriction. This study was undertaken to compare the effects of long-term nitric oxide synthase inhibition during pregnancy on blood pressure and fetal weight between Sprague-Dawley rats from outbred colonies of two different suppliers. STUDY DESIGN: Osmotic minipumps were inserted on day 10 or day 17 of pregnancy in Sprague-Dawley rats obtained from Charles River Laboratories, Inc, Wilmington, Mass, or Harlan Sprague Dawley, Inc, Indianapolis, Ind. The pumps were set to deliver vehicle only (control group) or N omega-nitro-L -arginine methyl ester (a nitric oxide synthase inhibitor) at a rate of 50 mg/d until postpartum day 7. Systolic blood pressures were measured daily with the tail-cuff method. Neonatal weights and survival were recorded. RESULTS: N omega-nitro-L -arginine methyl ester infusion initiated on gestational day 10 increased blood pressure relative to control levels in all rats studied. Harlan rats were much more sensitive to the hypertensive effect of N omega-nitro-L -arginine methyl ester. When N omega nitro-L -arginine methyl ester infusion was initiated on gestational day 17, blood pressure increased only in Harlan rats. Pups born to Harlan rats treated with N omega-nitro-L -arginine methyl ester had lower birth weights and a higher stillbirth rate than did pups of Charles River rats. The degree of hypertension was significantly correlated with the deleterious effects of N omega-nitro-L arginine methyl ester on the fetuses. CONCLUSION: Within the same strain of rats the effects of long-term nitric oxide synthase inhibition on blood pressure and fetal outcome depended on the original animal colony, with animals from Harlan Sprague Dawley being more sensitive than those from Charles River Laboratories. This difference in response between animals from different suppliers is most likely caused by genetic differences inbred into the strain. In addition to explaining some of the reported inconsistencies between laboratories, these results may also provide insights into the genetic basis of preeclampsia. PMID- 11262474 TI - Risk factors for sudden intrauterine unexplained death: epidemiologic characteristics of singleton cases in Oslo, Norway, 1986-1995. AB - OBJECTIVE: The epidemiologic characteristics of unexplained stillbirths are largely unknown or unreliable. We define sudden intrauterine unexplained death as a death that occurs antepartum and results in a stillbirth for which there is no explanation despite postmortem examinations, and we present risk factors for this type of stillbirth in singleton gestations. STUDY DESIGN: Singleton antepartum stillbirths (n = 291) and live births (n = 582) in Oslo were included and compared with national data (n = 2025 and n = 575,572, respectively). Explained stillbirths (n = 165) and live births in Oslo served as controls for the cases of sudden intrauterine unexplained death (n = 76) in multiple logistic regression analyses. RESULTS: One fourth of stillbirths remain unexplained. The risk of sudden intrauterine unexplained death (1/1000) increased with gestational age, high maternal age, high cigarette use, low education, and overweight or obesity. Primiparity and previous stillbirths or spontaneous abortions were not associated with sudden intrauterine unexplained death. CONCLUSIONS: Risk factors for sudden intrauterine unexplained death are identifiable by basic antenatal care. Adding unexplored stillbirths to the unexplained ones conceals several risk factors and underlines the necessity of a definition that includes thorough postmortem examinations. PMID- 11262475 TI - Significant fetal-maternal hemorrhage after termination of pregnancy: implications for development of fetal cell microchimerism. AB - OBJECTIVE: Recent reports that an association exists between fetal cell microchimerism and autoimmune disease has increased interest in the postpartum persistence of fetal cells. The purpose of this study was to determine, by means of quantitative polymerase chain reaction amplification, whether a significant fetalmaternal hemorrhage occurs after elective termination of pregnancy. STUDY DESIGN: Blood samples were obtained from 23 women who underwent termination of pregnancy immediately before venipuncture; these samples were subjected to analysis by quantitative polymerase chain reaction amplification with the use of Y-chromosome primers. There were 21 male and 2 female fetuses. Results were equilibrated to 16 mL and analyzed by a weighted linear regression analysis to evaluate the correlation between detected fetal nucleated cell equivalents and gestational weeks. RESULTS: Among the 21 known male fetuses, the median number of detected fetal nucleated cell equivalents was 1552 (range, 50-37,618). The female fetuses had no fetal nucleated cell equivalents detected. A positive dependence of male fetal nucleated cell equivalents on gestational age was shown (P <.001). CONCLUSION: Analysis by quantitative polymerase chain reaction amplification demonstrated a large fetal-maternal transfusion after elective abortion. Consideration of the biologic consequences of pregnancy and the potential for future development of fetal cell microchimerism must now extend to a larger population of women. PMID- 11262476 TI - Developmental changes in reactivity of small femoral arteries in the fetal and postnatal baboon. AB - OBJECTIVE: We evaluated in vitro responsiveness of small arteries (internal diameter, 300 microm) from the femoral vascular bed of normal fetal (0.75-1.0 gestation) and neonatal (43-46 days) baboons to investigate whether the transition from fetal to neonatal life was associated with functional alterations in vasoconstrictor and vasodilator responses. STUDY DESIGN: The maximum response and sensitivity to potassium and to the constrictor agonists norepinephrine and U46619 (a thromboxane mimetic) were studied by in vitro myography. Vasodilator responses to the endothelium-dependent dilators acetylcholine and bradykinin were also investigated. RESULTS: The maximum response to norepinephrine and U46619 and to potassium increased with gestational age, whereas the sensitivity to these vasoconstrictors was similar in all groups studied. In contrast, acetylcholine- and bradykinin-induced relaxation (median effective concentration and maximum response) did not change with age. CONCLUSION: Receptor-mediated responses to a catecholamine, a prostanoid, and 2 endotheliumdependent vasodilators are similar in the fetal and neonatal baboon. The increase in maximal constriction with development, which is probably associated with growth or maturation of vascular smooth muscle, is likely to be a functionally important aspect in the development of cardiovascular function. PMID- 11262477 TI - The impact of maternal ketonuria on fetal test results in the setting of postterm pregnancy. AB - OBJECTIVE: The aim of this study was to determine whether ketonuria, a commonly assessed urinary marker of maternal starvation and dehydration, is associated with abnormal fetal test results in the setting of postterm pregnancy. STUDY DESIGN: During a 4-year period (January 1993-December 1996), a total of 3655 visits for antepartum maternal-fetal testing of postterm pregnancies (> or =41 weeks' gestation) occurred at our institution. Maternal assessment included vital signs and urinalysis. The presence and degree of maternal ketonuria was correlated against abnormal results of fetal heart rate tests, nonstress tests, amniotic fluid index measurements, and biophysical profile scores performed on the same day. RESULTS: There were 3601 encounters suitable for inclusion in the study. Clinically detectable ketonuria occurred in 10.9% of the patients studied. Patients with clinically detectable ketonuria were at increased risk relative to patients without ketonuria for abnormal outcomes during postterm testing, including the presence of oligohydramnios (24% vs. 9.3%; P<.0001 ), nonreactive nonstress tests (6.2% vs. 2.15%; P<.0001), and fetal heart rate decelerations (14% vs 9.2%; P =.0039 ). CONCLUSION: Maternal ketonuria among patients with postterm pregnancy was associated with a >2-fold increase in the occurrence of oligohydramnios, a 3-fold increase in nonreactive nonstress tests, and a significant increase in fetal heart rate decelerations. Further studies are required to evaluate the potential benefits of treating ketonuria before fetal testing. PMID- 11262478 TI - Fetal loss rate associated with cordocentesis at midgestation. AB - OBJECTIVE: The aim of this study was to assess the risk of fetal loss attributable to cordocentesis at midgestation. STUDY DESIGN: A cohort study was conducted during the period 1989-1999. Women undergoing cordocentesis between 16 and 24 weeks' gestation with singleton pregnancies without obvious fetal anomaly were recruited into the study group. The control subjects were selected prospectively on a one-to-one basis with strict matching for maternal age, parity, gestational age at recruitment, and socioeconomic status. Both groups were prospectively followed up until delivery. RESULTS: A total of 1281 women with successful cordocentesis and their matched control subjects were recruited to the study. After exclusion of some pairs because of loss to follow-up or fetal malformations or severe disease necessitating termination of pregnancy, 1020 matched pairs were available to be compared with respect to fetal loss rate and pregnancy outcomes. The fetal loss rate was significantly higher among the study group (3.2% vs. 1.8%; P <.05, McNemar test). However, there were no significant differences in other obstetric complications between the study and control groups. CONCLUSION: The incremental fetal loss rate associated with cordocentesis at midgestation was about 1.4%. PMID- 11262479 TI - The prediction and prevention of intrapartum fetal asphyxia in term pregnancies. AB - OBJECTIVE: This study was undertaken to examine the roles of clinical risk scoring, electronic fetal heart rate monitoring, and fetal blood gas and acid base assessment in the prediction and prevention of intrapartum fetal asphyxia in term pregnancies. STUDY DESIGN: The outcomes of 166 term pregnancies with biochemically confirmed fetal asphyxia (umbilical artery base deficit at delivery, >12 mmol/L) were examined. This population included 83 pregnancies delivered abdominally matched with 83 pregnancies delivered vaginally. Antepartum and intrapartum clinical risk factors and neonatal complications were documented. Fetal assessments included fetal heart rate patterns in the fetal heart rate record and fetal capillary blood gas and acid-base assessments. Fetal asphyxia was classified as mild, moderate, or severe on the basis of umbilical artery base deficit (cutoff >12 mmol/L) and neonatal encephalopathy and other organ system complications. RESULTS: Fetal asphyxial exposures were as follows: mild, 140; moderate, 22; and severe, 4. Intervention and delivery during the first or second stage of labor occurred in 98 of the 166 pregnancies. Predictive fetal heart rate patterns were the primary indication leading to intervention and delivery during the first or second stage of labor. Clinical risk factors when present were secondary indications in the clinical decision to intervene. Fetal blood gas and acid-base assessment was a useful supplementary test in 41 pregnancies. Intervention and delivery may have prevented the progression of mild asphyxia in 78 pregnancies and may have modified the degree of moderate or severe asphyxia in 20 pregnancies. CONCLUSION: Although fetal heart rate patterns will not discriminate all asphyxial exposures, continuous fetal heart rate monitoring supplemented by fetal blood gas and acid-base assessment can be a useful fetal assessment paradigm for intrapartum fetal asphyxia. Such an assessment paradigm will not prevent all cases of moderate or severe fetal asphyxia. However, prediction and diagnosis with intervention and delivery during the first or second stage of labor could prevent the progression of mild asphyxia to moderate or severe asphyxia in some cases. PMID- 11262480 TI - Developmental changes in tolerance to transient intrauterine ischemia in rat cerebral mitochondria. AB - OBJECTIVE: Mitochondfial respiratory activities were measured in neonatal rat brain to compare the influence of transient intrauterine ischemia in the preterm fetus with that in the term fetus and to evaluate the effect of alpha-phenyl-N tert-butyl-nitrone treatment. STUDY DESIGN: Intrauterine ischemia was induced by a 30-minute occlusion of the right uterine artery. The control group consisted of term fetuses (20 days old) exposed to normoxia (n = 8) and ischemia (n = 8). For the investigation into maturity effect, preterm fetuses (14 days old) were exposed to normoxia (n = 8) or ischemia (n = 8), and for the alpha-phenyl-N -tert butyl-nitrone treatment investigation, term fetuses were exposed to ischemia with alpha-phenyl-N -tert-butyl-nitrone (n = 8). All subjects underwent cesarean delivery at 21 days of gestation, and the mitochondrial respiration was measured polarographically 1 hour after delivery. RESULTS: In the control group the neonatal cortical tissue exposed to ischemia showed a significant decrease in mitochondrial activities compared with those in normoxic control animals. In the preterm group the mitochondrial activities of ischemic fetuses were maintained close to normoxic levels. The neonatal mitochondrial deterioration caused by term ischemia was prevented by alpha-phenyl-N -tert-butyl-nitrone. CONCLUSION: The results indicate that preterm fetuses are more capable than term fetuses of maintaining mitochondrial function under conditions of transient intrauterine ischemia and suggest that oxygen derived free radicals may play a crucial role in the development of neonatal neurologic deficit. PMID- 11262482 TI - ST waveform changes during repeated umbilical cord occlusions in near-term fetal sheep. AB - OBJECTIVE: This study was undertaken to determine whether changes in the fetal ST waveform during repeated umbilical occlusion reflect the development of hypotension and acidosis. STUDY DESIGN: Chronically instrumented, near-term fetal sheep received 1-minute total umbilical cord occlusion either every 5 minutes for 4 hours (1:5 group, n = 8), or every 2.5 minutes until blood pressure fell <20 mm Hg on 2 successive occlusions (1:2.5 group, n = 8). RESULTS: Umbilical cord occlusion caused variable decelerations, with sustained hypertension in the 1:5 group and little change in acid-base status (pH = 7.34 +/- 0.07 after 4 hours). In contrast, the 1:2.5 group showed progressive hypotension and metabolic acidemia (pH 6.92 +/- 0.1 after the final occlusion). There was a marked increase in ST waveform height during occlusions; this increase was greater in the 1:2.5 group (P <.001), but there was overlap between the groups. ST waveform height between occlusions was significantly higher in the 1:2.5 group (P <.001) until negative and biphasic ST waveforms developed in these fetuses between occlusions in the final 30 minutes. CONCLUSION: ST waveform elevation occurs during umbilical cord occlusions but only crudely reflects the severity of hypoxia. Interocclusion waveform height may be a better reflection of the severity of hypoxia. The appearance of biphasic and negative waveforms between occlusions may be a useful marker for severe decompensation. PMID- 11262481 TI - Effect of gestational age and hypoxia on activity of ribonucleic acid polymerase in fetal guinea pig brain. AB - OBJECTIVE: The aim of this study was to determine the effect of gestational age and hypoxia on the activity of ribonucleic acid polymerase in fetal guinea pig brain. STUDY DESIGN: Fetal cerebral cortical neuronal nuclei were isolated at 40, 50, and 60 days (term) of gestation to determine the effect of gestational age on the activity of ribonucleic acid polymerase I, II, and III. Pregnant guinea pigs at 60 days' gestation were randomly assigned to a normoxic or hypoxic group to determine the effect of hypoxia on ribonucleic acid polymerase activity. The fetal neuronal nuclei were pooled from 6 pregnant animals in each group. In the normoxic group the pregnant guinea pigs were exposed to room air before delivery. In the hypoxic group delivery occurred after the pregnant guinea pig had been exposed to 7% oxygen for 60 minutes. The fetuses were delivered by cesarean, and the fetal cerebral cortical neuronal nuclei were isolated immediately. Ribonucleic acid polymerase activity was determined with nuclei suspended in a buffer containing adenosine triphosphate, guanosine triphosphate, cytidine triphosphate, and tritiated uridine triphosphate. Dactinomycin (actinomycin D) and polydeoxyadenylic-thymidylic acid were used to determine the activity of bound and free ribonucleic acid polymerase. alpha-Amanitin was used to determine the activity of ribonucleic acid polymerase II. RESULTS: The activity of total (bound and free) ribonucleic acid polymerase I and III increased from 85.4 +/- 9.4 fmol of tritiated uridine triphosphate incorporated per milligram of protein per hour at 40 days' gestation to 233.3 +/- 82.1 fmol at 50 days and to 343.4 +/- 231.6 fmol at 60 days (P =.02). Total ribonucleic acid polymerase II activity increased from 19.9 +/- 6.0 fmol of tritiated uridine triphosphate incorporated per milligram of protein per hour at 40 days to 123.8 +/- 53.0 fmol at 50 days and to 200.9 +/- 77.8 fmol at 60 days (P <.01). In the term fetal guinea pig brain the activity of bound ribonucleic acid polymerase I and III decreased from 116.8 +/- 107.2 fmol of tritiated uridine triphosphate incorporated per milligram of protein per hour under normoxic conditions to 92.8 +/- 76.0 fmol in hypoxic fetal brain, a decrease of 20.5%. Free ribonucleic acid polymerase I and III activity decreased from 199.2 +/- 115.2 fmol of tritiated uridine triphosphate incorporated per milligram of protein per hour in normoxic fetal brain to 132.0 +/- 66.4 fmol in hypoxic fetal brain, a decrease of 33.8%. Free ribonucleic acid polymerase II activity decreased from 62.4 +/- 70.4 fmol of tritiated uridine triphosphate incorporated per milligram of protein per hour in normoxic fetuses to 13.6 +/- 9.6 fmol in hypoxic fetal brain, a decrease of 78.2%. In contrast, however, in term fetal guinea pig brain, bound ribonucleic acid polymerase II activity increased from 8.0 +/- 10.4 fmol of tritiated uridine triphosphate incorporated per milligram of protein per hour under normoxic conditions to 35.2 +/- 8.8 fmol in hypoxic fetal brain, an increase of 340% (P <.01). CONCLUSION: The activity of ribonucleic acid polymerases I, II, and III increases throughout the latter half of gestation, from 40 to 60 days, in the fetal guinea pig brain. Hypoxia in utero is associated with a decrease in ribonucleic acid polymerase I and III activity. Although hypoxia is associated with a decrease in free ribonucleic acid polymerase II activity, we observed a marked increase in bound ribonucleic acid polymerase II activity, which may represent a hypoxia-induced alteration of gene expression. PMID- 11262483 TI - Screening for substance use in pregnancy: a practical approach for the primary care physician. AB - Our goal was to identify risk factors for substance use during pregnancy for primary care physicians so that we could assess a woman's risk of alcohol or illicit drug use. Participants were 2002 Medicaid-eligible pregnant women with < or =2 visits to prenatal care clinics in South Carolina and Washington State. Structured interviews were used to collect data. Logistic regressions and classification and regression trees identified predictors for pregnant women at high risk for substance use. Approximately 9% of the sample reported current use of either drugs or alcohol or both. Past use of alcohol or cigarettes, including during the month before pregnancy, most differentiated current drug or alcohol users from current nonusers. Our analysis suggests that primary care physicians can ask 3 questions in the context of a prenatal health evaluation to target women for referral to a full clinical assessment for drug and alcohol use. PMID- 11262484 TI - Prevention, diagnosis, and treatment of venous thromboembolic complications of gynecologic surgery. AB - Deep vein thrombosis and pulmonary embolism, collectively referred to as venous thromboembolic events, are a source of significant morbidity and mortality after gynecologic surgical procedures. In this literature review the advantages and disadvantages of various preventive measures for deep venous thrombosis, including low-molecular-weight heparins, are discussed. The most appropriate prophylactic methods for patients in varying risk categories are recommended. Current methods of diagnosing deep venous thrombosis and pulmonary embolism, including ultrasonography, venography, ventilation-perfusion scan, helical computed tomographic scan, and D -dimer measurement are then discussed. Finally, treatment modalities for deep venous thrombosis and pulmonary embolism, including heparin, low-molecular-weight heparin, warfarin, and thrombolytic therapy, are detailed. PMID- 11262485 TI - Umbilical cord vulnerability. PMID- 11262487 TI - Fetal heart rate monitoring: need for a stricter randomized study. PMID- 11262489 TI - Sildenafil citrate and sperm function. PMID- 11262491 TI - Lack of utility of amniotic fluid index in predicting perinatal morbidity and mortality. PMID- 11262493 TI - Progression of labor during induction. PMID- 11262495 TI - Vaginal birth after cesarean delivery? PMID- 11262497 TI - An appeal for a fair evaluation of the impact of elective induction of labor on population cesarean delivery rates. PMID- 11262499 TI - Potential for brief but severe intrapartum injury among neonates with early-onset seizures and elevated nucleated red blood cell counts. PMID- 11262501 TI - Questions about preeclampsia outcome predictors. PMID- 11262503 TI - A randomized trial of a dissonance-based eating disorder prevention program. AB - OBJECTIVE: As psychoeducational eating disorder prevention programs have not been shown to reduce bulimic pathology, we developed and evaluated a dissonance-based intervention for high-risk populations. METHOD: Young women (N = 87) with body image concerns were randomized to this intervention, which involves verbal, written, and behavioral exercises requiring them to critique the thin-ideal, or to a healthy weight management control group. Participants completed a baseline, termination, and 4-week follow-up survey. RESULTS: Participants in the dissonance intervention reported decreased thin-ideal internalization, body dissatisfaction, dieting, negative affect, and bulimic symptoms at termination and at 4-week follow-up. Unexpectedly, participants in the healthy weight management control group also reported some benefits. DISCUSSION: Taken in conjunction with past findings, these preliminary results suggest that the dissonance intervention, and to a lesser extent the healthy weight management intervention, may reduce bulimic pathology and risk factors for eating disturbances. PMID- 11262504 TI - Males with anorexia nervosa: a controlled study of eating disorders in first degree relatives. AB - OBJECTIVE: To compare lifetime rates of full and partial anorexia nervosa and bulimia nervosa in first-degree relatives of males with anorexia nervosa and in relatives of never-ill comparison subjects. METHODS: Rates of eating disorders were obtained for 747 relatives of 210 probands from personal structured clinical interviews and family history. Best-estimate diagnoses were determined blind to proband diagnosis and pedigree status. RESULTS: Full and partial syndromes of anorexia nervosa aggregated in female relatives of ill probands. For the full syndrome of anorexia nervosa, the crude relative risk was 20.3 among female relatives and for partial syndrome anorexia nervosa, the crude relative risk was 3.3. In contrast, bulimia nervosa was relatively uncommon among relatives of ill probands. CONCLUSION: Although anorexia nervosa in males is exceedingly rare, there is a pattern of familial aggregation that is highly similar to that observed in recent family studies of affected females. On the basis of these findings, there is no evidence that familial-genetic factors distinguish the occurrence of anorexia nervosa in the two sexes. PMID- 11262505 TI - Pathways mediating sexual abuse and eating disturbance in children. AB - OBJECTIVE: To examine the relationship between childhood maltreatment and eating disorders in a sample of children. METHOD: Twenty 10-15-year-old female children who were receiving treatment following reported childhood sexual abuse and 20 age matched controls were compared on a series of measures assessing eating disorder behaviors, body image concerns, substance use, mood, impulsive behavior, and self concept. RESULTS: Sexually abused children reported higher levels of eating disorder behaviors, impulsive behaviors, and drug abuse than controls. Furthermore, behavioral impulsivity provided the strongest mediational effect between a history of childhood sexual abuse and purging and restrictive dieting behavior. Drug use proved to be a significant secondary mediator of the childhood sexual abuse eating disorder behavior association. DISCUSSION: These data support the hypothesis that childhood sexual abuse is related to disordered eating in children, and extend similar findings that have been previously reported with adults. Behavioral impulsivity and drug use appear to be significant mechanisms that influence eating disorder behavior following childhood sexual abuse. PMID- 11262506 TI - Disordered eating and the transition to college: a prospective study. AB - OBJECTIVE: A longitudinal study was conducted to examine whether the transition to college changed eating disorder symptoms and related attitudes. METHOD: Participants were 342 women who completed an in-depth survey in the spring of their senior year of high school and again during their first year of college. We assessed changes in body self-perception, eating-related attitudes, and disordered eating classification (nondieter, dieter, problem dieter, subclinical eating disordered, or eating disordered on the basis of criteria for bulimia nervosa in the 4th ed. of the Diagnostic and Statistical Manual of Mental Disorders). RESULTS: Although participants viewed themselves as significantly heavier in their first year of college, dieting frequency and disordered eating classification in college did not differ from high school assessment. DISCUSSION: Evidence from this study indicates that disordered eating symptoms and attitudes are established before college. However, our findings also reveal that poor self image, dieting behaviors, and eating disorder symptoms are common among many young women, both before and during college. PMID- 11262507 TI - Exposure to westernization and dieting: a cross-cultural study. AB - OBJECTIVES: The study aimed to establish whether an index of exposure to westernization would predict dieting behavior over and above the predictors of body mass index (BMI) and social influences. The study also sought to compare dieting behaviors among adolescents from three different cultural backgrounds. METHOD: A total of 100 females from Beijing, China, 60 females of Chinese heritage living in Sydney, Australia, and 100 female Australians of no Chinese background were assessed. The exposure to westernization index incorporated the country of birth, the predominant language spoken at home, the country of birth of one's parents, and the country of residence. RESULTS: Exposure to westernization was found to be a significant predictor of dieting status. The westernization index remained an important predictor when BMI and social influences to diet were taken into account. Interestingly, the Chinese Australian girls dieted the least, although the Chinese girls living in China perceived more influence from their peers to diet, despite their lower BMI. CONCLUSION: The exposure to westernization index provides a useful assessment of important influences on dieting in adolescent females. PMID- 11262508 TI - Importance of size in defining binge eating episodes in bulimia nervosa. AB - OBJECTIVE: This study sought to determine if amount of food consumed is important in defining binge eating episodes in individuals with bulimia nervosa (BN). METHOD: Women (N = 30) with DSM-IV BN (OBN) and women (N = 25) who would have met DSM-IV criteria for BN except that their binge episodes were not objectively large (SBN) were recruited from the community. Subjects completed telephone interviews and questionnaires. RESULTS: Results demonstrated no significant differences between women with OBN and SBN in levels of dietary restraint, disinhibition, or hunger; no significant differences in general psychopathology; and significant differences in frequency of binge/purge episodes and impulsiveness. Differences in impulsiveness remained after controlling for frequency of binge/purge episodes. DISCUSSION: These results partially validate current diagnostic criteria for bulimia nervosa and elucidate one factor, impulsiveness, that may be important in understanding objective binge episodes in bulimia nervosa. PMID- 11262509 TI - Schema avoidance in bulimic and non-eating-disordered women. AB - OBJECTIVE: Models of bulimia lack a clear conceptualization of avoidance. This study considers the role of different domains of schema avoidance in bulimic disorders and examines the association of scores on the Young-Rygh Avoidance Inventory (YRAI) with bulimic pathology. METHODS: A total of 19 bulimic and 74 comparison women completed the YRAI and a measure of bulimic psychopathology. RESULTS: Bulimics scored significantly higher than the nonclinical women did on all YRAI scales. Greater reported use of avoidance was positively associated with bulimic attitudes, but only among the comparison group. At a dimensional level, behavioral/somatic avoidance was more strongly associated with bulimic pathology than cognitive/emotional avoidance, but the same was not true when differentiating groups. CONCLUSIONS: The YRAI is a robust measure of different domains of schema avoidance in understanding bulimic psychopathology. Clinically, the YRAI might be used to guide treatment for bulimic disorders. PMID- 11262510 TI - The role of perfectionism and excessive commitment to exercise in explaining dietary restraint: replication and extension. AB - The etiological complexity of the eating disorders has incited researchers to examine how personality characteristics and other variables operate jointly in the development of deviant eating patterns. OBJECTIVE: This study investigated the independent, interactive, and indirect prediction of dietary restraint by perfectionism and excessive commitment to exercise. METHOD: Multiple regression analyses designed to test moderating and mediating models were conducted on a sample of female university students (n = 269). RESULTS: Several dimensions of perfectionism, as well as excessive commitment to exercise, significantly and independently predicted dietary restraint in these women. There was no evidence for an interaction effect. Mediation analyses suggested that for selected dimensions of perfectionism, the direct relationship between perfectionism and dietary restraint is partially explained by excessive commitment to exercise. DISCUSSION: Interventions aimed at challenging perfectionistic standards in the context of dieting need to address not only one's self-standards, but one's perceptions of standards held by others. The mediating role of excessive exercise commitment pinpoints this variable as an alternative intervention target in the prevention of excessive dieting. PMID- 11262512 TI - Forbidden fruit: does thinking about a prohibited food lead to its consumption? AB - OBJECTIVE: The phenomenon of overeating the very foods that one is trying to resist is potentially consistent with both an ironic process account of overeating and a reactance account of the desire for "forbidden fruit." These two models are tested. METHOD: Participants in two studies were prohibited or not prohibited from eating a food, or they were encouraged to "choose" to avoid it. Food consumption, thoughts, and desire were assessed before and after the food was forbidden. RESULTS: Consistent with an ironic process account, participants' thoughts about the food increased, regardless of whether they were required to or chose to avoid it. Consistent with a reactance account, participants' desire for the food increased if they were required to avoid it, but not if they chose to avoid it. Participants did not, however, ultimately overeat the forbidden food. DISCUSSION: Neither increased thoughts nor enhanced desire for a food necessarily leads to overindulgence. PMID- 11262511 TI - Sexual orientation and eating psychopathology: the role of masculinity and femininity. AB - OBJECTIVE: Previous research suggests that eating disorders are related to homosexuality in men, although links with female sexual orientation are less clear. Appearance factors have generally been implicated in this relationship. However, previous studies have failed to consider the role of femininity, even though evidence suggests that this is a more critical factor than sexual preference. The aim of this study was to consider the relationship between gender role orientation and eating psychopathology in nonclinical men and women of different sexual orientations. METHOD: One hundred university students (40 homosexual; 60 heterosexual) completed the Bem Sex Role Inventory and the Eating Attitudes Test. RESULTS: For the group as a whole, there were links between femininity and high levels of eating psychopathology, whereas masculinity was associated with relatively healthy eating-related attitudes and behaviors. When considering the role of sexual orientation, these links were specific to homosexual men and women. CONCLUSIONS: In relation to homosexual men and women, the results support a model where femininity might be seen as a specific risk factor for eating disorders, whereas masculinity is likely to be a protective factor. Methodological and conceptual implications are discussed. PMID- 11262513 TI - Weight-related and shape-related self-evaluation in eating-disordered and non eating-disordered women. AB - OBJECTIVE: Weight- and shape-related self-evaluation refers to the process whereby an individual determines her self-worth based on an evaluation of her body weight and shape. This is a hallmark feature of both anorexia and bulimia nervosa, as specified in the 4th ed. of the Diagnostic and Statistical Manual of Mental Disorders. The purpose of this study was to further our understanding of weight-related self-evaluation in eating-disordered women. METHOD: Eating disordered patients, restrained eaters, and unrestrained eaters completed an experimenter-designed questionnaire that examines different dimensions of weight related self-evaluation (i.e., the Multidimensional Weight-Related Self Evaluation Inventory). RESULTS: Results revealed that weight-related self evaluation is a feature shared, to some extent, by both eating-disordered patients and restrained eaters. However, eating-disordered patients extend weight related self-evaluation to include more domains of self-esteem than did restrained eaters. DISCUSSION: These findings support a multidimensional approach to weight-related self-evaluation and further our understanding of the process of weight-related self-evaluation in eating-disordered patients. PMID- 11262514 TI - Changes in cue reactivity following treatment for bulimia nervosa. AB - OBJECTIVE: To examine changes in cue reactivity following cognitive-behavior therapy (CBT) for bulimia nervosa and to evaluate whether changes are associated with treatment modality or treatment outcome. METHOD: Subjects were 135 women (17 45 years old) with a current, primary diagnosis of bulimia nervosa. They were participants in a randomized clinical trial examining the additive efficacy of exposure and nonexposure-based behavior therapy to a core of CBT. Physiological, self-report, and behavioral measures of cue reactivity to individualized high risk binge foods were obtained at pretreatment and posttreatment. Primary, secondary, and tertiary outcome measures are reported for posttreatment. RESULTS: Bulimic patients experienced significant changes in cue reactivity following treatment. With the exception of salivary reactivity, patients experienced less reactivity at posttreatment. Changes in cue reactivity were not related to treatment modality, but were related to positive treatment outcome for self report measures of cue reactivity. DISCUSSION: Favorable treatment outcome among bulimic women is associated with low cue reactivity on self-report measures at posttreatment. PMID- 11262516 TI - Relationship between EDNOS and its subtypes and borderline personality disorder. AB - OBJECTIVE: The purpose of this study was to assess the prevalence of eating disorder not otherwise specified (EDNOS) and four well-defined subtypes of this disorder found in a sample of female borderline patients. METHOD: The lifetime prevalence of EDNOS and its various subtypes among 233 female borderline patients and 46 female Axis II comparison subjects was assessed using the Structured Clinical Interview for DSM-III-R Axis I disorders. RESULTS: Thirty-three percent of female borderline patients met DSM-III-R criteria for EDNOS at some point in their lives. Of these 76 women, 20% reported a pattern of restricting without low weight, 37% reported a pattern of binging without purging, 37% reported a pattern of purging without binging, and 33% reported a pattern of low weight without loss of menses. However, less than 25% of these 76 borderline women had ever met criteria for anorexia nervosa or bulimia nervosa. CONCLUSIONS: The results of this study suggest that EDNOS is a separate cluster of eating disorders among borderline women, rather than a prodromal or residual form of a more clear-cut case of anorexia or bulimia nervosa. PMID- 11262515 TI - Effect of inositol on bulimia nervosa and binge eating. AB - OBJECTIVES: This study aimed to determine whether inositol has therapeutic value in patients with bulimia nervosa and binge eating. METHOD: A double-blind crossover trial using 18 g inositol versus placebo was performed in 12 patients for 6 weeks in each arm. RESULTS: Inositol was significantly better than placebo on the Global Clinical Impression, the Visual Analogue Scale, and the Eating Disorders Inventory. DISCUSSION: Inositol is as therapeutic in patients with bulimia nervosa and binge eating as it is in patients with depression and panic and obsessive-compulsive disorders. This increases its parallelism with serotonin selective reuptake inhibitors. PMID- 11262517 TI - Perception of hunger to insulin-induced hypoglycemia in anorexia nervosa. AB - OBJECTIVE: We studied the effect of insulin-induced hypoglycemia on changes of hunger ratings in anorectic patients before and after cognitive-behavioral therapy. METHOD: The subjects were 17 females with restricting anorexia nervosa at low body weight (AN-R), 6 anorectic patients whose weight was restored after cognitive-behavioral therapy (AN-T), and 11 age-matched female controls. All subjects gave hunger ratings by linear visual analog technique before and after insulin or saline injection. RESULTS: Hunger ratings increased significantly 45 min after insulin injection in control females. However, ratings paradoxically decreased after insulin injection in AN-R females. They increased slightly after insulin injection in AN-T females, but the difference was not statistically significant. One-factor analysis of variance for the peak values of hunger ratings was significant. These values in control females were significantly higher than those in AN-R and AN-T females. DISCUSSION: These results suggest that perception of hunger to insulin-induced hypoglycemia in AN patients is disturbed. PMID- 11262519 TI - Bilateral osteonecrosis of the talus and "standing obsession" in a patient with anorexia nervosa. AB - METHOD: We report a case of a 24-year-old woman with anorexia nervosa (AN), obsessive-compulsive disorder (OCD), and osteoporosis. During 8 years of illness, her body mass index (BMI) fluctuated between 10 and 13,5 kg/m(2). She developed hyperactivity and the habit to stand for all daily activities. She did not allow herself to sit at any time, she had "standing obsession." A few weeks after her admission to an inpatient unit for eating disorders, she reported pain in both ankles. Results and Discussion The rheumatological diagnosis was bilateral osteonecrosis of the talus and generalized osteoporosis. The latter and excessive biomechanical stress could be interpreted as the cause for this serious and irreversible bone destruction. The following case shows how the psychiatric comorbidity of a patient suffering from an eating disorder with a chronic course can lead to severe somatic complications. PMID- 11262518 TI - Parental medical neglect in the treatment of adolescents with anorexia nervosa. AB - OBJECTIVE: Although childhood sexual abuse has been a frequent focus of research on eating disorders, other forms of maltreatment have been less commonly reported. Parental medical neglect is examined in this study as having serious consequences for the treatment and prognosis of patients with anorexia nervosa. METHOD: Two case studies illustrate parental interference with treatment in which Child Protective Services (CPS) had to be involved in compliance with state law. Two adolescent females who were admitted for treatment for anorexia nervosa are presented. RESULTS: In both cases, the parents refused to comply with the recommendations of the treatment team, placing their children's health in jeopardy. In compliance with reporting guidelines, CPS was notified in both cases. CONCLUSIONS: Clinicians who treat minors with anorexia nervosa must consider parental compliance with treatment. Indications for the involvement of CPS are outlined. Optimally, this notification can ensure that the patient and family receive the requisite treatment. PMID- 11262525 TI - Will systemic antivirals be sold over the counter? PMID- 11262527 TI - Prophylaxis for CMV should not now replace pre-emptive therapy in solid organ transplantation. AB - Pre-emptive therapy (PET) initiated on the basis of HCMV positivity in the blood using sensitive methods such as PCR, nucleic acid sequence based amplification or antigenaemia offers several advantages for the management of HCMV infection. These include the ability to target antiviral drug therapy to those most at risk of future disease, minimising drug exposure and maximising cost-benefit. In addition, allowing limited replication to occur also provides immune stimulation which will be important for future control of HCMV replication. In contrast, prophylaxis is a high-cost strategy which exposes all patients to potentially toxic drugs, does not facilitate immune priming and leads to the development of late HCMV infection and disease in high-risk patients. PMID- 11262526 TI - Prophylaxis for CMV should now replace pre-emptive therapy in solid organ transplantation. AB - Cytomegalovirus is a significant cause of morbidity and mortality in transplantation. Controversy exists concerning whether prophylactic or pre emptive therapy is the optimal strategy for preventing CMV disease. In addition, CMV impacts the transplanted graft, transplant recipient and transplant programme beyond just causing CMV disease; thus questioning whether 'asymptomatic' CMV replication should also be prevented. In this Forum article, prophylactic therapy is advocated as the preferred approach for preventing CMV disease. Prophylactic therapy has a large body of supportive controlled clinical studies demonstrating its efficacy and cost effectiveness. In addition, prophylactic therapy has the benefit of preventing other herpes viruses and other opportunistic superinfections by reducing the immunosuppressive effects of CMV. Moreover, a small but growing body of information suggests that prophylactic therapy may also have a beneficial effect on organ outcomes, including rejection. In contrast, pre emptive therapy is limited by its reliance on intensive surveillance, which presents logistical difficulties and requires perfect patient compliance. Ambiguity still exists concerning the best surveillance method and its effect on patient-care costs. Each proposed diagnostic approach has limitations, which are affected by the prevalence of CMV in the population studied, the particular assay employed, and the frequency of surveillance. The suggested benefits of pre emptive therapy, such as decreased cost, fewer adverse medication effects and less antiviral resistance have not been adequately proven in head-to-head clinical studies. We therefore support the proposition that transplant patients at risk for any level of CMV replication will significantly benefit from effective antiviral prophylaxis. PMID- 11262528 TI - Role of free radicals in viral pathogenesis and mutation. AB - Oxygen radicals and nitric oxide (NO) are generated in excess in a diverse array of microbial infections. Emerging concepts in free radical biology are now shedding light on the pathogenesis of various diseases. Free-radical induced pathogenicity in virus infections is of great importance, because evidence suggests that NO and oxygen radicals such as superoxide are key molecules in the pathogenesis of various infectious diseases. Although oxygen radicals and NO have an antimicrobial effect on bacteria and protozoa, they have opposing effects in virus infections such as influenza virus pneumonia and several other neurotropic virus infections. A high output of NO from inducible NO synthase, occurring in a variety of virus infections, produces highly reactive nitrogen oxide species, such as peroxynitrite, via interaction with oxygen radicals and reactive oxygen intermediates. The production of these various reactive species confers the diverse biological functions of NO. The reactive nitrogen species cause oxidative tissue injury and mutagenesis through oxidation and nitration of various biomolecules. The unique biological properties of free radicals are further illustrated by recent evidence showing accelerated viral mutation by NO-induced oxidative stress. NO appears to affect a host's immune response, with immunopathological consequences. For example, NO is reported to suppress type 1 helper T cell-dependent immune responses during infections, leading to type 2 helper T cell-biased immunological host responses. NO-induced immunosuppression may thus contribute to the pathogenesis of virus infections and help expansion of quasispecies population of viral pathogens. This review describes the pathophysiological roles of free radicals in the pathogenesis of viral disease and in viral mutation as related to both nonspecific inflammatory responses and immunological host reactions modulated by NO. PMID- 11262529 TI - Molecular epidemiology of respiratory syncytial virus. AB - Human respiratory syncytial virus (RSV) is the major cause of lower respiratory tract disease in infants. It is unusual in that it causes repeated infections throughout life. Despite considerable efforts there is as yet no satisfactory vaccine available. This paper reviews the molecular epidemiology of the RSV and describes the complex genotypic structure of RSV epidemics. The evolution of the virus is discussed, with particular reference to the antigenic and genetic variability of the attachment glycoprotein. PMID- 11262530 TI - Mucosal immune network in the gut for the control of infectious diseases. AB - The common mucosal immune system (CMIS) consists of an integrated cross communication pathway of lymphoid tissues made up of inductive and effector sites for host protection against pathogenic microorganisms. Major effector molecules of the CMIS include IgA antibodies and cytokines, chemokines and their corresponding receptors. Secretory IgA (S-IgA), the major immunoglobulin, is induced by gut-associated lymphoreticular tissue (GALT)-derived B cells with the help of Th1- and Th2-type CD4(+) T lymphocytes. Cytotoxic T lymphocytes (CTLs) in the mucosal epithelium, a subpopulation of intraepithelial lymphocytes (IELs), also help maintain the mucosal barrier. The CMIS is unique in that it can provide both positive and negative signals for the induction and regulation of immune responses in both the mucosal and systemic compartments after oral or nasal antigen exposure. Prevention of infection through mucosal surfaces can be achieved by the CMIS through connections between inductive (e.g. GALT) and effector tissues. When vaccine antigens are enterically administered together with mucosal adjuvants [e.g. cholera toxin (CT), heat-labile toxin produced by Escherichia coli (LT) and IL-12], antigen-specific Th1/Th2 and IgA B cell responses are induced simultaneously in the mucosal effector compartment. Since these antigen-specific immune responses are not generated by oral vaccine without mucosal adjuvant, safe and effective adjuvants for the induction of antigen specific S-IgA and CTL responses are essential for the development of mucosal vaccines for protection against infectious diseases. Finally, recent findings suggest the presence of a CMIS-independent IgA induction pathway, which also must be considered in the development of mucosal vaccines. PMID- 11262531 TI - Can the proximal isovelocity surface area method calculate stenotic mitral valve area in patients with associated moderate to severe aortic regurgitation? Analysis using low aliasing velocity of 10% of the peak transmitral velocity. AB - To assess the ability of the proximal isovelocity surface area (PISA) method to accurately measure the stenotic mitral valve area (MVA), and to assess whether aortic regurgitation (AR) affects the calculation, we compared the accuracy of the PISA method and the pressure half-time (PHT) method for determining MVA in patients with and without associated AR by using two-dimensional echocardiographic planimetry as a standard. The study population consisted of 45 patients with mitral stenosis. Seventeen of the 45 patients had associated moderate-to-severe AR. The PISA method was performed using low aliasing velocity (AV) of 10% of the peak transmitral velocity, which provided the most accurate estimation of MVA when compared with planimetry. The maximal radius r of the PISA was measured from the orifice to blue-red aliasing interface. Using the PISA method, MVA was calculated as (2pir(2)) x theta / 180 x AV/Vmax, where theta was the inflow angle formed by mitral leaflets, AV was the aliasing velocity (cm/sec), and Vmax was the peak transmitral velocity (cm/sec). MVA by the PISA method correlated well with planimetry both in patients with AR (r = 0.90, P < 0.001, SEE = 0.17 cm(2)) and without AR (r = 0.92, P < 0.001, SEE = 0.16 cm(2)). However, MVA by the PHT method did not correlate as well with planimetry (r = 0.57, P < 0.05, SEE = 0.37 cm(2)) in patients with associated AR, and the PHT method produced a significant overestimation (24%) of MVA obtained by planimetry in these patients. We conclude that the PISA method allows accurate estimation of MVA and is not influenced by AR. PMID- 11262532 TI - Eccentric mitral regurgitation jets among patients having sustained inferior wall myocardial infarction. AB - A strong association has been recognized between partial or complete mitral leaflet flail and highly eccentric mitral regurgitation jets. In light of anecdotal observation of eccentric mitral regurgitation apparently due to geometric and functional changes accompanying inferior wall myocardial infarction, the present study was performed to systematically study the eccentricity of mitral regurgitation jets complicating nonacute inferior wall myocardial infarction. Forty-eight consecutive patients with evidence of prior isolated inferior wall myocardial infarction and at least moderate mitral regurgitation but without other valvular, annular, chordal, or ventricular pathology potentially contributory to mitral regurgitation were studied. Mitral regurgitation jets were characterized with respect to eccentricity and anterior versus posterior direction. Regurgitant jet and mitral leaflet position were quantified relative to the mitral annulus. Five of 48 patients (10.4%) had eccentric jets, of which four were directed posterior and one anterior. Although not reaching statistical significance, patients with eccentric jets tended to have somewhat smaller left atrial size (41.2 +/- 7.8 vs 47.2 +/- 9.3 mm, P = 0.17) and left ventricular size (51.5 +/- 3.4 vs 55.1 +/- 7.8 mm, P = 0.13), and higher left ventricular ejection fraction (0.52 +/- 0.11 vs 0.46 +/- 0.09, P = 0.25) compared with patients with noneccentric jets. Leaflet position relative to the mitral annulus was significantly different among patients with eccentric compared with noneccentric posterior jets (54 +/- 10 degrees vs 33 +/- 11 degrees, P = 0.02), implying greater leaflet restriction toward the left ventricular apex. In conclusion, approximately one in 10 patients with isolated inferior wall myocardial infarction and at least moderate mitral regurgitation was found to have marked eccentricity of the regurgitant jet. Leaflet position was more apically displaced among patients with eccentric jets, suggesting greater leaflet restriction in systole. The finding of a highly eccentric posterior mitral regurgitation jet can be due to inferior wall myocardial infarction with posterior leaflet restriction as well as partial or complete anterior mitral leaflet flail. PMID- 11262533 TI - Prognostic implications of right atrial ischemic dysfunction in patients with biventricular inferior infarction: transesophageal echocardiographic analysis. AB - In order to determine the effect of right atrial dysfunction on clinical outcome, six patients with inferior myocardial infarction with extension to right ventricle and right atrium involving only obstructions of the right coronary artery were examined with transesophageal echocardiography (TEE) at the time of the event. Five of the patients were reexamined 15 to 55 months later. Two patients underwent thrombolysis and maintained ratios of right-to-left ventricular diameters of less than 1, as well as normal convexity of the interatrial septum. One patient had spontaneous reperfusion of the right coronary artery, reduction in right ventricular diameter, and normalization of interatrial septum. Another patient underwent delayed angioplasty and manifested a diminished wall movement score (WMS) in the follow-up echocardiogram. One patient died during his first hospitalization with significant right ventricular dilatation, inverted convexity of the interatrial septum, and right atrial thrombosis. The last patient died during follow-up with right ventricular dilatation, increased WMS, right atrial akinesis, and inverted interatrial convexity. Serial TEE examination of patients with infarction of the left ventricular inferior wall is a safe technique for determining the degree of the extension of the ischemic process to the right chambers. PMID- 11262534 TI - Lack of agreement between left ventricular volumes and ejection fraction determined by two-dimensional echocardiography and contrast cineangiography in postinfarction patients. AB - OBJECTIVE: To assess the agreement between left ventricular (LV) volumes and ejection fraction (EF) determined by two-dimensional echocardiography (2-D echo) and by cineangiography in postinfarction patients. DESIGN: LV end-diastolic and end-systolic volumes indexed (EDVI and ESVI) to body surface area as well as EF were determined by both methods in all patients. SETTING: Multicenter trial conducted in five university hospitals. PATIENTS: 63 patients, 61 male, two female, mean age 55.5 +/- 10.4 years, suffering from a recent myocardial infarction. Eighty-one pairs of measurements were available. METHODS: The results of biplane 2-D echo measures, using apical four-chamber (4C) and two-chamber (2C) views were compared to those of a 30 degrees right anterior oblique cineangiography projection, using either the apical method of discs or the area length 2-D echo method. Moreover, eyeball EF was estimated at 2-D echo and cineangiography, and was compared to the conventional methods. The agreement between results was assessed by the Bland and Altman method. RESULTS: The agreement between 2-D echo and cineangiography results was poor. Mean differences (MD) were -21.8 (EDVI, ml/m(2)), -9.5 (ESVI, ml/m(2)), and -0.9 (EF, %), respectively for 2-D echo method of discs versus cineangiography, and -23.2, 9.3, and -5.7 for area-length 2-D echo versus cineangiography. For EF (%), MD was -3.6 for eyeball cineangiography versus cineangiography, -1.3 for eyeball 2-D echo versus method of discs, and +0.30 for eyeball 2-D echo versus area-length 2 D echo, respectively. Two-dimensional echo is likely to underestimate LV volumes compared to cineangiography, especially for largest volumes. Even for EF, discrepancies are large, with a lack of agreement of 21%-25% between conventional methods, but agreement is better between eyeball EF and usual methods. CONCLUSIONS: Even with modern echocardiographic devices, agreement between 2-D echo and cineangiography-derived LV volumes and EF remains moderate, and both methods must not be considered interchangeable in clinical practice. PMID- 11262535 TI - Effects of verapamil in normal elderly individuals with left ventricular diastolic dysfunction. AB - Treatment with oral verapamil for 3 to 4 days has been found to enhance left ventricular (LV) diastolic filling in elderly subjects as assessed by radionuclide angiography. However, there are no Doppler echocardiographic studies to assess the long-term effect of verapamil in normal elderly subjects. Thirteen healthy elderly subjects (mean age, 64 +/- 7 years; 8 men and 5 women) with LV diastolic dysfunction underwent this placebo-controlled cross-over trial. The effect of verapamil on LV diastolic function was assessed by treadmill exercise test and Doppler echocardiography at baseline, and after each 3-month treatment period (placebo or verapamil 120 mg once daily), separated by a 1-week washout period before cross-over. Blood pressure, heart rate, LV ejection fraction, LV mass, and cardiac output were unaltered by placebo or verapamil. The exercise time was similar at baseline (11.4 +/- 2.4 min) and after placebo treatment (11.4 +/- 2.3 min) but significantly increased (P < 0.05) after verapamil treatment (12.3 +/- 2.0 min). The ratio of mitral A wave duration/pulmonary venous atrial systolic reversal duration increased after verapamil treatment (1.12 +/- 0.08) compared to placebo (0.93 +/- 0.06, P < 0.05) and baseline (0.89 +/- 0.09), which had similar durations. The isovolumic relaxation time (IVRT) was significantly decreased (P < 0.05) from 85 +/- 13 msec at baseline and 87 +/- 13 msec with placebo to 73 +/- 9 msec with verapamil. The results of this study suggest that in normal elderly patients with Doppler evidence of diastolic dysfunction, 3 months treatment with verapamil can increase exercise tolerance and improve LV diastolic function. PMID- 11262536 TI - Doppler tissue imaging to evaluate early myocardium damage in patients with undetermined form of Chagas' disease and normal echocardiogram. AB - Chagas' disease can lead to severe and potentially lethal damage of cardiac function. Thus, the identification of contractile abnormalities in asymptomatic patients can be important for risk stratification in those patients. Doppler tissue imaging (DTI) is a new diagnostic modality for the study of regional and longitudinal contractility of the left ventricle. However, DTI has not been used for the assessment of Chagas' cardiopathy. The purpose of this study was to identify abnormalities related to longitudinal contractility by means of DTI in patients with an undetermined form of Chagas' disease and with normal echocardiogram and Doppler studies. Forty patients were studied, including 21 chagasic ones with normal electrocardiographic, radiologic, and echocardiographic studies, and 19 normal control individuals. All of the patients were submitted to DTI to evaluate longitudinal contractility in the various myocardial segments, including assessment of systolic and diastolic velocities and isovolumic contraction time (IVCT). Similar values were observed among the various systolic function rates in both groups, except for the IVCT along the septal wall, which was significantly higher in the chagasic group. DTI enabled the early detection of contractile abnormalities in patients with an undetermined form of Chagas' disease and with normal echocardiogram. Such abnormalities were particularly striking in the interventricular septum. The present study could be useful in risk stratification for those patients. PMID- 11262537 TI - Proximal isovelocity surface area (PISA) as a noninvasive method for the estimation of the shunt quantification in perimembranous ventricular septal defects. AB - This study was designed to assess the reliability of the proximal isovelocity surface area (PISA) method for the estimation of shunt quantification in perimembranous ventricular septal defects (PVSD). The study group was composed of 30 patients (age 11 +/- 7 years, 13 female) with PVSD. The shunt flow (Qp-Qs) and the ratio of the pulmonary flow to the systemic flow (Qp/Qs) were calculated by spectral Doppler and catheterization. The Qp-Qs, the defect area (DA), and the shunt volume (SV) were obtained by the PISA method. The PISA method estimated the DA (cm(2)/m(2)), the SV (cm(3)/m(2)), and the Qp-Qs (L/min/m(2)) to be equal to (2 x pi x R(2) x NL)/(V(max) x Body surface area), DA x TVI(shunt), and to SV x Heart rate, respectively (R is the distance of the maximal PISA from the first aliasing line to the left ventricular side of the defect, NL is the nyquist limit, and V(max) and TVI(shunt) are the peak velocity and time-velocity integral of transdefect Doppler tracing obtained by continuous-wave Doppler). The PISA method (3.4 +/- 1.5 L/min/m(2)) underestimated the Qp-Qs according to spectral Doppler (r = 0.96, P < 0.001; mean difference -0.74 +/- 0.61 L/min/m(2); SEE = 0.11 L/min/m(2), P < 0.001) and catheterization (r = 0.92, P < 0.001; mean difference -0.45 +/- 0.7 L/min/m(2); SEE = 0.13 L/min/m(2), P < 0.001). The correlations between the PISA findings (Qp-Qs, DA, SV) and the catheterization Qp/Qs (r = 0.86, 0.84, and 0.86; P < 0.001, respectively), or between these and the spectral Doppler Qp/Qs (r = 0.80, 0.80, and 0.79; P < 0.001, respectively) were significant. The accuracies of the PISA findings in identifying large defects were high (0.90, 0.93, and 0.90 for cut-off values of Qp-Qs = 3.67 L/min/m(2), DA = 0.44 cm(2)/m(2), and SV = 43 cm(3)/m(2), respectively). As a result, the PISA method can be a simple and reliable alternative to the spectral Doppler method in the identification of large shunts in PVSD. PMID- 11262538 TI - In vitro measurement accuracy of three-dimensional ultrasound. AB - OBJECTIVES: We sought to validate distance and volume measurements in three dimensional (3-D) ultrasound images. BACKGROUND: Even with the latest equipment, it is not known how accurate 3-D echocardiographic measurements are. METHODS: Six models were imaged in ethanol solution and two within a tissue phantom using a mechanical rotation device rotating in 1 degrees intervals and a real-time 3-D scanner. Distance and volume measurements (n = 60) were performed in two dimensional (2-D) and 3-D images using TomTec and InViVo software. RESULTS: Distance measurements had a mean total error between 1.12% and 2.31% for Acuson (2.5 MHZ, 3 MHZ, and 4 MHZ) and Hewlett Parkard (HP) fusion frequencies h and m, HP fusion harmonic B in the axial, and between 3.5% and 4.9% in the lateral dimension. HP Harmonic A and B, Volumetrics (2.5 MHZ), and HP fusion Harmonic A exhibited significantly higher differences to reality with a mean difference between 5.1% and 8.9% in the axial and between 6.2% and 7.9% in the lateral direction. Axial 2-D measurements were not different from real dimensions except Volumetrics model 1. In the lateral axis, all imaging modalities were different from reality except the fusion harmonic modus B. Using the HP fusion frequency h and HP fusion Harmonic B-mode, volume measurements in 3-D images significantly underestimated reality, while Acuson's fundamental frequency 3.5 MHZ was not different from real volumes. CONCLUSION: Three-dimensional visualization using different ultrasound settings results in different accuracy. PMID- 11262539 TI - Rapid formation of left atrial appendage thrombus after unsuccessful cardioversion of atrial fibrillation. AB - The acute effect of failed attempts of cardioversion on left atrial (LA) and left atrial appendage (LAA) functions are generally considered benign and no adverse effects have been reported. We report on a subject who had rapid formation of a fresh, mobile thrombus in the LAA despite unsuccessful cardioversion and therapeutic anticoagulation. PMID- 11262540 TI - Left atrial and ventricular ball thrombi complicating rheumatic heart disease with combined mitral and aortic stenosis. AB - A 42-year-old woman with chronic mitral stenosis was admitted for progressive dyspnea, palpitation, and weakness of lower extremities. Echocardiography revealed rheumatic, thickened, and stenotic mitral and aortic valves, and two free-floating ball thrombi were detected in the left atrium and ventricle, respectively. She died suddenly the next day, probably due to mitral or aortic outflow obstruction by the ball thrombi. We believe that the occurrence of free floating ball thrombi in both the left atrium and left ventricle concomitantly has never been reported. PMID- 11262541 TI - Three-dimensional echocardiography of left atrial appendage thrombus. AB - Transesophageal echocardiography (TEE) is superior to transthoracic echocardiography (TTE) in evaluating the left atrial appendage (LAA) as a potential cardiac source of embolus in patients with stroke. We describe two such patients in whom a TEE and subsequently a three-dimensional (3-D) reconstruction of the LAA were performed. These case reports show that 3-D echocardiography provides better visualization of LAA anatomy, and that a 3-D description of LAA morphology may be the basis of describing normal and abnormal LAA. PMID- 11262542 TI - Myocardial contusion presented as acute myocardial infarction after chest trauma. AB - A 46-year-old male patient developed an acute myocardial infarction and congestive heart failure following blunt chest trauma. Electrocardiogram (ECG) revealed acute anterior myocardial infarction. Echocardiography showed akinesis of interventricular septum, dyskinesis in apical anterior wall, and severe impairment of left ventricular overall systolic function. Coronary angiography revealed normal coronary arteries. The patient followed a low-intensity physical medicine rehabilitation program. Follow-up was without new complications or deterioration of congestive heart failure. Five months later the patient presented with fulminant acute pulmonary edema and cardiogenic shock. Cardiopulmonary resuscitation was unsuccessful. PMID- 11262543 TI - Squamous cell carcinoma masquerading as a thoracic intramural hematoma. AB - Transesophageal echocardiography (TEE) is an excellent tool for the diagnosis of thoracic aortic pathology. However, not all lesions of the aortic wall represent primary pathology of the aorta. This case presents an elderly, hypertensive patient with back pain and crescentic thickening of the aortic wall that proved to represent external aortic infiltration by lung cancer. Invasion of the aortic wall mimicking intramural hematoma should be considered the differential diagnosis of patients with dissection-like symptoms and crescentic aortic lesions. PMID- 11262544 TI - Discovery of a parachute mitral valve complex (Shone's anomaly) in an adult. AB - Parachute mitral valve complex is an unusual congenital anomaly described by Shone et al. It is characterized by a parachute deformity of the mitral valve associated with additional forms of left heart anomalies, such as aortic valvular stenosis and coarctation of the aorta. Fewer than 50 cases of Shone's complex have been reported in the literature, and it has only been observed in children. We report the case of a 33-year-old man who was referred to our department because of atrial fibrillation. Echocardiographic evaluation and aortogram evidenced a Shone's complex, including a parachute mitral valve anomaly, an aortic bicuspid valvular anomaly, and a coarctation of the aorta. PMID- 11262545 TI - Aorto-right ventricular outflow tract fistula: presentation 1 year after stab wound to the heart. PMID- 11262546 TI - Right coronary artery to left ventricle fistula. PMID- 11262547 TI - Transesophageal echocardiographic diagnosis of traumatic rupture of the noncoronary cusp of the aortic valve. AB - We report a patient with traumatic aortic valve injury in whom a large defect in the noncoronary cusp of the aortic valve was clearly visualized by multiplane TEE and confirmed at surgery. PMID- 11262548 TI - Recertification of respiratory therapists' intubation skills one year after initial training: an analysis of skill retention and retraining. AB - Allied health personnel and nonanesthesiologist physicians often undergo training in tracheal intubation but then may actually use the skill relatively infrequently. This study assessed retention of skills one year after initial training and identified specific areas of knowledge critical to successful performance of intubation. Eleven respiratory therapists on the staff of a 253 bed hospital, each of whom had been trained one year previously in airway management, were evaluated. Prior to returning to the operating room for skills assessment and recertification, each respiratory therapist took a 21-question written exam. Therapists then went to the operating room and a trained observer (anesthesiologist) monitored the intubations performed to see whether critical steps were followed, while a second observer monitored a checklist of skills performed. The attending anesthesiologist recertified the therapist only when all steps were correctly performed and the intubation was successful. There was a poor correlation (r = -0.25, p > 0.1) between the number of intubations performed by the therapists for emergencies in the previous year and the number of intubations needed to be recertified. There was a negative correlation (r = -0.8, p < 0.05) between the score on the written test and the number of intubations required for recertification-a higher score meant fewer intubations were needed to achieve recertification. First-pass success occurred significantly more frequently if all skills tested were performed correctly (50/75 first-pass successes had all skills performed correctly vs 10/28 for failed first-pass, p < 0.01). The most common errors were levering the blade on the upper teeth (12/91) and tube not inserted from the right side of the mouth (28/104). When the blade was levered, 8 of 10 intubations failed. When the tube was not inserted from the right side of the face, 6 of 12 failed. The useful findings of this study are: (1) occasional performance of intubation did not ensure skill maintenance; (2) cognitive and procedural abilities correlated, suggesting benefits to study as well as to practical training; and (3) two specific mistakes were associated with a high incidence of failure. PMID- 11262549 TI - The impact of turnover among respiratory care practitioners in a health care system: frequency and associated costs. AB - BACKGROUND: Retention of respiratory therapists (RTs) is a desired institutional goal that reflects department loyalty and RTs' satisfaction. When RTs leave a department, services are disrupted and new therapists must undergo orientation and training, which requires time and expense. Despite the widely shared goal of minimal turnover, neither the annual rate nor the associated expense of turnover for RTs has been described. STUDY PURPOSE: Determine the rate of RT turnover and the costs related to training new staff members. METHODS: The Cleveland Clinic Health System is composed of 9 participating hospitals, which range from small, community-based institutions to large, tertiary care institutions. To elicit information about annual turnover among RTs throughout the system, we conducted a survey of key personnel in each of the hospitals' respiratory therapy departments. To calculate the costs of training, we reviewed the training schedule for an RT joining the Respiratory Therapy Section at the Cleveland Clinic Hospital. Cost estimates reflect the duration of training by various supervisory RTs, their respective wages (including benefit costs), and educational materials used in training and orientation. RESULTS: Turnover rates ranged from 3% to 18% per year. Five of the 8 institutions from which rates were available reported rates greater than 8% per year. The rate of annual turnover correlated significantly with the ratio of hospital beds to RT staff (Pearson r = 0.784, r(2) = 0.61, p = 0.02). The cost of training an RT at the Cleveland Clinic Hospital totaled $3,447.11. CONCLUSIONS: Turnover among respiratory therapists poses a substantial problem because of its frequency and expense. Greater attention to issues affecting turnover and to enhancing retention of RTs is warranted. PMID- 11262550 TI - Assessment of aspiration in patients with tracheostomies: comparison of the bedside colored dye assessment with videofluoroscopic examination. AB - BACKGROUND: Aspiration is a serious clinical concern in patients with long-term artificial airways. The purpose of this study was to determine the reliability of a bedside colored dye assessment of aspiration in tracheostomized patients and to determine its comparability to a more sophisticated videofluoroscopic study. METHODS: This was a prospective, blinded comparison study conducted in a large, urban, university teaching hospital. We studied 20 consecutive patients who underwent tracheostomy for bronchial hygiene needs and who were referred for videofluorographic evaluation for suspected oropharyngeal dysphagia and possible aspiration. Excluded were patients unable to follow verbal commands and those requiring mechanical ventilatory support. All patients were brought to the videofluorography suite for colored dye assessment for aspiration and videofluorographic assessment of oropharyngeal swallow. A nurse, blinded to the results of videofluorographic swallow study, performed colored dye assessments for aspiration. Speech-language pathologists, blinded to the results of the colored dye assessments, interpreted simultaneous (preliminary) and subsequent complete (final) videofluorographic evaluations of swallow. RESULTS: The colored dye aspiration assessments and the videofluoroscopic studies were compared for the frequency of aspiration detection. Sensitivity and specificity were determined using standard methods. Seven patients showed no aspiration on either the colored dye test or videofluoroscopic examination. Eight patients were judged to aspirate by videofluorography but not by the colored dye test. Five patients were judged to aspirate by both the colored dye test and videofluorography. The data indicate that the colored dye test for aspiration carries a low sensitivity of 38% (95% confidence interval = +/- 7%), but a high specificity of 100%. The videofluoroscopic study detected a significantly greater frequency of aspiration than did the colored dye test (p < 0.01). CONCLUSIONS: The colored dye test for aspiration can provide useful information when positive, but because there is a significant false negative rate, decisions made on the basis of a negative test must be made with caution. PMID- 11262551 TI - Heliox delivery with noninvasive positive pressure ventilation: a laboratory study. AB - BACKGROUND: There is clinical interest in the use of heliox (helium-oxygen mixture) during noninvasive positive pressure ventilation (NPPV), but delivery of heliox with ventilators designed for NPPV has not been reported. We studied helium concentration ([He]) when an 80%:20% helium:oxygen mixture (heliox) was used with 5 NPPV ventilators (Knightstar, Quantum, BiPAP S/T-D30, Sullivan, and BiPAP Vision). METHODS: A simulated spontaneous breathing lung model was connected to the ventilators with a circuit incorporating a standard leak. Heliox flows of 0, 5, 10, and 18 L/min and oxygen flows of 0 and 10 L/min were titrated into the system at either a proximal position near the lung model or a distal position near the ventilator (titration method). Because the BiPAP Vision has an oxygen delivery module, it was also studied using heliox connected to the air inlet of an oxygen blender, with the blender outlet connected to the oxygen module of the ventilator (blender method). All ventilators were evaluated in spontaneous/timed mode at inspiratory/expiratory pressures of 10/5, 15/5, and 20/5 cm H(2)O. After 5 minutes, [He], oxygen concentration, and pressure in the lung model were recorded. RESULTS: Heliox flow, NPPV settings, site of heliox infusion, and type of ventilator significantly (p < 0.05) affected [He]. [He] was > 60% when heliox flow was 18 L/min in some combinations of settings. The BiPAP S/T-D30 and Quantum occasionally functioned erratically. The BiPAP Vision (blender method) ventilator performed erratically with heliox unless the exhalation port test was bypassed on startup. The addition of heliox flow had no important effect on inspiratory or expiratory positive airway pressure on those breaths during which the ventilators functioned correctly. CONCLUSION: Heliox flow was the most important determinant of [He] when using heliox with NPPV. With heliox there was a potential for ventilator malfunction in some conditions. The clinical implications of these findings remain to be determined. PMID- 11262552 TI - Patient-ventilator interactions during volume-support ventilation: asynchrony and tidal volume instability--a report of three cases. AB - During pressure-support ventilation, tidal volume (V(T)) can vary according to the level of the patient's respiratory effort and modifications of the thoraco pulmonary mechanics. To keep V(T) as constant as possible, the Siemens Servo 300 ventilator proposes an original modification of pressure-support ventilation, called volume-support ventilation (VSV). VSV is a pressure-limited mode of ventilation that uses V(T) as a feedback control: the pressure support level is continuously adjusted to deliver a preset V(T). Thus, the ventilator adapts the inspiratory pressure level, breath by breath, to changes in the patient's inspiratory effort and the mechanical thoraco-pulmonary properties. The clinician sets V(T) and respiratory frequency, and the ventilator calculates a preset minute volume. It has been shown that ineffective respiratory efforts can occur during pressure-support ventilation. PMID- 11262553 TI - Weaning to extubation directly from high-frequency oscillatory ventilation in an infant with cystic lung disease and persistent air leak: a strategy for lung protection. AB - We report the successful weaning and extubation of an infant from a SensorMedics 3100A high-frequency oscillator without returning to conventional ventilation. A 7-week-old term infant with respiratory syncytial virus bronchiolitis complicated by cystic pulmonary lesions repeatedly failed attempts to return to conventional ventilation from high-frequency oscillatory ventilation (HFOV) for weaning, because of recurrent pneumothoraces. A computed tomography of the chest revealed multiple well defined cysts of various sizes involving both lungs. Therefore, weaning to extubation from HFOV was proposed as a way of preventing further air leak. The weaning strategy consisted of a technique we refer to as "sprinting." Using this method, the patient was successfully extubated directly from HFOV, with no complications. A follow-up computed tomography of the chest showed marked improvement in the size of the cystic lesions. The patient was discharged home with no need for home oxygen therapy. PMID- 11262554 TI - Metabolic acidosis. AB - Metabolic acidosis occurs in a number of diseases and even certain normal activities such as heavy exercise. It arises from increased endogenous acid production, exogenous acid (or acid-precursor) administration, base losses, and depression of renal acid secretion. Although the magnitude of acidosis is important, the ultimate pathophysiological impact of any metabolic acidosis is defined by the rate of change and the specific cause of the acidosis. This review discusses whole body, organ, and cellular effects of metabolic acidosis, its diagnosis by pathophysiologic categories, and treatment. The diagnosis is made by a synthesis of the clinical history, physical examination, other hematological values, serum and urinary chemistries, and arterial blood gases and electrolytes. Calculation of the anion and osmolal gaps can be effectively used to further narrow the diagnostic possibilities. Supportive care and therapy directed at the cause of the metabolic acidosis are the mainstays of treatment, since most acidotic states will spontaneously correct once the initiating cause is removed or reversed. Theoretical and clinical evidence are discussed for alkalinizing agents, whose use remains controversial except in the treatment of metabolic acidosis associated with hyperkalemia and certain drug or toxin ingestions. PMID- 11262556 TI - Respiratory acidosis. AB - Respiratory acidosis, or primary hypercapnia, is the acid-base disorder that results from an increase in arterial partial pressure of carbon dioxide. Acute respiratory acidosis occurs with acute (Type II) respiratory failure, which can result from any sudden respiratory parenchymal (eg, pulmonary edema), airways (eg, chronic obstructive pulmonary disease or asthma), pleural, chest wall, neuromuscular (eg, spinal cord injury), or central nervous system event (eg, drug overdose). Chronic respiratory acidosis can result from numerous processes and is typified by a sustained increase in arterial partial pressure of carbon dioxide, resulting in renal adaptation, and a more marked increase in plasma bicarbonate. Mechanisms of respiratory acidosis include increased carbon dioxide production, alveolar hypoventilation, abnormal respiratory drive, abnormalities of the chest wall and respiratory muscles, and increased dead space. Although the symptoms, signs, and physiologic consequences of respiratory acidosis are numerous, the principal effects are on the central nervous and cardiovascular systems. Treatment for respiratory acidosis may include invasive or noninvasive ventilatory support and specific medical therapies directed at the underlying pathophysiology. PMID- 11262555 TI - Metabolic alkalosis. AB - Metabolic alkalosis is a primary pathophysiologic event characterized by the gain of bicarbonate or the loss of nonvolatile acid from extracellular fluid. The kidney preserves normal acid-base balance by two mechanisms: bicarbonate reclamation, mainly in the proximal tubule, and bicarbonate generation, predominantly in the distal nephron. Bicarbonate reclamation is mediated mainly by a Na(+)-H(+) antiporter and to a smaller extent by the H(+)-ATPase (adenosine triphosphate-ase). The principal factors affecting HCO3(-) reabsorption include effective arterial blood volume, glomerular filtration rate, chloride, and potassium. Bicarbonate regeneration is primarily affected by distal Na(+) delivery and reabsorption, aldosterone, arterial pH, and arterial partial pressure of carbon dioxide. To generate metabolic alkalosis, either a gain of base or a loss of acid must occur. The loss of acid may be via the gastrointestinal tract or via the kidney. Excess base may be gained by oral or parenteral HCO3(-) administration or by lactate, acetate, or citrate administration. Factors that help maintain metabolic alkalosis include decreased glomerular filtration rate, volume contraction, hypokalemia, hypochloremia, and aldosterone excess. Clinical states associated with metabolic alkalosis are vomiting, mineralocorticoid excess, the adrenogenital syndrome, licorice ingestion, diuretic administration, and Bartter's and Gitelman's syndromes. The effects of metabolic alkalosis on the body are variable and include effects on the central nervous system, myocardium, skeletal muscle, and liver. Treatment of this disorder is simple, once the pathophysiology of the cause is delineated. Therapy consists of reversing the contributory factors that are promoting the alkalosis and, in severe cases, administration of carbonic anhydrase inhibitors, acid infusion, and low bicarbonate dialysis. PMID- 11262557 TI - Respiratory alkalosis. AB - Respiratory alkalosis is an extremely common and complicated problem affecting virtually every organ system in the body. This article reviews the various facets of this interesting problem. Respiratory alkalosis produces multiple metabolic abnormalities, from changes in potassium, phosphate, and calcium, to the development of a mild lactic acidosis. Renal handling of the above ions is also affected. The etiologies may be related to pulmonary or extrapulmonary disorders. Hyperventilation syndrome is a common etiology of respiratory alkalosis in the emergency department setting and is a diagnosis by exclusion. There are many cardiac effects of respiratory alkalosis, such as tachycardia, ventricular and atrial arrhythmias, and ischemic and nonischemic chest pain. In the lungs, vasodilation occurs, and in the gastrointestinal system there are changes in perfusion, motility, and electrolyte handling. Therapeutically, respiratory alkalosis is used for treatment of elevated intracranial pressure. Correction of a respiratory alkalosis is best performed by correcting the underlying etiology. PMID- 11262558 TI - Approach to patients with acid-base disorders. AB - Disorders of acid-base balance are commonly encountered in clinical practice and can have a substantial impact on the prognosis of the patient. Moreover, identification of a particular acid-base disturbance can provide a clue to an underlying disorder. Proper evaluation and treatment of acid-base disorders requires a systematic and analytic approach including: (1) assess the accuracy of the acid-base values using the Henderson equation or Henderson-Hasselbalch equation, (2) obtain a complete history and physical examination, (3) calculate the serum anion gap, (4) identify the primary acid-base disturbance and determine whether a simple or mixed disturbance is present, (5) examine serum electrolytes and additional laboratory data, and (6) measure urine pH and urine electrolytes and calculate the urine anion and osmolal gaps. Strict adherence to these principles will enable the clinician to diagnose the acid-base disturbance in the majority of cases. To illustrate these principles, 5 cases of patients with acid base disturbances are analyzed. PMID- 11262566 TI - [Radiographic clinical correlation - Case 2/2001 - Instituto do Coracao do Hospital das Clinicas da FMUSP ]. PMID- 11262568 TI - Changes in the profile of lipoprotein subfractions associated with hormone replacement therapy. AB - OBJECTIVE: To report the effects of 2 regimens of hormone replacement therapy during the postmenopausal period on the profile of the major lipoprotein subfractions (HDL, LDL, and VLDL). METHODS: We carried out a cohort study in 38 postmenopausal patients who were starting their hormone replacement therapy due to gynecological indications, for a period of 12 weeks. Analysis of lipoprotein subclasses was performed through nuclear magnetic resonance spectroscopy. RESULTS: Hormone replacement therapy cause an increase in the proportion of larger subfractions of VLDL and HDL (p=0.008 and 0.03, respectively) and in the proportion of larger particles of VLDL due to a 36% increase in the levels of larger particles (p=0.004), concomitantly with a 15% reduction in the levels of smaller particles (p=0.04). In regard to HDL, the increase occurred only a 17% increase in the levels of larger particles (p=0.002). No significant change occurred in the distribution pattern of LDL subfractions. CONCLUSION: The proportion of larger subfractions of VLDL and HDL increases after hormone replacement therapy. The increase in the proportion of larger particles of VLDL occurs due to an increase in the levels of the larger subclasses concomitantly with a reduction in the smaller particles. However, an increase in the proportion of larger particles of HDL occurs only due to an increase in the levels of the larger subfractions. PMID- 11262569 TI - Contribution of Doppler echocardiography to the evaluation of systolic and diastolic function of obese women versus a control group. AB - OBJECTIVE: To study by doppler echocardiography the cardiac systolic and diastolic functions of health, uncomplicated obese subjects. METHODS: Fifty-nine obese women with an average body mass index (BMI) of 35 kg/m2 were evaluated and compared with 19 subjects with an average BMI of 23 kg/m2 (control group). RESULTS: In the obese group, a clear tendency was observed toward higher systolic pressure, increased wall thickness and, consequently, myocardial mass, elevation on the circumference stress of the left ventricular wall, and an indisputable presence of diastolic abnormalities. Filling abnormalities were observed with impaired relaxation, with prolonged isovolumic relaxation time (IVRT) and augmented atrium contribution representing early indexes of cardiac dysfunction when systolic performance is still normal. CONCLUSION: Obesity is generally a chronic condition, and doppler echocardiography can be used as a noninvasive instrument for early evaluation of left ventricular diastolic indexes. PMID- 11262570 TI - Hypertension in a female nursing staff--Pattern of occurrence, diagnosis, and treatment. AB - OBJECTIVE: To report the pattern of occurrence, diagnosis, and treatment of hypertension in a female nursing staff of an emergency hospital. METHODS: We carried out a cross-sectional study that included interviews and blood pressure measurements of 494 nursing professionals at an emergency hospital in the city of Salvador, in the state of Bahia, Brazil. We considered hypertensive all individual with blood pressure > or = 140/90 mmHg or normal pressure if on regular treatment. RESULTS: We found a prevalence of hypertension of 36.4%. Only 18.3% of the individuals ignored their hypertensive condition, and 64.2% admitted not being having regular treatment. Of those individuals who were having treatment, 69.4% had elevated blood pressure on examination. The major reasons for not being on treatment was the occasional elevation of blood pressure (22.2%) and medical counseling (20.0%). CONCLUSION: The results point to the need to introduce hypertension control measures in this occupational group, because of the magnitude of the disease and the potential impact on diffusion of knowledge and measures to control hypertension. PMID- 11262571 TI - Unsupported valvuloplasty in children with congenital mitral valve anomalies. Late clinical results. AB - OBJECTIVE: To analyze late clinical evolution after surgical treatment of children, with reparative and reconstructive techniques without annular support. METHODS: We evaluated 21 patients operated upon between 1975 and 1998. Age 4.67+/ 3.44 years; 47.6% girls; mitral insufficiency 57.1% (12 cases), stenosis 28.6% (6 cases), and double lesion 14.3% (3 cases). The perfusion 43.10+/-9.50 min, and ischemia time were 29.40+/-10.50 min. The average clinical follow-up in mitral insufficiency was 41.52+/-53.61 months. In the stenosis group (4 patients) was 46.39+/-32.02 months, and in the double lesion group (3 patients), 39.41+/-37.5 months. The echocardiographic follow-up was in mitral insufficiency 37.17+/-39.51 months, stenosis 42.61+/-30.59 months, and in the double lesion 39.41+/-37.51 months. RESULTS: Operative mortality was 9.5% (2 cases). No late deaths occurred. In the group with mitral insufficiency, 10 (83.3%) patients were asymptomatic (p=0.04). The majority with mild reflux (p=0.002). In the follow-up of the stenosis group, all were in functional class I (NYHA); and the mean transvalve gradient varied between 8 and 12 mmHg, average of 10.7 mmHg. In the double lesion group, 1 patient was reoperated at 43 months. No endocarditis or thromboembolism were reported. CONCLUSION: Mitral stenosis repair has worse late results, related to the valve abnormalities and associated lesions. The correction of mitral insufficiency without annular support showed good long-term results. PMID- 11262572 TI - Oxygen therapy, continuous positive airway pressure, or noninvasive bilevel positive pressure ventilation in the treatment of acute cardiogenic pulmonary edema. AB - OBJECTIVE: To compare the effects of 3 types of noninvasive respiratory support systems in the treatment of acute pulmonary edema: oxygen therapy (O2), continuous positive airway pressure, and bilevel positive pressure ventilation. METHODS: We studied prospectively 26 patients with acute pulmonary edema, who were randomized into 1 of 3 types of respiratory support groups. Age was 69+/-7 years. Ten patients were treated with oxygen, 9 with continuous positive airway pressure, and 7 with noninvasive bilevel positive pressure ventilation. All patients received medicamentous therapy according to the Advanced Cardiac Life Support protocol. Our primary aim was to assess the need for orotracheal intubation. We also assessed the following: heart and respiration rates, blood pressure, PaO2, PaCO2, and pH at beginning, and at 10 and 60 minutes after starting the protocol. RESULTS: At 10 minutes, the patients in the bilevel positive pressure ventilation group had the highest PaO2 and the lowest respiration rates; the patients in the O2 group had the highest PaCO2 and the lowest pH (p<0.05). Four patients in the O2 group, 3 patients in the continuous positive pressure group, and none in the bilevel positive pressure ventilation group were intubated (p<0.05). CONCLUSION: Noninvasive bilevel positive pressure ventilation was effective in the treatment of acute cardiogenic pulmonary edema, accelerated the recovery of vital signs and blood gas data, and avoided intubation. PMID- 11262573 TI - Primary neoplasms of the heart. Clinical and histological presentation of 50 cases. AB - OBJECTIVE: To analyze clinical and histologic findings of 50 patients with primary neoplasms of the heart in a tertiary referral center. METHODS: From 1980 to 1998, we retrospectively analyzed 50 patients, 32 of whom were females, whose ages ranged from 9 to 73 years (mean age = 44.16+/-18 years). RESULTS: Most tumors were located in the left side of the heart (72%), myxoma being the most common (84%) histologic type. The other histologic types found were as follows: fibroma (4%), lipoma (2%), rhabdomyosarcoma (2%), hemangioma (2%), sarcoma (2%), angiosarcoma (2%), and lymphoma (2%). Diagnosis was established by echocardiography in 94% of the cases. Clinical findings were as follows: dyspnea (36%), weight loss (20%), palpitations (18%), chest pain (16%), fever (8%), and arthralgia (6%). All patients with thromboembolic phenomena (10%) had left atrial myxoma. Approximately 20% of the patients were asymptomatic at the initial clinical assessment. CONCLUSION: Primary cardiac tumors are a rare entity with diverse clinical and histologic findings, requiring, therefore, a high level of clinical suspicion. PMID- 11262574 TI - Remission of heart failure through endoluminal repair of femoral arteriovenous fistula with the use of a covered stent. AB - We report the case of a 21-year-old male with high-output heart failure due to a femoral arteriovenous fistula caused by a firearm wound. A new balloon expandable stent covered with polytetrafluorethylene was implanted in the artery to occlude the arteriovenous fistula. The fistula was immediately occluded and the artery remained patent. On the following day, the patient felt much better, with no symptoms of heart failure. Additional follow-up is required to assure the usefulness of this less invasive procedure in the treatment of arteriovenous fistulas. PMID- 11262576 TI - Calcified aneurysms in coronary arteries of a 48-year-old patient. AB - This is a case report of a 48-year-old female patient with a compatible history of Kawasaki disease during childhood, who was admitted to the emergency coronary unit with unstable angina pectoris. Coronary angiography identified two coronary aneurysms, one causing right coronary occlusion and the other causing severe obstruction of the left anterior descending coronary artery. Coronary artery bypass surgery was indicated. PMID- 11262577 TI - [Clinical-radiographic correlation. Case 3/2001 - Instituto do Coracao do Hospital das Clinicas da FMUSP]. PMID- 11262575 TI - Postoperative necrotizing fasciitis of the thorax in cardiac surgery. AB - Necrotizing fasciitis is a rare soft tissue infection and a life-threatening emergency, often fatal. Its incidence and management are described plentifully in the medical literature regarding the most common anatomical sites involved like the abdomen, lower and upper limbs, and perineum. However, available data and case reports of chest wall necrotizing fasciitis after thoracic procedures are scarce, mainly after major cardiac operations. We report and discuss a case of necrotizing fasciitis of the chest wall occurring in the immediate postoperative period of a cardiac procedure, and include a brief review of the concepts, pathophysiology, and treatment reported in the medical literature. We emphasize the need for early diagnosis and urgent and effective surgical debridement. Of importance is the fact that we have not found any references in the literature to cases similar or equal to the one we describe here, which occurred in the postoperative period of a cardiac procedure. PMID- 11262578 TI - The brain decade in debate: III. Neurobiology of emotion. AB - This article is a transcription of an electronic symposium in which active researchers were invited by the Brazilian Society of Neuroscience and Behavior (SBNeC) to discuss the advances of the last decade in the neurobiology of emotion. Four basic questions were debated: 1) What are the most critical issues/questions in the neurobiology of emotion? 2) What do we know for certain about brain processes involved in emotion and what is controversial? 3) What kinds of research are needed to resolve these controversial issues? 4) What is the relationship between learning, memory and emotion? The focus was on the existence of different neural systems for different emotions and the nature of the neural coding for the emotional states. Is emotion the result of the interaction of different brain regions such as the amygdala, the nucleus accumbens, or the periaqueductal gray matter or is it an emergent property of the whole brain neural network? The relationship between unlearned and learned emotions was also discussed. Are the circuits of the former the underpinnings of the latter? It was pointed out that much of what we know about emotions refers to aversively motivated behaviors, like fear and anxiety. Appetitive emotions should attract much interest in the future. The learning and memory relationship with emotions was also discussed in terms of conditioned and unconditioned stimuli, innate and learned fear, contextual cues inducing emotional states, implicit memory and the property of using this term for animal memories. In a general way it could be said that learning modifies the neural circuits through which emotional responses are expressed. PMID- 11262579 TI - RNA and DNA aptamers as potential tools to prevent cell adhesion in disease. AB - Recent research has shown that receptor-ligand interactions between surfaces of communicating cells are necessary prerequisites for cell proliferation, cell differentiation and immune defense. Cell-adhesion events have also been proposed for pathological conditions such as cancer growth, metastasis, and host-cell invasion by parasites such as Trypanosoma cruzi. RNA and DNA aptamers (aptus = Latin, fit) that have been selected from combinatorial nucleic acid libraries are capable of binding to cell-adhesion receptors leading to a halt in cellular processes induced by outside signals as a consequence of blockage of receptor ligand interactions. We outline here a novel approach using RNA aptamers that bind to T. cruzi receptors and interrupt host-cell invasion in analogy to existing procedures of blocking selectin adhesion and function in vitro and in vivo. PMID- 11262581 TI - ClC-5 chloride channel and kidney stones: what is the link? AB - Nephrolithiasis is one of the most common diseases in the Western world. The disease manifests itself with intensive pain, sporadic infections, and, sometimes, renal failure. The symptoms are due to the appearance of urinary stones (calculi) which are formed mainly by calcium salts. These calcium salts precipitate in the renal papillae and/or within the collecting ducts. Inherited forms of nephrolithiasis related to chromosome X (X-linked hypercalciuric nephrolithiasis or XLN) have been recently described. Hypercalciuria, nephrocalcinosis, and male predominance are the major characteristics of these diseases. The gene responsible for the XLN forms of kidney stones was cloned and characterized as a chloride channel called ClC-5. The ClC-5 chloride channel belongs to a superfamily of voltage-gated chloride channels, whose physiological roles are not completely understood. The objective of the present review is to identify recent advances in the molecular pathology of nephrolithiasis, with emphasis on XLN. We also try to establish a link between a chloride channel like ClC-5, hypercalciuria, failure in urine acidification and protein endocytosis, which could explain the symptoms exhibited by XLN patients. PMID- 11262580 TI - Fever induction pathways: evidence from responses to systemic or local cytokine formation. AB - The immune and central nervous systems are functionally connected and interacting. The concept that the immune signaling to the brain which induces fever during infection and inflammation is mediated by circulating cytokines has been traditionally accepted. Administration of bacterial lipopolysaccharide (LPS) induces the appearance of a so-termed "cytokine cascade" in the circulation more or less concomitantly to the developing febrile response. Also, LPS-like fever can be induced by systemic administration of key cytokines (IL-1 beta, TNF-alpha, and others). However, anti-cytokine strategies against IL-1 beta or TNF-alpha along with systemic injections of LPS frequently lead to attenuation of the later stages of the febrile response but not of the initial phase of fever, indicating that cytokines are rather involved in the maintenance than in the early induction of fever. Within the last years experimental evidence has accumulated indicating the existence of neural transport pathways of immune signals to the brain. Because subdiaphragmatic vagotomy prevents or attenuates fever in response to intraperitoneal or intravenous injections of LPS, a role for vagal afferent nerve fibers in fever induction has been proposed. Also other sensory nerves may participate in the manifestation of febrile responses under certain experimental conditions. Thus, injection of a small dose of LPS into an artificial subcutaneous chamber results in fever and formation of cytokines within the inflamed tissue around the site of injection. This febrile response can be blocked in part by injection of a local anesthetic into the subcutaneous chamber, indicating a participation of cutaneous afferent nerve signals in the manifestation of fever in this model. In conclusion, humoral signals and an inflammatory stimulation of afferent sensory nerves can participate in the generation and maintenance of a febrile response. PMID- 11262582 TI - Effect of epithelial debridement on human cornea proteoglycans. AB - Corneal transparency is attributed to the regular spacing and diameter of collagen fibrils, and proteoglycans may play a role in fibrillogenesis and matrix assembly. Corneal scar tissue is opaque and this opacity is explained by decreased ultrastructural order that may be related to proteoglycan composition. Thus, the objectives of the present study were to characterize the proteoglycans synthesized by human corneal explants and to investigate the effect of mechanical epithelial debridement. Human corneas unsuitable for transplants were immersed in F-12 culture medium and maintained under tissue culture conditions. The proteoglycans synthesized in 24 h were labeled metabolically by the addition of (35)S-sulfate to the medium. These compounds were extracted by 4 M GuHCl and identified by a combination of agarose gel electrophoresis, enzymatic degradation with protease and mucopolysaccharidases, and immunoblotting. Decorin was identified as the main dermatan sulfate proteoglycan and keratan sulfate proteoglycans were also prominent components. When the glycosaminoglycan side chains were analyzed, only keratan sulfate and dermatan sulfate were detected (approximately 50% each). Nevertheless, when these compounds were (35)S-labeled metabolically, the label in dermatan sulfate was greater than in keratan sulfate, suggesting a lower synthesis rate for keratan sulfate. (35)S-Heparan sulfate also appeared. The removal of the epithelial layer caused a decrease in heparan sulfate labeling and induced the synthesis of dermatan sulfate by the stroma. The increased deposit of dermatan sulfate proteoglycans in the stroma suggests a functional relationship between epithelium and stroma that could be related to the corneal opacity that may appear after epithelial cell debridement. PMID- 11262583 TI - Effect of a leucine-supplemented diet on body composition changes in pregnant rats bearing Walker 256 tumor. AB - Cancer patients present high mobilization of host protein, with a decrease in lean body mass and body fat depletion occurring in parallel to neoplastic growth. Since leucine is one of the principal amino acids used by skeletal muscle for energy, we investigated the changes in body composition of pregnant tumor-bearing rats after a leucine-supplemented diet. Sixty pregnant Wistar rats divided into six groups were fed a normal protein diet (18%, N) or a leucine-supplemented diet (3% L-leucine, L). The pregnant groups were: control (CN), Walker 256 carcinoma bearing rats (WN), control rats pair-fed with tumor-bearing rats (pfN), leucine supplemented (CL), leucine-supplemented tumor-bearing (WL), and leucine supplemented rats pair-fed with tumor-bearing rats (pfL). At the end of pregnancy, all animals were sacrificed and body weight and tumor and fetal weight were determined. The carcasses were then analyzed for water, fat and total, collagen and non-collagen nitrogen content. Carcass weight was reduced in the WN, WL, pfN and pfL groups compared to control. The lean body mass and total carcass nitrogen were reduced in both tumor-bearing groups. Despite tumor growth and a decrease in fetal weight, there was a slight decrease in collagen (7%) and non collagen nitrogen (8%) in the WL group compared with the WN group which showed a decrease of 8 and 12%, respectively. Although the WL group presented severe tumor growth effects, total carcass nitrogen and non-collagen nitrogen were particularly higher in this leucine-supplemented group compared to the WN group. These data suggest that the leucine-supplemented diet had a beneficial effect, probably attenuating body wasting. PMID- 11262584 TI - Catecholamine-induced vasoconstriction is sensitive to carbonic anhydrase I activation. AB - We studied the relationship between alpha- and beta-adrenergic agonists and the activity of carbonic anhydrase I and II in erythrocyte, clinical and vessel studies. Kinetic studies were performed. Adrenergic agonists increased erythrocyte carbonic anhydrase as follows: adrenaline by 75%, noradrenaline by 68%, isoprenaline by 55%, and orciprenaline by 62%. The kinetic data indicated a non-competitive mechanism of action. In clinical studies carbonic anhydrase I from erythrocytes increased by 87% after noradrenaline administration, by 71% after orciprenaline and by 82% after isoprenaline. The increase in carbonic anhydrase I paralleled the increase in blood pressure. Similar results were obtained in vessel studies on piglet vascular smooth muscle. We believe that adrenergic agonists may have a dual mechanism of action: the first one consists of a catecholamine action on its receptor with the formation of a stimulus receptor complex. The second mechanism proposed completes the first one. By this second component of the mechanism, the same stimulus directly acts on the carbonic anhydrase I isozyme (that might be functionally coupled with adrenergic receptors), so that its activation ensures an adequate pH for stimulus-receptor coupling for signal transduction into the cell, resulting in vasoconstriction. PMID- 11262585 TI - Bone mineral density of the lumbar spine of Brazilian children and adolescents aged 6 to 14 years. AB - The authors performed a study of bone mass in eutrophic Brazilian children and adolescents using dual-energy X-ray absorptiometry (DXA) in order to obtain curves for bone mineral content (BMC) and bone mineral density (BMD) by chronological age and correlate these values with weight and height. Healthy Caucasian children and adolescents, 120 boys and 135 girls, 6 to 14 years of age, residents of Sao Paulo, Brazil, were selected from the Pediatric Department outpatient clinic of Hospital Sao Paulo (Universidade Federal de Sao Paulo). BMC, BMD and the area of the vertebral body of the L2-L4 segment were obtained by DXA. BMC and BMD for the lumbar spine (L2-L4) presented a progressive increase between 6 and 14 years of age in both sexes, with a distribution that fitted an exponential curve. We identified an increase of mineral content in female patients older than 11 years which was maintained until 13 years of age, when a new decrease in the velocity of bone mineralization occurred. Male patients presented a period of accelerated bone mass gain after 11 years of age that was maintained until 14 years of age. At 14 years of age the mean BMD values for boys and girls were 0.984 and 1.017 g/cm2, respectively. A stepwise multiple regression analysis of paired variables showed that the "vertebral area-age" pair was the most significant in the determination of BMD values and the introduction of a third variable (weight or height) did not significantly increase the correlation coefficient. PMID- 11262586 TI - Intestinal permeability in strongyloidiasis. AB - The objective of the present study was to assess intestinal permeability in patients with infection caused by Strongyloides stercoralis. Twenty-six patients (16 women and 10 men), mean age 45.9, with a diagnosis of strongyloidiasis were evaluated. For comparison, 25 healthy volunteers (18 women and 7 men), mean age 44.9, without digestive disorders or intestinal parasites served as normal controls. Intestinal permeability was measured on the basis of urinary radioactivity levels during the 24 h following oral administration of chromium labeled ethylenediaminetetraacetic acid ((51)Cr-EDTA) expressed as percentage of the ingested dose. The urinary excretion of (51)Cr-EDTA was significantly reduced in patients with strongyloidiasis compared to controls (1.60 +/- 0.74 and 3.10 +/ 1.40, respectively, P = 0.0001). Intestinal permeability is diminished in strongyloidiasis. Abnormalities in mucus secretion and intestinal motility and loss of macromolecules could explain the impaired intestinal permeability. PMID- 11262587 TI - Fighting by sleep-deprived rats as a possible manifestation of panic: effects of sodium lactate. AB - Increased fighting is an effect of desynchronized sleep deprivation (DSD) in rats, and recently this behavior has been suggested to be spontaneous panic and equivalent to panic disorder. In the present study we tested this hypothesis by evaluating the effect of sodium lactate on this aggressiveness, because this substance is recognized to induce spontaneous panic attacks in patients. A total of 186 male albino Wistar rats, 250-350 g, 90-120 days of age, were submitted to DSD (multiple platform method) for 0, 4, or 5 days. At the end of the deprivation period the rats were divided into subgroups respectively injected intraperitoneally with 1.86, 2.98 and 3.72 g/kg of 1 M sodium lactate, or 1.86 and 3.72 g/kg of 2 M sodium lactate. The control animals were submitted to the same procedures but received equivalent injections of sodium chloride. Regardless of DSD time, sleep-deprived animals that received sodium lactate presented a significantly higher mean number of fights (0.13 +/- 0.02 fights/min) and a longer mean time spent in confrontation (2.43 +/- 0.66 s/min) than the controls (0.01 +/- 0.006 fights/min and 0.12 +/- 0.07 s/min, respectively; P<0.01, Student t-test). For the sodium lactate group, concentration of the solution and time of deprivation increased the number of fights, with the mean number of fights and mean duration of fighting episodes being greater with the 2.98 g/kg dose using 1 M lactate concentration. These results support the hypothesis that fighting induced by DSD is probably a spontaneous panic manifestation. However, additional investigations are necessary in order to accept this as a promising animal model for studies on panic disorder. PMID- 11262588 TI - Psychometric properties of the Portuguese version of the State-Trait Anxiety Inventory applied to college students: factor analysis and relation to the Beck Depression Inventory. AB - The psychometric properties of the Portuguese version of the trait form of the State-Trait Anxiety Inventory (STAI-T) and its relation to the Beck Depression Inventory (BDI) were evaluated in a large Brazilian college student sample containing 845 women and 235 men. STAI-T scores tended to be higher for women, singles, those who work, and subjects under 30 years. Factor analysis of the STAI T for total sample and by gender yielded two factors: the first representing a mood dimension and the second being related to worrying or cognitive aspects of anxiety. In order to study the relation between anxiety and depression measures, factor analysis of the combination of the 21 BDI items and the 20 STAI-T items was also carried out. The analysis resulted in two factors that were analyzed according to the tripartite model of anxiety and depression. Most of the BDI items (measuring positive affectivity and nonspecific symptoms of depression) were loaded on the first factor and four STAI-T items that measure positive affectivity. The remaining STAI-T items, all of them measuring negative affect, remained in the second factor. Thus, factor 1 represents a depression dimension and factor 2 measures a mood-worrying dimension. The findings of this study suggest that, although widely used as an anxiety scale, the STAI-T in fact measures mainly a general negative affect. PMID- 11262589 TI - Apoptosis of sensory neurons and satellite cells after sciatic nerve transection in C57BL/6J mice. AB - The rate of axonal regeneration, after sciatic nerve lesion in adult C57BL/6J mice, is reduced when compared to other isogenic strains. It was observed that such low regeneration was not due just to a delay, since neuronal death was observed. Two general mechanisms of cell death, apoptosis and necrosis, may be involved. By using the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) technique, we demonstrated that a large number of sensory neurons, as well as satellite cells found in the dorsal root ganglia, were intensely labeled, thus indicating that apoptotic mechanisms were involved in the death process. Although almost no labeled neurons or satellite cells were observed one week after transection, a more than ten-fold increase in TUNEL labeling was detected after two weeks. The results obtained with the C57BL/6J strain were compared with those of the A/J strain, which has a much higher peripheral nerve regeneration potential. In A/J mice, almost no labeling of sensory neurons or satellite cells was observed after one or two weeks, indicating the absence of neuronal loss. Our data confirm previous observations that approximately 40% of C57BL/6J sensory neurons die after sciatic nerve transection, and indicate that apoptotic events are involved. Also, our observations reinforce the hypothesis that the low rate of axonal regeneration occurring in C57BL/6J mice may be the result of a mismatch in the timing of the neurons need for neurotrophic substances, and production of the latter by non neuronal cells in the distal stump. PMID- 11262590 TI - The mechanism of gentisic acid-induced relaxation of the guinea pig isolated trachea: the role of potassium channels and vasoactive intestinal peptide receptors. AB - We examined some of the mechanisms by which the aspirin metabolite and the naturally occurring metabolite gentisic acid induced relaxation of the guinea pig trachea in vitro. In preparations with or without epithelium and contracted by histamine, gentisic acid caused concentration-dependent and reproducible relaxation, with mean EC(50) values of 18 microM and E(max) of 100% (N = 10) or 20 microM and E(max) of 92% (N = 10), respectively. The relaxation caused by gentisic acid was of slow onset in comparison to that caused by norepinephrine, theophylline or vasoactive intestinal peptide (VIP). The relative rank order of potency was: salbutamol 7.9 > VIP 7.0 > gentisic acid 4.7 > theophylline 3.7. Gentisic acid-induced relaxation was markedly reduced (24 +/- 7.0, 43 +/- 3.9 and 78 +/- 5.6%) in preparations with elevated potassium concentration in the medium (20, 40 or 80 mM, respectively). Tetraethylammonium (100 microM), a nonselective blocker of the potassium channels, partially inhibited the relaxation response to gentisic acid, while 4-AP (10 microM), a blocker of the voltage potassium channel, inhibited gentisic acid-induced relaxation by 41 +/- 12%. Glibenclamide (1 or 3 microM), at a concentration which markedly inhibited the relaxation induced by the opener of ATP-sensitive K(+) channels, levcromakalim, had no effect on the relaxation induced by gentisic acid. Charybdotoxin (0.1 or 0.3 microM), a selective blocker of the large-conductance Ca(2+)-activated K(+) channels, caused rightward shifts (6- and 7-fold) of the gentisic acid concentration-relaxation curve. L-N(G)-nitroarginine (100 microM), a NO synthase inhibitor, had no effect on the relaxant effect of gentisic acid, and caused a slight displacement to the right in the relaxant effect of the gentisic acid curve at 300 microM, while methylene blue (10 or 30 microM) or ODQ (1 microM), the inhibitors of soluble guanylate cyclase, all failed to affect gentisic acid induced relaxation. D-(P)-Cl-Phe(6),Leu(17)[VIP] (0.1 microM), a VIP receptor antagonist, significantly inhibited (37 +/- 7%) relaxation induced by gentisic acid, whereas CGRP (8-37) (0.1 microM), a CGRP antagonist, only slightly enhanced the action of gentisic acid. Taken together, these results provide functional evidence for the direct activation of voltage and large-conductance Ca(+2) activated K(+) channels, or indirect modulation of potassium channels induced by VIP receptors and accounts for the predominant relaxation response caused by gentisic acid in the guinea pig trachea. PMID- 11262591 TI - Lack of antidiabetic effect of a Eugenia jambolana leaf decoction on rat streptozotocin diabetes. AB - Streptozotocin-diabetic rats were treated for 17 days with a decoction of Eugenia jambolana (Myrtaceae) leaves (15%, w/v) as a substitute for water. Body weight, food and fluid intake, urine volume, glycemia, urinary glucose and urea were evaluated every 5 days. The animals were sacrificed by decapitation and blood samples collected for the determination of glycemia, serum cholesterol, HDL cholesterol, triglycerides and angiotensin-converting enzyme. The weight of adipose and muscle tissues was also determined. There were no statistically significant differences between treated and untreated rats for any of the biochemical or physiological parameters. We conclude that, at least in this experimental model, Eugenia jambolana leaf decoction has no antidiabetic activity. PMID- 11262592 TI - Acute phenobarbital administration induces hyperalgesia: pharmacological evidence for the involvement of supraspinal GABA-A receptors. AB - The aim of the present study was to determine if phenobarbital affects the nociception threshold. Systemic (1-20 mg/kg) phenobarbital administration dose dependently induced hyperalgesia in the tail-flick, hot-plate and formalin tests in rats and in the abdominal constriction test in mice. Formalin and abdominal constriction tests were the most sensitive procedures for the detection of hyperalgesia in response to phenobarbital compared with the tail-flick and hot plate tests. The hyperalgesia induced by systemic phenobarbital was blocked by previous administration of 1 mg/kg ip picrotoxin or either 1-2 mg/kg sc or 10 ng icv bicuculline. Intracerebroventricular phenobarbital administration (5 microg) induced hyperalgesia in the tail-flick test. In contrast, intrathecal phenobarbital administration (5 microg) induced antinociception and blocked systemic-induced hyperalgesia in this test. We suggest that phenobarbital may mediate hyperalgesia through GABA-A receptors at supraspinal levels and antinociception through the same kind of receptors at spinal levels. PMID- 11262594 TI - Experimental myocardial hypertrophy induced by a minimally invasive ascending aorta coarctation. AB - Ascending aorta coarctation was produced by a minimally invasive technique in rabbits. Animal mortality was 5%. Morphometric and hemodynamic parameters were evaluated. A parabiotically isolated heart model was used to assess the hemodynamic parameters. Left ventricular weight/body weight ratio and muscle area showed clear evidence of hypertrophy when compared to control. The hemodynamic changes in the isolated heart model suggested decreased diastolic and systolic function in the coarcted group. The present model produced hypertrophy with low mortality rates as a result of its less invasive nature. PMID- 11262593 TI - Effect of chronic oral administration of a low dose of captopril on sodium appetite of hypothyroid rats. Influence of aldosterone treatment. AB - Rats rendered hypothyroid by treatment with methimazole develop an exaggerated sodium appetite. We investigated here the capacity of hypothyroid rats (N = 12 for each group) to respond to a low dose of captopril added to the ration, a paradigm which induces an increase in angiotensin II synthesis in cerebral areas that regulate sodium appetite by increasing the availability of circulating angiotensin I. In addition, we determined the influence of aldosterone in hypothyroid rats during the expression of spontaneous sodium appetite and after captopril treatment. Captopril significantly increased (P<0.05) the daily intake of 1.8% NaCl (in ml/100 g body weight) in hypothyroid rats after 36 days of methimazole administration (day 36: 9.2 +/- 0.7 vs day 32: 2.8 +/- 0.6 ml, on the 4th day after captopril treatment). After the discontinuation of captopril treatment, daily 1.8% NaCl intake reached values ranging from 10.0 +/- 0.9 to 13.9 +/- 1.0 ml, 48 to 60 days after treatment with methimazole. Aldosterone treatment significantly reduced (P<0.05) saline intake before (7.3 +/- 1.6 vs day 0, 14.4 +/- 1.3 ml) and after captopril treatment. Our results demonstrate that, although hypothyroid rats develop a deficiency in the production of all components of the renin-angiotensin-aldosterone system, their capacity to synthesize angiotensin II at the cerebral level is preserved. The partial reversal of daily 1.8% NaCl intake during aldosterone treatment suggests that sodium retention reduces both spontaneous and captopril-induced salt appetite. PMID- 11262595 TI - Calcium signalling in secretory cells. AB - Stimulation of secretory cells with muscarinic agonists leads to an increase in the intracellular Ca (2+)concentration ([Ca (2+)]( i)), which activates protein secretion through exocytosis and causes closure of gap junctions between adjacent cells. In addition, the increase in [Ca (2+)](i) activates three different kinds of ion channels: large K(+) channels, Cl(-) channels and non-specific cation channels. The opening of those channels leads to an increase of [Na(+ )] and a decrease of [Cl(-)] and [K(+) ] in the cell. The two components that contribute to the increase in [Ca (2+)]( i) are calcium release from intracellular stores, localised in the endoplasmic reticulum and calcium influx through the plasma membrane. Several models for the regulation of [Ca (2+)](i) have been proposed, including a recently suggested model whereby a distinct pathway involving arachidonic acid is added to the well-established capacitative model. Different hypotheses concerning coupling between the intra-cellular calcium stores and membrane channels co-exist. In addition to a historical overview, recent developments and future challenges are discussed in this review. PMID- 11262596 TI - Force during stretches of rat skeletal muscles after hypertonia at short and long lengths. AB - Following injection of tetanus toxin into rat gastrocnemius muscle to produce hypertonia, plantar flexor muscles were allowed to shorten (S, n=5) without restraint or held lengthened (L, n=3) by splinting. Saline injected rats served as control (n=5). One week after injection, peak forces during 3 stretches with passive muscles and acute isometric force deficits produced by 15 stretches of electrically stimulated muscles were examined under pentobarbital anesthesia. Isometric force and mass of plantar flexors were similar in S rats but 16% lower in L rats compared to control. Peak passive forces were highest in S rats but not different between L rats and control. At the end of the stretch protocol, isometric force deficits were 26% larger in S rats compared to L rats and 17% smaller in L rats compared to control. Acute isometric force deficits produced by stretches of active skeletal muscles were dependent on the muscle length maintained during hypertonia. Our animal model could be used to test rehabilitation interventions during hypertonia of skeletal muscles. PMID- 11262597 TI - Walking at moderate speed with heel-less shoes increases calf blood flow. AB - This study was designed to examine the physiological and biochemical effects of wearing heel-less shoes over a wide range of walking speeds. Six male students wearing alternately regular shoes and heel-less shoes walked at the constant speeds of 60, 80, 100 and 120 m/min for 10 min on a treadmill at 0% grade. The average heart rate was higher during heel-less shoe trials than when subjects walked in regular shoes at each speed, but differences were not significant. The calf blood flow showed its highest mean value at 80 m/min when subjects walked in heel-less shoes, and at 100 m/min when they walked in regular shoes. However, at walking speeds higher than these, calf blood flow decreased for wearers of both types of shoes. The calf blood flow after 80 m/min was higher when walking in heel-less rather than regular shoes. Blood lactate concentration after walking in heel-less shoes at 120 m/min was significantly higher than basal level, but after walking in regular shoes it was unchanged from the level before walking. Noradrenaline concentration at 120 m/min while walking in heel-less shoes was significantly higher than while walking in regular shoes. In conclusion, walking exercise in heel-less shoes induced an increase of the calf blood flow at a moderate speed, and increased glycogen metabolism and noradrenaline secretion at a faster speed. PMID- 11262598 TI - Structural and functional responses of the bullfrog urinary bladder to distension caused by hydrostatic pressure gradients. AB - The responses to mucosal pressure elevation (physiological pressure: PP) were compared to responses to serosal pressure elevation (non-physiological pressure: NPP) in bullfrog urinary bladders (Rana catesbeiana). The bladders were mounted on vertical chambers as flat sheets. Distension was applied with 98.07 Pa. pressure gradients. PP resulted in increases in transepithelial electrical potential difference (TEP) and short-circuit current (SCC). Electrical resistance (R), urea permeability (P(urea)) and net water flux (J( v)) were not effected. NPP resulted in decreases in TEP (38%), SCC (13%), and R (36%). While P(urea) (97%) and J(v) (96%) increased. PP caused little or no change in the electron microscopic structure of frog bladder while NPP caused irreversible dilation of the lateral intercellular spaces. There were no observable changes in tight junctions under PP or NPP. The subepithelial elements of the bladder became detached from the epithelial layer during NPP suggesting a role for them during PP. PMID- 11262599 TI - Effect of oral L-arginine administration for three weeks in two kidney-two clip hypertensive rats. AB - Nitric oxide (NO) has been identified as an effective vascular relaxant. This study analyses the contribution of the precursor L-arginine (L-arg) by oral administration in two kidney-two clip hypertension in the rat (2K-2C). Two groups were studied: sham (SH, n=21) and hypertensive (HT, n=15). After 4 weeks of surgery, a group of rats remained as controls (SHc and HTc, respectively), while others were supplemented with L-arg (1.25 g/L) in drinking water (SHa and HTa) for 3 weeks. Blood pressure was significantly increased in 2K-2C rats but remained unchanged after L-arg treatment. Plasma nitrite/nitrate concentrations were not different among groups. The contractile response of aorta to KCl, serotonin and the protein kinase C (PKC) stimulant, phorbol 12,13-dibutyrate (PDBu) was also evaluated. Higher contractile responses to PDBu (p<0.001) and lower relaxation to acetylcholine (Ach 10(-6) M, p<0.05 and 10(-5)M, p<0.02) were observed in aortic rings of HTc vs SHc; L-arg supplementation significantly diminished tension development to all agonists (p<0.05) but failed to modify the lower relaxation to Ach in HTa. Thromboxane (TxA(2)) - synthesis in rings of HTc was higher than in SHc under basal conditions (p<0.05). In the groups with supplement of L-arg, PDBu significantly stimulated prostacyclin (PGI(2)) synthesis more in HTa rats than in SHa ones (p<0.05). To conclude: 1) L-arg fails to modify hypertension development in 2K-2C rats; and 2) L-arg exerts a beneficial effect on the vascular wall, by reducing contractility in rings from HTa rats; it also improved PGI(2) synthesis under PDBu stimulation. 3) greater PKC activation and TxA(2) production rather than lower NO availability might result in systemic hypertension in 2K-2C rats. PMID- 11262600 TI - Effects of morphine and naloxone on glucose metabolism in uterine strips from ovariectomized and non-ovariectomized restricted diet rats. AB - The effect of underfeeding over glucose metabolism in uteri isolated from ovariectomized and non-ovariectomized rats subjected to a restricted diet for 25 days (50% of the normal food intake), was studied. Underfeeding decreases (14)CO(2) formation from U(14) C-glucose in intact animal uteri. While in ovariectomized rats (25 days), the effect is the opposite. The addition of morphine 10(-6) M to the medium does not affect rats fed ad libitum. However, (14)CO(2) levels increase significantly in intact animals receiving a restricted diet. In ovariectomized rats morphine does not show any activity, regardless of the type of diet rats were subjected to. None of the rat groups seems to be sensitive to naloxone 10(-6) M. The s.c. injection of morphine (4 mg.kg (-1)) increases glucose metabolism only in intact rats provided with a restricted diet, while naloxone (2.5 mg.kg (-1) ) produces a decrease of ( 14)CO(2) in ovariectomized underfed animals. To conclude, morphine either 'in vivo' or 'in vitro' is active only in uteri from intact rats subjected to underfeeding. Naloxone produces a decrease in (14)CO(2) production, particularly when it is s.c. injected to ovariectomized rats undergoing a dietary restriction. Since the uterus does not react to naloxone, the effect of the opiod blocker may be the result of endogenous opioids originated in other tissues. PMID- 11262601 TI - Genistein inhibits osmotic activation of Na(+) /H( +) exchange in human platelets. AB - The osmotic shrinkage is an important activator of the Na(+)/H( *) exchanger. The intracellular signaling mechanisms by which shrinkage changes intracellular pH have not been fully elucidated. In human platelets, the removal of calcium did not prevent the osmotic activation of the exchanger. The increase of pH(i) after an hyperosmotic stress was reduced by W-7 (63 micromol l(-1)), and by ML-7 (25 micromol l(-1)), inhibitors of responses mediated by calmodulin or by myosin light chain kinase, but the high concentrations needed suggested that non specific effects could be involved. Although the exchanger was quiescent during preincubation in hypertonic sodium free solutions, some steps of the signal transduction chain that links the shrinkage to the exchanger activation suffers a modification. Therefore, upon exposure to isotonic sodium-containing media, the rate of recovery from acid loads was increased. The presence of genistein (100 micromol l( -1)) during the preincubation inhibited this activation of Na(+)/H( +) exchanger. We propose that shrinkage induce activation of tyrosine kinases, which in turn leads to the activation of Na(+)/H(+) exchanger and contributes to the restoration of cell volume in human platelets. PMID- 11262602 TI - Regulation of phosphofructokinase-1 on submandibular salivary glands of rats after isoproterenol administration. AB - The purpose of this investigation was to study the effect of isoproterenol (IPR) treatment on the regulation of phosphofructokinase-1 of submandibular salivary glands of rats. The animals were divided into control and experimental groups. In the first set of experiments, the rats received 5 mg of IPR/kg b.w. and were sacrificed at 24 hours after 1, 2, 3 and 4 doses. The content of fructose-2,6 bisphosphate (Fru-2,6-P(2)) and the activity of 6-phosphofructo-2-kinase (PFK-2) (active and total) were determined. The Fru-2,6-P(2) content was found to be reduced and the activity of PFK-2 (active and total) showed differences from the control. The active/total ratio, was higher for the group of one dose sacrificed 12 hours after the agonist injection as compared to the control. In the other groups, there were reductions which varied from 25 to 33%. In the second set of the experiment, the animals were injected with 23.0 mg of IPR/kg b.w. and were sacrificed from 5 up to 720 minutes after the administration of the agonist. After the sacrifice, salivary gland samples were analyzed for Fru-2,6-P(2). Again, a reduction in the metabolite content was observed. Using beta and alpha receptor blockers, it was found that both inhibited only partially the effect of IPR. The purification of PFK-1 up to homogeneity, from submandibular glands of rats which received 5 mg of IPR/mg b.w. as well as from the control, was performed and the Km and state of phosphorylation were determined. Rats from the group sacrificed 12 hours after the injection of the agonist showed the lowest Km for Fru-6-P. Animals which received 3 doses of IPR showed the highest phosphate content/mol of enzyme. Experiments of dephosphorylation of the purified PFK-1 from this latter group revealed that the presence of the phosphate groups influence the kinetic properties of the enzyme. PMID- 11262603 TI - Influences of graded dose of melatonin on the levels of blood glucose and adrenal catecholamines in male roseringed parakeets (Psittacula krameri ) under different photoperiods. AB - Effects of daily evening (just before the onset of darkness in a 24 h light dark cycle) administration of graded doses (25, 50, or 100 microg/100 g body wt./day for 30 days) of melatonin on the concentrations of blood glucose and adrenal catecholamines were studied in sexually active male roseringed parakeets under natural (NP; approximately 12L: 12D) and artificial long (LP; 16L: 8D) and short (SP; 8L: 16D) photoperiods. Blood samples and adrenal glands were collected from each bird during the mid-day on the following day of the last treatment. The concentrations of glucose in blood and epinephrine (E) and norepinephrine (NE) in the adrenals were measured. The results of the study indicated that exogenous melatonin induces hypo- or hyperglycemia depending on the dose of hormone administered as well as to the length of photoperiod to which birds were exposed. The levels of E and NE in the adrenals were shown also to vary in relation to photoperiod and the dose of melatonin administered. But the nature of the influence of melatonin becomes different under altered photoperiodic conditions. It appears that short photoperiods are more effective than long photoperiods as a modulator of glycemic and adrenal catecholaminergic responses to exogenous melatonin. A statistically significant correlation between the levels of blood glucose and that of E and NE in the adrenals was found in the control birds, but not in the melatonin treated birds. The results suggested that the responses of blood glucose and adrenal catecholamines to the treatment with melatonin in the roseringed parakeets may not be dependent on each other. PMID- 11262604 TI - Characterization of the insulin-signaling pathway in lacrimal and salivary glands of rats. AB - PURPOSE: Insulin has been acknowledged as a mediator of several physiological events in lacrimal and salivary glands. We investigated the presence of insulin receptors and of insulin-induced autophosphorylation of the insulin receptor and activation of elements involved in the early steps of insulin signaling in lacrimal and salivary glands of rats. METHODS: Lacrimal and salivary glands of Wistar rats were removed and processed for immunohistochemistry using anti insulin receptor and anti-IGF-1 receptor antibodies. The activation of insulin receptors following insulin treatment, and the involvement of insulin receptor substrates-1 and -2, Shc, JAK-2 and STAT-1, were analyzed by immunoprecipitation, followed by SDS-PAGE and immunoblotting of rat lacrimal and salivary glands after exposure to insulin. RESULTS: Insulin and IGF-1 receptors were present in rat lacrimal and salivary glands and were located predominantly in the cytoplasm and plasma membrane. Functional studies demonstrated that insulin induced a dose dependent phosphorylation of the insulin receptor, IGF-1R, insulin receptor substrates-1 and -2, Shc, and STAT-1. In rats with streptozotocin-induced diabetes mellitus there was a significant reduction in insulin-induced insulin receptor and STAT-1 phosphorylation in the lacrimal gland but not in the salivary gland; there was no influence on Shc phosphorylation in either tissue. CONCLUSIONS: The present results indicate that insulin and IGF-1 receptors are expressed in lacrimal and salivary glands, and that insulin can induce the phosphorylation of its receptor and activate elements involved in the early steps of insulin signaling in both tissues. PMID- 11262605 TI - Immunolocalization of Na-K-ATPase, Na-K-Cl and Na-glucose cotransporters in the conjunctival epithelium. AB - PURPOSE: To investigate possible regional expression of transport systems in the conjunctival epithelium given distinct differences in morphological appearance between the bulbar and palpebral epithelia as well as variations found in the proportions of Na absorptive versus Cl secretory activities in electrophysiological studies. METHODS: Mouse monoclonal antibodies against the alpha1-subunit of Na-K-ATPase and Na-K-Cl cotransporter (NKCC1) and a rabbit polyclonal antibody against the Na-glucose cotransporter (SGLT1) were used in immunoblotting and immunofluorescent labeling of frozen fixed sections isolated from either the bulbar and palpebral regions of the conjunctiva. RESULTS: Western blot analysis clearly demonstrated the presence of Na-K-ATPase, NKCC1 and SGLT1 proteins in both bulbar and palpebral conjunctiva. Indirect immunofluorescence studies on bulbar and palpebral conjunctival portions revealed intense staining by the anti-NKCC1 and anti-alpha1-Na-K-ATPase antibodies exclusively at the basolateral surfaces, whereas the anti-SGLT1 antibody was used with porcine conjunctiva to elicit strong and unambiguous staining along the apical plasma membrane. CONCLUSIONS: Proteins that mediate the transport activities of the Na-K ATPase and Na-K-Cl cotransporter are uniformly distributed throughout the palpebral and bulbar regions of the rabbit conjunctival epithelium. Although the Na-glucose cotransporter could be detected in immunoblots of the rabbit, this cotransporter appears to be uniformly distributed as well, based upon immunohistochemical sections of the pig conjunctiva. Thus, it appears likely that mechanisms for Cl secretion and Na absorption reside in both bulbar and palpebral segments of the conjunctival epithelium. PMID- 11262606 TI - Effect of taurolidine on the normal eyelid and conjunctival flora. AB - PURPOSE: The effectiveness of the topical taurolidine was evaluated in eradicating or reducing microorganisms in the normal flora of human eyes in a randomized controlled study and analyzed also the irritating effects of taurolidine on the ocular surface. METHODS: One hundred and twenty eyes of 110 patients awaiting cataract surgery were randomly divided into four groups consisting of 30 eyes each. The first group received 0.05% taurolidine, the second received 0.3% gentamicin, the third received vehicle eyedrops and the fourth received saline to the preoperative eye four times daily for two days. Cultures were obtained from the eyelids and conjunctivas of all subjects prior to the therapy and again at the end of 48 hours. Micro-biological identification and colony counts were performed by standard laboratory methods, and the results were compared. The patients were clinically evaluated for symptoms and signs at the end of therapies. RESULTS: Taurolidine and gentamicin produced a significant decrease from the basal bacteriological state: the number of colonies (p < 0.01 for taurolidine, p < 0.01 for gentamicin) was reduced by both agents. Staphylococcus epidermidis was the most common microorganism isolated before therapy, and the number of its colonies was significantly reduced in taurolidine treated (p < 0.001) and gentamicin-treated (p < 0.01) subjects. There was no significant difference in terms of the irritating effects for all therapies tested (p > 0.05). CONCLUSIONS: Taurolidine solution with its unique properties is an effective antimicrobial agent for reducing the number of bacteria in the flora of the eye. Taurolidine appears to be well tolerated and offers promise as a potential new antimicrobial drug. PMID- 11262607 TI - Release of complement regulatory proteins from ocular surface cells in infections. AB - PURPOSE: The decay accelerating factor (DAF or CD55) and the membrane inhibitor of reactive lysis (MIRL or CD59), two complement regulatory proteins that protect self cells from autologous complement-mediated injury, are attached to corneal and cqonjunctival epithelial cells by glycosylphos-phatidylinositol (GPI) anchors. We sought to 1) determine the frequency with which bacteria recovered from patients with infections of the eye elaborate factors that can remove these surface proteins from ocular cells, 2) determine the spectrum of bacteria from other sites that have similar effects, and 3) establish the time interval required for reconstitution of the two regulators. METHODS: Culture supernatants of 18 ocular isolates [P. aeruginosa (n = 3), S. marcescens (n = 1), S. epidermidis (n = 9), and S. aureus (n = 5)], and > 100 other clinical specimens isolated in the hospital's microbiology laboratory [P. mirabilis (n = 1), S. aureus (n = 65), S. epidermidis (n = 24), B. cereus (n = 12), H. influenzae (n = 15), and Enterobacter sp. (n = 21)] were incubated at 37 degrees C for various times with conjunctival epithelial cells, conjunctival fibroblasts or HeLa cells and the release of DAF and CD59 proteins from the surfaces of the cells analyzed by 2-site immunoradiometric assays and by Western blotting. The kinetics of recovery of DAF and CD59 expression on the cells was measured by flow cytometry. RESULTS: DAF and/or CD59 release from the cell monolayers varied from < 5% to > 99% at as much as a 1:81 dilution of the supernatant from some bacteria. On conjunctival epithelial cells, more than 8 hr was required for 44% recovery of DAF expression and for 50% recovery of CD59 expression. CONCLUSIONS: Bacteria produce phospholipases and/or other enzymes which can efficiently remove DAF and CD59 from ocular cell surfaces. This phenomenon may correlate with their in vivo pathogenicity. PMID- 11262608 TI - Altered gene expression in lymphocytes of patients with normal-tension glaucoma. AB - BACKGROUND: In glaucoma there is a loss of retinal ganglion cells. There is evidence that this loss can occur by apoptosis. The signal transduction leading to retinal ganglion cell apoptosis in glaucoma is not yet clear. The present study compares the gene expression in lymphocytes of normal tension glaucoma patients (NTG-patients) with the one of healthy controls. METHODS: Subtractive hybridization was used to compare mRNA in lymphocytes of six vasospastic NTG patients with six age and sex matched healthy subjects. RESULTS: Genes coding for p53-protein, NTP (neural thread protein) and 20 S proteasome subunit XAPC7 were overexpressed, whereas those coding for XPGC (Xeroderma pigmentosum gene), the survivin protein as well as one type of ABC transport protein were underexpressed. CONCLUSION: In comparison to healthy controls, patients with vasospastic NTG seem to over- as well as under-express certain genes in their lymphocytes. PMID- 11262609 TI - Expression of involucrin by ocular surface epithelia of patients with benign and malignant disorders. AB - PURPOSE: Keratinization of the ocular surface epithelium is associated with various disorders impairing vision. We immunohistochemically determined whether the ocular surface epithelia express involucrin, and whether its expression pattern may differ in benign vs. malignant disorders. Expression of cytokeratins was also examined to provide further information relative to the epithelial differentiation. METHODS: We evaluated 17 specimens; 6 specimens of the normal ocular surface epithelia, 3 specimens from cases of conjunctival intraepithelial neoplasia (CIN), 6 of conjunctival squamous cell carcinoma (SCC) and 2 of conjunctivae from cases of superior limbic keratoconjunctivitis (SLK). RESULTS: Corneal epithelium exhibited intracellular immunoreactivity for involucrin. Four of the 6 specimens of bulbar conjunctival epithelium showed involucrin immunoreactivity in the perimembranous region, whereas the fornical conjunctiva was negative. Cornified envelope in SLK specimens was positive for involucrin. The CIN showed its immunoreactivity in the perimembranous region in all levels of the hyperproliferative epithelium without keratinization, i.e., similar to the bulbar conjunctiva. The neoplastic cells of well-differentiated SCC showed involucrin in the perimembranous region, and those of moderately- to poorly differentiated SCC have involucrin in their cytoplasm. The expression pattern of cytokeratins was unrelated to grade of malignancy in ocular SCC. CONCLUSION: The epithelia of normal subjects and of CIN expresses involucrin without keratinization. In contrary, the keratinized SLK epithelium markedly expresses involucrin in the cornified envelope. The subcellular immunolocalization of involucrin in the ocular SCC may help in evaluating the differentiation, i.e., malignancy, of neoplastic cells. PMID- 11262610 TI - Normal expression levels of cathepsins, protease inhibitors, and Sp1 in conjunctival tissues from patients with keratoconus. AB - PURPOSE: In keratoconus corneas, it has been shown that levels of degradative enzymes and transcription factor Sp1 are elevated and those of inhibitors are reduced, especially in the epithelial layer. This study is to determine whether the biochemical abnormalities identified in corneas also exist in conjunctival tissues. METHODS: Conjunctival tissues were collected from normal subjects and from patients with keratoconus, senile cataract, and other corneal diseases. Immunohistochemical staining for cathepsins B and G, alpha 1-proteinase inhibitor, alpha 2-macro-globulin and Sp1 was performed. RESULTS: The epithelium of all conjunctival specimens showed immunoreactivity toward the antibodies. The staining for cathepsins B and G, and the inhibitors was mostly cytoplasmic, while that for Sp1 was nuclear. The staining intensity in keratoconus specimens was all within normal range. CONCLUSIONS: These results suggest that the abnormalities in cathepsins B and G, protease inhibitors and Sp1 identified in keratoconus corneas are not manifested in the conjunctiva. Keratoconus appears to be a disease localized to the cornea. PMID- 11262611 TI - Immunohistology of kerato-epithelin in corneal stromal dystrophies associated with R124 mutations of the BIGH3 gene. AB - PURPOSE: To investigate corneal deposits associated with kerato-epithelin (KE) in three corneal dystrophies harboring mutations at Arg-124 in the BIGH3 gene. METHODS: Six patients with Avellino corneal dystrophy (ACD) associated with R124H, one patient with superficial granular corneal dystrophy (SGCD) associated with R124L, and seven patients with lattice corneal dystrophy type 1 (CDL1) associated with R124C were examined. Corneal buttons obtained during keratoplasties were stained with Masson's trichrome and with Congo red, and immunostained with antibodies specific for N-terminal and C-terminal portions of KE (KE-15 and KE-2, respectively). RESULTS: In all corneas with ACD, subepithelial to midstromal deposits of granular material stained with KE-2 and KE-15. However, deep stromal deposits containing amyloid reacted with KE-2, but not KE-15. Granular deposits in the subepithelial layer observed in SGCD stained intensely with KE-2 and KE-15. In all corneas with CDL1, subepithelial and midstromal amyloid deposits stained with KE-2; these deposits did not stain with KE-15. Deposits between the epithelial layer and Bowman's layer stained with Masson's trichrome but not with Congo red in five of the seven corneas; these deposits were stained with both KE-2 and KE-15. CONCLUSIONS: Deposits in corneal buttons involved by ACD, SGCD, and CDL1 included forms of the BIGH3 gene product, KE. An N-terminal sequence of KE may be related to formation of amyloid associated with R124 mutations. PMID- 11262613 TI - Anti-rat ICAM-1 antibody does not influence the course of experimental melanin induced uveitis. AB - PURPOSE: Experimental melanin-induced uveitis (EMIU) is a T cell-mediated rat model of acute anterior uveitis. We investigated the possibility of preventing the inflammation with monoclonal antibody directed against rat intercellular adhesion molecule 1 (ICAM-1). METHODS: To induce EMIU, Lewis rats were immunized with bovine ocular melanin extract (250-500 microg). Each day from day 6 post immunization, rats were injected intraperitoneally with anti-ICAM-1 monoclonal antibody (IA29) or normal mouse serum, and examined with a slit-lamp biomicroscope. On the first day of clinical inflammation, intravital microscopy of iris vasculature was performed on each animal following intraperitoneal injection of rhodamine 6G. At the peak of clinical inflammation, rats were killed, and eyes were examined histologically. Binding potency of IA29 was tested by flow cytometry using concanavalin A-stimulated rat T cells. Immunohistochemical staining was used to detect IA29 on rat uveal vascular endothelium. RESULTS: The ability of IA29 to bind T cell blasts was present to a 1:2000 dilution, and IA29 was readily detectable on uveal vascular endothelium following systemic administration. However, there was no significant difference (p > 0.05) in incidence, time of onset, or severity or histological appearance of EMIU for the rats treated with IA29 when compared with the control rats. Intravital microscopy revealed sticking of leukocytes in the iris vasculature in both groups. CONCLUSIONS: We were unable to demonstrate an inhibitory effect of anti-rat ICAM-1 antibody on the outcome of EMIU. Our observations may reflect a redundancy in the adhesion molecule profile responsible for this uveitis. PMID- 11262612 TI - Association of ocular pressure and optic disc cup volume with red blood cell sodium-potassium ATPase inhibition. AB - PURPOSE: To determine if there were significant differences between the number of red blood cell ouabain binding sites in normals and untreated ocular hypertensives plus one open-angle glaucoma patient. METHODS: We measured the binding of (3)H ouabain to erythrocyte membranes of 23 normals, 25 ocular hypertensives and one open-angle glaucoma. We also measured the levels of plasma cortisol and digoxin in these subjects. Characteristics of cupping of the optic disc and thickness of the retinal nerve fiber layer, as well as area of optic disc pallor of these subjects were measured by stereophotogrammetry and by computerized image analysis from single and stereo photographs. RESULTS: The number of (3)H ouabain binding sites was observed to be significantly less in the ocular hypertensives and one glaucoma compared to the normals (p = 0.0009). In multi-variate analyses, to determine what other factors affected this difference, there was a significant negative association with mean intraocular pressure (p = 0.003) (average of both eyes) and total cup volume (average of both eyes) (p = 0.005), diagnosis of ocular hypertension and glaucoma (p = 0.0005) and male gender (p = 0.019). There was a significant positive association with plasma cortisol levels (p = 0.048), and diastolic blood pressure (p = 0.037). CONCLUSIONS: The number of (3)H ouabain binding sites in red blood cells decreases significantly with increasing ocular pressure and increasing cup volume indicating the possible presence of an increased systemic endogenous digoxin-like inhibitor and/or difference in the isozymes of Na(+), K(+)-ATPase which may be associated with increased levels of plasma cortisol in ocular hypertensives and glaucomas. PMID- 11262614 TI - Vasoactive intestinal peptide (VIP) exacerbates endotoxin-induced uveitis (EIU) in mice. AB - PURPOSE: We have previously shown that inhibition of the proinflammatory cytokine tumor necrosis factor- (TNF)-alpha exacerbates the inflammatory process of EIU. To further examine this paradoxic phenomenon, we investigated here the effect on EIU of VIP, a neuropeptide that inbibits TNF-alpha production. METHODS: VIP was injected concurrently with endotoxin at doses that induce EIU or lethality in mice. Severity of EIU was measured by counting infiltrating cells in eye sections, at 1 or 5 days post endotoxin injection. Survival of mice was monitored periodically, while serum levels of TNF-alpha, interleukin-(IL)-1beta and IL-10 were determined by caputure ELISA. RESULTS: Treatment with VIP exacerbated EIU but provided partial protection from the lethal endotoxin effect. VIP treatment also reduced serum levels of TNF-alpha and IL-1beta, but increased levels of IL 10. CONCLUSION: This study further established the paradoxical observation that EIU is exacerbated by lowering the levels of circulating pro-inflammatory cytokines, in particular TNF-alpha. PMID- 11262615 TI - Isolation and characterization of a Ca(2+)-activated chloride channel from human corneal epithelium. AB - PURPOSE: Transparency of the cornea is maintained through the activity of secretory mechanisms in the epithelium and endothelium, which offset the tendency of the stroma to imbibe fluid and swell. These secretory mechanisms establish osmotic gradients thereby providing the osmotic driving forces for coupled fluid transport from the stroma into both the tears and the anterior chamber. To further characterize the mechanism of epithelial Cl secretion, we cloned a cDNA encoding a Ca(2+)-dependent chloride channel, an abundant mRNA in human corneal epithelium. We investigated the abundance of all known human chloride channels in corneal epithelium to identify those responsible for regulating chloride conductance in this tissue. METHODS: For the isolation of a full-length cDNA clone, a probe was selected from a set of expressed sequenced tag (EST) clones classified as unique to corneal epithelium (http://bodymap. ims.u-tokyo.ac.jp). The expression patterns of the corresponding gene encoding novel chloride channel gene in human cornea and other tissues were examined by reverse transcription polymerase chain reaction (RT-PCR). Quantitative PCR was performed to clarify the expression level of the novel chloride channel gene in cornea relative to that in other human tissues. RESULTS: We cloned a new Ca(2+)-activated chloride channel, CLCA2, from corneal epithelium. The full length cDNA clone encoded 943 amino acids with 62% identity to bovine Ca(2+)-activated chloride channel. The CLCA2 gene mapped to human chromosome 1p32. Quantitative expression analysis by RT-PCR showed that it is the most abundant chloride channel in corneal epithelium. CONCLUSION: High and tissue specific expression of the CLCA2 gene in human corneal epithelium implies an important role in corneal transparency maintenance. PMID- 11262616 TI - Comparison of various calpain inhibitors in reduction of light scattering, protein precipitation and nuclear cataract in vitro. AB - PURPOSE: To compare effects of calpain inhibitors on in vitro light-scattering in rat lens soluble protein and calcium-ionophore (A23187)-induced cataract formation in cultured rat lenses. METHODS: Rat lens soluble protein was hydrolyzed for 24 hours by activation of endogenous lens calpain. Ten calpain inhibitors were tested in this model at 10 and 25 microM concentration. As an index of protein precipitation, light scattering was measured daily at 405 nm for 8 days. Lens proteins were analyzed by isoelectric-focussing. Subsequently, rat lenses were cultured for 5 days with 10 microM A23187. Calpain inhibitors (SJA6017, MDL28170, AK295 and PD150606), which inhibited light-scattering were tested at 100 microM concentration in this model. Cataract evaluation, isoelectric-focussing and calcium determinations were performed. RESULTS: At 25 microM concentration AK295, SJA6017, E-64, PD-150606 and MDL28170 produced greater than 25% inhibition of light-scattering. Isoelectric-focussing revealed that addition of Ca(2+) produced characteristic crystallin proteolysis and aggregation patterns. AK295, SJA6017, MDL28170 and E64c prevented these changes. Lenses cultured in A23187 exhibited nuclear cataract, elevated calcium and proteolysis and aggregation of crystallins. Co-culture with SJA6017, MDL28170 and E64c reduced A23187-induced nuclear opacities, proteolysis and aggregation of crystallins without affecting increased total calcium. CONCLUSIONS: Endogenous calpain-activation model and A23187-induced cataract model can be used sequentially to screen calpain inhibitors for potential anti-cataract activity. Proteolytic changes in lens cortex after exposure to A23187 are also due to calpain activation. AK295, SJA6017 and MDL28170 possess efficacy against calcium induced models of rodent cataracts. Use of calpain inhibitors represents a promising approach to cataract therapy. PMID- 11262617 TI - Immunohistochemical localization of MYOC/TIGR protein in the trabecular tissue of normal and glaucomatous eyes. AB - PURPOSE: To examine immunohistochemically the localization of myocilin/trabecular meshwork inducible glucocorticoid response (MYOC/TIGR) protein in the glaucomatous and normal trabecular meshworks. METHODS: Trabecular tissues were used from one eye with late-onset goniodysgenetic glaucoma, three with primary open angle glaucoma (one of which had the MYOC/TIGR gene mutation), two with exfoliation glaucoma and one without glaucoma. For light microscopic immunohistochemistry, frozen sections were stained by the avidin-biotin complex method using anti-MYOC/TIGR polyclonal antibody. For electron microscopic immunohistochemistry, the pre-embedding method using the same antibody was performed. Double immunostaining using both anti-MYOC/TIGR and anti-type VI collagen antibodies was done by the immunofluorescence method. RESULTS: With light microscopy, immunoreactivity was seen in the whole trabecular meshwork of each of the specimens. No notable differences were detected in staining among the types of glaucoma, or between the eyes with and those without the gene mutation. Under electron microscopy, immunoreaction products were observed not only in the cytoplasm of the trabecular cells but also in the extracellular matrix, where staining was associated with the long-spacing collagen, fine granular materials and possibly microfibrils. With double immunohistochemistry, MYOC/TIGR was colocalized with type VI collagen in the trabecular meshwork. CONCLUSIONS: In glaucomatous and normal trabecular meshworks, the MYOC/TIGR protein is distributed in the extracellular matrix colocalizing with type VI collagen. PMID- 11262618 TI - Effect of shear stress on the hydraulic conductivity of cultured bovine retinal microvascular endothelial cell monolayers. AB - The shear stress of flowing blood on endothelial cells increases water transport (hydraulic conductivity, Lp) in several vascular beds in vivo and has been hypothesized to play a role in elevating vascular transport in ocular diseases such as diabetic retinopathy. The purpose of this study is to determine the response of Lp to varying levels of shear stress using an in vitro model of the blood-retinal barrier: bovine retinal endothelial cells (BRECs) grown on polycarbonate filters. The study also addresses the role of nitric oxide (NO) and other downstream effectors in mediating shear-induced changes in water transport. A step change in shear stress of 10 dyn/cm(2) did not produce a significant change in Lp over 3 hours, whereas a 20 dyn/cm(2) step change elevated Lp by 14.6 fold relative to stationary controls at the end of 3h of shear exposure. 20 dyn/cm( 2) of shear stress stimulated the endothelial monolayers to release nitric oxide in a biphasic manner and incubation of the BRECs with a nitric oxide synthase (NOS) inhibitor, L-NMMA, significantly attenuated the shear-induced Lp response. These experiments demonstrate that NO is a key signaling molecule in the pathway linking shear stress and Lp in BRECs. A widely studied pathway downstream of NO involves the activation of guanylate cyclase (GC), guanosine 3', 5' -- cyclic monophosphate (cGMP) and protein kinase G (PKG). It was observed that incubation of BRECs with the GC inhibitor, LY83583 (10 microM) or the PKG inhibitor, KT5823 (1 microM) did not significantly alter the shear-induced Lp response. Also the cGMP analogue, 8-br-cGMP (1mM), did not affect the baseline Lp over 4h. These results demonstrate that shear stress elevates hydraulic conductivity in BRECs through a signaling mechanism that involves NO but not the GC/cGMP/PKG pathway. PMID- 11262619 TI - Active drug targeting with immunoconjugates to choroidal neovascularization. AB - PURPOSE: Active drug targeting with monoclonal antibody to neovascular vessels may be a potential treatment for choroidal neovascularization (CNV) in age related macular degeneration (AMD). Endoglin (CD105) is a proliferating endothelial cell marker with excellent potential for targeting. The goals of this study were to investigate the expression of CD105 in CNV membranes surgically excised from patients with AMD and CNV lesions induced by intense laser photocoagulation in a cynomolgus monkey and to evaluate the in vitro effect of immunoconjugates on endothelial cells. METHODS: CNV membranes were surgically excised from 10 patients with AMD. Experimental CNV was induced by intense laser photocoagulation in a cynomolgus monkey. Immunolocalization of CD105 on frozen sections of CNV lesions was studied by immunohistochemical evaluation. Anti-von Willebrand's factor antibody was used as an endothelial cell marker. The cytotoxic effect of immunoconjugates of anti-CD105 monoclonal antibody and dextran binding mitomycin C on human umbilical vein endothelial cells (HUVECs) was evaluated in vitro. RESULTS: Endothelial cells demonstrated strong immunoreactivity of CD105 in all surgically excised CNV membranes. In the monkey eye, CD105-positive cells were detected only in CNV lesions but not in normal chorioretinal tissues. Immunoconjugates with anti-CD105 monoclonal antibody showed a specific inhibitory effect on proliferating HU-VECs. CONCLUSIONS: These results suggest that anti-CD105 monoclonal antibody-mediated drug targeting has a potential to treat CNV in AMD. PMID- 11262620 TI - Matrix metalloproteinases and tissue inhibitors of metalloproteinases of fibrous humans lens capsules with intraocular lenses. AB - PURPOSE: We located immunohistochemically the matrix metalloproteinases (MMP) -1, -2, -3 and -9 and the tissue inhibitors of matrix metalloproteinases (TIMP) -1 and -2 in the fibrous capsule of patients with intraocular lenses (IOLs). METHODS: During vitreoretinal surgery in 10 patients we obtained post-cataract surgery lens capsules with or without an IOL. The mean interval between the previous cataract operation and the extraction of the specimens was 35.2 months (range: 2-120 months). Circular sections of the anterior capsule with lens epithelial cells (LECs) were also obtained during cataract surgery. Specimens were processed for immunohistochemical identification of MMPs and TIMPs by light microscopy. RESULTS: While all the members of MMPs and TIMPs were not detected in the normal anterior capsules, they were detected in the ECM and/or LECs on the lens capsules extracted within 18 months after IOL implantations in all of the 4 patients, but were not observed in specimens obtained 18 months or longer postoperatively. In LECs of 1 capsule specimen 10 years postoperatively, MMP-1, but not other MMPs and TIMPs, was detected. CONCLUSIONS: MMPs and TIMPs were detected in the ECM and/or LECs on post-cataract surgery capsules. These proteins may be remodeling the newly deposited ECM and regulating LEC behavior on residual lens capsules in the early phase of healing after cataract surgery. PMID- 11262621 TI - Modulation of early response gene expression by prostaglandins in cultured rat retinal pigment epithelium cells. AB - PURPOSE: To explore the role of prostaglandins (PGs) as modulators of retinal pigment epithelium (RPE) rod outer segment (ROS)-phagocytosis and ROS phagocytosis-induced gene expression. METHODS: RPE cells in primary cell culture were pre-incubated with PGE( 2), PGD(2), PGF(2)alpha, PGJ(2), 15-deoxy-Delta( 12,14)-PGJ(2) or U-46619 (stable analog of thromboxane A(2)), and fed with a suspension of ROS. Expression of zif-268 and tis-1 mRNA was determined by Northern blotting. DNA-binding activity of TIS-1 protein was assessed by electrophoretic mobility shift assay. Concentration of PGE (2) and PGD (2) in the tissue culture medium was measured by enzyme immuno-assay. Phagocytis-tosis was quantified by counting of double-immunostained bound and ingested ROS. RESULTS: PGE 2, the most potent of PGs, strongly elevated both basal and ROS-phagocytosis induced levels of tis-1 mRNA, while significantly inhibiting both basal and phagocytosis-induced expression of zif-268 mRNA. PGD(2), PGJ(2) and 15-deoxy Delta(12,14)-PGJ( 2) elevated ROS-phagocytosis-induced, but not basal, expression of tis-1 mRNA expression. PGF(2alpha) super-induced both phagocytosis-induced and basal tis-1 mRNA expression. U-46619 and carbaprostacyclin had no effect on expression of tis-1 mRNA. PGE(2) was the only PG to affect zif-268 expression. Exogenous PGE(2), PGD( 2) and PGF(2alpha), when added to the medium at 1-microM concentrations, significantly inhibited ingestion of ROS, with PGE(2) being the most potent PG affecting ROS-phagocytosis. CONCLUSIONS: PGs act as selective regulators of phagocytosis-induced transcription factor gene expression in RPE cells, as well as of ROS-phagocytosis itself. This modulation may help to ensure specificity in the differential activation of target genes by ROS-phagocytosis receptor-mediated signal transduction in RPE cells. PMID- 11262622 TI - Photosensitized light-induced damage of IRBP (interphotoreceptor retinoid-binding protein): effects on binding properties. AB - PURPOSE: To determine if IRBP (interphotoreceptor retinoid-binding protein) is damaged following irradiation by visible light in the presence of bound all-trans retinal. METHODS: Following irradiation of the IRBP-all-trans retinal complex, the retinal was removed and damage to IRBP measured as loss of titratable thiol groups, loss of tryptophan fluorescence, and changes in retinol-binding-induced fluorescence. RESULTS: IRBP irradiated by itself showed only minimal loss of tryptophan fluorescence; this loss was substantially increased by irradiation in the presence of all-trans retinal. Thiol groups and retinol-binding activity were also shown to be reduced. The damage to IRBP seemed to involve photosensitization by the all-trans retinal, which was in turn protected from bleaching by the IRBP. The binding affinity was shown to be reduced ten-fold following irradiation. CONCLUSION: In the eye, IRBP can stabilise vitamin A and debatably may be responsible for transport of different forms of vitamin A between the photoreceptor cells and pigment epithelium. If this is the case, it would play a key role in rhodopsin regeneration after bleaching. IRBP also appears to be necessary to sustain photoreceptor cells. Light was shown to cause photosensitized damage to IRBP, and thus might impair the regeneration process and photoreceptor viability. PMID- 11262623 TI - Flow cytometry for quantification of retrogradely labeled retinal ganglion cells by Fluoro-Gold. AB - PURPOSE: To count retrogradely labeled retinal ganglion cells by Fluoro-Gold. METHODS: Retinal ganglion cells were retrogradely labeled using bilateral injections of Fluoro-Gold into the superior colliculus. One week after injections, retinas were dissociated and immunolabeled using specific antibody against Fluoro-Gold. The Fluoro-Gold labeled cells were then counted using flow cytometry. RESULTS: Flow cytometry revealed that approximately 7% of the dissociated retinal cells were ganglion cells retrogradely labeled by Fluoro-Gold (Fig. 1B). Based on the total count of retinal cells per eye, the total number of retinal ganglion cells was estimated at approximately 131,250 +/- 2,542 per rat eye. The coefficient of variation of counts was calculated as 1.98%. CONCLUSIONS: The use of flow cytometry facilitates simple, reproducible and rapid quantification of virtually all of the retinal ganglion cells labeled by Fluoro Gold in a single rat eye. PMID- 11262629 TI - Field homology: a meaningful definition. AB - Field homology refers to populations of cells that derive from evolutionarily conserved regions of embryos but are distributed across sets of adult morphological structures that cannot be placed in one-to-one correspondance. The concept of field homology has proven especially attractive to comparative neurologists because it allows them to deal with the fact that sets of nuclei or nuclear subdivisions often cannot be compared on a one-to-one basis across phyletic groups. However, the concept of field homology has recently come under criticism. It has been argued that field homology is theoretically impossible because it requires sequences of developmental stages to be both evolutionarily conserved and evolutionarily modified. It has also been argued that field homology allows overly vague comparisons of adult morphological structures, fails to account for homologous structures that derive from non-homologous embryonic sources, and establishes overly rigid links between embryonic and adult morphology. All of these criticisms may be adequately addressed by explaining field homology in terms of differentiation. The present paper explains field homology in terms of differentiation using the amniote dorsal thalamus to illustrate major points. It is concluded that field homology is a meaningful concept when defined in terms of differentiation, applied to appropriate cases, and properly limited in its comparisons of adult structures. PMID- 11262628 TI - Effect of gravity on the interaction between the avian germ and neighbouring ooplasm in inverted egg yolk balls. AB - The developmental capacities of an avian germ (from before symmetrization to the moment of laying) are strongly diminished after inversion of its egg yolk ball followed by culture in egg white. Our present experiments show that even when the avian germ is completely horizontally inverted (without an upper or lower border) below its egg yolk ball before symmetrization, symmetrization and gastrulation phenomena take place. The germ grows slower and becomes smaller than after normal incubation. After culture of inverted unincubated germs, localized on freshly laid eggs, the closure of the neural tube is impaired and it remains open over a long distance. Although a primitive streak (PS) develops, mesoderm migration (mainly from the lateral part of the area pellucida) is also impaired. On sections through the germinal disc one can see the abnormal upward migration into the depth of the ooplasm and yolk of cells from the germ wall and the development of large cellular extensions encircling the yolk globules. Most prominent is the loss of contact between the superficial cell layers and the deep layer elements (junctional endoblast and yolk endoblast in the area opaca). Large areas without deep layer elements (even visible on surface micrographs) develop in the area vasculosa and area vitellina interna. The margin of overgrowth grows and extends normally over the egg yolk ball. An autoradiographic study after labelling of the yolk layers in inverted egg yolks reveals that mainly compression of the peripheral subgerminal and perigerminal ooplasm takes place. This suggests that the compression by the neighbouring yolk and upwards growth of cells are at the origin of the impaired development. After return to the normal upward orientation of the germ on the topmost part of the egg yolk ball, a more or less pronounced restoration to normal development takes place (depending on the duration of the inversion period and the age of the germ). PMID- 11262630 TI - Electronmicroscopic observations on the visceral and parietal rat's pleura after contralateral pneumonectomy. AB - The mechanism of compensatory growth and healing of the pleura remains unresolved. Contralateral visceral and parietal (diaphragmatic and costal) pleura were investigated by transmission electron microscopy, following an experimental pneumonectomy (EP). Fifteen young-adult Wistar rats were divided into three groups and with survival times of 1, 5 and 8 days respectively after EP. Three sham-operated (thoracic cavity opened and closed) and three unoperated rats served as controls. One day following EP the superficial mesothelial cells have more microvilli and microvesicles, but a lower number of specialized contacts. Multiplication of extravasal cells leads to an increase of the thickness of the layer over the basal lamina and of the submesothelial layer. Five days after EP the superficial cells show a stratified arrangement in longer sectors of both pleural sheets. Along with typical mesothelial cells there are three new populations of cells: (1) with an abundant granular endoplasmic reticulum and secretory granules, (2) with fibroblast-like characteristics and (3) with a more extensive lysosomal system. The submesothelial layer is thickened due to newly formed blood vessels and collagen bundles. Eight days after EP the mesothelial cells build multi-row arrangement sectors and surround intercellular dilatations covered with microvilli. 'Activated' high mesothelial cells characterize the monolayer sectors. The submesothelial layer remains thicker due to larger collagen bundles and elastic fibers. The changes in the mesothelium and in the connective tissue layer suggest the existence of two periods. The first one is characterized by different mesothelial cell populations, new vasculogenesis and starting of fibrillogenesis. In the second period there are 'activated' mesothelial cells, pleural villi, groups of lymphatic lacunae and significant fibrillogenesis. PMID- 11262631 TI - Immunohistochemical study of the distribution of endocrine cells in the gastrointestinal tract of the camel (Camelus bactrianus). AB - The regional distribution and relative frequency of endocrine cells in the gastrointestinal tract of the camel, Camelus bactrianus, were investigated using immunohistochemical methods. Ten types of immunoreactive (IR) endocrine cells were identified in this study. Among these cell types, only serotonin- and somatostatin-IR cells were detected in almost all regions of the gastrointestinal tract. Most of the cell types showed peak density in the pyloric gland region. The others showed restricted distribution: gastrin, cholecystokinin (CCK), motilin, bovine pancreatic polypeptide (BPP), and (gastric) substance P in the stomach; gastrin, CCK, BPP, gastric inhibitory polypeptide (GIP), glucagon, peptide tyrosine tyrosine (PYY) and substance P in the small intestine; and CCK, motilin, BPP, and PYY in the large intestine. Fundamentally the distribution pattern of endocrine cells in the gastrointestinal tract of the camel is similar to that of cattle. The distribution and frequency of endocrine cells in the glandular sac region are the same as those of the cardiac gland. PMID- 11262632 TI - Judging the future: Whose fault will it be? AB - This paper looks at the future from the perspective of the way in which present thinking can influence what the future might be. It assumes that history shapes the future and that the present generation is in a position to shape it. It looks at the future of medicine as a science and a professional discipline, of health care as policy and politics, of culture and ideology as forces shaping medicine and health care, and of biomedical ethics as an influential source of wisdom and perspective. The paper argues that a strong future for bioethics requires a broad rather than a reductionistic vision of its proper work. PMID- 11262633 TI - Doctor does not know best: why in the new century physicians must stop trying to benefit patients. AB - While twentieth-century medical ethics has focused on the duty of physicians to benefit their patients, the next century will see that duty challenged in three ways. First, we will increasingly recognize that it is unrealistic to expect physicians to be able to determine what will benefit their patients. Either they limit their attention to medical well-being when total well-being is the proper end of the patient or they strive for total well-being, which takes them beyond their expertise. Even within the medical sphere, they have no basis for choosing among the proper medical goals for medicine. Also, there are many plausible strategies for relating predicted benefits to harms, and physicians cannot be expert in picking among these strategies. Second, increasingly plausible ethical systems recognize that in some cases, patient benefit must be sacrificed to protect patient rights including the right to the truth, to have promises kept, to have autonomy respected, and to not be killed. Third, ethics of the next century will increasingly recognize that some patient benefits must be sacrificed to fulfill duties to others - either the duty to serve the interests of others or other duties such as keeping promises, telling the truth, and, particularly, promoting justice. Physicians in the twenty-first century will be seen as having a new, more limited duty to assist the patient in pursuing the patient's understanding of the patient's interest within the constraints of deontological ethical principles and externally imposed duties to promote justice. The result will be a duty to be loyal to the consumer of health care with the recognition that often this will mean that the physician is not permitted to pursue the physician's understanding of the patient's well-being. PMID- 11262634 TI - The twilight of "medicine" and the dawn of "health care": reflections on bioethics at the turn of the millennium. AB - The traditional paradigm of medicine assumes that health is a natural given depending on a body's intrinsic teleology, and that medicine aims at restoring or preserving health, making a physician only an "assistant to nature." I argue that nowadays this paradigm is becoming obsolete, because the concept of health is no longer a "natural given" and interventions on the human body attempt not only to help nature's teleology, but also to change it whenever doing so can satisfy human needs and wants. We should abandon the term "medicine" and adopt the term "health care" to mark such an epoch-making transition, analogous to that marking the passage from "alchemy" to "chemistry." PMID- 11262635 TI - Children as research subjects: a dilemma. AB - A complex problem exists about how to promote the best interests of children as a group through research while protecting the rights and welfare of individual research subjects. The Nuremberg Code forbids studies without consent, eliminating most children as subjects, and the Declaration of Helsinki disallows non-therapeutic research on non-consenting subjects. Both codes are unreasonably restrictive. Another approach is represented by the Council for the International Organizations of Medical Science, the U.S. Federal Research Guidelines, and many other national policies. They allow research ethics committees or institutional review boards to authorize studies with acceptable balances of likely benefits and harms, but neither clarify how to balance them nor explain the meaning of pivotal concepts, like "minimal risk." Paths to the improvement of balancing or consequentialist approaches include (1) improving standardizing of risk assessment, (2) rejecting crude utilitarianism, (3) identifying and justifying normative or moral judgments, and (4) acknowledging extra-regulatory thresholds and deontological or non-negotiable duties to children. PMID- 11262638 TI - Finely crafted distinctions and the art of clinical ethics. AB - Making finely crafted distinctions and deploying them in intellectually rigorous and clinically applicable judgments define, to a considerable degree, the art of clinical ethics. The papers in this "Clinical Ethics" number of the Journal of Medicine and Philosophy demonstrate the art of clinical ethics in their consideration of respect for autonomy vs. respect for persons, the role of risk in triggering assessment of decisional capacity vs. the role of risk in the concept and assessment of decisional capacity, intention vs. foresight in the clinical management of ectopic pregnancy, preserving life vs. relieving suffering in physician-assisted suicide, and what is essential vs. non-essential in defining the "core areas" of ethics education for members of hospital ethics committees. PMID- 11262639 TI - Persuasion as respect for persons: an alternative view of autonomy and of the limits of discourse. AB - The article calls for a departure from the common concept of autonomy in two significant ways: it argues for the supremacy of semantic understanding over procedure, and claims that clinicians are morally obliged to make a strong effort to persuade patients to accept medical advice. We interpret the value of autonomy as derived from the right persons have to respect, as agents who can argue, persuade and be persuaded in matters of utmost personal significance such as decisions about medical care. Hence, autonomy should and could be respected only after such an attempt has been made. Understanding suffering to a significant degree is a prerequisite to sincere efforts of persuasion. It is claimed that a modified and pragmatic form of discourse is the necessary framework for understanding suffering and for compassionately interacting with the frail. PMID- 11262640 TI - On risk and decisional capacity. AB - Limits to paternalism are, in the liberal democracies, partially defined by the concepts of decision-making capacity/incapacity (mental competence/incompetence). The paper is a response to Ian Wilks's (1997) recent attempt to defend the idea that the standards for decisional capacity ought to vary with the degree of risk incurred by certain choices. Wilks's defense is based on a direct appeal to the logical features of examples and analogies, thus attempting to by-pass earlier criticisms (e.g., Culver & Gert, 1990) of risk-based standards. Wilks's argument is found wanting on the grounds that he misconstrues the logic of such capacity, especially in accounting for conceptual and pragmatic ties with issues of decisional authority. A diagnosis is offered as to the source of Wilks's error (the assumption that mental competence is a species of wider genus of "competence"), and an alternative way of accounting for risk within the predominant contemporary legal framework is sketched. PMID- 11262641 TI - Moral absolutism and ectopic pregnancy. AB - If one accepts a version of absolutism that excludes the intentional killing of any innocent human person from conception to natural death, ectopic pregnancy poses vexing difficulties. Given that the embryonic life almost certainly will die anyway, how can one retain one's moral principle and yet adequately respond to a situation that gravely threatens the life of the mother and her future fertility? The four options of treatment most often discussed in the literature are non-intervention, salpingectomy (removal of tube with embryo), salpingostomy (removal of embryo alone), and use of methotrexate (MXT). In this essay, I review these four options and introduce a fifth (the milking technique). In order to assess these options in terms of the absolutism mentioned, it will also be necessary to discuss various accounts of the intention/foresight distinction. I conclude that salpingectomy, salpingostomy, and the milking technique are compatible with absolutist presuppositions, but not the use of methotrexate. PMID- 11262642 TI - Do physicians have an inviolable duty not to kill? AB - An important part of the debate over physician-assisted suicide concerns moral duties that are specific to physicians. It is sometimes argued that physicians, by virtue of special commitments rooted in the nature of their profession, may never intentionally kill a patient, and that therefore, whether or not assisted suicide may be justifiable, it can never be right for a physician to take part in such an act. I examine four types of argument that have been offered in support of this conclusion, and find that none succeeds. Each attempts to show why the duty to conserve life must be unconditional for physicians, yet a consideration of the ways in which contemporary medicine has evolved shows that such a duty is now no more fundamental to the profession than a duty to relieve suffering, which may in some cases override it. PMID- 11262643 TI - The breadth of bioethics: core areas of bioethics education for hospital ethics committees. AB - The multidisciplinary nature of bioethics can result in narrow "sub-specialists" within the field, whose work reflects the issues and concerns most relevant to their "home discipline." This can result in work which is insensitive to the important ways in which particular areas of bioethics are interrelated, and which (while viable in the context of the sub-specialty) is not viable in a broader context. The narrow focus of many healthcare ethics committees on issues directly related to clinical patient care can exacerbate this problem. Increasingly, issues in the clinical care of patients cannot be separated from issues in research, organizational ethics, and public policy. I argue that these problems call for a need to identify "core" areas for bioethics education. This is especially true for education of hospital ethics committees, which increasingly face complex cases involving concerns that fall outside traditional patient care issues. I then consider nine areas examined in detail in A Companion to Bioethics edited by Helga Kuhse and Peter Singer, as potential candidates for "core" areas of bioethics education. At the same time, I evaluate the range of issues examined in each area of the book, in the context of the book's ability to provide an introduction to each area. PMID- 11262645 TI - Further delineation of the facial 13q14 deletion syndrome in 13 retinoblastoma patients. AB - Thirteen years ago, Motegi and colleagues (J Med Genet 1987;24:696-697) summarized the specific facial phenotype of six Japanese retinoblastoma patients with interstitial 13q14 deletions. Among a series of 228 propositi with retinoblastoma referred to the Lausanne Retinoblastoma Clinic for treatment and genetic counseling between 1986 and 1997, 13 (5.7%) were diagnosed with a cytogenetic de-novo 13q14 deletion. We confirm the presence of the reported facial phenotype in our population of Caucasian patients and describe additional clinical traits, thus extending the facial phenotype associated with the 13q14 deletion. Del(13q14) comprises, among others, cranial anomalies, frontal bossing, deeply grooved and long philtrum, depressed and broad nasal bridge, bulbous tip of the nose, thick lower lip, thin upper lip, broad cheeks, and large ears and lobules. Recognition of this particular facial appearance was instrumental in the genetic diagnosis of 13q deletions and in the presymptomatic diagnosis of retinoblastoma in a significant number of our cases. Identification of this phenotype in a retinoblastoma patient allows for efficient diagnosis of recurrence in his progeny and/or sibship, while its ignorance will compromise genetic counseling due to the possible difficulties in detecting large deletions by standard molecular mutation analysis. Recognition of this syndrome in newborns without known familial risk for retinoblastoma is even more important as it is a clear warning sign that indicates immediate ophthalmic examination. PMID- 11262644 TI - Molecular characterization of the deletion in retinoblastoma patients with 13q14 cytogenetic anomalies. AB - We investigated the molecular deletions of twelve patients presenting with retinoblastoma and a cytogenetic abnormality including band 13q14. Dinucleotide markers spanning the complete chromosome 13 as well as two intragenic markers were analyzed in patients and their two parents. The deletion was considered confirmed when one heterozygous allele was missing, potential when a homozygous allele was observed in continuity with a clearly deleted allele, and noninformative when a homozygous allele was observed adjacent to a nondeleted region. The patients could be classified into three groups based on their cytogenetic abnormalities. In group 1, the cytogenetic deletion was restricted to band13q14 with confirmed or potential molecular deletions extending from D13S328 to D13S153. Although a possible common centromeric deletion breakpoint could exist for three of the patients and a common telomeric deletion breakpoint for two, the cytogenetic deletion was different for most of them. Group 2 included patients with a cytogenetic deletion extending up to 13q22. At the molecular level, the telomeric breakpoints were between the RB1 gene and D13S156. Here again, it is quite unlikely that a common telomeric breakpoint was responsible for the deletion. Group 3 consisted of special cases with either a paracentric inversion or a complex translocation. The cytogenetic abnormalities around 13q14 correlate with the molecular deletions that were observed in this study. Associated malformations cannot be easily predicted from the size of the deletions. PMID- 11262646 TI - Evaluation of RLBP1 in 50 autosomal recessive retinitis pigmentosa and 4 retinitis punctata albescens Spanish families. AB - Defects in retinal vitamin A metabolism or in genes expressed in the retinal pigment epithelium (RPE) are related to nonsyndromic retinitis pigmentosa (RP). The RLBP1 gene encodes the cellular retinaldehyde-binding protein which, in the RPE and Muller cells of the retina, is thought to play a role in retinoid metabolism and visual pigment regeneration. We describe a study of the involvement of the RLBP1 gene in 50 autosomal recessive retinitis pigmentosa (ARRP) and four retinitis punctata albescens Spanish families. Cosegregation and homozygosity studies using an intragenic polymorphism and three close markers (D15S116, D15S127, and D15S130) ruled out RLBP1 as the cause of ARRP in 26 pedigrees. In the remaining families, SSCP analysis of the coding region and sequencing of the abnormal migrating bands did not detect any disease-causing mutation. These results indicate that mutations in the RLBP1 gene are not responsible for the ARRP or retinitis punctata albescens in this set of Spanish families. We did, however, identify two frequent polymorphisms (3'UTR + 167 G > T, T: 0.23 and G: 0.77; IVS6 + 20 T > C, T: 0.36 and C: 0.64), a silent substitution (S218S), and a rare variant (5'UTR-101 G > A). PMID- 11262647 TI - EFEMP1 is not associated with sporadic early onset drusen. AB - The early onset of multiple drusen in the posterior pole of the retina is characteristic of a group of macular dystrophies often referred to as dominant or radial drusen. At least two forms, Doyne honeycomb retinal dystrophy (DHRD) and Malattia Leventinese (MLVT), are associated with a single missense mutation (R345W) in the gene encoding the EGF-containing fibulin-like extracellular matrix protein-1 (EFEMP1) and are now thought to represent a single entity. Here, we present a further evaluation of the role of EFEMP1 in the pathogenesis of sporadic forms of early onset drusen. We analyzed all coding exons of the EFEMP1 gene by SSCP analysis in 14 unrelated individuals with early onset of multiple drusen and no apparent family history of the disease. In this patient group, we did not detect the R345W mutation or any other disease-associated mutation. Three different polymorphisms and two intragenic polymorphic repeats were present in similar frequencies in the patients and control individuals. We conclude that EFEMP1 is unlikely to be involved in the disease in this patient group. This suggests that mutations in a different as yet unknown gene or genes may lead to the early onset drusen phenotype. PMID- 11262648 TI - Molecular and clinical characterization of a patient with a chromosome 4p deletion, Wolf-Hirschhorn syndrome, and congenital glaucoma. AB - Wolf-Hirschhorn syndrome is a developmental disorder associated with hemizygous deletion of the distal short arm of chromosome 4. We have identified a patient affected with Wolf-Hirschhorn syndrome and early onset glaucoma. Five other patients with Wolf-Hirschhorn syndrome and early onset glaucoma or ocular anomalies associated with early onset glaucoma have been previously described, suggesting that the association with Wolf-Hirschhorn syndrome is not coincidental. The infrequent association of early onset glaucoma suggests that the chromosomal region commonly deleted in Wolf-Hirschhorn patients does not contain genes responsible for early onset glaucoma. In this study, we performed a molecular characterization of the deleted chromosome 4 to determine the extent of the deletion in an attempt to begin to identify the chromosomal region responsible for the associated glaucoma. Using microsatellite repeat markers located on 4p, we determined that the deletion spanned a 60-cM region including the minimal Wolf-Hirschhorn region. The proximal breakpoint occurred between markers D4S3045 and D4S2974. These results support the hypothesis that patients with Wolf-Hirschhorn syndrome and early onset glaucoma may have large deletions of 4p that include a gene(s) that may be responsible for a dominant form of congenital glaucoma. PMID- 11262649 TI - Phenotype associated with an R120X nonsense mutation in the RP2 gene in a Japanese family with X-linked retinitis pigmentosa. AB - We examined a Japanese family with X-linked retinitis pigmentosa (RP) associated with a nonsense mutation, R120X, in the RP2 gene. The 26-year-old proband presented at the age of seven years with a two-year history of night blindness. Visual disability worsened with increasing age. At age 24, visual acuity was 0.08 in both eyes. Testing for refractive error indicated mild myopia. Visual fields showed bilateral-constriction to 10 degrees. He had central macular areolar sclerosis in both eyes. Two maternal uncles had vision of light perception to hand movement in their early forties together with dense bilateral cataracts. The ocular phenotype of this family with R120X was considered severe; reported phenotypes associated with this mutation have not been uniform. PMID- 11262650 TI - Laboratory methods in ophthalmic genetics: obtaining DNA from patients. AB - DNA samples are the fundamental research substrate in genetics. Although methodology and cost-effectiveness in molecular biology have improved dramatically, collecting biological samples and extracting DNA continue to be expensive and time-consuming steps of genetic research. This article reviews the issues surrounding the choice of biological samples for methods of DNA extraction as well as the storage and transport of biological and DNA samples for genetic studies. PMID- 11262651 TI - Cytokine gene expression in different strains of mice with endotoxin-induced uveitis (EIU). AB - PURPOSE: The kinetics of various cytokines in the eye plays a critical role in endotoxin-induced uveitis (EIU). This study examined the cytokine kinetics and susceptibility of EIU in four mice strains. METHODS: Four strains of TLR-4 or Toll-like receptor-4 (Lps, lipopolysaccharide-susceptible) gene-positive mice (C3H/HeN of H-2(k), C57/B6 of H-2(b), Balb/C of H-2(d), and 129/J of H-2(b)) were injected subcutaneously with either lipopolysaccharide (LPS) in phosphate buffered saline (PBS) or PBS alone in two repeated experiments. Mice were sacrificed 1, 3, 6, 24 (1 d), 72 (3 d), 120 (5 d), or 168 (7 d) hours after LPS injection. Ocular histology and reverse transcriptase-polymerase chain reaction (RT-PCR) to detect ocular interleukin-1 alpha (IL-1 alpha), IL-6, tumor necrosis factor-alpha (TNF-alpha), and granulocyte-macrophage colony-stimulating factor (GM-CSF) mRNA were performed. Serum IL-1 alpha, IL-1 beta, IL-6, and TNF-alpha levels were measured using enzyme-linked immunosorbent assay (ELISA). RESULTS: No ocular inflammation was present in any mice within six hours after LPS injection. Only the C3H/HeN mice developed a biphasic ocular inflammatory response (1 d and 5 d), during which all proinflammatory cytokine messages were expressed. In the other three strains with minimal (129/J and Balb/C) to mild (C57/B6) EIU that peaked at 1 d, IL-6 mRNA was barely detectable in C57/B6 and Balb/C; GM-CSF mRNA was also present in C57/B6. Serum IL-1 alpha, IL-1 beta, IL-6, and TNF-alpha were high in all EIU mice within six hours after LPS injection. Control mice did not develop uveitis or measurable cytokine messages. CONCLUSION: In the most susceptible strain, C3H/HeN, EIU was biphasic and correlated to multiple proinflammatory cytokines released in the eye. The less susceptible mice strains exhibited a monophasic response to LPS that may result from no cytokine cascade. PMID- 11262652 TI - Utility of existing Vogt-Koyanagi-Harada syndrome diagnostic criteria at initial evaluation of the individual patient: a retrospective analysis. AB - The diagnosis of Vogt-Koyanagi-Harada syndrome is hampered by its variable manifestations and the lack of unique ancillary and laboratory findings. In this study, the diagnostic criteria established in 1978 by the American Uveitis Society (AUS) are retrospectively analyzed via their application to a population of 71 consecutive patients, using only those features present at the initial evaluation. All patients were previously diagnosed with Vogt-Koyanagi-Harada syndrome based on the clinical features and course of the disease combined with fluorescein angiography with or without ultrasonography in selected cases. Mean age of all patients was 36.7 years +/- 15.1 years. Fifty (70%) were female and 45 (65%) were Hispanic. Patients presenting acutely, subacutely, and in the chronic stages met the AUS criteria for Vogt-Koyanagi-Harada syndrome in 56%, 48%, and 58% of cases, respectively. Allowance for variation in features, incomplete cases, and modification of the disease by treatment might increase the sensitivity of the criteria. While diagnostic criteria in general are useful for establishing the likelihood that a patient has a disease, the current AUS criteria for the diagnosis of Vogt-Koyanagi-Harada syndrome do not seem adequate. PMID- 11262653 TI - Clinical features of human T-lymphotropic virus type 1 uveitis: a long-term follow-up. AB - To investigate the clinical manifestations of human T-lymphotropic virus type-1 uveitis (HU), 112 HU patients who were followed up periodically for more than one year were retrospectively analyzed with respect to their ophthalmological and systemic complications. The gender ratio (female/male ratio) of the HU patients was 2.0 and the initial complications were foggy vision in 34.5%, ocular floaters in 33.3%, and blurred vision in 15.5%. As for the ocular symptoms, the majority (78.6%) of patients were classified as intermediate uveitis with vitreous inflammation. Recurrence of uveitis episodes was seen in one half of the patients (51.8%); 12 patients had more than six uveitis episodes. The interval of uveitis episodes varied from two weeks to 10 years. Nearly one half of the patients (43.8%) had ocular complications: e.g., cataract in 22 patients, persistent vitreous opacities in 17 patients, and glaucoma in 16 patients. Although the visual prognosis was essentially good, 11 patients had poor visual prognosis (<0.1). The causes of poor vision in these patients were cataract, cystoid macular edema, epiretinal membrane, and optic nerve atrophy. Of the 112 HU patients, two developed HTLV-I-associated myelopathy (TSP/HAM) after the onset of HU, while none developed adult T-cell leukemia. Sixteen HU patients had a previous history of Graves' disease and a past history of methimazole therapy, while Graves' disease was found in another HU patient only after HU onset and methimazole was not administered before the onset of HU. The present data of long term follow-up indicate that (1) HU causes various ocular complications and its visual prognosis can be poor, (2) TSP/HAM can be induced even after the onset of HU, and (3) methimazole is not a risk factor of HU after Graves' disease. PMID- 11262654 TI - Ocular and central nervous system lymphoma: clinical features and diagnosis. AB - PURPOSE: To evaluate the clinical, angiographic, and cytopathologic features of ocular and central nervous system (CNS) lymphoma. PATIENTS AND METHODS: Retrospective study of 44 patients over a 10-year period. RESULTS: A total of 36 women and six men, mean age 54 years (range: 36-90 years), were included. The mean time interval between onset of ocular symptoms and diagnosis was 40 months (range: 1-144 months). Ocular involvement was bilateral in 84% of the cases. Laser flare photometry readings averaged 9.6 photons/ms (2.9-78.3 photons/ms). Vitritis was constant. Funduscopy revealed RPE abnormalities in 60.49% of the cases and punctuate retinal infiltrates in 33.5%. The most common findings with fluorescein angiography were window defects and hypofluorescent round lesions. Patients had CNS involvement in 66% of the cases. Cytologic examination of the vitreous samples showed high-grade B lymphoma in 86% of the cases. Interleukin-10 dosage, when performed, showed elevated levels averaging 2352 pg/ml in all vitreous samples. Molecular biology based on PCR confirmed the diagnosis in 12 patients. Treatment included systemic chemotherapy alone or associated with radiotherapy in various regimens. Fourteen patients died during follow-up. Only 12 patients were in complete remission. CONCLUSION: The prognosis of the disease remains poor. However, the new diagnostic tools and therapeutic strategies may improve the diagnostic delay and the survival outcome. PMID- 11262655 TI - Uveitis in children. AB - PURPOSE: To summarize the prevalence and patterns of uveitis in children. METHODS: Pertinent articles were reviewed. RESULTS: Children constitute 5-10% of the patients with uveitis seen at tertiary referral centers, and girls appear to develop uveitis slightly more frequently than boys. Among all children with intraocular inflammation, anterior uveitis accounts for 30-40%, posterior uveitis accounts for 40-50%, intermediate uveitis accounts for 10-20%, and diffuse uveitis accounts for 5-10%. The most common cause of anterior uveitis is juvenile idiopathic arthritis (JIA), whereas the most frequent type of posterior uveitis is toxoplasmic retinochoroiditis. Most cases of intermediate and diffuse uveitis are bilateral, chronic, and idiopathic. The most common causes of vision loss in children with uveitis are cataract, band keratopathy, glaucoma, and cystoid macular edema. Up to one-third of the children with uveitis are left with severely impaired vision as a result of these complications. CONCLUSIONS: Uveitis is an important cause of ocular morbidity in children. Prompt diagnosis and treatment is essential to minimize the risk of long-term vision loss. PMID- 11262656 TI - Ocular manifestations of infection with the human immunodeficiency virus in an African pediatric population. AB - PURPOSE: To describe the ocular manifestations of HIV/AIDS infection in an African pediatric population. METHODS: From 1984 to 1990, all children with HIV infection attending the Department of Pediatrics of the 'Centre Hospitalier de Kigali', Rwanda, were referred to the Department of Ophthalmology for ophthalmic examination. RESULTS: A total of 162 HIV-infected children were examined. The overall rate of ophthalmic involvement was 54%. The most common finding was a perivasculitis of the peripheral retinal vessels, observed in 38% of the patients. Cytomegalovirus (CMV) infection of the retina was diagnosed in three patients. Isolated cotton-wool spots of the retina were not observed. Ophthalmic herpes zoster and conjunctival xerosis responding to vitamin A administration were each seen in two patients. One third of a subset of children tested for lacrimal function had evidence of decreased tear secretion. CONCLUSION: Our data, in agreement with other series reported in the literature, indicate that cotton wool spots and CMV retinitis, the most common ocular manifestations of HIV/AIDS in adults, are much less prevalent in children. The high incidence of perivasculitis in the present series, not observed or only seen in a few cases in other series, suggests that this ocular sign is more prevalent in African children. Our working hypothesis is that perivasculitis of the retinal vessels, lymphoid interstitial pneumonitis, parotitis, and lacrimal gland involvement are the expression of a diffuse infiltrative lymphocytosis syndrome, similar to what has been described in adults. PMID- 11262657 TI - Sensitivity of indocyanine green angiography for the follow-up of active inflammatory choriocapillaropathies. AB - BACKGROUND: Inflammatory choriocapillaropathies (choriocapillaritis) correspond to the clinical spectrum of lesions of the fundus, including acute posterior multifocal placoid pigment epitheliopathy (APMPPE), multiple evanescent white dot syndrome (MEWDS), multifocal choroiditis (MC), and other rarer entities caused by inflammatory disturbances of choriocapillaris perfusion. The aim here was to study the sensitivity of indocyanine green (ICG) angiography in investigating and following inflammatory choriocapillaropathies. PATIENTS AND METHODS: Patients with inflammatory choriocapillaropthies were included who had had a dual fluorescein and ICG angiography as well as visual field testing (Goldman or computerized perimetry) at presentation and on follow-up visits. ICG angiography was performed according to a routine angiographic protocol used for inflammatory diseases and was correlated with fundus examination, fluorescein angiography, and visual field testing. RESULTS: Three patients with MEWDS, two with APMPPE, and two with MC were included. The visual field alterations in all seven patients were well correlated with the extent of the hypofluorescent areas seen on ICG angiography, whereas they were badly correlated with fluorescein angiographic signs and their evolution. The visual field in MEWDS was particularly well correlated with the importance of peripapillary hypofluorescence seen on ICG angiography. In MC, the evolution of new lesions was well demonstrated by ICG angiography and well correlated with visual symptoms and visual fields, but was barely detected on fundus examination and by fluorescein angiography. CONCLUSIONS: ICG angiographic signs were shown to be closely correlated with visual function (visual field testing). This was not the case for either fundus examination or fluorescein angiography. ICG angiography appears as a very sensitive follow-up parameter in inflammatory choriocapillaropathies, giving morphological information on the evolution of the disease and on the response to treatment when therapy is indicated. PMID- 11262658 TI - Vitrectomy in the management of uveitis. AB - OBJECTIVE: To review the indications for vitrectomy in uveitis cases. PATIENTS AND METHODS: Charts of patients seen at the uveitis clinic of the Jules Gonin Eye Hospital from January 1993 to August 1998 and who had undergone vitrectomy were reviewed. Patients with infectious uveitis occurring within three months after intraocular cataract surgery were excluded. The types of uveitis were recorded and indications for vitrectomy were analyzed. RESULTS: A total of 630 patients were examined at the uveitis clinic. Fifty-one of these patients (51 eyes, 8.1%) were referred for vitrectomy and were included in this study. Vitrectomy was performed for three reasons: 1) to treat the complications of uveitis (90%), including vitreous opacification in 35 eyes (69%), retinal detachment in seven eyes (14%), epimacular membrane in seven eyes (14%), and dense opacification of the posterior capsule after cataract surgery in six eyes (12%)(the mean delay between uveitis and vitrectomy in this group was 8.4 years); 2) for diagnostic purposes in 19 eyes (37%); and 3) to remove confined infectious foci in 16 eyes (31%) and allow a thorough intraocular distribution of antibiotics. Visual acuity improved in 41 patients (80.4%), remained unchanged in three (5.8%), and decreased in seven (13.7%) because of secondary or persistent retinal detachment or cystoid macular edema. CONCLUSION: Vitrectomy was indicated to treat the complications of uveitis, to provide vitreous for diagnostic purposes, and to allow a better diffusion of intraocular antibiotics. Long-standing uveitis did not seem to be influenced by vitrectomy. PMID- 11262659 TI - Efficacy of interferon alfa-2a in severe and refractory uveitis associated with Behcet's disease. AB - PURPOSE: To evaluate the efficacy of interferon alfa-2a (IFN alfa) in severe uveitis associated with Behcet's disease, that is refractory to steroids and conventional immunosuppressive agents. PATIENTS AND METHODS: Patients with Behcet's disease (according to the International Study Group criteria), who relapsed despite steroids and immunosuppressive agents, were included in this retrospective study. Ophthalmological examination, laser flare photometry, and fluorescein angiography associated with laboratory tests were performed at regular intervals. IFN alfa (3 millions units thrice a week) was injected subcutaneously. RESULTS: Eight patients (sex ratio: 1) were included between May 1995 and January 1999. The mean age was 29.1 years (14-54 years) and the disease was present between 11 and 167 months before the administration of IFN alfa. IFN alfa was efficient in all cases with a mean follow-up of 22 months (10-55 months). Steroids were tapered from a mean dosage of 47 mg/d to a mean dosage of 8.5 mg/d. Ocular inflammation was controlled and visual acuity improved in all cases. Treatment was generally well tolerated despite a constant but transient flu-like syndrome. IFN alfa was tapered in three patients and stopped in one case without any relapse after five months. CONCLUSIONS: Within the limitations of this retrospective study, low-dose IFN alfa seems to be well tolerated, promising in the management of refractory forms of uveitis due to Behcet's disease, and effective in allowing a reduction of steroid dosage. A prospective controlled study is necessary to confirm these preliminary results. PMID- 11262660 TI - Precise monitoring and differentiation of inflammatory events by indocyanine green (ICG) angiography in a case of recurrent posterior sarcoid uveitis. AB - PURPOSE: By providing information on choroidal lesions, indocyanine green (ICG) angiography is complementary to fluorescein angiography in the workup of posterior uveitis. The aim here was to illustrate practically the suitability of performing dual fluorescein/ICG angiograms in a demonstrative case of recurrent presumed posterior sarcoid uveitis. METHODS: Sequential dual fluorescein/ICG angiograms were performed for the workup and follow-up of a case of recurrent presumed sarcoid uveitis. RESULTS: Dual fluorescein and ICG angiograms performed during the first recrudescence of inflammation showed mainly retinal involvement with cystoid macular edema, diffuse posterior pole retinal hyperfluorescence, and papillitis, whereas the second recurrence after tapering oral corticosteroid therapy showed new choroidal lesions of the posterior pole in the absence of significant retinal involvement. CONCLUSIONS: Dual fluorescein/ICG angiography clearly allowed the differentiation of two different inflammatory events, one involving the retina and the other involving the choroid, at the origin of similar clinical manifestations. PMID- 11262663 TI - The molecular biology of retinoblastoma. AB - Retinoblastoma, a rare pediatric eye tumor, has served as an important model for the heritable predisposition to cancer. The retinoblastoma protein, Rb, functions as a tumor suppressor by controlling progression through the cell cycle. Rb function is regulated primarily by its phosphorylation state, which is determined by the complex interaction of multiple kinases and their inhibitors that together form the 'Rb pathway'. This pathway has been found to be functionally inactivated in almost all types of cancer. Despite recent advances in our understanding of Rb function, the precise role of Rb loss in the development of retinoblastoma remains unclear. Recent work in genetically altered mice has suggested that an additional mutation in another gene is required for retinal tumor formation. An alternative model presented here is based on the noncell-autonomous functions of Rb contributing to tumorigenesis. PMID- 11262664 TI - The role of cytokines in corneal immunopathology. AB - Cytokines play an important role in the pathogenesis of various corneal diseases and during corneal graft rejection. Furthermore, cytokines may also play a role in the maintenance of the integrity of the normal cornea. This review focuses on the effects of several cytokines in corneal immunopathology, including the type of the corneal immune response, angiogenesis, chemotaxis, apoptosis, wound healing, corneal disease, and transplantation. It may provide clues for the future treatment of corneal disease and corneal transplantation rejection. PMID- 11262665 TI - Ocular pANCA antigens are expressed in nonpigmented ciliary body epithelium and are conserved in multiple mammalian species. AB - PURPOSE: pANCA marker autoantibody is expressed by a subset of patients with anterior uveitis. A recombinantly isolated pANCA monoclonal antibody, Fab 5-3, identifies ocular expression of corresponding pANCA antigens in human ciliary body and retina. In this study, Fab 5-3 was used to explore pANCA antigen expression in ocular tissues of multiple mammalian species and identify the ciliary body cell type expressing the pANCA antigen. METHODS: Ocular tissues were obtained from several mammalian species and evaluated for expression of the pANCA (Fab 5-3) antigen(s) using immunohistochemistry and Western analysis of tissue extracts. Additionally, primary cultures of nonpigmented and pigmented rabbit ciliary body epithelium were analyzed for pANCA expression using immunofluorescence and Western analysis. RESULTS: Ocular pANCA (Fab 5-3) antigen expression was observed by immunohistochemistry only in the cytoplasm of retinal ganglion cells and ciliary body epithelium. Retinal antigen expression was conserved in all species examined. Ciliary body expression was observed in human, rabbit, rat, and mouse, but not in pig or cow. Antigen expression in the rabbit ciliary body was restricted to the nonpigmented layer as defined in primary cultures of nonpigmented and pigmented ciliary body epithelium. Immunoreactive proteins in both the human and rabbit included a 32-33 kDa doublet (histone H1), and novel 80 and 100 kDa proteins. CONCLUSIONS: This study identifies ocular pANCA antigen expression in multiple mammalian species localized to the retinal ganglion cell layer and the non-pigmented ciliary body epithelium. The present study also establishes novel 80 and 100 kDa proteins which may correspond to the cytoplasmic antigens detected in situ and can be further characterized biochemically and immunologically using small animal model systems. PMID- 11262666 TI - Use of methotrexate in the management of sight-threatening uveitis. AB - Managing chronic inflammatory immune-mediated eye diseases currently involves the non-specific suppression of the immune system. Oral corticosteroids are still the chief component of this therapy, but additional immunosuppressive agents are often indicated to control the disease as well as to avoid the side effects of long-term therapy with high doses of corticosteroids. Many different drugs are available including cyclosporine, azathioprine and occasionally cyclophosphamide, chlorambucil, or more recently mycophenolate mofetil and tacrolimus (FK-506). Pulsed low-dose oral methotrexate (MTX) is now widely used in rheumatological disease and has been introduced in the management of sight-threatening uveitis. Some reports using low doses advocate its use as an alternative when multiple drug therapy is necessary or to help lower the dose of other drugs. In our group of 11 patients with long-standing disease, adding MTX to the treatment regimen allowed control of the inflammation with a reduction of the corticosteroid dose in more than 50% of the cases and decreased the number of disease relapses in 45%. These results indicate that MTX is useful as an alternative option when a second/third agent is needed, achieving a better control of the disease in some cases. PMID- 11262668 TI - Transient visual symptoms in systemic lupus erythematosus and antiphospholipid syndrome. AB - PURPOSE: To review the potential pathogenic mechanisms of transient visual symptoms (TVS) in the course of systemic lupus erythematosus (SLE) and antiphospholipid syndrome (APS), to discuss the most common clinical features associated with the occurrence of TVS, and to explore possible treatment options for these patients. METHODS: The literature regarding the clinical and laboratory characteristics of SLE and APS patients experiencing TVS is reviewed from 1979 onward. A brief review of the wide spectrum of ophthalmologic features occurring in SLE and APS is also provided. RESULTS: Data emerging from the review process point to thromboembolism as the most probable cause of TVS in SLE and APS. Thromboembolisms are likely induced by cardiac valve abnormalities and should be treated with anticoagulant drugs. CONCLUSION: While progress has been made in understanding the association of TVS with SLE and APS, further investigation is needed to clarify this interesting relationship. PMID- 11262667 TI - Outcome of vitrectomy for retained lens fragments after phacoemulsification. AB - PURPOSE: To evaluate the incidence of complications and the visual outcome of pars plana vitrectomy in patients with retained lens fragments in the vitreous cavity after phacoemulsification. METHODS: A retrospective chart review of 85 patients who underwent vitrectomy for removal of retained lens fragments at the Barnes Retinal Institute/Washington University Medical Center between 1990 and 1998. RESULTS: At the time of presentation, uveitis (n = 57, 67.1%), increased intraocular pressure >25 mmHg (n = 44, 51.8%), and corneal edema (n = 42, 49.4%) were frequently observed. The initial visual acuity was 20/200 or worse in 61 (71.8%) eyes. However, the final visual acuity after vitrectomy, with 10.1 months follow-up, was 20/40 or better in 44 (51.8%) eyes. The major complication observed was retinal detachment, which was present in seven (8.2%) eyes: four before vitrectomy and three after vitrectomy. Visual outcome after vitrectomy and cataract extraction was compared among three groups based on the timing of the second surgery: < or =7 days postcataract extraction; 8-30 days postcataract extraction; and >30 days postcataract extraction. No statistically significant difference in final visual acuity was observed between the three intervals. CONCLUSIONS: The major complication associated with vitrectomy for retained lens fragments in the vitreous cavity after phacoemulsification was retinal detachment. The timing of vitrectomy did not affect the final visual acuity outcome. Visual prognosis was most closely related to the presence of age-related macular degeneration and cystoid macular edema. The type of intraocular lens did not influence the visual outcome. Management with vitrectomy yielded favorable visual results in most patients with retained lens fragments. PMID- 11262669 TI - Conjunctival biopsy in the diagnosis of ocular sarcoidosis. AB - Sarcoidosis is a multisystem granulomatous disease of unknown etiology that can affect almost every organ in the body, particularly the lungs, skin, eyes, and thoracic lymph nodes. A definitive diagnosis of sarcoidosis requires that a biopsy be performed. A specimen can be obtained from any affected ocular structure, including conjunctiva, lacrimal gland, eyelid skin, and orbit. Among them, conjunctival biopsy has been suggested as a sensible and safe procedure for confirming suspected sarcoidosis. We describe three patients in whom ocular symptoms were the sole initial manifestations of sarcoidosis and who were diagnosed based on the results of a conjunctival biopsy. We also discuss the efficacy of and indications for conjunctival biopsy. PMID- 11262670 TI - Incidence of and risk factors for proliferative retinopathy and its association with blindness among diabetes clinic attenders. AB - BACKGROUND: Proliferative diabetic retinopathy (PDR), a prevalent late-stage complication of diabetes, is associated with severe visual loss. The objectives of this report were to estimate the incidence of and risk factors for PDR using routinely collected data from a clinical information system at University Hospital Nottingham for insulin- and non-insulin-dependent (insulin-treated and non-insulin-treated) diabetes separately. We also attempted to assess the risk of blindness in these diabetes clinic attenders. METHODS: During a mean (standard deviation (SD)) follow-up period of 5.1 (2.9) (range 0.5-12.4) years, 3482 diabetic patients (1915 male and 1567 female) from three outpatient clinics at University Hospital, Nottingham were examined. The mean (SD) age of the participants was 49.3 (17.9) years with a mean duration of diabetes of 7.1 (8.7) years at registration. RESULTS: Among the 3482 patients who attended the clinic at least twice in the period 1979-1992, and who were free of PDR at registration, the overall incidence of PDR was 16.2 per 1000 person-years, based on 17,618 person-years of follow-up. The incidence rate of PDR was nearly three times as high among patients with non-proliferative diabetic retinopathy (NPDR) as in those without any retinopathy (42.1 vs. 15.0 per 1000 person-years). Based on a Cox's Proportional Hazards Model, significant independent predictors of PDR recorded at baseline were glycosylated haemoglobin (HbA1), systolic blood pressure, and longer duration of diabetes for patients without PDR or any retinopathy among insulin-dependent patients. Longer duration of diabetes was the only independent predictor of PDR for patients without PDR or any retinopathy in both insulin- and non-insulin-treated non-insulin-dependent patients. These clinic-based data clearly indicate the higher risk of PDR in non-insulin dependent patients. Gender, age, BMI, creatinine, proteinuria and cigarette smoking, had no significant independent association with PDR when other covariates were considered in all groups. The risk of blindness was greater among those with PDR than those with NPDR in all three types of diabetes, but was substantial even for those without retinopathy. CONCLUSION: These data are of value in identifying those diabetes clinic attenders who may be most at risk. PMID- 11262671 TI - Lens thickness and five-year cumulative incidence of cataracts: The Beaver Dam Eye Study. AB - PURPOSE: To evaluate whether lens thickness is related to incidence of cataracts. METHODS: Lens thickness was measured from slit-lamp photographs of the lens at the time of the prevalence evaluation in the Beaver Dam Eye Study. Incident cataract was determined by grading standard slit-lamp and retroillumination photographs of the lens at the baseline and five-year follow-up examinations. Medical history was obtained and blood pressures, height and weight were measured according to protocol. RESULTS: Lens thickness was positively associated with incident nuclear cataract and inversely associated with incident cortical cataract after accounting for age, sex, diabetes status, hypertension, heavy drinking and cigarette smoking. CONCLUSIONS: Lens thickness is related to incidence of cataracts. Mechanisms to explain these relationships require further laboratory and epidemiologic investigation. PMID- 11262672 TI - Prevalence of amblyopia and associated refractive errors in an adult population in Victoria, Australia. AB - The study aimed to describe the prevalence of amblyopia and associated refractive errors among an adult Australian population. The Visual Impairment Project (VIP) is a population-based study of age-related eye disease in the state of Victoria, Australia. Data were collected through standardised interviews and orthoptic and ophthalmic dilated examinations. Amblyopia was defined as best-corrected visual acuity of 6/9 or worse in the absence of any pathological cause. The participants were 3,265 urban residents and 1,456 rural residents of the VIP ranging in age from 40-92 years (mean = 59 years; 53% female). The prevalence of unilateral amblyopia was 3.06% (95% C.I. 2.59, 3.53). Amblyopia was not found to be statistically different by age group (p=0.096), gender (p=0.675), or place of birth (p=0.14). Anisometropia was statistically more common (p<0.001) in amblyopic cases (51.1%) compared to the normal population (9.7%), and 54% of amblyopic eyes had visual acuity of worse than 6/12. Amblyopia is a significant cause of unilateral reduced visual acuity in a population aged 40 years and older. Anisometropia was more prevalent and the degree of anisometropia was greater in the amblyopic group compared with the normal population. Oblique astigmatism was more prevalent in the amblyopic group compared with the normal population. PMID- 11262673 TI - Evaluating the effectiveness of a vision rehabilitation intervention using an objective and subjective measure of functional performance. AB - PURPOSE: To test the hypothesis that vision rehabilitation using optometry, occupational therapy and social work services increases patients' functional ability and to assess whether involving families in the intervention results in more successful outcomes. METHODS: We conducted an outcome study of 97 patients new to the Vision Rehabilitation Service. Subjects were between the ages of 19 and 91 years, with a median age of 76. Their visual acuities were 20/100 or worse in the better eye, with 50% of the subjects having acuities worse than 20/200. Macular degeneration was the most prevalent diagnosis. Subjects were assigned to either an individually focused (n=48) or a family focused (n=49) intervention. The outcome measure was change in function, as assessed by speed and accuracy of performance (objective measure) and by the patients' self-reports of difficulty and dependency in performing daily activities (subjective measures). Data were collected before and after the intervention. RESULTS: Most patients had documented improvement after rehabilitation on both objective (p=.0001) and subjective (decreased dependency, p=.01) measures of function. The sample size did not provide adequate statistical power to show differences between family focused and individually focused interventions. CONCLUSIONS: This study documents significant improvement after vision rehabilitation for a predominantly elderly population. Patients in both family and individually focused interventions improved comparably. PMID- 11262674 TI - Traditional couching is not an effective alternative procedure for cataract surgery in Mali. AB - INTRODUCTION: In Mali, more cataract patients receive sight-restoring surgery using a traditional "couching" procedure (the lens inside the vitreous body) than by modern cataract surgery. In order to evaluate the relative effectiveness and other outcomes of the traditional procedure compared to the modern surgical intervention, we conducted a population-based survey in a rural district of Mali in 1996. METHOD: A total of 99,800 persons from 160 villages were eligible to be included in the sample. All individuals operated for cataract by a modern procedure were checked for visual acuity and questioned regarding their clinical history, the cost of the surgery and their satisfaction with the surgery immediately following the operation and presently. Each patient was paired with one person operated by a traditional cataract surgical procedure. RESULTS: From a total population of 99,800 we found 85 individuals (0.085%) who had been operated by intracapsular extraction (ICCE) without lens implantation and we paired these with 82 individuals operated by the traditional method and by a local healer. In both groups, males were predominant (74.4% in the modern group and 61.3% in the traditional) and the median age was 65 and 68 years, respectively. Men with a higher social status (defined as administrative or religious authority) were slightly more common among those operated by ICCE (18.9%) than among those operated by the traditional healer (4.4%). Nearly half (47.6%) of the patients operated by couching did not know that a modern alternative existed. The mean cost to the patient of the two procedures was similar; with traditional couching costing on average US$ 42.10 and modern surgery (including transport and drugs) costing US$ 52.40. The traditional healer was often paid partially in kind and the price paid varied according to the patient's ability to pay. The clinical results differed greatly between the two methods. After aphakic correction of eyes operated by ICCE, 5.3% had good vision (33/18), 76.8% had low vision (33/60 and <3/18) and 17.9% were still blind (<3/60). Of eyes operated by traditional couching, none had good vision, 29.1% had low vision and 70.9% were blind. The level of satisfaction was high (89.7%) among persons operated in an ophthalmic center by the modern method, and relatively low (22.6%) among persons operated traditionally. DISCUSSION: In Mali, two types of providers offer two different interventions to treat cataract-blind persons. This study suggests that the couching method used by traditional healers is relatively expensive and ineffective. It is also potentially dangerous although this study did not address this question specifically. It is important that health policy makers and medical authorities do what they can to prevent traditional healers from performing the couching procedure, as well as informing the population about the existence of a more-effective and safer alternative. However, while more effective and safer, the results obtained by ICCE are not excellent either. Further, it is important to improve the quality of ophthalmic services in order to provide cataract patients with the best, most accessible and least expensive services possible. PMID- 11262675 TI - Parental age in Indian patients with sporadic hereditary retinoblastoma. AB - There is a consistent correlation between sporadic hereditary retinoblastoma and parental age. It has been proven beyond doubt that the birth rank is correlated with parental age. In the present study, a test for the effect of birth rank was performed in order to assess the risk of developing retinoblastoma with increased parental age. The study of the effect of birth rank showed a significant association between sporadic retinoblastoma (bilateral and unilateral) and late para, indicating that fresh germline mutations must have taken place in some of the sporadic cases. An investigation of the effect of birth rank on familial cases, obtained from published papers and our own series, showed that familial retinoblastoma is significantly associated with early para, suggesting early parental age. Further analysis of the mean paternal and maternal ages of sporadic cases (bilateral and unilateral) showed that the mean paternal age of sporadic bilateral (sporadic hereditary) cases was higher than that of sporadic unilateral cases (p<0.05). No such correlation was seen with mean maternal age. Thus, the present study shows that a high paternal age may be associated with sporadic bilateral (sporadic hereditary) retinoblastoma. PMID- 11262678 TI - Reliability of the VCM1 Questionnaire when administered by post and by telephone. AB - PURPOSE: To assess the reliability of different methods of administration of the VCM1 vision-related quality-of-life questionnaire by: a) comparing responses obtained by post to responses obtained in a research clinic and b) comparing responses obtained by telephone to responses obtained in a research clinic. METHOD: Questionnaire responses given in advance by post (96 subjects) or by telephone (92 subjects) were compared to those subsequently given at a visit to a research clinic. The questionnaire included the VCM1 and two other questions commonly used in surveys of visual impairment (reading small print and recognising a face across the street). RESULTS: Similar levels of vision-related quality-of-life (VR-QOL) impairment were reported by post and in the research clinic. However, the participants in the telephone test group reported less VR QOL impairment by telephone than they subsequently reported in the clinic (P = 0.0001). The mean score difference between telephone and clinic administration was 3.2% of the VCM1 questionnaire scale. Lower social class (P = 0.002) and increasing duration of interview (P = 0.003) were associated with a tendency to under-report VR-QOL impairment by telephone. Interference with reading small print (P = 0.0001) and recognising a face across the street (P = 0.0001) were also under-reported by telephone. CONCLUSIONS: Telephone interviewing caused a general bias towards under-reporting of visual problems which was not confined to the VCM1. Care is required when planning outcome studies and questionnaire surveys to ensure that different methods of questionnaire administration produce comparable results. PMID- 11262679 TI - The economic impact of ophthalmic services for persons with diabetes in the Canadian Province of Nova Scotia: 1993-1996. AB - PURPOSE: If undetected and untreated, diabetic retinopathy can lead to severe vision loss and irreversible blindness, imposing both clinical and economic costs to patients and society. The purpose of this study, therefore, was to determine both the direct and indirect costs of providing ophthalmic, social and rehabilitative services for persons with diabetic eye disease. METHODS: Persons with diabetes and seen by ophthalmologists in the Canadian Province of Nova Scotia were ascertained from provincial health records for the years 1993-1996, inclusive. In addition, utilization data from community and blindness rehabilitation agencies located throughout the Province of Nova Scotia for the same time period were also obtained and analyzed. A cost-of-illness analysis of ophthalmic and additional services for persons with diabetes was then performed using the available data sources. RESULTS: The total cost of direct and indirect ophthalmic, disability and rehabilitative care for persons with diabetes mellitus in Nova Scotia over the study period, 1993-1996, was estimated to be CDN $10,521,816.58. Direct costs amounted to CDN $1,416,355.05 (13.46%), indirect costs due to lost productivity reached CDN $5,072,831.85 (48.22%), rehabilitation services cost CDN $251,204.08 (2.38%), disability payments amounted to CDN $979,992.00 (9.32%), while lost wages due to disability payments measured a further CDN $2,801,433.60 (26.62% of total costs). CONCLUSIONS: Over the period reviewed, ophthalmic costs rose by a factor 3.2 greater than that observed for goods and services captured by the Canadian Consumer Price Index. Further analyses over a longer follow-up period are required to definitively establish whether or not there is a long-term upward trend in ophthalmic costs. PMID- 11262680 TI - Statistical analysis when dealing with astigmatism: assessment of different spherocylindrical notations. AB - Ophthalmic epidemiological studies frequently deal with ocular refractive errors, which are commonly expressed in the form sphere/cylinder x axis. However, this representation has been shown not to be the most suitable one for performing statistical analysis. Although alternative analytical and graphic methods to represent this kind of data have been developed, these formalisms have often gone unnoticed by researchers, despite their usefulness and versatility. Besides, there has been no discussion of how each of them fits in with a particular type of study. In this paper, several mathematical representations of dioptric power are revisited in a comprehensive way. The aim is to encourage researchers in ophthalmology and optometry to use these formalisms in their epidemiological studies, thus profiting from their exactitude and simplicity. Consequently, the emphasis is not on complicated mathematical derivations but on how to use these representations. Their potential and suitability in different applications is analyzed in detail. In addition, some examples are presented to illustrate the mathematical methods considered. PMID- 11262682 TI - Uncorrected binocular distance visual impairment in U.S. Hispanic children and adolescents. AB - PURPOSE: To assess and compare uncorrected binocular distance visual impairment rates in U.S. Hispanic children and adolescents. METHODS: Data from the Hispanic Health and Nutrition Examination Survey, 1982-1984, were analyzed for 6-19 year old Cuban-Americans (n = 317), Mexican-Americans (n = 2519), and Puerto Ricans (n = 988). Visual acuity was assessed using Sloan Letters or Landolt Rings. RESULTS: Prevalence rates of uncorrected binocular distance visual impairment (20/30 or worse) were 15.5%, 14.9%, and 23.6% for Cuban-Americans, Mexican-Americans, and Puerto Ricans, respectively. After adjusting for age and gender, the differences between Puerto Ricans and both Cuban-Americans and Mexican-Americans were significant (p < 0.05). Children 6-12 years of age had lower visual impairment rates than 13-19 year-old adolescents. Girls had higher age-adjusted visual impairment rates than boys; these gender differences were statistically significant among Mexican-Americans (OR = 1.6, 95% CI = 1.1, 2.2) and Puerto Ricans (OR = 1.7, 95% CI = 1.2, 2.4). CONCLUSIONS: Among Hispanics, Puerto Rican children and adolescents have the highest prevalence rate of uncorrected binocular distance visual impairment; older age and female gender are associated with higher rates of uncorrected visual impairment. PMID- 11262681 TI - Gender and blindness: a meta-analysis of population-based prevalence surveys. AB - BACKGROUND: Many individual surveys of blindness have reported slightly higher rates of blindness for women. In order to gain a continent-by-continent and global sense of the burden of blindness by sex we conducted a meta-analysis of published, population-based surveys of blindness. METHOD: Published reports were collected using a predetermined search protocol involving commercial electronic databases, hand-searching of references and direct contact with researchers. Studies were included that were population-based, included clinical examination and had a minimum sample size of 1000. The studies were critically appraised to determine methodological rigour. Data were analysed using the Cochrane Collaboration Review Manager. RESULTS: The overall odds ratio (age-adjusted) of blind women to men is 1.43 (95% CI 1.33-1.53), ranging from 1.39 (95% CI 1.20 1.61) in Africa, 1.41 (95% CI 1.29-1.54) in Asia, and 1.63 (95% CI 1.30-2.05) in industrialised countries. There was good homogeneity of findings from Africa, Asia, and the industrialised countries. Globally, women bear excess blindness compared to men. In these surveys, overall, women account for 64.5% of all blind people. The excess of blindness in women was marked among the elderly and not due only to differential life expectancy. CONCLUSION: The excess burden of blindness among women is likely due to a number of factors, which are different in industrialised countries compared to developing countries. Particular attention to gender differences in blindness is needed in the creation of targets for blindness reduction and in the development of interventions. PMID- 11262683 TI - The optimal stimulus to elicit suppression in small-angle convergent strabismus. AB - PURPOSE: To determine the optimal stimulus duration as well as the most appropriate luminance profile to elicit suppression in small-angle convergent strabismus. METHODS: In 10 subjects with small-angle convergent strabismus, using a device allowing binocular viewing and peripheral fusion, we determined what the optimal stimulus would be to elicit suppression. Three control subjects were also included in the study. Stimuli were shown randomly in the central 3 degrees of the visual field of either eye. Stimulus durations were varied in seven steps from 50 to 1000 ms and three luminance-time profiles were used: square wave, triangle and half-sinus, thus yielding 21 different stimuli. The peak light intensity was the same for all stimuli. RESULTS: Suppression, defined as the difference in the threshold sensitivities under monocular vs. binocular viewing, was found with our test device in five of the ten subjects, and ranged between 3 and 33 dB. Suppression was deepest with triangular or half-sinusoidal stimuli of 400 ms duration. Square wave stimuli elicited the smallest amount of suppression. CONCLUSION: Stimuli with a gradual increment and decrement, like triangular or half-sinusoidal stimuli, with a duration of 400 ms are the most effective to elicit suppression PMID- 11262684 TI - Preliminary report: monocular spatial localization in children with strabismic amblyopia. AB - Defective spatial localization is an important feature of strabismic amblyopia. Based on our experience from testing adult strabismics under various test conditions, we developed a test for assessing vertical alignment in strabismic children. Patients had to align a vertical test line with the apices of two vertically arranged reference triangles, under the control of both the dominant eye and the amblyopic eye. Means and standard deviations of several judgements represent systematic errors and uncertainty of alignment. We tested 27 strabismic and 34 age-matched control children aged 4.5-10 years. Control children showed a scatter of mean systematic alignment around the correct position of up to 7 minarc. In the amblyopic eyes of strabismic children, uncertainty was consistently higher than in the eyes of the control children. Systematic errors outside the normal range frequently occurred. In children tested repeatedly during occlusion therapy, uncertainty decreased as visual acuity improved. In several cases we observed changes of systematic vertical alignment during therapy, sometimes unexpectedly in the sense of a change in the direction of mislocalization or an initial increase and later decrease of errors. Thus, children with strabismic amblyopia show spatial localization deficits which are similar to those of adult strabismic amblyopes. Both spatial uncertainty and systematic distortions are susceptible to change due to enforced use of the amblyopic eye during occlusion therapy. PMID- 11262685 TI - Minimal invasive decompression of the orbit in Graves' orbitopathy. AB - BACKGROUND: Surgical decompresssion of the orbit may be considered as a suitable form of therapy if, as a result of the increased volume inside the orbit, there are motility disorders such as diplopia or a progressive decrease in visual acuity. In view of the fact that this operation will not cure the underlying disease, the treatment should be as mild as possible. METHODS: In five subjects with Graves' orbitopathy we managed to extend the volume of the intraconal orbit by microsurgical liposuction. We carried out a lateral canthotomy to approach the orbit behind the globe. After decompression of the soft tissue, the lateral palpebral ligaments were refixed. To assess the scale of the functional rehabilitation we compared preoperative parameters (visual acuity, Hertel's index, visual field, motility) with the postoperative results. RESULTS: In all cases we found a significant improvement of position and motility without signs of diplopia. There was a postoperative increase in the visual field and visual acuity was 0.4 and the protusion of the globe could be decreased by 3-6 mm (Hertel's index). Furthermore, the ocular hypertension we found preoperatively could no longer be detected after the operation. CONCLUSIONS: Microsurgical decompression of the soft tissue via an approach from behind the globe proved to be a very gentle alternative to conventional methods of orbital decompression because of the satisfying functional and esthetic rehabilitation in selected cases. PMID- 11262686 TI - Stimulus deprivation amblyopia in human congenital ptosis: a study of 100 patients. AB - AIM: To investigate the frequency of stimulus deprivation amblyopia (SDA) in comparison with other reasons for amblyopia in human congenital ptosis. METHODS: The frequency and causes of amblyopia were evaluated in the 200 eyes of 100 patients. Congenital ptosis was present in 128 eyes (72 unilateral, 28 bilateral). The age at investigation was one year and older, with an average of 11 years and 10 months. Amblyopia was defined as best corrected visual acuity less than 1.0 or a difference between the two eyes of at least 0.2. The following causes of amblyopia were identified: amblyopiogenic refractive errors: astigmatism > or = 1 dpt, anisometropia > or = 1 dpt (79% cycloplegia) and strabismus. In cases with no other reasons for amblyopia, SDA was assumed. Statistical analysis was performed using the chi-square and the sign tests. RESULTS: The overall incidence of amblyopia in ptotic eyes was 89/128 (70%). In 3.9% of the cases (5/128; 2 eyes with unilateral and 3 eyes with bilateral ptosis) we assumed SDA. A comparison of ptotic eyes with (unilateral: n = 35, bilateral: n = 34) and without covered optical axis revealed the following: in the case of unilateral ptosis, amblyopia was found more often in ptotic eyes with covered optical axis: 30 out of 35 vs. 24 out of 37 (p = 0.06); in the case of bilateral ptosis this difference was significant: 27 out of 34 vs. 8 out of 22 (p < 0.05). In the case of SDA, the optical axis was covered in only a single eye, in a patient with bilateral ptosis. There was no difference in the incidence of anisometropia: 19 out of 53 vs. 14 out of 47 (p = 0.52). Astigmatism was found more frequently in ptotic eyes with covered optical axis in unilateral ptosis: 23 out of 35 vs. 16 out of 37 (p = 0.06) but not in bilateral ptosis: 21 out of 34 vs. 13 out of 22 (p > 0.9). Strabismus was found significantly more frequently in ptotic eyes with covered optical axis: 13 out of 35 vs. 4 out of 37 (p < 0.05) in unilateral ptosis and 7 out of 34 vs. 1 out of 22 (p = 0.13) in bilateral ptosis. CONCLUSION: In contrast to the classical animal models of stimulus deprivation amblyopia, this entity is rare in human congenital ptosis, perhaps because of the counter effect of compensating head posture. Disruption of fusion resulting in strabismus might be an additional indirect cause of amblyopia in congenital ptosis. Prophylactic amblyopia treatment in ptosis cases is important as long as no testing of visual acuity is possible in a child. PMID- 11262687 TI - Preliminary report: Dynamic stereopsis in patients with impaired binocular function. AB - In this study, 46 strabismic patients aged between 9 and 58 years were tested for dynamic stereopsis in the peripheral visual field with up to 20 degrees eccentricity. Squint angles ranged from +30 to -36 degrees. The effect of surgical realignment of the visual axes on dynamic stereopsis was tested before and after surgery in 40 of these patients. Of the 46 patients, 23 had esotropia and 23 exotropia. A test device was used which presented two projected squares in polarized light (each square being perceived monocularly through a polarization filter) in horizontal motion, thus creating a three-dimensional impression. Patients were tested qualitatively for dynamically stereoactive fields of vision and quantitatively for the threshold value needed to create a three-dimensional impression. We found residual dynamic stereopsis in 30% of patients who had no central static stereopsis. 56% of the patients improved after surgery, either through a significant (p < 0.01) gain of stereoactive fields or through a decrease in threshold values. This emphasizes the necessity of creating a test device suitable for everyday clinical use. Strabismus surgery is especially beneficial in connection with traffic and sports medicine, regardless of the effect on classical tests. PMID- 11262688 TI - Frequency of reading disability caused by ocular problems in 9- and 10-year-old children in a small town. AB - INTRODUCTION AND PURPOSE: In most children referred to our department with a diagnosis of dyslexia, we have found an ocular disorder that had not been detected during previous ophthalmologic examinations. Exophoria and/or hypoaccommodation were the most common cause. Some of these children needed eye muscle surgery to improve the reading problems. However, these patients represent a selection. Therefore, we performed a field study to determine the percentage of children with reading disability caused by ocular disorders and the percentage of children with real dyslexia in a normal population. This was made possible by an examination of most pupils in the 4th grade of the three primary schools in a small German town. The co-author and a very experienced orthoptist performed all of the examinations. RESULTS: Eighty-nine out of 127 children were examined. Of these, 16 (18%) had reading problems (2 girls and 14 boys). Most of them had accommodation problems: six (6.7%) suffered from an uncorrected hypoaccommodation, three children did not wear their prescribed glasses, one child had not been prescribed any glasses yet and one child had the wrong glasses. Two children suffered from pathophoria: one from eso- and the other from exophoria compensated by accommodative convergence. In 3 (3.4%) children no ocular cause could be found. These children may have true dyslexia. CONCLUSION: Of the 89 children examined, 16 (18%) had reading problems and only 3/16 had no detectable ophthalmologic explanation. Hypoaccommodation was the most common cause of reading problems (in 6 of 16). In most of the cases it had not been diagnosed before. In all of these children the reading ability improved markedly with the proper refractive correction, bifocals or prisms. PMID- 11262689 TI - Preliminary report: analgesia with Remifentanil for complicated eye muscle surgery. AB - The authors report a new anesthetic technique that is especially suitable for the surgery of complicated disorders of ocular motility. Analgesia is achieved by an intravenous dose of Remifentanil, an opioid that is normally used together with propophol as a form of total intravenous anesthesia (TIVA) for general anesthesia. The advantage of Remifentanil is a rapid increase (1-2 min.) and decrease (3-4 min.) of the effect. Eye muscle surgery can then be performed on a patient who is free of pain and slightly sedated. After the planned eye muscle surgery, while the patient is still under anesthesia, the muscle sutures are tied provisionally. Afterwards, the squint angle can be measured with a cover test and the patient is questioned about the presence of diplopia. If the position of the eye is not satisfactory, the knots can be loosened and the muscle can be fixated in another position. PMID- 11262690 TI - The cost-effectiveness of screening strategies for amblyopia: a preliminary report. AB - Five screening strategies for amblyopia in different age groups were compared according to a decision-analytical model from the perspective of the health insurance funds. Our findings indicate that the costs per detected case of amblyopia range from about 1200 DM to 3000 DM (613 Euro to 1534 Euro). The two most cost-effective screening strategies were to screen high-risk children up to the age of one by ophthalmologists and to screen all children up to the age of one by ophthalmologists. The screening of high-risk children identifies only about a third of all affected children in this age group, when compared with the number of cases detected by screening all children up to the age of one. However, the average cost per detected case of amblyopia among high-risk children is lower than the cost of screening all children in this age range. PMID- 11262694 TI - An approach to the surgical management of total oculomotor nerve palsy. AB - The goal of the procedure was to keep the eyes of patients with total oculomotor palsy in the straight ahead position by means of surgery on the horizontal and inferior rectus muscles in one session, without involving the superior oblique muscles. Six patients underwent surgery for total oculomotor nerve palsy. All of the surgical procedures were carried out on the muscles of the paralytic eye. We performed hemi-hangback recession of the lateral rectus and resection of the medial rectus for exotropia in all patients. Depending on the magnitude of vertical deviation, the insertions of the horizontal rectus muscles were moved upward, alone or in combination with hemi-hangback recession of the inferior rectus. The mean preoperative horizontal deviation was 66.6 PD. Two years after the operation, the horizontal deviation was measured to be 11.6 PD. Similarly, the mean preoperative vertical deviation of 16 PD decreased to 6.6 PD in two years. This procedure did not disturb normally functioning superior oblique and lateral rectus muscles. Subjectively, all of the patients were satisfied with their alignment two years after the operation. We are of the opinion that this technique is a safe, simple and effective procedure and can be regarded as a first-choice operation in total oculomotor palsy. If one fails to maintain the eye position with this procedure, one can still perform a second operation on the superior oblique muscle, which remains untouched in our procedure. PMID- 11262695 TI - Cocaine abuse, generalized myasthenia, complete external ophthalmoplegia, and pseudotonic pupil. AB - We present the case of a 29-year-old woman with generalized myasthenia. Myasthenia with complete external ophthalmoplegia was unmasked by cocaine abuse. It was associated with changes of the pupillary motility, including light-near dissociation and positive 0.1% pilocarpine test. Treatment with acetylcholinesterase inhibitors improved the patient's condition rapidly, and led to complete normalization of extraocular movements and pupillary function. To our knowledge, this is the fourth case of cocaine-related myasthenia, and the first case of myasthenia with pseudotonic pupil. PMID- 11262696 TI - Ophthalmodynamometry: a reliable method for measuring intracranial pressure. AB - Under physiological conditions, the pressure in the central retinal vein is equal to or higher than the intracranial pressure (ICP) because the cerebrospinal fluid (CSF) passes the sheath of the optic nerve before draining into the cavernous sinus. The optic nerve sheath is where the ICP affects the retinal venous pressure. Ophthalmodynamometry (ODM) is a useful method for determining the central retinal artery pressure. While papilledema and a lack of venous pulsations are commonly used as a vague indication of the ICP, ODM may be advantageous for determining the pressure in the central retinal vein. Until now, however, the venous pressure has never been compared with the intracranial pressure. In the present study, the pressure in the central retinal vein was recorded in 31 patients while the ICP was simultaneously being recorded for various reasons. The results demonstrate a linear correlation (r = 0.968) between the pressure in the central retinal vein and the ICP. This correlation is of great practical value since until now, reliable intracranial pressure monitoring has only been possible by invasive means, by placing a probe either in the brain parenchyma or the ventricle. Ophthalmodynamometry is useful for momentary assessment of the ICP, can easily be repeated, and may be used whenever an elevated ICP is suspected in hydrocephalus, brain tumors and after head injury. However, it is not suitable for continuous ICP monitoring. PMID- 11262697 TI - Preliminary report: prescription of prism-glasses by the Measurement and Correction Method of H.-J. Haase or by conventional orthoptic examination: a multicenter, randomized, double-blind, cross-over study. AB - In a multicenter, randomized, double-blind, cross-over study in the Netherlands, the effectiveness of (prism-)glasses prescribed by the Measurement and Correction Method of H.-J. Haase (MKH) was compared to that of glasses prescribed by conventional orthoptic examination. Nine pairs of MKH-optometrists and orthoptists recruited patients who primarily presented with asthenopia, and each prescribed the patient (prism-)glasses. A questionnaire for asthenopia was developed that rated headache and tired eyes as 0-7 days per week and none-light medium-severe, respectively. Light sensitivity, problems with focusing, near-work problems and burning eyes were each rated as: never-occasionally-often-always. A patient was eligible if he scored 'medium', 'often' or '5 days a week' twice; or 'medium' (etc.) once and 'light' (etc.) twice. Controls, in contrast to the patients, typically answered 'none' or 'never' to half of the complaints, but 37% of them would have passed the admission criteria. Among other criteria were: 18 to 40 years of age, horizontal angle < 4 degrees, vertical < 1.7 degrees, acuity > or = 0.8, stereopsis threshold disparity < 120". Seventy-two patients fulfilled all criteria and returned sufficient questionnaires. They wore the first glasses for six weeks, were without glasses for two weeks, and then wore the second glasses for six weeks. At the start, halfway and at the end of each 6-week period, questionnaires were filled out; 97% were returned. Only 19 of the orthoptists' glasses contained prisms (14 horizontal, 5 vertical; horizontal average of all glasses 0.49 PD, vertical 0.05 PD). Five of the orthoptists' glasses were plano. All MKH glasses contained prisms, 53 of 72 both horizontal and vertical, 18 only horizontal and one only vertical (horizontal average of all glasses 2.83 PD, vertical 0.79 PD). The starting levels of complaints were high and both glasses improved complaints dramatically. The starting levels were lower, but not significantly, in the second 6-week period and improvement was less outspoken. Because of these differences, the two periods had to be evaluated separately. The primary outcome of the study was defined as the difference between the effect of the MKH glasses and that of the orthoptists' glasses in the first and second 6-week periods. For problems with focusing, in the first 6-week period, and for tired eyes, in the second 6-week period, the difference exceeded the difference that had been defined as clinically significant (one day per week less headache or half the distance light-medium or half the distance occasionally often), but it did not reach statistical significance. The statistical power was approximately 0.7 for demonstrating this clinically significant difference. Statistical significance was not reached in multivariate repeated measure ANOVA either. Forty-four patients preferred to keep the MKH glasses, 25 the orthoptists' glasses, including one plano. It is striking that 25% of the patients did not prefer the glasses that, according to the questionnaire, improved their complaints the most. A year after the study, the questionnaire was sent again to all patients: The levels of complaints after a year were similar to those at the end of the second 6-week period, whether they had preferred the MKH or the orthoptists' glasses, and were similar to the levels in controls. The most conspicuous finding was that both glasses improved the complaints dramatically. Apart from the prisms, other reasons could be: spherical and cylindrical correction, improved wearing comfort of the frame, placebo effect, Hawthorne effect and regression to the mean. PMID- 11262698 TI - Binocular 3-D video-oculography. AB - A video technique is described to record ocular motions and positions of both eyes simultaneously in all degrees of freedom. This non-invasive method allows a 3-D positional analysis with free gaze directions and head tilts for measuring all ocular degrees of freedom, including torsion. PMID- 11262699 TI - Unilateral congenital oculomotor nerve palsy, optic nerve hypoplasia and pituitary malformation: a preliminary report. AB - A newborn male presented with complete external third nerve palsy of his right eye immediately at birth. Pediatric examination and MRI of the skull revealed no abnormalities. At the age of six weeks, strabismus surgery was performed to facilitate amblyopia treatment. The muscles appeared small and fibrotic. At the age of ten weeks, a brow suspension of the upper lid and a second strabismus surgery were performed. The amblyopia treatment and patching, applied for half of the waking hours over a period of six weeks, were unsuccessful. At the age of six months, a relative pallor of the right optic nerve head became evident. At the age of three years, at a new examination because of growth deficiency, a second MRI revealed defects involving the pituitary region. We concluded that extraocular muscle abnormality or oculomotor nerve palsy was present together with optic nerve dysplasia and pituitary gland malformation. PMID- 11262700 TI - American Board of Clinical Neuropsychology special presentation: The American Board of Clinical Neuropsychology (ABCN), 2000 update. AB - This paper updates neuropsychologists on the process of obtaining board certification in clinical neuropsychology through the American Board of Clinical Neuropsychology (ABCN), a specialty board operating under the auspices of the American Board of Professional Psychology (ABPP). At this time, the ABPP and ABCN have certified 406 clinical neuropsychologists, which makes it the largest board certification organization in clinical neuropsychology. This article details the advantages of board certification through the ABCN and the four steps which must be passed in order to obtain board certification. These steps are: credential review, written examination, work sample, and oral examination. PMID- 11262701 TI - Comparison of the CERAD and CVLT list-learning tasks in Alzheimer's disease. AB - This investigation examined the relationship of the word list from the CERAD neuropsychological battery to the California Verbal Learning Test (CVLT) in a sample of 138 subjects with Probable Alzheimer's disease (AD). Results revealed modest but statistically significant associations between the two measures on many key variables. Total words learned showed the strongest association, with lower correlations for delayed recall, intrusion errors, and recognition variables. As expected, the CERAD and CVLT assess similar aspects of verbal learning in patients with AD. However, the modest level of many of the correlations suggests that caution should be exercised in applying the same interpretive strategies derived on more comprehensive measures to shorter ones. PMID- 11262702 TI - Norms for the Wisconsin Card Sorting Test in 6- to 11-year-old children in Taiwan. AB - The main aims of this study were to develop norms for the Wisconsin Card Sorting Test in 6- to 11-year-old children in Taiwan; to explore the effect of sex, age, birth order, number of siblings, and parental education on WCST performance in 6- to 11-year-old children; and to make a comparison of WCST performance between children in Taiwan and the USA. The results of this comparison of developmental norms of school children in Taiwan and the United States may facilitate the WCST as a clinical or research instrument in combination with other test procedures to assess aspects of cognitive and neuropsychological functioning of school children. PMID- 11262703 TI - Factor analysis of computerized and traditional tests used in mild brain injury research. AB - The present study examines the relation between a set of computerized neuropsychological measures, Automated Neuropsychological Assessment Metrics (ANAM), and a set of traditional clinical neuropsychological tests. Both sets of tests have been employed in recent studies of mild brain injury. Factor analysis and stepwise regression indicate that both sets of tests measure similar underlying constructs of cognitive processing speed, resistance to interference, and working memory. The present findings indicate strong concordance between computerized and traditional neuropsychological measures and support the construct validity of ANAM and similar procedures. PMID- 11262704 TI - Differential impact of executive dysfunction on verbal list learning and story recall. AB - The California Verbal Learning Test (CVLT) and the Logical Memory (LM) subtest from the Wechsler Memory Scale-Revised (WMS-R) are generally thought to be interchangeable measures of verbal memory. However, little is known about the effects of executive dysfunction on these tasks. The present study involved 96 patients referred for neuropsychological evaluation who were classified as having either significant executive dysfunction (SED) or minimal executive dysfunction (MED) based on the number of impaired executive tasks. Results showed that the SED group performed significantly worse on CVLT total words learned and most of the recall conditions compared to the MED patients (p <.01). However, performance on both immediate and delayed LM did not significantly differentiate the groups. CVLT measures of semantic clustering, perseveration, intrusions, and false positive errors did not appear to account for the group differences. The current study strongly suggests that the CVLT and the LM subtest are differentially associated with executive dysfunction, and argues for the inclusion of both types of tasks in a comprehensive neuropsychological evaluation. PMID- 11262705 TI - Performance of older depressed patients on two cognitive malingering tests: false positive rates for the Rey 15-item memorization and dot counting tests. AB - To our knowledge, no investigations have been undertaken to determine whether depression impacts performance on two commonly used tests to detect malingering of cognitive symptoms, the Rey 15-item Memorization Test and the Rey Dot Counting Test. This is a critical issue because of the high rate of depressive symptoms in patients with neurological conditions. It was hypothesized that depressed individuals, especially those with more severe depression, might be at risk for failing the tests, because these patients exhibit mild deficits in mental speed, visual perceptual/spatial skills, and visual memory, abilities required for successful completion of the malingering tests. However, examination of test performance in 64 older participants with major depression generally revealed very low false positive rates for most test scores, and severity of depression was unrelated to test scores. These results add to accumulating data supporting the validity of these cognitive malingering tests by documenting few false positive identifications. PMID- 11262707 TI - Elderly norms for the Hopkins Verbal Learning Test-Revised. AB - The present study evaluates the effects of age, education, and gender in a representative sample of older adults and provides normative data for community dwelling elderly. Age and gender had significant effects on HVLT-R performance. We provide age- and gender-adjusted normative data. Surprisingly, education level did not affect HVLT-R performance, indicating that education-adjusted norms are not necessary for this measure within this age range. We evaluated a subsample of subjects census-matched on age, education, and gender. These subjects did not differ in overall performance from our entire sample. Therefore, the normative data provided in this paper can be considered to be census-comparable for age, education, and gender. PMID- 11262706 TI - Normative data on neuropsychological tests for very old adults living in retirement villages and hostels. AB - Normative data on neuropsychological tests for very old adults living in retirement villages and hostels are under-represented in the literature. This study reports normative data on the Mini-Mental State Examination, Digit Span Forwards, Digit Span Backwards, the Digit Symbol Substitution Test, the Controlled Oral Word Association Test, the Stroop Neuropsychological Screening Test and the National Adult Reading Test. Age and education showed moderate correlations with neuropsychological test performance. For all tests except the Stroop, differences between residents of retirement villages and hostels were explained by age and education. Men performed better on the NART than women, but this difference was eliminated when education was controlled for statistically. PMID- 11262708 TI - Limited accuracy of premorbid intelligence estimators: a demonstration of regression to the mean. AB - Regression-based premorbid intelligence estimators have been devised by Barona, Reynolds, and Chastain (1984), Barona and Chastain (1986), Hamsher (1984), Krull, Scott, and Sherer (1995; the Oklahoma Premorbid Intelligence Estimate: OPIE), and Vanderploeg, Schinka, and Axelrod (1996; BEST-3 approach), but little is known of their relative accuracy, particularly in outer ranges of intellectual ability (e.g., below-average, superior, etc.). Towards this end, the Wechsler Adult Intelligence Scale-Revised (WAIS-R) was administered to 150 neurologically normal adults, and estimated VIQ, PIQ, and FSIQ scores were computed according to each regression method. Results showed that methods based solely on demographic factors were most susceptible to meanward regression, rendering them poor estimators of IQ scores in outer ranges. Although the OPIE and BEST-3 performed somewhat better, their accuracy remained relatively weak. The findings suggest that regression-based estimates of premorbid IQ are very susceptible to error, particularly in outer ranges of intellectual function. PMID- 11262709 TI - What Persian Gulf War syndrome? AB - Recent reports of physical and neuropsychological syndromes putatively associated with service in the Persian Gulf War and ostensibly providing evidence for Gulf War Syndrome (GWS) are critically reviewed. Major methodological weaknesses are identified in the studies and it is contended that there is no solid evidence for GWS at this time. Suggestions are given for future investigations of symptoms associated with service during the Gulf War which may accurately lead to a tangible identification of a war-related illness entity. PMID- 11262710 TI - Gulf War illness research: separating the wheat from the chaff. AB - Clinical research of veterans' illnesses from the neuropsychology and medical literature are reviewed. Some studies reveal no significant findings, while others indeed detect a higher incidence of clinical and laboratory abnormalities in veterans of Operations Desert Storm and Desert Shield. Neuropsychological deficits are negligible and more often associated with affective, than cognitive, disruption. Some explanations of the results are offered, as are recommendations regarding the utility of clinical research. PMID- 11262711 TI - Through the looking glass: Where is the thought? A reply to Rodriguez-Menendez. AB - In reply to my presidential address, Rodriguez-Menendez questions my comments about poorer performance on the national licensing exam by graduates of Psy.D. programs and professional schools than by Boulder-model Ph.D. programs. He goes on to confirm that their scores are indeed significantly lower, but suggests this is not important. Apart from this apparent confirmation, the remainder of his assertions that Psy.D. programs indeed provide good scientific training appear to be contraindicated by both the model and the performance of graduates of those programs. Finally, the need for standards is again asserted, with a reiteration of the need to ask who benefits from attacks on upholding standards: the patient and profession or the one who is doing the attacking. PMID- 11262712 TI - Division 40 special presentation: listing of training programs in clinical neuropsychology--2000. PMID- 11262714 TI - Two-tailed versus one-tailed base rates of discrepancy scores in the WAIS-III. AB - Tables included in the WAIS-III report the frequency of discrepancies between IQ and index scores independent of the directionality of the score. If an examiner does not have any a priori hypothesis about which skill may be more developed and which skills may be weaknesses, it is appropriate to use these tables as they are. In this case the examiner would be looking for unusually large discrepancies between scores irrespective of direction. When an examiner has a hypothesis about which skills may be weaknesses and strengths for an individual, the frequencies that are based upon an absolute value of the discrepancy will cause examiners to overestimate the frequency of the occurrence of the discrepancy score in question. Sattler and Ryan (1998) and Tulsky, Zhu, and Vasquez (1998) suggested dividing the frequencies reported in the WAIS-III tables in half to obtain the correct base rate. This suggestion is tested in this paper. New observed frequency tables were derived from the WAIS-III and WMS-III standardization samples and these frequencies were compared against the estimated frequencies using the method described by Sattler and Ryan (1998) and Tulsky et al. (1998). The differences between these two methods were calculated and are, for the most part, insignificant. In light of the similarity between the methods, the implications of using observed frequencies versus estimated frequencies is discussed. PMID- 11262715 TI - Co-norming the WAIS-III and WMS-III: Is there a test-order effect on IQ and memory scores? AB - Test-order effect on the WAIS-III and WMS-III scores was evaluated using the WMS III standardization sample. Participants completed the standardization editions of the WAIS-III and WMS-III in one session, with the tests administered in roughly counterbalanced order. Repeated measure MANOVA analyses were conducted to determine if there was an overall test-order effect for subtest, index, or IQ scores. No significant test-order effects were found for either the WAIS-III index or IQ scores or for the WMS-III index scores. At the subtest level, the majority of the WAIS-III and WMS-III subtests did not show a significant test order effect. The exceptions were Digit Span and Digit Symbol-Coding on the WAIS III and Faces II and Logical Memory II on the WMS-III. Although statistically significant test-order effects were found on these subtests, the effect sizes were small. This study indicates that the test-order effect is not a potential threat to the internal validity of the WAIS-III and WMS-III normative data. The practical implications of the current study are discussed. PMID- 11262716 TI - Comparability of the expanded WMS-III standardization protocol to the published WMS-III among right and left temporal lobectomy patients. AB - We examined whether differences between the expanded standardization protocol (SP) used to derive norms for the final published version (PB) of the Wechsler Memory Scale - Third Edition (WMS-III; Wechsler, 1997a) would result in differences on the Primary Indexes in a neurologic sample. Specifically, we examined the comparability of the performances of 63 patients with temporal lobectomy (TL) who were administered either the expanded SP protocol (n = 33: 22 left TL and 11 right TL) or the PB battery (n = 30: 11 left TL and 19 right TL). Patients who were administered the SP or PB were comparable in terms of age, sex, education, seizure duration, postsurgical seizure status, and Full Scale IQ. Postoperative intervals were significantly longer for the SP group, although correlational analyses demonstrated no significant relationship between postoperative follow-up interval and WMS-III performance. A series of t tests revealed no significant differences on any of the eight Primary Index scores between patients taking the two versions of the WMS-III for either left or right TL groups. Furthermore, repeated measures analyses of variance failed to show significant differences on modality-specific memory scores between the SP and PB for the left and right TL groups. The current study indicates that temporal lobectomy patients obtained comparable scores on the two versions of the WMS-III. PMID- 11262717 TI - Could test length or order affect scores on letter number sequencing of the WAIS III and WMS-III? Ruling out effects of fatigue. AB - The Letter Number Sequencing subtest of the WAIS-III and WMS-III was administered at the end of the standardization edition of the WMS-III. It was not administered as part of the WAIS-III standardization battery. Nevertheless, the subtest was included in the published version of the WAIS-III. This study examines differences between examinees administered the Letter Number Sequencing subtest at three different times during a psychological battery: (1) as part of the published battery, (2) as part of the WMS-III when the WMS-III was administered as the first test in a sequence, and (3) as part of the WMS-III standardization when the WAIS-III was administered immediately preceding the WMS-III. The participants were 372 examinees ( n = 124 in each condition) who were matched on key demographic variables. A repeated measures MANOVA yielded no difference in subtest scores when administered in any of these conditions. The results show no evidence of fatigue or ordering effects on the Letter Number Sequencing subtest. PMID- 11262718 TI - Ten-year follow-up survey of clinical neuropsychologists: part II. Private practice and economics. AB - Analyses of a 10-year follow-up survey of clinical neuropsychologists demonstrated significant changes in employment settings away from institutions, placing a clear majority of the field in private practice settings in 1999 (Sweet, Moberg, & Suchy, 2000). The present paper compares characteristics of practices and beliefs of clinical neuropsychologists who work in institutions versus private practice, using data from 1989, 1994, and 1999. Previous survey data had not been analyzed along the dimension of work setting. Among the significant findings are differences in age, referral sources, hours per week engaged in specific professional activities (clinical, neuropsychological, forensic, supervisory, research, teaching), ages of patients, type and frequency of data gathered in assessments, hours spent per evaluation, use of an assistant to gather data, and journal subscriptions. Economic changes within the last 5 years have had a differential impact for the two groups in terms of yearly income and hourly reimbursement. However, approximately half of the neuropsychologists in both groups have increased hours performing clinical work, hours performing administrative duties, and patient load to compensate for economic changes in the last 5 years. Decreases in clinical research and teaching activities are apparent in both groups, but in different amounts. PMID- 11262719 TI - Influence of demographic variables on neuropsychological test performance after traumatic brain injury. AB - The validity of correcting for demographic variables when considering neuropsychological test scores was evaluated in a sample of 136 patients with traumatic brain injury (TBI) who had been screened carefully for premorbid or comorbid confounding factors. When considered in concert with neurological variables, age and education accounted for a significant proportion of the variance in raw scores on the Category Test and the Trail Making Test in the complete sample. Gender did not affect level of test performance. Correcting neuropsychological test scores for demographic variables did not significantly alter their success in identifying patients with severe TBI, but did lead to greater accuracy when classifying individuals with mild-moderate TBI. This investigation concluded that norms that consider the demographic background of the individual are likely to reflect more accurately the neuropsychological status of patients with TBI than interpretations that are based exclusively on raw data. PMID- 11262720 TI - Cognitive impairment with no dementia (CIND): longitudinal studies, the findings, and the issues. AB - Identification of persons at risk for developing dementia is of increasing importance as the proportion of persons over the age of 65 years grows globally. This review examines the neuropsychological literature specifically addressing the concept of impaired cognitive functioning of insufficient magnitude to warrant a diagnosis of dementia and its meaning with respect to the development of dementia. Although the most obvious finding in the literature is that persons with impaired cognitive functioning have varied outcomes, it is clear that a significant proportion of persons with mild cognitive impairment progress to dementia over a 1- to 2-year interval and approximately 50% progress to dementia by 5 years. The best and most commonly identified predictors of decline to dementia include age and lower baseline performance on neuropsychological measures (e.g., measures of memory). In discussing these findings, issues related to sample definition, sample selection, and methodology are identified and recommendations for future research are provided. PMID- 11262721 TI - Normative data on the Boston Naming Test and two equivalent 30-item short forms. AB - Because of the significance of the Boston Naming Test (BNT) in the differential diagnosis of the dementias, especially Alzheimer's disease, adequate norms from community-dwelling elderly individuals are essential. The present study describes the development of two new empirically derived equivalent short forms (30 items each) of the test. Normative data for the total BNT and the two equivalent 30 item halves based on item difficulty are presented using the performance of 314 community-dwelling individuals aged 65 and over. Age and education norms are presented using an overlapping midpoint interval strategy. PMID- 11262722 TI - Myths of neuropsychology: intelligence, neurometabolism, and cognitive ability. AB - Recently, Dodrill (1999) revised a previously described "Myth of neuropsychology" (1997) to state: "Just as below average performances on neuropsychological tests are found when intelligence is below average, to that same degree above average performances on neuropsychological tests are expected when intellectual abilities are above average." This study addresses the relationship between intellectual and neuropsychological performance in the context of Magnetic Resonance Spectroscopy (MRS) measurements of the neurometabolite N-acetylaspartate (NAA). When subjects were stratified by Full Scale IQ (Average, High Average, Superior) they differed significantly in terms of total neuropsychological performance [F(2,47) = 17.63; p <.001] and the neuronal marker NAA [F(2,47) = 3.25; p <.05]. Regression analysis across groups demonstrated that FSIQ and NAA were independently related to Total z-score [F(1,47) = 29.43; p <.0001] and accounted for over half the variance (r(2) of model =.56). The concurrent relationship of FSIQ and NAA to total neuropsychological performance suggests that the relationship between measures sensitive to intellectual ability and neuropsychological performance is real, and does not reflect arbitrary psychometric or scaling properties of the WAIS-III. PMID- 11262723 TI - Prospective study of the MMPI-2 correction factor after mild head injury. AB - Gass (1991) proposed a correction factor composed of 14 MMPI-2 items that were characteristically endorsed by patients with closed-head injury. Their frequency of occurrence suggested that the items reflected the neurological rather than emotional consequences of head injury. The current study was designed to evaluate the interpretive significance of correction factor items after mild head trauma. Patients were examined immediately upon hospitalization and followed prospectively for at least 3 months. Correction factor items were endorsed more frequently during acute hospitalization than in the MMPI-2 standardization sample. At follow-up, none of the items were endorsed more often by patients with chronic mild head injury than by uninjured controls. These results suggest that the correction factor is sensitive to the acute neurological consequences of mild head trauma, but that these symptoms can typically be expected to resolve. Chronic endorsement of the items in this population is therefore most likely related to psychological factors. PMID- 11262724 TI - The salience of visuospatial and organizational skills in reproducing the Rey Osterreith Complex Figure in subjects with high and low IQs. AB - The salience of visuospatial and organizational skills in copying the Rey Osterreith Complex Figure (ROCF) was examined in 44 adults diagnosed with Attention Deficit Hyperactivity Disorder (ADHD). Subjects were divided into a high average and above IQ group (n = 21) and an average and below IQ group (n = 23). Multiple regressions were conducted for both groups, with the ROCF copy as the dependent variable and WAIS-R Block Design and Stern Fragmentation scores as predictor variables. Results indicated that, for the high IQ group, visuospatial skills were more salient in predicting ROCF copy scores. By contrast, for the low IQ group, organizational skills were more salient for predicting ROCF copy score. Our findings are discussed in relation to the Boston Process Approach. PMID- 11262726 TI - Research strategies in autism: a story with two sides. PMID- 11262727 TI - Radiologic classification of malformations of cortical development. AB - Malformations of cerebral cortical development are common birth defects that can cause delayed development, epilepsy, focal neurologic deficits, and mental retardation. Rational classification of these disorders is essential for proper prognosis, genetic testing and counseling, and investigation of the underlying molecular causes. A rational approach to this classification is a framework based on whether these disorders are the result of abnormal cell proliferation or apoptosis, abnormal migration of immature neurons, or abnormal horizontal and radial orientation in the cortex. Superimposed on this framework are subclassifications that are based on topology of the malformation, associated central nervous system (CNS) or extra-CNS malformations, and results of molecular genetic testing. Characteristics that correlate with and enforce this system of classification can be identified by modern neuroimaging studies. PMID- 11262728 TI - Molecular genetics of human microcephaly. AB - Human microcephaly comprises a heterogeneous group of conditions that are characterized by a failure of normal brain growth. Microcephaly can be caused by many injurious or degenerative conditions, or by developmental malformations in which the growth of the brain is impaired as a result of defects in pattern formation, cell proliferation, cell survival, cell differentiation, or cell growth. These latter forms of congenital microcephaly are frequently inherited, usually as recessive traits, and are associated with mental retardation and sometimes epilepsy. Some of the genes that cause congenital microcephaly are likely to control crucial aspects of neural development, and may also be involved in the evolutionary explosion of cortical size that characterizes primates. There has recently been a rapid advance in the use of genetic mapping techniques to identify genetic loci responsible for microcephaly. Although several loci have been mapped, the condition is clearly genetically and clinically heterogeneous. PMID- 11262729 TI - Cerebral gyral dysplasias: molecular genetics and cell biology. AB - The promise of genetics has been partly realized in our understanding of human brain development as this relates to disorders of gyral formation. Cerebral gyral dysplasias are disorders of brain formation that result in phenotypes with the common feature of abnormal cerebral gyri. This review emphasizes the recent progress made in understanding the human lissencephalies and related disorders. LIS1 heterozygous loss-of-function deletions and point mutations, as well as Doublecortin mutations in males, lead to a very similar phenotype, termed type 1 lissencephaly. Additionally, Doublecortin mutations in females lead to a more variable subcortical band heterotopia. Given the similarities between the lissencephaly phenotypes that result from aberrations in these genes, it is important to review the genetics of these disorders. In order to begin to understand the cell biology of the LIS1 protein and the Doublecortin protein, potentially interacting pathways need to be emphasized. Another human genetic disorder with an interestingly similar phenotype has a mouse correlate that has been well characterized. This surprising finding may lead to further understanding of LIS1 protein and of Doublecortin protein. Furthermore, mouse modeling of the aforementioned human disorders now holds promise for enabling us finally to understand the formation of the most complex organ that nature has produced - the human brain. PMID- 11262730 TI - Molecular genetic aspects of the phakomatoses: tuberous sclerosis complex and neurofibromatosis 1. AB - The phakomatoses are a diverse group of diseases characterized by skin lesions in early childhood followed by the development of tumors in many other organs. Tuberous sclerosis complex and neurofibromatosis 1 are of special interest to the neurologist because of their prominent neuro-oncological and neuro-developmental consequences. The cloning of genes responsible for these two diseases has led to the identification of causative mutations, an understanding of basic cellular pathophysiology and the development of animal models. Current laboratory investigations are focused on bringing clinical relevance to these findings, including the prospects of molecular diagnostics and rational therapeutics. PMID- 11262732 TI - Homeobox gene mutations and brain-stem developmental disorders: learning from knockout mice. AB - Analysis of mice that carry targeted inactivations of Hox, Nkx and Phox2 homeobox genes revealed their involvement in regional patterning of brain-stem territories, in specification of neuronal identity, in establishment of appropriate patterns of connectivity and in control of neurotransmission. The specific abnormalities generated by such mutations may provide clues to the genetic basis and cellular mechanisms that are involved in human brain-stem developmental disorders. PMID- 11262731 TI - Molecular genetics of Rett syndrome and clinical spectrum of MECP2 mutations. AB - Rett syndrome, a neurodevelopmental disorder that is a leading cause of mental retardation in females, is caused by mutations in the X-linked gene encoding methyl-CpG-binding protein 2 (MeCP2). MECP2 mutations have subsequently been identified in patients with a variety of clinical syndromes ranging from mild learning disability in females to severe mental retardation, seizures, ataxia, and sometimes neonatal encephalopathy in males. In classic Rett syndrome, genotype-phenotype correlation studies suggest that X chromosome inactivation patterns have a more prominent effect on clinical severity than the type of mutation. When the full range of phenotypes associated with MECP2 mutations is considered, however, the mutation type strongly affects disease severity. MeCP2 is a transcriptional repressor that binds to methylated CpG dinucleotides throughout the genome, and mutations in Rett syndrome patients are thought to result in at least a partial loss of function. Abnormal gene expression may thus underlie the phenotype. Discovering which genes are misregulated in the absence of functional MeCP2 is crucial for understanding the pathogenesis of this disorder and related syndromes. PMID- 11262733 TI - Future trends in epileptology. PMID- 11262734 TI - The process of epileptogenesis: a pathophysiological approach. AB - Several recent advances have contributed to our understanding of the processes associated with mesial temporal lobe epilepsy in humans and in experimental animal models. Common pathological features between the human condition and the animal models may indicate a fundamental involvement of the given pathology in the process of epileptogenesis. PMID- 11262735 TI - Current trends in electroencephalography. AB - Several recent articles re-emphasize the value of clinical electrophysiology: in localizing epileptogenesis, predicting effectiveness of epilepsy surgery, and disclosing a mechanism of benign Rolandic epilepsy of childhood.A review of the role of EEG in the diagnosis of epilepsy indicated that epileptiform activity will appear in 50% of initial awake recordings of adults with epilepsy and in 85% of subjects undergoing two recordings. This contrasts with the appearance of spikes in only 4 of 1000 normal persons. Several studies focused on the value of electroencephalography in extratemporal epilepsy: 62% of patients with neocortical epilepsy had at least one localizing ictal EEG; occipital and temporal neocortical seizures were localized in a greater proportion than frontal or parietal attacks. Interictal spikes, if unifocal, always arose from the epileptogenic region in a study of their seizure localizing value. Such congruence augured for better seizure control by focal resection in two studies reviewed herein. Studies indicating the value of interictal temporal lobe spikes and scalp-recorded seizures in lateralising a temporal seizure focus are reviewed. One study found EEG to be slightly more reliable for lateralization of temporal epileptogenesis than MRI. In patients with benign Rolandic seizures, enhanced motor evoked potentials (MEPs) were obtained from transcranial magnetic stimulation when this was applied 50-80 msec after electrical stimulation of the thumb whereas this interval inhibited the MEP in normal subjects. This suggests that afferent cutaneous input abnormally and synchronously activates a large population of sensory neurons; such activation is subsequently transmitted to the motor cortex to produce the focal spikes in this condition.Finally, advances in non-invasive technology have redefined and limited the need for invasive monitoring in children with intractable seizure disorders. PMID- 11262736 TI - Epilepsy therapy: issues for women and men. AB - It has been suggested that polycystic ovary syndrome is a common finding in women treated with valproate. However, in a recent study this suggestion could not be confirmed. There is currently no clear evidence that valproate contributes to the development of the polycystic ovary syndrome. Focal epileptic discharges may have an impact on the hypothalamic-pituitary-ovarian or -testicular axis. In the case of successful epilepsy surgery the impact of epilepsy on endocrine functioning may cease. This may lead to a normalization of disturbed menstrual cycles in women, and leads to a post-surgical increase of serum androgens in men. Both findings are supplemented by the results of animal experiments. Children exposed to antiepileptic drugs during pregnancy show a normal psychomotor and cognitive development. However, newly developed as well as traditional antiepileptic drugs increase the risk that a child exposed to these drugs during pregnancy will develop a malformation. PMID- 11262738 TI - Memory and epilepsy: characteristics, course, and influence of drugs and surgery. AB - Memory processing in humans is essential for consciousness, cognitive-behavioral development and individual biography. In epilepsy, declarative memory functions show characteristic patterns of impairment when mesiotemporal and associated neocortical structures are affected by lesions, ongoing epileptic activity, or the undesired effects of conservative or operative treatment. Major issues are thus the etiology, onset and course of memory impairment, as well as the prevention of further memory decline during treatment. New input in the field has resulted from improved imaging techniques, sophisticated experimental study designs, more selective surgical approaches, and new antiepileptic drugs. PMID- 11262737 TI - Therapy of status epilepticus in adults and children. AB - Clinical studies of the treatment of status epilepticus are extremely difficult to carry out, therefore a paucity of new clinical studies have been reported. Much of the progress regarding the therapy of status epilepticus has come from a better understanding of the epidemiology of status epilepticus and its consequences and from laboratory studies of experimental status. Status epilepticus has been used as an experimental tool to study epileptogenesis, but from such studies have come insights that can be applied to the therapy of status epilepticus itself. This review will focus on information from epidemiological, experimental, and clinical studies of status epilepticus, which may contribute to the improved treatment of this life-threatening disorder. PMID- 11262739 TI - Psychiatric issues in epilepsy. AB - In recent years there has been considerable research interest at the interface between epilepsy and psychiatry. Topics of interest include the epidemiology of psychiatric co-morbidity in epilepsy; clinical syndromes at this interface and their classification; the relationship between cognitive dysfunction and psychiatric co-morbidity; biological mechanisms that mediate such co-morbidity, especially with developments in imaging and genetic research; the association between temporal lobe surgery, vagus nerve stimulation, and other non pharmacological treatments, and the development of such co-morbidity; the contribution of anticonvulsant drugs towards the development of psychiatric co morbidity; quality of life and other psychosocial issues; and non-epileptic attack disorder. In this review, papers on these psychiatric issues in epilepsy, with a focus on those published in the past year (October 1999 to October 2000) are critically evaluated, and some important current issues at this interface are considered in detail. PMID- 11262741 TI - Proximal Humerus Fracture. PMID- 11262740 TI - Assessing the severity of seizures and epilepsy: which scales are valid? AB - A review of the literature on methods of seizure or epilepsy severity assessments resulted in tabulation of the seizure rating scales known as US Department of Veterans Affairs (VA), Chalfont-National Hospital, Liverpool, Hague, and others. Each of the scales reviewed has some advantages, but none of them appears to be adequate to assess seizure or epilepsy severity. Most of the scales use similar components of seizures to evaluate severity. However, the disadvantages of each scale outweigh its usefulness. New approaches are needed to assess seizure severity for individuals and for use as an outcome measure after intervention such as surgery or medication changes. PMID- 11262756 TI - Why the US Department of Agriculture should be allowed to insist on inclusion of rodents and birds in the Animal Welfare Act. PMID- 11262757 TI - Novel pyrogen tests based on the human fever reaction. The report and recommendations of ECVAM Workshop 43. European Centre for the Validation of Alternative Methods. European Centre for the Validation of Alternative Methods. PMID- 11262758 TI - The establishment of human research tissue banking in the UK and several western European countries. The report and recommendations of ECVAM Workshop 44. PMID- 11262759 TI - The importance of the prediction model in the validation of alternative tests. AB - An overview is presented of the validation process adopted by the European Centre for the Validation of Alternative Methods, with particular emphasis on the central role of the prediction model (PM). The development of an adequate PM is considered to be just as important as the development of an adequate test system, since the validity of an alternative test can only be established when both components (the test system and the PM) have successfully undergone validation. It is argued, however, that alternative tests and their associated PMs do not necessarily need to undergo validation at the same time, and that retrospective validation may be appropriate when a test system is found to be reliable, but the case for its relevance remains to be demonstrated. For an alternative test to be considered "scientifically valid", it is necessary for three conditions to be fulfilled, referred to here as the criteria for scientific relevance, predictive relevance, and reliability. A minimal set of criteria for the acceptance of any PM is defined, but it should be noted that required levels of predictive ability need to be established on a case-by-case basis, taking into account the inherent variability of the alternative and in vivo test data. Finally, in view of the growing shift in emphasis from the use of stand-alone alternative tests to alternative testing strategies, the importance of making the PM an integral part of the testing strategy is discussed. PMID- 11262760 TI - The red blood cell phototoxicity test (photohaemolysis and haemoglobin oxidation): EU/COLIPA validation programme on phototoxicity (phase II). AB - In the EU/COLIPA validation programme on "Photoirritation in vitro", two core tests and a number of mechanistically based tests were carried out to examine their suitability as regulatory tests for phototoxicity testing. In the meantime, one core test, the 3T3 neutral red uptake phototoxicity test (NRU PT) has been validated and has been accepted by ECVAM and the European Commission. The second core test, the red blood cell phototoxicity test (Photo-RBC test), has passed through a prevalidation process during this programme. This test protocol combines two endpoints, photohaemolysis and met-haemoglobin (met-Hb) formation. These endpoints are determined by measuring changes in the optical density of the haemoglobin spectrum at 525 nm and 630 nm, respectively. In addition, a prediction model was inserted into the Standard Operating Procedure (SOP) with two cut-off values: a photohaemolysis factor (PHF) > or = 3.0 for photohaemolysis, and a deltaOD(max) > or = 0.05 for met-Hb formation. Three laboratories agreed to implement the SOP and to perform the study by testing 30 selected test chemicals (25 phototoxicants and 5 non- phototoxic chemicals). The outcome of the study presents a good overall fit, including acceptable accuracy, sensitivity, and positive predictivity. The specificity and the negative predictivity are comparably low, due to the low number of non-phototoxic substances among the test chemicals. Further analysis of the data showed that the transfer of the SOP from between laboratories could have been more efficient. The results, especially of the lead laboratory, clearly indicate that an experienced laboratory can handle the SOP with high predictivity for phototoxicants and non phototoxic substances. Finally, it was concluded that the combined Photo-RBC test can be considered as a second in vitro test, which can be used advantageously to obtain some mechanistic information, in particular on photodynamic effects on cellular proteins and biomembranes. PMID- 11262761 TI - Toxic effects of chromium acetate hydroxide on cells cultivated in vitro. AB - Many human activities, particularly industrial ones, result in an ever-growing production of toxic waste materials. The dynamics of the toxic effects of chromium acetate hydroxide, which is found in high concentrations in a waste sediment produced in the Czech Republic, were assessed by using a battery of in vitro tests carried out on two cell lines: L-929 (mouse fibroblasts) and Hep 2 (human laryngeal cells). Various markers of cell damage were assessed by phase contrast, video and fluorescence microscopy, fluorometry, and DNA analysis. Chromium acetate hydroxide, over a concentration range of 1-0.02mol/l induced immediate cell death by fixation, whereas, at 0.002mol/l, the treated cells died in a much slower, more discrete manner. All the detected markers of cell damage, whether immediate or slow, clearly demonstrated that the cells died by necrosis. On the other hand, test concentration of 0.001mol/l appeared to constitute a threshold at which no pathological changes of Hep 2 cells were observed over 96 hours. We conclude that chromium acetate hydroxide has a high toxic potential in vitro, which should be considered when studying the toxicity of waste materials containing it. PMID- 11262762 TI - Collagen type I gel cultures of adult rat hepatocytes as a screening induction model for cytochrome P450-dependent enzymes. AB - Albumin secretion, expression of cytochrome P450 dependent mono-oxygenases (CYPs) and their inducibility by well-known inducers were evaluated during 1 week in collagen type I gel sandwich and immobilisation cultures of adult primary rat hepatocytes. Albumin secretion increased during culture time and, following an initial decrease, CYP biotransformation activities remained stable for at least 7 days. Better preservation results were observed in the collagen gel sandwich culture than in the immobilisation model. The inducibility of CYPs by beta naphthoflavone (beta-NF), 3- methylcholanthrene (3-MC), phenobarbital (PB) and dexamethasone (DEX) was studied in both collagen gel hepatocyte cultures. Exposure of the cells to either 5microM 3-MC or 25 microM beta-NF, added to the culture medium, resulted in strong increases of CYP1A1/2 activity in both culture models. Treatment with PB (3.2 mM) resulted in an increase in the CYP2B activity and a higher hydroxylation of testosterone in the 16alpha-position (CYP2B1/2 and CYP2C11), the 7alpha-position (CYP2A1/2), and the 6beta-position (CYP3A1). DEX (10 microM) markedly increased testosterone 6beta- and 7alpha-hydroxylation. Expression and induction experiments of CYP proteins exposed to these molecules confirmed the results of the CYP activity measurements. The patterns of CYP induction in collagen gel cultures of rat hepatocytes were similar to those observed in vivo. Consequently, collagen gel cultures and, more specifically, collagen gel sandwich cultures seem to be suitable as in vitro models for evaluating xenobiotics as potential inducers of CYP-enzymes. PMID- 11262763 TI - Communication of the three Rs within establishments: some problems and solutions. Report of a LASA Alternatives Section meeting. Laboratory Animal Science Association. AB - A meeting was organised by the Laboratory Animal Science Association (LASA) Alternatives Section to provide a forum in which ideas for strategies to raise awareness of the Three Rs could be discussed. One of the main objectives of the meeting was to identify effective strategies which could feasibly be implemented at establishments. In general, there are two main ways in which Three Rs information can be obtained and communicated. Individuals can be assigned to a role with specific responsibilities (for example, a named alternatives expert), or committees can be established, with a remit to find ways to reduce, refine and/or replace animal use (for example, an alternatives committee). The meeting involved invited speakers who are engaged in obtaining and communicating Three Rs information, and this report presents a summary of the advantages and limitations of various approaches that are being undertaken. PMID- 11262764 TI - Gender inequality and severe malnutrition among children in a remote rural area of Bangladesh. AB - Bangladesh typifies many south-eastern countries where female children experience inferior health and uncertain survival, especially after the neonatal period. This paper attempts to study the gender inequality in nutritional status and the effects of various socioeconomic, demographic, and health-programme factors on gender inequality in a remote rural area of Bangladesh. Measurements of mid-upper arm circumference (MUAC) were taken from 2,016 children aged less than 5 years (50.8% male, 49.2% female) in 1994. Children were characterized as severely malnourished if MUAC was < 125 mm. Independent variables included various characteristics of children, households, and mothers. Average MUAC for all children was 130 mm; 33% were severely malnourished. Of the severely-malnourished children, 54.2% were female, and 45.8% were male. The gender gap persisted in the multivariate situation, with female 1.44 times more likely to be severely malnourished. Other variables with a statistically significant relationship included the age of children, acceptance of DPT1, and education of household heads. The persistence of such a gender discrimination now when the country has achieved a lot in terms of child survival is striking. The issue is important and demands appropriate corrective actions. PMID- 11262765 TI - Age- and cause-specific childhood mortality in Lombok, Indonesia, as a factor for determining the appropriateness of introducing Haemophilus influenzae type b and pneumococcal vaccines. AB - Using age and cause-specific childhood mortality in Lombok, Indonesia, as a factor for determining the appropriateness of introducing Haemophilus influenzae type b (Hib) and pneumococcal vaccines, the study describes a cross-sectional, hamlet-level mortality survey in 40 of 305 villages in Lombok Island, Indonesia. Causes of death were assessed with a standardized verbal-autopsy questionnaire. One thousand four hundred ninety-nine births and 141 deaths occurring among children aged less than 2 years were identified, with 43% of deaths occurring during the first 2 months of life. The infant mortality rate was 89 (95% CI: 75, 104) per 1,000 live-births. All mortality rates are reported per 1,000 live births. To examine children whose deaths could potentially have been prevented through vaccination with Hib or pneumococcal vaccine, deaths due to acute respiratory infection (ARI) and central nervous system (CNS) infections among children, aged 2-23 months, were analyzed. ARI and CNS infections caused 58% (mortality rate: 31 per 1,000 live-births; 95% CI: 23, 41) and 17% (mortality rate: 9 per 1,000 live-births; 95% CI: 5, 16), respectively, of all deaths within this age group. Between the ages of 2 and 23 months, 5% of all babies born alive died of ARI, and another 1% died of CNS infections. Our results indicate that current efforts to reduce childhood mortality should focus on reducing ARI and meningitis. These efforts should include evaluating the impact of Hib and pneumococcal vaccines within the routine Expanded Programme on Immunization system. PMID- 11262766 TI - Treatment of childhood diarrhoea in Nigeria: need for adaptation of health policy and programmes to cultural norms. AB - A community survey of treatment regimens for acute diarrhoea in children was carried out in 10 villages in the Ona Ara Local Government Area of Oyo State, Nigeria, using a combination of qualitative (focus-group discussions) and quantitative (weekly surveillance of diarrhoea) methods. Focus-group discussions were conducted with parents of children aged less than 5 years, while a surveillance of diarrhoea among 550 children of same age was carried out during a 6-month period. The findings of the study showed that not all types of diarrhoea were recognized as illnesses, and only those considered to be illnesses were treated. Treatment often involved an adhoc group which comprised adults who were present at the time the illness occurred (including parents, neighbours, relatives, and elders). Certain beliefs and practices, such as associating types of diarrhoea with occupation or ethnic groups, categorizing the severity on perceived causes, and withholding certain foods during episodes of diarrhoea, were common factors in decision-making for seeking treatment. Antimicrobial agents were used in the case of 46.8% of 205 diarrhoeal episodes, and 28.5% were not at all treated. The usual practice of focusing on a target group, such as mothers, during educational interventions may need to be modified in communities where nearly every adult has a role in decision-making in relation to health. The need to adapt health policy and programmes to cultural norms should be addressed to improve the impact of programmes. PMID- 11262767 TI - Prevalence and risk factors of hepatitis B virus, hepatitis C virus, and human immunodeficiency virus infections among drug addicts in Bangladesh. AB - This cross-sectional study investigated the prevalence and risk factors of hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus (HIV) infections among 266 drug users attending a drug-addiction treatment centre in Dhaka, Bangladesh, from November 1996 to April 1997. Of the 266 addicts, 129 were injectable drug users (IDUs), and 137 were non-injectable drug users (non-IDUs). The seroprevalences of hepatitis B virus surface antigen (HBsAg), anti-HBc, anti-HBs, and anti-HCV antibodies among the IDUs were 8 (6.2%), 41 (31.8%), 15 (11.6%), and 32 (24.8%), and among the non-IDUs were 6 (4.4%), 33 (24.1%), 9 (6.6%), and 8 (5.8%) respectively. None of the drug users were positive for anti-HIV antibody. Although the prevalence of HBV infection did not significantly differ between the IDUs and the non-IDUs, the prevalence of HCV infection was significantly higher among the IDUs. Among the IDUs, the prevalence of both HBV and HCV infections was associated with sharing of needles and longer duration of injectable drugs used. The seroprevalence of HBV infection in both IDUs and non-IDUs was significantly higher among those who had a history of extramarital and premarital sex. The prevalence of HCV infection was not associated with sexual promiscuity. There was no association between the seroprevalence of HBV and HCV infections and age. Active preventive programmes focusing on educational campaigns among the youths against substance abuse should be undertaken. PMID- 11262768 TI - Risk factors and gender differentials for death among children hospitalized with diarrhoea in Bangladesh. AB - To identify risk factors for death among children with diarrhoea, a cohort of 496 children, aged less than 5 years, admitted to the intensive care unit of a diarrhoeal disease hospital in Bangladesh, was studied during November 1992-June 1994. Clinical and laboratory records of children who died and of those who recovered in the hospital were compared. Deaths were significantly higher among those who had altered consciousness, hypoglycaemia, septicaemia, paralytic ileus, toxic colitis, necrotizing enterocolitis, haemolytic-uraemic syndrome, invasive or persistent diarrhoea, dehydration, electrolyte imbalances, and malnutrition. Females experienced a 2-fold higher risk of death than males (p = 0.003). Several indices of severe infections were identified more frequently among females than males. Females with severe infections were less frequently brought to the hospital than their male counterparts. The time lapse between onset of symptoms and hospital admission was significantly higher in females than males. This study suggests initiation of programmes to alleviate social disparity between genders for healthcare in poor communities. The study-results may also help physicians identify either prognostic indicators or risk factors for death among children hospitalized with severe illnesses associated with diarrhoea. PMID- 11262769 TI - Bacteriological quality of weaning food and drinking water given to children of market women in Nigeria: implications for control of diarrhoea. AB - Bacteriological quality of weaning food and drinking water given to 2 groups of children aged < or = years was evaluated by estimating bacterial cell count. One group consisted of those taken to market and the other of those left at home in the care of older siblings or house-helps. Bacterial counts (geometric mean) ranged from 5.02 +/- 1.82 to 8.70 +/- 1.0 log10 cfu per g or mL of food, and from 1.15 +/- 1.67 to 6.53 +/- 0.81 log10 cfu per g or 100 mL of water. Analysis of variance showed no significant difference in counts between types of food and between meals (breakfast and lunch). Bacterial contamination increased significantly with storage time, and was, in all circumstances except the water samples, significantly higher in foods given to children left at home. Reheated leftover foods also had significantly higher bacterial load than the freshly cooked food. Coliform count varied significantly with source of drinking water. Poor hygiene standard (inferred from bacterial contamination) was generally observed among mothers weaning < or = 2-year-old children, while they were engaged in trading activities in the market, thus exposing their children to high risk of diarrhoea. Hygiene was significantly poorer in weaning of children left at home in the care of older siblings or house-helps. This implies that, in spite of their trading activities in the market, mothers still take better care of their babies than the older siblings or house-helps who may be inexperienced. These mothers may need education on childcare and food hygiene to suit to their trading activities, for example, during their monthly meetings. There is also a need to establish ORT (oral rehydration therapy) corners in the markets as part of the municipal services. This can be used not only for efficient and quick management of diarrhoea in the market but also for reinforcing hygiene education. PMID- 11262770 TI - Circulation of group A rotavirus subgroups and serotypes in Pune, India, 1990 1997. AB - Group A rotavirus-positive stool specimens, collected from 432 hospitalized patients of all age groups with diarrhoea during 1990-1997 from Pune, India, were characterized for subgroups (SGs) and G serotypes (1, 2, 3, 4, 6, 8, and 10). ELISA for subgrouping was carried out by employing subgroup I and II-specific monoclonal antibodies (MAbs). For serotyping, MAbs against G1 (Ku), G2 (S2), G3 (Yo), and G4 (ST-3) were used. In addition, MAbs against G3 (RV-3), G8 (B37), G6 (bovine U.K.), and G10 (B223) were also employed. Of the 432 specimens, 174 (40.27%) belonged to subgroup I, 187 (43.29%) to subgroup II, 15 (3.47%) to subgroup I and II, and 56 (12.96%) did not react to MAbs specific to subgroup I and subgroup II MAbs. Of the 432 specimens, 111 (25.69%) reacted to one of the MAbs used. Thirty-five of the 111 specimens were serotyped as G1, 34 as G2, and 42 as G3, G4, G6, G8, and G10. Sixty-seven (21%) specimens gave dual reaction mainly to MAbs against G6, G10; G2, and G4, and in several other combinations. Forty-seven specimens (10.88%) showed multireactivities. A large number of specimens (47.92%) did not show any reactivity with MAbs employed in this study, and remained non-serotypeable. Subgroup I was found to be more common in Pune, and most specimens negative for subgroup I and II were non-serotypeable. The results implicate the need for characterization of unusual and non-typeable strains before undertaking any rotaviral vaccine studies in India. PMID- 11262771 TI - Use of mid-upper arm circumference for evaluation of nutritional status of children and for identification of high-risk groups for malnutrition in rural Bangladesh. AB - Measurements of mid-upper arm circumference (MUAC) of 8,881 children were considered cross-sectionally to determine the effects of diarrhoea, breast feeding, and birth-spacing on the nutritional status of children in rural Matlab, Bangladesh. It was observed that age was one of the most significant determinants of child nutrition. The younger children (< 2 years) had significantly higher levels of severe malnutrition than the children aged 2 years or older. Children who had diarrhoea during the last 12 months prior to the study had significantly (p < 0.001) higher severe malnutrition than the children who did not suffer from diarrhoea. Children born with a longer interval after birth of an elder sibling (24+ months) and who were breastfed for a longer duration (2-3 years) were less likely to be severely malnourished than those who were born with a shorter birth interval or who terminated breast-feeding prior to 2 years of age. Education of mothers, housing space, family size, religion, and sex of children had significant effects on the nutritional status of children. Results of the study suggest that MUAC is a potential anthropometric indicator of child nutrition. PMID- 11262773 TI - Study launched to help predict and manage serious cancer-treatment side effects. PMID- 11262772 TI - Is your Website ADA compliant? PMID- 11262774 TI - Intertrochanteric fractures in adults younger than 40 years of age. AB - This study reviewed 66 intertrochanteric fractures in patients younger than 40 years old (average 33.0 years old; range 17-40 years old). In contrast to the usual population with intertrochanteric fractures, the factors male predominance (46/66), less pre-injury comorbidity (9/66), more outdoor high energy trauma (47/66), and more associated injuries (32/66) were evident. The distribution of associated injuries was wide. Some of them were life threatening. According to Boyd's classification, 20 were type I, 24 were type II, 13 were type III, and 9 were type IV. Twenty-nine were stable, and 37 were unstable. Stratified by the mechanism of injury, the difference in distribution between the subgroups was significant (p = 0.027, two-tail Fisher's exact test). Simple falls only caused Boyd type I and II fractures. Boyd type III or IV fractures were found more often after vehicular trauma or falls from a height. All the intertrochanteric fractures healed on average 70.5 days (range 31-213 days) after operation. The fractures resulting from vehicular trauma or fall from a height healed significantly more slowly (p = 0.02, univariant log-rank test). There were 6 intertrochanteric fracture-related complications. The mechanism of injury determines the character of intertrochanteric fractures in young adults. Given tougher bone stock, better healing ability, and less co-morbidity, proper management can lead to healing of all intertrochanteric fractures. The extent of functional recovery was also determined by the associated injuries. PMID- 11262775 TI - Bone mineral density after total joint arthroplasty of lower extremities in rheumatoid arthritis patients. AB - We studied the effects on the axial bone mass of total joint arthroplasty (TJA) for lower extremities in 48 female rheumatoid arthritis (RA) patients by using dual-energy X-ray absorptiometry (DXA). Twenty-nine postmenopausal RA patients treated only with nonsteroidal anti-inflammatory drugs (NSAIDs) served as controls. They were studied for an average duration of 63 months. The reduction in the bone mineral density (BMD) of the lumbar spine (L2-4) was significant in both groups (p < 0.01-0.05), but it was not statistically different between the two groups. The BMD of the femoral neck decreased significantly in both groups (p < 0.01-0.05) after 2 years, but it was not statistically different between the two groups. Our data suggest that TJA slowed the rapid axial bone loss usually associated with advanced RA. PMID- 11262776 TI - Walking ability and activities of daily living after limb salvage operations for malignant bone and soft-tissue tumors of the lower limbs. AB - We evaluated the number of steps, activities of daily life (ADL) score, Enneking score, active range of motion and muscle strength by muscle manual testing for function in lower limbs after reconstructive procedures in surgical treatment of tumors. The 56 patients with 20 malignant bone tumors and 36 malignant soft tissue tumors averaged 7119 +/- 3563 steps per day, or 69.8% of the control group. The average ADL score of patients was 14.0 +/- 4.1 points (70.0%), and the average Enneking score 20.4 +/- 6.0 points (68.0%). The scores of the bone tumor group were lower than those of the soft-tissue tumor group. These scores were not correlated with the range of motion. The number of steps and ADL score were correlated with Enneking score (coefficient 0.52 and 0.84, respectively). The number of steps and the ADL score appear to be useful, as is Enneking score. PMID- 11262777 TI - The Kapandji technique for fixation of distal radius fractures--a biomechanical comparison of primary stability. AB - The goal of this study was to compare Kapandji-K-wiring and established K-wiring techniques of the distal radius fracture for primary stability in a biomechanical model: dorsal K-wiring according to Kapandji using different angles of the K wire, parallel and diagonal alignment of the K-wires. A new testing system which uses a synthetic material enabled us to carry out the cantilever bending test. By application of a lower load, the Kapandji procedure shows a higher reactive torque and stiffness. A higher reaction force of the other techniques, especially of the parallel wiring, are only observable under high-grade bending stress. Application of the Kapandji procedure with K-wires at a smaller angle to the axis of the radius results in the highest primary stability of the procedures investigated in the essential range of initial deformation. PMID- 11262778 TI - The role of computed tomography in cervical spinal injury due to diving. AB - We studied both the clinical features and computed tomography (CT) findings in 25 patients with a cervical spinal injury related to diving. In all patients the X rays, including anteroposterior, lateral and open mouth views, were normal. The clinical features included headache, dizziness, without an alteration in the state of consciousness. In 5 patients, the CT spinal scan revealed cervical spinal injury unrecognised on X-ray. In 4 of them, the mechanism of the trauma was a direct injury to the head when it hit the bottom of the pool; in 1 patient, the mechanism was indirect injury transmitted from the stretched hands to the cervical spine. We conclude that the cervical spinal injury caused by diving should be evaluated selectively by CT spinal scan in patients with direct cervical injury, even if the clinical and roentgenographic results appear negative. PMID- 11262780 TI - Results of open repair of the rotator cuff--a long-term review of 79 shoulders. AB - In 72 patients with 79 tears of the rotator cuff that had been completely repaired by open surgery, the outcome was evaluated on the basis of history, Constant functional score and radiography. At a mean follow-up period of 6.75 years, the Constant score was 71.5 points on average, showing a high correlation with the patients' subjective satisfaction, but the score was not a reliable indicator of recurrence. The larger the cuff tear, the poorer the result was. However, deterioration was not related to the duration of follow-up or the patient's age. The presence of acromioclavicular arthrosis also had no influence on the overall result. A comparison of follow-up radiographs with those obtained immediately after surgery revealed (despite postoperative flat subacromial resection) evidence of heterotopic bone formations or ossifications in nearly half of the patients. However, except for individual cases, this had no significant influence on the clinical result or the Constant score. PMID- 11262779 TI - Correction of kyphotic deformity before and after transection of the anterior longitudinal ligament--a cadaver study. AB - With a custom-made measuring unit, two separate experiments, involving six and five cadaveric torsos with intact rib cages and sternums, respectively, were carried out to determine the effect of the transection of the anterior longitudinal ligament with and without osteodiscectomy and its influence on the thoracic kyphosis. The open or thoracoscopically assisted anterior release, as part of the operative treatment of scoliosis or kyphosis, usually consists of a transection of the anterior longitudinal ligament (ALL) and an additional discectomy. A complete osteodiscectomy, however, is not always possible with a minimally invasive approach. As part of our biomechanical research, we attempted to quantify the amount of correction achievable with a defined force prior to and following the isolated transection of the anterior longitudinal ligament. The aim of the study was to clarify whether or not an isolated transection of the anterior longitudinal ligament is sufficient to obtain an adequate anterior release of the spine. In the surgical treatment of kyphotic deformities, anterior release of the spine is performed in the form of a transection of the ALL and discectomy. Recently, video-assisted thoracic surgery has become increasingly popular in spine surgery. As part of this change in surgical technique, the question has arisen as to what extent an isolated transection of the ALL provides an adequate release of the thoracic spine. Eleven human spines were retrieved from fresh cadavers, dissected, and attached to a specially constructed apparatus. The spine was attached to the construct at the twelfth vertebral body. C6 and C7 were fixed in synthetic resin. We installed the instruments in such a manner as to reproducibly apply a torsional moment of 10 Nm to the spine. Motion was only permitted in the sagittal plane. Segmental transections of the ALL were carried out from T3 to T7. For comparison, the sagittal Cobb angle was also documented following an anterior release combined with an osteodiscectomy. With the isolated transection of the ALL, an average correction of the sagittal Cobb angle of 4 degrees in each functional spinal motion segment was recorded. In comparison, the additional osteodiscectomy led to a further average increase of only 2 degrees per level. The measurements performed on human cadavers showed that the isolated transection of the ALL leads to a sufficient anterior release of the thoracic spine, allowing a correction of the kyphotic deformity. The release with a concomitant osteodiscectomy represents a more time-consuming and more invasive procedure resulting in only a slightly greater amelioration of the sagittal Cobb angle, while being associated with a greater patient morbidity. PMID- 11262781 TI - Elbow arthropathy in hemophilia. AB - Twenty-one affected elbows in 14 known hemophilic patients undergoing treatment at the Hemophilia Clinic of our institution between 1994 and 1997 were evaluated using the clinical evaluation score of the Orthopaedic Advisory Committee of the World Federation of Hemophilia and radiological examination using the Pettersson score. The mean age of patients was 16.2 years (range 10-25 years), and all patients except 1 had severe hemophilia with factor VIII levels less than 1% of normal. The mean duration of disease at the time of presentation was 12.8 years. The mean clinical score was 6, while the mean radiological score was 4.9. There was a positive correlation between the clinical and radiological scores (P < 0.0001). However, the clinical and radiological scores did not correlate with either duration of disease or joint bleeding score. Two new radiological signs of obliteration of the capitello-lateral epicondyle groove with rounding off of the lateral aspect of the distal humerus and lipping of the medial trochlear surface were also recognized in hemophilic arthropathy of the elbow. PMID- 11262782 TI - Evaluation of hydroxyapatite implants in vertebral bodies and extremities by contrast-enhanced magnetic resonance imaging. AB - This pilot study evaluated hydroxyapatite (HA) implants (Endobon) into bone with magnetic resonance imaging (MRI). Nineteen patients (median age 57 years; range 18-67 years) have been evaluated. Eight received granulated HA into vertebral bodies after trauma, while 11 received HA blocks into extremity bones after trauma (n = 8) or fibrous dysplasia (n = 2). In a 1.5-T MR scanner, transversal T1-weighted, flash, 2-dimensional images were obtained. Signal intensities were measured in regions of interest (ROI) from the centre and periphery of HA, trabecular bone, fat and air before and after gadolinium (Gd) contrast enhancement, and the signal-to-noise ratio (SNR) was calculated. After Gd application the SNR in HA increased significantly, peripherally (11.7 without vs 18.2 with Gd, P = 0.003) more than in the centre (7.9 without vs 11.9 with Gd, P = 0.007). The SNR of the HA was higher in patients with block implants compared with granulated HA (P > 0.008). After Gd application, granulated HA enhances significantly less than HA blocks. During insertion of HA, the granulated form is compressed more and is therefore more compact than the HA block. This might hinder integration into the bone structure and the blood supply. SNR of the HA margin was higher than centrally, which might be due to granulomatous tissue after trauma and/or operation or beginning marginal integration. PMID- 11262783 TI - Erythropoietin for autologous blood donation in total hip arthroplasty patients. AB - Forty patients who were scheduled for a total hip arthroplasty were enrolled in a prospective study and were randomly divided into two groups. Group 1 received recombinant human erythropoietin (300 U/kg twice a week), and group 2 received placebo. The medication was started 2 weeks before the operation, and only one dose of medication was given after the operation. Autologous blood was administered at the same time as the medication until the hemoglobin level sank to 10 g/dl. Forty-eight and 49 units of autologous blood were collected in group 1 and group 2, respectively. Intraoperative homologous blood was transfused only to patients in group 2. Seven and 13 units of allogenic blood were transfused into group 1 and group 2 patients during the postoperative period, respectively. There were no any significant differences between the groups in terms of early postoperative hemoglobin level and amount of autologous blood collected. However, the increase of the reticulocyte count in patients who received erythropoietin was significantly higher than in the group 2 patients. The study showed that short-term and low-dose erythropoietin usage strongly stimulates the bone marrow. Erythropoietin administration and preoperative autologous blood donation diminished the total units of allogenic blood required during the intraoperative or postoperative period. Autologous blood administration without concurrent erythropoietin did not stimulate the bone marrow adequately. PMID- 11262784 TI - Tailor's bunion: results of a scarf osteotomy for the correction of an increased intermetatarsal IV/V angle. A report on ten cases with a 1-year follow-up. AB - INTRODUCTION: The aim of this study was to analyze clinical and radiological results of scarf osteotomy for the correction of an increased intermetatarsal IV/V angle in patients with symptomatic tailors bunion. PATIENTS AND METHODS: Between 1997 and 1998, we performed a scarf osteotomy for the correction of an increased intermetatarsal IV/V angle (IMA) in ten cases. The indication was a painful prominence of the fifth metatatarsal with an increased IMA. Fixation of the osteotomy was performed with two 1.7-mm titanium miniscrews. Mobilization was allowed with full weight-bearing, with a forefoot relief orthosis. Clinical results were evaluated with the forefoot scoring system (ffss). The determination of the IMA was performed with weight-bearing dorsoplantar radiographs. RESULTS: All osteotomies healed within the first 6 postoperative weeks. Removal of the screws was not necessary in any case. The mean preoperative ffss was 29.5 points. At the last follow-up, the mean value of the ffss was 73 points and no patient presented a painful prominence above the fifth metatarsal head. The mean IMA was reduced significantly from 10.3 to 6.8 degrees. DISCUSSION: The scarf osteotomy is an adequate surgical procedure for the correction of an increased IMA in patients with symptomatic tailor's bunion. PMID- 11262785 TI - Effect of granulocyte-macrophage colony-stimulating factor on treatment of acute osteomyelitis. An experimental investigation in rats. AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine that affects the various developmental steps of hematopoietic cells and enhances the phagocytic activity of these cells. The effect of GM-CSF on acute osteomyelitis, developed in rats, was investigated. For this purpose, osteomyelitis was firstly developed through the direct inoculation of Staphylococcus aureus into rat tibial metaphysis. Twenty-four rats in which diagnosis of osteomyelitis was histopathologically established were divided into two groups. Antibiotic only was given to the first group, and antibiotic as well as GM-CSF to the second group. Rats were followed up for 3 months with plain radiographs and scintigraphic methods using 67Ga-citrate. Material obtained from the rats that had been killed at the end of the 3rd month were histopathologically investigated. One rat in the first group died. In another rat, chronic osteomyelitis developed and fracture was observed. In 12 rats of the second group, physical examination, plain radiographs, and histopathologic findings were normal. In scintigraphic studies with 67Ga-citrate, when the pre- and posttreatment value of the same groups were evaluated by the Mann-Whitney U-test, the mean values at 48 h after treatment were found to be significant (P < 0.05), indicating a decrease in the 2nd group (experimental group). In conclusion, the antibiotics were effective in the elimination of infection only together with neutrophils. In this manner, infections may be eliminated by strengthening the host's defense mechanism as well as by administering antibiotics. We believe that an adequate number of long term studies will shed light on this issue. Besides we consider that this factor will be more important in the study of chronic osteomyelitis. PMID- 11262786 TI - Cervical fracture of the anterior and posterior elements without evidence of neurological deficit. A report of three cases. AB - We present three cases of cervical spinal fracture, involving two columns without an obvious neurological deficit. Usually if two of three columns are fractured, the injury is considered unstable structurally and clinically. Fortunately our cases did not involve sensory or motor impairment because of an enlargement of the spinal canal. PMID- 11262787 TI - Total knee arthroplasty in a patient with chronic occlusion of the superficial femoral artery. AB - In a 67-year-old patient with severe valgus gonarthrosis and chronic occlusion of the superficial femoral artery on the same side, total knee replacement was performed without preceding angioplasty because the collateral circulation was intact. No tourniquet was used. To leave the peripatellar arterial ring intact on one side in this case of lateral patellar maltracking, a lateral approach was used. In this approach, a lateral release forms part of the approach itself. To achieve gentle eversion of the patella, the tibial tubercle was osteotomized. One year postoperatively, the patient was satisfied with the outcome and showed no clinical signs of any vascular deterioration. It is concluded that total knee replacement may be possible in the presence of chronic occlusion of the superficial femoral artery provided that the collateral circulation is intact. PMID- 11262788 TI - Human embryos in vitro: pioneer illustrations of oocyte maturation, fertilization, cleavage and blastulation. PMID- 11262789 TI - Oocytes. PMID- 11262790 TI - Fertilization. PMID- 11262791 TI - The embryo. PMID- 11262792 TI - Embryo and oocyte cryopreservation. PMID- 11262793 TI - The blastocyst. PMID- 11262794 TI - Intracytoplasmic sperm injection (ICSI). PMID- 11262795 TI - Assisted hatching. PMID- 11262796 TI - Preimplantation genetic diagnosis: polar body and embryo biopsy. PMID- 11262797 TI - [Donor site digital subtraction angiography before mandible reconstruction with free fibula transplantation]. AB - The use of fibula free flap has become established as a reliable and popular method for reconstruction of segmental mandibular defects. Vascular anomalies of the donor site may compromise the blood supply of the flap, may jeopardize perfusion of the foot or cause technical difficulties during the procedure. Clinical examination of the circulation in the lower extremities may have relatively normal results. Preoperative vascular imaging detects aberrant cases and thus may contraindicate the use of fibula free flap or may alter the surgical plan. Authors performed 64 preoperative lower-limb digital subtraction angiographies (DSA) in a series of 39 consecutive patients clinically judged to be satisfactory candidates for free fibula transfer. DSA detected vascular anomalies in 10 extremities (15.6%). Regarding the high potential for significant donor site morbidity authors consider vascular imaging essential part of preoperative evaluation. PMID- 11262798 TI - [Comparative epidemiologic examination of caries in adolescents]. AB - The aim of the study was to assess the caries prevalence and oral hygiene in groups of adolescents living in two Hungarian towns and to establish correlations with socio-economical factors. Examinations of caries and oral hygiene were performed in 586 14-16-year-old children. The percentage of caries free children was 5.1%, the DMFT and DMFS values were 6.97 +/- 4.67 and 9.95 +/- 7.94 respectively. The VPI index was 41.6 +/- 32.7% (mean +/- S.D.), and showed positive correlation with the caries prevalence. The DMFT and DMFS values, as well as VPI decreased as the educational level of the father had increased. The number of siblings had a worsening effect on DMFT, DMFS and VPI values. PMID- 11262799 TI - [Lethal complication of an odontogenic infection developing after tooth extraction in a patient with untreated diabetes. Case report]. AB - The extraction of a tooth led to the acute exacerbation of existing chronic osteomyelitis then phlegmon, and sepsis as a result. Persisting coma developed after three reanimations of the patient, on account of septic shock. Finally, in more than two weeks following the extraction the patient died due to multiorgan failure. This case description wishes to call attention to the risks of complications of diabetes, to summarize the relevant documents of diagnosis and treatment of osteomyelitis and phlegmon and the data of mortality. Hopefully we offer some useful advice to the general dentist about patients with immunosuppressive diseases. PMID- 11262800 TI - [Perspectives of dentistry in the 21st century]. AB - The topics reviewed are based on those published in a recent USA journal, including the following: demographic trends of oral diseases, changes in the trends of oral diseases, minimally invasive dentistry, the development of bond systems, endodontics in the 21st century, trends in surgical and non-surgical periodontal treatments, esthetic ceramics in the 21st century, total dentures: present trends. The author is attaching comments to each of the topics on the present situation in Hungary, hoping to challenge a debate in the following numbers of Fogorvosi Szemle. PMID- 11262801 TI - [Reproducibility of the laser fluorescence method for the diagnosis of occlusal caries. Clinical study]. AB - The aim of the study was to assess the reproducibility of recently developed laser fluorescence method for occlusal caries detection in vivo. 71 non-cavitated premolar and molar teeth were examined repeatedly by three dentists using DIAGNOdent (KaVo) instrument. The presence of caries has been established on the classification of Lussi, created on the basis of "in vitro" investigations. The reproducibility of the measurements were expressed in rank correlation of Spearman. The intra- and interexaminer reliability was also determined by Kappa statistics. The Spearman correlation coefficiencies were fairly high (0.738 0.949) The intraobserver and interobserver coincidencies reflected by Kappa values (0.60-0.88 and 0.44-0.77, respectively) were also convincing. PMID- 11262802 TI - [Evaluation of clinical data and immuno-modulating treatment of patients with oral lichen planus]. AB - The number of patients with oral lichen planus increases all over the world. The treatment of this disease is not yet definitive. Although the etiology is not clear until now increasing number of data show relations between the environment and immune-responsiveness. In this study the data (medical history and laboratory test results) of 33 lichen oris patients were processed. The prevalence of diabetes mellitus was much higher in lichen patients, than in persons without lichen oris. Likewise, higher transaminase serum levels in lichen patients were found, and there were two HCV positive cases among them. In contrast to the literary data instead of immune suppressive treatment the immune modulant treatment has been applied: 78% of beta levamisole treated and 90% of interferon alpha treated patients were cured. Only lichen patients without symptoms of general diseases could be stabilized. PMID- 11262803 TI - [Urinary incontinence in the elderly: what management for which patient?]. AB - THERAPEUTIC OPTIONS: Appropriate management of urinary incontinence in the elderly basically depends on the patient's medical status and degree of dependence and the type of incontinence. For outpatients, secondary effects of drugs limit their use, in favor of behavioral intervention and pelvic floor training, with good success. Recently proposed surgical approaches offer a promising alternative for the aged population. TEAM MANAGEMENT: Assessment and management of institutional urinary incontinence should be systematically elaborated by the medical care team. Potentially reversible conditions and precipitating factors may be causing or contributing to the incontinence. Their correction and bladder training are the principal items of the management scheme. PMID- 11262804 TI - [Hypercalcemia in the elderly]. AB - A COMMON FINDING: Hypercalcemia is not rare among elderly patients. Hyperparathyroidism and neoplasia are the most frequent causes of hypercalcemia in old patients. Symptoms due to hypercalcemia are usually non specific in old subjects, leading to consider easily this diagnosis and to measure plasma calcium level. DIAGNOSIS: Biological diagnosis of hypercalcemia is not always obvious in old patients because of frequently decreased plasma albumin levels leading to lower plasma total calcium level. Thus, it is always necessary to calculate plasma total calcium level corrected by albumin in order not to underestimate hypercalcemia in elderly subjects. PROGNOSIS: The short-term risk of hypercalcemia is acute hypercalcemia, which may be lifethreatening. The long-term risk of hypercalcemia is renal failure. TREATMENT: When hypercalcemia is due to primary hyperparthyroidism, the treatment of choice is surgery. However, for old patients with high surgical risk surgery with local anesthesia or ultrasonically guided percutaneous ethanol injection into parathyroid adenoma can be proposed. PMID- 11262805 TI - [Osteoporosis in the elderly. Practical prevention and treatment]. AB - A MAJOR PUBLIC HEALTH ISSUE: Osteoporosis in the elderly limits independence and quality of life. Preventive and curative treatment should be adapted to the patient's age. THERAPEUTIC OBJECTIVES: The goal of preventive like curative treatment is to reduce the risk of fracture. Risk can best be assessed from personal history of fracture or with bone densitometry. THERAPEUTIC METHODS: Non drug methods are based on physical activity, diet, reduction of alcohol intake or smoking, and limiting the risk of falls. Drugs used include calcium, vitamin D, hormone replacement therapy and bisphosphonates. THERAPEUTIC STRATEGY: Prevention programs should focus first on non-drug methods, adapted to the patient's age. For drug regimens, hormone replacement therapy is rarely used after 70 years of age while calcium and vitamin D are widely used. Drugs inhibiting bone resorption, e.g. bisphosphonates, are added on for curative treatment. After the age of 80 years, the calcium vitamin D combination alone is useful. FOLLOW-UP: A simple surveillance scheme should include a check-up of renal function every year in patients taking bisphosphonates. It is most difficult to assess treatment efficacy on an individual basis. PMID- 11262806 TI - [Rapid regression of prolonged pagophagia after treatment of iron deficiency]. AB - BACKGROUND: Pica is an eating disorder characterized by the irrepressible and elective craving for food and non-food substances. Pagophagia is a particular form characterized by an ingestion of ice, often associated with iron deficiency. We report a case of intense and prolonged pagophagia. We also analyse literature about the nature of the link between pica and iron deficiency. CASE REPORT: A forty-two year old woman was admitted for severe anaemia secondary to iron deficiency possibly related to chronic gynaecological bleeding. A diet investigation revealed a pagophagia of about eighty ice cubes per day developing over five years. Iron supplement corrected the anaemia and the pagophagia disappeared in less than fifteen days without recurrence in the following year. CONCLUSION: The disappearance of pagophagia after the correction of iron deficiency supports the theory that pagophagia could well be a symptom of iron deficiency. PMID- 11262807 TI - [Antivitamins K]. PMID- 11262808 TI - [Antivitamins K]. PMID- 11262809 TI - [Methicillin-resistant Staphylococcus colonization. Impact on morbidity, mortality, and glycopeptide use]. PMID- 11262810 TI - [Ribavirin in combination with interferon alpha-2b in the treatment of chronic hepatitis C]. PMID- 11262811 TI - [Depression in the elderly. Clinical aspects]. AB - DIFFICULT DIAGNOSIS: Depression in the elderly can take on many often misleading aspects. Sadness may be considered legitimate or "normal" for an elderly person. Depression may masquerade as an organic disorder where somatic complaints, pain and anxiety predominate. All these different clinical forms may mislead the clinician. THE MASK OF HYPOCHONDRIA: A tendency to hypochondria, found in more than one-half of all depressed elderly subjects, may be reinforced by bouts of complementary examinations. The patient is convinced of having an unrecognized organic disease. The mask of hypochondria must be considered with special care because it is a major risk factor for attempted and successful suicide. THE MASK OF DELUSIONS: Elderly patients often develop a state of melancolia-like depression with delusions. Delusions may be congruent with the predominant depressed mood, for example a guilt feeling for an act never committed, or inversely, non-congruent with the thymic state (persecution, negation delusin), for example Cotard syndrome where the patient is persuaded that his/her organs are malfunctioning or have disappeared. Despite these impressive mood disorders that often incite prescription of a neuroleptic, these elderly patients respond favorably to antidepressor treatment. PMID- 11262813 TI - [Depression in the elderly. Medical treatment]. AB - THE ADVENT OF DRUG THERAPY: In 1957, the advent of two compounds caught the attention of clinicians and psychopharmacologist: G 22355 or Tofranil, the leading drug in the tricyclic family, and iproniazid or Marsilid, a non-sedative monoamine oxidase inhibitor. WHAT IS AN ANTIDEPRESSOR DRUG?: Antidepressor drugs are not effective in all domains simultaneously: it takes 7 to 10 days before an effective equilibrium of mood is reached. Adverse effects often occur earlier, but generally fade out or are well tolerated when the antidepressor effect becomes patent. CASE-BY-CASE PRESCRIPTION: Twenty-four different antidepressor drugs are available in France, classed among imipramines, non-imipramines and non IMAO, serotonine and noradrenaline reuptake inhibitors, and specific serotonin reuptake inhibitors. Each class has its own contraindications, adverse effects and precautions for use. Normothymic drugs and mood' regulators are a separate category that constitute considerable progress for the treatment of relapse and recurrent depression. PMID- 11262812 TI - [Depression in the elderly. Principles of treatment]. AB - DRUG THERAPY: Drug therapy must be reassessed regularly to avoid overdosing or poor observance. Use of multiple drugs must be carefully monitored. PSYCHOTHERAPY: Listening to the patient and establishing an understanding friendly relationship are essential to successful psychotherapy in the depressed elderly. SUPPORT: Repulsion is common because old age evokes death. The reactions of family and friends may take on many forms, sometimes leading to counter transfer. EIGHT SIMPLE RULES FOR PRESCRIPTION: Ask key questions. Treat what is essential. Prescribe few drugs starting at low dose. Check observance. Favor good diet and hygiene. Help the patient accept the aging process. Avoid pseudo placebos. Use appropriate dose timing. PMID- 11262814 TI - [Depression in the elderly. Rigorous evaluation of drug treatment]. AB - CONSENSUS: There is a general consensus on two points: it takes about 2 months for symptoms to resolve; treatment for 4 to 6 months thereafter reduces the risk of relapse. The same dose should be used for both periods. Likewise for prophylaxis against recurrent depression (long-term treatment for several years). QUALITY CARE: There are three major risks in elderly patients with depression: relapse, chronic depression, suicide. Improved quality care for depressive patients is a major public health issue. It is essential to recognize affected patients who require regular care in order to reduce the risk of non-observance (30-70% of the cases). PRESCRIPTION RULES: Differentiate minor depression from major depression. Wait 7 days before evaluating drug efficacy. Be aware of the risk of desinhibition. See the patient and his/her relations often. Take steps to prevent suicide using appropriate drugs. Favor psychological counseling. Use single-drug regimens preferentially. Prescribe for a sufficiently long period (life?). PMID- 11262815 TI - [Obliterating thromboangiitis: contribution of magnetic resonance angiography]. PMID- 11262817 TI - [Computerized tomography assessment of femoral and tibial rotation in total knee prosthesis]. AB - PURPOSE: CT assessment of the axial deviation of the femoral and tibial prosthetic components in total knee arthroplasty. MATERIAL AND METHODS: January to July 1999, seventeen patients, 10 males and 7 females, mean age 66 years (standard deviation +/- 4) were examined after total knee arthroplasty. Exclusion criteria were prosthesis loosening and severe (equal or superior to 7 degrees) varus or valgus deviation. All patients were examined with knee radiography in the standing position completed by axial projections of patella and by CT scanning. We used a modification of Berger technique and carried out comparative CT scans extended lower limbs and acquisitions perpendicular to the mechanical axis of the knee, from the femoral supracondylar region down to the plane crossing the distal end of the tibial prosthetic component. Reference lines were then drawn electronically on given scanning planes to reckon the axial deviation of the femoral and tibial prosthetic components. RESULTS: Six patients, one female and 5 males, with normal rotational values of femoral and tibial prosthetic components presented no clinical symptoms. Eight patients, 4 females and 4 males, with abnormal values presented the following clinical symptoms: medial impingement, (incomplete) dislocation patella, and lateral instability. One female patient with a normal rotational value of femoral prosthetic component and an altered value of tibial prosthetic component presented medial impingement. Finally two patients, one female and one male, were absolutely asymptomatic although the rotational values of the two prosthetic components were beyond the normal range. CONCLUSIONS: Total knee arthroplasty is presently a standard treatment for many conditions involving this joint. There are several possible postoperative complications, namely fractures, dislocations (a)septic loosening and femoropatellar instability. The latter condition is the most frequent complication among implant failures and is caused by bad orientation of the femoral and tibial components on frontal and axial planes. We measured the orientation of the prosthetic components introducing a CT procedure which modifies the uniarticular with four scans introduced by Berger. The new method uses Berger's parameters and the CT study of both joints by means of Helical CT. With a single examination lasting less than 4 minutes and with the patient in a more comfortable position, we can obtain: 1) comparative and simultaneous assessment of the contralateral joint; 2) several scans to better define Berger's parameters and to accomplish measurement of the rotational deviation with higher precision and markedly decreasing the error margin. The analysis of the results confirms the international literature findings and especially the fundamental importance in positioning both prosthetic components within normal values, as emphasized by the relationship between the clinical symptoms and the rotational degree of the femoral and tibial prosthetic components. PMID- 11262816 TI - [Ankle impingement syndrome]. AB - PURPOSE: The ankle impingement syndrome depends on many factors (fiber or bone production changes) manifested with pain and limited range of movement of the tibiotarsal joint. We tried to classify the various causes and sites of impingement syndromes with MRI. MATERIALS AND METHODS: A cohort of 42 selected patients underwent a 2-year orthopedic follow-up. All patients were examined with MRI using both a low field permanent dedicated magnet at 0.2 Tesla (Artoscan, Esaote Biomedica, Genoa, Italy) and a high field General Electric unit at 1.0 T; sequences and views were chosen according to the condition studied. Gd-DTPA was administered in 26/42 patients. All patients underwent arthroscopy. RESULTS: Twenty-one patients had positive symptoms in the anterolateral region of the ankle and 14 of them had lateral changes. In 13 patients we found fibrous tissue (meniscoid lesion) or hypertrophy of the synovial tissue. An intra-articular body was observed in one patient. An osteophyte was found in 5 patients at the level of the anterior margin of the tibia, with the presence of reactive synovial tissue. Three of 18 patients with posterior pain had a traumatic injury of the posterior exterior tubercle of the astragalus, 7 had a fracture of the os trigonum and 2 had small sclerotic foci formations in calceneal site; chondropathy with sclerosis of subchondral bone was diagnosed in 2 patients. Posterior bone impingement was observed in the remaining 4 patients. Two patients had synovial impingement in posterolateral site. A posterior plica synovialis was seen in 1 patient. Fibrotic-scar tissue was observed in one case, in the subtalus region (impingement synovialis subtalaris). The administration of intra-articular Gd-DTPA provided better definition of the fibrous tissue and the intra-articular free bodies. Modest vascular enhancement of the tissues was seen in 9 of 8 patients receiving the contrast agent. In the other two cases, where signal tissue was low, no signal enhancement was observed after the contrast agent administration. DISCUSSION: Based on integrated MR, clinical and arthroscopic findings we classified tibiotarsal joint impingement syndromes into three types, namely: 1) bone impingement; 2) fibrous impingement where both site and grade are considered; 3) impingement synovialis subtalaris. CONCLUSIONS: MRI appears to be a fundamental diagnostic imaging tool in depicting and detailing the various patterns and sites of the impingement syndrome of the tibiotarsal joint thus allowing an objective classification. PMID- 11262819 TI - [Anatomo-radiologic correlations in spontaneous hematoma of the rectus abdominis muscles]. AB - PURPOSE: Rectus sheath hematomas are a frequent but sometimes misdiagnosed disease in patients under anti-coagulative drugs, hemodialysis, or simply in the elderly. The most frequent localization is in the lower part of the abdomen: the explanation lies in the anatomy of the abdominal wall, especially in the arcuate line of the rectus sheath. Aim of this work is to explain the reason of the almost constant location correlating the anatomy with the CT features. ANATOMIC CONSIDERATIONS: The rectus abdominis muscle lies between the aponeuroses of the transverse and oblique muscles which form the so called rectus sheath. This arrangement is found from the costal arch to a level approximately between the umbilicus and the pubic symphisis, where the rear layer of the rectus sheath ends with a curved edge, called the arcuate or semicircular line of Douglas. Beneath this line the aponeuroses of the three muscles pass in front of the rectus which is separated from the peritoneum only by the fascia trasversalis, a thin connective layer between the rectus and the preperitoneal fat. In this lower aspect of the muscle the perforating branches of the inferior epigastric artery running in the preperitoneal fat may rupture causing a large hematoma widely spreading in this loose space. MATERIAL AND METHODS: 11 cases of rectus sheath hematoma diagnosed over 5 years were reviewed. They were referred to US because of a rapidly growing palpable mass or painful swelling of the abdominal wall with acute anemia. Sonography was performed in 11 patients and CT in 7. RESULTS: 10 hematomas were located in the lower third of the rectus muscle below the arcuate line in the pelvis, 1 was in the upper third of the muscle: the vast majority of pelvic hematomas is easily accounted for by the peculiar anatomy of the region. DISCUSSION: The diagnosis of hematoma of the rectus abdominis, sometimes misleading, should be included as a differential in all the patients who present with acute abdominal pain and blood loss. The anatomy of abdominal wall correlates well with CT findings and explains the reason why most hematomas are found in the lower third of the muscle. CONCLUSIONS: The diagnosis, whether clinical or based on imaging findings, needs accurate pathoanatomic knowledge of the anterior abdominal wall. Once the diagnosis has been confirmed (by US or CT) patients should be treated conservatively as those that are operated are at risk of developing complications, mainly hemorrhagic. PMID- 11262818 TI - [Ultrasonography features of the diaphragmatic crura: normal anatomy and its variants]. AB - PURPOSE: To report the various US patterns of the diaphragmatic crura and the changes occurring during the different phases of respirations. The diaphragm has two US patterns: the central membranous part appears highly reflective while the posterior, upper and lateral muscular portions are hypoechoic and thick. The crura can sometimes appear quite bulky, which appearance is easy to misinterpret. MATERIAL AND METHODS: We carried out a three-stage work: first we reviewed the US examinations of 23 subjects with a nodular appearance of the posteromedial bundles and studied the changes in thickness during respiration. Second we studied the diaphragmatic crura in 30 subjects aged 18-71 years, 15 men and 15 women. We used a commercially available unit with sector and convex 3.5 MHz probes at baseline and during breath hold and acquired multiple parasagittal and transverse scans. The crura thickness was measured in all patients. Last, we studied the diaphragmatic regions of 10 patients with right pleural effusion and of 8 patients with associated ascites and pleural effusion using 2.0-5.0 MHz convex phased-array transducers. RESULTS: We found focal thickening of the crura in 11 of 23 patients with US findings of diaphragmatic nodules, but only in deep inspiration. The thickening was 1.5-2.2 cm long and maximum thickness was 10 mm. In the other 12 subjects we found 9 small lobules in the right and 3 in the left crus. In the anatomic study, we observed a 3-band appearance of the diaphragmatic crura, probably referable to muscle bundles, in 30 subjects on sagittal images, in 12 on coronal images and in 28 on anterior transverse images. The diaphragmatic crura were identified in 26 subjects only. The left posterior crus was identified in 29 subjects on left coronal images and in 15 on anterior transverse images; it was demonstrated on anterior sagittal images in close proximity to the aorta in only 4 subjects. Right crus thickness, measured on sagittal scans, ranged 3-10 mm in deep inspiration and 1-4 mm in expiration while the left crus was 3-6 mm in inspiration and 1-2 mm in expiration. The length of the right crus, studied in the preaortic portion, ranged from 7 cm in deep inspiration to 9.7 cm in expiration while the left one was 6.5 to 8.8 cm. The right lateral diaphragmatic bundles were seen in 28 subjects only on repeated subcostal oblique scans and the the left ones in 11 subjects only. Finally the thin anterior bundles were shown on parasagittal images in 13 cases in the right side and in 2 in the left. A 2-band appearance of the diaphragm was seen in 10 patients with pleural effusion and in 8 patients with associated ascites. A single band was found only in the tendinous portion of the diaphragm. DISCUSSION AND CONCLUSIONS: US is presently considered the imaging method of choice in the assessment of changes in thickness and length of the diaphragmatic crura. These structures have different US patterns and can sometimes appear quite bulky and thus be easily mistaken for other anatomic or abnormal structures; orthogonal scans may be required for the differential diagnosis. PMID- 11262820 TI - [Breast neoplasms detected only with ultrasonography: incidental finding or clinical evidence?]. AB - INTRODUCTION: Mammography is the only technique of proven efficacy in the early diagnosis of breast cancer, even though its sensitivity is much lower in breasts that are dense or with a high parenchymal-stromal component. In the past malignant breast nodules detected at US in patients with negative mammographic and physical findings were considered incidental findings, but more recent papers report increasing numbers of breast cancers detected only at US. PURPOSE: We investigated the yield of US performed as a diagnostic complement in asymptomatic women with mammographic findings that were either negative or poorly readable because of dense breast. MATERIAL AND METHODS: We examined 13 women 37 to 55 years old (mean 47): 9 of them were asymptomatic and 4 had poorly specific physical findings. The patients underwent physical examination, mammography, US, microhistologic biopsy with 14G needles under US guidance and surgery. RESULTS: Fourteen breast lesions 7.0-15 mm in diameter were detected only by US. Mammography (2 or 3 standard views) was negative in all cases. The lesions detected only by US (10% of all carcinomas) were typified with US-guided needle biopsy and finally confirmed surgically. DISCUSSION AND CONCLUSIONS: Though obtained in a small series, our results seem to suggest that US should be included in the diagnostic work-up, especially of women with dense breast. Also, any hypoechoic lesion detected at breast US in clinically asymptomatic women with negative mammographic findings should be further investigated with US, needle aspiration or core biopsy to make the final diagnosis. PMID- 11262821 TI - [Acquired immunodeficiency syndrome and pulmonary Mycobacterium xenopi infection. Role of computerized tomography]. AB - INTRODUCTION: Mycobacterium xenopi is one of the most common agents responsible for nontubercular mycobacterial pulmonary disease on AIDS patients. These lesions have been studied with conventional radiography, while CT has been used in patients with aspecific mycobacterioses or non-AIDS pulmonary conditions from Mycobacterium xenopi. PURPOSE: We investigated the yield of CT in the study of lung lesions from Mycobacterium xenopi in AIDS patients. MATERIAL AND METHODS: We examined 12 AIDS patients with pulmonary lesions from Mycobacterium xenopi, patients age ranged 30 to 46 years. All patients had CD4 blood levels lower than 250 cells/mL and Mycobacterium xenopi in the sputum. All patients underwent a standard chest radiograph and a CT examination. CT images were evaluated by three radiologists independently and the definitive diagnosis was made in the presence of a fourth radiologist. RESULTS: Chest CT showed parenchymal consolidation in 66% of cases, associated with bilateral basal bands in 16% of cases. Consolidation was unilateral in 41% of cases and most frequently involved the right lower lobe. Bilateral reticular interstitial involvement was seen in the patients (41%). Micronodules in 1 patient (8%) and mediastinal adenopathy in 33% of cases. Two patients had pre-xisting emphysema and 1 had bronchiectasis. DISCUSSION AND CONCLUSIONS: The frequency of lung disease from Mycobacoerium xenopi has increased because of the spreading of the HIV infection. Such lung lesions in AIDS patients are aspecific in appearance and localization, which the clinical radiologist needs to consider to address treatment planning. The frequent finding of parenchymal consolidation and the absence of cavitary lesions may be referred to the poor capability of AIDS to produce an adequate inflammatory response. The lung lesions tend to distribute in the lower lobes unilaterally. Adenopathy was also a frequent finding. CT plays a fundamental role in studying the chest of these patients because it permits to locate lung lesions with higher accuracy than conventional radiography and to detect adenopathies, micronodules reticular interstitial involvement and bronchiectases. PMID- 11262822 TI - [Use of virtual endoscopy with computerized tomography in the identification of colorectal neoplasms. Prospective study with symptomatic patients]. AB - INTRODUCTION: Aim of this study was to evaluate the sensitivity of virtual colonoscopy (CT colonography) in the identification of colorectal cancer and to define the limitations and the advantages of this imaging modality, as well as indications to the examination. MATERIAL AND METHODS: We examined prospectively 62 symptomatic patients aged 36 to 82 years (28 women and 34 men). All patients underwent both conventional and virtual colonoscopy on the same day; the conventional examination allowed exploration of the entire colon. RESULTS: Conventional colonoscopy identified 89 lesions 3-50 mm in diameter, namely 84 benign and 5 malignant lesions. No lesions were identified in 12 patients. CT colonography identified 52 of the 89 lesions, with 57.1% diagnostic accuracy. There were 11 false positives (82.5% positive predictive value and 52.2% specificity) and 37 false negatives (24.5% negative predictive value and 58.4% sensitivity). Sensitivity was significantly higher (85.7%) for polyps > or = 1 cm. CONCLUSIONS: Virtual colonoscopy is an imaging modality with good diagnostic yield, well tolerated by patients and with great potentials for further development. We suggest that the examination be performed in symptomatic patients who cannot undergo total colonoscopy or refuse the other imaging modalities. Further studies are warranted in larger series of patients, possibly introducing it in screening programs. PMID- 11262823 TI - [Multiphasic helical computerized tomography of hepatocarcinoma. Assessment after chemoembolization]. AB - PURPOSE: To report our personal experience with the addition of contrast-enhanced multiphase helical CT to unenhanced CT (Lipiodol CT) in the evaluation of patients with hepatocellular carcinoma treated with chemoembolization and to analyze the present role of oily agent CT. MATERIAL AND METHODS: We retrospectively reviewed the examinations of 42 consecutive patients submitted to global chemoembolization over a 2-year period. CT was performed 18-30 days after the treatment. The Lipiodol CT study was carried out with volume acquisitions. We considered as nodules all well-defined areas with dense oily agent uptake; uptake itself was classified as: 0 = absent, I = lower than 10% of the tumor volume, II = lower than 50%, III = higher than 50%, IV = homogeneous. Contrast-enhanced helical CT was performed with the 2-phase technique in 28 patients and with the 3 phase technique in 14; we considered as nodules all well-defined and relatively homogeneous areas with hyperattenuation in the arterial phase and hypo isoattenuation in the portal and/or delayed phase, or with hypo-isoattenuation in the arterial phase and in the portal and/or delayed phase. RESULTS: Lipiodol CT permitted to recognize 65 nodules (1-5/patient, mean 1.5), namely 15 grade I, 21 grade II, 20 grade III and 9 grade IV. Multiphase CT identified 6 additional nodules in 5 patients, 5 hypervascular and 1 hypovascular, and better assessed the correct morphology and volume of grade I nodules. Only 4 of 6 nodules missed on Lipiodol CT showed oily agent uptake after a new chemoembolization session. Moreover after retreatment, carried out in 6 of 9 patients with grade I uptake (11 nodules in all), we found persistence of the grade I pattern in 5 nodules, grade II in 5, and grade III in 1. CONCLUSION: Lipiodol CT may miss liver nodules and underestimate the volume of nodules with poor uptake. Though Lipiodol CT should still be considered slightly more sensitive than multiphase CT, in our opinion this technique has several limitations, as also shown in recent literature papers, and its clinical applications should be reduced. Multiphase helical studies may provide useful information and should be performed routinely in patients treated with chemoembolization. The present availability of alternative tools such as contrast-enhanced Doppler US and MRI should also be stressed and their potential role investigated. PMID- 11262824 TI - [Ultrasonography in the diagnosis of post-pubertal epidemic parotitis and its complications]. AB - PURPOSE: To assess the yield of US in the study of salivary glands and other organs involved in post-pubertal mumps. PATIENTS AND METHODS: We examined 68 patients with serologically proven post-pubertal mumps (age range 14-34 years). All patients were symptomatic, with fever and salivary gland swelling in 25 cases, marked hyperamylasemia in 32, epigastric pain in 9, unilateral scrotal swelling and/or pain in 19 cases and acute bronchitis in 1 case. All patients underwent US of salivary glands, neck lymph nodes, abdomen and scrotum with 48 hours of admission. RESULTS: Salivary glands: Parotid and submandibular glands showed normal echotexture in all patients. The parotid glands also showed multiple hypeoechoic intraparenchymal lymph nodes which were, ovoid or rounded, with smooth margins and a central hyperechoic area, with diameter ranging 3-14 mm (mean 5.4). No intraparenchymal lymph nodes were observed in submandibular glands. Neck: All patients had enlarged submandibular lymph nodes (maximum diameter ranging 5-22 mm; mean 11 mm); swelling was always bilateral and it was symmetric in 19/68 patients (30%) versus asymmetric because of prevailing right side involvement (more numerous and bigger nodes) in the other 47/68 cases (70%). All lymph nodes showed a benign pattern, with an ovoid or elongated shape, homogeneous hypoechoic echotexture and a hyperechoic hilum. Abdomen: The pancreas showed normal volume and normal parenchymal echotexture in all patients. Liver and spleen were always normal. Testes: US showed mild unilateral hydrocele in 10 cases, hydrocele and unilateral swelling of epidymis in 5 cases, hydrocele and swelling of both epidymis and didymis with inhomogeneous echotexture because of intraparenchymal hypeoechoic areas in 2 cases. There were no US changes in 2 cases. CONCLUSIONS: US of the salivary glands shows a specific pattern in post pubertal mumps which has never been reported for other salivary gland diseases. In contrast US signs in other organs are not specific. PMID- 11262825 TI - [Imaging quality of a digital Konica Regius 336 system for thoracic radiology]. AB - INTRODUCTION: Digital radiographic systems permit to optimize execution, depiction and storage of radiological images. Since a Regius 336 digital system (Konica Corp., Tokyo, Japan) devoted to chest radiography was recently installed in the Radiology Department of S. Anna Hospital in Como, Italy, we investigated its performance relative to image quality. MATERIAL AND METHODS: Konica Regius 336 is a computed radiography system made of a phosphorescence detector plate which is scanned with an infrared semiconductor laser beam. The radiographic image obtained from the detector is subjected to image processing, which allows a stable output and the nonlinear curve typical of conventional radiographic systems. Image quality was assessed based on the following parameters: dose, contrast, noise, and spatial resolution. As reference, we assessed the same parameters on a Cronex 88 analogic chest-changer (DuPont Pharma, North Billerica, Mass, USA). RESULTS: The Regius 336 air kerma values were always higher than the analogic ones (about 10%), both with and without a chest phantom; noise was also greater than in analogic images, sometimes even doubled. The optical densities of a step wedge and the spatial resolution of the digital chest-changer are independent of the X-ray tube voltage consequent to broader optical latitude. Inversely, the analogic images of the wedges show great optical density variability as a function of the X-ray tube voltage (in a range of 2). The modulation transfer functions of the two systems have the same trend. DISCUSSION AND CONCLUSIONS: The performance of the Konica Regius 336 is nearly equivalent to that of an analogic system. The main advantages of the digital system are a standard output, lower consumption of radiographic films, higher productiveness and better image quality standard level. PMID- 11262826 TI - [Meniscal ossicle: radiographic and MR findings. Report of a case]. PMID- 11262827 TI - [Role of color Doppler ultrasonography in the diagnosis and monitoring of renal venous thrombosis and of the inferior vena cava in a newborn girl with factor V mutation. A case report]. PMID- 11262828 TI - [Left inferior vena cava, with hemiazygos continuation and atypical flow into the superior vena cava. A case report]. PMID- 11262829 TI - [Iatrogenic intestinal invagination in the adult: computerized tomography features. Report of a case]. PMID- 11262830 TI - [A case of colo-colic invagination in a patient with left colon stenosis ]. PMID- 11262831 TI - [Iatrogenic diverticulum at the level of the Douglas pouch in patient treated with Longo hemorrhoidectomy. A case report]. PMID- 11262832 TI - [Spontaneous regression of hepatocellular carcinoma. Report of a case]. PMID- 11262834 TI - [Unusual case of malignant degeneration of multicystic pancreas in von Hippel Lindau syndrome]. PMID- 11262833 TI - [Sclerotherapy of hepatic cysts with alcohol. New percutaneous technique]. PMID- 11262835 TI - [Primitive neuroectodermal tumor of the kidney. Magnetic resonance features of a case]. PMID- 11262836 TI - [Cystic parathyroid adenoma of the mediastinum with primary hyperparathyroidism. Report of a case]. PMID- 11262837 TI - [Ultrasonography of a case of angiomatous lymphoid hyperplasia (Castleman's disease) of the retroperitoneum]. PMID- 11262838 TI - [Idiopathic retroperitoneal fibrosis. Report of a case studied with spiral computerized tomography]. PMID- 11262839 TI - [The infant's effort]. PMID- 11262840 TI - [Sialolithiasis]. PMID- 11262841 TI - [The meeting of radiology and literature]. PMID- 11262843 TI - [Chronic renal failure and pregnancy]. AB - The women who are suffering from chronic renal failure in an advanced stage have a deficient fertility but they are not sterile. Hemodialysis has improved considerably the fertility of these patients. The aim of this study is to give the results of our experience, from 1990 to 1996, about pregnancies among the uremic patients, dialysed or no and to make a literature review about this subject. We have noticed that pregnancies in the dialysis patients are rare and their evolution is precarious. We have also noticed more miscarriage or pregnancy interruption. Complications are frequent. Mothers have a high risk of hemorrhagic accident (ablatio placentae), of anemia aggravation, of thrombosis of the vascular approach and a high risk of liver anomalies (gravidic cholestasis). The fetus suffers from the maternal anemia and from chronic hypoxia. He's threatened by hydramnios in the case of bad volemic supervision. The intra uterine delayed developement and the prematurity are usual. The absence of high blood pressure and a residual renal function are representing the favourable elements of the good march of pregnancy. A therapeutic intensification is necessary in order to lead this pregnancies to a viable term. The management is heavy not only for the nephrologic, the obstetrical and the neonatal physicians, but also for the patient who is the only one who can decide to continue or to interrupt the pregnancy. It seems better to inform the patient rather than to procure her abortion by proposing her an effective and inoffensive contraceptive method meanwhile to be pregnant after renal transplantation. PMID- 11262842 TI - [Comparison of different protocols of ovulation induction, by GnRH agonists and chorionic gonadotropin]. AB - OBJECTIVE: The aim of this study was to determine the best way of using a gonadotropin-releasing hormone agonist (GnRHa) for triggering ovulation and to analyse the reasons for short luteal phases. MATERIALS AND METHODS: Thirteen different regimens of GnRH-a administration were used to trigger ovulation using different dosages and either one, two or three administrations: triptorelin, buserelin spray, buserelin subcutaneously, leuprolide and nafarelin in 231 treatment cycles. Pregnancy rate and luteal phase duration were compared with those of a control group for whom ovulation was triggered with hCG. RESULTS: Ovulation with supraphysiologic elevation of both FSH and LH was achieved in the 13 GnRHa groups. For the five main groups analysed, GnRHa produced shorter and inadequate luteal phases than did hCG but no difference was found between agonists. Pregnancy rates were not statistically different between the agonist groups or in comparison with the hCG group. CONCLUSION: The use of GnRHa to trigger ovulation is efficient, despite short luteal phases, and has proven its utility in comparison with hCG. As the different modes of stimulation appear to yield comparable results, the cost of treatment should be a significant element to take into consideration. PMID- 11262844 TI - [Comparison of 2 therapeutic strategies for severe endometriosis, in young women counsulting for sterility or pain. Results in cases of chronic pelvic pain]. AB - AIM OF THE STUDY: Compare two medical strategies associated to surgery in women requiring for chronic pelvic pain due to stage III-IV endometriosis. MATERIAL AND METHODS: Two different patient groups, A (N 27) and B (N 41), requiring for chronic pelvic pain, associated with AFS stage III-IV endometriosis, operated on from 1992 to 1997, were compared. The medium age was 35 and 34 years, respectively. Pelvic pain, classified in three stages, was similar in both groups but they were more AFS stage IV in group A, 67% than in group B, 46% (p < 0.01). Both groups had similar operative procedure: laparoscopic resection of deep endometriotic nodules or endometriomas, plus destruction of small superficial lesions using CO2 laser (A) or bipolar coagulation (B). Associated medical strategy was different: group A: operative laparoscopy without preoperative treatment and in 25% a second laparoscopy taking place after two-three months of LHRH analogues; no postoperative treatment; group B, operative laparoscopy taking place after ovarian blockage with three-six weeks of Diane (Androcur + ethinyl estradiol), then two-three months of analogue postoperative treatment immediately followed by long term progestoid treatment in order to prevent recurrences in women without pregnancy desire. RESULTS: After one year, 6/27 (22%) of A and 3/41 (7%) of B had no follow-up. Of the followed patients, a complete improvement was observed in 18/21 (86%) A, 33/38 (87%) B, moderate pelvic pain continued in 2/21 (10%) A, 4/38 (11%) B, and the treatment was in failure in 1/21 (5%) A, 1/38 (3%) B, without significant difference. After two years, 67% of A and 76% of B had a follow-up and the corresponding rates of complete improvement are 72% (A), 87% (B), incomplete improvement: 22% (A), 10% (B) and failure: 6% (A), 3% (B). The difference is lightly significant (p < 0.05) and remains so if patients without follow-up are considered as failures. There was no persistence nor recurrence of endometriosis nor endometrioma two years after the surgery was completed. CONCLUSION: Since there were more stage IV endometriosis in group A than in B, the different medical strategies and particularly the long term postoperative treatment used in B seem have little influence on results. However, these data was obtained in women of medium age > 30, with a relatively short follow-up. It should be of interest to compare in a prospective multicentric study the long term follow-up of two cohorts of young women operated on for stage III-IV endometriosis, receiving or not a long term medical treatment after surgery in order to prevent recurrences. PMID- 11262845 TI - [Personality traits of men with Klinefelter syndrome and their partners]. AB - The aim of the study was to find whether personality traits of men with Klinefelter syndrome and their partners (group 1-n = 17) differ from those of couples affected by idiopathic infertility (group 2; n = 16) and from those of fertile couples (group 3 n = 17). We further investigated the attitudes of the three groups towards pregnancy, labour and sexuality to find potential differences among the three groups. Besides, we verified the hypotheses of below average or low average intelligence of men with Klinefelter syndrome, and of inferior quality of social life in these men. The data were collected using the interview on medical history, the questionnaire on attitudes towards pregnancy, labour and sexuality (S-S-G), the personality questionnaire MMPI-2. The Raven progressive matrices were used only in group 1. The results show that men with Klinefelter syndrome and their partners do not differ significantly from the couples with idiopathic infertility (group 2), having some shizoide traits in their personality structure and mostly negative attitudes towards pregnancy, labour and sexuality. However, a significant difference has been found between the Klinefelter syndrome group and the fertile couples group. The hypothesis of below average intelligence has not been confirmed, but the quality of social life of men with Klinefelter syndrome has been found inferior. We may thus conclude that in the management of infertile couples in whom the man has been affected by Klinefelter syndrome, the personality structure, importantly affecting the outcome of treatment, should be taken into consideration. PMID- 11262846 TI - [Deliverance of emergency contraception, in 1988, in family planning and education clinics (FPEC) of Val de Marne]. AB - OBJECTIVES: To facilitate access to emergency contraception (EC). To allow the nurses in the family planning clinics to deliver it during the doctor's absence. METHODS: For one year, 1998, 1102 women requested EC in the 38 family planning clinics participating in the study. This study evaluated the utilisers and the circumstances under which dispensation occurred. RESULTS: The users of EC were young, 45% under 18 years and 90% under 25 years. There was a marked difference between the contraception the women declared they used and that which they actually did during their last episode of sexual intercourse. Women requested EC in case of condom breakage or slipping (49%), forgotten pill (8%), or after unprotected intercourse (43%). Nurses personally received 809 requests for EC, and 293 women were received by the family planning doctor. In 77% of the cases, EC was given by the nurses, and for the others after medical opinion. But only 7% had a medical "problem" (contraindications to estrogen-progestin, or cycle disorder). Among the 823 women for whom information was obtained, 22 unwanted pregnancies were observed, 18 of these patients decided to have an abortion. 97.3% efficacity. CONCLUSION: Making EC more easily available in family planning clinics with dispensation by nurses does no harm and may reduce the rate of unwanted pregnancy. PMID- 11262847 TI - [In response to the article by M. Levert et al. Human Papillomavirus typing in routine cervical screening. Results on a series of 3778 patients. Gynecol Obstet Fertil 2000; 28: 722-8]. PMID- 11262848 TI - [In response to the article by J. Saias-Magnan et al. Gynecol Obstet Fertil 2000; 28: 896-903]. PMID- 11262849 TI - [In response to the article by E. Verdy. Evaluation of thromboembolic risk after ovarian stimulation: what balance? What prevention? Gynecol Obstet Fertil 2000; 28: 875-9]. PMID- 11262851 TI - [Endovaginal ultrasonography of the uterine cervix in asymptomatic populations at high risk and at low risk for premature birth: to do or not to do?]. PMID- 11262850 TI - [Phytoestrogens. Therapeutic use]. PMID- 11262852 TI - [Role of diagnostic laparoscopy within the framework of infertility evaluation. For the systematic practice! ]. PMID- 11262853 TI - [Role of diagnostic laparoscopy within the framework of infertility evaluation: against the systematic practice]. PMID- 11262854 TI - [The First World War and the explosion and morbidity of venereal diseases. V. The therapies]. PMID- 11262855 TI - [Serologically different couples are not the same! Fertility of serologically different couples]. PMID- 11262856 TI - [Can donor insemination be optimised?]. AB - OBJECTIVE: To compare the different donor insemination technics. MATERIAL AND METHOD: Analysis of the published studies about donor insemination which value the effectiveness of Intra Cervical Insemination (ICID) and Intra Uterine Insemination (IUID), the interest of ovulation induction, the possible complications, and the cost-effectiveness ratio. RESULTS: The meta-analysis of the Cochrane data base (10 comparative studies IUID versus ICID, 2568 donor insemination cycles) lead to a pregnancy rate per cycle (PRC) 17.77% with IUID versus 7.68% with ICID. The odds ratio is 2.63 (CI from 1.85 to 3.73). With these PRC, the direct cost per evolutive pregnancy is 54,780 F with ICID and 25,675 F with IUID. CONCLUSION: If it is possible to propose ICID to patient with an excellent regularity of ovulation. IUID with ovulation induction by gonadotropins is today the gold standard, and more especially as the law restrict the number of donor inseminations. Indeed, the IUID is two or three times more effective than ICID, consume the half of sperm straws, use a semen of moderate quality, there is no complication provided that the cycle is cancelled if there is more than two mature follicles and the cost-effectiveness ratio is greatly in favour with IUID. PMID- 11262857 TI - Evaluation & usefulness of a immunochromatographic test for rapid detection of Plasmodium falciparum infection. AB - Rapid test (Parachek Pf) based on detection of HRP-2 protein specific to Plasmodium falciparum by immunochromatographic technique was evaluated. Prevalence of infection was 8.5%. The test was 100% sensitive & 99.5% specific on comparison with light microscopy. The test is useful for making 'on the spot' diagnosis. PMID- 11262858 TI - An analysis of the teratagenic effects that could possibly be due to alcohol consumption by pregnant mothers. AB - It can be concluded that alcohol is definitely harmful to the developing fetus. The effect can manifest in various ways, the most extreme of which is a condition called Fetal Alcohol Syndrome (FAS). The diagnosis of maternal alcoholism leading onto cases of FAS is difficult due to absence of accurate diagnostic tests. The diagnosis of FAS in a child is easier by a proper examination. There is no specific treatment of FAS in a child. The only management is by institution of corrective and rehabilitative measures. The exact mechanism of the teratogenic action of alcohol is not known. It is probably due to the harmful effect of alcohol on the epiblast layer of the bilaminar germ disc. In the absence of adequate knowledge regarding FAS, not much can be done to remedy the deleterious effects of alcohol. Hence, a word of advice to all pregnant women is to avoid drinking during pregnancy. PMID- 11262860 TI - Clinical significance of lipid profile in cancer patients. AB - In the present study, we have examined lipid profile in normal healthy age matched control and patients with various malignancies. Analysis of data revealed that total lipids, cholesterol and HDL cholesterol levels are inversely associated with incidence of cancer where as triglycerides levels were significantly elevated in cancer patients. Electrophoretic separation of lipoproteins revealed a significant decrease in the mean values of alpha fraction in patients with malignancy when compared with the corresponding control group. The other fractions beta and pre-beta did not show any change in the mean values in patients with cancer as compared to the normal corresponding control group. PMID- 11262859 TI - Evaluation of rapid kits for detection of HIV, HBsAg and HCV infections. AB - Diagnostic kits (J. Mitra Co. Ltd) for rapid detection of HIV, HBsAg & HCV were evaluated against respective enzyme immunoassay (Ortho Diagnostics). All rapid kits were 100% specific for HIV, HBsAg & HCV. Sensitivity for HIV, HCV & HBsAg was 80%, 87.5% & 93.4% respectively. PMID- 11262862 TI - Medical problems during pregnancy. PMID- 11262861 TI - Clinical significance of cancer antigen, CA 15.3 in breast cancer. AB - Breast cancer is the leading type of cancer in women. It is commonly accepted that the earlier the detection of the disease, the better the prognosis. Therefore, the present study was undertaken to evaluate the clinical significance of tumour marker CA 15-3 along with Carcinoembryonic antigen (CEA) in India women population. Tumour marker CA 15-3 concentrations reflected the tumour burden in a better way than CEA. However, the use of CEA as an additional conventional marker improved the Clinical efficiency of the marker CA 15-3. PMID- 11262867 TI - Milk thistle: effects on liver disease and cirrhosis and clinical adverse effects. PMID- 11262868 TI - DNA methylation in health and disease. AB - DNA methylation has recently moved to centre stage in the aetiology of human neurodevelopmental syndromes such as the fragile X, ICF and Rett syndromes. These diseases result from the misregulation of genes that occurs with the loss of appropriate epigenetic controls during neuronal development. Recent advances have connected DNA methylation to chromatin-remodelling enzymes, and understanding this link will be central to the design of new therapeutic tools. PMID- 11262870 TI - Tuning in to perfect pitch. PMID- 11262869 TI - Neuronal specification in the spinal cord: inductive signals and transcriptional codes. AB - Neural circuits are assembled with remarkable precision during embryonic development, and the selectivity inherent in their formation helps to define the behavioural repertoire of the mature organism. In the vertebrate central nervous system, this developmental program begins with the differentiation of distinct classes of neurons from progenitor cells located at defined positions within the neural tube. The mechanisms that specify the identity of neural cells have been examined in many regions of the nervous system and reveal a high degree of conservation in the specification of cell fate by key signalling molecules. PMID- 11262871 TI - Microbial genome analysis: insights into virulence, host adaptation and evolution. AB - Genome analysis of microbial pathogens has provided unique insights into their virulence, host adaptation and evolution. Common themes have emerged, including lateral gene transfer among enteric pathogens, genome decay among obligate intracellular pathogens and antigenic variation among mucosal pathogens. The advent of post-genomic approaches and the sequencing of the human genome will enable scientists to investigate the complex and dynamic interplay between host and pathogen. This wealth of information will catalyse the development of new intervention strategies to reduce the burden of microbial-related disease. PMID- 11262872 TI - Genome evolution. Arabidopsis 4, tomato 1. PMID- 11262873 TI - The origins, patterns and implications of human spontaneous mutation. AB - The germline mutation rate in human males, especially older males, is generally much higher than in females, mainly because in males there are many more germ cell divisions. However, there are some exceptions and many variations. Base substitutions, insertion-deletions, repeat expansions and chromosomal changes each follow different rules. Evidence from evolutionary sequence data indicates that the overall rate of deleterious mutation may be high enough to have a large effect on human well-being. But there are ways in which the impact of deleterious mutations can be mitigated. PMID- 11262874 TI - Molecular profiling of human cancer. AB - Traditionally, tumours have been categorized on the basis of histology. However, the staining pattern of cancer cells viewed under the microscope is insufficient to reflect the complicated underlying molecular events that drive the neoplastic process. By surveying thousands of genes at once, using DNA arrays, it is now possible to read the molecular signature of an individual patient's tumour. When the signature is analysed with clustering algorithms, new classes of cancer emerge that transcend distinctions based on histological appearance alone. Using DNA arrays, protein arrays and appropriate experimental models, the ultimate goal is to move beyond correlation and classification to achieve new insights into disease mechanisms and treatment targets. PMID- 11262875 TI - Diversification of haematopoietic stem cells to specific lineages. AB - Diverse types of blood cell (lineages) are produced from rare haematopoietic stem cells that reside in the bone marrow. This process, known as haematopoiesis, provides a valuable model for examining how genetic programs are established and executed in vertebrates, and also how homeostasis of blood formation is altered in leukaemias. So, how does an apparently small group of critical lineage restricted nuclear regulatory factors specify the diversity of haematopoietic cells? Recent findings not only indicate how this may be achieved but also show the extraordinary plasticity of tissue stem cells in vivo. PMID- 11262876 TI - Immunogenetics. Not just another editor. PMID- 11262878 TI - The mouse that eats less but gains weight. PMID- 11262877 TI - Horticulture: the font for the baptism of genetics. AB - This year marks the centenary of the rediscovery of the laws of heredity, and their introduction to the English-speaking world. Here I introduce the main events and the characters who figure in this story before turning to the task of this essay--to ask why it was that support in England for the emerging science of genetics, or Mendelism as it was then called, came chiefly from horticulture, and was only belatedly accepted into the mainstream of British academic biology. PMID- 11262879 TI - Genomics in the public domain: strategy and policy. AB - The public domain has been conspicuous in media accounts of public and private sector initiatives to complete the sequence of the human genome. The issue of whether the human genome will be freely available to the public or privately held as a proprietary resource has captured the attention of the scientific, trade and popular press, the financial markets, and even heads of state. Although some media commentary has framed the issue as a conflict between ethics and greed, strategic considerations go a long way towards explaining the timing and quality of information disclosures on both sides of the public-private divide. PMID- 11262880 TI - Evo-devo: the evolution of a new discipline. AB - The history of life documented in the fossil record shows that the evolution of complex organisms such as animals and plants has involved marked changes in morphology, and the appearance of new features. However, evolutionary change occurs not by the direct transformation of adult ancestors into adult descendants but rather when developmental processes produce the features of each generation in an evolving lineage. Therefore, evolution cannot be understood without understanding the evolution of development, and how the process of development itself blases or constrains evolution. A revolutionary synthesis of developmental biology and evolution is in progress. PMID- 11262881 TI - A mean spleen gene. PMID- 11262882 TI - Developmental biology. New balancing tricks. PMID- 11262883 TI - Evo-devo. A head with no torso. PMID- 11262884 TI - Gene expression. The nonsense police. PMID- 11262886 TI - [Cataract surgery in primary open angle glaucoma]. AB - Cataract surgery has a moderate lowering effect on the intraocular pressure. When glaucoma seems controlled by medication, phacoemulsification gives a chance of improving the situation. Those eyes are most likely to get an intraocular pressure rise by retention of visco-elastic substance. PMID- 11262885 TI - [Surgical indications in coexisting cataracts and glaucoma]. AB - Cataract surgery in glaucoma patients remains a controversial subjects. Indication of surgery depends on a lot of clinical parameters: diagnosis, state, evolution of glaucoma as well as compliance with medical treatment--surgical procedures of cataract and glaucoma--sites of the surgery--use of antifibrosis agents and surgeon's experience. As cataract extraction alone decreases the intraocular pressure in open angle glaucoma and mainly in uncomplicated closed angle glaucoma and trabeculectomy alone reduces the intraocular pressure more than combined surgery with less complications we recommended the following surgical options: Cataract extraction alone in patients with controlled open angle glaucoma and in patients with closed angle glaucoma. A two step procedure: filtering surgery followed by cataract extraction in patients with poorly controlled open angle glaucoma or mixed closed angle glaucoma. Ambulatory surgery and topical anesthesia permit a two stages surgery with less inconveniences. A combined procedure in patients with a chronic closed angle glaucoma where filtering procedure alone is associated with important complications. Actually, the best surgical cataract procedure is phacoemulsification with a small supero corneal incision and implantation of a foldable intraocular lens. The best filtering procedure remains trabeculectomy, or the new non penetrating trabecular surgery for experimented surgeons, in the superior quadrant. In the future new surgical procedures and new safe and non toxic pharmacologic drugs which modulate wound healing could be found in order to increase the efficacity and indications of combined surgery. PMID- 11262887 TI - [Non perforating trabecular surgery with reticulated hyaluronic acid implant]. AB - Non perforating trabecular surgery (NPTS) with reticulated hyaluronic acid implant (Skgel) allows aqueous humor to leave anterior chamber through a thin trabeculo-Descemet's membrane into a sclerocorneal space filled with Skgel implant and then via the outflow physiological channels. Good intraocular pressure results are obtained with less or without external filtration decreasing the incidence of per- and postoperative complications found after trabeculectomy. This surgery is actually only indicated for primary open angle glaucoma, the trabeculectomy still remaining the gold standard procedure for the other glaucoma cases. PMID- 11262889 TI - [Combined surgery for cataract and glaucoma: phacoemulsification and nonpenetrating surgery]. PMID- 11262888 TI - [Chronic open angle glaucoma and cataract: when is it necessary to perform combined surgery?]. AB - The combined surgical treatment of cataract and glaucoma has a long and controversial history (1). The guidelines proposed by the European Glaucoma Society facilitate however the therapeutic decisions (2). If the patient with open-angle glaucoma is stabilized with medical treatment it is often sufficient to operate the cataract alone and continue the medical treatment afterwards. If the patient is not stabilized or if the medical treatment is not well tolerated, it is suggested to perform a combined procedure. If visual functions are threatened by the glaucoma it is better to perform the filtering procedure first. We shall overview these options and elaborate on each surgical technique. PMID- 11262890 TI - [Prevention and treatment of complications due to combined surgery for glaucoma and cataract]. AB - Complications of combined glaucoma and cataract surgery are frequent. It is why prevention is necessary, based on rightness of the operative indication, the most appropriate technique and the quality of the execution, main topics reviewed in this paper. When prevention has not been sufficient and complications occur, it is necessary to recognize their causes. Complications may occur early or late, may not induce any clinical consequence or on the contrary induce serious ones. The most frequent complications and their treatment are reported. A technique of surgical revision, useful either in case of filtration failure or in case of severe hypotony, is proposed and discussed according to the most recent data in the literature. PMID- 11262891 TI - [A proposed variation for glaucoma surgery]. AB - When a trabeculectomy does not reduce the intraocular pressure, it is usually due to sealing of the scleral flap to the adjacent sclera, while the trabeculectomy itself remains open. A triangular excision in the scleral flap seems likely to keep the fistula in function. Deep sclerectomy does not lead to major hypotony, "the other complication" of trabeculectomy. However, the non perforating sclerectomy, under a sutured superficial scleral flap, could in the long term become inefficient. I propose a deep sclerectomy with controlled perforation under a notched superficial flap. PMID- 11262892 TI - [Cataract extraction after angle closure glaucoma]. AB - The best therapeutic approach to angle closure with cataract is phacoemulsification and foldable implant. The surgery is performed as soon as the acute crisis is over. Both dispersive and cohesive visco-elastic substance are used in the narrow anterior chamber. Poor mydriasis is best managed by means of iris hooks. PMID- 11262893 TI - [The financing of medical research and what we should know before writing grant applications]. PMID- 11262894 TI - [Familial hyperlipoproteinemias--correlations between phenotypes and genotypes]. AB - Within the grant project patients with familial hyperlipoproteinaemias have been examined. The examination was performed in the oldest lipid clinic and research laboratory in the world. The classification of lipid metabolism disorders was based upon a detailed biochemical analysis of plasma lipids including electrophoresis and assessment of apolipoprotein levels. Then optimal treatment regimen could be established. The project was aimed to evaluate the efficacy of different treatment regimens in different types of hyperlipoproteinaemias. Biochemical parameters and mainly the impact of treatment of hyperlipoproteinaemia on morphology and function of the vessel wall was monitored. The non-invasive ultrasound measurement of the intima thickness of carotid arteries was used. For more precise diagnosis of genetically determined disorders of lipid metabolism a large scale of methods of molecular biology was introduced. These methods enable confirmation of familial hypercholesterolaemia, familial defective apolipoprotein B-100 or studying polymorphism of apolipoprotein E. The effort of the authors of the project was to maximally utilise the results of basic and applied research in formulating recommendations for everyday practice of physicians. PMID- 11262895 TI - [Metaphylaxis in the first occurrence of urolithiasis]. AB - BACKGROUND: Urolithiasis is a socially important disease with a high tendency to recurrences. By specific medicamentous metaphylaxis it is possible to achieve an as high as tenfold reduction of recurrences, however, regular check-ups focussed on possible side-effects are required. The objective of the project was to assess prospectively the effectiveness of non-medicamentous metaphylaxis in patients with the first kidney stone. In 113 patients, a fluid and specific dietary regimen were recommended based on a comprehensive diagnosis of metabolic disorders (Table 1) and the regimen was modified with regard to results of metabolic check-up examinations after 6, 18, and 36 months (group 1). Ninety-four patients were recommended a fluid and non-specific dietary regimen after a limited metabolic evaluation with a subsequent check-up after 36 months (group 2). The two groups were clinically comparable. A stone recurrence developed in 7 (6%) patients of the group 1 and in 18 (19%) of the group 2 (p < 0.01). The difference was even greater after including the growth of the concrement (7% and 23%). The patients with a recurrence or growth of a stone had more frequently weddellite in the concrement and had also more frequently a bilateral residual or conservatively managed urolithiasis (p < 0.0001). Half the recurrencies were asymptomatic. The development of metabolic disorders in the group 1 indicated a gradual decrease of uric acid in serum and urine (p < 0.01) although it was not yet significant after 6 months. There was also a significant increase of calciuria (p < 0.01), most probably in conjunction with a regular calcium intake. CONCLUSION: Specific non-medicamentous metaphylaxis with an adequate calcium intake leads to a lower incidence of stone recurrences than a non-specific fluid and dietary regimen. It is justified to introduce the specific metaphylaxis in all patients after the first diagnosis of a kidney stone. It ensures subsequent adjustment of the individual diet which the patient is more likely to adhere to than to general non-specific instructions. PMID- 11262896 TI - [Pharmacologic control of hypophyseal tumors: interactions of estrogens, thyroid hormones, growth and anti-growth factors]. AB - Estrogens are involved in anterior pituitary (AP) growth and tumor transformation of AP. Central dopaminergic and noradrenergic systems are probably most important systems in regulation of AP growth. Estrogen-induced AP growth is associated with decreased metabolism of central catecholamines and increased dopaminergic DA-2 receptors. Application of thyroid hormones or methylene blue prevents both estrogen-induced catecholaminergic inhibition and dopamine DA-2 receptors increment in the AP. The alone given methylene blue increases the dopaminergic activity and the binding sites for dopamine. The study of interaction of natural regulators or synthetic compounds with estrogen-induced pituitary growth will be of value to understand better mechanisms of pituitary tumor formation and possibly find new approaches towards treating patients with these tumors. PMID- 11262897 TI - [Pronuclear and antinuclear factors in the pathogenesis of cholesterol cholelithiasis]. AB - Contrary to hitherto published results, the authors provided evidence of significant pronucleation activity in the protein fraction which is not linked to concanavaline A. Delipidation or proteolysis markedly reduce the pronucleation activity of this fraction. Albumin was identified as the main protein in this fraction. The lipid-protein complex formed by albumin and lipids had a high pronucleation and crystallization activity in relation to cholesterol. Calcium ions increased the crystalization activity. Complexes formed by proteins and lipids can be vectors of the main pronucleation activity in bile. In investigations of the main cholesterol fraction the authors provided evidence that only part of so-called pronucleation proteins is linked to vesicles--i.e. IgM, IgA and biliary glycoprotein BGP I and II. The authors assume that only proteins firmly linked to vesicles can participate in the process of cholesterol crystallization. Biliary glycoprotein BGP I and II was present in vesicles and when added into a model bile it presented a high pronucleation activity. Biliary glycoprotein is a new hitherto not identified pronucleation protein in bile. PMID- 11262898 TI - [Papillomaviruses and human tumors]. AB - The report summarizes the main results obtained in the course of our research project. The results of immunological and epidemiological studies provide further proofs that human papillomaviruses (HPV) are the etiological agents in cervical neoplasia. In addition, they raise hopes that immunological methods may be utilized in diagnostics of cervical cancer and for monitoring the clinical course of this disease in the near future. Since the etiological relationship between HPV and cervical carcinoma seems to be proven beyond reasonable doubt, the development of prophylactic and therapeutic vaccines has become the dominant of the contemporary HPV reseach. For studying immune reactions against HPV-induced tumours we developed a model of HPV16-transformed rodent cells. PMID- 11262899 TI - [Selected phytoestrogens with potential beneficial effects on risk of certain environmental diseases]. AB - Evidence has been accumulated that some phytoestrogens act as protective factors against development of cancer and also cardiovascular diseases. These are phytoestrogens of isoflavone and lignane series, found especially in soy. Beneficial effect of these compounds may be explained by a complexity of their actions at various levels: they interact with estrogen receptors, some of them are inhibitors of the key enzymes responsible in the final effect for cell growth and proliferation, and, due to their chemical nature they are scavengers of free radicals. In the presented work the authors, in collaboration with Finnish partners from the University of Helsinki, developed original immunoassays for determination of main phytoestrogens of isoflavone series--daidzein, genistein, formononoetin, biochanin A, their metabolite equal and lignane enterolactone. The methods are sensitive enough for follow-up of actual levels of phytoestrogens in serum. By using these methods, the levels of phytoestrogens in Czech population have been established. The group of patients suffering from osteoporosis has been investigated, too. Significantly lower levels of isoflavonoids in comparison with sex- and age-matched healthy subject have been found in the patients. The methods have also enabled us to follow up the dynamics of these compounds in the organism, as well as to determine their content in food and its sources. The original detection and quantification of four above mentioned isoflavonoids in beer is an example. PMID- 11262900 TI - [The spiral Z-stent with a knit dacron coating in endoluminal therapy of aneurysms of the abdominal aorta]. AB - The authors present their own experience with construction, experimental testing and clinical application of endoluminal grafts. In the first part of the project, a new selfexpandable stentgraft was constructed on the basis of spiral Z stent. Spiral Z stent was covered by ultrathin polyester sleeve. Next, stentgraft was tested on animal model of abdominal aortic aneurysm. On the basis of experimental results endoluminal grafting of AAA was started in Czech Republic. PMID- 11262901 TI - [Determination of DNA adducts in persons with occupational exposure to alkenes]. AB - DNA adducts induced by alkenes were studied in in vitro systems (cell cultures) and in experimental animals. Properties of DNA adducts, their relationship with other parameters of genotoxic effect and DNA repair were followed. Based on these results, human biomonitoring studies were conducted in order to elucidate the effect of exposure on enhanced levels of DNA and haemoglobin adducts, single strand breaks in DNA and HPRT mutant frequencies. Surprisingly, no correlation was found between primary DNA lesions and HPRT mutant frequencies, suggesting that no simple quantitative relationship could be drawn between primary DNA damage and mutagenesis. In order to understand mechanisms of genotoxic effects of xenobiotics, further studies aimed at individual susceptibility, individual repair capacity and the role of specific DNA adducts in mutagenesis, are required. PMID- 11262902 TI - [Significance of schistosome lectins in the host allergic reaction evoked by penetration of cercaria]. AB - Cercarial dermatitis is a skin allergic response caused by larval stages (cercariae) of the trematode family Schistosomatidae. In the Czech Republic the main causative agents of the disease are bird schistosomes of the genus Trichobilharzia. In the past it was supposed that cercariae of animal schistosomes die soon after the penetration into the human skin. However, we observed a migration and partial development of T. szidati cercariae within a nonspecific (mouse) host. The initial development of parasites in mice was similar to that observed in a specific host (duck) as well as to that described in human schistosomes causing schistosomiasis. After penetration, the transformation of T. szidati cercaria to schistosomulum is characterised by ultrastructural and biochemical changes, resulting in formation of a new parasite surface as well as in differences in specific binding of various lectins and host immunoglobulins to parasite surface components. It seems that the transformation of parasites represents a part of their immune evasion within the infected host. It was observed that T. szidati cercariae possess lectins localized on the parasite surface and in postacetabular penetration glands. It is supposed that, after the penetration into the skin, when glycocalyx disappears and gland material is expelled, the parasite surface may serve as an activator of the alternative complement pathway. The postacetabular gland components have probably a lytic function and facilitate migration of parasites through the skin. Moreover, the gland content is considered to play a role in shedding of surface antigens and in changes of parasite tegumental architecture. PMID- 11262903 TI - [The Internal Grant Agency of the Czech Ministry of Health from 1997 to 2000]. PMID- 11262905 TI - [Pericentric inversion of human chromosomes and its risks] ]. AB - Pericentric inversions of human chromosomes represent rearrangements are formed between two breaks on the short and on long arms of the chromosome with following rotation and new connection of the separated segment in the reversed position. The abnormality does not result in most of the carriers to any clinical manifestations. However, the basic risk the carriers of such inversion are exposed is the possibility of formation of a recombinant aneusomy--later transformation of the inverted chromosome during gametogenesis. Conception by the recombinant gamete usually results in spontaneous abortion or to a birth of seriously affected individual. The risk of recombination has to be in every newly registered inversion individually considered. The larger part of chromosome is taken into the pericentic inversion, the smaller is the extent of resulting duplication and smaller is the deficiency of chromosomal parts, which results from the recombination. The higher is then the viability of the affected foetus. In families with detected recombination, chromosomal examination is fully recommended. Prenatal examination is also indicated when the transformation is listed among the risk inversions or it has larger extent then recorded inversions. Beside the risk inversions, also the "safe" inversions exist, which include minor and frequently occurring transformation of chromosome No 2- inv(2)(p11q13) and inversion of chromosome 10--inv(10)(p11q21). PMID- 11262904 TI - [Rhinitis--still a real problem]. AB - Paper brings about findings on rhinitis--definition, classification and clinical picture. New discoveries concern mycotica infection and some methods of investigation. Differential diagnosis of rhinosinusitis includes several diseases. Author presents some possibilities of therapy of rhinitis and includes relations between upper and lower respiratory airways. PMID- 11262906 TI - [Deletion of the long arm of chromosome 5 in patients with hematologic malignancies]. AB - Deletion of part of the long arm of chromosome 5 (5q-) is one of the most common structural aberrations in patients with myeloid disorders. The deletion is interstitial and the deleted segment is variable in size and breakpoint localization. Precise analysis of chromosomal breakpoints proved that band 5q31 is the most common deleted region. Extensive molecular studies have been performed to identify one or several tumor suppressor gene(s) in this critical region. Although these genes have not been identified as yet, the candidate genes are being intensively studied. Isolation and characterization of tumor suppressor genes will lead to the understanding of molecular mechanisms of normal hematopoiesis and that of leukemic transformation. PMID- 11262907 TI - [Injuries of the upper and middle thirds of the face. Analysis of the cause of injury] ]. AB - BACKGROUND: From the point of traumatologist, the size, locality and direction of the injuring force give the extent of the injury of facial skeleton. The aim of the paper is to present an overview on aetiology of injuries of the upper and medial third of face in relation with the incidence, gender and age of patients. METHODS AND RESULTS: 282 adult patients (208 males and 74 females, average age 40 years) and 25 children (18 boys and 7 girls, average age 9 years) were treated since 1.1.1998 till 31.8.1999. 25 of them (9%, 19 males and 6 females) had fracture of the middle third of the face. In 5 patients (20%), fractures were accompanied by commotion of the brain. Conservative treatment was used in 16 patients (64%) and surgical approach in 9 of them (36%). Isolated fractures of nasal bones were diagnosed in 230 adult patients (82%, 168 males, 62 females, 25 children). In 19 adult patients (8%) the fracture was open, in 25 of them the reposition of dislocated bones was done in general anaesthesia. 27 patients (10%, 21 males, 6 females) had polytrauma, accompanied by the facial bones fractures of the upper and middle third. Facial fractures of the upper and middle third were caused by violence of other person (criminal injury) in 15 patients (60%), downfall in 66 patients (29%), sport injury in 38 patients (17%), traffic injury in 16 patients (7%). The most common cause of isolated fracture of nasal bones in children was downfall in 12 patients (48%), violence of other person or schoolmate in 7 patients (28%) sport injury in 5 patients (20%), traffic injury in 1 patient (4%). In 27 persons with polytrauma and facial fractures of the upper and middle third, the injury was caused by traffic accident in 13 patients (48%), downfall in 8 patients (30%), sport injury in 2 patients (7.4%), criminal injury in 2 patients (7.4%), work injury in 1 patients (7.4%). CONCLUSIONS: Polytrauma belongs to the most serious injuries both due to their extent and their significance because in most of them the life of patients is directly threatened. Criminal injuries represent very common cause of the facial fractures of the upper and middle third with maximum prevalence in males of the third decade. Part of those injuries belong to polytrauma. PMID- 11262908 TI - [Use of the paraxanthine/caffeine ratio in the saliva of patients with liver cirrhosis]. AB - BACKGROUND: Caffeine elimination and monitoring of its metabolites after a single peroral administration in used to evaluate liver metabolic function. The aim of the study was to use the paraxanthine/caffeine ratio in saliva to evaluate liver function in patients with liver cirrhosis. Results were correlated with various procedures evaluating the elimination rate of caffeine. METHODS AND RESULTS: The study group consisted of 7 patients with compensated liver cirrhosis and 10 healthy volunteers, all individuals were given a single 280 mg dose of caffeine perorally. Concentration of salivary caffeine was determined in each person with high performance liquid chromatography after 4, 6, 8 and 12 hours after application. The paraxanthine/caffeine ratio was evaluated at the 6-hour interval. Estimated clearance (CL), half-life (t1/2) and volume of distribution (Vd) were calculated from 2 or 4 concentrations of the drug. The metabolic ratio of the patients (x = 0.15 +/- 0.07) was significantly lower then the control group (x = 0.55 +/- 0.29). Using a four-point procedure we acquired the following values: patients--CL 2.39 +/- 2.36 l/h, t1/2 = 90.18 +/- 168.44 h; healthy volunteers--CL = 8.10 +/- 4.58 l/h, t1/2 = 8.60 +/- 4.12 h (p < 0.05). Statistically significant correlation was found between the metabolic ratio and CL value (rs = 0.83) or between half-life (rs = -0.72). Similar results were also obtained using the two-point method. CONCLUSION: In patients with liver cirrhosis we found significantly lower paraxanthine/caffeine ratio which correlates with lowered elimination of caffeine. Its evaluation enables non-invasive assessment of liver metabolic function from a single sample of saliva. PMID- 11262909 TI - [Ethics of working with DNA in health care]. AB - The reasoning on DNA as a biological sample of the particular status is based on the author's experience with DNA isolation, analysis and interpretation of genotyping results. The author discusses the nature of the information contained in DNA molecule, he specifies who uses this information and in what way the information is or should be used. The different standpoints concerning the ethical codex of patient's rights versus DNA handling are stressed. PMID- 11262910 TI - [Palliative therapy of bone metastases using radiopharmaceuticals. Recommendations for diagnostic and therapeutic care in nuclear medicine]. AB - The paper is presented as one of recommendations of diagnostic and therapeutic care in the field of nuclear medicine intended especially for the clinical needs of physicians indicating palliative treatment of metastases into bones using radiopharmaceuticals. It provides a brief review of principles, indications, the very performance of interventions, clinical interpretations, possible risks and contraindications including the clinical and social-economic meaning. PMID- 11262911 TI - [Historic medical anniversaries in 2001]. PMID- 11262912 TI - [Does moderate alcohol drinking decrease the incidence and mortality rate in ischemic heart disease?]. AB - Moderate drinking of alcohol decreases the progress of atherosclerosis, cardiovascular morbidity and mortality rate and total mortality. The mechanisms of action have not been clarified yet, but changes of lipid metabolism, antioxidative effect and changes in hemostasis are accused to play the major role. Moderate drinking leads to the increase of HDL cholesterol and decrease of LDL cholesterol. Antioxidants are distributed throughout the skin of grapes and therefore are present in higher concentration in red wine. Alcohol decreases the fibrinogen level and increases tPA, inhibits platelet aggregation and reduces factor VIIc. It positively influences stress, fear, anxiety and depression. Optimal daily consumption should be 20 to 40 g in men and half of it in women. Everyday drinking is important. There is no big difference between drinking beer, wine or drinks. The most crucial is to keep the moderate level of consumption. PMID- 11262913 TI - [Care of the diseased life]. AB - Care of the diseased patients is an integral part of the clinical intervention centred on the benefit of the patient. Author defines the conception of "care" as a component of medical activities. The characteristic features of the care and nursing are given. The specific problems of the providers of the care and those who accept it are discussed. Major differences between the cure (therapy) and care (nursing) and their mutual relations are considered. The compensatory character of nursing in situations when the therapy is ineffective is stressed. PMID- 11262914 TI - [Radiosurgical treatment of hypophyseal adenomas with the gamma knife: results in a group of 163 patients during a 5-year period]. AB - BACKGROUND: Gamma knife radiosurgery of pituitary adenomas is considered to be very perspective. It can be a very useful complement of traditional microsurgery, pharmacotherapy or fractionated radiotherapy which are seldom a sufficient treatment on their own. The modern radiosurgery does not offer the experience representative enough in this indication. We can offer results of medium long follow-up for tumor growth and hormonal hypersecretion of pituitary adenomas in a relatively large series of patients. METHODS AND RESULTS: We have analyzed a group of 163 patients with pituitary adenoma treated with gamma knife during 5 years and followed 12-60 months, median 24 months after irradiation. An antiproliferative effect has been achieved in 1-2 years using the minimal dose to the margin 16-35 Gy, median 20 Gy in all our patients who were controlled by MRI (n = 126 patients). One half of these adenomas evidently decreased their size. Our effective antiproliferative dose was safe for the surrounding structures. The hormonal normalization has been achieved at 50.4% from 133 hypersecreting adenomas (39/91 = 43% of acromegalics, 11/13 = 85% of patients with Cushing's disease, 2/9 = 22% of patients with Nelson's syndrome, 11/18 = 61% of prolactinomas). The median latency was 12 months. The minimal dose to the margin was 10-45 Gy, median 35 Gy. Rare side effects were provoked only by increasing the dose to influence the hypersecretion-the development of partial hypopituitarism in 3.1% of patients, the panhypopituitarism in 0.6% of patient and there was 1 hemianopic visual field defect (0.6%). CONCLUSIONS: Radiosurgery by gamma knife has a similar value for pituitary adenomas as microsurgery has with different distribution of advantages and drawbacks. This makes it suitable for the combined treatment where pharmacotherapy has its place under special conditions. Fractionated radiotherapy has now a marginal importance. PMID- 11262915 TI - [Surgical treatment of recurrent urethral strictures in males after unsuccessful operations]. AB - BACKGROUND: Subvesical obstructions of any origin represent a frequent and serious disorder occurring predominantly in males. Often it brings incontinence and/or erectility dysfunction, infection of urinary tract. Relapses of the acute pyelonephritis can turn into chronic tubulointersticial one and terminate in the renal insufficiency. To treat strictures, dilation, intermittent catheterization and recently stent introduction were used. Most suitable appears a stent from composite polymers. The aim of our work was to test properties of stents developed in the Institute of Macromolecular Chemistry ASCR. METHODS AND RESULTS: Stents from composite polymers, which are non-toxic, not-irritable can swell in body fluids and have mechanical properties similar to that of silicone rubber. Properties of the material are functionally graded and the casting or repousse from the material can subsequently change its shape. Ten patients (males, aged 25 to 78 years) with long urethral strictures in its bulbocavernous part (50%) were treated with this method. Strictures were caused by pelvical fractures (4 times), prostate hypertrophy surgery (4 times), prolonged catheterizations (2 times). All patients were followed for 16 to 26 month and had no severities. CONCLUSION: Our results indicate that stent from composite polymers and silicone may have long acting effects without irritation or crust formation and beneficially effected healing of the spongio-fibrous process. PMID- 11262917 TI - [Diagnosis of Helicobacter pylori with the 13C-labeled urea breath test: study methodology]. AB - Helicobacter pylori is one of the most common causes of chronic bacterial infection in humans, and it is associated with many diseases of the upper gastrointestinal tract. The 13C urea breath test (13C-UBT) is a simple, non invasive and global test for Helicobacter pylori detection. The test reflects the hydrolysis of 13C-labelled urea by Helicobacter pylori urease. The 13C-UBT is the gold standard test for Helicobacter pylori infection. Since the original description (in 1987) several modifications of 13C-UBT have been published to simplify and optimise the test. However, neither Standardised European Protocol nor Standard US Protocol were accepted. This paper gives the methodology of the 13C-UBT based on eur own study and on the review of the literature. PMID- 11262916 TI - [Derivatives of dehydroepiandrosterone and their changes during a one-day starvation test]. AB - BACKGROUND: Relationship between dehydroepiandrosterone (DHEA), its sulphate (DHEAS) and various components of metabolic syndrome X have been recently discussed in several papers. Originally only DHEA or DHEAS have been considered to be responsible for all of the effects. At present mainly DHEA hydroxyderivatives (particularly 7-hydroxyisometers) are assumed to be responsible for those effects. METHODS AND RESULTS: 68 obese subjects (28 males) aged 42.6 +/- 11.1 years, with average BMI 35.7 +/- 11.6 kg/m2 were examined. Relationship between 7-alpha and 7-beta-(OH)-DHEA and various components of metabolic syndrome X have been followed in a pilot epidemiological study. Fluctuations of the hydroxyderivates level during one-day starvation test were investigated in a group of 11 obese females and compared with that of the control group (12 lean subjects with BMI 23.1 +/- 2.4 kg/m2). From the view of the metabolic syndrome X, the negative correlation between the serum levels of 7-beta (OH)-DHEA and insulinemia (r = -0.28; p = 0.23), glycemia (r = -0.48; p < 0.001), serum level of uric acid (r = -0.35; p = 0.02) and opposite the positive correlation between the serum level of HDL-cholesterol (r = 0.42; p < 0.01) should be pointed out. The negative correlation between 7-alpha-(OH)-DHEA and age and BMI was noticed (correlation for 7-beta-(OH)-DHEA was similar). No other statistically significant correlation was found among the other monitored parameters. In 11 obese females the dynamic changes of the above-mentioned DHEA hydroxyderivatives during one-day starvation test were monitored. Changes were compared with the control group (12 lean females). Significant increases in DHEAS, 7-beta-(OH)-DHEA, and sex hormone binding globulin (SHBG) levels were observed during this test. Significant differences in dynamic changes (before and after the test) between obese and lean group have been found only in DHEAS and SHBG. CONCLUSION: We suppose that 7-beta-(OH)-DHEA (more than 7-alpha-(OH)-DHEA) is specifically related to the metabolic syndrome X and that its claimed anti glucocorticoid effect in the immune response can play some role in its metabolic effects. PMID- 11262918 TI - The ponderosa pine ecosystem and environmental stress: past, present and future. PMID- 11262919 TI - Changes in physiological attributes of ponderosa pine from seedling to mature tree. AB - Plant physiological models are generally parameterized from many different sources of data, including chamber experiments and plantations, from seedlings to mature trees. We obtained a comprehensive data set for a natural stand of ponderosa pine (Pinus ponderosa Laws.) and used these data to parameterize the physiologically based model, TREGRO. Representative trees of each of five tree age classes were selected based on population means of morphological, physiological, and nearest neighbor attributes. Differences in key physiological attributes (gas exchange, needle chemistry, elongation growth, needle retention) among the tree age classes were tested. Whole-tree biomass and allocation were determined for seedlings, saplings, and pole-sized trees. Seasonal maxima and minima of gas exchange were similar across all tree age classes. Seasonal minima and a shift to more efficient water use were reached one month earlier in seedlings than in older trees because of decreased soil water availability in the rooting zone of the seedlings. However, carbon isotopic discrimination of needle cellulose indicated increased water-use efficiency with increasing tree age. Seedlings had the lowest needle and branch elongation biomass growth. The amount of needle elongation growth was highest for mature trees and amount of branch elongation growth was highest for saplings. Seedlings had the highest biomass allocation to roots, saplings had the highest allocation to foliage, and pole sized trees had the highest allocation to woody tissues. Seedlings differed significantly from pole-sized and older trees in most of the physiological traits tested. Predicted changes in biomass with tree age, simulated with the model TREGRO, closely matched those of trees in a natural stand to 30 years of age. PMID- 11262920 TI - Use of a simulation model and ecosystem flux data to examine carbon-water interactions in ponderosa pine. AB - Drought stress plays an important role in determining both the structure and function of forest ecosystems, because of the close association between the carbon (C) and hydrological cycles. We used a detailed model of the soil-plant atmosphere continuum to investigate the links between carbon uptake and the hydrological cycle in a mature, open stand of ponderosa pine (Pinus ponderosa Dougl. ex Laws.) at the Metolius river in eastern Oregon over a 2-year period (1996-1997). The model was parameterized from local measurements of vegetation structure, soil properties and meteorology, and tested against independent measurements of ecosystem latent energy (LE) and carbon fluxes and soil water content. Although the 2 years had very different precipitation regimes, annual uptake of C and total transpiration were similar in both years, according to both direct observation and simulations. There were important differences in ratios of evaporation to transpiration, and in the patterns of water abstraction from the soil profile, depending on the frequency of summer storms. Simulations showed that, during periods of maximum water limitation in late summer, plants maintained a remarkably constant evapotranspirative flux because of deep rooting, whereas changes in rates of C accumulation were determined by interactions between atmospheric vapor pressure deficit and stomatal conductance. Sensitivity analyses with the model suggest a highly conservative allocation strategy in the vegetation, focused belowground on accessing a soil volume large enough to buffer summer droughts, and optimized to account for interannual variability in precipitation. The model suggests that increased allocation to leaf area would greatly increase productivity, but with the associated risk of greater soil water depletion and drought stress in some years. By constructing sparse canopies and deep rooting systems, these stands balance reduced productivity in the short term with risk avoidance over the long term. PMID- 11262922 TI - Ecosystem respiration in a young ponderosa pine plantation in the Sierra Nevada Mountains, California. AB - We estimated total ecosystem respiration from a ponderosa pine (Pinus ponderosa Dougl. ex Laws.) plantation in the Sierra Nevada Mountains near Georgetown, California, from June to October, 1998. We apportioned ecosystem respiration among heterotrophic, root, stem and foliage based on relationships for each component that considered microclimate and vegetation characteristics. We measured each respiration component at selected sampling points, and scaled the measurements up to the ecosystem based on modeled relationships. Over the study period, total mean ecosystem respiration was 5.7 +/- 1.3 mumol m-2 s-1 (based on daily mean), comprising about 67% from soil-surface CO2 efflux, 10% from stem and branch respiration and 23% from foliage respiration. Shrub leaves contributed about 24% to total foliage respiration, and current-year needles (1998 age class) accounted for 40% of total tree needle respiration. Root respiration accounted for 47% of soil-surface CO2 efflux. We conclude that ecosystem respiration can be estimated based on daily mean air and soil temperatures through exponential relationships with r2 values of 0.85 and 0.87, respectively. When based on both air and soil temperatures, about 91% of the variation in total ecosystem respiration could be explained by a linear regression. PMID- 11262921 TI - Carbon dioxide and water vapor exchange by young and old ponderosa pine ecosystems during a dry summer. AB - We investigated key factors controlling mass and energy exchange by a young (6 year-old) ponderosa pine (Pinus ponderosa Laws.) plantation on the west side of the Sierra Nevada Mountains and an old-growth ponderosa pine forest (mix of 45- and 250-year-old trees) on the east side of the Cascade Mountains, from June through September 1997. At both sites, we operated eddy covariance systems above the canopy to measure net ecosystem exchange of carbon dioxide and water vapor, and made concurrent meteorological and ecophysiological measurements. Our objective was to understand and compare the controls on ecosystem processes in these two forests. Precipitation is much higher in the young plantation than in the old-growth forest (1660 versus 550 mm year-1), although both forests experienced decreasing soil water availability and increasing vapor pressure deficits (D) as the summer of 1997 progressed. As a result, drought stress increased at both sites during this period, and changes in D strongly influenced ecosystem conductance and net carbon uptake. Ecosystem conductance for a given D was higher in the young pine plantation than in the old-growth forest, but decreased dramatically following several days of high D in late summer, possibly because of xylem cavitation. Net CO2 exchange generally decreased with conductance at both sites, although values were roughly twice as high at the young site. Simulations with the 3-PG model, which included the effect of tree age on fluxes, suggest that, during the fall through spring period, milder temperatures and ample water availability at the young site provide better conditions for photosynthesis than at the old pine site. Thus, over the long term, the young site can carry more leaf area, and the climatic conditions between fall and spring offset the more severe limitations imposed by summer drought. PMID- 11262923 TI - Blue wild-rye grass competition increases the effect of ozone on ponderosa pine seedlings. AB - Individual ponderosa pine (Pinus ponderosa Dougl. ex Laws.) seedlings were grown in mesocosms with three densities of blue wild-rye grass (Elymus glaucus Buckl.) (equivalent to 0, 32 or 88 plants m-2) to determine if the presence of a natural competitor alters the response of ponderosa pine seedlings to ozone. After 3 years of ozone exposure, grass presence reduced total ponderosa pine dry mass by nearly 50%, whereas ozone alone had no significant effect on ponderosa pine growth. The combination of ozone and grass further reduced needle, stem and branch dry mass significantly below that induced by grass competition alone. Root:shoot ratios increased in response to the combined grass and ozone treatments. Grass competition significantly reduced soluble sugar concentrations in all ponderosa pine tissue components examined. Starch concentrations were highly variable but did not differ significantly between treatments. Ozone significantly reduced soluble sugar concentrations in fine roots and stems. In the absence of grass, ozone-treated seedlings tended to have higher tissue N concentrations than controls. In the presence of grass, ozone-treated seedlings had lower N concentrations than controls, resulting in a significant interaction between these two stresses in 1- and 2-year-old needles. Needle C:N ratios decreased in response to grass competition, as a result of increased N concentration and no change in C concentration. The opposite response was observed in ozone-treated seedlings as a result of decreased N concentrations, indicating that ozone-treated seedlings were unable to take up or retain as much nitrogen when grown in the presence of grass. We conclude that ponderosa pine seedlings are more susceptible to ozone when grown in competition with blue wild rye grass. PMID- 11262924 TI - Multivariate patterns of biochemical responses of Pinus ponderosa trees at field plots in the San Bernardino Mountains, southern California. AB - Most environmental stress conditions promote the production of potentially toxic active oxygen species in plant cells. Plants respond with changes in their antioxidant and photoprotective systems. Antioxidants and pigments have been widely used to measure these responses. Because trees are exposed to multiple man made and natural stresses, their responses are not reflected by changes in single stress markers, but by complex biochemical changes. To evaluate such response patterns, explorative multivariate statistics have been used. In the present study, 12 biochemical variables (chloroplast pigments, state of the xanthophyll cycle, alpha-tocopherol, ascorbate and dehydroascorbate, glutathione and oxidized glutathione) were measured in previous-year needles of field-grown Pinus ponderosa Dougl. ex Laws. The trees were sampled in two consecutive years in the San Bernardino Mountains in southern California, where a pollution gradient is overlaid by gradients in natural stresses (drought, altitude). To explore irradiance effects, needle samples were taken directly in the field (sun exposed) and from detached, dark-adapted branches. A principal component analysis on this data set (n = 80) resulted in four components (Components 1-4) that explained 67% of the variance in the original data. Component 1 was positively loaded by concentrations of alpha-tocopherol, total ascorbate and xanthophyll cycle pools, as well as by the proportion of de-epoxides in the xanthophyll cycle. It was negatively loaded by the proportion of dehydroascorbate in the ascorbate pool. Component 2 was negatively loaded by chlorophyll concentrations, and positively loaded by the ratios of lutein and beta-carotene to chlorophyll and by the de epoxidation state of the xanthophyll cycle. Component 3 was negatively loaded by GSH concentrations and positively loaded by the proportions of GSSG and tocopherol concentrations. Component 4 was positively loaded by neoxanthin and negatively loaded by beta-carotene. The four components could be assigned to the concerted action of the biochemical protection system: high scores on Component 1 represent highly activated antioxidative defense, changes in pigment composition are represented in Components 2 and 4, and the glutathione system, which is important for antioxidant regeneration, is represented in Component 2. Although Component 1 scores were generally higher (indicating activation of antioxidant defense) in light-adapted needles relative to dark-adapted needles, they were also site dependent with increased scores at sites with less pollution, but higher natural stress impacts. High scores of Components 2 and 3 at the highest elevation site, which was only moderately polluted, indicated an increase in photoprotection by pigments and activation of the glutathione system. Significant differences between light- and dark-adapted needles in Components 2 and 3 were only found at the site with the highest pollution. Use of accumulated variables (components) instead of single biochemical variables enabled recognition of response patterns at particular sites and a better comparison with results of other studies is expected. Typical response patterns could be assigned to particular environmental stress combinations, providing a means of assessing potential biological risks within individual forest stands. PMID- 11262925 TI - Response of stomatal conductance to drought in ponderosa pine: implications for carbon and ozone uptake. AB - To gain insight into the limitations imposed by a typical Mediterranean-climate summer drought on the uptake of carbon and ozone in the ponderosa pine (Pinus ponderosa Dougl. ex Laws.) ecosystem, we compared diurnal trends in leaf physiology of young trees in a watered and a control plot located in the Sierra Nevada Mountains, CA, USA (Blodgett Forest, 38 degrees 53' N, 120 degrees 37' W, 1315 m elevation). Predawn water potential of trees in the watered plot remained above -0.3 MPa throughout the growing season, whereas it dropped in the control plot from -0.24 to -0.52 MPa between late May and mid-August. Photosynthesis and stomatal conductance of trees in the watered plot were relatively insensitive to atmospheric vapor pressure deficit (VPD), whereas gas exchange of trees in the control plot varied with changes in soil water, VPD and temperature. Although the 1998 growing season was abnormally wet, we saw a pronounced drought effect at the control site. Over the 2 months following the onset of watering, carbon and ozone uptake were measured on three days at widely spaced intervals. Carbon uptake per unit leaf area by 1-year-old foliage of trees in the control plot was 39, 35 and 30% less, respectively, than in the watered plot, and estimated ozone deposition per unit leaf area (ozone concentration times stomatal conductance) was 36, 46 and 41% less. PMID- 11262927 TI - Strategic planning: going beyond the basics. AB - Boards are discovering that many of the strategies of the '90s that took their organizations far afield from their core mission did not pan out as intended. Now they need to take a new strategic approach--combining the basics with a view toward a dynamic health care environment. PMID- 11262926 TI - The paper monster. Corporate compliance and the board. AB - Fraud and abuse lawsuits and settlements are getting bigger, and the DOJ shows no signs of letting up on its scrutiny of health care organizations. That's why a strong compliance program is your best defense, and it all starts with the board. PMID- 11262928 TI - Self-assessment: a powerful learning tool. PMID- 11262929 TI - Governance. Terminating board term limits. PMID- 11262930 TI - Technology. Hands-off care. PMID- 11262931 TI - How many hats are too many? AB - In rural communities, where the pool of leaders is limited, boards often struggle with conflicts of interest. Trustees and governance experts offer tips on how to prevent, or at least deal with, a potential problem that can undermine effective governance. PMID- 11262932 TI - Stress-induced DNA duplex destabilization in transcriptional initiation. AB - Stress-induced destabilization of the DNA double helix (SIDD) is involved in several mechanisms by which transcription is regulated. This paper describes a computational method for predicting the locations and extents of destabilization as functions of DNA sequence and imposed superhelical stress. This method is used to investigate several transcriptional regulatory events. These include IHF mediated activation of gene expression in E. coli, the bimodal control of the initiation of transcription from the human c-myc gene, and the determination of the minimal requirements for transcriptional activity in yeast. Collaborations with experimental groups have established the central role of SIDD in each of these processes. PMID- 11262933 TI - SAMIE: statistical algorithm for modeling interaction energies. AB - We are investigating the rules that govern protein-DNA interactions, using a statistical mechanics based formalism that is related to the Boltzmann Machine of the neural net literature. Our approach is data-driven, in which probabilistic algorithms are used to model protein-DNA interactions, given SELEX and/or phage data as input. In the current report, we trained the network using SELEX data, under the "one-to-one" model of interactions (i.e. one amino acid contacts one base). The trained network was able to successfully identify the wild-type binding sites of EGR and MIG protein families. The predictions using our method are the same or better than that of methods existing in the literature. However our methodology offers the potential to capitalise in quantitative detail, as well as to be used to explore more general model of interactions, given availability of data. PMID- 11262934 TI - BioProspector: discovering conserved DNA motifs in upstream regulatory regions of co-expressed genes. AB - The development of genome sequencing and DNA microarray analysis of gene expression gives rise to the demand for data-mining tools. BioProspector, a C program using a Gibbs sampling strategy, examines the upstream region of genes in the same gene expression pattern group and looks for regulatory sequence motifs. BioProspector uses zero to third-order Markov background models whose parameters are either given by the user or estimated from a specified sequence file. The significance of each motif found is judged based on a motif score distribution estimated by a Monte Carlo method. In addition, BioProspector modifies the motif model used in the earlier Gibbs samplers to allow for the modeling of gapped motifs and motifs with palindromic patterns. All these modifications greatly improve the performance of the program. Although testing and development are still in progress, the program has shown preliminary success in finding the binding motifs for Saccharomyces cerevisiae RAP1, Bacillus subtilis RNA polymerase, and Escherichia coli CRP. We are currently working on combining BioProspector with a clustering program to explore gene expression networks and regulatory mechanisms. PMID- 11262935 TI - A structure-based approach for prediction of protein binding sites in gene upstream regions. AB - The challenge of identifying DNA regulatory sequences based on sequence information only has been emphasized in view of the fast accumulation of new genes in the databases. While most predictive algorithms are based on multiple alignments of already known binding sites, here we examine the usefulness of a novel approach that is based on structural information of the protein-DNA complex. It has already been shown that specific recognition between a protein and its DNA target is achieved by stereo-chemical complementarity between the protein amino acids and the DNA bases. The proposed computational scheme uses crystallographic information to define the set of amino acid-base contacts between the proteins of a given DNA-binding protein family and their DNA targets. The compatibility of a given protein to bind to putative regulatory DNA sequences is then evaluated by knowledge-based parameters for amino acid-base interactions. By this procedure gene upstream regions may be screened for potential binding sites for regulatory proteins. Predictions are demonstrated for the E. coli cyclic AMP receptor protein (CRP) which recognizes the DNA via the helix-turn helix motif, and for various Zif268-like proteins which belong to the Cys2His2 zinc finger family. The advantages and limitations of this approach are discussed. PMID- 11262936 TI - Promoter region-based classification of genes. AB - In this paper we consider the problem of extracting information from the upstream untranslated regions of genes to make predictions about their transcriptional regulation. We present a method for classifying genes based on motif-based hidden Markov models (HMMs) of their promoter regions. Sequence motifs discovered in yeast promoters are used to construct HMMs that include parameters describing the number and relative locations of motifs within each sequence. Each model provides a Fisher kernel for a support vector machine, which can be used to predict the classifications of unannotated promoters. We demonstrate this method on two classes of genes from the budding yeast, S. cerevisiae. Our results suggest that the additional sequence features captured by the HMM assist in correctly classifying promoters. PMID- 11262937 TI - Nucleotide substitutions and the evolution of duplicate genes. AB - This paper describes software created to search for and analyze pairs of duplicate genes within a genome. The process is based on a program that uses aligned amino acid sequences to generate a corresponding alignment of the underlying nucleotide sequences and perform a codon by codon comparison of the nucleotides. Observed numbers of nucleotide substitutions can be used to make inferences about the ages of gene duplication events and the effects of natural selection acting on duplicate genes. PMID- 11262938 TI - An algorithm for statistical alignment of sequences related by a binary tree. AB - An algorithm is presented that allows the calculation of the probability of a set of sequences related by a binary tree that has evolved according to the Thorne Kishino-Felsenstein model (1991) for a fixed set of parameters. There are two ideas underlying this algorithm. Firstly, a markov chain is defined that generates ancestral sequences and their alignment at two neighboring nodes in a tree. Secondly, a stochastic walk on the binary tree, that defines a markov chain generating ancestral sequences and their alignment at the internal nodes in the tree is described. The running time of this algorithm is O(l2 kappa), where l is the geometric average of the sequence lengths and kappa the number of sequences- leaves at the binary tree. This could be improved to O(l kappa). PMID- 11262940 TI - Analyzing site heterogeneity during protein evolution. AB - New computational models of the kinetics of natural site substitutions in proteins are described based on the underlying physical chemical properties of the amino acids. The corresponding reduction in the number of adjustable parameters allows us to analyze site-heterogeneity. Applying this evolutionary model to various data sets allows us to identify the important factors constraining molecular evolution, providing insight into the relationship between amino acid properties and protein structure. PMID- 11262939 TI - Assessment and management of single nucleotide polymorphism genotype errors in genetic association analysis. AB - Single nucleotide polymorphisms (SNP) may be used in case-control designs to test for association between a marker (the SNP) and a disease. However, such designs usually assume that the genotype data are reported without error. We propose a method, the reduced penetrance model method (RPM) that allows for errors in a case-control design, as compared to the full penetrance model method (FPM), that assumes data are errorless. Pearson's chi 2 applied to a 2 x 2 contingency table is the test statistic considered. Additionally, we provide a likelihood method to estimate error rates using SNP genotype data in CEPH pedigrees. We test our method (RPM) against the standard method (FPM) using simulated data. All SNP loci are assumed to have two alleles, coded 1 and 2. We consider three pairs of error rates, two different sample sizes, and two sets of allele frequencies for the SNP locus. SNP genotype data in two populations are simulated under a null hypothesis (allele frequencies equal in both populations) and under an alternative hypothesis (allele frequencies differ between two populations). The total number of simulations is 24; 12 simulations under the null hypothesis, and 12 simulations under the alternative. The significance level threshold is 5%. For the null case, 9/12 (75%) of the simulations show no increase in type I error under RPM, while 3/12 (25%) show a slight increase (rejecting the null for at most 7% of the replicates). There is no increase in the type I error rate for FPM method, which can also be shown analytically. For the alternative case (power), there is a consistent increase in power for the RPM method as compared to FPM method, and average increase of 0.02 for the simulations considered. When sample sizes are large there is virtually no difference in power between RPM and FPM methods. Also, the RPM method provides consistently more accurate allele frequency estimates for the various populations. Our likelihood method to estimate error rates with CEPH pedigrees provides good estimates on average. The largest difference between a true error rate and our average estimated error rate is 0.006. However, there is a fair amount of variability in the estimates, suggesting the need for multiple experiments or larger numbers of CEPH pedigrees. Researchers may use the methods presented in this paper to (1) estimate error rates for their automated genotyping process, and (2) allow for such errors in association analyses, thereby increasing power to detect differences between allele frequencies in case and control populations when errors are present. PMID- 11262941 TI - Markov chain Monte Carlo computation of confidence intervals for substitution rate variation in proteins. AB - We suggest a method implemented in a computer program, immodestly dubbed TSUNAMI, that allows us to compare two homologous protein subfamilies with respect to the distribution of substitution rates along sequences. This study furthers our earlier work on a wavelet model of rate variation (1). The current approach allows sensitive detection of subtle discordances in the selection patterns between two protein subfamilies. In addition to performing fast computation of the maximum posterior probability estimates of the relative substitution rates, the method can select the most appropriate number of wavelet parameters for a particular dataset. TSUNAMI is based on a Markov chain Monte Carlo technique, and appears to be more applicable to larger datasets than is the full likelihood based approach. PMID- 11262942 TI - A model for phylogenetic inference using structural and chemical covariates. AB - We investigated whether or not evolutionary change in DNA sequence data was homogeneous across different classes of base pairs. DNA sequences for eight protein-coding mitochrondrial genes were obtained for 38 vertebrate taxa from GenBank. Each nucleotide site in the alignment was classified according to a number of covariates, including its codon position, genetic code degeneracy, and hydrophobicity. The evolutionary transition matrix for each base was estimated by tracing implied character changes under parsimony on a known phylogenetic tree. Canonical variates analyses of the inferred transition matrices were performed for each gene to determine whether or not different classes of bases behaved similarly. We found five distinct clusters of transition matrices that could be roughly defined by combinations of codon position and degeneracy. This pattern was consistent among all genes. A stochastic model of rate variation based on the interaction of the covariates was developed to assess the statistical significance of the clusters. The five-group classification was found to explain significantly more sequence variation than did a codon only classification, a codon degeneracy classification, or a codon and degeneracy classification. The same five-group classification was found for all genes tested, suggesting a common process underlying the molecular evolution of the mitochondrial genome. These results confirm that there are classes of base pairs that evolve differently, and suggest that models of sequence evolution that incorporate covariate information may be useful in developing nucleotide substitution models that more accurately reflect evolutionary history. PMID- 11262943 TI - Maximum likelihood analysis of adaptive evolution in HIV-1 gp120 env gene. AB - Every functional protein appears to have some conserved amino acids which are critically important to the basic structure and function of the protein and thus under purifying selection. Some proteins also have variable amino acids which, when changed, offer a selective advantage, and thus undergo adaptive evolution. These amino acids are also important to the structure and function of the protein, although in a different way. It seems that the selective pressure in every protein varies among sites. Maximum likelihood models developed recently for comparison of silent and replacement nucleotide substitution rates allow for variable selective pressures among amino acid sites, and provide a powerful approach to studying the evolutionary process of protein-coding genes. This paper applies the likelihood models to analyze a data set of 186 HIV-1 gp120 env gene sequences for comparison with a previous analysis of the same data set. The maximum likelihood analysis identified a number of sites under positive selection, some in the conserved regions of the protein. PMID- 11262944 TI - ViewFeature: integrated feature analysis and visualization. AB - Visualization interfaces for high performance computing systems pose special problems due to the complexity and volume of data these systems manipulate. In the post-genomic era, scientists must be able to quickly gain insight into structure-function problems, and require flexible computing environments to quickly create interfaces that link the relevant tools. Feature, a program for analyzing protein sites, takes a set of 3-dimensional structures and creates statistical models of sites of structural or functional significance. Until now, Feature has provided no support for visualization, which can make understanding its results difficult. We have developed an extension to the molecular visualization program Chimera that integrates Feature's statistical models and site predictions with 3-dimensional structures viewed in Chimera. We call this extension ViewFeature, and it is designed to help users understand the structural Features that define a site of interest. We applied ViewFeature in an analysis of the enolase superfamily; a functionally distinct class of proteins that share a common fold, the alpha/beta barrel, in order to gain a more complete understanding of the conserved physical properties of this superfamily. In particular, we wanted to define the structural determinants that distinguish the enolase superfamily active site scaffold from other alpha/beta barrel superfamilies and particularly from other metal-binding alpha/beta barrel proteins. Through the use of ViewFeature, we have found that the C-terminal domain of the enolase superfamily does not differ at the scaffold level from metal-binding alpha/beta barrels. We are, however, able to differentiate between the metal-binding sites of alpha/beta barrels and those of other metal-binding proteins. We describe the overall architectural Features of enolases in a radius of 10 Angstroms around the active site. PMID- 11262945 TI - Analyzing sensory systems with the information distortion function. AB - The nature and information content of neural signals have been discussed extensively in the neuroscience community. They are important ingredients in many theories on neural function, yet there is still no agreement on the details of neural coding. There have been various suggestions about how information is encoded in neural spike trains: by the number of spikes, by temporal correlations, through single spikes, or by spike patterns in one, or across many neurons. The latter scheme is most general and encompasses many others. We present an algorithm which can recover a coarse representation of a pattern coding scheme, through quantization to a reproduction set of smaller size. Among many possible quantizations, we choose one which preserves as much of the informativeness of the original stimulus/response relation as possible, through the use of an information-based distortion function. This method allows us to study coarse but highly informative models of a coding scheme, and then to refine them when more data becomes available. We shall describe a model in which full recovery is possible and present example for cases with partial recovery. PMID- 11262946 TI - Sequence analysis with the Kestrel SIMD parallel processor. AB - Computer aided sequence analysis is a critical aspect of current biological research. Sequence information from the genome sequencing projects fills databases so quickly that humans cannot examine it all. Hence there is a heavy reliance on computer algorithms to point out the few important nuggets for human examination. Sequence search algorithms range from simple to complex, as does the representation of the biological data. Typically though, simple algorithms are used on the simplest of data representations because of the large computational demands of anything more complex. This leads to missed hits because the simple search techniques are often not sufficiently sensitive. Here we describe the implementation of several sensitive sequence analysis algorithms on the Kestrel parallel processor, a single-instruction multiple-data (SIMD) processor developed and built at UCSC. Performance of the Smith-Waterman and Hidden Markov Model algorithms, with both Viterbi and Expectation Maximization methods ranges from 6 to 20 times faster than standard computers. PMID- 11262947 TI - A new algorithm for the alignment of multiple protein structures using Monte Carlo optimization. AB - We have developed a new algorithm for the alignment of multiple protein structures based on a Monte Carlo optimization technique. The algorithm uses pair wise structural alignments as a starting point. Four different types of moves were designed to generate random changes in the alignment. A distance-based score is calculated for each trial move and moves are accepted or rejected based on the improvement in the alignment score until the alignment is converged. Initial tests on 66 protein structural families show promising results, the score increases by 69% on average. The increase in score is accompanied by an increase (12%) in the number of residue positions incorporated into the alignment. Two specific families, protein kinases and aspartic proteinases were tested and compared against curated alignments from HOMSTRAD and manual alignments. This algorithm has improved the overall number of aligned residues while preserving key catalytic residues. Further refinement of the method and its application to generate multiple alignments for all protein families in the PDB, is currently in progress. PMID- 11262948 TI - Gene verification and discovery by Walking Tree Method. AB - The Walking Tree Method [3, 4, 5, 18] is an approximate string alignment method that can handle insertions, deletions, substitutions, translocations, and more than one level of inversions all together. Moreover, it tends to highlight gene locations, and helps discover unknown genes. Its recent improvements in runtime and space use extends its capability in exploring large strings. We will briefly describe the Walking Tree Method with its recent improvements [18], and demonstrate its speed and ability to align real complete genomes such as Borrelia burgdorferi (910724 base pairs of its single chromosome) and Chlamydia trachomatis (1042519 base pairs) in reasonable time, and to locate and verify genes. PMID- 11262949 TI - Predicting gene function from gene expressions and ontologies. AB - We introduce a methodology for inducing predictive rule models for functional classification of gene expressions from microarray hybridisation experiments. The basic learning method is the rough set framework for rule induction. The methodology is different from the commonly used unsupervised clustering approaches in that it exploits background knowledge of gene function in a supervised manner. Genes are annotated using Ashburner's Gene Ontology and the functional classes used for learning are mined from these annotations. From the original expression data, we extract a set of biologically meaningful features that are used for learning. A rule model is induced from the data described in terms of these features. Its predictive quality is fine-turned via cross validation on subsets of the known genes prior to classification of unknown genes. The predictive and descriptive quality of such a rule model is demonstrated on the fibroblast serum response data previously analysed by Iyer et. al. Our analysis shows that the rules are capable of representing the complex relationship between gene expressions and function, and that it is possible to put forward high quality hypotheses about the function of unknown genes. PMID- 11262951 TI - A multithreaded parallel implementation of a dynamic programming algorithm for sequence comparison. AB - This paper discusses the issues involved in implementing a dynamic programming algorithm for biological sequence comparison on a general-purpose parallel computing platform based on a fine-grain event-driven multithreaded program execution model. Fine-grain multithreading permits efficient parallelism exploitation in this application both by taking advantage of asynchronous point to-point synchronizations and communication with low overheads and by effectively tolerating latency through the overlapping of computation and communication. We have implemented our scheme on EARTH, a fine-grain event-driven multithreaded execution and architecture model which has been ported to a number of parallel machines with off-the-shelf processors. Our experimental results show that the dynamic programming algorithm can be efficiently implemented on EARTH systems with high performance (e.g., speedup of 90 on 120 nodes), good programmability and reasonable cost. PMID- 11262950 TI - Unsupervised learning from complex data: the matrix incision tree algorithm. AB - Analysis of large-scale gene expression data requires novel methods for knowledge discovery and predictive model building as well as clustering. Organizing data into meaningful structures is one of the most fundamental modes of learning. DNA microarray data set can be viewed as a set of mutually associated genes in a high dimensional space. This paper describes a novel method to organize a complex high dimensional space into successive lower-dimensional spaces based on the geometric properties of the data structure in the absence of a priori knowledge. The matrix incision tree algorithm reveals the hierarchical structural organization of observed data by determining the successive hyperplanes that 'optimally' separate the data hyperspace. The algorithm was tested against published data sets yielding promising results. PMID- 11262952 TI - Molecular fingerprinting on the SIMD parallel processor Kestrel. AB - In combinatorial library design and use, the conformation space of molecules can be represented using three-dimensional (3-D) pharmacophores. For large libraries of flexible molecules, the calculation of these 3-D pharmacophoric fingerprints can require examination of trillions of pharmacophores, presenting a significant practical challenge. Here we describe the mapping of this problem to the UCSC Kestrel parallel processor, a single-instruction multiple-data (SIMD) processor. Data parallelism is achieved by simultaneous processing of multiple conformations and by careful representation of the fingerprint structure in the array. The resulting application achieved a 35+ speedup over an SGI 2000 processor on the prototype Kestrel board. PMID- 11262953 TI - Percolation clustering: a novel approach to the clustering of gene expression patterns in Dictyostelium development. AB - We present a novel approach to the clustering of gene expression patterns based on the mutual connectivity of the patterns. Unlike certain widely used methods (e.g., self-organizing maps and K-means) which essentially force gene expression data into a fixed number of predetermined clustering structures, our approach aims to reveal the natural tendency of the data to cluster, in analogy to the physical phenomenon of percolation. The approach is probabilistic in nature, and as such accommodates the possibility that one gene participates in multiple clusters. The result is cast in terms of the connectivity of each gene to a certain number of (significant) clusters. A computationally efficient algorithm is developed to implement our approach. Performance of the method is illustrated by clustering both constructed data and gene expression data obtained from Dictyostelium development. PMID- 11262954 TI - Quality control in manufacturing oligo arrays: a combinatorial design approach. AB - The advent of the DNA microarray technology has brought with it the exciting possibility of simultaneously observing the expression levels of all genes in an organism. One such microarray technology, called "oligo arrays", manufactures short single strands of DNA (called probes) onto a glass surface using photolithography. An altered or missed step in such a manufacturing protocol can adversely affect all probes using this failed step, and is in general impossible to disentangle from experimental variation when using such a defective array. The idea of designing special quality control probes to detect a failed step was first formulated by Hubbell and Pevzner. We consider an alternative formulation of this problem and use a combinatorial design approach to solve it. Our results improve over prior work in guaranteeing coverage of all protocol steps and in being able to tolerate a greater number of unreliable probe intensities. PMID- 11262955 TI - Using meta computing tools to facilitate large-scale analyses of biological databases. AB - Given the high rate at which biological data are being collected and made public, it is essential that computational tools be developed that are capable of efficiently accessing and analyzing these data. High-performance distributed computing resources can play a key role in enabling large-scale analyses of biological databases. We use a distributed computing environment, Legion, to enable large-scale computations on the Protein Data Bank (PDB). In particular, we employ the Feature program to scan all protein structures in the PDB in search for unrecognized potential cation binding sites. We evaluate the efficiency of Legion's parallel execution capabilities and analyze the initial biological implications that result from having a site annotation scan of the entire PDB. We discuss four interesting proteins with unannotated, high-scoring candidate cation binding sites. PMID- 11262957 TI - Textquest: document clustering of Medline abstracts for concept discovery in molecular biology. AB - We present an algorithm for large-scale document clustering of biological text, obtained from Medline abstracts. The algorithm is based on statistical treatment of terms, stemming, the idea of a 'go-list', unsupervised machine learning and graph layout optimization. The method is flexible and robust, controlled by a small number of parameter values. Experiments show that the resulting document clusters are meaningful as assessed by cluster-specific terms. Despite the statistical nature of the approach, with minimal semantic analysis, the terms provide a shallow description of the document corpus and support concept discovery. PMID- 11262956 TI - Including biological literature improves homology search. AB - Annotating the tremendous amount of sequence information being generated requires accurate automated methods for recognizing homology. Although sequence similarity is only one of many indicators of evolutionary homology, it is often the only one used. Here we find that supplementing sequence similarity with information from biomedical literature is successful in increasing the accuracy of homology search results. We modified the PSI-BLAST algorithm to use literature similarity in each iteration of its database search. The modified algorithm is evaluated and compared to standard PSI-BLAST in searching for homologous proteins. The performance of the modified algorithm achieved 32% recall with 95% precision, while the original one achieved 33% recall with 84% precision; the literature similarity requirement preserved the sensitive characteristic of the PSI-BLAST algorithm while improving the precision. PMID- 11262958 TI - Bidirectional incremental parsing for automatic pathway identification with combinatory categorial grammar. AB - As the importance of automatically extracting and analyzing various natural language assertions about protein-protein interactions in biomedical publications is recognized, many uses of natural language processing techniques are proposed in the literature. However, most proposals to date make rather simplifying assumptions about the syntactic aspects of natural language due to various reasons including efficiency. In this paper, we describe an implemented system that utilizes combinatory categorical grammar known to be competent in modeling natural language, with a controlled mechanism for the parser to operate bidirectionally and incrementally. We discuss the performance of the system on a large set of abstracts in Medline with quite encouraging results. PMID- 11262959 TI - Event extraction from biomedical papers using a full parser. AB - We have designed and implemented an information extraction system using a full parser to investigate the plausibility of full analysis of text using general purpose parser and grammar applied to biomedical domain. We partially solved the problems of full parsing of inefficiency, ambiguity, and low coverage by introducing the preprocessors, and proposed the use of modules that handles partial results of parsing for further improvement. Our approach makes it possible to modularize the system, so that the IE system as a whole becomes easy to be tuned to specific domains, and easy to be maintained and improved by incorporating various techniques of disambiguation, speed up, etc. In preliminary experiment, from 133 argument structures that should be extracted from 97 sentences, we obtained 23% uniquely and 24% with ambiguity. And 20% are extractable from not complete but partial results of full parsing. PMID- 11262960 TI - Mutual information analysis as a tool to assess the role of aneuploidy in the generation of cancer-associated differential gene expression patterns. AB - Most human tumors are characterized by: (1) an aberrant set of chromosomes, a state termed aneuploidy; (2) an aberrant gene expression pattern; and (3) an aberrant phenotype of uncontrolled growth. One of the goals of cancer research is to establish causative relationships between these three important characteristics. In this paper we were searching for evidence that aneuploidy is a major cause of differential gene expression. We describe how mutual information analysis of cancer-associated gene expression patterns could be exploited to answer this question. In addition to providing general guidelines, we have applied the proposed analysis to a recently published breast cancer-associated gene expression matrix. The results derived from this particular data set provided preliminary evidence that mutual information analysis may become a useful tool to investigate the link between differential gene expression and aneuploidy. PMID- 11262961 TI - Using graphical models and genomic expression data to statistically validate models of genetic regulatory networks. AB - We propose a model-driven approach for analyzing genomic expression data that permits genetic regulatory networks to be represented in a biologically interpretable computational form. Our models permit latent variables capturing unobserved factors, describe arbitrarily complex (more than pair-wise) relationships at varying levels of refinement, and can be scored rigorously against observational data. The models that we use are based on Bayesian networks and their extensions. As a demonstration of this approach, we utilize 52 genomes worth of Affymetrix GeneChip expression data to correctly differentiate between alternative hypotheses of the galactose regulatory network in S. cerevisiae. When we extend the graph semantics to permit annotated edges, we are able to score models describing relationships at a finer degree of specification. PMID- 11262962 TI - Reverse engineering of metabolic pathways from observed data using genetic programming. AB - Recent work has demonstrated that genetic programming is capable of automatically creating complex networks (such as analog electrical circuits and controllers) whose behavior is modeled by linear and non-linear continuous-time differential equations and whose behavior matches prespecified output values. The concentrations of substances participating in networks of chemical reactions are also modeled by non-linear continuous-time differential equations. This paper demonstrates that it is possible to automatically create (reverse engineer) a network of chemical reactions from observed time-domain data. Genetic programming starts with observed time-domain concentrations of input substances and automatically creates both the topology of the network of chemical reactions and the rates of each reaction within the network such that the concentration of the final product of the automatically created network matches the observed time domain data. Specifically, genetic programming automatically created metabolic pathways involved in the phospholipid cycle and the synthesis and degradation of ketone bodies. PMID- 11262963 TI - Development of a system for the inference of large scale genetic networks. AB - We propose a system named AIGNET (Algorithms for Inference of Genetic Networks), and introduce two top-down approaches for the inference of interrelated mechanism among genes in genetic network that is based on the steady state and temporal analyses of gene expression patterns against some kinds of gene perturbations such as disruption or overexpression. The former analysis is performed by a static Boolean network model based on multi-level digraph, and the latter one is by S-system model. By integrating these two analyses, we show our strategy is flexible and rich in structure to treat gene expression patterns; we applied our strategy to the inference of a genetic network that is composed of 30 genes as a case study. Given the gene expression time-course data set under the conditions of wild-type and the deletion of one gene, our system enabled us to reconstruct the same network architecture as original one. PMID- 11262964 TI - Representation and simulation of biochemical processes using the pi-calculus process algebra. AB - Despite the rapidly accumulating body of knowledge about protein networks, there is currently no convenient way of sharing and manipulation of such information. We suggest that a formal computer language for describing the biomolecular processes underlying protein networks is essential for rapid advancement in this field. We propose to model biomolecular processes by using the pi-Calculus, a process algebra, originally developed for describing computer processes. Our model for biochemical processes is mathematically well-defined, while remaining biologically faithful and transparent. It is amenable to computer simulation, analysis and formal verification. We have developed a computer simulation system, the PiFCP, for execution and analysis of pi-calculus programs. The system allows us to trace, debug and monitor the behavior of biochemical networks under various manipulations. We present a pi-calculus model for the RTK-MAPK signal transduction pathway, formally represent detailed molecular and biochemical information, and study it by various PiFCP simulations. PMID- 11262965 TI - Nutrient-related analysis of pathway/genome databases. AB - We present an algorithm that solves two related problems in the analysis of metabolic networks stored within a pathway/genome database. (1) The Forward Propagation Problem: given a set of nutrients that are inputs to the metabolic network, what compounds will be produced by the metabolic network? (2) The Backtracking Problem: given the results of a forward propagation, and given a set of essential compounds that are not produced as a result of the forward propagation, what precursors must be supplied to produce those essential compounds? A program based on this algorithm is applied to the EcoCyc database, which is a pathway/genome database for E. coli that consists of annotated genomes and the metabolic reactions and pathways associated with the known gene products. The inputs to the program are a description of the metabolic network of an organism (EcoCyc), a set of nutrients corresponding to a known minimal growth medium, and a list of essential compounds to be produced. The program "fires" the microorganism's metabolism contained in the database and predicts all synthesized and nonsynthesized essential compounds, along with the missing precursors required to produce the latter. When applied to the EcoCyc database, the program identifies a number of missing precursors that indicate incomplete regions of the database. Thus the program results can be used to evaluate existing pathway databases like EcoCyc. PMID- 11262966 TI - Detecting gene relations from Medline abstracts. AB - Research in bioinformatics in the past decade has generated a large volume of textual biological data stored in databases such as MEDLINE. It takes a copious amount of effort and time, even for expert users, to manually extract useful information embedded in such a large volume of retrieved data and automated intelligent text analysis tools are increasingly becoming essential. In this article, we present a simple analysis and knowledge discovery method that can identify related genes as well as their shared functionality (if any) based on a collection of relevant retrieved relevant MEDLINE documents. The relative computational simplicity of the proposed method makes it possible to process and analyze large volumes of data in a short time. Hence, it significantly contributes to and enhances a user's ability to discover such embedded information. Two case studies are presented that indicate the usefulness of the proposed method. PMID- 11262967 TI - On reporting fold differences. AB - As we enter an age in which genomics and bioinformatics make possible the discovery of new knowledge about the biological characteristics of an organism, it is critical that we attempt to report newly discovered "significant" phenotypes only when they are actually of significance. With the relative youth of genome-scale gene expression technologies, how to make such distinctions has yet to be better defined. We present a "mask technology" by which to filter out those levels of gene expression that fall within the noise of the experimental techniques being employed. Conversely, our technique can lend validation to significant fold differences in expression level even when the fold value may appear quite small (e.g. 1.3). Given array-organized expression level results from a pair of identical experiments, our ID Mask Tool enables the automated creation of a two-dimensional "region of insignificance" that can then be used with subsequent data analyses. Fundamentally, this should enable researchers to report on findings that are more likely to be in nature truly meaningful. Moreover, this can prevent major investments of time, energy, and biological resources into the pursuit of candidate genes that represent false positives. PMID- 11262968 TI - A comparison of genetic network models. AB - With the completion of the sequencing of the human genome, the need for tools capable of unraveling the interaction and functionality of genes becomes extremely urgent. In answer to this quest, the advent of microarray technology provides the opportunity to perform large scale gene expression analyses. Recently, genetic networks were proposed as a possible methodology for modeling genetic interactions. Since then, a wide variety of different models have been introduced. However, it is, in general, unclear what the strengths and weaknesses of each of these approaches are and where these models overlap and differ. This paper compares different genetic modeling approaches that attempt to extract the gene regulation matrix from expression data. A taxonomy of continuous genetic network models is proposed and the following important characteristics are suggested and employed to compare the models: (1) inferential power; (2) predictive power; (3) robustness; (4) consistency; (5) stability and (6) computational cost. Where possible, synthetic time series data are employed to investigate some of these properties. PMID- 11262969 TI - A nonparametric scoring algorithm for identifying informative genes from microarray data. AB - Microarray data routinely contain gene expression levels of thousands of genes. In the context of medical diagnostics, an important problem is to find the genes that are correlated with given phenotypes. These genes may reveal insights to biological processes and may be used to predict the phenotypes of new samples. In most cases, while the gene expression levels are available for a large number of genes, only a small fraction of these genes may be informative in classification with statistical significance. We introduce a nonparametric scoring algorithm that assigns a score to each gene based on samples with known classes. Based on these scores, we can find a small set of genes which are informative of their class, and subsequent analysis can be carried out with this set. This procedure is robust to outliers and different normalization schemes, and immediately reduces the size of the data with little loss of information. We study the properties of this algorithm and apply it to the data set from cancer patients. We quantify the information in a given set of genes by comparing its distribution of the score statistics to a set of distributions generated by permutations that preserve the correlation structure among the genes. PMID- 11262970 TI - PIES, a protein interaction extraction system. AB - We consider the problem of extracting, manipulating, and managing biological pathways, especially protein-protein interaction pathways. We discuss here the Protein Interaction Extraction System (PIES). PIES is contructed on top of three main technologies: Kleisli, BioNLP, and Graphviz. Kleisli is a broad-scale data integration system that we use for downloading Medline abstracts and for general manipulation and management of pathway/interaction databases. BioNLP is a natural language-based information extraction module that we use for analysing Medline abstracts and to extract precise protein-protein and other interaction information. Graphviz is a graphical layout package developed for directed graphs that we use for visualization of the extracted pathways. PIES can be augmented with various means for extracting protein interaction information from sequence databases, for example, by using Kleisli's power to integrate sequence comparison tools to detect gene fusion events in sequence databases. PMID- 11262971 TI - Simulating the growth of viruses. AB - To explore how the genome of an organism defines its growth, we have developed a computer simulation for the intracellular growth of phage T7 on its E. coli host. Our simulation, which incorporates 30 years of genetic, biochemical, physiological, and biophysical data, is used here to study how the intracellular resources of the host, determined by the specific growth rate of the host, contribute toward phage development. It is also used to probe how changes in the linear organization of genetic elements on the T7 genome can affect T7 development. Further, we show how time-series trajectories of T7 mRNA and protein levels generated by the simulation may be used as raw data to test data-mining strategies, specifically, to identify partners in protein-protein interactions. Finally, we suggest how generalization of this work can lead to a knowledge driven simulation for the growth of any virus. PMID- 11262972 TI - Scaling of accuracy in extremely large phylogenetic trees. AB - The accuracy of phylogenetic inference was examined in simulated data sets up to nearly 10,000 taxa, the size of the largest set of homologous genes in existing molecular sequence databases. Even with a simple search algorithm (maximum parsimony without branch swapping), the number of characters needed to estimate 80% of a tree correctly can scale remarkably well at optimal substitution rates (on the order of log N, where N is the number of taxa). In other words, the number of taxa in an analysis can be doubled and only an arithmetic increase in the number of characters is required to maintain the same level of accuracy. Even substitution rates that are much higher than normally used in phylogenetic studies did not affect the scaling too adversely. However, scaling is usually worse than log N for more stringent levels of accuracy. Moreover, errors are not distributed randomly throughout the tree. Shallow nodes are remarkably easy to reconstruct and display favourable log-linear scaling. The deepest nodes are extremely difficult to reconstruct accurately, even with branch swapping, and the scaling is poor. Therefore, the strategy of sequencing large numbers of homologous genes may not always provide global solutions to extreme phylogenetic problems and alternative strategies may be required. PMID- 11262973 TI - Exact algorithms for computing pairwise alignments and 3-medians from structure annotated sequences (extended abstract). AB - Given the problem of mutation saturation in ancient molecular sequences, there is great interest in inferring phylogenies from higher-order types of molecular data that change more slowly, such as genomic organization and the secondary and tertiary structures of ribosomal RNA and proteins. In this paper, we define edit distances based on two representations of RNA secondary structure, arc annotation and hierarchical string annotation, and give algorithms for computing these distances on pairs of annotated sequences, aligning pairs of annotated sequences, and computing 3-median annotated sequences from triples of annotated sequences. The 3-median algorithms can be used as part of a well-known iterative heuristic for inferring phylogenies. All given algorithms are adapted from algorithms for computing longest common annotated subsequences of pairs of annotated sequences. PMID- 11262974 TI - A tree obscured by vines: horizontal gene transfer and the median tree method of estimating species phylogeny. AB - A phylogeny is a tree graph representation of genealogical relationships between biological objects. It is of general interest to estimate the phylogeny of whole organisms (species trees) using bio-molecular sequences. When multiple sequences are available for each organism such as with whole genome data, individual phylogenies estimated by each molecule (gene trees) may not be concordant. The lack of concordance may be due to actual biological mechanisms such as horizontal transfer of the molecules. Here, we present a new phylogeny estimation method designed to estimate the species tree despite such horizontal transfer. It uses the idea that horizontal transfer distorts distance relationships between pairs of species but a median estimate of the distances is robust to such distortions. We demonstrate the utility of our method using a simulation study. PMID- 11262975 TI - A new implementation and detailed study of breakpoint analysis. AB - Phylogenies derived from gene order data may prove crucial in answering some fundamental open questions in biomolecular evolution. Yet very few techniques are available for such phylogenetic reconstructions. One method is breakpoint analysis, developed by Blanchette and Sankoff for solving the "breakpoint phylogeny." Our earlier studies confirmed the usefulness of this approach, but also found that BPAnalysis, the implementation developed by Sankoff and Blanchette, was too slow to use on all but very small datasets. We report here on a reimplementation of BPAnalysis using the principles of algorithmic engineering. Our faster (by 2 to 3 orders of magnitude) and flexible implementation allowed us to conduct studies on the characteristics of breakpoint analysis, in terms of running time, quality, and robustness, as well as to analyze datasets that had so far been considered out of reach. We report on these findings and also discuss future directions for our new implementation. PMID- 11262976 TI - A new algorithm for analysis of within-host HIV-1 evolution. AB - A new algorithm for inferring the evolution of within-host viral sequences is presented. A sequential-linking approach is developed so that a longitudinal phylogenetic tree can be reconstructed from sequential molecular data that are obtained at different time points from the same host. The algorithm employs a codon-based model, which uses a Markov process to describe substitutions between codons, to calculate nonsynonymous and synonymous substitution rates and to distinguish positive selection and neutral evolution. The algorithm is applied to a data set of the V3 region of the HIV-1 envelope genes sequenced at different years after the infection of a single patient. The results suggest that this algorithm may provide a more realistic description of viral evolution than traditional evolutionary models, because it accounts for both neutral and adaptive evolution, and reconstructs a longitudinal phylogenetic tree that describes the dynamic process of viral evolution. PMID- 11262977 TI - Determining significant fold differences in gene expression analysis. AB - A typical use for RNA expression microarrays is comparing the measurement of gene expression of two groups. There has not been a study reproducing an entire experiment and modeling the distribution of reproducibility of fold differences. Our goal was to create a model of significance for fold differences, then maximize the number of ESTs above that threshold. Multiple strategies were tested to filter out those ESTs contributing to noise, thus decreasing the requirements of what was needed for significance. We found that even though RNA expression levels appears consistent in duplicate measurements, when entire experiments are duplicated, the calculated fold differences are not as consistent. Thus, it is critically important to repeat as many data points as possible, to ensure that genes and ESTs labeled as significant are truly so. We were successfully able to use duplicated expression measurements to model the duplicated fold differences, and to calculate the levels of fold difference needed to reach significance. This approach can be applied to many other experiments to ascertain significance without a priori assumptions. PMID- 11262978 TI - Efficiencies of genes and accuracy of tree-building methods in recovering a known Drosophila genealogy. AB - Phylogenetic hypotheses generated from seven Drosophila mitochondrial genomes support a well-corroborated genealogy with a single evolutionary history. These mitochondrial data form a model system for investigating the efficiency of genes and accuracy of different tree-building methods in recovering a well-supported genealogy. We consider 15 genes (13 protein-coding and 2 rRNAs) and 83 tree building methods (27 distance, 4 parsimony, 50 maximum likelihood, and 2 Bayesian). Among the 15 genes examined, ND4 recovered the true genealogy most efficiently (82 out of 83 methods). Generally, maximum likelihood models enforcing a clock most accurately reclaim the true genealogy. Surprisingly, however, this method fails to recover the well-supported topology for more than half of the genes. Additional studies are required to test the generality of the results presented here. PMID- 11262979 TI - An NMR-based quenched hydrogen exchange investigation of model amyloid fibrils formed by cold shock protein A. AB - Acid-denatured cold shock protein A (CspA) self-assembles into polymers with properties typical of amyloid fibrils. In the present work, a quenched hydrogen exchange experiment was used to probe the interactions of CspA fibrils with solvent. Exchange was initiated by incubating suspensions of fibrils in D2O, and quenched by flash freezing. Following lyophilization, exchange-quenched samples were dissolved in 90% DMSO/10% D2O, giving DMSO-denatured monomers. Intrinsic exchange rates for denatured CspA in 90% DMSO/10% D2O (pH* 4.5) were sufficiently slow (approximately 1 x 10(-3) min-1) to enable quantification of NMR signal intensity decays due to H/D exchange in the fibrils. Hydrogen exchange rate constants for CspA fibrils were found to vary less than 3-fold from a mean value of 5 x 10(-5) min-1. The uniformity of rate constants suggests that exchange is in the EX1 limit, and that the mechanism of exchange involves a cooperative dissociation of CspA monomers from fibrils, concomitant with unfolding. Previous studies indicated that the highest protection factors in native CspA are approximately 10(3), and that protection factors for the acid-denatured monomer precursors of CspA fibrils are close to unity. Because exchange in is in the EX1 regime, it is only possible to place a lower limit of at least 10(5) on protection factors in CspA fibrils. The observation that all amide protons are protected from exchange indicates that the entire CspA polypeptide chain is structured in the fibrils. PMID- 11262980 TI - Collective reorientational motion and nuclear spin relaxation in proteins. AB - Significant progress in NMR methodology for measuring spin-relaxation data at many different 15N and 13C sites in proteins demands new and increasingly sophisticated ways of data interpretation. Recent work of our group concerning the use of anisotropic and reorientational collective motional models for spin relaxation interpretation is briefly reviewed and a number of important aspects of collective reorientational motional models are discussed at the example of a 11 ns molecular dynamics computer simulation of the protein ubiquitin. PMID- 11262981 TI - The protein non-folding problem: amino acid determinants of intrinsic order and disorder. AB - To investigate the determinants of protein order and disorder, three primary and one derivative database of intrinsically disordered proteins were compiled. The segments in each primary database were characterized by one of the following: X ray crystallography, nuclear magnetic resonance (NMR), or circular dichroism (CD). The derivative database was based on homology. The three primary disordered databases have a combined total of 157 proteins or segments of length.30 with 18,010 residues, while the derivative database contains 572 proteins from 32 families with 52,688 putatively disordered residues. For the four disordered databases, the amino acid compositions were compared with those from a database of ordered structure. Relative to the ordered protein, the intrinsically disordered segments in all four databases were significantly depleted in W, C, F, I, Y, V, L and N, significantly enriched in A, R, G, Q, S, P, E and K, and inconsistently different in H, M, T, and D, suggesting that the first set be called order-promoting and the second set disorder-promoting. Also, 265 amino acid properties were ranked by their ability to discriminate order and disorder and then pruned to remove the most highly correlated pairs. The 10 highest ranking properties after pruning consisted of 2 residue contact scales, 4 hydrophobicity scales, 3 scales associated with.-sheets and one polarity scale. Using these 10 properties for comparisons of the 3 primary databases suggests that disorder in all 3 databases is very similar, but with those characterized by NMR and CD being the most similar, those by CD and X-ray being next, and those by NMR and X-ray being the least similar. PMID- 11262982 TI - Serum immunoglobulins and immunoglobulin G subclasses with recurrent wheezing. AB - In this study serum immunoglobulins (Ig) and IgG subclasses were measured in 42 patients (ranging 9 month-6 year) with recurrent wheezing and in 37 healthy children determined the relationship between serum Igs and recurrent wheezing. Patients were divided into two groups according to the age [9 month-2 year (n: 15), and 2-6 year (n: 27)]. In the patients placed in 9-24 month age group, serum IgG4 level was found to be lower than controls (p < 0.05). But there was not a significant difference in mean serum concentrations of total IgG, IgA, IgM, IgE, IgG1, IgG2 and IgG3 subclasses between the groups (P > 0.05). In the 25 month-6 year age group the mean IgE level was increased compared to the control while IgG3 and IgG4 levels were decreased (p < 0.05). On the other hand, in the 9-24 month age group there was no significant difference between the patients and controls for IgG subclasses deficiency (P > 0.05). However, significant difference in IgG subclasses deficiency was present between the patients and controls in the 25 month-6 year group (P < 0.001). In conclusion, our findings suggest that wheezing in childhood may be associated with low IgG3 and/or IgG4, and in older children high IgE level may be a part of pathogenetic mechanism in patients with recurrent wheezing. PMID- 11262984 TI - Immune functions in splenectomized thalassaemic children. AB - A prospective study to assess the immune functions in splenectomized thalassaemic children. Children were those registered in the Thalassemia major. There were 10 splenectomized children (Group 1), 10 non-splenectomized children and 6 age matched control (Group 3). All children were shown to be HIV seronegative. The mean concentrations of serum IgG and IgA were higher in Group 1 as compared to Groups 2 and 3 but the differences were not statistically significant. Nitroblue tetrazolium (NBT) dye reduction by stimulated polymorphonuclear leukocytes was normal in both study and control groups and the differences were not statistically significant. However, NBT reduction in the unstimulated state was much higher in Group 2 as compared to Groups 1 and 3. Phytohaemagglutinin induced mitogen proliferation was normal in all 3 groups. Children in Group 1 not only had a significantly higher absolute lymphocyte count but also had a lower CD4/CD8 ratio as compared to Groups 2 and 3. Splenectomy does appear to alter the immune status of thalassemic children but the exact mechanism by which this occurrence is not clear. PMID- 11262983 TI - Antipyretic effects of nimesulide, paracetamol and ibuprofen-paracetamol. AB - The antipyretic effect of nimesulide has not been adequately compared with paracetamol and ibuprofen-paracetamol combination in children. Hence, a randomized, double blind, and parallel groups' design and multicenter study was conducted on children with respiratory tract infections. Eighty-nine patients with temperatures above 38.5 degrees C were randomly administered nimesulide (1.5 mg/kg/dose), paracetamol (10.0 mg/kg/dose), or ibuprofen-patients combination (10.0 mg/kg/dose), thrice daily for five days. The axillary temperature was recorded at the baseline and at different time intervals post administration of drugs. The hematological and biochemical investigations were performed at the basal level and at the end of the treatment period. The adverse drug reactions were monitored during the trial. All the drugs produced a significant fall in temperature as compared to their respective basal values (p < 0.001). However, on looking at the change in temperatures at different time intervals from the respective basal levels, no significant difference was found among all the drugs. Surprisingly, nimesulide had a tendency to raise serum glutamate pyruvate transaminase and serum glutamate oxaloacetate transaminase levels as compared to its baseline values. There was no marked adverse effect of the drugs on other hematological and biochemical parameters investigated. No other serious adverse reaction occurred in the study. Ibuprofen-paracetamol combination, nimesulide, and paracetamol had almost similar antipyretic effects in children. PMID- 11262985 TI - Adolescent behaviour regarding reproductive health. AB - A cross-sectional household survey was undertaken in rural areas of district Sirmaur, Himachal Pradesh, to assess the knowledge, beliefs and practices of adolescents about reproductive health. Six hundred and forty three unmarried adolescents aged 15-19 years were selected by 30-cluster sampling method from 2400 households. They were interviewed using a semi-structured schedule. Fifty six per cent were girls. More girls (14%) were illiterate than boys (6%). Most of the boys (88%) and 58% girls knew that a female conceives through sexual intercourse. Seventy seven per cent girls and eighty seven per cent boys were aware of at least one contraceptive method. Majority of the girls (71%) and boys (82.5%) favoured termination of an unwanted pregnancy. About one-fourth respondents considered husband responsible for infertility and for sex of the baby. Boys considered night emission, poor body built and less growth of hair as reproductive health problems, whereas, girls were worried about menstruation and inadequate breast development. Almost 6% boys reported use of a contraceptive method indicating existence of pre-marital sexual activity. Knowledge on reproductive health is low and there is a big gap between actual and desired practices. PMID- 11262986 TI - An audit of phototherapy units. AB - Audit of phototherapy aims to examine their efficacy and functioning. Twenty-four centres providing neonatal care were visited by a team of two persons. Data form were completed with information regarding age of phototherapy units, its buildup in relation to number of tubelights, type of light and irradiance provided. Total of 58 units examined had a wide variety in relation to their build. There were only 21 units (36.2%) in which all the lights were in working order. Only five of the units (8.6%) had the recommended special blue lights. Only 18 of the units (31%) provided an acceptable level of irradiance. Phototherapy demonstrates a dose response relationship. By not providing optimum irradiance, the efficacy is compromised. This prolongs hospital stay and treatment costs. Acceptable standards should be insisted upon in purchase and maintenance of medical equipments. PMID- 11262987 TI - Oxygen therapy in pediatrics. AB - Oxygen therapy is the most important aspect of supportive care in the management of a critically ill child. Knowledge of the physiology of oxygenation is a key to the proper oxygen therapy. High flow systems are more dependable devices for oxygenation and their use needs to be stressed. Patients on oxygen need close monitoring. Ventilatory support and Continuous Positive Airway Pressure (CPAP) is mandatory in some patients in addition to oxygen therapy for the prevention and treatment of hypoxia. PMID- 11262988 TI - Burden of genetic disorders in India. AB - India, like other developing countries, is facing an accelerating demographic switch to non-communicable diseases. In the cities congenital malformations and genetic disorders are important causes of morbidity and mortality. Due to the high birth rate in India a very large number of infants with genetic disorders are born every year almost half a million with malformations and 21,000 with Down syndrome. In a multi-centric study on the causes of referral for genetic counselling the top four disorders were repeated abortions (12.4%), identifiable syndromes (12.1%), chromosomal disorders (11.3%) and mental retardation (11%). In a more recent study in a private hospital the top reasons for referral were reproductive genetics (38.9%)--comprising prenatal diagnosis, recurrent abortions, infertility and Torch infections--mental retardation +/- multiple congenital anomalies (16.1%), Down syndrome (9.1%), thalassemia/haemophilia (8.8%), and muscle dystrophy/spinal muscular atrophy (8.4%). The disorders for which prenatal has been done over an 18-month-period are given. A recent study carried out in three centers (Mumbai, Delhi and Baroda) on 94,610 newborns by using a uniform proforma showed a malformation frequency of 2.03%, the commonest malformations are neural tube defects and musculo-skeletal disorders. The frequency of Down syndrome among 94,610 births was 0.87 per 1000, or 1 per 1150. Screening of 112,269 newborns for aminoacid disorders showed four disorders to be the commonest--tyrosinemia, maple syrup urine disease and phenylketonuria. Screening of cases of mental retardation for aminoacid disorders revealed four to be the commonest--hyperglycinemia, homocystinuria, alkaptonuria, and maple syrup urine disease. Metabolic studies of cases of mental retardation in AIIMS, Delhi and KEM Hospital, Mumbai, demonstrated that common disorders were those of mucopolysaccharides, lysosomes, Wilson disease, glycogen storage disease and galactosemia. It is estimated that beta- thalassemia has a frequency at birth of 1:2700, which means that about 9,000 cases of thalassemia major are born every year. Almost 5200 infants with sickle cell disease are born every year. Disorders, which deserve to be screened in the newborn period, are hypothyroidism and G-6-PD deficiency, while screening for aminoacid and other metabolic disorders could presently be restricted to symptomatic infants. PMID- 11262989 TI - Prenatal diagnosis and fetal therapy--what lies in future? AB - Prenatal diagnosis has traditionally occurred between the 15th and 20th weeks of gestation. However, the capabilities of screening and diagnostic tools have advanced substantially over the past 10 years. Recent advances in the science of prenatal diagnosis allow for the evaluation of an affected embryo or an abnormal cell line prior to gestation within the womb via preimplantation diagnosis. The technique can be used for any genetic condition which can be detected with a chromosome-specific probe. At the current time, noninvasive second trimester maternal serum screening with either 2 or 3 serum analytes is associated with a 60-70% detection rate of Down syndrome. Although these particular serum markers are not useful during the first trimester, the fetoplacental secretory products free beta-hCG and pregnancy-associated plasma protein-A (PAPP-A) appear to be meaningful clinically when measured between 8 and 13 weeks of gestation, yielding similar first trimester detection rates as the current second trimester screening programme. The addition of nuchal translucency measurements as an independent predictor of fetal aneuploidy may further increase the detection rate of Down syndrome to 80%. Open fetal surgery is now possible under highly selective circumstances in which the fetal condition is considered life-threatening and the prognosis is extremely poor. Surgical intervention may be appropriate for congenital cystic adenomatoid malformation, bronchopulmonary sequestration, congenital diaphragmatic hernia, and possibly for myelomeningocele. PMID- 11262990 TI - Evaluation of newborns with skeletal dysplasias. AB - Skeletal dysplasia are a relatively common and distinct group of genetic disorders. Even though abnormalities of bone growth are the major clinical manifestations, the pathological process may involve in any body system. Many of these disorders have dire consequences such as neonatal death and most will have life-long physical and psychosocial morbidity associated with them. Recent advances in genetic research have identified the genes underlying the primary defects in several common skeletal dysplasias such as achondroplasia and diastrophic dysplasia. While these advances are clearly important in developing better therapy and a cure for those conditions, the role of the pediatrician as the diagnostician has remained unchanged. In this article we describe how a systematic approach using the simplest of tools--clinical assessment and radiograph--can arrive at a diagnosis in most instances of newborns with skeletal dysplasias. The key features that are essential for establishing a diagnosis for most of the entities encountered in the newborn are described along with our general approach to the evaluation of the radiographs. PMID- 11262992 TI - Muscular dystrophies. AB - The muscular dystrophies (MD) are a heterogenous group of genetically determined, variably inherited primary disorders of muscle that progress differently. The various forms can be distinguished by the combination of clinical, genetic and pathologic criteria, confirmation of the muscle biopsy should be with immunohistochemical staining rather than histological only. These and the gene deletion studies in the families have become essential diagnostic criteria. PMID- 11262991 TI - Neural tube defects: prevention by folic acid and other vitamins. AB - Folic acid has been demonstrated in clinical trials to reduce significantly the recurrence (and probably occurrence) of neural tube defects (NTD). In the U.K., there has been no decline in prevalence of NTD since the publication of the findings with folic acid. This article examines a series of questions relating to the action of folic acid, with emphasis on the use of mouse models as a source of experimental information which cannot easily be obtained by direct study of humans. Several mouse genetic NTD models exhibit sensitivity to prevention by folic acid, whereas other mice which develop morphologically similar NTD are resistant. Folic acid normalises neurulation in the sensitive mouse strains, providing evidence for a direct effect on the developing embryo, not on the pregnant female: Mouse studies do not support the proposed action of folic acid in encouraging the in utero demise of affected fetuses (i.e. terathanasia). Polymorphic variants of several folate-related enzymes have been shown to influence risk of NTD in humans and an inherited abnormality of folate metabolism has been demonstrated in one mouse NTD model. However, the biochemical basis of the action of folic acid in preventing NTD remains to be determined in detail. NTD in one folate-resistant mouse strain can be prevented by myo-inositol, both in utero and in vitro, raising the possibility of a therapeutic role also in humans. Gene-gene interactions seem likely to underlie the majority of NTD, suggesting that poly-therapy involving folic acid and other agents, such as myo inositol, may prove more effective in preventing NTD than folic acid treatment alone. PMID- 11262993 TI - Ellis-Van Creveld syndrome associated with nodular sclerosing Hodgkin's disease and nephrotic syndrome. AB - An 11-year-old male child presented with multiple congenital anomalies, gradually increasing swelling on the left side of neck for one month along with generalized swelling of body and passage of scanty urine for fifteen days. On examination, the child had multiple congenital anomalies and cervical lymphadenopathy. Cardiac examination revealed a pansystolic murmur grade III/VI in left lower parasternal area. Laboratory evaluation revealed significant hypoalbuminemia and hypercholesterolemia, 24 hours urinary protein was 116 mg/hour/m2. Histopathological examination of left cervical lymphnode revealed Hodgkin's disease (Nodular Sclerosing type), 2D echocardiography revealed presence of a single atrium. A diagnosis of Ellis-van Creveld syndrome with nodular sclerosing Hodgkin's disease and nephrotic syndrome was kept. This association, to the best of our knowledge has not been previously reported. PMID- 11262995 TI - Hypospadias-hypertelorism syndrome. AB - A young male child presented with hypospadias. Examination revealed additional anomalies including hypertelorism and upslanting of palpebral fissures, suggesting a diagnosis of hypospadias-hypertelorism syndrome. The case is reported because of its rarity and some unusual features. PMID- 11262994 TI - Duodeno-jejunal hemangiomatosis. AB - Diffuse duodeno-jejunal hemangiomatosis in children is a rare cause of bilious vomiting. In the clinical approach to bilious vomiting, tumors of the duodenum come at the end of the differential list--not to mention the rarity of hemangiomatosis. To our knowledge, isolated duodeno-jejunal hemangiomatosis as a cause of bilious vomiting in children is being reported for the first time. We analyse the various imaging modalities available to reach a clinical diagnosis. PMID- 11262996 TI - Extensive gliomas of visual tract in a patient of neurofibromatosis-I. AB - Although bilateral optic nerve gliomas are commonly found in patients with neurofibromatosis I, extensive gliomas involving the entire visual tracts, bilaterally are relatively rare. Usually the optic radiations are spared. We report a case of a 2-year-old child with extensive disease of bilateral visual pathways with involvement of the hypothalamus manifesting as obesity. PMID- 11262997 TI - MR imaging in cervico-thoracic lymphangioma. PMID- 11262998 TI - Authors' reply to "Plasmodium ovale infection". PMID- 11262999 TI - [ACE inhibition in patients with myocardial infarct and ventricular dysfunction: inappropriate application of therapy standards in patient samples]. AB - Several reports indicate the benefit of ACE inhibitors for patients with left ventricular systolic dysfunction after acute myocardial infarction (MI). We sought to determine the implementation of the treatment guidelines in patient samples from the general population. Furthermore we aimed to identify patient characteristics associated with the use of ACE inhibitors. Screening of two MI registries allowed the identification of 226 MI patients with left ventricular dysfunction. Patients were considered to be eligible for ACE inhibitor therapy when a EF < or = 40% was documented in the patient records of cardiac rehabilitation clinics (REG-MI, n = 147) or detected by standardised echocardiography (KORA, n = 78). On average 5.5 years following MI, a standardised questionnaire and a detailed medical history was obtained. Specifically, information was collected regarding current medication and potential contraindications for ACE inhibitors. MI patients with LV dysfunction received ACE inhibitors in 62% (REG-MI) and 45% (KORA). The doses prescribed were substantially smaller than target doses used in the large-scale studies (REG-MI: 40 +/- 4%, KORA: 23 +/- 3%, % of target doses). Only 13% (REG-MI) and 3% (KORA) received more than 50% of the target dosage. Additionally, actual doses of the most frequently used ACE inhibitors were significantly different (captopril: 23 +/- 2%, enalapril: 42 +/- 5% of target doses). The likelihood of receiving ACE inhibitors was significantly higher in patients with written recommendation for such medication (odds ratio 6.02, confidence interval 1.93-20.16) and in patients visiting cardiologists (odds ratio 3.69, confidence interval 1.26-11.07) as revealed by multivariate analysis of the REG-MI database. Despite national and international guidance, a large proportion of MI patients with left ventricular dysfunction is not receiving ACE inhibitors, and when used, the doses prescribed are markedly smaller than target doses used in clinical trials that established the utility of these drugs. Medical care by cardiologists and written recommendation of ACE inhibition in patient records were independent predictors of a more appropriate prescription of ACE inhibitors. PMID- 11263000 TI - Angioscopic predictors of restenosis following coronary angioplasty--the impact of yellow smooth plaques. AB - Despite angioscopy being used for more than 10 years, data regarding the prognostic significance are still limited. This study evaluated the prognostic relevance of the angioscopic lesion morphology and plaque colour on restenosis rate following coronary angioplasty. Out of 66 patients with coronary angioscopy prior to an angioplasty procedure, 46 patients with successful balloon dilation and 16 patients with stenting were included into the study. Angioscopic plaque morphology and plaque colour were correlated with the anginal status, the angiographic lesion morphology, the procedural result, and the restenosis rate during six months follow-up. Clinical follow-up was obtained from all patients, re-angiography was performed in 61%. Angioscopically complicated lesions were more frequent in patients with unstable versus stable angina (63% versus 28%, p < 0.005) and ACC/AHA type B2/C versus A/B1 stenoses (86% versus 7%, p < 0.03). In addition yellow plaques were more often seen in unstable versus stable angina (80% versus 50%, p < 0.02) and in type B2/C versus A/B1 lesions (81% versus 47%, p < 0.005). There were no deaths or myocardial infarctions during follow-up. Restenosis (n = 11) occurred significantly more frequent in patients with smooth, yellow lesions (37%) compared to all other lesion morphologies (9%, p < 0.02). Logistic regression analysis revealed angioscopically smooth, yellow plaques (p < 0.05) and angiographically type B2/C lesions (p < 0.03) as independent predictors for restenosis. We conclude that angioscopically smooth, yellow plaques covered by an intact inner vessel surface are associated with a higher incidence of restenosis following coronary angioplasty, potentially indicating a higher proliferative response after a mechanical trauma of such lesions. PMID- 11263001 TI - [Advances in interventional occlusion of persistent ductus arteriosus: comparison of results using different occlusion devices]. AB - This report describes our results with transcatheter closure of patent ductus arteriosus between March 1993 and May 2000 including our early experience with the Amplatzer duct occluder. One hundred and sixty-six consecutive procedures were performed in 160 patients. The mean age was 6.8 years (range 0.8 to 26.2), mean weight 24.3 kg (range 7.7 to 84.0). Single or multiple coils were implanted successfully in 114 cases, a Rashind double umbrella in 35 patients, and an Amplatzer duct occluder in 16 patients. After failure to implant coils in one patient, a second attempt with a Rashkind double umbrella was successful. Complete closure of the patent ductus arteriosus was achieved in a total of 148 patients (92.5%), mean fluoroscopy time was 13.7 min (range 3.1 to 126 min). In 144 patients (90.0%), occlusion without residual shunting was achieved by the first interventional approach. Angiography showed immediate closure in 44%, echocardiography within 48 h revealed complete closure in additional 28%. In a further 18%, a residual shunt disappeared spontaneously in the following months. In five patients a second procedure was performed to terminate residual shunting after placement of a Rashkind occluder system by coils. In three patients, the residual shunt resolved. In 12 patients (7.5%) echocardiography showed a residual shunt 2-1259 days after interventional approach. Occlusion rates of the different devices were 83% for the Rashkind occluder, 92% for detachable coils, and 100% for the Amplatzer duct occluder. Coil embolisation into the pulmonary artery occurred in two patients, interventional removal was successful in one of them. There were no further complications. According to our experience interventional occlusion of patent ductus arteriosus is highly effective and associated with a low complication rate. Transcatheter closure using the Amplatzer duct occluder seems to be safe and effective also in small children with a body weight of 8 kg and a large ductus diameter. Compared to the total group of patients, the occlusion rate of the Amplatzer duct occluder was significantly higher (p = 0.005), and of the Rashkind occluder system significantly lower (p = 0.026). Therefore, we recommend the use of detachable coils in patients with small ductus (diameter < or = 2 mm) and the Amplatzer duct occluder in those with a larger ductus. Transcatheter closure of the patent ductus arteriosus according to this regimen should achieve occlusion rates above 95%. PMID- 11263002 TI - [Double aortic arch: clinical aspects, diagnosis and therapy in children and adults]. AB - Double aortic arch is a rare vascular anomaly which usually causes tracheal and esophageal compression in the first few months of life. During the last 30 years, 7 children, 2 to 24 months old, and one 29-year-old woman with double aortic arches have been treatedatour institution. Symptoms, diagnosis and treatment of these patients were evaluated. Dyspnoe, stridor, recurrent pulmonary infections, feeding problems and failure to thrive were the leading symptoms. Despite typical symptoms from early childhood, the diagnosis was missed in our adult patient. Typical compression of the esophageus and the trachea was visualized by esophagography by 7 and bronchoscopy/-graphy by 6 patients. Angiography was performed in all children, whereas magnetic resonance angiography and computed tomography were done in the adult patient. Resection of the smaller aortic arch, left in 3 and right in 5, through a right or a left posterolateral thoracotomy was uncomplicated and fully resolved the symptoms in all patients. Typical symptoms in early childhood should lead to prompt diagnosis and surgical treatment of double aortic arch. Surgical resection of the smaller aortic arch should also be performed in oligosymptomatic patients to prevent complications later. Preoperative angiography can be replaced by the less invasive magnetic resonance imaging and computed tomography. PMID- 11263003 TI - [Aortic root abscess without involvement of the aortic valve: diagnosis and therapy in a 2.5-year-old child]. AB - Although formation of an aortic root abscess is a frequent complication of aortic valve endocarditis in adults, this complication has been rarely observed in children. In the majority of cases it has been described in children without underlying congenital heart disease. Due to the rarity of this complication, diagnosis and treatment is frequently delayed in childhood. We report a 2 1/2 year old girl who developed pericardial effusion in the course of pneumonia. Echocardiographic examinations, which were performed because of the pericardial effusion, revealed after 6 days the development of a cystic structure posterior to the aortic root. There was a perforation of this aortic root abscess to the left ventricular outflow tract; the aortic and mitral valves however were normal without endocarditic vegetations. Surgery was performed on the 10th day following a rapid increase in the size of the abscess. During surgery the abscess was drained and the perforation to the left ventricle was closed with direct sutures. Intraoperative transesophageal echocardiography confirmed a good surgical result. Blood cultures remained negative; in the material from the abscess however we found staphylococcus aureus. The postoperative course was uneventful. Our case demonstrates the necessity of detailed and repeated echocardiographic examinations in children with possible symptoms of bacterial endocarditis (in our case pericardial effusion) as well as the requirement of cultures of the abscess for identification of the infective organism. Intraoperative transesophageal echocardiography allows exact description of an aortic root abscess, its relation to other cardiac structures and immediate evaluation of the surgical result. PMID- 11263004 TI - [Late stent thrombosis after intracoronary brachytherapy. A case report and review of the literature]. AB - Intracoronary irradiation is currently the most promising approach to reduce restenosis after percutaneous transluminal coronary angioplasty. Meanwhile numerous data are available concerning efficacy and safety of this novel method. These data confirm the results of preclinical studies that reported a dramatic reduction of neo-intima proliferation and negative remodeling. However, the number of reports on an elevated incidence of late stent thrombosis (> 30 days post intervention) are increasing. It is commonly suggested that the delayed neo intima formation within vascular stents is responsible for this new phenomenon. We report the case of a 48-year-old man who underwent coronary irradiation therapy after stent placement in a de-novo/restenotic lesion. Despite an explicit recommendation of a combined anti-aggregatory therapy consisting of ticlopidine and acetysalicylic acid for at least 6 months, ticlopidine was withdrawn after 4 weeks. Two weeks later, the patient was readmitted to an external hospital with an acute myocardial infarction and successfully treated with thrombolysis. The angiographic and intravascular control, which was conducted after another two weeks, showed absolutely no neointima formation within the implanted stent. Thus, a late thrombotic occlusion of the implanted stent appears most likely to be the cause underlying the myocardial infarction. This case underlines, together with other existing reports, the importance of a prolonged, combined anti-aggregatory therapy after stent placement and subsequent intracoronary irradiation. PMID- 11263005 TI - ["Speaking" and "writing"!]. PMID- 11263006 TI - [Recommendations for an extensive risk decrease for patients with coronary disease, vascular diseases and diabetes. Issued by the Executive Committee of the German Society of Cardiology, Heart and Circulation Research, reviewed on behalf of the Clinical Cardiology Commission by the Prevention Project Group]. PMID- 11263007 TI - [Splice site mutations and atherosclerosis: mechanisms and prediction models]. AB - Nucleotide variants in genes of the lipid metabolism influence the risk of premature atherosclerosis. Ten percent of all single nucleotide substitutions in these genes involve splice sites. The effects of these changes on mRNA splicing and phenotypic severity, however, are not inherently obvious from the nucleotide sequence. This review presents various genes of lipid metabolism with splicing mutations known to influence the risk of premature atherosclerosis. Mechanisms of pre-mRNA splicing are illustrated and different models for prediction of the effect of nucleotide substitutions on splice-site function are presented. The role of information theory-based models is emphasized along with its role for prediction of splice-site function and phenotypic severity of atherosclerosis. PMID- 11263008 TI - [Vascular hardening: molecular biology and biophysics]. PMID- 11263009 TI - Topographic distribution of peripheral arteriopathy in non-diabetics and type 2 diabetics. AB - Peripheral arteriopathy (PA) in type 2 diabetics carries a worse prognosis compared to non-diabetics likely related to a more aggressive course of the macroangiopathy, presence of a prominent microangiopathy and worse results following revascularizations. In addition, the presence of Monckeberg's disease in diabetics has an additional negative impact. Interestingly, PA in diabetics compared to non-diabetics is redistributed towards periphery involving predominantly the lower leg arteries. Based on the evidence we suggest that diabetic PA represents a distinct form of a systemic vascular disease characterized by myointimal thickening of macro- and microvascular beds associated with acceleration of the common variety atherosclerosis and in some cases Monckeberg's media sclerosis. Despite the systemic involvement specific localizing factors determine the severity and clinical relevance of the diabetic PA in individual vascular beds. In this paper some of the potential localizing factors for diabetic PA are presented and briefly discussed. PMID- 11263010 TI - [The immunologic homunculus in rheumatoid arthritis. A new viewpoint of immunopathogenesis in rheumatoid arthritis and therapeutic consequences]. AB - Autoreactivity plays a major role in the pathogenesis of RA. The rheumatoid factor has been and still is for now more than 50 years the only autoreactivity that is clinically applied in the diagnosis of RA. This well reflects the current way of thinking that a single antigen or a single cause drives an individual into disease. Although by now many other autoantigens and autoreactivities have been described, their discovery was always on the search for the one and only autoreactivity that causes RA. This includes also immune reactivities directed against xenogenic antigens. But, none of the known RA-associated autoreactivities is present in all RA patients and none of them occurs exclusively in RA. Thus, the observed sensitivities and specificities are well below 100%. Therefore, RA has often been postulated to consist of various immunological subentities with similar clinical symptoms. Nevertheless, none of the autoreactivities correlates with a distinct clinical feature or course of disease. It is about time to say good-bye to the idea that a single antigen or immunoreactvity causes and maintains rheumatoid arthritis. In this paper we present RA as the clinical outcome of an immune system that has shifted from a healthy to an autoimmune steady state. This is accomplished by many different reactivities and autoreactivities that occur either in parallel or one after the other. The entirety of the known RA-associated reactivities and (auto)antigens is presented in detail. The major RA-relevant autoantigens comprise BiP, citrulline, the Sa antigen, hnRNP A2, p205, IgG, calpastatin, calreticulin, collagen and the shared HLA-DR epitope. The accumulation of factor--involving autoreactivities, cytokines, environmental and genetic factors--that challenge the normal regulatory mechanisms of the immune system lead to a regulatory catastrophe. In individuals developing the clinical features of RA the immune system has been regulated to a new--autoimmune--steady state. This attractor "rheumatoid arthritis" has many features of what has originally been described by Irun Cohen as the immunological homunculus: The healthy immune system is configured such as to direct its attention to major self-antigens. Thus it creates an autoreactivity to many autoantigens as a prerequisite for regulatory mechanisms that are sufficient to control them. The shift from the normal to rheumatoid attractor involves the inflammatory cytokines TNF-alpha, IL-1 and IL-6, autoreactive T- and B cells directed at a variety of synovial and systemic antigens, activated dendritic cells and macrophages, tissue destruction and genetic factors such as the association with shared epitope. Environmental factors involved may also, but do not necessarily, include infection. With the appearance of clinical features of RA, naive, potentially autoreactive T cells infiltrate the synovial compartment and become activated by dendritic cells and other APCs. The autoantigenic peptides that are presented to these T cells are derived from inflammatory cell and tissue destruction as well as from tissue repair and remodeling processes. These T cells proliferate and either provide help to B cells with the specificity to the same antigens or cause direct cytopathic tissue damage. Thereby, more and novel antigens are generated, released and presented again to naive or primed autoreactive T cells. These processes involving cytokines, tissue destruction and autoreactive T cells are sufficient to maintain RA even without the permanent presence of a triggering agent. The recursive autoimmune processes are well consistent with the finding of the many different autoreactivities in RA and their respective sensitivities and specificities. The massive influx of T cells into the arthritic joint is accompanied by the anergization of over 90% of T cells in this compartment--which further substantiates the concept of the RA attractor within the self-regulating immune system. Thereby, the RA-attracted immune system is not able to completely downregulate the inflammation and the local tissue damage/repair. Thus, the immune system is permanently stimulated and suddenly by chance shifts to a stable state different from the healthy system--reaching the wide fields of rheumatoid arthritis which in itself is self-sustaining as the healthy state before disease onset. PMID- 11263011 TI - [Are there effective dietary recommendations for patients with rheumatoid arthritis?]. AB - Patients with rheumatic diseases frequently ask the physician for diet recommendations. Although much has been written about this subject, scientifically validated studies investigating the impact of certain diets on rheumatoid arthritis are scant and often inconclusive. Elimination diets or total fasting is believed to eliminate food ingredients that cause or aggravate arthritis. In contrast, supplementation with fish oil, gamma-linoleic acid or vitamin E is directed at anti-oxidative and anti-inflammatory effects of these food compounds. So far, both approaches have failed to reveal a significant benefit with respect to objective signs of inflammation. Supplementation with other vitamins such as vitamin A and C, or with trace elements like selenium and zinc are of no proven influence on the disease activity as well. There is a higher request for calcium and vitamin D in patients with active RA under steroid treatment to prevent osteoporosis. In addition, patients with active RA have a slightly increased risk for cardiovascular events. Therefore, cholesterol lowering diets and drugs should be applied early. PMID- 11263012 TI - [Help status and help prospects of severely handicapped patients with rheumatoid arthritis]. AB - Although increased efforts have been made, rheumatoid arthritis still leads to severe disability and dependence on external help in about one-third of the patients. Relatively little is known on how help-dependent RA patients manage everyday life, to what extent they need help, by whom the help is given, what the patients' future help perspectives are and what kind of patient- and resource related characteristics are associated with unmet need. METHODS: By means of standardized interviews a representative sample of severly disabled RA patients was investigated with respect to present life situation, functional capacity, amount of help needed, relationship of caregiver and patient, health status of caregiver, stability of the help situation, housing preferences and help-seeking behaviour. The patients were classified as being either in definite need of help or independent of external help. RESULTS: Patients with comparable disabilities living on their own had only one-third of the help of those living together with others. They received help slightly more often from non-family caregivers, such as visiting nurses or privately paid household help, than from family members. Although many patients had rather uncertain perspectives for the future, the majority vehemently rejected living in a retirement home or in a home for the disabled. This refusal to live in an institution was independent of functional capacity, adequacy of the present help situation and the financial situation. Unmet need was highly associated with the age and health status of the caregiver as well as the help-seeking behaviour of the dependent person, but not with functional status or available resources. CONCLUSION: Given an appropriate home, the majority of conditions for a self-determined life for help-dependent RA patients are satisfied by the present outpatient help and care systems in Germany, but patients and their care givers need to be encouraged to take advantage of the available help and care facilities. PMID- 11263013 TI - [Listeria arthritis in chronic polyarthritis during low dose prednisolone and methotrexate therapy. Case report and review of the literature]. AB - We report about a patient with polyarticular rheumatoid arthritis taking methotrexat and 5 mg prednisolone who developed in the course of a RA flare a septic arthritis in the right shoulder. Listeria monocytogenes could be identified as the causative bacteria. Clinically, the Listeria-induced septic arthritis could not be differentiated from rheumatoid arthritis; fever was not present. The synovial analysis showed a granulocytic effusion with 19,000 cells/ml; there was no microbiological growth within the first 24 hours. Only the low glucose level indicated a possible septic arthritis. After 48 hours, gram positive bacterial growth was evident and Listeria monocytogenes could be isolated after 72 hours. Therapy was initiated by antibiotic treatment and arthrotomy with synovectomy followed by extensive irrigation which proved effective in bacterial elimination but joint destruction resulted. During the whole course, Listeria antibodies were negative and proved to be too insensitive. The incidence of Listeria-induced arthritis is very low; a review of the literature revealed only 24 reported cases. It occurs primarily in patients with rheumatic diseases under immunosuppression and in prosthetic joints. The diagnosis is based on cultural detection. It is important to cultivate synovial effusions for longer than 24 hours in order to identify Listeria. This is of relevance since Listeria serology is not sensitive. PMID- 11263014 TI - [5-HT3 receptor antagonists as a new therapeutic principle in rheumatology?]. PMID- 11263015 TI - The assessment and prevention of suicide for the 21st century: the Air Force's community awareness training model. AB - Professional practice standards and ethical obligations in the realm of suicide and risk management have been discussed for the last several decades. In the civilian sector, this discussion has taken the form of malpractice case law, the development of numerous assessment tools and practice guidelines for clinicians, and some attempts to describe possible models of prevention. In the Air Force, concern regarding suicide and risk management has evolved into a formalized program of community awareness and education that has been testing the boundaries of suicidal risk detection, assessment, and support facilitation. This article briefly describes this program, its success, and its implications for both active duty and civilian populations. Guidelines for mental health practice standards in risk management and suicide assessment are also discussed. PMID- 11263017 TI - Inpatient hospitalization. PMID- 11263016 TI - The contingency medical force: chronic challenge, new solution. AB - To keep pace with the changing requirements of the U.S. Army's combat doctrine, the U.S. Army Medical Department continually modifies its combat health support doctrine and unit organizations. This includes creating more capable, deployable, and mobile units. Unfortunately, as units become more capable, they become less mobile and deployable. As a result, striking a proper balance between capability, mobility, and deployability poses a significant challenge. In 1998, the 212th Mobile Army Surgical Hospital designed a rapidly deployable, air transportable medical module capable of supporting a brigade-sized contingency force (approximately 3,000 personnel) with level or echelon I to III medical care in an austere and ambiguous environment. This module, known as the contingency medical force (CMF), also provides command and control capabilities for this initial medical force and the transition to a more robust health care structure. Conducted over an 8-month period, the design process began with a staff exercise using the deliberate planning process model and culminated in a validation exercise monitored by external observers/controllers at the Combat Maneuver Training Center in Germany. This article describes the planning process, development, and initial deployment of the CMF. The CMF was then deployed on short notice to Albania in support of Task Force Hawk, the Army component of Joint Task Force Noble Anvil. PMID- 11263018 TI - Doxycycline-induced esophageal ulceration. PMID- 11263019 TI - An initiative to retain reserve soldiers failing to meet weight and physical fitness standards: the Wisconsin Army National Guard experience. AB - This paper presents the Wisconsin Army National Guard's attempt to retain soldiers failing to meet weight and annual physical fitness test standards. Soldiers failing or at risk of failing weight and fitness standards attend a wellness program one weekend per month for three consecutive months. Instruction includes topics in exercise training, nutrition, general wellness, stress reduction, and motivational lectures. A total of 324 soldiers who completed the program were evaluated for retention rates. At 48 months, graduates of the program had a 55% retention rate. This program is cost effective and soldier caring. PMID- 11263020 TI - The prevalence and importance of sexual concerns among female military beneficiaries. AB - BACKGROUND: Sexuality is an important part of health, quality of life, and general well-being, yet studies suggest that less than half of patients' sexual concerns are known by their physicians and that physicians are unaware of how common sexual concerns are among patients. The objective of this study was to determine the type and prevalence of sexual concerns among a randomly selected sample of women enrolled for health care at a military community hospital. METHODS: A randomized mail survey was used. Of 593 eligible participants, 232 responded (39%). Main outcome measures were self-reported sexual concerns and sociodemographic data. RESULTS: A total of 90.9% of women reported one or more sexual concerns. Most frequently reported sexual concerns were lack of interest (88.6%), difficulty with orgasm (81.2%), body image concerns (80.4%), inadequate lubrication (76.1%), dyspareunia (75.1%), needing information about sexual issues (75.1%), and unmet sexual needs (63.4%). More than half (65.2%) reported concerns of physical or sexual abuse, and more than half (53.1%) reported sexual coercion at some point in their lives. CONCLUSIONS: The results demonstrate a high prevalence of sexual health concerns for women enrolled for health care in a military community hospital. The implication for clinical practice is that sexual health inquiry should be a regular and important part of health care maintenance. PMID- 11263021 TI - Pain management in the special operations environment: regional anesthetics. AB - Pain relief is an essential component of combat casualty care. For the injured soldier, analgesia is not only a matter of comfort. Alleviating pain may allow the soldier to remain quiet when noise discipline is at a premium. It may also allow that person to continue to move, thus avoiding detection and potentially permitting the mission to carry on. Regional anesthetics provide an alternative to systemic medications and thus may avoid a clouded sensorium, limit narcotic administration, and provide superior pain relief. Standard local anesthetics and newer agents with potential field applicability are discussed along with their side effect profiles. Simple nerve block techniques that can be used by Army Special Forces medics, Navy SEAL and Reconnaissance corpsmen, and Air Force pararescuemen in the far forward environment are described step by step. The advantages of these regional anesthetic methods should make their use a must for every special operations medical care provider. PMID- 11263022 TI - Effect of fibrin bandage fibrinogen concentration on blood loss after grade V liver injury in swine. AB - OBJECTIVE: To determine the effect of fibrinogen concentration of dry fibrin bandages on blood loss after grade V liver injury. METHODS: Twenty-four pigs were used. Grade V liver injuries were induced and treated with dry fibrin bandages containing 0, 4, 8, or 15 mg fibrinogen/cm2. Animals were monitored for 60 minutes. Blood loss, fluid use, hematological data, and hemostasis were assessed. RESULTS: Post-treatment blood losses (mean and 95% confidence interval [CI]) were 1,560 mL (356-6,844), 372 mL (65-2,134), 225 mL (51-992), and 127 mL (22-732) in the 0-, 4-, 8-, and 15-mg groups, respectively. Only the 15-mg group had results significantly lower than the 0-mg group (p < 0.05). Blood loss was negatively related to fibrinogen concentration (p < 0.05). CONCLUSION: Fibrinogen concentration was inversely related to blood loss after grade V liver injury. The 15-mg formulation was the only one that significantly reduced blood loss. PMID- 11263023 TI - Functional gastrointestinal disorders among soldiers in peacetime versus out-of area missions. AB - Functional gastrointestinal syndromes are chronic disorders of the abdomen with an absence of organic findings. The aim of this study was to compare the frequency and symptomatology of functional abdominal syndromes in soldiers during an out-of-area mission versus during peacetime at home. We examined 124 soldiers who sought medical care for abdominal symptoms at the German Field Hospital Trogir, Croatia. The control group consisted of 113 soldiers who were referred with abdominal symptoms to the Central Hospital of German Armed Forces Koblenz, Germany. After excluding an organic disease, the diagnosis of a functional disorder was made. Fourteen percent of the Implementation Force soldiers had symptoms of a functional syndrome. At home, the frequency of functional gastrointestinal disorders was 50%, significantly greater than the rate during the out-of-area mission (p < 0.0001). We conclude that functional gastrointestinal disorders are more rare during out-of-area missions than during peacetime. They are probably as frequent away from home as at home, but "health care seeking" is less frequent under the stressful conditions of out-of-area missions. PMID- 11263024 TI - War abdominal trauma: usefulness of Penetrating Abdominal Trauma Index, Injury Severity Score, and number of injured abdominal organs as predictive factors. AB - OBJECTIVE: To analyze predictive factors for developing complications or lethal outcome in war abdominal trauma. DESIGN: Retrospective study. METHODS: We analyzed 93 cases of war penetrating abdominal trauma treated at the General Hospital Karlovac, Croatia. The following potential predictor variables were analyzed: age, sex, type of wound, Penetrating Abdominal Trauma Index (PATI), Injury Severity Score (ISS), and number of injured abdominal organs (NIAO). RESULTS: A total of 10.8% of wounded patients died and 25.8% developed complications. The overall average number of injured intra-abdominal organs was 2.0; in the group of patients with complications, the average was 3.0, and in the group of deceased patients, the average was 3.5. The most frequently injured organs were the small and large bowels. The significant predictors of developing complications as well as death outcome were the PATI, the ISS, and the NIAO. The best diagnostic efficiency (79.57%) for predicting complications was with the NIAO, whereas the best model for the prediction of death outcome combined all three variables (Z = -13.0776 + 0.1561 x PATI + 0.281 x ISS - 0.5234 x NIAO), with diagnostic efficiency of 92.47%. CONCLUSION: These models may be used as important prognostic factors in war abdominal injuries. PMID- 11263026 TI - Identifying hypertension using the Ohio Blood Pressure History Survey. AB - Although medical survey studies often rely on self-reported symptoms to establish the presence or absence of clinical conditions in respondents, recent findings suggest that surveys that assess a broad range of symptoms may have limited sensitivity in detecting specific clinical conditions such as hypertension. The present study evaluated the accuracy of a blood pressure history survey mailed to 800 men and women who had received treatment at a military medical facility in the previous year. Compared with their medical records, patient reports of a previous diagnosis of hypertension exhibited an overall accuracy of 94.2%. This high level of overall accuracy was associated with equally high proportions of correct identifications of high blood pressure histories (sensitivity = 95.4%) and normal blood pressure histories (specificity = 92.4%). Our findings indicate that the Ohio Blood Pressure History Survey is a highly accurate measure of hypertension history among active and retired military personnel. PMID- 11263025 TI - The impact of stroke on world leaders. AB - PURPOSE: Earlier studies by our unit documented frequent disability in world leaders resulting from stroke but did not quantify the incidence of cerebrovascular accidents. We sought to identify the frequency and impact of strokes in world leaders. METHODS: Using various sources, we identified world leaders who had sustained strokes while in office from 1970 to 1999 and tabulated information on symptoms and subsequent ability to lead. RESULTS: Twenty leaders were identified who had sustained strokes during the study period, for an incidence of 0.444 strokes/100 leaders/year. Half of the affected leaders lost their political power within the year; most had persistent disabilities, which included motor, speech, cognitive, and emotional deficits. CONCLUSION: Strokes in world leaders may be slightly less common than expected based on studies of Western populations of similar age, but they are often devastating to a political career. Nonetheless, loss of political power is not inevitable. PMID- 11263027 TI - Multiplexing for the detection of multiple biowarfare agents shows promise in the field. AB - Standard amplification of nucleic acids, or polymerase chain reaction (PCR), is replacing the more traditional microbiological assays in the detection of biological agents. However, standard PCR is designed as a one program-to-one agent amplification method, and not knowing what agents to test for makes this approach time consuming. During a field training exercise to detect four biowarfare agents using the standard PCR method, we conducted an additional experiment that reduced the diagnostic time to one-fourth. By reprogramming the four amplification programs to one program and preparing a cocktail containing the four different primer sets for the agents (known as multiplexing, or mPCR), we were able to amplify the four genetically different biological agents simultaneously. This is the first time a military unit has performed this kind of field testing, and it shows promise for the early detection of multiple biowarfare agents. PMID- 11263029 TI - Factors associated with depression on a hospital ship deployed during the Persian Gulf War. AB - Investigators surveyed health care providers (N = 250) deployed to the Persian Gulf on the USNS Comfort hospital ship days before the beginning of the Persian Gulf War in 1990. In this article, we identify factors associated with the development of depression during deployment. Age, gender, negative life events, stress from trauma-related work demands, and occupational experience with the dying and the dead were significant predictors of depression. Military training, although not associated with the experience of depression, was negatively correlated with concern about injury. PMID- 11263028 TI - Navy medicine: a health care leadership blueprint for the future. AB - Health care organizations face unprecedented fiscal, regulatory, and social challenges. Future executive leadership of health care organizations must be extremely effective for an organization to remain vital. To identify the characteristics that successful leaders of the future will require, 11 senior health care executives in the U.S. Navy Medical Department were invited to describe (1) desirable characteristics of future health care leaders, (2) how to identify individuals with executive potential, and (3) how to prepare individuals to become successful health care executives. The findings provide a blueprint for leadership development for leaders in health care organizations. PMID- 11263030 TI - Deep water running: an effective non-weightbearing exercise for the maintenance of land-based running performance. AB - Deep water running (DWR) has become a well-recognized from of cardiovascular conditioning for injured athletes and has been used successfully to maintain running performance. DWR provides for decreased stress and weightbearing to injured tissue and joints, allows for maintenance of cardiovascular fitness and a training effect, and offers greater specificity of exercise in relation to running. During a 22-month period, 181 active duty Army soldiers, placed on temporary profiles for injuries that precluded them from their regular weightbearing physical fitness activities, participated in a DWR program. Injuries to the back, knee, and ankle were the most common reasons for referral to the program. This article reviews the physiological characteristics of DWR, specifics of DWR program design, DWR mechanics, and the advantages of DWR over other aerobic forms of exercise to maintain land running performance in military personnel on temporary non-weightbearing profiles. PMID- 11263031 TI - Determination of daily living activities of retired officers. AB - The aim of this cross-sectional study was to assess the daily living activities and health status of retired military officers residing in Ankara, Turkey (N = 865). The study participants were scored according to their activities of daily living (ADL) and instrumental activities of daily living (IADL). For this purpose, ADL and IADL indexes were used. Groups formed as a result of the answers to these questionnaires were compared with each other according to their distinct features. It was found that 88% of the retired officers could perform all ADL and IADL without any help. With respect to performing IADL with disabilities, a statistically significant difference was found between married and unmarried retirees and between age groups (p < 0.01). The health status of retired officers and their ADL and IADL performance status were found to be better than those found in studies performed among other elderly population groups in Turkey. PMID- 11263032 TI - Abnormal eating behaviors in female reserve officer training corps cadets. AB - The objective of this study was to evaluate the prevalence of abnormal eating behaviors in Reserve Officer Training Corps (ROTC) female cadets. A total of 310 female ROTC cadets participated in a prospective, cross-sectional study during summer training at Fort Lewis, Washington. All subjects completed the Eating Disorder Inventory and a supplemental questionnaire. Because of time constraints, clinical interviews were not administered. Of the 310 ROTC cadets, 20% met the screening criteria for being at risk for an eating disorder. The cadets at risk for eating disorders had significantly higher Drive for Thinness, Bulimia, and Body Dissatisfaction subscale scores and were more dissatisfied with their weight than cadets not at risk. There was no significant difference in reported ideal body weight and exercise intensity between the two groups. In the female ROTC cadet population evaluated, 20% practiced abnormal eating behaviors and were at risk for developing an eating disorder. PMID- 11263033 TI - Treatment of severe thrombocytopenia in alloimmunized, transfusion-refractory patients. AB - A significant proportion of patients with hematologic malignancies who are exposed to multiple transfusions will develop alloantibodies to platelet human leukocyte antigens (HLA), resulting in poor responses to subsequent platelet transfusions. Transfusion of HLA-identical platelets is an effective method of platelet support in these patients, but perfectly HLA-matched platelets are often not available. In this paper, we review the recent literature on platelet transfusion support in alloimmunized individuals and illustrate alternative management strategies with cases from our own practice. PMID- 11263034 TI - Diagnosing exertional rhabdomyolysis: a brief review and report of two cases. AB - Exertional rhabdomyolysis is a potentially dangerous condition that involves release of intracellular contents from skeletal muscle in concentrations that may cause renal or other systemic complications. The purpose of the two case reports presented is to assist clinicians in recognizing this condition and in considering its predisposing factors. This paper describes two patients who, in the presence of several predisposing risk factors, developed exertional rhabdomyolysis. After diagnoses of rhabdomyolysis were reached, both patients were admitted to a local hospital for several days of monitoring and treatment. After 1 to 2 months of activity modification, both patients successfully resumed full physical activity and military duty. PMID- 11263035 TI - Delayed interval delivery military style. AB - INTRODUCTION: Intentionally delaying the delivery of a second twin from a previable state to a gestational age when survival is possible is a heroic measure whose outcome is unpredictable. We report the case of delayed interval delivery in a patient transported via air evacuation to a tertiary care center. CASE REPORT: A 31-year-old gravida 3 para 0020 at 18 weeks with a twin gestation presented to a medical treatment facility for evaluation of uterine cramps. She subsequently delivered Twin A. With cessation of labor, the patient was air evacuated to a medical center for continued care. Seven weeks later, she delivered a viable male infant. DISCUSSION: The treatment of multiple gestations presenting with preterm labor or rupture of membranes remains expectant. When delayed delivery of a previable second twin is undertaken, appropriate care includes the use of antibiotics, tocolytics, and cervical cerclage. Anticipation of preterm birth warrants continued care in a tertiary care center offering neonatal intensive care. PMID- 11263036 TI - [Multifocal intraocular lenses. A review]. AB - Modern cataract surgery has developed tremendously during the past 10-15 years. Improved surgical techniques, as well as improved implant materials and designs, have enlarged patient profiles and indications for cataract surgery. This also created much higher expectations from the patients' site. The loss of accommodation is loss of quality of life for presbyopic and especially young pseudophakic patients. Therefore cataract surgery with multifocal IOL implantation is not only of academic interest, but reflects demands and expectations of our patients. Multifocal IOLs have been implanted since 1986, starting with 2-3 zone refractive and diffractive designs. Due to surgical techniques of that time MIOL decentration and surgically induced astigmatism were possible complications. In addition reduced contrast sensitivity and increased glare were common problems of MIOL because of their optical principles. New developments in this field in recent years such as the multizonal, progressive refractive MIOL in combination with improved surgical techniques have overcome those initial problems. Therefore, modern multifocal IOLs can be considered not only for correction of aphakia but also for refractive purposes. PMID- 11263038 TI - [Antimicrobial decontamination of corneal donor material: infection prevention and quality assurance]. AB - BACKGROUND: Microbiological examinations in eye banks have found an increased contamination rate in preservation media. We studied the effect of prolonged submersion time for decontaminating donor globes. MATERIALS AND METHODS: We retrospectively analyzed the primary contamination of conjunctival smears in 76 cornea donors. The submersion time of donor eyes in PVP iodine solution before preparation was prolonged from 1 min to 5 min, and the contamination rate of storage vessels was compared. RESULTS: In 13 of the 76 conjunctival smears we found no contamination. Before prolonging submersion time, the preservation medium was contaminated in 15 cases, but after 5 min no contamination was observed. CONCLUSION: Prolonging the submersion time of donor globes from 1 to 5 min was effective. The model presented here provides guidelines for existing eye banks as well as for those yet to be established. PMID- 11263037 TI - [Morphological evaluation of posterior capsule opacification in diffractive multifocal intraocular lenses]. AB - BACKGROUND: Diffractive multifocals belong to the first generation of multifocal intraocular lenses. Dure to their optical principle of diffraction multifocals separate the incoming light on two foci (41%) with 18% loss of scattered light. Therefore, reduced contrast sensitivity and glare have been frequently described with this lens. PATIENTS AND METHODS: We evaluated 42 eyes of 25 patients (age at surgery 63.8 +/- 8.8 years) 3 years after implantation of a Pharmacia 811E diffractive multifocal (MIOL). They were tested for functional results including contrast and glare (Mesoptometer II). In addition 22 eyes were evaluated for posterior capsule opacification (PCO) using digitalized retroilluminations photographs and the EPCP image analysis program. RESULTS: Average visual acuity was 0.77 +/- 0.20 (uncorrected distance), 0.93 +/- 0.21 (corrected distance), 0.94 +/- 0.13 uncorrected near) und 0.98 +/- 0.08 (best corrected near). PCO values quantified by EPCO were 1.13 +/- 0.59 (Range 0.07 to 2.1). On average 18.1 +/- 14.9% of the optic area showed an opacification grade 1, 35.8 +/- 26.6% grade 2 and 7.7 +/- 17.1% grade 3. Contrast sensitivity showed no patient with contrast level 7 or better for monocular evaluation, but 30% for binocular testing. Only 4% met the criteria for night driving when tested for glare. Spearman correlation did not reveal any significant correlation between PCO-values and visual acuity, contrast sensitivity or glare (all p > 0.24, all r < 0.22). CONCLUSIONS: Patients with diffractive multifocal IOLs showed excellent results for near and distance visual acuity three years after implantation. However, contrast and glare results were poor, probably due to a rather high PCO-rate. Future developments of the lens type should include PCO-reducing factors, such as sharp edge optics and foldable materials. PMID- 11263039 TI - [Keratography (corneal tattooing) in corneal leukoma]. AB - BACKGROUND: Since antiquity attempts have been made to minimize disfigurement and stigmatization of patients with leukoma. Keratography is a relatively new method for imprinting color pigments into the corneal stroma with an entomological needle. MATERIALS AND METHODS: Keratography was performed in 20 patients at the University Eye Hospital, Munich, between November 1997 and September 1999. Patients had either a leukoma that did not tolerate prothesis or had another cloudy corneal disease. The operation was carried out in our outpatient clinic under local anesthesia. RESULTS: There was a single operation in three patients, two operations in nine, three operations in six, and four operations in two. Postoperatively 33% of patients complained of pain. No postoperative bacterial keratitis or perforation occurred. CONCLUSION: All patients were highly satisfied. The long-term stability of color pigments must still be evaluated. PMID- 11263040 TI - [Examination of the cornea using optical coherence tomography]. AB - INTRODUCTION: This study evaluated the clinical use of optical coherence tomography (OCT) for two-dimensional representation of the cornea. PATIENTS AND METHODS: Noncontact slitlamp-adapted OCT was used in selected cases to evaluate pathologically altered corneas and to measure the central corneal thickness and curvature. RESULTS: OCT provided correlation between differences in reflection and morphological changes. Scar tissue resulted in hyper-reflective light scattering, whereas cystic lesions were hyporeflective. Precise biomorphometry also allowed representation of intrastromal and retrocorneal changes. Central corneal thickness measured by OCT yielded reproducible values and corneal curvature could be calculated from the optical signals of the corneal surface. CONCLUSIONS: OCT provides high-resolution representation of the cornea and exact evaluation of its morphology, thickness, and curvature. Due to the noncontact, simple, and rapid examination using the slitlamp the corneal OCT method is a promising additional diagnostic modality. PMID- 11263041 TI - [Determination of retinal thickness in relation to the age and axial length using optical coherence tomography]. AB - PURPOSE: It is unknown whether the thickness of the retina depends on axial length or on age. We therefore used optical coherence tomography (OCT) to study this relationship. METHODS AND MATERIALS: We recruited 159 subjects aged 13-92 years (205 eyes) without macular pathology. OCT measurements included three horizontal scans and one vertical scan through the fovea. Axial length was determined by an analog high-resolution biometric unit. RESULTS: There was no correlation between retinal thickness and either axial length or age. Mean retinal thickness in the fovea was 142 +/- 18 microns. In the nasal retina thickness was significantly increased to 266 +/- 17 microns, compared to 249 +/- 18 microns in the temporal retina. Retinal thickness in subjects' two eyes was significantly correlated. CONCLUSIONS: Since retinal thickness does not depend upon age or length of the eye, no corrections are necessary when analyzing pathological retinal thickening, such as in diabetic retinal disease. PMID- 11263042 TI - [Cavitation bubble formation during Erbium:YAG laser vitrectomy]. AB - BACKGROUND: Clinical and experimental studies demonstrated the potential advantages of Erbium:YAG laser vitrectomy for posterior segment surgery. However, a detailed knowledge on the laser-tissue interaction is needed for an optimization of new ophthalmic applications. The aim of this experimental work was to investigate the cavitation bubble formation during Erbium:YAG laser vitrectomy and to find optimized laser parameters and the best geometry of the aspiration port of the microsurgical probe. MATERIALS AND METHODS: We investigated the formation of cavitation bubbles in water by high-speed photography. The output energy at the quartz tip reached up to 50 mJ and the laser pulse duration ranged from 50 to 300 microseconds. Various commercially available microsurgery probes were investigated regarding the extent of the cavitation bubbles. RESULTS: The threshold for laser-induced cavitation bubble formation was found to be 0.32 +/- 0.1 mJ for a laser pulse duration of 130 microseconds and a core diameter of 320 microns of the quartz fiber tip. The length of the cavitation bubbles increases with the laser pulse energy up to a length of 1.6 mm at a pulse energy of 10 mJ. In contrast, the size of the vapor bubbles decreases with an increase of the laser pulse duration. A slit-shaped aspiration port led to a 50% smaller volume of the cavitation bubble exiting the port compared with a circular aspiration port. CONCLUSIONS: The optimized laser parameters and microsurgery probe geometry may significantly decrease the risk of intraoperative ocular damages during Erbium:YAG laser vitrectomy. PMID- 11263043 TI - [Automated kinetic perimetry using different stimulus velocities]. AB - INTRODUCTION: Because kinetic perimetry is performed manually, it is difficult to standardize. This study evaluated the effect of different velocities of various stimuli on the results of automated kinetic perimetry. MATERIALS AND METHODS: Twelve ophthalmologically healthy subjects were tested with stimuli III4, I4, I2, I1 using the Twinfield perimeter at velocities of 1-7 degrees/s. RESULTS: Results of stimuli III4 and I4 were not dependent on velocity. Concerning stimuli I2 and especially I1 the isopters were narrower for increasing velocity, and variance in results was greater above 4 degrees/s. CONCLUSION: The recommended velocity of 2 degrees/s is often exceeded in daily clinical practice, which explains part of the variation in findings between studies. Automated kinetic perimetry could ensure a constant, slower velocity and thus improve standardization. PMID- 11263044 TI - [Laser-assisted transcanalicular dacryocystorhinostomy. Initial results]. AB - BACKGROUND: Endoscopes for direct examination of the mucosa and disorders of the lacrimal drainage system have been available for four years. This had led to the wish to be able to treat lacrimal disorders by endoscopes and laser. PATIENTS AND METHODS: Since September 1997 we have treated 48 patients by laser-assisted transcanalicular dacryocystorhinostomy (DCR) using a KTP laser, including all those with stenosis to the nasolacrimal duct and all those with postsaccal stenosis. A bony window with diameter of 5 x 5 mm was created. Bicanalicular intubation into the nose was performed and was left for 3-6 months. RESULTS: We found 40 patients to have patent lacrimal pathway and no symptoms, 4 watering in cold weather, and 4 a restenosis. CONCLUSIONS: The KTP-laser is sufficiently powerful to create bony windows at least 5 mm in diameter, and a transcanalicular laser-assisted DCR is therefore possible. PMID- 11263045 TI - [Autogenous bone transplants in repair of the traumatically damaged orbit. Functional and esthetic results in 42 cases]. AB - BACKGROUND: Autogenous bone transplantation is a well established aspect of craniofacial traumatology. This study evaluated functional and esthetic success in repairing traumatically damaged orbits by autogenous bone transplantation. PATIENTS AND METHODS: We repaired 102 osseous orbital defects by autogenous bone transplantation, 98 by calvarium split graft and 4 by iliac crest transplant. Of 42 surgical patients 34 were followed up. RESULTS: Autogenous bone transplants have proven themselves for repair of traumatically damaged orbits. Calvarium split grafts are particularly suitable for this purpose. Good esthetic and functional results can usually be achieved with primary osseous defect repair. However, more protracted functional defects can often no longer be helped. CONCLUSION: Autogenous bone transplants can be broadly recommended for providing primary care. PMID- 11263046 TI - [Immunology-related chronic progressive cicatricial conjunctival diseases: diagnosis, therapy and prognosis]. AB - PURPOSE: We reviewed the records of patients with chronic cicatricial disease of the conjunctiva for differences in prognosis between clinical and histopathological subgroups of the disease and in therapeutic options. PATIENTS: In 30 patients (58 eyes) with an average age of 69 years (52-86) chronic cicatricial disease of the conjunctiva was diagnosed clinically. Only in 22/30 patients conjunctival biopsies could be performed. The correlation of histopathological and immunohistochemical diagnoses with clinical course under systemic immunosuppression was studied. RESULTS: 15/22 biopsies led to a classification into different subgroups. Under systemic immunosuppression disease ceased to progress for a mean time of 15 months in 13 of 15 patients with positive biopsies and in 9 of 15 without classification. The results after cyclosporine A therapy (4 of 5 patients stabilized after a mean of 27.5 months) and mycophenolate mofetil (8 of 11 patients stabilized at a mean of 7.8 months) were better than those after therapy with dapsone, azathioprine and cyclophosphamide. CONCLUSIONS: Histopathological and immunohistochemical examinations led to a classification in two-thirds of the patients with clinical aspects of chronic cicatricial disease of the conjunctiva. There was no correlation between different histopathological subgroups, success of therapy and prognosis of the disease. There is little hope in using new systemic immunosuppression such as cyclosporine A and mycophenolate mofetil. PMID- 11263047 TI - [Quality management according to DIN EN ISO 9001 at a university eye hospital]. AB - BACKGROUND: Quality assurance is an integral part of modern microsurgical ophthalmology. Health care laws also mandate overall quality management. MATERIALS AND METHODS: In recent years we have standardized the preexisting features of quality management according DIN EN ISO 9001 and have integrated previously missing features. RESULTS: Establishing quality management according to ISO 9001 is possible even at a university eye hospital and department of ophthalmology. Certification according to ISO 9001 specifications was granted in April 1999. The major difficulty was in translating industrial norms to the context of an eye hospital. It was also difficult to overcome skepticism towards quality-assurance measures that lie beyond ophthalmological quality control. CONCLUSION: It is useful and feasible to establish a quality management system at German university eye hospitals and departments of ophthalmology. Certification according to ISO 9001 is one possibility to make a quality management system transparent and evaluable both inside and outside the hospital. PMID- 11263048 TI - [Transscleral and transpupillary laser coagulation in proliferative sickle-cell retinopathy]. AB - BACKGROUND: Transscleral diode laser photocoagulation is a new method for treating proliferative sickle-cell retinopathy (PSR). Other treatments include transpupillary laser photocoagulation and transscleral cryocoagulation CASE REPORT: We report two patients with sickle-cell disease and PSR, one treated by transpupillary argon laser coagulation and the other by transscleral diode laser scatter photocoagulation. CONCLUSION: Transpupillary and transscleral laser photocoagulation are effective and safe in the treatment of PSR. Transscleral photocoagulation is an alternative treatment method in eyes with PSR and cloudy optical media or with poor mydriasis in which transpupillary coagulation cannot be performed. PMID- 11263049 TI - [Intracorneal granulomatous inflammation]. AB - INTRODUCTION: We report a patient suffering from lattice corneal dystrophy and with a corneal granuloma 8 months after phototherapeutic keratectomy (PTK). CASE REPORT: PTK was performed in a 33-year-old man with hereditary lattice corneal dystrophy. An intracorneal tumor was found in the right eye 8 months after treatment. There was no indication of previous foreign body injury. Detailed clinical examinations yielded no evidence of the cause or nature of the tumor. For this reason an excision was performed. RESULTS: Histological examination revealed a dense inflammatory infiltration of the deeper corneal stroma with epithelioid cells, histiocytes, lymphocytes, eosinophilic leukocytes, and giant cells of the Touton type. There was no indication of a generalized granulomatous inflammation. CONCLUSION: A monosymptomatic, intracorneal, granulomatous infiltration is very rare and has not previously been described in a case of lattice corneal dystrophy. This may have been a case of xanthogranuloma or sarcoidosis. Strangely, the granuloma occurred only several months after PTK; nevertheless, a pathogenetic connection seems unlikely. PMID- 11263050 TI - [Posterior uveitis: sarcoidosis or tuberculosis]. AB - PURPOSE: To demonstrate the difficulties of the differential diagnosis between tuberculosis and sarcoidosis as the cause of posterior uveitis. CASE REPORTS: A 56-year-old woman suffered from bilateral anterior uveitis, snow-ball like infiltrates in the vitreous, and peripheral retinochoroidal granulomas with marked exudation shown in fluorescein angiography. Angiotensin-converting enzyme, as a marker of sarcoidosis, was elevated; the tuberculin test, however, was negative. Chest X-ray revealed an infiltrate and numerous smaller granulomas. The presumptive diagnosis was sarcoidosis. Surprisingly, in the biopsy of the pulmonal lesion tubercle bacilli were detected by Ziehl-Neelsen staining. Thus, a diagnosis of pulmonal and also retinochoroidal tuberculosis was made. After tuberculostatic therapy the choroidal lesions healed off. In a second case, a 30 year-old man suffered from bilateral panuveitis with candle wax exudates near the retinal vessels. Chest X-ray revealed lymphomas in the hilus, and the lymph node biopsy showed granulomas with epitheloid cells, indicating sarcoidosis. Detection of mycobacterium tuberculosis by culture or histological criteria was negative. Only in the PCR was mycobacterium tuberculosis DNA detectable. Tuberculostatic therapy had no benefit. Under therapy with steroids, however, pulmonal and ophthalmologic findings rapidly disappeared. CONCLUSION: The difficult differential diagnosis between sarcoidosis and tuberculosis cannot always be made by laboratory tests or diagnostic imaging alone. Clinical manifestations, including response to therapy, are essential. PMID- 11263052 TI - [Multifocal ERG in glimmer vision]. PMID- 11263051 TI - [Acute papilledema. A 69-year-old patient with acute bilateral papilledema]. PMID- 11263054 TI - Worker autonomy: the foundation of shared governance. PMID- 11263053 TI - [Photodynamic therapy: Recommendations for indication and treatment]. PMID- 11263055 TI - Administrative support for addressing staff nurses' ethical concerns regarding staffing. PMID- 11263056 TI - Is it time to eliminate nursing education departments? PMID- 11263057 TI - Honoring commitments. PMID- 11263058 TI - Determining indirect caregivers' contribution to patient care. AB - It is critical to measure contributions of staff who are not direct caregivers so that organizations can continually maximize resource. Understanding the consumer's perceptions is necessary to identify activities that contribute to the perception of an effective CNE. An awareness of the values placed by consumers on education services helps the educators to focus their efforts on those activities with the greatest perceived value. If educators put their efforts toward valued activities, their services would be frequently used and the staff would be more willing to accept the information, thereby benefiting from the educator's expertise. Learning how a role directly benefits patient care helps nurses in leadership positions meet institutional objectives. It allows nurse leaders to ensure that the role is, in fact, contributing to care, and it is doing so to the fullest extent. This builds institutional support and value for the role. The process of evaluating the benefit also allows the educators to gain support and credibility among consumers and other individuals within the institution. This perpetuates the increased utilization and benefit of the role. Results of this project can be used as a guide in evaluating various roles. Understanding activities that are valued by consumers enables staff in the roles that are being evaluated to determine how and where they should focus their efforts. This is especially critical as staff is being asked to do more work with less time and must establish priorities in their ongoing workload. Finally, it is vital to identify ways of turning the invisible work that indirect caregivers provide into visible work. PMID- 11263059 TI - Care centered organizations, Part 2. The changing role of the nurse executives. AB - In the first (February 2001) of a three-part series on the role of nursing in care centered organizations, the rationale for establishing service lines, the process for determining governance structure, and the changing role of nurse executives in a large academic medical center reorganized around service-line care delivery were discussed. In this second article, the author describes the nursing governance structure in two academic healthcare institutions after their redesign into care centers. PMID- 11263060 TI - Lessons learned while collecting ANA indicator data. AB - Realizing the importance of linking nursing's contribution to quality patient care, a pilot study was conducted to determine whether data regarding the quality indicators proposed by the American Nurses' Association (ANA) could be collected from five acute-care inpatient units at one medical center that is part of a multisite managed care system. Although it was determined that data regarding the ANA quality indicators could be collected at the study site, a variety of unanticipated findings emerged. These findings reflect both discrepancies and congruities between how the investigative team expected the ANA indicators to operate versus what was actually experienced. The lessons learned while collecting ANA indicator data are shared to assist future users and to advance the evolution of the ANA indicators. PMID- 11263061 TI - Lessons learned while collecting ANA indicator data. The American Nurses Association responds. AB - In 1994, the American Nurses Association (ANA) began a multiphase initiative known as Nursing's Safety and Quality Initiative (the Initiative) to address concerns about patient safety and quality of care. The Initiative has three basic components: research, education, and policy. PMID- 11263062 TI - What should we expect from California's minimum nurse staffing legislation? AB - In 1999, California passed the first legislation in the United States to establish minimum staffing levels in hospitals for registered nurses (RNs) and licensed vocational nurses. The author provides estimates of the increase in RN expenditures required by this mandate, by hospital size and for regions of California. Issues related to the implementation of minimum ratios also are discussed. Attention must be paid to other staffing regulations, special concerns of rural hospitals, the possibility that minimum ratios result in lower RN staffing, and the effect of the nursing shortage on the ability of hospitals to meet requirements. PMID- 11263063 TI - BSN by 2010. A California initiative. AB - The Association of California Nurse Leaders has developed an initiative to require the baccalaureate in nursing as the credential for entry into practice as a registered nurse by the year 2010 in the state of California. When nursing is compared to other healthcare professions, such as pharmacy, physical therapy, and occupational therapy, it becomes obvious that educational requirements for nurses must be updated. Nursing leaders have developed a 10-year action plan to change the entry-level educational requirements for California nurses. PMID- 11263065 TI - Fundamentals of capillary electrochromatography. PMID- 11263064 TI - Tel-eNurse Practice. Quality of care and patient outcomes. AB - As a growing specialty, telephone nursing practice requires definition, standardization, and identification of quality indicators and nursing-sensitive outcomes. An informal study was conducted to explore the relationship between telephone nursing quality indicators-assessment, critical thinking, use of protocols, and continuity of care-found in documentation and nursing-sensitive patient outcomes. Findings provide insight into the telephone process of care and application of critical thinking reflected in documentation and greater understanding of the complexity of telephone nursing practice and integration of care and outcomes. PMID- 11263066 TI - Molecularly imprinted extraction materials for highly selective sample cleanup and analyte enrichment. PMID- 11263067 TI - Biomembrane chromatography: application to purification and biomolecule-membrane interactions. PMID- 11263069 TI - High-performance liquid chromatography: trace metal determination and speciation. PMID- 11263068 TI - Transformation of analytes for electrochemical detection: a review of chemical and physical approaches. PMID- 11263070 TI - Temperature-responsive chromatography. PMID- 11263071 TI - Carrier gas in capillary gas-liquid chromatography. PMID- 11263073 TI - Membrane extraction techniques for sample preparation. PMID- 11263072 TI - Catechins in tea: chemistry and analysis. PMID- 11263074 TI - Design of rapid gradient methods for the analysis of combinatorial chemistry libraries and the preparation of pure compounds. PMID- 11263075 TI - [Clinical and genetic consequences of ring chromosome, a structural genome mutation]. AB - Ring chromosome is a peculiar genetic anomaly which may cause phenotypic abnormalities even without a loss of genetic material. The ring formation of chromosome ends induces difficulties in the sister chromatid separation at cell division, resulting in increased cell death rate through the generation of secondary aneuploid cells. Based on the in vitro analyses of own cases and a review of more than 200 case reports published in the literature, the author presents some conclusions on "ring syndrome", a non-specific clinical manifestation of ring formation of any chromosome, consisting of the lack of (or poorly expressed) organic manifestations, mild or borderline mental retardation, and a severe growth failure which is not explained by organic, biochemical, or endocrine findings. It is also suggested that the phenotypic manifestation of a ring chromosome is even less expressed when the ring is transmitted to the next generation. Based on observations of ring behaviour and the clinical phenotypes in a 3-generational family with a mosaic supernumerary ring, it is not unrealistic to assume that there is a transgenerational increase in the severity of the clinical manifestation resulting from a less efficient elimination process against the ring (cytogenetic anticipation). PMID- 11263076 TI - [The use of vinpocetine in chronic disorders caused by cerebral hypoperfusion]. AB - The clinical signs and symptoms of so-called "cerebrovascular insufficiency" or "cerebral vascular dysfunction" have the characteristics of those of chronic cerebral hypoperfusion. The clinical features of chronic cerebral hypoperfusion often show the symptoms of cognitive impairment and organic psychosyndromes. Cerebral hypoperfusion could be found in dementias of different origin (subcortical arteriosclerotic leucoencephalopathy [Binswanger], vascular dementia, Alzheimer's disease, etc.). Pathological changes caused by chronic cerebral hypoperfusion often confined only to the white matter (demyelisation, glial activation, damage of oligodendroglial cells, as well as scattered cell death). Each therapy has an influence on the biochemical and pathophysiological alterations caused by chronic cerebral hypoperfusion can be used with reason in these disorders. The mechanism of action of vinpocetine is interfering on many aspects with the biochemical and pathophysiological processes attributable to chronic cerebral hypoperfusion, independently of the original alteration responsible for hypoperfusion. This fact might give an explanation on the beneficial effect of vinpocetine on clinical signs and symptoms of chronic cerebrovascular insufficiency. PMID- 11263077 TI - [New aspects in the classification of cutaneous lymphomas]. AB - Authors discuss the classification of primary cutaneous lymphomas created by the Cutaneous Lymphoma Study Group of the European Organization for Research and Treatment of Cancer (EORTC) in 1996, which is based on the clinical, histological, immunohistochemical and genetic features of cutaneous lymphomas. Unlike the previous histologic classifications it contains well-defined disease entities characterized by their clinical and histological picture, clinical outcome, behaviour and therapeutic response. This classification does not use the term of low grade or high grade lymphoma, but introduces the indolent, aggressive and provisional subgroups in the T-cell lymphomas, and indolent, intermediate and provisional subgroups in the B-cell group. Authors demonstrate the EORTC classification by their own cases calling the attention to the clinical and therapeutic difference between nodal and extranodal lymphomas, and discuss the up to-date therapeutic possibilities. PMID- 11263078 TI - [Meningeal melanocytoma]. AB - The authors report on a case of melanocytoma surgically removed from the craniocervical meninges of a 59-year-old man. Although the excision had been incomplete, the patient showed a disease-free course extending well over ten years. On histology, the tumor consisted of moderately cellular arrays of spindle shaped melanocytes with a vaguely angiocentric whorling tendency, and without evidence of infiltrative growth. Electron microscopy identified tumor cells as ones bearing dendritic processes with complex melanosomes. The latter showed histochemical properties of melanosomal melanin, as well as immunoreactivity for the melanosome-associated markers HMB-45, and MELAN-A. Hallmarks of meningial differentiation were, at the same time, absent. The MIB-1 proliferation rate of the lesion, as assessed in a simultaneous testing of a panel including primary and metastatic central nervous system melanomas, as well as a uveal melanoma remained inferior to 1.5 percent. The data presented and a critical review of the literature suggest that meningeal melanocytoma is a mostly benign nevus-like lesion of neural crest cells with a very limited, although not discountable, margin for aggressive growth. PMID- 11263079 TI - [Life, curses and healing in the Kalevala]. PMID- 11263080 TI - [Relations between Frigyes Verzar and the Nobel Prize winner Wilhelm Einthoven, the inventor of the galvanometer]. PMID- 11263081 TI - [Madarasz Street Children's Hospital--past, present and future]. PMID- 11263082 TI - Fish saga continues. PMID- 11263083 TI - In one HealthNews article, you wrote that vitamin C can lower your blood pressure, and in another you said that it increases the intima-media thickness (IMT), thereby increasing the risk of heart disease. Should I or shouldn't I take extra vitamin C? PMID- 11263084 TI - My doctor prescribed a brand-name drug, but the pharmacist filled the prescription with a generic version. Will the generic work, and is it safe? PMID- 11263086 TI - Understanding off-label drug usage. PMID- 11263085 TI - What is urinary incontinence? PMID- 11263087 TI - More good news on glucosamine. PMID- 11263088 TI - Radiation routs restenosis. PMID- 11263089 TI - Listing Listeria's health risks. PMID- 11263090 TI - High blood pressure: two types? PMID- 11263091 TI - Hip fractures: do preventive drugs work for all? PMID- 11263092 TI - Timing is key when treating SAD. PMID- 11263093 TI - New IUD approved. PMID- 11263094 TI - Hair dye and cancer. PMID- 11263095 TI - Taking control of menopause. PMID- 11263096 TI - Surgeon-performed ultrasound imaging in acute surgical disorders. AB - As the role of the general surgeon continues to evolve, the surgeon's use of ultrasound imaging will surely influence practice patterns, particularly for the evaluation of patients in the acute setting. With the use of real-time imaging, the surgeon receives "instantaneous" information to augment the physical examination, to narrow the differential diagnosis, or to initiate an intervention. With select ultrasound examinations, the surgeon can rapidly evaluate adult and pediatric patients with an acute abdomen, especially those patients who are hypotensive. In the hands of the surgeon, this noninvasive, bedside tool can assess more accurately the presence, depth, and extent of an abscess, confirm complete aspiration, or diagnose wound dehiscence before it is apparent on physical examination. Ultrasound imaging is so accurate for the diagnosis of pyloric stenosis that it has essentially replaced the upper gastrointestinal series in most institutions. The surgeon's use of ultrasound imaging to detect a pleural effusion has virtually supplanted the lateral decubitus radiograph. Furthermore, an ultrasound-guided thoracentesis not only facilitates the procedure but improves its safety. As surgeons become more facile with ultrasound imaging, it is anticipated that other uses will develop to further enhance its value for the assessment of patients in the acute setting. PMID- 11263097 TI - An update on the Americans with Disabilities Act: implications for health and human services delivery. AB - The Americans with Disabilities Act (ADA) was signed into law nearly a decade ago. The ADA is designed to protect persons with disabilities from discrimination in nearly every aspect of American life. This article focuses on a number of recent court rulings that help to clarify important components of the ADA. It is critical for health and human services professionals to be aware of these recent rulings because they affect the delivery of services. It is concluded that professionals working in the health and human services should have ongoing training and education concerning the changing legal interpretations of the ADA. Many of these changes have important implications for service delivery. PMID- 11263098 TI - A life cycle model of public policy issues in health care: the importance of strategic issues management. AB - Public policy affects health and social services organizations. Senior management has a responsibility to prevent inappropriate demands of stakeholders from predominating and to influence the outcome of public policy to the benefit of their organization through the strategic issues management process. This article presents a public policy issue life cycle model, life-cycle stages and suggested strategies, paths issues can take in the life cycle, and factors that affect issue paths. An understanding of these dynamics can aid senior managers in shaping and changing public policy issues and lessening external environment threats to their organization. PMID- 11263099 TI - Down and out in America: children and health care. AB - One of the worst forms of inequality in the health care field is the inequality suffered by children. In 1996, over 14.5 million children lived in poverty in the United States. Children who live in poverty are less likely to have health insurance and have less access to health care and thus are more likely to suffer negative outcomes in health care. The Congress of the United States in 1997 enacted the State Children's Health Insurance Program (S-CHIP) to expand health insurance coverage for children. This paper examines the major features of the program, actions undertaken by the state governments under this program to expand health insurance coverage for children, and provides some preliminary analysis of the potential positive and negative impact of the program. PMID- 11263100 TI - Access to primary care: the role of race and income. AB - The study has two objectives: (1) to examine the racial differences in access provided by Municipal Health Services Clinics to the Medicare low-income beneficiaries, i.e., those also eligible for Medicaid (dual eligibles), and compare it with access provided to non-dually eligible patients; (2) to examine the racial and income disparities in access for primary and ancillary care services. The Municipal Health Services Program (MHSP) started in 1979 in five cities with the objective to improve access to primary care services. The study's method for measuring access combines use and need of care in a single index. The study finds that the clinics provided better access to dual eligibles than to non dual eligibles and to nonwhite than to white dual eligibles. However, this was a result of higher use of ancillary services by the clinic nonwhite patients. This was particularly true in large clinics such as Baltimore, where inadequate targeting of the low-income group and higher ancillary use were more significant problems than racial disparity in access. PMID- 11263101 TI - Providing behavioral incentives for improved health in aging and Medicare cost control: a policy proposal for Universal Medical Savings Accounts. AB - This paper examines policy options for addressing health care challenges posed by the aging of the baby boom generation. Universal Medical Savings Accounts (UMSAs) are proposed. UMSAs are defined-contribution vouchers coupled with medical savings accounts. The proposal includes significant equity protections for those with low income/wealth, including balance billing limits and stop-loss protections, together with subsidies for risk-adjustment. The policy would control costs while promoting quality, accessible, and affordable health care. UMSAs provide new behavioral incentives, both for cost-conscious health care decision-making and for healthy lifestyle choices. PMID- 11263102 TI - Lawrence-Douglas County Community Health Facility. Lawrence, Kansas. PMID- 11263103 TI - Hire ups. PMID- 11263104 TI - Dispense account. PMID- 11263105 TI - Spotlight. Managed care systems. PMID- 11263107 TI - I-medicine and the radiologist. PMID- 11263106 TI - Linking affiliates electronically. AB - PROBLEM: Store-and-forward clinical messaging system was cumbersome and inadequate to support modern technology. SOLUTION: Installation of a secure Web based system that automates communication among physicians, hospitals, clinical laboratories and other departments. RESULTS: Time savings for office personnel and physicians; access by physicians to clinical information anytime, anywhere; consolidation of lab results into one report; customized form--filing capability. KEY TO SUCCESS: Comprehensive and ongoing training to ensure user acceptance. PMID- 11263108 TI - PET screening of carotid occlusion. PMID- 11263109 TI - Looking within: Part 5. "Roentgen sure has gone crazy". PMID- 11263110 TI - Developing your strategy. PMID- 11263111 TI - Collective identity. PMID- 11263112 TI - Identifying bone mass and muscular changes. AB - The aim of this investigation was to examine the impact of a six-month high intensity strength-training program on lumbar bone mineral density (BMD), trunk and lower limb strength in a population of Australian women aged 50 years and over. A subject pool of 44 women were recruited and randomly allocated into either strength training (n = 19) or active control (n = 25) groups. All subjects trained twice weekly in either a 50 minute supervised strength training session that progressed from 60% one repetition maximum (1RM) to 90% 1RM or a 50 minute group walk session. Measurements included a lumbar (L2-L4) BMD scan: peak isokinetic trunk strength and a dynamic 1RM squat as a measure of lower body strength. No significant group differences in lumbar BMD were evident at the completion of training. However, a significant (p < 0.05) within group change was apparent for the active control group as lumbar BMD decreased 1.7% below baseline testing. A significant (p < 0.05) group difference was evident with the strength trained group increasing peak isokinetic trunk strength (19.3%) and 1RM squat strength (34.4%) above that of the active controls. It was concluded that strength training provides an effective means for increasing trunk and lower limb strength in women over 50 years. The impact of strength training on lumbar BMD was not conclusive in the present study. PMID- 11263114 TI - What makes surgical services successful at the top hospitals. PMID- 11263115 TI - Two hospitals named best workplaces. PMID- 11263113 TI - Shortage of anesthesia staff curtails surgery, pain services. PMID- 11263116 TI - Pulling the staff back from burnout. PMID- 11263117 TI - Managing cases in the afternoon. PMID- 11263118 TI - NPO status: have ASCs changed policies? PMID- 11263119 TI - Planning for HIPAA's privacy rule. PMID- 11263120 TI - 'Education day' is update for ASC staff. PMID- 11263121 TI - To treat or not to treat? Identifying ethical dilemmas in EMS. PMID- 11263122 TI - Sub-Q emphysema. Understanding the pathophysiology of subcutaneous emphysema. PMID- 11263123 TI - Low-flow oxygen for stroke patients. PMID- 11263124 TI - Doubts about Ipecac. PMID- 11263125 TI - Who's to blame? PMID- 11263126 TI - Australian medications. PMID- 11263127 TI - Take a 2nd look at your first impression. PMID- 11263128 TI - Conquer difficult airways. Strategies to help you identify & control a problem airway. PMID- 11263129 TI - Rescuing intubation. Simple techniques to improve airway visualization. PMID- 11263131 TI - Tragedy, terror & triumph. PMID- 11263130 TI - Crackdown on crics. Needle cricthryotomy & percutaneous transtracheal jet ventilation explained. PMID- 11263132 TI - Guide to prehospital surgical cricothyrotomy. PMID- 11263134 TI - Talk radio. The art of verbalizing visual images. PMID- 11263133 TI - Pre-packaged devices save time. PMID- 11263135 TI - Ethical challenges in everyday practice for healthcare lawyers. AB - A lawyer representing healthcare clients confronts numerous ethical issues in day to-day practice. The authors, practicing healthcare attorneys, first give a quick overview of the history of today's rules of legal ethics, and then turn to hypothetical (but realistic) scenarios to examine counsel's duties under various circumstances. The Article concludes with an examination of the overriding duties of confidentiality and privilege, which guide the analysis of ethical concerns in all areas. PMID- 11263136 TI - The reasonable physician standard: the new malpractice standard of care? AB - We are in the midst of a tremendous, but essentially unacknowledged, shift in the standard applicable in medical malpractice cases across the United States. The author provides a preliminary survey of this fluid area of the law, and provides rationales for the changes. At the same time, it is not yet clear whether the net impact of these changes will be for the better or for the worse--particularly in light of the simultaneous increase in societal emphasis of cost-conscious care. PMID- 11263137 TI - Business Associate Agreements Inventory. AB - This Business Associate Agreement Inventory is a checklist for use by covered entities who use or disclose individually identifiable health information with or to their Business Associates. The checklist is designed to assist the covered entities to review their contracts with each respective Business Associate and determine whether the contracts contain all of the provisions required by the Health Insurance Portability and Accountability Act regulations. The regulations addressed in the checklist include the Transactions and Code Sets Standards (effective October 16, 2000), the proposed Security Standards, and the final Privacy Regulations (published December 28, 2000). PMID- 11263138 TI - The emperor has no clothes: examining the employee exception to the anti-referral laws. AB - The Anti-Referral laws are omnipresent in the practice of health law. Perhaps as a result of their wide scope in application, those laws have had a number of unintended consequences. This Article closely examines one aspect of those laws: the exception applicable to otherwise forbidden conduct that takes place between employer and employee. The author concludes that this exception does nothing to advance the policy of the laws, has led to much confusion, and has had the unintended consequence of accelerating the business consolidation of physicians with each other and with clinics and hospitals. Consequently, she asserts that this exception should be repealed. PMID- 11263139 TI - Extending due process protections in managed care credentialling to hospital privileges. AB - The Article analyzes two recent state court decisions granting due process rights to physicians deselected from managed care networks. The author applauds these decisions and argues that managed care organizations wishing to deselect a physician should be required to demonstrate (1) that they have a legitimate reason for doing so relating to quality of care, economic factors, or administrative considerations, and (2) that the deselection will not unduly affect the quality of healthcare available in the network. In addition, the author contends that these same due process requirements may be applied to the closely analogous area of hospital staff privileges in situations in which the privileges of hospital-based practitioners are tied to employment, or the grant or termination of exclusive contracts. PMID- 11263140 TI - Managing medical staff disability issues: liability of private hospitals under ADA Titles I and III. AB - This Article examines the extent to which private hospital are liable for discrimination against medical staff members with disabilities, under the Americans with Disabilities Act ("ADA"). Specifically, the discussion focuses on the ways in which Title I, covering employment relationships, and Title III, covering places of public accommodation, apply to hospitals and their medical staff physicians. With respect to Title I, the author focuses on possible liability with respect to independent contractor physicians who have staff privileges at a hospital. The focus with respect to Title III involves claims filed by physicians against hospitals as places of public accommodation. The author concludes that the courts have applied the ADA in a manner broader than intended by Congress, and that private hospitals should assume that both Title I and Title III are applicable to staff privilege decisions. Therefore, any action that adversely affects a disabled physician should be supported by well documented, objective evidence of a nondiscriminatory reason for that action. PMID- 11263141 TI - [Anatomy and operation of agger nasi ethmoidal cells and frontal recess]. AB - To reduce postoperative recurrency of nasal polyps and sinusitis, the anatomy of lateral wall of nasal cavity was studied in body head. It was found that there was a considerable wide area between the anterior attachment of the middle turbinate and the roof of ethmoidal sinus, just where the agger nasi ethmoidal cells and frontal recess lying. And also the medial wall of agger nasi ethmoidal cell is just above the anterior attachment of middle turbinate. In our operations, the bony structure above the anterior attachment of middle turbinate was resected to open the medial wall of agger nasi ethmoidal cells and the frontal recess, the lesions in them was removed carefully to reobtain a well drainage. 15 cases were followed up for 0.5 to 1.5 years and the result was satisfying. PMID- 11263142 TI - [Sudden sensorineural hearing loss and jugular bulb diverticulum]. AB - The relationship between high placed jugular bulb (diverticulum) and inner ear disorder is not well known. Three of 19 patients with sudden sensorineural hearing loss (SSHL) treated in 1995 had right side jugular bulb diverticulum revealed by CT scan and MRA. One of the 3 SSHL cases complicated with delayed endolymphatic hydrops. The exact mechanism of causation of inner ear symptom is not clear, but may partly be due to pressure effects with the jugular fossa encroaching on inner ear structure such as the cochlear aqueduct and vestibular aqueduct, and due to turbular flow in the diverticulum striking the inner ear. It is needed to further study the influence of the diverticulum on the inner ear. PMID- 11263143 TI - [Quantitative and objective evaluation of vocal function after vertical hemilaryngectomy]. AB - The lately invented instrument, phono laryngograph, which can evaluate objectively vocal function, was used to determine separately in 30 normal adults and 30 patients with laryngeal carcinoma treated primarily by vertical hemilaryngectomy from 1984 to 1994. The results showed that mean air flow rate, intensity, MPT, have significant difference between surgical groups and normal group (P < 0.01). There is pitch difference between surgical groups and normal group (P < 0.05). We also found that the determined values have significant difference between postoperative two years and postoperative ten years groups (P < 0.01). PMID- 11263144 TI - [Significance of bcl-2 gene breaking and bcl-2 protein expression in nasopharyngeal carcinoma]. AB - In order to study the relationship between bcl-2 gene and nasopharyngeal carcinoma. The two hot breakpoints of bcl-2 gene and bcl-2 protein expression were examined by in situ hybridization and immunohistochemistry technique in 41 nasopharyngeal carcinoma(NPC) tissues. bcl-2 gene breakpoint were found in 4 cases of NPC(positive rate 9.8%). 34 cases of NPC expressed bcl-2 protein, the positive incidence was 82.9%. No statistically significant differences was found in histological grades. All benign lesions were negative for bcl-2 gene breaking and bcl-2 protein expression. There was no corresponding relation between bcl-2 protein expression and bcl-2 gene breaking. The results implicated that bcl-2 gene breaking may not play an important role in the pathogenesis of NPC. It also suggested that overexpression of bcl-2 protein may involved in carcinogenesis of nasopharyngeal epithelium. PMID- 11263146 TI - [Nasopharyngeal carcinoma and dermatomyositis (analysis of 12 cases)]. AB - The onset relationship between dermatomyositis (DM) and malignant tumor, especially nasopharyngeal carcinoma (NPC), was approached. Approximately 90 cases of DM were admitted in the hospital. Among them 15 cases were complicated with malignant tumor of which 12 cases were complicated with NPC (account for 80% of DM with complicated carcinoma). In addition to treat these patients with prednisone and antibiotics, radiation therapy has been applied for NPC and metastasis in the neck. During the hospitalizations, one patient died, eleven patients' NPC was controlled, and the DM was completely resolved or improved. There is an obvious relationship between DM and carcinoma. DM may improve or even subside with good short term prognosis, as long as the carcinoma is controlled successfully. PMID- 11263145 TI - [Anatomical consideration for the injury of dacryocyst and nasolacrimal duct during intranasal sinus surgery]. AB - Based on dissection and measurement on 20 adult head cadavers, there is the close relationship between lacrimal duct and lateral wall of the nasal cavity. The cells of anterior ethmoid sinus were classified into three degrees, according to the relationship between the anterior ethmoid sinus and the lacrimal sac fossa. The average distance is 6.74 +/- 1.72 mm between the nasolacrimal duct and the upper part of the uncinate process, 3.44 +/- 0.75 mm and 5.50 +/- 3.73 mm from the nasolacrimal duct to ethmoidal infundibulum and the maxillary ostium. The orifice of the duct most commonly opens under the insertion of the inferior turbinate anteriorly. To perform intranasal sinus surgery, the safest area and key operation approach were discussed. PMID- 11263147 TI - [Analysis for the auditory brainstem responses of human with wavelet transform]. AB - To find out the value of wavelet transform when it is applied in analysis of human's auditory brainstem responses (ABR). The subjects was divided into two groups, one group is normal hearing subjects (14 cases, 28 ears) and another is sensorial hearing loss subjects (17 cases, 27 ears). ABR of the two groups was collected and represented with wavelet transform respectively. The signals of pre and post being transferred was contrasted and dealt with statistics. Similar to the analysis of guinea pig's ABR, the wavelet transform scale 3 is mostly adequate to the analysis of human's ABR. The time's field after transforming is not changed and the emerged rate of wave I is increased, IV-V compound is divided clearly. So the measurement of I-V range is more precise. Analysing human's ABR with wavelet transform is more precise in measurement of I-V range than with former methods. PMID- 11263149 TI - [Study on contractile properties of the posterior cricoarytenoid muscle after delayed reinnervation]. AB - To study date on the contractile properties of posterior cricoarytenoid muscle after delayed reinnervation of different reinnervated methods. Twenty four dogs were reinnervated at 0,4,5,6,10 and 12 month interval following recurrent laryngeal nerve via the phrenic nerve anastomosed to the recurrent laryngeal nerve after cutting the adductor branch and ansa cervicalis-sternothyroid muscle pedicle implanted into the posterior cricoarytenoid muscle. After 6 months, a series of contractions were recorded from each side in twenty living dogs. The results showed that contractile force of reinnervated muscle decreased gradually with the time of denervation, but contractile force of muscle was no significantly difference between reinnervated side of nerve anastomosed group in 4 months after denervated and normal side, and it was significantly difference between nerve anastomosed group and nerve-muscle pedicle implanted group at some time of delayed reinnervation. The contractile time of reinnervated side of two operated groups was similar to that of normal side. The conclusion demonstrated that the contractile properties can indicate exactly reinnervated degree of muscle, and the earlier reinnervation was performed, the better curative effect was. PMID- 11263148 TI - [The experimental research of inner ear metabolism and electrical physiology of autoimmune sensorineural hearing loss]. AB - Inquire into the mechanism of inner ear pathological physiology in autoimmune sensorineural hearing loss (ASHL). With the auditory electric-physiological techniques and enzyme-histochemical method, the change of inner ear hearing function and enzyme activity were observed. These animals, which threshold of auditory nerve compound active potential (CAP) and cochlear microphonic potential(CM) heightening evidently, showed that the amplitude of endolymphatic potential(EP) (include-EP) bring down in various degrees, which was related to the change of the active of Na(+)-K(+)-ATPase and SDH in vascularis stria and endolymphatic sac. The abnormality of enzymes metabolism in inner ear tissues, which following autoimmune inflammation damage, is the pathological foundation of hearing dysfunction. PMID- 11263150 TI - [The study of ultrastructure of atrial natriuretic peptides immunoreactive in cochlear of guinea pigs]. AB - To further explore ultrastructure of atrial natriuretic peptides immunoreactive (ANP-IR) products in cochlear of guinea pig, and secretory type, ultrastructure of ANP-IR products of stria vascularis were studied with immunoelectronic microscopy. The result indicated: ANP-IR granules were seen in the cytoplasm, more obviously at the ends of the nucleus. There were two types of granule: storage and secretory granule. The study suggests that ANPs were secreted by rupture of restriction membrane, not by exocytosis secretion. PMID- 11263151 TI - [Effects of salicylates on the physiological function of cochlea]. PMID- 11263152 TI - [Parapharyngeal space neoplasms and surgical treatment]. AB - This paper reports surgical treated 66 cases of parapharyngeal space neoplasms. They consist of primary 52 and secondary 14 cases, which were 59 benign and 7 malignant tumors. The surgical procedures which have been used are oral approach in 5 cases, the parotid approach in 5 cases, the cervical approach in 47 cases, the cervical approach with mandibular swing in 7 cases and the postaurical C incision approach in 2 cases. The procedures and indications of the cervical approach or (and) its combination with mandibular swing are outlined. It is considered that these two surgical procedures can provide very safe and efficacious approaches to the removal of the primary parapharyngeal space neoplasms encountered. PMID- 11263153 TI - [Comparative studies of ciliary epithelial morphological and radiologic findings in the osteomeatal complex]. AB - In this study, we evaluated quantitatively preoperative coronal CT scan in 30 nasal cavities of 22 patients with chronic sinusitis who underwent functional endoscopic sinus surgery. The ciliated area of the osteomeatal complex (OMC) from these patients was quantitatively observed by scanning electron microscope and image analysis. Our results showed a positive correlation between ciliary epithelial pathological changes in OMC and disease degree of this area in CT scan. PMID- 11263154 TI - [The valuation of paranasal sinus coronal CT scanning in endoscopy sinus surgery]. AB - We applied coronal CT scanning of paranasal sinus to 260 patients with chronic sinusitis before they were performed endoscopy sinus surgery. The correct diagnostic rate was 99.2% in our study. The positive findings of CT were compared with that from normal person. By analyzing, we consider that coronal CT scanning can clearly reveal various anatomic variation for all sinuses, also, it is valuable in evaluating the difficulties of endoscopic sinus surgery and preventing dangerous operative complications. PMID- 11263155 TI - [The study of PCNA expression and AgNORs counting adult-onset laryngeal papilloma]. AB - Avidin-Biotin-peroxidase Complex technique (ABC) and silver staining technique were used to study the relationship between proliferative activity and the biological status of adult-onset laryngeal papilloma. The proliferating index of PCNA positive cells was closely related to AgNORs count. The two indices were significantly different among these groups (P < 0.05). CONCLUSION: PCNA expression and AgNORs count are useful in distinguishing between benign and malignant laryngeal neoplasms, monitoring premalignancy and observing tumor biological activity. PMID- 11263156 TI - [A study of the expression of c-myc onoprotein in laryngeal cancer and different distant adjacent mucosa]. AB - Using citric-LSAB-microwave immunohistochemical technique, the expression of c myc was studied in different regions of larynx including laryngeal squamous cell carcinoma (LSCC), border area, adjacent mucosa which was 0.5, 1.0, 1.5 and 2.0 cm away from cancer 30 cases and four cases of normal laryngeal mucosa. The results showed that over-expression of c-myc in LSCC was found in 29 of 30 (96.7%), which was higher than that in normal laryngeal mucosa (P < 0.01). A significant different expression of c-myc can be seen at the regions of 2.0 and 1.5 cm distant from the tumor (P < 0.05). The expression of c-myc increased as tissue progressed in sequence from normal mucosa, adjacent mucosa to carcinoma. The c myc expression did not correlate with site of tumor, stage and histological grade (P > 0.05). The results indicate that over-expression of c-myc is involved in genesis of LSCC, and may be an early event in the development of human squamous cell carcinoma of the larynx. The tissue adjacent to the tumor may have a potential malignancy. PMID- 11263157 TI - [The clinical feature of laryngeal cancer in Yanbian area of China]. AB - The pathogenic factors of laryngeal cancer in Yanbian area of China were investigated by analyzing the clinical characteristic of laryngeal cancer patients treated in Yanbian hospital from February 1981 to February 1992. The results showed that the peak period for the occurrence of this disease was at the ages between 50 and 69. The ratio of female patients to males was much lower than that found in Northeast Area of China. The onset of laryngeal cancer had been increasing since 1982 and remained high even in 1990s. The penetrating site was most frequently found in supraglottic portion, and the squamous cell carcinoma was the most common pathologic type. Laryngeal cancer was related to cigarette smoking to which the female patients were more sensitive than males. However, no definite reference was found between drinking and laryngeal cancer. PMID- 11263159 TI - [Clinical analysis of hoarseness after thyroidectomy]. AB - The present paper reports 86 cases of hoarseness after thyroidectomy. In 37 cases, glottic paralysis was confirmed. Among them the injury of recurrent laryngeal nerve were 89.91% (33/37). In 33(36 side) cases of recurrent laryngeal nerve paralysis, left injury was 20 and right was 16. Referring to the literature author consider that: 1. the recurrent laryngeal nerve was injured easy by thyroidectomy because that thyroid gland was located closely with recurrent laryngeal nerve in neck; 2. recurrent laryngeal nerve injury after thyroidectomy was related to the character of thyroid gland tumor and times of operations; 3. incidence of superior laryngeal nerve injure in thyroidectomy was rare; 4. following up 16 cases of glottic paralysis, most of all (13/16) hoarseness was improved with the health side vocal cords overcompensation. PMID- 11263160 TI - [Changes in distortion product otoacoustic emissions and hair cells after noise exposure in guinea pigs]. AB - In order to observe the changes in distortion product otoacoustic emissions (DPOAE) and hair cells after noise exposure, sixteen healthy guinea pigs were used. The animals were divided into 3 groups. Group 1 was normal animals. Group 2 and 3 were animal right and 7 days after exposed to 115 dB SPL simulated submarine engine room noise for 4 hours, respectively. The DPOAE audiogram and I/O function were measured before and after exposure. The changes of hair cells were observed by light microscope and scanning electron microscope. The amplitudes of DPOAE disappeared right after noise exposure(P < 0.01) and recovered to normal at 7 days after exposure (P > 0.05). The light microscopy and scanning electron microscopy showed the stereocilia of the outer hair cells were disarrangement and some disappeared at 2,3,4 kHz regions of the cochlea. The results indicated that the normal outer cells can regulate and compensate the function of some damaged outer hair cells which result in a normal DPOAE in mild damaged cochlea. PMID- 11263158 TI - [The influence of vascular endothelial growth factor and its receptor on the growth of laryngeal cancer cell]. AB - The function of VEGF in laryngeal carcinoma was studied by observing either the expressions of VEGF and its receptor flk-1 in laryngeal carcinoma, or the effect of anti-VEGF antibody and anti-VEGF receptor antibody on the growth of human laryngeal cancer cell line HEP-2 in vitro. The expression of VEGF and flt in the specimen of laryngeal carcinoma were determined by standardized immunohistochemistry (Elite ABC method). Anti-VEGF antibody and anti-VEGF receptor antibody were also added at various concentrations to the culture medium, respectively, and the growth of HEP-2 was measured by MTT assay. The immunohistochemical staining showed that the expressions of VEGF and flt were detected both in carcinoma cells and endothelial cells. The growth of HEP-2 cell line in vitro was suppressed when adding various concentration of anti-VEGF antibody and anti-VEGF receptor antibody to the culture medium. The laryngeal cancer cells can secret VEGF and there exists VEGF-binding site on laryngeal cancer cell. Both blockading VEGF with anti-VEGF antibody and blockading VEGF binding site with anti-VEGF receptor antibody could inhibit HEP-2 growth. PMID- 11263161 TI - [The effection of obstructing OCB with strychnine on the guinea pig's DPOAE]. AB - The strychnine was used to obstruct the oliver cochlear bundle (OCB) in order to explore the effect of the efferent system on distortion product otoacoustic emission (DPOAE) of guinea pig. The result showed that the DPOAE were not changed after strychnine were administrated. It is concluded that the efferent pulses of the CNS does not affect on DPOAE in silent circumstance. PMID- 11263162 TI - [Cochlear and retrocochlear deafness induced by immunity with different inner ear tissue antigens]. AB - OBJECTIVES: To inquire into whether the different inner ear tissue antigens could cause different kind of hearing loss, and to find out the disorder positions of auditory system. METHODS: The basilar membrane (BM), spiral ligament (SL), and spiral ganglion (SG) of guinea pigs were removed for making antigens, respectively. Then, we used these antigens to immune guinea pigs. The special humour and cellular immune reaction, hearing function, and inner ear histopathological changes were observed. RESULTS: In BM-antigen and SL-antigen immune group, the various degrees of cochlear microphonic potential disturbance, recruitment, and immune pathological inflammation in cochlear duct and stria vascularis were found. In SG-antigen immune group, auditory nerve compound action potential changes were prominent, and the inner ear pathological damage mainly existed in cochlear axis vessels or surround areas, and SG. CONCLUSIONS: It was confirmed that the inner ear antigens come from different part of auditory system, which could cause cochlear or retrocochlear autoimmune disease. PMID- 11263163 TI - [Chronic sinusitis: definition and pathogenic factors]. PMID- 11263164 TI - [Expression of endothelin-1 and endothelin-1 mRNA in placental villus of pregnancy induced hypertension and the changes of plasma endothelin-1]. AB - OBJECTIVE: To investigate the role of endothelin (ET-1) in the pathogenesis of pregnancy induced hypertension (PIH). METHODS: In 70 cases of PIH the plasma concentration of ET-1 was measured by RIA, and the expressions of ET-1 and its mRNA in placental villus were determined by immunohistochemistry and in situ hybridization respectively. 30 normal pregnant women (NP) served as control group. RESULTS: (1) The Plasma concentration of ET-1 was significantly higher in PIH than that in NP. There was a positive correlation between the plasma ET-1 level and mean arterial pressure in PIH. (2) The expressions of ET-1 and ET-1 mRNA were found in syncytiotrophoblast of placental villus and in the endothelium of fetal vessels. These positive signals were higher in PIH than in NP. CONCLUSION: The injury of endothelium of fetal and placental blood vessels was one of the causes of PIH. The placenta may play an important role in the pathogenesis of PIH. PMID- 11263165 TI - [Clinical study on pathologic changes of umbilical cord vessels in pregnancy induced hypertension]. AB - OBJECTIVE: To evaluate the relationship between the pathologic features of umbilical cord vessels and the severity of disease in pregnancy induced hypertension (PIH). METHODS: The umbilical cord vessels of forty-five women with PIH and thirty normal term-pregnant women (control group) were observed under light microscopy; the three parts of cord i.e. near placenta, near fetus and middle section of twenty women with PIH selected randomly were observed as well. RESULTS: There were no different pathologic changes in the 3 parts of umbilical cord in PIH; Forty-five women with PIH were categorized into three groups according to the degree of lesions: no lesion (n = 6), mild (n = 13) and severe (n = 26). The pathologic changes of umbilical vessels in moderate and severe PIH were more severe than those in mild PIH (P < 0.05, P < 0.01). However, there were no apparent pathologic changes in the control. The pathologic changes of umbilical vessels in PIH had significant relationship with mean arterial pressure (MAP), neonatal outcome and urinary protein. CONCLUSIONS: The pathologic changes of umbilical vessels can reflect the characteristic lesion of PIH and are parallel with the severity of disease; and there is a homogeneity in pathomorphology of umbilical cord. PMID- 11263166 TI - [Study of the plasma adrenomedullin value in pregnancy induced hypertension patients]. AB - OBJECTIVE: To investigate the relationship between adrenomedullin(ADM) and pregnancy induced hypertension(PIH). METHOD: The ADM values in the plasma from 12 normal term-pregnant women and 26 women with PIH were determined by specific radioimmunoassay. RESULT: (1) The ADM value in the PIH (24.67 +/- 1.27) ng/L increased significantly than that of normal pregnancy (19.16 +/- 1.17) ng/L (P < 0.05). (2) There were significant difference between severe(30.00 +/- 1.17) ng/L moderate (24.80 +/- 0.70) ng/L and mild (19.38 +/- 2.65) ng/L PIH groups (P < 0.001). (3) Significant positive correlation was found between plasma ADM value and mean arterial pressure (r = 0.775, P < 0.001). CONCLUSION: The ADM level is closely associated with PIH and may plan an important role in PIH. PMID- 11263167 TI - [The permeability of the cell membrane in the human uterine smooth muscle at term pregnancy]. AB - OBJECTIVE: To study the changes of the permeability of the uterine smooth muscle cell membrane at the various stages of labor, and comparing with the onset of labor. METHODS: 18 women (38 to 41 gestational weeks) were divided into three groups: late pregnancy not in labor, prelabor, active labor, and six cases in each group. The myometrium tissues in the corpus and lower segment of the uterus were studied by the Lanthanum tracing method under transmission electron microscopy (TEM). RESULTS: Lanthanum entered into the smooth muscle cells, deposited in the mitochondria. The positive rates of the smooth muscle cells of lower segment by lanthanum tracing were markedly higher than that of corpus in late pregnancy group (P < 0.05), the difference were not found between lower segment and corpus in prelabor group (P > 0.05). The positive rates in corpus were significant higher than that of lower site in the active labor group (P < 0.05); Comparing corpus to corpus, the positive rates were not significantly different between late pregnancy and prelabor group (P > 0.05), then in active labor group it were markedly higher than those in the other two groups (P < 0.05, respectively); but in lower segment, the positive rates in late pregnancy group were significant higher than that in prelabor and active labor group (P < 0.05 respectively), no difference was found between prelabor and active labor (P > 0.05). The lanthanum deposition in the late pregnancy and prelabor groups was mainly showed in larger mitochondria within the smooth muscle cell while in active labor group, it laid on the outer and inner membrane and cristae of all the mitochondria. CONCLUSION: The permeability of uterine smooth muscle cell membrane at term, pregnancy increases, especiany in uterine corpus, which showed a close correlation with the onset of labor. PMID- 11263168 TI - [Association of glucokinase gene with gestational diabetes mellitus in Chinese]. AB - OBJECTIVE: To evaluate the role of glucokinase(GCK) gene in the pathogenesis of gestational diabetes mellitus (GDM) in Chinese. METHODS: Two microsatellite polymorphisms, GCK1 and GCK2 which located at approximately 10 Kb 3' and 6 Kb 5', respectively, of the human glucokinase gene on chromosome 7p13, were genotyped in 40 unrelated gestational diabetics and 43 controls. RESULTS: Four alleles (A, B, C, D) and seven genotypes were identified at the GCK1 locus. There was no significant difference in allele and genotype frequency between GDM and control groups at the GCK1 locus. For GCK2, four alleles(1,2,3,4) and eight genotypes were detected. When compared with control subjects, the GDM group had a much less frequency of the allele 2 (51.3% vs 69.8%, chi 2 = 5.965, P = 0.015), and a much more frequency of allele 3(31.3% vs 17.4%, chi 2 = 4.321, P = 0.038). Nine haplotypes of GCK1 and GCK2 were observed and haplotype B/2 was much less in GDM group(1.9% vs 19.4%, RR = 0.078 6, P = 0.003). Among the GDM patients, comparing with carriers of other alleles, the carriers of allele 3 of GCK2 locus had significantly elevated fasting and 2 hour's blood glucose levels during OGTT, and their insulin levels at 2nd and 3rd hour during the OGTT test were significantly decreased. Our results suggest that GCK gene was associated with Chinese GDM, and haplotype of GCK1/GCK2 B/2 was a protective factor for GDM. PMID- 11263169 TI - [Detection of fetal DNA in maternal plasma using the nested polymerase chain reaction]. AB - OBJECTIVE: To search for a new method of non-invasive prenatal gene diagnosis. METHODS: A single-copy human DYS14 gene of Y-chromosome of fetal DNA sequence was amplified by nested polymerase chain reaction (PCR) from twelve pregnant maternal plasma (12-40 weeks). A 239 bp and 198 bp specific fragment were obtained. The maternal plasma samples of twelve pregnant women were used directly for nested PCR. RESULTS The fragment was identified in 8 of 10 male-bearing pregnant women plasma. The diagnostic accordance rate was 80% (8/10), 6 to 8 women gave positive signals in two consecutive amplifications, 2 of 8 women gave positive signals in the second amplification. The rate of positive was increased greatly by nested PCR (from 60% to 80%). None of the other 2 female-bearing pregnant women had positive results. The final accuracy of 83.3% (10/12) was attained in all cases. CONCLUSION: The finding of circulating fetal DNA in maternal plasma may have new implications for non-invasive prenatal gene diagnosis, and the nested PCR possesses the advantages of sensitivity and specificity which improves the clinical application. PMID- 11263170 TI - [Recurrent epithelial ovarian cancer]. AB - OBJECTIVE: To evaluate the factors of influence on recurrent epithelial ovarian cancer and to make strategy of treatment. METHODS: Retrospective study of 189 cases of ovarian cancer admitted in our hospital from Jan, 1987 to Dec, 1997. All of these cases had postoperative pathological diagnosis. RESULTS: Of 31 recurrent cases, 19 cases had first cytoreductive surgery in our hospital, while other 12 cases were done in other hospitals. According to FIGO criteria, stage I, II 4 cases (12.9%), III, IV 27 cases (87.1%). The mean recurrent time of no-residual disease was 17.2 months, while that of residual disease < or = 2 cm was 10.1 months (P < 0.01). The mean recurrent time of those having chemotherapy > or = 6 cycles was 13.1 months, while that of those having chemotherapy < 6 cycles was 10.1 months (P > 0.05). The five-year survival rate of 18 cases having secondary cytoreductive surgery was 27.3%, while that of 4 cases having chemotherapy only was 7.1% (P < 0.01). The five-year survival rate of 7 cases abandoning any therapy was 4.8% (P < 0.01). 2 cases had only irradiation therapy. CONCLUSIONS: Histology, stage, degree of tumor differentiation, chemotherapy and the residual disease of cytoreductive surgery all influence recurrence of epithelial ovarian cancer. Secondary cytoreductive surgery along with chemotherapy can raise recurrent epithelial ovarian cancer's five-year survival rate. PMID- 11263171 TI - [Contraceptive efficacy of Sino-female condom: comparison with condom]. AB - OBJECTIVE: To compare the contraceptive efficacy of Sino-female condom with condom. METHODS: 603 volunteer couples were randomly divided into two groups: 304 couples using female condom for contraception, and 299 using condom. Using lifetable method and log rank test, we compared the pregnancy rates and other discontinuation rates after follow-up for 6 months in two groups. RESULTS: No abnormal findings of cervical and vaginal smears were detected before and after this clinical trial in all 603 women. The follow-up rates at 6 months were 99.01% and 99.67% in the female condom group and condom group, respectively. The 6-month gross cumulative pregnancy rates were 1.06 and 1.69 per 100 women and the discontinuation rates due to allergy were 1.39 and 0.34, respectively. No difference was statistically significant (P > 0.05). However, the discontinuation rate for other causes in the female condom group was significantly higher than that in the condom group (P < 0.01). The main cause was that more than half of subjects were used to applying condom before this study. CONCLUSION: The contraceptive efficacy of Sino-female condom is as same as that of condom, and its clinical use is quite safe. PMID- 11263172 TI - [An introductory clinical trial of three-monthly injectable contraceptive-depot medroxyprogesterone acetate]. AB - OBJECTIVE: To evaluate contraceptive efficacy, continuative rate of use, side effects and acceptability of depot medroxyprogesterone acetate (DMPA) injectable contraception preparation in Chinese women. METHOD: This was an open study. One thousand nine hundred and eighty-five women acquiring contraception received DMPA injection after obtaining informed consent and were followed-up every 3-month up to one year. RESULTS: The total experience accumulated was 1,731.7 women-year with DMPA for 1,985 users. The one year continuation rate was 77.4%. Three subjects became pregnant, the one year accumulative life table failure rate was 0.2%. The accumulative discontinuation rates at one year for other reasons were amenorrhoea (4.8%), bleeding related (13.4%), other medical reasons (2.6%), other personal reasons (2.1%) and lost to follow-up (1.6%). The main complaints were irregular bleeding, amenorrhoea and prolonged bleeding. Lactating women had significantly lower rate of complaint. CONCLUSION: DMPA is a highly effective long-acting contraceptive, it could have good acceptability, especially for lactating women, under the well consultation. PMID- 11263173 TI - [The evaluation of 156 cycles of intracytoplasmic sperm injection in the management of male infertility and idiopathic infertility]. AB - OBJECTIVE: The aim of this study was to examine, retrospectively, the efficacy of intracytoplasmic sperm injection (ICSI) in the treatment of male infertility and idiopathic infertility. METHODS: From November 1995 to January 1998, 140 couples (156 cycles) received ICSI because of severe male factor infertility or idiopathic infertility in our assisted reproductive technology center. The treatment cycles were divided into two groups, group I included 74 cycles done before May 1997, group II included 82 cycles done after May 1997. Mature oocytes were selected for microinjection. RESULTS: No differences were observed between the two groups in the number of mature oocyte and the total cleavage rate. Oocyte survival rate, normal fertilization rate, embryos of good morphology rate and clinical pregnancy rate were significantly higher in group II than that in group I. CONCLUSION: ICSI procedure does affect the quality of embryos. With the improvement of ICSI technique, ICSI should be the treatment of choice for male infertility and idiopathic infertility. PMID- 11263174 TI - [Regulation of nitric oxide and vascular endothelial growth factors on placental function and fetal development]. PMID- 11263175 TI - [Telomeres, telomerase and neoplasms]. PMID- 11263176 TI - [Combined intrauterine and extrauterine gestations]. PMID- 11263177 TI - [Tubal catheterization for transvaginal gamete intrafallopian transfer: an alternative technique for non-tubal infertility]. AB - OBJECTIVE: To evaluate the outcome of transvaginal gamete intrafallopian transfer (TV-GIFT) for infertile patients with non-tubal factors. METHODS: Controlled ovarian hyperstimulation was used for all patients. Transvaginal ultrasound guided oocyte retrieval followed by tubal catheterization for transvaginal gamete intrafallopian transfer of 3.3 +/- 2.1 oocytes (range 1-8) and 200,000 sperms in 50 microliters culture medium. RESULTS: The TV-GIFT was used in total of 23 treatment cycles. Of 23 cycles performed for TV-GIFT, the procedure was completed easily in 18 and difficult in 5, resulting in 5 clinical pregnancies(21.7% per cycle), 2 of which resulted in abortion, 2 ongoing pregnancy one was ectopic pregnancy. No pregnancy occurred in four cycles with only one oocyte transferred. CONCLUSIONS: Simple and cost-effective TV-GIFT may achieve a higher pregnancy rate. Compared with laparoscopically directed GIFT, tubal catheterization for TV GIFT is safe and less invasive. It is an alternative for non-tubal infertility before entering the IVF-ET program. PMID- 11263178 TI - [The relationship of serum hepatitis B virus DNA load in HBsAg positive pregnant women to the intrauterine infection of newborns]. AB - OBJECTIVE: To study the relationship of serum hepatitis B virus (HBV) DNA load in HBsAg positive pregnant women to intrauterine infection of their newborns. METHODS: Serum HBV DNA were determined by dot-blot hybridization in 185 HBsAg positive pregnant women. Serum HBsAg were tested by ELISA in their newborns within 24 hours after birth. RESULTS: The prevalence of intrauterine HBV infection of the newborns was associated with the HBV DNA level of the mothers. With the increase of serum HBV DNA load, the risk of fetal intrauterine infection was increasing. CONCLUSION: Fetal exposure to high level of maternal HBV DNA is one of the important determinant of intrauterine infection. PMID- 11263179 TI - [Study on the relationship between epidermal growth factor and fetal growth retardation]. AB - OBJECTIVE: To evaluate the significance of epidermal growth factor (EGF) in the development of fetal growth retardation (IUGR). METHODS: EGF concentrations in amniotic fluid, maternal and fetal blood from 86 pregnant women in the third trimester were determined by radioimmunoassay (RIA). These subjects were divided into control, large-for gestational age (LGA) group and IUGR group according to neonatal birth weights. EGF concentrations in blood samples taken from umbilical vein and artery of 11 controls were also analyzed. RESULTS: No significant differences of EGF concentrations existed in amniotic fluid, maternal and fetal blood between control and LGA group (P > 0.2 for all). Compared to the control, EGF concentration of IUGR group in amniotic fluid decreased but those in maternal and fetal blood increased significantly (P < 0.05 for all). EGF concentration of cord serum in IUGR group was significantly higher than that in LGA group (P < 0.05), but no differences of EGF levels in amniotic fluid and maternal blood existed between the two groups (P > 0.05, for all). EGF in blood samples taken from umbilical vein and artery were at the same level (P > 0.1). CONCLUSIONS: EGF in amniotic fluid, maternal and fetal blood may originate from different sources, and EGF may play a role in the development of IUGR. PMID- 11263180 TI - [Serum concentration of interleukin-6 in maternal serum and its expression in amniochorionic membranes of preterm premature rupture of membrane]. AB - OBJECTIVE: To investigate the relationship between interleukin-6 (IL-6) and preterm premature rupture of membranes (PPROM). METHODS: Serum IL-6 in 20 cases of normal pregnancy (control group) and 30 cases with PPROM in which 14 had moderate and severe chorioamnionitis (group 1), 16 had no chorioamnionitis (group 2) were measured by enzyme-linked immunosorbent assay (ELISA), and IL-6 protein in fetal membranes were determined by immunohistochemistry technique. RESULTS: The level of serum IL-6 in group 1 (155.17 ng/L) was significantly higher than that in group 2 (79.11 ng/L) and control group (70.15 ng/L) (P < 0.01). Expression of the IL-6 had more positive signs in chorion with chorioamnionitis, especially in infiltrating inflammatory cells. CONCLUSION: IL-6 associated with the onset of preterm labor caused by infection. PMID- 11263181 TI - [The changes of plasma endothelin-1 and calcitonin gene-related peptide levels in patients with pregnancy induced hypertension]. AB - OBJECTIVE: To study the changes and clinical significance of plasma endothelin (ET-1) and calcitonin gene-related peptide (CGRP) levels in patients with pregnancy induced hypertension (PIH). METHODS: Plasma ET-1 and CGRP were studied by immunoradiological method in 60 patients with pregnancy induced hypertension (PIH) and 23 normal pregnant women and 20 normal non-pregnant women. RESULTS: The levels of plasma ET-1 in patients with PIH were significantly higher than those in healthy pregnancies (P < 0.01). There was a positive correlation between the plasma ET-1 level and degree of PIH; the higher the ET-1 level, the severer the PIH. The levels of plasma CGRP in patients with moderate and severe PIH decreased significantly than those in healthy pregnancy. There was a negative correlation between the plasma CGRP level and ET-1 level. CONCLUSION: The results suggest that plasma ET-1 and CGRP concentration could be used as indicators for the severity of PIH. CGRP may be one of the antagonists of ET-1 in the pathogenesis of PIH. PMID- 11263182 TI - [Study of autoimmunity in progeny of pregnant women with systemic lupus erythematosus]. AB - OBJECTIVE: To study the effect of systemic lupus erythematosus (SLE) on physical, mental development and plasma antibody level of SLE in their progenies. METHODS: Routine physical examinations of 49 children from 48 SLE mothers were conducted. Compared immuno-fluorescence anti-nuclear antibody (IFANA), anticardiolipin (ACL), extractable nuclear antigen (ENA) and anti-ds-DNA plasma levels of SLE mothers and their progenies with that levels during pregnancy and in umbilical blood. RESULTS: The physical development (height and weight) in 47 out of 49 children were within normal range while the remaining 2 were in the lower limit. The autoimmune antibodies were all negative in the umbilical blood with autoimmune negative mothers, while the anti-ribonucleoprotein (anti-RNP), anti Smith surface antigen (anti-SSA), anti-specific soluble ribonucleic acid (anti SSB) and ACL could be transferred to fetus through placenta. During follow up study, compared the autoimmune positive rates in progenies with that of mothers, the positive rates of IFANA and anti-ds-DNA decreased significantly (P < 0.01), while no changes in ACL. Compared the autoimmune positive rates in progenies with that of their own umbilical levels, the positive rates of IFANA, anti-RNP, anti SSA decreased significantly (P < 0.01), while no difference existed in ACL. Boys showed faster disappearance of autoimmune positive rates than that of girls. CONCLUSIONS: SLE did not show significant effects on the physical development of their progenies. Most autoimmune antibodies existed in umbilical blood were transferred through placenta during pregnancy and would disappear within 9 years after birth. Autoimmune antibodies decreased quicker in boys, and it indicated that girls should be follow-up more carefully. Autoimmune antibodies in the umbilical blood is an easy method for the screening of SLE progeny. PMID- 11263183 TI - [Effects of castor oil-diet on the synthesis of prostaglandin E2 in pregnant rats]. AB - OBJECTIVE: To determine effects of castor oil-diet on the synthesis of prostaglandin E2(PGE2) and explore the mechanism of labor induced by castor oil diet in pregnant rats. METHODS: Pregnant rats were gavaged castor oil-diet in 18 and 19 days of gestation. At the time of death, blood of portal and peripheral veins, intestinal mucosa, amnion, amniotic cells and placenta were collected, and the tissues were cultured in the presence of ricinoleic acid or indomethacin. The concentrations of PGE2 in the media or blood were measured by RIA methods. RESULTS: The PGE2 levels in the portal vein increased, while the PGE2 levels in peripheral blood had no changes significantly, the PGE2 levels in the tissues of the intestinal mucosa, placenta, amnion and amniotic cells were increased significantly; Ricinoleic acid stimulated the synthesis of PGE2 in the above tissues in vitro, which had the positive correlations with the dose of ricinoleic acid and its lasting time. Indomethacin inhibited the synthesis of PGE2 in-vitro. CONCLUSION: The increased synthesis of PGE2 in the intrauterine tissues is a key of the initiation of labor induced by castor oil-diet, and ricinoleic acid in castor oil-diet might be the active component which induced the initiator of labor. PMID- 11263184 TI - [Expression of endothelial nitric oxide synthase mRNA in human placenta]. AB - OBJECTIVE: To determine the localization and type of nitric oxide synthase in human placenta. METHODS: By polymerase chain reaction and in situ hybridization. The eNOS mRNA expression was observed in 10 cases of human normal term placenta and cord. RESULTS: In human normal term placenta, positive staining of eNOS was evident in the syncytiotrophoblast and the endothelium of umbilical artery and vein, positive staining also presented in the endothelium of stem villous vessels, but it was absent in the endothelium of terminal villous capillary. CONCLUSION: eNOS is present in syncytiotrophoblast and endothelium of stem villi vessels, and it can synthesize nitric oxide which results in the increase of nitric oxide in pregnancy. PMID- 11263185 TI - [Application of fluorescence insitu hybridization technique for prenatal diagnosis of chromosome abnormality in amniotic cells]. AB - OBJECTIVE: To study the technique and diagnostic value of fluorescence in situ hybridization (FISH) in chromosome abnormality for prenatal diagnosis. METHODS: Amniocenteses were performed in 34 pregnant women of 16-23 gestational weeks with indications for prenatal diagnosis. The amniotic fluid samples were cultured in Chang's medium. The metaphase chromosomes were hybridized in situ with the human centromere probes, alpha-satellites DNA probes of X, Y, 13, 21, 18 chromosomes and all primer chromosome probes. These probes were conjugated by Biotin and Dioxin then. The treated slides were examined and taken photos under the fluoromicroscope. RESULTS: By FISH technique, normal karyotypes were shown in 31 cases: 16 cases of 46, XY, 15 cases of 46, XX. Abnormalities were found in 3 patients which were 47, XY + 21; 47, XY + 18 and 45, X/46, X, r (X) respectively. CONCLUSION: Using FISH technique to detect chromosome abnormalities in amniotic fluid for prenatal diagnosis is a reliable method which is rapid and accurate. It has higher sensitivity and specificity in finding chromosome structure abnormalities and marking chromosome DNA sequences. PMID- 11263186 TI - [Clinicopathologic and immunohistochemical features of vulvar Paget diseases]. AB - OBJECTIVE: To improve understanding the factors of development, recurrence and prognosis of vulvar Paget disease (VP). METHODS: Clinicopathologic analyses were performed in 14 patients with VP treated in our hospital from Jan 1959 to Dec 1996. Immunohistochemical stainings for CK7, 34 beta E12, carcinoembryonic antigen (CEA), C-erbB2 and HMB45 were carried out in 6 cases. RESULTS: The mean duration from manifestation of symptoms to diagnosis was 57.1 months. The local recurrent rate after operation was 69%. The mean period from the initial operation to recurrence was 68.8 months. 2 cases out of 4 intraepithelial VP cases recurred with positive surgical margins, but no recurrence in of the other 4 cases with negative surgical margins. Immunohistochemical stainings for CK7 and CEA in 6 cases with VP showed strong positive, whereas for 34 beta E12 and HMB45 in all 6 cases were negative, and for C-erbB2 in 2 cases were positive. CONCLUSIONS: The findings that CK7 and CEA positive in Paget cell of VP, whereas 34 beta E12 and HMB45 negative, suggests that Paget cell of VP possesses characteristics of adenocarcinoma. The recurrence of VP correlates with positive surgical margins. Long term follow-up after operation is essential for VP. The prognosis of intraepithelial and minimally invasive VP was favourable, adjunctive radiotherapy after operation was effective for invasive VP in our studied cases. PMID- 11263187 TI - [Clinical report of six cases of vaginal sarcomas]. AB - OBJECTIVE: To evaluate the clinical characteristics and treatment methods of primary sarcoma of the vagina. METHOD: Retrospectively analysed the clinical data of 6 patients with vaginal sarcoma including 2 leiomyosarcoma, 1 angiosarcoma, 1 lymphoma, 1 alveolar soft tissue sarcoma and 1 rhabdomyosarcoma. RESULTS: The age of the patients was 7-52 year-old. 3 of them were treated by local resection combined with chemotherapy and radiotherapy and alive over 5-year. The rest 3 died of this disease within two years and a half after diagnosis. CONCLUSION: Local resection combined with chemotherapy and radiotherapy is the main therapeutic approach for vaginal sarcoma. PMID- 11263188 TI - [Clinical analysis of 25 cases of primary vaginal malignant melanoma]. AB - OBJECTIVE: To investigate the influence of tumor size and different treatment method on the prognosis of primary vaginal malignant melanoma. METHOD: The clinical data of 25 patients with primary malignant melanoma of the vagina admitted to our hospital from Dec. 1964 to Oct. 1997 were analysed retrospectively. RESULTS: The overall 2-year and 5-year survival rate were 21.4% and 5.4%, respectively. The mean survival time for patients with tumor diameter < or = 2 cm was 27.7 months and for patients with tumor > 2 cm, 9.7 months (P < 0.05). The mean survival time was 19.5 months for the 17 cases treated with surgery and 8.3 months for another 8 cases who did not received surgery (P < 0.05). Multi-variate Cox regression with tumor size, surgical treatment, chemotherapy and immunotherapy also showed that tumor diameter and surgery were significant variables statistically for survival time (both P < 0.05). CONCLUSION: The prognosis of the primary malignant melanoma of vagina is poor and can be improved if the disease could be diagnosed early and combined modality therapy with emphasis on surgery be given. PMID- 11263189 TI - [Effect of the herpes simplex virus I-thymidine kinase gene-ganciclovir system on the transplant of human ovarian cancer on the omentum of nude mice]. AB - OBJECTIVE: To investigate the therapeutic effect of the herpes simplex virus I thymidine kinase gene (HSV1-tk)/ganciclovir (GCV) system on human ovarian cancer. METHODS: The nude mice tumors were formed by injection of human ovarian cancer cell line AO and AO/HSV1-tkc (AO cells carried with HSV1-tk gene which was constructed in China) subcutaneously, and then were transplanted to the omentum of nude mice. The filter-passing culture fluid of VPC/HSV1-tkc was injected daily into the peritoneum of the nude mice in the group of in vivo. Finally, all of the three groups of nude mice (control, in vivo, ex vivo) accepted the treatment of GCV. RESULTS: The inhibitory rate of the RV/HSV1-tkc/GCV to AO tumors were 46.8%, and, only could the residual microscopic tumors be seen in most of the nude mice omentum transplanted with AO/HSV1-tkc tumor after GCV treatment. CONCLUSIONS: GCV could more effectively inhibit the growth of the human ovarian tumors carried with HSV1-tk gene which was transplanted onto the nude mice omentum than which transplanted subcutaneously; The application of the HSV1-tk/GCV system would be very promising in the gene therapy of ovarian cancer. PMID- 11263190 TI - [The growth effect of granulocyte colony-stimulating factors on ovarian carcinoma cell lines]. AB - OBJECTIVE: To evaluate the growth effects of granulocyte colony-stimulating factors (G-CSF) on ovarian carcinoma cell lines. METHODS: We investigated the possible growth effects of G-CSF on three ovarian carcinoma cell lines (SKOV3, 3AO, and CAOV3). Cell proliferation and viability was measured by XTT colorimetric assay. Flow cytometer (FCM) and G-CSF receptor mRNA were measured for response cell lines, the response cell lines were xenotransplanted into athymic mice, to investigated in vivo growth under systemic subcutaneous infusion of G-CSF. RESULTS: G-CSF showed no growth-stimulating effects in 3AO and CAOV3 cell lines. In SKOV3, an increase in growth (< 5%) was seen by XTT assay at 100 ng/ml and 1000 ng/ml after 24 hours treatment, but no significant proliferation was observed by FCM. No G-CSF receptor mRNA was detected by RT-polymerase chain reaction, and no significant alteration of the tumor growth was by G-CSF. CONCLUSION: Although, the above results suggest that G-CSF may act as growth factors in a few ovarian carcinoma cell lines, yet the worry about tumor growth stimulation by G-CSF as potential hazards for their clinical application in ovarian carcinoma patients as an adjuvant with cytotoxic chemotherapy is not necessary. PMID- 11263191 TI - [Cytotoxicities of low dose anticancer agents combining lymphokine activated killer cell against ovarian adenocarcinoma cell line SKOV3]. AB - OBJECTIVE: To investigate whether low-dose anticancer agents could increase the sensitivity of ovarian adenocarcinoma cell line SKOV3 to lymphokine activated killer cell (LAK). METHODS: After SKOV3 cells were pretreated by low dose anticancer agents Taxol, cis-diamminedichloroplatin(CDDP), 5-fluorouracilum(5-FU) for 18 hours, the sensitivity of SKOV3 to LAK was detected by four 51Cr release assay. And the percentage of SKOV3 adhesion to LAK and intercellular adhesion molecule-1 (ICAM-1) expression on SKOV3 were detected by improved Grimm's assay and FACS respectively. RESULTS: After pretreatment of SKOV3 cell with 1.5 micrograms/ml Taxol, 4 micrograms/ml CDDP, 25 micrograms/ml 5-FU or without anticancer agents as control for 18 hours, the cytotoxicities of Interleukin-2 activated LAK against them were 29.7%, 45.9%, 37.2% and 28.5% respectively. The conjugation rates of SKOV3 and LAK were 20.1%, 26.1%, 24.9% and 18.7% respectively. The positive rates of ICAM-1 expression were 52.5%, 65.5%, 68.1% and 49.7% respectively. CDDP and 5-FU increased ICAM-1 expression significantly and the sensitivity of SKOV3 cell to LAK cell lysis was well related to the ICAM 1 expression. CONCLUSION: The results indicate that some low dose anticancer agents can increase the sensitivity of cancer cells to LAK cells and it would be useful in clinical practice. PMID- 11263192 TI - [Osteoarthropathy in pregnancy]. PMID- 11263193 TI - [A study on C-erbB2, nm23 and p53 expressions in epithelial ovarian cancer and their clinical significance]. AB - OBJECTIVE: To investigate the expressions of C-erbB2, nm23 and p53 in epithelial ovarian cancer and their clinical significance. METHODS: Expression of C-erbB2, nm23 and p53 proteins was retrospectively studied by immunohistochemistry, and Kaplan-Meier Method and Cox's proportional hazard regression model were used to analyze the relationship of their expressions with prognosis. RESULTS: (1) The expression rate of C-erbB2 was 31.4%, and was in connection with clinical stage. (2) The expression rate of nm23 was 38.6%, while it maybe correlated with pelvic lymph node metastasis. (3) The expression rate of p53 was 34.3%, and the rate was higher in serous ovarian cancer than in mucinous. (4) There was no correlation among the expression of the three proteins. (5) Survival follow-up data showed that mean survival time was shorter in patients with C-erbB2 expression than those without; and nm23 and p53 were not significantly related to survival time. Multivariate Cox's proportional hazard regression model analysis suggest that C erbB2 expression clinical stage and residual tumor size were three variates significantly contributing to patients prognosis. CONCLUSIONS: C-erbB2 expression has a close relationship with malignancy degree of epithelial ovarian cancer, and significantly contributes to prognosis. p53 expression correlates with its subtype and nm23 with its status of pelvic lymph node metastasis, however, neither contributes to prognosis. PMID- 11263195 TI - [A rational selection of retroperitoneal lymphadenectomy for advanced epithelial ovarian cancer]. AB - OBJECTIVE: To evaluate the rational application of retroperitoneal lymphadenectomy to advanced epithelial ovarian cancer. METHODS: 42 patients of advanced epithelial ovarian cancer were treated by retroperitoneal lymphadenectomy. Two groups were divided according to the residual disease post operation. A: 26 patients with the residual disease < 2 cm; B: 16 patients > or = 2 cm. The regime of combined chemotherapy was the same in the two groups after operation. Clinical stage and pathologic grade showed no difference. RESULTS: The 5-year survival rate was 53.8% (14/26) in A and 12.5% (2/16) in B. There was a significant difference between the two groups (P < 0.001). CONCLUSIONS: The survival rate could be greatly improved for advanced epithelial ovarian cancer through retroperitoneal lymphadenectomy when the residual disease was smaller than 2 cm. This procedure would not be performed if the residual disease was larger than or equal to 2 cm. PMID- 11263194 TI - [Diagnosis and treatment of endodermal sinus tumor of the vagina]. AB - OBJECTIVES: To describe the clinical characteristics and the role of chemotherapy in endodermal sinus tumor of the vagina. METHOD: Two patients with endodermal sinus tumor of the vagina were presented focusing on the clinical manifestations and outcome of the chemotherapy. RESULTS: Patient's age was quite young, 2 and 3 years old respectively. Vaginal bleeding and a polypoid and fragile tumor of the vagina were main clinical features. Elevated serum alpha-FP was found before chemotherapy and dropped dramatically to normal if the tumor was sensitive to chemotherapy. Diagnosis was made by pathology and alpha-FP immunohistochemical staining. Both two patient was well responded to cisplatin vincristine bleomycin (PVB) and cisplatin etoposide bleomycin (PEB) chemotherapy. Clinical and pathological complete remission was obtained after 2-3 courses of chemotherapy without radical surgery and radiotherapy. CONCLUSIONS: Endodermal sinus tumor of vagina in infant was very sensitive to the chemotherapy. Serum alpha-FP was very useful in diagnosis and monitoring of the disease. PMID- 11263196 TI - [Effect of liposome-C-erbB2 antisense oligodeoxynucleotides on the growth and the chemotherapeutic drug sensitivity of human ovarian cancer transplanted in the omentum of nude mice]. AB - OBJECTIVE: To investigate the effects of liposome-C-erbB2 antisense phosphorothioate oligodeoxynucleotides(S-ODNs) on the growth and the chemotherapeutic drug sensitivity of human ovarian cancer transplanted in the omentum of nude mice. METHODS: Human ovarian cancer transplanted in the omentum of nude mice model was established, then divided into four groups: control group, experimental group, chemotherapeutic group, and experimental + chemotherapeutic group. Different treatments were given respectively. The weight of nude mice was observed and the morphology of tumor cells was observed by electromicroscope. RESULTS: The growth inhibitory rate in the experimental group was 37%. There were more heterochromatins in the treated tumor cells. The growth inhibitory rate in the experimental + chemotherapeutic group was increased to 50%. There was no obvious alteration in the weight of experimental group. CONCLUSION: The data in this study suggest that antisense therapy is an useful method of gene therapy in ovarian cancer; Moreover, it could enhance the effectiveness of antitumor drug. PMID- 11263197 TI - [The influences of interleukin-1 beta converting enzyme expression on the biologic characteristics of ovarian cancer cells]. AB - OBJECTIVE: To investigate the effects of interleukin-1 beta converting enzyme (ICE) gene on the biologic characteristics of ovarian cancer cells. METHODS: A 1.3 Kilobase human ICE cDNA with 5' and 3' untranslated sequences was cloned into the EcoR I site of pLXSN retrovirus vector in sense orientation. After that, the recombinant plasmid pLXSN-ICE was transferred to virus-packing cell PA317 by electroporation method. And then the retrovirus containing human ICE cDNA generated by these PA317 cells were used to transfect human ovarian cancer cell lines, AO and 3AO. RESULTS: After being transfected with these retrovirus, the number of the clones obtained after G418 resistance selection is far less than that of controlled groups with plasmid pLXSN. The growth rate of these transfected cells was significantly suppressed in comparison with the parental cell line. Apoptosis was also found in these cells. After the transfection cancer cells were inoculated subcutaneously in nude mice, the tumor growth was significantly inhibited. CONCLUSION: It suggest that human ICE gene can suppress the phenotype and tumorigenicity in nude mice of ovarian cancer cell lines AO and 3AO. PMID- 11263198 TI - [Advances in the study of mechanism of estrogen in the regulation of bone metabolism]. PMID- 11263199 TI - [Issues in prevention and treatment of postmenopausal osteoporosis]. PMID- 11263200 TI - [Measurement of tibia bone content and fracture threshold by quantitative ultrasonography in normal and fractured women in Taiyuan]. AB - OBJECTIVE: To establish the reference range of tibia bone content in normal female of Taiyuan area and to explore the diagnostic criterion for osteoporotic fracture threshold by quantitative ultrasonography (QUS). METHODS: The tibia bone contents (QUS value) of 1,681 normal women (aged 7-92 years) and 63 fractured women (aged 46-91) in Taiyuan were measured by QUS. RESULTS: The QUS value increased with age in normal women before age 30, and peaked at age 30-40, then decreased and reached significance after age 50. There was a negative correlation between QUS value and duration after menopause (P < 0.01). Taken the mean QUS value of normal female aged 20-40 years minus 2.5 s (3,667 m/s) as a cutoff threshold, 93.7% of the 63 fractured women had their QUS value below this threshold. CONCLUSION: It is suggested that the above cutoff QUS value may be considered as the risk threshold of osteoporotic fracture in women. PMID- 11263201 TI - [Bone mineral density investigation of normal women in Changsha by dual energy X ray absorptiometry and its clinical implications]. AB - OBJECTIVE: To establish reference values of bone mineral density (BMD) in normal female. METHODS: 1,257 healthy women aged 15 to 96 years from Changsha were involved. BMD measurements were taken at various skeletal sites (anteroposterior and lateral lumbar spine, hip and forearm) by Hologic QDR4500A dual energy X-ray absorptiometry (DEXA). RESULTS: (1) The BMD value of Ward's triangle and major trochanter peaked early at the age of 20-24, while that of the one-third mid distal forearm peaked late at 40-44 years old. (2) Significant loss of BMD ( 6.4%, -10.8%) was found at lateral lumbar spine and Ward's triangle respectively before age 45. The acceleration of bone loss at various sites appeared both at age 45-64 and 75 with the maximum magnitude at Ward's triangle (64.4%) and minimum at one-third mid-distal forearm (29.0%-34.3%). (3) The rank of osteoporosis detection rate from high to low after age 40 was lateral lumbar spine (27.8%), forearm, Ward's triangle and anteroposterior lumbar spine (10.5%) and femur neck or major trochanter (10.1%). CONCLUSION: BMD measurements are necessary in women after age of 40 years. The most sensitive sites for detecting osteoporosis are lateral lumbar spine, Ward's triangle and forearm. PMID- 11263202 TI - [The profile of low bone mass in amenorrhea with elevated follicle stimulating hormone]. AB - OBJECTIVE: To observe the characteristics of low bone mass in amenorrhea with elevated follicle stimulating hormone(FSH). METHODS: Amenorrhea patients with elevated FSH: primary amenorrhea (PA) 18 cases, secondary amenorrhea (SA), 171 cases and age matched control with normal menstruation (Nor) 180 cases. The descriptive parameters were: estradiol (E2), alkaline phosphatase (ALP), urinary excretion of calcium to creatinine ratio (Ca/Cr), the cortical bone mineral density (CBMD) at right radius measured by single photon absorptimetry (SPA) and the trabecular bone mineral density (TBMD) at lumbar vertebra body measured by quantitative computerized tomography (QCT). RESULTS: The experiment had shown the average E2 level in amenorrhea patients to be < 150 pmol/L. Significantly higher ALP and Ca/Cr values than the Nor group. In the SA group, the CBMD value was (655 +/- 69) mg/cm2, which was significantly lower than the Nor group's value of (677 +/- 56) mg/cm2 (3.2% lower, P < 0.01). The TBMD value is (145 +/- 26) mg/cm3, which is significantly lower than the Nor group's value of (192 +/- 28) mg/cm3 (24.5%, P < 0.001). The disparity with the Nor group was even greater in the PA group (11.1% and 35.7% lower, respectively). The BMD of the amenorrhea patients were negatively linearly correlated with their amenorrhea age. CONCLUSIONS: The serum estradiol level in amenorrhea patients with high FSH was so low that their bone turnover was increased which led to the insufficient bone accumulation and dramatically dropping of TBMD. Its extent was related to the initial age and the duration of ovarian failure. PMID- 11263203 TI - [Study of morphologic effects of 4 Chinese herbs by bone histomorphometry in ovariectomized rats]. AB - OBJECTIVE: To study the therapeutic effects of Chinese herbs Drynaria, Cynomorium, Epimedium and Chain Fern on osteoporotic ovariectomized (OVX) rat models. METHODS: 54 Wistar female rats at the age of 3 months were randomly divided into 2 groups: control groups and experimental group. The control group included: normal groups (N), shamed oprations group(S) and ovariectomized group (OVX) (10 in each group); The experimental group: group A (Drynaria), group B (Cynomorium), group C (Epimedium) and group D (Chain Fern) (6 in each group). Each of the experimental groups was fed separately with the 4 Chinese herbs (0.25 g/100 g B.W/day) respectively by mouth every day for 2 months. Double tetracycline labels were carried out on these animals before the sacrifice. Their left tibia was reserved for the preparation of bone specimen and was studied by bone histomorphometry. RESULTS: The percentage of trabecular volume in total bone volume (TV/TBV) decreased significantly in OVX rats as compared with shame group (P < 0.01). However, after treatment with Drynaria or Epimedium, TV/TBV increased significantly as compared with that before treatment (P < 0.05). The surface area of tetracycline label in Drynaria group lowered and osteoid percentage increased in Epimedium group. CONCLUSIONS: The 4 Chinese herbs have some therapeutic effects on osteoporotic rat models. Drynaria and Epimedium appeared to work better. PMID- 11263204 TI - [Study of nylestriol effect on bone histomorphometric parameters in ovariectomized rats]. AB - OBJECTIVE: To investigate the response of trabecular bone of ovariectomized (OVX) rats to nylestriol. METHODS: Thirty-three female Wistar rats were randomly divided into three groups, 11 rats in each: nylestriol-treated group (E), OVX and the control (C). The OVX rats were used as a model for osteoporosis. After being fed with nylestriol for 3 months, all rats were sacrificed. The effect of nylestriol on bone microarchitecture and dynamics were studied by bone histomorphometry. RESULTS: The data suggest that trabecular bone volume, mean trabecular width, density and cortical thickness were remarkably reduced in OVX group (P < 0.05) while tetracycline labeled surface, osteoid surface and bone turnover rate were increased in comparison with that in C group (P < 0.05). The bone turnover rate and trabecular bone volume were improved with the treatment of nylestriol for 3 months in the OVX rats(P < 0.05). CONCLUSIONS: Trabecular bone volume and bone turnover rate can be improved by 3 month administrations of nylestriol in osteoporotic rat models. PMID- 11263205 TI - [The impacts of sex hormones on histomorphometric and histological appearances of bone in ovariectomized rats]. AB - OBJECTIVES: To demonstrate the histomorphometric and histological changes of bone 3 weeks after bilateral ovariectomy in rats and to investigate the impacts of 4 different hormone replacement therapies on the bone histomorphometric, histological appearances. METHODS: Bilateral ovariectomies were done on 41 female rats and sham operations on other 9 (sham group) respectively. After 3 weeks, 4 different treatments: i.e. Livial, Gevrine, Premarin, Weinian were initiated separately on each 8 ovariectomized rats for another 3 weeks. The remaining 9 were served as controls (OVX group). All rats were sacrificed either 3 weeks after ovariectomy/sham operation or at the end of hormone therapies. Their femoral bone mineral density (BMD) were measured by dual energy X-ray absorptiometry (DEXA). Specimens of proximal femur were embedded undecacifide for histomorphometric analysis and of distal femoral metaphysics were procured for scanning electron microscope (SEM) and pathologic examinations. RESULTS: (1) Three weeks after OVX, the femoral BMD, mean cortical thickness decreased significantly while the number of osteoclast increased significantly as compared with sham group. The trabecular became thinner and irregular which changed the bone microstructure in three dimension. (2) After treatment of 4 different preparations, the above parameters restored to various extents to the sham operation levels. Among them, there was greater increase of femoral BMD on the Livial and Gevrine group as compared with Premarin and Weinian group (P < 0.05). CONCLUSIONS: Bilateral ovariectomy induced increased osteoclast activity and bone turnover, therefore caused accelerated bone loss. Treatment with combined sex hormones preparation could inhibit bone absorption and stimulate bone formation, especially those containing androgenic activity could increase the BMD. PMID- 11263206 TI - [Application of gonadotropin-releasing hormone agonist for triggering ovulation in high risk gonadotropin stimulating cycles of infertile polycystic ovary syndrome patients]. AB - OBJECTIVE: To evaluate the efficacy and safety of gonadotropin-releasing hormone agonist (GnRH-a) instead of hCG for triggering ovulation in high risk gonadotropin stimulating cycles of infertile polycystic ovary syndrome (PCOS) patients. METHODS: GnRH-a was given for triggering follicular final maturation and ovulation in 18 gonadotropin stimulating cycles of 14 PCOS patients with mean serum estradiol (E2) level of 8,379 +/- 2,958 pmol/L. Their outcomes and complications were analysed. RESULTS: Ovulation achieved in 15 (83.3%) treated cycles, 4 (22.2%) became pregnant. Only 1 developed moderate ovarian hyperstimulation syndrome (OHSS) and another 1 had multiple pregnancy. CONCLUSIONS: The use of GnRH-a instead of hCG in high risk gonadotropin stimulating cycles is able to successfully induce ovulation and pregnancy and decrease the incidence of severe OHSS and multiple gestation. PMID- 11263207 TI - [Methotrexate combined chemotherapy for chemorefractory gestational trophoblastic tumour]. AB - OBJECTIVE: To evaluate the response rate and toxicity of etopside, methotrexate, kengshengmycin, cyclophosphamide and vincristine (EMA/CO) regimen for women with chemorefractory gestational trophoblastic tumour. METHODS: Fifty-one patients with chemorefractory gestational trophoblastic tumour were treated by EMA/CO regimen. Operative excision and (or) selective arterial infusion as adjuncts to chemotherapy were performed in a selected subset of patients. The response and toxicity were assessed after treatment. RESULTS: Fifty-one patients received 352 cycles of the study regimen. The median number of courses for each patient was 6.9. Thirty-three cases (64.7%) achieved a complete remission while 14 patients (27.5%) had a partial remission, 4 cases showed nonresponse. Two (6.7%) of 30 complete responders developed recurrences. The main complications for EMA/CO chemotherapy were myelosuppression and gastrointestinal symptoms. CONCLUSION: The EMA/CO regimen is an effective treatment for chemorefractory gestational trophoblastic tumour, and the chemotherapeutic results can be further improved while combined with arterial infusion chemotherapy and surgery in the selected patients. PMID- 11263208 TI - [Neurilemmoma of the pharynx: 4 cases report]. AB - This paper reported 4 cases of neurilemmoma of the pharynx so as to introduce our clinical experience. Tumor body of 3 cases lay parapharyngeal space, and other one did retropharyngeal space. Among 4 cases, tumor body of two cases derived from vagus nerve, one from hypoglossus nerve, and one from pharyngeal plexus likely. The diagnosis of these cases were verified by operation and pathological observation. They didn't recurred after 3 years of following-up. The points for attention in diagnosis and treatment of this disease were discussed. PMID- 11263209 TI - [The use of bi-pedical rotatory door muscle-skin flap reconstruction in extended partial laryngectomy for late (T3 and T4) glottic cancer]. AB - The purpose is to radical treatment for late glottic cancer by surgery, to restore the essential function of the larynx, 62 patients of late (T3 and T4) glottic cancer were treated by extended vertical partial and subtotal laryngectomy. At the same time, an appropriate method of reconstruction of laryngeal function by bi-pedical rotatory door muscle-skin flap was presented. The decannulation rate was 87.1% and 85.0% cases enjoyed satisfactory voice. All cases resumed normal mouth-food-taking. The conclusion is that selective treated with extended partial laryngectomy is effective for T3 and T4 glottic cancer. PMID- 11263211 TI - [Voice rehabilitation by mucosa tube performed after near-total and total laryngectomy for the treatment of advanced laryngeal carcinoma]. AB - This paper introduced a new operative method which would rehabilitated voice for the advanced laryngeal carcinoma patients after near-total and total laryngectomy. All 13 patients had lost chance of partial laryngectomy. Near-total laryngectomy were performed for 10 patients, only the survivor arytenoid of healthy side was preserved, a mucosa tube was sutured by using healthy survivor arytenoid and a mucosa strip which was connected to the trachea. Total larynx of three patients were resected, a mucosa tube was sutured by using postarytenoid and neighbor mucosa. The postoperative follow-up was 2 mouth to 6 years, all patients had near normal voice and normal swallow function. PMID- 11263210 TI - [Rescue and prevention of serious systemic toxicity following local anesthesia to mucous membranes of tetracaine]. AB - Tetracaine remains a popular agent in clinical for the purpose of local anesthesia to mucous membranes. Its potential for systemic absorption and resulting toxicity has been seldom emphasized. One case of fatality from tetracaine pharyngeal local anesthesia for fibro-laryngoscopic procedure is described. The pharmacologic basis, clinical features, rescue and prevention for systemic toxicity is discussed. PMID- 11263212 TI - [Speech reconstruction after laryngectomy by using modified Pearson's operation]. AB - Since 1990, 16 patients received speech reconstruction after laryngectomy by using modified Pearson's operation. Among them, T3, T4 laryngeal cancer 12 cases, T4 hypopharyngeal cancer 4 case, 14 were male and 2 were female, the eldest was 73 years old and the youngest 51 years old, and the average age was 58 years. In the operation, the tracheo-pharyngeal speech tube were sewn up with mucosa remained at laryngopharynx. During 1-5 years follow up, the speech function and swallow function were satisfactory. The 3 and 5-year survival rates were 77.78% and 60.00% respectively. They obtained a good long-term speech function except one. It showed that speech reconstruction with the modified Pearson's operation has important value to advanced pharyngeal and laryngeal carcinoma. PMID- 11263213 TI - [Bacteriological study of chronic maxillary sinusitis in adults and observation of susceptibility to antibiotics]. AB - The unilateral mucosal samples were taken from 42 patients and were cultured for bacteria. Meanwhile, the antimicrobial susceptibility was determined for 11 antibiotics and the beta-lactamase-producing bacteria(beta LPB) were examined. In 85.71% of all samples, bacterial growth was present. Anaerobic bacteria were present in 21.43% of cases. Predominant aerobic bacteria were alpha-hemolytic streptococci and coagulase-negative staphylococci. Predominant anaerobic bacteria were peptostreptococcus species and bacteroides species. The beta LPB were isolated from 40% patients, which were responsible for the resistance to some antibiotics. The antimicrobial susceptibility testing showed that the antibiotics containing beta-lactamase inhibitor, third-generation cephalosporins and third generation quinolones had good efficacy. Our findings indicated the bacterial infection may not play the key role in the development of chronic sinusitis and the bacterial infection may not play the key role in the development of chronic sinusitis and the resistance to some antibiotics due to the high prevalence of beta-lactamase production must be considered when the antibiotics therapy is taken. PMID- 11263215 TI - [ABR feature of medial and old patients with hyperlipidemia]. AB - Sixteen medial and old patients with essential hyperlipidemia who were tested auditory brain-stem response (ABR) with click repetition rate 20/s and 40/s, respectively. In comparison with 12 medial and old people with normallipidemia and 10 health people, the IL and IPL of ABR of the hyperlipidemia group are longer than that of other two groups. There is statistical significant difference among the 3 groups of ABR date. The result showed that the hearing function of the medial and old patients with hyperlipidemia dropped potentially in early phase. Increasing the click repetition rate can improve the detection probability of positive reaction of ABR. PMID- 11263214 TI - [Cholesterol granuloma of the maxillary sinus: 2 cases report]. AB - The paper firstly represents two cases of cholesterol granuloma of the maxillary sinus in China. The common symptoms of the disease include nasal obstruction and bone erosion in severe case. The disease is related to obstruction of the antral ostia, hemorrhage into polypoidal mucosal disease or cholesterol precipitation in the walls of antral cyst. A review of the literature on cholesterol granuloma suggests that poor ventilation in a closed space and long-standing inflammation with effusion or hemorrhage contribute to the formation of cholesterol granuloma. Long-lasting cure is achieved by radical maxillary sinusotomy which is the main therapeutic means. PMID- 11263216 TI - [Chronic suppurative otitis media complicated diffuse serous labyrinthitis: 4 cases report]. AB - We reported 4 cases with rare diffuse serous labyrinthitis who were from 1,112 patients with dangerous otitis media in recent 20 years. The diagnostic key points and operative opportunity were discussed. We think that it is very effective to do radical mastoidectomy as soon as possible, to use vasodilator neurotrophic drugs and glucocorticoid hormone. PMID- 11263217 TI - [Repeat operations for recurrent trigeminal neuralgia after microvascular decompression]. AB - For the study of cause and treatment of recurrent trigeminal neuralgia, repeat operations were performed in 12 patients with recurrent trigeminal neuralgia after microvascular decompression. Among repeat operations, there was vascular loop recompression at trigeminal root in 10 patients, negative exploration in 2 patients. The pain relief was achieved after neurovascular redecompression or neurolysis in all patients. The patients had no recurrent pain during a follow-up from 6 months to 4 years. The causes of recurrent trigeminal neuralgia after microvascular decompression and the questions related to repeat operations were discussed. PMID- 11263218 TI - [A study on DPOAE in patients with diabetes mellitus]. AB - In present experiment we tested DPOAE in 19 patients with diabetes mellitus and compared with control subjects. Results showed that DPOAE (f2 above 3 kHz) disappeared in 2 patients (3 ears) indicating cochlear lesion, both of DPOAE contralateral suppressive effects and selective attention effects significantly reduced in 4 patients (7 ears), and reversed in 3 patients (6 ears) indicating disorders existing in cortex topdown and efferent of MOCS. It was suggested that tests of DPOAE and suppression (induced by CAS and selective attention) may be useful for making early diagnosis of cochlear-pathy and auditory efferent disorders (cortex and MOCS) before retinalpathy happened. This study on DPOAE provided important evidence for early diagnosis and early treatment of diabetic complications. PMID- 11263219 TI - [Clinical study of uvulopalatopharyngoplasty treatment for obstructive sleep apnea syndrome]. AB - In order to investigate the treatment of obstructive sleep apnea syndrome (OSAS), we have performed uvulopalatopharyngoplasty (UPPP) operations on 60 cases. The clinical results showed that 50.0 percent of the operated patients somewhat improved and 16.7 percent unimproved. The total success rate was 83.3%. The key ways; 1. Proper indication; 2. Using local anaesthesia rather than general anaesthesia; 3. Intraoperative monitor ECG, blood pressure and oxygen saturation; 4. Preoperative physician consultation if patients complicated with cardiopulmonary problems. UPPP is one of the effective methods to treat OSAS, especially for OSAS with stricture in pharyngeal cavity. It has less damage, less operative complications and so on. PMID- 11263220 TI - [Diagnosis and treatment of Ramsay Hunt syndrome (a report of 39 cases)]. AB - Thirty-nine cases with Ramsay Hunt syndrome were presented, in which 23 cases were firmly diagnosed early, and others were misdiagnosed to be Bell's palsy in 9 cases, sudden sensorineural hearing loss in 5, herpes zoster pharyngitis in 1 and acute suppurative otitis media in 1, respectively. All patients were treated with prednisone or dexamethasone for 3 weeks. The results of treatment were as follows: complete recovery in 27 cases, residual facial paralysis in 12 patients, in which 11 had sensorineural hearing loss. We concluded that: 1. When patients present idiopathic facial paralysis associated with objective sensorineural hearing loss, Hunt syndrome should be suspected even in the absence of vesicles. 2. Treatment with steroid and antiviral agent is needed. There is no significant difference in the effects between oral and intravenous steroid therapy. 3. The poor prognosis may relate with severe facial paralysis accompanying severe hearing loss. 4. Acoustic stapedius reflex in patients with mild or no hearing loss is useful for defining the involved sites and evaluating the prognosis. PMID- 11263221 TI - [Nasal packing and packing materials chosing after ESS (with clinical analysis of 769 cases)]. AB - Nasal packing after endoscopic sinus surgery is an important factor affecting the outcome of the operation. In order to discuss the clinical effect of no packing method and packing with different materials, we summarized our six years packing experiences of 769 cases (993 nasal cavities) undergoing ESS. The paper disclosed that if operation cavities didn't involve in middle turbinate, inferior turbinate and nasal septum, no packing method could be chosen; Otherwise, packing with different materials could be chosen, both methods could control the postoperative hemorrhage in a safe range. But in order to reducing patients' pain and operation cavities' reaction, it suggests operators had better choose no packing method or packing with absorbable hemostate gauze. PMID- 11263222 TI - [Histopathological study of surgical resection margins of laryngeal carcinoma]. AB - 64 specimens of laryngeal carcinoma were studied by using the step-serial whole organ section and histopathologic examination of the region around tumor. We discussed the situation of lesions spreading from tumor to resection margins, the survival rate of different types of resection margins, the common position of positive resection margins and the correlative factors of positive resection margins combining with histopathology, clinical expression and follow-up results. We concluded that resection margins more than 5 mm far from visible tumor is suitable. PMID- 11263223 TI - [Specific IgG and specific IgG subclass in allergic rhinitis]. AB - Using enzyme-linked immunosorbent assay, we detected the levels of total IgE, caddis fly specific IgE and caddis fly specific IgG subclass in 60 patients with caddis fly allergic rhinitis. The results showed that the levels of SIgE (30.60 +/- 0.78) of patients were significantly higher than that of the controls (P < 0.001). Same results were received in the levels of specific IgG2 and IgG4. The differences of specific IgE and specific IgG4 between the groups before treatment and after treatment were also significant (Specific IgE before treatment: 30.60 +/- 0.78, after treatment: 1.58 +/- 0.44, P < 0.001; specific IgG4 before treatment: 383.31 +/- 96.48, after treatment: 233.68 +/- 80.94, P < 0.001). We conclude that specific IgG subclass has obviously etiologic specificity and sensitivity. PMID- 11263224 TI - [The surgical management of the carotid artery involved with the head and neck cancer]. AB - To discuss the possibility of resection of the carotid artery involved with head and neck cancer without reconstruction. 5 cases of tumor fixed to segment of the carotid artery block were resected, among them 4 without reconstruction and only 1 artery anastomosed. None of them experienced cerebrovascular complications. 3 cases survived two years long, and 2 is now under follow-up. The selective resection of the carotid artery without reconstruction was practical if well prepared. PMID- 11263225 TI - [Examination of eustachian tube function in chronic suppurative otitis media]. AB - Discussing something about the methods of examining the eustachian tube function of chronic suppurative otitis media. The examination of the eustachian tube function were given on 96 cases of chronic suppurative otitis media with Valsalva method, tympanium drip and acoustic impedance method. Though the former two methods were imperfect, the combination of them was similar to the acoustic impedance method in outcome, which is easy and convenient and doesn't need expensive instrument. The combination of the former two methods in result could get rid of the imperfection of each of them and could evaluate the eustachian tube function in accuracy, which is fit for basic-level hospital. PMID- 11263226 TI - [The exchange and significance of the glucocorticoid receptor of circum circulative blood lymphocyte of cases with allergic rhinitis]. AB - It is found that the exceptional unity of blood Gr (Glucocorticoid Receptor) and F (Hydrocortisone) decreased obviously (P < 0.01) after the determination on them of circum-circulative blood lymphocyte of cases with allergic rhinitis. They showed positive interrelation, which means that the decrease of F and GR inhibited the anti-allergic action so that allergic rhinitis may occur. PMID- 11263227 TI - [Effect of furosemide on the carbonic anhydrase activity in vestibule]. AB - In order to explore the mechanism that the furosemide dehydrates and improves the vestibular function in the endolymphatic hydrops. The effects of furosemide on the carbonic anhydrase activity in vestibule were studied by using histocytochemistry and image analysis. The results demonstrated that the furosemide can obviously inhibit the carbonic anhydrase activity. There was no significant difference between the normal ear and the ear with endolymphatic hydrops in the location of the carbonic anhydrase in the vestibule. It suggested that one of the dehydrant mechanisms is inhibition of carbonic anhydrase activity. PMID- 11263228 TI - [Determination of IL-6 in serum and perilymph during inner ear immune response]. AB - During secondary inner ear immune responses against keyhole limpet hemocyanin (KLH), to understand the change of interleukin-6 (IL-6) levels in perilymph and serum, IL-6 levels in perilymph and serum were investigated using an immunoassay. No IL-6 levels were detectable at Day 0 in perilymph. The earliest perilymph IL-6 levels were observed after 6 h, peaking at 24 h, and then decreasing gradually. IL-6 level remnants were detected at 7 days in perilymph. In contrast, during this observation period, low IL-6 levels were only detectable in perilymph from control ears, disappearing after 72 h. IL-6 levels don't change in serum. Previous study has identified the endolymphatic sac (ES) lags behind the earlier appearance of intercellular adhesion molecule-1 (ICAM-1) in the spiral modiolar vein and spiral ligament during immune response. The present study not only provides further support for the existence of an inner ear immune response which is regulated by cytokines, but also supports that cytokines controlling expression of adhesion molecules are released probably by cells which located out of ES. PMID- 11263229 TI - [A research for basic properties of distortion product otoacoustic emissions in normally hearing subjects]. AB - With ILO-92 Otodynamics Analyzer, distortion product otoacoustic emissions (DPOAEs) at the 2 f1-f2 frequency were recorded from 86 normally hearing ears in response to four groups of different primary levels (L1 = L2 = 75 dB SPL; L1 = L2 = 65 dB SPL; L1 = L2 = 60 dB SPL; L1 = 65 dB SPL, L2 = 50 dB SPL). The average DPOAEs-gram demonstrated a bilobed contour with two peaks at approximately 1.5 kHz and 5 kHz and "notch" between 2.5 kHz and 3 kHz. Moreover, the contour of DPOAEs-gram had no obvious changes with sex, ear or primary stimulus levels differences. While the primary levels increased, the amplitude of DPOAEs gradually increased, which was about 55-65 dB SPL below the primary levels. At each frequency from 1-6 kHz, statistical analysis showed that there existed significantly different DPOAEs level under four groups of different primary stimulus levels. PMID- 11263230 TI - [Application of coronal CT scan and three-dimensional reconstruction in rhinology]. AB - To display the structure of the Ostiomeatal Complex (OMC) more clearly, and provide clearer and more living imaging for safe functional endoscopic sinus operation and thoroughly eliminating the focus of disease, coronal CT scanning and 3-dimensional reconstruction were performed in 168 sinusitis patients (290 sides) and 107 patients with sinusitis and nasal polyposis (175 sides). The CT findings were compared with endoscopic examination and normal people were taken as control. The found of coronal CT and endoscopic examination are consistent in recognizing mucosal thickening within nasal cavity and sinuses. Three-dimensional CT is more living than common coronal CT in displaying the uncinate process and turbinates. Our conclusion is that coronal CT scan processes high values for recognizing nasal sinus disease, anatomical variants, disease extent and adjacent structures of OMC. Three-dimensional reconstructed CT make the image of OMC more stereoscopic, audio-visual and living. PMID- 11263231 TI - [Observation on large doses of urokinase in treatment of sudden deafness]. AB - From September 1992 to July 1997 56 cases (59 ears) of sudden deafness were treated with urokinase 20,000 unit a day for ten days. At the same time also using Papaverine hydrochloride, ATP and CoA. The effective rate of treatment was 86.44%. But for the control group of 48 cases treated with only Papavrine hydrochloride, ATP and CoA, the effective rate of treatment was 63.32%. There was significant difference between the two groups (P < 0.01). In addition, urokinase is safe and reliable in treating sudden deafness and has not be found any evident side-effect. PMID- 11263232 TI - [The influence of posture on nasal geometry]. AB - To investigate the effect of posture on nasal cavity geometry, nasal cavity geometry were measured by acoustic rhinometry in 35 healthy adults (17 males and 18 females, 26-50 years old, mean age 32 years) after 5 min in changing postures of head and body. The minimum cross-sectional area and volume between the nostril and 8 cm posteriorly were measured on both sides. When changing from sitting to lateral recumbent with the wider side of the nasal cavity downwards both total volume and total minimum cross-sectional area decreased 18%, volume and minimum cross-sectional area of the wider sides showed that decreased 32% and 26%. When changing from lateral recumbent with the wider side of the nose downwards to lateral recumbent with the narrower side downwards, volume and minimum cross sectional area of the wider sides and total volume showed that increased 41%, 24% and 12%. Our findings indicate that there is a complex response of the nasal cavity mucosa to posture changing. This factor should be taken into account in rhinological studies of environmental, clinical and pharmacological conditions. PMID- 11263233 TI - [Voice measurement with computer and analysis in normal adult]. AB - Acoustic parameters of 90 normal adults (20-50 years old) were measured with computer. The results showed that fundamental frequency (F0), frequency perturbation quotient (FPQ) and amplitude perturbation quotient (APQ) between male and female in same ages were statistically significant difference. F0 and FPQ were bigger but APQ was smaller in female. FPQ and APQ were significantly related to F0. OCT wasn't obviously different among sexes or ages. The article indicated that voice of adults and the vibration of vocal cords had distinctive characteristics in different sexes or mode of phonation, which could provide objective methods and grounds for clinical voice evaluation. PMID- 11263234 TI - [Trans-ethmoidal optic nerve decompression in traumatic optic neuropathy]. AB - To evaluate the outcome of trans-ethmoidal optic nerve decompression (TOND). METHOD: Between November 1984 and May 1996, 35 patients with traumatic optic neuropathy (TON) underwent TOND. While 6 patients were treated with high dose corticosteroids (HDC). RESULTS: 13(37.14%) cases of 35 patients who underwent TOND showed improvement in vision. 1 (16.67%) of 6 patients who were treated with HDC showed improvement in vision. Of the 12 patients with some initial vision, 8 patients underwent TOND and 7(87.5%) patients showed improvement in vision. Another 4 patients treated with HDC, 1(25.0%) patient showed improvement in vision. Of the 29 patients who presented with no light perception, 27 patients underwent TOND and 6(22.22%) patients showed improvement in vision. 2 patients treated with HDC failed to get vision improvement. CONCLUSION: TOND can improve vision in patients with TON. It is rational to perform surgical decompression early. Using of HDC before and after surgery may improve the outcome of TON. No light perception and absence of waveform on visual evoked response is not contradict, some patients may show improvement in vision. PMID- 11263235 TI - [Significance of medical imaging findings in sphenoid sinus disease to pituitarism]. AB - To explore the relationship between medical imaging finding in sphenoid sinus disease and pituitarism, medical images in sphenoid sinus of 7 patients with pituitarism were analysed. Three cases had mass, and the other 4 cases had inflammatory disease in sphenoid sinus. However, their pituitary images were all normal. Sphenoid sinus disease is significant to pituitarism, but may be overlooked. Radiologists must pay more attention to imaging finding in sphenoid sinus. PMID- 11263236 TI - [Subjective and objective of evaluation of UPPP in the treatment of obstructive sleep apnea syndrome in short-term follow-up]. AB - Uvulopalatopharyngoplasty (UPPP) has become a widely accepted method for treating obstructive sleep apnea syndrome (OSAS). The authors studied 52 OSAS patients treated with UPPP and assessed pre- and postoperative parameters including subjective clinical analysis and objective evaluation of computer test. 34(65.38%) OSAS patients felt satisfied of improving snoring sound 3-6 months later following UPPP. 46 patients reported an improvement in their symptoms of daytime somnolence and morning headache. 41 patients underwent follow-up sleep analysis computer (SAC) study; 19 of them (46.34%) had a reduction in AHI more than 50%; meeting our criteria for surgical success. All 41 patients had a significant reduction of total duration of apnea at sleep stage. The study suggests that the effectiveness of UPPP in treating OSAS is limited and subjective clinical symptoms following UPPP do not correlate with the objective finding of SAC; so it is important to stress the significance of follow-up study and necessity to further therapy. PMID- 11263237 TI - [Cervical necrotizing fasciitis]. AB - Necrotizing fasciitis is a severe fatal soft tissue infection characterized by necrosis of fascia. It was caused by polymicrobial infections with aerobe and anaerobe. Although it commonly involved abdomen, extremities and perineum, it might also occur in head and neck. One case of CNF arising from pharyngolaryngitis was reported. A review of the literature with the clinical presentations, bacteriology, diagnosis and treatment was presented. The key to successful management is early recognition, broad-spectrum antibiotics and prompt aggressive surgical intervention with medical support and hyperbaric oxygen therapy. PMID- 11263238 TI - [Study on TGF beta 1, TGF beta 2, TGF beta 3 expression in the chick basilar papilla following gentamicin toxicity]. AB - The beta-type transforming growth factors (TGF beta s) are secreted proteins, which play an important role in regulation of cell proliferation and differentiation in the embryonic inner ear. In order to probe into the effect of TGF beta s on the hair cell regeneration, expression of TGF beta 1, TGF beta 2 and TGF beta 3 proteins were examined by using immunohistochemistry in the chicken basilar papilla during hair cell regeneration following gentamicin ototoxicity. Ten-day-old chickens received daily subcutaneous injection of gentamicin sulfate 50 mg/kg of ten consecutive days. The animals were allowed to survive 1,3,7,14,21 and 28 days before sacrifice and preparation for examination of the expression of TGF beta 1, TGF beta 2 and TGF beta 3 proteins. Immunostaining results demonstrated that TGF beta 2 and TGF beta 3 proteins were observed in the damaged region of basilar papilla. TGF beta 2 and TGF beta 3 proteins positive cells were limited to the lumenal nuclear layer within the damaged region. TGF beta 1 protein positive cell was not found in our study. These results indicated that TGF beta 2 and TGF beta 3 proteins might play a role in regulating proliferation of the supporting cells immigrated into the lumenal nuclear layer during hair cell regeneration. PMID- 11263239 TI - [The effect of endothelium-derived relaxation factor on specific property of vasomotion of cochlea in guinea pig]. AB - By using intravital microscopy measurement with image processing system, continuous diameter changes were measured by self-tracking with computer, the present study is to investigate specific property of vasomotion in cochlear microvessels and its responsibility to the endothelium-derived relaxation factor (EDRF/NO). The results demonstrated that the vasomotion of cochlear arteriole existed in normal condition in a small pattern. It was found that the vasomotion was significantly enhanced after administration of L-arginine(which is donor of EDRF/NO), and L-arginine induces a sequence of changes in cochlear blood flow. There were significant statistic difference before and after the drug administration. The conclusion suggest that EDRF/NO play an important role in the increase of blood flow by improvement of vasomotion. PMID- 11263240 TI - [Effects of testing parameters on the result of tympanometry]. PMID- 11263241 TI - Endothelium-dependent hyperpolarization of vascular smooth muscle cells. AB - In response to various neurohumoral substances endothelial cells release nitric oxide (NO) and prostacyclin, and produce hyperpolarization of the underlying vascular smooth muscle cells, possibly by releasing another factor termed endothelium-derived hyperpolarizing factor (EDHF). NO and prostacyclin stimulate smooth muscle soluble guanylate and adenylate cyclase respectively and can activate, depending on the vascular tissue studied, ATP-sensitive potassium (KATP) and large conductance calcium-activated potassium channels (BKCa). Furthermore, NO directly activates BKCa. In contrast to NO and prostacyclin, EDHF mediated responses are sensitive to the combination of charybdotoxin plus apamin but do not involve KATP or BKCa. As hyperpolarization of the endothelial cells is required to observe endothelium-dependent hyperpolarization, an electric coupling through myoendothelial gap junctions may explain the phenomenon. An alternative explanation is that the hyperpolarization of the endothelial cells causes an efflux of potassium that in turn activates the inwardly rectifying potassium conductance and the Na+/K+ pump of the smooth muscle cells. Therefore, in some vascular tissue K+ could be EDHF. Endothelial cells produce metabolites of the cytochrome P-450-monooxygenase that activate BKCa, and induce hyperpolarization of coronary arterial smooth muscle cells. Whether or not EDHF could be an epoxyeicosatrienoic acid is still a matter of debate. The elucidation of the mechanism underlying endothelium-dependent hyperpolarizations and the discovery of specific inhibitors of the phenomenon are prerequisite for the understanding of the physiologic role of this alternative endothelial pathway involved in the control of vascular tone in health and disease. PMID- 11263242 TI - Basic fibroblast growth factor up-regulates the expression of vascular endothelial growth factor in primary cultured rat astrocytes. AB - AIM: To examine the effect of recombinant human basic fibroblast growth factor (bFGF) on the expression of vascular endothelial growth factor (VEGF) in primary cultured rat astrocytes. METHODS: Semiquantification PCR (SQ-PCR) and immunocytochemistry were used to investigate the effect of bFGF on VEGF mRNA level and protein level, respectively. RESULTS: Treatment with bFGF dose dependently increased the VEGF mRNA level in astrocytes. The up-regulation of VEGF mRNA induced by bFGF (10 micrograms/L) was detected as short as 3-h treatment. The increase of VEGF mRNA level reached the maximum after 24-h treatment with bFGF. The immunocytochemical staining showed that the VEGF protein level in astrocytes also increased after the cells were incubated with bFGF. CONCLUSION: bFGF induced a marked time- and concentration-dependent increase in VEGF expression in primary cultured astrocytes, suggesting that the effect of bFGF on angiogenesis in brain may act partly by up-regulating VEGF expression in astrocytes. PMID- 11263243 TI - Effects of Veratrum nigrum alkaloids on central catecholaminergic neurons of renal hypertensive rats. AB - AIM: To study the central hypotensive mechanism of Veratrum nigrum L var ussurience Nakai alkaloids (VnA) in renal hypertensive rats(RHR). METHODS: The quantitative method of immunocytochemistry (ICC) was used to observe and detect the effect of VnA (30 micrograms.kg-1, i.v.) on activity of central catecholaminergic (CA) neurons of C1, C2, A1, and A5 areas in RHR. RESULTS: VnA increased the immunoreactivity (IR) of tyrosine 3-monooxygenase (TM) immunopositive (IP) neurons of C1, C2, and A5 areas in RHR experimental group compared with RHR control group [positive units: (1.9 +/- 0.4), (1.18 +/- 0.23), (1.2 +/- 0.4) vs (0.15 +/- 0.22), (0.31 +/- 0.16), (0.69 +/- 0.20), respectively]; IR of TM-IP neurons of C1 and C2 areas in RHR control group was decreased compared with sham-operated group [positive units: (0.15 +/- 0.22), (0.31 +/- 0.16) vs (1.45 +/- 0.29), (1.36 +/- 0.25), respectively]. CONCLUSION: VnA increased the activity of central CA neurons in RHR to exert its hypotensive effect. PMID- 11263244 TI - Molecular modeling on human CCR5 receptors and complex with CD4 antigens and HIV 1 envelope glycoprotein gp120. AB - AIM: To investigate the interaction between human CCR5 receptors (CCR5) and HIV-1 envelope glycoprotein gp120 (HIV-1 gp120) and HIV-1 receptor CD4 antigens (CD4). METHODS: The structurally conserved regions (SCR) of human CCR5 was built by the SYBYL/Biopolymer module using the corresponding transmembrane (TM) domain of bacteriorhodopsin (bR) as the template. The coordinates for amino-terminal residue sequence, and carboxyl-terminal residue sequence, extracellular and cytoplasmic loops were generated using LOOP SEARCH algorithm. Subsequently the structural model was merged into the complex with HIV-1 gp120 and CD4. RESULTS: Human CCR5 interacted with both an HIV-1 gp120 and CD4. The N-terminal residues (especially Met1 and Gln4) of human CCR5 contacted with CD4 residues, mainly with one span (56-59) of CD4 in electrostatic interaction and hydrogen-bonds. The binding sites of human CCR5 were buried in a hydrophobic center surrounded by a highly basic periphery. On the other hand, direct interatomic contacts were made between 7 CCR5 residues and 6 gp120 amino-acid residues, which included van der Waals contacts, hydrophobic interaction, and hydrogen bonds. CONCLUSION: The interaction model should be helpful for rational design of novel anti-HIV drugs. PMID- 11263246 TI - High glucose enhances H2O2-induced apoptosis in bovine aortic endothelial cells. AB - AIM: To investigate the effect of high glucose on hydroperoxide (H2O2)-induced apoptosis in cultured bovine aortic endothelial cells (BAEC). METHODS: BAEC were cultured and passaged in normal glucose (5.5 mmol.L-1, NG) and high glucose (25 mmol.L-1, HG). Morphologic changes and quantification of apoptotic cells were determined under fluorescence microscope after H2O2-treated BAEC for 24 h with Hoechst 33258 staining. DNA fragmentation was visualized by agarose gel electrophoresis. The expression of phospho-p38 Ca(2+)-calmodulin dependent protein kinase (CCDPK, formerly called MAPK) was measured by Western blotting. RESULTS: H2O2 elicited typical apoptotic morphologic changes (chromatic condensation, nucleus fragmentation). At 100 -300 mumol.L-1, both NG- and HG-BAEC incubated with H2O2 for 24 h increased cell apoptosis and phospho-p38 CCDPK expression in a concentration-dependent manner. In HG-BAEC, H2O2 induced DNA fragmentation at a lower concentration than that in NG-BAEC, and the apoptotic cell count in HG-BAEC was also higher than that of NG-BAEC (P < 0.05). Similarly, the expression of phospho-p38 CCDPK induced by H2O2 was up-regulated in HG-BAEC (P < 0.05). CONCLUSION: High glucose enhances H2O2-induced apoptosis in BAEC, which is related to high expression of phospho-p38 CCDPK. PMID- 11263245 TI - Antagonistic action of caffeine against LY294002-induced apoptosis in cerebellar granule neurons. AB - AIM: To study the effect of caffeine on apoptosis induced by inhibition of 1 phosphatidylinositol 3-kinase in cerebellar granule neurons. METHODS: Cerebellar granule neurons culture, agar gel electrophoresis, and stress-activated protein kinase (SAPK)/c-Jun N-terminal protein kinase (JNK) assay kit to measure SAPK/JNK activity. RESULTS: LY294002 evoked apoptosis concentration-dependently in cerebellar granule neurons. But death resulting from LY294002 was prevented by caffeine in a concentration-dependent manner. The survival effect of caffeine was not affected by inhibitors of ryanodine-sensitive Ca2+ release, nor was it inhibited by L-type channel blockers and N-methyl-D-aspartate (NMDA) receptor blocker. In addition, RP-cAMP, H89, and KN62 were not able to inhibit the protective effect of caffeine. Phosphorylation of c-Jun was necessary for the induction of apoptosis induced by LY294002 in cerebellar granule neurons. But caffeine directly inhibited the activation of JNK and decreased phospho-c-Jun in granule neurons. CONCLUSION: Caffeine inhibited the activation of JNK and decreased the phosphorylation of c-Jun to protect granule neurons from LY294002 induced apoptosis. PMID- 11263247 TI - Molecular modeling on solvent effect and interaction mechanism of fentanyl analogs to mu-opioid receptor. AB - AIM: To do theoretical study about solvation effect and interaction mechanism of fentanyl analogs (FA) to mu opioid receptor (microOR). METHODS: Flexible docking (FlexiDock) was performed by using the possible active conformations of FA and optimized 3D structure of mu opioid receptor. Binding energies were calculated. Comparative molecular force field analysis (CoMFA) and quantitative structure activity relationship (QSAR) studies were carried out based on results of flexible docking. Solvation effects were considered by studying interaction of FA with water molecules. Partial least square (PLS) analysis was used to calculate regression equation for analgesic activities using binding energies as descriptive factor. RESULTS: 1) Binding conformations of these analogs derived by flexible docking were reasonable. 2) It was most possible for the FA to exist in water solution in the form of binding conformations. 3) Energetic calculation and QSAR analysis showed a good correlation between the calculated binding energies of FA and their analgesic activities. 4) Based on the 3D-model, the possible interaction mechanism of FA with mu opioid receptor can be illustrated reasonably. CONCLUSION: The nature of the correlation between the binding affinities and analgesic activities of FA was explained by our modeling result. PMID- 11263248 TI - Cell proliferation and Ca(2+)-calmodulin dependent protein kinase activation mediated by alpha 1A- and alpha 1B-adrenergic receptor in HEK293 cells. AB - AIM: To examine the ability of alpha 1-AR subtypes on proliferation and Ca(2+) calmodulin dependent protein kinase (CCDPK, formerly called MAPK) activation in transfected human embryo kidney 293 (HEK293) cells. METHODS: pREP8/alpha 1A-AR, pREP4/alpha 1B-AR, and pREP9/alpha 1D-AR were transfected, respectively, into HEK293 cells by calcium phosphate precipitation. The expression of alpha 1-AR was detected by radioligand binding assays. DNA synthesis was measured by [3H]thymidine incorporation. CCDPK activity was determined by immunoprecipitation method and myelin basic protein was used as substrate. RESULTS: Three clonal HEK293 cell lines stably expressing alpha 1A- or alpha 1B- or alpha 1D-AR were chosen and characterized by radioligand binding assay with receptor densities of about 0.6 nmol.g-1. Treatment with norepinephrine (NE) in the presence of propranolol for 24 h increased DNA synthesis in HEK293/alpha 1A- or HEK293/alpha 1B-AR cells concentration-dependently, with EC50 values of 48.8 nmol.L-1 (95% confidence limits 9.7-246 nmol.L-1) and 8.4 nmol.L-1 (95% confidence limits 2.1 32.9 nmol.L-1), respectively. The increase of DNA synthesis induced by NE 10 mumol.L-1 was 201% +/- 28% and 269% +/- 44% of basal, and the activation of CCDPK was 171% +/- 84% and 292% +/- 92% of basal in HEK293/alpha 1A-AR and HEK293/alpha 1B-AR cells, respectively. Preincubation with prazosin completely abolished NE induced CCDPK activation in HEK293/alpha 1A- and alpha 1B-AR cells. Those changes were not found in HEK293/alpha 1D-AR cells. CONCLUSION: The activation of alpha 1A- or alpha 1B-AR but not alpha 1D-AR induces cell proliferation. PMID- 11263249 TI - Inhibitory effects of dauricine on potassium currents in guinea pig ventricular myocytes. AB - AIM: To study the effects of dauricine(Dau) on the rapidly activating component (IKr), the slowly activating component (IKs) of the delayed rectifier potassium current, and the inward rectifier potassium current (IKl) in guinea pig ventricular myocytes. METHODS: Single myocytes were dissociated by enzymatic dissociation method. The currents were recorded with the whole-cell configuration of the patch-clamp technique. RESULTS: (1) Dau 1, 3, 10, 30, and 100 mumol.L-1 blocked IKr and tail current (IKr-tail) in a concentration-dependent manner. The IC50 for block of IKr-tail was 16 (95% confidence limits: 13-22) mumol.L-1. The time constant of IKr-tail deactivation was (140 +/- 38) ms in the control and (130 +/- 26) ms in the presence of Dau 30 mumol.L-1 (n = 6 cells from 3 animals, P > 0.05). (2) Dau 1-100 mumol.L-1 produced concentration-dependent blocks of IKs and tail current (IKs-tail). The IC50 value for block of IKs-tail was 33 (95% confidence limits: 24-46) mumol.L-1. The time constant of IKs-tail deactivation was (92 +/- 18) ms in the control and (84 +/- 16) ms in the presence of Dau 30 mumol.L-1 (n = 8 cells from 4 animals, P > 0.05). (3) Addition of Dau 30 mumol.L 1 induced block of IKs and IKs-tail (n = 7 cells from 3 animals). The degree of block of IKs and IKs-tail depended on test potentials, increasing with more positive depolarizations. (4) Dau 20 mumol.L-1 blocked mainly inward component of IKl and reduced the reversal potential from -72 mV (control) to -78 mV (n = 6 cells from 3 animals). CONCLUSION: (1) Dau inhibited IKs, but not the process of IKs deactivation. (2) Dau blocked IKr, but not the process of deactivation. (3) Dau had a blocking effect on IKl. PMID- 11263250 TI - Huperzine A ameliorates the impaired memory of aged rat in the Morris water maze performance. AB - AIM: To determine the memory-improving properties of huperzine A in aged rats with memory impairments naturally occurring or induced by scopolamine. METHODS: Morris water maze was used to investigate the effects of huperzine A on the acquisition and memory impairments. RESULTS: During 7-day acquisition trials, aged rats took longer latency to find the platform. Huperzine A (0.1-0.2 mg/kg, s.c.) could significantly reduce the latency. In the probe trials on the eighth day, huperzine A (0.1, 0.2 and 0.4 mg/kg, s.c.) significantly increased the time in the quadrant where plateform had disappeared in aged rats. In the acute experiment, scopolamine (0.1 mg/kg, i.p.) significantly impaired spatial memory in the trained aged rats. Huperzine A (0.4 mg/kg, s.c.) significantly reversed the memory deficits induced by scopolamine. CONCLUSION: Huperzine A ameliorates the impaired memory naturally occurring or induced by scopolamine in aged rats. PMID- 11263251 TI - Effect of batroxobin against dog heart ischemia/reperfusion injury. AB - AIM: To study the effect of batroxobin(Bat) on dog heart ischemia/reperfusion (I/R) injury. METHODS: Dog heart I/R injury was induced by occluding the left anterior descending coronary artery for 30 min and restoring blood perfusion for 90 min. Bat was intravenously injected before heart ischemia and 15 min before reperfusion. Plasma creatine kinase (CK), lactate dehydrogenase (LDH), and myocardial malondiaedehyde (MDA) concentrations were measured. The pathologic changes of I/R myocardium were observed. RESULTS: Bat reduced the mortality rate of I/R dog (I/R group 65.0% vs Bat-I group 30.0% and Bat-II group 28.6%, P < 0.05). Myocytes of I/R heart showed intracellular edema, damaged mitochondria, and concentrated nucleus. Bat decreased these changes. In Bat-I and Bat-II group, plasma CK and LDH level were reduced, the +dp/dtmax and -dp/dtmax at 30 min after ischemia and 90 min after reperfusion were elevated, and left ventricular end dilation pressure (LVEDP) was lowered. The myocardial MDA contents were decreased by 42.3% and 38.1% (P < 0.01) in Bat-I and Bat-II group, respectively. CONCLUSION: Bat may exert an apparent role against dog heart ischemia/reperfusion injury and improve myocardial function. PMID- 11263252 TI - Protective effects of bilobalide on amyloid beta-peptide 25-35-induced PC12 cell cytotoxicity. AB - AIM: To study the effect of bilobalide, a terpene extracted from the leaves of Ginkgo biloba, on beta-amyloid peptide fragment 25-35 (A beta 25-35)-induced PC12 cell cytotoxicity. METHODS: 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide and lactate dehydrogenase assay were used to measure the viability of PC12 cells. Thiobarbituric acid-reactive substances were measured to determine lipid peroxidation of cells. Antioxidant enzymes in PC12 cells were detected. RESULTS: Treatment of PC12 cells with A beta 25-35 (100 mumol.L-1) for 24 h caused a great decrease in cell viability (P < 0.01 compared with control). Bilobalide 25-100 mumol.L-1 dose-dependently attenuated the cytotoxic effect of A beta 25-35. Bilobalide also inhibited A beta 25-35 (100 mumol.L-1)-induced elevation of lipid peroxidation and decline of antioxidant enzyme activities. CONCLUSION: Bilobalide protected PC12 cells from A beta 25-35-induced cytotoxicity. PMID- 11263253 TI - Application of secondary structure prediction in antisense drug design targeting protein kinase C-alpha mRNA and QSAR analysis. AB - AIM: To optimize the design of antisense drug targeting protein kinase C-alpha (PKC-alpha) mRNA and obtain better antisense drugs than ISIS3521 that is undergoing clinical trials. METHODS: RNAstructure (version 3.21, 1999) was utilized to predict the optimal and suboptimal secondary structures of human PKC alpha mRNA (GenBank, X52479), and 29 antisense phosphorothioate oligodeoxynucleotides (S-ODN) targeting the secondary structural elements, 3 partly matched S-ODN and 1 scrambled 3521 were designed. ISIS3521 was set as positive control. Mean (n = 3-5) 50% inhibitory effects on proliferation of A549 cells (IC50) of S-ODN were evaluated. Free energies (delta G degree 37) relating to the target secondary structural elements were calculated according to the nearest neighbor model. The quantitative structure-activity relationship (QSAR) analysis through multiple regression was obtained by SPSS. RESULTS: Three S-ODN; (5'-AGCCCA-GCCGCTTGGCTGGG-3', 5'-AGGAGTGCAGCTGC-GTCAAG-3', 5'-TCAGAGGG ACTGATGACTTT-3') had lower IC50[(48 +/- 7), (50 +/- 4), (64 +/- 2.7) nmol.L-1, respectively] than that of ISIS3521 [(81 +/- 25) nmol.L-1]. The number of bases comprising the target secondary structural element bulge loop, internal loop, and knot, the free energy of S-ODN (delta G degree 37S), and reaction (delta G degree 37R) were important parameters in QSAR equation. In the multiple regression, R was 0.68, P = 0.0193. Not tally with the equation, two S-ODN (5'-TCAAATGGAGG CTGCCCGGC-3', 5'-AAAACGTCAGCCATGGTCCC-3') with favorable target structures and delta G degree 37 did not behave good activities. CONCLUSION: Computer aided design was helpful to obtain S-ODN with better in vitro effect than current positive drug. The degree of instability of secondary structural elements and delta G degree 37 were important factors for drug activity. Other important factors needed for further investigation. PMID- 11263254 TI - Effect of metallothionein on tolerance of nitroglycerin in rats. AB - AIM: To assess whether metallothionein (Met) could improve the nitroglycerin tolerance in vivo. METHODS: Nitrate tolerance was induced by 2-d treatment of nitroglycerin (Nit) patch (0.05 mg.h-1). Endogenous Met production was induced by pretreatment of ZnCl2 and coadministration of intravenous Met (15 mg.kg-1.d-1) 2 d with Nit in tolerant rats. The induction of Met production was confirmed by the assay of liver and plasma Met levels. RESULTS: ZnCl2 induced large amount of endogenous Met production in liver and plasma in Nit + ZnCl2 group than that in control group, (89 +/- 4) micrograms/g tissue vs (11.0 +/- 2.4) micrograms/g tissue in liver, P < 0.01, and in plasma (85 +/- 6) micrograms.L-1 vs (71 +/- 6) micrograms.L-1, P < 0.01. There was no significant difference in plasma Met levels in Nit and control groups [(75 +/- 6) micrograms.L-1 vs (71 +/- 6) micrograms.L-1, P > 0.05]. The endogenous Met production enhanced the hypotensive response in Nit + ZnCl2 group [(15.7 +/- 0.8) kPa vs (11.5 +/- 0.6) kPa, n = 6, P < 0.05]. The maximal vessel relaxation induced by sodium nitroprusside (SNP) was the same in 4 different groups but the highest EC50 (concentration which produces 50% of the maximal response to SNP) was found in tolerant group [(42 +/- 9) nmol.L-1, P < 0.01]. CONCLUSION: Exogenous Met or induction of endogenous Met production antagonize the development of Nit tolerance. PMID- 11263255 TI - Antagonistic effects of 3 sesquiterpene lactones from Atractylodes macrocephala Koidz on rat uterine contraction in vitro. AB - AIM: To study the effects of three sesquiterpene lactones: atractylenolide I (8,9 dehydroasterolide, B), 4,15-epoxy-8 beta-hydroxyasterolide (C), and atractylenolide III (8 beta-hydroasterolide, D) from Atractylodes macrocephala Koidz, on rat isolated uterus smooth muscle. METHODS: Rat isolated uteri bathed in De Jalon I solution were used; acetylcholine (ACh), CaCl2, and oxytocin (Oxy) were used to evoke the contraction of uterus. RESULTS: B, C, and D 28 or 56 mumol.L-1 inhibited the spontaneous movement of uterus, reducing their rest force, contractile force, and movement ability. B 28 or 56 mumol.L-1 also slowed down the frequency of uterus spontaneous contraction, but C or D did not. B, C, or D 28 and 56 mumol.L-1 inhibited the uterine spasm induced by Oxy and ACh. Likewise, Ver 0.28 mumol.L-1, B, C, and D 28 or 56 mumol.L-1 relieved the contraction mediated by CaCl2 in high-KCl solution, but B, C, or D had not marked influence on the maximal response of uterus to CaCl2. CONCLUSION: B, C, and D inhibit the movement of uterus smooth muscle, and the mechanism is related to the inhibition of cholinergic system as well as Ca2+ movement. PMID- 11263256 TI - Bradykinin B1 receptor in isolated human umbilical vein: an experimental model of the in vitro up-regulation process. AB - Bradykinin (BK) B1 receptors are not normally expressed in physiological conditions but could be induced in immunopathological states. Molecular approaches have confirmed that BK B1 receptor gene is transcriptionally induced in injured tissues. In these situations, the cytokine network and other proinflammatory mediators are close linked to BK B1 receptor expression. In this article, we describe the functional characterization of the BK B1 receptor up regulation process in the isolated human umbilical vein and the pharmacological tools employed to demonstrate the de novo synthesis of these receptors. BK B1 receptors are up-regulated in a time- and protein synthesis-dependent process. Furthermore, in this tissue we have demonstrated the close link between the BK B1 receptor sensitization and proinflammatory cytokines, such as interleukin-1 beta and tumor necrosis factor-alpha. We also discuss the possible relationship between nuclear factor-kappa B and BK B1 receptor induction in human umbilical vein. PMID- 11263257 TI - Recent advances in GABAB receptors: from pharmacology to molecular biology. AB - Bicuculline-insensitive receptors for the inhibitory neurotransmitter gamma aminobutyric acid (GABA), GABAB receptors, are a distinct subclass of receptors that mediate depression of synaptic transmission and contribute to neuronal inhibition. When activated, these receptors reduce transmission at excitatory and inhibitory synapses, as a result of an increase in K+ conductance, or a decrease in voltage-dependent Ca2+ currents. They are also linked to G-proteins, or intracellular effector systems in a very complex manner. The recent development of highly specific and potent agonists and antagonists for these receptors has led to a much better understanding of their physiology and pharmacology, including their heterogeneity, as well as their molecular biology. Over the past year, expression and cloning studies have contributed to major advances in characterizing GABAB receptor structure, with the discovery of the amino acid sequences of GABABR1a/R1b splice variants and GABABR2 receptors. These isoforms are widely distributed throughout the nervous system, and can be functionally expressed. Importantly, GABABR2 receptors can form a heteromeric assembly with GABABR1 proteins to operate as a heterodimer that displays robust coupling to inward-rectifying K+ channels, as well as inhibition of forskolin-stimulated adenylate cyclase activity. Further insights underlying the mechanisms of GABAB receptor functions can now be gained, leading ultimately to the therapeutic potential of drugs acting at these sites. It is increasingly clear that new information on GABAB receptor molecular structure will provide a plethora of targets for pharmaceutical intervention in areas such as drug addiction, nociception and absence seizures. This review summarizes the renewed efforts, and highlights the recent advances emerging in this field. PMID- 11263258 TI - Learning deficits induced by 4 belladonna alkaloids are preferentially attenuated by tacrine. AB - AIM: To examine the antagonism of tacrine on the amnesic effects of scopolamine (Sco), anisodine (AT3), atropine (Atr), and anisodamine (Ani). METHODS: Cognitive functions and locomotor activities were determined using two sessions of step through and open-field tests, respectively. Mice were injected with one of the belladonna alkaloids (0.05-50 mumol.kg-1, i.p.) and tacrine (50 mumol.kg-1, s.c.) 30 min before the first session. RESULTS: Tacrine completely blocked the avoidance-learning deficit caused by Sco 0.5 mumol.kg-1, AT3 and Atr 5 mumol.kg 1, or Ani 50 mumol.kg-1. But tacrine partly antagonized the learning deficit induced by Sco 5-50 mumol.kg-1 or Atr and AT3 50 mumol.kg-1. The avoidance-memory deficit caused by Sco 0.05-5 mumol.kg-1 or Atr 5 mumol.kg-1 was completely or partly attenuated by tacrine, which did not antagonize the memory deficit elicited by Sco and Atr 50 mumol.kg-1, AT3 5 and 50 mumol.kg-1, and Ani 50 mumol.kg-1. During the acquisition, the locomotor activity of the mice was inhibited by tacrine. This reduction was completely antagonized by Sco 0.5-50 mumol.kg-1, AT3 5-50 mumol.kg-1, Atr 5-50 mumol.kg-1, and only partly antagonized by AT3 and Atr 0.5 mumol.kg-1 or Ani 50 mumol.kg-1. CONCLUSION: Compared with the avoidance-memory deficit, the avoidance-learning deficit caused by belladonna alkaloids is more preferentially attenuated by tacrine. PMID- 11263259 TI - Prostacyclin-induced relaxations of small porcine pulmonary arteries are enhanced by the basal release of endothelium-derived nitric oxide through an effect on cyclic GMP-inhibited-cyclic AMP phosphodiesterase. AB - AIM: To study the interactions between prostacyclin and endothelium-derived nitric oxide in porcine pulmonary arteries. METHODS: Rings of 5th order of porcine pulmonary arteries were studied in vitro for the measurement of tension and the content in cyclic nucleotides. RESULTS: Prostacyclin, given exogenously, caused endothelium-potentiated relaxations (inhibition of phenylephrine contraction) that were inhibited by the inhibitors of the L-arginine nitric oxide pathway, oxyhemoglobin and N omega-nitro-L-arginine. These inhibitors did not affect the tension in rings without endothelium. Cyclic GMP-concentrations were not increased above basal concentrations in the presence of prostacyclin. Increases were seen with acetylcholine and sodium nitroprusside. Prostacyclin stimulated cyclic AMP concentrations did not reach statistical significance compared to controls. The addition of 8-bromo-cyclic GMP to prostacyclin, however, increased the cyclic AMP content. The nitric oxide synthase inhibitor, nitro-L-arginine (NLA), reduced the prostacyclin-stimulated cyclic AMP content to basal level. Inhibition of cyclic GMP-inhibited cyclic AMP phosphodiesterase by 8 bromo-cyclic GMP or amrinone (a specific inhibitor of this enzyme) potentiated the prostacyclin-induced relaxations in rings without endothelium to a magnitude similar to that observed in rings with endothelium. CONCLUSION: These data suggest that the augmentation by the endothelium of the prostacyclin-induced relaxation of porcine pulmonary arteries is secondary to the inhibition of cyclic GMP-inhibited cyclic AMP phosphodiesterase by basally released endothelium derived nitric oxide. PMID- 11263260 TI - Effects of exogenous testosterone on isolated rabbit corpus cavernosum penis. AB - AIM: To study the effects of exogenous excess of testosterone on the constricting effect of phenylephrine and endothelium-dependent and -independent relaxing effects of different agonists in the corpus cavernosum penis (CCP). METHODS: Specimens of the CCP were obtained from rabbits testosterone for 1 and 2 months and untreated for 2 months after testosterone-treatment for 2 months. Preparations were mounted between two parallel platinum electrodes in organ baths. Responses to phenylephrine, carbachol, and sodium nitroprusside were obtained by adding the reagent cumulatively to the bath. RESULTS: The phenylephrine-induced contractions were decreased with no change in agonist potency (pD2 value) after both 1 and 2 month testosterone-treatment and did not return to control values in corpus cavernosum obtained from rabbits untreated for 2 months after testosterone-treatment for 2 months. Testosterone treatment for 1 or 2 months increased the endothelium-dependent relaxations induced by carbachol and decreased the relaxations elicited by electric stimulation but did not affect the relaxations induced by sodium nitroprusside. These relaxant responses to carbachol and electric stimulation did not return to control values in corpus cavernosum obtained from rabbits untreated for 2 months after testosterone treatment for 2 months. There were no significant changes in the pD2 values calculated by agonist-induced relaxation responses in all testosterone-treatment groups compared with control group. CONCLUSION: The exogenous excess of testosterone plays an important role in erectile function by a direct action on the relaxant and contractile responses of CCP. PMID- 11263261 TI - Dopamine-glutamate interaction in rat striatal slices: changes of CCDPK II, PKA, and LDH activity by receptor-mediated mechanisms. AB - AIM: To study the effects of dopamine (DA) and glutamate (Glu) and their receptor agonists/antagonists on Ca2+/calmodulin-dependent protein kinase II (CCDPK II), cyclic AMP-dependent protein kinase A (PKA) activities and the LDH release in rat striatal slices, and to examine the interaction between DA and Glu transmitter systems in striatum. METHODS: The activities of CCDPK II, PKA, and the release of LDH were determined with the 32P-incorporation and colorimetry respectively in rat striatal slices. RESULTS: (1) Exogenous DA, D1 receptor agonist SKF 38393 and D2 receptor agonist LY 171555 reduced CCDPK II activity in striatal slices; Glu also inhibited CCPPK II activity in a concentration-dependent manner. NMDA receptor antagonist MK-801 could antagonize the inhibitory effect of SKF 38393 and LY 171555 on the CCDPK II activity. D1 and D2 receptor antagonists SCH 23390 and spiperone could also antagonize the decrease of CCDPK II activity induced by Glu; (2) DA and SKF 38393 markedly increased PKA activity in striatal slices, which was reduced by MK-801; (3) DA and Glu increased the release of LDH from the striatal neurons in a concentration-dependent manner. MK-801 antagonized the increase of LDH induced by DA. Spiperone, rather than SCH 23390, could reduce the release of LDH from striatal neuron in the presence of Glu. CONCLUSION: The interaction between DA and Glu transmitter systems is found in the regulation of the CCDPK II and PKA activities and cell function in the striatum. PMID- 11263262 TI - Bilobalide promotes expression of glial cell line-derived neurotrophic factor and vascular endothelial growth factor in rat astrocytes. AB - AIM: To study the effects of bilobalide on the expression of glial cell line derived neurotrophic factor (GDNF) and vascular endothelial growth factor (VEGF) in rat astrocytes in vitro. METHODS: Semiquantification polymerase chain reaction (SQ-PCR) was used to investigate GDNF and VEGF mRNA expression in the astrocytes after bilobalide (5, 15, 50, 100 mumol.L-1) treatment. Immunohistochemistry method was used to detect GDNF and VEGF protein expression in cells treated with bilobalide 50 mumol.L-1 for 24 h. RESULTS: GDNF and VEGF mRNA increased markedly after astrocytes were treated with bilobalide 50 mumol.L-1 for 12 h. GDNF and VEGF protein were detected in the cytoplasm of astrocytes after the cells were treated with bilobalide 50 mumol.L-1 for 24 h. CONCLUSION: Bilobalide induced GDNF and VEGF expression in the cultured astrocytes. PMID- 11263263 TI - Corticosterone impairs cultured hippocampal neurons and facilitates Ca2+ influx through voltage-dependent Ca2+ channel. AB - AIM: To investigate the effect of corticosterone (Cor) on the viability of cultured hippocampal neurons as well as voltage-dependent Ca2+ channel (VDCC) on the membrane of the hippocampal neurons. METHODS: The primary cultured hippocampal neurons were prepared and the viability of hippocampal neurons was determined by MTT assays. Inward Ca2+ currents of VDCC on the membrane of the hippocampal neurons were measured with the whole-cell patch-clamp technique. RESULTS: Treatment with Cor concentration-dependently reduced the survival of hippocampal neurons. The IC50 of Cor was 3.2 mumol.L-1. Neurons from cerebral cortex were affected only by high concentrations of Cor (10 mumol.L-1 and 0.1 mmol.L-1) with the IC50, 85 mumol.L-1, 20 times larger than the former. Whole cell patch-clamp experiment showed that Cor (1 mumol.L(-1)-0.1 mmol.L-1) sprayed to the surface of the hippocampal neurons instantly facilitated Ca2+ influx through VDCC with the maximal elevation of 53%, 191%, and 84% above the baseline respectively and this effect was shown to be concentration-independent. In addition, changing the membrane potentials from -40 mV to -10 mV did not affect the facilitating effect of Cor on the Ca2+ influx, indicating that Cor-induced Ca2+ influx was membrane potential-independent. CONCLUSION: Cor markedly facilitated Ca2+ influx into the hippocampal neurons, which may be one of the important mechanisms underlying the neurotoxicity of Cor to hippocampus. PMID- 11263264 TI - Influence of batroxobin on cerebral ischemia-reperfusion injury in gerbils. AB - AIM: To study the effects of batroxobin (Bat) on neurons survival, neurobehavioral test, ATP levels and hydroxyl radical outputs in hippocampus during forebrain ischemia-reperfusion in gerbils. METHODS: The forebrain ischemia was induced by occluding the bilateral common carotid arteries for 10 min in gerbils, and ATP levels and 2, 3-dihydroxybenzoic acid (DHBA) outputs were assayed by HPLC. The neurons survival were assessed by histology, and behavioral tests of gerbils were assessed by open field test. RESULTS: The number of neurons survival in Ir at d 7 postischemic insult were (7 +/- 4)% of sham-operated gerbils, much less than that in Bat (45 +/- 16)%. The levels of explore activities of ischemic gerbils was 175% and 159% of sham-operated gerbils at d 3 and d 6 postischemic insult, much more than that in Bat (120% d 3 and 140% d 6). Hippocampal ATP levels in Ir were 64% of sham-operated gerbils at reperfusion 60 min, much less than that in Bat I and II (82% and 89% respectively). The hippocampal 2,3-DHBA outputs in Ir increased by 4.5 folds of sham-operated gerbils at reperfusion 60 min, but the 2,3-DHBA outputs in Bat I and Bat II were only 2.6 and 2.4 folds respectively. CONCLUSION: Bat possesses the inhibitory effects on DND and OH. production following cerebral ischemia-reperfusion in gerbils. PMID- 11263265 TI - Effects of TMB-8 on constriction of pial arteries in rats. AB - AIM: To study the effects of 8-(N,N-diethylamino)-n-octyl-3,4,5 trimethoxybenzoate (TMB-8) on constriction of pial arteries (PA). METHODS: The change of PA in rats was observed continuously and directly through cranial window using microcircular image-shearing system. RESULTS: Nimodipine (Nim) 1 mumol.L-1 produced dilatation of PA immediately and the maximal response occurred in 2 min. But the diameter was not changed by TMB-8 50 mumol.L-1. PA diameter decreased immediately after the application of CSF containing KCl. TMB-8 25, 50 mumol.L-1 apparently inhibited KCl-induced PA constriction. When persistent constriction was evoked by 5-HT, diameter of PA were increased in a concentration dependent manner after application of TMB-8, which was inhibited by N omega-nitro L-arginine methyl ester (L-NAME). CONCLUSION: TMB-8 inhibited the contraction induced by 5-HT or KCl in cerebral artery, probably via its calcium antagonization and nitric oxide release. PMID- 11263266 TI - High glucose impairs endothelium-dependent relaxation in rabbit aorta. AB - AIM: To study the effects of high glucose on endothelium-dependent relaxation (EDR) and the action of L-arginine, superoxide dismutase (SOD), or glucose re normalization in aorta. METHODS: Measurement of EDR of the isolated rabbit thoracic aortic rings. RESULTS: Elevated glucose (25 mmol.L-1) caused profound impairment of acetylcholine (ACh)-induced relaxation, EC50: 1.6 mumol.L-1 (95% CL: 7.9 nmol.L(-1)-6.3 mumol.L-1) vs normal glucose (5.5 mmol.L-1) EC50: 0.08 mumol.L-1 (95% CL: 0.02 mumol.L(-1)-0.3 mumol.L-1) (P < 0.01), which not reversed followed by a further 24 h incubation in normal glucose M199, EC50: 2.0 mumol.L-1 (95% CL: 0.2 pmol.L(-1)-12.5 mumol.L-1). However, aortic rings incubated with mannitol (19.5 mmol.L-1) relaxed to ACh normally. L-arginine 1 mmol.L-1 or SOD 150 U.L-1 restored ACh relaxation in elevated glucose to normal, EC50: 0.16 mumol.L-1 (95% CL: 0.04 mumol.L(-1)-0.8 mumol.L-1) and 0.16 mumol.L-1 (95% CL: 0.03-0.63 mumol.L-1). The relaxation in response to sodium nitroprusside was not different between rings exposed to normal or elevated glucose. CONCLUSION: Hyperglycemia impaired EDR, which was not reversible by glucose re-normalization, increased free radical production and altered L-arginine metabolism were involved in this endothelium dysfunction. PMID- 11263267 TI - Effect of PKC-zeta mediating Ang II-stimulated activation of CCDPK on rat cardiac fibroblast proliferation. AB - AIM: To investigate whether the effect of angiotensin (Ang) II or epidermal growth factor (EGF) on cardiac fibroblast proliferation involved in activation of extracellular signal-regulated kinase (ERK) 1/2 or Ca(2+)-calmodulin dependent protein kinase(CCDPK) mediated by protein kinase C (PKC)-zeta. METHODS: Relative activity of CCDPK was measured by Western blotting. DNA synthesis was assayed by [3H]thymidine incorporation. RESULTS: PDBU caused no decrease in Ang II- and 10% FCS-stimulated CCDPK activity and DNA synthesis. In contrary, 65% or 75% EGF- or tetradecanoylphorbol acetate (TDPA, formally called PMA)--stimulated CCDPK activity and 38% or 42% [3H]thymidine incorporation treated by PDBU were inhibited, respectively. Meanwhile 70% and 72% CCDPK activities induced by Ang II and EGF were inhibited by PD 98059, respectively. CONCLUSION: PKC-zeta mediated Ang II-induced activation of CCDPK and cardiac fibroblast proliferation. PMID- 11263268 TI - Anti-HIV-1 activity of trichobitacin, a novel ribosome-inactivating protein. AB - AIM: To determine whether trichobitacin, a novel ribosome-inactivating protein purified from the root tubers of Trichosanthes kirilowii, possesses the anti-HIV activity. METHODS: The inhibition of syncytial cell formation induced by human immunodeficiency virus type 1 (HIV-1) was determined under microscope, reduction of HIV-1 p24 antigen expression level was measured by ELISA, and decrease in numbers of HIV-1 antigen positive cells in acutely and chronically infected cultures were detected by indirect immunofluorescence assay. RESULTS: Trichobitacin was found to greatly suppress syncytial cell formation induced by HIV-1 and to markedly reduce both expression of HIV-1 p24 antigen and the number of HIV antigen positive cells in acutely but not chronically HIV-1 infected culture. The median inhibitory concentration (IC50) in inhibition of syncytial cell formation and HIV antigen positive cells were 5 micrograms.L-1 (95% confidence limits: 1.3-20 micrograms.L-1) and 0.09 mg.L-1 (95% confidence limits: 0.011-0.755 mg.L-1), respectively. CONCLUSION: Trichobitacin is a novel ribosome inactivating protein with anti-HIV-1 activity. PMID- 11263269 TI - Analysis of drug interactions in combined drug therapy by reflection method. AB - AIM: To set up a new method to analyze multidrug interaction in combined drug therapy. METHODS: Based on a dose-effect curve of the combined drugs and the equieffective test, a new mathematical model was set as Q = (Eo - Et)/L (-1 < Q < 1 addition, Q < or = 1 antagonism, Q > or = 1 synergism) where Eo = an observed value (or its fitted value) of combined effect, Et = an expected value of combined effect, and L = an equieffective cutoff between Eo and Et, decided by the equieffective criterion of a special field, the number of data points, and the experimental error. The different types of experimental data were analyzed by the new model. RESULTS: This reflection method could deal with data in combined drug therapy, unnecessary to distinguish between independent and similar action, or exclusive and non-exclusive case among drugs. The number of drugs for combination did not need to be limited. But the experimental data should be enough to fit a dose-effect curve of drugs in combination. If every dose-effect curve of drugs for combination was made, a series of Q values was obtained from all levels of dose-effect for a systematic analysis. To large animal or human experiment, the points of dose-effect of each drug used alone could be reduced to even 1 point. The results of analysis took the criterion of a special field and laboratory error into account in this method. CONCLUSION: The reflection method is an effective method for analysis of multidrug interaction in combined drug therapy. PMID- 11263270 TI - Effect of kanglemycin C on lymphocyte proliferation induced by tetradecanoylphorbol acetate and ionomycin. AB - AIM: To compare the suppressions of kanglemycin C (Kan) with that of cyclosporine (Cyc) on lymphocyte proliferations induced by tetradecanoylphorbol acetate (TPA) with ionomycin (IM), and concanavalin A (Con A). METHODS: Cell proliferation was quantified with 3-(4, 5-dimetyl-thiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) or [3H]thymidine ([3H]TdR) incorporating method. Calcineurin (CN) activity was measured with molybdate dye colorimetry. RESULTS: Kan (8, 40, 80, and 400 nmol.L-1) strongly suppressed splenocyte proliferation induced by TPA and IM, and the suppressive effect of Kan gradually decreased along with the increasing concentrations of TPA (1-100 micrograms.L-1) and IM (125-500 micrograms.L-1). But, the suppression of Cyc on the splenocyte proliferation induced by TPA was mild. Cyc suppressed CN activity of mouse splenocytes stimulated by Con A stronger than Kan. Moreover, Kan and Cyc strongly suppressed spleen enriched T cell proliferation induced by Con A and TPA + IM, and the suppression of Kan on proliferation was partly attenuated by exogenous IL-2. CONCLUSION: Kan competitively suppressed the proliferation induced by TPA and IM, and Cyc mainly suppressed IM part of the proliferation induced by TPA and IM. PMID- 11263271 TI - Nicotine and brain disorders. AB - During the last decade, brain nicotinic acetylcholine receptors were extensively characterized from electrophysiological and pharmacological points of view. These receptors play important roles in memory and cognition and participate in the pathogenesis of several brain disorders (Parkinson's and Alzheimer's diseases, Tourette's syndrome, schizophrenia, depression, attention deficit disorder). In the same diseases, clinical studies showed that nicotine had beneficial effects, both as therapeutic and prophylactic agent. This review presents recent data concerning the structure and properties of neuronal nicotinic receptors, their involvement in the pathogenesis of various brain disorders and the beneficial effects of nicotine as therapeutic agent. PMID- 11263272 TI - A clinical investigation on garlicin injectio for treatment of unstable angina pectoris and its actions on plasma endothelin and blood sugar levels. AB - To investigate the therapeutic effects and mechanisms of garlicin for treatment of unstable angina pectoris (UAP), garlicin injectio was intravenously dripped 60 mg/day in 34 cases for 10 days. Nitroglycerine was used in 21 cases of the control group. The results showed that the total effective rates in improving symptoms and electrocardiogram after garlicin treatment were respectively 82% and 62%, and that the plasma endothelin and blood sugar levels were markedly lowered in cases with hyperglycemia. PMID- 11263273 TI - Clinical observation on the long-term therapeutic effects of traditional Chinese medicine for treatment of liver fibrosis. AB - 48 patients with liver fibrosis due to hepatitis B were treated for 2 years with the drugs for tonifying the kidney, supplementing qi, cooling and invigorating the blood and detoxification. The symptoms were markedly improved, and serum ALT and bilirubin were recovered and kept normal in most of the cases. The mean levels of serum hyaluronic acid, procollagen peptide III and circulating immune complex were decreased and returned to normal after the treatment. B ultrasonography showed that the portal vein kept in normal size in 82% of the patients, the enlarged portal vein diminished in diameter, and the enlarged spleen reduced. PMID- 11263274 TI - Effects of xin mai tong capsule on vasoregulatory peptides in the patients of coronary heart disease. AB - In order to inquire into the therapeutic effects of Xin Mai Tong Capsule ([symbol: see text]) on coronary heart disease with myocardial ischemia, 40 patients were randomly divided into two groups (Xin Mai Tong group and the control group). The plasma endothelin (ET) levels in the two groups of patients were markedly higher than that of the healthy people (P < 0.001), and the calcitonin gene related peptide (CGRP) was similar to that of the healthy people (P > 0.05). After treatment, ET and symptomatic scores in the two groups decreased markedly (P < 0.01), and their S-T segments were elevated obviously (P < 0.01). But the decrease of ET and symptomatic scores and elevation of S-T segment in Xin Mai Tong group were superior to those of the control group (P < 0.05-0.01). The CGRP level in the control group did not vary obviously post treatment, but it increased markedly (P < 0.01) with the addition of Xin Mai Tong Capsule in Xin Mai Tong group. PMID- 11263275 TI - Effects of huang qi wu wu decoction on plasma proteins in 70 cases of chronic pulmonary heart disease. AB - Simple immune agar diffusion test was used to assay the contents of 12 plasma proteins in 70 cases of chronic pulmonary heart disease treated by Huang Qi Wu Wu Decoction ([symbol: see text]), with the other 70 cases who were not given Huang Qi Wu Wu Decoction as the control group. The total clinical effective rate in the treatment group was 90.0%, while that in the control group was 75.7%, with a statistically significant difference between the two groups (P < 0.05). In the treatment group, the levels of prealbumin, transferrin and fibronectin elevated obviously after treatment, and the contents of C-reactive protein, ceruloplasmin, haptoglobin, alpha 1-antitrypsin and alpha 1-acid glycoprotein decreased markedly (P < 0.01). In the control group, only the levels of ceruloplasmin and C-reactive protein decreased significantly (P < 0.05). It is shown that Huang Qi Wu Wu Decoction may enhance the therapeutic effects for pulmonary heart disease, regulate the metabolism of plasma proteins, and improve the life quality of the patients. PMID- 11263276 TI - A clinical study on treatment of diabetic peripheral neuropathy with tang zhi min capsules. PMID- 11263277 TI - Treatment of neurofibromatosis with Chinese herbal drugs. PMID- 11263278 TI - Acupuncture treatment of Parkinson's disease--a report of 29 cases. AB - It can be concluded that acupuncture possesses definite therapeutic effectiveness for Parkinson's disease, which is mainly represented by improvement in the clinical symptoms and signs, delaying of the disease's progression, decrease in the dosage of anti-parkinsonian drug, and expectant treatment of the complications and symptoms induced by the drug side-effects. PMID- 11263279 TI - Acupuncture treatment of 42 cases of insufficient blood supply to vertebrobasilar artery. PMID- 11263280 TI - Remission of abdominal colic during enteroscopy by injecting vitamin K3 into Zusanli acupoint. PMID- 11263281 TI - Fifty-six cases of protrusion of lumbar intervertebral disc treated by penetration and oral administration of Chinese decoction plus traction. AB - Fifty-six cases of the protrusion of the lumbar intervertebral disc in the treatment group were treated by drug-penetration and oral administration of traditional Chinese decoction plus traction, and the other 35 cases in the control group by oral administration of Chinese decoction and traction. The results showed that the cure rate in the treatment group was 83.9%, and that in the control group was 57.1%, with a statistically significant difference between the two groups (P < 0.01), indicating that the former is a more effective therapy for protrusion of the lumbar intervertebral disc. PMID- 11263282 TI - Treatment of protrusion of lumbar intervertebral disc by massotherapy under anesthesia--a report of 64 cases. PMID- 11263283 TI - Twenty one cases of vertebral-artery-type cervical spondylosis treated with acupuncture and moxibustion. PMID- 11263285 TI - Twenty-five cases of intractable cutaneous pruritus treated by auricular acupuncture. PMID- 11263284 TI - Evaluation of therapeutic effect of maneuver-dominated method in 30 cases of cervical spondylotic myelopathy. AB - Thirty cases of cervical spondylotic myelopathy (CSM) were treated by a maneuver dominated non-surgical therapy. Eighteen cases were recovered to grade E according to the criteria set by the American Spinal Injury Association. The effect was definite. Indications and contraindications of the maneuver were proposed on the basis of the pathogenesis of CSM and the principles of this manual method. PMID- 11263286 TI - Treatment of decortical state of child encephalitis with scalp acupuncture and the effects on EEG and BEAM. PMID- 11263287 TI - The analgesic action of semen coicis on severe functional dysmenorrhea--a sequential trial observation. AB - The analgesic effect of Semen Coicis was observed with the sequential trial in 26 cases of severe functional dysmenorrhea. The results showed that the markedly effective rate was 90%, which was much better than that of the control group treated by indomethacin plus subcutaneous injection of atropine (P < or = 0.01). PMID- 11263289 TI - The regulatory action of the modified Yu Ping Feng Tang on cellular immunity in mice under amputation-induced stress. AB - To approach the action of modified Yu Ping Feng Tang ([symbol: see text] Jade Screen Decoction) on cellular immunity, an experiment was conducted in mice under amputation-induced stress. On the 3rd day after amputation, acute atrophy was found in the thymus, the reactivities of T- and B-lymphocytes to Con-A and LPS were decreased, the IL-2 content and its activity reduced and the activity of NK cells lowered. The high, moderate and low concentrations of the modified Yu Ping Feng (YPF) Decoction all have antagonistic action on the above manifestations of immune inhibition. PMID- 11263288 TI - Effects of batroxobin on spatial learning and memory disorder of rats with temporal ischemia and the expression of HSP32 and HSP70. AB - The effect of Batroxobin on spatial memory disorder of left temporal ischemic rats and the expression of HSP32 and HSP70 were investigated with Morri's water maze and immunohistochemistry methods. The results showed that the mean reaction time and distance of temporal ischemic rats in searching a goal were significantly longer than those of the sham-operated rats and at the same time HSP32 and HSP70 expression of left temporal ischemic region in rats was significantly increased as compared with the sham-operated rats. However, the mean reaction time and distance of the Batroxobin-treated rats were shorter and they used normal strategies more often and earlier than those of ischemic rats. The number of HSP32 and HSP70 immune reactive cells of Batroxobin-treated rats was also less than that of the ischemic group. In conclusion, Batroxobin can improve spatial memory disorder of temporal ischemic rats; and the down regulation of the expression of HSP32 and HSP70 is probably related to the attenuation of ischemic injury. PMID- 11263290 TI - The therapeutic ways for chronic liver diseases accompanied by diseases of the endocrine and mammary glands. PMID- 11263291 TI - Acupuncture treatment of sciatica. PMID- 11263292 TI - [A study of the correlations between asthma and airway responsiveness]. AB - OBJECTIVE: To evaluate the effect of airway responsiveness on asthma and to study the causes of asthma. METHODS: The 641 nuclear families were confirmed by indicator cases with asthma in three counties. Investigation and methacholine challenge test were conducted for all relatives of the nuclear families. The data were by EPI5.01 and the odds ratio (OR) was employed to evaluate the correlation between asthma and airway responsiveness. RESULTS: 72 percent of persons with asthma history showed airway hyperresponsiveness, but only 70 per cent of persons with airway hyperresponsiveness suffered asthma. The risk for asthma in relatives with positive airway responsiveness was 5.8 times higher than those with negative airway responsiveness (P > 0.01) among the nuclear families. CONCLUSION: Airway hyperresponsiveness is one of the risk factors of asthma. PMID- 11263293 TI - [The distribution of substance P in the lungs of asthmatic guinea-pigs and the influence of dexamethasone on the distribution]. AB - OBJECTIVE: To investigate the distribution of substance P(SP) in the lungs of asthmatic guinea pigs and the effects of dexamethasone on its distribution. METHODS: Guinea pigs were divided into 3 groups: asthma group, dexamethasone treated group and control group. Rabbit serum against SP, immunohistochemical ABC method and glucose-DAB-nickel technique for staining and computer image analysis were used in this study. RESULTS: There was distribution of SP-IR positive fibres in airways in asthma group than the other two groups. The positive fibres, emerged like a string of beads, were present as large fibre bundles in the smooth muscle layer and basement membrane. Additionally, in the asthma group SP-IR positive fibres were detected on the walls of respiratory bronchioles and alveolar ducts in contrast to the other groups. There was no difference in distribution and morphological characteristic between the control group and the dexamethasone-treated group. CONCLUSION: Repeated antigen challenges, which cause the allergic inflammation of airways, may result in the growth of SP-containing nerve fibres in the airway and synthesis of SP in neurons. These may be involved in the persistence and exacerbation of airway inflammation in asthma. Dexamethasone can reverse the distribution of SP-IR positive fibres to the normal status in airway walls of asthma guinea pigs. PMID- 11263294 TI - [Changes of thromboxane A2 receptors and their mRNA expression and effects of their antagonist phenol red in the lung of acute lung injury of rats]. AB - OBJECTIVE: To investigate the changes of thromboxane A2 receptors and the influence of their antagonist phenol red, and to study in molecular level the mechanism of action of thromboxane A2 in acute lung injury. METHODS: Acute lung injury model of rats was produced by oleic acid, and phenol red (5 mg/kg, 50 mg/kg) was injected via vein 5 minutes before oleic acid injection. Changes of thromboxane A2 receptors and their mRNA expression in the lung were determined at 6 hours after oleic acid injection. RESULTS: Down-regulation of thromboxane A2 receptors were observed, but their mRNA transcription increased significantly in the lung after oleic acid injection. The injection of phenol red before oleic acid could induce down-regulation of thromboxane A2 receptors and abolish the increase of their mRNA transcription after oleic acid injection. Phenol red could reduce LBI, RLW and improve PaO2. Histological examination of lung tissues demonstrated that the degree of lung injury had been reduced significantly. CONCLUSION: Thromboxane A2 plays an important role plays in pathological processes of acute lung injury. Phenol red in vivo could effectively reduce severity of acute lung injury by blocking thromboxane A2 receptors and regulating the expression of thromboxane A2 receptors. PMID- 11263295 TI - [Studies on genes associated with biologic behavior in human lung cancer]. AB - OBJECTIVE: To study alteration of several genes in the process of primary lung cancer. METHODS: A series of 59 lung cancer specimens were analyzed for p53, myc oncogene family and mdrl gene by DNA/PCR sequencing, immunohistochemistry and RT PCR methods. RESULTS: p53 mutation or/and protein accumulation were found in 37 of 57(65%) cases. Overexpression of myc family oncogenes and mdr1 gene were 27/46 (59%) and 15/48 (31%), respectively. The results also showed that there was no significant correlation between p53 alteration and tumor size, metastasis, stage and relapse, but there was a significant correlation between overexpression of myc family oncogene and these factors. Overexpression of mdrl gene was detected in NSCLC, especially in adenocarcinoma, and was not associated with metastasis and stage of lung cancer. It was also found that aberration of both p53 and myc family oncogenes occurred in 19/30(63%) cases; the relapse rate was 76%. Both overexpression of mdrl and myc gene was 62%; the relapse rate was 83%. CONCLUSION: p53, myc and mdrl genes in cooperation may be involved in the process of lung cancer, but prognostic determinant is myc gene overexpression. PMID- 11263296 TI - [Clinical usefulness of a new tumor marker CYFRA21-1 in patients with lung cancer]. AB - OBJECTIVE: To set the serum cut-off value of CYFRA21-1 in lung cancer patients in china, and to evaluate the clinical usefulness of CYFRA21-1 as a new tumor marker of lung cancer. METHODS: CYFRA21-1 levels were measured with two monoclonal antibodies (Ks19.1 and BM19.21) in the serum of 72 patients with lung cancer, 50 patients with benign lung diseases and 33 post-therapy lung cancer patients. To set the cut-off value, ROC curve was use. RESULTS: (1) Serum concentrations of CYFRA21-1 in patients with lung cancer were significantly higher than those with benign lung diseases (P < 0.001). CYFRA21-1 levels in patients with squamous cell carcinoma were significantly higher than those in patients with any other subtypes. The more advanced the clinical stages, the higher the levels of CYFRA21 1. (2) If the threshold of CYFRA21-1 was set at 5.5 micrograms/L, its sensitivity and specificity for lung cancer were 63% and 94%, respectively. The sensitivity for squamous cell carcinoma was 78%, which was the highest among all the pathological subtypes. (3) The sensitivity of CYFRA21-1 in non-small cell lung cancer increased with the advancement of clinical stages. (4) CYFRA21-1 could be a good index in monitoring patient's condition and predicting prognosis for lung cancer. (5) When CYFRA21-1 was used as a screening index for lung cancer, the threshold of CYFRA21-1 should be set at the upper left corner of the receiver operating characteristic curve of stage I-II. The value of CYFRA21-1 in early diagnosis for non-small cell lung cancer was found limited. CONCLUSION: CYFRA21-1 is a sensitive and specific tumor marker of non-small cell lung cancer, especially of squamous cell carcinoma. PMID- 11263297 TI - [The clinical significance of biological characters on non-small cell lung cancer]. AB - OBJECTIVE: To study the biological characters of 137 cases of surgically treated NSCLC and their correlation with stages, types and survival rates. METHOD: Electronmicroscopy, immunohistochemistry and FCM were employed in this prospective, randomized study. K-P curve, COX univariance and multivariance were used for statistic analysis. RESULTS: (1) Electronmicroscopy showed that 40% of squamous cancer and 40% of adenocarcinoma diagnosed microscopically were mixed type. (2) The positive rate of p53 expression increased in advanced stage of NSCLC, and the survival rate was lower in cases with positive p53 expression. (3) A higher survival rate was seen in cases with positive Hras expression. (4) Lower survival rates were seen in cases with abnormal DI and PI. (5) Miscellaneous: expression rates of c-erbB2 and PCNA were related to adenocarcinoma and mixed type electronmicroscopically confirmed respectively. CONCLUSION: From this study p53, Hras, DI, PI, c-erbB2 and PCNA were found related to stages, types and survival rates, respectively. PMID- 11263298 TI - [Study on diagnostic value of analyzing argyrophilic nucleolar organizer regions in benign and malignant pleural effusions]. AB - OBJECTIVE: To evaluate the diagnostic value of argyrophilic nucleolar organizer regions (AgNOR) measurement in benign and malignant pleural effusions. METHODS: Pleural effusions from 50 patients with proven malignant disease, 30 patients with benign disease were studied using the one-step silver staining method. Six cytologically atypical samples were also included in this study. RESULTS: The mean AgNOR count in malignant cells was significantly higher than that in benign mesothelial cells (7.6 +/- 1.4 vs 2.3 +/- 0.7, P < 0.01). The morphological features of AgNOR in malignant cells were mainly of diffuse type (80%), while in benign mesothelial cells, mainly of nucleolar type (90%). Among the six cytologically atypical samples, four were in the malignant range, two were in the benign limits, and these were proved by histopathology. CONCLUSION: AgNOR study may be useful in differential diagnosis of benign and malignant pleural effusions. PMID- 11263299 TI - [Cloning and mapping of the rpoB gene in M. tuberculosis]. AB - OBJECTIVE: To clone the rpoB gene of M. tuberculosis. METHODS: Screening the rpoB gene of M. tuberculosis from M. tuberculosis genomic DNA library by using the rpoB gene conservative region PCR product as a probe. RESULTS: 411 bp products were seen after amplification of H37Ra DNA. The cloning and sequencing results indicated that the 411 bp products were homogeneous to M. tuberculosis rpoB gene. 12,000 clones of M. tuberculosis genomic DNA library were screened by hybridization using the 411 bp product as a probe and 7 clones seemed positive, 3 clones were real positive after the second hybridization. 1 of them carried a 3.8 kb insert fragment. The preliminary restriction map of the 3.8 kb segment was made. The regions which were found homogeneous to the probe to the end of this cloned fragment were 1 and 2.8 kb. CONCLUSION: In comparison with the open reading frame of H37Rv rpoB gene, the 3.8 kb segment cloned here covers larger portion of entire rpoB gene of M. tuberculosis. This study will provide a useful tool to study the resistant mechanism of rifampin and other rifamycin compounds to M. tuberculosis. PMID- 11263301 TI - [Lung volume reduction surgery for diffuse emphysema]. PMID- 11263300 TI - [Report on side effects of antituberculous agents]. AB - OBJECTIVE: To illustrate the side effects of antituberculous agents and the principles to cope with them. METHODS: Various side effects resulting from antituberculous agents were confirmed directly or indirectly in 62 cases. The distribution, time of presentation, and manifestation of the side effects were observed and analyzed. RESULTS: Allergic reaction showed highest frequency in 37 cases (60%). Serious side effects, which were mainly caused by rifamycin agents, occurred within 2 hours after administration. Overdosage tended to increase the occurrence of side effects. Different agents might result in serious side effects at the same time or in succession. CONCLUSION: Once serious side effects appear, the agents causing the effects must be stopped and suitable remedies should be introduced immediately or the consequences might be disastrous. Repeated use of rifamycin agents may aggravate allergic reaction, thus detailed history of previous treatments must be recorded. For patients with hepatitis, allergy history, alcoholics and weak senile patients, close observation is required during the process of treatment. For patients allergic to more than one drug, no matter the reaction is serious or not, the principle is to stop the used drugs and introduce desensitization therapy immediately. If the allergen is to be identified, the testing dosage should be less than 10 per cent of the original dosage, along with prepared procedures for emergen tresale. Repeated tests are not suggested for patients with serious allergic reaction. PMID- 11263302 TI - [Concurrent radiotherapy and chemotherapy for non-small cell lung cancers]. PMID- 11263304 TI - [Breast reconstruction using pedicled three-lobe transverse rectus abdominis musculocutaneous flap]. AB - OBJECTIVE: An innovated transverse rectus abdominis musculocutaneous flap, the three-lobe TRAM flap was designed for breast reconstruction. METHODS: A triangle shaped skin over the upper part of the rectus abdominis muscle was included in the conventional TRAM flap for the purpose of adding more skin for breast reconstruction and enhancing blood supply of the skin portion of the TRAM flap. RESULTS: In this group of six patients subjected to mastectomy and radiation therapy for breast cancer, complete survival of the three-lobe TRAM flap was obtained in 4 cases. Two patients had minor skin necrosis and fat liquefaction. Better breast contour and contracture release of the axilla were also achieved. CONCLUSION: This innovation of conventional TRAM flap for breast reconstruction has some advantages. It is especially useful for patients complicated with radiation lesions on the chest and axilla areas. PMID- 11263303 TI - [Prophylactic and therapeutic effects of beclomethasone inhalation on bronchial hyperresponsiveness and asthma: a randomized, double-blind, parallel-group controlled trial]. AB - OBJECTIVE: To better understand the effects of corticosteroid inhalation on asthma and asymptomatic bronchial hyperresponsiveness (BHR). METHODS: A double blind, randomized study was conducted to compare the effects of beclomethasone dry powder (BDP, 600 micrograms/day) inhalation with placebo on BHR and asthmatic symptoms in 59 students (12-18 yrs) for one year. RESULTS: After one year of treatment, lgPD20-FEV1 increased significantly in asthmatics in the BDP group (0.385 +/- 0.424 vs 1.187 +/- 0.603 mumol histamine). Thirty percent of asthmatics in BDP group and 86% in the control group developed symptoms in this year (P = 0.076). There was no significant difference in lgPD20-FEV1 between the BDP and the control group in asymptomatic BHR students. However, none in the BDP group of asymptomatic BHR students developed asthmatic symptoms compared with 3 cases (15%) in control group. The accumulative symptom score showed significant difference between the BDP group and the control group in asymptomatic BHR students (1.50 +/- 2.54 vs 5.58 +/- 6.22, P < 0.01). CONCLUSION: The results suggested that BDP could significantly relieve the severity of BHR and symptoms in asthmatics, and could have some prophylactic effects on the development of asthma in those with asymptomatic BHR. PMID- 11263305 TI - [Primary experience in inframammary crease reconstruction]. AB - OBJECTIVE: To improve the result of augmentation mammaplasty. METHODS: The authors selected 15 patients whose primary breast volume was less than 70 ml. During augmentation mammaplasty on these patients, the inframammary creases were reconstructed by lifting and fixation under the pectorales major. The paper elaborated on the principle, design and method of the operation. RESULTS: Postoperative follow-up for 6 to 12 months showed good results. CONCLUSION: This method is simple, of rational design and leaving no scar on the skin. PMID- 11263306 TI - [Prevention and treatment of eyelid retraction and ectropion following lower eyelid blepharoplasty with tarsal tuck procedure]. AB - OBJECTIVE: To prevent or decrease eyelid retraction and ectropion following lower eyelid blepharoplasty. METHODS: The tarsal tuck procedure was performed during lower eyelid blepharoplasty to tighten the lower eyelid. This method had been used since 1991. RESULTS: After the operation, the lower eyelid was tensional and steady. The complications of eyelid retraction and ectropion were diminished. CONCLUSION: The pathologic basis of the eyelid bag is that the supporting tissues become lax so that the lower eyelid and lateral canthus move downwards. The exact aim of the tarsal tuck procedure is to correct these pathologic changes. PMID- 11263307 TI - [Statistical analysis of complications of the frontalis aponeurosis flap for correction of complete blepharoptosis]. AB - OBJECTIVE: The cause of and prophylactic measures for the complications of the frontalis aponeurosis flap suspension for correction of blepharoptosis were investigated to improve the operative results. METHODS: Five hundred and thirteen patients who were found to have complications after frontalis aponeurosis flap suspension for blepharoptosis were studied by follow-up examinations. RESULTS: Fourteen kinds of postoperative complications were found and cured. CONCLUSION: Most of the postoperative complications following blepharoptosis correction can be prevented and well treated. The frontalis aponeurosis flap technique is reliable for correction of complete blepharoptosis. PMID- 11263308 TI - [Surgical treatment of maxillary protrusion by osteotomy of both anterior jaws]. AB - OBJECTIVE: In order to study the simple and practical surgical treatment of maxillary protrusion. METHODS: The wedge-shaped, subapical osteotomy of the maxilla and mandible was performed under local anesthesia. The procedure should not enter the nasal cavity. Some patients had preoperative or postoperative orthodontic treatment. RESULTS: The operation results in 46 patients were all satisfactory in terms of facial configuration and occlusion relationship. No complications occurred. CONCLUSION: This method is simple and less traumatic with good exposure of operative field and larger scope of movement. It is one of the ideal methods of surgical orthodontics for mild or medial maxillary and mandibular deformities. PMID- 11263309 TI - [Monitoring of serum lidocaine concentration in tumescent liposuction infiltrated with high-dosage lidocaine and its clinical significance]. AB - OBJECTIVE: This study was carried out to determine the maximum and safe dosage of lidocaine administered in tumescent liposuction. METHODS: In 14 cases of tumescent liposuction using high-dosage, low-concentration lidocaine as local infiltration anesthetic, serum lidocaine concentrations were monitored with automated fluorescence polarization analysis. RESULTS: The time-concentration curve together with clinical manifestations indicated that in tumescent liposuction, lidocaine, under a concentration of 0.1 mg/ml mixed with 1/1,000,000 to 1/2,000,000 epinephrine, could be administered with a dosage of 35 mg/kg body weight as a local infiltration anesthetic while the serum lidocaine was still within the safety limit and no signs of intoxication manifested. The resulting diminution both in pain and blood loss during the operation enhanced the overall operation safety and suction volume. CONCLUSION: This study provides the basis for high-dosage lidocaine administration in tumescent liposuction. PMID- 11263310 TI - [The medialis pedis flap based on the medial fasciocutaneous branches of the dorsal pedal artery]. AB - OBJECTIVE: This is to report the design and application of the medialis pedis flap. METHODS: Based on the medial fasciocutaneous branches of the dorsal pedal artery, a medialis pedis flap can be raised. The flap had been used in 18 patients with soft tissue defects of the dorsum pedis, ankle region, lower leg or hand. RESULTS: The anatomic observation of the blood supply to the medial region of the foot showed that a constant distribution of fasciocutaneous branches issued from the dorsal pedal artery was an important blood supply to this area. All flaps used in the 18 patients survived completely with satisfactory results and minimal morbidity at the donor site. CONCLUSION: We believe that the medial fasciocutaneous branches of the dorsal pedal artery are the reliable pedicle of the medialis pedis flap. The flap is useful for reconstruction of the soft tissue defects around the foot, ankle, lower leg or hand. PMID- 11263311 TI - [Repairing the soft tissue defect in the heel with a double-bridge pattern flap of the posterior ankle]. AB - OBJECTIVE: This is to introduce a method for repairing the soft tissue defect in the heel. METHODS: The double-bridge pattern flap in the posterior ankle region was designed with the pedicles of the flap being in each side of the malleoli. According to the defect, the lower bridge-pattern flap with an arc-shaped upper margin was designed first. Then the upper bridge-pattern flap was designed with a "lambda"-shaped margin. The flap was raised with the deep fascia and advanced down to cover the defect. In dissecting the flap, care should be taken not to damage the posterior tibial nerve and blood vessel bundle. The flap was used in 37 patients. RESULTS: All the flaps survived except 2, in whom partial necrosis occurred in the upper bridge-pattern flap; they healed after dress changings. Follow-up of more than one year indicated that no ulcer took place in all the transferred flaps. CONCLUSION: The blood supply and elasticity of the flap are good. Its sensation and resistance to friction and shearing are satisfactory. Distinguishable distance of two points in the flaps was 13 to 14 mm. PMID- 11263313 TI - [Conversion from a tridimensional surface to a plane for measuring expanded skin area with computer-aided technique]. AB - OBJECTIVE: To seek a reliable and practical method to measure the expanded skin area. METHODS: In our technique, the three-dimensional surface overlaid with the expanded skin was converted to a flat one through a special surface molding. This irregular plane area was scanned digitally to the computer and measured by means of counting its total pixels. With the areas of the underlying base and the defect measured in the same way, we could determine whether the extra tissue after expansion was sufficient for clinical repair at any stage of expansion. RESULTS: The deviation of this protocol was less than 3 percent at most in our preliminary reliability demonstration. CONCLUSION: This protocol can be used to estimate the expanded skin area in the clinical and experimental applications. PMID- 11263312 TI - [The study on the role of apoptosis suppressive gene bcl-2 in the pathogenesis of hemangioma]. AB - OBJECTIVE: To elucidate the relationship between bcl-2 and hemangioma. METHODS: We investigated the expression of bcl-2 in 38 samples of hemangioma (19 cases of capillary hemangioma, 10 cases of cavernous hemangioma, 9 cases of racemose hemangioma) and 6 samples of normal skin by immunohistochemical SP method. RESULTS: The results showed that the expression of bcl-2 was higher in cavernous and racemose hemangioma than in the controls, the expression of bcl-2 was lower in capillary hemangioma than in the controls. The differences were statistically significant. CONCLUSION: It can be deduced from the results that there is an intimate relationship between bcl-2 and the growth of cavernous and racemose hemangioma, and the lower expression of bcl-2 in capillary hemangioma shows that capillary hemangioma may disappear naturally. PMID- 11263314 TI - [The clinical application of triamcinolone to accelerate soft tissue expansion]. AB - OBJECTIVE: To speed up the expansion procedure. METHODS: Triamcinolone was injected during tissue expansion in 26 cases. RESULTS: The study proved that triamcinolone shortened the injection interval to 32 +/- 6 hours. The mean time of full expansion was shortened to 20 +/- 5 days in the experimental group. The injection interval and mean time of full expansion were 6 +/- 2 days and 42 +/- 8 days respectively in the control group (P < 0.01). The immediate retraction ratios of expanded skin of the two groups were 17% and 26% respectively. CONCLUSION: This new method is feasible in clinical application. PMID- 11263315 TI - [Allotransplantation of cryopreserved fetal cartilage in plastic surgery]. AB - OBJECTIVE: To study the applicability of homologous fetal cartilage in plastic surgery. METHODS: Limb and costal cartilage were taken from the dead fetus. The fetal cartilage was then frozen at -80 degrees C and preserved in -196 degrees C liquid nitrogen with a cryoprotective agent. A total of 38 patients received fetal cartilage transplantation. Of them there were 24 saddle nose, 4 auricle defects, 6 secondary deformities of cheiloschisis, 4 mandible deformities. RESULTS: Through follow-up of three and a half years, it was found that none of the transplanted cartilage was absorbed, deformed or rejected due to immunological reaction. The contours were satisfactory except 2 cases with auricle reconstruction. CONCLUSION: The clinical application of homologous fetal cartilages is of value in plastic surgery. PMID- 11263316 TI - [Preliminary experimental study on free radicals in expanded soft tissue]. AB - OBJECTIVE: To inquire into the formation of free radicals during tissue expansion and its influence on tissue expansion. METHOD: Expanders of 20 ml were placed beneath the dorsal skin of SD rats, and the conventional expansion course took about 23 to 25 days. The activity of SOD and the content of MDA in skin were determined prior to the last expansion, 1 h, 6 h, 12 h and 24 h after the last expansion, respectively. Meanwhile, the effect of SOD on expansion area and length was observed. RESULT: The activity of SOD and the content of MDA at 1 h, 6 h, and 12 h after expansion were significantly different from those before expansion, while the activity of SOD and the content of MDA at 24 h after expansion were almost the same as those before expansion. The difference of expansion area and length between SOD and NS groups was significant, especially during 16 to 24 days. CONCLUSION: Tissue expansion produces free radicals and SOD can increase expansion area and length. The study provides a clue to find a way for safe, rapid and effective soft tissue expansion. PMID- 11263317 TI - [Experimental study on the mechanisms of enhancement of aerobic glycolysis of muscles in burns and sepsis--the verification of the existence and the enhancement of the aerobic glycolysis of muscles in early postburn period and sepsis]. AB - OBJECTIVE: To verify if aerobic glycolysis exists in the muscle cells in normal rats and to analyze the changes in aerobic glycolysis in the muscle cells in the early postburn period and in septic states. METHOD: Using septic model of rats, we established the in vitro muscle incubation system with sufficient oxygen supply as well as the NADH fluoremetric method for the detection of trace amount of lactate in the samples. Extensor digitorium longus (EDL) and soleus muscles which represent two types of muscle fiber were studied. RESULTS: In the early postburn period as well as in septic states, the lactate production of muscle cells was significantly elevated as compared to the normal controls even though the muscles were incubated in a fully oxygenated media. The levels of aerobic glyoclysis, as well as its changes in postburn period and in septic states, vary depending on the difference in the fiber composition of the muscles. CONCLUSION: Muscle cells might develop a kind of metabolic enhancement which is referred to as aerobic glycolysis rather than the metabolic defect which results from tissue hypoperfusion and hypoxia in the early postburn period as well as in septic states. This provides us a special insight to elucidate the mechanisms of the metabolic derangement in the early postburn period and in sepsis. PMID- 11263318 TI - [Anatomical basis for the newly developed facelifting]. AB - OBJECTIVE: To investigate the relationship between the SMAS and the facial nerve divisions in the cheek area. METHOD: We dissected 12 cadaver heads(24 sides). RESULTS: 1. SMAS distributed over the middle face. It became thinner as it extended forward and there was a small aperture lateral to the mouth. The branches of facial nerve lay directly beneath the parotidomassetric fascia after emerging from the parotid gland. 2. The frontal branch penetrated the deep fascia to the SMAS at about 0.5 cm below the zygomatic arch. 3. Some buccal branches went through the cheek fat pad while the others lay on its surface directly under the thin SMAS. 4. There was constantly a zygomatic ramus went into the upper one third of the zygomaticus. It innervated the lower and lateral orbicularis oculi muscle in 9 of 24 sides (37.5%) of the cadaver heads, zygomaticus major and minor and orbicularis oculi muscle in 8 of 24 sides (33.3%), and the zygomaticus major and minor in 7 of 24 sides (29.2%). CONCLUSION: During facelifting, wide dissection of the SMAS should be done directly above the parotidomassetric fascia and 0.5 cm below the zygomatic arch. While approaching the middle face, the dissection should be limited to the lower two third of the zygomaticus, followed by elevating the zygomatic fat. The authors dissect the nasolabial fold through the lower eyelid incision, then the dissection should go downwards, under the orbicularis oculi muscle, to the nasolabial area. In a clinical situation, care must be taken not to damage the facial nerve trunk, and dissection of the upper one third of the zygomaticus should be avoided. PMID- 11263319 TI - [The expression of ICAM-1 on the keratinocytes in second degree human burn skin]. AB - OBJECTIVE: To investigate the significance of changes in intercellular adhesion molecule-1 (ICAM-1) expression on keratinocytes(KCs) during wound healing of second degree burn. METHODS: With immunohistochemistry technique, the ICAM-1 expression in second degree burn wounds in patients were investigated at four different postburn periods. The normal skins from 6 burn patients (BNG), the normal skins from 8 healthy volunteers (HVG) and the scar tissues (SG) from 8 patients served as controls. RESULTS: There was poor expression of ICAM-1 on basal cells in normal skins of healthy volunteers, while the expression of ICAM-1 enhanced in normal skins of burn patients. On basal cells in second degree burn wounds, the expression of ICAM-1 slightly increased in the first week postburn, obviously enhanced in the second week, and in the third or fourth week, the expression continued to keep strong. In addition, the ICAM-1 in KCs showed polar distribution, and the expression of ICAM-1 markedly enhanced on the multilayer KCs in neogenetic epithelium and the positive cells arranged columnarly. CONCLUSION: The enhanced expression of ICAM-1 on KCs in second degree burn wounds might be related to invasion of inflammatory cells beneath wound. It might play a role in inducing proliferation and migration of KCs and accelerating epithelization. PMID- 11263320 TI - [Effects of thermal injuries on electron transport chains of rat myocardial mitochondria]. AB - OBJECTIVE: The electron transport chain alterations of myocardial mitochondria and cardiac dysfunction were studied after severe burn injury. METHODS: Contraction properties of male Sprague-Dawley rat cardiac muscle were investigated at 2, 4, 6 hours after 30% TBSA full-thickness thermal injury. Meanwhile, changes in electron transport chains of myocardial mitochondria were measured. Mitochondria were obtained by differential centrifugation. Succinate respiratory chains and NADH-respiratory chains were assayed polarographically and spectrophotometrically in isolated myocardial mitochondria respectively. RESULTS: The results showed that thermal injury led to decrease in the activities of two respiratory chains. At 2 h postburn, the activities of NADH-cytochrome C reductase and cytochrome oxidase declined significantly as compared with that of the sham-operated group, and at 4 h postburn, all activities of succinate-Co. Q reductase, succinate-cytochrome C reductase, NADH-Co. Q reductase were much lower than those of the sham-operated group. Along with the electron transport chain alterations of myocardial mitochondria, there were decrease in myocardial contractile function in burned rats. CONCLUSION: These results imply that weakness of myocardial contractile function resulting in the decrease in cardiac output may be associated with the impairment of utilization of oxygen in myocardial mitochondria following burn injury. PMID- 11263321 TI - [Expression of adhesion molecules beta 1 integrin on the surface of fibroblasts in wound healing]. AB - OBJECTIVE: To investigate the relationship between the expression of adhesion molecules integrin beta 1 and the synthesis of collagen in wound healing. METHODS: In this study, integrin beta 1 expression on fibroblasts derived from granulated wound and from normal skin was compared by the technique of immunoelectron microscope and flow cytometry. The differences of ultrastructure between the two groups of fibroblasts were also observed. RESULTS: The results showed that the expression of alpha 5 and beta 1 subunits on fibroblasts derived from granulated wound were higher than those from normal skin. Endoplasmic reticulum attached with more ribosome in the former cells was richer than that in the latter cells. CONCLUSION: These results suggest that the expression of adhesion molecules integrin beta 1 appears to be associated with the synthesis of collagen in wound healing. PMID- 11263322 TI - [The origin of calcitonin gene-related peptide (CGRP) in burned skin of rats]. AB - OBJECTIVE: To investigate the origin of calcitonin gene related-peptide (CGRP) in skin burn wound. METHODS: Immunohistochemistry technique was used to determine change in distributive density in CGRP-containing nerve fibers in the wound, wound margin, remote intact skin of rats during the first 96 hours postburn. RESULTS: It was shown that the distributive density of CGRP-containing nerve fibers in all areas decreased at 15 min postburn, but the density recovered earlier in wound margins compared to other sites. In addition, CGRP immunoreactive positive cells, which emigrated from blood vessel in dermis underneath the wound and wound margin, were related to CGRP-containing nerve fibers at 12 hours. Stronger staining intensity to CGRP was observed in those cells and they disintegrated and released CGRP immunoreactive materials into the dermis at 24 hours. Thereafter, those cells disappeared. CONCLUSION: CGRP not only is released from cutaneous nerve, but also is synthesized and released by immune cells in response to burns. PMID- 11263323 TI - [Effect of hypertonic saline/dextran 70 on delayed resuscitation of dogs with burn shock]. AB - OBJECTIVE: To investigate the effect of hypertonic saline/dextran 70 on delayed resuscitation of burn shock. METHODS: Eighteen mongrel dogs with 35% TBSA, third degree burn were used in this study. Lactated Ringer's (LR) or 7.5% NaCl+ 6% dextran 70 (HSD) was given for resuscitation 6 h postburn. The volumes and rates of fluid infusion were controlled basically on the urinary output of 1.0 ml.kg 1.h-1 and cardiac output (CO) of 70%-80% of preburn values. The volume load, +dp/dtmax, -dp/dtmax, CI,DO2 and VO2 were obtained to evaluate the effect of HSD resuscitation. RESULTS: The resuscitated volume of HSD was 30.56% less during first 24 h postburn and 59.50% less at 4 h after resuscitation than LR's. The +dp/dtmax, CI,DO2 and VO2 were increased significantly with HSD infusion at 2 h, 1 h and 0.5 h after resuscitation compared with LR's. CONCLUSION: HSD could expand plasma volume significantly with small quantity. The cardiac contractility was enhanced and the oxygen delivery, oxygen consumption were increased in delayed resuscitation of burn shock. PMID- 11263324 TI - [Rehabilitation of 269 fingers with electrical burns]. AB - OBJECTIVE: To summarize the clinical feature of the fingers with electrical burns. METHODS: 269 fingers with electrical burns were analysed in the characteristics of injury and the types of repair. RESULTS: Fingers with electrical burns were often multiple, characterizing the complexity of the wound repair. Operation time should be determined by different conditions. Usually the fingers with electrical burns should be operated early but the circumferential wound surface should be depressed or eschar excised during early stage in order to avoid ischemia of distal segment. When the circulation of the finger distal to the injury is improved, operation should be done to repair wound surface. CONCLUSION: Many kinds of flaps might be used for repair of fingers with electrical burns. The flaps from the same finger should be the first choice, followed by flaps from the same hand and distant flaps. PMID- 11263325 TI - [Effects of transforming growth factor beta-1 on cicatrix formation]. PMID- 11263327 TI - [Efficient expression of multidrug resistance gene mdr1 in retroviral vector under control of an internal ribosome entry site]. AB - OBJECTIVE: To test the efficiency of human mdr1 gene expression under the control of an internal ribosome entry site (IRES). METHODS: The expression retroviral vector Halmdr1 was constructed from pSXLC/pHa system which contains an IRES from encephalomyocarditis virus. The vector was introduced into ecotropic packaging cells GP + E86 by liposome-mediated transfection. The retrovirus producing cells were obtained by 50 micrograms/L vincristine selection. The success of transfer and expression of mdr1 gene were determined by polymerase chain reaction (PCR) and flow cytometry (FCM). RESULTS: Virus in the supernatant of the producer cells, in which the integration of mdr1 gene was confirmed by PCR, was titrated to 2.0 x 10(5) cfu/ml. The selected producer cells exhibited a 24-52-fold increase of resistance to vincristine, daunorubicin and taxol in comparison with control cells GP + E86. The expression of mdr1 gene was confirmed by both reverse transcription PCR at RNA level and FCM at protein level, although the HaImdr1 transfectants showed somewhat less expression of P-gp when compared to that with cap-dependent translation of Ha mdr1. CONCLUSION: mdr1 gene can be effectively translated and expressed under the control of IRES in cap-independent manner, it may be used as a dominant selectable marker in bicistronic vectors for gene therapy. PMID- 11263326 TI - [The distribution of the frontal branch of the facial nerve and its significance in facelift]. AB - OBJECTIVE: To further understand the distribution and course of the frontal branch of the facial nerve in the temporal and zygomatic areas. METHODS: Facial dissection was performed on 9 cadaver heads(18 sides). RESULTS: It was found that the frontal of the facial nerve had 3 to 7 divisions (average 5) instead of one. The divisions of the frontal branch showed no constant course and distributed nearly all over the zygomatic arch. We divided the zygomatic arch into 3 equal parts and found that the middle third had the highest density of the branch distribution, accounting for 44.44% of the total divisions; the anterior third and the lateral third possessed 38.89% and 16.66% of the total divisions respectively. These divisions entered deeply into the temporoparietal fascia(the superficial temporal fascia). The divisions in the anterior third of the zygomatic arch innervate the lower and lateral orbicularis oculi muscle; the divisions in the middle third innervate the lateral orbicularis oculi muscle and the frontal muscle; the divisions in the lateral third innervate the frontal muscle and the anterior auricularis. CONCLUSION: We recommend discriminating the layers of the soft tissue over the zygomatic arch, not limiting the dissection range of the zygomatic arch to avoid damaging the nerve divisions while doing subperiosteal dissection of the zygomatic arch in facelifting. PMID- 11263328 TI - [Construction of retrovirus vector of bcr/abl mRNA cleaving ribozyme gene and its effects on K562 cells]. AB - OBJECTIVE: To investigate the effect of bcr/abl fusion gene on the growth of chronic myeloid leukemia(CML) cells and to explore the feasibility of ribozyme in CML gene therapy. METHODS: A hammerhead ribozyme DNA targeting the bcr/abl (b3a2) transcript was synthesized, and recombinated into retroviral vector pLXSN forming pLRZXSN recon. By lipofectin mediated DNA transfection technique, pLRZXSN was introduced into K562 cells. The effects of the ribozyme on the growth of K562 cells and apoptosis were studied by leukemic colony assay, flow cytometry (FCM), reverse transcript-polymerase chain reaction (RT-PCR), detection of DNA fragmentation and electron microscope. RESULTS: 1. The number of K562 cell colony was inhibited by 85% after the cells were transfected for 48 hours. 2. The expression of bcr/abl mRNA and the fusion protein P210 was decreased sharply in K562 cells transfected with pLRZXSN for 48 and 72 hours, respectively. 3. The characteristics of apoptosis was revealed in K562 cells transfected with pLRZXSN, i.e. sub-G1 peak, DNA fragmentation and morphological changes. CONCLUSION: The ribozyme was capable of inhibiting the proliferation of K562 cells and inducing the cell apoptosis. PMID- 11263329 TI - [Effect of WT1 gene expression on cell growth and proliferation in myeloid leukemia cell lines]. AB - OBJECTIVE: To explore the effect of WT1 antisense oligonucleotide(AS-oligo) on cell proliferation and apoptosis in myeloid leukemia cell lines. METHODS: K562 and HL-60 cells were cultivated with WT1 AS-oligo. The cell proliferation, apoptosis, cell cycle and gene expression were examined by MTT colorimetry, FACS and RT-PCR. RESULTS: WT1 AS-oligo could inhibit the proliferation of K562 cell and induce apoptosis of K562 and HL-60 cells. On the contrary, the growth of HL 60 cells and the expression of WT1, mdm2 and bcl-2 genes were unaffected. CONCLUSION: WT1 gene is related to the proliferation and apoptosis of leukemic cells. WT1 gene could suppress cell apoptosis independent of status of p53 and bcl-2 genes. It might play an role in leukemogenesis. PMID- 11263330 TI - [Enhancement of apoptotic sensitivity induced by UV irradiation on the p53 transducted K562 cells]. AB - OBJECTIVE: To answer whether wild-type p53 can sensitize K562 cell apoptosis induced by UV irradiation. METHODS: K562 cells transducted with retrovirus encoding wild-type p53 were irradiated by ultraviolet for different time and then cultured for different time. Apoptosis was detected by TDT end labelling technique, DNA fragmentation and MTT assay. RESULTS: TDT end labelling and DNA fragmentation showed that apoptosis induced by 8 min UV-irradiation differed significantly between K562-neo and K562-p53 cells. CONCLUSION: Wild-type p53 can enhance apoptosis induced by UV-irradiation on K562 cells. PMID- 11263331 TI - [Experimental study on mice scheduled-bone marrow transplantation]. AB - OBJECTIVE: To enhance the grafting efficiency of bone marrow transplantation. METHODS: Lethally irradiated recipient Kunming mice were transplanted with bone marrow cells from normal Kunming mice either in one large (10(7)) number (BMT group) or in four separate small (10(5)-10(6)) numbers (scheduled BMT group) and the survival rate, hematopoiesis reconstitution and acute graft versus host disease (aGVHD) were compared between the two groups. RESULTS: Both BMT and SBMT (10(5) x 4) groups obtained the same survival rate of 30%. In SBMT (10(6) x 4) group, the peripheral WBC count, bone marrow nucleated cells and CFU-E, CFU-GM, CFU-S and CFU-F yields all returned to normal 17 days after irradiation. The degree of GVHD in SBMT group was less severe than that in BMT group, and the survival rate (60%) was significantly higher than that in BMT group (30%). CONCLUSION: SBMT can significantly enhance the grafting efficiency. PMID- 11263332 TI - [Establishment and biological characterization of a transplantable BALB/c mouse erythroblastic leukemia model]. AB - OBJECTIVE: Biological characterization of a BALB/c mouse transplantable erythroblastic leukemia model EL9611. METHODS AND RESULTS: The original erythroblastic leukemia (EL) mouse was obtained by 7,12-DMBA induction. The spleen cell suspension from the original EL mouse was injected into the same inbred BALB/c strain mice. After successive transplantation for 20 generations, the incidence of EL was 100% without spontaneous remission. There was a linear relationship between the survival time of the EL mice and the number of leukemic cells inoculated. When 10(6) leukemic cells were inoculated intravenously, the EL mice survived 10.2 +/- 1.3 days. Invasion of leukemic cells involved mainly in the bone marrow, the spleen and the liver. There was no obvious infiltration of leukemic cells in lymph-nodes and thymus. In peripheral blood, there were a large number of leukemic cells and most of them were basophilic or polychromatophilic erythroblasts. At the advanced stage, the mice developed anemia and thrombocytopenia. The reaction of the leukemic cells for hemoglobin staining was positive or weakly positive. While the peroxydase reaction was negative, Thy-1 antigen and Fc-recepter were not presented. No virus-like particles was found in the cells under electron microscope. EL9611 leukemia model was sensitive to some antitumor drugs used in clinical therapy. CONCLUSION: The establishment of EL9611 leukemia model offered a useful tool for studying proliferation and differentiation of erythroid cells, as well as pathogenesis and experimental treatment of non-virus erythroid malignancy. PMID- 11263333 TI - [Effect of intravenous injection of interleukin-8 on mouse hematopoietic progenitor cell mobilization]. AB - OBJECTIVE: To study the mobilization effects of recombinant human IL-8(rhIL-8) on mouse peripheral blood hematopoietic progenitors (HPCs). METHODS: A daily single dose of rhIL-8 was intravenously injected into normal or G-CSF-treated mice for two days. The number of colony-forming unit-Mix (CFU-Mix) in peripheral blood was analyzed by colony assay, while the number of c-kit+ Sca-1+ HPCs was examined by immunofluorescence staining. In addition, colony forming unit of spleen (CFU-S) and marrow repopulating ability (MRA) cells were examined by transplanting lethally irradiated mice. RESULTS: Single dose of rhIL-8 (30 micrograms) significantly increased the numbers of CFU-Mix[(194 +/- 61) x 10(3)/L] and c-kit+ Sca-1+ cells [(3186 +/- 517) x 10(3)/L] in the peripheral blood as compared with that of normal saline controls [(7 +/- 3) x 10(3)/L and (644 +/- 341) x 10(3)/L] at 15 minutes after intravenous injection. G-CSF could also increase peripheral CFU-Mix and c-kit+ Sca-1+ cells to (790 +/- 48) x 10(3)/L and (10,729 +/- 2339) x 10(3)/L, respectively, after subcutaneously injection for two days, this increase was further augmented by rhIL-8 injection. IL-8 alone or in combination with G CSF could significantly increase the number of CFU-S and MRA cells in circulating blood. CONCLUSION: IL-8 alone or in combination with G-CSF could significantly mobilize circulating HPCs. PMID- 11263334 TI - [Effect of HHV-6 on expression of M-CSF in patients with hematological disorders]. AB - OBJECTIVE: To study the relationship between HHV-6 infection and abnormal expression of M-CSF in bone marrow mononucleated cells (MNC) from patients with hematological disorders. METHODS: M-CSF expression level were detected in 30 specimens of MNC from patients with hematological diseases by ABC immunocytochemical staining and reverse transcription polymerase chain reaction methods. RESULTS: The percentage and score of M-CSF antigen positive cells, and percentage of M-CSF mRNA expressing samples derived from HHV-6 infected group were 36.9% +/- 24.9%, 45.9 +/- 31.4 and 83.3%, respectively, which were significantly higher than those of control (20.3% +/- 19.4%, 25.3 +/- 24.2 and 50.0%, respectively) (P < 0.001). HHV-6 could promote M-CSF production to a large extent in MNC from leukemic patients than that from patients with benign hematological disorders. M-CSF existed mainly on membrane or in cytosol of granulocytes and monocytes. CONCLUSION: HHV-6 infection in vitro could promote M CSF production in MNC from patients with hematological diseases. PMID- 11263335 TI - [Study on hepatitis G virus infection in 63 posttransfusion patients with hematological diseases]. AB - OBJECTIVE: To study the hepatitis G virus(HGV) infection in posttransfusion patients. METHODS: HGV RNA and HCV RNA were detected by RT-PCR, and anti-HGV and HBsAg were assayed by ELISA in 63 posttransfusion patients with blood diseases. RESULTS: The positive rates of HGV RNA, HCV RNA and anti-HGV were 7.9%, 46.0% and 6.3%, respectively. HGV infection was usually accompanied by HCV infection and elevation of ALT. CONCLUSION: HGV as well as HCV may transmitted by blood transfusion. PMID- 11263336 TI - [Hematopoietic stem cell transplantation technique and its applications]. PMID- 11263337 TI - [Studies of tumor necrosis factor-alpha and red cell immunity state in multiple myeloma]. AB - OBJECTIVE: To explore factors in the pathogenesis of multiple myeloma (MM). METHODS: The plasma level of tumor necrosis factor-alpha (TNF alpha) was measured by immunoassay in 21 MM patients, and red cell immunologic function was assayed in 16 patients. RESULTS: There was no difference in TNF alpha levels among patients with either fulminant MM or stable MM and normal controls. The rosette formation rate of red cell C3 bR and red cell immune complex was significantly lower in MM patients than in the controls. CONCLUSION: MM patients might have impaired monocyte-phagocyte system and red cell immune function. PMID- 11263338 TI - [Report of 18 cases lymphoblastic lymphoma]. AB - OBJECTIVE: To identify the clinical, pathological, and immunological characteristics of lymphoblastic lymphoma (LBL). METHODS: The relation of clinical features, cellular morphology and immuno-phenotypes to the treatment outcome and prognosis was analyzed in 18 LBL patients. RESULTS: The patients were 15 males and 3 females with a median age of 27 years. Fourteen cases showed bone marrow infiltration, 10 lymphoma leukemia and 9 mediastinal tumor. Fifteen cases were of convoluted cell type expressed T cell marker, three patients were of B cell type and all achieved complete remission. The 5 year survival rate of the 18 cases was 11.2% with a median survival duration of 10.1 months. CONCLUSION: LBL was found predominantly in male with a younger onset age, and was characterized by involvement of bone marrow and mediastina. The majority of LBL patients were of convoluted cell type expressed T cell marker. The patients showed poor response to therapy, while B-cell type showed a better prognosis than T-cell type. PMID- 11263339 TI - [The expression of CD44, PCNA, vWF: Ag in non-Hodgkin's lymphoma and its relationship with the grade of malignancy]. AB - OBJECTIVE: To study the expression of CD44, proliferating cell nuclear antigen (PCNA), vWF: Ag in normal lymphatic tissue and non-Hodgkin's lymphoma (NHL) and its implication. METHODS: Immunohistochemical assay was performed with a LSAB kit. RESULTS: 1. In normal lymphatic tissues, mature appearing small lymphocytes were positive for CD44, while proliferating immature lymphocytes in germinal center were negative. 2. High grade and intermediate grade NHL samples were strongly positive for CD44, the positive rate in the former was higher than in the latter (P < 0.05). In the ten low grade lymphoma samples, only two were weakly positive and, the rest were negative. The expression of PCNA showed the same manner as CD44. 3. The microvessels in NHL increased with the histological malignant grade. CONCLUSION: The expression of CD44 in normal lymphatic tissues is different from that in NHL. Combination of CD44 expression with PCNA expression and microvessel density seems a reliable parameter for judging malignant degree of NHL. PMID- 11263340 TI - [Clinical analysis of 164 cases Coombs test positive autoimmune hemolytic anemia]. AB - OBJECTIVE: To analyze the clinical features and treatment outcomes of autoimmune hemolytic anemia (AIHA). METHODS: Coombs test positive AIHA patients were retrospectively analyzed. RESULTS: Patients with AIHA were predominantly female, secondary AIHA accounted for 49% of the total cases, mainly associated with connective tissue diseases. In most cases of AIHA, the anti-red cell antibodies were IgG plus C3, while patients with IgG plus IgM had more severe clinical manifestations. The response rate of adrenocortical hormones and cyclosporin A (CsA) was 90.9%. CONCLUSION: Adrenocorticosteroids is still the first choice of therapy for AIHA, CsA can improve the response rate. PMID- 11263341 TI - [Study on gene expression of TGF beta 1 and its receptor in leukemia cells and the serum TGF beta 1 level in the patients with acute leukemia]. AB - OBJECTIVE: To investigate the mechanism by which TGF beta 1 inhibits the growth of leukemic cells and the significance of serum TGF beta 1 level in acute leukemia(AL) patients. METHODS: RT-PCR was used to detect gene expression of cytokines including IL-1, IL-3, IL-6, TGF beta 1 and their receptors (R) in leukemic and normal cells. Leukemic colony assay was carried out to determine the effect of TGF beta 1 on the growth of leukemic cells and the serum TGF beta 1 level in 33 AL patients were measured by ELISA. RESULTS: TGF beta 1 was found to significantly inhibit the proliferation of leukemia cells of HL-60, K562 and DAMI cell lines. Furthermore, a variety of leukemia cell lines including HEL, HL-60, K562, U937, DAMI, MEG-01, HUT78 and CA were found to express mRNAs for TGF beta 1 and its receptor. In addition, TGF beta 1 was found to be able to evidently down regulate the expression of mRNAs for IL-6, IL-6R, IL-3, GM-CSFR in DAMI cells and, interestingly, up-regulate TGF beta 1 gene expression in the cells per se before the cells showed DNA fragmentation, a molecular hallmark for cell apoptosis. It was shown that serum TGF beta 1 levels were significantly decreased in leukemic patients, restored to normal in the patients achieved complete remission, and tended to decrease again in the recurrent patients. CONCLUSION: 1. TGF beta 1 may act as an inhibitory autocrine factor in the proliferation of leukemia cells; 2. the mechanism by which TGF beta 1 inhibits the growth of leukemia cells involves its down-regulating expression of positive cytokines and receptors and up-regulating TGF beta 1 expression in leukemia cells per se; 3. serum TGF beta 1 is a valuable parameter in monitoring the prognosis of leukemia and weakened control of TGF beta 1 on leukemia cells may play an important role in the pathogenesis of AL. PMID- 11263342 TI - [TNF-alpha production and antitumor effect of rhIL-2-activated bone marrow cells]. AB - OBJECTIVE: To explore the mechanism of the cytotoxic effects of bone marrow cells activated with recombinant human interleukin-2(rhIL-2). METHODS: The cytotoxicity of rhIL-2-activated bone marrow (ABM) and the TNF-alpha level in the supernant of ABM culture were measured after activated for 24 and 48 hours. RESULTS: The cytotoxic activity against HL-60 cells and the level of TNF-alpha were significantly increased in ABM as compared with that in non-ABM(P < 0.01). Meanwhile, there was a significantly positive relationship between the TNF-alpha levels in supernant and cytotoxicities of ABM. CONCLUSION: TNF-alpha produced by rhIL-2 activated bone marrow cells involved in the antitumor effect of ABM. PMID- 11263343 TI - [Effect of the IL-6 transfected bone marrow stromal cells on the recovery of hematopoietic functions in bone marrow transplanted mice]. AB - OBJECTIVE: To study the effect of the IL-6 transfected bone marrow stromal cell, QXMSC1 IL-6, on the hematopoietic reconstitution in bone marrow transplanted mice. METHODS: Bone marrow transplantation model was established by infusing hematopoietic cells combined with QXMSC1 IL-6 into syngeneic recipient mice through the tail vein. CFU-S, CFU-GM, CFU-E, BFU-E, and peripheral blood pictures were assayed. RESULTS: QXMSC1 IL-6 could increase the CFU-S, CFU-GM, CFU-E and BFU-E yields in BMT mice, and accelerate the recovery of peripheral blood cells. CONCLUSION: QXMSC1 IL-6 can improve the hematopoietic reconstitution in bone marrow transplanted mice. PMID- 11263344 TI - [Expression of thrombopoietin related genes in patients with idiopathic thrombocytopenic purpura]. AB - OBJECTIVE: To explore the expression of thrombopoietin (Tpo) and its receptor c mpl in bone marrow from patients with idiopathic thrombocytopenic purpura(ITP). METHODS: Non-radioactive in situ hybridization was used to detect tpo, c-mpl mRNA. Immune cytochemistry was used to detect MPL on megakaryocyte. RESULTS: The expression of tpo, c-mpl mRNA in bone marrow and MPL on megakaryocyte in patients with ITP was increased. CONCLUSION: c-mpl mRNA was regulated at transcriptional level. A negative feedback loop was involved in the regulation of megakaryopoiesis in bone marrow. MPL may be involved in the regulation of circulating Tpo. PMID- 11263345 TI - [Study of the etiologic pathogenesis of multiple myeloma]. PMID- 11263346 TI - [Will chronic nonproductive cough with bronchial hyperresponsiveness be cough variant asthma?]. AB - OBJECTIVE: The clinical characteristics of the patients suffered from chronic nonproductive cough with bronchial hyperresponsiveness were analyzed, so as to evaluate the value of measurement of bronchial responsiveness in diagnosis of cough variant asthma (CVA). METHOD: 124 patients with chronic nonproductive cough were divided into two groups after the measurement of bronchial responsiveness: cough bronchial hyperresponsiveness positive group (CBH-P, n = 35) and cough bronchial hyperresponsiveness negative group (CBH-N, n = 33). Routine pulmonary function, positive rate of antigen-skin pitting lest, count of blood eosinophil cells, level of blood IgE, positive rate of prednisone test and the number of development of classical asthma in two-year following up were studied. RESULT: The percentage of FEV1.0 in group CBH-P was significantly lower than in group CBH N, but antigen-skin pitting test, count of blood eosinophil cells, prednisone test and the number of development of classic asthma in two-year following up were all higher than in the latter group. In group CBH-P, there was no significant difference in positive rate of antigen-skin pitting test, routine pulmonary function, reacting threshold(Dmin) and wheezing threshold (DCW/Dmin) between the patients who developed classic asthma and those who did not. CONCLUSION: The measurement of bronchial responsiveness is important for the diagnosis of CVA, but it is not the only diagnostic criteria and should be combined with other clinical data. PMID- 11263347 TI - [Study on the pathogenesis of free radical in nocturnal asthma]. AB - OBJECTIVE: To examine the relationship between the pathogenesis of nocturnal asthma and free radicals. METHOD: The changes of the chemiluminescence of lymphocytes (Ly-cl) and polymorphonuclear leukocytes (PMN-cl) were studied by the cell chemiluminescence method. Glutathione peroxidase (GSH-Px) and malondialdehyde (MDA) levels also were determined respectively. RESULT: The Ly-cl and PMN-cl in patients with nocturnal asthma on the second day 2:00 am were significantly higher than that of the second day 2:00 pm (P < 0.01) and higher than that of normal group (P < 0.01). The GSH-px were decreased and MDA were increased, and there were significant differences compared with normal group (P < 0.001, P < 0.01 respectively). CONCLUSION: There is an obvious difference on metabolism of free radicals on day and night in nocturnal asthma. The results support that the free radicals have an important role in pathogenesis of nocturnal asthma. PMID- 11263349 TI - [Effect of adrenomedullin on bronchoconstriction responses induced by histamine]. AB - OBJECTIVE: To observe the effect of synthetic human adrenomedullin (AM) on anesthetized guinea pigs bronchoconstriction induced by histamine. METHOD: The guinea pigs were divided into saline group; isoproterenol groups at the dosages of 0.03 microgram/kg, 0.3 microgram/kg, 3 micrograms/kg and 30 micrograms/kg; adrenomedullin groups at the dosages of 0.02 microgram/kg, 0.2 microgram/kg, 2 micrograms/kg and 20 micrograms/kg. There were seven guinea pigs a group. Guinea pigs were anesthetized by sodium pentobarbital, normal saline group, isoproterenol and adrenomedullin at different dose i.v., while histamine (10(-4) mol/L) were vasolated and delivered into the airway. Body plethysmogroph was used to measure airway resistance and lung compliance. RESULT: The increment of airway resistance and decrement of lung compliance induced by histamine were inhibited significantly by isoproterenol at the dosages of 0.3 microgram/kg, 3 micrograms/kg and 30 micrograms/kg (P < 0.05). The increase of airway resistance was inhibited significantly by adrenomedullin at the dosages of 2 micrograms/kg and 20 micrograms/kg (P < 0.05). The decrease of lung compliance induced by histamine was inhibited significantly by adrenomedullin at 0.2 microgram/kg, 2 micrograms/kg and 20 micrograms/kg (P < 0.05). CONCLUSION: Adrenomedullin inhibited the bronchoconstriction induced by histamine significantly. PMID- 11263348 TI - [A model for exercise-induced asthma in guinea pigs with lipopolysaccharide and metyrapone]. AB - OBJECTIVE: To investigate the relationship between airway inflammation and exercise-induced asthma (EIA), an experimental model for EIA in guinea pigs was developed. METHOD: 27 guinea pigs was divided into 4 groups. A) LPS + MET exercise group (n = 7) had been pretreated with lipopolysaccharide (LPS 1 mg/kg, i.p.) and metyrapone (MET 50 mg/kg, i.p.) on the first day. Then lung resistance (RL) and dynamic compliance (Cdyn) were measured before and after exercise on the fourth day. B) LPS + MET non exercise group (n = 6) that pretreated with LPS and MET had nonexercise on the 4th day. C) NS exercise group (n = 7) pretreated with normal saline (NS 1 mg/kg, i.p.) on the 1st day. RL and Cdyn were measured before and after exercise on the 4th day. D) NS nonexercise group (n = 7) that pretreated with NS had no exercise on the 4th day. RESULT: In LPS+MET exercise group RL increased and Cdyn reduced significantly after exercise. In another 3 control groups there were no such changes either in RL or in Cdyn. CONCLUSION: These observations suggest that the treatment with LPS and MET could make a model for exercise-induced asthma. PMID- 11263350 TI - [The experimental and clinical study of heliox in treating bronchial asthma]. AB - OBJECTIVE: To evaluate the effects of breathing helium-oxygen(heliox: 79% He-21% O2) on bronchial asthma by way of experimental and clinical observations. METHOD: Six mongrel dogs were randomly inspired air or heliox after dripping 2% methacholine in its' tracheas controlled by 900C ventilator. During the experimental course, respiratory mechanical limits, hemodynamic index and blood gas values were observed. Eight asthmatic patients inspired heliox with assisted respirator of recycle-used helium, and the data of pulmonary function tests, arterial blood gas values of patients were recorded. RESULTS: The results demonstrated that heliox could reduce airway resistance, airway pressure and work of breathing, improve lung effective compliance, but had no effects on pulmonary circulation and systemic circulation. Heliox could ameliorate oxygen diffusion and CO2 eliminating, but the duration was very short after stopping breathing heliox. The assisted respirator of recycle-used helium could save helium distinctively, and the clinical application demonstrated that the instrument may be useful for pulmonary obstructive diseases. CONCLUSION: Heliox can reduce airway pressure, ameliorate oxygen diffusion and CO2 eliminating, so breathing heliox might be one of effective ways for asthmatic patients. PMID- 11263351 TI - [Polymerase chain reaction associated with dig-labelled DNA hybridization of mip gene to identify new isolated Legionella pneumophila strains]. AB - OBJECTIVE: By testing mip gene, one highly conservative virulence gene of Legionella pneumophila (Lp), as targets of polymerase chain reaction (PCR) and Digoxin-labelled probe hybridization to establish a rapid, specific and sensitive gene analysis method that can identify new isolated suspected bacteria strains as Lp. METHOD: Abstracting all the CDC reference Lp strains' and none-Lp Legionella strains' DNA, as well as the domestic isolated Lp strains' and some non Legionella control bacteria's DNA as templates, then were processed. All the positive PCR products(if no PCR amplicons were available, the original whole DNA should be detected) were hybridized with Digoxingen-labelled mip gene probe in dot-blot procedure. RESULT: All the tested Lp strains had positive PCR and hybridization results, at the same time, all none-Legionella bacteria and none-Lp Legionella strains got negative results. 6 of 26 isolates were identified as Lp strains by this method. CONCLUSION: Such a Lp strain identification procedure shows high specificity and sensitivity (nearly 100%, in this study), and mean while can be completed in a relative short time. Surely, this method can be largely available and has a potential value in diagnosis of clinical Legionellosis cases and pursuing the pathogen during Legionnaires' disease outbreaks. PMID- 11263352 TI - [The effect of c-myc antisense oligodeoxynucleotides on proliferation of pulmonary arterial smooth muscle cells stimulated by hypoxic endothelial cells conditioned medium]. AB - OBJECTIVE: To study the effect of c-myc antisense oligodeoxynucleotides on proliferation of pulmonary arterial smooth muscle cells (SMC) stimulated by hypoxic endothelial cells conditioned medium (HECCM). METHOD: After pulmonary arterial SMC were stimulated by HECCM, Northern blot technique was used for detection of expression of c-myc, and 3H-TdR incorporation and cell growth assay for detection of proliferation of SMC. RESULT: HECCM promoted the expression of c myc and proliferation of SMC significantly. Antisense-ODNs significantly inhibited the expression of c-myc and proliferation of SMC stimulated by HECCM, whereas sense ODNs didn't affect the expression of c-myc and proliferation of SMC. CONCLUSION: HECCM promote the proliferation of SMC by increasing the expression of c-myc, antisense ODNs inhibits the proliferation of SMC by downregulating the expression of c-myc gene. PMID- 11263353 TI - [A molecular and epidemiology study in patients with ventilator-associated pneumonia]. AB - OBJECTIVE: To evaluate causative agents in patients with ventilator-associated (VA) pneumonia epidemiologically, then to provide useful suggestion for diagnosis and treatment of VA pneumonia. METHOD: Prospectively, a protected specimens brush was used to obtain the secretion of lower respiratory tract of 65 patients, who had been receiving mechanical ventilation or tracheostomy for more than 72 hours. At the same time, other samples were collected from the relevant places, including pharyngeal and gastric juice of patients as well as other persons, ward's air. The secretion obtained were cultured with a quantitative method. Then all bacteria isolated were studied with the analysis of pattern of plasmids and chromosomal restriction endonuclease. RESULT: It was showed that the route of infection of the Gram-negative bacilli in VA pneumonia (19 cases) was intrinsic, a retrograde colonization from patient's stomach, that was the pattern of clonization from stomach to pharynx, then into lower respiratory tract, and the Gram-positive staphylococcus spread mainly through the ward's air, then directly into lower respiratory tract, or extrinsic (20 cases). CONCLUSION: The Gram positive staphylococcus is also major disease-producing germs in VA pneumonia, and the infection routes of G+ and G- bacteria might be different. PMID- 11263354 TI - [Expression and prognostic relation of cathepsin D in non-small cell lung cancer tissues and lymph nodes]. AB - OBJECTIVE: To study the expression and prognostic significance of cathepsin D(CD) in the patients with lung cancer. METHOD: A rapid immunohistochemical method (streptavidin-peroxidase conjugated method, SP) was used to detect CD expression of the NSCLC patients in 64 cases lung cancer tissues and paired lymph nodes samples. RESULT: The results showed positive expression rate of CD was 66% (42/64) in the lung cancer tissues, of which squamous cell cancer was 57% (17/30), adenocarcinoma was 74%(25/34). In the metastatic lymph nodes, the expression of CD was 57%(20/35). Expression of CD was higher in the patients with stage III-IV than in that with stage I-II (P < 0.05), of which was markedly higher in adenocarcinoma with lymph nodes metastasis than in which lymph nodes negative. CONCLUSION: Higher expression of CD is significantly related to the stage of the disease and lymph node metastasis in the adenocarcinoma. The expression of CD may be a good marker of prognosis in NSCLC. PMID- 11263355 TI - [The morphological and functional changes of pulmonary intravascular macrophages induced by LPS]. AB - OBJECTIVE: To study the roles of pulmonary intravascular macrophages (PIMs) on infective acute lung injury (ALI). METHOD: Porcine pulmonary blood vessels were flushed by modified Morton's method, PIMs were isolated by adhesion method and identified with the features at both light and electron microscopic level. The activity of IL-1 beta, and content of IL-6, IL-8 and TNF alpha in the culture supernatants were measured by thymocyte proliferation or ELISA, respectively. RESULT: Enlarged and increased pseudopods, and increased amount of lysosomes and phagosomes were found in the PIM stimulated with lipopolysaccharide (LPS, 10 micrograms/ml); the releases of TNF alpha, IL-1 beta, IL-6 and IL-8 were increased significantly as compared with the level of pre-stimulation of LPS (P < 0.01), and reaching their peaks at 1 h, 2 h, 4 h and 6 h after LPS stimulation, respectively. CONCLUSION: The isolation of porcine PIMs can be completed with modified Morton's method; the phagocytosis and secretion of PIMs are more active after LPS stimulation. In the pathogenesis of ALI, TNF alpha and IL-1 beta may play an important role at its early stage; however, the IL-6 and IL-8 may be associated with the pathophysiological changes at the later stage of ALI. PMID- 11263356 TI - [Analysis and application of polypeptide antigens of 13 mycobacterium strains]. AB - OBJECTIVE: To analyze and compare polypeptide antigens of 13 Mycobacterium strains and try to find out tuberculosis associated antigens. METHOD: Polypeptide antigens were analyzed by using SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblots techniques. Specific antibodies in serum or CSF samples obtained from 575 tuberculosis patients, 450 healthy individuals and 250 non-tuberculosis patients were detected by using immunoblots technique with purified specific antigen. RESULTS: The results showed that polypeptide of H37Rv, H37Ra, BCG, Bovis strains had the similar polypeptide bands and the similar immunogenicity in major polypeptide antigens. Molecular weight of major polypeptide antigens were 76,000, 65,000, 34,000-40,000, 28,000 and 25,000. The 76,000, 65,000 and 25,000 polypeptides were found in all the 13 Mycobacterium strains and they were common antigens probably. Purified specific antigen was used in serodiagnosis of tuberculosis and the sensitivity was 89.7%, while specificity was 95.7%. CONCLUSION: Polypeptide bands of Mycobacterium were found different and Mycobacterium strains could be identified by SDS-PAGE technique. The results suggested that specific antigen of H37Rv strain for antibody detection was a useful supplementary tool for diagnosis of extrapulmonary tuberculosis and bacteria-negative pulmonary tuberculosis. PMID- 11263357 TI - [Relationship between catalase activity, KatG gene and isoniazid-resistance in M. tuberculosis]. AB - OBJECTIVE: To investigate the relationship between catalase activity, KatG gene, gene mutation and INH-resistance in M. tuberculosis. METHOD: Catalase activities in 58 M. tuberculosis isolates were tested, and KatG gene mutations were detected further by PCR-SSCP. RESULT: None of INH-sensitive isolates lacked KatG sequences and all expressed catalase. 8(25%) INH-resistant isolates did not express catalase and 3(10%) lacked KatG gene, most of which were strains of high levels of resistance (MIC > 50 micrograms/ml). 8 INH-resistant isolates were analyzed further by PCR-SSCP and all were found to have KatG gene mutation. CONCLUSION: These findings indicated that lacking of catalase activity and KatG gene deletion occurred mainly in those highly INH-resistant strains, gene mutation other than complete deletion of KatG gene may be the major mechanism of INH-resistance. PMID- 11263358 TI - [Detection of Mycobacterium tuberculosis DNA in peripheral blood mononuclear cells from patients with pulmonary tuberculosis by polymerase chain reaction technique]. AB - OBJECTIVE: To evaluate the clinical value of detection of Mycobacterium tuberculosis DNA (MTB-DNA) in peripheral blood mononuclear cells (PBMC) for diagnosis of pulmonary tuberculosis. METHOD: Polymerase chain reaction (PCR) technique was used to amplify gene of 240 bp DNA fragment prepared by Triton X 100 method, and some factors affecting PCR result were also analysed. RESULT: In blood samples of 89 patients with pulmonary tuberculosis and in sputum samples of 84 patients with pulmonary tuberculosis, the positive rates of PCR were 73% and 57% respectively, and the total positive rate of combinative detection of blood and sputum samples was 87% in 84 pulmonary tuberculosis patients. In 30 non tuberculosis patients, 3 showed MTB-DNA positive. CONCLUSION: PCR technique prepared by Triton X-100 method is rapid, simple, specific, and sensitive for detection of MTB-DNA in PBMC, and its sensitivity and accuracy could be increased in combination with sputum MTB examination. Detection of MTB-DNA in PMBC is of value in diagnosis of pulmonary tuberculosis. PMID- 11263359 TI - [Analysis of reasons failed focal debridement in 109 patients with spinal tuberculosis]. AB - OBJECTIVE: To explore reasons failed focal debridement in treating spinal tuberculosis patients and methods to cope with them. METHOD: One hundred nine patients with spinal tuberculosis who failed initial focal debridement from 1987 to 1996 were reviewed. RESULT: The reasons of operational failure were found to be the use of unreasonable regimens and existence of drug resistant strains; inappropriate operational indication, approach and treatment during operation; lesions involving several vertebrae, skipping destruction and complicating with other kinds of extrapulmonary tuberculosis. CONCLUSION: The key to successful treatment of the disease is to choose appropriate operative indication and approach, discard spinal cord pressure and rebuild spinal stability on the basis of reasonable chemotherapy regimen. PMID- 11263360 TI - [Primary prevention of bronchial asthma]. PMID- 11263361 TI - [Tumor suppressor gene CDKN2 and lung cancers]. PMID- 11263362 TI - A glimpse of hepato-biliary-pancreatic surgery in China. PMID- 11263363 TI - Bletilla striata as a vascular embolizing agent in interventional treatment of primary hepatic carcinoma. AB - OBJECTIVE: To evaluate Bletilla striata as a vascular embolizing agent in interventional treatment of primary hepatic carcinoma. METHODS: We made a vascular embolizing agent from a Chinese medicinal herb, Bletilla striata (bai ji). From October 1991 to January 1995, 56 patients with primary hepatic carcinoma were treated by hepatic arterial chemoembolization with Bletilla striata angioembolus, and 50 patients with primary hepatic carcinoma who were treated by conventional gelfoam embolization served as controls. All patients were followed up for 10-48 months. RESULTS: Embolization with the Bletilla striata powder produced extensive and permanent vascular obstruction, resulting in marked shrinkage of the tumor and significant decrease of serum alpha fetoprotein (AFP) level. After embolization, less collateral circulation formed later. The treatment interval was prolonged to an average of 7 months. The 1-, 2- and 3-year survival rates were 81.9%, 44.9%, and 33.6%, respectively, with a mean survival period of 19.8 months. All the clinical parameters were better than those in the control group treated by conventional gelfoam embolization. CONCLUSION: Bletilla striata angioembolus is a good vascular embolizing agent for treating PHC. PMID- 11263364 TI - Mechanism of overproduction of plasma prostacyclin in portal hypertensive rats. AB - OBJECTIVE: To evaluate the role of increased portal pressure and portosystemic shunting in elevated level of prostacyclin (PGI2) in portal hypertension. METHODS: Thirty-six male Sprague-Dawley rats were divided into four groups: prehepatic portal hypertension (PHPH, 8 rats), intrahepatic portal hypertension (IHPH, 9), end-to-side portacaval shunt (PCS, 8), and sham-operated controls (SO, 11). Two weeks after surgery, free portal pressure (FPP) was measured; systemic and splanchnic hemodynamics was studied by radioactive microsphere technique and blood sample from the femoral artery was obtained to measure the level of plasma 6-keto-PGF1 alpha with radioimmunoassay. RESULTS: The FPP (mmHg) in IHPH, PHPH, PCS and SO rats was 13.10 +/- 1.02, 12.10 +/- 1.52, 3.0 +/- 0.82 and 6.86 +/- 0.69, respectively. The value of FPP was significantly increased in IHPH, PHPH rats and significantly decreased in PCS rats when compared to SO rats. Cardiac index (CI) and portal venous inflow (PVI) were in the order of PCS > PHPH > IHPH > SO rats. Portosystemic shunting (PSS) in PCS, PHPH, IHPH was 99.7 +/- 0.29%, 76.02 +/- 20.62% and 30.34 +/- 10.18%, respectively. The concentrations of plasma 6-keto-PGF1 alpha (ng/ml) in PHPH, IHPH, PCS and SO rats were 6.93 +/- 2.43, 5.09 +/- 2.27, 2.36 +/- 1.01 and 1.56 +/- 0.61, respectively. The concentrations of plasma PGI2 in PHPH, IHPH and PCS rats were significantly higher than those in SO rats. Furthermore, the concentrations of plasma PGI2 in PHPH and IHPH rats were also significantly higher than those in PCS rats. Moreover, a closed positive correlation existed between plasma PGI2 and FPP (r = 0.67, P < 0.001). CONCLUSIONS: The results of the present study suggest that the elevated PGI2 in portal hypertension is mainly due to the overproduction of PGI2 in vascular epithelium cells induced by increased portal pressure, whereas portosystemic shunting and liver dysfunction play a secondary role. In addition, the results of this study do not support that PGI2 mediated the hyperhemodynamics in portal hypertension. PMID- 11263365 TI - Detection of hepatitis C virus RNA sequences in hepatic portal cholangiocarcinoma tissue by reverse transcription polymerase chain reaction. AB - OBJECTIVE: To detect hepatitis C virus (HCV) RNA sequences in the hepatic portal cholangiocarcinoma tissues and their relationship. METHODS: RNA was extracted from paraffin-embedded cholangiocarcinoma tissues of 6 patients by guanidinium method, subjected to reverse transcription and then amplified by double PCR technique using nested primers from the highly conserved 5' noncoding region of HCV genome. RESULTS: HCV RNA of 5' NT sequences was found in the hepatic portal cholangiocarcinoma tissues of 5 out of 6 (83%) patients. CONCLUSIONS: HCV RNA sequences present with high infectious rate in cholangiocarcinoma, and reverse transcription polymerase chain reaction (RT-PCR) assay using primers derived from 5' NT region of HCV sequence is most useful in detecting HCV infection. The development of cholangiocarcinoma awaits further studies. PMID- 11263366 TI - Influence of octapeptide of cholecystokinin, vasoactive intestinal peptide and substance P on dynamics of biliary system and cardiovascular system. AB - OBJECTIVE: To examine the effects of cholecystokinin octapeptide (CCK-OP), vasoactive intestinal peptide (VIP) and substance P (SP) on the dynamics of the biliary system and cardiovascular system, and the relationship between the dynamics of the biliary system and cardiodynamics. METHODS: In 91 anesthetized guinea pigs, a triple-lumen, side-hole perfusion catheter (1.0 mmOD) was inserted through the duodenal papilla into the common bile duct (CBD) and the sphincter of Oddi (OS). An end-hole PE-50 catheter was inserted into the left ventricle of heart through the left jugular artery. The left ventricle of heart motility, OS motility and CBD pressure were recorded during the intravenous administration of CCK-OP, VIP, SP and the combination of CCK-OP and VIP. RESULTS: Intravenous CCK OP increased the fasted OS motility index (MI), decreased the basal pressure in OS, increased CBD pressure, and inhibited the motility of the left ventricle of heart. VIP alone showed no significant effect on the biliary system and cardiovascular system, but when infused together with CCK-OP, it inhibited the effects of CCK-OP on both systems. Exogenous SP acted like CCK-OP on both biliary system and cardiovascular system, but less potently. CONCLUSIONS: The gastrointestinal peptides have important effects on both biliary system and cardiovascular system. There is an important negative correlation between CBD pressure and the motility of the left ventricle of the heart during the infusion of peptides. PMID- 11263367 TI - Effects of proglumide, a gastrin receptor antagonist, on human large intestine carcinoma SW480 cell line. AB - OBJECTIVE: To explore the effects of proglumide, a gastrin receptor antagonist, on the amount of viable cells, synthesis of DNA and protein, and cell proliferation cycle in human large intestine carcinoma SW480 cell line in order to provide experimental basis for treatment of large intestine carcinoma using proglumide. METHODS: Large intestine carcinoma SW480 cells at logarithmic growth stage were cultivated with different concentrations of proglumide for different periods of time, then the amount of viable cells was determined by MTT colorimetric analysis. The SW480 cells were cultivated with proglumide, pentagastrin, proglumide + pentagastrin for the same period of time, then the contents of DNA and protein and the cell proliferation cycle were determined by flow-cytometry. RESULTS: The amount of viable cells, synthesis of DNA and protein, distribution of cell cycle, and proliferation index (PI) in the proglumide group did not differ significantly from those in the pentagastrin group (P > 0.05). The amount of viable cells in the pentagastrin group was significantly higher than that in the pentagastrin group (P < 0.01). In the proglumide + pentagastrin group the amount of viable cells, synthesis of DNA and protein, amount of S and G2M phase cells, and PI were all significantly lower than those in the pentagastrin group (all P < 0.01), and the amount of G0/G1 phase cells was significantly higher than that in the pentagastrin group (P < 0.01), but none of the above differed from those in the control group (all P > 0.05). CONCLUSIONS: Proglumide has no obvious effect on the growth of human large intestine carcinoma SW480 cell line, but can inhibit the growth-promoting effect of pentagastrin on large intestine carcinoma cells. The mechanism may be that proglumide inhibits the promoting effect of pentagastrin on the synthesis of DNA and protein of carcinoma cells, and then inhibits carcinoma cell growth from G0/G1 phase to S and G2M phase. PMID- 11263368 TI - Experimental study on liver microcirculation disturbance following transplantation and the protective effect of prostaglandin E1 in the rat. AB - OBJECTIVE: To determine the effect of PGE1 on liver microcirculation disturbance following orthotopic liver transplantation in rats. METHODS: Forty male adult Wistar rats were divided randomly into 3 groups. Eight transplantations were established in both the experimental and control group, while in the sham group, the liver was dissected like in the experimental group, but no resection was performed. In the experimental group, PGE1(0.5 microgram/kg.min-1) was injected intravenously into the donor before the operation, and added (1 mg/L) to the flush and preservation fluid, while PGE1 was replaced by normal saline in the control group. Confocal laser scan microscopy, biochemical test, and optical and electronic microscopy were used. RESULTS: In the control group the reperfusion state was poor, leukocyte infiltration appeared in the center of lobule, and transaminase rose after transplantation. In the experimental group distinctive improvement was seen as compared with the control group (P < 0.05). Histological findings showed progressive degeneration and necrosis following transplantation in the control group, while in the experimental group the histological changes were improved to some degree by the use of PGE1. CONCLUSIONS: In liver transplantation, ischemic reperfusion damage may lead to hepatic microcirculation disturbance, which is the major cause of graft failure. Infusing PGE1 into the donor intravenously before ischemia and adding PGE1 to the cold storage fluid could improve hepatic microcirculation, and thus reducing ischemic reperfusion damage in liver transplantation. PMID- 11263369 TI - Androgen receptor in primary hepatocellular carcinoma and its clinical significance. AB - OBJECTIVE: To assess retrospectively the clinical value of androgen receptor (AR) levels in primary hepatocellular carcinoma (HCC) as a prognostic factor of the disease. METHODS: Fresh HCC tissue and the surrounding liver tissue were obtained surgically from 32 patients with HCC, and preserved in liquid nitrogen. The levels of AR in all specimens were determined by radio-ligand binding assay. RESULTS: The median level of AR was 42.8 fmol/mg protein in the cancerous tissue and 48.3 fmol/mg protein in the surrounding non-cancerous liver tissue. The overall survival rate of the patients with AR < 30 fmol/mg protein in either HCC or the non-cancerous liver was significantly higher than that of the patients with AR > or = 30 fmol/mg protein (P < 0.05 and P < 0.01, respectively). The relative risk on prognosis was 3.27 (P < 0.01) for AR level in HCC and 6.06 (P < 0.001) for AR level in the non-cancerous tissue. The main prognostic factors except the tumor size were not different between the group with higher AR level and that with lower AR level in HCC. The AR level in HCC had a positive correlation with the tumor size (r = 0.44, P < 0.05). CONCLUSIONS: AR can be detected in HCC and the AR status might be a prognostic parameter that provides additional predictive information on the survival. Different AR status might define a real difference of biological characteristics between HCCs. PMID- 11263371 TI - Tumor angiogenesis related to growth pattern and lymph node metastasis in early gastric cancer. AB - OBJECTIVE: To investigate the correlation between angiogenesis and tumor growth pattern as well as the lymph node metastasis to reveal the significance of vascularity in the early stage of gastric cancer. METHODS: 97 specimens from patients with early gastric cancer were studied by immunohistochemical method using anti-factor VIII related antigen antibody. RESULTS: Microvessel count was related to tumor growth pattern. The mean vessel count was higher in superficially spreading and penetrating types of tumors. Lymph node metastasis was correlated to microvessel count. Tumors with lymph node metastasis had higher microvessel counts than those without lymph node metastasis. CONCLUSIONS: In the early stage of gastric carcinoma, angiogenesis is correlated with tumor growth pattern and lymph node metastasis. Identification of tumors with high density of vascularization is beneficial for closer follow-up and adjuvant therapy. PMID- 11263370 TI - Pericardial devascularization combined with Latarjet's innervation in the treatment of patients with portal hypertension. AB - OBJECTIVE: To study the effect of pericardial devascularization combined with Latarjet's innervation on portal hypertension. METHODS: Forty-eight patients undergoing pericardial devascularization combined with Latarjet's innervation were compared with 57 patients with devascularization. Clinical results were evaluated by postoperative portal vein pressure, postoperative rebleeding, operative mortality, abdominal distension, sudden diarrhea, and gastric retention. RESULTS: The incidence of rebleeding, mortality, abdominal distension, sudden diarrhea, and gastric retention was 2%, 6.3%, 6.3%, 4.2% and 0% respectively in pericardial devascularization combined with Latarjet's innervation, and 12.5%, 12.3%, 24.6%, 15.6% and 14% respectively in devascularization. CONCLUSIONS: Pericardial devascularization combined with Latarjet's innervation preserves the normal function of gastric emptying. This method plays an important role in maintaining nutritional supply and digestive function. PMID- 11263372 TI - Horizontal platform supported total hip replacement. AB - OBJECTIVE: To investigate the effect of horizontal platform supported (HPS) prosthesis on cementless total hip replacement for surgical treatment of arthritis in a long-term follow-up study. METHODS: Clinical and radiographic follow-up of 65 consecutive primary cementless porous coated HPS total hip replacements, which were implanted in 60 patients between 1982 and 1989, was carried out. RESULTS: The 53 hips in 50 patients were evaluated both clinically and radiographically. The average follow-up was 6.8 years (range: 5 to 12 years). According to the Harris hip score, 52 (98%) of the hips had sustained an excellent or good result with an average score of 92. At the time of final assessment, no patient experienced anterior thigh pain. There were two revisions performed early in the series due to technical failures. Radiographically, osteolysis and bone remodelling were assessed. CONCLUSION: The findings suggest that the clinical and radiographic results after cementless total hip replacement for primary hip arthroplasty may be favourably influenced by the use of the proximal stress loaded HPS femoral component. PMID- 11263373 TI - Heterotopic ossification of the hip after spinal cord injury. AB - OBJECTIVE: To analyse the risk factors of heterotopic ossification after spinal cord injury. METHODS: Of 104 patients with spinal cord injury, 47 (45.2%) were complicated by heterotopic ossification of the hip. These patients with heterotopic ossification were compared with 57 patients without heterotopic ossification. RESULTS: Statistical analysis showed that complete paraplegia, spasticity and pressure sore were significantly (P < 0.05) or highly significantly (P < 0.01) related to heterotopic ossification formation. The coexistence of two or three of these factors in the paraplegic patients was found to significantly (P < 0.01) enhance the risk of heterotopic ossification formation. CONCLUSION: When the spinal cord injured patients have single or multiple risk factors, they should be predicted for susceptibility of heterotopic ossification. PMID- 11263375 TI - Pharmacokinetics of intravenous arsenic trioxide in the treatment of acute promyelocytic leukemia. AB - OBJECTIVE: To study the pharmacokinetics and metabolism of arsenic trioixide (As2O3) and its main side effects. METHOD: As2O3 was administered intravenously at the dose of 10 mg per day for the treatment of 8 relapsed acute promyelocytic leukemia (APL) patients. The arsenic content was measured by Gas-phase chromatography. RESULTS: The plasma maximal concentration (Cpmax) was 0.94 +/- 0.37 mg/L (x +/- s), time to peak concentration (Tp) was 4 hours, plasma distribution half-time (t1/2 alpha) and elimination half-time (t1/2 beta) were 0.89 +/- 0.29 hours and 12.13 +/- 3.31 hours, respectively. Apparent distribution volume (Vc) was 3.83 +/- 0.45 L, system clearance (CLs) was 1.43 +/- 0.17 L/h, and area under curve (AUC) was 7.25 +/- 0.97 mg.h/L. The continuous administration of As2O3 did not alter its pharmacokinetic behaviors. During As2O3 treatment, 24-hour arsenic content in urine accounted for 1%-8% of the daily dose (10 mg). When arsenic accumulation in hair and nail increased continuously, the peak concentration could be five to seven-fold higher than that of pre-treatment. Importantly, arsenic contents in both urine and hair or nail declined gradually after drug withdrawal. No bone marrow suppression or severe organ-impairment was found. CONCLUSION: As2O3 is a relatively safe and effective remedy in the treatment of patients with relapsed APL, in spite of certain degree of arsenic accumulation in some tissues. PMID- 11263374 TI - Inhibiting effect of murine double minute-2 oncogene on apoptosis induced by retroviral-mediated wild-type p53. AB - OBJECTIVE: To investigate the effects of wild-type p53 (wtp53) on inducing apoptosis by restoring wtp53 expression in pancreatic adenocarcinoma cell line (PC-2) which contains mutant p53, and the interaction between murine double minute-2 (MDM2) and wtp53 in pancreatic adenocarcinoma. METHODS: A recombinant retroviral vector expressing wild-type p53 was constructed and packaged by packaging cell line PA317 cells using calcium phosphate coprecipitation method. The supernatant of the packaged cells PA317 was used to transfect the pancreatic carcinoma cell line (PC-2), then a transformed cell line PC-2/swtp53 was established. The recombinant vector pCMV-MDM2 was transduced into PC-2/swtp53 cell line by lipofectin-mediated method, a double transfected cell line (PC 2/swtp53/pCMV-MDM2) was formed. To determine the integration and expression of exogenous wtp53 gene in the transfected cells, polymerase chain reaction (PCR), Western blot and immunoprecipitation analyses were performed. Apoptosis was analyzed by means of flow cytometry, in situ terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) analysis and DNA agarose gel electrophoresis. RESULTS: Transduction with the retroviral vector resulted in integration and expression of wtp53 gene in host cells. The apoptotic cells in PC 2/swtp53 and PC-2/swtp53/pCMV-neo cell lines were 12.1%-12.9%, while the double transfected cell line, PC-2/swtp53/pCMV-MDM2, showed less (3.2%) apoptotic cells than its parent cell lines. CONCLUSIONS: Restoration of expression of wild-type p53 with a retroviral vector can increase programmed cell death of pancreatic adenocarcinoma cells (PC-2) containing mutant p53. The overexpression of MDM2 protein has a negative regulating role in wtp53-induced apoptosis in PC-2 cell. PMID- 11263376 TI - Cotransduction of tyrosine hydroxylase and aromatic L-amino acid decarboxylase genes into cultured striatal cells using adeno-associated virus vectors. AB - OBJECTIVE: To examine whether tyrosine hydroxylase (TH) and aromatic L-amino acid decarboxylase (AADC) genes can be cotransduced into the same target striatal cells using adeno-associated virus (AAV) vectors, and to determine whether the cotransduction would result in better biochemical change than the TH gene alone. METHODS: TH and AADC genes were cotransduced into cultured striatal cells with separate AAV vectors. Expressions of TH and AADC were detected by immunocytochemistry; intracellular catecholamine levels were assayed by high performance liquid chromatography (HPLC). RESULTS: TH and AADC genes were efficiently cotransduced into the striatal cells. Specifically, the coexpression of TH and AADC resulted in more effective dopamine production compared with the TH gene alone. CONCLUSION: Using AAV vectors, coexpression of TH and AADC in the striatal cells might be a useful approach to gene therapy for Parkinson's disease. PMID- 11263377 TI - Single-strand conformation polymorphism for analysis of genomic variability of hepatitis C virus nonstructure 5A region. AB - OBJECTIVE: To establish a convenient method to detect the genomic population with hepatitis C virus (HCV) at nonstructure 5A (NS5A) region and to determine the correlation between the genomic population complexity at NS5A region and disease stage. METHODS: The sera from 52 patients with chronic hepatitis C virus infections were analysed using single strand conformation polymorphism (SSCP). In the SSCP, an asymmetric polymerase chain reaction (PCR) was carried out on the 455 bp products of the first round PCR at the NS5A region and the number of band of single strand deoxyribonucleic acid (DNA) which reacted with complemental DNA probe specific for the NS5A region after gel electrophoresis was analyzed. RESULTS: In 90% patients with chronic persistent hepatitis, the bands of single strand DNA was limited to one, and in those with chronic active hepatitis or liver cirrhosis, two or more bands of DNA were frequently detected. In about half of patients with hepatocellular-carcinoma, three or more bands were found. The number of bands increased with the progression of liver disease. The multivariate analysis showed that the progression of liver disease was the independent factor of viral diversity (P < 0.025) and was not related to the age, sex, the route of infection and the titer of hepatitis C virus ribonucleic acid (HCV RNA). CONCLUSION: These results suggest that the genomic variability of HCV at NS5A region increases with the progression of liver disease, and this may be closely related to the clinical features of type C liver disease. PMID- 11263378 TI - Atrial sensing and pacing with a single pass ventricular lead. AB - The use of single lead for atrial synchronous ventricular (VDD) pacing in patients with high grade atrioventricular (AV) block and normal sinus node function is an acceptable alternative to dual chamber (DDD) pacing. Implantation and follow up procedures are simplified, and cost is usually reduced by more than the cost of an additional atrial lead. With the use of either diagonally arranged dipole or closely spaced ring electrodes, reliable atrial sensing can be achieved using differential atrial amplifier and high atrial sensitivity. Also oversensing is infrequently observed using provocation tests and dynamic recordings, clinical undersensing is unusual and minimized by programming to the highest atrial sensitivity. However, as atrial pacing is not possible, loss of AV synchrony and rate response may occur for unrecognized or progressive sinus node disease and lower rate limit. The development of single lead dual chamber pacing system may overcome this limitation. Recent studies have demonstrated that atrial pacing can be effective either with the use of a special pacing lead configuration or via floating atrial electrode with a novel stimulation method. Overlapping biphasic impulse (OLBI) can reduce atrial pacing threshold. Early clinical experience suggested that this new pacing method can provide effective and reliable atrial pacing with a relatively low incidence of diaphragmatic pacing. Thus the problem of atrial sensing is solved with a single pass lead but further long term evaluation is required to assess the efficacy and feasibility of new instrumentation for single lead dual chamber pacing. PMID- 11263379 TI - A symptomatic Rathke's cleft cyst: preoperative diagnosis. PMID- 11263380 TI - Primary omental torsion in a child: mimic acute appendicitis. PMID- 11263381 TI - [Evaluation of accuracy of Healthdyne Nightwatch to monitor sleep breathing disorder]. AB - OBJECTIVE: To study the advantage of computerized portable polysomnograph (Healthdyne Night Watch, NW) compared to standard paper scoring polysomnograph (PSG). METHOD: NW was used to monitor the same patient with sleep apnea syndrome at the same night and compared with standard PSG. RESULT: Apnea and hypopnea index (AHI) recorded by PSG was 28.6 +/- 6.9 versus 28.7 +/- 7.0 recorded by NW. Mean lowest SaO2 recorded by PSG was 73.5% +/- 2.8% versus 74.5% +/- 3.2% recorded by NW. Using a PSG value of AHI > or = 5 and > or = 10 as the diagnostic criteria, the diagnostic agreement of NW was 93.7% and 95.3%, respectively. CONCLUSION: The computerized portable polysomnograph (NW) saves time and labour. It could be used in patient's home or ward and could replace PSG in most situations. PMID- 11263382 TI - [Clinical study of physiologic work of breathing as a weaning parameter in mechanically ventilated patients]. AB - OBJECTIVE: To evaluate the physiologic work of breathing (WOBphy) as a predicting Parameter of weaning and extubation. METHODS: WOBphy was obtained by the patient's work of breathing (WOBt), minuting the imposed work of breathing (WOBimp). In weaning trial, if the patients could not meet predefined extubation criteria, but WOBphy < 0.70 J/L, the patients were also extubated immediately. RESULTS: In a group of 28 patients who met the predefined criteria (group 1), 27 patients were successfully extubated. In another group of 13 patients who could not meet the criteria, but WOBphy < 0.70 J/L (group 2), extubation was successful in 12 patients. WOBt and WOBimp were significantly higher in group 2 (1.37 +/- 0.50 and 0.82 +/- 0.37 J/L) than group 1 (0.96 +/- 0.38 and 0.55 +/- 0.27 J/L, P < 0.05), while WOBphy was higher in group 2, but did not reach statistical significance. CONCLUSIONS: WOBphy < 0.70 J/L may be safe as the weaning and extubation criterion and enables earlier extubation. PMID- 11263383 TI - [Clinical significance of cytokine and eosinophil cationic protein concentrations in sputum of asthmatic patients]. AB - OBJECTIVE: To examine whether levels of inflammatory cytokines and eosinophil cationic protein (ECP) in the sputum reflect the severity of bronchial asthma. METHOD: We collected sputum expectorated spontaneously from 15 asthmatics with acute attacks of moderate to severe degree (MS group) and 10 subjects with acute attacks of mild asthma (M group). The interleukin (IL)-5(35 ng/L) tumor necrosis factor (TNF)-alpha(M 149 +/- 59 ng/L, MS 267 +/- 147 ng/L), soluble IL-2 receptor (sIL-2R) (M 348 +/- 107 kU/L, MS 488 +/- 127 kU/L) levels in the sputum were measured with enzyme-linked immunosorbent assay, and sputum ECP (M 127 +/- 95 micrograms/L, MS 278 +/- 150 micrograms/L) concentration were measured by Immuno CAP System. RESULT: Sputum IL-5, TNF-alpha, sIL-2R, ECP concentrations in moderate to severe patients were significantly higher than in mild subjects. CONCLUSION: These findings suggest that inflammatory cytokines and mediator levels are detectable in the sputum from asthmatics and they might participate in the exacerbation of asthma. PMID- 11263384 TI - [Study on endothelin-1 positive expression and quantitative analysis in lung cancer]. AB - OBJECTIVE: In order to study endothelin-1 (ET-1) positive expression in lung cancer and the relationship between the ET-1 quantitative analysis and the types, grades of lung cancer. METHOD: ET-1 positive expression and quantitative analysis were detected using avidin-biotin-peroxidase complex method and image analysis technology. RESULT: The ET-1 positive expression were mainly located in the cytoplasm in 104 cases with various types of lung cancer. The positive rate of ET 1 in adenocarcinoma, squamous-cell carcinoma and large-cell-lung cancer were 71.4%, 57.1% and 40.0% respectively. The positive rate of ET-1 in small-cell-lung cancer was 21.4%, significantly lower than others. The results of image analysis on adenocarcinoma and squamous-cell carcinoma showed that the lower lung cancer differentiation, the higher ET-1 quantitative. CONCLUSION: There is ET-1 positive expression in all of types of lung cancer, but there is a high ET-1 positive expression in adenocarcinoma and squamous-cell carcinoma. The results of image analysis indicated that ET-1 quantitive expression was somehow related to the differentiation degree of neoplasm, so ET-1 quantitative analysis may be as a monitoring index of adenocarcinoma and squamous-cell carcinoma growing. PMID- 11263385 TI - [Diagnosis and management of pulmonary arteriovenous fistula]. AB - OBJECTIVE: To evaluate the causes, characteristics of clinical manifestations and the complications of pulmonary arteriovenous fistula, to analyse the advantages and disadvantages of diagnostic methods, and to improve the efficacy of treatment. METHODS: Through reviewing reference articles and 22 cases. RESULTS: Arteriovenous fistulae in 12 of 22 cases were removed by pulmonary resection, of whom 9 were completely recovered, 1 recovered significantly, 1 got better in early stage but could not be follow-up 6 months later, 1 died one year after operation. Five out of the 22 cases were surgically intervened with other non pulmonary operations, of whom 4 recovered but the clinical appearances of PAVF were not obviously improved, 1 died shortly after operation. The others of 22 cases (5 cases) were not specially treated and not be follow-up. CONCLUSION: Echocardiography is the most simple, efficient method for diagnosis of PAVF. Pulmonary angiography can identify the size, location and profile of the PAVF. The results of removal of the fistulae are excellent, however the candidates should be selected strictly. PMID- 11263386 TI - [Further implementation and study of the strategy and measures of tuberculosis control]. PMID- 11263387 TI - [Comparative study on efficacy of regimens including streptomycin or ethambutol]. AB - OBJECTIVE: To evaluate the advantages and disadvantages of regimens of 2E3H3R3Z3/4H3R3(EMB regimen) and 2S3H3R3Z3/4H3R3(SM regimen) in tuberculosis control program. METHOD: Retrospective, cross-sectional and prospective studies were carried out in Shijiazhuang city, Hebei province from January 1994 to June 1996. RESULT: There was no significant difference between the two regimens in efficacy, relapse rate and full course supervision. The EMB regimen was found more applicable than the SM regimen, and the SM regimen caused more side effects than the EMB regimen. Streptomycin skin test had a 4.5% positive rate, and using SM costs 84% more than using EMB. One of the drawbacks found in the SM regimen was that only in 42.9% of the rural sanitation units the disinfection standard could be fulfilled, and the patients preferred the EMB regimen to the SM regimen. CONCLUSION: The EMB regimen is more applicable than the SM regimen in the tuberculosis control program. PMID- 11263388 TI - [Detection of mycobacterium tuberculosis by AmpliSensor-PCR technique and its clinical application]. AB - OBJECTIVE: To explore the value of Amplisensor-PCR method in diagnosis of tuberculosis. METHOD: The samples from 784 patients with tuberculosis, 160 patients with lung cancer were detected by using AmpliSensor-PCR, PCR, smear and culture methods, and the results were compared. RESULT: The sensitivity of AmpliSensor-PCR technique was obviously higher than that of smear and culture (P < 0.01). The specificity of AmpliSensor-PCR method was higher than that of PCR method. CONCLUSION: Amplisensor-PCR method can be determined to detect the lower limit by standard curve, through which the original amount of target-DNA in samples can be calculated. AmpliSensor-PCR method shows higher specificity and sensitivity. It is of clinical value in diagnosis of pulmonary tuberculosis and particularly of extrapulmonary tuberculosis. PMID- 11263389 TI - [Clinical significance of serum anti-mycobacterium tuberculosis antibody in diagnosis of tuberculosis]. AB - OBJECTIVE: To evaluate the significance of measuring serum antilipoarabinomannan IgG (LAM-IgG), an anti-Mycobacterium tuberculosis antibody, in diagnosis of tuberculosis. METHOD: Sensitivity and specificity of the antibody in diagnosis of tuberculosis were detected. Results of the antibody detection, sputum smear and PPD examination were compared, and the relationship between the level of the antibody and the situation of the disease was also observed. RESULT: The sensitivity of the antibody in diagnosis of tuberculosis was found to be 71.9%, while the specificity 91.9%. The consistent rates of the antibody with sputum smear and PPD test were 80.2% and 61.6% respectively. The level of the antibody was related with the situation of the disease. CONCLUSION: Detection of the antibody is helpful for diagnosing tuberculosis and can be used to evaluate the prognosis of the disease. PMID- 11263390 TI - [Analysis of lobar pneumonic tuberculosis]. AB - OBJECTIVE: To heighten the awareness of lobar pneumonic tuberculosis(or tuberculous pneumonia or acute pneumonic tuberculosis). METHOD: 10 cases with lobar pneumonic tuberculosis were reviewed. RESULT: All the patients showed acute onsets and 9 of them had a continuous high fever, and their WBC was not found higher than 10 x 10(9)/L. All of the patients' chest radiographs showed a consolidation in one or two lobars, and in 30% of the patients pleural effusions were found. Significant roent genographic changes could occur in short duration. All the patients were not sensitive to common antibiotics, and the shadow could enlarge in short time. Exudative foci could be absorbed shortly after antituberculosis chemotherapy, and no cavitation was found. CONCLUSION: An early transbroncho-lung biopsy (TBLB) and (or) brushing smear may be advantageous to early diagnosis and treatment of this illness. PMID- 11263391 TI - [Treatment of thoracolumbar tuberculosis complicated with paraplegia]. AB - OBJECTIVE: To explore characteristics of thoracolumbar tuberculosis complicated with paraplegia, relationship between efficacy and approach of operation, and to discuss indication and applicability of conservative treatment. METHOD: Forty eight cases with thoracolumbar tuberculosis complicated with paraplegia were classified into two groups. Thirty-two cases in the operational group were treated with extrapleural, extraperitoneal approach and transpleural, extraperitoneal approach. Sixteen cases in the nonoperational group were treated conservatively. RESULT: The recovered cases in the operational and the nonoperational groups respectively accounted for 75% and 88%. CONCLUSION: Thoracolumbar tuberculosis complicated with paraplegia can be cured by either operative or conservative treatment, and the efficacy of operation directly related with operational approach. PMID- 11263392 TI - [A study on enzymatic activities of bronchoalveolar lavage fluid in patients with interstitial lung diseases]. AB - OBJECTIVE: To evaluate the relationship between enzymatic activities in bronchoalveolar lavage fluid(BALF) and interstitial lung diseases(ILDs). METHOD: Cellular components and levels of superoxide dismutase (SOD), glutathione peroxidase(GSH-PX), angiotensin converting enzyme(ACE) and lactate dehydrogenase(LDH) in BALF in 30 cases of ILDs: including 18 patients with idiopathic pulmonary fibrosis (IPF) and 12 patients with sarcoidosis (Sarc) and 9 healthy controls were determined. RESULT: (1) The levels of BALF-SOD, GSH-PX were significantly decreased(P < 0.01), and BALF-ACE, LDH were markedly increased (P < 0.05) in patients with IPF, and BALF-ACE were also increased in patients with Sarc(P < 0.05); (2) There were good correlation between ACE and percentage of lymphocyte (r1 = 0.6574, P < 0.05), and ratio of CD4+/CD8+ (r2 = 0.9544, P < 0.001) in BALF of Sarc group. CONCLUSION: Determining enzymatic activities could be helpful to study pathogenesis and diagnosis of ILDs, BALF-ACE may be as a good marker of disease activity of Sarcoidosis. PMID- 11263393 TI - [The tuberculin test in intrathoracic sarcoidosis]. AB - OBJECTIVE: To highlight the diagnosis and treatment of intrathoracic sarcoidosis. METHOD: The clinical and laboratory data of 35 cases of intrathoracic sarcoidosis in Tianjin thoracic hospital from 1991 to 1996 were reviewed. Tuberculin tests for 35 cases of sarcoidosis were carried out and the results were evaluated. Simultaneously, the results have been compared with that of literature published. RESULT: Of 35 cases of intrathoracic sarcoidosis 17 cases were tuberculin positive, 4 were strongly positive, positive tuberculin reactivity (60%) was significantly higher than that published. CONCLUSION: The results showed a considerably higher tuberculin positive rate, which may be due to the difference of positive tuberculin reactivity in different country or regions, age of patients and expression of gamma delta T cell receptor. The authors suggested that "sarcoidosis should be diagnosed with negative tuberculis reastivity" has to be reconsidered. PMID- 11263394 TI - [Occurrence of hyperventilation syndrome in Chinese patients]. AB - OBJECTIVE: To evidence the occurrence of hyperventilation syndrome in Chinese patients. METHOD: Case report and literature review. RESULT: Three cases with manifest hyperventilation syndrome were reported. The diagnosis was based on the presence of several suggestive complaints occurring in a context of stress, and the reproduction of the most important complaints by the hyperventilation provocation test. Organic diseases as a cause of the symptoms had been excluded. Breathing therapy reducing the tendency to hyperventilate by acquiring an abdominal breathing pattern, with slowing down of expiration, markedly reduced complaints in the three patients. CONCLUSION: Hyperventilation syndrome occurs also in Chinese patients. PMID- 11263396 TI - [Proceedings of the 4th European Congress of Oto-Rhino-Laryngology Head and Neck Surgery. Berlin, Germany, 2000]. PMID- 11263395 TI - Molecular basis of morphogenesis. Proceedings of a discussion meeting. 16-17 November 1999. PMID- 11263397 TI - Issue dedicated to Professor Jan Bubenik, M.D., D.Sc. PMID- 11263398 TI - 6th European Platelet, Granulocyte and Red Cell Immunobiology meeting. September 2000. Abstracts. PMID- 11263399 TI - Precancerous lesions of the digestive tract. The 18th Annual Symposium of the European Cancer Prevention Organization. Maastricht, The Netherlands. 12-14 October 2000. Abstracts. PMID- 11263400 TI - Proceedings of the 29th International Symposium on Growth Hormone and Growth Factors in Endocrinology and Metabolism. Marrakech, Morocco, 7-8 April 2000. PMID- 11263401 TI - From the Centers for Disease Control and Prevention. Serious adverse events attributed to nevirapine regimens for postexposure prophylaxis after HIV exposures--worldwide, 1997-2000. PMID- 11263402 TI - From the Centers for Disease Control and Prevention. Public health dispatch: outbreak of poliomyelitis--Dominican Republic and Haiti, 2000. PMID- 11263403 TI - From the Centers for Disease Control and Prevention. Recommendations from meeting on strategies for improving global measles control, May 11-12, 2000. PMID- 11263404 TI - From the Centers for Disease Control and Prevention. Foodborne outbreak of group A rotavirus gastroenteritis among college students--District of Columbia, March April 2000. PMID- 11263405 TI - JAMA patient page. Breastfeeding. PMID- 11263406 TI - Nonsteroidal anti-inflammatory drugs and the gastrointestinal tract: consensus and controversy. Proceedings of a symposium. September 23-25, 1999, Porto Cervo, Sardinia, Italy. PMID- 11263407 TI - Special issue in honour of Professor Mats Almgren on the occasion of his 60th birthday. PMID- 11263408 TI - Iron deficiency anemia: reexamining the nature and magnitude of the public health problem. Proceedings of a conference. May 21-24, 2000. Belmont, Maryland, USA. PMID- 11263409 TI - Ecological Management of Post-Mining Areas. Proceedings of a meeting. Cottbus, Germany, March 1999. PMID- 11263410 TI - Cardiovascular Radiation Therapy III. February 17-19, 1999. Washington, DC, USA. Abstracts. PMID- 11263411 TI - Ulrich Wolf dedication volume. PMID- 11263413 TI - Letter in answer to Oliver's article published in atherosclerosis 2000; 150:1-12. Investigations of the Medical Research Council's Social Medicine Unit. PMID- 11263412 TI - Arachidonic acid specifically regulates binding of S100A8/9, a heterodimer complex of the S100 class of calcium binding proteins, to human microvascular endothelial cells. PMID- 11263414 TI - Can dendritic cells be exploited for therapeutic intervention in atherosclerosis? PMID- 11263415 TI - Biological cardiovascular risk factors and plasma homocysteine levels in the general Dutch population. PMID- 11263416 TI - Renovascular hypertension detected by magnetic resonance arteriography. PMID- 11263417 TI - From the Centers for Disease Control and Prevention. Hypothermia-related deaths- Suffolk County, New York, January 1999-March 2000, and United States, 1979-1998. PMID- 11263419 TI - JAMA patient page. Down syndrome. PMID- 11263418 TI - From the Centers for Disease Control and Prevention. Outbreak of Ebola hemorrhagic fever--Uganda, August 2000-January 2001. PMID- 11263420 TI - [Formation of a new association: the AFJCMF (French Association of Young Maxillofacial Surgeons]. PMID- 11263423 TI - Direct access to specialists not necessarily more expensive. PMID- 11263424 TI - Folic acid antagonists during pregnancy and risk of birth defects. PMID- 11263425 TI - Folic acid antagonists during pregnancy and risk of birth defects. PMID- 11263426 TI - Folic acid antagonists during pregnancy and risk of birth defects. PMID- 11263427 TI - Folic acid antagonists during pregnancy and risk of birth defects. PMID- 11263428 TI - Screening for lung cancer. PMID- 11263421 TI - Policy statements adopted by the Governing Council of the American Public Health Association, November 15, 2000. PMID- 11263429 TI - Screening for lung cancer. PMID- 11263430 TI - Screening for lung cancer. PMID- 11263431 TI - Roads to breast cancer. PMID- 11263432 TI - Roads to breast cancer. PMID- 11263433 TI - Bilateral atrial myxomas associated with hyperpigmented skin lesions. PMID- 11263434 TI - Hematuria in a patient with an ileal conduit and hepatic cirrhosis. PMID- 11263435 TI - Needle length and incidence of local reactions to immunization. Needle gauge is more important than needle length. PMID- 11263436 TI - Comments on: Recommendations on colorectal cancer (CRC)screening in the European Union. Advisory Committee on Cancer Prevention. Eur J Cancer 2000, 36, 1473-1478. PMID- 11263437 TI - A phase II trial of bryostatin 1 in patients with non-Hodgkin's lymphoma. AB - Bryostatin 1 is a naturally occurring macrocyclic lactone with promising antitumour and immunomodulatory function in preclinical and phase I clinical investigations. In this phase II study, 17 patients with progressive non Hodgkin's lymphoma of indolent type (NHL), previously treated with chemotherapy, received a median of 6 (range 1-9) intravenous infusions of 25 microg/m(2) bryostatin 1 given once weekly over 24 hours. In 14 evaluable patients no responses were seen. Stable disease was attained in one patient for 9 months. The principal toxicities were myalgia and phlebitis. Treatment was discontinued early because of toxicity alone (phlebitis) in 2 patients, toxicity in addition to progressive disease in 3 patients (myalgia and phlebitis n = 2; thrombocytopenia n = 1) and progressive disease in 5 patients. The results fail to demonstrate efficacy of this regimen of bryostatin 1 in the treatment of NHL. In light of preclinical data that demonstrate synergy between bryostatin 1 and several cytotoxic agents and cytokines, clinical studies to investigate bryostatin 1 in combination are warranted. We also present data to demonstrate that central venous lines may be used in future studies to avoid phlebitis. PMID- 11263438 TI - CD38 expression is a poor predictor for VH gene mutational status and prognosis in chronic lymphocytic leukemia. PMID- 11263439 TI - Factor XIIIV34L is not an additional genetic risk for venous thrombosis in factor V Leiden carriers. PMID- 11263440 TI - Clinical and bone marrow effects of interferon alfa therapy in myelofibrosis with myeloid metaplasia. PMID- 11263441 TI - Radiation-induced leukemia. PMID- 11263442 TI - Criteria for diagnosis of acute megakaryocytic leukemia. PMID- 11263443 TI - T-cell apoptosis in HIV-1-infected individuals receiving highly active antiretroviral therapy. PMID- 11263444 TI - ALK is not expressed in Hodgkin disease. PMID- 11263445 TI - CD38 expression and Ig VH gene mutation in B-cell chronic lymphocytic leukemia. PMID- 11263447 TI - Photo finish for Barrett's esophagus, other digestive tract lesions? PMID- 11263448 TI - GERD remains an intriguing enigma. PMID- 11263446 TI - [Hair loss]. PMID- 11263449 TI - Prognosis of patients with a recurrent acute myocardial infarction before and in the reperfusion era--a national study. AB - BACKGROUND: Patients with recurrent acute myocardial infarction (AMI) are at increased risk for morbidity and mortality. We compared the outcome of patients with recurrent AMI hospitalized in coronary care units in the prereperfusion and reperfusion eras. METHODS: The study population comprised 2 large-scale cohorts with recurrent AMI: (1) 1415 (24%) of 5839 consecutive patients with AMI hospitalized in 1981 to 1983 (Secondary Prevention Reinfarction Israeli Nifedipine Trial [SPRINT] Registry) and (2) 1093 (25%) of 4317 patients with AMI from three national surveys performed in 1992 to 1996. RESULTS: Patients in the 1990s had significantly lower rates of heart failure and cardiogenic shock. The 7 day mortality declined from 18% in 1981-1983 to 10% in 1992-1996 (adjusted odds ratio [OR] 0.57 [0.44-0.75]), the 30-day mortality rate from 26% to 16% (OR 0.56 [0.44-0.71]), and the 1-year mortality rate from 39% to 26% (adjusted hazard ratio [HR] 0.64 [0.54-0.75]), respectively. In the 1992-1996 cohort, the adjusted risk of 7-day, 30-day, and 1-year mortality for patients with recurrent AMI treated with thrombolysis in comparison to patients without thrombolysis was OR 1.69 (1.07-2.65), 1.52 (1.03-2.23), and HR 1.18 (0.90-1.55), respectively. The mortality rate among patients treated with early percutaneous transluminal coronary angioplasty/coronary artery bypass grafting was 3% versus 12% at 7 days (OR 0.36 [0.16-0.73]), 7% versus 18% at 30 days (OR 0.45 [0.25-0.77]), and 16% versus 29% at 1 year (HR 0.64 [0.46-0.96]), in comparison to patients without revascularization. CONCLUSION: The prognosis of patients with recurrent AMI improved significantly during the reperfusion era. Although thrombolysis may have a limited therapeutic effect among patients with recurrent AMI, an interventional approach seems more appropriate when indicated. A randomized trial of thrombolysis versus early revascularization is needed in patients with recurrent AMI. PMID- 11263450 TI - AIDS vaccine shows promise after years of frustration. PMID- 11263451 TI - Severity of overdose after restriction of paracetamol availability. Study's results conflict with those of other papers. PMID- 11263452 TI - Severity of overdose after restriction of paracetamol availability. Restriction has not reduced admissions with self poisoning. PMID- 11263453 TI - Severity of overdose after restriction of paracetamol availability. Why hasn't strategy for minimising paracetamol poisoning been enacted? PMID- 11263455 TI - Guidelines for prevention of falls in people aged over 65. Health improvement plans must incorporate falls and osteoporosis strategies. PMID- 11263454 TI - Guidelines for prevention of falls in people aged over 65. Guidelines should state that assessment of vision is important. PMID- 11263456 TI - Unicef is crucial in promoting and supporting breast feeding. PMID- 11263457 TI - Evidence and belief in attention deficit hyperactivity disorder. Narrow focus of editorial was disappointing. PMID- 11263458 TI - Evidence and belief in attention deficit hyperactivity disorder. Reintroduction of methylphenidate in Italy needs careful monitoring. PMID- 11263459 TI - Twins and asthma. Difference in admission rates may be due to other factors. PMID- 11263460 TI - Eradicating Helicobacter pylori in non-ulcer dyspepsia may not be cost effective. PMID- 11263461 TI - [Continued ultrasonic follow-up of children infected with Schistosoma haematobium after treatment with praziquantel]. AB - During a Schistosoma haematobium morbidity control program in Niger, we conducted a survey to describe rhe resolution of lesions after treatment with praziquantel. to determine reinfection rates and to define retreatment schedules. 114 schoolchildren (7-15 years old) living in an hyperendemic village underwent 10 successive examinations over 34 months following an initial evaluation and the administration of 40 mg/kg of praziquantel. All children, whether apparently infected with S. haematobium or not, were treated. Egg output, microhaematuria, visual aspect of urine and abnormalities of the urinary tract by ultrasound were assessed. The initial prevalence tif infection was 74.5%. Reinfection began 5 months after treatment and the final prevalence was 47.1%. Bladder abnormalities decreased rapidly, but incompletely, probably due to reinfestation: initial prevalence: 89.5%). Their prevalence increased 8 months after treatment to 72.4% at month 34. Dilatations of the upper urinary tract regressed more slowly but constantly until the end of the study (initial prevalence: 43%; 4.6% at month 34), Three years after treatment, despite reinfection, the general morbidity level (prevalence and severity of lesions) was lower than at baseline in our cohort, which would suggest the advantage of a long interval between mass treatments in the epidemiological context of our survey. PMID- 11263462 TI - Unreliability of PAP smears to diagnose female genital schistosomiasis. PMID- 11263463 TI - Neurological long-term sequelae after spinal anaesthesia in a tropical setting: a case control study. AB - Spinal anaesthesia (SA) is an important form of anaesthesia in tropical countries. It is considered to have few long-term complications or sequelae, although this hypothesis has not been proven in a rural tropical setting. In a case control study we found SA to he a significant risk factor for lower back pain, reflex abnormalities and muscular atrophy and mild impairment of muscle power in patients examined between 3 months and 4 years after SA. These long-term sequelae need to be confirmed in a larger prospective study employing all possible neurophysiological and neuroimaging techniques. PMID- 11263464 TI - Inhibitory effects of 9-anilinoacridines on Plasmodium falciparum gametocytes. AB - Two gametocyte-producing isolates of Plasmodium falciparum, KT1 arid KT3, were cultivated in vitro. On day 11 of cultivation, pure gametocytes containing stage II, III and IV were used to test the gametocytocidal activity of 9 anilinoacridines that had previously demonstrated their activity against the asexual stage of the parasite. After drug exposure for 48 h, gametocytes were maintained without drugs for another 2 days before thin films were prepared for parasite counting. Gametocytocidal activities of 13 analogs of 9-anilinoacridine were observed with 50% inhibitory concentrations in the range of 0.6 microM to> 100 microM The most active compound was 1'-CH2NMe2-9-anilinoacridine. Anilinoacridine derivatives with 3,6-diamino substitution had reduced gametocytocidal activity in contrast to their enhancing effect against the asexual forms. Morphological abnormalities of gametocytes were observed following drug exposure. PMID- 11263465 TI - Therapeutic injections in Pakistan: from the patients' perspective. AB - OBJECTIVE: To investigate the behaviour, knowledge of risks, and attitudes towards injections among patients at a clinic in Karachi. METHODS: In March 1995, trained staff administered a structured questionnaire to 198 consecutive new adult patients attending a university clinic in Karachi, Pakistan. RESULTS: Half (97:49%) of the patients received injections at their last visit to a health care provider. 3.5% had received 10 or more injections in the last year. 64% felt that injections were more powerful and were willing to pay more for them than for pills. 84% preferred pills or advice over injections if told they were equally effective, 83% believed that a used needle could transmit a fatal disease, and 86% believed that it is usually possible to get better without an injection. 91% reported that the doctor always recommends an injection; few patients (9%) ever asked for one. Injections were given without much regard for the chief complaint of the patient. Sonic needles (n = 21) for the injection came from bowls of water: of those from closed packets (n = 116), 68 were 'cleaned' by wiping ot placing them in water. 91% of patients (180) knew at least one risk of reuse of needles. Patients who knew three or more risks of using unclean needles were 0.14 times as likely to have had more than five injections per year in the last 5 years hut only if the patients had s or more years of education. CONCLUSION: Patients receive injections from doctors in Pakistan frequently, indiscriminately and often without proper safety precautions. They are aware of both positive and negative aspects of injections but are likely to do what the doctor suggests. Interventions to reduce risky overuse of injections should focus on patients' general education and knowledge of the risks of injections to empower them to choose healthier therapies. PMID- 11263466 TI - Peritoneal dialysis in an infant with type 1 diabetes and hyperosmolar coma. AB - Hyperosmolar coma which is characterized by severe hyperglycemia in absence of chetosis is very rare in pediatric age with only 11 cases reported in the literature. The outcome of the condition is usually poor with mental retardation being the most common event. Here a case of hyperosmolar coma is described in a female of three months of age who was treated with peritoneal dialysis 11 hours after admittance to hospital. This female patient has been receiving insulin from three months of age and today at the age of 10 years she leads a normal life despite being on insulin therapy. A very low level of C-peptide (<0.3 ng/ml) clearly confirms she is affected by Type 1 diabetes. To our knowledge this is the first case report of hyperosmolar coma in a neonate with Type 1 diabetes who survived this condition without late neurological consequences. PMID- 11263467 TI - Non-secreting atypical parathyroid adenoma. AB - Parathyroid tumors can be divided in adenomas and carcinomas, usually detected by hypercalcemia. We report a case of parathyroid adenoma in a young man, who complained of a pressure in the left neck region. Physical examination revealed a firm mass in the neck, without lymphnodes. Although Ca (9.7 mg/dl), phosphorus (3.3 mg/dl) and intact-PTH (49 pg/ml) were normal, imaging techniques (computed tomography scan and sestamibi substraction scan) suggested that the mass could arise from the parathyroid gland. Histology and immune staining for chromogranin and parathyroid hormone confirmed the parathyroid nature of the mass. Histological criteria defined the lesion as an atypical parathyroid adenoma. We review the pathology, diagnosis and treatment of parathyroid adenomas in its non secreting atypical form. PMID- 11263468 TI - Extensive sterile abscess in an invasive fibrous thyroiditis (Riedel's thyroiditis) caused by an occlusive vasculitis. AB - Riedel's thyroiditis is a rare disease determined by an invasive fibrosclerotic transformation of the thyroid gland. It may be one manifestation of multifocal fibrosis with still unknown etiology. Because it mimics carcinoma, a biopsy must be performed to get the correct diagnosis. The condition is self-limiting when confined to the neck. Prognosis depends on the extent of extracervical fibrosclerosis. We present a patient with a huge cervical and mediastinal, unilateral thyroid mass expanding to the aortic curve, which led to tracheal deviation and compression with symptoms of stridor and dyspnea. These symptoms continued under a course of high-dose steroids; thus an operation was necessary to relieve the airway obstruction and limit inflammation. Intraoperative and pathological findings showed an inflammatory infiltration of the adjacent neck muscles and a sterile abscess caused by an occlusive vasculitis. Therefore, hemithyroidectomy had to be performed instead of a local limited resection. PMID- 11263470 TI - Parathyroidectomy for asymptomatic primary hyperparathyroidism (PHPT): is it worth the risk? PMID- 11263471 TI - Decrease of serum leptin levels in adult male with idiopathic hypogonadotropic hypogonadism (IHH) treated with pulsatile gonadotropin-releasing hormone (GnRH). PMID- 11263469 TI - Effects of amiodarone administration during pregnancy on neonatal thyroid function and subsequent neurodevelopment. AB - Amiodarone, a benzofuranic derivative, iodine-rich drug, has been used in pregnancy for either maternal or fetal tachyarrhythmias. Amiodarone, its main metabolite (desethylamiodarone) and iodine are transferred, albeit incompletely, through the placenta, resulting in a relevant fetal exposure to the drug and iodine overload. Since the fetus acquires the capacity to escape from the acute Wolff-Chaikoff effect only late in gestation, the iodine overload may cause fetal/neonatal hypothyroidism and goiter. Among the reported 64 pregnancies in which amiodarone was given to the mother, 11 cases (17%) of hypothyroidism in the progeny (10 detected at birth, 1 in utero) were reported, 9 non-goitrous (82%) and 2 (18%) associated with goiter. Hypothyroidism was transient in all cases, and only 5 infants were treated short-term with thyroid hormones. Only 2 newborns had transient hyperthyroxinemia, associated with low serum TSH concentrations in one. Neurodevelopment assessment of the hypothyroid infants, when carried out, showed in some instances mild abnormalities, most often reminiscent of the Non verbal Learning Disability Syndrome; however, these features were also reported in some amiodarone-exposed euthyroid infants, suggesting that there might be a direct neurotoxic effect of amiodarone during fetal life. Breast-feeding was associated with a substantial ingestion of amiodarone by the infant, but in the few cases followed it did not cause changes in the newborn's thyroid function. In conclusion, amiodarone therapy during pregnancy may cause fetal/neonatal hypothyroidism and, less frequently, goiter. Thus, the use of amiodarone in pregnancy should be limited to maternal/fetal tachyarrhythmias which are resistant to other drugs or life-threatening. If amiodarone is used during gestation, a careful fetal/neonatal evaluation of thyroid function and morphology is warranted. It seems prudent to advise that fetal/neonatal hypothyroidism be treated, as soon as the diagnosis is made, even in utero, to avoid neurodevelopment abnormalities, although the latter may occur independently of hypothyroidism. If breast-feeding is allowed, careful evaluation of the infant's thyroid function and morphology is required because of the continuing exposure of the infant to the drug. PMID- 11263472 TI - Endocrinology and art. "A ferocious cretin with voluminous goiter". Morazzone, circa 1600. PMID- 11263474 TI - Side-effects of iodized oil administration in patients with simple goiter. AB - The objective of this study was to determine side-effects associated with iodized oil injection in patients with simple goiter. In an iodine-deficient population, 3420 patients with simple goiter, who were not taking supplemental iodine, were chosen for this study. They received a single intramuscular injection of 1 ml iodized oil, containing 480 mg iodide. Clinical and laboratory evaluations were performed every 3 months for one year and every 6 months for the next 4 years. The incidence of hypo- and hyperthyroidism was 0.6% each, with equal prevalence in both sexes. Most cases of hypo- and hyperthyroidism were observed during the first 5 months after the injection. Eight cases of hyperthyroidism were asymptomatic. A further 8 patients had overt thyrotoxicosis and required treatment with methimazole for 18 months. Recurrence of hyperthyroidism was observed in one patient. Five hypothyroid patients were diagnosed only by abnormal thyroid function tests, and 4 cases needed no treatment. Others received T4 treatment for a mean of 14.5 months. Among 14 T4-treated patients, recurrence of hypothyroidism occurred in 7 patients after treatment was discontinued. Twenty nine patients (0.8%) were afflicted with dermatologic complications. The most common dermatologic side-effect was urticarial reaction. In 15 subjects, skin lesions appeared 8 to 14 days after injection. It is concluded that side-effects of iodized oil injection are rare, and in most cases the complications are transient and self-limited. The occurrence of iodine induced hyperthyroidism following iodized oil administration is close to the ratio observed in spontaneous thyrotoxicosis. PMID- 11263473 TI - Recombinant human IGF-I does not modify the ACTH and cortisol responses to hCRH and hexarelin, a peptidyl GH secretagogue, in humans. AB - An inhibitory influence of insulin-like growth factor-I (IGF-I) on hypothalamus pituitary-adrenal (HPA) axis has been hypothesized. In fact, it has been reported that the rhGH (recombinant human GH)-induced IGF-I increase inhibits both cortisol and GH response to MK-0677, a non-peptidyl GH secretagogue in animals. The aim of this study was to further clarify the inhibitory role, if any, of IGF I on corticotroph function. We studied the effect of rhIGF-I (recombinant human IGF-I; 20 microg/kg s.c. at -180 min) or placebo on the ACTH and cortisol responses to hCRH (human CRH; 2.0 microg/kg i.v. at 0 min) or hexarelin (HEX; 2.0 microg/kg i.v. at 0 min), a peptidyl GHS, in normal young women. The effect of rhIGF-I on the GH response to HEX was also studied. The subjects were six normal young women [age: 26-35 yr; body mass index (BMI): 19-23 kg/m2] in their early follicular phase. The results showed that after s.c. rhIGF-I administration, circulating IGF-I levels increased approximately 77%, peaking at -60 min and persisting similar up to +120 min. The mean ACTH, cortisol and GH concentrations did not change from -180 to 0 min when evaluated after both placebo or rhIGF-I. CRH and HEX induced similar ACTH (peak vs baseline, mean+/-SE: 47.5+/-10.9 vs 21.3+/-3.0 pg/ml and 30.3+/-6.9 vs 19.2+/-3.8 pg/ml, respectively; p<0.04) and cortisol responses (177.5+/-5.4 vs 109.3+/-10.3 microg/l and 149.4+/-12.3 vs 119.8+/-16.4 microg/l, respectively, p<0.04). RhIGF-I pretreatment did not modify the ACTH and cortisol responses to hCRH (46.0+/-13.8 pg/ml and 181.1+/-16.9 microg/l, respectively) as well as those to HEX (28.8+/-5.0 pg/ml and 144.1+/ 16.2 microg/l, respectively). On the other hand, the GH response to HEX was clearly reduced by rhIGF-I (23.9+/-4.7 vs 64.7+/-14.8 microg/l, p<0.05). Our findings show that rhIGF-I-induced increase of circulating IGF-I levels exerts negative feedback action on somatotroph secretion, while it does not modify the corticotroph and the adrenal responsiveness to CRH or hexarelin. PMID- 11263475 TI - Intestinal permeability in adult patients with growth hormone deficiency. AB - Pathological disruption of the intestinal mucosa increases the paracellular pathway, leading to an increase in the penetration of large molecules. Since growth hormone (GH) has a trophic intestinal effect, we used a double marker test to enable examination of intestinal permeability, which reflects the state of integrity of the intestinal mucosa. We recruited 22 adult patients, mean age 54+/ 13.3 years, with GH deficiency due to partial or total hypopituitarism. None had received GH treatment at any time, although they were all in optimized replacement therapy. A control group was composed of 19 healthy age-matched relatives. The intestinal permeability test was performed with lactulose (5 g) and mannitol (1 g) after an oral load of 100 ml of aqueous solution. The urinary lactulose/mannitol ratio and the percentages of lactulose and mannitol excreted were determined on a 5-h urine collection. There were no significant differences between the patients and the control group in the lactulose/mannitol ratio (0.087+/-0.059 vs 0.077+/-0.064, respectively) or in the urinary excretion percentages of lactulose (0.067+/-0.048% vs 0.073+/-0.070%, respectively) or mannitol (5.127+/-3.269% vs 5.068+/-2.985%, respectively). In conclusion, no increase in intestinal permeability was detected in patients with GH deficiency, so that in spite of the known trophic effects of GH on the epithelial crypt cells, there was no intestinal hyperpermeability in these patients. PMID- 11263476 TI - Adrenocorticotropin levels do not change during early recovery of transsphenoidal surgery for ACTH-secreting pituitary tumors. AB - In patients with ACTH-secreting pituitary tumor the peri-tumoral normal corticotrophs were supposed to be suppressed by cronic hypercortisolemia since frequently they develop transient secondary adrenal insufficiency after pituitary tumor resection and during early postoperative days. We evaluated the ACTH dynamics during transsphenoidal surgery in 16 patients with ACTH-secreting pituitary tumors (6 cured by surgery, 8 not cured Cushing's disease patients and 1 cured by surgery and 1 not cured Nelson's syndrome patients) and tested the hypothesis that in these patients, ACTH secretion from the peri-tumoral normal corticotrophs is inhibited and hence removal of the entire tumor should result in subtle postoperative reduction in plasma ACTH. Blood samples for ACTH determination were obtained from 14 Cushing's disease patients immediately before pituitary gland manipulation and 10, 30, 60, 90, 120, 150 and 300 min after pituitary tumor resection and on postoperative day one. In Nelson's syndrome patients the blood sample was obtained only after tumor removal. All patients received intravenous hydrocortisone during surgery and on the first postoperative day. Patients were considered cured by surgery if they presented adrenal insufficiency after hydrocortisone withdrawal. Mechanical pituitary manipulation induced increase in ACTH level. In all 14 Cushing's disease patients (cured and not cured), mean plasma ACTH levels were significantly greater 10 min after pituitary tumor resection (54.4+/-12.8 pmol/l) than in the premanipulation period (ACTH=26.3+/-5.3 pmol/l) (p=0.005). In Cushing's disease patients, the ACTH levels did not change significantly until 300 min after pituitary tumor resection either in those 6 patients cured by surgery (at 10 min after pituitary tumor resection ACTH was 54.4+/-12.8 pmol/l for all 14 Cushing's disease patients and at 300 min after tumor removal ACTH was 39.0+/-12.6 pmol/l for cured and 41.3+/ 15.7 pmol/l for not cured Cushing's disease patients). The ACTH level also persisted high until 300 min after complete pituitary tumor resection in one cured patient with Nelson's syndrome. ACTH level does not change in the early recovery period after ACTH-secreting pituitary tumor, even in those cured patients, and probably peri-tumoral normal corticotrophs are not completely suppressed by cronic hypercortisolemia (and acute glucocorticoid administration) when these patients are under intense stress, like transsphenoidal surgery. Mechanical pituitary manipulation may induce ACTH release in patients with ACTH secreting pituitary tumors but probably does not interfere in the maintenance of high ACTH-levels during the early postoperative period, since ACTH half-life is only 8-15 min. In patients with ACTH-secreting pituitary tumors, the behavior of the human hypothalamic-pituitary-adrenal system during transsphenoidal surgery does not conform to the specifications of a negative feedback mechanism. PMID- 11263477 TI - HLA DQA1*0501 and DRB1*0301 antigens do not independently convey susceptibility to Graves' disease. AB - Genes of, or closely associated to, the HLA complex are assumed to contribute to the genetic predisposition of Graves' disease. The aim of this study was to investigate the presence of the HLA DQA1*0501 and DRB1*0301 antigens in Greek patients with Graves' disease. In addition, we tried to establish if there is any association between these antigens and any of the clinical manifestations of the disease. We examined 117 patients with Graves' disease and 104 healthy controls. DNA was extracted from peripheral lymphocytes and the HLA DQA1*0501 and DRB1*0301 genomic regions were amplified by PCR and characterized by hybridization with sequence specific oligonucleotides (SSO). Two of the patients had a positive family history for Graves' disease and 46 had clinical thyroid eye disease (TED). The frequencies of both DQA1*0501 and DRB1*0301 antigens were significantly increased in patients compared to controls (relative risk [RR] 4.2 and 4.5 for each antigen respectively). Neither of these two antigens was an independent risk factor for Graves' disease. However, the combination of both these HLA antigens resulted in a striking increase in the RR for development of Graves' disease especially in females (RR/F=27, RR/M=8.4). No association was found between these antigens and positive family history or the presence of TED. These data suggest that HLA DQA1*0501 and DRB1*0301 antigens are not independent risk factors for the development of Graves' disease. On the contrary, the presence of both these alleles results in a significant increase in the RR for the development of Graves' disease in the Greek population, particularly in females. PMID- 11263479 TI - Improved yield and functionality of parathyroid cells separated by using collagenase-digestion with cold pre-incubation. AB - Preparation of cells from solid organs often induces a functional impairment due to the proteolytic cell damage by the applied digestive enzyme like collagenase, trypsin or dispase. To preserve the tissue and to enhance the yield of cells, Laue et al. reported an islet cell isolation with pre-incubation at 4 C permitting the enzyme to diffuse into the tissue and explicite activity equally throughout the whole particle. The aim of this study was to investigate whether this procedure can be applied to parathyroid glands. Therefore porcine parathyroid glands were dissected into 1 mm3 pieces. Subsequently one group of these pieces was incubated 22 h at 4 C in 2 mg/ml collagenase before activating the enzyme by elevating the temperature to 37 C for 30 min. The other group was incubated directly at 37 C for 30 min. The yield of cells and their viability was assessed by light-microscopy and staining with trypan-blue. After the cells were immobilized in barium-alginate and cultivated for 7 days, the function was tested by incubation in different calcium concentrations and PTH-measurement. Finally, the viability was assessed by histology. Using a cold pre-incubation with collagenase, a significantly higher number of isolated cells was retrieved compared with collagenase-digestion without pre-incubation. The viability was about 100% and did not differ between both groups. After immobilization and cultivation the viability decreased to less than 30%, with and without pre incubation. In contrast to viability the PTH-secretion of the cells differed significantly between both procedures. By pre-incubation with collagenase at 4 C a gentle method is presented resulting in an enhancement of yield and function of single cells of parathyroid glands. PMID- 11263478 TI - Pituitary adenomas in childhood and adolescence. Clinical analysis of 10 cases. AB - Pituitary adenomas in childhood and adolescence constitute 2-6% of all operated pituitary adenomas. We report the clinical features, treatment and follow-up of 10 pediatric patients affected by pituitary adenomas. All patients underwent clinical evaluation, endocrine tests, magnetic resonance imaging and visual field assessment. Follow-up ranged from 8 to 132 months (median 52.6). All patients were older than 10 years of age; 60% were males. In 50% the initial complaints were headache and/or visual impairment, all except one had clear evidence of endocrine dysfunction. Ninety percent were macroadenomas. According to hormone measurements and immunostaining 50% were prolactinomas, 20% were pure GH secreting and 30% were non-functioning adenomas. Prolactinomas in two females were successfully treated with cabergoline. The other patients underwent surgery: three prolactinomas are still being treated with dopamine agonists and a GH secreting adenoma is being treated with octreotide LAR and cabergoline. Two patients were also treated with conventional radiotherapy. Treatments were completely successful in 50% of patients: these have normal hormone secretion, full pubertal development, no significant tumor mass and normal visual field. Hypersecretion of prolactin persists in two cases; partial or complete hypopituitarism is present in four, relevant tumor remnant in another four and impairment of visual field is present in two cases. In conclusion, pediatric adenomas occur mostly in pubertal age, are prevalently macroadenomas and clinically functioning. Medical therapy should be preferred for secreting adenomas, but in some cases, notably prolactinomas in males, surgery and eventual radiotherapy may be needed. PMID- 11263481 TI - The secret ways of scientists. PMID- 11263482 TI - A cost/benefit analysis. About the precautionary principle. PMID- 11263480 TI - CDY1 analysis in infertile patients with DAZ deletions. AB - The DAZ (deleted in azoospermia) gene family is considered the major AZFc ("azoospermia factor" c) candidate responsible for male infertility. However, other genes have been recently mapped to this region and they could contribute to the AZFc phenotype. In particular, the testis-specific CDY1 (chromadomain protein 1) gene has one copy within the DAZ cluster and another one at its distal end. Therefore, this gene could be associated with the spermatogenic damage observed in DAZ-deleted patients since at least one CDY1 copy is invariably absent in these patients. In this study, we investigated whether selected infertile patients affected by different testiculopathies caused by DAZ deletions retained CDY1 and if a genotype-phenotype relation could be demonstrated. We found 3 out of 17 patients with absence of both CDY1 copies, while 14 patients have only one CDY1 copy absent. Two out of the 3 patients with deletion of both copies of CDY1 were affected by severe hypospermatogenesis while one patient presented Sertoli cell-only syndrome. Therefore, there was no clear relation between genotype and phenotype, and the loss of the distal copy of CDY1 does not seem to worsen the phenotype of infertile patients with deletion of the DAZ gene cluster. However, a possible contribution of CDY1 in determining the spermatogenic alteration could not be excluded. PMID- 11263483 TI - Molecular biology, China and the West. 15 years of teaching at the Max Planck Guest Laboratory in Shanghai. PMID- 11263484 TI - Creating a knowledge-based society. An interview with Noel Treacey, Minister for Science, Technology and Commerce of the Republic of Ireland. PMID- 11263485 TI - Same degree, same effort? A patchwork of differing PhD requirements throughout Europe disadvantages graduate students and compromises the quality of science. PMID- 11263486 TI - Trust, sensitivities and science. Scientists, policy makers and representatives of interest groups met in Brussels to discuss the future of genetic research in Europe. PMID- 11263487 TI - Embryo emergent: elucidating the cell biology of development. The Santa Cruz Conference on Developmental Biology 2000. PMID- 11263488 TI - The cytoskeleton: from regulation to function. Conference: the 15th Meeting of the European Cytoskeleton Forum. PMID- 11263489 TI - Proteolysis in Alzheimer's disease. Can plasmin tip the balance? PMID- 11263490 TI - Limiting DNA replication to once and only once. AB - In Escherichia coli cells, the origin of chromosomal replication is temporarily inactivated after initiation has occurred. Origin sequestration is the first line of defence against over-initiation, providing a time window during which the initiation potential can be reduced by: (i) titration of DnaA proteins to newly replicated chromosomal elements; (ii) regulation of the activity of the DnaA initiator protein; and (iii) sequestration of the dnaA gene promoter. This review represents the first attempt to consider together older and more recent data on such inactivation mechanisms in order to analyze their contributions to the overall tight replication control observed in vivo. All cells have developed mechanisms for origin inactivation, but those of other bacteria and eukaryotic cells are clearly distinct from those of E. coli. Possible differences and similarities are discussed. PMID- 11263491 TI - The beta clamp targets DNA polymerase IV to DNA and strongly increases its processivity. AB - The recent discovery of a new family of ubiquitous DNA polymerases involved in translesion synthesis has shed new light onto the biochemical basis of mutagenesis. Among these polymerases, the dinB gene product (Pol IV) is involved in mutagenesis in Escherichia coli. We show here that the activity of native Pol IV is drastically modified upon interaction with the beta subunit, the processivity factor of DNA Pol III. In the absence of the beta subunit Pol IV is strictly distributive and no stable complex between Pol IV and DNA could be detected. In contrast, the beta clamp allows Pol IV to form a stable initiation complex (t 1/2 approximately 2.3 min), which leads to a dramatic increase in the processivity of PoI IV reaching an average of 300-400 nucleotides. In vivo, the beta processivity subunit may target DNA Pol IV to its substrate, generating synthesis tracks much longer than previously thought. PMID- 11263492 TI - Mapping of a human centromere onto the DNA by topoisomerase II cleavage. AB - We have mapped the positions of topoisomerase II binding sites at the centromere of the human Y chromosome using etoposide-mediated DNA cleavage. A single region of cleavage is seen at normal centromeres, spanning approximately 50 kb within the centromeric alphoid array, but this pattern is abolished at two inactive centromeres. It therefore provides a marker for the position of the active centromere. Although the underlying centromeric DNA structure is variable, the position of the centromere measured in this way is fixed relative to the Yp edge of the array, and has retained the same position for >100,000 years. PMID- 11263493 TI - Escherichia coli cell cycle control genes affect chromosome superhelicity. AB - We have used ethidium bromide titration for direct measurement of the changes in the negative supercoiling of Escherichia coli chromosome caused by mutations inactivating the cell cycle functions mukB and seqA. The amounts of the intercalative agent required to relax the supercoiled chromosome in mukB and seqA mutants were lower and higher, respectively, than for the wild-type parent, confirming that these cell cycle genes modulate the topology of the E. coli chromosome. Plasmid superhelicity measured in these mutant strains showed similar effects albeit of reduced magnitude. As the effects of mukB and seqA mutations were not restricted to the chromosome alone, MukB and SeqA proteins possibly interact with factors involved in the maintenance of intracellular DNA topology. To our knowledge, this is the first direct demonstration of the influence of mukB and seqA genes on the superhelicity of the E. coli chromosome. PMID- 11263495 TI - The yeast mitotic cyclin Clb2 cannot substitute for S phase cyclins in replication origin firing. AB - Cyclin-dependent kinases (CDKs) drive the cell cycle, central to which is the accurate control of chromosome replication. In Saccharomyces cerevisiae, six closely related B-type cyclins (Clb1-6) drive the events of S phase and mitosis. Either Clb5 or Clb6 can activate early-firing replication origins, whereas only Clb5 can activate late origins. Clb1-4 are expressed later in the cell cycle. Whether Clb cyclins differ only in timing of expression, or else impart different kinase specificities is under ongoing investigation. This study shows that the expression of Clb2 during S phase in cells lacking Clb5 failed to rescue late origin activation. Early expression of Clb2 in cells lacking both Clb5 and Clb6 did not activate early origins on schedule to restore the correct S phase entry time. Therefore, Clb2 cannot drive timely activation of either early or late replication origins, demonstrating that Clb2-directed CDK has a specificity distinct from that driven by Clb5 and Clb6. PMID- 11263494 TI - The murine SNF5/INI1 chromatin remodeling factor is essential for embryonic development and tumor suppression. AB - The assembly of eukaryotic DNA into nucleosomes and derived higher order structures constitutes a barrier for transcription, replication and repair. A number of chromatin remodeling complexes, as well as histone acetylation, were shown to facilitate gene activation. To investigate the function of two closely related mammalian SWI/SNF complexes in vivo, we inactivated the murine SNF5/INI1 gene, a common subunit of these two complexes. Mice lacking SNF5 protein stop developing at the peri-implantation stage, showing that the SWI/SNF complex is essential for early development and viability of early embryonic cells. Furthermore, heterozygous mice develop nervous system and soft tissue sarcomas. In these tumors the wild-type allele was lost, providing further evidence that SNF5 functions as a tumor suppressor gene in certain cell types. PMID- 11263496 TI - Messenger RNAs that are not synthesized by RNA polymerase II can be 3' end cleaved and polyadenylated. AB - The poly(A) tail of influenza virus mRNAs is synthesized by the viral RNA polymerase by reiterative copying of a U5-7 sequence near the 5' end of the viral RNA (vRNA) template. We have engineered a vRNA molecule by replacing its viral U6 poly(A) site with a negative-sense eukaryotic polyadenylation signal. The vRNA was transcribed by the viral RNA polymerase and the transcription product was processed by the cellular 3' end processing machinery in vivo. According to the current model, 3' end processing of eukaryotic pre-mRNAs is coupled to cellular RNA polymerase II (pol II) transcription; thus only RNAs synthesized by pol III are believed to be polyadenylated efficiently. Our results show that the cellular polyadenylation machinery is nevertheless able to recognize and process RNA transcripts that are not synthesized by pol II, indicating that synthesis by pol II is not an absolute requirement for 3' end processing in vivo. PMID- 11263497 TI - APC-mediated downregulation of beta-catenin activity involves nuclear sequestration and nuclear export. AB - Mutational inactivation of adenomatous polyposis coli (APC) initiates most colon carcinomas. APC functions include targeting cytoplasmic beta-catenin, a Wnt pathway mediator, for proteolysis. Although APC shuttles between cytoplasm and nucleus, the role of nuclear APC protein, particularly with respect to nuclear beta-catenin levels and activity, remains unclear. Here, we demonstrate that APC lacking functional nuclear localization signals (NLSs) or nuclear export signals (NESs) does not effectively downregulate nuclear beta-catenin levels; neither does wild-type APC when nuclear export is blocked. While APC bearing mutated NLSs could not downregulate beta-catenin-mediated transcriptional activation, APC lacking NESs remained active. Consistent with the hypothesis that nuclear APC lacking NESs can inhibit beta-catenin function by sequestration, we show that endogenous APC and beta-catenin proteins interact within the nucleus. These data demonstrate that nuclear APC binding to beta-catenin, and then inducing its nuclear export, plays a critical role in the control of nuclear beta-catenin levels and activity. PMID- 11263498 TI - The PACT domain, a conserved centrosomal targeting motif in the coiled-coil proteins AKAP450 and pericentrin. AB - AKAP450 (also known as AKAP350, CG-NAP or Hyperion) and pericentrin are large coiled-coil proteins found in mammalian centrosomes that serve to recruit structural and regulatory components including dynein and protein kinase A. We find that these proteins share a well conserved 90 amino acid domain near their C termini that is also found in coiled-coil proteins of unknown function from Drosophila and fission yeast. Fusion of the C-terminal region from either protein to a reporter protein confers a centrosomal localization, and overexpression of the domain from AKAP450 displaces endogenous pericentrin, suggesting recruitment to a shared site. When isolated from transfected cells the C-terminal domain of AKAP450 was associated with calmodulin, suggesting that this protein could contribute to centrosome assembly. PMID- 11263499 TI - Brain plasmin enhances APP alpha-cleavage and Abeta degradation and is reduced in Alzheimer's disease brains. AB - The proteolytic processing of amyloid precursor protein (APP) has been linked to sphingolipid-cholesterol microdomains (rafts). However, the raft proteases that may be involved in APP cleavage have not yet been identified. In this work we present evidence that the protease plasmin is restricted to rafts of cultured hippocampal neurons. We also show that plasmin increases the processing of human APP preferentially at the alpha-cleavage site, and efficiently degrades secreted amyloidogenic and non-amyloidogenic APP fragments. These results suggest that brain plasmin plays a preventive role in APP amyloidogenesis. Consistently, we show that brain tissue from Alzheimer's disease patients contains reduced levels of plasmin, implying that plasmin downregulation may cause amyloid plaque deposition accompanying sporadic Alzheimer's disease. PMID- 11263500 TI - Frequent loss of 1p32 region but no mutation of the p18 tumor suppressor gene in meningiomas. AB - After chromosome 22 and NF2 inactivation, the loss of chromosome 1p is one of the most frequent abnormalities encountered in meningiomas. However the putative tumor suppressor gene located on 1p inactivated in meningiomas has still to be identified. We screened 68 meningiomas for LOH on chromosome 22 and 1. We found 34 LOH on the NF2 region on chromosome 22 (50%) and 19 LOH on 1p (28%), 16 being associated with loss of chromosome 22. Partial deletions delimited a candidate region located between D1S234 and D1S2797. The p18INK4C tumor suppressor gene, a member of the genes family coding for inhibitors of cyclin-dependent kinases, is located in this region. To determine whether p18 is involved in development of meningiomas, we performed a mutation analysis of the p18 gene and a search for homozygous deletion in the 19 meningiomas with 1p loss. Sequencing analysis of the p18 gene revealed one polymorphism, but no somatic mutations and no homozygous deletions were found. These results confirm that the loss of chromosome 1p32 is a frequent feature in meningiomas, however the p18 tumor suppressor gene which is located in this region, does not seem to be involved. PMID- 11263501 TI - Relationships between choline magnetic resonance spectroscopy, apparent diffusion coefficient and quantitative histopathology in human glioma. AB - This study sought to correlate quantitative presurgical proton magnetic resonance spectroscopic imaging (1H-MRSI) and diffusion imaging (DI) results with quantitative histopathological features of resected glioma tissue. The primary hypotheses were (1) glioma choline signal correlates with cell density, (2) glioma apparent diffusion coefficient (ADC) correlates inversely with cell density, (3) glioma choline signal correlates with cell proliferative index. Eighteen adult glioma patients were preoperatively imaged with 1H-MRSI and DI as part of clinically-indicated MRI evaluations. Cell density and proliferative index readings were made on surgical specimens obtained at surgery performed within 12 days of the radiologic scans. The resected tissue location was identified by comparing preoperative and postoperative MRI. The tumor to contralateral normalized choline signal ratio (nCho) and the ADC from resected tumor regions were measured from the preoperative imaging data. Counts of nuclei per high power field in 5-10 fields provided a quantitative measure of cell density. MIB-1 immunohistochemistry provided an index of the proportion of proliferating cells. There was a statistically significant inverse linear correlation between glioma ADC and cell density. There was also a statistically significant linear correlation between the glioma nCho and the cell density. The nCho measure did not significantly correlate with proliferative index. The results indicate that both ADC and spectroscopic choline measures are related to glioma cell density. Therefore they may prove useful for differentiating dense cellular neoplastic lesions from those that contain large proportions of acellular necrotic space. PMID- 11263502 TI - Drug resistance-associated factors in primary and secondary glioblastomas and their precursor tumors. AB - Malignant gliomas are largely resistant to current chemotherapeutic strategies often displaying a multidrug-resistant phenotype. Mechanisms involved in drug resistance are reduced cellular drug accumulation through membrane efflux pumps, drug detoxification as well as alterations in drug target specificity. In 27 primary and 17 secondary glioblastomas and their astrocytic precursor tumors, we studied the immunohistochemical expression profile of P-glycoprotein (P-gp), multidrug resistance-associated protein (MRP), lung resistance-related protein (LRP), metallothionein, and topoisomerase II alpha. Glial tumor cells in all glioblastomas showed constant up-regulation of LRP, MRP, and topoisomerase II alpha. P-gp was found in 90% of the primary and 60% of the secondary glioblastomas. In precursor tumors, these drug resistance-related factors were expressed in varying proportions. Metallothionein, also found in normal and activated astrocytes, was retained in all neoplastic phenotypes. Furthermore, metallothionein, P-gp, LRP, and topoisomerase II alpha were strongly expressed by normal and neoplastic vessels which may confer to impaired penetration of therapeutic agents through the blood-brain and blood-tumor barrier. However, the expression profiles of drug resistance-related proteins neither differed between primary and secondary glioblastomas nor revealed any correlation to precursor or recurrent tumors. Nevertheless, inhibition of these factors may be promising approaches to the management of malignant gliomas. PMID- 11263504 TI - Second line chemotherapy with docetaxel in patients with recurrent malignant glioma: a phase II study. AB - The objective of the study was to assess the efficacy of docetaxel in recurrent supratentorial malignant gliomas. The sample size of the study was determined by the Gehan's method for a response rate of 20% and a beta error of 5%. In the first step 14 patients (age 27-69, median 50; Karnofsky index 50-90, median 75) with recurrent malignant glioma after surgery, radiotherapy and nitrosourea, were enrolled (12 glioblastomas, 2 anaplastic astrocytomas). Docetaxel at the initial dose of 80 mg/m2 was administered every 3 weeks until progression or unacceptable toxicity. A total of 41 cycles was administered. Patients received a median of two cycles (range 1-6). No complete or partial response was observed. Therefore, according to the design of the study, no additional patients were enrolled and the trial was terminated. Two stabilizations were observed (14 and 15 weeks). Median TTP was 7 weeks (44 days). Median overall survival from recurrence was 26.5 weeks (6.4 months). Grade 3-4 neutropenia was observed in 8 patients (57%) but no life-threatening toxicity was observed. Other toxicities were uncommon and mild. Dose reduction was performed in 5 patients. This study suggests that docetaxel displayed no significant activity in patients with malignant recurrent gliomas. PMID- 11263503 TI - A case of spinal glioblastoma multiforme: immunohistochemical study and review of the literature. AB - A 31-year old female underwent subtotal resection of a spinal glioblastoma multiforme (GBM) at level D 10/11 in June 1997. Immunohistochemistry revealed increased MIB-1 labeling index and accumulation of p53 protein. Routine MRI in February 1998 showed multiple tumors of the lumbar spinal cord. At open biopsy, diffuse infiltration of multiple radices was seen. Histologically and immunohistochemically, the tumor was similar to the primary. In May 1998, MRI revealed multiple intracranial metastases and meningeal involvement. The patient died in June 1998, 13 months after the onset of symptoms. The lifes of patients with spinal gliomas are not endangered by direct compression of the brain stem, and systemic metastases are extremely uncommon with gliomas. Yet, survival times in the reported case and in the literature are not better than with cerebral localization. Analysis of the present case and a survey of the literature indicate that CSF involvement and consecutive intracranial seeding determine the prognosis of patients with spinal GBM. Thus, regular monitoring of CSF-cytology and/or spinal MRI appear to be advisable in spinal GBM. PMID- 11263505 TI - Cytogenetic profile of primary pituitary germinoma. AB - We present a case of a germinoma in the sellar region of a 10-year-old female patient who presented with a history of polydipsia, polyuria and visual disturbances. The tumor was resected and histologically analyzed. Interphase cytogenetics was performed using chromosome specific (peri)-centromeric DNA probes for all the somatic and X chromosomes on fresh frozen tissues. Fluorescent in situ cell hybridization demonstrated accumulated cytogenetic abnormalities involving significant alterations of chromosome 1, 4, 5/19 and 15. The child was treated postoperatively by radiation and now appears well with only minor neurological deficits. At 3-year follow-up no recurrent tumor mass could be demonstrated. PMID- 11263506 TI - The conditional survival statistics for survivors with primary supratentorial astrocytic tumors. AB - BACKGROUND: Relative survival rates can offer a general description of tumor outcome and, traditionally, are used for surveillance and comparison purposes. However, they are not informative for individual tumor survivors. Conditional survival estimates can calculate the probability of surviving next some years given survival to a specific period of time after craniotomy for individual tumor survivors. However, clinically, they have not been used for predicting the tumor outcome. METHODS: We calculated conditional probabilities of survival within 6 years after craniotomy in 112 patients with primary supratentorial astrocytic tumors and evaluated factors affecting the survival time more than 2 years after craniotomy. RESULTS: Our data showed that the conditional probability of survival can predict yearly survival rate when patients survive for a specific period of time. The conditional survival rates within 6 years after craniotomy were always higher than those evaluated by relative survival rates. Overall, the longer the patients survived, the higher the conditional probabilities of surviving sixth year postoperatively were. CONCLUSION: Our study demonstrates the conditional probabilities of survival have good availability and are important estimates for individual tumor survivors. PMID- 11263507 TI - Cerebral metastases as first symptom of cancer: a clinico-pathologic study. AB - Symptomatic brain metastases of carcinomas in patients without a previously diagnosed malignancy are frequent in neurosurgical series. Such tumors often lack distinctive morphological characteristics so that the routine histological examination can be unsuccessful in identifying the site of origin. Objectives of the present study were to evaluate the frequency of brain metastases as the only manifestation of an unknown primary cancer by the retrospective analysis of a series of consecutively operated single cerebral metastases; to verify the efficacy of clinical investigations in detecting the site of origin; to investigate whether the primary site can be identified by the immunohistochemical study of the neurosurgical specimens. Antibodies to the following antigens were used: carcinoembryonic antigen (CEA), carbohydrate antigen (CA) 19.9, CA 125, BCA 225, cytokeratin 20, PSA, HMB-45. Out of 181 patients operated for single cerebral metastasis of carcinoma, 99 (54.7%) were in patients without any previously diagnosed systemic neoplasm. In 26.7% the primary remained undiagnosed after clinical investigations, in 9 cases even at autopsy. PSA and HMB45 antibodies specifically identified metastases from prostate carcinomas and skin melanomas, respectively. No other specific immunophenotype was identified; the immunoreactivity of the single cases was more or less suggestive for a primary site. Precocious metastases of lung carcinomas expressed CEA more frequently than late metastases. It has been hypothesized that CEA plays some role as a contact mediating device. CEA expression can have some link with the tendency to metastasize precociously to the brain. No major difference of p53 and k-ras expression has been found in precocious versus late brain metastases. PMID- 11263508 TI - Psychophysiological evaluation of short-term neurotoxicity after prophylactic brain irradiation in patients with small cell lung cancer: a study of event related potentials. AB - BACKGROUND: The aim of this study was to show, whether a certain prophylacting applicable radiation affects the cognition, particularly, the specific cognitive components P50, N100, P300 and N400 of auditory event related potentials (ERPs) during a short memory test. METHODS AND MATERIALS: Eleven patients with small cell lung cancer (SCLC), who had presented complete response of disease after chemotherapy and radical radiotherapy in the lung, were prescribed to receive a prophylacting cranial irradiation (PCI) with a 6 MeV linear accelerator. The dose schedule was consisting of a total dose up to 30 Gy in 10 fractions, within 12 days (5 days a week). The psychophysiological approach before and after PCI was assessed by measurements of the auditory ERPs during a short memory performance using the digit-span Wechsler test. Components of ERP were recorded from 15 scalp electrodes. Additionally, symptomatology of depression and anxiety were assessed using Zung Self-Rating Depression Scale and Spielberger Anxiety Inventory, respectively, for pre- and post-PCI. RESULTS: No significant difference was noticed pre- and post-radiotherapy of all particular level of psychophysiological analysis concerning both the latencies and the amplitudes of ERPs auditory components P50, N100, P300 and N400 (P > 0.05, Wilcoxon signed test). Additionally, no changes were found with regard to behavioral performance (memory recall), depression symptomatology and state anxiety, according to pre- and post radiation measurements. However, the self-reported depression symptomatology showed that the patients presented moderate depression. CONCLUSION: No short-term psychophysiological neurotoxicity was detected with this PCI schedule using these instruments, lending additional support to evidence suggesting the benefit of this certain PCI schedule for patients with SCLC. PMID- 11263510 TI - Concomitant tuberculous and pyogenic brain abscess. PMID- 11263509 TI - Tuberculosis treatment: dangerous regimens? PMID- 11263511 TI - Pulmonary disease in HIV-infected African children. AB - Childhood human immunodeficiency virus (HIV) infection is common in most regions of sub-Saharan Africa. Acute and chronic respiratory diseases are major causes of morbidity and mortality in HIV-infected children. They represent a significant added burden in a region where diagnostic capabilities are limited and management decisions are often made on the basis of clinical guidelines alone. Pneumocystis carinii pneumonia is now recognised as an important cause of acute severe pneumonia and death in HIV-infected infants. However, there are few data on incidence and aetiology for more treatable conditions such as bacterial pneumonia. The association of pulmonary tuberculosis and HIV infection is uncertain, and the diagnosis is further confused by the presence of lymphoid interstitial pneumonitis and other chronic HIV-related pulmonary disease. This article reviews the literature and highlights the urgent need for further research in order to improve clinical management and appropriate interventions. PMID- 11263513 TI - Prevalence of drug-resistant tuberculosis in an HIV endemic area in northern Thailand. AB - SETTING: Chiang Rai Province in Northern Thailand, where human immunodeficiency virus (HIV) infection has been prevalent since the 1990s. OBJECTIVE: To observe the prevalence of drug-resistant tuberculosis (TB) and investigate the factors related to the level of drug resistance in an HIV endemic area. DESIGN: Population-based surveillance study covering the whole province. METHOD: Drug susceptibility testing was performed at the Thai Ministry of Public Health laboratory for all sputum smear-positive TB patients diagnosed in hospitals in Chiang Rai Province over a 25-month period in 1996-1998. Patient characteristics were obtained through interview by trained personnel. HIV testing was performed with informed consent. RESULTS: Among the 1077 incident patients without previous history of treatment, the proportion of patients with resistance to isoniazid was 13.2%, 10.8% to rifampicin, 15.6% to streptomycin, and 5.8% to ethambutol. Multidrug resistance (MDR), i.e., resistance to at least both isoniazid and rifampicin, was observed in 6.3%. Factors associated with primary MDR-TB were HIV positivity (OR 2.2, 95%CI 1.3-3.9), age <50 years (OR 2.0), and treatment in the provincial hospital (OR 2.3), compared to patients treated in the community and private hospitals. Stratified analysis shows a significantly high prevalence of primary MDR-TB among HIV-positive patients treated in the provincial hospital against HIV-negative patients or HIV-positive patients in other hospitals. CONCLUSION: The prevalence of primary MDR-TB in this area was high. It is necessary to strengthen TB control activities in order to reduce the burden of MDR-TB. PMID- 11263514 TI - Short-course instead of long-course chemotherapy for smear-negative patients in sub-Saharan Africa. AB - The use of short-course chemotherapy (SCC) in directly-observed treatment, short course (DOTS) programmes in sub-Saharan Africa was often restricted to patients with infectious and serious forms of tuberculosis, because of high costs of such regimens. With reduced drug prices and wide-scale substitution of thiacetazone by ethambutol in the continuation phase of treatment, various short-course regimens are now available at the same or even lower costs than long-course regimens. Several DOTS programmes are considering extending access to short-course chemotherapy to non-infectious patients, or have done so already. The authors provide an overview of the issues regarding the debate on the introduction of universal SCC in national tuberculosis control programmes in low-income countries in sub-Saharan Africa. They advise on a low-risk strategy to avoid the emergence of rifampicin resistance as a consequence of the wide availability of rifampicin associated with universal short-course, and strengthening of the health system to maintain high performance levels in diagnosis and treatment. PMID- 11263512 TI - Gender difference in delays to diagnosis and health care seeking behaviour in a rural area of Nepal. AB - SETTING: Directly observed treatment for tuberculosis using a short-course regimen (DOTS) was introduced in a rural area of Nepal. All new patients assigned to DOTS from mid-December 1997 to mid-June 1999 were eligible for the study. OBJECTIVE: To examine delays in tuberculosis (TB) diagnosis and compare health care seeking behaviour between men and women. DESIGN: A cross-sectional analysis of patient interviews. RESULTS: Women were found to have a significantly longer total delay before diagnosis of tuberculosis (median 2.3 months for men, 3.3 months for women). When they visited traditional healers first, women had a significantly longer delay than men from the first visit to health care providers to diagnosis (median 1.5 months for men, 3.0 months for women). More women (35%) visited traditional healers before diagnosis than men (18%), and were more likely to receive more complicated charms from traditional healers. Men tended to visit the government medical establishment first if they knew that free TB treatment was available, but women did not. CONCLUSION: Women were more likely to visit and to believe in traditional healers; this might lead to the longer delays experienced before TB diagnosis. PMID- 11263515 TI - Low rate of emergence of drug resistance in sputum positive patients treated with short course chemotherapy. AB - SETTING: Tuberculosis Research Centre, Chennai. OBJECTIVE: To study the emergence of drug resistance during treatment and relapse among sputum positive pulmonary tuberculosis patients treated with short-course chemotherapy regimens. DESIGN: Retrospective analysis of randomised clinical trials using the following regimens: 2HRZE7/6HE7, 2HRZE2/4HRE2, 2HRZE3/4HR2 and 3HRZE3/3HR2. Emergence of resistance was analysed in patients with unfavourable response/relapse based on culture and susceptibility reports. RESULTS: Of 1817 patients studied, 1435 (79%) had susceptible strains prior to treatment; 2% of these had an unfavourable response, 7% relapsed and 1% had emergence of resistance to isoniazid, rifampicin, or both. In 320 patients with initial isoniazid resistance, 19% had an unfavourable response and 13% relapsed, while resistance to rifampicin emerged in 11%. Treatment outcomes were similar whether patients received three or two drugs in the continuation phase. Data on resistance to ethambutol and pyrazinamide were not available. CONCLUSION: In this study, the overall emergence of resistance to rifampicin occurred in only 2% of patients, despite the high level (18%) of initial resistance to isoniazid. Thus, standardised short-course treatment carries only a minimal risk of emergence of rifampicin resistance. PMID- 11263517 TI - Drug-resistant tuberculosis in foreign-born persons from Mexico, the Philippines, and Vietnam-United States, 1993-1997. AB - SETTING: Foreign-born persons in the United States represent a growing proportion of the nation's tuberculosis (TB) cases. OBJECTIVE: To characterize drug resistance patterns in foreign-born TB patients from the three most common birth countries. DESIGN: A descriptive analysis of national TB surveillance data for 1993-1997. TB case reports for foreign-born persons who were at least 15 years old and born either in Mexico (6221), the Philippines (3624), or Vietnam (3351) were included. RESULTS: Among those with no prior history of TB, the proportions with isoniazid-resistant TB and MDR-TB (resistance to at least isoniazid and rifampin) were 9.2% and 1.6% for persons from Mexico, 13.7% and 1.4% for those from the Philippines, and 17.8% and 1.4% for those from Vietnam. Levels of isoniazid resistance and MDR-TB did not change during the 5-year study period. Levels of isoniazid resistance decreased with older age for persons with no prior TB from all three countries; however, rates of MDR-TB did not vary with age. Persons with <1 year of residence in the US were more likely to have MDR-TB; however, duration of residence in the US was not associated with isoniazid resistance. CONCLUSION: Increased drug resistance in younger and more recent arrivals suggests that vigorous efforts to prevent further development of MDR-TB in the three countries are essential. PMID- 11263516 TI - Resistance of Mycobacterium tuberculosis to four first-line anti-tuberculosis drugs in Japan, 1997. AB - SETTING: Five years after the last survey of drug-resistant tuberculosis in Japan, a nationwide survey was conducted by the Tuberculosis Research Committee. OBJECTIVE: To determine the prevalence of and risk factors for resistance to four first-line anti-tuberculosis drugs. DESIGN: Cultures were obtained from patients hospitalized at 78 hospitals in different districts of Japan throughout a 6-month period, 1 June-30 November 1997. Drug susceptibility testing was carried out at the Research Institute of Tuberculosis, Tokyo, one of the supranational reference laboratories of the WHO/IUATLD global project. RESULTS AND CONCLUSION: Among patients with no prior treatment, resistance to any of the four drugs was found in 10.3%, and the prevalence of primary multidrug resistance (MDR) was 0.8%. The prevalence of acquired resistance was 42.4% for any of the four drugs and 19.7% for MDR, indicating a high prevalence rate compared with those reported in the WHO/IUATLD global project. About 73% of resistant isolates from new cases were resistant to one drug, while 64.3% of resistant isolates from the re-treatment cases were resistant to two or more drugs (P < 0.0001). No significant differences in resistance rates by sex, age group, nationality, district, and/or accompanying diseases were observed in any of the new or re-treatment cases. Other factors associated with the high prevalence in re-treatment cases remain to be determined. PMID- 11263518 TI - Outbreak of multidrug-resistant tuberculosis at a methadone treatment program. AB - SETTING: An out-patient methadone treatment program MTP). OBJECTIVE: To investigate transmission of multidrug-resistant tuberculosis (MDR-TB) in the MTP. DESIGN: Cases were defined as MTP clients or staff who developed TB between 1 January 1994 and 1 January 1996, with at least one positive culture for Mycobacterium tuberculosis resistant to isoniazid and rifampin. Contacts were identified, located and evaluated. RESULTS: Thirteen cases of MDR-TB occurred among 462 clients and staff. One fifth (6/30) of the members of a counseling group for human immunodeficiency virus (HIV) infected clients developed MDR-TB. Individuals known to be HIV positive were at greater risk for TB than those who were HIV negative (RR 5.2, 95%CI 1.2-22.7). Of 449 clients and staff identified as contacts, 393 (87.5%) were located and screened. Among those with a negative baseline tuberculin skin test, 18.5% (56/303) were skin test converters. Attendance at the MTP during a period when the index case was infectious was associated with an increased risk of conversion (RR 2.5, 95%CI 1.1-6.0). CONCLUSION: Extensive transmission of MDR-TB occurred at an out-patient MTP serving numerous clients with HIV infection. This outbreak underscores the importance of developing effective strategies to prevent TB transmission in this setting. PMID- 11263519 TI - The management of anti-tuberculosis drug-induced hepatotoxicity. AB - SETTING: A tuberculosis ward in a chest disease teaching hospital. OBJECTIVE: To compare the efficacy of two different retreatment protocols on hepatotoxicity recurrence in tuberculosis treatment. DESIGN: In a prospective, randomised study, 45 patients with new tuberculosis developed hepatotoxicity after anti tuberculosis treatment. Patients in Group I (n = 20) were retreated with a drug regimen consisting of isoniazid, rifampicin, ethambutol and streptomycin administered by gradually increasing the number and dosage of the drugs. Patients in Group II (n = 25) were retreated with the same regimen (isoniazid, rifampicin, pyrazinamide and ethambutol) in the same dosages throughout. RESULTS: Hepatotoxicity recurred in respectively zero and six (24%) patients in Groups I and II (P = 0.021). Of the six patients with recurrence of hepatitis, one could not be followed up. The other five received the same retreatment protocol as Group I. By the end of retreatment, all patients were cured. CONCLUSION: The recurrence rate of hepatotoxicity in the retreatment of tuberculosis is higher in the reintroduction of a full-dose regimen including pyrazinamide, which causes more hepatotoxicity than gradual reintroduction of a regimen without pyrazinamide. PMID- 11263520 TI - Non-linear pharmacokinetics of rifampicin in healthy Asian Indian volunteers. AB - OBJECTIVE: To characterise the pharmacokinetics of rifampicin (RMP) in healthy Asian Indian volunteers after oral administration of commercially marketed reference formulations. DESIGN: Two separate studies were conducted. In Study 1, 12 volunteers were administered a single 450 mg sugar-coated tablet, and in Study 2, 11 volunteers were administered a 30 ml suspension (100 mg/5 ml) equivalent to a 600 mg dose of RMP. Blood samples were collected at 0 hours and then at 0.25, 0.5, 0.75, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 10 and 12 hours post dose, and plasma concentrations were assayed by high performance liquid chromatography. RESULTS: Non-compartmental analysis indicated that the mean (coefficient of variation [%CV]) values for the tablet for Cmax, Tmax and AUC(0-infinity) were 8.59 (53.66) microg/ml, 1.58 h and 49.54 (61.79) microg x h/ml, respectively. The corresponding mean (%CV) values for the suspension were 14.76 (24.14) microg/ml, 1.45 h and 119.12 (28.24) microg x h/ml. Two-compartment analysis indicated that the mean (%CV) values for Cl/F, beta and t(1/2beta) were respectively 197.51 (46.58) (ml/h)/kg, 0.2153 (32.01) h(-1) and 3.57 (34.85) h, at the 450 mg dose (tablet), and were significantly different from the respective 94.76 (33.96) (ml/h)/kg, 0.1210 (33.66) h(-1) and 5.44 (16.69) h at the 600 mg dose (suspension) (P < 0.05). In addition, there was a significant linear correlation (r = 0.60, P < 0.005, n = 21) between Cmax and the elimination half-life, indicating a concentration-dependent increase in half-life. CONCLUSION: Rifampicin obeys two-compartment kinetics with zero order absorption, and exhibits non-linear saturable elimination kinetics as well as large inter individual variability. PMID- 11263521 TI - Utility of blood cultures and incidence of mycobacteremia in patients with suspected tuberculosis in a South African infectious disease referral hospital. AB - SETTING: A 500-bed government referral institution for patients with tuberculosis and other infectious diseases in Gauteng, South Africa. OBJECTIVES: To assess the usefulness of BACTEC blood cultures over and above that of other microbiological methods for the diagnosis of tuberculosis in patients who are suspected of suffering from tuberculosis. DESIGN: Mycobacterial blood cultures were obtained from patients presenting with symptoms suspicious of tuberculosis and where there was no clinical evidence of other infectious etiologies, and from patients who had failed tuberculosis treatment. RESULTS: Sixteen (22%) of 71 patients included in the study were positive for Mycobacterium tuberculosis on blood culture, while seven (10%) were positive for M. avium complex (MAC). Twelve (75%) of the patients with tuberculosis and positive blood cultures were however also positive for acid-fast bacilli on sputum smears and eight (50%) were initially diagnosed clinically and radiographically as localized pulmonary tuberculosis. Blood cultures positive for mycobacteria were only found among patients with human immunodeficiency virus infection (HIV). CONCLUSIONS: Bacteremia with M. tuberculosis complex was detected in HIV-infected patients with suspected tuberculosis, even in patients presenting with localized pulmonary infection on initial clinical assessment. Among patients with suspected tuberculosis, blood cultures were useful in diagnosing unsuspected MAC disease, but did not add to the diagnostic yield of conventional tests for tuberculosis used routinely, namely sputum microscopy and culture, or occasional biopsy specimens. PMID- 11263522 TI - Isolation of Mycobacterium bovis from human cases of cervical adenitis in Tanzania: a cause for concern? AB - SETTING: Pastoralist communities in the Northern and Southern zones of Tanzania. DESIGN: Observational study. OBJECTIVES: To determine the involvement of Mycobacterium bovis in tuberculosis cases presenting at tuberculosis (TB) clinics in rural areas in these zones. METHODS: A total of 149 tuberculosis cases identified on the bases of clinical manifestation were sampled. Appropriate specimens were cultured on two Loweinstein Jensen slants with respectively glycerol and pyruvate added. Forty-one isolates were cultured and subjected to biochemical typing. RESULTS: Overall, 31 (70.5%) of the mycobacterial isolates recovered from all forms of tuberculosis were identified as M. tuberculosis, seven (16.0%) were identified as M. bovis, and six (13.6%) were other mycobacterial species. There was a significantly higher isolation rate (P < 0.05) of M. bovis among strains recovered from extra-pulmonary (26.8%) than pulmonary tuberculosis samples (4.3%). CONCLUSION: Based on these findings, it is imperative that M. bovis be considered as a pathogen of concern to people living in rural areas of Tanzania. Further work is required to establish a zoonotic link between cattle and the people in these communities who rear them. PMID- 11263523 TI - High rates of tuberculosis infection among children from Ciutat Vella District, Barcelona, 1996-1997. AB - With tuberculosis (TB) rates of over 160/100,000 in 1997, Ciutat Vella District, Barcelona, is the main community focus of TB in the city. For the purpose of TB screening, 415 children >2 years old from that district received a tuberculin skin test (TST); 27 (6.6%) (95%CI 4.5-9.3) were found to be infected but disease free. The frequency of a positive TST increased significantly with age, from 0% in the 2-4 year age group to 14.6% in 10-14 year olds. Three culture-positive source adults, two of them sputum smear-positive, who were not previously known were traced from six TST-positive children. Previous BCG vaccination was not associated with a positive TST. These data support the use of universal TST screening in children living in Ciutat Vella District, Barcelona, as a means of identifying and treating TB cases. PMID- 11263525 TI - Risk factors for hepatotoxicity during anti-tuberculosis chemotherapy in Asian populations. PMID- 11263524 TI - A spoonful of sugar...: improving adherence to tuberculosis treatment using financial incentives. AB - OBJECTIVE: To determine whether incentives increase adherence to directly observed therapy (DOT) for tuberculosis (TB) treatment. METHODS: The TB program gave a five-dollar grocery coupon for each DOT appointment kept to 55 patients who had missed at least 25% of DOT doses over a 4-week period. Treatment completion rates were compared with an historic control group of 52 patients who began treatment a year earlier, who would have been eligible for incentives but did not receive them. RESULTS: Incentive program patients were more likely than control patients to complete therapy within 32 weeks (OR 5.73, 95%CI 2.25-14.84) and 52 weeks (OR 7.29, 95%CI 2.45-22.73). CONCLUSION: Patient incentives can increase adherence to DOT in TB programs. PMID- 11263526 TI - Highly efficient transduction of endothelial cells by targeted artificial virus like particles. AB - Targeting the tumor vasculature by gene therapy is a potentially powerful approach, but suitable vectors have not yet been described. We have designed a new type of liposomal vector, based on the composition of anionic retroviral envelopes, that is serum-resistant and nontoxic. These artificial virus-like envelopes (AVEs) were endowed with a cyclic RGD-containing peptide as a targeting device for the a(v)beta3-integrin on tumor endothelial cells (ECs). The packaging of plasmid DNA complexed with low-molecular-weight, nonlinear polyethyleneimine into these AVEs yielded artificial virus-like particles (AVPs) that transduced ECs with efficiencies of up to 99%. In contrast, transduction of a variety of other cell types by these RGD-AVPs was comparably inefficient under the same experimental conditions. This EC selectivity was mediated, in part, but not exclusively, by the RGD ligand, as suggested by the reduced, but still relatively high, transduction efficiency seen with AVPs lacking RGD. The interaction of anionic lipids of the AVPs with ECs may therefore contribute to the observed selective and highly efficient transduction of this cell type. These findings suggest that the targeted AVE technology is a useful approach to create highly efficient nonviral vectors. PMID- 11263527 TI - B7.1 expression eliminates tumor resistance to IL-12 gene therapy. AB - IL-12 gene therapy results in tumor regression in some, but not all, murine models. We hypothesized that expression of B7.1 on the tumor cell surface was necessary for IL-12-mediated tumor regression. In addition, we hypothesized that all cells must express B7.1 for this to be effective. To evaluate this hypothesis, tumor nodules were established in mice with either wild-type B16 melanoma or with B16 melanoma modified to express B7.1. IL-12 cDNA was transferred to the tumor by particle-mediated gene transfer. All tumors modified to express B7.1 regressed completely after IL-12 cDNA treatment. When the percent of B7.1-transfected B16 cells was decreased to 50%, no animals survived after treatment. Animals rendered tumor-free were then challenged with wild-type B16. Fifty percent of mice was protected from this tumor challenge. Expression of CD28 (the stimulatory B7.1 ligand) was significantly increased in both CD8(+) T cells and natural killer cell populations of mice rejecting tumor challenge compared to mice with tumor growth. These results suggest that the costimulatory molecule B7.1 is required for initial tumor sensitivity to IL-12 gene therapy and that protection from subsequent challenge with B7.1 (-) tumor is mediated by CD28(+) immune effector cells. PMID- 11263528 TI - Immune-dependent distant bystander effect after adenovirus-mediated suicide gene transfer in a rat model of liver colorectal metastasis. AB - Gene transfer of the herpes simplex virus thymidine kinase (HSV-tk) gene sensitizes tumor cells to the toxic effect of ganciclovir (GCV). The toxic effect of GCV extends to nontransduced surrounding cells by a metabolic process known as the bystander effect. A distant bystander effect, which involves anatomically separated tumors, has been reported in vivo. Our aim was to evaluate and characterize such distant effect in a rat model of colorectal tumors implanted in the liver using adenovirus to carry the HSV-tk gene. Two colorectal tumors were implanted in two distinct liver lobes of the liver. One of the tumor was transduced with an adenoviral vector containing HSV-tk gene. The volumes of the tumors were monitored after GCV treatment. Implication of the immune system was studied histologically and after in vivo manipulations. After GCV administration, the nontransduced distant tumor regressed partially or completely in the experimental group. Immunohistochemical analysis revealed the presence of CD8+ lymphocytes in the distant lesion. HSV-tk/GCV-induced immune response against tumors was evidenced by an adoptive transfer assay (Winn assay) and the distant bystander effect was blunted after CD8+ lymphocytes depletion. However, the survival rates for treated animals were not improved. These findings demonstrate that an immune-mediated effective distant bystander effect can be obtained after limited adenoviral-mediated transfer of the HSV-tk gene. PMID- 11263529 TI - Human prostate carcinoma cells as targets for herpes simplex virus thymidine kinase-mediated suicide gene therapy. AB - To evaluate human prostate carcinoma cells as targets for herpes simplex virus thymidine kinase (HSV-TK) -mediated gene therapy, we tested the utility of different viral vectors on three human cell lines DU-145, LNCaP, and PC-3. Our viral vectors were carrying a fusion gene of HSV-TK and green fluorescent protein for accurate determination of the gene transfer rate and its contribution to the treatment outcome in each case. We observed that adenoviral and lentiviral vectors were efficient vehicles for all the cell lines, whereas Semliki Forest virus and Sindbis virus vectors yielded only a few percent of transgene-positive cells. Despite sufficient gene transfer rates (25-45%) in the ganciclovir (GCV) sensitivity experiment, only DU-145 cells were efficiently destroyed under clinically relevant GCV concentrations. This was shown to be due to low level of "bystander effect" in PC-3 and LNCaP cells. Our data demonstrate that human prostate tumors can be good targets for adenovirus- or lentivirus-mediated HSV TK/GCV gene therapy, but each tumor should be investigated for gene transfer rate and bystander effect to warrant a sufficient treatment result. PMID- 11263530 TI - Canarypox virus expressing wild type p53 for gene therapy in murine tumors mutated in p53. AB - The antitumor activity of a recombinant canarypox virus expressing wild type murine p53 (ALVAC-p53) was investigated in two murine syngeneic tumors harboring an endogenous p53 mutation (CMS4 and TS/A). Direct intratumor injections of ALVAC p53 in CMS4 pre-established subcutaneous tumors induced total tumor regression in 66% of mice. Furthermore, 100% of the cured mice was protected against a contralateral subsequent challenge with the parental tumor cells. The intravenous treatment of experimental lung metastasis by ALVAC-p53 also induced significant tumor growth inhibition in both models. The antitumor effect of ALVAC-p53 was only observed in immunocompetent animals and was associated with the generation of a specific antitumor immune response. ALVAC-p53 induced the expression of a functional p53 wild type protein as demonstrated by up-regulation of p21waf1 and induction of apoptosis. A vaccine strategy using intravenous or subcutaneous ALVAC-p53/NYVAC-p53 prime boost protocol failed to induce CTL against p53 wild type used as target tumor antigen, and failed to protect mice against challenge with the mutated tumor cells. The mechanism of the curative and protective effects observed after direct intratumor injections results from the induction of a specific antitumor response directed against other antigens than p53. Our results suggest that the local induction of tumor apoptosis, combined with the adjuvant effect of ALVAC vector, enhances the immunogenicity of the intratumor environment and allows induction of specific antitumor immune response. PMID- 11263531 TI - Gamma-rays enhance rAAV-mediated transgene expression and cytocidal effect of AAV HSVtk/ganciclovir on cancer cells. AB - Adeno-associated virus (AAV) vector has several unique properties suited for gene therapy applications. However, relatively low efficiency of transgene expression, which is mainly due to a limited second-strand synthesis from the single-stranded AAV genome, can be a problem in some applications that require potent gene expression such as antitumor applications. Recently, gamma-ray irradiation has been reported to enhance the second-strand synthesis of the AAV genome, and consequently transgene expression. We demonstrate here that an AAV vector harboring the herpes simplex virus type-1 thymidine kinase (HSVtk) is able to kill cancer cells more efficiently when used in combination with gamma-ray irradiation. A human maxillary sinus cancer cell line, NKO-1, was efficiently killed in combination with HSVtk transduction and ganciclovir (GCV), as expected. More importantly, gamma-ray irradiation of practical dosages augmented the cytocidal effect of the HSVtk/GCV system. Southern analysis indicated that gamma rays enhanced the double-strand synthesis of the rAAV genome in NKO-1 cells. These findings suggest that the combination of rAAVtk/GCV suicide gene therapy with radiotherapy has synergistic effects in the treatment of cancers and may lead to a reduction of the potential toxicity of both rAAVtk/GCV and gamma-ray irradiation. PMID- 11263533 TI - The genetic revolution in artificial reproduction: a view of the future. AB - With the completion of the human genome project, micro-array technology offers the potential to open up a whole new vista in assisted reproduction. In the next 10-20 years we will be able to screen each human embryo for all numerical chromosomal abnormalities as well as many genetic diseases. Micro-array analysis may permit the screening of multiple alleles for monogenetic diseases and polygenic diseases, including diabetes, hypertension and schizophrenia. In the near future, it may be possible to assess an individual's genetic predisposition for cardiovascular disease, all types of cancer and infectious diseases. In the distant future, it may even be possible to screen for any genetic trait, e.g. stature, baldness, obesity, hair colour, skin colour or even IQ. Although it is still uncertain what molecular genetic tools may be available, we can be sure that some of these trends will have major consequences on the future of assisted reproduction and society at large. PMID- 11263532 TI - Possible factors affecting the development of oocytes in in-vitro maturation. AB - To date, pregnancy rates from oocytes matured in-vitro (IVM) have been much lower than those with in-vivo stimulated maturation. In order to improve the developmental potential of IVM oocytes, we studied the effect of three possible factors on pregnancy rates: (i) priming in vivo with FSH before aspiration; (ii) the time interval of maturation in vitro, and (iii) timing the aspiration by monitoring the serum concentrations of oestradiol and inhibin A. In all experiments, oocyte retrieval was performed transvaginally and oocytes were matured individually in culture medium (TCM 199) under oil. Intracytoplasmic sperm injection (ICSI) was carried out on all metaphase II oocytes. Suitable embryos (maximum of two) were replaced after culturing for 2-3 days in IVF medium. Endometrial priming consisted of 2 mg 17beta-oestradiol, taken orally three times a day from the day of oocyte retrieval, and intravaginal progesterone suppositories initiated 2 days later. In the first experiment, 20 women were randomly allocated to two groups: group I (n = 10 cycles) received no stimulation, while group II (n = 10 cycles) received recombinant FSH 150 IU/day for 3 days, initiated on day 3. FSH priming did not affect the rates of maturation, fertilization or cleavage, and no effect was seen on embryo development. The second experiment included 48 patients undergoing 55 unstimulated cycles. The effects of IVM periods of 28 and 36 h were compared. Shortening the IVM period did not compromise subsequent embryo development. The third study analysed the results of maturation of oocytes obtained in 87 cycles in 75 unstimulated normal women, after a leading follicle of 10 mm in diameter and an endometrial thickness of at least 5 mm were observed. A pregnancy rate of 12.6% (11/87) per aspiration and 17.4% (11/63) per transfer was obtained. Serum concentrations of oestradiol and inhibin A were evaluated retrospectively. Significantly more pregnancies were obtained in cycles with a detected increase in the concentration of oestradiol from day 3 to the day of aspiration (19% per aspiration) compared with cycles without such an increase (0% per aspiration). A higher pregnancy rate was observed after an increase in inhibin A concentration (24 versus 0%). In conclusion, monitoring the timing of aspiration may potentially improve the developmental potential of immature oocytes. FSH priming did not affect either cleavage rate or embryo development. Shortening the maturation period from 36 to 28 h did not compromise subsequent embryonic development. PMID- 11263534 TI - Cryopreservation of embryos and oocytes: obstetric outcome and health in children. AB - This paper provides an overview of the effects of cryopreservation on obstetric outcome and child development. Cryopreservation of embryos has no apparent negative impact on perinatal outcome and early infant development. The available data do not indicate an elevated congenital malformation rate. Intracytoplasmic sperm injection and cryopreservation is comparable with conventional in-vitro fertilization and cryopreservation. It remains unclear if embryo freezing poses long-term risks to children so conceived. There are several potential advantages of oocyte freezing which, however, is still a research procedure. PMID- 11263535 TI - Aspects of natural cold tolerance in ectothermic animals. AB - Polar, alpine and temperate ectothermic (cold-blooded) animals encounter temperatures below the melting point of their body fluids either diurnally or seasonally. These animals have developed a number of biochemical and physiological adaptations to survive the low temperatures. The problems posed to the animals during cold periods include changes in membrane and protein structure due to phase changes in these molecules, changes in electrolyte concentrations and other solutes in the body fluids as well as changes in metabolism. Cold tolerant ectothermic animals can be divided into two groups depending which of two 'strategies' they employ to survive the low temperatures: freeze-tolerant animals which survive ice formation in the tissues and freeze-avoiding animals which tolerate the low temperatures but not crystallization of the body fluids. The adaptations are mainly directed towards the control or avoidance of ice formation and include the synthesis of low mol. wt cryoprotectants, ice nucleating agents and antifreeze proteins. However, some of the adaptations such as the synthesis of low mol. wt cryoprotectants are also more specific in their mechanism, e.g. direct stabilizing interaction with membranes and proteins. The mechanisms employed by such animals may offer ideas and information on alternative approaches which might be usefully employed in the cryopreservation of cells and tissues frequently required in assisted reproductive technology. PMID- 11263536 TI - Human oocyte maturation in vitro is stimulated by meiosis-activating sterol. AB - In-vitro studies in mouse oocytes have shown that the C-29 endogenously occurring sterol FF-MAS (follicular fluid meiosis-activating sterol) is a potent inducer of meiotic maturation leading to increased fertilization rates. We have used synthetic FF-MAS to induce meiotic maturation in immature human oocytes aspirated from polycystic ovarian syndrome patients. The patients were asked to give written consent to donate half of their aspirated oocytes to investigate the influence of culture conditions on maturation kinetics. The oocytes were aspirated from follicles 8-12 mm in diameter under ultrasound guidance after initial treatment with a gonadotrophin-releasing hormone agonist and s.c. injections of recombinant FSH for 3 days. The other half of the oocytes remained outside this present study. They were reserved for the patients' benefit and were fertilized with appropriate embryo stages being transferred. Fertilization and transfer were not attempted for the study oocytes. Synthetic sterol FF-MAS was added to the culture media at a concentration of 20 micromol/l and nuclear maturation was compared to a control group of oocytes cultured in media only supplemented with vehicle (TCM-199 supplemented with 0.2% ethanol v/v); thus no additional hormones, growth factors, serum or follicle fluid were added. In 31 cycles, oocytes were randomly allocated to one of seven treatment groups: fixed immediately upon aspiration (0 h group) or after in-vitro maturation culture in the presence or absence of FF-MAS for 22, 30 or 40 h respectively. A total of 81 oocytes were processed for light microscopy. The optimal timing of maturation was observed following 30 h of in-vitro culture, when 67% of FF-MAS-treated oocytes had completed nuclear maturation to the metaphase-II stage compared to 29% in the control group. The maturation time of 30 h appeared significantly superior to both 22 and 40 h, but only in the presence of FF-MAS. Cumulus expansion was most profound in the FF-MAS group after 30 h whereas all oocytes had shed the cumulus investment after 40 h. Our observations indicate that FF-MAS positively influences the absolute frequency and the kinetics of human oocytes undergoing nuclear maturation. PMID- 11263537 TI - In-vitro production of cattle embryos: problems with pregnancies and parturition. AB - Using complex media containing serum and somatic cells, pregnancy rates following transfer of single, day 7 advanced-stage blastocysts approached 60%, while pregnancy rates of morulae or day 8 blastocysts were substantially lower. Pregnancies resulting from in-vitro-derived embryos were characterized by the following features: sex ratio skewed in favour of males; increased spontaneous abortion rate throughout gestation; reduced intensity of labour in recipients; and increases in birth weights, dystocias, calf mortality, and fetal abnormalities. In an attempt to improve the normality of pregnancies, a field trial was conducted within the commercial in-vitro programme at Em Tran. Following fertilization in vitro, equal numbers of zygotes were put into a Menezo's B2-buffalo rat liver cell (B2-BRL) coculture system with (S+) or without (S-) 10% serum for the first 72 h of in-vitro culture. On day 4, all embryos were moved to fresh medium and cultured to day 7 in B2-BRL with serum. The efficiency of blastocyst production from oocytes (19.6 versus 17.8%) and the pregnancy rate at 60 days (47.8 versus 47.1%) did not differ between S+ and S-. Likewise, there was no difference between S+ and S- in the percentage of male calves (53.9 versus 54.3%), abortions (13.1 versus 11.9%), percentage of live calves (78.8 versus 81.4%), or congenital abnormalities (4.3 versus 3.3%). PMID- 11263538 TI - Bovine embryo culture in vitro: new developments and post-transfer consequences. AB - The past decade has seen a significant shift away from co-culture systems for cattle blastocyst production. In particular, recent adoption of sequential media systems has increased performance. However, wholly defined systems, such as the replacement of albumin with nonbiological macromolecules, fail to reproduce the nutritive role that this molecule has during development. Cattle blastocysts developed in protein-free medium are metabolically compromised. A further new concept is the use of metabolic inhibitors to stimulate embryo development in vitro. Non-toxic levels of NaN3, 2,4-dinitrophenol or very low oxygen atmospheres (approximately 2%) significantly increase both the yield (by approximately 10 20%) and the quality of blastocysts when these treatments are applied during the peri-compaction period in vitro. Nevertheless, there are also negative consequences of cattle embryo culture, such as fetal oversize and/or significant post-day 35 fetal loss. We have recently found that much of this loss is due to failure of normal allantoic development within the conceptus. Early fetal development is supported by vascularization within the yolk sac, but from day 35 to day 110, loss occurs through poor nutrient supply and an inability to remove nitrogenous wastes, leading to fetal death around day 35. The cause of disrupted allantois development has not been identified as yet, but may share a common 'cause-effect' mechanism with the fetal oversize syndrome. PMID- 11263539 TI - In-utero overgrowth in ruminants following embryo culture: lessons from mice and a warning to men. AB - Unusually large offspring have been born in ruminants following the transfer to recipients of embryos that have either been subjected to some form of manipulation, e.g. nuclear transfer, or have been exposed to an unusual in-vivo or in-vitro environment. Overgrowth syndromes have been reported in other species, including humans and mice, but these have arisen from chromosomal abnormalities and spontaneous or experimentally induced genetic mutations. Overgrowth phenotypes across the species, however, exhibit many common features, including alterations in organ and tissue development, and placental anomalies. Our current working hypothesis is that the causative agent(s) alter(s) the expression of a gene or genes associated with growth and development. Imprinted genes have been implicated in this syndrome because: (i) similar phenotypes are observed in both humans and mice when the expression of such genes has been altered; and (ii) they may be more vulnerable to epigenetic modification during the period (oocyte to blastocyst) when embryos are cultured in vitro. Evidence supporting this hypothesis is reviewed and the implications for assisted reproduction in humans discussed. PMID- 11263540 TI - Risks of in-vitro production of cattle and swine embryos: aberrations in chromosome numbers, ribosomal RNA gene activation and perinatal physiology. AB - In cattle, in-vitro production (IVP) of embryos has become a standardized technique; however, increased frequencies of calving problems and larger calves have been reported. In swine, IVP has resulted in only a limited number of piglets. In this paper we present information on cattle and swine embryos produced in vitro by oocyte maturation, fertilization and further embryo culture to the blastocyst stage in vitro. Control in-vivo developed embryos were collected after superovulation. The cattle embryos were processed for fluorescence in-situ hybridization (FISH) with two chromosome-specific probes to detect numerical chromosome aberrations. The swine embryos were processed for transmission electron microscopy and immunocytochemistry with an antibody against RNA polymerase I [essential for ribosomal RNA (rRNA) gene transcription] in order to highlight the post-fertilization development of the nucleolus as a marker for rRNA gene activation. The FISH analyses of the cattle embryos revealed that 72% of IVP blastocysts were mixoploid, i.e. contained both diploid and polyploid cells, versus 25% in vivo. Chromosome abnormalities were observed from the 2-cell stage onwards. The immunocytochemical analyses of the swine embryos revealed that during in-vivo development, RNA polymerase I became localized to multiple foci in the developing nucleoli late during the 4-cell stage. This focal localization of RNA polymerase I was not observed in IVP embryos. In conclusion, IVP embryos may display aberrations in chromosome numbers and rRNA gene activation. The significance of these deviations for fetal and perinatal viability, however, remains unknown. The survival of most calves derived from IVP indicates that a considerable number of these embryos are able to compensate for the adverse effects of the in-vitro procedures. PMID- 11263541 TI - The future of male infertility management and assisted reproduction technology. AB - Intracytoplasmic sperm injection (ICSI) is undoubtedly a powerful, and sometimes the only effective, form of infertility treatment. Nonetheless, it is a non specific treatment that, combined with increasingly heroic techniques to recover male germinal cells, has led to perceptions of men as just providers of gametes in the infertility equation. In response to this nihilist attitude, where women are investigated extensively and scant attention is paid to men, there is a re emerging awareness of andrology--particularly in countries with limited healthcare resources. Structured management strategies, using diagnostic information to recognize causative factors amenable to simpler, even systemic, therapies with reasonable chances of pregnancy rather than resorting prematurely to assisted reproduction technology, represent rational, cost-effective approaches to infertility management. Furthermore, genetic testing (particularly cystic fibrosis gene defects and Y-chromosome microdeletions) is essential for couples to make fully informed decisions on their options. Recognition that free radical-induced damage to the sperm genome (e.g. from smoking or in-vitro sperm manipulation) underlies deleterious paternal effects on preimplantation development promotes further synergy between andrology and embryology. Although societies strike different balances between considerations of affordability and cost-effectiveness of assisted reproduction technology, ICSI represents a last resort, to be used when less-invasive, lower-cost treatments have been deemed inappropriate or have failed. Consequently, rather than assisted reproduction technology eliminating the need for andrology, the future will see increasingly tighter integration of multidisciplinary infertility care, embracing careful diagnosis and patient education before obtaining truly informed consent and embarking upon cost-effective treatment. PMID- 11263542 TI - Malformations reported in chorionic villus sampling exposed children: a review and analytic synthesis of the literature. AB - PURPOSE: To determine whether the frequency of vascular disruption defects, other than limb defects, is increased in reports of chorionic villus sampling (CVS) exposed children compared with an unexposed population. METHODS: Only studies that reported the total number of CVS-exposed pregnancies and details of pregnancy outcome, including all the malformations, were included. Twenty-five articles met these criteria. RESULTS: The frequencies of gastroschisis, intestinal atresias, and clubfoot were significantly increased among the CVS exposed infants as compared with the baseline unexposed population. The frequencies of other vascular disruption defects, including Poland sequence, amniotic band sequence, and cleft lip/cleft palate, were not increased. CONCLUSION: CVS-exposed children have an increased frequency of intestinal atresia, gastroschisis, and clubfoot compared with the nonexposed population. The fact that an increased frequency of other defects attributed to vascular disruption was not found may be due to under-ascertainment, misclassification, or "lumping" of the defects identified in previous studies. PMID- 11263543 TI - CF carrier testing in a high risk population: anxiety, risk perceptions, and reproductive plans of carrier by "non-carrier" couples. AB - PURPOSE: The risk perceptions, psychological status and reproductive plans of 52 carrier by "noncarrier" (mutation screen negative) couples is the subject of this report. METHODS: Cystic fibrosis (CF) carrier testing was offered to relatives of individuals with CF. RESULTS: In this population testing was not associated with any significant adverse psychological effects, reproductive uncertainty, or inaccurate risk perceptions. CONCLUSIONS: The results of this study have important implications in light of the recent NIH CF Consensus Panel recommendations that CF carrier testing be offered to all high risk adults and all couples planning a pregnancy or seeking prenatal testing. PMID- 11263544 TI - Cytogenetic analysis using telemedicine consultation: an improved means of providing expert cross-coverage. AB - PURPOSE: We used telemedicine in providing cross-coverage for a clinical cytogenetics laboratory. A genetic teleconsultation system was used to provide expert cross-coverage for a laboratory in a neighboring metropolitan region for a 6 month period while the usual provider of these services was on military reserve duty. METHODS: The teleconsultation system was a commercially available Perceptive Scientific Instruments (PSI) workstation. Five hundred thirty-nine cytogenetic cases were performed during the study period in the home laboratory in Baltimore, Maryland. RESULTS: Karyotypes and supporting metaphase spreads were transmitted by modem to the covering director, whereas work sheets and reports were faxed. Physical transfer of data was not necessary, and turn-around-time was not increased. CONCLUSION: This ability to employ a remote part time director has significant benefits for the laboratory with an absentee director for short or even extended periods of time. We conclude that the use of telemedicine in clinical cytogenetics proved to be an efficient and a cost effective means of providing genetics services to a region during a time of cross-coverage need. PMID- 11263546 TI - Resurgent bacterial sexually transmitted disease among men who have sex with men- King County, Washington, 1997-1999. AB - During the late 1980s and early 1990s, King County, Washington (1998 population: 1.6 million), experienced a substantial epidemic of infectious syphilis (i.e., primary, secondary, and early latent). Subsequently, reported cases of infectious syphilis declined to six cases in 1995 and one in 1996; five of the 1995 cases and the case in 1996 were believed to have been acquired outside King County. However, in 1997, sustained spread of syphilis was reestablished in King County. To determine whether this reemergence was associated with changes in the epidemiology of other sexually transmitted diseases (STDs), Public Health-Seattle and King County (PHSKC) analyzed notifiable STD data for 1997-1999. This report summarizes the results of this analysis, which indicate that infectious syphilis among men who have sex with men (MSM) in King County increased to 46 cases during January-June 1999, and chlamydia and gonorrhea also increased among MSM attending public health clinics. PMID- 11263547 TI - Inadvertent use of Bicillin C-R for treatment of syphilis--Maryland, 1998. AB - In October 1998, the Maryland Department of Health and Mental Hygiene (MDH) was notified that a public sexually transmitted disease (STD) clinic in a county (county A) had used a nonrecommended preparation to treat syphilis patients during January-October 1998. The clinic had been inadequately treating syphilis patients or syphilis contacts with Bicillin C-R (a mixture of 1.2 million units [MU] benzathine penicillin G [BPG] and 1.2 MU procaine penicillin G), rather than with Bicillin L-A (2.4 MU BPG). Compared with short-acting procaine penicillin G, BPG has a longer half-life considered essential for effective syphilis treatment because it yields sustained spirochetecidal levels needed to treat the slowly reproducing agent of syphilis, Treponema pallidum. The inadvertent use of Bicillin C-R, which contains only half the recommended dose of BPG for syphilis, was recognized by a health-care provider at the STD clinic in a neighboring county (county B) approximately 1 month after county B had borrowed BPG from county A. This report summarizes the investigation of the use of Bicillin C-R to treat STD patients in county A and discusses the frequency of Bicillin C-R use in STD clinics nationwide. Findings of this investigation indicate that inadvertent Bicillin C-R use is more frequent than previously known and that preventive measures should be taken to minimize such use. PMID- 11263545 TI - Medium chain acyl-CoA dehydrogenase deficiency human genome epidemiology review. AB - Medium chain acyl-CoA dehydrogenase (MCAD) is a tetrameric flavoprotein essential for the beta-oxidation of medium chain fatty acids. MCAD deficiency (MCADD) is an inherited error of fatty acid metabolism. The gene for MCAD is located on chromosome one (1p31). One variant of the MCAD gene, G985A, a point mutation causing a change from lysine to glutamate at position 304 (K304E) in the mature MCAD protein, has been found in 90% of the alleles in MCADD patients identified retrospectively. There is a high frequency of MCADD among people of Northern European descent, which is believed to be due to a founder effect. MCADD is inherited in an autosomal recessive manner. Of patients clinically diagnosed with MCADD, 81% who have been identified retrospectively are homozygous for K304E, and 18% are compound heterozygotes for K304E. Clinical data on the probability of clinical disease indicates that MCADD patients are at risk for the following outcomes: hypoglycemia, vomiting, lethargy, encephalopathy, respiratory arrest, hepatomegaly, seizures, apnea, cardiac arrest, coma, and sudden and unexpected death. Long-term outcomes include developmental and behavioral disability, chronic muscle weakness, failure to thrive, cerebral palsy, and attention deficit disorder (ADD). Differences in clinical disease specific to allelic variants have not been documented. Factors that may increase risk for disease onset or modify disease severity are age when the first episode occurred, fasting, and presence of infection. Acute attacks must be treated immediately with appropriate intravenous doses of glucose. For those diagnosed, long-term management of the disease includes preventing stress caused by fasting and maintaining a high carbohydrate, reduced-fat diet, and carnitine supplementation. Hospitalization costs attributable to morbidity and mortality from MCADD are unknown; MCADD is not a diagnosis in the International Classification of Disease, 10th Revision (ICD-10) codebook. Furthermore, the penetrance of the MCAD genotypes is unknown; there appears to be a substantial number of asymptomatic MCADD individuals and some uncertainty regarding which individuals will manifest symptoms and which individuals will remain asymptomatic. Several technologies are available to detect MCADD. Diagnostic technologies include DNA-based tests for K304E mutations using the polymerase chain reaction (PCR), and the detection of abnormal metabolites in urine. Screening technologies include tandem mass spectrometry (MS/MS), which detects abnormal metabolites mostly in blood. State programs are beginning to offer screening in newborns for MCADD using MS/MS. In addition, a private company currently offers voluntary supplemental newborn screening for MCADD to birthing centers. PMID- 11263548 TI - Availability of hepatitis B vaccine that does not contain thimerosal as a preservative. AB - On August 27, 1999, Merck Vaccine Division (Merck & Co., Inc., West Point, Pennsylvania) received approval from the Food and Drug Administration (FDA) of a supplement to Merck's license application to include the manufacture of single antigen preservative-free hepatitis B vaccine (Recombivax HB, Pediatric); distribution is expected to begin September 13, 1999. In addition, SmithKline Beecham Biologicals (SmithKline Beecham, Philadelphia, Pennsylvania), expects to make single-antigen preservative-free hepatitis B vaccine (Engerix-B, Pediatric) available in the near future. Further product information will be provided when it becomes available. Product packaging and labels will indicate that these vaccines do not contain preservative. PMID- 11263549 TI - Transplantation tolerance and mixed chimerism: at the frontier of clinical application. AB - Although the persistence of donor-type hematopoietic cells in low numbers (microchimerism) is well established in some transplant recipients, its relevance for graft acceptance is still a matter of debate. On the other hand, clonal deletion of donor-specific alloreactive cells associated with mixed chimerism (macrochimerism) has reliably produced long-term graft tolerance in pre-clinical models. So far, the cytoablative conditioning regimens required to achieve mixed chimerism have hampered the clinical development of such protocols. Here, we discuss recent observations suggesting that the deliberate induction of hematopoietic cell chimerism might become a feasible strategy to achieve transplantation tolerance in clinics. PMID- 11263550 TI - Acute diverticulitis in heart- and lung transplant patients. AB - Significant gastrointestinal complications have been observed in patients following heart- and lung transplantation. These complications can occur in the immediate post-operative period or remote from the time of transplantation. We retrospectively reviewed the medical records of 268 consecutive patients who received either heart- or lung transplants at Henry Ford Hospital between 1985 and 1998. Two hundred and thirty-three patients received heart transplants and 35 underwent lung transplantation. Two patients developed acute diverticulitis post transplant, both requiring surgery. Management of acute diverticulitis in the heart- and lung transplant population requires a high index of suspicion. Early and aggressive diagnosis is mandatory. Surgical intervention must be prompt when indicated, with meticulous attention to surgical technique. With appropriate intervention, reasonable outcomes can be expected. PMID- 11263551 TI - Calcineurin-inhibitor induced pain syndrome (CIPS): a severe disabling complication after organ transplantation. AB - Bone pain after transplantation is a frequent complication that can be caused by several diseases. Treatment strategies depend on the correct diagnosis of the pain. Nine patients with severe pain in their feet, which was registered after transplantation, were investigated. Bone scans showed an increased tracer uptake of the foot bones. Magnetic resonance imaging demonstrated bone marrow oedema in the painful bones. Pain was not explained by other diseases causing foot pain, like reflex sympathetic dystrophy, polyneuropathy, Morton's neuralgia, gout, osteoporosis, avascular necrosis, intermittent claudication, orthopaedic foot deformities, stress fractures, and hyperparathyroidism. The reduction of cyclosporine- or tacrolimus trough levels and the administration of calcium channel blockers led to relief of pain. The Calcineurin-inhibitor Induced Pain Syndrome (CIPS) is a rare but severe side effect of cyclosporine or tacrolimus and is accurately diagnosed by its typical presentation, magnetic resonance imaging and bone scans. Incorrect diagnosis of the syndrome will lead to a significant reduction of life quality in patients suffering from CIPS. PMID- 11263552 TI - Endoluminal laser-Doppler measurements of jejunal perfusion in patients undergoing liver transplantation. AB - Patients undergoing liver transplantation are at risk of developing the multiple organ dysfunction syndrome, and attention has been focused on the pathogenic role of decreased gastro-intestinal mucosal perfusion. The aim of this study was to investigate the use of laser-Doppler flowmetry for determination of jejunal perfusion. In 10 patients an endoluminal laser-Doppler catheter was positioned with the tip in the jejunum for continuous measurements of jejunal perfusion. The anhepatic phase was associated with a progressive decrease in jejunal perfusion to 49 (40/65)% (P < 0.01) of dissection phase value. At the end of surgery the jejunal perfusion had increased to 134 (103/158)% (P < 0.01) of dissection phase jejunal perfusion. The endoluminal laser-Doppler technique was found to be easily applicable for continuous monitoring of jejunal perfusion, and the technique could prove valuable in detecting gastro-intestinal hypoperfusion in patients undergoing liver transplantation. PMID- 11263553 TI - Analysis of potential porcine endogenous retrovirus (PERV) transmission in a whole-organ xenotransplantation model without interfering microchimerism. AB - The question whether porcine xenografts can lead to porcine endogenous retrovirus (PERV) infection of recipients is critical for the evaluation of the safety of pig-to-man xenotransplantation. Unfortunately, polymerase chain reaction (PCR) based analysis of potential PERV infections in nonhuman-primate whole-organ xenotransplantation models is hampered by false positive results due to chimeric porcine cells. To avoid the inherent analytical problem of xenomicrochimerism, we developed a non-life-supporting pig-to-primate kidney xenotransplantation model: porcine kidneys were transplanted, whereas the functioning recipient kidneys remained in situ. Subsequent to rejection (after 2 hours to 15 days), xenografts were removed, and recipients remained alive for up to 287 days. Immunosuppressive therapy based on cyclophosphamide, cyclosporine, and steroids was maintained for 28 days after transplantation. Using appropriate PCR assays, xenochimerism was found in tissue samples and partly even in peripheral blood leukocytes (PBLs) while the porcine kidneys were in situ. After graft removal, xenochimerism was no longer detectable, thus allowing analysis for possible PERV transmission. PMID- 11263554 TI - Transplantation of a single kidney per se does not lead to late graft dysfunction. AB - In unraveling the pathogenesis of chronic transplant dysfunction (CTD), non alloantigen specific factors, as ischemia/reperfusion and renal mass have been suggested to play a role in the process. The aim of the present study was to investigate the effect of the transplantation procedure per se on the development of CTD in a syngeneic kidney transplant model in the rat. Kidney transplantation was performed with the BN rat as donor and recipient, the recipient kidneys having been removed. Unilaterally nephrectomized (UNx) and native BN rats served as controls. Renal function was determined monthly (proteinuria and glomerular filtration rate/100 g body weight; GFR). The follow-up period was until 52 weeks post-transplantation. Histomorphological analysis of CTD according to the BANFF criteria was carried out. Immunohistochemical staining was performed to identify infiltrating cells (CD4, CD8, and ED1) and the expression of MHC class II and ICAM-1. Isografts had a minor, constant proteinuria during follow-up, which did not differ from that of UNx: 27 +/- 10 vs. 29 +/- 2 mg/24 h at week 52. Unilateral nephrectomy led to a significant reduction of the GFR, which was about 80% of that of native rats. The GFR of isografts did not differ from that of UNx rats. Histomorphology of renal isografts was comparable to UNx and native kidneys; some glomerulopathy and tubular atrophy leading to a total BANFF-score of 2.6 +/- 0.5. In native BN kidneys, few CD4+ cells and ED-1+macrophages (mphi) were found; MHC class II was constitutively expressed on the proximal tubules and ICAM-1 on the glomeruli and peritubular capillaries. UNx-kidneys showed a similar pattern. Isografts had significantly more CD4+ cells and Mphi, mainly localized in the glomeruli, and a more intense ICAM-1 expression in the glomeruli and interstitium. Transplantation of one kidney in itself does not lead to CTD. PMID- 11263555 TI - Hemorrhagic colitis due to a novel Escherichia coli serotype (O121:H19) in a transplant patient. AB - Infection due to enterohemorrhagic Escherichia coli (EHEC) has not been described in immunosuppressed patients. We recently saw a case of EHEC infection caused by a novel Shiga toxin II-producing Escherichia coli serotype (O121:H19) that caused hemorrhagic colitis in a patient with renal and cardiac transplants. The patient's signs, symptoms, and colon pathology were similar to reports of EHEC infection in immunocompetent patients. This case suggests that the immunosuppressed state may not alter the clinical presentation or histopathologic findings of this disorder. Assays for EHEC are not routinely done at most hospitals. Therefore, clinicians caring for transplant patients should be aware of the typical clinical presentation of EHEC infection, so that they can initiate appropriate laboratory investigation in suspected cases. PMID- 11263557 TI - Laparoscopic live donor nephrectomy for transplantation: urgent need for standardising procedures. PMID- 11263556 TI - Portal vein angioplasty using a transjugular, intrahepatic approach for treatment of extrahepatic portal vein stenosis after liver transplantation. AB - Symptomatic portal vein stenosis is an uncommon complication after liver transplantation. Portal vein angioplasty has been successfully established for treatment of portal vein stenosis using mesenteric or percutaneous, transhepatic approaches. We herein report on a patient who suffered from variceal bleeding due to portal hypertension 3 months after liver transplantation. After successful endoscopic sclerotherapy, an extrahepatic portal vein stenosis was diagnosed, and portal vein angioplasty was considered as primary therapeutic option. Instead of mesenteric or percutaneous, transhepatic approaches, we adopted a transjugular, intrahepatic access to introduce a 14-mm balloon catheter into the portal vein. Using this technique, angioplasty was successfully performed. After intervention, no further episodes of variceal bleeding occurred. We favour the transjugular, intrahepatic technique for portal vein angioplasty because it does not require general anesthesia, in contrast to the mesenteric approach, and it reduces the risk of intra-abdominal bleeding, compared to the percutaneous, transhepatic approach. PMID- 11263559 TI - Successful transplantation of a divided horse-shoe kidney following prolonged donor hypotension and long-distance transportation. PMID- 11263558 TI - Procalcitonin increase after anti-CD3 monoclonal antibody therapy does not indicate infectious disease. PMID- 11263560 TI - Arterialization of portal conduit in pancreas transplantation. PMID- 11263561 TI - Impact of cyclosporine and low-dose steroid therapy on insulin sensitivity and beta-cell function in patients with long-term liver grafts. AB - To examine whether factors controlling glucose tolerance, i.e., insulin sensitivity (SI) and first-(phi1) and second-phase insulin secretion (phi2), are impaired in after orthotopic liver transplantation (OLT), they were assesssed in patients that had undergone OLT for cirrhosis (n = 10) with cyclosporin A and low dose steroid therapy (5 mg prednisone per day) and were compared with those of healthy matched control subjects (n = 10). These factors were determined by means of computer-based analysis of frequently sampled intravenous glucose tolerance tests (FSIGTT). Glucose and insulin profiles (posthepatic insulin) did not differ between both groups, whereas C-peptide levels (prehepatic insulin) were elevated in the transplant group after the FSIGTT, indicating an increased hepatic insulin degradation. SI and (phi1 did not differ between both groups. phi2, however, was significantly enhanced (23.94 +/- 2.63 vs 13.88 +/- 1.25 min(-1), P < 0.05). These results indicate that cyclosporine and low-dose steroid therapy do not impair SI and phi1. However, enhanced phi2 compensates the increased hepatic insulin clearance. PMID- 11263562 TI - Selective glycopeptide mapping of erythropoietin by on-line high-performance liquid chromatography--electrospray ionization mass spectrometry. AB - Selective glycopeptide mapping of recombinant human erythropoietin (rhEPO) used as a model glycoprotein was successfully carried out by on-line high-performance liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) using a Vydac C18 column eluted in acetonitrile-1 mM ammonium acetate, pH 6.8. rhEPO expressed in a Chinese hamster ovary clone was exhaustively digested into four glycopeptides and nine peptides with endoproteinase Glu-C. Both glycopeptides and peptides were eluted with trifluoroacetic acid as the eluent, whereas only glycopeptides were eluted selectively with ammonium acetate in the following order: N38, N24, 0126, and N83. Furthermore, many glycoforms included in each glycopeptide were found to be separated by differences in the numbers of sialic acid and N-acetyllactosaminyl repeats. Twenty, 16 and 22 different N-linked oligosaccharides were determined at Asn24, 38, and 83, respectively, and two different O-linked oligosaccharides were observed at Ser126. Our method is simple, rapid, and useful for determining the carbohydrate structures at each glycosylation site and for elucidating the site-specific carbohydrate heterogeneity. PMID- 11263563 TI - Sensitive gas chromatographic--mass spectrometric screening of acetylated benzodiazepines. AB - GC-MS screening conditions were developed for 15 low-dosed benzodiazepines, covering alprazolam, flunitrazepam, flurazepam, ketazolam, lorazepam and triazolam, and the corresponding metabolites alpha-hydroxyalprazolam, 4 hydroxyalprazolam; 7-aminoflunitrazepam, desmethylflunitrazepam, 7 aminodesmethylflunitrazepam; hydroxyethylflurazepam, N-desalkylflurazepam; oxazepam and alpha-hydroxytriazolam, respectively. Benzodiazepines are analyzed on a polydimethylsiloxane column in both the scan and the multiple ion monitoring modes using on-column injection to attain maximal sensitivity. The reactive compounds are acetylated with pyridine and acetic anhydride for 20 min. The derivatives are stable for at least 4 days. The relative standard deviation observed with standard compounds at the low nanogram-level ranged from 1.13 to 4.87% within-day and from 1.12 to 4.94% between-day. Unequivocal identification potential, high chromatographic resolution and sensitivity are combined with minimal thermal degradation. The presented screening conditions provide the basis for a unique routine screening method for low-dosed benzodiazepines with a broad polarity range. PMID- 11263564 TI - Modified mass action law-based model to correlate the solubility of solids and liquids in entrained supercritical carbon dioxide. AB - The solubility of solids and liquids in supercritical CO2 with added entrainers was modeled with a modified version of the equation of Chrastil to include the effect of entrainers. By considering the formation of the solute-entrainer solvent complexes an equation is obtained which predicts an exponential increase of solubility with fluid density and/or entrainer concentration. The correlating model was tested by non-linear regression through a computerized iterative process for several systems where an entrainer was present. Four experimental parameters are easily regressed from experimental data, hence the corresponding properties of components such as chemical potentials or critical parameters are not needed. Instead of its simplicity, this thermodynamical model provided a good correlation of the solubility enhancement in the presence of entrainer effect. PMID- 11263565 TI - On-line packed column supercritical fluid chromatography--microwave-induced plasma atomic emission. AB - Interfacing and evaluation of packed column supercritical fluid chromatography (SFC)-microwave-induced plasma atomic emission detection (AED) is described. Via a flow splitter and an integral restrictor, efficient transfer of solutes from column to detector without band broadening is obtained. Variation of CO2 flow rate during pressure gradients has little influence on both AED signal and baseline drift while it provides similar sensitivity as in capillary SFC. Continuous introduction of CO2 in the plasma reduces the available range of emission domains; nevertheless the region of detection which is free of CO2 interferences allows selective detection of C1 and Br as reported in this paper. reserved. PMID- 11263566 TI - Single step on-column frit making for capillary high-performance liquid chromatography using sol-gel technology. AB - One step frit-making in packing fused-silica capillary column of high-performance liquid chromatography was developed using sol-gel technology. Frit fabrication procedure was quite simple without sintering. On-column frit was formed through gelling of sol solution with packing materials, silica gel, and jointing the particles together with capillary wall through bonded and immobilized networks. Solvent types and proportions in sol solution were selected. And the sol solution compositions as well as amount of silica gel particles were also optimized to achieve maximum strength. Such an on-column frit of 250 microm in diameter is capable of resistant packing pressure up to 500 bars in ultra-sonic bath action. Chemical resistance to solvents and extreme pHs were also tested. Scanning electron micrograms of the frit profile showed that the evolving sol-gel network joined particles to each other and onto the column wall. Routine runs in reversed phase mode, the frits of several columns proved to be effective enough to resist pressures without collapse. PMID- 11263567 TI - Interaction of surfactants with homologous series of peptides studied by reversed phase thin-layer chromatography. AB - The relative strength of interaction between anionic (SDS) and nonionic surfactant (octaethoxylated oleyl alcohol, GEN) and homologous series of peptides was determined by reversed-phase thin-layer chromatography (RP-TLC) carried out on alumina layers impregnated with paraffin oil. The relative strength of interaction was calculated and was correlated with the physicochemical parameters of peptides. It was established that each peptide interacted with both surfactants and with their mixture (1:1, m/m). The relative strength of interaction depended on the number of amino acid units in the peptide, side chain bulk and electronic properties and hydrophobicity of the amino acids. The impact of individual parameters highly depended on the character of surfactant. The data prove that the retention order of peptides can be modified by adding different surfactants and surfactant mixtures to the mobile phase resulting in improved separation. PMID- 11263569 TI - Rapid enantiomeric separation of polychlorinated biphenyls by electrokinetic chromatography using mixtures of neutral and charged cyclodextrin derivatives. AB - Electrokinetic chromatography with cyclodextrin derivatives (CD-EKC) was used to achieve the rapid enantiomeric separation of chiral polychlorinated biphenyls (PCBs). Thirteen of the 19 chiral PCBs stable at room temperature were individually separated into their two enantiomers by using 2 morpholinoethanesulfonic acid (MES) buffer (pH 6.5) containing carboxymethylated gamma-cyclodextrin (CM-gamma-CD) as pseudostationary phase mixed with beta cyclodextrin (beta-CD) or permethylated beta-cyclodextrin (PM-beta-CD). Urea was also added to increase the solubility of PCBs and cyclodextrins in the aqueous separation buffer. Several experimental parameters such as the nature, concentration, and pH of the buffer, nature and concentration of the cyclodextrin derivatives used, and the addition of different additives were studied in order to improve the enantiomeric separation. In addition, the effect of some instrumental parameters such as separation temperature and applied voltage was also investigated. PCBs were enantiomerically separated in less than 12 min by using a 50 mM MES buffer (pH 6.5) containing 20 mM CM-gamma-CD, 10 mM beta-CD or 20 mM PM-beta-CD, and 2 M urea at a temperature of 45 degrees C and an applied voltage of 20 kV. PMID- 11263568 TI - Separation of monomethyl-benz[a]anthracene isomers using cyclodextrin-modified electrokinetic chromatography. AB - Cyclodextrin-modified electrokinetic chromatography (CD-EKC) was investigated for the separation of 12 monomethylbenz[a]anthracene (MBA) isomers. Combined use of a polymeric surfactant, poly(sodium 10-undecenyl sulfate) (poly-SUS), with various types of neutral cyclodextrins (CDs) [beta-CD, gamma-CD, dimethyl-beta-CD (DM beta-CD), trimethyl-beta-CD (TM-beta-CD) and hydroxypropyl-beta-CD (HP-beta-CD)] were successful in CD-EKC separation of the MBA isomers. Baseline resolution of 10 of the 12 isomers, except for 9-MBA and 2-MBA, was achieved with gamma-CD at pH 9.75. The beta-CD, gamma-CD, and beta-CD derivatives (DM-beta-CD, TM-beta-CD, HP-beta-CD) were found to have different resolution and selectivity. Additionally, the tR/t0 values of isomers were found to be dependent on the type and concentration of the CD additives. In general, tR/t0 values of MBA isomers decrease with an increase in the concentration of beta-CD derivatives, whereas the reversed was true when the concentrations of native beta-CD and gamma-CD were varied. The combination of 5 mM gamma-CD, 0.5% (w/v) poly-SUS, 35% (v/v) acetonitrile at a pH of 9.75 provided the best selectivity and resolution of the MBA isomers with a separation time of 110 min. However, the use of 30 mM DM-beta CD under similar EKC conditions resulted in much faster separation (ca. 16 min) of 10 MBA isomers. PMID- 11263570 TI - Optimization of separation and migration behavior of chloropyridines in micellar electrokinetic chromatography. AB - The separation and migration behavior of pyridine and eight chloropyridines, including three monochloropyridines, four dichloropyridines, and 2,3,5 trichloropyridine were investigated by micellar electrokinetic chromatography using either sodium dodecyl sulfate (SDS) as an anionic surfactant or SDS-Brij 35 mixed micelles. Various parameters such as buffer pH, SDS concentration, Brij 35 concentration and methanol content that affect the separation were optimized. Complete separation of these chloropyridines was optimally achieved with a phosphate buffer containing SDS (30 mM) and methanol (10%, v/v) at pH 7.0. The resolution and selectivity of analytes could be considerably affected by the addition of methanol and/or Brij 35 to the background electrolyte. The migration order of these chloropyridines depends primarily on their hydrophobicity. However, electrostatic interactions may also play a significant role in the determination of the migration order of the positional isomers of chloropyridines. PMID- 11263571 TI - Prediction of retention times for anions in linear gradient elution ion chromatography with hydroxide eluents using artificial neural networks. AB - The feasibility of using an artificial neural network (ANN) to predict the retention times of anions when eluted from a Dionex AS11 column with linear hydroxide gradients of varying slope was investigated. The purpose of this study was to determine whether an ANN could be used as the basis of a computer-assisted optimisation method for the selection of optimal gradient conditions for anion separations. Using an ANN with a (1, 10, 19) architecture and a training set comprising retention data obtained with three gradient slopes (1.67, 2.50 and 4.00 mM/min) between starting and finishing conditions of 0.5 and 40.0 mM hydroxide, respectively, retention times for 19 analyte anions were predicted for four different gradient slopes. Predicted and experimental retention times for 133 data points agreed to within 0.08 min and percentage normalised differences between the predicted and experimental data averaged 0.29% with a standard deviation of 0.29%. ANNs appear to be a rapid and accurate method for predicting retention times in ion chromatography using linear hydroxide gradients. PMID- 11263572 TI - Ionophoretic studies on mixed metal--nitrilotriacetate--penicillamine complexes. AB - Paper ionophoresis is described for the study of equilibria in a mixed ligand complex system in solution. This method is based on the movement of a spot of metal ion in an electric field with the complexants added in the background electrolyte at pH 8.5. The concentration of the primary ligand (nitrilotriacetate) was kept constant, while that of the secondary ligand (penicillamine) was varied. The stability constants of the metal nitrilotriacetate-penicillamine complexes have been found to be 6.26 +/- 0.09 and 6.68 +/- 0.13 (log K values) for Al3+ and Th4+ complexes, respectively, at 35 degrees C and ionic strength 0.1 M. PMID- 11263573 TI - Quantitative FTIR detection in size-exclusion chromatography. AB - With the increasing popularity of evaporative interfaces, detection using Fourier Transform Infrared (FTIR) spectrometry in the mid-infrared region is becoming more important in size-exclusion chromatography (SEC). FTIR spectrometry is a powerful, and potentially very widely applicable, method for obtaining chemical functional group information for each molecular size fraction. Quantitative evaluation of polymer composition across the SEC chromatogram can provide more accurate characterization of heterogeneous polymer samples for problem solving and for material specification. The evaporative interface removes the SEC mobile phase at the exit of the column and deposits the eluting polymer as a continuous film stripe or as a series of discrete films on infrared transparent substrates. Initially this detection approach was used only for qualitative analysis. More recently, it is being used quantitatively. Previously we demonstrated that the quality of the film generated by the evaporative interface was critical to determining the suitability of the resulting FTIR spectra for quantitative analysis. In a continuation of this work, the objective of this paper is to develop a procedure for obtaining valid quantitative results for polymer blends with the interface. Experimental topics include improving the quality of polymer films by post-SEC treatments, off-line FTIR calibrating using other means to obtain high quality polymer films, and utilizing in-line SEC detectors in calibration. Interpretation aspects focus upon peak fitting of FTIR spectra, linear regression, partial least squares, and data pre-processing. PLS prediction with internal calibration using the second derivative of solvent-annealed film spectra was found to provide the best compromise between processing time, accuracy and precision. PMID- 11263574 TI - Computer-assisted high-performance liquid chromatography method development with applications to the isolation and analysis of phytoplankton pigments. AB - We used chromatography modeling software to assist in HPLC method development, with the goal of enhancing separations through the exclusive use of gradient time and column temperature. We surveyed nine stationary phases for their utility in pigment purification and natural sample analysis. For purification, a complex algal matrix was separated on an efficient monomeric column, from which partially purified fractions were collected and purified on polymeric columns that exaggerated resolution between pigments of interest. Additionally, we feature an HPLC method that is simple, fast, demonstrates excellent transferability and is ideal for quantitative analysis of pigments in dilute natural water samples. PMID- 11263575 TI - Characterization of polystyrene and polyisoprene by normal-phase temperature gradient interaction chromatography. AB - Temperature gradient interaction chromatography (TGIC) is applied to the characterization of polyisoprene (PI) and polystyrene (PS) using normal-phase (NP) stationary phase--bare silica or diol bonded silica. Tetrahydrofuran isooctane mixtures are used as a mobile phase. PI and linear and star shaped PS samples are successfully fractionated in terms of the molecular mass with a high resolution comparable to that of reversed-phase (RP) HPLC. Temperature dependence of the retention shows that the enthalpy of adsorption of PS to the stationary phase is exothermic. In addition, some characteristic features of the NP-TGIC system relative to those of RP-TGIC are presented, which include a high sensitivity on the polar end group and the simultaneous size-exclusion chromatographic and TGIC characterization of PS and PI mixtures. PMID- 11263576 TI - Quantitative determination of rocuronium in human plasma by liquid chromatography electrospray ionization mass spectrometry. AB - Liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) was used for the quantification of the neuromuscular blocking agent rocuronium in human plasma. Verapamil was used as internal standard. The samples were subjected to a dichloromethane liquid-liquid extraction after ion pairing of the positively charged ammonium compound with iodide prior to LC-MS. Optimized conditions involved separation on a Symmetry Shield RP-18 column (50 x 2.1 mm, 3.5 microm) using a 15-min gradient from 10 to 90% acetonitrile in water containing 0.1% trifluoroacetic acid at 250 microl/min. Linear detector responses for standards were observed from 25 to 2,000 ng/ml. The extraction recovery averaged 59% for rocuronium and 83% for the internal standard. The limit of quantification (LOQ), using 500 microl of plasma, was 25 ng/ml. Precision ranged from 1.3 to 19% (LOQ), and accuracy was between 92 and 112%. In plasma samples, at 20 and 4 degrees C, rocuronium was stable at physiological pH for 4 h; frozen at -30 degrees C it was stable for at least 75 days. The method was found suitable for the analysis of samples collected during pharmacokinetic investigations in humans. PMID- 11263577 TI - Determination of drug residues in water by the combination of liquid chromatography or capillary electrophoresis with electrospray mass spectrometry. AB - Methods for the determination of drug residues in water have been developed based on the combination of liquid chromatography (LC) or capillary electrophoresis (CE) with mass spectrometry (MS). For HPLC-MS two types of interfaces (pneumatically assisted electrospray ionization interface or an atmospheric pressure chemical ionization interface, respectively) were employed and compared in terms of detection limits. 2 mM Ammonium acetate at pH 5.5 and a methanol gradient was used for the HPLC-MS allowing the separation of a number of drugs such as paracetamol, clofibric acid, penicillin V, naproxen, bezafibrate, carbamazepine, diclofenac, ibuprofen and mefenamic acid. A 20 mM ammonium acetate solution, pH 5.1 was employed for the separation of clofibric acid, naproxen, bezafibrate, diclofenac, ibuprofen and mefenamic acid by CE-MS. Sample pretreatment was performed by solid-phase extraction (SPE) for HPLC-MS or by a combination of liquid-liquid extraction and SPE for CE-MS. The applicability of both the HPLC-MS and CE-MS method was demonstrated for several river water samples. PMID- 11263578 TI - New strategies for the determination of phenylurea pesticides by gas chromatography with hot splitless inlet systems. AB - Direct gas chromatographic methods to analyse phenylurea pesticides are discouraged by the thermal instability of these compounds, that in conventional hot splitless inlet systems leads to extensive and irreproducible formation of isocyanates and amines. However a careful control of the operating conditions, like the inlet temperature, the pressure and the presence of suitable chemical additives (as acetic acid, low-molecular-mass amines, organic anhydrides) can either: (i) minimise the thermal decomposition enabling the direct GC-MS analysis of phenylureas, or (ii) lead to reproducible conversion to isocyanates. Experimental design was employed to study the effect of the experimental variables on the thermal transformation of phenylurea pesticides in splitless inlet system. Two strategies were alternatively optimised: (i) the minimisation of degradation reactions to increase the signal of phenylureas; (ii) the maximisation of the degradation to isocyanates that are in turn determined. The maximal yields in isocyanate were obtained with high inlet temperatures, low carrier flows in the injection phase and the presence of acetic anhydride. By contrast, the use of relatively low inlet temperatures, high carrier flows during the injection and the presence of an amine maximise the response of the parent compounds. PMID- 11263579 TI - New polymeric sorbent for the solid-phase extraction of chlorophenols from water samples followed by gas chromatography-electron-capture detection. AB - A polymeric material, polyaniline, was employed as a new sorbent for solid-phase extraction (SPE) of some environmental pollutants from water samples. Chlorophenols were extracted from aqueous samples by SPE using 120 mg polyaniline and determined by gas chromatography with electron-capture detection. The acetate esters of these phenols were formed by the direct addition of acetic anhydride to the organic extractant in the presence of K2CO3. Different conditions were applied to obtain higher retaining capacity and breakthrough volumes. The results compared with those obtained by other groups. The RSD for a river water sample spiked at sub-ppb level was lower than 10% (n = 3) and detection limits were between 3 and 110 ng(-1). PMID- 11263580 TI - Characterisation of fatty acids in biological oil samples using comprehensive multidimensional gas chromatography. AB - Comprehensive multidimensional gas chromatography can adequately resolve very complex mixtures of analytes such as the fatty acid mixtures which are contained in, e.g., fish and vegetable oils. Well-ordered patterns are obtained in the two dimensional separation plane which can be used to tentatively identify peaks when no standard is available. The technique which can also be used for quantification, i.e., quantitative ratio analysis, should be especially useful for fingerprinting purposes. Unravelling the composition of complex mixtures such as fish oils appears to be highly rewarding. PMID- 11263581 TI - Should we change our dietary advice on cancer prevention? PMID- 11263582 TI - Current thoughts on the histopathogenesis of gastric cancer. PMID- 11263585 TI - Viruses and alcohol in the pathogenesis of primary hepatic carcinoma. PMID- 11263586 TI - Surveillance for hepatocellular carcinoma in cirrhosis: is it cost-effective? PMID- 11263587 TI - Breast cancer risk factors: PCB congeners. AB - The chronic effects of polychlorinated biphenyls (PCBs) are a public health concern, and a potential relationship with breast cancer has been postulated. The purpose of this study was to examine the possible relationship between PCBs and breast cancer. All women (134) treated by excision biopsy because of breast lump at Reina Sofia University Hospital, Cordoba, Spain over a period of 10 months were included in our study. They were all administered a questionnaire by interview, calculation of body mass index, histopathological examination of excised mass and chemical estimation of PCB congener levels in breast fat. The collected samples were from 65 (48.5%) women with benign lesions and 69 (51.5%) with malignant lesions. The variables associated with malignant lesions on univariate analysis were age, lactation period, overweight, PCB n-28 and PCB n 52. On the multivariate analysis PCB n-28 was found to be the most important risk factor (OR 9.6, 95% CI 3.8-24.4). Other risk factors were identified as age, drinking alcohol, low parity and overweight. If these findings can be confirmed in a large study population, however, they may have important implications for breast cancer risk. PMID- 11263588 TI - Review of anthropometric factors and breast cancer risk. AB - Epidemiological evidence implicating anthropometric risk factors in breast cancer aetiology is accumulating. For premenopausal women, breast cancer risk increases with increasing height, but decreases with higher weight or body mass index, and no association with increased central adiposity exists. For postmenopausal women, an increased risk of breast cancer is found with increasing levels of all the anthropometric variables including height, weight, body mass index, waist-hip ratio, waist circumference and weight gain. Weight loss appears to decrease risk, particularly if it occurs later in life. Breast size may be a risk factor for breast cancer, however, the current evidence is inconclusive. Several hypothesized biologic mechanisms exist to explain how anthropometric factors influence breast cancer risk. Obesity may increase levels of circulating endogenous sex hormones, insulin and insulin-like growth factors that all, in turn, increase breast cancer risk. Genetic predisposition to obesity and to specific body fat distributions are also implicated. With obesity, there are increased levels of fat tissue that can store toxins and can serve as a continuous source of carcinogens. Recommendations for future research on anthropometric factors and breast cancer are provided. Sufficient evidence exists to support strategies to avoid weight gain throughout life as a means of reducing postmenopausal breast cancer risk. PMID- 11263589 TI - Risk of breast cancer in relation to anthropometry, blood pressure, blood lipids and glucose metabolism: a prospective study within the Malmo Preventive Project. AB - Insulin resistance may be a risk factor for breast cancer, possibly through increased levels of oestrogens or insulin-like growth factor I. Insulin resistance has been associated with obesity, hypertension, dyslipidaemia and impaired glucose tolerance. We studied the relation of these factors to breast cancer risk in a prospective cohort study of 9738 women. Menopausal status was defined a priori, and 112 cases of invasive breast carcinoma occurred in women who were premenopausal at baseline and 157 cases in subjects who were peri/postmenopausal. Relative risks (RR) for breast cancer were calculated by Cox's proportional hazards analysis for different quartiles of height, weight, body mass index, blood pressure, pulse rate and serum levels of total cholesterol, triglycerides, fasting blood glucose and glucose at 120 min after an oral dose of glucose. Peri/postmenopausal women had a significantly increased age adjusted relative risk of breast cancer associated with height (RR = 1.78 for the highest versus lowest quartile), and the RR was increased over quartiles of cholesterol levels (P-value for trend: 0.05). No other significant associations were found. Adjustments for potential confounding factors or restriction of the analysis to cases and person-years before 55 years of age (premenopausal women), or after 55 years (peri/postmenopausal women), did not change PMID- 11263590 TI - Cyclin D1 protein overexpression and gene amplification in benign breast tissue and breast cancer risk. AB - Cyclin D1 amplification and/or protein overexpression have been observed not only in breast cancer but also in the putative early stages of breast neoplasia. In a case-control study nested within a cohort of 4888 women, we investigated whether the occurrence of cyclin D1 gene amplification and/or protein overexpression in benign breast tissue might identify women at increased risk of subsequent breast cancer development. Cases were 92 women with a histological diagnosis of benign breast disease who subsequently developed breast cancer. Five controls (women with benign breast disease who had not developed breast cancer by the date of diagnosis of the corresponding case) were selected randomly for each case from those non-cases available within strata defined by screening centre, National Breast Screening Study (NBSS) study arm, year of birth and age at diagnosis of benign breast disease. Paraffin blocks of benign tissue were suitable for immunostaining for 71 cases and 293 controls. Sufficient DNA for analysis was obtained from a total of 356 subjects (69 cases, 287 controls). The benign breast tissues and breast cancers were immunostained for cyclin D1 and also analysed for the presence of cyclin D1 gene amplification by differential polymerase chain reaction (PCR). Fifteen cases and 60 controls showed evidence of cyclin D1 immunostaining, and 12 cases and 29 controls showed cyclin DL gene amplification. There was essentially no association between cyclin D1 protein overexpression in benign breast tissue and risk of subsequent breast cancer (adjusted odds ratio (OR) 1.06; 95% confidence interval (CI) 0.56-2.02). After adjustment for potential confounding, there was a statistically non-significant 40% increase in risk of breast cancer in association with cyclin D1 gene amplification (adjusted OR 1.41; 95% CI 0.62-3.22). As multiple genetic changes are required to develop breast cancer, it may not be until the cascade of molecular alterations leading to breast cancer development is understood that identification of biomarkers of breast cancer risk will be possible. PMID- 11263591 TI - Determinant factors for diagnostic delay in operable breast cancer patients. AB - Randomized trials of mammographic screening have provided strong evidence that early diagnosis and treatment of breast cancer can reduce the specific mortality. Moreover, in a recent systematic review of published studies, delays of 3-6 months between symptom onset and treatment have been clearly found to be associated with lower survival rates for breast cancer patients. The aim of this study was to examine delays registered among breast cancer patients in southern Italy, in order to recognize their determining factors so as to provide women with a better opportunity for survival. The variables examined were age (< 50, 50 64, > or = 65 years), education (< or = 5, > 5 school years); symptom status at first presentation (symptomatic or asymptomatic); date of first symptom presentation; date of first consultation with a health provider; the type of health provider consulted; tumour size and nodal status according to the pTNM system. Time intervals were categorized into: < 1 month, 1-3 months and > 3 months for patient and medical delay; 1-3 months, 3-6 months, > 6 months for overall delay. Patient delay was associated with age and education: a higher risk was found for women of over 65 years age (odds ratio (OR) 2.1, 95% confidence interval (CI) 1.2-3.5) and with < or = 5 years school attendance (OR 3.3, 95% CI 2.0-5.6). Medical delay was seen to be associated with the professional figure: significant differences were found between senologists (oncologists exclusively dedicated to breast cancer operation) and other specialists (OR 3.5, 95% CI 1.5 8.4). Young age and symptomatic presentation were found to be high risk factors. Concerning tumour size in overall delay, in cases where the tumour was > 2 cm the OR was 2.4 (95% CI 1.5-3.7). Our study suggests that diagnostic delay can be reduced by providing more efficient training programmes for members of the medical profession and by producing educational training programmes targeted specifically at each age category (i.e. in older women more attention to education in prevention; in younger women correct information about mammography and specialized structures). PMID- 11263592 TI - Stomach cancer-related mortality. AB - In Japan stomach cancer remains the leading cause of cancer-related mortality. We analysed the annual mortality rate of stomach cancer in relation to age, gender and life expectancy in Japan between 1970 and 1995. The adjusted stomach cancer related mortality rates decreased from 88.9 in 1970 to 45.4 per 100,000 in 1995 in males and from 46.5 to 18.5 per 100,000 in females. The male-female ratio for stomach cancer-related mortality in all ages was 1.9-2.5 during this 25-year period, and the mortality rate was higher in females than in males at young age. The negative contribution to life expectancy for stomach cancer in males was 0.65 years and 0.42 years in females, which is consistent with a higher mortality rate in males. This negative contribution was 41.8% of total cancer in 1970 and 39.4% in 1995 in males and 34.4% and 16.0%, respectively, in females. Our results demonstrated the need to take into consideration the characteristics of stomach cancer in young women and the effects of ageing when designing programmes aimed at prevention and control of this malignancy. PMID- 11263593 TI - Human leukocyte antigens related to Epstein-Barr virus-associated gastric carcinoma in Japanese patients. AB - To assess the association between specific types of human leukocyte antigen and the risk of Epstein-Barr virus (EBV)-associated gastric carcinoma, serological typing for major histocompatibility complex class I and class II antigens was performed for 110 EBV-positive and 155 EBV-negative gastric carcinoma cases. In class I analysis, the frequency of B59 in the EBV-positive cases was higher than for the EBV-negative cases (odds ratio (OR) 3.06; 95% confidence interval (CI) 1.02-9.23). For class II antigens, DQ3 and DR9 frequencies in the EBV-positive cases were higher (OR 1.94; 95% CI 1.16-3.24 and OR 1.93; 95% CI 1.11-3.37, respectively), whereas DR11 frequency was lower than found in the EBV-negative cases (OR 0.10; 95% CI 0.01-0.79). After adjusting for multiple comparisons, only DR11 frequency remained significantly lower in the EBV-positive cases (P = 0.04), and the association of DQ3 was marginally significant (P = 0.05). These results suggest that the presence of DR11-restricted cytotoxic T cells (CTLs) related to EBV-associated gastric carcinoma, or a deficiency of DR11 and a high frequency of DQ3 may be genetic markers for a population at greater risk of EBV-associated gastric carcinoma. However, further extensive studies to more cases and DNA typing are needed because our findings in this study are exploratory. PMID- 11263594 TI - Epidemiology and prevention of bladder cancer. PMID- 11263595 TI - Use of cytokeratins 7 and 20 in determining the origin of metastatic carcinoma of unknown primary, with special emphasis on lung cancer. AB - Metastatic carcinoma of unknown primary is a common problem, accounting for up to 10-15% of all solid tumours at presentation. Proper identification of the site of origin has prognostic and therapeutic significance. Prior immunohistochemical methods to identify the site of origin have been useful in a limited number of cases. Differential cytokeratin staining may be useful in this setting, particularly in identifying metastases from lung cancer. We have identified 144 cases of metastatic carcinoma of unknown primary to bone, lung or liver at Brigham and Women's Hospital between 1 January 1997 and 1 July 1998. Cytokeratin (CK) 7 and CK20 were used in 75 of these cases to narrow down the possible sites of the primary tumours. All of these cases were ambiguous as to the site of the primary tumour. Forty-five cases were CK7+/CK20-, 15 cases were CK7-/CK20-, 9 cases were CK7-/CK20+ and 6 cases were CK7+/CK20+. Three of the cases were selected for detailed presentation and discussion as well as a discussion of the pertinent literature. Overall, the CK7+/CK20- phenotype favours a lung primary, the CK7+/CK20+ phenotype strongly favours transitional cells (urothelial) carcinoma, the CK7-/CK20+ phenotype favours colorectal carcinoma, while the CK7 /CK20- profile is not helpful. PMID- 11263596 TI - Familial risks in invasive and in situ cervical cancer by histological type. AB - The Swedish Family-Cancer Database was used to analyse familial relationships in mothers and daughters with invasive and in situ cervical cancers by histological type during the years 1958-1996, including a total of 21,727 and 191,081 cases, respectively. Familial standardized incidence ratios (SIRs) were calculated separately for mothers and daughters and for invasive and in situ squamous cell carcinoma (SCC) and adenocarcinoma. Familial risks were about 2.0 in invasive SCC and less in in situ SCC. Limited analyses could be carried out on adenocarcinoma because the number of cases was small. However, familial risks were not much smaller in families where only SCC was diagnosed compared with those where both SCC and adenocarcinoma were present. A comparison of cancers between mothers and daughters showed an association between cervical cancer and SCC of skin, and between cervical cancer and smoking-related cancers. The familial risks were unaffected in Poisson regression analysis on many possible intervening variables. The data suggest that host factors modulate an individual's response to human papillomavirus infections. PMID- 11263597 TI - Epidemiology of adenocarcinoma and squamous cell carcinoma of the oesophagus. PMID- 11263598 TI - The UK National Barrett's Oesophagus Registry (UKBOR): aims and progress. PMID- 11263599 TI - Reactivation of ischemic events in acute coronary syndromes: results from GUSTO IIb. Gobal Use of Strategies To Open occluded arteries in acute coronary syndromes. AB - OBJECTIVES: We sought to determine the incidence of and risk factors for thrombotic events early after discontinuing antithrombin therapy in patients with acute coronary syndromes. BACKGROUND: Discontinuation of treatment with heparin and other thrombin inhibitors in patients with unstable coronary syndromes has related to clinical and biochemical evidence of early reactivation of thrombosis. METHODS: We studied 8,943 of the 12,142 patients with acute coronary syndromes enrolled in the Global Use of Strategies To Open occluded arteries in acute coronary syndromes trial of hirudin versus heparin. We excluded patients who received no study drug, lacked timing data, died or had myocardial (re)infarction [(re)MI] during study-drug infusion, or began heparin treatment within 2 h after treatment with the study drug was stopped. We assessed the incidence and timing of (re)MI by type and timing of antithrombin treatment. RESULTS: In all, 215 patients (2.4%) suffered (re)MI, 49 within 12 h of antithrombin therapy discontinuation and 166 between hour 12 and hospital discharge. The duration of infusion did not differ between the hirudin and heparin groups. The rate of early re(MI) after drug therapy discontinuation was significantly higher in patients given heparin versus hirudin (0.8% vs. 0.3%, p = 0.002). Patients with (re)MI had higher mortality at 30 days (23.6% vs. 2.4%, p = 0.001) and 1 year (35.2% vs. 6.7%, p = 0.001) compared with patients without (re)MI. CONCLUSIONS: The incidence of (re)MI was clustered within 12 h of heparin therapy discontinuation, with the greatest risk within 4 h. There was no evidence of early reactivation of thrombotic events after hirudin. Patients who had (re)infarction had worse outcomes. Better understanding of the mechanism and possible prevention of recurrent thrombosis is needed. PMID- 11263600 TI - Survival of patients with diabetes and multivessel coronary artery disease after surgical or percutaneous coronary revascularization: results of a large regional prospective study. Northern New England Cardiovascular Disease Study Group. AB - OBJECTIVES: We sought to assess survival among patients with diabetes and multivessel coronary artery disease (MVD) after percutaneous coronary intervention (PCI) and after coronary artery bypass grafting surgery (CABG). BACKGROUND: The Bypass Angioplasty Revascularization Investigation (BARI) demonstrated that diabetics with MVD survive longer after initial CABG than after initial PCI. Other randomized trials or observational databases have not conclusively reproduced this result. METHODS: A large, regional database was linked to the National Death Index to assess five-year mortality. Of 7,159 consecutive patients with diabetes who underwent coronary revascularization in northern New England during 1992 to 1996, 2,766 (38.6%) were similar to those randomized in the BARI trial. Percutaneous coronary intervention was the initial revascularization strategy in 736 patients and CABG in 2,030. Cox proportional hazards regression was used to calculate risk-adjusted hazard ratios (HR) and 95% confidence intervals (CI 95%). RESULTS: Patients who underwent PCI were younger, had higher ejection fractions and less extensive coronary disease. After adjusting for differences in baseline clinical characteristics, patients with diabetes treated with PCI had significantly greater mortality relative to those undergoing CABG (HR = 1.49; CI 95%: 1.02 to 2.17; p = 0.037). Mortality risk tended to increase more among 1,251 patients with 3VD (HR = 2.02; CI 95%: 1.04 to 3.91; p = 0.038) than among 1,515 patients with 2VD (HR = 1.33; CI 95%: 0.84 to 2.1; p = 0.21). CONCLUSIONS: In this analysis of a large regional contemporary database of patients with diabetes selected to be similar to those enrolled in the BARI trial, five-year mortality was significantly increased after initial PCI. This supports the BARI conclusion on initial revascularization of patients with diabetes and MVD. PMID- 11263601 TI - Coronary artery bypass graft or percutaneous coronary interventions in patients with diabetes: another nail in the coffin or "too close to call?". PMID- 11263602 TI - Predictors of diffuse and aggressive intra-stent restenosis. AB - OBJECTIVES: This study was performed to investigate the causes of diffuse and aggressive intra-stent restenosis. BACKGROUND: Although restenosis is usually considered to be a dichotomous variable, there is clinical relevance to the severity of restenosis. It is not known which variables are predictive of diffuse or aggressive intra-stent restenosis. METHODS: A consecutive series of 456 coronary lesions with in-stent restenosis was evaluated for the type of restenosis using quantitative coronary angiography. Restenosis was defined as > or = 50% diameter stenosis at follow-up angiography, diffuse restenosis as a follow-up lesion length > or = 10 mm and aggressive restenosis as either an increase in lesion length from the original lesion or a restenotic narrowing tighter than the original. Clinical, anatomic and procedural characteristics were evaluated for lesions associated with these types of restenosis. RESULTS: Diffuse restenosis was associated with a smaller reference artery diameter, longer lesion length, female gender, longer stent length and the use of coil stents. Aggressive restenosis was more common in women, with the use of Wallstents and with long stent to lesion length ratios. Aggressive restenosis occurred earlier and was more closely associated with symptoms and myocardial infarctions than nonaggressive restenotic lesions. CONCLUSIONS: Markers for diffuse restenosis were also important markers for the presence of any restenosis. A long stent to lesion length ratio is an important marker for aggressive restenosis. When severe forms of in-stent restenosis occur, they tend to present earlier and with more symptoms, including myocardial infarction. More careful consideration of the type of in-stent restenosis may aid in identifying when alternative strategies may be useful. PMID- 11263603 TI - Edge stenosis and geographical miss following intracoronary gamma radiation therapy for in-stent restenosis. AB - OBJECTIVES: We sought to determine the relationship between geographical miss (GM) and edge restenosis (ERS) following intracoronary radiation therapy. BACKGROUND: Edge restenosis may be a limitation of intracoronary irradiation to prevent in-stent restenosis (ISR). Inadequate radiation source coverage of the injured segment (GM) has been proposed as a cause of ERS. We studied the relationship between GM and ERS following 192Ir treatment of ISR. METHODS: There were 100 patients with native vessel ISR in WRIST (Washington Radiation for In Stent Restenosis Trial), in which patients with ISR were first treated with conventional techniques and then randomized to gamma irradiation (192Ir) or placebo. Geographical miss was defined as segments proximal or distal to the treated lesion that were subjected to injury during primary intervention but were not covered by the radiation source. RESULTS: Geographical miss was documented in 56 of 164 edges (34%). Edge restenosis was noted at eight of 80 radiated edges and in four of 84 placebo edges. In the irradiated group, ERS was observed in 21% of edges with GM and in 40% of edges without GM (p = 0.035). In contrast, in the placebo group, ERS was observed in only 7% of edges with GM and in 4% of edges without GM (p = NS). The late edge lumen loss was higher in the irradiated group with GM as compared to placebo with GM (0.74 +/- 0.57 vs. 0.41 +/- 0.50 mm, p = 0.016). CONCLUSIONS: Edge restenosis following gamma irradiation treatment of ISR is related to GM: a mismatch between the segment of artery injured during the primary catheter-based intervention and the length of the radiation source. PMID- 11263604 TI - Preintervention arterial remodeling affects clinical outcome following stenting: an intravascular ultrasound study. AB - OBJECTIVES: The study was done to elucidate the relationship between baseline arterial remodeling and clinical outcome following stenting. BACKGROUND: The impact of preintervention arterial remodeling on subsequent vessel response and clinical outcome has been reported following nonstent coronary interventions. However, in stented segments, the impact of preintervention remodeling on clinical outcome has not been clarified. METHODS: Preintervention remodeling was assessed in 108 native coronary lesions by using intravascular ultrasound (IVUS). Positive remodeling (PR) was defined as vessel area (VA) at the target lesion greater than that of average reference segments. Intermediate or negative remodeling (IR/NR) was defined as VA at the target lesion less than or equal to that of average reference segment. Remodeling index expressed as a continuous variable was defined as VA at the target lesion site divided by that of average reference segments. RESULTS: Positive remodeling was present in 59 (55%) and IR/NR in 49 (45%) lesions. Although final minimal stent areas were similar (7.76 +/- 1.80 vs. 8.09 +/- 1.90 mm2, p = 0.36), target vessel revascularization (TVR) rate at nine-month follow-up was significantly higher in the PR group (22.0% vs. 4.1%, p = 0.01). By multivariate logistic regression analysis, higher remodeling index was the only independent predictor of TVR (p = 0.02). CONCLUSIONS: Lesions with PR before intervention appear to have a worse clinical outcome following IVUS-guided stenting. Intravascular ultrasound imaging before stenting may be helpful to stratify lesions at high risk for accelerated intimal proliferation. PMID- 11263605 TI - Prognostic value of exercise echocardiography in 2,632 patients > or = 65 years of age. AB - OBJECTIVES: We sought to determine the prognostic value of exercise echocardiography in the elderly. BACKGROUND: Limited data exist regarding the prognostic value of exercise testing in the elderly, a population which may be less able to exercise and is at increased risk of cardiac death. METHODS: Follow up (2.9 +/- 1.7 years) was obtained in 2,632 patients > or = 65 years who underwent exercise echocardiography. RESULTS: There were 1,488 (56%) men and 1,144 (44%) women (age 72 +/- 5 years). The rest ejection fraction was 56 +/- 9%. Rest wall motion abnormalities were present in 935 patients (36%). The mean work load was 7.7 +/- 2.3 metabolic equivalents (METs) for men and 6.5 +/- 1.9 METs for women. New or worsening wall motion abnormalities developed with stress in 1,082 patients (41%). Cardiac events included cardiac death in 68 patients and nonfatal myocardial infarction in 80 patients. The addition of the exercise electrocardiogram to the clinical and rest echocardiographic model provided incremental information in predicting both cardiac events (chi-square = 77 to chi square = 86, p = 0.003) and cardiac death (chi-square = 71 to chi-square = 86, p < 0.0001). The addition of exercise echocardiographic variables, especially the change in left ventricular end-systolic volume with exercise and the exercise ejection fraction, further improved the model in terms of predicting cardiac events (chi-square = 86 to chi-square = 108, p < 0.0001) and cardiac death (chi square = 86 to chi-square = 99, p = 0.004). CONCLUSIONS: Exercise echocardiography provides incremental prognostic information in patients > or = 65 years of age. The best model included clinical, exercise testing and exercise echocardiographic variables. PMID- 11263606 TI - Predictive value of systolic and diastolic function for incident congestive heart failure in the elderly: the cardiovascular health study. AB - OBJECTIVES: We sought to assess the ability of echocardiographic indices of systolic and diastolic function to predict incident congestive heart failure (CHF). BACKGROUND: Noninvasive indices of subclinical systolic and/or diastolic dysfunction that can be used to identify patients in a transition phase between normal cardiac function and clinical CHF would be valuable. Though midwall shortening and Doppler mitral inflow patterns are seemingly well suited to predict subsequent CHF, the predictive value of these indices has not been investigated. METHODS: We studied 2,671 participants in the Cardiovascular Health Study who were free of coronary heart disease, CHF or atrial fibrillation. Clinical and quantitative echocardiographic data were obtained in all participants. RESULTS: At a mean follow-up of 5.2 years (range 0 to 6 years), 170 participants (6.4% of the cohort) developed CHF. Although 96% of these participants had normal or borderline ejection fraction (EF) at baseline, only 57% had normal or borderline EF at the time of hospitalization. In multivariate modeling, fractional shortening at the endocardium (relative risk [RR] 1.85 per 10-unit decrease, confidence interval [CI] 1.27 to 2.39), fractional shortening at the midwall (RR 1.29 per five-unit decrease, 95% CI 1.11-1.51) and peak Doppler peak E (RR 1.15 for each 0.1 M/s increment; CI 1.02 to 1.21) independently predicted incident CHF. Both high and low Doppler E/A ratios were predictive of incident CHF. CONCLUSIONS: Roughly half the occurrences of CHF in this population are associated with normal or borderline EF. Echocardiographic findings suggestive of subclinical contractile dysfunction and diastolic filling abnormalities are both predictive of subsequent CHF. The standard (FSendo) and refined (FSmw) parameters of systolic function performed similarly in this regard, though subjects with left ventricular hypertrophy and depressed FSmw are at particularly high risk for subsequent CHF. PMID- 11263607 TI - Prognostic value of Doppler echocardiographic mitral inflow patterns: implications for risk stratification in patients with chronic congestive heart failure. AB - OBJECTIVES: This prospective study tested whether transmitral flow patterns add incremental value to peak oxygen consumption (VO2) in determining the prognosis of patients with chronic congestive heart failure (CHF) and systolic dysfunction. BACKGROUND: Peak VO2 is an objective marker of functional capacity and is routinely used as a criterion to identify heart transplant candidates. Diastolic dysfunction limits functional capacity, but its prognostic importance relative to that of peak VO2 is unknown. METHODS: Peak VO2 and mitral inflow velocities were prospectively measured in 311 consecutive patients (mean age 54 years, 84% male) with impaired left ventricular function (ejection fraction <40%; 88 patients with ischemic and 223 with dilated cardiomyopathy) who were evaluated for heart transplant candidacy. RESULTS: During a mean follow-up period of 512 +/- 314 days, 65 patients died and 43 patients underwent heart transplantation. Diastolic filling patterns, peak VO2 and left ventricular end-diastolic diameters were independent predictors of cardiac mortality. In patients with peak VO2 < or = 14 ml/min per kg body weight, the outcome was markedly poorer in the presence of restrictive filling patterns as compared with their absence (two-year survival rate 52% vs. 80%). Similarly, despite peak VO2 levels >14 ml/min per kg, the outcome was less favorable in the presence of restrictive filling patterns (two year survival rate 80% vs. 94%). A risk-stratification model based on the identified independent noninvasive predictors separated groups into those with high (93%), intermediate (65%) and low (39%) two-year survival rates. CONCLUSIONS: Transmitral flow patterns add incremental value to peak VO2 in determining the prognosis of patients with CHF and impaired systolic function. PMID- 11263608 TI - Angiotensin converting enzyme (ACE) and non-ACE dependent angiotensin II generation in resistance arteries from patients with heart failure and coronary heart disease. AB - OBJECTIVES: We sought to demonstrate non-angiotensin converting enzyme (ACE) dependent angiotensin II (AII) generating pathways in resistance arteries from patients with chronic heart failure (CHF). BACKGROUND: Non-ACE dependent AII generation occurs in resistance arteries from normal volunteers. Inhibition of non-ACE dependent AII generation may have therapeutic potential in CHF. METHODS: Resistance arteries were dissected from gluteal biopsies from patients with coronary heart disease (CHD) and preserved left ventricular function and from patients with CHF. Using wire myography, concentration response curves to angiotensin I (AI) and AII were constructed in the presence of 1) vehicle, 2) chymostatin [an inhibitor of chymase], 3) enalaprilat, and 4) the combination of chymostatin and enalaprilat. RESULTS: In resistance arteries from patients with CHD, the vasoconstrictor response to AI was not inhibited by either inhibitor alone (chymostatin [p > or = 0.05] or enalaprilat [p > or = 0.05]) but was significantly inhibited by the combination (p < 0.001). In arteries from patients with CHF, AI responses were inhibited by enalaprilat (p < 0.05) but not by chymostatin alone (p > 0.05). The combination ofchymostatin and enalaprilat markedly inhibited the response to AI (p < 0.001) to a greater degree than enalaprilat alone (p < or = 0.01). CONCLUSIONS: Non-ACE dependent AII generating pathways exist in resistance arteries from patients with both CHF and CHD. In resistance arteries from patients with CHD, inhibition of either the ACE or chymase pathway alone has no effect on AII generation, and both pathways must be blocked before the vasoconstrictor action of AI is inhibited. In CHF, blockade of ACE results in marked inhibition of responses to AI, but this is enhanced by coinhibition of chymase. These studies suggest that full suppression of the renin angiotensin system cannot be achieved by ACE inhibition alone and provide a rationale for developing future therapeutic strategies. PMID- 11263609 TI - Preservation of venous endothelial function in the forearm venous capacitance bed of patients with chronic heart failure despite arterial endothelial dysfunction. AB - OBJECTIVES: The goal of this study was to assess whether endothelial dysfunction occurs in the forearm venous capacitance bed of patients with chronic heart failure (CHF) and to determine the role of nitric oxide (NO) in modulating venous tone. BACKGROUND: Control of venous tone is crucially important in CHF. More than 70% of blood volume lies in the venous capacitance beds. Therefore, small changes in venous tone may markedly affect cardiac filling pressures and cardiac output. METHODS: Venous tone was measured using radionuclide forearm venous plethysmography in 24 patients with CHF and 16 age-matched controls. The effect of basal NO activity on venous tone was assessed by infusing N-monomethyl-L arginine 12 mg/min and stimulated NO using carbachol 15 microg/min. Brachial artery flow-mediated dilation was assessed by ultrasonic wall-tracking. RESULTS: Blockade of basal NO release caused a significant and similar venoconstriction in patients (9.6 +/- 1.8%, p < 0.01) and controls (6.6 +/- 1.7%, p < 0.01). Carbachol-induced venodilation was significant and similar in patients (36.8 +/- 3.9%, p < 0.001) and controls (40.7 +/- 3.9%, p < 0.001). Brachial artery flow mediated dilation was impaired in patients compared with controls (2.0 +/- 0.6% vs. 7.5 +/- 2.5%, p < 0.01). CONCLUSIONS: Our data indicate that, despite marked impairment of the function of the arterial endothelium, there is preservation of both basal and stimulated NO release in the forearm venous capacitance bed. This may provide important insights into mechanisms of endothelial dysfunction in CHF and the potential for novel therapy. PMID- 11263610 TI - Echocardiography predicts embolic events in infective endocarditis. AB - OBJECTIVES: The aim of our study was to assess the value of transesophageal echocardiography (TEE) in predicting embolic events (EEs) in a large group of patients with definite endocarditis according to the Duke criteria, including silent embolism. BACKGROUND: The value of echocardiography in predicting embolism in patients with endocarditis remains controversial. Some studies reported an increased risk of embolism in patients with large and mobile vegetations, whereas other studies failed to demonstrate such a relationship. METHODS: Multiplane transesophageal echocardiograms of 178 consecutive patients with definite infective endocarditis (IE) were analyzed. The incidence of embolism was compared with the echocardiographic characteristics (localization, size and mobility) of the vegetations. To detect silent embolism, cerebral and thoraco-abdominal scans were performed in 95% of patients. RESULTS: Among 178 patients, 66 (37%) had one or more EEs. There was no difference between patients with and without embolism in terms of age, gender and left valve involved. On univariate analysis, Staphylococcus infection, right-side valve endocarditis and vegetation length and mobility were significantly related to EEs. A significant higher incidence of embolism was present in patients with vegetation length >10 mm (60%, p < 0.001) and in patients with mobile vegetations (62%, p < 0.001). Embolism was particularly frequent among 30 patients with both severely mobile and large vegetations (> 15 mm) (83%, p < 0.001). On multivariate analysis, the only predictors of embolism were vegetation length (p = 0.03) and mobility (p = 0.01). CONCLUSIONS: Our study shows that the presence of vegetations on TEE is predictive of embolism and that the morphologic characteristics of vegetations are helpful in predicting EEs in both mitral and aortic valve IE. It also suggests that early operation may be recommended in patients with vegetations > 15 mm and high mobility, irrespective of the degree of valve destruction, heart failure and response to antibiotic therapy. PMID- 11263611 TI - Echocardiography predicts embolic events in infective endocarditis. PMID- 11263612 TI - Cardiocyte cytoskeleton in patients with left ventricular pressure overload hypertrophy. AB - OBJECTIVES: We sought to determine whether the cardiocyte microtubule network densification characteristic of animal models of severe pressure overload cardiac hypertrophy occurs in human patients. BACKGROUND: In animal models of clinical entities causative of severe right and left ventricular (LV) pressure overload hypertrophy, increased density of the cellular microtubule network, through viscous loading of active myofilaments, causes contractile dysfunction that is normalized by microtubule depolymerization. These linked contractile and cytoskeletal abnormalities, based on augmented tubulin synthesis and microtubule stability, progress during the transition to heart failure. METHODS: Thirteen patients with symptomatic aortic stenosis (AS) (aortic valve area = 0.6 +/- 0.1 cm2) and two control patients without AS were studied. No patient had aortic insufficiency, significant coronary artery disease or abnormal segmental LV wall motion. Left ventricular function was assessed by echocardiography and cardiac catheterization before aortic valve replacement. Left ventricular biopsies obtained at surgery before cardioplegia were separated into free and polymerized tubulin fractions before analysis. Midwall LV fractional shortening versus mean LV wall stress in the AS patients was compared with that in 84 normal patients. RESULTS: Four AS patients had normal LV function and microtubule protein concentration; six had decreased LV function and increased microtubule protein concentration, and three had borderline LV function and microtubule protein concentration, such that there was an inverse relationship of midwall LV fractional shortening to microtubule protein. CONCLUSIONS: In patients, as in animal models of severe LV pressure overload hypertrophy, myocardial dysfunction is associated with increased microtubules, suggesting that this may be one mechanism contributing to the development of congestive heart failure in patients with AS. PMID- 11263613 TI - Pulmonary artery pulse pressure and wave reflection in chronic pulmonary thromboembolism and primary pulmonary hypertension. AB - OBJECTIVES: The purpose of this time-domain study was to compare pulmonary artery (PA) pulse pressure and wave reflection in chronic pulmonary thromboembolism (CPTE) and primary pulmonary hypertension (PPH). BACKGROUND: Pulmonary artery pressure waveform analysis provides a simple and accurate estimation of right ventricular afterload in the time-domain. Chronic pulmonary thromboembolism and PPH are both responsible for severe pulmonary hypertension. Chronic pulmonary thromboembolism and PPH predominantly involve proximal and distal arteries, respectively, and may lead to differences in PA pressure waveform. METHODS: High fidelity PA pressure was recorded in 14 patients (7 men/7 women, 46 +/- 14 years) with CPTE (n = 7) and PPH (n = 7). We measured thermodilution cardiac output, mean PA pressure (MPAP), PA pulse pressure (PAPP = systolic - diastolic PAP) and normalized PAPP (nPAPP = PPAP/MPAP). Wave reflection was quantified by measuring Ti, that is, the time between pressure upstroke and the systolic inflection point (Pi), deltaP, that is, the systolic PAP minus Pi difference, and the augmentation index (deltaP/PPAP). RESULTS: At baseline, CPTE and PPH had similar cardiac index (2.4 +/- 0.4 vs. 2.5 +/- 0.5 l/min/m2), mean PAP (59 +/- 9 vs. 59 +/- 10 mm Hg), PPAP (57 +/- 13 vs. 53 +/- 13 mm Hg) and nPPAP (0.97 +/- 0.16 vs. 0.89 +/- 0.13). Chronic pulmonary thromboembolism had shorter Ti (90 +/- 17 vs. 126 +/- 16 ms, p < 0.01) and higher deltaP/PPAP (0.26 +/- 0.01 vs. 0.09 +/- 0.07, p < 0.01). CONCLUSIONS: Our study indicated that: 1) CPTE and PPH with severe pulmonary hypertension had similar PA pulse pressure, and 2) wave reflection is elevated in both groups, and CPTE had increased and anticipated wave reflection as compared with PPH, thus suggesting differences in the pulsatile component of right ventricular afterload. PMID- 11263614 TI - Patients at lower risk of arrhythmia recurrence: a subgroup in whom implantable defibrillators may not offer benefit. Antiarrhythmics Versus Implantable Defibrillator (AVID) Trial Investigators. AB - OBJECTIVES: The goal of this study was to identify subgroups of arrhythmia patients who do not benefit from use of the implantable cardiac defibrillator (ICD). BACKGROUND: Treatment of serious ventricular arrhythmias has evolved toward more common use of the ICD. Since estimates of the cost per year of life saved by ICD therapy vary from $25,000 to perhaps $125,000, it is important to identify patient subgroups that do not benefit from the ICD. METHODS: Data for 491 ICD patients enrolled in the Antiarrhythmics Versus Implantable Defibrillators Study were used to create a hazards model relating baseline factors to time to first recurrent arrhythmia. The model was used to predict the hazard for recurrent arrhythmia among all trial patients. A priori cut points provided lower and higher recurrent arrhythmia risk strata. For each stratum the incremental years of life due to ICD versus antiarrhythmic drug therapy were calculated. RESULTS: Factors that predicted recurrent arrhythmia were: ventricular tachycardia as the index arrhythmia, history of cerebrovascular disease, lower left ventricular ejection fraction, a history of any tachyarrhythmia before the index event and the absence of revascularization after the index event. Survival times (over a follow-up of three years) were identical in each arm of the lowest risk sextile (survival advantage 0.03 +/- 0.12 [se] years), while the survival advantage for patients above the first sextile was 0.27 +/- 0.07 (se) years (two-sided p = 0.05). CONCLUSIONS: Patients presenting with an isolated episode of ventricular fibrillation in the absence of cerebrovascular disease or history of prior arrhythmia who have undergone revascularization or who have moderately preserved left ventricular function (left ventricular ejection fraction > 0.27) are not likely to benefit from ICD therapy compared with amiodarone therapy. PMID- 11263616 TI - Dofetilide: is the treatment worse than the disease? PMID- 11263615 TI - Pause-dependent polymorphic ventricular tachycardia during long-term treatment with dofetilide: a placebo-controlled, implantable cardioverter-defibrillator based evaluation. AB - OBJECTIVES: To compare the incidence of pause-dependent polymorphic ventricular tachycardia (PVT) in patients with implantable cardioverter-defibrillators (ICDs) randomly assigned to the QT-prolonging antiarrhythmic dofetilide or placebo. BACKGROUND: Drug-related torsade de pointes (TdP) is usually recognized within days of initiating therapy, but its incidence during long-term therapy is unknown. METHODS: We assessed the frequency of TdP and ICD electrograms compatible with TdP in a multicenter study that randomized ICD patients to placebo (n = 87) or dofetilide (n = 87). As reported elsewhere, the number of patients with a primary trial end point (ICD intervention for VT or ventricular fibrillation) was similar in the two groups. For this analysis, a qualifying event was TdP (on electrocardiogram) or an intracardiac electrogram showing pause dependent PVT. RESULTS: A total of 620 electrograms obtained in 131 patients were analyzed blindly by prospectively defined criteria for episodes of pause dependent polymorphic VT. These were identified in 15/87 (17%) patients receiving dofetilide and 5/87 (6%) patients on placebo (p < 0.05). Five of these episodes were early (<3 days), all of which were TdP on dofetilide. There were 15 late events, 10 on dofetilide and five on placebo (p = 0.29). The median time to a late event was 22 days (range 6 to 107 days) for dofetilide and 99 days (range 34 to 207 days) for placebo. CONCLUSIONS: Pause-dependent PVT was more common among patients receiving dofetilide, although total VT incidence was similar in the two groups. These data suggest that in ICD patients either long-term dofetilide therapy is associated with an increased risk of TdP or the drug alters VT morphology. PMID- 11263617 TI - Up-regulation of inositol 1,4,5 trisphosphate receptor expression in atrial tissue in patients with chronic atrial fibrillation. AB - OBJECTIVES: We examined whether patients with atrial fibrillation (AF) have alterations in atrial inositol 1,4,5 trisphosphate receptors (IP3 receptors). BACKGROUND: Abnormal intracellular Ca2+ homeostasis occurs in chronic AF. The intracellular Ca2+ concentration is regulated by ryanodine and IP3 receptors. We recently reported alterations in ryanodine receptors in atrial tissue from patients in chronic AF. METHODS: We analyzed IP3 receptor expression in the right atrial myocardium from 13 patients with mitral valvular disease (MVD) with AF (MVD/AF), five patients with MVD who had normal sinus rhythm (MVD/NSR) and eight control patients with NSR (tissue obtained during coronary artery bypass surgery). Hemodynamic and echocardiographic data were obtained preoperatively, and an immunohistochemical study was performed on atrial tissue. RESULTS: The relative expression level of IP3 receptor protein was significantly greater in MVD/AF (0.75 +/- 0.26) than it was in MVD/NSR (0.42 +/- 0.13, p < 0.01), and both were significantly above control (0.14 +/- 0.08). The relative expression level of IP3 receptor messenger RNA was significantly greater in the MVD/AF group (0.028 +/- 0.008) than it was in the control group (0.015 +/- 0.004, p < 0.01), but patients with MVD/AF did not differ from patients with MVD/NSR (0.020 +/- 0.006). The relative expression levels of IP3 receptor protein and messenger RNA were higher in patients with left atrial dimension > or = 40 mm, pulmonary capillary wedge pressure > or = 10 mm Hg and right atrial pressure > or = 5 mm Hg. Inositol 1,4,5 trisphosphate receptors were over-expressed in the cytosol and at the nuclear envelope of atrial myocytes in MVD. CONCLUSIONS: Since chronic mechanical overload of the atrial myocardium increased IP3 receptor expression, especially in patients with chronic AF, up-regulation of IP3 receptors may be important in modulating intracellular Ca2+ homeostasis and initiating or perpetuating AF. PMID- 11263618 TI - Ultrafast three-dimensional contrast-enhanced magnetic resonance angiography and imaging in the diagnosis of partial anomalous pulmonary venous drainage. AB - OBJECTIVES: The purpose of our study was to evaluate patients with suspected anomalous pulmonary veins (APVs) and atrial septal defects (ASDs) using fast cine magnetic resonance imaging (MRI) and ultrafast three-dimensional magnetic resonance angiography (MRA). BACKGROUND: Precise anatomic definition of anomalous pulmonary and systemic veins, and the atrial septum are prerequisites for surgical correction of ASDs. Cardiac catheterization and transesophageal echocardiography (TEE) are currently used to diagnose APVs, but did not provide complete information in our patients. METHODS: Twenty consecutive patients with suspected APVs were studied by MRA after inconclusive assessment by catheterization, TEE or both. The MRI images were acquired with a fast cine sequence and a novel ultrafast three-dimensional sequence before and after contrast injection. RESULTS: Partial anomalous pulmonary venous drainage was demonstrated in 16 of 20 patients and was excluded in four patients. Magnetic resonance imaging correctly diagnosed APVs and ASDs in all patients (100%) who underwent surgery. For the diagnosis of APVs, the MRI and catheterization results agreed in 74% of patients and the MRI and TEE agreed in 75% of patients. For ASDs, MRI agreed with catheterization and TEE in 53% and 83% of patients, respectively. CONCLUSIONS: Fast cine MRI with three-dimensional contrast-enhanced MRA provides rapid and comprehensive anatomic definition of APVs and ASDs in patients with adult congenital heart disease in a single examination. PMID- 11263619 TI - Dilated cardiomyopathy in isolated congenital complete atrioventricular block: early and long-term risk in children. AB - OBJECTIVES: We sought to identify the risk factors predicting the development of dilated cardiomyopathy (DCM) in patients with isolated congenital complete atrioventricular block (CCAVB). BACKGROUND: Recently evidence has emerged that a subset of patients with CCAVB develop DCM. METHODS: This was a retrospective study of 149 patients with CCAVB who had heart size and left ventricular (LV) function assessed by echocardiography and chest radiography over a follow-up period of 10 +/- 7 years. RESULTS: Nine patients developed DCM at the age of 6.5 +/- 5 years. No definite cause could be identified. In these nine patients, CCAVB was diagnosed in eight at 23 +/- 2.3 weeks gestation and in one at birth. Maternal SSA/SSB antibodies were confirmed in seven of the nine patients. Pacemakers were implanted in eight patients in the first month and in one patient at five years of age. The initial left ventricular end-diastolic dimension (LVEDD) was in the 96th +/- 2.6 percentile and the cardiothoracic (CT) ratio was 64 +/- 3.8% in the nine patients who developed DCM, and differed significantly in patients with CCAVB (p < 0.005) who did not develop DCM. The LVEDD and CT ratio did not decrease in the patients with CCAVB and DCM, but decreased significantly in the patients with CCAVB without DCM (p < 0.001) once pacing was initiated. Two patients with DCM died within two months of diagnosis; one patient is neurologically compromised; two patients received a heart transplant; and four patients are listed for heart transplantation. CONCLUSIONS: Isolated CCAVB is associated with a long-term risk for the development of DCM. Risk factors may be SSA/SSB antibodies, increased heart size at initial evaluation and the absence of pacemaker-associated improvement. PMID- 11263620 TI - Quantification of renal blood flow with contrast-enhanced ultrasound. AB - OBJECTIVES: The goal of this study was to determine the ability of contrast enhanced ultrasound (CEU) to quantify renal tissue perfusion. BACKGROUND: The kinetics of tracers used to assess renal perfusion are often complicated by countercurrent exchange, tubular transport or glomerular filtration. We hypothesized that, because gas-filled microbubbles are pure intravascular tracers with a rheology similar to that of red blood cells, CEU could be used to quantify renal tissue perfusion. METHODS: During a continuous venous infusion of microbubbles (SonoVue), regional renal perfusion was quantified in nine dogs using CEU by destroying microbubbles and measuring their tissue replenishment with intermittent harmonic imaging. Both renal blood volume fraction and microbubble velocity were derived from pulsing-interval versus video-intensity plots. The product of the two was used to calculate renal nutrient blood flow. Renal arterial blood flow was independently measured with ultrasonic flow probes placed directly on the renal artery and was increased using dopamine and decreased by placement of a renal artery stenosis. RESULTS: An excellent correlation was found between cortical nutrient blood flow using microbubbles and ultrasonic flow probe-derived renal blood flow (r = 0.82, p < 0.001) over a wide range (2.5 fold) of flows. CONCLUSIONS: Ultrasound examination during microbubble infusion can be used to quantify total organ as well as regional nutrient blood flow to the kidney. PMID- 11263621 TI - Regional asynchrony during acute myocardial ischemia quantified by ultrasound strain rate imaging. AB - OBJECTIVES: We propose a new method to easily quantify asynchronous wall motion due to postsystolic shortening (PSS). We also studied the relationship of the spatial and temporal extent of PSS to the extent of myocardium at ischemic risk after variable duration of ischemia. BACKGROUND: Postsystolic shortening is a sensitive marker of asynchrony during ischemia. Current techniques for detection of asynchrony are either subjective, or invasive and time-consuming. Strain rate imaging (SRI) can noninvasively depict PSS as prolonged compression/expansion crossover. METHODS: Nineteen open-chest pigs were scanned from apical views, before and after left anterior descending coronary artery occlusion. Strain rates were derived offline from tissue Doppler velocity cineloops. The time from electrocardiographic R-wave to the occurrence of compression/expansion crossover (TCEC) was calculated. Prolonged TCEC during ischemia was identified using a standardized analysis and both spatial (% of left ventricle) and temporal extent were quantified. The extent of myocardium at risk was measured in seven animals from dye-stained specimens. RESULTS: Prolonged TCEC was found in all ischemic segments. There was a good correlation (r = 0.91; p < 0.001) and good agreement between the spatial distributions of prolonged TCEC and myocardium at risk. The extent of myocardium at risk was better approximated by TCEC measurement (36 +/- 7% vs. 39 +/- 8%, respectively; p = NS) than by wall motion analysis (47 +/- 17%, p < 0.05). The duration of occlusion did not prolong TCEC. CONCLUSIONS: Prolonged TCEC consistently occurs in ischemic myocardium and is apparently not affected by the duration of ischemia. Standardized analysis of TCEC in SRI closely quantifies the extent of ischemic myocardium. This new method may be a useful tool in other cardiac conditions associated with regional diastolic asynchrony. PMID- 11263623 TI - President's page: cardiologists under stress: the response of the American College of Cardiology. PMID- 11263622 TI - Atherosclerotic aortic component quantification by noninvasive magnetic resonance imaging: an in vivo study in rabbits. AB - OBJECTIVES: We sought to demonstrate the ability that noninvasive in vivo magnetic resonance imaging (MRI) has to quantify the different components within atherosclerotic plaque. BACKGROUND: Atherosclerotic plaque composition plays a critical role in both lesion stability and subsequent thrombogenicity. Noninvasive MRI is a promising tool for the characterization of plaque composition. METHOD: Thoracic and abdominal aortic atherosclerotic lesions were induced in rabbits (n = 5). Nine months later, MRI was performed in a 1.5T system. Fast spin-echo sequences (proton density-weighted and T2-weighted [T2W] images) were obtained (in-plane resolution: 350 x 350 microns, slice thickness: 3 mm). Magnetic resonance images were correlated with matched histopathological sections (n = 108). RESULTS: A significant correlation (p < 0.001) was observed for mean wall thickness and vessel wall area between MRI and histopathology (r = 0.87 and r = 0.85, respectively). The correlation was also present on subanalysis of the thoracic and upper part of the abdominal aorta, susceptible to respiratory motion artifacts. There was a significant correlation for plaque composition (p < 0.05) between MRI and histopathology for the analysis of lipidic (low signal on T2W, r = 0.81) and fibrous (high signal on T2W, r = 0.86) areas with Oil Red O staining. T2-weighted images showed greater contrast than proton density-weighted between these different components of the plaques as assessed by signal intensity ratio analysis with the mean difference in signal ratios of 0.47 (S.E. 0.012, adjusted for clustering of observations within lesions) being significantly different from 0 (t1 = 39.1, p = 0.016). CONCLUSIONS: In vivo noninvasive high resolution MRI accurately quantifies fibrotic and lipidic components of atherosclerosis in this model. This may permit the serial analysis of therapeutic strategies on atherosclerotic plaque stabilization. PMID- 11263624 TI - Pulse pressure--a review of mechanisms and clinical relevance. AB - The goal of this study was to review the origin, clinical relevance and treatment of pulse pressure (PP). Elevated PP is increasingly being recognized as a risk factor for cardiovascular, particularly coronary, disease. Pulse pressure is discussed in terms of both Windkessel and distributive models of the arterial circulation. Pulse pressure arises from the interaction of cardiac ejection (stroke volume) and the properties of the arterial circulation. An increased stiffness of the aorta and large arteries leads to an increase in PP through a reduction in arterial compliance and effects on wave reflection. A number of factors are known to influence arterial wall behavior and, therefore, PP. In addition to the effects of aging and blood pressure on arterial wall elasticity, there is some evidence that atherosclerosis, per se, amplifies these effects. Thus, the relationship between PP and coronary disease may be bidirectional. A number of dietary and lifestyle interventions have been shown to modify large artery behavior. These include aerobic exercise training and consumption of n-3 fatty acids. Conversely, strength training is associated with an increase in arterial stiffness and a higher PP. The effects of antihypertensive medication have been extensively studied, but many studies are difficult to interpret because of concomitant change in blood pressure, and to a lesser degree, heart rate. However a number of studies do suggest direct arterial wall effects, particularly for angiotensin-converting enzyme inhibitors. A distributed compliance model of the arterial circulation provides a framework for understanding the causes, effects and potential treatment of elevations in PP. PMID- 11263626 TI - Development and validation of the Ontario acute myocardial infarction mortality prediction rules. AB - OBJECTIVES: To develop and validate simple statistical models that can be used with hospital discharge administrative databases to predict 30-day and one-year mortality after an acute myocardial infarction (AMI). BACKGROUND: There is increasing interest in developing AMI "report cards" using population-based hospital discharge databases. However, there is a lack of simple statistical models that can be used to adjust for regional and interinstitutional differences in patient case-mix. METHODS: We used linked administrative databases on 52,616 patients having an AMI in Ontario, Canada, between 1994 and 1997 to develop logistic regression statistical models to predict 30-day and one-year mortality after an AMI. These models were subsequently validated in two external cohorts of AMI patients derived from administrative datasets from Manitoba, Canada, and California, U.S. RESULTS: The 11-variable Ontario AMI mortality prediction rules accurately predicted mortality with an area under the receiver operating characteristic (ROC) curve of 0.78 for 30-day mortality and 0.79 for one-year mortality in the Ontario dataset from which they were derived. In an independent validation dataset of 4,836 AMI patients from Manitoba, the ROC areas were 0.77 and 0.78, respectively. In a second validation dataset of 112,234 AMI patients from California, the ROC areas were 0.77 and 0.78 respectively. CONCLUSIONS: The Ontario AMI mortality prediction rules predict quite accurately 30-day and one year mortality after an AMI in linked hospital discharge databases of AMI patients from Ontario, Manitoba and California. These models may also be useful to outcomes and quality measurement researchers in other jurisdictions. PMID- 11263627 TI - Measuring the effectiveness of medical care delivery. PMID- 11263625 TI - Stenting versus thrombolysis in acute myocardial infarction trial (STAT). AB - OBJECTIVES: We sought to directly compare primary stenting with accelerated tissue plasminogen activator (t-PA) in patients presenting with acute ST elevation myocardial infarction (AMI). BACKGROUND: Thrombolysis remains the standard therapy for AMI. However, at some institutions primary angioplasty is favored. Randomized trials have shown that primary angioplasty is equal or superior to thrombolysis, while recent studies demonstrate that stent implantation improves the results of primary angioplasty. METHODS: Patients presenting with AMI were randomly assigned to primary stenting (n = 62) or accelerated t-PA (n = 61). The primary end point was the composite of death, reinfarction, stroke or repeat target vessel revascularization (TVR) for ischemia at six months. RESULTS: The primary end point was significantly reduced in the stent group compared with the accelerated t-PA group, 24.2% versus 55.7% (p < 0.001). The event rates for other outcomes in the stent group versus the t-PA group were as follows: mortality: 4.8% versus 3.3% (p = 1.00); reinfarction: 6.5% versus 16.4% (p = 0.096); stroke: 1.6% versus 4.9% (p = 0.36); recurrent unstable ischemia: 9.7% versus 26.2% (p = 0.03) and repeat TVR for ischemia: 14.5% versus 49.2% (p < 0.001). The median length of the initial hospitalization was four days in the stent group and seven days in the t-PA group (p < 0.001). CONCLUSIONS: Compared with accelerated t-PA, primary stenting reduces death, reinfarction, stroke or repeat TVR for ischemia. In centers where facilities and experienced interventionists are available, primary stenting offers an attractive alternative to thrombolysis. PMID- 11263628 TI - Preparing staff to enhance active participation of adults with severe disabilities by offering choice and prompting performance during a community purchasing activity. AB - The purpose of this study was to evaluate the effects of a multicomponent staff training package on the number of choice responses and performance responses made by adults with disabilities in a community purchasing activity. The multicomponent training package included an inservice (written manuals, a verbal explanation of the information, role play activities, and video examples) performance feedback sessions in the community context, and self-monitoring instruction. Primary data were collected on how each staff offered choices and prompted performance with an individual with severe disabilities in three different fast food restaurants per week. Secondary data were collected on the number of choices individuals with disabilities made and the level of their performance during a purchase in a fast food restaurant. The findings showed that all four of the staff did not give opportunities for choices and used intrusive prompting or performed the skill for the person in baseline, but mastered these skills the first probe after the training sessions. In addition, the staff generalized offering choices and prompting performance across settings and adults with disabilities and maintained the skills. Also, the adults with disabilities increased the number of choice responses they made as well as their level of performance (compared to baseline) after the staff received the intervention. PMID- 11263629 TI - Use of microswitches and speech output systems with people with severe/profound intellectual or multiple disabilities: a literature review. AB - Microswitches and speech output systems are two forms of technology which have been used with people with severe/profound intellectual or multiple disabilities to help them reduce their isolation and interact with the surrounding world (i.e., thus obtaining environmental stimulation independently or requesting it efficiently). This paper reviews the studies which used microswitches and speech output systems with the aforementioned people during the 1986-1999 period, and discusses the research findings and the practicality of these two forms of technology. Some relevant issues for future research are also pointed out. PMID- 11263630 TI - Self-management of instruction cues for occupation: review of studies with people with severe and profound developmental disabilities. AB - Helping people with severe and profound developmental disabilities acquire and maintain constructive occupation is an objective of great practical importance. During the last 15-20 years, studies directed at this goal have largely relied on five strategies of self-management of instruction cues. Those strategies consist of the use of (1) picture cues presented on sets of cards, (2) picture cues stored in computer-aided systems, (3) object cues attached to cards, (4) verbal cues stored in audio recording devices, and (5) self-verbalizations. This paper reviews the aforementioned strategies and discusses their overall effectiveness and their suitability (practicality). The paper also points out some relevant issues for future research. PMID- 11263631 TI - Predicting the persistence of severe self-injurious behavior. AB - Information was collected on 95 people with mental retardation who had been identified seven years previously as showing severe self-injurious behavior. At follow up 71% of participants were still showing self-injurious behavior of a severity which presented a management problem for care staff. The occurrence of specific topographies of self-injury was extremely stable among the group showing persistent self-injury. Finally, self-injury status at follow-up was predicted with 76% accuracy by a logistic regression model containing three variables: site of injury (higher persistence being shown by people exhibiting head directed self injury); reported (greater) stability of self-injury when first identified; and (younger) age. PMID- 11263632 TI - The prevalence of challenging behaviors: a total population study. AB - A total population study was undertaken in two areas of England to identify the situation and characteristics of people reported to exhibit challenging behaviors. We found that: (1) challenging behaviors are shown by 10-15% of people with mental retardation who are in contact with educational, health or social care services for people with mental retardation; (2) the most common forms of challenging behaviors reported were 'other' behavior (shown by 9%-12% of all people screened), aggression (7%), destructive behavior (4%-5%) and self-injury (4%); (3) the majority of people identified showed two or more of these four general forms of challenging behavior; (4) approximately two-thirds of the people identified were boys/men; (5) close to two-thirds of the people identified were adolescents or young adults; (6) approximately 50% of the people identified as showing more demanding challenging behavior were living with their families; (7) people who showed more demanding challenging behavior were more likely to need greater levels of assistance in eating, dressing and washing, be incontinent and have more restricted expressive and receptive communication. PMID- 11263633 TI - Overnight delay in the reduction of supracondylar fractures of the humerus in children. PMID- 11263634 TI - The effect of surgical timing on the perioperative complications of treatment of supracondylar humeral fractures in children. AB - BACKGROUND: The purpose of this study was to evaluate the perioperative complication rates associated with early surgical treatment (eight hours or less following injury) and delayed surgical treatment (more than eight hours following injury) of displaced supracondylar humeral fractures in children. METHODS: Fifty two patients had early surgical treatment and 146 patients had delayed surgical treatment of a displaced supracondylar humeral fracture. The perioperative complication rates of the two groups were compared with the use of bivariate and multivariate statistical methods. RESULTS: There was no significant difference between the two groups with respect to the need for conversion to formal open reduction and internal fixation (p = 0.56), pin-track infection (p = 0.12), or iatrogenic nerve injury (p = 0.72). No compartment syndromes occurred in either group. Power analysis revealed that our study had an 86% power to detect a 20% difference between the two groups if one existed. CONCLUSIONS: We were unable to identify any significant difference, with regard to perioperative complication rates, between early and delayed treatment of displaced supracondylar humeral fractures. Within the parameters outlined in our study, we think that the timing of surgical intervention can be either early or delayed as deemed appropriate by the surgeon. PMID- 11263635 TI - Arthroscopic resection of the distal aspect of the clavicle with concomitant subacromial decompression. AB - BACKGROUND: Arthroscopic subacromial decompression and arthroscopic resection of the acromioclavicular joint as separate procedures have been well documented. However, there is little information on the success rate of resection with concomitant decompression. In this study, we retrospectively evaluated the results of a consecutive group of patients who underwent arthroscopic resection of the acromioclavicular joint with concomitant subacromial decompression. METHODS: We evaluated the surgical results in thirty-one consecutive patients (thirty-two shoulders) with acromioclavicular pathology with concomitant subacromial impingement. The mean age of the patients at the time of surgery was thirty-six years (range, eighteen to sixty-seven years). Twenty-five patients, including four professional athletes, were actively involved in sports activities. The mean duration of follow-up was four years and ten months (range, three to eight years). The follow-up examination included clinical evaluation, chart review, radiographic analysis, and isokinetic testing of both upper extremities. RESULTS: Of the twenty-five patients who participated in sports, twenty-two (including the four professional athletes) returned to their previous level of sports activity. Twenty-six patients had no pain, three reported mild pain on strenuous repetitive overhead activity, two (both weight-lifters) had occasional pain in the acromioclavicular joint and the lateral aspect of the shoulder with bench-pressing, and two (both baseball players) had mild pain in the posterior aspect of the shoulder with throwing. All of the patients were satisfied with the results. In the absence of a complete rotator cuff tear, isokinetic strength-testing of both upper extremities failed to demonstrate any weakness of the involved shoulder. The mean functional score for individual activities was 2.7 points (range, 2.1 to 3.0 points) preoperatively and 3.9 points (range, 3.6 to 4.0 points) postoperatively (p = 0.0001). No patient had superior migration of the clavicle. The amount of distal clavicular resection averaged 9 mm (range, 7 to 15 mm). One patient had heterotopic ossification at the resection site, with mild pain on direct palpation of the acromioclavicular joint and on strenuous overhead activity. Five patients had calcification at the anterior deltoid insertion into the acromion that was asymptomatic, with no impingement on overhead activity and no pain on direct palpation. CONCLUSIONS: We found excellent results with arthroscopic resection of the acromioclavicular joint and concomitant subacromial decompression. When this procedure is performed on properly selected patients, the results are similar to those of an open approach. PMID- 11263636 TI - Prolonged enoxaparin therapy to prevent venous thromboembolism after primary hip or knee replacement. Enoxaparin Clinical Trial Group. AB - BACKGROUND: Patients undergoing hip or knee joint replacement are at risk for venous thromboembolic complications for up to twelve weeks postoperatively. We evaluated the efficacy and safety of a prolonged post-hospital regimen of enoxaparin, a low-molecular-weight heparin, in this patient population. METHODS: Following elective total hip or knee replacement, 968 patients received subcutaneous enoxaparin (30 mg twice daily) for seven to ten days, and 873 were then randomized to receive three weeks of double-blind outpatient treatment with either enoxaparin (40 mg once daily) or a placebo. The primary efficacy end point was the prevalence of objectively confirmed venous thromboembolism or symptomatic pulmonary embolism during the double-blind phase of treatment. RESULTS: Of the 873 randomized patients, 435 underwent elective total hip replacement and 438 underwent elective total knee replacement. Enoxaparin was superior to the placebo in reducing the prevalence of venous thromboembolism in patients treated with hip replacement: 8.0% (eighteen) of the 224 patients treated with enoxaparin had venous thromboembolism compared with 23.2% (forty-nine) of the 211 patients treated with the placebo (p < 0.001; odds ratio, 3.62; 95% confidence interval, 2.00 to 6.55; relative risk reduction, 65.5%). Enoxaparin had no significant benefit in the patients treated with knee replacement: thirty-eight (17.5%) of the 217 patients treated with enoxaparin had venous thromboembolism compared with forty-six (20.8%) of the 221 patients treated with the placebo (p = 0.380; odds ratio, 1.24; 95% confidence interval, 0.76 to 2.02; relative risk reduction, 15.9%). Symptomatic pulmonary embolism developed in three patients, one with a hip replacement and two with a knee replacement; all had received the placebo. There was no significant difference in the prevalence of hemorrhagic episodes or other types of toxicity between the enoxaparin and placebo-treated groups. CONCLUSIONS: Prolonging enoxaparin thromboprophylaxis following hip replacement for a total of four weeks provided therapeutic benefit, by reducing the prevalence of venous thromboembolism, without compromising safety. A similar benefit was not observed in patients treated with knee replacement. PMID- 11263637 TI - Proximal femoral allografts for reconstruction of bone stock in revision arthroplasty of the hip. A nine to fifteen-year follow-up. AB - BACKGROUND: Revision of a femoral component in a patient who has severe bone loss is a complex problem that is likely to increase with the increasing numbers of patients who have multiple revision hip arthroplasties. A valuable option in such a situation is use of a long-stem prosthesis that is cemented to a proximal femoral allograft but not to the host bone. METHODS: Between April 1984 and December 1989, sixty-three total hip arthroplasties in sixty consecutive patients were revised with a proximal femoral allograft-prosthesis construct. The average length of the allograft was 15 cm. The average age of the patients at the time of the revision was 62.5 years. All patients had undergone at least one previous total hip arthroplasty, and an average of 3.8 previous total hip arthroplasties had been performed in the series. Each patient was assigned a modified Harris hip score. Radiographs were examined for trochanteric union, allograft-host union, endosteal and periosteal resorption, component loosening, and fracture. RESULTS: At an average of eleven years (range, nine years and four months to fifteen years) after the revision, forty-five patients were alive, fourteen patients had died, and one patient had been lost to follow-up. The patients who had died or had been lost to follow-up had had a total of fifteen allografts (24%) and had been followed for an average of five years and seven months (range, two years and four months to eight years). The average preoperative Harris hip score for the sixty-three hips was 30 points (range, 6 to 65 points). At the latest follow-up evaluation, the average score for the hips with the original graft in situ was 71 points (range, 47 to 95 points). Five hips failed because of infection, and four of them were successfully revised. Three hips failed because of aseptic loosening, at an average of ten years and three months; two were successfully revised, and the third was awaiting revision at the time of writing. An additional operation was performed in three hips with allograft-host nonunion and in two with dislocation. Success was defined as a postoperative increase in the Harris hip score of greater than 20 points, a stable implant, and no need for additional surgery related to the allograft at the time of the review. The success rate for all hips was 78% (forty-nine of sixty-three) after an average of nine years of follow-up. The success rate for the patients who were alive at the time of follow-up was 77% (thirty-seven of forty-eight hips) after an average of eleven years of follow-up. CONCLUSIONS: The clinical and radiographic results at an average of eleven years after revision hip arthroplasty with a proximal femoral allograft are encouraging. This report represents our early experience; improvements in the technique have been made. We believe that this technique provides a viable option for treatment of the difficult problem of severe femoral bone loss. PMID- 11263638 TI - The use of calcium sulfate in the treatment of benign bone lesions. A preliminary report. PMID- 11263639 TI - Primary cementless total hip arthroplasty in octogenarians. Two to eleven-year follow-up. AB - BACKGROUND: Cementless total hip arthroplasty is an accepted alternative to total hip arthroplasty with cement in younger patients, but it remains controversial for elderly patients. The purpose of this study was to evaluate the clinical and radiographic outcomes of cementless total hip arthroplasty with use of a proximally coated stem in patients who were at least eighty years of age at the time of the operation. METHODS: One hundred and twenty-three cementless total hip replacements were performed for the treatment of osteoarthritis in 114 patients between the ages of eighty and eighty-nine years. Seven patients (eight hips) died within two years after the surgery, seventeen patients (eighteen hips) died more than two years postoperatively but were not followed for at least two years, and five hips were lost to follow-up; this left ninety-two hips in eighty-six patients for review. The mean duration of follow-up was five years (range, two to eleven years). For the clinical evaluation, the Charnley modification of the Merle d'Aubigne and Postel scale was used. In addition, preoperative and postoperative Harris hip scores were available for sixty-nine hips. Seventy-eight hips were followed radiographically for two years or more. The focus of the radiographic evaluation was the status of the fixation of the femoral and acetabular components as well as cup wear. RESULTS: Perioperative medical complications occurred in association with 24% (thirty) of the 123 operations, but there were no deaths. The mean Charnley scores for pain and function for the ninety-two hips that were followed clinically for at least two years improved by 3.0 and 1.4 points, respectively. The sixty-nine hips for which preoperative and postoperative Harris hip scores were available had a mean improvement of 42 points, with a mean score of 82 points at the last follow-up evaluation. Mild thigh pain was present in four patients, but it did not limit their activity. There were no femoral component revisions. All of the femoral components were radiographically stable and had bone ingrowth. No acetabular component failed by loosening, but 41% (thirty) of the seventy-three hips with radiographs available for measurement of wear showed polyethylene wear. Of the seventy-eight cups that were followed radiographically for two years or more, 4% (three) were associated with lysis, but none had been revised. CONCLUSIONS: Cementless fixation in the elderly is safe, effective, and durable at the time of two to eleven-year follow up. PMID- 11263641 TI - Six-pin halo fixation and the resulting prevalence of pin-site complications. AB - BACKGROUND: In spite of the many advances in halo application technique, the prevalence of complications associated with the use of halo fixation remains high, particularly at the pin sites. Many practitioners do not use more than four pins for halo application in adults because they believe that it increases the risk of complications. The purpose of this study was to investigate the use of six pins in halo application, in order to determine if the extra pins increased fixation strength without increasing the overall pin-site complication rate. METHODS: The first part of our study consisted of force-deflection tests conducted on models of the skull fitted with either a four or a six-pin halo to determine if the six-pin halo provided greater fixation strength. Each skull model was placed in a servocontrolled hydraulic test machine; an axial distraction force was then applied until failure occurred. The second part of the study was a retrospective analysis of sixty-three patient records to document the prevalence of pin-site complications in patients treated with a six-pin halo system; these findings were then compared with established complication rates associated with four-pin halos. RESULTS: In the force-deflection tests, the mean load to failure of the six-pin halo construct (2879 N [647 lb]) showed the system to be significantly stronger (p = 0.0033) than the four-pin halo construct (1681 N [378 lb]). Of the sixty-three patient records reviewed, five (8% [95% confidence interval, 1% to 15%]) revealed pin-loosening; no infection was recorded for these five patients. One of the sixty-three patients had redness and erythema at "multiple sites," but these areas healed well. Another presented with infection at all six sites; this was recorded as an allergic reaction. CONCLUSIONS: Six-pin halo fixation results in greater halo strength and cervical spine stabilization without increasing the risk of pin-site complications. CLINICAL RELEVANCE: Our findings are relevant for current clinical practice as the high complication rates associated with halo application have deterred some practitioners from using this type of fixation. The use of six pins, along with an improved protocol for halo application and care, may contribute to a more successful treatment outcome with fewer complications. PMID- 11263640 TI - Adenoviral delivery of LIM mineralization protein-1 induces new-bone formation in vitro and in vivo. AB - BACKGROUND: The LIM mineralization protein-1 (LMP-1) gene encodes for an intracellular protein that induces the expression of several bone growth factors. The purpose of the present study was to determine the feasibility and the optimal dose of adenoviral delivery of the LMP-1 cDNA to promote spinal fusion. METHODS: A replication-deficient human recombinant adenovirus was constructed with the LMP 1 cDNA driven by a cytomegalovirus promoter. In phase 1, an in vitro dose response experiment was performed to determine the optimal adenovirus-LMP-1 (AdLMP-1) concentration and infection time. In phase 2, nine rabbits had a single level posterolateral arthrodesis of the lumbar spine with implantation of a carrier matrix loaded with bone-marrow-derived buffy-coat cells that had been infected for ten minutes with adenovirus containing the cDNA for LMP-1 (AdLMP-1) or beta-galactosidase (AdBgal). In phase 3, posterolateral arthrodesis of the spine was performed with implantation of cells infected with AdLMP-1 (ten rabbits) or cells infected with an empty adenovirus that did not contain LMP-1 cDNA (ten rabbits) and the results were compared. In this phase, peripheral-blood derived buffy-coat cells were used instead of bone-marrow-derived cells and a collagen-ceramic-composite sponge was used as the carrier. RESULTS: In phase 1, the in vitro dose-response experiment showed that a multiplicity of infection of 0.25 plaque-forming units per cell was the most efficient dose. In phase 2, the implants that had received cells infected with AdLMP-1 induced a solid, continuous spinal fusion mass at five weeks. In contrast, the implants that had received cells infected with AdBgal or a lower dose of AdLMP-1 induced little or no bone formation. In phase 3, a solid spinal fusion was observed at four weeks in all ten rabbits that had received cells infected with AdLMP-1 and in none of the ten rabbits that had received cells infected with the empty adenovirus. Biomechanical and histological testing of the AdLMP-1-treated specimens revealed findings that were consistent with a high-quality spinal fusion. CONCLUSIONS: Adenoviral delivery of LMP-1 cDNA promotes spinal fusion in immune-competent rabbits. PMID- 11263642 TI - Transtibial amputees from the Vietnam War. Twenty-eight-year follow-up. AB - BACKGROUND: The long-term functional outcome following lower-extremity amputation is not well documented. I ascertained the functional outcome and health status of patients who had sustained a unilateral transtibial amputation as a result of a battlefield injury. METHODS: The records of 123 patients who had been treated at Valley Forge Army General Hospital during the Vietnam War for a diagnosis of isolated transtibial amputation due to a battlefield injury were reviewed. Group 1 had an isolated transtibial amputation, and Group 2 had at least one other major injury (another major long-bone fracture of the lower extremity, burns covering >20% of the body surface area, or a chest, abdominal, face, or head wound) in addition to the transtibial amputation. Seventy-two (59%) of the patients were enrolled in the study: twenty-eight were in Group 1 and forty-four, in Group 2. Data were collected about employment status, marital status, whether the patient had children, and use of psychological support services. The Short Form-36 (SF-36) health survey was used to compare Group 1 and Group 2, individually and combined, with age and gender-matched controls. Scaled scores for the two groups (control and amputation) were compared with use of the Student t test (two-tailed). RESULTS: Tripping a land mine or booby trap caused 65% of the injuries. The average age at the time of follow-up was forty-eight years. The average time to follow-up was twenty-eight years. Only the prevalence of the use of psychological support services differed significantly between Groups 1 and 2 (21% compared with 50%; p = 0.015). The results of the SF-36 health survey for Groups 1 and 2 were 81.6 and 58.2, respectively, for physical function, 82.7 and 33.1 for role physical, 81.4 and 50.9 for bodily pain, 74.1 and 58.7 for general health, 67.1 and 51.5 for vitality, 89.1 and 70.4 for social function, 88.1 and 56.0 for role emotional, and 79.5 and 64.0 for mental health. The average scaled scores for Group 1 were similar to those for the age and gender-matched controls, but the scores for Group 2 were significantly lower (p < or = 0.001) than those for the age and gender-matched controls in all categories. CONCLUSIONS: Group-1 patients led relatively normal lives after sustaining a transtibial amputation in battle. The addition of another major injury (Group 2) appears to have significant long-term consequences with regard to SF-36 scores and the need for psychological care. PMID- 11263643 TI - Tibial post wear in posterior stabilized total knee arthroplasty. An unrecognized source of polyethylene debris. AB - BACKGROUND: With extensive use of posterior stabilized total knee arthroplasty implants, it is increasingly important to assess the mechanical performance of this design alternative. The purpose of this study was to examine the wear patterns at the femoral cam-tibial post interface in a series of posterior stabilized prostheses retrieved at revision arthroplasty. METHODS: Qualitative and quantitative wear analysis was performed over the surface of the stabilizing posts from twenty-three retrieved total knee components that had been implanted for a mean of 35.6 months (range, 2.3 to 107.2 months). The implants were designs from four different manufacturers. Digital images of the anterior, posterior, medial, and lateral surfaces of the tibial post were made for quantitative analysis and determination of a post wear score. Wear was characterized with a grading system that isolates adhesive, abrasive, and fatigue wear, inferring a weighted score from an estimation of generated polyethylene debris. RESULTS: Evidence of wear or damage was observed on all twenty-three of the stabilizing posts, including those revised because of infection. On the average, 39.9% (range, 18.5% to 60%) of the post surface demonstrated some form of deformation, with adhesive wear, or burnishing, being the predominant wear mechanism. Seven posts (30%) exhibited severe damage with gross loss of polyethylene. The wear caused premature failure and early revision of two components: one of these failures was related to isolated post wear and the other, to severe post wear and subsequent fracture. Overall, wear was primarily posterior, but wear over the anterior, medial, and lateral surfaces was also notable. CONCLUSIONS: The cam post articulation in posterior stabilized implants can be an additional source of polyethylene wear debris. The variability in wear patterns observed among designs may be due to differences in cam-post mechanics, post location, and post geometry. The surgeon should be aware that the cam-post interface is not an innocuous articulation, and manufacturers should be motivated to produce implants that maintain the function of the post while limiting wear and surface damage. PMID- 11263644 TI - Loosening and osteolysis with the press-fit condylar posterior-cruciate substituting total knee replacement. AB - BACKGROUND: Aseptic loosening and osteolysis are rarely associated with cemented posterior-cruciate-substituting total knee replacements. Consequently, there is a paucity of information on this topic. METHODS: After a mean follow-up interval of fifty-six months (range, thirty-seven to eighty-nine months), sixteen (2.9%) of 557 posterior-cruciate-substituting primary total knee replacements were revised by a single surgeon because of loosening and osteolysis. Clinical, radiographic, and retrieval analyses were conducted to determine the mechanism of loosening and to identify associated risk factors. RESULTS: All sixteen knees (fifteen patients) were rated as good or excellent at one year after the primary replacement, with mean clinical and functional Knee Society scores of 95 and 86 points, respectively. Nine of the fifteen patients who had a revision because of loosening and osteolysis had had a total knee arthroplasty on the contralateral side compared with only 18% of the patients who did not have a revision (p = 0.026). No evidence of transmission of substantial anteroposterior stresses from the posterior-cruciate-substituting mechanism was found. All twelve retrieved knee implants, however, had damage to the lateral and medial side walls of the polyethylene posterior-cruciate-substituting post. Damage to the inferior surface of the polyethylene inserts had a rotational pattern, with the axis of rotation in the medial compartment. Surface damage in a rotational pattern was also present on the superior and inferior surfaces of the titanium tibial base-plates. CONCLUSIONS: In the knees in our study, rotational forces were generated by impingement of the side walls of the intercondylar box on the polyethylene post. Such box-post impingement can occur throughout the range of motion. Rotational stresses are transmitted to the modular interfaces and to the metal-cement interfaces, resulting in loosening and osteolysis. A reduction in rotational constraint would be desirable. Patients with bilateral total knee replacement may be at increased risk for this type of loosening. PMID- 11263645 TI - The use of structural allograft for uncontained defects in revision total knee arthroplasty. A minimum five-year review. AB - BACKGROUND: To our knowledge, the medium to long-term outcome after revision knee arthroplasty with structural allograft augmentation for reconstruction of uncontained defects has not been determined. The purpose of the present study was to assess the outcome for patients managed with such a procedure. METHODS: We prospectively followed fifty patients who had fifty-two revision knee replacements with sixty-six structural grafts performed at three institutions. Twenty-nine knees (twenty-seven patients) were independently evaluated at a mean of 96.9 months (range, sixty to 189 months) by an investigator who had not been involved in the index procedure. Twelve knees (23%) had a repeat revision at a mean of 70.7 months (range, twenty-six to 157 months). The allograft was retained in two of these patients. Eleven patients died at a mean of ninety-three months (range, sixty-one to 128 months) after the procedure; the structural allograft and implants were intact, and the patients were not awaiting revision at the time of death. RESULTS: Clinical evaluation revealed that the mean modified Hospital for Special Surgery knee score had improved from 32.5 points preoperatively to 75.6 points at the time of the review and the mean range of motion had increased from 60.5 degrees preoperatively to 88.6 degrees. Failure was defined as an increase of less than 20 points in the modified Hospital for Special Surgery knee score at the time of the review or the need for an additional operation related to the allograft. Thirteen knee replacements failed, yielding a 75% success rate. Five knees had graft resorption, resulting in implant loosening. Four knee replacements failed because of infection, and two knees had nonunion between the host bone and the allograft. Two knees (one patient) did not have a 20-point improvement in the knee score. The survival rate of the allografts was 72% (95% confidence interval, 69% to 75%) at ten years. On radiographic analysis, none of the surviving grafts had severe resorption, one had moderate resorption, and two had mild resorption. One knee had a loose tibial component, and three knees had nonprogressive tibial radiolucent lines. All four knees were asymptomatic. CONCLUSIONS: Our results demonstrate that allografts used in revision knee replacement in patients with the difficult problem of massive bone loss have an encouraging medium-term rate of survival. PMID- 11263647 TI - The use of a Gore-Tex soft-tissue patch to repair large full-thickness defects after subtotal sternectomy. A report of three cases. PMID- 11263646 TI - The efficacy of low-pressure lavage with different irrigating solutions to remove adherent bacteria from bone. AB - BACKGROUND: Recent studies have suggested that high-pressure irrigation may have adverse effects on bone. However, the use of low-pressure irrigation may not remove all adherent bacteria from bone. The type of irrigating solution may be an important factor in the removal of adherent bacteria with pulsatile lavage. In this study, we compared the effects of various irrigating solutions on the number and function of osteoblasts and osteoclasts and we examined the effectiveness of these solutions in removing adherent bacteria from bone. METHODS: To examine the effect of irrigating solutions on the number and activity of osteoblasts, we isolated calvarial cells from newborn C57BI/6 mice and exposed the cells to equivalent concentrations of ethanol, povidoneiodine, liquid soap, antimicrobial wash (50 U/L of bacitracin), or chlorhexidine gluconate, for two, ten, or twenty minutes. The cells were then cultured in the presence of bone-nodule-enhancing medium (beta-glycerophosphate and ascorbic acid) for twenty-one days. The medium was changed every three or four days. Mineralized nodules were stained with alizarin red S, and osteoblasts were stained with a histochemical stain for alkaline phosphatase. Osteoclasts were identified with tartrate-resistant acid phosphatase staining. In a second experiment, canine cortical tibiae were contaminated with Staphylococcus aureus for six hours and subjected to different irrigating solutions with or without low-pressure lavage. Bacterial colony forming units were quantitated under each set of conditions. RESULTS: Each solution resulted in a time-dependent decrease in the number of calvarial osteoblasts and osteoclasts compared with that in the controls. The 1% soap solution resulted in greater preservation of both alkaline-phosphatase activity and bone-nodule formation than did the other solutions. Moreover, the soap solution preserved the number of osteoclasts to the greatest extent. The povidone iodine and chlorhexidine-gluconate solutions resulted in the largest decline in bone-nodule formation, alkaline-phosphatase activity, and number of osteoclasts. Low-pressure pulsatile lavage with the soap solution removed the most bacteria from the contaminated tibia when compared with either the soap solution alone or low-pressure irrigation with saline solution. CONCLUSIONS: Our findings suggest that certain solutions may be more effective in removing bacteria from bone than mechanical irrigation with saline solution alone. Among the various solutions examined, the soap solution preserved the number and activity of osteoblasts the most. Low-pressure lavage with the soap solution resulted in the greatest removal of adherent bacteria from bone. PMID- 11263648 TI - Extreme rotational malunion of the humerus. A case report. PMID- 11263649 TI - Metal sensitivity in patients with orthopaedic implants. AB - All metals in contact with biological systems undergo corrosion. This electrochemical process leads to the formation of metal ions, which may activate the immune system by forming complexes with endogenous proteins. Implant degradation products have been shown to be associated with dermatitis, urticaria, and vasculitis. If cutaneous signs of an allergic response appear after implantation of a metal device, metal sensitivity should be considered. Currently, there is no generally accepted test for the clinical determination of metal hypersensitivity to implanted devices. The prevalence of dermal sensitivity in patients with a joint replacement device, particularly those with a failed implant, is substantially higher than that in the general population. Until the roles of delayed hypersensitivity and humoral immune responses to metallic orthopaedic implants are more clearly defined, the risk to patients may be considered minimal. It is currently unclear whether metal sensitivity is a contributing factor to implant failure. PMID- 11263650 TI - Why orthopaedic surgeons leave full-time academic positions for private practice. PMID- 11263651 TI - Histological diagnosis and tibial neuromas. PMID- 11263652 TI - Diagnostic bone-density testing. PMID- 11263653 TI - Diagnostic coding and cohort data. PMID- 11263654 TI - Anterior instrumentation for the treatment of spinal tuberculosis. PMID- 11263655 TI - Orthopaedic POEMS and reliable research. PMID- 11263656 TI - Callus formation after bony resection in diabetic patients. PMID- 11263657 TI - Residency training. PMID- 11263658 TI - The Residency Review Committee for orthopaedic surgery. Establishing the standard for quality education. PMID- 11263659 TI - What's new in hand surgery. PMID- 11263660 TI - Bone morphogenetic proteins: from basic science to clinical applications. PMID- 11263661 TI - Smad-Runx interactions during chondrocyte maturation. AB - BACKGROUND: Intracellular signaling triggered by bone morphogenetic proteins (BMPs) results in activated Smad complexes that regulate transcription of BMP responsive genes. However, the low specificity of Smad binding to regulatory sequences implies that additional tissue-specific transcription factors are also needed. Runx2 (Cbfal) is a transcription factor required for bone formation. We have examined the role of Smads and Runx2 in BMP induction of type X collagen, which is a marker of chondrocyte hypertrophy leading to endochondral bone formation. METHODS: Pre-hypertrophic chondrocytes from the cephalic portion of the chick embryo sternum were placed in culture in the presence or absence of rhBMP-2. Cultures were transiently transfected with DNA containing the BMP responsive type X collagen promoter upstream of the luciferase gene. The cultures were also transfected with plasmids, causing over-expression of Smads or Runx2, or both. After 24-48 hours, cell extracts were examined for levels of luciferase expression. RESULTS: In the presence of BMP-2, chondrocytes over-expressing BMP activated Smadl or Smad5 showed significant enhancement of luciferase production compared with that seen with BMP alone. This enhancement was not observed with over-expression of Smad2, a transforming growth factor beta (TGF-beta)-activated Smad. Overexpression of Runx2 in BMP-treated cultures increased transcriptional activity to levels similar to those seen with Smads 1 or 5. When chondrocytes were simultaneously transfected with both Runx2 and Smad 1 or 5, promoter activity was further increased, indicating that BMP-stimulated Smad activity can be augmented by increasing the levels of Runx2. CONCLUSIONS: These results implicate the skeletal tissue transcription factor Runx2 in regulation of chondrocyte hypertrophy and suggest that maximal transcription of the type X collagen gene in pre-hypertrophic chondrocytes involves interaction of BMP stimulated Smads with Runx2. CLINICAL RELEVANCE: Many skeletal abnormalities are associated with impaired regulation of chondrocyte hypertrophy in growth plates. These studies demonstrate that both BMP-activated Smads and Runx2 levels can modulate chondrocyte transition to hypertrophy. PMID- 11263662 TI - Growth/differentiation factor-5 (GDF-5) and skeletal development. AB - BACKGROUND: Growth/differentiation factor-5 (GDF-5) has been shown to be essential for normal appendicular skeletal and joint development in humans and mice. In brachypod, a Gdf-5 gene mouse mutant, the defect is first apparent during early chondrogenesis, with the cartilage blastema already reduced in size by E12.5. This defect is associated with changes in the expression of cell surface molecules. METHODS: To understand further how GDF-5 controls cartilage formation, we first mapped the expression of the Gdf-5 gene during skeletal development (please note that the abbreviation for the gene is given in italics and the abbreviation for the protein expressed by the gene is given in capital letters). Subsequently, we over-expressed GDF-5 in the developing chick embryo using a replication competent retrovirus, RCAS(BP). We determined its effects on skeletal development by histological examination and its effects on early growth by autoradiography of proliferating cells. In addition, we examined the effect of GDF-5 on chondrogenic differentiation using micromass and single cell suspension cultures of limb mesenchymal cells, RESULTS: These studies show that the Gdf-5 gene is expressed in the early cartilage condensation, the perichondrium, and the joint interzone. Over-expresSion of GDF-5 in chick limb buds, during the condensation stage or later when the skeletal elements have formed, increased the size of the affected elements. In both cases, the increase in size was associated with an increase in cell number and, at later stages, this was correlated with an increase in S-phase cells. In vitro studies showed that GDF-5 could increase cell adhesiveness, and this may be a mechanism through which GDF-5 initiates condensation formation. CONCLUSION: These studies show that GDF-5 acts at two stages of skeletal development and by two distinct mechanisms. First, GDF-5 promotes the initial stages of chondrogenesis by promoting cell adhesion, which is consistent with the expression of Gdf-5 in the cartilage condensation. Second, GDF-5 can increase the size of the skeletal elements by increasing proliferation within the epiphyseal cartilage adjacent to its expression within the joint interzone. PMID- 11263663 TI - Transcriptional regulation by Smads: crosstalk between the TGF-beta and Wnt pathways. AB - BACKGROUND: Several studies have shown that cooperation between transforming growth factor beta (TGF-beta) and Wnt/wingless signaling pathways plays a role in controlling certain developmental events. These factors elicit their biological effects through distinct pathways in which TGF-beta and Wnt signaling induce activation of the transcriptional regulators Smads and lymphoid enhancer binding factor/T-cell-specific factor (LEF/TCF), respectively. To understand the mechanism for cooperativity between these pathways, we have investigated the molecular mechanism for this synergistic effect. METHODS: Transcriptional assays were conducted by transient transfection of HepG2 cells with use of luciferase reporter constructs. Protein/protein interaction studies were conducted in vitro with the use of glutathione-S-transferase pull-down assays and in intact cells by immunoprecipitation and immunoblotting. RESULTS: We show that Smads physically interact with LEF1/TCF transcription factors and that specific DNA binding sites in the Xenopus twin promoter are required for synergistic activation by TGF-beta and Wnt pathways. In addition, we demonstrate that TGF-beta-dependent activation of LEF1/TCF target genes can occur independently of beta-catenin, an essential component of the Wnt signaling pathway. CONCLUSIONS: TGF-beta and Wnt signaling pathways can independently or cooperatively regulate LEF1/TCF target genes. This suggests that the cooperation between these pathways may be important for the specification of cell fates during development. PMID- 11263664 TI - SIP1 (Smad interacting protein 1) and deltaEF1 (delta-crystallin enhancer binding factor) are structurally similar transcriptional repressors. AB - BACKGROUND: Smad proteins are intracellular mediators of transforming growth factor-beta (TGFbeta) signalling that regulate gene expression by interacting with different classes of transcription factors including DNA-binding multi-zinc finger proteins. One of these, Smad interacting protein 1 (SIP1), is a novel two handed zinc-finger protein that displays strong similarity with the transcriptional repressor delta-crystallin enhancer binding factor (deltaEF1). Here, we summarize what is known about the mechanism of action of both proteins and their role in vertebrate embryogenesis. Our data are discussed together with the present knowledge on other zinc-finger containing Smad interacting proteins. METHODS: The activities and function of SIP1 have been analysed through documentation of expression patterns, the effect of over-expression of SIP1 on target-gene expression, and promoter studies using Xenopus embryos. Moreover, S1P1/Smad complexes and their association with target promoter DNA were analyzed in biochemical studies. RESULTS: SIP1 is a transcriptional repressor displaying overlapping DNA binding specificities with deltaEF1. An in vivo target of SIP1 in Xenopus is a gene required for the formation of mesoderm, Brachyury (XBra). Our data indicate that SIP1 is required to confine XBra gene expression to the mesoderm. Furthermore, the expression pattern in Xenopus invites us to speculate that SIP1 plays a role in specification/differentiation of neuroectoderm. Unlike deltaEF1, SIP1 can bind directly to activated receptor regulated Smads (R-Smads) and recruit them to the DNA. This indicates that Smads may modulate the activity of SIP1 as a transcriptional repressor. CONCLUSIONS: Our data point to a role of SIP1 in developmental processes regulated by members of the TGFbeta family such as induction of mesoderm (mediated through activin-like signalling) and inhibition of neuroectoderm formation (mediated by bone morphogenetic proteins [BMPs]). Whereas SIP1 could act in TGFbeta signal transduction by virtue of interaction with activated R-Smads, genetic studies in the mouse indicate that deltaEF1 may act in certain TGFbeta pathways-i.e., BMPs and growth and differentiation factors (GDFs)-as well. The molecular mechanisms by which these transcriptional repressors act, as well as the function of the SIP1/Smad interaction, remain to be elucidated. CLINICAL RELEVANCE: Mutations in components of the TGFbeta signalling pathways have been associated with disease and congenital malformations. We anticipate that identification of Smad interacting transcription factors including SIP1 and their targets will help us to understand the molecular basis of certain pathologies. PMID- 11263665 TI - Intimate relationship between TGF-beta/BMP signaling and runt domain transcription factor, PEBP2/CBF. AB - BACKGROUND: When C2C12 pluripotent mesenchymal precursor cells are treated with transforming growth factor-beta1 (TGF-beta1), terminal differentiation into myotubes is blocked. Treatment with bone morphogenetic protein-2 (BMP-2) not only blocks myogenic differentiation but also induces osteoblastic differentiation. However, the molecular mechanisms governing the ability of TGF-beta and BMP to induce ligand-specific responses and inhibit myogenic differentiation are not known. The objective of our studies was to elucidate the molecular mechanisms that block myoblastic differentiation and induce osteoblastic differentiation in C2C12 cells. METHODS: Induction of RUNX2/PEBP2alphaA/Cbfa1 by TGF-beta and BMP was examined by electrophoretic mobility shift assay (EMSA) and Northern blot analysis. C2C12 cells stably expressing RUNX2 or Smad, or both, were established, and the role of these genes in the process of osteoblastic differentiation was analyzed by examining the expression of osteoblast-specific markers. RESULTS: Treatment of C2C12 with TGF-beta and BMP-induced RUNX2/PEBP2alphaA/Cbfa1, a global regulator of osteogenesis. Cooperation between RUNX2 and BMP-activated Smad induced osteoblastic differentiation. CONCLUSIONS: Both TGF-beta and BMP activate transcription of RUNX2, which is sufficient to inhibit myogenesis. To induce osteogenesis, BMP-induced RUNX2 must cooperate with BMP-activated Smads. PMID- 11263666 TI - BMP3: to be or not to be a BMP. AB - BACKGROUND: Bone morphogenetic proteins (BMPs) are osteogenic but also have diverse functions during development. BMP3 is a major component of osteogenin, which has osteogenic activity. However, recombinant BMP3 (rhBMP3) has no apparent osteogenic function, raising the possibility that BMP3 has no bone-inducing activity or that the recombinant material is not properly processed. To resolve this apparent discrepancy, we utilized a retroviral system to study the effects of BMP3 in vitro. In addition, we generated Bmp3-deficient mice to elucidate the function of BMP3 in vivo. METHODS: Retroviral as well as mammalian expression constructs were utilized to express BMP3 and to create BMP3 conditioned medium. Alkaline phosphatase (ALP) activity and transcriptional response assays were used to monitor the ability of BMP3 to elicit either a BMP-like or a transforming growth factor beta (TGF-beta)/activin-like response in osteoblastic cell lines. Finally, mice deficient in BMP3 were generated to investigate BMP3 function in vivo. RESULTS: BMP3 was unable to induce an osteogenic response in W-20-17, MC3T3 E1, or C3H10T1/2 cells, although all three cell lines were responsive to BMP2. However, BMP3 inhibited responsiveness to BMP2 in these assays, suggesting that BMP3 antagonizes BMP2 signaling. This inhibition did not occur through inhibition of binding of BMP2 to its receptors. BMP3 activated the TGF-beta/activin-pathway in these cells, suggesting that BMP3 exerts its inhibiting effects by activating a signaling pathway that antagonizes the BMP pathway. To examine the potential functional consequences of BMP3 action, Bmp3-/- mice, which lack BMP3, were generated. On an outbred genetic background, Bmp3-/- mice are viable and show no obvious skeletal phenotype as embryos or neonates. However, adult mice exhibit twice as much trabecular bone as do their wild-type littermates. This observation is consistent with our in vitro observations suggesting that BMP3 is an inhibitor of osteogenesis in vitro and of bone formation in vivo. CONCLUSIONS: BMP3 is an inhibitor of osteogenic BMPs and can signal through a TGF-beta/activin pathway. The ability of BMP3 to antagonize BMP2 activity may thus be a consequence of competition for signaling components common to TGF-beta/activin and BMP pathways. BMP3, the most abundant BMP in demineralized bone, may therefore play an essential role as a modulator of the activity of osteogenic BMPs in vivo. CLINICAL RELEVANCE: Therapies to accelerate bone healing usually utilize administration of exogenous BMP either in recombinant form or by gene therapy approaches. It is conceivable that the potency of osteogenic BMPs would be increased by inhibiting the activation of antagonistic signaling pathways or by increasing levels of rate-limiting signaling components shared by both BMP and TGF-beta/activin pathways. PMID- 11263667 TI - The effect of BMP on the expression of cytoskeletal proteins and its potential relevance. PMID- 11263668 TI - The crystal structure of the BMP-2:BMPR-IA complex and the generation of BMP-2 antagonists. AB - BACKGROUND: Bone morphogenetic proteins (BMPs) and growth and differentiation factors (GDFs) belong to the large transforming growth factor-beta (TGF-beta) superfamily of multifunctional cytokines. Signaling of the BMPs requires the binding of the BMP to the BMP cell surface receptors BMPR-IA, BMPR-IB, and BMPR II. Similar to other cytokines, members of the TGF-beta superfamily exhibit stringent specificity in their ligand-receptor interactions, which may be a reason for the qualitative and quantitative differences in cellular responses. To understand how BMPs and GDFs activate their receptors, it is important to determine structure and binding mechanisms of ligand-receptor complexes. We have used BMP-2 as a key representative of the BMPs to identify the epitopes for type I and type II receptor binding by mutational interaction analyses and have solved the crystal structure of a BMP2:BMPR-IA receptor ectodomain complex. METHODS: To identify amino acid side chains involved in receptor binding, a collection of in vitro mutagenized human BMP-2 variants was prepared and subjected to interaction analyses with use of the receptor ectodomains of BMPR-IA, BMPR-II, and ActR-II immobilized on a biosensor system. The biological activity of the BMP-2 variants was measured by BMP-2 dependent expression of alkaline phosphatase (ALP) in C2C12 cells. For crystallization, a complex of BMP-2 and the ectodomain of BMPR-IA was formed in solution, purified, and crystallized as described(12). RESULTS: The ligand-receptor interaction analysis of the BMP-2 variants identified distinct epitopes for type I and type II receptor binding. Because the structure of TGF beta-like proteins has been compared with that of an open hand, the binding epitope for the type I receptor was-on the basis of its location-termed "wrist" epitope. The crystal structure of the BMP-2:BMPR-IA ectodomain complex revealed a key feature of the ligand-receptor interaction: a large hydrophobic residue (Phe85) within a hydrophobic patch of BMPR-IA fit into a hydrophobic pocket composed of residues of both BMP-2 monomers. A second epitope identified by alanine mutagenesis scanning was termed the "knuckle" epitope on the basis of its location on the outer side of the "finger" segments of BMP-2. Mutations in either the wrist epitope or the knuckle epitope produced variants with altered biological activities. Variants with antagonistic properties were exclusively generated by mutations in the knuckle epitope of BMP-2. CONCLUSIONS AND CLINICAL RELEVANCE: The identification and characterization of the two receptor binding epitopes in BMP-2 provide new insight into the primary steps of BMP-receptor activation. Because of the structural similarities between members of the TGF beta superfamily, it can be assumed that the data presented in this work are transferable to other TGF-beta receptor systems. Because of the association with various diseases, the generation of antagonists of other TGF-beta superfamily members might generate potent tools for basic research and therapeutic approaches. PMID- 11263669 TI - Bone morphogenetic protein-7 modulates genes that maintain the vascular smooth muscle cell phenotype in culture. AB - BACKGROUND: The vasculature is an important component in the musculoskeletal system, and vascularization is a key event in the development of normal cartilage and bone formation. Blood vessels deliver nutrients, oxygen, and precursor cells to maintain the structural and functional integrity of joints and soft and hard tissues. Therefore, agents that help to inhibit proliferation and retain the phenotype of vascular smooth muscle cells (SMCs) are of critical importance. In this study, we examined the capacity of bone morphogenetic protein-7 (BMP-7) to inhibit the proliferation of SMCs and maintain their phenotype. METHODS: A thymidine-incorporation assay was used to monitor the proliferative activity of SMCs on stimulation with platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta), agents known to be stimulatory for these cells. Reverse transcriptase-polymerase chain reaction (RT-PCR), Northern blot analysis, and enzyme-linked immunosorbent assay (ELISA) were used to monitor the modulation of various genes and gene products. Immunolocalization of SMC specific markers was also performed. RESULTS: BMP-7 inhibited both serum-stimulated and growth factor-induced (PDGF-BB and TGF-beta1) SMC growth, as measured by 3H-thymidine uptake and cell number, in primary human aortic smooth muscle (HASM) cell cultures. The addition of BMP-7 stimulated the expression of developmentally regulated as well as SMC-specific markers, namely, Id-1 and Id-2, alpha-actin, and SMC-specific heavy-chain myosin, as examined by semiquantitative and quantitative RT-PCR and by Northern blot analysis. Additionally, BMP-7 exhibited anti-inflammatory activity by downregulating intercellular adhesion molecule-1 (ICAM-1) expression. The collagen type III/I ratio that becomes lower with the transdifferentiation of SMCs into myofibroblasts is maintained in BMP-7-treated cultures compared with untreated controls. Studies on the mechanism of action indicate that BMP-7 treatment induces cyclin-dependent kinase-2 inhibitor, p21, which was inhibited during PDGF-BB-induced proliferation of SMCs. Finally, BMP-7 upregulates the expression of the inhibitory Smads, Smad6 and Smad7, which are known to inhibit TGF-beta superfamily signaling. CONCLUSIONS: These results suggest that BMP-7 maintains the expression of the vascular SMC phenotype. Thus, BMP-7 may prevent vascular proliferative disorders and potentially could act as a palliative agent following damage to the vasculature. CLINICAL RELEVANCE: In musculoskeletal disorders in which the vasculature plays an important role, BMP-7 may be of benefit as an anti-inflammatory and anti-proliferative agent for vascular endothelium and help maintain vascular integrity. PMID- 11263671 TI - Alosetron and the rapid component of delayed rectifying potassium current in cardiac cells. AB - Some drugs acting on 5-hydroxytryptamine receptors inhibit the rapid component of delayed rectifying potassium currents (I(Kr)) in cardiac muscle cells. This is associated with lengthening of the QT interval in the cardiac cycle and can lead to fatal arrhythmias. We investigated whether alosetron, a novel 5HT3 antagonist proposed for treatment of irritable bowel syndrome (IBS), blocks I(Kr) in guinea pig cardiac myocytes. I(Kr) was isolated under whole-cell voltage clamp, and was identified by its sensitivity to the selective I(Kr) antagonist E4031. Cisapride (10(-6) M) inhibited the E4031-sensitive current while alosetron (10(-10)-10(-6) M) had no effect on I(Kr). We also found that alosetron did not inhibit I(Ks). Therefore, use of alosetron for treatment of IBS should not be confounded by long QT syndrome. PMID- 11263670 TI - Paradoxical effects of copper and manganese on brain mitochondrial function. AB - Defects in the mitochondrial genome have been associated with Parkinson's and Alzheimer's disease, and apoptosis can be triggered by the presence of energetically compromised mitochondria. Thus, in this study we have examined whether the divalent cations Cu2+ and Mn2+ could influence mitochondrial function in vitro. Mitochondrial electron transport was dose and time dependently reduced by Cu2+ to a greater extent with succinate as a substrate. Following a 60 min preincubation period, Mn2+ dose dependently inhibited electron transport to a greater extent with lactate and malate. In contrast, paradoxical effects were seen following a 5 min preincubation period with Mn2+. Cu2+ dose-dependently reduced NADH-dependent lactate dehydrogenase (LDH) activity, with almost complete inhibition apparent at 10 microM. An initial induction of LDH by 10 microM Mn2+ was partially reversed by higher concentrations of the metal. Cu2+ dose dependently reduced flavin adenine dinucleotide (FAD)-dependent monoamine oxidase A (MAO-A) activity in a time-independent manner, with an IC50 value approximately 20 microM, whereas Mn2+ had no effect. In conclusion, it is proposed that Cu2+ and Mn2+ have differential effects on nicotinamide adenine dinucleotide (NAD) and FAD-dependent mitochondrial enzymes at the level of the essential cofactors. Cu2+ appears to exert an inhibitory effect on both NAD and FAD-dependent enzymes, but predominantly against the latter, including MAO-A and succinate dehydrogenase. The complex responses to Mn2+ may be due to dose-related effects on the interconversion of NAD and NADH and reversible enzymatic reactions employing this nucleotide cofactor. PMID- 11263672 TI - Adhyperforin as a contributor to the effect of Hypericum perforatum L. in biochemical models of antidepressant activity. AB - The present paper describe investigations which demonstrate that hyperforin is not the only phloroglucinol derivative in extracts of the medicinal plant Hypericum perforatum L., which possess a biological activity. Hyperforin was the major lipophilic constituent in two different extracts, whereas the amount of adhyperforin was approximately 10 times lower. Adhyperforin, like hyperforin, is a potent inhibitor of the uptake of dopamine, serotonin and noradrenaline. Neither hyperforin nor adhyperforin inhibited binding of the cocaine analogue, [3H]WIN 35,428 to the dopamine transporter. However, the known antidepressives imipramine, nomifensine and fluoxetine all inhibited binding of [3H]WIN 35,428, indicating that hyperforin and adhyperforin do not bind to the same site on the dopamine transporter as these compounds. Furthermore, hyperforin and adhyperforin did not prevent dopamine binding, but inhibited dopamine translocation. Our studies further support recent reports suggesting that the effect of hyperforin on uptake of monoamines is probably not caused by a direct effect of hyperforin on known sites on the transporters. PMID- 11263673 TI - Ramadan fasting alters endocrine and neuroendocrine circadian patterns. Meal-time as a synchronizer in humans? AB - Muslims must refrain from eating, drinking, smoking, and sexual relations from sunrise to sunset during the month of Ramadan. Serum concentrations of melatonin, steroid hormones (cortisol, testosterone), pituitary hormones (prolactin, LH, FSH, GH, TSH) and thyroid hormones (free thyroxin and free triiodothyronine) were documented around the clock at six 4-hourly intervals before Ramadan began and on the twenty-third day of Ramadan (daytime fasting). Time series were analysed with repeated measures ANOVA. Statistically significant differences were found in some variables: the nocturnal peak of melatonin was diminished and may have been delayed; there was a shift in the onset of cortisol and testosterone secretion; the evening peak of prolactin was enhanced, FSH and GH rhythmic patterns were affected little or not at all by Ramadan fasting and only the serum TSH rhythm was blunted over the test time span. These data show that daytime fasting, modifications in sleep schedule and psychological and social habits during Ramadan induce changes in the rhythmic pattern of a number of hormonal variables. PMID- 11263674 TI - Decreased responsiveness of gluconeogenesis to the modulation by sulfonylureas in hepatocytes isolated from obese (fa/fa) Zucker rats. AB - The influence of the hypoglycemic agent glipizide (0-100 microM) on the rate of gluconeogenesis from lactate, as well as on the levels of fructose 2,6 bisphosphate, has been investigated in hepatocytes isolated from genetically obese (fa/fa) Zucker rats and from their corresponding lean (Fa/-) littermates. As compared to lean rat hepatocytes, liver cells isolated from obese animals showed a lower rate of basal gluconeogenesis (0.9 +/- 0.2 vs 5.4 +/- 0.5 micromol of lactate converted to glucose/g cell x 30 min, n=4) and higher levels of fructose 2,6-bisphosphate (11.5 +/- 1.0 vs 5.9 +/- 0.4 nmol/g cell, n=8-9). In lean rat hepatocytes, the presence of glipizide in the incubation medium caused a dose-dependent inhibition of the rate of lactate conversion to glucose (maximal inhibition=46%; EC50 value=26 microM), and simultaneously raised the cellular content of fructose-2,6-bisphosphate (maximal increment=40%; EC50 value=10 microM). In contrast, in hepatocytes isolated from obese rats, the inhibition of gluconeogenesis and the increment in fructose-2,6-bisphosphate levels elicited by glipizide were significantly reduced (maximal effects of 22 and 13%, respectively). Similarly, the activation of glycogen phosphorylase and the increase in hexose 6-phosphate levels in response to glipizide were less marked in obese rat hepatocytes than in liver cells isolated from lean animals. These results demonstrate that the efficacy of sulfonylureas as inhibitors of hepatic gluconeogenesis is reduced in the genetically obese (fa/fa) Zucker rat. PMID- 11263676 TI - The driving license examination as a stress model: effects on blood picture, serum cortisol and the production of interleukins in man. AB - We have studied the following stress model: the tension caused by sitting for the theoretical part of the driving license examination. Volunteers were investigated twice, after their driving license examination and after a (stress-free) control session. The effects of the stress were investigated by studying the blood picture (differential counts), serum concentration of cortisol, and cytokine production in stimulated blood cells. Relationships between the subjective perception of stress and the physiological reaction were also investigated. This stress induced significant increase in the concentrations of cortisol and hemoglobin, and in the values of hematocrit and MCV, and in the lipopolysaccharide-induced release of IL-1beta and -6. The subjective feelings of irritability and wakefulness were also significantly higher after the exam. A significant relationship was found between the changes in the stimulated production of IL-1beta and irritability. The responsiveness to psychological stress might be influenced by the temporary mood of the subjects. PMID- 11263675 TI - Pretreating rats with hyperoxia attenuates ischemia-reperfusion injury of the heart. AB - Oxidative stress may precondition the heart. The present study investigated whether hyperoxia elicits a preconditioning-like response. Rats were kept in a hyperoxic (>95% O2) environment for 60 or 180 minutes. Hearts were Langendorff perfused immediately or 24 hours after hyperoxia, and exposed to 25 minutes of global ischemia and 60 minutes of reperfusion. Whole blood was sampled after 60 and 180 minutes of hyperoxia for oxidative stress markers. Hearts were sampled immediately or 24 hours after hyperoxia for measurement of antioxidants, lipid peroxidation products, heat shock protein 72 and endothelial nitric oxide synthase. At the end of reperfusion after 1 h hyperoxia, infarct size was determined by tetrazolium staining. Hyperoxia increased serum levels of conjugated dienes, reduced serum antioxidative protection, reduced reperfusion arrhythmias in most groups, and improved myocardial function. Infarct size was reduced from 45% of myocardial tissue in controls to 22% in treated animals. The myocardial activity of antioxidant enzymes, content of heat shock protein 72, and endothelial nitric oxide synthase in myocardial tissue were not influenced. In conclusion, hyperoxia induces a low-graded systemic oxidative stress, improves postischemic cardiac function and reduces infarct size. The mediators of protection remain to be determined. PMID- 11263677 TI - Ligand binding profiles of delta-opioid receptor in human cerebral cortex membranes: evidence of delta-opioid receptor heterogeneity. AB - In this study, receptor binding profiles of opioid ligands for subtypes of opioid delta-receptors were examined employing [3H]D-Pen2,D-Pen5-enkephalin ([3H]DPDPE) and [3H]Ile(5,6)-deltorphin II ([3H]Ile-Delt II) in human cerebral cortex membranes. [3H]DPDPE, a representative ligand for delta1 sites, labeled a single population of binding sites with apparent affinity constant (Kd) of 2.72 +/- 0.21 nM and maximal binding capacity (Bmax) value of 20.78 +/- 3.13 fmol/mg protein. Homologous competition curve of [3H]Ile-Delt II, a representative ligand for delta2 sites, was best fit by the one-site model (Kd = 0.82 +/- 0.07 nM). Bmax value (43.65 +/- 2.41 fmol/mg) for [3H]Ile-Delt II was significantly greater than that for [3H]DPDPE. DPDPE, [D-Ala2,D-Leu5]enkephalin (DADLE) and 7 benzylidenaltrexone (BNTX) were more potent in competing for the binding sites of [3H]DPDPE than for those of [3H]Ile-Delt II. On the other hand, deltorphin II (Delt II), [D-Ser2,Leu5,Thr6]enkephalin (DSLET), naltriben (NTB) and naltrindole (NTI) were found to be equipotent in competing for [3H]DPDPE and [3H]Ile-Delt II binding sites. These results indicate that both subtypes of opioid delta receptors, delta1 and delta2, exist in human cerebral cortex with different ligand binding profiles. PMID- 11263678 TI - Increased ACTH and cortisol secretion after interleukin-alpha injection in the common marmoset (Callithrix jacchus jacchus). AB - We have studied the effect of intravenous injection of interleukin-1 (dose range: from 0.25 to 4.5 microg/kg of body weight) on plasma ACTH and cortisol levels in the marmoset, a primate paradygm of peripheral glucocorticoid resistance. Blood sampling were collected and body temperature recorded 0, 15, 30, 60, 120, 180, 240 and 300 min after injection. Interleukin-1 stimulated secretion of ACTH in a dose-dependent fashion. Maximal secretion occurred 120 min after injection, and lasted up to 240 min. Plasma ACTH levels returned to baseline 300 min after interleukin-1 injection. Plasma cortisol levels were related to ACTH levels. Body temperature elevation, which occurred 10-15 min after injection was dose dependent, and lasted 3 h. Results suggest that the pyrogenic effect of interleukin is associated, in the marmoset, with integrated activation of the hypothalamic-pituitary-adrenal axis. In light of the proneness of marmosets to hyperimmune disorders, our data are consistent with the hypothesized central biological role of IL-1, as well as the pathophysiological relevance of the neuro endocrine-immune cross-talk during the acute phase response. PMID- 11263679 TI - A reduction of tumor necrosis factor-alpha in paw exudate of lipopolysaccharide treated rats by nimesulide. AB - TNF-alpha is considered to play a pivotal role in many inflammatory and illness states. In our previous study we found that nimesulide, a COX-2 selective inhibitor, prevents the lipopolysaccharide-induced elevation in plasma TNF-alpha in rats. The present study was undertaken to elucidate the effect of nimesulide on TNF-alpha levels in the paw exudate of rats pretreated with lipopolysaccharide (LPS). Rats were injected (subplantar) with LPS (100 microg/paw) in the right hind, which resulted in a prominent increase in paw exudate TNF-alpha levels, peaked at one hour post injection. In rats pretreated with nimesulide (30 mg/kg, i.p.), the elevation in TNF-alpha in the paw exudate was significantly reduced, as compared to the LPS treated group. These results further stress the multiple antiinflammatory effects of nimesulide. PMID- 11263680 TI - Metabolic activation of mitomycin C by NADPH-ferredoxin reductase in vitro. AB - Mammalian NADPH-ferredoxin reductase (EC 1.18.1.2) functions in the mitochondrial electron transport chain for cytochrome P-450-dependent steroid hydroxylation. Significant homology of three-dimensional structure exists in the surroundings of FAD between NADPH-ferredoxin reductase and NADH-cytochrome b5 reductase. The latter is involved in the bioreduction of mitomycin C (MC), a prototype antitumor agent. In this study, we assessed the capacity of NADPH-ferredoxin reductase to activate MC. Mitomycin C increased the NADPH oxidase activity of NADPH-ferredoxin reductase. In the absence of ferredoxin, the Km value of NADPH-ferredoxin reductase for MC was 73.5 +/- 2.3 microM. While in the presence of 500 nM ferredoxin, a Lineweaver-Burk plot exhibited a biphasic curve. NADPH-ferredoxin reductase-mediated reduction of MC resulted in the formation of an alkylated complex of 4-(p-nitrobenzyl) pyridine and an increase in plasmide DNA single strand breaks under hypoxic conditions. With the addition of 500 nM ferredoxin, the amount of the alkylated complex of 4-(p-nitrobenzyl) pyridine and the plasmide DNA single-strand breaks increased by 40% and 37%, respectively. However, neither alkylated complex of 4-(p-nitrobenzyl) pyridine nor DNA strand breaks was observed in the presence of SOD and catalase under aerobic conditions. These findings demonstrate that NADPH-ferredoxin reductase is capable of catalyzing the bioactivation of mitomycin C under hypoxic conditions in vitro. PMID- 11263681 TI - A comparison of HIV-1 integrase inhibition by aqueous and methanol extracts of Chinese medicinal herbs. AB - The aqueous and methanol extracts of twenty herbs traditionally used in Chinese medicine were screened for anti-HIV-1 integrase activity in a non-radioactive ELISA-based HIV-1 integrase assay. The screening was performed at an herb extract concentration of 50 microg/ml. It was found that most of the aqueous and methanol herb extracts could elicit strong inhibition of HIV-I integrase activity. The inhibition was most likely due to tannins or polyphenolics in the herb extracts. In most of the herb extracts, 40-80% of the anti-HIV-1 integrase activity could be removed after passing through a minicolumn of polyamide resin. After removal of polyphenolic compounds, the methanol extract of Paeonia suffruticosa still exerted potent inhibition of HIV-1 integrase (EC50 = 15 microg/ml) and the aqueous extract of Prunella vulgaris caused moderate inhibition (EC50 = 45 microg/ml). The results support the view that herbs represent a rich source of anti-HIV compounds. PMID- 11263682 TI - Athletics for people with disabilities. PMID- 11263683 TI - Brain dysmorphology in individuals with severe prenatal alcohol exposure. AB - Our previous studies revealed abnormalities on structural MRI (sMRI) in small groups of children exposed to alcohol prenatally. Microcephaly, disproportionately reduced basal ganglia volume, and abnormalities of the cerebellar vermis and corpus callosum were demonstrated. The present study used sMRI to examine in detail the regional pattern of brain hypoplasia resulting from prenatal exposure to alcohol using a higher resolution imaging protocol and larger sample sizes than reported previously. Fourteen participants (mean 11.4 years; eight females, six males) with fetal alcohol syndrome (FAS) and 12 participants (mean 14.8 years; four females, eight males) with prenatal exposure to alcohol (PEA) but without the facial features of FAS were compared to a group of 41 control participants (mean 12.8 years, 20 females, 21 males). Findings of significant microcephaly and disproportionately reduced basal ganglia volumes in the FAS group were confirmed. Novel findings were that in FAS participants, white matter volumes were more affected than gray matter volumes in the cerebrum, and parietal lobes were more affected than temporal and occipital lobes. Among subcortical structures, in contrast to the disproportionate effects on caudate nucleus, the hippocampus was relatively preserved in FAS participants. Differences between the PEA group and controls were generally non-significant; however, among a few of the structures most affected in FAS participants, there was some evidence for volume reduction in PEA participants as well, specifically in basal ganglia and the parietal lobe. There were no group differences in cerebral volume asymmetries. Severe prenatal alcohol exposure appears to produce a specific pattern of brain hypoplasia. PMID- 11263684 TI - Corpus callosum morphology of Williams syndrome: relation to genetics and behavior. AB - As the largest interhemispheric commissure in the brain, the corpus callosum is of particular interest in disorders that may preferentially affect white matter development such as Williams syndrome (WS). Individuals with WS possess a remarkable array of neurobehavioral peaks and valleys, including deficits in visuospatial ability, mathematics, and attention, but with relative preservation of language and affect. Our study measured the corpus callosum and its primary subdivisions using high-resolution MRI in 20 individuals with WS (13 females and seven males; mean age 28.5, SD 8.3 years; range 19 to 44 years) and 20 age- and sex-matched control participants (mean age 28.5, SD 8.2 years; range 19 to 48 years). Total midsagittal corpus callosum area was reduced (F=4.5, p=0.04, df=36) in the WS population. The area of the splenium (F=12.4, p=0.001, df=36) and isthmus (F=9.4, p=0.004, df=36) were disproportionately reduced in WS beyond the absolute reduction of the entire corpus callosum. These reductions are in concordance with other neuroanatomical findings of decreased parietooccipital volumes as well as the observed visuospatial problems associated with WS. PMID- 11263685 TI - The 2nd to 4th digit ratio and autism. AB - It has been suggested that autism may arise as the result of exposure to high concentrations of prenatal testosterone. There is evidence that the ratio of the lengths of the 2nd and 4th digit (2D:4D) may be negatively correlated with prenatal testosterone. We measured 2D:4D in 95 families recruited via the National Autistic Society, UK. The sample comprised a total 72 children with autism (62 males, 10 females; age range 2 to 14 years), including 23 children (20 males, three females) with Asperger syndrome (AS), 34 siblings, 88 fathers, 88 mothers and sex- and age-matched control participants. We found that the 2D:4D ratios of children with autism, their siblings, fathers and mothers were lower than population normative values. Children with AS, who share the social and communicative symptoms of autism but have normal or even high IQ, had higher 2D:4D ratios than children with autism but lower ratios than population normative values. There were positive associations between 2D:4D ratios of children with autism and the ratios of their relatives. Children with autism had lower than expected 2D:4D ratios and children with AS higher ratios than expected in relation to their fathers' 2D:4D ratio. It was concluded that 2D:4D ratio may be a possible marker for autism which could implicate prenatal testosterone in its aetiology. PMID- 11263686 TI - Neurocognitive stability in Asperger syndrome, ADHD, and reading and writing disorder: a pilot study. AB - Boys with Asperger syndrome (n=20), attention-deficit-hyperactivity disorder (n=20), and reading and writing disorder (n=20) were followed up and retested on several neuropsychological measures 1 to 2 years after initial assessments. Wechsler Intelligence Scale for Children (WISC-III) Full Scale, Verbal, and Performance IQ scores remained stable for all diagnostic groups. Kaufman factors and 'fluid' and 'crystallized' abilities were also stable measures. Subtest stability over time, was slightly more variable. There was a tendency for the group with Asperger syndrome to deteriorate over time with respect to logical reasoning abilities. Measures of executive function/attention ('go-no-go' and 'conflict' tests) showed good test-retest stability in all diagnostic groups. This is the first study of its kind. PMID- 11263687 TI - Attentional and neuromotor deficits in ADHD. AB - In order to classify attention-deficit-hyperactivity disorder (ADHD) in 11-year old children, the role of specific attentional and motor deficits was examined. Participants comprised 22 children with ADHD (19 male, 3 female; median age 11 years, range 8.8 to 13.5 years) and 20 control children (17 male, 3 female; median age 10.6 years, range 8.2 to 12.6 years). Neuromotor assessment indicated that while both groups needed more time to complete finger compared to hand movements, this increase was more pronounced in children with ADHD. Reaction-time testing with continuous-force recording identified both motor and attentional deficits in children with ADHD. Longer intervals between force onset and force peak, and higher rate of responses with multiple force peaks (particularly in the bilateral condition) revealed specific deficits in the speed and quality of their motor output. Increase in errors and variability of force onsets indicated attentional deficits. Prediction analysis indicated that force-onset variability contributed significantly to group classification which was 85.7% correct. Neither neuromotor assessment nor specific motor deficits contributed significantly to classification, indicating that pure motor-speed measures play a minor role in characterizing ADHD in this age range. PMID- 11263688 TI - Test-retest reliability of balance tests in children with cerebral palsy. AB - To investigate intrasession and intersession reliability of balance tests in children with or without disabilities, 50 children without disabilities (ND) and 36 children with cerebral palsy (CP) aged from 5 to 12 years were tested. Intrasession reliability of postural stability of the Smart Balance Master System and one-leg standing test were assessed in both groups and intersession reliability of the Smart Balance Master System and balance subtest of the Bruininks-Oseretsky Test of Motor Proficiency (BOTMP) were assessed in ND children. Intersession reliability of the postural stability test in ND children, obtained using the Smart Balance Master System, was of moderate to good reliability in centre target (CT), sway vision (SV), eyes open and sway surface (EOSS), and sway vision and sway surface (SVSS; ICC 0.72 to 0.84). In children with CP, intrasession reliability was high in CT (ICC 1). One-leg standing tests in both groups also had moderate to good intersession reliability (ICC 0.56 to 0.99). Agreement of failure score of lateral rhythmic shifting (LRS) at 1 second and 2 seconds pace was 85% and 93% respectively in ND children. Within the balance subtest of BOTMP, only two items had 100% agreement. Results suggest that postural stability tests in four conditions (CT, SV, EOSS, and SVSS), LRS, one leg standing, and walking on a line are reliable and can be used to monitor balance control in ND children. Postural stability in CT condition and one-leg standing test are reliable in children with CP. Further study is needed to establish more reliable balance tests for children with CP. PMID- 11263689 TI - Relation between objectively measured feeding competence and nutrition in children with cerebral palsy. AB - This study aimed to investigate the prevalence of undernutrition in children with cerebral palsy (CP) and to determine the relation with feeding ability. Ninety children with CP from special needs schools were examined. Undernutrition was diagnosed on one or more of the following criteria: weight <2nd centile, triceps or subscapular skinfold measurement <3rd centile, mid-arm circumference <5th centile. Feeding competence was scored with respect to seven specific oromotor tasks using the Multidisciplinary Feeding Profile. Thirty-six participants (40%) had diplegia, 29 (33%) quadriplegia, 13 (14%) hemiplegia, and 12 participants (13%) had dyskinetic CP. Age ranged from 2.6 to 18.7 years (mean 10.8 years). Forty-six percent (41 of 90) were undernourished. In all aspects of feeding, those undernourished had lower feeding competence scores compared to adequately nourished children (p<0.002). Each modality of feeding competence correlated significantly to the centiles of weight, triceps or subscapular skinfold measurement and mid-arm circumference (p<0.02). A positive association of weight, triceps skinfold measurement, and mid-arm circumference with chewing ability was present independent of other feeding modalities (p<0.05). Undernutrition was common in this group and was associated with poorer feeding ability. PMID- 11263690 TI - Auditory function at 14 years of age of very-low-birthweight. AB - The aim of the study was to determine audiological function at 14 years of age of very-low-birthweight (VLBW < or = 1500 g) children compared with a cohort of normal birthweight (NBW > 2499 g) children. Participants were consecutive surviving preterm children of birthweight < 1000 g born between 1977 and 1982 (n=86) and of birthweight 1000 to 1500 g born between 1980 and 1982 (n=124) and randomly selected NBW children born between 1981 and 1982 (n=60). Audiometric tests included pure tone audiometry, tympanometry, stapedius muscle reflexes, and measures of central auditory processing. Psychometric tests included measures of IQ, academic achievement, and behaviour. There were no significant differences in rates of hearing impairment, abnormal tympanograms, figure-ground problems, or digit recall between VLBW children and NBW control children. VLBW children had higher rates of some central auditory processing problems, which in turn were associated with poorer intellectual, academic, and behavioural progress. PMID- 11263691 TI - Role of vision on early motor development: lessons from the blind. AB - For a better understanding of the contribution vision makes to the development of other sensory systems and to movement and posture, we studied effects of early blindness by examining video recordings of 14 totally blind infants. Infants were born at term or preterm and showed no evidence of brain damage. During preterm and term periods no noticeable changes in motor activity were observed. Around 2 months postterm all infants showed clear delay in head control and abnormal, exaggerated type of 'fidgety movements'. Later, postural control was characterized by a prolonged period of ataxic features. Results indicate a lack of normal calibration exerted by vision on proprioceptive and vestibular systems. Early visuomotor coordination such as coordinated eye-head scanning and head orientating were present but disappeared after several weeks. PMID- 11263693 TI - Physical fitness levels of persons with cerebral palsy. PMID- 11263692 TI - Fetal valproate syndrome and autism: additional evidence of an association. AB - Autism has been described in association with a variety of medical and genetic conditions. We previously reported on a patient whose clinical phenotype was compatible with both fetal valproate syndrome (FVS) and autism. Here we present five additional patients with FVS and autism. In all five of our patients, there was evidence of cognitive deficits, manifestations of autism, and typical phenotypic characteristics of FVS. The association between this known teratogen and autism has both clinical and research implications. PMID- 11263694 TI - Development of parainfectious opsoclonus in an infant by a non-humoral immune mechanism. PMID- 11263695 TI - When chronic disability meets acute stress: psychological and functional changes. PMID- 11263696 TI - Consolidating international relations. PMID- 11263697 TI - What is your diagnosis? Tracheal collapse. PMID- 11263698 TI - Measurement of the back fat layer in beagles for estimation of obesity using two dimensional ultrasonography. AB - The purpose of this study was to select a suitable location for measuring the subcutaneous fat layer in beagles as an indicator of excess body fat deposition. A location suitable for such a measurement should meet five conditions--it should be easy to: (1) apply the probe, (2) detect the anatomical character of the measurement location, and (3) obtain a reproducible ultrasonogram at all times. In addition, the fat layer should be (4) thick enough for measuring, and (5) located in an area that varies markedly with the degree of obesity. Ultrasonograms made in the transverse planes on the top of the spinous process of the sixth lumbar vertebra, the seventh lumbar vertebra and the first sacral vertebra were found to be suitable for measurement. The depth and area of the back fat layer at these locations were closely related to the degree of obesity. PMID- 11263699 TI - Bone allografts and adjuvant cisplatin for the treatment of canine appendicular osteosarcoma in 18 dogs. AB - The results achieved in 18 dogs following the use of frozen bone cortical allografts for limb-sparing resection of non-metastatic canine appendicular osteosarcoma are presented. Three to five cisplatin doses (70 mg/m2) were administered, starting the day after surgery. The mean and median survival times were 478 and 266 days (range 80 to 2,611 days), respectively. The survival rate was 94 per cent at three months, 78 per cent at six months, 35 per cent at 12 months, 23 per cent at 18 months and 19 per cent at 24 months; the disease-free interval was 80 to 1,246 days (mean 365 days, median 266 days). Lung metastasis developed in 55 per cent of the dogs within one year. Complications were observed in 14/18 dogs (78 per cent), comprising local recurrence (28 per cent), allograft infection (39 per cent) and implant failure (11 per cent). Despite complications, limb sparing is a useful alternative to amputation in selected cases of appendicular osteosarcoma. PMID- 11263700 TI - Effect of long-term intermittent periodontal care on canine periodontal disease. AB - The periodontal health status was assessed in two groups of dogs which had received different levels of periodontal care over a two-year period. The dental group received regular dental scaling and polishing, and intermittent daily tooth brushing, while the control group received no periodontal care. All dogs developed gingivitis, and two (one from each group) showed evidence of incipient periodontitis. The dental group had a reduced gingivitis index (GI) compared with the control group only when they had received daily tooth brushing before the GI assessment. When the dogs had not had their teeth brushed for four weeks before the assessment, the GI was not significantly different to that in the control dogs. This suggests that continual periodontal care throughout life is of great importance and questions the benefits of intermittent oral care. The GI of the palatal and lingual surfaces in all dogs was significantly higher than the GI of the buccal surfaces. Thus, all tooth surfaces may need to be cleaned to achieve optimal periodontal health. PMID- 11263701 TI - Effect of propofol at two injection rates or thiopentone on post-intubation apnoea in the dog. AB - Ventilatory effects at induction of anaesthesia were studied following intubation in 66 dogs anaesthetised using thiopentone (10 mg/kg) or propofol (4 mg/kg, injected rapidly or 4 mg/kg, injected slowly). Acepromazine and morphine preanaesthetic medication was administered, and anaesthesia was maintained with halothane in nitrous oxide and oxygen. The time from connection of the breathing system to the first breath was measured. Apnoea was defined as cessation of spontaneous respiration for 15 seconds or longer. Respiratory rate and minute volume were measured for the first five minutes of anaesthesia. Propofol was associated with a greater incidence of apnoea than thiopentone (59 per cent and 64 per cent compared with 32 per cent), but this difference was not statistically significant. Time to first breath was significantly longer with propofol than thiopentone and longest with the slower injection of propofol (P<0.05) (median of four seconds for thiopentone, 19.5 seconds for the propofol rapid injection, and 28.8 seconds for the propofol slow injection). In conclusion, the induction agent and speed of injection affect the incidence and duration of post-intubation apnoea. PMID- 11263702 TI - Cutaneous alternariosis in a cat. AB - A 10-year-old male domestic shorthaired cat had a chronic, slowly enlarging subcutaneous mass on the right side of its nose. At the time of presentation, the nasal airflow was severely impeded on the affected side. The cat had been treated medically with various drugs. Oral itraconazole had been the most effective in reducing the size of the mass, but had caused hepatotoxicity and had to be withdrawn. The mass was finally removed surgically, and a diagnosis of granulomatous cellulitis caused by Alternaria alternata (phaeohyphomycosis) was established, based on histopathology and fungal isolation. There has been no recurrence of the lesion after 21 months and the cat remains clinically well at the time of writing. Subcutaneous phaeohyphomycosis caused by A alternata has not, to the authors' knowledge, been previously described in small domestic animals in the UK. PMID- 11263703 TI - Atlantoaxial cartilaginous exostosis causing spinal cord compression in a mature Bernese mountain dog. AB - Cartilaginous exostosis developed in the atlantoaxial region of a three-and-a half-year-old Bernese mountain dog. The dog exhibited ataxia in the hindlimbs and flailing movements in the forelimbs. On survey radiographs of the cervical spine there was a focal calcified mass between the dorsal arch of the atlas and the spinous process of the axis. Lumbar myelography revealed severe dorsal spinal cord compression. The mass was removed surgically and the dog made a complete recovery. Histopathology of the excised mass was consistent with a diagnosis of cartilaginous exostosis. PMID- 11263705 TI - Atresia of the distal external acoustic meatus in a Bouvier des Flandres. AB - A five-year-old male Bouvier des Flandres was presented with chronic otalgia of approximately four and a half years' duration. Atresia of the external acoustic meatus was diagnosed. A total ear canal ablation and bulla osteotomy were performed. All symptoms of otalgia resolved within two weeks of surgery. PMID- 11263704 TI - Canine symmetrical lupoid onychodystrophy: a retrospective study with particular reference to management. AB - The records of six dogs in which a diagnosis of symmetrical lupoid onychodystrophy (SLO) had been made were examined retrospectively. The age at onset ranged from six months to eight years. All the dogs had been presented with onycholysis, onychomadesis, onychalgia and onychodystrophy. The diagnosis of SLO was confirmed in all cases by histological examination. Histological features were similar in all cases and included hydropic degeneration of the basal cell layer, pigmentary incontinence and a cell-rich interface dermatitis. Response (defined as good, partial or failure) to various therapies was compared. Treatments (as initial therapy or following previous treatment failure) included essential fatty acids (EFA) (three cases), a combination of tetracycline and nicotinamide (four cases) and azathioprine and/or prednisolone (one case each). EFA therapy resulted in one good response, one partial and one failure, tetracycline and nicotinamide in two good responses, one partial and one failure, and azathioprine and/or prednisolone in one good and one partial response. Although all treatments were successful in some cases, none was universally effective. PMID- 11263706 TI - Checklist of infections that may be imported into the UK by the travelling pet. PMID- 11263708 TI - Assessing pain in older adults. PMID- 11263707 TI - Solving the neurological dilemma. PMID- 11263709 TI - Is the clinical expression of late-life depression influenced by brain changes? MRI subcortical neuroanatomical correlates of depressive symptoms. AB - BACKGROUND: "Vascular depression" has recently been proposed. It is characterized by magnetic resonance imaging (MRI) T2-weighted subcortical lesions, a late onset of first episode of depression, and reduced heritability; a cerebrovascular etiology is suggested. The validity of "vascular depression" might be strengthened if an association was found between the subcortical lesions used to define it and particular depressive symptoms. METHODS: A blinded cross-sectional examination of DSM-III-R depressive symptoms (American Psychiatric Association, 1987) and MRI T2-weighted subcortical lesions in 44 patients with late-life depression. RESULTS: Many associations were found; however, because of multiple comparisons, their significance is viewed with caution. The most robust finding was that psychomotor retardation was independently related to total white-matter score. The odds of showing psychomotor retardation was increased 1.9 times for every point increase in severity of white-matter change. CONCLUSION: In late-life depression the clinical expression of the depression is influenced by the pattern of MRI T2-weighted subcortical lesions. This gives some validity to the concept of an MRI-defined "vascular" subtype of late-life depression and strengthens the argument for including neuroimaging in the classification of late-life depression. PMID- 11263710 TI - Development of a shorter version of the geriatric depression scale for visually impaired older patients. AB - OBJECTIVE: To validate a shorter version of the Geriatric Depression Scale (GDS) for older, visually impaired patients. PARTICIPANTS: Subjects were 70 visually impaired adults over age 65 who were presenting for services at a low vision clinic. METHOD: Subjects were interviewed by a geriatric nurse practitioner. A structured clinical interview was used to ascertain major depression, and the 15 item GDS was used to assess depressive symptoms. A multiple logistic regression was performed in which the dependent variable was clinical diagnosis of major depression and the independent variables were the 15 GDS items. Four items were significant, and were used to form the GDS-Abbreviated (GDS-A) scale. Sensitivity and specificity analyses were performed on various combinations of these four items to generate an effective cutoff score. RESULTS: Endorsing any two or more of the following four items--a) dissatisfied with life, (b) feeling helpless, (c) reporting problems with memory, and (d) lost activities and interests-yielded the best results with a sensitivity of .71 and a specificity of .88. This GDS-A cutoff score better differentiates depressed from nondepressed individuals than the cutoff score of 5 that is recommended for the GDS-15. CONCLUSION: The GDS-A's short format and strong discriminating ability make it an effective, convenient tool for screening visually impaired, older patients for depression. PMID- 11263711 TI - Advanced parental age: a risk factor for Alzheimer's disease or depression in the elderly? AB - BACKGROUND: Advanced parental age has been suggested as a risk factor for Alzheimer's disease (AD) as well as for other psychiatric disorders. In the present investigation, a sample of gerontopsychiatric patients was examined for a possible parental age effect. STUDY POPULATION AND METHODS: Eighty-three patients with AD, 154 elderly patients with depressive episodes, and 48 comorbid patients (AD and depressive episode) as well as 107 age-matched healthy control subjects from the general population were included in the investigation. Information on the years of birth of the parents was derived from personal or family history information. RESULTS: The mean maternal and paternal ages at the time of birth of the index subject were not significantly different for the different diagnostic subgroups or for the control sample. CONCLUSION: There was no evidence in our sample that advanced parental age increases the risk of AD or depression in the elderly. PMID- 11263712 TI - Feasibility and effectiveness of treatments for depression in elderly medical inpatients: a systematic review. AB - To determine the feasibility and effectiveness of treatments for depressed elderly medical inpatients, MEDLINE was searched for potentially relevant articles published from January 1987 to August 1997, using the keywords "depression or depressive disorder" (exploded) and "aged." The bibliographies of relevant articles were searched for additional references. Fifteen reports met the following inclusion criteria: (a) published in English or French; (b) minimum age criterion of 55 and over or mean age 65 and over; (c) subjects admitted to the medical service of an acute care hospital; (d) used accepted criteria for depression; (e) examined treatment(s) for depression; and (f) reported outcomes as a depression diagnosis and/or symptom level. Information was abstracted independently from each article by two reviewers, tabulated, and compared. The limited evidence suggests contraindications to treatment in 38% to 87% of subjects who received a heterocyclic antidepressant compared to 4% of subjects who received the selective serotonin reuptake inhibitor (SSRI) fluoxetine; rates of discontinuation and study completion were similar for heterocyclics, the SSRIs, and psychostimulants. All of the treatments (including social support/psychotherapy) appeared to be at least modestly effective in the short term. PMID- 11263713 TI - Disruptive vocalization and depression in older nursing home residents. AB - Screaming and other types of disruptive vocalization are commonly observed among nursing home residents. Depressive symptoms are also frequently seen in this group, although the relationship between disruptive vocalization and depressive symptoms is unclear. Accordingly, we sought to examine this relationship in older nursing home residents. We undertook a controlled comparison of 41 vocally disruptive nursing home residents and 43 nonvocally-disruptive nursing home residents. All participants were selected to have Mini-Mental State Examination (MMSE) scores of at least 10. Participants had a mean age of 81.0 years (range 63 97 years) and had a mean MMSE score of 17.8 (range 10-29). Nurse ratings of disruptive vocalization according to a semioperationalized definition were validated against the noisy behavior subscale of the Cohen-Mansfield Agitation Inventory. Subjects were independently rated for depressive symptoms by a psychiatrist using the Dementia Mood Assessment Scale, the Cornell Scale for Depression in Dementia, and the Depressive Signs Scale. Vocally disruptive nursing home residents scored significantly higher than controls on each of these three depression-in-dementia scales. These differences remained significant when the effects of possible confounding variables of cognitive impairment, age, and sex were removed. We conclude that depressive symptoms are associated with disruptive vocalization and may have an etiological role in the generation of disruptive vocalization behaviors in elderly nursing home residents. PMID- 11263714 TI - Delirium in elderly people without severe predisposing disorders: etiology and 1 year prognosis after discharge. AB - BACKGROUND: The etiologic factors of delirium have been frequently studied in hospitalized elderly patients who usually have an underlying disorder, i.e., hip fracture or dementia predisposing to delirium. The etiologic factors of delirium and prognosis in healthy elderly remain unstudied. The aim of our study was to detect the primary and additional etiologic factors contributing to delirium among community-dwelling healthy elderly people without predisposing disorders to delirium and to evaluate 1-year prognosis after discharge to home. METHOD: The study subjects consisted of 51 community-dwelling people over 65 years of age, without severe underlying disorders predisposing to delirium, admitted consecutively to the hospital because of a delirious state. The diagnosis of delirium was based on the DSM-III-R criteria. After discharge to home, the subjects were followed up for 1 year. RESULTS: The most important primary causes of delirium were infections in 22 cases (43%) and cerebrovascular attacks in 13 cases (25%). After the 1-year follow-up period, 10 patients (20%) had been taken into long-term care and 5 patients (10%) had died. DISCUSSION: The plausible etiologic factor of delirium was detected in all cases. Among healthy elderly people, infections and cerebrovascular attacks were the most important etiologic factors for delirium. After discharge to home, 30% of the patients had to be taken into long-term care or had died within 1 year of the delirium. PMID- 11263715 TI - Identifying dementia in the primary care practice. AB - BACKGROUND: The purpose of this study was to evaluate the utility (i.e., positive and negative predictive value) of the 7 Minute Screen in identifying patients with probable Alzheimer's disease (AD) in a primary care practice. A second objective was to estimate the number of undiagnosed AD patients in a typical primary care practice. METHODS: One hundred thirty-seven successive admissions (96%) of patients over the age of 60 to a primary care practice over a 53-day period who completed informed consent documents were administered the 7 Minute Screen. All patients who screened positive (n = 13) and a random sample of those who screened negative (n = 26) returned for full diagnostic evaluation. Positive predictive value (PPV) and negative predictive value (NPV) of the 7 Minute Screen were determined using the criterion standard of clinical diagnosis established by examination, history, and laboratory studies. Test-retest reliability and time for administration were also determined. RESULTS: Of the 137 patients evaluated, 13 screened positive and 124 screened negative. Eleven of the 13 patients who screened positive were willing to return to the primary care practice for follow up evaluation. A random sample of 26 patients who screened negative all agreed to return for follow-up evaluation. Of the 11 patients who screened positive and who returned for evaluation, 10 were subsequently diagnosed with probable AD. The remaining patient was diagnosed with mixed dementia. The caregivers of the two patients who refused to return were contacted and both indicated that the patients were having significant cognitive problems as verified by an activities of daily living scale. Of the 26 patients who screened negative, 25 were judged to be cognitively normal and the 26th was judged to have mild cognitive impairment. DISCUSSION: In successive admissions of patients over the age of 60 in a primary care practice, the 7 Minute Screen showed a PPV of 91% and an NPV of 96% in identifying patients who were subsequently identified with AD or other dementing disorder. These data suggest that this may be a useful instrument in identifying patients who should undergo diagnostic evaluation for AD and other dementing disorders. Additionally, extrapolation from the data in this practice suggests that there may be between 75 and 100 AD patients in the typical primary care practice, many of whom may not be diagnosed. PMID- 11263716 TI - Agitation in nursing home residents: the role of gender and social context. AB - We investigated the relationship among gender of resident, staff social interaction, and agitation in 46 (31 male and 15 female) nursing home residents with clinically significant agitation. Direct observations were conducted of resident behaviors and environmental contextual events using a computer-assisted, real-time observational system. The system recorded frequency, duration, and temporal sequencing of events. Results show that female residents displayed almost three times the amount of agitation as male residents (35% vs. 13% of total observation time, respectively), although men in the study were more likely to receive psychoactive drugs for their agitation. Staff spent similar amounts of time verbally interacting and touching male and female residents. Sequential analyses were conducted to examine the likelihood of staff verbal and touch interactions both preceding and following resident agitation using Bakeman and Quera's (1995) SDIS-GSEQ program. Results suggest that staff touch and verbal interaction elicit agitation in a significant proportion of residents. Once agitation occurs, staff were likely to respond by interacting verbally, but not physically, with the resident. PMID- 11263717 TI - Similarities in the disturbances in cortical information processing in alcoholism and aging: a pilot evoked potential study. AB - OBJECTIVE: To examine the hypothesis that chronic alcohol use causes accelerated aging of the brain. METHODS: The auditory evoked potentials (EPs) were compared in three groups of 10 subjects each: (a) middle-aged individuals meeting DSM-IV criteria for alcohol dependence, (b) age- and gender-matched group of healthy individuals, and (c) an older (>65 years) group of gender-matched healthy individuals. Multiple levels of cortical information processing were examined using EPs. Early stages of information processing, related to sensory gating and stimulus classification (P50, N100/P200), were studied using a paired-click paradigm. Later stages of information processing associated with memory upgrading and identification of novel stimuli (P300) were studied using an oddball paradigm. RESULTS: The amplitude and latency of the P300 of the alcoholic patients and the older healthy subjects differed significantly from those of the younger healthy group. Both groups showed changes that have been reported in association with aging. A tendency towards decreased sensory gating in later stages of information processing was noted in the aged healthy individuals. CONCLUSIONS: These data suggest that alcohol dependence may accelerate the aging process. The tendency towards a sensory gating deficit during the attentive phase of information processing in older healthy subjects requires further investigation because it may be a marker for an increased proneness to developing psychotic symptoms in that group. PMID- 11263718 TI - Risperidone in the treatment of patients with Alzheimer's disease with negative symptoms. AB - INTRODUCTION: Negative symptoms such as diminished initiative, drive, motivation, and emotional reactivity have been described in patients with Alzheimer's disease (AD). The purpose of this study was to retrospectively analyze the efficacy and tolerability of risperidone for the treatment of clinically significant positive and negative symptoms in AD. METHODS: We reviewed the charts of 50 community residing AD patients who had been treated in a specialized university-based dementia management clinic. Clinical data comparing baseline and 12 weeks of treatment were obtained by reviewing a series of rating scales that were recorded as part of a comprehensive behavioral assessment. RESULTS: Reviewed subjects had a mean age of 79.7 6 years and a mean of 12 +/- 3.6 years of school. Seventy percent of the subjects were female and the majority was White. The mean dose of risperidone prescribed was 1.3 +/- 0.6 mg per day (range from 0.5 mg to 3.0 mg). After 12 weeks of treatment, the severity of positive and negative symptoms was significantly reduced. Importantly, improvement in negative symptoms with the use of risperidone appeared to be independent of a positive treatment effect on positive symptoms. Risperidone had insignificant effects on both cognitive status and the emergence of extrapyramidal symptoms. CONCLUSION: This retrospective study demonstrates that risperidone appears to be efficacious in the treatment of clinically significant positive and negative symptoms in patients with AD. PMID- 11263719 TI - Serotonin 5-HT2A receptor binding in platelets from patients with Alzheimer's disease or vascular dementia. AB - It is well known that abnormalities in the brain serotonin system exist in patients with dementia. The present study was performed in order to investigate whether a peripheral serotonin system marker, the platelet 5-HT2A receptor, is affected in dementia. Thirty-eight patients with Alzheimer's disease (AD), 13 patients with vascular dementia, and 40 healthy controls were included in the study. There were no significant differences in receptor density for 5-HT2A receptor binding between the groups. Affinity of the radioligand to the receptor was significantly lower in AD than in vascular dementia and in the controls (p = .006 and p = .003, respectively), whereas there was no significant difference between the vascular dementia group and the control group. In 12 patients, treatment with citalopram was started due to depression or agitation. This treatment significantly reduced the Behavioral Pathology in Alzheimer's Disease Rating Scale scores (p = .001), but did not affect the platelet 5-HT2A receptor status. There was no correlation between 5-HT2A receptor status before treatment and the therapeutic effect of citalopram. The study indicates that platelet 5 HT2A receptor status is of limited value as a peripheral marker in dementia. PMID- 11263720 TI - Psychotic symptoms in Alzheimer's disease are not associated with more severe neuropathologic features. AB - Psychotic symptoms in Alzheimer's disease (AD) have been associated with increased rates of cognitive impairment and functional decline. Prior studies have been conflicting with regard to whether AD patients with psychosis (AD+P) have evidence of more severe neuropathologic findings at postmortem exam. We examined the severity of neuritic plaques and neurofibrillary tangles in six brain regions--middle frontal cortex, hippocampus, inferior parietal cortex, superior temporal cortex, occipital cortex, and transentorhinal cortex-in 24 AD+P subjects and 25 matched AD subjects without psychosis (AD-P). All analyses controlled for the presence of cortical Lewy bodies, and corrected for multiple comparisons. We found no significant associations between neuritic plaque and neurofibrillary tangle severity and AD+P, and no significant associations with any individual psychotic symptom. The association of AD+P with a more rapidly progressive course of AD appears to be mediated by a neuropathologic process other than increased severity of plaque and tangle formation. PMID- 11263721 TI - Heterothermy in elephant shrews, Elephantulus spp. (Macroscelidea): daily torpor or hibernation? AB - The physiological parameters of heterothermy (e.g. minimum body temperature and oxygen consumption, percentage metabolic reduction, and bout length) were measured in two species of Elephantulus elephant shrews (Elephantulus myurus and Elephantulus rozeti; Macroscelidea) as a function of ambient temperature. Both species displayed deep torpor whereby the body temperatures of ca. 5 degrees C and oxygen consumption as low as 2% of basal metabolic rate were attained. Torpor bout length (n = 57 bouts) never exceeded 24 h. These data are characteristic of both hibernation (minimum body temperature and metabolism) and daily torpor (bout length), and argue that these two physiological responses may not necessarily have separate evolutionary origins. PMID- 11263722 TI - Daily torpor in elephant shrews (Macroscelidea: Elephantulus spp.) in response to food deprivation. AB - Patterns of daily torpor were measured in response to photoperiod and food restriction at a constant temperature (18 degrees C) in two species of elephant shrew (Macroscelidea), Elephantulus rozeti (from Morocco) and Elephantulus myurus (from southern Africa). Body temperature was monitored continuously for ca. 3 months using temperature-sensitive telemeters. Under short photoperiods (8:16 L:D), both species entered spontaneous torpor on an ad libitum diet, but showed a higher frequency of induced torpor when food was restricted. Under long photoperiods (16:8 L:D), E. myurus could be induced to enter daily 'summer' torpor. A total of 378 torpor bouts were measured, none of which were longer in duration than 18 h. Under short photoperiods, arousal from torpor was associated with the onset of the photoperiod, whereas the time of entry was variable throughout the scotophase. However, E. myurus tended to phase shift torpor from the photophase to the scotophase under long photoperiods, despite displaying weak circadian amplitudes of body temperature indicative of a photophase rest phase. Both species lacked well-defined circadian amplitudes of body temperature, a pattern thought to be associated with polyphasic activity cycles characteristic of several Elephantuluis species. It was concluded that these and other patterns of torpor shown by Elephantulus show similarities with other small Afrotropical insectivores inhabiting semi-arid habitats or unpredictable environments. PMID- 11263723 TI - Water and electrolyte homeostasis and kidney function of desert-dwelling marsupial wallabies in Western Australia. AB - Prolonged drought, necessitating conservation of water, is one of the major environmental challenges faced by many Australian marsupials. Radioactive isotopes of water and sodium were used to assess the ability of two species of marsupial wallabies to maintain water and electrolyte balance during periods of extreme water deprivation in the arid Pilbara region of Western Australia. The spectacled hare-wallaby, Lagorchestes conspicillatus, has the lowest mass specific rate of water turnover at 27.5 ml.kg(-0.82).day(-1) yet reported for any mammal and was two to three orders of magnitude lower than that of the Rothschild's rock-wallaby, Petrogale rothschildi. Studies of renal function show that the hare-wallaby conserves water by producing a highly concentrated urine under the influence of lysine vasopressin (LVP), the anti-diuretic hormone (ADH) in macropodid marsupials. In contrast, rock-wallabies show unusual renal responses to water deprivation, with no change in LVP levels and a limited response to water deprivation involving a reduction in renal plasma flow and glomerular filtration rate, with no significant change in tubular function. Both species are able to maintain water and electrolyte homeostasis during periods of drought, highlighting the efficacy of their differing adaptive solutions to the problem of water scarcity, although the hare-wallaby is superior to the rock wallaby in this respect. Rock-wallabies appear to rely primarily on behavioural rather than physiological responses for their survival in the Pilbara and appear to be more vulnerable to extinction in the event of significant habitat modification. The secure nature of their rock habitat, however, means that they have suffered less than hare-wallabies in the recent past. PMID- 11263725 TI - Erythrocyte osmotic fragility of red (Macropus rufus) and grey (Macropus fuliginosus and Macropus giganteus) kangaroos and free-ranging sheep of the arid regions of Australia. AB - The mean corpuscular fragility (MCF) of erythrocytes may reflect phylogenetic characteristics as well as an animal's ability to respond to the osmotic challenges associated with cyclic dehydration and rehydration. This type of ecophysiological stress is commonly encountered by animals living in arid regions and low MCF may contribute to their ability to survive and thrive in these xeric habitats. The eastern grey kangaroo has only in recent times extended its range into the arid zone, and is considered a more mesic inhabitant than the red kangaroo. We therefore compared the ability of eastern grey kangaroos and red kangaroos to handle prolonged periods of water restriction, as well as the MCF of the erythrocytes of free-ranging red, eastern grey and western grey kangaroos found at the Fowlers Gap field station. In addition, the MCF of free-ranging sheep inhabiting the same pastures were used as an experimental control; they are phylogenetically unrelated yet are subjected to the same acclimatisation stresses. While red kangaroos exhibited greater tolerance of dehydration compared to eastern grey kangaroos, the MCF of all three kangaroo species was similar and more resilient to osmotic stresses (MCF, 130 mosmol/kg) than erythrocytes of sheep (MCF, 220 mosmol/kg). The MCF did not change with water restriction, however, the erythrocytes of long-term captive populations fed a comparatively better quality diet were more resistant to osmotic shock than the free-ranging animals. Phylogenetic commonality rather than ecophysiological responses to life in the arid zone appeared to influence MCF. The MCF values of sheep corresponded to that of other ovines; similarly the MCF of kangaroos concurred regardless of their preferred habitats. ecological history and differential success in the arid zone. PMID- 11263724 TI - Characterisation of cAMP-dependent protein kinase isoforms in the brain of the crab Chasmagnathus. AB - In the crab Chasmagnathus learning model, systemic administration of cAMP analogues that are specific activators or inhibitors of cAMP-dependent protein kinase (PKA) proved to respectively facilitate or impair long-term retention. The aims of the present work were to analyse PKA activity distribution in the crab brain and to characterise PKA isoforms. The neuropils from the eyestalk showed higher levels of induced PKA activity when compared with other neuropils of the central nervous system. Two PKA isoforms, homologous to mammalian PKA I and PKA II, were detected from central brain protein extracts using DEAE chromatography. Only PKA II was found in lateral protocerebrum extracts, suggesting a role of this isoform in the processing of visual inputs and in the integration of this information with other sensory inputs. PKA I was observed to be ten-fold more sensitive to cAMP than PKA II. cGMP induced a high activation of both PKA isoforms, similar to that obtained with cAMP. PKA I showed a two-fold greater sensitivity for cGMP than PKA II. An autophosphorylation assay was performed and a protein of 55 kDa, corresponding to phosphorylated R II regulatory subunit, was detected. The presence of a PKA I isoform with high sensitivity for cAMP in the central brain suggests a role of this subtype in long-term memory. PMID- 11263726 TI - Bioenergetics and RNA/DNA ratios in the common carp (Cyprinus carpio) under hypoxia. AB - Hypoxia caused by eutrophication occurs over large areas in aquatic systems worldwide. Common carp (Cyprinus carpio) exposed to hypoxia (1 mg.O2.l(-1) and 2 mg.O2.l(-1)) for 1 week showed a significant reduction in feeding rate, respiration rate, faecal production and nitrogenous excretion compared to those maintained at normoxia (7 mg.O2.l(-1)). Fish exposed to hypoxia showed negative scope for growth (SfG), but no significant difference in the specific growth rate was revealed after 1 week in both hypoxic groups. A significant reduction in RNA/DNA ratio was, however, clearly evident in the white muscle of the 1 mg.O2.l( 1) treatment group, but not in the 2 mg.O2.l(-1) treatment group. Both specific growth rate and RNA/DNA ratio were significantly reduced when fish were exposed to severe hypoxia (0.5 mg.O2.l(-1)) for 4 weeks. At all levels of hypoxia, growth reduction was accompanied by a significant decrease in RNA/DNA ratio in white muscle. Covariance analysis showed no significant difference between the slope of RNA/DNA ratio and growth rate under normoxic conditions and 0.5 mg. O2.l(-1) for 4 weeks (F= 1.036, P > 0.326), as well as 1.0 mg.O2.l(-1) and 2.0 mg.O2.l(-1) for 1 week (F = 0.457, P > 0.5), indicating that the RNA/DNA ratio serves as a biomarker of growth under all oxygen levels, at least under controlled experimental conditions. SfG also appears to be more sensitive than the RNA/DNA ratio in responding to hypoxia in fish. PMID- 11263727 TI - Maintenance nitrogen requirement of an obligate nectarivore, the honey possum, Tarsipes rostratus. AB - A nitrogen balance feeding trial was carried out with the marsupial honey possum, Tarsipes rostratus, using four pollen-honey diets varying in nitrogen content from 9.4 mg.g(-1) to 2.3 mg.g(-1) dry matter. The dietary maintenance nitrogen requirement (MNR) was determined by regression analysis as 89 +/- 21 mg N.kg( 0.75).day(-1) and the truly digestible MNR was 79 mg N.kg(-0.75).day(-1). Regressing nitrogen balance on daily nitrogen intake separately for ten males and seven females revealed that the slopes of the fitted lines did not vary significantly, but the difference in the intercepts approached significance. This suggests that the MNR for females may be lower than that of males. The nitrogen digestibility of the diet was 76% and the biological value (BV) was 58%. A comparison of the MNR of the honey possum with that of other marsupials shows that it is indeed much lower than that of herbivorous macropodid marsupials and is close to that of the sap- and gum-feeding sugar glider, Petarurus breviceps. The endogenous urinary nitrogen excretion (EUN) of the honey possum was 42 mg N.kg(-0.75.day(-1) and a regression analysis with other published data showed that the EUN per unit basal heat production is significantly lower than that of eutherian mammals. Measurements of the actual feeding rates of animals in the field, taken together with the low MNR, do not lend support to the hypothesis that the honey possum exhibits a reduced rate of reproduction due to a deficiency in dietary nitrogen. It is possible that the quality of nitrogen provided by pollen, as reflected in its composition of essential amino acids, may be a limiting factor. PMID- 11263728 TI - Energetics of calling and metabolic substrate use during prolonged exercise in the European treefrog Hyla arborea. AB - Rates of oxygen consumption and carbon dioxide release were measured in calling and resting European tree frogs using open-flow-through respirometry. The energetic cost of calling was high with an average of 1.076 ml O2/(g.h) at average call rates of 8,000 calls/h. The maximum factorial metabolic scopes averaged 24 with momentary peak values ranging between 5 and 41. There was a threefold difference in O2-consumption between individual males calling at the same rate. Respiratory quotients indicated that both lipids and carbohydrates were used to fuel calling. Carbohydrates provided the major fuel (69% on average) with dependence on carbohydrates increasing with call rate. In contrast to marathon runners, there was no shift in metabolic substrate use over a calling period of 2-3 h. PMID- 11263729 TI - The role of dietary fatty acids in the evolution of spontaneous and facultative hibernation patterns in prairie dogs. AB - The white-tailed prairie dog is a spontaneous hibernator which commences deep torpor bouts during early fall while the black-tailed prairie dog is a facultative hibernator that will only enter shallow torpor when stressed by cold and food deprivation. Plant oils rich in polyunsaturated fatty acids (PUFAs) enhance the duration and depth of mammalian torpor. Thus, we tested the hypothesis that black-tailed prairie dogs sampled in the field have less PUFAs in their diets and that the enhancement of torpor bouts by this species on a diet higher in PUFA is less profound than that by white-tailed prairie dogs. Individuals of both species fed a high PUFA diet: (1) entered torpor earlier, (2) had lower torpor body temperatures and (3) had longer bouts of torpor, compared to those on a low PUFA diet. However, the magnitude of this change was similar for both species. Additionally, the PUFA compositions of white adipose tissue (WAT) samples taken from individuals in the field were identical, indicating that diet PUFA contents for these two species were also equivalent. Therefore, while high PUFA diets can enhance hibernation by these species, it does not appear to explain the differences between spontaneous and facultative strategies. The rate of lipid peroxidation during torpor, however, was significantly higher in the WAT from white-tailed prairie dogs. Ancestral prairie dog species are spontaneous hibernators. Natural selection may have favored shallow, facultative hibernation with lower lipid peroxidation rates in the black-tailed prairie dogs as they radiated from the Rocky Mountains into the Great Plains. PMID- 11263730 TI - Perspective: transposable elements, parasitic DNA, and genome evolution. AB - The nature of the role played by mobile elements in host genome evolution is reassessed considering numerous recent developments in many areas of biology. It is argued that easy popular appellations such as "selfish DNA" and "junk DNA" may be either inaccurate or misleading and that a more enlightened view of the transposable element-host relationship encompasses a continuum from extreme parasitism to mutualism. Transposable elements are potent, broad spectrum, endogenous mutators that are subject to the influence of chance as well as selection at several levels of biological organization. Of particular interest are transposable element traits that early evolve neutrally at the host level but at a later stage of evolution are co-opted for new host functions. PMID- 11263731 TI - Sex-linked expression of a sexually selected trait in the stalk-eyed fly, Cyrtodiopsis dalmanni. AB - Recent theoretical and empirical work has suggested that the X chromosome may play a special role in the evolution of sexually dimorphic traits. We tested this idea by quantifying sex chromosome influence on male relative eyespan, a dramatically sexually selected trait in the stalk-eyed fly, Cyrtodiopsis dalmanni. After 31 generations of artificial sexual selection on eyespan:body length ratio, we reciprocally crossed high- with low-line flies and found no evidence for maternal effects; the relative eyespan of F1 females from high- and low-line dams did not differ. However, F1 male progeny from high-line dams had longer relative eyespan than male progeny from low-line dams, indicating X linkage. Comparison of progeny from a backcross involving reciprocal F1 males and control line females confirmed X-linked inheritance and indicated no effect of the Y chromosome on relative eyespan. We estimated that the X chromosome accounts for 25% (SE = 6%) of the change in selected lines, using the average difference between reciprocal F1 males divided by the difference between parental males, or 34%, using estimates of the number of effective factors obtained from reciprocal crosses between a high and low line. These estimates exceed the relative size of the X in the diploid genome of a male, 11.9% (SE = 0.3%), as measured from mitotic chromosome lengths. However, they match expectations if X-linked genes in males exhibit dosage compensation by twofold hyperactivation, as has been observed in other flies. Therefore, sex-linked expression of relative eyespan is likely to be commensurate with the size of the X chromosome in this dramatically dimorphic species. PMID- 11263732 TI - The evolution of trade-offs: effects of inbreeding on fecundity relationships in the cricket Gryllus firmus. AB - The evolution of traits is modulated by their interrelationships with each other, particularly when those relationships result in a fitness trade-off. In this paper we explore the consequences of genetic architecture on functional relationships between traits. Specifically, we address the consequences of inbreeding on these relationships. We show that the linear regression between two traits will not be affected if there is no dominance genetic variance in either trait, whereas the intercept but not the slope of the regression will change if there is dominance genetic variance in one trait only. We test the latter hypothesis using fecundity relationships in the cricket Gryllus firmus. Data from pedigree analysis and an inbreeding experiment show that there is significant dominance genetic variance in fecundity, but not head width (an index of body size) or dorsal longitudinal muscle (DLM) mass. Fecundity increases with head width, but decreases with DLM mass. As predicted, the intercepts of the regressions of fecundity on these two morphological traits decrease with inbreeding, but there is little or no change in slope. Gryllus firmus is wing dimorphic, with the macropterous (LW) morph having a lower fecundity than the micropterous (SW) morph. We hypothesize that the difference in fecundity arises primarily because of a competition for resources in the LW females between DLM maintenance (i.e., mass) and egg production. As a consequence, we predict that the fecundity within each morph should decline linearly with the inbreeding coefficient at the same rate in both morphs. The result of this will be a change in the relative fitness of the two morphs, that of the SW morph increasing with inbreeding. This prediction is supported. These results indicate that trade-offs will evolve and such changes will affect evolutionary trajectories by altering the pattern of relationships among fitness components. PMID- 11263733 TI - Molecular biogeography of cave life: a study using mitochondrial DNA from bathysciine beetles. AB - This study focuses on phylogenetic relationships in two distinct species assemblages of cave-dwelling beetles with similar disjunct distributions in the Pyrenees and Sardinia. One assemblage contains six species in the genera Ovobathysciola (four species) and Patriziella (two species) on Sardinia and one species of Anillochlamys in the Pyrenees. Species within the two Sardinian genera co-occur in the same karst area. Although, they are believed to be each others closest relative, they have very different body types (globular body with short appendages in Ovobathysciola; elongated body with long appendages in Patriziella), which are believed to reflect different degrees of adaptation to cave life. The other assemblage of Bathysciine beetles includes three species in the genus Speonomus in the Pyrenees and one on Sardinia. All the species are rare and many are endangered. One issue of particular interest was whether Ovobathysciola and Patriziella are reciprocally monophyletic or whether each of the Patriziella species evolved independently from the co-occurring Ovobathysciola species, as the similar morphology of the Patriziella species might be due to convergence rather than common descent. Based on DNA sequences of the cytochrome oxidase I (COI) region of the mtDNA, neither scenario was supported. Rather, the two Patriziella species are sister taxa embedded within the Ovobathysciola radiation. In addition, the well-dated geological history of this region allowed us to calibrate absolute rates of COI evolution, the first such estimates for any insect. Finally this study suggests that the evolutionary acquisition of typical cave adaptations (e.g., elongated body and appendages) may occur at about the same rate as loss of traits (e.g., eyes and pigmentation) associated with cave life. PMID- 11263734 TI - Isolation by distance in the Atlantic cod, Gadus morhua, at large and small geographic scales. AB - Genetic isolation by distance (IBD) has rarely been described in marine species with high potential for dispersal at both the larval and adult life-history stages. Here, we report significant relationships between inferred levels of gene flow and geographic distance in the Atlantic cod, Gadus morhua, at 10 nuclear restriction-fragment-length-polymorphism (RFLP) loci at small regional scales in the western north Atlantic region (< 1,600 km) that mirror those previously detected over its entire geographic range (up to 7,300 km). Highly significant allele frequency differences were observed among eight northwestern Atlantic populations, although the mean FST for all 10 loci was only 0.014. Despite this weak population structuring, the distance separating populations explained between 54% and 62% of the variation in gene flow depending on whether nine or 10 loci were used to estimate Nm. Across the species' entire geographic range, highly significant differences were observed among six regional populations at nine of the 10 loci (mean FST = 0.068) and seven loci exhibited significant negative relationships between gene flow and distance. At this large geographic scale, natural selection acting in the vicinity of one RFLP locus (GM798) had a significant effect on the correlation between gene flow and distance, and eliminating it from the analysis caused the coefficient of determination to increase from 17% to 62%. The role of vicariance was assessed by sequentially removing populations from the analysis and was found to play a minor role in contributing to the relationship between gene flow and distance at either geographic scale. The correlation between gene flow and distance detected in G. morhua at small and large spatial scales suggests that dispersal distances and effective population sizes are much smaller than predicted for the species and that the recent age of populations, rather than extensive gene flow, may be responsible for its weak population structure. Our results suggest that interpreting limited genetic differences among populations as reflecting high levels of ongoing gene flow should be made with caution. PMID- 11263736 TI - Mating systems, sperm competition, and the evolution of sexual dimorphism in birds. AB - Comparative analyses suggest that a variety of factors influence the evolution of sexual dimorphism in birds. We analyzed the relative importance of social mating system and sperm competition to sexual differences in plumage and body size (mass and tail and wing length) of more than 1,000 species of birds from throughout the world. In these analyses we controlled for phylogeny and a variety of ecological and life-history variables. We used testis size (corrected for total body mass) as an index of sperm competition in each species, because testis size is correlated with levels of extrapair paternity and is available for a large number of species. In contrast to recent studies, we found strong and consistent effects of social mating system on most forms of dimorphism. Social mating system strongly influenced dimorphism in plumage, body mass, and wing length and had some effect on dimorphism in tail length. Sexual dimorphism was relatively greater in species with polygynous or lekking than monogamous mating systems. This was true when we used both species and phylogenetically independent contrasts for analysis. Relative testis size was also related positively to dimorphism in tail and wing length, but in most analyses it was a poorer predictor of plumage dimorphism than social mating system. There was no association between relative testis size and mass dimorphism. Geographic region and life history were also associated with the four types of dimorphism, although their influence varied between the different types of dimorphism. Although there is much interest in the effects of sperm competition on sexual dimorphism, we suggest that traditional explanations based on social mating systems are better predictors of dimorphism in birds. PMID- 11263735 TI - Phylogeography of the tailed frog (Ascaphus truei): implications for the biogeography of the Pacific Northwest. AB - Tailed frogs are distributed in high-gradient streams within the disjunct mesic forests of the Pacific Northwest and represent the basal lineage of the anurans. We sequenced 1,530 nucleotides of the mitochondrial cytochrome b and NADH dehydrogenase subunit two genes from 23 populations and used parsimony, maximum likelihood, and nested-clade analyses to estimate relationships among populations and infer evolutionary processes. We found two divergent haplotype clades corresponding with inland Rocky Mountain populations and coastal populations and separated by up to 0.133 substitutions per site. Within the coastal assemblage, haplotypes formed clades by mountain range with 0.010-0.024 substitutions per site divergence among populations. Inland haplotypes exhibited minimal genetic structure, with the exception of 0.021 substitutions per site distance between populations from the East Fork of the South Fork of the Salmon River and all other inland haplotypes. The magnitude of divergence between inland and coastal populations, as well as the paleobotanical record, suggest isolation of these lineages occurred during the late Miocene to early Pliocene, probably in response to the rise of the Cascade Mountains. Genetic structure within coastal and inland populations is consistent with isolation in refugia during the late Pliocene and early Pleistocene. Closely related inland haplotypes reflect range expansion following glaciation. The depth of divergence between inland and coastal populations supports the persistence of mesic forests within the inland Pacific Northwest throughout the Pleistocene and is congruent with patterns found in several other mesic forest species. Based on mitochondrial divergence and previous allozyme and morphological data, we recommend recognition of inland populations as a distinct species, Ascaphus montanus. PMID- 11263737 TI - The evolution of sexual size dimorphism in the house finch. III. Developmental basis. AB - Sexual size dimorphism of adults proximately results from a combination of sexually dimorphic growth patterns and selection on growing individuals. Yet, most studies of the evolution of dimorphism have focused on correlates of only adult morphologies. Here we examined the ontogeny of sexual size dimorphism in an isolated population of the house finch (Carpodacus mexicanus). Sexes differed in growth rates and growth duration; in most traits, females grew faster than males, but males grew for a longer period. Sexual dimorphism in bill traits (bill length, width, depth) and in body traits (wing, tarsus, and tail length; mass) developed during different periods of ontogeny. Growth of bill traits was most different between sexes during the juvenile period (after leaving the nest), whereas growth of body traits was most sexually dimorphic during the first few days after hatching. Postgrowth selection on juveniles strongly influenced sexual dimorphism in all traits; in some traits, this selection canceled or reversed dimorphism patterns produced by growth differences between sexes. The net result was that adult sexual dimorphism, to a large degree, was an outcome of selection for survival during juvenile stages. We suggest that previously documented fast and extensive divergence of house finch populations in sexual size dimorphism may be partially produced by distinct environmental conditions during growth in these populations. PMID- 11263738 TI - An unusual sex-determination system in South American field mice (Genus Akodon): the role of mutation, selection, and meiotic drive in maintaining XY females. AB - The mechanism of sex determination in mammals appears highly conserved: the presence of a Y chromosome triggers the male developmental pathway, whereas the absence of a Y chromosome results in a default female phenotype. However, if the Y chromosome fails to initiate the male pathway (referred to as Y*), XY* females can result, as is the case in several species of South American field mice (genus Akodon). The breeding genetics in this system inherently select against the Y* chromosome such that the frequency of XY* females should decrease rapidly to very low frequencies. However, in natural populations of Akodon, XY* females persist at substantial frequencies; for example, 10% of females are XY* in A. azarae and 30% in A. boliviensis. We develop a mathematical model that considers the potential roles of three evolutionary forces in maintaining XY* females: Y-to-Y* chromosome transitions (mutation), chromosome segregation distortion (meiotic drive), and differential fecundity (selection). We then test the predictions of our model using data from breeding colonies of A. azarae. We conclude that any single force is inadequate to maintain XY* females. However, a combination of segregation bias of the male and female Y chromosomes during spermatogenesis/oogenesis and increased fecundity in XY* females could account for the observed frequencies of XY* females. PMID- 11263739 TI - A genetic interpretation of ecologically dependent isolation. AB - Hybrids may suffer a reduced fitness both because they fall between ecological niches (ecologically dependent isolation) and as a result of intrinsic genetic incompatibilities between the parental genomes (ecologically independent isolation). Whereas genetic incompatibilities are common to all theories of speciation, ecologically dependent isolation is a unique prediction of the ecological model of speciation. This prediction can be tested using reciprocal transplants in which the fitness of various genotypes is evaluated in both parental habitats. Here we expand a quantitative genetic model of Lynch (1991) to include two parental environments. We ask whether a sufficient experimental design exists for detecting ecologically dependent isolation. Analysis of the model reveals that by using both backcrosses in both parental environments, environment-specific additive genetic effects can be estimated while correcting for any intrinsic genetic isolation. Environment-specific dominance effects can also be estimated by including the F1 and F2 in the reciprocal transplant. In contrast, a reciprocal transplant comparing only F1s or F2s to the parental species cannot separate ecologically dependent from intrinsic genetic isolation. Thus, a reduced fitness of F1 or F2 hybrids relative to the parental species is not sufficient to demonstrate ecological speciation. The model highlights the importance of determining the contribution of genetic and ecological mechanisms to hybrid fitness if inferences concerning speciation mechanisms are to be made. PMID- 11263740 TI - Sex among the flowers: the distribution of plant mating systems. AB - Previous reviews of plant outcrossing rate survey data have agreed that predominant selfing and predominant outcrossing are alternative stable states of mating system evolution. We reanalyzed the most recent data and plot outcrossing rates as a continuous variable rather than as a class variable. Wind-pollinated species are indeed bimodal. However, the shape of the distributions for animal pollinated species reveals that intermediate rates of outcrossing are common (49% of species fall between 20% and 80% outcrossing). Consequently, we suggest that mating system is best considered a continuous rather than a discrete character of plant populations. PMID- 11263741 TI - Locomotor performance of Drosophila melanogaster: interactions among developmental and adult temperatures, age, and geography. AB - We explored the extent to which a phenotypic trait (walking speed) of Drosophila melanogaster is influenced by population, developmental temperature, adult temperature, and age. Our goals were to estimate the importance of these factors and to test the beneficial acclimation hypothesis. We measured speed of flies from two populations (the Congo and France) that developed at different temperatures (18, 25, and 29 degrees C) and were tested at different temperatures (18, 25, and 29 degrees C) and ages (2, 7, 13 days). Not surprisingly, speed increased strongly with test temperature. Speed was generally greatest for flies reared at an intermediate developmental temperature, contrary to the beneficial acclimation hypothesis, which predicts that speed would be greatest when influenced by interactions involving population. For example, speed was greatest for flies from France that developed at a low temperature, but for flies from the Congo that developed at a high temperature. The impact of developmental temperature declined with age. Surprisingly, speed actually increased with age for flies raised and maintained at a low temperature, but decreased with age for flies raised and maintained at an intermediate or at a high temperature. Thus, walking performance is highly dynamic phenotypically, complicating potential attempts to predict responses to selection on performance. PMID- 11263742 TI - Parental and developmental temperature effects on the thermal dependence of fitness in Drosophila melanogaster. AB - Cross-generational effects refer to nongenetic influences of the parental phenotype or environment on offspring phenotypes. Such effects are commonly observed, but their adaptive significance is largely unresolved. We examined cross-generational effects of parental temperature on offspring fitness (estimated via a serial-transfer assay) at different temperatures in a laboratory population of Drosophila melanogaster. Parents were reared at 18 degrees C, 25 degrees C, or 29 degrees C (Tpar) and then their offspring were reared at 18 degrees C, 25 degrees C, or 29 degrees C (Toff) to evaluate several competing hypotheses (including an adaptive one) involving interaction effects of parental and offspring temperature on offspring fitness. The results clearly show that hotter parents are better; in other words, the higher the temperature of the parents, the higher the fitness of their offspring, independent of offspring thermal environment. These data contradict the adaptive cross-generational hypothesis, which proposes that offspring fitness is maximal when the offspring thermal regime matches the parental one. Flies with hot parents have high fitness seemingly because their own offspring develop relatively quickly, not because they have higher fecundity early in life. PMID- 11263743 TI - Models of sexual selection on a quantitative genetic trait when preference is acquired by sexual imprinting. AB - The evolution of a quantitative genetic trait under stabilizing viability selection and sexual selection is modeled for a polygynous species in which female mating preferences are acquired by sexual imprinting on the parents and by exposure to the surviving population at large. Stabilizing viability selection acts equally on both sexes in the case of a sexually monomorphic trait and on males only in the case of a dimorphic trait. A genetically fixed sensory or perceptual bias defines the origin of the scale on which the trait is measured, and the possibility is incorporated that female preferences may deviate asymmetrically from the familiar-either toward or away from this origin. When viability selection is strong relative to sexual selection, the models predict that the mean trait value will evolve to the viability optimum. With intermediate ratios of the strength of viability to sexual selection, a stable equilibrium can occur on either side of this viability optimum, depending on the direction of asymmetry in female preferences. When viability selection is relatively weak and certain other conditions are also satisfied, runaway selection is predicted. PMID- 11263744 TI - Evolutionary adaptation to temperature. VIII. Effects of temperature on growth rate in natural isolates of Escherichia coli and Salmonella enterica from different thermal environments. AB - Are enteric bacteria specifically adapted to the thermal environment of their hosts? In particular, do the optimal temperatures and thermal niches of the bacterial flora reflect seasonal, geographic, or phylogenetic differences in their hosts' temperatures? We examined these questions by measuring the relationship between the temperature-dependent growth rates of enteric bacteria in a free-living ectothermic host. We sampled two species of enteric bacteria (Escherichia coli and Salmonella enterica) from three natural populations of slider turtles (Trachemys scripta elegans) seasonally over two years. Despite pronounced differences in turtle body temperatures at different seasons and in different locations, we found no evidence that the thermal growth profiles of these bacteria mirrored this variation. Optimal temperatures and maximal growth rates in rich medium were nearly the same for both bacterial species (35-36 degrees C, 2.5 doublings per hour). The thermal niche (defined as the range of temperatures over which 75% of maximal growth rate occurred) was slightly higher for E. coli (28.5-41.0 degrees C) than for S. enterica (27.7-39.8 degrees C), but the niche breadth was about the same for both. We also measured the thermal dependence of growth rate in these same bacterial species isolated from mammalian hosts. Both bacterial species had temperatures of maximal growth and thermal niches that were about 2 degrees C higher than those of their respective conspecifics sampled from turtles; niche breadths were not different. These data suggest that these bacterial species are thermal generalists that do not track fine-scale changes in their thermal environments. Even major differences in body temperatures, as great as those between ectothermic and endothermic hosts, may result in the evolution of rather modest changes in thermal properties. PMID- 11263745 TI - Patterns of association between crucifers and their flower-mimic pathogens: host jumps are more common than coevolution or cospeciation. AB - Morphological and molecular phylogenies of animal parasites have often shown parallel cladogenesis, supporting hypotheses of coevolution. Few studies of the phylogenetic history for plants and their pathogens exist. Gene-for-gene interactions suggest that plant pathogens ought to have similar phylogenetic histories as their hosts. However, high dispersability combined with an inability to choose to leave if an inappropriate host has been landed on could increase the likelihood of host jumps and thus decrease phylogenetic congruence between plant pathogens and their hosts. In this study, I examined the pattern of association between the flower-mimicking crucifer rusts and their hosts by comparing independent host phylogenies (based on both cpDNA trnL-F introns and nuclear internal transcribed spacer [ITS] sequences) with that of their rust pathogens (based on ITS sequences). The expectation was that if the pathogens coevolved or cospeciated with their hosts, then their phylogenies should be congruent. Host tracking coevolution can be differentiated from cospeciation by examining the times of divergence: If the pathogens are younger than the hosts, then it is likely that host tracking has occurred. For the crucifer rusts and their hosts, there was little evidence of parallel cladogenesis, suggesting that both cospeciation and coevolutionary tracking are rare. Instead, the most common pattern was one of host jumps to geographically associated taxa. There are at least three factors that may have contributed to the geographic structuring of the data. First, along the east-west transect stretching from the Rocky Mountains to California, large differences in rainfall and the timing of rainfall may reduce long-distance gene flow. Second, although dispersal of infectious spores is by wind, sexual reproduction of these fungi depends on insects, which move short distances. Third, host shifts are most likely to occur to geographically available taxa. Any species that grows adjacent to infected plants will be exposed to millions of spores, and the probability of eventual infection by a new mutant increases with greater exposure. Thus, patterns of association between the crucifers and their flower-mimic pathogens reflect jumps to geographically available hosts, which are not necessarily those that are most closely related. PMID- 11263747 TI - Host-plant adaptation in an herbivorous marine amphipod: genetic potential not realized in field populations. AB - Evolutionary responses of herbivores to their host plants depend not only on selection from plants, but also on the genetic basis of traits relating to host use. The genetic basis of such traits has been investigated extensively among terrestrial insect herbivores, but has received almost no attention among marine herbivores. We tested whether performance traits in the herbivorous marine amphipod Peramphithoe parmerong display heritable variation and, for the first time for a marine herbivore, whether selection has resulted in local adaptation to host plants on two spatial scales. Peramphithoe parmerong displayed heritable genetic variation for survival on two host macroalgae, the high-quality Sargassum linearifolium and the poor-quality Padina crassa, and for growth on S. linearifolium. Differences in performance on different hosts thus have the potential to select for differential use of hosts by this amphipod. Despite this potential, there was no evidence among field populations of local adaptation to host algae on either scale tested: between hosts within a site or among sites differing in algal species composition. Within a site, amphipods were not more likely to prefer or perform better on the host on which they were collected. Similarly, amphipods collected from sites in which P. crassa was present were not more likely to perform well on this host than amphipods collected from sites where this alga was not found. Ecological factors that may explain the persistence of P. parmerong on P. crassa and the possibility of phylogenetic constraints on host use by P. parmerong are discussed. PMID- 11263746 TI - A molecular phylogenetic analysis of reproductive trait evolution in the soft coral genus Alcyonium. AB - The soft coral genus Alcyonium is among the most reproductively diverse invertebrate taxa known: The genus includes species that vary both in mode of reproduction (including broadcast spawners, internal brooders, and external brooders) and sexual expression (gonochores, hermaphrodites, and a unisexual parthenogen). Such diversity offers a unique opportunity to examine associations between reproductive and morphological traits in a phylogenetic context. We used an approximately 900-bp sequence of the nuclear ribosomal gene complex spanning the internal transcribed spacer (ITS) regions to construct a molecular phylogeny for 14 European and North American species of Alcyonium onto which we mapped the known distribution of reproductive and morphological traits. The phylogeny suggests that hermaphroditism or parthenogenesis has evolved independently at least twice in this genus, and always in internally brooding species. Broadcast spawning and external brooding only occur in species with large colony size, whereas all species with small colony size brood their larvae internally. Internal brooding and small size appear to be ancestral in this genus; if this is the case, an association between broadcast spawning and large colony size has evolved independently in at least two clades. This tendency of small adults to brood their larvae while large adults broadcast spawn them into the plankton has been observed in a variety of solitary invertebrate taxa, but to date has not been documented in any other colonial invertebrates. Moreoever, it has been suggested that organisms with a colonial growth form should not experience the allometric constraints on brood space that have been proposed to explain the association between adult size and mode of reproduction in solitary organisms. Unlike many other colonial groups, however, module (polyp) size is strongly correlated with colony size in Alcyonium, and constraints on brooding may be imposed by module, rather than colony, allometry. The very close genetic relationship (< 1% sequence divergence) and shared polymorphisms among A. digitatum (a large, gonochoric broadcast spawner), A. siderium, and A. sp. A (intermediate-sized and small hermaphroditic, internal brooders) suggest that evolutionary transitions between broadcast spawning and brooding and between gonochorism and hermaphroditism can occur easily and rapidly in this group. PMID- 11263748 TI - Cryptic reproductive isolation in the Drosophila simulans species complex. AB - Forms of reproductive isolation that act after copulation but before fertilization are potentially important components of speciation, but are studied only infrequently. We examined postmating, prezygotic reproductive isolation in three hybridizations within the Drosophila simulans species complex. We allowed females to mate only once, observed and timed all copulations, dissected a subset of the females to track the storage and retention of sperm, examined the number and hatchability of eggs laid after insemination, counted all progeny produced, and measured the longevity of mated females. Each of the three hybridizations is characterized by a different set of cryptic barriers to heterospecific fertilization. When D. simulans females mate with D. sechellia males, few heterospecific sperm are transferred, even during long copulations. In contrast, copulations of D. simulans females with D. mauritiana males are often too short to allow sperm transfer. Those that are long enough to allow insemination, however, involve the transfer of many sperm, but only a fraction of these heterospecific sperm are stored by females, who also lay fewer eggs than do D. simulans females mated with conspecific males. Finally, when D. mauritiana females mate with D. simulans males, sperm are transferred and stored in abundance, but are lost rapidly from the reproductive tract and are therefore used inefficiently. These results add considerably to the list of reproductive isolating mechanisms in this well-studied clade and possibly to the list of evolutionary processes that could contribute to their reproductive isolation. PMID- 11263749 TI - Species-specific genitalic copulatory courtship in sepsid flies (Diptera, Sepsidae, Microsepsis) and theories of genitalic evolution. AB - Males of Microsepsis eberhardi and M. armillata use their genitalic surstyli to rhythmically squeeze the female's abdomen with stereotyped movements during copulation. Squeezing movements did not begin until intromission had occurred and, contrary to predictions of the conflict-of-interest hypothesis for genitalic evolution, did not overcome morphological or behavioral female resistance. Contrary to predictions of the lock-and-key hypothesis, female morphology was uniform in the two species and could not mechanically exclude the genitalia of either species of male. The complex pattern of squeezing movements differed between the two species as predicted by the sexual selection hypothesis for genitalic evolution. Also, evolutionarily derived muscles and pseudoarticulations in the male's genitalic surstyli facilitated one type of movement, whose patterns were especially distinct. The data support the hypothesis that the male surstyli evolved to function as courtship devices. PMID- 11263750 TI - The mouse light-dark paradigm: a review. AB - 1. The light/dark paradigm is based on the innate aversion of rodents to brightly illuminated areas and on the spontaneous exploratory behaviour of the animals, applying mild stressors i.e. novel environment and light. The test apparatus consists of a small dark secure compartment (one third) and a large illuminated aversive compartment (two thirds). 2. The test was developed with male mice. The strain, weight and age may be crucial factors. 3. The extent to which an anxiolytic compound can facilitate the exploratory activity depends on the baseline level in the control group. Differences between the type and severity of external stressors might account for variable results reported by different laboratories. 4. In conclusion, the black and white test may be useful to predict anxiolytic-like or anxiogenic-like activity in mice. Transitions have been reported to be an index of activity-exploration because of habituation over time and the time spent in each compartment to be a reflection of aversion. Classic anxiolytics (benzodiazepines) as well as the newer anxiolytic-like compounds (e.g. serotonergic drugs) can be detected using this paradigm. It has the advantages of being quick and easy to use, without requiring the prior training of animals. PMID- 11263751 TI - Anticipation, imprinting, trinucleotide repeat expansions and psychoses. AB - 1. Since 1991, approximately 20 trinucleotide repeat expansion type neurodegenerative disorders have been reported. They are clinically characterized by anticipation, i.e., worsening severity or earlier age at onset with each succeeding generation for an inherited disease, and imprinting, i.e., a process whereby specific genes are differentially marked during parental gametogenesis, resulting in the differential expression of these genes in the embryo and adult. 2. The phenomenon of anticipation in psychoses has been pointed out since the 19th century; however, it was ignored because no one knew the genetic mechanism underlying this type of inheritance pattern at the time, and because of several possible biases. 3. The discovery of trinucleotide repeat expansion diseases has reawakened interest in the phenomenon of anticipation in psychiatric diseases. Anticipation has been confirmed in schizophrenia, mood disorders, and anxiety disorders in much more sophisticated manners, although still not perfectly. 4. Molecular approaches as well as clinical ones have been taken to reveal the involvement of trinucleotide repeat expansion mechanism in psychoses by means of direct analyses of candidate genes, RED and DIRECT. Most efforts have been made for CAG type trinucleotide repeats. So far, direct analyses have failed to reveal pathogenic gene(s). There were several positive RED data at first, however, nowadays there seems to be a tendency of much more negative results. The DIRECT results did not support trinucleotide repeat expansions mechanism in psychoses either. One plausable explanation for the 'false positive' result is the presence of CAG trinucleotide repeats which are highly polymorphic but not associated with an obvious abnormal phenotype. Screening for trinucleotide repeats other than ones of the CAG type remained to be performed. PMID- 11263752 TI - Overview and perspective on the therapy of Alzheimer's disease from a preclinical viewpoint. AB - 1. Drugs effective in Alzheimer's disease (AD) should have several aims: to improve the cognitive impairment, control the behavioural and neurological symptoms, delay the progression of the disease, and prevent the onset. In order to attain these targets, cell and animal models are needed on which to test pathogenetic hypothesis and demonstrate the potential effectiveness of new drugs. This overview examines the results obtained in animal models. They are the link between the molecular and biochemical studies on the disease and the reality of human pathology. 2. The development of animal models reproducing the complexity of AD pathogenetic mechanisms and clinical symptoms still represents a challenge for the preclinical investigators. Moreover, the succession of different animal models well documents the progressive widening of our knowledge of the disease with the identification of new therapeutic targets. 3. The main animal models are listed, and their contribution to the understanding of the pathogenic mechanisms and development of the drugs presently used in AD therapy is described. Moreover, their role in the study of future drugs is analysed 4. Preclinical studies on cholinesterases and animal models mimicking the cholinergic hypofunction occurring in AD have been instrumental in developing cholinesterase inhibitors, which are the only recognised drugs for the symptomatic treatment of AD. 5. Artificially created beta-amyloid (A beta) deposits in normal rats, and transgenic mice overexpressing amyloid precursor protein (APP) are the models on which the future treatment are tested. They are aimed to prevent formation of A beta deposits or its transformation in neuritic plaques. 6. Models of brain inflammation, aging animals, and models of brain glucose and energy metabolism impairment make it possible to identify and assess the activity of anti inflammatory agents, antioxidants, ampakines and other potentially active agents. 7. It is concluded that the present level of information on AD could never have been reached without preclinical studies, and the development of new drugs will always require extensive preclinical investigations. PMID- 11263754 TI - Neuroscience research in AIDS. AB - 1. The human immunodeficiency virus invades the central nervous system early after infection where it later gives rise to cognitive, motor, and behavioral manifestations in children and adults. 2. Ranging from mild impairments to frank dementia, CNS manifestations can be diagnosed and measured with standard neuropsychological test batteries. 3. Great strides have been made with treatment: CNS manifestations are treatable, as are depression, psychosis, and delirium which sometimes accompany HIV disease at different stages. 4. With startling advances in antiretroviral therapy and lower mortality, patients face a constellation of new concerns stemming from HIV's transformation to a more chronic disease. 5. There are many compelling research directions ahead, including the psychosocial impact of living with HIV as a chronic disease, the development of medications expressly targeted to the CNS, and basic research on neuropathogenesis, including trafficking of virus into the CNS. PMID- 11263753 TI - Role of tonically-active neurons in the control of striatal function: cellular mechanisms and behavioral correlates. AB - 1. The striatum is primarily involved in motor planning and motor learning. Human diseases involving its complex circuitry lead to movement disorders such as Parkinson's disease (PD) and Huntington's disease (HD). Moreover the striatum has been involved in processes linked to reward, cognition and drug addiction. 2. The high content of acetylcholine (ACh) found in the striatum is due to the presence of cholinergic interneurons. The intrinsic electrical and synaptic properties of these interneurons have been recently characterized. However, their functional significance is far from being fully elucidated. 3. In vivo electrophysiological experiments from behaving monkeys have identified these cholinergic interneurons as "Tonically Active Neurons" (TANs). They are activated by presentation of sensory stimuli of behavioral significance or linked to reward. 4. Experimental evidence showed that integrity of the nigrostriatal dopaminergic system is essential for TANs to express learned activity. 5. PD is known to be due to the loss of the nigrostriatal dopaminergic pathway and the ensuing imbalance between the content of dopamine and acetylcholine in the striatum. This evidence supports the hypothesis that cholinergic interneurons, or TANs, play a key role in the modulation of striatal function. PMID- 11263755 TI - Computational approaches to the architecture and operations of the prefrontal cortical circuit for working memory. AB - 1. This article reviews recent progress in the computational studies towards the architecture and operations of the prefrontal cortical circuit, which are keys to understand the mechanisms of working memory processing. 2. The recurrent excitatory connections form closed-loop circuits, which contribute to the sustainment of delay-period activity. These connections subserve the cortical amplification of the activity. 3. Recent experimental studies (Wilson et al. 1994; Rao et al. 1999, 2000) suggested that at least two architectonically distinct types of intracortical inhibition, isodirectional and cross-directional inhibition, play significant roles in the formation of memory fields. 4. Computer simulations of a prefrontal cortical circuit model (Tanaka 1999, 2000a) showed that the isodirectional inhibition in the model regulated the amplitude of memory fields (i.e., the maximum firing rate) while the cross-directional inhibition contributed to the sharpening of the memory fields or the tuning curves. 5. The above characteristics enable the prefrontal cortical circuit to control memory fields, which would be necessary to general working memory processing. It would also be interesting to know whether different subtypes of the interneurons have distinct roles. 6. Another important issue is how neuromodulators contribute to working memory processing. Recent computer simulations by Durstewitz et al. (1999, 2000) showed that stronger dopamine action required stronger intervening input to destroy working memory, suggesting that dopamine contributes to the stabilization of working memory representation. 7. Further elucidation of these issues based on more detailed anatomical data of the cortical circuitry would make the architecture and operations of the prefrontal cortical circuit be more clearly described. PMID- 11263756 TI - Review of the next generation of Alzheimer's disease therapeutics: challenges for drug development. AB - 1. AD is believed to stem from dysfunctional cholinergic signaling in the regions of the brain associated with memory and cognition. 2. The occurrence of AD in afflicted individuals correlates with an increase in the accumulation of A beta rich senile plaques and neurofibrillary tangles in the brain. 3. Currently, the only FDA-approved AD therapies are a group of acetylcholinesterase inhibitors which slow the turnover of the neurotransmitter acetylcholine in the synapse. 4. Many other compounds which target other aspects of the disease, such as reducing neuronal damage and limiting oxidation, are in clinical trials. These include monoamine oxidase (MAO-B) inhibitors, NSAIDs, antioxidants and estrogen, among others. 5. Recent research discoveries have more completely defined the molecular nature of AD, and are generating new approaches for treatment. One idea is to limit the ability of the protein tau to become phosphorylated in hopes that this will limit the formation of neurofibrillary tangles in the brain. 6. A separate approach that is being pursued is to prevent formation and accumulation of A beta plaques. This may be accomplished by either regulating gamma-secretase activity, or using anti-beta-amyloid antibodies to reduce the size of existing plaques. 7. Employing improved procedural and technological approaches during clinical trials, such as bridging studies, dynabridge studies and PET analysis, promises to streamline the drug development process. 8. The use of biomarkers and MRI analysis may be an effective means by which to identify the disease early. Consequently, early intervention treatment therapies may be an effective way of delaying onset of the disease. 9. Long term AD studies, particularly those focusing on the MCI population, are likely to provide statistically valid results using a smaller study population. PMID- 11263757 TI - Pharmacological evidence supporting a role for galanin in cognition and affect. AB - 1. Galanin is localized in brain pathways involved in both cognition and affect. 2. Galanin has inhibitory actions on a variety of memory tasks including the Morris water maze, delayed nonmatching to position, T-maze delayed alternation, starburst maze, passive avoidance, active avoidance, and spontaneous alternation. 3. Galanin may inhibit learning and memory by inhibiting neurotransmitter release and neuronal firing rate. 4. Two signal transduction mechanisms through which galanin exerts its inhibitory actions are the inhibition of phosphatidyl inositol hydrolysis and the inhibition of adenylate cyclase. 5. Galanin released during periods of burst firing from noradrenergic locus coeruleus terminals in the ventral tegmental area (VTA) may lead to symptoms of depression through inhibition of dopaminergic VTA neurons. 6. Intraventricular galanin has anxiolytic effects in a punished drinking test. Intra-amygdala galanin has anxiogenic effects in a punished drinking test. PMID- 11263758 TI - On the role of prefrontal cortex glutamate for the antithetical phenomenology of obsessive compulsive disorder and attention deficit hyperactivity disorder. AB - 1. The objective of the present study was to compare the phenomenology and pathophysiology of obsessive compulsive disorder (OCD) and attention deficit hyperactivity disorder/deficits in attention, motor control and perception (ADHD/DAMP). 2. Through detailed studies of the literature on OCD and ADHD/DAMP, the phenomenology of these two conditions is compared, and possible underlying pathophysiological mechanisms involving interactions between glutamate, dopamine, serotonin and acetylcholine are discussed, with emphasis on OCD. The present paper also discusses possible mechanisms of action for current pharmacological treatments of OCD and ADHD, as well as possible future treatment strategies for these disorders. 3. OCD and ADHD/DAMP are common neuropsychiatric conditions which in many regards appear to be each other's antipodes with respect to clinical manifestations, associated personality traits and brain biochemistry, notably prefrontal cortical glutamate activity. Future pharmacological treatments of these disorders may involve manipulations with glutamate, dopamine D , serotonin 2A and nicotine receptors. 4. It appears that OCD is a hyperglutamatergic and ADHD a hypoglutamatergic condition, with prefrontal brain regions being especially affected. PMID- 11263759 TI - Nicotine induced behavioral locomotor sensitization. AB - 1. Nicotine behavioral sensitization of locomotor activity was investigated in adult female Sprague Dawley rats. Five different experiments were performed with nicotine in various doses of 0.1, 0.32, or 1.0 mg/kg i.p. These included: 1) effects of daily nicotine for 6 days, 2) effects of once per week nicotine for 3 weeks, 3) effects of MK-801 on nicotine-induced locomotor activity, 4) effects of dexamethasone on nicotine-induced locomotor activity, 5) induction of tolerance to nicotine-induced locomotor sensitization and lack of cross tolerance to caffeine. 2. Locomotor activity was measured with a photoelectric computerized system. The first dose of nicotine (0.32 mg/kg) induced marked locomotor depression. Once daily injection of 0.32 mg/kg of nicotine for 6 days produced tolerance to its depressant effects and sensitized the rats to its stimulant effects. Three once weekly doses of 0.32 mg/kg of nicotine also produced tolerance to its depressant effects and some locomotor stimulation. 3. Daily pretreatment for 5 days with a dose of 0.18 mg/kg of MK-801 i.p. partially antagonized the locomotor depressant and stimulant actions of nicotine. 4. Dexamethasone (1 mg/kg i.p.) daily pretreatment barely reduced nicotine locomotor depression and only very slightly enhanced locomotor stimulation. 5. Accumulating doses of 0.32 and 1.0 mg/kg b.i.d. of nicotine produced tolerance to its locomotor stimulant effects in rats previously sensitized to 0.32 mg/kg. There was no cross-tolerance to 32 mg/kg of caffeine citrate in previously sensitized animals tolerant to the stimulant effects of nicotine. PMID- 11263760 TI - Alzheimer's disease: current and future therapeutic perspectives. AB - 1. A better understanding of the pathophysiology of AD has been made possible through population-based epidemiological studies, human genetic and post-mortem studies, leading to a number of testable hypothesis towards delaying progression. 2. A number of disease milestones have been identified as therapeutic targets, such as conversion from MCI to diagnosable dementia. 3. Clinicians caring for patients with AD have currently available a number of symptomatic drugs, and will have in the future the ability to predict the risk for asymptomatic individuals to develop AD, and provide advice towards prevention. PMID- 11263761 TI - Brain structures and neurotransmitters regulating aggression in cats: implications for human aggression. AB - 1. Violence and aggression are major public health problems. 2. The authors have used techniques of electrical brain stimulation, anatomical-immunohistochemical techniques, and behavioral pharmacology to investigate the neural systems and circuits underlying aggressive behavior in the cat. 3. The medial hypothalamus and midbrain periaqueductal gray are the most important structures mediating defensive rage behavior, and the perifornical lateral hypothalamus clearly mediates predatory attack behavior. The hippocampus, amygdala, bed nucleus of the stria terminalis, septal area, cingulate gyrus, and prefrontal cortex project to these structures directly or indirectly and thus can modulate the intensity of attack and rage. 4. Evidence suggests that several neurotransmitters facilitate defensive rage within the PAG and medial hypothalamus, including glutamate, Substance P, and cholecystokinin, and that opioid peptides suppress it; these effects usually depend on the subtype of receptor that is activated. 5. A key recent discovery was a GABAergic projection that may underlie the often-observed reciprocally inhibitory relationship between these two forms of aggression. 6. Recently, Substance P has come under scrutiny as a possible key neurotransmitter involved in defensive rage, and the mechanism by which it plays a role in aggression and rage is under investigation. 7. It is hoped that this line of research will provide a better understanding of the neural mechanisms and substrates regulating aggression and rage and thus establish a rational basis for treatment of disorders associated with these forms of aggression. PMID- 11263762 TI - Involvement of the hypothalamic--pituitary--adrenal/gonadal axis and the peripheral nervous system in rheumatoid arthritis: viewpoint based on a systemic pathogenetic role. AB - From the compendium presented above, the following statements become evident: 1) Inappropriately low secretion of cortisol in relation to inflammation is a typical feature of the inflammatory disease in patients with RA. 2) The secretion of adrenal androgens is significantly reduced, which is a problem in postmenopausal women and elderly men due to a lack of downstream sex hormones. 3) Serum levels of testosterone are markedly reduced in RA. 4) Sympathetic nerve fibers are markedly reduced in the synovial tissue of patients with RA, whereas proinflammatory sensory fibers (substance P) are present. 5) Substance P serves to continuously sense painful stimuli in the periphery, and the nociceptive input from the inflamed joint shows a large amplification in the spinal cord. This leads to continuous pain with stabilization of the afferent sensory input and continuous release of proinflammatory substance P into the lumen of the joint. From these facts it is obvious that alterations of the systemic antiinflammatory feedback systems contribute significantly to the pathogenesis of RA. Disease therapy directed at these alterations must provide a mechanism to replace the adrenal glands (glucocorticoids), the gonadal glands (androgens), and the sympathetic nervous system (adenosine increase by low-dose MTX, sulfasalazine, and salicylates) in order to integrate their immunosuppressive effects at the local site of synovial inflammation. Although local processes of the adaptive immune system are important in pathogenesis in the acute phase of RA, these mechanisms may be less important during the chronic phase of the disease in the absence of a specific trigger. We believe that a defect of systemic antiinflammatory feedback systems is an important factor in the perpetuation of RA. This review reinforces the belief that combined therapeutic approaches on a neuroendocrine immune basis are of crucial importance in a pathogenetically oriented therapy of RA. PMID- 11263763 TI - Treatment of polyarteritis nodosa, microscopic polyangiitis, and Churg-Strauss syndrome: where do we stand? PMID- 11263764 TI - A randomized two-year study of the effects of dynamic strength training on muscle strength, disease activity, functional capacity, and bone mineral density in early rheumatoid arthritis. AB - OBJECTIVE: To evaluate the impact of a 2-year program of strength training on muscle strength, bone mineral density (BMD), physical function, joint damage, and disease activity in patients with recent-onset (<2 years) rheumatoid arthritis (RA). METHODS: In this prospective trial, 70 RA patients were randomly assigned to perform either strength training (all major muscle groups of the lower and upper extremities and trunk, with loads of 50-70% of repetition maximum) or range of motion exercises (without resistance) twice a week; all were encouraged to engage in recreational activities 2-3 times a week. All patients completed training diaries (evaluated bi-monthly) and were examined at 6-month intervals. All were treated with medications to achieve disease remission. Maximum strength of the knee extensors, trunk flexors and extensors, and grip strength was measured with dynamometers. BMD was measured at the femoral neck and lumbar spine by dual x-ray densitometry. Disease activity was determined by the Disease Activity Score, the extent of joint damage by the Larsen score, and functional capacity by the Health Assessment Questionnaire (HAQ); walking speed was also measured. RESULTS: Sixty-two patients (31 per group) completed the study. Strength training compliance averaged 1.4-1.5 times/week. The maximum strength of all muscle groups examined increased significantly (19-59%) in the strength training group, with statistically significant improvements in clinical disease activity parameters, HAQ scores, and walking speed. While muscle strength, disease activity parameters, and physical function also improved significantly in the control group, the changes were not as great as those in the strength training group. BMD in the femoral neck and spine increased by a mean +/- SD of 0.51 +/- 1.64% and by 1.17 +/- 5.34%, respectively, in the strength-training group, but decreased by 0.70 +/- 2.25% and 0.91 +/- 4.07% in the controls. Femoral neck BMD in the 17 patients with high initial disease activity (and subsequent use of oral glucocorticoids) remained constantly at a statistically significantly lower level than that in the other 45 patients. CONCLUSION: Regular dynamic strength training combined with endurance-type physical activities improves muscle strength and physical function, but not BMD, in patients with early RA, without detrimental effects on disease activity. PMID- 11263765 TI - Increased mortality in adults with a history of juvenile rheumatoid arthritis: a population-based study. AB - OBJECTIVE: To assess mortality in a population-based cohort of adults with a history of juvenile rheumatoid arthritis (JRA). METHODS: The Rochester Epidemiology Project database was used to identify all cases of JRA diagnosed among Rochester, Minnesota residents under the age of 16 between January 1, 1960 and December 31, 1993. Fifty-seven patients in this cohort are now adults (ages 18-53 years, mean age 34.3 years), and this subgroup was contacted for a long term followup study. The average length of followup from the time of diagnosis was 25.6 years. RESULTS: Four deaths occurred in this cohort of 57 adults with a history of JRA. All 4 deceased patients had other autoimmune illnesses and died of complications of these diseases. The observed frequency of 4 deaths was significantly greater (P < 0.0026 by one-sample log-rank test) than the 1 death that would be expected among Minnesota whites of similar age and sex, and corresponds to a mortality rate of 0.27 deaths per 100 years of patient followup compared with an expected mortality rate of 0.068 deaths per 100 years of followup in the general population. CONCLUSION: The results indicate a significant, unexpected increase in mortality in this population-based cohort of adults with a history of JRA in comparison with the rate in the general population. The deaths in this group were all associated with other autoimmune disorders, suggesting that special emphasis should be given to the diagnosis and treatment of other autoimmune diseases, including immunodeficiencies, in JRA patients. The frequency of deaths in this cohort suggests that JRA patients are at substantial risk for mortality, and highlights the need for longitudinal followup and care into adulthood. PMID- 11263766 TI - Indirect medical costs in early rheumatoid arthritis: composition of and changes in indirect costs within the first three years of disease. AB - OBJECTIVE: To investigate 1) the magnitude of indirect costs, 2) changes in cost components, and 3) correlations between changes in cost and social, clinical, and occupational variables within the first 3 years of rheumatoid arthritis (RA). METHODS: We evaluated the indirect costs per person-year in 133 consecutive gainfully employed out-patients with early RA, in a prospective multicenter followup study. Costs due to RA-related sick leave, work disability, and other work loss were assessed using the human capital approach. Variables associated with reduction in lost productivity were tested by multivariate logistic regression analysis. RESULTS: Mean +/- SEM annual indirect costs were $11,750 +/- 1,120 per person. During the 3-year period of observation, a marked reduction in the costs associated with sick leave was seen, which exceeded the increase in costs due to work disability and other work loss. This phenomenon resulted in an overall reduction in indirect costs of 21%. The final logistic regression model of reduced loss of productivity included 3 variables: no problems with standing (odds ratio [OR] 7.1), no problems with working speed (OR 4.1), and no problems with outdoor work (OR 3.1). CONCLUSION: High indirect costs in early RA were demonstrated. An overall decrease of costs can be seen in the first 3 years, due to the reduction in sick leave. Since the absence of problems due to strenuous working conditions was found to be associated with a reduction in indirect costs, it is assumed that early intensified vocational rehabilitation, apart from controlling disease activity by adequate treatment, might help to reduce indirect costs. PMID- 11263767 TI - Influence of shared epitope-negative HLA-DRB1 alleles on genetic susceptibility to rheumatoid arthritis. AB - OBJECTIVE: Most patients with rheumatoid arthritis (RA) express the shared epitope (SE). It is not known whether SE-negative HLA-DRB1 alleles influence the development of RA. This study examined the influence of SE-negative HLA-DR alleles (DRB1*X) on the development of RA in 3 different French populations. METHODS: HLA-DRB1 alleles were defined by polymerase chain reaction with sequence specific oligonucleotide hybridization or sequence-specific primers. SE-negative alleles were classified according to the electric charge of their P4 pocket. HLA DRB1 alleles *0103, *0402, *07, *08, *11 (except *1107), *12, and *13 have a neutral or negative P4 charge and are called DRB1*XP4n. HLA-DRB1*03, *0403, *0406, *0407, *0901, *1107, *14, *15, and *16 have a positive P4 charge and are called DRB1*XP4p. RESULTS: Among the SE-negative subjects, DRB1 genotypes with 1 or 2 DRB1*XP4n alleles were significantly overrepresented in the control subjects compared with the RA patients, whereas DRB1*XP4p/XP4p genotypes were equally represented in the patients and controls. In single-dose SE-positive subjects, SE/XP4n genotypes were equally represented in the patients and controls. However, SE/XP4p genotypes were significantly overrepresented in the RA patients. CONCLUSION: The DRB1*X allele polymorphism influences susceptibility to RA. Alleles that have a neutral or negative electric charge in their P4 pocket (DRB1*XP4n), such as DRB1*0103, *0402, *07, *08, *11 (except *1107), *12, and *13, protect against RA. Alleles that have a positive electric charge in their P4 pocket (DRB1*XP4p), such as DRB1*03, *0403, *0406, *0407, *0901, *1107, *14, *15, and *16, have no influence on the predisposition to RA. PMID- 11263768 TI - Lymph draining from foot joints in rheumatoid arthritis provides insight into local cytokine and chemokine production and transport to lymph nodes. AB - OBJECTIVE: Rheumatoid arthritis (RA) is characterized by inflammatory reactions in joints and adjacent tissues unaccompanied by clinically evident changes in lymphatics and lymph nodes draining the inflamed areas. The explanation for this phenomenon, which contrasts with infectious processes in joints and soft tissues that evoke major changes in the lymphatic system, is unclear. To determine which inflammatory factors produced in the joints of RA patients are transported in lymph to lymph nodes, we measured levels of immunoglobulins, cytokines, and chemokines in prenodal lymph from the foot joints of RA patients and quantified their rate of transport to regional lymph nodes. METHODS: Lymph was collected from the cannulated lymphatics draining the foot joints, tendons, fascia, and skin of 20 RA patients. Lymph flow rate and concentrations of proteins and immunoglobulins were measured. Cytokine and chemokine levels were quantified by enzyme-linked immunosorbent assays. Results were compared with those obtained in 20 control subjects. RESULTS: In the cannulated vessel, the mean +/- SEM lymph flow rate in RA patients was almost 2-fold that in control subjects (22.6 +/- 3.2 ml/24 hours versus 13.2 +/- 1.1 ml/24 hours; P < 0.01). Lymph concentrations of total protein, IgG, and IgM were 1.80 +/- 0.14 gm/dl, 384 +/- 45 mg/dl, and 32.0 +/- 1.5 mg/dl, respectively, in RA patients and 1.66 +/- 0.14 gm/dl, 238 +/- 32 mg/dl, and 15.0 +/- 1.3 mg/dl, respectively, in control subjects. The corresponding lymph:serum (L:S) ratios were 0.21 +/- 0.02, 0.22 +/- 0.02, and 0.15 +/- 0.02, respectively, in RA patients and 0.22 +/- 0.02, 0.19 +/- 0.02, and 0.11 +/- 0.02, respectively, in control subjects. The L:S ratios of <1 and the absence of significant differences between groups suggested a lack of local production of immunoglobulins. In RA patients, lymph concentrations (in pg/ml) were as follows: interleukin-1beta (IL-1beta) 14.8 +/- 3.9, IL-6 511 +/- 143, tumor necrosis factor alpha (TNFalpha) 9.9 +/- 1.1, IL-1 receptor antagonist (IL 1Ra) 4,274 +/- 737, IL-10 13.3 +/- 4.4, IL-8 846 +/- 174, IL-15 6.2 +/- 0.9, granulocyte-macrophage colony-stimulating factor (GM-CSF) 2.30 +/- 0.15, vascular endothelial growth factor (VEGF) 80.4 +/- 8.6, and macrophage inflammatory protein 1alpha (MIP-1alpha) 171 +/- 34. In control subjects, these values were as follows: IL-1beta 1.50 +/- 0.25, IL-6 79.0 +/- 14.6, TNFalpha 4.4 +/- 1.1, IL-1Ra 208 +/- 52, IL-10 0.0, IL-8 216 +/- 83, IL-15 5.00 +/- 0.45, GM-CSF 0.40 +/- 0.05, VEGF 42.0 +/- 2.4, and MIP-1alpha 3.4 +/- 1.7 (P < 0.05 versus RA patients for all except IL-15). The L:S ratio was >1 in all RA patient samples for IL 1beta, IL-6, IL-1Ra, IL-8, GM-CSF, IL-10, IL-15, TNFalpha, and MIP-1alpha, indicating local production of cytokines. Great variability in lymph cytokine concentrations, presumably reflecting differences in the intensity of local inflammation, was not reflected in serum cytokine concentrations. Intravenously infused methylprednisolone decreased lymph cytokine levels to normal within 12 hours. In contrast, their concentrations in serum showed little or no change. CONCLUSION: High lymph concentrations of cyto kines and chemokines, exceeding those in serum, were found in RA patients. The L:S concentration ratios of > 1 indicate the local production of these cytokines and chemokines in the inflamed tissues. High flow rates of lymph containing high cytokine concentrations through the regional lymph nodes are likely to affect node lymphocytes and dendritic cells. Analysis of cytokines in lymph should provide insight into events in inflamed tissues in RA and in regional lymph nodes. PMID- 11263769 TI - Levels of interleukin-18 and its binding inhibitors in the blood circulation of patients with adult-onset Still's disease. AB - OBJECTIVE: Interleukin-18 (IL-18) is a proinflammatory cytokine that is involved in immunologically mediated tissue damage, but its bioactivity is regulated in vivo by its soluble decoy receptor, IL-18 binding protein (IL-18BP). This study was undertaken to determine levels of IL-18 and IL-18 binding inhibition in the blood of patients with adult-onset Still's disease (ASD). METHODS: Serum concentrations of IL-18 in ASD patients were compared by enzyme-linked immunosorbent assay (ELISA) with those in patients with other systemic rheumatic diseases and healthy controls. The biologically active mature protein of IL-18 was detected by Western blot analysis with anti-IL-18 antibody and its induction of interferon-gamma (IFNgamma) secretion from IL-18-responding human myelomonocytic KG-1 cells. The inhibitory activity on IL-18 binding to its receptor was determined by 125I-IL-18 binding inhibition assay using the Chinese hamster ovary cell line transfected with a murine IL-18 receptor (CHO-K1/mIL 18R). RESULTS: Concentrations of serum IL-18 were extremely elevated in patients with active ASD compared with those in patients with rheumatoid arthritis, systemic lupus erythematosus, systemic sclerosis, polymyositis/dermatomyositis, Sjogren's syndrome, or healthy individuals. Levels of IL-18 were found to correlate with serum ferritin values and disease severity in ASD. Western blot analysis revealed that serum samples from patients with active ASD contained an 18-kd polypeptide of IL-18, corresponding in size to the mature form. Accordingly, the samples were able to induce IFNgamma secretion from KG-1 cells, which was largely abolished by neutralizing anti-IL-18 antibody. However, the level of IL-18 bioactivity was more than 10-fold weaker than the concentration of IL-18 protein measured by ELISA. Serum samples from patients with active ASD showed an inhibitory effect on the binding of 125I-IL-18 to CHO-K1/mIL-18R cells, and this activity was associated with elevation of IL-18. CONCLUSION: These data indicate that systemic overproduction of IL-18 may be closely related to the pathogenesis of ASD, despite the restriction on its inflammatory activity by IL 18 binding inhibitors such as IL-18BP. The disease activity appears to be determined on the basis of the relative levels of IL-18 and its specific inhibitors. PMID- 11263770 TI - Shift toward T helper 1 cytokines by type II collagen-reactive T cells in patients with rheumatoid arthritis. AB - OBJECTIVE: To investigate the impact of type II collagen (CII)-reactive T cells on the Th1/Th2 cytokine balance in patients with rheumatoid arthritis (RA). METHODS: T cell proliferative responses to bovine CII were examined in synovial fluid mononuclear cells (SFMC) and peripheral blood mononuclear cells (PBMC) by mixed lymphocyte culture. CII-reactive T cell lines were generated from the SFMC and PBMC. Interferon-gamma (IFNgamma), interleukin-12 (IL-12), and IL-4 were measured by enzyme-linked immunosorbent assay in the SF, sera, and culture supernatants of PBMC and SFMC that had been stimulated with CII. RESULTS: The frequency of CII-reactive T cells was higher in the PBMC from RA patients than in that from osteoarthritis patients and healthy control subjects. In RA patients, CII-reactive T cells were more prevalent in SFMC than in PBMC. The mean level of IFNgamma and the ratio of IFNgamma to IL-4 were significantly higher in the culture supernatants of T cells stimulated with CII; these differences were more prominent in SFMC. Levels of IL-12 in the culture supernatants of SFMC and PBMC stimulated with CII were significantly higher than those in unstimulated supernatants. T cell responsiveness correlated well with the level of type 1 cytokines in culture supernatants from RA T cells stimulated with CII. In the CII reactive cell lines, the increased production of IFNgamma was consistent with clonal expansion. CONCLUSION: CII-reactive T cells are more abundant in SFMC than in PBMC and are strongly associated with a shift toward Thl cytokine in the inflamed joints of RA patients. Our results suggest that a skewing toward type 1 cytokines by CII-reactive T cells may play an important role in the chronic inflammatory process of RA. PMID- 11263771 TI - Infection efficiency of type 5 adenoviral vectors in synovial tissue can be enhanced with a type 16 fiber. AB - OBJECTIVE: To obtain an adenoviral vector with increased infection efficiency in the synovial tissue compared with conventional vectors based on adenovirus serotype 5 (Ad5), without compromising the specificity of infection. METHODS: Coxsackie adenovirus receptor (CAR) expression was assessed in cultured synoviocytes. Chimeric adenoviruses based on Ad5 but carrying the DNA encoding the fiber of adenovirus from subgroup B (Adll, 16, 35) or D (Ad24, 28, 33, 45, or 47) were constructed and produced on PER.C6 cells. The gene transfer efficiency of these chimera was tested on cultured synoviocytes and peripheral blood mononuclear cells (PBMC). RESULTS: No surface expression of CAR protein was observed on synoviocytes. CAR messenger RNA expression of synoviocytes was found to be low. Of all fiber chimeric vectors tested, vectors carrying the fiber of Ad16 (Ad5.fib16) were most potent, yielding approximately150 times increased transgene expression in cultured synoviocytes compared with those of Ad5. Flow cytometry showed that the increase in transgene expression was caused by the transduction of higher percentages of synoviocytes and higher gene expression per synoviocyte. Experiments with 500 virus particles/cell of Ad5.GFP or Ad5.fib16.GFP resulted in an infection efficiency of 0.6% and 1% in PBMC and 43% and 76% in synoviocytes, respectively. CONCLUSION: Synoviocytes hardly express CAR, which hampers Ad5-mediated gene transfer. Ad5.fib16 is superior to Ad5 vectors for transducing synoviocytes, without compromising the specificity of infection. Our data suggest that Ad5.fib16-mediated gene transfer to synovial tissue improves the therapeutic window. PMID- 11263772 TI - Osteopontin: an intrinsic inhibitor of inflammation in cartilage. AB - OBJECTIVE: To identify extracellular and intraarticular matrix components that are differentially expressed in normal and osteoarthritis (OA)-affected cartilage and to investigate their functions with respect to regulation of mediators of inflammation. METHODS: Differential-display reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of a pool of messenger RNA (mRNA) from 10 human OA cartilage samples and 5 normal cartilage samples was performed using arbitrary primers. Confirmatory analysis of the up-regulated transcripts of fibronectin (FN) and osteopontin (OPN) was performed by RT-PCR of individual RNA samples from a separate set of donors. The effect of recombinant OPN (or anti-OPN antiserum) on chondrocyte function was examined by analyzing the spontaneous or interleukin 1 (IL-1)-induced release of nitric oxide (NO) and prostaglandin E2 (PGE2) from human OA-affected cartilage under ex vivo conditions. RESULTS: Up-regulation (300 700%) of FN and OPN mRNA was observed in human OA-affected cartilage as compared with normal cartilage. Functional analysis of the role of OPN in OA cartilage showed that 1) Addition of 1 microg/ml (20 nM) of recombinant OPN to human OA affected cartilage under ex vivo conditions inhibited spontaneous and IL-1beta induced NO and PGE2 production, and 2) neutralization of intraarticular OPN with anti-OPN antiserum augmented NO production. CONCLUSION: The data indicate that one of the functions of intraarticular OPN, which is overexpressed in OA cartilage, is to act as an innate inhibitor of IL-1, NO, and PGE2 production. These findings suggest that the production of pleiotropic mediators of inflammation that influence cartilage homeostasis, such as NO and PGE2, is regulated by the interaction of chondrocytes with differentially expressed proteins within the extracellular matrix. PMID- 11263773 TI - Matrix metalloproteinase and proinflammatory cytokine production by chondrocytes of human osteoarthritic cartilage: associations with degenerative changes. AB - OBJECTIVE: To examine by immunohistochemistry the relative distributions of 6 matrix metalloproteinases (MMPs 1, 2, 3, 8, 9, and 13) and the 2 proinflammatory cytokines interleukin-1beta (IL-1beta) and tumor necrosis factor alpha (TNFalpha) in osteoarthritic (OA) cartilage compared with normal, age-matched articular cartilage. METHODS: Articular cartilage samples were obtained from the tibial plateau of OA knees removed at arthroplasty and from normal, nonarthritic, knees obtained at autopsy. Specimens were promptly fixed in Carnoy's fixative, processed, embedded in paraffin, sectioned, and examined by immunohistochemistry for MMP and cytokine production. In addition, human articular chondrocytes (HAC) were treated in vitro with either IL-1beta, TNFalpha, or phorbol myristate acetate (PMA) to assess their potential to produce each of the MMPs, as determined by Western blotting and gelatin zymography. RESULTS: Immunodetection of the collagenases (MMPs 1, 8, and 13) and stromelysin 1 (MMP-3) was demonstrated in a proportion of chondrocytes in the superficial zone of almost all of the OA specimens that had degenerative matrix changes. The gelatinases (MMPs 2 and 9) were also demonstrated by immunohistochemistry but were not so prominent. IL-1beta- and TNFalpha-positive chondrocytes were also observed in a proportion of cells in the superficial zones of OA specimens. Much less immunostaining for MMPs and cytokines was observed in the deep zone of all OA specimens, where the cartilage matrix and chondrocyte morphology appeared normal. In contrast, full-thickness normal cartilage specimens showed virtually no immunostaining for these MMPs or cytokines. Confirmation that chondrocytes can produce these 6 MMPs was obtained from HAC cultures treated with either IL-1beta, TNFalpha, or PMA; conditioned medium from activated HAC contained all the MMPs demonstrated by immunohistochemistry. Dual immunolocalization studies of OA cartilage specimens demonstrated the coexpression of IL-1 with MMP-8 by individual chondrocytes in situ. CONCLUSION: These results indicate that the superficial zone of OA cartilage specimens, which is characterized by fibrillations, chondrocyte clusters, and degenerative matrix changes, contains a variable proportion of cells that immunostain for IL-1beta, TNFalpha, and 6 different MMPs. These observations support the concept that cytokine-MMP associations reflect a modified chondrocyte phenotype and an intrinsic process of cartilage degradation in OA. PMID- 11263774 TI - Peroxisome proliferator--activated receptor gamma activators inhibit interleukin 1beta-induced nitric oxide and matrix metalloproteinase 13 production in human chondrocytes. AB - OBJECTIVE: To determine the effects of peroxisome proliferator-activated receptor gamma (PPARgamma) agonists on interleukin-1 (IL-1) induction of nitric oxide (NO) and matrix metalloproteinase 13 (MMP-13) in human chondrocytes. METHODS: PPARgamma expression and synthesis in human chondrocytes were determined by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively. Chondrocytes were cultured with IL-1beta, tumor necrosis factor alpha (TNFalpha), and IL-17 in the presence or absence of PPARgamma agonists, and NO and MMP-13 synthesis and expression levels were measured. Transient transfection experiments were performed with the 7-kb inducible NO synthase (iNOS) and 1.6-kb MMP-13 human promoters, as well as with the PPARgamma expression vector and the activator protein 1 (AP-1) and nuclear factor kappaB (NF-kappaB) reporter constructs. RESULTS: RT-PCR and immunohistochemical analysis revealed that human chondrocytes expressed and produced PPARgamma. Treatment of chondrocytes with PPARgamma ligands BRL 49653 and 15-deoxy-delta12,14-prostaglandin J2 (15d-PGJ2), but not with PPARalpha ligand Wy 14643, decreased IL-1beta-induced NO and MMP-13 production in a dose dependent manner. In addition, both iNOS and MMP-13 messenger RNA were inhibited in the presence of 15d-PGJ2. The inhibitory effect of PPARgamma activation was not restricted to IL-1beta, since TNFalpha- and IL-17-induced NO and MMP-13 production were also inhibited by 15d-PGJ2. In transient transfection experiments, we showed that a constitutively active form of mitogen-activated protein kinase kinase kinase 1 (AMEKK-1) induced the MMP-13 and iNOS human promoter activity. This process was reduced by 15d-PGJ2 and further inhibited by cotransfection with a PPARgamma expression vector. Similarly, in a PPARgamma dependent manner, 15d-PGJ2 inhibited deltaMEKK-1-induced AP-1- and NF-kappaB luciferase reporter plasmid activation. CONCLUSION: The findings of this study demonstrate that PPARgamma agonists inhibit IL-1beta induction of both NO and MMP 13 in human chondrocytes. The inhibition occurs at least at the transcriptional level through a PPARgamma-dependent pathway, probably by interfering with the activation of AP-1 and NF-kappaB. PMID- 11263777 TI - Immunoglobulin Vkappa light chain gene analysis in patients with Sjogren's syndrome. AB - OBJECTIVE: Patients with Sjogren's syndrome (SS) have characteristic lymphocytic infiltration of the salivary glands with a previously reported predominance of Vkappa-bearing B cells and produce a variety of autoantibodies, indicating that there is a humoral autoimmune component in this syndrome. This study was undertaken to determine whether there are primary deviations of immunoglobulin V gene usage, differences in somatic hypermutation, defects of selection, or indications for perturbances of B cell maturation in SS. METHODS: Individual peripheral B cells from patients with SS were analyzed for their Ig V gene usage, and the findings were compared with results in normal controls. RESULTS: Molecular differences, as reflected by findings in the nonproductive Vkappa repertoire of the patients, were identified by an enhanced usage of Jkappa2 gene segments and a lack of mutational targeting toward RGYW/WRCY sequences compared with controls. A greater usage of Vkappa1 family members and a reduced frequency of Vkappa3 gene segments in the productive repertoire suggested differences in selection, possibly driven by antigen. Overall positive selection for mutations, especially for replacements in the complementarity-determining region and for mutations in RGYW/WRCY, similar to that found in controls, was detected. CONCLUSION: Disturbances of strictly regulated B cell maturation, during early B cell development as indicated by prominent Jkappa2 gene usage and during germinal center reactions as indicated by a lack of targeting of the hypermutation mechanism, might contribute to the emergence of autoimmunity in SS. PMID- 11263776 TI - The Fcgamma receptor IIIA-158F allele is a major risk factor for the development of lupus nephritis among Caucasians but not non-Caucasians. AB - OBJECTIVE: To determine whether inheritance of Fcgamma receptor (FcgammaR) alleles conferring lower affinity for IgG binding increases the risk of developing lupus nephritis. METHODS: We compared the frequency of low-affinity alleles of two FcgammaR polymorphisms (FcgammaRIIA and FcgammaRIIIA) among 235 patients with systemic lupus erythematosus (SLE) and proven nephritis (nephritis patients) and among 352 SLE patients with no evidence of renal disease (non nephritis control subjects). The ethnic distribution of patients in the study was 49% Caucasian, 20% Hispanic, 17% Asian/Pacific Islander, 12% African American, and 2% from other ethnic groups. All patients were genotyped for the FcgammaRIIA 131R/H and FcgammaRIIIA-158V/F polymorphisms. We used contingency table analysis to compare allele and genotype distributions for nephritis patients and nonnephritis control subjects, including ethnic-specific strata. Multivariate logistic regression analyses included sex and disease duration as covariates. RESULTS: Univariate and multivariate analyses demonstrated a striking association between the low-affinity FcgammaRIIIA-158F allele and FF genotype and the risk of nephritis among Caucasians, but not among non-Caucasians (multivariate odds ratio [OR] 2.6 for Caucasians with FF genotype), (P = 0.0017). This association was even stronger among Caucasians with severe nephritis (OR 4.4, P < 0.0001). In contrast, inheritance of the low-affinity FcgammaRIIA-131R allele (and RR genotype) was not associated with an increased risk of lupus nephritis among any of the ethnic groups examined. CONCLUSION: The FcgammaRIIIA-158F allele is a major risk factor for the development of lupus nephritis among Caucasians, but not among non-Caucasians. These results suggest that ethnic variation is critical in defining the specific genetic factors underlying the pathogenesis of SLE, and they have important prognostic and therapeutic implications as well. PMID- 11263775 TI - Cyclic tensile strain suppresses catabolic effects of interleukin-1beta in fibrochondrocytes from the temporomandibular joint. AB - OBJECTIVE: To discern the effects of continuous passive motion on inflamed temporomandibular joints (TMJ). METHODS: The effects of continuous passive motion on TMJ were simulated by exposing primary cultures of rabbit TMJ fibrochondrocyte monolayers to cyclic tensile strain (CTS) in the presence of recombinant human interleukin-1beta (rHuIL-1beta) in vitro. The messenger RNA (mRNA) induction of rHuIL-1beta response elements was examined by semiquantitative reverse transcriptase-polymerase chain reaction. The synthesis of nitric oxide was examined by Griess reaction, and the synthesis of prostaglandin E2 (PGE2) was examined by radioimmunoassay. The synthesis of proteins was examined by Western blot analysis of the cell extracts, and synthesis of proteoglycans via incorporation of 35S-sodium sulfate in the culture medium. RESULTS: Exposure of TMJ fibrochondrocytes to rHuIL-1beta resulted in the induction of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2), which were paralleled by NO and PGE2 production. Additionally, IL-1beta induced significant levels of collagenase (matrix metalloproteinase 1 [MMP-1]) within 4 hours, and this was sustained over a period of 48 hours. Concomitant application of CTS abrogated the catabolic effects of IL-1beta on TMJ chondrocytes by inhibiting iNOS, COX-2, and MMP-1 mRNA production and NO, PGE2, and MMP-1 synthesis. CTS also counteracted cartilage degradation by augmenting expression of mRNA for tissue inhibitor of metalloproteinases 2 that is inhibited by rHuIL-1beta. In parallel, CTS also counteracted rHuIL-1beta-induced suppression of proteoglycan synthesis. Nevertheless, the presence of an inflammatory signal was a prerequisite for the observed CTS actions, because fibrochondrocytes, when exposed to CTS alone, did not exhibit any of the effects described above. CONCLUSION: CTS acts as an effective antagonist of rHuIL-1beta by potentially diminishing its catabolic actions on TMJ fibrochondrocytes. Furthermore, CTS actions appear to involve disruption/regulation of signal transduction cascade of rHuIL-1beta upstream of mRNA transcription. PMID- 11263778 TI - Polymorphisms of the Ro52 gene associated with anti-Ro 52-kd autoantibodies in patients with primary Sjogren's syndrome. AB - OBJECTIVE: To screen for the Ro52 gene encoding the 52-kd Ro autoantigen for possible mutations and polymorphisms associated with primary Sjogren's syndrome (SS). METHODS: The restriction enzyme fragment-single-strand conformation polymorphism method was used to search for mutations and polymorphisms in the Ro52 gene in 97 patients with primary SS and 72 healthy control subjects. The results were verified by automated DNA sequencing and natural or amplification created restriction site tests. RESULTS: A single-nucleotide polymorphism (SNP) was discovered in intron 3 (137 bp upstream of exon 4). The C/T genotype was significantly more prevalent among patients who were positive for anti-Ro 52-kd (20 of 38) than among healthy controls (9 of 72) (P = 0.00003); significant differences were not seen in patients who were negative for anti-Ro 52-kd. Furthermore, the frequency of the T allele in this position among groups of anti Ro 52-kd-positive patients, anti-Ro 52-kd-negative patients, and control subjects was significantly increased in the patients who were positive for anti-Ro 52-kd compared with the controls. CONCLUSION: We present the results of a complete screening for the Ro52 gene in patients with primary SS and the results of an association study. An SNP in intron 3 was found to be strongly associated with the presence of anti-Ro 52-kd autoantibodies in primary SS. This finding is interesting in light of the fact that an alternative messenger RNA is made by deleting exon 4, which encodes a putative leucine zipper domain, to generate a shorter version of the Ro 52-kd protein. PMID- 11263779 TI - Long-term outcome of mothers of children with isolated heart block in Finland. AB - OBJECTIVE: To study the long-term outcome of mothers of children with isolated heart block in a defined population. METHODS: We reviewed the Finnish hospital registries for patients born between 1950 and 1999 who had been diagnosed as having isolated heart block before the age of 15 years. We identified 101 children with isolated congenital heart block (CHB) and 55 with isolated heart block detected after the newborn period. Eighty-three (91%) of the 91 mothers of children with CHB and 48 (87%) of the 55 mothers of children with heart block detected after the newborn period were studied according to a protocol defining clinical characteristics (mean 9.9 years, range 0-49 years, and mean 22.9 years, range 4-47 years after the index delivery, respectively). Maternal survival was compared with survival in an age-matched population of normal Finnish women. RESULTS: Before the index delivery, 29 (37%) of the 78 surviving mothers of children with CHB had a self-reported clinical diagnosis of a chronic autoimmune disease, and 55 (71%) had had symptoms, signs, or abnormal laboratory findings suggesting an underlying subclinical disease. Of the 23 mothers who were completely asymptomatic before the index delivery, 10 (13% of the surviving mothers) remained so after a mean followup of 9.6 years (range 0-21 years). In mothers of children with CHB, clinical characteristics different from those of healthy mothers were photosensitivity, fatigue, dry eyes, and dry mouth. Forty eight (58%) of these 83 mothers developed an autoimmune disease during followup. The most common diagnosis was primary Sjogren's syndrome (22 definite, 11 probable), followed by systemic lupus erythematosus (SLE). The standardized mortality ratio of mothers of children with CHB was 5.1, and 3 of the 5 deaths were associated with SLE. Mothers of children with heart block detected after the newborn period had similar symptoms and signs of autoimmune diseases as the healthy controls, and their standardized mortality ratio was 1.9. CONCLUSION: Primary Sjogren's syndrome, either definite or subclinical, is the predominant autoimmune disorder in mothers of children with CHB. Mothers of children with isolated heart block detected after the newborn period do not, as a group, have clinical features suggestive of autoimmune diseases. PMID- 11263780 TI - The Disability Index of the Health Assessment Questionnaire is a predictor and correlate of outcome in the high-dose versus low-dose penicillamine in systemic sclerosis trial. AB - OBJECTIVE: To explore the clinical implications of a score of > or =1.0 on the Disability Index of the Health Assessment Questionnaire (HAQ DI) at the first patient visit, and to examine the implications of improvement in HAQ DI score over 2 years in a cohort of systemic sclerosis (SSc) patients with diffuse cutaneous scleroderma. METHODS: SSc skin and visceral involvement was assessed in 134 SSc patients with diffuse scleroderma (mean +/- SD disease duration of 10 +/- 4 months) when they entered a multicenter drug trial and again 2 years later. Mortality and the occurrence of scleroderma renal crisis were assessed for a mean +/- SD of 4.0 +/- 1.1 years. Logistic and linear regression analyses were used to examine the relationship of the baseline HAQ DI score to morbidity, mortality, and visceral involvement, as well as the relationship of changes in the HAQ DI score to changes in physical examination, laboratory, and functional variables over 2 years. RESULTS: A baseline HAQ DI score of > or =1.0 was predictive of mortality (odds ratio 3.22, 95% confidence interval 1.097-9.468) over 4 years. Multivariate linear regression demonstrated that a model which included the erythrocyte sedimentation rate at baseline (P = 0.005) and changes at 2 years in the swollen joint count (P = 0.002), total skin score (P = 0.005), and white blood cell count (P = 0.005) best explained the change in HAQ DI score over 2 years (R2 = 0.528). The HAQ DI score and total skin score at baseline were highly correlated (correlation coefficient 0.368), as were changes in the HAQ DI score and the total skin score over 2 years (correlation coefficient 0.492). Although the HAQ DI score was heavily influenced by hand dysfunction at baseline and at 2 years, improvement (reduction) in the HAQ DI score over 2 years was related to factors other than hand dysfunction. CONCLUSION: A baseline HAQ DI score of > or =1.0 predicted mortality over 4 years. Improvement in the HAQ DI score in these patients with diffuse scleroderma was associated with improvement in skin thickening, hand function, oral aperture, lung function, signs of arthritis, serum creatinine level, and the investigator's global assessment of improvement. The HAQ DI is a self-administered questionnaire that SSc patients can complete easily and rapidly and that gives the practicing physician important information about prognosis, patient status, and changes in disease course over time. PMID- 11263781 TI - Cytochrome P2 polymorphisms and susceptibility to scleroderma following exposure to organic solvents. AB - OBJECTIVE: To determine whether there are specific cytochrome P450 (CYP2) alleles that increase susceptibility to scleroderma in individuals who have been exposed to organic solvents. METHODS: CYP alleles at 2 loci, 2E1 and 2C19, were compared in 7 patients who had developed scleroderma after exposure to solvents versus 71 patients with scleroderma without solvent exposure ("sporadic" disease) and 106 population controls. RESULTS: The 2E1*3 allele was found in 2 of the 7 patients who had been exposed to organic solvents, with a greater frequency than occurred in either the disease controls or the population controls (odds ratio [95% confidence interval] 9.1 [1.5-59.1] and 10.2 [1.8-62.2], respectively). All 7 patients with solvent exposure carried the 2C19EM genotype, compared with 89% of patients with sporadic scleroderma. CONCLUSION: Our results suggest that alleles at CYP loci may be involved in increasing susceptibility to scleroderma among subjects who have been exposed to organic solvents. PMID- 11263782 TI - Long-term followup of polyarteritis nodosa, microscopic polyangiitis, and Churg Strauss syndrome: analysis of four prospective trials including 278 patients. AB - OBJECTIVE: To determine the long-term outcome of patients with polyarteritis nodosa (PAN), microscopic polyangiitis (MPA), and Churg-Strauss syndrome (CSS), to compare the long-term outcome with the overall French population, to evaluate the impact on outcome of the type of vasculitis, prognostic factors, and treatments administered at diagnosis, and to analyze treatment side effects and sequelae. METHODS: Data from PAN, MPA, and CSS patients (n = 278) who were enrolled between 1980 and 1993 were collected in 1996 and 1997 and analyzed. Two prognostic scoring systems, the Five-Factors Score (FFS) and the Birmingham Vasculitis Activity Score (BVAS), were used to evaluate all patients at the time of diagnosis. RESULTS: The mean (+/- SD) followup of the entire population was 88.3 +/- 51.9 months (range 3 days to 192 months). Of the 85 deaths recorded, at least 41 were due to progressive vasculitis or its consequences. Death rates reflected disease severity, as assessed by the FFS (P = 0.004) and the BVAS (P < 0.0002), and the 2 scores were correlated (r = 0.69). Relapses, rarer in hepatitis B virus (HBV)-related PAN (7.9%) than in MPA (34.5%) (P = 0.004), occurred in 56 patients (20.1%) and did not reflect disease severity. Survival curves were similar for the subpopulation of 215 patients with CSS, MPA, and non HBV-related PAN who were given first-line corticosteroids (CS) with or without cyclophosphamide (CYC). However, CS with CYC therapy significantly prolonged survival for patients with FFS scores > or =2 (P = 0.041). Relapse rates were similar regardless of the treatment regimen; only patients treated with CS alone had uncontrolled disease. CYC was associated with a greater frequency of side effects (P < 0.00001). CONCLUSION: Rates of mortality due to PAN (related or unrelated to HBV), MPA, and CSS reflected disease severity and were higher than the mortality rate in the general population (P < 0.0004). Rates of relapse, more common in MPA than HBV-related PAN patients, did not reflect disease severity. Survival rates were better among the more severely ill patients who had received first-line CYC. Based on these findings, we recommend that the intensity of the initial treatment be consistent with the severity of the disease. The use of the FFS and BVAS scores improved the ability to evaluate the therapeutic response. PMID- 11263783 TI - Invasiveness of synovial fibroblasts is regulated by p53 in the SCID mouse in vivo model of cartilage invasion. AB - OBJECTIVE: In vitro data suggest that the tumor suppressor p53 is critically involved in the regulation of proliferation and apoptosis in fibroblast-like synoviocytes (FLS). Based on evidence that abnormalities in p53 expression and function are found in rheumatoid arthritis (RA), we analyzed whether inhibition of p53 using gene transfer with the human papilloma virus type 18 (HPV-18) E6 protein results in an increased cellularity and invasiveness of synovial fibroblasts in vivo. METHODS: RA and normal FLS were transduced with a pLXSN based construct encoding for the HPV-18 E6 protein or with the pLXSN vector alone. After selection with G418, FLS were coimplanted with normal human cartilage under the renal capsule of SCID mice. Parental, nontransduced cells were used as additional controls. After 60 days, the implants were removed, and FLS invasion into the cartilage, perichondrocytic degradation, and cellularity were assessed. RESULTS: Nontransduced and mock-transduced RA FLS exhibited characteristic invasion into the cartilage (mean +/- SEM scores 2.2 +/- 0.3 and 2.4 +/- 0.2, respectively). Invasion was increased significantly in the E6 transduced RA FLS (mean score 3.1 +/- 0.3; P < 0.05). Inhibition of p53 also resulted in an increase in cellularity. Parental and mock-transduced normal FLS did not exhibit significant invasion (mean score 1.5 +/- 0.1 and 1.4 +/- 0.3, respectively), but transduction with E6 resulted in clear invasiveness (mean score 2.4 +/- 0.4) as well as increased cellularity. CONCLUSIONS: The data suggest that inhibition of endogenous p53 leads to increased invasiveness and cellularity of RA FLS and may also transform normal FLS to cells that display an aggressive, RA FLS-like behavior. Therefore, abnormalities such as somatic mutations in the p53 tumor suppressor may contribute to synovial hyperplasia and invasion in RA. PMID- 11263784 TI - Variations in susceptibility to proteoglycan-induced arthritis and spondylitis among C3H substrains of mice: evidence of genetically acquired resistance to autoimmune disease. AB - OBJECTIVE: To identify and screen the level of arthritis susceptibility in C3H murine strains known to be resistant to proteoglycan (aggrecan)-induced arthritis, and to measure and correlate various immunologic and inflammatory parameters with susceptibility to either arthritis or spondylitis in various C3H substrains. METHODS: Mice of 10 C3H substrains (subcolonies) were immunized with cartilage proteoglycan (aggrecan) for induction of arthritis. Animals were assessed for clinical symptoms, and the peripheral joints and spine were studied by histologic methods. Proteoglycan-specific T cell responses (T cell proliferation and production of interleukin-2 [IL-2], interferon-y, and IL-4) and the B cell response to lipopolysaccharide (LPS) were measured in spleen cell cultures. Serum levels of heteroantibodies and autoantibodies as well as various cytokines (IL-6, IL-10, IL-12, and tumor necrosis factor alpha) and soluble CD44 were determined by enzyme-linked immunosorbent assay. RESULTS: Immunization with cartilage proteoglycan induced severe arthritis in the C3H/HeJCr substrain (95 100% incidence), whereas the original parent mice of the C3H/HeJ colony were resistant to proteoglycan (aggrecan)-induced arthritis. Furthermore, the progressive polyarthritis that is characteristic in susceptible C3H/HeJCr mice was accompanied by progressive inflammation around the spine. In subsequent experiments, 10 different C3H colonies with largely identical genetic backgrounds (all originating from the National Institutes of Health or Jackson Laboratory) exhibited extreme differences in susceptibility. Although none of the laboratory findings, including LPS hyporesponsiveness, immunologic parameters, and inflammatory markers, showed a correlation with susceptibility or resistance in the C3H/HeJCr and C3H/HeJ substrains, respectively, significant differences were found when all arthritic C3H mice were compared with all nonarthritic animals, regardless of their substrain origin. CONCLUSION: Because many of the C3H substrains lost arthritis susceptibility or acquired resistance, our results suggest that a preferred site for a mutation(s) in a gene(s) in a relatively upstream position of the inflammatory cascade is present. This is the first autoimmune model that exhibits extreme differences in arthritis susceptibility in the same murine strain, and is therefore a valuable tool for identification of arthritis-susceptible (or arthritis-suppressive) genes. PMID- 11263785 TI - Elevated proapoptotic Bax and caspase 3 activation in the NOD.scid model of Sjogren's syndrome. AB - OBJECTIVE: Salivary gland epithelial cells in patients with Sjogren's syndrome (SS) and in NOD and NODscid mice express Fas and Fas ligand, and these cells die from apoptosis. To elucidate the intracellular molecular mechanisms responsible for this salivary gland epithelial cell apoptosis, expression of the Bcl-2 family of proteins (Bcl-2, Bcl-xL, Bax) and caspase (caspases 3 and 8) was studied in young (ages 8-10 weeks) and old (ages 17-28 weeks) NOD and NOD.scid mice. METHODS: Sections of frozen salivary gland tissue were obtained from NOD and NOD.scid mice and from the lip biopsy material of SS patients. Immunohistochemistry or Western blot analysis was performed to assess the apoptotic-associated proteins. RESULTS: Levels of Bax and caspase 3 were elevated in the epithelial cells of glands from old NOD mice, but not in those from young NOD mice. In contrast, epithelial cells from both young and old NOD.scid mice exhibited strong expression of Bax and caspase 3. Western blot analysis showed that the activated form of caspase 3 was increased 2-5-fold in the glands from old NOD, old NOD.scid, and young NOD.scid mice compared with those from young NOD mice. Caspase 3 was also significantly elevated (P < 0.01) in SS patients whose focus scores were grade 3 or 4. In the SS patients' biopsy tissue and in the mouse glands, cells with fragmented DNA were positive for caspase 3. CONCLUSION: These results demonstrate that salivary gland epithelial cells in NOD and NOD.scid mice overexpress the proapoptotic molecules Bax and caspase 3. Bax could be the gene responsible for initiation of caspase activation, epithelial cell destruction, and lymphocyte glandular localization in SS. PMID- 11263786 TI - Cysteine protease activity is up-regulated in inflamed ankle joints of rats with adjuvant-induced arthritis and decreases with in vivo administration of a vinyl sulfone cysteine protease inhibitor. AB - OBJECTIVE: Cysteine proteases are postulated to play a role in tissue destruction in the joints of animals with arthritis. The purpose of the present study was to confirm the concept that cysteine proteases are enzymes involved in the pathology of rheumatoid arthritis (RA). METHODS: Arthritis was induced in Lewis rats by adjuvant injection (adjuvant-induced arthritis [AIA] model) and scored for inflammation. At necropsy, the rear paws were either fixed in formalin and assigned a histologic score (based on synovial cell proliferation, cartilage erosion, bone erosion, and fibroproliferative pannus) or frozen, cryosectioned, and assayed for enzyme activity either by in situ cytochemical staining with a post-azo-coupling method using a chromogenic substrate (Z-arg-arg-MNA) or by a novel assay placing the tissue section directly in a cuvette using the fluorogenic substrate Z-arg-arg-AMC. RESULTS: Enzymatic activity, measured either in frozen sections in situ or in the cuvette assay, was positively correlated with joint destruction (r = 0.7) and inflammation (r = 0.8). Activity was not inhibited significantly by Pefabloc (a serine protease inhibitor), EDTA (a metalloprotease inhibitor), or pepstatin A (an aspartyl protease inhibitor) but was inhibited by E-64 and vinyl sulfone irreversible inhibitors of cysteine proteases. The effect of one of the vinyl sulfone cysteine protease inhibitors, Mu-Leu-HomoPhe-vinylsulfone, was tested in vivo by dietary administration at 2.2 mg/kg/day in the AIA model; this resulted in a significant decrease in inflammation and in the amount of cysteine protease activity measured in the joint tissue. CONCLUSION: Cysteine protease activity levels increase in the diseased state and may be an important target for designing small molecule inhibitors to reduce the inflammation and tissue destruction associated with RA. PMID- 11263787 TI - Activation of a fibroblast-specific enhancer of the proalpha2(I) collagen gene in tight-skin mice. AB - OBJECTIVE: Reporter transgenes were introduced into the type 1 tight-skin (Tsk1/+) mouse model of scleroderma to test the hypothesis that fibroblast specific genetic programs are activated in fibrosis. METHODS: Transgenes harboring upstream fragments of the 5' flanking region of the mouse proalpha2(I) collagen gene (Col1a2), linked to a 400-bp minimal Col1a2 promoter driving an Escherichia coli beta-galactosidase (LacZ) reporter gene, were introduced into Tsk1/+ mice by breeding. Expression of these transgenes, which function as lineage-specific markers of fibroblast differentiation, was compared between the Tsk-LacZ mice and non-Tsk littermates. Responsiveness of these constructs to the profibrotic cytokine, transforming growth factor beta1 (TGFbeta1), was investigated by transient transfection of reporter constructs in tissue-culture cells. RESULTS: There was significant activation of reporter genes harboring the upstream enhancer in Tsk1/+ mice starting from 1 week of age. This was maximal at 6 weeks old (mean +/- SD 237 +/- 24% of non-Tsk controls; P= 0.001). Recombinant TGFbeta1 significantly activated reporter genes regulated by the upstream enhancer in transient transfection, and Tsk-LacZ fibroblasts showed elevated LacZ expression in tissue culture. CONCLUSION: These data suggest that activating signals in Tsk1/+ mice may act via fibroblast-specific regulatory elements within the murine Col1a2 gene. Although TGFbeta has been implicated in the pathogenesis of fibrosis, and reporter genes regulated by the upstream enhancer appear to be TGFbeta responsive in vitro, our results suggest that fibroblast-specific pathways may also be involved. PMID- 11263788 TI - Development of the tight-skin phenotype in immune-deficient mice. AB - OBJECTIVE: To determine if cutaneous thickening, a major phenotypic feature of the tight-skin (Tsk) mutation, could develop in an immune-deficient mouse. METHODS: Experimental crosses among different strains of mice were conducted to create mice that were genetically Tsk/+, and that were also homozgyous for a mutation at the Prkdc(scid) locus and thus lacked mature T and B lymphocytes. Skin samples prepared from experimental and control genotypic groups of mice were evaluated for skin thickness. RESULTS: The data showed that the Tsk/+ mice developed the Tsk phenotype in the absence of a functional immune system. CONCLUSION: Mature T and B cells are not required for the development of the cutaneous thickening in mice carrying the Tsk mutation. PMID- 11263789 TI - Prolonged remission without treatment after autologous stem cell transplantation for refractory childhood systemic lupus erythematosus. PMID- 11263790 TI - Up-regulated expression of transforming growth factor beta receptors in dermal fibroblasts in skin sections from patients with localized scleroderma. PMID- 11263791 TI - Updating the American College of Rheumatology preliminary classification criteria for systemic sclerosis: addition of severe nailfold capillaroscopy abnormalities markedly increases the sensitivity for limited scleroderma. PMID- 11263792 TI - Possible role of antiidiotypes in the loss of anti-topoisomerase I antibodies: comment on the article by Kuwana et al. PMID- 11263793 TI - Mortality from coronary heart disease and acute myocardial infarction--United States, 1998. AB - Despite improved clinical care, heightened public awareness, and widespread use of health innovations, coronary heart disease (CHD) remains the leading cause of death in the United States, and the decline in rates from CHD that began during the 1960s slowed during the 1990s. This report provides national and state specific death rates for CHD and for acute myocardial infarction (AMI). During 2001, approximately 1.1 million persons are expected to have a CHD event. Prevention remains the key strategy for reducing CHD mortality. PMID- 11263794 TI - Impact of the 1999 AAP/USPHS joint statement on thimerosal in vaccines on infant hepatitis B vaccination practices. AB - On July 8,1999, the American Academy of Pediatrics (AAP) and the U.S. Public Health Service (PHS) jointly recommended reducing infant exposure to thimerosal, a commonly used vaccine preservative that contains mercury. Specific recommendations were made to postpone the first hepatitis B vaccine dose until 2 6 months of age for infants born to hepatitis B surface antigen (HBsAg)-negative (i.e., not hepatitis B virus [HBV]-infected) women. Infants born to HBsAg positive (i.e., HBV-infected) women, or to women whose HBsAg status was unknown, were recommended to receive postexposure prophylaxis with the first dose of hepatitis B vaccine administered within 12 hours of birth. By mid-September 1999, when adequate supplies of preservative-free hepatitis B vaccine became available, PHS advocated a return to previous infant hepatitis B vaccination practices, including administering the first dose of hepatitis B vaccine to newborns in hospitals that had discontinued the practice. In 2000, preliminary assessments of the impact of these policy changes on routine hepatitis B vaccination practices were conducted by public health officials in Wisconsin, Oklahoma, Oregon, and Michigan. This report summarizes the results of these analyses, which indicate that many hospitals in Wisconsin have not reinstated policies to ensure routine administration of hepatitis B vaccine to newborns despite the availability of preservative-free hepatitis B vaccine, that the number of hepatitis B vaccine doses given to newborns in Oklahoma and Oregon has declined, and that an unvaccinated Michigan infant died from fulminant hepatitis B. Restoring routine newborn hepatitis B vaccination practices may require active advocacy by professional and government groups. PMID- 11263795 TI - Risk for meningococcal disease associated with the Hajj 2001. AB - Every year approximately two million pilgrims from more than 140 countries gather in Saudi Arabia for a pilgrimage to the holy places of Islam known as the Hajj. Coinciding with the Hajj pilgrimage during March 2000, Saudi Arabian health officials identified an outbreak of meningococcal disease; a substantial proportion of the isolates were the bacterial strain Neisseria meningitidis serogroup W-135. Four cases of meningococcal disease subsequently were identified among the estimated 15,000 pilgrims returning to the United States, their close contacts, and community. In addition, approximately 400 cases of meningococcal disease caused by N. meningitidis serogroup W-135 wereidentified worldwide during 2000. Whether an outbreak of meningococcal disease will recur in 2001 is unknown. PMID- 11263796 TI - Food, water, insulin, and the upright position. PMID- 11263797 TI - Prominent Q waves in a preoperative ECG. PMID- 11263798 TI - New approaches to pulmonary hypertension. AB - Careful evaluation will often reveal secondary--and perhaps reversible--factors contributing to pulmonary hypertension. For primary disease, there are now a variety of treatments, ranging from calcium channel blockers to lung transplantation. PMID- 11263799 TI - COX-2 inhibitors and the kidney. AB - Prostaglandins produced by the COX-2 isoform of cyclooxygenase may be important for renal function--and perhaps even more so in diseases that render the kidney highly prostaglandin-dependent. COX-2 inhibitors may thus have much the same renal risks that a standard NSAID does. Several clinical trials have already focused on the new drugs' renal effects. PMID- 11263800 TI - Cancer screening in the elderly. AB - A number of disease- and patient-specific factors need to be taken into account when cancer screening is considered in an older patient. They include the impact of aging on the cancer's biology and screening test performance, the patient's remaining years of life and candidacy for further diagnostic testing and available therapies, potential barriers to compliance with screening, and the patient's values and preferences about the screening. PMID- 11263801 TI - Eating disorders: 2. Bulimia nervosa. PMID- 11263802 TI - Blood compatibility--a perspective. AB - This perspective on blood- materials interactions is intended to introduce the set of papers stemming from the symposium, "Devices and Diagnostics in Contact with Blood: Issues in Blood Compatibility at the Close of the 20th Century," organized on August 4-6, 1999 at the University of Washington by the University of Washington Engineered Biomaterials (UWEB) Engineering Research Center. This article outlines some of the history of blood contacting materials, overviews the work that has originated at the University of Washington over the past 28 years, speculates on the origins of the controversies on blood compatibility and considers the issues that should be addressed in future studies. PMID- 11263803 TI - In vitro and in vivo studies of PEO-grafted blood-contacting cardiovascular prostheses. AB - The initial step of thrombus formation on blood-contacting biomaterials is known to be adsorption of blood proteins followed by platelet adhesion. Poly(ethylene oxide) (PEO) has been frequently used to modify biomaterial surfaces to minimize or prevent protein adsorption and cell adhesion. PEO was grafted onto a number of biomaterials in our laboratory. Nitinol stents and glass tubes were grafted with PEO by priming the metal surface with trichlorovinylsilane (TCVS) followed by adsorption of Pluronic and y-irradiation. Nitinol stents were also coated with Carbothane for PEO grafting. Chemically inert polymeric biomaterials, such as Carbothane, polyethylene, silicone rubber, and expanded polytetrafluoroethylene (e-PTFE), were first adsorbed with PEO-polybutadiene-PEO (PEO-PB-PEO) triblock copolymers and then exposed to gamma-irradiation for covalent grafting. For PEO grafting to Dacron (polyethylene terephthalate), the surface was sequentially treated with PEO-PB-PEO and Pluronics followed by gamma-irradiation. In vitro studies showed substantial reduction in fibrinogen adsorption and platelet adhesion to the PEO-grafted surfaces compared with control surfaces. Fibrinogen adsorption was reduced by 70-95% by PEO grafting on all surfaces, except for e PTFE. The platelet adhesion corresponded to the fibrinogen adsorption. When the PEO-grafted surfaces were tested ex vivo/in vivo, however, the expected beneficial effect of PEO grafting was inconsistent. The beneficial effect of the PEO grafting was most pronounced on the PEO-grafted nitinol stents. Thrombus formation was reduced by more than 85% by PEO grafting on metallic stents. Only moderate improvement (i.e. 35% decrease in platelet deposition) was observed with PEO-grafted tubes of polyethylene, silicone rubber, and glass. For PEO-grafted heart valves made of Dacron, however, no effect of PEO grafting was observed at all. It appears that the extent of thrombus formation on PEO-grafted biomaterials was directly related to the extent of tissue damage during implantation surgery. Platelets can be activated and form aggregates in the bulk blood, and the formed platelet aggregates may be able to deposit on the PEO monolayer overcoming its repulsive property. Our studies indicate that the testing of in vitro platelet adhesion should include adhesion of large platelet aggregates, in addition to adhesion of individual platelets. Furthermore, the surface modification methods should be improved over the current monolayer grafting concept so that the repulsive force by the grafted PEO layers is large enough to prevent adhesion of platelet aggregates formed in the bulk blood before arriving at the biomaterial surface. PMID- 11263805 TI - Use of simple and complex in vitro models for multiparameter characterization of human blood-material/device interactions. AB - Medical devices, intended for blood contacting applications, undergo extensive in vitro testing followed by animal and clinical feasibility studies. Besides the use of materials known to be intrinsically blood-compatible, the surface of such devices is often modified with a coating in order to improve the performance characteristics during blood exposure. In vitro evaluation of blood-device interactions accompanies the product development cycle from the early design phase using basic material geometries until final finished-product testing. Specific test strategies can vary significantly depending on the end application, the particular study objectives and variables of interest, and cost. To examine the degree to which findings derived from two different in vitro approaches complement one another, this report contrasts findings from a simple multipass loop model with findings from a simulated cardiopulmonary bypass (CPB) model. The loop model consists of tubular test materials, with and without surface modification, formed into valved Chandler loops. The CPB model has an oxygenator with and without surface modification connected to a reservoir and a blood pump. The surface modifications studied in this report are the Carmeda BioActive Surface and Duraflo II heparin coatings. Common blood parameters in the categories of coagulation, platelets, hematology, and immunology were monitored in each model. Ideal models employ the optimal level of complexity to study the design variables of interest and to meet practical cost considerations. In the case of medical device design studies, such models should also be predictive of performance. In the more complex and realistic simulated CPB model, experimental design and cost factors prevented easy/optimum manipulation of critical variables such as blood donor (use of paired samples) and heparin level. Testing in the simpler loop model, on the other hand, readily offered manipulation of these variables, and produced findings which overlapped with observations from the more complex CPB model. Thus, the models described here complimented one another. Moreover, conclusions from consistent findings, such as favorable responses associated with the heparin coatings, between the two models were considered to be more robust. PMID- 11263804 TI - Exploiting the current paradigm of blood-material interactions for the rational design of blood-compatible materials. AB - The paradigm of tissue material interactions, which holds that protein adsorption is the first event following contact and determines the later interactions of cells, is invoked to propose a design strategy for biocompatibility. Control of protein interactions is the key element, and it is suggested that nonspecific protein adsorption must be prevented while the adsorption of specific proteins that are expected to result in appropriate bioactivity must be promoted. Modification with polyethylene oxide has been investigated extensively as a means of preventing nonspecific adsorption. Examples of proteins that could be targeted for specific adsorption are antithrombin III to prevent coagulation and albumin to minimize platelet adhesion. Two examples of surfaces designed for specific adsorption from the author's laboratory are discussed: the incorporation of thrombin binding peptides to give a thrombin scavenging surface, and the incorporation of lysine to give a plasminogen specific surface with the potential to dissolve clots. PMID- 11263806 TI - What really is blood compatibility? AB - The criteria for nonthrombogenicity are classically defined as long clotting times and minimal platelet deposition. The inability to point to unequivocal progress in the development of truly nonthrombogenic materials, highlights the inadequacy if not actually invalidity of these criteria. Our approach is to define nonthrombogenicity in terms of: (1) a thrombin production rate constant, kp < 10(-4) cm s(-1); (2) low platelet consumption and low degree of platelet activation (e.g., microparticle formation); (3) perhaps some platelet spreading; and (4) low complement and leukocyte activation. Only when the target becomes clear, will it be possible to identify clear strategies for producing the materials we need. PMID- 11263807 TI - Antithrombogenic polymer alloy composed of 2-methacryloyloxyethyl phosphorylcholine polymer and segmented polyurethane. AB - To evaluate the antithrombogenicity of a new polymeric biomaterial in vivo, a polymer alloy tube composed of poly[2-methacryloyloxyethyl phosphorylcholine(MPC) co-2-ethylhexyl methacrylate](PMEH) polymer and a segmented polyurethane (SPU) was prepared by a solvent evaporation method on a Teflon rod from a homogeneous solution containing both the PMHE and SPU. The composition of the PMEH vs the SPU was 10 wt%. The inner and outer surfaces of the polymer alloy tubing were characterized by X-ray electron spectroscopic (XPS) measurements. The MPC units were located on the inner surface of the polymer alloy tubing rather than the outer surface. After immersion in aqueous media, a higher concentration of the MPC units was observed on both surfaces. Selective staining of the MPC units with osmium tetraoxide was carried out to observe the morphology of the PMEH domain on the surface of the polymer alloy. There were large-sized PMEH domains on the inner surface of the tubing but small-sized domains were found on the outer surface. This result was in good agreement with the XPS results. Blood compatibility of the polymer alloy was evaluated by observation of fibrinogen adsorption and platelet adhesion from human plasma. A lot of fibrinogen was adsorbed and many platelets adhered to the inner surface of the original SPU tubing. On the other hand, the PHEH/SPU polymer alloy tubing suppressed these adsorptions and adhesions. When the PMEH/SPU polymer alloy tubing was implanted into a rabbit's artery, thrombus could not be observed even after a 7-day implantation but the original SPU tubing was almost totally occluded only after a 90-min implantation due to serious thrombus deposition on the surface. These results clearly indicated that the PMEH in the SPU matrix acted as an antithrombus reagent by suppression of protein adsorption and platelet adhesion and activation. Particularly, the MPC units played a significant role in this function. PMID- 11263808 TI - Flow cytometric evaluation of material-induced platelet and complement activation. AB - Flow cytometry is used to characterize the activation state of platelets and leukocytes within whole blood after contact for 4 h at 37 degrees C with various materials under conditions of low shear. The contact involved adding heparinized whole blood to small diameter tubes that were connected to two arms extending from a rocking platform. For all surfaces (polyethylene, polypropylene, Silastic, PVA hydrogel) tested there was strong evidence of platelet activation in the bulk blood: platelet-derived microparticles. P-selectin expression and platelet leukocyte aggregates. Only contact with PVA hydrogel surfaces led to dramatic increases in CD11b up-regulation on monocytes and neutrophils that was inhibited by complement inhibition (sCRI). Flow cytometry was also used to evaluate the effectiveness of various agents to inhibit material-induced complement activation. The assay involved incubating 10 microm polystyrene beads for 1 h with serum at 37 degrees C before isolating the beads so as to label them with a monoclonal antibody against a neoantigen on SC5b-9. The beads were then identified by flow cytometry and the fluorescence associated with their SC5b-9 level recorded. The ability of C1-INH, pentamidine and benzamidine to moderately inhibit SC5b-9 levels suggests a role for classical complement activation in material-induced complement activation. PMID- 11263809 TI - Active tissue factor shed from human arterial smooth muscle cells adheres to artificial surfaces. AB - Through a series of in vitro assays, this study outlines a flow-mediated process by which active tissue factor (TF), the prime initiator of coagulation, may be transferred from the plasma membrane of vascular smooth muscle cells (VSMCs) to that of artificial surfaces such as those typically associated with intravascular implants. Studies with quiescent and activated rat VSMCs demonstrated that pathologically high shear stresses (tau(w) = 250 dyn cm(-2)) resulted in the loss of TF activity from the cell surface. Subsequent experiments with human VSMCs showed that VSMCs continuously release active TF into their extracellular medium, presumably in the form of lipid vesicles or microparticles, and that fluid shear stress (tauw = 50 dyncm(-2)) or chemical agonists (A23187) can significantly accelerate this release. Experiments with a wide array of polymeric and metallic materials showed that the TF shed from VSMCs was able to adhere to these surfaces and promote the activation of coagulation factor X (FX) at the material surface. Extracellular TF bound strongly to both uncoated and human plasma coated surfaces under a wide range of hemodynamic shear stresses (0-20 dyncm(-2)). When an extracellular, VSMC-derived TF mixture was perfused over Ti 6-4 surfaces, the adhesion of TF was found to be time-dependent, gradually accumulating on the material surface over time. Thus an important criterion in the design or success of intravascular devices may be related to their ability to interact with TF, shed from cell surfaces. This is especially important as TF may lead to thrombotic complications, the products of which may also increase cellular proliferation. PMID- 11263810 TI - The influence of thrombus components in mediating bacterial adhesion to biomaterials. AB - Thrombosis and infection represent the two largest limiting factors determining the long term success of implanted biomaterials. Infections associated with biomaterials are difficult to treat, and appear to evade the host defense systems. Mechanisms relating infection to thrombosis are described. Investigations into the role of receptors in mediating adhesion to thrombi are also discussed, in addition to strategies to reduce bacterial adhesion to biomaterial surfaces. PMID- 11263811 TI - Blood biocompatibility analysis in the setting of ventricular assist devices. AB - Ventricular assist devices (VADs) are increasingly applied to support patients with advanced cardiac failure. While the benefit of VADs in supporting this patient group is clear, substantial morbidity and mortality occur during the VAD implant period due to thromboembolic and infective complications. Efforts at the University of Pittsburgh aimed at evaluating the blood biocompatibility of VADs in the clinical, animal, and in vitro setting over the past decade are summarized. Emphasis is placed on understanding the mechanisms of thrombosis and thromboembolism associated with these devices. PMID- 11263812 TI - On the blood compatibility of end-point immobilized heparin. PMID- 11263813 TI - Angiogenic mechanisms of endothelialization of cardiovascular implants: a review of recent investigative strategies. AB - Both cardiovascular implants and therapeutic interventions on native arteries fail due to biologic responses occurring at the blood/prosthesis/arterial wall and tissue/prosthesis/arterial wall interfaces, resulting in the failure modes of thrombosis and myointimal hyperplasia. Systemic pharmacologic approaches including use of anti-coagulant and anti-platelet agents have significant untoward side effects and have not resulted in a dramatic impact on failure modes in many applications, including small diameter vascular grafts. Local delivery of therapeutic agents via surface attachment with defined release kinetics may alter thrombogenicity and/or myointimal hyperplasia. Therapeutic agents may include a spectrum of biologic agents from peptides to endothelial cells. Efficient attachment and release of these agents in biologically active form is dependent upon improved methods of surface modification. The intended action of the biologic agent may similarly be impacted by the surface and bulk characteristics of the underlying biomaterial. It is often assumed, without concrete data. that surface re-endothelialization may have a beneficial impact on both thrombogenicity and myointimal hyperplasia. New clinical data on endothelial cell seeding has been supportive. Spontaneous re-endothelialization may be stimulated via an induced directed angiogenesis resulting in trans-interstitial capillarization and surface endothelialization. Recent advances in therapeutic angiogenesis have suggested the power of angiogenic factors to induce neovascularization of ischemic tissue beds. These concepts have been used to surface modify prosthetic devices with either VEGF or FGF and both in vitro and animal data suggest a potent stimulation of surface re-endothelialization. Neither of these growth factors is likely to be ideal. VEGF is relatively endothelial cell specific but is a relatively weak endothelial cell mitogen. FGF 1 and FGF-2 are more potent mitogens but are less cell specific. Recent work has led to the generation of mutant growth factors via site-induced mutagenesis and results of several such FGF mutants on endothelial cell and smooth muscle cell proliferative response have been studied. The use of 'designer growth factors' on cardiovascular implants and on manipulated native vessels may have a significant positive impact on re-endothelialization and thereby on the failure modes of thrombosis and myointimal hyperplasia. PMID- 11263814 TI - Pregnancy rate in beef heifers after synchronization to either random or programmed estrous cycles. AB - We hypothesized that heifers in diestrus at the beginning of a Syncro-Mate-B (SMB) regimen would have higher pregnancy rates to AI than heifers not in diestrus and that administration of a PGF2alpha analogue 11 d before a SMB regimen would increase pregnancy rates to AI. In both replicate years of Exp. 1, heifers (n = 150) were classified by stage of the estrous cycle at the beginning of a SMB regimen (d 0). Following implant removal (d 9), heifers were artificially inseminated 12 h after the onset of estrus (95.5% in estrus by 72 h). Blood samples were collected for progesterone (P4) analysis on d 0, 9, and 20. Pregnancy rates did not differ between yr 1 and 2. Pregnancy rate for heifers classified in diestrus (53.6%; n = 69) was higher (P = 0.06) than for heifers in metestrus (43.7%; n = 48). Pregnancy rate for proestrus (44.4%; n = 18) heifers was not different from that for heifers in the metestrus or diestrus groups. Mean plasma P4 concentration was affected by both treatment and day. Pregnancy rate was higher (P < 0.01) for heifers with P4 > 1 ng/mL plasma (51.6%; n = 120) than for heifers with P4 < or = 1 ng/mL plasma (23.3%; n = 30) on d 0. In Exp. 2, beef heifers (Santa Cruz; n = 195) were allotted to two treatments. Heifers (n = 98) in the control group were administered a conventional SMB treatment. Heifers (n = 97) in the PGF group were injected with PGF2alpha 11 d (d -11) before a SMB regimen. Progesterone concentration was determined from blood samples collected on d -11, -2, 0, and 9. All heifers were artificially inseminated 48 to 50 h after implant removal. At the beginning of the SMB regimen (d 0), a greater (P < 0.05) percentage of PGF (74.2%) than of control heifers (59.2%) were in diestrus (P4 > 1 ng/mL). Mean P4 concentration was not affected by treatment or day x treatment but differed (P < 0.05) among days. Pregnancy rate of cycling heifers was similar for PGF (36%) and control heifers (35.9%). Pregnancy rate was higher (P < 0.01) for heifers with P4 > 1 ng/mL plasma (37.6%) than for heifers with P4 < or = 1 ng/mL plasma (18.5%) on d 0. These results support the hypothesis that fertility is enhanced when a progestin synchrony regimen is initiated during diestrus, but methods to program estrous cycles to increase fertility warrant investigation. PMID- 11263815 TI - Oxytocin-induced cervical dilation and cervical manipulation in sheep: effects on laparoscopic artificial insemination. AB - The difficulty of cervical penetration severely limits the use of transcervical AI (TAI) in sheep, and trauma from cervical manipulation (CM) may reduce fertility after TAI. We investigated the effects of cervical dilation using exogenous oxytocin (OT) to facilitate TAI and its effects on reproductive variables after laparoscopic AI (LAI). Estrus was synchronized by inserting pessaries impregnated with 6alpha-methyl-17alpha-hydroxyprogesterone acetate (60 mg) for 12 d. In Exp. 1, we determined whether OT and CM before LAI affected the interval from pessary removal to ovulation and fertilization rate. Crossbred ewes (n = 16) were assigned to 1) saline-CM or 2) OT-CM. In Exp. 2, effects of OT and CM on lambing rates were evaluated with white-faced ewes (n = 220) in a 2 x 2 factorial experiment: 1) saline-sham CM; 2) saline-CM; 3) OT-sham CM; and 4) OT CM. In both studies, eCG (400 IU i.m.) was injected at pessary removal, and LAI was performed 48 to 52 h later. In Exp. 1, ewes received i.v. either 400 USP units of OT or 20 mL of saline at 30 to 60 min before LAI, and CM was administered as for TAI. Beginning 32 h after pessary removal and continuing at 8 h intervals, ovaries were examined with ultrasonography to estimate time of ovulation. Treatment in Exp. 1 did not affect combined ovum/embryo recovery rate (69%), but OT-CM decreased fertilization rate (47 vs 59%; P < 0.05). The OT tended to reduce the interval to ovulation (OT, 59 h vs saline, 66 h; P < 0.06). The OT x CM interaction in Exp. 1 was not significant. For Exp. 2, approximately 25 min before sham CM or CM, 200 USP units of OT or 10 mL of saline was injected i.v. The LAI was performed immediately after sham CM or CM. At 10 to 12 d after AI in Exp. 2, ewes were mated with Suffolk rams. Blood was collected between 24 and 26 d after AI for pregnancy-specific protein B (PSPB) RIA. The PSPB pregnancy and lambing rates were both 62% in saline-sham controls. The CM did not affect pregnancy (69%) or lambing rate (64%). The OT treatment decreased (P < 0.05) PSPB pregnancy (59%) and lambing rates (56%) in OT-sham ewes and pregnancy and lambing rates in CM ewes (both 43%). Neither CM nor OT before LAI affected lambing rates to next estrus, indicating no long-term damage to the cervix or uterus. In summary, CM did not affect fertility after LAI, but OT decreased lambing rate independent of CM. If OT will not be usable for TAI, it may still be a tool for training TAI personnel. PMID- 11263816 TI - Influence of undegraded intake protein on reproductive performance of primiparous beef heifers maintained on stockpiled fescue pasture. AB - The objectives of this study were to evaluate the effects of pre- and postpartum undegraded intake protein (UIP) supplementation on body condition score (BCS), BW, calf weight, milk production, serum IGF-I concentrations, and postpartum interval in primiparous beef heifers (n = 44). Heifers were maintained on endophyte-free stockpiled tall fescue (11.7% CP, 38% ADF) and individually fed supplement daily beginning 60 d prepartum. Pre- and postpartum supplements provided 19.3% CP, 83.4% TDN (UIP); 14.1% CP, 84.1% TDN (Control); 21.5% CP, 81.5% TDN (UIP); and 14.6% CP, 81.4% TDN (Control); respectively. Blood meal (146 g/d) was the source of UIP. Six heifers were removed from the study due to calf loss unrelated to treatment; therefore, postpartum measurements are based on 19 animals per treatment. Statistical analyses using ANOVA and a split-plot design revealed no effects of treatment (P > 0.2) on BCS, BW, calf weight, milk production, or postpartum interval. There tended to be a treatment x time interaction on BCS (P < 0.09) with UIP heifers having higher BCS than Control at wk 5, 7, and 9 postpartum. There was a treatment x time interaction on serum IGF I (P < 0.06) during the first 35 d postpartum. In UIP heifers, serum IGF-I was greater at calving compared with Control heifers (117.5 vs 92.4 ng/mL, respectively); however, these differences were not related to changes in BCS or BW. Although serum IGF-I concentrations were increased at calving in heifers receiving UIP, there were no treatment effects on postpartum interval (P > 0.7). During the first 30 d postpartum, IGF-I differed (P < 0.01) among heifers with postpartum intervals defined as short, < 50 d (128.9 ng/mL); medium, 51 to 65 d (115.2 ng/mL); and long, 66 to 130 d (52.9 ng/mL). When analyzed as a regression, a 1 ng/mL increase in IGF-I (UIP and Control heifers) at calving (P < 0.05) and throughout the postpartum period (P < 0.01) corresponded to a decrease in postpartum interval of 0.13 d. Based on the results of this study, the inclusion of UIP in diets for primiparous heifers and its effects on postpartum interval warrant further evaluation. PMID- 11263817 TI - Effects of feeding high-oil corn to beef steers on carcass characteristics and meat quality. AB - To assess the effects of feeding high-oil corn on carcass characteristics and meat quality, 60 yearling steers were fed high concentrate diets containing either control corn (82% of diet), high-oil corn (82% of diet), or high-oil corn at a concentration that was isocaloric with the control diet (74% of diet). After being fed for 84 d, steers were slaughtered. At 72 h postmortem, carcass data were collected and rib sections from five steers grading U.S. Choice and five steers grading U.S. Select from each treatment were collected, vacuum packaged, and aged for 14 d. Three steaks (2.54 cm thick) were removed from each rib for Warner-Bratzler shear force measurement, sensory appraisal, and fatty acid composition analyses. Data were analyzed with treatment as the main effect for the carcass data and treatment, quality grade, and two-way interaction in the model for the longissimus data. The two-way interaction was nonsignificant (P > 0.05) for all variables tested. No differences were detected (P > 0.05) in carcass measurements except for marbling scores and quality grades, both of which were greater (P < 0.05) for carcasses from steers fed the high-oil corn. Overall, 78% of steers fed the high-oil corn graded U.S. Choice compared with 47% for the control and 67% for isocaloric group. Shear force and sensory properties of the longissimus were not different (P > 0.05) among treatments. Steaks from U.S. Choice carcasses rated higher (P < 0.05) for tenderness and tended to rate higher (P < 0.10) for juiciness. Feeding the isocaloric and high-oil diets increased (P < 0.05) linoleic acid, arachidonic acid, and the total PUFA content of lipid extracted from the longissimus. Saturated fatty acid percentage was lowest (P < 0.05) for high-oil corn and highest (P < 0.05) for control, with isocaloric being intermediate. Feeding high-oil corn increased (P < 0.05) pentadecyclic acid, margaric acid, and total odd-chain fatty acid content. Feeding high-oil corn in finishing beef cattle diets enhanced intramuscular lipid deposition and increased unsaturation of fatty acids of the longissimus. PMID- 11263818 TI - Intervals from norgestomet withdrawal and injection of equine chorionic gonadotropin or P.G. 600 to estrus and ovulation in ewes. AB - Synchronization of estrus and ovulation is essential for AI of ewes during a predetermined time frame, and progestogen-eCG treatments are typically used to prepare the ewes. However, eCG is not readily available in the United States, but P.G. 600 (400 IU of eCG and 200 IU of hCG) is available. Thus, we conducted a study to determine the effects of eCG and P.G. 600 on the timing of estrus and ovulation after progestogen withdrawal. Ewes were assigned to two replicates of an experiment with the following treatments: 1) 3-mg norgestomet implant (i.e., one-half of a Syncro-Mate-B [SMB] implant) for 10 d, plus 2 mL of saline i.m. at SMB removal (n = 11); 2) 3-mg SMB implant for 10 d, plus 400 IU of eCG i.m. at SMB removal (n = 13); and 3) 3-mg SMB implant for 10 d, plus P.G. 600 i.m. at implant removal (n = 9). On d 6 after SMB insertion, PGF2alpha was used to induce luteolysis. Beginning 12 h after implant removal, vasectomized rams were used at 12-h intervals to check for estrus. When a ewe was detected in estrus, each ovary was evaluated ultrasonically. Ovaries were evaluated again 16 h later and then at 8-h intervals until ovulation. Treatment altered the interval from implant removal to estrus (less [P < 0.05] in SMB + eCG than in the other two groups) and to ovulation (greatest [P < 0.05] in SMB). However, the treatment x replicate interaction was significant for the intervals from implant removal to estrus (P < 0.03) and from implant removal to ovulation (P < 0.05). An inconsistent response in the SMB-treated ewes seemed to be primarily responsible for the interaction. The intervals to estrus and to ovulation for the SMB-treated ewes were shorter (P < 0.05) in Replicate 1 than in Replicate 2. Also, both intervals seemed to be less consistent between replicates for the SMB + P.G. 600- than for the SMB + eCG treated ewes; that is, eCG seemed to increase the predictability of the intervals to estrus and to ovulation. Neither the main effects of treatment and replicate nor their interaction were significant for the interval from estrus to ovulation (38.4 /- 3.3 h), size of the ovulatory follicle (7.7 +/- 0.8 mm), or ovulation rate (1.6 +/- 0.2). We concluded from this experiment that eCG is a better choice than P.G. 600 as the gonadotropin to use at the time of progestogen withdrawal to prepare ewes for AI during a predetermined interval. PMID- 11263819 TI - Genetic evaluation of carcass traits in Simmental-sired cattle at different slaughter end points. AB - Our objectives were to estimate genetic parameters for carcass traits and evaluate the influence of slaughter end point on estimated breeding values (BV). Data provided by the American Simmental Association were divided into three sets: 1) 9,604 records of hot carcass weight (CW) and percentage retail cuts (PRC), 2) 6,429 records of CW, PRC, and marbling score (MS), and 3) 1,780 records of CW, PRC, MS, fat thickness (FT), and longissimus muscle area (LMA). Weaning weights (WW) from animals with carcass data and from their weaning contemporaries were used. Data were analyzed with a multiple-trait animal model and REML procedures to estimate genetic parameters and BV on an age-, CW-, MS-, or FT-constant basis. The model for carcass traits included fixed contemporary group and covariates for breed, heterozygosity, and slaughter end point and random additive direct genetic and residual effects. Weaning weight was preadjusted for founder effects, direct and maternal heterosis, age of dam, and age of calf. The model for WW included fixed contemporary group and random additive direct genetic, maternal genetic, maternal permanent environment, and residual effects. Heritabilities from data set 1 were 0.34 for CW and 0.25 for PRC on an age-constant basis and 0.25 for PRC on a CW end point. Heritabilities for data set 2 were 0.35, 0.24, and 0.36 for CW, PRC, and MS, respectively, on an age-constant basis. Data set 2 heritabilities were 0.25 for PRC and 0.34 for MS on a CW-constant basis and 0.33 for CW and 0.25 for PRC at a constant MS end point. Heritabilities on an age constant basis for data set 3 were as follows: CW, 0.32; PRC, 0.09; MS, 0.12; FT, 0.10; and LMA, 0.26. Heritability estimates for data set 3 on a CW-, MS-, and FT constant basis were similar to those on an age-constant basis. Heritabilities were 0.12 for PRC, 0.12 for MS, 0.14 for FT, and 0.22 for LMA on a CW-constant basis; 0.30 for CW, 0.09 for PRC, 0.10 for FT, and 0.28 for LMA at a constant MS end point; and 0.33, 0.17, 0.13, and 0.29 for CW, PRC, MS, LMA on a FT-constant basis. Genetic correlations among traits varied across groups and end points but suggested that it should be possible to select for improved lean yield without sacrificing quality grade. Correlations were calculated among BV computed at different end points. Adjustment to various end points resulted in some changes in BV and reranking of sires, especially for PRC; however, the number of records available had a larger influence than slaughter end point. PMID- 11263820 TI - Estimation of genetic covariances with method R. AB - Method R is a simple and computationally inexpensive method for estimating (co)variances. The objective of the study was to investigate properties of Method R for estimation of (co)variance components with emphasis on covariance estimation. Theoretical Method R formulas were developed for simplified single variate and bivariate models. In single-trait models, the curve of the regression of Method R was continuous and monotonic and its slope depended on the amount of information on each animal and on the variance ratio. The curve became steeper as the number of records per animal decreased. For covariance, the curve of the regression was monotonic but not continuous. However, a regression coefficient of 1 still corresponded to the correct covariance. Similar curves were observed in analyses of simulated data sets. Because of the observed discontinuity, algorithms implementing Method R that require a continuous regression curve would not work in models with covariances. An alternative algorithm was based on a transformation matrix obtained by multiplying a matrix of numerators with the inverse of a matrix of denominators of the regression factors. Such an algorithm converged reliably for all models tested. Method R can be modified to estimate covariances in models too large for other methods. PMID- 11263821 TI - Mapping of multiple quantitative trait loci by simple regression in half-sib designs. AB - Detection of QTL in outbred half-sib family structures has mainly been based on interval mapping of single QTL on individual chromosomes. Methods to account for linked and unlinked QTL have been developed, but most of them are only applicable in designs with inbred species or pose great demands on computing facilities. This study describes a strategy that allows for rapid analysis, involving multiple QTL, of complete genomes. The methods combine information from individual analyses after which trait scores for a specific linkage group are adjusted for identified QTL at other linkage groups. Regression methods are used to estimate QTL positions and effects; permutation tests are used to obtain empirical threshold values. The description of the methods is complemented by an example of the combined analysis of 28 bovine chromosomes and their associations with milk yield in Finnish Ayrshire cattle. In this example, the individual analysis revealed five suggestive QTL affecting milk yield. Following the strategy presented in this paper, the final combined analysis showed eight significant QTL affecting milk yield. This clearly demonstrates the potential gain of using the combined analysis. The use of regression methods, with low demands on computing resources, makes this approach very practical for total genome scans. PMID- 11263822 TI - Identification of quantitative trait loci affecting reproduction in pigs. AB - The objective of this research was to identify chromosomal regions harboring QTL affecting reproduction in pigs. A three-generation resource population was developed by crossing low-indexing pigs from a randomly selected control line (C) with high-indexing pigs of a line selected for increased index of ovulation rate and embryonic survival (I). Differences between Lines I and C at Generation 10 were 6.7 ova and 3.3 fetuses at 50 d of gestation and 3.1 fully formed and 1.6 live pigs at birth. Phenotypic data were collected on F2 females, born in three replicates, for ovulation rate (n = 423), age at puberty (n = 295), litter size (n = 370), and number of nipples (n = 428). Litter-size data included number of fully formed, live, stillborn, and mummified pigs. Grandparent, F1, and F2 animals were genotyped for 151 microsatellite markers distributed across all 18 autosomes and the X chromosome. Genotypic data were available on 423 F2 females. Average spacing between markers was 19.3 Kosambi centimorgans. Calculations of logarithms of odds (LOD) scores were by least squares, and fixed effects for sire dam combination and replicate were included in the models. Genome-wide significance level thresholds of 5% and 10% were calculated using a permutation approach. There was evidence (P < 0.05) for QTL affecting ovulation rate on SSC9, age at puberty on SSC7 and SSC8, number of nipples on SSC8 and SSC11, number of stillborn pigs on SSC5 and SSC13, and number of fully formed pigs on SSC11. There was evidence (P < 0.10) for additional QTL affecting age at puberty on SSC7, SSC8, and SSC12, number born live on SSC11, and number of nipples on SSC1, SSC6, and SSC7. Litter size is lowly heritable and sex-limited. Therefore, accuracy of selection for litter size may be enhanced by marker-assisted selection. Ovulation rate and age at puberty are laborious to measure, and thus marker-assisted selection may provide a practical and efficient method of selection. PMID- 11263823 TI - The "new perception" of animal agriculture: legless cows, featherless chickens, and a need for genuine analysis. AB - A growing popular literature has created a "New Perception" of animal agriculture by depicting commercial animal production as 1) detrimental to animal welfare, 2) controlled by corporate interests, 3) motivated by profit rather than by traditional animal care values, 4) causing increased world hunger, 5) producing unhealthy food, and 6) harming the environment. Agricultural organizations have often responded with public relations material promoting a very positive image of animal agriculture and denying all six of the critics' claims. The public, faced with these two highly simplistic and contradictory images, needs knowledgeable research and analysis to serve as a basis for public policy and individual choice. Scientists and ethicists could provide such analysis. In some cases, however, scientists and ethicists have themselves produced misleading, polarized, or simplistic accounts of animal agriculture. The problems in such accounts include the repetition of unreliable information from advocacy sources, use of unwarranted generalizations, simplistic analysis of complex issues, and glossing over the ethical problems. The New Perception debate raises important and complex ethical issues; in order to provide useful guidance, both scientists and ethicists must consider these issues as research problems that are worthy of genuine investigation and analysis. PMID- 11263824 TI - The influence of farmers' behavior on calves' reactions to transport and quality of veal meat. AB - The relationships between farmers' behavior toward veal calves, calves' responses to handling and transport, and veal meat quality were assessed. Two groups of 10 veal units were selected based on previous observed farmers' behavior toward the calves: one group consisted of farmers who had shown predominantly "positive" behavior toward the calves, and the other group of farmers had shown predominantly "negative" behavior. Calves were observed for their reactions to people at the unit, and 20 calves per veal unit were transported either directly to the slaughterhouse or subjected to additional transport consisting of a supplementary 20-min transport with additional unloading and loading. The effort needed to load the calves onto the truck and their behavior during loading was observed. During loading and unloading, and during lairage at the slaughterhouse, potentially traumatic incidents (falling down, hits against structures, slips) were recorded, and heart rate and cortisol measurements were taken. Carcasses were evaluated on their weight, color, conformation, pH, and bruise level. A meat sample was taken from the longissimus thoracis muscle for physical, chemical, and sensory analysis. Calves originating from "positive behavior" units showed fewer fear responses to people at the veal unit, needed less effort to be loaded to the truck, had lower heart rates during loading and unloading, and had fewer incidents at the slaughterhouse than calves from "negative behavior" units (P < 0.05). Carcasses from calves from "positive behavior" units were paler, and analyses of the meat sample revealed lower pH, moisture level, and redness compared to carcasses from calves from "negative behavior" units (P < 0.05). Additional transport led to a lower cortisol level after transport and to higher carcass pH values at slaughter compared to direct transport (P < 0.05) but did not affect meat quality. We concluded that farmers' positive behavior toward veal calves during rearing is likely to reduce the emotional responses of calves to handling and transport and to lead to fewer incidents, compared to negative behavior. This reduction of calves' emotional responses seems to be the reason for improved veal meat color. PMID- 11263825 TI - Porcine leptin alters insulin inhibition of lipolysis in porcine adipocytes in vitro. AB - The present study determined whether porcine leptin can alter the lipolytic rate in porcine adipocytes produced in vitro. The stromal-vascular cell fraction of neonatal subcutaneous adipose tissue was isolated by collagenase digestion, filtration, and subsequent centrifugation. These stromal-vascular cells were seeded on 25-cm2 tissue culture flasks and proliferated to confluency in 10% fetal bovine serum in DMEM/F12 (50:50). Cultures were differentiated using 2% pig serum + 10 mM isobutyl methylxanthine + 1 microM dexamethasone for 48 h. This medium was replaced with 5% pig serum + 1 microM insulin to promote lipid filling of adipocytes for 7 d. Adipocyte-containing cultures were incubated overnight in serum-free medium and then used for experiments. Acute experiments assessed lipolysis in cultures exposed to porcine leptin (0 to 1,000 ng/mL medium) for 2 h. Chronic experiments used cultures incubated with 100 ng porcine leptin/mL of medium for 72 h prior to lipolysis measurements. Direct effects of leptin were examined by incubating cultures in DMEM/F12, 25 mM HEPES, 3% bovine serum albumin, 20 mU of adenosine deaminase/mL of medium in the presence of 0 to 1,000 ng of porcine leptin/mL of medium. Indirect effects of leptin were examined using the same incubation medium but also supplemented with 1 microM isoproterenol +/- 10 nM insulin in the presence of 0 to 1,000 ng of porcine leptin/mL of medium. Media glycerol concentration was measured at the end of 2-h incubations. Acute leptin exposure induced up to a 76% increase in lipolysis (P < 0.05) but had no effect on insulin's inhibition of lipolysis. Chronic exposure to leptin produced up to a 56% increase in lipolysis (P < 0.05) and reduced insulin's inhibition ofisoproterenol-stimulated lipolysis by up to 31% (P < 0.05). These data demonstrate leptin functions to promote the partitioning of energy away from lipid accretion within porcine adipose tissue by promoting lipolysis directly and indirectly by reducing insulin-mediated inhibition of lipolysis. PMID- 11263826 TI - National market cow and bull beef quality audit-1999: a survey of producer related defects in market cows and bulls. AB - The 1999 National Market Cow and Bull Beef Quality Audit comprised face-to-face interviews with industry representatives (n = 49); in-plant evaluations of cattle in holding pens (n = 3,969), carcasses on harvest floors (n = 5,679), and in carcass coolers (n = 4,378); and a strategy workshop. Face-to-face interviews suggested that the beef industry was most frequently concerned about the presence of antibiotic residues in carcasses, presence of lead shot in carcasses, and price discovery for carcasses following excessive trimming of bruises and testing due to arthritic joints, pathogens, or antibiotic residues. Although live animal evaluations determined that 73.4% of beef cows, 60.8% of dairy cows, 63.7% of beef bulls, and 70.9% of dairy bulls did not exhibit evidence of lameness, losses due to lameness were greater (P < 0.05) than in the 1994 National Non-Fed Beef Quality Audit. In-plant audits revealed that 88.9, 10.3, and 88.2% of cow carcasses and 18.9, 21.2, and 52.9% of bull carcasses had inadequate muscling, arthritic joints, and at least 1 bruise, respectively, all of which resulted in greater (P < 0.05) losses than the same defects in 1994. Audits revealed that 88.9% of cow carcasses and 18.9% of bull carcasses were lightly muscled, resulting in greater (P < 0.05) losses for cow carcasses, and similar (P > 0.05) losses for bull carcasses, than the same defect in the 1994 audit. Also, 14.5 and 30.8% of cow carcasses and 6.9 and 5.9% of bull carcasses had excess external fat and yellow-colored external fat, respectively, which was an improvement (P < 0.05) over 1994 results. In aggregate, 24.1, 19.2, 7.2, 6.7, 9.5, and 1.1% of livers, tripe, hearts, heads, tongues, and whole cattle or carcasses, respectively, were condemned and 60.6, 2.4, and 46.5% of cattle had hide damage from latent defects, insect damage, and brands, respectively. Condemnation rates were generally lower (P < 0.05), but tongue condemnations and frequency of branded hides were higher (P < 0.05) than in 1994. Producers should promote value in cows and bulls by managing to minimize quality defects, monitoring health and condition, and marketing in a timely manner. Using these techniques, producers might have recaptured $13.82, $27.50, and $27.50, respectively, for each cow or bull harvested in 1999. PMID- 11263827 TI - Antioxidant status affects color stability and tenderness of calcium chloride injected beef. AB - The objectives of this study were to determine whether vitamin E supplementation influences color and tenderness of beef injected with calcium chloride. Market heifers (n = 12) were fed a standard finishing diet with minimal levels of vitamin E (NE group). Another 12 market heifers were fed the NE diet with the inclusion of 1,000 IU/d of DL-alpha-tocopherol per animal for the last 125 d on feed (E group). Animals were slaughtered after 125 d on the diets and upon reaching an ultrasound backfat thickness > 10 mm. Half of the longissimus muscles from each treatment group (NE and E) were pumped to 10% over the original weight with 250 mM CaCl2 (Ca) at 24 h postmortem. Remaining muscles (NE and E) were pumped to 10% over the original weight with water (NC) at 24 h postmortem. After equilibrating overnight, steaks (2.54 cm) were overwrapped with O2-permeable film and stored for 7 d after injection. Hunter "L," "a," and "b" values were obtained each day of storage. Trained panelists evaluated color on d 1, 4, and 7 after injection. 2-Thiobarbituric acid-reactive substances (TBARS) values were measured on d 1 and 7 after injection. Warner-Bratzler (W-B) shear force values and trained sensory panel evaluations at 1, 3, and 7 d after injection were obtained. Immunoblotting techniques were used to monitor the 30-kDa degradation product of troponin-T at 1, 3, and 7 d after injection. At 4 d after injection, E/Ca steaks were the least discolored (P < 0.05). The E/Ca steak TBARS values were not significantly different from values for NE/NC steaks at 7 d after injection, whereas NE/Ca steaks had greater (P < 0.05) TBARS values after 7 d following injection compared with all other groups. Treatment with Ca resulted in higher off-flavor scores (P < 0.05). The E/Ca samples had the most rapid tenderization and proteolysis of all treatment groups. Warner-Bratzler shear values were lower in the E/Ca samples than in the E/NC samples at 1, 3, and 7 d after injection (P < 0.05). No difference in shear force was noted between NE/Ca and NE/NC samples at any time point. No difference in sensory tenderness was noted between NE/Ca and NE/NC samples at 1 d after injection. However, Ca-injected samples (NE/Ca and E/Ca) were rated as being significantly more tender than their uninjected counterparts (NE/NC and E/NC) at 3 and 7 d after injection. Injection of CaCl2 may result in more rapid and immediate tenderization if beef from animals supplemented with vitamin E is used. Vitamin E incorporation into muscle tissue may potentiate the action of exogenously added calcium by protecting the calpains from oxidation. PMID- 11263828 TI - A survey of beef muscle color and pH. AB - The objectives of this study were to define a beef carcass population in terms of muscle color, ultimate pH, and electrical impedance; to determine the relationships among color, pH, and impedance and with other carcasses characteristics; and to determine the effect of packing plant, breed type, and sex class on these variables. One thousand beef carcasses were selected at three packing plants to match the breed type, sex class, marbling score, dark-cutting discount, overall maturity, carcass weight, and yield grade distributions reported for the U.S. beef carcass population by the 1995 National Beef Quality Audit. Data collected on these carcasses included USDA quality and yield grade data and measurements of muscle color (L*, a*, b*), muscle pH, and electrical impedance of the longissimus muscle. About one-half (53.1%) of the carcasses fell within a muscle pH range of 5.40 to 5.49, and 81.3% of the carcasses fell within a longissimus muscle pH range of 5.40 to 5.59. A longissimus muscle pH of 5.87 was the approximate cut-off between normal and dark-cutting carcasses. Frequency distributions indicated that L* values were normally distributed, whereas a* and b* values were abnormally distributed (skewed because of a longer tail for lower values, a tail corresponding with dark-cutting carcasses). Electrical impedance was highly variable among carcasses but was not highly related to any other variable measured. Color measurements (L*, a*, b*) were correlated (P < 0.05) with lean maturity score (-.58, -.31, and -.43, respectively) and with muscle pH (-.40, -.58, and -.56, respectively). In addition, fat thickness was correlated with muscle pH and color (P < 0.05). There was a threshold at approximately .76 cm fat thickness, below which carcasses had higher muscle pH values and lower colorimeter readings. Steer carcasses (L* = 39.62, a* = 25.20, and b* = 11.03) had slightly higher colorimeter readings (P < 0.05) than heifer carcasses (L* = 39.20, a* = 24.78, and b* = 10.80) even though muscle pH was not different between steer and heifer carcasses. Dairy-type carcasses (pH = 5.59, L* = 37.56, a* = 23.40, and b* = 9.68) had higher muscle pH values and lower colorimeter readings than either native-type (pH = 5.50, L* = 39.55, a* = 25.13, and b* = 11.00) or Brahman-type (pH = 5.46, L* = 39.75, a* = 25.17, and b* = 11.05) carcasses (P < 0.05). PMID- 11263829 TI - Evaluation of the Tendertec beef grading instrument to predict the tenderness of steaks from beef carcasses. AB - Four experiments were conducted, using carcasses from cattle identified for anticipated variability in tenderness (Exp. 1, 2, and 3) and carcasses selected for variability in physiological maturity and marbling score (Exp. 4), to evaluate the ability of the Tendertec Mark III Beef Grading Probe (Tendertec) to predict tenderness of steaks from beef carcasses. In Exp. 1, 2, and 3, longissimus steaks were aged for different periods of time, cooked to a medium degree of doneness (70 degrees C), and evaluated for Warner-Bratzler shear force (WBS) and trained sensory panel ratings. In Exp. 4, longissimus steaks were aged 14 d and cooked to 60, 65, 70, 75, or 80 degrees C for WBS tests and to 65 or 75 degrees C for sensory panel evaluations. Tendertec output variables were not correlated with 1) 24-h calpastatin activity, steak WBS (following 1, 4, 7, 14, 21, or 35 d of aging), or d-14 sensory panel tenderness ratings in Exp. 1 (n = 467 carcasses) or 2) 14-d WBS in Exp. 2 (n = 202 carcasses). However, in Exp. 3 (n = 29 carcasses), Tendertec output variables were correlated (P < 0.05) with tenderness of steaks aged 1, 21, 28, or 35 d, and we were able to separate carcasses into groups yielding tough, acceptable, and tender steaks. In Exp. 4 (n = 70), Tendertec output variables were correlated (P < 0.05) with steak WBS at 60 degrees C and with steak ratings for muscle fiber tenderness, connective tissue amount, and overall tenderness at 65 degrees C, but these relationships weakened (P > 0.05) as degree of doneness increased. Consequently, Tendertec output variables only were effective for stratifying carcasses according to tenderness when steaks from those carcasses in Exp. 4 were cooked to a rare or medium-rare degree of doneness. Although Tendertec was able to sort carcasses of older, mature cattle based on tenderness of steaks at some cooked end points, it failed to detect tenderness differences in steaks derived from youthful carcasses consistently, and was thus of limited value as an instrument for use in improving the quality, consistency, and uniformity of the U.S. fed-beef supply. PMID- 11263830 TI - Effect of diets containing n-3 fatty acids on muscle long-chain n-3 fatty acid content in lambs fed low- and medium-quality roughage diets. AB - In two experiments, each with 32 cross-bred ([Merino x Border Leicester] x Poll Dorset) wether lambs (26 to 33 kg weight range), animals were randomly assigned to one of four treatments. A mixture of lucerne chaff:oaten chaff was used as a basal diet, offered in different ratios. Animals were allowed to consume on a free-access basis in Exp. 1 or 90% of ad libitum intake in Exp. 2 in order to provide a low- (6.5 MJ ME/d) and medium- (9.5 MJ ME/d) quality basal diet, respectively. Isoenergetic amounts of lipid supplements, fish meal (80 g DM), canola meal (84 g DM), and soy meal (75 g DM) were tested in Exp. 1. In Exp. 2, fish meal (9% DM), unprotected rapeseed (7% DM), and protected canola seed (6% DM) were fed as supplements. At the end of 53-d (Exp. 1) or 46-d (Exp. 2) experimental periods, lambs were slaughtered at a commercial abattoir and at 24 h postmortem longissimus thoracis (LT) muscle was collected for the analysis of fatty acid (FA) composition of structural phospholipid and storage triglyceride fractions. Fish meal diet increased LT muscle long-chain n-3 FA content by 27% (P < 0.02) in Exp. 1 and 30% (P < 0.001) in Exp. 2 compared with lambs fed the basal diet, but fish meal decreased (P < 0.01) the n-6 FA content only in Exp. 1. Soy meal and protected canola seed diets increased (P < 0.01) LT muscle n-6 FA content but did not affect long-chain n-3 FA content. Longissimus thoracis muscle long-chain n-3 FA were mainly deposited in structural phospholipid, rather than in storage triglyceride. In both Exp. 1 and Exp. 2, the ratio of n-6:n-3 FA in LT muscle was lowest (P < 0.01) in lambs fed fish meal supplement compared with all other treatments. Protected canola seed diet increased the ratio of n-6:n-3 FA (P < 0.01) and PUFA:saturated fatty acid (P < 0.03) content from those animals fed the basal, fish meal, and unprotected rapeseed diets in Exp. 2. This was due to an increase in muscle n-6 FA content, mainly linoleic acid, of both phospholipid (P < 0.001) and triglyceride (P < 0.01) fractions and not to an increase in muscle n-3 FA content. The results indicate that by feeding fish meal supplement, the essential n-3 FA can be increased while lowering the ratio of n-6:n-3 content in lamb meat to an extent that could affect nutritional value, attractiveness, and the economic value of meat. PMID- 11263831 TI - True ileal digestibility of amino acids in sow's milk for 17-day-old pigs. AB - The digestibility of amino acids in sow's milk consumed by young pigs is currently unknown because of difficulties associated with collecting an adequate quantity of milk, and also problems in cannulating suckling pigs. A total of 14 kg of sow's milk was collected, two soluble indigestible markers (Co-EDTA and YbC13) were added, and the milk was fed to four pigs at 17 d of age that were fitted with a simple T-cannula at the terminal ileum. Another four cannulated pigs were offered a similar amount of a 20% DM liquid diet based on enzymatically hydrolyzed casein and lactose to assess endogenous amino acid losses. All pigs were fed about 875 g of each diet per day in 10 hourly meals from 0700 to 1700. Following 2 d of adaptation, ileal digesta were collected from 0800 to 1800 for 2 d. Diets and digesta were analyzed for amino acids using appropriate hydrolysis and preoxidation procedures. Average nitrogen true digestibility was 88%, whereas amino acid true digestibilities ranged from 84% (cystine and threonine) to 100% (methionine, histidine, and glutamic acid); the average for all amino acids was 92 +/- 4%. Based on average values, true digestibility of essential amino acids was not different from that of nonessential amino acids (P > 0.10). In whole milk, amino acids found in abundance in whey proteins (i.e., cystine, glycine, and threonine) were less (P < 0.05) digestible than amino acids predominating in casein proteins (i.e., glutamic acid, proline, and methionine). When true ileal digestible amino acid concentrations in sow's milk were expressed as ratios to digestible lysine, it appeared that threonine, tryptophan, and arginine were lower than what might be considered optimal. In conclusion, amino acids in sow's milk were highly digestible, but most of the amino acids had true ileal digestibility values significantly less than 100%. PMID- 11263832 TI - Effects of dietary conjugated linoleic acid in nursery pigs of dirty and clean environments on growth, empty body composition, and immune competence. AB - Early-weaned pigs (n = 64) averaging 5.3 +/- 0.3 kg and distributed into two environments (dirty and clean) were used to evaluate effects of conjugated linoleic acid (CLA) on growth performance, immune competence, and empty body composition. A factorial (2 x 4) arrangement within a split-plot design, with four littermate pigs as the experimental unit for the environment, pig within litter as the experimental unit for dietary treatment, and d-0 body weight used as covariate, were used in data analysis. Diets were formulated to contain CLA at 0, 0.67, 1.33, or 2% and to exceed the NRC (1988) nutrient needs of pigs. Animals were given ad libitum access to feed for 7 wk in three phases (I, 1 to 2; II, 3 to 5; and III, 6 to 7 wk). Within phases, diets were isocaloric and isonitrogenous. In Phase I, as dietary CLA concentration increased, ADG and ADFI decreased linearly (P < 0.05 and P < 0.02, respectively). In Phase II, upon adaptation to dietary CLA supplementation, ADG increased quadratically (603, 623, 622, and 548 g/d; P < 0.01), ADFI decreased linearly (873, 840, 867, and 717 g/d; P < 0.02) and gain:feed ratio tended to increase linearly (691, 742, 715, and 763; P < 0.07). In Phase III, no differences in growth performance were attributed to either dietary or environmental treatments. The poor health status associated with the dirty environment induced a growth suppression; pigs in the clean room had a greater cumulative ADG (P < 0.01) and ADFI (P < 0.01) than pigs in the dirty room. In Phase I, lower plasma urea nitrogen levels observed in pigs found in the dirty room (P < 0.03) indicated a lower protein intake caused by a lower ADFI. The effects of dietary CLA on peripheral phenotypic profiles of lymphoytes did not appear until d 42. However, as indicated by the growth suppression of pigs in the dirty room, the negative effects of the environmental challenge on pig health and growth had already appeared during phase I. On d 42, CLA induced a linear increase in percentages of CD8+ lymphocytes (21.7, 22.3, 28.0, and 32.7%; P < 0.001). These data suggest that a 42-d dietary CLA supplementation preceding a disease challenge could have prevented disease associated growth suppression. Also, CLA-mediated amelioration of particular infectious diseases will depend on which CD8+ T cell subset (i.e., CD8alphaalpha immunoregulatory or CD8alphabeta-cytotoxic) is most influenced by dietary CLA supplementation. PMID- 11263833 TI - Effects of betaine on growth, carcass characteristics, pork quality, and plasma metabolites of finishing pigs. AB - An experiment was conducted to determine the effect of dietary betaine (0, 0.125, 0.250, or 0.500%) on growth, carcass traits, pork quality, plasma metabolites, and tissue betaine concentrations of cross-bred finishing pigs. Four replications of three pigs (two barrows and one gilt) each were used for each treatment. The basal diet contained 0.85 (69 to 88 kg BW) or 0.65% Lys (88 to 115 kg BW). Overall ADG and gain:feed were not affected (P > 0.10) by betaine, but overall ADFI was decreased (quadratic, P < 0.05; 0 vs betaine, P < 0.01) by betaine; pigs fed 0.250% betaine had the lowest ADFI. Loin muscle area, average back-fat, dressing percentage, percentage lean, total fat, lean:fat, and leaf fat weight were not affected (P > 0.10) by betaine. Tenth-rib backfat thickness was decreased (quadratic, P < 0.05; 0 vs betaine, P < 0.05); pigs fed 0.250% betaine had the lowest 10th-rib backfat thickness. Carcass length was increased (linear, P < 0.05; 0 vs betaine, P < 0.10) as the level of betaine was increased. Fat-free lean, lean gain per day, ham weight, ham fat-free lean, and ham percentage lean were increased (quadratic, P < 0.10), but percentage fat, total ham fat, percentage ham fat, and butt-fat thickness were decreased (quadratic, P < 0.10); these traits were respectively highest or lowest in pigs fed 0.250% betaine. Thaw loss and 24-h pH were increased (quadratic, P < 0.10; 0 vs betaine, P < 0.05) and cook loss was decreased (linear, P < 0.05) in pigs fed betaine. The CIE L* value for the biceps femoris was decreased (quadratic, P < 0.10; 0 vs betaine, P < 0.10); pigs fed 0.250% betaine had the lowest CIE L* value. Subjective color, firmness-wetness, marbling, percentage moisture and bound water of the loin muscle, and shear force were not affected (P > 0.10) by betaine. Betaine was not detectable (< 0.07 mg/g) in the loin muscle of pigs fed 0% betaine, but betaine was detectable and relatively constant in pigs fed 0.125, 0.250, or 0.500% betaine (0.22, 0.17, and 0.21 mg/g, respectively). Plasma urea N, total protein, albumin, triglycerides, and HDL cholesterol concentrations were not affected (P > 0.10). Plasma total cholesterol (linear, P < 0.10) and NEFA (quadratic, P < 0.10) were increased in pigs fed betaine. Betaine improved carcass traits when provided at 0.250% of the diet and improved some aspects of pork quality. PMID- 11263834 TI - Transport stress modulates adrenocorticotropin secretion from peripheral bovine lymphocytes. AB - The influence of transport stress on the secretion of adrenocorticotropin (ACTH) from peripheral bovine lymphocytes was evaluated by exposing cows to short- (30 min) or long-term (14 h) transport. After transporting animals for 14 h they were given a mandatory rest for 24 h in two different situations, either by off loading them and allowing them to rest in a stall or by keeping them in the truck. Blood samples were withdrawn before and after transport and after the rest period in long-term transported cows and before and after transport in cows transported for 30 min. Peripheral blood lymphocytes were separated and cultured for 72 h in serum-free medium. Adrenocorticotropin was measured using highly sensitive and specific immunoradiometric assay in culture supernates. We noticed no effect of short-term transport on ACTH secretion from lymphocytes. The ACTH concentration in animals transported for 14 h increased (P < or = 0.01) from 4.72 +/- 0.48 pg x mL(-1)/2 x 10(6) lymphocytes before the transport to 8.24 +/- 1.40 pg x mL(-1) directly after the transport. When animals were off-loaded and rested in a stall for 24 h, ACTH secretion from cultured lymphocytes returned to the basal value of 4.24 +/- 0.31 pg x mL(-1), whereas the animals rested in the truck had ACTH levels of 8.9 +/- 1.43 pg x mL(-1). Phytohemagglutinin, a plant lectin that stimulates lymphocytes, did not affect the lymphocytic ACTH secretion in this study. Heart rate and rectal temperature measured telemetrically increased in cows directly after 14 h of transport but decreased to pretransport values in cows rested for 24 h in cows rested in stalls and those rested inside the truck. This experiment is the first to show lymphocytic ACTH secretion in cows, and the results indicate that ACTH secretion from peripheral lymphocytes could be used as a reliable measurement in stress studies. PMID- 11263835 TI - Pituitary hormone and insulin responses to infusion of amino acids and N-methyl D,L-aspartate in horses. AB - Thirty-nine adult light horse mares, geldings, and stallions were used in two experiments to assess the pituitary hormone and insulin responses to infusions of arginine, aspartic acid, lysine, glutamic acid, and N-methyl-D,L-aspartate (NMA). In Exp. 1, 27 horses were assigned to one of three infusion treatments: 1) physiological saline (1 L); 2) 2.855 mmol of arginine/kg BW in 1 L of water; or 3) 2.855 mmol of aspartic acid/kg BW in 1 L of water. In Exp. 2, 12 horses were assigned, in a multiple-square 4 x 4 Latin square design, to one of four infusion treatments: 1) 2 mL of saline/kg BW; 2) 2.855 mmol of lysine/kg BW in water; 3) 2.855 mmol of glutamic acid/kg BW in water; or 4) 1 mg of NMA/kg BW in water. In Exp. 1, an acute (within 20 min) release of growth hormone (GH) was induced (P = 0.002) by aspartic acid. In contrast, acute release of prolactin (P = 0.001) and insulin (P = 0.002) was induced only by arginine; moreover, the arginine effect on insulin was present only in mares (P = 0.011). In Exp. 2, an acute release of GH was induced (P = 0.001) by glutamic acid and NMA. In males, the glutamic acid induced GH release was greater than that of NMA; in mares, the NMA-induced GH release was greater than that of glutamic acid (P = 0.069). Both lysine and glutamic acid induced (P = 0.001) acute release of prolactin, whereas an acute release of insulin was elicited (P = 0.002) only by lysine. The NMA-induced LH response was due almost entirely to the response in mares and stallions (P = 0.016), and the NMA-induced FSH release was due almost entirely to the response in mares (reproductive status effect; P = 0.004). In the horse, aspartic acid, glutamic acid, and NMA seem to stimulate GH release; arginine and lysine seem to stimulate prolactin and insulin release; and NMA seems to stimulate LH and FSH release. It seems that N-methyl-D-aspartate glutamate receptors are involved in controlling GH, LH, and FSH secretion, whereas other mechanisms are involved with prolactin secretion. These results also indicate that gonadal steroids interact with amino acid-induced pituitary hormone release in adult horses. PMID- 11263836 TI - Influence of pasture sward height and concentrate supplementation on intake, digestibility, and grazing time of lactating beef cows. AB - To establish the effect of sward height (SH) and concentrate supplementation on performance of grazing cattle, 24 crossbred Angus beef cows (535 kg BW) and calves (114 kg BW) were grouped by weight and calving date. They were randomly assigned to two SH treatments, either 4 to 8 cm or 8 to 11 cm, and fed three levels of supplement, high, low, or none, consisting of 6.24, 3.12, and 0 kg x animal(-1) x d(-1), respectively. The experiment was repeated over three 15-d periods in 1996: May (P1), June/July (P2), and August (P3). No SH x supplement level x period or SH x supplement level interactions (P > 0.10) were evident for responses tested. Cows on lower SH had greater (P < 0.08) DMI but spent an additional 1.3 h/d (P < 0.01) grazing compared with cows on higher SH. Sward height had no influence (P > 0.10) on forage DM digestibility (DMD). Forage DMI, DMD, and grazing time (GT) decreased (P < 0.05) as supplementation increased. Nonetheless, supplemented cows consumed more total DMI (P < 0.08) than unsupplemented cows. Cows consumed 2.4 kg/d more forage DM (P < 0.01) in P1 and P2 than in P3. Cows grazed 1.3 h/d (P < 0.01) less in P1 than in P2 and P3. Grazing efficiency (DMI/h GT) declined as supplementation increased and grazing season advanced to P3 (P < 0.01). Decreased forage DMI and grazing efficiency with increasing supplementation suggests that supplemented cattle should be able to maintain productivity while grazing at SH lower than unsupplemented cattle. PMID- 11263837 TI - Optimization of rate and efficiency of dietary nitrogen utilization through the use of animal by-products and(or) urea and their effects on nutrient digestion in Holstein steers. AB - The objective of this N balance study was to determine the potential for improving the efficiency and rate of dietary N utilization in Holstein steers by feeding an amino acid-balanced mixture of animal by-product protein sources in combination with urea. The Beef NRC 1996 Model Level 2 was used to formulate a corn-based (86:14 concentrate-hay) control diet with soybean meal as the primary N supplement that would provide ME and metabolizable protein (MP) allowable ADG of 1.4 kg in 250-kg steers with an estrogenic implant and fed an ionophore. A combination of porcine meat and bone meal, fish meal, hydrolyzed feather meal, and blood meal was also formulated as an undegradable intake protein (UIP) blend to complement those amino acids (AA) derived from microbial protein synthesis. Four steers with an average initial BW of 259 kg were assigned in a 4 x 4 Latin square design to treatments consisting of control, two levels of UIP inclusion (2.6 and 5.2%; DM basis) in combination with urea, and a negative control "urea diet" containing no UIP and no SBM. The steers were fed at hourly intervals 95% of ad libitum intake and were injected with 500 microg of estradiol-17beta twice daily. Nitrogen intakes were 155, 160, 162, and 145 g/d, and N balances were 47, 51, 42, and 47 g/d when the 0, 2.6, 5.2% UIP and the urea diets were fed, respectively. Nitrogen balance was reduced with the 5.2% UIP diet (P < 0.05), and was less than the capacity estimate derived from abosmasal casein infusion studies. Apparent N digestibilities averaged 69%, but DM, OM, and nonstructural carbohydrate digestibilities were significantly reduced for the urea diet. Feeding 5.2% UIP in the diet reduced (P < 0.05) the biological value from 46 to 38%, which was accompanied by a significant elevation of plasma urea N. Results indicate that genetic capacity for N retention was approximately 51 g/d. Results demonstrate that use of an AA-balanced blend of animal by-product protein sources did not improve the efficiency of dietary N usage when added to corn-based diets formulated with the Beef NRC 1996 Model Level 2 to meet nutrient requirements of rapidly growing steers. Using urea as the only N supplement achieved equal rate and efficiency of N use. PMID- 11263838 TI - Technical note: use of laser diffraction for particle size distribution measurements in duodenal digesta. AB - Duodenal samples from three lactating cows were used to measure the particle size distribution by wet sieving and by laser diffraction (LD). The median particle size was calculated in the size range where the methods overlapped, and the median particle size was not different between the methods (P = 0.98). The particle size data were also fitted to a log normal distribution function. The logarithmic mean particle size (log10mean) tended to be larger for wet sieving (P = 0.06), and the logarithmic standard deviation (log10SD) was similar for the two methods (P = 0.32). In addition, duodenal samples from the same three animals were fractionated into five different size classes by wet sieving. Particle size distributions were measured by LD and by microscopic image analysis (MIA) for each of the size classes. The log10mean particle size calculated by LD was inside the sieve size range of the four smallest size classes. Log10mean particle size calculated by LD was smaller than log10mean calculated by MIA for two size classes (106 to 300 microm and 300 to 600 microm), and LD gave generally broader particle size distributions than MIA. This study showed that duodenal particle size distributions measured by LD and wet sieving were similar and LD is therefore an alternative method for particle size distribution measurements in postruminal digesta. PMID- 11263839 TI - Soybean hulls as a primary ingredient in forage-free diets for limit-fed growing cattle. AB - In Exp. 1, 300 heifers (260 kg initial BW) were used to compare growth performance of cattle fed forage-free diets containing predominantly soybean hulls with that of cattle receiving roughage- and corn-based diets and to determine whether cattle fed soybean hull-based diets would respond to supplementation with methionine hydroxy analogue (MHA), lipid-coated betaine, or concentrated separator by-product (CSB; a source of betaine). Treatments included 1) a roughage-based diet fed at 2.75% of BW, 2) a corn-based diet fed at 1.5% of BW, 3) a corn-based diet fed at 2.25% of BW, 4) a soybean hull-based diet fed at 1.5% of BW (SH1.5), 5) a soybean hull-based diet fed at 2.25% of BW (SH2.25), 6) SH1.5 top-dressed with 11.4 g/d Alimet (10 g/d MHA), 7) SH2.25 top-dressed with 11.4 g/d Alimet, 8) SH2.25 top-dressed with 7 g/d of a lipid-coated betaine product (4.2 g/d betaine), and 9) SH2.25 top-dressed with 250 g/d CSB (15.5 g/d betaine). Supplemental MHA, betaine, and CSB did not change DMI, ADG, or gain:feed ratio for cattle fed soybean hulls. Heifers fed soybean hull-based diets gained 29% slower (P < 0.05) and had 27% lower gain:feed ratios than heifers fed the corn-based diets. Cattle fed soybean hull-based diets had gains that were lower (P < 0.05) than those of cattle fed the roughage-based diets, but gain:feed ratios were similar because cattle were fed less of the soybean hull based diets. Roughage-fed cattle had similar gains but 25% lower (P < 0.05) gain:feed ratios than cattle fed the corn-based diets. In Exp. 2, degradation by ruminal microbes of betaine in anhydrous betaine, betaine-HCl, feed-grade betaine, lipid-coated betaine, and CSB was evaluated in vitro using ruminal inocula collected from steers fed a high-grain or high-roughage diet. The roughage diet led to less betaine disappearance than the grain diet. More betaine was degraded from CSB than from other sources, perhaps because sugars provided by CSB stimulated fermentation, but no large differences occurred among the other four sources. Betaine from all sources was extensively degraded, although some betaine may escape ruminal degradation. PMID- 11263840 TI - Rapid communication: sequence of a quail calpain cDNA. PMID- 11263841 TI - Rapid communication: mapping of oxytocin (OXT) to the central region of bovine chromosome 13 by linkage analysis using SSCP. PMID- 11263842 TI - Rapid communication: three new allelic sequences at the ovine MHC class II DQA1 locus. PMID- 11263843 TI - Rapid communication: nucleotide sequence of the porcine beta3-adrenergic receptor gene. PMID- 11263844 TI - What are 'tissue ACE inhibitors,' and should they be used instead of other ACE inhibitors? PMID- 11263846 TI - A 46-year-old man with dyspnea. PMID- 11263845 TI - Can angiotensin II receptor blockers be used in patients who have developed a cough or angioedema as a result of taking an ACE inhibitor? PMID- 11263847 TI - Making the most of cholesterol-lowering margarines. AB - Used as a substitute for normal dietary intake of saturated fatty acids, margarines containing plant sterols can cause a modest reduction in serum total cholesterol and low-density lipoprotein cholesterol levels. They have been shown effective in patients with mild hypercholesterolemia, but they are also useful in the general population. PMID- 11263848 TI - A 35-year-old man with recurrent aseptic meningitis. PMID- 11263849 TI - Phenylpropanolamine and stroke: the study, the FDA ruling, the implications. AB - Following a recent case-control study that linked the use of phenylpropanolamine (PPA) in diet aids to the risk of hemorrhagic stroke, the Food and Drug Administration requested that drug companies stop marketing products that contain PPA. Dozens of over-the-counter and prescription diet aids and cough and cold remedies will need to be reformulated or discontinued. This paper reviews the study and its implications for physicians. PMID- 11263850 TI - The challenge of irritable bowel syndrome: creating an alliance between patient and physician. AB - The most important component of the treatment of irritable bowel syndrome (IBS) is to establish a therapeutic physician-patient relationship, coupled with patient education. We describe a stepwise approach to management, including judicious use of invasive tests, and setting realistic treatment goals that address the dominant symptoms, their severity, and psychosocial factors. PMID- 11263851 TI - Evolving cardiovascular applications for magnetic resonance imaging. AB - Improvements in magnetic resonance imaging (MRI) have increased its value in existing cardiovascular applications and opened the door to new uses. The image quality and spatial resolution of cardiovascular MRI is better than that of most other noninvasive imaging procedures. In addition, MRI can measure and map biochemical markers diminished by ischemic damage. PMID- 11263852 TI - Managing menopause after breast cancer: balancing risks and benefits. AB - Many women now survive breast cancer, but find themselves at increased risk of menopausal complications. How to manage menopause after breast cancer is a complex issue, given that estrogen has a role in the development of breast cancer and valid concerns exist about estrogen replacement therapy in patients who have had breast cancer. This article explores the relationship between estrogens and breast cancer and discusses management options for a variety of menopausal complications in breast cancer survivors. PMID- 11263853 TI - Management of asymptomatic left ventricular dysfunction. AB - Asymptomatic left ventricular dysfunction should be treated as an early stage on the continuum that is chronic heart failure. The author presents the clinical trial data on which current management with angiotensin-converting enzyme inhibitors and beta-blockers is based. Issues surrounding screening are also discussed. PMID- 11263854 TI - Shedding: how to manage a common cause of hair loss. AB - Although telogen effluvium, or shedding-the most common type of diffuse hair loss in both women and men-is usually self-limiting, the condition may become chronic if the trigger is not identified and corrected. The authors discuss the physiologic and emotional triggers, clinical presentation, diagnosis, and management strategies, including the importance of patient education and reassurance. PMID- 11263855 TI - Reliability and limitations of cytochemistry in diagnosis of acute myeloid leukemia. Minireview. AB - Accurate characterization of leukemic blast cells is an important prerequisite of the precise diagnosis of acute myeloid leukemia and has a great impact on therapy and prognosis. The purpose of this review is to consider the present possibilities and limitations of enzyme cytochemistry and to emphasize how cytochemistry may contribute to the final classification and differential diagnosis of acute myeloid leukemia. The role of conventional enzyme cytochemistry, either dominant or subsidiary, in the discrimination of acute myeloid leukemia subgroups is discussed. The survey confirms the necessity of immunological marker analysis in the accurate diagnosis of minimally differentiated myeloid leukemias and acute leukemia of megakaryocytic lineage. In these cases, the cytochemical evaluation provides insufficiently relevant information regarding blast cell origin. On the other hand, cytochemical investigation is appreciated to be dominant over immunophenotyping in characterizing majority of acute myeloid leukemia subgroups, because of the availability of standardized and sufficiently specific cytochemical reactions and, because of the lack of specificity of the many of immunological markers against myeloid antigens. The immunocytochemical, cytogenetic, molecular biology and electron microscopic studies shortly mentioned in this review supplement the information for correct diagnosis of acute myeloid leukemia. PMID- 11263857 TI - Expression of the non-classical HLA-G antigen in tumor cell lines is extremely restricted. AB - It has been proposed that tumor cells frequently associated with partial or total loss of HLA class Ia expression may abnormally express HLA-G class Ib antigen. Such peculiar HLA class I expression would allow tumor cells to escape not only from CD8+T but also from NK-cell cytotoxicity. We studied the cell surface expression of HLA-G using flow cytometry with two HLA-G specific monoclonal antibodies (87G, 01G). The JEG-3 choriocarcinoma cell line, which constitutively expresses HLA-G antigens was used as a positive control. We did not detect the cell-surface HLA-G antigens in the following 75 tumor cell lines: melanoma (22), neuroblastoma (7), retinoblastoma (1), glioma (2), breast carcinoma (3), ovarian carcinoma (3), cervical carcinoma (1), colon carcinoma (3), bladder carcinoma (2), hepatocarcinoma (1), sarcoma (2) and leukemia cell lines: T-lymphocytes (6), B-lymphocytes (13) and myelo-monocytes (9). We found that some myelomonocytic cell lines express on their surface high affinity FcgammaRI (CD64) that may result in the binding of HLA-G specific mabs to their cell surface even in the absence of HLA-G molecules. Our panel of HLA-G negative tumor cell lines accommodated 62 cell lines for which similar analysis have not been reported and also contained 13 cell lines with total or partial loss of HLA class Ia molecules. Our observation imply that under normal culture conditions the cell surface HLA-G reactive with 87G and 01G mabs is absent in most tumor cell lines of different origin. PMID- 11263858 TI - Oxidative/antioxidative effects of different lignin preparations on DNA in hamster V79 cells. AB - Endogenous oxidative damage to DNA is thought to be an important etiologic factor in the development of chronic diseases such as cancer. Many products of the vegetable kingdom have been suggested to limit oxidative damage to DNA in humans. To this group belong lignins, polyphenols present in all plants (including edible plants). The aim of this study was to examine oxidative/antioxidative effects of different lignin preparations on mammalian DNA. In addition to a water-soluble sulfur-free lignin 1 which was obtained by fractionation of hardwood hydrolysate, we investigated lignin 2 (obtained by oxidation of lignin 1), lignin 3 (prepared by the extraction of lignin 2 with a mixture ethanol-water 3:1), lignin 4 (Na salt of lignin 3) and lignin 5 (prepared by extraction of lignin 2 with diethylether). Our results showed that only the original lignin 1 did not increase substantially the level of DNA damage. Lignins 2, 3, 4 and 5 increased both the level of frank DNA strand breaks + alkali-labile sites and the level of FPG-sensitive sites representing oxidative damage to DNA. Lignin 1 was further tested for its antioxidative activity against DNA base modifications generated by visible light+photosensitizer. Obtained results confirmed the oxygen species scavenging activity of lignin 1. PMID- 11263856 TI - A double germline mutations in the APC and p53 genes. AB - Germline mutation in the APC gene is required for the initiation of the development of familial adenomatous polyposis (FAP). According to Fearon and Vogelstein model, further somatic mutations in the K-ras oncogene, DCC gene and p53 tumor suppressor gene are prerequisite for development of colon carcinoma. We have found that the germline mutations in the DNA isolated from lymphocytes of an 18 years old girl with extraordinary expressive phenotype in codons 1060-1061 of the APC gene result in truncation of the APC protein. The mutation in codons 12 and 13 of the K-ras oncogene was not detected, but another germline mutation was found in codon 210 of the p53 gene. Furthermore, no one of these germline mutations was detected in the DNA of peripheral blood lymphocytes of the patient's 21 years old healthy sister. Until now, there has been no evidence about the expressive phenotype due to mutation in codons 1060-1061 of the APC gene; the role of germline missense mutation in codon 210 of the p53 gene in the FAP malignant process remains to be elucidated too. The effect of the combination of germline mutation in two different tumor suppressor genes in the progress of disease is discussed. PMID- 11263859 TI - Failure of carboxymethylglucan to inhibit oxidative DNA damage induced by hydroxyl radicals or singlet oxygen. AB - The ability of carboxymethylglucan (CMG), a high molecular water-soluble derivative of glucan, was evaluated to act as a scavenger of reactive oxygen species. Hydrogen peroxide and methylene blue plus visible light, well-defined oxidant factors, were used as a model agents for induction ofoxidative DNA damage in CaCo-2 cells. Both hydrogen peroxide and visible light gave rise to dose dependent increase of DNA damage mediated by hydroxyl radicals (*OH) or singlet oxygen (1O2), respectively. While DNA lesions generated by hydrogen peroxide dominated by strand breakage, exposure of CaCo-2 cells to visible light led mainly to base modifications sensitive to formamidopyrimidine DNA-glycosylase (Fpg). Neither CMG nor ascorbic acid, a known antioxidant, induced any DNA damage in CaCo-2 cells. Pretreatment of cells with ascorbic acid prior to H2O2 or visible light exposure resulted into statistically significant reduction of DNA lesions induced by particular agent. However, pretreatment of CaCo-2 cells with CMG in concentration range from 0.01 microM to 1 microM reduced neither the level of strand breaks induced by hydrogen peroxide nor the number of Fpg-sensitive base modifications generated by visible light. PMID- 11263860 TI - Co-expression of GFAP, vimentin and cytokeratins in GL-15 glioblastoma cell line. AB - Glial fibrillary acidic protein (GFAP), vimentin (Vi) and cytokeratin (CK) intermediate filament (IF) proteins were studied in glioblastoma cell line GL-15. The immunofluorescence staining revealed strong positive staining for vimentin in all cultured cells. Approximately 20% of analyzed cells showed strong and 50% moderate intensity of staining for GFAP. About 3% of all cells were positively stained with a mixture of anti-CK monoclonal antibodies. The expression of all IF was not in relation to the cell density or days in vitro after passage. The double immunofluoresce revealed that all CK-positive cells express GFAP and vimentin. This study demonstrates the heterogeneity of the clonal GL-15 glioma cell line which consists in three immunocytochemically distinct cell types: Vi+/GFAP-/CK-, Vi+/GFAP+/CK-, and Vi+/GFAP+/CK+. These findings give further evidence about the expression of non-glial IF in cultured glioma cells. PMID- 11263861 TI - Radiation-induced DNA damage and repair evaluated with 'comet assay' in human ovarian carcinoma cell lines with different radiosensitivities. AB - Radiation-induced DNA damage and kinetics of DNA repair was evaluated in three human ovarian carcinoma cell lines (i.e. CH-1, A-2780 and SKOV-3) with different sensitivities to ionizing radiation and radiation-induced apoptosis with the aid of single cell gel electrophoresis (SCGE, the comet assay). A good correlation was found between the initial level of DNA breaks and radiation induced apoptosis in CH-1 and SKOV-3 cell lines. While the radiation-sensitive CH-1 cell line manifested the highest level of initial DNA breakage and a significant delay in DNA break rejoining, the inverse correlation was found in the radiation-resistant cell line SKOV-3. Intermediate initial level of breaks was induced in the A-2780 cell line characterized by the intermediate sensitivity to X-ray radiation in comparison to CH-1 and SKOV-3 cells, however, the kinetics of DNA repair was comparable with radiation-resistant cell line SKOV-3. Our data suggest that the comet assay could be a promising tool for prediction of intrinsic cell radiosensitivity. This method might be considered as a supplementary technique to the more reliable but time consuming clonogenic assay. PMID- 11263862 TI - Mismatch repair in xenopus egg extracts is not strand-directed by DNA methylation. AB - The efficiency of Xenopus laevis egg extract to repair T:G and A:C mismatched base pairs in unmethylated, hemimethylated and fullymethylated heteroduplexes was investigated. Filamentous phage M13mp18 and its derivative M13mp18/MP-1 (C changed to T inside the sequence dCC*C GGG, at the position 6248) were used for heteroduplexes construction. The three origins of mismatched base-pairs in the eukaryotic DNA are mimicked by in vitro methylation: hemimethylated DNA (me-/me+) for replication errors; unmethylated (me-/me-) and fully methylated DNA (me+/me+) for recombination heteroduplexes, and fullymethylated also for locally, spontaneously deaminated 5-methylcytosine (5meC) to T, generating the exclusively T:G mismatch. The methylations were in CpG dinucleotides, mostly characteristic ofeukaryotic cells [5, 24]. In vitro methylation was done by HpaII methylase which methylate central C of dCCGG sequence in the manner of eukaryotic methylation. The position of mismatched bases was chosen so that correction of mismatched bases in any strand would create the sequence for one of the "diagnostic" restriction endonucleases, either BstNI or MspI. Correction efficiency was about 10(8) repair events per egg equivalent. Correction in favor of C:G base pair restoration occurred regardless of the T:G or C:A mispairs, with almost equal efficiency. Repair of T:G to T:A was up to 10 times less efficient comparing to C:G, and repair of C:A to T:A was in our experimental system undetectable. No significant difference in repair efficiency of mismatched bases situated in unmethylated, hemimethylated or fullymethylated heteroduplexes indicate methylation-independent repair of mismatched bases in X. laevis oocite extracts. PMID- 11263863 TI - Correlation of clinical picture (event free survival and overall survival) in childhood acute leukemia patients with immunophenotype and chromosomal abnormalities. AB - In a group of 102 children with different immunological subtypes of acute leukemia, both lymphoblastic and nonlymphoblastic, the clinical parameters - event free survival and overall survival were correlated with numerical and structural chromosomal abnormalities. In a group of 80 ALL patients genetic abnormalities were observed in 40 patients, from those 19 of numerical type, 17 of structural type and 4 with both, numerical and structural anomalies. From the whole ALL group observed 23 patients (28.75%) died. In 10 died patients genetic abnormalities were found and in 6 cases less mature T-phenotype ALL has been documented. It seems, therefore, that immature T-phenotype with pathological karyotypes of all types of genetic anomalies presents the most risk group of patients of which all children died. ALL patients, as a whole, with pathological karyotype have shown significantly lower event free survival rate, comparing to the group of ALL patients with normal karyotype. Overall survival rate was also lower in the first group, but statistically not significant. In T-ALL patients, in both groups, with and without pathological karyotype, event free survival rate and overall survival rate were also lower in the first group, but statistically not significant. In B-ALL patients with pathological karyotypes vs. normal ones overall survival rate was lower in the first group, but statistically not significant. There was no difference in overall survival rate in these patients between pathological and normal karyotypes. In ANNL group of patients pathological karyotype was observed in 14 of them, with numerical anomalies in 6 patients, structural in 4 patients and both of them - numerical + structural in 4 children. From the whole ANLL group observed 11 (50%) patients died during the follow-up period (9 in relapse and 2 of treatment complications). From 11 died patients in 81.8% pathological karyotype was present. The prevalence of pathological karyotypes was observed in less mature M0-M2 ANLL subtype (71.4%). ANLL patients with pathological karyotype have shown significantly lower event free survival rate (in one of the two statistical log-rank analyses), comparing to the group of ANLL patients with normal karyotype. Overall survival rate was also lower in the first group, but statistically not significant. The presence/absence of CD34 marker expression in blast cells of our group of acute leukemia patients did not show any difference in event free survival and overall survival rates. PMID- 11263864 TI - Diazenes as modificators of drug-resistance in tumor cells. AB - To overcome the drug resistance, which is the major obstacle in the successful treatment of cancer patients, various compounds have been tested. Glutathione is one of the most promising targets for modulation. In the present study, we examined the influence of five new synthesized compounds--diazenes on the reduction of the intracellular level of GSH. Further, we investigated their ability to increase the cytotoxicity of cisplatin, vincristine and doxorubicin. In experiments human parental cervical (HeLa) and laryngeal (HEp2) carcinoma cells and their drug-resistant cell sublines (HeLaCA and CK2, respectively) were used. Intracellular GSH content was examined spectrophotometrically by the procedure developed by Tietze. The cell sensitivity to drugs was determined using a modified colorimetric MTT assay. Results showed that the rate of reduction of GSH concentration was dependent on the cell type and the type of diazenes. We did not find a correlation between the reduction in GSH level and increased cytotoxicity to selected anticancer drugs. Nevertheless, we found that: a) diazenes LV-35 and VZ-19 increased the cytotoxicity of cisplatin in HEp2 cells, b) diazene MG-19 potentiated the cytotoxicity of vincristine in HEp2 cells, and c) diazene VZ-19 in HeLaCA cells. These data suggest that specific combination of diazene and anticancer drug may be useful in the treatment of certain tumor types. PMID- 11263865 TI - Quality of life in cancer patients treated by chemotherapy. AB - Many studies connected with different aspects of the quality of life (QL) have been increasingly reported. In this study we have been used FACT questionnaire to estimate QL in three groups of cancer patients receiving chemotherapy. Questionnaires were completed by 177 patients. Adverse effects of treatment severely influence the cancer patients QL. The most significant worsening of QL was noticed in the group of lung cancer patients receiving the most emetogenic chemotherapy(i.e. cis-platinum, vepesid). In other two groups (gastric and colorectal cancer patients) side effects of chemotherapy caused relatively less QL deterioration. This finding implicates possibility of intensifying the course of treatment (dosage and duration) assuming there is no hematopoetic insufficiency. PMID- 11263866 TI - CD44 and its v6 spliced variant in lung carcinomas: relation to NCAM, CEA, EMA and UP1 and prognostic significance. AB - CD44 is a polymorphic family of cell surface glycoproteins that was recently reported to have important role in cell adhesion and migration as well as modulation of cell-matrix interactions. Thus, expression of CD44 has been proposed to be associated with malignant behavior of tumors like invasive growth and formation of metastasis. The expression of CD44s and its v6 isoform (CD44v6) was determined immunohistochemically in 106 lung tumors of various histophenotypes, degrees of differentiation, and clinical stages. The results were compared with the expression of NCAM, CEA, EMA and UP1 and with clinicopathological parameters including patients' survival. CD44s was expressed in all histophenotypes of non-small cell lung carcinomas (NSCLC) with tendency being squamous cell lung carcinoma (SqCC) > bronchioloalveolar adenocarcinoma (BAC) > conventional adenocarcinoma (ConAC) (91, 66.7 and 38.9%, respectively). Almost identical distribution of positivity revealed CD44v6 in all three subgroups of NSCLC mentioned above (91, 66.7 and 36.1%, respectively). In the subgroup of neuroendocrine tumors, CD44s and CD44v6 were restrictedly expressed in small cell lung carcinomas (2/14 tumors), while all 3 typical carcinoids were strongly positive for these markers. Expression of NCAM and CEA was significantly higher in adenocarcinoma subgroup than those in SqCC subgroup (45.7 and 75% vs. 14.8 and 39%, respectively). NCAM expression was also significantly different in BACs and in ConACs (69.2 vs. 36.4%, p < 0.05). The expression of CD44 was related to the differentiation of SqCC. The carcinomas with keratinization were CD44 positive. Adenocarcinomas producing mucin were CD44 negative. The expression of CD44, NCAM, CEA, EMA and UP1 did not correlate with lymph node metastasis and disease stage. CD44V6 was the only marker that its expression was closely related to patients' survival. The absence CD44v6 but not CD44s in NSCLC group was associated with significantly longer survival of patients compared to patients with CD44v6 positive tumors. This difference was even higher in tumors negative for CD44v6 and simultaneously NCAM and/or CEA positive. The data of this study suggest that CD44v6 might be an independent prognostic factor in NSCLC. Moreover, our data give another evidence of diverse role of CD44 in the differentiation and progression of non-small cell lung carcinomas and neuroendocrine carcinomas of the lung. PMID- 11263867 TI - Colorectal carcinoid tumors--own experience. AB - Symptomatology, diagnostics and treatment problems in 5 patients with colorectal carcinoid are presented. From 1974 to 1999 in the Clinic of Endocrinological and General Surgery of the Medical University of Lodz, 3001 patients underwent surgery due to acute appendicitis and 431 for colorectal cancer. Among them, there were 5 patients in whom the histological examination revealed colorectal carcinoid. The carcinoids were localized in the appendix in 4 patients and in the left colon flexure in 1 patient. The mean age of these 5 carcinoid patients at the time of diagnosis was 38.4 years (range 18-72 yr). The female-to-male ratio amounted to 4:1. The symptoms of all 5 patients was not typical for carcinoid of the colon. In four surgery was performed for acute appendicitis and one patient complained of chronic obstipation and pain in the left epi- and mesogastrium. The double-contrast examination of the large intestine revealed tumor of the left colon flexure. Four carcinoid patients with the signs of acute appendicitis had emergency surgery. The carcinoid tumors were diagnosed microscopically after surgery only. In 3 of them the tumor extended beyond the appendix and a reoperation was performed. In one patient with the tumor of small diameter (5 mm) involving only the mucosa and submucosa a reoperation was not indicated. In 3 reoperated patients right hemicolectomy with regional lymphadenectomy was performed. The patient with the tumor of the left colon flexure diagnosed preoperatively underwent radical surgery with regional lymphadenectomy. The postoperative histological examination of the tumor confirmed carcinoid. No carcinoid metastases were found in lymph nodes of all studied cases. Until today, all 5 carcinoid patients are alive with no signs of local reccurrence or distant metastases over the 1-20 year follow-up period. PMID- 11263868 TI - Recent highlights in hemicarcerand chemistry. AB - Since Donald J. Cram's first synthesis of a carcerand, which permanently entrapped a single guest molecule, many other carcerands and hemicarcerands have been synthesized and studied. Slowly, we begin to understand the role of the template in the formation of hemicarceplexes and carceplexes, why some hemicarceplexes are more stable than others, and how guests enter and exit the inner phases of molecular containers. In this Account we discuss our new insight in the chemistry of molecular containers, as well as recent highlights in through shell chemistry and in the inner-phase stabilization of reactive intermediates. PMID- 11263869 TI - Metric engineering of soft molecular host frameworks. AB - The self-assembly and solid-state structures of host-guest inclusion compounds with lamellar architectures based on a common building block, a resilient hydrogen-bonded sheet consisting of guanidinium ions and sulfonate moieties of organodisulfonate "pillars", are described. The pillars connect adjacent sheets to generate galleries with molecular-scale cavities occupied by guest molecules. The size, shape, and physicochemical character of the inclusion cavities can be systematically adjusted by interchanging framework components while maintaining the lamellar architecture, enabling prediction and control of crystal lattice metrics with a precision that is unusual for "crystal engineering". The reliability of the lamellar architecture is a direct consequence of conformational flexibility exhibited by these hosts that, unlike rigid systems, enables them to achieve optimal packing with guest molecules. The adaptability of these hosts is further reflected by an architectural isomerism that is driven by guest templating during assembly of the inclusion compounds. Host frameworks constructed with various pillars display metric interdependences among specific structural features that reveal a common mechanism by which these soft frameworks adapt to different guests. This unique feature facilitates structure prediction and provides guidance for the design of inclusion compounds based on these hosts. PMID- 11263870 TI - Copper delivery by metallochaperone proteins. AB - Copper is an essential element in all living organisms, serving as a cofactor for many important proteins and enzymes. Metallochaperone proteins deliver copper ions to specific physiological partners by direct protein-protein interactions. The Atx1-like chaperones transfer copper to intracellular copper transporters, and the CCS chaperones shuttle copper to copper,zinc superoxide dismutase. Crystallographic studies of these two copper chaperone families have provided insights into metal binding and target recognition by metallochaperones and have led to detailed molecular models for the copper transfer mechanism. PMID- 11263871 TI - Substituent effects on the reactivity of the silicon-carbon double bond. AB - Laser flash photolysis of various organosilicon compounds such as aryl-, vinyl-, and alkynyldisilanes, silacyclobutanes and silacyclobutenes, and alpha silylketenes and -diazomethanes leads to the formation of reactive silenes which can be detected directly in solution, allowing detailed studies of the kinetics and mechanisms of their reactions with nucleophiles. Over 30 transient silenes have now been studied by these methods, providing the opportunity to systematically assess the effects of substituents at silicon and carbon on the reactivity of the Si=C bond. PMID- 11263872 TI - Is the CO adduct of myoglobin bent, and does it matter? AB - Early reports of a severely bent CO adduct in myoglobin inspired the idea that heme proteins discriminate against CO, relative to O(2), via steric hindrance imposed by a distal histidine residue. Recent results showing that the bound CO is only slightly distorted do not by themselves overthrow the steric hypothesis, because the steric energy could be stored in displacements of the protein. However, experimental data on site-directed mutants show that the main determinant of ligand affinity changes is the polarity of the binding pocket and that H-bonding by the distal histidine accounts for about 85% of the O(2)/CO discrimination while steric hindrance accounts for the remaining 15%. PMID- 11263873 TI - The crotonase superfamily: divergently related enzymes that catalyze different reactions involving acyl coenzyme a thioesters. AB - Synergistic investigations of the reactions catalyzed by several members of an enzyme superfamily provide a more complete understanding of the relationships between structure and function than is possible from focused studies of a single enzyme alone. The crotonase (or enoyl-CoA hydratase) superfamily is such an example whereby members catalyze a wide range of metabolic reactions but share a common structural solution to a mechanistic problem. Some enzymes in the superfamily have been shown to display dehalogenase, hydratase, and isomerase activities. Others have been implicated in carbon-carbon bond formation and cleavage as well as the hydrolysis of thioesters. While seemingly unrelated mechanistically, the common theme in this superfamily is the need to stabilize an enolate anion intermediate derived from an acyl-CoA substrate. This apparently is accomplished by two structurally conserved peptidic NH groups that provide hydrogen bonds to the carbonyl moieties of the acyl-CoA substrates and form an "oxyanion hole". PMID- 11263874 TI - Dynamics of photoinduced charge transfer and hole transport in synthetic DNA hairpins. AB - The dynamics of photoinduced charge separation and charge recombination processes in synthetic DNA hairpins have been investigated by means of femtosecond transient absorption spectroscopy. The driving force and distance dependence of charge-transfer processes involving singlet acceptors and nucleobase donors are consistent with a single-step superexchange mechanism in which the electronic coupling between the donor and acceptor is strongly distance dependent. The dynamics of reversible hole transport between a primary guanine donor and nearby GG or GGG sequences has also been determined and establishes that these sequences are very shallow hole traps. PMID- 11263875 TI - Model studies for molecular recognition of carbonic anhydrase and carboxypeptidase. AB - Model studies using Zn(2+) complexes of various derivatives of macrocyclic triamines ([12]aneN(3)) and tetraamines (cyclen) have been found to be useful in elucidating and understanding the intrinsic properties of substrate or inhibitor recognition by zinc ions at the active centers of carbonic anhydrase and carboxypeptidase. PMID- 11263876 TI - Dendrimer-encapsulated metal nanoparticles: synthesis, characterization, and applications to catalysis. AB - This Account reports the synthesis and characterization of dendrimer-encapsulated metal nanoparticles and their applications to catalysis. These materials are prepared by sequestering metal ions within dendrimers followed by chemical reduction to yield the corresponding zerovalent metal nanoparticle. The size of such particles depends on the number of metal ions initially loaded into the dendrimer. Intradendrimer hydrogenation and carbon-carbon coupling reactions in water, organic solvents, biphasic fluorous/organic solvents, and supercritical CO2 are also described. PMID- 11263877 TI - Molecular sieve catalysts for the regioselective and shape- selective oxyfunctionalization of alkanes in air. AB - Framework-substituted, molecular-sieve, aluminophosphate, microporous solids are the centerpieces of a new approach to the aerobic oxyfunctionalization of saturated hydrocarbons. The sieves, and the few percent of the Al(III) sites within them that are replaced by catalytically active, transition-metal ions in high oxidation states (Co(III), Mn(III), Fe(III)), are designed so as to allow free access of oxygen in to and out of the interior of these high-area solids. Certain metal-substituted, molecular sieves permit only end-on approach of linear alkanes to the active centers, thereby favoring enhanced reactivity of the terminal methyl groups. By optimizing cage dimension, with respect to that of the hydrocarbon reactant, as well as adjusting the average separation of active centers within a cage, and by choosing the sieve with the appropriate pore aperture, highly selective conversions such as n-hexane to hexanoic acid or adipic acid, and cyclohexane to cyclohexanol, cyclohexanone, or adipic acid, may be effected at low temperature, heterogeneously in air. PMID- 11263878 TI - Is water a friend or foe in organometallic chemistry? The case of group 13 organometallic compounds. AB - This Account summarizes the recent developments in the hydrolysis chemistry of Group 13 trialkyl and triaryl compounds. Emphasis has been placed on the results obtained by us on (a) 1H NMR investigations of controlled hydrolyses of AlMes3 and GaMes3, (b) low-temperature isolation of water adducts of triaryl compounds of aluminum and gallium, (c) synthesis and structural characterization of new polyhedral alumoxanes and galloxanes, and (d) the search for an easy way to synthesize well-defined crystalline methylalumoxanes by deprotonation of the hydroxides with alkyllithium reagents. The systematic studies on the hydrolysis of tBu3Al carried out by Barron et al. are also discussed in order to elucidate the roles of (i) reaction temperature, (ii) solvent medium, and (iii) source of water molecules, in building up hitherto unknown alumoxane clusters. The role of water impurity in organometallic reactions involving a Group 13 alkyl and other ligands (such as silanetriols and phosphorus acids) to build molecular clusters has also been discussed. PMID- 11263879 TI - Experimental strategies for combinatorial and high-throughput materials development. AB - As high-throughput experimental techniques have become common in the area of materials research, entirely new types of experimental strategies have appeared. The kinds of problems, the desired outcomes, and the appropriate patterns are significantly different from those associated with conventional experimentation. Classical experimental design (design of experiments, DOE) strategies grew up in a period of slow, laborious, error-prone experimentation; a modern high throughput laboratory can test more materials in a week than was previously done in a year. The goal of this Account is to identify and critically discuss some of the strategies that are being developed and used in this new, exciting area of research. PMID- 11263880 TI - Behavioral patterns of heterometallic cuboidal derivatives of [M3Q4(H2O)9]4+ (M = Mo, W; Q = S, Se). AB - This Account reports recent progress in the study of some approximately 20 heterometal derivatives of [Mo3S4(H2O)9]4+ with reference also to W and Se analogues. Single cubes (3:1) and corner-shared double cubes (6:1), as well as dimers of the 3:1 single cubes, are considered. A classification of the heterometals as subtypes A, B, and C is introduced. PMID- 11263881 TI - Molecule-mimetic chemistry and mesoscale self-assembly. AB - Molecules are structured aggregates of atoms joined by chemical bonds; crystals are aggregates of molecules, interacting covalently or noncovalently. The work described in this Account uses molecules, crystals, and other forms of atomic/molecular matter to suggest principles that can be used in generating structured aggregates of millimeter-scale components, interacting through capillary interactions. The properties of these aggregates--that is, their "chemistry"--mimic aspects of the chemistry of molecules. PMID- 11263882 TI - New materials in hydrothermal synthesis. AB - In this Account we describe the hydrothermal synthesis of some new materials including microporous crystals, ionic conductors, complex oxides and fluorides, low-dimensional aluminophosphates, inorganic-organic hybrid materials, and particularly condensed materials such as diamond and inorganic helical chains. Hydrothermal synthesis in biology and environment sciences is also introduced. The increasing interest in hydrothemal synthesis derives from its advantages in terms of high reactivity of reactants, easy control of solution or interface reactions, formation of metastable and unique condensed phases, less air pollution, and low energy consumption. PMID- 11263883 TI - Formation of novel polymeric nanoparticles. AB - We have found that not only block copolymers but also ionomers can self-assemble in a selective solvent to form surfactant-free nanoparticles. The self-assembly can be induced by chemical reaction, polymer-polymer complexation, and microphase inversion in addition to the temperature. A recently developed microwave method for the preparation of uniform surfactant-free polymeric nanoparticles is also reviewed. Our results have revealed that for a given dispersion, the particle surface area occupied per stabilizer (surfactant, polymer chains, and ionic groups) is close to a constant. PMID- 11263884 TI - Novel 3',5'-cyclic nucleotide analogue: adenosine 3',5'-cyclic boranomonophosphate. AB - A general procedure for the first synthesis of a 3',5'-cyclic boranomonophosphate was established. Specifically, adenosine 3',5'-cyclic boranomonophosphosphate (cyclic AMPB, 4c), a P-borane (BH(3)) analogue of adenosine 3',5'-cyclic monophosphate (cAMP), was synthesized via a phosphite approach in good yield. The method is also applicable for syntheses of natural cAMP and its phosphorothioate analogue. The two diasteromers of cyclic AMPB 4c were separated, and their chemical structures were established via spectroscopic methods. PMID- 11263885 TI - Synthesis and characterization of fluorescent cobalamin (CobalaFluor) derivatives for imaging. AB - Fluorescent derivatives of cobalamin have been prepared by linking fluorophores to cobalamin through a propylamide spacer. Fluorescein, naphthofluorescein, and Oregon Green derivatives have been prepared in good yield by reaction of the fluorophore NHS-ester with beta-(3-aminopropyl)cobalamin to form fluorescent cobalamin conjugates (CobalaFluors) that are potentially suitable for the in vitro and in vivo imaging of transcobalamin receptors on cancer cells. PMID- 11263886 TI - Metal-catalyzed release of supported boronic acids for C-C bond formation. AB - The viability of solid-supported boronic acids as reagents for Suzuki couplings and nucleophilic additions to aldehydes and enones was successfully demonstrated. This metal-catalyzed cleavage strategy allows the synthesis of a series of functionalized biphenyl products, benzylic alcohols, and beta-substituted ketones. PMID- 11263887 TI - Use of a conformational radical clock for evaluating alkyllithium-mediated cyclization reactions. AB - The reductive lithiation of nitrile 9 led to the cyclic product 11 as a single diastereomer in 42% ee. The intermediate radical is a conformational radical clock. The radical lifetime can be determined from the optical purity of the product 11. We show that the lifetime of the intermediate radical is too brief to allow a radical cyclization, and thus the cyclization proceeds through an alkyllithium intermediate. PMID- 11263888 TI - Synthesis of biaryls via unusual deoxygenative dimerization of 1,4-epoxy-1,4 dihydroarenes catalyzed by palladium complexes. AB - Treatment of various 1,4-epoxy-1,4-dihydroarenes with trichlorosilane in toluene in the presence of a palladium complex affords the corresponding biaryls in good to excellent yields. The process appears to occur via a novel palladium-catalyzed hydrosilylative dimerization of 1,4-epoxy-1,4-dihydroarenes and subsequent elimination of HOSiCl(3) and H(2)O. PMID- 11263889 TI - Ethyl(methyl)dioxirane as an efficient reagent for the oxidation of nucleoside phosphites into phosphates under nonbasic anhydrous conditions. AB - A convenient method for the oxidation of nucleoside phosphites into phosphates under nonbasic and nonaqueous conditions using commercially available ethyl(methyl)dioxirane has been developed. This oxidation is effective with both N-protected and N-unprotected strategies. PMID- 11263890 TI - Synthesis of polyacetylenic acids isolated from Heisteria acuminata. AB - Four linear polyacetylenic compounds were synthesized. Pentadeca-6,8,10-triynoic acid 1 and octadeca-8,10,12-triynoic acid 2 were synthesized by using acetylene coupling reactions. The syntheses of (Z)-hexadec-11-en-7,9-diynoic acid 3 and (Z) octadec-12-en-7,9-diynoic acid 4 by using vinylic telluride coupling reactions were accomplished. PMID- 11263891 TI - The asymmetric catalytic aldol reaction of allenolates with aldehydes using n fluoroacyl oxazaborolidine as the catalyst. AB - The asymmetric catalytic aldol reaction of silyl allenolates with aldehydes has been achieved by using N-C(3)F(7)CO oxazaborolidine as the catalyst. The fluoroacyl group of the catalyst was found to be crucial for control of enantioselectivity. The reaction provides the first enantioselective approach to beta-halo Baylis-Hillman-type adducts. PMID- 11263892 TI - Enantioselective syntheses of (13)C-labeled (2R)- and (2S)-phytochromobilin dimethyl ester. AB - (2R)- and (2S)-phytochromobilin dimethyl ester have been prepared in enantiomerically pure form, specifically (13)C-labeled at C(10) or C(15). PMID- 11263893 TI - A new synthesis of chlorins. AB - A new synthesis of chlorins has been developed, based upon the acid-catalyzed condensation of dialdehydes AB with dipyrromethanes CD. PMID- 11263894 TI - Directed deprotonation-transmetalation as a route to substituted pyridines. AB - Regioselective C-4 deprotonation of 3-bromopyridine, followed by Li/Zn transmetalation and Pd-mediated coupling processes, provides a flexible entry to 4-substituted and 3,4-disubstituted pyridines. Application of a similar sequence to 2-bromopyridine (with LDA as base) provides 2,3-disubstituted pyridines, but using lithium 2,2,6,6-tetramethylpiperidide (LiTMP) provides access to both the 2,3- and 2,4-disubstituted isomers. PMID- 11263895 TI - Design and synthesis of inhibitors of adenosine and AMP deaminases. AB - Nucleosides and nucleotides which are able to undergo covalent hydration in the aglycone ring system are potential inhibitors of the enzymes adenosine deaminase (ADA) and AMP deaminase, respectively. Calculations of the enthalpy of covalent hydration and of enzyme binding energy have been used to design new inhibitors of ADA. The ribosyl triazolotriazine 16, which was synthesized as a result of these calculations, exists predominantly as the covalent hydrate 18 in water and is a potent inhibitor of mammalian ADA (IC(50) 50 nM). PMID- 11263896 TI - Synthesis of a 3,4,5-trimethoxybenzoyl ester analogue of epigallocatechin-3 gallate (EGCG): a potential route to the natural product green tea catechin, EGCG. AB - The synthesis of a trimethoxybenzoyl ester (D-ring) analogue of epigallocatechin 3-gallate (EGCG) is described. The versatile synthesis route can be used to synthesize A, B, and D ring analogues of EGCG and involves a key cyclization of the chalcone to the 3-flavene. This synthesis provides a possible route to the polyphenolic green tea natural product EGCG. PMID- 11263897 TI - Synthetic studies of the HIV-1 protease inhibitive didemnaketals: stereocontrolled synthetic approach to the key mother spiroketals. AB - The stereocontrolled synthesis of (2S,4R,6R,8S,10S,1'R,1' 'R) 2(acetylhydroxymethyl)-4,10-dimethyl-8(isopropenylhydroxymethyl)-1,7 dioxaspiro[5,5]undecane (4a) and its C1' '-epimer (4b), the key mother spiroketals of the HIV-1 protease inhibitive didemnaketals from the ascidian Didemnum sp., has been carried out through multisteps from the natural (R)-(+) pulegone, which involved the diastereoselective construction of four chiral carbon centers(C-2, C-6, C-8, and C-1') by intramolecular chiral induce. PMID- 11263898 TI - A strategy for probing the autonomy of cross-domain stereochemical communication in glycoconjugates. AB - Glycoproteins contain carbohydrate and peptide sectors. As a model for studying whether there exists stereochemical "communication" between the two domains, we prepared two glycopeptides differing only in the absolute stereochemistry of the peptide domain (L-peptide vs D-peptide). High-field NMR spectroscopy revealed that there are distinct and measurable differences, indicating that the two domains are at some level interactive. PMID- 11263899 TI - A new supported reagent for the photochemical generation of radicals in solution. AB - A new polymer-supported radical source was developed by loading an N-hydroxy thiazole 2(3)-thione on a Wang resin. This new supported reagent can be employed for a solid-phase version of the Hunsdiecker reaction or to liberate free alkoxy radicals, in a variant of the "catch and release" technique, under very mild conditions (irradiation with a discharge lamp) and simplifying the purification procedure. PMID- 11263900 TI - Palladium-catalyzed arylation of enynes and electron-deficient alkynes using diaryliodonium salts. AB - A new single-pot procedure for the synthesis of aryl alkynes is described. Palladium catalyzes the coupling reaction of diaryliodonium compounds with enynes and electron-deficient alkynes to give aryl alkynes in good yields. PMID- 11263901 TI - Partial reduction of annulated heterocycles as a general route to medium rings containing oxygen and nitrogen. AB - The preparation of annulated furans and pyrroles is described as part of a general strategy for the synthesis of medium ring heterocycles. After Birch reduction, the corresponding dihydro compounds were oxidatively cleaved to produce medium ring ethers and amines in an efficient manner. This methodology was successfully applied to the formation of eight- and nine-membered cyclic ethers and nine-membered cyclic amines. Attaching a chiral auxiliary (bismethoxymethylpyrrolidine) to the furan allowed the formation of nine-membered ethers in 95% ee. PMID- 11263902 TI - 3,8-Diazabicyclo[3.2.1]octan-2-one peptide mimetics: synthesis of a conformationally restricted inhibitor of farnesyltransferase. AB - A new synthesis of the 3,8-diazabicyclo[3.2.1]octan-2-one framework is described. Transannular enolate alkylation of piperazinone derivatives provides a flexible route to highly constrained bicyclic peptidomimetic synthons with substitution at the Calpha position. The chemistry was used to produce a conformationally constrained farnesyltransferase inhibitor, which aided the elucidation of enzyme bound conformation. PMID- 11263903 TI - Catalytic asymmetric coupling of 2-naphthols by chiral tridentate oxovanadium (IV) complexes. AB - A series of chiral oxovanadium(IV) complexes derived from tridentate N-3,5 substituted and N-3,4-benzo- and N-5,6-benzo-salicylidene-alpha-amino acids can serve as efficient catalysts for the enantioselective oxidative couplings of various 3-, 6-, and 7-substituted 2-naphthols under O(2). The best scenario involves the use of a vanadyl complex arising from 2-hydroxy-1-naphthaldehyde and valine (or phenylalanine) in CCl(4), leading to BINOLs in good yields (75-100%) and with enantioselectivities of up to 68%. PMID- 11263904 TI - Novel porphyrin-cryptand cyclic systems: receptors for saccharide recognition in water. AB - Two macrocyclic porphyrin sandwich systems, 4 and 5, together with the linear compound 6 have been prepared and examined as saccharide receptors. The cyclic porphyrin-cryptand conjugates 4 and 5 bind saccharides efficiently in highly competitive media with a preference for trisaccharides probably due to a complementary topology of hydrophobic and hydrophilic solvating regimes with respect to the sugar guests. PMID- 11263905 TI - A novel multicomponent synthesis of polysubstituted 5-aminooxazole and its new scaffold-generating reaction to pyrrolo[3,4-b]pyridine. AB - A novel three-component synthesis of 5-amino oxazole (1) is reported. Its subsequent reaction with alpha,beta-unsaturated acyl chloride leads to polysubstituted pyrrolopyridine (2). A triple domino process, acylation/IMDA/retro-Michael cycloreversion, was involved in the latter process. The methodology allows the quick preparation, from simple and readily available inputs, of highly functionalized title compounds not easily accessed by other methods. PMID- 11263906 TI - Donor-acceptor interactions in crystal engineering. AB - The molecular complex formed between 4-methyltolane and bis(4-N methylpyridinium)ethyne ditriflate is reported. The X-ray crystal structure indicates that the crystalline superstructure consists of infinite zigzag ribbons of interlocked donor-acceptor complexes separated by triflate counterions. PMID- 11263907 TI - Synthesis of acetylenic cyclophanes via intramolecular self-assembly: evidence of perfluorophenyl-phenyl quadrupole interactions in the solution state. AB - Reported herein is an example of a solution-state cross-coupling cyclization with an outcome mediated by perfluorophenyl-phenyl electrostatic interactions. PMID- 11263908 TI - Cu2+-induced blue shift of the pyrene excimer emission: a new signal transduction mode of pyrene probes. AB - A pentiptycene-bispyrenyl system (1) has been synthesized and investigated as a fluorescent chemosensor for metal ions. A novel blue shift along with an intensity enhancement of the pyrene excimer emission is observed for 1 in the presence of Cu(2+). Such a new signal transduction mode of pyrene probes results from the formation of a static pyrene excimer that has very different characteristics from its dynamic counterpart. PMID- 11263909 TI - Synthesis of an eight-membered cyclic pseudo-dipeptide using ring closing metathesis. AB - Ring closing metathesis of diallylglycine 6 provided cyclic Z-olefin 7 in 80% yield. The reaction was promoted by substitution of the amide nitrogen with the 2,4-dimethoxybenzyl group allowing for the required cis diallylglycine amide rotamer. Removal of the protecting groups provided cyclic dipeptide 2, a constrained scaffold useful in peptidomimetic research. PMID- 11263910 TI - Versatile synthons for optically pure alpha-amino aldehydes and alpha-amino acids: (+)- and (-)-4,5-dialkoxy-2-oxazolidinones. AB - Both enantiomers of trans-5-benzyloxy-4-methoxy- (BMOx) and trans-4,5-dimethoxy-2 oxazolidinones (DMOx), which are readily accessible from simple 2-oxazolone heterocycles, represent good candidates for a new class of chiral synthons for use in the preparation of optically pure alpha-amino aldehydes and alpha-amino acids, respectively. PMID- 11263911 TI - Chlorodicyclohexylborane-mediated aldol additions of alpha,alpha'-dioxygenated ketones. AB - Boron aldol additions of variously O-protected alpha,alpha'-dioxygenated ketones using dicyclohexylboron chloride and a tertiary amine have been investigated. The stereoselectivity of the process was dependent on the protecting group on the alpha-oxygen atoms. Notably, ketones with bulky silyloxy groups gave syn aldols, most likely via Z enolates. This rules out the participation of chelates during the enolization process, at least in the presence of such sterically crowded protecting groups. An alternative explanation is offered. PMID- 11263913 TI - Palladium-catalyzed tandem bis-allylation of isocyanates. AB - A tandem bis-allylation of p-toluenesulfonyl isocyanate can be achieved by palladium-catalyzed three-component coupling reaction with allylstannanes and allyl chlorides. A high level of regioselectivity can be obtained by the appropriate choice of the allylic substituents. The reaction mechanism and the regiochemistry of the reaction can be explained by formation of an amphoteric bis allylpalladium intermediate. This bis-allylpalladium intermediate undergoes an initial electrophilic attack on one of the allyl moieties followed by a nucleophilic attack on the other. PMID- 11263912 TI - Synthesis of (+/-)-epoxysorbicillinol using a novel cyclohexa-2,5-dienone with synthetic applications to other sorbicillin derivatives. AB - A novel route to epoxysorbicillinol as well as dimers of sorbicillin is reported. The synthesis is-in principle-amenable to enantioselectivity. The key step is an oxidative dearomatization to produce a stable and highly malleable p-quinol intermediate, which undergoes a highly diastereoselective epoxidation. PMID- 11263915 TI - Highly fructose selective transport promoted by boronic acids based on a pentaerythritol core. AB - We have designed and synthesized a highly lipophilic boronic acid (11) with a molecular shape that makes it much more effective at carrying sugars through organic membranes than a previously used steroidal boronic acid. The corresponding diboronic acid (12) was also found to transport fructose ahead of glucose with a very high selectivity (7.6:1.0). Modeling suggests that 12 is able to carry two fructose molecules at once in a complex stabilized through hydrogen bonding and ion pairing. PMID- 11263914 TI - Iodine monochloride/silver trifluoromethanesulfonate (ICI/AgOTf) as a convenient promoter system for O-glycoside synthesis. AB - The novel promoter system iodine monochloride/silver trifluoromethanesulfonate (ICl/AgOTf) was evaluated with various thioglycoside donors and saccharide acceptors, and O-glycosides were obtained in 46-82% yield. Several practical advantages of the ICl/AgOTf system over known promoter systems were observed, such as convenient handling of the reagents and absence of byproducts related to N-succinimide. PMID- 11263916 TI - Mechanism of pH-dependent photolysis of aliphatic amino acids and enantiomeric enrichment of racemic leucine by circularly polarized light. AB - It has been proposed that the origin of biological homochirality may be the result of irradiation of a racemic sample of amino acids by circularly polarized light (CPL). To determine the mechanism of enantiomeric enrichment, the irradiation of aliphatic amino acids by CPL was undertaken. An enantiomerically enriched sample (e.g., L isomer enrichment from r-CPL) was found to result from the preferential excitation/decomposition of one enantiomer over another via a Norrish Type II mechanism (leucine, valine, and isoleucine), with the enantiomeric excess dependent on the degree of protonation of the amino/carboxylic acid moiety. PMID- 11263917 TI - Synthesis and properties of peptide nucleic acids containing a psoralen unit. AB - We prepared the psoralen PNA unit from 8-methoxypsoralen and synthesized various PNAs containing psoralen by a typical (t)()Boc method. PNAs containing psoralen (P-PNA) at strand end formed a stable duplex with complementary DNA. The hybridization of P-PNA with complementary DNA resulted in a considerable decrease of the psoralen fluorescence. PMID- 11263918 TI - Asymmetric synthesis of the northern half C1-C16 of the bryostatins. AB - Starting from 8-oxabicyclo[3.2.1]oct-6-en-3-one and racemic 2,2-dimethyl-8 oxabicyclo[3.2.1]oct-6-en-3-one, the C1-C16 segment of the bryostatins has been synthesized in 30 steps and 9% overall yield (17 steps longest linear sequence). Fragment coupling by dithiane strategy and protecting group manipulations provided an advanced chemodifferentiated northern half segment. PMID- 11263919 TI - Epoxides and aziridines from diazoacetates via ylide intermediates. AB - An effective methodology is reported for stereospecific epoxidation and aziridination via carbonyl ylide intermediates using rhodium(II) acetate catalyzed reactions of phenyl- and styryldiazoacetates with aldehydes, ketones, or imines. PMID- 11263920 TI - Synthesis of (Z)-vinylsilanes with high diastereoselectivity by using samarium diiodide. AB - beta-Elimination of O-acetyl 1-chloro-1-trimethylsilylalkan-2-ols 1 was achieved by using samarium diiodide as a metaling reagent and afforded the corresponding (Z)-vinylsilanes with high stereoselectivity. The starting compounds 1 were easily prepared by treatment of different aldehydes with (chlorolithiomethyl)trimethylsilane and further acetylation. PMID- 11263921 TI - Efficient catalysis of a Diels-Alder reaction by metallo-vesicles in aqueous solution. AB - Vesicles have been prepared from a cyclic phosphate ester (5,5-di-n-dodecyl-2 hydroxy-1,3,2-dioxaphosphorinan-2-one) with copper(II) counterions (Cu(dDP)(2)). They form a highly efficient aqueous Lewis acid catalyst system. The reaction of two azachalcon derivatives (1a, 1b) with cyclopentadiene (2) was studied to elucidate the catalytic potential of this system. PMID- 11263923 TI - An aldol approach to the synthesis of the EF fragment of spongistatin 1. AB - A synthesis of the C29-C51 fragment of spongistatin 1, containing the E and F rings, has been completed. The approach relies on four diastereoselective aldol additions and an asymmetric glycolate alkylation to establish eight of the eleven stereogenic centers. The intact chlorodiene side chain was appended by a Lewis acid catalyzed addition of an allylstannane to an epoxy enol ether. PMID- 11263922 TI - Synthesis of a tripeptide derivative containing the Phe-Arg hydroxyethylene dipeptide isostere. AB - The protected hydroxyethylene dipeptide isostere of Phe-Arg and the tripeptide derivative 1 were synthesized as components of potential peptidase inhibitors. Key to the success of these syntheses is selective rhodium-catalyzed hydroboration in the presence of a readily reduced lactone. A convenient one-pot conversion of azides to diprotected guanidines was developed on the basis of the Staudinger reaction. PMID- 11263924 TI - A concise enantioselective synthesis of antimalarial febrifugine alkaloids. AB - Reaction of (S)-2-(tert-butyldiphenylsilyloxy)-5-(mesyloxy)pentanal with hydroxylamine in allyl alcohol brought about simultaneous 1,3-dipolar cycloaddition of the resulting nitrone to allyl alcohol to give three diastereoisomeric adducts, from which (+)-febrifugine and (+)-isofebrifugine, potent antimalarial alkaloids, were synthesized. PMID- 11263925 TI - Lewis acid catalyzed intramolecular Diels-Alder reactions of acyclic (Z) substituted 1,3-dienes. AB - Lewis acid catalyzed intramolecular Diels-Alder reactions of trienes (E,E,Z)-1a d, (E,E,Z)-4a-d, and (E,Z,Z)-7a,b are described. Trienes containing enal or enone dienophiles cyclize in excellent yield under mild conditions using substoichiometric amounts of MeAlCl(2), in most cases with high levels of diastereoselectivity. The thermal IMDA reactions of 1a, 4a, and 7a require forcing conditions and proceed in low yield with reversed stereoselectivity in the cases of 1a and 4a. PMID- 11263927 TI - BRCA: the breast, ovarian, and other cancer genes. PMID- 11263928 TI - A genetic epidemiological study of carcinoma of the fallopian tube. AB - OBJECTIVE: The goal of this work was to evaluate the importance of genetic factors in the etiology of fallopian tube cancer. METHODS: All pathologically confirmed cases of fallopian tube cancer diagnosed in Ontario from 1990 to 1998 were identified from the records of the Ontario Cancer Registry. Living patients were approached to provide information about their family history and to provide a blood sample for testing for mutations in BRCA1 and BRCA2. RESULTS: A modest increase in the risk of ovarian cancer (relative risk (RR) = 2.2; 95% confidence interval (CI) = 0.4, 6.3) and of early-onset breast cancer (RR = 2.4; 95% CI = 0.6, 6.1) was observed in the first-degree relatives of the fallopian cancer cases. Five of the forty-four cases were positive for a mutation in BRCA1 (11%) and two were positive for a BRCA2 mutation (5%). Five of eighteen women diagnosed at or before age 55 were positive (28%). Two of the seven mutation carriers had a strong family history of breast and ovarian cancer, and three carriers had a modest family history. Three of the forty-four cases were Jewish, and of these, two carried a founder mutation characteristic of this population. CONCLUSIONS: Fallopian tube carcinoma should be considered to be a clinical component of the hereditary breast-ovarian cancer syndrome, and may be associated with BRCA1 and BRCA2 mutations. Genetic evaluation should be offered to women who present with fallopian tube carcinoma. It is important to consider the risk of fallopian tube carcinoma when prophylactic oophorectomy is performed in high-risk women. PMID- 11263929 TI - Cancer risk in young women at risk of hereditary nonpolyposis colorectal cancer: implications for gynecologic surveillance. AB - OBJECTIVES: The lifetime risk of endometrial and ovarian cancers in hereditary nonpolyposis colorectal cancer (HNPCC) is up to 60 and 12%, respectively, in addition to the high risk of colorectal cancer. International guidelines recommend surveillance of those at risk with colonoscopy every 1--2 years from age 20--25 years and annual gynecologic surveillance from 25--35 years for women. We reviewed our own experience to see whether these recommendations were appropriate. METHODS: Pedigrees of 120 HNPCC families registered with the Familial Bowel Cancer Service at The Royal Melbourne Hospital were reviewed. Ninety families met our criteria for HNPCC and were included in the study. Pedigrees were analyzed to identify early-age onset colorectal and gynecologic cancers and to calculate cumulative incidence of both cancer types for at-risk women (HNPCC-affected and first-degree female relatives of affected family members) for comparison with the general population. RESULTS: Colorectal cancer occurred in 434 individuals, endometrial cancer in 43, and ovarian cancer in 24. Gynecologic and colorectal cancers were diagnosed at similar ages (mean 49.3 versus 51.8 years; P = 0.25), with 5 (7.1%) gynecologic cancers diagnosed by 35 years. Cumulative incidences of gynecologic and colorectal cancers to ages 25, 30, 35, and 40 years were substantially higher among at-risk women than in the general population. CONCLUSIONS: Consideration should be given to offering gynecologic cancer surveillance from the age of 25 years, as for colorectal cancer. However, this approach should be individualized as it has yet to be demonstrated that surveillance reduces the mortality of gynecologic cancer in HNPCC. PMID- 11263931 TI - Overexpression of CD44 variant 6 in human endometrial cancer and its prognostic significance. AB - OBJECTIVE: We evaluated the prognostic significance of CD44 isoform CD44 V6 in endometrial cancer. METHOD: Immunohistochemistry was used to determine the expression of CD44 V6 in 78 randomly selected endometrial cancer specimens. RESULTS: CD44 V6 was detected in 53% (41/78) of the tumor samples. Clinicopathological evaluation revealed an inverse correlation with CD44 V6 expression and conventional poor prognosticators such as lower segment involvement, nuclear and structural grade, myometrial invasion, serosal involvement, lymph-vascular space invasion, and adnexal metastasis. Furthermore, the CD44 V6-negative group was found to be associated have a higher recurrence rate and a shorter disease-free survival. CONCLUSION: These findings indicate that absence of CD44 V6 expression in cases of endometrial cancer might be coupled with a more aggressive course. PMID- 11263930 TI - Cervical dysplasia in women infected with the human immunodeficiency virus (HIV): a correlation with HIV viral load and CD4+ count. AB - OBJECTIVES: The incidence of cervical dysplasia and carcinoma is known to be increased in HIV-infected women. In addition, there is a positive correlation between HIV viral load (VL), CD4+ count, and opportunistic infections, as well as the incidence of various malignancies. This study compares HIV VL and CD4+ count with the presence of cervical dysplasia, as well as with the degree of severity of dysplasia. METHODS: A retrospective chart review of 350 HIV-infected women with polymerase chain reaction (PCR) quantitation of viral load was performed to identify 82 women with biopsy-proven cervical dysplasia and 25 women without any significant cervical pathology. The highest plasma VL within a year of the patients' cervical pathology and corresponding CD4+ count was selected and compared with cervical pathology. Univariate and multivariate statistical analysis using Student's t test and logistic regression analysis was used to analyze the significance of other risk factors such as age, race, smoking history, history of illicit drug use, and prior sexually transmitted disease as well as of viral load and CD4+ count. RESULTS: Of 82 cases of cervical dysplasia, 33 (40.24%) were mild (CIN I), 47 (57.32%) were either moderate or severe (CIN II III) dysplasia, and 2 demonstrated invasive squamous cell carcinoma (2.44%). A significant statistical difference was found when comparing either HIV plasma VL or CD4+ T-cell counts with the presence of cervical dysplasia on biopsy (P < 0.005). However, only CD4+ count was identified as an independent risk factor for the presence of cervical dysplasia after multivariate analysis. CONCLUSION: In our population, there is a significant correlation between VL and CD4+ count and the presence of cervical dysplasia. However, VL does not appear to be an independent risk factor for cervical dysplasia in this population of HIV-infected women. PMID- 11263932 TI - Phase I trial of ifosfamide and 24-h infusional paclitaxel in pelvic malignancies: a Gynecologic Oncology Group study. AB - OBJECTIVE: The goal of this work was to develop a combination chemotherapy regimen consisting of ifosfamide and paclitaxel to be evaluated in the management of gynecologic malignancies. METHODS: The Gynecologic Oncology Group conducted a Phase I trial of the regimen, initially with paclitaxel (24-h infusion) delivered on Day 1 and ifosfamide (1 h) administered (with Mesna) over the subsequent 4 days. All patients received granulocyte colony-stimulating factor (G-CSF) starting 24 h after the Day 5 chemotherapy. Treatment was repeated on a 28-day schedule. A cohort of patients also received the alternate sequence of ifosfamide (4 days) followed by paclitaxel. RESULTS: Twenty-two patients were evaluated. Even at the lowest dose level tested (paclitaxel 135 mg/m(2) followed by ifosfamide 1 g/m(2)/day x 4 days) grade 4 neutropenia was almost universal, despite the routine use of G-CSF. The alternate drug administration sequence resulted in marrow suppression of similar severity. CONCLUSION: The combination of 24-h infusional paclitaxel with ifosfamide delivered over 4 days results in severe neutropenia, despite the administration of G-CSF, and is not recommended for further evaluation. In view of the known activity of the two agents in several malignancies, including cervix cancer, it would be reasonable to investigate the delivery of the agents employing alternative treatment schedules predicted to result in less severe marrow suppression (e.g., 3-h infusional paclitaxel). PMID- 11263933 TI - Allelic deletion mapping on chromosome 6q and X chromosome inactivation clonality patterns in cervical intraepithelial neoplasia and invasive carcinoma. AB - OBJECTIVE: Loss of heterozygosity (LOH) profiles and X chromosome inactivation patterns are analyzed in 42 patients with cervical intraepithelial neoplasias (CIN), including low-grade (CIN1) and high-grade (CIN2, CIN3) lesions, and 22 patients with invasive cervical carcinomas. METHOD: Laser capture microdissection was utilized to procure pure matched normal and lesional cells from each case. Sixteen microsatellite markers on four chromosomal arms, 6q21-q25.1, 8p21, 13q12.3--q13, and 17q12--q21, were amplified for LOH, as well as the HUMARA locus for X chromosome inactivation analysis. Eight additional markers spanning the long arm of chromosome 6 were utilized in all cases showing LOH on this arm and in which further tissue material was available for microdissection. RESULTS: Fifty-five percent of carcinomas showed deletions on chromosome bands 6q21- q25.1, 43% on 13q12.3--q13, and 40% on 17q12--q21. Deletions on 6q were identified in CIN3 (40%), CIN2 (37%), and CIN1 (10%), on 13q in CIN3 (33%) and CIN2 (33%), and rarely on chromosomal arm 17q. Finer 6q mapping revealed that marker D6S310 (q22) represented the centromeric and marker D6S255 (q25--q16) the telomeric boundary of deletion. A second, telomeric area of deletion at marker D6S281 (q27) was also identified. Monoclonal X chromosome inactivation patterns were identified in 12/13 cancers, 13/14 CIN3, 5/10 CIN2, and 0/6 CIN1. CONCLUSIONS: Two areas of deletion on chromosome 6q were identified in cervical tumors, suggesting the presence of tumor suppressor gene(s) inactivated in this neoplasia. LOH on this arm were identified early during cervical tumor progression. LOH on 13q and 17q also occur in cervical cancers. X chromosome inactivation patterns suggest that CIN develops into a monoclonal lesion during progression from CIN1 to CIN3. PMID- 11263934 TI - Positron emission tomography with (18)F-fluorodeoxyglucose of uterine sarcoma: a comparison with magnetic resonance imaging and power Doppler imaging. AB - OBJECTIVE: The effectiveness of positron emission tomography with (18)F fluorodeoxyglucose (FDG-PET) for diagnosis of uterine sarcoma was evaluated in comparison to the effectiveness of magnetic resonance (MR) imaging and power Doppler imaging. METHOD: The cases of five Osaka City University Hospital patients diagnosed with uterine sarcoma based on histopathological examination, in whom FDG-PET, MR imaging, and power Doppler imaging studies had been performed preoperatively, were reviewed. A comparative study of the usefulness of these three imaging modalities for diagnosis of sarcoma was conducted. Tumors comprised three leiomyosarcomas, one endometrial stromal sarcoma, and one carcinosarcoma. RESULTS: FDG-PET examinations were 100% positive for the five sarcomas; MR imagings were 80% positive (four of five cases), and US was 40% positive (two of five cases). The mean standardized uptake value of the sarcomas was 4.5 +/- 1.3. CONCLUSION: The sarcoma lesions were clearly imaged by FDG-PET. FDG-PET may be a most useful diagnostic method for uterine sarcoma. PMID- 11263935 TI - High incidence of positive peritoneal cytology in low-risk endometrial cancer treated by laparoscopically assisted vaginal hysterectomy. AB - OBJECTIVE: Laparoscopically assisted vaginal hysterectomy (LAVH) has evolved into an alternative form of surgical management in the treatment of low-risk endometrial cancer. The purpose of this study was to determine whether low-risk endometrial cancer patients are subject to a higher incidence of positive peritoneal cytology when treated with LAVH compared to total abdominal hysterectomy (TAH). METHODS: We retrospectively reviewed the medical records of patients with low-risk endometrial cancer (grade 1--2 endometrioid type with no evidence of extrauterine spread or grade 3 with <50% myometrial invasion (MI), no cervical or adnexal involvement, and negative lymph nodes when sampled) treated at Memorial Sloan-Kettering Cancer Center from January 1993 to September 1999. We compared 131 patients treated with LAVH to 246 controls who underwent TAH. The two groups were compared for known prognostic factors including grade, MI, vascular space involvement, and lower uterine segment extension. RESULTS: The mean age of patients who underwent LAVH (61 years) was similar to that of the controls (62 years). Fourteen (10.3%) of the patients treated with LAVH had positive peritoneal cytology compared to only 7 (2.8%) of the control population. Factors including FIGO grade, myometrial invasion, and preoperative hysteroscopy did not influence the final results. When stratifying for these factors, the odds ratios of having positive peritoneal washings in those patients treated by LAVH were 5.2, 5.2, and 3.7, respectively. CONCLUSION: Treatment of low-risk endometrial cancer by LAVH is associated with a significantly higher incidence of positive peritoneal cytology. This may be due to the retrograde dissemination of cancer cells into the peritoneal cavity during uterine manipulation. The clinical significance of these findings is yet to be determined. PMID- 11263936 TI - Serum levels of macrophage colony-stimulating factor in trophoblastic disease. AB - OBJECTIVES: The aims of this study were to measure levels of colony stimulating factor (CSF-1) in patients with trophoblastic disease, to determine whether such measurement may be useful to supplement measurement of the prognostically reliable human chorionic gonadotrophin (hCG), and to assess whether measurement of CSF-1 may be helpful in predicting requirement for chemotherapy in patients with hydatidiform mole. METHODS: Serial weekly serum samples were selected for CSF-1 assay from representative diagnostic groups of patients with trophoblastic disease: hydatidiform-mole with spontaneous resolution, low-risk post hydatidiform-mole trophoblastic tumor, partial hydatidiform mole, high-risk metastatic gestational trophoblastic tumor, primary ovarian choriocarcinoma, and placental site trophoblastic tumor. hCG was measured by an in-house radioimmunoassay that measures all parts of the hCG molecule. CSF-1 was measured by radioimmunoassay with (125)I-labeled recombinant CSF-1. The upper level of normal CSF-1 was taken as 8 ng/ml. RESULTS: In this study of 45 patients with trophoblastic disease, some very high levels of CSF-1 were encountered. In a few patients there was dramatic correlation with hCG. Generally, however, CSF-1 and hCG did not correlate. CSF-1 was frequently not elevated when hCG was still significantly elevated and conversely CSF-1 was elevated when hCG was negative. CONCLUSION: The measurement of CSF-1 does not appear to be useful in managing trophoblastic disease as it does not correlate with the level of hCG. Occasionally, high levels of CSF-1 were found in patients with trophoblastic disease. PMID- 11263937 TI - Psychosocial adjustment in gynecologic cancer survivors: a longitudinal study on risk factors for maladjustment. AB - OBJECTIVE: The objective was to describe the change in psychosocial state over time and to identify risk factors for maladjustment in gynecologic cancer survivors. Awareness of these issues is important for planning supportive care services for cancer patients. METHODS: A longitudinal prospective study of patients with newly diagnosed gynecologic cancer using individual patients as their own control was performed. Patients were interviewed after confirmation of the diagnosis and were reassessed at 6 and 18 months after completion of treatment and with no evidence of recurrent disease. Psychological adjustment was measured by self-rating on self-esteem, outlook on life, self-role, and femininity. Neuroticism and anxiety were assessed using a neuroticism score and the Hamilton Anxiety Scale. Depressive symptoms were questioned directly. Social adjustment was assessed by changes in working capacity or work status, leisure activity, marital relationship, and sexual activity. RESULTS: Seventy-four women participated. Adjustment problems did not occur in the majority of patients. Psychosocial adjustment was different for patients receiving different types of treatments. Improvement in feminism (P = 0.050) and neuroticism (P = 0.010) was observed for patients receiving chemotherapy and deterioration was observed in patients treated with surgery. Deterioration in neuroticism was associated with lower education level (P = 0.032). With religious belief, there was better family support and more significant improvement in social activity (P = 0.038). CONCLUSION: Most patients adapted well. Patients at risk for psychosocial maladjustment include those who are treated surgically, less educated, and without religious belief. PMID- 11263938 TI - Risk of endometrial carcinoma associated with BRCA mutation. AB - OBJECTIVE: Inherited mutations in the BRCA1 or BRCA2 genes are associated with a greatly increased lifetime risk of breast and ovarian cancers and a modestly increased risk of several other cancer types. Several case reports of endometrial carcinoma in women with a BRCA mutation have led to speculation regarding the effect of these genes on the risk of endometrial cancer. The purpose of this study was to test the hypothesis that germline mutation of a BRCA gene is associated with an increased risk of endometrial carcinoma. METHODS: A retrospective cohort of 199 consecutive Ashkenazi Jewish patients with endometrial carcinoma was identified from a 12-year period at this institution. All were genotyped for the three BRCA founder mutations (185delAG and 5382insC in BRCA1 and 6174delT in BRCA2) that exist in this population, and the case frequency was compared to the known population frequency of these mutations. Additionally, endometrial carcinomas occurring in patients with BRCA mutations were assessed for somatic loss of the wild-type BRCA allele. RESULTS: Germline BRCA mutations were identified in 3 (1 in BRCA1 and two in BRCA2) of 199 (1.5%) patients, compared to a frequency of 2.0% in this population generally. A relative risk of endometrial carcinoma associated with BRCA mutation, as estimated by the odds ratio, was calculated as 0.75 (95% CI = 0.24--2.34; P = 0.6). Loss of the wild-type BRCA allele was observed in two of three tumors associated with a BRCA mutation. CONCLUSIONS: For individuals with a germline BRCA mutation, the lifetime risk of endometrial carcinoma is not increased. PMID- 11263939 TI - Two BRCA1-positive epithelial ovarian tumors with metastases to the central nervous system: a case report. AB - BACKGROUND: Cerebral metastasis secondary to ovarian cancer is a rare phenomenon. While no clear relationship to known prognostic factors is found, others suggest this as a biologically diverse behavior of ovarian cancer. CASES: In a pilot study, 37 invasive epithelial ovarian cancer samples were analyzed to detect the frequency of BRCA1/BRCA2 mutations in the south of Sweden (results published). A retrospective follow-up revealed that 2 of these (2/37; 5.4%) patients developed central nervous system metastases during the course of their disease. Both patients had advanced surgical stage disease at the time of diagnosis, with histopathological serous type tumors that were negative for estrogen and progesterone receptors. One of these patients carried a germline BRCA1 mutation, whereas a somatic BRCA1 mutation was identified in the other patient. CONCLUSIONS: To the best of our knowledge the molecular genetic profile of these tumors is not found in the literature and it is suggested that such analyses could provide some insight for a better understanding of this rare phenomenon. PMID- 11263940 TI - Recurrent carcinoma in situ of a neovagina. AB - BACKGROUND: Development of carcinoma in situ in a neovagina is rare. CASE: A case of carcinoma in situ of a neovagina complicated by recurrence after ablative therapy is discussed. Recurrence occurred within 4 months of initial therapy, and a total vaginectomy was performed after the patient declined other therapeutic options. CONCLUSION: Recurrent carcinoma in situ of a neovagina can be successfully treated by surgical excision. PMID- 11263941 TI - Tamoxifen as systemic treatment of advanced breast cancer during pregnancy--case report and literature review. AB - BACKGROUND: When patients with metastatic breast cancer become pregnant, management is complicated by the potential harms of drug treatment to the fetus and by the potential effects of the pregnancy on the cancer. Chemotherapy is considered optimal systemic anti-cancer therapy from the second trimester, while tamoxifen has been considered inappropriate due to concerns over possible teratogenesis and lack of efficacy. CASE: We report a patient who became pregnant concurrent with the identification of metastatic breast cancer and who elected to continue her pregnancy with tamoxifen as sole systemic anti-cancer therapy. The pregnancy was difficult, but a normal child was delivered and the mother responded to subsequent hormone manipulation. The putative teratogenic effects of tamoxifen and the mechanisms underlying tamoxifen resistance in this setting are discussed. CONCLUSIONS: The use of tamoxifen in pregnancy is complex, but is not necessarily associated with fetal harm and may be considered a therapeutic option in selected cases. PMID- 11263942 TI - Clear cell variant of malignant melanoma of the uterine cervix: a case report and review of the literature. AB - BACKGROUND: A rare variant of malignant melanoma of the uterine cervix mimicking clear cell carcinoma or clear cell sarcoma is described. CASE: A 33-year-old Japanese woman was admitted to the hospital complaining of genital discharge and lower back pain. The stage was FIGO IIB and radical hysterectomy and pelvic lymphadenectomy were done. Pathological examination, immunohistochemical studies of melanin granules, and molecular analysis of the EWS/ATF-1 fusion gene were also done. A diffuse proliferation of amelanotic clear cells was detected in the uterine cervix. Tumor cells were positive for HMB 45, Melan-A (MART-1), and S-100 protein and negative for epithelial markers. The EWS/ATF-1 fusion gene was not detected. CONCLUSION: This is apparently the first report of a case of clear cell melanoma of the uterine cervix. Despite its rarity, this variant of malignant melanoma should be considered when diagnosing clear cell neoplasms of the uterine cervix. PMID- 11263943 TI - Vacuum-assisted closure for cutaneous gastrointestinal fistula management. AB - BACKGROUND: Cutaneous gastrointestinal (GI) fistulas are a challenging complication in the oncologic patient population. The fistulous effluent is difficult to manage and adversely alters quality of life. Nonsurgical management of enteric fistulas is successful in 30% of cases, requiring at least 4 to 6 weeks. Recently a new technology has been developed to expedite wound healing. The Vacuum-Assisted Closure (VAC) method is a subatmospheric pressure technique that has been demonstrated in laboratory and clinical studies to significantly improve wound healing. Here we report its use in the successful medical management of a cutaneous GI fistula. CASE: A 63-year-old woman with advanced ovarian cancer developed an extensive complex cutaneous GI fistula in an open healing wound. She was treated with total parental nutrition and the VAC device, which resulted in complete closure of the fistula. CONCLUSION: We propose that the VAC device may be a useful adjunct for the medical management of cutaneous GI fistulas. PMID- 11263944 TI - Uterine papillary serous carcinoma presenting as distant lymph node metastasis. AB - BACKGROUND: Uterine papillary serous carcinomas are highly aggressive malignancies that often present with high-stage disease. We report two cases that presented initially as distant metastatic disease. One case was found incidentally at the time of axillary dissection for breast cancer and the second case in the workup of a neck mass. CASES: Clinicopathologic review of the patient material including review of routine H&E pathology and immunohistochemical studies of the patients tumors was performed. Both cases showed high-grade papillary carcinomas with psammoma bodies metastatic to lymph nodes in the axilla or neck. Sampling of the endometrium in these patients confirmed primary uterine papillary serous carcinoma. Patients were treated with adjuvant chemotherapy. CONCLUSIONS: Metastatic uterine papillary serous carcinoma presenting initially in distant sites is an unusual manisfestation of this highly aggressive tumor. This tumor should be considered in the differential diagnosis when patients present with metastatic high-grade papillary serous carcinomas and the primary site is unknown. PMID- 11263945 TI - Re: van Nagell JR Jr, et al. The efficacy of transvaginal sonographic screening in asymptomatic women at risk for ovarian cancer. Gynecol Oncol 2000; 77: 350-6. PMID- 11263946 TI - Re: van Nagell JR Jr, et al. The efficacy of transvaginal sonographic screening in asymptomatic women at risk for ovarian cancer. Gynecol Oncol 2000; 77: 350-6. PMID- 11263948 TI - Doxil-induced radiation recall: a cause for false-positive PET scan findings. PMID- 11263949 TI - Lactoferrin: a tamoxifen-responsive protein in normal and malignant human endometrial cells in culture. AB - We investigated the possible role of the estrogen-regulated protein lactoferrin (Lf) in the response of isolated normal human endometrial epithelial cells (NHEC) and established human endometrial carcinoma (EC) cell lines to tamoxifen (TAM). Using confocal laser scanning microscopy and a monospecific antibody, Lf was localized to the cytoplasm of normal and EC cells. Antibody neutralization of secreted Lf inhibited, whereas exogenous Lf (0--100 microg/ml) enhanced, cell proliferation in both classes of cells. Treatment of NHEC with TAM inhibited cell growth via a protein kinase-C-mediated pathway, concomitant with a reduction in the staining intensity for Lf. Importantly, in EC cells, TAM greatly enhanced the staining intensity for Lf, but did not affect cell growth. We propose that stable expression of Lf protein by EC cells may impart a survival advantage to these cells, which may, in part, account for the resistance of these cells to tamoxifen. PMID- 11263950 TI - Functional and morphological differences between human alveolar and interstitial macrophages. AB - Macrophages play an essential role in pulmonary host defense. They are, however, a heterogeneous cell population located in different lung compartments. This study was designed to elucidate differences between two macrophage populations obtained from the human lung, i.e., alveolar macrophages (AM) and interstitial macrophages (IM). Macroscopically tumor-free lung segments from nine patients undergoing lobectomy or pulmectomy were studied. All patients had a diagnosis of primary lung cancer. AM were recovered by bronchoalveolar lavage and IM were isolated by mechanical fragmentation of the lavaged lung segments followed by enzymatic treatment. The cell fractions were analyzed with respect to morphology (transmission electron microscopy) and function (phagocytosis). The cells in the IM fraction were smaller (7.6 +/- 1.8 microm (mean +/- SD) compared with 16.0 +/- 4.1 microm) and morphologically more heterogeneous than those in the AM fraction. Interestingly, a considerable portion of the cells in the IM fraction had a typical AM-like appearance. Despite this, the AM fraction had a higher phagocytic activity compared to IM, with faster attachment and ingestion processes (P <0.001 for both). We conclude that the heterogeneity of human lung macrophages must be taken into consideration when their role in the inflammatory response is studied. PMID- 11263951 TI - Adeno-associated virus is associated with a lower risk of high-grade cervical neoplasia. AB - Adeno-associated virus (AAV) is a ubiquitous human helper-dependent parvovirus which may interact with human papillomaviruses (HPV) to modify a woman's risk of cervical neoplasia. This analysis was nested in a cohort study of low-income women receiving Pap smears as part of their family planning services. We selected cases (55 with high-grade cervical squamous intraepithelial lesions (HSIL) and 162 with low-grade LSIL) and controls (96 women with normal cervical cytology) and analyzed cervical DNA for AAV, using PCR amplification/dot blot hybridization, and HPV, using hybrid capture I. AAV positivity was associated with a significantly reduced risk of HSIL (age and HPV-adjusted odds ratio (aOR) = 0.32) yet not with LSIL (aOR = 0.78); 53.8% of HSIL, 66.9% of LSIL, and 70.7% of controls were AAV+. AAV appears to interact with HPV to reduce SIL risk; relative to the HPV-/AAV+ exposure, the respective aORs for HSIL and HPV+/AAV-, HPV+/AAV+, and HPV-/AAV+ were 17.0, 6.9, and 3.5. AAV+ was not associated with age, race, HPV status, or sexual or reproductive risk factors. These results strongly suggest that AAV may play a protective or inhibitory role in late stage cervical carcinogenesis. This conclusion needs to be verified in additional epidemiologic studies. PMID- 11263952 TI - High-risk HPVs and risk of cervical neoplasia: a nested case-control study. AB - The purpose of this nested case-control study was to estimate the risk of SIL development among a cohort of women providing cervical samples as part of their family planning visit at baseline in 1991-1992. All women had normal cervical cytology (N = 2905) at baseline and provided a cervical sample for subsequent HPV typing. Among this cohort, 426 women developed SIL (22 HSIL and 404 LSIL), 619 developed atypia, and 1860 remained cytologically normal. Two controls per case were sampled from those who remained normal. PCR-based methods with L1 consensus primers were used to assess high-risk HPV positivity. Having an oncogenic HPV type at baseline was associated with an almost fourfold increased risk of HSIL development (relative risk (RR) = 3.8; 95% CI, 1.5--9.0) and a 70% increased risk of LSIL development (RR = 1.7; 95% CI, 1.2--2.3%). The association between HPV positivity and SIL development was strongest in the first year of follow-up (RR = 9.2 for HSIL and 2.5 for LSIL development). The decline in HPV-associated SIL risk may be a function of having only one measure of HPV positivity (at baseline). PMID- 11263953 TI - Stage-specific alternative cplicing of CD44 and alpha 6 beta 1 integrin in colorectal tumorigenesis. AB - Recent reports suggest that cancerogenesis induces changes in alternative processing of human genes. However, little is known about the regulation of alternative splicing during malignant transformation. Therefore, we examined changes in alternative splicing of two different adhesion molecules, alpha 6 beta 1 integrin and CD44, in multiple stages of colon tumorigenesis. Using semiquantitative RT-PCR it is shown that the alternatively spliced isoforms of both adhesion molecules, alpha 6A and -B and CD44v6, are significantly upregulated in colorectal adenoma (n = 20) compared to normal colon mucosa (n = 32) (P < 0.01). Although beta1 isoforms were expressed in almost all tissues, there was a significant increase in the intensity of gene expression of beta 1A compared to beta 1B (P <0.05) in adenoma tissue. Interestingly, CD44v6 and alpha 6 variant isoforms were downregulated in carcinoma tissue (n = 28) compared to adenoma. These results establish a link between neoplastic transformation and alternative splicing of cell adhesion molecules. Furthermore, these data suggest that colon epithelial cells carrying splice variants of adhesion molecules might acquire a selective growth advantage during early tumorigenesis. PMID- 11263954 TI - Coronary heart disease, hypercholesterolemia, and atherosclerosis. I. False premises. AB - Lipid-rich caseous debris of advanced lesions stimulated interest in the role of cholesterol and lipids in atherosclerosis. Lipid-containing arterial lesions in cholesterol-overfed animals (cholesterolosis) and xanthomatous vascular lesions in subjects with familial hypercholesterolemia were then misrepresented as being atherosclerotic and led to the development of the hypercholesterolemic/lipid hypothesis. It is untenable that cholesterol, an essential multifunctional metabolite, is pathogenic at all blood levels and hypercholesterolemia is not prerequisite for human or experimental atherosclerosis. Serum cholesterol levels display a poor correlation with atherosclerosis at autopsy and with unreliable national coronary heart disease (CHD) mortality in each sex. Atherosclerosis topography and its iatrogenic production in humans and experimentally in herbivores by hemodynamic means both support a biomechanical causation and preclude causality by any circulating humoral factor. CHD, not a specific disease, is a nonspecific complication of many diseases including atherosclerosis and cannot be equated with coronary atherosclerosis due to differences in pathology and pathogenesis. Thus, extrapolations from CHD risk factors or correlations with fallacious vital statistics to atherosclerosis are invalid. It follows that the hypercholesterolemic/lipid hypothesis evolving from false premises, misuse of CHD, scientific misrepresentation, and fallacious data has no legitimate basis. PMID- 11263956 TI - Human retinal S-antigen: T cell epitope mapping in posterior uveitis patients. AB - Uveitis, an intraocular inflammatory disease which affects the uveal tract and the retina of the eye in humans, is one of the major causes of visual impairment. Posterior uveitis is often associated with inflammation of the retina and vitreous. Unfortunately, etiological diagnosis of the disease is not possible in the majority of patients. It is generally felt that an autoimmune mechanism may be involved in so-called idiopathic cases. The role of retinal S-antigen, its 20 linear peptides spanning the entire sequence, and 2 additional peptides, known to be uveitopathogenic in experimental animals, was studied in 26 patients with uveitis. Lymphocyte proliferative response was tested in vitro to identify the epitopes of S-antigen involved and to establish their role in the pathogenesis of uveitis. Of 26 uveitis patients tested, 11 showed a significant T cell proliferative response in vitro to at least 1 antigen used. None among the controls showed any response to the peptides or native S-antigen used in this study. We have found that uveitis patients respond most frequently to peptide 4 (61--80), peptide 5 (81--100), peptide 8 (141--160), peptide 9 (161--180), peptide 12 (221--240), and peptide 13 (241--260) of the human S-antigen. These results further confirm that autoimmunity to retinal S-antigen may play a role in the etiopathogenesis of a subset of patients with idiopathic uveitis. PMID- 11263955 TI - Coronary heart disease, hypercholesterolemia, and atherosclerosis. II. Misrepresented data. AB - Early development of the hypercholesterolemic/lipid hypothesis of atherosclerosis was based on false premises including fallacious national mortality rates and misrepresentation of the vascular lesions in cholesterol-overfed animals and monogenic hypercholesterolemias (MH). Nonspecific coronary heart disease (CHD) was inappropriately used as a surrogate of atherosclerosis, unmeasured and unseen. Causality was assumed and implied by classifying statistical correlates of CHD as atherogenic risk factors. These faults were compounded by methodological errors, pooling of all causes of CHD, a large clinical diagnostic error, biased age selection of cohorts leading to confounding by age and MH, and emphasis on population and cohort mean values which conceal heterogeneity within cohorts and are inapplicable to individuals. Overzealous investigators neglected to review the premises and relevant pathology on which the hypothesis was based or to reconcile valid criticisms, inconsistencies, and invalidation of CHD epidemiology by pathological, experimental, and iatrogenic evidence. Statistical data, pertaining to CHD but with no scientific applicability to atherosclerosis, progressively imparted to readers a misleading perception of the relationship of serum cholesterol to CHD. Concurrently the statistical serum cholesterol range was unjustifiably abandoned. The evidence establishes that the lipid hypothesis of atherosclerosis lacks scientific basis. PMID- 11263958 TI - Acute doxorubicin (adriamycin) dosage does not reduce cardiac protein synthesis in vivo, but decreases diaminopeptidase I and proline endopeptidase activities. AB - Anthracycline antibiotics are effective anticancer agents but their use is limited due to unwanted adverse side effects. The toxic effects of doxorubicin (adriamycin) include the development of defined cardiac lesions leading to cardiomyopathy in some patients. This has been reported to be due to reductions in cardiac protein synthesis. However, virtually all of these previous studies have failed to consider the specific radioactivity of the precursor pool in their measurements or have carried out their studies in vitro. To further resolve the above we measured fractional rates of cardiac protein synthesis using the "flooding dose" method in rats treated with adriamycin (5 mg/kg body wt). Controls were identically treated and injected with saline. At 2.5 or 24 h after adriamycin injection, rates of protein synthesis were measured with a flooding dose of l-[4-(3)H]phenylalanine. Measurements included free (S(i)) and protein bound (S(b)) phenylalanine-specific radioactivities, the protein synthetic capacity (RNA/protein ratio; C(s)), the fractional rates of protein synthesis calculated from the ratio S(b)/S(i), and the protein synthetic efficiency calculated from the ratio k(s)/C(s). Complementary analyses included assays of lysosomal (cathepsins B, D, H, and L and diaminopeptidases I and II) and cytoplasmic proteases (alanyl aminopeptidase, arginyl aminopeptidase, leucyl aminopeptidase, diaminopeptidase IV, tripeptidyl aminopeptidase, and proline endopeptidase). These enzymes constitute the most active proteases in this tissue and represent an index of protein degradation capacity in cardiac muscle. The results showed that in 2.5-h dosed rats, adriamycin had no effect on S(i), S(b), C(s), k(s), or k(RNA) (P > 0.05, not significant (NS) in all instances). In 2.5-h dosed rats, levels of diaminopeptidase I activity were reduced (P < 0.05), whereas the activities of other proteases were not significantly altered (NS in all instances). In 24-h dosed rats, adriamycin reduced cardiac S(b) (P < 0.001), which would normally be interpreted as a reduction in protein synthesis. However, S(i) was also decreased in 24-h adriamycin-injected rats (P < 0.025%). C(s) was not changed (NS). Consequently, the calculated k(s) and k(RNA) values were not significantly affected in 24-h adriamycin-dosed rats (NS). There were also significant reductions in proline endopeptidase activities in rats exposed for 24 h to adriamycin. The activities of other proteases were not significantly affected at this time point (NS in all instances). In conclusion, adriamycin reduces amino acid labeling of cardiac proteins, an effect that is a consequence of altered free phenylalanine-specific radioactivities. There was some evidence of limited altered intracellular proteolysis. PMID- 11263957 TI - Plasminogen activator and plasminogen activator inhibitor gene expression in human saphenous vein organ culture. AB - We describe the expression of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI-1) in saphenous vein culture. Smooth muscle cells (SMC) are quiescent in fresh tissue, whereas these cells acquire a proliferating phenotype when venous segments are cultured in the presence of serum. t-PA and PAI-1 were localized immunohistochemically and quantified using biochemical techniques. t-PA and PAI-1 mRNA was quantified by reverse transcription polymerase chain reaction (RT--PCR) assay. Immunostaining showed an increase in the positivity of proliferating cell nuclear antigen (PCNA) from 10 day tissue culture. Tissue sections from fresh vein showed minimal t-PA and maximal PAI-1 immunostaining. In contrast, 10-day cultures showed an increase in t-PA and a decline in PAI-1 staining. Biochemical analysis revealed a 118% increase in t-PA and a 50% decrease in PAI-1 antigen levels from 10-day tissue cultures. RT--PCR demonstrated that the mRNA encoding t-PA increased, while PAI-1 decreased after 10 days of culture. In conclusion, venous culture showed an up regulation of t-PA and a repression of PAI-1 gene expression during SMC proliferation in the vessel wall. The PAI-1 repression observed in venous culture is in contrast to the situation observed in human atheroma. A shift in the t PA/PAI-1 balance may have a role in vascular remodeling. PMID- 11263959 TI - Mechanisms of adrenal damage induced by 7,12-dimethylbenz (alpha) anthrancene in female Sprague--Dawley rats. AB - The mechanisms of adrenal damage induced by 7,12-dimethylbenz (alpha) anthrancene (DMBA) in 50-day-old female Sprague--Dawley rats were investigated. A single dose of DMBA, either fed (30 mg) per os or injected (6 mg) in a caudal vein, caused inner cortical cell death (cells of the zonae fasciculata and reticularis) by an apoptotic mechanism. Apoptotic cells were identified by cell morphology, and terminal dUTP nick end labeling (TUNEL)-positive cells were seen at 12 hrs post DMBA, reached a maximum at 36 h, and were accompanied by blood congestion followed by massive hemorrhage leading to post-apoptotic necrosis at 48 and 72 h. The apoptotic cascade involved the up-regulation of Bax, the down-regulation of Bcl-2, and the activation of caspase-3. At 72 h, regeneration as evidenced by the appearance of 5-bromo-2'-deoxyuridine-positive cells began to occur in the damaged inner cortical zones, with the cells proliferating toward the medulla thereafter. Regenerative cells expressed cytochrome P450 11 beta hydroxylase. The damage was repaired but calcification appeared at 2 weeks post-DMBA, leaving bow shaped lesions in some adrenals. PMID- 11263960 TI - Deletion polymorphism of DNASE1L1, an X-linked DNase I-like gene, in acid maltase deficiency disorders. PMID- 11263961 TI - Chimeric synthetic peptides from the nucleocapsid p24 protein of human immunodeficiency virus type-1. AB - Two chimeric synthetic peptides incorporating antigenic sequences from N-terminal (peptide C14) (134-163) and C-terminal (peptide C15) (335-364) of the p24 protein of human immunodefiency virus (HIV- 1), were synthesized. Peptides C14-GG-C15 and C15-GG-C14 represented sequences from the p24 protein in both possible orders, separated by two glycine residues as arm spacers. These peptides were evaluated as antigen in an Ultramicroenzyme-linked immunosorbent assay (UMELISA) using sera of HIV-1-infected individuals (n = 16) with different titers of antibodies and the specificity was evaluated with healthy blood donors (n = 20). The results were compared to plates coated with monomeric peptides C14 and C15. The chimeric peptide C14-GG-C15 was the most antigenic. Those results may be related to the peptide structure, the sequence order in the chimeric peptide, and epitope accessibility to the antibodies. This chimeric peptide would be very useful for HIV-1 diagnostics. PMID- 11263963 TI - Rkp1/Cpc2, a fission yeast RACK1 homolog, is involved in actin cytoskeleton organization through protein kinase C, Pck2, signaling. AB - The Rkp1/Cpc2, a fission yeast RACK1 homolog, interacted with Pck2, one of the known PKC homologs, in vivo and in vitro. The rkp1-deletion mutants (Deltarkp1) are elongated and the pck2-deletion mutant (Deltapck2) showed abnormal morphology. The double-deletion mutant (Deltarkp1Deltapck2) showed more aberrant cell shapes and was sensitive to high salt concentration. Both Deltarkp1 and Deltapck2 cells were sensitive to latrunculin B (Lat B) which inhibits actin polymerization. The cells expressing the human RACK1 homolog complemented the latrunculin B sensitivity of Deltarkp1 indicating that human RACK1 is a functional homolog of Rkp1/Cpc2. We propose that Rkp1/Cpc2 may function as a receptor for Pck2 in the regulation of actin cytoskeleton organization during cell wall synthesis and morphogenesis of Schizosaccharomyces pombe. PMID- 11263962 TI - Different Rab GTPases associate preferentially with alpha or beta GDP dissociation inhibitors. AB - GDIs (GDP-dissociation inhibitors) bind to Rab GTPases and mediate their membrane targeting and recycling. In vitro, most Rabs can bind to either of the major isoforms of GDI, leading to the assumption that the proportion of each specific Rab/GDI complex in vivo reflects the relative abundance of the alpha versus beta forms of GDI. Here we show that when human teratocarcinoma cells (Ntera2) are induced to differentiate into postmitotic neurons (NT2N), there is a major change in the proportion of GDIalpha relative to GDIbeta. Under these conditions, certain Rab GTPases associate preferentially with either GDIalpha or GDIbeta, irrespective of the relative abundance of the GDI isoform. These findings suggest that heretofore unrecognized functional specificity may exist between the two major forms of GDI. PMID- 11263964 TI - Allelic loss mapping and physical delineation of a region harboring a thymic lymphoma suppressor gene on mouse chromosome 16. AB - Our previous mapping of allelic loss in gamma-ray induced thymic lymphomas in F(1) hybrid and backcross mice between BALB/c and MSM strains identified three regions with high frequencies of allelic loss which probably harbor a tumor suppressor gene. One region, Tlsr7, exists near the D16 Mit122 locus on chromosome 16. This study has further localized Tlsr7 by constructing a physical map and scanning a total of 587 thymic lymphomas. The map consists of 13 overlapping BAC clones and isolation of BAC-derived polymorphic probes leads to fine mapping of allelic losses. Eleven lymphomas show informative breakpoints of allelic loss regions relative to the flanking markers on the map. Pulsed-field gel electrophoresis of NotI digests of the clones shows that the commonly lost region is localized within an approximately 300 kb interval near D16Mit192. This map is invaluable to facilitate the identification of genes in the Tlsr7 region. PMID- 11263965 TI - Cloning and expression of glucose 3-dehydrogenase from Halomonas sp. alpha-15 in Escherichia coli. AB - The gene encoding glucose 3-dehydrogenase (G3DH) from Halomonas sp. alpha-15 was cloned and expressed in Escherichia coli. An open reading frame of 1686 nucleotides was shown to encode G3DH. The flavine adenine dinucleotide binding motif was found in the N-terminal region of G3DH. The deduced primary structure of G3DH showed about 30% identity to sorbitol dehydrogenase from Gluconobacter oxydans and 2-keto-d-gluconate dehydrogenases from Erwinia herbicola and Pantoea citrea. The folding prediction of G3DH suggested that the 3D structure of G3DH was similar with cholesterol oxidase from Brevibacterium sterolicum or glucose oxidase from Aspergillus niger. PMID- 11263966 TI - Probing local conformational changes during equilibrium unfolding of firefly luciferase: fluorescence and circular dichroism studies of single tryptophan mutants. AB - Firefly luciferase is a monomeric protein composed of two globular domains. There is a wide cleft between the two domains. The N-terminal domain can be further divided into A-, B-, and C-subdomains. Previous studies showed that in vitro unfolding of firefly luciferase induced by guanidinium chloride can be described as a four-state equilibrium with two inactive intermediates (Herbst, R., et al. (1997) J. Biol. Chem. 272, 7099-7105). In order to monitor spectroscopically the conformational changes that occur in the different domains and subdomains during the multi-state unfolding process, we constructed a series of single-tryptophan mutants. These mutants were purified and characterized and shown to retain essentially all of the structural properties of the wild-type luciferase. Under equilibrium conditions, the unfolding of each mutant protein were studied by means of fluorescence and circular dichroism. The results show that different conformational changes occur in specific regions, suggesting a sequential unfolding process for firefly luciferase. Under 2.5 M GdmCl, whereas the N-domain unfolds partially holding half of the secondary structure content, the C-domain unfolds almost completely. In the equilibrium intermediate I(2), the secondary structure might stem mostly from the A- and B- subdomains. PMID- 11263968 TI - L-selectin tyrosine phosphorylates cbl and induces association of tyrosine phosphorylated cbl with crkl and grb2. AB - L-Selectin-mediated rolling of leukocytes on endothelial cells is an important step for lymphocyte homing and an early event in the immune response to pathogens or inflammatory stimuli. We have previously elucidated intracellular signaling cascades upon L-selectin engagement resulting in activation of Ras, Rac and JNK as well as cytoskeletal changes, oxygen release, ceramide synthesis and receptor capping. Activation of the src-tyrosine kinase p56lck is followed by phosphorylation of the L-selectin molecule and MAP-K. Here we show a tyrosine kinase dependent phosphorylation of the Cbl adapter protein after L-selectin engagement in lymphocytes. Phosphorylation of Cbl was absent in Jurkat cells that are pharmacologically treated with tyrosine kinase inhibitors and in lck deficient JCaM cells. There is an activation induced association of tyrosine phosphorylated Cbl with Grb2 and CrkL, respectively, but not CrkII. Therefore, the adapter protein Cbl plays a role in L-selectin signaling and might modulate immune function by the specific recruitment of signaling molecules to multiprotein complexes. PMID- 11263967 TI - Expression of dominant negative form of PAX4 in human insulinoma. AB - The paired-homeodomain transcription factor PAX4 is expressed in the early pancreas, but is later restricted to beta cells and not expressed in mature islets, suggesting an important role of PAX4 in differentiation and development of pancreatic islet. Here we show that PAX4 mRNA was highly expressed in human insulinoma tissues, whereas little if any mRNA was expressed in normal islets. Furthermore, this insulinoma associated expression of PAX4 mRNA was accompanied with expression of its novel variant form (PAX4v). PAX4v was generated by alternative splicing lacking the exon 7, and containing intact paired and homeo domain followed by novel 35 amino acids. PAX4v reversed the wild-type PAX4 mediated repression of the insulin promoter in cotransfection assays. PAX4v may play a role to antagonize the wild-type PAX4 function in human insulinoma. These data imply a role of PAX4 and PAX4v expression in tumorigenesis and development of insulinoma. PMID- 11263969 TI - Involvement of a lysine residue in the active site of a thermostable xylanase from Thermomonospora sp. AB - A highly thermostable xylanase (Xyl I) produced by Thermomonospora sp. was purified to homogeneity and was classified as a family 10 xylanase based on its molecular weight (38,000 Da) and isoelectric point (4.1). K2d analysis showed that the secondary structure of Xyl I was made up of 38% alpha-helix and 10% beta sheet. The optimal temperature for the activity of Xyl I was 80 degrees C. Xyl I was highly thermostable with half-lives of 86, 30, and 15 min at 80, 90, and 100 degrees C respectively. Xyl I was stable in an expansive pH range of 5 to 10 with more than 75% residual activity. Our present investigation using o-phthalaldehyde (OPTA) as the chemical initiator for fluorescent chemoaffinity labeling and trinitrobenzenesulphonic acid (TNBS) as chemical modifier have revealed the presence of a single lysine residue in the active site of Xyl I. The high pK value for the basic limb of the pH profile reflects the ionization of a lysine residue. The higher K(m) values and similar k(cat) values of the TNBS modified enzyme in comparison to native enzyme and the substrate protection against OPTA and TNBS, suggested the presence of the lysine residue in the substrate-binding site. PMID- 11263970 TI - Skeletal muscle sodium channel is affected by an epileptogenic beta1 subunit mutation. AB - The syndrome of generalized epilepsy with febrile seizures plus type 1 (GEFS+) has been associated to the gene SCN1B coding for the sodium channel beta1 subunit (Wallace, R. H. et al. (1998) Nature Genetics 19, 366-370). In patients, a mutation of the cysteine 121 to trpyptophane (C121W) would cause a lack of modulatory activity of the beta1 subunit on sodium channels expressed in the brain, rendering neurons hyperexcitable. We have confirmed that the normal beta1 modulation of type-IIA adult brain alpha subunits (BIIA) expressed in frog oocytes is defective in C121W. We observed that the mixture of wild-type and mutant beta1 subunits is less effective than wild-type alone, suggesting that the mutant beta1 subunit does bind the alpha subunit. However, we also observed a similar lack of modulation by C121W of the in adult skeletal muscle alpha subunit (SkM1). This finding is in contrast with the simple idea that the mutational effect observed in the oocyte expression system is the principal physiopathological correlate of GEFS+, because no skeletal muscle symptoms have been reported in GEFS+ patients. We conclude that the manifestation of the pathological phenotype is conditioned by the presence of susceptibility genes and/or that the frog oocyte expression system is inadequate for the study of the mutant beta1 subunit physiopathology. PMID- 11263971 TI - Abc protein transport of MRI contrast agents in canalicular rat liver plasma vesicles and yeast vacuoles. AB - The mechanism of excretion into bile of hepatospecific magnetic resonance imaging (MRI) contrast media employed labeled Gd-reagents EOB.DTPA, BOPTA, B 20790 (iopanoate-linked), and B 21690 (glycocholate-linked) for measurement in rat liver canalicular plasma membrane vesicles and yeast vacuoles. The presence of ATP gave threefold greater transport of B 20790 and B 21690 than of EOB.DTPA and BOPTA. In yeast vacuoles the ATP stimulatory effect was eightfold with B 20790 and fivefold greater for B 21690, whereas in YCF1- or YLLO115w-deleted yeast cells the transport was significantly reduced and absent from double mutants, YCF1 and YLLO15w. The transport was similar in wild-type and deletant cells for B 21690; taurocholate gave 85% inhibition. These data suggest that bilary secretion of structurally related MRI agents depend on molecular structure. The findings are suggestive as of possible value for clinical diagnosis of inherited hyperbilirubinemias and other liver disorders. PMID- 11263972 TI - Identification of multiple gephyrin variants in different organs of the adult rat. AB - The neurotransmitter receptor anchoring protein gephyrin is encoded by a highly mosaic gene whose primary transcript is subject to extensive alternative splicing. Gephyrin mRNAs are widely expressed in various mammalian tissues, and gephyrin has been implicated in neuron-specific and general metabolic functions. Using a novel affinity isolation procedure, we report the identification of different gephyrin variants in various organs of the adult rat. In particular, polypeptides of 52, 56, 60, and 91 kDa were detected in addition to the previously characterized 93-kDa protein. Our results suggest tissue-specific functional differences between gephyrin variants. PMID- 11263973 TI - Kinase-inactive G-protein-coupled receptor kinases are able to attenuate follicle stimulating hormone-induced signaling. AB - Homologous desensitization of G-protein-coupled receptors (GPCR) is thought to occur in several steps: binding of G-protein-coupled receptor kinases (GRKs) to receptors, receptor phosphorylation, kinase dissociation, and finally binding of beta-arrestin to phosphorylated receptors and functional uncoupling of the associated Galpha protein. It has recently been reported that GRKs can inhibit Galphaq-mediated signaling in the absence of phosphorylation of some GPCRs. Whether or not comparable phosphorylation-independent effects are also possible with Galphas-coupled receptors remains unclear. In the present study, using the tightly Galphas-coupled FSR receptor (FSH-R) as a model, we observed inhibition of the cAMP-dependent signaling pathway using kinase-inactive mutants of GRK2, 5, and 6. These negative effects occur upstream of adenylyl cyclase activation and are likely independent of GRK interaction with G protein alpha or beta/gamma subunits. Moreover, we demonstrated that, when overexpressed in Cos 7 cells, mutated GRK2 associates with the FSH activated FSH-R. We hypothesize that phosphorylation-independent dampening of the FSH-R-associated signaling could be attributable to physical association between GRKs and the receptor, subsequently inhibiting G protein activation. PMID- 11263974 TI - The Zrc1 is involved in zinc transport system between vacuole and cytosol in Saccharomyces cerevisiae. AB - The ZRC1 gene encodes a multicopy suppressor of zinc toxicity in Saccharomyces cerevisiae; however, previously we found that the expression of ZRC1 was induced when the intracellular zinc level was decreased. Zrc1 has six putative transmembrane domains and we determined that a Zrc1-GFP fusion protein was localized to the vacuolar membrane. The steady state level of intracellular zinc in a zrc1Delta mutant cultured in the zinc-abundant medium was lower than that in wild type. No distinct difference was observed in the basal activity of glyoxalase I, which is a cytosolic enzyme requiring zinc for catalytic function and is used here as a marker for cytosolic zinc-availability, between wild type and zrc1Delta mutant, although the activity was decreased much greater extent in the zrc1Delta mutant if the cells were exposed to the metal-limited medium. Similarly, the basal expression level of ZRC1-lacZ reporter gene in zrc1Delta mutant was the same as that in wild type; however, the fold of induction of ZRC1 lacZ expression in zrc1Delta mutant under the zinc-limited conditions was higher than that in the wild type. Based on these results, we present a tentative model for the function of Zrc1 as a mechanism to maintain the zinc homeostasis in yeast. PMID- 11263975 TI - Cytosolic Ca(2+) signal is involved in regulating UV-induced apoptosis in hela cells. AB - Results of recent studies using BAPTA/AM have raised a serious question on whether Ca(2+) signal is truly involved in regulating the progression of apoptosis. To resolve this question, we examined the differential effects of three different Ca(2+) signaling blockers (BAPTA/AM, membrane-impermeant BAPTA, and heparin) on UV-induced apoptosis in HeLa cells. We found that although the membrane-permeable form of BAPTA (i.e., BAPTA/AM) could not inhibit cell death, the membrane-impermeant form of BAPTA, loaded into the cytosol by electroporation, clearly protected cells from entering apoptosis. Furthermore, when we injected heparin to block Ca(2+) release from the endoplasmic reticulum (ER) to cytosol, apoptosis was greatly suppressed. These findings strongly suggest that elevation of cytosolic Ca(2+) is part of the signal that drives the progression of apoptosis. The negative result of BAPTA/AM is probably due to its dual effect on subcellular Ca(2+) distribution; besides suppressing the Ca(2+) elevation in cytosol, BAPTA/AM can also enter into the ER to reduce the free Ca(2+) level there. The depletion of Ca(2+) in ER is believed to stimulate apoptosis and thus would counterbalance the protection effect of BAPTA/AM in suppressing the cytosolic Ca(2+) elevation. PMID- 11263977 TI - Molecular cloning and expression of the novel splice variants of K(+) channel interacting protein 2. AB - Two cDNAs encoding the splice variants of K(+) channel-interacting protein 2 (KChIP2) recently reported as human KChIP2 have been identified from rat, mouse, and human heart by RT-PCR. A longer variant, KChIP2L encodes a protein of 270 amino acids, which has a 50-amino-acid insertion in N-terminus in comparison with a shorter one, KChIP2S. Interestingly, both KChIP2S and KChIP2L (KChIP2S/L) but not the original KChIP2 were expressed in human heart and umbilical vein endothelial cells (HUVECs). KChIP2S transcripts but not KChIP2L were predominantly expressed in rat, mouse, and human heart and HUVECs, whereas both transcripts were expressed at low levels in other tissues such as brain, aorta, and kidney. Using chimeric proteins of green fluorescence protein (GFP) fused to the N-terminus of KChIP2S/L, the interactions between Kv4.3 and KChIP2S/L were analyzed in native and Kv4.3-expressed HEK293 cells. Specific localization of GFP fused KChIP2S/L proteins on or near cell membrane was observed only in Kv4.3 expressed HEK293 cells. PMID- 11263976 TI - Soluble apyrases release adp during ATP hydrolysis. AB - A soluble form of CD39 was expressed and purified from High-Five insect cells. The soluble CD39 is a monomer with a molecular weight of 54,000. The k(cat) and K(m) of the purified soluble CD39 were 4.6 s(-1) and 12 microM for ATP and 1.3 s( 1) and 7 microM for ADP as substrates, respectively. One nucleotide binding site was detected on the monomer only in the presence of Ca(2+). In contrast to the membrane bound CD39, soluble CD39 released ADP as an intermediate during ATP hydrolysis, as did the soluble potato apyrase. PMID- 11263979 TI - Efficient separation of Thermus aquaticus EF-Tu functional complexes. AB - A new method for fast separation of the main functional complexes of the elongation factor Tu from Thermus aquaticus has been developed. Binary complexes EF-Tu * GDP and EF-Tu * GDPNP as well as the ternary complex EF-Tu * GDPNP * Leu approximately tRNA were separated from each other by means of HPLC on a hydrophobic sorbent TSK-Gel Phenyl 5PW in a reverse gradient of ammonium sulfate. This technique is suitable for monitoring EF-Tu activity, characterisation of the ratio between different EF-Tu forms in cell extracts, and isolation of individual EF-Tu complexes for structural and functional investigations. In order to illustrate the potentials of the method, we used HPLC on a TSK-Gel Phenyl 5PW matrix to determine the ratio between affinities of GDP and GDPNP for EF-Tu. We found that K(a)(GDP) is about 27 times higher than K(a)(GDPNP) at 37 degrees C, the value being close to the one reported for Thermus thermophilus EF-Tu. PMID- 11263978 TI - Distribution of cellular isoform of prion protein in T lymphocytes and bone marrow, analyzed by wild-type and prion protein gene-deficient mice. AB - In this study, the authors investigated normal cellular prion protein (PrP(C)) expression on murine immune systems using prion protein gene-deficient mouse as negative control. Immunocytes expressing PrP(C) in adult and fetal mice were detected by flow cytometry with the monoclonal antibody against PrP(C), 6H4. Cells from thymus and bone marrow reacted positively with 6H4, while spleen cells, peritoneal cells, peripheral blood leukocytes, and intestinal intraepithelial lymphocytes were nonreactive. In thymus, PrP(C) was observed in CD4(-)CD8(-) double-negative thymocytes. PrP(C+) cells of double-negative thymocytes belonged to the CD3(-) subset, but not to the CD3(+) subset. Triple negative PrP(C+) thymocytes expressed CD44 or CD25 antigens. Furthermore, PrP(C) was observed in c-kit(+) bone marrow cells. In fetuses, PrP(C+) cells were observed in the liver and thymus at day 16.0 and 15.0 of gestation, respectively. These results demonstrated that PrP(C) is expressed on immature immunocytes. PMID- 11263980 TI - Molecular cloning and characterization of MFRP, a novel gene encoding a membrane type Frizzled-related protein. AB - The Frizzled-type cysteine-rich domain (CRD) is a binding motif for soluble-type glycoprotein WNTs, which play key roles in embryogenesis and carcinogenesis. Here, we have cloned and characterized a novel gene MFRP, encoding a type II transmembrane protein with CRD. In addition to CRD, two tandem-repeats containing the Cubilin domain approximately the MFRP domain were present in the extracellular region of MFRP. Although MFRP was homologous to Corin, FZDs, and SFRPs in CRD, amino-acid identities between CRD in MFRP and CRDs in these molecules were less than 40%. The MFRP gene on 11q23 consisted of at least 13 exons. The 4.0-kb MFRP was not detected by Northern blot analysis in normal tissues other than adult and fetal brain. The MFRP mRNA was undetectable in seven gastric cancer cell lines, seven brain tumor cell lines, and other eight tumor cell lines. Regional distribution of the MFRP mRNA in human brain was further investigated, and MFRP was found to be expressed strongly in medulla oblongata, and weakly in hippocampus and corpus callosum. Thus, MFRP with CRD might play key roles in medulla oblongata as a regulator of the WNT signaling pathway. PMID- 11263981 TI - Molecular cloning, genomic organization, and expression of a C-type (Manduca sexta-type) allatostatin preprohormone from Drosophila melanogaster. AB - The insect allatostatins are a diverse group of neuropeptides that obtained their names by their inhibitory actions on the corpora allata (two endocrine glands near the insect brain), where they block the biosynthesis of juvenile hormone (a terpenoid important for development and reproduction). Chemically, the allatostatins can be subdivided into three different peptide groups: the large group of A-type (cockroach-type) allatostatins, which have the common C-terminal sequence Y/FXFGLamide; the B-type (cricket-type) allatostatins, which have the C terminal sequence W(X(6))Wamide in common; and a single allatostatin that we now call C-type allatostatin that was first discovered in the moth Manduca sexta, and which has a nonamidated C terminus, and a structure unrelated to the A- and B type allatostatins. We have previously cloned the preprohormones for the A- and B type allatostatins from Drosophila melanogaster. Here we report on the cloning of a Drosophila C-type allatostatin preprohormone (DAP-C). DAP-C is 121 amino acid residues long and contains one copy of a peptide sequence that in its processed form has the sequence Y in position 4) from the Manduca sexta C-type allatostatin. The DAP-C gene has three introns and four exons and is located at position 32D2-3 on the left arm of the second chromosome. Northern blots show that the gene is strongly expressed in larvae and adult flies, but less in pupae and embryos. In situ hybridizations of larvae show that the gene is expressed in various neurons of the brain and abdominal ganglia and in endocrine cells of the midgut. This is the first publication on the structure of a C-type allatostatin from insects other than moths and the first report on the presence of all three types of allatostatins in a representative of the insect order Diptera (flies). PMID- 11263982 TI - Identification and structure of the nerve growth factor binding site on TrkA. AB - Nerve growth factor (NGF) is involved in the development and maintenance of the nervous system and has been implicated as a possible therapeutic target molecule in a number of neurodegenerative diseases, especially Alzheimer's disease. NGF binds with high affinity to the extracellular region of a tyrosine kinase receptor, TrkA, which comprises three leucine-rich motifs (LRMs), flanked by two cysteine-rich clusters, followed by two immunoglobulin-like (Ig-like) domains. We have expressed the second Ig-like domain as a recombinant protein in E. coli and demonstrate that NGF binds to this domain with similar affinity to the native receptor. This domain (TrkAIg(2)) has the ability to sequester NGF in vitro, preventing NGF-induced neurite outgrowth, and in vivo, inhibiting NGF-induced plasma extravasation. We also present the three-dimensional structure of the TrkAIg(2) domain in a new crystal form, refined to 2.0 A resolution. PMID- 11263983 TI - Endoglin expression in human and rat mesangial cells and its upregulation by TGF beta1. AB - Endoglin is a component of the TGF-beta receptor complex present in the kidney at the human glomerular mesangium. Since the cellular origin of the glomerular endoglin is unknown, in the present study we investigated the expression of endoglin in mesangial cells in culture, as well as their response to TGF-beta1. Western and Northern blot analysis identified the expression of endoglin protein and mRNA transcript in both human and rat mesangial cells. Flow cytometry and immunocytochemistry analyses revealed that endoglin is present on the cell membrane. Exogenous TGF-beta1 stimulated not only the expression of collagen alpha1 (I) I and TGF-beta1, but also that of endoglin. These data provide the first evidence for the expression of endoglin in mesangial cells, as well as its upregulation by TGF-beta1, thus suggesting that endoglin may have a role in modulating the effects of TGF-beta1 on the glomerular mesangium. PMID- 11263984 TI - In vitro differentiation of human marrow stromal cells into early progenitors of neural cells by conditions that increase intracellular cyclic AMP. AB - Human marrow stromal cells (hMSCs) are multipotential stem cells that can be differentiated into bone, cartilage, fat, and muscle. In the experiments here, we found that undifferentiated cultures of hMSCs express some markers characteristic of neural cells such as microtubule-associated protein 1B (MAP1B), neuron specific tubulin (TuJ-1), neuron-specific enolase (NSE), and vimentin. By treating hMSCs with 0.5 mM isobutylmethylxanthine (IBMX)/1 mM dibutyryl cyclic AMP (dbcAMP) for 6 days, about 25% of the hMSCs differentiated into cells with a typical neural cell morphology and with increased levels of both NSE and vimentin. The data suggested that the hMSCs may have been differentiated into early progenitors of neural cells in vitro under conditions that increase the intracellular level of cAMP. PMID- 11263985 TI - Activation-dependent surface expression of LOX-1 in human platelets. AB - Lectin-like oxidized LDL receptor-1 (LOX-1) was initially identified as an oxidized LDL receptor in aortic endothelial cells. Here we identified LOX-1 mRNA and protein in human platelets in addition to recent findings on the expression in macrophages and smooth muscle cells. The presence of LOX-1 was further confirmed in the megakaryocytic cell lines. Flow cytometric analyses revealed that LOX-1 was exposed on the surface of platelets in an activation-dependent manner. Consistently, the activation-dependent binding of OxLDL to platelets was mostly inhibited by anti-LOX-1 antibody. Immunohistochemistry of the atherosclerotic plaque from a patient with unstable angina pectoris (UAP) revealed accumulation of LOX-1 protein at the site of thrombus. As LOX-1 recognizes and binds activated platelets, exposure of LOX-1 on activated platelets surface might assist thrombosis formation. PMID- 11263987 TI - Galectin-1 is a component of neurofilamentous lesions in sporadic and familial amyotrophic lateral sclerosis. AB - In amyotrophic lateral sclerosis (ALS), abnormal accumulation of neurofilaments induces pathological changes such as axonal spheroids, cord-like neurite swellings, and perikaryal conglomerate inclusions in degenerating motor neurons of the spinal cord, and the accumulation seems to cause motor neuron degeneration in this disease. Such ALS lesions were intensely labeled with HepSS-1, a monoclonal antibody to heparan sulfate. Since the identification of HepSS-1 immunoreactive substance seems to be an important step for understanding the molecular pathology of ALS, we purified the substance from human spinal cord tissue to homogeneity. Amino acid sequence of the protein was consistent with that of galectin-1. Immunohistochemistry using antibodies against recombinant human galectin-1 showed that galectin-1 was accumulated in these lesions in ALS. Although HepSS-1 was believed to be specific for heparan sulfate, it reacted with recombinant human galectin-1 which has no heparan sulfate moiety. The results show that galectin-1 is a component of the neurofilamentous lesions in ALS. Since galectin-1 has axonal regeneration-enhancing activity, the abnormal accumulation of galectin-1 to the lesions seems to be related to the pathological process of ALS. PMID- 11263986 TI - Effect of hyperthermia on premature intracellular trypsinogen activation in the exocrine pancreas. AB - Hyperthermia, raising the body temperature from normal to above 40 degrees C, has been shown to prevent pancreatitis in an experimental animal model of the disease, but the underlying cellular mechanisms of this protection remain unknown. We induced controlled hyperthermia in either laboratory rats and isolated pancreatic acini or, alternatively, raised the temperature of pancreatic homogenates in vitro from 37 to 41 degrees C. In vitro controlled hyperthermia of up to 41 degrees C increased the autoactivation-induced and enterokinase-induced trypsinogen activation as well as free trypsin activity. Conversely, in whole animal studies and in living acinar cells hyperthermia reduced or abolished premature intracellular trypsinogen activation in a time- and temperature dependent manner and this protective effect was independent of either de novo protein synthesis, interference with acinar cell signal transduction, or confirmational changes in pancreatic trypsinogen. We conclude that hyperthermia, in a manner that is independent of the synthesis of pancreatic chaperone or heat shock proteins, can directly abolish the earliest initiating event involved in the onset of pancreatitis, namely the premature and intracellular activation of digestive zymogens. PMID- 11263988 TI - Efficient gene delivery into dendritic cells by fiber-mutant adenovirus vectors. AB - Recent studies have demonstrated the usefulness of dendritic cells (DCs) genetically modified by adenovirus vectors (Ad) to immunotherapy, while sufficient gene transduction into DCs is required for high doses of Ad. The RT PCR analysis revealed that the relative resistance of DCs to Ad-mediated gene transfer is due to the absence of Coxsackie-adenovirus receptor expression, and that DCs expressed adequate alpha(v)-integrins. Therefore, we investigated whether fiber-mutant Ad containing the Arg-Gly-Asp (RGD) sequence in the fiber knob can efficiently transduce and express high levels of the LacZ gene into DCs. The gene delivery by fiber-mutant Ad was more efficient than that by conventional Ad in both murine DC lines and normal human DCs (NHDC). Furthermore, NHDC transduced with fiber-mutant Ad and conventional Ad at 8000-vector particles/cell resulted in a 70-fold difference in beta-galactosidase activity. We propose that alpha(v)-integrin-targeted Ad is a very powerful tool with which to implement DC based vaccination strategies. PMID- 11263989 TI - Accumulation of LOX-1 ligand in plasma and atherosclerotic lesions of Watanabe heritable hyperlipidemic rabbits: identification by a novel enzyme immunoassay. AB - LOX-1 (lectin-like oxidized LDL receptor) is a newly identified cell surface receptor for oxidized LDL mainly expressed in endothelial cells. Recombinant soluble LOX-1(LOX-Fc) was generated by fusing the extracellular domain of LOX-1 with the Fc portion of IgG. A novel sandwich enzyme immunoassay specific for LOX 1 ligand is designed, using LOX-Fc and anti-apoB antibody. This immunoassay was used to determine LOX-1 ligand activity in normal and Watanabe heritable hyperlipidemic (WHHL) rabbit plasma. LOX-Fc was further applied for staining of atherosclerotic lesions of WHHL rabbits. LOX-1 ligand levels were significantly elevated in the plasma of hyperlipidemic rabbits compared with controls. Furthermore, LOX-1 ligand activity was detected in the atherosclerotic lesions in situ. These results support the potential roles of LOX-1 interacting with its ligand in the pathogenesis of atherosclerosis, which is enhanced in hyperlipidemia. PMID- 11263990 TI - Hsp27-induced MMP-9 expression is influenced by the Src tyrosine protein kinase yes. AB - The small heat shock protein hsp27 is associated with aggressive tumor behavior in certain subsets of breast cancer patients. Previously we demonstrated that hsp27 overexpression in breast cancer cells increased in vitro and in vivo invasiveness, suggesting that hsp27 influences the metastatic process. To investigate this role for hsp27, we have utilized MDA-MB-231 breast cancer cells that overexpress hsp27 and cDNA expression array technology. We demonstrate that hsp27 overexpression up-regulates MMP-9 expression and activity and down regulates Yes expression. Furthermore, our results suggest that Yes may be involved in regulating MMP-9 expression, as well as in vitro invasion. Reconstitution of Yes expression by transfection into hsp27-overexpressing cells decreased MMP-9 expression, and increased in vitro invasiveness, abrogating the phenotype conferred by hsp27 overexpression. Therefore, our results provide a new potential mechanism by which hsp27 affects the metastatic cascade-through regulation of MMP-9 and Yes expression. PMID- 11263991 TI - SNARE proteins are critical for regulated exocytosis of ECP from human eosinophils. AB - The SNARE hypothesis, describing a protein assembly-disassembly pathway, was recently proposed for the sequential steps of synaptic vesicle docking, activation, and fusion. To determine if SNARE proteins are involved in regulated exocytosis in eosinophils, the presence and functional role of SNAREs was examined in human blood eosinophils. Immunoblotting, subcellular fractionation, and immunocytochemistry documented that vesicle-associated membrane protein-2 (VAMP-2), a vesicle-SNARE, was expressed in human eosinophils. Syntaxin 4 and SNAP-25 were also detected. Sequencing of cloned RT-PCR products amplified from a domain conserved among VAMP isoforms revealed identity only to VAMP-2 but not to VAMP-1 or cellubrevin. Functional experiments revealed that tetanus toxin pretreatment, which cleaved VAMP-2 in eosinophils, significantly inhibited both IgE receptor- and phorbol ester-mediated exocytosis of eosinophil cationic protein (ECP) from streptolysin-O-permeabilized eosinophils. Thus, these results strongly suggest a critical role of SNAREs in regulated exocytosis in eosinophils. PMID- 11263992 TI - Genes highly expressed in the early phase of murine graft-versus-host reaction. AB - Graft-versus-host reaction (GVHR) is a complex process initiated upon allorecognition. For detection of early molecular events in GVHR, we first assessed time courses with respect to symptoms and serum interferon (IFN)-gamma levels and then used the differential display method to compare gene transcript patterns during the early phase between acute lethal GVHR mice and syngeneic controls. In the GVHR mice, high expression levels of seven genes encoding the following molecules were detected: TGTP/Mg21 (an IFN-gamma-related signaling molecule), vitronectin, Nedd5 (a mammalian septin), manganese superoxide dismutase, activin betaC subunit, PRCC (a papillary renal cell carcinoma associated molecule), and an uncharacterized gene corresponding to a mouse expressed sequence tag (EST). The expression levels of most genes peaked before the symptomatological onset and the peak of IFN-gamma levels. Thus, gene expression monitoring may characterize the inductive process of GVHR and aid in the development of gene-based diagnostics and therapies. PMID- 11263993 TI - A novel homologue of the TIAP/m-survivin gene. AB - The inhibitor of apoptosis (IAP) proteins comprise a highly conserved gene family that prevents cell death in response to a variety of stimuli. TIAP/m-survivin, a murine homologue of human Survivin, is a member of the IAP family. TIAP/m survivin has one baculovirus IAP repeat (BIR) and lacks a C-terminal RING finger motif. Here we identified the genomic DNA region (TIAP-2) that is homologous to the TIAP/m-survivin gene by a low stringency genomic DNA hybridization. The region is on the chromomsome 9 which is distinct from that (chromosome 11) of the TIAP/m-survivin gene, and contains DNA sequence similar to a part of the BIR and the 3' side of the TIAP/m-survivin gene and the sequence homology between them is 92%. Expression of TIAP-2 mRNA was detected in various murine tissues by RT-PCR. Although expression of TIAP/m-survivin mRNA is upregulated in synchronized cells at S to G2/M phase of the cell cycle, expression of TIAP-2 mRNA was constant in the cell cycle, suggesting the different role of TIAP-2 from that of TIAP/m survivin. PMID- 11263994 TI - Expression of chloride channel, ClC-5, and its role in receptor-mediated endocytosis of albumin in OK cells. AB - By using Western blot and RT-PCR analyses, the expression of ClC-5, a member of the ClC family of voltage-gated chloride channels, and its mRNA was detected in OK cells. The effect of chloride channel inhibitors on receptor-mediated endocytosis of albumin was examined in OK cells and compared to that of vacuolar H(+)-ATPase inhibitors. Accumulation of fluorescein-isothiocyanate (FITC) albumin, a receptor-mediated endocytosis marker, was inhibited by 5-nitro-2-(3 phenylpropylamino)-benzoic acid (NPPB), a chloride channel inhibitor, in a concentration-dependent fashion. In contrast, uptake of FITC-inulin, a fluid phase endocytosis marker, was not affected by NPPB. Other chloride channel inhibitors, 4,4'-diisothiocyanatostilbene-2-2'-disulfonic acid and diphenylamine 2-carboxylic acid, also inhibited FITC-albumin uptake. NPPB, as well as a vacuolar H(+)-ATPase inhibitor bafilomycin A(1), caused a decrease in the affinity and in the maximal velocity of FITC-albumin uptake. These results suggest that chloride channel, most likely ClC-5, plays an important role in the receptor-mediated endocytosis of albumin in OK cells. PMID- 11263995 TI - Toward the three-dimensional structure of the Escherichia coli DNA-binding protein H-NS: A CD and fluorescence study. AB - The DNA-binding protein H-NS compacts DNA and acts as a specific transcription factor regulating the expression of various bacterial genes. The small abundant protein binds to curved DNA without apparent sequence specificity and the exact nature of its DNA interaction is still unknown. H-NS lacks any common DNA-binding or oligomerization motif and except for a C-terminal fragment of the protein no high resolution structural information is available today. Since the complete structure of H-NS is of considerable interest for understanding its versatile regulatory features, and in lack of high-resolution data for the complete molecule, we have combined circular dichroism (CD) and fluorescence measurements to collect secondary- and higher-order structural information on H-NS. Comparison of CD analyses of wild type H-NS and functional defective mutants allowed assigning secondary structure elements to the N-terminal oligomerization domain of the protein. Moreover, according to fluorescence energy-transfer data we calculate a 45 A distance between the DNA-binding and the oligomerization domain of H-NS. PMID- 11263996 TI - Polyamine cytotoxicity in the presence of bovine serum amine oxidase. AB - The toxicity of extracellular spermine, determined in the presence of fetal calf serum, was studied using three cell lines: FM3A, L1210, and NIH3T3 cells. Amine oxidase in fetal calf serum produces aminodialdehyde generating acrolein spontaneously, H(2)O(2), and ammonia from spermine. Spermine toxicity was prevented by aldehyde dehydrogenase, but not by catalase. Similar concentrations of spermine and acrolein were needed to produce toxicity. Other aldehydes (formaldehyde, acetaldehyde, and propionaldehyde) and hydrogen peroxide were less toxic than acrolein. Spermidine and 3-aminopropanal, which produces acrolein, also exhibited severe cytotoxicity. The degree of cytotoxicity of spermine, spermidine, and 3-aminopropanal was nearly parallel with the amount of acrolein produced from each compound. Thus, it was deduced that acrolein is a major toxic compound produced from polyamines (spermine and spermidine) by amine oxidase. PMID- 11263997 TI - Proline suppresses Rubisco activity by dissociating small subunits from holoenzyme. AB - Proline caused irreversible inhibition (involving reduction in V(max) without altering K(m) for RuBP) in Rubisco activity. Proline-induced suppression in Rubisco activity did not exceed beyond approximately 65% of the original activity even upon exposure to higher levels of proline for prolonged duration. However, NaCl-induced reduction in Rubisco activity was reversible. Native PAGE analysis of Rubisco-incubated with proline showed the presence of two distinct bands corresponding to approximately 430 and approximately 28 kDa, but that incubated with NaCl showed a single band. SDS-PAGE analysis revealed that the approximately 430- and approximately 28-kDa bands represent octamers of large subunits and dimers of small subunits, respectively. These results demonstrated for the first time that proline suppresses Rubisco activity by bringing about dissociation of the small subunits from the octamer core of large subunits, probably by weakening hydrophobic interactions between them. PMID- 11263998 TI - Intact LTP and fear memory but impaired spatial memory in mice lacking Ca(v)2.3 (alpha(IE)) channel. AB - To investigate the functional roles of the Ca(v)2.3 (alpha(1E)) channel in hippocampal CA1 pyramidal neurons, we studied in vitro synaptic properties and in vivo behaviors of the Ca(v)2.3 gene deficient mice. The Ca(v)2.3 channel mRNA was identified in the hippocampal formation of the wild-type mouse by in situ hybridization. The basic excitatory synaptic transmission and long-term potentiation by theta-burst stimulation were intact in CA1 region of Ca(v)2.3-/- mice. We performed two forms of behavioral tests to examine the hippocampus dependent function, i.e., emotional and spatial learning tests. The Ca(v)2.3-/- mice were able to establish and maintain fear memories. Although general improvement in the performance of Morris water maze test was seen in Ca(v)2.3-/- mice, they displayed an obvious impairment in the probe test. These results suggest that the Ca(v)2.3 channel plays some role in formation of the accurate spatial memory but not of the fear memory. PMID- 11263999 TI - Genomic and functional characteristics of novel human pancreatic 2P domain K(+) channels. AB - We isolated three novel 2P domain K(+) channel subunits from human. The first two subunits, TALK-1 and TALK-2, are distantly related to TASK-2. Their genes form a tight cluster of 25 kb on chromosome 6p21.1-p21.2. The corresponding channels produce quasi-instantaneous and non-inactivating currents that are activated at alkaline pHs. These currents are sensitive to Ba(2+), quinine, quinidine, chloroform, halothane, and isoflurane but are not affected by TEA, 4-AP, Cs(+), arachidonic acid, hypertonic solutions, agents activating protein kinases C and A, changes of internal Ca(2+) concentrations, and by activation of G(i) and G(q) proteins. TALK-1 is exclusively expressed in the pancreas. TALK-2 is mainly expressed in the pancreas, but is also expressed at a lower level in liver, placenta, heart, and lung. We also cloned a third subunit, named hTHIK-2 which is present in many tissues with high levels again in the pancreas but which could not be functionally expressed. PMID- 11264001 TI - Exercise decreases cytosolic aconitase activity in the liver, spleen, and bone marrow in rats. AB - Effects of strenuous exercise on cytosolic aconitase activity (CAA) were investigated in this study. Female Sprague-Dawley rats were randomly assigned to four groups: S1 (Sedentary), S2 (Sedentary + L-NAME [N-nitro-l-arginine methyl ester]), E1 (Exercise), and E2 (Exercise + L-NAME). Rats in the E1 and E2 groups swam for 2 h/day for 3 months. L-NAME (an inhibitor of NOS) in drinking water (1 mg/ml) was administered to rats in the S2 and E2 groups for the same period. At the end of the third month, the CAA in the liver, spleen, and bone marrow cells was measured. In the exercise group (E1), CAA in the liver, spleen, and bone marrow cells was 19.99 +/- 1.49, 1.61 +/- 0.13, and 0.59 +/- 0.09 mU/mg protein, respectively. These values were significantly lower than the corresponding sedentary values in the S1 group (33.96 +/- 1.38, 3.96 +/- 0.19, and 3.20 +/- 0.18 mU/mg protein) (P < 0.01, 0.001, and 0.001, respectively). The treatment of L-NAME led to a significant increase in tissue CAA in the sedentary rats (S2). Also, the significantly higher CAA in the liver, spleen, and bone marrow cells was found in the exercised rats treated with L-NAME (E2) (29.50 +/- 1.27, 2.89 +/ 0.25, and 1.34 +/- 0.20 mU/mg) than without L-NAME (E1) (P < 0.01, 0.01, 0.05, respectively). However, the values in the E2 group were still significantly lower than those in the S1 group (P < 0.05, 0.01, and 0.001, respectively). This indicates that L-NAME treatment can partly recover the decreased CA in tissues in the exercised rats. These results provide evidence for the existence of the increased activity of IRP1 (iron regulatory protein 1) that is probably induced by the increased nitric oxide production in the strenuously exercised rats. PMID- 11264000 TI - Reactive oxygen species-related induction of multidrug resistance-associated protein 2 expression in primary hepatocytes exposed to sulforaphane. AB - Expression of multidrug resistance-associated protein 2 (MRP2), an efflux pump contributing to biliary secretion of xenobiotics, was investigated in primary rat and human hepatocytes exposed to sulforaphane, a naturally-occurring chemopreventive agent. Northern blot indicated that sulforaphane increased MRP2 mRNA levels in primary rat hepatocytes; it also induced expression of drug metabolizing enzymes such as glutathione S-transferase A1/2 isoforms and NAD(P)H:quinone oxidoreductase in a dose-response and time-course manner similar to that observed for the upregulation of MRP2 transcripts. This sulforaphane related increase of MRP2 mRNAs paralleled increased expression of 190 kD MRP2 protein as assessed by Western blotting; it was fully abolished by the transcription inhibitor actinomycin D. MRP2 induction was associated with increased cellular production of reactive oxygen species (ROS) and addition of dimethyl sulfoxide, that reduced sulforaphane-related formation of ROS, and also decreased MRP2 mRNA levels in sulforaphane-treated primary rat hepatocytes; this suggests that sulforaphane-mediated production of ROS may contribute to MRP2 induction. This link between ROS and MRP2 regulation was further supported by the increase of MRP2 expression occurring in response to t-butylhydroquinone, known to regulate drug metabolizing enzymes through ROS formation. In addition to rat cells, primary human hepatocytes exposed to sulforaphane also displayed induced MRP2 expression evidenced at both mRNA and protein levels. All these observations strongly support the conclusion that the export pump MRP2 can be classified among the detoxifying proteins that are regulated by sulforaphane and that are thought to contribute, at least in part, to its anticarcinogenic properties. PMID- 11264002 TI - Apaf-1XL is an inactive isoform compared with Apaf-1L. AB - Apaf-1 plays a crucial role in the cytochrome c/dATP-dependent activation of caspase-9 and -3. We found that the human myeloid leukemic K562 cells were more resistant to cytochrome c-induced activation of caspase-9 and -3 in a cell-free system compared with the human T-lymphoblastic subclone CEM/VLB(100) cells. Apaf 1 cDNA sequencing revealed an additional insert of 11 aa between the CARD and CED 4 (ATPase) domains in K562 cells, which was identical to the sequence of Apaf 1XL. Immunoprecipitation of Apaf-1 with caspase-9 after a cell-free reaction demonstrated that Apaf-1XL in the K562 cell line showed a lower binding ability to caspase-9 compared with Apaf-1L protein. The resistance of K562 cells to cytochrome c-dependent apoptosis may be partly due to this Apaf-1XL form. These results suggest that the additional insert between CARD and CED-4 domains might affect Apaf-1 recruitment of caspase-9 during apoptosis. PMID- 11264003 TI - Uncoupling protein-2 participates in cellular defense against oxidative stress in clonal beta-cells. AB - The role of uncoupling protein-2 (UCP-2) in beta-cells is presently unclear. We have tested the notion that UCP-2 participates in beta-cell defense against oxidants. Expression of the UCP-2 gene in clonal beta-cells (INS-1) was decreased by 45% after 48 h of culture with vitamin E and selenite. When INS-1 cells were exposed to 200 microM H(2)O(2) for 5 min, the cell viability (MTT assay) decreased to 85 +/- 1, 61 +/- 1, 40 +/- 2, and 39 +/- 2% of control when measured respectively 30 min, 2 h, 6 h, and 16 h after H(2)O(2) exposure. At corresponding time points UCP-2 mRNA levels were 1.01 +/- 0.09, 1.53 +/- 0.15 (P < 0.05), 1.44 +/- 0.18 (P = 0.06), and 1.12 +/- 0.09 fold of control, i.e., transiently increased. We next tested whether overexpression of UCP-2 could enhance resistance of beta-cells toward H(2)O(2) toxicity. A cotransfection method using EGFP as a suitable marker and a human cDNA UCP-2 construct was used for transient overexpression of UCP-2. Transfected cells expressed the gene about 30-fold more than normal cells. After exposure to H(2)O(2) (200 micrometer, 5 min), the survival of UCP-2 overexpressing cells was measured 30-45 min later by flow cytometry. Survival was 13 +/- 0.05% higher than control (EGFP only) cells, P < 0.004 for difference. The results indicate that oxidative stress induces UCP-2 expression in beta-cells, and that UCP-2 serves a role in beta-cell defense against oxidative stress. PMID- 11264004 TI - Osteoclast differentiation factor modulates cell cycle machinery and causes a delay in s phase progression in RAW264 cells. AB - Osteoclast differentiation factor (ODF) induces differentiation of mouse RAW264 cells to mature osteoclasts. To understand the mechanism controlling a coupling between withdrawal from the cell cycle and differentiation, we examined cell cycle progression and expression profiles of cell cycle regulatory genes at the initial phase in committed cells. ODF rapidly converted the hyperphosphorylated form of the retinoblastoma protein (pRb) into the hypophosphorylated form. The p21 protein was induced by ODF treatment in the same time course with that of dephosphorylation of pRb, followed by a sharp decline. After this period, a delayed entry of the S phase started accompanying the induction of CycD3 and cdk6 in differentiating cells. Hydroxyurea treatment indicated that the S phase entry was a prerequisite for osteoclast formation. Thus, ODF induces pleiotropic effects on cell cycle regulatory genes in RAW264 cells during the initial phase of the differentiation process to osteoclasts. PMID- 11264005 TI - Asymmetric arrangement of auxiliary subunits of skeletal muscle voltage-gated l type Ca(2+) channel. AB - Highly purified L-type Ca(2+) channel complexes containing all five subunits (alpha(1), alpha(2), beta, gamma, and delta) and complexes of alpha(1)-beta subunits were obtained from skeletal muscle triad membranes by three-step purification and by 1% Triton X-100 treatment, respectively. Their structures and the subunit arrangements were analyzed by electron microscopy. Projection images of negatively stained Ca(2+) channels and alpha(1)-beta complexes were aligned, classified and averaged. The alpha(1)-beta complex showed a hollow trapezoid shape of 12 nm height. In top view, four asymmetric domains surrounded a central depression predicted to form the channel pore. The complete Ca(2+) channel complex exhibited the cylindrical shape of 20 nm in height binding a spherical domain on one edge. Further image analysis of higher complexes of the Ca(2+) channel using a monoclonal antibody against the beta subunit showed that the alpha(1)-beta complex forms the non-decorated side of the cylinder, which can traverse the membrane from outside the cell to the cytoplasm. Based on these results, we propose that the Ca(2+) channel exhibits an asymmetric arrangement of auxiliary subunits. PMID- 11264006 TI - 5-Fluorouracil suppression of NF-KappaB is mediated by the inhibition of IKappab kinase activity in human salivary gland cancer cells. AB - We have recently shown that 5-Fluorouracil (5-FU) suppresses the transcription factor NF-kappaB in human salivary gland cancer cells (cl-1) by mediating upregulation of IkappaB-alpha expression. However, the precise mechanism involved in this action has not yet been elucidated. IkappaB kinases (IKK-alpha and IKK beta) are the key components of the IKK complex that mediates activation of NF kappaB in response to external stimuli such as cytokines. In addition, NF-kappaB inducing kinase (NIK) and mitogen-activated protein kinase kinase kinase 1 (MEKK 1), both of which are the upstream kinases for the IKKs, interact with and activate the IKKs. Thus, we investigated the molecular mechanisms involved in the suppression of NF-kappaB by 5-FU. Although 5-FU did not affect the expression levels of IKKs, NIK, or MEKK-1, IKK activity in cl-1 cells was suppressed at both 6 h and 12 h after treatment with 2 microgram/ml 5-FU. Moreover, when cells were treated with various concentrations of 5-FU for 12 h, the concentration of 2 microgram/ml efficiently inhibited the IKK activity as compared to 1, 5, or 10 microgram/ml. The expression of Fas-associated death domain-like interleukin 1 converting enzyme-inhibitory protein (FLIP), which acts as an inhibitor of an initiator caspase (caspase-8), was down-regulated by 5-FU treatment in cl-1 cells. Apoptosis, as evidenced by cleavage of poly(ADP-ribose) polymerase through the action of an executioner caspase (caspase-3), was also clearly observed. Thus, these results suggest that 5-FU induction of apoptosis in cl-1 cells may be mediated by suppression of NF-kappaB via inhibition of IKK activity. PMID- 11264007 TI - Microelectrospray ionization analysis of noncovalent interactions within the electron transferring flavoprotein. AB - Cofactor associations within the electron transferring flavoprotein (ETF) were studied in real time using microelectrospray ionization-mass spectrometry (muESI MS). Initial analysis of porcine (pETF) and human ETF (hETF) revealed only the holoprotein. When muESI-MS source energies were increased, both pETF and hETF readily lost AMP. Analysis of hETF and pETF in methanol revealed intact alpha- and beta-subunits, and beta-subunit with AMP. The pETF also contained beta subunit with FAD and beta-subunit with both cofactors. In contrast to crystal structure predictions, AMP dissociates more readily than FAD, and the pETF beta subunit has an intimate association with FAD. This work demonstrates the complementarity of muESI-MS with NMR X-ray and optical spectroscopy in the analysis of noncovalent complexes. PMID- 11264008 TI - HARP induces angiogenesis in vivo and in vitro: implication of N or C terminal peptides. AB - HARP (heparin affin regulatory peptide) is a growth factor displaying high affinity for heparin. In the present work, we studied the ability of human recombinant HARP as well as its two terminal peptides (HARP residues 1-21 and residues 121-139) to promote angiogenesis. HARP stimulates endothelial cell tube formation on matrigel, collagen and fibrin gels, stimulates endothelial cell migration and induces angiogenesis in the in vivo chicken embryo chorioallantoic membrane assay. The two HARP peptides seem to be involved in most of the angiogenic effects of HARP. They both stimulate in vivo angiogenesis and in vitro endothelial cell migration and tube formation on matrigel. We conclude that HARP has an angiogenic activity when applied exogenously in several in vitro and in vivo models of angiogenesis and its NH(2) and COOH termini seem to play an important role. PMID- 11264009 TI - Molecular cloning of a novel crustacean member of the aldoketoreductase superfamily, differentially expressed in the antennal glands. AB - Biochemical studies on ecdysteroid metabolism in arthropods suggest that aldoketoreductase enzymes (AKRs) may be involved in this pathway, but very few molecular data are available on these oxidoreductases in invertebrates. Looking for such enzymes in the crayfish Orconectes limosus, we have used a PCR strategy with primers deduced from a recent insect 3beta-reductase sequence, and from mammalian 5beta-reductase sequences. A full-length cDNA, corresponding to a putative AKR, was isolated from crayfish antennal gland. This cDNA contains an open-reading frame of 1008 bp, encoding a predicted protein of 336 amino acids. Northern blots indicated a restricted expression of the transcript in the antennal glands, quite constant during the molting cycle, and in situ hybridization demonstrated a strong expression of the transcript in the labyrinth. This is to date the first member of the AKRs superfamily characterized in a crustacean species, and the putative function of the corresponding enzyme is discussed. PMID- 11264010 TI - Expression profiling of acetaminophen liver toxicity in mice using microarray technology. AB - Drug-induced hepatotoxicity causes significant morbidity and mortality and is a major concern in drug development. This is due, in large part, to insufficient knowledge of the mechanism(s) of drug-induced liver injury. In order to address this problem, we have evaluated the modulation of gene expression within the livers of mice treated with a hepatotoxic dose of acetaminophen (APAP) using high density oligonucleotide microarrays capable of determining the expression profile of >11,000 genes and expressed sequence tags (ESTs). Significant alterations in gene expression, both positive and negative, were noted within the livers of APAP treated mice. APAP-induced toxicity affected numerous aspects of liver physiology causing, for instance, >twofold increased expression of genes that encode for growth arrest and cell cycle regulatory proteins, stress-induced proteins, the transcription factor LRG-21, suppressor of cytokine signaling (SOCS)-2-protein, and plasminogen activator inhibitor-1 (PAI-1). A number of these and other genes and ESTs were detectable within the liver only after APAP treatment suggesting their potential importance in propagating or preventing further toxicity. These data provide new directions for mechanistic studies that may lead to a better understanding of the molecular basis of drug-induced liver injury and, ultimately, to a more rational design of safer drugs. PMID- 11264011 TI - Cytochrome C is a potent catalyst of dichlorofluorescin oxidation: implications for the role of reactive oxygen species in apoptosis. AB - The generation of reactive oxygen species has been suggested to occur at increased rates during apoptosis, but the validity and significance of this remain contentious. In several key studies levels of reactive oxygen species have been monitored using the intracellular probe dichlorofluorescin (DCFH(2)), which undergoes oxidation to the fluorescent dichlorofluorescein (DCF). We report here that cytochrome c, which is released from mitochondria during cell death, is a potent catalyst of DCF formation. In a model system using xanthine oxidase to generate superoxide radicals, the rate of DCF formation was insensitive to changes in the rate of superoxide production over a 17-fold range, but extremely sensitive to nanomolar concentrations of cytochrome c. Thus we conclude that the DCF fluorescence observed in dying cells is a reflection of increased cytosolic cytochrome c. Moreover, we suggest that the suppression of DCF formation by the anti-apoptotic oncoprotein Bcl-2, which has been suggested to have antioxidant properties, can be explained on the basis of its prevention of mitochondrial cytochrome c release. PMID- 11264012 TI - UCP3 expressed in yeast is primarily localized in extramitochondrial particles. AB - Previously it was concluded (1) that, differently from UCP1, on expression in Saccharomyces cerevisiae, UCP3, and UCP3 short (UCP3s) are in a deranged state, allowing for unregulated uncoupling. Here we show that the bulk of UCP3 and UCP3s is in extramitochondrial aggregates whether expressed with high or medium expression vectors. The evidence is based on the insolubility of most UCP3 and UCP3s in nonionic detergents such as Triton X100, in contrast to UCP1. Using very high expression vector, macroscopic evidence for extramitochondrial UCP3 containing particles is a viscous white sediment surrounding the mitochondrial fraction which contains UCP3 as inclusion body type aggregate. Together with the previous data it is concluded that uncoupling due to small amounts of incorporated, deranged, and nucleotide insensitive UCP3 prevents incorporation of the bulk of UCP3 into mitochondria. This finding also provides a simple and stringent assay for the state of heterologously expressed in mitochondrial membrane proteins. PMID- 11264014 TI - Carbon monoxide complex of cytochrome b(5) at acidic pH. AB - CO complex of cyt b(5) generated at acidic pH is investigated by absorption, resonance Raman (RR), and far UV CD measurements. The Soret maximum wavelength blue-shifted to 420 nm with other absorption bands observed around 540 and 570 nm for reduced cyt b(5) upon interaction with CO at acidic pH (pH 3.1-3.5). Under this condition, the iron-carbon stretching RR band was observed at 529 cm(-1) (520 cm(-1) for C(18)O), which indicated formation of a heme&bond;CO adduct with a histidine as an axial ligand. Heme dissociated from the reduced cyt b(5) protein at pH approximately 3.5, whereas its rate decreased under CO atmosphere compared with N(2) atmosphere, due to formation of a heme&bond;CO adduct with a histidine as an axial ligand. PMID- 11264013 TI - Polycystin-2 is a novel cation channel implicated in defective intracellular Ca(2+) homeostasis in polycystic kidney disease. AB - Mutations in polycystins-1 and -2 (PC1 and PC2) cause autosomal dominant polycystic kidney disease (ADPKD), which is characterized by progressive development of epithelial renal cysts, ultimately leading to renal failure. The functions of these polycystins remain elusive. Here we show that PC2 is a Ca(2+) permeable cation channel with properties distinct from any known intracellular channels. Its kinetic behavior is characterized by frequent transitions between closed and open states over a wide voltage range. The activity of the PC2 channel is transiently increased by elevating cytosolic Ca(2+). Given the predominant endoplasmic reticulum (ER) location of PC2 and its unresponsiveness to the known modulators of mediating Ca(2+) release from the ER, inositol-trisphosphate (IP(3)) and ryanodine, these results suggest that PC2 represents a novel type of channel with properties distinct from those of the other Ca(2+)-release channels. Our data also show that the PC2 channel can be translocated to the plasma membranes by defined chemical chaperones and proteasome modulators, suggesting that in vivo, it may also function in the plasma membrane under specific conditions. The sensitivity of the PC2 channel to changes of intracellular Ca(2+) concentration is deficient in a mutant found in ADPKD patients. The dysfunction of such mutants may result in defective coupling of PC2 to intracellular Ca(2+) homeostasis associated with the pathogenesis of ADPKD. PMID- 11264015 TI - Asp-Ala-His-Lys (DAHK) inhibits copper-induced oxidative DNA double strand breaks and telomere shortening. AB - Both DNA and the telomeric sequence are susceptible to copper-mediated reactive oxygen species (ROS) damage, particularly damage attributed to hydroxyl radicals. In this study, ROS-induced DNA double strand breaks and telomere shortening were produced by exposure to copper and ascorbic acid. Asp-Ala-His-Lys (DAHK), a specific copper chelating tetrapeptide d-analog of the N-terminus of human albumin, attenuated DNA strand breaks in a dose dependent manner. d-DAHK, at a ratio of 4:1 (d-DAHKCu), provided complete protection of isolated DNA from double strand breaks and, at a ratio of 2:1 (d-DAHKCu), completely protected DNA in Raji cells exposed to copper/ascorbate. Southern blots of DNA treated with copper/ascorbate showed severe depletion and shortening of telomeres and Raji cell treated samples showed some conservation of telomere sequences. d-DAHK provided complete telomere length protection at a ratio of 2:1 (d-DAHKCu). The human albumin N-terminus analog, d-DAHK, protects DNA and telomeres against copper-mediated ROS damage and may be a useful therapeutic adjunct in ROS disease processes. PMID- 11264016 TI - Ca(2+)-induced changes of surfactin conformation: a FTIR and circular dichroism study. AB - Previous NMR studies on surfactin proposed two gamma or beta-turn-containing conformers while recent CD studies described beta-sheets and alpha-helices in surfactin. Since these data were not obtained in the same conditions, the conformation of surfactin was reinvestigated by FTIR spectroscopy, a diagnostic method for beta-sheets. In trifluoroethanol, the FTIR spectra of surfactin and its diester are compatible with gamma and/or beta-turn(s) and the differences in their CD spectra show the importance of the Glu(1) and Asp(5) COOH groups in stabilizing the lipopeptide conformation. The calcium-induced spectral changes of both lipopeptides suggest a first binding of the divalent ions to the surfactin COOH groups (until calcium-lipopeptide mole ratio reached 1) followed by bulk conformational changes (at higher mole ratios). In Tris buffer at pH 8.5, the FTIR amide I band shape, without the typical 1610-1628 and 1675-1695 cm(-1) bands, ascertains the absence of beta-sheets. PMID- 11264017 TI - NMR identification of heavy metal-binding sites in a synthetic zinc finger peptide: toxicological implications for the interactions of xenobiotic metals with zinc finger proteins. AB - Lead (Pb), mercury (Hg), and cadmium (Cd) are toxic and interfere with protein metal-binding sites. The Cys(2)/His(2) zinc finger is a structural motif required for sequence-specific DNA binding and is present in zinc finger transcription factors (ZFP): Sp1, Egr-1, and TFIIIA. Neurotoxic studies have shown that heavy metals directly inhibit the DNA binding of ZFP and result in adverse cellular effects. Recently, we demonstrated the ability of heavy metals to alter the DNA binding of a synthetic Cys(2)/His(2) finger peptide (Razmiafshari and Zawia, Toxicol. Appl. Pharmacol. 166, 1-12, 2000). To determine the precise site of interactions between heavy metals and this protein domain, Pb, Hg, Cd, and Ca were reconstituted with the synthetic apopeptide and studied by one- and two dimensional NMR spectroscopy. In the presence of Zn, Cd, Hg, and Pb, but not Ca, distinct peptide NMR signal changes in the aliphatic region were observed and attributed to metal-cystiene interactions. However, chemical shifts indicative of metal-histidine binding were elicited by all the metals in the peptide's aromatic region. Chemical shift assignments and sequential connectivity were established in the presence and absence of Zn, Pb, and Ca through TOCSY and NOESY spectra. Cysteine and histidine residues showed a distinct change in their amide and beta resonances in the presence of Zn and Pb, suggesting the metal-ligand binding sites were near these residues. However, Ca led to no significant spectral changes in these regions, suggesting that it is not actively involved in the binding site. These studies reveal this structure as a mediator of metal-induced alterations in protein function. PMID- 11264018 TI - Hexachlorobenzene-induced eosinophilic and granulomatous lung inflammation is associated with in vivo airways hyperresponsiveness in the Brown Norway rat. AB - We investigated whether the eosinophilic and granulomatous lung pathology that develops in Brown Norway (BN/SsNOlaHsd) rats upon feeding hexachlorobenzene (HCB) is associated with nonspecific in vivo airways hyperresponsiveness (AHR) to methacholine. To this end, female BN/SsNOlaHsd rats were exposed to diets with no supplementation or diets supplemented with 450 mg HCB per kg feed. On days 7 or 21 of exposure in vivo airways hyperresponsiveness to increasing concentrations of methacholine was assessed both by whole body plethysmography and by visual scoring. In addition, lungs were lavaged to count and differentiate lavage cells, and skin and lungs were processed for histology. Lungs of the control rats showed some scattered microgranulomas and by 3 weeks of control diet some rats showed rather extensive granuloma formation and perivascular and peribronchiolar infiltration of eosinophils, as well as increased responsiveness to methacholine. Oral exposure to HCB for 7 days caused a moderate perivasculitis, but no increase of total serum IgE levels and no AHR to methacholine was found. Prolonged HCB exposure for 21 days resulted in severe and extensive eosinophilic and granulomatous lung inflammation, a threefold increase of total serum IgE levels, and marked cholinergic AHR in all rats. Correlation analysis revealed a significant relation between the AHR and lung inflammation, as judged by granuloma formation and increased numbers of eosinophilic granulocytes in the lung interstitium, particularly around the bronchi and bronchioli. No correlation was observed between serum IgE levels and AHR. Data suggest that HCB induces AHR by stimulating eosinophilic lung inflammation and that the preexistent microgranulomas may predispose to development of the HCB-induced lung pathology. PMID- 11264019 TI - Effects of Stachybotrys chartarum on surfactant convertase activity in juvenile mice. AB - We have shown recently that alveolar type II cells are sensitive to exposure to Stachybotrys chartarum spores, both in vitro and in an in vivo juvenile mouse model. In mice, this sensitivity is manifest in part as a significant increase in the newly secreted, biologically active, heavy aggregate form of alveolar surfactant (H) and the accumulation of the lighter, "metabolically used", biologically inactive alveolar surfactant forms (L(vivo)) in the interalveolar space. Conversion of the heavy, surface-active alveolar surfactant to the light metabolically used, nonsurface active forms is believed to involve the activity of an enzyme, namely convertase, which is thought to be derived from lamellar bodies (LB) in alveolar type II cells. The purpose of this study was to evaluate the effects of S. chartarum spores on mouse H and LB convertase activity by measuring their rates of conversion to L(vivo) using the in vitro surface area cycling technique. It was determined whether there were concurrent changes in the protein and phospholipid concentrations of the raw bronchoalveolar lavage fluid (RL) and LB fractions that could be correlated with changes in convertase activity. Conversions of H to L(vivo) in untreated control mice and saline-, isosatratoxin F-, and Cladosporium cladosporioides-exposed mice were not significantly different (p > 0.05). However, conversion from H to L(vivo) in the mice exposed to S. chartarum spores was significantly higher than all other treatment groups (p < 0.001). LB to L(vivo) conversions in untreated and saline exposed mice were not significantly different, although they were significantly higher than the H to L(vivo) conversions in these two animal treatment groups (p < 0.005), which supports the position that LB is a source of convertase activity in animals. LB to L(vivo) conversion from C. cladosporioides-, isosatrotoxin F-, and S. chartarum-exposed mice were all significantly depressed (p < 0.003) compared to the LB to L(vivo) conversion values obtained from untreated and saline-exposed mice. Protein concentrations in RL, H, L(vivo), and LB from mice exposed to S. chartarum spores were significantly elevated compared to those from the other treatment groups (p < 0.001). Protein concentration in H isolated from C. cladosporioides-exposed mice was also significantly elevated above untreated and saline control animal levels. Phospholipid concentrations in H isolated from S. chartarum-exposed mice were significantly elevated compared to those from other treatment groups, while LB phospholipid concentrations were significantly increased compared to saline and untreated control animal groups. These results show that S. chartarum spores significantly alter convertase activity in both the H and LB surfactant fractions in juvenile mice and that these changes can be related to changes in protein and phospholipid concentrations in alveolar lavage fractions. As surfactant promotes lung stability by reducing the surface tension of the air-alveolar interface, these results further support our position that inhalation exposure to S. chartarum spores in exposed individuals may lead to altered surfactant metabolism, and possibly to lung dysfunction through diminished alveolar surfactant surface tension attributes, and lung stability. PMID- 11264020 TI - Fumonisin B1 promotes aflatoxin B1 and N-methyl-N'-nitro-nitrosoguanidine initiated liver tumors in rainbow trout. AB - Laboratory studies have described the carcinogenicity of fumonisin B1 (FB1) in rodents and epidemiological evidence suggests an association between FB1 (a mycotoxin produced by Fusarium moniliforme) and cancer in humans. This study was designed to reveal in rainbow trout, a species with very low spontaneous tumor incidence, if FB1 was (i) a complete carcinogen, in the absence of an initiator; (ii) a promoter of liver tumors in fish initiated as fry with aflatoxin B1 (AFB1); and (iii) a promoter of liver, kidney, stomach, or swim bladder tumors in fish initiated as fry with N-methyl-N'-nitro-nitrosoguanidine (MNNG). FB1 was not a complete carcinogen in trout. No tumors were observed in any tissue of fish fed diets containing 0, 3.2, 23, or 104 ppm FB1 for a total of 34 weeks (4 weeks FB1 exposure, 2 weeks outgrowth on control diet, followed by 30 weeks FB1 diet) in the absence of a known initiator. FB1 promoted AFB1 initiated liver tumors in fish fed > or = 23 ppm FB1 for 42 weeks. A 1-week pretreatment of FB1 did not alter the amount of liver [3H]AFB1 DNA adducts, which suggests that short-term exposure to FB1 will not alter phase I or phase II metabolism of AFB1. In MNNG initiated fish, liver tumors were promoted in the 104 ppm FB1 treatment (42 weeks), but FB1 did not promote tumors in any other tissue. Tumor incidence decreased in kidney and stomach in the 104 ppm FB1 treatment of MNNG-initiated trout. The FB1 promotional activity in AFB1-initiated fish was correlated with disruption of sphingolipid metabolism, suggesting that alterations in associated sphingolipid signaling pathways are potentially responsible for the promotional activity of FB1 in AFB1-initiated fish. PMID- 11264021 TI - Attenuation of paraquat-induced dopaminergic toxicity on the substantia nigra by (-)-deprenyl in vivo. AB - (-)-Deprenyl (DEP) had been shown to slow of progression of Parkinson's disease (PD). The present study sought to determine whether DEP would attenuate the nigrostriatal system damage induced by intranigral administration of the herbicide paraquat (PQ) as a model of parkinsonism in vivo. Neurochemical and behavioral observations of Wistar rats were the focus of our study. In the neurochemical observation, the PQ injected in the rats caused dose-dependent depletion of dopamine (DA) in the ipsilateral striata. The coadministration of DEP with PQ partially increased the striatal DA level. The prediction of the striatal DA levels was calculated by regression coefficients obtained from multiple linear regression (r(2) = 0.82): DA level (% of control) = 103.34 - 9.58 PQ (nmol) + 0.79 DEP (nmol). It was demonstrated that the high dose of 20 nmol DEP could significant attenuate the PQ (5 nmol)-elicited dopaminergic toxicity (p < 0.05). In the behavioral observation, the intranigral injection of PQ into the rats caused a rotation behavior contralateral to the lesioned side in response to apomorphine administration (0.5 mg/kg, sc). This apomorphine-induced rotational behavior could also be attenuated significantly by coadministration of DEP (20 nmol) and PQ (5 nmol) compared with PQ-treated (5 nmol) animals (p < 0.05). The above observations indicate that DEP could provide a protective effect on the moderate injury elicited by PQ toxicity of the nigrostriatal dopaminergic system. DEP might be a useful therapeutic agent in treating patients with early-stage PD. PMID- 11264022 TI - Distinct roles of tumor necrosis factor-alpha and nitric oxide in acute liver injury induced by carbon tetrachloride in mice. AB - Macrophages are known to release a number of different inflammatory mediators with cytotoxic potential. In the present studies we analyzed the role of two macrophage-derived mediators, tumor necrosis factor-alpha (TNF-alpha) and nitric oxide, in liver injury induced by carbon tetrachloride (CCl4). Treatment of mice with CCl4 resulted in a dose- and time-dependent induction of centrilobular hepatic necrosis. This was observed within 12 h with 0.3 ml/kg CCl4 and was correlated with increases in serum transaminase levels. CCl4 administration also caused increases in hepatic TNF-alpha mRNA expression and serum TNF-alpha levels, as well as inducible nitric oxide synthase (NOS II) protein expression in the liver. To study the role of TNF-alpha and nitric oxide in hepatotoxicity, we used knockout mice lacking the gene for the 55-kDa TNF-alpha receptor (TNFR1/p55), the TNF-alpha cytokine, or NOS II. We found that CCl4 was significantly less effective in inducing hepatotoxicity in mice lacking TNFR1/p55 or the TNF-alpha cytokine. In contrast, CCl4-induced liver injury was increased in knockout mice lacking the gene for NOS II. This was associated with an increase in hepatic TNF alpha mRNA expression and serum TNF-alpha levels. These data suggest that the hepatoprotective effects of nitric oxide in this model may be due in part to inhibition of TNF-alpha. PMID- 11264023 TI - Selenium modifies the metabolism and toxicity of arsenic in primary rat hepatocytes. AB - Arsenic and selenium are metalloids with similar chemical properties and metabolic fates. Inorganic arsenic (iAs) has been shown to modify metabolism and toxicity of inorganic and organic selenium compounds. However, little is known about effects of selenium on metabolism and toxicity of iAs. The present work examines the effects of selenite (Se(IV)) on the cellular retention, methylation, and cytotoxicity of trivalent iAs, arsenite (iAs(III)), in primary cultures of rat hepatocytes. The concurrent exposure to Se(IV) (0.1 to 6 microM) inhibited methylation and/or significantly increased cellular retention of iAs(III) in cultured cells. The ratio of the methylated metabolites produced from iAs(III), dimethylarsenic (DMAs) to methylarsenic (MAs), decreased considerably in cells treated with Se(IV), suggesting that synthesis of DMAs from MAs may be more susceptible to inhibition by Se(IV) than is the production of MAs from iAs(III). The 24-h preexposure to 2 microM Se(IV) had a similar but less pronounced inhibitory effect on the methylation of iAs(III) in cultured cells. The exposure to 2 microM Se(IV) alone for up to 24 h had no effect on the viability of cultured hepatocytes. However, concurrent exposure to 2 microM Se(IV) increased the cytotoxicity of iAs(III) and its mono- and dimethylated metabolites that contain trivalent arsenic, MAs(III) and DMAs(III). These data suggest that pre- or coexposure to inorganic selenium may enhance the toxic effects of iAs, increasing its retention in tissues and suppressing its methylation, which may be a pathway for detoxification of iAs. PMID- 11264024 TI - Kinetics of propylene oxide metabolism in microsomes and cytosol of different organs from mouse, rat, and humans. AB - Kinetics of the metabolic inactivation of 1,2-epoxypropane (propylene oxide; PO) catalyzed by glutathione S-transferase (GST) and by epoxide hydrolase (EH) were investigated at 37 degrees C in cytosol and microsomes of liver and lung of B6C3F1 mice, F344 rats, and humans and of respiratory and olfactory nasal mucosa of F344 rats. In all of these tissues, GST and EH activities were detected. GST activity for PO was found in cytosolic fractions exclusively. EH activity for PO could be determined only in microsomes, with the exception of human livers where some cytosolic activity also occurred, representing 1-3% of the corresponding GST activity. For GST, the ratio of the maximum metabolic rate (V(max)) to the apparent Michaelis constant (K(m)) could be quantified for all tissues. In liver and lung, these ratios ranged from 12 (human liver) to 106 microl/min/mg protein (mouse lung). Corresponding values for EH ranged from 4.4 (mouse liver) to 46 (human lung). The lowest V(max) value for EH was found in mouse lung (7.1 nmol/min/mg protein); the highest was found in human liver (80 nmol/min/mg protein). K(m) values for EH-mediated PO hydrolysis in liver and lung ranged from 0.83 (human lung) to 3.7 mmol/L (mouse liver). With respect to liver and lung, the highest V(max)/K(m) ratios were obtained for GST in mouse and for EH in human tissues. GST activities were higher in lung than in liver of mouse and human and were alike in both rat tissues. Species-specific EH activities in lung were similar to those in liver. In rat nasal mucosa, GST and EH activities were much higher than in rat liver. PMID- 11264025 TI - Association of tumor necrosis factor-alpha and interleukin-1 gene polymorphisms with silicosis. AB - Silicosis, an interstitial lung disease prevalent among miners, sand blasters, and quarry workers, is manifested as a chronic inflammatory response leading to severe pulmonary fibrotic changes. Proinflammatory cytokines, such as TNFalpha and IL-1, produced in the lung by type II epithelial cells and alveolar macrophages, have been strongly implicated in the formation of these lesions. Recently, a number of single nucleotide polymorphisms (SNPs), which quantitatively affect mRNA synthesis, have been identified in the TNFalpha promoter and IL-1 gene cluster and their frequency is associated with certain chronic inflammatory diseases. To assess the role of these SNPs in silicosis, we examined their frequency in 325 ex-miners with moderate and severe silicosis and 164 miners with no lung disease. The odds ratio of disease for carriers of the minor variant, TNFalpha (-238), was markedly higher for severe silicosis (4.0) and significantly lower for moderate silicosis (0.52). Regardless of disease severity, the odds ratios of disease for carriers of the IL-1RA (+2018) or TNFalpha (-308) variants were elevated. There were no significant consistent differences in the distribution of the IL-1alpha (+4845) or IL-1beta (+3953) variants with respect to disease status. In addition, several significant gene gene and gene-gene-environment interactions were observed. Different associations between moderate cases and controls versus severe cases and controls were also observed in a number of these multigene comparisons. These studies suggest that gene-environment interactions involving cytokine polymorphisms play a significant role in silicosis by modifying the extent of and susceptibility to disease. PMID- 11264026 TI - Role of Mycoplasma genitalium and Ureaplasma urealyticum in acute and chronic nongonococcal urethritis. AB - One hundred fourteen heterosexual men with acute nongonococcal urethritis (NGU) and 64 patients without NGU were studied. We determined that Chlamydia trachomatis and Mycoplasma genitalium were strongly associated with acute NGU after controlling, by means of multivariate analysis, for age, race, sexual lifestyle, and coinfection (odds ratio [OR], 13.0, 95% confidence interval [CI], 2.6-64.5; and OR, 17.9, 95% CI, 2.0-160, respectively). Eighty-six men with acute NGU reattended at least once 10-92 days after treatment; 59 (69%) of these 86 men had urethritis. Seven men had M. genitalium detected during 10-92 days of follow up, and all had urethritis. Ureaplasmas were not associated with acute NGU in multivariate analysis, but their detection was associated with the presence of urethritis during follow-up (P=.014). Ureaplasmas or M. genitalium were associated with both chronic NGU, which was defined as urethritis that occurred 30-92 days after the commencement of treatment (P=.028), and chronic NGU with symptoms or signs (P=.005). PMID- 11264027 TI - Meningococcal disease among children who live in a large metropolitan area, 1981 1996. AB - Neisseria meningitidis is an important cause of serious bacterial infections in children. We undertook a study to identify meningococcal infections of the blood, cerebrospinal fluid, or both of children in a defined geographic area to describe the burden of disease and the spectrum of illness. We reviewed the medical records of all children aged <18 years who had meningococcal infections at the 4 pediatric referral hospitals in Boston, Massachusetts, from 1981 through 1996. We identified 231 patients with meningococcal disease; of these 231 patients, 194 (84%) had overt disease and 37 (16%) had unsuspected disease. Clinical manifestations included meningitis in 150 patients, hypotension in 26, and purpura in 17. Sixteen patients (7%) died. Although meningococcal disease is devastating to a small number of children, we found that the burden of pediatric disease that it caused at the 4 pediatric referral centers in this geographic region was limited; that patients with overt meningococcal disease are most likely to have meningitis; and that individual practitioners are unlikely to encounter a patient with unsuspected meningococcal disease. PMID- 11264028 TI - Outbreak of cyclosporiasis associated with basil in Missouri in 1999. AB - During the summer of 1999, an outbreak of cyclosporiasis occurred among attendees of 2 events held on 24 July in different counties in Missouri. We conducted retrospective cohort studies of the 2 clusters of cases, which comprised 62 case patients. The chicken pasta salad served at one event (relative risk [RR], 4.25; 95% confidence interval [CI], 1.80-10.01) and the tomato basil salad served at the other event (RR, 2.95; 95% CI, 1.72-5.07) were most strongly associated with illness. The most likely vehicle of infection was fresh basil, which was included in both salads and could have been grown either in Mexico or the United States. Leftover chicken pasta salad was found to be positive for Cyclospora DNA by means of polymerase chain reaction analysis, and 1 sporulated Cyclospora oocyst was found by use of microscopy. This is the second documented outbreak of cyclosporiasis in the United States linked to fresh basil and the first US outbreak for which Cyclospora has been detected in an epidemiologically implicated food item. PMID- 11264030 TI - Current understanding and management of chronic hepatosplenic suppurative brucellosis. AB - To outline the characteristics and define appropriate management of chronic hepatosplenic suppurative brucellosis (CHSB), 905 patients with brucellosis were analyzed. Sixteen episodes of CHSB (14 in the liver and 2 in the spleen) were found in 15 patients. Six patients had had previous remote brucellosis. Twelve patients presented with systemic symptoms, and 12 with local symptoms. Cultures of blood samples yielded negative results in all cases except 1, and the results of cultures of pus specimens were positive for Brucella melitensis in only 2 cases. All patients showed calcium deposits surrounded by a hypodense area on computed tomography. Patients often had low titers of agglutinating antibody. In patients who were receiving conservative management, early response was successful in 50% and late response was successful in 33.3%. In the patients who underwent surgery and concomitant antibiotic therapy, early and late response was successful in 100%. Thus, CHSB mainly represents a local reactivation of previous brucellosis. Its diagnosis may be difficult to establish and surgery may be required to cure many patients. PMID- 11264029 TI - Ophthalmologic, visceral, and cardiac involvement in neonates with candidemia. AB - A retrospective review of 86 neonates with candidemia hospitalized from January 1989 through June 1999 was conducted to determine the frequency of ophthalmologic, visceral, or cardiac involvement. Retinal abnormalities were observed in 4 (6%) of the 67 infants in whom indirect ophthalmoscopy examination was performed. Abdominal ultrasound abnormalities were detected in 5 (7.7%) of 65 infants. Echocardiogram revealed thrombi or vegetations in 11 (15.2%) of 72 infants. Age at onset, presence of central venous catheters, and species of Candida were not predictors for involvement at these sites. Infants with candidemia that lasted > or =5 days were more likely to demonstrate ophthalmologic, renal, or cardiac abnormalities than those with a shorter duration. Infants with involvement of these organs received larger cumulative doses of amphotericin B than those without detectable abnormalities. Because complication of disseminated candidiasis by eye, renal, or cardiac involvement has therapeutic implications, and because risk factors for candidemia inadequately predict these complications, evaluations are indicated for all neonates with candidemia. PMID- 11264031 TI - Candida dubliniensis at a cancer center. AB - Candida dubliniensis, a germ tube-positive yeast first described and identified as a cause of oral candidiasis in patients with acquired immunodeficiency syndrome in Europe in 1995, has an expanding clinical and geographic distribution that appears to be similar to that of the other germ tube-positive yeast, Candida albicans. This study determined the frequency, clinical spectrum, drug susceptibility profile, and suitable methods for identification of this emerging pathogen at a cancer center in 1998 and 1999. Twenty-two isolates were recovered from 16 patients with solid-organ or hematologic malignancies or acquired immunodeficiency syndrome. Two patients with cancer had invasive infection, and 14 were colonized with fungus or had superficial fungal infection. All isolates produced germ tubes and chlamydospores at 37 degrees C, did not grow at 45 degrees C, and gave negative reactions with d-xylose and alpha-methyl-d-glucoside in the API 20 C AUX and ID 32 C yeast identification systems. Phenotypic identification was confirmed by molecular beacon probe technology. All isolates were susceptible to the antifungal drugs amphotericin B, 5-fluorocytosine, fluconazole, itraconazole, and ketoconazole. PMID- 11264032 TI - Streptococcus pneumoniae nasopharyngeal carriage prevalence, serotype distribution, and resistance patterns among children on Lombok Island, Indonesia. AB - Few data exist on childhood pneumococcal carriage prevalence, serotype distribution, and resistance patterns for Indonesia, the world's fourth most populous country. During August 1997, nasopharyngeal samples were collected from a population-based, island-wide sample of 484 healthy children (age, 0-25 months) from Lombok Island, Indonesia. Two hundred twenty-one pneumococcal isolates were identified, for a carriage prevalence of 48%; 66% of isolates were of serogroup or serotype 6, 23, 15, 33, or 12. All isolates were susceptible to penicillin and cefotaxime. Twelve percent of the isolates were nonsusceptible to sulfamethoxazole or chloramphenicol and 4% were nonsusceptible to both of these drugs. Nonsusceptible organisms were most frequently serogroup or serotype 6, 12, and 33. Lombok has a moderate pneumococcal carriage prevalence and a relatively low proportion of resistant isolates. At least 3 of the 5 most common serogroups and serotypes and 2 of the 3 most common nonsusceptible serogroups and serotypes are not included in the current 7-valent pneumococcal conjugate vaccine. PMID- 11264033 TI - Measuring and interpreting associations between antibiotic use and penicillin resistance in Streptococcus pneumoniae. AB - Studies of the relationship between antibiotic use and resistance in pneumococci have produced conflicting results, reflecting differences in study design, setting, and measures of association used. Mathematical models of pneumococcal transmission dynamics provide a framework for interpreting and reconciling these studies. The model predicts, and the review of published studies confirms, that treatment often has little effect in increasing an individual's absolute risk of carrying/being infected by penicillin-resistant Streptococcus pneumoniae (PRSP). However, treatment substantially increases a patient's risk of carriage of/infection by PRSP relative to that of penicillin-susceptible S. pneumoniae (PSSP). The appropriate measure of association depends on the question of interest. Antibiotic use can substantially increase the prevalence of risk in the community as a whole, even when there is a small or nonexistent effect of treatment on the absolute risk that a treated individual will carry a resistant organism. Recommendations for the design and analysis of future studies of antibiotic treatment and pneumococcal resistance are proposed. PMID- 11264034 TI - Methodological principles of case-control studies that analyzed risk factors for antibiotic resistance: a systematic review. AB - Case-control studies that analyze the risk factors for antibiotic-resistant organisms have varied epidemiological methodologies, which may lead to biased estimates of antibiotic risk factors. A systematic review of case-control studies that analyzed risk factors for antibiotic-resistant organisms addressed 3 methodological principles: method of control group selection, adjustment for time at risk, and adjustment for comorbid illness. A total of 406 abstracts were reviewed. Thirty-seven studies met the inclusion and exclusion criteria and were reviewed and evaluated for the 3 methodological principles. Thirteen (35%) of 37 studies chose the preferred control group. Eleven adjusted for time at risk. Twenty-seven adjusted for comorbid illness. Future studies need to consider more closely the optimization of control group selection, adjusting for confounding caused by time at risk, and adjusting for confounding caused by comorbid illness. PMID- 11264035 TI - Sexually transmitted diseases in travelers. AB - Prevention of sexually transmitted diseases (STDs) is a low priority among travel clinic services, despite increasing evidence that travelers have an increased risk of acquiring such infections. A proportion of 5%-50% of short-term travelers engage in casual sex while abroad, and this rate is even higher among long-term travelers. Few publications are available on STD preventive interventions among travelers. Education and counseling are recognized as key components of risk reduction. New efforts should be put forth with regard to identifying effective tools to promote safer sexual behaviors and to reduce the spread of infection by promoting condom use. Travelers at increased risk should be identified for targeted interventions; research to validate proposed markers of increased risk is prospectively needed. Hepatitis B infection is the only STD that is preventable by vaccination. The feasibility and cost-effectiveness of STD screening in travelers after exposure is a virtually unexplored field, though it may represent an important component of STD control strategies in developed countries. PMID- 11264036 TI - CHAT oral polio vaccine was not the source of human immunodeficiency virus type 1 group M for humans. AB - A book published in 1999 hypothesized that the scientists who worked with the CHAT type 1 attenuated poliomyelitis strain, tested in the former Belgian Congo in the late 1950s, had covertly prepared the vaccine in chimpanzee kidney cells contaminated with a simian immunodeficiency virus, which evolved into human immunodeficiency virus type 1 group M. This article summarizes the results of the investigation conducted by the author to determine the legitimacy of the accusation. Testimony by eyewitnesses, historical documents of the time, epidemiological analysis, and analysis of ancillary phylogenetic, virological, and polymerase chain reaction data all indicate that this hypothesis is false. PMID- 11264037 TI - New beta-lactamases in gram-negative bacteria: diversity and impact on the selection of antimicrobial therapy. AB - Of the 340 discrete beta-lactamases that have been identified, the most important groups of enzymes that are continuing to proliferate include the plasmid-encoded cephalosporinases, the metallo-beta-lactamases, and the extended-spectrum beta lactamases. Resistance to specific beta-lactam-containing antimicrobial agents frequently can be traced to a single beta-lactamase, but this task is becoming more difficult for the clinical microbiology laboratory. Other factors, such as multiple beta-lactamase production, transferable multidrug-resistance genes, alterations in outer-membrane porins, and possible antibiotic efflux, all may contribute to a resistance phenotype. Appreciation of these factors may help the physician make a more informed decision when choosing therapy to try to avoid selection of even more pathogenic strains. PMID- 11264038 TI - Effect of highly active antiretroviral therapy on the serological response to additional measles vaccinations in human immunodeficiency virus-infected children. AB - Children infected with human immunodeficiency virus (HIV) often lose their vaccine-induced antibody to measles virus. Before highly active antiretroviral therapy (HAART), an additional immunization against measles infrequently resulted in protective antibodies. The antibody response to an additional measles-mumps rubella (MMR) vaccination was compared in 28 HIV-infected children who lacked protective antibody to measles virus and were undergoing HAART or non-HAART regimens. Serostatus was measured by automated enzyme-linked immunoassay. Nine (64.3%) of 14 children undergoing HAART, compared with 3 (21.4%) of 14 in the non HAART group, had antibody to measles virus after the additional vaccination with MMR (P=.027). The groups showed no significant difference in CD4 cell values. Ten of 14 HAART patients had undetectable levels of HIV. The mean HIV load for the HAART group was 27,700 copies/mL (median, <400 copies/mL); for the non-HAART group, it was 86,000 copies/mL (median, 9000 copies/mL). Thus, HAART improves the response to an additional MMR vaccination, which is consistent with immune system reconstitution. PMID- 11264039 TI - Concentrations of human immunodeficiency virus type 1 (HIV-1) RNA in cerebrospinal fluid after antiretroviral treatment initiated during primary HIV-1 infection. AB - In 6 patients with primary human immunodeficiency virus type 1 (HIV-1) infection, concentrations of HIV-1 RNA and beta(2)-microglobulin were monitored in cerebrospinal fluid (CSF) and in plasma during antiretroviral therapy. Four patients had neurological symptoms. At baseline, the CSF of 5 patients had detectable levels of HIV-1 RNA (median, 3.68 log(10) copies/mL; range, <2.60-5.67 log(10) copies/mL), and the CSF of 3 patients had elevated levels of beta(2) microglobulin. After 8 weeks of treatment, the median concentrations of HIV-1 RNA in CSF had decreased to <2.60 log(10) copies/mL (range, <1.60-3.00 log(10) copies/mL; P=.04) and in plasma to 3.07 log(10) copies/mL (range, 2.57-3.79 log(10) copies/mL; P=.03). Median concentration of beta(2)-microglobulin in CSF had decreased to 1.2 mg/L (range, 0.9-1.7 mg/L; P=.06) and, in plasma, to 1.7 mg/L (range, 1.1-2.2 mg/L; P=.03). After 48 weeks, HIV-1 RNA concentrations in 1 patient were still 1.97 log(10) copies/mL in CSF and 1.51 log(10) copies/mL in plasma, although beta(2)-microglobulin concentrations in CSF and plasma had normalized after 8 weeks. PMID- 11264040 TI - Discordant HHV8 detection in a young HIV-negative patient with Kaposi's sarcoma and sarcoidosis. AB - Human herpesvirus 8 (HHV8), which has been suggested as the causal agent of Kaposi's sarcoma (KS), has also been implicated in the pathogenesis of sarcoidosis. We describe a patient affected concomitantly by sarcoidosis and KS. HHV8 sequences were detected with PCR only on KS lesions, whereas sarcoid tissues did not harbor HHV8 DNA. Immune dysfunction related to sarcoidosis may have facilitated the oncogenic role of HHV8 and the development of KS. PMID- 11264041 TI - Myositis resulting from disseminated cryptococcosis in a patient with hepatitis C cirrhosis. AB - We report a case of myositis that resulted from disseminated cryptococcosis in a patient with hepatitis C cirrhosis. One year after cessation of treatment, the patient remains symptom free with negative results of serum cryptococcal antigen tests and negative culture results. PMID- 11264042 TI - Neurosyphilis presenting as herpes simplex encephalitis. AB - We report the case of a 55-year-old man with neurosyphilis that presented with features of herpes simplex encephalitis. Neurosyphilis is easily diagnosed and treated and should be included in the differential diagnosis of herpes simplex encephalitis. PMID- 11264043 TI - Is vancomycin resistance in enterococci predictive of inferior outcome of enterococcal bacteremia? PMID- 11264044 TI - Estimating the cost of nosocomial candidemia in the united states. PMID- 11264046 TI - Reassuring safety profile of moxifloxacin. PMID- 11264047 TI - Are enzyme-linked immunosorbent assay and immunoblot assay independent in immunodiagnosis of infectious diseases? PMID- 11264048 TI - Environmental exposure to Toxocara as a possible risk factor for asthma: a clinic based case-control study. AB - The zoonotic ascarid Toxocara has been suggested as a possible etiologic agent of asthma. We conducted a clinic-based case-control study to examine whether the zoonotic infection acquired by ingesting Toxocara eggs is associated with asthma in children. Blood samples were collected from children aged 2-15 years, 95 of whom had asthma and 229 of whom did not have asthma. Risk factors for asthma and Toxocara infection were assessed by a questionnaire given to each child's parent or legal guardian. Blood samples were tested for the presence of Toxocara antibodies, using an enzyme-linked immunosorbent assay. No significant association was found between Toxocara infection and asthma. Significant associations were found between asthma and risk factors and between Toxocara infection and risk factors. High prevalence of Toxocara infection was noted among Hispanic children of Puerto Rican descent. PMID- 11264050 TI - Corynebacterium jeikeium native valve endocarditis following femoral access for coronary angiography. AB - We present a unique case of rapidly fatal native aortic-valve endocarditis due to Corynebacterium jeikeium, with inoculation as a complication of repeated femoral vascular access for coronary angiography. PMID- 11264049 TI - Non-serogroup O:1 Vibrio cholerae bacteremia and cerebritis. AB - We describe a case of non-serogroup O:1 Vibrio cholerae bacteremia and cerebritis in a 41-year-old Thai man with alcoholism who presented with fever and cellulitis of the right ankle. He was successfully treated with parenteral cefotaxime and then was switched to treatment with oral ciprofloxacin. PMID- 11264052 TI - When parties and candidates collide: citizen perception of house candidates' positions on abortion. AB - When candidates assume issue positions opposite those of their sponsoring political party do citizens recognize these positions? Relatedly, what role do candidates' actual issue positions play in citizens' perceptions of their issue positions? Examining citizens' perceptions of 1996 and 1998 House candidates' position on abortion, this research finds that citizens' perceptions are shaped largely by partisan and, to a lesser extent, gender stereotypes. However, candidates' individuating positions on abortion influence perceptions of the candidates' position, but the effects are considerably stronger for perceptions of Republican candidates. Democratic candidates are likely to adopt anti-abortion positions in districts characterized by lower than average levels of political awareness and education, reducing the likelihood their party-contradicting position is accurately perceived. In contrast, Republican candidates adopt a pro choice position in districts characterized by high education and political awareness, increasing the likelihood their position is accurately perceived. PMID- 11264051 TI - Resolution of chronic parvovirus b19-induced anemia, by use of highly active antiretroviral therapy, in a patient with acquired immunodeficiency syndrome. AB - We present what is, to our knowledge, the second case report of a patient with acquired immunodeficiency syndrome and transfusion-dependent anemia caused by persistent infection with parvovirus B19. The patient's anemia was successfully treated, and she demonstrated serologic evidence of eradication of parvovirus after having received effective therapy for human immunodeficiency virus infection. PMID- 11264053 TI - Overreporting voting: why it happens and why it matters. AB - The key to understanding why people overreport is that those who are under the most pressure to vote are the ones most likely to misrepresent their behavior when they fail to do so. Among all nonvoters, the most likely to overreport are the more educated, partisan, and religious, and those who have been contacted and asked to vote for a candidate. The greater the concentration of African-American and Latino nonvoters in a district, the greater the probability of overreporting in those districts, both among those in the relevant minority group and among white Anglos. White nonvoters are more likely to overreport in the Deep South than elsewhere. Overreporting matters: using reported votes in place of validated votes substantially distorts standard multivariate explanations of voting, increasing the apparent importance of independent variables that are related in the same direction to both overreporting and voting and sharply decreasing the apparent importance of independent variables related in opposing directions to those two variables. PMID- 11264054 TI - Event history calendars and question list surveys: a direct comparison of interviewing methods. AB - The research reported in this article provides the first direct experimental comparison between Event History Calendar (EHC; N=309; 84.4 percent response rate) and standardized state-of-the-art question list (Q-list; N=307; 84.1 percent response rate) interviewing methodologies. Respondents and 20 interviewers were randomly assigned to EHC and Q-list interviews that were conducted via telephone in the spring of 1998. All interviews asked for retrospective reports on social and economic behaviors that occurred during the calendar years of 1996 and 1997. Using data from the same respondents collected 1 year earlier on events reported during 1996 as a standard of comparison, the quality of retrospective reports on 1996 events from the 1998 administration of EHC and Q-list interviews was assessed. In comparison to the Q-list, the EHC condition led to better-quality retrospective reports on moves, income, weeks unemployed, and weeks missing work resulting from self illness, the illness of another, or missing work for these reasons in combination with other ones. For reports of household members entering the residence, and number of jobs, the EHC led to significantly more overreporting than the Q-list. Contingent on additional research that examines a wider range of reference periods and different modes of interviewing, the EHC may become a viable and potentially superior method to the Q-list in the collection of self-reported retrospective information. PMID- 11264055 TI - Do phone calls increase voter turnout?: a field experiment. PMID- 11264056 TI - Innumeracy about Minority Populations: African Americans and Whites Compared. PMID- 11264057 TI - Validation of Time-of-Voting-Decision Recall. PMID- 11264058 TI - The polls-review: a late campaign swing or a failure of the polls? The case of the 1998 quebec election. PMID- 11264059 TI - The Polls-Trends: American Ambivalence toward China. PMID- 11264060 TI - Yin and yang: the paradox of advances in imaging. PMID- 11264061 TI - Localized cystic disease of the kidney. AB - OBJECTIVE: Localized cystic disease of the kidney is a benign nonsurgical condition. Its imaging and clinical features are characterized and differentiated from autosomal dominant polycystic kidney disease, multilocular cystic nephroma, and cystic neoplasm. MATERIALS AND METHODS: Localized cystic disease was diagnosed in 18 patients on the basis of a review of imaging studies, clinical histories, and pathologic proof in four of the 18 patients. Average age at diagnosis was 54 years (age range, 24-83 years). Fifteen of the patients (83%) were men. CT was performed on 18 patients, sonography on nine, excretory urography on six, arteriography on four, and MR imaging on two. RESULTS: Localized cystic disease was unilateral in all patients and characterized by multiple cysts of various sizes separated by normal (or atrophic) renal tissue in a conglomerate mass suggestive of cystic neoplasm. In some patients, involvement of the entire kidney, which was suggestive of unilateral autosomal dominant polycystic kidney disease, was seen. No cysts were seen in the contralateral kidney in 14 patients, and only one or two scattered small cysts were present in four patients. Clinical presentations included hematuria, flank pain, palpable abdominal mass, and localized cystic disease as an incidental finding. None of the patients had a family history of autosomal dominant polycystic kidney disease. Ten patients underwent follow-up (follow-up range, 1-12 years); nine patients underwent imaging follow-up and one patient underwent clinical follow up, which showed stability of disease. Four patients underwent nephrectomy for suspected renal neoplasm. CONCLUSION: Familiarity with localized cystic disease of the kidney and its imaging findings is important to avoid unnecessary surgery and to differentiate the disease from autosomal dominant polycystic kidney disease. PMID- 11264063 TI - Trends in case-mix-adjusted use of radiology resources at an urban level 1 trauma center. AB - OBJECTIVE: The objective of our study was to determine the utilization rates of diagnostic radiology services at an urban level 1 trauma center. MATERIALS AND METHODS: This was an observational study of imaging use patterns from 1993 to 1998. Data were segregated by patient type and imaging procedure. Annual hospital admissions were adjusted for severity of illness using the Health Care Financing Administration's case-mix index. Per-patient imaging trends for the emergency department and outpatients were assessed using a ratio of the total number of procedures to the number of patient visits. Linear regression models were used to assess the strength of associations between resource use, measured as relative value units (RVUs), and independent variables (calendar year, patient type, and examination type). RESULTS: The RVUs for all imaging increased 53% for inpatients, 69% for outpatients, and 85% in the emergency department. No significant trend for use was found for the aggregate of inpatient imaging. There was a significant increase in the inpatient MR imaging RVUs (p = 0.04). No significant trend was found for the aggregated outpatient imaging RVUs. The trends were significant for angiography (p = 0.006), MR imaging (p = 0.002), and sonography (p = 0.04). The aggregated emergency room imaging RVUs showed a significant increase over time (p < 0.03). CONCLUSION: The number of imaging procedures increased during the study period. There was no overall trend toward increasing use of imaging in inpatients once an adjustment for severity of illness was made. Increases in patient visit-adjusted emergency department use of CT, sonography, and nuclear medicine procedures resulted from changes in practice patterns. For emergency department and outpatient settings, adjusting for the number of patient visits explains a significant portion of the increase in utilization. PMID- 11264064 TI - Remote sonographic interpretation using a laser printer network: system performance and diagnostic accuracy in actual clinical practice. AB - OBJECTIVE: The purpose of our study was to evaluate the technical and clinical performance of remote sonographic interpretation using a laser printer network connecting a community-based imaging center and an academic medical center. SUBJECTS AND METHODS: During a 3-month period, 161 consecutive sonographic examinations were performed at a community-based imaging center and all 161 patients were enrolled in the study. Seventy-one (44%) of 161 examinations were interpreted on-site at the community-based imaging center, and 90 (56%) of 161 were transmitted over a T-1 line to an academic medical center where the static images were interpreted remotely. For the purposes of this study, the examination time was defined as the interval from the time the technologist started to scan the patient to the time the patient was dismissed from the radiology department. Examination times were recorded for each patient. Follow-up was available for 92 (57%) of 161 studies. Sensitivity and specificity for studies interpreted at the community-based imaging center and at the academic medical center were calculated. RESULTS: The mean examination time for pelvic sonographic examinations interpreted at the academic medical center (43 min) was significantly longer than for scans interpreted at the community-based imaging center (31 min) (p < 0.01). However, no significant difference was noted in the examination time for abdominal sonography. For all examinations interpreted on site at the community-based imaging center for which follow-up was available, the sensitivity and specificity were 95% and 100%, respectively. For all examinations interpreted remotely at the academic medical center for which follow-up was available, the sensitivity and specificity were 93% and 90%, respectively. No significant difference was seen in the sensitivity (p = 1.00) or specificity (p = 0.24) of studies interpreted on-site versus remotely. CONCLUSION: Static sonographic images can be interpreted remotely without loss of sensitivity, but with decreased specificity. However, more time must be allotted for performing pelvic sonography when these examinations are to be interpreted remotely. PMID- 11264065 TI - Radiologists' productivity in the interpretation of CT scans: a comparison of PACS with conventional film. AB - OBJECTIVE: We compared radiologists' times in the interpretation of CT using hardcopy films with the interpretation using a soft-copy picture archiving and communication system (PACS) computer workstation. MATERIALS AND METHODS: One hundred CT examinations were selected at random and reviewed by four board certified radiologists experienced in soft-copy interpretation. We performed time motion analysis to determine the total time required to display, interpret, and dictate the individual findings of CT using conventional hard-copy interpretation on a viewbox and soft-copy interpretation, using a four-monitor high-resolution (2048 x 1536 pixel) workstation. RESULTS: Time-motion analysis showed a reduction of 16.2% in the overall time required for soft-copy interpretation of CT compared with that of film. Time savings with soft-copy interpretation were observed for all four participating radiologists. The benefit of soft-copy interpretation was increased for examinations in which there were comparison studies. CONCLUSION: We found that soft-copy interpretation of CT using a PACS workstation requires less time than interpretation using conventional film hung on a viewbox. The transition to filmless imaging has the potential to improve radiologists' productivity and report-turnaround time. PMID- 11264067 TI - Iodine-131 and the pregnant patient. PMID- 11264066 TI - Using intranasal midazolam spray to prevent claustrophobia induced by MR imaging. AB - OBJECTIVE: Up to 37% of patients undergoing MR imaging examinations experience moderate to severe levels of anxiety that necessitate the termination of the procedure in 5-10% of patients. Although the clinical use of MR imaging has increased, effective procedures to handle claustrophobia are lacking. We evaluated the effectiveness of intranasally administered midazolam spray in preventing claustrophobic responses of patients undergoing MR imaging. SUBJECTS AND METHODS: Fifty-four patients scheduled for MR imaging were included in this prospective study. Anxiety and sedation of patients were evaluated before drug administration, immediately before MR imaging, and at the end of the procedure. The Spielberger State-Trait Anxiety Inventory, the visual analogue scale of anxiety, and a five-point sedation scale were used. Half the patients received intranasal spray applications of 4 mg midazolam, whereas the other patients received a placebo, in a randomized, double-blind study design (six sprayings of 0.5% midazolam solution or NaCl 0.9%, respectively). The intensity of the sensation of burning of the nasal mucosa was rated by patients using a three point scale (no, slight, or strong burning). The quality of scan images was evaluated by a radiologist using a five-point scale (0 = extremely poor, 5 = excellent). RESULTS: No cancellations occurred with patients who received midazolam, whereas four of 27 patients receiving placebo panicked and terminated the scanning procedure. The initial anxiety and sedation scores did not differ between the groups. Patients who received midazolam spray were more sedated and less anxious immediately before entering the MR scanner and reported a more intense slight transient burning of the nasal mucosa than those in the placebo group. The quality of the MR image was higher in the midazolam group. CONCLUSION: A sizeable reduction in MR imaging-related anxiety and improved MR image quality were seen with patients who received intranasal midazolam spray. With the exception of transient burning of the nasal mucosa, no adverse effects were reported. This simple and safe method is useful in sedating patients for MR imaging and other minor procedures. PMID- 11264069 TI - The research framework. PMID- 11264070 TI - Prediction rule for characterization of hepatic lesions revealed on MR imaging: estimation of malignancy. AB - OBJECTIVE: Our aims were to establish factors that are most predictive of hepatic lesion malignancy and to formulate a prediction rule. MATERIALS AND METHODS: A cross-sectional study of 227 abdominal MR imaging examinations revealed 85 lesions in 67 patients (29 men, 38 women; age range, 29-78 years; mean age, 51.4 years) who were being examined for primary malignancy (n = 42) or unknown lesion characterization (n = 25). All were referred for MR imaging after CT or sonography. Patient demographics (age, sex, history of malignancy), lesion size and morphology, quantitative T2 calculation, and pattern of enhancement on gadopentetate dimeglumine administration were evaluated for predictive ability. RESULTS: Thirty-two liver lesions were malignant (eight colon cancer, five breast cancer, four cervical cancer, three renal cancer, three lung cancer, and nine miscellaneous cancers), 53 were benign (37 hemangiomas, 15 cysts, and one focal nodular hyperplasia). Calculated T2 relaxation times (mean +/- standard deviation [SD]) were as follows: malignant tumors (91.72 +/- 21.9 msec), hemangiomas (136.1 +/- 26.3 msec), cysts (284.1 +/- 38.2 msec) (p < 0.001). Logistic regression analysis indicated that lesion size and sex and age of patient were not significant independent predictors (p > 0.05). However, the combination of a history of malignancy, T2 value, and gadopentetate dimeglumine-enhancement pattern allowed generation of a prediction rule with an area under the receiver operating characteristic curve of 0.95. The patient's weight, lesion morphology, and cell type of the primary malignancy did not provide additional predictive information (p > 0.2). CONCLUSION: We recommend using the combination of T2 quantification and patient history of malignancy before deciding to administer gadopentetate dimeglumine for optimal lesion characterization, especially for equivocal lesions with T2 values between 90 and 130 msec. These factors allowed the construction of a prediction rule for lesion characterization. PMID- 11264071 TI - Primary biliary cirrhosis: MR imaging findings and description of MR imaging periportal halo sign. AB - OBJECTIVE: This study reviews the prevalence of MR imaging abnormalities seen in 21 consecutive patients with primary biliary cirrhosis before transplantation and describes a new MR imaging sign in these patients: the MR imaging periportal halo sign. CONCLUSION: Abdominal adenopathy was present in 62% of the patient population, and none of the patients with adenopathy had a known malignancy. Findings associated with end-stage cirrhosis and portal hypertension were seen and included ascites (62%), splenomegaly (71%), portosystemic collaterals (57%), portal vein thrombosis (5%), and hepatocellular carcinoma (5%). The MR imaging periportal halo sign was seen in 43% of patients with primary biliary cirrhosis, but none of the patients in a sex- and age-matched cohort of 21 patients with cirrhosis not caused by primary biliary cirrhosis had the finding. Statistical analysis of these results produced a t score of 3.97 and a p value of less than 0.001, suggesting that this new MR imaging sign is highly specific for the diagnosis of primary biliary cirrhosis. PMID- 11264072 TI - Periportal contrast enhancement and abnormal signal intensity on state-of-the-art MR images. PMID- 11264074 TI - Three-dimensional portography using multislice helical CT is clinically useful for management of gastric fundic varices. AB - OBJECTIVE: This study seeks to evaluate three-dimensional (3D) helical CT portography as a tool for examining patients with gastric fundic varices. SUBJECTS AND METHODS: We compared 3D helical CT portography and conventional angiographic portography in 30 consecutive patients with gastric fundic varices. We assessed whether 3D helical CT portography is useful in selecting patients and in evaluating the results of balloon-occluded retrograde transvenous obliteration. RESULTS: Three-dimensional helical CT portography simultaneously depicted second or third branches of the intrahepatic portal vein and provided images of entire portosystemic collaterals. On 3D helical CT portography, gastric fundic varices were seen in 30 patients (100%), left gastric veins in 19 (63%), posterior gastric veins or short gastric veins in 28 (93%), gastrorenal shunts in 27 (90%), paraumbilical veins in three (10%), and inferior phrenic veins in two patients (7%). Findings of 3D helical CT portography and conventional angiographic portography were in close agreement. However, in four patients, posterior gastric veins or short gastric veins were not seen on conventional angiographic portography images of the spleen, but they were clearly revealed on 3D helical CT portography. Treatment was successful in all patients except one. Three-dimensional helical CT portography could easily evaluate therapeutic results. CONCLUSION: Three-dimensional helical CT portography proved so effective that it can be considered a less invasive alternative than conventional angiographic portography in assessing portosystemic collaterals. CT portography is useful in selecting candidates from patients with gastric fundic varices for retrograde transvenous obliteration and also in evaluating therapeutic results. PMID- 11264075 TI - Volumetric mangafodipir trisodium-enhanced cholangiography to define intrahepatic biliary anatomy. PMID- 11264076 TI - Doppler sonography of hepatic arterial blood flow velocity after percutaneous transhepatic portal vein embolization. AB - OBJECTIVE: This study was conducted to elucidate the changes in hepatic arterial blood flow after portal vein embolization. SUBJECTS AND METHODS: We prospectively measured the flow velocity and resistive index of the common, right, and left hepatic arteries, using Doppler sonography, in 21 patients who underwent embolization of the right portal vein. The measurements were performed before and 1, 3, 5, 7, and 14 days after embolization. We assessed the changes in liver volume with a volumetric study using CT. RESULTS: After embolization, flow velocity in the common hepatic artery increased significantly (p < 0.0001). Flow velocity in the right hepatic artery also increased significantly (p < 0.0001), with a significant decrease in resistive index (p < 0.0001). The flow velocity and resistive index of the left hepatic artery were unchanged. The increase in flow velocity in the right hepatic artery significantly correlated with that in the common hepatic artery (r = 0.514, p < 0.05). The calculated volume of the embolized right hepatic lobe significantly (p < 0.0001) decreased, from 685 +/- 32 cm(3) before embolization to 568 +/- 28 cm(3) after embolization. The atrophy rate of the right hepatic lobe significantly correlated with the increase in flow velocity in the right hepatic artery (r = 0.700, p < 0.0005). CONCLUSION: Portal vein embolization induces an increase in hepatic arterial blood flow velocity in the embolized hepatic segments, resulting from an increase in common hepatic arterial flow, but not from a steal phenomenon due to decreased hepatic arterial blood flow in the nonembolized hepatic segments. This observation may be explained by the simple mechanical effect of interposing a slower flowing stream (portal flow) in the path of a faster flowing stream (arterial flow). PMID- 11264077 TI - Ability of MR cholangiography to reveal stent position and luminal diameter in patients with biliary endoprostheses: in vitro measurements and in vivo results in 30 patients. AB - OBJECTIVE: Our goal was to evaluate the ability of MR cholangiography to show stent position and luminal diameter in patients with biliary endoprostheses. MATERIALS AND METHODS: Susceptibility artifacts were evaluated in vitro in three different stent systems (cobalt alloy-based, nitinol-based, and polyethylene) using two breath-hold sequences (rapid acquisition with relaxation enhancement, half-Fourier acquisition single-shot turbo spin echo) on a 1.5-T MR imaging system. The size of the stent-related artifact was measured, and the relative stent lumen was calculated. In vivo stent position and patency were determined in 30 patients (10 cobalt alloy-based stents, five nitinol-based stents, and 15 polyethylene stents). RESULTS: In vitro, the susceptibility artifact of the cobalt stent caused complete obliteration of the stent lumen. The relative stent lumens of the nitinol-based and polyethylene stents were 38-50% and 67-100%, respectively. In vivo, all stents were patent at the time of imaging. The position of the cobalt alloy-based stent could be determined in nine of 10 patients, but stent patency could not be evaluated. Stent position of nitinol stents could not be adequately evaluated in any of the five patients, and internal stent diameter could be visualized in only one patient. In nine of 15 patients, the fluid column within the implanted polyethylene stent was seen on MR cholangiography. CONCLUSION: The internal stent lumen could be visualized in most patients with an indwelling polyethylene stent, but not in patients with cobalt alloy- or nitinol-based stents. PMID- 11264078 TI - Osler-Weber-Rendu disease: visualizing portovenous shunting with three dimensional sonography. PMID- 11264079 TI - Making sense of mucin-producing pancreatic tumors. PMID- 11264080 TI - Three-dimensional double-contrast MR colonography: a display method simulating double-contrast barium enema. PMID- 11264081 TI - Comparative assessment of CT and sonographic techniques for appendiceal imaging. AB - OBJECTIVE: We performed a comparative assessment of CT and sonographic techniques used to assess appendicitis. MATERIALS AND METHODS: One hundred patients with clinically suspected acute appendicitis were examined with sonography, unenhanced focused appendiceal CT, complete abdominopelvic CT using IV contrast material, focused appendiceal CT with colonic contrast material, and repeated sonography with colonic contrast material. Each sonogram was videotaped for subsequent interpretation by three radiologists and two sonographers. The mean sensitivity, specificity, positive and negative predictive values, inter- and intraobserver variability, and diagnostic confidence scores of all observers were used for comparative performance assessments. The three CT examinations were filmed and interpreted separately by four radiologists. Patient discomfort was assessed on a 10-point scale for each radiologic study. Diagnoses were confirmed by pathologic evaluation of resected appendixes or clinical follow-up for a minimum of 3 months after presentation. RESULTS: Twenty-four of the 100 patients had positive findings for acute appendicitis. Both sonographic techniques had high specificity (85-89%) and comparable accuracy (73-75%) but low sensitivity (33-35%) and inter- and intraobserver variability (kappa = 0.15-0.20 and 0.39-0.42, respectively). Unenhanced focused appendiceal CT, abdominopelvic CT, and focused appendiceal CT with colonic contrast material all significantly outperformed sonography (p <0.0001), with sensitivities of 78%, 72%, and 80%; specificities of 86%, 91%, and 87%; and accuracies of 84%, 87%, and 85%, respectively. Abdominopelvic CT gave the greatest confidence in cases with negative findings (p = 0.001), and focused appendiceal CT with colonic contrast material gave the greatest confidence for cases with positive findings (p = 0.02). In terms of inter- and intraobserver variability, focused appendiceal CT with colonic contrast material yielded the highest, and unenhanced focused appendiceal CT the lowest, agreement (interobserver kappa = 0.45 vs. 0.36 and intraobserver kappa = 0.85 vs. 0.76, respectively) (p <0.05). Colonic contrast material was unsuccessfully advanced into the cecum in 18% of patients and leaked in another 24%. Patient discomfort was greatest with focused appendiceal CT using colonic contrast material and least with unenhanced focused appendiceal CT (p <0.05). CONCLUSION: A standard abdominopelvic CT scan is recommended as the initial examination for appendicitis in adult patients. However, focused appendiceal CT with colonic contrast material material should be used as a problem-solving technique in difficult cases. PMID- 11264083 TI - Value of double-contrast barium enema performed immediately after incomplete colonoscopy. AB - OBJECTIVE: The purpose of this study was to evaluate the ease, completeness, and clinical utility of double-contrast barium enema (DCBE) performed immediately after incomplete colonoscopy. SUBJECTS AND METHODS: During a 30-month period, a prospective study was performed in 103 patients (79 women, 24 men) to determine the ease and completeness of DCBE immediately after failed colonoscopy and any additional useful information provided by the enema. The ease with which DCBE was performed was graded from 1 (easy) to 10 (difficult). RESULTS: DCBE revealed the entire colon in 97 patients (94%). Incomplete DCBE was a result of obstruction and incontinence in three patients each. The mean score for ease of performing DCBE was 5.0. In 14 patients (14%), significant additional diagnostic information was provided by the immediate DCBE. In eight patients, abnormalities were identified on DCBE that had not been seen at colonoscopy (five malignant neoplasms, one diverticular mass, two extrinsic masses, and multiple strictures). In four patients, a suspected colonoscopic abnormality was excluded with DCBE findings; and in two patients, a colonoscopic abnormality was further characterized with DCBE. CONCLUSION: Immediate DCBE after incomplete colonoscopy allows complete colonic evaluation in most cases, often adds vital diagnostic information, and eliminates repeated bowel preparation and unnecessary delay in diagnosis. PMID- 11264084 TI - Carcinoid tumors of the stomach: a clinical and radiographic study. AB - OBJECTIVE: Our purpose is to describe associated and coexistent diseases of gastric carcinoid tumors, the unique biologic behavior of these tumors, the appearance of these tumors on fluoroscopic and CT images, and the radiologic management of these neoplasms. CONCLUSION: First, multiple gastric carcinoid tumors are associated with enterochromaffin-like cell hyperplasia, chronic atrophic gastritis, and pernicious anemia and have a low risk of malignancy. Second, solitary gastric carcinoid tumors, or gastric carcinoid tumors associated with multiple endocrine neoplasia-type I (MEN-I) and Zollinger-Ellison syndrome, have a higher potential for metastatic disease. Third, the radiologic appearance and management of these tumors depend on the clinical background of the patient. PMID- 11264086 TI - Wide-mouthed sacculations in the esophagus: a radiographic finding in scleroderma. PMID- 11264087 TI - The spectrum of abdominal venous CT findings in blunt trauma. PMID- 11264088 TI - Ultrafast MR imaging of the pelvic floor. AB - OBJECTIVE: The aim of this study was to compare pelvic floor anatomy and laxity at rest and on straining (Valsalva's maneuver) using dynamic ultrafast MR imaging in women who were continent versus those with stress incontinence differing in obstetric history. MATERIALS AND METHODS: Thirty continent women were divided into three equal groups (nulliparous, previous cesarean delivery, previous vaginal delivery) and compared with 10 women with stress-incontinence with a history of at least one vaginal delivery. MR imaging of the pelvic floor at rest and on maximal strain was performed, using axial T2-weighted fast spin-echo images followed by sagittal ultrafast T2-weighted single-shot fast spin-echo sequences. Mean population age (age range, 22-45 years; mean +/- SD, 36 +/- 5.4 years), was similar in the four groups, as was parity in the three parous groups. RESULTS: Mean distances between the bladder floor and pubococcygeal line at rest did not differ between the four groups. On straining, bladder floor descent was 1.1 +/- 0.9, 1.0 +/- 1.1, and 1.9 +/- 0.9 cm in continent nulliparous, cesarean delivery, and vaginal delivery women, respectively, versus 3.2 +/- 1.0 cm in incontinent women (p = 0.0005). Cervical descent was greater in incontinent versus nulliparous women (p = 0.0019). Bladder floor descent was greater in the continent vaginal delivery group than in continent cesarean delivery control patients (p = 0.04). In patients with stress incontinence, symptoms did not correlate with amplitude of descent. The right levator muscle was thinner overall than the left, regardless of frequency direction (p = 0.001). CONCLUSION: Ultrafast MR imaging using the T2-weighted single-shot fast spin-echo sequence allows dynamic evaluation of the pelvic compartments at maximal strain with no need for contrast medium. Pelvic floor laxity and supporting fascia abnormalities were most common in patients with stress incontinence followed by continent women with a history of vaginal delivery. The results are therefore compatible with the hypothesis of vaginal delivery as a contributory factor to stress incontinence in older parous women. PMID- 11264090 TI - Differences of renal parenchymal attenuation for acutely obstructed and unobstructed kidneys on unenhanced helical CT: a useful secondary sign? AB - OBJECTIVE: The purpose of this study was to ascertain whether the difference in attenuation frequently noted on unenhanced helical CT scans between a patient's acutely obstructed kidney and the unobstructed kidney is a reliable secondary sign of acute renal obstruction. CONCLUSION: In 95% of patients with acute renal obstruction, the affected kidney was less dense than the unobstructed kidney. When visually detected by radiologists using CT, this difference in density was at least two standard deviations above normal, making it a reliable secondary sign for acute obstruction. PMID- 11264091 TI - Giant cystic Schmorl's nodes: imaging findings in six patients. AB - OBJECTIVE: We describe the imaging findings of an unusual type of Schmorl's node appearing as giant cystlike lesion of the vertebral bodies. CONCLUSION: Giant cystic Schmorl's nodes are unusual entities; their radiologic appearance differs dramatically from the classic description and is diagnostically challenging. However, the appearance of these nodes on conventional radiographs and especially on MR images is characteristic. Knowledge of this entity would help to eliminate unnecessary invasive diagnostic or therapeutic procedures. PMID- 11264092 TI - Volume rendering of tendon-bone relationships using unenhanced CT. AB - OBJECTIVE: Clinically, three-dimensional CT of the extremities is most often used to display bony anatomy. However, by combining unenhanced CT with volume rendering computer graphics, visualization of relationships between bone and soft tissue structures such as muscle tendon is also possible. The aims of this study were to quantify CT attenuation values of peripheral tendon, muscle, and bone on unenhanced CT and to develop custom opacity transforms on the basis of the attenuation measurements to effectively depict tendon-muscle-bone relationships. CONCLUSION: The mean attenuation of peripheral tendon ( approximately 100 H) is distinctly higher than that of muscle ( approximately 60 H) enabling high-quality volume rendering of muscle-tendon-bone relationships with unenhanced CT. High frequency (bone) CT reconstruction algorithms commonly used for extremity CT produce approximately twofold higher image noise and inferior three-dimensional renderings compared with those based on less noisy standard or soft-tissue reconstruction algorithms. These concepts can be used to uniquely reveal tendon muscle-bone relationships for clinical, scientific, and educational purposes. PMID- 11264093 TI - A complex atlantoaxial fracture with craniocervical instability: a case with bilateral type 1 dens fractures. PMID- 11264094 TI - Musculoskeletal imaging with multislice CT. PMID- 11264096 TI - Radiologic placement of implantable chest ports in pediatric patients. AB - OBJECTIVE: We evaluated the technical success and complications associated with radiologic placement of implantable chest ports in children for long-term central venous access. MATERIALS AND METHODS: Between May 1, 1996 and January 11, 2000, 29 chest ports were placed in 28 children (15 girls, 13 boys; age range, 2-17 years; mean, 11.7 years). The patient's right internal jugular vein was used for access in 93% (27/29) of the procedures, and a collateral neck vein was used as a conduit to recanalize the central veins in two procedures because of bilateral jugular and subclavian vein occlusion. All procedures were performed in interventional radiology suites. Both real-time sonography and fluoroscopy were used to guide venipuncture and port insertion. Follow-up data were obtained through the clinical examination and electronic review of charts. RESULTS: Technical success was 100%. Fourteen percent of the catheters were removed prematurely, including one catheter removed 17 days after placement because the patient's blood cultures were positive for Candida albicans. No patients experienced hematoma, symptomatic air embolism, symptomatic central venous thrombosis, catheter malposition, or pneumothorax. The median number of days for catheter use by patients was 280 days (total, 9043 days; range, 17-869 days). The rate of confirmed catheter-related infection was 14% or 0.04 per 100 venous access days. One catheter occluded after 132 days. CONCLUSION: In pediatric patients, radiologists can insert implantable chest ports using real-time sonographic and fluoroscopic guidance with high rates of technical success and low rates of complication. PMID- 11264095 TI - Rate of abnormal osteoarticular radiographic findings in pediatric patients. AB - OBJECTIVE: The objective of our study was to assess the rate of abnormal radiographic findings in the most frequent osteoarticular locations of traumatic injury in a pediatric population. SUBJECTS AND METHODS: During two periods of 12 weeks each, all patients admitted to the pediatric emergency department for osteoarticular trauma who underwent radiography were prospectively included in this study. A connection was drawn between the rate of abnormal radiographic findings for the seven most frequently radiographed locations and the clinical findings. RESULTS: Of 3128 locations of trauma in 2470 children, only 22% of the radiographic examinations were considered to reveal abnormal findings. In decreasing order, the hand and fingers, the ankle, the wrist, the knee, the elbow, the foot and toes, and the forearm were the most frequently examined locations. The rate of abnormal findings was 25.7% for the hand and fingers, 9.0% for the ankle, 42.5% for the wrist, 9.5% for the knee, 33.3% for the elbow, 18.3% for the foot, and 43.2% for the forearm. When only the direct sign of fracture was taken into account, these rates decreased for the ankle and knee to 2.6% and 1.9%, respectively. There was always a significant link between the degree of clinical suspicion and the rate of abnormal radiographic findings. However, fewer than 50% of the cases with high clinical suspicion of fracture were radiographically confirmed. CONCLUSION: It appears necessary, especially in cases of lower limb trauma, to evaluate clinical tests, including the implementation of the Ottawa ankle rules, to reduce the number of unnecessary radiographic examinations. This reduction will improve some parameters of children's quality of life and will significantly decrease the cost of emergency care. PMID- 11264097 TI - High-frequency linear array transducers for neonatal cerebral sonography. PMID- 11264098 TI - The anterior iliac separation: alternative index for pelvic morphometry in fetuses with Down syndrome. AB - OBJECTIVE: The goal of this study was to assess the diagnostic use of an anterior iliac separation measurement as an alternative index for the iliac angle in the assessment of fetal pelvic morphometry. SUBJECTS AND METHODS: In 358 fetuses, the anterior iliac separation, iliac length, and iliac angle were prospectively measured on antenatal sonography. All measurements were obtained at two axial levels (superior and inferior). The gestational age of the fetus was recorded. The anterior iliac separation was normalized by iliac length, and coefficients of variation were calculated for all measurements. The effects of axial level and gestational age were assessed in a linear regression model. The diagnostic use of the anterior iliac separation relative to that of the iliac angle was assessed in a comparison of 24 fetuses with Down syndrome and 247 non-Down syndrome fetuses. RESULTS: The anterior iliac separation was less variable than the iliac angle at both superior and inferior levels. There were statistically significant effects for gestational age and axial level on both the anterior iliac separation and the iliac angle, but there was no significant effect for either factor when the anterior iliac separation was normalized by the iliac length. Comparing Down and non-Down syndrome fetuses, we found that the normalized anterior iliac separation had discriminating power similar to the iliac angle. CONCLUSION: The linear measurement of the anterior iliac separation has diagnostic properties similar to the iliac angle and is subject to less measurement variability. This simpler measurement may be particularly useful when normalized by the iliac length. PMID- 11264100 TI - Auditory functional MR imaging. PMID- 11264101 TI - Imaging cerebral vasculitis in refractory epilepsy using [(11)C](R)-PK11195 positron emission tomography. PMID- 11264102 TI - Comparison of sonography and CT for differentiating benign from malignant cervical lymph nodes in patients with squamous cell carcinoma of the head and neck. AB - OBJECTIVE: We compared the ability of sonography and CT to differentiate benign from malignant cervical lymph nodes in patients with squamous cell carcinoma of the head and neck. MATERIALS AND METHODS: We analyzed 209 cervical nodes (102 metastatic and 107 nonmetastatic) from 62 patients with head and neck cancer. These nodes were topographically correlated by node between images and surgical specimens, and accordingly between sonography and CT. RESULTS: The area under the receiver operating characteristic curve (A(z) value) for the overall impressions of metastatic or nonmetastatic nodes was significantly greater for sonography (power Doppler sonography plus gray-scale sonography, 0.97 +/- 0.005; gray-scale sonography, 0.95 +/- 0.004) than for CT (0.87 +/- 0.018). Receiver operating characteristic curve analysis also showed that the greater ability of sonography to depict the internal architecture of the nodes (A(z) value, 0.96 +/- 0.006) compared with CT (A(z) value, 0.81 +/- 0.027) significantly contributed to the better performance of sonography compared with CT in diagnosing metastatic nodes in the neck. On the other hand, size criterion (the short-axis diameter) was equally predictive in sonography and CT. The greater contributions of internal architectures relative to the size criterion of the node in the sonographic assessment for metastatic nodes were further evidenced by the findings that sonography provided higher sensitivity and specificity than CT did, whereas the cutoff points for the short-axis diameter in both tests were equivalent. CONCLUSION: Sonography performed significantly better than CT in depicting cervical metastatic nodes. Sonography could be a useful adjunct to CT in surveying cervical metastatic nodes. PMID- 11264103 TI - Treatment of acute cholecystitis in non-critically ill patients at high surgical risk: comparison of clinical outcomes after gallbladder aspiration and after percutaneous cholecystostomy. AB - OBJECTIVE: This study was performed to compare the clinical outcome after gallbladder aspiration with that after percutaneous cholecystostomy in non critically ill patients with acute cholecystitis who were at high risk from surgery. MATERIALS AND METHODS: Medical records of 53 consecutive non-critically ill, high-surgical-risk patients admitted with acute cholecystitis between July 1995 and July 1999 were reviewed. Thirty-one had gallbladder aspiration and 22 had percutaneous cholecystostomy. The primary outcome measure of clinical response within 72 hr and the secondary outcome measures of overall positive response rate, complication rate, time to resolution, and rate of recurrence of acute cholecystitis were compared between the two groups. RESULTS: Gallbladder aspiration and percutaneous cholecystostomy were technically successful in 30 (97%) and 21 (97%) patients, respectively; of these, 23 (77%) and 19 (90%) patients responded clinically within 72 hr (p > 0.2). Complications occurred in three patients (12%) after percutaneous cholecystostomy and in none after gallbladder aspiration (p < 0.05). No significant difference was noted in the other secondary outcome measures of the two groups. CONCLUSION: We found no significant difference in the clinical outcomes of gallbladder aspiration and percutaneous cholecystostomy in the treatment of acute cholecystitis in high surgical-risk patients who are not critically ill. However, we found gallbladder aspiration to be significantly safer. Therefore, gallbladder aspiration should be the procedure of choice in high-risk patients with acute cholecystitis who are not critically ill, and percutaneous cholecystectomy should be reserved as a salvage procedure if gallbladder aspiration is technically or clinically unsuccessful. PMID- 11264105 TI - Using dual-detector helical CT angiography to detect deep venous thrombosis in patients with suspicion of pulmonary embolism: diagnostic value and additional findings. AB - OBJECTIVE: The purpose of this study was to assess the value of dual-slice helical CT angiography in detecting deep venous thrombosis in patients in whom acute pulmonary embolism was suspected and to describe the additional extrathoracic findings. SUBJECTS AND METHODS: Sixty-five consecutive patients were examined for suspected pulmonary embolism using helical CT of the chest (2.7 mm collimation; table speed, 7.5 mm/sec; 100-140 mL of contrast medium injected at a rate of 3 mL/sec) followed by CT of the lower limbs (6.5-mm collimation; table speed, 10 mm/sec) without any additional contrast medium injection. Sequential scanning of the abdomen was performed using 10-mm collimation and an interval of 40 mm. Color Doppler sonography of the lower limbs was done within 24 hr of CT by two radiologists who were unaware of CT findings. Results of CT venography were compared with those of Doppler sonography and with phlebography or repeated focalized sonography in cases of discrepancy. RESULTS: Twenty-two patients had pulmonary embolism revealed on chest CT. Sixteen patients had a deep venous thrombosis. Thirteen patients with pulmonary embolism had a deep venous thrombosis. Three patients with deep venous thrombosis had no pulmonary embolism. Sensitivity and specificity for diagnosing deep venous thrombosis with CT was 93% and 97%, respectively (kappa = 0.88). Additional extrathoracic findings were observed in four patients. CONCLUSION: Combined CT venography with dual-slice scanning is an accurate method to diagnose deep venous thrombosis that may reveal additional imaging findings in some patients with possible pulmonary embolism. PMID- 11264104 TI - Sonographically guided percutaneous thrombin injection for treatment of a vein graft pseudoaneurysm. PMID- 11264106 TI - Treatment of chylothorax: percutaneous catheterization and embolization of the thoracic duct. PMID- 11264107 TI - Thromboembolic disease: variability of interobserver agreement in the interpretation of CT venography with CT pulmonary angiography. AB - OBJECTIVE: The objective of this study was to determine interobserver agreement in the diagnosis of acute deep venous thrombosis on CT venography performed in addition to CT pulmonary angiography. SUBJECTS AND METHODS: One hundred forty-six CT venograms of 144 patients (mean age, 61.74 years) clinically suspected of having pulmonary embolism were analyzed prospectively and independently by two experienced thoracic and body imaging radiologists and later by consensus of the two radiologists. The CT venography protocol consisted of 5-mm-thick axial images at 20-mm intervals from the popliteal fossa to the renal veins. Images were acquired 3-4 min after the start of 100-150 mL of undiluted contrast medium administration at 4 mL/sec. Thirteen venous segments were analyzed in each patient. There were 1586 analyzable venous segments. RESULTS: Interobserver agreement, with the patient as the unit of analysis, was moderately good (kappa, 0.59; 95% confidence interval [CI], 0.39-0.78). Kappa values were similar for CT venography studies performed with 150 mL of contrast medium and 4-min delay (kappa, 0.62; 95% CI, 0.30-0.88) and with 3-min delay and 100 mL of contrast medium (kappa, 0.56; 95% CI, 0.32-0.80). Interobserver disagreement occurred in 17 (12%) of 146 CT venography studies. Findings of 11 CT venography studies were interpreted as negative, and six were interpreted as positive after consensus interpretation. CONCLUSION: Interobserver agreement for deep venous thrombosis with CT venography is moderately good. PMID- 11264109 TI - Lower extremity venous sonography in the high-risk cancer population: one leg or two? AB - OBJECTIVE: We correlated the diagnostic yield of unilateral and bilateral lower extremity venous sonograms in a high-risk cancer population with the clinical indication for the examination. MATERIALS AND METHODS: Reports from 433 bilateral and 619 unilateral lower extremity Doppler sonograms obtained over an 18-month period in patients with cancer were retrospectively reviewed, and clinical indication and findings were determined. RESULTS: Overall, 228 (22%) of 1052 examinations revealed deep venous thrombosis (DVT): 83 (19%) of 433 bilateral and 145 (23%) of 619 unilateral. Among studies performed for unilateral symptoms (pain, edema, or postorthopedic procedure), 23% (135/581) of unilateral and 27% (44/162) of bilateral studies revealed DVT. Among these 44 bilateral studies with positive findings performed for unilateral symptoms, there were 30 DVT in the symptomatic side, 12 bilaterally, and two in the asymptomatic side alone. Ten percent (11/110) of the bilateral studies performed for bilateral symmetric symptoms revealed DVT. Among studies performed for bilateral asymmetric symptoms, 13% (1/8) of the unilateral and 8% (2/25) of the bilateral studies revealed DVT; both bilateral studies showed positive findings in the more symptomatic side. Among studies performed for suspected or proven pulmonary embolus, 20% (23/113) of bilateral and 54% (7/13) of unilateral studies had positive findings. CONCLUSION: In a high-risk cancer population, the incidence of DVT in patients with unilateral symptoms is more than twice that of patients with bilateral symptoms. Because DVT isolated to an asymptomatic lower extremity is rare (1%), bilateral sonographic examination is generally unnecessary with unilateral lower extremity symptoms. PMID- 11264110 TI - Bronchiolitis obliterans with organizing pneumonia versus chronic eosinophilic pneumonia: high-resolution CT findings in 81 patients. AB - OBJECTIVE: The objective of this research was to compare high-resolution CT findings of bronchiolitis obliterans with organizing pneumonia (BOOP) with those of chronic eosinophilic pneumonia (CEP) and to determine whether high-resolution CT can differentiate the two. MATERIALS AND METHODS: We retrospectively reviewed high-resolution CT scans of 38 patients with BOOP and 43 patients with CEP. Without knowledge of the diagnosis, two radiologists evaluated the frequency and distribution of high-resolution CT findings in both groups of patients and made a diagnosis using a three-point scale of confidence. RESULTS: Nodules, nonseptal linear or reticular opacities, and bronchial dilatation were significantly more common in BOOP than in CEP (31.6% vs. 4.7%, p < 0.005; 44.7% vs. 9.3%, p < 0.001; and 57.9% vs. 25.6%, p < 0.005, respectively). Septal line thickening was more frequent in CEP than in BOOP (72.1% vs. 39.5%, p < 0.005). Peribronchial distribution of consolidation was more frequent in BOOP than in CEP (28.9% vs. 9.3%, p < 0.05). A correct diagnosis was made in 69.7% of cases, and the diagnostician was confident in 21.7%. Interobserver agreement was good (kappa = 0.6). CONCLUSION: Although several of the high-resolution CT findings of BOOP and CEP are different, these diseases are differentiated with confidence in only a small percentage of cases. PMID- 11264111 TI - Topographic anatomy of the vertebral venous system in the thoracic inlet. PMID- 11264112 TI - Electron beam CT in the diagnosis of recurrent cardiac lipoma. PMID- 11264113 TI - In vitro high-resolution helical CT of small axillary lymph nodes in patients with breast cancer: correlation of CT and histology. AB - OBJECTIVE: This study seeks to determine whether high-resolution in vitro helical CT can show the internal structure of small axillary nodes and to establish the CT characteristics of benign versus metastatic axillary nodes in patients with breast cancer. SUBJECTS AND METHODS: We obtained in vitro helical CT images of 212 nodes excised from 19 patients with breast cancer. The longest mean size was 5.9 mm, and the range was 0.5 to 26.5 mm. The hilar and cortical characteristics, the size, and the ratio of the longest axis to the shortest axis were evaluated. CT findings were correlated with histologic findings. RESULTS: Pathologic assessment of excised nodes with a central low-density hilum visualized on CT showed arteries, veins, lymphatic sinuses, and fatty tissue. A peripheral high density cortex on CT contained mostly lymphatic tissue. Abnormal (eccentric, irregular) cortices were observed in malignant nodes (p<0.0001). Marked differences were observed among the proportions of benign and malignant nodes when the ratio of the longest axis to the shortest axis was less than 2 and an abnormal cortex was observed. CT could also detect extracapsular lymph node extension. CONCLUSION: In vitro high-resolution helical CT can detect the internal structure of small nodes. Morphologic changes detected on helical CT help distinguish benign from malignant nodes. PMID- 11264114 TI - Physician training requirements in sonography. PMID- 11264115 TI - Clip placement during sonographically guided breast biopsy. PMID- 11264116 TI - "Mini brain" of plasmacytoma. PMID- 11264117 TI - Sonography versus CT for monitoring radiofrequency ablation. PMID- 11264118 TI - Possible duplicate publication: AJR 2000;174:1698 and Clin Orthop 2000;373:311 316, 318-319. PMID- 11264119 TI - The radiology job market: good news and bad news for residents. PMID- 11264120 TI - The architecture of the corneal stroma. PMID- 11264121 TI - Discontinuing anticytomegalovirus therapy in patients with cytomegalovirus retinitis and AIDS. PMID- 11264122 TI - Are ophthalmologists overtrained? PMID- 11264123 TI - Can we eliminate trachoma? PMID- 11264124 TI - The Bhaktapur eye study: ocular trauma and antibiotic prophylaxis for the prevention of corneal ulceration in Nepal. AB - AIMS: To determine the incidence of ocular trauma and corneal ulceration in the district of Bhaktapur in Kathmandu Valley, and to determine whether or not topical antibiotic prophylaxis can prevent the development of ulceration after corneal abrasion. METHODS: A defined population of 34 902 individuals was closely followed prospectively for 2 years by 81 primary eye care workers who referred all cases of ocular trauma and/or infection to one of the three local secondary eye study centres in Bhaktapur for examination, treatment, and follow up by an ophthalmologist. All cases of ocular trauma were documented and treated at the centres. Individuals with corneal abrasion confirmed by clinical examination who presented within 48 hours of the injury without signs of corneal infection were enrolled in the study and treated with 1% chloramphenicol ophthalmic ointment to the injured eye three times a day for 3 days. RESULTS: Over the 2 year period there were 1248 cases of ocular trauma reported in the population of 34 902 (1788/100 000 annual incidence) and 551 cases of corneal abrasion (789/100 000 annual incidence). The number of clinically documented corneal ulcers was 558 (799/100 000 annual incidence). Of the 442 eligible patients with corneal abrasion enrolled in the prophylaxis study, 424 (96%) healed without infection, and none of the 284 patients who were started on treatment within 18 hours after the injury developed ulcers. Four of the 109 patients (3.7%) who presented 18-24 hours after injury developed infections, and 14 (28.6%) of the 49 patients who presented 24-48 hours subsequently developed corneal ulceration. CONCLUSIONS: Ocular trauma and corneal ulceration are serious public health problems that are occurring in epidemic proportions in Nepal. This study conclusively shows that post-traumatic corneal ulceration can be prevented by topical application of 1% chloramphenicol ophthalmic ointment in a timely fashion to the eyes of individuals who have suffered a corneal abrasion in a rural setting. Maximum benefit is obtained if prophylaxis is started within 18 hours after injury. PMID- 11264126 TI - The a-wave of the dark adapted electroretinogram in glaucomas: are photoreceptors affected? AB - AIMS: To evaluate whether the a-wave of the dark adapted flash electroretinogram (ERG) is affected by glaucomatous damage. METHODS: ERGs were recorded in 20 patients (age 33-65 years) with advanced glaucomas (primary and secondary open angle and low tension glaucomas) and 20 normals using a ganzfeld stimulus. After 30 minutes of dark adaptation and pupil dilatation to at least 7.5 mm in diameter, luminance response functions were obtained presenting white flashes of increasing scotopic luminance (the highest flash intensity being 9.4 cd/s/m2, the lowest being 5.75 log units below it) with an interflash interval of 5 seconds. For each scotopic luminance, the responses of four flashes were averaged. The a wave's amplitude was measured at 10, 11, and 12 ms. Within the glaucoma group, correlations between the interocular differences of the a-wave's amplitude and the mean deviation of a static perimetry (Octopus 500 perimeter, program G1) were computed for all flash intensities. Between normals and glaucomas, the a-wave's amplitude was compared for all flash intensities (paired t test). RESULTS: Within the glaucoma group, the interocular differences of the a-wave's amplitudes correlated significantly with the differences of the MD for flash intensities of 9.4, 5.3, 1.7, and 0.5 cd/s/m2. The a-wave's amplitude was significantly lower in the glaucoma compared with the normal group (p <0.005) for flash intensities of 9.4 and 5.3 cd/s/m2. CONCLUSION: These electrophysiological results imply that also the outer retinal structures, especially the photoreceptors, may be affected by glaucomatous damage. PMID- 11264125 TI - The epithelial flap for photorefractive keratectomy. AB - BACKGROUND/AIMS: Epithelial debridement for photorefractive keratectomy (PRK) is associated with pain, slower visual recovery, and may be aetiological in haze production. The aim of this study was to assess the clinical results of a new technique involving raising and replacing of an epithelial flap in photorefractive keratectomy. METHODS: A prospective, non-randomised, comparative, paired eye trial was performed in 72 eyes of 36 patients who underwent PRK with a Nidek EC-5000 excimer laser. For epithelial debridement before PRK, the eyes were divided into two groups. The first eye of each patient was treated with 20% ethanol debridement and the second eye with an epithelial flap which was replaced after treatment. PRK was carried out with the same laser and nomogram in both groups by the same surgeon. Visual and refractive outcome of PRK treatment was compared in both groups. RESULTS: The mean (SD) preoperative mean spherical equivalent (MSE) was -3.61 (1.38) dioptres (D) (range -1.00 D to -7.88 D) with no significant difference between the two groups. After a mean follow up period of 62.6 weeks (range 52-70) the final MSE was +0.07 (0.61) D (range -5.50 D to +4.50 D) in the debridement group and -0.24 (0.43) D in the epithelial flap group. There was no statistically significant difference between the two groups in postoperative MSE. The best corrected visual acuity was better in the epithelial flap group at all visits; this difference was statistically significant (p<0.05). The corneal haze was less in the epithelial flap group and this difference was also statistically significant (p<0.05). CONCLUSIONS: Managing the corneal epithelium as a hinged flap with 20% ethanol is a safe technique with faster visual rehabilitation and reduced haze compared with debridement of the epithelium with alcohol. Further studies need to be performed to compare pain levels postoperatively with the epithelial flap and epithelial debridement. PMID- 11264127 TI - The b-wave of the dark adapted flash electroretinogram in patients with advanced asymmetrical glaucoma and normal subjects. AB - AIMS: To evaluate whether the b-wave of the dark adapted flash electroretinogram (ERG) is affected by glaucomatous damage. METHODS: ERGs were recorded in 35 patients aged 33-65 years with advanced asymmetrical glaucomas (interocular difference of perimetric defects (mean deviation) >2 dB between the two fellow eyes of the glaucoma patients, primary and secondary open angle and low tension glaucomas) and 17 normal subjects matched for age and sex using white flashes of a xenon discharge tube in a Ganzfeld stimulator. After 30 minutes of dark adaptation luminance response functions were obtained using flashes of increasing scotopic luminance (highest 9.4 cd/s/m2, lowest 5.5 log units below it). The parameters Vmax, n, and K of the Naka-Rushton equation were computed from the measurement values based on the usual fitting procedure. These parameters, together with b-wave amplitudes and implicit times for all flash intensities, were compared interocularly and between the normal subjects and those with glaucoma. Correlations were computed between interocular differences of the mean deviation and interocular differences of Vmax, n, K, b-wave amplitudes, and implicit times between the two fellow eyes of the patients with asymmetrical glaucomatous damage. RESULTS: Implicit times were significantly longer (p<0.005) in the glaucoma patients than in the normal group for flash intensities of 9.4, 5.3, 1.7, 0.53, and 0.17 cd/s/m2. b-Wave amplitudes did not differ significantly between the two study groups. Comparing the two fellow eyes of each patient with glaucoma, Vmax was significantly higher in the less damaged eye than in the more damaged eye. The interocular differences in the mean deviation correlated significantly with the interocular differences in the b-wave amplitudes, implicit times, and Vmax. CONCLUSIONS: These results suggest that glaucomas can lead to electrophysiologically measurable damage of the inner nuclear layer. PMID- 11264128 TI - Changes of cone electroretinograms to colour flash stimuli after successful retinal detachment surgery. AB - AIM: To examine the changes in the short wavelength (S) and mixed long (L) and middle (M) wavelength sensitive cone (L,M-cone) electroretinograms (ERGs) after successful retinal detachment surgery. METHODS: Cone ERGs elicited by different colour flashes were recorded from 19 eyes with unilateral rhegmatogenous retinal detachment treated successfully by conventional buckling surgery. Ganzfeld colour flashes on a bright white background were used to elicit S-cone and L,M-cone ERGs. The ratio (operated eye/fellow eye) of the S-cone b-wave elicited by a 450 nm stimulus and the ratio (operated eye/fellow eye) of the L,M-cone b-wave elicited by a 633 nm stimulus were evaluated preoperatively and 1, 3, and 6 months after surgery. RESULTS: Preoperatively, no significant difference was observed between the ratio of the S-cone ERG amplitudes and the ratio of the L,M cone ERG amplitudes. Postoperatively, the ratio of the L,M-cone ERGs increased significantly over the preoperative value (p=0.001) but the ratio of the S-cone ERG did not improve. There were significant differences between the ratios of the S-cone and the L,M-cone ERGs at 1, 3, and 6 months after surgery. The postoperative recovery of the S-cone ERG was significantly greater in eyes treated within 4 weeks after the onset of the detachment than in eyes treated later than 4 weeks. CONCLUSIONS: These results indicate that the impairment of the L,M-cone system caused by retinal detachment may be reversible. However, the S-cone system may have more profound permanent damage. PMID- 11264129 TI - Assessment of early retinal changes in diabetes using a new multifocal ERG protocol. AB - AIMS: To assess early functional retinal changes in diabetics without retinopathy, a new multifocal stimulus paradigm was used that emphasises fast adaptive response contributions. METHODS: 25 normal control subjects (25 eyes) and 11 diabetics without retinopathy (22 eyes) served as subjects. Stimulation and analysis were performed with Veris Science 4.0. A stimulation protocol was used that combines regular multifocal flicker stimulation with a periodic "global" flash inserted between the multifocal stimuli. The multifocal stimuli were presented four video frames apart. The global flash covered the entire screen in the third frame of the four frame interval. The remaining two frames were dark. The periodic global flashes could only contribute to the focal responses if they were affected by the multifocal stimulation. A non-linear component induced by the interaction of the focal and global flashes was observed. The differences between control subjects and diabetics were assessed in both the multifocal responses and their induced effect on the following global flashes. RESULTS: The responses to focal flashes were reduced significantly in diabetics matched in age to the control subjects. The induced components showed large intersubject variability in controls and patients, and did not differ significantly between the two groups. CONCLUSION: The periodic global flashes produce a greater multifocal response reduction in diabetics than in normals, indicating impairment in the rate or magnitude of recovery from the bright preceding stimulus. The new stimulation protocol reveals early changes in retinal function of diabetics. PMID- 11264130 TI - Ophthalmic manifestations of tuberous sclerosis: a population based study. AB - BACKGROUND/AIMS: Tuberous sclerosis complex (TSC) has retinal and non-retinal ophthalmic manifestations. This study was designed to determine the prevalence of the ophthalmic manifestations and of refractive errors in a population of patients with TSC. METHODS: 179 patients identified were in a prevalence study of TSC in the south of England and 107 of these agreed to full ophthalmic examination which was successful in 100. Ophthalmic examination included examination of the eyelids, cover test, examination of the irides, dilation funduscopy using both direct and indirect ophthalmoscopy, and refraction using retinoscopy. Myopia was defined as a spherical equivalent <-0.5D and hyperopia as a spherical equivalent >+0.5D. RESULTS: Retinal hamartomas were seen in 44 of the 100 patients. The commonest morphological type of hamartoma seen was the flat, translucent lesion in 31 of the 44 patients (70%). The multinodular "mulberry" lesion was seen in 24 of the 44 patients (55%) and the transitional type lesion was seen in four of the 44 patients (9%). Punched out areas of retinal depigmentation were seen in 39 of the 100 patients but only six of 100 controls. 27% of eyes were myopic, 22% were hyperopic, and 27% had astigmatism >0.75D. Of the non-retinal findings, 39 patients had angiofibromas of the eyelids, five had non-paralytic strabismus, and three had colobomas. CONCLUSION: Apart from the higher prevalence of flat retinal hamartomas, the findings of this study compare closely with previous large clinic based series of TSC patients. Refractive findings were similar to previous studies of a similarly aged non-TSC population. This is the first series to document the statistically significant association of punched out chorioretinal depigmentation with TSC and the authors believe that it should be looked for as an aid to diagnosis. PMID- 11264131 TI - Retinal dystrophies caused by mutations in RPE65: assessment of visual functions. AB - AIMS: To characterise the disease in patients with mutations in RPE65. METHODS: Individuals from two families were studied clinically. RESULTS: 13 and 20 year old compound heterozygote individuals from one family with R234X and 1121delA mutations showed nystagmus, macular dystrophy and low contrasted spots in the fundus. Some heterozygotes had macular drusen. A 40 year old compound heterozygote individual from another family with L22P and H68Y mutations had few bone spicule pigment deposits and macular atrophy. CONCLUSION: Compound heterozygote individuals had severe rod-cone dystrophies featuring few pigment deposits in the fundus, pigment epithelium atrophy, and early involvement of the macula, with variations in severity leading to the diagnosis of Leber's congenital amaurosis or retinitis pigmentosa. Macular drusen in heterozygotes carrying a null allele may reflect the decreased capacity in the RPE65 function. PMID- 11264132 TI - Spontaneous reversal of nystagmus in the dark. AB - AIM: To report five children with horizontal jerk nystagmus in whom eye movement recordings in the dark revealed a spontaneous reversal in the direction of the nystagmus beat. Three patients were blind in one eye and were diagnosed as having a manifest latent nystagmus (MLN), and two patients had strabismus and congenital nystagmus (CN). METHODS: Eye movements were recorded using DC electro-oculography with simultaneous video recording, including infrared recording in total darkness. RESULTS: Four patients had decelerating velocity slow phase jerk nystagmus when recorded under natural lighting conditions; the fifth case had accelerating velocity and linear slow phase jerk nystagmus. Under absolute darkness, nystagmus reversed in direction of beat with a mixture of linear and decelerating velocity slow phase waveforms. One child with unilateral anophthalmos could wilfully reverse the beat direction of his nystagmus by trying to look with his blind eye in the light and dark. CONCLUSIONS: These observations support the theory that LN/MLN beat direction is determined by the "presumed" viewing eye and may be consciously controlled. The spontaneous reversal of beat direction in the dark suggests eye dominance is predetermined. Eye movement recordings identified mixed nystagmus waveforms indicating that CN (accelerating velocity slow phases) and LN/MLN (linear/decelerating velocity slow phases) coexist in these subjects. PMID- 11264133 TI - The development of a "reduced logMAR" visual acuity chart for use in routine clinical practice. AB - BACKGROUND/AIMS: The advantages of logMAR acuity data over the Snellen fraction are well known, and yet existing logMAR charts have not been adopted into routine ophthalmic clinical use. As this may be due in part to the time required for a logMAR measurement, this study was performed to determine whether an abbreviated logMAR chart design could combine the advantages of existing charts with a clinically acceptable measurement time. METHODS: The test-retest variability, agreement (with the gold standard), and time taken for "single letter" (interpolated) acuity measurements taken using three prototype "reduced logMAR" (RLM) charts and the Snellen chart were compared with those of the ETDRS chart which acted as the gold standard. The Snellen chart was also scored with the more familiar "line assignment" method. The subjects undergoing these measurements were drawn from a typical clinical outpatient population exhibiting a range of acuities. RESULTS: The RLM A prototype chart achieved a test-retest variability of +/-0.24 logMAR compared with +/-0.18 for the ETDRS chart. Test-retest variability for the Snellen chart was +/-0.24 logMAR using clinically prohibitive "single letter" scoring increasing to +/-0.33 with the more usual "line assignment" method. All charts produced acuity data which agreed well with those of the ETDRS chart. "Single letter" acuity measurements using the prototype RLM charts were completed in approximately half the time of those taken using the ETDRS and Snellen charts. The duration of a Snellen "line assignment" measurement was not evaluated. CONCLUSION: The RLM A chart offers an acceptable level of test retest variability when compared with the gold standard ETDRS chart, while reducing the measurement time by half. Also, by allowing a faster, less variable acuity measurement than the Snellen chart, the RLM A chart can bring the benefits of logMAR acuity to routine clinical practice. PMID- 11264134 TI - The specific architecture of the anterior stroma accounts for maintenance of corneal curvature. AB - AIM: To analyse the human corneal stroma in extreme hydration to discover if its structure is responsible for corneal stability. METHODS: Corneas in several hydration states were used: postmortem control corneas (PM; n=3), corneas left for 1 day in phosphate buffered saline (PBS; n=4), and corneas left for 1 day (n=4), 2 days (n=4), 3 days (n=2), and 4 days (n=4) in deionised water. All corneas were fixed under standardised conditions and processed for light and electron microscopy. In addition, two fresh corneas from the operating theatre were studied which were processed 6 months after storage in sodium cacodylate buffer. RESULTS: After 1 day in deionised water maximal stromal swelling was reached which did not change up to 4 days. The stroma of deionised water corneas (1400 microm) was much thicker than that of PBS corneas (650 microm) and PM corneas (450 microm). Deionised water treatment led to disappearance of all keratocytes leaving only remnants of nuclei and large interlamellar spaces. In these specimens the distance between the collagen fibres had increased significantly, but the diameter of the collagen fibres did not seem to be affected. A remarkable observation was that the most anterior part of the stroma (100-120 microm) in all deionised water specimens and those stored for 6 months in buffer was not swollen, indicating that the tightly interwoven anterior lamellae are resistant to extreme non-physiological hydration states. CONCLUSIONS: The rigidity of the most anterior part of the corneal stroma in extreme hydration states points to an important role in maintenance of corneal curvature. Since a large part of this rigid anterior part of the stroma is either removed (PRK) or intersected (LASIK), it is possible that in the long run patients who underwent refractive surgery may be confronted with optical problems. PMID- 11264135 TI - Suppression of interleukin 1alpha and interleukin 1beta in human limbal epithelial cells cultured on the amniotic membrane stromal matrix. AB - AIMS: Amniotic membrane (AM) transplantation reduces inflammation in a variety of ocular surface disorders. The aim of this study was to determine if AM stroma suppresses the expression of the IL-1 gene family in cultured human corneal limbal epithelial cells. METHODS: Human corneal limbal epithelial cells were cultured from limbocorneal explants of donor eyes on plastic or on the AM stroma. Transcript expression of IL-1alpha, IL-1beta, IL-1 receptor antagonist (RA), and GAPDH was compared with or without addition of lipopolysaccharide to their serum free media for 24 hours using RNAse protection assay (RPA). Their protein production in the supernatant was analysed by ELISA. RESULTS: Expression of IL 1alpha and IL-1beta transcripts and proteins was significantly reduced by cells cultured on the AM stromal matrix compared with plastic cultures whether lipopolysaccharide was added or not. Moreover, expression of IL-1 RA by cells cultured in the lipopolysaccharide-free medium was upregulated by AM stromal matrix. The ratio between IL-1 RA and IL-1alpha protein levels in AM cultures was higher than in plastic cultures. CONCLUSIONS: AM stromal matrix markedly suppresses lipopolysaccharide induced upregulation of both IL-1alpha and IL 1beta. These data may explain in part the effect of AM transplantation in reducing ocular surface inflammation, underscoring the unique feature of the AM as a substrate for tissue engineering. PMID- 11264136 TI - A simple corneal perfusion chamber for drug penetration and toxicity studies. AB - AIMS: Corneal perfusion chambers are important tools in the development and assessment of ophthalmic drugs. The aim of this study was to design and test a modified perfusion chamber suitable for topical application of drugs to isolated corneoscleral preparations, and which allowed continuous monitoring of endothelial cell function. METHODS: A polycarbonate and stainless steel perfusion chamber was designed to clamp corneas in a horizontal plane suitable for topical drug delivery. Endothelial cell function was assessed by ultrasonic pachymetry and specular microscopy during perfusion. Epithelial barrier function was assessed by penetration of fluorescein. Leakage was examined by measuring penetration of a large protein, IgG. Tissue architecture after perfusion was examined by conventional histology. RESULTS: Corneas maintained a functionally and morphologically intact endothelial monolayer during perfusion periods of up to 14 hours. The epithelial barrier function was well preserved. The tissue clamp sealed the preparation effectively against leakage of macromolecules. CONCLUSION: The new chamber device forms a reliable tool for in vitro drug penetration and toxicity studies in isolated perfused corneoscleral tissue. PMID- 11264137 TI - Clinicopathological correlation of retinal pigment epithelial tears in exudative age related macular degeneration: pretear, tear, and scarred tear. AB - AIMS: To analyse the histopathology of vascularised pigment epithelial detachments and tears of the retinal pigment epithelium (RPE) in age related macular degeneration (AMD). METHODS: The light microscopic architecture of 10 surgically removed subretinal specimens-three vascularised pigment epithelial detachments, four recent tears, and three scarred tears as a manifestation of AMD were studied and correlated with the angiographic findings. RESULTS: Recent tears: a large fibrovascular membrane was found to be originally situated in Bruch's membrane. About half of the surface of the fibrovascular tissue was denuded of RPE and diffuse drusen. The RPE and diffuse drusen had retracted and rolled up, covering a neighbouring part of the intra-Bruch's fibrovascular membrane. The rolled up RPE and diffuse drusen were not interspersed with fibrovascular tissue but lay superficial to the intra-Bruch's fibrovascular membrane itself. Scarred tears: a collagen capsule surrounded the rolled up diffuse drusen and RPE. Fibrovascular tissue was found inside the rolled up material, predominantly at its choroidal side. CONCLUSION: The area of choroidal neovascularisation associated with a vascularised pigment epithelial detachment and a tear of the RPE may be larger than was hitherto thought or indicated by fluorescein angiography. This neovascular tissue may be present within the bed of the RPE tear, as well as at the site of the scrolled up RPE. PMID- 11264139 TI - Suspension of anticytomegalovirus maintenance therapy following immune recovery due to highly active antiretroviral therapy. AB - AIM: To describe the authors' experience with discontinuation of anti cytomegalovirus (CMV) maintenance therapy in patients showing immune recovery following highly active antiretroviral therapy (HAART). METHODS: Retrospective analysis of the records of 41 patients who presented with CMV retinitis and whose maintenance therapy was discontinued from March 1997 to December 1999. RESULTS: 41 patients had their anti-CMV therapy discontinued. The mean follow up after discontinuation of maintenance therapy in April 2000 was 20.4 months. At the time of discontinuation of maintenance therapy the lowest CD4+ count was 143 cells x 10(6)/l and only three patients had detectable HIV viral load. No reactivation or progression was seen in any of these patients after suspension of maintenance therapy. CONCLUSION: The anti-CMV maintenance therapy could be discontinued safely in patients with CD4+ above 150 cells x 10(6)/l although close follow up remains necessary especially in patients whose CD4+ count drops below this level. PMID- 11264138 TI - Intravitreal invading cells contribute to vitreal cytokine milieu in proliferative vitreoretinopathy. AB - AIM: To examine the contribution of infiltrating cells in the local production of cytokines within the vitreous of patients with proliferative vitreoretinopathy (PVR). METHODS: The presence of mRNA coding for IL-6, IL-8, IL-1beta, IL-1alpha, TNFalpha, IFNgamma, IL-12, and HPRT was investigated in 25 vitreous samples from patients with PVR, 11 vitreous samples from patients with retinal detachment (RD) not complicated by PVR, and 10 vitreous samples from patients with macular hole (MH). A quantitative reverse transcriptase polymerase chain reaction (RT-PCR) using an internal competitor was used to investigate these samples. From these samples, 15 PVR, 8 RD, and 8 MH were analysed for the protein levels of the same cytokines using enzyme linked immunosorbent assay (ELISA). Spearman correlation was used to test any association between mRNA and cytokine protein levels, as an indicator of the contribution these cells make to the intravitreal cytokine milieu. RESULTS: A strong correlation was found between mRNA and their respective cytokine levels (protein products) for IL-6, IL-8, IL-1beta, IL-1alpha, TNFalpha, IFNgamma (Spearman r = 0.83, 0.73, 0.67, 0.91, 0.73, and 0.73 respectively), but not for IL-12. The median levels of IL-6, IL-8, IL-1beta, and IFNgamma mRNA and their respective cytokines were significantly higher (p <0.05) in patients with PVR than in those with macular hole. There was no statistically significant difference in the median levels of IL-1alpha mRNA between PVR and MH but the cytokine IL-1alpha was detected at a significantly higher level in PVR compared with MH patients. Between PVR and RD patients, there was no statistically significant difference in mRNA levels for all the investigated cytokines (p >0.05) except for IL-6 where there was a statistical significance (p= 0.038). In contrast, the median levels of IL-6, IL-8, and IL-1beta cytokines were significantly higher (p <0.05) in patients with PVR than in those with RD, whereas for IL-1alpha and IFNgamma no significant statistical difference was detected between PVR and RD patients (p >0.05). When results of RD and MH patients were compared, a statistical difference was only detected in mRNA levels of INFgamma (p = 0.008). However, no difference was detected for INFgamma (protein product) or for any of the other cytokines between RD and MH patients. CONCLUSION: Levels of both protein and mRNA encoding IL-6, IL-8, IL-1beta, and IFNgamma is significantly increased in vitreous samples from patients with PVR. The strong correlation between ELISA detectable cytokines (protein products) and their respective mRNA levels suggest that intravitreal, invasive cells are the major source of these cytokines, with the exception of IL-12. Cells invading the vitreous do not appear to locally produce IL-12 mRNA. This would appear to implicate cells peripheral to the vitreal mass as the major source of this cytokine. PMID- 11264140 TI - Cyclodiode laser therapy for painful, blind glaucomatous eyes. AB - AIMS: To determine the ability of cyclodiode laser treatment to relieve discomfort in painful blind glaucomatous eyes. METHODS: 30 eyes underwent cyclodiode to reduce intraocular pressure (IOP) and relieve pain. Patients graded their pre-cyclodiode and post-cyclodiode pain. RESULTS: After a minimum follow up of 6 months, a single cyclodiode treatment lowered mean IOP from 51 mm Hg (95% CI plus or minus 3.7 mm Hg) to 26 mm Hg (95% CI plus or minus 5.8 mm Hg) providing pain relief in 73.3% (22/30). After retreatment of six eyes, mean IOP was reduced to 22 (95% CI plus or minus 5.3) mm Hg and pain relief was obtained in 96.7% (29/30). For eyes achieving pain relief after one treatment, IOP was reduced by >30% in 81.0% (17/21). For eyes not achieving pain relief after one treatment, IOP was reduced by >30% in only 22.2% (2/9) (p=0.0042, Fisher's exact test). CONCLUSION: Cyclodiode was highly successful in providing pain relief in painful blind hypertensive glaucomatous eyes. The best predictor of successful pain relief was IOP reduction of > 30% from baseline. PMID- 11264141 TI - A comparative study of proxymetacaine-fluorescein and lignocaine-fluorescein use during applanation tonometry. AB - AIMS: To evaluate the relative merits of proxymetacaine-fluorescein (PROX-FLU) and lignocaine-fluorescein (LIG-FLU) when used to perform applanation tonometry. METHODS: This prospective, masked, double blind study assessed several aspects of the tonometry process-the duration of the stinging sensation and degree of discomfort, the extent of reflex lacrimation induced, the need for subsequent tear film manipulation to ensure an accurate tonometry reading, and total time to complete tonometry-for each preparation. RESULTS: PROX-FLU caused significantly less discomfort and reflex lacrimation than LIG-FLU and accurate tonometry was more rapidly completed when it was used. PROX-FLU was preferred by 98% of the study patients. CONCLUSION: PROX-FLU is a well tolerated and useful alternative to the more widely used LIG-FLU mixture. PMID- 11264142 TI - Management of inferior retinal breaks during pars plana vitrectomy for retinal detachment. AB - AIMS: To determine whether it is necessary to support inferior retinal breaks with a scleral explant during pars plana vitrectomy (PPV) for rhegmatogenous retinal detachment (RD). METHODS: A prospective study was carried out on nine eyes of nine consecutive patients undergoing PPV for primary RD with associated inferior retinal breaks and no significant proliferative vitreoretinopathy. RESULTS: Eight eyes were successfully reattached with a single operation. No cases presented with redetachment because of failed closure of the original inferior breaks. CONCLUSIONS: It is not necessary to support inferior retinal breaks with a scleral explant during PPV for primary RD repair in selected cases. PMID- 11264144 TI - Acute posterior vitreous detachment. PMID- 11264143 TI - Treatment of subfoveal choroidal neovascularisation in age related macular degeneration: focus on clinical application of verteporfin photodynamic therapy. PMID- 11264146 TI - Plasma glycolipids levels: new factors regulating the protein C anticoagulant pathway and determining thrombotic risk. PMID- 11264150 TI - Plasma glucosylceramide deficiency as potential risk factor for venous thrombosis and modulator of anticoagulant protein C pathway. AB - To assess the relationship between venous thrombosis and plasma glucosylceramide (GlcCer) or phosphatidylethanolamine (PE), plasma levels of GlcCer and PE were determined for 70 venous thrombosis patients referred for evaluation and 70 healthy blood donors. The mean GlcCer level, but not the PE level, was lower in patients versus controls (4.9 vs 6.5 microg/mL [P =.0007] and 66 vs 71 microg/mL [P =.48], respectively). As a measure of relative risk, the odds ratio for deep vein thrombosis in subjects with GlcCer levels below the 10th percentile of controls was 5.7 (95% CI, 2.3-14). To assess the influence of glycolipids on anticoagulant response to activated protein C (APC):protein S in modified prothrombin time assays, the effects of depleting endogenous plasma GlcCer by glucocerebrosidase treatment or of adding exogenous purified GlcCer or other neutral glycolipids to plasma were tested. Glucocerebrosidase treatment reduced plasma sensitivity to APC:protein S in parallel with GlcCer reduction. Exogenously added GlcCer and the homologous Glc-containing globotriaosylceramide (Gb3Cer), but not galactosylceramide, dose-dependently prolonged clotting times of normal plasma in the presence, but not absence, of APC:protein S, which suggests that GlcCer or Gb3Cer can enhance protein C pathway anticoagulant activity. In studies using purified proteins, inactivation of factor Va by APC:protein S was enhanced by GlcCer alone and by GlcCer in multicomponent vesicles containing phosphatidylserine and phosphatidylcholine. These results suggest that the neutral glycolipids GlcCer and Gb3Cer may directly contribute to the anticoagulant activity of the protein C pathway and that deficiency of plasma GlcCer may be a risk factor for venous thrombosis. (Blood. 2001;97:1907-1914) PMID- 11264151 TI - How I treat patients with von Willebrand disease. AB - Von Willebrand disease (vWD) is a frequent inherited disorder of hemostasis that affects both sexes. Two abnormalities are characteristic of the disease, which is caused by a deficiency or a defect in the multimeric glycoprotein called von Willebrand factor: low platelet adhesion to injured blood vessels and defective intrinsic coagulation owing to low plasma levels of factor VIII. There are 2 main options available for the treatment of spontaneous bleeding episodes and for bleeding prophylaxis: desmopressin and transfusional therapy with plasma products. Desmopressin is the treatment of choice for most patients with type 1 vWD, who account for approximately 70% to 80% of cases. This nontransfusional hemostatic agent raises endogenous factor VIII and von Willebrand factor 3 to 5 times and thereby corrects both the intrinsic coagulation and the primary hemostasis defects. In patients with the more severe type 3 and in most patients with type 2 disease, desmopressin is ineffective or is contraindicated and it is usually necessary to resort to plasma concentrates containing both factor VIII and von Willebrand factor. Concentrates treated with virucidal methods should be preferred to cryoprecipitate because they are equally effective and are perceived as safer. (Blood. 2001;97:1915-1919) PMID- 11264152 TI - Eotaxin is a natural antagonist for CCR2 and an agonist for CCR5. AB - Eotaxin is a potent inducer of eosinophil chemotaxis and was considered as a selective ligand of the CC chemokine receptor 3 (CCR3), which is expressed on eosinophils, basophils, and Th2 lymphocytes. This study shows that eotaxin also interacts with CCR2 and CCR5 and can, thus, affect the responses of monocytes, which express both receptors. In human monocytes pretreatment with eotaxin decreased responsiveness to MCP-1, a selective ligand for CCR2, as well as to RANTES and MIP-1 beta, which bind to CCR5. Similar effects were obtained with transfected cells expressing CCR2 or CCR5, but here a difference became apparent: Eotaxin triggered CCR5 at a concentration of 100 nM but not CCR2 even at 1 microM, suggesting an antagonistic effect on this receptor. In agreement with this observation, eotaxin induced internalization of CCR5 but not of CCR2 in human monocytes and transfected cells. Binding studies showed that eotaxin displaces (125) I-MCP-1 from monocytes in a concentration-dependent manner, and functional experiments showed that eotaxin inhibits MCP-1-induced chemotaxis and enzyme release. The results demonstrate that eotaxin is a CCR5 agonist and a CCR2 antagonist. The present findings suggest a role of eotaxin in the fine-tuning of cellular responses occurring at sites of allergic inflammation, in which both MCP 1 and eotaxin are produced. (Blood. 2001;97:1920-1924) PMID- 11264153 TI - Polymorphism in the fractalkine receptor CX3CR1 as a genetic risk factor for coronary artery disease. AB - Coronary atherosclerosis is a major cause of death in industrialized countries. Monocytes, which play a key role in atherosclerosis, migrate into the vessel wall, presumably guided by specific chemoattractant and adhesion molecules. A compelling candidate for this role is the chemokine receptor CX3CR1, which is expressed on monocytes and acts as either a chemotactic receptor or an adhesion molecule, depending on whether its ligand, fractalkine, is presented free or membrane bound. A common variant of CX3CR1 was recently identified, encoded by the alleles I249 and M280, which form a common I(249)M(280) haplotype. When CX3CR1 genotypes were analyzed in 151 patients with acute coronary syndromes and in 249 healthy controls, CX3CR1 I249 heterozygosity was associated with a markedly reduced risk of acute coronary events, independent of established acquired coronary risk factors (eg, smoking, diabetes). The adjusted odds ratio for this allele was 0.43 (95% confidence interval, 0.26-0.72; P =.001). Consistent with this, functional analysis of peripheral blood mononuclear cells showed that CX3CR1 I249 heterozygosity was associated with a significant decrease in the number of fractalkine binding sites per cell. The results show that CX3CR1 I249 is an independent genetic risk factor for coronary artery disease and that CX3CR1 may be involved in the pathogenesis of atherosclerotic disease. (Blood. 2001;97:1925-1928) PMID- 11264154 TI - Clonotypic polymerase chain reaction confirms minimal residual disease in CLL nodular PR: results from a sequential treatment CLL protocol. AB - Patient-tumor-specific oligonucleotides were generated for the detection of minimal residual disease (MRD) in a highly specific and sensitive clonotypic polymerase chain reaction (cPCR). The clone-specific region of highest diversity, CDR-III, was PCR amplified and sequenced. Nested CDR-III clonotypic primers were used in a semi-nested cPCR with a sensitivity of at least 1 in 10(5) cells. Patients with protocol-eligible Rai intermediate or high-risk chronic lymphocytic leukemia (CLL) received induction with fludarabine 25 mg/m(2) per day for 5 days every 4 weeks for 6 cycles, followed by consolidative high-dose cyclophosphamide (1.5, 2.25, or 3g/m(2)). cPCR was performed on peripheral blood and bone marrow mononuclear cells. All 5 patients achieving a clinical partial remission (PR) studied by cPCR were positive. Five patients achieved nodular PR (nPR) (residual nodules or suspicious lymphocytic infiltrates in a bone marrow biopsy as the sole suggestion of residual disease). Five of 5 patients with nPR were cPCR positive. In contrast, flow cytometry for CD5-CD19 dual staining and kappa--lambda clonal excess detected MRD in only 3 of the same 5 nPR patients, all of whom were cPCR positive, and immunohistochemistry detected MRD in only 1 of 4 assessable patients. Three of 7 CR patients evaluable by cPCR had MRD. Only 1 CR patient had MRD by flow cytometry; that patient was also cPCR positive. These data support the conclusions that nodular PR in CLL represents MRD and that clonotypic PCR detects MRD in CLL more frequently than flow cytometry or immunohistochemistry. (Blood. 2001;97:1929-1936) PMID- 11264156 TI - A prospective randomized phase II trial of GM-CSF priming to prevent topotecan induced neutropenia in chemotherapy-naive patients with malignant melanoma or renal cell carcinoma. AB - We conducted a phase II randomized trial of recombinant granculocyte-macrophage colony-stimulating factor (GM-CSF) administered before topotecan chemotherapy to determine whether it could prevent myelosuppression and to determine the antitumor activity of this topoisomerase I inhibitor in 53 patients with metastatic malignant melanoma and renal cell cancer. All patients received GM-CSF after topotecan at a dose of 250 microg/m(2) daily for at least 8 days. Patients randomly assigned to receive GM-CSF priming were treated with GM-CSF at 250 microg/m(2) twice daily for 5 days before treatment. Twenty-five patients were randomly assigned to receive GM-CSF priming and 28 to receive topotecan without priming. The primary analysis was restricted to the protective effects seen during the first cycle of therapy. Grade 4 neutropenia occurred in 8 of 23 patients (35%) and grade 3 neutropenia in 5 of 23 patients (22%) randomized to GM CSF priming, whereas 18 of 26 (69%) and 5 of 26 (19%) patients experienced grade 4 or 3 neutropenia, respectively, without GM-CSF priming (P =.0074). The mean duration of neutropenia was reduced by GM-CSF priming: grade 3 neutropenia from 5.2 +/- 0.7 to 2.8 +/- 0.7 days (P =.0232) and grade 4 neutropenia from 2.7 +/- 0.6 to 1.1 +/- 0.4 days (P = 0.0332). The protective effects of GM-CSF extended to the second cycle of treatment. The incidence of febrile neutropenia was also reduced. Chemotherapy-induced anemia and thrombocytopenia were similar in both groups. One partial response was seen in a patient with melanoma, and one patient with renal cell cancer had complete regression of pulmonary metastases and was rendered disease-free by nephrectomy. (Blood. 2001;97:1942-1946) PMID- 11264155 TI - Modulation of endothelial cell activation in sickle cell disease: a pilot study. AB - The vessel wall endothelium undoubtedly plays a role in the vascular pathobiology of sickle cell disease. This pilot study tested the feasibility of using an inhibitor of nuclear factor (NF)-kappa B, a transcription factor, to modify the endothelial activation state of patients with this vascular disease. For a total of 7 separate drug exposure tests, 3 subjects with sickle cell disease took sulfasalazine (given orally at 1 g every 8 hours), and the activation state of their circulating endothelial cells (CECs) was assessed using immunofluorescence microscopy. Companion studies were also performed using sulfasalazine in sickle transgenic mice to verify its effect simultaneously on both CECs and vessel wall endothelium. Both CECs and tissue vessel wall endothelium in sickle mice have an activated phenotype. In these mice sulfasalazine significantly reduced CEC expression of vascular cell adhesion molecule (VCAM), intracellular adhesion molecule (ICAM), and E-selectin, and it correspondingly reduced expression of these molecules in some tissue vessels. In humans with sickle cell disease, sulfasalazine significantly reduced CEC expression of VCAM, ICAM, and E-selectin, but it did not reduce expression of tissue factor. Addition of a second transcription factor inhibitor, salsalate, did not change this result. This pilot study suggests that endothelial cell activation state can be modified and down regulated in vivo by sulfasalazine. (Blood. 2001;97:1937-1941) PMID- 11264157 TI - Bone marrow transplantation corrects osteopetrosis in the carbonic anhydrase II deficiency syndrome. AB - Carbonic anhydrase II (CAII), found in renal tubules, brain, and osteoclasts, is critical in acid-base homeostasis and bone remodeling. Deficiency of CAII gives rise to a syndrome of osteopetrosis, renal tubular acidosis (RTA), and cerebral calcification with associated developmental delay. It is inherited in an autosomal recessive fashion and found most frequently in the Mediterranean region and the Middle East. We report 2 related Irish families with clinically severe CAII deficiency in whom the gene mutation has been fully elucidated. Two children, one from each family, have undergone allogeneic bone marrow transplantation because of severe progressive visual and hearing loss. The older 2 children had already developed cerebral calcification and marked visual loss at the time of diagnosis and were treated symptomatically. Post-transplantation evaluation at 2 and 3 years demonstrates histologic and radiologic resolution of their osteopetrosis with stabilization of hearing and vision. Both children remain developmentally delayed and continue to have RTA, and the older child has now developed cerebral calcification. Allogeneic bone marrow stem cell replacement cures the osteoclast component of CAII deficiency and retards the development of cerebral calcification, but it appears to have little or no effect on the renal lesions. (Blood. 2001;97:1947-1950) PMID- 11264158 TI - Induction of cytotoxic T lymphocyte and antibody responses to enhanced green fluorescent protein following transplantation of transduced CD34(+) hematopoietic cells. AB - Genetic modification of hematopoietic stem cells often results in the expression of foreign proteins in pluripotent progenitor cells and their progeny. However, the potential for products of foreign genes introduced into hematopoietic stem cells to induce host immune responses is not well understood. Gene marking and induction of immune responses to enhanced green fluorescent protein (eGFP) were examined in rhesus macaques that underwent nonmyeloablative irradiation followed by infusions of CD34(+) bone marrow cells transduced with a retroviral vector expressing eGFP. CD34(+) cells were obtained from untreated animals or from animals treated with recombinant human granulocyte colony-stimulating factor (G CSF) alone or G-CSF and recombinant human stem cell factor. Levels of eGFP expressing cells detected by flow cytometry peaked at 0.1% to 0.5% of all leukocytes 1 to 4 weeks after transplantation. Proviral DNA was detected in 0% to 17% of bone marrow--derived colony-forming units at periods of 5 to 18 weeks after transplantation. However, 5 of 6 animals studied demonstrated a vigorous eGFP-specific cytotoxic T lymphocyte (CTL) response that was associated with a loss of genetically modified cells in peripheral blood, as demonstrated by both flow cytometry and polymerase chain reaction. The eGFP-specific CTL responses were MHC-restricted, mediated by CD8(+) lymphocytes, and directed against multiple epitopes. eGFP-specific CTLs were able to efficiently lyse autologous CD34(+) cells expressing eGFP. Antibody responses to eGFP were detected in 3 of 6 animals. These data document the potential for foreign proteins expressed in CD34(+) hematopoietic cells and their progeny to induce antibody and CTL responses in the setting of a clinically applicable transplantation protocol. (Blood. 2001;97:1951-1959) PMID- 11264159 TI - Human homologues of Delta-1 and Delta-4 function as mitogenic regulators of primitive human hematopoietic cells. AB - Delta-mediated Notch signaling controls cell fate decisions during invertebrate and murine development. However, in the human, functional roles for Delta have yet to be described. This study reports the characterization of Delta-1 and Delta 4 in the human. Human Delta-4 was found to be expressed in a wide range of adult and fetal tissues, including sites of hematopoiesis. Subsets of immature hematopoietic cells, along with stromal and endothelial cells that support hematopoiesis, were shown to express Notch and both Delta-1 and Delta-4. Soluble forms of human Delta-1 (h Delta-1) and h Delta-4 proteins were able to augment the proliferation of primitive human hematopoietic progenitors in vitro. Intravenous transplantation of treated cultures into immune-deficient mice revealed that h Delta-1 is capable of expanding pluripotent human hematopoietic repopulating cells detected in vivo. This study provides the first evidence for a role of Delta ligands as a mitogenic regulator of primitive hematopoietic cells in the human. (Blood. 2001;97:1960-1967) PMID- 11264160 TI - Erythroblasts are a source of angiogenic factors. AB - In adult bone marrow, mature erythroblasts are produced within structures called erythroblastic islands and then cross the endothelial barrier to reach circulation. Erythroblastic islands are composed of a central macrophage surrounded by maturing erythroblasts. In this study, it is shown that erythroid cells, but not the other mature hematopoietic cells, coexpress 2 angiogenic factors, vascular endothelial growth factor A (VEGF-A) and placenta growth factor (PlGF). Secretion of both VEGF-A and PlGF increases during in vitro erythroid differentiation. Erythroblast-conditioned medium can induce both migration of monocytes and endothelial cells and the permeability of endothelial cells. These effects are inhibited by anti-PlGF and/or anti-VEGF antibodies. Finally, it is shown that VEGF-A and PlGF proteins are expressed by bone marrow erythroblasts in vivo. Angiogenic factors secreted by erythroblasts may promote interactions either with macrophages in erythroblastic islands or with endothelial cells that would facilitate the passage of erythroid cells through the endothelial barrier. (Blood. 2001;97:1968-1974) PMID- 11264161 TI - Cross-talk between alpha(4)beta(1)/alpha(5)beta(1) and c-Kit results in opposing effect on growth and survival of hematopoietic cells via the activation of focal adhesion kinase, mitogen-activated protein kinase, and Akt signaling pathways. AB - Erythroid progenitor cells (EPCs) are deficient in mice lacking either the ligand stem cell factor (SCF), its receptor c-Kit, or beta(1)-integrins. In nonhematopoietic cells, integrins and receptor tyrosine kinases can collaborate to modulate cellular functions, providing evidence for cross-talk between signals emerging from these cell surface molecules. Using specific recombinant fibronectin peptides that contain the binding site for the integrin alpha(4)beta(1) (FN-H296) or alpha(5)beta(1) (FN-CH271) or both alpha(4)beta(1) and alpha(5)beta(1) (FN-CH296), this study investigated the effect of adhesion alone, or in combination with activation of c-Kit, on functional and biochemical outcomes in an EPC line, G1E-ER2, and primary EPCs. G1E-ER2 cells and primary EPCs cultured on FN-CH271 in the presence of c-Kit activation led to a significant increase in proliferation in comparison with cells grown on FN-H296 or FN-CH296. G1E-ER2 cells cultured on FN-H296 or FN-CH296 resulted in significant cell death in comparison to cells grown on FN-CH271. Activation of c Kit enhanced the survival of G1E-ER2 cells grown on FN-H296 or FN-CH296; however, the rescue was only partial. The reduced survival of G1E-ER2 cells on FN-H296 correlated with reduced activation of Akt and expression of Bcl-2 and Bcl-x(L), whereas increase in proliferation on FN-CH271 correlated with significantly enhanced and sustained activation of focal adhesion kinase (FAK) and extracellular-regulated kinase (ERK) pathways. These data demonstrate that adhesion-induced signals emanating from ligation of alpha(4)beta(1) and alpha(5)beta(1) result in distinct biologic outcomes, including death via alpha(4)beta(1) and survival/proliferation via alpha(5)beta(1). (Blood. 2001;97:1975-1981) PMID- 11264162 TI - CD9 and megakaryocyte differentiation. AB - It is shown that the tetraspanin CD9 has a complex pattern of distribution in hematopoietic cells and is heterogeneously expressed on human bone marrow CD34(+) cells. CD34(high)CD38(low)Thy1(+) primitive progenitors are contained in the population with intermediate CD9 expression, thus suggesting that CD9 expression may precede CD38 appearance. Cell sorting shows that colony-forming unit (CFU) GEMM and CFU-GM are present in high proportions in this fraction and in the fraction with the lowest CD9 expression. Cells with the highest level of CD9 are committed to the B-lymphoid or megakaryocytic (MK) lineages, as shown by the co expression of either CD19 or CD41/GPIIb and by their strong potential to give rise to CFU-MK. In liquid cultures, CD9(high)CD41(neg) cells give rise to cells with high CD41 expression as early as 2 days, and this was delayed by at least 3 to 4 days for the CD9(mid) cells; few CD41(high) cells could be detected in the CD9(low) cell culture, even after 6 days. Antibody ligation of cell surface CD9 increased the number of human CFU-MK progenitors and reduced the production of CD41(+) megakaryocytic cells in liquid culture. This was associated with a decreased expression of MK differentiation antigens and with an alteration of the membrane structure of MK cells. Altogether these data show a precise regulation of CD9 during hematopoiesis and suggest a role for this molecule in megakaryocytic differentiation, possibly by participation in membrane remodeling. (Blood. 2001;97:1982-1989) PMID- 11264163 TI - Basis of hematopoietic defects in platelet-derived growth factor (PDGF)-B and PDGF beta-receptor null mice. AB - Platelet-derived growth factor (PDGF)-B and PDGF beta-receptor (PDGFR beta) deficiency in mice is embryonic lethal and results in cardiovascular, renal, placental, and hematologic disorders. The hematologic disorders are described, and a correlation with hepatic hypocellularity is demonstrated. To explore possible causes, the colony-forming activity of fetal liver cells in vitro was assessed, and hematopoietic chimeras were demonstrated by the transplantation of mutant fetal liver cells into lethally irradiated recipients. It was found that mutant colony formation is equivalent to that of wild-type controls. Hematopoietic chimeras reconstituted with PDGF-B(-/-), PDGFR beta(-/-), or wild type fetal liver cells show complete engraftment (greater than 98%) with donor granulocytes, monocytes, B cells, and T cells and display none of the cardiovascular or hematologic abnormalities seen in mutants. In mouse embryos, PDGF-B is expressed by vascular endothelial cells and megakaryocytes. After birth, expression is seen in macrophages and neurons. This study demonstrates that hematopoietic PDGF-B or PDGFR beta expression is not required for hematopoiesis or integrity of the cardiovascular system. It is argued that metabolic stress arising from mutant defects in the placenta, heart, or blood vessels may lead to impaired liver growth and decreased production of blood cells. The chimera models in this study will serve as valuable tools to test the role of PDGF in inflammatory and immune responses. (Blood. 2001;97:1990-1998) PMID- 11264164 TI - In vitro cytotoxic effects of a tyrosine kinase inhibitor STI571 in combination with commonly used antileukemic agents. AB - The BCR/ABL tyrosine kinase has been implicated in the pathogenesis of chronic myelogenous leukemia (CML) and Philadelphia chromosome-positive (Ph(+)) acute lymphoblastic leukemia (ALL). STI571 is a novel anticancer agent that selectively inhibits the BCR/ABL tyrosine kinase. The cytotoxic effects of STI571 were studied in combination with antileukemic agents against Ph(+) leukemia cell lines, KU812, K-562, TCC-S, and TCC-Y. The cells were exposed to STI571 and to other agents simultaneously for 5 or 7 days. Cell growth inhibition was determined by MTT assay. The cytotoxic effects in combinations at the inhibitory concentration of 80% level were evaluated by the isobologram. STI571 produced synergistic effects with recombinant and natural alpha-interferons in 2 of 3 and 3 of 3 cell lines, respectively. STI571 produced additive effects with hydroxyurea, cytarabine, homoharringtonine, doxorubicin, and etoposide in all 4 cell lines. STI571 with 4-hydroperoxy-cyclophosphamide, methotrexate, or vincristine produced additive, antagonistic, and synergistic effects in 3 of 4 cell lines, respectively. These findings suggest that the simultaneous administration of STI571 with other agents except methotrexate would be advantageous for cytotoxic effects against Ph(+) leukemias. Among them, the simultaneous administration of STI571 and alpha-interferons or vincristine would be highly effective against Ph(+) leukemias and these combinations would be worthy of clinical trials. In contrast, the simultaneous administration of STI571 with methotrexate would have little therapeutic efficacy. Although there are gaps between in vitro studies and clinical trials, the present findings provide useful information for the establishment of clinical protocols involving STI571. (Blood. 2001;97:1999-2007) PMID- 11264165 TI - Comparison of effects of the tyrosine kinase inhibitors AG957, AG490, and STI571 on BCR-ABL--expressing cells, demonstrating synergy between AG490 and STI571. AB - STI571 (formerly CGP57148) and AG957 are small molecule inhibitors of the protein tyrosine kinase (PTK) p145(abl) and its oncogenic derivative p210(bcr-abl). AG490 is an inhibitor of the PTK Janus kinase 2 (JAK2). No direct comparison of these inhibitors has previously been reported, so this study compared their effects on factor-dependent FDC-P1, 32D, and MO7e cells and their p210(bcr-abl)-expressing factor-independent derivatives. STI571 was a more potent inhibitor of (3)H thymidine incorporation in p210(bcr-abl)-expressing cells than was AG957, and it showed superior discrimination between inhibitory effects on parental cell lines and effects on their p210(bcr-abl)-expressing derivatives. Assays performed with and without growth factor demonstrated that STI571 but not AG957 reversed the p210(bcr-abl)-driven factor independence of cell lines. p210(bcr-abl)-expressing cells were less sensitive to AG490 than to AG957 or STI571. However, for p210(bcr abl)-expressing clones from all 3 cell lines, synergistic inhibition was demonstrated between STI571 and concentrations of AG490 with no independent inhibitory effect. Inhibition of nucleic acid synthesis with AG957 treatment was associated with reduced cell numbers, reduced viability, and small pyknotic apoptotic cells. At concentrations of STI571 that reversed the p210(bcr-abl) factor-independent phenotype, STI571 treatment and growth factor deprivation together were sufficient to induce apoptosis. This study concludes that, for the cell lines studied, (1) STI571 is a more potent and more selective inhibitor of a p210(bcr-abl)-dependent phenotype than AG957; (2) AG490 synergizes with STI571 to enhance its inhibitory effect on p210(bcr-abl)-driven proliferation; and (3) the combination of p210(bcr-abl)-tyrosine kinase inhibition and growth factor signal withdrawal can be sufficient to induce apoptotic death of transformed cells. (Blood. 2001;97:2008-2015) PMID- 11264166 TI - Evidence for the presence of murine primitive megakaryocytopoiesis in the early yolk sac. AB - During mouse embryogenesis, primitive erythropoiesis occurs in blood islands of the yolk sac (YS) on the seventh day of gestation. This study demonstrated for the first time the presence of unique primitive megakaryocytic (Mk) progenitors in the early YS, which disappeared by 13.5 days postcoitum (dpc). When 7.5 dpc YS cells were incubated in the presence of stem cell factor (SCF), interleukin (IL) 3, IL-6, erythropoietin (EPO), thrombopoietin (TPO), and granulocyte colony stimulating factor in methylcellulose clonal culture, not only erythroid bursts but also megakaryocyte colonies were observed. The megakaryocytes in the colonies matured to proplatelet stages and produced platelets as early as day 3 of culture, much earlier than those from adult bone marrow, although their ploidy class was lower. These megakaryocytes were stained with acetylcholine esterase, and expressed platelet glycoprotein (GP)Ib beta, GPIIIa, and platelet factor 4 by reverse transcription-polymerase chain reaction analysis. The analysis of hemoglobin types in erythrocytes obtained from hematopoietic multilineage colonies containing the megakaryocytes indicated that the Mk progenitors originated from primitive hematopoiesis. The primitive Mk progenitors formed colonies in the absence of any cytokines in fetal bovine serum (FBS)-containing culture, and SCF, IL-3, EPO, and TPO significantly enhanced the Mk colony formation. In FBS-free culture, however, no colony formation was induced without these cytokines. Because megakaryocytes were detected in 8.5-dpc YS, these unique primitive Mk progenitors may rapidly mature and give rise to platelets to prevent hemorrhage in the simultaneously developing blood vessels until definitive hematopoiesis begins to produce platelets. (Blood. 2001;97:2016-2022) PMID- 11264167 TI - Different expression of CD41 on human lymphoid and myeloid progenitors from adults and neonates. AB - The glycoprotein (Gp) IIb/IIIa integrin, also called CD41, is the platelet receptor for fibrinogen and several other extracellular matrix molecules. Recent evidence suggests that its expression is much wider in the hematopoietic system than was previously thought. To investigate the precise expression of the CD41 antigen during megakaryocyte (MK) differentiation, CD34(+) cells from cord blood and mobilized blood cells from adults were grown for 6 days in the presence of stem cell factor and thrombopoietin. Two different pathways of differentiation were observed: one in the adult and one in the neonate cells. In the neonate samples, early MK differentiation proceeded from CD34(+)CD41(-) through a CD34( )CD41(+)CD42(-) stage of differentiation to more mature cells. In contrast, in the adult samples, CD41 and CD42 were co-expressed on a CD34(+) cell. The rare CD34(+)CD41(+)CD42(-) cell subset in neonates was not committed to MK differentiation but contained cells with all myeloid and lymphoid potentialities along with long-term culture initiating cells (LTC-ICs) and nonobese diabetic/severe combined immune-deficient repopulating cells. In the adult samples, the CD34(+)CD41(+)CD42(-) subset was enriched in MK progenitors, but also contained erythroid progenitors, rare myeloid progenitors, and some LTC-ICs. All together, these results demonstrate that the CD41 antigen is expressed at a low level on primitive hematopoietic cells with a myeloid and lymphoid potential and that its expression is ontogenically regulated, leading to marked differences in the surface antigenic properties of differentiating megakaryocytic cells from neonates and adults. (Blood. 2001;97:2023-2030) PMID- 11264168 TI - Signal transduction pathways involved in soluble fractalkine-induced monocytic cell adhesion. AB - Fractalkine displays features that distinguishes it from the other chemokines. In particular, besides its chemoattractant action it promotes, under physiologic flow, the rapid capture and the firm adhesion of a subset of leukocytes or intervenes in the neuron/microglia interaction. This study verified that indeed the human monocytic MonoMac6 cell line adheres to fibronectin-coated filters in response to soluble fractalkine (s-FKN). s-FKN stimulates, with distinct time courses, extracellular signal-related kinases (ERK1 and ERK2) and stress activated protein kinases (SAPK1/JNK1 and SAPK2/p38). Both p60 Src and p72 Syk were activated under s-FKN stimulation with a rapid kinetic profile compatible with a downstream regulation on the mitogen-activated protein kinase (MAPK) congeners. The use of specific tyrosine kinase inhibitors revealed that the ERK pathway is strictly controlled by Syk, whereas c-Src up-regulated the downstream SAPK2/p38. In contrast, the SAPK1/JNK1 pathway was not regulated by any of these nonreceptor tyrosine kinases. The s-FKN-mediated increased adherence of MonoMac6 cells was partially inhibited by SB202190, a broad SAPKs inhibitor, PD98059, an MEK inhibitor, LY294002, a phosphatidyl inositol 3-kinase inhibitor, and a pertussis toxin-sensitive G protein. These data highlight that the integration of a complex array of signal transduction pathways is necessary to complete the full s-FNK-dependent adherence of human monocytic cells to fibronectin. (Blood. 2001;97:2031-2037) PMID- 11264169 TI - Importance of leucine zipper domain of mi transcription factor (MITF) for differentiation of mast cells demonstrated using mi(ce)/mi(ce) mutant mice of which MITF lacks the zipper domain. AB - The mi transcription factor (MITF) is a basic helix-loop-helix leucine zipper (bHLH-Zip) transcription factor that is important for the development of mast cells. Mast cells of mi/mi genotype express normal amount of abnormal MITF (mi MITF), whereas mast cells of tg/tg genotype do not express any MITFs. Mast cells of mi/mi mice show more severe abnormalities than those of tg/tg mice, indicating that the mi-MITF possesses the inhibitory function. The MITF encoded by the mi(ce) mutant allele (ce-MITF) lacks the Zip domain. We examined the importance of the Zip domain using mi(ce)/mi(ce) mice. The amounts of c-kit, granzyme B (Gr B), and tryptophan hydroxylase (TPH) messenger RNAs decreased in mast cells of mi(ce)/mi(ce) mice to levels comparable to those of tg/tg mice, and the amounts were intermediate between those of +/+ mice and those of mi/mi mice. Gr B mediates the cytotoxic activity of mast cells, and TPH is a rate-limiting enzyme for the synthesis of serotonin. The cytotoxic activity and serotonin content of mi(ce)/mi(ce) mast cells were comparable to those of tg/tg mast cells and were significantly higher than those of mi/mi mast cells. The phenotype of mi(ce)/mi(ce) mast cells was similar to that of tg/tg mast cells rather than to that of mi/mi mast cells, suggesting that the ce-MITF had no functions. The Zip domain of MITF appeared to be important for the development of mast cells. (Blood. 2001;97:2038-2044) PMID- 11264170 TI - Human skin-derived mast cells can proliferate while retaining their characteristic functional and protease phenotypes. AB - Human mast cells in adult tissues have been thought to have limited, if any, proliferative potential. The current study examined mast cells obtained from adult skin and cultured in serum-free medium with recombinant human stem cell factor. During the first 4 weeks of culture, the percentages of mast cells increased from 10 to almost 100. After 8 weeks, a 150-fold increase in the number of mast cells was observed. When freshly dispersed mast cells were individually sorted onto human fibroblast monolayers and cultured for 3 weeks, one or more mast cells were detected in about two thirds of the wells, and in about two thirds of these wells the surviving mast cells showed evidence of proliferation, indicating most mast cells in skin can proliferate. Such mast cells all expressed high surface levels of Kit and Fc epsilon RI, each of which were functional, indicating IgE was not required for Fc epsilon RI expression on mast cells. Such mast cells also retained the MC(TC) protease phenotype of mast cells that normally reside in the dermis. After 4 to 8 weeks of culture these mast cells degranulated in response to substance P and compound 48/80, characteristics of skin-derived mast cells that persist outside of the cutaneous microenvironment. (Blood. 2001;97:2045-2052) PMID- 11264171 TI - Identification of polymorphisms in the promoter and the 3' region of the TAFI gene: evidence that plasma TAFI antigen levels are strongly genetically controlled. AB - Thrombin-activable fibrinolysis inhibitor (TAFI) is a recently described carboxypeptidase that is potentially involved in the regulation of fibrinolysis by decreasing plasminogen binding to the fibrin surface. This role makes the TAFI gene a good candidate in atherothrombotic diseases. The great interindividual variability of plasma TAFI antigen levels is poorly explained by lifestyle characteristics, thus suggesting its genetic determination. To test this hypothesis, the promoter and the 3'-untranslated region of the TAFI gene were screened for polymorphisms, and their contribution to the variability of plasma TAFI antigen levels was evaluated. Seven new polymorphisms are described, 5 in the promoter (C-2599G, -2345 2G/1G, A-1690G, G-1102T, and G-438A) and 2 in the 3'UTR (C+1542G and T+1583A). All these polymorphisms were in strong linkage disequilibrium with each other and with the previously described Ala147Thr polymorphism. They generated 4 main haplotypes, accounting for 80% of all observed haplotypes. In univariate analyses, all polymorphisms were associated with plasma TAFI Ag levels and, individually, contributed to a large fraction of plasma TAFI Ag levels, ranging from 20% to 52%. In a stepwise regression analysis including all polymorphisms, several combinations remained significantly and independently associated with plasma TAFI Ag levels: C+1542G associated with Ala147Thr, T+1583A, or -2345 2G/1G explaining 61.6%, 60.2%, and 58.1% of the variance, respectively. These findings clearly demonstrate that circulating levels of TAFI are strongly determined by polymorphic variations in the promoter and the 3'UTR of the TAFI gene. (Blood. 2001;97:2053-2058) PMID- 11264172 TI - Expression and characterization of von Willebrand factor dimerization defects in different types of von Willebrand disease. AB - Dimerization defects of von Willebrand factor (vWF) protomers underlie von Willebrand disease (vWD) type 2A, subtype IID (vWD 2A/IID), and corresponding mutations have been identified at the 3' end of the vWF gene in exon 52. This study identified and expressed 2 additional mutations in this region, a homozygous defect in a patient with vWD type 3 (C2754W) and a heterozygous frameshift mutation (8566delC) in a patient with vWD type 2A, subtype IIE. Both mutations involve cysteine residues that we propose are possibly essential for dimerization. To prove this hypothesis, transient recombinant expression of each of the 2 mutations introduced in the carboxy-terminal vWF fragment II and in the complete vWF complementary DNA, respectively, were carried out in COS-7 cells and compared with expression of vWD 2A/IID mutation C2773R and the wild-type (WT) sequence in COS-7 cells. Recombinant WT vWF fragment II assembled correctly into a dimer, whereas recombinant mutant fragments were monomeric. Homozygous expression of recombinant mutant full-length vWF resulted in additional dimers, probably through disulfide bonding at the amino-terminal multimerization site, whereas recombinant WT vWF correctly assembled into multimers. Coexpression of recombinant mutant and recombinant WT vWF reproduced the multimer patterns observed in heterozygous individuals. Our results suggest that a common defect of vWF biosynthesis--lack of vWF dimerization--may cause diverse types and subtypes of vWD. We also confirmed previous studies that found that disulfide bonding at the vWF amino-terminal is independent of dimerization at the vWF carboxy terminal. (Blood. 2001;97:2059-2066) PMID- 11264173 TI - TRAIL expression by activated human CD4(+)V alpha 24NKT cells induces in vitro and in vivo apoptosis of human acute myeloid leukemia cells. AB - Human Valpha24NKT cells are activated by alpha-galactosylceramide (alpha-GalCer) pulsed dendritic cells in a CD1d-dependent and a T-cell receptor-mediated manner. Here, we demonstrate that CD4(+)V alpha 24NKT cells derived from a patient with acute myeloid leukemia (AML) M4 are phenotypically similar to those of healthy donors and, in common with those derived from healthy donors, express tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) when the cells are activated by alpha-GalCer-pulsed dendritic cells but not prior to activation. We also show that myeloid leukemia cells from patients with AML M4, but not from patients with AML M0 or M1, undergo apoptosis following culture with TRAIL expressing autologous or allogeneic healthy donor V alpha 24NKT cells. Apoptosis of AML M4 leukemia cells from patient peripheral blood was almost completely blocked by a neutralizing monoclonal antibody against TRAIL, indicating that TRAIL on V alpha 24NKT cells is essential for the induction of apoptosis in AML M4 leukemia cells. A nonobese diabetic-severe combined immunodeficient human leukemia (AML M4) model showed that human activated CD4(+)V alpha 24NKT cells induced apoptosis of human leukemia cells in vivo. This is the first evidence that activated V alpha 24NKT cells express TRAIL and that TRAIL causes apoptosis of monocytic leukemia cells from patients with AML M4 in vitro and in vivo. Adoptive immune therapy with activated V alpha 24NKT cells, or other strategies to increase activated V alpha 24NKT cells in vivo, may be of benefit to patients with AML M4. (Blood. 2001;97:2067-2074) PMID- 11264174 TI - Inhibitory effect on natural killer activity of microphthalmia transcription factor encoded by the mutant mi allele of mice. AB - The mouse mi locus encodes a basic-helix-loop-helix-leucine zipper-type transcription factor, microphthalmia transcription factor (MITF). Mice of mi/mi genotype express a mutant form of MITF (mi-MITF), whereas mice of tg/tg genotype have a transgene in the 5' flanking region of the mi gene and do not express MITF. Although the mi/mi mouse is deficient in natural killer (NK) activity, it was found that the tg/tg mouse was normal in this respect. To know the cause, spleen cells of both genotypes were compared. Although the proportion of spleen cells expressing an NK cell marker, NK1.1, was comparable in both mice, the proportion of large granular lymphocytes decreased only in mi/mi mice. The difference between mi/mi and tg/tg mice was reproducible in the culture supplemented with interleukin-2. Moreover, the perforin gene expression was reduced in mi/mi-cultured spleen cells. Wild-type (+) MITF transactivated, but mi MITF suppressed, the perforin gene promoter through the NF-P motif, a strong cis acting element. However, neither +-MITF nor mi-MITF bound the NF-P motif. Instead, 2 nuclear factors that bound the NF-P motif were retained in the cytoplasm of mi/mi-cultured spleen cells. In addition, overexpression of mi-MITF resulted in cytoplasmic retention of the 2 NF-P motif-binding factors in cytotoxic T lymphocytes. The presence of mi-MITF rather than the absence of + MITF appeared to lead to poor transactivation of the NF-P motif by intercepting NF-P motif-binding factors. This inhibitory effect of mi-MITF may cause the deficient cytotoxicity of NK cells in mi/mi mice. (Blood. 2001;97:2075-2083) PMID- 11264175 TI - BCR-ABL down-regulates the DNA repair protein DNA-PKcs. AB - This study demonstrates in both stable and inducible BCR-ABL-expressing hematopoietic cells a down-regulation of the major mammalian DNA repair protein DNA-PKcs by BCR-ABL. Similar results were found in BCR-ABL CD34(+) cells from patients with chronic myelogenous leukemia (CML). DNA-PKcs down-regulation is a proteasome-dependent degradation that requires tyrosine kinase activity and is associated with a marked DNA repair deficiency along with increased sensitivity to ionizing radiation. The conjunction of a major DNA repair deficiency and a resistance to apoptosis, both induced by BCR-ABL, provides a new mechanism to explain how secondary genetic alterations can accumulate in CML, eventually leading to blast crisis. The down-regulation of DNA-PKcs was reversible in CD34(+) CML cells suggesting that this approach might offer a novel and powerful therapeutic strategy in this disease, especially to delay the blast crisis. (Blood. 2001;97:2084-2090) PMID- 11264176 TI - Cyclin A1 directly interacts with B-myb and cyclin A1/cdk2 phosphorylate B-myb at functionally important serine and threonine residues: tissue-specific regulation of B-myb function. AB - Cyclin A1 is tissue-specifically expressed during spermatogenesis, but it is also highly expressed in acute myeloid leukemia (AML). Its pathogenetic role in AML and in the cell cycle of leukemic blasts is unknown. B-myb is essential for G1/S transition and has been shown to be phosphorylated by the cyclin A2/cdk2 complex. Here it is demonstrated that cyclin A1 interacts with the C-terminal portion of B myb as shown by glutathione S-transferase (GST) precipitation. This interaction is confined to cyclin A1 because binding could not be detected between cyclin A2 and B-myb. Also, cdk2 was not pulled down by GST-B-myb from U937 lysates. In addition, co-immunoprecipitation of cyclin A1 and B-myb in leukemic cells evidenced protein interaction in vivo. Baculovirus-expressed cyclin A1/cdk2 complexes were able to phosphorylate human as well as murine B-myb in vitro. Tryptic phosphopeptide mapping revealed that cyclin A1/cdk2 complexes phosphorylated the C-terminal part of B-myb at several sites including threonine 447, 490, and 497 and serine 581. These phosphorylation sites have been demonstrated to be important for the enhancement of B-myb transcriptional activity. Further studies showed that cyclin A1 cooperated with B-myb to transactivate myb binding site containing promoters including the promoter of the human cyclin A1 gene. Taken together, the data suggest that cyclin A1 is a tissue specific regulator of B-myb function and activates B-myb in leukemic blasts. (Blood. 2001;97:2091-2097) PMID- 11264177 TI - Nucleotide sequence, transcription map, and mutation analysis of the 13q14 chromosomal region deleted in B-cell chronic lymphocytic leukemia. AB - Deletions of the 13q14 chromosome region are associated with B-cell chronic lymphocytic leukemia (B-CLL) and several other types of cancer, suggesting the presence of a tumor suppressor gene. In previous studies the minimal region of deletion (MDR) was mapped to a less than 300-kilobase (kb) interval bordered by the markers 173a12-82 and 138G4/1.3R. For the identification of the putative tumor suppressor gene, the entire MDR (approximately 347 kb) has been sequenced, and transcribed regions have been identified by exon trapping, EST-based full length complementary DNA cloning, database homology searches, and computer assisted gene prediction analyses. The MDR contains 2 pseudogenes and 3 transcribed genes: CAR, encoding a putative RING-finger containing protein; 1B4/Leu2, generating noncoding transcripts; and EST70/Leu1, probably representing another noncoding gene (longest open reading frame of 78 codons). These genes have been sequenced in 20 B-CLL cases with 13q14 hemizygous deletion, and no mutations were found. Moreover, no somatic variants were found in the entire MDR analyzed for nucleotide substitutions by a combination of direct sequencing and fluorescence-assisted mismatch analysis in 5 B-CLL cases displaying 13q14 monoallelic deletion. The nondeleted allele of the CAR and EST70/Leu1 genes was expressed in B-CLL specimens, including those with monoallelic loss, whereas no expression of 1B4/Leu2 was detectable in B-CLL, regardless of the 13q14 status. These results indicate that allelic loss and mutation of a gene within the MDR is an unlikely pathogenetic mechanism for B-CLL. However, haplo-insufficiency of one of the identified genes may contribute to tumorigenesis. (Blood. 2001;97:2098 2104) PMID- 11264178 TI - Synergistic induction of apoptosis in human leukemia cells (U937) exposed to bryostatin 1 and the proteasome inhibitor lactacystin involves dysregulation of the PKC/MAPK cascade. AB - Cotreatment with a minimally toxic concentration of the protein kinase C (PKC) activator (and down-regulator) bryostatin 1 (BRY) induced a marked increase in mitochondrial dysfunction and apoptosis in U937 monocytic leukemia cells exposed to the proteasome inhibitor lactacystin (LC). This effect was blocked by cycloheximide, but not by alpha-amanitin or actinomycin D. Qualitatively similar interactions were observed with other PKC activators (eg, phorbol 12-myristate 13 acetate and mezerein), but not phospholipase C, which does not down-regulate the enzyme. These events were examined in relationship to functional alterations in stress (eg, SAPK, JNK) and survival (eg, MAPK, ERK) signaling pathways. The observations that LC/BRY treatment failed to trigger JNK activation and that cell death was unaffected by a dominant-interfering form of c-JUN (TAM67) or by pretreatment with either curcumin or the p38/RK inhibitor, SB203580, suggested that the SAPK pathway was not involved in potentiation of apoptosis. In marked contrast, perturbations in the PKC/Raf/MAPK pathway played an integral role in LC/BRY-mediated cell death based on evidence that pretreatment of cells with bisindolylmaleimide I, a selective PKC inhibitor, or geldanamycin, a benzoquinone ansamycin, which destabilizes and depletes Raf-1, markedly suppressed apoptosis. Furthermore, ERK phosphorylation was substantially prolonged in LC/BRY-treated cells compared to those exposed to BRY alone, and pretreatment with the highly specific MEK inhibitors, PD98059, U0126, and SL327, opposed ERK activation while protecting cells from LC/BRY-induced lethality. Together, these findings suggest a role for activation and/or dysregulation of the PKC/MAPK cascade in modulation of leukemic cell apoptosis following exposure to the proteasome inhibitor LC. (Blood. 2001;97:2105-2114) PMID- 11264179 TI - Identification of novel markers for monitoring minimal residual disease in acute lymphoblastic leukemia. AB - To identify new markers of minimal residual disease (MRD) in B-lineage acute lymphoblastic leukemia (ALL), gene expression of leukemic cells obtained from 4 patients with newly diagnosed ALL was compared with that of normal CD19(+)CD10(+) B-cell progenitors obtained from 2 healthy donors. By cDNA array analysis, 334 of 4132 genes studied were expressed 1.5- to 5.8-fold higher in leukemic cells relative to both normal samples; 238 of these genes were also overexpressed in the leukemic cell line RS4;11. Nine genes were selected among the 274 overexpressed in at least 2 leukemic samples, and expression of the encoded proteins was measured by flow cytometry. Two proteins (caldesmon and myeloid nuclear differentiation antigen) were only weakly expressed in leukemic cells despite strong hybridization signals in the array. By contrast, 7 proteins (CD58, creatine kinase B, ninjurin1, Ref1, calpastatin, HDJ-2, and annexin VI) were expressed in B-lineage ALL cells at higher levels than in normal CD19(+)CD10(+) B cell progenitors (P <.05 in all comparisons). CD58 was chosen for further analysis because of its abundant and prevalent overexpression. An anti-CD58 antibody identified residual leukemic cells (0.01% to 1.13%; median, 0.03%) in 9 of 104 bone marrow samples from children with ALL in clinical remission. MRD estimates by CD58 staining correlated well with those of polymerase chain reaction amplification of immunoglobulin genes. These results indicate that studies of gene expression with cDNA arrays can aid the discovery of leukemia markers. (Blood. 2001;97:2115-2120) PMID- 11264181 TI - Kaposi sarcoma-associated herpesvirus infects monotypic (IgM lambda) but polyclonal naive B cells in Castleman disease and associated lymphoproliferative disorders. AB - In a previous study, it was shown that the Kaposi sarcoma-associated herpesvirus (KSHV) was specifically associated with monotypic (IgMlambda) plasmablasts in multicentric Castleman disease (MCD). The plasmablasts occur as isolated cells in the mantle zone of B-cell follicles but may form microlymphoma or frank plasmablastic lymphoma. To determine the clonality and cellular origin of the monotypic plasmablasts, the rearranged Ig genes in 13 patients with KSHV-related MCD, including 8 cases with microlymphomas and 2 with frank lymphomas, were studied. To investigate the role of the interleukin 6 (IL-6) receptor signaling in the pathogenesis of MCD and associated lymphoproliferative disorders, viral IL 6 and human IL-6 receptor expression was examined. KSHV-positive plasmablasts were polyclonal in MCD-involved lymphoid tissues in all cases and microlymphomas in 6 of 8 cases. Monoclonal KSHV-positive plasmablasts were seen in microlymphomas of 2 cases and in both frank lymphomas. Despite their mature phenotype, KSHV-positive plasmablasts did not harbor somatic mutations in the rearranged Ig genes, indicating origination from naive B cells. Viral IL-6 was expressed in 10% to 15% of KSHV-positive plasmablasts, whereas the human IL-6 receptor was expressed in most KSHV-positive cells. Thus, KSHV infects monotypic but polyclonal naive B cells and is associated with a range of lymphoproliferative disorders from polyclonal isolated plasmablasts and microlymphomas to monoclonal microlymphoma and frank plasmablastic lymphomas in MCD patients. Activation of the IL-6 receptor signaling pathway may play a role in differentiation of KSHV-infected naive B cells into plasmablasts and development of lymphoproliferative lesions. (Blood. 2001;97:2130-2136) PMID- 11264180 TI - Endothelial cell activation by myeloblasts: molecular mechanisms of leukostasis and leukemic cell dissemination. AB - Leukostasis and tissue infiltration by leukemic cells are poorly understood life threatening complications of acute leukemia. This study has tested the hypothesis that adhesion receptors and cytokines secreted by blast cells play central roles in these reactions. Immunophenotypic studies showed that acute myeloid leukemia (AML) cells (n = 78) of the M0 to M5 subtypes of the French-American-British Cooperative Group expressed various amounts of adhesion receptors, including CD11a, b, c/CD18, CD49d, e, f/CD29, CD54, sCD15, and L-selectin. The presence of functional adhesion receptors was evaluated using a nonstatic adhesion assay. The number of blast cells attached to unactivated endothelium increased by 7 to 31 times after a 6-hour exposure of endothelium to tumor necrosis factor (TNF) alpha. Inhibition studies showed that multiple adhesion receptors--including L selectin, E-selectin, VCAM-1, and CD11/CD18--were involved in blast cell adhesion to TNF-alpha-activated endothelium. Leukemic cells were then cocultured at 37 degrees C on unactivated endothelial cell monolayers for time periods up to 24 hours. A time-dependent increase in the number of blasts attached to the endothelium and a concomitant induction of ICAM-1, VCAM-1, and E-selectin were observed. Additional experiments revealed that endothelial cell activation by leukemic myeloblasts was caused by cytokine secretion by blast cells, in particular TNF-alpha and IL-1 beta, and direct contacts between adhesion receptors expressed by blast cells and endothelial cells. Thus, leukemic cells have the ability to generate conditions that promote their own adhesion to vascular endothelium, a property that may have important implications for the pathophysiology of leukostasis and tissue infiltration by leukemic blast cells. (Blood. 2001;97:2121-2129) PMID- 11264183 TI - Evidence for linkage of familial Diamond-Blackfan anemia to chromosome 8p23.3-p22 and for non-19q non-8p disease. AB - Diamond-Blackfan anemia (DBA) is a rare congenital hypoplastic anemia that usually presents early in infancy and is inherited in 10% to 20% of cases. Linkage analysis has shown that DBA in many of both dominant and recessive DBA families mapped to chromosome 19q13.2 leading to the cloning of a gene on chromosome 19q13.2 that encodes a ribosomal protein, RPS19. However, subsequently, mutations of the RPS19 gene have only been identified in 25% of all patients with DBA. This study analyzed 14 multiplex DBA families, 9 of which had 19q13.2 haplotypes inconsistent with 19q linkage. A genome-wide search for linked loci suggested the presence of a second DBA locus in a 26.4-centimorgan (cM) interval on human chromosome 8p. Subsequently, 24 additional DBA families were ascertained and all 38 families were analyzed with additional polymorphic markers on chromosome 8p. In total, 18 of 38 families were consistent with linkage to chromosome 8p with a maximal LOD score with heterogeneity of 3.55 at D8S277 assuming 90% penetrance. The results indicate the existence of a second DBA gene in the 26.4-cM telomeric region of human chromosome 8p23.3-p22, most likely within an 8.1-cM interval flanked by D8S518 and D8S1825. Seven families were inconsistent with linkage to 8p or 19q and did not reveal mutations in the RPS19 gene, suggesting further genetic heterogeneity. (Blood. 2001;97:2145-2150) PMID- 11264182 TI - Human T-cell leukemia virus type I oncoprotein Tax represses Smad-dependent transforming growth factor beta signaling through interaction with CREB-binding protein/p300. AB - Human T-cell leukemia virus type I (HTLV-I) Tax is a potent transcriptional regulator that can activate or repress specific cellular genes and that has been proposed to contribute to leukemogenesis in adult T-cell leukemia. Previously, HTLV-I- infected T-cell clones were found to be resistant to growth inhibition by transforming growth factor (TGF)-beta. Here it is shown that Tax can perturb Smad dependent TGF-beta signaling even though no direct interaction of Tax and Smad proteins could be detected. Importantly, a mutant Tax of CREB-binding protein (CBP)/p300 binding site, could not repress the Smad transactivation function, suggesting that the CBP/p300 binding domain of Tax is essential for the suppression of Smad function. Because both Tax and Smad are known to interact with CBP/p300 for the potentiation of their transcriptional activities, the effect of CBP/p300 on suppression of Smad-mediated transactivation by Tax was examined. Overexpression of CBP/p300 reversed Tax-mediated inhibition of Smad transactivation. Furthermore, Smad could repress Tax transcriptional activation, indicating reciprocal repression between Tax and Smad. These results suggest that Tax interferes with the recruitment of CBP/p300 into transcription initiation complexes on TGF-beta-responsive elements through its binding to CBP/p300. The novel function of Tax as a repressor of TGF-beta signaling may contribute to HTLV I leukemogenesis. (Blood. 2001;97:2137-2144) PMID- 11264184 TI - Reduced oxidative-stress response in red blood cells from p45NFE2-deficient mice. AB - p45NF-E2 is a member of the cap 'n' collar (CNC)-basic leucine zipper family of transcriptional activators that is expressed at high levels in various types of blood cells. Mice deficient in p45NF-E2 that were generated by gene targeting have high mortality from bleeding resulting from severe thrombocytopenia. Surviving p45nf-e2(-/-) adults have mild anemia characterized by hypochromic red blood cells (RBCs), reticulocytosis, and splenomegaly. Erythroid abnormalities in p45nf-e2(-/-) animals were previously attributed to stress erythropoiesis caused by chronic bleeding and, possibly, ineffective erythropoiesis. Previous studies suggested that CNC factors might play essential roles in regulating expression of genes that protect cells against oxidative stress. In this study, we found that p45NF-E2-deficient RBCs have increased levels of reactive oxygen species and an increased susceptibility to oxidative-stress-induced damage. Deformability of p45NF-E2-deficient RBCs was markedly reduced with oxidative stress, and mutant cells had a reduced life span. One possible reason for increased sensitivity to oxidative stress is that catalase levels were reduced in mutant RBCs. These findings suggest a role for p45NF-E2 in the oxidative-stress response in RBCs and indicate that p45NF-E2 deficiency contributes to the anemia in p45nf-e2(-/-) mice. (Blood. 2001;97:2151-2158) PMID- 11264186 TI - Stroke in hemoglobin (SD) sickle cell disease with moyamoya: successful hydroxyurea treatment after cerebrovascular bypass surgery. AB - An 11-year-old boy with hemoglobin sickle disease (HbSD), bilateral stenosis of the intracranial carotid arteries, and moyamoya syndrome had recurrent ischemic strokes with aphasia and right hemiparesis. His parents (Jehovah's Witnesses) refused blood transfusions. After bilateral extracranial-intracranial (EC-IC) bypass surgery, hydroxyurea treatment increased hemoglobin F (HbF) levels to more than 30%. During a follow-up of 28 months, flow velocities in the basal cerebral arteries remained stable, neurologic sequelae regressed, and ischemic events did not recur. This is the first report of successful hydroxyurea treatment after bypass surgery for intracranial cerebral artery obstruction with moyamoya syndrome in sickle cell disease. The patient's religious background contributed to an ethically challenging therapeutic task. (Blood. 2001;97:2165-2167) PMID- 11264185 TI - Integrin-associated protein is an adhesion receptor on sickle red blood cells for immobilized thrombospondin. AB - The adhesive protein thrombospondin (TSP) potentially mediates sickle (SS) red blood cell (RBC) adhesion to the blood vessel wall, thereby contributing to vaso occlusive crises in sickle cell disease. We previously reported that SS RBCs bind to immobilized TSP under flow conditions, whereas normal (AA) red cells do not. However, the SS RBC receptors that mediate this interaction are largely unknown. Here it is reported that integrin-associated protein (IAP), or CD47, mediates the adhesion of these cells to immobilized TSP under both flow and static conditions. A peptide derived from the C-terminal IAP binding site of TSP also supports sickle cell adhesion; adhesion to this peptide or to TSP is inhibited specifically by the anti-IAP monoclonal antibody, 1F7. Furthermore, these data suggest that IAP on SS RBCs is structurally different from that expressed on AA RBCs but that IAP expression levels do not vary between AA and SS RBCs. This structural difference may contribute to the enhanced adhesion of SS RBCs to immobilized TSP. These results identify IAP as a TSP receptor on SS RBCs and suggest that this receptor and its binding site within TSP represent potential therapeutic targets to decrease vaso-occlusion. (Blood. 2001;97:2159-2164) PMID- 11264188 TI - Autoantibodies to alpha(IIb)beta(3) in patients with chronic immune thrombocytopenic purpura bind primarily to epitopes on alpha(IIb). AB - Chronic immune thrombocytopenic purpura (ITP) is an autoimmune disease caused by platelet destruction resulting from autoantibodies against platelet surface proteins, particularly platelet glycoprotein IIb/IIIa (alpha(IIb)beta(3)). To localize the auto-epitopes on platelet alpha(IIb)beta(3), the binding of autoantibodies to Chinese hamster ovary (CHO) cells expressing either alpha(IIb)beta(3) or alpha(v)beta(3) was studied. Thirteen of 14 ITP autoantibodies bound only to CHO cells expressing alpha(IIb)beta(3). Because these 2 integrins have the same beta chain (beta(3)), these results show that most epitopes in chronic ITP are dependent on the presence of glycoprotein alpha(IIb.) (Blood. 2001;97:2171-2172) PMID- 11264187 TI - Fusion AML1 transcript in a radiation-associated leukemia results in a truncated inhibitory AML1 protein. AB - AML1 is a transcription factor that is essential for normal hematopoietic development. It is the most frequent target for translocations in acute leukemia. Recently, fluorescence in situ hybridization was used to identify a novel syndrome of radiation-associated secondary acute myelogenous leukemia that had AML1 translocations. Using polymerase chain reaction, the AML1 fusion transcript was isolated from the patient who had a t(19;21) radiation-associated leukemia. The AML1 gene is fused out of frame to chromosome 19 sequences, resulting in a truncated AML protein bearing the DNA binding domain but not the transcriptional activation domain. This fusion AML1 protein functions as an inhibitor of the normal AML1 protein. (Blood. 2001;97:2168-2170) PMID- 11264189 TI - Detection of viral interleukin-6 in Kaposi sarcoma-associated herpesvirus-linked disorders. AB - Expression of a viral interleukin-6 (vIL-6) has been detected in certain Kaposi sarcoma (KS)--associated herpesvirus positive (KSHV(+)) lesions. The release of vIL-6 systemically and its contribution to the pathogenesis of HIV-related malignancies was studied. Serum vIL-6 was detected in 13 (38.2%) of 34 HIV(+) patients with KS, in 6 (85.7%) of 7 HIV(+) patients with primary effusion lymphoma (PEL) and/or multicentric Castleman disease (MCD), and in 18 (60.0%) of 30 HIV(+), mostly homosexual, individuals without KS, MCD, or PEL. By contrast, serum vIL-6 was detected in only 3 (23.1%) of 13 patients with classic KS, 1 (2.5%) of 40 blood donors from the United States, and 4 (19.0%) of 21 blood donors from Italy. Circulating vIL-6 levels were associated with HIV(+) status (P <.0001). However, within the HIV(+) cohort, serum vIL-6 levels were not associated with the occurrence of KSHV-associated malignancies (P =.43). (Blood. 2001;97:2173-2176) PMID- 11264190 TI - Expression of tumor-suppressor genes interferon regulatory factor 1 and death associated protein kinase in primitive acute myelogenous leukemia cells. AB - Previous studies indicate that human acute myelogenous leukemia (AML) arises from a rare population of leukemic stem cells. Cells of this nature can initiate and maintain leukemic cell growth in both long-term cultures and nonobese diabetic/severe combined immune-deficient mice. To characterize the biology of primitive AML cells, gene expression screens were performed with 7 primary AML and 3 normal specimens. For each sample, stem cell populations (CD34(+)/CD38(-)) were isolated and used to synthesize radiolabeled complementary DNA (cDNA). AML vs normal probes were then hybridized to cDNA arrays containing genes related to cancer and apoptosis. Of approximately 1400 genes analyzed, 2 tumor-suppressor genes were identified that were overexpressed in all 7 of the AML CD34(+)/CD38(-) cell populations: death-associated protein kinase and interferon regulatory factor 1. Expression of each gene was confirmed by reverse-transcription polymerase chain reaction and immunoblot analysis. It is proposed that tumor suppressor proteins play a role in the biology of primitive AML cells. (Blood. 2001;97:2177-2179) PMID- 11264191 TI - Closing the gap between professional teaching and practice. PMID- 11264192 TI - Thromboprophylaxis after replacement arthroplasty. PMID- 11264193 TI - Abdominal obesity and the "hypertriglyceridaemic waist" phenotype. PMID- 11264194 TI - Reforming the GMC. PMID- 11264195 TI - Death through selfishness and failure of imagination. PMID- 11264196 TI - Regulators accused of bias against cannabis based medicines. PMID- 11264197 TI - Plastic surgeon sacked days before employment tribunal due to begin. PMID- 11264198 TI - Cancer registries fear collapse. Need for patient consent for cancer registration creates logistical nightmare. PMID- 11264199 TI - Drug company to sell AIDS drugs at less than cost price. PMID- 11264202 TI - Indian scientists warn of "mutant measles" virus. PMID- 11264203 TI - Rubella vaccine may be safe in early pregnancy. PMID- 11264204 TI - Doctors give guarded response to 100m pounds sterling for GP services. PMID- 11264205 TI - Company played down drug's risks, report says. PMID- 11264206 TI - Effectiveness and economic evaluation of a nurse delivered home exercise programme to prevent falls. 1: Randomised controlled trial. AB - OBJECTIVES: To assess the effectiveness of a trained district nurse individually prescribing a home based exercise programme to reduce falls and injuries in elderly people and to estimate the cost effectiveness of the programme. DESIGN: Randomised controlled trial with one year's follow up. SETTING: Community health service at a New Zealand hospital. PARTICIPANTS: 240 women and men aged 75 years and older. INTERVENTION: 121 participants received the exercise programme (exercise group) and 119 received usual care (control group); 90% (211 of 233) completed the trial. MAIN OUTCOME MEASURES: Number of falls, number of injuries resulting from falls, costs of implementing the programme, and hospital costs as a result of falls. RESULTS: Falls were reduced by 46% (incidence rate ratio 0.54, 95% confidence interval 0.32 to 0.90). Five hospital admissions were due to injuries caused by falls in the control group and none in the exercise group. The programme cost $NZ1803 (523 pound sterling) (at 1998 prices) per fall prevented for delivering the programme and $NZ155 per fall prevented when hospital costs averted were considered. CONCLUSION: A home exercise programme, previously shown to be successful when delivered by a physiotherapist, was also effective in reducing falls when delivered by a trained nurse from within a home health service. Serious injuries and hospital admissions due to falls were also reduced. The programme was cost effective in participants aged 80 years and older compared with younger participants. PMID- 11264207 TI - Effectiveness and economic evaluation of a nurse delivered home exercise programme to prevent falls. 2: Controlled trial in multiple centres. AB - OBJECTIVES: To assess the effectiveness of trained nurses based in general practices individually prescribing a home exercise programme to reduce falls and injuries in elderly people and to estimate the cost effectiveness of the programme. DESIGN: Controlled trial with one year's follow up. SETTING: 32 general practices in seven southern New Zealand centres. PARTICIPANTS: 450 women and men aged 80 years and older. INTERVENTION: 330 participants received the exercise programme (exercise centres) and 120 received usual care (control centres); 87% (371 of 426) completed the trial. MAIN OUTCOME MEASURES: Number of falls, number of injuries resulting from falls, costs of implementing the programme, and hospital costs as a result of falls. RESULTS: Falls were reduced by 30% in the exercise centres (incidence rate ratio 0.70, 95% confidence interval 0.59 to 0.84). The programme was equally effective in men and women. The programme cost $NZ418 (121 pound sterling) (at 1998 prices) per person to deliver for one year or $NZ1519 (441 pound sterling) per fall prevented. Fewer participants had falls resulting in injuries, but there was no difference in the number who had serious injuries and no difference in hospital costs resulting from falls in exercise centres compared with control centres. CONCLUSIONS: An individually tailored exercise programme, delivered by trained nurses from within general practices, was effective in reducing falls in three different centres. This strategy should be combined with other successful interventions to form part of home programmes to prevent falls in elderly people. PMID- 11264209 TI - Understanding the clinical dilemmas that shape medical students' ethical development: questionnaire survey and focus group study. PMID- 11264211 TI - Cancer registries fear collapse. BUPA wants to ensure systematic transfer of data. PMID- 11264208 TI - Benzodiazepines and hip fractures in elderly people: case-control study. AB - OBJECTIVE: To determine whether benzodiazepines are associated with an increased risk of hip fracture. DESIGN: Case-control study. PARTICIPANTS: All incident cases of hip fracture not related to traffic accidents or cancer in patients over 65 years of age. 245 cases were matched to 817 controls. SETTING: Emergency department of a university hospital. MAIN OUTCOME MEASURES: Exposure to benzodiazepines and other potential risk or protective factors or lifestyle items. RESULTS: The use of benzodiazepines as determined from questionnaires, medical records, or plasma samples at admission to hospital was not associated with an increased risk of hip fracture (odds ratio 0.9, 95% confidence interval 0.5 to 1.5). Hip fracture was, however, associated with the use of two or more benzodiazepines, as determined from questionnaires or medical records but not from plasma samples. Of the individual drugs, only lorazepam was significantly associated with an increased risk of hip fracture (1.8, 1.1 to 3.1). CONCLUSION: Except for lorazepam, the presence of benzodiazepines in plasma was not associated with an increased risk of hip fracture. The method used to ascertain exposure could influence the results of case-control studies. PMID- 11264210 TI - Sex differences in speed of emergence and quality of recovery after anaesthesia: cohort study. PMID- 11264212 TI - Model for directly assessing and improving clinical competence and performance in revalidation of clinicians. PMID- 11264213 TI - Treatment of obesity: need to focus on high risk abdominally obese patients. PMID- 11264214 TI - ABC of diseases of liver, pancreas, and biliary system. Pancreatic tumours. PMID- 11264215 TI - Postpsychiatry: a new direction for mental health. PMID- 11264216 TI - Promoting health and function in an ageing population. PMID- 11264217 TI - Using internet to access confidential patient records. Information about NHSnet was incorrect. PMID- 11264219 TI - Postoperative pressure sores after epidural anaesthesia. Good nursing care should prevent pressure sores. PMID- 11264218 TI - Undertreatment of heart failure has high cost to patients. PMID- 11264220 TI - Menorrhagia. Underlying bleeding disorders need to be ruled out. PMID- 11264221 TI - Menorrhagia. Sexual history needs to be taken. PMID- 11264223 TI - Menorrhagia. Ten minutes may not be enough. PMID- 11264222 TI - Postoperative pressure sores after epidural anaesthesia. Informed nursing care is needed. PMID- 11264224 TI - Postoperative pressure sores after epidural anaesthesia. Anaesthetists and their teams should examine heels of patients. PMID- 11264225 TI - Rights involve responsibilities for patients. PMID- 11264226 TI - Postoperative pressure sores after epidural anaesthesia. Staff needs to recognise patients are at risk. PMID- 11264227 TI - Evidence for a non-adrenoceptor, imidazoline-mediated contractile response to oxymetazoline in the porcine isolated rectal artery. AB - Imidazoline derivatives are known to elicit responses through both alpha(2) adrenoceptor and non-adrenoceptor, imidazoline sites, though as yet there are no examples of the latter on vascular smooth muscle. In the presence of 0.3 microM prazosin, neither UK-14304 (0.01 - 3 microM) nor oxymetazoline (0.01 - 30 microM) caused a significant contraction of the porcine isolated rectal artery, a preparation with a low density of alpha(2)-adrenoceptors. In the presence of a combination of U46619 and forskolin, however, both agonists produced concentration-dependent contractions. Pretreatment with phenoxybenzamine (3 microM) abolished responses to UK-14304, but left those elicited by oxymetazoline largely unaffected. The putative I(3) imidazoline antagonist 2-(2,3 dihydro-2 benzofuranyl)-2-imidazole (KU-14R, 10 microM) caused a 6 fold rightward displacement of the phenoxybenzamine-insensitive concentration - response curve to oxymetazoline. Our data indicates that non-adrenoceptor, imidazoline sites, pharmacologically similar to the I(3) imidazoline site on islet cells, mediate vasoconstriction in the porcine isolated rectal artery. PMID- 11264229 TI - Effects of anti-oestrogens and beta-estradiol on calcium uptake by cardiac sarcoplasmic reticulum. AB - 1. Tamoxifen and a group of structurally similar non-steroidal, triphenolic compounds inhibit the oestrogen receptor. In addition to this action, these anti oestrogens are known to inhibit some types of plasma membrane ion channels and other proteins through mechanisms that do not appear to involve their interactions with the estrogen receptor but could be the result of their effect on membrane lipid structure or fluidity. 2. We studied the effects of beta estradiol and three anti-oestrogens (tamoxifen, 4-hydroxytamoxifen and clomiphene) on Ca(2+) uptake into sarcoplasmic reticulum (SR) vesicles isolated from canine cardiac ventricular tissue. 3. The antiestrogens all inhibit SR Ca(2+) uptake in a concentration-dependent manner (order of potency: tamoxifen > 4-hydroxytamoxifen > or = clomiphene). Although these compounds rapidly inhibit net Ca(2+) uptake they do not have a similar rapid effect on the ATPase activity of the SR Ca pump. beta-estradiol has no effect on Ca(2+) uptake nor does it alter the inhibitory action of tamoxifen on the SR. 4. The differences in the effects of beta-estradiol and the anti-oestrogens on cardiac SR Ca(2+) uptake do not correlate with differences in the ways in which these compounds have been reported to interact with membrane lipids. Our results are consistent, however, with direct effects on a membrane protein (possibly an SR Cl(-) or K(+) channel). PMID- 11264230 TI - Inhibition of aggregation of rabbit and human platelets induced by adrenaline and 5-hydroxytryptamine by KB-R7943, a Na(+)/Ca(2+) exchange inhibitor. AB - 1. We investigated the effect of KB-R7943, a Na(+)/Ca(2+) exchange inhibitor, on the aggregation response induced by adrenaline and 5-hydroxytryptamine (5-HT), alone or in combination in human and rabbit platelets in the presence or absence of ouabain. 2. KB-R7943 inhibited aggregation induced by the combination of adrenaline and 5-HT in a concentration-dependent manner. The IC(50) values of KB R7943 were 4.2+/-2.0 or 3.0+/-0.7 microM with washed rabbit platelets with or without ouabain pretreatment, respectively. 3. In platelet-rich human plasma, the aggregation was biphasic. The IC(50) value of KB-R7943 was 17.2+/-4.4 microM for the first phase aggregation. 4. KB-R7943 did not inhibit the first phase of aggregation induced by adrenaline alone, or the monophasic aggregation induced by 5-HT alone. 5. The aggregation of rabbit platelets depended on the presence of K(+) in the medium, and K(+)-dependent and K(+)-independent Ca(2+) influx were observed in resting platelets. Ouabain treatment increased only the K(+) dependent but not the K(+)-independent Ca(2+) influx. 6. KB-R7943 inhibited K(+) dependent Ca(2+) influx with or without ouabain pretreatment, but not K(+) independent Ca(2+) influx. 7. From these results, we conclude that KB-R7943 inhibits the adrenaline plus 5-HT induced aggregation of rabbit and human platelets by inhibiting K(+)-dependent Na(+)/Ca(2+) exchange (NCKX). Our results suggest that NCKX plays an important role in platelet aggregation. PMID- 11264228 TI - Chronic vascular toxicity of doxorubicin in an organ-cultured artery. AB - 1. We investigated the chronic effects of doxorubicin (DXR) on morphological and functional changes in the rabbit mesenteric artery using an organ culture system. 2. In arteries cultured with 0.3 microM DXR for 7 days, the contractions induced by noradrenaline, but not those induced by endothelin-1 or high K(+), were strongly inhibited. This reaction was followed by a decrease in the induction of the alpha(1A)-adrenoceptor without any change in the mRNA level. Inhibition of noradrenaline-induced contractions by DXR was attenuated by superoxide dismutase, and alpha(1A)-adrenoceptor protein expression recovered. 3. In the arteries cultured with 1 microM DXR for 7 days, contractions induced by endothelin-1 or high K(+) and absolute force in the permeabilized muscles were also inhibited. Morphological examinations revealed the existence of concentrated nuclei and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) positive smooth muscle cells, and internucleosomal DNA fragmentation was also detected, indicating the induction of apoptosis. 4. In the arteries cultured with 10 microM DXR for 7 days, nuclear swelling, karyolysis and random DNA fragmentation indicative of necrosis were observed, and muscle contractility was abolished. 5. These results suggest that 0.3 microM DXR selectively down regulates the alpha(1A)-adrenoceptor protein expression, resulting in a decrease in the noradrenaline-induced contraction. This down-regulation may be at least partly due to the production of a superoxide radical. DXR also caused a decrease in muscle contractility followed by apoptotic changes at 1 microM and necrotic changes at 10 microM. These changes might be responsible for the disturbance of the circulatory system that is often observed during the course of repetitive chemotherapy. PMID- 11264231 TI - Social isolation modifies nicotine's effects in animal tests of anxiety. AB - 1. These experiments determined whether the housing conditions of rats influenced the effects of nicotine in two animal tests of anxiety, social interaction and elevated plus-maze tests. 2. In animals housed singly for 7 days, (-)nicotine (0.025 mg kg(-1) s.c.) was ineffective, but 0.05, 0.1 and 0.25 mg kg(-1) (s.c.) significantly increased the time spent in social interaction, without changing locomotor activity, thus indicating anxiolytic actions. (-)Nicotine (0.45 mg kg( 1) s.c.) significantly reduced social interaction, indicating an anxiogenic effect. 3. However, in group-housed animals, (-)nicotine (0.025 mg kg(-1) s.c.) had a significant anxiolytic effect in the social interaction test, but 0.01, 0.05, 0.1, 0.25 and 0.45 mg kg(-1) were ineffective. (-)Nicotine (1 mg kg(-1)) reduced motor activity and social interaction in the group-housed animals. 4. In the elevated plus-maze, the time-course and the dose-response curve to nicotine were investigated. In both singly- and group-housed rats, (-) nicotine (0.1 - 0.45 mg kg(-1) s.c.) decreased the per cent entries into, and per cent time spent on, the open arms, indicating anxiogenic effects. 5. The housing condition influenced the time course, with significant effects at 5 and 30 min after injection in group-housed rats, and significant effects at 30 and 60 min in singly-housed rats. 6. In the social interaction test there was no difference in the scores of the first and last rats removed from group cages, whereas the order of removal from the cages did affect the scores in the elevated plus-maze. 7. These results provide further evidence that the two animal tests model distinct states of anxiety, and show how social isolation powerfully modifies both anxiolytic and anxiogenic effects of nicotine. PMID- 11264232 TI - Xenopus tropicalis oocytes as an advantageous model system for the study of intracellular Ca(2+) signalling. AB - 1. The purpose of this study was to compare oocytes from the pipid frogs Xenopus tropicalis and Xenopus laevis, with respect to their utility for studying Ca(2+) signalling mechanisms and for expression of heterologous proteins. 2. We show that X. tropicalis oocytes possess an intracellular Ca(2+) store that is mobilized by inositol (1,4,5) trisphosphate (IP(3)). Ca(2+) signalling is activated by endogenous lysophosphatidic acid receptors and cytosolic Ca(2+) activates a plasma membrane chloride conductance. The spatiotemporal organization of cytosolic Ca(2+) signals, from the microscopic architecture of elementary Ca(2+) 'puffs' to the macroscopic patterns of Ca(2+) spiking are closely similar to the local and global patterns of Ca(2+) release previously characterized in oocytes from X. laevis. 3. By injecting X. tropicalis oocytes with cDNA encoding an ER-targeted fluorescent protein construct, we demonstrate the capacity of the X. tropicalis oocyte to readily express heterologous proteins. The organization of ER is polarized across the oocyte, with the IP(3)-releaseable store targeted within an approximately 8 microm wide band that circumscribes the cell. 4. We conclude that the X. tropicalis oocyte shares many of the characteristics that have made oocytes of X. laevis a favoured system for studying Ca(2+) signalling mechanisms. Moreover, X. tropicalis oocytes display further practical advantages in terms of imaging depth, Ca(2+) signal magnitude and electrical properties. These further enhance the appeal of X. tropicalis as an experimental system, in addition to its greater amenability to transgenic approaches. PMID- 11264233 TI - Effect of vanilloid drugs on gastrointestinal transit in mice. AB - 1. We have studied the effect of capsaicin, piperine and anandamide, drugs which activate vanilloid receptors and capsazepine, a vanilloid receptor antagonist, on upper gastrointestinal motility in mice. 2. Piperine (0.5 - 20 mg kg(-1) i.p.) and anandamide (0.5 - 20 mg kg(-1) i.p.), dose-dependently delayed gastrointestinal motility, while capsaicin (up to 3 mg kg(-1) i.p.) was without effect. Capsazepine (15 mg kg(-1) i.p.) neither per se affected gastrointestinal motility nor did it counteract the inhibitory effect of both piperine (10 mg kg( 1)) and anandamide (10 mg kg(-1)). 3. A per se non effective dose of SR141716A (0.3 mg kg(-1) i.p.), a cannabinoid CB(1) receptor antagonist, counteracted the inhibitory effect of anandamide (10 mg kg(-1)) but not of piperine (10 mg kg( 1)). By contrast, the inhibitory effect of piperine (10 mg kg(-1)) but not of anandamide (10 mg kg(-1)) was strongly attenuated in capsaicin (75 mg kg(-1) in total, s.c.)-treated mice. 4. Pretreatment of mice with N(G)-nitro-L-arginine methyl ester (25 mg kg(-1) i.p.), yohimbine (1 mg kg(-1), i.p.), naloxone (2 mg kg(-1) i.p.), or hexamethonium (1 mg kg(-1) i.p.) did not modify the inhibitory effect of both piperine (10 mg kg(-1)) and anandamide (10 mg kg(-1)). 5. The present study indicates that the vanilloid ligands anandamide and piperine, but not capsaicin, can reduce upper gastrointestinal motility. The effect of piperine involves capsaicin-sensitive neurones, but not vanilloid receptors, while the effect of anandamide involves cannabinoid CB(1), but not vanilloid receptors. PMID- 11264234 TI - Docosahexaenoic acid improves long-term potentiation attenuated by phospholipase A(2) inhibitor in rat hippocampal slices. AB - 1. We investigated the possible involvement of phospholipase A(2) (PLA(2)) and its products in long-term potentiation (LTP) in the CA1 neurotransmission of rat hippocampal slices. 2. Inhibitors of Ca(2+)-independent PLA(2) (iPLA(2)) prevented the induction of LTP without affecting the maintenance phase of LTP whereas Ca(2+)-dependent PLA(2) inhibitors were virtually ineffective, which suggests a pivotal role of iPLA(2) in the initiation of LTP. 3. We then investigated the effect of docosahexaenoic acid (DHA) and arachidonic acid (AA) on BEL (bromoenol lactone, an iPLA(2)-inhibitor) -impaired LTP, and found that either DHA or AA abolished the effect of BEL. However, DHA did not restore BEL attenuated LTP when applied after the tetanus. DHA per se affected neither the induction nor maintenance of LTP. Linoleic acid had no effects, either. 4. These results suggest that DHA is crucial for the induction of LTP and that endogenously released DHA during tetanus is sufficient to trigger the formation of LTP. PMID- 11264235 TI - Potential anxiolytic- and antidepressant-like effects of MPEP, a potent, selective and systemically active mGlu5 receptor antagonist. AB - 1. Several lines of evidence suggest a crucial involvement of glutamate in the mechanism of action of anxiolytic and/or antidepressant drugs. The involvement of group I mGlu receptors in anxiety and depression has also been proposed. Given the recent discovery of a selective and brain penetrable mGlu5 receptor antagonists, the effect of 2-methyl-6-(phenylethynyl)-pyridine (MPEP), i.e. the most potent compound described, was evaluated in established models of anxiety and depression. 2. Experiments were performed on male Wistar rats or male Albino Swiss or C57BL/6J mice. The anxiolytic-like effects of MPEP was tested in the conflict drinking test and the elevated plus-maze test in rats as well as in the four-plate test in mice. The antidepressant-like effect was estimated using the tail suspension test in mice and the behavioural despair test in rats. 3. MPEP (1 - 30 mg kg(-1)) induced anxiolytic-like effects in the conflict drinking test and the elevated plus-maze test in rats as well as in the four-plate test in mice. MPEP had no effect on locomotor activity or motor coordination. MPEP (1 - 20 mg kg(-1)) did shorten the immobility time in a tail suspension test in mice, however it was inactive in the behavioural despair test in rats. 4. These data suggest that selective mGlu5 receptor antagonists may play a role in the therapy of anxiety and/or depression, further studies are required to identify the sites and the mechanism of action of MPEP. PMID- 11264236 TI - Galpha(14) links a variety of G(i)- and G(s)-coupled receptors to the stimulation of phospholipase C. AB - 1. The bovine Galpha(14) is a member of the G(q) subfamily of G proteins that can regulate phospholipase Cbeta isoforms but the extent to which Galpha(14) recognizes different receptor classes is not known. 2. Galpha(14) was cotransfected with a variety of receptors in COS-7 cells, and agonist-induced stimulation of phospholipase C was then measured. 3. Activation of the type 2 but not type 1 somatostatin receptor in cells coexpressing Galpha(14) stimulated the accumulation of inositol phosphates; functional expression of both subtypes of somatostatin receptors was determined by the ability of somatostatin to inhibit cyclic AMP accumulation. 4. Among the three opioid receptors (mu, delta, and kappa), only the delta receptor was capable of stimulating IP formation when coexpressed with Galpha(14) in COS-7 cells. 5. A panel of G(i)- and G(s)-linked receptors was screened for their ability to stimulate IP accumulation via Galpha(14). The adenosine A(1), complement C5a, dopamine D(1), D(2) and D(5), formyl peptide, luteinizing hormone, secretin, and the three subtypes of melatonin (mt1, MT2, and Xenopus) receptors were all incapable of activating Galpha(14), while the alpha(2)- and beta(2)-adrenoceptors were able to do so. 6. Galpha(14)-mediated stimulation of phospholipase Cbeta was agonist dose dependent. These data demonstrate that although Galpha(14) can interact with different classes of receptors, it is much less promiscuous than Galpha(15) or Galpha(16). PMID- 11264237 TI - Evidence for functionally active protease-activated receptor-4 (PAR-4) in human vascular smooth muscle cells. AB - 1. This study investigates, whether in addition to the protease-activated receptor-1 (PAR-1), PAR-4 is present in vascular smooth muscle cells (SMC) of the human saphenous vein and whether this receptor is functionally active. PAR-1 and PAR-4 are stimulated by thrombin and by the synthetic peptides SFLLRN and GYPGQV, respectively. 2. mRNAs for both, PAR-1 and PAR-4, were detected in the SMC by using reverse transcriptase polymerase chain reaction (RT - PCR). 3. Treatment of the SMC with GYPGQV (200 microM) resulted in a transient increase in free intracellular calcium. This calcium signal was completely abolished after a preceding challenge with thrombin (10 nM), indicating homologous receptor desensitization. 4. Stimulation of the SMC with 10 nM thrombin or 200 microM SFLLRN caused a time-dependent activation of the extracellular signal-regulated kinases-1/2 (ERK-1/2) with a maximum at 5 min. In contrast, 100 nM thrombin as well as 200 microM of GYPGQV induced a prolonged phosphorylation of ERK-1/2 with a maximum at 60 min. These data suggest that PAR-1 and PAR-4 are activated by thrombin at distinct concentrations and with distinct kinetics. 5. GYPGQV stimulated [(3)H]-thymidine incorporation in SMC. At 500 microM, the peptide increased DNA synthesis 2.5 fold above controls. A comparable mitogenic effect was obtained after stimulation of the SMC by 10 nM thrombin or 100 microM SFLLRN, respectively. 6. These data indicate that a functionally active PAR-4 is present in SMC and, in addition to PAR-1, might contribute to thrombin-induced mitogenesis. PMID- 11264238 TI - Role of endothelin and vasopressin in DOCA-salt hypertension. AB - 1. The relative roles of endothelin (ET) and vasopressin (AVP) in the regulation of blood pressure (BP), cardiac output (CO) and total peripheral resistance (TPR) were investigated in the early stages (24 - 31 days) of development of hypertension in the conscious deoxycorticosterone acetate (DOCA)-salt hypertensive rat model. 2. BP was recorded with radiotelemetry devices and CO with ultrasonic transit-time probes. TPR was calculated from the BP and CO recordings. The contributions of endogenous ET and AVP were studied by infusing [d(CH(2))(5)(1),O-Me_Tyr(2),Arg(8)]-vasopressin, a V(1)-receptor antagonist, and bosentan, a mixed ET(A)/ET(B) receptor antagonist (Study 1). Vascular responsiveness was estimated from the changes in TPR evoked by i.v. infusions of ET-1 and AVP (Study 2). 3. In study 1, infusion of bosentan reduced TPR and BP dramatically in DOCA-salt hypertensive rats but not in SHAM control rats, and this effect was greater when the AVP system had been blocked. In contrast, the V(1) receptor antagonist alone failed to change TPR and BP in DOCA-salt hypertensive rats. However, subsequent infusion of the V(1) receptor antagonist during the plateau phase of the response in bosentan pretreated DOCA-salt hypertensive rats led to significant decreases in both BP and TPR. 4. In study 2, TPR and BP responses to ET-1, but not AVP, were greater in DOCA-salt rats than in control rats. CO responses to ET-1 or AVP were similar in the two groups. 5. The results suggest that both ET and AVP play a role in the maintenance of TPR and BP; when one system is blocked the other compensates. However, the magnitude of the contribution to the hypertensive state appears greater for ET than for AVP. Enhanced vascular responses to ET appear to contribute to this greater role. PMID- 11264239 TI - Heterogeneous increases of cytoplasmic calcium: distinct effects on down regulation of cell surface sodium channels and sodium channel subunit mRNA levels. AB - 1. Long-term (> or = 12 h) treatment of cultured bovine adrenal chromaffin cells with A23187 (a Ca(2+) ionophore) or thapsigargin (TG) [an inhibitor of sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA)] caused a time- and concentration-dependent reduction of cell surface [(3)H]-saxitoxin (STX) binding capacity, but did not change the K:(D:) value. In A23187- or TG-treated cells, veratridine-induced (22)Na(+) influx was reduced (with no change in veratridine EC(50) value) while it was enhanced by alpha-scorpion venom, beta-scorpion venom, or Ptychodiscus brevis toxin-3, like in nontreated cells. 2. The A23187- or TG induced decrease of [(3)H]-STX binding was diminished by BAPTA-AM. EGTA also inhibited the decreasing effect of A23187. A23187 caused a rapid, monophasic and persistent increase in intracellular concentration of Ca(2+) ([Ca(2+)](i)) to a greater extent than that observed with TG. 2,5-Di-(t-butyl)-1,4-benzohydroquinone (DBHQ) (an inhibitor of SERCA) produced only a rapid monophasic increase in [Ca(2+)](i), without any effect on [(3)H]-STX binding. 3. Reduction in [(3)H]-STX binding capacity induced by A23187 or TG was attenuated by Go6976 (an inhibitor of conventional protein kinase C) or calpastatin peptide (an inhibitor of calpain). When the internalization rate of cell surface Na(+) channels was measured in the presence of brefeldin A (an inhibitor of vesicular exit from the trans-Golgi network), A23187 or TG accelerated the reduction of [(3)H]-STX binding capacity. 4. Six hours treatment with A23187 lowered Na(+) channel alpha- and beta(1)-subunit mRNA levels, whereas TG had no effect. 5. These results suggest that elevation of [Ca(2+)](i) caused by A23187, TG or DBHQ exerted differential effects on down-regulation of cell surface functional Na(+) channels and Na(+) channel subunit mRNA levels. PMID- 11264240 TI - Brain alpha(2)-adrenoceptors in monoamine-depleted rats: increased receptor density, G coupling proteins, receptor turnover and receptor mRNA. AB - 1. This study was designed to assess the molecular and cellular events involved in the up-regulation (and receptor supersensitivity) of brain alpha(2) adrenoceptors as a result of chronic depletion of noradrenaline (and other monoamines) by reserpine. 2. Chronic reserpine (0.25 mg kg(-1) s.c., every 48 h for 6 - 14 days) increased significantly the density (B(max) values) of cortical alpha(2)-adrenoceptor agonist sites (34 - 48% for [(3)H]-UK14304, 22 - 32% for [(3)H]-clonidine) but not that of antagonist sites (11 - 18% for [(3)H] RX821002). Competition of [(3)H]-RX821002 binding by (-)-adrenaline further indicated that chronic reserpine was associated with up-regulation of the high affinity state of alpha(2)-adrenoceptors. 3. In cortical membranes of reserpine treated rats (0.25 mg kg(-1) s.c., every 48 h for 20 days), the immunoreactivities of various G proteins (Galphai(1/2), Galphai(3), Galphao and Galphas) were increased (25 - 34%). Because the high-affinity conformation of the alpha(2)-adrenoceptor is most probably related to the complex with Galphai(2) proteins, these results suggested an increase in signal transduction through alpha(2)-adrenoceptors (and other monoamine receptors) induced by chronic reserpine. 4. After alpha(2)-adrenoceptor alkylation, the analysis of receptor recovery (B(max) for [(3)H]-UK14304) indicated that the increased density of cortical alpha(2)-adrenoceptors in reserpine-treated rats was probably due to a higher appearance rate constant of the receptor ((Delta)r=57%) and not to a decreased disappearance rate constant ((Delta)k=7%). 5. Northern- and dot-blot analyses of RNA extracted from the cerebral cortex of saline- and reserpine treated rats (0.25 mg kg(-1), s.c., every 48 h for 20 days) revealed that reserpine markedly increased the expression of alpha(2a)-adrenoceptor mRNA in the brain (125%). This transcriptional activation of the receptor gene expression appears to be the cellular mechanism by which reserpine induces up-regulation in the density of brain alpha(2)-adrenoceptors. PMID- 11264241 TI - Agonist trafficking of G(i/o)-mediated alpha(2A)-adrenoceptor responses in HEL 92.1.7 cells. AB - 1. The ability of 19 agonists to elevate Ca(2+) and inhibit forskolin-induced cyclic AMP elevation through alpha(2A)-adrenoceptors in HEL 92.1.7 cells was investigated. Ligands of catecholamine-like- (five), imidazoline- (nine) and non catecholamine-non-imidazoline-type (five) were included. 2. The relative maximum responses were similar in both assays. Five ligands were full or nearly full agonists, six produced 20 - 70% of the response to a full agonist and the remaining eight gave lower responses (< 20%) so that their potencies were difficult to evaluate. 3. Marked differences in the potencies of the agonists with respect to the two measured responses were seen. The catecholamines were several times less potent in decreasing cyclic AMP than in increasing Ca(2+), whereas the other, both imidazoline and ox-/thiazoloazepine ligands, were several times more potent with respect to the former than the latter response. For instance, UK14,304 was more potent than adrenaline with respect to the cyclic AMP response but less potent than adrenaline with respect to the Ca(2+) response. 4. All the responses were sensitive to pertussis toxin-pretreatment. Also the possible role of PLA(2), beta-adrenoceptors or ligand transport or metabolism as a source of error could be excluded. The results suggest that the active receptor states produced by catecholamines and the other agonists are markedly different and therefore have different abilities to activate different signalling pathways. PMID- 11264242 TI - Activity of mu- and delta-opioid agonists in vas deferens from mice deficient in MOR gene. AB - 1. Mice lacking the mu-opioid receptor have been recently generated. Centrally mediated responses of mu-opioid agonists are suppressed whereas some of the delta opioid responses are preserved in these mutant mice. 2. The vas deferens bioassay has been used in this study to investigate the functional activity at a peripheral level of mu- and delta-opioid agonists in mice lacking mu-opioid receptors. 3. The different mu-opioid agonists evaluated, morphine, DAMGO, dermorphin and [Lys(7)]-dermorphin produced an inhibitory response in vas deferens from wild-type mice but had no relevant activity on vas deferens from mutant mice. 4. The selective delta-opioid agonists DPDPE, BUBU, deltorphin I, deltorphin II and [D-Met(2)]-deltorphin induced inhibitory effects in vas deferens from both wild-type and mutant mice. However, the biological activities of these ligands were slightly reduced in preparations from mutant mice. The inhibitory responses of all these delta-opioid agonists were prevented by the administration of the selective delta-opioid antagonist naltrindole. 5. These data indicate that delta-opioid agonists, but not mu-opioid agonists, are biologically active in vas deferens from mice lacking mu-opioid receptors. The decreased response of delta-agonists in mutant mice suggests that some cooperativity may exist between mu- and delta-opioid receptors in these vas deferens preparations. PMID- 11264243 TI - Tedisamil and dofetilide-induced torsades de pointes, rate and potassium dependence. AB - 1. Tedisamil is a bradycardiac agent that prolongs the QT interval of the ECG and prevents cardiac arrhythmias. Given this profile, tedisamil might be expected to have proarrhythmic actions similar to Class III antiarrhythmic drugs. To address this question, the actions of dofetilide and tedisamil were examined in rabbit isolated hearts in which bradycardia was induced by AV ablation. 2. The QT interval was prolonged in a reverse rate-dependent fashion by dofetilide (3 and 30 nM) and tedisamil (0.3 and 3 microM). 3. Torsades de pointes was observed in 1/7 hearts treated with 3 nM dofetilide and 0/7 hearts treated with 0.3 microM tedisamil. The incidence of torsades de pointes was increased to 5/7 in hearts treated with 30 nM dofetilide and to 7/7 in hearts treated with 3 microM tedisamil (both P < 0.05 vs control). 4. The actions of 30 nM dofetilide and 3 microM tedisamil were also examined in hearts paced at 50, 100, 200 and 50 beats min(-1) successively. Both drugs caused torsades de pointes in 5/5 hearts paced at 50 beats min(-1); however, the incidence was reduced to 0/5 during pacing at 200 beats min(-1). Thus, drug-induced proarrhythmia was bradycardia-dependent. 5. Drug-induced prolongation of the interval between the peak and end of the T-wave (QTa-e) was reverse rate-dependent and was associated with the occurrence of torsades de pointes (r = 0.91, P < 0.01). 6. The results suggest that tedisamil, like dofetilide, presents a risk for development of torsades de pointes. PMID- 11264244 TI - Serum constituents can affect 2'-& 3'-O-(4-benzoylbenzoyl)-ATP potency at P2X(7) receptors. AB - 1. 2'-& 3'-O-(4-benzoylbenzoyl)-ATP (BzATP) is the prototypic agonist for P2X(7) receptors. In this study we demonstrate that bovine serum albumin (BSA) can affect the potency of BzATP at P2X receptors. 2. BzATP potency (pEC(50)) to stimulate ethidium accumulation in cells expressing recombinant P2X7 receptors varied between 6.5 and 4, depending upon the species orthologue studied and ionic conditions employed. BSA (0.1 - 1 mg ml(-1)) and foetal bovine serum (FBS, 1 - 10% v v(-1)) inhibited responses to BzATP but only when the BzATP pEC(50) exceeded 5. 3. BSA did not block ATP-stimulated ethidium accumulation, suggesting its effects were independent of P2X(7) receptor blockade. 4. BSA did not cause breakdown of nucleotides, although FBS (10% v v(-1)) exhibited appreciable nucleotidase activity and caused significant breakdown of ATP. 5. In the presence of BSA, lipids such as 11-((5-dimethylaminonaphthalene-1 sulphonyl)amino)undecanoic acid (DAUDA) and arachidonic acid (AA) markedly increased BzATP potency. Lipids had no affect on ATP potency in the presence of BSA and had little effect on responses to BzATP in the absence of BSA. 6. These results suggested that the reduction in BzATP potency by BSA was due to BzATP binding to BSA and that lipids prevented this binding. Consistent with this hypothesis, BzATP inhibited binding of the fluorescent lipid, DAUDA, to BSA. 7. In conclusion, BSA and lipids can markedly affect BzATP potency at P2X(7) receptors but this is probably a consequence of BzATP binding to BSA. This finding has important implications when using BzATP in vivo or in the presence of albumin. PMID- 11264245 TI - Mechanism of airway hyperresponsiveness to adenosine induced by allergen challenge in actively sensitized Brown Norway rats. AB - 1. We have explored the role of allergen sensitization and challenge in defining the response of the airways of the Brown Norway (BN) rat to adenosine. 2. In naive animals or in rats sensitized to ovalbumin (OA) adenosine induced only weak bronchoconstrictor responses. Challenge of sensitized animals with OA induced a marked airway hyperresponsiveness to adenosine which was not seen with methacholine or bradykinin. 3. The augmented bronchoconstrictor response to adenosine was not affected by acute bivagotomy or atropine nor mimicked by an i.v. injection of capsaicin. It was, however, blocked selectively by disodium cromoglycate methysergide or ketanserin and reduced in animals treated sub chronically with compound 48/80. 4. The augmented response to adenosine was associated with increases in the plasma concentrations of both histamine and 5 hydroxytryptamine (5-HT), which were attenuated by pretreatment with disodium cromoglycate, and degranulation of mast cells in the lung. 5. Parenchymal strips from lungs removed from sensitized rats challenged with OA gave augmented bronchoconstrictor responses to adenosine relative to strips from sensitized animals challenged with saline. Responses were inhibited by methysergide and disodium cromoglycate. 6. These data demonstrate a marked augmentation of the bronchoconstrictor response to adenosine in actively sensitized BN rats challenged with OA. The augmented response is primarily a consequence of mast cell activation, leading to the release of 5-HT, which in turn induces bronchoconstriction. Our data further suggest the involvement of a discrete lung based population of mast cells containing and releasing mainly 5-HT and brought into play by prior exposure to allergen. PMID- 11264246 TI - A nitric oxide-dopamine link pathway in organum vasculosum laminae terminalis of rat brain exerts control over blood pressure. AB - 1. Experiments were carried out to explore the possible role played by the nitric oxide (NO) and dopamine (DA) system in the organum vasculosum laminae terminalis (OVLT) of rat brain in arterial pressure regulation. 2. Intracerebroventricular (ICV) administration of NO donors such as hydroxylamine or sodium nitroprusside (SNP) caused an up to 59 mmHg decrease in blood pressure (BP) and a decrease in DA release (measured by nafion coated carbon fibre electrodes in combination with voltammetry) in the OVLT. In contrast, ICV administration of N(G)-nitro-L arginine methyl ester (L-NAME; a constitutive NO synthase inhibitor) or 7 nitroindazol (a neuronal NO synthase inhibitor) caused an up to 98 mmHg increase in BP and an increase in DA release in the OVLT. 3. Intra-OVLT injection of amphetamine (0.1 - 0.3 mg), SKF 38393 (a DA D(1) receptor agonist; 0.01 - 0.03 mg), or apomorphine (a DA D(2,3) receptor agonist; 0.01 - 0.03 mg) caused an increase in BP. On the other hand, intra-OVLT injection of SCH23390 (a DA D(1) receptor antagonist; 0.005 - 0.020 mg) or haloperidol (0.005 - 0.020 mg) caused a decrease in BP. 4. The pressor effects induced by intra-OVLT administration of L NAME were attenuated by pretreatment with intra-OVLT injection of haloperidol, SCF23390, or 6-hydroxydopamine. In the contrast, the hydroxylamine-, 8-Br-cGMP- or SNP-induced depressor effects were attenuated by pretreatment with intra-OVLT injection of amphetamine, SKF 38393 or apomorphine. 5. The data suggest that activation of a NO-DA link pathway within the OVLT of rat brain exerts control over blood pressure. PMID- 11264247 TI - Mitogenic effect of oxidized low-density lipoprotein on vascular smooth muscle cells mediated by activation of Ras/Raf/MEK/MAPK pathway. AB - 1. It has been demonstrated that oxidized low-density lipoprotein (OX-LDL) is a risk factor in atherosclerosis by stimulating vascular smooth muscle cell (VSMC) proliferation. However, the mechanisms of OX-LDL-induced cell proliferation are not completely understood. Therefore, we investigated the effect of OX-LDL on cell proliferation associated with mitogen-activated protein kinase (MAPK) activation in rat cultured VSMCs. 2. Both native-LDL (N-LDL) and OX-LDL induced a time- and concentration-dependent incorporation of [(3)H]-thymidine in VSMCs. 3. OX-LDL induced time- and concentration-dependent phosphorylation of p42/p44 MAPK. Pretreatment of these cells with pertussis toxin or U73122 attenuated the OX-LDL induced responses. 4. Pretreatment with PMA for 24 h, preincubation with a PKC inhibitor staurosporine or the tyrosine kinase inhibitors, genistein and herbimycin A for 1 h, substantially reduced [(3)H]-thymidine incorporation and p42/p44 MAPK phosphorylation induced by OX-LDL. 5. Removal of Ca(2+) by BAPTA/AM or depletion of the internal Ca(2+) pool by thapsigargin significantly inhibited OX-LDL-induced [(3)H]-thymidine incorporation and p42/p44 MAPK phosphorylation. 6. OX-LDL-induced [(3)H]-thymidine incorporation and p42/p44 MAPK phosphorylation was inhibited by PD98059 (an inhibitor of MEK1/2) and SB203580 (an inhibitor of p38 MAPK) in a concentration-dependent manner. 7. Overexpression of dominant negative mutants of Ras (H-Ras-15A) and Raf (Raf-N4) significantly suppressed MEK1/2 and p42/p44 MAPK activation induced by OX-LDL and PDGF-BB, indicating that Ras and Raf may be required for activation of these kinases. 8. These results suggest that the mitogenic effect of OX-LDL is mediated through a PTX-sensitive G protein-coupled receptor that involves the activation of the Ras/Raf/MEK/MAPK pathway similar to that of PDGF-BB in rat cultured VSMCs. PMID- 11264248 TI - Effects of mitiglinide (S 21403) on Kir6.2/SUR1, Kir6.2/SUR2A and Kir6.2/SUR2B types of ATP-sensitive potassium channel. AB - 1. We have investigated the mechanism of action of the novel anti-diabetic agent mitiglinide (S 21403) on Kir6.2/SUR1, Kir6.2/SUR2A and Kir6.2/SUR2B types of ATP sensitive potassium (K(ATP)) channel. These possess a common pore-forming subunit, Kir6.2, and different regulatory sulphonylurea receptor (SUR) subunits. It is believed that they correspond to native K(ATP) channels in pancreatic beta cells, heart and non-vascular smooth muscle, respectively. 2. Kir6.2 was coexpressed with SUR1, SUR2A or SUR2B in Xenopus oocytes and macroscopic currents were recorded in giant inside-out membrane patches. Mitiglinide was added to the intracellular membrane surface. 3. Mitiglinide inhibited Kir6.2/SUR currents at two sites: a low-affinity site on Kir6.2 and a high-affinity site on SUR. Low affinity inhibition was similar for all three types of K(ATP) channel but high affinity inhibition was greater for Kir6.2/SUR1 currents (IC(50), 4 nM) than for Kir6.2/SUR2A or Kir6.2/SUR2B currents (IC(50), 3 and 5 microM, respectively). 4. Inhibition of Kir6.2/SUR1 currents was only slowly reversible on the time scale of electrophysiological experiments. 5. Kir6.2/SUR1-S1237Y currents, which previously have been shown to lack high affinity tolbutamide inhibition, resembled Kir6.2/SUR2 currents in being unaffected by 100 nM but blocked by 10 microM mitiglinide. 6. Our results show that mitiglinide is a high-affinity drug that shows a 1000 fold greater affinity for the beta-cell type than the cardiac and smooth muscle types of K(ATP) channel, when measured in excised patches. PMID- 11264250 TI - Endothelium-dependent vasorelaxation independent of nitric oxide and K(+) release in isolated renal arteries of rats. AB - 1. We investigated whether K(+) can act as an endothelium-derived hyperpolarizing factor (EDHF) in isolated small renal arteries of Wistar-Kyoto rats. 2. Acetylcholine (0.001 - 3 microM) caused relaxations that were abolished by removal of the endothelium. However, acetylcholine-induced relaxations were not affected by the nitric oxide (NO) synthase inhibitor N:(omega)-nitro-L-arginine methyl ester (L-NAME, 100 microM), by L-NAME plus the soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ, 1 microM) or by L NAME plus the cyclo-oxygenase inhibitor indomethacin (10 microM). In rings precontracted with high-K(+)(60 mM) physiological salt solution in the presence of L-NAME, acetylcholine-induced relaxations were abolished. 3. L-NAME-resistant relaxations were abolished by the large-conductance Ca(2+)-activated K(+) channel inhibitor charybdotoxin plus the small-conductance Ca(2+)-activated K(+) channel inhibitor apamin, while the inward rectifier K(+) channel inhibitor Ba(2+) or the gap junction inhibitor 18alpha-glycyrrhetinic acid had no effect. Acetylcholine induced relaxation was unchanged by ouabain (10 microM) but was partially inhibited by a higher concentration (100 microM). 4. In half of the tissues tested, K(+)(10 mM) itself produced L-NAME-resistant relaxations that were blocked by ouabain (10 microM) and partially reduced by charybdotoxin plus apamin, but not affected by 18alpha-glycyrrhetinic acid or Ba(2+). However, K(+) did not induce relaxations in endothelium-denuded tissues. 5. In conclusion, acetylcholine-induced relaxations in this tissue are largely dependent upon hyperpolarization mechanisms that are initiated in the endothelium but do not depend upon NO release. K(+) release cannot account for endothelium-dependent relaxation and cannot be an EDHF in this artery. However, K(+) itself can initiate endothelium-dependent relaxations via a different pathway from acetylcholine, but the mechanisms of K(+)-induced relaxations remain to be clarified. PMID- 11264249 TI - Tonabersat (SB-220453) a novel benzopyran with anticonvulsant properties attenuates trigeminal nerve-induced neurovascular reflexes. AB - 1. The effects of tonabersat (SB-220453) were evaluated on trigeminal nerve ganglion stimulation-induced sensory-autonomic neurovascular reflexes in the anaesthetized cat. Comparisons were made to intravenous administration of carabersat (SB-204269), and to valproate, gabapentin and lamotrigine following intraduodenal administration. 2. There were no effects on resting blood pressure, heart rate, carotid blood flow or carotid vascular resistance for any compound evaluated. 3. Trigeminal nerve ganglion stimulation increased carotid blood flow by 65% and reduced vascular resistance by 41% with minimal effect on blood pressure (< 10%) and no effect on heart rate. Intravenous infusion of tonabersat or carabersat (both 3.4 micromol h(-1)) produced time related reductions in stimulation-induced responses with a maximal inhibition (relative to control) of 30 +/- 7% (n=4), at 240 min for tonabersat and 33+/-4% (n=3) at 180 min for carabersat. Tonabersat (11.5 micromol h(-1)) produced a similar inhibitory effect (32 +/- 9%, n=4) after 120 min of infusion. 4. Following intraduodenal administration of tonabersat, the maximal inhibition of nerve stimulation-induced responses was 55 +/- 4% at 120 min (n=4) for tonabersat 10 mg kg(-1), and 24+/-2% after 180 min for 1 mg kg(-1) (n=4). 5. Intraduodenal administration of sodium valproate (10 or 100 mg kg(-1) n=4/group) had no effect on neurovascular reflexes. Maximal inhibition of nerve ganglion-stimulated reductions in carotid vascular resistance were observed at 150 min for lamotrigine (50 mg kg(-1), 52+/ 12%, n=4) and gabapentin (100 mg kg(-1), 17+/-13%, n=3). Lamotrigine 10 mg kg(-1) produced 22+/-11% (n=3) inhibition after 180 min. 6. These data demonstrate blockade of trigeminal parasympathetic reflexes with tonabersat, carabersat and other anticonvulsants. These agents may therefore have therapeutic benefit in conditions where this type of reflex is evident. PMID- 11264251 TI - Effects of specific inhibition of cyclo-oxygenase-1 and cyclo-oxygenase-2 in the rat stomach with normal mucosa and after acid challenge. AB - 1. Effects of the cyclo-oxygenase (COX)-1 inhibitor SC-560 and the COX-2 inhibitors rofecoxib and DFU were investigated in the normal stomach and after acid challenge. 2. In healthy rats, neither SC-560 nor rofecoxib (20 mg kg(-1) each) given alone damaged the mucosa. Co-treatment with SC-560 and rofecoxib, however, induced severe lesions comparable to indomethacin (20 mg kg(-1)) whereas co-administration of SC-560 and DFU (20 mg kg(-1) each) had no comparable ulcerogenic effect 5 h after dosing. 3. SC-560 (20 mg kg(-1)) inhibited gastric 6 keto-prostaglandin (PG) F(1alpha) by 86+/-5% and platelet thromboxane (TX) B(2) formation by 89+/-4% comparable to indomethacin (20 mg kg(-1)). Rofecoxib (20 mg kg(-1)) did not inhibit gastric and platelet eicosanoids. 4. Intragastric HCl elevated mucosal mRNA levels of COX-2 but not COX-1. Dexamethasone (2 mg kg(-1)) prevented the up-regulation of COX-2. 5. After acid challenge, SC-560 (5 and 20 mg kg(-1)) induced dose-dependent injury. Rofecoxib (20 mg kg(-1)), DFU (5 mg kg( 1)) and dexamethasone (2 mg kg(-1)) given alone were not ulcerogenic but aggravated SC-560-induced damage. DFU augmented SC-560 damage 1 but not 5 h after administration whereas rofecoxib increased injury after both treatment periods suggesting different time courses. 6. Gastric injurious effects of rofecoxib and DFU correlated with inhibition of inflammatory PGE(2). 7. The findings show that in the normal stomach lesions only develop when both COX-1 and COX-2 are inhibited. In contrast, during acid challenge inhibition of COX-1 renders the mucosa more vulnerable suggesting an important role of COX-1 in mucosal defence in the presence of a potentially noxious agent. In this function COX-1 is supported by COX-2. In the face of pending injury, however, COX-2 cannot maintain mucosal integrity when the activity of COX-1 is suppressed. PMID- 11264252 TI - The effect of SB-269970, a 5-HT(7) receptor antagonist, on 5-HT release from serotonergic terminals and cell bodies. AB - 1. The presence of 5-HT(7) receptor mRNA and protein in 5-HT neurons suggests that this receptor may act as a 5-HT autoreceptor. In this study, the effect of the 5-HT(7) receptor antagonist, SB-269970 ((R)-1-[3-hydroxy phenyl)sulfonyl]-2 [2-(4-methyl-1-piperidinyl)ethyl]pyrrolidine), was investigated on 5-HT release in the guinea-pig and rat cortex and the rat dorsal raphe nucleus (DRN), using the techniques of in vitro [(3)H]-5-HT release or fast cyclic voltammetry, respectively. 2. Cortical slices were loaded with [(3)H]-5-HT and release was evoked by electrical stimulation. 5-CT inhibited the evoked release of [(3)H]-5 HT in a concentration-dependent manner. SB-269970 had no significant effect on [(3)H]-5-HT release while the 5-HT(1B) receptor antagonist, SB-224289 significantly potentiated [(3)H]-5-HT release. In addition, SB-269970 was unable to attenuate the 5-CT-induced inhibition of release while SB-224289 produced a rightward shift of the 5-CT response, generating estimated pK(B) values of 7.8 and 7.6 at the guinea-pig and rat terminal 5-HT autoreceptors respectively. 3. Rat DRN slices were electrically stimulated and the evoked 5-HT efflux detected by voltammetric analysis. 8-OH-DPAT inhibited evoked 5-HT efflux and was fully reversed by WAY 100635. SB-269970 had no effect on either 5-HT efflux per se or 8 OH-DPAT-induced inhibition of 5-HT efflux. In addition, 5-CT inhibited 5-HT efflux in a concentration-dependent manner. SB-269970 was unable to attenuate the 5-CT-induced inhibition of 5-HT efflux. 4. In conclusion, we were unable to provide evidence to suggest a 5-HT autoreceptor role for 5-HT(7) receptors. However, investigations with more selective 5-HT(7) receptor agonists are needed to confirm the data reported here. PMID- 11264253 TI - Cyclo-oxygenase and lipoxygenase pathways in mast cell dependent-neurogenic inflammation induced by electrical stimulation of the rat saphenous nerve. AB - 1. We investigated the role of arachidonic acid metabolism and assessed the participation of mast cells and leukocytes in neurogenic inflammation in rat paw skin. We compared the effect of lipoxygenase (LOX) and cyclo-oxygenase (COX) inhibitors on oedema induced by saphenous nerve stimulation, substance P (SP), and compound 48/80. 2. Intravenous (i.v.) pre-treatment with a dual COX/LOX inhibitor (RWJ 63556), a dual LOX inhibitor/cysteinyl-leukotriene (CysLt) receptor antagonist (Rev 5901), a LOX inhibitor (AA 861), a five-lipoxygenase activating factor (FLAP) inhibitor (MK 886), or a glutathione S-transferase inhibitor (ethacrynic acid) significantly inhibited (40 to 60%) the development of neurogenic oedema, but did not affect cutaneous blood flow. Intradermal (i.d.) injection of LOX inhibitors reduced SP-induced oedema (up to 50% for RWJ 63556 and MK 886), whereas ethacrynic acid had a potentiating effect. 3. Indomethacin and rofecoxib, a highly selective COX-2 inhibitor, did not affect neurogenic and SP-induced oedema. Surprisingly, the structurally related COX-2 inhibitors, NS 398 and nimesulide, significantly reduced both neurogenic and SP-induced oedema (70% and 42% for neurogenic oedema, respectively; 49% and 46% for SP-induced oedema, respectively). 4. COX-2 mRNA was undetectable in saphenous nerves and paw skin biopsy samples, before and after saphenous nerve stimulation. 5. A mast cell stabilizer, cromolyn, and a H(1) receptor antagonist, mepyramine, significantly inhibited neurogenic (51% and 43%, respectively) and SP-induced oedema (67% and 63%, respectively). 6. The co-injection of LOX inhibitors and compound 48/80 did not alter the effects of compound 48/80. Conversely, ethacrynic acid had a significant potentiating effect. The pharmacological profile of the effect of COX inhibitors on compound 48/80-induced oedema was similar to that of neurogenic and SP-induced oedema. 7. The polysaccharide, fucoidan (an inhibitor of leukocyte rolling) did not affect neurogenic or SP-induced oedema. 8. Thus, (i) SP-induced leukotriene synthesis is involved in the development of neurogenic oedema in rat paw skin; (ii) this leukotriene-mediated plasma extravasation might be independent of mast cell activation and/or of the adhesion of leukocytes to the endothelium; (iii) COX did not appear to play a significant role in this process. PMID- 11264254 TI - Intracellular Angiotensin II and cell growth of vascular smooth muscle cells. AB - 1. We recently demonstrated that intracellular application of Angiotensin II (Angiotensin II(intr)) induces rat aorta contraction independent of plasma membrane Angiotensin II receptors. In this study we investigated the effects of Angiotensin II(intr) on cell growth in A7r5 smooth muscle cells. 2. DNA-synthesis was increased dose-dependently by liposomes filled with Angiotensin II as measured by [(3)H]-thymidine incorporation at high (EC(50)=27+/-6 pM) and low (EC(50)=14+/-5 nM) affinity binding sites with increases in E(max) of 58+/-4 and 37+/-4% above quiescent cells, respectively. Cell growth was corroborated by an increase in cell number. 3. Extracellular Angiotensin II (10 pM - 10 microM) did not modify [(3)H]-thymidine incorporation. 4. Growth effects of Angiotensin II(intr) mediated via high affinity sites were inhibited by liposomes filled with 1 microM of the non-peptidergic antagonists losartan (AT(1)-receptor) or PD123319 (AT(2)-receptor) or with the peptidergic agonist CGP42112A (AT(2)-receptor). E(max) values were decreased to 30+/-3, 29+/-4 and 4+/-2%, respectively, without changes in EC(50). The Angiotensin II(intr) effect via low affinity sites was only antagonized by CGP42112A (E(max)=11+/-3%), while losartan and PD123319 increased E(max) to 69+/-4%. Intracellular applications were ineffective in the absence of Angiotensin II(intr). 5. Neither intracellular nor extracellular Angiotensin I (1 microM) were effective. 6. The Angiotensin II(intr) induced growth response was blocked by selective inhibition of phosphatidyl inositol 3 kinase (PI-3K) by wortmannin (1 microM) and of the mitogen-activated protein kinase (MAPK/ERK) pathway by PD98059 (1 microM) to 61+/-14 and 4+/-8% of control, respectively. 7. These data demonstrate that Angiotensin II(intr) induces cell growth through atypical AT-receptors via a PI-3K and MAPK/ERK -sensitive pathway. PMID- 11264255 TI - Inhibition of the cyclin D1/E2F pathway by PCA-4230, a potent repressor of cellular proliferation. AB - 1. Tight control of cellular growth is essential to ensure normal tissue patterning and prevent pathological responses. Excessive vascular smooth muscle cell (VSMC) proliferation is associated with the pathophysiology of atherosclerosis and restenosis post-angioplasty. Thus, drug targeting of pathological VSMC growth may be a suitable therapeutic intervention in vascular proliferative diseases. 2. In the present study, we investigated the mechanisms underlying VSMC growth arrest induced by the pharmacological agent PCA-4230. Addition of PCA-4230 to cultured VSMCs blocked the induction of cyclin D1 and cyclin A expression normally seen in serum-restimulated cells. Moreover, PCA-4230 inhibited cyclin-dependent kinase 2 (CDK2) activity and abrogated hyperphosphorylation of the retinoblastoma (Rb) gene product. Similarly, PCA-4230 dependent growth arrest of transformed cell lines correlated with reduced level of cyclin D1 protein and inhibition of CDK2 activity. Consistent with these findings, PCA-4230 repressed serum-inducible cyclin A promoter activity, and overexpression of either cyclin D1 or E2F1 efficiently circumvented this inhibitory effect. Importantly, adenovirus-mediated overexpression of E2F1 restored S-phase entry in PCA-4230-treated VSMCs, demonstrating that PCA-4230 represses cyclin A gene expression and VSMC growth via inhibition of the cyclin D1/E2F pathway. 3. Because of its ability to inhibit the growth of human VSMCs and transformed cell lines, future studies are warranted to assess whether PCA 4230 may be a suitable therapeutic intervention for the treatment of hyperproliferative disorders, including cardiovascular disease and cancer. PMID- 11264256 TI - Functional characterization of rat submaxillary gland muscarinic receptors using microphysiometry. AB - 1. Muscarinic cholinoceptors (MChR) in freshly dispersed rat salivary gland (RSG) cells were characterized using microphysiometry to measure changes in acidification rates. Several non-selective and selective muscarinic antagonists were used to elucidate the nature of the subtypes mediating the response to carbachol. 2. The effects of carbachol (pEC(50) = 5.74 +/- 0.02 s.e.mean; n = 53) were highly reproducible and most antagonists acted in a surmountable, reversible fashion. The following antagonist rank order, with apparent affinity constants in parentheses, was noted: 4-DAMP (8.9)= atropine (8.9) > tolterodine (8.5) > oxybutynin (7.9) > S-secoverine (7.2) > pirenzepine (6.9) > himbacine (6.8) > AQ RA 741 (6.6) > methoctramine (5.9). 3. These studies validate the use of primary isolated RSG cells in microphysiometry for pharmacological analysis. These data are consistent with, and extend, previous studies using alternative functional methods, which reported a lack of differential receptor pharmacology between bladder and salivary gland tissue. 4. The antagonist affinity profile significantly correlated with the profile at human recombinant muscarinic M(3) and M(5) receptors. Given a lack of antagonists that discriminate between M(3) and M(5), definitive conclusion of which subtype(s) is present within RSG cells cannot be determined. PMID- 11264259 TI - Coping strategies in the self-management of chronic heart failure. AB - BACKGROUND: A patient's psychological adaptation to heart failure can influence its impact on his or her life. However, attempts to understand how patients cope mentally with severe emotional strain have led to inconsistent use of a plethora of concepts, making communication and clinical care difficult. OBJECTIVES: The aim of the present study was to develop a framework for conceptualizing how patients with chronic heart failure cope mentally with their illness, and then use the framework to suggest how GPs can facilitate patient self-care. METHODS: We systematically reduced and reassembled the narrative texts of personal, semi structured interviews until their interpretation was complete. The interviews were conducted during late 1999 with 62 heart failure patients under GP care in 30 practices across central Auckland, New Zealand. RESULTS: Our framework describes four coping strategies: avoidance, disavowal, denial and acceptance. Disavowal provides a distinct coping strategy through which patients, who basically understand the threat to their life situations, seek hope through positively reconstructing this threat. Use of this strategy was highly salient regardless of patients' age, the length of time since their recorded diagnosis or the degree of self-reported limitation of recent physical function due to heart failure. Only over age 70 were avoidance and acceptance also highly salient among patients whose heart failure was diagnosed at least 3 years previously and had mildly limited their recent physical function. CONCLUSION: Many different heart failure patients use disavowal to palliate the emotional strain and find hope. Disavowal is not a problem to deal with but a process GPs can facilitate by implementing a range of suggested strategies through methods such as story telling. PMID- 11264257 TI - The effect of ABT-702, a novel adenosine kinase inhibitor, on the responses of spinal neurones following carrageenan inflammation and peripheral nerve injury. AB - 1. Adenosine (ADO) receptor activation modulates sensory transmission in the dorsal horn. Little is known about the circumstances underlying release of the purine. The present study was conducted to investigate the effect of a novel and potent non-nucleoside adenosine kinase (AK) inhibitor, ABT-702, on the responses of dorsal horn neurones to selected peripheral stimuli. ABT-702 is orally effective to reduce behavioural signs of nociception in models of acute, inflammatory, and neuropathic pain. 2. Electrophysiological recordings were made from wide dynamic range (WDR) neurones in halothane-anaesthetized rats. ABT-702 was given subcutaneously following either carrageenan inflammation or peripheral nerve injury (L5/L6 spinal nerve ligation). Comparisons were made between carrageenan and uninjected control animals, and similarly between spinal nerve ligated (SNL) and sham operated animals. 3. ABT-702 produced inhibition of the postdischarge, wind-up and C-fibre evoked responses in both carrageenan and nerve injured animals. Furthermore, the mechanical and thermal evoked responses were similarly reduced in SNL rats. Overall, ABT-702 produced a significantly greater inhibition of these responses in SNL rats as compared to sham controls. Similarly ABT-702 tended to produce greater effects after carrageenan inflammation, however this did not reach significance. 4. Protection of endogenous adenosine by ABT-702 therefore produces a marked inhibition of the noxious evoked neuronal activity in inflamed and neuropathic rats. Our results demonstrate a plasticity in the endogenous adenosine-mediated inhibitory system following SNL and provide a possible basis for the use of this compound for the treatment of neuropathic and other persistent pain states. PMID- 11264260 TI - Supporting GPs whose performance gives cause for concern: the North Trent experience. AB - BACKGROUND: The North Trent scheme to address the problem of GPs whose performance gives cause for concern was implemented in 1997. This paper describes the structure and process of the scheme and evaluates the main outcomes. METHODS: We used non-participant observation and semi-structured interviews with representatives of the seven Health Authorities (HAs) of North Trent including medical and prescribing advisors and senior primary care managers. Twenty-one GPs who were members of the Performance Review Quartets (PRQs) were also interviewed. Qualitative data were analysed using a constant comparative method to identify emergent themes. RESULTS: Performance indicators were agreed between HAs and the profession in the seven North Trent localities. The scheme identified 18 GPs whose performance gave cause for concern, of whom 15 GPs in six practices received a formal visit. Educational plans were agreed and implemented with three GPs. The remaining 12 received administrative and clinical support. Three of the 18 GPs initially refused to co-operate with the scheme. Two of these have since agreed a practice visit following a visit by a senior local medical committee representative. The performance indicators used in the scheme have not been specific to individual GPs except those in single-handed practices. Some indicators used by PRQs related to cost effectiveness rather than quality of care for individual patients. Current resources were adequate for the small number of underperforming GPs identified by the scheme. CONCLUSIONS: The North Trent scheme has identified a number of underperforming GPs, 83% of whom have been willing to participate in a supportive intervention. The scheme will need some modification with the advent of primary care trusts and the proposed assessment and support centres. PMID- 11264261 TI - A study of GP referrals to a family cancer clinic for breast/ovarian cancer. AB - OBJECTIVES: The aim of this study was to investigate the appropriateness of primary care referrals to the Oxford Regional Genetics Service on account of a family history of breast and/or ovarian cancer and to explore GPs' expectations following a referral. METHODS: Fifty consecutive GP referrers were sent a questionnaire post-referral, and their referral letters were reviewed. RESULTS: The study achieved a high response rate (94%) and showed that many GPs did not know which patients warrant referral to the genetics service and that they had unrealistic expectations of what happens at the clinic. CONCLUSIONS: If GPs are to fulfil their gatekeeper role effectively, and possibly become more involved in the delivery of genetic services in the future, it is clear that they require further education in this area. PMID- 11264262 TI - Guidelines for referral to a regional genetics service: GPs respond by referring more appropriate cases. AB - OBJECTIVES: The aim of this study was to see whether guidelines on whom to refer to a regional genetics service could improve the appropriateness of referrals to the service. It also aimed to assess whether the genetic clinic assessment of risk agreed with that described in the GP letter. METHODS: Referral guidelines were sent to all Oxfordshire GPs and the subsequent content of the referral letters was analysed. A retrospective assessment of referral letters sent during the 6 months before dissemination was also made. RESULTS: The study showed that post-guidelines, fewer "lower risk" referrals were made and that the description of the risk in the GP letter improved, so that a greater proportion of genetic clinic risks agreed with those described in the GP letter. CONCLUSION: The use of referral guidelines can help GPs to act as gatekeeper for referrals to secondary care. PMID- 11264263 TI - An inner city GP unit versus conventional care for elderly patients: prospective comparison of health functioning, use of services and patient satisfaction. AB - BACKGROUND: GP units are generally nurse-led wards, where GPs have direct admitting rights and retain clinical responsibility for their patients. While GP led wards are not new, they are relatively uncommon in urban areas. In addition, there has been little comparative evaluation of this type of service. OBJECTIVES: The aim of the present study was to compare patients admitted to an inner city GP unit with comparable patients in conventional care (e.g. district nursing, nursing/residential homes, acute care of the elderly wards) in terms of mental and physical functioning, use of health and social services and patient satisfaction. METHODS: Study group patients were those admitted to the GP unit; comparison group patients were identified by GP practices or conventional services who had agreed to participate in the study. Suitable patients were aged 65 years or over and fitted the eligibility criteria for the GP unit. Patients were interviewed at three time points: admission to either the GP unit or conventional care, and at 1 and 3 months after admission. Baseline comparability was assessed by demographic and medical data, cognitive function, mental state, social support, use of health and social services, and mental and physical functioning (SF-12). Mental and physical functioning and use of health and social services were compared between the groups over time. Patient satisfaction with their care was also compared between groups. RESULTS: Change in the mental and physical functioning between patients on the GP unit (n = 67) and those in conventional care (n = 60) did not differ when the groups were compared at any of the three time points. However, the mental function of patients in the GP unit significantly improved between admission and 1 month after admission (P: < 0.05). This effect was not sustained at 3 months after admission. GP unit patients were consistently more positive about the care they received than patients receiving conventional care; this included communication and information, staff, care and the facilities. Both groups of patients were high users of health and social services, with similar patterns of use in both groups, which did not alter over time. CONCLUSIONS: Patients who received care on the GP unit experienced a similar physical outcome to patients in conventional settings; however, they appeared to enjoy a short-term improvement in mental functioning and were consistently more positive about the quality of their care. This study has important policy implications with regard to planning future intermediate care services and will be of particular interest to health service planners and those responsible for clinical governance. PMID- 11264264 TI - Patient-perceived benefits of and barriers to using out-of-hours primary care centres. AB - BACKGROUND: The rapid growth of GP co-operatives has encouraged the development of primary care centres, but little is known about patients' views and experiences of these new forms of out-of-hours service delivery. OBJECTIVES: This study was designed to understand patients' views, expectations and experiences of attending an out-of-hours primary care centre which was part of an inner London GP co-operative. METHODS: Systematic samples of patients using the out-of-hours service received semi-structured interviews covering the decision to contact the service, expectations and experience of the service and, if relevant, the experience of travelling to the primary care centre. Interviews were conducted by telephone between 7 and 10 days after patient contact. RESULTS: Interviews were completed with 55.4% (72/130) of sampled patients who were primary care centre attenders, 50.0% (47/94) of those receiving telephone advice and 45.3% (53/117) of those receiving a home visit. Most attenders of the primary care centre said that they were satisfied with the consultation (90.0%, 65) and were able to get all the help they needed (83%, 60). The speed of being seen and the opportunity of having a face-to-face consultation were key benefits identified. For some, this outweighed difficulties experienced in attending the centre, including arranging transport, caring for other children, managing several children on the journey and travelling while ill. The main barriers patients identified for not wanting to attend the primary care centre included feeling too ill to travel, having other dependants to care for or lacking transportation. CONCLUSIONS: While primary care centres offer patients speedy access to face-to-face consultations, there are a range of obstacles which are encountered. Those who are socially disadvantaged appear likely to experience greatest difficulty, raising concerns about equity in access to services. Out-of-hours services may need to give consideration to patient transport and a more flexible approach to visiting at home if such inequities are to be avoided. PMID- 11264265 TI - Patients' experiences of receiving telephone advice from a GP co-operative. AB - BACKGROUND: The use of the telephone to deliver health care advice has increased considerably in recent years. Little research has been carried out to explore the experience of patients who receive such advice and its acceptability. OBJECTIVES: The aim of this study is to describe the expectations of patients, or third party callers, who had contacted a GP out-of-hours co-operative and their satisfaction with telephone advice received. METHODS: Semi-structured interviews were conducted by telephone 7-10 days after contact with one inner city GP co operative. RESULTS: A total of 47 telephone consultations were followed up with an interview. Of these, 23 (48.9%) callers had expected to be offered a home visit when they called. Reasons for wanting a home visit were either to do with the nature of the condition and its perceived severity, problems in being able to attend the primary care centre and the risks of travel, or because of problems in communicating over the telephone. Satisfaction with telephone consultations centred mostly on the doctor being able to provide reassurance and give adequate time to allay concerns. The most common reasons given for dissatisfaction were the caller feeling that the doctor could not make a correct diagnosis without having seen the patient, or the caller being made to feel that they were wasting the doctor's time. Many patients were anxious about their ability to describe symptoms over the telephone, or understand and follow the advice that they received. CONCLUSIONS: There appears to be a need for patients to be better informed about the service they can expect to receive from GP co-operatives. Recent developments such as NHS Direct may have an influence on the telephone consultation rate to GP co-operatives. PMID- 11264266 TI - Contradictions in the medical encounter: female sexual dysfunction in primary care contacts. AB - BACKGROUND: Over the past two decades, primary care physicians have been encouraged to participate in the management of sexual disturbances. Women with type 2 diabetes, often treated by GPs, are at high risk of experiencing sexual dysfunction. OBJECTIVE: Very few qualitative studies have described the impact of sexual dysfunction on the diabetic women experiencing it. Our aim was, therefore, to explore the effects, if any, of type 2 diabetes on "womanhood and intimacy" and investigate whether women wish to receive medical attention for their sexual disturbances. METHODS: We used a purposeful sample of middle-aged and older women (44-80 years) diagnosed with type 2 diabetes (n = 33). Methods triangulation was employed: focus group interviews were combined with observer data and a structured, anonymous questionnaire. We performed content analysis, with co researcher control for systematic bias during the coding process. RESULTS: Personal characteristics, such as age, sex, experience and attitude of the doctor, the specialty considered to be appropriate (GP versus gynaecologist) and circumstances (time and privacy) in the primary care setting appeared to significantly influence women's willingness to discuss--if at all--sexual matters with physicians. CONCLUSION: GPs should aim to create an open atmosphere to encourage discussion of female sexual dysfunction in the consultation room. However, women with sexual problems might benefit more from peer help through patient or women's organizations. The role of GPs might therefore consist of supporting these services and identifying female sexual dysfunction in type 2 diabetes, a problem that middle-aged and older women have difficulty communicating. PMID- 11264267 TI - Prescribing rates for psychotropic medication amongst east London general practices: low rates where Asian populations are greatest. AB - OBJECTIVES: The aim of this study was to examine the contribution of Asian ethnicity to the variation in rates of practice prescribing for antidepressant and anxiolytic medication, taking into account other population and practice organizational factors. METHODS: A practice-based cross-sectional survey was carried out of the prescribing of antidepressants and anxiolytics (daily defined dosages) in 164 general practices. The study was set in East London and the City Health Authority, which includes the multiethnic inner London boroughs of Hackney, Tower Hamlets, Newham and the City of London. The main outcome measures were the annual prescribing rates for each group of drugs, calculated as the total annual daily defined dosages divided by the practice population, and the ratio of antidepressant/ anxiolytic annual prescribing rates. RESULTS: Prescribing rates for antidepressants showed a 25-fold variation between practices; this was greater for anxiolytics. The median annual prescribing rate for all antidepressants combined was 4.13 (interquartile range 2.50-5.88). For all anxiolytics and hypnotics combined the median annual prescribing rate was 3.55 (interquartile range 1.71-6.36). Univariate analysis showed that Asian ethnicity alone accounted for 28% of the variation in antidepressant prescribing and 20.5% of the variation in the anxiolytic prescribing. A backwards multiple regression model using 10 explanatory practice and population variables accounted for 47.7% of the variance in antidepressant prescribing and 34% of the variance in the anxiolytic prescribing. CONCLUSION: In practices where the proportion of Asian patients is high, both antidepressant and anxiolytic prescribing is low. This is important for understanding interpractice prescribing variation and for setting levels of drug budgets. This study confirms that the low rates of non psychotic disorders presented by Asian populations is not a selective feature of access to secondary care, but is evident in the prescribing behaviour of GPs. Uncertainty remains as to how much this is due to a lower prevalence rate, "culture-bound syndromes" or practical difficulties in diagnosis and management within the general practice setting. PMID- 11264268 TI - Health from the patient's point of view. How does it relate to the physician's judgement? AB - OBJECTIVES: The purpose of this study was to evaluate the relationship of self rated health to a measure of physical status, based on a professional rating of the individual's health from a strictly physical point of view. METHODS: A random selection of 407 people over the age of 20 from the north-western catchment area of greater Stockholm were invited in 1995 to a physical examination, including a self-report questionnaire with questions about self-rated health, lifestyle, psychosocial factors and quality of life. A measure of physical health on a 5 point graded scale was constructed using the information from the records of the physical examination as a base. RESULTS: Self-rated health and the professional ratings of health coincided in approximately 60% of the cases. There were a relatively large number of cases where the ratings were contradictory. The correlation between the scales was 0.45. Comparison between the two ratings with respect to association with potential determinants showed that physical factors naturally explained most of the variances in physical health, whereas social and mental well-being and somatic conditions (women) were the most important explanatory variables for self-rated health. Irrespective of whether they had "favourable" or "unfavourable" health, those with "poor" self-rated health also had perceived lower social and mental well-being, less appreciation, more somatic conditions (women) and worse coping abilities (men). CONCLUSIONS: With mental, psychosocial and social problems becoming more pronounced in sickness patterns for primary care patients, self-rated health could be a helpful device, especially when time resources for consultations are short. This measure could also give a more global view of the patient's situation when effectivity and rationality can be a threat to a holistic view of the patient. PMID- 11264269 TI - Choosing not to immunize: are parents making informed decisions? AB - BACKGROUND: Childhood immunization is an important aspect of childhood preventive health, world wide, with programmes such as the Expanded Programme on Immunization organized by the World Health Organization. Unlike other countries, the immunization programme is not compulsory in the UK, and the decision whether to immunize a child or not is parental. OBJECTIVE: The objective of this study was to explore the decision-making process of parents who have chosen not to have their children immunized. METHODS: This was a qualitative study, using semi structured interviews with parents either in their own homes or at their places of work. The study was set in an inner city area with a high level of deprivation. The district immunization co-ordinator and health visitors within the area referred parents to the researcher. Parents subsequently were selected using purposive maximum variation sampling. Data were analysed using consistent and systematic review. An initial coding frame that was derived from the first few transcripts was revised and developed through its application to subsequent transcripts. The final stage of analysis involved comparing the data using the revised coding frame for drawing conclusions and verification. RESULTS: Interviews were completed with 13 parents. Parents discussed their perceptions of childhood diseases and immunization, and the risk-benefit analysis that occurred between the two. All parents identified the risk of side effects as a reason for choosing not to immunize. A proposed model of the decision-making process that represented the experiences of the parents in this study is presented. In response to the question of immunization, three actions were described by parents: a routine response, an emotional response and delaying the decision by entering a questioning stage followed by a cyclical process of seeking and evaluating information. Key to this model was a stage of reflection that most parents described irrespective of their initial action in response to the question of immunization. Parents also discussed their responsibilities in terms of the consequences of their decisions. Health professionals were perceived as providing unbalanced information that was an obstacle in decision making. CONCLUSION: The parents included in this study had chosen not to immunize at least one of their children. Most parents felt they had made an informed decision, based on an assessment of the risks and benefits of immunization and an acceptance of responsibility for that decision. Health professionals were not perceived as providers of balanced information. It is therefore important that parents have easy access to accurate information concerning the pros and cons of treatment, and have the opportunity to discuss their concerns with health professionals. PMID- 11264270 TI - Climacteric complaints in the community. AB - BACKGROUND: At the onset of the climacteric, healthy middle-aged women present with a variety of complaints, especially in general practice. In these first years of entering the menopause, vaginal blood loss alters from irregular periods to complete amenorrhoea. According to these different menstrual patterns, we can distinguish a pre-, peri- and postmenopausal phase. It could be useful to know whether specific climacteric complaints are related to these different phases. OBJECTIVE: The aim of this study was to investigate the relationship between climacteric complaints and the menstrual pattern during the menopausal transition in a population-based cross-sectional survey of healthy middle-aged women. METHODS: All women aged 47-54 years, living in the city of Eindhoven, were invited to participate in the Eindhoven Osteoporosis Study (EPOS); 6648 (78%) agreed to participate. All women completed a questionnaire concerning climacteric complaints. Climacteric status was defined by menstrual history. Odds ratios (ORs) were obtained for the relationship between climacteric status and climacteric complaints. Multiple logistic regression analysis was carried out, with climacteric status as the dependent variable. RESULTS: Of the 27 items in the questionnaire concerning climacteric complaints, seven were significantly different between all three climacteric phases (P: < 0.1). After multiple logistic regression analysis, comparing peri- and premenopause, only flushing (OR 5.9) was significantly different. Between post- and perimenopause, seven symptoms appeared to be different: three urogenital complaints [vaginal dryness (OR 1.6), vaginal discharge (OR 0.4) and pain during intercourse (OR 1.9)], three vasomotor symptoms [daytime sweating (OR 1.4), night-time sweating (OR 0.7) and flushing (OR 1.9)] and, finally, insomnia (OR 1.3). When comparing post- and premenopause, flushing (OR 13.4), insomnia (OR 2.1) and depressed mood (OR 0.6) were significantly different, in addition to three urogenital symptoms: vaginal dryness (OR 2.6), vaginal discharge (OR 0.3) and pain during intercourse (OR 2.1). CONCLUSION: The major findings of the study are that flushing is strongly associated with the transition from pre- to perimenopause, while urogenital complaints, daytime sweating and insomnia are more prominent in the transition from peri- to postmenopause. PMID- 11264271 TI - The management of thyroid disease by GPs. AB - BACKGROUND: Good medical practice depends on a collaborative relationship between a GP and a targeted specialist. OBJECTIVE: The aim of the present study was to assess knowledge and management by GPs of common endocrine disorders such as thyroid diseases. METHODS: We submitted to all the GPs (622) of the Province of Lecce an anonymous questionnaire with 11 questions which aimed to evaluate methods of approach to (questions 1 and 2) and knowledge about (questions 3-11) thyroid diseases. RESULTS: (i) Most GPs (72.1%) evaluate thyroid function on the basis of a clinical suspicion and perform preliminary investigations before referring the patient to a specialist. (ii) The ratio between right and wrong answers was significantly higher for four questions, significantly lower for one question and distributed by chance for four questions. (iii) The degree of knowledge strictly corresponds to the GP's attitude to patient's management. CONCLUSIONS: For a thyropathic patient to be diagnosed rapidly and treated efficiently, it is necessary to disseminate knowledge of standardized protocols to ensure a better utilization by both the GP and the endocrinologist of their respective competences. PMID- 11264272 TI - Turkish women's knowledge of osteoporosis. AB - BACKGROUND: Preventive measures including patient education can reduce hip fractures related to osteoporosis. Sometimes osteoporosis can be diagnosed with fractures or with a serious health problem, and most women are probably unaware of the risk factors which can be changed by prevention. The first step in preventing osteoporosis in women should be to make them aware of the risk factors. OBJECTIVES: Our aim was to determine Turkish women's knowledge about and attitudes to osteoporosis and its prevention. METHODS: A total of 311 women who applied to the Family Medicine department of the Middle East Technical University Medical Center were asked to fill in a questionnaire about osteoporosis. Only 270 of the 311 women who completed the entire questionnaire were included in the study. RESULTS: Nearly 90% of the women surveyed thought they were somewhat familiar with osteoporosis. However, >65% were unaware that the disease is directly responsible for disabling hip fractures, and >40% were unable to identify significant risk factors. Only 36% of the respondents could correctly identify the calcium-rich foods among the choices. CONCLUSION: According to our survey, a considerable number of the Turkish women in our settlement are unaware of the risk factors and the consequences of osteoporosis. Therefore, the women have inadequate knowledge of osteoporosis. There should be information resources easily accessible for the patients. The most important organizational incentives for providing patient information are further health promotion by the health authorities and the support of family physicians and the primary health care team. PMID- 11264273 TI - Factors influencing contraceptive use in Tehran. AB - BACKGROUND: Despite reluctance to conceive, approximately 30% of couples do not use any method of contraception. Health concerns, side effects, failure of the method and some demographic issues such as education, age, residential region and number of living children have a major effect on contraceptive use. OBJECTIVE: The aim of the present study was to determine those factors which influence contraceptive use in Tehran. METHODS: Data from the project "The Study of the Effectiveness of Postpartum Consultation about Family Planning on Contraceptive Practice during 2 years after Parturition in University Hospitals of Tehran in 1996" were applied for the analysis of those factors which influence contraceptive use by Iranian couples. A total of 4177 women of reproductive age who gave birth in one of the 12 hospitals in Tehran during the 24 hours following the interview of the initial study and had at least one living child were enrolled in the present study. The questionnaire used included some questions about socio-demographic status, fertility history, knowledge of contraceptives and the source of this knowledge, and previous contraception practice and its effectiveness. RESULTS: Using a logistic regression model, it was found that age, women's level of education, their husbands' level of education and previous familiarity with contraceptive methods were the most significant factors influencing contraceptive use. CONCLUSIONS: It is suggested that health policy makers strengthen the family planning services through providing appropriate counselling in family planning clinics. PMID- 11264274 TI - Antibiotic prescribing in acute infections of the nose or sinuses: a matter of personal habit? AB - BACKGROUND: A proper understanding of how and why GPs prescribe antibiotics in general practice is essential for the design of strategies aimed at making prescribing more rational. OBJECTIVE: The intention of this study is to contribute to such understanding by investigating which elements are important in the GP's decision to prescribe antibiotics for patients with acute infectious complaints of the nose and/or sinuses. METHODS: During their training in general practice, students observed the following elements while attending encounters between their trainer-GP and patients with a runny nose, blocked nose or cough: patient characteristics, contact characteristics, signs and symptoms, diagnosis and prescriptions. Information on practice characteristics and characteristics of the trainer-GP were collected. Data were analysed using multiple logistic regression and multiple linear regression. RESULTS: A total of 722 cases were analysed with the following results: the best independent predictor of an antibiotic prescription is the individual antibiotic prescribing rate (IAPR), which expresses the personal habit of the GP in prescribing antibiotics [adjusted odds ratio (OR) 5.27, 95% confidence interval (CI) 3.22-8.62]. Others are the diagnostic labels "sinusitis" (adjusted OR 2.80, 95% CI 1.2-6.49) and "flu-like syndrome" (adjusted OR 0.08, 95% CI 0.01-0.45), and the sign "sinus tenderness" (adjusted OR 4.37, CI 2.15-8.89). The antibiotic prescribing behaviour intensifies with an increasing tendency to prescribe medication in general (beta = 0.46, P: < 0.00) and with an increasing defensive attitude (beta = 0.22, P: < 0.05). CONCLUSIONS: Whether or not a patient with an acute infection of the nose and/or sinuses will be handed an antibiotic prescription seems to depend more on the attending doctor's prescribing behaviour than on the clinical picture. Further qualitative research into attitudes which may be related to a high tendency to prescribe antibiotics consequently is of the utmost importance. PMID- 11264275 TI - Barriers in the care of patients who have experienced a traumatic event: the perspective of general practice. AB - BACKGROUND: Previous research has indicated that GPs encounter barriers in the care of patients who have experienced a traumatic event. OBJECTIVES: The aims of the present study were to map barriers GPs encounter in the care of patients who experience a traumatic event and solutions for these barriers, and to estimate the influence of GP characteristics on the number of barriers experienced. METHODS: Telephone interviews were conducted among a sample of 500 Dutch GPs stratified by sex. Topics covered barriers in the care of victims of: accidents, incest in the past, ongoing physical or sexual abuse of adults, and ongoing physical or sexual abuse of children. RESULTS: The response rate was 44%. GPs are regularly confronted with patients who have experienced a traumatic event. GPs experience 10% barriers in care of patients who have difficulties getting over an accident, 13% in the care of incest victims, 16% in the care of adults who are physically or sexually abused, and 20% in the care of physically or sexually abused children. Most of the GPs recently updated their knowledge of care of victims of traumatic events, but still the majority feel in need of additional expert training. CONCLUSION: GPs experience the greatest number of barriers in the care of children who are abused. GP characteristics were not related to the number of barriers. However, seeing more victims was related to fewer barriers. To facilitate GP care of victims of traumatic events, GP training and continuing medical education should focus especially on skills education regarding the detection and initial treatment of traumatic events of ongoing physical or sexual abuse. PMID- 11264276 TI - Evidence-based guideline for the primary care management of stable angina. AB - This is an updated version of the first North of England Stable Angina Guideline (1,2) and summarizes the full guideline. (3) This paper presents all the recommendations within the guideline; and where these are new or substantially altered from the original version, it also presents a summary of the supporting evidence. The aims and methods of development (summarized in Box 1) of this guideline are unchanged from the original version, to which readers are directed for more detail. The research questions raised during the development of this guideline are shown in Box 2. PMID- 11264277 TI - Evidence-based guideline on the primary care management of asthma. AB - This is an updated version of the first North of England Asthma Guideline (1,2) and summarizes the full guideline. (3) This paper presents all the recommendations within the guideline and, where these are new or substantially altered from the original version, it also presents a summary of the supporting evidence. The aims and methods of development (summarized in Box 1) of this guideline are unchanged from the original version, to which readers are directed for more detail. The research questions raised during the development of this guideline are shown in Box 2. PMID- 11264278 TI - Selections from current literature: Recent developments in human HIV-1 vaccine. AB - This article reviews recent changes in the design of HIV-I vaccine. The safety information for current vaccines has been established, and future ethical considerations are reviewed. Of recent significance, gp120 envelope vaccines are being combined with canarypox vectors in an effort to elicit a broad immune response. This will probably be the aim of future research for an ever-growing problem. A total of 10 143 AIDS cases have occurred in the USA in the year 2000.(1) The international burden is grimmer. For example, approximately 13% of South Africans between 20 and 64 years old are HIV positive. Predictions estimate this number to rise to almost 30% by 2010.(2) With many different HIV serotypes, a worldwide effective HIV vaccine is not in the near future.(3) However, the scientific community has bolstered its effort by strengthening resources and developing national and international collaboration groups focused in developing a safe and effective HIV vaccine. PMID- 11264280 TI - Vitronectin functions as a cofactor for rapid inhibition of activated protein C by plasminogen activator inhibitor-1. Implications for the mechanism of profibrinolytic action of activated protein C. AB - Activated protein C (APC) is a natural anticoagulant in plasma that down regulates the coagulation cascade by degrading factors Va and VIIIa. In addition to its anticoagulant function, APC is also known to possess a profibrinolytic property. This property of APC has been attributed to its ability to neutralize PAI-1, thereby increasing the concentration of tissue plasminogen activator in plasma leading to up-regulation of the fibrinolytic cascade. This hypothesis, however, has not been well established, since the concentration of PAI-1 in plasma is low, and its reactivity with APC is very slow in a purified system. Here we demonstrate that vitronectin enhances the reactivity of PAI-1 with APC approximately 300-fold making PAI-1 the most efficient inhibitor of APC thus far reported (k(2) = 1.8 x 10(5) m(-)1 s(-)1). We further show that PAI-1 inhibition of the Glu(192) --> Gln mutant of APC is enhanced approximately 40-fold, independent of vitronectin, suggesting that vitronectin partially overcomes the inhibitory interaction of PAI-1 with Glu(192). Additionally, we show that PAI-1 inhibition of the Lys(37)-Lys(38)-Lys(39) --> Pro-Gln-Glu mutant of APC is severely impaired, suggesting that, similar to tissue plasminogen activator, the basic 39-loop of APC plays a critical role in the reaction. Together, these results suggest that vitronectin functions as a cofactor to promote the profibrinolytic activity of APC. PMID- 11264281 TI - Tumor necrosis factor-alpha selectively induces MnSOD expression via mitochondria to-nucleus signaling, whereas interleukin-1beta utilizes an alternative pathway. AB - Mitochondrial levels of the anti-oxidant enzyme, manganese superoxide dismutase (MnSOD), are dramatically elevated in response to stimulation with tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and lipopolysaccharide (LPS). However, the precise intracellular signaling pathways responsible for this inducible expression are poorly understood. MnSOD expression in pulmonary epithelial and endothelial cells, treated with inflammatory mediators and various inhibitors, was studied by Northern analysis. The mitochondrial electron transport chain inhibitors, antimycin A and myxothiazol, selectively blocked TNF alpha-inducible expression of MnSOD but not that of IL-1beta or LPS, indicating different signaling pathways. N-Acetylcysteine could reliably decrease inducible MnSOD expression by TNF-alpha, but not IL-1, linking reactive oxygen species (ROS) to the TNF-alpha signaling pathway. Elevated levels of arachidonic acid have been demonstrated previously to generate mitochondrial ROS. A specific cytoplasmic phospholipase A(2) inhibitor reduced stimulated MnSOD expression by TNF-alpha, but not by IL-1beta, further supporting the role of ROS. Other investigators have shown that MnSOD expression may be regulated by NF-kappaB. Our results with a specific inhibitory kappa-kinase inhibitor indicate that NF-kappaB modulates IL-1beta signaling but not the TNF-alpha pathway. Thus, we have demonstrated that although inducible MnSOD transcription may appear similar at the messenger RNA level, the intracellular signaling pathways are differentially regulated. PMID- 11264282 TI - Tts, a processive beta-glucosyltransferase of Streptococcus pneumoniae, directs the synthesis of the branched type 37 capsular polysaccharide in Pneumococcus and other gram-positive species. AB - The type 37 capsule of Streptococcus pneumoniae is a homopolysaccharide built up from repeating units of [beta-d-Glcp-(1-->2)]-beta-d-Glcp-(1-->3). The elements governing the expression of the tts gene, coding for the glucosyltransferase involved in the synthesis of the type 37 pneumococcal capsular polysaccharide, have been studied. Primer extension analysis and functional tests demonstrated the presence of four new transcriptional start points upstream of the previously reported tts promoter (ttsp). Most interesting, three of these transcriptional start points are located in a RUP element thought to be involved in recombinational events (Oggioni, M. R., and Claverys, J. P. (1999) Microbiology 145, 2647-2653). Transformation experiments using either a recombinant plasmid containing the whole transcriptional unit of tts or chromosomal DNA from a type 37 pneumococcus showed that tts is the only gene required to drive the biosynthesis of a type 37 capsule in S. pneumoniae and other Gram-positive bacteria, namely Streptococcus oralis, Streptococcus gordonii, and Bacillus subtilis. The Tts synthase was overproduced in S. pneumoniae and purified as a membrane-associated enzyme. These membrane preparations used UDP-Glc as substrate to catalyze the synthesis of a high molecular weight polysaccharide immunologically identical to the type 37 capsule. In addition, UDP-Gal was also a substrate to produce type 37 polysaccharide since a strong UDP-Glc-4'-epimerase activity is associated to the membrane fraction of S. pneumoniae. These results indicated that Tts has a dual biochemical activity that leads to the synthesis of the branched type 37 polysaccharide. PMID- 11264283 TI - Changes in the lipid turnover, composition, and organization, as sphingolipid enriched membrane domains, in rat cerebellar granule cells developing in vitro. AB - In the present paper, we report on the properties of sphingolipid-enriched domains of rat cerebellar granule cells in culture at different stages of neuronal development. The major lipid components of these domains were glycerophospholipids and cholesterol. Glycerophospholipids were 45-75% and cholesterol 15-45% of total lipids of the domains. This corresponded to 5-17% of total cell glycerophospholipids and 15-45% of total cell cholesterol. Phosphatidylcholine, mainly dipalmitoylphosphatidylcholine, was 66-85% of all the glycerophospholipids associated with these domains. Consequently, the palmitoyl residue was significantly enriched in the domains. The surface occupied by these structures increased during development. 40-70% of cell sphingolipids segregated in sphingolipid-enriched membrane domains, with the maximum ganglioside density in fully differentiated neurons. A high content of ceramide was found in the domains of aging neurons. Then, the sphingolipid/glycerophospholipid molar ratio was more than doubled during the initial stage of development, whereas the cholesterol/glycerophospholipid molar ratio gradually decreased during in vitro differentiation. Phosphorylated phosphoinositides, which were scant in the domains of undifferentiated cells, dramatically increased during differentiation and aging in culture. Proteins were minor components of the domains (0.1-2.8% of all domain components). Phosphotyrosine-containing proteins were selectively recovered in the sphingolipid-enriched domain. Among these, Src family protein tyrosine kinases, known to participate to the process of neuronal differentiation, were associated with the sphingolipid-enriched domains in a way specific for the type of kinase and for the developmental stage of the cell. Proteins belonging to other signaling pathways, such as phosphoinositide 3-kinase and its downstream target, Akt, were not associated with the domains. PMID- 11264284 TI - Two Menkes-type atpases supply copper for photosynthesis in Synechocystis PCC 6803. AB - Synechocystis PCC 6803 contains four genes encoding polypeptides with sequence features of CPx-type ATPases, two of which are now designated pacS and ctaA. We show that CtaA and PacS (but not the related transporters, ZiaA or CoaT) facilitate switching to the use of copper (in plastocyanin) as an alternative to iron (in cytochrome c(6)) for the carriage of electrons within the thylakoid lumen. Disruption of pacS reduced copper tolerance but enhanced silver tolerance, and pacS-mediated restoration of copper tolerance was used to select transformants. Disruption of ctaA caused no change in copper tolerance but reduced the amount of copper cell(-1). In cultures supplemented with 0.2 microm copper, photooxidation of cytochrome c(6) (PetJ) was depressed in wild-type cells but remained elevated in both Synechocystis PCC 6803(ctaA) and Synechocystis PCC 6803(pacS). Conversely, plastocyanin transcripts (petE) were less abundant in both mutants at this [copper]. Synechocystis PCC 6803(ctaA) and Synechocystis PCC 6803(pacS) showed increased iron dependence with impaired growth in deferoxamine mesylate (iron chelator)-containing media. Double mutants also deficient in cytochrome c(6), Synechocystis PCC 6803(petJ,ctaA) and Synechocystis PCC 6803(petJ,pacS), were viable, but the former had increased copper dependence with severely impaired growth in the presence of bathocuproinedisulfonic acid (copper chelator). Analogous transporters are likely to supply copper to plastocyanin in chloroplasts. PMID- 11264285 TI - Modulation of cellular iron metabolism by hydrogen peroxide. Effects of H2O2 on the expression and function of iron-responsive element-containing mRNAs in B6 fibroblasts. AB - Cellular iron uptake and storage are coordinately controlled by binding of iron regulatory proteins (IRP), IRP1 and IRP2, to iron-responsive elements (IREs) within the mRNAs encoding transferrin receptor (TfR) and ferritin. Under conditions of iron starvation, both IRP1 and IRP2 bind with high affinity to cognate IREs, thus stabilizing TfR and inhibiting translation of ferritin mRNAs. The IRE/IRP regulatory system receives additional input by oxidative stress in the form of H(2)O(2) that leads to rapid activation of IRP1. Here we show that treating murine B6 fibroblasts with a pulse of 100 microm H(2)O(2) for 1 h is sufficient to alter critical parameters of iron homeostasis in a time-dependent manner. First, this stimulus inhibits ferritin synthesis for at least 8 h, leading to a significant (50%) reduction of cellular ferritin content. Second, treatment with H(2)O(2) induces a approximately 4-fold increase in TfR mRNA levels within 2-6 h, and subsequent accumulation of newly synthesized protein after 4 h. This is associated with a profound increase in the cell surface expression of TfR, enhanced binding to fluorescein-tagged transferrin, and stimulation of transferrin-mediated iron uptake into cells. Under these conditions, no significant alterations are observed in the levels of mitochondrial aconitase and the Divalent Metal Transporter DMT1, although both are encoded by two as yet lesser characterized IRE-containing mRNAs. Finally, H(2)O(2)-treated cells display an increased capacity to sequester (59)Fe in ferritin, despite a reduction in the ferritin pool, which results in a rearrangement of (59)Fe intracellular distribution. Our data suggest that H(2)O(2) regulates cellular iron acquisition and intracellular iron distribution by both IRP1-dependent and -independent mechanisms. PMID- 11264286 TI - Cytochrome c release occurs via Ca2+-dependent and Ca2+-independent mechanisms that are regulated by Bax. AB - Release of cytochrome c from mitochondria is a key initiative step in the apoptotic process, although the mechanisms regulating this event remain elusive. In the present study, using isolated liver mitochondria, we demonstrate that cytochrome c release occurs via distinct mechanisms that are either Ca(2+) dependent or Ca(2+)-independent. An increase in mitochondrial matrix Ca(2+) promotes the opening of the permeability transition (PT) pore and the release of cytochrome c, an effect that is significantly enhanced when these organelles are incubated in a reaction buffer that is based on a physiologically relevant concentration of K(+) (150 mm KCl) versus a buffer composed of mannitol/sucrose/Hepes. Moreover, low concentrations of Ca(2+) are sufficient to induce mitochondrial cytochrome c release without measurable manifestations of PT, though inhibitors of PT effectively prevent this release, indicating that the critical threshold for PT varies among mitochondria within a single population of these organelles. In contrast, Ca(2+)-independent cytochrome c release is induced by oligomeric Bax protein and occurs without mitochondrial swelling or the release of matrix proteins, although our data also indicate that Bax enhances permeability transition-induced cytochrome c release. Taken together, our results suggest that the intramitochondrial Ca(2+) concentration, as well as the reaction buffer composition, are key factors in determining the mode and amount of cytochrome c release. Finally, oligomeric Bax appears to be capable of stimulating cytochrome c release via both Ca(2+)-dependent and Ca(2+)-independent mechanisms. PMID- 11264287 TI - 1-Deoxy-D-xylulose-5-phosphate synthase, a limiting enzyme for plastidic isoprenoid biosynthesis in plants. AB - The initial step of the plastidic 2C-methyl-D-erythritol 4-phosphate (MEP) pathway that produces isopentenyl diphosphate is catalyzed by 1-deoxy-d-xylulose 5-phosphate synthase. To investigate whether or not 1-deoxy-d-xylulose-5 phosphate synthase catalyzes a limiting step in the MEP pathway in plants, we produced transgenic Arabidopsis plants that over- or underexpress this enzyme. Compared with non-transgenic wild-type plants, the transgenic plants accumulate different levels of various isoprenoids such as chlorophylls, tocopherols, carotenoids, abscisic acid, and gibberellins. Phenotypically, the transgenic plants had slight alterations in growth and germination rates. Because the levels of several plastidic isoprenoids correlate with changes in 1-deoxy-D-xylulose-5 phosphate synthase levels, we conclude that this enzyme catalyzes one of the rate limiting steps of the MEP biosynthetic pathway. Furthermore, since the product of the MEP pathway is isopentenyl diphosphate, our results suggest that in plastids the pool of isopentenyl diphosphate is limiting to isprenoid production. PMID- 11264289 TI - Different regulatory mechanisms modulate the expression of a dinoflagellate iron superoxide dismutase. AB - Regulation of antioxidant enzymes is critical to control the levels of reactive oxygen species in cell compartments highly susceptible to oxidative stress. In this work, we studied the regulation of a chloroplastic iron superoxide dismutase (Fe-SOD) from Lingulodinium polyedrum (formerly Gonyaulax polyedra) under different physiological conditions. A cDNA-encoding Fe-SOD was isolated from this dinoflagellate, showing high sequence similarity to cyanobacterial, algal, and plant Fe-SODs. Under standard growth conditions, on a 12:12-h light-dark cycle, Lingulodinium polyedrum Fe-SOD exhibited a daily rhythm of activity and cellular abundance with the maximum occurring during the middle of the light phase. Northern analyses showed that this rhythmicity is not related to changes in Fe SOD mRNA levels, indicative of translational regulation. By contrast, conditions of metal-induced oxidative stress resulted in higher levels of Fe-SOD transcripts, suggesting that transcriptional control is responsible for increased protein and activity levels. Daily (circadian) and metal-induced up-regulation of Fe-SOD expression in L. polyedrum are thus mediated by different regulatory pathways, allowing biochemically distinct changes appropriate to oxidative challenges. PMID- 11264288 TI - Vacuolar localization of oligomeric alpha-mannosidase requires the cytoplasm to vacuole targeting and autophagy pathway components in Saccharomyces cerevisiae. AB - One challenge facing eukaryotic cells is the post-translational import of proteins into organelles. This problem is exacerbated when the proteins assemble into large complexes. Aminopeptidase I (API) is a resident hydrolase of the vacuole/lysosome in the yeast Saccharomyces cerevisiae. The precursor form of API assembles into a dodecamer in the cytosol and maintains this oligomeric form during the import process. Vacuolar delivery of the precursor form of API requires a vesicular mechanism termed the cytoplasm to vacuole targeting (Cvt) pathway. Many components of the Cvt pathway are also used in the degradative autophagy pathway. alpha-Mannosidase (Ams1) is another resident hydrolase that enters the vacuole independent of the secretory pathway; however, its mechanism of vacuolar delivery has not been established. We show vacuolar localization of Ams1 is blocked in mutants that are defective in the Cvt and autophagy pathways. We have found that Ams1 forms an oligomer in the cytoplasm. The oligomeric form of Ams1 is also detected in subvacuolar vesicles in strains that are blocked in vesicle breakdown, indicating that it retains its oligomeric form during the import process. These results identify Ams1 as a second biosynthetic cargo protein of the Cvt and autophagy pathways. PMID- 11264290 TI - Conversion of Glu-plasminogen to Lys-plasminogen is necessary for optimal stimulation of plasminogen activation on the endothelial cell surface. AB - When Glu-plasminogen is bound to cells, plasmin (Pm) formation by plasminogen (Pg) activators is markedly enhanced compared with the reaction in solution. It is not known whether the direct activation of Glu-Pg by Pg activators is promoted on the cell surface or whether plasminolytic conversion of Glu-Pg to the more readily activated Lys-Pg is necessary for enhanced Pm formation on the cell surface. To distinguish between these potential mechanisms, we tested whether Pm formation on the cell surface could be stimulated in the absence of conversion of Glu-Pg to Lys-Pg. Rates of activation of Glu-Pg, Lys-Pg, and a mutant Glu-Pg, [D646E]Glu-Pg, by either tissue Pg activator (t-PA) or urokinase (u-PA) were compared when these Pg forms were either bound to human umbilical vein endothelial cells (HUVEC) or in solution. ([D646E]Glu-Pg can be cleaved at the Arg(561)-Val(562) bond by Pg activators but does not possess Pm activity subsequent to this cleavage because of the mutation of Asp(646) of the serine protease catalytic triad.) Glu-Pg activation by t-PA was enhanced on HUVEC compared with the solution phase by 13-fold. In contrast, much less enhancement of Pg activation was observed with [D646E]Glu-Pg ( approximately 2-fold). Although the extent of activation of Lys-Pg on cells was similar to that of Glu Pg, the cells afforded minimal enhancement of Lys-Pg activation compared with the solution phase (1.3-fold). Similar results were obtained when u-PA was used as activator. When Glu-Pg was bound to the cell in the presence of either t-PA or u PA, conversion to Lys-Pg was observed, but conversion of ([D646E]Glu-Pg to ([D646E]Lys-Pg was not detected, consistent with the conversion of Glu-Pg to Lys Pg being necessary for optimal enhancement of Pg activation on cell surfaces. Furthermore, we found that conversion of [D646E]Glu-Pg to [D646E]Lys-Pg by exogenous Pm was markedly enhanced ( approximately 20-fold) on the HUVEC surface, suggesting that the stimulation of the conversion of Glu-Pg to Lys-Pg is a key mechanism by which cells enhance Pg activation. PMID- 11264291 TI - Inflammatory mediators potentiate ATP-gated channels through the P2X(3) subunit. AB - The P2X(3) receptor is an ATP-gated ion channel predominantly expressed in nociceptive neurons from the dorsal root ganglion. P2X(3) receptor channels are highly expressed in sensory neurons and probably contribute to the sensation of pain. Kinetics of P2X(3) currents are characterized by rapid desensitization (<100 ms) and slow recovery (>20 s). Thus, any mechanism modulating rate of desensitization and/or recovery may have profound effect on susceptibility of nociceptive neurons expressing P2X(3) to ATP. Here we show that currents mediated by P2X(3) receptor channels and the heteromeric channel P2X(2/3) composed of P2X(2) and P2X(3) subunits are potentiated by the neuropeptides substance P and bradykinin, which are known to modulate pain perception. The effect is mediated by the respective neuropeptide receptors, can be mimicked by phorbol ester and blocked by inhibitors of protein kinases. Together with data from site-directed mutagenesis our results suggest that inflammatory mediators sensitize nociceptors through phosphorylation of P2X(3) and P2X(2/3) ion channels or associated proteins. PMID- 11264292 TI - Control of electron transfer in nitric-oxide synthases. Swapping of autoinhibitory elements among nitric-oxide synthase isoforms. AB - To clarify the role of the autoinhibitory insert in the endothelial (eNOS) and neuronal (nNOS) nitric-oxide synthases, the insert was excised from nNOS and chimeras with its reductase domain; the eNOS and nNOS inserts were swapped and put into the normally insertless inducible (iNOS) isoform and chimeras with the iNOS reductase domain; and an RRKRK sequence in the insert suggested by earlier peptide studies to be important (Salerno, J. C., Harris, D. E., Irizarry, K., Patel, B., Morales, A. J., Smith, S. M., Martasek, P., Roman, L. J., Masters, B. S., Jones, C. L., Weissman, B. A., Lane, P., Liu, Q., and Gross, S. S. (1997) J. Biol. Chem. 272, 29769-29777) was mutated. Insertless nNOS required calmodulin (CaM) for normal NOS activity, but the Ca(2+) requirement for this activity was relaxed. Furthermore, insert deletion enhanced CaM-free electron transfer within nNOS and chimeras with the nNOS reductase, emphasizing the involvement of the insert in modulating electron transfer. Swapping the nNOS and eNOS inserts gave proteins with normal NOS activities, and the nNOS insert acted normally in raising the Ca(2+) dependence when placed in eNOS. Insertion of the eNOS insert into iNOS and chimeras with the iNOS reductase domain significantly lowered NOS activity, consistent with inhibition of electron transfer by the insert. Mutation of the eNOS RRKRK to an AAAAA sequence did not alter the eNOS Ca(2+) dependence but marginally inhibited electron transfer. The salt dependence suggests that the insert modulates electron transfer within the reductase domain prior to the heme/reductase interface. The results clarify the role of the reductase insert in modulating the Ca(2+) requirement, electron transfer rate, and overall activity of nNOS and eNOS. PMID- 11264293 TI - Imidazole glycerol phosphate synthase from Thermotoga maritima. Quaternary structure, steady-state kinetics, and reaction mechanism of the bienzyme complex. AB - Imidazole glycerol phosphate synthase, which links histidine and de novo purine biosynthesis, is a member of the glutamine amidotransferase family. In bacteria, imidazole glycerol phosphate synthase constitutes a bienzyme complex of the glutaminase subunit HisH and the synthase subunit HisF. Nascent ammonia produced by HisH reacts at the active site of HisF with N'-((5' phosphoribulosyl)formimino)-5-aminoimidazole-4-carboxamide-ribonucleotide to yield the products imidazole glycerol phosphate and 5-aminoimidazole-4 carboxamide ribotide. In order to elucidate the interactions between HisH and HisF and the catalytic mechanism of the HisF reaction, the enzymes tHisH and tHisF from Thermotoga maritima were produced in Escherichia coli, purified, and characterized. Isolated tHisH showed no detectable glutaminase activity but was stimulated by complex formation with tHisF to which either the product imidazole glycerol phosphate or a substrate analogue were bound. Eight conserved amino acids at the putative active site of tHisF were exchanged by site-directed mutagenesis, and the purified variants were investigated by steady-state kinetics. Aspartate 11 appeared to be essential for the synthase activity both in vitro and in vivo, and aspartate 130 could be partially replaced only by glutamate. The carboxylate groups of these residues could provide general acid/base catalysis in the proposed catalytic mechanism of the synthase reaction. PMID- 11264294 TI - Pregnancy-associated plasma protein-A2 (PAPP-A2), a novel insulin-like growth factor-binding protein-5 proteinase. AB - A novel metalloproteinase with similarity to pregnancy-associated plasma protein A (PAPP-A), which we denoted PAPP-A2, has been identified. Through expression in mammalian cells we showed that recombinant PAPP-A2 polypeptide of 1558 residues resulted from processing of a 1791-residue prepro-protein. Unlike PAPP-A, PAPP-A2 migrated as a monomer (of 220 kDa) in non-reducing SDS-polyacrylamide gel electrophoresis. The prepro-parts of PAPP-A2 and PAPP-A are not homologous, but mature PAPP-A2 shares 45% of its residues with PAPP-A. Because PAPP-A specifically cleaves insulin-like growth factor-binding protein (IGFBP)-4, one of six known modulators of IGF-I and -II, we looked for a possible PAPP-A2 substrate among the members of this family. We showed that PAPP-A2 specifically cleaved IGFBP-5 at one site, between Ser-143 and Lys-144. In contrast to the cleavage of IGFBP-4 by PAPP-A that strictly requires the presence of IGF, the cleavage of IGFBP-5 by PAPP-A2 was IGF-independent. Recent data firmly establish PAPP-A and IGFBP-4 as an important functional pair in several systems. Because of its close relationship with PAPP-A, both structurally and functionally, PAPP-A2 is a likely candidate IGFBP-5 proteinase in many tissues and conditioned media where IGFBP-5 proteolysis has been reported. PMID- 11264295 TI - Neurofilaments consist of distinct populations that can be distinguished by C terminal phosphorylation, bundling, and axonal transport rate in growing axonal neurites. AB - We examined the steady-state distribution and axonal transport of neurofilament (NF) subunits within growing axonal neurites of NB2a/d1 cells. Ultrastructural analyses demonstrated a longitudinally oriented "bundle" of closely apposed NFs that was surrounded by more widely spaced individual NFs. NF bundles were recovered during fractionation and could be isolated from individual NFs by sedimentation through sucrose. Immunoreactivity toward the restrictive C-terminal phospho-dependent antibody RT97 was significantly more prominent on bundled than on individual NFs. Microinjected biotinylated NF subunits, GFP-tagged NF subunits expressed after transfection, and radiolabeled endogenous subunits all associated with individual NFs before they associated with bundled NFs. Biotinylated and GFP tagged NF subunits did not accumulate uniformly along bundled NFs; they initially appeared within the proximal portion of the NF bundle and only subsequently were observed along the entire length of bundled NFs. These findings demonstrate that axonal NFs are not homogeneous but, rather, consist of distinct populations. One of these is characterized by less extensive C-terminal phosphorylation and a relative lack of NF-NF interactions. The other is characterized by more extensive C-terminal NF phosphorylation and increased NF-NF interactions and either undergoes markedly slower axonal transport or does not transport and undergoes turnover via subunit and/or filament exchange with individual NFs. Inhibition of phosphatase activities increased NF-NF interactions within living cells. These findings collectively suggest that C-terminal phosphorylation and NF-NF interactions are responsible for slowing NF axonal transport. PMID- 11264296 TI - Postsynaptic calcium transients evoked by activation of individual hippocampal mossy fiber synapses. AB - Control of Ca(2+) within dendritic spines is critical for excitatory synaptic function and plasticity, but little is known about Ca(2+) dynamics at thorny excrescences, the complex spines on hippocampal CA3 pyramidal cells contacted by mossy fiber terminals of dentate granule cell axons. We have monitored subthreshold stimulus-dependent postsynaptic Ca(2+) transients in optically and ultrastructurally characterized complex spines and find that such spines can act as discrete units of Ca(2+) response. In contrast to the more common "simple" spines, synaptically evoked Ca(2+) transients at complex spines have only a small NMDA receptor-dependent component and do not involve release of calcium from internal stores. Instead, they result mainly from AMPA receptor-gated Ca(2+) influx through voltage-activated calcium channels on the spine; these channels provide graded amplification of the response of thorny excrescences to individual mossy fiber synaptic events. PMID- 11264297 TI - Propagation of intercellular calcium waves in retinal astrocytes and Muller cells. AB - Intercellular Ca(2+) waves are believed to propagate through networks of glial cells in culture in one of two ways: by diffusion of IP(3) between cells through gap junctions or by release of ATP, which functions as an extracellular messenger. Experiments were conducted to determine the mechanism of Ca(2+) wave propagation between glial cells in an intact CNS tissue. Calcium waves were imaged in the acutely isolated rat retina with the Ca(2+) indicator dye fluo-4. Mechanical stimulation of astrocyte somata evoked Ca(2+) waves that propagated through both astrocytes and Muller cells. Octanol (0.5 mm), which blocks coupling between astrocytes and Muller cells, did not reduce propagation into Muller cells. Purinergic receptor antagonists suramin (100 microm), PPADS (20-50 microm), and apyrase (80 U/ml), in contrast, substantially reduced wave propagation into Muller cells (wave radii reduced to 16-61% of control). Suramin also reduced wave propagation from Muller cell to Muller cell (51% of control). Purinergic antagonists reduced wave propagation through astrocytes to a lesser extent (64-81% of control). Mechanical stimulation evoked the release of ATP, imaged with the luciferin-luciferase bioluminescence assay. Peak ATP concentration at the surface of the retina averaged 78 microm at the stimulation site and 6.8 microm at a distance of 100 microm. ATP release propagated outward from the stimulation site with a velocity of 41 microm/sec, somewhat faster than the 28 microm/sec velocity of Ca(2+) waves. Ejection of 3 microm ATP onto the retinal surface evoked propagated glial Ca(2+) waves. Together, these results indicate that Ca(2+) waves are propagated through retinal glial cells by two mechanisms. Waves are propagated through astrocytes principally by diffusion of an internal messenger, whereas waves are propagated from astrocytes to Muller cells and from Muller cells to other Muller cells primarily by the release of ATP. PMID- 11264298 TI - The influence of glutamate receptor 2 expression on excitotoxicity in Glur2 null mutant mice. AB - AMPA receptor (AMPAR)-mediated ionic currents that govern gene expression, synaptic strength, and plasticity also can trigger excitotoxicity. However, native AMPARs exhibit heterogeneous pharmacological, biochemical, and ionic permeability characteristics, which are governed partly by receptor subunit composition. Consequently, the mechanisms governing AMPAR-mediated excitotoxicity have been difficult to elucidate. The GluR2 subunit is of particular interest because it influences AMPAR pharmacology, Ca(2+) permeability, and AMPAR interactions with intracellular proteins. In this paper we used mutant mice lacking the AMPAR subunit GluR2 to study AMPAR-mediated excitotoxicity in cultured cortical neurons and in hippocampal neurons in vivo. We examined the hypothesis that in these mice the level of GluR2 expression governs the vulnerability of neurons to excitotoxicity and further examined the ionic mechanisms that are involved. In cortical neuronal cultures AMPAR-mediated neurotoxicity paralleled the magnitude of kainate-evoked AMPAR-mediated currents, which were increased in neurons lacking GluR2. Ca(2+) permeability, although elevated in GluR2-deficient neurons, did not correlate with excitotoxicity. However, toxicity was reduced by removal of extracellular Na(+), the main charge carrier of AMPAR-mediated currents. In vivo, the vulnerability of CA1 hippocampal neurons to stereotactic kainate injections and of CA3 neurons to intraperitoneal kainate administration was independent of GluR2 level. Neurons lacking the GluR2 subunit did not demonstrate compensatory changes in the distribution, expression, or function of AMPARs or of Ca(2+)-buffering proteins. Thus GluR2 level may influence excitotoxicity by effects additional to those on Ca(2+) permeability, such as effects on agonist potency, ionic currents, and synaptic reorganization. PMID- 11264299 TI - S100beta interaction with tau is promoted by zinc and inhibited by hyperphosphorylation in Alzheimer's disease. AB - The zinc-binding protein S100beta has been identified as an interacting partner with the microtubule-associated protein tau. Both proteins are individually affected in Alzheimer's disease (AD). S100beta, is overexpressed in the disease, whereas hyperphosphorylated tau constitutes the primary component of neurofibrillary tangles. In this study, we examine factors that modulate their binding and the potential role the complex may play in AD pathogenesis. Zinc was identified as a critical component in the binding process and a primary modulator of S100beta-associated cellular responses. Abnormally phosphorylated tau extracted from AD tissue displayed a dramatically reduced capacity to bind S100beta, which was restored by pretreatment with alkaline phosphatase. In differentiated SH-SY5Y cells, exogenous S100beta was internalized and colocalized with tau consistent with an intracellular association. This was enhanced by the addition of zinc and eliminated by divalent metal chelators. S100beta uptake was also accompanied by extensive neurite outgrowth that may be mediated by its interaction with tau. S100beta-tau binding may represent a key pathway for neurite development, possibly through S100beta modulation of tau phosphorylation and/or functional stabilization of microtubules and process formation. S100beta tau interaction may be disrupted by hyperphosphorylation and/or imbalances in zinc metabolism, and this may contribute to the neurite dystrophy associated with AD. PMID- 11264300 TI - Caspase-8 is an effector in apoptotic death of dopaminergic neurons in Parkinson's disease, but pathway inhibition results in neuronal necrosis. AB - Caspase-8 is a proximal effector protein of the tumor necrosis factor receptor family death pathway. In the present human postmortem study, we observed a significantly higher percentage of dopaminergic (DA) substantia nigra pars compacta neurons that displayed caspase-8 activation in Parkinson's disease (PD) patients compared with controls. In an in vivo experimental PD model, namely subchronically 1,2,3,6-tetrahydropyridine-treated mice, we also show that caspase 8 is indeed activated after exposure to this toxin early in the course of cell demise, suggesting that caspase-8 activation precedes and is not the consequence of cell death. However, cotreatment of 1-methyl-4-phenylpyridinium-intoxicated primary DA cultures with broad-spectrum and specific caspase-8 inhibitors did not result in neuroprotection but seemed to trigger a switch from apoptosis to necrosis. We propose that this effect is related to ATP depletion and suggest that the use of caspase inhibitors in pathologies linked to intracellular energy depletion, such as PD, should be cautiously evaluated. PMID- 11264301 TI - Multiple distinct signal pathways, including an autocrine neurotrophic mechanism, contribute to the survival-promoting effect of depolarization on spiral ganglion neurons in vitro. AB - We have shown previously that BDNF, neurotrophin-3 (NT-3), chlorphenylthio-cAMP (cpt-cAMP) (a permeant cAMP analog), and membrane depolarization promote spiral ganglion neuron (SGN) survival in vitro in an additive manner, depolarization having the greatest efficacy. Expression of both BDNF and of NT-3 is detectable in cultured SGNs after plating in either depolarizing or nondepolarizing medium. These neurotrophins promote survival by an autocrine mechanism; TrkB-IgG or TrkC IgG, which block neurotrophin binding to, respectively, TrkB and TrkC, partially inhibit the trophic effect of depolarization. The mitogen-activated protein kinase kinase inhibitor PD98059 and the phosphatidylinositol-3-OH kinase inhibitor LY294002 both abolish trophic support by neurotrophins but only partially inhibit support by depolarization. Inhibition by these compounds is not additive with inhibition by Trk-IgGs. The cAMP antagonist Rp-adenosine-3',5' cyclic-phosphorothioate (Rp-cAMPS) abolishes survival attributable to cpt-cAMP but has no effect on that attributable to neurotrophins, nor do inhibitors of neurotrophin-dependent survival affect survival attributable to cpt-cAMP. However, Rp-cAMPS does partially inhibit depolarization-dependent survival, an inhibition that is additive with that by Trk-IgGs, PD98059, or LY294002. Moreover, Rp-cAMPS prevents depolarization-dependent survival of PC12 cells maintained in subthreshold levels of NGF. Inhibition of Ca(2+)/calmodulin dependent protein kinases (CaMKs) with KN-62 reduces SGN survival independently of Rp-cAMPS, Trk-IgGs, and LY294002 and additively with them. Combined inhibition of Trk, cAMP, and CaMK signaling prevents depolarization-dependent survival. Thus, survival of SGNs under depolarizing conditions involves additivity among a depolarization-independent autocrine pathway, a cAMP-dependent pathway, and a CaMK-dependent pathway. PMID- 11264302 TI - D1/D5 dopamine receptor activation differentially modulates rapidly inactivating and persistent sodium currents in prefrontal cortex pyramidal neurons. AB - Dopamine (DA) is a well established modulator of prefrontal cortex (PFC) function, yet the cellular mechanisms by which DA exerts its effects in this region are controversial. A major point of contention is the consequence of D(1) DA receptor activation. Several studies have argued that D(1) receptors enhance the excitability of PFC pyramidal neurons by augmenting voltage-dependent Na(+) currents, particularly persistent Na(+) currents. However, this conjecture is based on indirect evidence. To provide a direct test of this hypothesis, we combined voltage-clamp studies of acutely isolated layer V-VI prefrontal pyramidal neurons with single-cell RT-PCR profiling. Contrary to prediction, the activation of D(1) or D(5) DA receptors consistently suppressed rapidly inactivating Na(+) currents in identified corticostriatal pyramidal neurons. This modulation was attenuated by a D(1)/D(5) receptor antagonist, mimicked by a cAMP analog, and blocked by a protein kinase A (PKA) inhibitor. In the same cells the persistent component of the Na(+) current was unaffected by D(1)/D(5) receptor activation-suggesting that rapidly inactivating and persistent Na(+) currents arise in part from different channels. Single-cell RT-PCR profiling showed that pyramidal neurons coexpressed three alpha-subunit mRNAs (Nav1.1, 1.2, and 1.6) that code for the Na(+) channel pore. In neurons from Nav1.6 null mice the persistent Na(+) currents were significantly smaller than in wild-type neurons. Moreover, the residual persistent currents in these mutant neurons-which are attributable to Nav1.1/1.2 channels-were reduced significantly by PKA activation. These results argue that D(1)/D(5) DA receptor activation reduces the rapidly inactivating component of Na(+) current in PFC pyramidal neurons arising from Nav1.1/1.2 Na(+) channels but does not modulate effectively the persistent component of the Na(+) current that is attributable to Nav1.6 Na(+) channels. PMID- 11264303 TI - Neurofilaments are nonessential to the pathogenesis of toxicant-induced axonal degeneration. AB - Axonal neurofilament (NF) accumulations occur before development of symptoms and before other pathological changes among idiopathic neurodegenerative diseases and toxic neuropathies, suggesting a cause-effect relationship. The dependence of symptoms and axonal degeneration on neurofilament accumulation has been tested here in a transgenic mouse model (Eyer and Peterson, 1994) lacking axonal NFs and using two prototypic toxicant models. Chronic acrylamide (ACR) or 2,5-hexanedione exposure resulted in progressive and cumulative increases in sensorimotor deficits. Neurobehavioral tests demonstrated similar expression of neurotoxicity in transgenic (T) mice and their nontransgenic (NT) littermates (containing normal numbers of axonal NFs). Axonal lesions were frequently observed after exposure to either toxicant. Quantitation of ACR-induced lesions demonstrated the distal location of pathology and equal susceptibility of T and NT axons. We conclude that axonal NFs have no effect on neurotoxicity and the pattern of pathology in these mammalian toxic neuropathies. These results also suggest that the role of neurofilament accumulation in the pathogenesis of neurodegenerative diseases requires careful evaluation. PMID- 11264304 TI - Changes in microtubule stability and density in myelin-deficient shiverer mouse CNS axons. AB - Altered axon-Schwann cell interactions in PNS myelin-deficient Trembler mice result in changed axonal transport rates, neurofilament and microtubule associated protein phosphorylation, neurofilament density, and microtubule stability. To determine whether PNS and CNS myelination have equivalent effects on axons, neurofilaments, and microtubules in CNS, myelin-deficient shiverer axons were examined. The genetic defect in shiverer is a deletion in the myelin basic protein (MBP) gene, an essential component of CNS myelin. As a result, shiverer mice have little or no compact CNS myelin. Slow axonal transport rates in shiverer CNS axons were significantly increased, in contrast to the slowing in demyelinated PNS nerves. Even more striking were substantial changes in the composition and properties of microtubules in shiverer CNS axons. The density of axonal microtubules is increased, reflecting increased expression of tubulin in shiverer, and the stability of microtubules is drastically reduced in shiverer axons. Shiverer transgenic mice with two copies of a wild-type myelin basic protein transgene have an intermediate level of compact myelin, making it possible to determine whether the actual level of compact myelin is an important regulator of axonal microtubules. Both increased microtubule density and reduced microtubule stability were still observed in transgenic mouse nerves, indicating that signals beyond synaptogenesis and the mere presence of compact myelin are required for normal regulation of the axonal microtubule cytoskeleton. PMID- 11264305 TI - A depletable pool of adenosine in area CA1 of the rat hippocampus. AB - Adenosine plays a major modulatory and neuroprotective role in the mammalian CNS. During cerebral metabolic stress, such as hypoxia or ischemia, the increase in extracellular adenosine inhibits excitatory synaptic transmission onto vulnerable neurons via presynaptic adenosine A(1) receptors, thereby reducing the activation of postsynaptic glutamate receptors. Using a combination of extracellular and whole-cell recordings in the CA1 region of hippocampal slices from 12- to 24-d old rats, we have found that this protective depression of synaptic transmission weakens with repeated exposure to hypoxia, thereby allowing potentially damaging excitation to both persist for longer during oxygen deprivation and recover more rapidly on reoxygenation. This phenomenon is unlikely to involve A(1) receptor desensitization or impaired nucleoside transport. Instead, by using the selective A(1) antagonist 8-cyclopentyl-1,3-dipropylxanthine and a novel adenosine sensor, we demonstrate that adenosine production is reduced with repeated episodes of hypoxia. Furthermore, this adenosine depletion can be reversed at least partially either by the application of exogenous adenosine, but not by a stable A(1) agonist, N(6)-cyclopentyladenosine, or by endogenous means by prolonged (2 hr) recovery between hypoxic episodes. Given the vital neuroprotective role of adenosine, these findings suggest that depletion of adenosine may underlie the increased neuronal vulnerability to repetitive or secondary hypoxia/ischemia in cerebrovascular disease and head injury. PMID- 11264306 TI - LTD induction in adult visual cortex: role of stimulus timing and inhibition. AB - One Hertz stimulation of afferents for 15 min with constant interstimulus intervals (regular stimulation) can induce long-term depression (LTD) of synaptic strength in the neocortex. However, it is unknown whether natural patterns of low frequency afferent spike activity induce LTD. Although neurons in the neocortex can fire at overall rates as low as 1 Hz, the intervals between spikes are irregular. This irregular spike activity (and thus, presumably, irregular activation of the synapses of that neuron onto postsynaptic targets) can be approximated by stimulation with Poisson-distributed interstimulus intervals (Poisson stimulation). Therefore, if low-frequency presynaptic spike activity in the intact neocortex is sufficient to induce a generalized LTD of synaptic transmission, then Poisson stimulation, which mimics this spike activity, should induce LTD in slices. We tested this hypothesis by comparing changes in the strength of synapses onto layer 2/3 pyramidal cells induced by regular and Poisson stimulation in slices from adult visual cortex. We find that regular stimulation induces LTD of excitatory synaptic transmission as assessed by field potentials and intracellular postsynaptic potentials (PSPs) with inhibition absent. However, Poisson stimulation does not induce a net LTD of excitatory synaptic transmission. When the PSP contained an inhibitory component, neither Poisson nor regular stimulation induced LTD. We propose that the short bursts of synaptic activity that occur during a Poisson train have potentiating effects that offset the induction of LTD that is favored with regular stimulation. Thus, natural (i.e., irregular) low-frequency activity in the adult neocortex in vivo should not consistently induce LTD. PMID- 11264307 TI - Afferent input is necessary for seasonal growth and maintenance of adult avian song control circuits. AB - The neural circuits that regulate song behavior in adult songbirds undergo pronounced seasonal changes in morphology, primarily in response to changes in plasma testosterone (T). Most song nuclei have T receptors. We asked whether seasonal growth and maintenance of nuclei within these circuits are direct responses to the effects of T or its metabolites or are mediated indirectly via the effects of T on afferent nuclei. Photosensitive white-crowned sparrows were exposed to one of three treatments. (1) The neostriatal nucleus HVc (also known as the "high vocal center") was lesioned unilaterally, and the birds were exposed to long-day (LD) photoperiods and breeding levels of T for 30 d. (2) Birds were exposed to LD plus T (LD+T) for 30 d; then HVc was lesioned, and the birds were killed after an additional 30 d exposure to LD+T. (3) HVc was lesioned, and the sparrows were housed on short-day (SD) photoperiods in the absence of T treatment for 30 d. In both LD+T groups, the direct efferent targets of HVc, the robust nucleus of the archistriatum (RA) and area X, were smaller ipsilateral to the lesion. The lesion did not prevent growth of the hypoglossal motor nucleus, which does not receive direct afferent input from HVc. RA and area X were also smaller ipsilateral to the lesion in the SD birds. These results indicate that afferent input is required both for the growth of adult song circuits in response to typical breeding photoperiod and hormone conditions and for the maintenance of efferent nuclei in either their regressed or enlarged states. PMID- 11264308 TI - Synapse formation is arrested in retinal photoreceptors of the zebrafish nrc mutant. AB - We describe the effects of a recessive mutation on visual behavior, the electroretinogram (ERG), and photoreceptor structure in zebrafish. At 6 d post fertilization (dpf), no optokinetic reflex could be elicited in no optokinetic response c (nrc) mutant animals under any test condition. The animals exhibited ERG responses at 5-7 dpf that were markedly abnormal and could be categorized into two groups. The first showed an initial negative a-wave followed by a delayed positive b-wave of small amplitude. Often a second ERG-like response was recorded after the initial b-wave. The second group showed only a large negative a-wave; an initial b-wave was not evident. In most recordings additional oscillatory waves varying in number, amplitude, and time course were observed. Multiple responses at the cessation of long-duration flashes were also observed. Light and electron microscopy revealed that the cone photoreceptor pedicles of nrc fish were highly abnormal. Although the appropriate number of synaptic ribbons formed in these terminals, they "floated" in the terminal, unassociated with postsynaptic processes or arciform densities. The few processes invaginating the nrc pedicles resembled those of horizontal cells. Invaginating bipolar cell processes were rare, but basal contacts were observed on pedicle surfaces. The severity of the mutation did not change between 6 and 8 dpf, showing that there is neither a delay in development nor a degeneration of the terminals; rather, nrc pedicle development appears arrested. Bipolar cell terminals in the inner plexiform layer made normal ribbon synapses; thus, the mutation appears to affect only the outer retina. PMID- 11264309 TI - GABA expression dominates neuronal lineage progression in the embryonic rat neocortex and facilitates neurite outgrowth via GABA(A) autoreceptor/Cl- channels. AB - GABA emerges as a trophic signal during rat neocortical development in which it modulates proliferation of neuronal progenitors in the ventricular/subventricular zone (VZ/SVZ) and mediates radial migration of neurons from the VZ/SVZ to the cortical plate/subplate (CP/SP) region. In this study we investigated the role of GABA in the earliest phases of neuronal differentiation in the CP/SP. GABAergic signaling components emerging during neuronal lineage progression were comprehensively characterized using flow cytometry and immunophenotyping together with physiological indicator dyes. During migration from the VZ/SVZ to the CP/SP, differentiating cortical neurons became predominantly GABAergic, and their dominant GABA(A) receptor subunit expression pattern changed from alpha4beta1gamma1 to alpha3beta3gamma2gamma3 coincident with an increasing potency of GABA on GABA(A) receptor-mediated depolarization. GABA(A) autoreceptor/Cl(-) channel activity in cultured CP/SP neurons dominated their baseline potential and indirectly their cytosolic Ca(2+) (Ca(2+)c) levels via Ca(2+) entry through L-type Ca(2+) channels. Block of this autocrine circuit at the level of GABA synthesis, GABA(A) receptor activation, intracellular Cl(-) ion homeostasis, or L-type Ca(2+) channels attenuated neurite outgrowth in most GABAergic CP/SP neurons. In the absence of autocrine GABAergic signaling, neuritogenesis could be preserved by depolarizing cells and elevating Ca(2+)c. These results reveal a morphogenic role for GABA during embryonic neocortical neuron development that involves GABA(A) autoreceptors and L-type Ca(2+) channels. PMID- 11264310 TI - Evidence for the involvement of Tiam1 in axon formation. AB - In cultured neurons, axon formation is preceded by the appearance in one of the multiple neurites of a large growth cone containing a labile actin network and abundant dynamic microtubules. The invasion-inducing T-lymphoma and metastasis 1 (Tiam1) protein that functions as a guanosine nucleotide exchange factor for Rac1 localizes to this neurite and its growth cone, where it associates with microtubules. Neurons overexpressing Tiam1 extend several axon-like neurites, whereas suppression of Tiam1 prevents axon formation, with most of the cells failing to undergo changes in growth cone size and in cytoskeletal organization typical of prospective axons. Cytochalasin D reverts this effect leading to multiple axon formation and penetration of microtubules within neuritic tips devoid of actin filaments. Taken together, these results suggest that by regulating growth cone actin organization and allowing microtubule invasion within selected growth cones, Tiam1 promotes axon formation and hence participates in neuronal polarization. PMID- 11264311 TI - Short-range guidance of olfactory bulb axons is independent of repulsive factor slit. AB - During development, mitral cells, the major output neurons of the olfactory bulb, project their axons caudolaterally into the telencephalon and form the lateral olfactory tract (LOT). Two types of guidance cues have been suggested for this projection. First, a long-range factor Slit, which is secreted from the septum, repels mitral cell axons into a caudolateral direction. Second, the pathway of mitral cell axons contains a subset of neurons designated as lot cells, which guide the axons through short-range interactions. It is not clear how these two guidance cues relate to each other and how they share the physiological roles. Here we examined the behavior of mitral cell axons in organotypic culture on ectopic application of Slit and inhibition of endogenous Slit signaling. The results suggested that the short-range guidance cue in the LOT pathway functions independently from Slit. Furthermore, our results showed that removal of the septum and inhibition of Slit signaling did not affect the projection of mitral cell axons. Although the septum and exogenous Slit can repel olfactory bulb axons, our results cast doubts on the physiological relevance of the septum and endogenous Slit in guiding the projection of mitral cell axons. PMID- 11264312 TI - Formation and function of synapses with respect to Schwann cells at the end of motor nerve terminal branches on mature amphibian (Bufo marinus) muscle. AB - A study has been made of the formation and regression of synapses with respect to Schwann cells at the ends of motor nerve terminal branches in mature toad (Bufo marinus) muscle. Synapse formation and regression, as inferred from the appearance and loss of N-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styryl) pyridinium dibromide (FM1-43)-stained vesicle clusters, occurred at the ends of terminal branches over a 16 hr period. Multiple microelectrodes placed in an array about FM1-43 blobs at the ends of terminal branches detected the electrical signs of neurotransmitter being released onto receptors. Injection of a calcium indicator (Oregon Green 488 BAPTA-1) into the motor nerve with subsequent imaging of the calcium transients, in response to stimulation, often showed a reduced calcium influx in the ends of terminal branches. Injection of a fluorescent dye into motor nerves revealed the full extent of their terminal branches and growing processes. Injection of the terminal Schwann cells (TSCs) often revealed pseudopodial TSC processes up to 10-microm-long. Imaging of these TSC processes over minutes or hours showed that they were highly labile and capable of extending several micrometers in a few minutes. Injection of motor nerve terminals with a different dye to that injected into their TSCs revealed that terminal processes sometimes followed the TSC processes over a few hours. It is suggested that the ends of motor nerve terminals in vivo are in a constant state of remodeling through the formation and regression of processes, that TSC processes guide the remodeling, and that it can occur over a relatively short period of time. PMID- 11264313 TI - Dendritic spines lost during glutamate receptor activation reemerge at original sites of synaptic contact. AB - During cerebral ischemia, neurons undergo rapid alterations in dendritic structure consisting of focal swelling and spine loss. We used time-lapse microscopy to determine the fate of dendritic spines that disappeared after brief, sublethal hypoxic or excitotoxic exposures. Dendrite and spine morphology were assessed in cultured cortical neurons expressing yellow fluorescent protein or labeled with the fluorescent membrane tracer, DiI. Neurons exposed to NMDA, kainate, or oxygen-glucose deprivation underwent segmental dendritic beading and loss of approximately one-half of dendritic spines. Most spine loss was observed in regions of local dendritic swelling. Despite widespread loss, spines recovered within 2 hr after termination of agonist exposure or oxygen-glucose deprivation and remained stable over the subsequent 24 hr. Recovery was slower after NMDA than AMPA/kainate receptor activation. Time-lapse fluorescence imaging showed that the vast majority of spines reemerged in the same location from which they disappeared. In addition to spine recovery, elaboration of dendritic filopodia was observed in new locations along the dendritic shaft after dendrite recovery. Spine recovery did not depend on actin polymerization because it was not blocked by application of latrunculin-A, which eliminated filamentous actin staining in spines and blocked spine motility. Throughout spine loss and recovery, presynaptic and postsynaptic elements remained in physical proximity. These results suggest that elimination of dendritic spines is not necessarily associated with loss of synaptic contacts. Rapid reestablishment of dendritic spine synapses in surviving neurons may be a substrate for functional recovery after transient cerebral ischemia. PMID- 11264314 TI - Long-term memory is facilitated by cAMP response element-binding protein overexpression in the amygdala. AB - At least two temporally and mechanistically distinct forms of memory are conserved across many species: short-term memory that persists minutes to hours after training and long-term memory (LTM) that persists days or longer. In general, repeated training trials presented with intervening rest intervals (spaced training) is more effective than massed training (the same number of training trials presented with no or short intervening rest intervals) in producing LTM. LTM requires de novo protein synthesis, and cAMP response element binding protein (CREB) may be one of the transcription factors regulating the synthesis of new proteins necessary for the formation of LTM. Here we show that rats given massed fear conditioning training show no or weak LTM, as measured by fear-potentiated startle, compared with rats given the same amount of training but presented in a spaced manner. Increasing CREB levels specifically in the basolateral amygdala via viral vector-mediated gene transfer significantly increases LTM after massed fear training. The enhancing effect of CREB overexpression on LTM formation is shown to be specific in terms of biochemistry, anatomy, time course, and the training procedure used. These results suggest that CREB activity in the amygdala serves as a molecular switch for the formation of LTM in fear conditioning. PMID- 11264315 TI - Leptin-induced nuclear translocation of STAT3 immunoreactivity in hypothalamic nuclei involved in body weight regulation. AB - Leptin is involved in the hypothalamic control of food intake and body weight. Fos immunohistochemistry has been used to functionally map leptin target neurons involved in these regulatory processes. However, only a subset of hypothalamic neurons expressing the long form of the leptin receptor (Ob-Rb) also coexpress the neuronal activation marker Fos after leptin stimulation. To functionally map all leptin target neurons, regardless of whether leptin-mediated neuronal activation or inhibition occurs, we immunohistochemically investigated the leptin induced nuclear translocation of the signal transducer and activator of transcription molecule STAT3, which represents a crucial step in the regulation of leptin-dependent gene expression. As proven by colocalization studies with the nuclear 4',6-diamidino-2-phenylindole dilactate stain, intracerebroventricular leptin treatment, but not intracerebroventricular application of pyrogen-free saline, induced a time-dependent nuclear translocation of STAT3 immunoreactivity in hypothalamic nuclei, with strong nuclear STAT3 signals detectable in the arcuate nucleus, the lateral hypothalamus, and the ventromedial and dorsomedial hypothalamic nuclei. This leptin-induced STAT3 translocation pattern proved to be distinct from that induced by interleukin-6, another cytokine using STAT3 in its signaling pathway. Combined immunohistochemical STAT3 and Fos detection after leptin treatment revealed a higher number of STAT3-positive than Fos-positive cell nuclei in the aforementioned hypothalamic structures and showed that Fos immunoreactivity colocalized only in a subset of all leptin-responsive STAT3 nuclei. These results suggest that the detection of nuclear STAT3 immunoreactivity represents a new neuroanatomical tool to functionally map central leptin actions. They further support the importance of ventrally located caudal hypothalamic structures representing the main leptin targets involved in body weight regulation. PMID- 11264316 TI - Agonist-induced internalization and trafficking of cannabinoid CB1 receptors in hippocampal neurons. AB - Agonist-induced internalization of G-protein-coupled receptors is an important mechanism for regulating receptor abundance and availability at the plasma membrane. In this study we have used immunolabeling techniques and confocal microscopy to investigate agonist-induced internalization and trafficking of CB(1) receptors in rat cultured hippocampal neurons. The levels of cell surface CB(1) receptor immunoreactivity associated with presynaptic GABAergic terminals decreased markedly (by up to 84%) after exposure to the cannabinoid agonist (+) WIN55212, in a concentration-dependent (0.1-1 microm) and stereoselective manner. Inhibition was maximal at 16 hr and abolished in the presence of SR141716A, a selective CB(1) receptor antagonist. Methanandamide (an analog of an endogenous cannabinoid, anandamide) also reduced cell surface labeling (by 43% at 1 microm). Differential labeling of cell surface and intracellular pools of receptor demonstrated that the reduction in cell surface immunoreactivity reflects agonist induced internalization and suggests that the internalized CB(1) receptors are translocated toward the soma. The internalization process did not require activated G-protein alpha(i) or alpha(o) subunits. A different pattern of cell surface CB(1) receptor expression was observed using an undifferentiated F-11 cell line, which had pronounced somatic labeling. In these cells substantial CB(1) receptor internalization was also observed after exposure to (+)-WIN55212 (1 microm) for relatively short periods (30 min) of agonist exposure. In summary, this dynamic modulation of CB(1) receptor expression may play an important role in the development of cannabinoid tolerance in the CNS. Agonist-induced internalization at presynaptic terminals has important implications for the modulatory effects of G-protein-coupled receptors on neurotransmitter release. PMID- 11264317 TI - In vitro eye-blink classical conditioning is NMDA receptor dependent and involves redistribution of AMPA receptor subunit GluR4. AB - The classically conditioned vertebrate eye-blink response is a model in which to study neuronal mechanisms of learning and memory. A neural correlate of this response recorded in the abducens nerve can be conditioned entirely in vitro using an isolated brainstem-cerebellum preparation from the turtle by pairing trigeminal and auditory nerve stimulation. Here it is reported that conditioning requires that the paired stimuli occur within a narrow temporal window of <100 msec and that it is blocked by the NMDA receptor antagonist d,l-2-amino-5 phosphonovaleric acid. Moreover, there is a significant positive correlation between the levels of conditioning and greater immunoreactivity with the glutamate receptor 4 (GluR4) AMPA receptor subunit in the abducens motor nuclei, but not with NMDAR1 or GluR1. It is concluded that in vitro classical conditioning of an abducens nerve eye-blink response is generated by NMDA receptor-mediated mechanisms that may act to modify the AMPA receptor by increasing GluR4 subunits in auditory nerve synapses. PMID- 11264318 TI - Behavioral and anatomical correlates of chronic episodic hypoxia during sleep in the rat. AB - The role played by chronic episodic hypoxia (EHYP) in the neurocognitive morbidity of obstructive sleep apnea (OSA) is unknown. Sleep recordings, Morris water maze experiments, and immunohistochemistry for NMDA NR1 glutamate receptor, c-fos protein, and apoptosis [nuclear immunoreactivity for single-stranded DNA and terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling assay] were conducted in EHYP-exposed Sprague Dawley male rats. Exposures consisted of up to14 d in an environmental chamber in which O(2) concentrations were cycled between 10 and 21% every 90 sec or 30 min during 12 hr of daylight. For the remaining 12 hr, EHYP rats breathed room air, while controls spent 14 d in room air. Although EHYP induced significant disruption of sleep architecture during the initial day of exposure, sleep patterns normalized thereafter. Marked increases in apoptosis occurred in the CA1 hippocampal region (sevenfold) and cortex (Cx; eightfold) after 1-2 d of EHYP but not in CA3 and were followed by decreases toward normoxic levels by 14 d. Double labeling for NMDA NR1 and c-fos revealed marked architectural disorganization in CA1 and Cx with increases in c-fos over time. Rats exposed to EHYP displayed significantly longer escape latencies and swim path lengths to escape a hidden platform during 12 training trials given over 2 d. Differences in the performances of EHYP and control rats, although reduced, persisted after 14 d of recovery. We conclude that EHYP is associated with marked cellular changes over time within neural regions associated with cognitive functions. Furthermore, EHYP impaired performance during acquisition of a cognitive spatial task without affecting sensorimotor function. Such changes may underlie components of the learning and memory impairments found in OSA. PMID- 11264319 TI - Human brain regions involved in heading estimation. AB - Observer motion in a stationary visual environment results in an optic flow pattern on the retina, which in simple situations can be used to determine the direction of self motion or heading. The present study, using positron emission tomography (PET) and functional magnetic resonance imaging (fMRI), investigated the human cerebral activation pattern, elicited when subjects viewing a ground plane optic flow pattern actively judged heading. Several successive experiments controlled for visual input, visuospatial attention, and motor response effects. Results indicate that the network specifically involved in heading consists of only two motion sensitive areas: human MT/V5+, including an inferior satellite, and dorsal intraparietal sulcus area (DIPSM/L), predominantly in the right hemisphere, plus a dorsal premotor region bilaterally. These results suggest possible homologies with the dorsal part of the medial superior temporal area and area 7a in the monkey. PMID- 11264320 TI - Coincidence detection or temporal integration? What the neurons in somatosensory cortex are doing. AB - To assess the impact of thalamic synchronization on cortical responsiveness, we used conditional cross-correlation analysis to measure the probability of neuronal discharges in somatosensory cortex as a function of the time between discharges in pairs of simultaneously recorded neurons in the ventrobasal thalamus. Among 26 neuronal trios, we found that thalamocortical efficacy after synchronous thalamic activity was nearly twice as large as the efficacy rate obtained when pairs of thalamic neurons discharged asynchronously. Nearly half of these neuronal trios displayed cooperative effects in which the cortical discharge probability after synchronous thalamic events was larger than could be predicted from the efficacy rate of individual thalamic discharges. In these cases of heterosynaptic cooperativity, thalamocortical efficacy declined to asymptotic levels when the interspike intervals were >6-8 msec. These results indicate that thalamic synchronization has a significant impact on cortical responsiveness and suggest that neuronal synchronization may play a critical role in the transmission of sensory information from one brain region to another. PMID- 11264321 TI - Sex difference and steroid modulation of pheromone-induced immediate early genes in the two zones of the mouse accessory olfactory system. AB - Two anatomically and neurochemically distinct zones within the vomeronasal organ (VNO) and accessory olfactory bulb (AOB) have been identified that are responsible for the detection of pheromones. Using markers to distinguish between apical and basal neurons of the VNO neuroepithelium and rostral versus caudal AOB glomeruli, we examined immediate early gene immunoreactivity (IEG-IR) in gonadectomized, steroid-treated mice in response to pheromones of male and female conspecifics. After exposure of estradiol-treated females to soiled male bedding, more VNO neurons in the basal than the apical layer exhibited IEG-IR compared with VNO neurons of estradiol-treated males. Conversely, whereas soiled female bedding failed to induce IEG-IR in VNO neurons of estradiol-treated males or females, both apical and basal neurons were activated in testosterone-treated males. Male and female pheromones also activated mitral and granule cells in the AOBs of all subjects, but responses to different pheromones were distributed across the boundary of the rostral and caudal regions. These data show that differences in the response of males and females to the same pheromonal stimulus are found in the sensory neurons of the VNO. We propose that centrifugal, noradrenergic inputs to VNO neurons, which may differ in the two sexes and respond differently to adult sex steroids, modulate sensitivity to pheromonal stimulation. PMID- 11264322 TI - Olfactory fingerprints for major histocompatibility complex-determined body odors. AB - Recognition of individual body odors is analogous to human face recognition in that it provides information about identity. Individual body odors determined by differences at the major histocompatibility complex (MHC or H-2) have been shown to influence mate choice, pregnancy block, and maternal behavior in mice. Unfortunately, the mechanism and extent of the main olfactory bulb (MOB) and accessory olfactory bulb (AOB) involvement in the discrimination of animals according to H-2-type has remained ambiguous. Here we study the neuronal activation patterns evoked in the MOB in different individuals on exposure to these complex, biologically meaningful sensory stimuli. We demonstrate that body odors from H-2 disparate mice evoke overlapping but distinct maps of neuronal activation in the MOB. The spatial patterns of odor-evoked activity are sufficient to be used like fingerprints to predict H-2 identity using a novel computer algorithm. These results provide functional evidence for discrimination of H-2-determined body odors in the MOB, but do not preclude a role for the AOB. These data further our understanding of the neural strategies used to decode socially relevant odors. PMID- 11264323 TI - The synaptic architecture of AMPA receptors at the cone pedicle of the primate retina. AB - Cone pedicles, the output synapses of cone photoreceptors, transfer the light signal onto the dendrites of bipolar and horizontal cells. Cone pedicles contain between 20 and 45 ribbon synapses (triads) which are the release sites for glutamate, the cone transmitter. Several hundred postsynaptic dendrites contact individual cone pedicles, and we studied the glutamate receptors expressed and clustered at these contacts, particularly the AMPA receptor subunits. Using immunocytochemistry and confocal imaging we were able to resolve individual triads within the cone pedicles by light microscopy. We studied their differences in L/M- and S-cones, and we counted the number of triads per pedicle across the retina. The presynaptic matrix protein bassoon, the synapse-associated membrane protein P84, and peanut agglutinin were used to specifically label synaptic ribbons, invaginating dendrites of horizontal cells and invaginating dendrites of ON-cone bipolar cells, respectively. Pre- and post-embedding immunocytochemistry and electron microscopy were used to localize the AMPA receptor subunits at the cone pedicle base. They were aggregated at three different postsynaptic sites: at horizontal cell invaginating contacts, at bipolar cell flat contacts, and at desmosome-like junctions underneath the cone pedicles. We also performed double labeling experiments with the triad-specific markers and the antibodies against the AMPA receptor subunits. AMPA receptors were preferentially expressed by horizontal cells, and to a lesser extent by OFF-cone bipolar cells. We did not observe any cone-selective expression of AMPA receptor subunits postsynaptic to L/M- or S-cones, suggesting AMPA receptors are not the key to understanding trichromatic signaling in the primate retina. PMID- 11264324 TI - Postlearning consolidation of birdsong: stabilizing effects of age and anterior forebrain lesions. AB - Birdsong is a learned, sequenced motor skill. For the zebra finch, learned song normally remains unchanging beyond early adulthood. However, stable adult song will gradually deteriorate after deafening (Nordeen and Nordeen, 1992), indicating an ongoing influence of auditory feedback on learned song. This plasticity of adult song in response to deafening gradually declines with age (Lombardino and Nottebohm, 2000), suggesting that, after song learning, there continue to be changes in the brain that progressively stabilize the song motor program. A qualitatively similar stabilization of learned song can be precipitated artificially by lesions of a basal ganglia circuit in the songbird anterior forebrain (Brainard and Doupe, 2000), raising the question of whether and how these two forms of song stabilization are related. We investigated this issue by characterizing the deterioration of song that occurs after deafening in young adult birds and the degree to which that deterioration is reduced by age or by lesions of the anterior forebrain that were directed at the lateral portion of the magnocellular nucleus of the anterior neostriatum (LMAN). In most respects, LMAN lesions stabilized song to a significantly greater extent than did aging; whereas old-deafened birds eventually exhibited significant deterioration of song, lesioned-deafened birds generally did not differ from controls. The one exception was for song tempo, which was significantly stabilized by age, but not by LMAN lesions. The results indicate that LMAN lesions do not simply mimic a normal aging process, and likewise suggest that the anterior forebrain pathway continues to play a role even in the residual song plasticity that is observed after the age-dependent stabilization of song. PMID- 11264325 TI - Basolateral amygdala-nucleus accumbens interactions in mediating glucocorticoid enhancement of memory consolidation. AB - Systemic or intracerebral administration of glucocorticoids enhances memory consolidation in several tasks. Previously, we reported that these effects depend on an intact basolateral nucleus of the amygdala (BLA) and efferents from the BLA that run through the stria terminalis (ST). The BLA projects directly to the nucleus accumbens (NAc) via this ST pathway. The NAc also receives direct projections from the hippocampus and, therefore, may be a site of convergence of BLA and hippocampal influences in modulating memory consolidation. In support of this view, we found previously that lesions of either the NAc or the ST also block the memory-modulatory effect of systemically administered glucocorticoids. The present experiments examined the effects of lesions of the NAc or the ST on the memory-modulatory effects of intracerebral glucocorticoids on inhibitory avoidance training. Microinfusions of the specific glucocorticoid receptor agonist 11beta,17beta-dihydroxy-6,21-dimethyl-17alpha-pregna-4,6-trien-20yn-3-one (RU 28362; 1.0 or 3.0 ng) into either the BLA or the hippocampus of male Sprague Dawley rats administered immediately after training enhanced the 48 hr retention performance in a dose-dependent manner. Bilateral lesions of the NAc or the ST alone did not affect retention performance but blocked the memory enhancement induced by intra-BLA or intrahippocampal glucocorticoid receptor agonist administration. These findings indicate that the BLA-NAc pathway plays an essential role in mediating glucocorticoid effects on memory consolidation and suggest that the BLA interacts with hippocampal effects on memory consolidation via this pathway. PMID- 11264326 TI - Limbic-cortical-ventral striatal activation during retrieval of a discrete cocaine-associated stimulus: a cellular imaging study with gamma protein kinase C expression. AB - We investigated the neuronal activation associated with reexposure to a discrete cocaine-associated stimulus using in situ hybridization to quantify the expression of the plasticity-regulated gene, gamma protein kinase C (gamma PKC), in the limbic-cortical-ventral striatal system. Groups of rats were trained to self-administer cocaine paired with a light stimulus (Paired) or paired with an auditory stimulus but also receiving light presentations yoked to those in the Paired group (Unpaired). Additional groups received noncontingent cocaine-light pairings (Pavlovian) or saline-light pairings (Saline) that were yoked to the Paired group. After acquisition of self-administration by the Paired and Unpaired groups, all groups had a 3 d drug- and training-free period before being reexposed to noncontingent presentations of the light conditioning stimulus during a 5 min test session in the training context. There were four major patterns of results for regional gamma PKC expression 2 hr later. (1) Changes occurred only in groups in which the light was predictive of cocaine. (2) Increases were seen in the amygdala, but decreases were seen in the medial prefrontal cortex. (3) No changes were seen in the hippocampus. (4) Although changes were observed in the basal and central nuclei of the amygdala and the prelimbic cortex in both the Paired and Pavlovian groups, additional changes were observed in the nucleus accumbens core, lateral amygdala, and anterior cingulate cortex in the Pavlovian group. These results suggest not only that regionally selective alterations in gamma PKC expression are an index of the retrieval of Pavlovian associations formed between a drug and a discrete stimulus, but also that a distinct neural circuitry may underlie Pavlovian stimulus-reward associations in cocaine-experienced rats. PMID- 11264327 TI - Contribution of endogenous enkephalins to the enhanced analgesic effects of supraspinal mu opioid receptor agonists after inflammatory injury. AB - This study examined a mechanism responsible for the enhanced antihyperalgesic and antinociceptive effects of the mu opioid receptor agonist (ORA) [D-Ala(2), NMePhe(4), Gly(5)-ol]enkephalin (DAMGO) microinjected in the rostroventromedial medulla (RVM) of rats with inflammatory injury induced by injection of complete Freund's adjuvant (CFA) in one hindpaw. In rats injected with CFA 4 hr earlier, microinjection of the mu opioid receptor antagonist D-Phe-Cys-Tyr-D-Trp-Arg-Thr Pen-Thr-NH(2) (CTAP) in the RVM antagonized both the marginal enhancement of the potency of DAMGO and its antinociceptive effect. The delta opioid receptor antagonist naltriben (NTB) was without effect. In rats injected with CFA 2 weeks earlier, CTAP antagonized the effects of DAMGO to a lesser extent. However, NTB completely prevented the enhancement of the potency of DAMGO, whereas it did not antagonize DAMGO's antinociceptive effects. Microinjection of NTB alone, but not CTAP in the RVM of CFA-treated rats, enhanced the hyperalgesia present in the ipsilateral hindpaw and induced hyperalgesia in the contralateral, uninjured hindpaw. These results suggest that persistent inflammatory injury increased the release in the RVM of opioid peptides with preferential affinity for the delta opioid receptor, which can interact in a synergistic or additive manner with an exogenously administered mu opioid receptor agonist. Indeed, the levels of [Met(5)]enkephalin and [Leu(5)]enkephalin were increased in the RVM and in other brainstem nuclei in CFA-treated rats. This increase most likely presents a compensatory neuronal response of the CNS of the injured animal to mitigate the full expression of inflammatory pain and to enhance the antinociceptive and antihyperalgesic effects of exogenously administered mu opioid receptor analgesics. PMID- 11264328 TI - CNS region-specific oxytocin receptor expression: importance in regulation of anxiety and sex behavior. AB - The oxytocin receptor (OTR) is differentially expressed in the CNS. Because there are multiple mechanisms by which the OTR can be transcriptionally induced, we hypothesized that differences in OTR expression may be explained by activation of distinct signal transduction pathways and may be critical for the control of anxiety and sex behaviors. To determine the regulation and functional significance of this expression, we infused female rats with modifiers of protein kinases before assaying for behavior and oxytocin receptor binding. In the ventromedial nucleus of the hypothalamus (VMH), estrogen-dependent induction of oxytocin receptors required protein kinase C activation, and oxytocin infused here promoted female sex behavior but had no effect on anxiety. In contrast, dopamine controlled tonic oxytocin receptor expression in the central nucleus of the amygdala (cAmyg) through activation of protein kinase A, and oxytocin infused here was anxiolytic but had no effect on female sex behavior. Therefore, we have identified brain region-specific regulation of the OTR in the VMH and cAmyg. Distinct signal transduction pathways regulating receptor expression and binding in each brain region may mediate in part the ability of oxytocin to exert these differential behavioral effects. PMID- 11264329 TI - Extinction training regulates tyrosine hydroxylase during withdrawal from cocaine self-administration. AB - Chronic exposure to drugs of abuse is known to modulate tyrosine hydroxylase (TH) levels in the mesolimbic dopamine system. In this study, 12 d of cocaine self administration in rats (4 hr/d) reduced TH immunoreactivity by 29% in the nucleus accumbens (NAc) shell, but not core, after a 1 week withdrawal period. In contrast, TH immunoreactivity in the NAc was completely restored in animals that experienced extinction training (4 hr/d) during the same withdrawal period. Extinction training also increased TH levels in the ventral tegmental area (VTA) by 45%, whereas TH was not altered in the VTA by cocaine withdrawal alone. Thus, extinction-induced normalization of NAc TH levels could involve increased TH synthesis, stability, and/or transport from the VTA to the NAc. A similar extinction training regimen failed to alter TH levels in the NAc or VTA of rats trained to self-administer sucrose pellets, indicating that TH regulation in cocaine-trained animals is not a generalized effect of extinction learning per se. Rather, these data suggest that neuroadaptative responses during cocaine withdrawal ultimately are determined by a complex interaction between chronic drug exposure and drug-seeking experience. The ability of extinction training to restore NAc TH levels is hypothesized to accelerate recovery from dopamine depletion and anhedonia during cocaine withdrawal. PMID- 11264330 TI - The in vitro fate of rabbit fetal brain cells after acute in vivo hypoxia. AB - In the investigation of ischemia-induced brain damage, traditional methods using histopathology estimate brain cell death at a time remote from ischemic insult. These observations fail to take into account endogenous repair processes or ongoing injury cascades like apoptosis. The cells that are injured but not killed initially are the population most amenable to rescue. The hypothesis was that in vivo uterine ischemia-reperfusion would result in more cell death and apoptosis in fetal brain cells cultured in vitro. Near-term, 29 d gestation, pregnant New Zealand White rabbits were subjected to repetitive uterine ischemia for a cumulative time of 40 min ischemia and 20 min reperfusion. Immediately after uterine ischemia, the fetal brains were removed and dissociated into a cell suspension. The ischemic group had more cell death than non-ischemic controls as assessed by Trypan Blue exclusion and propidium iodide (PI) uptake on a flow cytometer. Aliquots of cells were plated and cultured for 24 and 48 hr. The ischemic group had significantly more cell death (propidium iodide) than non ischemic controls at 24 hr and significantly more apoptosis, as assessed by annexin-V binding in cells at 24 hr and caspase-3 activity at 48 hr. Fewer cells attached to the culture plates at 48 hr in the ischemia group. After uterine ischemia, certain fetal brain cells die immediately, and other cells undergo ongoing damage resulting in necrosis and apoptosis that is manifest later. This method offers insight into the fate of those cells and provides a tool for assessing interventions to decrease cell injury. PMID- 11264331 TI - Effect of patient gender on outcome in two forms of short-term individual psychotherapy. AB - This study examined the relationship of patient gender and outcome for two forms (interpretive, supportive) of short-term, individual psychotherapy. Female and male patients (N=89) were randomly assigned to either interpretive or supportive therapy. Outcome was measured in the areas of depression, anxiety, and general symptomatic distress. A significant interaction effect between patient gender and form of therapy was found for measures of depression and general symptomatic distress at post-therapy. Male patients had better outcome in interpretive therapy than in supportive therapy. Female patients had better outcome in supportive therapy than in interpretive therapy. The findings suggest that patient gender may be differentially influential with different forms of short term therapy. PMID- 11264332 TI - Early transference interventions with male patients in psychotherapy. AB - Transcripts of early sessions for 7 personality-disordered male subjects participating in an ongoing naturalistic long-term dynamic psychotherapy project were rated for therapist interventions and alliance. Early transference interpretations were followed by increased defensiveness even when there was a solid alliance. Omitting transference interpretations in the face of an early negative transference was equally problematic. However, the rapid sequence of early transference and defense interpretations, or early defense interpretations alone, enhanced therapeutic work without increasing defensiveness. Caretaking of the alliance after early interpretive work was also investigated. Two different styles of handling affect emerged from the sample. PMID- 11264334 TI - Patterns of consistency and deviation in therapists' countertransference feelings. AB - The author addressed the question of consistency in psychotherapists' countertransference feelings. Research findings have indicated that the therapist's own personal feeling style may be more important than the patient's impact on the therapist's feelings. In this study, the feelings of 9 psychotherapists toward 28 patients were followed by using checklist self-report after each session during moderately long psychotherapies. ANOVAs and discriminant analyses showed that the therapists were very consistent in their feeling style over different patients and over time. The consistency in feelings toward the individual patients was smaller. Deviations from consistency are analyzed, and their importance for the understanding of different aspects of the countertransference is discussed. It is suggested that a meaningful use of the countertransference concept ought to be based on systematic identifications of recurrent and deviant patterns in the therapist's reactions. PMID- 11264333 TI - Adding group psychotherapy to medication treatment in dysthymia: a randomized prospective pilot study. AB - Patients with dysthymia have been shown to respond to treatment with antidepressant medications, and to some degree to psychotherapy. Even patients successfully treated with medication often have residual symptoms and impaired psychosocial functioning. The authors describe a prospective randomized 36-week study of dysthymic patients, comparing continued treatment with antidepressant medication (fluoxetine) alone and medication with the addition of group therapy treatment. After an 8-week trial of fluoxetine, medication-responsive subjects were randomly assigned to receive either continued medication only or medication plus 16 sessions of manualized group psychotherapy. Results provide preliminary evidence that group therapy may provide additional benefit to medication responding dysthymic patients, particularly in interpersonal and psychosocial functioning. PMID- 11264336 TI - Interpersonal psychotherapy adapted for the group setting in the treatment of postpartum depression. AB - Interpersonal psychotherapy (IPT) has demonstrated efficacy in the individual treatment of antepartum and postpartum depression. The current investigation extends prior work by examining the efficacy of a group IPT approach for the treatment of postpartum depression. Depression scores of 17 women diagnosed with postpartum depressive disorder (DSM-IV criteria) decreased significantly from pre to post-treatment. Follow-up assessments at 6 months revealed continuation of the treatment effect. Results indicate that IPT adapted for a group model has positive implications for the treatment of postpartum depression, demonstrating both short-term and longer-term effects in the reduction of depressive symptomatology. Study limitations include the small sample size, absence of control group, possible bias in therapist's assessments, and lack of monitoring adherence, which may have jeopardized the accuracy of the results. PMID- 11264335 TI - A randomized controlled trial comparing moclobemide and moclobemide plus interpersonal psychotherapy in the treatment of dysthymic disorder. AB - The authors compared the outcomes of 35 outpatients with dysthymic disorder randomized to receive either treatment with moclobemide and interpersonal therapy (IPT) or moclobemide and routine clinical management. Diagnosis was based on the ICD-10 symptom checklist. Patients were evaluated by trained raters using the 17 item Hamilton Rating Scale for Depression (Ham-D), Montgomery-Asberg Depression Rating Scale (MADRS), Global Assessment of Functioning, and Quality of Life and Satisfaction Questionnaire at baseline, 12, 24, and 48 weeks. Patients in both treatment groups showed statistically significant improvement in all measures across time. There was a nonsignificant trend toward lower scores on Ham-D and MADRS for patients in the moclobemide plus IPT group. Longer, better-powered trials should be carried out to study the efficacy of IPT plus antidepressant medication in the treatment of dysthymic disorder. PMID- 11264337 TI - Attachment theory and psychotherapy research. PMID- 11264339 TI - Human dendritic cells are activated by dengue virus infection: enhancement by gamma interferon and implications for disease pathogenesis. AB - The ability of dendritic cells (DCs) to shape the adaptive immune response to viral infection is mediated largely by their maturation and activation state as determined by the surface expression of HLA molecules, costimulatory molecules, and cytokine production. Dengue is an emerging arboviral disease where the severity of illness is influenced by the adaptive immune response to the virus. In this report, we have demonstrated that dengue virus infects and replicates in immature human myeloid DCs. Exposure to live dengue virus led to maturation and activation of both the infected and surrounding, uninfected DCs and stimulated production of tumor necrosis factor alpha (TNF-alpha) and alpha interferon (IFN alpha). Activation of the dengue virus-infected DCs was blunted compared to the surrounding, uninfected DCs, and dengue virus infection induced low-level release of interleukin-12 p70 (IL-12 p70), a key cytokine in the development of cell mediated immunity (CMI). Upon the addition of IFN-gamma, there was enhanced activation of dengue virus-infected DCs and enhanced dengue virus-induced IL-12 p70 release. The data suggest a model whereby DCs are the early, primary target of dengue virus in natural infection and the vigor of CMI is modulated by the relative presence or absence of IFN-gamma in the microenvironment surrounding the virus-infected DCs. These findings are relevant to understanding the pathogenesis of dengue hemorrhagic fever and the design of new vaccination and therapeutic strategies. PMID- 11264340 TI - Sequence requirements for Sindbis virus subgenomic mRNA promoter function in cultured cells. AB - The Sindbis virus minimal subgenomic mRNA promoter (spanning positions -19 to +5 relative to the subgenomic mRNA start site) is approximately three- to sixfold less active than the fully active -98 to +14 promoter region. We identified two elements flanking the -19 to +5 region which increase its transcription to levels comparable to the -98 to +14 region. These elements span positions -40 to -20 and +6 to +14 and act synergistically to enhance transcription. Nine different virus libraries were constructed containing blocks of five randomized nucleotides at various positions in the -40 to +14 region. On passaging these libraries in mosquito cells, a small subset of the viruses came to dominate the population. Sequence analysis at the population level and for individual clones revealed that in general, wild-type bases were preferred for positions -15 to +5 of the minimal promoter. Base mutagenesis experiments indicated that the selection of wild-type bases in this region was primarily due to requirements for subgenomic mRNA transcription. Outside of the minimal promoter, the -35 to -29 region contained four positions which also preferred wildtype bases. However, the remaining positions generally preferred non-wild-type bases. On passaging of the virus libraries on hamster cells, the -15 to +5 region again preferred the wild-type base but most of the remaining positions exhibited almost no base preference. The promoter thus consists of an essential central region from -15 to +5 and discrete flanking sites that render it fully active, depending on the host environment. PMID- 11264341 TI - Two functionally distinct forms of a retroviral receptor explain the nonreciprocal receptor interference among subgroups B, D, and E avian leukosis viruses. AB - Subgroups B, D, and E avian leukosis viruses (ALV-B, -D, and -E) share the same chicken receptor, TVB(S1), a tumor necrosis factor receptor (TNFR)-related protein. These viruses, however, exhibit nonreciprocal receptor interference (NRI): cells preinfected with ALV-B or ALV-D are resistant to superinfection by viruses of all three subgroups, whereas those pre-infected by ALV-E are resistant only to superinfection by other subgroup E viruses. In this study, we investigated the basis of this phenomenon by characterizing the interaction of TVB(S1) with ALV-B Env or ALV-E Env. Sequential immunoprecipitation analysis using surface envelope immunoglobulin fusion proteins revealed the existence of two separate types of TVB(S1) that are encoded by the same cDNA clone. One form, designated the type 1 receptor, is specific for ALV-B and ALV-E. The other form, the type 2 receptor, is specific for ALV-B. We show that a protein consisting of only the first and second extracellular cysteine-rich domains of TVB(S1) is capable of forming both receptor types. However, the third extracellular cysteine rich domain is required for efficient formation of the type 1 receptor. We also demonstrate that heterogeneous N-linked glycosylation cannot explain the difference in activities of the two receptor types. The existence of two types of TVB(S1) explains the NRI pattern between ALV-B and -E: subgroup B viruses establish receptor interference with both receptor types, whereas subgroup E viruses interfere only with the type 1 receptor, leaving the type 2 receptor available to mediate subsequent rounds of ALV-B entry. The formation of a TVB receptor type that is specific for cytopathic ALV may also have important implications for understanding how some subgroups of ALV cause cell death. PMID- 11264342 TI - Antiviral effect of N-butyldeoxynojirimycin against bovine viral diarrhea virus correlates with misfolding of E2 envelope proteins and impairment of their association into E1-E2 heterodimers. AB - The iminosugar N-butyldeoxynojirimycin (NB-DNJ), an endoplasmic reticulum alpha glucosidase inhibitor, has an antiviral effect against bovine viral diarrhea virus (BVDV). In this report, we investigate the molecular mechanism of this inhibition by studying the folding pathway of BVDV envelope glycoproteins in the presence and absence of NB-DNJ. Our results show that, while the disulfide dependent folding of E2 glycoprotein occurs rapidly (2.5 min), the folding of E1 occurs slowly (30 min). Both BVDV envelope glycoproteins associate rapidly with calnexin and dissociate with different kinetics. The release of E1 from the interaction with calnexin coincides with the beginning of E1 and E2 association into disulfide-linked heterodimers. In the presence of NB-DNJ, the interaction of E1 and E2 with calnexin is prevented, leading to misfolding of the envelope glycoproteins and inefficient formation of E1-E2 heterodimers. The degree of misfolding and the lack of association of E1 and E2 into disulfide-linked complexes in the presence of NB-DNJ correlate with the dose-dependent antiviral effect observed for this iminosugar. PMID- 11264343 TI - Direct and indirect regulation of cytokine and cell cycle proteins by EBNA-2 during Epstein-Barr virus infection. AB - We have studied the pathways of regulation of cytokine and cell cycle control proteins during infection of human B lymphocytes by Epstein-Barr virus (EBV). Among 30 cytokine RNAs analyzed by the RNase protection assay, tumor necrosis factor alpha (TNF-alpha), granulocyte colony-stimulating factor, lymphotoxin (LT), and LTbeta were found to be regulated within 20 h of EBV infection of primary B cells. Similar results were obtained using the estrogen-regulated EBNA 2 cell line EREB2.5, in which RNAs for LT and TNF-alpha were induced within 6 h of activation of EBNA-2. Expression of Notch also caused an induction of TNF alpha RNA. The induction of TNF-alpha RNA by EBNA-2 was indirect, and constitutive expression of either LMP-1 or c-myc proteins did not substitute for EBNA-2 in induction of TNF-alpha RNA. Cyclin D2 is also an indirect target of EBNA-2-mediated transactivation. EBNA-2 was found to activate the cyclin D2 promoter in a transient-transfection assay. A mutant of EBNA-2 that does not bind RBP-Jkappa retained some activity in this assay, and activation did not depend on the presence of B-cell-specific factors. Deletion analysis of the cyclin D2 promoter revealed that removal of sequences containing E-box c-myc consensus DNA binding sequences did not reduce EBNA-2-mediated activation of the cyclin D2 promoter in the transient-transfection assay. The results indicate that cytokines are an early target of EBNA-2 and that EBNA-2 can regulate cyclin D2 transcription in EBV-infected cells by mechanisms additional to the c-myc pathway. PMID- 11264344 TI - Lentivirus vector-mediated hematopoietic stem cell gene transfer of common gamma chain cytokine receptor in rhesus macaques. AB - Nonhuman primate model systems of autologous CD34+ cell transplant are the most effective means to assess the safety and capabilities of lentivirus vectors. Toward this end, we tested the efficiency of marking, gene expression, and transplant of bone marrow and peripheral blood CD34+ cells using a self inactivating lentivirus vector (CS-Rh-MLV-E) bearing an internal murine leukemia virus long terminal repeat derived from a murine retrovirus adapted to replicate in rhesus macaques. In vitro cytokine stimulation was not required to achieve efficient transduction of CD34+ cells resulting in marking and gene expression of the reporter gene encoding enhanced green fluorescent protein (EGFP) following transplant of the CD34+ cells. Monkeys transplanted with mobilized peripheral blood CD34+ cells resulted in EGFP expression in 1 to 10% of multilineage peripheral blood cells, including red blood cells and platelets, stable for 15 months to date. The relative level of gene expression utilizing this vector is 2- to 10-fold greater than that utilizing a non-self-inactivating lentivirus vector bearing the cytomegalovirus immediate-early promoter. In contrast, in animals transplanted with autologous bone marrow CD34+ cells, multilineage EGFP expression was evident initially but diminished over time. We further tested our lentivirus vector system by demonstrating gene transfer of the human common gamma chain cytokine receptor gene (gamma(c)), deficient in X-linked SCID patients and recently successfully used to treat disease. Marking was 0.42 and.001 HIV-1 vector DNA copy per 100 cells in two animals. To date, all EGFP- and gamma(c) transplanted animals are healthy. This system may prove useful for expression of therapeutic genes in human hematopoietic cells. PMID- 11264345 TI - DNase protection analysis of retrovirus integrase at the viral DNA ends for full site integration in vitro. AB - Retrovirus intasomes purified from virus-infected cells contain the linear viral DNA genome and integrase (IN). Intasomes are capable of integrating the DNA termini in a concerted fashion into exogenous target DNA (full site), mimicking integration in vivo. Molecular insights into the organization of avian myeloblastosis virus IN at the viral DNA ends were gained by reconstituting nucleoprotein complexes possessing intasome characteristics. Assembly of IN-4.5 kbp donor complexes capable of efficient full-site integration appears cooperative and is dependent on time, temperature, and protein concentration. DNase I footprint analysis of assembled IN-donor complexes capable of full-site integration shows that wild-type U3 and other donors containing gain-of-function attachment site sequences are specifically protected by IN at low concentrations (<20 nM) with a defined outer boundary mapping ~20 nucleotides from the ends. A donor containing mutations in the attachment site simultaneously eliminated full site integration and DNase I protection by IN. Coupling of wild-type U5 ends with wild-type U3 ends for full-site integration shows binding by IN at low concentrations probably occurs only at the very terminal nucleotides (<10 bp) on U5. The results suggest that assembly requires a defined number of avian IN subunits at each viral DNA end. Among several possibilities, IN may bind asymmetrically to the U3 and U5 ends for full-site integration in vitro. PMID- 11264347 TI - Characterization of the CD154-positive and CD40-positive cellular subsets required for pathogenesis in retrovirus-induced murine immunodeficiency. AB - Genetically susceptible C57BL/6 (B6) mice that are infected with the LP-BM5 isolate of murine retroviruses develop profound splenomegaly, lymphadenopathy, hypergammaglobulinemia, terminal B-cell lymphomas, and an immunodeficiency state bearing many similarities to the pathologies seen in AIDS. Because of these similarities, this syndrome has been called murine AIDS (MAIDS). We have previously shown that CD154 (CD40 ligand)-CD40 molecular interactions are required both for the initiation and progression of MAIDS. Thus, in vivo anti CD154 monoclonal antibody (MAb) treatment inhibited MAIDS symptoms in LP-BM5 infected wild-type mice when either a short course of anti-CD154 MAb treatment was started on the day of infection or a course was initiated 3 to 4 weeks after LP-BM5 administration, after disease was established. Here, we further characterize this required CD154-CD40 interaction by a series of adoptive transfer experiments designed to elucidate which cellular subsets must express CD154 or CD40 for LP-BM5 to induce MAIDS. Specifically with regard to CD154 expression, MAIDS-insusceptible B6 nude mice reconstituted with highly purified CD4+ T cells from wild-type, but not from CD154 knockout, B6 donors displayed clear MAIDS after LP-BM5 infection. In contrast, nude B6 recipients that received CD8+ T cells from wild-type B6 donors did not develop MAIDS after LP-BM5 infection. B6 CD40 knockout mice, which are also relatively resistant to LP-BM5 induced MAIDS, became susceptible to LP-BM5-induced disease after reconstitution with highly purified wild-type B cells but not after receiving purified wild-type dendritic cells (DC) or a combined CD40+ population composed of DC and macrophages obtained from B6 SCID mouse donors. Based on these and other experiments, we thus conclude that the cellular basis for the requirement for CD154-CD40 interactions for MAIDS induction and progression can be accounted for by CD154 expression on CD4+ T cells and CD40 expression on B cells. PMID- 11264348 TI - Inhibition of human immunodeficiency virus type 1 (HIV-1) replication by HIV-1 based lentivirus vectors expressing transdominant Rev. AB - Retrovirus vectors expressing transdominant-negative mutants of Rev (TdRev) inhibit human immunodeficiency virus type 1 (HIV-1) replication by preventing the nuclear export of unspliced viral transcripts, thus inhibiting the synthesis of Gag-Pol, Env, and genomic RNA. The use of HIV-1-based vectors to express TdRev would have the advantage of allowing access to nondividing hematopoietic cells. It would also provide additional levels of protection by sequestering the viral regulatory proteins Tat and Rev, competing for encapsidation into wild-type virions, and inhibiting reverse transcription. Here we describe HIV-1-based vectors that express TdRev. These vectors contain mutations in the splicing signals or replacement of the Rev-responsive element by the simian retrovirus type 1 constitutive transport element, making them less sensitive to the inhibitory effects of TdRev. In addition, overexpression of Rev and the use of an HIV-1 helper plasmid that drives high levels of Gag-Pol synthesis were used to transiently overcome the inhibition by TdRev of the synthesis of Gag-Pol during vector production. SupT1 cells transduced with these vectors were more resistant to HIV-1 replication than cells transduced with Moloney murine leukemia virus based vectors expressing TdRev. Furthermore, we show that these vectors can be mobilized by the wild-type virus, reducing the infectivity of virions escaping inhibition and conferring protection against HIV-1 replication to previously untransduced cells. PMID- 11264346 TI - Differential CD4/CCR5 utilization, gp120 conformation, and neutralization sensitivity between envelopes from a microglia-adapted human immunodeficiency virus type 1 and its parental isolate. AB - Human immunodeficiency virus type 1 (HIV-1) infects and induces syncytium formation in microglial cells from the central nervous system (CNS). A primary isolate (HIV-1(BORI)) was sequentially passaged in cultured microglia, and the isolate recovered (HIV-1(BORI-15)) showed high levels of fusion and replicated more efficiently in microglia (J. M. Strizki, A. V. Albright, H. Sheng, M. O'Connor, L. Perrin, and F. Gonzalez-Scarano, J. Virol. 70:7654-7662, 1996). The parent and adapted viruses used CCR5 as coreceptor. Recombinant viruses demonstrated that the syncytium-inducing phenotype was associated with four amino acid differences in the V1/V2 region of the viral gp120 (J. T. C. Shieh, J. Martin, G. Baltuch, M. H. Malim, and F. Gonzalez-Scarano, J. Virol. 74:693-701, 2000). We produced luciferase-reporter, env-pseudotyped viruses using plasmids containing env sequences from HIV-1(BORI), HIV-1(BORI-15), and the V1/V2 region of HIV-1(BORI-15) in the context of HIV-1(BORI) env (named rBORI, rB15, and rV1V2, respectively). The pseudotypes were used to infect cells expressing various amounts of CD4 and CCR5 on the surface. In contrast to the parent recombinant, the rB15 and rV1V2 pseudotypes retained their infectability in cells expressing low levels of CD4 independent of the levels of CCR5, and they infected cells expressing CD4 with a chimeric coreceptor containing the third extracellular loop of CCR2b in the context of CCR5 or a CCR5 Delta4 amino terminal deletion mutant. The VH-rB15 and VH-rV1V2 recombinant viruses were more sensitive to neutralization by a panel of HIV-positive sera than was VH-rBORI. Interestingly, the CD4-induced 17b epitope on gp120 was more accessible in the rB15 and rV1V2 pseudotypes than in rBORI, even before CD4 binding, and concomitantly, the rB15 and rV1V2 pseudotypes were more sensitive to neutralization with the human 17b monoclonal antibody. Adaptation to growth in microglia--cells that have reduced expression of CD4 in comparison with other cell types--appears to be associated with changes in gp120 that modify its ability to utilize CD4 and CCR5. Changes in the availability of the 17b epitope indicate that these affect conformation. These results imply that the process of adaptation to certain tissue types such as the CNS directly affects the interaction of HIV-1 envelope glycoproteins with cell surface components and with humoral immune responses. PMID- 11264349 TI - Deleterious effects of hepatitis delta virus replication on host cell proliferation. AB - Hepatitis delta virus (HDV) infection and spread in vivo are dependent upon coinfection by hepatitis B virus (HBV), and dual HDV/HBV infection is frequently more severe than HBV infection alone, raising the possibility that HDV infection may be deleterious to cells. Here we have examined the effects of HDV replication on the long-term growth of cultured cells. Our results show that most cells transfected with HDV cDNA do not give rise to stable cell lines expressing viral antigens or replicative intermediates; in addition, cotransfection of HDV replicons with a plasmid vector expressing a hygromycin resistance marker results in a dose-dependent impairment of hygromycin-resistant colony formation. When cells transfected with replication-competent HDV cDNA are followed prospectively, a progressive decline in viral RNA replication and a steady decrease in the proportion of cells expressing delta antigen are observed. However, in transient transfection assays, no evidence was found to link HDV replication to apoptosis or to cell cycle arrest, nor did HDV replication confer on host cells enhanced sensitivity to inducers of apoptosis. Thus, HDV replication does not appear to be acutely cytotoxic. However, in dividing cells HDV replication is associated with a subtler growth disadvantage, leading to selection in culture for cells displaying diminished HDV expression. This effect would not be expected to cause hepatitis in vivo but might contribute to impaired liver regeneration in the setting of ongoing hepatocellular injury. PMID- 11264350 TI - Characterization of endogenous avian leukosis viruses in chicken embryonic fibroblast substrates used in production of measles and mumps vaccines. AB - Previous findings of low levels of reverse transcriptase (RT) activity in chick cell-derived measles and mumps vaccines showed this activity to be associated with virus particles containing RNA of both subgroup E endogenous avian leukosis viruses (ALV-E) and endogenous avian viruses (EAV). These particles originate from chicken embryonic fibroblast (CEF) substrates used for propagating vaccine strains. To better characterize vaccine-associated ALV-E, we examined the endogenous ALV proviruses (ev loci) present in a White Leghorn CEF substrate pool by restriction fragment length polymorphism. Five ev loci were detected, ev-1, ev 3, ev-6, ev-18, andev-19. Both ev-18 and ev-19 can express infectious ALV-E, while ev-1, ev-3, and ev-6 are defective. We analyzed the full-length sequence of ev-1 and identified an adenosine insertion within the pol RT-beta region at position 5026, which results in a truncated RT-beta and integrase. We defined the 1,692-bp deletion in the gag-pol region of ev-3, and we found that in ev-6, sequences from the 5' long terminal repeat to the 5' pol region were absent. Based on the sequences of the ev loci, RT-PCR assays were developed to examine expression of ALV-E particles (EV) in CEF supernatants. Both ev-1- and ev-3-like RNA sequences were identified, as well as two other RNA sequences with intact pol regions, presumably of ev-18 and ev-19 origin. Inoculation of susceptible quail fibroblasts with CEF culture supernatants from both 5-azacytidine-induced and noninduced CEF led to ALV infection, confirming the presence of infectious ALV-E. Our data demonstrate that both defective and nondefective ev loci can be present in CEF vaccine substrates and suggest that both ev classes may contribute to the ALV present in vaccines. PMID- 11264352 TI - Characterization of intracellular reverse transcription complexes of human immunodeficiency virus type 1. AB - To examine the early events of the life cycle of human immunodeficiency virus type 1 (HIV-1), we analyzed the intracellular complexes mediating reverse transcription isolated from acutely infected cells. Partial purification of the reverse transcription complexes (RTCs) by equilibrium density fractionation and velocity sedimentation indicated that two species of RTCs are formed but only one species is able to synthesize DNA. Most of the capsid, matrix, and reverse transcriptase (RT) proteins dissociate from the complex soon after cell infection, but Vpr remains associated with the RTC. The RTCs isolated 1, 4, and 7 h after infection are competent for reverse transcription in vitro, indicating that a small proportion of RT remains associated with them. HIV RTCs isolated early after infection have a sedimentation velocity of approximately 560S. Later, different species with a sedimentation velocity ranging from 350S to 100S appear. Nuclear-associated RTCs have a sedimentation velocity of 80S. Shortly after initiation of reverse transcription, the viral strong-stop DNA within the RTC is sensitive to nuclease digestion and becomes protected when reverse transcription is almost completed. PMID- 11264351 TI - Degradation of p21cip1 in cells productively infected with human cytomegalovirus. AB - Human cytomegalovirus (HCMV) stimulates arrested cells to enter the cell cycle by activating cyclin-dependent kinases (Cdks), notably Cdk2. Several mechanisms are involved in the activation of Cdk2. HCMV causes a substantial increase in the abundance of cyclin E and stimulates translocation of Cdk2 from the cytoplasm to the nucleus. Further, the abundance of the Cdk inhibitors (CKIs) p21cip1/waf1 (p21cip1) and p27kip1 is substantially reduced. The activity of cyclin E/Cdk2 increases as levels of CKIs, particularly p21cip1, fall. We have previously shown that these phenomena contribute to priming the cell for efficient replication of HCMV. In this study, the mechanisms responsible for the decrease in p21cip1 levels after HCMV infection were investigated by measuring p21cip1 RNA and protein levels in permissive human lung (LU) fibroblasts after HCMV infection. Northern blot analysis revealed that p21cip1 RNA levels increased briefly at 3 h after HCMV infection and then decreased to their nadir at 24 h; thereafter, RNA levels increased to about 60% of the preinfection level. Western blot analysis demonstrated that the relative abundance of p21cip1 protein roughly paralleled the observed changes in initial RNA levels; however, the final levels of protein were much lower than preinfection levels. After a transient increase at 3 h postinfection, p21cip1 abundance declined sharply over the next 24 h and remained at a very low level through 96 h postinfection. The disparity between p21cip1 RNA and protein levels suggested that the degradation of p21cip1 might be affected in HCMV-infected cells. Treatment of HCMV-infected cells with MG132, an inhibitor of proteasome-mediated proteolysis, provided substantial protection of p21cip1 in mock-infected cells, but MG132 was much less effective in protecting p21cip1 in HCMV-infected cells. The addition of E64d or Z-Leu-Leu-H, each an inhibitor of calpain activity, to HCMV-infected cells substantially increased the abundance of p21cip1 in a concentration-dependent manner. To verify that p21cip1 was a substrate for calpain, purified recombinant p21cip1 was incubated with either m calpain or mu-calpain, which resulted in rapid proteolysis of p21cip1. E64d inhibited the proteolysis of p21cip1 catalyzed by either m-calpain or mu-calpain. Direct measurement of calpain activity in HCMV-infected LU cells indicated that HCMV infection induced a substantial and sustained increase in calpain activity, although there was no change in the abundance of either m- or mu-calpain or the endogenous calpain inhibitor calpastatin. The observed increase of calpain activity was consistent with the increases in intracellular free Ca2+ and phospholipid degradation in HCMV-infected LU cells reported previously from our laboratory. Considered together, these results suggest that the increase in calpain activity observed following HCMV infection contributes significantly to the reduction of p21cip1 levels and the resultant cell cycle progression. PMID- 11264353 TI - Region of herpes simplex virus type 1 latency-associated transcript sufficient for wild-type spontaneous reactivation promotes cell survival in tissue culture. AB - The latency-associated transcript (LAT) is the only abundant herpes simplex virus type 1 (HSV-1) transcript expressed during latency. In the rabbit eye model, LAT null mutants do not reactivate efficiently from latency. We recently demonstrated that the LAT null mutant dLAT2903 induces increased levels of apoptosis in trigeminal ganglia of infected rabbits compared to LAT+ strains (G.-C. Perng, C. Jones, J. Ciacci-Zarella, M. Stone, G. Henderson, A. Yokht, S. M. Slanina, F. M. Hoffman, H. Ghiasi, A. B. Nesburn, and C. S. Wechsler, Science 287:1500-1503, 2000). The same study also demonstrated that a plasmid expressing LAT nucleotides 301 to 2659 enhanced cell survival of transfected cells after induction of apoptosis. Consequently, we hypothesized that LAT enhances spontaneous reactivation in part, because it promotes survival of infected neurons. Here we report on the ability of plasmids expressing different portions of the 5' end of LAT to promote cell survival after induction of apoptosis. A plasmid expressing the first 1.5 kb of LAT (LAT nucleotides 1 to 1499) promoted cell survival in neuro-2A (mouse neuronal) and CV-1 (monkey fibroblast) cells. A plasmid expressing just the first 811 nucleotides of LAT promoted cell survival less efficiently. Plasmids expressing the first 661 nucleotides or less of LAT did not promote cell survival. We previously showed that a mutant expressing just the first 1.5 kb of LAT has wild-type spontaneous reactivation in rabbits, and a mutant expressing just the first 811 nucleotides of LAT has a reactivation frequency higher than that of dLAT2903 but lower than that of wild-type virus. In addition, mutants reported here for the first time, expressing just the first 661 or 76 nucleotides of LAT, had spontaneous reactivation indistinguishable from that of the LAT null mutant dLAT2903. In summary, these studies provide evidence that there is a functional relationship between the ability of LAT to promote cell survival and its ability to enhance spontaneous reactivation. PMID- 11264354 TI - Definitive assignment of proton selectivity and attoampere unitary current to the M2 ion channel protein of influenza A virus. AB - The viral ion channel protein M2 supports the transit of influenza virus and its glycoproteins through acidic compartments of the cell. M2 conducts endosomal protons into the virion to initiate uncoating and, by equilibrating the pH at trans-Golgi membranes, preserves the native conformation of acid-sensitive viral hemagglutinin. The exceptionally low conductance of the M2 channel thwarted resolution of single channels by electrophysiological techniques. Assays of liposome-reconstituted M2 yielded the average unitary channel current of the M2 tetramer--1.2 aA (1.2 x 10(-18) A) at neutral pH and 2.7 to 4.1 aA at pH 5.7- which activates the channel. Extrapolation to physiological temperature predicts 4.8 and 40 aA, respectively, and a unitary conductance of 0.03 versus 0.4 fS. This minute activity, below previous estimates, appears sufficient for virus reproduction, but low enough to avert abortive cytotoxicity. The unitary permeability of M2 was within the range reported for other proton channels. To address the ion selectivity of M2, we exploited the coupling of ionic influx and efflux in sealed liposomes. Metal ion fluxes were monitored by proton counterflow, employing a pH probe 1,000 times more sensitive than available Na+ or K+ probes. Even low-pH-activated M2 did not conduct Na+ and K+. The proton selectivity of M2 was estimated to be at least 3 x 10(6) (over sodium or potassium ions), in agreement with electrophysiological studies. The stringent proton selectivity of M2 suggests that the cytopathology of influenza virus does not involve direct perturbation of cellular sodium or potassium gradients. PMID- 11264355 TI - Modulation of different human immunodeficiency virus type 1 Nef functions during progression to AIDS. AB - The human immunodeficiency virus type 1 (HIV-1) Nef protein has several independent functions that might contribute to efficient viral replication in vivo. Since HIV-1 adapts rapidly to its host environment, we investigated if different Nef properties are associated with disease progression. Functional analysis revealed that nef alleles obtained during late stages of infection did not efficiently downmodulate class I major histocompatibility complex but were highly active in the stimulation of viral replication. In comparison, functional activity in downregulation of CD4 and enhancement of HIV-1 infectivity were maintained or enhanced after AIDS progression. Our results demonstrate that various Nef activities are modulated during the course of HIV-1 infection to maintain high viral loads at different stages of disease progression. These findings suggest that all in vitro Nef functions investigated contribute to AIDS pathogenesis and indicate that nef variants with increased pathogenicity emerge in a significant number of HIV-1-infected individuals. PMID- 11264356 TI - AlaArg motif in the carboxyl terminus of the gamma(1)34.5 protein of herpes simplex virus type 1 is required for the formation of a high-molecular-weight complex that dephosphorylates eIF-2alpha. AB - The gamma(1)34.5 protein of herpes simplex virus (HSV) type 1 functions to prevent the shutoff of protein synthesis mediated by the double-stranded-RNA dependent protein kinase PKR. This is because gamma(1)34.5 associates with protein phosphatase 1 (PP1) through its carboxyl terminus, forming a high molecular-weight complex that dephosphorylates the alpha subunit of translation initiation factor eIF-2 (eIF-2alpha). Here we show that Val193Glu and Phe195Leu substitutions in the PP1 signature motif of the gamma(1)34.5 protein abolished its ability to redirect PP1 to dephosphorylate eIF-2alpha and replication of mutant viruses was severely impaired. The gamma(1)34.5 protein, when expressed in Sf9 cells using a recombinant baculovirus, was capable of directing specific eIF 2alpha dephosphorylation. Deletions of amino acids 258 to 263 had no effect on activity of gamma(1)34.5. However, deletions of amino acids 238 to 258 abolished eIF-2alpha phosphatase activity but not PP1 binding activity. Interestingly, deletions in the AlaArg motif of the carboxyl terminus disrupted the high molecular-weight complex that is required for dephosphorylation of eIF-2alpha. These results demonstrate that gamma(1)34.5 is functionally active in the absence of any other HSV proteins. In addition to a PP1 binding domain, the carboxyl terminus of gamma(1)34.5 contains an effector domain that is required to form a functional complex. PMID- 11264357 TI - Egress of alphaherpesviruses: comparative ultrastructural study. AB - Egress of four important alphaherpesviruses, equine herpesvirus 1 (EHV-1), herpes simplex virus type 1 (HSV-1), infectious laryngotracheitis virus (ILTV), and pseudorabies virus (PrV), was investigated by electron microscopy of infected cell lines of different origins. In all virus-cell systems analyzed, similar observations were made concerning the different stages of virion morphogenesis. After intranuclear assembly, nucleocapsids bud at the inner leaflet of the nuclear membrane, resulting in enveloped particles in the perinuclear space that contain a sharply bordered rim of tegument and a smooth envelope surface. Egress from the perinuclear cisterna primarily occurs by fusion of the primary envelope with the outer leaflet of the nuclear membrane, which has been visualized for HSV 1 and EHV-1 for the first time. The resulting intracytoplasmic naked nucleocapsids are enveloped at membranes of the trans-Golgi network (TGN), as shown by immunogold labeling with a TGN-specific antiserum. Virions containing their final envelope differ in morphology from particles within the perinuclear cisterna by visible surface projections and a diffuse tegument. Particularly striking was the addition of a large amount of tegument material to ILTV capsids in the cytoplasm. Extracellular virions were morphologically identical to virions within Golgi-derived vesicles, but distinct from virions in the perinuclear space. Studies with gB- and gH-deleted PrV mutants indicated that these two glycoproteins, which are essential for virus entry and direct cell-to-cell spread, are dispensable for egress. Taken together, our studies indicate that the deenvelopment-reenvelopment process of herpesvirus maturation also occurs in EHV 1, HSV-1, and ILTV and that membrane fusion processes occurring during egress are substantially different from those during entry and direct viral cell-to-cell spread. PMID- 11264358 TI - Activation of membrane fusion by murine leukemia viruses is controlled in cis or in trans by interactions between the receptor-binding domain and a conserved disulfide loop of the carboxy terminus of the surface glycoprotein. AB - Cell entry of retroviruses is initiated by the recognition of cellular receptors and the subsequent membrane fusion between viral and cellular membranes. These two steps are mediated by the surface (SU) and transmembrane (TM) subunits of the retroviral envelope glycoprotein (Env), respectively. Determinants regulating membrane fusion have been described throughout SU and TM, but the processes coupling receptor recognition to fusion are still elusive. Here we establish that a critical interaction is formed between the receptor-binding domain (RBD) and the major disulfide loop of the carboxy-terminal domain (C domain) of the murine leukemia virus SU. Receptor binding causes an alteration of this interaction and, in turn, promotes further events of Env fusion activation. We characterize mutations which, by lowering this interaction and reducing the compatibility between the RBD and C domains of Env glycoprotein chimeras, affect both Env fusogenicity and sensitivity to receptor interference. Additionally, we demonstrate that suboptimal interactions in such mutant Env proteins can be compensated in trans by soluble RBDs in a manner that depends on their compatibility with the C domain. Our results therefore indicate that RBD/C domain interactions may occur in cis, via the proper RBD of the viral Env itself, or in trans, via a distinct RBD expressed by virion-free Env glycoproteins expressed endogenously by the infected cells or provided by neighboring Env trimers. PMID- 11264359 TI - Reassortment in vivo: driving force for diversity of human rotavirus strains isolated in the United Kingdom between 1995 and 1999. AB - The G and P genotypes of 3,601 rotavirus strains collected in the United Kingdom between 1995 and 1999 were determined (M. Iturriza-Gomara et al., J. Clin. Microbiol. 38:4394-4401, 2000). In 95.4% of the strains the most common G and P combinations, G1P[8], G2P[4], G3P[8], and G4P[8], were found. A small but significant number (2%) of isolates from the remaining strains were reassortants of the most common cocirculating strains, e.g., G1P[4] and G2P[8]. Rotavirus G9P[6] and G9P[8] strains, which constituted 2.7% of all viruses, were genetically closely related in their G components, but the P components of the G9P[8] strains were very closely related to those of cocirculating strains of the more common G types (G1, G3, and G4). In conclusion, genetic interaction by reassortment among cocirculating rotaviruses is not a rare event and contributes significantly to their overall diversity. PMID- 11264360 TI - Role of alpha/beta interferon in Venezuelan equine encephalitis virus pathogenesis: effect of an attenuating mutation in the 5' untranslated region. AB - Venezuelan equine encephalitis virus (VEE) is an important equine and human pathogen of the Americas. In the adult mouse model, cDNA-derived, virulent V3000 inoculated subcutaneously (s.c.) causes high-titer peripheral replication followed by neuroinvasion and lethal encephalitis. A single change (G to A) at nucleotide 3 (nt 3) of the 5' untranslated region (UTR) of the V3000 genome resulted in a virus (V3043) that was avirulent in mice. The mechanism of attenuation by the V3043 mutation was studied in vivo and in vitro. Kinetic studies of virus spread in adult mice following s.c. inoculation showed that V3043 replication was reduced in peripheral organs compared to that of V3000, titers in serum also were lower, and V3043 was cleared more rapidly from the periphery than V3000. Because clearance of V3043 from serum began 1 to 2 days prior to clearance of V3000, we examined the involvement of alpha/beta interferon (IFN-alpha/beta) activity in VEE pathogenesis. In IFN-alpha/betaR(-/-) mice, the course of the wild-type disease was extremely rapid, with all animals dying within 48 h (average survival time of 30 h compared to 7.7 days in the wild-type mice). The mutant V3043 was as virulent as the wild type (100% mortality, average survival time of 30 h). Virus titers in serum, peripheral organs, and the brain were similar in V3000- and V3043-infected IFN-alpha/betaR(-/-) mice at all time points up until the death of the animals. Consistent with the in vivo data, the mutant virus exhibited reduced growth in vitro in several cell types except in cells that lacked a functional IFN-alpha/beta pathway. In cells derived from IFN alpha/betaR(-/-) mice, the mutant virus showed no growth disadvantage compared to the wild-type virus, suggesting that IFN-alpha/beta plays a major role in the attenuation of V3043 compared to V3000. There were no differences in the induction of IFN-alpha/beta between V3000 and V3043, but the mutant virus was more sensitive than V3000 to the antiviral actions of IFN-alpha/beta in two separate in vitro assays, suggesting that the increased sensitivity to IFN alpha/beta plays a major role in the in vivo attenuation of V3043. PMID- 11264361 TI - Genetic analysis of a poliovirus/hepatitis C virus chimera: new structure for domain II of the internal ribosomal entry site of hepatitis C virus. AB - Internal ribosomal entry sites (IRESs) of certain plus-strand RNA viruses direct cap-independent initiation of protein synthesis both in vitro and in vivo, as can be shown with artificial dicistronic mRNAs or with chimeric viral genomes in which IRES elements were exchanged from one virus to another. Whereas IRESs of picornaviruses can be readily analyzed in the context of their cognate genome by genetics, the IRES of hepatitis C virus (HCV), a Hepacivirus belonging to Flaviviridae, cannot as yet be subjected to such analyses because of difficulties in propagating HCV in tissue culture or in experimental animals. This enigma has been overcome by constructing a poliovirus (PV) whose translation is controled by the HCV IRES. Within the PV/HCV chimera, the HCV IRES has been subjected to systematic 5' deletion analyses to yield a virus (P/H710-d40) whose replication kinetics match that of the parental poliovirus type 1 (Mahoney). Genetic analyses of the HCV IRES in P/H710-d40 have confirmed that the 5' border maps to domain II, thereby supporting the validity of the experimental approach applied here. Additional genetic experiments have provided evidence for a novel structural region within domain II. Arguments that the phenotypes observed with the mutant chimera relate solely to impaired genome replication rather than deficiencies in translation have been dispelled by constructing novel dicistronic poliovirus replicons with the gene order [PV]cloverleaf-[HCV]IRES-Deltacore-R-Luc-[PV]IRES-F Luc-P2,3-3'NTR, which have allowed the measurement of HCV IRES-dependent translation independently from the replication of the replicon RNA. PMID- 11264362 TI - Palindromic sequence plays a critical role in human foamy virus dimerization. AB - The retroviral RNA genome is dimeric, consisting of two identical strands of RNA linked near their 5' ends by a dimer linkage structure. Previously it was shown that human foamy virus (HFV) RNA transcribed in vitro contained three sites, designated SI, SII, and SIII, which contributed to the dimerization process (O. Erlwein, D. Cain, N. Fischer, A. Rethwilm, and M. O. McClure, Virology 229:251 258, 1997). To characterize these sites further, a series of mutants were designed and tested for their ability to dimerize in vitro. The primer binding site and a G tetrad in SI were dispensable for dimerization. However, a mutant that changed the 3' end of SI migrated slower on nondenaturing gels than wild type RNA dimers. The sequence composition of the SII palindrome, consisting of 10 nucleotides, proved to be critical for in vitro dimerization, since mutations within this sequence or replacement of the sequence with a different palindrome of equal length impaired in vitro dimerization. The length of the palindrome also seems to play an important role. A moderate extension to 12 nucleotides was tolerated, whereas an extension to 16 nucleotides or more impaired dimerization. When nucleotides flanking the palindrome were mutated in a random fashion, dimerization was unaffected. Changing the SIII sequence also led to decreased dimer formation, confirming its contribution to the dimerization process. Interesting mutants were cloned into the infectious molecular clone of HFV, HSRV 2, and were transfected into BHK-21 cells. Mutations in SII that reduced dimerization in vitro also abolished virus replication. In contrast, constructs containing mutations in SI and SIII replicated to some extent in cell culture after an initial drop in viral replication. Analysis of the SIM1 mutant revealed reversion to the wild type but with the insertion of an additional two nucleotides. Analysis of cell-free virions demonstrated that both replication competent and replication-defective mutants packaged nucleic acid. Thus, efficient dimerization is a critical step for HFV to generate infectious virus, but HFV RNA dimerization is not a prerequisite for packaging. PMID- 11264363 TI - CD4+ T-cell effectors inhibit Epstein-Barr virus-induced B-cell proliferation. AB - In immunodeficient hosts, Epstein-Barr virus (EBV) often induces extensive B-cell lymphoproliferative disease and lymphoma. Without effective in vitro immune surveillance, B cells infected by the virus readily form immortalized cell lines. In the regression assay, memory T cells inhibit the formation of foci of EBV transformed B cells that follows recent in vitro infection by EBV. No one has yet addressed which T cell regulates the early proliferative phase of B cells newly infected by EBV. Using new quantitative methods, we analyzed T-cell surveillance of EBV-mediated B-cell proliferation. We found that CD4+ T cells play a significant role in limiting proliferation of newly infected, activated CD23+ B cells. In the absence of T cells, EBV-infected CD23+ B cells divided rapidly during the first 3 weeks after infection. Removal of CD4+ but not CD8+ T cells also abrogated immune control. Purified CD4+ T cells eliminated outgrowth when added to EBV-infected B cells. Thus, unlike the killing of EBV-infected lymphoblastoid cell lines, in which CD8+ cytolytic T cells play an essential role, prevention of early-phase EBV-induced B-cell proliferation requires CD4+ effector T cells. PMID- 11264365 TI - Adaptation of influenza A viruses to cells expressing low levels of sialic acid leads to loss of neuraminidase activity. AB - Influenza A viruses possess two virion surface proteins, hemagglutinin (HA) and neuraminidase (NA). The HA binds to sialyloligosaccharide viral receptors, while the NA removes sialic acids from the host cell and viral sialyloligosaccarides. Alterations of the HA occur during adaptation of influenza viruses to new host species, as in the 1957 and 1968 influenza pandemics. To gain a better understanding of the contributions of the HA and possibly the NA to this process, we generated cell lines expressing reduced levels of the influenza virus receptor determinant, sialic acid, by selecting Madin-Darby canine kidney cells resistant to a lectin specific for sialic acid linked to galactose by alpha(2-3) or alpha(2 6) linkages. One of these cell lines had less than 1/10 as much N acetylneuraminic acid as its parent cell line. When serially passaged in this cell line, human H3N2 viruses lost sialidase activity due to a large internal deletion in the NA gene, without alteration of the HA gene. These findings indicate that NA mutations can contribute to the adaptation of influenza A virus to new host environments and hence may play a role in the transmission of virus across species. PMID- 11264366 TI - Actin rearrangement-inducing factor of baculoviruses is tyrosine phosphorylated and colocalizes to F-actin at the plasma membrane. AB - In previous studies we have identified actin rearrangement-inducing factor 1 as an early gene product of Autographa californica multicapsid nuclear polyhedrosis virus that is involved in the remodeling of the actin cytoskeleton. We have constructed viral recombinants with a mutated Arif-1 open reading frame that confirm the causal link of Arif-1 expression and the actin rearrangement observed as accumulation of F-actin at the plasma membrane at 3 to 7 h postinfection. Infection with Arif mutant viruses leads to the loss of actin accumulation at the plasma membrane in TN-368 cells, although in the course of infection, early actin cables and nuclear F-actin are observed as in wild-type-infected cells. By immunofluorescence studies, we have demonstrated the localization of Arif-1 at the plasma membrane, and confocal imaging reveals the colocalization to F-actin. Accordingly, the approximately 47-kDa Arif-1 protein is observed exclusively in membrane fractions prepared at 4 to 48 h postinfection, with a decrease at 24 h postinfection. Phosphatase treatment suggests that Arif-1 is modified by phosphorylation. Antibodies against phosphotyrosine precipitate Arif-1 from membrane fractions, indicating that Arif-1 becomes tyrosine phosphorylated during the early and late phases of infection. In summary, our results indicate that functional Arif-1 is tyrosine phosphorylated and is located at the plasma membrane as a component of the actin rearrangement-inducing complex. PMID- 11264364 TI - Route of simian immunodeficiency virus inoculation determines the complexity but not the identity of viral variant populations that infect rhesus macaques. AB - A better understanding of the host and viral factors associated with human immunodeficiency virus (HIV) transmission is essential to developing effective strategies to curb the global HIV epidemic. Here we used the rhesus macaque simian immunodeficiency virus (SIV) animal model of HIV infection to study the range of viral genotypes that are transmitted by different routes of inoculation and by different types of viral inocula. Analysis of transmitted variants was undertaken in outbred rhesus macaques inoculated intravenously (IV) or intravaginally (IVAG) with a genetically heterogeneous SIVmac251 stock derived from a well-characterized rhesus macaque viral isolate. In addition, we performed serial IV and IVAG passage experiments using plasma from SIV-infected macaques as the inoculum. We analyzed the V1-V2 region of the SIV envelope gene from virion associated RNA in plasma from infected animals by the heteroduplex mobility assay (HMA) and by DNA sequence analysis. We found that a more diverse population of SIV genetic variants was present in the earliest virus-positive plasma samples from all five IV SIVmac251-inoculated monkeys and from two of five IVAG SIVmac251 inoculated monkeys. In contrast, we found a relatively homogeneous population of SIV envelope variants in three of five monkeys inoculated IVAG with SIVmac251 stock and in two monkeys infected after IVAG inoculation with plasma from an SIV infected animal. In some IVAG-inoculated animals, the transmitted SIV variant was the most common variant in the inoculum. However, a specific viral variant in the SIVmac251 stock was not consistently transmitted by IVAG inoculation. Thus, it is likely that host factors or stochastic processes determine the specific viral variants that infect an animal after IVAG SIV exposure. In addition, our results clearly demonstrate that the route of inoculation is associated with the extent and breadth of the genetic complexity of the viral variant population in the earliest stages of systemic infection. PMID- 11264368 TI - Comparison of cell cycle arrest, transactivation, and apoptosis induced by the simian immunodeficiency virus SIVagm and human immunodeficiency virus type 1 vpr genes. AB - All primate lentiviruses known to date contain one or two open reading frames with homology to the human immunodeficiency virus type 1 (HIV-1) vpr gene. HIV-1 vpr encodes a 96-amino-acid protein with multiple functions in the viral life cycle. These functions include modulation of the viral replication kinetics, transactivation of the long terminal repeat, participation in the nuclear import of preintegration complexes, induction of G2 arrest, and induction of apoptosis. The simian immunodeficiency virus (SIV) that infects African green monkeys (SIVagm) contains a vpr homologue, which encodes a 118-amino-acid protein. SIVagm vpr is structurally and functionally related to HIV-1 vpr. The present study focuses on how three specific functions (transactivation, induction of G2 arrest, and induction of apoptosis) are related to one another at a functional level, for HIV-1 and SIVagm vpr. While our study supports previous reports demonstrating a causal relationship between induction of G2 arrest and transactivation for HIV-1 vpr, we demonstrate that the same is not true for SIVagm vpr. Transactivation by SIVagm vpr is independent of cell cycle perturbation. In addition, we show that induction of G2 arrest is necessary for the induction of apoptosis by HIV-1 vpr but that the induction of apoptosis by SIVagm vpr is cell cycle independent. Finally, while SIVagm vpr retains its transactivation function in human cells, it is unable to induce G2 arrest or apoptosis in such cells, suggesting that the cytopathic effects of SIVagm vpr are species specific. Taken together, our results suggest that while the multiple functions of vpr are conserved between HIV-1 and SIVagm, the mechanisms leading to the execution of such functions are divergent. PMID- 11264367 TI - CCR5, CXCR4, and CD4 are clustered and closely apposed on microvilli of human macrophages and T cells. AB - The chemokine receptors CCR5 and CXCR4 act synergistically with CD4 in an ordered multistep mechanism to allow the binding and entry of human immunodeficiency virus type 1 (HIV-1). The efficiency of such a coordinated mechanism depends on the spatial distribution of the participating molecules on the cell surface. Immunoelectron microscopy was performed to address the subcellular localization of the chemokine receptors and CD4 at high resolution. Cells were fixed, cryoprocessed, and frozen; 80-nm cryosections were double labeled with combinations of CCR5, CXCR4, and CD4 antibodies and then stained with immunogold. Surprisingly, CCR5, CXCR4, and CD4 were found predominantly on microvilli and appeared to form homogeneous microclusters in all cell types examined, including macrophages and T cells. Further, while mixed microclusters were not observed, homogeneous microclusters of CD4 and the chemokine receptors were frequently separated by distances less than the diameter of an HIV-1 virion. Such distributions are likely to facilitate cooperative interactions with HIV-1 during virus adsorption to and penetration of human leukocytes and have significant implications for development of therapeutically useful inhibitors of the entry process. Although the mechanism underlying clustering is not understood, clusters were observed in small trans-Golgi vesicles, implying that they were organized shortly after synthesis and well before insertion into the cellular membrane. Chemokine receptors normally act as sensors, detecting concentration gradients of their ligands and thus providing directional information for cellular migration during both normal homeostasis and inflammatory responses. Localization of these sensors on the microvilli should enable more precise monitoring of their environment, improving efficiency of the chemotactic process. Moreover, since selectins, some integrins, and actin are also located on or in the microvillus, this organelle has many of the major elements required for chemotaxis. PMID- 11264370 TI - Replication of naturally occurring woodchuck hepatitis virus deletion mutants in primary hepatocyte cultures and after transmission to naive woodchucks. AB - Woodchuck hepatitis virus (WHV) mutants with core internal deletions (CID) occur naturally in chronically WHV-infected woodchucks, as do hepatitis B virus mutants in humans. We studied the replication of WHV deletion mutants in primary woodchuck hepatocyte cultures and in vivo after transmission to naive woodchucks. By screening 14 wild-caught, chronically WHV-infected woodchucks, two woodchucks, WH69 and WH70, were found to harbor WHV CID mutants. Consistent with previous results, WHV CID mutants from both animals had deletions of variable lengths (90 to 135 bp) within the middle of the WHV core gene. In woodchuck WH69, WHV CID mutants represented a predominant fraction of the viral population in sera, normal liver tissues, and to a lesser extent, in liver tumor tissues. In primary hepatocytes of WH69, the replication of wild-type WHV and CID mutants was maintained at least for 7 days. Although WHV CID mutants were predominant in fractions of cellular WHV replicative intermediates, mutant covalently closed circular DNAs (cccDNAs) appeared to be a small part of cccDNA-enriched fractions. Analysis of cccDNA-enriched fractions from liver tissues of other woodchucks confirmed that mutant cccDNA represents only a small fraction of the total cccDNA pool. Four naive woodchucks were inoculated with sera from woodchuck WH69 or WH70 containing WHV CID mutants. All four woodchucks developed viremia after 3 to 4 weeks postinoculation (p.i.). They developed anti-WHV core antigen (WHcAg) antibody, lymphoproliferative response to WHcAg, and anti-WHV surface antigen. Only wild-type WHV, but no CID mutant, was found in sera from these woodchucks. The WHV CID mutant was also not identified in liver tissue from one woodchuck sacrificed in week 7 p.i. Three remaining woodchucks cleared WHV. Thus, the presence of WHV CID mutants in the inocula did not significantly change the course of acute self-limiting WHV infection. Our results indicate that the replication of WHV CID mutants might require some specific selective conditions. Further investigations on WHV CID mutants will allow us to have more insight into hepadnavirus replication. PMID- 11264369 TI - Y2, the smallest of the Sendai virus C proteins, is fully capable of both counteracting the antiviral action of interferons and inhibiting viral RNA synthesis. AB - An open reading frame (ORF) overlapping the amino-terminal portion of the Sendai virus (SeV) P ORF in the +1 frame produces a nested set of carboxy-coterminal proteins, C', C, Y1, and Y2, which are referred to collectively as the C proteins. The C proteins are extremely versatile triple-role players; they counteract the antiviral action of interferons (IFNs), inhibit viral RNA synthesis, and are involved in virus assembly. In this study, we established HeLa cell lines stably expressing the C, Y1, and Y2 proteins individually and examined the capacities of these cells to circumvent the antiviral action of alpha/beta IFN (IFN-alpha/beta) and IFN-gamma and to inhibit viral transcription. The assay protocols included monitoring of IFN-alpha/beta-mediated signaling by interferon stimulated response element-driven reporter gene expression and of the antiviral state induced by IFN-alpha/beta and IFN-gamma and measurement of reporter gene expression from an SeV minigenome, as well as quantification of SeV primary transcripts. When necessary, the activities measured were carefully normalized to the expression levels of the respective C proteins in cells. The data obtained clearly indicate that the smallest protein, Y2, was as active as the C and Y1 proteins in both counteracting the antiviral action of IFNs and inhibiting viral transcription. The data further show that intracellular transexpression of either C, Y1, or Y2 rendered HeLa cells moderately or only poorly permissive for not only wild-type SeV but also 4C(-) SeV, which expressed none of the four C proteins. On the basis of these findings, the roles of SeV C proteins in the natural life cycle are discussed. PMID- 11264371 TI - Human neuron-committed teratocarcinoma NT2 cell line has abnormal ND10 structures and is poorly infected by herpes simplex virus type 1. AB - Herpes simplex virus type 1 (HSV-1) immediate-early regulatory protein ICP0 stimulates the initiation of lytic infection and reactivation from quiescence in human fibroblast cells. These functions correlate with its ability to localize to and disrupt centromeres and specific subnuclear structures known as ND10, PML nuclear bodies, or promyelocytic oncogenic domains. Since the natural site of herpesvirus latency is in neurons, we investigated the status of ND10 and centromeres in uninfected and infected human cells with neuronal characteristics. We found that NT2 cells, a neuronally committed human teratocarcinoma cell line, have abnormal ND10 characterized by low expression of the major ND10 component PML and no detectable expression of another major ND10 antigen, Sp100. In addition, PML is less extensively modified by the ubiquitin-like protein SUMO-1 in NT2 cells compared to fibroblasts. After treatment with retinoic acid, NT2 cells differentiate into neuron-like hNT cells which express very high levels of both PML and Sp100. Infection of both NT2 and hNT cells by HSV-1 was poor compared to human fibroblasts, and after low-multiplicity infection yields of virus were reduced by 2 to 3 orders of magnitude. ICP0-deficient mutants were also disabled in the neuron-related cell lines, and cells quiescently infected with an ICP0-null virus could be established. These results correlated with less efficient disruption of ND10 and centromeres induced by ICP0 in NT2 and hNT cells. Furthermore, the ability of ICP0 to activate gene expression in transfection assays in NT2 cells was poor compared to Vero cells. These results suggest that a contributory factor in the reduced HSV-1 replication in the neuron related cells is inefficient ICP0 function; it is possible that this is pertinent to the establishment of latent infection in neurons in vivo. PMID- 11264372 TI - Requirements for the nuclear-cytoplasmic translocation of infected-cell protein 0 of herpes simplex virus 1. AB - Earlier studies have shown that wild-type infected-cell protein 0 (ICP0), a key herpes simplex virus regulatory protein, translocates from the nucleus to the cytoplasm of human embryonic lung (HEL) fibroblasts within several hours after infection (Y. Kawaguchi, R. Bruni, and B. Roizman, J. Virol. 71:1019-1024, 1997). Translocation of ICP0 was also observed in cells infected with the d120 mutant, in which both copies of the gene encoding ICP4, the major regulatory protein, had been deleted (V. Galvan, R. Brandimarti, J. Munger, and B. Roizman, J. Virol. 74:1931-1938, 2000). Furthermore, a mutant (R7914) carrying the D199A substitution in ICP0 does not bind or stabilize cyclin D3 and is retained in the nucleus (C. Van Sant, P. Lopez, S. J. Advani, and B. Roizman, J. Virol. 75:1888 1898, 2001). Studies designed to elucidate the requirements for the translocation of ICP0 between cellular compartments revealed the following. (i) Translocation of ICP0 to the cytoplasm in productive infection maps to the D199 amino acid, inasmuch as wild-type ICP0 delivered in trans to cells infected with an ICP0 null mutant was translocated to the cytoplasm whereas the D199A-substituted mutant ICP0 was not. (ii) Translocation of wild-type ICP0 requires a function expressed late in infection, inasmuch as phosphonoacetate blocked the translocation of ICP0 in wild-type virus-infected cells but not in d120 mutant-infected cells. Moreover, whereas in d120 mutant-infected cells ICP0 was translocated rapidly from the cytoplasm to the nucleus at approximately 5 h after infection, the translocation of ICP0 in wild-type virus-infected cells extended from 5 to at least 9 h after infection. (iii) In wild-type virus-infected cells, the MG132 proteasomal inhibitor blocked the translocation of ICP0 to the cytoplasm early in infection, but when added late in infection, it caused ICP0 to be relocated back to the nucleus from the cytoplasm. (iv) MG132 blocked the translocation of ICP0 in d120 mutant-infected cells early in infection but had no effect on the ICP0 aggregated in vesicle-like structures late in infection. However, in d120 mutant infected cells treated with MG132 at late times, proteasomes formed a shell-like structure around the aggregated ICP0. These structures were not seen in wild-type virus or R7914 mutant-infected cells. The results indicate the following. (i) In the absence of beta or gamma protein synthesis, ICP0 dynamically associates with proteasomes and is translocated to the cytoplasm. (ii) In cells productively infected beyond alpha gene expression, ICP0 is retained in the nucleus until after the onset of viral DNA synthesis and the synthesis of gamma2 proteins. (iii) Late in infection, ICP0 is actively sequestered in the cytoplasm by a process mediated by proteasomes, inasmuch as interference with proteasomal function causes rapid relocation of ICP0 to the nucleus. PMID- 11264373 TI - Requirements for assembly of poliovirus replication complexes and negative-strand RNA synthesis. AB - HeLa cells were transfected with several plasmids that encoded all poliovirus (PV) nonstructural proteins. Viral RNAs were transcribed by T7 RNA polymerase expressed from recombinant vaccinia virus. All plasmids produced similar amounts of viral proteins that were processed identically; however, RNAs were designed either to serve as templates for replication or to contain mutations predicted to prevent RNA replication. The mutations included substitution of the entire PV 5' noncoding region (NCR) with the encephalomyocarditis virus (EMCV) internal ribosomal entry site, thereby deleting the 5'-terminal cloverleaf-like structure, or insertion of three nucleotides in the 3Dpol coding sequence. Production of viral proteins was sufficient to induce the characteristic reorganization of intracellular membranes into heterogeneous-sized vesicles, independent of RNA replication. The vesicles were stably associated with viral RNA only when RNA replication could occur. Nonreplicating RNAs localized to distinct, nonoverlapping regions in the cell, excluded from the viral protein-membrane complexes. The absence of accumulation of positive-strand RNA from both mutated RNAs in transfected cells was documented. In addition, no minus-strand RNA was produced from the EMCV chimeric template RNA in vitro. These data show that the 5'-terminal sequences of PV RNA are essential for initiation of minus-strand RNA synthesis at its 3' end. PMID- 11264374 TI - Hepatitis B virus X protein acts as a tumor promoter in development of diethylnitrosamine-induced preneoplastic lesions. AB - Chronic infection with hepatitis B virus (HBV) is one of the major etiological factors in the development of human hepatocellular carcinoma. Transgenic mice that express the HBV X protein (HBx) have previously been shown to be more sensitive to the effects of hepatocarcinogens. Although the mechanism for this cofactor role remains unknown, the ability of HBx to inhibit DNA repair and to influence cell cycle progression suggests two possible pathways. To investigate these possibilities in vivo, we treated double-transgenic mice that both express HBx (ATX mice) and possess a bacteriophage lambda transgene with the hepatocarcinogen diethylnitrosamine (DEN). Histological examination of liver tissue confirmed that DEN-treated ATX mice developed approximately twice as many focal lesions of basophilic hepatocytes as treated wild-type littermates. Treatment of mice with DEN resulted in a six- to eightfold increase in the mutation frequency (MF), as measured by a functional analysis of the lambda transgene. HBx expression was confirmed by immunoprecipitation and Western blotting and was associated with a modest 23% increase in the MF. Importantly, the extent of hepatocellular proliferation in 14-day-old mice, as measured by the detection of proliferating cell nuclear antigen and by the incorporation of 5 bromo-2'-deoxyuridine, was determined to be approximately twofold higher in ATX livers than in wild-type livers. These results are consistent with a model in which HBx expression contributes to the development of DEN-mediated carcinogenesis by promoting the proliferation of altered hepatocytes rather than by directly interfering with the repair of DNA lesions. PMID- 11264375 TI - Evaluation of interactions of human cytomegalovirus immediate-early IE2 regulatory protein with small ubiquitin-like modifiers and their conjugation enzyme Ubc9. AB - The human cytomegalovirus (HCMV) major immediate-early protein IE2 is a nuclear phosphoprotein that is believed to be a key regulator in both lytic and latent infections. Using yeast two-hybrid screening, small ubiquitin-like modifiers (SUMO-1, SUMO-2, and SUMO-3) and a SUMO-conjugating enzyme (Ubc9) were isolated as IE2-interacting proteins. In vitro binding assays with glutathione S transferase (GST) fusion proteins provided evidence for direct protein-protein interaction. Mapping data showed that the C-terminal end of SUMO-1 is critical for interaction with IE2 in both yeast and in vitro binding assays. IE2 was efficiently modified by SUMO-1 or SUMO-2 in cotransfected cells and in cells infected with a recombinant adenovirus expressing HCMV IE2, although the level of modification was much lower in HCMV-infected cells. Two lysine residues at positions 175 and 180 were mapped as major alternative SUMO-1 conjugation sites in both cotransfected cells and an in vitro sumoylation assay and could be conjugated by SUMO-1 simultaneously. Although mutations of these lysine residues did not interfere with the POD (or ND10) targeting of IE2, overexpression of SUMO 1 enhanced IE2-mediated transactivation in a promoter-dependent manner in reporter assays. Interestingly, many other cellular proteins identified as IE2 interaction partners in yeast two-hybrid assays also interact with SUMO-1, suggesting that either directly bound or covalently conjugated SUMO moieties may act as a bridge for interactions between IE2 and other SUMO-1-modified or SUMO-1 interacting proteins. When we investigated the intracellular localization of SUMO 1 in HCMV-infected cells, the pattern changed from nuclear punctate to predominantly nuclear diffuse in an IE1-dependent manner at very early times after infection, but with some SUMO-1 protein now associated with IE2 punctate domains. However, at late times after infection, SUMO-1 was predominantly detected within viral DNA replication compartments containing IE2. Taken together, these results show that HCMV infection causes the redistribution of SUMO-1 and that IE2 both physically binds to and is covalently modified by SUMO moieties, suggesting possible modulation of both the function of SUMO-1 and protein-protein interactions of IE2 during HCMV infection. PMID- 11264376 TI - Biogenesis of the Semliki Forest virus RNA replication complex. AB - The nonstructural (ns) proteins nsP1 to -4, the components of Semliki Forest virus (SFV) RNA polymerase, were localized in infected cells by confocal microscopy using double labeling with specific antisera against the individual ns proteins. All ns proteins were associated with large cytoplasmic vacuoles (CPV), the inner surfaces of which were covered by small invaginations, or spherules, typical of alphavirus infection. All ns proteins were localized by immuno electron microscopy (EM) to the limiting membranes of CPV and to the spherules, together with newly labeled viral RNA. Along with earlier observations by EM autoradiography (P. M. Grimley, I. K. Berezesky, and R. M. Friedman, J. Virol. 2:326-338, 1968), these results suggest that individual spherules represent template-associated RNA polymerase complexes. Immunoprecipitation of radiolabeled ns proteins showed that each antiserum precipitated the other three ns proteins, implying that they functioned as a complex. Double labeling with organelle specific and anti-ns-protein antisera showed that CPV were derivatives of late endosomes and lysosomes. Indeed, CPV frequently contained endocytosed bovine serum albumin-coated gold particles, introduced into the medium at different times after infection. With time, increasing numbers of spherules were also observed on the cell surfaces; they were occasionally released into the medium, probably by secretory lysosomes. We suggest that the spherules arise by primary assembly of the RNA replication complexes at the plasma membrane, guided there by nsP1, which has affinity to lipids specific for the cytoplasmic leaflet of the plasma membrane. Endosomal recycling and fusion of CPV with the plasma membrane can circulate spherules between the plasma membrane and the endosomal-lysosomal compartment. PMID- 11264377 TI - Herpes simplex virus type 1 promoter activity during latency establishment, maintenance, and reactivation in primary dorsal root neurons in vitro. AB - A neonatal rat dorsal root ganglion-derived neuronal culture system has been utilized to study herpes simplex virus (HSV) latency establishment, maintenance, and reactivation. We present our initial characterization of viral gene expression in neurons following infection with replication-defective HSV recombinants carrying beta-galactosidase and/or green fluorescent protein reporter genes under the control of lytic cycle- or latency-associated promoters. In this system lytic virus reporter promoter activity was detected in up to 58% of neurons 24 h after infection. Lytic cycle reporter promoters were shut down over time, and long-term survival of neurons harboring latent virus genomes was demonstrated. Latency-associated promoter-driven reporter gene expression was detected in neurons from early times postinfection and was stably maintained in up to 83% of neurons for at least 3 weeks. In latently infected cultures, silent lytic cycle promoters could be activated in up to 53% of neurons by nerve growth factor withdrawal or through inhibition of histone deacetylases by trichostatin A. We conclude that the use of recombinant viruses containing reporter genes, under the regulation of lytic and latency promoter control in neuronal cultures in which latency can be established and reactivation can be induced, is a potentially powerful system in which to study the molecular events that occur during HSV infection of neurons. PMID- 11264378 TI - Canine and feline parvoviruses can use human or feline transferrin receptors to bind, enter, and infect cells. AB - Canine parvovirus (CPV) enters and infects cells by a dynamin-dependent, clathrin mediated endocytic pathway, and viral capsids colocalize with transferrin in perinuclear vesicles of cells shortly after entry (J. S. L. Parker and C. R. Parrish, J. Virol. 74:1919-1930, 2000). Here we report that CPV and feline panleukopenia virus (FPV), a closely related parvovirus, bind to the human and feline transferrin receptors (TfRs) and use these receptors to enter and infect cells. Capsids did not detectably bind or enter quail QT35 cells or a Chinese hamster ovary (CHO) cell-derived cell line that lacks any TfR (TRVb cells). However, capsids bound and were endocytosed into QT35 cells and CHO-derived TRVb 1 cells that expressed the human TfR. TRVb-1 cells or TRVb cells transiently expressing the feline TfR were susceptible to infection by CPV and FPV, but the parental TRVb cells were not. We screened a panel of feline-mouse hybrid cells for susceptibility to FPV infection and found that only those cells that possessed feline chromosome C2 were susceptible. The feline TfR gene (TRFC) also mapped to feline chromosome C2. These data indicate that cell susceptibility for these viruses is determined by the TfR. PMID- 11264380 TI - Separation of human immunodeficiency virus type 1 replication from nef-mediated pathogenesis in the human thymus. AB - Human immunodeficiency virus type 1 (HIV-1) is frequently attenuated after long term culture in vitro. The attenuation process probably involves mutations of functions required for replication and pathogenicity in vivo. Analysis of attenuated HIV-1 for replication and pathogenicity in vivo will help to define these functions. In this study, we examined the pathogenicity of an attenuated HIV-1 isolate in a laboratory worker accidentally exposed to a laboratory-adapted HIV-1 isolate. Using heterochimeric SCID-hu Thy/Liv mice as an in vivo model, we previously defined HIV-1 env determinants (HXB/LW) that reverted to replicate in vivo (L. Su, H. Kaneshima, M. L. Bonyhadi, R. Lee, J. Auten, A. Wolf, B. Du, L. Rabin, B. H. Hahn, E. Terwilliger, and J. M. McCune, Virology 227:46-52, 1997). Here we further demonstrate that HIV-1 replication in vivo can be separated from its pathogenic activity, in that the HXB/LW virus replicated to high levels in SCID-hu Thy/Liv mice, with no significant thymocyte depletion. Restoration of the nef gene in the recombinant HXB/LW genome restored its pathogenic activity, with no significant effect on HIV-1 replication in the thymus. Our results suggest that in vitro-attenuated HIV-1 lacks determinants for pathogenicity as well as for replication in vivo. Our data indicate that (i) the replication defect can be recovered in vivo by mutations in the env gene, without an associated pathogenic phenotype, and (ii) nef can function in the HXB/LW clone as a pathogenic factor that does not enhance HIV-1 replication in the thymus. Furthermore, the HXB/LW virus may be used to study mechanisms of HIV-1 nef-mediated pathogenesis in vivo. PMID- 11264379 TI - Ability of the V3 loop of simian immunodeficiency virus to serve as a target for antibody-mediated neutralization: correlation of neutralization sensitivity, growth in macrophages, and decreased dependence on CD4. AB - To better define the effects of sequence variation and tropism on the ability of the simian immunodeficiency virus SIVmac V3 loop to act as a target of antibody mediated neutralization, a series of experiments were performed. Three SIV strains, SIVmac239, SIVmac316, and SIVmac155/T3, each with defined differences in env sequence and tropism, were used to construct a panel of viruses chimeric for a portion of envelope that includes the V2 and V3 regions. Peptides with sequences corresponding to the V3 loops of the parental viruses were used to immunize rabbits. The polyclonal rabbit antibodies and plasma from SIVmac239 infected animals were then used to assess the neutralization sensitivity of the parental and chimeric viruses. One of the parental viruses, SIVmac316, which is able to replicate to high titer in alveolar macrophages and can infect cells in a CD4-independent fashion, was highly sensitive to neutralization by plasma from SIVmac-infected rhesus macaques, with average 50% neutralization titers of 1:20,480; this same strain was also sensitive to neutralization by the anti-V3 loop peptide sera. Other parental and chimeric viruses were less sensitive to neutralization with this same panel of antibodies, but as seen with SIVmac316, those viruses that were able to productively replicate in alveolar macrophages were more sensitive to antibody-mediated neutralization. To further define the amino acids involved in increased sensitivity to neutralization, a panel of viruses was constructed by changing envelope residues in SIVmac316 to the corresponding SIVmac239 amino acids. The increased neutralization sensitivity observed for SIVmac316 was mapped principally to three amino acid changes spread throughout gp120. In addition, the increased sensitivity to neutralization by V3 directed antibodies correlated with the ability of the various viruses to replicate to high levels in alveolar macrophage cultures and a CD4-negative cell line, BC7/CCR5. These results demonstrate that the V3 loop of SIVmac Env can act as an efficient target of neutralizing antibodies in a fashion that is highly dependent on sequence context. In addition, these studies suggest a correlation between decreased dependence on CD4 and increased sensitivity to antibody mediated neutralization. PMID- 11264381 TI - Sequence requirements for interaction of human herpesvirus 7 origin binding protein with the origin of lytic replication. AB - As do human herpesvirus 6 variants A and B (HHV-6A and -6B), HHV-7 encodes a homolog of the alphaherpesvirus origin binding protein (OBP), which binds at sites in the origin of lytic replication (oriLyt) to initiate DNA replication. In this study, we sought to characterize the interaction of the HHV-7 OBP (OBP(H7)) with its cognate sites in the 600-bp HHV-7 oriLyt. We expressed the carboxyl terminal domain of OBP(H7) and found that amino acids 484 to 787 of OBP(H7) were sufficient for DNA binding activity by electrophoretic mobility shift analysis. OBP(H7) has one high-affinity binding site (OBP-2) located on one flank of an AT rich spacer element and a low-affinity site (OBP-1) on the other. This is in contrast to the HHV-6B OBP (OBP(H6B)), which binds with similar affinity to its two cognate OBP sites in the HHV-6B oriLyt. The minimal recognition element of the OBP-2 site was mapped to a 14-bp sequence. The OBP(H7) consensus recognition sequence of the 9-bp core, BRTYCWCCT (where B is a T, G, or C; R is a G or A; Y is a T or C; and W is a T or A), overlaps with the OBP(H6B) consensus YGWYCWCCY and establishes YCWCC as the roseolovirus OBP core recognition sequence. Heteroduplex analysis suggests that OBP(H7) interacts along one face of the DNA helix, with the major groove, as do OBP(H6B) and herpes simplex virus type 1 OBP. Together, these results illustrate both conserved and divergent DNA binding properties between OBP(H7) and OBP(H6B). PMID- 11264384 TI - Direct in vitro binding of full-length human immunodeficiency virus type 1 Nef protein to CD4 cytoplasmic domain. AB - The Nef protein of the simian and human immunodeficiency viruses is known to directly bind and downregulate the CD4 receptor. Although the molecular mechanism is well understood, direct binding of Nef and CD4 is difficult to demonstrate and is believed to be of low affinity. Applying nuclear magnetic resonance and fluorescence spectroscopy, we biophysically reevaluated the CD4-Nef complex and found the dissociation constant to be in the submicromolar range. We conclude that additional, so far disregarded residues in the N terminus of Nef are important for interaction with CD4. PMID- 11264383 TI - Close but distinct regions of human herpesvirus 8 latency-associated nuclear antigen 1 are responsible for nuclear targeting and binding to human mitotic chromosomes. AB - Human herpesvirus 8 is associated with all forms of Kaposi's sarcoma, AIDS associated body cavity-based lymphomas, and some forms of multicentric Castleman's disease. Herpesvirus 8, like other gammaherpesviruses, can establish a latent infection in which viral genomes are stably maintained as multiple episomes. The latent nuclear antigen (LANA or LNAI) may play an essential role in the stable maintenance of latent episomes, notably by interacting concomitantly with the viral genomes and the metaphase chromosomes, thus ensuring an efficient transmission of the neoduplicated episomes to the daughter cells. To identify the regions responsible for its nuclear and subnuclear localization in interphase and mitotic cells, LNAI and various truncated forms were fused to a variant of green fluorescent protein. This enabled their localization and chromosome binding activity to be studied by low-light-level fluorescence microscopy in living HeLa cells. The results demonstrate that nuclear localization of LNAI is due to a unique signal, which maps between amino acids 24 and 30. Interestingly, this nuclear localization signal closely resembles those identified in EBNA1 from Epstein-Barr virus and herpesvirus papio. A region encompassing amino acids 5 to 22 was further proved to mediate the specific interaction of LNA1 with chromatin during interphase and the chromosomes during mitosis. The presence of putative phosphorylation sites in the chromosome binding sites of LNA1 and EBNA1 suggests that their activity may be regulated by specific cellular kinases. PMID- 11264382 TI - RGD tripeptide of bluetongue virus VP7 protein is responsible for core attachment to Culicoides cells. AB - Bluetongue virus (BTV) is an arthropod-borne virus transmitted by Culicoides species to vertebrate hosts. The double-capsid virion is infectious for Culicoides vector and mammalian cells, while the inner core is infectious for only Culicoides-derived cells. The recently determined crystal structure of the BTV core has revealed an accessible RGD motif between amino acids 168 to 170 of the outer core protein VP7, whose structure and position would be consistent with a role in cell entry. To delineate the biological role of the RGD sequence within VP7, we have introduced point mutations in the RGD tripeptide and generated three recombinant baculoviruses, each expressing a mutant derivative of VP7 (VP7-AGD, VP7-ADL, and VP7-AGQ). Each expressed mutant protein was purified, and the oligomeric nature and secondary structure of each was compared with those of the wild-type (wt) VP7 molecule. Each mutant VP7 protein was used to generate empty core-like particles (CLPs) and were shown to be biochemically and morphologically identical to those of wt CLPs. However, when mutant CLPs were used in an in vitro cell binding assay, each showed reduced binding to Culicoides cells compared to wt CLPs. Twelve monoclonal antibodies (MAbs) was generated using purified VP7 or CLPs as a source of antigen and were utilized for epitope mapping with available chimeric VP7 molecules and the RGD mutants. Several MAbs bound to the RGD motif on the core, as shown by immunogold labeling and cryoelectron microscopy. RGD specific MAb H1.5, but not those directed to other regions of the core, inhibited the binding activity of CLPs to the Culicoides cell surface. Together, these data indicate that the RGD motif present on BTV VP7 is responsible for Culicoides cell binding activity. PMID- 11264385 TI - Propagation of rat parvovirus in thymic lymphoma cell line C58(NT)d and subsequent appearance of a resistant cell clone after lytic infection. AB - Rat parvovirus (RPV) is nonpathogenic in rats but causes persistent lymphocytotropic infection. We found that RPV was propagated in rat thymic lymphoma cell line C58(NT)D and induced apoptosis. Interestingly, a resistant subclone, C58(NT)D/R, from surviving cells after lytic infection had differentiated phenotypic modifications, such as increased cell adherence, resistance to apoptosis, and suppressed tumorigenicity. PMID- 11264386 TI - Nef-induced CD4 downregulation: a diacidic sequence in human immunodeficiency virus type 1 Nef does not function as a protein sorting motif through direct binding to beta-COP. AB - The Nef protein from the human immunodeficiency virus (HIV) induces CD4 cell surface downregulation by interfering with the endocytic machinery. It has been recently proposed that binding of HIV type 1 Nef to the beta subunit of COPI coatomers participated in the Nef-induced CD4 downregulation through recognition of a novel diacidic motif found in the C-terminal disordered loop of Nef (V. Piguet, F. Gu, M. Foti, N. Demaurex, J. Gruenberg, J. L. Carpentier, and D. Trono, Cell 97:63-73, 1999). We have mutated the glutamate residues which formed this motif in order to document this observation. Surprisingly, mutation of the diacidic sequence of Nef did not significantly affect its ability (i) to interact with beta-COP, (ii) to downregulate CD4 cell surface expression, and (iii) to address an integral resident membrane protein containing Nef as the cytoplasmic domain to the endocytic pathway. Our results indicate that these acidic residues are not involved in the connection of Nef with the endocytic machinery through binding to beta-COP. Additional studies are thus required to characterize the residues of Nef involved in the binding to beta-COP and to evaluate the contribution of this interaction to the Nef-induced perturbations of membrane trafficking. PMID- 11264388 TI - Degenerate immunogenicity of an HLA-A2-restricted hepatitis B virus nucleocapsid cytotoxic T-lymphocyte epitope that is also presented by HLA-B51. AB - The recent identification of hepatitis B virus (HBV) epitopes restricted by multiple HLA alleles has greatly expanded the epitope repertoire available for T cell-mediated therapeutic vaccine development. The HLA-B51-restricted peptide HBc19-27 is particularly interesting because it is located entirely within the HLA-A2-restricted HBc18-27 epitope. Here we show that HLA-B51-restricted cytotoxic T lymphocytes specific for HBc19-27 from a patient with acute HBV infection were also able to lyse HLA-B51-positive target cells pulsed with HBc18 27 and to produce gamma interferon when stimulated by that peptide, implying that HBc18-27 can be presented by HLA-B51 as well as by HLA-A2. These results demonstrate the concept of degenerate immunogenicity across HLA class supertype boundaries in a human viral disease setting. In addition, they could facilitate the development of an epitope-based therapeutic vaccine to terminate chronic HBV infection that could provide a broad and diverse population coverage with a single peptide. PMID- 11264389 TI - Amino acid deletion at codon 67 and Thr-to-Gly change at codon 69 of human immunodeficiency virus type 1 reverse transcriptase confer novel drug resistance profiles. AB - The potential roles of an amino acid deletion at codon 67 (Delta67) and a Thr-to Gly change at codon 69 (T69G) in the reverse transcriptase of human immunodeficiency virus (HIV) type 1 in drug sensitivity and relative replication fitness were studied. Our results suggest that the Delta67 and T69G changes can be categorized as mutations associated with multidrug resistance. The combination of both mutations with an L74I change (Delta67+T69G/L74I) leads to a novel 3' azido-3'-deoxythymidine resistance motif and compensates for impaired HIV replication. PMID- 11264387 TI - A single amino acid substitution in nonstructural protein 3A can mediate adaptation of foot-and-mouth disease virus to the guinea pig. AB - The genetic changes selected during the adaptation of a clonal population of foot and-mouth disease virus (FMDV) to the guinea pig have been analyzed. FMDV clone C S8c1 was adapted to the guinea pig by serial passage in the animals until secondary lesions were observed. Analysis of the virus directly recovered from the lesions developed by the animals revealed the selection of variants with two amino acid substitutions in nonstructural proteins, I(248)-->T in 2C and Q(44)- >R in 3A. On further passages, an additional mutation, L(147)-->P, was selected in an important antigenic site located in the G-H loop of capsid protein VP1. The amino acid substitution Q(44)-->R in 3A, either alone or in combination with the replacement I(248)-->T in 2C, was sufficient to give FMDV the ability to produce lesions. This was shown by using infectious transcripts which generated chimeric viruses with the relevant amino acid substitutions. Clinical symptoms produced by the artificial chimeras were similar to those produced by the naturally adapted virus. These results obtained with FMDV imply that one or very few replacements in nonstructural viral proteins, which should be within reach of the mutant spectra of replicating viral quasispecies, may result in adaptation of a virus to a new animal host. PMID- 11264391 TI - Langat virus M protein is structurally homologous to prM. AB - Langat (LGT) virus M protein has been generated in a recombinant system. Antiserum raised against the LGT virus M protein neutralizes tick-borne encephalitis serocomplex flaviviruses but not mosquito-borne flaviviruses, indicating that the M protein is exposed on the surface of virions. The antiserum recognizes intracellular LGT virus prM/M and binds to prM and M in Western blots of whole-cell lysates and purified virus, respectively. These data suggest that the prM and M proteins are structurally similar under native conditions and support the hypothesis that the "pr" portion of prM facilitates proper folding of the M protein for expression on the virion surface. PMID- 11264390 TI - RNA replication from the simian virus 5 antigenomic promoter requires three sequence-dependent elements separated by sequence-independent spacer regions. AB - We have previously shown for the paramyxovirus simian virus 5 (SV5) that a functional promoter for RNA replication requires proper spacing between two discontinuous elements: a 19-base segment at the 3' terminus (conserved region I [CRI]) and an 18-base internal region (CRII) that is contained within the coding region of the L protein gene. In the work described here, we have used a reverse genetics system to determine if the 53-base segment between CRI and CRII contains additional sequence-specific signals required for optimal replication or if this segment functions solely as a sequence-independent spacer region. A series of copyback defective interfering minigenome analogs were constructed to contain substitutions of nonviral sequences in place of bases 21 to 72 of the antigenomic promoter, and the relative level of RNA replication was measured by Northern blot analysis. The results from our mutational analysis indicate that in addition to CRI and CRII, optimal replication from the SV5 antigenomic promoter requires a third sequence-dependent element located 51 to 66 bases from the 3' end of the RNA. Minigenome RNA replication was not affected by changes in the either the position of this element in relation to CRI and CRII or the predicted hexamer phase of NP encapsidation. Thus, optimal RNA replication from the SV5 antigenomic promoter requires three sequence-dependent elements, CRI, CRII and bases 51 to 66. PMID- 11264392 TI - Biophysical characterization and vector-specific antagonist activity of domain III of the tick-borne flavivirus envelope protein. AB - The molecular determinants responsible for flavivirus host cell binding and tissue tropism are largely unknown, although domain III of the envelope protein has been implicated in these functions. We examined the solution properties and antagonist activity of Langat virus domain III. Our results suggest that domain III adopts a stably folded structure that can mediate binding of tick-borne flaviviruses but not mosquito-borne flaviviruses to their target cells. Three clusters of phylogenetically conserved residues are identified that may be responsible for the vector-specific antagonist activity of domain III. PMID- 11264393 TI - Herpesvirus saimiri open reading frame 50 (Rta) protein reactivates the lytic replication cycle in a persistently infected A549 cell line. AB - Herpesviruses occur in two distinct forms of infection, lytic replication and latent persistence. In this study, we investigated the molecular mechanisms that govern the latent-lytic switch in the prototype gamma-2 herpesvirus, herpesvirus saimiri (HVS). We utilized a persistently HVS-infected A549 cell line, in which HVS DNA is stably maintained as nonintegrated circular episomes, to assess the role of the open reading frame 50 (ORF 50) (Rta) proteins in the latent-lytic switch. Northern blot analysis and virus recovery assays determined that the ORF 50a gene product, when expressed under the control of a constitutively active promoter, was sufficient to reactivate the entire lytic replication cycle, producing infectious virus particles. Furthermore, although the ORF 50 proteins of HVS strains A11 and C488 are structurally divergent, they were both capable of inducing the lytic replication cycle in this model of HVS latency. PMID- 11264394 TI - Human immunodeficiency virus type 1 particles pseudotyped with envelope proteins that fuse at low pH no longer require Nef for optimal infectivity. AB - We have investigated the effects of Nef on infectivity in the context of various viral envelope proteins. These experiments were performed with a minimal vector system where Nef is the only accessory protein present. Our results support the hypothesis that the route of entry influences the ability of Nef to enhance human immunodeficiency virus (HIV) infectivity. We show that HIV particles pseudotyped with Ebola virus glycoprotein or vesicular stomatitis virus glycoprotein (VSV-G), which fuse at low pH, do not require Nef for optimal infectivity. In contrast, Nef significantly enhances the infectivity of virus particles that contain envelope proteins that fuse at neutral pH (CCR5-dependent HIV Env, CXCR4 dependent HIV Env, or amphotropic murine leukemia virus Env). In addition, our results demonstrate that virus particles containing mixed CXCR4-dependent HIV and VSV-G envelope proteins show a conditional requirement for Nef for optimal infectivity, depending on which protein is allowed to facilitate entry. PMID- 11264396 TI - Maximum-likelihood approach for gene family evolution under functional divergence. AB - According to the observed alignment pattern (i.e., amino acid configuration), we studied two basic types of functional divergence of a protein family. Type I functional divergence after gene duplication results in altered functional constraints (i.e., different evolutionary rate) between duplicate genes, whereas type II results in no altered functional constraints but radical change in amino acid property between them (e.g., charge, hydrophobicity, etc.). Two statistical approaches, i.e., the subtree likelihood and the whole-tree likelihood, were developed for estimating the coefficients of (type I or type II) functional divergence. Numerical algorithms for obtaining maximum-likelihood estimates are also provided. Moreover, a posterior-based site-specific profile is implemented to predict critical amino acid residues that are responsible for type I and/or type II functional divergence after gene duplication. We compared the current likelihood with a fast method developed previously by examples; both show similar results. For handling altered functional constraints (type I functional divergence) in the large gene family with many member genes (clusters), which appears to be a normal case in postgenomics, the subtree likelihood provides a solution that is computationally feasible and robust against the uncertainty of the phylogeny. The cost of this feasibility is the approximation when frequencies of amino acids are very skewed. The potential bias and correction are discussed. PMID- 11264397 TI - Cytochrome b phylogeny and the taxonomy of great apes and mammals. AB - In the Linnaean system of classification, the generic status of a species is part of its binomial name, and it is therefore important that the classification at the level of genus is consistent at least in related groups of organisms. Using maximum-likelihood phylogenetic trees constructed from a large number of complete or nearly complete mammalian cytochrome b sequences, I show that the distributions of intrageneric and intergeneric distances derived from these trees are clearly separated, which allows the limits for a more rational generic classification of mammals to be established. The analysis of genetic distances among hominids in this context provides strong support for the inclusion of humans and chimpanzees in the same genus. It is also of interest to decipher the main reasons for the possible biases in the mammalian classification. I found by correlation analysis that the classification of mammals of large body size tends to be oversplit, whereas that of small mammals has an excess of lumping, which may be a manifestation of the larger difficulty in finding diagnostic characters in the classification of small animals. In addition, and contrary to some previous observations, there is no correlation between body size and rate of cytochrome b evolution in mammals, which excludes the difference in evolutionary rates as the cause of the observed body size taxonomic bias. PMID- 11264395 TI - Suboptimal nucleotides in the infectious, pathogenic simian immunodeficiency virus clone SIVmac239. AB - We analyzed virus sequences in two monkeys infected with SIVmac239 and two monkeys infected with SHIVnef that maintained high, persisting viral loads. Sequence changes were observed consistently at four loci in all four animals: a single nucleotide change in the Lys-tRNA primer binding site in the 5' long terminal repeat; two nucleotide changes that resulted in two amino acid changes in the pol gene product; and a single nucleotide change in the region of the simian immunodeficiency virus genome where the rev and env genes overlap, resulting in changes in the predicted amino acid sequences of both gene products. None of these mutations were seen in short-term cultures of CEMx174 cells infected with SIVmac239 or SHIVnef. At all four positions in all four animals, the new sequences represented consensus sequences for primate lentiviruses, whereas the inoculum sequences at these four loci have either never been or rarely been reported outside of SIVmac239. Thus, although cloned SIVmac239 is consistently pathogenic and consistently induces high viral load set points, it is clearly less than optimal at these four nucleotide positions. PMID- 11264398 TI - Speciation and intrasubspecific variation of Bornean orangutans, Pongo pygmaeus pygmaeus. AB - Mitochondrial DNA control region sequences of orangutans (Pongo pygmaeus) from six different populations on the island of Borneo were determined and analyzed for evidence of regional diversity and were compared separately with orangutans from the island of Sumatra. Within the Bornean population, four distinct subpopulations were identified. Furthermore, the results of this study revealed marked divergence, supportive evidence of speciation between Sumatran and Bornean orangutans. This study demonstrates that, as an entire population, Bornean orangutans have not experienced a serious genetic bottleneck, which has been suggested as the cause of low diversity in humans and east African chimpanzees. Based on these new data, it is estimated that Bornean and Sumatran orangutans diverged approximately 1.1 MYA and that the four distinct Bornean populations diverged 860,000 years ago. These findings have important implications for management, breeding, and reintroduction practices in orangutan conservation efforts. PMID- 11264399 TI - Models of sequence evolution for DNA sequences containing gaps. AB - Most evolutionary tree estimation methods for DNA sequences ignore or inefficiently use the phylogenetic information contained within shared patterns of gaps. This is largely due to the computational difficulties in implementing models for insertions and deletions. A simple way to incorporate this information is to treat a gap as a fifth character (with the four nucleotides being the other four) and to incorporate it within a Markov model of nucleotide substitution. This idea has been dismissed in the past, since it treats a multiple-site insertion or deletion as a sequence of independent events rather than a single event. While this is true, we have found that under many circumstances it is better to incorporate gap information inadequately than to ignore it, at least for topology estimation. We propose an extension to a class of nucleotide substitution models to incorporate the gap character and show that, for data sets (both real and simulated) with short and medium gaps, these models do lead to effective use of the information contained within insertions and deletions. We also implement an ad hoc method in which the likelihood at columns containing multiple-site gaps is downweighted in order to avoid giving them undue influence. The precision of the estimated tree, assessed using Markov chain Monte Carlo techniques to find the posterior distribution over tree space, improves under these five-state models compared with standard methods which effectively ignore gaps. PMID- 11264400 TI - Isolation of homeodomain-leucine zipper genes from the moss Physcomitrella patens and the evolution of homeodomain-leucine zipper genes in land plants. AB - Homeobox genes encode transcription factors involved in many aspects of developmental processes. The homeodomain-leucine zipper (HD-Zip) genes, which are characterized by the presence of both a homeodomain and a leucine zipper motif, form a clade within the homeobox superfamily and were previously reported only from vascular plants. Here we report the isolation of 10 HD-Zip genes (named PPHB:1-PPHB:10) from the moss Physcomitrella patens. Based on a phylogenetic analysis of the 10 PPHB: genes and previously reported vascular plant HD-Zip genes, all of the PPHB: genes except Pphb3 belong to three of the four HD-Zip subfamilies (HD-Zip I, II, and III), indicating that these subfamilies originated before the divergence of the vascular plant and moss lineages. Pphb3 is sister to the HD-Zip II subfamily and has some distinctive characteristics, including the difference of the a(1) and d(1) sites of its leucine zipper motif, which are well conserved in each HD-Zip subfamily. Comparison of the genetic divergence of representative HD-Zip I and II genes showed that the evolutionary rate of HD-Zip I genes was faster than that of HD-Zip II genes. PMID- 11264401 TI - Evolutionary dynamics of the T-cell receptor VB gene family as inferred from the human and mouse genomic sequences. AB - The diversity of T-cell receptors is generated primarily by the variable-region gene families, each of which is composed of a large number of member genes. The entire genomic sequence of the variable region (VB) of the T- cell receptor beta chain from humans and mice has become available. To understand the evolutionary dynamics of the VB gene family, we conducted a phylogenetic analysis of all VB genes from humans and mice, as well as a detailed analysis of internal DNA duplications in the human genomic VB region. The phylogenetic tree obtained shows that human and mouse VB genes intermingle extensively rather than forming two separate clusters and that many gene duplications occurred both before and after the divergence between primates and rodents. Analyzing the genomic maps of transposable elements (e.g., LINEs and SINEs) and relic VB genes in the VB gene region, we present evidence that a 20-kb VB region duplicated tandemly four times in the human lineage during the last 32 Myr, and 6 out of the 15 VB genes in this region have become nonfunctional during this period. Our results show that the VB gene family is subject to evolution by a birth-and-death process rather than to concerted evolution. PMID- 11264402 TI - Evolutionary relationships among "jakobid" flagellates as indicated by alpha- and beta-tubulin phylogenies. AB - Jakobids are free-living, heterotrophic flagellates that might represent early diverging mitochondrial protists. They share ultrastructural similarities with eukaryotes that occupy basal positions in molecular phylogenies, and their mitochondrial genome architecture is eubacterial-like, suggesting a close affinity with the ancestral alpha-proteobacterial symbiont that gave rise to mitochondria and hydrogenosomes. To elucidate relationships among jakobids and other early-diverging eukaryotic lineages, we characterized alpha- and beta tubulin genes from four jakobids: Jakoba libera, Jakoba incarcerata, Reclinomonas americana (the "core jakobids"), and Malawimonas jakobiformis. These are the first reports of nuclear genes from these organisms. Phylogenies based on alpha-, beta-, and combined alpha- plus beta-tubulin protein data sets do not support the monophyly of the jakobids. While beta-tubulin and combined alpha- plus beta tubulin phylogenies showed a sister group relationship between J. libera and R. americana, the two other jakobids, M. jakobiformis and J. incarcerata, had unclear affinities. In all three analyses, J. libera, R. americana, and M. jakobiformis emerged from within a well-supported large "plant-protist" clade that included plants, green algae, cryptophytes, stramenopiles, alveolates, Euglenozoa, Heterolobosea, and several other protist groups, but not animals, fungi, microsporidia, parabasalids, or diplomonads. A preferred branching order within the plant-protist clade was not identified, but there was a tendency for the J. libera-R. americana lineage to group with a clade made up of the heteroloboseid amoeboflagellates and euglenozoan protists. Jakoba incarcerata branched within the plant-protist clade in the beta- and the combined alpha- plus beta-tubulin phylogenies. In alpha- tubulin trees, J. incarcerata occupied an unresolved position, weakly grouping with the animal/fungal/microsporidian group or with amitochondriate parabasalid and diplomonad lineages, depending on the phylogenetic method employed. Tubulin gene phylogenies were in general agreement with mitochondrial gene phylogenies and ultrastructural data in indicating that the "jakobids" may be polyphyletic. Relationships with the putatively deep branching amitochondriate diplomonads remain uncertain. PMID- 11264403 TI - Positive and negative selection in the DAZ gene family. AB - Because a microdeletion containing the DAZ gene is the most frequently observed deletion in infertile men, the DAZ gene was considered a strong candidate for the azoospermia factor. A recent evolutionary analysis, however, suggested that DAZ was free from functional constraints and consequently played little or no role in human spermatogenesis. The major evidence for this surprising conclusion is that the nonsynonymous substitution rate is similar to the synonymous rate and to the rate in introns. In this study, we reexamined the evolution of the DAZ gene family by using maximum-likelihood methods, which accommodate variable selective pressures among sites or among branches. The results suggest that DAZ is not free from functional constraints. Most amino acids in DAZ are under strong selective constraint, while a few sites are under diversifying selection with nonsynonymous/ synonymous rate ratios (d(N)/d(S)) well above 1. As a result, the average d(N)/d(S) ratio over sites is not a sensible measure of selective pressure on the protein. Lineage-specific analysis indicated that human members of this gene family were evolving by positive Darwinian selection, although the evidence was not strong. PMID- 11264404 TI - Giardia lamblia expresses a proteobacterial-like DnaK homolog. AB - We identified a novel gene encoding molecular chaperone HSP70 in the amitochondriate parasite Giardia lamblia. The predicted protein is similar to bacterial DnaK and mitochondrial HSP70s. The gene is transcribed and translated at a constant level during trophozoite growth and encystation. Alignment of the sequence with a data set of cytosolic, endoplasmic reticulum (ER), mitochondrial, and DnaK HSP70 homologs indicated that the sequence was extremely divergent and contained insertions unique to giardial HSP70s. Phylogenetic analyses demonstrated that this sequence was distinct from the cytosolic and ER forms and was most similar to proteobacterial and mitochondrial DnaKs. However, a specific relationship with the alpha proteobacterial and mitochondrial sequences was not strongly supported by phylogenetic analyses of this data set, in contrast to similar analyses of cpn60. These data neither confirm nor reject the possibility that this gene is a relic of secondary mitochondrial loss; they leave open the possibility that it was acquired in a separate endosymbiotic event. PMID- 11264405 TI - Two Cyp19 (P450 aromatase) genes on duplicated zebrafish chromosomes are expressed in ovary or brain. AB - Cytochrome P450 aromatase (Cyp19) is an enzyme catalyzing the synthesis of estrogens, thereby controlling various physiological functions of estrogens. We isolated two cyp19 cDNAs, termed cyp19a and cyp19b, respectively, from zebrafish. These genes are located in linkage groups 18 and 25, respectively. Detailed gene mapping indicated that zebrafish linkage groups 18 and 25 may have arisen from the same ancestral chromosome by a chromosome duplication event. Cyp19a is expressed mainly in the follicular cells lining the vitellogenic oocytes in the ovary during vitellogenesis. Cyp19b is expressed abundantly in the brain, at the hypothalamus and ventral telencephalon, extending to the olfactory bulbs. The expression of duplicated cyp19 genes at two different tissues highlights the evolutionary significance of maintaining two active genes on duplicated zebrafish chromosomes for specific functions in the ovary and the brain. PMID- 11264407 TI - Differential selection after duplication in mammalian developmental genes. AB - Gene duplication provides the opportunity for subsequent refinement of distinct functions of the duplicated copies. Either through changes in coding sequence or changes in regulatory regions, duplicate copies appear to obtain new or tissue specific functions. If this divergence were driven by natural selection, we would expect duplicated copies to have differentiated patterns of substitutions. We tested this hypothesis using genes that duplicated before the human/mouse split and whose orthologous relations were clear. The null hypothesis is that the number of amino acid changes between humans and mice was distributed similarly across different paralogs. We used a method modified from Tang and Lewontin to detect heterogeneity in the amino acid substitution pattern between those different paralogs. Our results show that many of the paralogous gene pairs appear to be under differential selection in the human/mouse comparison. The properties that led to diversification appear to have arisen before the split of the human and mouse lineages. Further study of the diverged genes revealed insights regarding the patterns of amino acid substitution that resulted in differences in function and/or expression of these genes. This approach has utility in the study of newly identified members of gene families in genomewide data mining and for contrasting the merits of alternative hypotheses for the evolutionary divergence of function of duplicated genes. PMID- 11264406 TI - Evolutionary history of microsatellites in the obscura group of Drosophila. AB - The evolutionary origins of microsatellites are not well understood. Some investigators have suggested that point mutations that expand repeat arrays beyond a threshold size trigger microsatellites to become variable. However, little empirical data has been brought forth on this and related issues. In this study, we examine the evolutionary history of microsatellites in six species within the obscura group of Drosophila, tracing changes in microsatellite alleles using both PCR product size and sequence data. We found little evidence supporting a general role of point mutations triggering initial microsatellite expansion, and no consistent threshold size for expansion was observed. Flanking region length variation was extensive when alleles were sequenced in distantly related species, and some species possessed altogether different repeat arrays between the same primer binding sites. Our results suggest extreme caution in using microsatellite allele sizes for phylogenetic analyses or to infer divergences between populations. PMID- 11264408 TI - Evolution of nuclear- and mitochondrial-encoded subunit interaction in cytochrome c oxidase. AB - Mitochondrial DNA (mtDNA)-encoded proteins function in eukaryotes as subunits of respiratory complexes that also contain nuclear DNA (nDNA)-encoded subunits. The importance of functional interactions between mtDNA- and nDNA-encoded proteins was previously demonstrated by testing the survivability of cybrid cells or individuals containing nDNA and mtDNA from different populations or species. This report focuses on the multisubunit respiratory complex cytochrome c oxidase (COX), made up of both mtDNA-encoded and nDNA-encoded subunits. A combination of evolutionary and crystallographic data is employed to determine whether rates of nonsynonymous substitutions have been higher, the same, or lower for residues in close proximity that are encoded by a different genome (nDNA or mtDNA). This determination is performed by simply taking the ratio, called the interaction ratio i, of the nonsynonymous substitution rate of the close-contact residues to the nonsynonymous substitution rate of the noncontact residues. We assume that the close-contact residues (which are more likely to interact) are functionally important and that, therefore, amino acid replacements among these residues cannot escape the scrutiny of natural selection. i = 1 indicates that the close contact residues have been under neither greater purifying selection nor greater positive selection than the noncontact residues as a specific consequence of their being encoded by separate genomes. i < 1 indicates that the close-contact residues have been under greater purifying selection but less positive selection than have the noncontact residues. Conversely, i > 1 indicates that the close contact residues have been under less purifying but greater positive selection than have the noncontact residues. i < 1 may be referred to as a constraining interaction; i.e., the close-contact residues compared with the noncontact residues appear to be under greater structural-functional constraints. On the other hand, i > 1 may be referred to as an optimizing interaction; i.e., apparently many different amino acid replacements are required to optimize this subunit's interaction with the other subunit. A major finding is that the nDNA encoded residues in close physical proximity to mtDNA-encoded residues evolve more slowly than the other nuclear-encoded residues (and thus display a constraining interaction), whereas the mtDNA-encoded residues in close physical proximity to nDNA-encoded residues evolve more rapidly than the other mitochondrial-encoded residues (and thus display an optimizing interaction). A possible reason for this striking difference between the nuclear- and mitochondrial-encoded COX subunits in how their functional interaction evolves is discussed. PMID- 11264409 TI - Multimeric hemoglobin of the Australian brine shrimp Parartemia. AB - The hemoglobin molecule of the commercially important brine shrimp Artemia sp. has been used extensively as a model for the study of molecular evolution. It consists of nine globin domains joined by short linker sequences, and these domains are believed to have originated through a series of duplications from an original globin gene. In addition, in Artemia, two different polymers of hemoglobin, called C and T, are found which differ by 11.7% at the amino acid level and are believed to have diverged about 60 MYA. This provides a set of data of 18 globin domain sequences that have evolved in the same organism. The pattern of amino acid substitution between these two polymers is unusual, with pairs of equivalent domains displaying differences of up to 2.7-fold in total amino acid substitution. Such differences would reflect a similar range of molecular-clock rates in what appear to be duplicate, structurally equivalent domains. In order to provide a reference outgroup, we sequenced the cDNA for a nine-domain hemoglobin (P) from another genus of brine shrimp, Parartemia zietziana, which differs morphologically and ecologically from Artemia and is endemic to Australia. Parartemia produces only one hundredth the amount of hemoglobin that Artemia produces and does not upregulate production in response to low oxygen partial pressure. Comparison of the globin domains at the amino acid and DNA levels suggests that the Artemia globin T gene has accumulated substitutions differently from the Parartemia P and Artemia C globin genes. We discuss the questions of accelerated evolution after duplication and possible functions for the Parartemia globin. PMID- 11264410 TI - Vertebrate serpins: construction of a conflict-free phylogeny by combining exon intron and diagnostic site analyses. AB - A combination of three independent biological features, genomic organization, diagnostic amino acid sites, and rare indels, was used to elucidate the phylogeny of the vertebrate serpin (serine protease inhibitor) superfamily. A strong correlation between serpin gene families displaying (1) a conserved exon-intron pattern and (2) family-specific combinations of amino acid residues at specific sites suggests that present-day vertebrates encompass six serpin gene families which evolved from primordial genes by massive intron insertion before or during early vertebrate radiation. Introns placed at homologous positions in the gene sequences in combination with diagnostic sequence characters may also constitute a reliable kinship indicator for other protein superfamilies. PMID- 11264411 TI - Distribution of the bilbo non-LTR retrotransposon in Drosophilidae and its evolution in the Drosophila obscura species group. AB - The bilbo element is a non-LTR retrotransposon isolated from Drosophila subobscura. We conducted a distribution survey by Southern blot for 52 species of the family Drosophilidae, mainly from the obscura and melanogaster groups. Most of the analyzed species bear sequences homologous to bilbo from D. subobscura. In the obscura group, species from the same species subgroup also share similar Southern blot patterns. To investigate the phylogenetic relationship among these elements, we analyzed eight copies of a short sequence of the element from several species of the obscura group. The obtained phylogram agrees with the phylogeny of the species, which suggests vertical transmission of the element. PMID- 11264412 TI - Phylogeny, function, and evolution of the cupins, a structurally conserved, functionally diverse superfamily of proteins. AB - The cupin superfamily is a group of functionally diverse proteins that are found in all three kingdoms of life, Archaea, Eubacteria, and Eukaryota. These proteins have a characteristic signature domain comprising two histidine- containing motifs separated by an intermotif region of variable length. This domain consists of six beta strands within a conserved beta barrel structure. Most cupins, such as microbial phosphomannose isomerases (PMIs), AraC- type transcriptional regulators, and cereal oxalate oxidases (OXOs), contain only a single domain, whereas others, such as seed storage proteins and oxalate decarboxylases (OXDCs), are bi-cupins with two pairs of motifs. Although some cupins have known functions and have been characterized at the biochemical level, the majority are known only from gene cloning or sequencing projects. In this study, phylogenetic analyses were conducted on the conserved domain to investigate the evolution and structure/function relationships of cupins, with an emphasis on single- domain plant germin-like proteins (GLPs). An unrooted phylogeny of cupins from a wide spectrum of evolutionary lineages identified three main clusters, microbial PMIs, OXDCs, and plant GLPs. The sister group to the plant GLPs in the global analysis was then used to root a phylogeny of all available plant GLPs. The resulting phylogeny contained three main clades, classifying the GLPs into distinct subfamilies. It is suggested that these subfamilies correlate with functional categories, one of which contains the bifunctional barley germin that has both OXO and superoxide dismutase (SOD) activity. It is proposed that GLPs function primarily as SODs, enzymes that protect plants from the effects of oxidative stress. Closer inspection of the DNA sequence encoding the intermotif region in plant GLPs showed global conservation of thymine in the second codon position, a character associated with hydrophobic residues. Since many of these proteins are multimeric and enzymatically inactive in their monomeric state, this conservation of hydrophobicity is thought to be associated with the need to maintain the various monomer- monomer interactions. The type of structure-based predictive analysis presented in this paper is an important approach for understanding gene function and evolution in an era when genomes from a wide range of organisms are being sequenced at a rapid rate. PMID- 11264413 TI - Local changes in GC/AT substitution biases and in crossover frequencies on Drosophila chromosomes. AB - I present here evidence of remarkable local changes in GC/AT substitution biases and in crossover frequencies on Drosophila chromosomes. The substitution pattern at 10 loci in the telomeric region of the X chromosome was studied for four species of the Drosophila melanogaster species subgroup. Drosophila orena and Drosophila erecta are clearly the most closely related species pair (the erecta complex) among the four species studied; however, the overall data at the 10 loci revealed a clear dichotomy in the silent substitution patterns between the AT biased- substitution melanogaster and erecta lineages and the GC-biased substitution yakuba and orena lineages, suggesting two or more independent changes in GC/AT substitution biases. More importantly, the results indicated a between- loci heterogeneity in GC/AT substitution bias in this small region independently in the yakuba and orena lineages. Indeed, silent substitutions in the orena lineage were significantly biased toward G and C at the consecutive yellow, lethal of scute, and asense loci, but they were significantly biased toward A and T at sta. The substitution bias toward G and C was centered in different areas in yakuba (significantly biased at EG:165H7.3, EG:171D11.2, and suppressor of sable). The similar silent substitution patterns in coding and noncoding regions, furthermore, suggested mutational biases as a cause of the substitution biases. On the other hand, previous study reveals that Drosophila yakuba has about 20-fold higher crossover frequencies in the telomeric region of the X chromosome than does D. melanogaster; this study revealed that the total genetic map length of the yakuba X chromosome was only about 1.5 times as large as that of melanogaster and that the map length of the X-telomeric y-sta region did not differ between Drosophila yakuba and D. erecta. Taken together, the data strongly suggested that an approximately 20- fold reduction in the X-telomeric crossover frequencies occurred in the ancestral population of D. melanogaster after the melanogaster-yakuba divergence but before the melanogaster-simulans divergence. PMID- 11264414 TI - Estimating mutation rate and generation time from longitudinal samples of DNA sequences. AB - We present in this paper a simple method for estimating the mutation rate per site per year which also yields an estimate of the length of a generation when mutation rate per site per generation is known. The estimator, which takes advantage of DNA polymorphisms in longitudinal samples, is unbiased under a number of population models, including population structure and variable population size over time. We apply the new method to a longitudinal sample of DNA sequences of the env gene of human immunodeficiency virus type 1 (HIV-1) from a single patient and obtain 1.62 x 10(-2) as the mutation rate per site per year for HIV-1. Using an independent data set to estimate the mutation rate per generation, we obtain 1.8 days as the length of a generation of HIV-1, which agrees well with recent estimates based on viral load data. Our estimate of generation time differs considerably from a recent estimate by Rodrigo et al. when the same mutation rate per site per generation is used. Some factors that may contribute to the difference among different estimators are discussed. PMID- 11264415 TI - Molecular evolution of the teosinte branched gene among maize and related grasses. AB - Several authors have proposed that changes in a small number of regulatory genes may be sufficient for the evolution of novel morphologies. Recent analyses have indicated that teosinte branched1 (tb1), a putative bHLH transcription factor, played such a role during the morphological evolution of maize from its wild ancestor, teosinte. To address whether or not tb1 played a similar role during the evolution of the Andropogoneae, the tribe to which maize belongs, and to examine the rate and pattern of tb1 evolution within this tribe, we analyzed tb1 like sequences from 23 members of the Andropogoneae and five other grasses. Our analysis revealed that the TB1 protein evolves slowly within three conserved domains but rapidly outside these domains. The nonconserved regions of the gene are characterized by both a high nonsynonymous substitution rate and frequent indels. The ratio of nonsynonymous substitutions per nonsynonymous site (d(N)) to synonymous substitutions per synonymous site (d(S)) was not significantly greater than 1.0, providing no evidence for positive selection. However, the d(N)/d(S) ratio varied significantly among lineages and was high compared with those of other plant nuclear genes. Variation in the d(N)/d(S) ratio among the Andropogoneae could be explained by unequal levels of purifying selection among lineages. Consistent with this interpretation, the rate of nonsynonymous substitution differed along several lineages, while the synonymous substitution rate did not differ significantly. Finally, using tb1, we examined phylogenetic relationships within the Andropogoneae. The phylogeny suggests that the tribe underwent a rapid radiation during its early history and that the monoecious Andropogoneae are polyphyletic. PMID- 11264416 TI - Molecular evolution of transferrin: evidence for positive selection in salmonids. AB - Transferrins are iron-binding proteins that are involved in iron storage and resistance to bacterial disease. Previous work has shown that nonsynonymous-to synonymous-site substitution ratios (d(n)/d(s) ratios) between transferrin genes from some salmonid species were significantly greater than 1.0, providing evidence for positive selection at the transferrin gene. The purpose of the current study was to put these earlier results in a broader evolutionary context by examining variation among 25 previously published transferrin sequences from fish, amphibians, and mammals. The results of the study show that evidence for positive selection at transferrin is limited to salmonids-d(n)/d(s) ratios estimated for nonsalmonid lineages were generally less than 1.0. Within the salmonids, approximately 13% of the transferrin codon sites are estimated to be subject to positive selection, with an estimated d(n)/d(s) ratio of approximately 7. The three- dimensional locations of some of the selected sites were inferred by comparing these sites to homologous sites in the bovine lactoferrin crystallographic structure. The selected sites generally fall on the outside of the molecule, within and near areas that are bound by transferrin-binding proteins from human pathogenic bacteria. The physical locations of sites estimated to be subject to positive selection support previous speculation that competition for iron from pathogenic bacteria could be the source of positive selection. PMID- 11264417 TI - Extreme length and length variation in the first ribosomal internal transcribed spacer of ladybird beetles (Coleoptera: Coccinellidae). AB - DNA sequences of the first ribosomal internal transcribed spacer (ITS1) were isolated from 10 ladybird beetle species (Coleoptera: Coccinellidae) representing four subfamilies (Coccinellinae, Chilocorinae, Scymninae, and Coccidulinae). The spacers ranged in length from 791 to 2,572 bp, thereby including one of the longest ITS1s and exhibiting one of the most extreme cases of ITS1 size variation in eukaryotes recorded to date. The causes of length variation were therefore analyzed. Almost no putatively homologous sequence similarities were identified for the taxa included. The only exception was for the subfamily Coccinellinae, which yielded sequence similarities in six regions of approximately 550 nucleotide positions, primarily at the 5' and 3' ends of ITS1. The majority of differences in ITS1 length between taxa could be attributed to the presence of repetitive elements with comparatively long repeat units. Repetition arose several times independently and was confined to the middle of the spacer which, in contrast to the 5' and 3' ends, had not been inferred in previous studies to be subject to functional constraints. These elements were characterized by high rates of evolutionary change, most likely as a result of high substitution rates in combination with inefficient homogenization across repeats. The repeated origin and subsequent divergence of "long" repetitive elements should thus be assumed to be an important factor in the evolution of coccinellid ITS1. PMID- 11264418 TI - Dating the origin of the African human T-cell lymphotropic virus type-i (HTLV-I) subtypes. AB - To investigate the origin of the African PTLV-I virus, we phylogenetically analyzed the available HTLV-I and STLV-I strains. We also attempted to date the presumed interspecies transmissions that resulted in the African HTLV-I subtypes. Molecular-clock analysis was performed using the Tamura-Nei substitution model and gamma distributed rate heterogeneity based on the maximum-likelihood topology of the combined long-terminal-repeat and env third-codon-position sequences. Since the molecular clock was not rejected and no evidence for saturation was found, a constant rate of evolution at these positions for all 33 HTLV-I and STLV I strains was reasonably assumed. The spread of PTLV-I in Africa is estimated to have occurred at least 27,300 +/- 8,200 years ago. Using the available strains, the HTLV-If subtype appears to have emerged within the last 3,000 years, and the HTLV-Ia, HTLV-Ib, HTLV-Id, and HTLV-Ie subtypes appear to have diverged between 21,100 and 5,300 years ago. Interspecies transmissions, most probably simian to human, must have occurred around that time and probably continued later. When the synonymous and nonsynonymous substitution ratios were compared, it was clear that purifying selection was the driving force for PTLV-I evolution in the env gene, irrespective of the host species. Due to the small number of strains in some of the investigated groups, these data on selective pressure should be taken with caution. PMID- 11264420 TI - Uninformative characters and apparent conflict between molecules and morphology. PMID- 11264419 TI - Inferring admixture proportions from molecular data: extension to any number of parental populations. AB - The relative contribution of two parental populations to a hybrid group (the admixture proportions) can be estimated using not only the frequencies of different alleles, but also the degree of molecular divergence between them. In this paper, we extend this possibility to the case of any number of parental populations. The newly derived multiparental estimator is tested by Monte Carlo simulations and by generating artificial hybrid groups by pooling mtDNA samples from human populations. The general properties (including the variance) of the two-parental estimator seem to be retained by the multiparental estimator. When mixed human populations are considered and hypervariable single-locus data are analyzed (mtDNA control region), errors in the estimated contributions appear reasonably low only when highly differentiated parental populations are involved. Finally, the method applied to the hybrid Canary Island population points to a much lower female contribution from Spain than has previously been estimated. PMID- 11264421 TI - Evolutionary rates of duplicate genes in fish and mammals. PMID- 11264422 TI - Implications for bat evolution from two new complete mitochondrial genomes. PMID- 11264423 TI - The limits of viability. PMID- 11264424 TI - Nothing endures but change. PMID- 11264425 TI - Is periventricular leukomalacia an axonopathy as well as an oligopathy? AB - Periventricular leukomalacia is a white matter disorder, the neonatal cranial ultrasound images of which predict long-term developmental limitations among preterm infants. The vulnerability of oligodendrocytes has led to the hypothesis that oligodendrocytes suffer the primary damage, with axonal damage occurring as a consequence. In this article, we discuss the differential role of oligodendrocytes and axons in this disorder's etiology, offering analogies from the multiple sclerosis and hydrocephalus literature. We conclude that it is too early to view periventricular leukomalacia exclusively as a consequence of oligodendrocyte damage and/or maldevelopment. PMID- 11264426 TI - Renal development and adult hypertension. PMID- 11264428 TI - Food deprivation limits insulin secretory capacity in postpubertal rats. AB - ABSTRACT Because prenatal and perinatal undernutrition are associated with type 2 diabetes later in life, we posed the question whether nutrient deprivation during puberty would also result in a decreased ability to secrete insulin. Chronically catheterized, unstressed Sprague Dawley rats, fed ad libitum, were studied before puberty (Pre, n = 14) and after puberty (Post, n = 8). Moderately caloric restricted rats (fed 70% of the control diet, n = 9), were studied after puberty. Insulin secretion was assessed using a hyperglycemic clamp at a glucose concentration of 300 mg/dL, or with a primed continuous infusion of intralipid (plasma FFA levels approximately 1.5 mM) at a plasma glucose concentration of 200 mg/dL. Stimulated insulin levels increased in Post rats by 3- to 4-fold compared with Pre rats (from 4.6 +/- 0.4 ng/mL Pre to 12.8 +/- 0.7 ng/mL Post, and from 4.5 +/- 0.4 ng/mL Pre to 15.8 +/- 0.7 ng/mL Post, respectively, p < 0.001, at a glucose concentration of 300 mg/dL, and 200 mg/dL with intralipid). Caloric restriction prevented any rise in insulin secretion (3.8 +/- 0.5 and 4.6 +/- 0.5 ng/mL in the caloric-restricted rats at glucose concentrations of 300 mg/dL and 200 mg/dL with intralipid, respectively). A semiquantitative reverse transcriptase PCR procedure was used to assess basal and stimulated insulin mRNA levels. Caloric restriction did not compensate by enhancing insulin mRNA levels in response to glucose stimulation. Moderate food deprivation during puberty reduced the capacity of the pancreas to secrete insulin in response to different nutrient stimuli. We hypothesize that puberty has an important role in beta-cell maturation and any major nutrient modification may have deleterious consequences later in life. PMID- 11264427 TI - Maternal protein restriction suppresses the newborn renin-angiotensin system and programs adult hypertension in rats. AB - Restriction of maternal protein intake during rat pregnancy produces offspring that are hypertensive in adulthood, but the mechanisms are not well understood. Our purpose was to determine whether this adult hypertension could be programmed during development by suppression of the fetal/newborn renin-angiotensin system (RAS) and a consequent reduction in nephron number. Pregnant rats were fed a normal protein (19%, NP) or low-protein (8.5%, LP) diet throughout gestation. Birth weight was reduced by 13% (p < 0.0005), and the kidney/body weight ratio was reduced in LP pups. Renal renin mRNA levels were significantly reduced in newborn LP pups; renal renin concentration and renin immunostaining were suppressed. Renal tissue angiotensin II levels were also suppressed in newborn LP (0.079 +/- 0.002 ng/mg, LP versus 0.146 +/- 0.016 ng/mg, NP, p < 0.01). Mean arterial pressure in conscious, chronically instrumented adult offspring (21 wk) was higher in LP (135 +/- 1 mm Hg, LP versus 126 +/- 1 mm Hg, NP, p < 0.00007), and GFR normalized to kidney weight was reduced in LP (p < 0.04). The number of glomeruli per kidney was lower in adult LP offspring (21,567 +/- 1,694, LP versus 28,917 +/- 2,342, NP, p < 0.03), and individual glomerular volume was higher (1.81 +/- 0.16 10(6) microm(3), LP versus 1.11 +/- 0.10 10(6) microm(3), NP, p < 0.005); the total volume of all glomeruli per kidney was not significantly different. Thus, perinatal protein restriction in the rat suppresses the newborn intrarenal RAS and leads to a reduced number of glomeruli, glomerular enlargement, and hypertension in the adult. PMID- 11264429 TI - Intrauterine growth restriction induces up-regulation of cerebral aromatic amino acid decarboxylase activity in newborn piglets: [18F]fluorodopa positron emission tomographis study. AB - There is evidence that intrauterine growth restriction (IUGR) is associated with altered dopaminergic function in the immature brain. However, the relevant enzyme activities have not been measured in the living neonatal brain together with brain oxidative metabolism. Therefore, fluorine-18-labeled 6-fluoro-L-3,4 dihydroxyphenylalanine (FDOPA) was used together with positron emission tomography to estimate the activity of the aromatic amino acid decarboxylase in the brain of 10 newborn IUGR piglets (2 to 5 d old; body weight, 908 +/- 109 g) and in 10 normal-weight (3 to 5 d old; body weight, 2142 +/- 373 g) newborn piglets. The regional transport of FDOPA to the brain and the clearance rate of labeled metabolites from brain tissue were broadly similar in the two groups. However, the regional rate constant for back flux from the brain was markedly increased in IUGR piglets for striatum (72%) and frontal cortex (83%) (p < 0.05). Furthermore, the rate constant for conversion of FDOPA to fluorodopamine was markedly increased (between 48% in cerebellum and 91% in mesencephalon, p < 0.05) in all brain regions of IUGR piglets studied. Thus, it is suggested that IUGR induces an up-regulation of aromatic amino acid decarboxylase activity that is not related to alterations in brain oxidative metabolism. PMID- 11264430 TI - Markers of type I and type III collagen turnover, insulin-like growth factors, and their binding proteins in cord plasma of small premature infants: relationships with fetal growth, gestational age, preeclampsia, and antenatal glucocorticoid treatment. AB - Disorders affecting fetal growth are commonly associated with premature birth. IGFs and their binding proteins (IGFBPs) are potent regulators of fetal growth. In vitro evidence suggests that they regulate collagen turnover. Collagen turnover can be monitored by serum markers of type I collagen synthesis (PINP) and degradation (ICTP) and a marker of type III collagen synthesis (PIIINP). We examined whether these markers in fetal circulation reflect intrauterine growth and maturity, and whether any interrelationship exists between them and fetal IGFs and IGFBPs in preterm infants before 32 wk of gestation. Cord plasma PINP, ICTP, PIIINP, IGF-I, IGF-II, IGFBP-1, and IGFBP-3 were determined for 98 preterm infants. To express birth weight in units adjusted for gestational age, a birth weight SD score (SDS) was calculated. Negative correlations existed between gestational age and PINP (r = -0.43; p < 0.0001), ICTP (r = -0.34; p = 0.002), and PIIINP (r = -0.34; p = 0.0001). Positive correlations existed between birth weight SDS and PINP (r = 0.40; p = 0.0002) and ICTP (r = 0.48; p < 0.0001) but not PIIINP. Moreover, birth weight SDS was positively correlated with IGF-I (r = 0.58; p < 0.0001) and IGFBP-3 (r = 0.44; p < 0.0001) and negatively correlated with IGF-II (r = -0.36; p = 0.003) and IGFBP-1 (r = -0.50; p < 0.0001). Gestational age correlated with IGFBP-3 (r = 0.25; p = 0.03). In preeclampsia, IGF-I was lower (p = 0.002) and IGFBP-1 higher (p < 0.0001), also after adjustment for fetal size. The number of antenatal glucocorticoid treatments was associated with lower ICTP (p = 0.04), higher IGF-I (p = 0.002), lower IGF-II (p = 0.02), lower IGFBP-1 (p = 0.05), and higher IGFBP-3 (p = 0.004), also after adjustment for potential confounders. In multiple regression analysis, the factors significantly associated with PINP (R:(2) = 0.47) were gestational age and IGF-I, and those associated with ICTP (R:(2) = 0.54) were IGF-I, gestational age, and antenatal glucocorticoid treatment. We conclude that IGF-I may be involved in regulation of type I collagen turnover in the growing fetus. Cord blood PINP and ICTP reflect both fetal growth and maturity and deserve evaluation as potential indicators of postnatal growth velocity in preterm infants, whereas PIIINP reflects fetal maturity. PMID- 11264431 TI - Effects of altitude versus economic status on birth weight and body shape at birth. AB - The compelling evidence linking small size at birth with later cardiovascular disease has renewed and amplified a clinical and scientific interest in the determinants of fetal growth. Although the effects of maternal nutrition on fetal growth have been extensively studied, comparatively little is known about the effects of maternofetal hypoxia. This study tested the hypothesis that in highland regions, high altitude rather than maternal economic status is associated with reduced and altered fetal growth by investigating the effects of high altitude versus economic status on birth weight and body shape at birth in Bolivia. Bolivia is geographically and socioeconomically unique. It contains several highland (>3500 m above sea level) and lowland (<500 m) cities that are inhabited by very economically divergent populations. Birth weight, body length, and head circumference were compared between a high- (n = 100) and low- (n = 100) income region of La Paz (3649 m; largest high-altitude city) and a high- (n = 100) and low- (n = 100) income region of Santa Cruz (437 m; largest low-altitude city). In addition, the frequency distribution across the continuum of birth weights was plotted for babies born from high- and low-income families in La Paz and Santa Cruz. Mean birth weights were lower in babies from La Paz than in babies from Santa Cruz in both high- and low-income groups. The cumulative frequency curve across all compiled birth weights was shifted to the left in babies from La Paz compared with those from Santa Cruz, regardless of economic status. The frequency of low birth weight (<2500 g) was higher in babies from La Paz than from Santa Cruz in both high- and low-income groups. In addition, at high altitude but not at low altitude, high income was associated with an increase in the head circumference:birth weight ratio. These findings suggest that high altitude rather than economic status is associated with low birth weight and altered body shape at birth in babies from Bolivia. PMID- 11264432 TI - Cerebral glucose metabolism measured by positron emission tomography in term newborn infants with hypoxic ischemic encephalopathy. AB - Total and regional cerebral glucose metabolism (CMRgl) was measured by positron emission tomography with 2-((18)F) fluoro-2-deoxy-D-glucose ((18)FDG) in 20 term infants with hypoxic ischemic encephalopathy (HIE) after perinatal asphyxia. All infants had signs of perinatal distress, and 15 were severely acidotic at birth. Six infants developed mild HIE, twelve moderate HIE, and two severe HIE during their first days of life. The positron emission tomographic scans were performed at 4-24 d of age (median, 11 d). One hour before scanning, 2-3.7 MBq/kg (54-100 microCi/kg) (18)FDG was injected i.v. No sedation was used. Quantification of CMRgl was based on a new method employing the glucose metabolism of the erythrocytes, requiring only one blood sample. In all infants, the most metabolically active brain areas were the deep subcortical parts, thalamus, basal ganglia, and sensorimotor cortex. Frontal, temporal, and parietal cortex were less metabolically active in all infants. Total CMRgl was inversely correlated with the severity of HIE (p < 0.01). Six infants with mild HIE had a mean (range) CMRgl of 55.5 (37.7-100.8) micromol.min(-1).100 g(-1), 11 with moderate HIE had 26.6 (13.0-65.1) micromol.min(-1).100 g(-1), and two with severe HIE had 10.4 and 15.0 micromol.min(-1).100 g(-1), respectively. Five of six infants who developed cerebral palsy had a mean (range) CMRgl of 18.1 (10.2-31.4) micromol.min(-1).100 g(-1) compared with 41.5 (13.0-100.8) micromol.min(-1).100 g(-1) in the infants with no neurologic sequela at 2 y. We conclude that CMRgl measured during the subacute period after perinatal asphyxia in term infants is highly correlated with the severity of HIE and short-term outcome. PMID- 11264434 TI - Bilirubin exerts additional toxic effects in hypoxic cultured neurons from the developing rat brain by the recruitment of glutamate neurotoxicity. AB - Both hypoxia and bilirubin are common risk factors in newborns, which may act synergistically to produce anatomical and functional disturbances of the CNS. Using primary cultures of neurons from the fetal rat brain, it was recently reported that neuronal apoptosis accounts for the deleterious consequences of these two insults. To investigate the influence of hypoxia, bilirubin, or their combination on the outcome of neuronal cells of the immature brain, and delineate cellular mechanisms involved, 6-d-old cultured neurons were submitted to either hypoxia (6 h), unconjugated bilirubin (0.5 microM), or to combined conditions. Within 96 h, cell viability was reduced by 22.7% and 24.5% by hypoxia and bilirubin, respectively, whereas combined treatments decreased vital score by 34%. Nuclear morphology revealed 13.4% of apoptotic cells after hypoxia, 16.2% after bilirubin, and 22.6% after both treatments. Bilirubin action was specifically blocked by the glutamate receptor antagonist MK-801, which was without effect on the consequences of hypoxia. Temporal changes in [(3)H]leucine incorporation rates as well as beneficial effects of cycloheximide reflected a programmed phenomenon dependent upon synthesis of selective proteins. The presence of bilirubin reduced hypoxia-induced alterations of cell energy metabolism, as reflected by 2-D-[(3)H]deoxyglucose incorporation, raising the question of free radical scavenging. Measurements of intracellular radical generation, however, failed to confirm the antioxidant role of bilirubin. Taken together, our data suggest that low levels of bilirubin may enhance hypoxia effects in immature neurons by facilitating glutamate-mediated apoptosis through the activation of N:-methyl-D-aspartate receptors. PMID- 11264433 TI - Early biochemical indicators of hypoxic-ischemic encephalopathy after birth asphyxia. AB - Hypoxic-ischemic encephalopathy (HIE) after perinatal asphyxia is a condition in which serum concentrations of brain-specific biochemical markers may be elevated. Neuroprotective interventions in asphyxiated newborns require early indicators of brain damage to initiate therapy. We examined brain-specific creatine kinase (CK BB), protein S-100, and neuron-specific enolase in cord blood and 2, 6, 12, and 24 h after birth in 29 asphyxiated and 20 control infants. At 2 h after birth, median (quartiles) serum CK-BB concentration was 10.0 U/L (6.0-13.0 U/L) in control infants, 16.0 U/L (13.0-23.5 U/L) in infants with no or mild HIE, and 46.5 U/L (21.4-83.0 U/L) in infants with moderate or severe HIE. Serum protein S 100 was 1.6 microg/L (1.4-2.5 microg/L) in control infants, 2.9 microg/L (1.8-4.7 microg/L) in asphyxiated infants with no or mild HIE, and 17.0 microg/L (3.2-34.1 microg/L) in infants with moderate or severe HIE 2 h after birth. No significant difference was detectable in serum neuron-specific enolase between infants with no or mild and moderate or severe HIE 2 and 6 h after birth. A combination of serum protein S-100 (cutoff value, 8.5 microg/L) and CK-BB (cutoff value, 18.8 U/L) 2 h after birth had the highest predictive value (83%) and specificity (95%) of predicting moderate and severe HIE. Cord blood pH (cutoff value, <6.9) and cord blood base deficit (cutoff value, >17 mM) increase the predictive values of protein S-100 and CK-BB. We conclude that elevated serum concentrations of protein S-100 and CK-BB reliably indicate moderate and severe HIE as early as 2 h after birth. PMID- 11264435 TI - Variations in the promoter region of the apolipoprotein A-1 gene influence plasma lipoprotein(a) levels in Asian Indian neonates from Singapore. AB - We studied the influence of two DNA polymorphisms (-75 bp G/A and +83 bp C/T) in the promoter region of the apolipoprotein A-1 (apoA1) gene on cord plasma level of lipoprotein(a) [Lp(a)] in 1076 newborns of both genders from the three major ethnic groups in Singapore-Chinese, Malays, and Asian Indians. The frequency of the A: allele at -75 bp in the Indians was significantly lower than the Chinese and Malays. There was no linkage disequilibrium between the two sites studied. Both polymorphic sites were not significantly associated with any lipid factors except for Lp(a) levels in the Asian Indians. The AA and CC homozygotes were significantly associated with lower Lp(a) levels. These associations were specific only to the male Indian neonates. The genetic variations at the -75 and +83 bp explained 6.9% and 7.2%, respectively, of the total variability of plasma Lp(a) levels at birth in the Asian Indians. The Lp(a) levels were also significantly different between composite genotypes in the order GG/TT > GA/CT > GG/CT > GA/CC > GG/CC > AA/CC. The effects of the two polymorphisms seem to be additive as the composite genotypes were able to explain 14% of the Lp(a) variance, equivalent to the sum of the two constituent sites. Our results showed that there is significant ethnic- and gender-specific influence of the apoA1 gene on plasma Lp(a) levels at birth that is inherent and independent of known gene environment interactions. PMID- 11264436 TI - Role of oxidant stress in the permeability transition induced in rat hepatic mitochondria by hydrophobic bile acids. AB - Hydrophobic bile acids may cause hepatocellular necrosis and apoptosis during cholestatic liver diseases. The mechanism for this injury may involve mitochondrial dysfunction and the generation of oxidant stress. The purpose of this study was to determine the relationship of oxidant stress and the mitochondrial membrane permeability transition (MMPT) in hepatocyte necrosis induced by bile acids. The MMPT was measured spectrophotometrically and morphologically in rat liver mitochondria exposed to glycochenodeoxycholic acid (GCDC). Freshly isolated rat hepatocytes were exposed to GCDC and hepatocellular necrosis was assessed by lactate dehydrogenase release, hydroperoxide generation by dichlorofluorescein fluorescence, and the MMPT in cells by JC1 and tetramethylrhodamine methylester fluorescence on flow cytometry. GCDC induced the MMPT in a dose- and Ca(2+)-dependent manner. Antioxidants significantly inhibited the GCDC-induced MMPT and the generation of hydroperoxides in isolated mitochondria. Other detergents failed to induce the MMPT and a calpain-like protease inhibitor had no effect on the GCDC-induced MMPT. In isolated rat hepatocytes, GCDC induced the MMPT, which was inhibited by antioxidants. Blocking the MMPT in hepatocytes reduced hepatocyte necrosis and oxidant stress caused by GCDC. Oxidant stress, and not detergent effects or the stimulation of calpain like proteases, mediates the GCDC-induced MMPT in hepatocytes. We propose that reducing mitochondrial generation of reactive oxygen species or preventing increases in mitochondrial Ca(2+) may protect the hepatocyte against bile acid induced necrosis. PMID- 11264437 TI - Normal clinical outcome in untreated subjects with mild hyperphenylalaninemia. AB - ABSTRACT There is international consensus that patients with phenylalanine (Phe) levels <360 microM on a free diet do not need Phe-lowering dietary treatment whereas patients with levels >600 microM do. Clinical outcome of patients showing Phe levels between 360 and 600 microM in serum on a free nutrition has so far only been assessed in a small number of cases. Therefore, different recommendations exist for patients with mild hyperphenylalaninemia. We investigated in a nationwide study 31 adolescent and adult patients who persistently displayed serum Phe levels between 360 and 600 microM on a normal nutrition with a corresponding genotype. Because of limited accuracy of measurements, Phe levels should be looked on as an approximation, but not as an absolute limit in every instance. In addition to serum Phe levels, the assessment program consisted of comprehensive psychological testing, magnetic resonance imaging of the head, (1)H magnetic resonance spectroscopy, and genotyping. We found a normal intellectual (intelligence quotient, 103 +/- 15; range, 79-138) and educational (school performance and job career) outcome in these subjects as compared with healthy control subjects (intelligence quotient, 104 +/- 11; range, 80-135). Magnetic resonance imaging revealed no changes of cerebral white matter in any patient, and (1)H magnetic resonance spectroscopy revealed brain Phe levels below the limit of detection (<200 microM). In the absence of any demonstrable effect, dietary treatment is unlikely to be of value in patients with mild hyperphenylalaninemia and serum Phe levels <600 microM on a free nutrition, and should no longer be recommended. Because of a possible late-onset phenylketonuria, Phe levels of untreated patients should be monitored carefully at least during the first year of life. Nevertheless, problems of maternal phenylketonuria should still be taken into account. PMID- 11264438 TI - Impairment of the Golgi GDP-L-fucose transport and unresponsiveness to fucose replacement therapy in LAD II patients. AB - Leukocyte adhesion deficiency type II is an autosomal recessive syndrome characterized by generalized reduction of L-fucose in glycoconjugates; the specific molecular defect is still undefined. The most important clinical symptoms include severe growth and mental retardation and severe immunodeficiency. Patients from two ethnic groups have been reported, i.e. Arab and Turkish. We have observed that GDP-L-fucose transport into Golgi vesicles was specifically impaired in an Arab patient, with a significant reduction of the V:(max) but no significant differences in the K:(m) from control and parents. GDP L-fucose transport showed simple saturation kinetics in all samples. Transport of UDP-galactose, UDP-N:-acetylglucosamine, and CMP-sialic acid was comparable into vesicles from the Arab patient, parents, and control. These kinetic parameters probably account for the failure to obtain any clinical and biochemical response to fucose therapy in Arab patients. This contrasts both with the distinctive kinetic properties of GDP-L-fucose transport and with the success of fucose therapy, which have been recently reported in one patient of Turkish origin. Accordingly, the biochemical properties of GDP-L-fucose transport into the Golgi are consistent with different variants of leukocyte adhesion deficiency type II that are probably the result of different molecular defects. PMID- 11264439 TI - Differential appearance of T cell subsets in the large and small intestine of neonatal mice. AB - We examined the appearance of intestinal intraepithelial lymphocytes (IEL) during the first 12 wk of life to gain insight into postnatal factors that contribute to the differences found between IEL in the large and small intestines of adult mice. Intestinal T cells were very infrequent at birth, but increased in number in the large and small intestine during the first 4 wk of life and then stabilized. The small intestinal epithelium at 2 wk of age contained mostly T cell receptor (TCR) alphabeta+, CD2+ T cells, unlike IEL in adult mice, which were composed of nearly equal proportions of CD2-, TCR alphabeta+ and TCR gammadelta+ cells. Between 2 and 3 wk of age, TCR gammadelta+, CD2- IEL increased greatly in the small intestine, whereas TCR alphabeta+ cells expressing CD2 decreased. By contrast, IEL in the large intestine at 2 and 3 wk of age were mostly TCR alphabeta+, CD2+ T cells similar to large intestinal IEL in adult mice. And finally, the expression of CD69 increased earlier and to higher levels on TCR alphabeta+ and TCR gammadelta+ IEL in the small intestine than in the large intestine. Our results demonstrate that IEL in the large and small intestine are phenotypically similar during suckling and that differences between these populations are established after weaning. Furthermore, the earlier accumulation of IEL with an activated adult IEL phenotype in the small intestine suggests that these T cells mature or expand in the gut and contribute to the maturation of immune function during postnatal life in mice. PMID- 11264440 TI - TNFalpha decreases gluconeogenesis in hepatocytes isolated from 10-day-old rats. AB - Gluconeogenesis decreases during septic shock, but its mechanism is not well known. Tumor necrosis factor alpha (TNF-alpha), which is a key cytokine in septic shock, can increase GLUT1 gene expression and glucose uptake in muscles and fatty tissues. TNF-alpha does not alter the metabolism of hepatocytes in which GLUT2 is the predominant glucose transporter. However, GLUT1 is the predominant glucose transporter in hepatocytes of 10-d-old rats. Thus, we hypothesized that TNF-alpha might increase glucose uptake and glycolysis in those cells, and decrease gluconeogenesis. In the present study, hepatocytes isolated from 10-d-old rats were incubated with TNF-alpha at the concentrations of 0, 0.98, 9.8, 98, and 980 ng/mL to evaluate TNF-alpha effects on gluconeogenesis and glucose uptake. TNF alpha increased glucose uptake (41.1 +/- 8 to 114 +/- 21.4 micromol/10(6) cells at the concentration of 980 ng/mL of TNF-alpha) in a dose-dependent manner, and decreased gluconeogenesis (98.2 +/- 8.2 to 1.1 +/- 3.2 micromol/10(6) cells at the concentration of 980 ng/mL of TNF-alpha) in a dose-dependent manner. The decrease of glucokinase mRNA and GLUT1 mRNA abundance correlated with glucose uptake (r = 0.988 and 0.997, respectively), and the decrease of phosphoenolpyruvate carboxykinase mRNA abundance correlated with the decrease of gluconeogenesis (r = 0.972). The decrease of gluconeogenesis by TNF-alpha correlated with the increase of glucose uptake (r = -0.988). We concluded that TNF-alpha reciprocally suppressed gluconeogenesis in hepatocytes isolated from 10 d-old rats. PMID- 11264441 TI - Heart rate response to transient chemoreceptor stimulation in term infants is modified by exposure to maternal smoking. AB - Modulation of heart rate (HR) during transient hyperoxia, hypoxia, and hypercapnia was studied in 46 healthy term infants on 103 occasions (postnatal d 2 to 82). Twenty-three infants had smoking mothers (median, 11 cigarettes/d). Transient chemoreceptor stimuli (100% O(2), 15% O(2), or 3% CO(2)) were presented repeatedly during quiet sleep. Beat-by-beat HR and breath-by-breath ventilation were recorded continuously. The coherently averaged HR and ventilation responses to each stimulus were calculated for each infant at each age. Outcome variables (HR change from baseline to end of stimulation, maximum HR change, and time to half-maximum) were analyzed by ANOVA. Overall, HR declined during hyperoxia (median change, 4.2 beats/min) and rose during hypoxia (median change, 4.2 beats/min) and hypercapnia (median change, 4.6 beats/min). The percentage change in HR was positively correlated with the percentage change in ventilation (p < 0.001). Increasing number of cigarettes smoked by the mother was correlated with deeper HR declines and smaller HR rises (p = 0.02). For the population as a whole, the HR response lagged 3.8 s behind the ventilatory response during hyperoxia and hypoxia (p < 0.001), whereas during hypercapnia there was no significant lag. The lag in HR response in the smoke-exposed group was 2.5 s greater than that in the control group for all three stimuli (p = 0.001), and the difference increased with the number of cigarettes smoked by the mother (p < 0.01). Both pulmonary reflexes and the type of the chemoreceptor stimulus seemed to influence HR. Maternal smoking affected the magnitude and time-course of the HR response in a dose-dependent manner. PMID- 11264442 TI - Antenatal betamethasone administration decreases the lung hyaluronan concentration in preterm rabbit pups. AB - Maternal treatment with corticosteroids before preterm delivery is effective in reducing the incidence of respiratory distress syndrome and neonatal mortality. We hypothesized that corticosteroids might lower the lung hyaluronan concentration. Twenty-five rabbit dams (term = 31 d) with timed pregnancies were injected s.c. with 0.75 mg of betamethasone or saline (controls) 1 d before delivery. In addition, two dams delivered at 25 d of gestation were injected with 0.75 mg of betamethasone on two consecutive days before delivery. A total of 238 live pups were delivered by preterm cesarean section at 25, 27, 28, or 29 d of gestation and killed immediately. Their lung hyaluronan concentrations were measured with a radiometric assay, and wet/dry lung weight ratios were determined. Lungs of rabbit pups exposed antenatally to betamethasone and delivered at 25 or 27 d of gestation, but not at 28 or 29 d, displayed significantly (p = 0.001 and p = 0.008, respectively) lower hyaluronan concentrations than control pups, accompanied by less intense subepithelial staining for hyaluronan in alveolar walls. There was no significant difference in wet/dry lung weight ratio between pups exposed to one dose of betamethasone and controls. Antenatal corticosteroid exposure lowers the lung hyaluronan concentration in preterm rabbit pups delivered at 25 or 27 d of gestation, but not in those delivered at 28 or 29 d. PMID- 11264443 TI - Spontaneous breathing during partial liquid ventilation in animals with meconium aspiration. AB - Partial liquid ventilation (PLV) has been shown to improve gas exchange in paralyzed animals and humans with lung disease. The present study tests the hypothesis that PLV improves gas exchange in spontaneously breathing animals with meconium aspiration supported by proportional assist ventilation. Twenty-five adult anesthetized intubated rabbits with experimental meconium aspiration were randomized to gas ventilation (GV) or PLV while being supported by proportional assist ventilation. Minute ventilation, tidal volume, respiratory rate, mean airway pressure, heart rate, and mean arterial and pulmonary arterial pressure were recorded continuously. Every 30 min, arterial blood gases were obtained, and lung compliance, airway resistance, work of breathing, and cardiac output were measured. Animals were sacrificed after 5 h to obtain lung histology. More PLV animals survived until the end of the study period. PaO(2) (14.5 +/- 4.5 versus 25.6 +/- 6.7 kPa; p < 0.01; GV versus PLV) and lung compliance (4.3 +/- 0.4 versus 6.1 +/- 1.2 mL.kPa(-1).kg(-1); p < 0.001) were improved during PLV, resulting in a lower work of breathing (5.3 +/- 2.8 versus 3.5 +/- 1.5 mL.kPa.kg( 1); p < 0.05) and less need for ventilatory support. Minute ventilation and respiratory rate were higher during GV versus PLV, resulting in a slightly lower PaCO(2) (3.9 +/- 0.5 versus 4.5 +/- 0.7 kPa; p < 0.05). Histologic evaluation showed more atelectasis, inflammatory changes, and hemorrhage in GV animals. Other parameters measured were similar. We conclude that PLV improves oxygenation, lung compliance, and survival and results in less lung injury in spontaneously breathing animals with meconium aspiration when supported by proportional assist ventilation. PMID- 11264444 TI - Measurement of functional residual capacity in rabbits and children using an ultrasonic flow meter. AB - A sulfur hexafluoride (SF(6)) washin/washout technique was developed using an ultrasonic flowmeter to measure functional residual capacity (FRC) during mechanical ventilation. The ultrasonic flowmeter measures simultaneously flow and molar mass of the mainstream gas. Ventilation distribution was studied using moment ratios analysis (alveolar-based mean dilution number). Accuracy and precision of the measurement technique were tested in a mechanical lung model, and the method's sensitivity to changes of FRC was assessed in seven ventilated rabbits and six children. In the mechanical lung model with a volume range from 10 to 60 mL, the mean error of FRC measurement was 0.096 +/- 0.9 mL (range, 0-2 mL). In seven rabbits (mean body weight, 3.6 kg), measurements of FRC and alveolar-based mean dilution number were made at positive end-expiratory pressures (PEEP) of 0, 3, and 6 cm H(2)O. The mean coefficient of variation of 66 FRC-measurements was 5.5% (range, 0-15.3%). As the applied PEEP increased, mean FRC per kilogram body weight increased from 13.3 +/- 3.4 mL/kg (PEEP of 0 cm H(2)O) to 16.7 +/- 3.6 mL/kg (PEEP of 3 cm H(2)O) and to 20.8 +/- 4.3 mL/kg (PEEP of 6 cm H(2)O). Alveolar-based mean dilution number decreased accordingly from 1.94 +/- 0.42 (PEEP = 0; mean +/- SD), to 1.91 +/- 0.45 (PEEP = 3) and to 1.59 +/ 0.35 (PEEP = 6). In the six children, as applied PEEP increased, mean FRC per kilogram increased from 21.1 +/- 4.51 mL/kg (PEEP = 0), to 22.4 +/- 1.8 mL/kg (PEEP = 5) and 27.2 +/- 3.4 mL/kg (PEEP = 10). FRC measurement using the ultrasonic flowmeter is accurate and simple to use in ventilated and spontaneously breathing children. PMID- 11264445 TI - Evidence for an innate immune response in the immature human intestine: toll-like receptors on fetal enterocytes. AB - The intestinal epithelium is an active participant in the mucosal immune response against luminal pathogens. Microorganisms and their cell wall products, i.e. lipopolysaccharide (LPS), can stimulate the enterocyte to produce an innate immune response with the increased production of IL-8 via an activation of the transcription factor NFkappaB. The innate response mechanism, however, has not been understood until the recent description of a family of human toll-like receptors (hTLR) on immune cells that interact with LPS and modulate the IL-8 response via an intracellular signal transduction pathway similar to that of the IL-1 receptor family. Accordingly, in this study we have sought to determine the constitutive and regulated expression of hTLR on a nonmalignant human fetal primary small intestinal cell line (H4 cells) and on small intestinal samples of ileum from human fetuses (age 18-21 wk). Specimens were examined by reverse transcription PCR, Western blot analysis, and immunofluorescence for hTLR2 and hTLR4 mRNA and protein and to determine whether their expression was regulated by LPS or by an endogenous inflammatory stimulus, IL-1beta. hTLR2 and hTLR4 were expressed constitutively on H4 cells and on human fetal small intestinal enterocytes, predominantly on the basolateral surface of crypt enterocytes. Inflammatory stimuli appeared to regulate hTLR transcription (IL-1beta increased both hTLR2 and hTLR4 whereas LPS decreased hTLR4) and possibly translation (qualitative observations). The presence of hTLR on human fetal enterocyte suggests a mechanism for the innate immune response to pathogens and could provide the basis for further study of the accentuated inflammatory response in age-dependent gastrointestinal diseases such as necrotizing enterocolitis. PMID- 11264446 TI - Effective selective head cooling during posthypoxic hypothermia in newborn piglets. AB - Selective head cooling has been proposed as a neuroprotective intervention after hypoxia-ischemia in which the brain is cooled without subjecting the rest of the body to significant hypothermia, thus minimizing adverse systemic effects. There are little data showing it is possible to cool the brain more than the body. We have therefore applied selective head cooling to our hypoxia-ischemia piglet model to establish whether it is possible. Nine piglets were anesthetized, and brain temperature was measured at the surface and in the superficial (0.2 cm) and deep (1.7-2.0 cm) gray matter. Rectal (6-cm depth), skin, and scalp temperatures (T) were recorded continuously. Lowering T-rectal from normothermia (39 degrees C) to hypothermia (33.5-33.8 degrees C) using a head cap perfused with cold (6-24 degrees C) water was undertaken for up to 6 h. To assess the impact of the 45-min hypoxia-ischemia insult on the effectiveness of selective head cooling, four piglets were cooled both before and after the insult, and four, only afterward. During selective head cooling, it was possible to achieve a lower T-deep brain than T-rectal in all animals both before and after hypoxia. However, this was only possible when overhead body heating was used. The T-rectal to T-deep brain gradient was significantly smaller after the insult (median, 5.3 degrees C; range, 4.2-8.5 degrees C versus 3.0 degrees C; 1.7-7.4 degrees C; p = 0.008). During rewarming to normothermia, the gradient was maintained at 4.5 degrees C. We report for the first time a study, which by direct measurement of deep intracerebral temperatures, validates the cooling cap as an effective method of selective brain cooling in a newborn animal hypoxia-ischemia model. PMID- 11264447 TI - Growth hormone testing and the short child. PMID- 11264448 TI - Toxicology comes of age. AB - This paper contains recollections of some of the people and events that influenced the development of toxicology as an academic discipline. It also describes my experiences in pharmacology at the University of Chicago and the University of Kansas Medical Center and concludes with speculation concerning the future of toxicology. Moderation in all things/Ne quid nimis. --Terence in Andria PMID- 11264449 TI - Anesthetics and ion channels: molecular models and sites of action. AB - The mechanisms of general anesthesia in the central nervous system are finally yielding to molecular examination. As a result of research during the past several decades, a group of ligand-gated ion channels have emerged as plausible targets for general anesthetics. Molecular biology techniques have greatly accelerated attempts to classify ligand-gated ion channel sensitivity to general anesthetics, and have identified the sites of receptor subunits critical for anesthetic modulation using chimeric and mutated receptors. The experimental data have facilitated the construction of tenable molecular models for anesthetic binding sites, which in turn allows structural predictions to be tested. In vivo significance of a putative anesthetic target can now be examined by targeted gene manipulations in mice. In this review, we summarize from a molecular perspective recent advances in our understanding of mechanisms of action of general anesthetics on ligand-gated ion channels. PMID- 11264450 TI - Tumor cell death induced by topoisomerase-targeting drugs. AB - DNA topoisomerases are double-edged swords. They are essential for many vital functions of DNA during normal cell growth. However, they are also highly vulnerable under various physiological and nonphysiological stresses because of their delicate act on breaking and rejoining DNA. These stresses (e.g. exposure to topoisomerase poisons, acidic pH, and oxidative stresses) can convert DNA topoisomerases into DNA-breaking nucleases, resulting in cell death and/or genomic instability. The importance of topoisomerase-mediated DNA cleavage in tumor cell death and carcinogenesis has been recognized. This review focuses on recent findings concerning the molecular mechanisms of the stress responses to topoisomerase-mediated DNA damage. The involvement of ubiquitin/26S proteasome and SUMO/UBC9 in these processes, as well as the role of topoisomerase cleavable complexes in apoptotic cell death are discussed. PMID- 11264451 TI - The clinical pharmacology of L-arginine. AB - L-Arginine (2-amino-5-guanidinovaleric acid) is the precursor of nitric oxide, an endogenous messenger molecule involved in a variety of endothelium-mediated physiological effects in the vascular system. Acute and chronic administration of L-arginine has been shown to improve endothelial function in animal models of hypercholesterolemia and atherosclerosis. L-Arginine also improves endothelium dependent vasodilation in humans with hypercholesterolemia and atherosclerosis. The responsiveness to L-arginine depends on the specific cardiovascular disease studied, the vessel segment, and morphology of the artery. The pharmacokinetics of L-arginine have recently been investigated. Side effects are rare and mostly mild and dose dependent. The mechanism of action of L-arginine may involve nitric oxide synthase substrate provision, especially in patients with elevated levels of the endogenous NO synthase inhibitor asymmetric dimethylarginine. Endocrine effects and unspecific reactions may contribute to L-arginine-induced vasodilation after higher doses. Several long-term studies have been performed that show that chronic oral administration of L-arginine or intermittent infusion therapy with L-arginine can improve clinical symptoms of cardiovascular disease in man. PMID- 11264452 TI - Pharmacogenomics: unlocking the human genome for better drug therapy. AB - There is great heterogeneity in the way humans respond to medications, often requiring empirical strategies to find the appropriate drug therapy for each patient (the "art" of medicine). Over the past 50 years, there has been great progress in understanding the molecular basis of drug action and in elucidating genetic determinants of disease pathogenesis and drug response. Pharmacogenomics is the burgeoning field of investigation that aims to further elucidate the inherited nature of interindividual differences in drug disposition and effects, with the ultimate goal of providing a stronger scientific basis for selecting the optimal drug therapy and dosages for each patient. These genetic insights should also lead to mechanism-based approaches to the discovery and development of new medications. This review highlights the current status of work in this field and addresses strategies that hold promise for future advances in pharmacogenomics. PMID- 11264453 TI - Phenobarbital response elements of cytochrome P450 genes and nuclear receptors. AB - Phenobarbital (PB) response elements are composed of various nuclear receptor (NR)-binding sites. A 51-bp distal element PB-responsive enhancer module (PBREM) conserved in the PB-inducible CYP2B genes contains two NR-binding direct repeat (DR)-4 motifs. Responding to PB exposure in liver, the NR constitutive active receptor (CAR) translocates to the nucleus, forms a dimer with the retinoid X receptor (RXR), and activates PBREM via binding to DR-4 motifs. For CYP3A genes, a common NR site [DR-3 or everted repeat (ER)-6] is present in proximal promoter regions. In addition, the distal element called the xenobiotic responsive module (XREM) is found in human CYP3A4 genes, which contain both DR-3 and ER-6 motifs. Pregnane X receptor (PXR) could bind to all of these sites and, upon PB induction, a PXR:RXR heterodimer could transactivate XREM. These response elements and NRs are functionally versatile, and capable of responding to distinct but overlapping groups of xenochemicals. PMID- 11264454 TI - Regulation and role of adenylyl cyclase isoforms. AB - At least nine closely related isoforms of adenylyl cyclases (ACs), the enzymes responsible for the synthesis of cyclic AMP (cAMP) from ATP, have been cloned and characterized in mammals. Depending on the properties and the relative levels of the isoforms expressed in a tissue or a cell type at a specific time, extracellular signals received through the G-protein-coupled receptors can be differentially integrated. The present review deals with various aspects of such regulations, emphasizing the role of calcium/calmodulin in activating AC1 and AC8 in the central nervous system, the potential inhibitory effect of calcium on AC5 and AC6, and the changes in the expression pattern of the isoforms during development. A particular emphasis is given to the role of cAMP during drug and ethanol dependency and to some experimental limitations (pitfalls in the interpretation of cellular transfection, scarcity of the invalidation models, existence of complex macromolecular structures, etc). PMID- 11264455 TI - The basic and clinical pharmacology of nonpeptide vasopressin receptor antagonists. AB - The neurohypophysial hormone arginine vasopressin (AVP) is a cyclic nonpeptide whose actions are mediated by the stimulation of specific G protein--coupled membrane receptors pharmacologically classified into V1-vascular (V1R), V2-renal (V2R) and V3-pituitary (V3R) AVP receptor subtypes. The random screening of chemical compounds and optimization of lead compounds recently resulted in the development of orally active nonpeptide AVP receptor antagonists. Potential therapeutic uses of AVP receptor antagonists include (a) the blockade of V1 vascular AVP receptors in arterial hypertension, congestive heart failure, and peripheral vascular disease; (b) the blockade of V2-renal AVP receptors in the syndrome of inappropriate vasopressin secretion, congestive heart failure, liver cirrhosis, nephrotic syndrome and any state of excessive retention of free water and subsequent dilutional hyponatremia; (c) the blockade of V3-pituitary AVP receptors in adrenocorticotropin-secreting tumors. The pharmacological and clinical profile of orally active nonpeptide vasopressin receptor antagonists is reviewed here. PMID- 11264456 TI - Novel effects of nitric oxide. AB - Nitric oxide (NO), a simple free radical gas, elicits a surprisingly wide range of physiological and pathophysiological effects. NO interacts with soluble guanylate cyclase to evoke many of these effects. However, NO can also interact with molecular oxygen and superoxide radicals to produce reactive nitrogen species that can modify a number of macromolecules including proteins, lipids, and nucleic acids. NO can also interact directly with transition metals. Here, we have reviewed the non--3',5'-cyclic-guanosine-monophosphate-mediated effects of NO including modifications of proteins, lipids, and nucleic acids. PMID- 11264457 TI - Interactions between monoamines, glutamate, and GABA in schizophrenia: new evidence. AB - In spite of its proven heuristic value, the dopamine hypothesis of schizophrenia is now yielding to a multifactorial view, in which the other monoamines as well as glutamate and GABA are included, with a focus on neurotransmitter interactions in complex neurocircuits. The primary lesion(s) in schizophrenia does not necessarily involve any of these neurotransmitters directly but could deal with a more general defect, such as a faulty connectivity of developmental origin. Nevertheless, a precise identification of neurotransmitter aberrations in schizophrenia will probably provide clues for a better understanding of the disease and for the development of new treatment and prevention strategies. PMID- 11264458 TI - Properties and biological activities of thioredoxins. AB - The mammalian thioredoxins are a family of small (approximately 12 kDa) redox proteins that undergo NADPH-dependent reduction by thioredoxin reductase and in turn reduce oxidized cysteine groups on proteins. The two main thioredoxins are thioredoxin-1, a cytosolic and nuclear form, and thioredoxin-2, a mitochondrial form. Thioredoxin-1 has been studied more. It performs many biological actions including the supply of reducing equivalents to thioredoxin peroxidases and ribonucleotide reductase, the regulation of transcription factor activity, and the regulation of enzyme activity by heterodimer formation. Thioredoxin-1 stimulates cell growth and is an inhibitor of apoptosis. Thioredoxins may play a role in a variety of human diseases including cancer. An increased level of thioredoxin-1 is found in many human tumors, where it is associated with aggressive tumor growth. Drugs are being developed that inhibit thioredoxin and that have antitumor activity. PMID- 11264459 TI - Regulation, function, and tissue-specific expression of cytochrome P450 CYP1B1. AB - Cytochrome P450 CYP1B1 is a relatively recently identified member of the CYP1 gene family. The purpose of this commentary is to review the regulatory mechanisms, metabolic specificity, and tissue-specific expression of this cytochrome P450 and to highlight its unique properties. The regulation of CYP1B1 involves a variety of both transcriptional and post-transcriptional mechanisms. CYP1B1 can metabolize a range of toxic and carcinogenic chemicals in vitro but in some cases with a unique stereoselectivity. Estradiol 4-hydroxylation appears to be a characteristic reaction catalyzed by human CYP1B1. However, there are considerable species differences regarding the regulation, metabolic specificity, and tissue-specific expression of this P450. In humans CYP1B1 is overexpressed in tumor cells, and this has important implications for tumor development and progression and the development of anticancer drugs specifically activated by CYP1B1. PMID- 11264460 TI - Physiological functions of cyclic ADP-ribose and NAADP as calcium messengers. AB - Cyclic ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP) are two Ca(2+) messengers derived from NAD and NADP, respectively. Although NAADP is a linear molecule, structurally distinct from the cyclic cADPR, it is synthesized by similar enzymes, ADP-ribosyl cyclase and its homolog, CD38. The crystal structure of the cyclase has been solved and its active site identified. These two novel nucleotides have now been shown to be involved in a wide range of cellular functions including: cell cycle regulation in Euglena, a protist; gene expression in plants; and in animal systems, from fertilization to neurotransmitter release and long-term depression in brain. A battery of pharmacological reagents have been developed, providing valuable tools for elucidating the physiological functions of these two novel Ca(2+) messengers. This article reviews these recent results and explores the implications of the existence of multiple Ca(2+) messengers and Ca(2+) stores in cells. PMID- 11264461 TI - Use of biomarkers and surrogate endpoints in drug development and regulatory decision making: criteria, validation, strategies. AB - In the future, biomarkers will play an increasingly important role in all phases of drug development, including regulatory review. However, only a few of these biomarkers will become established well enough to serve in regulatory decision making as surrogate endpoints, thereby substituting for traditional clinical endpoints. Even generally accepted surrogate endpoints are unlikely to capture all the therapeutic benefits and potential adverse effects a drug will have in a diverse patient population. Accordingly, combinations of biomarkers probably will be needed to provide a more complete characterization of the spectrum of pharmacologic response. In the future, pharmacogenomic approaches, including those based on differential expression of gene arrays, will provide panels of relevant biomarkers that can be expected to transform the drug development process. PMID- 11264462 TI - Cellular responses to DNA damage. AB - Cells are constantly under threat from the cytotoxic and mutagenic effects of DNA damaging agents. These agents can either be exogenous or formed within cells. Environmental DNA-damaging agents include UV light and ionizing radiation, as well as a variety of chemicals encountered in foodstuffs, or as air- and water borne agents. Endogenous damaging agents include methylating species and the reactive oxygen species that arise during respiration. Although diverse responses are elicited in cells following DNA damage, this review focuses on three aspects: DNA repair mechanisms, cell cycle checkpoints, and apoptosis. Because the areas of nucleotide excision repair and mismatch repair have been covered extensively in recent reviews, we restrict our coverage of the DNA repair field to base excision repair and DNA double-strand break repair. PMID- 11264463 TI - Antisense oligonucleotides: promise and reality. AB - Antisense oligonucleotides have been used for more than a decade to downregulate gene expression. Phosphodiester oligonucleotides are nuclease sensitive, and the more nuclease-resistant phosphorothioate oligonucleotides are now in common use in the laboratory and have entered clinical trials. However, these molecules are highly bioactive and may inhibit gene expression by more than one mechanism. Although some dramatic successes have been demonstrated, it can still be difficult to properly interpret experimental data derived from the use of this class of oligonucleotide. This review discusses some of these issues with particular reference to a major area of current interest--inhibition of bcl-2 expression in tumor cells. PMID- 11264464 TI - Cancer chemoprevention using natural vitamin D and synthetic analogs. AB - Substantial epidemiologic data support a role for vitamin D in cancer prevention. However, dose-limiting hypercalcemic effects have proved a major obstacle to the development of natural vitamin D as a cancer chemopreventive. Structure-activity studies have sought to disassociate the toxicities and chemopreventive activities of vitamin D, and a number of synthetic deltanoids (vitamin D analogs) have shown considerable promise in this regard. Several such compounds have chemopreventive efficacy in preclinical studies, as does natural vitamin D. Data supporting further development of agents of this class include in vitro and in vivo evidence of antiproliferative, proapoptotic, prodifferentiating and antiangiogenic activities. Ongoing studies are aimed at further defining the molecular mechanisms through which vitamin D and synthetic deltanoids affect gene expression and cellular fate. Additional efforts are focused on establishing the chemopreventive index (efficacy vs toxicity) of each synthetic deltanoid. PMID- 11264465 TI - Metabolism of fluorine-containing drugs. AB - This article reviews current knowledge of the metabolism of drugs that contain fluorine. The strategic value of fluorine substitution in drug design is discussed in terms of chemical structure and basic concepts in drug metabolism and drug toxicity. PMID- 11264466 TI - Ca(2+)/CaM-dependent kinases: from activation to function. AB - Calmodulin (CaM) is an essential protein that serves as a ubiquitous intracellular receptor for Ca(2+). The Ca(2+)/CaM complex initiates a plethora of signaling cascades that culminate in alteration of cellular functions. Among the many Ca(2+)/CaM-binding proteins to be discovered, the multifunctional protein kinases CaMKI, II, and IV play pivotal roles. Our review focuses on this class of CaM kinases to illustrate the structural and biochemical basis for Ca(2+)/CaM interaction with and regulation of its target enzymes. Gene transcription has been chosen as the functional endpoint to illustrate the recent advances in Ca(2+)/CaM-mediated signal transduction mechanisms. PMID- 11264467 TI - Lysophospholipid receptors. AB - Lysophospholipids (LPs), including lysophosphatidic acid and sphingosine 1 phosphate, produce many cellular effects. However, the prolonged absence of any cloned and identified LP receptor has left open the question of how these lipids actually bring about these effects. The cloning and functional identification of the first LP receptor, lp(A1)/vzg-1, has led rapidly to the identification and classification of multiple orphan receptors/expression sequence tags known by many names (e.g. edg, mrec1.3, gpcr26, H218, AGR16, nrg-1) as members of a common cognate G protein-coupled receptor family. We review features of the LP receptor family, including molecular characteristics, genomics, signaling properties, and gene expression. A major question for which only partial answers are available concerns the biological significance of receptor-mediated LP signaling. Recent studies that demonstrate the role of receptor-mediated LP signaling in the nervous system, cardiovascular system, and other organ systems indicate the importance of this signaling in development, function, and pathophysiology and portend an exciting time ahead for this growing field. PMID- 11264468 TI - Interindividual variability in inhibition and induction of cytochrome P450 enzymes. AB - Drug interactions have always been a major concern in medicine for clinicians and patients. Inhibition and induction of cytochrome P450 (CYP) enzymes are probably the most common causes for documented drug interactions. Today, many pharmaceutical companies are predicting potential interactions of new drug candidates. Can in vivo drug interactions be predicted accurately from in vitro metabolic studies? Should the prediction be qualitative or quantitative? Although some scientists believe that quantitative prediction of drug interactions is possible, others are less optimistic and believe that quantitative prediction would be very difficult. There are many factors that contribute to our inability to quantitatively predict drug interactions. One of the major complicating factors is the large interindividual variability in response to enzyme inhibition and induction. This review examines the sources that are responsible for the interindividual variability in inhibition and induction of cytochrome P450 enzymes. PMID- 11264469 TI - Neurotrophic and neuroprotective actions of estrogens and their therapeutic implications. AB - Originally known for its regulation of reproductive functions, estradiol, a lipophilic hormone that can easily cross plasma membranes as well as the blood brain barrier, maintains brain systems subserving arousal, attention, mood, and cognition. In addition, both synthetic and natural estrogens exert neurotrophic and neuroprotective effects. There is increasing evidence that estrogen actions are mediated by nongenomic as well as direct and indirect genomic pathways. Although in vitro models have provided the most extensive evidence for neurotrophic and neuroprotective actions to date, there are also in vivo studies that support these actions. PMID- 11264470 TI - Genetic variations and polymorphisms of G protein-coupled receptors: functional and therapeutic implications. AB - G protein-coupled receptors (GPCRs) represent a major class of proteins in the genome of many species, including humans. In addition to the mapping of a number of human disorders to regions of the genome containing GPCRs, a growing body of literature has documented frequently occurring variations (i.e. polymorphisms) in GPCR loci. In this article, we use a domain-based approach to systematically examine examples of genetic variation in the coding and noncoding regions of GPCR loci. Data to date indicate that residues in GPCRs are involved in ligand binding and coupling to G proteins and that regulation can be altered by polymorphisms. Studies of GPCR polymorphisms have also uncovered the functional importance of residues not previously implicated from other approaches that are involved in the function of GPCRs. We predict that studies of GPCR polymorphisms will have a significant impact on medicine and pharmacology, in particular, by providing new means to subclassify patients in terms of both diagnosis and treatment. PMID- 11264471 TI - Drug treatment effects on disease progression. AB - Degenerative diseases are characterized by a worsening of disease status over time. The rate of deterioration is determined by the natural rate of progression of the disease and by the effect of drug treatments. A goal of drug treatment is to slow disease progression. Drug treatments can be categorized as symptomatic or protective. Symptomatic treatments do not affect the rate of disease progression whereas protective treatments have the ability to slow disease progression down. Many current methods for describing disease progression have two common drawbacks: a linear relationship between time and disease status is assumed, and within- and between-subject variability is ignored. Disease progress models combined with pharmacokinetic pharmacodynamic models and hierarchical random effects statistical models provide insights into understanding the time course and management of degenerative disease. PMID- 11264473 TI - Pharmacology of the lower urinary tract. AB - The functions of the lower urinary tract, to store and periodically release urine, are dependent on the activity of smooth and striated muscles in the urinary bladder, urethra, and external urethral sphincter. This activity is in turn controlled by neural circuits in the brain, spinal cord, and peripheral ganglia. Various neurotransmitters, including acetylcholine, norepinephrine, dopamine, serotonin, excitatory and inhibitory amino acids, adenosine triphosphate, nitric oxide, and neuropeptides, have been implicated in the neural regulation of the lower urinary tract. Injuries or diseases of the nervous system, as well as drugs and disorders of the peripheral organs, can produce voiding dysfunctions such as urinary frequency, urgency, and incontinence or inefficient voiding and urinary retention. This chapter will review recent advances in our understanding of the pathophysiology of voiding disorders and the targets for drug therapy. PMID- 11264472 TI - Prostanoid receptors: subtypes and signaling. AB - Cyclooxygenases metabolize arachidonate to five primary prostanoids: PGE(2), PGF(2 alpha), PGI(2), TxA(2), and PGD(2). These autacrine lipid mediators interact with specific members of a family of distinct G-protein-coupled prostanoid receptors, designated EP, FP, IP, TP, and DP, respectively. Each of these receptors has been cloned, expressed, and characterized. This family of eight prostanoid receptor complementary DNAs encodes seven transmembrane proteins which are typical of G-protein-coupled receptors and these receptors are distinguished by their ligand-binding profiles and the signal transduction pathways activated on ligand binding. Ligand-binding selectivity of these receptors is determined by both the transmembrane sequences and amino acid residues in the putative extracellular-loop regions. The selectivity of interaction between the receptors and G proteins appears to be mediated at least in part by the C-terminal tail region. Each of the EP(1), EP(3), FP, and TP receptors has alternative splice variants described that alter the coding sequence in the C-terminal intracellular tail region. The C-terminal variants modulate signal transduction, phosphorylation, and desensitization of these receptors, as well as altering agonist-independent constitutive activity. PMID- 11264474 TI - Role of osteopontin in cellular signaling and toxicant injury. AB - Osteopontin (OPN) is a glycosylated phosphoprotein found in all body fluids and in the proteinaceous matrix of mineralized tissues. It can function both as a cell attachment protein and as a cytokine, delivering signals to cells via a number of receptors including several integrins and CD44. Expression of OPN is enhanced by a variety of toxicants, especially those that activate protein kinase C. In its capacity as a signaling molecule, OPN can modify gene expression and promote the migration of monocytes/macrophages up an OPN gradient. It has both inflammatory and anti-inflammatory actions. Some experiments suggest that it may inhibit apoptosis, possibly contributing to the survival of cells in response to toxicant injury. Elevated OPN expression often correlates with malignancy and has been shown to enhance the tumorigenic and/or metastatic phenotype of the cancer cell. Recent studies have revealed that OPN plays critical roles in bone remodeling and cell-mediated immunity. PMID- 11264475 TI - Compartmentation of G protein-coupled signaling pathways in cardiac myocytes. AB - There is a large body of functional data that supports the existence of subcellular compartmentation of the components of cyclic AMP action in the heart. Data from isolated perfused hearts and from purified ventricular myocytes imply a fixed and hormone-specific spatial relationship amongst components of cyclic AMP synthesis, response, and degradation. Available data demonstrate that within a cardiac myocyte, not all cyclic AMP gains access to all cyclic AMP-dependent protein kinase (PKA), that not all PKA interacts with all possible cellular substrates of PKA, and that only a subset of the myocyte's phosphodiesterases (PDEs) may degrade cyclic AMP after a given synthetic stimulus. Molecular mechanisms contributing to compartmentation are being discovered: localization of receptors, G proteins, and adenylyl cyclases in caveolar versus noncaveolar regions of the sarcolemma; localization of PKA by A-kinase anchoring proteins; localization of PKA substrates, PDE isoforms, and phosphoprotein phosphatases in discrete subcellular regions; and differential regulation of multiple isoforms of adenylyl cyclase, phosphoprotein phosphatase, and PDE in distinct subcellular compartments. PMID- 11264476 TI - Molecular approach to adenosine receptors: receptor-mediated mechanisms of tissue protection. AB - Adenosine accumulation during ischemia and inflammation protects tissues from injury. In ischemic tissues adenosine accumulates due to inhibition of adenosine kinase, and in inflamed tissues adenosine is formed from adenine nucleotides that are released from many cells including platelets, mast cells, nerves, and endothelium. Nucleotides are rapidly converted to adenosine by a family of ecto nucleotidases including CD39 and CD73. Activation of A(1) and possibly A(3) adenosine receptors (ARs) protects heart and other tissues by preconditioning through a pathway including protein kinase C and mitochondrial K(ATP) channels. Activation of A(2A) receptors limits reperfusion injury by inhibiting inflammatory processes in neutrophils, platelets, macrophages and T cells. Adenosine produces proinflammatory responses mediated by receptors that vary among species; A(3) and A(2B) receptors mediate degranulation of rodent and human or canine mast cells, respectively. Novel adenosine receptor subtype-selective ligands have recently been developed. These include MRS1754 (A(2B) blocker), MRS1220 (A(3) blocker), MRE 3008F20 (human A(3) blocker), MRS1523 (rat A(3) blocker), and ATL146e (A(2A) agonist). These new pharmacologic tools will help investigators to sort out how adenosine protects tissues from injury and to identify new therapeutic agents that hold promise for the treatment of inflammatory and ischemic diseases. PMID- 11264477 TI - Molecular targets of lithium action. AB - Lithium is highly effective in the treatment of bipolar disorder and also has multiple effects on embryonic development, glycogen synthesis, hematopoiesis, and other processes. However, the mechanism of lithium action is still unclear. A number of enzymes have been proposed as potential targets of lithium action, including inositol monophosphatase, a family of structurally related phosphomonoesterases, and the protein kinase glycogen synthase kinase-3. These potential targets are widely expressed, require metal ions for catalysis, and are generally inhibited by lithium in an uncompetitive manner, most likely by displacing a divalent cation. Thus, the challenge is to determine which target, if any, is responsible for a given response to lithium in cells. Comparison of lithium effects with genetic disruption of putative target molecules has helped to validate these targets, and the use of alternative inhibitors of a given target can also lend strong support for or against a proposed mechanism of lithium action. In this review, lithium sensitive enzymes are discussed, and a number of criteria are proposed to evaluate which of these enzymes are involved in the response to lithium in a given setting. PMID- 11264479 TI - Endothelin system: the double-edged sword in health and disease. AB - The endothelin system consists of two G-protein-coupled receptors, three peptide ligands, and two activating peptidases. Its pharmacological complexity is reflected by the diverse expression pattern of endothelin system components, which have a variety of physiological and pathophysiological roles. In the vessels, the endothelin system has a basal vasoconstricting role and participates in the development of diseases such as hypertension, atherosclerosis, and vasospasm after subarachnoid hemorrhage. In the heart, the endothelin system affects inotropy and chronotropy, and it mediates cardiac hypertrophy and remodeling in congestive heart failure. In the lungs, the endothelin system regulates the tone of airways and blood vessels, and it is involved in the development of pulmonary hypertension. In the kidney, it controls water and sodium excretion and acid-base balance, and it participates in acute and chronic renal failure. In the brain, the endothelin system modulates cardiorespiratory centers and the release of hormones. More advanced functional analysis of the endothelin system awaits not only additional pharmacological studies using highly specific endothelin antagonists but also the generation of genetically altered rodent models with conditional loss-of-function and gain-of-function manipulations. PMID- 11264478 TI - Molecular basis of ethnic differences in drug disposition and response. AB - Ethnicity is an important demographic variable contributing to interindividual variability in drug metabolism and response. In this rapidly expanding research area many genetic factors that account for the effects of ethnicity on pharmacokinetics, pharmacodynamics, and drug safety have been identified. This review focuses on recent developments that have improved understanding of the molecular mechanisms responsible for such interethnic differences. Genetic variations that may provide a molecular basis for ethnic differences in drug metabolizing enzymes (CYP 2C9, 2C19, 2D6, and 3A4), drug transporter (P glycoprotein), drug receptors (adrenoceptors), and other functionally important proteins (eNOS and G proteins) are discussed. A better understanding of the molecular basis underlying ethnic differences in drug metabolism, transport, and response will contribute to improved individualization of drug therapy. PMID- 11264480 TI - Neurokinin(1) receptor antagonists as potential antidepressants. AB - Selective, nonpeptide antagonists for tachykinin receptors first became available ten years ago. Of the three known tachykinin receptors, drug development has focused most intensively on the substance P-preferring receptor, neurokinin(1) (NK(1)). Although originally studied as potential analgesic compounds, recent evidence suggests that NK(1) receptor antagonists may possess antidepressant and anxiolytic properties. If confirmed by further controlled clinical studies, this will represent a mechanism of action distinct from all existing antidepressant agents. As reviewed in this chapter, the existing preclinical and clinical literature is suggestive of, but not conclusive, concerning a role of substance P and NK(1) receptors in the pathophysiology of depression and/or anxiety disorders. The ongoing clinical trials with NK(1) receptor antagonists have served as an impetus for much needed, basic research in this field. PMID- 11264481 TI - Iodine deficiency as a cause of brain damage. AB - This editorial reviews the impact of iodine deficiency (1) on thyroid function in pregnant women and neonates and (2) on the neurointellectual development of infants and children. All degrees of iodine deficiency (mild: iodine intake of 50 99 microg/day, moderate: 20-49 microg/day, and severe: <20 microg/day) affect thyroid function of the mother and the neonate as well as the mental development of the child. The damage increases with the degree of the deficiency, with overt endemic cretinism as the severest consequence. Maternal hypothyroxinaemia during early pregnancy is a key factor in the development of the neurological damage in the cretin. Selenium deficiency combined with iodine deficiency partly prevents the neurological damage but precipitates severe hypothyroidism in cretins. Iodine deficiency results in a global loss of 10-15 IQ points at a population level and constitutes the world's greatest single cause of preventable brain damage and mental retardation. PMID- 11264483 TI - The immunocompromised patient and transfusion. PMID- 11264484 TI - Review of the current methods in the diagnosis and treatment of scaphoid fractures. AB - If neglected or misdiagnosed, non-union of a scaphoid fracture will almost inevitably progress to radiographic and symptomatic osteoarthritis of the wrist with subsequent morbidity and lifelong disability, especially in young males in which the fracture is more common. Fractures of the scaphoid bone are the most common fractures of the carpus and second in occurrence among fractures of the wrist. The diagnosis and treatment are not simple. Familiarity with different imaging methods and treatment options is required. The treatment in most cases is conservative and will lead to uneventful union, but an operation may be needed in certain cases primarily and in the treatment of non-union. The current literature on the diagnosis and treatment of scaphoid fractures is reviewed, and the authors try to make a clear and concise picture of this complex and sometimes controversial field. PMID- 11264482 TI - Cholesterol gallstones: from epidemiology to prevention. PMID- 11264485 TI - Ethical, professional, and legal obligations in clinical practice: a series of discussion topics for postgraduate medical education. Introduction and topic 1: informed consent. PMID- 11264486 TI - Smoking and diabetes in Chinese men. AB - Smoking is a major cardiovascular risk factor and cause of death. Diabetes mellitus is also associated with an increased mortality and morbidity. Evidence concerning whether smoking increases the incidence of diabetes remains conflicting. Glycaemic status and smoking habits were analysed in 3718 Chinese subjects in order to assess the possible association between smoking and risk of diabetes in the Chinese population. The World Health Organisation 1998 criteria were used for the diagnosis of glucose intolerance. Smoking was defined as current cigarette smoking or ex-smoking without regard to daily consumption. The smoking habits of the studied subjects were correlated with glycaemic status. There were 3003 (80.8%) women and 715 (19.2%) men. The mean age (SD) was 38.4 (12.8) years (median 35.0, range 12-88 years). Of the 3718 subjects, 786 (21.1%) had diabetes, 708 (19.1%) had impaired glucose tolerance, and 2224 (59.8%) had normal results. Of the 3003 women, only 87 (2.9%) were smokers. The female smokers were younger, heavier, and had higher alcohol consumption than non smokers. The prevalence of diabetes was similar between female smokers and non smokers after adjustment for age, body mass index, family history of diabetes, and alcohol. Of the 715 men, 175 (24.5%) were smokers. The male smokers were younger, had lower blood pressure, and higher alcohol consumption. After adjustment for age, body mass index, family history of diabetes and alcohol, the male smokers had lower blood pressure, higher one hour plasma glucose, and more diabetes. Using logistic regression analysis (stepwise forward) with age, body mass index, alcohol, smoking, and family history of diabetes as independent variables to predict the risk of having diabetes, age and body mass index are independently associated with diabetes in both men and women. In addition, smoking is independently associated with the risk of diabetes in men, the odds ratio (95% confidence interval, CI) being 1.705 (1.106 to 2.630). Family history of diabetes is independently associated with the risk of diabetes in women, and the odds ratio (95% CI) is 1.643 (1.314, to 2.053). In conclusion, it was found that smoking is independently associated with diabetes after adjustment for age, body mass index, alcohol, and family history of diabetes in Hong Kong Chinese men, the odds ratio being 1.7. The prevalence of smoking in Hong Kong Chinese women is low and its association with diabetes is inconclusive. PMID- 11264487 TI - Is there a dissociative process in sleepwalking and night terrors? AB - The enduring and contentious hypothesis that sleepwalking and night terrors are symptomatic of a protective dissociative mechanism is examined. This is mobilised when intolerable impulses, feelings and memories escape, within sleep, the diminished control of mental defence mechanisms. They then erupt but in a limited motoric or affective form with restricted awareness and subsequent amnesia for the event. It has also been suggested that such processes are more likely when the patient has a history of major psychological trauma. In a group of 22 adult patients, referred to a tertiary sleep disorders service with possible sleepwalking/night terrors, diagnosis was confirmed both clinically and polysomnographically, and only six patients had a history of such trauma. More commonly these described sleepwalking/night terrors are associated with vivid dream-like experiences or behaviour related to flight from attack. Two such cases, suggestive of a dissociative process, are described in more detail. The results of this study are presented largely on account of the negative findings. Scores on the dissociation questionnaire (DIS-Q) were normal, although generally higher in the small "trauma" subgroup. These were similar to scores characterising individuals with post-traumatic stress disorder. This "trauma" group also scored particularly highly on the anxiety, phobic, and depression scales of the Crown-Crisp experiential index. In contrast the "no trauma" group scored more specifically highly on the anxiety scale, along with major trends to high depression and hysteria scale scores. Two cases are presented which illustrate exceptional occurrence of later onset of sleepwalking/night terrors with accompanying post-traumatic symptoms during wakefulness. It is concluded that a history of major psychological trauma exists in only a minority of adult patients presenting with sleepwalking/night terror syndrome. In this subgroup trauma appears to dictate the subsequent content of the attacks. However, the symptoms express themselves within the form of the sleepwalking/night terror syndrome rather than as rapid eye movement sleep related nightmares. The main group of subjects with the syndrome and with no history of major psychological trauma show no clinical or DIS-Q evidence of dissociation during wakefulness. The proposition that, within the character structure of this group, the mechanism still operates but exclusively within sleep remains a possibility. PMID- 11264488 TI - Unexplained groin pain: safety and reliability of herniography for the diagnosis of occult hernias. AB - A retrospective study of our initial experience of herniography in a district general hospital is presented. A total of 43 herniograms were performed in 41 patients (median age 57, range 16-77, 27 males, 14 females) over a two year period. Four herniograms were unsuccessful due to failed intraperitoneal contrast injection, of which two were repeated (success rate 90.5%). A total of 25 groin hernias were identified radiologically (two on the asymptomatic side). Twenty one patients underwent surgery and a hernia was confirmed in 19 (true positive rate 90.5%). Sixteen herniograms were considered negative and after a median follow up of 28 months (range 16-42 months), none of these patients have developed a hernia. There were no major complications. It is concluded that herniography is a safe and reliable method of determining or excluding the presence of an occult groin hernia. PMID- 11264489 TI - Proctocolitis in breast fed infants: a contribution to differential diagnosis of haematochezia in early childhood. AB - Dietary protein induced proctocolitis in exclusively breast fed infants is rarely taken into consideration as a cause of rectal bleeding or blood streaked stool in the neonatal period and early infancy. Eleven babies are presented in whom it is believed that bleeding through the rectum was due to proctocolitis as a result of allergy triggered by cows' milk protein transferred to the infants via the breast milk. Colonoscopy was performed in five infants, revealing benign eosinophilic proctocolitis. Standard treatment was the exclusion of the allergen from the mother's diet. Resolution of visible rectal bleeding took place within 72 to 96 hours after elimination of the offending protein from the mother's diet. PMID- 11264491 TI - Dysphagia and thoracoabdominal aneurysm. AB - Two elderly patients who presented with gradually progressive dysphagia are described. Investigations excluded an intraluminal obstruction and showed extrinsic compression of the oesophagus by an aneurysmal aorta. Surgery was not performed and they were successfully managed with a liquid diet. PMID- 11264490 TI - Adenocarcinoma of the gastro-oesophageal junction with a synchronous carcinoid of the duodenum. AB - Duodenal carcinoids are rare tumours. There is an increased incidence of primary carcinomas, especially in the gastrointestinal tract, which occur synchronously with gastrointestinal tract carcinoids. However, the synchronous occurrence of adenocarcinoma of the gastro-oesophageal junction with a duodenal carcinoid has not been previously described. A case report is presented, with discussion of carcinoid tumours and management when occurring synchronously with non-carcinoid tumours. PMID- 11264492 TI - Staphylococcus lugdunensis endocarditis. AB - A case of Staphylococcus lugdunensis endocarditis is presented with low back pain suggesting a secondary bone focus of infection. An umbilical skin lesion may have been an additional embolic phenomenon. The case highlights the aggressive nature of S lugdenensis endocarditis compared with other coagulase negative staphylococci and its association with native heart valves. In addition the importance of full identification of coagulase negative staphylococci isolated from patient samples in a case of suspected S lugdenensis infection is emphasised. Antibiotic treatment may be insufficient alone in the treatment of S lugdenensis endocarditis and early recourse to surgical intervention and valve replacement should therefore be considered. PMID- 11264493 TI - It could only happen to a doctor--Haemophilus aphrophilus septicaemia complicated by a prevertebral infection after dental work. AB - A 53 year old man presented with severe neck pain and a flu-like illness; he had recently returned from Sri Lanka and had had dental treatment six days before illness onset. Blood culture showed infection by Haemophilus aphrophilus. Magnetic resonance imaging was performed and exploratory surgery undertaken. The prevertebral cervical fascia was inflamed but no abscess identified. He was treated with antibiotics and made an uneventful recovery. PMID- 11264494 TI - Does history repeat itself in medicine? PMID- 11264495 TI - Systemic sarcoidosis with spleen involvement. PMID- 11264496 TI - Diagnostic issues in systemic lupus erythematosis. PMID- 11264497 TI - Optic disc oedema in first degree relatives with different macrovascular risk factors (type 1 diabetes and hypertension). PMID- 11264498 TI - An unusual palmoplantar pigmentation. PMID- 11264499 TI - An unusual cause of tremor in an elderly man. PMID- 11264500 TI - A 35 year man with acromegaly and neck stiffness. PMID- 11264501 TI - Joint pains, hoarseness, and deafness. PMID- 11264502 TI - Helicobacter heilmannii. PMID- 11264503 TI - Chronic pulmonary suppuration. PMID- 11264504 TI - A transient pleural effusion. PMID- 11264524 TI - Direct detection of galactic halo dark matter. AB - The Milky Way galaxy contains a large, spherical component which is believed to harbor a substantial amount of unseen matter. Recent observations indirectly suggest that as much as half of this "dark matter" may be in the form of old, very cool white dwarfs, the remnants of an ancient population of stars as old as the galaxy itself. We conducted a survey to find faint, cool white dwarfs with large space velocities, indicative of their membership in the galaxy's spherical halo component. The survey reveals a substantial, directly observed population of old white dwarfs, too faint to be seen in previous surveys. This newly discovered population accounts for at least 2 percent of the halo dark matter. It provides a natural explanation for the indirect observations, and represents a direct detection of galactic halo dark matter. PMID- 11264505 TI - A young woman with muscle weakness. PMID- 11264525 TI - Observation of vortex lattices in Bose-Einstein condensates. AB - Quantized vortices play a key role in superfluidity and superconductivity. We have observed the formation of highly ordered vortex lattices in a rotating Bose condensed gas. These triangular lattices contained over 100 vortices with lifetimes of several seconds. Individual vortices persisted up to 40 seconds. The lattices could be generated over a wide range of rotation frequencies and trap geometries, shedding light on the formation process. Our observation of dislocations, irregular structure, and dynamics indicates that gaseous Bose Einstein condensates may be a model system for the study of vortex matter. PMID- 11264526 TI - A Bose-Einstein condensate of metastable atoms. AB - We report the realization of a Bose-Einstein condensate of metastable atoms (helium in the lowest triplet state). The excitation energy of each atom with respect to the ground state is 20 electron volts, but inelastic processes that would destroy the sample are suppressed strongly enough in a spin-polarized sample to allow condensation. Our detection scheme takes advantage of the metastability to achieve detection of individual atoms as well as of the decay products of inelastic processes. This detection opens the way toward new studies in mesoscopic quantum statistical physics, as well as in atomic quantum optics. PMID- 11264527 TI - Noble gases in mantle plumes. PMID- 11264529 TI - Squeezed states in a Bose-Einstein condensate. AB - We report manipulation of the atom number statistics associated with Bose Einstein condensed atoms confined in an array of weakly linked mesoscopic traps. We used the interference of atoms released from the traps as a sensitive probe of these statistics. By controlling relative strengths of the tunneling rate between traps and atom-atom interactions within each trap, we observed trap states characterized by sub-Poissonian number fluctuations and adiabatic transitions between these number-squeezed states and coherent states of the atom field. The quantum states produced in this work may enable substantial gains in sensitivity for atom interference-based instruments as well as fundamental studies of quantum phase transitions. PMID- 11264528 TI - Space-geodetic constraints on glacial isostatic adjustment in Fennoscandia. AB - Analysis of Global Positioning System (GPS) data demonstrates that ongoing three dimensional crustal deformation in Fennoscandia is dominated by glacial isostatic adjustment. Our comparison of these GPS observations with numerical predictions yields an Earth model that satisfies independent geologic constraints and bounds both the average viscosity in the upper mantle (5 x 10(20) to 1 x 10(21) pascal seconds) and the elastic thickness of the lithosphere (90 to 170 kilometers). We combined GPS-derived radial motions with Fennoscandian tide gauge records to estimate a regional sea surface rise of 2.1 +/- 0.3 mm/year. Furthermore, ongoing horizontal tectonic motions greater than approximately 1 mm/year are ruled out on the basis of the GPS-derived three-dimensional crustal velocity field. PMID- 11264530 TI - Electrochromic nanocrystal quantum dots. AB - Incorporating nanocrystals into future electronic or optoelectronic devices will require a means of controlling charge-injection processes and an understanding of how the injected charges affect the properties of nanocrystals. We show that the optical properties of colloidal semiconductor nanocrystal quantum dots can be tuned by an electrochemical potential. The injection of electrons into the quantum-confined states of the nanocrystal leads to an electrochromic response, including a strong, size-tunable, midinfrared absorption corresponding to an intraband transition, a bleach of the visible interband exciton transitions, and a quench of the narrow band-edge photoluminescence. PMID- 11264531 TI - In situ measurement of grain rotation during deformation of polycrystals. AB - Texture evolution governs many of the physical, chemical, and mechanical properties of polycrystalline materials, but texture models have only been tested on the macroscopic level, which makes it hard to distinguish between approaches that are conceptually very different. Here, we present a universal method for providing data on the underlying structural dynamics at the grain and subgrain level. The method is based on diffraction with focused hard x-rays. First results relate to the tensile deformation of pure aluminum. Experimental grain rotations are inconsistent with the classical Taylor and Sachs models. PMID- 11264532 TI - Turing-type patterns on electrode surfaces. AB - We report stationary, nonequilibrium potential and adsorbate patterns with an intrinsic wavelength that were observed in an electrochemical system with a specific type of current/electrode-potential (I-phi(DL)) characteristic. The patterns emerge owing to the interplay of a self-enhancing step in the reaction dynamics and a long-range inhibition by migration currents rather than by diffusion. Theoretical analysis revealed that this self-structuring of the electrode occurs in all electrochemical systems with an S-shaped I-phi(DL) characteristic in wide and well-accessible parameter ranges. This unusual pattern forming instability in electrochemical systems has all the characteristics of the mechanism proposed by Turing in 1952 in the framework of an early theory of morphogenesis. Our finding might account for structure formation in certain biological systems that have gradients in the electric potential and may open new paths for fabricating patterned electrodes. PMID- 11264533 TI - Biogenic carbon cycling in the upper ocean: effects of microbial respiration. AB - Food-web processes are important controls of oceanic biogenic carbon flux and ocean-atmosphere carbon dioxide exchange. Two key controlling parameters are the growth efficiencies of the principal trophic components and the rate of carbon remineralization. We report that bacterial growth efficiency is an inverse function of temperature. This relationship permits bacterial respiration in the euphotic zone to be computed from temperature and bacterial production. Using the temperature-growth efficiency relationship, we show that bacterial respiration generally accounts for most community respiration. This implies that a larger fraction of assimilated carbon is respired at low than at high latitudes, so a greater proportion of production can be exported in polar than in tropical regions. Because bacterial production is also a function of temperature, it should be possible to compute euphotic zone heterotrophic respiration at large scales using remotely sensed information. PMID- 11264534 TI - Variation of crystal dissolution rate based on a dissolution stepwave model. AB - A formulation based on defect-generated dissolution stepwaves of the variation of dissolution rate with the degree of undersaturation is validated by near-atomic scale observations of surfaces, Monte Carlo simulations, and experimental bulk dissolution rates. The dissolution stepwaves emanating from etch pits provide a train of steps similar to those of a spiral but with different behavior. Their role in accounting for the bulk dissolution rate of crystals provides a conceptual framework for mineral dissolution far from equilibrium. Furthermore, the law extends research to conditions closer to equilibrium and predicts a nonlinear decrease in the rate of dissolution as equilibrium is approached, which has implications for understanding artificial and natural processes involving solid-fluid reactions. PMID- 11264536 TI - Receptor-mediated activation of heterotrimeric G-proteins in living cells. AB - Receptor-mediated activation of heterotrimeric GTP-binding proteins (G-proteins) was visualized in living Dictyostelium discoideum cells by monitoring fluorescence resonance energy transfer (FRET) between alpha- and beta- subunits fused to cyan and yellow fluorescent proteins. The G-protein heterotrimer rapidly dissociated and reassociated upon addition and removal of chemoattractant. During continuous stimulation, G-protein activation reached a dose-dependent steady state level. Even though physiological responses subsided, the activation did not decline. Thus, adaptation occurs at another point in the signaling pathway, and occupied receptors, whether or not they are phosphorylated, catalyze the G protein cycle. Construction of similar energy-transfer pairs of mammalian G proteins should enable direct in situ mechanistic studies and applications such as drug screening and identifying ligands of newly found G-protein-coupled receptors. PMID- 11264535 TI - Regulation of vegetative phase change in Arabidopsis thaliana by cyclophilin 40. AB - During its development, a plant shoot progresses from a juvenile to an adult phase of vegetative growth and from a reproductively incompetent to a reproductively competent state. In Arabidopsis, loss-of-function mutations in SQUINT (SQN) reduced the number of juvenile leaves and had subtle effects on inflorescence morphology but had no effect on flowering time or on reproductive competence. SQN encodes the Arabidopsis homolog of cyclophilin 40 (CyP40), a protein found in association with the Hsp90 chaperone complex in yeast, mammals, and plants. Thus, in Arabidopsis, CyP40 is specifically required for the vegetative but not the reproductive maturation of the shoot. PMID- 11264537 TI - Length of the flagellar hook and the capacity of the type III export apparatus. AB - Length determination in biology generally uses molecular rulers. The hook, a part of the flagellum of motile bacteria, has an invariant length. Here, we examined hook length and found that it was determined not by molecular rulers but probably by the amount of subunit protein secreted by the flagellar export apparatus. The export apparatus shares common features with the type III virulence-factor secretion machinery and thus may be used more widely in length determination of structures other than flagella. PMID- 11264538 TI - Preferential localization of effector memory cells in nonlymphoid tissue. AB - Many intracellular pathogens infect a broad range of host tissues, but the importance of T cells for immunity in these sites is unclear because most of our understanding of antimicrobial T cell responses comes from analyses of lymphoid tissue. Here, we show that in response to viral or bacterial infection, antigen specific CD8 T cells migrated to nonlymphoid tissues and were present as long lived memory cells. Strikingly, CD8 memory T cells isolated from nonlymphoid tissues exhibited effector levels of lytic activity directly ex vivo, in contrast to their splenic counterparts. These results point to the existence of a population of extralymphoid effector memory T cells poised for immediate response to infection. PMID- 11264539 TI - Memory extinction, learning anew, and learning the new: dissociations in the molecular machinery of learning in cortex. AB - The rat insular cortex (IC) subserves the memory of conditioned taste aversion (CTA), in which a taste is associated with malaise. When the conditioned taste is unfamiliar, formation of long-term CTA memory depends on muscarinic and beta adrenergic receptors, mitogen-activated protein kinase (MAPK), and protein synthesis. We show that extinction of CTA memory is also dependent on protein synthesis and beta-adrenergic receptors in the IC, but independent of muscarinic receptors and MAPK. This resembles the molecular signature of the formation of long-term memory of CTA to a familiar taste. Thus, memory extinction shares molecular mechanisms with learning, but the mechanisms of learning anew differ from those of learning the new. PMID- 11264540 TI - Activity-dependent transfer of brain-derived neurotrophic factor to postsynaptic neurons. AB - Neurotrophins such as brain-derived neurotrophic factor (BDNF) are thought to be transferred from post- to presynaptic neurons and to be involved in the formation and plasticity of neural circuits. However, direct evidence for a transneuronal transfer of BDNF and its relation to neuronal activity remains elusive. We simultaneously injected complementary DNAs of green fluorescent protein (GFP) tagged BDNF and red fluorescence protein into the nucleus of single neurons and visualized expression, localization, and transport of BDNF in living neurons. Fluorescent puncta representing BDNF moved in axons in the anterograde direction, though some moved retrogradely, and transferred to postsynaptic neurons in an activity-dependent manner. PMID- 11264541 TI - Interference by huntingtin and atrophin-1 with cbp-mediated transcription leading to cellular toxicity. AB - Expanded polyglutamine repeats have been proposed to cause neuronal degeneration in Huntington's disease (HD) and related disorders, through abnormal interactions with other proteins containing short polyglutamine tracts such as the transcriptional coactivator CREB binding protein, CBP. We found that CBP was depleted from its normal nuclear location and was present in polyglutamine aggregates in HD cell culture models, HD transgenic mice, and human HD postmortem brain. Expanded polyglutamine repeats specifically interfere with CBP-activated gene transcription, and overexpression of CBP rescued polyglutamine-induced neuronal toxicity. Thus, polyglutamine-mediated interference with CBP-regulated gene transcription may constitute a genetic gain of function, underlying the pathogenesis of polyglutamine disorders. PMID- 11264542 TI - Two-state allosteric behavior in a single-domain signaling protein. AB - Protein actions are usually discussed in terms of static structures, but function requires motion. We find a strong correlation between phosphorylation-driven activation of the signaling protein NtrC and microsecond time-scale backbone dynamics. Using nuclear magnetic resonance relaxation, we characterized the motions of NtrC in three functional states: unphosphorylated (inactive), phosphorylated (active), and a partially active mutant. These dynamics are indicative of exchange between inactive and active conformations. Both states are populated in unphosphorylated NtrC, and phosphorylation shifts the equilibrium toward the active species. These results support a dynamic population shift between two preexisting conformations as the underlying mechanism of activation. PMID- 11264543 TI - A clinician's perspective on clinical trials. PMID- 11264544 TI - Predicting response to carvedilol for the treatment of heart failure: a multivariate retrospective analysis. AB - BACKGROUND: Carvedilol has been shown to decrease the progression of heart failure and improve left ventricular function and survival in patients with a left ventricular ejection fraction (LVEF) less than 35%. However, not all patients respond uniformly to this therapy. We proposed to identify variables that could, potentially, be used to predict response to carvedilol therapy as measured by the change in LVEF after treatment (Delta LVEF), and to identify pretreatment variables associated with hospitalization for heart failure after carvedilol therapy. METHODS AND RESULTS: A retrospective analysis of 98 patients treated with open-label carvedilol for a mean period of 16 months was performed by using bivariate and step-wise multivariate analyses. Bivariate analysis showed a positive correlation of Delta LVEF with heart rate at baseline (P =.001). There was a negative correlation of Delta LVEF with baseline LVEF (P <.01), diabetes mellitus (P =.04), and ischemic cardiomyopathy (P =.0002). Multivariate analysis showed a positive correlation of Delta LVEF with heart rate at baseline (P =.01) and a negative correlation with initial LVEF (P =.02) and ischemic cardiomyopathy (P =.006). Variables associated with hospitalization after initiation of carvedilol therapy were New York Heart Association (NYHA) classification (P =.001), lower extremity edema (P =.001), presence of an S3 (P =.02), hyponatremia (P =.02), elevated blood urea nitrogen (BUN) (P =.002), atrial fibrillation (P =.001), diabetes mellitus (P =.02), and obstructive sleep apnea (P =.009). CONCLUSIONS: Heart failure patients with the lowest LVEF or the highest heart rate at baseline had the greatest gain in LVEF after treatment with carvedilol. Patients with ischemic cardiomyopathy derived less benefit. Patients with clinical evidence of decompensated heart failure were at greater risk for hospitalization after initiation of carvedilol therapy. PMID- 11264545 TI - Radiographic measurements of cardiac size as predictors of outcome in patients with dilated cardiomyopathy. AB - BACKGROUND: Cardiac dilatation is a predictor of poor outcome in patients with dilated cardiomyopathy. Whereas cardiac chamber dimensions or volumes can be assessed by various noninvasive and invasive techniques, simple chest radiography also may provide a valuable assessment of cardiac size. METHODS AND RESULTS: To determine the relative power of radiographic heart measurements for predicting outcome in dilated cardiomyopathy, we retrospectively studied 88 adult patients with chest radiographs obtained within 35 days of echocardiography. Standard radiographic variables were measured for each patient, and the cardiothoracic (CT) ratio, frontal cardiac area, and volume were calculated. During a mean 4.1 year follow-up, 62 of the 88 (71%) patients died. CT ratio was the best predictor of mortality among the radiographic cardiac measurements. By multivariate analysis, a model including echocardiographic ejection fraction, New York Heart Association (NYHA) functional class, and history of heart failure was highly predictive of survival. When added to this model, CT ratio also was independently associated with mortality, but not radiographic cardiac area or volume. When radiographic variables were each added to CT ratio, they did not add incremental predictive value to the model that included CT ratio alone. Echocardiographic measurement of left ventricular (LV) size, especially when indexed for body size, was independently predictive of outcome, but it did not supersede the predictive power of CT ratio. CONCLUSION: The simply derived radiographic CT ratio is a useful predictor of outcome in patients with dilated cardiomyopathy and compares favorably with other clinical and selected echocardiographic variables. PMID- 11264546 TI - A rapid test for B-type natriuretic peptide correlates with falling wedge pressures in patients treated for decompensated heart failure: a pilot study. AB - OBJECTIVES: To determine if changes in B-type natriuretic peptide (BNP) levels can accurately reflect acute changes in pulmonary capillary wedge pressure during treatment of decompensated heart failure. BACKGROUND: Tailored therapy of decompensated congestive heart failure with hemodynamic monitoring is controversial. Other than the expense and complications of Swan-Ganz catheters, its use in titration of drug therapy has no conclusive end point. Because BNP reflects both elevated left ventricular pressure and neurohormonal modulation and has a short half-life, we hypothesized that levels of BNP would decline in association with falling wedge pressures. Final BNP levels would perhaps signify a new set point of neuromodulation. METHODS AND RESULTS: Twenty patients with decompensated New York Heart Association (NYHA) class III-IV congestive heart failure (CHF) undergoing tailored therapy were studied. BNP levels were drawn every 2 to 4 hours for the first 24 hours (active treatment phase) and then every 4 hours for the next 24 to 48 hours (stabilization period). Hemodynamic data was recorded simultaneously. In 15 patients whose wedge pressure responded to treatment in the first 24 hours, there was a significant drop in BNP levels (55%) versus nonresponders (8%). There was a significant correlation between percent change in wedge pressure from baseline per hour and the percent change of BNP from baseline per hour (r = 0.79, P <.05). When the wedge pressure was kept at a stable, low level during the stabilization phase, BNP levels continued to fall another 37% (937 +/- 140 pg/mL at 24 hours to 605 +/- 128 pg/mL). Patients who died (n = 4) had higher final BNP levels (1,078 +/- 123 pg/mL v 701 +/- 107 pg/mL). CONCLUSIONS: The data suggest that rapid testing of BNP may be an effective way to improve the in-hospital management of patients admitted with decompensated CHF. Although BNP levels will not obviate the need for invasive hemodynamic monitoring, it may be a useful adjunct in tailoring therapy to these patients. PMID- 11264547 TI - Left ventricular function after a new pregnancy in patients with peripartum cardiomyopathy. AB - BACKGROUND: A new pregnancy is usually discouraged in patients with peripartum cardiomyopathy (PPCM), particularly when there is persistent left ventricular dysfunction. This study was undertaken to evaluate left ventricular systolic function after a new pregnancy in patients with PPCM. METHODS AND RESULTS: Nine of 44 patients with PPCM became pregnant and were selected for this study. Two patients were lost to follow-up, 1 immediately after the new pregnancy diagnosis, and the other 1 after the latest delivery, and, thus, were excluded. The remaining 7 patients had regular clinical and obstetric examinations until delivery, continued follow-up, and were submitted to echocardiography 6 to 12 months thereafter. Pregnancy was relatively well tolerated in the patients, and they gave birth to 7 healthy newborns. After this latest pregnancy, 4 patients with heart failure functional class II and 2 patients with functional class III remained unchanged. A patient, initially in functional class III, improved and was then in functional class II. Although left ventricular end-diastolic diameter did not change (61 to 58 mm), left ventricular end-systolic dimension decreased (50 to 47 mm, P =.008), resulting in a significant increase in left ventricular fractional shortening (19% to 23%, P =.02). CONCLUSION: Although based only in a small number of patients, the present results suggest that cardiac function does not deteriorate during a new pregnancy in patients with PPCM. PMID- 11264548 TI - Risks of repeat pregnancy after peripartum cardiomyopathy: double jeopardy. PMID- 11264549 TI - Blunted peripheral vasodilatory response is a hallmark of progressive deterioration in mild to moderate congestive heart failure. AB - BACKGROUND: Several reports have shown that dilatory response to acetylcholine (ACh) and nitroprusside (SNP) is blunted in the limb vasculature in patients with congestive heart failure (CHF). However, it is not yet known whether this vascular dysfunction is related to clinical outcome. We have examined the relationship between peripheral vasodilatory response and prognosis of CHF. METHODS AND RESULTS: A total of 46 patients with mild to moderate CHF were enrolled (mean age 56 years). Changes in forearm blood flow (FBF) during intra arterial infusion of ACh and SNP were determined by plethysmography. FBF changes above baseline for each dose were cumulated and used as an index of endothelium dependent (ACh) response and endothelium-independent (SNP) response, respectively. During the follow-up period (mean 32 months), 9 patients were admitted to the hospital for treatment of worsening refractory CHF, and 6 patients died suddenly or developed life-threatening arrhythmia. By Kaplan-Meier analysis, when all cardiac events were included, no significant differences were observed between any levels of vascular response in terms of prognosis. However, when deterioration events were analyzed separately, patients with SNP responses below the median (7.4 mL/min/dL) had significantly higher rates of hospital admission caused by worsening CHF than those with above the median responses (P <.05). This relationship was not found between ACh response and clinical outcome. By Cox multivariate analysis, blunted vasodilatory response to SNP was a significant predictor of worsening CHF (chi(2) = 3.95; P <.05). CONCLUSION: This study has shown that patients with mild to moderate CHF showing a blunted vascular response to SNP rather than ACh were admitted to the hospital more frequently because of deterioration of CHF. This finding suggests that changes in vascular smooth muscle and/or vascular structure in the peripheral vasculature may be a critical element in the worsening of CHF. PMID- 11264550 TI - Atrial fibrillation impairs endothelial function of forearm vessels in humans. AB - BACKGROUND: Although there have been many studies on the effects of atrial fibrillation (AF) on cardiac function, few studies have been done on its effects on endothelial function. The present study was designed to examine the effects of AF on endothelial function in human subjects. METHODS AND RESULTS: Changes in forearm blood flow (FBF) induced by acetylcholine and nitroglycerin were measured by using plethysmography in 14 patients with lone AF, 13 patients with AF and underlying heart disease, and 12 normal control subjects. In the patients, these measurements were repeated after cardioversion. Although baseline FBF was the same in the 3 groups, acetylcholine-induced increases in FBF were significantly smaller in both patient groups than in the control group, and FBF increases were particularly depressed in AF patients with underlying heart disease. After restoration of sinus rhythm by cardioversion, FBF response to the highest dose of acetylcholine increased by 46% in patients with lone AF (n = 10) and by 90% in AF patients with underlying heart disease (n = 11). Nitroglycerin-induced vasodilatation was the same in all 3 groups and was not affected by cardioversion. CONCLUSIONS: These findings suggest that endothelium-dependent vasodilatation is impaired by AF and improves after sinus rhythm is restored. PMID- 11264552 TI - Prospective evaluation of an outpatient heart failure management program. AB - BACKGROUND: Although considerable effort has been devoted to the follow-up of hospitalized patients, the effectiveness and process of heart failure outpatient management have not been well demonstrated. METHODS AND RESULTS: All new patients referred to the program from April 1997 to September 1998 were followed and managed by comprehensive strategies including preemptive hospitalization. Quality of life (QOL) and patients' self-care adherence behaviors were measured at baseline, 3 months, and 6 months. Clinical outcomes were compared for the 6 months before and 6 months after referral. A total of 108 patients were recruited. Patients' self-care knowledge score was improved over time (difference score = 0.9, P <.01). The proportion of patients weighing themselves daily increased by 24% (P =.02). The proportion of patients with New York Heart Association (NYHA) class III to IV was 67.6% at baseline and 49.1% at 6 months (P =.01). Compared with 6 months before referral, the program intervention was accompanied by a 52% reduction in the risk of hospitalization for cardiovascular causes (56.1% v 27.2%, P <.001) and a 72% reduction in emergency room visits (53.6% v 14.5%, P <.01). The total hospital admissions for cardiovascular causes decreased by 59% from 94 to 39; the total emergency room visits decreased by 77% from 83 to 19. The patients' QOL was improved over time with a change score of 11.2 (P <.001) at 3 months and 10.7 (P <.001) at 6 months. CONCLUSION: Our study shows the effectiveness of this heart failure outpatient management program. PMID- 11264551 TI - Long-acting natriuretic peptide, vessel dilator, and kaliuretic peptide enhance urinary excretion rate of albumin, total protein, and beta(2)-microglobulin in patients with congestive heart failure. AB - BACKGROUND: Albumin excretion is increased in persons with congestive heart failure (CHF), but the mechanism of this increase is unknown. Atrial natriuretic peptide (ANP) does not correlate with this albumin excretion, but the other 3 cardiac hormones derived from the ANP prohormone have never been investigated as to whether they can enhance albumin and/or protein excretion in persons with CHF. METHODS AND RESULTS: Long-acting natriuretic peptide (LANP), vessel dilator, and kaliuretic peptide (100 ng/kg body weight/min) given intravenously for 60 minutes to NYHA Class III CHF patients (n = 24) increased the albumin excretion rate 2- to 7-fold (P <.001) and total protein excretion rate 2- to 5-fold (P <.001). These peptide hormones similarly enhanced beta(2)-microglobulin, a specific marker of proximal tubular reabsorption, excretion rate 25- to 40-fold (P <.0005) at the end of their respective infusions. Three hours after stopping their respective infusions, the beta(2)-microglobulin excretion rate was still 11- to 33-fold (P <.0005) increased. CONCLUSIONS: LANP, vessel dilator, and kaliuretic peptide each enhance albumin and total protein excretion in persons with CHF. Part of the mechanism of this enhanced protein excretion is the inhibition of proximal tubular reabsorption of protein as shown by the beta(2)-microglobulin data. PMID- 11264553 TI - Attenuated natriuretic response to adrenomedullin in experimental heart failure. AB - BACKGROUND: The recently discovered vasodilating and positive inotropic peptide, adrenomedullin (ADM), has strong natriuretic actions. ADM-induced natriuresis is caused by an increase in glomerular filtration rate and a decrease in distal tubular sodium reabsorption. Although ADM is activated in human and experimental heart failure, the role of ADM in the kidney in heart failure remains undefined. METHODS AND RESULTS: The present study was performed to determine the renal hemodynamic and urinary excretory actions of exogenously administered ADM in a canine model of acute heart failure produced by rapid ventricular pacing. Experimental acute heart failure was characterized by a decrease in cardiac output and an increase in pulmonary capillary wedge pressure with an increase in plasma ADM concentration. Intrarenal infusion of ADM (1 and 25 ng/kg/min) induced an increase in urinary sodium excretion in the normal control dogs (change in urinary sodium excretion [Delta UNaV], +94.5 microEq/min during 1 ng/kg/min ADM infusion and +128.1 microEq/min during 25 ng/kg/min ADM infusion). In the acute heart failure dogs, intrarenal ADM infusion resulted in an attenuated increase in urinary sodium excretion (Delta UNaV, +44.8 microEq/min during 1 ng/kg/min ADM infusion and +51.8 microEq/min during 25 ng/kg/min ADM infusion). Both glomerular and tubular actions of ADM were attenuated in the acute heart failure group compared with responses in the normal control group. CONCLUSION: The present study shows that the renal natriuretic responses to ADM are markedly attenuated in experimental acute heart failure. This study provides insight into humoral mechanisms that may promote sodium retention in heart failure via a renal hyporesponsiveness to natriuretic actions of ADM. PMID- 11264554 TI - Expression and activity of pulmonary endothelin converting enzyme in heart failure: relation to endothelin biosynthesis and receptor distribution. AB - BACKGROUND: Although reduced pulmonary clearance of endothelin-1 (ET-1) has been suggested to contribute to increased circulating levels in congestive heart failure (CHF), the regulation of the pulmonary ET system with CHF remains to be defined. Accordingly, the aim of the present study is to investigate the expression and activity of the ET system with the development of CHF. METHODS AND RESULTS: Pulmonary tissue samples were collected from pigs with pacing CHF (240 bpm, 3 wks, n = 10) and controls (n = 10). The pulmonary messenger RNA (mRNA) and protein levels of endothelin converting enzyme-1 (ECE-1) subisoforms, ET-1, and ET receptor profiles were determined. The gene expression of ET-1 precursor, ECE 1a, and ET(A) was upregulated 4-, 3-, and 2-fold, respectively, with CHF. Pulmonary tissue ET-1 was increased to 13 +/- 2 fmol/mg protein from control values of 5 +/- 1 fmol/mg protein (P <.05), and ECE-1 activity was augmented from 3,264 +/- 665 fmol/mg protein in control animals to 14,073 +/- 654 fmol/mg protein per hour in CHF animals (P <.05). The ET(B) receptor density decreased, whereas ET(A) receptors were increased in CHF, indicating a shift in the ET(A) to ET(B) ratio. CONCLUSIONS: Both the increased synthesis and the decreased clearance of ET-1 via ET(B) receptors may contribute to the increased systemic and pulmonary ET-1 levels in CHF. PMID- 11264555 TI - Nesiritide for the treatment of decompensated heart failure. AB - Nesiritide (human recombinant B-type natriuretic peptide) binds to receptors in the vasculature, kidney, and other organs to mimic the actions of endogenous natriuretic peptides. Intravenous infusion of nesiritide has been studied in more than 1,700 patients with acute decompensated heart failure (HF). Nesiritide causes potent, dose-related vasodilation that is rapid in onset and sustained for the duration of drug infusion. There is balanced arterial and venous dilation as reflected by decreases in systemic vascular resistance, systemic arterial pressure, pulmonary capillary wedge pressure, right atrial pressure, and mean pulmonary arterial pressure. Vasodilation occurs without a change in heart rate and is associated with increases in stroke volume and cardiac output. Nesiritide may promote diuresis because of a direct natriuretic action, increased cardiac output, and/or decreased aldosterone levels. In patients hospitalized for decompensated HF, nesiritide improves symptoms and is well tolerated. The major adverse effect is dose-related hypotension. Nesiritide is thus an attractive new vasodilator that should be valuable in the treatment of patients hospitalized for acute decompensated HF. PMID- 11264557 TI - Health Check: helping patients make healthy choices. PMID- 11264556 TI - What's up doc? PMID- 11264558 TI - The price of life. PMID- 11264559 TI - Previous exposure to Chlamydia pneumoniae, Helicobacter pylori and other infections in Canadian patients with ischemic heart disease. AB - OBJECTIVE: Previous exposures to Chlamydia pneumoniae (CP), Helicobacter pylori (HP) or cytomegalovirus (CMV) have been associated with atherosclerotic heart disease. These associations were studied in Canadian patients, and the exposure to five infections measured. DESIGN: Case-control study. SETTING: In the coronary care units (Hamilton General site, Henderson General site, McMaster University Medical Centre site of the Hamilton Health Sciences Corporation and St Joseph's Hospital) and from the regional angiography suite (Hamilton General site), Hamilton, Ontario. PATIENTS AND METHODS: One hundred seven consecutive patients presenting with myocardial infarction or unstable angina (coronary care unit patients), or with previous angina or myocardial infarction (angiography suite patients), were compared with 107 family practice or outpatient clinic control subjects. INTERVENTIONS: Cardiovascular risk factors were measured, as was serology for CP, HP, CMV, adenovirus and hepatitis A virus. Statistical analysis was by logistic regression, adjusted for age and sex. RESULTS: Exposure to CP was more frequent in patients than in control subjects (85.4% versus 70.3%, adjusted odds ratio [OR] 2.3, 95% CI 1.1 to 5.1, P=0.03). Dividing CP immunoglobulin G absorbance into quarters, with the lowest quarter as the reference group, the adjusted ORs were 2.8, 3.0 and 4.3, respectively, for the second, third and fourth quarters (P=0.001 for trend). The seroprevalences of HP (61.7%), CMV (64.0%), adenovirus (75.6%) and hepatitis A virus (59.2%) were high, with no association with disease. CONCLUSIONS: An association was found between heart disease and previous exposure to CP, with a stepwise increase in ORs at higher antibody levels, whereas no association was found with HP, CMV or other infections. A prospective validation of this association is needed. PMID- 11264560 TI - Comparison of midazolam with or without fentanyl for conscious sedation and hemodynamics in coronary angiography. AB - OBJECTIVE: To compare the hemodynamic and sedative effects of midazolam - with or without fentanyl combination - with placebo in coronary angiography. DESIGN: Prospective, double-blind, randomized study. SETTING: University medical centre. PATIENTS: All patients undergoing coronary angiography. INTERVENTIONS: Demographic data, hemodynamic variables, sedation and anxiety scores, amnesia, patient and cardiologist satisfaction, and adverse effects were evaluated and compared among coronary angiography patients taking midazolam, midazolam and fentanyl, or placebo before the procedure. MAIN RESULTS: Ninety patients scheduled for coronary angiography were randomly assigned into three groups: a midazolam-placebo group (group MP), a midazolam-fentanyl group (group MF) and a placebo group (group P). Hemodynamic stability was better in each sedation group (groups MP and MF) than in group P. Sedation scores, anxiolysis, and patient and cardiologist satisfaction were not different between the sedation groups. CONCLUSIONS: Both techniques of conscious sedation - midazolam and midazolam with fentanyl - are satisfactory for coronary angiography where hemodynamic stability and patient cooperation are required. In such procedures, local anesthesia without sedation may lead to hypertension and increase overall morbidity. PMID- 11264561 TI - Correlation of remote ST segment depression and coronary anatomy during acute coronary occlusion. AB - BACKGROUND: The appearance of remote ST segment depression (RSTD) on an electrocardiogram (ECG) is associated with more extensive infarction and a worse clinical outcome than when RSTD is absent. OBJECTIVE: To determine whether RSTD predicts coronary anatomy during acute coronary occlusion. It was hypothesized that RSTD is associated with the occlusion of a proximal lesion, an extensive artery and an artery without distal collateralization. PATIENTS AND METHODS: In 113 consecutive patients with single vessel disease undergoing percutaneous transluminal coronary angioplasty (PTCA), 12-lead ECGs (recorded at baseline and during balloon inflation) and angiographical data were analyzed independently. Patients with ST segment elevation in the primary territory and RSTD (greater than 1 mm ST depression at 80 ms after the J point) (group A) were compared with patients without RSTD (group B). Proximal lesions were defined as lesions located in the segments proximal to the acute marginal branch, first diagonal artery or first obtuse marginal branch. An extensive right coronary artery (RCA) was one that supplied the posterolateral wall; an extensive left anterior descending (LAD) artery was one that supplied the inferoapical wall; and an extensive circumflex artery was one that supplied the posterior descending artery. RESULTS: Fifty-four patients (48%) had PTCA of the proximal vessels, 43 patients (38%) had extensive target vessels and 11 patients (9.7%) had collaterals. Target vessels included 33% in RCA, 44% in LAD artery and 23% in circumflex artery. Forty-five patients (40%) developed RSTD during balloon inflation (group A). Patients in group A were more likely to have extensive vessels on the angiogram than those in group B (group A 49%, group B 31%; P=0.05). None of the patients in group A had collaterals to the culprit artery, while 16% of patients in group B did (P=0.003). The two groups were not significantly different with respect to the number of proximal lesions (group A 58%, group B 42%; P=0.08). Analysis performed according to the target artery revealed that RSTD was associated with occlusion of an extensive RCA during RCA occlusion (extensive RCA in group A 100%, group B 57%; P=0.006). For the LAD artery, RSTD was associated with proximal lesions (group A 74%, group B 41%; P=0.02) and absence of collaterals (group A 100%, group B 74%; P=0.01). CONCLUSIONS: During acute coronary occlusion, the presence of RSTD on 12-lead ECG was specific for the absence of collaterals. The presence of RSTD during RCA occlusion was strongly associated with an extensive RCA, suggestive of posterolateral wall ischemia. During LAD artery occlusion, the presence of RSTD was associated with proximal occlusion, which resulted in ischemia of the LAD artery and the major diagonal artery territories. PMID- 11264562 TI - Predicting and explaining cardiac rehabilitation attendance. AB - BACKGROUND: A variety of factors influence patients' health behaviour; these are patterns of practitioner practice, patient characteristics and availability of resources. OBJECTIVES: To examine patient-related factors (demographic, health, psychosocial characteristics) that may influence patients' attendance at cardiac rehabilitation programs and their subsequent behaviour change. PATIENTS AND METHODS: A prospective cohort design was used. Three hundred four acute myocardial infarction and/or coronary artery bypass graft surgery patients from a tertiary care centre in a Western Canadian city were enrolled to participate in telephone interviews at two weeks and again at approximately six months after their hospital discharge. Measures of self-efficacy and behaviour performance for cardiac health maintenance and role resumption, motivation and social support were used at both interview times. A survey focusing on factors influencing patients' choices to attend cardiac rehabilitation programs was also administered at the interview six months after discharge. RESULTS: Attendance at cardiac rehabilitation programs is not associated with patients' risk factor status, and elderly and rural-living patients are at particular risk for nonattendance. CONCLUSIONS: Systematic mechanisms to guide the appropriate referral of patients to this health care resource and administer secondary prevention initiatives to those with limited access to resources need to be a priority in cardiovascular health care. PMID- 11264563 TI - Prosthetic valve fungal endocarditis due to histoplasmosis. AB - Fungal endocarditis is associated with severe patient morbidity and mortality. Unfortunately, fungal endocarditis is difficult to diagnose because fungal pathogens are uncommonly isolated from routine blood cultures. Histopathological examination of surgically excised cardiac valves, peripheral emboli and systemic ulcers may be useful in identifying pathogens as etiological agents of culture negative endocarditis. The authors describe a 63-year-old man who had culture negative endocarditis. Multiple echocardiograms showed progression of the vegetations with valve stenosis despite treatment with multiple antimicrobials. He had multiple peripheral emboli before surgery. Disseminated histoplasmosis was diagnosed by bone marrow culture. Yeast organisms consistent with histoplasma were shown in the vegetations of his excised mitral valve prosthesis. The patient was treated with amphotericin and has been doing well in the two years since his surgery. The diagnosis and management of fungal endocarditis are emphasized. PMID- 11264564 TI - A new method for quantification of spatial and temporal parameters of endocardial motion: evaluation of experimental infarction using magnetic resonance imaging. AB - BACKGROUND: With the development of high-resolution myocardial imaging there has evolved a need for automated techniques that can accurately quantify regional function. OBJECTIVE: To develop a new method for quantification of spatial and temporal parameters of endocardial motion. DESIGN: Magnetic resonance images were analyzed using a unique, shape-based approach that tracks endocardial surface motion at defined points through the cardiac cycle by minimizing the bending energy. SETTING: Animal instrumentation was performed in the Nuclear Cardiology Experimental Research Laboratory at Yale University, New Haven, Connecticut. Magnetic resonance imaging was performed at the Yale New Haven Hospital Center. ANIMALS: Eight mongrel canines were used. INTERVENTIONS: Electrocardiograph-gated gradient-echo magnetic resonance images were obtained before and after occlusion of the left anterior descending coronary. Thirty-two points along automatically defined endocardial contours were tracked. Average displacements and cumulative path lengths were computed from end-diastole for each point over the entire cardiac cycle. The average cumulative path length was computed for each of four quarters of systole for the normal, border and infarct zones. Shape-based parameters of systolic motion were compared with the centreline approach. Infarct zone was defined by postmortem histochemical staining. MAIN RESULTS: Displacement and cumulative path length over the cardiac cycle decreased significantly in the infarct and border zones (P<0.05), but did not change in the normal zone (P was not significant). Temporal changes in motion were observed in all zones. Displacement measured using the shape-based algorithm was more consistent than cumulative path length when compared with systolic motion measured using the centreline method. CONCLUSIONS: An automated, shape-based approach permits quantitative evaluation of both spatial and temporal parameters of regional endocardial motion from high-resolution electrocardiograph-gated images. Analysis of endocardial motion and cumulative motion over the entire cardiac cycle discriminated infarcted from normal and border regions. PMID- 11264565 TI - Entry sites for coronary angiography and therapeutic interventions: from the proximal to the distal radial artery. AB - When coronary angiography was introduced at the Montreal Heart Institute, Montreal, Quebec, in 1964, a cutdown arteriotomy of the proximal radial artery 2 to 3 cm distal to its origin was selected as the standard access site used instead of the brachial artery approach described by Mason Sones Jr. The risk of symptomatic local thrombosis requiring surgical care appeared less at that site on the basis of collateral circulation to the hand by the ulnar and palmar arches. Attempts to replace the time-consuming cutdown arteriotomy by a percutaneous transarterial approach led successively to the axillary, femoral and brachial arteries and finally, over 20 years later, to the radial artery. The transradial approach for diagnostic angiography, first reported in 1989 and adapted by others several years later for angioplasty, is now used in over 44 countries. The only contraindication for this approach is the rare, inadequate ulnar artery collateral circulation to the hand, clinically recognized by an abnormal Allen test. The transradial approach is advantageous in that there is an easier and safer postprocedural hemostasis at the entry site, it is preferred by patients and ambulation within hours is compatible with an outpatient procedure. PMID- 11264566 TI - Myxoid leiomyosarcoma of the left atrium: a rare malignancy of the heart and its comparison with atrial myxoma. AB - Primary tumours of the heart are rare. The majority of these tumours are benign, with myxomas located in the left atrium being the most common form. Almost all malignant tumours are sarcomas and occur preferentially in the right side of the heart. An exception to this rule is leiomyosarcoma, a rare form of primary cardiac sarcoma that occurs predominantly in the left atrium, as does cardiac myxoma. The case of a 53-year-old woman who presented with symptoms of mitral valve stenosis and pulmonary hypertension is reported. Cardiac catheterization, angiography and echocardiography revealed a left atrial mass that was interpreted as atrial myxoma. At the time of operation, the myxoid appearance of the tumour mass further supported this assumption. The tumour, including a wide rim of atrial septum, was removed with cautery. Histopathological examination unexpectedly showed that the tumour was not an atrial myxoma but rather a myxoid variant of a primary leiomyosarcoma. Immunohistochemistry and electron microscopy confirmed the diagnosis. Local radiotherapy was considered but deemed contraindicated in view of the longstanding pulmonary hypertension. Two months after excision, a repeat echocardiogram indicated recurrence of tumour in the left atrium, and the patient died a few days later. The preferential left atrial location and the frequently myxoid appearance of primary leiomyosarcomas of the heart make it particularly difficult to differentiate them preoperatively from atrial myxomas. The authors recommend resection of all atrial myxoid tumours with a wide (at least 1 cm) margin, combined with intraoperative frozen section diagnosis, because complete surgical resection appears to correlate with prolonged survival in the few reported cases of atrial leiomyosarcomas. In cases of incomplete initial resection or local recurrence in the absence of metastatic disease, heart transplantation may be a valid option in appropriately selected patients. PMID- 11264567 TI - The clinical pathological conference. PMID- 11264568 TI - Difficult decisions regarding percutaneous endoscopic gastrostomy placement. PMID- 11264569 TI - Hepatocyte growth factor in patients with three different stages of chronic liver disease including hepatocellular carcinoma, cirrhosis and chronic hepatitis: an immunohistochemical study. AB - BACKGROUND AND AIMS: The specific role of hepatocyte growth factor in liver disease is unknown. The presence and density of this factor in patients with three different stages of liver disease were investigated, with the aim of assessing its prognostic significance. PATIENTS AND METHODS: Liver specimens from patients with chronic hepatitis (n=20), cirrhosis (n=20), hepatocellular carcinoma (n=30) and normal livers (n=20) were immunohistochemically stained to determine the presence and density of hepatocyte growth factor. RESULTS: There were significantly more hepatocyte growth factor-positive Kupffer and Ito cells in all three diseased groups than in the control group. Also, there was significantly more positive staining in chronic hepatitis specimens than in specimens from the cirrhosis, hepatocellular carcinoma and control groups (P<0.05). The hepatoma cells in 10 of the hepatocellular carcinoma cases stained positive, but none of the hepatocytes in the chronic hepatitis, cirrhosis and normal liver specimens stained. It was only possible to assess nonmalignant hepatocytes adjacent to the hepatocellular carcinoma in the four resection specimens, and no staining for hepatocyte growth factor was observed in these areas. There was no statistical association between density of hepatocyte growth factor and histological activity index in chronic hepatitis, or between density of hepatocyte growth factor and grade of hepatocellular carcinoma. CONCLUSIONS: Similar to some previous reports, this study revealed that hepatoma cells can also express this growth factor. Immunohistochemical detection of hepatocyte growth factor may prove to be a useful method of diagnosing hepatocellular carcinoma in challenging cases. PMID- 11264570 TI - The relevance of apoptosis for cellular homeostasis and tumorigenesis in the intestine. AB - Intestinal epithelium is a rapidly renewing tissue in which cell homeostasis is regulated by a balance among proliferation, growth arrest, differentiation and apoptosis (programmed cell death). Until recently, studies on oncogenesis have focused on the regulation of cell proliferation. The recognition that apoptosis must be understood to comprehend how appropriate cell numbers are maintained and how alterations in any part of the equation can contribute to malignancy has led to an explosion of research in this field. The first half of this review gives an overview of morphology and mechanisms of apoptosis, emphasizing key areas of genetic control such as the bcl-2 family and p53. The second half of the review focuses on the role of apoptosis in normal cellular homeostasis and tumorigenesis in the gastrointestinal epithelium. The importance of understanding the molecular biology of apoptotic pathways in cancer therapy and future directions are also addressed. PMID- 11264571 TI - Serological and molecular testing in viral hepatitis: an update. AB - The routine serological diagnoses of the three major forms of viral hepatitis - A, B and C - as well as delta hepatitis, are important in the evaluation of acute and chronic viral hepatitis. Increasingly, molecular virology is also being used to evaluate patients with chronic hepatitis C, with genotype and viral load testing to plan therapy. PMID- 11264572 TI - Liver fibrosis and altered matrix synthesis. AB - Liver fibrosis represents the uniform response of liver to toxic, infectious or metabolic agents. The process leading to liver fibrosis resembles the process of wound healing, including the three phases following tissue injury: inflammation, synthesis of collagenous and noncollagenous extracellular matrix components, and tissue remodelling (scar formation). While a single liver tissue injury can be followed by an almost complete restitution ad integrum, the persistence of the original damaging noxa results in tissue damage. During the establishment of liver fibrosis, the basement membrane components collagen type IV, entactin and laminin increase and form a basement membrane-like structure within the space of Disse. The number of endothelial fenestrae of the sinusoids decreases. These changes of the sinusoids are called 'capillarization' because the altered structure of the sinusoids resembles that of capillaries. At the cellular level, origin of liver fibrogenesis is initiated by the damage of hepatocytes, resulting in the recruitment of inflammatory cells and platelets, and activation of Kupffer cells, with subsequent release of cytokines and growth factors. The hepatic stellate cells seem to be the primary target cells for these inflammatory stimuli, because during fibrogenesis, they undergo an activation process to a myofibroblast-like cell, which represents the major matrix-producing cell. Based on this pathophysiological mechanism, therapeutic methods are developed to inhibit matrix synthesis or stimulate matrix degradation. A number of substances are currently being tested that either neutralize fibrogenic stimuli and prevent the activation of hepatic stellate cells, or directly modulate the matrix metabolism. However, until now, the elimination of the hepatotoxins has been the sole therapeutic concept available for the treatment of liver fibrogenesis in humans. PMID- 11264573 TI - Pneumatic dilation in achalasia. AB - Pneumatic dilation is the most common first-line therapy for the treatment of achalasia. The aim of dilation is a controlled disruption of circular muscle fibres of the lower esophageal sphincter to reduce the functional obstruction. Several types of dilators and different dilation techniques are used, but the achieved results are similar. The mean success rate is about 80% in the short term, but some patients need redilation in the further course (particularly young patients). Best long term results are obtained if the lower esophageal sphincter pressure can be reduced below 10 mmHg. Major complications are rare after pneumatic dilation; the most serious complication is esophageal perforation, which occurs at a mean rate of about 2.5%. Considering the pros and cons of other effective forms of treatment of achalasia (esophagomyotomy and intrasphincteric injection of botulinum toxin), pneumatic dilation is still the treatment of choice in the majority of patients with achalasia. PMID- 11264574 TI - Ulcerative colitis of the appendix ('ulcerative appendicitis') mimicking acute appendicitis. AB - Appendiceal involvement in ulcerative colitis may occur in the setting of either diffuse or distal disease, and is usually diagnosed incidentally at the time of proctocolectomy. The present patient had a rare case of 'ulcerative appendicitis' occurring on a background of clinically quiescent ulcerative colitis, and presented with the signs and symptoms of acute appendicitis. PMID- 11264575 TI - To save crystallization data. PMID- 11264576 TI - Structures of the B1 domain of protein L from Peptostreptococcus magnus with a tyrosine to tryptophan substitution. AB - The three-dimensional structure of a tryptophan-containing variant of the IgG binding B1 domain of protein L has been solved in two crystal forms to 1.7 and 1.8 A resolution. In one of the crystal forms, the entire N-terminal histidine tag region was immobilized through the coordination of zinc ions and its structural conformation along with the zinc coordination scheme were determined. However, the ordering of the histidine tag by zinc does not affect the overall structure of the rest of the protein. Structural comparisons of the tryptophan containing variant with an NMR-derived wild-type structure, which contains a tyrosine at position 47, reveals a common fold, although the overall backbone root-mean-square difference is 1.5 A. The Y47W substitution only caused local rearrangement of several side chains, the most prominent of which is the rotation of the Tyr34 side chain, resulting in a 6 A displacement of its hydroxyl group. A small methyl-sized cavity bounded by beta-strands 1, 2 and 4 and the alpha-helix was found in the structures of the Y47W-substituted protein L B1 domain. This cavity may be created as the result of subsequent side-chain rearrangements caused by the Y47W substitution. These high-resolution structures of the tryptophan-containing variant provide a reference frame for the analysis of thermodynamic and kinetic data derived from a series of mutational studies of the protein L B1 domain. PMID- 11264577 TI - Atomic resolution structures of trypsin provide insight into structural radiation damage. AB - Radiation damage is an inherent problem in protein X-ray crystallography and the process has recently been shown to be highly specific, exhibiting features such as cleavage of disulfide bonds, decarboxylation of acidic residues, increase in atomic B factors and increase in unit-cell volume. Reported here are two trypsin structures at atomic resolution (1.00 and 0.95 A), the data for which were collected at a third-generation synchrotron (ESRF) at two different beamlines. Both trypsin structures exhibit broken disulfide bonds; in particular, the bond from Cys191 to Cys220 is very sensitive to synchrotron radiation. The data set collected at the most intense beamline (ID14-EH4) shows increased structural radiation damage in terms of lower occupancies for cysteine residues, more breakage in the six disulfide bonds and more alternate conformations. It appears that high intensity and not only the total X-ray dose is most harmful to protein crystals. PMID- 11264578 TI - Three-dimensional structure of human RNase 1 delta N7 at 1.9 A resolution. AB - Human pancreatic ribonuclease 1 (RNase 1) is considered to be the human counterpart of bovine pancreatic RNase A. Truncation of seven amino-acid residues from the amino-terminal sequence resulted in RNase 1 Delta N7, which has a reduced ribonucleolytic activity and a lower affinity for the human placental RNase inhibitor (PRI). This RNase 1 variant has been cloned, heterologously overexpressed, purified and crystallized. Its crystal structure has been determined and refined using data to 1.9 A resolution. The molecule displays the alpha + beta folding topology typical of members of the RNase A superfamily. The main distinct features found in RNase 1 Delta N7 are basically located in three loops affecting the fitting of the enzyme to the active site of subtilisin and the shape of the B2 subsite. These changes, taken with the lack of the catalytically active residue Lys7, may explain the reduced affinity of RNase 1 Delta N7 for PRI and the low ribonucleolytic activity of the protein when compared with the native enzyme. PMID- 11264579 TI - The structure of human mitochondrial branched-chain aminotransferase. AB - X-ray crystal structures of three forms of human mitochondrial branched-chain aminotransferase (BCAT) were solved by molecular-replacement methods, using Escherichia coli BCAT as the search model. The enzyme is a homodimer and the polypeptide chain of each monomer has two domains. The small domain is composed of residues 1--175 and the large domain is composed of residues 176--365. The active site is close to the dimer interface. The 4'-aldehyde of the PLP cofactor is covalently linked to the epsilon-amino group of the active-site lysine, Lys202, via a Schiff-base linkage in two of the structures. In the third structure, the enzyme is irreversibly inactivated by Tris. The overall fold of the dimer in human mitochondrial BCAT is similar to the structure of two bacterial enzymes, E. coli BCAT and D-amino acid aminotransferase (D-AAT). The residues lining the putative substrate-binding pocket of human BCAT and D-AAT are completely rearranged to allow catalysis with substrates of opposite stereochemistry. In the case of human mitochondrial branched-chain aminotransferase, a hydrogen-bond interaction between the guanidinium group of Arg143 in the first monomer with the side-chain hydroxyl of Tyr70 in the second monomer is important in the formation of the substrate-binding pocket. PMID- 11264580 TI - X-ray structure of bovine pancreatic phospholipase A2 at atomic resolution. AB - Using synchrotron radiation and a CCD camera, X-ray data have been collected from wild-type bovine pancreatic phospholipase A(2) at 100 K to 0.97 A resolution allowing full anisotropic refinement. The final model has a conventional R factor of 9.44% for all reflections, with a mean standard uncertainty for the positional parameters of 0.031 A as calculated from inversion of the full positional least squares matrix. At 0.97 A resolution, bovine pancreatic phospholipase A(2) reveals for the first time that its rigid scaffolding does not preclude flexibility, which probably plays an important role in the catalytic process. Functionally important regions (the interfacial binding site and calcium-binding loop) are located at the molecular surface, where conformational variability is more pronounced. A cluster of 2-methyl-2,4-pentanediol molecules is present at the entrance of the hydrophobic channel that leads to the catalytic site and mimics the fatty-acid chains of a substrate analogue. Bovine pancreatic phospholipase A(2) at atomic resolution is compared with previous crystallographic structures and with models derived from nuclear magnetic resonance studies. Given the high structural similarity among extracellular phospholipases A(2) observed so far at lower resolution, the results arising from this structural analysis are expected to be of general validity for this class of enzymes. PMID- 11264581 TI - Structures and comparison of the Y98H (2.0 A) and Y98W (1.5 A) mutants of flavodoxin (Desulfovibrio vulgaris). AB - The structures for two mutants at the Tyr98 site of Desulfovibrio vulgaris flavodoxin have been determined. The first, a tyrosine-to-histidine (Y98H) variant, was determined at the moderately high resolution of 2.0 A, while the tyrosine-to-tryptophan variant (Y98W) yielded very high resolution data (beyond 1.5 A) allowing a detailed look at the water structure, alternate side-chain conformations and the planarity of the FMN. Both structures were solved by molecular replacement beginning with the native (P2A) coordinates as a starting point. The Y98H variant of D. vulgaris flavodoxin crystallizes in space group P2(1)2(1)2(1), with unit-cell parameters a = 41.96, b = 61.45, c = 57.04 A, while the Y98W mutant adopts space group P2(1), with a = 41.29, b = 55.82, c = 32.52 A, beta = 100.68 degrees. Refinement for both mutants utilized PROLSQ followed by, for the high-resolution Y98W structure, anisotropic refinement as implemented in SHELXL. Final R factors of 17% for the Y98H mutant and 9.8% for the Y98W mutant were obtained. For the high-resolution (1.5 A) Y98W mutant, 31,010 unique reflections were collected from a single crystal. The final model includes 273 solvent molecules, with eight side chains assuming multiple conformations. At this resolution, the detailed conformation of the FMN can be observed, with both a bow and twist being noted. A comparison is made between the two mutants and the different oxidation states of the native flavodoxin. Although both mutants show similar E(2) (oxidized/semiquinone) one-electron redox potentials to the native, the E(1) (semiquinone/hydroquinone) redox potential for the Y98H mutant is significantly different from that of the Y98W variant and the native protein. The surprising similarity in the folding of the polypeptide chain 60--64 between the two mutants and the reduced states of the native is discussed. The interaction between O61 and N5 in the flavin is discussed because of the new conformation of this loop. PMID- 11264582 TI - Identification of a small-molecule binding site at the dimer interface of the HIV integrase catalytic domain. AB - Integration of the reverse-transcribed HIV cDNA into the host DNA is a required step in viral replication. The virus-encoded integrase protein catalyzes the initial DNA breaking and joining reactions that mediate cDNA integration. Here, the identification by X-ray crystallography of a small-molecule binding site on the integrase catalytic domain is reported. The small-molecule family studied consists of a core of arsenic or phosphorus surrounded by four aromatic groups. Two arsenic derivatives were visualized bound to integrase. In each case, two molecules bound at symmetry-related sites on the catalytic domain dimer interface. The first compound studied, tetraphenyl arsonium, did not inhibit integrase. However, a synthetic compound substituting a catechol for one of the phenyl rings, dihydroxyphenyltriphenylarsonium, bound to the same site and did inhibit the enzyme. Changes in the vicinity of the catalytic site were seen with the inhibitory compound only, potentially explaining its mechanism of action. Further substituting phosphonium for arsonium yielded a compound with an IC(50) in the low micromolar range. These findings may be useful in designing new inhibitors of integrase, which is at present the only one of the three HIV enzymes for which clinically useful inhibitors are not available. PMID- 11264583 TI - Structure of the C-terminal sterile alpha-motif (SAM) domain of human p73 alpha. AB - p73 is a homologue of the tumour suppressor p53 and contains all three functional domains of p53. The alpha-splice variant of p73 (p73 alpha) contains near its C terminus an additional structural domain known as the sterile alpha-motif (SAM) that is probably responsible for regulating p53-like functions of p73. Here, the 2.54 A resolution crystal structure of this protein domain is reported. The crystal structure and the published solution structure have the same five-helix bundle fold that is characteristic of all SAM-domain structures, with an overall r.m.s.d. of 1.5 A for main-chain atoms. The hydrophobic core residues are well conserved, yet some large local differences are observed. The crystal structure reveals a dimeric organization, with the interface residues forming a mini four helix bundle. However, analysis of solvation free energies and the surface area buried upon dimer formation indicated that this arrangement is more likely to be an effect of crystal packing rather than reflecting a physiological state. This is consistent with the solution structure being a monomer. The p73 alpha SAM domain also contains several interesting structural features: a Cys-X-X-Cys motif, a 3(10)-helix and a loop that have elevated B factors, and short tight inter-helical loops including two beta-turns; these elements are probably important in the normal function of this domain. PMID- 11264584 TI - Growth kinetics, diffraction properties and effect of agarose on the stability of a novel crystal form of Thermus thermophilus aspartyl-tRNA synthetase-1. AB - Growth kinetics and diffraction properties of monoclinic crystals of eubacterial Thermus thermophilus aspartyl-tRNA synthetase-1 (AspRS-1) prepared in the presence of polyethylene glycol and agarose are studied. Their solubility and two dimensional phase diagram are compared with those of orthorhombic crystals which grow in the presence of sodium formate or ammonium sulfate. The growth mechanism of the novel crystals was monitored by atomic force microscopy. The gel stabilizes the crystal lattice under the cryogenic conditions used for structure determination at high resolution. PMID- 11264585 TI - Conformational stabilization and crystallization of the SecA translocation ATPase from Bacillus subtilis. AB - SecA is the peripheral membrane-associated subunit of the enzyme complex 'preprotein translocase' which assists the selective transport of presecretory proteins into and across bacterial membranes. The SecA protein acts as the molecular motor that drives the translocation of presecretory proteins through the membrane in a stepwise fashion concomitant with large conformational changes accompanying its own membrane insertion/retraction reaction cycle coupled to ATPase activity. The high flexibility of SecA causes a dynamic conformational heterogeneity which presents a barrier to growth of crystals of high diffraction quality. As shown by fluorescence spectroscopy, the T(m) of the endothermic transition of cytosolic SecA from Bacillus subtilis is shifted to higher temperatures in the presence of 30% glycerol, indicating stabilization of the protein in its compact membrane-retracted conformational state. High glycerol concentrations are also necessary to obtain three-dimensional crystals suitable for X-ray diffraction analysis, suggesting that stabilization of the conformational dynamics of SecA may be required for effective crystallization. The SecA crystals grow as hexagonal bipyramids in the trigonal space group P3(1)12; they possess unit-cell parameters a = 130.8, b = 130.8, c = 150.4 A at 100 K and diffract X-rays to approximately 2.70 A using a high-flux synchrotron radiation source. PMID- 11264586 TI - Solvent behaviour in flash-cooled protein crystals at cryogenic temperatures. AB - The solvent behaviour of flash-cooled protein crystals was studied in the range 100--180 K by X-ray diffraction. If the solvent is within large channels it crystallizes at 155 K, as identified by a sharp change in the increase of unit cell volume upon temperature increase. In contrast, if a similar amount of solvent is confined to narrow channels and/or individual cavities it does not crystallize in the studied temperature range. It is concluded that the solvent in large channels behaves similarly to bulk water, whereas when confined to narrow channels it is mainly protein-associated. The analogy with the behaviour of pure bulk water provides circumstantial evidence that only solvent in large channels undergoes a glass transition in the 100--180 K temperature range. These studies reveal that flash-cooled protein crystals are arrested in a metastable state up to at least 155 K, thus providing an upper temperature limit for their storage and handling. The results are pertinent to the development of rational crystal annealing procedures and to the study of temperature-dependent radiation damage to proteins. Furthermore, they suggest an experimental paradigm for studying the correlation between solvent behaviour, protein dynamics and protein function. PMID- 11264587 TI - Map self-validation: a useful discriminator of phase correctness at low resolution. AB - A new map-validation procedure is based on the correlation-coefficient agreement between the observed structure-factor magnitudes and their extrapolated values from suitably modified electron-density maps from which they have been each in turn systematically excluded. The correlation coefficient tends to a maximum as the phase errors in a map are reduced. This principle was used to resolve the single-wavelength anomalous scattering (SAS) and single-derivative isomorphous replacement (SIR) phase ambiguity for a number of error-free trial structures. Applications employing real data sets tend to be more difficult owing to data incompleteness and errors affecting the construction of the Argand diagram. PMID- 11264588 TI - Crystallization of Clonorchis sinensis 26 kDa glutathione S-transferase and its fusion proteins with peptides of different lengths. AB - A Clonorchis sinensis 26 kDa glutathione S-transferase (CsGST) and its fusion proteins containing 14 and 48 amino-acid peptides at the N-terminus have been crystallized using polyethylene glycol monomethylether 550 as a precipitant. Crystals of the three proteins show very similar crystal properties: they diffract to at least 2.3 A resolution and belong to the orthorhombic space group P2(1)2(1)2(1). The unit-cell parameters of CsGST crystals were a = 66.64 (1), b = 68.91 (1), c = 123.41 (2) A, which are very close to those of the crystals of the two fusion proteins. In addition, CsGST fusion proteins containing varying extents of N-terminal-extended peptides are incorporated into a crystal, indicating that the extended peptides have little effect on crystal packing. These results suggest that the crystallization system of CsGST/peptide fusion protein may be generally applicable to obtain crystals of small peptides. PMID- 11264590 TI - Expression, purification and preliminary crystallographic studies of human ketohexokinase. AB - Ketohexokinase (KHK; E.C. 2.7.1.3) catalyses the (reversible) phosphorylation of fructose to fructose-1-phosphate. KHK is the first enzyme in a specialized catabolic pathway metabolizing dietary fructose to the glycolytic intermediate glyceraldehyde-3-phosphate. Mutations inactivating KHK underlie the metabolic disorder essential fructosuria. The primary structure of KHK shows no significant homology to other mammalian hexokinases. It is most similar to prokaryotic ribokinases, but catalyses a distinct phosphorylation reaction. Recombinant human KHK has been crystallized in the orthorhombic form (space group P2(1)2(1)2 or P2(1)2(1)2(1)). Single crystals of this polymorph suitable for X-ray diffraction have been obtained by vapour diffusion using 2-propanol and MPD as precipitants (pH 7.5). The crystals have unit-cell parameters a = 93.4, b = 121.5, c = 108.4 A. Diffraction data were collected to 4.3 A resolution. The asymmetric unit contains four protein molecules. PMID- 11264589 TI - Crystallization and preliminary X-ray analysis of dmpFG-encoded 4-hydroxy-2 ketovalerate aldolase--aldehyde dehydrogenase (acylating) from Pseudomonas sp. strain CF600. AB - The final two steps of the meta-cleavage pathway for catechol degradation in Pseudomonas sp. strain CF600 involve the conversion of 4-hydroxy-2-ketovalerate to pyruvate and acetyl coenzyme A by the enzymes 4-hydroxy-2-ketovalerate aldolase and NAD(+)-dependent acylating aldehyde dehydrogenase. Biochemical studies indicate that these two enzymes comprise a bifunctional heterodimer (DmpFG, molecular mass 71 kDa) and suggest that the product of the aldolase reaction is transferred to the dehydrogenase active site via a channeling mechanism. Crystals of the DmpFG complex grow in multiple fan-like clusters of thin plates by the hanging-drop method and are improved by streak-seeding. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 102.0, b = 140.7, c = 191.3 A, and diffract to 2.1 A resolution. The asymmetric unit contains four DmpFG heterodimers. Heavy-atom derivative screening identified three isomorphous derivatives. PMID- 11264591 TI - Crystallization and preliminary X-ray crystallographic analysis of the human type 3 3 alpha-hydroxysteroid dehydrogenase at 1.8 A resolution. AB - In androgen-sensitive target tissues, 3 alpha-hydroxysteroid dehydrogenase regulates the androgen receptor (AR) activity by catalyzing the inactivation of 5 alpha-dihydrotestosterone (the most natural potent androgen) to 5 alpha androstane-3 alpha,17 beta-diol. In this report, the crystallization of a human prostatic type 3 3 alpha-hydroxysteroid dehydrogenase, a member of the aldo-keto reductase superfamily, is described. Two different crystal forms of the complex between the human type 3 3 alpha-HSD, NADP(+) and testosterone have been obtained using PEG as precipitant. Crystal form I, which diffracts to 1.6 A, belongs to the monoclinic space group P2(1), with unit-cell parameters a = 55.07, b = 87.15, c = 76.88 A, beta = 107.37 degrees and two subunits in the asymmetric unit. A complete data set has been collected at 1.8 A. Crystal form II, which diffracts to 2.6 A, belongs to the rhombohedral space group R32, with unit-cell parameters a = b = 143.59, c = 205.86 A, alpha = beta = 90, gamma = 120 degrees and two subunits in the asymmetric unit. PMID- 11264592 TI - Human phosphoglucose isomerase: expression, purification, crystallization and preliminary crystallographic analysis. AB - Phosphoglucose isomerase (PGI) is the second enzyme in the glycolytic pathway and catalyzes an aldose-ketose isomerization. Outside the cell, PGI has been found to function as both a cytokine and as a growth factor. The human pgi gene was cloned and the expressed enzyme was purified to homogeneity. Isomorphous crystals were obtained under two conditions and belong to the P2(1)2(1)2(1) space group, with unit-cell parameters a = 80.37, b = 107.54, c = 270.33 A. A 94.7% complete data set was obtained and processed to a limiting resolution of 2.6 A. The asymmetric unit contains two hPGI dimers according to density calculations, a self-rotation function map and molecular-replacement solution. PMID- 11264593 TI - Structural studies on the cobra venom factor: isolation, purification, crystallization and preliminary crystallographic analysis. AB - Cobra venom factor (CVF) is the complement-activating protein in cobra venom. It is a three-chain glycoprotein with a molecular weight of 149,000 Da. In serum, CVF forms a bimolecular enzyme with the Bb subunit of factor B. The enzyme cleaves C3 and C5, causing complement consumption in human and mammalian serum. CVF is frequently used to decomplement serum to investigate the biological functions of complement and serves as a tool to investigate the multifunctionality of C3. Furthermore, CVF bears the potential for clinical application to deplete complement in situations where complement activation is involved in the pathogenesis of disease. CVF was isolated from Indian cobra (Naja naja naja) venom. The protein was crystallized at room temperature using the sitting-drop vapour-diffusion technique. The crystals diffract to 2.7 A resolution and belong to the tetragonal space group P4(1), with unit-cell parameters a = b = 62.7, c = 368.1 A. PMID- 11264594 TI - Crystallization and preliminary crystallographic studies of a phospholipase A2 from the venom of the Brazilian snake Bothrops moojeni. AB - A phospholipase A(2) purified from the venom of the snake Bothrops moojeni has been crystallized by vapour-diffusion techniques in hanging drops at 291 K. The crystals, which were grown in the absence of Ca(2+), belong to the cubic system, space group P432, with unit-cell parameters a = b = c = 91.86 A, and contain one molecule in the asymmetric unit (V(M) = 2.71 A(3) Da(-1)). X-ray diffraction experiments provide data to 2.35 A resolution collected on a rotating-anode home source at cryogenic temperatures. The structure has been solved via molecular replacement techniques using a single monomer of the crystallographic structure of the phospholipase from the Western diamondback rattlesnake (Crotalus atrox) as a search model. PMID- 11264595 TI - Preliminary crystallographic studies of EcTI, a serine proteinase inhibitor from Enterolobium contortisiliquum seeds. AB - Enterolobium contortisiliquum trypsin inhibitor (EcTI) belongs to the Kunitz family of plant inhibitors, which are widely distributed in nature, especially in plant seeds. EcTI is composed of two polypeptide chains with a total of 174 residues, homologous to other inhibitors from the same family. EcTI crystals, which were obtained with the acupuncture-gel technique, diffract to 2.0 A resolution and belong to space group P2(1), with unit-cell parameters a = 37.12, b = 38.42, c = 54.08 A, beta = 98.08 degrees. Molecular-replacement techniques using Erythrina caffra trypsin inhibitor (PDB code 1tie) as the search model indicate one monomer in the asymmetric unit. The secondary-structure content of EcTI was determined by circular dichroism spectroscopy, yielding values compatible with the expected topology. PMID- 11264596 TI - Crystallization and functional analysis of a soluble deglycosylated form of the human costimulatory molecule B7-1. AB - The interactions of B7-1 with CD28 and CTLA-4 modulate the course of human immune responses, making B7-1 an important target for developing structure-based therapeutics. B7-1 is, however, one of the most heavily glycosylated proteins found at the leukocyte cell surface, complicating the structural analysis of this molecule. Methods for the production, crystallization and selenomethionine labelling of a soluble deglycosylated form of this molecule are described. The protein readily forms both tetragonal plate and bipyramidal crystals belonging to space groups I4(1)22, with unit-cell parameters a = b = 56.9, c = 298.7 A, and P4(1)22 (or P4(3)22), with unit-cell parameters a = b = 89.0, c = 261.9 A, respectively. The I4(1)22 and primitive crystal forms diffract to 2.7 and 3.5 A, respectively. Surface plasmon resonance-based assays indicate that the ligand binding properties of sB7-1 are unaffected by deglycosylation. Since none of the methods relied on any special structural properties of sB7-1, it is proposed that this novel combination of procedures could in principle be adapted to the systematic analysis of many other glycoproteins of structural or functional interest. PMID- 11264597 TI - The crystals of a mannose-specific jacalin-related lectin from Morus nigra are merohedrally twinned. AB - MornigaM, a lectin from Morus nigra, belongs to the mannose-binding subgroup of the family of jacalin-related plant lectins. It was crystallized in the P6(5) space group, with unit-cell parameters a = b = 110.74, c = 159.28 A. The partially merohedrally twinned crystals could be detwinned and a subsequent molecular-replacement solution could be found using the coordinates of jacalin. Preliminary analysis clearly shows the tetrameric assembly of this protein. Furthermore, data from MornigaM crystals soaked in a mannose solution were collected. PMID- 11264598 TI - Crystallization and preliminary X-ray crystallographic analysis of the surE protein from Thermotoga maritima. AB - The surE protein from Thermotoga maritima is a 247-residue protein of unknown function. Its homologues are well conserved among both the eubacteria and the archaea. It has been overexpressed in soluble form in Escherichia coli. The protein has been crystallized at 296 K using 2-propanol as a precipitant. X-ray diffraction data have been collected to 1.9 A resolution using synchrotron radiation. The crystals belong to the trigonal space group P3(1)21 (or P3(2)21), with unit-cell parameters a = b = 115.96, c = 78.60 A, alpha = beta = 90, gamma = 120 degrees. The asymmetric unit contains two monomers of the surE protein, with a corresponding V(M) of 2.72 A(3) Da(-1) and a solvent content of 54.7%. PMID- 11264599 TI - Purification, crystallization and preliminary X-ray studies of thermostable alkaline phosphatase from Thermus sp. 3041. AB - Thermostable alkaline phosphatase from Thermus sp. 3041 has been expressed in Escherichia coli, purified and crystallized. The crystals belong to space group P2(1)22(1), with unit-cell parameters a = 57.7, b = 69.9, c = 111.5 A. Diffraction data were collected to 2.54 A with a completeness of 91.1% (87.8% for the last shell), an R(merge) value of 0.105 (0.312) and an I/sigma(I) value of 9.5 (3.6). PMID- 11264601 TI - Twin pregnancy: the role of ultrasound in management. AB - Determination of chorionicity is one of the most important issues in the management of twin pregnancy. Modern ultrasound equipment has made it possible to accurately assess placentation already in the first trimester with the lambda sign. With regard to prenatal diagnosis, it is important to know the chorionicity in order to calculate the risk of chromosomally abnormal fetuses. Accurate chorionicity offers the obstetricians the opportunity to observe the monochorionic twins more intensively than is required for twins with dichorionic placentation. This review gives an update of the state of the art for clinicians caring for twin pregnancies. PMID- 11264600 TI - Crystallization and preliminary X-ray diffraction studies of the guanylate kinase like domain of PSD-95 protein from rat. AB - The PSD-95 (postsynaptic density-95) protein, one of the members of the MAGUK (membrane-associated guanylate kinase) family, is composed of three PDZ domains, one SH3 domain and one guanylate kinase-like (GK) domain. The GK domain mediates the scaffolding function of PSD-95 by protein--protein interaction. Here, the GK domain was subcloned, expressed as an intein fusion protein, purified without the intein and then crystallized at room temperature by the hanging-drop vapour diffusion method using PEG 8000 as a precipitant. The complete native data set was collected to a resolution of 2.35 A using flash-cooling. The crystals belong to the primitive tetragonal space group P4(3) (or P4(1)), with unit-cell parameters a = b = 70.03 (4), c = 37.64 (1) A. PMID- 11264602 TI - Screening for Chlamydia trachomatis in asymptomatic women in Hungary. An epidemiological and cost-effectiveness analysis. AB - BACKGROUND: A multicenter survey was carried out in order to determine the prevalence and risk factors for Chlamydia trachomatis infection in the population of asymptomatic women in Hungary. Results were used to carry out a cost effectiveness analysis of screening for chlamydial infection in women with asymptomatic genital infections. METHODS: The non-amplified nucleic acid hybridization method (PACE 2 Gen-Probe) was used to diagnose C. trachomatis and Bayes' theorem was applied to assess the prevalence of the infection. Multiple logistic regression analysis was performed to differentiate the risk factors for chlamydial infections. RESULTS: According to the test, the prevalence of Chlamydia trachomatis among 1300 pregnant women was 4.5%. The sensitivity and specificity of the test are estimated to be 70% and 99%, respectively. After Bayes' correction, the overall estimated prevalence of chlamydial infection was 5.1%. There were significant differences in proportions of chlamydial infection in different regions, and also in different age groups and different family status groups. The highest rate was for women aged below 20 years: 16.9%. Cost effectiveness analysis, with associated sensitivity analysis was carried out for women aged below 20 years. Three screening strategies were compared: using the ELISA method, using amplified Gen-Probe method and no screening. The amplified Gen-Probe method was best provided, the infection prevalence exceeded 16.7%, the PID rate exceeded 24% and the probability of tubal infertility in untreated women exceeded 25%. CONCLUSION: We conclude that screening with amplified Gen-Probe assays (followed by treatment of positive patients) is the preferred screening strategy for young women in Hungary. PMID- 11264603 TI - Effects of umbilical arterial resistance on its arterial blood flow velocity waveforms. AB - BACKGROUND: We propose to clarify the effect of umbilical arterial resistance on umbilical arterial blood flow patterns in normal pregnant women. SUBJECTS: Pulsed Doppler examination was administered to 145 pregnant women of between 28 and 31 weeks of normal gestation without any prenatal complications. METHODS: The arterial flow velocity waveforms from two different arteries, one in the intra abdominal portion and the other at the placental insertion, were sampled with the color flow mapping method, and the peak systolic velocity and the resistance index were calculated. RESULTS: The peak systolic velocity and resistance index on the placental side were significantly lower than those of the fetal side. CONCLUSION: A major difference exists between the umbilical arterial flow velocity waveforms on the fetal and placental sides, which might be caused by difference in the umbilical arterial resistance. PMID- 11264604 TI - Warm tub bathing during labor: maternal and neonatal effects. AB - AIM: To study possible detrimental maternal and neonatal effects of immersion in warm water during labor. DESIGN: Prospective randomized controlled bathing during first stage of labor vs no bathing. SETTING: Obstetrical departments at a university hospital and two central hospitals. PRIMARY END-POINT: Referral of newborns to NICU. MATERIAL AND METHODS: Randomization took place by means of sealed opaque envelopes at each delivery unit. Preconditions for participation in the study were: singleton parturient wishing to bathe, a gestational duration of at least 35 weeks+0 days, a planned vaginal delivery, normal admission test, regular contractions and cervix dilated to at least 3-4 cm. Parturients randomized to the 'no bath' control group were allowed to use a shower. Rupture of the membranes was not a contra-indication to participation. Those excluded from randomization were women with intra-uterine growth retardation, meconium stained amniotic fluid, or in the event that the tub was occupied by another randomized parturient. MAIN RESULTS: On average, parturients stayed in the tub for 50-60 min. No significant difference was seen regarding the referral rate to NICU among 612 cases vs 625 controls, OR 0.8; 95% CL 0.2, 3.1. The OR for epidural analgesia was 1.0; 95% CL 0.8, 1.3. Nor was any significant difference seen in the rate of perineal tear grade III-IV (OR 1.3), instrumental delivery (OR 1.1), cesarean section (OR 1.8), or maternal post partum stay on the ward. During the neonatal period, no significant difference was seen in the number of newborns with Apgar <7 at 5 min (4 vs 5), neonatal distress (OR 2.2) or tachypnea (OR 1.0). CONCLUSION: In the present study no negative effects of bathing during labor could be discerned. The results indicate that expectant mothers wishing to bathe during labor may do so without jeopardizing their own, or their newborns' wellbeing after birth. PMID- 11264606 TI - Socio-economic, demographic and obstetric risk factors for late fetal death of unknown etiology in Lithuania: a case--referent study. AB - BACKGROUND: The purpose of this study was to evaluate the association between third trimester unexplained prelabor fetal deaths and various socio-economic, demographic and obstetric factors in Lithuania. METHODS: A case-referent study on 58 women with third trimester fetal death and 116 women with live fetus at term was carried out. Inclusion criteria for women in the first group (cases) were: prelabor fetal death of unknown etiology, singleton pregnancy >26 weeks of gestation and intact fetal membranes. For each case two referent women were recruited, admitted during the same period in active phase of labor at term (>37 weeks of gestation) with intact fetal membranes and fetus alive. Data were obtained by interview, anthropometry and by reviewing the medical records. Several potential socio-economic, demographic and obstetrical risk factors for unexplained fetal death were investigated. RESULTS: Univariate analyses determined several factors that were associated with fetal death of unknown etiology: low educational level, single marital status, low income, etc. After secondary logistic regression analysis only three independent variables remained significantly associated with otherwise unexplained stillbirth: small for gestational age fetus (OR 29.6; 95% CI 6.2-141.6), low income (OR 7.4; 95% CI 3.1 17.6), and maternal white blood cell count more than 16,000/mm3 (OR 5.4; 95% CI 1.4-21.6). Body mass index, smoking, occupation of women and other evaluated parameters were not confirmed to be significant risk factors. CONCLUSION: Small for gestational age fetus, low income and elevated maternal white blood cell count are factors significantly associated with late prelabor fetal death in Lithuania. PMID- 11264605 TI - Fear during labor. AB - BACKGROUND: The aims of the present study were to compare primiparous and multiparous women's experiences of fear of delivery during an early stage of active labor (cervix dilatation 3-5 centimeters) and to study whether fear of delivery, measured during the early stage of active labor, was a predictor of the amount of pain relief received during the remaining part of labor (cervix dilatation 5 cm - partus), of the duration of the remaining part of labor, and of the occurrence of instrumental vaginal delivery and emergency cesarean section. METHOD: Thirty-five primiparous and 39 multiparous women answered the Delivery Fear Scale (DFS) once during the early stage of labor and before they had received any pain relief. RESULTS: Primiparous women reported higher levels of fear than multiparous women did. Fear during the first phase of labor predicted only the total amount of pain relief received during labor. CONCLUSION: The clinical implications of the study are that the delivery staff should consider women's fear during labor and pay attention especially to primiparous women's increased risk of higher levels of fear during an early stage of active labor, as compared with multiparous women's. The challenge for staff of a delivery ward is to support the woman in labor in a way that decreases fear, which in turn might reduce the woman's need of pain relief. PMID- 11264607 TI - Correlation between serum testosterone levels and peripartal mood states. AB - BACKGROUND: We conducted a prospective study at the Department of Obstetrics and Gynecology, University Hospital of Vienna to investigate associations between serum testosterone levels and maternal peripartal mood states. METHODS: Two hundred and fifty-two pregnant women at term (38 to 40 weeks' gestation) took part in the study. Blood samples for plasma testosterone levels and other biochemicals were obtained prepartum, and on the 1st and 3rd day postpartum. Mood was assessed with the McNair Profile of Mood States (POMS) at term pregnancy and daily from the first day after delivery until discharge from the hospital. RESULTS: The final study population consisted of 193 women. Serum testosterone levels correlated significantly with maternal depression scores, both pre- and post partum (at term r=0.148, p=0.04; 1st day postpartum r=0.156, p=0.03; and 2nd day postpartum r=0.186, p=0.02, respectively). Testosterone concentrations also correlated with anger prepartum (r=0.164, p=0.02) and on the third day after delivery (r=0.188, p=0.02). No significant correlation between testosterone concentration and fatigue and vigor both pre- and post partum, respectively were found. CONCLUSION: Serum testosterone levels correlate with depression and anger in the first postpartum days. PMID- 11264608 TI - Preimplantation genetic diagnosis (PGD): the Gothenburg experience. AB - BACKGROUND: A program for preimplantation genetic diagnosis of pre-embryos from patients with hereditary disorders was set up in our unit at Sahlgrenska University Hospital in 1994. The majority of the patients were carriers of X chromosome linked disorders; a few patients were translocation carriers. In this paper we describe our experiences of our first 36 cycles, 30 gender determinations and six analyses of embryos with possible translocations. METHODS: Conventional hormone replacement treatment with intracytoplasmic sperm injection to fertilize the eggs followed by blastomere biopsy and fluorescent in situ hybridization at the eight cell stage was used for sexing as well as detection of translocations. RESULTS: Out of the 30 cycles in 13 patients for gender determination, blastomere biopsies could be carried out in 25 cycles. Transfer of normal female embryos (XX) was performed in 18 cycles, resulting in five pregnancies (pregnancy rate 27.8%) and an implantation rate of 20% per transfer. Three girls have been born. Hence the take home baby rate was 16.7% per transfer and 10% per started cycle. Six cycles (three patients) for detection of translocations in embryos were performed. Diagnosis was possible in four cycles. Transfer of normal embryos was carried out in one cycle. No pregnancy was achieved. CONCLUSION: Successful PGD in its clinical application demands close collaboration between a large group of specialists. Even so, the success rate is considerably lower than after conventional IVF or ICSI procedures. Taking into account the stress caused to the parents facing late interruption of pregnancy following conventional prenatal diagnosis we are convinced that this technique is well worthwhile continuing and refining. PMID- 11264609 TI - Recovery from vaginal hysterectomy compared with laparoscopy-assisted vaginal hysterectomy: a prospective, randomized, multicenter study. AB - OBJECTIVE: To evaluate short-term recovery of vaginal hysterectomy with those of laparoscopic assisted vaginal hysterectomy performed in a prospective, randomized multicentric study. STUDY DESIGN: Eighty patients referred for hysterectomy for benign pathology were randomized to either vaginal hysterectomy (40 patients) or laparoscopic assisted vaginal hysterectomy (40 patients). Inclusion criteria were uterine size larger than 280 g and one or more of the following: previous pelvic surgery, history of pelvic inflammatory disease, moderate or severe endometriosis, concomitant adnexal masses, and indication for adnexectomy. No upper limit of uterine size was set. All the laparoscopic and the vaginal hysterectomies were done under endotracheal general anesthesia. RESULTS: There was no statistically significant difference in terms of patient's age, parity, postmenopausal state, indication for surgery and mean uterine weight between the 2 groups. Laparoconversion was performed in three women in the laparoscopic group. Operative time was significantly shorter in the vaginal versus the laparoscopic groups 108+/-35 and 160+/-50 respectively (p<0.001). The use of paracetamol, non steroidal anti-inflammatory drugs, and opioid during hospitalization were similar in the 2 groups. There was no difference in the 1st day hemoglobin level drop, time of passing gas and stool, or hospital stay between the 2 groups. CONCLUSION: In contrast with earlier reports, there was no difference in short-term recovery between patients undergoing vaginal or laparoscopic hysterectomy. No advantage was found performing laparoscopic assisted vaginal hysterectomy in comparison with the standard vaginal hysterectomy. PMID- 11264610 TI - Intrauterine insemination with donor semen. An evaluation of prognostic factors based on a review of 1131 cycles. AB - OBJECTIVE: To identify prognostic factors influencing the outcome of infertility treatment using intrauterine insemination with donor semen (IUI-D). DESIGN: Retrospective study of all patients undergoing IUI-D between August 1st, 1990 and July 31st, 1998. SETTING: University-affiliated infertility clinic. PATIENTS: Three hundred and five couples undergoing 1131 IUI-D treatment cycles. MAIN OUTCOME MEASURES: Type of hormonal treatment, number of follicles, length of follicular phase, endometrial pattern, female age, infertility diagnosis and semen quality related to clinical pregnancy rate, cumulative birth rate and multiple gestations. RESULTS: Throughout the nine year period the overall clinical pregnancy rate per cycle was 22.3%, with an increase from 12.9% in 1990 to 34.6% in 1998. The multiple birth rate was 20.6%. The birth rate per couple was 61.1% after a mean of 3.2 treatment cycles. The pregnancy rate was highest in the first treatment cycle and the cumulative birth rate rose only slightly after the sixth treatment cycle. The following parameters were positively and significantly correlated to a successful outcome of IUI-D: i) the first treatment cycle - compared to the following up to six treatment cycles; ii) number of mature follicles - more than one - at the time of insemination, however, with an unacceptable high rate of multiple pregnancies when more than 3 mature follicles were present; iii) time of insemination after the 12th day in the cycle; iv) insemination after ovulation has occurred and; v) female age under 30 years. CONCLUSIONS: IUI-D is a simple and inexpensive treatment giving acceptable pregnancy rates for up to six treatment cycles if at least 2 mature follicles have developed at the time of insemination, which implies that hormonal ovarian stimulation and induction of ovulation is used, and ovulation has occurred at the time of insemination, which ought to take place after cycle day (cd) 12 with at least two million motile spermatozoa. PMID- 11264611 TI - Laparoscopically assisted vaginal management of deep endometriosis infiltrating the rectovaginal septum. AB - BACKGROUND: Two aims: 1) To assess the results of laparoscopically assisted vaginal management of deep endometriosis infiltrating the rectovaginal septum (RVS); 2) to pinpoint the differences between this procedure and that used for deep endometriotic lesions located on the uterosacral ligaments (USL). METHODS: Descriptive retrospective study. Twenty-nine consecutive patients operated for deep endometriosis infiltrating the RVS were included in this series. RESULTS: One patient only (3.5%) presented a major complication of the recto-vaginal fistula type. After a one step reoperation under anesthesia, the post operative history was uncomplicated and no sequelae are to be deplored. With respect to dysmenorrhea (DM), deep dyspareunia (DP) and chronic pelvic pain (CPP), there was an improvement in respectively 91.7% (22 patients), 100% (24 patients) and 92.9% (13 patients) of cases. For each of these 3 symptoms the median score according to the visual analog scale was significantly lower after the operation (for DM: 7.6+/-2.0 versus 1.7+/-2.6; for DP 7.5+/-1.9 versus 0.5+/-1.1; for CPP 5.9+/-2.8 versus 1.4+/-3.2) (p<0.0001). CONCLUSIONS: These results demonstrate that provided the surgeon is highly skilled in laparoscopy, operative laparoscopy is efficient for the treatment of patients presenting painful symptoms related to deep endometriotic infiltrating the RVS. From the technical point of view the rectum must be freed, leaving the deep endometriotic nodule attached to the posterior wall of the vagina. Resection of the whole lesion requires the posterior wall of the vagina to be resected, whereas ureterolysis is often unnecessary. So for lesions located on the RVS the vagina is opened systematically, unlike the situation when resecting deep endometriotic lesions infiltrating the USL. Deep pelvic endometriosis is not synonymous with endometriosis of the RVS. Lesions truly infiltrating the RVS represent only a small proportion of all deep endometriosis lesions. PMID- 11264612 TI - Radiotherapy alone for invasive vaginal cancer: outcome with intracavitary high dose rate brachytherapy versus conventional low dose rate brachytherapy. AB - PURPOSE: Our aim was to compare the role of remote afterloaded high-dose-rate brachytherapy (HDRB) with traditional low-dose-rate brachytherapy (LDRB) for patients with invasive primary vaginal carcinoma. METHODS: The study group comprised 190 patients with invasive carcinoma of the vagina. The patients were staged according to the International Federation of Gynecology and Obstetrics (FIGO) staging system. Eighty patients were treated with intracavitary high-dose rate iridium 192 brachytherapy with or without external beam therapy. These patients are compared with a historical group of 110 patients treated with intracavitary low-dose-rate radium 226 or cesium 137 brachytherapy with or without external beam therapy. RESULTS: No significant differences were found for stages, tumor grade or location between the two groups. Crude 5-year survival for all patients was 41% in the former LDRB group, 81% in stage I and 43% in stage II. Overall actuarial 3-year survival and disease-specific survival rates for all patients in the HDRB series were 51% and 66%, respectively. Disease-specific 3 year survival attained 83% in stage I and 66% in stage II. There were no significant differences in local and distant recurrences between the treatment modalities. The comparison of treatments with or without external beam radiation and of complications showed no significant differences between the HDRB and LDRB series. CONCLUSION: With HDRB and its advantages of decreased radiation exposure and patient immobilization and precise positioning, treatment results to be obtained are at least similar to traditional LDRB for primary vaginal cancer. PMID- 11264613 TI - Peritoneal fluid concentrations of interleukin-4 in relation to the presence of endometriosis, its stage and the phase of the menstrual cycle. AB - BACKGROUND: Interleukin-4 (IL-4) is a cytokine with both stimulatory and inhibitory effects on the inflammatory system such as macrophage inhibition and T cell activation. It is known to regulate several monocyte functions, including inhibition of the synthesis of cytokines such as IL-1, IL-6 and TNF-alpha as well as potentiating IL-8. METHOD: In an attempt to clarify the association between IL 4 and endometriosis, we measured the concentration of IL-4 in the peritoneal fluid of 52 women; 24 with endometriosis and 28 with no endometriosis, controlling for the phase of the cycle and the stage of disease. RESULTS: There was no difference in the concentrations of IL-4 between women with (n=28) and without endometriosis (n=24). No difference was found between the IL-4 concentrations in women with different stages of endometriosis. Levels of IL-4 did not show a difference according to the phase of the cycle in either group. CONCLUSION: Our results indicate no association between peritoneal fluid levels of IL-4 and endometriosis and hence suggest that IL-4 is not involved in the pathogenesis of endometriosis. PMID- 11264614 TI - Tumor cell spillage to the vaginal cavity and vaginal stump during the surgery of endometrial carcinoma. AB - BACKGROUND: Our purpose was to investigate the spillage of the endometrial carcinoma cells to the vaginal cavity and vaginal stump during the surgery. METHODS: Cytologic examination of specimens obtained from the vaginal cavity and vaginal stump during surgery performed on 15 patients with endometrial carcinoma. And the risk factors of carcinoma cell spillage to the vaginal cavity were investigated. RESULTS: Carcinoma cells from the vaginal cavity were negative for nine patients and positive for six patients. Those from the vaginal stump were negative for l4 patients and positive for one patient. In one patient, both were positive. Total abdominal hysterectomy might be one risk of carcinoma cell spillage to the vaginal cavity compared with modified radical hysterectomy and radical hysterectomy. CONCLUSIONS: Endometrial carcinoma cells were spilled into the vaginal cavity in not a few cases and their spillage from there to the vaginal stump could follow during the surgery. PMID- 11264615 TI - A woman with isolated prolactin deficiency. PMID- 11264617 TI - Chromatometra or chromatocolpos--technical considerations in the management of obstructed mullerian duct anomalies. PMID- 11264616 TI - Osteoclast-like giant cells in leiomyomatous tumors of the uterus. A case report and review of the literature. AB - Osteoclast-like giant cells (OLGC) in leiomyomatous tumors of the uterus are rarely seen, and their significance is unknown. We present a case of a large leiomyomatous tumor in which OLGC were found in only few sections showing leiomyosarcoma whereas the majority of sections revealed a leiomyoma. Though radically operated, the patient died a few months later with recurrent tumor in the pelvis and metastases to the lungs. PMID- 11264618 TI - A potential cause of hyperemesis gravidarum: evidence from the sex ratio of associated infants. PMID- 11264620 TI - The associated offspring sex ratios and cause(s) of hyperemesis gravidarum. PMID- 11264621 TI - Making childbirth a normal process. PMID- 11264622 TI - Maternal analgesia during labor disturbs newborn behavior: effects on breastfeeding, temperature, and crying. AB - BACKGROUND: Newborns not exposed to analgesia, when placed on the mother's chest, exhibit an inborn prefeeding behavior. This study was performed to assess the effects of different types of analgesia during labor on the development of spontaneous breastfeeding movements, crying behavior, and skin temperature during the first hours of life in healthy term newborns. METHODS: Video recordings were made of 28 newborns who had been dried and placed in skin-to-skin contact between their mother's breasts immediately after delivery. The video recordings were analyzed blindly with respect to infant exposure to analgesia. Defined infant behaviors were assessed every 30 seconds. Group 1 mothers (n = 10) had received no analgesia during labor, group 2 mothers (n = 6) had received mepivacaine via pudendal block, and group 3 mothers (n = 12) had received pethidine or bupivacaine or more than one type of analgesia during labor. RESULTS: All infants made finger and hand movements, but the infant's massagelike hand movements were less frequent in infants whose mothers had received labor analgesia. A significantly lower proportion of group 3 infants made hand-to-mouth movements (p < 0.001), and a significantly lower proportion of the infants in groups 2 and 3 touched the nipple with their hands before suckling (p < 0.01), made licking movements (p < 0.01), and sucked the breast (p < 0.01). Nearly one-half of the infants, all in groups 2 or 3, did not breastfeed within the first 2.5 hour of life. The infants whose mothers had received analgesia during labor had higher temperatures (p = 0.03) and they cried more (p = 0.05) than infants whose mothers had not received any analgesia. CONCLUSIONS: The present data indicate that several types of analgesia given to the mother during labor may interfere with the newborn's spontaneous breast-seeking and breastfeeding behaviors and increase the newborn's temperature and crying. PMID- 11264623 TI - Postpartum maternal oxytocin release by newborns: effects of infant hand massage and sucking. AB - BACKGROUND: Newborns placed skin-to-skin with their mothers show an inborn sequence of behavior similar to that seen in other mammals. The purpose of this study was to make a detailed exploration of hand movements and sucking behavior in healthy term newborns who were placed skin-to-skin on their mothers' chests, and to study maternal oxytocin release in relation to these behaviors. METHODS: Ten vaginally delivered infants whose mothers had not been exposed to maternal analgesia were video-recorded from birth until the first breastfeeding. Video protocols were developed based on observations of the videotapes. Each infant's hand, finger, mouth, and tongue movements, positions of the hand and body, and sucking behavior were assessed every 30 seconds. Maternal blood samples were collected every 15 minutes, and oxytocin levels were analyzed by radioimmunoassay. A statistical test for establishing the relationship between maternal oxytocin levels and infants' hand movements or sucking behavior was developed. RESULTS: Infants used their hands to explore and stimulate their mother's breast in preparation for the first breastfeeding. A coordinated pattern of infant hand and sucking movements was also identified. When the infants were sucking, the massagelike hand movements stopped and started again when the infants made a sucking pause. Periods of increased massagelike hand movements or sucking of the mother's breast were followed by an increase in maternal oxytocin levels (p < 0.005). CONCLUSIONS: The findings indicate that the newborns use their hands as well as their mouths to stimulate maternal oxytocin release after birth, which may have significance for uterine contraction, milk ejection, and mother-infant interaction. PMID- 11264624 TI - What babies teach us: the essential link between baby's behavior and mother's biology. PMID- 11264626 TI - Perinatal depression: a randomized controlled trial of an antenatal education intervention for primiparas. AB - BACKGROUND: Depression can be an unexpected and distressing companion for a woman during the major life transition of becoming a mother for the first time. Researchers now demonstrate that approximately 50 percent of women will experience perinatal distress. Therefore, the etiology and management of perinatal depression is essential for a quality care of childbearing women. The objectives of this study were to develop an education intervention tailored to the information needs of primiparous women about perinatal depression, to deliver this intervention antenatally, and to conduct a randomized controlled trial to determine the effect of the antenatal education intervention in the reduction of postnatal depression. METHOD: A prospective, randomized controlled trial of the education intervention (n = 206) was conducted at three sites in Australia. The outcome of changes in mood state was measured by the Profile of Mood States questionnaire once antenatally (12-28 wk), and twice postnatally (8-12 and 16-24 wk); social support and demographic data were also collected. The education package was administered to the intervention group at the antenatal assessment of mood. RESULTS: A significant and steady reduction in scores (overall and on the subscales) was observed over time for both groups that showed significant improvement in symptoms of depression. No difference was detected when comparing the intervention group with the control group. Additional multivariate regression analyses revealed no relevant influence of social support or demographic variables. CONCLUSIONS: Women in both the study and control groups were more depressed antenatally than postnatally. The finding that the education intervention made no difference challenges the two strongly held tenets of health education in childbearing women-that depression can be reduced through education and that antenatal education interventions can endure into the postnatal period. PMID- 11264625 TI - Reducing postnatal pain from perineal tears by using lignocaine gel: a double blind randomized trial. AB - BACKGROUND: Perineal pain is one of the most common causes of maternal morbidity in the early puerperium. Several randomized trials have shown that topical application of local anesthetics is effective in reducing postepisiotomy pain, but no randomized study has assessed the efficacy of local anesthetics for other perineal trauma. This study investigated if topically applied 2 percent lignocaine gel was an effective treatment for this group of women. METHODS: A double-blind placebo controlled trial was conducted in a regional teaching hospital in the northwest of England. One hundred and forty-nine women who had sustained a first- or second-degree tear were allocated by sealed envelopes to the lignocaine gel or placebo group. The primary outcome was self-reported pain at 24 hours postdelivery as measured on a numerical rating scale (pain score). Secondary outcomes included pain scores at 48 hours, the need for oral analgesia, and maternal satisfaction. Based on a pilot study, we calculated that 128 women were required to detect a 25 percent difference in pain scores between the two groups with 80 percent power (alpha = 0.05). The pain scores of women in each trial arm were compared using the unpaired t test and 95 percent confidence intervals. RESULTS: Women using lignocaine gel had lower average pain scores, although this only reached statistical significance at 48 hours after delivery (p = 0.023). In general, women liked using the study gel. No difference was found in consumption of oral analgesia. CONCLUSIONS: This study suggested that lignocaine gel may be effective on the second postnatal day. Further research is required to assess the optimum timing of this intervention and the population that would most benefit from its use. PMID- 11264627 TI - Implementing change: becoming baby-friendly in an inner city hospital. AB - The Baby-Friendly Hospital Initiative of the United Nations Children's Fund and the World Health Organization dramatically raises breastfeeding rates when implemented. To date, only 27 of the 16,000 Baby-Friendly hospitals worldwide are located in the United States. Barriers to becoming Baby-Friendly in the United States include the strength of the infant formula industry, suboptimal clinician knowledge, and the need to implement significant change throughout an institution. This paper describes how Boston Medical Center, an inner-city teaching hospital in Boston with approximately 1800 births per year, overcame numerous obstacles and, in December 1999, became the first Baby-Friendly hospital in Massachusetts. PMID- 11264628 TI - Effective care in pregnancy and childbirth: a synopsis. PMID- 11264629 TI - Pregnant women's perceptions of their nurse's role during labor and delivery. AB - BACKGROUND: Little has been studied about pregnant women's perceptions of their nurse's role during labor and delivery. The objective of this study was to determine nulliparous pregnant women's expectations of their nurse's role during labor and delivery as expressed during the last trimester of pregnancy. METHOD: Nulliparous women in childbirth classes were asked on a questionnaire, "What do you think your nurse's role will be during labor and delivery? You may list as many things as you wish." RESULTS: Fifty-seven completed surveys were collected. The women listed a total of 174 items. Approximately 29 percent of the nursing tasks listed by the nulliparous women were related to providing them with physical comfort and emotional support, 24 percent related to providing informational support, almost 21 percent were related to providing technical nursing care, and 21 percent related to monitoring of the baby, mother, or labor progress; approximately 5 percent related to indirect care (outside the room). CONCLUSION: The expectations of women in our study were in contrast with findings from two previous work sampling studies, in which nurses provided much less time giving women physical comfort, emotional support, and informational support than would have been expected by women in our study. Fulfilling women's expectations about childbirth can increase women's satisfaction with their birth experiences. Further studies can help maternity caregivers learn more about women's expectations. PMID- 11264631 TI - Sheila Kitzinger's and Jenny Kitzinger's letter from Europe: childbirth and breastfeeding in the British media. PMID- 11264630 TI - Preemies on steroids: a new iatrogenic disaster? PMID- 11264632 TI - Cytological monitoring of peripheral blood, bronchoalveolar lavage fluid, and transbronchial biopsy specimens during acute rejection and cytomegalovirus infection in lung and heart--lung allograft recipients. AB - STUDY OBJECTIVES: Acute rejection and cytomegalovirus (CMV) infection are important complications after lung and heart--lung transplantation. We sought to investigate whether acute rejection and CMV infection demonstrated as CMV antigenemia had an effect on the cell profiles of peripheral blood (PB), bronchoalveolar lavage fluid (BAL-F), or TBB histology. PATIENTS AND DESIGN: In this prospective study, composition of cells in PB, BAL-F, and TBB samples from 20 lung or heart-lung transplantation patients were analyzed during episodes of acute rejection or CMV antigenemia. Rejection was graded according to the International Society for Heart and Lung Transplantation criteria. As controls, samples with no evidence of rejection or infection were used. To evaluate the effect of time on cellular findings, samples were divided into three groups according to time after transplantation: 1--30, 31--180, and more than 180 d after transplantation. RESULTS: Acute rejection was associated with mild blood basophilia (p<0.05; specificity 94%, sensitivity 42%). In BAL-F during rejection, the number of basophils (p<0.05), eosinophils (p<0.05), and lymphocytes (p<0.05; specificity 77%, sensitivity 64%) was increased compared to controls during the post-operative month 1. Later-occurring rejections were associated with increased amounts of neutrophils in BAL-F (p<0.05; specificity 82%, sensitivity 74%). In TBB histology, acute rejections were associated with perivascular and/or peribronchial infiltration of lymphocytes (p<0.001) and plasma cells (p<0.05) compared to controls. In our patients receiving gancyclovir prophylaxis, CMV antigenemia did not significantly alter the cell profiles in PB and BAL-F nor the inflammatory cell picture in TBB histology. CONCLUSION: TBB histology remains the 'gold standard' for diagnosing rejection in lung and heart-lung transplantation patients, as the inflammatory cell findings in TBB specimens are highly specific for rejection. The cellular changes associated with rejection, mild PB basophilia and increased proportions of lymphocytes in early- and neutrophils in later occurring rejection, observed in BAL-F cannot be considered specific for rejection, but may warrant clinical suspicion of rejection. PMID- 11264633 TI - Influence of diabetes mellitus on patient and graft survival in recipients of kidney transplantation. AB - AIMS: To investigate the outcomes in patients who have pre-existing diabetes and those who develop post-transplant diabetes mellitus (PTDM). METHODS: We retrospectively reviewed the charts of 939 patients who received a first functioning renal transplant in the cyclosporine (CyA) era between 1984 and 1999. RESULTS: Sixty-six (7%) patients had renal failure due to insulin-dependent diabetes mellitus (IDDM) and 7 (0.8%) patients due to non-insulin-dependent diabetes mellitus (NIDDM). Ten (1.1%) patients had coexistent diabetes and 48 (5.1%) recipients developed PTDM. The mean graft survival for the patients with PTDM was 9.7 yr versus 11.3 yr for the non-diabetic patients, while mean graft survival was 10.1 yr for patients with IDDM and 2.9 yr with NIDDM and 8.3 yr for those with coexistent diabetes (p=ns). However, there was a statistically significant difference in patient survival between patients who developed PTDM and in those who did not develop this complication. The mean survivals of patients with IDDM, NIDDM, coexistent diabetics and PTDM were 8.4, 3.7, 8.6 and 10.3 yr, respectively. The mean survival of the patients without pre-existing diabetes or PTDM was 12.8 yr (p<0.001). The survival of patients older than 55 yr with PTDM was no different to the control group. However, in those younger than 55 yr, PTDM was associated with a higher risk of death (relative risk of 2.54, p<0.001). Fifty percent of patients with IDDM developed acute rejection episodes, whereas rejection rate was 57.1% in NIDDM group, 50.0% in the PTDM group, 20.0% in the coexistent diabetes group and 44.3% in the control group (p=ns). CONCLUSION: Patient survival, but not graft survival, was adversely affected by both pre-existing diabetes and by PTDM, particularly in those with an age less than 55 yr. PMID- 11264634 TI - Clotrimazole increases tacrolimus blood levels: a drug interaction in kidney transplant patients. AB - In order to substantiate a previous case report of a drug interaction between tacrolimus and clotrimazole, we randomly assigned tacrolimus-treated renal allograft recipients to therapy with either clotrimazole or nystatin for oral thrush prophylaxis immediately following transplantation. Patients receiving other agents known to interact with cytochrome P450 were excluded from the study. The clotrimazole group consisted of 17 patients and the nystatin group, which served as the control group, consisted of 18 patients. An oral loading dose (approximately 0.3 mg/kg) of tacrolimus was given pre-operatively. Post transplant, tacrolimus (approximately 0.15 mg/kg) was orally administered twice daily. Clotrimazole therapy consisted of a 10-mg troche administered three times daily. Nystatin therapy consisted of the oral suspension (5 mL) administered as a 'swish and swallow' four times daily. We evaluated tacrolimus trough blood levels and tacrolimus doses on days 1, 3, 5, and 7 following transplantation. On post transplant day 1, mean tacrolimus trough levels did not differ between clotrimazole- and nystatin-treated patients. Mean tacrolimus blood trough levels were significantly higher in clotrimazole-treated patients on days 3, 5, and 7 post-transplant, 42+/-14, 53+/-7, and 33+/-17 ng/mL, respectively, compared to 15+/-8, 15+/-7, and 14+/-6 ng/mL in nystatin-treated patients (p<0.05). The mean tacrolimus dose was significantly lower in the clotrimazole group by day 7 post transplant (p<0.05). We conclude that clotrimazole therapy may cause a significant rise in tacrolimus trough blood levels. Recognition of this potential drug interaction is essential to minimize tacrolimus-associated toxicities in the early post-transplant period. PMID- 11264635 TI - Eurotransplant Senior Program 'old for old': results from 10 patients. AB - More frequently there is the need for renal transplantation of older patients. Against the background of an increasing number of old donors and recipients, Eurotransplant Leiden started the Eurotransplant Senior Program (ESP) 'old for old' in 1999. The ESP works with donors and recipients both over 65 yr. The kidneys are transplanted with short cold ischaemia time regardless of the human leukocyte antigen (HLA) compatibility. Compatibility of blood groups, negative crossmatch and less than 5% cytotoxic antibodies are required. First experiences from 10 patients at Heinrich Heine University hospital are reported here. The course of 10 transplanted patients is described from January 1999 until November 1999 (28.4+/-15.8 wk). Age of donor and recipient, cause of dialysis and concomitant diseases from recipients, function of the transplanted kidney and complications are analysed. Immunosuppression consisted initially of cyclosporin A, mycophenolic acid and steroids. The results of these 10 patients were compared to 14 patients who were transplanted according to the ordinary Eurotransplant criteria (Eurotransplant Kidney Allocation System) in the same period of time. Kidneys from six donors (70.5+/-3.3 yr) were transplanted to 10 different recipients (66.9+/-2.2 yr). The control group consisted of 14 patients (47.6+/ 14.4 yr) who received kidneys from 14 donors (48.3+/-10.1 yr). One double kidney transplantation was performed in the senior group, i.e. two kidneys from a marginal donor were transplanted to one recipient ('two in one'). In the ESP group, cold ischaemia time was reduced by 5 h and mean of HLA mismatches was more than doubled. Mean length of hospitalisation of ESP and control groups was 47.2+/ 28.2 and 34.2+/-11.6 d, respectively. Intraoperatively, no complications were seen, post-operative care was performed on a normal ward. ESP patients suffered more often from delayed graft function, which led to further need for haemodialysis for 11.2 d. Finally, 9 of 10 patients acquired a satisfactory renal graft function. A total of 13 biopsies were performed in eight cases. Altogether seven acute rejections in 6 patients were found (four interstitial, one vascular, one interstitial+vascular, one clinical). The 9 patients with sufficient renal graft function were discharged with a mean serum creatinine level of 2.3+/-0.5 mg/dL (control: 1.9+/-0.8 mg/dL). Comparing these 10 recipients to a control group consisting of 14 patients, the results are comparable and encouraging. In conclusion, the short-term results of the ESP are promising. Nevertheless, the post-operative care requires more attention due to several complications. Though the HLA compatibility was not considered, all rejections were coped with effectively. Quality of life was improved. PMID- 11264636 TI - Inferior liver allograft survival from cadaveric donors >50 years of age? AB - The growing imbalance between the number of cadaveric organ donors and recipients has led to an increasing use of high-risk donors as an option to expand the donor pool. The aim of this study was to evaluate our experience with the use of older liver (donor>50 yr of age) allografts. The medical records, postreperfusion biopsies and laboratory results were reviewed of the 393 patients who underwent orthotopic liver transplantation between 1986 and 1997. The outcome of the 61 patients who received older livers (OL) was compared to that of the other 332 recipients. Increasing use of OL was evident from 1992 onwards. Recipients of OL were older than recipients of younger livers (YL, p<0.001) and more commonly had underlying chronic viral hepatitis (CVH) or fulminant hepatic failure (p<0.05). Patient and allograft survival were only slightly less in recipients of OL versus YL (p=NS). Although postperfusion biopsies showed more damage in OL than YL allografts (p<0.05), this was not associated with increased primary graft failure. OL allografts can be transplanted with acceptable results into recipients without the concern of early allograft loss. SUMMARY OF ARTICLE: This report of one centre's experience with 61 recipients of older donor liver allografts identifies recipient factors that may also have a negative impact on allograft outcome. These factors include a diagnosis of either CVH or fulminant hepatic failure at the time of transplantation. Postreperfusion biopsies of older donor allografts tend to show more damage, but this is not associated with primary non-function. PMID- 11264637 TI - Decreased effect of immunosuppression on immunocompetence in African--Americans after kidney and liver transplantation. AB - Several studies indicate that the poorer outcomes for African--Americans after transplantation may be due to decreased effectiveness of immunosuppressive agents. Using an in vitro test of immunocompetence (IMC), we measured the effects of immunosuppression on African-American, compared with Caucasian, kidney or liver transplantation recipients. The IMC result was the highest of three mixed lymphocyte culture responses using validated stimulator cell pools. A total of 293 tests were done in Caucasians and 144 in African--Americans. Overall, the IMC for African--Americans was 38, compared with 19 for Caucasians (p<0.01). This decreased effect of immunosuppression (higher IMC) was the same for liver as for kidney transplant recipients, occurred at the 2--3-yr interval, and was largely in patients of tacrolimus (FK506), with a strong but not significant trend in cyclosporine (CYA) recipients. The two groups were on the same nominal immunosuppression and FK506 and CYA levels were not different. We conclude that African-Americans retain more immune responsiveness on equivalent dose immunosuppression, notable particularly in years 2--3 after transplantation when earlier graft loss occurs in this group. PMID- 11264638 TI - Enhanced cyclosporine-itraconazole interaction with cola in lung transplant recipients. AB - BACKGROUND: Invasive aspergillosis is a major cause of morbidity and mortality in lung transplant recipients (LTR), occurring in up to 15% of patients post transplant. The 14% aspergillus incidence at the Cleveland Clinic Foundation prompted institution of universal prophylaxis with oral itraconazole (ICZ) in 1997. We report our experience with two protocols of ICZ administration in non cystic fibrosis LTR and the interaction with cyclosporine (CSA). METHODS: Group 1 patients (n=12) were administered ICZ capsules in a fasting or fed state, with or without a histamine-2 (H-2) receptor antagonist or proton pump inhibitor. Group 2 patients (n=12) received the same protocol as group I, but in a fed state with a carbonated beverage (cola) to increase acidity in the stomach to enhance absorption of ICZ. The ICZ dose was 200 mg/d, given as one daily dose. A historical control group (n=10) did not receive chemoprophylaxis with ICZ. CSA daily doses, dose intervals, concentration, cost, and random ICZ levels were documented over a 4-month period of time and compared using generalized estimating equations. RESULTS: The daily CSA mg/kg/d dose decreased over time in all three groups, but no differences were found between the three groups. The CSA dosing interval over time was significantly prolonged in group 2 compared to group 1 or the control group (p< or =0.003). Over time, there was no difference in CSA concentration between all groups. There was no difference in cost over time between the three groups; however, the mean cost of CSA therapy was significantly lower in group 2 compared to the control group (p=0.025). Group 2 administered ICZ with cola had greater random blood concentrations of ICZ (p=0.019). CONCLUSIONS: ICZ capsules administered in a fed state with a cola resulted in greater random levels of ICZ, a decrease in cost/d of CSA, and a prolongation of CSA dosing interval. Although daily CSA dosage trended lower in group 2, it did not reach statistical significance. We believe these changes in CSA dosing over time reflect increased absorption of ICZ and recommend verifying ICZ absorption with an itraconazole level, especially when CSA intervals are not prolonged. PMID- 11264639 TI - Differential influence of azathioprine and mycophenolate mofetil on the disposition of basiliximab in renal transplant patients. AB - Pharmacokinetic sampling was performed in two multicenter trials in which basiliximab (anti-CD25 monoclonal antibody) was administered with triple immunosuppression consisting of cyclosporine microemulsion, corticosteroids, and either azathioprine or mycophenolate mofetil. Blood samples were collected over 12 wk post-transplant from 31 azathioprine-treated and 66 mycophenolate mofetil treated patients. Empirical Bayes estimates of each patient's basiliximab disposition parameters were derived and the duration of CD25 saturation was estimated as the time over which serum concentrations exceeded 0.2 microg/mL as confirmed by flow cytometry measurements. Basiliximab clearance was 29+/-14 mL/h when coadministered with azathioprine and 18+/-8 mL/h with mycophenolate mofetil. Both were significantly lower compared with a clearance of 37+/-15 mL/h from a previous study of basiliximab with dual therapy (p<0.001). As a consequence of the lower clearance of basiliximab, the durations of CD25 saturation were prolonged in the presence of azathioprine (50+/-20 d; range, 13--84) and mycophenolate mofetil (59+/-17 d; range, 28--94) compared with dual therapy (36+/ 14 d; range, 12--91). A total of 27 acute rejection episodes occurred during the first 6 months in the two studies. Durations of CD25 saturation were not different in these patients compared with those who remained rejection-free in each study. A single patient among 57 who were screened developed anti-idiotype antibodies to basiliximab. The average duration of CD25 saturation was prolonged by 39 and 64% in the presence of azathioprine and mycophenolate mofetil, respectively. This graded effect was also observed for basiliximab clearance and may be due in part to a differentially reduced humoral response to basiliximab. Nonetheless, the range of CD25 saturation durations and basiliximab clearances did not extend outside the range when basiliximab was used with dual therapy in the absence of these agents. Hence, no dosing adjustment is deemed necessary when basiliximab is used in triple immunosuppressive therapy including either azathioprine or mycophenolate mofetil. PMID- 11264640 TI - Sequential anastomosis of accessory renal artery to inferior epigastric artery in the management of multiple arteries in live related renal transplantation: a critical appraisal. AB - In live related renal transplant program, management of multiple renal arteries (MRA) is technically demanding and used to be considered a relative contraindication because of increased risk of vascular and urologic complications. We present a retrospective analysis of the outcome of grafts with MRA and suggest certain guidelines. Of the 680 live related kidney transplantations done, 53 allografts had MRA. Cases were grouped according to the reconstruction technique: group A, MRA reconstructed ex vivo into a single renal artery (n=27); group B, MRA with multiple anastomoses in vivo (n =13); group C, MRA with sequential revascularization using inferior epigastric artery (n=11). We compared serum creatinine, acute tubular necrosis, rejection rates and the rewarm ischemia time between the three groups. Overall patient survival and graft survival were excellent (100 and 96%). Mean serum creatinine at 1 yr did not differ significantly between the three groups. Rewarm ischemia time was significantly less in group C (p<0.01). Incidence of acute tubular necrosis and rejection episodes was also less in group C although the difference was statistically significant only between group C and group B. We conclude that allografts with MRA can be used successfully in a live related renal transplantation program. Bench reconstruction should be done whenever possible. For reconstruction of an accessory vessel, inferior epigastric artery with sequential revascularization is recommended. PMID- 11264641 TI - Cyclosporin A withdrawal in live related renal transplantation: long-term results. AB - Cyclosporin A (CsA) withdrawal after 1 yr of stable graft function has been shown to be beneficial in cadaveric renal transplantation. This strategy could be even more suitable for 'immunologically advantaged' grafts as in live related renal transplantation. We report the long-term outcome of patients in a live related transplantation programme undergoing early (between 1989 and 1992) and late (1993 onwards) CsA withdrawal as compared with those on long-term low dose CsA (1993 onwards). Two-hundred and fifty-two patients were divided into three groups based on the following immunosuppressive protocol: group ECyW (n=99), early CsA withdrawal (9 months after transplantation); group LCyW (n=44), late CsA withdrawal (median 16 months, range 13--22 months after transplantation); and group LDCy (n=109), long-term low dose CsA. The median period of follow-up was 66 months after transplantation (range 43--84 months). There was no difference in the actuarial 6-yr patient or graft survival among the three groups. Acute rejection episodes were more frequent in ECyW (54.4%) than in LDCy (31.8%) and LCyW (23.8%) (p=0.001). The risk of developing late (> or =9 months) acute rejection was highest in ECyW 32/99 (32.3%) as compared with LCyW 8/44 (18.4%; p=0.08) and LDCy 8/109 (7.3%; p=0.0001). Of the 32 ECyW patients who developed acute rejection episodes after CsA withdrawal, 13 (40.6%) lost their grafts either due to uncontrolled acute rejection or to chronic rejection. Chronic rejection was higher in ECyW (24%) than in LCyW (11%; p=0.04) and LDCy (17%; p=0.17). Antihypertensive requirement was highest in patients maintained on low dose CsA. Graft function, as measured by serum creatinine levels, was significantly better in LCyW (1.24+/-0.4 mg%) as compared with ECyW (1.49+/-0.5 mg%) and LDCy (1.48+/-0.6 mg%). Early CsA withdrawal after live related renal transplantation is associated with a significant risk of acute rejection and subsequent chronic rejection. Slow withdrawal after 1 yr is safe and more economical than the long-term administration of low dose CsA. PMID- 11264642 TI - The use of intravenous tacrolimus and mycophenolate mofetil as induction and maintenance immunosuppression in simultaneous pancreas--kidney recipients with previous transplants. AB - Clinical trials using quadruple immunosuppression that include the combination of tacrolimus (TAC) and mycophenolate mofetil (MMF) have been shown to reduce the incidence of acute rejection episodes in simultaneous pancreas-kidney (SPK) transplantation. In attempting to obtain a low rejection rate without antibody induction therapy, we proceeded with the combination of TAC intravenous (i.v.), MMF, and steroids as induction therapy and as primary immunosuppression for recipients with previous transplants. In this study, we analyzed 10 patients who received previous transplants, treated with low-dose TAC i.v. as induction therapy. Group A consisted of 6 patients with previous transplants that underwent SPK and group B consisted of four recipients with previous SPK that underwent cadaveric kidney transplants. For group A, the previous transplants were: living related kidney (LRK) followed by islet cell (IC) transplant (n=2), LRK transplant (n=1), cadaver kidney (CAD) and IC transplant (n=1), SPK (n=1), and three previous CAD kidney transplants (n=1). In group A, all six kidneys were lost due to recurrent diabetic nephropathy, IC possibly to rejection, and the pancreas due to thrombosis. In group B with previous SPK transplants, three recipients lost their kidney to chronic rejection and one to long-term use of a nephrotoxic antibiotic. Currently, in all group A and B patients, the kidney and the pancreas are functioning, although 1 patient in group A developed type 2 diabetes (normal fasting C-peptide). Two patients in group A developed three rejection episodes that responded to steroid treatment. The results indicate the TAC i.v. in combination with oral TAC, MMF, and steroids offer effective induction therapy in patients with previous transplants. PMID- 11264643 TI - Insulin, blood pressure and elevated Na+/H+ exchange activity--novel therapeutic implications. PMID- 11264644 TI - HIV-1 infection: new treatment paradigms. PMID- 11264645 TI - Activation of Na+/H+ exchanger is associated with hyperinsulinemia in borderline hypertensive rats. AB - Activation of Na+/H+ exchanger (NHE) is known to be related to elevated blood pressure in hyperinsulinemia. To test whether there is the change in NHE activity in insulin resistance, we measured NHE activity of platelets in fructose-induced hyperinsulinemia in Wistar-Kyoto rats (WKY), in borderline hypertensive rats (BHR), and in spontaneously hypertensive rats (SHR). All rats were fed a 60% fructose diet for 4 weeks to induce hyperinsulinemia and hypertriglyceridemia. Intracellular pH (pHi) was measured with a pH-sensitive fluorescent dye 2'7'-bis (2-carboxyethyl)-5-carboxyfluorescein acetoxymethyl ester. NHE activity was evaluated by the recovery of pHi following addition of sodium propionate (Vmax). Measurement of intracellular calcium ([Ca2+]i) was performed using fura2/acetoxymethylester. Systolic blood pressure in fructose diet BHR elevated significantly greater than that in control diet BHR with the increase of both [Ca2+]i and Vmax. In WKY, there was no significant increase in systolic blood pressure and [Ca2+]i except Vmax in a fructose diet. Vmax in control diet SHR was greater than in control diet WKY and BHR, and we found no additional increase in Vmax with a fructose diet in SHR. In BHR, a high salt diet increased systolic blood pressure and Vmax to a similar degree as a fructose diet or a high salt combined with a fructose diet. Plasma insulin concentration correlated positively with Vmax in WKY and BHR, but not SHR. A fructose diet induces hyperinsulinemia and elevates blood pressure in BHR. Hyperinsulinemia appears to activate NHE in a different manner in SHR, and might be associated with an elevation in blood pressure in BHR. PMID- 11264646 TI - Moderate exercise, postprandial lipaemia and triacylglycerol clearance. AB - Moderate intensity exercise reduces postprandial triacylglycerol (TG) concentrations. We tested whether this reflects increased TG clearance. Eight normotriglyceridaemic men, aged 48.3 +/- 7.3 years (mean +/- SD), performed two oral fat tolerance tests (blood samples taken in the fasted state and for six hours after a high-fat meal containing 1.00 g fat, 0.97 g carbohydrate, 58 kJ energy kg-1 fat-free body mass) and two intravenous fat tolerance tests (blood samples in the fasted state and after a bolus injection of Intralipid, 0.1 g fat kg-1 body mass). The afternoon before one oral and one intravenous test, subjects walked briskly for 90 min; no exercise was performed before the control tests. Prior exercise reduced fasting TG concentration similarly in the oral (16 +/- 7 %) (mean +/- SEM) and intravenous (18 +/- 7 %) tests, and reduced postprandial TG concentrations in the oral test by 18 +/- 6 % (all P < 0.05). However, prior exercise did not increase Intralipid clearance (disappearance curve slopes: control, 4.69 +/- 0.49 % min-1; exercise, 4.85 +/- 0.40 % min-1). These data suggest that mechanisms other than increased TG clearance mediate the lower postprandial TG concentrations seen after moderate exercise. PMID- 11264647 TI - Somatostatin receptor subtype 2 and 5 in human GH-secreting pituitary adenomas: analysis of gene sequence and mRNA expression. AB - The role of somatostatin receptor subtypes 2 and 5 (SSTR2 and SSTR5) in determining the secretory and proliferative phenotype as well as the sensitivity to somatostatin analogue treatment is not clearly established. We quantified the expression of SSTR2 and SSTR5 mRNA using a semiquantitative reverse transcription polymerase chain reaction (RT-PCR) in 19 human growth hormone (GH) -secreting adenomas. Tumour characteristics and in vivo sensitivity to somatostatin analogues were assessed; tumours were screened for Gsalpha gene mutations. PCR products of SSTR2 and SSTR5 DNA from tumours resistant to somatostatin analogues were directly sequenced. All tumours expressed both SSTR2 and SSTR5 mRNA at variable levels. No significant correlation between SSTR2 and SSTR5 expression and the presence of Gsalpha mutation, GH levels, or tumour size and invasiveness was observed. A negative correlation between SSTR2 and SSTR5 mRNA levels was observed (r = 0.5; P < 0.05). No significant correlation between the levels of SSTR2 and SSTR5 expression and the in vivo responsiveness to somatostatin analogues was observed, although a tendency to a low SSTR2 expression in resistant tumours was found. No mutations in the coding or bordering regions of either SSTR2 or SSTR5 adenomatous DNA from patients totally or partially resistant to somatostatin analogues were found. The study shows that the different expression of SSTR2 and SSTR5 in GH-secreting adenomas is not significantly correlated with the secretory and proliferative phenotype, although the large, hypersecretory tumours and those with a poor sensitivity to somatostatin analogues seem to express low levels of SSTR2 mRNA. Moreover, both SSTR2 and SSTR5 DNA from tumours resistant to somatostatin analogues were found to possess intact coding sequences. PMID- 11264648 TI - Vasorelaxant effects of oestradiols on guinea pigs: a role for gender differences. AB - Various studies have shown vasorelaxation properties for 17alpha- and 17beta oestradiol. Here, we studied the effects of gender difference as well as oestrous cycle on oestradiol-induced vasorelaxation in mesenteric arteries from male and female guinea-pigs and in main uterine arteries from female guinea-pigs in vitro. Both 17alpha- and 17beta-oestradiol (0.5-20 micromol L-1) induced concentration dependent relaxation of both mesenteric and uterine arteries preconstricted with either noradrenaline (NA; 10 micromol L-1) or KCl (125 mmol L-1) from both day-7 and day-15 female guinea-pigs. 17beta-oestradiol was more potent in relaxing arteries precontracted with NA than those pretreated with KCl, while 17alpha oestradiol was more effective in relaxing those arteries precontracted with KCl. In mesenteric arteries from male animals, 17alpha-oestradiol was significantly (P < 0.001) more potent on arteries precontracted with NA than KCl, an effect opposite to that seen on arteries from female animals. However, both 17alpha- and 17beta-oestradiol were more potent in relaxing arteries from male animals compared with their female counterparts. These data indicate a possible role for gender differences in the vascular effects of oestradiols. PMID- 11264649 TI - IgG subclasses of anti Saccharomyces cerevisiae antibody in inflammatory bowel disease. AB - Elevation of serum anti Saccharomyces cerevisiae antibody (ASCA) has been reported in patients with Crohn's disease. We analysed the subclasses of Immunoglobulin (Ig) G reaction in ASCA in sera from patients with inflammatory bowel disease, healthy controls, and patients with intestinal Behcet's disease. Serum samples were obtained from 29 patients with Crohn's disease, 30 patients with ulcerative colitis, 7 patients with intestinal Behcet's disease, and 12 healthy controls. Serum IgG subclasses IgG1, IgG2, IgG3, and IgG4 of ASCA were analysed using ELISA. IgG4 ASCA was significantly increased in patients with inflammatory bowel disease. In patients with intestinal Behcet's disease, IgG1, IgG3, and IgG4 ASCA were increased. Differential responses, in terms of subclasses in ASCA, were found in patients with inflammatory bowel disease and patients with intestinal Behcet's disease, which may represent different pathophysiologies of these intestinal inflammatory diseases. PMID- 11264650 TI - The 13C/2H-glucose test for determination of small intestinal lactase activity. AB - To diagnose hypolactasia, determination of lactase enzyme activity in small intestinal biopsy material is considered to be the golden standard. Because of its strongly invasive character and the sampling problems, alternative methods have been looked for. We analysed the 13C-glucose response in serum after consumption of 25 g of naturally enriched 13C-lactose. As an internal standard, 0.5 g of 2H-glucose was added and the 2H-glucose response in serum was measured simultaneously. The studies were performed in healthy volunteers with a background of genetically determined lactase nonpersistence (n = 12; low lactase activity) and lactase persistence (n = 27; high lactase activity). The results were compared with those of the lactose hydrogen breath test, the lactose 13CO2 breath test and the previously described 13C-lactose digestion test. After consumption of 13C-lactose and 2H-glucose, the mean ratio 13C-glucose/2H-glucose concentration in serum at 45-75 min was 0.26 +/- 0.09 in the low lactase activity group and 0.93 +/- 0.17 in the high lactase activity group (P < 0.01). Threshold of the ratio between digesters and maldigesters was calculated as 0.46. Accuracy of the new test was superior to all other tests. We conclude that the 13C/2H glucose test has the potential of determining the small intestinal lactase activity in vivo and of estimating the amount of lactose which is digested in the small intestine. PMID- 11264651 TI - Inhibition of nitric oxide synthesis by aminoguanidine increases intestinal damage in the acute phase of rat TNB-colitis. AB - The role of nitric oxide (NO) in the pathophysiology of inflammatory bowel diseases (IBD) is controversially discussed. The aim of the present study was to investigate the role of NO inhibition in the acute phase of rat 2,4,6 trinitrobenzenesulphonic acid (TNB)-colitis. To inhibit NO synthesis we used aminoguanidine (AG) as a selective inhibitor of inducible nitric oxide synthase (iNOS). TNB-colitis was induced in rats with and without pretreatment with AG (200 mg kg-1 body weight in the drinking water). The severity of colitis was observed over a period of 7 days. On days 1 and 2, AG reduced concentrations of plasma nitrate and nitrite as well as of portal 6-keto-prostaglandin 1alpha. AG pretreatment increased colonic damage and inflammatory response, assessed by colonic myeloperoxidase and serum lactate dehydrogenase activity, macroscopic damage score, tumour necrosis factor-alpha concentration in stool and colonic glutathione content. The AG-treated group showed a higher and prolonged nuclear factor kappaB (NF-kappaB)/Rel binding activity in the colon. We conclude that NOS inhibition by AG is not beneficial in acute intestinal inflammation. With regard to appropriate therapeutic strategies, NF-kappaB/Rel activation might be a more suitable target. PMID- 11264652 TI - Low frequency of p53 and ras mutations in bile of patients with hepato-biliary disease: a prospective study in more than 100 patients. AB - The diagnosis of biliary disease, namely malignant disorders, is frequently hampered by the inconclusive cytology. We investigated prospectively the frequency of molecular changes in p53 and ras compared with cytology in patients with primary or secondary hepato-biliary disease. We investigated 118 consecutive patients, aged 24-89 with the following clinical diagnoses: choledocho/cholecystolithiasis (28), cholangiocellular carcinoma (21), gall bladder tumor (8), liver metastasis (3), autoimmune disease (8), chronic pancreatitis (16), pancreatic carcinoma (11), papillary disease (4), hepatic cirrhosis (6), cholangitis (2), anomalies (2), and normal (9). Bile was aspirated during routine endoscopic retrograde cholangio pancreatography (ERCP) or percutaneous transhepatic cholangiography (PTC). DNA was prepared freshly from a native aliquot. p53 mutations were detected by polymerase chain reaction (PCR) for exons 5 through 8 followed by TGGE. PCR for ras mutations was performed as RFLP-PCR with sequencing. In four cases, mutations in p53 could be found in exons 6 and 7. Twenty-two samples showed ras mutations; ras mutations were found in choledocholithiasis (4/28), bile duct (5/21), gall bladder (3/8) and pancreatic (1/11) carcinoma, liver metastasis (3/3), ulcerative colitis (2/3), PSC (1/2), and chronic pancreatitis (1/16). Cytology was clearly positive in seven cases, suspicious in three other, inconclusive in six, and negative in the rest. The molecular analysis resulted in a sensitivity of 33% and specificity of 87%, respectively, for the diagnosis of a malignant condition. PCR for p53 and ras mutations may aid the diagnosis of primary and secondary (metastatic) hepatobiliary disease if a malignant condition of the bile ducts and the liver is suspected and cytology is inconclusive or negative. However, the incidence of p53 and ras mutations in bile seems less frequent than in other malignant conditions of the gastrointestinal tract and the pancreas and lower than in tissue, leaving a poor sensitivity and specificity. Nevertheless, the presence of a p53 and/or ras mutation per se supports a clinical suspicion of malignancy, even when the conventional cytology is negative or inconclusive. PMID- 11264653 TI - New drug combinations for the treatment of murine AIDS and macrophage protection. AB - The failure of highly active antiretroviral therapies (HAART) is mainly due to the existence of latent infected reservoirs, such as macrophages and resting CD4+ T cells. In this paper, we report the results that we obtained in a murine model of AIDS by alternating the administration of the lympholitic drug 2-Fluoro-ara AMP (Fludarabine) to eliminate the infected cells, with that of Azidothymidine (AZT) plus reduced glutathione (GSH) encapsulated in erythrocytes, to protect lymphocytes and macrophages not yet infected, respectively. Two groups of infected mice were treated as follows: one group was treated by alternating the administration of Fludarabine and AZT (treatment A), while the other group received the same treatment plus GSH-loaded erythrocytes given with AZT (treatment A + L-RBC). Fludarabine was administered intraperitoneally, AZT in the drinking water and GSH was encapsulated in erythrocytes by a procedure of hypotonic dialysis and isotonic resealing. The results obtained show that GSH loaded erythrocytes provide additive effects in all the parameters examined. Alternation of a lympholitic drug and antiretroviral drug is effective in reducing the progression of murine AIDS. Addition of a system to protect macrophages provides additive effects in almost all the parameters considered, confirming that combination therapies aimed at protecting different infectable cell compartments are better than treatments protecting mainly lymphocytes. PMID- 11264654 TI - Levels of vasodilators (SP, CGRP) and vasoconstrictor (NPY) peptides in early human burns. AB - An intact nociceptor system of primary afferent sensory nerves is important for the initiation of the inflammatory process and successful tissue repair. Dysfunction of this system could be a contributing factor for delayed wound healing in humans. We examined the levels of vasodilators [substance P (SP), calcitonin gene-related peptide (CGRP)] and a vasoconstrictor peptide [neuropeptide Y (NPY)] in the peripheral blood samples of patients with burns covering from 20 to 75% of body surface area. Thirteen patient samples were obtained immediately on admission (OA), which was within 12 h of the thermal injury, and 24 h post-admission (PA). Enzyme immunoassay techniques were used for the measurement of the neuropeptides. In addition, an inflammatory marker, tumour necrosis factor-alpha (TNF-alpha), and a myofibrillar protein, creatine kinase (CK), were examined and compared with levels in 13 control subjects. CGRP was high OA and the levels were maintained PA (P < 0.05). SP was also significantly high at both sampling times (P < 0.05). Although TNF-alpha and NPY were somewhat higher in the patients' samples than in the control samples, these levels were not statistically significant (P = NS). CK was higher OA (P < 0.01) than PA (P < 0.04), compared to controls. Plasma levels of SP and CGRP increased significantly in patients with thermal injuries. These peptides may yet be another group of neuromodulators playing a significant role in immune, pain, inflammatory and wound healing in burns. PMID- 11264655 TI - Effect of vitamin C on neutrophil function after high-intensity exercise. AB - High-intensity exercise leads to an increased risk of upper respiratory tract infections in athletes, which had been related to an exercise-induced impairment of neutrophil function. In this study, several indices of neutrophil function were analysed before and after a biathlon and the effect of oral vitamin C on neutrophil function was determined. Six athletes took 2 g vitamin C daily for 1 week prior to a biathlon and four athletes did not take any supplementation. Neutrophil phagocytosis was analysed by fluorescence microscopy and flow cytometry. Cytosolic calcium kinetics were assessed fluorometrically and neutrophil bactericidal ability was assessed by fluorescence microscopy. Reactive oxygen production was analysed by flow cytometry. Catecholamines were analysed by high-performance liquid chromatography. After high-intensity exercise there were significant reductions in the number of phagocytosed Escherichia coli per neutrophil and in neutrophil bactericidal ability. There was a significant exercise-dependent increase of catecholamines. There was no difference between the two groups of athletes. These results do not support the concept that vitamin C supplementation corrects neutrophil dysfunction after strenuous exercise. PMID- 11264656 TI - Closed-circuit organ perfusion technique for gene transfer into the lungs. An experimental trial on farm pigs. AB - In an attempt to develop gene therapy for lung diseases, we have explored a closed-circuit surgical perfusion method for gene transfer into the lung. For gene transfer we used a replication defective type 5 adenovirus carrying the E. coli beta-galactosidase gene as a reporter gene. The middle lobe of the right lung of eight young farm pigs was perfused in vivo via thoracotomy for up to 60 min with the viral solution. The gene transfer was performed using a closed circuit organ perfusion method in vivo. The efficiency of gene transfer was assessed visually by analysis of histologic sections after X-gal, PAS and immunohistochemical stainings. The lung perfusion resulted in transgene expression in the alveolar epithelial cells, capillary endothelial cells, airway epithelial cells and alveolar macrophages of the lung examined seven days after perfusion. The present results suggest that operatively performed closed-circuit warm lung perfusion method may be used for gene transfer in treatment of diseases that have pulmonary manifestations. PMID- 11264657 TI - Suppression of superantigen-induced lung injury and vasculitis by preadministration of human urinary trypsin inhibitor. AB - We examined whether the lung injury produced in rats by intraperitoneal injection of the superantigen, staphylococcal enterotoxin B (SEB), could be inhibited by intravenous preadministration of human urinary trypsin inhibitor (UTI), which exhibits multipotent inhibitory effects on serine proteinases such as plasmin, chymotrypsin, or human leukocyte elastase or cathepsin G, since preliminary experiments showed the ability of UTI to bind lipopolysaccharides and bacterial toxins. For ligand blotting analysis, four kinds of toxins were run on a slab gel and the binding of UTI to the toxins was visualized by immunoblotting. Lung tissue from 26 rats was used for immunohistochemistry using a mouse antirat CD 45 mAb and an antirat macrophage mAb. Lung tissue from 31 rats was used for measurement of myeloperoxidase activity before and after intraperitoneal injection of SEB, after infusion of PBS, UTI, PBS-SEB or UTI-SEB combination. Ten of the 26 rats described above were used for electron microscopy. Rat sera were used for measurement of TNF-alpha. Statistical analysis was performed using the Mann-Whitney U-test. Intraperitoneal injection of SEB caused an increase in the number of punctate areas of haemorrhage on the surface of the lung with time, and histological examination revealed lung injuries with different extents, vasculitis where inflammatory cells were concentrated, and infiltration of numbers of eosinophils into the alveolar septa. However, preadministration of UTI for rats markedly attenuated lung injury and vasculitis induced by intraperitoneal injection of SEB. This revealed, from a marked reduction in the number of inflammatory cells and the extent of injury, a marked inhibition of serum TNF-alpha production and reduction of myeloperoxidase content of rat lungs compared to controls. UTI may have defensive effects to infection by suppressing the early responses of stimulated cells to activated stimulus such as SEB as well as the release of stimulant-mediated cytokines via trapping of bacterial toxins. PMID- 11264658 TI - Developmental pattern of expression of the alpha chemokine stromal cell-derived factor 1 in the rat central nervous system. AB - Stromal cell-derived factor 1 (SDF-1) is an alpha-chemokine that stimulates migration of haematopoietic progenitor cells and development of the immune system. SDF-1 is also abundantly and selectively expressed in the developing and mature CNS, as we show here. At embryonic day 15, SDF-1 transcripts were detected in the germinal periventricular zone and in the deep layer of the forming cerebral cortex. At birth, granule cells in the cerebellum and glial cells of the olfactory bulb outer layer showed an SDF-1 in situ hybridization signal that decreased progressively within the next 2 weeks. In other regions such as cortex, thalamus and hippocampus, SDF-1 transcripts detected at birth progressively increased in abundance during the postnatal period. SDF-1 protein was identified by immunoblot and/or immunocytochemistry in most brain regions where these transcripts were detected. SDF-1 was selectively localized in some thalamic nuclei and neurons of the fifth cortical layer as well as in pontine and brainstem nuclei which relay the nociceptive response. The presence of SDF-1 transcripts in cerebellar granule cells was correlated with their migration from the external to the inner granular layers with disappearance of the signal when migration was completed. In contrast, SDF1 mRNA signal increased during formation of the hippocampal dentate gyrus and stayed high in this region throughout life. The selective and regulated expression of SDF-1 in these regions suggests a role in precursor migration, neurogenesis and, possibly, synaptogenesis. Thus this alpha chemokine may be as essential to nervous system function as it is to the immune system. PMID- 11264659 TI - Role of GAP-43 in mediating the responsiveness of cerebellar and precerebellar neurons to axotomy. AB - To determine whether the competence for axonal sprouting and/or regeneration in the cerebellar system correlates with GAP-43 expression, we have studied GAP-43 mRNA and protein expression in the postlesioned cerebellum and inferior olive. Purkinje cells transiently express GAP-43 during their developmental phase (from E15 to P5 in the rat) which consists of fast axonal growth and the formation of the corticonuclear projection. Adult Purkinje cells, which in control adult rats do not express GAP-43, are extremely resistant to the effects of axotomy but cannot regenerate axons. However, a late and protracted sprouting of axotomized Purkinje cells occurs spontaneously and correlates with a mild expression of GAP 43 mRNA. In contrast, inferior olivary neurons, despite their high constitutive expression of GAP-43, do not sprout but retract their axons and die after axotomy. Furthermore, mature Purkinje cells in cerebellar explants of transgenic mice that overexpress GAP-43 do not regenerate after axotomy, even in the presence of a permissive substrate (cerebellar embryonic tissue) and, contrary to the case in wild-type mice, they do not survive in the in vitro conditions and undergo massive cell death. These results show that the expression of GAP-43 is not only associated with axonal growth, but also with neuronal death. PMID- 11264660 TI - Defining the role of Drosophila lateral neurons in the control of circadian rhythms in motor activity and eclosion by targeted genetic ablation and PERIOD protein overexpression. AB - The ventral lateral neurons (LNvs) of the Drosophila brain that express the period (per) and pigment dispersing factor (pdf) genes play a major role in the control of circadian activity rhythms. A new P-gal4 enhancer trap line is described that is mostly expressed in the LNvs This P-gal4 line was used to ablate the LNvs by using the pro-apoptosis gene bax, to stop PER protein oscillations by overexpressing per and to block synaptic transmission with the tetanus toxin light chain (TeTxLC). Genetic ablation of these clock cells leads to the loss of robust 24-h activity rhythms and reveals a phase advance in light dark conditions as well as a weak short-period rhythm in constant darkness. This behavioural phenotype is similar to that described for disconnected1 (disco1) mutants, in which we show that the majority of the individuals have a reduced number of dorsally projecting lateral neurons which, however, fail to express PER. In both LNv-ablated and disco1 flies, PER cycles in the so-called dorsal neurons (DNs) of the superior protocerebrum, suggesting that the weak short period rhythm could stem from these PDF-negative cells. The overexpression of per in LNs suppresses PER protein oscillations and leads to the disruption of both activity and eclosion rhythms, indicating that PER cycling in these cells is required for both of these rhythmic behaviours. Interestingly, flies overexpressing PER in the LNs do not show any weak short-period rhythms, although PER cycles in at least a fraction of the DNs, suggesting a dominant role of the LNs on the behavioural rhythms. Expression of TeTxLC in the LNvs does not impair activity rhythms, which indicates that the PDF-expressing neurons do not use synaptobrevin-dependent transmission to control these rhythms. PMID- 11264661 TI - Neurochemical and electrophysiological evidence that 5-HT4 receptors exert a state-dependent facilitatory control in vivo on nigrostriatal, but not mesoaccumbal, dopaminergic function. AB - In this study we investigated, using in vivo microdialysis and single unit recordings, the role of serotonin4 (5-HT4) receptors in the control of nigrostriatal and mesoaccumbal dopaminergic (DA) pathway activity. In freely moving rats, the 5-HT4 antagonist GR 125487 (1 mg/kg, i.p.), without effect on its own, significantly reduced the enhancement of striatal DA outflow induced by 0.01 (-35%) and 0.1 (-66%), but not 1 mg/kg, s.c. haloperidol (HAL). Intrastriatal infusion of GR 125487 (1 microM) had no influence on basal DA outflow, but attenuated (-49%) the effect of 0.01 mg/kg HAL. Systemic administration of GR 125487 modified neither basal nor 0.01 mg/kg HAL-stimulated accumbal DA outflow. In halothane-anaesthetized rats, 1 or 10 mg/kg GR 125487, without effect by itself, failed to modify the changes in accumbal and striatal DA outflow elicited by electrical stimulation (300 microA, 1 ms, 20 Hz, 15 min) of the dorsal raphe nucleus. Finally, GR 125487 (444 microg/kg, i.v.), whilst not affecting basal firing of DA neurons within either the substantia nigra or the ventral tegmental area, reduced HAL-stimulated (1--300 microg/kg, i.v.) impulse flow of nigrostriatal DA neurons only. These results indicate that 5-HT4 receptors exert a facilitatory control on both striatal DA release and nigral DA neuron impulse flow only when nigrostriatal DA transmission is under activated conditions. Furthermore, they indicate that the striatum constitutes a major site for the expression of the control exerted by 5-HT4 receptors on DA release. In contrast, 5-HT4 receptors have no influence on mesoaccumbal DA activity in either basal or activated conditions. PMID- 11264662 TI - P2Y receptor-mediated inhibition of voltage-activated Ca(2+) currents in PC12 cells. AB - To search for inhibitory nucleotide receptors in the sympathoadrenal cell lineage of the rat, voltage-activated Ca(2+) currents were recorded in PC12 cells after differentiation with nerve growth factor. ADP and ATP, but not uridine nucleotides, reduced Ca(2+) current amplitudes and slowed activation kinetics. This effect was mediated by GTP binding proteins, as it was abolished by intracellular GDP beta S and after treatment with pertussis toxin. Furthermore, depolarizations preceding the activation of Ca(2+) currents abolished the ADP induced slowing of activation kinetics and attenuated its inhibitory action on current amplitudes. The modulatory effect of ADP was neither altered in the presence of adenosine receptor antagonists, nor mimicked by agonists at these receptors. In addition, the action of ADP was antagonized by reactive blue 2, but not by suramin or PPADS. Nucleotides tested for their inhibitory action on Ca(2+) currents displayed the following rank order of potency: 2-methylthio-ADP > or = 2 methylthio-ATP >> ADP beta S > ADP = ATP. When P2X receptors were blocked, the P2X agonists ATP and 2-methylthio-ATP still reduced Ca(2+) currents. The P2Y1 receptor antagonists adenosine-2'-phosphate-5'-phosphate and adenosine-3' phosphate-5'-phosphate did not alter the inhibitory action of ADP, whereas the Sp isomer of adenosine-5'-O-(1-thiotriphosphate) and 2'- and 3'-O-(4-benzoylbenzoyl) ATP showed significant antagonistic activity. These results demonstrate that PC12 cells express an as yet unidentified P2Y receptor with pharmacological characteristics similar to those of P2Y1. As receptor-dependent modulation of Ca(2+) channels is a key event in presynaptic inhibition, this receptor may correspond to previously described presynaptic nucleotide receptors mediating autoinhibition of sympathetic transmitter release. PMID- 11264663 TI - Fibroblast growth factor-2 is selectively modulated in the rat brain by E-5842, a preferential sigma-1 receptor ligand and putative atypical antipsychotic. AB - Fibroblast growth factor-2 (FGF-2) is a member of a large family of trophic factors whose expression is regulated under several conditions in different areas of the brain. The goal of our experiments was to determine whether the administration of 4-(4-fluorophenyl)-1,2,3,6-tetrahydro-1-[4-(1,2,4-triazol-1 il)butyl] pyridine citrate (E-5842), a sigma-1 receptor ligand and putative atypical antipsychotic, could regulate the expression of FGF-2. After chronic treatment with E-5842 (21 days, and the animals killed 24 h after the last administration), an up-regulation was observed of the expression of FGF-2 mRNA in the prefrontal cortex and the striatum, and a down-regulation of the expression of FGF-2 mRNA in the hypothalamus of the rat brain. Acute treatment with E-5842 (one single administration and animals killed 6 h later) up-regulated FGF-2 expression in the prefrontal cortex, the striatum, the hypothalamus and the hippocampus in a dose-dependent manner. The acute up-regulation was transient and disappeared 24 h after E-5842 administration. The induction of FGF-2 in the striatum after repeated administration has been described for clozapine, but our data concerning regulation in the prefrontal cortex suggest that this effect is unique to E-5852 among other antipsychotics. Given the neuroprotective activity of FGF-2, the data presented here might be relevant to the deficit in cognition and other symptoms that appear in schizophrenia. PMID- 11264664 TI - Synthesis of progesterone in Schwann cells: regulation by sensory neurons. AB - In peripheral nerves, progesterone synthesized by Schwann cells has been implicated in myelination. In spite of such an important function, little is known of the regulation of progesterone biosynthesis in the nervous system. We show here that in rat Schwann cells, expression of the 3 beta-hydroxysteroid dehydrogenase and formation of progesterone are dependent on neuronal signal. Levels of 3 beta-hydroxysteroid dehydrogenase mRNA and synthesis of [3H]progesterone from [3H]pregnenolone were low in purified Schwann cells prepared from neonatal rat sciatic nerves. However, when Schwann cells were cultured in contact with sensory neurons, both expression and activity of the 3 beta-hydroxysteroid dehydrogenase were induced. Regulation of 3 beta hydroxysteroid dehydrogenase expression by neurons was also demonstrated in vivo in the rat sciatic nerve. 3 beta-hydroxysteroid dehydrogenase mRNA was present in the intact nerve, but could no longer be detected 3 or 6 days after cryolesion, when axons had degenerated. After 15 days, when Schwann cells made new contact with the regenerating axons, the enzyme was re-expressed. After nerve transection, which does not allow axonal regeneration, 3 beta-hydroxysteroid dehydrogenase mRNA remained undetectable. The regulation of 3 beta-hydroxysteroid dehydrogenase mRNA after lesion was similar to the regulation of myelin protein zero (P0) and peripheral myelin protein 22 (PMP22) mRNAs, supporting an important role of locally formed progesterone in myelination. PMID- 11264665 TI - Overexpression of the myristoylated alanine-rich C kinase substrate decreases uptake and K(+)-evoked release of noradrenaline in the human neuroblastoma SH SY5Y. AB - The aim of this study was to investigate a possible role of the myristoylated alanine-rich C kinase substrate (MARCKS) in the mechanism of noradrenaline uptake and release in the human neuroblastoma cell line SH-SY5Y. A stable cell line showing a twofold overexpression of MARCKS was prepared by transfecting SH-SY5Y with pCEP4 containing MARCKS cDNA in the sense orientation. This cell line showed no changes in the expression of neurofilaments or markers of noradrenergic large dense-cored vesicles compared with both untransfected SH-SY5Y and SH-SY5Y transfected with pCEP4 only (mock transfected). Similarly, no differences in the rate of cell growth could be detected between these three cell lines. In contrast, specific uptake and depolarization-evoked (100 mM K(+)) release of noradrenaline from the cell line overexpressing MARCKS was inhibited by approximately 50% compared with mock-transfected SH-SY5Y. K(+)-evoked noradrenaline release enhanced by pretreatment with 12-O-tetradecanoylphorbol 13 acetate (100 nM) was also inhibited by 50%. In contrast, carbachol-evoked noradrenaline release was unaffected. Thus, in SH-SY5Y cells, overexpression of MARCKS leads to a decrease in the K(+)-evoked noradrenaline release possibly by increased actin cross-linking preventing the movement of noradrenaline containing large dense-cored vesicles to the plasma membrane in response to depolarization. PMID- 11264666 TI - Matrix metalloproteinase (MMP) system in brain: identification and characterization of brain-specific MMP highly expressed in cerebellum. AB - The matrix metalloproteinase (MMP) family, comprising more than 20 isoforms, modulates the extracellular milieu by degrading extracellular matrix (ECM) proteins. Because MMP is one of the few groups of proteinases capable of hydrolysing insoluble fibrillar proteins, they are likely to play crucial roles in regulating both normal and pathophysiological processes in the brain. However, little is yet known about their possible neuronal functions due presumably to their unusual redundancy and to the absence of a complete catalogue of isoforms. As an initial step in understanding the MMP system in the brain, we analysed an expression spectrum of MMP in rat brain using RT-PCR and discovered a novel brain specific MMP, MT5-MMP. MT5-MMP was the predominant species among the nongelatinase-type isoforms in brain. MT5-MMP, present in all brain tissues examined, was most strongly expressed in cerebellum and was localized in the membranous structures of expressing neurons, as assessed biochemically and immunohistochemically. In cerebellum, its expression was regulated developmentally and was closely associated with dendritic tree formation of Purkinje cells, suggesting that MT5-MMP may contribute to neuronal development. Furthermore, its stable postdevelopmental expression and colocalization with senile plaques in Alzheimer brain indicates possible roles in neuronal remodeling naturally occurring in adulthood and in regulating pathophysiological processes associated with advanced age. PMID- 11264667 TI - Photoreceptor plasticity in reaggregates of embryonic chick retina: rods depend on proximal cones and on tissue organization. AB - Plasticity of photoreceptors and their integration into epithelial structures homologous to an outer nuclear layer (ONL), was investigated in embryonic chick retinal cell reaggregates by immunohistochemistry using an antibody specific for red plus green cones (RG-cones) and an antibody for rods. If reaggregates are raised in the presence of pigmented epithelium (RPE), completely reconstructed, stratified retinal spheres are produced, where all rods and cones are integrated into an outer laminar ONL, similar to a normal retina. In the absence of RPE, 'rosetted' spheres form which contain internal rosettes homologous to an ONL. Only a minor fraction of cones and rods of 'rosetted' spheres are located within rosettes, while a larger fraction is diffusely displaced in nonorganized areas, thus, not contributing to an ONL-like epithelium. In both types of spheres, the total percentage of RG-cones was similar to the in vivo retina, indicating that expression of cones is autonomous. Following cones, after about one day, rods developed only within already existing RG-cone clusters. Thereby, the ratio of rods to RG-cones increases as the tissue organization decreases: for stratified spheres this ratio is, 0.50 (1 rod/2 cones; similar to mature retina); for rosettes, 0.74 (3 rods/4 cones) and for nonorganized areas, 1.09 (1 rod/1 cone) - a higher ratio under our conditions has never been detected. Thus, rod expression depends strictly on the presence of nearby cones; their relative numbers are distinctively adjusted according to the cytoarchitecture of the tissue environment. The biomedical implications of these findings are briefly discussed. PMID- 11264668 TI - Nonactivated microglia promote oligodendrocyte precursor survival and maturation through the transcription factor NF-kappa B. AB - We demonstrate a role for nonactivated rat microglia in the survival and maturation of oligodendrocyte precursor cells (OPCs). Media conditioned by nonactivated microglia increase the number of surviving galactocerebroside(+) (GalC(+)) oligodendrocytes in vitro at 48 h by inhibiting the apoptosis of OPCs and stimulating their maturation to GalC+ oligodendrocytes. These effects are not observed with medium conditioned by microglia activated with interferon-gamma (IFN-gamma). Conditioned medium from nonactivated microglia is associated with upregulation in OPCs of nuclear factor of kappa binding (NF-kappa B) p65 subunit. The use of antisense to the inhibitor of kappa binding (I kappa B) induces p65 subunit activation in OPCs and, in common with medium conditioned by nonactivated microglia, also inhibits OPC apoptosis and promotes cell maturation. Anti platelet-derived growth factor (PDGF) antibody abolishes this effect even though PDGF-A chain is expressed at similar levels within both nonactivated and IFN gamma-activated microglia and both conditioned media have similar levels of PDGF A chain bioactivity. However, only conditioned medium from nonactivated microglia recruit phosphatidyl-3-inositol (PI-3) kinase to the PDGF-alpha receptor and synergise with endogenous PDGF-A chain to increase NF-kappa B activation. These results suggest that, dependent on their state of activation, microglia produce soluble factors that promote oligodendrocyte development through an effect on the PDGF-alpha receptor-signalling pathway. PMID- 11264669 TI - Subfield-specific immediate early gene expression associated with hippocampal long-term potentiation in vivo. AB - It is not known whether NMDA receptor-dependent long-term potentiation (LTP) is mediated by similar molecular mechanisms in different hippocampal areas. To address this question we have investigated changes in immediate early gene and protein expression in two hippocampal subfields following the induction of LTP in vivo and in vitro. In granule cells of the dentate gyrus, LTP induced in vivo by tetanic stimulation of the perforant path was followed by strong induction of the immediate early genes (IEGs) Zif268, Arc and Homer. The increase in Zif268 mRNA was accompanied by an increase in protein expression. In contrast, we were unable to detect modulation of the IEGs Zif268, Arc, Homer and HB-GAM following induction of LTP by high-frequency stimulation of the commissural projection to CA1 pyramidal cells in vivo. In this pathway, we also failed to detect modulation of Zif268 protein levels. Zif268, Arc and Homer can be modulated in CA1 pyramidal cells approximately twofold after electroshock-induced maximal seizure, which demonstrates potential responsiveness to electrical stimuli. When LTP was induced in vitro neither CA1 pyramidal cells nor granule cells showed an increase in Zif268, Arc or Homer mRNA. However, in the slice preparation, granule cells have a different transcriptional state as basal IEG levels are elevated. These results establish the existence of subfield-specific transcriptional responses to LTP inducing stimulation in the hippocampus of the intact animal, and demonstrate that in area CA1-enhanced transcription of Zif268, Arc and Homer is not required for the induction of late LTP. PMID- 11264670 TI - Adrenoreceptor-mediated modulation of the spinal locomotor pattern during swimming in Xenopus laevis tadpoles. AB - This study focused on the contribution of different adrenoreceptor subtypes to the modulation of fictive swimming activity in a relatively simple, yet intact, lower vertebrate system, the immobilized Xenopus laevis tadpole and explored their possible role in mediating the noradrenergic modulation of spinal motor networks. In Xenopus embryos, near the time of hatching, activation of alpha(1) adrenoreceptors increased the duration of episodes of fictive swimming, whilst in larvae, 24 h after hatching, they were decreased. Activation of alpha(2) adrenoreceptors, however, markedly reduced episode duration at both developmental stages. Cycle periods in both stages were increased by the activation of alpha(1) and/or alpha(2) receptor subclasses, whereas beta adrenoreceptors were not apparently involved in the modulation of cycle periods or the duration of swim episodes. However, both beta and alpha(1) receptor activation decreased the intersegmental delay in the head-to-tail propagation of swimming activity, while alpha(2) receptors did not influence these rostro-caudal delays. Activation of neither alpha, nor beta, receptor subclasses had any consistent effect on the duration of ventral motor bursts. Our findings suggest that noradrenergic modulation of the swim-pattern generator in Xenopus tadpoles is mediated through the activation of alpha and beta adrenoreceptors. In addition, activation of particular receptor subclasses might enable the selective modulation of either the segmental rhythm generating networks, the intersegmental coordination of those networks or control at both levels simultaneously. PMID- 11264671 TI - Processing of facial emotional expression: spatio-temporal data as assessed by scalp event-related potentials. AB - Event-related potentials (ERPs) were recorded in 10 adult volunteers, who were asked to view pictures of faces with different emotional expressions, i.e. fear, happiness, disgust, surprise and neutral expression [Ekman, P. & Friesen, W.V. (1975). Pictures of Facial Affect. Consulting Psychologist Press, Palo Alto, CA]. ERPs were recorded during two different tasks with the same stimuli. Firstly, subjects were instructed to pay attention to the gender of the faces by counting males or females. Secondly, they had to focus on facial expressions by counting faces who looked surprised. The classical scalp 'face-related potentials', i.e. a vertex-positive potential and a bilateral temporal negativity, were recorded 150 ms after the stimulus onset. Significant differences were found, firstly between late-latency ERPs to emotional faces and to neutral faces, between 250 and 550 ms of latency and, secondly, among the ERPs to the different facial expressions between 550 and 750 ms of latency. These differences appeared only during the expression discrimination task, not during the gender discrimination task. Topographic maps of these differences showed a specific right temporal activity related to each emotional expression, some particularities being observed for each expression. This study provides new data concerning the spatio-temporal features of facial expression processing, particularly a late-latency activity related to specific attention to facial expressions. PMID- 11264672 TI - The D2 receptor is critical in mediating opiate motivation only in opiate dependent and withdrawn mice. AB - According to the dual systems model for opiate reward, dopamine mediates opiate motivation when an animal is in a deprived motivational state (i.e. opiate dependent and in withdrawal) and not when an animal is in a nondeprived state (i.e. previously drug-naive). To determine the role of the D2 dopamine receptor subtype in mediating opiate motivation, we examined the behaviour of N5 congenic D2 receptor knockout mice and their wild-type siblings in opiate-naive and opiate dependent and withdrawn place conditioning paradigms. Opiate-naive D2 receptor knockout mice demonstrated acquisition of morphine-conditioned place preference but failed to acquire place preference when conditioned in the deprived state. We propose that D2 receptor function is critical in mediating the motivational effects of opiates only when the animal is in an opiate-dependent and withdrawn motivational state. These findings also underscore the important influence of the genetic background to a given phenotype, as evidenced by the observation that increasing the allelic contribution from the 129/SvJ strain abolishes morphine place preference in C57BL/6 wild-type mice. PMID- 11264673 TI - Tuning for the orientation of spatial attention in dorsal premotor cortex. AB - We tested whether neuronal activity in the dorsal premotor cortex (PMd) reflected the orientation of selective spatial attention, as opposed to the target of a reaching movement, eye position and saccade direction. These four spatial variables were dissociated in two tasks, which both required that a monkey attend to a robot's location in order to know when to make a movement. However, the target of the reaching movement varied; it was the robot's location in one task, but a different location in the other task. Eye position was recorded, but not explicitly controlled. Of 199 PMd neurons sampled, 19% had activity related to eye position, and an overlapping 11% were related to saccade direction (totaling 24% of the PMd sample). Of the 152 PMd neurons that lacked oculomotor relationships, approximately 20% reflected the orientation of selective spatial attention. Attentional tuning may account, at least in part, for gaze-independent receptive fields and visuospatial, target or goal relationships in tasks involving stimulus-response incompatibility. PMID- 11264676 TI - Signal detection in amplitude-modulated maskers. I. Behavioural auditory thresholds in a songbird. AB - Vertebrates have evolved mechanisms to exploit amplitude modulations in background noise for improving signal detection. However, the mechanisms underlying this masking release are not yet well understood. Here we present evidence for masking release observed in European starlings (Sturnus vulgaris, Aves) that were trained in a Go/NoGo paradigm to report the detection of a short tone (20 ms) in 100% sinusoidally amplitude-modulated noise maskers (400 ms duration). Maskers centred at the tone frequency were composed of one, three, or five spectrally adjacent noise bands each of auditory filter bandwidth. Envelopes of the masking noise bands were either in-phase (i.e. coherent) or successively phase shifted by 90 degrees (i.e. incoherent). A release from masking of up to 28 dB was observed for detection of signals presented in dips of the envelope of coherent maskers compared with those presented in peaks of coherent maskers and in incoherent maskers. For maskers limited to one auditory filter (i.e. limited to the analysis channel tuned to the test signal) this masking release was about 10 dB less than that observed for maskers allowing a comparison across three or five auditory filters. This indicates that both within-channel cues and across channel cues are important for signal detection. These behavioural data provide the reference for the study of responses of auditory forebrain neurons in the same species reported in a companion paper [Nieder & Klump (2001) Eur. J. Neurosci., 13, 1033-1044]. PMID- 11264674 TI - GABA(A) receptors in the ventral tegmental area control bidirectional reward signalling between dopaminergic and non-dopaminergic neural motivational systems. AB - In the midbrain ventral tegmental area (VTA), both dopaminergic and nondopaminergic neural substrates mediate various behavioural reward phenomena. VTA GABAergic neurons are anatomically positioned to influence the activity of both the mesolimbic dopamine system and nondopamine efferents from the VTA. In order to examine the possible functional role of VTA GABA(A) receptors in neural reward processes, we performed discrete, bilateral microinjections of the GABA(A) receptor agonist, muscimol, or the GABA(A) receptor antagonist, bicuculline, into the VTA. Using a fully counterbalanced, unbiased conditioned place-preference paradigm, we demonstrate that activation of VTA GABA(A) receptors, with the GABA(A) receptor agonist muscimol (5--50 ng/microL), or inhibition of VTA GABA(A) receptors, with the GABA(A) receptor antagonist bicuculline (5--50 ng/microL), both produce robust rewarding effects. Furthermore, these rewarding effects can be pharmacologically dissociated: blockade of dopamine receptors with a dopamine receptor antagonist, alpha-flupenthixol (0.8 mg/kg; i.p.), or concurrent activation of VTA GABA(B) receptors with a GABA(B) receptor agonist, baclofen (70 ng/microL), blocked the rewarding properties of the GABA(A) receptor agonist, but had no effect on the rewarding properties of the GABA(A) receptor antagonist. These results suggest that, within the VTA, a single GABA(A) receptor substrate controls bidirectional reward signalling between dopaminergic and nondopaminergic brain reward systems. PMID- 11264675 TI - Maternal defence as an emotional stressor in female rats: correlation of neuroendocrine and behavioural parameters and involvement of brain oxytocin. AB - In order to study neuroendocrine and behavioural stress responses in female rats post partum we aimed to establish a relevant emotional stressor -- the maternal defence test based on maternal aggression of a lactating resident towards a virgin or lactating intruder approaching the cage. Exposure to maternal defence significantly elevated corticotropin (ACTH) and corticosterone responses of the residents and of virgin or lactating intruders, with an attenuated response in lactating residents and lactating intruders. Exposure to maternal defence increased plasma oxytocin in virgin intruders only. The aggressive behaviour displayed by the residents was directly correlated with the amount of defensive behaviour of the intruder and independent of the intruder's reproductive state. However, the amount of maternal and explorative behaviours displayed by the lactating residents was significantly higher when exposed to a lactating, compared to a virgin, intruder. ACTH responses in lactating residents exposed to virgin intruders were significantly correlated to the amount of offensive (direct correlation) and maternal (inverse correlation) behaviours they displayed. Plasma prolactin concentrations, elevated in lactating compared to virgin rats under basal conditions, were found to be reduced in the lactating residents and intruders in response to exposure to the maternal defence test, whereas it was unchanged in virgin intruders. To test for the involvement of brain oxytocin in neuroendocrine and behavioural responses of the lactating residents an oxytocin receptor antagonist (0.1 microg/5 microL) was infused icv 10 min prior to testing. This treatment increased basal, but not stress-induced, ACTH, corticosterone and oxytocin secretion. Whereas parameters of aggressive behaviour were unchanged, the antagonist reduced signs of maternal behaviour during maternal defence. In summary, the maternal defence test has been characterized as a relevant emotional stressor for female rats which is useful for studying neuroendocrine and emotional responses in females, in particular in the context of reproductive adaptations. PMID- 11264677 TI - Signal detection in amplitude-modulated maskers. II. Processing in the songbird's auditory forebrain. AB - In the natural environment, acoustic signals have to be detected in ubiquitous background noise. Temporal fluctuations of background noise can be exploited by the auditory system to enhance signal detection, especially if spectral masking components are coherently amplitude modulated across several auditory channels (a phenomenon called 'comodulation masking release'). In this study of neuronal mechanisms of masking release in the primary auditory forebrain (field L) of awake European starlings (Sturnus vulgaris), we determined and compared neural detection thresholds for 20-ms probe tones presented in a background of sinusoidally amplitude modulated (10-Hz) noise maskers. Responses of a total of 34 multiunit clusters were recorded via radiotelemetry with chronically implanted microelectrodes from unrestrained birds. For maskers consisting of a single noise band centred around the recording site's characteristic frequency, a substantial reduction in detection threshold (21 dB on average) was found when probe tones were presented during envelope dips rather than during envelope peaks. Such effects could also explain results obtained for masking protocols where the on frequency noise band was presented together with excitatory or inhibitory flanking bands that were either coherently modulated (in-phase) or incoherently modulated (phase-shifted). Generally, masking release for probe tones in maskers with flanking bands extending beyond the frequency range of a cell cluster's excitatory tuning curve was not substantially improved. Only some of the neurophysiological results are in agreement with behavioural data from the same species if only the average population response is considered. A subsample of individual neurons, however, could account for behavioural thresholds. PMID- 11264678 TI - Intrinsic connectivity of human auditory areas: a tracing study with DiI. AB - The human supratemporal plane contains the primary as well as several other auditory areas. We have investigated the intrinsic connectivity of these areas by means of antero- and retrograde labelling with the carbocyanin dye DiI (1,1' dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate). A total of 30 injections was placed in both hemispheres of four freshly fixed postmortem brains. Labelled neurons and axons were found in cortex around the injection. The retrograde labelling varied from faint to Golgi-like; most of the retrograde labelled neurons were layers II-III pyramids and only a few were nonpyramidal neurons. Labelled axons were dense in all layers near the injection site, while they became relatively rare in layer IV further away. The tangential spread of labelling differed among auditory areas. On Heschl's gyrus (corresponding to the primary auditory cortex and cytoarchitectonic areas TD and part of TB) intrinsic connectivity involved a relatively narrow part of cortex. They spread over larger parts of cortex in plana polare and temporale (areas TG, TA and the remaining part of TB). A number of injections also produced anisotropic labelling patterns. These results reveal differences in intrinsic connectivity between auditory areas. They suggest that intrinsic connections within the primary auditory area, area TD and part of TB that is on Heschl's gyrus, involve mainly nearby units or modules, probably with similar coding properties, whereas in surrounding areas, connections spread over more distant units and may play an important role in the integration of different auditory features. PMID- 11264679 TI - Dopamine and noradrenaline efflux in the rat prefrontal cortex after classical aversive conditioning to an auditory cue. AB - We used bilateral microdialysis in the medial prefrontal cortex (PFC) of awake, freely moving rats to study aversive conditioning to an auditory cue in the controlled environment of the Skinner box. The presentation of the explicit conditioned stimuli (CS), previously associated with foot shocks, caused increased dopamine (DA) and noradrenaline (NA) efflux. This conditioned response was dependent on the immediate pairing of the two stimuli; in the pseudoconditioned group that received an equal number of both stimuli, but in an unpaired fashion, no conditioned increases in efflux were observed. PMID- 11264680 TI - Place-cell firing does not depend on the direction of turn in a Y-maze alternation task. AB - Hippocampal place cells were recorded while rats solved a continuous spatial alternation task requiring short-term spatial memory. All cells that had a firing field on the stem of the Y-shaped maze were found to have a very similar pattern of discharge whether the rat was about to make a right or a left turn, and whether the preceding turn was a right or a left turn. Thus, the view that place cells encode a variety of events (including the direction of turns) useful for solving memory tasks is not well supported by the present data. We suggest several possible explanations to account for the discrepancy with other recent studies showing turn-related modulation of place-cell activity. PMID- 11264681 TI - Stimulation of corticospinal tract regeneration in the chronically injured spinal cord. AB - Acute spinal cord injury models have proved popular in studies aimed at identifying factors capable of influencing axonal regeneration within the central nervous system. In these models, the test factors (e.g. graft tissues or cells, antibodies, growth factors, etc.) are typically administered at the time of spinal cord injury. In this study, we use a rat chronic spinal cord injury model to identify possible factors which can stimulate regeneration of the chronically lesioned corticospinal tract axons. We demonstrate that surgical grafting of segments of autologous, preligated sural nerve, into the syrinx, stimulates sprouting and regeneration of the corticospinal tract as evidenced by the presence of anterograde labelled corticospinal tract processes within the cavity walls two or more weeks after treatment. Regrowing corticospinal processes were not observed within control animals. The anterogradely labelled corticospinal tract axons were found exclusively within the central grey tissue comprising the cavity walls with no regrowing corticospinal process observed within the white matter. A similar pattern of regeneration was observed following injection into the cavity of a suspension of minced autologous preligated sural nerve. Evidence of corticospinal tract regeneration was seen when either wheat germ agglutinin- horseradish peroxidase or biotinylated--dextran was used as an anterograde tracer. These data demonstrate that the chronically injured cortical motor neurons retain the capacity to regenerate for extended periods and that regeneration can be stimulated using grafts of minced, preligated autologous peripheral nerve tissue. PMID- 11264682 TI - GABA(A) and GABA(B) receptors on neocortical neurons are differentially distributed. AB - The distribution of functional neurotransmitter receptors on the surface of neurons is highly relevant for synaptic transmission and signal processing. To map functional GABA(A) and GABA(B) receptors on the somadendritic membrane of rat neocortical layer V pyramidal neurons we used patch-clamp recording in combination with infrared-guided laser stimulation to release gamma-aminobutyric acid (GABA) photolytically. The data strongly suggest that relatively more GABA(A) receptors are located at the apical dendrite and relatively more GABA(B) receptors near the soma. Such a specific distribution of GABA(A) and GABA(B) receptors may serve to compensate for differences in electrotonic voltage attenuation between GABA(A) and GABA(B) receptor-mediated inhibitory postsynaptic potentials (IPSPs). PMID- 11264683 TI - Headache in divers. AB - The increasing popularity of scuba diving has added a new category to the differential diagnosis of headache. Headache in divers, while uncommon and generally benign, can occasionally signify serious consequences of hyperbaric exposure such as arterial gas embolism, decompression sickness, and otic or paranasal sinus barotrauma. Inadequate ventilation of compressed gases can lead to carbon dioxide accumulation, cerebral vasodilatation, and headache. Other types of headache encountered in divers include exertional headache, cold stimulus headache, migraine, tension-type headache, acute traumatic headache, cervicogenic headache, carbon monoxide poisoning headache, and headache associated with envenomation. Correct diagnosis and appropriate treatment require a careful history and neurologic examination as well as an understanding of the unique physiologic stresses of the subaquatic environment. PMID- 11264684 TI - Naratriptan as short-term prophylaxis of menstrually associated migraine: a randomized, double-blind, placebo-controlled study. AB - OBJECTIVE: To determine the efficacy of naratriptan 1-mg and 2.5-mg tablets twice daily compared with placebo as short-term prophylaxis of menstrually associated migraine. BACKGROUND: Approximately 60% of women with migraine report headaches associated with their menstrual cycles. Results from an open-label study suggest that short-term administration of sumatriptan is useful in the prophylaxis of menstrually associated migraine. METHODS: A randomized, double-blind, three-arm, parallel-group, placebo-controlled study was conducted in women aged 18 years or older with a history of migraine with or without aura, as defined by the International Headache Society, of at least 6 months. Two dose strengths of naratriptan (1 mg, 2.5 mg) or identical-appearing placebo tablets (1:1:1) were administered twice daily for 5 days starting 2 days prior to the expected onset of menses across four perimenstrual periods. End points included the number of menstrually associated migraines, total migraine days, peak headache severity, lost work/activity time, migraine-related quality of life, and incidence of adverse events. RESULTS: Overall, the intent-to-treat population comprised 206 women (naratriptan 1 mg, n = 70; naratriptan 2.5 mg, n = 70, and placebo, n = 66); 171 women treated four perimenstrual periods. Significantly more perimenstrual periods per subject treated with naratriptan, 1 mg, were headache free compared with placebo (50% versus 25%, P =.003). Naratriptan, 1 mg, significantly reduced the number of menstrually associated migraines (2.0 versus 4.0, P <.05) and menstrually associated migraine days (4.2 versus 7.0, P <.01) compared with placebo. More patients treated with naratriptan, 1 mg, were headache-free across all treated perimenstrual periods compared with placebo (23% versus 8%). No difference in headache severity was observed in breakthrough headaches. The incidence and severity of adverse events was similar across treatment groups. Naratriptan, 2.5 mg, was not statistically superior to placebo for any measure. CONCLUSIONS: Naratriptan, 1 mg, with tolerability similar to placebo, is an effective, short-term, prophylactic treatment for menstrually associated migraine. PMID- 11264685 TI - Differentiating the efficacy of 5-HT(1B/1D) agonists. AB - OBJECTIVE: To examine, for a set of published clinical trials of serotonin (5 HT(1B/1D)) agonists as acute treatments for migraine, whether transformation of efficacy data into therapeutic gain (TG) or number needed to treat (NNT) is useful. BACKGROUND: Pivotal clinical trials of 5-HT(1B/1D) agonists in migraine use a primary end point of change in pain score from 3 or 2 to 1 or 0. Placebo response rates among such studies are variable. Meta-analytic comparisons of 5 HT(1B/1D) agonists often employ TG and NNT as efficacy measures. METHODS: Data from US product labeling or published sources were converted into TG (TG = active response rate [%] - placebo response rate [%]) and NNT (NNT = 1/TG). Pivotal clinical trial data were compared before and after transformation. RESULTS: Therapeutic gain ranged from 17.5% to 51%. The transformation of TG into NNT yielded no clinically significant difference in efficacy estimate for the range of 17.5% to 47% (N = 29 clinical trials). However, NNT and TG had a nonlinear relationship for some secondary end points. When the relationship between the standard primary and secondary end points was compared, the correlation of TG with clinical disability (Pearson coefficient R = 0.93) was stronger than for NNT. Placebo response rates correlated more strongly with NNT (R = 0.66) than active response rates (R = 0.42; N = 29 clinical trials), although both TG and NNT were sensitive to placebo response rate. CONCLUSIONS: Transforming efficacy rates into TG or NNT adds no new information to placebo-controlled trials. The variables, TG and NNT, should not be used to compare members of this class of drugs. Migraine therapies can only be compared using well-designed head-to-head studies and not by meta-analysis. Broader measures of efficacy should be used to describe and compare 5-HT(1B/1D) efficacy. PMID- 11264687 TI - Divalproex in the long-term treatment of chronic daily headache. AB - OBJECTIVE: The purpose of this study was to assess the safety and efficacy of divalproex sodium in the long-term treatment of chronic daily headache. Correlations between treatment variables were assessed. BACKGROUND: Controlled and open-label trials of divalproex sodium have previously demonstrated its efficacy and safety in the treatment of migraine and chronic daily headaches. These data were primarily short-term and did not examine interaction between treatment variables. METHODS: Retrospective chart review with data extraction was conducted from headache diaries of 642 current patients under treatment with divalproex sodium for chronic daily headaches. One hundred thirty-eight of the patients were treated with only divalproex sodium. Demographic variables including age, sex, initial and final body weights, adverse events, dose of divalproex sodium, duration of treatment, and the ability to differentiate their chronic daily headache into its migraine and tension-type headache components were analyzed. Baseline and end of study headache frequency indices were obtained. RESULTS: The mean improvement was 47%, with an improvement in migraine of about 65%. At least a 50% reduction in headache frequency was reported by 93 of the 138 patients receiving treatment with only divalproex sodium. No correlation between response and age, sex, duration of treatment, and the prescribed dose of divalproex sodium was demonstrated. Adverse events occurred in approximately 35% of the patients. None were severe. Women were more likely to experience adverse effects than men. Weight gain, however, occurred less commonly in women (mean, 1.9 lbs) than in men (mean, 7 lbs). Initial body weight and age did not correlate with the weight change. CONCLUSIONS: Divalproex sodium can be used for a prolonged period as a sole agent for the successful treatment of chronic daily headache. Nearly 75% of the patients had at least a 50% reduction in headache frequency, and adverse effects occurred in approximately one third. Weight gain was negligible and hepatotoxicity did not occur during treatment periods of up to 6 years. PMID- 11264688 TI - Headache characteristics and race in Singapore: results of a randomized national survey. AB - This study presents the first account of the racial differences in headache prevalence and characteristics in the Singapore population. A questionnaire was administered to 2096 individuals from a randomized sample of 1400 households to test the hypothesis that race was independently correlated with headache diagnosis and morbidity. The overall lifetime prevalence of headaches in the study population was 82.7%; this did not vary between racial groups. The modal age of headache onset in all races was in the second decade and was similar in all races. Multivariate analysis showed that headache morbidity was independent of age, sex, income level, marital status, shift duties, and educational level, and correlated only with race and a positive family history of severe headache. Non-Chinese were more likely to suffer from severe headaches than Chinese, were more likely to seek medical attention, and were more likely to require medical leave for their symptoms. Non-Chinese had more migrainous headaches than Chinese, although characteristics of headache both groups experienced that were unrelated to severity differed only in a few aspects. We conclude that racial factors account for differences in headache classification, perception of headache severity and health-seeking behavior. PMID- 11264686 TI - Efficacy and tolerability of rizatriptan 10 mg in migraine: experience with 70 527 patient episodes. AB - As patients who suffer from migraine need long-term treatment, the safety and consistent efficacy of such therapy is very important. Concurrent illness and additional medication can interfere with the treatment chosen for the attacks of migraine. The objective of this open-label study was to investigate the efficacy and tolerability of rizatriptan, in the treatment of up to three attacks of migraine, in the clinical setting. From October 1998 to July 1999, 6 174 doctors enrolled 33 147 patients into the study (26 644 women, 650 men). The mean age was 42.7 years. We were able to examine standardized migraine diaries relating to 25 501 patients and 70 537 migrainous episodes. Rizatriptan scored consistently high on efficacy and showed a consistently rapid onset. There was no evidence of tolerance to repeated use. An effect was reported within 1 hour of ingestion in 79% of attacks treated. In 27.8% of attacks, remission of headache was complete at 1 hour. Two hours after ingestion, 74% of attacks had subsided completely. Repeated administration of rizatriptan was well tolerated, and few adverse effects were seen. The most common unwanted effects were dizziness, weakness, fatigue, and nausea. No cardiovascular disturbance was seen. In the clinical setting, rizatriptan, 10 mg, is an effective and well-tolerated agent for the treatment of migraine attacks. Particularly noteworthy is the rapid onset of effect, with swift disappearance of headache. Rizatriptan has a favorable side effect profile, and, provided contraindications are observed, severe adverse cardiovascular complications are extremely unlikely. PMID- 11264690 TI - Does relaxation treatment have differential effects on migraine and tension-type headache in adolescents? AB - The present study evaluated the effects of relaxation treatment in 36 adolescents aged 13 to 18 years (mean +/- SD, 15.4 +/- 1.55 years) suffering from migraine or migraine and tension-type headache as compared with a waiting-list condition group. The subjects rated various characteristics of migraine and tension-type headache in a diary. Significant reductions were found for total headache sum (P <.05) and intensity scores of total headache activity (P <.05) as well as for migraine intensity (P <.05) for subjects treated with relaxation as compared with those in the waiting-list condition group. However, no significant differences between the two groups were found for tension-type headache. Fifty percent of the adolescents treated with relaxation training attained a clinically significant improvement, compared with 12% of those in the waiting-list condition. It is suggested that treatment goals for migraine and migraine occurring with tension type headache might be different and that relaxation training also might benefit from focusing on specific aspects of the two headache types. PMID- 11264689 TI - Prevalence of migraine in patients with systemic lupus erythematosus. AB - OBJECTIVE: To determine the prevalence of migraine in patients with systemic lupus erythematosus (SLE), and to examine the relationships between headache type and other clinical, serologic, and treatment features of the disease. BACKGROUND: Headaches are common in SLE and are a significant source of patient disability. The exact prevalence of headaches in patients with SLE is unknown. The classification of headache syndromes in SLE is also unclear. Previous studies were based on small numbers of patients and the headache types and criteria to define headache types varied widely. METHODS: Four hundred fourteen patients meeting American College of Rheumatology criteria for the diagnosis of SLE were sent the University of California, San Diego Migraine Questionnaire. Patients who completed the questionnaire had their medical records reviewed for constitutional, respiratory, cardiac, vascular, skin, musculoskeletal, other neuropsychiatric, hematologic, renal, and immunologic manifestations of the disease. Recent corticosteroid, nonsteroidal anti-inflammatory drug, antimalarial, and immunosuppressive medications were also recorded. RESULTS: One hundred eighty-six patients completed the questionnaire. Sixty-two percent of patients reported headaches: 39% met diagnostic criteria for migraine and 23% met criteria for nonmigrainous headache. Of the patients with migraine, 56% met criteria for migraine without aura and 44% met criteria for migraine with aura. There were no significant associations between headache type and other clinical, serologic, or treatment features of the disease. CONCLUSIONS: There is a high prevalence of migraine in patients with SLE, and patients should be routinely evaluated for migraine symptoms. PMID- 11264691 TI - Coping strategies in episodic and chronic tension-type headache. AB - OBJECTIVE: To study the importance of coping with illness strategies in tension type headache (TTH). BACKGROUND: The pathophysiology of TTH is complex, and coping with illness strategies might contribute to the transformation to a chronic form. METHODS: We examined 89 subjects (mean age, 45.6 +/- 14.8 years; range, 18 to 72 years) with episodic (n = 37) and chronic (n = 52) TTH. Patients were required to fill in a Freiburg Questionnaire of Coping with Illness (FQCI), a von Zerssen Depression Scale, quality-of-life questionnaires, and a headache home diary (over 4 weeks). In addition, pressure pain thresholds (temporal muscles) and total tenderness scores were obtained. RESULTS: Patients with chronic TTH exhibited poorer quality-of-life measures, slightly more depressive symptoms, and significantly stronger avoidance behavior and endurance strategies on FQCI scales F4 and F5 (P< .05). There was no difference between episodic and chronic TTH with respect to measures of muscle tenderness or pain thresholds. CONCLUSIONS: We conclude that disadvantageous coping with illness strategies might contribute to a transformation to chronic TTH. PMID- 11264692 TI - Opiate use to control bowel motility may induce chronic daily headache in patients with migraine. AB - OBJECTIVES: To investigate whether opiate overuse might cause chronic daily headache in those with migraine, we studied patients who were taking codeine (or other opiates) for control of bowel motility after colectomy for ulcerative colitis. BACKGROUND: Analgesic overuse is considered by many to be one factor which can result in the transformation of migraine into a chronic daily headache pattern. Most of the evidence for this comes from patients with migraine who are taking increasing amounts of analgesia for headache. Many of these patients revert to an intermittent migraine pattern once the analgesics are stopped. METHODS: Women who were 1 year postcolectomy for ulcerative colitis were identified in several colorectal surgery practices in Calgary. They were sent a questionnaire designed to determine if they had a history of migraine prior to surgery, if they currently had chronic daily headache, what medications they were taking to control bowel motility, and what medications they were taking for headache. RESULTS: Twenty-eight patients who met our inclusion criteria returned completed questionnaires. Eight of these exceeded the recommended limits for opiate use in patients with headache. Eight patients met diagnostic criteria for migraine. Two patients had chronic daily headache starting after surgery. Both used daily opiates beginning after their surgery, and both had a history of migraine. The other six patients who used opiates daily did not have a history of migraine and did not have chronic daily headache. All patients with migraine who used daily opiates to control bowel motility following surgery developed chronic daily headache after surgery. CONCLUSIONS: Patients with migraine who use daily opiates for any reason are at high risk of developing transformed migraine with chronic daily headache. This risk appears much lower in patients without a history of migraine who use opiates for nonpain indications. PMID- 11264693 TI - Hemiplegic migraine during pregnancy: unusual magnetic resonance appearance with SPECT scan correlation. AB - OBJECTIVE: This article discusses the pathophysiology and implications for treatment of hemiplegic migraine within a case study presentation. BACKGROUND: We evaluated a 31-year-old white woman for hemiplegia in her 36th week of pregnancy. She initially presented with severe headache, dysarthria, lethargy, and left sided numbness and weakness. Hemiplegic migraine remains a diagnosis made by exclusion; neurologic examination of these patients is localizing, but nonspecific. DESIGN: Magnetic resonance imaging and single photon emission computed tomography scanning were performed on this patient during an exacerbation of headache associated with dense hemiplegia. RESULTS: Magnetic resonance imaging showed a superficial cerebral hemispheric signal abnormality with enhancement. Single photon emission computed tomography scanning confirmed hyperperfusion of that hemisphere. CONCLUSIONS: We believe the imaging evidence in our patient suggests that hemiplegia was caused and sustained by hyperperfusion. This case lends supportive evidence to a primarily vasodilatory mechanism and hyperperfusion as an etiology of the paralysis in such headaches and perhaps migraine with aura. PMID- 11264694 TI - Transitional interpersonality thunderclap headache. AB - OBJECTIVE: To report a patient with multiple personality disorder who experienced severe acute headaches without warnings, solely during the transition between her host personality and her pain-prone personality. BACKGROUND: The initial detailed description of headache in multiple personality disorder was made by Packard and Brown and published in this journal 15 years ago. METHODS: Clinical history, neurologic examination, electroencephalogram, and brain magnetic resonance imaging. RESULTS: A 54-year-old Holocaust survivor with an established diagnosis of multiple personality disorder had recurrent, excruciating, acute ("thunderclap") headaches only when switching between her domineering personality and her pain-prone personality, who suffered from chronic back pain. None of her personalities otherwise suffer from headaches. Electroencephalogram and brain magnetic resonance imaging were normal. CONCLUSION: This is an independent and current confirmation of the existence of transitional headaches in a patient with multiple personality disorder. They may occur as an isolated event during the switch process and have features of benign thunderclap headache. PMID- 11264695 TI - Zolmitriptan versus sumatriptan comparison trial. PMID- 11264697 TI - "Herbal" remedies and patient protection. PMID- 11264699 TI - Migraine-associated seizure: a case of reversible MRI abnormalities and persistent nondominant hemisphere syndrome. PMID- 11264701 TI - Re: "Acute intracranial hemorrhage caused by acupuncture" (Choo DC, Yue G. Headache. 2000;40:397-398.). PMID- 11264703 TI - Central nervous system in cerebral malaria: 'Innocent bystander' or active participant in the induction of immunopathology? AB - Cerebral malaria (CM) is a major life-threatening complication of Plasmodium falciparum infection in humans, responsible for up to 2 million deaths annually. The mechanisms underlying the fatal cerebral complications are still not fully understood. Many theories exist on the aetiology of human CM. The sequestration hypo-thesis suggests that adherence of parasitized erythrocytes to the cerebral vasculature leads to obstruction of the microcirculation, anoxia or metabolic disturbances affecting brain function, resulting in coma. This mechanism alone seems insufficient to explain all the known features of CM. In this review we focus on another major school of thought, that CM is the result of an over vigorous immune response originally evolved for the protection of the host. Evidence in support of this second hypothesis comes from studies in murine malaria models in which T cells, monocytes, adhesion molecules and cytokines, have been implicated in the development of the cerebral complications. Recent studies of human CM also indicate a role for the immune system in the neurological complications. However, it is likely that multiple mechanisms are involved in the induction of cerebral complications and both the presence of parasitized erythrocytes in the central nervous system (CNS) and immunopathological processes contribute to the pathogenesis of CM. Most studies examining immunopathological responses in CM have focused on reactions occurring primarily in the systemic circulation. However, these also do not fully account for the development of cerebral complications in CM. In this review we summarize results from human and mouse studies that demonstrate morphological and functional changes in the resident glial cells of the CNS. The degree of immune activation and degeneration of glial cells was shown to reflect the extent of neurological complications in murine cerebral malaria. From these results we highlight the need to consider the potentially important contribution within the CNS of glia and their secreted products, such as cytokines, in the development of human CM. PMID- 11264704 TI - Binding of Griffonia simplicifolia 1 isolectin B4 (GS1 B4) to alpha-galactose antigens. AB - Glycoconjugates with terminal Galalpha1-3Galbeta1-4GlcNAc sequences (alpha galactosyl epitopes, natural xenoreactive antigens) are present on various tissues in pigs and are recognized by human anti-alphagalactosyl (alphaGal) antibodies1. Hence xenotransplantation (pig-to-human) would trigger immune reactions involving complement activation and lead to the hyperactute rejection of the graft. Xenoreactive antigens are often studied by using the lectin Griffonia simplicifolia 1 isolectin B4 (GS1 B4), which shows high affinity to galactose. We here estimate the specificity of GS1 B4 for detecting various galactosyl epitopes by measuring lectin binding to neoglycoproteins, thyroglobulin and pig skeletal muscle. Enzyme linked lectin assays confirmed that GS1 B4 was highly specific to alpha-galactosylated neoglycoproteins while the lectin did not detect a beta-galactosylated ligand. The length of the sugar chains influenced the lectin-carbohydrate interaction. A monosaccharide linked to serum albumin showed higher lectin affinity than did neoglycoproteins with di- and tri-alpha-galactosyl epitopes. When the carbohydrate was extended, as in the xenoreactive pentasaccharide (Galalpha1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc), the carbohydrate- lectin interaction was meagre. Not only the terminal, but also the subterminal sugar affected the lectin binding because the GS1 B4 affinity to Galalpha1-3Gal was much stronger than to Galalpha1-3GalNAc. In bovine and porcine thyroglobulin most alphaGal epitopes appear to be cryptic, but are unmasked by a heat denaturation. In pig skeletal muscle there was lectin reaction not only in the muscle capillaries, but also in the connective tissue and intracellularly in muscle fibres. In Western blots of isolated proteins from pig muscle at least three bands were strongly stained after incubation with lectin. PMID- 11264705 TI - Effect of localized cytokine dysregulation: accelerated rejection of IL-2 expressing skin grafts. AB - Transgenic mice were created in which a sheep keratin promoter directed the expression of IL-2 into the dermis. These KIL-2 transgenic mice were used to investigate the effects of localized IL-2 dysregulation on immune responses. Peripheral tolerance to skin antigens was not broken by in situ IL-2 expression because syngeneic KIL-2 skin grafts were not rejected. However, MHC Class I disparate skin grafts from KIL-2 donors were rejected faster (median survival time (MST) 12 days) than grafts of non-transgenic littermate skin (MST 18 days). In contrast, the kinetics of KIL-2 H-Y-disparate skin graft rejection (MST 14 days) did not differ significantly from controls (MST 16 days), suggesting that upregulation of IL-2 at the effector site could affect CD4+ T cell- independent, but not CD4+ T cell-dependent, responses. No effect on rejection kinetics was observed when wild type allogeneic skin was grafted onto transgenic mice that expressed bcl2 constitutively in their lymphocytes (MST of 14 days, both sets), indicating that this was not simply due to increased longevity of T cells within the IL-2 expressing graft. We therefore suggest that aberrant expression of IL-2 can accelerate helper-independent CD8+ T cell responses by increasing proliferation and/or differentiation of cytolytic T cells at the effector site. PMID- 11264706 TI - Tumour necrosis factor-alpha: the role of this multifunctional cytokine in asthma. AB - Tumour necrosis factor-alpha (TNF-alpha) is recognized as an important mediator in many cytokine- dependent inflammatory events. It is known that TNF-alpha is released in allergic responses from both mast cells and macrophages via IgE dependent mechanisms, and elevated levels have been demonstrated in the bronchoalveolar fluid (BALF) of asthmatic subjects undergoing allergen challenge. Inhaled TNF-alpha increases airway responsiveness to methacholine in normal and asthmatic subjects associated with a sputum neutrophilia. Additional data indicate that TNF-alpha can upregulate adhesion molecules, facilitate the immigration of inflammatory cells into the airway wall and activate pro-fibrotic mechanisms in the subepithelium. These data suggest that TNF-alpha plays a role in the initiation of allergic asthmatic airway inflammation and the generation of airway hyper-reactivity. In addition, polymorphisms of the TNF-alpha gene 5' untranslated region, particularly at -308 bp, have been described as being associated with asthma. This polymorphism is associated with increased levels of TNF-alpha, but as yet, no asthma studies have demonstrated a phenotypic difference between those individuals with the polymorphism and those with the wild type gene. The TNF receptors (TNF-R p55 and p75), also known as CD120a and b, have also been shown to be present in the lung, but their functional importance is only just emerging. In asthma, TNF may function as a pro inflammatory cytokine that causes the recruitment of neutrophils and eosinophils. Treatment directed specifically at a reduction in TNF-alpha activity may conceivably be useful as a glucocorticosteroid-sparing asthma therapy. PMID- 11264707 TI - Murine model of chronic human asthma. AB - Human asthma is associated with acute and chronic inflammation of the airway mucosa, accompanied by airway wall remodelling. Experimental models of acute allergic bronchopulmonary inflammation in mice are useful for investigation of immunological mechanisms and of cellular recruitment, but have significant limitations because they fail to reproduce a number of characteristic lesions of human asthma, while usually being associated with marked alveolitis. We have developed an improved murine model of asthma that exhibits almost all of the morphological and functional changes that typify the human disease, without any confounding alveolitis. This model has considerable potential for the investigation of pathogenetic mechanisms and potential treatments of chronic human asthma. PMID- 11264708 TI - A study of the normal temporal healing pattern and the mucociliary transport after endoscopic partial and full-thickness removal of nasal mucosa in sheep. AB - The aim of this study was to assess the temporal healing process of nasal epithelium after full- thickness and partial thickness mucosal removal in sheep. Healing was assessed by histologically examining serial biopsies of the healing wounds. The histology assessed the regeneration of epithelium and return of cilia. Mucociliary clearance was measured before and after injury. On day 84 post injury partial thickness injuries had 80.7% (SEM = 10.25) normal epithelium and 68.35% (SEM = 19.2) reciliation. Full-thickness wounds had 64.98% (SEM = 19.17) normal epithelium and 32.96% (SEM = 17.46) reciliation. On day 84 the difference for epithelium regeneration was not significant (P > 0.05), but reciliation was significant (P < 0.05). The baseline mucociliary clearance was 0.84 mm per second (SEM = 0.2) and did not differ significantly from either the partial thickness wound transport rate (2.49 mm/s; SEM = 1.02) or the full-thickness transport rate (0.9 mm/s; SEM = 0.37) (paired t-test P > 0.05). The time period (84 days) for evaluation of reciliation was insufficient, as reciliation appeared to be continuing. The healing process took place over a longer time period than what had been previously reported in the literature and this may account for the symptoms seen in the postoperative period in patients after sinus surgery. PMID- 11264710 TI - New approaches in the treatment of asthma. AB - Asthma is a common and complex inflammatory disease of the airways that remains incurable. Current forms of therapy are long term and may exhibit associated side effect problems. Major participants in the development of an asthma phenotype include the triggering stimuli such as the allergens themselves, cells such as T cells, epithelial cells and mast cells that produce a variety of cytokines including IL-5, GM-CSF, IL-3, IL-4 and IL-13 and chemokines such as eotaxin. Significantly, the eosinophil, a specialized blood cell type, is invariably associated with this disease. The eosinophil has long been incriminated in the pathology of asthma due to its ability to release preformed and unique toxic substances as well as newly formed pro-inflammatory mediators. The regulation of eosinophil production and function is carried out by soluble peptides or factors. Of these IL-5, GM-CSF and IL-3 are of paramount importance as they control eosinophil functional activity and are the only known eosinophilopoietic factors. In addition they regulate the eosinophil life span by inhibiting apoptosis. While one therapeutic approach in asthma is directed at inhibiting single eosinophil products such as leukotrienes or single eosinophil regulators such as IL-5, we believe that the simultaneous inhibition of more than one component is preferable. This may be particularly important with eosinophil regulators in that not only IL-5, but also GM-CSF has been repeatedly implicated in clinical studies of asthma. The fact that GM-CSF is produced by many cells in the body and in copious amounts by lung epithelial cells highlights this need further. Our approach takes advantage of the fact that the IL-5 and GM-CSF receptors (as well as IL-3 receptors) utilize a shared subunit to bind, with high affinity, to these cytokines and the same common subunit mediates signal transduction culminating in all the biological activities mentioned. By generating the monoclonal antibody BION-1 to the cytokine binding region of the common subunit (betac) we have shown that the approach of inhibiting IL-5, GM-CSF and IL-3 binding and the resulting stimulation of eosinophil production and function with a single agent is feasible. Furthermore we have used BION-1 as a tool to crystallize and define the structure of the cytokine binding domain of betac. This knowledge and this approach may lead to the generation of novel therapeutics for the treatment of asthma. PMID- 11264709 TI - Regulation of the inflammatory response in asthma by mast cell products. AB - In airways, mast cells lie adjacent to nerves, blood vessels and lymphatics, which highlights their pivotal importance in regulating allergic inflammatory processes. In asthma, mast cells are predominantly activated by IgE receptor cross linking. In response to activation, preformed mediators that are stored bound to proteoglycans, for example, TNF-alpha, IL-4, IL-13, histamine, tryptase and chymase, are released. New synthesis of arachidonic acid metabolites (leukotriene C4 (LTC4), leukotriene B4 (LTB4) and prostaglandin D2 (PGD2)) and further cytokines is stimulated. Mediators from degranulating mast cells are critical to the pathology of the asthmatic lung. Mast cell proteases stimulate tissue remodelling, neuropeptide inactivation and enhanced mucus secretion. Histamine stimulates smooth muscle cell contraction, vasodilatation and increased venular permeability and further mucus secretion. Histamine induces IL-16 production by CD8+ cells and airway epithelial cells; IL-16 is an important early chemotactic factor for CD4+ lymphocytes. LTC4, LTB4 and PGD2 affect venular permeability and can regulate the activation of immune cells. The best characterized mast cell cytokine in asthmatic inflammation is TNF-alpha, which induces adhesion molecules on endothelial cells and subsequent transmigration of inflammatory leucocytes. IL-13 is critical to development of allergic asthma, although its mode of action is less clear. PMID- 11264711 TI - Epithelium-fibroblast interactions in response to airway inflammation. AB - Dramatic changes to the architecture of the airway walls have been commonly described in the airways of patients with asthma, cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD). Much research has focused on how airway inflammation drives these structural changes, particularly in terms of the mechanisms/mediators that are involved, and a number of parallels are observed between the disease phenotypes. For example, the increased deposition of extracellular matrix (ECM) at focal sites in the airway wall is seen in asthma and all interstitial lung diseases that involve fibrosis. In addition, increased expression of a number of well characterized cytokines and growth factors, such as TGF-beta and epidermal growth factor (EGF) have been demonstrated in these diseases. However, the role of the lesser-known cytokines, including the leukaemia inhibitory factor (LIF) and other members of the IL-6 family of cytokines in the pathogenesis of airway remodelling and fibrosis is largely unknown. However, the use of genetic manipulation in vivo and more specific inhibitors/antibodies in vitro has now provided increasing evidence to support the hypothesis that a complex interaction exists between these cytokines, ECM and integrins in regulating the function of both epithelial cells and fibroblasts. PMID- 11264712 TI - Distinct spatial requirement for eosinophil-induced airways hyperreactivity. AB - T helper (Th)-2-derived cytokines and their involvement in the recruitment and activation of inflammatory cells crucially orchestrate asthma pathogenesis. A notable cellular component of this allergy-induced inflammation is the eosinophil. However, whether the eosinophil is an obligatory mediator for enhancing airways hyperreactivity (AHR) to cholinergic stimuli, a watershed of the asthmatic lung, is somewhat controversial. In this investigation we have endeavoured to define the spatial requirements for IL-4 and IL-13, and the downstream effector molecules, IL-5 and the CC chemokine eotaxin, for the recruitment of eosinophils and the development of AHR in a murine model of allergic pulmonary disease. These studies are of particular importance considering clinical trials, with either the soluble IL-4Ralpha subunit or a humanized anti-IL-5 antibody, are being conducted. Interestingly, our studies show that depletion of both IL-4 and IL-13 is necessary to ablate pulmonary eosinophilia and AHR, and that this may be attributed to the role these cytokines play in regulating the expression of the eosinophil- activating molecules, IL-5 and eotaxin. While it is clear that depletion of IL-5 diminishes pulmonary eosinophilia, we demonstrate in BALB/c mice that a deficiency in both IL-5 and eotaxin is necessary to abolish both the trafficking of eosinophils to the lung and AHR. However, in contrast to the neutrophil-rich inflammation observed in mice deficient in both IL-4 and IL-13, inflammation per se in mice deficient in both IL-5 and eotaxin is significantly attenuated. This suggests that asthma immunotherapy may be better directed towards the eosinophil- activating molecules IL-5 and eotaxin, rather than towards pleiotrophic molecules such IL-4 and IL-13, which are additionally important in modulating alternative inflammatory responses. PMID- 11264713 TI - New insights into the role of zinc in the respiratory epithelium. AB - Over the past 30 years, many researchers have demonstrated the critical role of zinc (Zn), a group IIb metal, in diverse physiological processes, such as growth and development, maintenance and priming of the immune system, and tissue repair. This review will discuss aspects of Zn physiology and its possible beneficial role in the respiratory epithelium. Here we have detailed the mechanisms by which Zn diversely acts as: (i) an anti-oxidant; (ii) an organelle stabilizer; (iii) an anti-apopototic agent; (iv) an important cofactor for DNA synthesis; (v) a vital component for wound healing; and (vi) an anti-inflammatory agent. This paper will also review studies from the authors' laboratory concerning the first attempts to map Zn in the respiratory epithelium and to elucidate its role in regulating caspase-3 activated apoptosis. We propose that Zn, being a major dietary anti oxidant has a protective role for the airway epithelium against oxyradicals and other noxious agents. Zn may therefore have important implications for asthma and other inflammatory diseases where the physical barrier is vulnerable and compromised. PMID- 11264714 TI - Role of exhaled nitric oxide in asthma. AB - Nitric oxide (NO), an evanescent atmospheric gas, has recently been discovered to be an important biological mediator in animals and humans. Nitric oxide plays a key role within the lung in the modulation of a wide variety of functions including pulmonary vascular tone, nonadrenergic non-cholinergic (NANC) transmission and modification of the inflammatory response. Asthma is characterized by chronic airway inflammation and increased synthesis of NO and other highly reactive and toxic substances (reactive oxygen species). Pro- inflammatory cytokines such as TNFalpha and IL-1beta are secreted in asthma and result in inflammatory cell recruitment, but also induce calcium- and calmodulin independent nitric oxide synthases (iNOS) and perpetuate the inflammatory response within the airways. Nitric oxide is released by several pulmonary cells including epithelial cells, eosinophils and macrophages, and NO has been shown to be increased in conditions associated with airway inflammation, such as asthma and viral infections. Nitric oxide can be measured in the expired air of several species, and exhaled NO can now be rapidly and easily measured by the use of chemiluminescence analysers in humans. Exhaled NO is increased in steroid-naive asthmatic subjects and during an asthma exacerbation, although it returns to baseline levels with appropriate anti-inflammatory treatment, and such measurements have been proposed as a simple non-invasive method of measuring airway inflammation in asthma. Here the chemical and biological properties of NO are briefly discussed, followed by a summary of the methodological considerations relevant to the measurement of exhaled NO and its role in lung diseases including asthma. The origin of exhaled NO is considered, and brief mention made of other potential markers of airway inflammation or oxidant stress in exhaled breath. PMID- 11264715 TI - Prevalence, morbidity and management of adult asthma in South Australia. AB - This paper reviews asthma-related data obtained between 1987 and 1997 from self report population surveys of adults in South Australia. A multistage, systematic, clustered area sample of adults (>15 years) was selected from a random sample of Australian Bureau of Statistics collector districts, and interviewed at home by trained health interviewers. The self-report prevalence of doctor-diagnosed asthma increased from 5.6% in 1987 to 12.2% in 1997. Morbidity measured as days lost from usual activities and nights awakened by asthma remained high, but hospitalization rates are trending down. The ownership of asthma action plans peaked in 1995 and has declined. The ownership of peak flow meters increased between 1992 and 1997, and the ownership of nebulisers remained constant. Evidence-based interventions are required to improve asthma management. PMID- 11264717 TI - Clock gene protein mPER1 is rhythmically synthesized and under cAMP control in the mouse pineal organ. AB - The mammalian clock gene Per1 is an important element of endogenous oscillators that control daily rhythms in central and peripheral tissues. Although such autonomous clock function is lost in the mammalian pineal gland during evolution, mPer1 mRNA and mPER1 protein were found to be strongly elevated in the mouse pineal organ during the dark period compared to daytime values. In vitro studies showed that mPer1 mRNA and mPER1 protein in mouse pineal gland are induced following the activation of a signalling pathway of fundamental importance for pineal physiology, the norepinephrine/cAMP/phosphoCREB cascade. mPER1 may function in the mouse pineal gland as a time-measuring molecule to participate in regulating rhythmic cellular responses in vivo. PMID- 11264718 TI - Rapid upregulation of aromatase mRNA and protein following neural injury in the zebra finch (Taeniopygia guttata). AB - The expression of aromatase (oestrogen synthase) within the vertebrate central nervous system (CNS) is key in the provision of local oestrogens to neural circuits. Aromatase expression appears to be exclusively neuronal under normal conditions. However, some in vitro studies suggest the presence of astrocytic aromatase in songbirds and mammals. Recently, aromatase in reactive astrocytes has been demonstrated in response to neural injury in the mammalian CNS. Since the glial aromatase expression first documented in cultures of the songbird telencephalon may reflect processes similar to those in response to mammalian neural injury, we investigated whether injury alters the pattern of aromatase expression in the zebra finch, a species with very high levels of forebrain aromatase expression. Adult males received a penetrating neural injury to the right hemisphere and were killed either 24 or 72 h later. Controls were anaesthetized and otherwise unmanipulated. We determined the expression of aromatase mRNA and protein using in situ hybridization and immunocytochemistry, respectively. Both the transcription and translation of aromatase is dramatically upregulated around the lesion site in response to neural injury in the zebra finch forebrain. This effect is robust and rapid, occurring within 24 h of the injury itself. Cells that upregulate aromatase appear to be reactive astrocytes based upon morphology. The hemisphere contralateral to the injury and both hemispheres in control birds showed the normal, exclusively neuronal pattern of aromatase expression. The upregulation of aromatase in astrocytes may provide high levels of oestrogen available to modulate processes such as CNS repair. Injury-induced upregulation of astrocytic aromatase may be a general characteristic of the injured vertebrate brain. PMID- 11264719 TI - Effect of antisense suppression of transforming growth factor-beta3 gene on lactotropic cell proliferation. AB - The mechanism by which oestrogen regulates lactotropic cell proliferation is not well understood. Recently it has been shown that a transforming growth factor beta (TGF-beta) gene-related peptide, TGF-beta3, is produced in the lactotropes and stimulates lactotropic cell proliferation in vitro. In this study, the role of this growth factor in oestrogen-induced lactotropic cell proliferation was determined in vitro using oligonucleotide designed to inhibit TGF-beta3 gene expression. We used the oligonucleotide in an antisense orientation, which is complementary to regions in the TGF-beta3 message. Oligonucleotides in sense and missense orientation were also used. We found that the antisense sequence that was effectively incorporated into pituitary cells produced a marked inhibition of the stimulatory action of oestrogen on lactotropic cell proliferation. Sense and missense oligonucleotides produced no significant effects on oestrogen-stimulated lactotropic cell proliferation. The growth-inhibitory effect of antisense oligonucleotide was blocked by TGF-beta3 peptide. These results suggest that TGF beta3 may be involved in the processes that mediate oestrogen-regulated lactotropic cell proliferation. PMID- 11264720 TI - Expression of corticotropin releasing factor (CRF), urocortin and CRF type 1 receptors in hypothalamic-hypophyseal systems under osmotic stimulation. AB - The expression of corticotropin releasing factor (CRF) and urocortin in hypothalamic magnocellular neurones increases in response to osmotic challenge. To gain a better understanding of the physiological roles of CRF and urocortin in fluid homeostasis, CRF, urocortin and CRF type 1 receptor (CRFR-1) gene expression was examined in the hypothalamic-hypophyseal system usingin situ and double-label in situ hybridization following chronic salt loading. CRFR-1 expression was further examined by immunohistochemistry and receptor binding. Ingestion of hypertonic saline by Sprague-Dawley rats for 7 days induced CRF mRNA exclusively in the oxytocin neurones of the magnocellular paraventricular nucleus (PVN) and the supraoptic nucleus (SON), but induced CRFR-1 mRNA in both oxytocin and vasopressin-containing magnocellular neurones. Hypertonic saline treatment also increased urocortin mRNA expression in the PVN and the SON. In the SON, urocortin was localized to vasopressin and oxytocin neurones but was rarely seen in CRF-positive cells. Changes in CRFR-1 mRNA expression in magnocellular neurones by hypertonic saline treatment were accompanied by changes in CRFR-1 protein levels and receptor binding. Hypertonic saline treatment increased CRFR-1 like immunoreactivity in the magnocellular PVN and SON, and decreased it in the parvocellular PVN. CRF receptor binding in the PVN and SON was also increased in response to osmotic stimulation. Finally, hypertonic saline treatment increased CRFR-1 mRNA, CRFR-1-like immunoreactivity and CRF receptor binding in the intermediate pituitary. These results demonstrate that the increase in the expression of CRF and urocortin message in magnocellular neurones induced by salt loading is accompanied by an increase in CRF receptor levels and binding in the hypothalamus and intermediate pituitary. Thus, CRF and urocortin may exert modulatory effects locally within magnocellular neurones as well as at the pituitary gland in response to osmotic stimulation. PMID- 11264721 TI - Insulin and glucose administration stimulates Fos expression in neurones of the paraventricular nucleus that project to autonomic preganglionic structures. AB - Insulin and glucose play a key role in the control of body energy homeostasis. However, the anatomical organization of the network of central insulin and glucose sensitive areas is still unclear. In the present study, we used a multiple-labelling technique combining retrograde tracing and Fos-like immunohistochemistry, to analyse the anatomical projections from hypothalamic neurones activated by the combined stimulus of insulin and glucose. After intraperitoneal injections of a bolus of insulin plus glucose, Fos-like immunoreactive neurones were observed in the paraventricular nucleus (PVN), ventromedial and arcuate nuclei, as well as the lateral hypothalamic area. In addition, neurones projecting to the autonomic preganglionic levels in the brainstem and spinal cord potentially involved in the control of glucose metabolism were identified by injections of fluorochrome tracers. Thus, Fluorogold was injected into the intermediolateral cell column of the lower spinal cord and Fast Blue was injected into the dorsal motor nucleus of the vagus. Perikarya of descending neurones were detected chiefly in the dorsal, medial and lateral parvocellular subnuclei and also in the posterior magnocellular subnucleus of the PVN. In contrast, insulin-glucose activated neurones in the PVN were observed mainly in the medial parvocellular and posterior magnocellular subnuclei. Fluorogold/Fos double-labelled neurones were only observed in the ventral zone of the medial parvocellular subnucleus. These data indicate that, within the PVN, there could be neurones responding to insulin glucose administration, which are involved in the sympathetic control of the classical regulatory structures of body energy homeostasis, such as the liver and pancreas, and which could play a role in the output of the neuronal circuitry controlling food intake. PMID- 11264722 TI - The differential regulation of CART gene expression in a pituitary cell line and primary cell cultures of ovine pars tuberalis cells. AB - The cocaine-amphetamine regulated transcript (CART) encodes for a protein which has an important role in the regulation of appetite and body weight. To date, no details of the molecular events and signal transduction pathways which regulate this gene are available. We report the identification of CART gene expression in the GH3 pituitary cell line. We have used activators of the cAMP or protein kinase C (PKC) signal transduction pathways to show that, in GH3 cells, CART is transcriptionally up-regulated by activators of the cAMP signal transduction pathway. We also identify CART gene expression in ovine pars tuberalis (PT) tissue and primary cell cultures. In PT cells in contrast to GH3 cells, CART gene expression is upregulated by activators of the PKC signal transduction pathway. Cultured cells have provided a valuable resource for the detailed analysis of specific regulatory mechanisms underlying transcriptional or translational regulation of genes, signal transduction events and many other cellular processes. GH3 and PT cells may therefore provide a resource for the further detailed molecular analysis of the events regulating CART gene expression and processing. PMID- 11264723 TI - 5Alpha-reductase type 2 and androgen receptor expression in gonadotropin releasing hormone GT1-1 cells. AB - Gonadal steroids are potent modulators of gonadotropin releasing hormone (GnRH) secretion, and androgen binding sites and 5alpha-reductase activity have been found in the immortalized GnRH secreting cell line GT1-1, suggesting the existence of a direct androgenic control of GnRH dynamics. Two isoforms of the 5alpha-reductase have been cloned with very different biochemical/functional properties: 5alpha-reductase type 1 (widely distributed in the body) and 5alpha reductase type 2 (confined in androgen target structures). We have analysed whether, in GT1-1, androgen binding sites are linked to "classical" androgen receptor, and which 5alpha-reductase isoform is active. Reverse transcriptase polymerase chain reaction analysis showed that the mRNAs coding for androgen receptor and for the two 5alpha-reductase isoforms are all expressed in GT1-1 cells. However, the 5alpha-reductase enzymatic reaction showed a peak of activity at a narrow pH around 5.5, the optimum for the 5alpha-reductase type 2. The affinity for testosterone, of the enzyme present in GT1-1 cells, was very similar to that observed for the recombinant type 2 isozyme expressed in yeasts. The data indicate that GT1-1 cells (i) express a "classical" androgen receptor and (ii) contain the 5alpha-reductase type 2 isoform, a specific marker of androgen responsiveness. PMID- 11264724 TI - Regulation by osmotic stimuli of galanin-R1 receptor expression in magnocellular neurones of the paraventricular and supraoptic nuclei of the rat. AB - Neurones of the supraoptic nucleus (SON) and the magnocellular and parvocellular divisions of the paraventricular nucleus (PVN) express galanin and [125I]galanin binding sites. Although the precise role(s) of galanin in these different cell populations is still unknown, it has been shown to regulate the electrophysiological, neurochemical and secretory activity of magnocellular neurones. In light of the well-described effects of hyperosmotic stimuli, such as salt-loading on magnocellular neurone activity and galanin synthesis and release, and the recent identification of multiple galanin receptors in brain, this study assessed the possible regulation of galanin receptor subtype expression in the PVN/SON of salt-loaded, dehydrated and food-deprived rats. Gal-R1 mRNA was abundant in the SON (and magnocellular PVN) of control rats and levels were increased in these same cells after 4 days of salt-loading (2% NaCl solution as drinking water) or water deprivation. The density of specific [125I]galanin(1-29) binding and the intensity of Gal-R1-like immunostaining were also increased in the characteristically enlarged, magnocellular neurones of the PVN and SON after these treatments. Gal-R2 mRNA was detected in the parvocellular PVN, but levels were not altered by the hyperosmotic stimuli. In contrast, food deprivation (4 days), which has been shown to reduce levels of several neurochemical markers in magnocellular neurones, produced a significant reduction in Gal-R1 (and galanin) mRNA levels in the SON, but no consistent change in neurone size, [125I]galanin binding levels, or Gal-R1 immunostaining. Along with previous findings from this and other laboratories, these data suggest that the expression of galanin and Gal R1 receptors is regulated in parallel with functional and morphological changes in hypothalamic magnocellular neurones. Furthermore, Gal-R1 immunoreactivity was primarily detected in somatodendritic areas and thus galanin may influence the activity of these cells, particularly vasopressin synthesis/release, via autocrine or paracrine activation of Gal-R1 receptors, especially during long lasting stimulation. PMID- 11264725 TI - Prevention of glucoprivic stimulation of corticosterone secretion by leptin does not restore high frequency luteinizing hormone pulses in rats. AB - We have previously determined that exogenous leptin prevents the inhibition of pulsatile luteinizing hormone (LH) release in the fasting rodent. The present study tested the hypothesis that the mechanism by which leptin facilitates high LH secretion is through an attenuation of the stress response produced by a deficit in energy. Because hypogonadotropism is associated with activation of the hypothalamic-pituitary-adrenal (HPA) axis during both metabolic stress and nonmetabolic stress, our approach included a comparison of whether exogenous leptin could prevent the rise in corticosterone produced by a nonmetabolic stress (immobilization for 2 h), as well as by a widely used metabolic stress (transient glucoprivation by 2-deoxyglucose, 2DG; 400 mg/kg, b.w., i.v.). Each stressor was applied to well-fed ovariectomized rats (n = 4-6 per group), 2 h after leptin (3 microg/g, b.w., i.p.) or vehicle administration. Blood samples were collected through an indwelling atrial cannula every 6 min for 1 h before and for 2 h after the stress treatment to measure LH, leptin and corticosterone. During metabolic stress (acute glucoprivation), circulating leptin decreased, corticosterone increased and LH decreased; leptin administration abolished the increase in corticosterone, but pulsatile LH secretion remained inhibited. In contrast, during nonmetabolic stress (immobilization), leptin secretion was unaffected, but circulating corticosterone increased and LH decreased; leptin treatment did not prevent either the increase in corticosterone or the decrease in LH secretion. An important overall finding is that leptin can differentially alter the HPA axis depending upon the type of stress. In addition, whether the pattern of leptin is altered depends upon the type of stress. Although a glucoprivic-induced decrease in endogenous leptin can be a stressor responsible for the increase in corticosterone secretion, a nonmetabolic stress-induced increase in corticosterone is not mediated by leptin. Moreover, our results reveal that the depression of LH secretion when leptin is low during reduced energy availability is not due to activation of the HPA axis. During an energy deficit, exogenous leptin could not restore high frequency LH secretion when HPA function was restored to normal. Finally, the inability of leptin to increase LH secretion in the face of 2DG supports the notion that the action of leptin is dependent upon the degree of glucose availability. PMID- 11264726 TI - Purinergic regulation of intracellular Ca2+ concentration of rat pituitary folliculo-stellate cells in primary culture. AB - Pituitary folliculo-stellate cells (FSCs) are glia-like cells in the anterior pituitary and are believed to modulate the activity of the pituitary endocrine cells. However, little is known what regulates the activity of FSCs. We hypothesized that ATP could act on FSCs, because ATP is coreleased with pituitary hormones from endocrine cells. To test this possibility, we examined the effect of ATP by measuring intracellular Ca2+ concentration [Ca2+]i of FSCs in primary culture. Both ATP and UTP increased the [Ca2+]i in a concentration-dependent manner in a range between 0.1 microM and 10 microM. The response was completely suppressed by thapsigargin, an inhibitior of endoplasmic reticulum Ca2+-ATPase, and was significantly suppressed by U-73122, an inhibitor of phospholipase C. The response was also suppressed by caffeine, a blocker of IP3 receptor, whereas that was not suppressed by ryanodine, an antagonist of ryanodine receptor. These results indicate that ATP increases [Ca2+]i of FSCs by activating phospholipase C via P2Y purinergic receptor and suggest that ATP would be one of paracrine factors to FSCs in the anterior pituitary. PMID- 11264728 TI - Sensitivity and specificity of third-generation hepatitis C virus antibody detection assays: an analysis of the literature. AB - This study assessed the sensitivity and specificity of third-generation serological hepatitis C diagnostic tests from an analysis of the literature. The literature analysis was run using criteria from McMaster University for the assessment of diagnostic tests. The selected studies were grouped according to the type of population at high and low risk for hepatitis C virus (HCV) infection and to the type of reference test. The homogeneity of the sensitivity and the specificity was tested in each group using a Fisher's exact test. Of 132 studies, 10 were selected. When the estimates were homogeneous, summary point estimates and confidence intervals were computed; when the estimates were heterogeneous, subgroup analysis was performed. The sensitivity of third-generation enzyme linked immunosorbent assay (ELISA3) was estimated at 98.9% (95% CI: 94-100%) in patients with chronic liver disease and at 97.2% (95% CI: 92-99%) in panels of sera. ELISA3 specificity was found at 100% in patients with chronic liver disease. The sensitivity of the third generation recombinant immunoblot assay (RIBA3) was assessed at 78.8% (95% CI: 65-89%) in haemodialysed patients. This analysis provides evidence for the good sensitivity and specificity of ELISA3 assays particularly in high risk patient groups and confirms their use for screening in these populations. Further studies are needed to assess properly RIBA3 in general population and in risk patients. PMID- 11264727 TI - Neuropeptide-Y: a possible mediator of prolactin-induced feeding and regulator of energy balance in the ring dove (Streptopelia risoria). AB - Although neuropeptide-Y (NPY) has been widely reported to be a potent stimulator of feeding activity and regulator of energy homeostasis, most of the supportive evidence for such effects has been gathered in mammalian species. This study characterized the orexigenic potency of NPY in an avian species, the ring dove, and measured changes in hypothalamic NPY-immunoreactive (NPY-ir) cell numbers in response to energy state fluctuations or intracranial administration of the potent orexigenic hormone prolactin. Food intake was significantly elevated in male doves at 1 h after intracerebroventricular (i.c.v.) injection of 0.25 and 0.5 microg NPY but not after injection of a higher dose (1.0 microg). In time course studies, food intake was increased at 1 h after i.c.v. injection of 0.5 microg NPY but was not elevated at 2, 3, or 4 h. The number of NPY-ir cell bodies in the infundibular region of the dove hypothalamus increased two to four-fold following acute food deprivation, chronic food restriction, or repeated i.c.v. injections of prolactin. No additive effects were observed when food restriction and prolactin treatment were combined. These findings suggest that NPY is involved in energy homeostasis in doves and are consistent with the hypothesis that prolactin-induced hyperphagia is mediated in part by NPY. PMID- 11264729 TI - Cell transformation induced by hepatitis C virus NS3 serine protease. AB - Persistent infection with hepatitis C virus (HCV) may lead to hepatocellular carcinoma (HCC). It has been suggested that HCV-encoded proteins are directly involved in the tumorigenic process. The HCV nonstructural protein NS3 has been identified as a virus-encoded serine protease. To study whether HCV NS3 has oncogenic activity, nontumorigenic rat fibroblast (RF) cells were stably transfected with an expression vector containing cDNA for the NS3 serine protease (nucleotides 3356-4080). The NS3 serine protease activity was determined in the transfected cells. The transfected cells grew rapidly and proliferated serum independently, lost contact inhibition, grew anchorage independently in soft agar and induced significant tumour formation in nude mice. Cells transfected with an expression vector containing a mutated NS3 serine protease (serine 139 to alanine at the catalytic site) showed no transforming abilities; their growth was dependent on serum and they did not grow anchorage independently in soft agar. Moreover, cells transfected with the NS3 serine protease and treated with the chymotrypsin inhibitors TPCK and PMSF (a serine protease inhibitor) lost their transforming feature. These results suggest that the NS3 serine protease of HCV is involved in cell transformation and that the ability to transform requires an active enzyme. PMID- 11264730 TI - Expression of interferon-alpha subtypes in peripheral mononuclear cells from patients with chronic hepatitis C: a role for interferon-alpha5. AB - Interferon (IFN)-alpha is a family of antiviral proteins encoded by different genes. The biological significance of the existence of various IFN-alpha subtypes is not clear. We have investigated the interferon system in chronic hepatitis C virus (HCV) infection, a disease that responds to interferon-alpha2 therapy in only a limited proportion of cases. We analysed the expression of interferon regulatory factor (IRF)-1, IRF-2, and IFN-alpha subtypes in nonstimulated and Sendai virus-stimulated peripheral blood mononuclear cells (PBMC) from HCV infected patients and healthy controls. We observed that the IRF-1 mRNA and IRF 1/IRF-2 ratios were increased in PBMC from hepatitis C patients with respect to normal subjects. Sendai virus stimulation of PBMC led to a significant increase in the levels of IRF-1, IRF-2 and IFN-alpha mRNAs and in the production of IFN alpha protein with respect to basal values in healthy controls as well as in patients with HCV infection. In addition, we found that while natural HCV infection induced increased IFN-alpha5 expression in PBMC, in vitro infection of these cells with Sendai virus caused a raise in the expression of IFN-alpha8 in both patients and normal controls. In summary, our results indicate that virus induced activation of the IFN system in human PBMC is associated with selective expression of individual IFN-alpha subtypes, IFN-alpha5 being the specific subtype induced in PBMC from patients with chronic HCV infection. PMID- 11264732 TI - Evolution of hepatitis C virus genome in chronically infected patients receiving ribavirin monotherapy. AB - Recent results of clinical trials suggest that combination of interferon and ribavirin exhibits an enhanced antiviral effect in the treatment of chronic hepatitis C. To investigate the effect of ribavirin on hepatitis C virus (HCV) infection, we analysed the evolution of the genetic heterogeneity of HCV in relation to the anti-HCV humoral response in patients treated by ribavirin alone. The study population included 35 patients with liver biopsy proven chronic hepatitis C infected with HCV genotype 1. Among them, 26 were treated with ribavirin for at least 12 months and nine untreated patients served as a control group. Serum samples were analysed before and at 6 and 12 months of therapy. Three regions of the HCV genome, i.e. HVR1, a domain of NS5A including part of the interferon sensitivity determining region (ISDR), and a segment of NS5B, were amplified by RT-PCR using specific primers. The PCR products were then studied using single-strand conformation polymorphism (SSCP) analysis followed by either direct sequencing, or cloning and sequencing. In parallel, the humoral anti-E1 response was studied using an ELISA (Innotest HCV E1Ab, Innogenetics). The results of HCV genome analysis showed no significant effect on the amino acid sequence evolution of the HVR1, NS5A and NS5B regions of HCV. Analysis of a phylogenetic tree from the major quasispecies variants showed the absence of correlation with ribavirin response, and the absence of selection of viral strains during ribavirin treatment. A trend towards a decrease in the anti-E1 Ab response was also observed. Altogether these results suggest that ribavirin may not exhibit a direct antiviral effect, but may trigger a favourable response to interferon by modulating the immune response against HCV. PMID- 11264731 TI - Human hepatocytes transplanted into genetically immunocompetent rats are susceptible to infection by hepatitis B virus in situ. AB - Immune tolerance of human cells without generalized immunosuppression was created in groups of normal fetal rats at 17 days of gestation by inoculation ip with primary human hepatocytes in utero. One day after birth, suspensions of human hepatocytes were transplanted via intrasplenic injection and one week later groups of rats were inoculated with hepatitis B virus (HBV). Tolerized rats that were transplanted with human hepatocytes and subsequently infected with HBV produced hepatitis B surface antigen (HBsAg) in serum beginning on day 3. Levels rose fivefold and remained stable at 0.75 pg/ml through at least 60 days. Of cells that stained positive for human serum albumin, approximately 30% were found to be also positive for HBsAg by immunohistochemistry. Serum HBV DNA was detectable from 1 to 15 weeks postinfection. Finally, covalently closed circular DNA, reflecting HBV replication, was found in liver and serum. Controls that were tolerized and not transplanted, but inoculated with HBV, as well as untreated controls, had no evidence of HBV gene expression or replication under identical conditions. The data support the conclusion that primary human hepatocytes transplanted into genetically immunocompetent rodent hosts, survive and maintain sufficient differentiation to produce human serum albumin and be infected by HBV. PMID- 11264733 TI - HVR1 quasispecies analysis from a long-term culture of hepatitis C virus in Hep G2 derived cells grown in a haemodialysis cartridge. AB - Studies on the in vitro hepatitis C virus (HCV) infection are hampered by the lack of an appropriate system to culture permissive cells to be continuously infected with HCV. Trypsinization required for cell passage can lead to possible temporary loss of permissiveness for infection, whereas refreshment of the medium can result in loss of infectious particles necessary for perpetuation of the infection; it is therefore very difficult to maintain a continuous HCV infection in cell cultures. A new infection method was designed and evaluated in order to prevent these unfavourable circumstances. A cell line derived from the human hepatoblastoma cell line Hep G2 was grown in the extracapillary space of a haemodialysis cartridge, in the presence of a HCV-positive inoculum, while the culture medium was recirculated through the intracapillary space, supplying the cells with nutrients and oxygen. HCV RNA could continuously be detected in the cells up to 77 days of culture. Sequence analysis of the HCV hypervariable region 1 (HVR1) revealed that 56% and 75%, respectively, of the clones obtained from the cells at day 20 and 40 after start of the infection were different from the clones obtained from the original inoculum and that certain nucleotide positions in this region were more susceptible to mutations, leading to an alteration in amino acid sequence. As none of these sequences were present in the clones from the inoculum, it is suggested that new HCV quasispecies have emerged as a result of viral replication in the hepatocytes in vitro. This system seems a valuable tool for the in vitro evaluation of antiviral drugs. PMID- 11264735 TI - Phylogenetic investigation for the risk of hepatitis C virus transmission to surgical and dental patients. AB - Blood loss during treatment carries a potential risk for the transmission of blood-borne pathogens in hospital patients. To determine whether nosocomial transmission of hepatitis C virus (HCV) occurs in surgical wards and dental hospitals, we tested anti-HCV antibodies and HCV RNA in sera from these patients and analysed the hypervariable region 1 (HVR1) sequence of HCV phylogenetically in the HCV RNA-positive patients. Five of 83 patients from a surgical ward were positive for HCV RNA, and six patients from one dental hospital and nine patients from a second were found to be positive for HCV RNA during the examination period. The HVR1 sequences were amplified from these patients' serum, and after subcloning, multiple clones of the HVR1 sequence from each patient were determined. The phylogenetic analysis of these sequences showed that HVR1 species from each patient could be classified into one to three genetic clusters of HVR1 quasi-species and that these clusters were independent of each other among patients. Thus, there was no evidence of HCV transmission in our study, and unrecognized transmission of HCV may be a rare event in surgical and dental patients at university hospitals. PMID- 11264734 TI - Impact of chronic hepatitis B and interferon-alpha therapy on growth of children. AB - Interferon-alpha (IFN) has been approved as treatment for children with chronic hepatitis B (CHB). The aims of this study were to assess the impact on children's growth of the disease itself and of IFN treatment. The growth of 142 children with CHB (70 IFN-treated, 72 untreated) was monitored for a minimum of one year. Regression analysis models were used to determine which of the variables most affected children's growth. After adjusting for racial differences, the population of 142 children with CHB had a mean baseline height for age percentile of 39 and a mean baseline weight for age percentile of 38, which were significantly different (P < 0.0001) from the 50th percentiles of their respective reference populations. The height for age Z score of untreated children was inversely correlated with serum hepatitis B virus DNA and aspartate aminotransferase levels, and the weight for age Z score was inversely correlated with serum hepatitis B virus DNA levels. While undergoing IFN therapy, children displayed a "U-shaped" growth pattern, such that height for age and weight for age Z scores at 3 or 6 months were lower than scores at baseline or 12 months. In this study the average child with CHB showed compromised growth even in the absence of IFN therapy. During IFN therapy, children's growth was temporarily disrupted. PMID- 11264736 TI - Hepatitis B surface antigen variants in vaccinees, blood donors and an interferon treated patient. AB - Variants of hepatitis B virus (HBV), with amino acid substitutions in the major antigenic "a" determinant of hepatitis B surface antigen (HBsAg), have been described mainly in vaccinated children. In the present study in addition to vaccinated children, we have investigated Chinese blood donors positive for anti HBc alone, and a patient with continuing liver disease after interferon-induced seroconversion to anti-HBs. Variants were detected in two of four children with break-through infections. One child had a double mutation (P142S and G145R) and the other a G145A substitution. Three of seven anti-HBc positive Chinese blood donors had a T131I substitution, whilst the interferon-treated patient had a treble amino acid substitution (P142S, G145R and N146D). The present results indicate that HBsAg variants may exist in individuals other than vaccinated children. PMID- 11264737 TI - Influence of butyric and lactic acids on the beta-glucuronidase activity of Clostridium perfringens. AB - The beta-glucuronidase activity of intact cells of Clostridium perfringens was not influenced by the presence of either 0.09 or 0.19% lactic or butyric acids. In contrast, the enhanced enzyme activity of intact cells due to sodium deoxycholate was significantly decreased by the presence of these acids. These results suggest the possibility that the development of cancer due to the intake of a high fat diet may be inhibited by the presence of organic acids produced by intestinal bacteria. PMID- 11264738 TI - Optimization of random amplification of polymorphic DNA analysis for molecular subtyping of Escherichia coli O157. AB - Random amplification of polymorphic DNA (RAPD) analysis using the polymerase chain reaction has proved to be a useful technique in the epidemiological investigation of micro-organisms but may suffer from a lack of reproducibility in poorly optimized protocols. In this study a method of obtaining reproducible genomic fingerprints using RAPD analysis of Escherichia coli O157 is described. By systematic optimization of reaction conditions and selection of suitable primers, reproducible and discriminatory profiles could be obtained from all E. coli O157 strains tested. In addition, two other methods of obtaining reproducible profiles from E. coli O157 strains without the need to purify genomic DNA are described. PMID- 11264739 TI - Prediction of conidial germination of Penicillium chrysogenum as influenced by temperature, water activity and pH. AB - AIMS: Conidial germination of Penicillium chrysogenum was carried out under operating conditions compatible with a pastries manufacturing process. METHODS AND RESULTS: A range, limited by two experimental values, was defined for each environmental factor tested: temperature (15 or 25 degrees C), water activity (0.75 or 0.85) and pH (3.5 or 5.5). A closed device was made, which maintained an equilibrium between water activity of the culture medium and atmospheric relative humidity during 25 days, to follow spore germination. The combined effects of temperature, water activity and pH on spore germination were studied by applying factorial design methodology. CONCLUSIONS: Higher rates of spore germination were associated with a high level of water activity. The incubation temperature also had a positive effect. A significant positive interaction between water activity and temperature was observed. Under these specific experimental conditions, pH did not have a significant effect on conidial germination. SIGNIFICANCE AND IMPACT OF THE STUDY: A model describing the behaviour of fungal conidia is proposed. PMID- 11264740 TI - Susceptibility of Burkholderia cepacia and other pathogens of importance in cystic fibrosis to u.v. light. AB - To investigate the potential usefulness of u.v. germicidal irradiation (UVGI) in preventing the spread of Burkholderia cepacia, an important pathogen in cystic fibrosis (CF), the in-vitro susceptibility of B. cepacia to UVGI was determined. Five strains were exposed to UVGI from a 7.2-W source. Burkholderia cepacia was less susceptible to UVGI than other important CF-related pathogens, namely Staphylococcus aureus and Pseudomonas aeruginosa, but was more susceptible than Stenotrophomonas maltophilia. No strain of B. cepacia survived longer than an 8 s exposure to UVGI, with doses required to achieve 1 log reduction in bacterial numbers ranging from 28.3 to 57.5 J m(-2). PMID- 11264741 TI - Molecular characterization of Bacillus anthracis using multiplex PCR, ERIC-PCR and RAPD. AB - AIMS: To investigate the molecular characterization of Bacillus anthracis strains by multiplex PCR, enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) and random amplification of polymorphic DNA (RAPD). METHODS AND RESULTS: Three primers were used to amplify the cya, cap and cereolysinAB genes in the multiplex PCR. Two distinct ERIC-PCR and RAPD fragments, which separated B. anthracis into two groups, were used as probes in Southern hybridization experiments. The probes hybridized only to the cya+ B. anthracis strains identified by the multiplex PCR. Nucleotide sequence analysis of the two cloned fragments showed they were from the pXO1 plasmid of B. anthracis. CONCLUSION: Multiplex PCR simultaneously identified isolates of the Bacillus cereus group and the B. anthracis virulence factors. ERIC-PCR and RAPD, combined with the Southern hybridization analyses, differentiated B. anthracis strains and separated them from the closely related B. cereus group bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: ERIC-PCR and RAPD assay could be effective in differentiating virulent from avirulent B. anthracis. Our results also show that the amplification of the large plasmids was allowed in the ERIC-PCR and RAPD assay. PMID- 11264742 TI - Partial characterization of polyfermenticin SCD, a newly identified bacteriocin of Bacillus polyfermenticus. AB - AIMS: To characterize polyfermenticin SCD, a newly identified bacteriocin of Bacillus polyfermenticus SCD. METHODS AND RESULTS: Bacillus polyfermenticus SCD was identified as a bacteriocin producer with a bactericidal activity against Bacillus subtilis IFO 12113. Polyfermenticin SCD, named tentatively as the bacteriocin produced by B. polyfermenticus SCD, showed a narrow spectrum of activity against Gram-positive and Gram-negative bacteria, a yeast and moulds. Production of polyfermenticin SCD in a 5 l jar fermenter followed typical kinetics of primary metabolite synthesis. The antibacterial activity of polyfermenticin SCD on sensitive indicator cells disappeared completely by treatment with proteinase K, which indicates its proteinaceous nature. Polyfermenticin SCD seemed to be very stable throughout the pH range of 2.0 to 9.0, and it was relatively heat labile compared with other bacteriocins. Direct detection of polyfermenticin SCD activity on SDS-PAGE suggested that it had an apparent molecular mass of about 14.3 kDa. CONCLUSIONS: Bacillus polyfermenticus SCD produced relatively heat-labile polyfermenticin SCD with a narrow spectrum of activity. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacillus polyfermenticus SCD is a commercial probiotic which has been used for the treatment of long-term intestinal disorders. New findings on polyfermenticin SCD will be valuable in the evaluation of commercial probiotics. Polyfermenticin SCD can be used to control Bacillus spoilage organisms as a biological control agent. PMID- 11264743 TI - The survival of Escherichia coli O157:H7 in slurry from cattle fed different diets. AB - AIMS: The survival characteristics of Escherichia Coli O157:H7 were investigated in bovine slurry from cattle fed two different diets: (i) silage and (ii) silage + concentrates. METHODS AND RESULTS: Slurry samples collected from freshly agitated tanks were inoculated at a level of log10 6.0 cfu g(-1) and stored in the laboratory at 10 degrees C. Over a 12 week storage period, a 3.5 and 5.5 log reduction was observed in slurry from cattle fed a silage and silage plus concentrate diet, respectively. CONCLUSIONS: The persistence of E. coli O157:H7 in slurry over a 3 month storage period indicates its potential for transmitting the organism back into the environment. SIGNIFICANCE AND IMPACT OF THE STUDY: The discussion concludes however, that despite pathogen survival in slurry, it may not represent a major source of transmission in the farm environment. PMID- 11264744 TI - Fermentation properties of gentio-oligosaccharides. AB - AIMS: To investigate the fermentation properties of gentio-oligosaccharides (GOS), as compared to fructo-oligosaccharides (FOS) and maltodextrin in mixed faecal culture. METHODS AND RESULTS: The substrates were incubated in 24 h batch culture fermentations of human faecal bacteria. Fluorescent in situ hybridization was used to determine changes in populations of bifidobacteria, lactobacilli, clostridia, bacteroides, streptococci and Escherichia coli. Gas and short-chain fatty acid (SCFA) production was also measured. GOS gave the largest significant increases in bifidobacteria, lactobacilli and total bacterial numbers during the incubations. However, FOS appeared to be a more selective prebiotic as it did not significantly stimulate growth of bacterial groups which were not probiotic in nature. GOS and maltodextrin produced the highest levels of SCFA. Lowest gas production was seen with GOS and highest with FOS. CONCLUSIONS: GOS possessed bifidogenic activity in vitro. Although fermentation of GOS was not as selective as FOS, gas production was lower. Gas production is often seen as an undesirable side effect of prebiotic consumption. SIGNIFICANCE AND IMPACT OF THE STUDY: The study has provided the first data on fermentation of GOS in mixed faecal culture. The study has also used molecular microbiology methods (FISH) to quantify bacterial groups. The data extend our knowledge of the selectivity of fermentation of oligosaccharides by the gut microflora. PMID- 11264745 TI - A directed approach to the selection of bacteria with enhanced catabolic activity. AB - AIMS: This study was aimed at selecting catabolicly-improved bacteria by in vitro evolution using a specially designed fermentor system. METHODS AND RESULTS: To facilitate this objective, genetic variation was induced by ultraviolet irradiation, and a selective pressure was subsequently exerted by gradual increases in the concentration of organic toxins. During a pilot experiment, a culture was forced to tolerate and catabolize a mixture of phenol and formaldehyde. The population developed a high resistance against formaldehyde and the specific degradation rate increased rapidly. Biochemical analysis of the mutants revealed an increase in the expression of enzymes involved in the pathway oxidizing formaldehyde. CONCLUSIONS: The fermentor system described is, in general, suitable for the selection of bacteria with enhanced catabolic activities. SIGNIFICANCE AND IMPACT OF THE STUDY: The procedure is an alternative to conventional genetic engineering, providing efficient and genetically stable strains suitable for applications in the field of environmental biotechnology. PMID- 11264746 TI - Detection of tabtoxin-producing strains of Pseudomonas syringae by PCR. AB - AIMS: The present study describes a system based on PCR to distinguish tabtoxin producing strains of Pseudomonas syringae from other Ps. syringae plant pathogens that produce chlorosis-inducing phytotoxins. METHODS AND RESULTS: Thirty-two strains of Ps. syringae and related species were examined. Two sets of PCR primers were developed to amplify genes (tblA and tabA) required for tabtoxin production. Only a PCR product of 829 bp or 1020 bp was produced in PCR reactions with the tblA or tabA primer sets, respectively, and cells from tabtoxin producing pathovars of Pseudomonas syringae. All known non-tabtoxin producing bacterial species failed to produce an amplification product with either primer set. CONCLUSIONS: PCR of genes required for tabtoxin production is a simple, rapid and reliable method for identifying tabtoxin-producing strains of Ps. syringae. SIGNIFICANCE AND IMPACT OF THE STUDY: The protocol can effectively distinguish tabtoxin-producing strains of Ps. syringae from other Ps. syringae pathovars and Ps. syringae pv. tabaci strains from other tabtoxin-producing Ps. syringae pathovars. PMID- 11264747 TI - A comparison of immunomagnetic separation and culture, Reveal and VIP for the detection of E. coli O157 in enrichment cultures of naturally-contaminated raw beef, lamb and mixed meat products. AB - AIMS: The aims of this study were (i) to evaluate the specificity and sensitivity of three previously described PCR assays for the detection of E. coli O157 and, (ii) to compare PCR, culture, and two visual immunoassays (VIAs), BioSign and Path-Stik, for detecting E. coli O157 after enrichment culture and immunomagnetic separation (IMS) performed on various naturally contaminated raw beef, lamb and mixed meat products. METHODS AND RESULTS: Twelve sorbitol non fermenting (SNF) verocytotoxin-producing (VT+) E. coli O157, 6 SNF VT- E. coli O157, 4 sorbitol fermenting (SF) VT+ E. coli O157, 3 SF VT- E. coli O157, 23 E. coli belonging to 17 other serogroups and 12 organisms of other species were used to check the specificity of PCR reactions. Only one primer pair generated amplimers only with E. coli O157 and was used for all subsequent work. After enrichment culture and on inoculated minced beef samples, PCR was as sensitive as culture for detecting 9 of the 12 strains of E. coli O157, but up to 4 log10 more sensitive than culture for detecting three strains. Of the 120 samples of naturally contaminated meat products examined, 80 (67%) were positive by PCR, 70 (58%) were positive by BioSign, 69 (58%) were positive by culture and 67 (56%) were positive by Path Stik. Eleven samples were positive by PCR and both VIAs, but negative by culture because culture plates were heavily overgrown with SF organisms making detection of any E. coli O157 present impossible. CONCLUSIONS: PCR and both VIAs compared well with culture of beads to CT-SMAC for detecting E. coli O157 after enrichment culture and IMS. PCR appeared to be the most sensitive method, but needed specialised equipment and was also the most expensive, laborious and technically demanding technique. Although lacking the sensitivity of PCR, the VIAs were of comparable sensitivity to culture and were extremely quick and easy to perform giving a result in less than 15 minutes. SIGNIFICANCE AND IMPACT OF THE STUDY: Culture techniques may fail to detect E. coli O157 retrieved by IMS due to overgrowth with other organisms. PMID- 11264748 TI - Development of Dot-ELISA for the detection of human rotavirus antigen and comparison with RNA-PAGE. AB - AIMS: Development of a simple, specific, rapid and inexpensive Dot-ELISA test for diagnosis of rotaviral antigen in stool samples. METHODS AND RESULTS: Hyperimmune rabbit antisera raised against SA-11 (Simian Agent-11) strain was used as primary antibody. The secondary antibody conjugate used was the goat anti-rabbit IgG alkaline phosphatase, and BCIP/NBT solution was used as substrate. Faecal extracts were diluted 10-fold and used for the detection of rotavirus antigen. RNA-PAGE was performed to compare the specificity and sensitivity of the diagnostic tests. Dot-ELISA positive samples were further confirmed by Western blot analysis. CONCLUSIONS: This Dot-ELISA test could be used as an alternative method for diagnosing rotaviral samples in the field. SIGNIFICANCE AND IMPACT OF THE STUDY: The Dot-ELISA test is simple, specific, rapid and cost effective. It is suitable for identifying a large number of samples obtained from epidemiological studies and hence, reducing the death rate of rotavirus-infected patients. PMID- 11264749 TI - Lactobacillus plantarum phytase activity is due to non-specific acid phosphatase. AB - Microbial phytases suitable for food fermentations could be obtained from lactic acid bacteria isolated from natural vegetable fermentations. Phytase activity was evaluated for six lactic acid bacteria cultures. Although the highest activity was found for Lactobacillus plantarum, the phytase activity was very low. Further characterization of the enzyme with phytate-degrading activity showed a molecular weight of 52 kDa and an optimum activity at pH 5.5 and 65 degrees C. Enzyme activity was due to a non-specific acid phosphatase which had a higher hydrolysis rate with monophosphorylated compounds such as acetyl phosphate that could explain the low phytase activity. PMID- 11264750 TI - Characterization of dextran-producing Leuconostoc strains. AB - Leuconostoc strains were characterized according to their antibiotic susceptibilities, carbohydrate fermentation profiles, sucrase activity patterns and plasmid content. All the strains tested were resistant to the antibiotics sulphathiazole, trimethoprim and vancomycin, and could be separated into two groups based on whether or not they could ferment melibiose and raffinose. Six Leuconostoc strains possessed plasmids and many produced unique sucrase activity patterns in polyacrylamide gels. These data will aid in distinguishing among physiologically similar dextran-producing leuconostocs, frequently used in research and industry. PMID- 11264751 TI - Optimum conditions for yeast protoplast release and regeneration in Saccharomyces cerevisiae and Candida tropicalis using gut enzymes of the giant African snail Achatina achatina. AB - Release of viable protoplasts of Saccharomyces cerevisiae and Candida tropicalis was achieved using fresh crude enzyme extracts of the giant African snail Achatina achatina. Optimum results of 2.8 x 10(6) protoplast ml(-1) were obtained when 1 g (wet wt) of cell slurry from the yeast strains was first treated with 1% beta-mercaptoethanol for 10 min and incubated with the undiluted crude enzyme for 120 min using 1.0 mol l(-1) sorbitol as osmotic stabilizer. Protoplast yield was enhanced with higher enzyme concentrations, longer digestion times and treatment of cells with beta-mercaptoethanol. Percentage regeneration of protoplast to viable cells in isotonic medium containing 0.01 mol l(-1) CaCl2 was in the range of 52-77%. These findings could be useful in the genetic manipulation of yeast of industrial importance. PMID- 11264752 TI - Copper sorption by native and modified pellets of wood-rotting basidiomycetes. AB - AIMS: The aim of this study was to investigate the biosorption of copper to the pellets of different wood-rotting fungal species. METHODS AND RESULTS: Copper sorption was studied in both batch and column arrangements. The optimum pH for copper sorption was between 3.5 and 4. In 100 mg l(-1) Cu (II), maximum qe values were found for Oudemansiella mucida (8.77 mg g(-1) dry wt), Lepista nuda (6.29 mg g(-1)), Pycnoporus cinnabarinus (5.08 mg g(-1)) and Pleurotus ostreatus (4.77 mg g(-1)). Both biomass yield and specific sorption were influenced by the composition of the fermentation broth. The results of column experiments showed that mycelial pellets of wood-rotting fungi can be considered as promising biosorbent material. CONCLUSIONS: Pellets of wood-rotting fungi showed the same or better copper sorption properties as those previously reported for lower fungi or filamentous bacteria, as well as good mechanical properties. PMID- 11264753 TI - Taxonomy of obligately homofermentative and facultatively heterofermentative lactobacilli in pig faeces. AB - AIMS: Identification and characterization of obligately homofermentative and facultatively heterofermentative strains of Lactobacillus spp. isolated from the faeces of pigs that had been raised under different conditions. METHODS AND RESULTS: The phenotypic relatedness of the isolated strains and reference strains were determined by numerical analysis of total soluble cell protein patterns and simple physiological and biochemical tests. Of the 23 strains isolated from faeces, nine were obligately homofermentative and 14 facultatively heterofermentative. The strains clustered at r > or = 0.61 with Lactobacillus amylovorus (seven strains), Lactobacillus crispatus (one strain), Lactobacillus plantarum (14 strains) and Lactobacillus intestinalis (one strain). CONCLUSIONS: Results obtained from the physiological and biochemical tests confirmed the identity of the isolates as determined by numerical analysis of total soluble cell protein profiles. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the association of Lact. crispatus and Lact. intestinalis with the gastro-intestinal tract of pigs. PMID- 11264754 TI - Effects of chilling on sampling of bacteria attached to swine carcasses. AB - Two microbiological sampling techniques, excision and sponge swabbing, were compared by determining counts of aerobic bacteria, coliforms and injured coliforms from 20 de-haired swine carcasses before and after chilling. Excised jowl skin produced significantly greater counts of the three types of bacteria than sponge swabs. Aerobic bacteria, coliforms and injured coliforms recovered by sponge swabbing carcasses before chilling were 11.6%, 0.9% and 11.0% of excised samples, respectively; the corresponding percentages recovered after chilling were 23.9%, 11.1% and 5.0%. Numbers of all bacteria present on the post-chill carcasses were substantially lower than on the pre-chill carcasses. Excision usually produced more countable plates for coliforms and injured coliforms on chilled carcasses than sponge swabbing and therefore, is more suitable in estimating low numbers of faecal bacteria on chilled carcasses. To explore the possible structural bases for these findings, skin samples were inoculated with 10(2)-10(7) cfu cm(-2) faecal bacteria and examined by scanning electron microscopy. Chilled samples showed bacteria and biofilm embedded in superficial crevices, which underlies a possible reason for the lower recovery of bacterial cells by the sponge swabbing. The study indicates that the differences between sampling techniques may be a result of the chilling process of swine carcasses. PMID- 11264755 TI - Sensitive plate assay for screening and detection of bacterial polyurethanase activity. AB - AIMS: A plate assay to screen and detect bacterial polyurethanase in agar medium containing a colloidal polyester-polyurethane and rhodamine B is presented. METHODS AND RESULTS: Substrate hydrolysis causes the formation of orange fluorescent halos visible upon u.v. irradiation. The logarithm of polyurethanase activity from a purified polyurethanase protein is linearly correlated with the diameter of halos, thereby allowing quantification of polyurethanase activities ranging from 0.81 to 7.29 Units. CONCLUSIONS: The potential advantages of this system are in identification and recovery of viable polyurethanolytic bacteria and quantification of polyurethanase activity. SIGNIFICANCE AND IMPACT OF THE STUDY: These advantages are derived largely from the intense fluorescence observed due to the hydrolysis of substrate reacting with rhodamine B allowing for the use of low substrate concentrations and corresponding decrease in time required detecting low levels of enzyme activity. PMID- 11264756 TI - Allergic bronchopulmonary aspergillosis: new concepts of pathogenesis and treatment. AB - Allergic bronchopulmonary aspergillosis (ABPA) is a condition that results from a hypersensitivity reaction to the fungus Aspergillus fumigatus. The purpose of the present review is to examine the pathogenesis of this condition and the evidence for treatments available. Allergic bronchopulmonary aspergillosis is characterized by an intense airway inflammation with eosinophils and the formation of mucus plugs. Clinically, there are periods of exacerbation and remission that may lead to proximal bronchiectasis and fibrotic lung disease. New evidence confirms the role of intense airway inflammation with eosinophils, but also suggests a role for interleukin (IL)-8/neutrophil-mediated inflammation in this process, and the potential deficiency of anti-inflammatory cytokines such as reduced IL-10. Treatment for ABPA has so far focused on corticosteroids to suppress eosinophilic airway inflammation. An expanding knowledge of the pathology of ABPA also suggests other therapies may be of potential benefit, particularly the use of azole antifungal agents. Allergic bronchopulmonary aspergillosis is itself an important complication of asthma and cystic fibrosis. A greater understanding of the condition is required to improve management and well-designed clinical trials need to be carried out to critically assess new and current treatments. In addition, the information gained from the studies of its pathogenesis has the potential to benefit our understanding of the disease processes in asthma and bronchiectasis. PMID- 11264757 TI - Influence of body fat distribution on oxygen uptake and pulmonary performance in morbidly obese females during exercise. AB - OBJECTIVE: The aim of this study was to determine the effects of fat distribution on aerobic and ventilatory response to exercise testing in morbidly obese (MO) females. METHODOLOGY: The study population consisted of 164 MO females, 55% (n = 90) with upper body or abdominal adiposity (UBD), as defined by waist-hip circumference ratio (WHR) > or = 0.80, and 45% (n = 74) with lower body fat distribution (LBD) (WHR < 0.80). An incremental exercise testing on cycle ergometer was performed to determine the effect of exercise on oxygen consumption (VO2), carbon dioxide production (VCO2), minute ventilation (VE), tidal volume (VT), respiratory rate (fb) and heart rate (HR). RESULTS: Upper body adiposity individuals had significantly higher VO2 and VCO2 than LBD subjects (P < 0.05) from 0 watt (W) of pedalling up to their anaerobic threshold (AT) and maximal exercise. VE was significantly higher in UBD subjects compared with LBD subjects, from 20 W during exercise up to AT and peak work levels (P < 0.05). Upper body adiposity group also had a significantly higher fb than the LBD group at rest, after each workload and at AT and peak exercise work rates (P < 0.05). VT was lower in UBD subjects at free pedalling and up to AT and peak workload with significant difference at 60 and 80 W (P < 0.05). The anaerobic threshold, expressed as work rate, was significantly lower in the UBD subjects (P < 0.05) and peak workload achieved did not differ significantly between the two groups. CONCLUSIONS: Upper body adiposity subjects had higher oxygen requirement, more rapid and shallow breathing, higher ventilatory demand, but lower anaerobic threshold than the LBD individuals during progressive exercise. It suggests that the cardiopulmonary endurance to exercise in MO patients with upper body fat distribution is lower than in those with lower body fat distribution. PMID- 11264758 TI - Clinical effects of pranlukast, an oral leukotriene receptor antagonist, in mild to-moderate asthma: a 4 week randomized multicentre controlled trial. AB - OBJECTIVE: Leukotriene antagonists are increasingly used in asthma management. Pranlukast is a new, orally active, selective inhibitor of CysLt1 leukotriene receptor. The present clinical trial was performed to study the effect and safety of pranlukast in mild-to-moderate asthma. METHODOLOGY: A randomized, double blind, placebo-controlled, parallel group study was performed in eight medical centres in Korea. Mild-to-moderate asthma patients who had been treated with beta2-agonists and/or inhaled corticosteroids were studied. The patients' symptoms were evaluated by asthma diary and twice-daily peak flow monitoring. RESULTS: Of the 206 patients enrolled, 197 were eligible for analysis. The pranlukast group (n = 98) showed statistically significant improvement in asthma symptoms, including asthma attack rate, daily living score, and morning and evening asthma scores. Pranlukast significantly reduced the consumption of beta2 agonist. Compared with the placebo group, forced vital capacity (FVC) and forced expiratory volume in 1 s (FEV1) were not significantly higher in the pranlukast group. Morning and evening peak expiratory flow (PEF) were significantly increased after pranlukast treatment at weeks 2 and 4 (380.8 +/- 10.1 L/min at baseline, 394.5 +/- 10.1 at week 2, 396.3 +/- 10.4 at week 4). There were no serious adverse reactions. CONCLUSION: Pranlukast, an oral leukotriene antagonist, was well tolerated and was effective for the management of mild-to moderate asthma. PMID- 11264759 TI - Penicillin resistance in Streptococcus pneumoniae in Isparta. AB - OBJECTIVE: The first case reports of infection with penicillin-resistant pneumococci were made in Australia in 1967 and South Africa in 1977. Since this time the increasing emergence of penicillin- resistant strains of Streptococcus pneumoniae have been a serious therapeutic problem. Therefore, the aim of the present study was to determine the penicillin resistance of S. pneumoniae strains isolated in the laboratory. The effect of procaine penicillin treatment against these strains was also investigated. METHODS: Sensitivity testing was done by disc diffusion method using oxacillin discs. Minimal inhibitory concentration (MIC) values were determined in tests with penicillin by the use of E-test (AB Biodisc, Solna, Sweden). Patients were treated with 2 x 800,000 U of i.m. procaine penicillin every 12 h for 10 days. RESULTS: Thirty-seven strains of S. pneumoniae were isolated from the sputa of adult patients who had pneumonia. Moderately resistant (0.12-1.00 microg/mL) and penicillin-sensitive (< or = 0.06 microg/mL) strains were identified in nine (24.3%) and 28 (75.7%) isolates, respectively. There were no high-level penicillin-resistant strains in the study. There was no therapeutic failure. CONCLUSION: These results suggest that procaine penicillin may still be useful in the empirical therapy of pneumococcal pneumonia. PMID- 11264760 TI - Efficacy of budesonide Turbuhaler compared with that of beclomethasone dipropionate pMDI in Japanese patients with moderately persistent asthma. AB - OBJECTIVE: The aim of the study was to compare the efficacy and safety of budesonide Turbuhaler with that of beclomethasone dipropionate (BDP) pMDI. METHODOLOGY: Three hundred and fifty adult asthma patients (mean age 52.7 years, mean baseline morning peak expiratory flow (PEF) 294 L/min (< 80% predicted normal)), taking BDP via pressurized metered-dose inhaler (pMDI), 400 microg daily for at least 2 months, were randomized in an open 6 week study to receive daily doses of either budesonide 100 microg or 400 microg twice daily via Turbuhaler or continued treatment with BDP, 100 microg four times daily. The primary efficacy variable was the mean change in morning PEF from baseline to the end of treatment. Outcome was also assessed using symptom scores and investigators' assessments employed in Japanese clinical trials. RESULTS: At the end of the 6 week treatment period, mean morning PEF improved significantly from baseline in both budesonide groups, 16 L/min and 33 L/min in the 200 microg and 800 microg groups, respectively, but not in the BDP group, 5 L/min. There was no significant difference between 200 microg budesonide and 400 microg BDP treatment in the effect on PEF (P = 0.29), but 800 microg budesonide was significantly superior to BDP (P < 0.001). Final assessment of improvement and usefulness ratings showed that both budesonide treatments were significantly superior to BDP (P < 0.001). All treatments were well tolerated. CONCLUSION: Budesonide Turbuhaler (200 microg) was as effective as 400 microg BDP pMDI. The efficacy of budesonide was improved significantly by increasing the dosage to 800 microg daily. The study design shows the importance of including a higher dose treatment group when comparing two formulations of inhaled corticosteroids in order to determine whether the treatments to be compared are on the steep part of the dose response curve. Without that information, comparative studies are usually inconclusive. PMID- 11264761 TI - Case management may reduce length of hospital stay in patients with recurrent admissions for chronic obstructive pulmonary disease. AB - OBJECTIVES: The aim of the study was to determine whether the case management of patients with recurrent hospital admissions for chronic obstructive pulmonary disease (COPD) can reduce hospital days without reducing quality of life. METHODOLOGY: Sixteen subjects (mean forced expiratory volume in 1 second; FEV1 0.64 L) with at least four admissions for COPD in the previous 2 years were case managed by a clinical nurse specialist. Admissions and hospital bed days were recorded before and after the introduction of case management, and compared with data for 16 controls at another hospital who received usual care. Quality of life was measured serially in the case-managed group. RESULTS: In the first year of case management, the number of hospital bed days fell to eight per patient from 22 per patient in the previous year. This was mainly due to a reduction in the length of stay from 5.6 to 3.5 days. In the control group length of stay did not change. Admissions in both groups declined. Case-managed patients had a significant improvement in their quality-of-life scores. CONCLUSIONS: In a group of patients with severe COPD and recurrent admissions, case management reduced the number of days in hospital while improving the quality of life. These findings need to be confirmed in a randomized, controlled trial. PMID- 11264762 TI - Enteral feeding in stable chronic obstructive pulmonary disease patients. AB - OBJECTIVE: The study aimed to compare the effectiveness of a defined formula diet with a blenderized diet on nutritional and respiratory function parameters and to determine the bacteriological load of the two formulations. METHODOLOGY: Seventeen patients, aged 50-75 years, admitted to the University of the Philippines-Philippine General Hospital for chronic bronchitis and/or emphysema, were studied. They were divided into two groups according to dietary regimens. Each group of patients received either the standardized commercial formula or the blenderized formula for 2 weeks. Evaluation of dietary intake, anthropometric measurements, laboratory examinations and lung function were assessed. Subjective evaluation (patient's and physician's assessment) was also sought. Microbiological examinations were performed on the prepared enteral formulas. RESULTS: There was a slight increase in weight and in pulmonary function in both groups but these results did not differ significantly. Possible formula contamination was confirmed. Furthermore, in the overall assessment, the physician and patients rated both formulas as comparable. PMID- 11264763 TI - New quinolone, grepafloxacin, inhibits Cl- secretion across bovine airway epithelium in culture. AB - OBJECTIVE: Transepithelial ion transport plays an important role in the regulation of the amount and the rheological properties of bronchial secretion. The effect of grepafloxacin (GPFX), a new quinolone agent, on bioelectrical properties of airway epithelium was determined. METHODOLOGY: Electrical properties of bovine tracheal epithelium cultured under an air-liquid interface condition were measured by the short-circuit technique. RESULTS: Addition of GPFX (100 microg/mL) to the mucosal side decreased short-circuit current (Isc) from 14.4 +/- 1.3 to 5.6 +/- 0.6 microA/cm2 (P < 0.001), and the response was accompanied by corresponding decreases in transepithelial potential difference and cell conductance. This effect was concentration dependent, and a similar response was also noted when GPFX was added to the submucosal side. The GPFX induced decrease in Isc was not altered by the Na+ channel blocker amiloride, but was inhibited by the Cl- channel blocker diphenylamine-2-carboxylate or Cl(-) free medium (P < 0.001, in each case). Furthermore, GPFX reduced Cl- conductance (P < 0.01) without affecting Na+ conductance of the epithelium. CONCLUSIONS: Grepafloxacin selectively inhibits Cl- secretion across tracheal epithelial cells, which may result in the inhibition of water secretion and, hence, the reduction of airway secretion. PMID- 11264764 TI - Indomethacin induced bulky lymphadenopathy and eosinophilic pneumonia. AB - Indomethacin is one of the most popular non-steroidal anti-inflammatory drugs (NSAID). Although NSAID occasionally provoke bronchospasm and hypersensitivity pneumonia, they seldom cause lymphadenopathy. This is the first report in which NSAID induced both eosinophilic pneumonia and bulky intrathoracic lymphadenopathy simultaneously. A 76-year-old Japanese man experienced high fever and dyspnoea after using an indomethacin suppository. Computed tomography scan of his chest revealed massive mediastinal and hilar lymphadenopathy along with diffuse infiltration in both lungs. He was diagnosed to have eosinophilic pneumonia because of eosinophilia in his peripheral blood and bronchoalveolar lavage fluid (BALF). Without using glucocorticoids, the pulmonary infiltration and lymphadenopathy subsided spontaneously. As the blastoid transformation test using the lymphocytes in his BALF was positive to indomethacin, we judged that both his eosinophilic pneumonia and mediastinal lymphadenopathy were due to a hypersensitivity reaction to indomethacin. An allergic reaction to NSAID should be considered as a rare cause of mediastinal lymphadenopathy. PMID- 11264765 TI - Pulmonary presentations of amyloidosis. AB - Respiratory tract involvement with amyloid is rare. We report eight cases of lower respiratory tract amyloidosis including a case of isolated pulmonary interstitial amyloidosis treated with chemotherapy, two cases of recurrent endobronchial amyloid with airway obstruction successfully treated with laser therapy and three cases of localized nodular pulmonary amyloidosis. The subjects with endobronchial and nodular amyloid demonstrated good long-term survival, while those with systemic or interstitial pulmonary amyloid had progressive disease and poor survival. Circulating monoclonal immunoglobulins were identified in five of the eight cases as the likely cause of the amyloid. PMID- 11264766 TI - Tuberculosis in Thailand. AB - Tuberculosis (TB) was expected to be eradicated by the end of this century. However, an increasing incidence of tuberculosis in many parts of the world has led to renewed interest in the disease. The pandemic of HIV infection has changed TB, an endemic disease, to an epidemic worldwide. In Thailand, tuberculosis cases and deaths reduced year after year, until 1992 when the cases began to increase as a result of HIV infection. The annual risk of infection in 1997 was estimated at 1.4%, with approximately 100 000 new TB cases developing each year. Fifteen per cent of tuberculosis patients are seropositive for HIV infection. Increasing antituberculosis drug resistance has been correlated with the high prevalence of HIV infection in some parts of the country. In 1995, cure rate of this disease was approximately 50% and, since 1996, in order to cope with the worsening situation, the National Tuberculosis Programme (NTP) has adopted Directly Observed Treatment, Short-course (DOTS). Despite the current economic turmoil of the country, the programme has now been expanded to cover over 400 of the 810 districts of Thailand. Also, the economic effects of tuberculosis at the household level in Thailand were recently studied. Tuberculosis is a chronic disease that commonly affects the lower socioeconomic classes. Some patients were unable to follow the treatment regimens because of the financial burden. The low case detection and treatment completion rates are, in part, due to the inability of poor patients to cope with the expenditure. PMID- 11264767 TI - The incidence of tuberculosis in a cohort of South-East Asian refugees arriving in Australia 1984-94. AB - We have used record linkage analysis to describe the incidence of tuberculosis in a cohort of 24 652 predominantly south-east Asian refugees who arrived in Sydney, Australia during the period 1984 to 1994. Cases that had been registered with the State Department of Health were confirmed by examination of case records. After an average follow-up interval of 10.3 years there were 189 cases of tuberculosis, equivalent to an average incidence rate of 74.9 cases per 100 000 person-years. The highest incidence rate was in 40-49 year olds and 47% of cases were in women. One hundred and twenty seven cases (67%) were pulmonary and, of these, 64 (50%) were direct smear positive. The incidence of tuberculosis in this cohort is similar to that observed among Vietnamese migrants to Australia and the USA and substantially higher than the incidence among people born in Australia. It is important to maintain awareness of the diagnosis of tuberculosis, especially in countries such as Australia, where the incidence in the general population is low but where there are large populations of migrants and refugees in whom a higher incidence is expected. PMID- 11264768 TI - Tuberculosis problems in the Asia-Pacific region. AB - Tuberculosis (TB) is the top killer of the productive age group in developing countries. More than half of cases in the world occur in Asia-Pacific region. The number of cases will increase in the next decades due to increase in urban poor population and HIV incidence, and poor access to health services and poor TB programme. The mainstay of effective strategy is to promote DOTS. DOTS population coverage is high, 58% in the Western Pacific Region but low; 29% in the southeast Asia region. Even among intermediate TB-burden countries in Asia, reduction of incidence has stagnated in recent years because of aging population, health problems in the urban poor population and influx of populations from high endemic areas. Indicators of successful TB programmes are high cure rate and low drug resistance rate. There is a strong correlation between the overall quality of TB control in the past and the current primary drug-resistant rate. To solve these problems, priority should be given to nationwide implementation of DOTS in high TB-burden countries with emphasis on ensuring availability of free anti-TB drugs and strengthening primary health care. General hospitals and private sector should be involved in control programmes to prevent drug resistant cases. PMID- 11264769 TI - Nitric oxide modulates interleukin-1beta and tumour necrosis factor-alpha synthesis, and disease regression by alveolar macrophages in pulmonary tuberculosis. AB - Pretreatment with nitric oxide synthase (NOS) inhibitors profoundly increases mortality, bacterial burden and pathological tissue damage in mice infected with Mycobacterium tuberculosis. Nitric oxide (NO) production is enhanced in alveolar macrophages (AM) of tuberculosis (TB) patients. Interleukin (IL)-1beta and tumour necrosis factor (TNF)-alpha released from AM are involved in the immune response to mycobacterial infection. The aim of the present study was to examine whether NO is implicated in IL-1beta and TNF-alpha synthesis by AM and related to the resolution of disease activity in TB patients. Purified AM were retrieved by bronchoalveolar lavage from TB patients and normal subjects, and cultured in the presence or absence of a NO inhibitor, NG-monomethyl-L-arginine (L-NMMA). The release of IL-1beta and TNF-alpha, and their mRNA expression were determined by enzyme-linked immunosorbent assay (ELISA) and northern analysis, respectively. The level of nitrite released into the culture medium was determined. The rate of disease regression was evaluated by serial chest radiography. The release of nitrite, IL-1beta and TNF-alpha was much greater from AM of TB patients than normal subjects. NG-monomethyl-L-arginine inhibited the production of nitrite as well as IL-1beta and TNF-alpha in TB patients. The mRNA expression for IL-1beta and TNF-alpha was upregulated in TB patients and was depressed by L-NMMA. Immunocytochemistry using a monoclonal antibody against nuclear factor-kappaB (NF kappaB) subunit p65 showed NF-kappaB was highly expressed and translocated to the nuclei of AM in TB patients, and was inhibited by L-NMMA. An inhibition of NF kappaB by pyrrolidine dithiocarbamate attenuated IL-1beta and TNF-alpha synthesis. More generation of NO from cultured AM increased the disease regression in TB patients. We conclude that the enhanced NO generation by AM of TB patients may play an autoregulatory role in amplifying the synthesis of pro inflammatory cytokines, probably through the activation of NF-kappaB. Nitric oxide may also play an important role in resistance to M. tuberculosis infection. PMID- 11264771 TI - Dialysis-related amyloidosis: history and clinical manifestations. AB - Dialysis-related amyloidosis (DRA) or beta(2)-microglobulin amyloidosis (A beta(2)M) is a unique type of amyloidosis that has been described in individuals with both long-standing chronic renal disease and end-stage renal disease (ESRD). It has been associated with serious complications that significantly add to the morbidity of long-term dialysis patients. The deposition of beta(2)M in amyloid fibrils in various joint and osteoarticular surfaces leads to the clinical complaints and findings typical of this disorder. However, a visceral form with systemic organ involvement has also been described. Despite advances in the understanding of this disorder and in the delivery of dialysis, the ability to alter the incidence of DRA and its course remains uncertain. PMID- 11264772 TI - Pathological aspects of beta(2)-microglobulin amyloidosis. AB - Histology remains the gold standard to diagnose beta(2)-microglobulin amyloidosis (A beta(2)M). Two diagnostic criteria are required: positive Congo red staining with typical birefringence under polarized light and immunostaining of amyloid deposits with a labeled anti-beta(2)M antibody. A beta(2)M is preferentially located in the joints. Small deposits are also found in various organs, mainly the heart and gastrointestinal tract. Pathologic studies have demonstrated a high prevalence of articular A beta(2)M early in the course of hemodialysis and peritoneal dialysis, antedating clinical manifestations by several years. The stages of beta(2)M amyloid formation have been delineated: beta(2)M amyloid deposits first on the surface of the cartilage, in the absence of macrophages (stage 1), and subsequently involves capsules and synovia (stage 2), with eventual recruitment of macrophages around large beta(2)M amyloid deposits (stage 3). Clinical manifestations are likely associated with the inflammation observed in stage 3. The factors triggering the fibrillar precipitation of beta(2)M remain unknown. Macrophages do not play a role: their presence is the consequence rather than the cause of beta(2)M amyloid deposits. Several substances coprecipitated with beta(2)M amyloid have been incriminated: highly sulfated glycosaminoglycans such as chondroitin or keratan sulfate, antiproteases such as alpha(2) macroglobulin, and apolipoprotein E. As yet, no definitive conclusion has been reached. PMID- 11264773 TI - Imaging techniques in the diagnosis of dialysis-related amyloidosis. AB - beta(2)-microglobulin amyloidosis (A beta(2)M) is a major determinant of morbidity in patients on dialysis treatment. Symptoms of A beta(2)M amyloid are mainly related to (peri-)articular amyloid deposition. Imaging techniques [i.e., joint ultrasonography, X-ray, computed tomography (CT), or magnetic resonance imaging (MRI) findings], as well as conventional bone scans, are helpful in the screening of local lesions but are relatively nonspecific and/or not sensitive enough. Scintigraphic techniques using radiolabeled serum amyloid P component (SAP) or the radiolabeled A beta(2)M precursor protein, beta(2)M, generate more specific results. A beta(2)M deposits have been visualized in several long-term hemodialysis patients by using (123)I-labeled SAP. However, this scan did not show tracer accumulation in some frequently involved sites such as hips or shoulders, and frequently labeled the spleen, which is usually spared from A beta(2)M deposits. Improvements in technical sensitivity and specificity could be achieved by scanning with (131)I-labeled beta(2)M: this technique detected tracer accumulations corresponding to the typical distribution pattern of A beta(2)M. Further, both the radiation exposure and the optical resolution of this latter scan have been refined by substituting (111)In for (131)I. In a final step we generated recombinant human beta(2)M (rh beta(2)M). While (111)In rh beta(2)M again failed to show significant tracer accumulation over joint regions in patients on short-term hemodialysis without evidence of A beta(2)M, local tracer accumulations similar to those observed with natural, (111)In-labeled beta(2)M could be demonstrated in long-term hemodialysis patients with evidence of A beta(2)M. In conclusion, scintigraphy for A beta(2)M with (111)In-labeled rh beta(2)M provides a homogeneous and safe recombinant protein source and represents a suitable detection method of beta(2)M amyloid deposits in dialysis patients. PMID- 11264774 TI - Nontransplant therapy for dialysis-related amyloidosis. AB - There is no specific treatment for dialysis-related amyloidosis (DRA). Available therapy is directed at removal of large quantities of beta(2)-microglobulin (beta(2)M) and palliation of symptoms. Plasma concentrations of beta(2)M in end stage renal disease (ESRD) depend on the degree of residual renal function, the type of blood purification therapy, and properties of the dialysis filtration membrane. Retention of beta(2)M appears to be a necessary, although not sufficient, condition for DRA. While preserving residual renal function is important, dialysis modality largely determines beta(2)M removal. Convective dialysis treatments (hemofiltration and hemodiafiltration) remove beta(2)M more efficiently than diffusive treatments (conventional dialysis). In addition, column adsorption of beta(2)M can extensively remove the molecule, as can nocturnal hemodialysis. Hemodialysis membrane properties that are particularly important with regard to beta(2)M removal include permeability, adsorptive capacity, and biocompatibility. As such, beta(2)M removal with highly permeable biocompatible membranes such as polysulfone and polyacrylonitrile is relatively large. Several studies have suggested that use of such membranes can significantly delay DRA development and may be useful in ameliorating DRA associated symptoms. Non-dialysis-related therapy for DRA is palliative and includes both medical and surgical therapies. Medical therapy includes low-dose corticosteroids and nonsteroidal anti-inflammatory drugs (NSAIDs). Surgical therapy consists of relief of carpal tunnel syndrome, or palliation of shoulder pain, destroyed weight-bearing joints, or spinal cord compression. DRA is a serious complication of long-term dialysis. It is important for nephrologists to recognize the condition and attempt to slow its progression. PMID- 11264775 TI - Dialysis-related amyloidosis after renal transplantation. AB - There is no effective therapy for dialysis-related amyloidosis (DRA). The restoration of renal function with a functional graft seems the most reasonable therapy for this disturbing disease. Although there are not many prospective series on the efficacy of renal transplantation for DRA, all the studies agree that most of the patients experienced a significantly clinical improvement of the articular symptoms after a successful renal transplantation. Nevertheless, radiologic (bone cysts) and histologic lesions did not disappear after long-term follow-up. PMID- 11264777 TI - Effect of hemodialysis membranes on beta 2-microglobulin amyloidosis. AB - Early after the identification of beta(2)-microglobulin amyloidosis (A beta(2)M) as the cause of carpal tunnel syndrome, it was thought that hemodialysis was a major cause in the development of the disease. It was subsequently shown that hemodialysis was not necessary for the development of dialysis-related amyloidosis; however, it was believed that the different dialysis membranes did modulate the progression of the disease. Current data demonstrate that hemodialysis fails to prevent or reverse the disease, but there is substantial evidence that high-flux, high-efficiency dialyzers slow its progression. Many factors related to hemodialysis have been evaluated in relation to A beta(2)M, including the effect of the bioincompatibility of the membrane, the capacity of the different membranes to remove beta(2)M, and the effect of reuse on beta(2)M levels. Moreover, there have been intensive efforts to evaluate, explore, and improve the different mechanisms in beta(2)M removal, with adsorption as a promising prospect. With the available evidence, it seems that the removal of beta(2)M by the membrane plays the most important role in modulating the disease outcome and rate of progression, although a large, long-term, multicentered and randomized study is still lacking to prove this relationship. However, it is possible that with the continuing advances in optimizing the beta(2)M removal efficiency of the different membranes, the frequency and severity of the disease can be substantially decreased. PMID- 11264776 TI - Modalities for the removal of beta 2-microglobulin from blood. AB - The long-term accumulation of beta(2)-microglobulin (beta(2)M) in patients with kidney failure results in a debilitating condition referred to as dialysis related amyloidosis (DRA). There have been few methods specifically designed to remove the large quantities of beta(2)M that are produced by the body. This article briefly reviews current modalities and concepts for the removal of beta(2)M from blood. The various approaches are classified according to the mechanism of beta(2)M clearance. The potential application of immunoadsorption, a biologically specific approach to remove macromolecules, in the treatment and understanding of DRA is discussed. PMID- 11264778 TI - Role of beta(2)-microglobulin in the immune response in renal osteodystrophy. AB - Renal osteodystrophy is the major cause of skeletal morbidity in dialysis patients. It is characterized by beta(2)-microglobulin (beta(2)M) amyloid deposition at the osteoarticular sites and a destructive arthropathy. beta(2)M is present on the surface of all nucleated cells as the small extracellular subunit of the major histocompatibility complex (MHC) class I molecule and actively participates in the immune response. Accumulating evidence suggests that beta(2)M plays a key role in the development of renal osteodystrophy through a T cell mediated inflammatory immune mechanism. PMID- 11264779 TI - Dynamic of beta(2)-microglobulin fibril formation and reabsorption: the role of proteolysis. AB - Dialysis-related amyloidosis (DRA) is caused by the deposition, in target tissues, of beta(2)-microglobulin (beta(2)M) in fibrillar conformation. Several reports indicate that fibrillar beta(2)M is chemically heterogeneous and such heterogeneity is partially related to the presence of truncated species of the protein. In association with the full-length species, a beta(2)M isoform lacking six N-terminal residues is present in all the samples of our collection of ex vivo fibrils. The pattern of proteolytic cleavage in amyloidosis and in other diseases is completely different, as demonstrated by the absence in fibrillar beta(2)M of the cleavage at lysine 58, which is contrary to that described in rheumatoid arthritis and other diseases. The role of limited proteolysis of beta(2)M in the pathogenesis of the disease is uncertain. However, we have shown that the apparently minor modification of the intact protein, such as the removal of N-terminal hexapeptide, is capable of dramatically affecting its stability, protection from proteolytic digestion, and enhance its capacity to make in vitro amyloid fibrils. The structure, folding dynamic, and function of the truncated species of beta(2)M, peculiar of DRA, could shed new light on the mechanism of beta(2)M fibril formation and reabsorption. PMID- 11264780 TI - Dialysis-related amyloidosis: pathogenesis focusing on AGE modification. AB - Dialysis-related amyloidosis (DRA) is a serious complication in long-term dialysis patients, and presents with carpal tunnel syndrome, cystic bone lesions, destructive spondylarthropathy, diffuse arthritis and periarthritis, systemic organ involvement, and dialysis-related spinal canal stenosis (DSCS). Recently a new concept of DSCS has been proposed that includes both destructive spondylarthropathy and myeloradiculopathy induced by extradural thickness. beta(2)-microglobulin (beta(2)M) amyloid was demonstrated to be modified with advanced glycation end products (AGEs) such as imidazolone, N(epsilon) (carboxymethyl)lysine (CML), and pentosidine. Imidazolone is a reaction product of arginine residue in proteins with 3-deoxyglucosone (3-DG), which is markedly accumulated in uremic serum. Imidazolone is generated under nonoxidative conditions, while CML and pentosidine are formed by oxidative processes. Immunoelectron microscopy demonstrated that AGEs were localized not only in dialysis amyloid but also in nonamyloid collagenous structures, supporting the hypothesis that AGE modification of collagen might have pathogenic relevance in the deposition of beta(2)M on collagen. Serum levels of AGEs are increased in uremic patients. The dimeric form of beta(2)M in the dialysate and urine of uremic patients is more susceptible to imidazolone modification as observed in dialysis amyloid. However, the major component of dialysis amyloid is a native form of beta(2)M, while AGE-modified beta(2)M and truncated beta(2)M are the minor components. Thus I propose that 3-DG and the other dicarbonyl compounds accumulating in uremic serum promote the modification of beta(2)M with AGEs mainly after deposition of beta(2)M as amyloid. For the prevention and treatment of DRA, beta(2)M should be efficiently eliminated from circulating blood by kidney transplantation, hemodialysis, or hemodiafiltration using high-flux membranes and an adsorbent (Lixelle) column. PMID- 11264781 TI - The role of the synovium and cartilage in the pathogenesis of beta(2) microglobulin amyloidosis. AB - The predilection for beta(2)-microglobulin (beta(2)M) amyloid deposition in articular structures is unique compared to other forms of amyloid; this article focuses on possible pathogenic mechanisms. The synovium and/or cartilage appear to be important in the pathogenesis of beta(2)M amyloidosis (A beta(2)M), as amyloid is not found in the shafts of long bones. The concentration of beta(2)M in the joint fluid parallels that in serum. Once in the joint space, evidence suggests that the beta(2)M binds to collagen in cartilage as the initial site of deposition. This binding may serve as the first step in subsequent amyloid formation, although this remains to be proven. beta(2)M has been shown to have many direct effects on synovial fibroblasts, including induction of the release of cytokines, metalloproteinases, cyclooxygenase-2, and vascular cell adhesion molecule-1 (VCAM-1). The release of these inflammatory mediators that lead to tissue degradation is also observed in other forms of arthritis. Thus beta(2)M itself may elicit the release of inflammatory mediators from synovial fibroblasts even in the absence of cellular infiltrates. PMID- 11264782 TI - The pathogenesis of beta(2)-microglobulin-induced bone lesions in dialysis related amyloidosis. AB - Dialysis-related amyloidosis (DRA), also referred to as beta(2)-microglobulin amyloidosis (A beta(2)M), is an important cause of morbidity in patients with chronic renal failure and in those who are on dialysis. Although DRA deposits from affected joints have been characterized as a unique amyloid fibril protein, beta(2)M, less is known about the pathologic role of beta(2)M as a mediator of bone and joint disease. Potential mechanisms for beta(2)M pathologic interaction in bone include bone growth factors, cytokines, and advanced glycation end products (AGEs). It appears that DRA is the result of a complex interaction between bone resorption and surrounding tissue destruction culminating in beta(2)M deposition and amyloid formation. More work is required to elucidate the relationship between beta(2)M accumulation and progressive tissue destruction. PMID- 11264783 TI - Cobalt blues. PMID- 11264785 TI - Asparagusuria. PMID- 11264784 TI - Pathogenesis of beta(2)-microglobulin amyloidosis: role of monocytes/macrophages. AB - beta(2)-microglobulin (beta(2)M) amyloidosis (A beta(2)M) is a serious, often incapacitating complication for patients undergoing long-term hemodialysis. Amyloid deposits composed of beta(2)M fibrils as the major constituent protein are mainly localized in joints and periarticular bone and lead to chronic arthralgias, carpal tunnel syndrome, and eventually destructive arthropathy. Although recent histologic studies have shown the accumulation of monocytes/macrophages around amyloid deposits, the factor(s) causing their infiltration and pathologic involvement have yet to be fully elucidated. Immunohistochemical staining reveals that macrophages in tenosynovial tissues express CD13, CD14, CD33, HLA-DR, and CD68 antigens on their surfaces and express interleukin (IL)-1 beta, tumor necrosis factor (TNF)-alpha, and IL-6. Many of these cells also express LFA-1 (CD11a/CD18), Mac-1 (CD11b/CD18), and VLA-4 (CD49d/CD29) on their surfaces. AGE-modified beta(2)M enhances chemotaxis of monocytes and stimulates macrophages to release bone-resorbing cytokines, such as IL-1 beta, TNF-alpha and IL-6. Via a RAGE-mediated pathway, AGE-modified, but not unmodified beta(2)M, significantly delays constitutive apoptosis of human peripheral blood monocytes. Monocytes survival in an advanced glycation end product (AGE) beta(2)M-containing microenvironment is associated with their phenotypic alteration into macrophage-like cells that generate more reactive oxygen species and elaborate greater quantities of IL-1 beta and TNF-alpha. Thus through regulation of their survival and differentiation, AGE beta(2)M in amyloid deposits may be able to influence the presence and quantity of infiltrated monocytes, and hence their biologic effects. PMID- 11264787 TI - Platelet activation markers and soluble adhesion molecules in patients with systemic lupus erythematosus. AB - We assessed the role of platelet activation markers (PMPs, Annexin V and CD62P on activated platelets), cytokines (IL-1 beta, IL-4, IL-6, IFN- gamma, GM-CSF, and TNF alpha ), and soluble factors (sIL-2R, TM, sHLA-1, beta(2) -m, sVCAM-1, sPECAM 1, sP-selectin and sE-selectin) in vascular damage related to SLE. There were differences in the levels of PMPs and platelet activation markers between the SLE patients and controls (PMPs: 493+/-82 vs. 328+/-36, p<0.05; plt-CD62P; 8.5%+/-1.2 % vs. 4.6%+/-0.7 %, p<0.05; plt-Annexin V: 11.3%+/-2.1 % vs. 4.9%+/-0.6 %, p<0.01). There were no differences in the levels of IFN- gamma between the groups. However, the levels of IL-1 beta, IL-4, IL-6, GM-CSF, TNF alpha, and soluble factors were higher in the SLE patients than in the controls. The levels of IL-4, IL-6, beta2 -m, sIL-2R, sVCAM-1, sP-selectin, and sE-selectin in SLE patients with elevated sTM levels were higher than those in the SLE patients without elevated sTM levels. On the other hand, elevations of sIL-2R, sVCAM-1, and sP-selectin were not found in patients with Behcet disease or rheumatoid arthritis. The levels of platelet CD62P, platelet annexin V, and PMP were significantly elevated in high-sTM patients. These findings suggest the possibility that activated platelets and cytokines participate in the pathogenesis of SLE in patients with elevated sTM levels. PMID- 11264786 TI - Linomide does not prevent spontaneous autoimmune thyroiditis in NOD mice. AB - Linomide is a potent immunomodulator and has been reported to prevent type 1 diabetes mellitus in non-obese diabetic (NOD) mice and to reduce the incidence of other autoimmune diseases in animal models. The mechanisms of action seem to involve antigen expression by down regulation of macrophage activity and to antagonise the activation of Th1 cells during the cellular immune response. With the purpose to investigate the effect of Linomide on the incidence of spontaneous autoimmune thyroiditis (AIT) in female NOD mice we administered Linomide in drinking water (100 mg/kg/day) to NOD mice from 5th to 19th week of age. The mice were sacrificed at the end of week 19. None of the mice developed diabetes during the study period. The incidence of thyroiditis was evaluated on paraffin HE stained sections and graduated on a scale from 0 to 4. Thirty-two percent of 37 mice treated with Linomide developed thyroiditis compared to 45% of 22 controls (p=0.31, chi2 =1.00). Among the mice who developed thyroiditis no difference in the degree of thyroiditis was found. Therefore no beneficial effect of Linomide on the incidence of spontaneous AIT in NOD mice could be demonstrated. PMID- 11264788 TI - Antibodies to SOX13 (ICA12) are associated with type 1 diabetes. AB - SOX13 is an islet cell autoantigen (ICA12), identified by antibody screening of an islet cDNA library, using sera from patients with Type 1 diabetes. We ascertained the frequency of antibody reactivity to SOX13 and compared it with other Type 1 diabetes autoantibody reactivities. Antibodies were measured by radioimmunoprecipitation (RIP) using (35) S labelled SOX13 expressed in rabbit reticulocyte lysate. Sera from 109 subjects with Type 1 diabetes, 29 with Type 2 diabetes, 144 with other autoimmune diseases and from 201 controls were tested for anti-SOX13, and results were compared with the frequency of antibodies to glutamic acid decarboxylase (anti-GAD), islet cell antigen 512 (anti-ICA512) and islet cell cytoplasm (ICA). Anti-SOX13 were detected in 20 (18.3%) of 109 subjects with Type 1 diabetes, and more frequently in adults than in children (29% vs 10%). Anti-SOX13 usually occurred with anti-GAD but rarely with anti ICA512. Seven sera positive for anti-SOX13 did not react with either GAD, ICA512 or islet cell cytoplasm indicating that anti-SOX13 represented a distinct population of antibodies. Reactivity to SOX13 represents a further autoantibody response in adults with Type 1 diabetes and may provide a useful disease marker in subjects in whom other autoantibody tests are negative. PMID- 11264789 TI - Low-dose streptozotocin induces sustained hyperglycemia in Macaca nemestrina. AB - The potential for using macaques to create a nonhuman primate diabetic model was investigated. The significant objectives were to determine a) prognosis of STZ induced permanent beta cell destruction in nonhuman primates, and b) the potential to use STZ treated animals in a model of autoimmune diabetes by following adoptively transferred lymphocytes into MHC identical macaques. Beta cell impairment was achieved by a single intravenous, low dose (10-40 mg/kg body weight) streptozotocin injection in a majority of pigtailed macaques (Macaca nemestrina). Multiple injections, even at low doses at close intervals affected liver and kidney functions in addition to beta cell destruction. Abnormal IVGTT were observed in all streptozotocin-treated animals, in some within a week to 10 days. The fasting blood glucose levels rose from <70 mg/dl in pre-STZ stage to above 400 mg/dl in severely diabetic macaques. Histological evidence suggests loss of beta cells when animals were euthanized within two to four weeks post-STZ treatment. Near complete destruction of beta cells was observed in animals maintained longer than three months on insulin. Donor T cells from STZ-treated animals were incubated overnight with 10U/ml IL-2 and 2.5 ug/ml PHA and then injected iv into a MHC-identical non-diabetic sibling. Three weeks later a second injection of donor PMBC labeled with vital dye Cell Tracker Green was given and the animal was euthanized after 24 hours. The recipient showed labeled donor T cells in the pancreas, spleen and peripheral blood, consistent with specific homing of activated lymphocytes from the diabetic donor. PMID- 11264791 TI - Hypothesis. Bystanders or bad seeds? Many autoimmune-target cells may be transforming to cancer and signalling "danger" to the immune system. AB - Autoimmune-target cells in autoimmune disease (AID) are usually construed as constitutionally normal healthy cells. A related assumption is that other cells in the body of AID patients, except for certain immunocytes, are healthy cells. An implication of that view is that any systemic pathology in organ-specific AID is related to metabolic derangements secondary to tissue destruction. However, much data on target and other cells in AID suggest widespread primary cellular defects. In insulin-dependent diabetes mellitus (IDDM), for example, many "complications" such as atherosclerosis, premature arterial stiffening, senescence of fibroblasts in vitro, and exuberant growth of smooth muscle and mesangial cells in vivo are not strictly attributable to glucose elevation. Also unexplained is the similar appearance of IDDM beta-cells and cells from insulinoma and why the prodromal phase of IDDM has many insulinoma-like features. While AID target cells have often been likened to neoplastic cells, investigators have rarely explored the possibility that autoimmunity in AID is fundamentally antineoplastic. This is likely because the dominant ideas in oncology and immunology-somatic mutation and clonal deletion, respectively-have prevented explanations for how normal immunity could detect transforming cells not expressing non-self antigens. New and less conventional theories of cancer and immunity have facilitated such an explanation. I use Rubin's "epigenetic" aging model of carcinogenesis and Matzinger's "danger" model of immunity to integrate the immunological and oncological sides of AID. In particular, I postulate that individuals suffering from AID have inherited many foci of prematurely aging cells. Those inherently damaged cells adapt to in vivo challenges by beginning to transform into cancer cells. However, as long as those stressed cells have not fully transformed, they will continue to signal "danger" to the innate immune system. The clinical outcome of that struggle between incipient neoplasia and immunity will vary depending upon the degree of tumor-proneness and resistance of the individual. Borrowing from cancer geneticist Henry Lynch, I postulate that tumor-resistance is inherited as a quantitative polygenic trait in direct proportion to tumor-proneness. I further contend that tumor-proneness and immunity are linked polygenic traits such that the greater one's tumor-proneness, the more powerful his/her antitumor immunity. I point to the shared DNA repair deficiency of certain cancer-prone syndromes and HLA-linked AID, their occasional co-occurrence, and their demonstrably exceptional immunity against solid tumors. I propose that HLA-linked AID constitute "chronic hypersensitivity syndromes" due to immunity's largely hidden battle to suppress multiple incipient neoplastic microfoci. Much of the physiopathology of AID is explicable as a sustained systemic response to threatened neoplastic transformation. PMID- 11264790 TI - Combinations of beta cell specific autoantibodies at diagnosis of diabetes in young adults reflects different courses of beta cell damage. AB - To explore the natural course of beta cell function in recent onset diabetes, a subgroup (n=157) of all incident cases (n=879) 15-34 years old, 1992-1993 in Sweden, and with positivity for at least one autoantibody of islet cell antibodies (ICA), glutamic acid decarboxylase antibodies (GADA) or tyrosine phosphatase antibodies (IA-2A) were followed prospectively for the first four years with annual analysis of C-peptide. The aim was to relate the course of beta cell function, measured as C-peptide, in early diabetes with the presence of different islet autoantibodies at diagnosis. We found that patients positive for ICA alone (n=11) had significantly higher C-peptide levels both at diagnosis and during the first three years compared with the other patients (n=146; p=0.022, p<0.001, p=0.004 and p=0.0022). Patients positive for GADA alone or in combination with other antibodies (n=125) had significantly lower C-peptide during the first three years after diagnosis compared with the other patients (n=32, p<0.001, p=0.0011 and p=0.0136). Patients with two or three autoantibodies had C-peptide levels similar to levels found in patients positive only for GADA. However, after four years, there were no significant differences between any of the groups of different autoantibody combinations. At diagnosis, 55% (86/157) of the patients had C-peptide levels above the lower normal range of 0.25 nmol/l, but the frequency of patients with beta cell function above this level decreased after two years to 41% (65/157; p=0.035) and after four years to 22% (35/157; p=0.0041). It is concluded that young adult diabetic patients positive only for ICA at diagnosis have a better preserved beta cell function with higher levels of C-peptide during the first three years compared with patients positive for GADA alone or in combinations with other autoantibodies. PMID- 11264792 TI - Peripheral T-cell tolerance defined through transgenic mouse studies. AB - Selection in the thymus restricted by MHC and self-peptide shapes the diverse reactivities of the T-cell population which subsequently seeds into the peripheral tissues, in anticipation of the universe of pathogen antigens to which the organism may be exposed. A necessary corollary is the potential for T-cell self-reactivity (autoimmunity) in the periphery. Transgenic mouse models in which transgene expression in the thymus is prevented or excluded, have been particularly useful for determining the immunological outcome when T-cells encounter transgene-encoded 'self' antigen in peripheral tissues. Data suggest that non-mutually exclusive mechanisms of T-cells 'ignoring' self-antigen, T-cell deletion, T-cell anergy and T-cell immunoregulation have evolved to prevent self reactivity while maintaining T-cell diversity. The peripheral T-cell repertoire, far from being static following maturation through the thymus, is in a dynamic stated determined by these peripheral selective and immunoregulatory influences. This article reviews the evidence with particular reference to CD8+ive T-cells. PMID- 11264793 TI - A Numerical Simulation of the Diffusion of Near Infrared Light Through Brain Tissue. AB - A numerical simulation of the transport of light energy in the near infrared region of the spectrum through human brain tissue is presented. This simulation models the use of near infrared spectroscopic techniques to quantify the levels of oxygen present in brain tissue. Successful application of the technique requires knowledge of the optical pathlength in the tissue, and it is the goal of this simulation to quantify the relationship of the optical pathlength and the oxygenation state of the tissue. Both implicit and explicit finite element schemes for unstructured grids are implemented and discussed. Several application simulations using three tissue grids of varying degrees of physiological accuracy are then conducted, and figures for the optical pathlength of light through the tissue at varying levels of oxygenation are computed. These results are then used to develop a quantitative relationship between the pathlength and the absorption parameter for the three tissue models which we explore. PMID- 11264794 TI - Quantification of Bone Microarchitecture with the Structure Model Index. AB - The deterioration of cancellous bone structure due to aging and disease is characterized by a conversion from plate elements to rod elements. Consequently the terms "rod-like" and "plate-like" are frequently used for a subjective classification of cancellous bone. In this work a new morphometric parameter called Structure Model Index (SMI) is introduced, which makes it possible to quantify the characteristic form of a three-dimensionally described structure in terms of the amount of plates and rod composing the structure. The SMI is calculated by means of three-dimensional image analysis based on a differential analysis of the triangulated bone surface. For an ideal plate and rod structure the SMI value is 0 and 3, respectively, independent of the physical dimensions. For a structure with both plates and rods of equal thickness the value lies between 0 and 3, depending on the volume ratio of rods and plates. The SMI parameter is evaluated by examining bone biopsies from different skeletal sites. The bone samples were measured three-dimensionally with a micro-CT system. Samples with the same volume density but varying trabecular architecture can uniquely be characterized with the SMI. Furthermore the SMI values were found to correspond well with the perceived structure type. PMID- 11264795 TI - Mixed and Penalty Finite Element Models for the Nonlinear Behavior of Biphasic Soft Tissues in Finite Deformation: Part I - Alternate Formulations. AB - This paper addresses finite element-based computational models for the three dimensional, (3-D) nonlinear analysis of soft hydrated tissues, such as the articular cartilage in diarthrodial joints, under physiologically relevant loading conditions. A biphasic continuum description is used to represent the soft tissue as a two-phase mixture of incompressible, inviscid fluid and a hyperelastic solid. Alternate mixed-penalty and velocity-pressure finite element formulations are used to solve the nonlinear biphasic governing equations, including the effects of a strain-dependent permeability and a hyperelastic solid phase under finite deformation. The resulting first-order nonlinear system of equations is discretized in time using an implicit finite difference scheme, and solved using the Newton-Raphson method. Using a discrete divergence operator, an equivalence is shown between the mixed-penalty method and a penalty method previously derived by Suh et al. [1]. In Part II [2], the mixed-penalty and velocity-pressure formulations are used to develop two-dimensional (2-D) quadrilateral and triangular elements and 3-D hexahedral and tetrahedral elements. Numerical examples, including those representative of soft tissue material testing and simple human joints, are used to validate the formulations and to illustrate their applications. A focus of this work is the comparison of alternate formulations for nonlinear problems. While it is demonstrated that both formulations produce a range of converging elements, the velocity-pressure formulation is found to be more efficient computationally. PMID- 11264796 TI - Adaptation Models of Anisotropic Bone. AB - A theoretical model and numerical methods were developed for testing different bone internal remodelling stimuli. The keystone of the study was the formulation of a stimulus based on the mechanical invariants of the stress tensor, which took into account bone non-homogeneity and anisotropy. A non-site specific remodelling rate equation was then used for the apparent density whereas anisotropy was fixed and evaluated from anatomic observations. An node-based semi-implicit algorithm with adaptive stepsize was implemented for solving the evolution equation. To preclude numerical artifacts (non-convergence, instability), a phase space description was proposed. As an illustration, the evolution of apparent density distribution surrounding the femoral stem after a Total Hip Replacement was simulated. Three stimuli were tested: the strain energy density stimulus, the octahedral shear stress stimulus, and an anisotropic plastic yield stress stimulus. PMID- 11264797 TI - Strain Concentrations Surrounding an Ellipsoid Model of Lacunae and Osteocytes. AB - Direct cell sensing of tissue matrix strains is one possible signaling mechanism for mechanically mediated bone adaptation. We utilized homogenization theory to estimate bone tissue matrix strains surrounding osteocytes using two sets of models. The first set of models estimated the strain levels surrounding the lacunae and canaliculi, taking into account variations in lamellar properties. The second set estimated strain levels in the osteocyte and the surrounding matrix for different cellular mechanical properties. The results showed that the strain levels found in and surrounding osteocytes, 1700 to 2700 microstrain (denoted as ue; 1 ue =.0001% strain), were significantly greater than the trabecular tissue level strains of {1325 ue, 287 ue, 87 ue} used for model input. Variation in lamellar properties did not affect strain levels, except at lamellar boundaries. Strain in and surrounding the osteocyte was not significantly affected by cellular stiffness ranging between 28 and 28, 000 Pascals (Pa). Strain levels surrounding lacunae and canaliculi were approximately equivalent. PMID- 11264798 TI - C/C++ Coding for Matrix Pseudo Inverses in Clinical Near Infrared Spectroscopy. AB - Near infrared spectroscopy is used clinically to investigate patterns of change in cerebral oxygenation. We have shown that differences reported between authors are likely the result of computer encoding errors in the manipulation of matrices. Current methods compute the inverse of a non-square matrix to derive chromophore concentration values, and solution of another non-square matrix to derive polynomial coefficients of a least squares best fit curve from which the first derivative can be used to estimate blood flow values. Encoding of these pseudo inverses involves too many nested looping steps to easily identify encoding errors. We have given C/C++ source code along with sample numerical values at the termination of each loop within the algorithm. This provides counter checking for future software development by other programmers, and also permits other investigators to report whether the software used for their experiments agrees with previously published material. PMID- 11264799 TI - A Three-Dimensional Musculoskeletal Model of the Human Knee Joint. Part 1: Theoretical Construct. AB - A three-dimensional model of the knee is developed to study the interactions between the muscles, ligaments, and bones during activity. The geometry of the distal femur, proximal tibia, and patella is based on cadaver data reported for an average-size knee. The shapes of the femoral condyles are represented by high order polynomials; the tibial plateaux and patellar facets are approximated as flat surfaces. The contacting surfaces of the femur and tibia are modeled as deformable, while those of the femur and patella are assumed to be rigid. Interpenetration of the femur and tibia is taken into account by modeling cartilage as a thin, linear, elastic layer, mounted on rigid bone. Twelve elastic elements describe the geometry and mechanical properties of the cruciate ligaments, the collateral ligaments, and the posterior capsule. The model is actuated by thirteen musculotendinous units, each unit modeled as a three-element muscle in series with tendon. The path of each muscle is approximated as a straight line, except where it contacts and wraps around bone and other muscles; changes in muscle paths are taken into account using data obtained from MRI. In the first part of this paper, the model is used to simulate passive knee flexion. Quantitative comparisons of the model results with experimental data reported in the literature indicate that the relative movements of the bones and the geometry of the ligaments and muscles in the model are similar to those evident in the real knee. In Part II, the model is used to describe knee-ligament function during anterior-posterior draw, axial rotation, and isometric knee-extension and knee-flexion exercises. PMID- 11264801 TI - Review: Diarthrodial Joints-Kinematic Pairs, Mechanisms or Flexible Structures? PMID- 11264800 TI - Further Roles of Geometry and Properties in the Mechanics of Saccular Aneurysms. AB - Rupture of intracranial saccular aneurysms continues to result in significant morbidity and mortality. Although it has long been thought that biomechanical factors play key roles in the genesis, growth, and rupture of these lesions, few analysis have employed realistic descriptions of the geometries and material properties. This paper presents parametric finite element studies for subclasses of elliptical and spherical lesions which complement those recently reported by Kyriacou and Humphrey. In particular, we show again that lesion shape, not size, is a primary determinant of aneurysmal wall stress. Moreover, material anisotropy and geometry can exhibit competing or synergistic effects on the stress fields - this suggests that these interactions may be important in the formulation of theories on lesion growth. Finally, we show that Laplace's equation (for spherical membranes) yields reasonable approximations for wall stress only for a very limited class of lesions. There is a need, therefore, for detailed analysis and thus more precise data on lesion geometry, material properties, and loading conditions. PMID- 11264802 TI - Mixed and Penalty Finite Element Models for the Nonlinear Behavior of Biphasic Soft Tissues in Finite Deformation: Part II - Nonlinear Examples. AB - This two-part paper addresses finite element-based computational models for the three-dimensional (3-D) nonlinear analysis of soft hydrated tissues, such as articular cartilage in diarthrodial joints, under physiologically relevant loading conditions. A biphasic continuum description is used to represent the soft tissue as a two-phase mixture of incompressible inviscid fluid and a hyperelastic, transversely isotropic solid. Alternate mixed-penalty and velocity pressure finite element formulations are used to solve the nonlinear biphasic governing equations, including the effects of strain-dependent permeability and a hyperelastic solid phase under finite deformation. The resulting first-order, nonlinear system of equations is discretized in time using an implicit finite difference scheme, and solved using the Newton-Raphson method. Details of the formulations were presented in Part I [1]. In Part II, the two formulations are used to develop two-dimensional (2-D) quadrilateral and triangular elements and three-dimensional (3-D) hexahedral and tetrahedral elements. Numerical examples, including those representative of soft tissue material testing and simple human joints, are used to validate the formulations and to illustrate their applications. A focus of this work is the comparison of the alternate formulations for nonlinear problems. While it is demonstrated that both formulations produce a range of converging elements, the velocity-pressure formulation is found to be more efficient computationally. PMID- 11264803 TI - Review: Computer Methods in Membrane Biomechanics. AB - The purpose of this paper is twofold: first, to review analytical, experimental, and numerical methods for studying the nonlinear, pseudoelastic behavior of membranes of interest in biomechanics, and second, to present illustrative examples from the literature for a variety of biomembranes (e.g., skin, pericardium, pleura, aneurysms, and cells) as well as elastomeric membranes used in balloon catheters and new cell stretching tests. Although a membrane approach affords great simplifications in comparison to the three-dimensional theory of nonlinear elasticity, associated problems are still challenging. Computer-based methods are essential, therefore, for performing the requisite experiments, analyzing data, and solving boundary and initial value problems. Emphasis is on stable equilibria although material instabilities and elastodynamics are discussed. PMID- 11264804 TI - Intra-Operative Analysis of Scoliosis Surgery in 3-D. AB - Scoliosis is a three-dimensional deformity characterized by coronal, sagittal and axial rotation of the spine. Surgical fusion of the spine is required in severe cases. Assessment of the surgical procedure requires enough accuracy and flexibility to allow planning of individual interventions or implant designs. Conventional 2-D radiography and even 3-D CT scanning have limitations for in depth analysis of scoliosis that limit the ability to see the three-dimensional deformity and expose the patient to considerable doses of radiation, respectively. Our stereophotogrammetric analysis is able to provide accurate, intra-operative measurement of vertebral movement during surgical manuevres. Stereophoto pairs taken at each stage of the operation and robust statistical techniques can be used to determine rotation, translation, goodness of fit, and overall spinal contour before, during, and after the surgical instrumentation. A demonstration of data available from this system is included. PMID- 11264805 TI - An Analysis and Comparison of Convergence and Uniqueness of Time-Independent Bone Adaptation Models. AB - Several stimuli are proposed in the bone remodeling theory. It is not clear, if a unique solution exists and if the result is convergent using a certain stimulus. In this study, the strain stimulus, strain energy stimulus and the von Mises stress stimulus for bone remodeling are compared and applied to a square plate model using the finite element method. In the plane stress state, the remodeling equilibrium equations are transformed into functions of only the principal strains and the graphs of these functions are drawn in a diagram using the principal strains as the variables of two coordinate axes. The equation of the sum of principal strain squared equal to a constant is a circle in the diagram. The remodeling equilibrium equation of the strain stimulus is a quadrangle fitting into the circle, the remodeling equilibrium equation of the strain energy stimulus is an ellipse and the remodeling equilibrium equation of the von Mises stress stimulus is also an ellipse close to the principal strains circle when we take the same constants in the above equations. Using the finite element method, two models are performed with the uniform initial elastic properties and with the semi-random initial distribution of the elastic properties. The principal strains as the final finite element results converge within 2% of the objective constant for all the different stimuli. The obtained Young's moduli of two models as the adaptation object are different but in equilibrium, i.e. the equilibrium solution of adaptation model is not unique. The principal strains can not be used to examine the uniqueness of solution, since two different solutions can have the same results of principal strains. Using a certain stimulus, certain initial properties and a certain iterative equation, the solution is unique in equilibrium. The results using the model in this study show also that the same results can be obtained using any of the three stimuli when a proper constant in each remodeling equilibrium equation is chosen. PMID- 11264806 TI - Bone Load Estimation for the Proximal Femur Using Single Energy Quantitative CT Data. AB - A density-based load estimation method was applied to determine femoral load patterns. Two-dimensional finite element models were constructed using single energy quantitative computed tomography (QCT) data from two femora. Basic load cases included parabolic pressure joint loads and constant tractions on the greater trochanter. An optimization procedure adjusted magnitudes of the basic load cases, such that the applied mechanical stimulus approached the ideal stimulus throughout each model. Dominant estimated load directions were generally consistent with published experimental data for gait. Other estimated loads suggested that loads at extreme joint orientations may be important to maintenance of bone structure. Remodeling simulations with the estimated loads produced density distributions qualitatively similar to the QCT data sets. Average nodal density errors between QCT data and predictions were 0.24 g/cm(3) and 0.28 g/cm(3). The results indicate that density-based load estimation could improve understanding of loading patterns on bones. PMID- 11264807 TI - Stability and Control of Human Trunk Movement During Walking. AB - A mathematical model has been developed to study the control mechanisms of human trunk movement during walking. The trunk is modeled as a base-excited inverted pendulum with two-degrees of rotational freedom. The base point, corresponding to the bony landmark of the sacrum, can move in three-dimensional space in a general way. Since the stability of upright posture is essential for human walking, a controller has been designed such that the stability of the pendulum about the upright position is guaranteed. The control laws are developed based on Lyapunov's stability theory and include feedforward and linear feedback components. It is found that the feedforward component plays a critical role in keeping postural stability, and the linear feedback component, (resulting from viscoelastic function of the musculoskeletal system) can effectively duplicate the pattern of trunk movement. The mathematical model is validated by comparing the simulation results with those based on gait measurements performed in the Biomechanics Laboratory at the University of Manitoba. PMID- 11264808 TI - Technical Note: Modelling Soft Tissue Using Biphasic Theory - A Word of Caution. AB - In recent years the biphasic approach initiated by Mow and coworkers, has been very popular in modelling soft, hydrated, cartilage tissues as well as other soft tissues, such as the brain. This work points out that due to the inherent inability of biphasic models in their present form to account for stress-strain rate dependence resulting from the viscoelasticity of the solid phase, the applicability of these models is limited to the loading conditions producing large relative velocities of phases. PMID- 11264809 TI - A Three-Dimensional Musculoskeletal Model of the Human Knee Joint. Part 2: Analysis of Ligament Function. AB - A three-dimensional model of the knee is used to study ligament function during anterior-posterior (a-p) draw, axial rotation, and isometric contractions of the extensor and flexor muscles. The geometry of the model bones is based on cadaver data. The contacting surfaces of the femur and tibia are modeled as deformable; those of the femur and patella are assumed to be rigid. Twelve elastic elements are used to describe the geometry and mechanical properties of the cruciate ligaments, the collateral ligaments, and the posterior capsule. The model is actuated by thirteen musculotendinous units, each unit represented as a three element muscle in series with tendon. The calculations show that the forces applied during a-p draw are substantially different from those applied by the muscles during activity. Principles of knee-ligament function based on the results of in vitro experiments may therefore be overstated. Knee-ligament forces during straight a-p draw are determined solely by the changing geometry of the ligaments relative to the bones: ACL force decreases with increasing flexion during anterior draw because the angle between the ACL and the tibial plateau decreases as knee flexion increases; PCL force increases with increasing flexion during posterior draw because the angle between the PCL and the tibial plateau increases. The pattern of ligament loading during activity is governed by the geometry of the muscles spanning the knee: the resultant force in the ACL during isometric knee extension is determined mainly by the changing orientation of the patellar tendon relative to the tibia in the sagittal plane; the resultant force in the PCL during isometric knee flexion is dominated by the angle at which the hamstrings meet the tibia in the sagittal plane. PMID- 11264810 TI - The Influence of Flow on the Concentration of Platelet Active Substances in the Vicinity of Mural Microthrombi. AB - The flow effect on the concentration of platelet active substances in the vicinity of a mural microthrombus is investigated numerically. A three dimensional model is employed in which the mural microthrombus is modelled as a semisphere attached to a plane surface. The description of the blood flow uses the three-dimensional incompressible Navier-Stokes equations for Newtonian fluids, and the mass transport and reaction kinetics are modelled applying a system of coupled convection-diffusion equations with reaction terms. The numerical approach employs the finite element method and a streamline upwind stabilization due to the high Peclet numbers. To verify the calculated concentrations a random walk model for the simulation of the convective diffusion of thrombogenic substances is employed. The results show flow recirculations directly upstream and downstream of the microthrombus which cause local concentration maxima in these regions. The resulting thrombin concentration exceeds the required level for platelet activation at shear rates and thrombus sizes under consideration. The resulting concentrations of thromboxane A(2) (TxA(2) ) and adenosine diphosphate (ADP) exceed the corresponding activation levels only at low shear rates and for large aggregates. For an aggregate diameter of 20 um at the shear rate [formula: see text] the maximum TxA(2) concentration at a distance from the wall of 3 um is 0.69 uM, which exceeds the required level of 0.6 uM. Increasing the shear rate to [formula: see text] causes a decrease of the maximum concentration to 0.43 uM. PMID- 11264811 TI - Loading Mode Interactions in Simulations of Long Bone Cross-Sectional Adaptation. AB - Although many bone adaptation theories have been formulated to address both trabecular and cortical adaptation, most applications have focused on trabecular adaptation. Thus far, no thorough investigations of the influence of different types of loading on predicted patterns of long bone cross-sectional adaptation have been reported. In the current study, we present a new model for long bone cross-sectional adaptation that incorporates axial, bending and torsional loading components. We found that bending moments have a strong potential to modulate cross-sectional geometry, but can produce unforseen (and unrealistic) geometric instabilities. Torsional moments have the ability to suppress these instabilities, suggesting that torsion may play a more significant role in guiding long bone development than previously recognized. Our results also call into question the concept of strict "remodeling equilibrium," suggesting that long bones do not necessarily approach a state of uniform mechanical stimulation. This modeling approach provides an additional perspective on experimental studies, and may lead to a greater understanding of the interaction between mechanics and biology in long bone adaptation. PMID- 11264812 TI - A Novel 3D Microstructural Model for Trabecular Bone: I. The Relationship between Fabric and Elasticity. AB - A novel 3D microstructural model is proposed to investigate the relationship between morphology and mechanical properties of trabecular bone. Open and closed cell geometries were selected with varying volume fractions and degrees of anisotropy that simulate the architectures of human cancellous bone over a broad range of anatomical locations. Finite element models of both cells were developed using beams and shells. Volume fraction and mean intercept length were calculated analytically and the effective elastic tensors were computed with linear tissue properties and periodic boundary conditions. Distinct, but strong relationships were obtained between fabric and the elastic tensors for open and closed cell geometries, which bound the experimental results obtained for human bone and support the relevance of the selected model to address trabecular bone fragility. PMID- 11264814 TI - A Novel 3D Microstructural Model for Trabecular Bone: II. The Relationship Between Fabric and the Yield Surface. AB - A novel 3D microstructural model was proposed and validated in part I of this publication. In part II, the model was used to identify the yield surface of a representative volume element of human trabecular bone as a function of volume fraction and degree of anisotropy. Finite element models of open and closed cells geometries were used to calculate effective yield stresses for a variety of loading cases with periodic boundary conditions. The postyield behaviour of the trabecular tissue was assumed from data available for cortical tissue. The yield stresses defined by a 0.2% offset in the global stress-strain curve were fit to an orthotropic Hill criterion and the parameters of the surface calculated. Similarly to the previous elastic analysis, distinct but strong relationships were obtained between volume fraction, fabric and the yield surface parameters for both the open and closed cell geometries. This finding suggests that volume fraction and fabric may be used to predict the initiation of mechanical damage in human trabecular bone at the continuum level. PMID- 11264813 TI - Quantitative Assessment of Polyethylene Component Position in a Mobile-Bearing Prosthetic Knee, Using a Single Radiograph. AB - Rationale and Objectives. To reduce tibio-femoral misalignment, the polyethylene bearing-component of a new knee prosthesis was allowed limited motion on the underlying metallic component. The object of the work presented here was to develop a suitable radiographic technique for quantifying the in-vivo position of the bearing. By collecting these data at discrete flexion angles, the functional operation of the prosthesis could be determined. Methods. The known geometries between landmarks on the two components were used to produce algorithms for reconstructing their spatial positions from a single radiograph. A custom designed computer program utilized these algorithms to determine the relative translation and rotation of the polyethylene component. Results. This technique produced typical errors of 0.54 mm translation and 0.56 degrees rotation between the polyethylene component and the underlying metallic component. Conclusions. A practical method has been developed for assessing mobile-bearing motion, in vivo. This method can be applied to other prosthetic devices, or combinations of components, once the requirement for identifiable landmarks has been addressed. Clinical Relevance. Skeletal and soft-tissue changes in the pathological knee may produce abnormal rotations and translations in the transverse tibial plane. This technique is intended both to validate the design philosophy of a mobile-bearing prosthesis and to provide additional data on any pathological motions, which will have implications for future prosthetic designs. PMID- 11264815 TI - Characterisation of Anisotropic and Non-linear Behaviour of Human Skin In Vivo. AB - The aim of this work is to characterise the in-plane mechanical behaviour of human skin in vivo. For this purpose the structural skin model proposed by Lanir [1] is employed and a mixed numerical-experimental method is developed. The numerical-experimental method is based on the confrontation of measured data from an experiment, with calculated data from a finite element model, eventually leading to the determination of some of the parameters of a constitutive model, in the present case Lanir's Skin Model. Since collagen, the main constituent of skin, dominates the anisotropic and non-linear behaviour of skin, the parameters of Lanir's Skin Model concerning the mechanical behaviour of the collagen fibres are estimated. In vivo experiments were carried out on the volar forearm. During the experiments, reaction forces and the displacement field at different states of deformation are measured. Both data sets are used for the determination of the parameters. PMID- 11264816 TI - Rhythmic Movement of a Pair of One-Link Arms: Coordination by Intermittent Control. AB - This paper considers the coordination and control of periodic movements of a pair of one-link arms. The system consists of two one-link arms each controlled by two muscle-like actuators. The muscle-like actuators are activated by simulated neural inputs. The model is simple enough to analyze, yet it embodies many aspects of human arms. Three attributes of the rhythmic coordinated movement of two arms, namely frequency, magnitude, and relative phase, are the only inputs to the controller. The controller uses mild co-activation and primarily activates the agonist. The effects of transmission delays, present in the reflex loop of physiological systems, also are modeled. The results of this research indicate the feasibility of controlling oscillatory body movements with short periods of activation. The result of many simulations, by varying the frequency or amplitude of the movement, indicate that the apparent lack of a simple relationship between neural control and desired behavior of the system should not be mistaken as evidence for the absence of a causal relationship between the activation patterns of the muscles and the desired behavior. Simulations of this system show stable oscillations at different frequencies and magnitudes even with additive noise and changes in the system mass. PMID- 11264817 TI - Simulation of the Process of Food Destruction During the Final Occlusal Stage Using a Finite Element Non-Linear Dynamic Analysis. AB - The purpose of this study was to evaluate food destruction in the small space between opposing teeth during the final stage of occlusion. The time related process of food destruction was simulated using a finite element non-linear dynamic analysis based on three dimensional occlusal co-ordinate data of the upper and lower teeth, masticatory movement data and occlusal force records. Food destruction was analysed in two inter-arch relationships. The first was normal occlusion (cusp-to-fossa) and the second was Angle class II (cusp-to-cusp). Food destruction was successfully demonstrated in both cases. PMID- 11264818 TI - Transient Analysis of the Biomechanics of the Human Head with a High-Resolution 3D Finite Element Model. AB - An investigation of the mechanical response of the human head subject to externally applied forces is analysed using the finite element method (FEM). The model's geometry description is based on individual 3D MRI datasets of the head. After segmentation the MRI data serves as input to a fully automated mesh generator. The underlying algorithms of the mesh generator are stable and fast. Isotropic and anisotropic meshes consisting of tetrahedra or brick elements are created with a predefined spatial resolution. The meshes are of high quality with regard to numerical stability of the FE calculation. Transient head impact simulations can be carried out on the basis of these meshes with a spatial resolution of up to 2 mm. The results of the impact simulations are validated against previously published experimental data obtained from cadaver tests. PMID- 11264819 TI - A Case Study on Human Upper Limb Modelling for Dynamic Simulation. AB - Modelling consists of developing a representation of the properties of an object or a phenomenon with respect to the goals of its analysis. In this paper, the procedure presented is that followed in the CHARM project for human upper limb modelling with respect to the project constraints on the model implementation and simulation. The objective is to develop a Comprehensive Human Animation Resource Model allowing the simulation of human motion, including the finite element simulation of soft tissue deformation and muscular contraction. Generally, models are presented based on assumptions left for a fortiori validation. The a priori considerations which lead to these assumptions are rarely detailed. Here, the aim is to form a basis for the choices and assumptions which are to be made prior to the validation. The analysis is based on the general approach for the modelling of multi-body deformable systems as well as on previous studies on the human upper limb. PMID- 11264820 TI - Fibre Orientation in Human Fetal Heart and Ventricular Mechanics : A Small Perturbation Analysis. AB - The study of the topological organisation of myocardial cells is a basic requirement for understanding the mechanical design of the normal and pathological heart. Anatomical observations show that cardiac muscle tissue has a highly specialized architecture. We have made new quantitative measurements of fibre orientation through the heart wall by means of polarized light analysis on some thick sections of human fetal heart embedded in a resin and polymerized. A small perturbation method to find an equilibrium solution in a cylindrical left ventricular (LV) geometry with fibres running on toroidal shells of revolution is used to investigate the mechanical behaviour of three human fetal hearts (FH) of 14, 20 and 33 weeks of gestational age. The results of fibre strains and stresses presented for end-systolic state show significant differences when compared to results of the cylindrical geometry with regular helicoidal fibres running on cylindrical surfaces. Moreover, the toroidal shells of revolution explain shear stresses and strains in the transverse plane which also exist in the adult heart. PMID- 11264821 TI - A Kinematic Model of the Upper Limb Based on the Visible Human Project (VHP) Image Dataset. AB - A kinematic model of the arm was developed using high-resolution medical images obtained from the National Library of Medicine's Visible Human Project (VHP) dataset. The model includes seven joints and uses thirteen degrees of freedom to describe the relative movements of seven upper-extremity bones: the clavicle, scapula, humerus, ulna, radius, carpal bones, and hand. Two holonomic constraints were used to model the articulation between the scapula and the thorax. The kinematic structure of each joint was based on anatomical descriptions reported in the literature. The three joints comprising the shoulder girdle - the sternoclavicular joint, the acromioclavicular joint, and the glenohumeral joint - were each modeled as a three degree-of-freedom, ideal, ball-and-socket joint. The articulations at the elbow and wrist - humeroulnar flexion-extension, radioulnar pronation-supination, radiocarpal flexion-extension, and radiocarpal radial-ulnar deviation - were each modeled as a single degree-of-freedom, ideal, hinge joint. Locations of the joint centers and joint axes were derived by graphically inspecting the three-dimensional surfaces of the reconstructed bones. The relative positions of the bones were defined by fixing a reference frame to each bone; the position and orientation of each reference frame were based on the positions of anatomical landmarks and on the shapes of the reconstructed bone surfaces. Tables are provided which specify the positions and orientations of the joint axes and the bone-fixed reference frames for the model arm. PMID- 11264822 TI - A Model of Bone Adaptation Using a Global Optimisation Criterion Based on the Trajectorial Theory of Wolff. AB - Julius Wolff originally proposed that trabecular bone was influenced by mechanical stresses during the formative processes of growth and repair such that trabeculae were required to intersect at right angles. In this work, we have developed an analytical parametric microstructural model, which captures this restriction. Using homogenisation theory, a global material model was obtained. An optimal structure constructed of the homogenised material could then be found by optimising a cost function accounting for both the structural stiffness and the biological cost associated with metabolic maintenance of the bone tissue. The formulation was applied to an example problem of the proximal femur. Optimal densities and orientations were obtained for single load cases. The situation of multiple loads was also considered. In this case, we observe that the alignment of principal strains with the material orthotropy direction is, in general, not possible for all load cases. Thus less restrictive microstructures (nonorthotropic) will yield higher structural stiffnesses than strictly orthotropic microstructures. PMID- 11264823 TI - Development and Physical Validation of a Finite Element Model of Total Hip Dislocation. AB - Component-on-component impingement, followed by levering of the femoral head, is a common mode of dislocation in total hip arthroplasty. While there have been many registry-based studies of dislocation incidence, confounding factors and sources of variability in the clinical domain make it difficult to identify specific parameter influences. A three dimensional nonlinear finite element model has been developed for the purpose of studying the dislocation event, to allow determination of how individual factors such as component design and clinical implantation position affect the propensity for dislocation. Also, a laboratory testing apparatus was constructed to provide physical validation of the computational model. The finite element model correctly predicted the range of motion observed in the physical apparatus to within 1%, and predicted the peak resisting moment to within 2.5%. Under even a light joint load of 200 N, the von Mises stresses developed in the polyethylene insert reached 13 MPa, and the contact stresses rose to as high as 30 MPa. These deleterious elevations occurred not only at the site of neck impingement, but also at the site of head egress from the liner. PMID- 11264824 TI - Numerical Estimation of the Centres of Rotation and Resistance in Orthodontic Tooth Movement. AB - The position of the centre of resistance (Cre) as well as the centre of rotation (Cro) of a tooth under a force-system is still an open question. This paper presents a reliable and efficient three-dimensional rigid-body finite element technique to accurately estimate these centres. The influence of not only the root length but also the root diameter, the thickness of the periodontal ligament, as well as its material properties on the position of the Cre and Cro is investigated. Additionally, an explanation is given for the meaning of the coefficient (0.068 h(2) ) involved in Burstone's theoretical formula which is generalised and is expressed as the ratio of the flexibilities of tooth support in translation and pure moment rotation, respectively. The former ratio determines the position of the centres of rotation as a function of the applied moment-to-force ratio (M/F) and the relevant curve remains an isosceles hyperbola for any arbitrary-shaped tooth. The present study focuses on single-rooted teeth, such as maxillary canines and maxillary incisors, but the proposed methodology is generally applicable to any tooth. PMID- 11264825 TI - A Porous Media Approach to Finite Deformation Behaviour in Soft Tissues. AB - The present work presents a porous medium formulation for the biomechanical analysis of soft tissues. An updated Lagrangian approach is developed to study the coupled effects of low speed flows of fluid phases, in partially or fully saturated conditions, and the finite deformation occurring in the solid matrix. The procedure developed allows both for the evaluation of coupled geometric and material non-linearities. The main theoretical and computational aspects of this multiphase formulation are discussed. The finite element method is used for the numerical solution of the resulting coupled system of equations. A reference case is reported with regard to healthy and degenerative phases of intervertebral segment. Results reported allow for a detailed interpretation of the formulation reliability, also by comparison with existing experimental data. In particular, the role played by the fluid on the load carrying mechanism is pointed out, thus stressing the importance of a multiphase approach to the overall behaviour of the spinal motion segment in time. PMID- 11264826 TI - Flow Characteristics in an Anatomically Realistic Compliant Carotid Artery Bifurcation Model. AB - A numerical model for the pulsatile blood flow in an anatomically realistic compliant model of the human carotid artery bifurcation has been developed. The geometric model has been generated on the basis of an optically digitized arterial cast. The effects of the geometrically realistic flow domain and of the wall distensibility on the flow characteristics are investigated. The description of the blood flow uses the time-dependent, three-dimensional, incompressible Navier-Stokes equations for non-Newtonian inelastic fluids. The calculation of the wall displacement uses geometrically non-linear shell theory. In an iteratively coupled approach the flow equations and the shell equations are numerically solved using the finite element method. The results show strongly skewed velocity profiles with high gradients at the internal and external divider walls downstream of the bifurcation. Flow separation and recirculation occur in the regions at the outer walls of the branching during systolic flow deceleration and at the diastolic flow minimum. Further the results show complex non-symmetric secondary motion in the carotid sinus due to the slightly non-planar branching of the geometrically realistic model. At the internal divider wall high shear stress can be observed, whereas at the outer internal wall low and oscillating shear stress occurs. The comparison of the results in the realistic model with results in a geometrically idealized model primarily points out differences concerning the flow recirculation and the secondary flow pattern. Comparing the results with results of a corresponding rigid wall model demonstrates a decrease of wall shear stress magnitude and a slight reduction of flow separation and recirculation at the outer sinus wall in the distensible model. The relative decrease of the axial wall shear stress maximum is 17% at the internal divider wall. At the outer sinus wall where the shear stress is low the decrease is in the range of up to 50%. PMID- 11264827 TI - Surface Temperature Distribution of a Breast With and Without Tumour. AB - Breast cancer is a common and dreadful disease in women. Regular screening helps in its early detection. At present the most common methods of screening are by self examination and mammography. The surface temperature distribution of the breast can also provide some information on the presence of tumour. This distribution has a relation to the size and location of tumour and can be seen using thermography, where the infrared radiation emitted from the surface of the breast is recorded and a thermal pattern obtained. Thermography is a non-invasive and an inexpensive tool which could be used for early detection. In order to simulate the surface temperature distribution, a two-dimensional model of female breast with and without a carcinoma is considered. The breast is modelled with varying layer thickness close to the actual shape and numerically solved using finite element analysis. Temperature profiles are obtained for a normal breast and for a malignant one by varying the tumour size, location and the blood flow rates. The results show that the surface temperature for a malignant breast is higher than that of a normal one. In addition the size and location of the tumour do have an effect on the surface temperature distribution. It can also be seen that tumour of different sizes placed at the same location would yield the same maximum temperature depending on the blood perfusion rate. PMID- 11264828 TI - A Dynamic Optimization Solution for Vertical Jumping in Three Dimensions. AB - A three-dimensional model of the human body is used to simulate a maximal vertical jump. The body is modeled as a 10-segment, 23 degree-of-freedom (dof), mechanical linkage, actuated by 54 muscles. Six generalized coordinates describe the position and orientation of the pelvis relative to the ground; the remaining nine segments branch in an open chain from the pelvis. The head, arms, and torso (HAT) are modeled as a single rigid body. The HAT articulates with the pelvis via a 3 dof ball-and-socket joint. Each hip is modeled as a 3 dof ball-and-socket joint, and each knee is modeled as a 1 dof hinge joint. Each foot is represented by a hindfoot and toes segment. The hindfoot articulates with the shank via a 2 dof universal joint, and the toes articulate with the hindfoot via a 1 dof hinge joint. Interaction of the feet with the ground is modeled using a series of spring-damper units placed under the sole of each foot. The path of each muscle is represented by either a series of straight lines or a combination of straight lines and space curves. Each actuator is modeled as a three-element, Hill-type muscle in series with tendon. A first-order process is assumed to model muscle excitation-contraction dynamics. Dynamic optimization theory is used to calculate the pattern of muscle excitations that produces a maximal vertical jump. Quantitative comparisons between model and experiment indicate that the model reproduces the kinematic, kinetic, and muscle-coordination patterns evident when humans jump to their maximum achievable heights. PMID- 11264829 TI - A New Least Squares Based Calibration for an Ultrasound Tomography Scanner Using Radial Image Processing. AB - In the context of the development of an ultrasound scanner where a probe turns around the part of the body to be studied, a new tomographic technique has been developed: the Ultrasound Reflection-mode Tomography Using Radial Image Processing (URTURIP Technique). This technique is used when a unique B-scan image is insufficient to correctly describe a cross-section. It utilises B-scan images obtained under different angles of view in the same plane to reconstruct a better cross-sectional image. Before using the URTURIP Technique, a calibration step is required to accurately determine the centre of rotation of the probe. To solve the calibration problem, a reference method has been developed, but it is very time-consuming. This paper presents a faster method based on a least squares fitting and is compared to the reference method in terms of accuracy and time consuming. PMID- 11264830 TI - Identification of the Constitutive Behaviour of Dental Composite Cements During Curing. AB - This paper is concerned with the qualitative and quantitative description of the constitutive behaviour of dental composites during the curing process. Both generalized Kelvin- and Maxwell-models are studied and the parameters characterizing these models are determined by the means of an identification procedure based on the matching of model calculations to experimental data. This procedure is illustrated by means of an example. It is found that the Maxwell model fits the experimental data better than the Kelvin Model. PMID- 11264831 TI - Finite Element Analysis of Simulations of Cancellous Bone Resorption. AB - A stochastic simulation of the resorption of cancellous bone has been developed and integrated with a finite element model to predict the resultant change in structural properties of bone as bone density decreases. The resorption represents the net imbalance of osteoclast and osteoblast activity that occurs in osteoporosis. A simple lattice structure of trabecular bone is considered, with an examination of the lattice geometry and discretization indicating that just five trabeculae need to be modelled. The results from the analysis show how the mechanical properties of the cancellous bone degrade with osteoporosis and demonstrate how the method can be used to predict the relationships between stiffness and density or porosity. PMID- 11264832 TI - Proximal Femoral Density Patterns are Consistent with Bicentric Joint Loads. AB - We developed an alternate method for density-based load estimation and applied it to estimate hip joint load distributions for two femora. Two-dimensional finite element models were constructed from single energy quantitative computed tomography (QCT) data. Load estimation was performed using five loading regions on the femoral head. Within each loading region, individual nodal loads, normal to the local surface, were supplied as input to the load estimation. An optimization procedure independently adjusted individual nodal load magnitudes in each region, and the magnitudes of muscle forces on the greater trochanter, such that the applied tissue stimulus approached the reference stimulus throughout the model. Dominant estimated load resultant directions were generally consistent with published experimental data for loads during gait. The estimated loads also suggested that loads near the extremes of the articulating surface may be important (even required) for development and maintenance of normal bone architecture. Estimated load distributions within nearly all regions predicted bicentric loading patterns, which are consistent with observations of hip joint incongruity. Remodeling simulations with the estimated loads predicted density distributions with features qualitatively similar to the QCT data sets. This study illustrates how applications of density-based bone load estimation can improve understanding of dominant loading patterns in other bones and joints. The prediction of bicentric loading suggests a very fine level of local adaptation to details of joint loading. PMID- 11264833 TI - A Three-Dimensional Finite Element Scheme to Investigate the Apparent Mechanical Properties of Trabecular Bone. AB - A computational technique is described for investigating the apparent mechanical properties of trabecular bone based on tissue geometry obtained from the marching cubes volume rendering scheme. Using this scheme, a 3D representation of the trabecular bone was extracted from two-dimensional cross-sections of the tissue originating from a quantitative serial sectioning procedure. Surface information consists of node coordinates and polygon connectivity in a 3D space. A custom, adaptive mesh generation technique using a normal offset was used to prepare 3D finite element volume meshes (4-node tetrahedral elements) of variable mesh density from the extracted surface geometry. Nine target mesh resolutions (32 um to 107 um) were examined for a (1.5 mmx 1.5 mmx 2 mm) volume of trabecular bone. A mesh density of 50,000 elements/mm(3) of bone tissue was found to be adequate for convergence of apparent (bulk) modulus for 1% uniaxial compression. For this convergent case, the maximum local normal compressive tissue stress was 400 MPa which was six hundred-fold greater than the computed apparent stress. Variation in the apparent modulus was less than 5% when Poisson's ratio values were varied between 0.1 and 0.4. Poisson's ratio values greater than 0.4 had a more marked effect on the apparent modulus. Based upon these results, approximately 1 million, 4-node tetrahedral elements are required to analyze a continuum scale model of trabecular bone (5 mm cube). PMID- 11264834 TI - Curvilinear Three-Dimensional Modeling of Spinal Curves with Dual Kriging. AB - In spinal deformation studies, three-dimensional reconstruction of the spine is frequently represented as a curve in space fitted to the vertebral centroids. Conventional interpolation techniques such as splines, Bezier and the least squares method are limited since they cannot describe precisely the great variety of spinal morphologies. This article presents a more general technique called dual kriging, which includes two mathematical constituents (drift and covariance) to adjust the interpolated functions to spinal deformity better. The cross validation technique was used to compare the parametric representations of spinal curves with different combinations of drift and covariance functions. Model validation was performed from a series of analytic curves reflecting typical scoliotic spines. Calculation of geometric torsion, a sensitive parameter, was done to evaluate the accuracy of the kriging models. The best model showed an absolute mean difference of 1.2 x 10(-5) (+/- 7.1 x 10(-5) ) mm(-1) between the analytical and estimated geometric torsions compared to 5.25 x 10(-3) (+/- 3.7 x 10(-2) ) mm(-1) for the commonly used least-squares Fourier series method, a significant improvement in spinal torsion evaluation. PMID- 11264835 TI - Elastohydrodynamic Lubrication of Porous Substrates. Application to Synovial Joint Analysis. AB - This work presents computational results for an elastohydrodynamic model with a porous wall. The model - inspired by the cartilage exudation phenomenon arising in living joints - allows the lubricating fluid to be transferred from the porous walls to the flow channel and viceversa. The system is represented through a strongly coupled set of nonlinear equations which involves hydrodynamic lubrication, plane elasticity and continuity in porous media. The solution technique employed is based on the finite element method, Newton iteration and parametric continuation processes. The predictions obtained uncover mechanisms that might be responsible for the extremely low friction coefficients found in synovial joints. PMID- 11264836 TI - The Obstacle-Set Method for Representing Muscle Paths in Musculoskeletal Models. AB - A computational method is introduced for modeling the paths of muscles in the human body. The method is based on the premise that the resultant muscle force acts along the locus of the transverse cross-sectional centroids of the muscle. The path of the muscle is calculated by idealizing its centroid path as a frictionless elastic band, which moves freely over neighboring anatomical constraints such as bones and other muscles. The anatomical constraints, referred to as obstacles, are represented in the model by regular-shaped, rigid bodies such as spheres and cylinders. The obstacles, together with the muscle path, define an obstacle set. It is proposed that the path of any muscle can be modeled using one or more of the following four obstacle sets: single sphere, single cylinder, double cylinder, and sphere-capped cylinder. Assuming that the locus of the muscle centroids is known for an arbitrary joint configuration, the obstacle set method can be used to calculate the path of the muscle for all other joint configurations. The obstacle-set method accounts not only for the interaction between a muscle and a neighboring anatomical constraint, but also for the way in which this interaction changes with joint configuration. Consequently, it is the only feasible method for representing the paths of muscles which cross joints with multiple degrees of freedom such as the deltoid at the shoulder. PMID- 11264837 TI - Analysis of Time-Varying Biological Data Using Rainflow Cycle Counting. AB - A wide range of biological investigations lead to time-history data. The characterization of such data can be difficult particularly in the presence of signal noise or superimposed signals. Several methods are described which can be brought to bear including FFT, thresholding, peak counting, and range counting. However, each of these approaches has significant disadvantages. In this paper we describe a novel method, known as rainflow cycle counting, for characterizing time varying biological time-history data in terms of spiking or oscillation amplitude and frequency. Rainflow counting is a straightforward algorithm for identifying complete cycles in the data and determining their amplitudes. The approach is simple, reliable, easily lends itself to automation, and robust even in the presence of superimposed signals or background noise. After describing the method, its use and behavior are demonstrated on three sample histories of intracellular calcium concentration in chondrocytes exposed to fluid shear stress. The method is also applied to a more challenging data set that has had an artificial random error included. The results demonstrate that the rainflow counting algorithm identifies signal oscillations and appropriately determines their amplitudes even when superimposed or distorted by background noise. These attractive properties make rainflow counting a powerful approach for quantifying and characterizing biological time histories. PMID- 11264838 TI - A Mathematical Formulation for 3D Quasi-Static Multibody Models of Diarthrodial Joints. AB - This study describes a general set of equations for quasi-static analysis of three-dimensional multibody systems, with a particular emphasis on modeling of diarthrodial joints. The model includes articular contact, muscle forces, tendons and tendon pulleys, ligaments, and the wrapping of soft tissue structures around bone and cartilage surfaces. The general set of equations governing this problem are derived using a consistent notation for all types of links, which can be converted conveniently into efficient computer codes. The computational efficiency of the model is enhanced by the use of analytical Jacobians, particularly in the analysis of articular surface contact and wrapping of soft tissue structures around bone and cartilage surfaces. The usefulness of the multibody model is demonstrated by modeling the patellofemoral joint of six cadaver knees, using cadaver-specific data for the articular surface and bone geometries, as well as tendon and ligament insertions and muscle lines of actions. Good accuracy was observed when comparing the model patellar kinematic predictions to experimental data (mean +/- stand. dev. error in translation: 0.63 +/- 1.19 mm, 0.10 +/- 0.71 mm, -0.29 +/- 0.84 mm along medial, proximal, and anterior directions, respectively; in rotation: -1.41 +/- 1.71 degrees, 0.27 +/- 2.38 degrees, -1.13 +/- 1.83 degrees in flexion, tilt and rotation, respectively). The accuracy which can be achieved with this type of model, and the computational efficiency of the algorithm employed in this study may serve in many applications such as computer-aided surgical planning, and real-time computer-assisted surgery in the operating room. PMID- 11264839 TI - Development and Experimental Validation of a Fluid/Structure-Interaction Finite Element Model of a Vacuum-Driven Cell Culture Mechanostimulus System. AB - A new fluid/structure-interaction finite element formulation is reported, by means of which reactive fluid stresses can be determined for what is currently the most widely used laboratory apparatus (the Flexercell Strain Unit) for delivering controlled in vitro mechanical stimuli to cultured cells. The apparatus functions by means of cyclic vacuum application to the undersurface of a membrane-like circular rubber substrate. When operated in its original embodiment (i.e., without axial constraint to substrate motion), the pulsatile vacuum causes appreciable pulsatile excursions (often several millimeters) of the substrate. The mechanical stimuli experienced by cells attached atop the substrate include not only substrate distention, but also potentially confounding reactive fluid stresses due to coupled motions of the overlying liquid culture nutrient medium. Since it is impractical to directly measure reactive fluid stress in such environments, a corresponding mathematical model has been developed. The formulation involves transient continuum finite element solutions for the nutrient medium flow field and for the deformation of the substrate, coupled at their mutual interface (the substrate culture surface). Besides the nonlinearities inherent in the flow field and substrate treatments per se, the numerical problem is complicated by the presence of moving boundaries at the nutrient free surface and at the nutrient/substrate interface, as well as by the need to enforce fluid/structure interaction throughout the duty cycle. Algorithmic considerations appropriate to achieving physically realistic numerical performance are reported, and a confirmatory laboratory validation experiment is described. PMID- 11264840 TI - Simulation of the Seated Postural Stability of Healthy and Spinal Cord-Injured Subjects Using Optimal Feedback Control Methods. AB - A two-dimensional, biomechanical computer model was developed, using the software package Working Model(TM), to simulate the postural control of seated individuals. Both able-bodied and spinal cord-injured subjects were represented. The model incorporated active control of the upper body through full-state feedback. Specifically, a linear quadratic regulator scheme was implemented in the model. Nonlinearities were included in the torque computations to mimic physiological constraints and disability. Interactions between the subject and the wheelchair were also included in the model. Simulation results were compared with those obtained from experiments in which the subjects had attempted to remain stable during the application of significant disturbance moments, similar to those experienced during braking in a vehicle. While subjects exhibited more complex control schemes, the model was able to simulate overall stability. Therefore, it is believed that the model could prove beneficial to future research examining the effects of various restraints on stability. PMID- 11264842 TI - Finite-Difference Time-Domain Simulator for Half-Space Bioacoustic Problems. AB - The finite-difference time-domain (FDTD) method provides a numerical technique for solving acoustic forward problems. The FDTD is particularly useful in geometries that encompass heterogeneities. Plane-wave modeling permits the simulation of many practical bioacoustic problems because the phase front in the focal zone of many acoustic transducers is nearly planar. The formulation presented in this paper provides a technique for solving half-space problems under local plane-wave illumination. PMID- 11264841 TI - Prediction of Individual Muscle Forces Using Lagrange Multipliers Method - A Model of the Upper Human Limb in the Sagittal Plane: I. Theoretical Considerations. AB - Analytical solutions of indeterminate problems formulated for biomechanical models with more than one degree of freedom (DOF) are rarely found. This paper is an extension of the investigations of a 1 DOF model (Raikova, 1996, J. Biomechanics, 763-772) for a more complex 3 DOF model. The proposed model of the human upper limb is in the sagittal plane and includes ten muscle elements, four of them being two-joint ones. The formulated optimization task is solved analytically using the method of Lagrange multipliers. It is supposed that the optimization function is complex but can be approximated by a weighted sum of the squared muscle forces, where the nature of the weight factors of the muscles are unknown. The proposed computational algorithm for determination of the unknown individual muscle forces and joint reactions is easily implemented and may be extended without difficulties for more DOF and muscles. The aim is to establish a means of investigation of the possible weight coefficients for different modelled situations, which will help in searching for their physiological interpretation and analytical description. PMID- 11264843 TI - A Powerful Algorithm to aid Cardiac Arrhythmia Diagnosis. AB - The Electrocardiogram (ECG), represents the electrical activity of the heart. It is characterised by a number of waves P, QRS, T which are correlated to the status of the heart activity. In this paper, the aim is to present a powerful algorithm to aid cardiac diagnosis. The approach used is based on a determinist method, that of the tree decision. However, the different waves of the ECG signal need to be identified and then measured following a signal to noise enhancement. Signal to noise enhancement is performed by a combiner linear adaptive filter whereas P, QRS, T wave identification and measurement are performed by a derivative approach. Results obtained on simulated and real ECG signals are shown to be highly, satisfactory in the aid of cardiac arrhythmia diagnosis, such as junctionnal escapes, blocks, etc. PMID- 11264844 TI - In Vivo Analysis of Heterogeneous Brain Deformation Computations for Model Updated Image Guidance. AB - Neurosurgical image-guidance has historically relied on the registration of the patient and preoperative imaging series with surgical instruments in the operating room (OR) coordinate space. Recent studies measuring intraoperative tissue motion have suggested that deformation-induced misregistration from surgical loading is a serious concern with such systems. In an effort to improve registration fidelity during surgery, we are pursuing an approach which uses a predictive computational model in conjunction with data available in the OR to update the high resolution preoperative image series. In previous work, we have developed an in vivo experimental system in the porcine brain which has been used to investigate a homogeneous finite element rendering of consolidation theory as a tissue deformation model. In this paper, our computational approach has been extended to include heterogeneous tissue property distributions determined from an image-to-grid segmentation scheme. Results produced under two different loading conditions show that heterogeneity in the stiffness properties and interstitial pressure gradients varied over a range of physiologically reasonable values account for 1-3% and 5-8% of the predicted tissue motion, respectively, while homogeneous linear elasticity is responsible for 60-70% of the surgically induced motion that has been recoverable with our model-based approach. PMID- 11264845 TI - Flow and Oxygen Transfer Characteristics of an Intravenous Membrane Oxygenator: A Computational Study. AB - Spectral element computational simulations of the conservation of mass, momentum and species equations are performed to investigate the flow and oxygen transfer characteristics of an Intravenous Membrane Oxygenator (IMO). The simulations consider a three-dimensional IMO computational model consisting of equally-spaced fibers, an elastic balloon with non-permeable walls positioned longitudinally within the vena cava, and a Newtonian and time-dependent incompressible flow. Flow characteristics and oxygen transfer parameters are determined for operating conditions of a stationary and a pulsating balloon. For the stationary balloon configuration the flow is two-dimensional, parallel, laminar and without secondary flows for the Reynolds number range of 5.7-455.2. Evaluations of the oxygen transfer characteristics for the stationary balloon indicate that the main transport mechanisms are diffusion and convection in the crosswise and streamwise directions, respectively. Additionally, evaluations of oxygen transfer rates and Sherwood numbers in this Reynolds number range indicate that the oxygen transfer rate reaches an asymptotic limit at relatively moderate Reynolds numbers. For the pulsating balloon, flow characteristic results demonstrate the existence of a strong secondary flow around the fiber, and between the balloon and the fiber. This secondary flow induces oscillatory crosswise and streamwise velocities and a seemingly random spanwise flow which enhances the flow mixing as well as the transport of oxygen from the fiber surface to the bulk flow. PMID- 11264846 TI - Prediction of Individual Muscle Forces Using Lagrange Multipliers Method - A Model of the Upper Human Limb in the Sagittal Plane: II. Numerical Experiments and Sensitivity Analysis. AB - Using the method of Lagrange multipliers an analytical solution of the optimization problem formulated for a two-dimensional, 3DOF model of the human upper limb has been described in Part I of this investigation. The objective criterion used is the following: [formula: see text], where F(i) -s are the muscle forces modelled and c(i) -s are unknown weight factors. This study is devoted to the numerical experiments performed in order to investigate which sets of the weight factors may predict physiologically reasonable muscle forces and joint reactions. A sensitivity analysis is also presented. The influence of: the gravity forces, different external loads applied to the hand, changes of the weight factors and of joint angle on the optimal solution is studied. A general conclusion may be drawn: using the above mentioned objective criterion, practically all motor tasks performed by the human upper limb may be described if the c(i) -s are properly chosen. These weight factors generally depend on the joint moments and must be different (their magnitudes as well as their signs) for agonistic muscles and for their antagonists. PMID- 11264847 TI - Finite Element Analysis of Tibial Implants - Effect of Fixation Design and Friction Model. AB - A three dimensional nonlinear finite element model was developed to investigate tibial fixation designs and friction models (Coulomb's vs nonlinear) in total knee arthroplasty in the immediate postoperative period with no biological attachment. Bi-directional measurement-based nonlinear friction constitutive equations were used for the bone-porous coated implant interface. Friction properties between the polyethylene and femoral components were measured for this study. Linear elastic isotropic but heterogeneous mechanical properties taken from literature were considered for the bone. The Tensile behaviour of polyethylene was measured and subsequently modeled by an elasto-plastic model. Based on the earlier finite element and experimental pull-out studies, pegs and screws were also realistically modeled. The geometry of every component was obtained through measurement. The PCA tibial baseplate with three different configurations was considered; one with three screws, one with one screw and two short inclined porous-coated pegs, and a third one with no fixation for the sake of comparison. The axial load of 2000N was applied through the femoral component on the medial plateau of articular insert. It was found that Coulomb's friction significantly underestimates the relative micromotion at the bone-implant interface. The lowest micromotion and lift-off were found for the design with screws. Relative micromotion and stress transfer at the bone-implant interface depended significantly on the friction model and on the baseplate anchorage configuration. Cortical and cancellous bones carried, respectively, 10-13% and 65 86% of the axial load depending on the fixation configuration used. The remaining portion was transmitted as shear force by screws and pegs. Normal and Mises stresses as well as contact area in the polyethylene insert were nearly independent of the baseplate fixation design. The Maximum Mises stress in the polyethylene exceeded yield and was found 1-2 mm below the contact surface for all designs. PMID- 11264848 TI - Use of Neural Networks to Correlate Spine and Rib Deformity in Scoliosis. AB - Artificial neural networks (ANN's) recognize patterns relating input and output data in a manner analogous to the function of biological neurons. Here, we show that ANN's can predict rib deformity in scoliosis more accurately than regression analysis. ANN's and linear regression models were developed to predict rib rotation from several combinations of input spinal indices including Cobb angle, vertebral rotation, apex location and orientation of the plane of maximal curvature. ANN's averaged 60% correct predictions compared to 34% for regression analysis. This study provides evidence for the utility of artificial neural networks in scoliosis research. These data lend credence to the use of ANN's in future work on the prediction of scoliotic spinal deformity from torso surface data, which would permit assessment of scoliosis severity with minimal use of harmful X-rays. PMID- 11264849 TI - Adaptive Finite Element Analysis of the Anisotropic Biphasic Theory of Tissue Equivalent Mechanics. AB - The nonlinear partial differential equations of the anisotropic biphasic theory of tissue-equivalent mechanics are solved with axial symmetry by an adaptive finite element system. The adaptive procedure operates within a method-of-lines framework using finite elements in space and backward difference software in time. Spatial meshes are automatically refined, coarsened, and relocated in response to error indications and material deformation. Problems with arbitrarily complex two-dimensional regions may be addressed. With meshes graded in high error regions, the adaptive solutions have fewer degrees of freedom than solutions with comparable accuracy obtained on fixed quasi-uniform meshes. The adaptive software is used to address problems involving an isometric cell traction assay, where a cylindrical tissue equivalent is adhered at its end to fixed circular platens; a prototypical bioartificial artery; and a novel configuration that is intended as an initial step in a study to determine bioartificial arteries having optimal collagen and cell concentrations. PMID- 11264850 TI - A Finite Element Approach for Skeletal Muscle using a Distributed Moment Model of Contraction. AB - The present paper describes a geometrically and physically nonlinear continuum model to study the mechanical behaviour of passive and active skeletal muscle. The contraction is described with a Huxley type model. A Distributed Moments approach is used to convert the Huxley partial differential equation in a set of ordinary differential equations. An isoparametric brick element is developed to solve the field equations numerically. Special arrangements are made to deal with the combination of highly nonlinear effects and the nearly incompressible behaviour of the muscle. For this a Natural Penalty Method (NPM) and an Enhanced Stiffness Method (ESM) are tested. Finally an example of an analysis of a contracting tibialis anterior muscle of a rat is given. The DM-method proved to be an efficient tool in the numerical solution process. The ESM showed the best performance in describing the incompressible behaviour. PMID- 11264851 TI - Sensitivity Analysis and Optimal Shape Design for Bone-Prosthesis Interfaces in a Femoral Head Surface Replacement. AB - A numerical optimization procedure has been applied for the shape optimal design of a femoral head surface replacement. The failure modes of the prosthesis that were considered in the formulation of the objective functions concerned the interface stress magnitude and the bone remodelling activity beneath the implant. In order to find a compromising solution between different requirements demanded by the two objective functions, a two step optimization procedure has been developed. Through step 1 the minimization of interface stress was achieved, through step 2 the minimization of bone remodelling was achieved with constraints on interface stresses. The results obtained provided an optimal design that generates limited bone remodelling activity with controlled interface stress distribution. The computational procedure was based on the application of the finite element method, linked to a mathematical programming package and a design sensitivity analysis package. PMID- 11264852 TI - Finite Element Analysis of Interbody Cages in a Human Lumbar Spine. AB - An innovative surgical procedure is vertebral stabilization by interbody cages. It is currently being used to separate and stabilize vertebral bodies and to promote bony fusion of the vertebrae onto or through the cages. This surgery, at some spine levels, can be performed through a laparoscope as an outpatient procedure with low morbidity. Because the procedure is new, little structural information is available on the interbody cages. The objective of this study was to evaluate the human lumbar spine stabilized by interbody cages biomechanically. The finite element method was used to compare cage designs by considering stresses in the cage and in the bone as well as relative displacements between the cage and the adjacent bone at the interface. The biomechanical evaluation considered different bone densities and considered axial, torsional, and bending loads on the lumbar spine. Stress analysis predicts local regions of stress concentration that could be damaging to cancellous bone and will likely require a remodeling response for local damage. This study predicts relative micromotion that could cause the bone resorption and fibrous tissue formation on the contact surfaces of the cage. The geometric constraints caused by the use of two cages will reduce the relative motion and therefore be more likely to allow bone ingrowth at the posterocentral contact region. Finite element analysis suggests that cages are a promising method for separation and stabilization of the vertebral bodies. PMID- 11264853 TI - A 3d-fem Simulation of Highest Stress Lines in Mandible Fractures by Elastic Impact. AB - In this article, the more usual mandible fracture areas are located by identifying the highest stress lines using a three-dimensional (tetrahedral) finite element method. By taking into account the temporomandibular contact and the inertia effects, the mathematical model is considered to be a dynamic Signorini's problem, that is, a dynamic variational inequality which is discretized in time following Newmark's method. So, in each time step a stationary variational inequality is solved by a penalty-duality algorithm. Finally, some numerical results obtained by simulating the more usual fractures in the human mandible are presented and compared with clinical experimental information. PMID- 11264854 TI - A Three-Dimensional Finite Element Analysis of Heat Transfer in the Forearm. AB - The finite element method was used to analyze heat transfer within a section of the forearm while exposed to different ambient conditions and with different metabolic states. The three-dimensional model accounts for the different material properties of bone, muscle and blood and incorporates a single artery-vein pair for counter-current heat exchange. The geometry of the model was developed from anatomical cross-sectional images of the forearm. The model was used to determine the effects or rest vs. exercise, free vs. forced surface convection and 0 degrees C vs. -20 degrees C external temperatures. The results of the model were compared to experimental data and the model exhibits qualitatively correct behaviour. This model can be used to study hyperthermia, burns and cryogenic freezing of tissue. PMID- 11264855 TI - Modelling the Brain for Rotational Loading: Shear Effects and Other Complications. AB - This study considers modelling the brain due to rotation of the skull where, at lower frequencies, the shear property of the material is important. Investigations reported here cover the effect of elastic and viscoelastic (lossy) cerebral material, the effect of the Falx protruding into the brain, the gap around the Falx and the brain filled with non viscous fluid in addition to different models of the Falx with bending or membrane stiffness. Analytical benchmark formulations are also described for the simple 2D plane strain in a cylinder produced by a half-sine rotation on the outer periphery which allows numerical (Finite Element) models to be validated. The results show the importance of the material properties, duration of loading and amplitude of loading as well as the influence of the partition. The results are shown for predicted maximum Principal strains in the models, as this may well be indicative of whether damage of the brain tissue occurs. PMID- 11264856 TI - Computational and Experimental Study of Performance of an Artificial Ligament: DacronTM yarn. AB - The success rate of reconstructing the Anterior Cruciate Ligament (ACL) with prosthetic ligaments is currently low both in humans and animals. The stress distribution in prosthetic ligaments that causes failure is very complex and not yet understood. Therefore, we have begun to develop a Finite Element Model of a prosthetic ACL. Here we describe the normal and contact stresses in Dacron(TM) yarn (a multi-fibrillar structure) using input data based on experimental measurements of the load and strain of six designed yarns. The results show that the normal and contact stresses in the fibres of the ACL yarn are directly proportional to the yarn strains. Increasing the twisting length (transverse deformation) of the yarn increases the normal stress in the fibres and the yarn modulus, but decreases the contact stresses between the fibres. The structural properties of a yarn are dependent on the specific arrangement of various filament types. Increasing the distance between the longitudinal (symmetry) axes of the filaments and the axis of symmetry of the yarn decreases the stresses. PMID- 11264857 TI - A Method for Numerical Simulation of Single Limb Ground Contact Events: Application to Heel-Toe Running. AB - The objective of this work was to develop a method to simulate single-limb ground contact events, which may be applied to study musculoskeletal injuries associated with such movements. To achieve this objective, a three-dimensional musculoskeletal model was developed consisting of the equations of motion for the musculoskeletal system, and models for the muscle force generation and ground contact elements. An optimization framework and a weighted least-squares objective function were presented that generated muscle stimulation patterns that optimally reproduced subject-specific movement data. Experimental data were collected from a single subject to provide initial conditions for the simulation and tracking data for the optimization. As an example application, a simulation of the stance phase of running was generated. The results showed that the average difference between the simulation and subject's ground reaction force and joint angle data was less than two inter-trial standard deviations. Further, there was good agreement between the model's muscle excitation patterns and experimentally collected electromyography data. These results give confidence in the model to examine musculoskeletal loading during a variety of landing movements and to study the effects of various factors associated with injury. Limitations were examined and areas of improvement for the model were presented. PMID- 11264858 TI - Mathematical Modeling and Numerical Simulation of Magnetic Susceptibility Artifacts in Magnetic Resonance Imaging. AB - The technique used to recognise information in Magnetic Resonance Imaging (MRI) is based on electromagnetic fields. A linearly varying field (around 10(-2) Tesla per meter) is added to a strong homogeneous magnetic field (order of magnitude of approximately one Tesla). When these fields are disturbed by the presence of a paramagnetic material, in the sample for instance, the resulting image is usually distorted, these distortions being termed artifacts. Our goal is to present a method, assuming the field disturbances are known, to construct the resulting images. A mathematical model of the MRI process is developed. The way the images are distorted in intensity and shape is explained and an algorithm to simulate magnetic susceptibility artifacts is deduced. PMID- 11264859 TI - Computational models of blood flow in the circle of Willis. AB - A two-dimensional, steady state model of the circle of Willis has been developed. To simulate the peripheral resistance of the cerebrovascular tree, blocks of porous media were used. Their effective resistance was kept constant, disregarding the effects of arterial auto-regulation. The model was then used to simulate different common abnormalities of the circle of Willis while a range of varying boundary conditions was imposed to the right internal carotid artery (ICA). The total flux was tabulated and compared favourably with both clinical measurements and other models of the circle of Willis. Relevant fluid dynamics effects were also observed and analysed. The present model demonstrates that the use of CFD can produce physiological results if the appropriate boundary conditions are used. We can provide clinicians with a priority list of the severity of the flux reduction for the considered abnormalities for different degrees of stenosis of the right ICA. From this study it is apparent that the redistribution of blood via the circle of Willis is mainly driven by changes in the vascular resistance of the brain rather than in the local arterial geometry. The use of valid peripheral resistances allows for a more realistic model of the circle of Willis but also highlights the need for more accurate means to estimate the vascular resistance of a patient. PMID- 11264860 TI - Image-guided surgery: from X-rays to virtual reality. AB - Since the discovery of X-rays, medical imaging has played a major role in the guidance of surgical procedures. While medical imaging began with simple X-ray plates to indicate the presence of foreign objects within the human body, the advent of the computer has been a major factor in the recent development of this field. Imaging techniques have grown greatly in their sophistication and can now provide the surgeon with high quality three-dimensional images depicting not only the normal anatomy and pathology, but also vascularity and function. One key factor in the advances in Image-Guided Surgery (IGS) is the ability not only to register images derived from the various imaging modalities amongst themselves, but also to register them to the patient. The other crucial aspect of IGS is the ability to track instruments in real time during the procedure, and to portray them as part of a realistic model of the operative volume. Stereoscopic and virtual-reality techniques can usefully enhance the visualization process. IGS nevertheless relies heavily on the assumption that the images acquired prior to surgery, and upon which the surgical guidance is based, accurately represent the morphology of the tissue during the surgical procedure. In many instances this assumption is invalid, and intra-operative real-time imaging, using interventional MRI, Ultrasound, and electrophysiological recordings are often employed to overcome this limitation. Although now in extensive clinical use, IGS is often currently perceived as an intrusion into the operating room. It must evolve towards becoming a routine surgical tool, but this will only happen if natural and intuitive human interfaces are developed for these systems. PMID- 11264861 TI - Reconstruction of laser-scanned 3D torso topography and stereoradiographical spine and rib-cage geometry in scoliosis. AB - Assessments of scoliosis are routinely done by means of clinical examination and full spinal x-rays. Multiple exposure to ionization radiation, however, can be hazardous to the child and is costly. Here, we explain the use of a noninvasive imaging technique, based on laser optical scanning, for quantifying the three dimensional (3D) trunk surface topography that can be used to estimate parameters of 3D deformity of the spine. The laser optical scanning system consisted of four BIRIS laser cameras mounted on a ring moving along a vertical axis, producing a topographical mapping of the entire torso. In conjunction with the laser scans, an accurate 3D reconstruction of the spine and rib cage were developed from the digitized x-ray images. Results from 14 scoliotic patients are reported. The digitized surfaces provided the foundation data to start studying concordance of trunk surface asymmetry and spinal shape in idiopathic scoliosis. PMID- 11264862 TI - Preliminary studies on the optimum shape of dental bridges. AB - Several pre-existing anterior and posterior dental bridge models using Finite elements and the new ceramic material In-Ceram have been developed. The mechanical behaviour of these models has been compared with optimised profiles obtained from a newly developed evolutionary algorithm known as Evolutionary Structural Optimisation (ESO). The results show that the mechanical behaviour of the bridges was mainly restricted by the properties of the porcelain veneer and the design of the bridges themselves. For the case of the anterior bridge, it was found that there existed a specific thickness of veneer that minimised the maximum principal stress. This was related to peak stresses that occurred at the bridge surface. Peak stresses also occurred in the material interface between the In-Ceram and the veneer. These extreme stresses were attributed to the notch size and shape. For the case of the posterior bridge, it was concluded that the shape of the bottom of the Pontic tooth is crucial in reducing the magnitude of the maximum principal tensile stress. The ESO process produced bridge designs which have uniformly stressed bridge surfaces, and which also have significantly lower maximum principal tensile stresses compared to the pre-existing designs (up to 44%). PMID- 11264863 TI - Musculoskeletal model of the upper limb based on the visible human male dataset. AB - A mathematical model of the human upper limb was developed based on high resolution medical images of the muscles and bones obtained from the Visible Human Male (VHM) project. Three-dimensional surfaces of the muscles and bones were reconstructed from Computed Tomography (CT) images and Color Cryosection images obtained from the VHM cadaver. Thirteen degrees of freedom were used to describe the orientations of seven bones in the model: clavicle, scapula, humerus, radius, ulna, carpal bones, and hand. All of the major articulations from the shoulder girdle down to the wrist were included in the model. The model was actuated by 42 muscle bundles, which represented the actions of 26 muscle groups in the upper limb. The paths of the muscles were modeled using a new approach called the Obstacle-set Method [33]. The calculated paths of the muscles were verified by comparing the muscle moment arms computed in the model with the results of anatomical studies reported in the literature. In-vivo measurements of maximum isometric muscle torques developed at the shoulder, elbow, and wrist were also used to estimate the architectural properties of each musculotendon actuator in the model. The entire musculoskeletal model can be reconstructed using the data given in this paper, along with information presented in a companion paper which defines the kinematic structure of the model [26]. PMID- 11264864 TI - A biomechanical model of the upper airways for simulating laryngoscopy. AB - This paper describes a three-dimensional finite element model of the human upper airways during rigid laryngoscopy. In this procedure, an anaesthetist uses a rigid blade to displace and compress the tongue of the patient, and then inserts a tube into the larynx to allow controlled ventilation of the lungs during an operation. A realistic model of the main biomechanical aspects involved would help anaesthetists in training and in predicting difficult cases in advance. For this purpose, the finite element method was used to model structures such as the tongue, ligaments, larynx, vocal cords, bony landmarks, laryngoscope blade, and their inter-relationships, based on data extracted from X-ray, MRI, and photographic records. The model has been used to investigate how the tongue tissue behaves in response to the insertion of the laryngoscope blade, when it is subjected to a variety of loading conditions. In particular, the mechanical behaviour of the soft tissue of the tongue was simulated, from simple linear elastic material to complex non-linear viscoelastic material. The results show that, within a specific set of tongue material parameters, the simulated outcome can be successfully related to the view of the vocal cords achieved during real laryngoscopies on normal subjects, and on artificially induced difficult laryngoscopy, created by extending the upper incisors teeth experimentally. PMID- 11264865 TI - Computer simulation of non-newtonian effects on blood flow in large arteries. AB - The influence of viscoelastic effects on blood flow in large arteries is studied numerically. The description of the blood flow uses the conservation of mass and momentum and a constitutive relation of Jeffreys' type (Oldroyd-B) and appropriate relations to describe the shear thinning behaviour. The steady flow studies are carried out in an axisymmetric tube with a local constriction modelling a stenosed blood vessel and in a three-dimensional 90 degrees curved tube. The numerical approach applies a decoupled technique where the computation of kinematics and stresses is separated. The governing equations are solved by means of an upwind stabilised Galerkin finite element method. The numerical results indicate significant influence of viscoelastic effects in the stenosed model. The flow through the curved tube shows minor quantitative viscoelastic influence. The influence of the shear thinning effect can be observed in both geometries. The results demonstrate that the viscoelastic behaviour of the local flow patterns in large arteries is dependent on the shape of the flow domain. PMID- 11264866 TI - Computation of a stabilizing set of feedback matrices of a large-scale nonlinear musculoskeletal dynamic model. AB - The purpose of this study is to present a general mathematical framework to compute a set of feedback matrices which stabilize an unstable nonlinear anthropomorphic musculoskeletal dynamic model. This method is activity specific and involves four fundamental stages. First, from muscle activation data (input) and motion degrees-of-freedom (output) a dynamic experimental model is obtained using system identification schemes. Second, a nonlinear musculoskeletal dynamic model which contains the same number of muscles and degrees-of-freedom and best represents the activity being considered is proposed. Third, the nonlinear musculoskeletal model (anthropomorphic model) is replaced by a family of linear systems, parameterized by the same set of input/output data (nominal points) used in the identification of the experimental model. Finally, a set of stabilizing output feedback matrices, parameterized again by the same set of nominal points, is computed such that when combined with the anthropomorphic model, the combined system resembles the structural form of the experimental model. The method is illustrated in regard to the human squat activity. PMID- 11264867 TI - Development of articular cartilage: what do we know about it and how may it occur? AB - Articular cartilage has a fundamental role in joint function. While much is known about its structure, organization and biomechanical properties, there is a very poor understanding of how articular chondrocytes develop during embryogenesis and acquire the unique ability to organize and maintain the articular tissue. Given that articular cartilage forms in close juxtaposition with the joint, here we review past studies on limb joint determination and morphogenesis and more recent studies on a number of factors thought to have roles in joint and epiphysis development. These factors include: the homeobox gene Barx-1; the bone morphogenetic protein (BMP) family member GDF-5; the growth factors HGF and PTHrP; and the transcription factor ERG. We summarize current thinking on how these factors participate in joint development and how some of these factors may influence development and behavior of epiphyseal chondrocytes. We also describe pertinent recent studies from our laboratories on ERG and the newly-identified alternatively spliced variant C-1-1, and finally propose a sequela of events that may subtend the process of determination and emergence of articular chondrocytes during limb synovial joint development. PMID- 11264868 TI - Localization of cathepsin D in human odontoclasts. a light and electron microscopical immunocytochemical study. AB - Odontoclasts are dentine and cementum resorbing cells whose relationship to bone resorbing osteoclasts is not clear. Like osteoclasts, they possess different cathepsins which are involved in mineralized tissue degradation during the tooth root resorption process in deciduous teeth. Whether cathepsin D, which in osteoclasts probably functions as an activator of other cathepsins, can be found in odontoclasts, has, however, not been investigated before. In order to determine its occurrence and localization, cathepsin D immunocytochemistry was applied to paraffin-embedded sections from 30 human deciduous tooth roots undergoing resorption. Using immunogold postembedding immunocytochemsitry on LR Gold embedded specimens, the distribution of cathepsin D was investigated at the ultrastructural level. We identified tartrate-resistent acid phosphatase-positive mono- and multinuclear odontoclasts near and on the periodontal surfaces of tooth roots. Nearly all of these cells showed cytoplasmic granular cathepsin D immunoreactivity. At the electron microscopical level, gold labelling was seen on vacuoles and vesicles of the odontoclasts, which were identified as secondary lysosomes and phagosomes. Extracellularly it was seen along the ruffled border and in neighboured resorption areas of dentine and cementum. These findings indicate that cathepsin D is secreted into the resorbing area of human odontoclasts in order to participate in degradation of mineralized tooth matrix, but may also function as an activator of other proteases in lysosomal organelles. PMID- 11264869 TI - Molecular and cell biology of skin wound healing in a pig model. AB - To define the pattern of change at the molecular and cellular levels during the healing of excisional skin wounds in the skeletally immature pig, mRNA levels for relevant molecules were assessed by semiquantitative RT-PCR using porcine specific primer sets and RNA isolated from normal skin and samples at various time post-wounding. Analysis of cellular change was assessed by DNA quantification and histology of tissue sections. The results demonstrated that the changes in the pattern of RNA and DNA content of the scar tissue were consistent with the observed increasing cellularity. The mRNA levels for collagen I, III, HSP47, IL-1, TGF-beta, MMP-1, -2 and -9, TIMP-1, -2, and-4, PAI-1, versican were significantly elevated during healing; levels for biglycan and fibromodulin were not significantly altered; and the mRNA levels for TIMP-3 were depressed. These findings suggest that skin wound healing is a series of complex matrix-cell interactions that involve cellular migration and inflammation, followed by proliferation of fibroblasts with new collagen synthesis, and lastly tissue remodeling of the scar. PMID- 11264870 TI - Possible involvement of altered RGD sequence in reduced adhesive and spreading activities of advanced glycation end product-modified fibronectin to vascular smooth muscle cells. AB - Although fibronectin (FN) modified by advanced glycation end products (AGEs) has been shown to contribute to the development of diabetic vascular complications through its reduced adhesive activity to vascular cells, little is known about changes in the cell binding domain of AGE-modified FN. Here we examined the mechanism of reduced adhesive and spreading activities of AGE-modified FN to vascular smooth muscle cells (SMCs), particularly the contribution of modification of Arg-Gly-Asp (RGD) sequence. Incubation with glucose caused not only the formation of N(epsilon) -carboxymethyllysine and pentosidine, but also polymerization of FN in a dose- and time-dependent manner. AGE-modified FN had significantly low adhesive and spreading activities to cultured SMCs. On the other hand, multimeric FN formed by disulfide bonds did not show any effect on either cell adhesion or spreading. The adhesive activity of type I collagen, one of the RGD sequence-containing proteins, to SMCs also decreased by AGE modification. The inhibitory effect of AGE-modification on cell adhesion was significantly greater in type I collagen than in FN. Although the extent of AGE modification of type I collagen was indistinguishable from that of FN, AGE modification decreased the arginine content of type I collagen by 69.5% and of FN by 30.6%, compared with their non-glycated forms. The addition of RGD peptides caused a decrease in adhesion of SMCs to non-glycated FN, but not to AGE-modified FN. Modification of RGD sequence with glyoxal eliminated its inhibitory effect on cell adhesion. Our results suggest that a marked decrease in adhesive and spreading activities of AGE-modified FN to SMCs might largely be due to a modification of its RGD sequence by AGE, thus suggesting a potential link between AGE modification of FN and the pathogenesis of diabetic angiopathy. PMID- 11264871 TI - Exploring the effects of hypermineralisation in bone tissue by using an extreme biological example. AB - The properties of bone tissue with very high or very low mineral levels attract attention because they allow researchers to comprehend more fully the mechanics, interaction and effects of mineral on collagen through a greater range of compositions than that found in the "ordinary". The bone tissue of the rostrum of the whale Mesoplodon densirostris is the densest bone known. We examined the composition, static and fatigue strength, hardness and toughness of this tissue and compared them to those of other less mineralised analogues. The rostrum bone has remarkably little organic matter and retains very little water in its native state, but its basic mineral stoichiometry is very similar to that of other bones. We present here updated versions of the microhardness vs. modulus and microhardness vs. mineral fraction relationships, which thanks to the rostrum have been produced for a considerably wider range than in the past. We found the rostrum to be extremely brittle with a toughness ratio in two perpendicular directions (along and across its length) similar to that of tissue of other "ordinary" long bones and we discuss the possible significance of our findings. PMID- 11264872 TI - The isolation and detection of non-collagenous proteins from the compact bone of the dinosaur Iguanodon. AB - This report describes the isolation of guanidinium chloride extractable protein from demineralised bone extracts obtained from the 125-130 mya dinosaur Iguanodon. Protein products were isolated in the Mr. range 5,000-66,000 using SDS PAGE and represent the first electrophoretically defined proteins isolated from dinosaur tissues. The levels of glycine, aspartate and serine tentatively suggest the presence of phosphoproteins. Hydroxylysine and hydroxyproline were not detected, confirming the presence of non-collagenous material. In addition the absence of ornithine confirmed lack of bacterial contamination. The relatively high level of leucine in the 2MNaCl NaCl fractions together with the abolition of alcian blue reactivity following protease-free chondroitinase digestion suggests the presence of proteoglycans. The study is of interest in describing the early proteins laid down in mineralised tissues for epitactic crystal growth and may provide evidence on evolutionary aspects of bone proteins. PMID- 11264889 TI - Uric acid mediates photodynamic inactivation of caprine alpha-2-macroglobulin. AB - Uric acid (2,6,8 trioxopurine), the end product of purine metabolism in mammalian systems, has shown a wide range of antioxidant properties including scavenging of hydroxyl radical and singlet oxygen. In this study we show that in the presence of visible light, uric acid disrupted caprine alpha-2-macroglobulin (alpha(2) M) structure and antiproteolytic function in vitro. Proteinase cleaves the bait region of caprine inhibitor inducing major conformational changes and entrapping the enzyme within its molecular cage. In contrast to native alpha(2) M, modified antiproteinase lost half of its antiproteolytic potential within 4 hours of uric acid exposure. The changes in uv-absorption spectra of the treated protein suggested possible spatial rearrangement of subunits or conformational change. Analysis of the mechanism by which alpha(2) M was inactivated revealed that the process was dependent on generation of superoxide anion and hydrogen peroxide. Our findings suggest that antiproteolytic activity of caprine alpha(2) M could be compromised via oxidative modification mediated by uric acid. Moreover, low concentrations of alpha(2) M were found to stimulate superoxide production by some unknown mechanism. PMID- 11264891 TI - Restoration of the cellular thiol status of peritoneal macrophages from CAPD patients by the flavonoids silibinin and silymarin. AB - During continuous ambulatory peritoneal dialysis (CAPD) the peritoneal immune cells, mainly macrophages, are highly compromised by multiple factors including oxidative stress, resulting in a loss of functional activity. One reason for the increase of inflammatory reactions could be an imbalance in the thiol-disulfide status. Here, the possible protective effects of the antioxidant flavonoid complex silymarin and its major component silibinin on the cellular thiol status were investigated. Peritoneal macrophages from dialysis fluid of 30 CAPD patients were treated with silymarin or silibinin up to 35 days. A time-dependent increase of intracellular thiols was observed with a nearly linear increment up to 2.5 fold after 96 hours, reaching a maximum of 3.5-fold after 20 days of culture. Surface-located thiols were also elevated. The stabilization of the cellular thiol status was followed by an improvement of phagocytosis and the degree of maturation as well as significant changes in the synthesis of IL-6 and IL-1ra. Furthermore, the treatment of peritoneal macrophages with flavonoids in combination with cysteine donors resulted in a shortened and more efficient time course of thiol normalization as well as in a further increased phagocytosis. In addition, GSH-depletion in thiol-deficient media simulating CAPD procedures led to intracellular thiol deficiency similar to the in vivo situation. It is concluded that treatment with milk thistle extracts silymarin and silibinin alone or, more effectively in combination with cysteine donors, provide a benefit for peritoneal macrophages of CAPD-patients due to a normalization and activation of the cellular thiol status followed by a restoration of specific functional capabilities. PMID- 11264890 TI - Protection of erythrocytes by the macrophage synthesized antioxidant 7,8 dihydroneopterin. AB - Neopterin and the reduced form, 7,8-dihydroneopterin (78NP) are pteridines released from macrophages when stimulated with gamma-interferon in vivo. The role of 78NP in inflammatory response is unknown though neopterin has been used clinically as a marker of immune cell activation, due to its very fluorescent nature. Using red blood cells as a cellular model, we demonstrated that micromolar concentrations can inhibit or reduce red blood cell haemolysis induced by 2,2'-azobis(amidinopropane)dihydrochloride (AAPH), hydrogen peroxide, or hypochlorite. One hundred microM 78NP prevented HOCl haemolysis using a high HOCl concentration of 5 micromole HOCl/10(7) RBC. Fifty microM 78NP reduced the haemolysis caused by 2 mM hydrogen peroxide by 39% while the same 78NP concentration completely inhibited haemolysis induced by 2.5 mM AAPH. Lipid peroxidation levels measured as HPLC-TBARS were not affected by addition of 78NP. There was no correlation between lipid oxidation and cell haemolysis suggesting that lipid peroxidation is not essential for haemolysis. Conjugated diene measurements taken after 6 and 12 hour exposure to hydrogen peroxide support the TBARS data. Gel electrophoresis of cell membrane proteins indicated 78NP might inhibit protein damage. Using dityrosine as an indicator of protein damage, we demonstrated 200 microM 78NP reduced dityrosine formation in H(2) O(2) /Fe(++) treated red blood cell ghosts by 30%. HPLC analysis demonstrated a direct reaction between 78NP and all three oxidants. Two mM hydrogen peroxide oxidised 119 nM of 78NP per min while 1 mM AAPH only oxidised 50 nM 78NP/min suggesting that 78NP inhibition of haemolysis is not due to 78NP scavenging the primary initiating reactants. In contrast, the reaction between HOCl and 78NP was near instant. AAPH and hydrogen peroxide oxidised 78NP to 7,8-dihydroxanthopterin while hypochlorite oxidation produced neopterin. The cellular antioxidant properties of 78NP suggest it may have a role in protecting immune cells from free radical damage during inflammation. PMID- 11264892 TI - Lucigenin chemiluminescence in human seminal plasma. AB - Seminal plasma protects spermatozoa from the detrimental effects of reactive oxygen species such as hydrogen peroxide. We investigated the lucigenin-dependent chemiluminescence in cell-free seminal plasma from andrological patients. The seminal plasma was separated from cells by centrifugation. In all seminal plasmas studied lucigenin-dependent chemiluminescence (LCL) was detected. The LCL showed a strong pH-dependence. The signal was stable if samples were stored at +4 degrees C for up to 4 days or up to 8 days at -80 degrees C. Filtration of the samples (0.45 and 0.22 microm pore size) did not lower their luminescence. The addition of superoxide dismutase (SOD) and ascorbic acid oxidase (AAO) lowered LCL nearly to baseline values while trolox and desferal showed moderate effect, whereas allopurinol had no effect. Electron paramagnetic resonance spectroscopy demonstrated ascorbyl radicals in seminal plasma. Physiological concentrations of ascorbic acid yielded SOD-inhibitable lucigenin-chemiluminescence. The nitroblue tetrazolium assay showed that ascorbic acid in buffer solution produced formazan. Superoxide-anion radicals were not detected in seminal plasma by the spin-trap DEPMPO due to their low steady state concentration. It is concluded that in seminal plasma ascorbate reacts with molecular oxygen yielding ascorbyl radicals and superoxide anion. If lucigenin is added to seminal plasma, reducing substances present, such as ascorbate, reduce lucigenin to the corresponding radical; this radical reacts with molecular oxygen and also forms O2-. So LCL in human seminal plasma results from the autoxidation of ascorbate and the oxidation of the reduced lucigenin. While the physiological relevance of the former mechanism is unknown, the latter is an artifact. PMID- 11264893 TI - Phenylbutazone radicals inactivate creatine kinase. AB - Creatine kinase (CK) was used as a marker molecule to examine the side effect of damage to tissues by phenylbutazone (PB), an effective drug to treat rheumatic and arthritic diseases, with horseradish peroxidase and hydrogen peroxide (HRP H(2)O2). PB inactivated CK during its interaction with HRP-H(2) O(2), and inactivated CK in rat heart homogenate. PB carbon-centered radicals were formed during the interaction of PB with HRP-H(2)O2. The CK efficiently reduced electron spin resonance signals of the PB carbon-centered radicals. The spin trap agent 2 methyl-2-nitrosopropane strongly prevented CK inactivation. These results show that CK was inactivated through interaction with PB carbon-centered radicals. Sulfhydryl groups and tryptophan residues in CK were lost during the interaction of PB with HRP-H(2)O2, suggesting that cysteine and tryptophan residues are oxidized by PB carbon-centered radicals. Other enzymes, including alcohol dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, but not lactate dehydrogenase, were also inactivated. Sulfhydryl enzymes seem to be sensitive to attack by PB carbon-centered radicals. Inhibition of SH enzymes may explain some of the deleterious effects induced by PB. PMID- 11264894 TI - Chemical evaluation of compounds as nitric oxide or peroxynitrite donors using the reactions with serotonin. AB - The aim of this work was to assess the capacities of some .NO-donors to release .NO, and consequently NOx in aerobic medium, or to give peroxynitrite. The method was based on the differential reactivity of serotonin (5-HT) with either NO(x) or peroxynitrite, leading in phosphate-buffered solutions to 4-nitroso- and 4-nitro 5-HT formation, respectively. Yields and formation rates of 5-HT derivatives with .NO-donor were compared to those obtained with authentic .NO or peroxynitrite in similar conditions. Aside from the capacity of diazenium diolates (SPER/NO and DEA/NO) to release .NO spontaneously, converting 5-HT exclusively to 4-nitroso-5 HT, all other .NO donors must undergo redox reactions to produce .NO. S nitrosoglutathione (GSNO) and sodium nitroprusside (SNP) modified 5-HT only in the presence of Cu2+, GSNO yielding 6 times more 4-nitroso-5-HT than SNP. Furthermore, in the presence of Cu+, the yield of .NO-release from GSNO was 45%. The molsidomine metabolite (SIN-1), which was presumed to release both .NO and O2(7-) at pH 7.4, reacted with 5-HT differently, depending on the presence of reductant or oxidant. Under aerobic conditions, SIN-1 acted predominantly as a 5 HT oxidant and also as a poor .NO and peroxynitrite donor (15% yield of .NO release and 14 % yield of peroxynitrite formation). The strong oxidant Cu2+, even in the presence of air oxygen, accelerated oxidation and increased .NO release from SIN-1 up to 86%. Only a small part of SIN-1 gave simultaneously .NO and O2(7 ) able to link together to give peroxynitrite, but other oxidants could enhance .NO release from SIN-1. PMID- 11264895 TI - Induction of heme oxygenase 1 in liver of spontaneously diabetic rats. AB - It has been suggested that diabetes induces an increase in oxidative stress; the increased expression of heme-oxygenase 1 (HO-1) in liver is believed to be a sensitive marker of the stress response. The aim of this study was to examine whether diabetes is able to induce HO-1 expression in liver. The specific mRNA was amplified by RT/PCR and calibrated with amplified beta-actin mRNA. The mRNA HO-1 levels in the liver of spontaneously diabetic rats were increased by 1.8 fold compared with non diabetics; this supports the hypothesis of weak but significant oxidative damage due to chronic hyperglycaemia. This work represents the first in vivo study exploring the semi-quantitative expression of HO-1 in the liver of spontaneously diabetic rats. PMID- 11264897 TI - Protective effects of vitamins C and E on the number of micronuclei in lymphocytes in smokers and their role in ascorbate free radical formation in plasma. AB - Cigarette smoke is widely believed to increase free radical concentrations causing subsequent oxidative processes that lead to DNA damage and hence, to several diseases including lung cancer and atherosclerosis. Vitamin C is a reducing agent that can terminate free-radical-driven oxidation by being converted to a resonance-stabilized free radical. To investigate whether short term supplementation with the antioxidants vitamin C and E decreases free-radical driven oxidation and thus decreases DNA damage in smokers, we determined the frequency of micronuclei in lymphocytes in 24 subjects and monitored the electron paramagnetic resonance signal of ascorbate free radical formation in plasma. Further parameters comprised sister-chromatid exchanges and thiobarbituric acid reactive substances. Twelve smokers and twelve non-smokers took 1000 mg ascorbic acid daily for 7 days and then 1000 mg ascorbic acid and 335.5 mg RRR-alpha tocopherol daily for the next 7 days. Baseline concentrations of both vitamins C and E were lower and baseline numbers of micronuclei were higher (p < 0.0001) in smokers than in non-smokers. After 7 days of vitamins C and E, DNA damage as monitored by the number of micronulei was decreased in both, smokers and non smokers, but it was more decreased in smokers as indicated by fewer micronuclei in peripheral lymphocytes (p < 0.05). Concomitantly, the plasma concentrations of vitamin C (p < 0.001) as well as the ascorbate free radical (p < 0.05) were increased. The corresponding values in non-smokers, however, did not change. Our findings show that increased ascorbate free radical formation in plasma after short-term supplementation with vitamins C and E can decrease the number of micronuclei in blood lymphocytes and thus DNA damage in smokers. PMID- 11264896 TI - The role of nitric oxide in paraquat-induced cytotoxicity in the human A549 lung carcinoma cell line. AB - Paraquat (PQ) is a well-known pneumotoxicant that exerts its toxic effect by elevating intracellular levels of superoxide. In addition, production of pro inflammatory cytokines has possibly been linked to PQ-induced inflammatory processes through reactive oxygen species (ROSs) and nitric oxide (NO). However, the role of NO in PQ-induced cell injury has been controversial. To explore this problem, we examined the effect of NO on A549 cells by exposing them to the exogenous NO donor NOC18 or to cytokines; tumor necrosis factor-alpha, interleukin-1 beta and interferon-gamma, as well as PQ. Although the exogenous NO donor on its own had no effect on the release of lactate dehydrogenase (LDH), remarkable release was observed when the cells were exposed to high concentrations of NOC18 and PQ. This cellular damage caused by 1 mM NOC18 plus 0.2 mM PQ was ascertained by phase contrast microscopy. On the other hand, NO derived from 25-50 microM NOC18 added into the medium improved the MTT reduction activity of mitochondria, suggesting a beneficial effect of NO on the cells. Incubation of A549 cells with cytokines increased in inducible NO synthase (iNOS) expression and nitrite accumulation, resulting in LDH release. PQ further potentiated this release. The increase in nitrite levels could be completely prevented by NOS inhibitors, while the leakage of LDH was not attenuated by the inhibition of NO production with them. On the other hand, ROS scavenging enzymes, superoxide dismutase and catalase, inhibited the leakage of LDH, whereas they had no effect on the increase in the nitrite level. These results indicate that superoxide, not NO, played a key role in the cellular damage caused by PQ/cytokines. Our in vitro models demonstrate that NO has both beneficial and deleterious actions, depending on the concentrations produced and model system used. PMID- 11264898 TI - Iron induces Bcl-2 expression in human dermal microvascular endothelial cells. AB - Iron is suspected to be involved in the induction and/or progression of various human tumors. The present study was designed to investigate the effects of iron on endothelial cells, keeping in mind that the homeostasis of microvessels plays a critical role in neo-angiogenesis. Applying a model of human dermal microvascular endothelial cell terminal differentiation and death induced by serum deprivation, we found that iron salts (iron chloride and ferric nitrilotriacetate) provided a survival advantage to endothelial cells. Using immunohistochemistry and Western Blot analysis, we found that the extended cellular life span induced by iron was paralleled by an increase of Bcl-2 protein expression. Taken together, these observations suggest that iron may give a survival advantage to endothelial cells and represent a novel mechanism through which iron may contribute to tumorigenesis. PMID- 11264899 TI - Characteristics and use as spin trapping agent of a beta-phosphorylated nitroso compound, DEPNP. AB - 2-(Diethylphosphonate)-nitrosopropane (DEPNP), prepared by oxidation of the corresponding aminophosphonate, was found to essentially exist as monomer in both water and organic solvents. The mechanisms of its degradation under 80 degrees C heating or visible light exposure were studied by EPR spectroscopy: its decomposition gave rise to paramagnetic by-products, which have been identified as DEPNP/.C(CH(3) )(2) [P(O)(OC(2) H(5) )(2) ] and DEPNP/.P(O)(OC(2) H(5) )(2) spin adducts. Despite this drawback, DEPNP was successfully used as spin trapping agents to scavenge various carbon - and phosphorus-centred free radicals both in aqueous and organic media, giving rise to intense EPR spectra characteristic of the species trapped. PMID- 11264901 TI - Nitroxide radicals. Controlled release from and transport through biomimetic and hollow fibre membranes. AB - Stable nitroxide radicals have found wide applications in chemistry and biology and they have some potential applications in medicine due to their antioxidant properties. Nitrocellulose filters impregnated with lipid-like substances are used as an imitation of biomembranes and could be used as a controlled drug release vehicle, while experiments with hollow fibres can be useful in the modelling of a drug delivery via blood vessels. This paper describes mechanisms of the nitroxide transport in four different model systems, i.e. a) exit of nitroxide into aqueous solution from porous nitrocellulose filters, impregnated with organic solvents, b) transport of nitroxides through the impregnated membrane from one into another aqueous solution, c) transport of nitroxides from bulk phase of organic solvents through the impregnated membrane into aqueous phase with ascorbic acid, and d) transport of nitroxides from liquid organic phase into aqueous solution through porous hollow fibres. The results are analysed in terms of mass transfer resistance of a membrane, organic and aqueous phase, based on nitroxide diffusion and distribution coefficients. Ascorbic acid reduced nitroxides in water and enhanced the rate of their transfer due to the decrease of transport resistance of unstirred aqueous layers. It is demonstrated that in the case of biomembranes the rate limiting step could be the transport through unstirred aqueous layers and membrane/water interface. PMID- 11264900 TI - Alteration of antioxidants during the progression of heart disease in streptozotocin-induced diabetic rats. AB - Involvement of oxidative stress is implicated in the progression of complication of diabetes mellitus. With respect to heart diseases, we have studied role of oxidative stress/antioxidants using rats treated with streptozotocin to induce diabetes (DM). Hemodynamic and echocardiographic measurements showed thickening of the wall and an increase in the internal dimension of the left ventricle (LV) in DM rats at 8th week. Decrease in diastolic posterior wall velocity and rate of LV pressure change, and increase in LV end diastolic pressures also proved cardiac dysfunction. These changes were further developed in DM rats after 12 weeks. Utilizing rat hearts at 8th and 12th weeks, the following estimations were performed. There was a decrease in the activity of Mn-superoxide dismutase (SOD), suggesting abnormal mitochondrial metabolism of reactive oxygen species. The level of glutathione (GSH) decreased concomitant with a decrease in the expression of gamma-glutamylcysteine synthetase (gamma-GCS). The expression of transforming growth factor-beta1 (TGF-beta1), known as a growth factor and a suppressor of GSH synthesis, elevated in DM rat hearts. Immunohistochemical estimation showed an increase in type IV collagen in DM hearts. Collectively, it was suggested a linkage between mitochondrial damage to generate reactive oxygen species and inactivation of Mn-SOD and elevation of the expression of TGF-beta1 to lead suppression of GSH synthesis and induction of fibrous change for the consequent cardiac dysfunction in DM. PMID- 11264902 TI - Protection against nitrofurantoin-induced oxidative stress by coelenterazine analogues and their oxidation products in rat hepatocytes. AB - Coelenterazine (3,7-dihydro-2-(p-hydroxybenzyl)-6-(p-hydroxyphenyl)-8 benzylimidazolo[1,2-a]pyrazin-3- one) is a substrate for the bioluminescence reaction in many marine animals. Recent work showed that CLZn, its synthetic analogue CLZm, and their common oxidation product coelenteramine (CLM) have strong antioxidative properties in acellular lipid peroxidation systems as well as in rat hepatocytes subjected to tert-butyl hydroperoxide (t-BHP). Here, we analyzed the ability of CLZm and several imidazolopyrazinone (IMPZs) analogues to protect primary cultures of rat hepatocytes against a nitrofurantoin (NF)-induced oxidative stress. Comparison of protection capabilities with reference antioxidants yielded the following ranking: CLZm >>> BHT >Trolox C((R)) > probucol > alpha-tocopherol. The comparison of CLZm with analogues lacking the phenol group in R(1) revealed no differences although the presence of this phenol conferred superior protection against t-BHP. CLM, as well as its methoxylated analogue mCLM which lacks chain-breaking properties, were equally potent in preventing cellular damage caused by NF. mCLM and alpha-naphthoflavone, an inhibitor of cytochrome P450 (CYP450) IAI, similarly protected cells against NF induced mortality and also equally inhibited EROD activity in methylcholanthrene induced hepatocytes. The inhibition of EROD by CLZm and CLM was less pronounced. We suggest that the extent of protection conferred by IMPZs against NF-toxicity reflects both the occurrence of antioxidative properties detoxifying ROS produced within cells and inhibitory actions on CYP450 isoforms involved in the bioreduction of NF. PMID- 11264903 TI - Addition of milk does not affect the absorption of flavonols from tea in man. AB - Tea is a major source of flavonols, a subclass of antioxidant flavonoids present in plant foods which potentially are beneficial to human health. Milk added to tea, a frequent habit in the United Kingdom, could inhibit absorption of tea flavonoids, because proteins can bind flavonoids effectively. Eighteen healthy volunteers each consumed two out of four supplements during three days: black tea, black tea with milk, green tea and water. A cup of the supplement was consumed every 2 hours each day for a total of 8 cups a day. The supplements provided about 100 micromol quercetin glycosides and about 60 - 70 micromol kaempferol glycosides. Addition of milk to black tea (15 ml milk to 135 ml tea) did not change the area under the curve of the plasma concentration-time curve of quercetin or kaempferol. Plasma concentrations reached were about 50 nM quercetin and 30 - 45 nM kaempferol. We conclude that flavonols are absorbed from tea and that their bioavailability is not affected by addition of milk. PMID- 11264904 TI - Hydroxytyrosol, as a component of olive mill waste water, is dose- dependently absorbed and increases the antioxidant capacity of rat plasma. AB - Hydroxytyrosol is the most potent phenolic antioxidant of olive oil and olive mill waste water (OMWW) and its biological activities have stimulated research on its potential role in cardiovascular protection. However, evidence of the absorption of OMWW phenolics and on their possible in vivo activity has, until now, never been provided. Three groups male Sprague-Dawley rats were administered 1, 5, or 10 mg/Kg of the OMWW extract, respectively, providing 41.4, 207, and 414 microg/Kg of hydroxytyrosol, respectively. Urine was collected for 24 h and the urinary levels of hydroxytyrosol were quantified by mass spectrometry. Hydroxytyrosol was dose-dependently (R(2) = 0.95) absorbed and excreted in the urines mostly as a glucuronide conjugate. Further, the administration of an hydroxytyrosol-rich OMWW extract (10 mg/kg) to the rats was also associated with an increase of their plasma antioxidant capacity. Future experiments will eventually further clarify its metabolic fate and its in vivo actions. PMID- 11264905 TI - Effect of a reduced base of support in standing and balance training on the soleus H-reflex. AB - The present study provides evidence that a reflex at the segmental level can adapt over a two hour experiment in a functionally appropriate manner in response to a balance training task. Subjects (N=9) received soleus (S) H-reflexes in blocks of seven trials while free standing on a normal base-of-support (NBOS) and while standing on a plafform with a reduced base-of-support (RBOS) in the sagittal plane. During the RBOS condition, the H-reflex served as a postural perturbation. Subjects were instructed to suppress the H-reflex when it was evoked, as an attempt to maintain a balanced state. Background EMG from the S and tibialis anterior muscles, the S M-wave, and stimulus current were maintained at a constant level during the experiment. Subjects initially received a block of NBOS trials, followed by 4 RBOS blocks (training), a second NBOS block, four additional RBOS blocks, and a third NBOS block with the protocol repeated on three different days (D1, D2 and D3) within the same week. The S H/M ratio was depressed 9% upon standing on the RBOS. With training the S H/M ratio decreased by 22% on Dl, 18% on D2 and 6% on D3. The ratio between the H-reflex and background S EMG (H-reflex gain) decreased 10% on D1, 40% on D2 and 23% on D3 when the first NBOS and first RBOS blocks were compared. Due to a slight increase in the S EMG across blocks, the H-reflex gain decreased considerably more across blocks than the H/M ratio. Although the S H/M ratio underwent an 7% decrease from D1 to D3, the differences were not significant. Individually, however, six of the nine subjects decreased their H/M ratios from 12-42% across days. The results may reflect the inception of longer-term adaptations of the segmental stretch reflex system. PMID- 11264906 TI - Acoustic startle eyeblink response after acute exercise. AB - The acoustic startle eyeblink response (ASER) is a useful probe for investigating central nervous system activity associated with emotional responses to aversive and appetitive stimuli. Though the ASER is sensitive to change in emotional arousal, the effect of acute physical exertion on ASER has not been reported. We examined changes in ASER amplitude and latency in 26 healthy young men (24+/-5 yr) after 20 min of cycling at light and hard intensities (40% and 75% VO2peak) and after 20 min of quiet rest. Mixed model ANCOVA, controlling precondition scores, indicated no effects for ASER amplitude or latency in either sedentary or active participants (p>.10). Our findings indicate that possible effects of acute exercise on potentiated startle or ASER responses elicited by positive or negative foreground stimuli should not be expected to be confounded by an altered baseline acoustic startle eyeblink response when measured in healthy young men. PMID- 11264907 TI - HIV adolescents show improved immune function following massage therapy. AB - HIV+adolescents (M CD4=466 mm3) recruited from a large urban university hospital's outpatient clinic were randomly assigned to receive massage therapy (n=12) or progressive muscle relaxation (n=12) two-times per week for 12 weeks. To assess treatment effects, participants were assessed for depression, anxiety and immune changes before and after treatment the 12 weeks treatment period. Adolescents who received massage therapy versus those who experienced relaxation therapy reported feeling less anxious and they were less depressed, and showed enhanced immune function by the end of the 12 week study. Immune changes included increased Natural Killer cell number (CD56) and CD56+CD3-. In addition, the HIV disease progression markers CD4/CD8 ratio and CD4 number showed an increase for the massage therapy group only. PMID- 11264908 TI - Functional connectivity and working memory in schizophrenia: an EEG study. AB - A leading hypothesis suggests that schizophrenic patients suffer from a disconnection syndrome. A failure in functional connectivity curtails the cortical integration and network activation needed to perform working memory tasks. Simulations with neural network models also indicate that connectivity is crucial for simulation of working memory asks. Multichannel EEG correlation coefficient estimations are considered as a reliable measurement of connectivity patterns among cortical regions. In this study EEG samples are obtained selectively at the delay epochs of a delayed response working memory task. Results of correlation-coefficient estimations indicate a lack of statistically significant changes between non-task and task conditions in frontal, certain parietal, temporal and central channels. These findings propose that schizophrenics probably "fail" to activate the neural networks of the fronto temporal regions. These are the networks involved in computation of the working memory task. Interestingly also good performers schizophrenics failed to activate these networks suggesting that the connectivity function is more relevant to the disorder than to task performance. If distinct deficits in cortical network activations would correlate with mental disorders it would be relevant to diagnosis and treatment of psychiatric disorders. PMID- 11264909 TI - Behavioral and morphological consequences of primary astrocytes transplanted into the rat cortex immediately after nucleus basalis ibotenic lesion. AB - Adult male rats received transplants of dissociated 30-day old cultured cortical astrocytes into the ipsilateral frontal and parietal cortex immediately after unilateral ibotenic acid lesion of the NBM or after sham injury. We hypothesized that transplants of astrocytes into the acetylcholine-deprived cortex might provide trophic support to terminals arising from damaged NBM neurons. Twenty four hours after transplantation and every other day for 11 days post surgery, the animals were tested for locomotion and habituation in an open field. NBM lesion reduced vertical movements only as compared to no lesion and no transplant counterparts. Nine days after surgery rats with NBM lesion and astrocyte transplants into the cortex were as impaired in the acquisition of a passive avoidance (PA) task as untreated counterparts. Animals with no lesions and transplants into the cortex also had significant PA acquisition deficits. All rats with ibotenic lesion were significantly impaired on PA retention as compared to rats with no lesions. Astrocyte-transplants survived up to 2 months after cortical implantation but these transplants produced severe laminar disruption and gliosis. This effect was greater in rats with NBM lesion than in intact animals with transplants into the cortex. These data show that astrocyte transplants do not promote functional recovery after NBM lesion and suggest an immune rejection of the astrocyte transplants by the host brain. PMID- 11264910 TI - Effects of testosterone and clomiphene on spectral EEG and visual evoked response in a young man with posttraumatic epilepsy. AB - The effects of testosterone and clomiphene on epilepsy was studied in a young man with posttraumatic seizures. In the control period, digital EEG and visual evoked potentials (VEPs) were recorded under carbamazapine therapy. After testosterone (T), seizures lessened and almost disappeared; the theta, delta, alpha, and beta powers decreased; VEPs increased. After clomiphene, VEPs considerably increased in size; clinical picture slightly improved; EEG power spectrum remained unchanged. It was suggested that T may be beneficial for epilepsy treatment by suppressing the EEG synchronization (slow wave activity) and attenuating the entropy state of the epileptic brain. PMID- 11264911 TI - Blockade of alpha2-adrenergic receptors markedly potentiates glutamate-evoked activity of locus coeruleus neurons. AB - The locus coeruleus (LC) is a small pontine nucleus with widely distributed efferents that supply the majority of norepinephrine in brain. Evidence from our laboratory has implicated alpha2-adrenergic receptors on LC neurons in regulating the magnitude of the bursting response of LC neurons to both sensory stimulation and systemically administered drugs. Sub-stimulation of alpha2-adrenergic receptors in the LC and the resulting hyper-responsiveness of LC neurons is thought to at least partially mediate depression in an animal model. The present study extends previous work in our laboratory by demonstrating that blockade of alpha2-adrenergic receptors also markedly augments the bursting response of LC neurons to locally-via microiontophoresis-applied glutamate. PMID- 11264912 TI - Neurological and emotional sequelae of exposure to ethylene oxide. AB - Subjects were 66 traumatic brain-injured (TBI) versus 22 neurotoxic-injured individuals (due to ethylene oxide - ETO). Among significant findings were higher cognitive functioning for the TBI group for FSIQ, VIQ and PIQ; elevated scores on MMPI scales 1, 2, 3 and 8 for both groups, and elevated anxiety scores on the MAACL for the ETO group relative to the TBI group. Results indicate a more severe impact of ETO exposure in the areas of intellectual functioning and anxiety. PMID- 11264913 TI - Sex differences in EEG correlates of a self-reported measure of hemisphere preference. AB - The present study explored sex differences in hemisphere preference (HP) assessed by the Preference Test (PT) and its EEG correlates. PT is a paper-and-pencil test designed to measure the extent of individuals relaying on right-hemisphere or left-hemisphere cognition. "Study 1" verified sex differences in HP only among subjects with statistical significant differences between mean and/or median of right- respect to left-HP abilities scores. No sex differences in the frequency of right- and left-HP among the 16 to 17% of a cohort of 1,057 young subjects (473 men and 584 women) with a significant HP were registered. Minimal sex differences were observed in relation to the magnitude of PT score. "Study 2" verified on 34 healthy adults (22 women and 12 men) the sex differences in correlation between PT-defined HP and a more direct index of hemisphere activation such as the alpha power asymmetry derived from resting EEG (vertex reference). In both sexes, PT scores were found to be related with frontal (0.54 women, 0.69 men), but not parietal, alpha power asymmetries. Higher positive correlation of PT score with frontal alpha ratios was reported in both sexes when the median difference of right- respect to left-HP abilities scores in PT score calculation was used. Overall, this study confirmed and extended the evidence on the association between PT-defined HP and frontal, but not parietal, alpha power asymmetries. No significant sex differences were registered in this pattern. PMID- 11264914 TI - On the duration of spatial fluency measures. AB - Various measures of spatial fluency have been developed and have been shown to be sensitive to right frontal lobe dysfunction. Patients with diffuse cerebral injuries (traumatic brain injury) also show impaired performance. Administration times for these tests range from three to five minutes. It is well known that longer tests provide more reliable data. In the present study, the base rates of unique designs and perseverative errors on the Five-Point Test were examined minute-by-minute for ten minutes, in a sample of healthy young adults (n=80). Contrasts between each minute showed significant decreases in number of unique designs to the ninth minute (significant at p<.001). Contrasts between each minute revealed significant increases (significant at p<.00l) in percentage of perseverative errors to the eighth minute. Data demonstrating the progressive decrease of unique designs and progressive increase of perseverative errors as a function of time have important implications for clinical practice. Optimal administration time is considered in the context of clinicians' objectives. PMID- 11264915 TI - Lower back pain is reduced and range of motion increased after massage therapy. AB - STUDY DESIGN: A randomized between-groups design evaluated massage therapy versus relaxation for chronic low back pain. OBJECTIVES: Treatment effects were evaluated for reducing pain, depression, anxiety and stress hormones, and sleeplessness and for improving trunk range of motion associated with chronic low back pain. SUMMARY OF BACKGROUND DATA: Twenty-four adults (M age=39.6 years) with low back pain of nociceptive origin with a duration of at least 6 months participated in the study. The groups did not differ on age, socioeconomic status, ethnicity or gender. METHODS: Twenty-four adults (12 women) with lower back pain were randomly assigned to a massage therapy or a progressive muscle relaxation group. Sessions were 30 minutes long twice a week for five weeks. On the first and last day of the 5-week study participants completed questionnaires, provided a urine sample and were assessed for range of motion. RESULTS: By the end of the study, the massage therapy group, as compared to the relaxation group, reported experiencing less pain, depression, anxiety and improved sleep. They also showed improved trunk and pain flexion performance, and their serotonin and dopamine levels were higher. CONCLUSIONS: Massage therapy is effective in reducing pain, stress hormones and symptoms associated with chronic low back pain. PRECIS: Adults (M age=39.6 years) with low back pain with a duration of at least 6 months received two 30-min massage or relaxation therapy sessions per week for 5 weeks. Participants receiving massage therapy reported experiencing less pain, depression, anxiety and their sleep had improved. They also showed improved trunk and pain flexion performance, and their serotonin and dopamine levels were higher. PMID- 11264916 TI - The clinical utility of force/displacement analysis of muscle testing in applied kinesiology. AB - Manual muscle testing procedures are the subject of a force and displacement analysis. Equipment was fabricated, tested, and employed to gather force, displacement, and time data for examining muscle test parameters as practiced by applied kinesiology (A.K.) clinicians. Simple mathematical procedures are used to process the data in an effort to find potential patterns of force and displacement which would correspond to the testing of strong and weak muscles on healthy subjects. Particular attention is paid to the leading edge of the force pulses, as most clinicians report that they derive most of their assessment from the initial thrust imparted on the patient's limb. An analysis of the simple linear regression of the slope of the leading edge of a force pulse reveals that a high dx/dF is indicative of a weak muscle test result (as perceived by the tester), and a low dx/dF is indicative of a strong muscle test. Thresholds for dx/dF are determined to discriminate between inhibited and facilitated muscle test results. The data lay the groundwork for future studies that examine the objectivity of A.K. muscle testing. PMID- 11264917 TI - Continuing cognitive impairment after isolated transient global amnesia. AB - Fourteen patients were investigated 3-4 days after end of their transient global amnesia (TGA) with a number of neuropsychological tests. Their performance was compared with that of a control group, matched for age, education, and profession. It was found that in spite of the common definition of TGA, impairments in both verbal and non-verbal long term memory and verbal fluency persisted and were in fact impaired to such a degree that it seemed unlikely that full recovery would have occurred within the next few days. We propose a major role of stress in the etiology and the recovery process of TGA and consider it likely that stress hormones are of major influence both in the triggering of TGA and the subsequent continuation of cognitive impairments. PMID- 11264918 TI - Synchronized feeding as a "conditioned stimulus" for overt seizures in chronically (limbic) epileptic rats: a model for "psychogenic seizures" with complex partial epilepsy. AB - Chronic limbic epilepsy was induced in male albino rats by a single systemic injection of lithium (3 mEq/kg) and pilocarpine (30 mg/kg). During the subsequent months the numbers of spontaneous, paroxysmal stereotyped episodes (analogous to Racine stages 4 and 5) were monitored. The numbers of these "overt seizures" increased within 10 min of the daily presentation of a food stimulus even though food was available ad libitum. The majority of the paroxysmal, stereotyped behaviours occurred within 1 min of the stimulus presentation; they were attenuated by oral prednisolone. Three rats displayed evidence of "conditioned seizures" to specific stimuli. The results suggest that the display of these behaviours can be synchronized and learned in contexts that are associated with the release of CRF (corticotrophin releasing factor) and may involve the disinhibited activity within the central amygdaloid nucleus of these rats. Implications for the occurrence of psychogenic seizures in patients with complex partial (limbic) epilepsy are discussed. PMID- 11264919 TI - Influence of photoperiod, laboratory caging and aging on plasma lipid response to an atherogenic diet among F1B hamsters. AB - The effects and interactions of photoperiod, animal caging, aging and diet on plasma lipid levels in male F1B hamsters were examined in the current study. Sixteen young and sixteen old animals were housed one or four per cage. Eight young animals from each housing group were placed in an animal room with either 12/12 h (PT-12) or 10/14 h (PT-10) light/dark cycle while the sixteen old animals were maintained under a PT-12 light cycle. Plasma cholesterol and triglyceride concentrations were determined in all animals after a 2-week period of acclimation on chow diet and following 4-week intervention on atherogenic diet. Baseline total cholesterol (TC) levels were 131+/-25 mg/dl and 142+/-39 mg/dl for young and old animals, respectively, while baseline triglyceride (Tg) levels were 202+/-48 mg/dl and 160+/-37 mg/dl respectively for the same animals. Following 4 weeks on an atherogenic diet, single-caged PT 12 animals had elevated but significantly lower TC levels than group-caged animals (161+/-30 mg/dl and 240 +/ 58 mg/dl, respectively) while single and group housed PT10 animals had TC levels of 296+/-75 mg/dl and 351+/-124 mg/dl, respectively. Similarly, plasma Tg levels rose to 330+/-100 mg/dl and 486+/-200 mg/dl in single and group housed PT12 animals (respectively) and to 668+/-270 mg/dl and 545+/-199 mg/dl in single and group housed PT10 animals (respectively). No significant changes related to atherogenic diet were observed in plasma TC or Tg levels in the older animals. Although caging conditions influence the cholesterol and triglyceride response to the atherogenic diet (p<.05), light cycle photoperiod seems to exert a greater effect (p<.005). In conclusion, photoperiod length dramatically affects diet induced plasma lipid concentrations in young male F1B hamsters, and thus needs be considered in experimental designs of animal-housed lipid research. PMID- 11264920 TI - Experimental production of illusory (false) memories in reconstructions of narratives: effect size and potential mediation by right hemispheric stimulation from complex, weak magnetic fields. AB - This experiment was designed to discern the proportion of false, inferential and verbatim memories that would be included in the reconstruction, one week later, of a 5 min narrative containing ambiguous but emotional content about a little boy. After 48 subjects were administered Spiegel's Hypnosis Induction Profile, they listened to the narrative, were exposed to one of four applications of transcerebral weak, complex magnetic fields for 30 min and then given either an accurate or inaccurate short summary of the story. One week later the group who received the erroneous summary reported more false memories about the original story than did the reference group; this treatment accommodated about 40% of the variance in numbers of false memories. Only an indicator of electrical lability within the temporal lobes (but not hypnotizability) was strongly associated with the numbers of inferential memories but not the numbers of false memories. The group that received transcerebral stimulation over the right hemisphere by a complex magnetic field and the erroneous summary reported three times the numbers of false memories compared to the other groups. Whereas verbatim memories showed a strong primacy effect inferential memories exhibited a strong recency effect (eta(2) =.66). PMID- 11264921 TI - Event-related potentials for category-specific information during passive viewing of faces and objects. AB - Normal subjects passively viewed an upright or inverted face or objects during recording of event-related potentials. Face inversion augmented N170 amplitude and latency in the temporal region, but only the latency in the parietal region. The same manipulation slowed down the onset of the P220 and caused disappearance of the N300, whereas none of these effects was seen after object inversion. Item specific processing of objects was observed, namely disappearance of the N190 and the appearance of a P170 wave in the left posterior hemisphere to one object but not the other. These results are concordant with the hypothesis of category specific processing during the recognition of faces and objects. PMID- 11264922 TI - Electrophysiological correlates of the visual after effect by means of visual evoked potentials. AB - An attempt was made to determine whether changes of electrical activity could be seen in the posterior cortex during an after image of high frequency luminance gratings. Steady state visual evoked potentials were recorded (midoccipital, right and left temporo-occipital sites) immediately after a period of visual adaptation (15 min) to the stimulus, while the subjects experienced the after image. During this illusion, frequencies of the fast Fourier transform spectra linked to the stimulation differed from the noise and were larger at temporo occipital sites than at the midoccipital one. In view of these results, the hypothesis that the after effect represents a short term storage of the temporal characteristics of the stimulus is evoked. PMID- 11264923 TI - Neuropsychological clusters within intelligence levels for learning disabled children. AB - The present study assessed 1142 learning disabled children with the Halstead Reitan Neuropsychological Battery and the Wechsler Intelligence Scale for Children-Revised. Subjects were divided into four groups based upon Full Scale IQ (i.e., 70-79; 80-89; 90-99 and 100-110). Subsequent clustering of the test data within each group suggested that while the students in the 70-79 IQ range were represented by a single Impaired Cluster, each of the other IQ groups had both an Impaired Cluster and a Non-Impaired Cluster. PMID- 11264924 TI - Neuropsychological predictors of the attainment of treatment objectives in substance abuse patients. AB - This study examined the contribution of neuropsychological functioning to the attainment of treatment objectives in substance abuse patients. Subjects were 85 patients enrolled in comprehensive, inpatient and outpatient substance abuse treatment at a VA Medical Center. Most subjects were diagnosed with Alcohol Dependence or Abuse, and nearly half were seeking treatment for Cocaine Dependence or Abuse. After acute detoxification, but before beginning individualized treatment, subjects were administered a neuropsychological screening battery to assess cognitive functioning and affective status. They then attended a variety of daily group therapies. Each therapy group had its own set of specific treatment objectives; on each treatment day, group therapists rated each patient's attainment of the specific objectives for their group. Groups included Assertiveness Training (Levels I and II), Stress Management (Levels I and II), Social Skills Training, Job Skills, Relapse Prevention (Levels I and II), Leisure Planning, Leisure Skills, Occupational Therapy, and 12-Step Study. Stepwise multiple regression indicated that the best predictors of overall objective attainment were better attention (WMS-R Digits Backwards) and less depressive symptomatology (Beck Depression Inventory). These results suggest that attention and mood have a modest yet significant impact on the success of treatment interventions for substance abuse patients. Thus, evaluation of cognitive as well as affective factors in substance abuse patients might be helpful in designing and implementing specialized interventions to maximize the likelihood of treatment success. PMID- 11264925 TI - The author's guide to controlling the photograph. PMID- 11264926 TI - Editorial. 1951. PMID- 11264927 TI - The developments in the occlusal patterns of artificial teeth. 1951. PMID- 11264928 TI - Procera AllCeram laminates: a clinical report. AB - The use of Procera AllCeram laminates for patients with discolored teeth has been described. In our experience, these laminates are simple to use and provide possibilities for excellent esthetics. PMID- 11264929 TI - A vibratory stimulation-based inhibition system for nocturnal bruxism: a clinical report. AB - For the single subject tested to date, the bruxism-contingent vibratory-feedback system for occlusal appliances effectively inhibited bruxism without inducing substantial sleep disturbance. Whether the reduction in bruxism would continue if the device no longer provided feedback and whether the force levels applied are optimal to induce suppression remain to be determined. PMID- 11264930 TI - Determining the accuracy of articulator interchangeability and hinge axis reproducibility. AB - STATEMENT OF PROBLEM: It has been reported that articulators are interchangeable, which means that a clinician should be able to use one articulator and send casts to a dental laboratory with the assurance that the casts will be remounted with positional accuracy on a similar articulator. PURPOSE: The purpose of the study was to determine whether mounted casts could be transferred from 1 articulator to another with positional accuracy and whether the hinge axis was reproducible in each of the articulators tested. MATERIAL AND METHODS: This study compared left and right second premolars and first molar occlusal contact areas with respective contact areas of like-mounted casts. Five calibrated Whip Mix 3040, 5 calibrated Girrbach Artex AL, and 5 calibrated KaVo Protar articulators were tested. Impact resistant resin casts mounted in occlusion on 1 articulator were transferred to 4 like articulators. Each of the 5 articulators of each brand was opened and closed 10 times. Ten vinyl polysiloxane right and left posterior interocclusal records of the occluded casts were made for each articulator. The use of a computerized image analysis program provided quantitative measurements of light transmitted through the occlusal records. A Kruskal-Wallis test was used for each of the 4 independent variables of the study (molar differences, premolar differences, left differences, right differences). By using a calibrated grid, a numerical assessment of positional changes was made in millimeters. RESULTS: None of the articulator systems was found to be exact, and no single articulator was an exact duplicate of another (P<.01). The Whip Mix articulator showed greater deviation both in hinge axis repeatability and in articulator interchangeability than the KaVo. The Artex articulator provided the most consistent hinge axis repeatability and interchangeability of the 3 brands of articulators. CONCLUSION: The Artex brand reproduced dental cast positions more consistently than the other articulators tested. PMID- 11264931 TI - Procedure for occlusal refinement of mounted definitive casts to reduce clinical time required for adjustment of occlusion. AB - The dental technician works with rigid casts and dies that do not tip or depress. The dentist must use fixed prostheses that have been fabricated in a laboratory setting in a clinical environment that is significantly different. Periodontal ligaments permit individual teeth to move within their boney housing. Mandibles tip and bend, but articulators cannot simulate all this activity. A relatively simple technique for adjusting inclines on the mounted definitive and opposing casts can compensate for many of the differences between rigid casts and subtle clinical tooth movement. This procedure can reduce required chairtime for adjusting the occlusion on crowns and fixed partial dentures. PMID- 11264932 TI - Masticatory muscle activity assessment and reliability of a portable electromyographic instrument. AB - STATEMENT OF PROBLEM: Masticatory muscle hyperactivity is thought to produce muscle pain and tension headaches and can cause excessive wear or breakage of restorative dental materials used in the treatment of prosthodontic patients. The quantification and identification of this type of activity is an important consideration in the preoperative diagnosis and treatment planning phase of prosthodontic care. PURPOSE: This study investigated the quantification process in complete denture/overdenture patients with natural mandibular tooth abutments and explored the reliability of instrumentation used to assess this parafunctional activity. MATERIAL AND METHODS: The nocturnal EMG activity in asymptomatic complete denture/overdenture subjects was assessed with and without prostheses worn during sleep. Because of the large variance within and between subjects, the investigators evaluated the reliability of the 3 instruments used to test nocturnal EMG activity in the sample. RESULTS: Electromyographic activity data of denture/overdenture subjects revealed no differences between prostheses worn versus not worn during sleep but demonstrated a very large variance factor. Further investigation of the instrumentation demonstrated a consistent in vitro as well as in vivo reliability in controlled laboratory studies. CONCLUSION: The portable EMG instrumentation used in this study revealed a large, uncontrollable variance factor within and between subjects that greatly complicated the diagnosis of parafunctional activity in prosthodontic patients. PMID- 11264933 TI - The CICERO system for CAD/CAM fabrication of full-ceramic crowns. AB - The CICERO method of crown fabrication consists of optically digitizing a gypsum die, designing the crown layer buildup, and subsequently pressing, sintering, and milling consecutive layers of a shaded high-strength alumina-based core material, a layer of dentin porcelain, and a final layer of incisal porcelain. Final finishing is performed in the dental laboratory. The CICERO method allows efficient production of all-ceramic restorations without compromising esthetics or function. This article reviews the process involved in the fabrication of a CICERO crown. PMID- 11264935 TI - Histomorphometric comparison of implant anchorage for two types of dental implants after 3 and 6 months' healing in baboon jaws. AB - STATEMENT OF PROBLEM: A complete understanding of dental implant prognosis requires better knowledge of the bone anatomy after implant healing. Such baseline data are necessary to compare against load-induced changes in anatomy. PURPOSE: The purpose of this article is to describe and compare measures of implant support (percentage [%] integration and percentage [%] bone area) for various implants in baboon jaws after healing times of 3 and 6 months. MATERIAL AND METHODS: Commercially pure titanium (cpTi) and titanium alloy (Ti-alloy) screw-shaped implants were placed in the posterior jaws of 9 female baboons after 2 months of postextraction healing. Specimens were harvested after 3 months (5 baboons: 8 cpTi, 7 Ti-alloy) and after 6 months (4 baboons: 8 cpTi, 8 Ti-alloy). Each implant provided 6 polished horizontal sections for data collection, which was accomplished from digitized images with the IMAGE analysis system (reliability at 1.6%). Three- and six-month data for each parameter were compared with the use of ANOVA (P<.01). RESULTS: The results revealed a significant increase in the % integration (cpTi 39.1 to 56.2; Ti-alloy 40.0 to 55.2) and the % bone area (cpTi 38.8 to 47.9; Ti-alloy 38.9 to 49.2) from 3 to 6 months for both implants. This significant increase was also true for comparisons by jaw for each implant material (P<.01 for overall and by jaw comparisons). CONCLUSION: A time-dependent increase in jawbone anchorage was measured in this nonhuman primate population, and it was shown that the 6-month maxillary data were comparable to the 3-month mandibular data. These results lend support to the clinical strategy of waiting longer to load implants in the maxilla. PMID- 11264934 TI - Examination of the implant-abutment interface after fatigue testing. AB - PURPOSE: This study examined potential differences in detorque values of abutment screws after fatigue testing when the dimensions between external implant hexagon and internal abutment hexagon were altered or the implant external hexagonal shape was eliminated. MATERIAL AND METHODS: Three subsets (N = 10) of NobelBiocare implants were assessed: (1) standard external hexagon (R), (2) modified hexagon (M), and (3) circular (C) platform geometry. Thirty Procera machined abutments with 25-degree angulated loading platforms were manufactured. Abutments were retained with gold Unigrip abutment screws tightened to 32 N/cm with an electronic torque controller. Vertical scribes across the implant abutment interface allowed longitudinal displacement evaluation. A carousel-type fatigue testing device delivered dynamic loading forces between 20 and 200 N for 5,000,000 cycles, or the approximate equivalent of 5 years in vivo mastication, through a piston to the abutment platform. Macroscopic and radiographic examination of the implant/abutment specimens was performed. The abutment screws were removed and the detorque values recorded. Bearing surfaces were examined microscopically. RESULTS: No abutment looseness or longitudinal displacements at the implant-abutment interface were noted. Radiographic examination demonstrated no indication of screw bending or displacement. The mean detorque values for R, M, and C were 14.40 +/- 1.84 N/cm, 14.70 +/- 1.89 N/cm, and 16.40 +/- 2.17 N/cm, respectively. The analysis of variance demonstrated significant differences between only designs R and C (P=.031). CONCLUSION: Increasing the vertical height, or degree of fit tolerance, between the implant external hexagon and the abutment internal hexagon or completely eliminating the implant external hexagon did not produce a significant effect on the detorque values of the abutment screws after 5,000,000 cycles in fatigue testing, or the equivalent of 5 years' of mastication for the implant/abutment specimens evaluated. PMID- 11264936 TI - Practical application of polyurethane and Velcro in maxillofacial prosthetics. AB - A procedure is described for the fabrication of an extraoral prosthesis with an acrylic resin substructure that retains a magnet sealed from the environment by a polyurethane liner. Velcro is used to enhance the bond of the acrylic substructure to the silicone prosthesis. This procedure results in improved retention of the acrylic resin substructure and protection of the magnet with an encapsulating polyurethane liner. PMID- 11264937 TI - Microleakage of endodontically treated teeth restored with fiber posts and composite cores after cyclic loading: a confocal microscopic study. AB - STATEMENT OF PROBLEM: The effectiveness of the seal obtained with carbon fiber posts and composite cores is still unclear. Both 3-step dental adhesives and self etching adhesive primers have been suggested as adhesive systems. PURPOSE: This confocal microscopic study evaluated the microleakage of teeth endodontically treated and restored with fiber posts and composites with 3 adhesive systems. MATERIAL AND METHODS: A total of 72 human mandibular premolars were endodontically treated and divided into 6 groups of 12 teeth each. The first 3 groups were treated with an endodontic sealer containing zinc oxide-eugenol (ZOE) and restored with temporary filling materials containing ZOE. The last 3 groups were treated with ZOE-free materials. Post spaces were prepared in the root canals. The first group treated with ZOE-based materials was restored with fiber posts cemented with zinc phosphate cement and composite cores without adhesive. The other 2 groups of ZOE-treated teeth were restored with fiber posts cemented with All Bond 2 and Panavia 21 dental adhesives, respectively. The last 3 groups were restored with fiber posts cemented with All Bond 2, Panavia 21, and Panavia Fluoro cement, respectively. The teeth were loaded intermittently at 2 cycles per second in a moist environment and, after 300,000 cycles, immersed in a solution of Rhodamine B dye for 48 hours. A confocal microscope was used to observe the teeth. The ratio between the length of the interfaces observed and the length of the dye penetration was evaluated. Two teeth from each group acted as controls and were not subjected to dynamic loads. RESULTS: All resin cement groups leaked significantly less than the group cemented with zinc phosphate cement. No statistically significant difference was found between the microleakage of teeth treated with ZOE-based and non-ZOE-based materials. Teeth restored with All Bond 2 dental adhesive leaked significantly less than those restored with Panavia cement. CONCLUSION: The 3-step dental adhesive (All Bond 2) resulted in a better marginal seal than that obtained with the self-etching primers (Panavia 21 and Panavia F). The use of endodontic sealers and temporary filling materials containing ZOE had no detrimental effect on the marginal seal of carbon fiber post/composite resin core restorations. PMID- 11264938 TI - In vitro microleakage of luting cements and crown foundation material. AB - STATEMENT OF PROBLEM: Microleakage is a concern for the long-term prognosis of a cemented crown and foundation. PURPOSE: The aims of this investigation were, first, to evaluate microleakage of zinc phosphate cement and resin-reinforced glass ionomer cement under ideal (dry) versus contaminated (wet) conditions, and second, to compare 3 foundations under both ideal and contaminated conditions. MATERIAL AND METHODS: One hundred forty extracted molar teeth were cleaned and mounted. Tooth preparations for complete veneer cast crowns were completed with a chamfer finish line. A mesial surface class II cavity preparation 4 mm wide buccolingually and 2 mm deep was made in each tooth. Seven restorative groups were formed: amalgam/cavity varnish, amalgam/dentinal bonding agent, and composite/dentinal bonding agent, each with dry and contaminated groups, and a seventh group of class II cavity preparations without foundations. Finish lines for crown margins were refined 1.5 mm gingival to the restoration. Artificial crowns were cast in type III gold. Treatment groups were divided into 4 cement groups: dry and contaminated zinc phosphate cement and dry and contaminated resin reinforced glass ionomer cement. The specimens were thermocycled and immersed in erythrosine B solution for 24 hours. Subsequently, they were rinsed, and their coronal portions were embedded in clear resin. Teeth were sectioned mesiodistally, and standard photomicrographs were made. The microleakage of each restoration and crown was measured. RESULTS: The least foundation microleakage was recorded for amalgam/dentinal bonding agents (ideal group) and composite/dentinal bonding agents (ideal group). The most microleakage was observed within the group without a foundation. In cement groups, the control and experiment sides were evaluated separately but displayed the same order of finding. The least leakage was recorded with resin-reinforced glass ionomer cement (ideal group); the most microleakage was noted with zinc phosphate cement (ideal group). An interaction was demonstrated on the experimental side between cements and the foundations (P=.0001). CONCLUSION: Within the experimental conditions of this study, less microleakage was recorded with resin-reinforced glass ionomer cement (ideal or contaminated) than with zinc phosphate cement (ideal or contaminated). There also was less microleakage evident with a foundation of silver amalgam or composite when a dentinal bonding agent was used under ideal conditions. PMID- 11264940 TI - Behavioral approaches to reduce hypersensitive gag response. PMID- 11264939 TI - Castability and resistance of ceramometal bonding in Ni-Cr and Ni-Cr-Be alloys. AB - STATEMENT OF PROBLEM: Because of its pathogenic potential, the importance of the use of beryllium with Ni-Cr alloys must be determined. PURPOSE: This study compared fundamental properties for the clinical use of Ni-Cr alloys, determining the advantage of the addition of beryllium, despite the involved risks. MATERIAL AND METHODS: Two Be-free commercial alloys (Vera Bond II and Wiron 99) and 1 Be free experimental alloy (E3) with Nb and/or Mo in their formulations and 1 experimental Ni-Cr alloy (E4) with 1.1% Be were submitted to ceramometal bonding resistance, castability, and hardness tests. RESULTS: Analysis of variance showed significant differences (P=.05) between Vera Bond II and Wiron 99 (Be-free) and the E4 alloy for the castability test. Vera Bond II, Wiron 99, and E4 presented higher castability values than E3. There were no statistically significant differences for the ceramometal bonding resistance test. The Kruskal-Wallis test showed significant differences (P=.01) among the alloys with Rockwell 30 T values for the hardness test. CONCLUSION: Within the limitations of this study, the presence of Be in Ni-Cr alloys was not necessary to guarantee the castability and the ceramometal bond resistance of the alloys tested. PMID- 11264941 TI - Simplified procedure for preparing removable dies. PMID- 11264942 TI - A modified rotary instrument for tightening the lingual locking screw of an implant-supported prosthesis. PMID- 11264944 TI - A method for establishing primary cultures of human gastric epithelial cells. AB - Long-term culture of human gastric epithelial cells has been difficult, and at present no normal human gastric epithelial cell lines are readily available. As part of our experiments to study pathogenesis of H. pylori, a bacterium that infects the stomach, we developed methods to culture normal human gastric epithelial cells. Primary cultures of human gastric epithelial cells can be established from gastric biopsies taken at upper G.I. endoscopy. Enzymatically isolated gastric epithelial-like cells are present in tight colonies on culture dishes within 24 hours of placing the cells in culture. Cells isolated stain positively for cytokeratin and produce neutral mucins, indicating that they are mucin secreting epithelial cells, consistent with gastric epithelial cells. Epithelial cells can be maintained up to 4 weeks in culture with evidence of DNA synthesis up through the first week of culture. PMID- 11264945 TI - Use of a micromanipulator for high-efficiency cloning of cells co-expressing fluorescent proteins. AB - The inclusion of the gene encoding the Green Fluorescent Protein (GFP) or its derivatives into dicistronic transfer vectors is a useful method to visually identify cells that have incorporated a specific gene of interest. By combining this approach with the use of a micromanipulator, we have developed a protocol for the one-step isolation of cells expressing a specific transgene from a pool of transfected cells. Target fluorescent cells could be identified and isolated even when they occurred at frequencies as low as 1/100,000. The use of Leibowitz L-15 serum- free medium and serum-coated non-charged petri dishes, along with minimal light exposure yielded maximal cell viability and high cloning efficiency (approximately 40%, on average) for a large number of cell lines, both adherent and suspension. Several variations of the basic method are presented, as well as guidelines for the choice of hardware components to implement our cloning workstation. PMID- 11264946 TI - Culturing endothelial cells of microvascular origin. AB - There is considerable divergence of opinion on the best methods for the isolation and in vitro culture of microvascular endothelium. Reports have either only described the isolation without mentioning culture conditions or have not fully defined the cell population being cultured. Even at the close of the 20th Century, the isolation and in vitro culture of endothelial cells of microvasculature origin still proves to be technically difficult and many questions remain. These questions need to be addressed by improvements to current methods of isolation and culture of Endothelial cells. A number of more 'high tech' approaches to this isolation are being explored currently. Use of a more definitive panel of antibodies for immunocytochemical characterization, should enable a more confident characterization of the endothelial cell preparations cultured in vitro. Cell adhesion molecules such as ELAM and VCAM can be used to assist in determining cell population purity. PMID- 11264947 TI - Growth of rainbow trout hemopoietic cells in methylcellulose and methods of monitoring their proliferative response in this matrix. AB - A technique for the clonal culture of rainbow trout leukocytes in a methylcellulose matrix can be used to identify growth factors and other substances affecting cell proliferation and development in fish. Methylcellulose supports colony formation by rainbow trout leukocytes isolated from the major hemopoietic organ, the pronephros. The addition of rainbow trout serum dramatically increased the number of colonies formed, scored by counting colonies. As an alternative measure of cell proliferation, 3H-thymidine incorporation by cells can be easily measured in methylcellulose cultures. This method requires only small amounts of test substances, is rapid, and is superior for assessing the growth-stimulating ability of some substances, such as bacterial lipopolysaccharide, which stimulated growth but not the formation of discreet colonies by rainbow trout cells. PMID- 11264948 TI - Establishment of a pure culture of distinct cell types from bovine placental cotyledon. AB - Methods are described to establish distinct cell cultures from bovine placental cotyledon. The villous tissue of the bovine placental cotyledon is collected and dissociated with 0.125% trypsin. The cells are then cultured in three different media: a serum-free medium, a growth factor supplemented medium, and a medium with 10% serum. A polygonal cell line grew out of the serum-containing medium, a fan-shaped cell line grew out of the serum-free medium, and an epitheloid cell line grew out of the growth factor supplemented medium. These cells maintained their morphology when grown in serum containing medium. The preference of distinct cells for different media in vitro reflects the in vivo physiological regulation of these cells. These distinct cultures re ideal to study the extrinsic and interactive factors in bovine placenta. PMID- 11264949 TI - Procedures for the preparation and culture of 'reconstructed' rainbow trout branchial epithelia. AB - Techniques for the in vitro 'reconstruction' of freshwater rainbow trout branchial epithelia using the primary culture of gill cells on permeable polyethylene terephthalate cell culture filter supports are described. Representing models of the freshwater fish gill, epithelia grown by two separate techniques are composed of branchial pavement cells with or without the inclusion of mitochondria-rich (MR) cells. The generation of epithelia consisting of pavement cells only (via a method called single seeded inserts = SSI) involves an initial period of flask culture during which time MR cells, that appear unable to attach to the culture flask base, are excluded from the general cell populace. Alternately, the generation of a heterogeneous epithelia consisting of both pavement cells and MR cells (via a method called double seeded inserts = DSI) is facilitated by the direct seeding of cells into cell culture filter inserts. Critical to this second procedure is the repeat seeding of filter inserts over a two day period. Repeat seeding appears to allow MR cells to nest amongst the attached cell layer generated by the first day's seeding. The use of cell culture filter supports allows free access to both the apical and basolateral compartment of the epithelium and is ideal for experimental manipulation. Cells are grown under symmetrical conditions (apical media/basolateral media) and epithelium growth is measured as a function of transepithelial resistance (TER). When the epithelia exhibit a plateau in growth they can be subjected to asymmetrical conditions (freshwater apical/media basolateral) in order to assess gill cell function as in vivo. PMID- 11264950 TI - Swimbladder gas gland cells of the European eel cultured in a superfusion system. AB - Swimbladder gas gland cells are polar epithelial cells which release acidic metabolites through the membranes of an extensive basolateral labyrinth, and secret surfactant via exocytosis at apical membranes. We have developed a method to establish primary cell cultures of gas gland cells in order to establish a model system for physiological analysis of gas gland cell function in vitro. Isolated gas gland cells attach to collagen S coated surfaces. Cells cultured in collagen S coated petri dishes were flat and showed no histological polarity. Cells cultured on Anodisc membranes in a superfusion system, in which the apical and basal side of the cells was supplied with a saline solution and with glucose containing DMEM cell culture medium, respectively, showed a clear polarity similar to the in vivo situation. Measurement of lactate release at the apical side and at the basal side revealed that these cells were functionally polar and secreted at least 70% of the lactate at their basal membranes. Gas gland cells could also be cultured in an air/liquid system, in which the apical membrane was exposed to humidified air. Cells cultured under these conditions released lactate only on the basal side and histologically were similar to cells cultured in the superfusion system. PMID- 11264951 TI - Applications of flow cytometry across species. PMID- 11264952 TI - Sperm chromatin structure assay is useful for fertility assessment. AB - The Sperm Chromatin Structure Assay (SCSA) serves as a tool for measuring clinically important properties of sperm nuclear chromatin integrity. The assay utilizes the metachromatic features of Acridine Orange (AO), a DNA probe, and the principles of flow cytometry (FCM). SCSA data are not well correlated with classical sperm quality parameters and have been solidly shown to predict sub/infertility. This assay is ideally suited to human and animal fertility clinics to assess male sperm DNA integrity as related to fertility potential and embryo development as well as effects of reproductive toxicants. A detailed description of the SCSA follows. PMID- 11264953 TI - Standardized flow cytometry gating in veterinary medicine. AB - Gating in flow cytometry is used to select subpopulations of cells for analysis. The technique is critical for subsequent analysis in order to select the population, free of debris and unrelated cells. Accurately quantifying subpopulations in clinical cases is necessary for correct diagnosis. Human lymphocytes are selected by backgating on populations of CD45+high CD14- cells. These reagents are not available widely across species. In veterinary medicine, markers to identify lymphocytes are usually limited to T-lymphocyte, CD4, CD8, and B-lymphocyte surface antigens. A standardized gating technique using a T lymphocyte antibody is described and is applicable across species where limited phenotype markers are available. PMID- 11264954 TI - Clinical applications of flow cytometry and cell immunophenotyping to companion animals (dog and cat). AB - Clinical applications of flow cytometry to certain diseases of the dog and cat are now possible. The utility of such applications for diagnosis, prognosis and follow-up are illustrated here by a number of examples: feline AIDS resulting from FIV infection, Leukocyte Adhesion Deficiency in Irish setters, deep pyoderma in German shepherds, Immune-mediated Thrombocytopenia, canine Systemic Lupus Erythematosus and Leishmaniasis, Leukemia and Lymphoma. PMID- 11264955 TI - Flow cytometric detection of host cell apoptosis induced by bacterial infection. AB - We detail two methods for detection of cell death induced by infection of a human monocytic cell line with invasive Campylobacter bacteria. Staining with a natural ligand for exposed phosphatidylserine residues coupled with propidum iodide discriminated between apoptosis and necrosis. Additionally, cells infected with a bacterial strain expressing green fluorescent protein stained with dye sensitive to mitochondrial membrane potential demonstrated a direct association of bacteria with dying cells. Analyses of cells stained by these methods employing flow cytometry enumerated proportions of cell populations undergoing either apoptosis or necrosis after bacterial infection in vitro. PMID- 11264956 TI - Rapid purification of transfected porcine muscle cells. AB - A detailed methodology is described for fluorescence-activated cell sorting (FACS) of porcine muscle cells that have been transfected to express green fluorescent protein (GFP). Cells are liberated from porcine skeletal muscle and primary cultures are transfected with DNA encoding GFP. Primary cultures are subjected to immunocytochemistry using a primary muscle-specific monoclonal antibody followed by incubation with a phycoerythrin-conjugated second antibody. Transfected myoblasts are sorted from fibroblasts using forward angle light scatter and ninety degree light scatter, phycoerythrin fluorescence, and GFP fluorescence. These procedures allow for isolation of genetically- engineered porcine muscle cells more rapidly than traditional clonal selection procedures. Consequently, FACS provides porcine myoblast populations that retain the majority of their replicative capacity and are not contaminated with non-myogenic cells. PMID- 11264957 TI - Methods for cell cycle analysis and detection of apoptosis of teleost cells. AB - Flow cytometric techniques have not been previously used on a routine basis to study teleost cell growth and development. In the present chapter, flow instrumentation and cell preparation protocols are given in order to provide evaluation criteria characteristic of different phases of the cell cycle. Flow cytometry is used as an analytical and diagnostic tool to measure DNA ploidy as well as to measure alterations in cell cycle profiles characteristic of random DNA fragmentation (necrosis) compared to patterned DNA cleavage (apoptosis). The types of information obtained by flow analysis include the visualization of cell subpopulations with differing DNA content. For each identified nuclei subpopulation, the parameters of population size, fractions of nuclei in each phase of the cell cycle and computation of DNA ratios can be discerned. Data are presented of ex vivo prepared teleost nonspecific cytotoxic cells (NCC) at resting phase compared to NCC undergoing DNA hypoploid changes characteristic of apoptosis. These cells are compared with a teleost tissue cultured cell line maintained under optimum cell growth conditions versus cells undergoing necrotic cellular pathology. Finally, the requirements for optimum flow analysis are described. Techniques including gating strategies, voltage and gain settings, discrimination options and data collection and interpretation are provided. PMID- 11264958 TI - Nonradiometric detection of cytotoxicity of teleost nonspecific cytotoxic cells. AB - The development of a sensitive, rapid and reliable nonradiometric cytotoxicity assay would significantly facilitate studies of teleost nonspecific cytotoxic cells. Such an assay would not require handling and disposal of radionuclides and it would not depend on secondary enzyme or colorimetric determinations. The requirements for this assay would consist of a one-step binding protocol which could detect early target cell membrane lesions and at very low effector:target cell ratios. In this chapter, we have developed a flow cytometry based cytotoxicity assay utilizing the uptake of propidium iodide (PI) into cells containing damaged (i.e. permeabilized) cell membranes. The basis of detection of cellular damage depended on flow discrimination of targets from effector cells by establishing 'scatter' gates from these mixtures. Teleost (catfish) anterior kidney NCC were mixed with human transformed targets (IM-9 and HL-60 cells) at effector:target cell ratios of 1, 5 and 10 and PI uptake was determined at 3 and 14 hours post-incubation. Percent specific uptake (PSU) was calculated by determining total binding capacity (TBC) (i.e. uptake of PI by cold acetone permeabilized target cells) and spontaneous binding capacity (SBC) (i.e. PI uptake by target cells incubated in media w/o effectors). This was represented by the formula PSU = [T - SBC/TBC - SBC] x 100 where T is the PI uptake of targets following addition of effector cells. Using this technique, NCC initiated target cell permeabilization as early as 30 minutes co-incubation (25:1 E:T ratio) and damaged membranes were detected in mixtures containing as few as 1:1 effector:target cell ratios. At 5:1 E:T ratios, greater than 50 PSU was determined following 14 hours co-incubation. Using these criteria, a new and sensitive cytotoxicity assay has been developed to determine NCC activity. PMID- 11264959 TI - Isolation of bovine mammary lymphocytes for fluorescent activated flow cytometry. AB - A detailed methodology is described for the isolation of T lymphocytes from bovine mammary gland secretions for flow cytometry. Mammary gland secretions are collected and centrifuged to separate the leukocytes from the mammary supernatant. The leukocytes are purified using a density gradient and diluted for fluorescent staining. Using tri-color fluorescent staining, the T lymphocytes are identified by monoclonal antibodies and stained with secondary antibodies. Flow cytometry is used to quantify the cellular subpopulations of the T lymphocytes. Proportional and statistical analysis of the flow cytometric data is conducted with computer software, CELLQuest and SAS. PMID- 11264960 TI - Evaluation of endothelial cell migration with a novel in vitro assay system. AB - In this study we introduce a novel in vitro 'oil-drop' assay system for the measurement of endothelial cell (EC) migration, based on the original concept of the Teflon fence assay (Pratt et al., 1984; Am. J. Pathol. 117: 349-354). An aliquot of 15-20,000 human umbilical vein EC (HUVEC) is pipetted through a layer of mineral oil. The cells readily attach, spread and migrate on the surface of a matrix-coated tissue culture dish as a confluent circular monolayer. Migration is measured as the net increase in the total area covered at 24 hours. We have used this system to quantify EC migration on matrices composed of a mixture of type I collagen and either von Willebrand factor (vWF) or fibronectin (FN) in the presence or absence of tumor necrosis factor alpha (TNFalpha). Plating efficiency on both vWF/collagen and FN/collagen, measured by counting cells after attachment and spreading, is about 80%. With this method, migration on vWF/collagen was about 6.4 mm(2) and 5.3 mm(2) for TNFalpha-treated and untreated HUVEC, respectively. HUVEC migration on FN/collagen was slightly greater - 6.4 mm(2) and 6.5 mm(2) with and without TNFalpha treatment, respectively. During the 24 hour time period, HUVEC numbers increased 30-40% on vWF/collagen, and 60-80% on FN/collagen, with increased proliferation observed with TNF-alpha treatment. EC proliferation could be completely inhibited by 2 mM hydroxyurea. This assay system has proven useful in our studies to quantify cell migration and proliferation. PMID- 11264961 TI - Establishment, cryopreservation, and growth of 11 cell lines prepared from a juvenile Hawaiian monk seal, Monachus schauinslandi. AB - Eleven cell lines were prepared from skin, snout, liver, kidney, lung, heart, brain, spleen, thyroid, urinary bladder, and periorbital soft tissue of a juvenile Hawaiian monk seal (Monachus schauinslandi). The cell grew at 37 degrees C in RPMI 1640 medium supplemented with 20% fetal bovine serum. These cell lines have been subcultured 11-27 times since their initiation in May 1997. Growth of the monk seal cells was serum-dependent and plating efficiencies ranged from 4 24%. These monk seal cells grew well in M199, L-15 and MEM commonly used for cultivation of animal and mammalian cells and retained 87% cell viability following storage for 2.5 years in liquid nitrogen. Karyotyping indicated that these monk seal-derived cell lines remained diploid with a chromosome count of 34 at their early passage (passage 9-13). These cell lines were tested for herpesvirus by polymerase chain reaction using degenerate oligonucleotide primers designed from the highly conserved region of herpesviral DNA polymerase gene and no specific detection occurred. These newly established cell lines are currently being used for the investigation of an eye disease occurring in captive monk seal pups in Oahu and will be available for future isolation and study of monk seal viruses. PMID- 11264962 TI - The Lords Report: a challenge to the profession. PMID- 11264963 TI - Use and expenditure on complementary medicine in England: a population based survey. AB - OBJECTIVES: Many claims are made that complementary medicine use is a substantial and growing part of health-care behaviour. Estimates of practitioner visits in the USA and Australia indicate high levels of use and expenditure. No reliable population-based estimates of practitioner use are available for the UK. METHODS: In 1998, a previously piloted postal questionnaire was sent to a geographically stratified, random sample of 5010 adults in England. The questionnaire focuses on practitioner contacts, but also asked about the purchase of over-the-counter remedies. Additional information was requested on socio-demographic characteristics, perceived health, and recent NHS resource use. Information on use included reason for encounter, expenditure, insurance, and location of visit. MAIN OUTCOMES MEASURES: Population estimates (by age group and sex) of lifetime use and use in the past 12 months for acupuncture, chiropractic, homoeopathy, hypnotherapy, medical herbalism, osteopathy. Estimates for two additional therapies (reflexology and aromatherapy), and homoeopathic or herbal remedies purchased over-the-counter. Estimates of annual out-of-pocket expenditure on practitioner visits in 1998. RESULTS: A crude response rate of 60% was achieved (adjusted response rate 59%). Responders were older and more likely to be female than non-responders. Usable responses (n = 2669) were weighted using the age/sex profile of the sample frame. From these adjusted data we estimate that 10.6% (95% CI 9.4 to 11.7) of the adult population of England had visited at least one therapist providing any one of the six more established therapies in the past 12 months (13.6% for use of any of the eight named therapies, 95% CI 12.3 to 14.9). If all eight therapies, and self-care using remedies purchased over the counter are included, the estimated proportion rises to 28.3% (95% CI 26.6 to 30.0) for use in the past 12 months, and 46.6% (95% CI 44.6 to 48.5) for lifetime use. All types of use declined in older age groups, and were more commonly reported by women than men (P < 0.01 for all comparisons). An estimated 22 million visits were made to practitioners of one of the six established therapies in 1998. The NHS provided an estimated 10% of these contacts. The majority of non-NHS visits were financed through direct out-of-pocket expenditure. Annual out-of-pocket expenditure on any of the six more established therapies was estimated at pound 450 million (95% CI 357 to 543). CONCLUSION: This survey has demonstrated substantial use of practitioner-provided complementary therapies in England in 1998. The findings suggest that CAM is making a measurable contribution to first contact primary care. However, we have shown that 90% of this provision is purchased privately. Further research into the cost-effectiveness of different CAM therapies for particular patient groups is now urgently needed to facilitate equal and appropriate access via the NHS. PMID- 11264965 TI - Acupuncture treatment: side effects and complications reported by Swedish physiotherapists. PMID- 11264964 TI - Iscador Qu for chronic hepatitis C: an exploratory study. AB - Five patients with chronic hepatitis C were treated for one year with the herbal immunomodulatory agent Iscador (Weleda, Schwabisch Gmund Germany). Two patients showed 6-20 fold decreases in viral load and normalisation of liver inflammation. The treatment was well tolerated; no serious side effects were observed. The quality of life improved on average. Iscador may have potential as a non toxic therapy for chronic hepatitis C treatment. PMID- 11264966 TI - A study of the use of acupuncture in physiotherapy. AB - OBJECTIVES: This two-phase study was designed to establish the current use of acupuncture within physiotherapy and to determine the opinions of those who received acupuncture therapy. DESIGN: Retrospective study and questionnaire survey. PATIENTS: Patients who attended an outpatient physiotherapy department over a 2-year period (phase 1, retrospective study of clinical records; n = 599). Patients who had received acupuncture treatment from outpatient physiotherapy (phase 2, patient survey; n = 200). MAIN OUTCOME MEASURE: Patient records and questionnaire. RESULTS: The patients who attended for outpatient physiotherapy were categorized into three main groups: low back pain, cervical/thoracic spine problems and soft-tissue injuries of peripheral joints. Acupuncture appeared to be used as a secondary form of treatment for these conditions, where other modalities failed rather than being used for best effect. The response rate to the questionnaire was 78%, of whom 60% stated that they had experienced pain relief following their acupuncture therapy, and 31% were still experiencing pain relief. The majority had achieved sufficient relief to carry out daily activities at home (80%) and at work (57%). Ninety-four per cent of respondents were either 'satisfied' or 'very satisfied' with their treatment. CONCLUSION: Further investigation is required to adequately assess the efficacy of acupuncture as a pain-relieving modality. PMID- 11264968 TI - The House of Lords report on complementary medicine: a summary. AB - The House of Lords Science and Technology Committee reviewed a large amount of oral and written evidence from a wide variety of sources in order to scrutinize CAM. In their published report, they propose that CAM be subdivided into three groups for operational purposes. They note that public satisfaction with CAM is high and use of CAM is increasing. Evidence is required that CAM has an effect above and beyond placebo, and once this is provided the public should have access to it and its potential benefits. Public interest is not protected by the current lack of regulation of CAM, which should be organized in an appropriate way by and for each therapy. Acupuncture and herbal medicine should be subject to statutory regulation, and possibly non-medical homeopathy. The regulatory status of herbal medicines is particularly unsatisfactory, and should be clarified and enforced in law. Training for CAM professionals should be standardized and independently accredited and, for many, should include basic biomedical science. Registered conventional health professionals should become more familiar with CAM. Research into CAM requires the same rigour as is required of conventional medicine, and recommendations are made as to how research could be encouraged, including pump priming by the National Health Service and Medical Research Council. The provision of information to the public and health professionals is inadequate and recommendations are made on how this should be improved. Those working in the best regulated CAM professions should work towards integration with conventional medicine. PMID- 11264967 TI - Complementary medicine education in Japanese medical schools: a survey. AB - OBJECTIVES: To evaluate the present state of complementary medicine (CM) education in Japanese medical schools. DESIGN: This investigation consisted of two studies: (1) a telephone survey to curricular office workers in September 1998; and (2) a self-completed questionnaire to representatives of sponsoring departments in July 1999. SETTINGS: All 80 medical schools for Western medicine. MAIN OUTCOME MEASURES: Presence of a CM course and sponsoring department. Titles of courses and teaching methods. RESULTS: The response rate to the telephone survey and self-completed questionnaire was 100 and 95%, respectively. Of 80 medical schools, CM was officially taught in 16 schools (20%). Of these 16 schools, there were 19 CM courses and the anesthesia department sponsored the most courses (six courses). All courses had oriental medicine titles such as acupuncture and Kampo except for one course. CONCLUSION: Twenty per cent of Japanese Medical Schools taught CM with predominantly oriental medicine themes. PMID- 11264969 TI - Randomized controlled trials: the control group dilemma revisited. AB - In some randomised controlled trials the nature of the therapy means that subjects cannot or should not be blinded. Such studies need careful design. Particular attention needs to be given to the choice of control group and the nature of the informed consent obtained from subjects, because these affect the precise research question being addressed. A survey of published studies was carried to investigate how these issues had been tackled. The paper summarizes key findings from the survey. If the research question is about the specific effect of a therapy sometimes a good case can be made for a second control group which is 'attention-controlled'. There is a need for more detailed justifications of such design decisions in published studies. PMID- 11264970 TI - Providing information on complementary and alternative therapies and practitioners by producing a directory of practitioners. AB - This article describes the experience of the authors in producing a booklet to provide information on complementary and alternative therapies (CAM) in their local area. The practical steps needed to produce a booklet are described, including ways of involving local general practitioners, meeting with local therapists, recruiting contact persons for each therapy, designing and producing a booklet and launching the final version. The feedback from general practitioners and CAM practitioners is reported. PMID- 11264971 TI - 'Neue Deutsche Heilkunde': complementary/alternative medicine in the Third Reich. AB - The aim of this article is to discuss complementary/alternative medicine (CAM) in the Third Reich. Based on a general movement towards all things natural, a powerful trend towards natural ways of healing had developed in the 19(th)century. By 1930 this had led to a situation where roughly as many lay practitioners of CAM existed in Germany as doctors. To re-unify German medicine under the banner of 'Neue Deutsche Heilkunde', the Nazi officials created the 'Heilpraktiker' - a profession which was meant to become extinct within one generation. The 'flag ship' of the 'Neue Deutsche Heilkunde' was the 'Rudolf Hess Krankenhaus' in Dresden. It represented a full integration of CAM and orthodox medicine. An example of systematic research into CAM is the Nazi government's project to validate homoeopathy. Even though the data are now lost, the results of this research seem to have been negative. Even though there are some striking similarities between today's CAM and yesterday's 'Neue Deutsche Heilkunde' there are important differences. Most importantly, perhaps, today's CAM is concerned with the welfare of the individual, whereas the 'Neue Deutsche Heilkunde' was aimed at ensuring the dominance of the Aryan race. PMID- 11264974 TI - Acupuncture on the WWW. PMID- 11264975 TI - Seventh International Symposium on Functional Medicine, Scottsdale, Arizona, May 2000. PMID- 11264976 TI - 7th Annual Symposium on Complementary Health Care, Exeter, UK 7-9 December 2000. PMID- 11264980 TI - Genes involved in initial steps of bile acid synthesis. AB - The mechanism and regulation of the degradation of cholesterol into bile acids has attracted increased interest, in particular after the recent discovery that nuclear receptors (farnesoid X receptor and liver X receptor) are involved in the regulation of bile acid synthesis. Recently, it has also been shown that the biosynthesis of bile acids is not exclusively restricted to the liver, and that degradation may start by a hydroxylation of cholesterol in the brain or in other extrahepatic organs. During the past 2 years the genes coding for three of the six enzymes catalysing the first steps in bile acid biosynthesis have been cloned and characterized. These genes and their gene products will be described here. PMID- 11264981 TI - Cholesterol metabolism in the brain. AB - The central nervous system accounts for only 2% of the whole body mass but contains almost a quarter of the unesterified cholesterol present in the whole individual. This sterol is largely present in two pools comprised of the cholesterol in the plasma membranes of glial cells and neurons and the cholesterol present in the specialized membranes of myelin. From 0.02% (human) to 0.4% (mouse) of the cholesterol in these pools turns over each day so that the absolute flux of sterol across the brain is only approximately 0.9% as rapid as the turnover of cholesterol in the whole body of these respective species. The input of cholesterol into the central nervous system comes almost entirely from in situ synthesis, and there is currently little evidence for the net transfer of sterol from the plasma into the brain of the fetus, newborn or adult. In the steady state in the adult, an equivalent amount of cholesterol must move out of the brain and this output is partly accounted for by the formation and excretion of 24S-hydroxycholesterol. This cholesterol turnover across the brain is increased in neurodegenerative disorders such as Alzheimer's disease and Niemann Pick type C disease. Indirect evidence suggests that large amounts of cholesterol also turn over among the glial cells and neurons within the central nervous system during brain growth and neuron repair and remodelling. This internal recycling of sterol may involve ligands such as apolipoproteins E and AI, and one or more membrane transport proteins such as members of the low density lipoprotein receptor family. Changes in cholesterol balance across the whole body may, in some way, cause alterations in sterol recycling and apolipoprotein E expression within the central nervous system, which, in turn, may affect neuron and myelin integrity. Further elucidation of the processes controlling these events is very important to understand a variety of neurodegenerative disorders. PMID- 11264982 TI - Xol INXS: role of the liver X and the farnesol X receptors. AB - Cholesterol and bile acid metabolism is tightly controlled by nuclear receptors. The liver X receptor, an oxysterol-activated nuclear receptor, limits cholesterol accumulation in the body both by stimulating reverse cholesterol transport and by inhibiting intestinal cholesterol absorption. The liver X receptor stimulates the adenosine triphosphate binding cassette transporter (types 1 and 8)-mediated cholesterol efflux from peripheral tissues to apolipoprotein AI and the expression of the cholesterol ester transfer protein, hence facilitating cholesterol transfer to the liver. In the liver, the liver X receptor alpha induces the cholesterol 7alpha-hydroxylase (CYP7A1) gene, which controls the rate limiting step in bile acid synthesis, the major cholesterol excretion pathway. The liver X receptor also limits cholesterol entry in the body by promoting cholesterol efflux from enterocytes into the intestinal lumen, again via an adenosine triphosphate binding cassette transporter type-mediated process. Whereas the liver X receptor is a master controller of cholesterol metabolism, the farnesol X receptor, a bile acid-activated receptor, coordinates bile acid homeostasis. Bile acids facilitate the solubilization of dietary lipids and their subsequent absorption. Bile acids enter the enterocyte through the ileal bile acid transporter and activate the farnesol X receptor, which upregulates the ileal bile acid binding protein, a carrier protein facilitating their re-uptake by the gut. Bile acids are then delivered into the portal blood and taken up in the hepatocytes by the sodium taurocholate co-transporting polypeptide. Inside the hepatocytes, activated farnesol X receptor will decrease further bile acid uptake by reducing the levels of sodium taurocholate co-transporting polypeptide, and stimulating the export of bile acid by increasing the expression of the bile salt export pump. Furthermore, the farnesol X receptor induces the small heterodimer partner, an atypical nuclear receptor, which attenuates bile acid synthesis by inhibiting the action of the orphan nuclear receptor, liver receptor homolog-1, which is a competence factor for CYP7A1 transcription. The farnesol X receptor hence stimulates bile acid re-uptake and controls bile acid production through a regulatory circuit involving both a nuclear receptor regulatory cascade and a number of specific transporter proteins. PMID- 11264983 TI - Acyl coenzyme A: cholesterol acyltransferase types 1 and 2: structure and function in atherosclerosis. AB - Two enzymes are responsible for cholesterol ester formation in tissues, acyl coenzyme A:cholesterol acyltransferase types 1 and 2 (ACAT1 and ACAT2). The available evidence suggests different cell locations, membrane orientations, and metabolic functions for each enzyme. ACAT1 and ACAT2 gene disruption experiments in mice have shown complementary results, with ACAT1 being responsible for cholesterol homeostasis in the brain, skin, adrenal, and macrophages. ACAT1 -/- mice have less atherosclerosis than their ACAT1 +/+ counterparts, presumably because of the decreased ACAT activity in the macrophages. By contrast, ACAT2 -/- mice have limited cholesterol absorption in the intestine, and decreased cholesterol ester content in the liver and plasma lipoproteins. Almost no cholesterol esterification was found when liver and intestinal microsomes from ACAT2 -/- mice were assayed. Studies in non-human primates have shown the presence of ACAT1 primarily in the Kupffer cells of the liver, in non-mucosal cell types in the intestine, and in kidney and adrenal cortical cells, whereas ACAT2 is present only in hepatocytes and in intestinal mucosal cells. The membrane topology for ACAT1 and ACAT2 is also apparently different, with ACAT1 having a serine essential for activity on the cytoplasmic side of the endoplasmic reticulum membrane, whereas the analogous serine is present on the lumenal side of the endoplasmic reticulum for ACAT2. Taken together, the data suggest that cholesterol ester formation by ACAT1 supports separate functions compared with cholesterol esterification by ACAT2. The latter enzyme appears to be responsible for cholesterol ester formation and secretion in lipoproteins, whereas ACAT1 appears to function to maintain appropriate cholesterol availability in cell membranes. PMID- 11264984 TI - Structure, function and regulation of the ABC1 gene product. AB - The role of the ATP-binding cassette transporter 1 (ABCA1) in cellular lipid efflux and high density lipoprotein metabolism has been recently documented by mutations in genetic HDL deficiency syndromes such as classical Tangier disease. Analysis of ABCA1 knockout mice and overexpression studies have established the importance of ABCA1 as a major determinant of HDL cholesterol in plasma. These studies also indicate that ABCA1 is critically involved in cellular trafficking of cholesterol and choline-phospholipids and in total body lipid homeostasis, such as intestinal cholesterol and fat-soluble vitamin absorption and in the modulation of steroidogenesis. First insights into the upregulation of ABCA1 gene expression by cellular cholesterol and cAMP have identified critical ABCA1 promoter elements, which bind the transcription factors liver X receptor, retinoid X receptor, Sp1 and E-box proteins. The finding that a lipid sensitive subgroup of ABC transporters is able to translocate cholesterol and phospholipids supports the concept that in ABCA1 deficiency, compensatory mechanisms possibly involving MDR1, MDR3 and MRP-family members could be active. This suggests that a network of ABC transporters involved in cellular lipid transport exists, which is under the tight control of energy pathways directly linked to high density lipoprotein metabolism and atherogenesis. PMID- 11264985 TI - Genetic basis of sitosterolemia. AB - The molecular mechanisms regulating the amount of dietary cholesterol retained by the body, as well as the body's ability to exclude other dietary sterols selectively, are poorly understood. An average Western diet will contain approximately 250-500 mg of dietary cholesterol and approximately 200-400 mg of non-cholesterol sterols, of which plant sterols are the major constituents. Approximately 50-60% of dietary cholesterol is absorbed and retained by the normal human body, but less than 1% of the non-cholesterol sterols are retained. There thus exists a subtle mechanism that allows the body to distinguish between cholesterol and non-cholesterol sterols. In sitosterolemia, a rare autosomal recessive disorder, affected individuals hyperabsorb and retain not only cholesterol but also all other sterols, including plant and shellfish sterols from the intestine. Consequently, patients with this disease have very high levels of plant sterols in the plasma, and develop tendon and tuberous xanthomas, accelerated atherosclerosis, and premature coronary artery disease. The STSL locus has been mapped to human chromosome 2p21. Mutations in two tandem ABC genes, ABCG5 and ABCG8, encoding sterolin-1 and -2, respectively, are now known to be mutant in sitosterolemia. The identification of these genes should now lead to a better understanding of the molecular mechanism(s) governing the highly selective absorption and retention of cholesterol by the body. Indeed, it is the very existence of this disease that has given credence to the hypothesis that there is a molecular pathway that regulates dietary cholesterol absorption and sterol excretion by the body. PMID- 11264986 TI - Very-low-density lipoprotein assembly and secretion. AB - The assembly of apolipoprotein B (apoB) into VLDL is broadly divided into two steps. The first involves transfer of lipid by the microsomal triglyceride transfer protein (MTP) to apoB during translation. The second involves fusion of apoB-containing precursor particles with triglyceride droplets to form mature VLDL. ApoB and MTP are homologs of the egg yolk storage protein, lipovitellin. Homodimerization surfaces in lipovitellin are reutilized in apoB and MTP to achieve apoB-MTP interactions necessary for first step assembly. Structural modeling predicts a small lipovitellin-like lipid binding cavity in MTP and a transient lipovitellin-like cavity in apoB important for nucleation of lipid sequestration. The formation of triglyceride droplets in the endoplasmic reticulum requires MTP however, their fusion with apoB may be MTP-independent. Second step assembly is modulated by phospholipase D and A2. Phospholipases may prime membrane transport steps required for second step fusion and/or channel phospholipids into a pathway for VLDL triglyceride production. The enzymology of VLDL triglyceride synthesis is still poorly understood; however, it appears that ACAT2 is the sole source of cholesterol esters for VLDL and chylomicron assembly. VLDL production is controlled primarily at the level of presecretory degradation. Recently, it was discovered that the LDL receptor modulates VLDL production through its interactions with nascent VLDL in the secretory pathway. PMID- 11264987 TI - Molecular mechanisms of apolipoprotein B mRNA editing. AB - A site-specific post-transcriptional cytidine to uridine deamination reaction is responsible for the production of apolipoprotein B48 in the mammalian small intestine. The molecular machinery responsible for apolipoprotein B RNA editing consists of apobec-1, an RNA-specific cytidine deaminase that functions in conjunction with a recently identified protein referred to as ACF/ASP. These proteins together represent the minimal editing enzyme, although other proteins may associate with the enzyme complex. Apobec-1 is a member of a supergene family of cytidine deaminases, with several homologs recently identified in the human genome. ACF/ASP is novel, and emerging information reveals interesting clues to its role in the apolipoprotein B RNA editing enzyme complex. PMID- 11264989 TI - Involvement of chemokine receptor 2 and its ligand, monocyte chemoattractant protein-1, in the development of atherosclerosis: lessons from knockout mice. AB - Blood monocytes are the precursors of the lipid-laden foam cells that are the hallmark of early atherosclerotic lesions, but the signals that initiate their recruitment to the vessel wall are poorly understood. Here, we review in vivo studies in genetically altered mice that support the notion that monocyte chemoattractant protein-1 (a member of the chemokine family of chemotactic cytokines) and chemokine receptor 2 (its cognate receptor) play important roles in this recruitment. An unexpected finding in chemokine receptor 2-knockout mice was the diminished production of interferon-gamma, which is a potent macrophage activator. The basis of this cytokine defect is not yet clear, but suggests that chemokines may influence atherosclerotic lesion development at several levels. Understanding the roles of chemokines and cytokines in atherogenesis may provide a basis for the development of future therapeutic agents that are aimed at interrupting monocyte recruitment and activation. PMID- 11264988 TI - Mouse models of atherosclerosis. AB - Atherosclerosis bears many features of a chronic inflammation that affects the intima of large and medium-sized arteries. In recent years apolipoprotein E deficient and LDL receptor-deficient mice have been used to examine the effects of various gene products on the development of atherosclerosis. In the present review the effects of genetics, apolipoprotein E, inflammatory gene modifiers, lipoprotein modifications, lipoprotein receptors, vessel wall expression of lipoprotein-metabolizing enzymes, and the atheroprotective role of HDL on atherosclerosis in these mice are discussed. The importance of examining lesions that are more advanced than fatty streaks and careful histologic and immunologic examination of lesion composition is emphasized. PMID- 11264991 TI - Nutrition and metabolism. PMID- 11264990 TI - Transcriptional regulatory mechanisms of the human apolipoprotein genes in vitro and in vivo. AB - The present review summarizes recent advances in the transcriptional regulation of the human apolipoprotein genes, focusing mostly, but not exclusively, on in vivo studies and signaling mechanisms that affect apolipoprotein gene transcription. An attempt is made to explain how interactions of transcription factors that bind to proximal promoters and distal enhancers may bring about gene transcription. The experimental approaches used and the transcriptional regulatory mechanisms that emerge from these studies may also be applicable in other gene systems that are associated with human disease. Understanding extracellular stimuli and the specific mechanisms that underlie apolipoprotein gene transcription may in the long run allow us to selectively switch on antiatherogenic genes, and switch off proatherogenic genes. This may have beneficial effects and may confer protection from atherosclerosis to humans. PMID- 11264992 TI - Genetics and molecular biology: genetic control of body weight. PMID- 11264993 TI - Lipid metabolism. PMID- 11264994 TI - Hyperlipidaemia and cardiovascular disease. PMID- 11264995 TI - Atherosclerosis: cell biology and lipoproteins--three distinct processes that control apolipoprotein-B secretion. PMID- 11264996 TI - Therapy and clinical trials. PMID- 11264997 TI - International perspectives on antiretroviral resistance. Phenotypic and genotypic resistance assays: methodology, reliability, and interpretations. AB - Phenotypic and genotypic resistance assays-methods to measure the susceptibility of HIV to drug therapy-are becoming widely available to clinicians for use in aiding long-term antiretroviral treatment planning. Phenotypic tests assess the actual response of a patient's viral isolate to individual antiretroviral drugs in culture, and genotypic tests sequence viral DNA, providing an inferred measure of resistance based on the evaluator's knowledge of established HIV-1 genetic mutational patterns for resistance to antiretroviral drugs. These tests, when used as intended, can provide information useful in planning successful antiretroviral regimens. Both types of assays have been shown to provide reliable and reproducible measures of resistance, with certain caveats: accuracy depends on the experience of the interpreter and laboratory; results from the available tests are not interchangeable, and clinically relevant thresholds of resistance have not been fully defined. Because of pharmacokinetic and cross-resistance issues, the extrapolation of single-drug data to combination regimens is difficult to accomplish and must be pooled with treatment and viral load history to best identify effective subsequent treatment regimens. This article describes the experience to date establishing the efficacy, accuracy, reproducibility, and limitations of the phenotypic and genotypic resistance assays used today. PMID- 11264998 TI - International perspectives on antiretroviral resistance. Nucleoside reverse transcriptase inhibitor resistance. AB - Nucleoside reverse transcriptase inhibitors (NRTIs) comprise the first class of drug with proven antiretroviral efficacy against HIV-1, and the first in which drug resistance was reported. Ongoing research in the area of NRTI resistance and cross-resistance contributes much to what we know about the failure of antiretroviral therapy. The genetic mutation patterns responsible for resistance to the available NRTIs have been well documented. This information is being used to plan rational drug therapy. Furthermore, it serves as the standard against which to evaluate response patterns to multiple-drug regimens, ultimately enabling more accurate prediction of outcome with combination therapies. Other features of NRTI resistance, such as the theoretic reversal of zidovudine resistance associated with the M184V mutation or the powerful influence of the Q151M multiple-drug resistance mutation, have revealed the unpredictable nature of HIV resistance and how much we still need to learn. Although NRTIs are the cornerstone of antiretroviral therapy at present and are used to control disease progression for extended periods, it is clear that eventually resistance occurs with all antiretroviral regimens. Future research into NRTI-resistance mutations, mutational interactions, treatment sequencing, and viral fitness and fidelity will continue to refine our understanding of drug resistance and improve our ability to delay or eliminate resistance and advance HIV control. PMID- 11264999 TI - International perspectives on antiretroviral resistance. Nonnucleoside reverse transcriptase inhibitor resistance. AB - Although understanding of nonnucleoside reverse transcriptase inhibitor (NNRTI) resistance is less clearly established than that of other classes of antiretroviral drugs, certain facts have been established. The treatment associated genetic mutation profiles of the available NNRTIs have been mapped, and resistance has been found to develop rapidly after initiation of NNRTI therapy. Despite the chemical diversity of the NNRTIs, cross-resistance among agents of this class is nearly universal. Although the viral replicative capacity ("fitness") of NNRTI-induced viral variants has not been extensively studied, available data suggest that NNRTI-selected mutations confer little damage to viral fitness, and thus a single point mutation produces a strain that is both resistant and fit. Furthermore, with continued therapy, viral evolution persists, creating species with greater numbers of mutations and higher level phenotypic resistance. Taken together, these facts suggest that continued use of NNRTIs after emergence of resistance will produce variants of complex mutational patterns that limit future treatment options, and, therefore, strong consideration should be given to discontinuing NNRTIs after virologic failure is confirmed. This article describes the scientific literature establishing the efficacy and limitations of NNRTI therapy and attempts to define a role for this class of drug in the long-term treatment of HIV-1 disease. PMID- 11265000 TI - International perspectives on antiretroviral resistance. Resistance to protease inhibitors. AB - The availability of protease inhibitors (PIs) and their combination with nucleoside reverse transcriptase inhibitors marked the passage of antiretroviral therapy (ART) from potential for control to effective suppression and thus substantially reduced rates of morbidity and mortality related to HIV. Even so, what was first hoped to be an immutable HIV DNA treatment target has proved to be prone to resistance mutations, with substitutions identified at more than 20 amino acid sites, which reduces PI susceptibility and increases resistance to treatment. The mutation patterns associated with each PI have been defined, and have been observed to occur at one of two locations: at or near the active site, or in the substrate cleavage site. The natural history of PI resistance has been extensively studied, and the genetic and cellular pathways are described in detail in this article. In addition, cross-resistance among PIs is now recognized to be fairly extensive, although the degree of cross-resistance varies with the number of mutations and the variants selected by drug pressure. Thus, it is still possible to salvage a response with another PI after a first regimen with another PI has failed. The extensive basic science and clinical experience with PIs in the fight against HIV are reviewed in this article, which provides data on resistance-mutation profiles, cellular resistance mechanisms, viral fitness studies, and clinical outcome trials with various first-line and subsequent regimens that contain PIs. It is hoped that the information provided will guide physicians in best using PIs as part of a logical and successful ART strategy. PMID- 11265001 TI - International perspectives on antiretroviral resistance. Clinical utility of resistance testing: retrospective and prospective data supporting use and current recommendations. AB - Data from retrospective and prospective studies support use of genotypic and phenotypic resistance assays to guide treatment changes when initial or subsequent antiretroviral regimens fail. Several retrospective studies have shown that response to antiretroviral therapy can be predicted based on genotypic analysis of HIV, with baseline genotypic evidence of resistance predicting virologic failure. Other retrospective analyses demonstrated that phenotypic drug sensitivity correlates with increased viral load suppression, particularly when virus remains sensitive to two or three drugs at initiation of the regimen. Furthermore, prospective studies such as VIRADAPT and Genotype-Assisted Antiretroviral Resistance Testing (GART) have substantiated that drug selection based on genotypic assay results yield superior viral suppression compared with empiric treatment assignment. One additional study suggested significant improvement in short-term virologic outcome when phenotypic testing was used to guide treatment selection. Based on these findings, resistance testing is currently recommended for patients with acute HIV infection, those who have failed one or more antiretroviral regimens, and pregnant women. Although these tests move toward becoming a standard of care, several research questions remain: the long-term benefit of resistance testing is not yet certain, the interpretation of specific genotypic resistance patterns needs to be better defined, and clinical cut-off points for phenotypic resistance need to be established. As these issues continue to be studied, resistance testing likely will prove a reliable tool to help plan successful ART strategies. PMID- 11265002 TI - International perspectives on antiretroviral resistance. Introduction. PMID- 11265004 TI - Surgical options for the treatment of severe tibial pilon fractures: a study of three techniques. AB - OBJECTIVE: To determine whether long-term results of one of three different management protocols for severe tibial pilon fractures offer advantages over the other two. DESIGN: In a retrospective study, patients were examined clinically and radiologically after internal fixation of severe tibial plafond fractures (i.e., 92 percent Type C fractures according to the AO-ASIF classification). SETTING: Department of Traumatology, Hanover Medical School. Level I trauma center. PATIENTS: Fifty-one of seventy-seven patients treated between 1982 and 1992 were examined clinically and radiologically at an average of sixty-eight months (range 13 to 130 months) after injury. INTERVENTIONS: The patients were treated in three different ways: primary internal fixation with a plate following the AO-ASIF principles (n = 15), which was reserved for patients with closed fractures without severe soft tissue trauma; one-stage minimally invasive osteosynthesis for reconstruction of the articular surface with long-term transarticular external fixation of the ankle for at least four weeks (n = 28); and a two-stage procedure entailing primary reduction and reconstruction of the articular surface with minimally invasive osteosynthesis and short-term transarticular external fixation of the ankle joint followed by secondary medial stabilization with a plate using a technique requiring only limited skin incisions (a reduced invasive technique) (n = 8). MAIN OUTCOME MEASUREMENTS: Objective evaluation criteria were infection rate, amount of posttraumatic arthritis, range of ankle movement, and number of arthrodeses. Subjective criteria were pain, swelling, and restriction of work or leisure activities. RESULTS: Because only closed fractures were treated by primary internal fixation with a plate, there was a statistically significant difference (p < 0.005) in the distribution of open fractures between the three treatment groups. Fracture classification in these groups were not significantly different. All but four fractures were classified as Type C lesions according to the AO-ASIF system. The soft tissue was closed in 63 percent (n = 32) and open in 37 percent (n = 19). No significant relationship could be found between the soft tissue damage and degree of arthritis or between the type of surgical treatment and extent of posttraumatic arthritis. However, none of the patients who required secondary arthrodesis (23 percent of all cases) were in the group who had undergone two step surgery (p < 0.05). The range of ankle movement was much greater in the two step group than in the others; these patients also had less pain, more frequently continued working in their previous profession, and had fewer limitations in their leisure activities. These differences did not reach statistical significance. The incidence of wound infection did not differ significantly among the three groups. CONCLUSIONS: On the basis of our results, we now prefer a two step procedure for the treatment of severe tibial pilon fractures with extensive soft tissue damage. In the first stage, primary reduction and internal fixation of the articular surface is performed using stab incisions, screws, and K-wires. Temporary external fixation is applied across the ankle joint. After recovery of the soft tissues, the second stage entails internal fixation with a medial plate using a reduced invasive technique. PMID- 11265005 TI - Retrograde versus antegrade nailing of femoral shaft fractures. AB - OBJECTIVES: To compare union rates and complications of retrograde intramedullary nailing of femoral shaft fractures with those of antegrade intramedullary nailing. DESIGN: Retrospective. SETTING: Level I trauma center. PATIENTS: Two hundred eighty-three consecutive adult patients with 293 fractures of the femoral shaft who underwent stabilization with antegrade or retrograde inserted femoral nails were studied. There were 140 retrograde nails and 153 antegrade nails. Twelve fractures in twelve patients were excluded (three in patients who died early in the postoperative period, three in patients because of early amputation, four in patients who were paraplegic, and two in patients who fractured through abnormal bone owing to metastatic carcinoma), leaving 134 fractures treated with retrograde nails and 147 treated with antegrade nails. One hundred four femurs treated with retrograde nails (Group R) and ninety-four femurs treated with antegrade nails (Group A) had sufficient follow-up and served as the two study groups. The average clinical follow-up was twenty-three months (range 6 to 66 months) for Group R and twenty-three months (range 5 to 64 months) for Group A. Both groups were comparable with regard to age, gender, number of open fractures, degree of comminution, mode of interlocking (i.e., static or dynamic), and nail diameter (p > 0.05). INTERVENTION: Retrograde intramedullary nails were inserted through the intercondylar notch of the knee, and antegrade nails were inserted through the pirformis fossa using standard techniques. MAIN OUTCOME MEASURES: Union, delayed union, nonunion, malunion, and complication rates. RESULTS: After the index procedure there were no significant differences in healing or incidence of malunion between Group R and Group A (p > 0.05). Healing after the index procedure occurred in ninety-one (88 percent) of the femurs in Group R and in eighty-four (89 percent) of the femurs in Group A. In Group R, there were seven delayed unions (7 percent) and six nonunions (6 percent). In Group A, there were four delayed unions (4 percent) and six nonunions (6 percent). Healing ultimately occurred in 100 (96 percent) femurs from Group R and in ninety-three (99 percent) femurs from Group A. In Group R, there were eleven malunions (11 percent), and in Group A, there were twelve malunions (13 percent). When patients with ipsilateral knee injuries were excluded, the incidence of knee pain was significantly greater for Group R patients (36 percent) than for Group A patients (9 percent) (p < 0.001). When patients with ipsilateral hip injuries were excluded, the incidence of hip pain was significantly greater for Group A patients (10 percent) than for Group R patients (4 percent) (p < 0.05). CONCLUSIONS: Retrograde and antegrade nailing techniques provided similar results in union and malunion rates. There were more complications related to the knee after retrograde nailing and more complications related to the hip after antegrade nailing. PMID- 11265006 TI - Blood flow changes to the femoral head after acetabular fracture or dislocation in the acute injury and perioperative periods. AB - OBJECTIVES: Acute blood flow to the femoral head has been postulated to be affected negatively by traumatic acetabular fracture or dislocation. To the best of our knowledge, a prospective study that has examined acute changes in blood flow to the femoral head with respect to the timing of reduction and the effect of open reduction and internal fixation after acetabular fracture or dislocations has not been performed. DESIGN AND SETTING: From June 1994 to February 1996, fifty-four consecutive patients with hip dislocations with or without fractures of the acetabulum were entered into this investigation. The patients were categorized into three groups: isolated dislocations, fractures or dislocations requiring open reduction and internal fixation, and isolated acetabular fractures without dislocation but requiring open reduction and internal fixation. Single photon emission computed tomography (SPECT) scans were obtained after relocations and preoperatively and postoperatively after open reduction and internal fixation of displaced acetabular fractures. RESULTS: The median dislocation time for all patients flow was 4.00 hours (range 1 to 24 hours). SPECT scanning showed a low blood flow pattern in five (9.25 percent) patients. A low blood flow pattern was seen in patients with early and late relocation times. Open reduction and internal fixation was not statistically associated with an avascular pattern of blood flow. Forty-two (78 percent) of our patients were available for follow-up, with an average of 24.3 months and a minimum of one year. There was one false positive, one false-negative, and thirty-eight true-negative scans. CONCLUSIONS: A global loss of scintillation in the femoral head as determined by SPECT scanning occurs in some patients with hip dislocations and fractures or dislocations of the acetabulum in the early injury period. Changes in blood flow occurred in patients with short (one hour) and long (twenty-four hours) dislocation times. However, the development of avascular necrosis could not be predicted by early SPECT scanning. Until further multicenter studies are performed, SPECT scanning cannot be recommended on an acute or routine basis to predict those patients who will develop avascular necrosis. Operative approaches for open reduction of the hip and internal fixation of acetabular fractures do not appear to affect blood flow to the femoral head. Although a golden time to relocation cannot be fully established from this study, early relocation is advised to decrease the potential risk of vascular spasm, scarring, and subsequent avascular necrosis. PMID- 11265007 TI - Fixation of periprosthetic femoral shaft fractures: a biomechanical comparison of two techniques. AB - OBJECTIVE: To determine which of two currently used techniques for the treatment of periprosthetic femoral shaft fractures provides the greater fixation rigidity and strength. DESIGN: A laboratory study using six matched pairs of femurs. METHODS: Embalmed femur prosthesis constructs had a simulated periprosthetic fracture created and were fixed with a plate with proximal cables and distal bicortical screws (Ogden concept) or two allograft struts and cables. Fixation stability was compared in various loading modalities before and after cycling. They were then tested to failure. OUTCOME MEASUREMENTS: Fixation rigidity was defined as the ratio of applied load to the amount of displacement at the fracture. RESULTS: In all loading modalities, the Ogden construct was more rigid than the allograft strut fixation. The Ogden construct required 1,295 newtons for failure and the allograft strut fixation required 950 newtons (p < 0.05). CONCLUSION: The Ogden construct provided a more rigid and stronger initial fixation of a periprosthetic fracture than did the allograft construct. PMID- 11265008 TI - Femoral neck stress fractures and metabolic bone disease. AB - OBJECTIVE: To determine whether metabolic bone disease plays a role in the cause of femoral neck stress fractures. STUDY DESIGN: Twenty-three patients with femoral neck stress fractures were enrolled prospectively in the study. Examination included computed tomography bone densitometry, trace mineral analysis, and histomorphometric analysis of the iliac crest in thirteen patients who underwent surgical treatment of their stress fractures. A control group of fifteen patients undergoing iliac crest bone grafting for scaphoid nonunions underwent similar examinations. SETTING: Tertiary military medical center. RESULTS: Patients with femoral neck stress fractures had lower bone mineral density than did control patients (p = 0.010), but no trace mineral deficiencies or consistent histomorphometric differences were noted. CONCLUSIONS: Bone mineral density is decreased in patients with femoral neck stress fractures. Despite observations of decreased bone mineral density in the stress fracture group, osteoporosis, as determined by histomorphometry, is not a consistent finding. PMID- 11265009 TI - Influence of the number of cortices on the stiffness of plate fixation of diaphyseal fractures. AB - OBJECTIVE: To determine the influence of the number of cortices of fixation on the stiffness of plate fixation of diaphyseal fractures. DESIGN: Canine experimental study. SETTING: Tertiary referral and teaching hospital in Toronto, Canada. PARTICIPANTS: Paired radii from fourteen skeletally mature, cross-bred dogs. MAIN OUTCOME MEASURE: One member of each pair of radii was tested intact as a control, and the other had a transverse osteotomy plated sequentially with five to ten cortices of fixation on either side of the simulated fracture. Dynamic compression plates and limited contact dynamic compression plates were used in two groups with seven paired radii each. Normalized torsional stiffness and four point bending stiffness were determined in the elastic range for the control and each of the plated constructs in both groups, using a materials testing machine. RESULTS: The authors found no significant difference between the stiffness of the dynamic compression plates and limited contact dynamic compression plates. With either plate of a given length, significantly increased torsional stiffness is achieved with end bicortical screws. For bending stability with the plate at right angles to the bending plane, even short plated constructs have a stiffness exceeding that of intact bone. CONCLUSIONS: For a transverse osteotomy with no fracture interdigitation, the bending rigidity with the plate at right angles to the bending plane is greater than the original stiffness of the bone for all constructs tested, with the exception of the limited contact dynamic compression plate with five cortices of fixation. The torsional rigidity of fixation only approaches the original rigidity of the bone for ten cortices of fixation with the dynamic compression plate and the limited contact dynamic compression plate. PMID- 11265010 TI - Improved intramedullary nail interlocking in osteoporotic bone. AB - OBJECTIVE: Intramedullary nail locking bolts often fail to gain purchase or cut out in osteoporotic bone. The biomechanical stability of a bladelike device that lowers intraosseous stress levels by distributing the load over a greater volume of bone was compared with conventional locking bolts in osteoporotic bone. METHODS: Standardized simulated comminuted supracondylar femoral fractures (segmental defect) in fresh-frozen paired osteoporotic (bone mineral density <200 milligrams per cubic centimeter) human cadaveric femurs were stabilized with a retrograde unreamed distal femoral nail and distally interlocked with conventional locking bolts or a bladelike device. The distal portions of the fixator-bone constructs were tested under axial load, and the stiffness and strength were compared (pairwise). RESULTS: Interlocking with a bladelike device was 41 percent stiffer (p = 0.01) and 20 percent stronger (p = 0.02) than that with conventional locking bolts. All posttesting radiographs showed compaction of the cancellous bone distal to the interlocking devices. Even after nail displacements of twelve millimeters, only a few locking bolts were plastically deformed and no bladelike device showed gross plastic deformation. CONCLUSION: This study showed the biomechanical benefits of increasing the bone-implant interface surface for improving the acute stiffness and strength of fracture fixation in osteoporotic cancellous bone. The fixator-bone construct withstood higher forces before failure in these fragile bones. PMID- 11265012 TI - Tibial portal placement: the radiographic correlate of the anatomic safe zone. AB - OBJECTIVE: To identify the radiographic correlate of the anatomic safe zone for tibial portal placement. DESIGN: Cadaveric, anatomic, and radiographic study using twenty cadaveric knees. Kirschner wires were placed in the anatomic safe zone. Anteroposterior and lateral radiographs were taken to evaluate the portal placement. SETTING: Anatomy laboratory. OUTCOME MEASUREMENTS: Radiographic measurements of Kirschner wires placed in the anatomic safe zone. RESULTS: The safe zone for tibial nail placement as seen on radiographs is just medial to the lateral tibial spine on the anteroposterior radiograph and immediately adjacent and anterior to the articular surface as visualized on the lateral radiograph. There is some variance on the anteroposterior radiograph but no variance on the lateral radiograph. CONCLUSIONS: The placement of tibial nails in the superior portion of the tibia in the documented position generates the least risk to the intraarticular structures of the knee. PMID- 11265011 TI - Biomechanical comparison of conventional open reduction and internal fixation versus calcium phosphate cement fixation of a central depressed tibial plateau fracture. AB - OBJECTIVE: To evaluate the effect of calcium phosphate bone cement on stability and strength of the fracture repair in a central depressed tibial plateau fracture cadaveric model. DESIGN: Paired human cadaveric tibial specimens. SETTING: Biomechanics laboratory. PATIENTS: Uniform pure depression fractures of lateral tibial plateau were created in twenty human cadaveric tibial specimens. INTERVENTION: The first part of the study used thirteen pairs of tibiae in two groups: a control group receiving the conventional treatment of morselized bone graft and two cancellous screws and an experimental group receiving calcium phosphate bone cement only. The second part of the study used seven pairs of tibiae in two experimental groups: one receiving calcium phosphate bone cement with a more extensive void preparation and the other group receiving calcium phosphate bone cement with a more extensive void preparation and two screws. MAIN OUTCOME MEASUREMENTS: Each tibia was loaded on a Material Testing Systems machine from twenty newtons to 250 newtons for 10,000 cycles to simulate immediate postoperative load transmission to the tibial plateau. Specimens were then loaded to failure to determine the ultimate strength of the reconstruction. Displacement of the articular fragment and stiffness at each cycle were measured during dynamic loading. Peak load, deformation at peak load, and resistance to depression were measured during the load to failure. RESULTS: The treatment of depressed tibia plateau fractures with a calcium phosphate cement provides equivalent or better stability than conventional open reduction and internal fixation in pure depression tibial plateau fractures. If the fracture void is prepared by eliminating the cancellous bone under the subchondral plate, the results are further improved. CONCLUSIONS: This study suggests that the non weight-bearing postoperative period may be significantly reduced without clinically significant articular collapse. PMID- 11265013 TI - One-year outcome after tibial shaft fractures: results of a prospective fracture registry. AB - OBJECTIVES: To describe the outcome in a consecutive series of patients with tibial shaft fractures and to determine whether prospective registration of fracture care produces useful data for clinical purposes. DESIGN: A prospective follow-up study. SETTING: A large teaching hospital in Stockholm, Sweden. PATIENTS: Sixty-four consecutive patients with a tibial shaft fracture. INTERVENTION: Patients were surgically treated according to the protocols at our institute and were followed up prospectively for one year. MAIN OUTCOME MEASUREMENTS: Clinical outcome, functional results, Short Form 36 Health Survey, Olerud Molander Ankle score, visual analogue scale. RESULTS: The fractures were classified as 42A (61 percent), 42B (31 percent), and 42C (8 percent). Forty three (67 percent) patients were treated with an interlocked tibial nail. The complication rate was low and associated with high-energy trauma. The quality of life according to the Short Form 36 Health Survey was diminished at four and twelve months after the injury, as compared with the preinjury status. Twelve months after the injury, 44 percent had not regained full function of the injured leg, although all but two of the patients had returned to preinjury working status. CONCLUSIONS: Although the complication rate was low, twelve months after the injury, nearly half the patients still experienced functional limitations related to the fracture, which was also reflected in the quality-of-life parameters. There were difficulties in retrieving data for this registry. We think that periodic, rather than continuous, registration of patient-related outcome after fracture treatment is more beneficial from a clinical and economic point of view. PMID- 11265014 TI - Effect of cutting flute design on cortical bone screw insertion torque and pullout strength. AB - OBJECTIVE: To determine the effect of the number and length of cutting flutes on the insertion torque and pullout strength for self-tapping 4.5-millimeter cortical bone screws. DESIGN: Screws were self-tapped in the diaphysis of human cadaver femurs. Each of the six screw types studied had different designs with varying cutting flute lengths and numbers. Bone mineral density, insertion torque, and pullout strength were measured. SETTING: The study was conducted at an experimental biomechanics laboratory associated with a university medical center. OUTCOME MEASUREMENTS: Insertion torque and pullout strength were normalized by the local bone mineral density. RESULTS: The mean normalized insertion torque of the design with four full-length cutting flutes was less than the design with three full-length flutes and the two designs with one-third length flutes (p < 0.05). The mean normalized pullout strength of the screw with four full-length flutes was significantly greater than that of all screws with fewer than three flutes (p < 0.05). CONCLUSIONS: Priorities for a cutting flute design should ideally include ease of screw insertion, minimal soft tissue irritation, and maximal screw holding power. Screws with more than two flutes were easier to insert and did not cause cortical damage during insertion. The screw with four full-length flutes showed a trend toward being the easiest to insert and having the greatest holding strength. PMID- 11265015 TI - Late arthroscopic debridement of metal fragments and synovectomy after penetrating knee joint injury by low-velocity missile: a report of two cases. AB - Retained metal debris from intraarticular missile injury to the knee may produce mechanical symptoms and synovitis. Arthroscopic debridement and thorough synovectomy can relieve symptoms and allow early return of function. PMID- 11265016 TI - Palmar lunate transscaphoid fracture-dislocation caused by palmar flexion injury. AB - The authors report a rare case of palmar lunate transscaphoid fracture dislocation resulting from a palmar flexion injury. After performing an open reduction of the lunate, they used a dorsal approach to fix the fractured scaphoid with a Herbert screw. A Kirschner wire fixation was also performed to stabilize the lunotriquetral joint. PMID- 11265017 TI - Traditional injury scoring underestimates the relative consequences of orthopedic injury. AB - OBJECTIVE: To demonstrate that patients with multiple injuries who have orthopedic injuries (ORTHO) face greater challenges regarding functional outcome than those without, to identify domains of postinjury dysfunction, and to illustrate the increasing discordance of functional recovery over time for ORTHO patients in relation to nonORTHO patients. METHODS: A convenience sample of adult blunt force trauma patients admitted to a Level I trauma center was evaluated at admission, and at 6 and 12 months after injury. Data were collected from the trauma registry (Trauma One), chart review, and interviews. Mailed surveys were completed 6 and 12 months after injury. The Short Form 36 (SF36) general health survey and the Sickness Impact Profile work scale (SIPw) were administered at both time points. Data are presented as mean +/- SEM or percent (%). To compare means, t tests were conducted, and Injury Severity Score (ISS) was controlled by linear regression before the evaluation of the role of ORTHO injury pattern on outcome measures. Significance is noted at the 95% confidence level (p < 0.05). RESULTS: The 165 patients studied averaged 37.2 +/- 1.1 years in age and were 67% men. The mean ISS was 14.4 +/- 0.6 and 61% had ORTHO injury. ORTHO patients were no different from nonORTHO in any measure of baseline status including the SIPw score and all domains of the SF36, except that the ISS was greater in the ORTHO group (15.6 +/- 0.96 vs. 12.7 +/- 0.73, p = 0.017). Baseline SF36 values were similar to national norms. Follow-up was 75% at 6 months, and 51% at 12 months. Those lost to follow-up differed only in that they were more likely to be men. Sixty-four percent had returned to work 12 months after injury. After controlling for ISS with linear regression, the ORTHO patients had worse scores on all physical measures of the SF36 (bodily pain, physical function, and role physical). By 12 months after injury, the relative dysfunction of the ORTHO patients had expanded to include the SIPw score (p = 0.016) and six of eight SF36 domains (bodily pain, physical function, role-physical, mental health, role emotional, and social function, all p < 0.05). CONCLUSION: Injury severity affects both mortality and the potentially more consequential issues of long-term morbidity. Patients with ORTHO injury have relatively worse functional recovery, and this worsens with time. As trauma centers approach the limits of achievable survival, new advances in trauma care can be directed more toward the quality of recovery for our patients. This will be contingent on further development of screening, scoring, and treatment systems designed to address issues of functional outcome across injury boundaries for those who survive. PMID- 11265018 TI - Antioxidant vitamin therapy alters burn trauma-mediated cardiac NF-kappaB activation and cardiomyocyte cytokine secretion. AB - BACKGROUND: This study examined the effects of antioxidant vitamins A, C, and E on nuclear transcription factor-kappa B (NF-kappaB) nuclear translocation, on secretion of inflammatory cytokines by cardiac myocytes, and on cardiac function after major burn trauma. METHODS: Adult rats were divided into four experimental groups: group I, shams; group II, shams given oral antioxidant vitamins (vitamin C, 38 mg/kg; vitamin E, 27 U/kg; vitamin A, 41 U/kg 24 hours before and immediately after burn); group III, burns (third-degree scald burn over 40% total body surface area) given lactated Ringer's solution (4 mL/kg/% burn); and group IV, burns given lactated Ringer's solution plus vitamins as described above. Hearts were collected 4, 8, 12, and 24 hours after burn to assay for NF-kappaB nuclear translocation, and hearts collected 24 hours after burn were examined for cardiac contractile function or tumor necrosis factor-alpha secretion by cardiomyocytes. RESULTS: Compared with shams, left ventricular pressure was lower in burns given lactated Ringer's solution (group III) (88 +/- 3 vs. 64 +/- 5 mm Hg, p < 0.01) as was +dP/dt max (2,190 +/- 30 vs. 1,321 +/- 122 mm Hg/s) and dP/dt max (1,775 +/- 71 vs. 999 +/- 96 mm Hg, p < 0.01). Burn injury in the absence of vitamin therapy (group III) produced cardiac NF-kappaB nuclear migration 4 hours after burn and cardiomyocyte secretion of tumor necrosis factor alpha, interleukin-1beta, and interleukin-6 by 24 hours after burn. Antioxidant therapy in burns (group IV) improved cardiac function, producing left ventricular pressure and +/-dP/dt (82 +/- 2 mm Hg, 1,880 +/- 44 mm Hg, and 1,570 +/- 46 mm Hg/s) comparable to those measured in shams. Antioxidant vitamins in burns inhibited NF-kappaB nuclear migration at all times after burn and reduced burn mediated cytokine secretion by cardiomyocytes. CONCLUSION: These data suggest that antioxidant vitamin therapy in burn trauma provides cardioprotection, at least in part, by inhibiting translocation of the transcription factor NF-kappaB and interrupting cardiac inflammatory cytokine secretion. PMID- 11265019 TI - Population-based study of hospital trauma care in a rural state without a formal trauma system. AB - OBJECTIVE: Formalized systems of trauma care are believed to improve outcomes in an urban setting, but little is known of the applicability in a rural setting. METHODS: We conducted a population-based analysis of hospital survival after trauma comparing an American College of Surgeons-verified Level I trauma center (TC) with the pooled results of 13 small community hospitals (CH) in a rural state with no formal trauma system. All patients admitted to any hospital within the state of Vermont over a 5-year period (1995-1999) with a trauma discharge diagnosis were included. Elderly patients with isolated femur fractures were excluded from the database. International Classification of Diseases Injury Severity Scores (ICISSs) were calculated for each patient and used to control for injury severity in an omnibus logistic regression model that included age, ICISS, and hospital type (TC vs. CH) as predictors of survival. Patients who died were characterized on the basis of ICISS into "expected" (ICISS < 0.25), "indeterminate" (ICISS = 0.26-0.50), and "unexpected" (ICISS > 0.5). RESULTS: In 16,354 trauma admissions over the 5-year period in the rural state of Vermont, 370 (2.2%) died. There were 5,964 (36%) admitted to TC. Patients admitted to TC were more injured (ICISS 0.94 vs. 0.96) and had a higher mortality (3.1% vs. 1.8). Overall, care at the CH provided an improved survival (odds ratio = 1.75, 95% confidence internal = 1.31-2.18, p = 0.000). However, in the more severely injured cohort of trauma patients (expected and indeterminate; n = 133), overall survival was higher in the TC (16% CH vs. 38% TC, p = 0.02, chi2). Because the TC was known to provide care equivalent to Major Trauma Outcome Study norms during this time period (Z = -0.03, M = 0.894), we believe this study confirms that trauma care throughout the state is in accordance with national norms. CONCLUSION: In a rural state, without a statewide formal trauma system, survival after trauma is no worse at CH than TC when corrected for injury severity and age. Future expenditures of resources might better be concentrated in other areas such as discovery or prehospital care to further improve outcomes. PMID- 11265020 TI - Computerized decision support for mechanical ventilation of trauma induced ARDS: results of a randomized clinical trial. AB - BACKGROUND: Variability and logistic complexity of mechanical ventilatory support of acute respiratory distress syndrome, and need to standardize care among all clinicians and patients, led University of Utah/LDS Hospital physicians, nurses, and engineers to develop a comprehensive computerized protocol. This bedside decision support system was the basis of a multicenter clinical trial (1993-1998) that showed ability to export a computerized protocol to other sites and improved efficacy with computer- versus physician-directed ventilatory support. The Memorial Hermann Hospital Shock Trauma intensive care unit (ICU) (Houston, TX; a Level I trauma center and teaching affiliate of The University of Texas Houston Medical School) served as one of the 10 trial sites and recruited two thirds of the trauma patients. Results from the trauma patient subgroup at this site are reported to answer three questions: Can a computerized protocol be successfully exported to a trauma ICU? Was ventilator management different between study groups? Was patient outcome affected? METHODS: Sixty-seven trauma patients were randomized at the Memorial Hermann Shock Trauma ICU site. "Protocol" assigned patients had ventilatory support directed by the bedside respiratory therapist using the computerized protocol. "Nonprotocol" patients were managed by physician orders. RESULTS: Of the 67 trauma patients randomized, 33 were protocol (age 40 +/- 3; Injury Severity Score [ISS] 26 +/- 3; 73% blunt) and 34 were nonprotocol (age 38 +/- 2; ISS 25 +/- 2; 76% blunt). For the protocol group, the computerized protocol was used 96% of the time of ventilatory support and 95% of computer generated instructions were followed by the bedside respiratory therapist. Outcome measures (i.e., survival, ICU length of stay, morbidity, and barotrauma) were not significantly different between groups. Fio2 > or = 0.6 and Pplateau > or = 35 cm H2O exposures were less for the protocol group. CONCLUSION: A computerized protocol for bedside decision support was successfully exported to a trauma center, and effectively standardized mechanical ventilatory support of trauma-induced acute respiratory distress syndrome without adverse effect on patient outcome. PMID- 11265021 TI - Plasma from aged stored red blood cells delays neutrophil apoptosis and primes for cytotoxicity: abrogation by poststorage washing but not prestorage leukoreduction. AB - BACKGROUND: Blood transfusion-particularly that of older stored red blood cells (RBCs)--is an independent risk factor for postinjury multiple organ failure. Immunomodulatory effects of RBC transfusion include neutrophil (PMN) priming for cytotoxicity, an effect exacerbated by longer RBC storage times. We have found that delayed PMN apoptosis in trauma patients is provoked by transfusion, independent of injury severity. We hypothesized that aged stored RBCs delay PMN apoptosis, but that prestorage leukodepletion or poststorage washing could abrogate the effect. METHODS: Healthy volunteers each donated 1 unit of blood. One half was leukodepleted, and RBC units were processed in the usual fashion and stored at 4 degrees C. Aliquots were removed on days 1, 14, 21, and 42 and the plasma fraction isolated. Selected aliquots were washed with normal saline before plasma isolation. PMNs harvested from healthy controls were incubated (5% CO2, 37 degrees C) with unmodified, leukoreduced, or washed RBC plasma (20% plasma/80% RPMI 1640), and apoptosis assessed by morphology after 24 hours. Apoptotic index (apoptotic PMNs/total PMNs) was compared. PMN priming for superoxide release was also assessed after plasma exposure. RESULTS: PMN apoptosis was delayed by RBCs stored for 21 or 42 days. Prestorage leukodepletion did not alter the effect. However, washing 42-day-old RBCs abrogated the effect. PMN priming for superoxide was provoked by stored packed RBCs in an identical pattern to delayed apoptosis. CONCLUSION: Plasma from stored RBCs-even if leukoreduced-delays apoptosis and primes PMNs. The effect becomes evident at 21 days and worsens through product outdate (42 days), but may be prevented by poststorage washing. Inflammatory agents contaminating stored blood likely mediate the effect. Modification of transfusion practices (e.g., giving fresher or washed RBCs or blood substitutes) may attenuate adverse immunomodulatory effects of transfusion in trauma patients. PMID- 11265022 TI - Acidic fibroblast growth factor attenuates the cytotoxic effects of peroxynitrite in primary human osteoblast precursors. AB - BACKGROUND: Skeletal injury and associated ischemia and inflammation induce the generation of pro-oxidants such as peroxynitrite (ONOO-), which has been demonstrated to induce apoptosis in several cell lines. Fibroblast growth factor (FGF-1) is important for coordinating osteogenesis and angiogenesis of osseous repair. In vitro studies were performed examining the effect of FGF-1 on human osteoblast progenitor stromal stem (HSS) cell proliferation, differentiation, and response to ONOO-. METHODS: HSS cells were isolated and growth kinetics determined in the presence and absence of FGF-1. The effect of FGF-1 on HSS cell expression of osteoblast-specific osteopontin and osteocalcin mRNA and protein was examined by reverse transcriptase polymerase chain reaction and Western blot techniques. To determine the sensitivity of HSS cells to ONOO- in the absence and presence of FGF-1 pretreatment, cells were exposed to varying concentrations of the oxidant and examined for cell death using quantitative fluorescence staining with fluorescein diacetate and propidium diacetate. RESULTS: Treatment of HSS cells with FGF-1 significantly enhanced cellular growth rates by 5 days (4.6 x 105 cells/mL vs. 3.1 x 105 cells/mL) and induced expression of both osteopontin and osteocalcin mRNA and protein. Exposure of HSS cells to ONOO- resulted in a dose- and time-dependent delayed cell death that was more characteristic of apoptosis than necrosis. Pretreatment of HSS cells with FGF-1 prevented ONOO- mediated apoptosis. CONCLUSION: In vitro, treatment of HSS cells with FGF-1 stimulates cell growth and induces expression of differentiation markers specific to osteoblasts. FGF-1 treatment renders osteoblast precursors resistant to the cytotoxic effects of ONOO-. These results suggest that FGF-1 promotes the progression of bone repair mechanisms by increasing the population of osteoblasts and imparting protection to the cell line from the hostile inflammatory environment. PMID- 11265023 TI - Resuscitation with a blood substitute abrogates pathologic postinjury neutrophil cytotoxic function. AB - BACKGROUND: Resuscitation with oxygen-carrying fluids is critically important in the patient with hemorrhagic shock caused by trauma. However, it is clear that a number of biologic mediators present in stored blood (packed red blood cells [PRBCs]) have the potential to exacerbate early postinjury hyperinflammation and multiple organ failure through priming of circulating neutrophils (PMNs). PolyHeme (Northfield Laboratories, Evanston, IL), a hemoglobin-based substitute that is free of priming agents, provides an alternative. We hypothesized that PMN priming would be attenuated in patients resuscitated with PolyHeme in lieu of stored blood. METHODS: Injured patients requiring urgent transfusion were given either PolyHeme (up to 20 units) or PRBCs. Early postinjury PMN priming was measured via beta-2 integrin expression, superoxide production, and elastase release. RESULTS: Treatment groups were comparable with respect to extent of injury and early physiologic compromise. PMNs from patients resuscitated with PRBCs showed priming in the early postinjury period by all three measures. No such priming was evident in patients resuscitated with PolyHeme. CONCLUSION: The use of a blood substitute in the early postinjury period avoids PMN priming and may thereby provide an avenue to decrease the incidence or severity of postinjury multiple organ failure. PMID- 11265024 TI - Ligamentous injuries of the cervical spine in unreliable blunt trauma patients: incidence, evaluation, and outcome. AB - BACKGROUND: The potential for ligamentous injury of the cervical spine (C-spine) may mandate prolonged neck immobilization via a hard cervical collar in the blunt trauma victim (BTV) with altered sensorium. We investigated the incidence of ligamentous C-spine injuries, and whether applying (post hoc) the practice management guidelines from the Eastern Association for the Surgery of Trauma (three radiograph views plus computed tomographic scan of C1-C2) would have detected the injuries. METHODS: The study was a 3-year retrospective review of BTVs admitted to the state's Primary Adult Resource Center for trauma from 1996 to 1998. Unreliable patients were defined as those with admission Glasgow Coma Scale score < 15. A rigorous algorithm to clear the C-spine was used. Pure ligamentous C-spine injury was defined as a C-spine having abnormal anatomic alignment, dislocation, subluxation, or listhesis, but without fracture. Demographics, diagnostic studies, presence of neurologic deficit, therapy, survival, and disposition were analyzed. RESULTS: There were 14,577 BTVs with 614 (4.2%) patients having C-spine injury. There were 2,605 (18%) unreliable patients, with 143 (5.5%) of these having C-spine injury, 129 (90%) having fracture and 14 (10% of BTVs; 0.5% of unreliable patients) having no fracture. Of the 14 unreliable patients with pure ligamentous C-spine injury, 13 had initial diagnosis by supine cross-table lateral radiograph. The one exception had a normal three-view radiographic series, but atlanto-occipital dislocation was diagnosed by computed tomographic scan. Eight patients had upper level injury (C0 C4) and six were lower (C4-C7). Four patients died within 30 minutes after admission, 4 underwent cervical fusion, and 6 were treated with collar only. Five (50%) of the survivors had no apparent neurologic deficit attributed to the C spine at admission. Nine patients remained institutionalized after discharge and one was discharged home. CONCLUSION: Ligamentous injuries without fracture of the C-spine are rare. Application of the practice management guidelines developed by the Eastern Association for the Surgery of Trauma for identifying C-spine instability is effective and should facilitate early removal of the cervical collar in unreliable patients. PMID- 11265025 TI - Outcome analysis of Pennsylvania trauma centers: factors predictive of nonsurvival in seriously injured patients. AB - BACKGROUND: The purpose of this study was to evaluate the impact of five trauma center characteristics on survival outcome in nine serious injury categories. METHODS: A retrospective analysis of prospectively collected data from 1992 to 1996 on patients older than 14 years of age from 24 accredited trauma centers in Pennsylvania was performed. Trauma center characteristics selected for evaluation were level of accreditation, volume of trauma admissions, presence of in-house trauma surgeons, presence of a surgical residency program, and presence of an on site medical school. Each of these characteristics was evaluated to determine its impact on survival in the selected serious injuries. A logistic regression model was then created to evaluate the most seriously injured patients as defined by A Severity Characterization of Trauma score of < 0.50. On the basis of the logistic regression model, odd ratios were calculated treating low volume as a significant risk factor for mortality. RESULTS: Of the 88,723 patients meeting registry criteria, 13,942 met the serious injury criteria. Independent analysis suggested that accreditation was beneficial regardless of level, volume of patients treated had a direct impact on survival outcome, and the presence of a surgical residency program may confer survival benefit. Of the 13,942 patients with serious injuries, those with A Severity Characterization of Trauma score of < 0.5 were selected for evaluation by logistic regression (n = 3,562). The logistic regression model, however, showed that only volume of patients treated had a consistent association with improved survival. Odds ratio analysis revealed low volume as a significant risk factor for mortality in seven of the nine injuries studied. CONCLUSION: In this analysis, only volume of patients treated had a direct impact on survival outcome. Accreditation, regardless of level, appears to be beneficial. PMID- 11265026 TI - Role of ultrasonography in penetrating abdominal trauma: a prospective clinical study. AB - BACKGROUND: Focused Assessment with Sonography for Trauma (FAST) is rapidly establishing its place in the evaluation of blunt abdominal trauma. However, no prospective study specifically evaluates its role in penetrating abdominal trauma. METHODS: Data were collected prospectively in 75 consecutive stable patients with penetrating trauma to the abdomen, flank, or back, from December 1998 to June 1999. Those with an obvious need for emergent laparotomy were excluded. FAST was performed as the initial diagnostic study on all patients. Wound location, type of weapon, and findings of diagnostic peritoneal lavage, triple-contrast computed tomographic scan, or laparotomy were recorded. The presence of peritoneal blood was noted. Data were analyzed using the chi(2) test. RESULTS: Of the 75 patients, there were 32 stab and 43 gunshot wounds. There were 66 male patients and 9 female patients; the mean age was 30 years; 41 had proven abdominal injury and 34 had no injury; and 21 patients had a positive FAST. Nineteen had peritoneal blood and injuries requiring repair at the time of laparotomy. There were two false-positive studies. Fifty-four patients had a negative FAST. In 32 patients, this was a true-negative study. Thirteen patients had a false-negative FAST and had peritoneal blood and significant injury on further evaluation. Nine patients had a negative FAST and no peritoneal blood but still had abdominal injuries requiring operative repair, including liver (four), small bowel (four), diaphragm (three), colon (three), and stomach (one). The overall sensitivity of FAST was 46% and the specificity was 94%. The positive predictive value was 90%, and the negative predictive value was 60%. CONCLUSION: FAST can be a useful initial diagnostic study after penetrating abdominal trauma. A positive FAST is a strong predictor of injury, and patients should proceed directly to laparotomy. If negative, additional diagnostic studies should be performed to rule out occult injury. PMID- 11265027 TI - Major intrahepatic bile duct injuries detected after laparotomy: selective nonoperative management. AB - BACKGROUND: Abdominal trauma causing major intrahepatic bile duct injury is a relatively uncommon occurrence. Most authorities recommend operative, usually resectional, management of these injuries when recognized, citing increased risks of complications and mortality with nonoperative management. However, very few data have been published to document the optimal management of these challenging injuries. METHODS: We present a series of five patients with significant hepatic injury and documented major bile duct injury managed at a single provincial trauma center. All of these patients had first- or second-order bile duct injuries diagnosed using endoscopic retrograde cholangiopancreatography and had developed complications caused by the ductal injury. RESULTS: In all patients, the bile duct injury and resulting complication were successfully managed by a combination of endoscopic drainage procedures and interventional radiology techniques. Average length of hospital stay for these patients was 45 days. All patients eventually attained preinjury functional status. CONCLUSION: Nonoperative techniques can be used to successfully manage selected patients and represent a reasonable alternative to operative intervention and resectional therapy, especially in the compromised patient. Extended length of stay is to be expected, but good outcomes can be achieved. PMID- 11265028 TI - Monitoring tissue oxygenation during resuscitation of major burns. AB - BACKGROUND: Because subcutaneous and splanchnic oxygenation indices are sensitive indicators of evolving hemorrhagic shock and adequacy of resuscitation, we postulated that these indices might have an equivalent role in the monitoring of severely burned patients. This observational study was undertaken to examine changes in tissue oxygenation indices during burn resuscitation. METHODS: Seven patients with major burns (54 +/- 21% total body surface area) were studied during the first 36 hours of fluid resuscitation. Silastic tubing was placed in the subcutaneous tissue just beneath both normal skin and deep partial thickness burn. Fiberoptic sensors inserted into the tubing measured subcutaneous oxygen and carbon dioxide tensions in the burnt skin (PO2scb and PCO2scb) and normal skin (PO2scn and PCO2scn) continuously. Gastric intramucosal pH (pHi) and the mucosal CO2 (PCO2m) gap were calculated using gastric tonometers. Mean arterial pressure, arterial pH, lactate, and pHi measurements were obtained for 36 hours. RESULTS: There were no significant differences in mean arterial pressure, arterial pH, or lactate concentrations throughout the study period, whereas indices of tissue oxygenation showed deterioration: pHi decreased from 7.2 +/- 0.1 to 6.7 +/- 0.3 (p = 0.06), the PCO2m gap increased from 12 +/- 17 to 108 +/- 123 mm Hg (p < 0.01), PO2scn decreased from 112 +/- 18 to 50 +/- 11 mm Hg (p < 0.01), PO2scb decreased from 62 +/- 23 to 29 +/- 16 mm Hg (p < 0.01), PCO2scn increased from 42 +/- 4 to 46 +/- 10 mm Hg (p = 0.2), and PCO2scb increased from 42 +/- 10 to 52 +/- 5 mm Hg (p = 0.05). CONCLUSION: Despite adequate global indices of tissue perfusion after 36 hours of resuscitation, tissue monitoring indicated significant deterioration in the splanchnic circulation and in the normal and burnt skin. PMID- 11265029 TI - Thermally injured patients are at significant risk for thromboembolic complications. AB - BACKGROUND: The incidence of thromboembolic complications such as deep vein thrombosis (DVT) and pulmonary embolism (PE) in thermally injured patients is considered sufficiently uncommon that routine prophylactic measures are not warranted. Nevertheless, the incidence of DVT/PE may be increasing. METHODS: The records of 1,300 patients admitted to our unit from January 1990 to June 1995 were reviewed. RESULTS: Twenty-three patients developed a clinically significant DVT, eight patients developed a PE, and two patients developed both a DVT and a PE, for an overall DVT/PE incidence of 2.9%. Four of 10 PEs were felt to be fatal. The DVT/PE patients were older (mean age, 42.6 vs. 28.7; p < 0.001) and had larger burns (37% vs. 18%, p < 0.001) than patients without evidence of DVT/PE. Body weight appeared to also influence DVT/PE rates, with obese patients (>30% over ideal body weight) having a higher incidence than patients with low or normal body weight (7.2 vs. 2.7%, p < 0.015). Age and total body surface area (TBSA) burn had a synergistic effect on DVT/PE risk, with the sum of age and TBSA burn exerting the strongest independent effect when discriminant function analysis was performed (p < 0.001). CONCLUSION: One can identify a population at increased risk of DVT/PE on the basis of the sum of age and TBSA burn, but prospective screening trials that assess all risk factors for DVT/PE should be performed before routine prophylaxis is used in thermally injured patients. PMID- 11265030 TI - Peripheral blood mononuclear cells exhibit hypercatabolic activity in response to thermal injury correlating with diminished MHC I expression. AB - BACKGROUND: Muscle wasting is one of the major consequences of severe injury or infection. Although the mechanisms underlying this hypercatabolic state are not completely characterized, it was hypothesized that other cells in the body would be similarly affected. In particular, we sought to determine whether lymphoid cell populations experienced increased protein turnover after burn injury in a fashion analogous to that seen in skeletal muscle. METHODS: BALB/c mice received either a 20% total body surface area burn or a control sham treatment. At days 1, 2, and 7 after treatment, skeletal muscle, peripheral blood, spleen, and lymph nodes were harvested from both groups. Protein synthesis and degradation rates were measured using 14C-phenylalanine incorporation and tyrosine release. Lymphocyte subpopulations (CD4 and CD8 T cells, macrophages, and B cells) and expression of major histocompatibility complex I (MHC I) molecules were assessed by flow cytometry. RESULTS: The burn model used in this study resulted in increased skeletal muscle protein turnover in the first 2 days after injury. Protein synthetic and degradation rates of peripheral blood mononuclear cells (PBMNCs) in burned mice also demonstrated comparable changes, but persisted through day 7. Splenocytes showed similar hypercatabolic effects, whereas lymph node cells showed no change. Cell viability analysis confirmed that the observed alterations were not caused by cell death. MHC I expression was depressed in tandem with the increased catabolic rate in PBMNCs. CONCLUSION: This study demonstrates that various lymphoid populations undergo protein catabolic changes similar to those characteristically observed in skeletal muscle, and these correlated with diminished MHC I expression. Moreover, PBMNCs exhibited prolonged sensitivity to burn injury, of a duration exceeding that observed in skeletal muscles or other lymphoid tissues. PMID- 11265031 TI - Objective estimates of the incidence and consequences of multiple organ dysfunction and sepsis after burn trauma. AB - BACKGROUND: Organ dysfunction and sepsis are frequent after major burn trauma, represent quantifiable consequences of the systemic response to injury, and may be important end points by which to measure treatment effectiveness. However, standard and widely applied methods for their measurement have not been applied to burn trauma victims. Therefore, the purpose of this study was to quantify these complications after burn trauma. METHODS: Patients with > or = 20% total body surface area burns admitted to a single center were prospectively enrolled. Standard sepsis criteria and multiple organ dysfunction (MOD) scores for the pulmonary, renal, cardiovascular, hepatic, and hematologic systems were determined. The incidence and risk factors for severe MOD (cumulative MOD score > or = 6) and severe sepsis were determined. The relationships between these complications and mortality and resource utilization were examined by univariate and multivariate analyses. RESULTS: A total of 85 patients were enrolled over 1 year. Severe MOD developed in 24 (28%) and severe sepsis or septic shock developed in 12 (14%). Both were associated with increasing age and burn size and were more likely to occur in men. Most patients who developed severe MOD or severe sepsis survived (71% and 67%, respectively), and both were associated with longer intensive care unit stays and duration of mechanical ventilation. CONCLUSION: According to simple and objective scoring systems, severe MOD and severe sepsis/septic shock are both related to burn size, age, and male sex. Both are related to intensive care unit length of stay and duration of mechanical ventilation. PMID- 11265032 TI - Prevalence and importance of pneumothoraces visualized on abdominal computed tomographic scan in children with blunt trauma. AB - BACKGROUND: Chest radiographs are routinely obtained for the identification of pneumothoraces in trauma patients. Computed tomographic (CT) scanning has a higher sensitivity for the detection of pneumothoraces, but the prevalence and importance of pneumothoraces detectable by CT scan but not by chest radiography in children sustaining blunt trauma is unclear. METHODS: We conducted a prospective observational cohort study of children less than 16 years old with blunt trauma undergoing both abdominal CT scan and chest radiography in the emergency department of a Level I trauma center over a 28-month period. All abdominal CT scans were interpreted by a single faculty radiologist. The chest radiographs of all patients with pneumothoraces detected on CT scan as well as a random sample of chest radiographs from pediatric blunt trauma patients without pneumothoraces on abdominal CT scan (in a ratio of four normals per pneumothorax) were reviewed by a second faculty radiologist. Both radiologists were masked to all clinical data as well as to the objective of the study. RESULTS: Five hundred thirty-eight children underwent both abdominal CT scan and chest radiography in the emergency department. Twenty patients (3.7%; 95% confidence interval [CI], 2.3-5.7%) were found to have pneumothoraces on CT scan. Of these 20 patients, 9 (45%; 95% CI, 23-68%) had pneumothoraces identified on initial chest radiography and 11 patients did not ("unsuspected pneumothoraces"). Twelve pneumothoraces were identified in these 11 patients; 6 were graded as minuscule and 6 as anterior according to a previously established scale. One patient with an unsuspected pneumothorax underwent tube thoracostomy. None of the 10 patients (0%; 95% CI, 0-26%) with unsuspected pneumothoraces who were managed without thoracostomy (including two patients who underwent positive pressure ventilation) had complications from their pneumothoraces. CONCLUSION: Less than half of pediatric blunt trauma patients with pneumothoraces visualized on abdominal CT scan had these pneumothoraces identified on initial chest radiograph. Patients with pneumothoraces identified solely on abdominal CT scan, however, uncommonly require tube thoracostomy. PMID- 11265033 TI - Blood alcohol concentration and management of road trauma patients in the emergency department. AB - BACKGROUND: The effects of blood alcohol on injury after crash are controversial, and safe limits are not settled. We examined if a positive blood alcohol concentration, even in a nontoxic range, affects management and outcome of injured patients after road crashes. METHODS: In this prospective cohort study, we recruited all adult subjects admitted to an emergency department within 4 hours after a road crash. Outcomes were mortality or expected permanent disability, and data related to patients' management. RESULTS: Alcohol-positive trauma patients were more frequently critical at admission (odds ratio [OR], 1.89; 95% confidence interval [CI], 1.18-3.02), and had an increased risk of combined mortality or expected permanent disability (OR, 1.67; 95% CI, 1.08 2.58), need for intensive care (OR, 1.87; 95% CI, 1.01-3.46), surgery (OR, 1.91; 95% CI, 1.37-2.66) and blood transfusions (OR, 2.09; 95% CI, 1.20-3.64), and acute medical complications (OR, 1.94; 95% CI, 1.33-2.85). All these events were explained by higher trauma severity. Only the risk of unsuspected injuries, diagnosed only at final evaluation, was independently associated with a positive blood alcohol concentration (OR, 4.98; 95% CI, 3.62-6.87), in addition to trauma severity and preexisting chronic conditions. Blood alcohol measurement significantly improved the accuracy in predicting unsuspected injuries, from 81.3% to 86.2%. CONCLUSION: In injured patients after a road crash, a positive blood alcohol concentration increases the chance that the final diagnosis will include more injuries than initially documented. More careful monitoring is needed in alcohol-positive trauma patients, independent of clinical status, injury severity, and overt symptoms of alcohol intoxication. PMID- 11265034 TI - Routine cervical spine radiography for trauma victims: Does everybody need it? AB - OBJECTIVE: The purpose of this study was to evaluate the indication for routine cervical spine radiography in trauma patients. METHODS: Prospective analysis of radiologic and clinical findings was performed during a 5-year period. Patients suitable for a clinical decision rule were reviewed separately. RESULTS: Of the 1,757 consecutive patients included in the study, 38 were diagnosed with a cervical spine injury. Of the 599 patients suitable for the clinical decision rule, 62 had midline cervical tenderness, including 2 with cervical spine injury. No additional cervical spine injuries were found during follow-up. CONCLUSION: It is within good practice, and it is also cost-effective, to obtain a cervical spine radiograph only on clinical parameters in trauma patients with no apparent bodily trauma and optimal parameters. With this clinical decision rule, 30.6% of all cervical spine series were redundant, and no (occult) spinal fractures would have been undetected. PMID- 11265035 TI - Basal release of nitric oxide and its interaction with endothelin-1 on single vessel hydraulic permeability. AB - BACKGROUND: Both endothelin-1 (ET-1) and nitric oxide (NO) are released by the endothelium and are implicated in modulating the permeability of the endothelial barrier. The present study was designed to examine the interaction between ET-1 and NO and its influence on microvascular permeability as well as the role of NO in maintaining microvascular permeability. To isolate the direct effect of ET-1 and NO, experiments were conducted under conditions where hydraulic and oncotic pressures were controlled. METHODS: Postcapillary venules in the rat mesentery were perfused in situ and paired measurements of hydraulic permeability (Lp) obtained using the modified Landis micro-occlusion method. The effect of basal endogenous NO was tested by measuring the effects of perfusion with the NO synthase inhibitor Nw-nitro-L-arginine-methyl-ester (L-NAME) (100 micromol/L) on Lp (n = 6). In addition, Lp measured after a 15-minute perfusion with L-NAME (100 micromol/L) was compared with measures of Lp obtained after perfusion with a combined mixture of L-NAME (100 micromol/L) and ET-1 (80 pmol/L) (n = 6). RESULTS: Units for Lp are mean +/- SE x 10(-8) cm x sec(-1) x cm H2O(-1). Under basal conditions, in the absence of exogenous ET-1, NO synthase inhibition led to a significant increase in Lp from 5.7 +/- 0.5 to 9.8 +/- 1.4 (p = 0.02). Compared with L-NAME alone, ET-1 + L-NAME significantly decreased Lp from 10.3 +/- 0.8 to 5.7 +/- 0.6 (p = 0.006). CONCLUSION: Constitutive release of NO from the microvascular endothelium plays a role in maintaining a basal level of microvascular permeability. Decreases in microvascular permeability seen with the administration of ET-1 are not mediated via the release of NO. These findings suggest important roles for ET-1 and NO in maintaining and modulating microvascular permeability. PMID- 11265036 TI - Nonunion after intramedullary nailing of humeral shaft fractures. AB - OBJECTIVE: To assess results of exchange nailing in nonunion after intramedullary (IM) nailing of humeral shaft fractures. METHODS: This was a retrospective study; 24 patients with nonunion after IM nailing of humeral shaft fractures were reviewed. In 13 cases, nonunion was treated using exchange nailing, and 11 patients were treated nonoperatively. Union was assessed from radiographs. Shoulder joint symptoms and function were assessed after a mean 4.7 years' follow up using Constant-Murley scoring and self-administered questionnaires devised by L'Insalata et al. RESULTS: Single or repeated exchange nailing resulted in union in 6 of 13 patients. Shoulder joint function was satisfactory (mean Constant Murley score of 72) for those patients whose fracture eventually united and poor (mean Constant-Murley score of 39) for those left with nonunion. CONCLUSION: Exchange nailing results in a poor union rate in nonunion after IM nailing of humeral shaft fractures. Permanent nonunion of the humeral shaft leaves the patient with severe disability. PMID- 11265037 TI - RAB-plate versus sliding hip screw for unstable trochanteric hip fractures: stability of the fixation and modes of failure--radiographic analysis of 218 fractures. AB - BACKGROUND: The sliding hip screw has gained considerable acceptance in the treatment of unstable trochanteric fractures. However, the new type of 120 degrees fixed angle blade-plate with a buttress rod (RAB-plate) showed encouraging clinical results. The purpose of this study was to assess stability of fixation and analyze modes of failure in unstable trochanteric hip fractures treated with these devices. METHODS: A retrospective radiographic review of 218 unstable fractures was performed. Linear and angular displacements of the major fragments and implant migration into the femoral head during healing were assessed. Additionally, adequacy of the reduction and the location of the implant within the femoral head as predictors of fixation failure were evaluated. RESULTS: The postreduction neck-shaft angle was maintained in the majority of the fractures in both groups. However, there was a significantly higher incidence of varus angulation by 10 degrees or more by the completion of healing among fractures treated with the sliding hip screw (p = 0.04). There was no statistically significant difference in vertical migration of the device into the femoral head between the implants used (p = 0.3). There was a significant relationship between failure of the fixation and varus reduction (p = 0.04) as well as screw/neck angle deviation more than 20 degrees in the lateral projection (p = 0.005) or if the implant was in a superior or posterior position (p = 0.02). CONCLUSION: The RAB-plate provided a more stable fixation, especially with regard to maintained postoperative alignment. However, positive predictors for fixation failure were identical for both devices. Here, the screw/neck angle deviation has had the strongest significance for prediction of fixation failure. PMID- 11265038 TI - Traumatic pleurocaval fistula: potential source of air embolism. PMID- 11265039 TI - Abducens nerve palsy and ipsilateral Horner syndrome: a predicting sign of intracranial carotid injury in a head trauma patient. PMID- 11265040 TI - Contralateral acute subdural hematoma occurring after evacuation of subdural hygroma: case report. PMID- 11265041 TI - Penetrating lung hernia with pulmonary evisceration: case report. PMID- 11265042 TI - Brachial plexus innervated, functional tensor fasciae latae muscle transfer for controlling a Utah Arm after dislocation of the shoulder caused by an electrical burn. PMID- 11265043 TI - Simultaneous fracture of the body of the hamate and the distal pole of the scaphoid. PMID- 11265044 TI - Acute thumb ischemia secondary to high-pressure injection injury: salvage by emergency decompression, radical debridement, and free hallux hemipulp transfer. PMID- 11265046 TI - Incomplete quadriplegia resulting from minor trauma: initial presentation of ossification of the posterior longitudinal ligament. PMID- 11265045 TI - Traumatic avulsion of the bony insertion of infraspinatus tendon. PMID- 11265047 TI - Endovascular therapy for priapism secondary to perineal trauma. PMID- 11265048 TI - Fatal cerebral fat embolism after open reduction and internal fixation of femur fracture. PMID- 11265049 TI - Re: Treatment of humeral shaft fractures by closed fixation using multiple intramedullary Kirschner wires. PMID- 11265050 TI - Scoring system for the prediction of mortality. PMID- 11265051 TI - Beneficial effects of a hospital bereavement intervention program after traumatic childhood death. AB - PURPOSE: An investigation of the experiences of parents grieving the traumatic death of their child, the initiatives that helped, and common parental concerns that would benefit from improved education. METHODS: From January 1, 1995, to December 31, 1998, 81 of 3,501 children admitted to our pediatric trauma center died. An attempt was made to enroll all parents. Interactions included family contact at hospital, home/funeral home visit within 1 month of death, educational meeting with parents and 15 supporters at a restaurant within 2 months of death, follow-up survey to parents/supporters, and final interview/survey with parents in 1999-2000. RESULTS: Seventy-seven families were enrolled; 59 families completed the educational meeting with supporters, and 245 parental supporters returned surveys. Supporters were likely to use proposed interventions (82%), were more accepting of the duration of grief (94%), and interacted with parents more often after the death (78%). Parents (n = 44) felt the hospital staff was appropriately sensitive to their child (90%), themselves (93%) and prepared them for their child's death (81%). Parents (n = 54) on behalf of 37 children have completed the final interview. Poor conceptualization of aspects of the medical care and brain death, and delayed regret for missing the opportunity to donate organs, were recurring themes. CONCLUSION: We conclude that parents' unanswered questions or misconceptions regarding brain death, organ donation, and their child's medical care adversely affect their grief; that "normal life" for parents is challenged as they struggle to establish a new sense of normal; and that hospital and trauma service personnel can positively impact the grieving process with appropriate training. PMID- 11265052 TI - Origins of timing errors in human sensorimotor coordination. AB - The authors analyzed fluctuations in timing errors when 8 human participants attempted to coordinate movement with external rhythmic signals. The temporal dynamics of the errors is usually described in terms of simple, self-correcting models. Here the authors demonstrate that timing errors are characterized by a 1/f(alpha) type of long memory process. The value of the exponent alpha differentiates different types of coordination states: synchronization and syncopation. More interesting, evidence was found that alpha can be changed when participants use different coordination strategies. Together with the authors' understanding of the generation mechanism for long memory processes, these results suggest that 1/f(alpha) type of long-range correlated timing errors are of higher cortical origin and are likely the outcome of distributed neural processes acting on multiple time scales. PMID- 11265053 TI - Cognitive activity shifts the attractors of bimanual rhythmic coordination. AB - The attractors of bimanual rhythmic coordination are given as the solutions of a motion equation in relative phase. How are those attractors affected by cognitive activity? In 3 experiments, participants (N = 10 in Experiments 1 and 2; N = 5 in Experiment 3) were required to produce in-phase or antiphase coordination while they either did or did not perform an information-reduction task. The average absolute deviation from in-phase (0 degrees ) and antiphase (180 degrees ) satisfying a particular parameterization of the motion equation was amplified by cognitive activity. That amplification of absolute phase shift was the same for both in-phase and antiphase coordination. Furthermore, the amplification (in degrees) increased linearly with the magnitude of cognitive activity (in bits). Cognitive activity had limited influence on the variability of relative phase and did not affect its average signed deviation. Collectively, the results suggest that cognitive activity produces a shift in the attractors of bimanual coordination dynamics that is directionally nonspecific and is independent of movement speed, detuning, and the in-phase-antiphase distinction. PMID- 11265054 TI - Response facilitation and inhibition in manual, vocal, and oculomotor performance: evidence for a modality-unspecific mechanism. AB - In 2 experiments (N = 10, Experiment 1; N = 16, Experiment 2), the authors investigated whether evidence for response facilitation and subsequent inhibition elicited by masked prime stimuli can be observed for output modalities other than manual responding. Masked primes were followed by target stimuli that required a 2-choice manual, saccadic, or vocal response. Performance was measured for compatible trials in which primes and targets were identical and for incompatible trials in which they were mapped to opposite responses. When primes were presented centrally, performance benefits were obtained for incompatible trials; whereas for peripherally presented primes, performance benefits were found in compatible trials. That pattern of results was obtained for manual responses and for saccadic eye movements (Experiment 1), demonstrating that those effects are not mediated by specialized dorsal pathways involved in visuomanual control. An analogous pattern of effects was found when manual and vocal responses were compared (Experiment 2). Because vocal responding is controlled by the inferotemporal cortex, that result shows that prime-target compatibility effects are not primarily mediated by the dorsal stream, but appear to reflect modality unspecific visuomotor links that allow rapid activation of motor responses that may later be subject to inhibition. PMID- 11265055 TI - Grasping a better understanding of the intrinsic dynamics of rhythmical and discrete prehension. AB - In this study, the authors examined the influence of the intrinsic dynamics of discrete and rhythmical prehension. Six adults underwent a scaling procedure in which the movement time was systematically increased so that it corresponded with 6 frequencies: 0.5, 0.75, 1.0, 1.25, 1.5, and 1.75 Hz. In posttests, participants moved at their own preferred pace. No differences were found in the relative time to final hand closure (T[rfc]) between the rhythmical and discrete conditions. The variability of T[rfc] was shown to be less at the preferred step of scaling than during the posttest. With the scaling technique, one can guide participants into more stable movement patterns than they can achieve when the metronome is not present, because, when the metronome is present, their movements become anchored to the external pacing cue. Those findings provide support for the use of a scaling technique to identify the influence of the intrinsic dynamics during rhythmical and discrete movements. PMID- 11265056 TI - Control strategies when intercepting slowly moving targets. AB - In 3 experiments, the authors investigated and described how individuals control manual interceptive movements to slowly moving targets. Participants (N = 8 in each experiment) used a computer mouse and a graphics tablet assembly to manually intercept targets moving across a computer screen toward a marked target zone. They moved the cursor so that it would arrive in the target zone simultaneously with the target. In Experiment 1, there was a range of target velocities, including some very slow targets. In Experiment 2, there were 2 movement distance conditions. Participants moved the cursor either the same distance as the target or twice as far. For both experiments, hand speed was found to be related to target speed, even for the very slowly moving targets and when the target-to cursor distance ratios were altered, suggesting that participants may have used a strategy similar to tracking. To test that notion, in Experiment 3, the authors added a tracking task in which the participants tracked the target cursor into the target zone. Longer time was spent planning the interception movements; however, there was a longer time in deceleration for the tracking movements, suggesting that more visually guided trajectory updates were made in that condition. Thus, although participants scaled their interception movements to the cursor speed, they were using a different strategy than they used in tracking. It is proposed that during target interception, anticipatory mechanisms are used rather than the visual feedback mechanism used when tracking and when pointing to stationary targets. PMID- 11265057 TI - Horizontal plane head stabilization during locomotor tasks. AB - Frequency characteristics of head stabilization were examined during locomotor tasks in healthy young adults(N = 8) who performed normal walking and 3 walking tasks designed to produce perturbations primarily in the horizontal plane. In the 3 walking tasks, the arms moved in phase with leg movement, with abnormally large amplitude, and at twice the frequency of leg movement. Head-in-space angular velocity was examined at the predominant frequencies of trunk motion. Head movements in space occurred at low frequencies (< 4.0 Hz) in all conditions and at higher frequencies (> 4.0 Hz) when the arms moved at twice the frequency of the legs. Head stabilization strategies were determined from head-on-trunk with respect to trunk frequency profiles derived from angular velocity data. During natural walking at low frequencies (< 3.0 Hz), head-on-trunk movement was less than trunk movement. At frequencies 3.0 Hz or greater, equal and opposite compensatory movement ensured head stability. When arm swing was altered, compensatory movement guaranteed head stability at all frequencies. Head stabilization was successful for frequencies up to 10.0 Hz during locomotor tasks. Maintaining head stability at high frequencies during voluntary tasks suggests that participants used feedforward mechanisms to coordinate head and trunk movements. Maintenance of head stability during dynamic tasks allows optimal conditions for vestibulo-ocular reflex function. PMID- 11265058 TI - Motor learning as a function of KR schedule and characteristics of task-intrinsic feedback. AB - Forty participants (age range = 18-35 years) practiced 1 of 2 versions of an aiming task (with or without spring resistance). Knowledge of results (KR) was provided to them either immediately or after a delay of 2 trials. Immediate KR led to significantly more accurate performance during the 80 trials in acquisition but significantly less accurate performance on a 40-trial retention test given 24 hr after practice. In addition, the spring version of the task was performed significantly less accurately than the no-spring version on the 24-hr retention test. Most important, a significant interaction on the 24-hr retention test revealed that performance of the no-spring version of the task, when KR had been given after a 2-trial delay, was significantly more accurate than performance of the other 3 combinations of task version and KR schedule. The results suggest that KR dependency in motor skill learning is related to familiarity with task-intrinsic feedback in addition to the schedule on which KR is presented. PMID- 11265059 TI - Static and phasic cross-talk effects in discrete bimanual reversal movements. AB - The authors examined the hypothesis that the phasic and the static cross-talk effects found in bimanual movements with different target amplitudes originate at different functional levels of motor control, which implies that the effects can be dissociated experimentally. When the difference between the short and the long amplitudes assigned to the 2 hands of 12 participants was decreased, the static effect disappeared, In contrast, the phasic effect, which can be observed only at short preparation intervals, did not disappear; although it became smaller in absolute terms, in relative terms it did not. In addition, the authors compared the time course of amplitude variability and examined the correlation between left hand and right hand amplitudes. The disappearance of the phasic amplitude assimilation at increasing preparation intervals turned out to be delayed relative to the decline of the correlation between amplitudes. That finding suggests that the assimilation of mean amplitudes and the correlation between left hand and right hand amplitudes are not fully equivalent indicators of intermanual interactions, but may indicate different kinds of inter-limb coupling. PMID- 11265060 TI - Spatial conceptual influences on the coordination of bimanual actions: when a dual task becomes a single task. AB - When the left and right hands produce 2 different rhythms simultaneously, coordination of the hands is difficult unless the rhythms can be integrated into a unified temporal pattern. In the present study, the authors investigated whether a similar account can be applied to the spatial domain. Participants (N = 8) produced a movement trajectory of semicircular form in single-limb and bimanual conditions. In the bimanual tasks, 1 limb moved above the other in the frontal plane. Bimanual unified tasks were constructed so that the spatial paths to be produced by the 2 limbs could be easily conceptualized as parts of a unified circle pattern. Bimanual distinct tasks availed a less obvious spatial pattern that would unify the 2 tasks, despite similar demands placed on the coordination dynamics in the 2 cases (e.g., the phase relations). The authors conclude that a dual task becomes a single task, and interlimb interference is reduced, when the spatial patterns produced by the 2 hands form a geometric arrangement that can be conceptualized as a unified representation. PMID- 11265061 TI - An algorithm for mapping positively selected members of quasispecies-type viruses. AB - BACKGROUND: Many RNA viruses do not have a single, representative genome but instead form a set of related variants that has been called a quasispecies. The sequence variability of such viruses presents a significant bioinformatics challenge. In order for the sequence information to be understood, the complete mutational spectrum needs to be distilled to a biologically relevant and analyzable representation. RESULTS: Here, we develop a "selection mapping" algorithm--QUASI--that identifies the positively selected variants of viral proteins. The key to the selection mapping algorithm is the identification of particular replacement mutations that are overabundant relative to silent mutations at each codon (e.g., threonine at hemagglutinin position 262). Selection mapping identifies such replacement mutations as positively selected. Conversely, selection mapping recognizes negatively selected variants as mutational "noise" (e.g., serine at hemagglutinin position 262). CONCLUSION: Selection mapping is a fundamental improvement over earlier methods (e.g., dN/dS) that identify positive selection at codons but do not identify which amino acids at these codons confer selective advantage. Using QUASI's selection maps, we characterize the selected mutational landscapes of influenza A H3 hemagglutinin, HIV-1 reverse transcriptase, and HIV-1 gp120. PMID- 11265062 TI - Fatal cardiac tamponade after emergency tension-free repair of a large paraesophageal hernia. AB - Paraesophageal hernia is an unusual disorder of the esophageal hiatus that may be associated with life-threatening mechanical problems. We report a case of a large paraesophageal hernia that presented with acute thoracic herniation and incarceration of the stomach. The patient underwent laparoscopic operation, including reduction of an intrathoracic stomach, hernial sac removal, and tension free repair of the hiatus with polytetrafluoroethylene (PTFE) mesh. The mesh was fixed with a straight hernia stapler. Postoperatively the patient developed a fatal cardiac tamponade secondary to a coronary vein laceration due to fixation of the mesh with the stapler. Different operative techniques and possibilities for prevention of the complication are discussed. PMID- 11265063 TI - Gallbladder melanoma mimicking acute acalculous cholecystitis. AB - Gallbladder (GB) melanoma is a rare entity with a dismal prognosis. Its primary or secondary status is difficult to establish in the absence of an overt cutaneous localization. We report herein the case of a misdiagnosed GB melanoma mimicking acute acalculous cholecystitis that was treated by means of laparoscopic cholecystectomy (LC). A 54-year-old man was referred to our institution for acute cholecystitis. Apart from the ablation of some nevocytic nevi 7 years before admission, the patient's medical history was unremarkable. The ultrasound (US) examination showed a slightly enlarged acalculous gallbladder with thickened walls and a well-circumscribed polypoid mass in the fundus. The patient was treated medically and referred to LC. At surgery, some satellite nodules were visualized in the GB hepatic bed. The GB was removed, and two hepatic nodules were excised. Histology showed a pT3 melanoma. The patient underwent an open hepatic wedge resection 3 weeks after laparoscopy. No recurrence was observed at 6-month follow-up. To date, only one case of melanoma of the gallbladder treated with LC has been reported. GB melanoma is a diagnostic challenge when there is no evidence of a primary lesion. However, the occurrence of acalculous cholecystitis and a GB polyp in patients with a positive history of mole ablation should alert surgeons to the possibility of a melanoma. PMID- 11265064 TI - Incidental discovery of pelvic cholelithiasis on diagnostic laparoscopy. AB - Laparoscopic cholecystectomy, although a safe technique for removal of the gallbladder, is associated with surgery-related complications. We report a complication caused by this procedure in a young woman, in whom a diagnostic laparoscopy led to the incidental finding of retained gallstones in the pelvis. A 30-year-old woman underwent diagnostic laparoscopy for exclusion of an ectopic pregnancy. She had undergone laparoscopic cholecystectomy 3 years before presentation. Diagnostic laparoscopy ruled out ectopic pregnancy, but yellow nodules were noted in her pelvis. Removal of these nodules was not attempted because of their close proximity to the ureter. A biopsy of these lesions confirmed amorphous material suggestive of gall stones. The finding was documented in the case notes and explained to the patient after the procedure. PMID- 11265065 TI - Intraabdominal abscess managed successfully via the laparoscopic approach. AB - A rare complication of laparoscopic fundoplication-an intraabdominal abscess located between the fundus and the caudate lobe of the liver-is described. A 41 year-old man had undergone a laparoscopic Nissen-Rossetti fundoplication for longstanding gastroesophageal reflux disease. On the 5th postoperative day, the patient's general condition became worse, and he developed intermittent-remittent fever (40 degrees C), an elevated white blood cell count (WBC), and an accelerated sedimentation rate. Evidence of leakage was excluded by Gastrografin swallow. The diagnosis was finally revealed by means of ultrasound and computed tomography (CT) scan, which showed an intraabdominal fluid collection with an air cap of ~10 cm in diameter situated between the diaphragmatic crura, the caudate lobe of the liver, and the gastric fundus. The location did not allow semi invasive management of the abscess, such as ultrasound or CT-guided puncture and drainage. On the 8th postoperative day, a laparoscopic exploration was performed utilizing the previous port sites. The adhesions were easily dissected, and evacuation of ~300 ml of white, dense fluid, and lavage and drainage were performed without intraabdominal dissemination of pus. The patient was discharged on the 12th postoperative day free of symptoms. Microbiological examination of the pus showed the presence of Peptostreptococcus. The patient remained symptom free. At 8 weeks postoperatively, barium swallow, endoscopy, 24-h pH monitoring, and stationary manometry of the esophagus yielded normal results. Because there was no direct evidence of leakage at the fundus, the development of the abscess was concluded to be due to the use of deep transmucosal stitches rather than seromuscular ones to create the wrap. The nonabsorbable multifilament suture material passing through the entire gastric wall could have facilitated bacterial contamination of the operative field. PMID- 11265067 TI - Intraperitoneal abscess after an undetected spilled stone. AB - Gallbladder perforation with loss of calculi in the abdomen is frequent during laparoscopic cholecystectomy and can cause serious late complications. We report on a 65-year-old woman who underwent laparoscopic cholecystectomy for gallbladder empyema, during which a stone spilled into the peritoneal cavity. The spilled gallstone was not noticed during the initial operation. Three months later, she reported left upper quadrant pain of recent onset without associated symptoms such as fever, nausea, or weight loss. On examination, a palpable 2-cm tender subcutaneous mass was found. Abdominal ultrasound demonstrated an incarcerated hernia, and computed tomography (CT) scan showed an intraperitoneal abscess located in the back of the anterior abdominal wall in the left upper quadrant, which contained a recalcification figure. The patient was brought to surgery, at which time an incision was made over the mass. A chronic abscess in the back of the abdominal wall, also spreading into the subfascial space, was drained, and purulent material was obtained with a large stone, 2.8 cm in diameter, which had become lodged in the rectus abdominis after an undetected stone spillage during laparoscopic cholecystectomy. The patient continued receiving antibiotic treatment for 7 days, recovered well, and was discharged 7 days after drainage of the abscess. PMID- 11265066 TI - Palliative treatment of recurrent obstructing gastric cancer. A case report of successful treatment with three self-expanding metallic stents. AB - Patients with advanced or recurrent gastric cancer affecting the upper and lower gastrointestinal tract usually experience obstructive symptoms, causing a severe compromise in their quality of life. Surgery may not be feasible because of the patient's precarious medical condition and multilevel tumor infiltration. When faced with these circumstances, surgeons have few options. Parenteral nutrition and comfort measures are utilized when surgical bypass is not a tenable option. We herein describe a unique case of multilevel upper and lower gastrointestinal obstruction secondary to recurrent gastric cancer. The patient was treated palliatively through a combined surgical, radiological, and endoscopic approach by implanting a series of self-expanding metallic stents. To our knowledge, there are no previous reports of successful management of simultaneous strictures of the upper and lower gastrointestinal tract using this technique. PMID- 11265068 TI - A mucinous tumor of the mesocolon with features of borderline malignancy. AB - Primary mucinous cystadenoma of the mesocolon is a rare tumor with an uncertain histogenesis. A 38-year-old woman was diagnosed with a 17-cm cystic lesion in the left abdomen, identified as a mucinous cystadenoma of the mesocolon. This type of tumor appears rarely in extraovarian sites. We believe that metaplasia, either celomic or mucinous, is the most likely pathogenic mechanism. On rare occasions, a borderline or invasive component may be present. PMID- 11265069 TI - Unexpected cyanosis in the surgical patient. AB - Benzocaine is a commonly used topical anesthetic present in many over-the-counter preparations. The development of methemoglobinemia, associated with the use of benzocaine, is potentially fatal. Methemoglobinemia remains unresponsive to the administration of oxygen alone and, in fact, results in greater tissue hypoxemia than the usual monitoring techniques indicate. We report two cases of benzocaine induced methemoglobinemia occurring 3 months apart at the same institution. A brief discussion regarding methemoglobin is presented. PMID- 11265070 TI - Multiendoscope-assisted treatment for blue rubber bleb nevus syndrome. AB - Blue rubber bleb nevus syndrome is characterized by gastrointestinal and cutaneous hemangiomas and gastrointestinal bleeding causing anemia. We report a unique case of this syndrome in an adult woman. It was associated with congenital heart disease, for which the patient underwent surgery at 12 months of age, and cutaneous hemangiomas, for which surgery was performed later in childhood. Gastrointestinal bleeding was diagnosed and treated when she was 21 years of age after a workup for iron deficiency anemia. Successful total resection of all gastrointestinal hemangiomas was performed by minimally invasive surgery with gastric, small intestinal, and colonic fiberscopy and laparoscopy. The postoperative course was uneventful. The patient could walk the day after surgery, and she was discharged from the hospital 14 days after surgery. Our experience and findings given in other reports suggest that total resection of hemangiomas should be the final goal and that minimal skin incision is preferable for this benign disease, with multiendoscope-assisted treatment to ensure that any hemangiomas remaining in the gastrointestinal tract are not overlooked. PMID- 11265071 TI - Lymph node metastases identified with mediastinoscopy in a patient with superficial carcinoma of the esophagus. AB - Superficial esophageal cancers limited to the lamina propria are not associated with lymph node metastases. Mediastinoscopic transhiatal esophagectomy was planned in a patient with widespread superficial cancer of the midthoracic esophagus. Sampling of the upper mediastinal lymph nodes revealed metastases. The operation was converted to a transthoracic esophagectomy with radical lymphadenectomy. Histopathologic examination of the resection specimen showed three metastatic lymph nodes, despite local invasion limited to the lamina propria. This is the first report of a patient with superficial esophageal cancer and lymph node metastases. PMID- 11265072 TI - Esophageal hypermotility associated with intramural pseudodiverticulosis. Primary esophageal disease or epiphenomena? AB - Esophageal intramural pseudodiverticulosis is a very rare disease of unclear etiology. The clinical picture is characterized by progressive dysphagia. Because of its frequent association with alcohol abuse and subsequent weight loss, it must be differentiated reliably from esophageal carcinoma. The diagnosis is established by the characteristic detection of multiple intramural contrast accumulations in the barium esophagogram. Additional endoscopic and endosonographic confirmation and histological examination are required to exclude a malignant tumor. Moreover, associated diseases are almost always present and should also be diagnosed by pH-metry, cytology, and esophageal manometry. Good and long-lasting therapeutic success can be achieved by bouginage of the stenosis with concomitant treatment of the associated esophageal diseases. Based on two case reports of patients with this disease, we discuss the unusual association with esophageal hypermotility as well as the symptoms, clinical course, therapy, and pathogenesis of the disease. PMID- 11265073 TI - Captopril-associated cholestasis complicating the management of pancreatic cancer. AB - Cholestatic jaundice is a rare complication associated with the use of the angiotensin -converting enzyme inhibitor captopril. The severity of the disease may range from cholestasis on liver histology to overt fulminant hepatic failure. This diagnosis is seldom considered in patients with pancreatic or biliary tract malignancy. We present a patient with unresectable adenocarcinoma of the pancreas whose jaundice decreased slowly over many weeks despite establishment of adequate endoscopic biliary drainage. The presence of captopril-associated cholestasis confounded confirmation of adequate biliary drainage. The absence of observed hepatic bile secretion at duodenoscopy, as seen in this patient, is a previously unreported endoscopic feature of this syndrome. PMID- 11265075 TI - Impact of aortic diameter on the outcome of surgical treatment of abdominal aortic aneurysm. AB - Several recent articles have discussed ultrasonographic surveillance of small abdominal aortic aneurysms (AAA). The purpose of this study was to evaluate the impact of lesion size on immediate morbidity and mortality after surgical treatment of AAA. More specifically we investigated whether the mortality rate was lower after treatment of AAA measuring PMID- 11265074 TI - Aortic aneurysm: construction of a life-size model by rapid prototyping. AB - Development of new endovascular techniques for repair of abdominal aortic aneurysm (AAA) requires the use of experimental models. Stereolithography is a rapid prototyping technique used in industry to prototype parts during the design phase. A stereolithography apparatus (STL) employs laser technology to build a digital model layer by layer with photopolymer resin. The purpose of this study was to use this technology to produce a life-size AAA model. Data were acquired by CT scan and stored in DICOM 3 format. Specifically designed software was used for 3-D imaging and conversion of data to a standard STL format. Two replicas were made: one to scale and the other 3 mm larger. The final model was made by pouring silicone rubber or polyurethane into the mold over the life-size model so as to obtain a sturdy, life-size, soft, transparent plastic casting. Arterial models made for living subjects with these rapid prototyping techniques can be used to simulate surgical procedures, calibrate imaging modalities, and design new stent grafts. PMID- 11265077 TI - Diagnosis and treatment of type II endoleak after stent placement for exclusion of an abdominal aortic aneurysm. AB - After endovascular treatment of AAA, regular clinical and radiologic surveillance is necessary for early diagnosis and treatment of mid-term and long-term complications. The purpose of this report was to evaluate the efficacy of magnetic resonance imaging (MRI) in screening for type II endoleaks and assessing the results of treatment by embolization. From March 1996 to November 1999, 64 patients with uncomplicated infrarenal abdominal aortic aneurysm (AAA) were treated by endovascular exclusion with a covered aortic stent. Radiological surveillance included plain abdominal roentgenogram (PAR), CT scan, and pelvioabdominal MRI at 1 month, 3 months, 6 months, and every 6 months thereafter. Arteriography was performed routinely after 1 year or sooner if an endoleak was suspected. Based on the results of this study, MRI seems to be more sensitive than CT scanning for detection of type II endoleaks. The negative predictive value of MRI is also better. In this series, all endoleaks were treated by embolization. In most cases, the maximum transverse diameter and maximum anteroposterior diameter decreased after embolization. Further follow-up will be necessary to confirm these findings. PMID- 11265078 TI - Comparison of color duplex ultrasound and computed tomography scan for surveillance after aortic endografting. AB - Endovascular repair of abdominal aortic aneurysms (AAA) requires regular surveillance for early detection of endograft failure. CT scanning is the gold standard surveillance procedure. The purpose of this study was to assess the reliability of color duplex ultrasound (CDU) in comparison to CT scanning for detection of endoleaks and changes in aneurysmal diameter. From November 1996 to September 1999, a total of 41 patients treated by aortic endografting underwent regular surveillance with both CT scanning and CDU. There were 39 men and 2 women with a mean age of 71 years (range, 50-83). Endovascular treatment involved deployment of a straight aorto-aortic stent in 6 cases, bifurcated stent in 33, and aorta-to-unilateral iliac artery stent in 2. Stent deployment failed in one case; the procedure was conversion to open surgery. Primary or secondary endoleaks were detected in 17 patients (42%). Our findings indicated that CDU is less reliable than the CT scan for detection of endoleaks, but that reliability of CDU for surveillance of aneurysmal diameter is fair. PMID- 11265079 TI - Long-term outcome of 121 iliofemoral endarterectomy procedures. AB - Iliofemoral endarterectomy (EA) is now considered by most vascular surgeons to be an obsolete technique that is difficult and unreliable. The purpose of this retrospective study was to reassess the place of iliofemoral EA on the basis of long-term outcome in our experience. From 1982 to 1995, we performed a total of 121 iliofemoral EA procedures on 98 patients with a mean age of 57 years. The anatomical presentation involved iliac occlusion in 55 cases and complex iliac stenosis in 63. The indication for treatment was critical ischemia in 28 cases. Operative mortality was nil. Major amputation was required in only one patient because of contralateral thrombosis during the procedure. Postoperative thrombosis requiring early thrombectomy occurred in five cases. At 5 and 10 years, actuarial rates were 77.6% and 61.3%, respectively, for survival, 98.3% and 90.1%, respectively, for limb salvage, 78.9% and 65.1%, respectively, for primary patency, and 88.2% and 73.8%, respectively, for secondary patency. On the basis of these findings, we consider iliofemoral EA to be a viable alternative to iliac bypass in patients ineligible for transluminal angioplasty. PMID- 11265076 TI - Endovascular treatment of occlusive lesions in the distal aorta: mid-term results in a series of 31 consecutive patients. AB - The purpose of this study was to evaluate the early and mid-term results of endovascular treatment of occlusive lesions in the distal aorta in a consecutive series of patients. Between February 1996 and March 1999, a total of 31 patients underwent transluminal procedures for treatment of occlusive atherosclerotic lesions located at the lower end of the aorta. Thirty patients presented with intermittent claudication and one had critical ischemia. Manifestations were bilateral in 26 cases and unilateral in 5. The lesion was confined to the lower aorta in 3 patients and extended to the common iliac arteries in 19, with predominant proximal lesions of the common iliac artery occurring in 9 patients. Fourteen patients had concurrent infracrural occlusive lesions. All patients underwent exclusive endovascular treatment without any associated open surgical procedure. The three patients with isolated aortic lesions were treated by angioplasty, followed by stent placement in two cases. The 19 patients with aortobiiliac lesions were treated by bilateral common iliac artery angioplasty according to the "kissing-balloon" technique; 7 of these patients also underwent aortic angioplasty. In these 19 patients, aortic stenting was performed in 3 cases and bilateral iliac stenting in 10 cases, including 3 in association with aortic stenting. The nine patients with a proximal lesion of the common iliac arteries were treated by angioplasty, followed by bilateral stenting in three cases and unilateral stenting in three cases. The findings of this study show that the mid-term anatomical and functional results of endovascular treatment for atherosclerotic lesions of the distal aorta are satisfactory. We recommend it as the initial treatment modality. PMID- 11265080 TI - Do internal iliac arteries contribute to vascularization of the descending colon during abdominal aortic aneurysm surgery? An intraoperative hemodynamic study. AB - The inferior mesenteric artery (IMA) is the nutrient artery for the descending colon. Colon ischemia after repair of abdominal aortic aneurysm (AAA) can be prevented by routine or elective revascularization of the IMA. In case of occlusion of the IMA, revascularization of the internal iliac artery (IIA) has been recommended but its effectiveness has never been documented. In this study, intraoperative hemodynamic monitoring of the IMA was performed to determine if the IIA contributed significantly to the region supplied by the IMA. From January 1998 to August 1999, a total of 223 patients underwent AAA repair at 11 vascular surgery centers. The IMA was occluded in 113 of these patients (51%). This study involves the other 110 patients (49%) with patent IMA. Study consisted of measuring residual systolic arterial pressure in the IMA (IMAP) immediately after AAA repair. To compensate for blood pressure variations, systolic pressure in the radial artery (RAP) was measured concurrently and the inferior mesenteric index (P) was calculated by dividing IMAP by RAP. Measurements were made before and during cross-clamping of the IIA to obtain two corresponding indexes-i.e., P1 and P2, respectively. Mean P1 and P2 were 0.61 (95% confidence interval, 0.8-0.4) and 0.58 (95% confidence interval, 0.55-0.61), respectively, with p PMID- 11265081 TI - Percutaneous transluminal angioplasty for management of critical ischemia in arteries below the knee. AB - Percutaneous transluminal angioplasty (PTA) can be performed safely in arteries below the knee by using current coaxial catheters. This study includes 37 consecutive patients treated between March 1992 and March 1999 by PTA for limb threatening infrageniculate occlusive artery disease. The mean duration of follow up was 28 months. Limb salvage was achieved in 32 patients. The actuarial limb salvage rate at 2 years was 87 +/- 6%. This study shows that PTA was a viable alternative to surgical treatment for management of critical lower extremity ischemia in carefully selected patients. Limb salvage rates after PTA and conventional surgical revascularization seem comparable. Based on these findings, we recommend that PTA be attempted, whenever possible, for initial treatment of patients presenting critical, limb-threatening ischemia due to isolated or multiple stenoses of below-knee arteries. PMID- 11265083 TI - Optimization of the resistance of arterial allografts to infection: comparative study with synthetic prostheses. AB - Arterial allografts can be used for in situ treatment of prosthetic graft infection. The purpose of this in vitro study was to compare the resistance of allografts and synthetic prostheses to infection by five strains of bacteria and to study antibiotic treatments designed to reduce allograft infection. Fresh and cryopreserved allografts were compared with synthetic prostheses made of various biomaterials including PTFE, plain Dacron, gelatine-sealed Dacron, and gelatine sealed, rifampicine-bonded Dacron. Allografts were used with or without treatment using an antibiotic containing gentamycine, lincomycine, and vancomycine. The bacterial strains tested were Escherichia coli, Staphylococcus aureus, slime producing Staphylococcus epidermidis, non-slime-producing Staphylococcus epidermidis, and Pseudomonas aeruginosa. Infection was evaluated by counting the number of adherent bacteria on the allograft or synthetic material after rinsing and ultrasonication. Statistical analysis was achieved using nonparametric Mann Whitney tests. Results showed that allografts not treated with antibiotics were highly susceptible to bacterial infection. Antibiotic treatment decreased infection. Application of antibiotic after thawing cryopreserved allografts led to a significant decrease. None of the biomaterials tested provided sufficient protection against bacteria resistant to the antibiotics used. PMID- 11265082 TI - Reduction of groin lymphatic complications by application of fibrin glue: preliminary results of a randomized study. AB - Lymphoceles and lymph fistulas are common complications after exposure of the common femoral artery in the Scarpa triangle because of operative transsection of overlying lymphatics. The purpose of this prospective randomized study was to determine the incidence of groin lymphatic complications and to assess the impact of routine application of fibrin glue on lymphatic structures and subcutaneous tissue prior to closure. All patients undergoing exposure of the common femoral artery in the Scarpa triangle were included in this study. They were divided into two groups according to closure technique. In group A, closure was performed without fibrin glue. In Group B, fibrin glue was applied to lymphatic structures prior to closure. The efficacy of fibrin glue application was estimated on the basis of two criteria: incidence of local complications and amount of lymphatic fluid in the Redon drain. The preliminary findings suggest that application of fibrin glue leads to a significant reduction in the incidence of lymphatic complications after femoral artery exposure in the Scarpa triangle. PMID- 11265084 TI - Infected aneurysms of neck and limb arteries: a retrospective multicenter study. AB - Infected aneurysms (IA) of neck and limb arteries are uncommon. This report describes the results of a retrospective study undertaken by the University Association for Surgical Research (AURC) to evaluate etiology, bacteriology, location, diagnostic features, and therapeutic methods associated with IA. A total of 58 IA in 52 patients were reviewed. The lesion was located in a lower extremity artery in 47 patients (81%), internal carotid artery in 7 (12%), and upper extremity artery in 4 (6%). Eleven patients had multilocular aneurysm (21%). Symptoms of local infection were observed in 43 patients (82.6%). Rupture or splitting was the presenting manifestation in 13 patients (25%). Primary IA following bacteremia or septicemia without endocarditis was the most common type of IA observed in 34 patients (65.3%). Twelve patients (23%) presented mycotic IA secondary to bacterial endocarditis. In the remaining six patients (11.5%), IA resulted from direct contamination or spreading from a contiguous infection site. Surgical treatment included ligation of the artery without reconstruction in 19 patients and exclusion bypass in 33 patients. The duration of antibiotic treatment ranged from 15 days to 3 months. No recurrence of aneurysm was observed but three patients developed bypass infection. Primary IA was associated with high mortality due to severe septicemia. PMID- 11265085 TI - Surgical exposure of superior sulcus lung tumors with vascular involvement. AB - Choice of exposure route for surgical excision of superior sulcus lung tumors depends on involvement at the thoracic inlet. From December 1985 to September 1999, we performed surgical treatment of superior sulcus tumors in 42 patients, including 22 with vascular involvement. Various exposure techniques were used, including a novel technique combining transverse supraclavicular cervicotomy and posterolateral thoracotomy in 11 cases, anterior transclavicular cervicothoracotomy in 7 cases, isolated posterolateral thoracotomy in 3 cases, and cervicosternotomy in 1 case. Vascular procedures consisted of subadventitial dissection of the subclavian artery in 5 patients, arterial resection-anastomosis in 7, and prosthetic bypass in 10. Postoperative mortality was 11.9% in the overall series of 42 patients (n = 5) and 9% (n = 2) in the subgroup of patients with vascular involvement. During follow-up, 13 patients died of tumor recurrence and 1 patient died of respiratory insufficiency. Actuarial 5-year survival was 22.7 +/- 17.5% overall and 18 +/- 17.9% in the subgroup of patients with vascular involvement. This study indicates that the combined exposure route with transverse supraclavicular cervicotomy and posterolateral thoracotomy was useful for treatment of superior sulcus lung tumors requiring lobectomy and pneumonectomy. PMID- 11265086 TI - Long-term results of venous revascularization for Paget-Schroetter syndrome in athletes. AB - The indications for surgical management of primary subclavian vein thrombosis are not agreed upon. This report describes our experience in the treatment of exertional thrombosis causing Paget-Schroetter syndrome in 10 athletes between 18 and 45 years of age. In seven patients urokinase was injected and this resulted in complete revascularization in three cases (42.9%). The remaining three patients were treated by anticoagulation using heparin with adjuvant Coumadin after resolution of thrombosis. Duplex ultrasonography and dynamic phlebography were performed to assess the results of drug treatment. Decompression of the thoracic outlet was performed secondarily using various techniques, including first-rib resection in 10 cases, scalenectomy in 9, and resection of a clavicular callus in 1 case. In six patients, persistent debilitating venous stasis required revascularization by axillojugular bypass in three cases, thrombectomy in two, and axillojugular anastomosis in one case. All patients were reexamined. Mean follow-up was 45 months. Symptomatic relief and vein patency was achieved in all cases. All patients were able to resume sports activity. In agreement with previous studies, our findings confirm the efficacy of immediate anticoagulation, thrombolysis, and complete decompression of the thoracic outlet. Should this approach fail to reestablish patency, axillosubclavian vein revascularization can provide good mid-term results. PMID- 11265087 TI - Compromised hemodynamics associated with multipedicular lesions of cerebral arteries. AB - The purpose of this study was to demonstrate that severe multipedicular lesions involving supraaortic trunks cause compromised cerebral hemodynamics with nonhemispheric symptoms (NHS) that can be relieved by surgical treatment. A total of 11 patients were prospectively included in the study. Regional cerebral blood flow (rCBF) and cerebral blood flow reactivity (CBFR) were measured by acetazolamide single photon emission computed tomoscintigraphy scans (SPECT) before and after surgery. Seven patients presented with isolated NHS and four presented with NHS associated with hemispheric symptoms. Lesions consisted of either high-grade (>75%) bilateral carotid artery stenosis associated with vertebral or subclavian artery lesions or high-grade (>75%) bilateral vertebral or subclavian artery stenosis associated with medium-grade (>50%) carotid lesions. All patients presented with a functional circle of Willis with no significant intracranial arterial lesions and no corticosubcortical atrophy. A total of 15 procedures were performed for revascularization of 19 arteries. The cumulative morbidity/mortality rate was nil. All revascularizations were patent on postoperative controls. Results from this study show that multipedicular lesions lead to hemodynamic changes affecting hemispheric and vertebrobasilar territories. Surgical treatment can improve or normalize cerebral hemodynamic abnormalities and relieve NHS. PMID- 11265088 TI - Treatment of nonmalignant obstructive iliocaval lesions by stent placement: mid term results. AB - This report describes mid-term results of endovascular treatment of obstructive iliocaval lesions. Between November 1995 and December 1999, a total of 15 patients were treated by angioplasty and stent placement in the iliac vein. These patients were divided into two groups. Group I consisted of six patients with acute iliofemoral thrombosis of less than 10 days duration, with associated caval involvement in three cases. Angioplasty was performed after surgical thrombectomy, and creation of an arteriovenous fistula as a one-stage procedure in four cases and as a two-stage procedure in two cases. The underlying chronic lesion was stenosis of the left iliocaval junction (Cockett syndrome) in five cases and retroperitoneal fibrosis in one. Group II comprised nine patients with chronic symptomatic stenosis or occlusion. The etiology was Cockett syndrome in seven cases, post-thrombotic syndrome in three cases, including two associated with Cockett syndrome, and retroperitoneal fibrosis in one case. The mean number of stents per patient was 1.5. The mean duration of follow-up was 23.5 months. Evaluation of clinical outcome according to CEAP criteria for chronic syndromes showed significant improvement. Given good mid-term findings, venous angioplasty with stent placement appears to be a safe and effective technique for treatment of acute or chronic obstructive iliocaval lesions. PMID- 11265089 TI - Internal to external carotid artery transposition to repair recurrent stenosis after carotid artery stenting. AB - Recently, carotid artery stenting (CAS) has emerged as a treatment option for carotid artery stenosis. Since the procedure is new, management of its complications is not standardized. This case report describes one method of arterial reconstruction after failed CAS. A 64-year-old male underwent CAS of his right internal carotid artery (ICA) for an asymptomatic 65% stenosis. Seven months later the stented area had narrowed to 95%. Arteriography revealed that the common and external carotid arteries (ECA) were free of disease so we elected to perform a transposition of the distal ICA onto the proximal ECA. The ECA and its branches were completely mobilized and the ascending pharyngeal and lingual arteries divided. The ICA was divided distal to the stent. Transection of the occipital artery provided an arteriotomy for an end ICA to side ECA anastamosis, thus preserving ECA flow. Postoperative surveillance after 8 months has revealed no recurrent stenosis. Operative repair of restenosis after CAS may be challenging if standard endarterectomy is not possible. Other options for reconstruction are feasible but if the common and external carotid arteries are disease-free, an ICA to ECA transposition provides a simple all-arterial repair that avoids bypass and prosthetic material. PMID- 11265091 TI - Cardiac morbidity and mortality following carotid endarterectomy: the importance of diabetes and multiple Eagle risk factors. AB - The objectives of this study were to (1) determine cardiac morbidity and mortality in patients undergoing carotid endarterectomy (CEA) and (2) to determine whether any Eagle risk factors and/or the indication for CEA are associated with a higher risk of an adverse cardiac event. The records of 123 male patients who underwent CEA were retrospectively reviewed. The Eagle risk factors for adverse cardiac events, indications for CEA, and adverse cardiac events were recorded and analyzed. In males undergoing CEA, the presence of diabetes or multiple Eagle risk factors significantly increases the risk of an adverse postoperative cardiac event. There is no difference in cardiac event rate between symptomatic and asymptomatic CEA. Thus, asymptomatic CEA should be approached with caution in these higher cardiac risk patients. PMID- 11265090 TI - Carotid body tumors: the role of preoperative embolization. AB - Resection of carotid body tumors (neck paragangliomas) carries inherent risks of injury to the cranial nerves and other structures as well excessive blood loss. Preoperative embolization has been used to lessen the morbidity in tumors that are larger than 2 cm in diameter. Two female patients presented for treatment with large asymptomatic carotid body tumors-one 4 cm and one 5 cm in diameter. Both patients had preoperative angiography the day before surgery that revealed the feeding arterial vessels so that successful embolization could be accomplished with gel. Success was judged by diminution of the angiographic blush. Both patients had an uneventful surgical excision the following day with the carotid body tumors being able to be resected periadventitially without damage to either the external or internal carotid artery. The cranial nerves were preserved in both patients and blood loss was only 200 cc in both cases. We conclude that preoperative embolization is an important adjunct in treating patients with large carotid body tumors. The surgical exploration proceeds much smoother, the blood loss is minimal, and patients have minimal morbidity. PMID- 11265092 TI - Homocysteine-associated acute mesenteric artery occlusion treated with thrombectomy and bowel resection. AB - Elevated plasma homocysteine is an acknowledged risk factor for arteriosclerotic occlusive disease, but little clinical evidence is available regarding its role in acute arterial thrombosis in the absence of an underlying lesion. A 45-year old man presented with an acute abdomen. A magnetic resonance arteriogram (MRA) showed occlusion of the superior mesenteric artery. At exploration, necrotic ileum was resected and the superior mesenteric artery was thrombectomized, restoring normal mesenteric flow. The plasma homocysteine level was 98.8 mmol/L, more than eight times the normal level. No embolic source was identified and an MRA and contrast arteriogram showed no residual occlusive disease in the superior mesenteric artery. Additional studies documented pernicious anemia, which was treated with cobalamin (vitamin B12) injections. This case provides further evidence of an association between hyperhomocysteinemia and acute arterial thrombosis. Hyperhomocysteinemia can result from easily correctible vitamin B12, B6, or dietary folate deficiencies. Plasma homocysteine levels should be determined in young individuals with acute arterial thrombosis whenever a hypercoagulable state is suspected. PMID- 11265093 TI - Primary aortoenteric fistula: computed tomographic diagnosis of an atypical presentation. AB - A primary aortoenteric fistula is a potentially devastating complication of untreated aortic aneurysmal disease. The clinical presentation can be confusing, leading to a delay in diagnosis. Computed tomography (CT) can greatly assist in establishing the diagnosis. An unusual case of a primary aortoenteric fistula with an atypical presentation is described. The patient presented with symptoms indicating an exacerbation of recurrent nephrolithiasis. No clinical history of an abdominal aortic aneurysm or previous history of gastrointestinal hemorrhage was reported. A CT scan demonstrated extravasation of arterial contrast into the duodenum. The aorta was repaired with an in-line prosthetic graft. A review of the literature regarding this rare entity and surgical options are presented. PMID- 11265094 TI - Penetrating ulceration of the infrarenal aorta: case reports of an embolic and an asymptomatic lesion. AB - Penetrating aortic ulceration is uncommon in the infrarenal aorta. We describe a patient with a penetrating infrarenal aortic ulcer manifesting as blue toe syndrome, and a second patient with a similar lesion identified as an incidental finding. These two patients were treated for penetrating infrarenal aortic ulceration within the past 9 months at two university-affiliated hospitals, a regional Veterans Administration Medical Center, and a County Medical Center. Both lesions demonstrated aneurysm changes with varying degrees of mural thrombus. The lesion filled with fresh thrombus proved labile, with embolization manifesting as blue toe syndrome. We support the aggressive treatment of aneurysmal penetrating aortic ulcer with aortic graft replacement to eliminate the potential for distal embolization and to obviate the risk of rupture and death. PMID- 11265095 TI - Renal artery endarterectomy for treatment of renovascular hypertension combined with infrarenal aortic reconstruction: analysis of surgical results. AB - From June 1995 to February 2000, 16 patients with renovascular hypertension had bilateral transaortic renal artery endarterectomy (RA TEA) combined with either infrarenal aortic aneurysm repair (8 patients) or infrarenal aortodistal bypass for occlusive disease (8 patients). Aortic clamp level for RA TEA was supraceliac in eight patients and suprarenal in eight patients with a mean clamp time of 19 min (range 14 to 25 min). Perioperative complications occurred in four patients. These included respiratory insufficiency with prolonged intubation (1 patient), prolonged intubation with transient renal failure requiring temporary dialysis (1 patient), acute thrombosis of right limb of aortofemoral bypass graft (1 patient) and major left hemispheric cerebrovascular accident (1 patient). Results from this contemporary patient series demonstrate acceptable perioperative morbidity and mortality when RA TEA for treatment of renovascular hypertension is combined with infrarenal aortic reconstruction. In this setting, either supraceliac or suprarenal aortic clamping for short time periods appears to be well tolerated. Clinical outcome is enhanced by salvage of renal function, decrease in medication requirement, and improvement in blood pressure control. PMID- 11265096 TI - An abnormal dipyridamole thallium/sestamibi fails to predict long-term cardiac events in vascular surgery patients. AB - Recent data demonstrate that dipyridamole-thallium (DTHAL) and sestamibi (DMIBI) are not predictive of adverse perioperative cardiac events in moderate-risk patients (one or more Eagle risk factors) undergoing major elective vascular surgery. Less data are available regarding the ability of DTHAL/DMIBI to predict adverse cardiac events on long term follow-up. We sought to determine whether an abnormal DTHAL/DMIBI is predictive of adverse cardiac events on long-term follow up in moderate-risk patients undergoing major elective vascular surgery. Patients were enrolled prospectively between June 1997 and June 1999 at West Los Angeles VA and Harbor-UCLA Medical Centers. Adverse cardiac events were defined as congestive heart failure (CHF), myocardial infarction (MI), unstable angina (USA), and ventricular arrhythmias. Follow-up was obtained via clinic visits, telephone calls, and chart review. We studied 75 patients (76% male, 24% female) with a mean age of 65 years. Operative procedures were primarily femorodistal (83%) and aortic (16%). DTHAL/DMIBI results were normal in 35 patients (47%), demonstrated reversible ischemia in 26 (35%), and showed a fixed defect alone in 14 (18%). From the follow-up results of this study we conclude that there is no association between a reversible ischemia or an abnormal (fixed or reversible) DTHAL/DMIBI and adverse cardiac events or mortality on long-term follow-up in moderate-risk patients who have undergone major vascular surgery. PMID- 11265099 TI - Videoconferencing in the provision of psychological services at a distance. AB - This paper reviews the literature on the provision of psychological services using videoconferencing. First, mental health assessments are considered in terms of both the initial interview and the use of scales for rating symptoms of mental state dysfunction, including psychosis, depression and anxiety. Ways to increase the reliability of initial assessment data collected by videoconference are provided, and the consumer's experience of receiving this service by videoconference is also considered. Research comparing the administration of psychometric tests in person and by videoconference is then reviewed, as is the client's experience of receiving this service by videoconference. Psychological interventions provided for individuals, families and groups by videoconference are also considered. Positive and negative experiences relate to issues of empathy, working alliance, a sense of control and a sense of presence. The levels of comfort and satisfaction expressed by both counsellors and clients with the use of the medium are discussed. Recommendations for how best to use videoconferencing for psychological interventions are offered and contraindications are reviewed. Videoconferencing for the purposes of supervision is also briefly covered. The legal issues associated with the use of videoconferencing to provide psychological services include consent, reimbursement, professional licensing and liability. It seems that videoconferencing is a new and potentially beneficial means of bringing psychological services to isolated communities. However, it may be necessary to explore the technique cautiously. There is a dearth of evidence regarding the reliability of psychological services provided using videoconferencing and consequently there is vast opportunity for further research. PMID- 11265100 TI - Patient referral by telemedicine: effectiveness and cost analysis of an Intranet system. AB - The clinical effectiveness and costs of telemedicine in improving the referral process from primary to secondary care were examined in an eight-month prospective, comparative study with one-year follow-up. The internal-medicine outpatient clinics of two Finnish district hospitals were compared--Peijas Hospital (PH) with telemedicine and Hyvinkaa Hospital (HH) without it. The three primary-care centres studied referred a total of 292 adult patients to the outpatient clinics. The population-based number of referrals to PH (7.5/1000) from primary-care centres was twice that to HH (3.8/1000). Thirty-seven per cent of referrals to PH included requests from general practitioners for on-line medical advice (teleconsultation). Forty-three per cent of the total number of intranet referrals resulted in outpatient visits at PH, compared with 79% in the outpatient clinic at HH. Only 18% of the patients receiving a teleconsultation ended up in the outpatient department of PH within one year. These visits were mainly due to progression of chronic disease. No deaths or missed diagnoses could be attributed to telemedicine, but one diagnosis was delayed. The direct costs of an outpatient clinic visit in internal medicine (EU211) were seven times greater per patient than for an e-mail consultation (EU32), with a marginal cost decrease of EU179 for every new intranet consultation. A cost-minimization analysis of the alternative interventions showed a net benefit of EU7876 in favour of the teleconsultation process. General practitioners sought an outpatient visit for 130 of their patients, and advice only for another 77. On-line advice was nonetheless given in 108 cases, and only 88 patient visits were arranged. Eleven referrals were declined. The cost difference between giving on-line medical advice for the 108 cases and a visit to the outpatient clinic for the other 88 was less costly (by EU4140) than investigating the 124 patients whose original clinic referrals to the PH were not declined. Productivity in the hospital increased over threefold by using e-mail consultations instead of traditional outpatient visits. The wide interactive use of the intranet referral system between secondary and primary care improved clinical effectiveness, lowered direct costs, increased productivity and was cost-effective. PMID- 11265101 TI - A follow-up study of remote trauma teleconsultations. AB - We conducted a follow-up study of patients who had attended a nurse-led minor accident and treatment service (MATS) and who had participated in a teleconsultation. Over three and a half years, 31,510 patients had attended the MATS unit and 1854 patients (5.9%) of these had participated in a teleconsultation. Of the 1854 telemedicine patients, 1199 had been referred to hospital or clinic and 1153 had actually attended. Retrospective examination of the relevant hospital records showed that in 25 cases (2%) the original telediagnosis was considered incorrect at face-to-face review and that treatment was either begun or changed in 264 cases (23%). All patients, including those discharged home after the teleconsultation, were sent a questionnaire about any changes to their injury. Of the 655 patients discharged home, a questionnaire response was obtained from 598 (91%). Following discharge, 43 of these patients had sought help from another health-care provider (the majority from their general practitioner). Of the 46 patients referred to hospital who did not keep their follow-up appointments, questionnaire results were obtained from 35 (76%). Nine of these patients had sought help from another health-care provider (the majority from their general practitioner) but there had been no change in diagnosis or treatment. Our findings suggest that teleconsultations are an effective means of delivering minor injuries care. PMID- 11265102 TI - The diagnostic acceptability of low-bandwidth transmission for tele-ultrasound. AB - Ultrasound recordings were made of 100 consecutive patients attending for obstetric examination in Peterhead and 100 patients attending for non-obstetric examination in Aberdeen. Two identical video-conferencing machines were used to transmit and receive the original ultrasound images at data rates of 384 kbit/s and 128 kbit/s, thus producing a total of three tapes for each case. Four experienced observers, who were blinded to the transmission bandwidth, each viewed 300 examinations and decided whether the images were acceptable or not for diagnosis. Almost 100% of the obstetric ultrasound images on the original recordings were considered diagnostically acceptable, compared with 93% of the 384 kbit/s transmissions and 44% of the 128 kbit/s transmissions. Similarly, 99% of the non-obstetric ultrasound images were considered acceptable, compared with 87% of the 384 kbit/s transmissions and 21% of the 128 kbit/s transmissions. For the obstetric ultrasound images the intra-observer diagnostic agreement was 93% (kappa = 0.89) between the original and the 384 kbit/s transmissions, and 78% (kappa = 0.63) between the original and the 128 kbit/s transmissions. For the non obstetric ultrasound images the respective intra-observer diagnostic agreements were 77% (kappa = 0.62) and 78% (kappa = 0.63). The quality of dynamic ultrasound images transmitted at 384 kbit/s was diagnostically acceptable, but was unsatisfactory at 128 kbit/s. PMID- 11265103 TI - Remote access to an expert system for infectious diseases. AB - An expert-system antibiotic information database was developed in order to help non-specialist doctors to choose the appropriate treatment for patients with infectious diseases. Fifty doctors conducted a pilot trial of the database, using modern access and the telephone network. During an eight-month study period, 1053 queries were received. The range of duration of the queries was 130-350 s. Of the queries, 473 (45%) were for particular patients with an infectious disease. The response rate to a questionnaire mailed out to the users at the end of the project was 100%. All doctors, even those who had limited experience with computers, found it easy to understand and to use the database. PMID- 11265104 TI - Telemedicine techniques can be used to facilitate the conduct of multicentre trials. AB - A multicentre randomized controlled trial was established in Pretoria, Bloemfontein and Edendale in South Africa, and coordinated from London. The purpose of the trial was to determine the efficacy of low-dose beta irradiation of glaucoma. Five communication modalities (telephone, fax, e-mail, videoconferencing and face-to-face meetings) were examined in terms of their benefits in a multicentre trial. The eight stages of the multicentre trial examined were: set-up and training, recruitment, standardization, patient management, data transmission, update and data dissemination, clinical follow-up and monitoring, and publication. On four-point Likert scales for rating the usefulness of the communication modalities at each of the eight stages of the trial (from 0 = not useful to 3 = very useful; maximum score 24) the telephone was given a total score of 10, fax 9, e-mail 13, videoconferencing 15 and face-to face meetings 9. Telemedicine techniques offer considerable benefits in the coordination of multicentre trials by improving data collection, maintaining the efficacy and monitoring of trials, while potentially offering reduced costs in terms of travel and time. The realtime scrutiny of patient records helps to ensure data uniformity and completeness of data collection. Videoconferencing was most useful when considered as one of several communication tools that can be used to improve the effectiveness of a service or process. PMID- 11265105 TI - Telemedical management of an odontoid peg fracture in the Shetland Isles. PMID- 11265106 TI - Guidelines for using videoconferencing in mental health services. PMID- 11265107 TI - Australian National Telehealth Think Tank. PMID- 11265108 TI - [Roles of Toll-like receptors and associated MD molecules in innate recognition of lipopolysaccharides]. PMID- 11265109 TI - [The mechanism and regulation of Fas-mediated apoptosis]. PMID- 11265110 TI - [The structure and physiological function of bHLH-PAS transcription factors]. PMID- 11265111 TI - [Regulation of glutathione transferase expression and function in stress response]. PMID- 11265112 TI - [Three families of phospholipase A2 inhibitory proteins derived from the blood of venomous snakes]. PMID- 11265113 TI - [Apoptosis-associated alteration of cell surface composion and its possible implication in immune regulation]. PMID- 11265114 TI - [Multifunctional peptide, adrenomedullin]. PMID- 11265115 TI - [Genetic and biochemical studies on the regulatory mechanism of the self resistance and biosynthesis of antibiotics]. AB - The following topics are described: 1. chemistry of beta-lactamases; 2. beta lactamases from Streptomyces including distribution of beta-lactamases in actinobacteria, properties of beta-lactamases from Streptomyces, cloning and regulatory mechanism of expression of beta-lactamase genes from Streptomyces and evolution and classification of beta-lactamases in general; 3. penicillin-binding proteins from Streptomyces including beta-lactam-producing- and non-producing strains and 4. eukaryotic-type protein kinases from Streptomyces including cloning of the genes and evolution and classification of eukaryotic-type protein kinases in general. PMID- 11265116 TI - Investigation of patients' demand for community pharmacies: relationship between pharmacy services and patient satisfaction. AB - We performed an investigation on the patients' demand for community pharmacy based on the analysis of questionnaire responses on community pharmacy services from the patients at 32 pharmacies in Tokyo and Osaka. In the previous study, we developed seven evaluation indices for pharmacy services, and showed that the functions most sought by patients in the "ideal pharmacies" were "Attitude of pharmacy/pharmacist", "Convenient hours" and "Information management". The objective of this study is to determine the relationship between these pharmacy functions and patient satisfaction by analyzing responses from the same questionnaire survey. Overall satisfaction score with the "pharmacy used today" was employed as the dependent variable, while the six factors derived from the 26 item evaluation scale in the questionnaire by factor analysis were used as the independent variables. As a result of analysis, it was found that four variables had a significant positive correlation with patient satisfaction, one had a significant inverse correlation, and one showed no significant correlation (p < 0.05). These results suggest that: attitude of the pharmacists such as general attitude and specialized activities of pharmacy/pharmacist such as providing information and explanations, and convenience of hours are not only judged to be important by patients, but also influence their satisfaction; comfortable facilities and availability of OTC drugs, while rated relatively low by patients in terms of importance, do influence their satisfaction; and convenience of location does not influence patient satisfaction. It was also indicated that insufficient inventories of prescribed medications have an impact upon patient satisfaction. This investigation offers evidence to provide patient-based pharmacy services. PMID- 11265117 TI - [Improvement of method to estimate guidance by pharmacists and trial to obtain standard pharmaceutical management and guidance services program]. AB - The level of understanding of taking medicine was examined at the start and the end of medicine-taking guidance, based on the estimation by pharmacists and inpatients. Six items for the estimation, such as "the way of taking medicines" were evaluated using a five-grade system. The significant improvement was observed in all items, suggesting that inpatients' understanding is improved by the guidance. A markedly positive correlation was found between the estimation by inpatients and that by pharmacists (p < 0.001). This confirms the appropriateness of estimation by pharmacists. Inpatients whose estimation was significantly different from that by pharmacists (difference of two grades or more) were examined to identify clear and potential problems with taking medicine. The problems were classified into such seven categories as "types of medicine". For each item with differences in estimation (two grades or more), problems were biased in some specific categories. On the basis of such a bias, a flow chart was prepared to clarify problems. In addition, a standard pharmaceutical management and guidance services program was drafted, in which measures in observation, treatment and education against problems by pharmacists were described according to the frequency of occurrence. Information about the program available on the Internet enables its use by other hospitals. PMID- 11265118 TI - [Effect of mouth wash on the removing fluticasone propionate delivered by dry powder inhaler in mouth]. AB - The effect of mouth wash on the removal of drug residues in both mouth and pharynx after the use of fluticasone propionate dry powder inhaler (FP-DPI) was studied. The concentration of FP in mouth wash after sprinkle and inhalation of Flutide 50, 100, 200 Rotadisk was determined by HPLC-UV. The total amount of the removed FP was measured by the sum of the concentration of FP in 5 times of mouth washes. The mouth wash procedures removed totally 79.3 +/- 4.4% (50 micrograms), 68.5 +/- 3.6% (100 micrograms), 69.3 +/- 3.4% (200 micrograms) of sprinkled amount of FP and 29.5 +/- 11.1% (50 micrograms), 35.6 +/- 6.6% (100 micrograms), 31.6 +/- 8.3% (200 micrograms) of inhaled, respectively. It was required for the removal of 90% of the totally recovered FP to do two times of mouth washes in each case. These data suggest that the mouth wash is an effective precaution for candidiasis induced by FP delivered by DPI. PMID- 11265120 TI - [Medical education for students in compulsory education: the conception of the preparation of three graded textbooks and preliminary evaluation of lectures using these textbooks]. AB - The school pharmacist in our hospital pharmacy used three graded textbooks about medicine for students at the Sukagawa School for the Health-Impaired (Fukushima Medical University Hospital Branch (H. I. school)). A revised textbook for 4th 6th grade elementary school students containing 12 important items of information about medicine, a new picture textbook for 1st-3rd grade elementary school students, and a new textbook containing practical data for junior high school students were prepared by supplementing original information with illustrations, simplified expression and large type face. Additionally, the pronunciation of Chinese characters was included in the textbook for the 1st-3rd grade elementary school students. In this study, 9 students from H. I. school and 37 students from Koyase junior high school took part in a questionnaire and an examination evaluating the usefulness of the lectures, and these textbooks, in regard to the student's recognition and understanding of medicine. Most students answered that the lectures and textbooks helped them to understand medicine. Furthermore, the results of the examination indicated that the students had a general understanding of medicine. In conclusion, we suggest that it is important for students in compulsory education to learn about medicine, and that according to the preliminary result of questionnaires and examinations, both the lectures and textbooks were useful to help the students to understand more about medicine. PMID- 11265119 TI - In vitro and in vivo evaluation of sustained release chitosan-coated ketoprofen microparticles. AB - Simple ketoprofen microspheres (MS) were prepared by the dry-in-oil method using ethylcellulose (EC) as a matrix polymer. Further, the microspheres modified by addition of polyethylene glycol (PEG) and hydroxypropyl cellulose (HPC), called MS-P and MS-H, respectively, were prepared. The in vitro release from MS, MS-P and MS-H was examined in the JP XIII second fluid, pH 6.8, at 37 degrees C and 60 rpm. Chitosan-coated ketoprofen microparticles (Chi-MP) were prepared by the precipitation of droplets of chitosan solution containing MS, and their adhesion to the rat small intestinal mucosa was tested. The plasma concentrations after duodenal administration were investigated for ketoprofen powder suspension, MS and Chi-MP. The particle size was raised with the increase in amount of ketoprofen added. The drug content and addition of PEG or HPC affected the drug release rate. The microspheres with moderate drug content, prepared by addition of modest amount of PEG, exhibited better gradual drug release. Chi-MP showed a good mucoadhesion. The maximum plasma concentration of ketoprofen for Chi-MP was less than one-third of that for ketoprofen powder suspension. Chi-MP tended to show the higher and steadier plasma level than MS. PMID- 11265122 TI - [Appearance of the hepatic artery in Multidetector-row CT using a test bolus injection]. AB - To separate the hepatic artery from the portal vein for the clear appearance of the hepatic artery, we used a test bolus injection in Multidetector-row CT. We examined 25 patients using the following method: 75 ml Iopamidol (370 mgI/ml), at a rate of 5 ml/sec, with scanning commencing 5 sec after the contrast medium reached the upper abdominal aorta. Successful separation could be obtained in almost all of the patients (88%). Separating the hepatic artery from the portal vein enabled us to evaluate the anatomy of the hepatic artery and the abnormality of blood flow in the liver. PMID- 11265121 TI - [No relation between angiotensin-converting enzyme (ACE) inhibitor-induced cough and ACE gene polymorphism, plasma bradykinin, substance P and ACE inhibitor concentration in Japanese patients]. AB - Persistent dry cough is well known as the most common side-effect of angiotensin converting enzyme (ACE) inhibitors. We examined the relationship between a cough and ACE gene polymorphism, plasma bradykinin (BK), substance P (SP) and ACE inhibitor concentrations in patients with hypertension or chronic nephritis. First, ACE genotyping was carried out in 96 patients, 42 with coughs and 54 without coughs, which had been treated with various kinds of ACE inhibitors. However, no significant difference in the ACE genotypes was observed between the two groups. Second, the plasma concentrations of BK, SP and ACE inhibitor were measured in 12 patients, which were treated with trandolapril at a daily dose of 1 mg for 4-33 weeks. In 3 patients, the cough was induced during the trandolapril therapy, while it was induced not in 9 patients. The plasma levels of BK and SP did not significantly change after trandolapril administration in the patients with and without coughs. Between the two groups, there were no significant differences in the plasma levels of BK and SP either before or after the trandolapril therapy. Also the plasma concentrations of trandolapril and trandolaprilat, the active metabolite of trandolapril, did not significantly differ between the two groups. These results suggest that there is no significant relationship between the ACE inhibitor-induced cough and ACE gene polymorphism, plasma BK, SP and ACE inhibitor concentrations in patients with hypertension or chronic nephritis. PMID- 11265123 TI - [Development of three-dimensional stereo viewer for high-resolution data]. AB - In order to visualize high-resolution three-dimensional (3D) data as a stereogram, a real-time volume-rendering system using a hardware graphic board and conventional PC was developed. A 256(3) data set could be visualized at a redrawing rate of 12 Hz, and a 512(3) data set at a rate of 2.5 Hz. It was demonstrated that stereogram visualization using volume graphic hardware architecture potentially enables rapid examination of high-resolution 3D data by changing visualization parameters such as level, window, transfer function for opacity, and color map or coordinate direction. PMID- 11265124 TI - [Multislice CT of the abdomen]. AB - With the advent of multislice CT, temporal and spatial resolution has been dramatically improved, enabling multi-phase dynamic CT of the abdomen with very thin slices. The image quality of multiplanar reconstruction (MPR) and CT angiography (CTA) has been markedly improved. These high-quality dynamic CT, MPR, and CTA images are an advantage of multislice CT of the abdomen. The CT protocol is very important in obtaining such high-quality images, and knowledge of the relation between the contrast medium and enhancement of each organ is essential. In this article, we present our multislice CT protocol for the abdomen as well as the results of our investigation using time-density-curve analysis. The clinical usefulness of MPR images and CTA of the abdomen are also discussed. PMID- 11265125 TI - [Gynecologic and obstetric disorders presenting with abdominal pain]. AB - This article briefly reviews the clinical settings and imaging findings of gynecologic/obstetric disorders presenting with abdominal pain. MR imaging is considered to be an excellent modality for the diagnosis of these disorders because of its high sensitivity to blood. Although CT allows images to be obtained within a shorter time than does MR imaging, the possibility of pregnancy should be excluded prior to the examination. Hemorrhagic ovarian cyst exhibits hyperintensity on T1-weighted images or hematocrit effect on CT or MR images. Hemoperitoneum associated with hemorrhagic cyst strongly indicates rupture of the cyst. Rupture of endometrial cysts and dermoid cysts can cause acute chemical peritonitis. In ectopic pregnancy, findings of hematosalpinx associated with strong enhancement of the tubal wall frequently contribute to the diagnosis. Torsion of the adnexa can be diagnosed by the pedicle between the ovary and uterus, and the lack of contrast enhancement. Tuboovarian abscess is recognized as a cystic mass having a thick wall that is strongly enhanced. Among complications of uterine leiomyoma, red degeneration shows characteristic MR findings, hyperintense rim on T1-WI, hypointense rim on T2-WI, and lack of contrast enhancement. Because of the high incidence of OHSS associated with normal pregnancy, CT is contraindicated. PMID- 11265126 TI - [Dynamic MR imaging of liver lesions with superparamagnetic iron oxide (SH-U 555A)]. AB - Dynamic MRI with SH-U-555 (SPIO) was evaluated. Dynamic MRI was performed for 17 patients with 22 lesions. Dynamic study with T2*-weighted imaging (T2* dynamic) and T1-weighted imaging (T1 dynamic) were performed in 8 cases (10 lesions) and 9 cases (12 lesions), respectively. T2* dynamic MR images were obtained before and 30, 90, 180, 270, 360, and 450 seconds and 31 minutes after the intravenous injection of SPIO, and T1 dynamic MR images were obtained before and 0, 40, 80, 120, 180, 240, 300, 360, 420, and 480 seconds and 28 minutes after the injection of SPIO. The signal intensity of each lesion was measured before and after the injection of SPIO. The enhancement ratio of the lesions was calculated and evaluated. The enhancement ratio of hypervascular lesions decreased rapidly in the first phase of T2* dynamic MRI, while that of hypovascular lesions decreased gradually. The enhancement ratio of hypervascular lesions increased in the first phase of T1 dynamic MRI and decreased gradually, while that of hypovascular lesions lacked the increase in the first phase, in contrast to hypervascular lesions. However, the changing of signal intensity could not be recognized on images with T2* dynamic and T1 dynamic study. In conclusion, quantitative analysis using the enhancement ratio made it possible to anticipate lesion vascularity. PMID- 11265127 TI - [Radiation therapy for brain metastases from breast cancer by histological classification]. AB - One hundred thirteen patients with metastatic brain tumor from breast cancer who were treated with external irradiation between 1989 and 1997 at Cancer Institute Hospital were studied. The patients were all histopathologically proven to have invasive ductal carcinoma (scirrhous type 54 cases, papillotubular type 18, solid tubular type 41). The patients were evaluated for efficacy and histopathological subtypes. The time interval between the diagnosis of breast cancer and brain metastases was 53.6 months for the scirrhous type, 75.0 months for the papillotubular type, and 35.5 months for the solid-tubular type. The time interval between the diagnosis of initial distant metastases and brain metastases was 14.3 months for the scirrhous type, 22.5 months for the papillotubular type, and 12.5 months for the solid-tubular type. Efficacy rates (CR + PR) for external irradiation of the brain metastases were 40.0% for the scirrhous type, 66.7% for the papillotubular type, and 36.6% for the solid-tubular type. The papillotubular type had a favorable efficacy rate compared with the other two types. Median survival time (MST) from the start of treatment for brain metastases and one-year survival rate were 5 months and 11.1% for the scirrhous type, 7 months and 41.5% for the papillotubular type, and 4 months and 28.3% for the solid-tubular type, respectively. No statistically significant difference between survival rates was observed among the histopathological types. Univariate analysis showed performance status, number of metastatic tumors, and existence of extracranial metastases without bony metastasis to be significantly related to prognosis. Multivariate analysis showed only extracranial metastases without bony metastases to be related to prognosis. PMID- 11265128 TI - [CT-guided needle biopsy of the lung: factors affecting risk of complications]. AB - PURPOSE: To analyze factors influencing the risk of complications associated with CT-guided percutaneous needle biopsy for lung lesions. MATERIALS AND METHODS: Sixty patients, aged 24-85 years (37 men and 23 women), underwent CT-guided needle biopsy. A definite diagnosis was made in 49 of 60 cases (81.7%), including 38 of 43 malignant lesions (88.4%) and 11 of 17 benign lesions (64.7%). Complications associated with biopsy were observed in 35 patients (58.3%). Major complications included pneumothorax (n = 26) and pulmonary hemorrhage (n = 20). Chest tube placement was needed in 5 (19.2%) of 26 pneumothorax cases (8.3% of all biopsies). RESULTS: The high frequency of pneumothorax (43.3%) in this series had several contributing factors, including the presence of pulmonary emphysema, lesion size, and traversal of aerated lung. Chest tube replacement was necessary more frequently in patients with pulmonary emphysema. The number of pleural passes, location of lesions, and size of needles were not correlated with the incidence of pneumothorax. CONCLUSION: The presence of pulmonary emphysema, lesion size, and traversal of aerated lung are the predominant risk factors for pneumothorax in patients with CT-guided lung biopsy. PMID- 11265130 TI - [Special categories of elderly patients with intractable neurological diseases (NANBYO) in Toyama, Japan]. AB - The purpose of this study was to elucidate special categories of elderly patients with intractable neurological diseases (NANBYO) in Toyama, Japan. This goal was accomplished through comparisons of data gathered in 1991 and 1997. Forty-eight (20 males, 28 females) with such diseases were recruited for the study and agreed to be interviewed. The surveys were performed by three public health center nurses. The same questionnaire was used in both years. The surveys show that the number of jobless patients and those who needed total support increased by approximately 20% from 1991 to 1997. In addition, it appears that the ratio of hospitalized patients and those who needed medical equipment installed at home increased considerably. The number of patients with caregivers increased by approximately 30%. As the caregivers got older, it was observed that the patients became more anxious about their health condition. Also, in both years, both patients and caregivers showed the highest concern about a sudden change in the patients' conditions. In conclusion, an assessment of patient and family needs is important for the development of a community health care system. It is also essential for all the parties in the field of health, medicine, and welfare, as well as volunteer groups, to have a good understanding of each other's services and to have a cooperative relationship coordinated by health centers through discussion of each individual case. PMID- 11265129 TI - [Selenium deficiency and brain functions: the significance for methylmercury toxicity]. AB - Selenium has been long recognized as one of the essential trace elements. Although many selenoproteins have been identified in the last decade, the physiological roles of Se and selenoproteins remain to be elucidated. Since iodothyronine deiodinases (DIs), which regulate the tissue levels of thyroid hormone, are (likely to be) selenoproteins, Se might have specific roles for developing brain. In fact, when rodents are depleted of Se perinatally, the thyroid hormone economy of the fetus is disturbed, which may lead to the abnormal development of the brain and to the abnormal postnatal behavior observed in Se deficient animals. When the animals were depleted of Se after weaning, when the role of thyroid hormone on brain development is minimal, neurochemical and neurophysiological alterations were found in the dopaminergic system. These postnatally-depleted rodents also showed abnormal open-field behavior, which was distinct from that observed with perinatally-depleted animals. The molecular events that convert Se-deficient status to these neurochemical, neurophysiological, and behavioral functions are largely unknown, and need to be further examined. The interaction between Se and mercury compounds has also been the focus of many research, but there have been few reports on the interaction between the physiological (nutritional) level of Se and the toxicity of prenatal methylmercury (MeHg). Experimental findings showed that Se-deficient rodents are more susceptible to the prenatal toxicity of MeHg. It is noteworthy that MeHg specifically altered the metabolism of Se in fetal/neonatal brain. Significance of the alteration of the activities of selenoenzymes such as glutathione peroxidase and DIs in animals by prenatal MeHg exposure are discussed in relation to the neurobehavioral toxicity of MeHg. PMID- 11265131 TI - [Structural analysis for psychosocial factors including health locus of control (HLC) and sense of coherence (SOC) associated with lifestyle-related diseases]. AB - The purpose of this study was to clarify psychosocial characteristics associated with preventive health behavior for lifestyle-related diseases. The author performed objective health examinations and gave questionnaires to 289 men (39.7 +/- 11.8 years, mean +/- SD) and 80 women (32.8 +/- 10.4 years) engaged in office work. Psychosocial factors included lifestyle and perceived stress, as well as the health locus of control (HLC) and sense of coherence (SOC) as newly developed indicators for health behavior. The principal component analysis for men did not extract lifestyle from the psychosocial structures. Multiple regression analysis showed that internal HLC (IHLC), chance HLC (CHLC), professional HLC (PHLC) and stress significantly contributed to SOC. Principal component analysis using psychosocial factors in women showed two psychosocial structures, i.e. the second principal (high SOC, high lifestyle, and low stress) and the 4th principal components (high supernatural HLC, and high PHLC). Both components were negatively correlated with systolic blood pressure. SOC was recognized to be negatively associated with age, stress, and total cholesterol, and positively with IHLC, FHLC, lifestyle, and gamma-GTP using multiple regression analysis for women. These results indicated a distinguishable sex difference regarding the involvement of psychosocial factors including HLC and SOC in objective health. SOC seems likely to be involved not in objective health, but closely with stress, suggesting a direct influence on mental health. Lifestyle should be divided into more detailed categories such as smoking and salt intake. Structural analysis of women suggests that SOC is involved directly or indirectly through lifestyle in objective health, different from men. To further clarify causal relationships between psychosocial factors and risk factors for lifestyle-related diseases, a longitudinal study is necessary based on these results. PMID- 11265132 TI - [Effects of ethanol on the nervous and vascular systems: the mechanisms of alcohol-induced hypertension]. AB - Many previous experimental and epidemiological studies have shown that alcohol consumption has a positive correlation with the incidence of hypertension. The effects of ethanol on the nervous and vascular systems in relation to the mechanisms of alcohol-induced hypertension proposed so far are reviewed here. Alcohol ingestion influences many pathophysiological functions which regulate blood pressure, as follows: 1) Sympathetic nervous activity is increased after drinking. 2) Ethanol acts directly on the contractility of vascular smooth muscle. Ethanol acutely contracts some arteries and increases their contractile responses to agonists, while it also displays inhibitory effects on vasocontractility in other arteries. Thus, ethanol has two opposite actions, both of which depend on the kinds of vessels and animal species used for the experiments. Intra- and extracellular Ca2+ mobilization and activation of the contractile apparatus have been suggested as mechanisms for ethanol's vasocontractile actions. 3) Chronic alcohol ingestion has been reported to induce a deficiency of blood and intracellular magnesium, which influences cellular Ca2+ homeostasis through attenuation of plasmalemmal ATPase activity. Direct alcohol effects on cardiovascular systems may not be involved in hypertension that develops after long-term habitual drinking. 4) Ethanol affects vascular endothelial functions, inhibiting endothelial NO- and EDHF-dependent vasorelaxations. 5) The serum levels of vasoactive substances such as cathecolamines, renin-aldosterone, prostacyclin, and endothelin have been reported to be affected by alcohol ingestion or ethanol in vitro. 6) In heavy drinkers, alcohol withdrawal results in an elevation of blood pressure due to sympathetic nervous stimulation. 7) Long-term heavy drinking often results in the development of insulin resistance and glucose intolerance, which in turn triggers hypertension. 8) The difference in the genetic polymorphism of acetaldehyde dehydrogenase among Japanese people may not be directly related to development of alcohol-induced hypertension. As mentioned above, alcohol shows multiple actions on various factors regulating blood pressure. More detailed and integrated mechanisms for alcohol-induced hypertension, which is not a homogeneous disease, remain to be clarified. PMID- 11265133 TI - [Effect of exercise habits and lifestyles on changes in physical fitness in medical college students: a 3-year follow-up study]. AB - This follow-up study investigated the relationship between changes in physical fitness and exercise habits during the 1st and 4th years in healthy medical college students. We also investigated the relation between exercise habits and the lifestyle factors during the same period of time. The subjects were 229 male and 126 female student volunteers who received physical fitness tests and questionnaires (lifestyles and exercise habits) in both the 1st and 4th years. The results were as follows: 1. A significant association between increment in physical fitness and exercise habits was shown by the fitness score in the 1st year, and the intensity and frequency of exercise in the 4th year in males. In females, an association was found in the intensity of exercise during the 1st year, and frequency of exercise during the 4th year. The intensity in 4th year among males (odds ratio(OR): 2.80, 95% confidence interval (CI): 1.33-5.87) and the frequency during the 4th year among females (OR: 5.63, 95% CI: 2.36-13.43) were associated with a higher odds ratios for improvement in physical fitness than other factors related to exercise habits in 4th year medical students. 2. In the association between exercise habits related to increment in physical fitness and other lifestyles, a significant association was admitted between the intensity of exercise and the frequency of eating breakfast during the 4th year in males (OR: 2.18, 95% CI: 1.11-4.28). These results suggested that the intensity of exercise during the 4th year in males and the frequency of exercise during the 4th year in females were associated with increments in physical fitness for follow-up. A lifestyle factor related to the intensity in exercise habits has been suggested to be the frequency of eating breakfast (more than 5 days/week) in males. PMID- 11265134 TI - [Resource oriented interventions in pregnancy]. AB - Premature labour and premature birth keep on being an unsolved problem in spite of diverse medical measures in the obstetrics. The system of risk catalogues did not meet the expectations and can lead even for the disconcertion from pregnant women. If one places the risks performed in the German mother passport as a basis, approximately 50% of all pregnant women would be risk cases. In current publications stress finds strong attention as a psychophysiological declaration model of premature birth. Own results refer to psychosomatic connections at premature birth. Social relief and here particularly the one through the babies father seems to be a preventive factor with regard to some premature birth. Under a resource-oriented visual angle the salutogenetic-model of Antonovsky is employed for care concepts of the primary and second prevention in the pregnancy as a basis. Specific target group offers for pregnant woman that focus also on the stabilization of their competence feeling, particularly next to knowledge transfer, relaxation training and learning of coping strategies, the activation of social nets and, where appropriate, early inclusion of the partner and offer of communication training for pregnant couples are described as a good complement to traditional prenatal care. A bearing away of the risk-model opens the resources-model for prenatal care new ways and is a challenge also for science. PMID- 11265135 TI - [Legal problems in an elective cesarean section]. AB - The requirements for a legal assessment of the Caesarean section on request can be deduced from the general criteria for the legitimacy of medical treatment: indication of treatment, adequacy of treatment (= quality) and informed consent. The problem of indication is of particular relevance under the aspect of criminal law. Even if one takes the view that a Caesarean section on request is an operation without indication this does not automatically amount to unconscionability as defined by sect. 226 a StGB (Criminal Code) since such an operation may be justified by the consent of the woman. This leads to the problem of comprehensive information on the risks of such an operation. Moreover, it appears to be questionable to reject the indication of an Caesarean section on request. The information requirements with reference to liability law can only be formulated with the background of the general information requirements for methods of delivery. The general principle that the greater the risk of operation and the more difficult the decision to be taken are, the more intensive should be the information provided, results in comprehensive duties of information in the case of the Caesarean section on request. At present, court rulings establish natural birth to be the first choice of treatment. However, if the risk/benefit assessment should lead to new results due to new medical findings in the future, this will require some discourse within the profession and possibly a new establishment of the medical standard. With regard to the professional aspect, the doctor in charge is entitled to refuse a Caesarean section on request. PMID- 11265136 TI - [Improved diagnosis through digital mammography with the assistance of computer algorithms]. AB - OBJECTIVE: The aim of this survey was to determine improved diagnostics through digital mammography with assistance of computer algorithms and new investigation techniques. MATERIAL AND METHODS: In general, the presented procedures are practicable with all digital mammography procedures. In our clinic the largest clinical experience exists with the only FDA certified full field mammography device Senograph 2000D (GE Medical Systems). RESULTS: First results from initial studies are shown, which are to evaluate the new procedures. The so-called post processing with visualization of microcalcifications, computer-assisted diagnosis, tomosynthesis, energy subtraction, contrast media in mammography and teleradiology are considered in particular. CONCLUSION: Digital mammography represents a promising procedure, opening new possibilities for improved diagnostic in mammography. PMID- 11265137 TI - Grandmothering and reproductive ageing--shifting the reproductive period. PMID- 11265138 TI - [Differential diagnosis of thrombocytopenia in pregnancy]. AB - Thrombocytopenia (< 150,000/microliter) is a common finding, occurring in 7-8% of pregnancies. Some conditions, such as gestational thrombocytopenia pose no maternal or fetal risks. Idiopathic thrombocytopenic purpura (ITP) is an acquired haematologic disorder, common among children and adults, with unknown etiology and autoimmune pathogenesis. The incidence of severe fetal and neonatal thrombocytopenia is very rare, and neonatal intracranial hemorrhage is unlikely to be related to the mode of delivery. Alloimmune thrombocytopenia occurs with an incidence of 1/1,000 livebirths and is induced by a maternal alloimmunization against fetal platelet antigens. The incidence of intracranial haemorrhage in the fetus and neonate is the highest for any immune thrombocytopenia. The HELLP syndrome is a severe, unpredictable and life-threatening complication of preeclampsia, characterized by a triad of hemolysis, elevated liver enzymes and low platelet counts. HELLP syndrome develops in the third trimester but can occur postpartum. Hemolytic uremic syndrome (HUS) and thrombotic thrombocytopenic purpura (TTP) are syndromes of microangiopathic hemolytic anemia, and thrombocytopenia. During pregnancy, TTP usually presents in the second trimester, whereas HUS develops in the postpartum period. Heparin-induced thrombocytopenia type II is a serious, immune-mediated complication of heparin therapy. PMID- 11265140 TI - When 'too much' is 'not enough'. PMID- 11265139 TI - Human papillomavirus (HPV) DNA in primary cervical cancer and in cancer free pelvic lymph nodes--correlation with clinico-pathological parameters and prognostic significance. AB - OBJECTIVE: To assess whether the presence of human papilloma virus (HPV) DNA and/or several genotypes of HPV DNA in primary cervical cancer and cancer free pelvic lymph nodes are correlated with several clinicopathological parameters of well-defined prognostic significance and whether virological parameters are predictors of long-term survival in cancer patients. PATIENTS AND METHODS: 223 cases of cervical cancer patients included in this retrospective study underwent follow-up evaluation. Survival and cause of death were examined for 204 (91.4%) patients, with a mean follow-up time of 4.4 years. HPV DNA was detected using the high sensitive polymerase chain reaction (PCR) method followed by HPV DNA sequencing for HPV genotyping. These results were correlated with well-defined clinicopathological parameters and survival data. RESULTS: HPV DNA was detected by PCR in 150 of 203 (73.4%) tissue specimens of cervical cancer patients. DNA sequence analysis revealed the presence of HPV 16 (n = 68, 45.3%), HPV 18 (n = 49, 32.6%) and rare HPV types (n = 33, 22.1%). HPV genotypes correlated significantly with histological tumor types, node status, blood vessel invasion and lymph space involvement. The presence of HPV DNA in cervical cancer as well as the genotype of HPV 16 could also be confirmed as significant prognostic factors in the univariate Cox Regression Analysis (RR 2.856, p < 0.003 resp. RR 3.444, p < 0.0001). The presence of HPV DNA in cancer free pelvic lymph nodes was significantly correlated to the concomitant manifestation of pelvic lymph node metastases (RR 3.1, p < 0.0001). In the multivariate analysis, however, HPV DNA in primary tumor and in negative pelvic lymph nodes failed to be of prognostic relevance. Exclusively, HPV 16 appears to impact independently on the overall survival in cervical cancer patients (RR 3.653, p < 0.002). CONCLUSION: The detection of HPV 16 genotype may play an important adjunct role in assessing prognosis of cervical cancer patients. The clinical impact of the presence of HPV DNA in primary tumors and cancer free pelvic lymph nodes remains to be investigated in further studies. The exact mechanisms by which HPV influence the prognosis of cervical cancer patients have to be defined. PMID- 11265141 TI - In medical negligence case, causation must be shown with a reasonable degree of medical certainty. PMID- 11265142 TI - The complete story about Medicare 'private contracts'. PMID- 11265144 TI - [Surgical therapy of carcinoma of the pancreas]. PMID- 11265143 TI - Percutaneous vertebroplasty: a new technique for treatment of painful compression fractures. AB - Vertebroplasty is an effective technique in which polymethyl methacrylate, a surgical cement, is injected into a vertebral body. This technique provides increased strength and pain relief in vertebrae weakened by osseous lesions as metastases, multiple myeloma, and aggressive hemangiomas, and from osteoporotic fractures. Infrequent serious complications are mainly related to polymethyl methacrylate leakage into the spinal canal and around vital soft tissue structures. Complications are decreased by careful fluoroscopic guidance and meticulous radiological technique. Indications should be dictated by a multidisciplinary team approach. PMID- 11265145 TI - [The role of laparoscopy in the identification of peritoneal carcinosis from abdominal neoplasms. Analysis of our initial experience]. AB - BACKGROUND: The authors reviewed the experience on the use of laparoscopy performed since January 1997 in malignant neoplasms at their institution. The aim of the study was to evaluate the real effectiveness of this procedure in the staging of abdominal neoplasms which were considered resectable at preoperative examinations and in particular in the detection of peritoneal metastases not evidenced with traditional imaging techniques. METHODS: Twenty-eight patients with malignant neoplasms: colo-rectum (15), stomach (5), pancreas (4), gallbladder (2), cardias (1), liver (1), were studied. All the patients were preoperatively examined with abdominal computed tomography (CT). In the 2 patients with gallbladder neoplasm a MR cholangiography was also performed. An explorative laparoscopy with peritoneal washing was then performed in all the patients. The diagnostic and therapeutic choices were subsequently done on the basis of laparoscopy results. RESULTS: Therapeutic approach was modified in 21% of cases, as a result of the detection of peritoneal metastases which were not evidenced with imaging examinations. On the contrary, peritoneal washing was not responsible of any preoperative evaluation. CONCLUSIONS: Laparoscopy performed in patients with abdominal neoplasms allows the detection of peritoneal micrometastases not previously evidenced through preoperative CT, thus modifying the therapeutic approach. PMID- 11265146 TI - [Histologic study of experimental spleen transplant in rats]. AB - BACKGROUND: The objective of this paper was to demonstrate that the grafts of cervical splenic transplantation on rats using our experimental model present a normal histological appearance. METHODS: Isogenic consanguineous Lewis rats 12 weeks old and weighing 250 gr. were used. Histological findings of a group of 25 cervical splenic grafts transplanted by means of splinting vascular venous microanastomoses and a group of 25 splenic grafts autotransplanted in the omentum were compared with a control group. The specimens were assigned according to a score of 0 to 4, following Moore's histological criteria. RESULTS: All grafts in transplanted and autotransplanted groups had a score of 3 or 4. Then, all splenic grafts from the transplanted group had histological findings very similar to a normal spleen. In the autotransplantation group, the percentage of grafts with a score 3 (60%) was superior to the transplantation group (46%). However, the transplantation group presented a percentage of score 4 (54%), superior to the autotransplantation group (40%). CONCLUSIONS: In our study all grafts from the cervical spleen transplantation group had histological findings very similar to a normal spleen. The percentage of spleens with histological normality in the transplantation group was superior to the autotransplantation group. However, there was no statistical significance. PMID- 11265147 TI - [The role of melatonin in the immediate postoperative period in elderly patients]. AB - BACKGROUND: Melatonin induces sleep and modulates immune system. Aim of the paper is to show a possible relation between impaired rhythm of melatonin secretion and the onset of postoperative septic complications and insomnia in old patients undergoing surgery. METHODS: Fifty old patients, aged from 60 to 94 years, have been studied; 39.5% of the patients had neoplastic disease. Melatonin serum levels have been evaluated by ELISA technique at 12 p.m., 3 a.m. 8 a.m. immediately before operation. Postoperative clinical findings of insomnia and septic complications have been recorded. RESULTS: The melatonin serum mean values of all the patients were 16.3 pg/ml at 12 p.m., 22.4 pg/ml at 3 a.m. and 7.1 pg/ml at 8 a.m. Neoplastic patients showed the higher values of melatonin (26.696 pg/ml, 33.143 pg/ml, 9.185 pg/ml), long-lived patients (> 90 years) the lower. The melatonin secretion curve of the old patients with postoperative insomnia (19.961 pg/ml, 20.297 pg/ml, 9.378 pg/ml) or postoperative septic complications (20.695 pg/ml, 16.183 pg/ml, 6.036 pg/ml) was significantly different compared with that of other patients. The peak was advanced, lower and decreased slowly in the midnight. CONCLUSIONS: The study seems to show a possible correlation between impaired rhythm of melatonin secretion and postoperative insomnia and postoperative sepsis in old patients undergoing surgery. PMID- 11265148 TI - [Surgical treatment of varicocele with inguinal microligation technique. 6-year experience]. AB - BACKGROUND: Varicocele is found approximately in 15% of the male population and is considered a major cause of infertility. Varicocele management include surgical (traditional or laparoscopic) or conservative techniques (sclerotherapy). The authors present their experience on microsurgical inguinal varicocelectomy. This technique has been adopted since 1992 to decrease the incidence of recidives of high spermatic vein ligation; it also permitted to use local or loco-regional anesthesia, reducing time of hospitalization and realizing a minimally invasive approach. METHODS: From 1992 to 1997, 433 microsurgical inguinal varicocelectomy with artery and lymphatic sparing have been performed at the Militar Hospital of Milan in 409 young men with idiopathic varicocele. All patients were discharged 24 hours after operation. Only those who lived particularly far from the hospital remained for 48 hours. RESULTS: Clinical controls were performed I, III, VI months after operation. At the third control (VI month), a new semen analysis was performed, and 65% of patients had an improvement of seminal characteristics. In 394 patients, a complete resolution of varicocele was observed; 4 patients had a recurrence of the pathology and 11 had a recidive. Seventy-three patients who presented a concomitant homolateral inguinal hernia were treated at the same time. CONCLUSIONS: The conclusion is drawn that microsurgical ligation of spermatic veins represents a good surgical option in the treatment of varicocele. It is a quite simple technique that guarantees a low risk of recidives, permits using local or loco-regional anesthesia and can be performed in day-surgery with good results, few complications and good short and long term results. PMID- 11265149 TI - [Transplant of livers from living relatives: selection of recipients and donors]. AB - Living relative liver transplantation is a valid alternative to cadaver transplantation especially at a time when the availability of organs cannot meet the requests of long waiting lists. This procedure was initially introduced in response to the shortage of organs for pediatric cases, but the rapid growth of demand for liver transplantation has led to its extension to the adult population. The procedure raises a number of ethical, logistic and technical questions. The ethical aspect has been widely debated and in order to be acceptable, the procedure must comply with three critical points: the need for innovation, an acceptable risk-benefit ratio and adequate informed consent. The technical aspect is essential for the success of the procedure. It calls for an extensive experience and know-how of hepatobiliary surgery on one hand, and the use of high-resolution vision on the other, an aspect which is crucial for the success of vascular anastomoses. The indications for living relative transplantation are the same as for standard transplants. The sole exception is for adult patients with 2A status who present advanced hepatic imbalance caused by chronic liver disease, thereby reducing the probability of success, above all because a living donor graft is always smaller compared to the ideal dimensions for the recipient. In view of the severe shortage of organs, living relative transplantation is an important alternative for both pediatric and adult patients. The challenge over the coming decades will be to extend living relative transplantation to a growing number of patients, without jeopardizing the health of the donor. PMID- 11265150 TI - [Surgery in the cirrhotic patient. Prognosis and risk factors]. AB - Patients with cirrhosis have reduced life expectancy. Surgery is often associated with clinical decompensation in this group of patients. The purpose of this paper is to study the surgical risk in cirrhotic patients undergoing nonderivative operations. Unfortunately, most of the studies in the literature about this problem are retrospective reviews with limitations. The conditions increasing surgical risk in cirrhotic patients are analysed. These include changes in the pharmacokinetics and pharmacodynamics of various drugs, altered hemostasis, poor resistance to infections, water retention, suture line insufficiency, chronic renal failure and congestive heart failure. Assessment of the disease stage in cirrhosis is very important, because the severity of hepatic abnormalities influences the prognosis. The Child-Pugh classification has been used extensively to risk-stratify patients with cirrhosis. However, the disregard for cardiorespiratory, renal, electrolyte balance and acid-base status limits its predictive accuracy. Recently a new scoring system, the Acute Physiology and Chronic Health Evaluation (APACHE III), has been introduced and seems to be superior to Child-Pugh for prognosticating short term survival of cirrhotic patients. In conclusion, surgery can be done safely only in cirrhotic patients with a good hepatic function. On the contrary, in patients with advanced cirrhosis, surgery causes a very high mortality. Finally, the patients with moderate hepatic failure can be operated only after a careful study of the disease and an adequate correction of the reversible risk factors. PMID- 11265151 TI - [Treatment of pain in the oncologic patient]. AB - Around 65-85% of cancer patients suffer from pain at advanced stages. Pain is often inadequately treated, although it can be controlled simply in the majority of cases. It is important to try and achieve a number of targets, including pain control at night, resting pain and pain during movement. Pain can be divided into somatic pain caused by the stimulation of traditional nociceptors, visceral pain and neuropathic pain caused by damaged nervous fibres. All three types may exist in the same patient. Drugs are the main method used to control oncological pain. The three main classes of drugs (FANS, opioid analgesics and adjuvant analgesics) are used individually or in combination. Given that the collateral effects of opioid analgesics may limit their value, they must be monitored to ensure careful treatment. The appropriate use of invasive treatment in patients with advanced disease who do not respond to oral therapy may alleviate cancer pain in 10-30% of cases. These adjuvant procedures are classified as blockades of autonomous nervous tissue, peripheral nerves and neuraxis. In conclusion, the ability to give an overall evaluation of a patient with pain, to ensure the component administration of analgesic drugs and to inform the patient and the family forms the basis of the treatment of pain in cancer. PMID- 11265152 TI - [Gastric metastasis from melanoma. Report of 2 surgically treated cases]. AB - Metastatic disease involving the gastrointestinal tract is a rare pathology and melanoma is the extra-intestinal neoplasm more frequently concerned. Two cases of gastric metastases, revealed by symptoms of upper gastrointestinal bleeding, are reported. In the first case the disease-free interval, after excision of the primary lesion located in the right lower limb, was 13 years; in the second case the primary lesion remained unknown, although it probably originated from a giant congenital nevus of the left foot. Both patients had been affected before by inguinal nodes metastases, treated by radical groin lymphadenectomy; the concomitant multiple metastases to other sites (adrenal glands, retro-peritoneum, liver, lung, small bowel, brain, ovaries) limited surgery to a likely prospect of palliation, conditioning an unfavourable prognosis. Resection of gastrointestinal metastases is justified for the relief of intestinal hemorrhage (as in these reported cases), perforation and obstruction, even if treatment of single non complicated lesions can have a curative intent. The conclusion is drawn that more aggressive diagnostic and staging procedures are indicated for the early detection of gastrointestinal metastases whenever non-specific abdominal symptoms and a history of melanoma are present. PMID- 11265153 TI - [Secondary costal myxoid chondrosarcoma. Presentation of a clinical case]. AB - A type of malignant neoplasm of not frequent observation is reported and the difficulties concerning the distinction from the benign counterpart are underlined, for which a detailed instrumental study is made necessary, integrated with pre and, above all, postoperative histologic analysis. A white male patient of 83, affected by plurirecidivant chondromyxoma of the rib, was admitted for local recurrence. The lesion objectively appeared not dissimilar from the preceding ones, as an ovalar mass fixed on the underlying plains. Radiologic and ultrasonographic examinations seemed to confirm the admission diagnosis; at the end the patient underwent a new intervention. Histologic examination of the lesion yielded a diagnosis of chondromyxosarcoma. The clinical case is made even more interesting by the finding, through the TC technique, preoperatively performed, of a metastatic repetition on the opposite side. The authors examine the problem of the early recognition of a malignant neoplasm developing, in a patient with a clinical history of recidival chondromyxoma. Physical examination and laboratory analysis are not useful in the assessment of the occurring transformation. At molecular level chondromyxosarcoma is characterized by several genomic rearrangements and mutations. Though primitive chondromyxosarcoma often involves the ribs, chondromyxoma's transformation into its malignant counterpart is not a common event. The development of distant metastasis characterizes long term cases. Given the lack of encouraging results about chemo and radio-therapy efficacy in influencing the natural course of the disease, a systematic approach will be made necessary in patient management. PMID- 11265154 TI - [Angiosarcoma of the breast after conservative surgery and radiotherapy for early breast carcinoma. Description of a case]. AB - The occurrence of an angiosarcoma of the residual breast after conservative surgery and adjuvant radiotherapy for early mammary carcinoma is a very rare event. In western countries only 57 cases have been published in the literature (5 in Italy) since the first described case in 1987. Radiotherapy seems to be the most important etiological factor in the development of the neoplasm. Diagnosis is often delayed, owing to the "benign" aspect of the lesion. The only effective treatment is residual mastectomy, because chemotherapy is ineffective. The prognosis is often dismal, because of the aggressive behaviour of the lesion in most cases. The case of a patient with a multicentric secondary angiosarcoma of the breast recently operated on is described. PMID- 11265156 TI - [Inflammatory pseudotumor of the lung. Diagnostic-therapeutic effectiveness of its radical resection]. AB - Clinical, therapeutical observations and experience in 3 cases of pulmonary inflammatory pseudotumors (PIP) are presented. A retrospective analysis is made of cases with pulmonary "mass" suspected as malignant tumor, resected in a general surgery department between 1988 and 1995, and finally diagnosed as inflammatory pseudotumor. Three of the 10 cases originally diagnosed as malignant lung tumor were inflammatory pseudotumor (30%). Pulmonary inflammatory pseudotumors, may be a pitfall diagnosing a lung mass and implicate legal problems. Surgical resection leads to the final diagnosis in doubtful cases. A wide resection has a diagnostic aim and may preserve healthy parenchyma. Clinicians, pathologists and surgeons should accurately inform patients with doubtful diagnosis of pulmonary malignancy. Any decision should be kept altogether either choosing the simple observation or the timely surgical diagnostic and therapeutical approach. PMID- 11265155 TI - [Association of deep venous thrombosis and popliteal pseudoaneurysm after arthroscopic meniscectomy]. AB - Popliteal pseudoaneurysm is an infrequent outcome of closed or penetrating injuries to the popliteal artery. An unusual case of iatrogenic right popliteal pseudoaneurysm in a female patient referred to our attention 20 days after arthroscopic meniscectomy is described. PMID- 11265157 TI - A medical sabbatical to England. PMID- 11265158 TI - James Mercer Green: southern gentleman, Confederate surgeon. PMID- 11265159 TI - Sojourn to Ethiopia. PMID- 11265161 TI - The newborn with hearing loss. PMID- 11265162 TI - Epidural hematoma associated with anticoagulant therapy. PMID- 11265160 TI - Stroke care and prevention in Georgia: current status and challenges. PMID- 11265163 TI - In vitro susceptibility of 137 Candida sp. isolates from HIV positive patients to several antifungal drugs. AB - Oropharyngeal candidiasis caused by various species of Candida is one of the most common infections in HIV seropositive or AIDS patients. Drug resistance among these yeasts is an increasing problem. We studied the frequency of resistance profile to fluconazole, itraconazole, ketoconazole, amphotericin B and terbinafine of 137 isolates of Candida sp. From HIV positive or AIDS patients with oropharyngeal candidiasis at Instituto de Inmunologia, U.C.V. and the Hospital "Jose Ignacio Baldo", Caracas Venezuela, using the well diffusion susceptibility test (Magaldi et al.). We found that nearly 10% of C. albicans isolates were primarily fluconazole resistant, 45% of C. albicans isolates from patients with previous treatment were resistant to fluconazole, of which 93% showed cross-resistance to itraconazole, and even about 30% of C. tropicalis (n = 13) were resistant to fluconazole and/or itraconazole. To this respect, several recent reports have been described antifungal cross-resistance among azoles. Therefore, we consider that C. tropicalis should be added to the growing list of yeast in which antifungal drug resistance is common. This report could be useful for therapeutic aspect in AIDS patients with oral candidiasis. PMID- 11265164 TI - Incidence of histoplasmin hypersensitivity in the Philippines. AB - Long-term residents of the Philippines were skin tested with histoplasmin skin test material. This study was conducted with 143 electric company (MERALCO) employees from Manila, Philippines. We found that 37 (26%) of the subjects were skin test positive. Characteristics of the positive group were: average age of 37 years; all except one were lifelong inhabitants of Metro Manila; 25 were male and 12 were female; one-half of the subjects reported extended contact with chickens. Despite these findings, histoplasmosis is considered to be a very rare disease in the Philippines. This survey indicates that Histoplasma capsulatum is sufficiently present in the Philippines to come in contact with one-fourth of the test population. This reinforces the hypothesis that histoplasmosis is present in the Philippines and is probably being misdiagnosed as granulomatous-inducing diseases such as tuberculosis, e.g., so-called "drug resistant" tuberculosis. We recommend larger surveys of this type and attempts to culture the etiologic agent from natural sources such as chicken and bat droppings. PMID- 11265165 TI - Incidence and severity of frogeye leaf spot and associated yield losses in soybeans in agroecological zone II of Zambia. AB - The incidence and severity of frogeye leaf spot of soybean (Glycine max (L.) Merr.) was studied in agroecological region II of Zambia during the 1997/98 crop growing season. A survey was conducted on farmers' fields on SCSI Kaleya, Magoye and Hernon-147 cultivars. Disease incidence and severity was assessed by monitoring disease increments at two weeks interval (beginning of January to April) from nine fields, three from each province. Soybean cultivars were evaluated for yield losses resulting from frogeye leaf spot. Field plots of each cultivar were either sprayed twice with benomyl (benlate) or not sprayed at all. The results showed that the incidence of frogeye leaf spot was highest in Southern province (5.1), followed by Lusaka province (4.9) while Central province had the lowest disease incidence (1.8). Values for area under disease progress curve (AUDPC) were significantly greater (P < 0.05) for Lusaka and Southern provinces than for Central province. Yields in benomyl protected plots ranged from 1444 kg ha-1 to 2320 kg ha-1 and were significantly different among the cultivars. Average yields of non protected plants were reduced by 30.5% for Kaleya, 35.6% for Hernon-147 and 37.2% for SCS1. Incidence and severity increased with time and varied depending on weather parameters and susceptibility of cultivars to the disease. Yield losses due to frogeye leaf spot occurred through a reduction in seed size. Differences in weather conditions and amount of inocula are believed to contribute to the observed variation in incidence and severity of the disease at different locations. PMID- 11265166 TI - Identification of deoxynivalenol, 3-acetyldeoxynivalenol and zearalenone in the galactose oxidase-producing fungus Dactylium dendroides. AB - The galactose oxidase-producing fungus Dactylium dendroides was re-identified as a Fusarium species. Fungi of this genus are well known for the production of mycotoxins. Verification of growth of this fungus on rice, corn and liquid medium described for the production of galactose oxidase is provided to determine whether the fungus could produce Fusarium toxins, namely, moniliformin, fusaric acid, fumonisin, zearalenone and the trichothecenes, deoxynivalenol, 3 acetyldeoxynivalenol, fusarenone, nivalenol, diacetoxyscirpenol, neosolaniol, and toxin T-2. Under the culture conditions used, deoxynivalenol, 3 acetyldeoxynivalenol and zearalenone were detected in the fungal culture medium. The finding is consistent with the hypothesis that the fungus is in fact a Fusarium species. PMID- 11265167 TI - Biodiversity and concentration of airborne fungi in a hospital environment. AB - The biodiversity and concentration of airborne fungi were monitored over a period of 6 months in a special-care unit of a hospital. Air sampling was performed in a corridor that was also accessible to visitors and in an adjacent bone-marrow transplantation (BMT) unit using an air sampler and two isolation media. Altogether, 98 fungal species could be identified, among them Aspergillus fumigatus and A. terreus as well as 48 other species reported as potential pathogens. The average contamination values of the corridor air ranged from 124 to 485 cfu m-3. Neither the degree of fungal air contamination nor the species composition inside the special care unit differed from those found in the corridor. By means of data obtained with a light-activated sensor, a possible influence of human activities on diurnal changes of fungal propagule concentration was shown. PMID- 11265168 TI - Ultrastructural changes in the muscles, midgut, hemopoietic organ, imaginal discs and Malpighian tubules of the mosquito Aedes aegypti larvae infected by the fungus Coelomomyces stegomyiae. AB - Fungi belonging to the genus Coelomomyces can infect mosquito larvae and develop within the larval hemocoel. To examine fungal development, Aedes aegypti larvae infected with Coelomomyces stegomyiae Keilin were fixed, embedded and sectioned for both light and electron microscopy. While fungal hyphae of C. stegomyiae did not invade cells other than the cuticular epithelial cells, they did penetrate a number of tissues including muscles, midgut, hemopoietic organ, imaginal discs, and Malpighian tubules. PMID- 11265169 TI - The Journal of Clinical Pathology goes online (http://www.jclinpath.com)! PMID- 11265170 TI - The causes and effects of fetal macrosomia in mothers with type 1 diabetes. PMID- 11265171 TI - Recommendations for HER2 testing in the UK. AB - Determining the HER2 status of breast carcinomas is a prerequisite for the use of the monoclonal antibody trastuzumab (Herceptin), which has recently been licensed for the treatment of metastatic disease. This necessitates a test based on archival material. The preferred analyses are immunohistochemistry with fluorescent in situ hybridisation (FISH) as a follow up test for ambiguous results. Guidelines have been developed for standardised, well controlled procedures for the provision of reliable results. A group of three reference laboratories has been established to provide advice, quality assurance, and materials, where needed. PMID- 11265172 TI - Down's syndrome screening: a controversial test, with more controversy to come! AB - By 1998, most health authorities offered antenatal screening for Down's syndrome, usually by biochemical methods. To date, the development of this form of screening has not been coordinated by a national body and, consequently, there are wide variations in practice between localities. Fortunately, many of these variations have not led to any noticeable inequality of health provision, but the wide variation in risk cut offs used by different centres does. Other variations merely lead to potentially unnecessary expenditure; whereas it is believed that adding extra tests to the screening procedure is beneficial (such as double test to triple test), statistical evaluation of the confidence intervals for the detection rates quoted indicates that there is no evidence that the extra test provides an increase in detection. The cervical screening programme has progressively improved, partly through the auspices of a national framework. A similar national approach would benefit Down's screening and is only now being considered: the national screening committee (NSC) is currently drafting recommendations. To ensure optimum screening performance, the NSC should specify the risk thresholds applied, the screening protocols to be used--that is, an opt in programme with a minimum (possibly even a maximum) of two biochemical analytes or a nuchal fold evaluation--and perhaps should even recommend national population parameters to be used for risk calculation. It might even be advisable for statistical work to be carried out to determine whether local derivation of medians is truly necessary. Furthermore, defined options for older women could be specified--for example, should all older patients have the option to proceed directly to amniocentesis if they wish or should National Health Service amniocentesis only be available for those with a "high risk" screening result. The difficulties that will face the NSC in deciding which screening policy to adopt are also considered; specifically, the lack of evidence to suggest that triple testing is superior to double testing, and the lack of evidence to prove the superiority of one analyte over another. This inadequacy of evidence is not from want of trying, but is caused by the problems of collecting enough data to provide statistical significance. Finally, there is one important difference between cervical and Down's syndrome screening that has a major impact on the advice given by any "expert"; namely, patents. Many aspects of Down's screening are subject to patents and, therefore, there is more potential for apparently uncontroversial decisions to rebound with future retrospective patent infringement claims. Thus, it would be sensible to insist that any member of a national body deciding upon Down's screening policy must fully disclose all potential conflicts of interest, both personal and family, before they are allowed to sit on the committee. Furthermore, if a national policy is decided upon, worldwide patent searches should be carried out to determine whether there are any possible unforeseen legal consequences of any recommendation. PMID- 11265174 TI - Chlamydia pneumoniae in vitro and in vivo: a critical evaluation of in situ detection methods. AB - AIMS: There is a considerable discrepancy between data from the detection of Chlamydia pneumoniae in atherosclerotic lesions obtained by means of immunocytochemistry and data obtained using the polymerase chain reaction. This study evaluated methods for the in situ detection and assessment of the viability of C pneumoniae bacteria. METHODS: Chlamydia pneumoniae membrane protein, heat shock protein 60, and lipopolysaccharide were detected by immunocytochemistry, and genomic DNA and 16S rRNA by in situ hybridisation in paraffin wax embedded sections of cultured HEp2 cells infected with C pneumoniae and of lungs from mice infected intranasally with C pneumoniae. RESULTS: Inclusions reactive for all three antigens, DNA, and 16S rRNA were seen in infected HEp2 cells, in all positive bronchus and alveolar epithelial cells, and in some of the positive infiltrate cells in the lungs of mice up to seven days after infection. In all alveolar macrophages and in the infiltrate cells positive for antigens only, the staining pattern was granularly dispersed throughout the cytoplasm up to seven days after infection. At 21 days after infection, only this granular staining pattern was seen for antigens in infiltrate cells and macrophages in the alveoli and bronchus associated lymphoid tissue. At this time point, DNA or 16S rRNA were detected sporadically, but always as inclusion-like staining. CONCLUSIONS: Because antigens with an inclusion-like staining were detected only together with DNA and 16S rRNA, this type of staining pattern suggested the presence of viable bacteria. Thus, the granular staining pattern of antigens in the absence of staining for DNA and 16S is most likely caused by non-viable bacteria. In conclusion, these methods are suitable for the in situ detection of C pneumoniae and the assessment of its viability. PMID- 11265173 TI - Primary pulmonary hypertension: the pressure rises for a gene. AB - Primary pulmonary hypertension (PPH) represents the end stage of a disruption of pulmonary vascular integrity, of unknown cause. Although PPH is associated with several systemic disorders, there have hitherto been few clues as to the aetiological factors responsible for the pathogenesis of this condition. As an example of the application of modern molecular genetics and positional cloning, this leader describes the range of studies currently under way, which aim to find the gene that underlies PPH, and summarises the implications of the identification of such a gene. PMID- 11265175 TI - Chlamydia pneumoniae antigens, rather than viable bacteria, persist in atherosclerotic lesions. AB - AIMS: To evaluate the nature of the presence of Chlamydia pneumoniae or of other members of the order Chlamydiales in atherosclerotic lesions. METHODS: Consecutive sections of 13 carotid artery specimens obtained at necropsy and of C pneumoniae infected HEp2 cells were analysed using: (1) immunocytochemistry (ICC) to detect C pneumoniae membrane protein; (2) in situ hybridisation (ISH) using a polymerase chain reaction (PCR) fragment of the omp1 gene to detect C pneumoniae specific DNA; (3) ISH using an oligonucleotide probe to detect Chlamydiales specific 16S rRNA; (4) PCR to detect C pneumoniae 16S rDNA; and (5) in situ DNA nick and labelling (TUNEL) to detect fragmented DNA. RESULTS: Staining by ICC and ISH of infected HEp2 cells showed characteristic inclusions. Chlamydia pneumoniae membrane protein was demonstrated in macrophages in advanced atherosclerotic lesions (six of six), but not in fatty streaks (none of two), or normal arteries (none of five). ISH assays using both probes and PCR were all negative, indicating the absence of both specific C pneumoniae DNA and Chlamydiales specific 16S rRNA. Only after treatment with DNAse I were uniformly sized dots demonstrated by the TUNEL assay in inclusions of infected HEp2 cells. The TUNEL assay showed a similar staining pattern in macrophages in five carotid artery specimens, of which four were also positive for C pneumoniae membrane protein. Both macrophage populations were morphologically similar and were similarly distributed. CONCLUSIONS: No evidence was obtained for the involvement of other members of the order Chlamydiales in atherosclerosis. The presence of C pneumoniae antigen in the absence of DNA and 16S rRNA suggests that antigens, rather than viable bacteria, persist in atherosclerotic lesions. PMID- 11265176 TI - Fetal macrosomia related to maternal poorly controlled type 1 diabetes strongly impairs serum lipoprotein concentrations and composition. AB - AIMS: To determine the effects of fetal macrosomia related to maternal type 1 diabetes on the lipid transport system. METHODS: Serum lipoprotein concentrations and composition and lecithin:cholesterol acyltransferase (LCAT) activity were investigated in macrosomic newborns (mean birth weight, 4650 g; SEM, 90) and their mothers with poorly controlled type 1 diabetes, in appropriate for gestational age newborns (mean birth weight, 3616 g; SEM, 68) and their mothers with well controlled type 1 diabetes, and macrosomic (mean birth weight, 4555 g; SEM, 86) or appropriate for gestational age (mean birth weight, 3290 g; SEM, 45) newborns and their healthy mothers. RESULTS: In mothers with well controlled type 1 diabetes, serum lipids, apolipoproteins, and lipoproteins were comparable with those of healthy mothers. Similarly, in their infants, these parameters did not differ from those of appropriate for gestational age newborns. Serum triglyceride, very low density lipoprotein (VLDL), apolipoprotein B100 (apo B100), and high density lipoprotein (HDL) triglyceride concentrations were higher, whereas serum apo A-I and HDL3 concentrations were lower in mothers with diabetes and poor glycaemic control than in healthy mothers. Their macrosomic newborns had higher concentrations in all serum lipids and lipoproteins, with high apo A-I and apo B100 values compared with appropriate for gestational age newborns. In macrosomic infants of healthy mothers, there were no significant differences in lipoprotein profiles compared with those of appropriate for gestational age infants. LCAT activity was similar in both groups of mothers and newborns. CONCLUSION: Poorly controlled maternal type 1 diabetes and fetal macrosomia were associated with lipoprotein abnormalities. Macrosomic lipoprotein profiles related to poor metabolic control of type 1 diabetes appear to have implications for later metabolic diseases. PMID- 11265178 TI - Audit and internal quality control in immunohistochemistry. AB - AIMS: Although positive and negative controls are performed and checked in surgical pathology cases undergoing immunohistochemistry, internal quality control procedures for immunohistochemistry are not well described. This study, comprising a retrospective audit, aims to describe a method of internal quality control for immunohistochemistry. A scoring system that allows comparison between cases is described. METHODS: Two positive tissue controls for each month over a three year period (1996-1998) of the 10 antibodies used most frequently were evaluated. All test cases undergoing immunohistochemistry in the months of April in this three year period were also studied. When the test case was completely negative for a given antibody, the corresponding positive tissue control from that day was examined. A marking system was devised whereby each immunohistochemical slide was assessed out of a possible score of 8 to take account of staining intensity, uniformity, specificity, background, and counterstaining. Using this scoring system, cases were classified as showing optimal (7-8), borderline (5-6), or unacceptable (0-4) staining. RESULTS: Most positive tissue controls showed either optimal or borderline staining with the exception of neurone specific enolase (NSE), where most slides were unacceptable or borderline as a result of a combination of low intensity, poor specificity, and excessive background staining. All test cases showed either optimal or borderline staining with the exception of a single case stained for NSE, which was unacceptable. CONCLUSIONS: This retrospective audit shows that immunohistochemically stained slides can be assessed using this scoring system. With most antibodies, acceptable staining was achieved in most cases. However, there were problems with staining for NSE, which needs to be reviewed. Laboratories should use a system such as this to evaluate which antibodies regularly result in poor staining so that they can be excluded from panels. Routine evaluation of immunohistochemical staining should become part of everyday internal quality control procedures. PMID- 11265177 TI - Is the detection of Mycobacterium tuberculosis DNA by ligase chain reaction worth the cost: experiences from an inner London teaching hospital. AB - AIMS: To evaluate the clinical usefulness and the costs of using a rapid, commercial ligase chain reaction test (LCx) to detect Mycobacterium tuberculosis directly from clinical samples. METHODS: A prospective study of 2120 routine clinical specimens from 1161 patients over a 13 month period. Investigations for mycobacterial disease by microscopy, culture, and the Abbott LCx assay were performed. Sequential LCx assays were monitored in a cohort of patients undergoing treatment. The costs of the assay were calculated using the WELCAN system. Sensitivity, specificity, and positive and negative predictive values of the LCx assay were compared with conventional tests. The performance of the assay in patients undergoing treatment and cost in terms of WELCAN units converted to pounds/annum was studied. RESULTS: The assay was 85%/88% sensitive and 98%/100% specific in culture confirmed/clinically confirmed cases of tuberculosis, respectively. The assay was not useful for the measurement of treatment outcomes. The test cost approximately 42,500 Pounds/annum. CONCLUSIONS: The assay is a rapid, sensitive, and specific adjunct to clinical diagnosis, especially in differentiating non-tuberculous mycobacteria. However, it does not differentiate old and treated tuberculosis from reactivated disease, it is not useful to monitor adherence to treatment, and it is expensive. PMID- 11265179 TI - Clinical use of WHO haemoglobin colour scale: validation and critique. AB - AIM: The World Health Organisation (WHO) haemoglobin colour scale has been developed as a simple, inexpensive clinical device for diagnosing anaemia when laboratory based haemoglobinometry is not available. In an initial validation study at several health centres, scale readings were compared with measurements of haemoglobin by the laboratory. This showed the scale to have 90% sensitivity and 70% specificity in identifying whether anaemia was present or not. In addition, when present, the degree of anaemia was correctly classified in clinical terms as moderate, pronounced, or severe, with an overall sensitivity of 60% and specificity of 88%. Errors were mainly marginal--that is, between two adjacent categories--but there were also some major discrepancies, such as a blood with a haemoglobin of 6-7 g/dl being read as normal or vice versa. Because this would compromise the scale's reliability in practice, this study was undertaken to identify the cause of the discrepancies and to reassess the performance of the scale. METHODS: Venous blood samples were collected into potassium EDTA from patients attending selected clinics at three South African hospitals with good laboratory facilities. A prototype of the device was used unsupervised by nursing staff, doctors, and phlebotomists, who were told to follow the printed instructions. The blood specimens were then immediately sent to the laboratory where haemoglobin was measured by standardised automated blood cell counters. Any discrepancies > 1 g/dl were recorded and the tests were repeated by the same operators under supervision of the investigators. RESULTS: Almost all the errors that occurred resulted from the incorrect use of the device, namely: inadequate or excessive blood, reading the results too soon or too late (beyond the limit of two minutes), poor lighting, or holding the scale at the wrong angle. The accuracy improved dramatically when the tests were repeated under supervision and these faults were avoided: 95% of readings were within 1 g/dl of the reference measurements, and 97% within 1.5 g/dl. Anaemia screening showed 96% sensitivity and 86% specificity. Clinical judgement of pallor was frequently wrong, whereas the scale gave the correct diagnosis in more than 97% of cases. CONCLUSION: The study confirmed the usefulness and reliability of the scale and its advantage over clinical signs for the diagnosis of anaemia, thus providing a clinically reliable near patient method in the absence of a laboratory. The instructions are easy to follow but must be strictly adhered to. PMID- 11265180 TI - Clear cell carcinoma of the ovary arising in a mucinous cystadenoma. AB - A 57 year old woman presented complaining of increasing abdominal swelling of six months duration. A mixed solid cystic left ovarian tumour measuring 24 cm in diameter was excised. Histology showed numerous cysts lined by benign mucinous epithelium blending imperceptibly into borderline clear cell and mucinous areas that in turn merged with an invasive clear cell carcinoma. To the best of our knowledge, this is the first reported case of clear cell carcinoma arising in a mucinous cystadenoma. The implications for the previously postulated pathogenesis of these tumours are discussed. PMID- 11265181 TI - Chronic myeloid leukaemia occurring in a patient with hairy cell leukaemia. AB - Occurrences of second malignancies in hairy cell leukaemia are well recognised. Most of these malignancies are either solid tumours or lymphoproliferative disorders. The association of myeloproliferative disorders with hairy cell leukaemia (HCL) is very rare. This report describes a case of a patient with HCL who after remaining in remission developed Philadelphia chromosome positive chronic myeloid leukaemia (CML), which rapidly transformed to acute lymphoblastic leukaemia with further cytogenetic abnormalities. PMID- 11265182 TI - Polymorphous low grade adenocarcinoma with distant metastases and deletions on chromosome 6q23-qter and 11q23-qter: a case report. AB - Polymorphous low grade adenocarcinomas (PLGAs) are thought to be indolent tumours that are localised preferentially to the palate and affect the minor salivary glands almost exclusively. Metastases to locoregional lymph nodes occur in only 6 10% of cases. Recently, two cases of PLGA with microscopically confirmed distant metastases have been reported. This study reports a third case of PLGA with histologically and immunohistochemically confirmed distant metastases. It is the first case with multiple pleural, as well as pulmonary parenchymal, metastases and metastases in cervical and paraoesophageal lymph nodes. In most cases, PLGAs are salivary gland tumours with limited potential to metastasis and a good prognosis after local treatment. However, the recently reported cases reveal that the tumour can give rise to widely spread metastases. To obtain more information about the incidence of distant metastases, periodic chest x ray examination during follow up is desirable. PMID- 11265183 TI - Pulmonary adenocarcinoma metastatic to pituitary craniopharyngioma. PMID- 11265185 TI - Response to the October 2000 veterinary medical question of the month. PMID- 11265184 TI - Parasitologic data incidental to survey of prevalence of Giardia in dogs. PMID- 11265186 TI - An ethicist's commentary on trying alternative therapy. PMID- 11265187 TI - Seroprevalence of infection with Mycobacterium avium subspecies paratuberculosis, bovine leukemia virus, and bovine viral diarrhea virus in maritime Canada dairy cattle. AB - The purpose of this study was to survey the seroprevalence of infection with the agents of production-limiting diseases in dairy cattle in New Brunswick, Nova Scotia, and Prince Edward Island. In 30 randomly selected herds per province, 30 cattle per herd were randomly selected and tested for antibodies to bovine leukemia virus (BLV) and Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis), while 5 unvaccinated cattle over 6 months of age were tested for antibodies to bovine viral diarrhea virus (BVDV). For BLV, 20.8% (15.8% to 27.0%) of cows were positive, and 70.0% (60.3% to 79.7%) of herds had at least one positive cow. In BLV-positive herds, the average BLV prevalence was 30.9% (24.8% to 37.2%). For M. paratuberculosis, 2.6% (1.8% to 3.9%) of cows were positive, and 16.7% (8.8% to 24.5%) of herds had at least 2 M. paratuberculosis positive cows. In M. paratuberculosis-positive herds, the average M. paratuberculosis prevalence was 8.5% (6.9% to 10.1%). For BVDV, 46.1% (35.5% to 56.7%) of herds had at least 1 BVDV-positive animal with a titer greater than or equal to 1:64. PMID- 11265188 TI - [Comparison of four protocols for preoperative preparation in cattle]. AB - This study was designed to evaluate 4 preoperative skin preparations, that is, more specifically, to compare the efficacy of chlorhexidine gluconate (CG) and povidone-iodine (PI), as well as 2 hair removal techniques (clipper alone or clipper followed by razor) for preoperative skin preparation in cattle. The 4 protocols resulted in a significant decrease in the number of bacterial colony forming units (cfu). Group 4 (clipping + shaving + CG) had a significantly lower number of preoperative cfu per gel plate compared with groups 1 (clipping + PI) and 3 (clipping + shaving + PI). Skin reaction frequency was significantly higher in groups 3 and 4 (47.8% for both protocols) than in groups 1 and 2 (clipping + PI or CG) (8.7% for both). Wound infection frequency was 4.3% (4/92) and no significant difference was observed between the 4 treatment groups. The 4 protocols tested were equivalent as to efficacy and satisfactorily decreased skin microflora. Clipping alone was shown to be preferable to clipping plus shaving as a method of hair removal in cattle, with fewer skin reactions and no more wound infections. PMID- 11265189 TI - Cutaneous metastases of a mammary carcinoma in a llama. AB - An 8-year-old, female llama was evaluated for nonhealing, ulcerative, cutaneous lesions, which also involved the mammary gland. Biopsies of the lesions distant from and within the mammary gland area revealed an aggressive carcinoma. The tumor was confirmed at necropsy to be a mammary gland adenocarcinoma with cutaneous metastasis. PMID- 11265190 TI - Incursion of epizootic hemorrhagic disease into the Okanagan Valley, British Columbia in 1999. AB - In September 1999, unusually high mortality rates in white-tailed deer and California bighorn sheep occurred in the southern Okanagan Valley. Necropsy and histopathologic findings were compatible with epizootic hemorrhagic disease (EHD); the presence of virus was not demonstrated. Subsequent serologic and polymerase chain reaction assays on sentinel cattle suggested an EHD virus incursion. PMID- 11265191 TI - Presumptive diagnosis of Clostridium botulinum type D intoxication in a herd of feedlot cattle. AB - Fifty-two feedlot cattle exhibited clinical signs suggestive of botulism. Clostridium botulinum type D organisms were recovered from ruminal fluid of 4 of the 5 affected animals tested and were isolated from bakery waste fed to the cattle. Clostridium botulinum type D has not been reported previously in Canadian cattle. PMID- 11265192 TI - Nasal dermoid sinus in an American cocker spaniel. AB - An American cocker spaniel was presented for a subcutaneous mass and draining tract located between its eyes. Contrast radiography and surgical excision showed communication of the tract with the left frontal sinus and rostral cerebral dura, respectively. A dermoid sinus was diagnosed by a combination of gross and histologic findings. PMID- 11265194 TI - Distribution of Streptococcus suis capsular types in 2000. PMID- 11265193 TI - Equine herpesvirus myeloencephalopathy in a 14-year-old quarter horse stallion. AB - A 14-year-old, quarter horse stallion was presented in lateral recumbency, unable to rise. Equine herpesvirus myeloencephalopathy was diagnosed, based on presentation, clinical signs, and the ruling out of other possibilities. After initial rapid improvements, ataxia remained, as did chronic cystitis secondary to bladder paralysis. He was euthanized after 2 months. PMID- 11265195 TI - Rhinocerebral zygomycosis in a sheep. PMID- 11265196 TI - Diagnostic ophthalmology. Epithelial inclusion cyst of the right cornea. PMID- 11265198 TI - [Molecular biology techniques in articles published in Revista Medica de Chile]. PMID- 11265197 TI - [New concepts in hypertension]. AB - The management of arterial hypertension has experienced great changes during the past years, based on the knowledge of its pathophysiology, as well as the development of new antihypertensive drugs. Recent studies show the effect of the dysfunction of the endothelium in hypertension, sleep apnea and a higher prevalence of primary hyperaldosteronism. The Sixth report of the Joint Committee makes a new classification of this pathology, including a new concept of the stratification of risk for each stage of the disease, and according to it a new way of treatment. This fact marks a new and a more aggressive behaviour to treat the hypertense patient. With respect to the treatment, the emphasis has been on the non-pharmacological measures, and the development of new antihypertensive drugs of long action and high through/peek rate. Prevention continues to be an important goal and main challenge for the clinician, given the high morbidity and mortality rate caused by hypertension. PMID- 11265200 TI - [Successful rescue of out-of-hospital airway emergency using the esophageal tracheal Combitube by non skilled personnel]. PMID- 11265201 TI - [Hepatitis C virus in a group of hematological and oncohematological patients]. AB - BACKGROUND: Little information is available in Chile about hepatitis C virus (HCV) in hematological and oncohematological patients. AIM: To evaluate the prevalence of hepatitis C virus markers in a group of hematological and oncohematological pediatric patients seen at Valdivia Regional Hospital. PATIENTS AND METHODS: Antibodies against virus C, determined by ELISA and viral RNA, determined using RT-polymerase chain reaction, were measured in 54 blood samples from children with hematological diseases (34 with Acute Lymphoblastic Leukaemia, 4 with Hodgkin Diseases, 4 with Haemolytic Anemia, 5 with Sarcomas, 2 with Non Hodgkin Lymphoma, 2 with Thrombocytopenic Purpura, 1 with an Ependimoma, one with a Wilms Tumor and 1 with a Von Willebrand Disease). RESULTS: All samples were negative for antibodies against hepatitis C virus. Viral RNA was found in four children, all with a diagnosis of acute lymphoblastic leukemia and who received chemotherapy and multiple transfusions. CONCLUSIONS: The prevalence of Viral RNA for hepatitis C virus in oncohematological patients in our study is high and associated with the use of chemotherapy and multiple transfusions. PMID- 11265202 TI - [Instrument for the functional evaluation of disability rehabilitation. Study on the reliability and clinical experience with the use of functional independence measure]. AB - BACKGROUND: The Functional Independence Measure (FIM) is an instrument widely used to assess the results and progress of a medical rehabilitation program. AIM: To assess the inter rater agreement in the application of FIM and show the clinical experience with its use in disabled patients enrolled in a rehabilitation program. PATIENTS AND METHODS: FIM was applied in 40 patients and the inter rater agreement was assessed, comparing raters with and without training in its use. Agreement was evaluated using Kappa test. Afterwards, the FIM was used to assess changes in 100 patients hospitalized and being rehabilitated at the Rehabilitation ward of a general hospital. RESULTS: Inter rater agreement is high in physical areas of the FIM and low in cognitive areas. Training significantly improves agreement in communication and cognitive areas. During a mean period of hospitalization of 38 days, FIM score changed from a mean of 47.2 to 92 points, with an improvement of 1.18 points per hospitalization day. The higher improvement was achieved in the physical area of the instrument. CONCLUSIONS: Training is required for a proper application of FIM. When used adequately, this instrument allows an assessment of the degree of disability and the changes obtained with rehabilitation programs. PMID- 11265203 TI - [Indoor air pollution in a zone of extreme poverty of La Pintana, Santiago Chile]. AB - BACKGROUND: Indoor pollution can be an important risk factor for human health, considering that people spend more than 60% of their time in their houses. AIM: To investigate indoor pollution in a zone of extreme poverty in Metropolitan Santiago. MATERIAL AND METHODS: During 24 h, carbon monoxide (CO), sulfur dioxide (SO2), respirable particulate matter (PM10), polycyclic aromatic hydrocarbons absorbed in PM5, temperature and humidity, were measured in the interior of 24 houses in a La Pintana, Santiago. RESULTS: The higher pollutant concentrations were observed during hours when heating was used, in houses that used coal (mean PM10 250 micrograms/m3, CO 42 ppm, SO2 192 pph) or firewood (mean PM10 489 micrograms/m3, CO 57 ppm, SO2 295 pph). In all houses, polycyclic aromatic hydrocarbons were detected and they came from the interior of the house and not from external filtered air. Coal, firewood and cigarette smoke were important sources of carcinogenic and kerosene and gas were sources of non carcinogenic polycyclic aromatic hydrocarbons. CONCLUSIONS: In the houses studied, the population was exposed to an accumulation of highly toxic pollutants, caused by a lack of ventilation. A high relative humidity also contributed to the growth of biological pollutants. PMID- 11265205 TI - [Strength of inspiratory muscles in chronic heart failure and chronic pulmonary obstructive disease]. AB - BACKGROUND: The maximal pressure generated by inspiratory muscles (PIMax) is an index of their strength which is diminished in both chronic obstructive pulmonary disease (COPD) and chronic heart failure (CHF). Although inspiratory muscle power output (IMPO), which includes both strength and velocity of shortening, has been shown to be reduced in COPD, there is no information regarding IMPO in CHF. AIM: To measure IMPO in patients with CHF and COPD. PATIENTS AND METHODS: We studied 9 CHF patients with functional capacity II and III and 9 patients with severe COPD. Eight normal subjects of similar ages were included as controls. Power output was measured using the incremental threshold loading test. RESULTS: Maximal IMPO was significantly reduced in both groups of patients. Power output developed with each increasing load was also diminished, basically as a consequence of a reduction in Vinsp. The degree of dyspnea at the end of the test was greater in COPD than in CHF patients and normal subjects. For a given level of power, dyspnea was also greater in patients than in normals subjects. There was no decrease in SpO2 during the test. CONCLUSIONS: IMPO is equally reduced in COPD and CHF patients. Power output is better related to dyspnea than PIMax, probably because of the inclusion of shortening velocity. PMID- 11265204 TI - [Differences in plasma antioxidants according to socioeconomic level in Chilean women]. AB - BACKGROUND: Free radical-mediated oxidative damage is a known initial event in atherogenesis. Cardiovascular disease is frequent in the Chilean population showing differences in the prevalence of risk factors of the disease according to socioeconomic level (SEL). AIM: To determine levels of antioxidants and lipid peroxides in Chilean women from different SEL. PATIENTS AND METHODS: Blood samples were taken from 81 women for measurements of plasma ascorbic acid, beta carotene, alpha-tocopherol, licopene, ubiquinol, glutathione, total plasma antioxidant capacity, and lipid peroxides (TBARS). RESULTS: Individuals in the lower SEL showed reduced levels of plasma beta-carotene, ascorbic acid, alpha tocopherol, and ubiquinol compared to women in the higher SEL. There were no differences between groups in the plasma levels of glutathione, total antioxidant capacity, or TBARS. CONCLUSIONS: The results could be explained in part by the higher consumption of fruits and vegetables in women from the upper SEL. PMID- 11265206 TI - [Radiofrequency catheter ablation of symptomatic isolated ventricular extrasystole in patients with a normal heart]. AB - INTRODUCTION: Premature ventricular depolarizations (PVDs) in patients without heart disease, are a frequent clinical problem that can cause important symptoms. Most commonly, this benign arrhythmia responds to treatment with antiarrhythmic drugs. However, occasionally PVDs are refractory to pharmacological treatment but they can be eliminated with radiofrequency catheter ablation. AIM: To show our experience with four patients in whom we used this method. MATERIAL AND METHOD: We studied three men and a woman, twelve to forty six years old. All of them were symptomatic, their EKG and echocardiogram were normal and they had been treated with several drugs without response. In three of them the PVDs had left bundle branch block morphology with inferior axis; the other patient had right bundle branch block morphology with superior axis. The origin of the PVDs was determined using pace mapping. RESULTS: Two of the patients had spontaneous PVDs; in the other two isoproterenol infusion was used to induce them. In three patients the origin of the PVDs was located in right ventricular outflow and in the other in the anterolateral region of the left ventricle. None had sustained atrial or ventricular arrhythmia. In all of them PVDs were eliminated. A patient presented a second morphology that could not be treated. None of the patients had complications and they were discharged within the next 24 hours. Three noted symptomatic improvement and after 18 months, only one had a probable recurrence of the arrhythmia. CONCLUSIONS: Radiofrequency catheter ablation can be successfully used to eliminate PVDs in severely symptomatic and drug-resistant patients. PMID- 11265207 TI - [Disabling malformations in Chile. Latin American Cooperative Study of Congenital Malformations (ECLAMC), 1982-1997]. AB - BACKGROUND: The ECLAMC (Estudio Colaborativo Latinoamericano de Malformaciones Congenitas) is an epidemiological surveillance program for congenital defects that operates in Chile since 1969. AIM: To communicate the frequency of disabling congenital defects in Chile in the period 1982-1997. MATERIAL AND METHODS: A review of the ECLAMC registry, choosing 12 congenital defects: amelia, limb amputations, limb reductions, arthrogryposis, hip luxation and subluxation, spina bifida, hydrocephaly, microcephaly, cephalocele, talipes equinovarus, Down syndrome and multiple abnormalities. RESULTS: In the study period, 283,403 births occurred and 7,917 newborns were malformed (7,654 born alive and 263 stillbirths). The congenital defects prevalence rates appeared higher in Chile than in other Latin American countries, specially among stillbirths. Among the studied maternity hospitals, the Clinical Hospital of the University of Chile, showed the higher prevalence of congenital defects. Rancagua and the Navy Hospital in Valparaiso have a high frequency of Down syndrome. Global rates in Chile and in the rest of ECLAMC for specific defects, do not have significant differences, except for hip subluxation, that has a lower incidence in Chile. CONCLUSIONS: The ECLAMC allows to have a good knowledge of the prevalence of congenital malformations in Latin America. PMID- 11265209 TI - [Study of 3 hypervariable loci in a mixed Chilean population]. AB - BACKGROUND: Genetic markers are useful to study evolution parameters in populations and to determine kinship. AIM: To characterize three short tandem repeat loci in a sample of Chilean subjects and compare them with Caucasian and Hispanic populations. MATERIAL AND METHODS: Three hundred ninety three unrelated subjects that were sent for genetic studies from courts of justice, were studied. The loci FESFPS, F13A01 and vWA in blood samples, were typified amplifying DNA by polymerase chain reactions. RESULTS: The three studied loci were highly polymorphic. F13A01 and FESFPS were in Hardy-Weinberg genetic equilibrium. A significant excess of heterozygotes was detected for vWA locus. There were no differences in allele frequencies, according to ethnic origins of last names. Allele frequencies for F13A01 and vWA loci were similar to those of Hispanic populations of Unites States and FESFPS loci was different. CONCLUSIONS: All three loci had a high efficiency for genetic identification tests according to the estimated a priory exclusion probability. PMID- 11265210 TI - [Hemorrhage caused by the rupture of a pancreatic pseudoaneurysm. Case report]. AB - We report a 52 year old man with a pancreatic pseudocyst, that was admitted with severe abdominal pain, severe vomiting, fever and malaise. The clinical picture was considered secondary to a pseudocyst infection and the patient was operated, draining the infected cyst performing a necrosis surgery and pancreatojejunostomy. Forty eight hours after the operation, an ostomy bleeding was detected. A upper mesenteric artery angiography showed two pseudoaneurysms in the gastroduodenal artery, that were embolized. Bleeding stopped initially, but seven days later, it reappeared. The patient was subjected to an emergency pancreatoduodenectomy. Postoperative evolution was uneventful and the patient was discharged two weeks later. Spontaneous bleeding of pseudoaneurysms secondary to chronic pancreatitis is a complication with a 15 to 40% mortality that must be bore in mind. PMID- 11265211 TI - [Metachronous pancreatic metastasis of a renal cell carcinoma. 3 new cases]. AB - We report two males and one female, aged 71, 81 and 73 years old respectively, previously operated of a renal cell carcinoma. During the follow up of the 71 years old male, the CAT scan showed three lesions in the pancreas. Also, the CAT scan in the female showed one lesion in the tail of the pancreas. The 81 years old male was admitted to the emergency room due to a upper gastrointestinal bleeding. Endoscopy showed a proliferating lesion in the second portion of the duodenum whose biopsy showed a clear cell carcinoma. The CAT scan showed also a tumoral mass in the head of the pancreas. All three patients were subjected to surgical resection of the tumors without postoperative complications or mortality. PMID- 11265212 TI - [Treatment of advanced heart failure by heart transplantation]. AB - BACKGROUND: Heart transplantation currently provides the most effective treatment for advanced heart failure. However, medical therapy for this condition has also improved, heart donors are scarce and the cost of the procedure is high. Therefore the indications and management of these patients need reevaluation. AIM: To analyze the results of 24 patients submitted to heart transplantation for end-stage heart failure needing repeated hospitalizations and i.v. inotropes for compensation. PATIENTS AND METHODS: The group was comprised by 21 men and 3 women with a mean age of 36.8 years, mean left ventricular ejection fraction 19 +/- 4.5%, mean systolic pulmonary artery pressure 48 +/- 13 mmHg (24-70) and mean pulmonary vascular resistance 2.6 Wood Units (1-5). Fourteen patients (58%) had a previous median sternotomy. Immunosuppression did not include induction therapy and steroids were discontinued early. RESULTS: Operative mortality was 4% at 30 days. Actuarial survival at one year was 90% and at 5 years 72%. Freedom from rejection at one year was 76% and at 5 years 50%. Freedom from infection was 70% at one year and 56.5% at five years. All patients with more than 3 months of follow-up were in functional class I. CONCLUSIONS: These results justify the proposed modifications for transplantation protocols. PMID- 11265213 TI - [Bilateral pheochromocytoma: laparoscopic surgery in 2 cases]. AB - Laparoscopic adrenalectomy, if done by skilled surgeons, is now the first choice for treating most adrenal tumors, including bilateral pheochromocytoma. We report two women, aged 35 and 34 years old, with bilateral adrenal pheochromocytoma successfully excised by laparoscopic surgery. Both had severe hypertension, high urinary catecholamine values (epinephrine + norepinephrine: 528 and 1083 ug/24 h) and bilateral adrenal tumors at CT scan. After 4 weeks of doxazosin treatment, a laparoscopic transperitoneal adrenalectomy was done (Gugner's technique), with surgical times of 7 and 5 hours respectively. Both patients received hydrocortisone and only the second one required one unit of packed cells. Postoperative evolution was uneventful and both patients were discharged at the fifth postoperative day. At two months of follow up, both patients are asymptomatic and normotensive. PMID- 11265214 TI - [The Chilean Association of Biomedical Journal Editors]. AB - On September 29th, 2000, The Chilean Association of Biomedical Journal Editors was founded, sponsored by the "Comision Nacional de Investigacion Cientifica y Tecnologica (CONICYT)" (the Governmental Agency promoting and funding scientific research and technological development in Chile) and the "Sociedad Medica de Santiago" (Chilean Society of Internal Medicine). The Association adopted the goals of the World Association of Medical Editors (WAME) and therefore it will foster "cooperation and communication among Editors of Chilean biomedical journals; to improve editorial standards, to promote professionalism in medical editing through education, self-criticism and self-regulation; and to encourage research on the principles and practice of medical editing". Twenty nine journals covering a closely similar number of different biomedical sciences, medical specialties, veterinary, dentistry and nursing, became Founding Members of the Association. A Governing Board was elected: President: Humberto Reyes, M.D. (Editor, Revista Medica de Chile); Vice-President: Mariano del Sol, M.D. (Editor, Revista Chilena de Anatomia); Secretary: Anna Maria Prat (CONICYT); Councilors: Manuel Krauskopff, Ph.D. (Editor, Biological Research) and Maritza Rahal, M.D. (Editor, Revista de Otorrinolaringologia y Cirugia de Cabeza y Cuello). The Association will organize a Symposium on Biomedical Journal Editing and will spread information stimulating Chilean biomedical journals to become indexed in international databases and in SciELO-Chile, the main Chilean scientific website (www.scielo.cl). PMID- 11265215 TI - [Investigation and development of new drugs: from the molecule to drug]. AB - There are many stages and a large investment of both time and money involved in the process of research and development before a new drug can be prescribed for clinical use. Of the thousands of new molecular entities, only one or two are approved for commercialization, after having endured a trajectory of 12 to 15 years in clinical trials in both animals and humans, demonstrating their therapeutic effectiveness and safety. There are three large administrators of medicines that control the process of new drug registration, the FDA--Food and Drug Administration of the USA being the largest and most well known. This article is based on their model and details the various stages that the molecule must undergo before finally being administered to patients. The future holds many exciting promises for new drug development with the advent of the human genome project and other highly advanced technological methods. However, the main challenge still remains, which is to guarantee the access of basic medicines to the majority of the world's population that is still without them. PMID- 11265216 TI - Structural molecules of connective tissue matrices. PMID- 11265217 TI - Anatomy of the immune system: facts and problems. AB - In the introductory section of this report, the anatomy of the immune system, from organs and tissues to molecules, will be reviewed briefly. Cell proliferation and differentiation in the central lymphoid organs (thymus and bone marrow) yield a repertoire of T- and B-cell clones that seed into peripheral lymphoid organs (spleen, lymph nodes and Mucosa-Associated Lymphoid Tissue, MALT), where humoral and cell-mediated antigen-specific immune responses occur. The stringent process of clonal selection in the central lymphoid organs implies deletion of inappropriate cells via apoptosis. In the peripheral lymphoid organs, the potential of unlimited activation and expansion of lymphocytes in response to antigens is primarily regulated by apoptosis and anergy. These events, on the one hand, are relevant to prevent autoimmunity and lymphoproliferative disorders; on the other hand, clonal deletion and anergy provide a detrimental escape to immune recognition of malignant cells. Two major inhibitory mechanisms of the immune response have emerged recently. One is linked to the existence of bona fide suppressor cells and cytokines; the other relies on the existence of inhibitory molecules expressed by T, B and NK cells, as well as by other leukocytes. In the studies herein reported, emphasis will be given to surface membrane molecules that down-regulate T-cell-mediated immune responses. These molecules control interactions between T cells and antigen presenting cells (APC's) or target (virus-infected or mutated) cells that have to be killed. Two sets of molecules exist that either upregulate (coactivation molecules) or down-regulate (inhibitory molecules) T-cell mediated responses. The latter aspect of the immune regulation, i.e. molecules that limit the expansion of T-cell clones following specific recognition of antigens will be considered in depth. Two inhibitory molecules, CD152 (CTLA-4) and CD85/LIR-1/ILT2 are expressed in all T cells, being largely confined within intracellular compartments of these lymphocytes when they are in a resting state, but ready to be shuttled to and from the plasma membrane when cells are activated following encounter with antigen. Membrane expression of the two inhibitory molecules is transient and is regulated by an internalization process directed to endosomal compartments and to receptor degradation and/or recycling. CTLA-4 and CD85/LIR-1/ILT2 play a pivotal role in T-cell homeostasis that follows any cell-mediated immune response; their localization and functional role will be thoroughly analyzed. In the last part of this study a major question will be faced, i.e. is the containment of the possibly unlimited expansion of the immune system due to a blockade of the cell cycle? Or, else, could be apoptosis the sole mechanism responsible? Experimental data in support of the latter contention will be provided. PMID- 11265218 TI - [Correlation between sensitivity to fosfomycin and the presence of penicillinase PSE-1 in Pseudomonas aeruginosa]. AB - A prospective survey was carried out during three three-weeks periods in May, October 1997 and October 1998 in 13 teaching hospitals. All non-repetitive isolates of P. aeruginosa collected were subject to serotypage and determination of the inhibiting minimal concentrations for ticarcillin, piperacillin, piperacillin + tazobactam, ceftazidime, imipenem, amikacin, ciprofloxacin and fosfomycin. Identification of the betalactamases and quantification of the cephalosporinase were done for the strains intermediate or resistant to ticarcillin. The most frequent serotypes were O: 6 (17%), O: 11 (13%), O: 1 (10%) and O: 12 (9%). Serotype O: 12 was the least susceptible to antibiotics except for fosfomycin. Whatever the serotype, 76% of P. aeruginosa strains with bla PSE 1 are susceptible to fosfomycin, when only 29.8% of non bla PSE-1 producing strains were susceptible to this antibiotic. Integron encoding bla PSE-1 could be implicated in susceptibility to fosfomycin of P. aeruginosa strains. The associations fosfomycin + imipenem or fosfomycin + ceftazidime could be proposed in case of infections due to P. aeruginosa O: 12. PMID- 11265219 TI - [Epidemic of Staphylococcus aureus nosocomial infections resistant to methicillin in a maternity ward]. AB - Methicillin-resistant Staphylococcus aureus (MRSA) nosocomial infections frequently occur in the hospital environment, but their incidence is less often observed in neonates. In the present investigation, seventeen cases were recorded over a nine-week period (two cases per week). Pulsed field gradient gel electrophoresis confirmed the clonal character of the strain. The hypothesis of manually-transmitted infection due to contamination from multiple sources was reinforced by the fact the epidemic persisted in spite of the elimination of the main human infectious source and an absence of risk factors determined by the case-control study. The role of environmental factors in the persistence of this outbreak of MRSA infection has been considered. PMID- 11265220 TI - [Evaluation of 7 endoscope washer-disinfectors: bactericidal activity of the disinfectants, antibacterial efficacity of paired washer-disinfector products]. AB - The authors studied seven automatic washer disinfectors for flexible endoscopes with two methods. The first method, a microdilution method, studied the bactericidal activity of the seven disinfectants against 21 strains: four reference strains, 14 hospital strains reported in the literature as nosocomial strains responsible for infections transmitted by flexible endoscopes and three vancomycin-resistant Enterococci. The ability of the seven automatic washer disinfectors to decontaminate flexible endoscopes following inoculation with four reference strains was studied with the second method. There were three kinds of results: the results of three automatic washer disinfectors in conformity with both methods; the results of three automatic washer disinfectors conformity with 1 method only; the results of one automatic washer disinfector without conformity with any methods. Both methods should be used for evaluation of automatic washer disinfector. Then, these results emphasize the necessity to modify the use of disinfectants and/or the systems of some automatic washer disinfectors. PMID- 11265221 TI - [Antibiotic resistance of Staphylococcus aureus in urban experience: 6 month study in Aquitaine]. AB - Antibiotic resistance of Staphylococcus aureus has been surveyed by eight city laboratories of the Aquitaine area, during a six month-period (january to june 1998). Antibiotic susceptibility testing has been performed by the disk diffusion method, and the results have been collected and analysed using the SIRscan system. After elimination of the redundant strains, a total of 747 isolates has been retained. They were mainly isolated from pus (64%) collected from patients of the community (40%) or hospitalized in 30 private clinics or nursing homes. The percentages of resistant strains (community/institutions) were: benzylpenicillin: 90% (87/92%), oxacillin: 39% (23/50%), kanamycin: 37% (22/47%); gentamicin: 13% (8/16%), tobramycin: 37% (21/47%), amikacin: 21% (13/27%); netilmicin: 6% (5/7%), erythromycin: 33% (30/35%), spiramycin: 72% (77/69%), lincomycin: 24% (17/29%), pristinamycin: 2% (1/2%), tetracycline: 17% (14/19%); pefloxacin: 40% (25/50%), fosfomycin: 9% (6/12%), rifampicin: 10% (7/13%), fusidic acid: 14% (11/15%), cotrimoxazole and vancomycin: 0%. Meticillin susceptible strains of S. aureus were mostly sensitive to other antibiotics (< or = 6% resistant strains, except for erythromycin: 22%). Among meticillin-resistant S. aureus, heterogeneous strains with a KT phenotype, and various resistance patterns to the remaining antibiotics were predominant (61%), compared to the homogeneous strains with a KTG phenotype and multiresistant to the other antibiotics (32%). The frequencies of resistant strains were highly variable depending on the specimen, the laboratory and the health care institution, except for cotrimoxazole and vancomycin which were always active. PMID- 11265222 TI - [Transient electrical birefringence study of the interaction between DNA and platinum compounds: cis-DDP, trans-DDP and TDP]. AB - The interaction between DNA and the platinum compounds cis-DDP, trans-DDP and TDP has been studied in aqueous solution at pH 7.0 by transient electric birefringence (TEB). Data was obtained on the electro-optical characteristics and hydrodynamic properties of these solutions. The specific interactions between each of the three platinum compounds and DNA were differentiated, and their binding affinity for DNA phosphate sites was as follows, in decreasing order of importance: TDP >> cis-DDP > trans-DDP. PMID- 11265224 TI - [In vitro inhibition of Chlamydia trachomatis growth by liposome-encapsulated cyclines]. AB - The antichlamydial activity of tetracycline (Tet) and doxycycline (Dox) encapsulated in cationic (CaL), anionic (AnL) and neutral (NtL) liposomes has been evaluated in vitro by adding serial dilutions of antibiotics (minimum inhibitory concentration, MIC: 0.12-0.007 microgram/ml; MBC: 4 to 0.25 micrograms/ml) to HeLa 229 cell monolayers inoculated with Chlamydia trachomatis L2/434/Bu (10(3) ufi/ml). Following 72 h incubation at 37 degrees C under a 5% CO2 atmosphere, the chlamydial inclusions were stained by the May-Giemsa method to determine the MICs. After a second and third passage, the MBC1 and MBC2 were determined in antibiotic-free medium. The chlamydial inclusions were then counted to assess the degree of growth inhibition at each antibiotic dilution tested for MBC1 and MBC2 determinations. The MIC, MBC1 and MBC2 of the various antichlamydial agents were as follows: Tet (0.12; 4; 4), AnL-Tet (0.01; 1; 1), NtL-Tet (0.03; 1; 2), Dox (0.06; 1; 2), CaL-Dox (0.03; 0.5; 2), AnL-Dox (0.01; 1; 2), and NtL-Dox (0.03; 0.5; 0.5). It was found that Tet and Dox liposome encapsulated antibiotics were more active than their non-encapsulated counterparts, and the inclusion count showed a higher inhibitory activity of the former antibiotics on chlamydial growth. The inhibition of chlamydial growth by AnL-Tet may be of bactericidal nature. In conclusion, liposome-encapsulated drugs could be of value in the treatment of chlamydial infections. PMID- 11265223 TI - [Anti-endomysium, anti-reticulin and anti-gliadin antibodies, value in the diagnosis of celiac disease in the child]. AB - Coeliac disease is associated with gluten intolerance in genetically predisposed subjects. Environmental factors, particularly of viral origin, may also play a major role. In this study, the presence of IgA class anti-endomysium antibodies (AEA-IgA), IgA class anti-reticulin antibodies (ARA-IgA) and IgA class anti gliadin antibodies (AGA-IgA) was investigated in 120 serum samples from 120 children (60 patients with coeliac disease and 60 control subjects). The AEA were detected by indirect immunofluorescence on sections of human umbilical cord. The ARA were also investigated by the same technique in rat kidney, liver and stomach. The AGA were determined by an enzyme-linked immunosorbent assay (ELISA). In the patients with coeliac disease, the sensitivity of AEA and ARA was 86% and 76% respectively. In both cases, the specificity was 100%. In children below two years of age, the sensitivity of AEA and ARA was too low, i.e., 57% and 35% respectively. In children aged between two and 15 years, the sensitivity of AEA and ARA was 95% and 89% respectively. The sensitivity of IgA class AGA was 86%, and their specificity was 83%. In this study population, these results show that IgA class AEA are interesting markers for the diagnosis of coeliac disease in the child, and could be used in screening for coeliac disease in a high-risk population. PMID- 11265226 TI - [New strategies for antiretroviral treatment in HIV infected patients]. AB - Medical care of HIV-infected patients has been greatly improved during the last four years with the combination of new highly active drugs and routine monitoring of plasma viral load. Three-drugs regimens including a protease inhibitor are the recommended treatments. The objective is the suppression of viral load below detectable levels and the correction of immune deficit. Non-nucleoside reverse transcriptase inhibitors can be used in new therapeutic regimens. To ameliorate efficacy of the antiretroviral treatment and to avoid viral resistance, the tolerance and the ability to adhere are an important challenge. PMID- 11265225 TI - [Gene therapy of X-linked severe combined immunologic deficiency (SCID-X1)]. AB - X-linked severe combined immunodeficiency (SCID-X1) is a recessive hereditary disorder in which early T and Natural Killer (NK) lymphocyte development is blocked. The genetic disorder results from mutations in the common gamma c chain that participates in several cytokine receptors including the interleukin-2 (Il 2), Il-4, Il-7, Il-9, Il-15 receptors. SCID-X1 offers a reliable model for gene therapy as it is a lethal condition that is, in many cases, curable by allogeneic bone marrow transplantation. We have shown that retrovirus-mediated transfer of the gamma c cDNA induced gamma c chain expression and restored the function of the high-affinity IL-2 receptor on SCI-X1 EBV-transformed B-cell lines. We have the designed culture conditions to study NK-cell and T-cell development of CD34+ hematopoietic progenitor cells. In the culture systems, gamma c transduced CD34+ marrow cells from two SCID-X1 patients were able to mature into CD56+ and/or CD16+ NK cells and into CD4+ TCR alpha beta+ T cells. These preclinical results set the basis for a clinical study of ex-vivo gamma c gene transfer into CD34+ cells from SCID-X1 patients. PMID- 11265227 TI - beta-Amyloid protein aggregation: its implication in the physiopathology of Alzheimer's disease. AB - beta-Amyloid protein (A beta), a 39-42 residue peptide resulting from the proteolytic processing of a membrane-bound beta-amyloid precursor protein (APP), is one of the major components of the fibrillar deposits observed in Alzheimer patients. A beta fibril formation is a complex process which involves changes in A beta conformation and self-association to form cross-beta pleated sheets, protofibrils, and fibrils. Since the aggregation of soluble A beta peptide into fibrils is viewed as a critical event in the physiopathology of Alzheimer's disease (AD), preventing, altering, or reversing fibril formation may thus be of therapeutic value. This review will focus on the current state of knowledge of A beta fibril formation, with special emphasis on physiological and exogenous inhibitors which may have a therapeutic potential. PMID- 11265228 TI - [Role and functional spectrum of HPTLC in a hospital pharmaceutical quality control program]. AB - As part of the development of a quality assurance program (QAP), a high performance thin layer chromatography (HPTLC) analysis unit was installed in the pharmacy department at Gustave-Roussy. The HPTLC-CAMAG consists of: 1) an HPTLC Vario development chamber for optimization of the mobile phases; 2) TLC Sampler III automated sample applicators; 3) solid teflon migration chambers, i.e., horizontal tanks that enable separation to be carried out either in sandwich or in saturation mode; 4) a TLC Scanner 3 densitometer controlled by CATS 4 software; and 5) a Pentium MMX 233 MHz personal computer with an external backup unit. HPTLC quantitative and qualitative analysis has now reached a remarkably high level of development and performance. The samples (aqueous or non-aqueous solutions) that are to be processed are automatically applied by spraying (50-300 nl) in calibrated bands of a few mm (with up to 64 3-mm bands per 10 x 20 cm plate) on high-performance stationary phases and of wide technological diversity. The chromatogram is obtained in 10 min, and run over a migration pathway of 5-6 cm. The plates are read by absorption-reflection or fluorescence-reflection at an ad hoc wavelength (190-800 nm), then the peak areas which have been scanned are calculated by the trapezoid method. The calibration curves are generated by Michaelis-Menten non-linear regression, and validated by internal quality control. The analytical yield is high, i.e., up to 50 assays and 250 determinations per day. HPTLC analysis covers a wide functional range, and can be used in the following ways: 1) as a teaching tool for separative analysis and GLP; 2) it is an invaluable method for the optimization of mobile phases and for the determination of absorption spectra and absorption maxima, with a view to developing HPLC methods in complex matrices; 3) it provides major support for post-production quality control of prescribed hospital preparations of all types, e.g., those connected with parenteral nutrition, chemotherapy, synthetic narcotic analgesia; and it can also be used for dry dosage analysis; 4) it is useful in pharmaceutical assessment, e.g., in studies on the physico-chemical characteristics of various substances, such as their identity, purity, concentration, stability and compatibility, particularly with regard to generic products; 5) it can contribute to monitoring the safety of medical apparatus and equipment via the analysis of container-content interactions; 6) it provides a qualification system for personnel and procedures for within- and between-center validation of GMP. Setting up such an HPTLC quality control unit requires a basic investment of about 0.9 MF or 70,000 US dollars for a cost of no more than 10 F or 1.5 US dollars (including tax) per routine assay. After 18 months in operation and 16,500 assays, the HPTLC analysis unit has become one of the mainstays of the Gustave-Roussy QAP. PMID- 11265229 TI - [Mycophenolate mofetil: a new approach by immunosuppressive treatment]. PMID- 11265230 TI - Carboplatin is effective therapy for young children with progressive optic pathway tumors: a Pediatric Oncology Group phase II study. AB - The Pediatric Oncology Group conducted a phase II study to evaluate the activity of carboplatin in children 5 years or younger with progressive optic pathway tumors (OPTs). Of the 51 patients accrued to this study, 1 was not eligible because the child was older than 6 years. Fifty patients were eligible and had either neuro-imaging or symptomatic evidence of progressive OPTs. Twenty-one of 50 had evidence of neurofibromatosis type I (NF-1). Therapy consisted of carboplatin 560 mg/m2 at 4-week intervals. Patients with stable disease or better after two courses were continued on therapy for 18 months or until progressive disease. Of the 50 eligible children, 39 had stable disease or better, and 34 completed the 18-month therapy. Our data are sufficient to conclude that the proportion of objective responses (complete, partial, or minor response or stable disease) exceeded 30% (P < 0.00001), and the approximate 95% confidence interval estimate of the objective response rate was 0.665 to 0.895. Twenty-one patients went off protocol because of progressive disease. Fifteen patients progressed during the 18-month therapy, and 6 patients progressed after completing therapy. Six children died with progressive disease. Major toxicities were neutropenia and thrombocytopenia, and 3 children experienced allergic reactions. Carboplatin is active and safe for the treatment of young children with progressive OPTs. The addition of other potentially active drugs may further increase the event-free survival for these children. PMID- 11265231 TI - Effect of disease burden on health-related quality of life in patients with malignant gliomas. AB - The burden imposed by disease recurrence in patients with high-grade gliomas is not well documented. We studied the frequency of self-report symptoms and the effects on health-related quality of life in patients who had recurrent glioblastoma multiforme or anaplastic astrocytoma and who had a Karnofsky performance score > or = 70. Patients completed the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire-Core 30 Items (QLQ C30) and the Brain Cancer Module (BCM20) before initiation of treatment for first recurrence of disease. Six symptoms (fatigue, uncertainty about the future, motor difficulties, drowsiness, communication difficulties, and headache) were reported with a frequency > 50% by both groups of patients. An additional two symptoms (visual problems and pain) were also reported with frequencies of > 50% by patients with recurrent glioblastoma multiforme. Most of the symptoms were likely due to recurrence, but previous radiation therapy and on-going corticosteroid treatment may have also been casual factors for fatigue, whereas uncertainty about the future and pain were probably nonspecific for brain cancer. Problems with motor functioning, vision, leg strength, and pain were reported more frequently by patients with recurrent glioblastoma multiforme than by those with recurrent anaplastic astrocytoma. Scores on health-related quality-of-life functioning scales were similar in the two groups. Finally, the scores for patients who had recurrent high-grade gliomas and a Karnofsky performance score > or = 70 were compared with the reported health-related quality of life scores of patients with other cancers. Their scores were similar to those of patients with metastatic cancers and worse than those of patients with localized cancers. PMID- 11265233 TI - Primary Hodgkin's disease of the CNS in an immunocompetent patient: a case study and review of the literature. AB - Primary Hodgkin's disease limited to the CNS is exceedingly rare. Little is known regarding etiologic risk factors, optimal management, and prognosis. A case of Hodgkin's disease confined to the CNS, with cerebrospinal fluid negative for cytology, is described in an immunocompetent patient previously treated for hyperthyroidism with 131I. The patient underwent craniotomy, with resection of two lesions in close proximity within the parenchyma of the temporoparietal lobe. Histopathology revealed classic nodular sclerosing Hodgkin's disease, without evidence of Epstein-Barr viral infection. Treatment included radiation to the whole brain with a boost to the tumor bed. The patient made a full neurologic recovery and remains free of disease recurrence 21 months after treatment. A literature review has identified only 9 additional cases. Seven of 8 evaluable patients remain alive and free of recurrence with a median follow-up of 13 months. The risk factors for this presentation remain undefined. Although follow up is short, radiotherapy alone appears to provide excellent disease-free survival. Chemotherapy may be reserved for patients with positive cerebrospinal fluid, extracranial disease, or subsequent relapse. PMID- 11265232 TI - Posttransplant primary CNS lymphoma. AB - The records and neuro-imaging studies of 8 cases of posttransplant primary CNS lymphoma (PT-PCNSL) diagnosed at Mayo Clinic Rochester between 1970 and 1998 were reviewed retrospectively. All patients received organ transplantation. Patients who had hematologic transplantation were not included in the analysis. The median and mean age of the 4 men and 4 women was 45 years (range, 34 to 50 years). The median duration of symptoms before diagnosis was 36 days (range, 5 to 98 days). At diagnosis, the neurologic examination was focally abnormal in 6 of 8 patients. Compared with the initial computed tomographic study, MRI showed 25 additional brain lesions. Only 43.7% of lesions enhanced with contrast agent; of those that did, all but one were heterogeneous. Ependymal contact occurred in 5 patients. MRI lesion burden increased proportionally to the interval between scans. Diagnostic tissue was obtained by stereotactic biopsy from 6 patients and by open biopsy from 2. Hemorrhage occurred in the biopsy area in 4 patients who had stereotactic biopsy and 2 died (all had normal coagulation studies). Slides available for review (7 patients) showed that the tumors were of CD20-positive lineage and were positive for Epstein-Barr virus, using in situ hybridization. Six patients died. Median survival for the cohort was 13 weeks. PT-PCNSL has clinical and imaging features distinct from typical PCNSL. In our series, (1) PT PCNSL presented nonspecifically and progressed rapidly, (2) stereotactic brain biopsy had significant morbidity, and (3) despite multimodal therapy, survival was poor. PMID- 11265234 TI - Cell division. Location, location, location. PMID- 11265238 TI - Cell signalling. Ready, steady, go! PMID- 11265236 TI - Plant development. De-pipping the Pippin. PMID- 11265239 TI - Cell cycle. A complex twist for BRCA2. PMID- 11265240 TI - Transcription. Dial S for silence. PMID- 11265244 TI - Endocytosis. Tent pegs for clathrin. PMID- 11265245 TI - Phosphorylation. The key to staying faithful. PMID- 11265246 TI - Themes and variations on ubiquitylation. AB - Ubiquitylation--the conjugation of proteins with a small protein called ubiquitin -touches upon all aspects of eukaryotic biology, and its defective regulation is manifest in diseases that range from developmental abnormalities and autoimmunity to neurodegenerative diseases and cancer. A few years ago, we could only have dreamt of the complex arsenal of enzymes dedicated to ubiquitylation. Why has nature come up with so many ways of doing what seems to be such a simple job? PMID- 11265247 TI - Antigen processing by the proteasome. AB - The proteasome is an essential part of our immune surveillance mechanisms: by generating peptides from intracellular antigens it provides peptides that are then 'presented' to T cells. But proteasomes--the waste-disposal units of the cell--typically do not generate peptides for antigen presentation with high efficiency. How, then, does the proteasome adapt to serve the immune system well? PMID- 11265248 TI - Regulation of cellular polyamines by antizyme. AB - Proteins that are degraded by the proteasome are first modified by a set of enzymes that attach multiple copies of ubiquitin to substrate lysines, but a tiny minority, including the polyamine-synthesizing enzyme ornithine decarboxylase, is handled differently. This enzyme is targeted for destruction by another protein- antizyme. Why does ornithine decarboxylase have its own dedicated destruction mechanism, how does it work, and is it the only protein to be targeted to the proteasome in this way? PMID- 11265249 TI - Protein regulation by monoubiquitin. AB - Multi-ubiquitin chains at least four subunits long are required for efficient recognition and degradation of ubiquitylated proteins by the proteasome, but other functions of ubiquitin have been discovered that do not involve the proteasome. Some proteins are modified by a single ubiquitin or short ubiquitin chains. Instead of sending proteins to their death through the proteasome, monoubiquitylation regulates processes that range from membrane transport to transcriptional regulation. PMID- 11265250 TI - SUMO, ubiquitin's mysterious cousin. AB - Covalent modification of cellular proteins by the ubiquitin-like modifier SUMO regulates various cellular processes, such as nuclear transport, signal transduction, stress response and cell-cycle progression. But, in contrast to ubiquitylation, sumoylation does not tag proteins for degradation, but seems to enhance their stability or modulate their subcellular compartmentalization. PMID- 11265251 TI - Molecular dissection of autophagy: two ubiquitin-like systems. AB - Recent analyses of the genes required for autophagy--intracellular bulk protein degradation--in yeast have revealed two ubiquitin-like systems, both of which are involved in the membrane dynamics of the process. Molecular dissection of these systems is now revealing some surprises. PMID- 11265252 TI - Biochemistry and molecular biology teaching over the past 50 years. AB - The revolution in molecular biology that began nearly 50 years ago has had an enormous impact in the lab, but it also triggered a quieter revolution in undergraduate teaching. How has it affected the content and teaching methodology of undergraduate courses, and is the training that students now receive in the molecular life sciences appropriate to their future career paths? PMID- 11265253 TI - The future of education in the molecular life sciences. AB - The changing landscape of education in biochemistry and molecular biology presents many challenges for the future, for students and educators alike. The exponential increase in knowledge, the genomics, proteomics and computing revolutions, and the merging of once separate fields in biology, chemistry, physics and mathematics, mean that we need to rethink how we should be preparing today's science undergraduates for the future. What do we need to change, and how will we implement it? PMID- 11265254 TI - Characteristics of repeated ambulance use in an urban emergency medical service system. AB - BACKGROUND AND PURPOSE: Although many studies have examined the reasons for repeated use of emergency medical service (EMS), little information is available concerning repeated ambulance use in Taiwan. This study evaluated the characteristics of repeated EMS ambulance use in an urban EMS system in Taiwan. METHODS: Data from a local EMS computerized database for the period from January 1996 through December 1998 were collected for analysis. All calls to the dispatch center that resulted in EMS transports were included. Repeat users were identified by matching the user name, sex, age, and home address. Transports were categorized according to how many times the patient was transported by ambulance during the 3-year period: single use, one time; repeated use, two or three times; or frequent use, more than three times. RESULTS: During the 36-month study period, there were 41,792 calls, with 13,076 non-transports (a non-transport rate of 31.3%). Of the 28,716 transports during the study period, 2,101 represented repeated or frequent use (7.3%); the rate of frequent use was 1.4% (406/28,716). The frequency of repeated use reached a daily first peak at 8:00 AM, with the second and third peaks at 1:00 PM and 7:00 PM. The mean age increased with increasing repeated use of transport (37.25 +/- 0.24 vs 41.55 +/- 1.03 vs 46.23 +/- 1.57 years, respectively; p < 0.001). The percentage of non-trauma missions increased with increasing repeated use of transport (26.3% vs 55.6% vs 73.2%; p < 0.001). Response time significantly increased for repeated use (analysis of variance [ANOVA], p < 0.001). The on-scene interval (time from arrival until departure) in the single-use group was shorter than in the repeated and frequent use groups (ANOVA, p < 0.005). CONCLUSIONS: The results of this study indicate that the characteristics of repeat users of EMS transport differ from those of single users. Characteristics of service time, reason for transport, and interval to each subsequent call varied among different groups of users. Studies of repeat use under a wider range of conditions such as in rural EMS systems and after implementation of a priority-dispatch system are needed to determine the implications of repeated ambulance use. PMID- 11265255 TI - Disability-adjusted life years for breast cancer patients in Taiwan. AB - BACKGROUND AND PURPOSE: The incidence and mortality of breast cancer in Taiwan have increased rapidly in the past several decades, but the societal impact of deaths and disabilities due to breast cancer has not been assessed. This study estimated the disability-adjusted life years (DALYs) for breast cancer patients during 1994, and compared the results with similar data from other areas of the world. METHODS: DALYs for breast cancer patients in Taiwan were calculated using the equation developed by Murray and Lopez. The incidence and mortality of breast cancer and the population structure were obtained from national statistics maintained by the Department of Health and the Ministry of the Interior. The age specific mean survival time for breast cancer patients was estimated using the exponential distribution from incidence-mortality linkage of the incidence file at National Taiwan University Hospital and the National Mortality File maintained by the Department of Health. RESULTS: There were 11,963 years of life lost (YLL) due to breast cancer during 1994, 2677 years lived with disability (YLD), and 14,640 DALYs. The YLL and DALYs per 1000 population (1.17 and 1.44) were in the middle of the world spectrum, while the YLD value per 1000 population (0.26) was closer to those of developed countries. The proportion of DALYs contributed by younger patients (< 45 years) was higher than in developed countries and similar to those in developing countries other than Sub-Saharan Africa. The DALYs per 1000 population of women younger than 45 years of age in Taiwan were also higher than those in India, China, other regions of Asia and Islands, Sub-Saharan Africa, and the Middle Eastern Crescent. CONCLUSIONS: The disability portion (YLD) of the DALYs for breast cancer patients in Taiwan was higher than in other regions of the world. Moreover, patients younger than 45 years contributed a higher proportion of DALYs than in developed countries. The DALY value per 1000 population younger than 45 years of age was also higher than in developing countries. These results suggest that health professionals should focus more attention on programs for education, screening, and treatment of younger women. PMID- 11265256 TI - Preoperative diagnosis by electron beam computed tomography and perioperative management of primary tracheal anomalies in tetralogy of Fallot. AB - PURPOSE: To investigate the usefulness of electron beam computed tomography (EBCT) in the preoperative detection and perioperative management of insidious concomitant primary tracheobronchial anomalies in patients with tetralogy of Fallot (TF). METHODS: From July 1995 to October 1999, 88 children (38 girls, 50 boys) with TF were enrolled in this study. EBCT examinations provided information needed to plan management. The final diagnoses of airway abnormalities were correlated with the findings of bronchoscopy in five patients and confirmed during surgery. RESULTS: Fourteen (16%) of the 88 patients had associated primary tracheobronchial anomalies. Nine patients had tracheal bronchi, which were combined with tracheobronchial stenosis in two patients and with tracheal stenosis in one patient. Two patients had tracheal diverticulum, which was combined with lower tracheal stenosis in one patient. Two patients had congenital tracheal stenosis. Tracheomalasia was found in one patient. Three patients with ventilation difficulties died postoperatively. Special attention was given to the care of the diseased airways perioperatively, and the remaining 11 patients had a smooth course of hospitalization and discharge. CONCLUSIONS: Our results show that EBCT provides good delineation of both cardiac and tracheobronchial anomalies and suggest that it should be used perioperatively to detect associated airway anomalies in TF, to facilitate the design of an appropriate ventilation care strategy. PMID- 11265258 TI - Antigenic specificity of anti-neutrophil cytoplasmic antibody. AB - PURPOSE: To determine the antigens recognized by sera containing classic anti neutrophil cytoplasmic antibodies (c-ANCAs) and perinuclear anti-neutrophil cytoplasmic antibodies (p-ANCAs). METHODS: A total of 160 serum samples (all from a reference laboratory) that were originally collected from different clinics for ANCA tests were examined for c-ANCA and p-ANCA by indirect immunofluorescence (IIF). All positive sera were further tested for reactivity to proteinase 3 (PR3), myeloperoxidase (MPO), lactoferrin (LF), and lysozyme (LZ) by enzyme linked immunosorbent assay (ELISA). In addition, sera from 110 patients with systemic lupus erythematosus (SLE), 51 patients with rheumatoid arthritis (RA), and 40 healthy subjects were also tested for reactivity to these antigens. RESULTS: HF detected ANCA in 81 (51%) of the 160 clinical serum samples. Of these 81 serum samples, 21 (26%) contained c-ANCA and 60 (74%) contained p-ANCA. P-ANCA was more commonly found in antinuclear antibody (ANA)-positive sera than in ANA negative sera (p < 0.01). Of the 21 serum samples positive for c-ANCA, 12 (57%) reacted to PR3, four (19%) to LF, four (19%) to LZ, and three (14%) to MPO on ELISA. By contrast, of the 60 sera positive for p-ANCA, 15 (25%) reacted to MPO, 13 (22%) to PR3, eight (13%) to LF, and four (7%) to LZ. The prevalence of ANCA specificities in serum samples from SLE patients were as follows: anti-PR3, 0%; anti-MPO, 1%; anti-LF, 27%; and anti-LZ, 29%. The prevalence of ANCA specificities in serum samples from RA patients were as follows: anti-PR3, 6%; anti-MPO, 2%; anti-LF, 8%; anti-LZ, 4%. CONCLUSION: Sera positive for c-ANCA and p-ANCA reacted to diverse cytoplasmic antigens from neutrophils. P-ANCA was found in 55% of ANA-positive serum samples. LF and LZ were most commonly found in serum samples from patients with SLE. PMID- 11265257 TI - Increased soluble Fas ligand concentration in tuberculous pleural effusion. AB - BACKGROUND AND PURPOSE: Tuberculous (TB) pleurisy results from a delayed hypersensitivity reaction involving macrophages, T-cells, and many cytokines (including tumor necrosis factor, interferon-gamma, and interleukin 1 and 2). Infection by Mycobacterium tuberculosis induces apoptosis in gamma/delta T-cells and macrophages. Fas ligand (FasL) is a type II membrane protein that plays an important role in the regulation of apoptosis and has an intimate relation with these cells and cytokines. A soluble form of FasL (sFasL) exists in a variety of human body fluids, including serum, pleural effusion, cerebral spinal fluid, and ocular fluid. Therefore, we hypothesized that Fas activity is elevated in TB pleurisy. This study investigated the concentration of sFasL in TB pleural effusions and compared it with expression of sFasL in various other pleural effusions. METHODS: Using an enzyme-linked immunosorbent assay, we investigated the sFasL concentrations of 80 pleural effusions from patients with various diagnoses. RESULTS: The median sFasL concentration in the TB pleural effusion group was 104.91 pg/mL (n = 32). This was significantly higher than values in the transudate group (median value, 20.02 pg/mL, n = 9, p < 0.001) and patients with malignant effusion associated with adenocarcinoma of the lung (median value, 23.29 pg/mL, n = 14, p < 0.001). Lymphoproliferative disease could not be distinguished from TB based on sFasL concentrations in pleural effusion. CONCLUSIONS: The sFasL concentration in TB pleural effusions is significantly higher than that in adenocarcinomatous pleural effusions, which are the most common malignant pleural effusions. This difference may serve as a diagnostic tool to differentiate these two most commonly encountered unexplained pleural effusions. Determination of the cellular source and the actual role of the abundant sFasL in TB pleurisy will require further investigation. PMID- 11265259 TI - Intellectual outcome of patients with congenital hypothyroidism detected by neonatal screening. AB - BACKGROUND AND PURPOSE: Mental retardation is a major sequela of delayed treatment for congenital hypothyroidism; congenital hypothyroidism can be treated early if detected with neonatal screening. We evaluated the intellectual outcomes of 62 patients with congenital hypothyroidism detected by neonatal screening at a major teaching hospital in northern Taiwan. The effects of thyroid pathology, age at the initiation of treatment, socioeconomic status, and severity of hypothyroidism on intellectual outcome were also analyzed. METHODS: All patients had euthyroid status at the time of intelligence testing. The Chinese Fourth Revision of the Binet-Simon Scales was used to evaluate the patients' intelligence between the ages of 3 and 6 years. RESULTS: The mean intelligence quotient (IQ) score was 102 +/- 18. Only four of the 62 patients were mentally retarded. Patients with lower initial serum thyroxine concentrations (T4; < 2 micrograms/dL) at the time of diagnosis of congenital hypothyroidism had significantly lower IQs (95 +/- 19, n = 26) than those with higher initial T4 concentrations (106 +/- 16, n = 36; p < 0.05). Patients with fewer than three ossification centers had lower IQs (91 +/- 20, n = 12) than those with three or more (104 +/- 17, n = 36; p < 0.05). Significantly lower IQs were also found in patients with a smaller femoral epiphysis area (< 0.1 cm2) (92 +/- 20, n = 15) than in those with larger epiphyses (106 +/- 15, n = 21; p < 0.05). The type of pathology (ectopia, athyrosis, dyshormonogenesis), age at the start of treatment (before or after 30 days of age), and socioeconomic status did not significantly affect the intellectual outcome. CONCLUSIONS: Our results indicate that intellectual outcome in Taiwanese patients with congenital hypothyroidism has been improved by neonatal screening and that the severity of hypothyroidism at diagnosis is the most important prognostic factor affecting intellectual outcome in these patients. PMID- 11265260 TI - Alstrom syndrome in two siblings. AB - Alstrom syndrome is a very rare autosomal recessive inherited disorder. Only 50 cases have been reported since the syndrome was first described in 1959. This syndrome is characterized by obesity, impaired glucose tolerance with insulin resistance, retinal degeneration, neurosensory deafness, acanthosis nigricans, hepatic dysfunction, and some endocrine disorders. The index case of this report was a 12-year-old girl who became blind at the age of 6 years as the result of progressively impaired vision. At the age of 12, diabetes mellitus was diagnosed and acanthosis nigricans presented in the neck, axilla, and groin regions. Her 10 year-old brother had similar symptoms. Electroretinography and audiometry disclosed generalized pigmentary epithelial change, decreased to absent cone and rod responses, and moderate sensorineural hearing loss in both siblings. Biochemistry and oral glucose tolerance tests showed diabetes mellitus, dyslipidemia, and hepatic dysfunction in the index case. Elevations of insulin, C peptide, and leptin concentrations were found in both siblings. Insulin resistance was also demonstrated in both siblings using the modified insulin suppression test with constant infusion of somatostatin and exogenous insulin. PMID- 11265261 TI - Use and abuse of surgical antibiotic prophylaxis in hospitals in Taiwan. AB - BACKGROUND AND PURPOSE: A large proportion of antibiotics used in hospitals is for surgical prophylaxis. We determined the prevailing practices and factors associated with the misuse of surgical antibiotic prophylaxis in hospitals in Taiwan. METHODS: In a systematic survey of the medical records of 629 patients from 14 hospitals who underwent clean or clean-contaminated surgery from September 1998 through March 1999, data on the timing and duration of, and reasons for, antibiotic use were collected and analyzed. RESULTS: Overall, 578 (92%) patients received antibiotics perioperatively; in 499 (79%) cases, antibiotics were used for surgical prophylaxis. Only 302 (61%) patients received prophylaxis within 1 hour before surgery. More than 70% of patients received more than 3 days of postoperative antibiotic prophylaxis. The most commonly used antibiotics were first-generation cephalosporins and aminoglycosides. Factors independently associated with inappropriately timed prophylaxis included surgery performed at a hospital with a greater proportion of intensive care unit beds (conditional odds ratio [OR] = 1.14 per 1% increase, 95% confidence interval [CI95%] 1.06-1.23; p < 0.01), surgery duration of at least 1 hour (OR = 0.40, CI95% 0.24-0.67; p < 0.01), orthopedic surgery (OR = 0.59, CI95% 0.36-0.98; p = 0.041), and cardiothoracic surgery (OR = 2.07, CI95% 1.14-3.77; p = 0.02). Risk factors for more than 3 days of prophylaxis included surgical placement of prosthetic material (OR = 2.26, CI95% 1.10-4.64; p = 0.03), the number of antibiotics used (OR = 1.99 per antibiotic, CI95% 1.26-3.13; p < 0.01), surgery duration of at least 1 hour (OR = 3.07, CI95% 1.45-6.51; p < 0.01), neurosurgery (OR = 4.57, CI95% 2.24-9.31; p < 0.01), and the use of oral antibiotics together with intravenous drugs (OR = 20.72, CI95% 10.72-40.07; p < 0.01). CONCLUSIONS: The results of this survey indicate that inappropriate use of surgical antibiotic prophylaxis is common in hospitals in Taiwan. Our results identify several problem areas, including incorrect timing, duration, and use of oral antibiotics for surgical prophylaxis, that require targeted physician education and public health interventions. PMID- 11265262 TI - Central venous catheter-induced atrial ectopic tachycardia with reverse alternating Wenckebach periods. AB - A centrally inserted venous catheter may cause atrial ectopic tachycardia. The association of atrial ectopic tachycardia with spontaneous reverse alternating Wenckebach periodicity has rarely been reported. We describe a 4-year-old boy with tetralogy of Fallot who developed atrial ectopic tachycardia with reverse alternating Wenckebach periods postoperatively after central venous catheter placement. All such episodes emerged from a 3:2 atrioventricular block, followed by runs of 2:1 atrioventricular block with progressive shortening of the conducted PR intervals. Normal sinus rhythm returned after the catheter was withdrawn to the superior vena cava. Reverse alternating Wenckebach periodicity may be a tachycardia-dependent physiologic phenomenon. PMID- 11265263 TI - Combined atrial and arterial switch operations for congenitally corrected transposition. AB - Conventional repair of congenitally corrected transposition of the great arteries (CCTGA) is directed at eliminating the associated defects and leaves the right ventricle in a systemic position. The long-term outcome of this procedure may involve deterioration of right ventricular function with tricuspid regurgitation and failure of the conduction system. We describe two consecutive patients with CCTGA, one of whom had apicocaval juxtaposition. The patients were aged 19 and 16 months, respectively, and both underwent a combination of atrial and arterial switch. These are the first two reported cases of successful completion of this type of operation in Taiwan. Our review of previously reported cases suggested that no significant difference exists in the outcome of patients with this condition who undergo either arterial switch or Rastelli-type repair plus atrial redirection. However, reported patients who underwent anatomic repair had lower early mortality, late mortality, and incidence of complete heart block than those who underwent conventional repair. The present two cases and our review of the literature suggest that, among patients with apicocaval juxtaposition, 1) Mustard operation is optimal for patients with small atrial volume; 2) one-and-one-half ventricular repair may be helpful to the outcome, especially when treatment is combined with Rastelli-type repair; and 3) excellent access to the ventricular septal defect through the tricuspid valve is afforded via a left atriotomy. From the present two cases and our review of the literature, we conclude that anatomic repair is superior to conventional repair of CCTGA in terms of protection against dysfunction and failure of the anatomic right ventricle, tricuspid valve, and conduction system. Long-term follow-up is mandatory. PMID- 11265264 TI - Growth pattern of renal angiomyolipoma on computed tomography: report of two cases. AB - The incidence of renal angiomyolipoma is increased in patients with tuberous sclerosis. Renal angiomyolipomas frequently exhibit both intratumoral and subcapsular or perirenal hemorrhage when they grow into the extrarenal space. Evidence of serial morphologic change is crucial to decision-making for surgical exploration in patients with growing renal angiomyolipomas. We report two cases of angiomyolipoma of the kidney with expansion of an angiomyolipoma into the subcapsular region. Neither patient had tuberous sclerosis and both developed bleeding when the mushroom-shaped tumor reached the extrarenal space. These cases suggest that expansion of the tumor into the subcapsular region is indicative of impending rupture of a renal angiomyolipoma. PMID- 11265265 TI - Recurrent cardiac myxoma with multiple distant metastasis and malignant change. AB - A 37 year-old female underwent open heart surgery for a left atrial myxoma. The post-operative course was uneventful and she was discharged two weeks later. She had regular monthly follow-up in the outpatient department until 10 months postoperatively when she was readmitted to the orthopedic ward for excision of a left ankle tumor. Two days after admission, she developed severe orthopnea. The initial diagnosis was heart failure, and she was transferred to the medical ward for treatment. Transthoracic and transesophageal echocardiography revealed a recurrent left atrial tumor. Because of acute obstruction of the mitral valve and deterioration of her condition, she underwent emergent open heart surgery. The recurrent atrial tumor was excised; histopathologic examination revealed a myxoid sarcoma. Multiple tumors were found on this admission, including a mass in the neck and in the left forearm; computed tomography revealed a brain tumor in the left posterior frontal lobe and a chest wall tumor. She died two months later. Recurrent cardiac myxoma with multiple distant metastasis may have a malignant potential. Because of the potential for tumor recurrence, long-term and regular follow-up is mandatory. PMID- 11265266 TI - Perceptions of change in the health care industry. AB - Cost containment, governmental regulations, and managed care are leading factors driving change in the delivery of health care services. Job insecurity and stress are frequently felt by the allied health professional in times of such change. This investigation measured the perceptions of change in the health care industry. A survey tool measuring attitudes and beliefs about the state of health care was distributed to health professionals in Arkansas. Voluntary responses were coded using a four-point Likert scale. Data were processed from an overall response rate of 34.6% (n = 203) using descriptive frequencies, analysis of variance (ANOVA), and a post-hoc Scheffe test to determine significances of differences between professional groups. Differences were found between professional groups' perceptions of the overall quality of health care, job security, and having a sense of control to positively impact the future of health care. When dealing with changes, professionals must maintain a balance of ethics and efficacy. They need to be agents of change, promoting quality and efficiency. Their role should be to shape health care, not simply to react to change. PMID- 11265267 TI - Practice autonomy among primary care physician assistants: the predictive abilities of selected practice attributes. AB - The practice autonomy of primary care physician assistants (PAs) is of interest to those organizing, financing, and delivering health services. This study examined the predictive abilities of practice attributes with respect to multidimensional aspects of practice autonomy (clinical decision making and prescriptive authority) in primary care PAs. A sample of 225 practicing PAs was used to construct the 16-item Physician Assistant Autonomy of Practice Instrument (PAAPI), which includes three subscales, routine prescriptive authority, advanced prescriptive authority, and clinical decision making. All were used as dependent variables in multiple regression analyses. The most significant correlates of practice autonomy included years in practice as a PA, years in practice with supervising physician, annual income from practice, recognition as the exclusive primary care provider for patients, primary practice in a rural county, and primary employment setting (single-specialty group practice). More primary care PAs continue to be used in under-served rural areas and in managed care. Organizational structure of the work setting may influence these PAs' practice autonomy. PMID- 11265268 TI - Respiratory therapists and critical-thinking behaviors: a self-assessment. AB - The purpose of this study was to assess critical-thinking behaviors of respiratory therapists through self-report. Using a quantitative survey research method, respiratory therapists rated themselves on seven critical thinking skills. The effects of personal variables on the self-assessments were also investigated. The respiratory therapists self-assessed their critical-thinking behaviors highest in the categories of prioritizing, troubleshooting, and communicating. Anticipating was self-assessed as the lowest-ranked critical thinking behavior. Age and educational level were found to have no effect on the self-assessed behaviors, while years of experience in respiratory care and gender were found to affect self-assessed troubleshooting, decision making, and anticipating. The results of this study suggest that educators and clinicians should consider learning strategies that incorporate the use of experience when targeting novice practitioners. PMID- 11265269 TI - Issues and dilemmas of developing a new faculty practice plan. AB - Due to changes in health care and increased knowledge of the public concerning health care, faculty need to assume a more active role in practice settings. Faculty involvement in the practice of their disciplines allows them to remain on the "cutting edge" as well as be models for their students. Most faculty practice plans will provide some external funding but more importantly will allow additional educational opportunities for the students as they work with the faculty. These plans allow faculty to interact with the community and strengthen the ties of the community and the academic institution. However, there are dilemmas in establishing faculty plans, for the quality of teaching cannot be threatened, and most universities expect their faculty also to be involved in scholarly activities. This article discusses 1) the process used in planning and developing a faculty practice plan; 2) details of the plan, to include faculty incentives, administrative cost, etc.; and 3) strategy for implementation. PMID- 11265270 TI - Distance learning in the health professions: on the verge of collapse or poised to soar? AB - Distance education provides universal access to education. While the issue of access to education is seemingly resolved, the question "what is the best way to teach?" remains. Thomas Jefferson espoused the creation of the academic village to foster broad, intensive scholarship. John Dewey, Ralph W. Tyler, Malcolm S. Knowles, and Alexander W. Astin echo Jefferson's ideals. To ensure excellence in distance learning, a disciplined rethinking of teaching and a reordering of academic priorities is essential. If consumer confidence in the academy is to be restored, there must be a return to collegial leadership in: defining institutional purpose and resource commitment, improving teacher competence, honing curricular content, perfecting interactive learning, selecting students, and designing outcome measures for both teaching and learning. Then, and only then, will distance learning be ready for "prime time." PMID- 11265271 TI - Physical therapy as primary health care: public perceptions. AB - This study investigated the public's knowledge of direct access and the role of physical therapists, and whether the public would consider using a physical therapist for primary care. Persons living in South Florida were selected at random by dialing telephone numbers. Using three-digit telephone number prefixes, four-digit suffixes were generated by rolling dice. When consent was obtained, the respondents' answers were recorded on a self-generated questionnaire. No knowledge of direct access was reported by 67.3% of the sample. Additionally, 57.4% of the sample had never been to a physical therapist. A substantial number of respondents (73.4%) stated that they would go directly to a physical therapist. Thus, the public poorly understands direct access and the role of the physical therapist. The members of the public might use physical therapists as primary care practitioners if they were aware of this option. PMID- 11265272 TI - Efficacy of telemedicine in occupational therapy: a pilot study. AB - The purpose of this pilot study was to determine the efficacy of using a less costly, low-bandwidth telemedicine system to evaluate occupational therapy clients in rural areas. Four residents (aged 63 +/- 10.2 years) from rural Tillery, North Carolina, were evaluated via either The Kohlman Evaluation of Living Skills or The Canadian Occupational Performance Measure. An occupational therapist in Tillery administered one of these tools, while a second occupational therapist at East Carolina University simultaneously scored the same tool by telemedicine link. Comparison of their responses revealed scoring differences in only one of four administered evaluations. Video images were insufficient for visualization of finer movements, but audio quality was excellent. The authors conclude that select occupational therapy evaluation data can be accurately transmitted and properly scored using low-bandwidth telemedicine systems. PMID- 11265273 TI - How do skilled nursing rehabilitation managers track efficiency and costs? AB - Financial and efficiency indicators have not been used extensively by skilled nursing (SNF) rehabilitation managers, but may prove useful in the prospective payment system (PPS)-dominated long-term care (LTC) environment. The purpose of this pilot study was to demonstrate a method for measuring the extent of SNF rehabilitation managers' use of volume, revenue, cost, and manpower indicators; and whether usages differ among occupational, physical, and speech therapy managers. The subjects were 74 occupational, 75 physical, and 72 speech therapy managers employed by a multinational health care corporation. A total of 221 Likert scale surveys were mailed to SNF rehabilitation managers to obtain data on use of 32 efficiency and financial indicators. The 32 indicators were in four categories: visit volume, revenue, costs, and manpower utilization. Twelve indicators were used regularly by at least 60% and three were used regularly by 85-100% of respondents. The response rate was 17.89%. Chi-square analyses were performed for therapy discipline and response frequency. None was significant at 0.05. Three of the payermix indicators demonstrated post-hoc significance in "regular" or "sometimes" use by the physical therapy managers. The results show discipline-specific differences among LTC rehabilitation managers in the extents of efficiency and financial indicator usage for decision making. Suggestions for future use of the survey instrument are provided. The method proved useful for demonstrating differences in the combinations and degrees of use of the 32 indicators. The results point to a need to educate SNF rehabilitation managers in cost-monitoring techniques. Recommendations for cost monitoring are provided. PMID- 11265274 TI - Salmonella enteritidis, Denmark. PMID- 11265275 TI - Wild poliovirus imported into Qinghai province, China. PMID- 11265277 TI - Ribonuclease inhibitors. PMID- 11265279 TI - Assays for the evaluation of HIV-1 integrase inhibitors. PMID- 11265278 TI - Producing soluble recombinant RNases and assays to measure their interaction with interferon-gamma in vitro. PMID- 11265280 TI - Analysis by HPLC of distributive activities and the synthetic (back) reaction of pancreatic-type ribonucleases. PMID- 11265281 TI - The direction of ribonucleases H by antisense oligodeoxynucleotides. PMID- 11265282 TI - The 2-5A/RNase L pathway and inhibition by RNase L inhibitor (RLI). PMID- 11265283 TI - Crystallization and crystal structure determination of ribonuclease A ribonuclease inhibitor protein complex. PMID- 11265284 TI - Methods for studying the interaction of barnase with its inhibitor barstar. PMID- 11265285 TI - Assaying in vitro refolding of RNases by mass spectrometry. PMID- 11265286 TI - Dissecting nucleases into their structural and functional domains. Mapping the RNA-binding surface of RNase III by NMR. PMID- 11265287 TI - Using electrostatics to define the active site of Serratia endonuclease. PMID- 11265288 TI - Ire1p: a kinase and site-specific endoribonuclease. PMID- 11265289 TI - Homology modeling and simulations of nuclease structures. PMID- 11265290 TI - Methods for determining activity and specificity of DNA binding and DNA cleavage by class II restriction endonucleases. PMID- 11265291 TI - Processivity of DNA repair enzymes. PMID- 11265292 TI - Engineered properties and assays for human DNase I mutants. PMID- 11265293 TI - Assays for human DNase I activity in biological matrices. PMID- 11265294 TI - Evaluation of retroviral ribonuclease H activity. PMID- 11265295 TI - Assays for detection of RNase A superfamily ribonucleases. PMID- 11265296 TI - Assay for antitumor and lectin activity in RNase homologs. PMID- 11265297 TI - Microtiter-plate assay and related assays for nonspecific endonucleases. PMID- 11265298 TI - Isolation and enzymatic activity of angiogenin. PMID- 11265299 TI - Preparation and preclinical characterization of RNase-based immunofusion proteins. PMID- 11265300 TI - Restriction endonucleases and their uses. PMID- 11265301 TI - Isolation and characterization of an unknown restriction endonuclease. PMID- 11265302 TI - Gene modification with hapaxoterministic restriction enzymes. Easing the way. PMID- 11265303 TI - Techniques to measure nucleic acid-protein binding and specificity. Nuclear extract preparations, DNase I footprinting, and mobility shift assays. PMID- 11265305 TI - Quantitating mRNAs with relative and competitive RT-PCR. PMID- 11265304 TI - Analyzing the developmental expression of sigma factors with S1-nuclease mapping. PMID- 11265306 TI - Detection and quantitation of mRNAs using ribonuclease protection assays. PMID- 11265307 TI - Expressing self-incompatibility RNases (S-RNases) in transgenic plants. PMID- 11265308 TI - Molecular cloning, tissue distribution, and chromosomal localization of the human homolog of the R2/Th/Stylar ribonuclease gene family. PMID- 11265309 TI - Introduction to DNA sequencing. PMID- 11265310 TI - Cycle sequencing. PMID- 11265311 TI - Cycle sequencing of polymerase chain reaction-amplified genomic DNA with dye labeled universal primers. PMID- 11265312 TI - Automated fluorescent DNA sequencing on the ABI PRISM 377. PMID- 11265313 TI - The universal primers and the shotgun DNA sequencing method. PMID- 11265314 TI - Automated fluorescent DNA sequencing on the ABI PRISM 310 Genetic Analyzer. PMID- 11265316 TI - Fluorescent sequencing for heterozygote mutation detection. PMID- 11265317 TI - Sequence databases and the Internet. PMID- 11265315 TI - Fluorescent sequencing protocols for the ALF. PMID- 11265318 TI - DNA sequencing by capillary array electrophoresis. PMID- 11265319 TI - M13 sequencing. PMID- 11265320 TI - Primer design and primer-directed sequencing. PMID- 11265321 TI - Direct sequencing of DNA produced in a polymerase chain reaction. PMID- 11265322 TI - Solid phase fluorescent sequencing of the CFTR gene. PMID- 11265323 TI - Shotgun DNA sequencing. PMID- 11265324 TI - [Mediastinal teratomas--diagnosis and therapy]. AB - The authors present an account on patients with a teratoma of the mediastinum who were operated at the Second Surgical Clinic, L. Pasteur Faculty Hospital in Kosice. In the course of 10 years (Jan. 1, 1990-Dec. 31 1999) 73 patients with tumours of the mediastinum were operated. In four the diagnosis of teratoma of the mediastinum was confirmed by histological examination (5.47%): three adult patients and one child. In two patients the tumour of the mediastinum was diagnosed accidentally during X-ray examination of the chest. In one female patient surgical revision was indicated on account of a relapse of the process. In the conclusion the authors emphasize that teratomas of the mediastinum are frequently asymptomatic, and in case the process is in the anterior or upper mediastinum, teratomas must be taken into account and removed as a whole during surgical intervention. PMID- 11265325 TI - [Cytokines in the diagnosis of peritoneal inflammation]. AB - Early diagnosis of peritoneal inflammation in the initial stage with equivocal clinical manifestations is not always simple. At present there are in the literature many new laboratory indicators which to a different extent describe the inflammatory process. In the present paper the authors focused attention on the practical importance of the use of cytokines in the diagnosis of peritoneal inflammations. They compare the diagnostic impact of clinical examination as compared with assessed plasma concentrations of inflammation promoting cytokines. The authors maintain that the diagnosis of secondary peritonitis is based on the clinical examination of the patient. Elevated values of PCT or IL-6 can support this clinical conclusion. In postoperative peritonitis dynamic investigation of cytokines is important. The diagnosis of tertiary peritonitis is based on laboratory findings. PMID- 11265326 TI - [Postoperative immunosuppression--a physiological process and source of complications. Minireview]. AB - Postoperative immunosuppression reflects non-specific neurohumoral stress responses to surgical trauma. Surgical stress in the early stage is characterised by a dysbalance of the adrenocortical response, the index of the formation of glucocorticoids and dehydroepiandrosterone is shifted to the glucocorticoids. This non-specific hormonal response has an impact on the balance between TH1 and TH2 lymphocyte response. Inhibition of the TH1 response occurs, the clinical reflection of which is a decline of cell-mediated immunity. Postoperative immunosuppression and dysbalance of cell-mediated and humoral immunity are physiological processes, part of the neurohumoral stress response to surgical trauma. Its potential pathological consequences, although proved so far only experimentally, make it on the one hand an important factor for evaluation of the risk of postoperative complications and on the other hand a possible of object of therapeutic intervention in oncological or high risk patients. PMID- 11265327 TI - [Polytetrafluoroethylene vascular prostheses and patches in pediatric cardiac surgery]. AB - The author reviews the indications for the use of vascular grafts, patches, membranes and stitches from polytetrafluoroethylene (PTFE) in pediatric cardiac surgery. Vascular grafts are used for construction of arterial shunts in neonates with complex cyanotic heart defects, for palliative reconstruction of interrupted aortic arch, for construction of extraanatomic aortic by-pass and total cavopulmonary connection. The PTFE patches are suitable for closure of septal defects, plastic reconstruction of the right and the left ventricular outflow tracts, correction of coarctation of the aorta and plastic repair of stenoses on the main pulmonary artery and its branches. PTFE stitches can be used for plastic reconstruction of atrioventricular valves. Membranes represent very good pericardial substitutes after complex surgeries with valve replacement or the use of a valved conduit, where reoperation must be suspected. In neonates and small children with signs of heart failure after surgery, it is possible to use PTFE membranes for transient wound cover when the chest is left opened. PMID- 11265328 TI - [Aneurysm of the superior mesenteric artery in a child]. AB - A case report of the ruptured aneurysma of the superior mesenteric artery at 12 year-old girl which was successfully treated by resection and substitution by means of venous graft. There is illustrated good long-term result more than 7 years after operation. Superior mesenteric artery aneurysma diagnosis in childhood is very rare. PMID- 11265329 TI - [Extracorporeal membrane oxygenation in the treatment of severe pulmonary hypertension in a neonate after surgery for laparoschisis]. AB - The authors describe the case of newborn with laparoschisis in whom severe idiopathic pulmonary hypertension during postoperative period developed and initiation of extracorporeal membrane oxygenation (ECMO) to maintain circulatory stability and adequate oxygenation was necessary. ECMO was performed for 75 hours with maximum extracorporeal support up to 50% of cardiac output (Biomedicus pump BP 50, Jostra oxygenator M8). Patient was successfully weaned and switched to conventional ventilation and nitric oxide inhalation with consequent extubation. No bleeding complications were observed during ECMO in connection with surgical repair of laparoschisis. PMID- 11265331 TI - [Ileus and intestinal perforation in premature infants--current trends in diagnosis and treatment]. AB - OBJECTIVE OF STUDY: a) assessment of the relationship between ileus of premature infants and the development of intestinal perforation in premature infants, b) suggestions of an optimal therapeutic procedure. In the study children (n = 50) are included a) with intestinal perforation in conjunction with impaired excretion of meconium (n = 22), b) with an ileus state based on obstruction of the ileum by a viscous meconium treated either surgically or conservatively (n = 28). Surgical treatment involved: a) establishment of a double ileostomy (n = 28), b) insertion of a T drain into the terminal ileum (n = 8), c) removal of meconium from the gut and its primary closure (n = 2). Conservative treatment in 11 children involved irrigography with an liquid contrast substance under X-ray control. The group of children with perforation was compared with the group of children without perforation, risk factors were evaluated by statistical methods. The necessity of ventilation (P = 0.051) and gestation age (P = 0.006) proved to be statistically significant risk factors for the development of perforation. Survival was not influenced by perforation. All 11 children treated conservatively survived, of 39 operated children 26 survived (66.7%). An early start of conservative treatment of ileus of premature infants reduces markedly the risk of intestinal perforation and can thus influence the survival of low birth weight neonates. PMID- 11265332 TI - [A kidney harvesting technic from a non-heart beating donor]. AB - The authors describe the technique of collection of kidneys from NHBD. They present their own protocol elaborated in Plzen for collection of kidneys from NHBD. For collection they use a special double-balloon catheter inserted into the aorta to ensure that the perfusion fluid will penetrate into visceral branches of the aorta. For the flow of the perfusion fluid a urinary catheter is used inserted via the femoral vein into the vena cava inferior. Immediately after the beginning of perfusion laparotomy is performed with immediate cooling of the kidney with ice. The authors give an account of their own experience and discuss further technical possibilities as regards collection of kidneys from NHBD. PMID- 11265330 TI - [Abdominal lymphangiomas in childhood]. AB - The authors evaluate their experience with the diagnosis and treatment of abdominal lymphangiomas during the period of 1995-1999. During the above period 6 girls and 4 boys with abdominal lymphangiomas were operated. The mean age at the time of operation was 5 years and 8 months. Ultrasonographic examination was made in all 10 patients, CT examination in 8. The surgical finding was lymphangioma of the mesenterium 4x, of the omentum 2x, of the adrenals 2x of the retroperitoneum 1x and intestinal lymphangiomatosis 1x. Macroscopically complete extirpation of the lymphangioma was possible in 9 patients and called for resection of the gut 3x and for adrenalectomy 2x. In one patient subtotal extirpation was performed. Postoperative follow-up did not reveal a relapse in any of the children. Abdominal cystic lymphangiomas are rare benign malformations of the lymphatic system. For preoperative differential diagnosis in typical cases USG is sufficient. The therapeutic method is complete surgical extirpation which is associated with a low incidence of relapses. PMID- 11265333 TI - [Donors with non-beating hearts--a new possibility in expanding the kidney donor program]. AB - In the Czech Republic in recent years a steady decline of renal transplantations was recorded due to shortage of organ donors. The authors present initial experience with the collection of organs from dead non heart beating donors (NHBD). In the course of less than two years this programme was introduced within the framework of a clinical experiment (grant IGA MofH CR ND/4781-3) at the Surgical Clinic, Faculty Hospital in Plzen. A total of 20 kidneys were collected from NHBD and another 12 kidneys were not collected, most frequently because of difficulties in communication and organization. The donors were grouped according to the Maastricht classification. The mean age of donors was 43.3 years, the mean period of warm ischaemia was 22 minutes and the serum creatinine level was 145 mumol/l. Of 20 collected kidneys a normal histological appearance was recorded in 14, the remainder had different degrees of damage due to inadequate rinsing. Based on hitherto assembled experience the authors emphasize perfect team collaboration in collection from NHBD and a precise technique of collection. The next stage of the clinical study will be the preparation proper for clinical transplantation of kidneys from NHBD. PMID- 11265334 TI - [Xanthogranulomatous pyelonephritis presenting as a Grawitz tumor in a 21-year old female patient. Case report] ]. AB - Xanthogranulomatous pyelonephritis is a chronic inflammatory disease of the kidney characterized by destruction and replacement of the renal parenchyma with granulomatous tissue containing lipid-filled macrophages. The disease affects mainly elders, more frequent females and is difficult to differentiate it from renal neoplasms by standard diagnostic imaging methods. This case report confirms it. PMID- 11265335 TI - [Valtrac--anastosmoses of the digestive tract] ]. AB - The author presents a brief review of anastomoses of the digestive tract using a biofragmentable BAR ring. A group of 300 anastomoses is discussed implemented from the beginning of February 1994 till May 2000. PMID- 11265336 TI - [Diagnosis and treatment of pelvic and perineal defects and fistulae and reconstructive surgery using a gluteus maximus flap]. AB - In the form of a comment some aspects of diagnosis and treatment of unhealed pelvic and perineal defects and fistulas were analysed. Based on author's own experience with the treatment of 2 females with postradiation rectovaginal fistulas and 2 males with perineal sinus after proctectomy the authors discuss some advantages and technical aspects of pedunculated myoplasty by the gluteus maximus muscle. PMID- 11265337 TI - [Reconstructive gluteus maximus muscle pedicled flaps in the treatment of resistant fistulae of the pelvic organs]. AB - The author describes a surgical method used to close poorly healing defects between the vagina and rectum, and the urethra and rectum resp. or persistent sinuses after proctectomy by means of a muscular flap of the gluteus maximus muscle. The results of treatment are demonstrated on a group of 7 patients operated by the author in the course of 26 years. In six patients the procedure was successful and led to healing of the defects. PMID- 11265339 TI - [Angio-organic ischemic syndromes--etiopathogenesis and differential diagnosis]. AB - The authors analyze in the submitted review the classification, etiopathogenesis and in particular the differential diagnostic principles of the main obliterating arterial diseases which condition angio-organ syndromes. The most frequent disease which causes angio-organ ischaemic syndromes worldwide is at present atherosclerosis which however is not the only nosological unit, and in clinical practice it is important to consider also other organically conditioned diseases of the vascular system. In addition to ischaemic hypoxia it is important to consider also all types and varieties non-angiogenic (non-vascular) hypoxia. In the management of angio-organ ischaemic syndromes the surgeon (angiosurgeon) must collaborate in particular with the angiologist because vascular diseases are conditioned by many risk factors of a medical character and their elimination and treatment are within the competence of internal medicine and its specialized branch--angiology. PMID- 11265338 TI - [Use of oxicellulose in neurosurgery--technical comments]. AB - In different surgical disciplines preparations which promote haemostasis are widely used. The latter comprise in particular preparations on the basis of collagen (e.g. Actifoam, Avitene), gelatin (Gelfoam, Spongostan) and oxidized celluloses (Traumacel, Surgicel). The application of these preparations in neurosurgery is very specific and has its laws. At the Neurosurgical Clinic, Faculty Hospital Brno the authors used since 1995 for brain surgery in particular Traumacel, i.e. oxidized cellulose in powder form. Since 1996 the author uses in brain surgery Surgical, extradurally Spongostan. The author summarizes his experience with the use of Surgical in 114 operations of the brain. He analyzes specific features of the use of Surgical in different diagnoses and its effectiveness in the prevention of haemorrhagic complications. At present the use of Surgical is an integral part of haemostasis in the majority of open operations of the brain in the author's department. PMID- 11265340 TI - [Cervical pulmonary hernia]. AB - Pulmonary hernia is in general a rare affection--in particular in the cervical region. Symptoms are as a rule slight and are encountered only in exceptional cases. The diagnosis is based on physical examination and imaging techniques. The authors submit a case-history of a cervical pulmonary hernia in a young woman. It was noteworthy because of marked complaints of the patient which involved professional damage. Therefore it had to be treated by surgery which was successful. PMID- 11265341 TI - [Role of peroperative coloscopy]. AB - The authors focus attention on the method of preoperative endoscopy which is useful in some operations of the digestive tract and evaluate retrospectively its contribution in their own department. They compare data on this method in the professional literature and consistent with them they recommend preoperative endoscopic examination in indicated cases as a safe, rapid and effective method for improving the standard of surgical care of patients and recommend this method in the algorithm of treatment. PMID- 11265342 TI - [Medical ethics at the turn of the second and third millennium]. PMID- 11265343 TI - [Methods of intraperitoneal placement of mesh in laparoscopic surgery of inguinal hernias (IPOM--Intraperitoneal Onlay Mesh)]. AB - BACKGROUND: Intraperitoneal onlay mesh is a new method of laparoscopic inguinal hernia repair shortening an operative time and reducing its cost, which is very safe and less burdening a patient. METHODS: 325 males and 37 females at the mean age of 47.3 years were operated on electively for symptomatic inguinal hernia by IPOM method. In all cases we used a polyester mesh impregnated with silicone. RESULTS: The mean operative time was 18 minutes for unilateral procedure and 28 minutes for bilateral one. There was no serious complication during the operation even the post-operative time and no recurrency following this type of the operation. CONCLUSION: IPOM hernioplasty can be a method of choice in the inguinal hernia repair because of this procedure is very simple, cheap and safe for a patient. We have to wait for a long-term results to validate the hitherto existing ones, even if they are very optimistic. PMID- 11265344 TI - [Results of laparoscopic implantation of peritoneal catheters]]. AB - The advantages of miniinvasive surgical techniques were proved in a number of surgical operations. In the submitted article the authors evaluate five years' experience with laparoscopic implantation of Tenckhoff's catheter for peritoneal dialysis. The authors implanted in 1995-1999 catheters by the laparoscopic route in 34 patients. The most frequent complication was early leak of the dialysate along the catheter (41%). After modification of the surgical technique early leak was observed in 11% of the patients. Escape of the catheter from the lesser pelvis was observed in 14.8% patients. The authors did not observe early infection of the tunnel along the catheter. It may be concluded that the laparoscopic technique of implantation of a peritoneal catheter was not associated with a higher incidence of complications than the laparotomy. The advantage of laparoscopic operation is earlier mobilisation and shorter hospitalization of the patient. Another advantage of this technique, tested by the authors, is the possibility of exact diagnosis possibly with an immediate single-stage plastic operation of hernias in the abdominal region. PMID- 11265345 TI - [Dislocation of the humerus--diagnosis and the Arlt method of reduction]. AB - Authors present 248 shoulder dislocations included recurrent dislocations in a 3 year period. All of dislocations were of anterior type except 3 posterior dislocations and 1 inferior or luxatio erecta. They describe and recommend method of reduction without local or general anesthesia according to Arlt, which was used in 234 cases with effectivity of 96.6%. Posterior dislocation can have a very subtle signs and is generally initially missed in more than 50% cases. Two X ray views in posterior dislocation are necessary to confirm diagnosis. Authors used in addition to anteroposterior transthoracic view with no posterior dislocation missed. PMID- 11265346 TI - [Complications of synthesis using the Medin sliding compression screw]. AB - In 1993-1999 in the Traumatological Hospital in Brno 141 syntheses DHS Medin were made in patients with fractures of the proximal femur. (Eighteen DHS syntheses of other manufacturers were also made. The latter were however not analyzed). During the primary operations twice revision was necessary on account of a technical mistake of the surgeon ad twice an early infection developed. Seven late complications occurred--4 times a pseudoarthrosis and three times an aseptic necrosis of the head of the femur. All complications were resolved by total replacement of the hip joint. In no instance failure of the implant was recorded. PMID- 11265347 TI - [Injuries of the cervical spine in patients with ankylosing spondylitis]. AB - A brittle and rigid, osteoporotic and injury prone "bamboo" spine which develops as a result of transformation of the normal spine in the final stage of Bekhterev's disease contains normal neural elements. Unstable fractures occur frequently even after a minimal injury and the morbidity and mortality is significantly higher than in the group of routine injuries of the cervical, spine, in particular as a result of pulmonary complications. Diagnosis of the fracture based on simple X-ray pictures is difficult, CT examination is essential. The authors present an account on their experience with three patients. They draw attention to problems of skeletal traction when the spinal axis is altered by disease and to risks associated with semiconservative treatment. The authors therefore recommend traction with a minimal load and with subsequent early decompression and stabilization by a combined anterior and posterior approach in a single session. By this approach secondary affection of neural structures in unstable fractures can be prevented, and early rehabilitation with verticalization without restriction of respiratory excursions by an orthesis then significantly reduces the risk of pulmonary complications. PMID- 11265348 TI - [Doppler scanning in aneurysm surgery]. AB - Subarachnoid hemorrhage still represents important medical problem, from both, ineffective prevention and high morbidity and mortality rate of the condition. On the other hand, majority of patients who survive initial hemorrhage and successful clipping of their aneurysm can live without major medical restrictions. Apart from surviving the initial attack the most important condition is properly clipped aneurysm. Doppler scanning control allows precise placement of the clip on the neck of an aneurysm. Since January 1997 through December 1999 the authors have surgically treated 204 aneurysms in 181 patient. In 40 surgeries the Doppler scanning was employed after the aneurysm complex was dissected free. Using 20 MHz PW Doppler probe the flow within the aneurysm and in the involved arteries was measured (usual depth 1.5 mm, range 1-2.5 mm). After the clipping the flow was evaluated again, to ensure proper exclusion of an aneurysm and good flow in involved arteries. The authors consider the Doppler scanning to be important technical aid in aneurysm surgery, especially in anatomically complex situations. PMID- 11265349 TI - Long-term management of cirrhosis. Appropriate supportive care is both critical and difficult. AB - Orthotopic liver transplantation has emerged as an important treatment option for patients with advanced liver disease. However, each year the number of new cases of cirrhosis exceeds the number of livers available for transplantation by a factor of 5 to 10. This translates into long waiting lists and restrictive criteria for selecting transplant recipients. Until advances in surgical technique or biotechnology increase the availability of organs for transplantation, the majority of patients with advanced liver disease will have to be managed medically for years--perhaps indefinitely. Early consultation with a liver transplant center can be helpful. The transplant hepatologist and surgeon can help with triage decisions, guide workup, provide advice about patient care, optimize the timing of transplantation, offer specialized diagnostic and therapeutic options, and help the treating physician stay abreast of the continuous changes in this complex field. In the final analysis, however, it is often the skill and diligence of the primary care physician in diagnosing liver disease, identifying and treating correctable causes, optimizing the patient's health and nutrition, and anticipating and preventing catastrophic complications that determine whether the patient lives or dies. PMID- 11265350 TI - Site-specific techniques of joint injection. Useful additions to your treatment repertoire. AB - Joint and soft-tissue aspiration and injection procedures are relatively easy to master once the physician has acquired an understanding of the techniques and subtleties of performing these procedures at certain sites. These techniques can be used to diagnose and treat many painful joint and soft-tissue conditions and are useful additions to the primary care physician's treatment repertoire. PMID- 11265351 TI - What caused this verrucous plaque? PMID- 11265352 TI - Hematologic and oncologic emergencies. Doing the most good in the least time. AB - Broad categories of emergency hematologic and oncologic situations are metabolic crises, compressions and obstructions, and symptomatic cytopenias. In each instance, a decision to intervene should be made on the basis of findings on diagnostic assessment in combination with prognostic information. Management should be directly proportional to the possibility for cure, significant remission, or improved quality of life. Numerous diseases, including potentially curable cancer, can be modified and patients' quality of life substantially improved with appropriate emergency intervention. Fortunately, the modern therapeutic arsenal provides many specific measures to manage these challenging clinical situations. PMID- 11265353 TI - HRT debate continues in eastern Europe. PMID- 11265354 TI - Domestic violence screening rate leaves much to be desired. PMID- 11265355 TI - Making a decision about ERT/HRT. Evidence to consider in initiating and continuing protective therapy. AB - Alzheimer's disease, CAD, and osteoporosis significantly affect the health and well-being of senior citizens in the United States. The fact that women have a longer life expectancy than men has led to the hypothesis that estrogen in some way imparts protection against these disease processes. Available data on the possible negative effect of estrogen on the development and progression of Alzheimer's disease are provocative but inconclusive. Thus, for the time being, they must remain no more than the basis of an attractive hypothesis. In contrast, available data suggest that ERT and HRT can reduce the risk of CAD, but this effect seems more preventive than therapeutic. Addition of a progestational agent to an estrogen regimen may blunt this effect. Although the medical literature contains very few data that address the issue of duration of therapy, logic would suggest that cessation of therapy would result in the loss of a protective effect. With regard to osteoporosis, ERT and HRT have clear beneficial effects in that they increase BMD and decrease fracture risk. There is good evidence that duration of therapy may be more important than dosage and that these effects rapidly dissipate with cessation of therapy. Finally, as with all medical interventions, ERT or HRT must be individualized for each patient. Although actual health hazards are few, adverse effects are common and the emotion charged, ever-evolving issue of the negative impact of ERT and HRT on breast cancer risk must always be considered before such therapy is instituted. PMID- 11265356 TI - The enigma of whiplash injury. Current management strategies and controversies. AB - How to diagnose and treat whiplash injuries, particularly in chronic cases, is one of the most controversial topics in medicine. Current diagnostic approaches cannot fully define the pathophysiologic mechanisms of these injuries, and traditional treatment regimens often fail to completely relieve symptoms. Dr Young reviews current management strategies and controversies and discusses potential methods of dealing with chronic cases. PMID- 11265357 TI - When to measure BMD in patients being treated for osteoporosis. PMID- 11265358 TI - Constipation. PMID- 11265359 TI - Let's have a meeting--at your place and mine. Videoconferencing can keep you out of those crowded airports. PMID- 11265360 TI - Postherpetic pain control with concomitant illness? PMID- 11265361 TI - Slowing the progression of CHF. Drug therapy to correct neurohormonal abnormalities. AB - As the population ages, the number of cases of congestive heart failure (CHF) is expected to climb. Primary care physicians will be increasingly called upon to treat patients with this serious cardiac derangement. In this article, Drs Ward and Anderson discuss the latest approaches to treatment, which are based on the current understanding that CHF results from left ventricular dysfunction, which causes a complex activation of multiple neurohormonal reflexes. PMID- 11265362 TI - Beta blockers for CHF. Adrenergic blockade dramatically reduces morbidity and mortality. AB - Several large clinical trials have shown that beta blockers can reduce morbidity and mortality in patients with CHF. Therefore, current guidelines for treatment of CHF now include beta blockers as standard therapy for patients with left ventricular systolic dysfunction (ejection fraction < or = 40%) and mild to moderate heart failure. Beta-blocker therapy for CHF should be started cautiously and increased gradually to avoid exacerbating symptoms of heart failure. At this time, data for therapy in patients with NYHA class I or IV symptoms are limited, and it is unclear whether all beta blockers confer benefit or whether some are better than others. Several trials are under way to answer these questions. Until more evidence is available, only those agents that have proved beneficial in mortality trials should be used to manage CHF. PMID- 11265363 TI - Surgical treatment of chronic heart failure. What to tell patients about heart saving options. AB - Patients with severe, chronic heart failure can be managed by several surgical techniques that lead to cure for some or provide a bridge to heart transplantation for others. Although transplantation is currently the only proved curative therapy for end-stage heart failure, the supply of donor hearts has not kept pace with the demand. Therefore, procedures such as reduction ventriculoplasty, transmyocardial laser revascularization, or dynamic cardiomyoplasty and the use of assist devices or artificial hearts hold promise for helping patients maintain heart function until a cure can be offered. PMID- 11265364 TI - Treatment for coccidioidomycosis in pregnancy? PMID- 11265365 TI - Pulmonary diagnostic quiz. Deciphering a breathtaking emergency. Pulmonary edema. PMID- 11265366 TI - What do the new antimicrobials offer? Weighing the advantages and disadvantages compared with traditional agents. AB - The newest antimicrobial drugs have broader spectrums of activity than their predecessors. But are they better than traditional antimicrobial agents? Several drugs recently approved by the US Food and Drug Administration (FDA) may be more effective against uncommon and antimicrobial-resistant pathogens, but their superior efficacy in treating common infections has yet to be proved. Dr Gleckman discusses the advantages and disadvantages of these new drugs and how they can best be used to avoid fostering the emergence of drug-resistant pathogens. PMID- 11265367 TI - The case for an ancient cause. Healthy People 2010. PMID- 11265369 TI - [45-year "Experiences in medicine" Professor Dr. of Medicine Jan Bozydar Latkowski]. PMID- 11265368 TI - Necrobiosis lipoidica. Indolent plaques may signal diabetes. PMID- 11265370 TI - [Historical outline of the Otolaryngologic Academic Medical Departments and Clinics in Lodz, particularly from 1989-2000]. PMID- 11265371 TI - [Theories attempting to explain the significance of the paranasal sinuses]. AB - The author presents 12 theories which try to evaluate the significance of paranasal sinuses. A vast amount of references on this subject and additionally this paper do not explain the role of paranasal sinuses in human. In the author's opinion the most probable is the amortizative and constructive theory based on antropometric studies and experimental head traumas. PMID- 11265372 TI - [Remarks concerning didactics based on personal experiences]. AB - The author described own experiences concerning didactics in a medical school, particularly elements of practical training in otolaryngology. PMID- 11265373 TI - [Incidence of pharyngocutaneous fistulas after laryngectomy depending on the method of nutritive drain insertion]. AB - In the article incidence of pharyngocutaneous fistulas after total laryngectomy was estimated depending on the method of nutritive drain insertion. The study was performed in 464 patients (64% of all laryngectomized) treated surgically in ENT Department of Medical University in Lodz from 1988 to 1997. It was stated that fistulas developed in 114 cases (24.6%). There were compared two groups of patients following laryngectomy: group I was consisted of 279 patients with nasogastric tube and group II numbered 163 subjects with using a different method of nutritive drain insertion by the upper pole of neck wound during laryngectomy. It was found that pharyngocutaneous fistulas developed statistically lower in group with drain inserted by the neck wound than in group with nasogastric tube (17.8% vs 27.9% p < 0.05). Moreover patients in the second group could not suffered from the unpleasant pain in the nose, and easier accepted this manner of nutrition. Satisfactory surgical aspects for the patients and their environment confirm usefulness of different method of insertion of the nutritive drain by the neck wound. PMID- 11265374 TI - [Evaluation of CD44 adhesion molecule, nm23 gene product expression and intensity of angiogenesis in patients with laryngeal cancer]. AB - One of the most important factor in prognosis of the patients with laryngeal cancer is presence of the metastases in lymph nodes of the neck. The main purpose of the paper was the evaluation of CD34 and FVIII antigens as angiogenesis markers, and nm23 protein and CD44 antigen expression as metastasis potential markers and description of their role in the tumour progression and making metastasis in the patients with laryngeal cancer. Paraffin-embedded tissue sections from 89 patients with laryngeal cancer were stained with a monoclonal antibody raised against CD34 and FVIII antigens, against nm 23 protein and against CD44 antigen. Measuring the density of the microvasculature in tumour was investigated. We found significant dependence between intensity of angiogenesis (IA) and pT, nodal metastasis, histological grading and survival. There were also significant correlation between nm23 protein expression and nodal metastasis, and between CD44 antigen expression and pT, nm23 protein expression and FVIII antigen expression. Evaluation of mentioned markers allowed to asses the aggressiveness of tumour cells and anticipate neck metastasis in the patients with laryngeal cancer. PMID- 11265375 TI - [Expression of selected markers for apoptosis, proliferation and metastasis in evaluation of laryngeal cancer invasiveness dynamics]. AB - Evaluation of the biology of laryngeal cancer cell is connected either with many process inside the cell or reactions between cancer cell itself and extracellular matrix. The main purpose in this paper was the evaluation of p53 protein, bcl-2 protein, Ki-67 antigen and CD44 adhesive molecule expressions in comparison to clinical and histopathological features in patients with laryngeal cancer. Paraffin-embedded tissue sections from 89 patients with laryngeal cancer were stained with a monoclonal antibody raised against p53 and bcl-2 proteins, Ki-67 and CD44 antigens using a peroxidase-labelled streptavidin-biotin kit. There were statistically significant relationships between p-53 protein over-expression and pT, histological grading, survival and Ki-67 and CD44 antigens expressions. There were no correlation between bcl-2 protein expression and clinical and histopathological features. We observed statistically significant correlation between Ki-67 expression and pT, histological grading, recurrences and survival. Expression of CD44 statistically significant correlated only with tumour size. We conclude that comparison of data covering mentioned tumour markers expression gives valuable evaluation of biological activity of cancer cells and may allow to create the immunological panel of tumour markers which simplify the prognosis about nodal metastases, recurrences and survival in patients with laryngeal cancer. PMID- 11265376 TI - [Audiobase--a computer system for monitoring and analyzing audiometric data in clinical and ambulatory practice]. AB - The amount of data in the field of otorhinolaryngology has rapidly increased in proportion to the growing number of patients. It is a very important issue to preserve their records and make them easily available. Authors present a computer based system for monitoring audiometric data of hearing impaired persons named Audiobase. The main elements of this system as: the personal data, otoscopic and audiometric examination have been described. This system was created to be used by staff without computer skills and therefore is largely "icon-driven". The main functions include patient record creation, update, and retrieval, as well as the generating of reports and graphical presentations. The system has already provided useful research material and is now beginning to fulfill an even more important role in patient follow-up and in evaluation of alternative treatment protocols. PMID- 11265377 TI - [Function of sensorimotor equilibrium function in elderly patients on the basis of vestibular evoked myogenic potentials (m.p.p.w.)]. AB - Observed elderly had general weakness of motor and mental function. The slower reactions to environment stimulus in elderly above 60 years result first of all from weakness of sensory receptors as visible, hearing, smell and taste. In this age group disturbing of balance system function demonstrated vertigo, dizziness, darkness of visible, disequilibrium was more encountered. Up to now there is not specified that "presbyequilibrium" (presbyvertigo, presbyastasis) as physiology ageing process causes this condition or destructive processes, infections, vascular insufficiency and neoplastic processes in inner ear and acoustic tracts and centres. Vestibular symptoms often are related to age by the doctors so for that reason basic and sometimes dangerous disease may be missed. Proposition of this paper is estimation of reflex function of balance system in connection with ageing process. To this end VEMPs were used and latencies and amplitudes of ipsilateral and contralateral VEMP recordings were calculated in young and elderly groups. Lower reflex efficiency in elderly disclosed longer latencies and lower aptitudes of p14 n21 of VEMPs than young people but without significant differences. PMID- 11265378 TI - [Evaluation of sensorineural hearing loss by late response of vestibular evoked myogenic potentials]. AB - The aim of this paper was assessment of usefulness of MPPW recording in diagnosis of hearing loss. Study of MPPW performed in 65 people, in these 22 cases with normal hearing (44 ears), 20 subjects with sensorineural hearing loss (27 ears), 10 examined with retrocochlear hearing loss (12 ears) and 10 patients with unilateral totally deafness. Differences of latency wave n34 p44 (later response of VEMPs) were stated in sensorineural and retrocochlear hearing loss with reference to latencies this wave in normal hearing subjects. Vestibular organ lesion may be not only diagnosed based on VEMPs results but also cochlear organ lesion can. PMID- 11265379 TI - Speech rehabilitation using a voice prostheses following laryngectomy. AB - The most serious consequence for patients following laryngectomy is the restriction of verbal communication. Since the introduction of laryngectomy significant concerns have already been focused on the field of speech rehabilitation. The operational procedures for the speech rehabilitation include training of the oesophageal voice speech and the voice prostheses. Speech prostheses are available in our hospital since 1983. The speech quality of the speech prostheses is compared with the classical oesophageal voice or to the voice by means of a Provox speech help. Bacteriological and mycological colonisation as a function of the length of implantation are defined. Our approach to the voice rehabilitation after a laryngectomy by use of a spacer during the laryngectomy has proven successful. As a result patients do not fall into a "hole" of non verbal communication. The aim of our efforts is always to create a functioning oesophageal voice after leaving the care of the hospital. PMID- 11265380 TI - Laryngeal chondrosarcoma with regard to the ultrastructure. AB - Most common among the laryngeal sarcomas is the chondrosarcoma. The difficulties in differentiation between benign and malignant behavior of cartilaginous tumors are responsible for the danger of misinterpretation of chondrosarcoma as chondroma. A case of a laryngeal chondrosarcoma is presented examined by light and electron microscopy to determine if the ultrastructure of chondrosarcoma could be helpful in the correct diagnosis. The cells and their formation play a more important role than the extracellular matrix in the differentiation of tumor's behavior. As special criterias of malignancy by electron microscopy are the presence of dominant mitochondria, fat vacuoles, dilated rough endoplasmatic reticulum and the irregular shape of chondrocytes. The knowledge of the ultrastructure of chondrosarcomas may be helpful to distinguish between a benign chondroma and low-grade chondrosarcoma, especially when only small tumor biopsies are valuable. PMID- 11265381 TI - [Manometric evaluation of disorders in oral and pharyngeal phases of swallowing in patients after partial laryngectomy for localized supraglottic cancer]. AB - Manometric examinations of swallowing were conducted on 81 patients after partial laryngectomy and on 35 subjects being a control group. Resection of piriform recess, a part of the base of the tongue, the hyoid bone or its part is the factor that causes intensified difficulty during swallowing and increase in the frequency of the occurrence of aspiration. The results of manometric examinations indicate that the shape and mobility of the tongue and the mobility of remaining after the surgery parts of the larynx have the greatest influence on the efficient swallowing in patients who have undergone partial laryngectomy due to cancer initially located in the supraglottic area. The larynx mobility is closely related to the remaining of the hyoid bone. The importance of remaining the possibly non-deformed structure and mobility of the tongue during partial laryngectomy involves the issue of reconstruction of defects occurred during the surgery. Manometric examinations confirm the effectiveness of the method involving reconstruction of defects in a part of the base of the tongue with a vascular pedicle flap of the submandibular gland. PMID- 11265382 TI - [Usefulness of chromosome instability analysis for selecting proper therapy of laryngeal cancer]. AB - A decision concerning of larynx cancer therapy is dependent on such factors as tumour location, its size, histology and occurrence of metastasis. The aim of the paper is to turn attention on usefulness of individual sensitivity of genetic information on mutagen-induced damage. The considerations are based on own author's experience and the literature data. Mutagen sensitivity is estimated by bleomycin test that is a measure of individual resistance to mutagens and carcinogens. The review is focused on DNA and chromosome sensitivity to radiation induced damage. The attempts to individualize radiotherapy basing on cytogenetic and molecular studies are presented. PMID- 11265383 TI - [Development of second primary malignant neoplasms in patients treated for laryngeal cancer in the Otolaryngology Clinic of the University School of Medical Sciences in Poznan 1981-1999]. AB - Second malignancies are the important cause shorting survival of patients with laryngeal cancer. We noted 51 patients with second malignant neoplasm after treatment for laryngeal cancer. The incidence of second neoplasms in lung, next in tonsils and the incidence of cancer of skin was the highest. Early detection of precancerous lesions would correct the prevention against second malignancies and by this way--correct the survival among patients with laryngeal cancer. The rate of incidence of the second primary tumors among patients after total laryngectomy with radiotherapy was 2.9%. PMID- 11265384 TI - [Expression of p53 antigen in laryngeal carcinoma]. AB - The results of immunohistochemical investigations of p53 presence in 50 patients with laryngeal carcinoma were presented. In the whole investigated group the presence of p53 protein in 90% of all investigated cases was observed. In patients with advanced clinical stage of laryngeal carcinoma higher expression of p53 protein comparing with patients with lower clinical stages was more frequently observed--but it wasn't significant. No significant difference in presence of p53 protein among the patients with a different stage of histological differentiation of carcinoma and among the patients with present and absent metastatic changes in regional lymph nodes was observed. PMID- 11265386 TI - [The value of endoscopic ultrasound examination for tongue neoplasms]. AB - The aim of the paper was to assess the value of endoral ultrasound in tongue tumors diagnostics. It was performed by the evaluation of the tumor advancement and its comparison with palpation, assessment of neoplastic tissue infiltration, detection of satellite foci surrounding the main tumor mass. Endoral ultrasound vs transcutaneous ultrasound was compared. Follow-up by means of transoral ultrasound was applied. The limitations of the method were presented. 65 patients with tongue tumors, treated in ENT Department K. Marcinkowski University School of Medical Sciences between 1995-1999 were under study. 9 patients with suspicion of local recurrence were included to the examined group. Not able value of the method was proved. In 81.5% of cases the method occurred to be much more precise than palpation and revealed the bigger dimension of the tumor then stated clinically. The satellite foci and small lesions, not possible to be detect by means of transcutaneous ultrasound, were find out and thanks to it treated radically. This method allows to differentiate the scarred and neoplastic tissue, especially in patients with clinical suspicion of relapse. The limitations of the method: trismus, big extension of the lesion, surface bleeding and gag reflex are to be listed. PMID- 11265385 TI - [A case of neuroma in the sublingual region]. AB - Neuromas are being mesenchymal tumors which are very rare in the mouth. The authors presented the case of the tumour of this localisation of 46-year old female, treated in the Department of Otolaryngology University School of Medical Sciences in Poznan, with suspicion of sublingual cyst. PMID- 11265387 TI - [Otoacoustic emission examinations in soldiers before and after shooting]. AB - Firearms are a common source of impulse noise that may potentially damage hearing organ. The purpose of this study was evaluation of the click evoked otoacoustic emission (TEOAE) and distortion-product otoacoustic emission (DPOAE) before and after shooting and comparison with conventional pure tone audiometry. Standard pure tone audiometry, tympanometry, TEOAE and DPOAE were made before and 10-15 minutes after shooting. Ten male soldiers (20 ears) were exposed to impulse noise from automatic gunfire (15 single rounds of live ammunition). They did not use any earplugs. The reduction amplitude of the TEOAE after shooting was found especially for the frequency 3,4 kHz for the right ear and 1 and 2 kHz for the left ear. The DPOAE greatest reduction concerned frequency 2,5 and 4 kHz for the left ear. All our shooters were right-handed and probably the asymmetrical effect resulted from the shooting posture. Any differences existing between the audiometric threshold before and after shooting were not noticed. Clinical experience with OAE indicates that it may play a role as a screening method for the soldiers exposed to noise and as a tool for monitoring early changes in cochlea. Emissions seem to be more sensitive for monitoring early cochlear changes after shooting than pure tone audiometry. PMID- 11265388 TI - [The usefulness of ultrasonic examination for diagnosis of congenital lateral and medial neck cysts]. AB - In the study results of ultrasound examination of 169 patients treated in ENT Department in Poznan with clinical suspicion of congenital neck cyst were analysed. It was made methodic evaluation of ultrasound examination and confrontation of ultrasound attributes characteristic for each type of cyst- complicated and uncomplicated by inflammation. Results were compared with intraoperative. PMID- 11265389 TI - [Vascular compression syndrome of the vestibulocochlear nerve--otolaryngologic and radiologic diagnosis]. AB - Vascular compression syndrome is the term used to classify a group of conditions though to be caused by the compression of cranial nerve by vessel. In the most cases the contact of vascular loop formed by the anterior inferior cerebellar artery (AICA) with the eight and facial nerve correlated with unilateral auditory symptoms or hemifacial spasms. The vascular compression syndrome of vestibulocochlear nerve in 8 patients treated in I ENT Clinic of Silesian Academy and MRI Department in Katowice was observed. All patients were otologic findings such as a tone audiometry, ENG, ABR and radiological diagnostics included MRI and angio MRI. The prospective analysis was performed. The results suggest that the unilateral sensorineural hearing loss, tinnitus, vestibular disorders and positive findings on magnetic resonance imaging are the most reliable evidence for the presence neurovascular compression syndrome of the eight cranial nerve. The MRI and otologic studies provided quite detailed information about topography of relationship between the blood vessels and cranial nerves in the crebellopontine cistern. PMID- 11265390 TI - [Evaluation of cochlear function by means of otoacoustic emission in children treated with cytostatic drugs in the course of bone marrow proliferative diseases]. AB - The aim of study was cochlear function estimation by means otoacoustic emission in children treated by leukaemia cytostatic drugs in course of the bone marrow proliferative diseases. The children after treatment of acute lymphoblastic leukemia or nongranulomatic malignant lymphoma according to BFM programme were examined in this study. Audiologic examination of children included tonal audiometry, tympanometry and otoacoustic emission as well. The results were compared to the findings of control group consisted of healthy children have never suffered from ear diseases and have never used ototoxic drugs. No significant differences in tonal audiometry and TEOAE were observed. Aggressive treatment of leukaemia does not result in hearing loss. PMID- 11265391 TI - [Noise from a car airbag as a cause of acute acoustic trauma]. AB - A case of acute acoustic trauma in 32 year male, caused by a noise from car airbag, was described. The symptoms occurring directly after the airbag shot were tinnitus and hearing loss in right ear. Patient was admitted to ENT Clinic in Gdansk just after 5 months after an accident. Treatment--vasodilatators and oxygen hyperbaric therapy--was ineffective. Review of literature, concerning the side effects of car airbag shot, was made. A mechanism of airbag action was presented as a source of short time noise. PMID- 11265392 TI - [Leopold Schroetter von Kristelli and Polish laryngologists]. AB - The researches of Leopold Schroetter von Kristelli, Karl Stoerk, Johann Schnitzler, Ottokar Chiari, Markus Hajek and others, the best Viennese rhinolaryngologists clarified our knowledge on rhinolaryngology. Leopold Schroetter, the famous rhinolaryngologist and internist, was a head of rhinolaryngological clinic in Vienna. He is shown as a precursor of treatment tracheas stenosis, he was also the author of many scientific papers and several books, coeditor of scientific medical magazines, a good teacher of a great number of physicians also from Polish territories. The Polish doctors: Samuel Meyerson, Wladyslaw Rudnicki and Alfred M. Sokolowski were a reporters from Schroetter's clinic. PMID- 11265393 TI - [Report from the 63rd Czech Congress of Otolaryngologic Head and Neck Surgeons. Olomuniec, June 7-9, 2000]. PMID- 11265394 TI - [Sentinel lymph node biopsy in breast cancer surgery--a review of the literature]. AB - Sentinel lymph node biopsy for breast cancer is reviewed herein. It has been recently investigated at many hospitals and institutes, mainly in Europe and the USA. Two lymphatic mapping methods using radiolabeled colloids and dyes are the most popular for detecting the target nodes, and seem to become a standard method of sentinel biopsy. A learning curve for identifying sentinel lymph nodes is important in improving the accuracy of detection. However, it should also be noted that a certain number of false negative cases are usually found. Furthermore, a highly reliable and suitable method to examine lymphatic metastasis has not yet been established. Large clinical trials need to be performed to confirm the optimum method for this new treatment for breast cancer. PMID- 11265395 TI - [Efforts to prevent adverse events in the United States--health care risk management and a fresh perspective on adverse events prevention]. AB - Not causing adverse events is never-ceasing issue in the health care field. However, the advances and greater specialization of medical technologies and the increasing number of elderly people, are all factors in the occurrence of adverse events. At the same time, greater efficiency is now demanded in the health care field, and the problem of preventing adverse events has become tougher than ever before. Given the situation, a fresh perspective on attempts to prevent adverse events may be important. One hint for such a new perspective is the health care risk management that is widely practiced in the health care field in the United States. This was introduced in the mid-1970s to counter the disputes and lawsuits at the time, but over the years the focus has shifted to the importance of prevention, and is now recognized as a means to work toward the assurance of quality of health care. Hints are also found in the suggestions related to adverse events prevention. In "To Err Is Human," published in November 1999 in the United States, includes proposals to "respect human limits in process design" and "promote effective team functioning," which are just the approaches we should adopt for a new perspective. I would also like to draw attention to the idea that there should be investigations into "developing effective mechanisms for identifying and dealing with unsafe practitioners" and the importance of "protecting voluntary reporting systems" that is mentioned. Adopting American methods unchanged to the health care system in Japan may not be appropriate, but the way of thinking and know-how from health care risk management, as well as the suggestions for adverse events prevention will provide us new perspectives on adverse events prevention, from which we should work toward a system of more efficient, and high-quality adverse events prevention. PMID- 11265396 TI - [The approaches to factors which cause medication error--from the analyses of many near-miss cases related to intravenous medication which nurses experienced]. AB - Given the complexity of the intravenous medication process, systematic thinking is essential to reduce medication errors. Two thousand eight hundred cases of 'Hiyari-Hatto' were analyzed. Eight important factors which cause intravenous medication error were clarified as a result. In the following I summarize the systematic approach for each factor. 1. Failed communication of information: illegible handwritten orders, and inaccurate verbal orders and copying cause medication error. Rules must be established to prevent miscommunication. 2. Error prone design of the hardware: Look-alike packaging and labeling of drugs and the poor design of infusion pumps cause errors. The human-hardware interface should be improved by error-resistant design by manufacturers. 3. Patient names similar to simultaneously operating surgical procedures and interventions: This factor causes patient misidentification. Automated identification devices should be introduced into health care settings. 4. Interruption in the middle of tasks: The efficient assignment of medical work and business work should be made. 5. Inaccurate mixing procedure and insufficient mixing space: Mixing procedures must be standardized and the layout of the working space must be examined. 6. Time pressure: Mismatch between workload and manpower should be improved by reconsidering the work to be done. 7. Lack of information about high alert medications: The pharmacist should play a greater role in the medication process overall. 8. Poor knowledge and skill of recent graduates: Training methods and tools to prevent medication errors must be developed. PMID- 11265397 TI - [What we must learn from recent medical accidents--experiences at the investigation committee]. AB - The current publicity of medical accidents in Japan started with two well-known accidents. On January 11, 1999, a cardiac patient underwent a pulmonary operation, while a pulmonary patient received cardiac surgery at Yokohama City University Hospital. On February 11, 1999, detergent was inadvertently injected into a venous line at Hiroo Hospital in Tokyo. The author was invited as an indifferent specialist to serve on the Investigation Committee for these two accidents. This article describes the lessons to be learned from these two accidents. PMID- 11265398 TI - [To promote hospital activities for the patient safety]. AB - Recently, increasing news reports of adverse events in health care suggest the need for reconsideration of the role and responsibility of health care in society. In this paper, after reviewing the history and the characteristics of the health care delivery system in Japan, central issues of patient safety management are sorted out. A good example of a team approach that has prevented progress of a serious disease is also presented. It is suggested that the total quality improvement activities are useful for the patient safety management. PMID- 11265399 TI - [Risk management and clinical path]. AB - In the risk management of medical practice, developing an efficient system to prevent medical accidents and trouble is required. For this purpose it is important to promote a cooperative team practice, to clarify each staff member's role, for each staff member to accurately carry out that role, for each staff member to always try to understand the patient's condition, and for staff members to ask each other whenever in doubt as to a patient's condition or the medical practice itself. These points for risk management share the same goal as the clinical path. The introduction of a clinical path is useful from the viewpoint of risk management. Electronic medical records and computer ordering systems in the medical practice should be developed as tools to decrease medical accidents. PMID- 11265400 TI - [Clinical result of thermochemoradiotherapy for advanced head and neck cancer]. AB - Since 1986, we have applied thermochemo (CDDP) therapy combined with radiotherapy to 18 patients with 25 advanced and/or recurrent head and neck cancers (thermochemoradiotherapy: TCR). In this report, the effects of TCR were compared with those of radiochemotherapy (RC) without hyperthermia for 22 patients with 27 cancers before 1985 in our department. Thermochemotherapy was performed twice a week, for a total of 8.8 times on average. Three kinds of heating system were used: a radiofrequency system, a microwave system, and a RF interstitial system. In the TCR group, 11 lesions (44.0%) showed a complete response (CR), 12 lesions (48.0%) a partial response (PR), and 2 lesions (8.0%) no change (NC). Therefore, the total response rate was 92.0%. After treatment with only RC, 5 lesions (18.5%) showed a CR, 12 lesions (44.5%) a PR, and 10 lesions (37.0%) NC, giving a total response rate of 63.0%. There was a significant difference in the CR and total response rate between these two groups. Furthermore, the 5-year cumulative local control and survival rates in the TCR group were 68.2% and 44.4%, and in the RC group were 22.2% and 18.2%, respectively. There was a significant difference in the local control rate between these two groups. In conclusion, thermochemoradiotherapy is an effective strategy for patients with advanced head and neck cancers. PMID- 11265402 TI - [Expression and pathophysiologic features of orotate phosphoribosyl transferase activity (OPRT) in gastric carcinoma]. AB - Orotate phosphoribosyl transferase (OPRT) is an enzyme that converts the pyrimidine fluoride-class anticancer drug 5-fluorouracil (5-FU) into the active nucleotide form. As such, it can be considered a primary enzyme in the first stage inhibiting DNA and RNA expression. The present study measured OPRT activity both in gastric carcinoma tissue and in surrounding normal tissue, and investigated the correlation between these findings and clinicopathologic characteristics in the patients. The study subjects were 20 patients with gastric carcinoma who were treated by surgical resection in our department. The relationship between OPRT activity in gastric carcinoma and surrounding normal tissue and patient age, sex, tissue type, extent of tumor invasion, extent of metastasis to the lymph nodes, lymphatic invasion and the existence of venous invasion of the gastric wall were investigated. The mean OPRT activity for all patients was 0.039 +/- 0.042 nmol/min/mg-prot in normal tissue and 0.120 +/- 0.099 nmol/min/mg-prot in tumor tissue, and the mean ORPT activity was significantly higher in tumor tissue than in surrounding normal tissue (p < 0.01). The OPRT ratio for tumor tissue/normal tissue (T/N) was significantly decreased as the invasiveness of the tumor increased, and was also significantly lower in patients with lymph node metastasis (p < 0.05) than in patients without lymph node metastasis. A decrease of OPRT activity in tumor tissue is a possible reason for the equivocal results of 5-FU-based chemotherapy in advanced gastric carcinoma. PMID- 11265403 TI - [Concurrent chemoradiotherapy for advanced cervical cancer--a pilot study]. AB - Recently, attempts have made to use radiotherapy in combination with chemotherapy in various solid tumors including cervical cancer. Twenty-four patients with locally advanced cervical cancer were treated with concurrent Carboplatin (16-24 mg/m2/day) or Nedaplatin (20 mg/m2/week) and conventional radiotherapy. Of 13 evaluable patients, there were nine complete responders and four partial responders. There was no renal damage or grade 4 hematological toxicity. Gastrointestinal adverse reactions were mild. One patient had grade 3 dermatologic toxicity after delayed radiation therapy. This pilot study suggests that daily Carboplatin or weekly Nedaplatin administered with standard radiation therapy is safe, well-tolerated, and thus may be useful as a radiation sensitizer in the treatment of locally advanced cervical cancer. PMID- 11265401 TI - [The histological antitumor effect and side effects of preoperative chemotherapy for patients with oral squamous cell carcinoma--comparison between low-dose and high-dose CDDP regimens]. AB - Preoperative chemotherapy should be effective against cancers and have few side effects that would prevent surgery. We investigated the histological effects and side effects of low- and high-dose CDDP chemotherapy against oral squamous cell carcinoma (SCC), and discuss the therapeutic benefits of each regimen. Thirty-six patients were divided into two groups as follows, in a non-randomized manner: A) low-dose CDDP (17 patients): CDDP 5 mg/m2/day + UFT 400 mg/day (day 1-5) (1 or 2 courses), B) high-dose CDDP (19 patients): CDDP 70-100 mg/m2/day (day 1) + peplomycin 5 mg/day (day 2-6) (1 or 2 courses). Curative surgery was conducted 1 week after protocol A or 2-3 weeks after protocol B. The histological antitumor effects were evaluated with Ohboshi & Shimosato's classification using surgical materials of primary tumors. In this classification, grade IIB, III and IV were as effective. Maximum histological effect was seen with grade IIB for regimen A and grade IV for regimen B. Four of 17 patients (23.5%) responded to regimen A and 13 of 19 patients (68.4%) to regimen B. Side effects, such as nausea, vomiting and myelosuppression, appeared with regimen B, but were seen little with regimen A. The 2-year survival rate was 93.3% with regimen A and 78.9% with regimen B. With regimen A, the 2-year survival rate of effective cases was 100% and that of ineffective cases was 91.7%. With regimen B, the rate was 92.3% and 50.0%, respectively. Effective cases showed good prognosis in both groups. The low-dose CDDP regimen was not so effective against primary tumors histologically, but the prognosis was good. The low-dose CDDP regimen appears to be useful for preoperative chemotherapy of oral SCC. PMID- 11265404 TI - [A lung cancer patient whose renal scintigram showed transient renal dysfunction after administration of contrast material for CT]. AB - We encountered a patient whose renal scintigram showed renal tubular damage four days after administration of contrast material for CT. As no secretion of radioisotope to the urinary tract was observed on the scintigram and the renogram did not show a secretionary phase, renal tubular damage was suspected. As the renal damage disappeared six days after administration of the contrast material, the damage seemed to be transient, disappearing within five days. Caution is needed with the use of renotoxic drugs such as anti-tumor drugs, especially within five days after administration of contrast materials. PMID- 11265405 TI - [A case of recurrent lung cancer successfully treated with vinorelbine and cisplatin]. AB - A 56-year-old female underwent lobectomy with ND2a lymph node dissection for left lung cancer in April 1999. Histopathological examination demonstrated moderately differentiated adenocarcinoma (pT2N2M0, stage III A). She received one course of a combination of etoposide and cisplatin as adjuvant therapy, followed by oral intake of UFT. In November 1999, a left para-aortic lymph node recurrence was found. She received radiation therapy (total 60 Gy) to the mediastinum. In April 2000, new lung and left supraclavicular lymph node recurrences were found. She received three courses of vinorelbine 20 mg/m2 (days 1, 8) and cisplatin 80 mg/m2 (day 1) followed by radiation therapy (total 50.4 Gy) to the left supraclavicular lesion. After the chemotherapy, a complete response (CR) of all metastatic lesions was achieved. Adverse reactions were grade 1 alopecia, grade 2 nausea/vomiting, grade 3 neutropenia, hypochromia, and injection site reaction. The combination of vinorelbine and cisplatin is a useful regimen in non-small cell lung cancer. PMID- 11265406 TI - [Effectiveness of intra-arterial infusion chemotherapy for solitary bone metastasis from lung cancer with radiotherapy--a case report]. AB - There have been few reports on intra-arterial infusion chemotherapy for metastatic bone cancer because the bone metastasis is multiple in almost all cases. However, selective intra-arterial infusion chemotherapy is thought to be more effective than systemic chemotherapy for solitary bone metastasis. The patient was a 47-year-old man who had been diagnosed with solitary metastasis of the right knee joint from lung cancer on the basis of various imaging studies and biopsy. The metastatic bone cancer showed rapid growth with systemic inflammatory response, and the patient's general condition became progressively worse. Therefore, radiotherapy alone consisting of 3 Gy of fraction x 5 fractions/week was initiated, but the metastatic lesion was a progressive disease (PD) at the middle point of radiotherapy (24 Gy), and we had no choice but to alter the therapy. Angiography showed dense tumor staining, so intra-arterial infusion chemotherapy was contemplated. Subsequently 15 mg/body of CDDP was administered persistently 5 days a week through a catheter placed in the right femoral artery that had been introduced via the left femoral artery. After 8 courses of this therapy (total dose 600 mg), the metastatic bone cancer was remarkably reduced in size and showed nearly a complete response (CR) on CT scan. This result suggests that intra-arterial infusion chemotherapy is very effective if there is only one bone metastasis lesion. PMID- 11265407 TI - [A case of primary malignant hemangiopericytoma of the lung with marked response to combination chemotherapy with cisplatin, ifosfamide and gemcitabine]. AB - A 51-year-old man was admitted because of complaints of cough and bloody sputa. A chest CT scan revealed a giant mass lesion in the right middle and lower lobes of the lung and mediastinal lymphadenopathy. Bronchoscopic findings showed a tumor which almost completely obstructed the intermediate bronchus. Histopathological examination of a biopsy specimen demonstrated malignant hemangiopericytoma. Two courses of chemotherapy that combined cisplatin, ifosfamide and gemcitabine were performed every 3 weeks. Both primary lesion and mediastinal lymph node metastases showed marked reduction and toxicity was manageable. PMID- 11265408 TI - [A case of breast cancer 5 cm in diameter treated with a breast preserving approach after preoperative intra-arterial chemotherapy]. AB - The patient was a 59-year-old female with a mass in the right breast (area C). At the initial examination, the mass was 5.0 x 4.5 cm on palpation, aspiration cytology was Class V, and a diagnosis of T2aN1aM0, Stage II breast cancer was made. Since the patient strongly desired a breast preserving treatment, a reduction in the size of the mass was attempted by local intra-arterial chemotherapy. Docetaxel (TXT) was administered at 60 mg into the internal thoracic artery and lateral thoracic artery at a rate of once a month for a total of 5 times. After the fifth treatment, the mass was reduced in size to 2.8 x 2.5 cm on palpation, and breast-preserving resection was then carried out. On histopathological examination, cancer was observed in an area of 3.0 x 2.2 cm. Careful follow-up is still needed, but preoperative intra-arterial chemotherapy is considered to be significant as a step before breast preserving surgery for breast cancer with a diameter of 3-5 cm. PMID- 11265409 TI - [A case of recurrent breast cancer successfully treated with docetaxel]. AB - A 50-year-old female underwent mastectomy for left breast cancer in June, 1991. She received tamoxifen for 36 months and tegafur for 30 months as adjuvant therapy. In November 1997, liver, lung and para-aortic lymph node recurrences were found, and we treated her six times with docetaxel 60 mg. After the chemotherapy, a complete response (CR) of all metastatic lesions was achieved and her serum CA15-3 level was decreased. Adverse reactions were grade 4 neutropenia, grade 2 alopecia, fever, and grade 1 edema. She received medroxyprogesterone acetate after the chemotherapy and has been well without re-growth of any metastases for over eight months. PMID- 11265411 TI - [A case of advanced gastric cancer complicated with liver metastases responding remarkably to combined chemotherapy with 5-fluorouracil and low-dose cisplatin, with UFT and low-dose cisplatin for maintenance on an outpatient basis]. AB - A 77-year-old man who had advanced gastric cancer with multiple liver metastases was treated by combined chemotherapy with 5-fluorouracil and low-dose cisplatin for 1 and half courses (1 course = 4 weeks). After this treatment, the primary gastric lesion was reduced, the liver metastases disappeared, and serum tumor marker levels decreased. After discharge, we administered a dose of 300 mg of UFT E orally every day, and 10 mg of CDDP intravenously once weekly on an outpatient basis. The patient has survived with a good quality of life. PMID- 11265410 TI - [A case of AFP-producing gastric cancer responding to low-dose CPT-11 and low dose cisplatin combination chemotherapy]. AB - Gastric cancers that produce alpha feto protein (AFP) usually have a poor prognosis. We report an AFP-producing gastric cancer that showed a partial response to low-dose CPT-11 and low-dose cisplatin combination chemotherapy. AFP producing gastric cancers successfully treated with chemotherapy have been reported, but to our knowledge this is the first report of successful treatment with low-dose CPT-11 and low-dose cisplatin combination chemotherapy. CASE: A 49 year-old woman who had gastric cardiac cancer with esophageal invasion was admitted to our institution. Since AFP-positive cells were demonstrated immunohistochemically in biopsy specimens and levels of AFP in serum were high, AFP-producing cancer was diagnosed. Because of metastasis to Virchow's node and the paraaortic lymph nodes, the tumor was considered unresectable. The patient's poor general condition necessitated chemotherapy with low toxicity and high efficacy. She was treated with low-dose CPT-11 and low-dose cisplatin combination chemotherapy. After two cycles of this treatment, the tumor volume and the serum levels of AFP had decreased markedly. The only side effect of the treatment was leukopenia. PMID- 11265412 TI - [Long-term survival of a patient with postoperative liver metastasis of stage IVa gallbladder cancer responding to hepatic arterial infusion chemotherapy]. AB - A 58-year-old man was diagnosed as having advanced gallbladder cancer (T4, P0, H0, N0, stage IVa) with direct invasion to the liver, transverse colon and duodenum. Therefore, extended cholecystectomy and bile duct resection with a partial resection of the transverse colon and the duodenum were performed in March 1992. Histopathological examination revealed moderately differentiated tubular adenocarcinoma of si, ly1, v1, hinf3, binf3, n0. Three years and eight months after the operation, multiple liver metastases were diagnosed by abdominal CT. Repeated hepatic arterial infusion chemotherapy with 5-FU 500 mg/body/w, MMC 4 mg/body/2w and EPI 40 mg/body/4w was performed starting in January 1996. Four months later, the lesions in the liver were reduced in size, and abdominal CT 10 months after the chemotherapy showed a partial response. However, the liver metastasis of the right lobe was enlarged on an abdominal CT in August 1997. Repeated hepatic arterial infusion chemotherapy with the same regimen was performed again starting in March 1998. Ten months later, the liver metastasis was slightly enlarged, but the greater part of the metastasis showed necrosis on an abdominal CT in January 1999. However, peritonitis carcinomatosa was observed later and the patient died 8 years after the operation. PMID- 11265413 TI - [Two cases of squamous cell carcinoma of the anal canal treated with chemoradiotherapy]. AB - We recently treated 2 patients with squamous cell carcinoma in the anal canal with bilateral inguinal nodal metastases using chemoradiotherapy. Chemotherapy (CT) consisted of 5-fluorouracil 700 mg/m2/day (continuous intravenously) on days 1-5 and cisplatin 50 mg/m2/day (continuous intravenously) on days 6-7. Chemotherapy was administered before the beginning of radiotherapy. In one patient, 2 cycles of CT were performed, and in the other patient 1 cycle only. The total radiation dose was 57.6 Gy to the primary lesion in each patient, and 53.6 Gy, 55.8 Gy to the nodal metastases, respectively. As a primary treatment response, CR was obtained in both patients. Acute grade 3-4 hematologic toxicities were observed in one patient. The patients have had 7 and 9 months survival without disease, and excellent function of the anal sphincter after treatment. PMID- 11265414 TI - [Prolonged use of paclitaxel for platinum-refractory ovarian cancer--brief report]. AB - The patient was a 41-year-old women with platinum-refractory ovarian cancer. She received 135-210 mg/m2 paclitaxel for 13 cycles. She received granulocyte colony stimulating factor (G-CSF) support to prevent severe granulocytopenia, and experienced little neurologic toxicity. Her quality of life was good throughout the prolonged period of use of paclitaxel. PMID- 11265415 TI - [All-trans retinoic acid combined with low-dose cytosine arabinoside treatment for acute myelogenous leukemia with trilineage myelodysplasia--a case report]. AB - AML with trilineage myelodysplasia (AML/TMDS) is recognized as having of poor prognosis due to its resistance to chemotherapy in comparison with de novo AML. An AML/TMDS patient who failed to respond to ordinary induction therapy achieved complete remission with combination therapy consisting of all-trans retinoic acid (ATRA) and low-dose Ara-C. No serious toxicity was observed. ATRA combined with low-dose Ara-C could be an alternative treatment for this kind of patient. PMID- 11265416 TI - [Early clinical trial of antineoplastic agents--recent perspectives of pharmacological analysis in phase I study]. AB - Importance pharmacological analysis has been widely recognized in conducting early clinical trials of antineoplastic agents, especially phase I trials, in recent years. Pharmacokinetic-pharmacodynamic analysis during dose escalation process is one of the extremely important research endpoints in phase I trials. In addition, measurement of protein unbound drug concentration is more informative for understanding pharmacological difference between patients and animals. Achievable plasma drug concentration which would be compared with preclinical results might be useful surrogate marker to decide optimum administration dose for further clinical investigation in conducting phase I trials of non-cytotoxic or molecular targeting drugs. PMID- 11265417 TI - [Diagnosis of appendicitis syndromes: what has changed]. PMID- 11265418 TI - [Did Ambrose Pare actually perform the first arterial ligation?]. PMID- 11265419 TI - [Myology, from ancient times to present days, or the emergence of a separate discipline]. PMID- 11265420 TI - [When should you consider a muscular disease?]. AB - Muscle diseases can take different clinical aspects and some unusual symptoms are often underestimated. Weakness of proximal limb muscles is the most frequent but distal weakness can be observed. Sometimes oculomotor, bulbar, facial, respiratory, paraspinal muscles or myocardium involvement may be the expression of a muscle disease. Similarly it is important to recognize a muscle disease in occurrence of neonatal hypotonia, muscle hypertrophy, muscle stiffness, transitory paralytic episodes, fatigability, myalgia, rhabdomyolysis or isolated increased creatine-kinases level. Biological and electrophysiological examinations have essentially a role in pointing a muscle disease. Muscle biopsy keeps the key role in the diagnosis. Muscle imaging allows mapping of the lesions and new techniques of NMR spectroscopy constitute an in vivo approach of muscle metabolism. PMID- 11265421 TI - [Mitochondrial and metabolic myopathies]. AB - Mitochondrial and metabolic myopathies constitute a group of disorders characterised by abnormal muscular metabolism of energy. Most of these disorders are genetically transmitted. Recent progress in the field has led to spectacular advances in their classification and the understanding of the mechanisms involved, particularly in mitochondrail myopathies. Diagnosis can be made et any age; the patient can present manifestations that can be misleading for the clinician. Lipid myopathies and glycogenoses usually present as a myopathic syndrome associated with cramps, spasm and myalgia, with fatigue on effort. Acute episodes of rhabdomyolysis on effort can occur, with an attendant risk of renal failure. Mitochondrial myopathies have multi-organ manifestations and muscular involvement is not always at the forefront. Although diagnosis may be suggested by clinical factors, it should be confirmed by teams and laboratories that specialize in muscular disorders. PMID- 11265422 TI - [Renewed interest in muscular dystrophy]. AB - In the last years, the molecular approach of the muscular dystrophies with mutated gene and deficient protein identification has completely renewed the knowledge of these myopathies. The combination of the following data 1. clinical phenotype, transmission mode; 2. evidence of muscle protein expression deficiency by immunocytochemical assay; and 3. gene mutation allow an unquestionable identification of the muscle disease. A new dystrophy classification has been established: thus 3 groups of autosomal recessive dystrophies have been isolated on molecular grounds, calpainopathies, sarcoglycanopathies, dysferlinopathies; each of these corresponding to a particular clinical phenotype. A suitable genetic council is now possible in most dystrophies. Finally, therapeutical prospects using "molecular repairing" may be now considered. PMID- 11265423 TI - [Inflammatory myopathies]. AB - Primary myositis (or inflammatory myopathies) comprises three main groups of diseases, based on clinical and immunohistochemical characteristics: polymyositis (PM), dermatomyositis (DM) and inclusion body myositis. Their clinical presentation and course are disparate, but a common characteristic is immune dysfunction-related inflammation of the striated muscles. Their etiologies are still not fully elucidated but associate environmental and, to a lesser degree, genetic factors. Nevertheless, considerable progress has recently been made in the understanding and management of these diseases. PMID- 11265424 TI - [Drug-induced and toxic myopathies]. AB - A large number of drugs and toxins may induce myopathic changes in several ways, they are probably more common than realized. The clinical and pathological features depend on the causative agent and on individual susceptibility to a given compound. Based on their pathologic mechanisms, there are 6 main categories of toxic myopathies: necrotizing myopathy mainly due to lipid-lowering drugs (fibrates and statines); vacuolar myopathy, usually associated with antimalarial agents; inflammatory myopathy induced by thiol derivatives; mitochondrial myopathy; steroid myopathy, and hypokaliemic myopathy. Toxic myopathies are usually reversible after discontinuation of the offending agent. Their prompt recognition may reduce their damaging effects or prevent a fatal outcome. Muscle biopsy can be very useful for the diagnosis of toxic myopathies. PMID- 11265425 TI - [Infectious myopathies]. AB - Infectious myopathies are rare acquired affections which have, generally, a good prognostic. Many types of viral infections can cause transient inflammatory myopathies. HIV myopathy may be present early in the HIV infection, but more often it is a complication of fully developed AIDS. Influenza virus myositis tend to be more severe in adults than in children. Group B coxsackie virus has been isolated from striated muscle of patients with epidemic myalgia. Parasitic infections of muscle include trichinosis, toxoplasmosis, and cysticercosis. Trichinosis is the most frequent parasitic myositis. The ocular, lingual or pharyngeal weakness and/or hypereosinophilia suggest the diagnosis. Pyomyositis, is a located zone of suppuration of muscle due to staphylococcus in 90% of the cases. It is a common occurrence in tropical climates, but has been recognized increasingly in temperate climates. PMID- 11265427 TI - [The prescriber and the pedagogue]. PMID- 11265426 TI - [Endocrine myopathies]. AB - Disturbances in the endocrine system induce a myopathy by acting on protein synthesis or on energetic metabolic pathways. Thus, a proximal myopathy is seen in hypercorticism, hyper- or hypothyroidism and acromegaly. On the other hand, endocrine disorders modify the transsarcolemmal balance in electrolytes, inducing generalized paresia, as in adrenal insufficiency or in thyrotoxic periodic paralysis. The diagnosis is usually easy if a muscle disorder occurs in a complete clinical feature of endocrinopathy but a myopathy may reveal it. Moreover, the steroid myopathy induced by iatrogenic glucocorticoid excess may lead to confusion in patients treated for an inflammatory myopathy. The treatment of endocrine myopathies is based on the correction of the hormonal disorder. PMID- 11265428 TI - [Blood and blood product transfusions. Immunological basis and indications]. PMID- 11265429 TI - [Acute anxiety attack. Diagnostic orientation and management in an emergency situation with drug dosage]. PMID- 11265430 TI - [Surveillance of patients in plaster casts]. PMID- 11265431 TI - [Soft tissue infections by anaerobic bacteria. Etiology, diagnosis, treatment]. PMID- 11265432 TI - [Hemoptysis. Diagnostic orientation]. PMID- 11265433 TI - [Red and/or painful eye. Diagnostic orientation]. PMID- 11265434 TI - [Acute osteoarticular infection of the limbs in children. Physiopathology, diagnosis, evolution, treatment]. PMID- 11265435 TI - The more things change.... PMID- 11265436 TI - Should videotaping be restricted during labor and birth? Pro. PMID- 11265437 TI - Should videotaping be restricted during labor and birth? Con. PMID- 11265438 TI - Fundal pressure during the second stage of labor. AB - The role of fundal pressure during the second stage of labor is controversial and can result in clinical disagreements between nurses and physicians. Clearly the time for resolution of this issue is not when there is a physician request at the bedside in front of the patient. A prospectively agreed upon plan specifying how this request will be addressed is ideal. In order to develop this plan, risks, benefits, and alternative approaches to the use of fundal pressure should be reviewed by an interdisciplinary perinatal team. Much of the data about maternal fetal injuries related to fundal pressure are not published for medical-legal reasons; however, anecdotal reports suggest that these risks exist. Unfortunately, it is therefore difficult to quantify with any degree of accuracy the exact number of maternal-fetal injuries that are directly related to use of fundal pressure to shorten an otherwise normal second stage of labor. However, there is enough evidence to suggest that if injury does occur when fundal pressure is used, there are significant medical-legal implications for the health care providers involved. This article will review what is currently known about fundal pressure including risks, benefits, and alternative approaches. In that context, suggestions will be offered for a safe approach to managing the second stage of labor. PMID- 11265439 TI - Taking the "ouch" out of injections for children. Using distraction to decrease pain. AB - PURPOSE: This research compared the effect of two forms of distraction on injection pain in a convenience sample of preschool children. DESIGN: A quasi experimental study of 105 children (53 girls and 52 boys) ages 4 to 6 years needing DPT immunizations. Data were collected at three sites: two school-based immunization clinics and one public health center with a walk-in immunization program. METHODS: Study children were randomly assigned to receive one of three treatments with their DTP injection: touch, bubble-blowing, or standard care. Prior to injection, a measure of medical fear was obtained (Child Medical Fear Scale) and pain was measured through use of the Oucher Scale. DATA ANALYSIS: Planned comparisons within analysis of variance (ANOVA) tested the differences in pain scores by treatment. Factorial ANOVA was used to determine the influence of age or gender on treatment, and the effect of medical fear on pain was analyzed using correlational statistics and factorial ANOVA. RESULTS: Both forms of distraction, touch and bubble-blowing, significantly reduced pain perception. There were no interaction effects of either age or gender. Fear was a significant covariate, but distraction was effective even when fear was not held constant. CLINICAL IMPLICATIONS: Distraction appears to be an effective method for decreasing injection pain in young children. It is an easy, practical nursing intervention to help children cope with this common, painful experience. PMID- 11265440 TI - Behavioral characteristics of infants with nonorganic failure to thrive during a play interaction. AB - PURPOSE: To examine the behavioral responses of infants with nonorganic failure to thrive (NOFTT) during play interactions with their mothers. DESIGN: Comparative descriptive. METHODS: The sample consisted of 31 infants; 17 with nonorganic failure to thrive (NOFTT) and 16 matched healthy controls. The infants were videotaped during a play interaction with their mothers. The behaviors exhibited by the infants were scored with the Parent-Child Early Relational Assessment. The environmental context of the play interaction was also rated for how play was initiated, maternal involvement, and the presence of chaos. RESULTS: Infants with NOFTT exhibited more difficult behaviors during play such as more negative affect, less vocalizing, and more gaze aversion. Mothers of the infants with NOFTT were less likely to remain involved during the play interaction. The environments of the infants with NOFTT were also found to be more chaotic during play. CLINICAL IMPLICATIONS: Assessment of the infant-mother interaction during play may provide insight into the interactions that occur during other caretaking activities. Strategies could be developed to assist the mother with interacting with her difficult infant. Future research could lead to interventions that could help improve the dynamics of the infant-mother interaction in infants with NOFTT. PMID- 11265441 TI - Premature rupture of the fetal membranes. An update for advanced practice nurses. AB - Advanced practice nurses in obstetric settings are frequently required to diagnose premature rupture of fetal membranes; co-management of care with physicians is becoming more common in many health care facilities. Therefore, Advanced practice nurses must have an in-depth understanding of this potentially severe obstetric complication. This article presents a review of the current literature focusing on the epidemiology, physiology, pathophysiology, prevention measures, subjective and objective assessment, diagnostic tests, and management of premature rupture of membranes. Psychosocial aspects of this event, often upsetting for the family, are also discussed. PMID- 11265442 TI - Infant feeding practices of low-income rural mothers. AB - PURPOSE: To examine infant feeding practices at 1 to 2 months of age and at 4 to 6 months in a rural population of infants at risk for failure to thrive. DESIGN: A descriptive/exploratory study with 52 mothers who were interviewed twice during the infant's first 6 months of life. Mothers were recruited from health care facilities in rural southeastern Kentucky. Mothers participated in two structured interviews about feeding practices conducted in health care clinics or in the home. RESULTS: At birth 52% of mothers chose to use formula, 41.2% chose breastfeeding, and 8% were both breastfeeding and formula feeding. By 1 month, 71% of mothers were formula feeding and only 29% were breastfeeding. At 4 to 6 months postpartum 80% of mothers were formula feeding and 20% were breastfeeding. Mothers with more children, higher family income, and more education were more likely to breastfeed. Almost all mothers began solid foods before the infant was 4 months old. Infants were fed table foods including mashed potatoes and gravy, and beverages such as apple juice, fruit juices, and soda. Mothers relied on health professionals for support for feeding decisions at the first interview; however, they relied more on the grandmother for support at the time of the second interview. CONCLUSIONS AND CLINICAL IMPLICATIONS: Breastfeeding mothers need additional support to continue breastfeeding beyond the first month. Mothers and grandmothers need education to discourage the practice of early introduction of inappropriate solid foods, including the practice of thickening bottles of formula with cereal. Nutrition teaching should be provided to mothers and grandmothers including how to select high nutrient, lower fat-weaning foods, and limiting infant intake of high-calorie drinks. PMID- 11265445 TI - [The regression model and the nonlinear autoregressive processes]. AB - In this article we review different regression models with fixed and random effects. We consider the usual methods, parametric and nonparametric, for the estimation in these models. We also see how the estimation in the autoregressive nonlinear model can be deduced of the regression, if the model satisfies some mixing conditions. PMID- 11265443 TI - Intimate partner violence. Mothers' perspectives of effects on their children. AB - PURPOSE: Intimate partner violence not only affects adults but also the children living within that "war zone." The present study expands our understanding about how children are affected when they observe violence in their own homes, as reported by their mothers. STUDY DESIGN: This descriptive study was conducted to describe mothers' perspectives of the impact of the violence on their children. A consecutive sample of 72 mothers attempting to file assault charges were interviewed in a private room by a registered nurse and were asked to describe the effect of witnessing intimate partner violence on their child's behavior. Each response was written verbatim by the interviewer. RESULTS: A majority (72%) of the mothers reported negative behaviors in their children that they believed were as a result of witnessing their mother's violent experiences. The most common negative traits were distress-indicating behaviors such as sleep disturbances, clinging, and fretful behaviors followed by problems with the abuser, problems in school, and problems with mother. CLINICAL IMPLICATIONS: Because intimate partner violence affects children, health care providers should become familiar with behaviors indicative of this problem. To promote the well being and development of children, recommendations for assessment and intervention for women experiencing intimate partner violence are discussed. PMID- 11265447 TI - [Synthesis of 3-carbethoxi-2-phenyl-4-quinolones. Potential antimitotics]. AB - The synthesis of the title compounds involved four main reactions: condensation between the acid chlorides and diethyl malonate to obtain the diethyl benzoylmalonates; transformation of these in diethyl chloroarylidenmalonates; the reaction of the substituted anilines with the chlorinated compounds and finally the thermal cyclization in diphenyl ether. PMID- 11265448 TI - Acetylation and characterization of banana (Musa paradisiaca) starch. AB - Banana native starch was acetylated and some of its functional properties were evaluated and compared to corn starch. In general, acetylated banana starch presented higher values in ash, protein and fat than corn acetylated starch. The modified starches had minor tendency to retrogradation assessed as % transmittance of starch pastes. At high temperature acetylated starches presented a water retention capacity similar to their native counterpart. The acetylation considerably increased the solubility of starches, and a similar behavior was found for swelling power. When freeze-thaw stability was studied, acetyl banana starch drained approximately 60% of water in the first and second cycles, but in the third and fourth cycles the percentage of separated water was low. However, acetyl corn starch showed lower freeze-thaw stability than the untreated sample. The modification increased the viscosity of banana starch pastes. PMID- 11265453 TI - Glyceraldehyde-3-phosphate dehydrogenase from Sulfolobus solfataricus. PMID- 11265454 TI - Nonphosphorylating glyceraldehyde-3-phosphate dehydrogenase from Thermoproteus tenax. PMID- 11265455 TI - Isocitrate dehydrogenase, malate dehydrogenase, and glutamate dehydrogenase from Archaeoglobus fulgidus. PMID- 11265456 TI - Aldehyde oxidoreductases from Pyrococcus furiosus. PMID- 11265458 TI - Acetyl-CoA synthetases I and II from Pyrococcus furiosus. PMID- 11265457 TI - 2-keto acid oxidoreductases from Pyrococcus furiosus and Thermococcus litoralis. PMID- 11265459 TI - Phosphate acetyltransferase and acetate kinase from Thermotoga maritima. PMID- 11265460 TI - Alcohol dehydrogenase from Sulfolobus solfataricus. PMID- 11265461 TI - Alcohol dehydrogenases from Thermococcus litoralis and Thermococcus strain ES-1. PMID- 11265462 TI - Alcohol dehydrogenase from Thermococcus strain AN1. PMID- 11265463 TI - Hydrogenases I and II from Pyrococcus furiosus. PMID- 11265464 TI - Fe-only hydrogenase from Thermotoga maritima. PMID- 11265465 TI - Ornithine carbamoyltransferase from Pyrococcus furiosus. PMID- 11265466 TI - Carbamoyl phosphate synthesis: carbamate kinase from Pyrococcus furiosus. PMID- 11265467 TI - Aspartate transcarbamoylase from Pyrococcus abyssi. PMID- 11265468 TI - Glutamate dehydrogenases from hyperthermophiles. PMID- 11265469 TI - Phosphoribosylanthranilate isomerase and indoleglycerol-phosphate synthase: tryptophan biosynthetic enzymes from Thermotoga maritima. PMID- 11265471 TI - Nicotinamide-mononucleotide adenylyltransferase from Methanococcus jannaschii. PMID- 11265470 TI - Nicotinamide-mononucleotide adenylyltransferase from Sulfolobus solfataricus. PMID- 11265472 TI - Alkaline phosphatase from Thermotoga neapolitana. PMID- 11265473 TI - Citrate synthase from hyperthermophilic Archaea. PMID- 11265474 TI - Dihydrofolate reductase from Thermotoga maritima. PMID- 11265475 TI - Tetrahydromethanopterin-specific enzymes from Methanopyrus kandleri. PMID- 11265476 TI - Ribulose-1,5-bisphosphate carboxylase/oxygenase from Thermococcus kodakaraensis KOD1. PMID- 11265477 TI - Respiratory enzymes from Sulfolobus acidocaldarius. PMID- 11265478 TI - ADP-dependent glucokinase and phosphofructokinase from Pyrococcus furiosus. PMID- 11265479 TI - Siroheme-sulfite reductase-type protein from Pyrobaculum islandicum. PMID- 11265480 TI - Dissimilatory ATP sulfurylase from Archaeoglobus fulgidus. PMID- 11265481 TI - Sulfite reductase and APS reductase from Archaeoglobus fulgidus. PMID- 11265482 TI - Hydrogen-sulfur oxidoreductase complex from Pyrodictium abyssi. PMID- 11265483 TI - Pyrophosphate-dependent phosphofructokinase from Thermoproteus tenax. PMID- 11265484 TI - Triose-phosphate isomerase from Pyrococcus woesei and Methanothermus fervidus. PMID- 11265485 TI - Phosphoglycerate kinase-triose-phosphate isomerase complex from Thermotoga neapolitana. PMID- 11265487 TI - An overview of canine reproduction services. Getting started. AB - A veterinarian desiring to increase his or her proficiency in canine reproduction needs to become competent in a variety of reproductive procedures. This article describes commonly performed procedures and gives an overview of how to develop a practice in canine reproduction. Once a veterinarian develops expertise in this area, the base of breeder clients in the practice should rapidly increase. PMID- 11265486 TI - Phosphoglycerate kinases from bacteria and archaea. PMID- 11265489 TI - A logical approach to infertility in the bitch. AB - This approach to infertility in the bitch describes what diagnostic methods to perform and what thought processes to consider at different phases of the estrous cycle. PMID- 11265488 TI - Ovulation timing. Concepts and controversies. AB - While the luteinizing hormone (LH) surge has long been accepted as the key event in the estrous cycle of the bitch, historically, there has been no practical way to identify it. In the past, the veterinary practitioner had to rely on general and/or subjective information received from vaginal cytology, physical examinations, and observations. With the recent development of in-clinic progesterone and LH assays, and the wider availability of laboratory quantitative progesterone assays, the LH surge can either be identified directly or estimated by the detection of changes in progesterone. As a result, ovulation time can now be predicted with high accuracy in a private practice setting. PMID- 11265490 TI - Disorders of the canine penis. AB - Function and anatomy of the canine penis are reviewed. Functional abnormalities of the penis described include lack of erection and lack of ejaculation. Physical abnormalities of the penis also are described, including paraphimosis. Diagnosis and treatment options are described. PMID- 11265491 TI - Clinical management of the subfertile stud dog. AB - Many subfertile stud dogs can sire pups with appropriate management. Determination of the area of the problem (libido, ability to breed, semen quality) is the first step. Each of these areas can often be improved or managed. A complete history, physical examination, and semen evaluation should be performed on every patient. In specific cases, additional diagnostics may be helpful, including a CBC, biochemistry profile, urinalysis, semen cultures, ultrasonography, and biopsy. Management of breeding, including ovulation timing and intrauterine insemination, can be vital in dogs with low spermatozoal numbers or motility. Treatment of underlying prostatic disease can dramatically improve semen quality and fertility. PMID- 11265493 TI - Transcervical insemination techniques in the bitch. AB - The benefits of using endoscopic TCI for frozen semen come from being able to achieve the same or better results without the need and risks of general anesthesia and surgery. The ability to do all fresh and chilled inseminations this way will certainly improve conception rates without the owner having to make a decision about exposing their bitch to the risks of anesthesia and surgery. The other potential uses open up a whole new field for canine theriogenology. Above all, the client response to the technique is overwhelmingly positive. At times the learning process will be discouraging, but the end result is worth the effort. The endoscope should not be treated as something special for frozen semen insemination but should be used at every opportunity in order to develop experience and expertise in all situations. PMID- 11265492 TI - Surgery of the canine vagina and vulva. AB - Accurate diagnosis of canine vaginal abnormalities often requires general anesthesia, vaginoscopy, and contrast radiography. Abdominal ultrasonography, thoracic radiography, computed tomography, and histopathology may also be advised for the workup of mass lesions before surgery. Many procedures such as episioplasty and resection of pedunculated vaginal masses or edematous tissue are easily performed with proper planning and equipment (e.g., electrocautery). Consideration should be given to referring more complicated procedures such as resection of large vaginal masses or vaginal stenoses to a board-certified surgeon. Finally, preoperative placement of a fentanyl patch and pre- or postoperative epidural analgesia are highly recommended for any vulvovaginal surgical procedure. PMID- 11265494 TI - Uterine and fetal monitoring in the bitch. AB - The use of uterine and fetal monitoring improves the outcome of canine obstetrics. Much of the guesswork of managing whelping can be eliminated. At normal term, absolute indications for cesarean section are detected with monitoring, before multiple fetal deaths or any serious maternal compromise occurs. Bitches with previous history of cesarean section may be able to whelp vaginally successfully, having medical intervention based on monitoring. The anxiety level of owners during whelping is diminished, and the level of participation of the veterinarian improves. PMID- 11265495 TI - Periparturient and neonatal anesthesia. AB - Small animal patients may need to be anesthetized in the periparturient period for emergency, nonobstetric reasons, elective ovariohysterectomy, or cesarean section. In each case, the physiologic changes in the dam must be accounted for in designing an anesthetic protocol, but the requirements of the fetuses will be different. Subsequent to birth, the neonatal animal may need to be anesthetized, and the unique physiology and pharmacology at this age is described. PMID- 11265496 TI - Neonatal critical care. AB - The first few minutes after a neonate's birth may determine the quality of its entire life. Immediate care includes prevention of hypothermia, clearing of nasal and oral passages, stimulation of ventilation and oxygenation, and, in a few cases, advanced life support. Any additional stress during the first weeks of life can also result in neonatal morbidity and mortality. Care of the diseased newborn must focus not only on treatment of the underlying disease but on aggressive supportive care. A safe, warm, clean, proper environment and adequate nutrition are essential. PMID- 11265497 TI - New concepts in pediatric nutrition. AB - The ultimate goal of feeding puppies and kittens is to ensure a healthy adult. The specific objectives, however, are to optimize growth, minimize risk factors for disease, and achieve optimal health and longevity. Minimum nutrient requirements are easiest to determine in growing animals using growth rates as the nutritional marker. These levels ensure a minimum level of good health in most animals. Nevertheless, the optimal nutrient levels for growth may not represent the optimal levels for other physiologic functions (e.g., immune function, disease prevention, behavior). Nutritional requirements for growing animals are being redefined using physiologic parameters other than growth rate. The most common causes of malnutrition in the neonate seem to be protein-energy deficiency or overnutrition in the perinatal period. Single micronutrient abnormalities are relatively uncommon. Nevertheless, the nutritional status during neonatal development is known to affect genetic expression and to have a lifelong impact. It is thus important to tailor the nutritional plan to the individual at each life stage and to remember that pediatric nutrition should start before conception. PMID- 11265498 TI - Congenital and inherited renal disease of small animals. AB - Congenital renal diseases are present at birth and may be determined genetically; familial renal disorders occur in related animals with a higher frequency than would be expected by chance, and frequently are inherited. The most common familial disorders in cats and dogs include renal amyloidosis, renal dysplasia, polycystic kidneys, basement membrane disorders, and tubular dysfunction (Fanconi's syndrome). This article alerts the veterinarian to commonly observed congenital and hereditary conditions of the kidneys in small animals. PMID- 11265499 TI - Diagnosis and treatment of juvenile endocrine disorders in puppies and kittens. AB - Endocrine and metabolic disorders affecting puppies and kittens from birth until 6 months of age may manifest as clinical problems related to growth, water metabolism (polydipsia or polyuria), or as episodic weakness. Endocrine and metabolic disorders that affect stature, such as pituitary or hypothyroid dwarfism, present to the veterinarian for assessment of delayed or aberrant growth. Conversely, juvenile-onset diabetes mellitus and diabetes insipidus cause excessive thirst, urination, and difficulty in house-breaking. PMID- 11265500 TI - Frustrating case presentations in canine theriogenology. AB - The practice of small animal theriogenology is rewarding, but frustrations exist concerning technologic advances as compared with other species. Reproductive clinicians striving to practice good quality medicine readily identify topics of common concern: causes that are not identified or therapeutics that are not available or applicable. Improved collaboration among theriogenologists specializing in small animal practice is evidenced by growing attendance at national and international scientific meetings, increased scientific publications, and internet communications. PMID- 11265501 TI - Canine molecular genetic testing. AB - Inherited diseases are common among dogs. Recent advances in molecular genetics provide the groundwork for the development of genetic tests for the diagnosis and prevention of inherited diseases. As a result of this progress, genetics should become an integral part of veterinary medicine. DNA tests are safe, easy to perform, and reliable if interpreted correctly. Genetic tests only need to be performed once in a dog's lifetime, because the results of DNA testing never change. Veterinarians should be prepared to understand genetic testing and counseling because they are becoming increasingly important to veterinary medicine. PMID- 11265502 TI - The epidemiology and genetics of congenital heart disease. AB - The studies summarized demonstrate that CHD is a common, major malformation. The genetic cause of each specific lesion is heterogeneous. In addition, different types of CHD can result from the same chromosomal alteration or from mutations in the same gene. Although one might predict that genotype influences clinical outcome, further studies are required. At this time, routine clinical diagnostic tests to identify the specific genetic cause are available in only a few cases, namely, those with abnormal karyotypes or those with a 22q11 deletion. In those cases with single-gene defects, genetic testing is not clinically available at this time and most likely will not become available until we can predict the significance of each mutation and until technologic advances are made that allow for large-scale, accurate screening. In the meantime, continued research on the genetic cause of CHD promises to augment our understanding of the mechanisms underlying the normal and abnormal development of the cardiac structures. These investigations also promise to augment our ability to counsel families on the recurrence risk with greater accuracy and, in the future, will allow the physician to modify his or her clinical management based on genetic cause. Finally, identifying the cause and understanding the disease mechanism allows for early intervention that may modify the degree of cardiac maldevelopment or avoid cardiac malformation altogether. PMID- 11265503 TI - Fetal diagnosis and management of congenital heart disease. AB - Fetal echocardiography has had a significant impact on the diagnosis and management of congenital heart disease. Although the fetal circulation is quite forgiving for many heart lesions, other flow abnormalities gravely affect fetal physiology. Serial studies have given us a window to observe the natural history of many lesions that progress during gestation. Although fetal cardiac surgery is not yet a viable alternative to neonatal repair, fetal intervention on noncardiac disorders has had limited success and the field is growing. Fetal diagnosis remains an integral part of the management of children with cardiac lesions. PMID- 11265504 TI - Transcatheter intervention in the neonate with congenital heart disease. AB - Over the past 30 years, interventional cardiology has developed as a distinct subspecialty, playing a major role in the management of infants with CHD. In the neonatal period, a wide variety of transcatheter interventions are performed routinely, either as palliation or therapy, as adjunct to surgery, or in place of surgical intervention. Among these are creation or enlargement of ASDs to allow atrial mixing; balloon valvotomy to treat congenital valvar stenoses; balloon angioplasty or stenting of stenotic vessels (pulmonary arteries, coarctation of aorta, or systemic or pulmonary veins) or postoperative anastomoses; closure of [figure: see text] unwanted vessels (congenital fistulae or collaterals); and other miscellaneous interventions. A wide variety of patients are candidates for these procedures, including those with transposition of the great arteries or other defects with transposition physiology, left atrial outlet obstruction and hypertension, severe valvar pulmonary or aortic stenosis, hypoplastic stenotic pulmonary arteries with severe symptomatology, severe coarctation of aorta and high surgical risks, large aortopulmonary collaterals or other hemodynamically significant unwanted vessels, acute thrombosis of certain surgical anastomoses, and many more. In experienced hands, these procedures are highly successful and safe, with a low morbidity and mortality (less than 1%). PMID- 11265506 TI - Fetal and neonatal arrhythmias. AB - Perinatal arrhythmias may occur either during fetal life or in the early neonatal period. These arrhythmias include both tachycardias and bradycardias. This article presents a brief overview of fetal and neonatal arrhythmias concentrating on their presentation, diagnosis, and treatment. PMID- 11265505 TI - Neonatal cardiac surgery. Anatomic, physiologic, and technical considerations. AB - Neonatal repair for all cardiac lesions is an attractive but as yet unattainable goal for the surgical team. We are obliged to consider both lesions that must be repaired in the absolute neonatal period, and those for which later repair is an option. This article serves as an update on some issues relating to neonatal heart surgery. The first section deals with selected general aspects of perioperative support. The second section discusses two representative lesions that illustrate many of the problems encountered in neonatal cardiac surgery: transposition of the great arteries and hypoplastic left heart syndrome. PMID- 11265507 TI - Pharmacologic considerations in the neonate with congenital heart disease. AB - Advances in knowledge about the developing cardiovascular system and compensatory physiologic changes that occur in infants with congenital heart disease have led to new approaches in the management of cardiac failure and arrhythmias. Information about the pharmacologic effects, pharmacokinetics, and pharmacodynamics of newer agents used in the management of congenital heart disease have led to more appropriate use of these medications to prolong survival and improve outcomes. PMID- 11265508 TI - Nursing considerations of the neonate with congenital heart disease. AB - Nursing care of the neonate with CHD incorporates a knowledge base of the anatomy and physiology of congenital heart defects, surgical repair, complications associated with CHD, diagnostic testing, medical therapy, and psychosocial support. The neonatal nurse is a vital member of the cardiac team in providing accurate assessments, implementing medical therapies, and supporting the family. PMID- 11265509 TI - Long-term developmental outcome of children with complex congenital heart disease. AB - As the mortality of neonatal and infant surgery for CHD continues to decrease, attention is now focused on long-term sequelae, especially later cognitive and neurologic function, in survivors. Although children with repaired or palliated CHD have an increased risk for neurocognitive deficits, most survivors are performing within the normal range for most standardized measures. Even those children at highest risk, such as patients with HLHS, are comparable with survivors of other congenital lesions, such as diaphragmatic hernia, or low-birth weight children. Continued efforts are underway to reduce cerebral injury before, during, and after congenital heart surgery. PMID- 11265510 TI - Cardiac surgery in the low-birth weight neonate. New approaches. AB - The LBW infant with congenital heart defects is likely to be at a higher risk for postoperative death and complications than their normal-sized counterparts because of the effects of prematurity and other organ system dysfunction. Current experience, however, does not suggest that waiting to obtain an arbitrary weight before surgery significantly improves outcome. Survival for low-weight infants undergoing complex repairs, such as those for interruption of the aortic arch and HLHS, is decreased compared with larger babies having the same operation at the same institution; however, the absolute survival rates are still quite acceptable. Weight alone should not be considered a contraindication to surgery for complex CHD. A careful investigation for the cause of the LBW must be undertaken (e.g., prematurity, infection, multiple congenital anomalies, and genetic abnormalities) and appropriate therapies initiated. The medical and surgical teams should collaborate to determine the optimal timing and type of surgery, with body weight being only of minor importance. PMID- 11265511 TI - Maternal issues affecting the fetus. AB - Maternal disorders and exposures that affect fetal cardiac structure and function are reviewed, emphasizing fetal echocardiographic diagnosis and monitoring, and approaches for in utero therapy. Maternal diabetes, hyperthyroidism, lupus erythematosis, epilepsy, congenital heart disease, infections, and drug exposures are considered. PMID- 11265512 TI - Radiologic interpretation of congenital heart disease. AB - Radiology in the evaluation of congenital heart disease has changed over the years but still has an important role to play. Although we can rarely make the definitive diagnosis of the intracardiac abnormalities, we can direct the clinicians' attention when cardiac disease is unsuspected and we can assist in the evaluation of complications of the disease or its treatment. PMID- 11265513 TI - Diagnosis and management of the newborn with suspected congenital heart disease. AB - In critical congenital heart lesions, the ultimate outcome depends on timely and accurate diagnosis of the structural anomaly, the evaluation and resuscitation of secondary organ damage, effective lesion-specific preoperative management, and the appropriate timing and type of surgery. Crucial to this process is continuous communication among medical, surgical, and nursing disciplines. PMID- 11265514 TI - [Post-tracheotomy pneumothorax. Is thoracic radiography required after each tracheotomy?]. AB - Pneumothorax complicating a tracheotomy is a rare event, but it should be clinically considered in order to the early treatment, mostly in urgent or pediatric cases. We report 2 cases in which pneumothorax complicated postoperatively the tracheotomy and required early clinic and radiologic diagnosis followed by a promptly and satisfactorily evacuated. We review the need to perform a chest X-Ray after every tracheotomy or only following those cases found to be at risk, as well some effective preventive measures. PMID- 11265515 TI - [Epley's reposition maneuver in the treatment of benign paroxysmal positional vertigo]. AB - We performed a prospective study over 19 patients to evaluate the efficacy of the Epley's particle repositioning maneuver for treatment of the benign paroxysmal positional vertigo (BPPV), 78.9% were cured through this procedure, so we find this method as very useful for improving the BPPV. PMID- 11265517 TI - [Sjogren syndrome in ORL. Diagnostic considerations]. AB - A presentation the history of a 51-year-old woman with xerostomia and keratoconjunctivitis sicca (KCS), developed in 10 months, investigations revealed the presence in serum of antibodies against cytoplasmic antigens SS-A (Ac anti Ro/SS-A), antinuclear antibodies (ANAs) and rheumatoid factor (RF). The Rose Bengal test was positive and in the salivary gammagraphy, made with pertecnate 99 mTc, it was observed a decrease of the captation and excretion of the designer for salivary glands. The histopathology and immunohistochemical study of minor salivary glands showed the presence of a focal lymphocitic sialadenitis (fsa) and a predominance of lymphocites CD4+. It was diagnosed as primary Sjogren's syndrome (PSS) and the patient treated with salivary substitutes, artificial tears and corticoids. We analyse the current diagnostic criteria of the group of study of the European Community for the Sjogren's syndrome (SS) and emphasize the importance of histologic and immunochemical studies, that together with the rest of complementary tests will led us to distinguish not only the different forms of the presentation of the illness but also those of all patients with pathologies which are nowadays very prevalent in our environment, such as the hepatitis C (HCV) an the human immune deficiency (HIV) virus infections. PMID- 11265516 TI - [Pharmacologic treatment of vertigo]. AB - The goal of the present work is to present a review about the main pharmacological groups that are actually used in the treatment of dizziness. PMID- 11265518 TI - [Dilated vestibular aqueduct syndrome in children. Report of a case]. AB - Enlarged vestibular aqueduct syndrome is a clinical condition characterized for a progressive perceptive deafness linked to a broadening of vestibular aqueduct greater than 1.5 mm without other otic abnormal structures. We report the case of a 2-year-old child with such congenital malformation. We have reviewed the actual literature and point out its clinical and physiopathologic features as well treatment possibilities of these cases. PMID- 11265519 TI - [Arguments in favor of hearing loss screening programs]. AB - In recent years criteria for the cochlear implant candidacy have been widened to childhood, therefore we are compelled to look for the earlier possible diagnosis of deafness. With this aim the AA. have reviewed the diagnostic process of hearing loss made during the last two decades and, consequently, be aware of the problems and planning solutions to avoid unnecesary delays in the treatment. PMID- 11265520 TI - [Jugular-tympanic paraganglioma: our experience in 2 years]. AB - The paper is a review of the jugulotympanic paragangliomata seen in our Hospital in a 2 years term. The AA. discuss every case and achieve a checking of several therapeutics procedures accepted for these growths. PMID- 11265521 TI - [Primary malignant melanoma of the oral cavity. Report of a case. Review of the literature]. AB - Malignant melanomas of the oral mucous membrane seems an infrequent pathology. They are characterized by a late diagnosis, bad prognosis and a disappointing treatment, because of the difficulties encountered for its total removal owing to complicated accessibility. We report one case of mucous membrane of the mouth in a 56 year-old man, diagnosed and operated in the General Hospital of Specialties "Ciudad de Jaen". The article end with a careful revision of the literature. PMID- 11265522 TI - [Therapy perspectives in subjective tinnitus]. AB - The AA. of this article have achieved a bibliographical perusal about treatment of subjective tinnitus, including even papers based on controlled clinical trials. Pharmacologic agents are settled on vasodilators of cochlear microcirculation (nimodipine, trimetazidine, Ginkgo biloba extract, misoprostol), lidocaine, the anxiolitics (alprazolam, corazepam) and the antidepressants (nortrityline). Comments sonorous amplification. Also are displayed, because of their benefits, the relaxation techniques (biofeeback, hypnotherapy, acupuncture and yoga) and psychological counseling. PMID- 11265523 TI - [Extramedullary plasmocytoma. Report of 5 cases]. AB - The extramedullary plasmacytoma account for 1-2% of the total number of plasma cell growths. 80 percent are originated on head and neck, beginning on submucous layer of the upper airways. Male are more frequent affected at sixth-seventh decade. Roentgentherapy chemotherapy is only worldwide accepted in cases of scattered disease, but when the cases are localizated the method is under assessment. Long tern follow up of patients is recommended, because there are cases which after many years of calm can evolve into a multiple myeloma. We report 5 cases: 3 women and 2 men, diagnosed as extramedullary plasmocytoma. In one of them, considered as nasal plasmacytoma, three years later was discovered an other plasmacytoma on a tibial epiphysis. The whole group was surgically treated with complementary rontgentherapy 4 of them. Afterwards no one showed multiple myeloma. PMID- 11265524 TI - [Is there such a thing as induced temporary disability?]. PMID- 11265525 TI - [Data about prevalence of female ejaculation]. PMID- 11265526 TI - [Arthritis and antithyroid agents]. PMID- 11265527 TI - [Todd paralysis: review of our caseload in the past 2 years]. PMID- 11265528 TI - [Drug consumption and prisons]. PMID- 11265529 TI - [Detection of depression prevalence according to the CIE-10 criteria versus the Hamilton test]. PMID- 11265530 TI - [Shopping until financial ruin. Buying euphoria or true addiction? (interview by Petra Eiden)]. PMID- 11265531 TI - [Therapy of distal esophageal carcinoma. It can also be done per endoscope]. PMID- 11265532 TI - [Differential diagnosis of acute arthritis. Quickly assessing infectious etiology]. AB - Acute arthritis of abrupt onset producing symptoms showing peak intensity within a matter of hours, have a largely uniform clinical presentation. Urogenital or gastrointestinal infections (= reactive arthritis) or infectious conditions (= septic genesis) in the patient's history may provide further information of relevance for the differential diagnosis, which includes--in particular in the case of the reactive, infectious/allergic forms--direct and serological demonstration of pathogens. In contrast, imaging procedures in the early stages are of little help in clarifying causes. PMID- 11265533 TI - [New therapy approaches in rheumatoid arthritis. Preventing early joint destruction]. AB - New perceptions about the cellular and molecular interactions involved in the inflammatory process have created a basis for specific treatment of rheumatoid arthritis. The therapeutic arsenal has now been expanded to include more selective inhibitors of cyclooxygenase, antibodies and antagonists of the cytokines, and an inhibitor of pyrimidine synthesis. Large clinical trials have demonstrated the efficacy of early immunomodulatory treatment aimed at preventing joint destruction, as well as the superiority of the combination therapy over monotherapy with a long-acting antirheumatic agent. To date, uniform criteria for the evaluation of the results of treatment are, however, lacking. PMID- 11265534 TI - [Emergencies in general practice, 3. Acute scrotum]. PMID- 11265535 TI - [Discoid lupus, recurrent aphthosis. New indications for thalidomide?]. PMID- 11265536 TI - [Dry, rough skin with extensive infection and eczema. Rapid regeneration with corticoids]. PMID- 11265537 TI - [Examination techniques in lumbar syndrome. 2: Evaluating the function of the lumbar spine in functional and static disorders]. PMID- 11265538 TI - [Therapy and prevention of migraine. New recommendations and their practical application]. PMID- 11265539 TI - [Observations on use of modern hypnotics. More rapid sleep onset, more frequent uninterrupted sleep and sleep duration]. PMID- 11265540 TI - [Diagnostic quiz. Young roofer with palpitations]. PMID- 11265541 TI - [Collective regress should disappear. And then testing that beams can bend]. PMID- 11265542 TI - [Transparency law: data protection in the forefront. Can they prevent the "transparent physician"?]. PMID- 11265543 TI - [Angiotensin II antagonism. Critical morning hours are covered]. PMID- 11265544 TI - [Bronchial asthma. A combination preparation improves patient compliance]. PMID- 11265545 TI - [When long-term marcumar administration is needed. Low-molecular-weight heparin bridges the surgery gap]. PMID- 11265546 TI - [Autism. Tormented children or parents' souls?]. PMID- 11265547 TI - [Effect of aortic valve replacent for aortic stenosis on cervical arterial blood flow]. AB - Aortic stenosis is known to modify initial upstroke time (IUT) of velocity in peripheral arteries and carotid velocities. The authors conducted a prospective study in 30 patients scheduled for aortic valve replacement for aortic stenosis. The goal was to establish postoperative correction of carotid flow disorders. In the preoperative period, a positive correlation (p < 0.01) was observed between IUT and mean pressure gradient, and a negative correlation (p < 0.02) between IUT and aortic valve area. Post-operatively, the authors observed a large decrease (p < 0.0001) of IUT, and higher (p < 0.05) systolic peaks of velocity (PSV) in all studied arteries. In this article, the authors confirmed the few previous studies which described preoperative velocity modifications in aortic stenosis population, but they also described for the first time their postoperative correction. Therefore, identifying these patterns of peripheral circulatory alterations is important and underestimation of carotid stenosis, currently estimated preoperatively, must be avoided. PMID- 11265548 TI - [Validation of quantitative parameters of paraprosthetic mitral valve regurgitation of the zone of convergence by transthoracic echocardiography]. AB - The study of the convergence zone by echocardiography is a validated method of quantification of native valve mitral regurgitation. However, there is little data concerning its applications to paraprosthetic mitral regurgitation. The aim of this study was to evaluate the method in this indication. Thirty consecutive patients (21 mechanical and 9 bioprostheses) with paraprosthetic mitral regurgitation quantified by transoesophageal echocardiography were included: 4 mild, 13 moderate and 13 severe. The regurgitant volume RV) and the regurgitant surface area (RSA) were calculated by the following formulae: RV = 2 pi.r2.Va.t.alpha/180 and RSA = RV/VTI (r: mid systolic radius of the convergence zone, Va: aliasing velocity, t: regurgitation time, alpha/180: the angular correction due to parietal stress, VTI: velocity time integral of the regurgitant flow). The feasibility of the calculation of the RV and RSA was 93 and 63% respectively. There was a statistically significant correlation between the RV and transoesophageal echocardiography (r: 0.85, p < 0.001), between RSA and transoesophageal echocardiography (r: 0.67, p < 0.05) and between RV and RSA (r: 0.95, p < 0.001). When severe paraprosthetic regurgitation was defined by a RV greater than 60 ml and RSA greater than 40 mm2, the concordance between RV, RSA and transoesophageal echocardiography was 75% and 74% respectively. Therefore, the study of the convergence zone provides an accurate evaluation of paraprosthetic mitral regurgitation by transthoracic echocardiography. PMID- 11265549 TI - [Evaluation of the ejection fraction tomoscintigraphy synchronised with an electrocardiogram]. AB - Electrocardiographic gated myocardial scintigraphy (gated SPECT) allows simultaneous evaluation of left ventricular perfusion and contractility. The aim of this study was to assess the impact of the type of tracer and temporal sampling on the measurement of the ejection fraction by gated SPECT. Eighty-six consecutive patients (59 men, 27 women, average age 62 +/- 11 years) with a history of myocardial infarction (anterior N = 50--inferior N = 36) were studied. All had measurement of the isotopic ejection fraction by equilibrium angioscintigraphy and by gated SPECT sampled at 8 (N = 45, 52%) or 16 images/cycle RR (N = 41, 48%). After filtered retroprojection, the ejection fraction was calculated by a validated method (QGS). A better correlation between gated SPECT and equilibrium angioscintigraphy was observed at 16 images/cycle (r = 0.90) compared with 8 (r = 0.80). The Bland-Altman curves showed improved accuracy of the measurement of the ejection fraction by gated SPECT with 16 compared with 8 images/cycles sampling (mean differences of 0.37 +/- 6.06% and 4.3 +/- 7.71%, respectively). Multivariable analysis showed that temporary sampling was the only parameter to significantly affect the difference between the two methods. There was no difference between the use of Thallium and that of 99mTc-sestamibi. Gated SPECT, therefore, provides an accurate measurement of the ejection fraction for routine clinical usage, providing the acquisition is made at 16 images/cycle. PMID- 11265550 TI - [Brain natriuretic peptide and heart failure]. AB - Brain natiuretic peptide (BNP) is a hormone secreted specifically by the left ventricular myocytes. Its concentration is correlated with the severity of symptomatic or asymptomatic left ventricular dysfunction. The measurement of BNP has several applications from the screening of populations to the monitoring of the effects of treatment and the evaluation of the prognosis of cardiac failure. The emergence of new methods of rapid measurement will enable its usage as a routine investigation in the near future. Large scale clinical trials are, however, required to confirm the hopes raised by this new marker of left ventricular dysfunction. PMID- 11265551 TI - [Hormone replacement therapy and cardiovascular risk in menopausal women]. AB - The incidence of cardiovascular disease (CVD) differs markedly with regard to gender and age. Women have CVD 10 years later than men. Estrogen seems to play a key role in mediating the risk of CVD in women. The relative risk of CVD is lower in women before menopause and equals that of men between the 6th and 7th decade. However the effects of hormone replacement therapy (HRT) on cardiovascular morbidity and/or mortality in postmenopausal women remain controversial. The results of observational studies and randomized trials in post menopausal women are reviewed in this article. They point out the absolute need of large scale prospective randomized clinical studies to quantify the effects of HRT for primary and secondary prevention of cardiovascular disease in particular in non American countries. PMID- 11265552 TI - [Stenosis of the superior caval canal after Mustard and Senning procedures: treatment by dilatation and stent implantation. Two case reports]. AB - The authors report two cases of stenosis of the superior caval canal after Mustard and Senning procedures for transposition of the great arteries in patients paced for atrial arrhythmia. During cardiac catheterisation, it was possible to treat the stenosis by perforation in one case and by balloon dilatation in the second, followed by the implantation of two stents to reestablish vascular patency. In one patient, two pacing catheters were implanted by an endovascular approach without complications three months after stenting; in the other case, epicardial pacing was required because the patient's condition could not wait for endothelialisation of the stent before implanting the pacing catheters. This technique of revascularisation may also be used in children with venous stenosis after implantation of endocavitary pacing catheters, in which the pacing system has to be changed. PMID- 11265554 TI - [Obstructive left triatrial heart in the adult. Echocardiographic appearances]. AB - The authors report the case of a 23-year old man who presented with signs of pulmonary hypertension due to an obstructive left triatrial heart in adulthood. Transthoracic and transoesophageal echocardiography showed a partially calcified intra-left atrial membrane perforated in its centre. Doppler analysis of flow through the membrane showed continuous systolo-diastolic flow at high velocity indicating haemodynamic obstruction. The systolic pulmonary artery pressure was estimated at 80 mmHg. The patient was treated by surgical excision of the intra left atrial membrane. PMID- 11265553 TI - [Use of Organon, a synthetic heparinoid, in two cardiopulmonary bypass procedures in the same patient sensitive to heparin]. AB - We report the case of a patient who underwent two cardiopulmonary bypass (CPB) procedures with Orgaran because of heparin-induced thrombocytopenia. A 38 years old man with ischemic mitral insufficiency was operated for coronary artery bypass and valvular replacement. The CPB was carried out with heparin. Heparin induced thrombocytopenia occured and was proven immunologically. Two months later, a new valvular replacement was performed because of paravalvular leak due to endocarditis. The Orgaran-CPB protocol was as follows: 5,000 units before cardiopulmonary bypass, 5,000 units in the priming volume, anti-Xa level between 0.9 and 1.1 units/mL, with injection of 1,500 units if necessary, no administration of protamine. One month later, a new valvular replacement was necessary and performed with the same protocol using Orgaran. No bleeding or thrombotic complication occurred. Orgaran is a safe and reliable anti-thrombotic substitute if anti-Xa activity is closely monitored. PMID- 11265555 TI - [Mitral stenosis secondary to Hurler's syndrome]. AB - Valvular disease in mucopolysaccharidosis type I-Hurler (MPS/1H) is relatively common, but mitral stenosis is very rare in this genetic abnormality. The authors describe the case of a 16-year old girl with Hurler's syndrome diagnosed at 4 years of age. The morphological features were characteristic: bridged nose, thickened lips, macroglassia, short neck (gargoylism, short, thick fingers and limitation of brachial and fore-arm flexion. She presented with stage II dyspnoea and paroxysmal nocturnal dyspnoea. Radiological and echocardiographic studies revealed severe mitral stenosis with haemodynamic complications requiring mitral valve replacement. Anatomopathological analysis of the mitral valve confirmed mucopolysaccharide deposits as the cause of this particular case of mitral stenosis. PMID- 11265556 TI - [Subacute infectious endocarditis due to the agent of cat scratch fever: Bartonella henselae]. AB - The diagnosis of severe mitral stenosis with left atrial thrombus was rectified at valvular replacement in a 48-year old immuno-competent man who was a cat owner. The mass in the left atrium was, in fact, a large endocarditic vegetation. Pre- and postoperative blood cultures were negative as was culture of the excised mitral valve. The diagnosis of infectious endocarditis (IE) due to Bartonella Henselae was made from a positive serological test (1600) and identification of the germ by genetic amplification. Antibiotic therapy was continued for 6 months and the patient was cured with a follow-up of 4 years. Bartonella Henselae IE is very rare (14 reported cases) and affects mainly the aortic valve, often giving rise to very large vegetations which, in half the cases, are complicated by systemic emboli. Germs like Batonella are sensitive to most antibiotics, especially the aminosides and macrolides. In Bartonella Henselae IE, valve replacement is the rule (13 out of 14 cases) and the prognosis is usually good. Sero-diagnosis of Bartonellosis should be part of the systematic investigation of all blood culture negative IE. PMID- 11265557 TI - [Atrial fibrillation: a time when everything goes]. PMID- 11265558 TI - [Economic study of carvedilol in heart failure. A cost effectiveness study in France]. AB - A programme of four phase III clinical trials carried out in the USA on 1094 patients showed that Carvedilol, associated with the usual bitherapy and eventually with digitalis, reduced the mortality and number of hospital admissions of patients with cardiac failure. These results, transposed to the French population, may be used to evaluate the economic advantages of Carvedilol by developing a cost-effectiveness study which consists in relating the direct expenses (drugs and hospital admissions) of each of the two strategies, with or without Carvedilol, to their respective mortalities. Hospital expenses were estimated with respect to the H.M.G. corresponding to each hospital stay at 1997 1998 values. The cost in the Carvedilol group was 2,823 FF per patient (including 1,491 FF for the drug itself) but 2,056 FF were economised in hospital expenses. With an increased cost of 767 FF but a 50% reduction in mortality corresponding to a difference in mortality of 45@1000, the cost-effectiveness of Carvedilol was 17,040 per life saved and 2,130 FF per additional year of life expectancy. A study of the sensitivity produced even more favourable results of Carvedilol. An evaluation of hospital expenses on the basis of AP-HP data indicates that the addition of Carvedilol is associated with a 4,425 FF reduction in hospital expenses, which makes it a cost saving strategy. PMID- 11265559 TI - [Percutaneous revascularization of multivessel coronary disease with systematic stent implantation. Immediate results and clinical follow-up and middle term angiography]. AB - The aim of this retrospective study was to assess the immediate and medium-term clinical and angiographic results of multiple angioplasty with stenting in 100 consecutive patients with multivessel coronary artery disease. The mean age of the population was 62 +/- 11 years. Two hundred and eight lesions were treated (2.5 +/- 0.7 per patient) with implantation of 1.14 +/- 0.4 stents per lesion. The angiographic success rate was 98.7%. There were 5 major complications in the hospital period: 3 deaths, including 2 of cardiac causes, one coronary bypass procedure and one Q wave myocardial infarction. During follow-up (17 +/- 6 months), eight patients died (5 of cardiac causes) and secondary revascularisation procedures were required in 22 patients. At 6 months, the angiographic restenosis rate was 32% per lesion, 28.8% per stent and 33 patients had at least one restenosis. PMID- 11265560 TI - [Observation studies are essential in many medical questions. But there is a risk of systemic errors]. PMID- 11265561 TI - [Fixation on randomization inhibits clinical therapeutic research]. PMID- 11265562 TI - [Comment 1. Caution for over-confidence in science]. PMID- 11265563 TI - [Comment 2. Randomized trials are still the golden standard]. PMID- 11265564 TI - [Albumin use as plasma expander within intensive care is doubtful]. PMID- 11265565 TI - [Molecular genetics of the long QT syndrome. Genes causing syncope and sudden death]. AB - Molecular genetic studies in congenital long QT syndrome have characterized genes and mechanisms of arrhythmias. At least six genes encoding cardiac potassium and sodium ionic channels have been described with several mutations in each gene. The altered function produces abnormal cardiac repolarization seen on ECG as prolongation of the QT-interval and T-wave abnormalities. This may increase the propensity for ventricular arrhythmias such as Torsade de Pointe, the cause of unexpected syncope and sudden death in young patients. Clinical manifestations vary depending on the genotype present. Gene-specific therapies have recently been tried. Initial therapy of choice for symptomatic and also asymptomatic children is administration of beta-blockers. PMID- 11265566 TI - [Young woman dies of water intoxication after taking one tablet of ecstasy. Today's drug panorama calls for increased vigilance in health care]. AB - Increasing numbers of reports demonstrate that low doses of ecstasy (3,4 methylenedioxymethamphetamine--MDMA) may induce life-threatening conditions, such as hyperthermia and water intoxication. These lethal states are rarely due to overdose, and young women seem to be at particular risk. This is a case report of a 20-year-old previously healthy Swedish girl. She died of water intoxication and cerebral edema approximately 24 hours after ingestion of one tablet of "ecstasy" at a rave club in Amsterdam. Clinical findings and laboratory data suggested a syndrome of inappropriate antidiuretic hormone secretion (SIADH) induced by MDMA in combination with excessive intake of water. This case illustrates the dire consequences of the present drug abuse panorama, if the increasing need for awareness of potentially lethal complications is not met within emergency health care services. PMID- 11265567 TI - [Depression and anxiety in the elderly still underdiagnosed. SSRI preparations in conjunction with psychotherapy provide effective treatment]. AB - Depression and anxiety disorders in the elderly are common and under-diagnosed. As depressed elderly people often present with more somatic than psychiatric symptoms, diagnosis is difficult for the general practitioner. The Geriatric Depression Scale can be used as a screening instrument for diagnosis in the elderly. The etiology of depression and anxiety disorders is multifactorial. Important risk factors are psychological stress, reduced absorption of essential nutrients such as folacin and vitamin B12, and biological changes in the brain associated with aging. Selective serotonin reuptake inhibitors (SSRIs) are the drugs of choice in the treatment of elderly people with depression and anxiety disorders. Currently, the most widely used SSRI is citalopram, which according to controlled trials has an effect not only on depressed mood but also on anxiety. The use of SSRIs combined with support and psychotherapy elicits a positive response in nearly 90% of elderly patients. In Sweden, the use of antidepressants is currently most common in the age group 75-80 years, expressed in DDD (defined daily doses/1,000 inhabitants). This indicates a fairly active treatment of the elderly in Sweden. PMID- 11265568 TI - [Blockade of TNFalpha--new therapeutic principle in severe rheumatoid arthritis]. AB - TNF-alpha is a proinflammatory cytokine. It has a key function in the inflammatory cascade both systemically and locally in the inflamed joints of patients affected by rheumatoid arthritis (RA). Treatment with two different "biological" drugs that block the proinflammatory capacity of TNF-alpha has recently been approved by the European drug authorities. This paper discusses experience gained in clinical trials and during the first year of treatment in Sweden using infliximab (anti-TNF-alpha monoclonal antibodies) and etanercept (recombinant TNF-alpha receptor fusion protein). PMID- 11265569 TI - [Clostridium difficile-associated diarrhea--a growing problem in geriatric care]. AB - From 1994 to 1998 the incidence of Clostridium difficile-associated diarrhoea (CDAD) in the Department of Geriatric Medicine, Huddinge University Hospital increased from 0.5% to 2.2% of all admissions. Corresponding figures for the whole hospital were 0.3% and 0.6%, respectively. The increase in CDAD at the Department of Geriatric Medicine was parallel with a more than doubled consumption of antibiotics. All geriatric patients with CDAD had been treated with antibiotics before onset of diarrhoea. Out of the antibiotic prescriptions 48% were a cephalosporin (mainly cefuroxim). In a matched reference group of geriatric patients 51% had been treated with antibiotics during the hospital stay. The patients with CDAD spent 27 +/- 14 days in hospital as compared to 13 +/- 9 days (P < 0.05) in the reference population. PMID- 11265570 TI - [Centralization or more money to health care services? A third way--use the technology!]. PMID- 11265571 TI - [Who is to take photos of accidents?]. PMID- 11265573 TI - [Computer tomography in brain concussion: neither risks nor benefits should be ignored]. PMID- 11265572 TI - [Competence examination for specialists in general practice is under development]. PMID- 11265574 TI - [Is there a need of cost limit in connection with X-ray examination in private practice?]. PMID- 11265575 TI - [Working dog club in medical education--or our need of logic]. PMID- 11265576 TI - [About the evil ideology of Nazism]. PMID- 11265577 TI - [Rohypnol--a narcotic drug]. PMID- 11265578 TI - [An unpleasant thought about circumcision]. PMID- 11265579 TI - Promoting health: intervention strategies from social and behavioral research. AB - This report, released by the Division of Health Promotion and Disease Prevention within the Institute of Medicine at the National Academy of Sciences, asserts that behavioral and social interventions such as health promotion and disease prevention offer great promise to reduce disease morbidity and mortality in the United States, but as yet their potential has not been recognized or tapped by the federal government. Two overarching recommendations are the need to address generic social and behavioral determinants of health rather than the clinical causes of disease and death, and the need to intervene at multiple levels of influence including the individual, interpersonal, institutional, community, and policy levels. Seven recommendations for intervention strategies, nine recommendations for research, and three recommendations for funding are offered. PMID- 11265580 TI - The effect of disease prevention and health promotion on workplace productivity: a literature review. AB - This report was prepared by the Center for Disease Prevention and Health Promotion in the Institute for Health and Productivity Management as part of an effort to improve understanding of the connection between employee health and performance and to begin to identify new strategies through which treating health as an investment in human capital can lead to greater business success. Computer database searches of peer-reviewed literature published between 1993 and 1998 and manual reviews of 20 journals were used to identify research on the link between employee health and performance. Data was extracted to summarize the overall findings on the magnitude of health problems addressed by health promotion and disease prevention programs, and the impact of interventions on improving health risk, reducing medical care cost, and improving worker performance. From this summary, major conclusions on early detection of disease, the impact of behavior change programs, and appropriate care-seeking were drawn. This systematic review is supplemented with summaries of 15 seminal articles and descriptions of five leading-practices programs. The influence of developments in work/family issues, complementary and alternative medicine, and quality of care and health outcomes research are briefly discussed. Finally, a conceptual framework for studying the impact of health and productivity is described. PMID- 11265582 TI - Role of uncondensed 1,2,4-triazine derivatives as biocidal plant protection agents--a review. AB - The role of uncondensed 1,2,4-triazine derivatives and the related compounds as biocidal plant protection agents such as herbicides, bactericidal, fungicidal, antimicrobial, protozacides, anticoccidal, parasiticides, insecticides, acaricdes and pesticides, is reviewed. PMID- 11265583 TI - Antimycobacterial activity of new quinoxaline-2-carbonitrile and quinoxaline-2 carbonitrile 1,4-di-N-oxide derivatives. AB - The compounds being reported in this paper have all been evaluated within the TAACF Antituberculosis Screen Program, and some of them have been shown to possess high growth inhibition activity against Mycobacterium tuberculosis and Mycobacterium avium in the run of the first and second level in vitro screenings. The three compounds which have shown a good SI (Selectivity Index) are 2b, 4b and 4d; in addition, 6,7-dimethyl-3-[4-(4'-nitrophenyl)piperazinl-yl]quinoxaline-2 carbonitrilo 1,4-di-N-oxide (4b) is currently being tested within the in vivo antituberculosis screening in view of its very good in vitro activity. PMID- 11265581 TI - Forces shaping the future of health promotion. PMID- 11265584 TI - [Strategies for the synthesis of nifedipine analog esters of l-ascorbic acid]. AB - The synthesis of L-ascorbyl 4-aryl-2,6-dimethyl-1,4-dihydropyridine-3,5 dicarboxylates was not successful using various modifications. The strategies are shown and 7 new L-ascorbic acid derivatives are synthesized, including the dispiro compound 14. PMID- 11265585 TI - HPLC enantiomeric resolution of nebivolol on normal and reversed amylose based chiral phases. AB - Racemic nebivolol, a beta-adrenergic blocker showing very promising beta adrenergic antagonist properties in comparison to other beta-adrenergic blockers has been resolved by HPLC under normal and reversed phase modes. The columns used were Chiralpak AD and Chiralpak AD-RH containing amylose tris (3,5-dimethyl phenyl carbamate) as the chiral selector. The mobile phases used were pure ethanol and 1-propanol. The flow rates used were 0.5, 1.0 and 1.5 ml/min. The best resolution was achieved at 0.5 ml/min. flow rate with ethanol and 1-propanol on both Chiralpak AD and Chiralpak AD-RH stationary phases. The values of infinity for both alcohols on Chiralpak AD were 1.38 while on Chiralpak AD-RH these values were 1.41 and 1.38 respectively. The values of Rs for ethanol and 1 propanol were 2.63 and 1.71 on Chiralpak AD and 1.73 and 1.76 on Chiralpak AD-RH respectively. PMID- 11265586 TI - Pelleted bioadhesive polymeric nanoparticles for buccal delivery of insulin: preparation and characterization. AB - The study was an attempt to develop an alternative buccal delivery system for insulin. Insulin bearing nanoparticles were prepared by the emulsion internal phase evaporation method. The effect of some formulation variables viz., polymer/drug ratio and emulsifier concentration was studied on particle size and entrapment efficiency. Nanoparticles were pelleted to impart three-dimensional structural conformity and coherence thereby facilitating buccal application. Solid lateral and horizontal sedimentaton in the pellet can be avoided by nanoparticulation and ensuring uniform drug distribution throughout the pellet. The in vitro studies of the pellets included bioadhesion and drug release profile. In vivo studies were performed on diabetic rats. A significant hypoglycemic response was observed after 7 h, without any detectable fluctuation in blood glucose profile and risk of hypoglycemia. PMID- 11265587 TI - Diazepam submicron emulsions containing soya-bean oil and intended for oral or rectal delivery. AB - Physically stable diazepam submicron emulsions were prepared using soya-bean oil. Diazepam concentration 4 mg/ml, suitable for rectal or oral delivery, was achieved in 20% emulsions. Mixture of egg lecithin (1.2%) and poloxamer (2.0%) has been chosen as the most suitable emulsifying agent. Composition of the emulsion may be supplemented with alpha-tocopherol and parabens. However, the system was not stable when either phenylethanol or chlorhexidine gluconate was added. Taste masking agents commonly used as food additives decreased stability of the preparation and were not efficient in elimination of a bitter taste of the drug-loaded emulsions. PMID- 11265588 TI - Chitosan based pentazocine microspheres for intranasal systemic delivery: development and biopharmaceutical evaluation. AB - Bioadhesive chitosan microspheres (Ms) of pentazocine (Pz) for intranasal systemic delivery were prepared with the aim of avoiding the first pass effect, and thus improving the bioavailability and achieving sustained and controlled blood level profiles, as an alternative therapy to injection and to obtain improved therapeutic efficacy in the treatment of chronic pain such as cancer, trauma and post-operative pain, etc. The formulation variables were drug loading, polymer concentration, stirring rate during crosslinking and oils. The microspheres (Ms) were subjected to evaluation for physical characteristics, such as particle size, incorporation efficiency, swelling ability, in vitro bioadhesion, in vitro drug release characteristics and in vivo performance in rabbits. Application of in vitro data to various kinetic equations indicated matrix diffusion controlled drug delivery from chitosan Ms. Drug loading, polymer concentration and stirring speed influenced the drug release profiles significantly while oils had negligible effect. In vivo studies indicated significantly improved bioavailability of Pz from Ms with sustained and controlled blood level profiles as compared to i.v., oral and nasal administration of drug solution. Good correlation was observed between in vitro and in vivo data. PMID- 11265589 TI - Studies on aging piroxicam-polyvinylpyrrolidone solid dispersions. AB - The stabilities of X-ray amorphous solid dispersions of piroxicam and polyvinylpyrrolidone (PVP) K-17 and PVP K-30 (1:5 and 1:4), respectively, were investigated after storage for 12 months. X-ray diffraction showed that in the aged solid dispersions piroxicam remained in the amorphous state. Fourier transform infrared (FTIR) spectroscopy indicated that the interactions between drug and PVP in aged solid dispersions are similar to those in freshly prepared samples. The dissolution rates of the X-ray amorphous solid dispersions during storage for 12 months at 45 degrees C and ambient temperature were examined. Very minor decreases in dissolution rates of aged solid dispersions were found which might be due to the coarsening of the particles. Dissolutions of these amorphous solid dispersions after aging for 12 months still showed an about 40-fold increase in dissolution in 5 min compared to pure drug. PMID- 11265590 TI - Age-related changes in skin permeability of hydrophilic and lipophilic compounds in rats. AB - The effect of age on intact and stripped skin permeability of lipophilic (ketoprofen and isosorbide dinitrate) and hydrophilic permeants (deuterium oxide and diclofenac sodium) was investigated using STD: Wistar male rats aged 5 to 180 days. The permeability of permeants through intact skin increased with increasing lipophilicity of the permeants at each age, indicating that the permselective property of rat skin is revealed even at 5-days-old. The permeability coefficients through intact skin decreased with increasing age, and the extent of these decreases was higher for lipophilic permeants than that for hydrophilic permeants. On the other hand, the stripped skin permeability of permeants was almost the same at each age, and with aging each permeability coefficient through stripped skin decreased up to 21 days, dramatically during 21-90 days and then gradually again to 180 days. The thickness of the stratum corneum and stripped skin increased according to age with faster growth during 21-90 days. The reciprocal of the mean thickness of stratum corneum and stripped skin correlated well with intact skin and stripped skin permeability (r > 0.9), respectively. These results clarified that the permselectivity of rat skin against lipophilicity of permeant exists at the latest from 5 days after birth. In addition, it is speculated that the thickness of skin is a large factor in the decrease of its permeability with age. PMID- 11265591 TI - In vivo modulation of the splenocyte yield and composition by female sex steroid hormones. AB - The study was designed to shed more light on the controversial role of the two main ovarian steroid hormones (i.e. estradiol and progesterone) in shaping the size and phenotypic characteristics of the splenic lymphocyte pool. For this purpose ovariectomized adult rats (OVX) were treated for 14 subsequent days with either estradiol or progesterone (to attain physiological concentrations of the hormones). Afterwards, the splenocyte yield, and overall number of splenocytes bearing TCR alpha beta receptor, CD4 and CD8 coreceptor were evaluated. Fourteen day-long ovarian hormone deprivation produced an increase in the splenic weight and splenocyte yield (on the account of a rise in the number of TCR alpha beta- cells), although the number of TCR alpha beta+ cells was reduced as a result of a decrease in the size of the CD4+ cell subpopulation. Replacement of either estradiol or progesterone prevented the increase in splenic weight and reduced the splenocyte yield to values significantly lower than that in sham-OVX rats. Both the treatments completely abolished the effect of ovariectomy on the size of TCR alpha beta- cell population, but had differential effects on that of TCR alpha beta+ cell population; estradiol did not affect its size, while progesterone caused a reduction on the account of a decrease in the numbers of both CD4+ and CD8+ cells. The results suggest that: a) estradiol and progesterone have similar effects on the size of the splenic B cell population and that replacement of either estradiol or progesterone can prevent the effects of ovariectomy on the size of this population and b) estradiol does not affect while progesterone reduces the size of splenic T cell population. Thus, replacement of none of them is able to compensate the removal of gonads. PMID- 11265592 TI - Role of lycopene in recovery of radiation induced injury to mammalian cellular organelles. AB - Whole body exposure of male rats to 7 Gy gamma irradiation increased lipid peroxidation in the liver resulting in biomembrane damage of subcellular structures and release of their enzymes. This is evidenced by increase of thiobarbituric acid-reactive substances (TBARS) in mitochondria, lysosomes and microsomes. This was associated with a decrease in activity of the enzymes specific for each subcellular fraction; namely, mitochondrial glutamate dehydrogenase (GDH), lysosomal beta-glucuronidase and microsomal glucose 6 phosphatase. This was paralleled by an increased activity of these enzymes in the cytosol. Rats were supplemented with lycopene, a carotenoid present in tomatoes (5 mg/kg weight/day), by gavage, for 7 days before exposure to 7 Gy gamma irradiation. This resulted in diminishing amount of TBARS recorded for each subcellular structure in the liver of irradiated animals. Significant amelioration in the decrease recorded for the activity of mitochondrial glutamate dehydrogenase, lysosomal beta-glucuronidase and microsomal glucose 6-phosphatase was observed. This was associated with significant amelioration in the increase recorded for the activity of these enzymes in the cytosol. It is postulated that lycopene could play an important role in the recovery of the integrity of biological membranes of the liver after radiation injury. PMID- 11265593 TI - Chemoprotective activity of boldine: modulation of drug-metabolizing enzymes. AB - Possible chemoprotective effects of the naturally occurring alkaloid boldine, a major alkaloid of boldo (Peumus boldus Mol.) leaves and bark, including in vitro modulations of drug-metabolizing enzymes in mouse hepatoma Hepa-1 cell line and mouse hepatic microsomes, were investigated. Boldine manifested inhibition activity on hepatic microsomal CYP1A-dependent 7-ethoxyresorufin O-deethylase and CYP3A-dependent testosterone 6 beta-hydroxylase activities and stimulated glutathione S-transferase activity in Hepa-1 cells. In addition to the known antioxidant activity, boldine could decrease the metabolic activation of other xenobiotics including chemical mutagens. PMID- 11265594 TI - New pentacyclic triterpenes from the roots of Hemidesmus indicus. AB - Phytochemical studies on the roots of Hemidesmus indicus resulted in the isolation of six new pentacyclic triterpenes including two oleanenes identified as olean-12-en-21 beta-yl acetate, and olean-12-en-3 alpha-yl acetate, three ursenes characterized as 16(17)-seco-urs-12,20(30)-dien-18 alpha H-3 beta-yl actetate, urs-20(30)-en-18 beta H-3 beta-yl acetate and 16(17)-seco-urs-12,20(30) dien-18-alpha H-3 beta-ol and a lupene formulated us lup-1,12-dien-3-on-21-ol including a known compound, beta-amyrin acetate, on the basis of spectroscopic techniques and chemical means. PMID- 11265595 TI - Further saponins from Fagonia cretica. AB - Three triterpenoid saponins including two new ones were isolated and identified from the aerial parts of Fagonia cretica. The new saponins were characterized as 3-O-[beta-D-xylopyranosyl(1-->2)alpha-L-arabinopyranosyl]27-hydroxyoleanolic acid 28-O-[beta-D-glucopyranosyl(1-->6)beta-D-glucopyranosyl]ester and 3 beta-O-[beta D-xylopyranosyl(1-->2)alpha-L-arabinopyranosyl] olean-12-en-27-al-28-oic acid 28 O-[beta-D-glucopyranosyl(1-->6)beta-D-glycopyranosyl] ester. The structures were determined by spectral analyses. The NMR assignments were made by means of HOHAHA, 1H-1H COSY, HMQC, HMBC spectra and NOE studies. PMID- 11265596 TI - [Maximilian Ehrenstein (1899-1968)--life and work]. AB - The pharmacist, chemist and food analyst Maximilian Ehrenstein prepared his doctoral thesis at the Georg-August-University Gottingen under the supervision of the later Nobel Prize winner Adolf Windaus. He has been a postdoctoral fellow with the two later Nobel Prize winners Paul Karrer at Zurich and Heinrich Wieland at Munich and worked in Berlin with Carl Mannich to obtain his habilitation for pharmaceutical chemistry. After his emigration to the United States he prepared the ground for the development of oral active progestational hormones, which finally led to the anti-baby pill by Carl Djerassi. He received the honorary doctorates from the Free University of Berlin (Dr. rer. nat. h. c.) and the University of Hamburg (Dr. med. h. c.). PMID- 11265597 TI - [N1-hetarylcarbonyl substituted amidrazones and 3,5-disubstituted 1,2,4-triazoles as potential antimyocobacterial agents]. PMID- 11265598 TI - Correlation of drug absorption with molecular charge distribution. PMID- 11265599 TI - A new cyclolanostanol arabinoside from the rhizome of Cimicifuga racemosa. PMID- 11265600 TI - Two new terpenoid glucosides from Clerodendrum serratum. PMID- 11265601 TI - Forum on managed care controls? PMID- 11265602 TI - "Diagnostic value of cervical discography". PMID- 11265603 TI - Practical experience with the Stoppa repair of ventral/incisional hernias. AB - BACKGROUND: Although there are several surgical techniques to repair ventral/incisional hernias, these recur as often as 50% of the time. We have reviewed our experience with a wide, preperitoneal inlay of a conforming mesh (the Stoppa repair) in the management of more difficult ventral/incisional hernias. METHODS: Retrospective chart review of all patients receiving ventral/incisional hernia repair at our institution between 29 December 1993 and 20 May 1998. Charts were analyzed for prior ventral/incisional hernia repairs, and risk factors for recurrence (morbid obesity, prior mesh infections, and asthma/chronic obstructive pulmonary disease). Follow-up was obtained through outpatient chart reviews and phone calls. RESULTS: There were 76 repairs in 67 patients. The mean length of hospital stay was 5.08 days (4.38 days primary, 5.3 days mesh, 5.5 days Stoppa). The mean length of operation time was 164 minutes (131 minutes primary, 141 minutes mesh, 231 minutes Stoppa). Forty-one percent of the patients who underwent a Stoppa repair had one or more risk factors for recurrence and 50% (mean of 1.9 prior repairs per patient) of the patients had prior repairs, compared to 26% and 36% (mean of 1.4 prior repairs per patient) for mesh patch repairs, and 33% and 21% (mean of 1 prior repair per patient) for primary repairs. The crude recurrence rates were 43.5% for primary repairs, 19.2% for mesh repairs, and 18.2% for Stoppa repairs. CONCLUSIONS: The Stoppa repair appears to be an effective technique for particularly complex ventral/incisional hernia repairs. In such repairs it achieves a recurrence rate comparable to that achieved by other mesh techniques for less complex hernias. PMID- 11265605 TI - Intracorporeal electrohydraulic lithotripsy in gallstone ileus. PMID- 11265604 TI - Intraperitoneal cisplatin-based chemotherapy for primary treatment of epithelial ovarian cancer. AB - INTRODUCTION: This study looks at the feasibility of intraperitoneal cisplatin based treatment in patients newly diagnosed with ovarian cancer in a community hospital setting and provides long-term follow-up of a cohort of patients. METHODS: Sixteen patients with epithelial ovarian cancer were studied. All patients underwent definitive surgical debulking. Patients were scheduled to be treated with six cycles of intraperitoneal cisplatin and either intravenous cyclophosphamide or intravenous doxorubicin at four week intervals. RESULTS: Fifteen patients were evaluable for response. All patients received at least three cycles of intraperitoneal therapy. All patients had an initial clinical response. Nine of 15 patients underwent second-look laparotomy; five of the nine patients had positive second-looks, none had residual macroscopic disease. Of the remaining six patients, five had clinical complete remissions and four are alive without recurrence. CONCLUSION: High-dose intraperitoneal cisplatin-based with sodium thiosulfate protection is generally well-tolerated and possibly an appropriate alternative first-line therapy for selected patients with epithelial ovarian cancer. PMID- 11265606 TI - Intravenous amiodarone in cardiac arrest. PMID- 11265607 TI - Firearm-related fatality surveillance in Hartford County, Connecticut. AB - OBJECTIVES: To evaluate the feasibility of implementing a firearm fatality surveillance system in Hartford County, Connecticut. METHODS: Medical examiner, police, and crime lab data were collected for firearm deaths occurring in Hartford County during 1997. Data included characteristics of victims, suspects, and firearms. We used standard criteria for evaluating an epidemiological surveillance system. RESULTS: The surveillance system detected 52 firearm-related fatalities; 31 were suicides and 21 were homicides. Handguns accounted for 50% of the suicides and 72% of the homicides. Sensitivity was 96%, specificity was 100%, representativeness adequate, simplicity enhanced by a common case identifier, flexibility constrained by the use of existing data, timeliness varied by data source, and system acceptable to all data sources. Estimated statewide cost is $200 per case, or $52,000 per year. CONCLUSION: Firearm injury surveillance in Hartford County is feasible and expansion to statewide coverage possible. The surveillance yielded considerable data at reasonable costs. PMID- 11265608 TI - [Association between soybean dust exposure, allergic sensitivity and profile of respiratory symptoms]. AB - The purpose of this study was to correlate soybean dust (SD) exposure, skin reactivity to soybean hull (SH) allergens, and symptoms of asthma and/or allergic rhinitis. A group of 365 subjects with asthma and/or allergic rhinitis and a control group of 50 individuals without respiratory symptoms were studied. The level of exposure to SD is defined as follows: 1) direct (DE); 2) indirect (ID), and 3) urban (UE). All subjects completed standard questionnaires. Skin tests with a SH extract and with common allergens were performed by the prick technique (SPT). Fifty-six (15.3%) patients and no subjects from control group had positive SPT (histamine index > or = 0.5) with a SH allergen extract. The percentages of positive SPT to SH extract were 38.7%, 20.3% and 8.4% in subjects with DE, IE and UE, respectively (p < 0.001). Monosensitization to SH was absent in all subjects. The percent of subjects with positive SPTs to mites (p < 0.01), pollen (p < 0.01) and molds (p < 0.05) were higher in subjects with a positive SPT to SH versus those with a negative SPT to SH. Sixty-six percent of subjects with DE and 13.6% of subjects with IE or UE reported respiratory symptoms after SD inhalation (Odds Ratio: 12.67 [2.4-74.9], p < 0.001). Compared to subjects exclusively sensitized to mites, patients sensitized to SH presented significantly different clinical characteristics. Soybean production has been increasing in Argentina during the last 20 years, determining an increase in the population exposed to chronic SD inhalation. This fact determines a high risk of sensitization and triggering of respiratory symptoms in atopic subjects. This study demonstrates that there is: 1) a high prevalence of skin reactivity to SH in subjects with asthma and/or allergic rhinitis from Argentina and that this prevalence is associated with the level of exposure to SD, and 2) an association between sensitivity to SH and severity of asthma. Measures to avoid release and inhalation of SD in rural areas from Argentina are needed. PMID- 11265609 TI - [The most uninspired intuition of an erudite]. PMID- 11265611 TI - [Cloning]. PMID- 11265610 TI - [Exactness and corrections]. PMID- 11265612 TI - [Antoni van Leeuwenhoek (1632-1723)]. PMID- 11265613 TI - [High prevalence of left anterior hemiblocking in the electrocardiogram in asymptomatic Chagas cardiomyopathy]. PMID- 11265614 TI - [Conscience objection]. PMID- 11265615 TI - [Detection of preclinical Cushing's syndrome in overweight type 2 diabetic patients]. PMID- 11265616 TI - [Mistreatment during medical residency. Residents' perception]. PMID- 11265617 TI - [Acute confusion syndrome in the hospitalized aged]. PMID- 11265618 TI - [Standard of birth weight for gestational age in 55706 healthy newborns in a public maternity of Buenos Aires]. AB - Birth-weight-for-gestational-age patterns in Argentina are scarce and outdated. The same study has been performed within our institution for 8 years already. Our hypothesis is that there could have been population changes with repercussions on fetal growth. The objectives were: 1) to determine new normal values of birth weight (BW)-for-gestational-age; 2) to study growth speed and acceleration, and 3) to compare these new results between trienniums. POPULATION: All liveborn babies between 1988-1998 (n = 67,857) were included. Those with BW lower than 500 g, gestational age (GA) lower than 25 weeks or mistakes in the appraisal of GA and outliers (birth-weight-for gestational-age > 2.5 DS of the median) were excluded. Those without maternal or obstetric history that could have influenced the BW were defined as "healthy newborns" (n = 55,706). The software Persi, that employs 34 of the 93 variables included in the Perinatal Clinical Record (SIP/OPS/OMS, Agustina v 5.1), was used. Birth weight median, standard deviation and error, coefficient of variation, skewness and kurtosis's coefficients, real and polynomial percentiles, standard distribution (Z), and the corresponding charts were generated in an automatic way for each gestational week and through the use of the method of least squares (polynomial models up to 4th grade). Results were as follows: maximum variability 15% as from the 30th week, maximum absolute speed in the 36th week (263 g/week) and a positive acceleration, up to the 36th week, and then a negative one (maximum -127 g/week2 in the 42nd week) was observed. Skewness varied between -0.247 (31st week) and 0.129 (38th week), and kurtosis was around 3, from which it can be inferred that the population has a normal distribution. Compared to the score z, the new curves showed a maximum error of 1.53% for the 10 percentile and 1.50% for the 50 percentile. By analyzing the data by trienniums (88-91, 91-93, 94-96 and 97-98) a growing mean BW (3243 +/- 539 g to 3286 +/- 508 g; p < 0.001) and 10 percentile (2600 to 2690 g, p < 0.001) trend was appreciated. IN CONCLUSION: new values of birth weight for-gestational-age were determined, and a secular increase trend of the mean birth weight (+43 g) was observed. PMID- 11265619 TI - [Characterization of 3 microsatellites of the cystic fibrosis gene in Argentine families]. AB - Argentine population is highly heterogeneous for the cystic fibrosis (CF) transmembrane regulator (CFTR) gene mutations. The study of 14 more common mutations identified both mutated alleles in only 51% of patients. This study confirmed the diagnosis of cystic fibrosis in these patients and enabled the detection of asymptomatic carriers in their families. However, in the remaining patients the direct molecular assay did not provide the necessary information for genetic counselling. To establish the mutated allele transmission in the affected families, negative for the most common mutations, three microsatellites (IVS17bTA, IVS8CA and IVS17bCA) located in intronic regions of CFTR gene were studied. In the 40 CF families analyzed, different allelic variants were detected: 15 for IVS17bTA, 10 for IVS8CA and 4 for IVS17bCA. Polymorphism information content and heterozygosity were high, except for IVS17bCA. By the simultaneous analysis of the three microsatellites we could counsel 100% of the families. Ours results show that these microsatellites are an excellent group of markers for linkage studies in cystic fibrosis families of the Argentine population. PMID- 11265620 TI - [Molecular analysis of the most frequent mutations associated with congenital adrenal hyperplasia secondary to 21-hydroxylase enzyme deficiency]. AB - Most cases (90%) of congenital adrenal hyperplasia (CAH) are secondary to steroid 21-hydroxylase enzyme deficiency (P450c21). In human, the P450c21 gene (CYP21B) is present along with a non functional pseudogene (CYP21A). These genes, located in chromosome 6, present a sequence homology of 98%. This high homology and the complexity of this gene locus brings about considerable difficulties in its molecular analysis and in the interpretation of the results. The aim of the present study was to elaborate an adequate strategy for the analysis of the most frequent mutations described in the CYP21B gene. A total of 77 patients with clinical and biochemical diagnosis of CAH secondary to P450c21 enzyme deficiency, as well as 170 unaffected relatives, were studied. They belonged to 73 unrelated families (146 chromosomes). The strategy allowed for the differentiation of patients with homozygous point mutations (PM), with PM in one allele and deletions, conversions, Ex3 or Cluster Ex6 PM in the other, even though parents were not always available for the study. Furthermore, it allowed for the discrimination of heterozygous deletions or conversions of the CYP21B gene from duplications of the non functional gene CYP21A, as well as CYP21B and A deletions from normal copies of the two genes. An exhaustive molecular analysis of this gene is necessary for an adequate characterization of the alterations present in this locus. PMID- 11265621 TI - Secretion from neuropeptide-treated splenocytes modifies ovarian steroidogenesis. AB - There are evidences for modulation of immune function by the sympathetic nervous system and its principal neurotransmitter norepinephrine (NE) through superior ovarian nerve (SON)-coeliac ganglion-noradrenergic postganglionic innervation of the spleen. Seven days after SON transection at 53 days of age, the rat splenocytes were isolated and then cultured for 48 h. These culture media, used to stimulate ovaries from 60-day-old intact rats (neither SON-transected nor sham operated) at diestrus 2 stage, in in vitro incubations, showed a decrease in progesterone release, an increase in estradiol release and no change in androstenedione release in relation to splenocyte culture media from control (sham-operated) rats. When splenocytes from SON transected (SON-t) rats were treated with vasoactive intestinal peptide (VIP) or neuropeptide Y (NPY), both at 10(-6) M for 24 h, their secretions increased the progesterone release while decreasing the estradiol release from the intact ovaries, compared with the secretions of untreated splenocytes from SON-t rats. Although the secretions of splenocytes treated with VIP decrease the androstenedione release from the ovaries, the treatment with NPY produced no change in hormone release. In the present paper the ovarian steroidogenic response, which was modified by the effects of an in vivo SON transection on spleen cells, was reverted by an in vitro system in which the splenocytes were treated with VIP or NPY. This could indicate that the spleen of SON-t rats does not receive those neuropeptides by neural route however, when they are added to splenocyte culture in vitro, the cell secretions revert the profile of steroid hormones released from the intact ovary. We also present functional evidence for modulation of the immune function by sympathetic nervous system and neurotransmitters other than NE. PMID- 11265622 TI - [Fatty acids of plasma and erythrocyte phospholipids in malnourished infants well fed by breast feeding or formula]. AB - Polyunsaturated fatty acids (PUFA) derived from essential fatty acids (EFA) play an important role in prenatal visual and neural development. Protein-energy malnutrition affects PUFA supply, and hence the synthesis of structural lipids during growth. Recently, some physiological studies reported abnormalities in the neurological functions of formula-fed infants relative to breast-fed. The purpose of our study was to assess whether fatty acid composition of the malnourished infant diet modifies plasma and erythrocyte phospholipid fatty acid composition. Three groups of full-term malnourished infants were selected according to their prior feeding. Two groups had received commercial formulas, one of them supplied with linoleic and alpha-linolenic acid, and the other supplied in addition with long chain PUFA from n-3 and n-6 series. The reference group of breast-fed infants was also enrolled. Plasma and erythrocyte phospholipid fatty acid composition was determined by gas-liquid chromatography. Those infants receiving formulas showed in plasma and erythrocyte phospholipids increased values in total saturated and monoethylenic fatty acids, and decreased values in polyunsaturated fatty acids from both n-6 and n-3 series, relative to that of breast-fed infants. These differences were more remarkable in the case of infants who received formula without PUFA. We conclude that in malnourished infants, a nutrient formula enriched with long chain fatty acids of n-6 and n-3 series could be helpful to achieve an erythrocyte and plasma fatty acid pattern similar to that obtained in breast-fed infants. PMID- 11265623 TI - Erythrophagocytosis assay in patients with autoimmune hemolytic anemia. AB - The aim of this paper is to evaluate the erythrophagocytosis assay (EA) in patients with autoimmune hemolytic anemia (AIHA). Direct antiglobulin test (DAT), indirect antiglobulin test (IAT) and EA were performed in blood samples from 46 patients with presumed AIHA. The EA was carried out incubating patients' erythrocytes and peripheral blood monocytes. A total of 200 monocytes were analysed to determine the percentage of active phagocytic cells (% APC). In 9 of these patients the applied treatment was evaluated by DAT, IAT and EA. In 14 transfusion requirements, the compatibility tests and EA were performed. For EA, patients' monocytes were incubated with erythrocytes from previously selected units sensitized with patients' sera. The % of APC was 32.1 +/- 1.7 in 35 patients with positive DAT and 17.8 +/- 1.3 in 11 patients with negative DAT. This last value was significantly higher than that with negative controls (3.7 +/ 0.3)(p < or = 0.01). As regards the applied treatment, patients with a successful response (n = 6) showed a significant decrease in the initial % APC (31.8 +/- 1.6 to 15.3 +/- 2.4; p < or = 0.05) while DAT and IAT remained positive. In those patients who required blood transfusion the compatibility tests were positive with all the units to be transfused, whereas the % APC varied for each one. Blood units were selected according to the lower % APC. PMID- 11265624 TI - [Magnetic resonance and clinical and electroencephalographical localization in focal epilepsy]. AB - Magnetic Resonance Imaging (MRI) is the method of choice to search for epileptogenic lesions. We correlated MRI findings with the epileptogenic zone (EZ) depicted by clinical and electroencephalographic (EEG) data. We studied 400 clinical records of patients who had been submitted to MRI studies and we analyzed, retrospectively, their ictal semiology, EEG characteristics and response to treatment. They were classified into 3 groups: A) temporal lobe epilepsy, B) frontal lobe epilepsy and C) parieto-occipital epilepsy. We included 155 patients: Group A) 68 cases (43.9%), 28 men (41.1%), mean age 32 +/- 11 years old, abnormal IMR in 44 (64.7%), refractory to treatment 48 (70.5%). Group B) 68 cases (43.9%), 38 men (55.8%), mean age 30 +/- 15 years old, abnormal IMR in 26 (38.2%), refractory to treatment 30 (44.1%). Group C) 19 cases (12.2%), 13 men (68.4%), mean age 27 +/- 11 years old, abnormal IMR in 11 (57.8%), refractory to treatment 12 (63.1%). Results showed that there were higher possibilities of detecting lesions which correlate with EZ in temporal than in frontal or parieto occipital lobes epilepsy. The chances to find abnormalities on the MRI were 5 times higher in refractory patients than in those who were non-refractory. PMID- 11265625 TI - [Endovascular treatment of partially clipped aneurysms]. AB - Partial clipping may occur in about 4% of surgical procedures. The risk of hemorrhage persists if the aneurysm is not completely excluded. Reoperations are often difficult, technically demanding and may carry an increased risk of complications. We report our experience with the use of Guglielmi detachable coils in the treatment of 9 aneurysm remnants. Five patients (55.6%) presented with a second subarachnoid hemorrhage. Eight of the aneurysms (88.9%) were located on the anterior circulation. Postoperative angiography showed complete occlusion in 8 cases (88.9%). Certain partial clipping types may assist and favor a stable coiling procedure allowing a more compact cast. On the other hand, the clip may interfere with the correct visualization of the neck. In this series, there was no neurological morbidity associated with the procedure. There were no hemorrhagic events during or after the embolization. Endovascular treatment of aneurysm remnants can be performed safely and may constitute a valuable option to microsurgery. PMID- 11265628 TI - [HIV infected macrophages isolated from HIV+ patients with undetectable viral load undergoing combined antiretroviral treatment]. AB - A new method of culture of peripheral blood leukocytes (PBMC) from HIV+ patients, in the absence of exogenous stimuli (allogeneic cells or cytokines) (PBMC w/s) was used for the detection of persistent viral infection in HIV patients who had undergone successful highly active antiretroviral therapy (HAART) lowering their viral burden to undetectable levels (< 50 RNA copies/ml). Infected cells were always of the monocyte/macrophage lineage (M). No infection could be detected in these patients using the classical system (co-culture with HIV-CMP activated with PHA and IL-2). Differences in the class of target cells (higher proportion of proliferating M and CCR5 expressing cells in the PBMC w/s system than in PBMC-PHA cultures) may determine the relative sensitivity of each technique to achieve successful isolation of HIV from different patients. PMID- 11265626 TI - [Criteria of low risk of mortality in children with neutropenia and fever during cancer chemotherapy]. AB - To validate the use of a lower-risk mortality profile in pediatric febrile neutropenia during anticancer therapy and to evaluate the efficacy of a sequential parenteral-oral antibiotic treatment for these children, a prospective study was conducted between May 1997 and December 1999. During this period 247 episodes in 215 patients were included in the present study. Children with neutropenia (ANC < 500/mm3) and fever (> 38 degrees C) due to anticancer therapy were eligible for the study if they presented the following lower-risk conditions: absence of severe co-morbidity factors, good clinical condition, no risk clinical foci, no bacteremia, and responsible parents. They were initially treated with inpatient parenteral short course of ceftriaxone and amikacin followed by ambulatory oral cefixime or ciprofloxacin to complete 7 days. Mean age was 64 (range: 8-200) months. The most common underlying malignant disease was acute lymphoblastic leukemia in 48% (118) of cases and 57% (141) of patients had an indwelling central venous catheter. Clinical evidence of infection was found in 47% (122) of children and the most common site was the upper respiratory tract (81%). Mean period of fever was 1.1 days (r: 1-8) and the duration of neutropenia was 3.9 days (r: 1-9). Sixty-one% (150) of children was discharged with neutropenia. Mean time of hospitalization was 1.5 days. Four clinical failures were detected (1.6%). They all were satisfactorily treated with a secondary treatment and none underwent any major complications or died. The lower risk profile used was safe and the sequential antibiotic therapy was adequate to manage febrile neutropenia in this subset of children. PMID- 11265627 TI - [Ultrastructural alterations in colonic mucosa of nifurtimox treated rats]. AB - Nifurtimox (Nfx) is a chemotherapeutic agent used in the treatment of acute Chagas' disease. Clinical and experimental studies with this nitroheterocyclic compound evidenced serious undesirable side effects. These were correlated with Nfx nitroreduction to a nitroanion radical followed by superoxide anion generation through a redox cycling process. The aim of this study was to verify whether the oral administration of Nfx to Sprague Dawley male rats (100 mg.kg-1, p.o.) produced any observable ultrastructural alteration in the cells of the colonic mucosa. Results showed that 24 h after Nfx administration there were observable alterations in this type of cells. They essentially consisted of moderate dilatation of their endoplasmic reticulum and intense dilatation of their Golgi complex. Already 1 and 3 h after Nfx administration, the original compound reached a concentration of 9.7 +/- 2.9 and 7.0 +/- 1.7 nmol.g-1 respectively in the colonic tissue. Studies on Nfx nitroreductase activity of colonic mucosa as determined spectrophotometrically and by HPLC methods showed that the microsomal fraction (from 0.72 +/- 0.29 to 0.26 +/- 0.04 nmol Nfx.min 1.mg-1 protein) but not the cytosol had the ability to nitroreduce Nfx. The results obtained show a correlation between the ultrastructural localization of injury and that of nitroreductase activity. The intense deleterious effects of Nfx in the Golgi apparatus suggest the potential occurrence of alterations in the synthesis/storage of secretory products of the colonic mucosa. PMID- 11265629 TI - [Rh system genotyping in amniotic fluid]. AB - The aim of this work was to determine the presence of the RHD gene in fetal cells obtained from amniotic fluid (AF). We studied 65 samples of AF, 11 from RhD- mothers sensitized with anti-D. The fetal origin of the DNA was confirmed with the analysis of 1 VNTR locus and 3 STR loci in DNA samples from AF and maternal blood. The RHD genotyping was performed in non contaminated samples (n = 62) using a multiplex PCR strategy that yields 3 amplification products from RhD+ phenotypes and 1 DNA fragment from RhD- phenotypes. We genotyped 54 RhD+ fetuses (8 from RhD- sensitized mothers) and 8 RhD- fetuses (3 from RhD- sensitized mothers). Fetal DNA genotyping allows the diagnosis, from a single amniocentesis, of fetuses at real risk of hemolytic disease of the newborn. When the fetus is determined to be RhD- all invasive procedures can be avoided. PMID- 11265630 TI - [Achromobacter xylosoxidans bacteremia in a patient with community-acquired pneumonia]. AB - Achromobacter xylosoxidans is a rare cause of bacteremia, and little information on treatment is available. The majority of patients who have developed Achromobacter bacteremia have presented predisposing causes to the infection. A case of community-acquired pneumonia and bacteremia due to A. xylosoxidans in a previously healthy patient is reported. Achromobacter is usually resistant to ampicillin, cephalosporins (1st, 2nd, and 3rd generation), aminoglycosides, and fluoroquinolones. Piperacillin, piperacillin-tazobactam, and trimethoprim sulfamethoxazole inhibit most isolates. PMID- 11265631 TI - [Kawasaki disease. Immunological evaluation of 26 cases]. AB - Kawasaki disease (KD) is an acute febrile vasculitis of childhood, characterized by multiple clinical and biochemical features of inflammation with special involvement of the heart. The activation of lymphocytes and monocytes/macrophages and their secreted soluble products, cytokines, play a central role in the pathogenesis of the disease. In this study we performed immunologic studies in 26 patients with KD. No constant pattern of serum levels of IgG, IgA, IgM, C3 and C4 fractions of complement measured by Nephelometry and neither autoantibodies, FAN and ANCA performed by indirect immunofluorescence were found in 22 patients in the acute stage. Variable percentages of CD3, CD4, CD8, CD20, CD56 and DR in peripheral mononuclear cells specifically stained and analysed by flow cytometry were seen among 25 patients in the acute stage. CD25 was elevated in 17/25 cases. Serum levels of TNF alpha performed by ELISA in 12 patients in acute stage were low. Intracellular cytokines such as TNF alpha, IL1 beta, IL2 and IFN gamma were measured in peripheral mononuclear cells of 15 patients in acute stage, in 5th and 30th days after gammaglobulin treatment, utilizing specific staining and analysis by flow cytometry showing no sole characteristic profile. In 2 patients there was an elevated percentage of TNF alpha and IL1 beta in monocytes during the convalescent stage; both had coronary sequelae. More research on this question is needed. In conclusion, immunologic studies showed an heterogeneous profile and no laboratory finding was registered in the acute stage that could be used as predictive factor of cardiovascular involvement. PMID- 11265632 TI - [Acquired partial lipodystrophy. Insulin resistance, hepatic lipase activity and small and dense LDL particles]. AB - Partial lipodystrophy (PLD) is an infrequent condition characterized by symmetric loss of subcutaneous adipose tissue in the upper or lower part of the body, although occasionally it affects only the extremities. In all cases it appears along with acantosis nigricans (AN), insulin resistance and impairment in the metabolism of lipids and carbohydrates. The case depicted pertains to a 49 year old female with no family history involving loss of adipose tissue in face and upper body. No fat in lower part of body was observed. The patient showed facial thinning at age 8, AN at 11 and gestational diabetes during her fourth pregnancy at 33. At present, the patient presents severe hyperglycemia and hyperinsulinemia with a marked insulin resistance. Type IV hyperlipoproteinemia (OMS), declined C HDL and Apo A1 and low C-LDL but with a high proportion of small and dense LDL particles were present. Non esterified fatty acids were high. Lipoprotein lipase and hepatic lipase activities are in the lower limit and increased respectively. Fraction C3 of the complement was diminished. No mutations were observed either in codons 170, 809 and 972 of the IRS-1 receptor or in codon 276 of the adrenergic beta 2 gene. PMID- 11265633 TI - [Galectins: a novel family of proteins involved in the regulation of the immune response. Implications in immunopathological processes]. AB - Galectins have emerged as a new family of closely related carbohydrate-binding proteins, which exert their functions by virtue of their ability to decipher glycocodes on complex glycoconjugates. They have been implicated in different immunological processes, such as lymphocyte adhesion, cytokine production, cell growth regulation, apoptosis and central and peripheral immune tolerance. In the present article we analyze the implications of this protein family in different immune pathologies with up- or down-regulated immune responses, such as autoimmune disorders, acute and chronic inflammation, allergic diseases, infection and metastases. The use of recombinant galectins or their antagonists will have future implications in the diagnosis, prognosis and treatment of these diseases, widening the horizons of molecular immunopathology. PMID- 11265634 TI - Phylogenetic conservation of the molecular and immunological properties of the chaperones gp96 and hsp70. AB - The heat shock proteins (HSP) gp96 and hsp70 have been shown to have a critical role in eliciting adaptive immune responses to cancers and viruses. This role derives from (i) their ability to chaperone antigenic peptides generated in the cells from which the HSP are isolated, and (ii) their capacity to interact with antigen presenting cells (APC) which re-present the HSP-chaperoned peptides in context of MHC I molecules. We have asked whether the immunological properties of HSP extend beyond the mammals to other phyla. We report here the serological, biochemical, genetic, and immunological characterization of the Xenopus gp96. Like mammalian gp96, Xenopus gp96 forms non-covalent complexes with peptides. Immunization with gp96 and hsp70 purified from Xenopus tumors, elicits potent and specific anti-tumor immunity, which is dependent on their ability to chaperone peptides in vivo. An immunogenic peptide chaperoned by the Xenopus gp96 can be processed and presented by mouse APC, to antigen-specific CD8+ T cells of mice. The remarkable conservation of these essential immunological properties of gp96 and hsp70 between amphibians and mammals suggests the importance of HSP in the evolution of the vertebrate immune system. PMID- 11265635 TI - The murine mutation scurfy (sf) results in an antigen-dependent lymphoproliferative disease with altered T cell sensitivity. AB - The scurfy (sf) murine mutation results in a rapidly fatal lymphoproliferative disease, causing death by 26 days. Mature CD4+ T cells which tested hyperresponsive to T cell receptor (TCR) stimulation are involved. When sf was bred onto a transgenic line (DO11.10) in which 75 - 95 % of the T cells express TCR for ovalbumin (OVA) 323 - 339, sf / Y OVA mice had prolonged lifespans and less severe clinical symptoms compared to controls. However, sf / Y OVA mice eventually developed disease and died with manifestations similar to those of the original sf strain. The Rag1 knockout (KO) mouse, which cannot produce mature T (or B) cells without the addition of functional transgenes, was chosen for further breeding. The combination of Rag1 KO, the OVA transgene, and sf produced mice with 100 % of their mature DO11.10 alpha beta T cells reactive strictly to OVA peptide. None of these Rag1 - / - sf / Y OVA mice developed the scurfy disease. They retained central deletion capability in vivo, but demonstrated an altered in vitro response to OVA peptide. These results indicate that mice without TCR for endogenous antigens do not develop scurfy symptoms, and are consistent with the hypothesis that the sf mutation requires antigen stimulation to manifest disease, perhaps via altered TCR sensitivity. PMID- 11265637 TI - Autoreactive isotype-specific T cells determine B cell frequency. AB - Suppressive activities involving T-B and T-T cell interactions are important to maintain immune system homeostasis. Negative control of IgG2ab+ B cells by anti IgG2ab T cells derived from Igha mice has been well documented. Nevertheless the real contribution of anti-IgG2ab T cells, endogenously matured in Ighb mice, in controlling IgG2ab+ B cell function has never been investigated. We previously generated anti-IgG2ab TCR-transgenic mice and showed that transgenic T cells were not deleted in the thymus and that they were responsible for a complete and chronic IgG2ab suppression. Here we show that T cells expressing high density of anti-IgG2ab TCR were positively selected in the thymus with a higher efficiency in animals expressing IgG2ab, reached peripheral lymphoid organs and negatively controlled IgG2ab serum levels. Moreover, anti-IgG2ab T cells transgenic for the single TCR chain, thus undergoing normal rearrangements and normal processes of selection, also reached the periphery and suppressed IgG2ab. Interestingly, concentration of IgG2ab in serum inversely correlated with the peripheral frequency of Ig-specific T cells. Finally, T cells able to suppress IgG2ab were obtained from Ighb non-transgenic mice, indicating that anti-2ab T cells are naturally present in the periphery of Ighb animals. We propose that IgG2ab specific T cells contribute to determine IgG2ab serum levels in Ighb mice. PMID- 11265636 TI - Strain-specific TCR repertoire selection of IL-4-producing Thy-1 dull gamma delta thymocytes. AB - Thy-1 dull gamma delta thymocytes constitute an unusual subset of mature TCR gamma delta cells which share with NK T cells the expression of cell surface markers usually associated with activated or memory cells and the simultaneous production of high levels of IL-4 and IFN-gamma upon activation. In DBA / 2 mice, Thy-1 dull gamma delta thymocytes express a restricted repertoire of TCR that are composed of the V1 gene product mainly associated with V6.4 chains exhibiting very limited junctional sequence diversity. In this study we have characterized this gamma delta T cell population in different strains of mice and show that Thy 1 dull gamma delta thymocytes are present in every strain tested, albeit at different frequencies. Moreover IL-4 production by gamma delta thymocytes is mainly confined to the Thy-1 dull population in every strain tested. Finally, the repertoire of TCR expressed by Thy-1 dull gamma delta thymocytes varies in different strain of mice, although a biased expression of Vgamma1 and Vdelta6 chains was observed in all strains studied. However, the extent of junctional diversity of the V1 and V6 chains expressed by Thy-1 dull gamma delta thymocytes varied from oligoclonal in DBA/2 mice to polyclonal in FVB/N mice. Thy-1 dull gamma delta thymocytes from mouse strains such as C3H/HeJ and BALB/c contain cells with diverse Vdelta6(D)Jdelta junctions together with cells with relatively homogeneous Vdelta6(D)Jdelta junctions, similar to those found in DBA/2. Thus, the Thy-1 dull gamma delta population appears to contain two subsets of cells which differ in the diversity of their TCR. PMID- 11265638 TI - Disruption of the IL-1beta gene diminishes acetylcholine receptor-induced immune responses in a murine model of myasthenia gravis. AB - Human autoimmune myasthenia gravis (MG) is associated with the IL-1beta TaqI RFLP allele 2. Individuals positive for this allele have high levels of inducible IL 1beta in their peripheral blood. Here, we have characterized MG induction and the immune response elicited by Torpedo acetylcholine receptor (AChR) immunization in wild-type and IL-1beta deficient (-/-) mice. Compared with wild-type mice, IL 1beta-/- mice were relatively resistant to induction of clinical experimental autoimmune myasthenia gravis (EAMG). Draining lymph node cells from IL-1beta-/- mice showed poor proliferative capacity upon AChR stimulation in vitro. Both Th1 (IFN-gamma, IL-2) and Th2 (IL-4) cytokine responses were reduced and levels of serum anti-AChR antibodies decreased in IL-1beta-/- mice compared to wild-type mice. Taken together, these results reveal a critical role for IL-1beta in the induction of MG in mice, and support a role for IL-1beta in the pathogenesis of MG in man. PMID- 11265639 TI - Identification of NKp80, a novel triggering molecule expressed by human NK cells. AB - The ability of NK cells to kill a wide range of tumor or virally infected target cells as well as normal allogeneic T cell blasts appears to depend upon the concerted action of multiple triggering NK receptors. In this study, using two specific monoclonal antibodies [(mAb) MA152 and LAP171], we identified a triggering NK receptor expressed at the cell surface as a dimer of approximately 80 kDa (NKp80). NKp80 is expressed by virtually all fresh or activated NK cells and by a minor subset of T cells characterized by the CD56 surface antigen. NKp80 surface expression was also detected in all CD3- and in 6 / 10 CD3+ large granular lymphocyte expansions derived from patients with lymphoproliferative disease of granular lymphocytes. In polyclonal NK cells, mAb-mediated cross linking of NKp80 resulted in induction of cytolytic activity and Ca2+ mobilization. A marked heterogeneity existed in the magnitude of the cytolytic responses of different NK cell clones to anti-NKp80 mAb. This heterogeneity correlated with the surface density of NKp46 molecules expressed by different NK clones. The mAb-mediated masking of NKp80 led to a partial inhibition of the NK mediated lysis of appropriate allogeneic phytohemagglutinin-induced T cell blasts, while it had no effect on the lysis of different tumor target cells, including T cell leukemia cells. These data suggest that NKp80 recognizes a ligand on normal T cells that may be down-regulated during tumor transformation. Molecular cloning of the cDNA coding for NKp80 revealed a type II transmembrane molecule of 231 amino acids identical to the putative protein encoded by a recently identified cDNA termed KLRF1. PMID- 11265640 TI - IgG subclass switch capacity is low in switched and in IgM-only, but high in IgD+IgM+, post-germinal center (CD27+) human B cells. AB - Recent studies have shown that in humans the germinal center reactions produce three types of V(D)J mutated B cells in similar proportions, i.e. Ig-switched, IgD-IgM+ (IgM-only) and IgD+IgM+ cells, and that together they form the CD27+ compartment of recirculating B cells. We investigated the Ig isotype switch capacity of these cells. Peripheral blood B subsets were sorted and IgG subclass secretion in presence or absence of IL-4 was compared in B cell assays which lead to Ig secretion in all (coculture with EL-4 thymoma cells) or only in CD27+ (CD40L stimulation) B cells. Already switched IgG+ B cells showed no significant sequential switch and IgM-only cells also had a low switch capacity, but IgD+CD27+ switched as much as IgD+CD27- B cells to all IgG subclasses. Thus, in switched B cells some alterations compromising further switch options occur frequently; IgM-only cells may result from aborted switch. However, IgD+CD27+ human B cells, extensively V(D)J mutated and "naive" regarding switch, build up a repertoire of B cells combining (1) novel cross-reactive specificities, (2) increased differentiation capacity (including after T-independent stimulation by Staphylococcus aureus Cowan I) and (3) the capacity to produce appropriate isotypes when they respond to novel pathogens. PMID- 11265641 TI - In situ cytokine therapy: redistribution of clonally expanded T cells. AB - Immunity to tumors relies on recirculating antigen-specific T cells. Whilst induction of antigen-specific T cells by immunotherapy has been convincingly proven, direct evidence for recirculation of such cells is still lacking. Here, employing a recently established in situ immunotherapy model for murine melanoma we directly demonstrate the redistribution of clonally expanded T cells. In this model IL-2 is targeted to the tumor microenvironment by means of specific antibody-IL-2 fusion proteins resulting in the expansion of T cells. The therapeutic effect of the fusion protein is not restricted to tumors expressing the targeted antigen, but extends to antigen negative variants of the tumor if present in the same animal. Analysis of the T cell infiltrate by quantitative reverse transcription-PCR revealed the presence of highly expressed TCR BV regions in both tumor variants. TCR clonotype mapping revealed that the high expressions of these regions were caused by clonal expansions and, notably, that these specific clonotypic TCR transcripts were identical in both tumors. Thus, T cell clones activated locally by targeted IL-2 therapy recirculate and mediate eradication of distant tumor sites not subjected to in situ cytokine therapy. PMID- 11265642 TI - Optimizing therapeutic strategies to inhibit circulating soluble target molecules with monoclonal antibodies: example of the soluble IL-6 receptors. AB - Therapeutic targeting of soluble molecules such as cytokines can be achieved with monoclonal antibodies (mAb). Anti-IL-6 mAb have been shown to form circulating complexes, resulting in the increase of the half-life of the cytokine in vivo. In IL-6-related diseases, the soluble human IL-6 receptors (shIL-6R), which have been shown to possess strong agonist activity, circulate in the plasma at a high concentration and must be neutralized. Their clearance was studied in mice that had been made to express circulating shIL-6R after i.p. grafting of mouse thymoma cells transfected with a gene coding for shIL-6R, treated with various anti-shIL 6R mAb recognizing different epitopes of the molecule. Injection of one anti-hIL 6R mAb stabilized the short-lived hIL-6R and led to their accumulation. The same result was observed when two mAb directed against two different epitopes of the hIL-6R were used. Clearance of the receptors was only achieved when three mAb specific for three different epitopes were injected. A permanent clearing of the hIL-6R could be obtained by repeated injections of the clearing mixture. No correlation was found between the ability of the mAb to clear the sIL-6R and to immunoprecipitate them in agarose gel. The F(ab')2 fragments lost the clearing ability of the intact mAb. These results clearly show that therapeutic clearance of sIL-6R by mAb need at least three mAb directed against three different epitopes of the molecule, a conclusion which is likely to apply for clearing any soluble target molecule. PMID- 11265643 TI - Neutrophil depletion exacerbates experimental Chagas' disease in BALB/c, but protects C57BL/6 mice through modulating the Th1/Th2 dichotomy in different directions. AB - To elucidate the roles of neutrophils in experimental Chagas' disease, we depleted the peripheral neutrophils in BALB/c and C57BL/6 mice with a monoclonal antibody 1 day before Trypanosoma cruzi infection. Neutrophil depletion in BALB/ c mice resulted in exacerbation of the disease and decreased expression of mRNA for Th1 cytokines, including IL-2 and IFN-gamma, IL-12p40 and TNF-alpha in their spleens after the infection, while a Th2 cytokine, IL-10, increased especially 1 day after infection. Neutrophils from infected BALB / c mice expressed mRNA for IL-12p40, IFN-gamma, TNF-alpha and Th1 chemoattractive chemokines, monokine induced by IFN-gamma (MIG) and macrophage inflammatory protein-1alpha (MIP-1alpha ). In contrast, in C57BL/6 mice neutrophil depletion induced resistance to the disease and enhanced the expression of the above Th1 cytokines, although IL-10 mRNA in neutrophil-depleted C57BL/6 mice was also higher than in control mice. Neutrophils from C57BL/6 mice did not express IL-12p40, IFN-gamma and MIG but expressed TNF-alpha, MIP-1alpha and IL-10. Therefore, neutrophils may play opposite roles in these two strains of mice with respect to protection versus exacerbation of T. cruzi infection, possibly through modulating the Th1/Th2 dichotomy in different directions. PMID- 11265644 TI - Beta1 integrin activation on human neutrophils promotes beta2 integrin-mediated adhesion to fibronectin. AB - Although the importance of beta1 integrin-mediated binding to adhesion molecules and extracellular matrix (ECM) molecules is well established for most types of leukocytes, the expression patterns and functional importance of beta1 integrins on neutrophils have remained controversial. Using flow cytometry, we found that human neutrophils express the alpha4, alpha5, alpha9 and beta1 integrin subunits. To examine whether the integrins VLA-4 (alpha4/beta1) and VLA-5 (alpha5/beta1) have a functional role on neutrophils, we studied adhesion to their ligand fibronectin. Treatment of neutrophils with antibody 8A2, which specifically binds and activates beta1 integrins, resulted in increased binding to fibronectin. However, addition of blocking mAb revealed that 8A2-induced adhesion did not depend on beta1 integrins, but on the beta2 integrin CD11b/CD18. Similarly, activation of beta1 integrins by 8A2 resulted in CD11b-dependent binding of neutrophils to fibrinogen. 8A2 treatment increased expression of an activation epitope of CD11b/CD18, which depended on phosphoinositide 3-OH kinase activity and an adequate concentration of intracellular free Ca2+. These data suggest that engagement of beta1 integrins on neutrophils results in a cross-talk signal that leads to activation of the beta2 integrin CD11b/CD18, followed by CD11b-mediated adhesion. As transmigrated neutrophils are surrounded by both beta1 and beta2 ligands in the ECM, this integrin cross-talk could play a role in modifying migration and cellular activation in inflamed tissues. PMID- 11265645 TI - Wnt signaling is required for thymocyte development and activates Tcf-1 mediated transcription. AB - T cell factor / lymphocyte enhancer factor (Tcf/Lef) transcription factors complex with the transcriptional co-activator beta-catenin to transduce Wnt signals in a variety of developmental systems. The prototypic family member Tcf-1 is highly expressed in T lineage cells. Tcf1-/- mice are defective in cell cycling of early thymocyte stages. Here, we show that the interaction of beta catenin with Tcf-1 is required for full thymocyte development. This interaction may be established by signals mediated by Wnt1 and Wnt4, leading to increased Tcf dependent transcriptional activity in thymocytes, as demonstrated in Tcf-LacZ reporter mice. Transduction of fetal thymocytes with Wnt1 and Wnt4 results in increased survival in an in vitro cell culture system. Retroviral expression of soluble Wnt receptor mutants that block Wnt signaling inhibits thymocyte development. These results imply an important role for the Wnt cascade in thymocyte development. PMID- 11265646 TI - Transforming growth factor-beta up-regulates CD40-engaged IL-12 production of mouse Langerhans cells. AB - Transforming growth factor (TGF)-beta is an immunosuppressive agent that is efficacious in suppressing a wide variety of cell-mediated immune responses. However, the direct effect of this cytokine on Langerhans cells (LC) has not been clarified. In this study, we examined its modulatory effects on the expression of co-stimulatory molecules and LC IL-12 production. A highly purified population of LC (>95%) was prepared from BALB/c mouse skin by the panning method using anti-I Ad mAb. Semiquantitative reverse transcription-PCR analysis showed that LC express TGF-beta receptor II mRNA. Interestingly, TGF-beta1 enhanced IL-12 p40 production of anti-CD40/IFN-gamma-stimulated LC, despite its down-regulatory effect on CD40 expression. A bioassay using an IL-12-dependent T cell line demonstrated the correlation of the IL-12 p40 level with the bioactivity of IL 12. More importantly, it was found that in contrast to TGF-beta, granulocyte/macrophage colony-stimulating factor (GM-CSF) strikingly inhibits IL 12 production of anti-CD40/IFN--stimulated LC and that the level of LC IL-12 production is determined by the relative amounts of TGF-beta1 and GM-CSF. Taken together, these results suggest that the two cytokines produced in the skin microenvironment, namely TGF-beta and GM-CSF, exert their important effects on LC function by regulating the secretion of IL-12, a cytokine influencing the Th1-Th2 balance. PMID- 11265647 TI - CpG motifs of DNA vaccines induce the expression of chemokines and MHC class II molecules on myocytes. AB - Determining how an immune response is initiated after in vivo transfection of myocytes with plasmids encoding foreign antigens is essential to understand the mechanisms of intramuscular (i. m.) genetic immunization. Since myocytes are facultative antigen-presenting cells lacking MHC class II and co-stimulatory molecules, it was assumed that their unique role upon DNA vaccination is to synthesize and secrete the protein encoded by the plasmid. Here we describe that i. m. injection of unmethylated CpG motifs induced the expression of chemokines (monocyte chemotactic protein-1) and MHC class II molecules on myocytes. Our results indicate that immunostimulatory DNA sequences (CpG motifs) of DNA vaccines augment synthesis of chemokine by myocytes with subsequent recruitment of inflammatory cells secreting IFN-gamma, a potent cytokine that up-regulates the expression of MHC class II molecules on myocytes. A myoblast cell line triple transfected with plasmids encoding MHC class II molecules and an immunodominant CD4 T cell epitope of influenza virus presented the endogenously synthesized peptide and activated specific T cells. These findings suggest that one mechanism for the immunogenicity of DNA vaccines consists in the presentation of peptides to CD4 T cells by in vivo plasmid-transfected myocytes. PMID- 11265648 TI - CD4+ T cells that evade deletion by a self peptide display Th1-biased differentiation. AB - We have examined factors governing the differentiation of autoreactive CD4+ T cells that have evaded deletion by a self peptide. Two lineages of transgenic mice (HA12 and HA104) expressing the influenza virus hemagglutinin (HA) were mated with TS1 mice that express a clonotypic T cell receptor (TCR) specific for the I-Ed-restricted determinant site 1 (S1) of HA. Thymocytes expressing high levels of the clonotypic TCR were deleted in both TS1xHA transgenic lineages. However, through allelic inclusion, thymocytes expressing low levels of the clonotypic TCR and high levels of endogenous TCR alpha-chains evaded deletion in TS1xHA12 and TS1xHA104 mice to graded degrees. When stimulated with S1 peptide in vitro, the non-autoreactive TS1 T cells were biased toward differentiation into Th2 effectors. By contrast, CD4+ T cells that evaded deletion in TS1xHA12 and TS1xHA104 mice were progressively biased toward Th1-like differentiation. Moreover, the effector cells from TS1xHA12 and TS1xHA104 mice secreted higher levels of IFN-gamma , on a per cell basis, than were secreted by their non autoreactive counterparts. Thus, CD4+ T cells that evade deletion by a self peptide can exhibit an intrinsic bias toward differentiation into Th1 effector cells. PMID- 11265649 TI - New version of didanosine. PMID- 11265650 TI - UNAIDS recommends therapy. PMID- 11265651 TI - Gene mutations identified. PMID- 11265652 TI - HAART and IL-2. PMID- 11265653 TI - 125 Gy or 135 Gy? Comments on the American Brachytherapy Society article by Beyer et al. IJROBP 2000;47:273-275. PMID- 11265654 TI - American Brachytherapy Society recommendations for clinical implementation of NIST-1999 standards for palladium-103 brachytherapy: in regard to Beyer et al. IJROBP 2000;47:273-275. PMID- 11265655 TI - In regard to the American Brachytherapy Society recommendations for 103palladium brachytherapy. Beyer et al. IJROBP 2000;47:273-275. PMID- 11265656 TI - In regard to Beyer, Nath, and Butler et al. IJROBP 2000;47:273-275. PMID- 11265657 TI - In regard to Altun et al. IJROBP 2000;47:401-404. PMID- 11265658 TI - Analysis of dose distribution in multiple-target gamma knife radiosurgery. PMID- 11265659 TI - [From genomics to therapeutics]. AB - The sequencing of the human genome will be achieved in the first years of the next century. This program is often presented as constituting a huge hope for medicine. It is sometimes expected that a disease-free world is a realistic prospect for tomorrow. Otherwise, the future of therapeutics is viewed as characterized by a personalized medicine in which each person will be preventively or curatively treated in function of its genetic make-up, assuring a maximal efficacy and the absence of toxicity. In fact, we have to be cautious, even if, indeed, progress are expected in chemotherapy and biological therapies. Expected difficulties will arise from the nature of the main diseases persisting in the developed countries, and from the economical situation in the developing ones. Fortunately for the modern doctors, their grand-grand children will still have the possibility to become themselves doctors: patients requiring treatments will still be there! PMID- 11265660 TI - Titanium alkali metal nitrido complexes. AB - Treatment of [(Ti(eta5-C5Me5)(mu-NH))3(mu3-N)] with alkali metal bis(trimethylsilyl)amido reagents in toluene afforded the complexes [M(mu3-N)(mu3 NH)2[Ti3(mu5-C5Me5)3(mu3-N)]]2 (M = Li (2), Na, (3), K (4)). The molecular structures of 2 and 3 have been determined by X-ray crystallographic studies and show two azaheterometallocubane cores [MTi3N4] linked by metal-nitrogen bonds. Reaction of the lithium derivative 2 with chlorotrimethylsilane or trimethyltin chloride in toluene gave the incomplete cube nitrido complexes [Ti3(eta5 C5Me5)3(mu-NH)2(mu-NMMe3)(mu3-N)] (M = Si (5), Sn (6)). A similar reaction with indium(I) or thallium(I) chlorides yielded cube-type derivatives [M(mu3-N)(mu3 NH)2[Ti(eta5-C5Me5)3(mu3-N)] (M=In (7), Tl (8)). PMID- 11265661 TI - Confidentiality for Teens. PMID- 11265662 TI - Indian Health Service oral health survey of American Natives. Preface. PMID- 11265663 TI - Sudden cardiac deaths rise 10% in young Americans. PMID- 11265664 TI - Severe endometriosis and apoptotic granulosa cells. PMID- 11265665 TI - Hyperinsulinemia: not involved in the development of PCO? PMID- 11265666 TI - The Chinese experience with elastomer vas deferens occlusion for reversible contraception. PMID- 11265667 TI - G20210A prothrombin gene mutation and other trombophilic polymorphisms in patients with portal or hepatic venous thrombosis. PMID- 11265668 TI - SPINK1 mutations in chronic pancreatitis. PMID- 11265669 TI - Mutations of the pancreatic secretory trypsin inhibitor (PSTI) gene in idiopathic chronic pancreatitis. PMID- 11265670 TI - Significance of genetic abnormalities after photodynamic therapy. PMID- 11265671 TI - Photodynamic therapy and genetic abnormalities. PMID- 11265672 TI - Optimal cases and sites to search for primary microbial agents in Crohn's disease. PMID- 11265673 TI - Image of the month. Von Hippel-Lindau (VHL) disease. PMID- 11265674 TI - Raynaud's phenomenon in workers exposed to vibration. PMID- 11265675 TI - Looking for the disorder in conduct disorder. AB - Besides their well-known externalizing behavior, children with conduct disorder (CD) often have additional impairments outside the criteria for the CD diagnosis. In a 5-year study of 984 treated children (ages 5-17 years), those with CD had an average of 2.2 primary diagnoses. Children with CD showed the worst problem and impairment scores in comparison with 11 common diagnoses. Compared with other treated children, children with CD achieved worse scores on 14 of 15 syndromes, including internalizing problems such as withdrawal and major depression. The average child with CD had larger relapse scores in the 1.5- to 3-year period after admission to treatment. This pattern, pervasive at intake and chronic in course, resembles a global disability more than a circumscribed problem managed with a narrow range of treatments specific to it. PMID- 11265676 TI - Transformation to acute leukaemia in an MDS patient harbouring bcr-abl and bcl 2 IgH rearrangements, without expression of either activated oncogene. PMID- 11265677 TI - Role of phenytoin in wound healing--a wound pharmacology perspective. AB - Topical agents used for the enhancement of wound healing are designed to act locally and, therefore, do not undergo classic systemic metabolic modification. This commentary reviews the potential role of a vulnerary agent, phenytoin, (PHT), from a wound pharmacology perspective. This agent may have the potential to alter the dynamics of wound healing, suggesting a therapeutic use for the stimulation of chronic wounds. Oral PHT therapy is used widely for the treatment of convulsive disorders, and about half the patients treated develop gingival overgrowth as a side-effect. This apparent stimulatory effect has prompted its assessment in wound healing. Investigations into the mechanisms of gingival overgrowth also provide clues to its action in wound healing, and important similarities and differences are discussed. It appears also that both gingiva and skin are important extrahepatic sites for xenobiotic metabolism, and analysis of the biochemical mechanisms should lead to the design of safer analogues for wound healing. On the other hand, differences between the pharmacokinetics of topical PHT in these tissue situations indicate that different formulations are required for gingival and cutaneous wound healing and during the changing course of wound healing itself. PMID- 11265678 TI - Human- and model-observer performance in ramp-spectrum noise: effects of regularization and object variability. AB - We consider detection of a nodule signal profile in noisy images meant to roughly simulate the statistical properties of tomographic image reconstructions in nuclear medicine. The images have two sources of variability arising from quantum noise from the imaging process and anatomical variability in the ensemble of objects being imaged. Both of these sources of variability are simulated by a stationary Gaussian random process. Sample images from this process are generated by filtering white-noise images. Human-observer performance in several signal known-exactly detection tasks is evaluated through psychophysical studies by using the two-alternative forced-choice method. The tasks considered investigate parameters of the images that influence both the signal profile and pixel-to pixel correlations in the images. The effect of low-pass filtering is investigated as an approximation to regularization implemented by image reconstruction algorithms. The relative magnitudes of the quantum and the anatomical variability are investigated as an approximation to the effects of exposure time. Finally, we study the effect of the anatomical correlations in the form of an anatomical slope as an approximation to the effects of different tissue types. Human-observer performance is compared with the performance of a number of model observers computed directly from the ensemble statistics of the images used in the experiments for the purpose of finding predictive models. The model observers investigated include a number of nonprewhitening observers, the Hotelling observer (which is equivalent to the ideal observer for these studies), and six implementations of channelized-Hotelling observers. The human observers demonstrate large effects across the experimental parameters investigated. In the regularization study, performance exhibits a mild peak at intermediate levels of regularization before degrading at higher levels. The exposure-time study shows that human observers are able to detect ever more subtle lesions at increased exposure times. The anatomical slope study shows that human-observer performance degrades as anatomical variability extends into higher spatial frequencies. Of the observers tested, the channelized-Hotelling observers best capture the features of the human data. PMID- 11265679 TI - Polarization and retinal image quality estimates in the human eye. AB - We have previously studied how polarization affects the double-pass estimates of the retinal image quality by using an imaging polarimeter [Opt. Lett. 24, 64 (1999)]. A series of 16 images for independent combinations of polarization states in the polarimeter were recorded to obtain the spatially resolved Mueller matrices of the eye. From these matrices, double-pass images of a point source for light with different combinations of incoming (first-pass) and outcoming (second-pass) polarization states were reconstructed and their corresponding modulation transfer functions were calculated. We found that the retinal image or, alternatively, the ocular aberrations, are nearly independent of the state of polarization of the incident light (in the first pass). This means that a significant improvement in the ocular optics by using a specific type of polarized light could not be achieved. However, quite different estimates of the retinal image quality are obtained for combinations of polarization states in both the first and the second passes in the double-pass apparatus. PMID- 11265680 TI - Dynamics of the eye's wave aberration. AB - It is well known that the eye's optics exhibit temporal instability in the form of microfluctuations in focus; however, almost nothing is known of the temporal properties of the eye's other aberrations. We constructed a real-time Hartmann Shack (HS) wave-front sensor to measure these dynamics at frequencies as high as 60 Hz. To reduce spatial inhomogeneities in the short-exposure HS images, we used a low-coherence source and a scanning system. HS images were collected on three normal subjects with natural and paralyzed accommodation. Average temporal power spectra were computed for the wave-front rms, the Seidel aberrations, and each of 32 Zernike coefficients. The results indicate the presence of fluctuations in all of the eye's aberration, not just defocus. Fluctuations in higher-order aberrations share similar spectra and bandwidths both within and between subjects, dropping at a rate of approximately 4 dB per octave in temporal frequency. The spectrum shape for higher-order aberrations is generally different from that for microfluctuations of accommodation. The origin of these measured fluctuations is not known, and both corneal/lenticular and retinal causes are considered. Under the assumption that they are purely corneal or lenticular, calculations suggest that a perfect adaptive optics system with a closed-loop bandwidth of 1-2 Hz could correct these aberrations well enough to achieve diffraction-limited imaging over a dilated pupil. PMID- 11265681 TI - Hyperefficient detection of targets in noisy images. AB - We compared human detection of visual targets in noisy images with that of a theoretically optimum matched filter. Using a small thin target with vertically aligned markers, we obtained hyperefficient detection as high as 91% as compared with the theoretical optimum, a value far exceeding the 30-50% value typically reported. When the markers were removed, detection efficiencies degraded to an average of 27%, even though subjects were aware that the target was always placed in the center of a reasonably small panel. Using a nine-alternative forced-choice experiment, we compared detection by human observers with a matched-filter computational observer on a trial-by-trial basis. With the markers present, when humans missed the correct panel, they most often chose the panel with the second highest decision variable output from the computational observer, suggesting that the template-matching model is a good one. To model results without the markers, we included location uncertainty and additional noise sources in the template matching of the computational observer. A location uncertainty of only 1 pixel, corresponding to a retinal distance of approximately 12 microm, a dimension of the order of the size of the receptive field of photoreceptors, explained the psychometric data. With the marker present, the model suggests that hyperefficient detection is obtained by limiting target location uncertainty to <6 microm. Together these results give important new insights into human visual detection mechanisms. PMID- 11265708 TI - The right ventricular tachycardias. AB - A variety of tachycardias originate from the right ventricle or use right ventricular structures as part of their circuit. They are characterized by a left bundle branch block pattern. Many of these tachycardias are relatively easy targets for radiofrequency catheter ablation. Ventricular tachycardia (VT) is the most common manifestation of arrhythmogenic right ventricular dysplasia, an often familial disease that can cause sudden death. Catheter ablation, antiarrhythmic drugs, or an implantable cardioverter-defibrillator may be used as therapy. Idiopathic right ventricular tachycardia has a benign course. It most often arises from the septal region of the right ventricular outflow tract. It commonly presents as nonsustained, repetitive monomorphic VT. The success rate of catheter ablation is greater than 90%. Bundle branch reentry occurs in patients with cardiomyopathy and His-Purkinje disease. It uses the right bundle branch anterogradely and the left bundle branch retrogradely. The QRS is very similar during VT and sinus rhythm. It can be cured by catheter ablation of the right bundle branch. VT seldom originates from the right ventricle in patients with coronary artery disease, idiopathic cardiomyopathy, or myocarditis. Atriofascicular (so-called Mahaim) fibers can sustain antidromic AV reentrant tachycardia. They represent an accessory AV node and His-Purkinje-like conduction system with atrial insertion in the right free wall near the tricuspid annulus and distal insertion directly into the right bundle branch. The accessory connection is ablated at the level of the tricuspid ring. PMID- 11265707 TI - Electrocardiographic features of inherited diseases that predispose to the development of cardiac arrhythmias, long QT syndrome, arrhythmogenic right ventricular cardiomyopathy/dysplasia, and Brugada syndrome. AB - Analysis of the 12-lead electrocardiogram (ECG) provides important diagnostic and prognostic information in the long QT syndrome. The clinical diagnosis of long QT syndrome is determined by the presence of a QTc > or = 0.44 sec. A normal QTc does not exclude a family member from being a genetic carrier. The ECG patterns of depolarization, the ST segment and shape of the T-wave can provide important clues as to the affected gene, particularly in conjunction with clinical information as to the precipitating causes of syncope or cardiac events. In arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D), the typical ECG pattern consists of T-wave inversion beyond lead V1. Evidence of right ventricular parietal block is manifest by a QRS duration in V1 > or = 110 msec and a longer QRS duration in the right then left precordial leads. Evidence of slow fractionated conduction is present as epsilon waves. The signal averaged ECG may show exceedingly long and low late potentials. Information regarding the risk of sudden death may also be obtained from the ECG. The ECG changes alone or in combination can provide strong evidence for the diagnosis of ARVC/D and helps to differentiate ARVC/D from right ventricular outflow tract (RVOT) tachycardia. The typical pattern of the ECG in the Brugada syndrome is ST segment elevation in the right precordial leads. This abnormality can be dormant and elicited by administration of drugs that cause Na channel blockade, such as ajmaline or type 1a or 1C antiarrhythmic drugs. Individuals who do not have the Brugada ECG findings at baseline but have this pattern induced by antiarrhythmic drugs are also at risk for sudden death. Further risk stratification may be obtained in the asymptomatic patients if ventricular fibrillation is induced at electrophysiological study. PMID- 11265709 TI - Genetics of brugada, long QT, and arrhythmogenic right ventricular dysplasia syndromes. AB - This article outlines the up-to-date understanding of the molecular basis of primary ventricular arrhythmias. Two disorders have recently been well described at the molecular level, the long QT syndromes and Brugada syndrome, and this article reviews the current scientific knowledge of each disease. A third disorder, arrhythmogenic right ventricular dysplasia, which is on the cusp of understanding, will also be described. PMID- 11265710 TI - Hemodynamic implications of left bundle branch block. AB - Left bundle branch block (LBBB), traditionally viewed as an electrophysiologic abnormality, is increasingly recognized for its profound hemodynamic effects. LBBB causes asynchronous myocardial activation, which, in turn, may trigger ventricular remodeling. Exercise nuclear studies frequently show reversible perfusion defects in the absence of obstructive coronary artery disease and some patients with intermittent LBBB develop angina coincident with the onset of LBBB. It is uncertain, however, if these phenomena are because of myocardial ischemia or ventricular asynergy. LBBB is associated with impaired systolic and diastolic function. In patients with dilated cardiomyopathy (DCM), LBBB is accompanied by progressive left ventricular (LV) dilatation and mitral regurgitation. It is not known whether LBBB is the cause or the consequence of LV dilatation. DCM patients with LBBB, as compared to those with normal intraventricular conduction, are more likely to have a nonischemic etiology, profound LV dilatation, lower ejection fraction, increased symptomatology, and shorter survival. Patients with DCM and acceleration-dependent LBBB may benefit from restoration of a narrow QRS complex by suppressing the heart rate with beta-blocker. There is extensive research underway in patients with DCM and LBBB to evaluate the short and long-term effects of normalization of ventricular activation sequence with high septal, LV, or biventricular pacing. PMID- 11265711 TI - Automated discrimination between atrial fibrillation and atrial flutter in the resting 12-lead electrocardiogram. AB - Computerized time-domain analysis of the QRST-subtracted 12-lead electrocardiogram (ECG) has been used successfully to determine several atrial activity patterns. These time-domain methods are particularly useful for low frequency signals such as those originating at the sinus node. However, high frequency atrial fibrillation (AFIB) and atrial flutter (AFL) waves can be better estimated by using spectral methods. In this study, we investigated the use of spectral entropy (SE) and spectral peak detection to distinguish fibrillatory from flutter activity in the QRST-subtracted ECG. In a set of 4,172 cardiologist overread ECGs, a computerized ECG analysis program (12SL MAC-Rhythm, GE-Marquette Medical Systems, Milwaukee, WI) detected 270 AFIB rhythms and 100 AFL rhythms. Compared to the cardiologist's reading, the AFIB versus AFL miss-classification error was 5.6%. The Fourier Transform was used to estimate the power spectral density of the QRST-subtracted ECG data. Individual lead spectra were then averaged and SE was computed for each of the ECGs originally called AFIB or AFL by the computer program. Additional criteria that included SE, spectral peak frequencies, and time-domain measures of atrial activity were then applied to discriminate between the 2 rhythms. Employing these criteria resulted in a decrease of miss-classification error to 2.5%. PMID- 11265712 TI - Multi-site dual surface monophasic action potential mapping of atrial repolarization in vivo: is atrial repolarization a two- or three-dimensional process? AB - Although the atrial free wall is a thin structure, atrial depolarization has been shown to have aspects of three-dimensionality. This study asks whether the same is true for atrial repolarization. By using a multi-element dual-surface probe, monophasic action potentials (MAPs) were recorded simultaneously at several opposing sites on the right atrial endocardial and epicardial surfaces in six open-chest pigs. The times of depolarization and repolarization were marked in recordings during sinus, paced rhythms, and during infusions of cold saline to the epicardial surface, which generated a temperature gradient across the atrial wall. Repolarization times were similar on endocardial and epicardial surfaces in some sites, but others showed significant differences during sinus and paced rhythms. Cold saline infusion produced a significant lengthening of MAP duration, and this was more pronounced on the atrial endocardial sites than on the epicardial sites. The observed differences in endocardial and epicardial repolarization times may be due to the presence of atrial pectinate muscles on the endocardial surface. These results suggest that in some regions atrial repolarization is a three-dimensional process. Possible limitations of this study include the fact that the depth of view of MAPs recorded from the atrial wall may extend to the opposing surface. PMID- 11265713 TI - Detection of atrial arrhythmia for cardiac rhythm management by implantable devices. AB - Implantable atrial defibrillators (IAD) should provide pacing therapy whenever appropriate (ie, typical atrial flutter) to minimize shock-related patient discomfort. Additionally, IADs should provide diagnostics regarding atrial arrhythmia type and frequency of occurrence to enable improved physician management of atrial arrhythmia. To achieve this, IADs should accurately classify atrial arrhythmia such as atrial fibrillation (AF) and atrial flutter (AFL) This article evaluates the performance of an algorithm, atrial rhythm classification (ARC), designed to classify AF and AFL. The ARC algorithm uses maximum rate, standard deviation, and range of the 12 most recent atrial cycle lengths to plot a point in a three-dimensional space. A decision boundary divides the space into 2 regions--faster/unstable atrial cycle lengths (AF) or slower/stable cycle lengths (AFL). Classifications are made on a sliding window of 12 consecutive cycles until the end of the episode is reached. In this way, continuous episode feedback is provided that can be used to help guide device therapy, measure arrhythmia type and frequency of occurrence. Bipolar (1-cm) electrogram episodes of AF (n = 16) and AFL (n = 7) were acquired from 20 patients and retrospectively analyzed using the ARC algorithm. The sensitivity and specificity in this study was 0.993 and 0.982, respectively. The ARC algorithm would have appropriately guided atrial therapy and minimized discomfort associated with defibrillation shocks in this small patient data set warranting further studies. The ARC algorithm may also be beneficial as a diagnostic tool to assist physician management of atrial arrhythmia. PMID- 11265714 TI - Evidence of heterogeneous remodeling in canine atrial fibrillation. AB - Although electrophysiologic changes occur during atrial remodeling, little is known how remodeling affects atrial fibrillation (AF) organization. We hypothesized that, in animals with long-term rapid atrial rates and a rapid ventricular response, AF would be more disorganized than in animals with rapid atrial rates only. In 8 dogs, chronic AF was created by 6 weeks of continuous rapid atrial pacing. In this group, the ventricular response to AF was spontaneous and unaltered. Twenty-one epochs of AF were epicardially mapped from the right and left atria. In 6 dogs, chronic AF was also created with rapid atrial pacing, however, the AV node was ablated and the ventricles were VVI paced at 80 BPM. Only 1 epoch of AF per dog was mapped. Atrial cycle length (CL) and spatial organization were compared. In chronic AF with a spontaneous ventricular rate, left atrial CL (96+/-14 ms) averaged 24 ms shorter than right atrial CL (121+/-18 ms) (P < .0001). With VVI pacing, AF CL was longer than in the dogs with the spontaneous ventricular rate. However, the left atrial CL (109+/-30 ms) was still significantly shorter than the right atrial CL (145+/-43 ms) (P < .001). Spatial organization values showed that during chronic AF with a spontaneous ventricular rate, the left atrium is more disorganized (2593+/-497) than the right atrium (2052+/-732) (P < .0001). With VVI pacing, the left atrium (2202+/-597) is still more disorganized than the right (1620+/-936) (P < .05). However, with VVI pacing, both atria appear less disorganized than dogs with VVI pacing. Atrial remodeling caused by heart failure that is superimposed on the remodeling due to rapid atrial rates causes the atria to be more disorganized than remodeling due to rapid atrial rates alone. However, in either case the left atrium is faster and more disorganized than the right atrium. Atrial fibrosis caused by the heart failure may increase the disorganization of AF activation. PMID- 11265715 TI - Dysrhythmia hazard after hospitalization for myocardial infarction: two ECG prognostic methods compared. AB - We retrieved reports of heart rate variability and signal-averaged electrocardiograms (SAECG) used to predict risk of a dysrhythmic event. From each report the number of cases with and without events was extracted to establish accurate values for true positive rate (tpr = sensitivity) and false positive rate (fpr = 1 minus specificity). For all the heart rate variability reports, these values were collected and tpr values were plotted versus fpr. The (fpr,tpr) data were summarized by a meta ROC graph using the method of Moses and Shapiro. A composite weighted mean value and 95% confidence interval were also derived. A summary meta-ROC curve for the SAECG reports was similarly obtained., Meta-ROC analysis of multiple reports better summarizes the performances of different prognostic methods and allows the effect of combining tests for a larger population to be simulated. PMID- 11265716 TI - Diagnostic accuracy of derived versus standard 12-lead electrocardiograms. AB - To compare the diagnostic yield of electrocardiograms (ECGs) recorded by 12 standard leads with that of 12-lead ECGs derived from 3 bipolar EASI leads, we analyzed pertinent ECG data for 290 normal subjects and 497 patients who had had a prior myocardial infarction (MI); the latter group comprised 36 patients with a non-Q MI, 282 patients with a Q-wave MI, and 179 patients with a history of ventricular tachycardia (VT). We first estimated statistically an optimal set of coefficients for deriving the 12 standard leads from EASI leads and assessed this transformation in terms of goodness of fit. To gauge the diagnostic information content of the recorded vs. derived 12-lead ECGs, we performed successively two group diagnostic classification--based on the Cardiac Infarction Injury Score (CIIS)--separating each of the patient subgroups from the normal group; the classification was repeated for 200 sets of patients selected randomly (with replacement), and the results were plotted as mean receiver operating characteristics. We found that derived 12-lead ECGs correlated well with the recorded ones, and reproduced faithfully the diagnostic features needed for the CIIS. When the CIIS was determined from features of the recorded standard 12 leads, its mean diagnostic performance (assessed in terms of area under the receiver operating characteristics curve) was 0.9004 for detecting non-Q MIs, 0.9546 for Q-wave MIs, and 0.9919 for MIs complicated by a history of VT. When, instead, features of derived 12 leads were used to determine the CIIS, diagnostic performance remained virtually unchanged (at 0.8905, 0.9531, and 0.9906, respectively). We conclude that, in our population, EASI-derived 12-lead ECGs contain nearly the same diagnostic information as standard 12-lead ECGs. PMID- 11265717 TI - Minimal lead sets for reconstruction of 12-lead electrocardiograms. AB - It may not always be possible to record all precordial leads of the standard 12 lead electrocardiogram (ECG). Especially in monitoring situations, a minimal lead set from which the 12-lead ECG can be reconstructed, would be valuable. This article assesses how well missing precordial leads could be synthesized from the remaining leads of the 12-lead ECG. A total of 2,372 diagnostic 12-lead ECG recordings were obtained from subjects with chest pain suggestive for acute myocardial infarction. Representative average beats were computed from the digital 12-lead ECG recordings with our Modular ECG Analysis System. The recordings were divided into a learning set and a test set. We considered all lead sets with one or more precordial leads removed, but always including limb leads I and II. By using the learning set, general reconstruction coefficients were computed to synthesize the missing precordial leads to each lead set. Performance of the synthesis was assessed by cross correlation between the original and the reconstructed leads. Also, patient-specific reconstruction coefficients were derived for each ECG in the test set and correlations were determined. High correlation coefficients were found with both reconstruction techniques. For different sizes of lead sets, the best patient-specific reconstructions had higher correlation values than the general reconstructions. For example, when 2 precordial leads were excluded, the best patient-specific median correlation was 0.994 compared to 0.963 for the best general reconstruction correlation. General reconstruction allows synthesis of 2 or 3 excluded precordial leads in good approximation. When patient-specific reconstruction can be applied, a minimal lead set including the limb leads and only 2 precordial leads suffices. PMID- 11265718 TI - Thoracic location of the lead with maximal ST-segment deviation during posterior and right ventricular ischemia: comparison of 18-lead ECG with 192 estimated body surface leads. AB - By using our database of continuous 18-lead electrocardiographic (ECG) recordings (standard + V3-5R + V7-9) during coronary angioplasty, we selected 68 patients with left circumflex balloon occlusions (posterior ischemia model) or proximal right coronary artery balloon occlusions (right ventricular IRV] ischemia model). ST-segment amplitudes (J + 60 ms) at preangioplasty baseline were subtracted from maximal ST amplitudes during balloon inflation to create a positive or negative change score (deltaST) for each of the 18 leads. DeltaST elevation was used to describe a change in the ST level in the positive direction from baseline, whether or not actual ST elevation from the isoelectric line was present. DeltaST depression was used to describe a change in the ST level in the negative direction from baseline, whether or not actual ST depression from the isoelectric line was present. ST amplitudes from 8 of the 12 standard leads were then used to estimate ST amplitudes at 192 body surface sites spanning the entire anterior and posterior thorax using the transformation technique of Lux. Thoracic distributions of the DeltaST values were displayed on a torso figure, including locations of the 18 lead locations and points of maximal ST elevation and depression. The 192 estimated body surface unipolar leads were compared with 18 lead ECGs (bipolar and unipolar). During 53 left circumflex occlusions, the maximal deltaST elevation was always located in the 18-lead ECG, with the most frequent locations at leads III, II (41%), V7-8 (34%), and V5-6 (25%). The maximal deltaST depression was located outside the 18-lead ECG (89%), with the most frequent locations above standard lead V2 (67%) and V3 (14%). During 16 proximal right coronary artery occlusions, the maximal deltaST elevation was always located in the 18-lead ECG, with the most frequent locations at leads III (81%) and V2-3R (13%). The maximal deltaST depression was located outside the 18 lead ECG (93%), with the most frequent locations above standard lead V2 (50%), V3 (14%), and V4 (14%). We conclude that maximal deltaST elevation is always located in the 18-lead ECG and maximal deltaST depression is frequently located outside of 18-lead ECG during left circumflex and proximal right coronary artery occlusions. Future studies are required to determine the bipolar leads for the 192 estimated body surface potential mapping leads. PMID- 11265719 TI - Derivation of an optimal lead set for measuring ectopic atrial activation from the pulmonary veins by using body surface mapping. AB - Atrial fibrillation is often initiated by atrial premature beats originating in the pulmonary veins. Non-invasive localization of these ectopic beats would be of significant value in guiding therapy. Body surface potential mapping was performed in nine patients undergoing invasive electrophysiologic study. Signals were recorded from 62 electrodes during pace mapping from each of the pulmonary veins. Optimal electrodes for localizing pulmonary vein activation were sequentially chosen. Seven optimal electrodes (6 anterior, 1 posterior) for recording ectopic atrial activation originating in the pulmonary veins were selected. The seven optimal electrode set performed better than the standard 9 electrode ECG at estimating the full body surface map (correlation 97 vs. 95.7%; p < 0.05). Seven optimally selected electrodes can estimate the body surface potential distribution during ectopic atrial activation orignating from the pulmonary veins. The ability of this electrode configuration to discriminate the site of origin of ectopic atrial beats requires prospective evaluation. PMID- 11265720 TI - Computing and visualizing electric potentials and current pathways in the thorax. AB - The long-term goal of electrocardiography is to relate electric potentials on the body surface with activities in the heart. Many previously reported studies have focused on direct links between heart and body surface potentials. The goals of this study were first to validate computational methods of determining volume potentials and currents with high-resolution experimental measurements and then to use interactive visualization of thoracic currents to understand features of the electrocardiographic fields from measured cardiac sources. We developed both simulation and experimental studies based on a realistic shaped torso phantom containing an isolated, perfused dog heart. Interventions included atrial pacing, single pacing and simultaneously pacing at multiple locations on the ventricles. Simulated torso volume potentials closely matched measured potentials in the torso-tank preparation (mean correlation coefficients of 0.95). Simulation further provided a means of estimating the current field in the torso from the computed torso volume potentials and the local geometric and conductive properties of the medium. Applying these techniques to the torso electric fields under a variety of pacing conditions, we have further demonstrated that thoracic current can provide many insights into the relationship between heart surface potential and body surface potentials. Specifically, we have shown that geometric factors including cardiac source configuration and location play an important role in determining to what extent electric activity in the heart is directly visible on the body surface electrocardiogram. The computation and visualization toolkit we developed in this study to explore current fields associated with cardiac events may provide new insights into electrocardiology. PMID- 11265721 TI - Continuous age-dependent normal limits for the pediatric electrocardiogram. PMID- 11265723 TI - A novel method to detect electrocardiographic electrode interchanges. PMID- 11265722 TI - Uncertainty of the electrocardiogram: old and new ideas for assessment and interpretation. AB - The electrocardiogram (ECG) is a highly complex, dynamic and stochastic phenomenon. Although it provides a valuable, noninvasive and rapid means of assessing cardiac state and its change, uncertainties in its measurement and variation in the underlying electrophysiology that generates the ECG make difficult further improvement in its reliability for detecting and monitoring cardiac pathologies and conditions. This article reviews the sources of variability and uncertainty in ECG measurement and interpretation, revisits some old ideas for dealing with them, and proposes some novel directions for improving accuracy of ECG assessment and interpretation. We shall explore relative information content of lead systems, representation of ECG signals and patterns, and estimation of ECG distributions from limited lead systems. In addition, we will compare strategies for measuring ECG information and suggest new paradigms for feature extraction that reduce the sensitivity of assessment accuracy to intrinsic and extrinsic measurement errors. Finally, we review the importance of including dynamic information in ECG assessment, both for interpreting current cardiac state as well as for monitoring its change and significance. PMID- 11265724 TI - A method to locate electrode placement. AB - A new method uses redundancy in the 12-lead electrocardiogram (ECG) to determine the angles to all of the electrodes used to record the ECG. No other transducers or signals are needed. The method, a matrix manipulation of the standard 12-lead, would be applicable to all existing ECGs already stored on hospital systems. The invention of this method was originally motivated by the slight differences seen between ECGs acquired by the standard resting electrode placement versus those acquired in a monitoring or exercise placement. An ECG signal is acquired in multiple channels. A covariance matrix is formed. From the eigenvector solution of the matrix, the angles between the eigenvectors and the original signal vectors are determined. The angles calculated for any ECG test are compared to reference angles to determine whether the electrodes are placed in the standard ECG electrode placement, an alternative electrode placement, or an incorrect electrode placement. PMID- 11265725 TI - Geometrical factors affecting the interindividual variability of the ECG and the VCG. AB - Various measures for quantifying the interindividual variability of the electrocardiogram and the vectorcardiogram in healthy subjects are presented. An analysis of factors that may cause this variability is performed, in particular of the geometrical factors of body size, heart size, heart position, and orientation. The results indicate that the variations in the magnitude of the electrocardiogram as observed through leads placed on the anterior thorax are dominated by the solid angle at which the outline of ventricular mass is seen from points on the thorax. Heart size and body size as such play only a secondary role. The limited spatial sampling of the anterior thorax directly overlaying the heart causes the mean values of all measures of amplitudes in women to be lower than in men. The vectorcardiogram magnitude was found to be much less dependent on overall geometry and heart position, and, hence, also to be less dependent on gender. PMID- 11265726 TI - Effects of heart position on the body-surface electrocardiogram. AB - Previous studies have examined the influence of body position, respiration, and habitus on body surface potentials. However, the authors could only estimate the sources of the effects they documented. Among the proposed origin of changes in body surface potentials from those studies were the position of the heart, alterations in autonomic tone, differences in ventricular blood volume, and variations in torso resistivity. The goal of this study was to investigate specifically the role of geometric factors in altering body surface potentials and the electrocardiogram. For this, we used experiments with an isolated, perfused dog heart suspended in a realistically shaped electrolytic torso tank. The experimental preparation allowed us to measure epicardial and tank surface potentials simultaneously, and then reconstruct the geometry of both surfaces. Our results mimicked some of the features described by previous investigators. However, our results also showed differences that included considerably larger changes in the peak QRS and T-wave amplitudes with heart movement than those reported in human studies. We detected smaller values of root-mean-squared variability from heart movements than those reported in a human study comparing body surface potentials during change in inspiration and body position. There was better agreement with relative variability, which in these studies ranged from 0.11 to 0.42, agreeing well with an estimate from human studies of 0.40. Our results suggest that the isolated heart/torso tank preparation is a valuable tool for investigating the effects of geometric variation. Furthermore, the geometric position of the heart appears to be a large source of variation in body surface potentials. The size of these variations easily exceeded thresholds used to distinguish pathologic conditions and thus such variations could have important implications on the interpretation of the standard electrocardiogram. PMID- 11265727 TI - Cellular basis for dispersion of repolarization underlying reentrant arrhythmias. AB - Substantial heterogeneity in ion channel density and expression exists in cells isolated from various regions of the heart. Cell-to-cell coupling in the intact heart, however, is expected to attenuate the functional expression of the ion channel heterogeneities. Due to limitations of conventional electrophysiological recording techniques, the extent to which cellular electrical heterogeneities are functionally present in intact myocardium remains unknown. High-resolution optical mapping with voltage-sensitive dyes was used to measure transepicardial and transmural repolarization gradients in the Langendorff perfused guinea pig ventricle and the canine wedge preperation, respectively. Diversity of repolarization kinetics in the transepicardial direction modulated dispersion of repolarization in a biphasic fashion as premature coupling interval was shortened. Moreover, modulation of repolarization paralleled arrhythmia vulnerability in a predictable fashion. Transmural optical mapping revealed significant gradients of repolarization across the ventricular wall that were markedly increased in a surrogate model of LQTS. Transmural gradients of repolarization in LQTS were associated with an enhanced susceptibility to TdP. Therefore, despite strong cell-to-cell coupling in the normal heart, heterogeneities in the ionic make-up of cells across the epicardial and transmural surfaces result in functional heterogeneities of repolarization leading to arrhythmias. PMID- 11265728 TI - False positive ECG reports of anterior myocardial infarction in women. AB - The prevalence of electrocardiographic poor R-wave progression was estimated by reviewing all electrocardiograms recorded in Glasgow Royal Infirmary over a 2 week period. It was found to be higher in women (19% vs. 11%) than in men. To investigate one possible reason, the effect of chest electrode positioning in women was thereafter examined. Eighty four women were recruited to a study in which chest electrodes were placed strictly in adherence with recommendations of using the 4th and 5th intercostal spaces as references and also using the more widely adopted technique of placing electrodes V3 to V6 under the left breast. R wave amplitudes were compared in V3 to V6 from both sets of recordings. It was found that measurements recorded on the breast by electrode V3 have a significantly smaller R wave magnitude compared to corresponding measurements below the breast, the mean difference being 34 (95% confidence interval [CI] of 7 to 60) microvolts. For V5 and V6, the reverse is true with measurements taken on the breast being larger, on average, than those taken below the breast by 119 (95% CI of 87 to 152) and 134 (95% CI of 108 to 160) microvolts respectively. For V4, there was no significant difference. Seventeen women with poor R wave progression suggestive of old anterior myocardial infarction had clinical data examined from which it was determined that 11 had a history suggestive of myocardial infarction, ie, the positive predictive value was 65% (95% CI of 42% to 87%). It was concluded that positioning of electrodes beneath rather than on top of the breast was not responsible for the increased prevalence of poor R wave progression in women and that the criterion of isolated poor R wave progression was too nonspecific to be of clinical value. PMID- 11265729 TI - ECG scores for a triage of patients with acute myocardial infarction transported by the emergency medical system. AB - Prehospital triage of cardiac patients for bypass from community hospitals to cardiac centers may improve survival. This article determines if electrocardiogram (ECG)-based scoring triage methods (Aldrich MI scoring, QRS distortion, and the TIMI classification) and location of infarct (via 12 lead ECG) are associated with mortality before and after adjusting for age, sex, and race. It is a retrospective study of 291 AMI adult patients transported by ambulance to community hospitals or cardiac centers. Patients with an ED chief complaint of chest pain or dyspnea, presence of MI as defined by ECG findings of 0.1 mV of ST segment elevation in two leads or positive CPK-MB were eligible for the study. The primary outcome variable was 2-year mortality as determined with a metropolitan Detroit tri-county death index. Logistic regression was used to calculate the unadjusted and adjusted odds ratios (with 95% CIs) of the predictor variables with mortality. Of the initial population selected for the study (n = 291), 229 patients were eligible for the analysis. The mean age was 66 years (SD of 14.4) with 63.8% being male and 54% being white. The overall mortality point estimate was 21.3% (95% CI of 15.2 to 27.3%). Aldrich scores and QRS distortion (yes/no) were not associated with mortality. Patients classified as a "high risk" for AMI per TIMI status were almost 3 times more likely to die than those at "low risk" and reached borderline statistical significance (P = .06) after adjusting for the covariates. Having an anterior infarct, as opposed to an inferior infarct, was significantly associated with death before and after adjusting for the covariates (Unadjusted OR = 2.6, Adjusted OR = 2.8). Properly training emergency medical system professionals in this area may prove useful for identifying higher risk AMI patients in the prehospital setting. PMID- 11265730 TI - A prediction model for prehospital triage of patients with suspected cardiac ischemia. AB - The American College of Cardiology recommends that patients with high risk acute myocardial infarction (AMI) be triaged to hospitals with percutaneous transluminal coronary angioplasty capability. However, there are no prehospital triage criteria to select candidates for bypassing community hospitals and being taken directly to "cardiac centers." This article assesses which independent variables predict death within 7 days in patients with suspected AMI transported by EMS. This is a retrospective study of 291 AMI patients transported by ambulance to 3 hospitals during 1996-1997. Included were patients who were (n = 244) > or =18 years of age, had a ED chief complaint of chest pain or dyspnea for whom we had mortality data. Mortality at 7 days, our primary outcome measure, was obtained by using a metropolitan Detroit tricounty death index records. Differences between the survivors and nonsurvivors were assessed using the Student's t-test and chi-square tests. Multiple triage criteria were assessed for optimal identification of high risk patients by constructing a logistic multivariate model. Among the study population, 15% died within 7 days (95% confidence interval (CI) 10.3-19.2), and this group represented 63.2% of all deaths over a 2 year surveillance period. Survivors, compared to nonsurvivors, were 14.1 years younger (P < or = .001) and more often men (P < or = 0.001). The dispatch time to ED arrival was less among survivors than nonsurvivors (42.8 vs. 50.6 min, P < or = .01). EMS vital signs differed by survivor status. Among survivors, HR was lower (-11.9 bpm; P < or = 0.01), RR was lower (-6.7 rpm; P < or = .001), SBP was higher (+14.5 mmHg; P < or = 0.05) and DBP was higher (+13.2 mm Hg; P < or = .01). A multivariate model identified the following as independent predictors of early mortality: female gender (OR = 2.3; P < or = .05), age > or =65 (OR = 5.9; P < or = .01), RR > or = 20 (OR = 4.6; P < or = .001), SBP < 120 (OR = 2.4; P < or = .05). The overall model was 86% sensitive and 53% specific with an area under the receiving operating characteristic curve of 0.8 (P < or = .001). A triage rule based on a multivariate model can identify the group at high risk of early cardiac death. This decision rule needs to be prospectively validated. PMID- 11265732 TI - The potential use of ECG-based prognostic instruments in clinical trials and cost effectiveness analyses of new therapies in acute cardiac ischemia. AB - The dramatic improvements in outcomes in acute cardiac ischemia because of therapeutic advances has led to "diminishing returns" with increasingly intensive therapies. This article explores the potential of electrocardiograph (ECG)-based prognostic instruments to identify patients likely to benefit from intense regimens, even in the absence of overall average benefit in the population, with 2 clinical examples: 1) Reperfusion therapy in acute myocardial infarction (AMI); and 2) anticoagulation/antiplatelet therapy in unstable angina. Based on previously developed, ECG-based prognostic instruments we explored the distribution of potential benefits in individual patients from increasingly intense therapy in both AMI and unstable angina. Predictions were obtained on community-based patient samples with both AMI and unstable angina to examine the distribution of effectiveness and cost-effectiveness. For both AMI and unstable angina, much of the benefit of intensifying therapy can be obtained by targeting a subgroup of patients that can be identified in multivariable dimensions by clinical and ECG characteristics. Treatment of these patients with more potent agents (such as hirudin or the glycoprotein inhibitors in unstable angina) is likely to be both effective and cost-effective. However, treatment of "low benefit" patients is unlikely to be effective or cost-effective, and some candidates for therapy are more likely to be harmed, than to benefit, by the more intensive regimens. Multivariable stratification can improve clinical and economic outcomes in acute cardiac ischemia, particularly when such models help identify "high benefit" patients early in their clinical course. Additionally, using validated models in the planning and execution of clinical trials of new therapies can improve the power of the trial and help target the therapies to patients most likely to benefit. PMID- 11265731 TI - Comparison of an automated thrombolytic predictive instrument to both diagnostic software and an expert cardiologist for diagnosis of an ST elevation acute myocardial infarction. AB - Because the electrocardiograms (ECGs) of patients with symptoms suggesting an acute thrombotic coronary occlusion are typically read by physicians relatively inexperienced in this skill, it is important to develop automated decision support. A Thrombolytic Predictive Instrument (TPI) is now available along with the standard diagnostic software in a commercially available electrocardiograph. This study evaluates the performance of the predictive software in comparison to both an expert cardiologist and standard diagnostic software. True sensitivity and specificity cannot be determined because acute coronary angiography was not performed. The specificities determined by this study were excellent (98% and 99%), and the sensitivities were very good (72% and 78%). These results that the TPI will be only rarely applied to patients who do not indeed have an acute coronary thrombosis. However, the reasons for even this small number of presumably falsely TPI positive patients should be determined and analyzed. It is unlikely that alterations of the thresholds for TPI activation will significantly improve on this very good level of sensitivity, without prohibitively decreasing specificity. PMID- 11265733 TI - The Nara ISCE 1999 meeting experience: progress report for 2000 and future directions. Presidental talk in Yosemite, California 2000. International Society for Computerized Electrocardiology. PMID- 11265734 TI - Cellular and ionic mechanisms responsible for the Brugada syndrome. AB - The Brugada syndrome is characterized by ST-segment elevation in the right precordial leads, V1-V3 (unrelated to ischemia or structural disease), normal QT intervals, RBBB pattern, and sudden cardiac death, particularly in men of Asian origin. An autosomal dominant mode of inheritance with variable penetrance is generally observed. The only gene mutations thus far linked to the Brugada Syndrome appear in the alpha subunit of the gene that encodes for the cardiac sodium channel, SCN5A. An outward shift in the balance of currents contributing to phase 1 of the right ventricular action potential is thought to underline to electrocardiographic manifestation of the syndrome. Strong sodium channel block, among other modalities, can accentuate the action potential notch in right ventricular epicardial cells, eventually leading to loss of the action potential dome. This results in the development of a large dispersion of repolarization within epicardium as well as between epicardium and endocardium, providing the substrate for the development of phase 2 and cirus movement reentry, which underline VT/VF. Therapy is directed at restoring the balance of current via inhibition of the transient outward current, Ito, and/or stimulation of inward calcium using beta adrenergic agonists, among several strategies. PMID- 11265735 TI - Pharmacological and device approach to therapy of inherited cardiac diseases associated with cardiac arrhythmias and sudden death. AB - A genetic origin in diseases like the long QT syndrome, the Brugada syndrome, or hypertrophic cardiomyopathy have been identified over the past years. These diseases have in common that they may result in sudden cardiac death of the patient. Recognition of patients based on their phenotype and application in clinical practice of the knowledge acquired on the genetic basis may have a major impact on how we approach them. In the long QT syndrome several mutations have been identified both in the sodium and in the potassium channels. The different electrophysiological effects of the mutations lead to a common phenotype: prolongation of the QT interval; but also to a common clinical impact: occurrence of malignant ventricular arrhythmias. Genetics should help us in treating in a more rational way our patients depending on the type of mutation. In the Brugada syndrome, mutations affecting the sodium channel have been so far identified. The results are electrophysiologically opposite to the ones observed in the long QT syndrome. Thus different mutations in the same gene lead to different functional consequences. Again, identification and study of the right mutation may lead to a more rational treatment directed to correct the malfunction of the channel. PMID- 11265736 TI - Clinical diagnosis and management strategies in arrhythmogenic right ventricular cardiomyopathy. AB - Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a myocardial disease, often familial, that is characterized by fibro-fatty replacement of the right ventricular (RV) myocardium. The most common clinical manifestations of ARVC consists of ventricular arrhythmias of RV origin, which may lead to sudden death mostly in young people and athletes, electrocardiograph depolarization/repolarization changes mostly localized to right precordial leads, and global and/or regional dysfunction and structural alterations of the RV. The diagnosis of ARVC may be difficult due to several problems with the specificity of the electrocardiograph abnormalities, the different potential etiologies of ventricular arrhythmias with a left bundle branch morphology, the assessment of the RV structure and function, and the interpretation of endomyocardial biopsy findings. Therefore, standardized diagnostic criteria have been proposed by the Study Group on ARVC of the European Society of Cardiology. According to these guidelines, the diagnosis of ARVC is based on the presence of major and minor criteria encompassing electrocardiograph, arrhythmic, morphofunctional, histopathologic, and genetic factors. Because the assessment of sudden death risk in patients with ARVC is still not well established, there are no precise guidelines to determine which are the patients who need to be treated and which is the best management approach. The therapeutic options include beta blockers, antiarrhythmic drugs, catheter ablation, and implantable cardioverter defibrillator. The implantable defibrillator is the most effective safe-guard against arrhythmic sudden death. However, its precise role in changing natural history of ARVC by preventing sudden and nonsudden death needs to be evaluated by a prospective study of a large series of patients. In patients in whom ARVC has progressed to severe RV or biventricular systolic dysfunction with risk of thromboembolic complications, treatment consists of current therapy for heart failure including anticoagulant therapy. In case of refractory congestive heart failure, the patients may become candidates for heart transplantation. PMID- 11265737 TI - Moving toward a new definition of acute myocardial infarction for the 21st century: status of the ESC/ACC consensus conference. European Society of Cardiology and American College of Cardiology. AB - The European Society of Cardiology and American College of Cardiology established their initial Joint Consensus Conference in July 1999 to develop a new definition of Acute Myocardial Infarction. This action was deemed necessary because of the development of new sensitive biochemical markers of myocardial necroses: Troponins T and I. There were 5 working groups, including one in Electrocardiography. The Conference adopted a definition that required only a history of "ischemic symptoms" and "a typical rise and fall" of at least one of the biochemical markers. The ECG Working Group strongly advised that a term distinctive from "myocardial infarction" such as "myocardial necrosette" be adopted as the diagnosis for an acute coronary event during which the peak biochemical marker level is below that which occurs when serial evolutionary ECG changes are observed. A pilot substudy from the GUSTO IIa Clinical Trial has identified the low end of the "ECG Change Range" to be: >2x the upper limit of normal for CK-MB, > 11 x for Troponin T, and >6 x for Troponin I. PMID- 11265738 TI - Evolving non-Q wave versus Q wave myocardial infarction after thrombolysis: a high risk population benefitting from early revascularization. A DANAMI substudy. PMID- 11265739 TI - ST-Segment resolution as a marker of epicardial and myocardial reperfusion after thrombolysis: insights from the TIMI 14 and in TIME-II trials. AB - Patients with persistent ST segment elevation after fibrinolysis are at high risk for death and congestive heart failure, even if normal epicardial flow has been restored. In the TIMI 14 trial, combination therapy with abciximab plus reduced dose alteplase enhanced the speed and efficacy of epicardial reperfusion. We also found that combination therapy provided an additional benefit in terms of myocardial reperfusion, as evidenced by greater ST resolution on serial 12-lead electrocardiograms. Specifically, the proportion of patients with complete (> or =70%) ST resolution was higher among patients receiving combination therapy than in those treated with alteplase alone (59% vs 37%; p < 0.0001). Even among patients with normal (TIMI grade 3) epicardial blood flow, combination therapy was associated with a significantly greater likelihood of complete ST resolution than was fibrinolysis alone (69% vs 44%; p = 0.0002). In conclusion, combination reperfusion therapy improved myocardial (microvascular) reperfusion, independent of epicardial flow, suggesting an additional mechanism by which abciximab may improve outcomes in patients with acute MI. PMID- 11265740 TI - The predischarge electrocardiographic pattern in anterior acute myocardial infarction: relation between evolutionary ST segment and T-wave configuration and prediction of myocardial infarct size and left ventricular systolic function by the QRS Selvester score. AB - Left ventricular systolic function, determined mainly by final infarct size, has a major influence on prognosis after acute myocardial infarction (MI). It was found that infarct size and left ventricular ejection fraction can be predicted using the Selvester QRS-score in patients not receiving reperfusion therapy. We assessed whether the predischarge QRS-score can be used for estimating infarct size and left ventricular ejection fraction in 51 patients with a first anterior MI who had received reperfusion therapy and whether considering the configuration of the ST-segments and T-waves will increase the accuracy of these predictions. All patients had received reperfusion therapy and had predischarge resting 99mTc sestamibi scan. We determined the Selvester QRS score using the electrocardiograms performed on the same day of the scan. In addition, we divided the patients into 3 groups: A: isoelectric ST and negative T-waves (n = 12); B: ST elevation (> or =0.1 mV) and negative T-waves (n = 23); and C: ST elevation (> or =0.1 mV) and positive T-waves (n = 16). The myocardial perfusion defect extent increased from group A to C (28.5+/-16.4%, 39.4+/-14.8%, and 45.3+/-15.8% in groups A, B, and C. respectively; P = .022). Similarly, the left ventricular ejection fraction decreased (41.7+/-11.6%, 38.4+/-8.1%, and 32.0+/-9.7%, respectively; P = .042) from group A to C. Overall, the correlation between the QRS-score and the myocardial perfusion defect extent (Rho = 0.249; P = .08), and ejection fraction (Rho = -0.229; P = .11) was not good. A statistically significant correlation between the myocardial perfusion defect size and the QRS score was found only in group A (Rho = 0.599, P = .04). In patients with a first anterior myocardial infarction who underwent reperfusion therapy, the predischarge QRS-score is predictive of infarct size only in those in whom ST elevation resolved completely. In patients with residual ST elevation the Selvester QRS-score is inaccurate in predicting infarct size and left ventricular ejection fraction upon discharge. PMID- 11265741 TI - Is there longitudinal dissociation in the undamaged his bundle? In vitro studies in the normal canine heart. AB - We investigated the concept of longitudinal dissociation in the His-Purkinje system in vitro. Hearts were excised from the eleven anesthetized dogs and a septal preparation containing the exposed His bundle and the entire right bundle branch and left bundle branch were displayed in a two-dimensional arrangement pinned to the bottom of a superfusion chamber. Tyrodes solution, gassed with 95% O2 and 5% CO2, at 37 degrees C was continuously passed over the preparation. Pacing was performed over a wide range of heart rates (30 to 180/min) from the proximal His bundle and by moving bipolar electrodes we monitored activation at various sites along the right and left bundle branch. The earliest site of muscle activation (27+/-2 msec) on the left septum was a relatively large area in the midposterior septal region; whereas, on the right septum the earliest site of activation (27+/-3 msec) was a relatively small zone at the base of the anterior papillary muscle, (p, N.S). The larger area of early activation on the left compared to the right is consonant with a left to right septal vector accounting for the Q wave in the standard bipolar leads in the normal heart and the loss of Q waves in left bundle branch block. We conclude that His-Purkinje and ventricular muscle activation is remarkably synchronous on both sides of the heart and accounts for optimal contractile function during His bundle or biventricular pacing compared to standard site ventricular pacing, particularly in patients with left ventricular dysfunction. PMID- 11265742 TI - Value of the ECG in suspected acute myocardial infarction with left bundle branch block. AB - Uncomplicated left bundle branch block (LBBB) is characterized by true ST-segment shifts resulting from delayed repolarization in the left ventricle with respect to the right ventricle. When acute coronary occlusions develop in the setting of previous or new LBBB, 12-lead eCG manifestations of injury may also appear. They consist of a more pronounced ST-segment elevation, of ST-segment deviations opposite to those of uncomplicated LBBB, or both. We have reported that the only 3 independent ECG signs of acute MI during LBBB among patients with chest pain or history of coronary disease are: ST elevation > or = 1 mm in leads with a positive QRS, ST-depression > or = 1 mm in V1 to V3, and ST elevation > or = 5 mm in leads with a negative QRS. In our study, the clinical prediction rule score values of these signs were 5; 3; and 2, respectively. A score > or = 3 made a diagnosis of MI with a 90% specificity and a score of 2 with > 80%, specificity. Recent validation studies have confirmed that the presence of any of these ECG signs is associated with a sensitivity of 44 to 79% and a specificity of 93 to 100%. Sensitivity increases if serial or previous ECGs are available for comparison. Interobserver agreement is very high. While current practice guidelines recommend thrombolysis for all patients with chest pain and LBBB, concern among physicians about hemorrhagic stroke prevents many of these patients from receiving timely treatment. In a population with LBBB and chest pain where our proposed ECG criteria were not ascertained, only 73% of eligible patients received thrombolysis; on the other hand, 48% of patients with no biochemical evidence of MI were thrombolyzed. For the latter group, the clinical prediction rule had a score of 0. Instead, 79% of patients with confirmed acute MI had a prediction rule score > or =2. Similar values applied to a subgroup of patients with serial ECGs. We propose that thrombolysis among patients with chest pain and LBBB be decided on the basis of a systematic ECG review to "rule patients in". This provision may result in both a significant reduction in the number of patients without infarction who receive thrombolysis and in timely treatment of those who do have MI. PMID- 11265743 TI - Left bundle branch block and right axis deviation: a report of 36 cases. AB - Right axis deviation (RAD) with left bundle branch block (LBBB) is a rare combination. From a database of 636,000 electrocardiograms we report a series of 36 patients with this disorder. The majority of subjects had dilated cardiomyopathy with biventricular enlargement. LBBB was fixed in 21 of 36 cases. It was freshly acquired, episodic, intermittent, or physiologic in 15 of 36. The RAD was episodic in 30 of 36; it was fixed and concurrent with LBBB in only 2 cases, and never episodically concurrent. Reported for the first time here were 4 of 36 cases in which the combination of LBBB and RAD was elicited with atrial premature impulses as a rare form of QRS aberration. In one case where the combination was intermittent, a clear relationship with freshly acquired intermittent posterior fascicular block was demonstrated. The possible relationship of the deviation with variable degrees of right ventricular overload is discussed. PMID- 11265744 TI - The key is in the genes, or is it...? With the human genome project completed, the question is 'what comes next'? PMID- 11265746 TI - The flight from European science. The attraction of the American research system for young European scientists. PMID- 11265745 TI - JBIC will pool Japanese genomics research. Japan's efforts to catch up in bioinformatics. PMID- 11265747 TI - Upwardly mobile proteins. Workshop: the role of HMG proteins in chromatin structure, gene expression and neoplasia. PMID- 11265748 TI - Networks of tumor suppressors. Workshop: tumor suppressor networks. PMID- 11265749 TI - Creating a transloxation. Engineering interchromosomal translocations in the mouse. PMID- 11265750 TI - Mini- and microsatellite expansions: the recombination connection. AB - It is widely accepted that the large trinucleotide repeat expansions observed in many neurological diseases occur during replication. However, genetic recombination has emerged as a major source of instability for tandem repeats, including minisatellites, and recent studies raise the possibility that it may also be responsible for trinucleotide repeat expansions. We will review data connecting tandem repeat rearrangements and recombination in humans and in eukaryotic model organisms, and discuss the possible role of recombination in trinucleotide repeat expansions in human neurological disorders. PMID- 11265751 TI - Inter-chromosomal recombination of Mll and Af9 genes mediated by cre-loxP in mouse development. AB - Chromosomal translocations are crucial events in the aetiology of many leukaemias, lymphomas and sarcomas, resulting in enforced oncogene expression or the creation of novel fusion genes. The study of the biological outcome of such events ideally requires recapitulation of the tissue specificity and timing of the chromosomal translocation itself. We have used the Cre-loxP system of phage P1 to induce de novo Mll-Af9 chromosomal recombination during mouse development. loxP sites were introduced into the Mll and Af9 genes on chromosomes 9 and 4, respectively, and mice carrying these alleles were crossed with mice expressing Cre recombinase. A resulting Mll-Af9 fusion gene was detected whose transcription and splicing were verified. Thus, programmed interchromosomal recombination can be achieved in mice. This approach should allow the design of mouse models of tumorigenesis with greater biological relevance than those available at present. PMID- 11265752 TI - Inducible chromosomal translocation of AML1 and ETO genes through Cre/loxP mediated recombination in the mouse. AB - Transgenic mice have been used to explore the role of chromosomal translocations in the genesis of tumors. But none of these efforts has actually involved induction of a translocation in vivo. Here we report the use of Cre recombinase to replicate in vivo the t(8;21) translocation found in human acute myeloid leukemia (AML). As in the human tumors, the murine translocation fuses the genes AML1 and ETO. We used homologous recombination to place loxP sites at loci that were syntenic with the break points for the human translocation. Cre activity was provided in mice by a transgene under the control of the Nestin promoter, or in cultured B cells by infecting with a retroviral vector encoding Cre. In both instances, Cre activity mediated interchromosomal translocations that fused the AML1 and ETO genes. Thus, reciprocal chromosomal translocations that closely resemble rearrangements found in human cancers can be achieved in mice. PMID- 11265753 TI - The DNA segregation mechanism of Epstein-Barr virus nuclear antigen 1. AB - Latent Epstein-Barr virus (EBV) genomes are maintained in human cells as low copy number episomes that are thought to be partitioned by attachment to the cellular mitotic chromosomes through the viral EBNA1 protein. We have identified a human protein, EBP2, which interacts with the EBNA1 sequences that govern EBV partitioning. Here we show that, in mitosis, EBP2 localizes to the condensed cellular chromosomes producing a staining pattern that is indistinguishable from that of EBNA1. The localization of EBNA1 proteins with mutations in the EBP2 binding region was also examined. An EBNA1 mutant (delta325-376) disrupted for EBP2 binding and segregation function was nuclear but failed to attach to the cellular chromosomes in mitosis. Our results indicate that amino acids 325-376 mediate the binding of EBNA1 to mitotic chromosomes and strongly suggest that EBNA1 mediates EBV segregation by attaching to EBP2 on the cellular mitotic chromosomes. PMID- 11265754 TI - Fate of mat1 DNA strands during mating-type switching in fission yeast. AB - The mating-type switching of the fission yeast, Schizosaccharomyces pombe, is highly regulated. Two consecutive asymmetric divisions are required to produce one mating-type switched cell among the four progeny. Using DNA density-gradient centrifugation we demonstrate that one-fourth of the mat1 DNA is not replicated by the conventional semi-conservative mode, but instead both DNA strands are synthesized de novo. Our data are consistent with a gene conversion event, initiated by a site- and strand-specific DNA break (SSB). We further demonstrate that the virgin switched mat1-containing chromatid no longer contained the nick, while it is reintroduced during the lagging strand synthesis of the mat1 locus on the sister chromatid. This finding establishes at the molecular level a firm experimental link between the phenotype and genotype in the process of asymmetric mating-type switching during mitotic divisions. PMID- 11265755 TI - Synergy between estrogen receptor alpha activation functions AF1 and AF2 mediated by transcription intermediary factor TIF2. AB - The activation function AF2 in the ligand-binding domain of estrogen receptors ER alpha and ER beta signals through the recruitment of nuclear receptor coactivators. Recent evidence indicates that coactivators, such as the transcription intermediary factor TIF2, also bind to and transactivate the N terminal AF1 function of the two ERs. We have generated TIF2 mutant proteins that are deficient in either AF1 or AF2 interaction and use these mutants to investigate the relative contribution of both AFs to TIF2 recruitment and transactivation. We observe that TIF2 is capable of interacting simultaneously with both the isolated N- and C-terminus of ER alpha in transfected mammalian cells and in vitro, indicating that TIF2 can bridge both receptor domains. The concomitant interaction of TIF2 with both AFs results in synergistic activation of transcription. Thus, synergy between ER alpha AF1 and AF2 is a result of the cooperative recruitment of TIF2 and/or other members of the p160 coactivator family. PMID- 11265756 TI - Decoding apparatus for eukaryotic selenocysteine insertion. AB - Decoding UGA as selenocysteine requires a unique tRNA, a specialized elongation factor, and specific secondary structures in the mRNA, termed SECIS elements. Eukaryotic SECIS elements are found in the 3' untranslated region of selenoprotein mRNAs while those in prokaryotes occur immediately downstream of UGA. Consequently, a single eukaryotic SECIS element can serve multiple UGA codons, whereas prokaryotic SECIS elements only function for the adjacent UGA, suggesting distinct mechanisms for recoding in the two kingdoms. We have identified and characterized the first eukaryotic selenocysteyl-tRNA-specific elongation factor. This factor forms a complex with mammalian SECIS binding protein 2, and these two components function together in selenocysteine incorporation in mammalian cells. Expression of the two functional domains of the bacterial elongation factor-SECIS binding protein as two separate proteins in eukaryotes suggests a mechanism for rapid exchange of charged for uncharged selenocysteyl-tRNA-elongation factor complex, allowing a single SECIS element to serve multiple UGA codons. PMID- 11265757 TI - RNA degradation buffers asymmetries of transcription in Arabidopsis mitochondria. AB - To understand better the relative contributions of transcriptional and post transcriptional processes towards the regulation of gene expression in plant mitochondria, we compared the steady state levels of RNAs with the respective transcriptional activities. All of the protein and rRNA coding genes of the Arabidopsis mitochondrial genome and several orfs were analyzed by run-on and northern experiments. rRNAs constitute the bulk of the steady state RNA in Arabidopsis mitochondria, but are (different from maize mitochondria) not equally prominent among the run-on transcripts. Their relatively low rate of active transcription is apparently compensated by their high stability. Run-on transcription values differ significantly between genes coding for different subunits of the same protein complex. The steady state RNA levels are considerably more homogeneous, indicating that high variations of transcription rates are counterbalanced by post-transcriptional processes. The relative amounts of the steady state transcripts for the different subunits in a given protein complex reflect the relative stoichiometries of the protein subunits much more closely than the respective transcriptional activities. Post-transcriptional RNA processing and stability thus contribute significantly to the regulation of gene expression in Arabidopsis mitochondria. PMID- 11265758 TI - TIF-IA, the factor mediating growth-dependent control of ribosomal RNA synthesis, is the mammalian homolog of yeast Rrn3p. AB - Cells carefully modulate the rate of rRNA transcription in order to prevent an overinvestment in ribosome synthesis under less favorable nutritional conditions. In mammals, growth-dependent regulation of RNA polymerase I (Pol I) transcription is mediated by TIF-IA, an essential initiation factor that is active in extracts from growing but not starved or cycloheximide-treated mammalian cells. Here we report the molecular cloning and functional characterization of recombinant TIF IA, which turns out to be the mammalian homolog of the yeast factor Rrn3p. We demonstrate that TIF-IA interacts with Pol I in the absence of template DNA, augments Pol I transcription in vivo and rescues transcription in extracts from growth-arrested cells in vitro. PMID- 11265759 TI - Regulation of intracellular dynamics of Smad4 by its leucine-rich nuclear export signal. AB - Smad family proteins play a pivotal role in transmitting the transforming growth factor-beta (TGF-beta) superfamily signals from the cell surface to the nucleus. In response to ligand stimulation, Smad4 forms a complex with respective receptor specific Smads, and the complex translocates into the nucleus and regulates gene expression. Thus, the nuclear entry of the Smad complex is one of the key steps in signal transduction. However, little is known about regulatory mechanisms for nucleocytoplasmic transport of Smads. Here we report identification of a functional, leucine-rich nuclear export signal (NES) in Smad4, which regulates subcellular distribution of Smad4. We then show evidence suggesting that the NES dependent cytoplasmic localization of Smad4 is important for ensuring optimal TGF beta responsivenesses in transcriptional activation. Moreover, we show that the NES of Smad4 is specifically inactivated by the stimulus-dependent hetero oligomerization with receptor-specific Smads during the TGF-beta-induced nuclear translocation of Smad4. Taken together, these results suggest an important regulatory role of the NES of Smad4 in TGF-beta signaling. PMID- 11265762 TI - Genocentric promises. PMID- 11265760 TI - The 3.7 A projection map of the glycerol facilitator GlpF: a variant of the aquaporin tetramer. AB - GlpF, the glycerol facilitator protein of Escherichia coli, is an archetypal member of the aquaporin superfamily. To assess its structure, recombinant histidine-tagged protein was overexpressed, solubilized in octylglucoside and purified to homogeneity. Negative stain electron microscopy of solubilized GlpF protein revealed a tetrameric structure of approximately 80 A side length. Scanning transmission electron microscopy yielded a mass of 170 kDa, corroborating the tetrameric nature of GlpF. Reconstitution of GlpF in the presence of lipids produced highly ordered two-dimensional crystals, which diffracted electrons to 3.6 A resolution. Cryoelectron microscopy provided a 3.7 A projection map exhibiting a unit cell comprised of two tetramers. In projection, GlpF is similar to AQP1, the erythrocyte water channel. However, the major density minimum within each monomer is distinctly larger in GlpF than in AQP1. PMID- 11265763 TI - Something for everyone. Horizontal gene transfer in evolution. PMID- 11265761 TI - Membrane raft microdomains mediate lateral assemblies required for HIV-1 infection. AB - HIV-1 infection triggers lateral membrane diffusion following interaction of the viral envelope with cell surface receptors. We show that these membrane changes are necessary for infection, as initial gp120-CD4 engagement leads to redistribution and clustering of membrane microdomains, enabling subsequent interaction of this complex with HIV-1 co-receptors. Disruption of cell membrane rafts by cholesterol depletion before viral exposure inhibits entry by both X4 and R5 strains of HIV-1, although viral replication in infected cells is unaffected by this treatment. This inhibitory effect is fully reversed by cholesterol replenishment of the cell membrane. These results indicate a general mechanism for HIV-1 envelope glycoprotein-mediated fusion by reorganization of membrane microdomains in the target cell, and offer new strategies for preventing HIV-1 infection. PMID- 11265764 TI - A European research identity. A talk with Philippe Busquin, Commissioner for Research of the European Commission. Interview by Holger Breithaupt, Russ Hodge and Frank Gannon. PMID- 11265765 TI - Graft-versus-host alloresponses to treat nonlymphohematopoietic tumors: is there a solid approach? PMID- 11265766 TI - Issues on clinical applications of cancer vaccines. PMID- 11265767 TI - Antitumor effect of immunizations with fusions of dendritic and glioma cells in a mouse brain tumor model. AB - The authors studied antitumor immunity conferred by fusions of dendritic and glioma cells in a mouse brain tumor model. Previous immunization with fusion cells (FCs) prevented tumor formation on challenge with glioma cells in the flank or in the brain. Efficacy was decreased when studies were performed in mice depleted of CD8+ cells. In a treatment model, FCs were injected subcutaneously after tumor development in the brain. The administration of FCs alone had limited effects on survival of mice bearing brain tumors. Importantly, however, administration of FCs and recombinant interleukin-12 (rIL-12) remarkably prolonged the survival of mice with brain tumors. Cytotoxic T lymphocyte activity against glioma cells from immunized mice was also stimulated by coadministration of FCs and rIL-12 compared with that obtained with FCs or rIL-12 alone. These data support the therapeutic efficacy of combining FC-based vaccine therapy and rIL-12. PMID- 11265768 TI - Cell therapy with preimmunized effector cells mismatched for minor histocompatible antigens in the treatment of a murine mammary carcinoma. AB - Cell therapy with allogeneic donor cells mismatched for minor histocompatible (MiHC) antigens was applied to a murine mammary carcinoma (4T1) model to test the feasibility of graft versus tumor (GVT) effect against metastatic epithelial tumor cells. BALB/c mice bearing a 4T1 tumor of BALB/c origin were given syngeneic or MiHC-mismatched splenocytes. GVT effects were determined in secondary recipients of adoptively transferred lung cells derived from primary hosts who had previously been inoculated intravenously with 4T1 cells, and injected with one of the following: 1) naive BALB/c splenocytes, 2) naive DBA/2 splenocytes, 3) 4T1-immune DBA/2 splenocytes, or 4) DBA/2 splenocytes immunized with host-derived BABL/c spleen cells. Naive DBA/2 splenocytes inhibited tumor growth only slightly and only slightly prolonged the survival of secondary recipients, in comparison with fully matched tumor/host BALB/c spleen cells. An efficient GVT reaction was demonstrated in vitro and in vivo with MiHC-mismatched DBA/2 splenocytes from mice presensitized by multiple injections of irradiated tumor or BALB/c-derived spleen cells. All 30 mice adoptively inoculated with lung cells from primary hosts that had previously been treated with these presensitized effector cells were tumor free for >250 days. Secondary recipients inoculated with lung cells from mice given naive BALB/c or DBA/2 spleen cells died of metastatic tumors within 33 to 46 days. These results suggest that preimmunized donor cells represent an effective tool against metastatic disease; hence, the next goal should be to control graft-versus-host disease while exploiting the GVT potential. PMID- 11265769 TI - A rapid, novel strategy to induce tumor cell-specific cytotoxic T lymphocyte responses using instant dentritomas. AB - The generation of fused cells between dendritic cells (DC) and tumor cells is a very effective approach for tumor antigen presentation in cancer immunotherapy. However, the application of this approach in clinical studies is limited by the need for established tumor cell lines and the time-consuming procedures for selecting and expanding the fused cells. In the current study, the authors report a rapid, novel approach to produce fused cells between DCs and primary tumor cells from patients with malignant melanoma. Peripheral blood DCs and a primary tumor cell culture were generated from the same patients, labeled with fluorescent green and red dyes, respectively, and fused. The fused cells were isolated by fluorescence-activated cell sorting. Because the fused cells do not need to be expanded, these cell hybrids have been named instant dendritomas. Fluorescence-activated cell sorting analysis showed that instant dendritomas express the key molecules for antigen presentation (HLA-A, B, C; HLA-DR; CD80; and CD86). In vitro studies have shown that instant dendritomas effectively activated autologous CD8+ T lymphocytes to proliferate and secret interferon gamma. More importantly, the activated CD8+ T lymphocytes effectively lysed the patients' primary tumor cells. This approach represents a practical clinical strategy for cancer immunotherapy. PMID- 11265770 TI - Dendritic cell infiltration in colon cancer. AB - We quantitatively evaluated dendritic cell (DC) infiltration in primary colorectal cancers from 44 patients and metastatic colorectal tumors from 13 patients using immunohistochemistry for the DC marker CD83, HLA-DR, and the DC activation molecules CD40 and CD86. Nearly all CD83+ cells were also HLA-DR+, CD40+, and CD86+, indicating that the DCs that infiltrate colon cancer in vivo express the activation and costimulatory molecules associated with a mature DC phenotype. The density of DCs in colorectal cancer primaries was three times lower than that seen in normal colonic mucosa (0.29 versus 0.84 CD83+ cells/ high power field (hpf), p < 0.001). Dendritic cells were rarely observed in metastatic tumors: DC density in metastases was sixfold lower than in colorectal primary tumors (0.05 versus 0.29 CD83+ cells/hpf, p < 0.001). Because cytokines have been shown, in vitro, to exert potent effects on DCs, we also evaluated the relationship between intratumor DC density and the expression of cytokines by tumor-infiltrating lymphocytes (TILs) and tumor cells. Expression of interleukin 10 and transforming growth factor beta by either TIL or tumor cells was not associated with decreased DC density or decreased expression of CD40 or CD86 on DCs. Tumor expression of vascular endothelial growth factor was associated with a more than twofold increase in DC density (p = 0.01). Patients who had a high proportion of TILs expressing tumor necrosis factor (TNF) had a greater intratumor mature DC density than patients with a low proportion of TNF + TIL (0.54 versus 0.21 CD83+ cells/hpf, p < 0.01). PMID- 11265771 TI - Expression of the chemokines IP-10 and Mig in IL-10 transduced tumors. AB - Several laboratories have reported marked tumor inhibition when the cytokine interleukin-10 (IL-10) is overexpressed as a transgene in a variety of tumor cells. To identify critical effector molecules, we compared the expression of the chemokine crg-2, the murine homolog of human inducible protein 10 (human IP-10) in murine mammary tumors derived from the transplantation of six IL-10 expressing clones of tumor cell line 66.1, parental 66.1, or 66-neo-cells. We observed increased levels of IP-10 mRNA in all IL-10-expressing tumors examined in comparison to 66-neo. IP-10 mRNA was not detected in parental 66.1 tumors. The closely related chemokine Mig (monokine induced by interferon-gamma [IFN-gamma]) was also induced in all IL-10-expressing tumors. Studies of cultured tumor cells in vitro show that mammary epithelial tumor cells, in the absence of host elements, can express IP-10 and Mig in response to induction with either lipopolysaccharide (LPS) or IFN-gamma alone. The combination of LPS plus IFN gamma resulted in even greater induction of IP-10 RNA. The kinetics of induction differ somewhat for the two chemokines, with IP-10 showing slower induction and less rapid decline. Because both Mig and IP-10 are chemotactic for tumor infiltrating lymphocytes, we examined the presence of CD4+ and CD8+ lymphocytes in these tumors. Consistent with the upregulation of Mig and IP-10, we saw significantly increased numbers of CD8+ cells and a lesser increase in CD4+ cells in tumors with elevated levels of both chemokines. PMID- 11265772 TI - Cytotoxicity of antiosteosarcoma recombinant immunotoxins composed of TP-3 Fv fragments and a truncated Pseudomonas exotoxin A. AB - Regrowth of drug-resistant tumor cells is responsible for approximately half of an unselected osteosarcoma population still dying of the disease despite aggressive combination therapy. Two monoclonal antibodies, TP-1 (immunoglobulin 2a) and TP-3 (immunoglobulin 2b) are available, which specifically recognize an antigen on osteosarcoma cells. In this work, we have fused the variable (V) genes of TP-3 to a truncated fragment of Pseudomonas exotoxin A, referred to as PE38. Two immunotoxins were made that differed in the Fv portion: TP-3(scFv)-PE38, which contains a peptide linker, and TP-3(dsFv)-PE38, which contains a disulfide bond for stabilization of the association between the V domains. Recombinant TP-3 immunotoxins were expressed in Escherichia coli and purified from inclusion bodies. We describe the design and expression of these immunotoxins, and their properties with regard to antigen binding, stability, and cytotoxicity. Toxicity studies were done in mice. We found that the immunotoxins exhibited very similar in vitro properties, whereas in vivo TP-3(dsFv)-PE38 was much better tolerated than TP-3(scFv)-PE38. PMID- 11265773 TI - Sequential 5-Aza-2 deoxycytidine-depsipeptide FR901228 treatment induces apoptosis preferentially in cancer cells and facilitates their recognition by cytolytic T lymphocytes specific for NY-ESO-1. AB - Global alterations in chromatin structure profoundly influence gene expression in thoracic neoplasms, silencing tumor suppressors while facilitating the expression of various cancer testis antigens such as NY-ESO-1. Although recent studies have shown that histone deacetylase inhibitors can potentiate tumor suppressor gene induction mediated by demethylating agents in cancer cells, the ability of these agents to augment cancer testis antigen expression have not been fully defined. The authors designed the current study to determine whether the histone deacetylase inhibitor, depsipeptide FR901228 (DP), could enhance NY-ESO-1 induction mediated by the DNA demethylating agent 5-Aza-2'-deoxycytidine (DAC) in cell lines established primarily from thoracic cancers. Quantitative reverse transcriptase polymerase chain reaction analysis revealed that, under exposure conditions potentially achievable in clinical settings, DAC dramatically induced NY-ESO-1 expression in cultured cancer lines. DP alone mediated negligible target gene induction but significantly augmented DAC-mediated induction of NY-ESO-1. After DAC or sequential DAC-DP treatment, HLA-A*0201 cancer cells were recognized by an HLA-A*0201 CTL specific for NY-ESO-1. Although sequential DAC/DP exposure did not uniformly enhance immune recognition of target cells compared with DAC alone, this treatment mediated profound induction of apoptosis in cancer cells but not normal human bronchial epithelia. The apoptotic effects of DAC, DP, or sequential DAC-DP did not correlate in an obvious manner with histology, or the magnitude of NY-ESO-1 induction in cancer cells. Although the mechanisms have not been fully defined, sequential DAC-DP treatment may be a novel strategy to augment antitumor immunity in cancer patients. PMID- 11265774 TI - Proinflammatory cytokines and CD40 ligand enhance cross-presentation and cross priming capability of human dendritic cells internalizing apoptotic cancer cells. AB - Human monocyte-derived dendritic cells (DC) can ingest apoptotic tumor cells (ATC) and present tumor-associated antigens (TAA) to T cells, leading to the generation of tumor-specific cytotoxic effector cells (Cancer Res 2000;60:3542 9). To further augment antitumor effector cell responses, attempts were made to modify antigen presentation and cross-priming of T cells by DC fed with ATC. Proinflammatory cytokines (PC), CD40 ligand (CD40L) and/or interferon-gamma (IFN gamma) were found to markedly enhance the immunogenicity of TAA presented by DC. While PC upregulated expression of major histocompatibility complex class I/II and costimulatory molecules on the surface of DC, CD40L +/- IFN-gamma increased interleukin (IL)- 12 and to a lesser extent, IL-15 production by DC. Additionally, lactacystin, a specific proteasome inhibitor, significantly abrogated the effects of IFN-gamma and, in part, also those of CD40L or PC. The ability of DC + ATC to cross-prime TAA-inexperienced ("naive") T cells was significantly enhanced by PC and CD40L or CD40L + IFN-gamma, but not by IFN-gamma alone. These results indicate that future vaccines for patients with cancer incorporating DC fed with ATC could be made more effective by the addition of proinflammatory cytokines or CD40L +/- IFN-gamma to improve the DC function. PMID- 11265776 TI - Neoadiuvant treatment with intravesical interleukin-2 for recurrent superficial transitional bladder carcinoma Ta-T1/G1-2. AB - The aim of this study was to evaluate the direct action of IL-2 on recurrent superficial transitional bladder carcinoma and the effect on recurrence rate. 27 patients were submitted to neoadjuvant treatment by intra-vesical instillation of recombinant IL-2 and to transurethral resection. We did not observe any effect on neoplasms but the recurrence rate was less than the expected one. It is possible that treatment of bladder carcinoma with intra-vesical instillation of IL-2 may promote immuno-prophilaxis. PMID- 11265775 TI - Mannan mucin-1 peptide immunization: influence of cyclophosphamide and the route of injection. AB - The mucin MUC1 is greatly increased in breast cancer and is a potential target for immunotherapy. In mice, MUCI conjugated to oxidized mannan (MUC1-mannan fusion protein [M-FP]) targets the mannose receptor and induces a high frequency of cytotoxic T lymphocytes and anti-tumor responses. On this basis, three phase I trials were performed in patients with adenocarcinoma to evaluate the toxicity and the immunologic responses to mannan MUCI. Forty-one patients with metastatic or locally advanced carcinoma of the breast (trial 1), colon (trial 2), and various adenocarcinomas (trial 3) received increasing doses of M-FP (1 to 300 microg). The immunizations were given at weekly intervals (weeks 1 to 3) and repeated in weeks 7 to 9. Cyclophosphamide (to increase cellular immunity) was given on weeks 1 and 4. M-FP was given intramuscularly in trial 1 and intraperitoneally in trial 2. No toxic effects occurred, and delayed-type hypersensitivity responses were present only as a microscopic lymphocytic infiltration. Overall, approximately 60% of the patients had high-titer MUC1 immunoglobulin G1 antibody responses, with the intraperitoneal route yielding approximately 10-fold higher responses. Cellular responses (proliferation, cytotoxic T cells, or CD8 T cells secreting tumor necrosis factor-alpha alphand interferon-gamma in response to MUC1 stimulation in vitro) were found in 28% of the patients, which was similar to that seen without cyclophosphamide. In most patients, disease progressed, but in five it remained stable. In addition, there were no objective responses. M-FP is not toxic and induces immune responses that were amplified by the intraperitoneal route of immunization. Cyclophosphamide was of no benefit. PMID- 11265777 TI - A prospective, randomized, double-blind, placebo-controlled trial evaluating the effect of nystatin on the development of oral irritation in patients receiving high-dose intravenous interleukin-2. AB - Interleukin-2 (IL-2) has been used to treat patients with metastatic melanoma and renal cell cancer for nearly two decades, and much progress has been made in ameliorating its adverse effects. One bothersome adverse effect, oral pain or oral irritation, is usually treated with an oral antifungal antibiotic, nystatin. The authors performed a prospective, randomized, double-blind, placebo-controlled trial involving 64 patients to evaluate the effect of prophylactic administration of nystatin or placebo on the development of oral irritation in patients receiving high-dose intravenous IL-2. No difference was found between patients randomized to receive nystatin or placebo in their rates of development of oral irritation, the severity of IL-2 adverse effects, the duration of their treatment, the rate of development of positive studies for oral yeast, or their pattern of experiencing other adverse effects. Thus, patients who receive high dose intravenous IL-2 should not be treated prophylactically with nystatin to prevent oral irritation, and clinicians should seek evidence of the presence of oral thrush before using antifungal agents to treat oral pain in these patients. PMID- 11265778 TI - Tumor-dendritic cell fusion technology and immunotherapy strategies. PMID- 11265779 TI - Chemically modified papain for applications in detergent formulations. AB - Papain was modified using succinic anhydride, and the modified papain so obtained was compared with the native papain for its activity and stability in detergents. This study was done using commercial enzyme detergents as references. It was found that modified papain retained activity comparable to the commercial enzyme detergents. Chemically modified papain may prove to be an inexpensive alternative to alkaline proteases that are used in detergents. PMID- 11265780 TI - Composted grape marc as growing medium for hypostases (Hypostases phyllostagya). AB - The use of composted grape marc (CGM) as a plant growth medium was investigated with Hypostases (Hypostases phyllostagya). Seven media were prepared using CGM mixed, in different ratios, with native peat and perlite. The following mixtures were used: 100% CGM, 75% CGM + 25% peat, 50% CGM + 50% perlite, 25% CGM + 75% peat, 50% CGM + 25%) peat + 25% perlite, 25%, CGM + 50%, peat + 25% perlite and 100% peat. The experiment was arranged in a randomized plot design with four replicates under greenhouse conditions. After a growing period of three months, some horticultural parameters were measured. Besides, some physical and chemical properties of the growing medium were determined. The mixtures of 50% CGM + 50% peat, 25% CGM + 75% peat and 100% peat were found to be most suitable based on the horticultural parameters. This was confirmed through the physical characteristics. Up to 50% composted grape marc can be used in mixtures with peat on account of its low cost and high nutrient content. PMID- 11265781 TI - Co-composting of chestnut burr and leaf litter with solid poultry manure. AB - A co-composting of chestnut burr and leaf litter mixed with solid poultry manure was assessed by comparison of several chemical, physicochemical and biological parameters. The final pH of the co-compost was 8.89 and the C/N ratio was 13. The germination index (GI) obtained using the co-compost varied with the seeds used. It was 155.35% for ryegrass seeds, 56.56% for wheat seeds and 100% for barley seeds. The co-compost was mature in 103 days from a biological point of view. PMID- 11265782 TI - Pig manure vermicompost as a component of a horticultural bedding plant medium: effects on physicochemical properties and plant growth. AB - This experiment was designed to characterize the physical, chemical and microbial properties of a standard commercial horticultural, greenhouse container, bedding plant medium (Metro-Mix 360), that had been substituted with a range of increasing concentrations (0%, 5%, 10%, 25%, 50% and 100% by volume) of pig manure vermicompost and to relate these properties to plant growth responses. The growth trials used tomatoes (Lycopersicon esculentum Mill.), grown in the substituted media for 31 days under glasshouse conditions, with seedling growth recorded in 20 pots for each treatment. Half of the tomato seedlings (10 pots per treatment) were watered daily with liquid inorganic fertilizer while the other half received water only. The percentage total porosity, percentage air space, pH and ammonium concentrations of the container medium all decreased significantly, after substitution of Metro-Mix 360 with equivalent amounts of pig manure vermicompost; whereas bulk density, container capacity, electrical conductivity, overall microbial activity and nitrate concentrations, all increased with increasing substitutions of vermicompost. The growth of tomato seedlings in the potting mixtures containing 100% pig manure vermicompost was reduced, possibly as a result of high soluble salt concentrations in the vermicompost and poorer porosity and aeration. The growth of tomato seedlings was greatest after substitution of Metro-Mix 360 with between 25% and 50% pig manure vermicompost, with more growth occurring in combinations of pig manure vermicompost treated regularly with a liquid fertilizer solution than in those with no fertilizer applied. Some of the growth enhancement in these mixtures seemed to be related to the combined effects of improved porosity, aeration and water retention in the medium and the high nitrate content of the substrate, which produced an increased uptake of nitrogen by the plant tissues, resulting in increased plant growth. When the tomato seedlings were watered daily with liquid inorganic fertilizer, substitution of Metro-Mix 360 with a very small amount (5%) of pig manure vermicompost resulted in a significant increase in the growth of tomato seedlings. Such effects could not be attributed solely to the nutritional or physical properties of the pig manure vermicompost. Therefore, it seems likely that the pig manure vermicompost provided other biological inputs, such as plant growth regulators into the container medium, that still need to be identified fully. PMID- 11265784 TI - Growth substrates made from duck excreta enriched wood shavings and source separated municipal solid waste compost and separates: physical and chemical characteristics. AB - Production and use of compost is an effective means to reduce wastes, and offers a large potential as growth substrates and source of nutrients. The objective of this study was to determine the physical and chemical characteristics of duck excreta enriched wood shavings (DMC) and source-separated municipal solid waste (MSW) composts and separates, and to assess the physical characteristics of growth substrates made from these two composts and selected substrates. MSW compost separates were the following sizes: F1 > 4 mm diameter, 2 mm < F2 < 4 mm, 1 mm < F3 < 2 mm and F4 < 1 mm. Growth substrates were prepared by mixing DMC and F2 and F3 MSW separates (M/M ratios). Growth substrates A-E consisted exclusively of 10-60% DMC and 20-60% of MSW separates F2 and F3. Growth substrates F-J, and K O were the same as substrates A E, with 15% M/M brick fragments or shredded plastic added as porosity agents, respectively. Growth substrates (BE/S) made of black earth (BE) and sandy loam soil (Ls) in a 1:4 (M/M) ratio, commercially available peat substrate (Pr) and an in-house sphagnum peat-based substrate (Gs) were used for comparison. Principal component analysis (PCA) showed that DMC was a better material than MSW with respect to porosity and water field capacity. MSW compost and separates differed by their relatively high levels of water-soluble and HCl-hydrolyzable N and increased advantageous water retention capacity. PCA also showed that substrates A-E exhibited porosity and water field capacity similar to those of Pr. Substrates F-J had porosity and water field capacity similar to those of BE/S, whereas substrates K O were more similar to Pr and to substrates A and B. The presented data indicate that DMC and MSW separates were complementary in providing good physical and chemical characteristics to the growth substrates. PMID- 11265783 TI - Decolourisation of molasses wastewater by cells of Pseudomonas fluorescens immobilised on porous cellulose carrier. AB - Pseudomonas fluorescens isolated from soil samples contaminated with molasses, decolourised molasses wastewater (MWW) samples up to 76% under non-sterile conditions in four days at 30 degrees C. Immobilised cells could be reused for decolourisation activity. However, in subsequent cycles, this was found to decrease from 76% to 50% and from 50% to 24%. Decolourisation activity was regenerated from 30% to 45% by recultivating the immobilised cells in a fresh growth medium. Cellulose carrier coated with collagen was found to be most effective carrier, which produced the highest decolourisation activity of 94% in a 4-day process. This carrier could be reused with 50% of the decolourisation activity retained until the seventh day. PMID- 11265785 TI - Chromate tolerant bacteria isolated from tannery effluent. AB - The occurrence of metal tolerant and antibiotic resistant organisms was investigated in tannery effluent. Seventy-seven isolates comprising heterotrophs (41) and coliforms (36) which were tolerant to chromate level of > 50 microg/ml were selected for detailed study. The majority of the coliforms were resistant to higher levels of chromate (200 microg/ml) whereas around 3% of the heterotrophs were resistant to Cr6+ at a level of > 150 microg/ml. All chromate tolerant heterotrophs were also tolerant to Cu2+ (100%) whereas only 58.53% coliforms were tolerant to Cu2+. Except in the case of Cd2+ a higher number of heterotrophs were found tolerant to other heavy metals tested. Both groups of isolates were found sensitive to mercury. Resistance to cephaloridine was more abundant (P < 0.001) in coliforms as compared to heterotrophs. On the other hand a significantly higher number (P < 0.01) of heterotrophs showed resistance to streptomycin and carbencillin. All coliforms were sensitive to chloramphenicol. Around 80%) and 31.70% of coliforms and heterotrophs exhibited a relationship to the combination of metals and antibiotics. Both heterotrophs and coliforms tolerant to Hg2+ were also resistant to polymixin-B. PMID- 11265786 TI - Three-dimensional outgrowth of a wood-rotting fungus added to a contaminated soil from a former gasworks site. AB - The capability of wood-rotting fungi (WRF) to colonise contaminated soil is an important fungal characteristic in the development of WRF-based soil bioremediation, it is also important to have methods that monitor the presence of the WRF in the soil. In this lab-scale study, it was shown that it was possible to re-capture, localise and identify a brown-rot fungus, Antrodia vaillantii, after it has been inoculated into, and grown in, a contaminated soil from a former gasworks site. The three-dimensional outgrowth of A. vaillantii was monitored by allowing it to grow into fungicide-treated wood baits, temporarily placed in the soil. After two weeks, the baits were withdrawn from the soil and surface sterilised with hydrogen peroxide to favour fungi growing inside baits, i.e., A. vaillantii. After subsequent plating of baits on selective agar medium the presence of A. vaillantii was confirmed with PCR/RFLP. A. vaillantii was found to be viable throughout the 54 days long study and exhibited a surface growth pattern similar to other well-known cord-forming basidiomycetes. Firstly, the upper part of the soil closest to the place of inoculation was colonised, however, over a period of time, the area of colonisation spread deeper into the soil. The detection method employed in the current study gave a conservative estimate of the fungal proliferation and did not require extensive sampling. Its use could be applicable in both applied research, such as soil bioremediation, and in pure microbial ecology studies. PMID- 11265787 TI - Detection of catabolic genes in indigenous microbial consortia isolated from a diesel-contaminated soil. AB - Bioremediation is often used for in situ remediation of petroleum-contaminated sites. The primary focus of this study was on understanding the indigenous microbial community which can survive in contaminated environment and is responsible for the degradation. Diesel. toluene and naphthalene-degrading microbial consortia were isolated from diesel-contaminated soil by growing on selective hydrocarbon substrates. The presence and frequency of the catabolic genes responsible for aromatic hydrocarbon biodegradation (xylE, ndoB) within the isolated consortia were screened using polymerase chain reaction PCR and DNA DNA colony hybridization. The diesel DNA-extract possessed both the xy/E catabolic gene for toluene, and the nah catabolic gene for polynuclear aromatic hydrocarbon degradation. The toluene DNA-extract possessed only the xylE catabolic gene, while the naphthalene DNA-extract only the ndoB gene. Restriction enzyme analysis with HaeIII indicated similar restriction patterns for the xylE gene fragment between toluene DNA-extract and a type strain, Pseudomonas putida ATCC 23973. A substantial proportion (74%) of the colonies from the diesel-consortium possessed the xylE gene, and the ndoB gene (78%), while a minority (29%) of the toluene consortium harbored the xylE gene. 59% of the colonies from the naphthalene consortium had the ndoB gene, and did not have the xylE gene. These results indicate that the microbial population has been naturally enriched in organisms carrying genes for aromatic hydrocarbon degradation and that significant aromatic biodegradative potential exists at the site. Characterization of the population genotype constitutes a molecular diagnosis which permits the determination of the catabolic potential of the site to degrade the contaminant present. PMID- 11265788 TI - Yeast production from virgin grape marc. AB - An alternative utilization of virgin grape marc (VGM) to produce SCP from S. ceretisiae is reported. A simple extraction method of fresh grape marc produces a sugar-rich solution: through fed-batch fermentation, a high-value yeast biomass instead of a low-value product like ethanol can be produced. Productivity and quality of yeast are similar to these obtainable from molasses. The convenience of yeast production from VGM is briefly discussed; it appears of great interest in south Italy and generally in grape-producing countries, specially if these lack relevant sources of fermentable sugars. PMID- 11265789 TI - Studies on the suitability of alginate-entrapped Chlamydomonas reinhardtii cells for sustaining nitrate consumption processes. AB - Some aspects of the suitability of alginate beads entrapping Chlamydomonas reinhardtii cells for nitrate consumption from nitrate-containing waters were studied and discussed. Among 14 different metal cations tested as gel bead stabilizing agents, only calcium and barium formed beads showing nitrate consuming activity. Pure calcium alginate cell entrapment resulted in the most suitable method for active cell immobilization compared to alginate-composite-gel beads based on poly-vinylcaprolactam (PVCL) and poly-vinylpyrrolidone (PVP). To perform a continuous nitrate consumption process, calcium alginate-entrapped cells were first grown in a 2.5 l airlift-loop reactor. A cell loading of about 150 microg Chl. g(-1) gel was achieved. Afterwards, five days nitrate consumption processes were performed and three different dilution rates were applied: (i) D < mu; (ii) D = mu; (iii) D > mu, where mu is the specific growth rate (h(-1)). The maximum consumption rates calculated for each dilution rate were: (i) 3.8, (ii) 6.4 and (iii) 7.2 mg nitrate mg(-1) Chl. h(-1). For low dilution rates (D < mu) some nitrite (< 300 microM) was excreted into the culture medium. However, this concentration of nitrite was not high enough to inhibit nitrate consumption. PMID- 11265790 TI - Influence of process variables in the ethanol pulping of olive tree trimmings. AB - A central composition design was developed to study the influence of process variables (temperature, pulping time and ethanol concentration) on the properties of the pulp produced (yield and holocellulose, alpha-cellulose and lignin contents) and the pH of the resulting wastewater, in the ethanol pulping of olive tree trimmings. The proposed equations reproduce the experimental results for the dependent variables with errors less than 5% for the holocellulose and alpha cellulose contents, yield and wastewater pH, and less than 15% for the lignin content. Obtaining pulp with acceptably high yield (37.6%), high holocellulose and alpha-cellulose contents (above 88.8% and 46.9%, respectively), and low lignin contents (below 7.2%), entails operating at a pulping temperature of 200 degrees C, using an ethanol concentration of 75% and a pulping time of 60 min. PMID- 11265791 TI - Oxidation of lignin in eucalyptus kraft pulp by manganese peroxidase from Bjerkandera sp. strain BOS55. AB - The white rot fungus Bjerkandera sp. strain BOS55 was shown in previous studies to cause high levels of kraft pulp bleaching and delignification under culture conditions in which manganese peroxidase (MnP) occurs as the dominant oxidative enzyme. In this study, the MnP of Bjerkadera was isolated and tested in vitro with eucalyptus oxygen-delignified kraft pulp (ODKP) based on measuring the reduction in kappa number as an indicator of lignin oxidation. The MnP preparation applied at 60 U/g pulp for 6 h caused a significant decrease of 11 13% in the kappa number in the ODKP under optimal conditions compared to parallel incubated controls lacking enzyme. The effects of MnP dosage, Mn2+ concentration, organic acid buffer selection, pH and H2O2 addition were evaluated. The optimal Mn2+ concentration range for lignin oxidation in ODKP was 100-500 microM. In the presence of low oxalate concentrations (0.3-2 mM) the Bjerkandera MnP also significantly reduced the kappa number of ODKP by 6% without any Mn. This observation is in agreement with the fact that purified Bjerkandera MnP has Mn independent activities. Under incubation conditions with added Mn2+, buffers composed of metal-complexing organic acids provided two-fold better kappa number reductions compared to the inert acetic acid. The optimal H2O2 dosage was found to be 0.017 micromol/min ml when added as semi-continuous pulses (every 30 min) or 0.2 micromol/min ml when generated continuously by glucose oxidase. Excess H2O2 caused severe inactivation of MnP during the incubations. Factors that improved the turnover of the enzyme, such as Mn2+ and metal-chelating acids, stabilized MnP against rapid inactivation. PMID- 11265792 TI - Anaerobically digested poultry slaughterhouse wastes as fertiliser in agriculture. AB - Chemical and physical analysis, 27-d plant growth assays with carrot (Daucus carota) and Chinese cabbage (Brassica campestris var. chinensis), and 5-d phytotoxicity assays with Chinese cabbage and perennial ryegrass (Lolium perenne) were used to investigate the suitability of anaerobically digested poultry slaughterhouse waste for fertiliser in agriculture and the effect of aerobic post treatment on the properties of the digested material. The digested material appeared to be rich in nitrogen. In 27-d assays with digested material as nitrogen source, carrots grew almost as well as those fertilised with a commercial mineral fertiliser used as reference, whereas, the growth of Chinese cabbage was inhibited. In further 5-d phytotoxicity assays, the digested material inhibited the germination and root growth of ryegrass and Chinese cabbage, apparently because of organic acids present in it. Aerobic post-treatment of the material reduced its phytotoxicity but, probably due to the volatilisation of ammonia, resulted in loss of nitrogen. PMID- 11265793 TI - Degumming of ramie fibers by alkalophilic bacteria and their polysaccharide degrading enzymes. AB - Three strains of alkalophilic bacteria, Bacillus sp. NT-39, NT-53 and NT-76, were selected for the degumming of ramie fibers and production of polysaccharide degrading enzymes. After 48 h of incubation with the strains, the loss of the gum might amount to 5.0% or more of the fibers and a number of polysaccharide degrading enzymes were secreted to the culture supernatants. The residual gum of the fibers decreased to 9.4% after 5 h of enzymatic degumming. Analysis of gum contents and enzyme activities revealed that pectate lyase and xylanase played an important role in the degradation of residual gum. Enzymatic degumming resulted in an increment of 5.4 ISO units in fiber brightness, whereas the reduction in bundle breaking tenacity of the fibers was less than 5.%. The results confirmed that degumming of ramie fibers by alkalophilic bacteria and their enzymes had substantial advantages. PMID- 11265794 TI - Decolourisation of synthetic and spentwash melanoidins using the white-rot fungus Phanerochaete chrysosporium JAG-40. AB - Phanerochaete chrysosporium JAG-40 was isolated from the soil samples saturated with spilled molasses collected from a sugar mill. This isolate decolourised synthetic and natural melanoidins present in spentwash in liquid fermentation; up to 80% in 6 days at 30 degrees C under aerobic conditions. A large inoculum size stimulated fungal biomass production, but this gave less decolourisation of pigment; 5% w/v (dry weight) mycelial suspension was found optimum for maximum decolourisation in melanoidin medium supplemented with glucose and peptone. Gel filtration chromatography showed that larger molecular weight fractions of melanoidin were decolourised more rapidly than small molecular weight fractions. PMID- 11265795 TI - Reaction coefficient (K) evaluation for full-scale facultative pond systems. AB - Facultative ponds have found application in wastewater treatment as an economical system where geographical locations are available at reasonable cost. Several design methods have been reported to describe the organic matter removal of a facultative pond and to determine the reaction coefficient (K) in general terms. Therefore, it is important to evaluate if these coefficient values can be used satisfactorily in regional cases. For this purpose the data of two full-scale facultative ponds located in Brazlandia and Samambaia, in the mid-west region of Brazil, were used. A correlation between applied COD load and reaction coefficient (K) was obtained based on a mathematical adjustment using the dispersed flow hydraulic model. The results provide a suggested regional design parameter for facultative ponds in this region in terms of domestic wastewater. PMID- 11265796 TI - Changing scenario of coronary artery disease. PMID- 11265797 TI - Radiofrequency ablation: a cure for tachyarrhythmias. AB - Radiofrequency (RF) ablation is a new modality of pennanently curing patients with various tachycardias using radiofrequency energy, a technique evolved in the past decade. RF ablation was performed on 913 patients with different tachyarrhythmias from April, 1994 to July, 1999. There were 491 men and 422 females aged 42 +/- 34 years (range 1 to 76 years). Supraventricular tachycardia (SVT) was present in 462 patients, accessory pathway mediated atrioventricular re entrant tachycardia (AVRT) in 355 patients (377 accessory pathways) and idiopathic ventricular tachycardia (VT) in 96 patients. Amongst the patients with SVT, 402 had atrioventricular nodal re-entrant tachycardia (AVNRT), 22 had atrial flutter, 20 had ectopic atrial tachycardia and 18 had atrial fibrillation. RF successfully abolished the tachycardia in 400/402 patients (99.5%) with AVNRT, 330/377 (87.5%) accessory pathways in patients with AVRT, 14/22 patients (63.6%) of atrial flutter, 18/20 patients (90%) of atrial tachycardia and 79/96 patients (82.3%) with idiopathicVT. Successful AV nodal ablation with pacemaker implantation was done in 10/18 patients with chronic atrial fibrillation with fast ventricular rate and tachycardia induced cardiomyopathy. AV nodal modulation for atrial fibrillation was tried in the remaining 8 patients and was successful in 4 (4/8). The overall success rate for all arrhythmias was 93.6%, and there was no mortality. At a follow-up of 6.8 +/- 5.4 months, there was a recurrence in 34/420 patients (8%), in whom successful re-ablation was performed. One patient with AVNRT and another with a parahisian pathway developed complete heart block and were given pacemakers. One patient developed inferior wall infarction on the next day post RF. There were 4 patients who had pericardial tamponade necessitating pericardiocentesis and 2 patients developed deep vein thrombosis, which was treated conservatively. Thus RF ablation is an effective, safe and curative therapy for various arrhythmias. PMID- 11265798 TI - Myocardial bridges. AB - Recently, many cases of sudden death on strenuous exercise attributed to muscular bridges on the coronary arteries have been reported in the literature. The incidence of such bridges, pooling all reports, is 5.4-85.7% of autopsies. In the present study, 30 hearts from postmortems and 76 hearts from departmental collections from embalmed cadavers (making a total of 106 hearts) were studied. Fifteen hearts (14.2%) showed muscular bridges. The length of the bridges varied from 4mm to 40mm, the majority being 10-19mm in length. Only two arteries viz, the left anterior descending artery and the left diagonal artery showed muscular bridges. PMID- 11265799 TI - Coronary artery disease epidemic in Indians: a cause for alarm and call for action. AB - Coronary artery disease (CAD) rates in urban areas in India are now 4-fold higher than in the United States (US) although the rates were similar in 1968. Both overseas and resident Indians have the highest rates of CAD, although almost half of them are life-long vegetarians. When compared to Whites, Blacks, Hispanics and other Asians, CAD rates among Indians worldwide are two to four times higher at all ages and five to ten times higher in those < 40 years of age. Although CAD is a fatal disease with no known cure, it is also highly predictable, preventable, and treatable. During the past 30 years, CAD rates halved in the US, Australia, Canada, France, Japan, and Finland. These vast reductions in CAD mortality are attributed to nationwide changes in specific risk factors that were identified through epidemiological research and addressed through population-based interventions, rather than extensive use of expensive technology. Reduction in risk factors explains most of the decline with modest contributions from advances in treatment. Ironically, the CAD rates doubled in India during the same period, primarily due to dietary changes associated with epidemiological transition from a rural sustenance economy to an urban market oriented economy. The impact of such changes appears to be greater in Indians than in other populations due to a genetic predisposition. Significant decline of CAD is readily achievable in India, by adopting a combined population-wide and high-risk primary prevention strategy. This requires concerted action by the medical profession, govemment, media, and the public. PMID- 11265800 TI - Prevention of coronary heart disease among Indians: focus on primary prevention. AB - Epidemiological transition with increasing life expectancy and demographic shifts in population age-profile combined with lifestyle related increases in the levels of cardiovascular risk factors is accelerating coronary heart disease (CHD) epidemic in India. As prospective cohort studies for evaluation of coronary risk factors do not exist, urban-rural differences in prevalence of coronary risk factors and case-control studies provide important information regarding coronary risk factors that need prevention to control the CHD epidemic. The risk factors more in urban Indians or associated with increased risk in case-control studies are: Sedentary lifestyle, smoking, truncal obesity, hypertension, hypercholesterolaemia, impaired glucose tolerance, insulin resistance and diabetes. Primordial prevention ie, prevention of risk factors can be achieved by encouragement of positive health behaviour and promotion of the concept of health as a social value. Special target groups are children, adolescents, family unit, under-privileged and high-risk groups. Behavioural and environmental changes relevant to primordial prevention are changes in eating patterns, drinking, smoking, physical activity and stress management. Primary prevention focuses on population and on high-risk groups. Specific high-risk subjects are those with family history of CHD, hypertension or diabetes or those having sedentary lifestyle, obesity, truncal obesity and biochemical coronary risk factors. The interventions are smoking cessation, increased physical activity, weight regulation, blood pressure control, lipid regulation and diabetes management. PMID- 11265801 TI - Current understanding of pathogenesis of coronary artery disease and its future implications. AB - Coronary artery disease (CAD) is the most important cause of morbidity and mortality in a population. Percutaneous coronary intervention and coronary artery by-pass grafting have greatly changed the treatment of CAD, still many questions remain unanswered. Atherosclerosis is a normal consequence of ageing but some patients may experience it at an earlier age. As regarding pathogenesis of atherosclerosis, it is described often as a focal process which is diffuse in nature primarily involving the vessel intima. Salient features of 'lesion prone areas' in atherosclerosis include increased endothelial permeability to an intimal accumulation of plasma proteins, including albumin, fibronogen and LDL. The clinical expression of atherosclerotic disease activities is determined by pathologic events leading to coronary thrombosis. A vulnerable plaque has the characteristics of: Extracellular lipid pool occupies a large proportion of overall plaque volume, the fibrous cap which separates the lipid core from luminal blood is thinner within, and high macrophage density. Typically these plaques cause < 50% cross-sectional stenosis of the artery. The contribution of CAD is clearly of both genetic and environmental in origin. An increase in shedding of cell adhesion molecules may be a characteristic of atherosclerotic lesion. There is also suggestion that plasminogen activation inhibitor type I (PAI-1) has an important role in atherogenesis. Angiogenic growth factors and their endothelial receptors function as major regulators of blood vessel formation. Thereapeutic angiogenesis can be achieved by exogenously adding VEGF and/or other angiogenic growth factors. PMID- 11265802 TI - Percutaneous transluminal myocardial revascularisation: current status and future perspectives. AB - Percutaneous transluminal myocardial revascularisation presently appears to be a potential palliative treatment for coronary artery disease, neither controlled with drugs nor amenable to available coronary revascularisation techniques. Ongoing trials will provide answer to short and long term efficacy. Recent developments using angiogenic growth factors appear very promising, and the role of growth factors as an adjunct to percutaneous transmyocardial revascularisation with laser remains to be seen. PMID- 11265803 TI - Cardiac tumors. AB - Primary cardiac tumours of the heart are rare. Primary cardiac neoplasms are benign in 75% cases. Among the benign lesions about 40% are myxomas. Approximately 25% of primary cardiac neoplasms are malignant and almost all are sarcomas and lymphomas of varying types. Clinical evaluation has been augmented by echocardiography, angiography, radionuclide studies, computed tomography and magnetic resonance imaging. Benign tumours are amenable to resection and in selected patients with malignant cardiac disease surgical intervention may be feasible. PMID- 11265804 TI - Predictive value of maximal stress test in patients showing atypical or no symptoms of ischaemic heart disease. AB - A study was carried out on 76 patients attending outpatients' department of medicine, Government Medical College, Nagpur. These patients had either (a) atypical angina or (b) no symptoms but certain factors for ischaemic heart disease. All cases were subjected to maximal stress test. Thirty-five cases (46.05%) showed a positive result on the basis of 'ST' segment depression of 2 mm or more. In the absence of facilities for coronary angiography, treadmill test is an important diagnostic tool for ischaemic heart disease. PMID- 11265805 TI - Diverse manifestation of arrhythmogenic right ventricular dysplasia in a family. AB - Arrhythmogenic right ventricular dysplasia (ARVD) is a cardiomyopathy of unknown cause associated with life-threatening arrhythmias. The purpose of this case report is two-fold: (i) To highlight the utility of a definite set of diagnostic criteria encompassing structural, histologic, electrocardiographic, arrhythmic and genetic factors in establishing diagnosis of ARVD in institutions like ours which are handicapped by lack of magnetic resonance imaging (MRI) and endomyocardial biopsy facilities, (ii) to present diverse arrhythmic manifestations in a single family. PMID- 11265806 TI - Early and long-term neuroendocrine effects of prenatal stress in male and female rats. AB - The effects of maternal stress, termed prenatal stress (PNS) on the neuroendocrine regulation of reproduction and the stress reactivity of offspring were studied in rats. PNS prevented the formation of sexual dimorphism in catecholamine levels, aromatase activity, and androgen 5alpha-reductase activity in the preoptic area of the brain and the mediobasal hypothalamus in 10-day-old rats. The morphological correlate of the functional lesions induced by PNS consisted of the elimination of gender-related differences in the volumes of neuron nuclei in the suprachiasmatic nucleus. Prenatal stress altered the stress and adrenergic reactivities of the hypothalamo-hypophyseal-adrenal system in mature males and females. The long-term effects of PNS were regarded as a consequence of the disruption of the hormone-neurotransmitter imprinting of the neuroendocrine system. PMID- 11265807 TI - Modular organization of the granular layer of the human cerebellar cortex during post-natal ontogenesis. PMID- 11265808 TI - The structural organization of the statocyst sensory system in nudibranch mollusks. PMID- 11265809 TI - The effects of a serotoninergic substrate of the nucleus accumbens on latent inhibition. AB - Latent inhibition is an effect consisting of a delay in the acquisition of a stimulus in response to quenching of its significance by repeated presentation (pre-exposure) before combination with the reinforcement. This phenomenon is used for studies of the mechanisms of attention. Experiments were performed on rats to determine whether latent inhibition could be formed in a passive avoidance response in conditions of blockade of serotoninergic terminals in the nucleus accumbens. After pre-exposure, sham-operated animals demonstrated a delay in acquisition of the stimulus as compared with animals not subjected to pre exposure. Bilateral injection of the neurotoxin 5,7-dihydroxytryptamine into the nucleus accumbens led to disruption of latent inhibition, which could in turn be prevented by systemic administration of haloperidol before training. The importance of serotoninergic terminals in the nucleus accumbens for latent inhibition is discussed, along with the mechanism of their interaction with the dopaminergic system during the formation of this effect. PMID- 11265810 TI - Optical recording of responses to odor in olfactory structures of the nervous system in the terrestrial mollusk Helix. AB - Electrophysiological methods and optical methods based on the use of potential sensitive dyes were used to record stable rhythmic oscillations of local potentials in the olfactory structure (procerebrum) of the pulmonate mollusk Helix: these oscillations were generally similar to those previously observed in slugs. Odor had the effect of transiently altering rhythmic oscillations to generate an individual pattern. This is the first study describing the recording of procerebrum potentials evoked by presentation of odor, with mapping of the areas of propagation of these potentials relative to the areas of propagation of rhythmic oscillations. The boundary of the propagation of the evoked potential was essentially similar to the projection of the neuropil, and rhythmic oscillations were recorded in the projection layer of procerebrum cell bodies. Evoked potential waves appeared in areas corresponding to the site at which the olfactory nerve enters the cerebral ganglion (of which the procerebrum forms a part) and were propagated in the procerebrum neuropil towards the cell body layer. Evoked potentials did not provoke out-of-phase waves of rhythmic oscillations. PMID- 11265812 TI - I. P. Pavlov's theory of higher nervous activity: landmarks and developmental trends. PMID- 11265811 TI - Reverberation of excitation in living "hippocampal formation-entorhinal cortex" slices from rats. Optical recording. AB - A vital potential-dependent dye was used to conduct optical recording of the electrical activity of the hippocampal formation in living slices of the rat brain including the hippocampal formation and the entorhinal cortex. These studies showed that single electrical stimuli applied to the entorhinal cortex, subiculum. and dentate gyrus produced responses in which waves of excitation passed across the hippocampal formation sequentially from the dentate gyrus, through CA3, to the CA1 field of the hippocampus. When GABAergic inhibition was partially blocked with picrotoxin, the first wave of excitation was immediately followed by several further waves in all zones of the hippocampal formation, with a constant shift in latency, which increased from the dentate gyrus to CA3 and CA1. Reverberation of excitation in the "hippocampal formation-entorhinal cortex" structure is regarded as the most probable cause for the appearance of these sequences of waves. PMID- 11265813 TI - Rhythmic alpha-activity generators in the human EEG. AB - This study was designed to locate the neuronal mechanisms of rhythmic activity in the human brain in the alpha-rhythm range using an equivalent dipole model. The experiments approach used here is based on using the "imposed rhythm" phenomenon, whereby a light stimulus flashing with a frequency close to the frequency of the individual alpha rhythm increases its power in the EEG spectrum. The results obtained showed that the activity of the structures generating the alpha rhythm could be differentiated by using a specific photostimulation frequency and were located by constructing a dipole model. Two sources of alpha-rhythm generation were found, located in the thalamic structures of the brain and operating over a narrow-band frequency range with maximum response resonance frequencies of 10.1 and 10.5 Hz. PMID- 11265814 TI - Generalized post-tetanic changes in excitatory postsynaptic and acetylcholine evoked currents in neurons in the common snail. AB - Changes in excitatory postsynaptic currents (EPSC) and influx currents induced by local iontophoretic application of acetylcholine (ACh currents) to the surfaces of the bodies of identified neurons LPa3 and RPa3 in the common snail after heteroreceptor tetanic stimulation were studied. The effects of rhythmic application of acetylcholine (0.5-1.0 times per sec for 10-40 sec) on the magnitude of EPSC were analyzed, along with the effects of tetanic stimulation of the nerves (n. intestinalis, n. pallialis dexter, n. pallialis sinister; 1-5 Hz for 1-2 min) on the amplitudes of the ACh current and the magnitudes of EPSC evoked by stimulation of another nerve. The following changes in conditioning responses were found over 1-1.5 h after tetanization: rhythmic application of ACh to the dorsal surface of the cell body resulted in potentiation of EPSC, and tetanic orthodromic stimulation potentiated the ACh current and induced heterosynaptic depression of EPSC. These post-tetanic changes were not local- they developed in those chemoreceptor zones of the membrane which were not activated by the preceding conditioning rhythmic stimulation. It is concluded that generalized post-tetanic changes in neuron responses are induced by activation of its subsynaptic and extrasynaptic mediator chemoreceptors. PMID- 11265815 TI - The effects of scopolamine on the spatial organization of cortical potentials in the rat brain. PMID- 11265816 TI - Changes in direct and interhemisphere responses of the pyramidal tract after tetanization of the cortex and lateral hypothalamus. PMID- 11265817 TI - The duration of persistence of the electrical characteristics of command neurons during acquisition of long-term sensitization in the common snail. PMID- 11265819 TI - The effects of prenatal stress on learning in rats in a Morris maze. AB - The offspring of female Wistar rats subjected to daily stress (they were placed in an unfamiliar social group for 1 h) during the last third of pregnancy were studied. The offspring of these females were tested for the ability to perform spatial orientation in a Morris water maze at the ages of two and four months. Prenatal stress had no effect on the ability of rats to learn in the Morris maze. However, two-month-old animals subjected to prenatal stress, unlike controls, demonstrated less flexibility in their behavioral strategy in solving the spatial orientation task. These animals were characterized by a clear tendency for their behavior to perseverate. By the age of four months, the differences between the control and prenatally stressed animals had disappeared. PMID- 11265820 TI - Changes in various measures of immune status in mice subject to chronic social conflict. AB - Levels of the major regulatory subpopulations of lymphocytes in the thymus and spleen and blood lymphocyte dehydrogenase levels were measured in male mice with aggressive and submissive patterns of behavior formed over 10 or 20 aggressive confrontations leading to repeated experience of victory or defeat. The results showed that repeated experience of social confrontation non-specifically increased the proportion of segmented neutrophils and lactate dehydrogenase activity in both participants in aggressive encounters, and decreased the numbers of CD4+ and CD8+ T-lymphocytes in the spleen. Succinate dehydrogenase specifically decreased in the lymphocytes of aggressive mice and increased in the lymphocytes of submissive mice. The proportion of CD4+ T-lymphocytes in victims' thymuses also decreased. Changes in metabolic measures and percentage ratios of lymphocyte contents were dynamic and depended on the duration of confrontational interactions. PMID- 11265818 TI - Anti-dopamine antibodies: effects on behavior in an "open field," pain sensitivity, CNS monoamine content, and functional activity of immunocytes in C57Bl/6 mice. AB - Single i.p. doses of anti-dopamine antibodies were given to C57Bl/6 mice. This resulted in inhibition of motor activity in a large proportion of the animals in the "open field" test, which lasted five days. Hyperalgesia, detected 1.5 h and 1 day after doses of antibody, was replaced by analgesia on day 5. There was a sharp reduction in the levels of dopamine and its metabolites in the cerebral cortex at 1 and 5 days; the serotonin level was increased 1 day after doses of antibody, and was significantly decreased at 5 days. There was no effect on cells of the immune system. The possible mechanisms of the neurotropic action of these antibodies are discussed. PMID- 11265821 TI - The effects of delta sleep-inducing peptide on the intensity of lipid peroxidation and xanthine oxidase activity in rat tissues during cold stress. AB - Exogenous delta sleep-inducing peptide given i.p. to intact rats at a dose of 12 microg/100 g decreased the levels of diene conjugates and Schiff bases in liver and brain tissues and had no effect on xanthine oxidase activity in these tissues. Cold stress was accompanied by increases in xanthine oxidase activity in rat liver and brain, with a consequent accumulation of diene conjugates and Schiff bases, as compared with intact animals. Preliminary administration of delta sleep-inducing peptide before three days of cold stress led to decreases in xanthine oxidase activity and lipid peroxidation products in the liver and brain, as compared with values in stressed rats. The protective effect of delta sleep inducing peptide in stress is discussed. PMID- 11265822 TI - The development of the scientific ideas of I. P. Pavlov on the goal reflex in studies of the mechanisms of biological motivation. AB - Data obtained in experimental studies developing I. P. Pavlov's concept of the "goal reflex" are presented. It is shown that purposive behavior in the systems organization of behavioral acts is constructed by the dominant motivation. This article discusses mechanisms determining the energy of biological motivations and their controlling component--the results of action acceptor. The role of reinforcement in the mechanisms forming molecular engrams of behavior on structures of the results of action acceptor is assessed. PMID- 11265823 TI - Changes in enzyme activity in brain structures in august rats in response to delta sleep peptide in conditions of prolonged administration of L dihydroxyphenylalanine. PMID- 11265824 TI - Responses of populations of mitochondria in rat sensorimotor cortex neurons to prolonged continuous exposure to low-frequency vibration. PMID- 11265825 TI - Researcher and researched--community perspectives: toward bridging the gap. AB - In the process of initiating a new community-based research project, the authors wanted to understand the experiences of community members and researchers in community-based research projects and to develop guidelines to improve future projects. They conducted qualitative, key informant interviews with 41 people involved at all levels of community-based research projects in Seattle. Respondents were identified using a snowball sampling technique. More problems than successes were discussed by informants, including dissatisfaction with the focus of research, which some said is marked by a lack of cultural appropriateness and relevance. Power imbalances, lack of trust, and communication difficulties impeded collaboration. According to respondents, many problems could be avoided if the community were involved from the beginning in setting research priorities and developing and implementing interventions. Meaningful collaboration between communities and researchers is characterized by early involvement of communities, power sharing, mutual respect, community benefit, and cultural sensitivity. PMID- 11265826 TI - "I was not invited to be a [CHW] ... I asked to be one": motives for community mobilization among women community health workers in Mexico. AB - Despite health educators' revitalized interest in community mobilization for health, little attention has been given to participants' motives for mobilizing. The purpose of this article is to contribute to the understanding of community mobilizing by analyzing the motives for mobilization among a group of women community health workers (CHWs), members of a community-based organization in Mexico. The study is guided by critical feminist and social constructivist theories. It aims at identifying the categories of motives used by women CHWs and exploring how these motives are created while presenting women's own voices. Women's motives fall into four categories: getting out, serving, learning, and women's betterment. These motives blend personal andpublic motives. The analysis suggests that mobilization for health may be enhanced by addressing both the personal satisfaction of individuals and the accomplishment of public goods. The study of motives may be useful for the recruitment and retention of participants in community mobilization efforts. PMID- 11265827 TI - Schoolwide effects of a multicomponent HIV, STD, and pregnancy prevention program for high school students. AB - Few studies have tested schoolwide interventions to reduce sexual risk behavior, and none have demonstrated significant schoolwide effects. This study evaluates the schoolwide effects of Safer Choices, a multicomponent, behavioral theory based HIV, STD, and pregnancy prevention program, on risk behavior, school climate, and psychosocial variables. Twenty urban high schools were randomized, and cross-sectional samples of classes were surveyed at baseline, the end of intervention (19 months after baseline), and 31 months afterbaseline. At 19 months, the program had a positive effect on the frequency of sex without a condom. At 31 months, students in Safer Choices schools reported having sexual intercourse without a condom with fewer partners. The program positively affected psychosocial variables and school climate for HIV/STD and pregnancy prevention. The program did not influence the prevalence of recent sexual intercourse. Schoolwide changes in condomuse demonstrated that aschool-based program can reduce the sexual risk behavior of adolescents. PMID- 11265828 TI - Project ARM: alcohol risk management to prevent sales to underage and intoxicated patrons. AB - Clear policies and expectations are key to increasing responsible service of alcohol in licensed establishments. Few training programs focus exclusively on owners and managers of alcohol establishments to reduce the risk of alcohol service. Project ARM: Alcohol Risk Management is a one-on-one consultation program for owners and managers. Participants received information on risk level, policies to prevent illegal sales, legal issues, and staff communication. This nonrandomized demonstration project was implemented in five diverse bars. Two waves of underage and pseudo-intoxicated purchase attempts were conducted pre- and postintervention in the five intervention bars and nine matched control bars. Underage sales decreased by 11.5%, and sales to pseudo-intoxicated buyers decreased by 46%. Results were in the hypothesized direction but not statistically significant. A one-on-one, outlet-specific training program for owners and managers is a promising way to reduce illegal alcohol sales, particularly to obviously intoxicated individuals. PMID- 11265829 TI - Older women and mammography screening behavior: do possible selves contribute? AB - This study sought to explore the contribution of the self-concept to older women's adherence to regular mammography screening behavior. The PRECEDE and health belief model concepts were incorporated with a measure of the women's future selves to determine whether the self-concept adds to our ability to predict screening. A self-administered questionnaire was completed by 210 community-dwelling women ages 50 to 75 years, recruited from urban and rural women's groups. Logistic regression analyses revealed that predictors of adherence were clinical breast examination, physician recommendation, age, barriers, benefits, feared health-related possible self, and self-efficacy in the feared domain. The addition of the self measures significantly improved the overall fit of the model. Implications for theory development, practice, and future research are discussed. PMID- 11265830 TI - Privatization of local health department services: effects on the practice of health education. AB - Local health departments (LHDs) are changing service delivery mechanisms to accommodate changes in health care financing and decreased public support for governmental services. This study examined the extent to which North Carolina LHDs privatized and contracted out services and the effects on the time spent on core functions of public health and activities of health educators. Questionnaires were mailed to the senior health educators in all LHDs. Sixty-nine responded, and 68% of LHDs had not privatized any services other than laboratory and home health. Clinical services were more commonly privatized than nonclinical services. Respondents perceived that privatization produces more time for LHDs to address the core public health functions and for health educators to engage in appropriate professional activities. Health educators in LHDs that had not privatized were more likely to be concerned about potential negative effects. This study suggests that privatization has generally had a positive effect on the roles of health educators in North Carolina LHDs. PMID- 11265831 TI - Searching for evidence about health education and health behavior interventions. AB - Evidence is fundamental to science, but finding the right evidence in health education and health behavior (HEHB) is often a challenge. The authors discuss some of the controversies about the types of evidence that should be considered acceptable in HEHB, the tension between the use of qualitative versus quantitative data, the need for measures of important but neglected constructs, and interpretation of data from experimental and nonexperimental research. This article discusses some of the challenges to the use of evidence and describes a number of strategies and some forces encouraging the use of evidence-based interventions. Finally, the authors suggest ways to improve the practice and dissemination of evidence-based HEHB. Ultimately, if evidence-based interventions are not disseminated, the interventions will not achieve their potential. The goal should be to develop more effective interventions and disseminate them to improve the public's health. PMID- 11265832 TI - Suicidology in Ireland--a missed opportunity. PMID- 11265833 TI - Family murder suicides in Kerala. PMID- 11265834 TI - Letter across the Pacific. PMID- 11265835 TI - Could "How-to-commit-suicide" book prevent suicide? PMID- 11265836 TI - Elevated rates of suicidal behavior in gay, lesbian, and bisexual youth. AB - Both clinical and epidemiological literature point to elevated rates of suicidal behaviors in gay, lesbian, and bisexual youth (GLBY). Recent North American and New Zealand studies of large populations (especially the US Youth Risk Behavior Surveys from several states) indicate that gay, lesbian, and bisexual adolescents (males in particular) can have rates of serious suicide attempts at least four times those of apparently heterosexual youth. There are various reasons why this figure is likely to be an underestimate. Reasons for these elevated rates of suicidal behavior include a climate of homophobic persecution in schools, and sometimes in family and community--values and actions that stigmatize homosexuality and that the youth who has not yet "come out" has to endure in silence. PMID- 11265837 TI - Suicide rates and mental health services in modern China. AB - Suicide rates in China have in the past been reported to be very low for a variety of historical and political reasons. In recent years, however, the reported suicide rates in China have increased alarmingly among certain age groups. This article reviews reports of the national rural suicide rates in China for 1992, gathered from the Annal of Chinese Public Health, which has previously never been reported publicly. The highest suicide rates occur in the rural areas and among young women and men over 60 years. These data reveal that suicide in China may have some unique characteristic associated with a variety of socio cultural variables, such as traditional culture, social class, economic status, health care levels, and interpersonal problems. The author shows that lack of mental health services in rural areas in China may be considered one of associated reasons to the high rural suicide rate in China. PMID- 11265838 TI - Double suicide and homicide-suicide in Switzerland. AB - Double suicide and homicide-suicide are infrequent and are different in psychopathology from that of a single suicide. However, precipitating factors for double suicide and homicide-suicide are similar to these found in single suicide. Depression, borderline disturbances, and narcissistic neuroses in combination with stressors such as physical illness, isolation, and social losses can lead to homicide-suicide. Epidemiological studies indicate that double suicide and homicide-suicide are influenced by ethnicity and cultural and social background. This study explores the situation in Switzerland between 1928-48 and 1971-1990. PMID- 11265839 TI - Education and debriefing: strategies for preventing crises in crisis-line volunteers. AB - Telephone crisis lines offer an important service to individuals in crisis. The accessibility as well as a lack of other means of support leads many individuals to call the line. The role of the volunteer is to listen and support the caller as well as provide information and referrals to other agencies. Agencies are presented with a high turnover of volunteers and are then faced with the task of recruiting and training replacements. Volunteers are often exposed to horrific accounts of human pain and suffering which may affect their personal thoughts, feelings, beliefs and actions and influence the decision to quit. Compassion fatigue is one term used for this inherent "cost of caring." Many factors contribute to this cost including the nature of crisis calls, the repeat caller, and personal coping mechanisms. Educating and debriefing the volunteer are two strategies that may prevent the onset of compassion fatigue and volunteer resignation. Debriefing is viewed as an effective strategy for volunteers as it has been found to be successfull in assisting other helpers in many different contexts to cope and deal with the traumatic events that they experience or hear about. PMID- 11265841 TI - The 8th European Symposium on Suicide and Suicidal Behavior. PMID- 11265840 TI - A mass-distributed CD-ROM for school-based suicide prevention. AB - Recent violent events in schools in the United States have underscored the need for an examination of efforts aimed at mass education of school professionals on issues related to violence prevention and crisis intervention. A CD-ROM ("Team Up to Save Lives: What Your School Should Know About Preventing Youth Suicide") was mass mailed to schools throughout the United States in January 1997. We conducted an initial evaluation of this intervention in the Chicago area and found that the scope of the CD's impact was relatively narrow: Only 39% of all schools with informants that were contacted actually reviewed the CD-ROM and provided feed back for the evaluation. Insufficient time, computer equipment, and training were the major obstacles to CD-ROM use and to participation in the evaluation. Among those reviewing the CD-ROM, overall perceptions regarding its utility were extremely favorable. Multiple instances in which the CD had been used to address actual crisis situations were identified. PMID- 11265843 TI - Left ventricular assist device bridge to recovery: a review of the current status. AB - The use of the left ventricular assist device as a bridge to recovery represents a new phenomenon. This article focuses on bridge-to-recovery in the settings of myocarditis and dilated cardiomyopathy with a review of the hemodynamic, neurohormonal, physiologic, cellular, and molecular changes of recovery during left ventricular assist device support. Despite numerous markers of success, there is a disconnect from the limited clinical successes that are reviewed. The current status and future options to increase the chances of success are highlighted. PMID- 11265842 TI - Living with loss. The Irish Association of Suicidology Conference, Derry, Northern Ireland, 1999. PMID- 11265844 TI - Bridging-to-recovery. AB - BACKGROUND: Patients with end-stage heart failure placed on a cardiac assist device show at least some degree of improvement of cardiac function. In a subgroup of selected patients, some hearts recovered considerable function. In these patients the device was removed and cardiac transplantation was no longer necessary. We report our long-term experience with these weaned patients. METHODS: As of today, 512 cardiac assist devices of various types (Berlin Heart, Berlin, Germany; Novacor, World Heart, Ottawa, Ontario, Canada; TCI, ThermoCardio Systems, Inc, Woburn, MA; DeBakey, Micromed Technology Inc, Houston, TX) were implanted in patients with end-stage heart failure in our institution. Of these, 95 patients belonged to a subgroup of patients with nonischemic, idiopathic, dilated cardiomyopathy who were implanted with a left ventricular support system (Novacor 84, TCI 10, Berlin Heart 1) between 1994 and 2000. All were routinely examined by echocardiography for improvement of cardiac function. The left ventricular diameter in diastole (LVIDd) and left ventricular ejection fraction (LVEF) served as the main parameters to assess changes in cardiac performance. Under the conditions of a running device, an LVIDd below 60 mm and an LVEF above 40% were the criteria to do further echocardiographic studies when the pump was turned off for up to 20 minutes. RESULTS: Twenty-eight patients (26 men, 2 women; ages 18 to 64 yrs; history of heart failure, 1 to 17 yrs) fulfilled the criteria of improved cardiac performance and were weaned from the device. Since then, 16 patients have continued "normal" heart function with follow-up times ranging from 1 month to 5.5 years (group B). Three patients died of noncardiac causes (group C). Eight patients were transplanted from 1 to 17 months later and one died on the waiting list (group A). Statistically significant differences between groups A and B were calculated for the duration of heart failure (9 versus 2 years, p = 0.0002). Differences in LVIDd before removal of the device (57 versus 51 mm, p = 0.0420), LVEF after 2 months of unloading (30 versus 49%, p = 0.0300), and LVEF preexplantation (43 versus 52%, p = 0.0001) were significant. Overall, 17% of the cohort of 95 patients were weaned successfully. CONCLUSIONS: Weaning from cardiac assist devices is feasible for selected patients; it saves donor hearts and is preferred to cardiac transplantation. However, as of today no reliable parameter predicts outcome after weaning and none determines the possibility of device removal before implantation in advance. PMID- 11265845 TI - HeartMate II left ventricular assist system: from concept to first clinical use. AB - The HeartMate II left ventricular assist device (LVAD) (ThermoCardiosystems, Inc, Woburn, MA) has evolved from 1991 when a partnership was struck between the McGowan Center of the University of Pittsburgh and Nimbus Company. Early iterations were conceptually based on axial-flow mini-pumps (Hemopump) and began with purge bearings. As the project developed, so did the understanding of new bearings, computational fluid design and flow visualization, and speed control algorithms. The acquisition of Nimbus by ThermoCardiosystems, Inc (TCI) sped developments of cannulas, controller, and power/monitor units. The system has been successfully tested in more than 40 calves since 1997 and the first human implant occurred in July 2000. Multicenter safety and feasibility trials are planned for Europe and soon thereafter a trial will be started in the United States to test 6-month survival in end-stage heart failure. PMID- 11265846 TI - Clinical results with the AB-180 left ventricular assist device. AB - BACKGROUND: This report reviews the initial clinical experience with the AB-180 ventricular assist device. METHODS: Between Dec 1997 and July 2000, the AB-180 was implanted in 17 patients at five institutions. The mean age was 52 years (range 21 to 68 years) and 14 of 17 were male. The indications for implantation were postcardiotomy shock (12 of 17, 70%), decompensated cardiomyopathy (2 of 17, 12%), viral myocarditis (2 of 17, 12%), and acute myocardial infarction (1 of 17, 6%). RESULTS: The mean duration of support was 8.5 days (range 1 to 28 days). In the group of 17 patients, 8 were weaned from the device and 2 underwent transplantation. Four of the weaned patients (4 of 8, 50%) and 1 of the transplant patients (1 of 2, 50%) survived. The overall weaning and survival rates were 58% (10 of 17) and 29% (5 of 17). There were no major device-related complications and no major device malfunctions. CONCLUSIONS: The AB-180 provides reliable circulatory support for reversible forms of heart failure. PMID- 11265847 TI - Research and development of an implantable, axial-flow left ventricular assist device: the Jarvik 2000 Heart. AB - Advances in technology and increased clinical need have led to the development of a new type of blood pump. The Jarvik 2000 Heart is an electrically powered, axial flow left ventricular assist device that has been developed during the past 13 years. Unlike first-generation left ventricular assist devices, which were developed in the 1970s and were designed to totally capture the cardiac output, the Jarvik 2000 is designed to normalize the cardiac output by augmenting the function of the chronically failed heart for extended periods. Design iterations have been tested in 67 animals, and clinical trials have recently begun. Three patients have received the Jarvik 2000 as a bridge to transplantation, and 1 patient is being supported permanently outside the hospital. All 4 patients have improved from New York Heart Association functional class IV to class I, and 2 of them have been discharged from the hospital after heart transplantation. The experimental and clinical results indicate that the Jarvik 2000 can provide physiologic support with minimal complications and is reliable, biocompatible, and easy to implant. PMID- 11265849 TI - Lessons learned from the first clinical implants of the DeBakey ventricular assist device axial pump: a single center report. AB - BACKGROUND: The bridge to transplantation with pulsatile mechanical assist devices became a standard procedure for patients deteriorating on the waiting list. Recently, continuous flow axial impeller pumps were introduced to clinical application offering new advantages. METHODS: From November 1998 till September 2000, 6 male patients (mean age 53 plus or minus 11 years) with end-stage left heart failure were implanted with a DeBakey ventricular assist device (VAD) axial flow pump for bridge to transplantation. RESULTS: Three patients were successfully transplanted after 74, 115, and 117 days, respectively. Two other patients died after 25 and 133 days. One patient is still on the device after 108 days. Because of modification of the implantation technique after the first 2 patients, mean pump-flow within the first 3 weeks was increased from 4.3 +/- 0.6 L/min to 6.7 +/- 0.3 L/min. Patients were put on regular bicycle-ergometer training and improved their exercise capacities up to a mean maximum oxygen consumption of 20.2 mL/kg/min. CONCLUSIONS: Initial implants of the DeBakey VAD demonstrated support properties comparable to pulsatile pumps but without significant restrictions for extended use. PMID- 11265848 TI - Clinical experience with the MicroMed DeBakey ventricular assist device. AB - BACKGROUND: The MicroMed DeBakey ventricular assist device (VAD) (MicroMed Technology, Inc, Houston, TX) is the first long-term axial flow circulatory assist device to be introduced into clinical trials as a bridge to transplantation. Clinical trials began in Europe in November 1998 and in the United States in June 2000. METHODS: To qualify for the study, the patients must be listed for cardiac transplantation and must have demonstrated profound cardiac failure. There were no exclusions to the MicroMed DeBakey VAD implant other than those patients who would typically be excluded from cardiac transplantation. RESULTS: As of September 2000, 51 patients have been implanted with the MicroMed DeBakey VAD. A detailed evaluation of the first 32 patients has been completed. With current data, the probability of survival at 30 days after VAD implant is 81%. CONCLUSIONS: The clinical trial demonstrated that the MicroMed DeBakey VAD is capable of providing adequate circulatory support in patients with severe heart failure, sufficient to recover and return to normal activities while awaiting a heart transplantation. Much has been learned about the function of the device and its continuous flow in humans. PMID- 11265850 TI - Current status of the AbioCor implantable replacement heart. AB - BACKGROUND: The AbioCor implantable replacement heart (IRH) has been developed as an alternative to transplant (ie, destination therapy). We report our experience with the AbioCor IRH in a bovine model at the University of Louisville. METHODS: Male Holstein cows were used (85 to 115 kg). The internal controller, battery, and secondary transcutaneous energy transfer coil were implanted in the right flank. After cardiopulmonary bypass, the thoracic unit was implanted orthotopically. After removal of air and weaning from cardiopulmonary bypass, the AbioCor was connected to internal components and energy transfer through transcutaneous energy transfer coils was achieved. RESULTS: Nineteen animals underwent implantation of the AbioCor IRH for a proposed 30-day duration. There were 6 deaths, none related to device malfunction. All animals demonstrated normal hemodynamics with normal pressures in the aorta, pulmonary artery, left atrium, and right atrium. There was no significant hemolysis and all animals demonstrated normal end organ function. The internal battery allowed for brief periods of untethered mobility. CONCLUSIONS: The AbioCor IRH has resulted in normal hemodynamics and normal end organ function without evidence of hemolysis in a bovine model. PMID- 11265851 TI - Modifications in surgical implantation of the Penn State electric total artificial heart. AB - BACKGROUND: Two modifications of the surgical implantation protocol for the Penn State Total Artificial Heart (ETAH) were evaluated: Phrenic nerve ischemia was prevented by minimizing dissection and traction; and hemostasis was augmented and ETAH cuff anastomoses reinforced by using fibrin glue. METHODS: Thirteen Holstein calves underwent orthotopic surgical implantation of the Penn State ETAH between February 1998 and August 2000. Mean hemodynamic and laboratory chemistry variables from the first postoperative week were compared between calves receiving the original (n = 7) and modified (n = 6) protocol. RESULTS: Calves assigned to the modified protocol displayed an improvement in the Po2/FiO2 ratio compared to original (419.4 +/- 17.5 vs 336.3 +/- 35.4, respectively; p = 0.05). All additional parameters were equivalent between groups. The percent survival of animals receiving the modified protocol at 2, 4, and 12 weeks was higher than that of animals that underwent the original protocol. Original-protocol calf deaths consisting of hemothorax (n = 3), and respiratory failure (n = 1) were prevented in the modified protocol. CONCLUSIONS: Our results suggest that manipulations in surgical protocol may promote increased survival in calves implanted with the Penn State ETAH. PMID- 11265852 TI - The LionHeart LVD-2000: a completely implanted left ventricular assist device for chronic circulatory support. AB - Management of patients with end-stage cardiac disease remains a vexing problem. Limitations in medical management and a fixed supply of donor organs for cardiac transplant have a continued impact on this growing population of patients. Mechanical circulatory support has proved very successful as a means of bridging patients to cardiac transplant when all medical options have been exhausted. The development of a chronic system of circulatory support has been underway at the Pennsylvania State University for nearly 30 years. These efforts have been recently merged with the industrial partnership with Arrow International toward the development of the LionHeart LVD-2000 (Arrow International, Reading, PA) completely implanted left ventricular support system. We present an overview of the system, details of implantation, a review of preclinical studies, and a synopsis of the first European implants. Early results have demonstrated the system to be safe, effective, and reliable. Transcutaneous energy transmission and the compliance chamber have been validated. PMID- 11265853 TI - The totally implantable novacor left ventricular assist system. AB - The Novacor Left Ventricular Assist System (LVAS) (Novacor Corp, Oakland, CA) was initially console-based and has been available since 1993 in a wearable configuration. It has been successfully used for the past 16 years as a bridge to cardiac transplantation in patients with end-stage congestive heart failure. The Stanford experience represents 53 patients (48 male, 5 female) with a mean age of 44 +/- 13 years (16 to 62) and a mean support time of 56 +/- 76 days (1 to 374). Complications with LVAS use consisted predominantly of bleeding (43%), infection, (30%), and embolic cerebrovascular events (24.5%). Sixty-six percent of the supported patients were successfully bridged to cardiac transplantation. In animal studies, 4 sheep had the totally implantable configuration in place for a cumulative duration of 1 year with 1 animal supported for 260 days. The next generation Novacor LVAS will be small, quiet, and fully implantable without the need for volume compensation. It will also provide physiologic pulsatile flow and will be fail-safe. PMID- 11265854 TI - The HeartSaver left ventricular assist device: an update. AB - BACKGROUND: Ventricular assist devices have been shown to be effective as bridges to transplantation and recovery for patients with end-stage heart failure. Current technology has been limited because of the need for percutaneous connections with controllers. The HeartSaver ventricular assist device (VAD) (World Heart Corporation, Ottawa, Ontario, Canada) was developed with the intention of having a completely implantable, portable VAD system. The system consists of an electrohydraulic blood pump, internal and external battery power, and a transcutaneous energy transfer and telemetry unit that allows for power transmission through the skin. Control of the device may be achieved locally or remotely through a variety of communication systems. METHODS: The device has been modified with the Series II preclinical version being available for in vitro (mock loop) and in vivo (bovine model) testing. RESULTS: Seventeen Series II devices have been functional on mock loops or other testing trials for an accumulated 900 days of operation. There have been eight acute experiments using a bovine model to test various components as they have become available from manufacturing. Mean pump output was 10.4 +/- 1.1 L/min in full-fill/full-eject mode. Changes in the last 24 months include (1) cannula redesign for better port alignment and integration of tissue valves; (2) battery redesign to convert to new lithium-ion cells; (3) optimized infrared information and electromagnetic inductance energy transmission through various skin thicknesses and pigmentation; and (4) improved reliability of internal and external controller hardware and software. CONCLUSIONS: Modifications have been required to optimize the HeartSaver VAD's performance. The final HeartSaver VAD design will be produced in the near future to allow for formal in vitro and in vivo testing before clinical implantation. PMID- 11265855 TI - A versatile intracorporeal ventricular assist device based on the thoratec VAD system. AB - BACKGROUND: As patients are supported for longer durations with paracorporeal Thoratec left ventricular and biventricular assist devices (longest durations: 515 and 457 days, respectively), there is a need for implantable options. METHODS: We are developing a small, simple, and versatile intracorporeal ventricular assist device (IVAD) for left, right, or biventricular support as an alternative to the large, implantable, pulsatile left ventricular assist device (LVAD) systems available today. The new device is based on the Thoratec paracorporeal VAD that has been used in more than 1,400 patients weighing from 17 to 144 kg and for durations exceeding 1 year including patient discharge (using the portable driver). RESULTS: The IVAD has the same blood flow path and Thoralon polyurethane blood pumping sac as the paracorporeal VAD, but the housing is a smooth contoured, polished titanium alloy. The IVAD has a new sensor to detect when the pump is full and empty, and is controlled with the Thoratec TLC-II portable VAD driver, which is a small, briefcase-sized, battery-powered, pneumatic control unit. A small flexible (9 mm OD) percutaneous pneumatic driveline for each VAD is tunneled out of the body from the LVAD or right VAD in a pre- or intraperitoneal position. Small size and simplicity are the major advantages of the new device. The IVAD weight (339 g) and implanted volume (252 mL) are approximately one-half that of the current implantable pulsatile electromechanical LVAD systems. CONCLUSIONS: The small size of the IVAD should not only allow support of a large range of patient sizes and body habitus, but also provide options for implantable left, right, or biventricular support. By implanting only the mechanically simple blood pump, the more complex control unit is external, where it can be serviced and replaced without surgery. The IVAD with the portable driver will be a viable alternative to large implanted electromechanical systems and should address a larger segment of the physically diverse patient population. PMID- 11265856 TI - Advanced mechanical circulatory support with the HeartMate left ventricular assist device in the year 2000. AB - This paper describes the HeartMate left ventricular assist device (ThermoCardiosystems Inc, Woburn, MA) technology, which has been successfully introduced into the clinical arena with more than 2,400 implants as of the year 2000. The review summarizes the clinical experience, and identifies the benefits and limitations of the current state-of-the-art technology of the leading implantable circulatory support system. PMID- 11265857 TI - Passive ventricular constraint for the treatment of congestive heart failure. AB - Progressive heart failure is characterized by loss of cardiac function associated with maladaptive changes in myocardial gene expression and neuroendocrine activity, leading to progressive increases in heart size. Elevated ventricular wall stress results from an increase in chamber size, and is thought to play a role in furthering development towards end-stage disease. Reduction of wall stress and stress mediated myocardial stretch may be an important means for mitigating heart failure progression. One possible approach to accomplish this goal is through passive support of the heart with the Cardiac Support Device. Results from preclinical and clinical evaluation give support to this premise. PMID- 11265858 TI - Direct cardiac compression for cardiogenic shock with the CardioSupport system. AB - Epicardial direct cardiac compression for cardiogenic shock avoids a blood surface interface with associated thromboembolic and immunologic sequelae and could be placed rapidly with technical ease. The Cardio Technologies device provides synchronized biventricular cardiac compression, is placed via a thoracotomy, and remains on the heart without need for sutures. In preclinical work, the system has successfully restored cardiac function to near normal in the setting of heart failure. The CardioSupport system offers an attractive and novel alternative for treating cardiogenic shock and is being prepared for upcoming clinical trials. PMID- 11265859 TI - An anatomically compatible, wearless, reliable, and nonthrombogenic centrifugal blood pump. PMID- 11265860 TI - The CorAid blood pump. PMID- 11265861 TI - Economics of ventricular assist devices: European view. AB - Among the treatment options for congestive heart failure, ventricular assist devices (VADs) are an emerging new technology, which may serve as a realistic therapeutic option once the complication rate is reduced. Whereas these devices were used in the past almost exclusively as a bridge to transplant, today permanent VAD and full-implantable VAD are considered for destination therapy. It is therefore necessary to compare this treatment modality with other current strategies with regard to not only current survival, quality of life, and complication metrics but also relative costs between treatments. Comparable studies evaluating treatment costs are rare and future research must focus on theses issues because of the demands of permanent cost containment. PMID- 11265862 TI - The cost of long-term LVAD implantation. AB - BACKGROUND: With increasing use of left ventricular assist devices (LVAD) worldwide, the economics of LVAD implantation have become an important focus of concern. Although these devices have high unit costs, they are the only hope for survival for a large group of terminally ill patients and are likely to have an expansion in indications for use. METHODS: We calculated the costs associated with long-term LVAD implantation. We used the ratio of cost-to-charges method to calculate hospital costs per resource category, market prices for drugs and device, and payments for physician services. RESULTS: Based on our experience with "bridge-to-transplantation" patients, we estimated average first-year costs to be $222,460 including professional fees and $192,154 excluding professional fees. The latter figure is comparable to average first-year costs for cardiac transplantation, which is $176,605 without professional fees at our institution. CONCLUSIONS: The costs of LVAD therapy will change after the first year of implantation, and device reliability and longevity will be important factors in determining these costs. Should the costs of LVAD therapy continue to track those of cardiac transplantation, devices will be cost-effective only if they offer similar efficacy to cardiac transplantation. PMID- 11265863 TI - Economics of devices. AB - BACKGROUND: The economics of devices used for mechanical circulatory support not only involve the patient, the provider, and society as a whole, but also, importantly, the developers and manufacturers of these new technologies. The combined effects of years of development and testing with significant regulatory, reimbursement, and acceptance barriers make this a capital-intensive and high risk endeavor. In addition, long-term circulatory support is, today, essentially limited to bridge to transplantation, a "market" of only $100 million. Competition is increasing, with new devices under development and entering clinical trials. CONCLUSIONS: Economic health for this new industry is dependent on expanding clinical indications to definitive or destination therapy, and perhaps other applications such as bridge to recovery and assisted medical therapies. PMID- 11265864 TI - Database: relevant or not. AB - Progress in the field of ventricular assist devices requires a more rigorous and systematic method of collecting outcomes data. A worldwide registry of device implants and results is proposed. With widespread participation, data from this registry would improve the identification of risk factors and complications, and allow for the creation of predictive models that would enhance patient selection. Professional societies should lead the development of a registry in close partnership with government and industry. Data collection using the Web, with rigorous security measures to protect patient privacy, would offer numerous advantages in efficiency and immediacy of communication for all participants. PMID- 11265865 TI - Healing the heart with ventricular assist device therapy: mechanisms of cardiac recovery. AB - As experience has grown with the use of mechanical circulatory support systems in patients with cardiogenic shock, many anecdotes have been noted where myocardial recovery occurred and devices could be removed with reasonable residual cardiovascular performance and resolution of the shock syndrome. Indeed, when first used, ventricular assist devices were inserted to bridge patients unable to be separated from cardiopulmonary bypass to eventual recovery. Many successes with ventricular support systems have been recorded in individuals with postcardiotomy cardiogenic shock, acute myocarditis, and in the periinfarction period where stunning of potentially viable myocardial tissue contributed to severe heart failure. From an experimental standpoint, recovery of myocyte function and restoration of more normal myocardial geometry and constitution have been noted. There are many explanations for this, but principally, benefit is related to amelioration of circulatory insufficiency with attenuation of perturbed humoral networks and reduction of myocardial wall stress. It is important to understand how ventricular assist device implantation in select advanced heart failure patients might precipitate recovery of depressed myocardial function. PMID- 11265866 TI - Right heart failure: best treated by avoidance. AB - Right heart failure continues to affect our clinical success with left ventricular assist device support. The inability to consistently predict the probability of the onset of right heart dysfunction contributes to this problem. We have developed an aggressive approach to the management of these patients in an attempt to decrease the incidence of this condition, which continues to carry a very high operative mortality. PMID- 11265867 TI - Mechanical circulatory support for acute heart failure. AB - Circulatory support devices are frequently required in postcardiotomy shock, postmyocardial infarction shock, and acute myocarditis. A panel of cardiac surgeons addressed the use of these devices in 4 patients. Cardiogenic shock after mitral valve replacement was considered best served by a left ventricular assist device (VAD) with apical rather than atrial cannulation. A left VAD should be placed first and a right VAD only if needed. Acute myocardial infarction shock was considered best treated with a left VAD with left ventricular cannulation to avoid thrombosis. If cardiac transplantation is an option, a long-term device must be considered. Young patients with acute fulminant myocarditis should be implanted with VADs in anticipation of recovery, and transplantation should be delayed. Patients with severe heart failure after coronary bypass grafting were considered best served by an extracorporal membrane oxygenation (ECMO) system or a VAD. Current postcardiotomy survival rates of postcardiotomy patients of 20% to 40% are worthwhile, but can be improved. Temporary devices such as ECMO can be changed to more long-term devices when necessary. PMID- 11265868 TI - Postcardiotomy mechanical support: risk factors and outcomes. AB - BACKGROUND: The need for postcardiotomy mechanical support is uncommon, with an incidence of 0.5%. METHODS: Multivariable logistic regression analysis of factors associated with postcardiotomy extracorporeal membrane oxygenation (ECMO) support was investigated in 19,985 patients, of whom, 97 required ECMO. RESULTS: Younger age, number of reoperations, emergency operation, higher creatinine, greater left ventricular dysfunction, and history of myocardial infarction were significant predictors. Overall survival was 35%, but significantly better (72%) in the subgroup converted to an implantable system and then bridged to transplantation. CONCLUSIONS: Patients at increased risk for mechanical support can be identified preoperatively and patient management modified as indicated. Improvement in postcardiotomy survival has been realized by bridging to transplantation. In nontransplant candidates, permanent support may be the only option for increasing survival. PMID- 11265869 TI - Management of acute cardiac failure with mechanical assist: experience with the ABIOMED BVS 5000. AB - BACKGROUND: Mechanical circulatory assist industries have developed ventricular assist devices (VAD) for short-, intermediate-, and long-term use. The purpose of this report is to describe the progress made with the ABIOMED Biventricular System (BVS) 5000 (ABIOMED, Inc, Danvers, MA) short-term VAD. METHODS: From June 1994 through August 2000, all cardiogenic shock patients who required short-term mechanical assist were supported with the ABIOMED BVS 5000. Insertion criteria included any condition that may potentially result in cardiac recovery. A formal algorithm for timing of insertion was established to standardize implantation criteria. RESULTS: A total of 45 patients were supported at Hahnemann University Hospital, Philadelphia, PA. There were 26 male and 19 female patients, with a mean age of 57.9 years (range 33 to 80 years). Devices were inserted for postcardiotomy shock in 36 patients (80%) and precardiotomy shock in 9 patients (20%). The average duration of support was 8.3 days (range 1 to 31 days). Overall, there were 22 (49%) patients weaned from support and 14 (31%) discharged from the hospital. For patients in whom the device was implanted in accordance with an established protocol (group A), the wean and discharge rates were 60% and 43%, respectively. The most common morbidities included bleeding and adverse neurologic events. CONCLUSIONS: The ABIOMED BVS 5000 VAD continues to be a valuable form of short-term mechanical assist for acute cardiogenic shock. The formation of a uniform VAD insertion algorithm has helped to standardize protocols in management. PMID- 11265870 TI - Mechanical circulatory support for patients with acute-fulminant myocarditis. AB - This report provides a review of mechanical circulatory support for patients in cardiogenic shock secondary to acute/fulminant myocarditis. Experience and outcomes with extracorporeal membrane oxygenation, left ventricular assist device support (ABIOMED, Thoratec, Thermo Cardiosystems, Novacor), and biventricular ventricular assist device support (ABIOMED, Thoratec) are described. Patients in cardiogenic shock secondary to acute myocarditis in its fulminant presentation can recover, surprisingly with normal cardiac function. An aggressive approach to the use of mechanical support is strongly justified. Survival, either by bridge to transplant or recovery, should approach 70%. Transplantation can often be avoided. PMID- 11265871 TI - The use of extracorporeal life support in adult patients with primary cardiac failure as a bridge to implantable left ventricular assist device. AB - BACKGROUND: Extracorporeal life support (ECLS) is an effective technique for providing emergent circulatory assistance, and may represent a life-saving option in patients who might not initially be considered a candidate for other forms of circulatory support (extracorporeal or implantable left ventricular assist device [LVAD]). In the setting of cardiac arrest, ECLS represents the only viable method of initiating circulatory support. However, ECLS has a number of disadvantages that include high complication rates (eg, stroke, bleeding) and a limited duration of potential support, which have prevented its widespread acceptance, particularly in the adult population. With the increased successful application of long-term implantable LVADs as a bridge to transplant, the major limitation of ECLS could be overcome by bridging patients to a long-term implantable LVAD ("bridge to bridge"), thereby reducing the reluctance to utilize ECLS when indicated. After acquisition of the HeartMate LVAD (Thermo Cardiosystems, Inc, Woburn, MA) we investigated the use of ECLS as a bridge to an implantable LVAD and subsequent transplantation in selected high-risk patients. METHODS AND RESULTS: From Oct 1, 1996 to Sept 30, 2000, 33 adult patients presenting with cardiac arrest or severe hemodynamic instability were placed on ECLS for the bridge to bridge indication. Of the 33 patients, 10 patients survived to LVAD implant, 1 was bridged directly to transplant, 5 weaned from ECLS, and 16 died on ECLS. Overall, 12 patients survived to discharge. One-year actuarial survival from the initiation of ECLS was 36%. One-year actuarial survival from the time of LVAD implant, conditional on surviving ECLS, was 80%. CONCLUSIONS: The 1-year survival of adult patients placed on ECLS and who subsequently survived to an implantable LVAD was favorable. These data support a strategy of ECLS to implantable LVAD bridge to heart transplant in adult patients who are in need of circulatory support and who are not initially candidates for other forms of mechanical support. The favorable results of this strategy support utilization of ECLS even in situations where myocardial recovery is thought to be unlikely. PMID- 11265872 TI - Indications and patient selection for mechanical ventricular assistance. AB - Cardiac assist devices have become an important component of transplantation programs as they successfully bridge unsalvageable patients who would otherwise die. The indications for a device can still be classified into short-term and long-term situations. The short-term indications have expanded into areas such as postcardiotomy failure, high-risk cardiac operations, and acute myocardial infarction with results that were not previously possible in the absence of a well-established mechanical assistance program. Appropriate patient selection remains challenging and perhaps the most important attribute of a successful ventricular assist program. Although few exact criteria can define patients who are not eligible, several considerations and screening scales can help determine a particular patient's suitability. Specific attention must be given to right heart function, neurologic status, existing infections, renal function, and transplantation suitability. The future of this field will not only be in technological advances with devices but in optimization of patient selection and expanding indications such as permanent replacement therapy. PMID- 11265873 TI - Comparison of the CardioWest total artificial heart, the novacor left ventricular assist system and the thoratec ventricular assist system in bridge to transplantation. AB - BACKGROUND: Device selection has historically been supported by minimal comparative data. Since 1994, we have implanted 43 patients with the CardioWest Total Artificial Heart (CW), 23 with the Novacor Left Ventricular Assist System (N), and 26 with the Thoratec Ventricular Assist System (T). This experience provides a basis for our device selection criteria. METHODS: We reviewed retrospectively the results for survival, stroke, and infection in the CW, N, and T groups. Statistical methods included the Student's t-test, chi2 analysis, and Kaplan-Meier actuarial survival curves. RESULTS: The T group patients were younger and smaller sized than the CW or N group. The CW group had the highest mean central venous pressure (CVP) and lowest mean cardiac index. Survival to transplantation was 75% for CW, 57% for N, and 38% for T. Multiple organ failure postimplant caused most deaths in the CW and T groups. Right heart failure and stroke caused most N deaths. Linearized stroke rates (event/patient-month) were 0.03 for CW, 0.28 for N, and 0.08 for T. Serious infections were found in 20% of CW, 30% of N, and 8% of T patients, but linearized rates showed little difference and death from infection was rare. CONCLUSIONS: The N device should be used in "stable" patients with body surface area (BSA) greater than 1.7 m2 and with minimal right heart failure. Unstable patients with biventricular failure should receive a CW if the BSA is greater than 1.7 m2 or a T if they are smaller. PMID- 11265874 TI - Device and patient management in a bridge-to-transplant setting. AB - BACKGROUND: A variety of sophisticated devices have been developed for mechanical circulatory support in patients bridged to cardiac transplantation. Based on 13 years' experience, we have developed specific protocols for patient selection and management for different devices. METHODS: The principal systems applied in the bridge-to-transplant cohort are the Thoratec ventricular assist device (n = 144, mean duration of support 53 +/- 57 days), the Novacor left ventricular assist system (LVAS) (n = 85, mean duration of support 154 +/- 15 days), and the HeartMate LVAS (n = 54, mean duration of support 143 +/- 142 days). The Thoratec device is used for biventricular assistance or if the duration of support is expected to be less than 6 months. For long-term support, either the Novacor or HeartMate LVAS are preferred. RESULTS: Despite careful postoperative patient management, this group of patients is prone to a variety of complications. Bleeding occurred in 22% to 35%, right heart failure in 15% to 26%, neurologic disorders in 7% to 28%, infection in 7% to 30%, and liver failure in 11% to 20% of patients. Complications varied with the device applied and the patient's preoperative condition. A total of 73 patients were discharged from hospital for a mean period of 184 days; this cumulative experience amounted to 37.5 patient years. CONCLUSIONS: The Novacor and the HeartMate systems offer the additional possibility of discharging patients during support if they fulfill certain criteria. The main reasons for rehospitalization were thromboembolic and infectious complications. PMID- 11265875 TI - Quality improvement guidelines for transjugular intrahepatic portosystemic shunts. SCVIR Standards of Practice Committee. PMID- 11265876 TI - Quality improvement guidelines for percutaneous permanent inferior vena cava filter placement for the prevention of pulmonary embolism. SCVIR Standards of Practice Committee. PMID- 11265877 TI - Emergency interventional stroke therapy. PMID- 11265878 TI - Should interventional radiologists be involved in acute stroke intervention? PMID- 11265881 TI - Percutaneous AngioJet thrombectomy in the management of extensive deep venous thrombosis. AB - PURPOSE: This study was undertaken to evaluate the efficacy of a percutaneous mechanical thrombectomy (PMT) device for rapid thrombus removal following deep venous thrombosis (DVT). MATERIALS AND METHODS: Over a 37-month period, 17 patients (14 women; mean age, 41 y +/- 20) with extensive DVT were treated with initial attempts at PMT with use of the AngioJet rheolytic thrombectomy device. Sites of venous thrombosis included lower extremities in 14 patients and upper extremity and brachiocephalic veins in three. The etiology for venous thrombosis was malignancy in seven, idiopathic etiology in three, May-Thurner syndrome and immobilization in three each, and oral contraceptive use and hypercoagulable disorder in one each. The primary endpoint was venographic evidence of thrombus extraction. Perioperative complications, mortality, and recurrence-free survival were also evaluated. RESULTS: After PMT, four of 17 patients (24%) had venographic evidence of >90% thrombus removal, six of 17 (35%) had 50%-90% thrombus removal, and seven of 17 (41%) had <50% thrombus extraction. Adjunctive thrombolytic therapy was used in nine of 13 patients with <90% thrombus extraction by PMT; six patients (35%) had contraindications to pharmacologic thrombolytic therapy. An underlying lesion responsible for the occlusion was uncovered in 10 patients (59%). Significant improvement in clinical symptoms was seen in 14 of 17 patients (82%). No complications were noted directly relating to the use of the AngioJet thrombectomy catheter. None of the patients were lost to follow-up during a mean of 8.9 months +/- 5.3 (range, 2-21 months). At 4 and 11 months, recurrence-free survival rates were 81.6% and 51.8%, respectively. CONCLUSION: PMT with adjunctive thrombolytic therapy is a minimally invasive, low risk therapeutic option in patients with extensive DVT, associated with clinical benefits including thrombus removal, patency, and relief of symptoms. PMID- 11265879 TI - Interventional therapy for pulmonary embolism. AB - Venous thromboembolism is a common cause of death. Acute massive pulmonary embolism (PE) is life-threatening and may require vigorous more invasive treatment. Several risk factors are related to increased incidence of massive PE. Anticoagulation is the most traditional treatment for PE but may not suffice in cases of massive PE. Systemic thrombolytic therapy, catheter-directed thrombolysis, percutaneous embolectomy, and more recently, percutaneous thrombus fragmentation techniques with a multitude of devices are now available to treat the most severe cases of massive PE. Successful treatment of PE includes implementation of a treatment protocol and the use of associated techniques and devices. PMID- 11265882 TI - Sarcomas metastatic to the liver: response and survival after cisplatin, doxorubicin, mitomycin-C, Ethiodol, and polyvinyl alcohol chemoembolization. AB - PURPOSE: To evaluate the response to and survival after chemoembolization with cisplatin, doxorubicin, mitomycin-C, Ethiodol, and polyvinyl alcohol for patients with sarcomas metastatic to the liver that are surgically unresectable. MATERIALS AND METHODS: Sixteen patients were treated. Primary tumors included 11 gastrointestinal leiomyosarcomas, two splenic angiosarcomas, one leiomyosarcoma of the broad ligament, one leiomyosarcoma of the inferior vena cava, and one malignant fibrous histiocytoma of the colon. Chemoembolization with cisplatin, doxorubicin, mitomycin-C, Ethiodol, and polyvinyl alcohol particles was performed 1-5 times at approximately monthly intervals (mean, 2.8). Pre- and posttreatment cross-sectional imaging was performed 1 month after completion of treatment and then every 3 months. Thirty-day response was graded according to World Health Organization/Eastern Cooperative Oncology Group criteria. Survival was calculated with use of Kaplan-Meier analysis. RESULTS: Two patients (13%) exhibited partial morphologic response, 11 patients (69%) were morphologically stable, and three (19%) demonstrated progression of disease 30 days after completion of treatment. Among the 13 responders, two underwent partial hepatectomy after initial treatment. Seven developed intrahepatic progression at a mean of 10 months and a median time of 8 months. The remaining four patients had no documented intrahepatic progression at the time of last imaging follow-up. Nine patients developed extrahepatic progression at a mean time of 6.3 months and a median time of 6 months, of whom four underwent additional surgical resection. Response to therapy was based on time of first intervention. Cumulative survival from time of diagnosis with use of Kaplan-Meier analysis was 81% at 1 year, 54% at 2 years, and 40% at 3 years. Median survival time was 20 months. Cumulative survival from initial chemoembolization was 67% at 1 year, 50% at 2 years, and 40% at 3 years, with a median survival time of 13 months. The thirty-day mortality rate was zero. CONCLUSION: Durable tumor response with chemoembolization is possible in this form of metastatic disease, which is highly resistant to systemic chemotherapy. PMID- 11265883 TI - Arterial embolotherapy for upper gastrointestinal hemorrhage: outcome assessment. AB - PURPOSE: To identify predictors of clinical outcome after arterial embolotherapy for upper gastrointestinal (UGI) hemorrhage. MATERIALS AND METHODS: Seventy-five consecutive patients (mean age, 62.5 y) underwent arterial embolization for acute UGI hemorrhage. Bleeding was detected at endoscopy and angiography in 22 patients, at endoscopy alone in 29 patients, and at angiography alone in 24 patients. As such, embolization was directed by angiography in 46 patients (61.3%) and by endoscopy (referred to as "blind" embolization) in 29 patients (38.7%). The embolic agents used were metallic coils, polyvinyl alcohol particles (size range, 355-710 microm), gelatin sponge, and tissue adhesive. Predictors of bleeding recurrence and mortality were analyzed with logistic regression and Cox models, respectively. RESULTS: The technical success rate of embolization was 98.7%. Primary clinical success was achieved in 57 patients (76%). Secondary clinical success occurred in five additional patients (82.5%) after repeat embolization. There were four (5.3%) complications: two cases of self-resolving duodenal ischemia, one hepatic infarct, and one inguinal hematoma. The periprocedural mortality rate was 34.6% (26 of 75), mostly related to underlying illness. Early recurrence of bleeding (within 30 days of embolization) was associated with coagulation disorders (international normalized ratio >1.5, partial thromboplastin time >45 seconds, or platelet count <80,000/microL; odds ratio, 19.46; P = .001) and with the use of coils as the only embolic agent (odds ratio, 7.73; P = .01). Cirrhosis and cancer shortened the overall survival of patients after embolic therapy. The mean patient follow-up time was 34.5 months. CONCLUSION: Arterial embolotherapy for UGI hemorrhage is safe, effective, and durable. Coagulopathy and the use of coils as the only embolic agent were associated with a higher risk of early bleeding recurrence. PMID- 11265884 TI - Treatment of malignant pleural effusions with tunneled long-term drainage catheters. AB - PURPOSE: To assess the effectiveness of tunneled pleural catheters (TPCs) in the treatment of malignant pleural effusions (MPEs). MATERIALS AND METHODS: Twenty eight patients with symptomatic MPEs had 31 hemithoraces treated with TPCs placed under image guidance. Chemical sclerotherapy had failed in two patients and two had symptomatic locules. Drainage was accomplished by intermittent connection to vacuum bottles. Pleurodesis was considered achieved when three consecutive outputs were scant and imaging showed no residual fluid. RESULTS: All catheters were successfully placed. Dyspnea improved in 94% (29 of 31 hemithoraces) at 48 hours and 91% (20 of 22 patients) at 30 days. Control of the MPE was achieved in 90% of hemithoraces (28/31), although five required ancillary procedures. Pleurodesis occurred in 42% (13 of 31) of hemithoraces, including both that underwent an earlier attempt at chemical sclerotherapy and one treated locule. Continued drainage without pleurodesis controlled the effusion in 48% (15 of 31). In only 7% was hospital time necessary for care related to the TPC. Early, transient catheter-related pain was common, but only three complications (in two patients) occurred. Neither of these altered patient care. CONCLUSIONS: Regardless of whether pleurodesis is achieved, TPCs provide effective long-term outpatient palliation of MPEs. PMID- 11265885 TI - CT findings after embolization for blunt splenic trauma. AB - PURPOSE: To determine complications after transcatheter embolization for blunt splenic injury as recognized with computed tomography (CT). MATERIALS AND METHODS: From March 1997 to January 2000, 80 patients underwent transcatheter embolization after blunt splenic injury, of whom 53 underwent abdominal CT examination before and after embolization. Preembolization CT scans were reviewed to determine grade of injury, and postembolization CT scans were reviewed to identify complications secondary to embolization. Arteriography results were reviewed to determine findings and method and location of embolization. RESULTS: Splenic infarcts occurred in 63% of patients after proximal embolization and in 100% of patients after distal embolization. Infarcts after distal embolization tend to be larger and occur just distal to the embolization material, whereas infarcts after proximal embolization tend to be smaller, multiple, and located in the periphery. Most infarcts resolved without sequelae. Seven patients developed gas within an infarct or subcapsular fluid collection. Two collections were drained and found to be sterile and one patient had a splenic abscess at laparotomy. CONCLUSIONS: Infarcts are common after splenic embolization. Gas may be present within an infarct after embolization with Gelfoam; however, the presence of air/fluid level is a better predictor of abscess. PMID- 11265886 TI - Biopsy of lung nodules with use of I-I device under intermittent CT fluoroscopic guidance: preliminary clinical study. AB - PURPOSE: To investigate the efficacy of computed tomography (CT) fluoroscopy and a new needle holder (the I-I device) in lung nodule biopsy. MATERIALS AND METHODS: The I-I device is made of acrylate resin and was used to keep the entire needle in the tomographic plane. This study consisted of biopsies of 79 lung nodules in 77 patients. The final diagnoses were malignant in 54 patients, benign in 23, and unconfirmed in two. The biopsy procedure time from the beginning of the CT fluoroscopy procedure to the removal of the needle was measured for 24 needle passes. The radiation dose on the physician's hand was measured in five cases with use of a thermoluminescence ring. RESULTS: Fifty-one malignant and 20 benign lesions were correctly diagnosed with histologic specimens (90%). In 58 of 77 patients (75%), the biopsy procedures were completed within a single breath hold. Pneumothorax occurred in 20 of 77 patients (26%) and chest tube insertion was required in five. The incidence of pneumothorax was significantly lower in patients who held their breath during biopsy procedures compared with those who did not (P < .0001; chi2 test). The biopsy procedure time ranged from 15 to 39 seconds (mean: 28.2 sec). The mean radiation dose on the physician's hand was 2 mSv/case. CONCLUSION: The diagnostic accuracy of biopsy with use of the I-I device under CT fluoroscopic guidance is comparable with that of the conventional method; however, a combination of CT fluoroscopy and the I-I device enables rapid biopsy procedures. PMID- 11265887 TI - Treatment of infrapopliteal occlusive disease by high-speed rotational atherectomy: initial and mid-term results. AB - PURPOSE: To assess the effectiveness and patency rates of high-speed rotational atherectomy (HSRA) for the treatment of infrapopliteal arterial occlusive disease. MATERIAL AND METHODS: During an 18-month period, a total of 19 infrapopliteal lesions in 15 consecutive patients were treated primarily by HSRA with use of the Rotablator device. Patients were followed up with documentation of clinical symptoms, standardized treadmill exercise, and Doppler sonography at 1, 3, and 6 months. Control angiography was performed 6 months after primary treatment. RESULTS: HSRA was initially successful in 14 of 15 patients, yielding an initial technical success rate of 94%. Percutaneous treatment induced an improvement of the ankle-brachial index (ABI) from 0.6 +/- 0.09 to 0.86 +/- 0.2 after intervention (P < .0001). Doppler analysis showed a mean ABI of 0.85 +/- 0.2 (P < .001) at 1 month, 0.72 +/- 0.2 (P = .012) at 3 months, and 0.7 +/- 0.2 (P = .08) at 6 months after initial therapy. Although six patients were lost to follow-up at various times, control angiography at 6 months was carried out in nine of 15 patients, allowing direct assessment of 12 of 19 treated lesions. Among six high-grade restenoses and five total occlusions in the treated vascular segments, only one arterial lumen (of 12) remained patent without presenting a hemodynamically relevant restenosis. These results led to termination of the study. CONCLUSION: Although HSRA for the treatment of infrapopliteal occlusive disease yields a very high initial technical success rate, mid-term results are extremely poor. Therefore, HSRA cannot be recommended for primary treatment of this type of lesion. PMID- 11265888 TI - Short-term patency and safety of an expanded polytetrafluoroethylene encapsulated endoluminal device at the venous anastomosis of a canine arteriovenous graft model. AB - PURPOSE: To determine the safety and short-term patency of a polytetrafluoroethylene (PTFE)-encapsulated carbon-lined endoluminal device (ED) deployed across the venous anastomosis of arteriovenous conduits. MATERIALS AND METHODS: Arteriovenous grafts (n = 16) were created between femoral arteries and veins in eight female canines and allowed to mature 30 days +/- 5 (SD). Five were excluded before implantation because of thrombosis or intragraft stenosis. Deployment was conducted in the remaining 11 anastomoses. Fistulography and intravascular ultrasound (IVUS) were performed before and after the procedure and 1 month postimplantation. Stent migration, apposition, and stenosis were evaluated. The angle of the anastomosis was compared before and after deployment and at follow-up. Mural thrombus thickness, percentage of surface covering, and percentage of endothelialization within the device were measured histologically. RESULTS: There was no significant migration. By explant, all devices were completely apposed. Stenosis occurred in three of nine grafts. The angle of the venous anastomosis decreased by 29.5 degrees (posteroanterior) and 32.4 degrees (oblique) after ED deployment. There was a further decrease of 6.1 degrees (posteroanterior) and 3.2 degrees (oblique) during the 4-week follow-up period. Hemostasis was difficult to achieve in this animal model. Five required more than 1 hour to achieve hemostasis manually. Six in three animals were closed with a Perclose device, achieving immediate hemostasis; however, three (one in each animal) re-bled intermittently 2 weeks after implantation for an average of 9.3 days. The puncture site of each graft that bled was radiographically shown abnormal. CONCLUSION: The ED can be deployed without stent migration and is completely apposed and patent after 4 weeks. Although bleeding was a problem with this animal model, delayed bleeding complications associated with puncture site abnormalities were seen only in grafts closed with a percutaneous suturing device. PMID- 11265889 TI - Effects of intraarterial thrombin in the swine model. AB - PURPOSE: To evaluate the effects of catheter-directed thrombin in the peripheral arterial circulation of swine. MATERIALS AND METHODS: Thrombin was injected into a single femoral artery in 20 domestic swine. Each of five animals from four dose groups received 50, 150, 250, or 1,000 U as a single dose. Bilateral femoral arterial flow was monitored for as long as 4 hours and evaluated relative to baseline and contralateral limb flow. Interval arteriographic results were evaluated by segmental patency and a numeric angiographic score. RESULTS: Mean baseline flow was 136 mL/min +/- 44, with an internal arterial diameter of 3.4 mm +/- 0.5. A transient increase in blood flow after thrombin administration was followed by diminished flow and thrombosis. These findings varied directly with dose and inversely with baseline flow. Angiographic and flow abnormalities generally improved with time and recovery was generally better in swine that received 50 or 1,000 U than in other groups. However, one animal that received 1,000 U (13.2 U/mL/min) developed stable, complete limb thrombosis. The degree of recovery varied with thrombin dose and thrombus location. At doses greater than 50 U (0.33 U/mL/min +/- 0.05), abnormalities were commonly persistent. Animals receiving the 150-U dose (1.33 U/mL/min +/- 0.41) had a higher incidence of persistent distal occlusion. Distal occlusions were less likely to resolve than proximal occlusions. CONCLUSIONS: The effect of intraarterial thrombin is directly related to dose and inversely related to baseline blood flow. In swine, a threshold for significant flow disruption and thrombosis exists above a dose of 50 U (0.33 U/mL/min +/- 0.05). A threshold dose for irreversible occlusion may also exist. Although small amounts of thrombin in a high-flow vessel may not cause significant complication, administration into the arterial circulation should be avoided. PMID- 11265891 TI - Fluoroscopically guided Norplant removal. AB - The Norplant contraceptive implant system is a commonly used method of contraception worldwide. Implant placement and removal are usually simple office based outpatient procedures. Norplant removal can occasionally become difficult, usually secondary to improper insertion. In these instances, we describe a method of Norplant removal that can easily be performed with use of high-resolution fluoroscopy with associated digital subtraction imaging. PMID- 11265890 TI - Daily catheter-directed single dosing of t-PA in treatment of acute deep venous thrombosis of the lower extremity. AB - The strong fibrin affinity of recombinant tissue plasminogen activator (rt-PA) theoretically obviates continuous infusion or replacement of t-PA after direct intrathrombic injection. This hypothesis led the authors to evaluate single daily catheter-directed injection of rt-PA as a thrombolytic treatment for acute deep vein thrombosis of the lower extremity. Once-daily injection of rt-PA was performed in large thrombosed veins (popliteal or larger) with use of pulse-spray catheters and in small thrombosed veins in patients' calves with use of 3-4-F coaxial catheters. Patients received only full systemic anticoagulation on his/her patient care unit. This dosing regimen has been tested in 10 patients (12 legs) with a maximum dose of 50 mg per leg per day. Extensive thrombolysis was achieved in nine patients and partial thrombolysis was achieved in one patient, at an average total dose of 106 mg of rt-PA per leg. Minor bleeding was seen in three patients and no transfusions were needed. Our technique and the rationale for this pilot study is the focus of this article. PMID- 11265892 TI - Transnasal removal of a biliary endoprosthesis with percutaneous transhepatic biliary catheter replacement. AB - The authors describe a novel fluoroscopically guided transnasal technique for removal of an endoscopically placed endobiliary prosthesis (EBP). A case in which the approach was applied is used to demonstrate its indications, clinical utility and possible benefits. The procedure entails simultaneous percutaneous replacement of the EBP with an internal/external biliary catheter over a through and-through transhepatic/transnasal access wire. A comparison of the technique's limitations with those of traditional approaches for endoscopic or percutaneous EBP removal and replacement is presented. PMID- 11265893 TI - Radiofrequency thermal ablation of a splenic metastasis. AB - Effective local ablation of large tumors with radiofrequency has been made possible by recent advancements. Tumor ablation with radiofrequency has been described mainly in the liver, but also recently in the kidney, adrenal gland, lung, and breast. A rapidly growing splenic metastasis from renal cell carcinoma was effectively treated percutaneously, with US guidance. Focal splenic disease may not be a common indication for ablation; however, further work is necessary to evaluate the safety and efficacy of this procedure in this setting. PMID- 11265894 TI - Crystallization when mixing contrast materials with ethanol for embolization of venous malformations. PMID- 11265895 TI - Re: augmented experimental pulse-spray thrombolysis with tissue plasminogen activator, enabling dose reduction by one or more orders of magnitude. PMID- 11265896 TI - A simple wire technique to correct malfunctioning permacaths. PMID- 11265897 TI - Tissue phantom for learning US-guided vascular punctures. PMID- 11265898 TI - Delayed diagnosis in pediatric blunt trauma. AB - OBJECTIVE: Identification of injuries of a traumatized patient is a mandate for the emergency department (ED) and the trauma team. Delayed diagnosis of injury in trauma patients leads to increased morbidity, mortality, dissatisfaction, and risk of litigation. Comparing children admitted for blunt trauma, with and without delay, this study examines risk factors for delayed diagnosis. METHODS: Delays in diagnosis from 1991 to 1996 were identified during prospective collection of trauma registry data. Controls were randomly selected from the trauma registry. Charts from both groups were retrospectively reviewed. RESULTS: Fifty-eight patients had 65 delays in diagnosis. Significant independent delay variables included: female, motor vehicle crash (MVC)-related mechanism, altered consciousness, higher injury severity score, and multiple injuries (P < 0.05). Trauma team activation, documentation of tertiary survey, and length of hospitalization were greater in patients with delay injuries (P < 0.05). Logistic regression identified MVC-related mechanism, female, facial, and extremity injuries as a combination of predictors. CONCLUSIONS: Delays occurred in 1% of patients. Trauma team care itself did not protect all patients from delay. Injury severity at presentation alone is not an adequate predictor of delayed diagnosis in the pediatric patient. A combination of variables was identified as negative predictors of delay. Further study is needed to validate these criteria, and determine if earlier diagnosis would effect quality. PMID- 11265899 TI - Accuracy of visual determination of neutral position of the immobilized pediatric cervical spine. AB - BACKGROUND: The definition of neutral position for the immobilized pediatric cervical spine is not well standardized. In this study, we attempted to determine whether 1) physicians and/or paramedics could accurately assess visually if the cervical spine was in a neutral position, 2) the visual assessments of the observers were in agreement, and 3) a radiographic Cobb angle would correlate with the visual determination. METHODS: Children presenting to a pediatric emergency department (ED) in full spinal immobilization were randomly selected (convenience sample) for this prospective study. The emergency physician and transporting paramedic independently determined positioning of the cervical spine. A radiologist, blinded to clinical information, determined Cobb angles from radiographs of the immobilized cervical spines. RESULTS: Of the 59 children studied, the evaluation of cervical spine position by the physician and paramedic correlated in 88% of the cases. For the 22 children with non-neutral Cobb angles (definition of neutral: between 5 degrees flexion and 5 degrees extension), observers agreed in 100% of the cases. However, in 21 of these cases (95%) the position was observed as neutral. CONCLUSIONS: Although visual determinations of neutral position of the cervical spine by two observers may correlate, radiographic studies demonstrate that neutral position was not achieved in 37% of the cases. PMID- 11265900 TI - The clinical presentation of pediatric pelvic fractures. AB - BACKGROUND: Few studies have addressed the presentation and clinical impact of pediatric pelvic fractures. We sought to describe pediatric blunt trauma patients with pelvic fracture (PF) and to evaluate the sensitivity and specificity of physical examination at presentation for diagnosis. METHODS: Retrospective analysis of all PF and control (NPF) patients from our pediatric institution over an 8-year period. RESULTS: A total of 174 patients (88 PF, 86 NPF) were included. Median patient age was 8 years (range, 3 months to 18 years), with 54% males. The most common mechanisms of injury for PF patients were automobile-related accidents (75%). There were 140 patients (87%) who were transported by air or ground medical services. At presentation, approximately 16% of PF patients had a Glasgow Coma score of <15, a mean Revised Trauma Score of 7.49, and a median Injury Severity Score (ISS) of 9. Thirty-one PF patients (35%) had an ISS of >15 indicating severe, multiple injuries. Sixty-eight PF patients (77%) had severe isolated injuries (Abbreviated Injury Scale 1990 value of >3); 11% of PF patients required transfusions, and 2% died. Fifteen PF patients (17% ) had no pelvic ring disruption; 39 (43%) had a single pelvic ring fracture, 22 (2%) had two pelvic ring fractures, 2 (2%) had acetabular fractures, and 10 (11%) had a combination of pelvic fractures. An abnormal physical examination of the pelvis was noted in 81 patients with PF (92% sensitivity, 95% confidence interval [CI] = 0.89-0.95), 15 NPF patients had an abnormal examination (79% specificity, 95% CI = 0.74 0.84). The positive predictive value of the pelvis examination was 0.84, and the negative predictive value was 0.89. The most common abnormal pelvis examination finding was pelvic tenderness in 65 PF patients (73%). A total of seven PF patients had a normal examination of the pelvis; four had a depressed level of consciousness (defined as GCS <15), and six patients had a distracting injury. CONCLUSIONS: Pediatric blunt trauma patients with pelvic fracture represent a severely injured population but generally have lower transfusion rates and mortality than noted in adult studies. The pelvis examination appears to be sensitive and specific in this retrospective study. However, an altered level of consciousness and/or distracting injuries may affect examination sensitivity and specificity. Based on this retrospective study, we cannot advocate eliminating pelvic radiographs in the severely injured, blunt trauma patient. Prospective studies are recommended. PMID- 11265901 TI - Child literacy promotion in the emergency department. AB - INTRODUCTION: Studies using primary health care settings to promote literacy have demonstrated success. Since socioeconomically disadvantaged children have less access to primary care, obtaining much of their medical care episodically in emergency departments (ED), the purpose of this study is to investigate the effectiveness of a simple literacy promotion program conducted in the ED and to determine its efficacy and if there is a difference in promoting literacy with a brochure alone versus a brochure plus a children's book. METHODS: Medical student study investigators enrolled patients aged 20 months to 7 years undergoing acute care treatment and evaluation in an ED. Subjects were randomized to a literacy promotion brochure versus a brochure plus a children's book. Phone follow-up interviews were conducted to survey the degree of parental reading taking place at home. RESULTS: Fifty-one families were enrolled, 8 could not be contacted for follow-up, which left 43 families. Twenty-eight were randomized to book + handout and 15 were randomized to handout alone. On follow-up phone interviews, no significant change in reading was demonstrated regardless of whether a brochure alone was given or a book was given with the brochure. CONCLUSION: Literacy promotion in the ED was not associated with any measurable improvement in parent child reading. This could mean that the ED is not an effective place to promote literacy or confounding factors in this study are responsible for failing to demonstrate a measurable benefit. PMID- 11265902 TI - Should parents accompany pediatric interfacility ground ambulance transports? Results of a national survey of pediatric transport team managers. PMID- 11265904 TI - Traumatic posterior dislocation of hip in children. AB - Traumatic posterior dislocation of the hip joint in children is an uncommon injury. It constitutes a true orthopedic emergency. It makes up over 80% of pediatric hip dislocations. In children, it can occur as a result of minimal trauma, which is attributed to a soft pliable acetabulum and ligamentous laxity. In skeletally mature adolescents, a greater force is required to dislocate the hip joint. Delay in reduction is associated with long-term complications such as avascular necrosis and degenerative arthritis. Avascular necrosis is related to the duration of dislocation. A poorer prognosis is associated with delay in reduction beyond 6 hours, advanced skeletal maturity, or multiple traumas. Prompt reduction minimizes complications. We report two cases of traumatic posterior dislocation of hip in children aged 3 and 14 years. Both were reduced within 6 hours of dislocation, and review at 6 months revealed normal examination and no evidence of any post-traumatic changes. Post-reduction treatment remains without a consensus. This review highlights the clinical presentation, management, and time-sensitive complications of the injury. PMID- 11265903 TI - Metal lawn and garden edging: the hidden knife? AB - OBJECTIVE: Lacerations account for many visits to the pediatric emergency department. We observed children presenting to local emergency departments in a large metropolitan area with lacerations incurred from metal lawn and garden edging, a landscaping tool. We sought to describe the severity of lacerations caused by metal edging, the characteristics of wound repair, and the need for subspecialty consultation. DESIGN: A retrospective chart review including all pediatric patients (< 18 years) presenting with lacerations caused by metal lawn and garden edging from January 1995 to October 1997 was performed. Patients were seen at one of three emergency departments in Colorado. RESULTS: One hundred twenty-six patients were enrolled (76% male, 24% female), with a median age of 9 years. The most frequent location of laceration was the foot (40%), followed by the knee (26%). The median length of laceration was 3 cm (range 1-22 cm). Sixteen patients (13%) received either intravenous or oral antibiotics, and six patients (5%) received orthopedic evaluation. CONCLUSIONS: Metal lawn and garden edging in landscaped neighborhoods presents a previously undescribed laceration danger to children. Some lacerations sustained from the metal lawn edging are extensive, receiving either multiple layer closure and/or the need for subspecialty consultation. PMID- 11265905 TI - Oral and airway sequelae after hair relaxer ingestion. PMID- 11265906 TI - Mucus plugging as a cause of acute lobar overdistension. AB - Acute lobar overdistension in children is usually indicative of foreign body aspiration, especially when a history of a choking episode is recalled. An unusual presentation of asthma as recurrent overdistension of the left lung in a toddler is described. The child had undergone two consecutive negative bronchoscopies with a presumptive diagnosis of foreign body aspiration; however, it was only the antiasthmatic treatment that resulted in an excellent outcome. PMID- 11265907 TI - A traumatic spinal epidural hematoma in an infant with hemophilia A. PMID- 11265909 TI - Fever education: does it reduce parent fever anxiety? AB - OBJECTIVE: The aims of the study were to determine the following: 1) if a fever education program (interactive or written) reduces parent fever anxiety; 2) if an interactive fever program was more effective as a teaching style than standard written material alone; and 3) if a fever program increases parent fever home management and reduces return emergency department (ED) visits. METHOD: A quasiexperimental, pretest and post-test pilot study examining parental fever anxiety was conducted at The Children's Hospital of Philadelphia. Eligible participants consisted of 87 parents and their children, aged 3 months to 5 years presenting with fever >38.4 degrees C, and without coexisting serious illness. RESULTS: Both the interactive fever education program and the standard written fever pamphlet were equally effective as teaching methods. Data revealed a 30% reduction in fever anxiety rated as moderate-severe on arrival to none-low post fever education, increased parent fever home management skills with correct use of thermometer and antipyretics, and reduced unnecessary return ED visits. CONCLUSION: Parents in the acute and nonacute care setting may benefit from an interactive fever education program that includes the definition and benefit of fever, the correct use of a thermometer, fever home management skills, and appropriate fever telephone follow-up. PMID- 11265910 TI - Pediatric pre-hospital advanced life support care in an urban setting. AB - OBJECTIVE: To describe pediatric advanced life support (PALS) in a single urban environment and clarify educational priorities for ALS pre-hospital providers and pediatric medical control physicians. METHODS: Retrospective observational review of all pediatric pre-hospital PALS transport and medical control records of the two-tiered, unified, municipal emergency medical service of the City of Boston (catchment area 590,000) over a 1-year period. RESULTS: Of the 555 pediatric patients receiving ALS transport, 38% were for respiratory emergencies, 24% for nonrespiratory medical emergencies, 19% for traffic-related blunt trauma, and 10% for penetrating trauma. Two percent involved cardiac arrests. The most frequent procedures performed were intravenous (IV) cannulation (n = 184, 33%), bag-mask ventilation (n = 28, 5%) and intubation (n = 15, 3%). Intraosseous access was only performed in three patients (0.5%). Fifty ALS providers in the EMS system averaged pediatric IV cannulation 3.7 times, intubation 0.3 times, and intraosseous access 0.06 times per provider per year. On-line medical control was requested in 28 % of PALS transports. The chief complaints managed by medical control closely mirrored the distribution of all ALS transports. The most frequent medication ordered by on-line medical control was additional nebulized albuterol after standing orders (off-line medical control) had been exhausted. CONCLUSIONS: A limited number of chief complaints make up the majority of PALS transports. Initial and continuing education for ALS providers needs to reflect the importance of these critical entities. Education for urban pre-hospital providers should reflect that certain procedures will be only executed every few years (eg, pediatric intubation) or once in the career of an ALS pre-hospital provider (eg, intraosseous access). With a limited amount of pediatric teaching time, paramedic education will have to strike a careful balance between teaching about the chief complaints most frequently encountered and teaching rare, high risk procedures that could provide maximal support for the uncommon critically ill child. On-line medical control physicians need to be prepared to direct and support the management by ALS pre-hospital providers for the chief complaints most frequently seen in pediatric patients. PMID- 11265911 TI - Trauma to wrist. PMID- 11265908 TI - Intracranial hemorrhage at the onset of thrombotic thrombocytopenic purpura in an infant: therapeutic approach and intensive care management. AB - Thrombotic thrombocytopenic purpura (TTP) is quite rare in infancy and must be treated intensively as a life-threatening disease. Diffuse vascular thromboses may occur, and neurologic involvement is a cornerstone of the diagnosis of TTP. We describe a case of an infant who presented with a sudden cerebral hemorrhage and subsequently developed the typical clinical features of TTP. Emergency treatment in the Pediatric Intensive Care Unit (PICU) consisted of plasma therapy and exchange-transfusion (EXT) to arrest the intravascular process and the exsanguinating blood loss. Exchange-transfusion is a life-saving procedure that is rarely performed after the neonatal age. PMID- 11265912 TI - Percutaneous endoscopic gastrostomy or skin-level gastrostomy tube replacement. PMID- 11265913 TI - Differentiating osteomyelitis from bone infarction in sickle cell disease. AB - This brief review discusses one possible approach to evaluating the sickle cell patient with bone pain. The major differential diagnoses include osteomyelitis and bone infarction. Based on previous studies, we provide an approach to assessing and treating patients with the possible diagnosis of osteomyelitis. An algorithm has been provided, which emphasizes the importance of the initial history and physical examination. Specific radiographic studies are recommended to aid in making the initial assessment and to determine whether the patient has an infarct or osteomyelitis. Differentiating osteomyelitis from infarction in sickle cell patients remains a challenge for the pediatrician. This algorithm can be used as a guide for physicians who evaluate such patients in the acute care setting. PMID- 11265914 TI - Pediatric emergency medicine: legal briefs. PMID- 11265915 TI - Tangential transplant issues. PMID- 11265916 TI - I'll have my N-acetylcysteine with an orange twist. PMID- 11265917 TI - Mediators of behavioral problems in 7-year-old children born after 24 to 28 weeks of gestation. AB - We tested the hypothesis that prematurity acts through its association with neuromotor and intellectual functioning to explain behavior problems at school age. Sixty-one extremely preterm (EP) very low birth weight (VLBW) children (< 29 wk and < 1,500 g) born in 1987-1990 and 44 normal birth weight children (NBW) (> 37 wk and > 2,500 g) were matched for age, sex, and socioeconomic status (SES). Mediator variables were evaluated at a hospital at 5 years and 9 months. Behaviors were evaluated at school at 7 years by peers, teachers, and parents. When compared with NBW children, EP/VLBW children had poorer IQ and neuromotor development. At school, EP/VLBW children were evaluated by peers as more sensitive/ isolated, and by teachers and parents as more inattentive and hyperactive than NBW. When mediators were introduced, the previously significant relation between prematurity and behavior problems disappeared. Hyperactive and inattentive behaviors were explained by a specific working memory factor for the latter, and by a general intellectual delay for the former, whereas sensitive/isolated behaviors were best explained by neuromotor delays. Inattentive behaviors were also related to family adversity. At school age, extreme prematurity had thus an indirect effect on behaviors via specific and nonspecific intellectual and neuromotor delays. PMID- 11265918 TI - Cognition, academic progress, behavior and self-concept at 14 years of very low birth weight children. AB - The aim of this study was to compare cognition, academic progress, behavior, and self-concept children of very low birth weight (VLBW, birth weight < 1501 g) born in the period 1980 to 1982 with randomly selected children of normal birth weight (NBW, birth weight > 2,499 g). At 14 years of age, 130 (84.4%) of 154 VLBW and 42 (70.0%) of 60 NBW children were assessed. Ten VLBW children and one NBW child who had cerebral palsy were excluded. VLBW children scored at a significantly lower level on all three composite scales of the Wechsler Intelligence Scale for Children, 3rd Edition. VLBW children were also significantly disadvantaged on more specific cognitive processes, including tests of visual processing and visual memory and on subtests reflecting learning and problem solving. Only in arithmetic was a difference between the groups discerned on tests of achievement. Significantly more VLBW children were rated by teachers as socially rejected and by their parents as having learning problems at school. VLBW children had significantly reduced self-esteem. VLBW children had more cognitive, academic, and behavioral problems and lower self-esteem at 14 years of age than NBW control subjects. PMID- 11265919 TI - Preschool language outcomes of children with history of bronchopulmonary dysplasia and very low birth weight. AB - A prospective follow-up of very low birth weight (VLBW) infants with and without bronchopulmonary dysplasia (BPD) and term control infants was conducted. The effects of BPD and VLBW on speech-language development and specific language impairment at 3 years of age were investigated, controlling for the effects of sociodemographic and other medical risk factors. Groups were compared on cognitive and speech-language outcomes using the Battelle Language and Bayley Mental Scales of Infant Development. Children with a history of BPD had lower receptive language skills than VLBW children without BPD, who in turn had lower receptive skills than term children. Children with a history of BPD also had lower expressive skills than the two comparison groups, whereas VLBW children without BPD did not differ in expressive language from term children. When IQ score was controlled, children with BPD demonstrated specific language impairment in receptive language. The presence of patent ductus arteriosis (PDA) was the best predictor of language deficits and the combined occurrence of PDA and BPD resulted in differentially lower language scores. Neurologic complications, low socioeconomic status, and minority race were also significant predictors of language delay. The findings emphasize the importance of considering both medical and sociodemographic factors in evaluating the risk of VLBW infants for poorer speech-language outcomes. PMID- 11265921 TI - An opportunity for office-based research. AB - Robert, a nearly 12-year-old boy, traveled an hour to see a new pediatrician. Robert's mom told the pediatrician that Robert had not been seen by a doctor for several years because "no one seems to be able to help him with his problem." Robert had been wetting the bed "ever since he was toilet-trained" at age 2 years. Robert wets the bed about 5 out of 7 nights. He never has daytime accidents. He did not have a history of urinary tract infection, dysuria, urgency, or increased frequency of urination. He has daily bowel movements and denied soiling or accidents. Robert's mom said he had "toilet-trained himself" at age 2 years. Both Robert's mom and maternal grandfather had nocturnal enuresis "into their teenage years." The pediatrician was surprised to learn that another physician had treated Robert with imipramine at age 5 years. The medication worked intermittently and Robert continued to take it for about a year. At age 6 years, Robert's parents saw an advertisement for a bed-wetting alarm. They purchased the alarm but found that Robert never woke up when the alarm sounded. At age 7 years, Robert saw a urologist who told him he would "outgrow the problem." A year later, the urologist prescribed desmopressin acetate (DDAVP) nasal spray, which Robert took on occasion during the next 2 years. Every time he stopped the DDAVP, he resumed wetting the bed. His parents never punished him for his accidents, but they did try restricting fluids after dinner and also woke Robert in the middle of the night and encouraged him to go to the bathroom. Neither of these strategies was successful. Robert said he was "frustrated" and wondered if "I would still be wetting the bed as a grown-up." The pediatrician explained the nature of enuresis to Robert and his mom, provided them with instructions and an order form for a bed-wetting alarm, and arranged a follow-up visit. The next day, during nursery rounds, he asked several of his colleagues about their approaches to the treatment of enuresis. A few used DDAVP, one found imipramine beneficial, and one preferred behavioral treatment with a bed-wetting alarm. The pediatrician became concerned that he had misread the literature on enuresis. He brought the question up at the next pediatric staff meeting at the local hospital. A lively discussion ensued as the physicians realized that they employed a variety of treatments for enuresis. Robert's pediatrician wondered why his colleagues were not using the alarm because the literature seemed to indicate it to be the preferred treatment for enuresis. He asked the group if they would be interested in talking about the issue further and perhaps trying to understand the reasons for their varied approaches to this problem. PMID- 11265920 TI - A prospective comparison of developmental outcome of children with in utero cocaine exposure and controls using the Battelle Developmental Inventory. AB - Children with in utero cocaine exposure may be at risk for adverse neurodevelopmental outcome. To evaluate such outcome in young children, we administered the Battelle Developmental Inventory (BDI) to a group of inner-city children with (COC) and without (CON) in utero cocaine exposure at ages 3 and 5 years. Sixty-five COC and 68 CON, similar at age of testing, were evaluated at both time points by examiners masked to child group status. Both groups scored poorly and worsened over time. Although Total BDI raw scores were lower in the COC group than in the CON group at 3 years, this difference was related to postnatal environmental factors, caregiver (p = .022), and home environment (p = .010), not to in utero cocaine exposure (p = .88). At 5 years, the Total BDI score was related to the home environment (p < .001) but not to the caregiver (p = .36) or in utero cocaine exposure (p = .83). We conclude that inner-city children are at risk for adverse developmental outcome regardless of in utero cocaine exposure. PMID- 11265922 TI - Down syndrome: advances in molecular biology and the neurosciences. AB - The entire DNA sequence for human chromosome 21 is now complete, and it is predicted to contain only about 225 genes, which is approximately three-fold fewer than the number initially predicted just 10 years ago. Despite this remarkable achievement, very little is known about the mechanism(s) whereby increased gene copy number (gene dosage) results in the characteristic phenotype of Down syndrome. Although many of the phenotypic traits show large individual variation, neuromotor dysfunction and cognitive and language impairment are observed in virtually all individuals. Currently, there are no efficacious biomedical treatments for these central nervous system-associated impairments. To develop novel therapeutic strategies, the effects of gene dosage imbalance need to be understood within the framework of those critical biological events that regulate brain organization and function. PMID- 11265923 TI - Findings from the NIMH Multimodal Treatment Study of ADHD (MTA): implications and applications for primary care providers. AB - In 1992, the National Institute of Mental Health and 6 teams of investigators began a multisite clinical trial, the Multimodal Treatment of Attention-Deficit Hyperactivity Disorder (MTA) study. Five hundred seventy-nine children were randomly assigned to either routine community care (CC) or one of three study delivered treatments, all lasting 14 months. The three MTA treatments-monthly medication management (usually methylphenidate) following weekly titration (MedMgt), intensive behavioral treatment (Beh), and the combination (Comb)-were designed to reflect known best practices within each treatment approach. Children were assessed at four time points in multiple outcome. Results indicated that Comb and MedMgt interventions were substantially superior to Beh and CC interventions for attention-deficit hyperactivity disorder symptoms. For other functioning domains (social skills, academics, parent-child relations, oppositional behavior, anxiety/depression), results suggested slight advantages of Comb over single treatments (MedMgt, Beh) and community care. High quality medication treatment characterized by careful yet adequate dosing, three times daily methylphenidate administration, monthly follow-up visits, and communication with schools conveyed substantial benefits to those children that received it. In contrast to the overall study findings that showed the largest benefits for high quality medication management (regardless of whether given in the MedMgt or Comb group), secondary analyses revealed that Comb had a significant incremental effect over MedMgt (with a small effect size for this comparison) when categorical indicators of excellent response and when composite outcome measures were used. In addition, children with parent-defined comorbid anxiety disorders, particularly those with overlapping disruptive disorder comorbidities, showed preferential benefits to the Beh and Comb interventions. Parental attitudes and disciplinary practices appeared to mediate improved response to the Beh and Comb interventions. PMID- 11265925 TI - Subjective refraction techniques in the frame of the three-dimensional dioptric space. AB - A novel heuristic approach to the well-known representation of the dioptric power in a three-dimensional space is presented. It is shown how this theoretical framework is ideal for discussing the principles of several subjective refraction methods. In particular, this formalism is used to justify the stenopaic slit refraction, the Barnes subjective refraction technique, and the Jackson cross cylinder procedure. In view of this analysis, some modifications to the traditional procedures are proposed. PMID- 11265924 TI - Microbial colonization of soft contact lenses over time. AB - PURPOSE: To compare the bacterial colonization of soft contact lenses in subjects for successively increasing periods, up to 13 nights of wear. The aim of this study was to determine whether increasing the length of lens wear predisposed subjects to high levels of microbial colonization of lenses. METHODS: Subjects (N = 20) were divided into those with a prior history of adverse events (N = 6), gram-negative bacterial carriers (N = 6), and those with no previous history (N = 8). RESULTS: There were no temporal changes in microbial colonization of lenses. Lenses from all wearers were colonized at least once during the study by gram positive bacteria at low numbers (<10 cfu/ml). Gram-negative bacteria colonized lenses at least once in 80% of all wearers. Lenses from gram-negative bacterial carriers were more frequently colonized by Staphylococcus aureus and Pseudomonas sp. compared with subjects with no previous history and subjects with a prior history of adverse events, respectively. Lenses from gram-negative bacterial carriers were less frequently colonized by a range of gram-positive bacteria compared with subjects with a prior history of adverse events. CONCLUSIONS: Increasing the length of lens wear up to 13 nights did not result in a predictable increase in bacterial colonization of contact lenses. Gram-positive bacteria were isolated frequently but in low numbers, whereas gram-negative bacteria were present sporadically. PMID- 11265926 TI - The slope of the psychometric function for Bailey-Lovie letter charts: defocus effects and implications for modeling letter-by-letter scores. AB - BACKGROUND: Visual acuity measurement often results in an imprecise endpoint because subjects correctly identify some but not all of the letters on one or more size levels on a letter chart. The extent of this transition zone from seeing to nonseeing can be described by probit size, which is calculated by performing Probit Analysis on letter chart data. There has been no previous research into the effects of optical defocus on letter chart probit size. METHODS: We tested 18 young visually normal subjects monocularly during three different defocus conditions: best spectacle correction (zero defocus) and +1.00 D and +2.00 D additions. Stimuli were Bailey-Lovie-style logarithm of the minimum angle of resolution (log MAR) letter charts constructed with a 0.05 logMAR size progression between size levels. Frequency of seeing data from these charts were used to calculate probit size. RESULTS: There were statistically significant effects of optical defocus on mean probit size. After Monte Carlo correction for bias, we believe that true mean values for probit size are about 0.07 logMAR for well-corrected subjects and up to 0.12 logMAR with optical defocus. CONCLUSION: The smaller probit size for well-corrected subjects should correspond to a sharper logMAR visual acuity endpoint and less intrasubject variability in logMAR acuity than for subjects with a larger probit size (optical defocus). Our modeling shows that these different probit sizes can also significantly affect letter-by-letter visual acuity scoring. PMID- 11265927 TI - Preschool vision screening: Maternal and Child Health Bureau and National Eye Institute Task Force on Vision Screening in the Preschool Child. PMID- 11265928 TI - Sieving named NEI Director. PMID- 11265929 TI - Postpartum proptosis with ophthalmopathy. AB - BACKGROUND: Postpartum thyroid disease presents in two forms: postpartum thyroiditis (PPT) and postpartum Graves' disease (PPGD). CASE REPORT: A case of postpartum thyroid dysfunction with ophthalmopathy is presented. DISCUSSION: In women diagnosed with Graves' disease during the ages of 20 to 35 years, 66% have a postpartum onset; women with a previous history of Graves' disease frequently relapse in the postpartum period. Additionally, up to 25 to 30% of postpartum women can develop permanent hypothyroidism from postpartum thyroiditis. The signs and symptoms of PPT and PPGD may be indefinite after delivery and often go undiagnosed. Because complications can be significant and may become irreversible, proper diagnosis and timely treatment is important. PMID- 11265930 TI - Granular corneal dystrophy: slitlamp biomicroscopic appearances in three generations of patients. AB - PURPOSE: The purpose of this case series is to show photographically the varying clinical appearance of granular corneal dystrophy in three generations of one family and to review the genetic basis of this and related conditions. CASE SERIES: We present cases for four affected individuals along with slitlamp biomicroscopic photographs. DISCUSSION: A review of the photographs and the literature suggests that the abnormal keratoepithelin first appears in the superficial cornea as faint subepithelial opacities. With time, these become arranged in the curved lines of a vortex pattern, after which the deposits become scattered in no particular pattern and at all levels of the cornea. In this family, corneal erosions are a regular feature. Mutations of the gene coding for keratoepithelin (beta ig-h3) may give rise to variable clinical manifestations. PMID- 11265931 TI - Beyond intraocular pressure: neuroprotective strategies for future glaucoma therapy. AB - BACKGROUND: All currently approved glaucoma medications are directed toward lowering intraocular pressure. However, it is apparent that there are pressure independent mechanisms associated with the development of glaucomatous optic neuropathy. There has been considerable effort to develop therapeutics that rescue the retinal ganglion cells from undergoing secondary degeneration after the original insult has occurred. This therapeutic strategy has been termed neuroprotection. METHODS: The literature was reviewed to examine the current knowledge of the degenerative cascade involved in glaucomatous damage, with emphasis on potential therapeutic targets for neuroprotective strategies. RESULTS: There are a number of promising areas of research for new glaucoma therapies including glutamate antagonists, calcium channel blockers, antioxidants, nitric oxide synthase inhibitors, neurotrophins, and anti-apoptotic agents. CONCLUSIONS: Glaucoma is a complex disease with a number of risk factors and mechanisms leading to ganglion cell death. Future glaucoma therapy will likely include neuroprotectants that could be used as an adjunct therapy with other medications designed to lessen the initial insult (i.e., intraocular pressure-lowering compounds). As the word neuroprotection becomes more popular, care must be taken in evaluating the research literature for clinically effective therapies. PMID- 11265932 TI - Hypoxic effects on the anterior eye of high-Dk soft contact lens wearers are negligible. AB - PURPOSE: To determine whether the eyes of high-Dk soft contact lens wearing subjects can be discriminated from non-contact lens wearing subjects. METHODS: This study was a prospective masked assessment of 32 subjects, 16 of whom wore experimental high-Dk soft contact lenses and 16 of whom did not wear contact lenses. Subjects wore high-Dk lenses on a 30-night replacement schedule for an average of 9 months. Tear film characteristics, staining and vascularization of the cornea, conjunctival staining, and the presence of microcysts in the corneal epithelium were assessed using slitlamp microscopy. The endothelium was examined for polymegethism. RESULTS: No differences were found between the two groups in any of the variables that were examined (p > 0.05) except that the high-Dk lens wearing group had about twofold more tear film debris and 2.5-fold more severe conjunctival staining (p < 0.05). CONCLUSIONS: Hypoxia-associated effects were not apparent in the eyes of subjects wearing experimental high-Dk soft contact lenses. Conjunctival staining can generally distinguish lens wearers from non lens wearers and can be used to discriminate between high-Dk lens wearing subjects and non-lens wearing subjects. PMID- 11265933 TI - Telemedicine and doctor-patient communication: an analytical survey of the literature. AB - The literature about the effect of telemedicine on doctor-patient communication was reviewed. A total of 38 studies were identified: six were surveys of provider and community attitudes; 21 were post-encounter surveys of participants in a medical consultation; and 11 were qualitative analyses of behaviour in a medical encounter. Twenty-one of the 38 investigations originated in the USA, six in the UK, four in Australia, three in Norway, two in Canada, one in Finland and one in Sweden. All were relatively recent. The findings from each study were coded according to 23 categories developed from the literature and a positive or negative rating was assigned to each of the 213 communication results. Approximately 80% of abstracted findings favoured telemedicine, with all but two of the 23 categories analysed (non-verbal behaviour and lack of touch) reporting more positive than negative results. Verbal content analysis is important for the development of interventions aimed at facilitating doctor-patient telecommunication. However, further research is necessary if the nature and content of the communication process are to be fully understood. PMID- 11265934 TI - A qualitative study of the organizational consequences of telemedicine. AB - The organizational consequences of telemedicine have frequently been mentioned in the telemedicine community, but there are few empirical studies. A study was therefore carried out of what happens in organizations when telemedicine is implemented. Qualitative interviews were undertaken with 30 persons working in teledermatology, telepsychiatry, a telepathology frozen-section service and tele otolaryngology. Almost all respondents reported numerous organizational changes, some important. Changes in work processes were the most common. Examples of the organizational consequences of telemedicine were organizational restructuring, new organizational units, changed mechanisms for internal coordination, different flows of patients through the health-care system, improved coordination of care, new job descriptions, relocation of the place of work, employment of personnel living far away from the workplace, effects on employees not directly involved in telemedicine, sharing of experiences, minor staffing changes, clinical teamwork independent of co-location, administrative meetings arranged by telemedicine, merger of organizations independent of location, less travel by staff (and patients), a possible beneficial effect on the quality of care, and limited opposition to the adoption of the technology. Telemedicine may be important in the future organization of the disciplines studied and in health-care generally. The infrastructure of electronic networks may play an important role for organizations as the volume of telemedicine activity increases and economies of scale are realized. PMID- 11265935 TI - Evaluation of telemedical orthopaedic specialty support to a minor accident and treatment service. AB - Over three and a half years there were 200 teleconsultations between emergency nurse practitioners at a minor accident and treatment service and the orthopaedic service of a main hospital. The main problems were fractures (93% of cases). The reasons for consultation fell almost equally into four groups: request for direct ward admission; discussion and decision about treatment; decision about the disposition of a case; and diagnosis. The technical quality of the majority of teleconsultations was considered satisfactory. Following the teleconsultation, 39% of patients were admitted to hospital, 6% were transferred to the accident and emergency department for a face-to-face consultation, and 56% were discharged and referred to a review clinic. Of the 200 cases, 193 needed teleradiology and the nurse practitioners and the orthopaedic registrars diagnosed all these cases correctly, as judged by the subsequent radiologist's report. Teleconsultations save time and prevent the unnecessary transfer of patients to main hospitals. PMID- 11265936 TI - General practitioner participants in a telemedicine trial: comparisons with their peers. AB - A study was conducted to determine whether general practitioners (GPs) participating in a telemedicine trial were self-selected enthusiasts for information technology compared with GPs in general. We compared two experimental groups of GPs in London and Wales (n = 126) who had volunteered to participate in a telemedicine trial and two randomly selected groups of GPs from the same areas (total n = 300) who acted as controls. A postal questionnaire was used and achieved a 79% response rate. There were no significant differences in the demographic characteristics of the experimental and control groups in London and Wales, except that the London telemedicine GPs (mean age 41.9 years) were significantly younger than the London controls (mean age 46.6 years). The control GPs were similar to the telemedicine GPs in terms of frequency of computer use and their responses to statements about their attitudes to computers. In the combined telemedicine and control groups, 85% and 77% respectively said that they enjoyed using computers. Telemedicine GPs used computers more frequently for administrative purposes but they shared the same attitudes towards information technology as GPs in general. In all situations where GPs could exercise personal choice, the control and experimental groups were similar. PMID- 11265937 TI - The role of telenursing in the provision of geriatric outreach services to residential homes in Hong Kong. AB - A residential nursing home in Hong Kong was linked to the community geriatric assessment team based in a regional hospital using videoconferencing equipment operating at 384 kbit/s. The feasibility of providing nursing services and their acceptability to users were evaluated over 12 months. There were 198 occupants of the nursing home and their mean age was 82 years (range 60-101). Services included patient education regarding the use of a metered dose inhaler, wound management and a falls prevention programme, together with assessment of clients' need for infirmary care and the risk of aspiration. The acceptability of the system to the clients and nursing home staff was also assessed. It was found that 89% of such services could be carried out via telemedicine, and only 11% required on-site visits. There was an increase in the proportion of patients correctly using inhalers as well as a reduction in the number of falls. More consultations were conducted by the nurse (an increase of 76% per month) and an additional 8.4 patients per month could be attended to by the nurse compared with 5 patients when on-site visits were used. Acceptability to clients and nursing home staff was good. The problem of lack of resources to support elderly residential care institutions makes service delivery via telemedicine appear economically attractive, as well as facilitating improvements in the quality of long-term care. PMID- 11265938 TI - Improvement in diabetes control with a monitoring system based on a hand-held, touch-screen electronic diary. AB - We conducted a six-month prospective interventional crossover study examining a computerized diabetes monitoring system (DMS) that conveyed dietary information. The objectives were to compare glycaemic control between intervention and control periods, and to assess patients' acceptance of the DMS. Nineteen patients were randomized into two groups, each using the DMS for three months and serving as the control group for another three months. The patients recorded information about their meal portions and blood glucose readings in a hand-held electronic diary. After transmitting the data to the DMS through a telephone modem, the patients received immediate feedback about the carbohydrate, protein and fat content of the meal, as well as the calorie content. A significant improvement in glycaemic control was achieved during intervention compared with control periods (mean HbA1C reduction of 0.825%). The DMS was also highly acceptable: 95% patients found it easy to operate while 63% found it useful. The DMS was thus a feasible model of telemedicine in diabetes care and a larger study is warranted to examine its cost-effectiveness. PMID- 11265939 TI - An internet-based blood pressure monitoring system for patients. AB - We developed a personal blood pressure monitoring system for patients with hypertension or hypotension. The system can be used to measure a patient's blood pressure at home and to transmit the data automatically to a hospital database via the Internet. The accuracy of blood pressure readings using the system was assessed by comparison with readings from a standard digital sphygmomanometer in four subjects. The measurement error for the systolic readings was 1.7-2.7% and for the diastolic readings 2.7-3.2%. The system therefore appears to be a promising means of assessing blood pressure remotely. PMID- 11265940 TI - Telemedicine as a support system to encourage breast-feeding in Northern Ireland. PMID- 11265942 TI - Computer clothes. PMID- 11265941 TI - A development and evaluation template for minor injuries telemedicine. PMID- 11265943 TI - Animal research in the post-genome era. PMID- 11265944 TI - Non-therapeutic research in the EU in adults incapable of giving consent? PMID- 11265945 TI - Transplantation from non-heart-beating donors. PMID- 11265946 TI - Shedding light on photopheresis. PMID- 11265947 TI - Panayiotopoulos syndrome: a common and benign childhood epilepsy. PMID- 11265948 TI - Is pacifier use a risk factor for otitis media? PMID- 11265949 TI - The universe weighed and found wanting. PMID- 11265950 TI - Transplantation of lungs from a non-heart-beating donor. AB - BACKGROUND: In animals, we have previously done successful lung transplantations using organs from non-heart-beating donors. We have also developed an ex-vivo system of assessing the function of such organs before transplantation. The next stage was to try the technique in human beings. Bearing in mind the sensitive ethical issues involved, our first aim was to find out what procedures would be acceptable, and to use the results to guide a clinical lung transplantation from a non-heart-beating donor. METHODS: The ethical acceptability of the study was gauged from the results of a broad information programme directed at the general public in Sweden, and from discussions with professionals including doctors, nurses, hospital chaplains, and judges. The donor was a patient dying of acute myocardial infarction in a cardiac intensive-care unit after failed cardiopulmonary resuscitation. The next of kin gave permission to cool the lungs within the intact body, and intrapleural cooling was started 65 min after death. Blood samples were sent for virological testing and cross matching. The next of kin then had time to be alone with the deceased. After 3 h, the body was transported to the operating theatre and the heart-lung block removed. The lungs were assessed ex vivo, and the body was transported to the pathology department for necropsy. RESULTS: No contraindications to transplantation were found, and the right lung was transplanted successfully into a 54-year-old woman with chronic obstructive pulmonary disease. The donor lung showed excellent function only 5 min after reperfusion and ventilation, and during the first 5 months of follow-up, the function of the transplanted lung has been good. INTERPRETATION: About half the deaths in Sweden are caused by cardiac and cerebrovascular disease. This group could be a potential source of lung donors. When all hospitals and ambulance personnel in Sweden have received training in non-heart beating lung donation, we hope that there will be enough donor lungs of good quality for all patients needing a lung transplant. PMID- 11265951 TI - Oral amiodarone for prevention of atrial fibrillation after open heart surgery, the Atrial Fibrillation Suppression Trial (AFIST): a randomised placebo controlled trial. AB - BACKGROUND: Beta-blockers and amiodarone reduce the frequency of atrial fibrillation after open-heart surgery but the effectiveness of oral amiodarone in older patients already receiving beta-blockers is unknown. We have assessed the efficacy of oral amiodarone in preventing atrial fibrillation in patients aged 60 years or older undergoing open-heart surgery. METHODS: We did a randomised, double-blind placebo-controlled trial in which patients undergoing open-heart surgery (n=220, average age 73 years) received amiodarone (n=120) or placebo (n=100). Patients enrolled less than 5 days before surgery received 6 g of amiodarone or placebo over 6 days beginning on preoperative day 1. Patients enrolled at least 5 days before surgery received 7 g over 10 days beginning on preoperative day 5. FINDINGS: Patients on amiodarone had a lower frequency of any atrial fibrillation (22.5% vs 38.0%; p=0.01; absolute difference 15.5% [95% CI 3.4-27.6%]), and there were significant differences in favour of the active drug for symptomatic atrial fibrillation (4.2% vs 18.0%, p=0.001), cerebrovascular accident (1.7% vs 7.0%, p=0.04), and postoperative ventricular tachycardia (1.7% vs 7.0%, p=0.04). Beta-blocker use (87.5% amiodarone vs 91.0% placebo), nausea (26.7% vs 16.0%), 30-day mortality (3.3% vs 4.0%), symptomatic bradycardia (7.5% vs 7.0%), and hypotension (14.2% vs 10.0%) were similar. INTERPRETATION: Oral amiodarone prophylaxis in combination with beta-blockers prevents atrial fibrillation and symptomatic fibrillation and reduces the risk of cerebrovascular accidents and ventricular tachycardia. PMID- 11265952 TI - Risk of colorectal cancer after breast cancer. AB - BACKGROUND: History of breast cancer has been reported as a risk factor for colorectal cancer in women. In view of the ambiguous nature of existing evidence and the growing interest in targeted colorectal cancer prevention, we sought to quantify this risk. METHODS: We used the Surveillance Epidemiology and End Results (SEER) database to estimate risk of colorectal cancer after breast-cancer diagnosis in women with first incident breast cancer between 1974 and 1995. Observed colon and rectal cancer risk was compared with that expected in the general population. We stratified comparisons by age at breast-cancer diagnosis, stage of cancer, ethnic origin of patient, and follow-up time. FINDINGS: Overall, women with previous breast cancer were 5% less likely (95% CI 1-9) to develop colon and 13% less likely (6-19) to develop rectal cancer than women in the general population. Stratified analyses suggested that the risk reductions observed for colon and rectal cancer were most pronounced for women with breast cancer diagnosed after age 65 years, in white women, women with local stage breast cancer, and women diagnosed in the later study years (1990-94). INTERPRETATIONS: Breast cancer does not increase subsequent colorectal cancer risk, and reduced risk was seen for certain subgroups of women. Because no biologically plausible endogenous protective factor has been identified, we suggest that reduced risk could stem from an accumulation of exposures that increase breast-cancer frequency but protect against colorectal cancer. PMID- 11265954 TI - Urinary sodium excretion and cardiovascular mortality in Finland: a prospective study. AB - BACKGROUND: The evidence that high salt intake increases the risk of cardiovascular disease has been challenged. We aimed to find out whether salt intake, measured by 24 h urinary sodium excretion, is an independent risk factor for cardiovascular disease frequency and mortality, and all-cause mortality. METHODS: We prospectively followed 1173 Finnish men and 1263 women aged 25-64 years with complete data on 24 h urinary sodium excretion and cardiovascular risk factors. The endpoints were an incident coronary and stroke event, and death from coronary heart disease, cardiovascular disease, and any cause. Each endpoint was analysed separately with the Cox proportional hazards model. FINDINGS: The hazards ratios for coronary heart disease, cardiovascular disease, and all-cause mortality, associated with a 100 mmol increase in 24 h urinary sodium excretion, were 1.51 (95% CI 1.14-2.00), 1.45 (1.14-1.84), and 1.26 (1.06-1.50), respectively, in both men and women. The frequency of acute coronary events, but not acute stroke events, rose significantly with increasing sodium excretion. When analyses were done separately for each sex, the risk ratios were significant in men only. There was a significant interaction between sodium excretion and body mass index for cardiovascular and total mortality; sodium predicted mortality in men who were overweight. Correction for the regression dilution bias increased the hazards ratios markedly. INTERPRETATION: High sodium intake predicted mortality and risk of coronary heart disease, independent of other cardiovascular risk factors, including blood pressure. These results provide direct evidence of the harmful effects of high salt intake in the adult population. PMID- 11265953 TI - Cognitive behaviour therapy for chronic fatigue syndrome: a multicentre randomised controlled trial. AB - BACKGROUND: Cognitive behaviour therapy (CBT) seems a promising treatment for chronic fatigue syndrome (CFS), but the applicability of this treatment outside specialised settings has been questioned. We compared CBT with guided support groups and the natural course in a randomised trial at three centres. METHODS: Of 476 patients diagnosed with CFS, 278 were eligible and willing to take part. 93 were randomly assigned CBT (administered by 13 therapists recently trained in this technique for CFS), 94 were assigned the support-group approach, and 91 the control natural course. Multidimensional assessments were done at baseline, 8 months, and 14 months. The primary outcome variables were fatigue severity (on the checklist individual strength) and functional impairment (on the sickness impact profile) at 8 and 14 months. Data were analysed by intention to treat. FINDINGS: 241 patients had complete data (83 CBT, 80 support groups, 78 natural course) at 8 months. At 14 months CBT was significantly more effective than both control conditions for fatigue severity (CBT vs support groups 5.8 [2.2-9.4]; CBT vs natural course 5.6 [2.1-9.0]) and for functional impairment (CBT vs support groups 263 [38-488]; CBT vs natural course 222 [3-441]). Support groups were not more effective for CFS patients than the natural course. Among the CBT group, clinically significant improvement was seen in fatigue severity for 20 of 58 (35%), in Karnofsky performance status for 28 of 57 (49%), and self-rated improvement for 29 of 58 (50%). Prognostic factors for outcome after CBT were a higher sense of control predicting more improvement, and a passive activity pattern and focusing on bodily symptoms predicting less improvement. INTERPRETATION: CBT was more effective than guided support groups and the natural course in a multicentre trial with many therapists. Our study showed a lower proportion of patients with improvement than CBT trials with a few highly skilled therapists. PMID- 11265955 TI - A lump in the mouth. PMID- 11265956 TI - Variant esp gene as a marker of a distinct genetic lineage of vancomycin resistant Enterococcus faecium spreading in hospitals. AB - In the USA, vancomycin-resistant Enterococcus faecium (VREF) is endemic in hospitals, despite lack of carriage among healthy individuals. In Europe, however, hospital outbreaks are rare, but VREF carriage among healthy individuals and livestock is common. We used amplified fragment-length polymorphism analysis to genotype 120 VREF isolates associated with hospital outbreaks and 45 non epidemic isolates from the USA, Europe, and Australia. We also looked for the esp virulence gene in these isolates and in 98 VREF from animals. A specific E. faecium subpopulation genetically distinct from non-epidemic VREF isolates was found to be the cause of the hospital epidemics in all three continents. This subpopulation contained a variant of the esp gene that was absent in all non epidemic and animal isolates. Identification of the variant esp gene will be important in guiding infection-control strategies, and the Esp protein could be a new target for antibacterial therapy. PMID- 11265957 TI - Enterococcal vanB resistance locus in anaerobic bacteria in human faeces. AB - While developing a rapid method to detect carriers of vancomycin-resistant enterococci (VRE), we found the vanB gene by PCR in 13 of 50 human faecal specimens that did not contain culturable VRE. Passaging under antibiotic selection allowed us to isolate two species of anaerobic bacteria that were vanB PCR positive, vancomycin resistant, and teicoplanin sensitive. Sequence analysis of the 16S rRNA genes showed that one isolate resembled Eggerthella lenta (98% identity), and the other Clostridium innocuum (92% identity). Southern hybridisation and nucleotide sequencing showed a vanB locus homologous to that in VRE. We propose that vanB resistance in enterococci might arise from gene transfer in the human bowel. PMID- 11265958 TI - Early detection of heart transplant patients with increased risk of ciclosporin nephrotoxicity. AB - Chronic nephrotoxic effects from ciclosporin are a common clinical complication after heart transplantation and frequently lead to progressive renal failure. There is no laboratory test to predict development of chronic renal failure in heart transplant patients. We analysed urinary retinol-binding protein (uRBP) concentration, to assess proximal tubular dysfunction, in 36 clinically stable heart transplant patients. We detected a subgroup of 13 patients who had high concentrations of uRBP, good renal function, and a high risk of developing progressive renal failure compared with patients with normal uRBP (relative risk 3.87, p=0.003). PMID- 11265959 TI - Infective cause of childhood leukaemia and wartime population mixing in Orkney and Shetland, UK. AB - In Orkney and Shetland (the UK's northernmost islands), during World War II, local people were outnumbered by servicemen stationed there in case of a northern invasion. Such rural-urban population mixing promotes contact between susceptible and infected individuals. We compared childhood leukaemia mortality in wartime and postwar cohorts of Orkney and Shetland children. Childhood leukaemia increased 3.6-fold, (p=0.001) in the wartime, but not in the postwar, cohort compared with national Scottish rates. These findings add to the evidence for infection as a cause of childhood leukaemia. PMID- 11265960 TI - Should visits to relatives carry a health warning? PMID- 11265962 TI - Competition prompts drug companies to cut antiretroviral drug prices. PMID- 11265961 TI - Gordon Brown unveils "good news" budget for health. PMID- 11265963 TI - Global population will increase to nine billion by 2050, says UN report. PMID- 11265964 TI - US newspaper charges research centre with misconduct. PMID- 11265965 TI - Ireland's government appeals free education for disabled children ruling. PMID- 11265966 TI - International group aims to advise countries on mental health policies. PMID- 11265967 TI - Teaching professional development in medical schools. AB - Doctors must increasingly be aware of what they should be, as well as what they should know. Professionalism, including a value system that supports the compassionate care of patients, is a means of encapsulating and prioritising these competing responsibilities. Accordingly, in this article, we assume that professionalism is an essential aspect of medical practice that needs to be taught to those entering medicine. We first describe critiques of professionalism and current challenges to it, in practice and in medical education. We then assess the current efforts of curriculum reform to incorporate professionalism and the methods used to teach it. Adopting and assessing such approaches to ensure that they are effective is of central importance in the education of future clinicians. PMID- 11265968 TI - Queen Margrethe II and mortality in Danish women. AB - All-cause mortality in women is declining in all western European countries, apart from in Danish women. All-cause mortality in Danish women, age-adjusted to 45-74 years was compared with all-cause mortality in women in Scotland and with the mean of the other countries of the European Union for 1970-96. The decline of all-cause mortality in Danish women stopped in 1978 whereas the decline continued in Scotland. In 1996, all-cause mortality was 48% higher in Denmark than that of the mean for the European Union countries. Many Danish women are smokers. Halting of the decline in mortality occurred about 5 years after the ascension to the throne of Denmark by Queen Margrethe II. The queen is very popular in Denmark and a known cigarette smoker. As a role model for women, the Queen's example could offer an explanation for the unusual mortality in Danish women. PMID- 11265969 TI - Growing pains of East Timor: health of an infant nation. AB - In August, 1999, three-quarters of East Timorese adults voted to end more than two decades of an Indonesian administration never recognised by the United Nations. The ensuing spree of violence and destruction by militia backed by the Indonesian military meant the birth of the fledgling nation became a complex humanitarian disaster. 1 year on, progress was heartening: a transitional government, a judiciary, and tax systems were in place, and East Timor was a proud competitor in the Sydney Olympic games. Rebuilding a country from ground level has brought a golden opportunity for fresh approaches. However, reconstruction is also a slow, complex, and sometimes controversial process at the mercy of multiple agendas. The health sector has seen basic care restored, establishment of a much-needed public-health service, and planning for the future health system. An innovative partnership between WHO/Roll Back Malaria and Merlin for post-conflict research has provided data to guide malaria control. The story of progress from humanitarian emergency to national health plan epitomises the triumphs and challenges of this newest nations' first 18 months. PMID- 11265970 TI - A place on the map. PMID- 11265971 TI - Dementia and statins. PMID- 11265972 TI - Dementia and statins. PMID- 11265973 TI - Dementia and statins. PROSPER study group. PMID- 11265974 TI - Dementia and statins. PMID- 11265975 TI - Dementia and statins. PMID- 11265976 TI - Dementia and statins. PMID- 11265977 TI - Deferiprone or fatal iron toxic effects? PMID- 11265978 TI - Predictive testing for Huntington's disease. PMID- 11265979 TI - Treatment of oxygen-induced hypercapnia. PMID- 11265980 TI - Treatment of oxygen-induced hypercapnia. PMID- 11265981 TI - Treatment of oxygen-induced hypercapnia. PMID- 11265982 TI - Animal testing in India. PMID- 11265983 TI - Trends in use of positron emission tomography. PMID- 11265984 TI - Subarachnoid haemorrhage. PMID- 11265985 TI - Fetal microchimerism and inflammatory myopathies. PMID- 11265986 TI - Pain in Anderson-Fabry's disease. PMID- 11265987 TI - Invasive brain tumour after radiosurgery. PMID- 11265988 TI - Anonymous testing for HIV in tuberculosis cases and contacts. PMID- 11265989 TI - Electrocardiography first for reducing cot death. PMID- 11265990 TI - Redefining goals for tobacco control. PMID- 11265991 TI - Fake antimalaria drugs in Cambodia. PMID- 11265992 TI - Women in advertising. PMID- 11265993 TI - Sizing up research. PMID- 11265994 TI - Transparency and openness: the Government's response to the BSE inquiry. PMID- 11265995 TI - Relationships in broiler chickens between lameness, liveweight, growth rate and age. AB - Thirteen genotypes of poultry were selected to represent a wide range of growth profiles and were fed either a non-limiting or Label Rouge diet. The birds' degree of lameness and liveweight were measured after 54 and 81 days. The birds reared on the Label Rouge diets were less lame than birds of the same genotype reared on the non-limiting diet. More traditional and slower growing genotypes tended to be less lame than the modern genotypes reared on the same feeding regimen. All the birds, irrespective of their genotype or diet, were less lame after 54 days than after 81 days. However, when liveweight was included in the analysis as a covariable, many of the differences disappeared. Only age at assessment significantly affected the walking ability of the birds, with the birds being approximately 0.6 units of gait score worse at 54 days of age than at 81 days. The regression coefficient between gait score and liveweight was 1.262 at 54 days and 1.128 at 84 days. The results indicated that liveweight was an important determinant of lameness in the diverse range of genotypes examined, that growth rate was also an important determinant of lameness and that younger birds may be more sensitive to differences in liveweight than older birds. PMID- 11265996 TI - Evaluation and optimisation of a target-controlled infusion system for administering propofol to dogs as part of a total intravenous anaesthetic technique during dental surgery. AB - The performance of a modified target-controlled infusion system was investigated in 16 dogs undergoing routine dental work, by comparing the predicted concentrations of propofol in venous blood samples with direct measurements; the optimum targets for the induction and maintenance of anaesthesia were also identified. The performance of a target-controlled infusion system is considered clinically acceptable when the median prediction error, a measure of bias, is not greater than +/-10 to 20 per cent, and the median absolute performance error, a measure of the accuracy, is not greater than 20 to 30 per cent. The results fell within these limits indicating that the system performed adequately. The optimal induction target was 3 microg/ml, and anaesthesia of adequate depth and satisfactory quality was achieved with maintenance targets of between 2.5 and 4.7 microg/ml propofol. The system was easy to use and the quality of anaesthesia was adequate for dental work. PMID- 11265997 TI - Variations in cartilage catabolism in different equine joints in response to interleukin-1 in vitro. AB - An explant system was used to investigate the hypothesis that cartilage from different equine joints might respond differently to challenge with interleukin 1alpha (IL-1alpha). Pairs of normal cartilage samples were taken from the metacarpophalangeal, proximal interphalangeal and distal interphalangeal joints of six horses. One of each pair was stimulated with 10 ng/ml human recombinant IL 1alpha for three days, and the supernatants and remaining cartilage explants were analysed for their total content of glycosaminoglycans. A significantly higher percentage of glycosaminoglycans was released from the cartilage of the proximal and distal interphalangeal joints than from the metacarpophalangeal joint. PMID- 11265998 TI - Streptococcus suis serotypes 3 to 28 associated with disease in pigs. PMID- 11265999 TI - Relationship between serum biotin concentration and moisture content of the sole horn in cows with clinical laminitis or sound hooves. PMID- 11266000 TI - Morbillivirus infection in a bottlenosed dolphin and a Mediterranean monk seal from the Atlantic coast of West Africa. PMID- 11266001 TI - Liver lobe torsion in an oriental small-clawed otter (Aonyx cinerea). PMID- 11266002 TI - Veterinary ethics: filling a gap in undergraduate education. PMID- 11266003 TI - Closure of the Thurso veterinary investigation centre. PMID- 11266004 TI - Closure of the Thurso veterinary investigation centre. PMID- 11266005 TI - Maedi visna infection in a flock in Yorkshire. PMID- 11266006 TI - Privilege to dispense. PMID- 11266007 TI - Ionophore toxicity in turkeys. PMID- 11266008 TI - Prevalence of Bartonella henselae in cats in the UK. PMID- 11266009 TI - Bovine viral diarrhoea virus serology: antibody response after vaccination. PMID- 11266010 TI - Fees and insurance. PMID- 11266011 TI - Angiostrongylus vasorum in dogs. PMID- 11266012 TI - Assisted reproduction in the dog. PMID- 11266013 TI - Cerebral activation patterns in patients with writer's cramp: a functional magnetic resonance imaging study. AB - Functional MRI (fMRI), visualizing changes in cerebral blood oxygenation, has to date not been performed either in patients with writer's cramp or in healthy subjects during writing. We compared the cerebral and cerebellar activation pattern of 12 patients with writer's cramp during writing with a group of 10 healthy subjects performing the same tasks over 30-s periods of rest or writing. Sixty echo planar imaging multislice datasets were analysed using SPM96 software. Data were analysed for each subject individually and groupwise for patients vs. controls. Healthy subjects showed a significant activation of the ipsilateral dentate nucleus, contralateral cerebellar hemisphere, contralateral primary sensorimotor cortex, and contralateral precentral gyrus during writing. Patients with writer's cramp showed significantly greater activation of the ipsilateral cerebellar hemisphere than controls. Also the activation in the primary sensorimotor cortex extended further caudally and anteriorly towards the premotor association area. Activation was observed in the thalamus during writing only among the patients. Our results indicate an increased basal ganglia output via the thalamus to the motor and premotor cortical areas in dystonia patients and support the notion of disinhibition of the motor cortex leading to cocontractions and dystonic postures. PMID- 11266014 TI - Spinal cord compression due to extramedullary haematopoiesis in two patients with thalassaemia: complete regression with blood transfusion therapy. AB - Extramedullary haematopoiesis, a common finding in thalassaemia, is rarely localized in the spinal cord and even more rarely has neurological manifestations. We present two patients suffering from thalassaemia (intermedia and beta homozygous) and paraparesis due to spinal cord compression from intrathoracic extramedullary haematopoietic masses. Diagnosis was based on magnetic resonance imaging findings, and treatment consisted of blood hypertransfusions. The majority of published cases have been successfully treated by radiation and in some cases by blood transfusions. Our two patients completely recovered, and there has been no recurrence during the 4 years since their treatment. Diagnosis and treatment of this rare condition are discussed. PMID- 11266015 TI - Epileptic and non-epileptic seizures in multiple sclerosis. AB - Knowledge concerning the relationship between multiple sclerosis and epilepsy is reviewed. Epidemiological studies have established that epileptic seizures are more frequent in multiple sclerosis than predicted by chance. Partial epilepsies with focal seizures often with atypical symptoms and with or without secondary generalisation are the usual pattern. In the survey special emphasis is laid on the direct correlation between paroxysmal phenomena and plaques now demonstrable by modern imaging techniques. These images have shown that epileptic seizures can be caused by cortical and subcortical lesions and by their accompanying oedema. We extend the review to non-epileptic paroxysmal symptoms, such as tonic spasm, which may be confused with epileptic seizures. As far as they are supported by data, recommendations for diagnosis and therapy are given. Open questions are identified and issues for further research are suggested. PMID- 11266016 TI - Thirty-seven CAG repeats in the androgen receptor gene in two healthy individuals. AB - X-linked recessive spinobulbar muscular atrophy (SBMA) is an adult-onset X-linked neurodegenerative disease, characterised by muscular atrophy, bulbar symptoms and endocrinological disturbances. SBMA is caused by the expansion of a CAG repeat in the androgen receptor gene. The maximum number of CAG repeats found in a healthy person is 35 while the minimum number of repeats found in SBMA patients is 38. We have identified a 46-year-old man from an SBMA family with 37 CAG repeats who until now is clinically unaffected. Interestingly, his 85-year-old mother who has the genotype 37/51 CAG repeats is clinically unaffected as well. These results suggest an exactly defined border between normal and disease alleles. PMID- 11266017 TI - Time estimation in Parkinson's disease: normal long duration estimation despite impaired short duration discrimination. AB - It has been claimed that patients with Parkinson's disease (PD) are deficient in estimating and reproducing time intervals in the range of seconds. This deficit is more severe when subjects are requested to count internally during the demanded intervals, and when the rate of internal counting is fast. The observed deficit might therefore reflect slow internal counting, i. e. motor slowness, rather than a specific deficit of temporal processing. However, PD patients also have a higher temporal discrimination threshold for sensory stimuli, a finding purportedly indicating a slow 'internal clock'. In this study we examined PD patients' processing of short durations (approx. 1s,'psychological present') and long durations (up to 48 s,'extended present'). In the first experiment the ability to discriminate between temporally overlapping presentations of two visual stimuli (darkened rectangles on the computer screen) in the range of one second was tested. In the second experiment PD patients' ability to estimate time intervals between 12 and 48 s was investigated. During these intervals, subjects were required to tap and read a random number on the computer screen at a rate of 1 Hz. We found that the patients were impaired at discriminating between short intervals. This deficit was independent of task difficulty and appeared to be based on an impairment of divided attention. Despite this deficit, the PD patients estimated time intervals up to 48 s as accurately as the controls. We suggest that time estimation, i. e. the feeling for the flow of time, is normal in PD patients despite impaired discrimination of brief intervals in the range of seconds. PMID- 11266018 TI - Phenotype assignment in symptomatic female carriers of X-linked adrenoleukodystrophy. AB - However, only very few studies have systematically investigated the extent and distribution of nervous and endocrine system involvement in female carriers of X ALD. To define the phenotype in symptomatic female carriers of X-ALD we performed a prospective study including eight symptomatic women who were followed for a mean period of 3.4+/-1.8 years (range 1-6) using standardized clinical examination protocols, magnetic resonance imaging and spectroscopy, evoked potential studies including visual, brainstem auditory, somatosensory and magnetic evoked potentials, neurographic recordings and endocrine studies. Spastic paraparesis and decreased vibration sense in the lower extremities were the most frequent clinical findings. Slightly hyperintense symmetric parieto occipital white matter lesions on magnetic resonance imaging were detectable in two of seven cases, and the N-acetylaspartate/choline ratios on magnetic resonance spectroscopy were decreased in three of seven patients. P40 latencies were abnormal in all patients, and central motor conduction times to the lower extremities in seven of eight patients. Prolonged latencies of brainstem auditory evoked potential waves III-V or interpeak latencies of waves I-III, I-V and III-V were detectable in all patients. The degree of walking impairment was positively correlated with the duration of clinical disease (r=0.58, P < 0.05) and inversely correlated with the N-acetylaspartate/choline ratios (r=0.85; P < 0.05). Neurographic recordings revealed only subtle abnormalities, suggesting that nervous system involvement in symptomatic female carriers of X-ALD is confined mainly to the central nervous system. No evidence of adrenal insufficiency was detected in any of the patients. PMID- 11266019 TI - Neuropsychological profiles of familial Alzheimer's disease associated with mutations in the presenilin 1 and amyloid precursor protein genes. AB - Patients with familial Alzheimer's disease and a subset known to have presenilin mutations were compared with sporadic cases on a comprehensive battery of cognitive tests. These included measures of memory, intelligence, language and perception. The three groups were very comparable, in terms of severity, on global measures of dementia. However, their profiles/patterns of cognitive impairment differed in two respects; the group with sporadic Alzheimer's disease were significantly more impaired on tests of object naming and object perception than either the group with familial Alzheimer's disease or group with familial Alzheimer's disease and presenilin mutations, yet they scored at a significantly higher level on the measure of verbal intelligence. This study provides further evidence of the heterogeneity of the disease process. PMID- 11266020 TI - Abnormal postexcitatory and interhemispheric motor cortex inhibition in writer's cramp. AB - Focal transcranial magnetic stimulation (TMS) of the motor cortex was used to study excitatory and inhibitory stimulation effects in 25 patients with writer's cramp and 25 healthy volunteers. We investigated excitatory and inhibitory corticospinally mediated motor effects in muscles contralateral to the stimulation side as well as interhemispheric inhibition of tonic motor activity in muscles ipsilateral to stimulation. Motor evoked potentials (MEPs) were recorded from both first dorsal interosseus muscles. Motor thresholds at rest and amplitudes and latencies of MEPs obtained during maximal contraction were always bilaterally normal. The duration of postexcitatory inhibition was significantly shortened (168+/-55 vs. 198+/-39 ms in normal subjects, P=0.001) and the duration of interhemispheric inhibition prolonged (30.3+/-6.6 vs. 26+/-3.9 ms in normal subjects, P < 0.001). Both observations would be compatible with a decreased inhibition of corticospinal and transcallosal outputs of the motor cortex. The results were not influenced by fatigue effects. Abnormal motor cortex inhibition seems to be a generalized phenomenon in writer's cramp since it was detected in both hemispheres and during a simple isometric motor task which did not evoke dystonic symptoms. PMID- 11266021 TI - Wernicke's encephalopathy in a patient with Crohn's disease: a pathological study. PMID- 11266022 TI - Multiple sclerosis-like disease in polyglandular autoimmune syndrome. PMID- 11266023 TI - Polyradiculopathy in the course of superficial siderosis of the CNS. PMID- 11266024 TI - Polyneuritis cranialis related to anti-GT1 a IgG antibody. PMID- 11266025 TI - Acetylsalicylic acid induced cessation of transient ischaemic attacks and microembolic signals detected by transcranial Doppler in a patient with essential thrombocythaemia. PMID- 11266026 TI - Magnetic stimulation during a somatosensory-precipitated epileptic seizure. PMID- 11266028 TI - Glial cell line-derived neurotrophic factor (GDNF) prevents neurodegeneration in models of Parkinson's disease. PMID- 11266027 TI - S.A. Kinnier Wilson (1878-1937). PMID- 11266029 TI - Disproportion of economic impact, research achievements and research support in digestive diseases in Canada. AB - OBJECTIVES: To assess the economic impact, research output and research support of digestive diseases, and to compare them to those of other common disease entities, specifically mental, cardiovascular, respiratory, and central nervous system diseases. METHODS: Economic burden was assessed with the use of (a) published Canadian government data of direct cost from 1963 to 1993, (b) data from the Canadian Institute of Health Information and (c) recent Canadian economic studies. Research achievements were assessed on the basis of (a) research training in Canadian units, (b) individual achievements by Canadian investigators and (c) contribution to meetings and reception of awards. Research support was assessed by reviewing (a) Canadian government publications, (b) the Association of Canadian Medical Colleges, (c) the Medical Research Council (MRC) of Canada, (d) charitable organizations and (e) the Canadian Association of Gastroenterology (CAG). RESULTS: Digestive diseases are responsible for 15% of the total direct economic burden of Canadian health costs, and this figure exceeds those for mental, cardiovascular, respiratory and central nervous system diseases. Hospital discharges for digestive diseases contribute 12% of all hospitalizations and 20% of all neoplasias. Digestive diseases cause short-term loss of productivity, costing $1.14 billion/yr and exceeding the costs of mental, cardiovascular, respiratory and central nervous system diseases. Eighty-one percent of Research Fellows trained in Canadian units entered academic positions, and 63% obtained operating grants. Canadian investigators made important contributions in all areas of digestive science and received major international awards. Government support for digestive diseases was less than that for cardiovascular and neurologic research. In contrast to the highest economic burden, university staffing and residents were fewer for digestive than for mental, cardiovascular, respiratory and neurologic diseases. The number of MRC grants decreased, mainly because of organizational problems. Most charitable organizations support research specifically oriented to the disease of their interest. The CAG was the major supporter of non-specified research. CONCLUSIONS: Digestive diseases are responsible for a major economic burden. Scientists in this field have established international recognition, but research support lags behind the need to correct the economic burden and to provide future generations of scientists in the digestive sciences. There is need for government to readdress this shortcoming and to review its method of support. PMID- 11266031 TI - Canadian research fellowship training programs in digestive sciences: achievements and challenges. AB - BACKGROUND: The Canadian Association of Gastroenterology (CAG) is committed to fostering the development of future Canadian investigators. Up to 1986, research fellowship support was obtained from the Medical Research Council (MRC) of Canada. Since that time, several peer-reviewed, industry-sponsored, CAG-supported research fellowships and a variety of independently funded awards have augmented this effort. In the same period, peer-reviewed operating grants (OGs) from the MRC and other agencies have been constrained. The aim of this study was to determine the success of CAG, MRC or any other Canadian research fellowships in the development of career investigators in digestive sciences and to identify factors influencing the outcomes of such training. METHODS: MRC records and the minutes of CAG annual meetings were reviewed to identify research fellowship support. Canadian program directors were requested to list research fellows affiliated with their groups between 1986 and 1997. Only fellowships providing at least 1 year of training were included. A 7-page questionnaire detailing biographic characteristics, the site and duration, and specific issues related to the quality of research training was sent to identified trainees. Significant associations between success in achieving an academic appointment or OG support and several variables of training were identified. RESULTS: Eighty-six research fellows were trained. Responses were obtained from 43 of them. The demographic characteristics of the whole group and the respondents were similar. Of the respondents, 81% of trainees obtained academic appointments. Fellowships longer than 1 year were associated with higher rates of academic posting, and MRC-funded fellows had greater success rates of academic appointments. Of eligible trainees 63% have obtained OG support. None of the other variables examined predicted success. Of the trainees responding, 85% valued the fellowship very highly. CONCLUSIONS: The establishment of the additional research fellowships has fostered the development of career investigators in digestive sciences. The high success rate of former trainees in obtaining academic appointments and OG support suggests that the fellowship programs are effective and appropriately oriented. The structure of the current programs does not require substantial revision. OG support for new investigators appears now to lag substantially. PMID- 11266030 TI - Microalbuminuria screening for patients having type 2 diabetes mellitus: who wants to participate? AB - BACKGROUND AND OBJECTIVES: "Difficult-to-recruit" patients are sometimes less compliant with their care, are more reluctant to seek medical attention and less likely to survive than their "easy-to-recruit" counterparts. They also tend to be excluded from clinical trials. The aim of this paper was to evaluate whether such differences extend to patients' willingness to be screened for diabetic nephropathy in a family practice setting. DESIGN: A cross-sectional study. SETTING: A Canadian university family practice unit. PATIENTS: Two hundred and forty-seven patients with type 2 (adult-onset) diabetes mellitus as identified by computer searches of patient records of approximately 12,000 patients in the family practice unit. INTERVENTION: A cross-sectional secondary preventive screening program obtained urine samples from all patients with type 2 diabetes mellitus, regardless of patients' willingness to participate. MAIN OUTCOME MEASURE: The prevalence of micro- and macroalbuminuria. RESULTS: Of the 247 patients identified, 186 (75%) easy-to-recruit enrollees agreed to participate in screening and 61 (25%) difficult-to-recruit non-enrollees initially declined to be screened. The non-enrollees were subsequently evaluated by their own family physicians as part of routine clinical care and the results were captured for analysis. Overall rates of albuminuria were similar in the easy- and difficult-to recruit groups (31% versus 38%, p = 0.151). The main predictors of albuminuria were female sex (odds ratio [OR] = 2.1, p = 0.021), duration of diabetes in years (OR = 1.05, p = 0.023), current use of angiotensin-converting enzyme inhibitor (OR = 2.26, p = 0.008) and number of diabetic complications (OR = 1.45, p = 0.028). CONCLUSIONS: There is little difference in the prevalence of albuminuria related to patients' willingness to participate in a screening program. Therefore, there are no disproportionate gains for family practice researchers who aggressively seek difficult-to-recruit patients in this set ting. In contrast, primary care doctors should make every effort to ensure optimal care to diabetic patients regardless of a patient's initial hesitancy. PMID- 11266032 TI - Parenteral vitamin B12 reduces hyperhomocysteinemia in end-stage renal disease. AB - OBJECTIVE: The authors found considerably lower plasma total homocysteine (tHcy) concentrations in patients with end-stage renal disease (ESRD) on maintenance hemodialysis, who routinely received high-dose parenteral vitamin B12, than in comparable patients receiving much higher doses of folic acid but only replacement-dose oral vitamin B12. They therefore sought prospective evidence that high-dose parenterally administered vitamin B12 may partially ameliorate renal failure-associated hyperhomocysteinemia. DESIGN: Open phase 2 clinical trial. SETTING: Outpatient hemodialysis unit. PATIENTS: Fourteen clinically stable patients on maintenance hemodialysis with normal baseline serum vitamin B12 concentrations. INTERVENTION: Three parenteral injections of 1 mg vitamin B12 given at 4-week intervals. OUTCOME MEASURES: Plasma tHcy and serum vitamin B12 concentrations were measured before, during and 7 months after the termination of vitamin B12 therapy. RESULTS: The mean (and standard error) baseline plasma tHcy was 26.5 (1.8) micromol/L. The plasma tHcy value fell successively after each vitamin injection to reach a value of 23.6 (1.6) micromol/L 1 month after the final injection (p < 0.05), while the serum vitamin B12 concentration increased from 471 (42) pmol/L to 890 (74) pmol/L (p < 0.05). Seven months after the final injection, the serum B12 concentration had fallen and tHcy had risen to near their original values. CONCLUSIONS: Three monthly vitamin B12 injections modestly but distinctly reduced tHcy concentrations in hemodialysis patients whose prior vitamin B12 status was normal. Randomized placebo-controlled clinical trials of longer duration and using larger or more frequent parenteral doses are indicated to determine whether administration of this safe and inexpensive vitamin can improve hyperhomocysteinemia in ESRD. PMID- 11266033 TI - Gene mutations can produce polymorphisms that alter minimal daily micronutrient requirements. PMID- 11266034 TI - Methylenetetrahydrofolate reductase. PMID- 11266036 TI - Nocturnal hemodialysis: an update. PMID- 11266035 TI - Understanding pre-term birth. PMID- 11266037 TI - The outcome of patients presenting with crush syndrome after the Marmara earthquake. AB - In this study, we evaluated the clinical and laboratory data of the patients presenting after the Marmara earthquake. Crush syndrome was diagnosed in 60 patients (30 M, 30 F, mean age: 31.3+/-13.8 years). They were buried under the rubble for a mean period of 12.3+/-15.1 hours. On admission, 27 patients were oligoanuric and the mean serum creatinine, creatinine phosphokinase and potassium levels were 4.4+/-3.2 mg/dl, 18453.1+/-24527.2 IU/L, and 4.9+/-1.7 mEq/L, respectively. The most frequent site of trauma was the lower extremity. Dialysis treatment was initiated in 40 patients (19 M, 21 F; mean age: 32.7+/-13.0 years). Mean number of hemodialysis sessions/patient was 8.9+/-6.8. Nine (23%) patients among the dialyzed and 4 (20%) among the non-dialyzed died leading to an overall mortality of 21.6%. This low mortality rate suggests that the death rate from acute renal failure due to crush syndrome could be decreased by extensive follow up. PMID- 11266038 TI - Evaluation of stress on the autonomic nervous system of patients on mechanical circulatory assist. AB - PURPOSE: The power density values of high (HF; 0.15 to 0.4 Hz) and low frequency (LF; 0.04 to 0.15 Hz) components of arterial pressure were adopted for evaluation of stress on the autonomic nervous system of patients supported by mechanical circulatory assist. METHODS: Power spectral analysis (PSA) of arterial pressure signals was carried out, and trends in changes in the spectral components were observed in 12 patients on mechanical circulatory support (IABP and/or PCPS) and without the support (n=10). RESULTS: The normalized LF was depressed initially and increased gradually in both groups of patients except for four patients on mechanical circulatory assist. Three patients among them consequently died. CONCLUSIONS: Depressed LF represented marked stress on ANS and prolongation of the depression was related to poor outcome of patients. PSA of systemic blood pressure offers a reasonable tool for evaluation of stress on the ANS and for prediction of the prognosis of patients with circulatory assist devices. PMID- 11266041 TI - Selective apheresis--time for a change. PMID- 11266039 TI - Regulation of perfusion pressure during cardiopulmonary bypass using sevoflurane. AB - In hypothermic cardiopulmonary bypass (CPB), various vasodilators are used to control the perfusion pressure. These agents, however, often decrease the pressure excessively, and the low perfusion pressure may persist until the end of CPB. In this study we evaluate the safety and characteristics of the regulation of perfusion pressure during CPB using a volatile anesthetic, sevoflurane which has an extremely low partition coefficient. Twenty adult patients who underwent cardiac surgery were studied. Sevoflurane was applied by a vaporizer inserted into the oxygenator gas supply line. Pump flows were fixed at 2.4 L/min/m2 during the hypothermic period. Sevoflurane concentration was adjusted to keep mean arterial pressure (MAP) between 40 and 70 mmHg during CPB. Hemodynamic and metabolic parameters were measured and compared to the group we previously treated with chlorpromazine. In all cases, MAP could be maintained adequately. In the sevoflurane group, systemic vascular resistance indices (SVRI) during the rewarming period and at the end of CPB were higher, and doses of norepinephrine needed at the end of CPB were significantly lower than in the chlorpromazine group. The regulation of perfusion pressure during CPB using sevoflurane was safe and could easily maintain adequate SVRI. PMID- 11266040 TI - The efficacy of nafamostat mesilate on the performance of a hybrid-artificial liver using a polyurethane foam/porcine hepatocyte spheroid culture system in human plasma. AB - Nafamostat mesilate (FUT) is a protease inhibitor of complement activation. The present study investigates whether FUT protects porcine hepatocytes from being injured by human plasma in a multi-capillary polyurethane foam packed-bed culture system (MC-PUF) such as the hybrid-artificial liver (PUF-HAL). Human plasmas with 1 mM of added ammonia were perfused using a small-scale PUF-HAL with porcine hepatocytes. FUT was continuously infused (10 microg/ml, 50 microg/ml). The ammonia detoxification was maintained in human plasma for 24 hours and for 48 hours with FUT which suppressed the rapid increase of asparaginic acid aminotransferase (AST) and alanine aminotransferase (ALT). After 60 hours of perfusion, hepatocyte spheroids completely collapsed in the human plasma, but a small amount of hepatocyte spheroid was maintained by FUT. The effect of FUT was slightly greater at 50 microg/ml than at 10 microg/ml. Our results suggest that FUT has protective effects against porcine hepatocytes in human plasma, and our PUF-HAL using porcine hepatocytes can function in human plasma for about 48 hours with FUT. PMID- 11266042 TI - Evaluation of systemic metal diffusion after spinal pedicular fixation with titanium alloy and stainless steel system: a 36-month experimental study in sheep. AB - It is known that titanium alloys cause more extensive local metallosis due to fretting corrosion than stainless steel implants. The aim of the present study was to investigate possible systemic metal releases (Ti, Al, V, Cr, Ni) in sheep where L4-L5 were implanted with titanium alloy (Ti6Al4V, ASTM F 136) and stainless steel (AISI 316 L). 16 sheep were used: 8 were implanted with Ti6Al4V and 8 with stainless steel. At 6, 12, 24 and 36 months, the following examinations were performed: histology, atomic absorption spectrophotometry (AAS) and scanning electron microscopy (SEM), on liver, lung, kidney, brain, spleen and lumbo-aortic lymph nodes. Hair, urine and arteria blood samples were also analysed by AAS before implantation and at sacrifices. A histologic and ultrastructural study was performed on peri-implant tissues, too. Particular attention was paid to avoid contamination from dissection instruments or use of containers. In basal and in samples at 6 and 12 months, no metals were found in blood, urine, hair or other target tissues of the animals implanted with either Ti6Al4V or stainless steel. Regarding Al, V, Co and Ni, negative results in all tissues and body fluids were obtained also at 24 and 36 months. On the contrary, Ti traces were found in lumbo-aortic lymph nodes and lungs of one sheep only (10 and 30 ng/g, respectively) at 24 months. At 36 months, a systemic diffusion of Ti was observed in all tissues of both sheep instrumented with Ti6Al4V (2-16.5 ng/g), except for body fluids and hair. Metal research in target tissues by light and SEM micro-probe analysis provided negative results. Current data suggest that the amount of Ti found in organs after stable pedicular fixation is extremely low and not biologically available. This observation would lead us to exclude the hypothesis of any toxic reaction and such a release seems to be due to the passive diffusion through lymphatic fluids. Additional studies are needed to confirm if this long-term release of Ti particles might cause tissue damage. PMID- 11266043 TI - Tissue engineering of the small intestine by acellular collagen sponge scaffold grafting. AB - Tissue engineering of the small intestine will prove a great benefit to patients suffering from short bowel disease. However cell seeding in tissue engineering, such as fetal cell use, is accompanied by problems of ethical issues, rejection, and short supply. To overcome these problems, we carried out an experimental study on tissue engineering of the small intestine by acellular collagen sponge scaffold grafting. We resected the 5 cm long jejunum from beagle dogs and reconstructed it by acellular collagen sponge grafting with a silicon tube stent. The graft was covered with the omentum. At 1 month after operation, the silicon stent was removed endoscopically. Animals were sacrificed 1 and 4 months after operation, and were examined microscopically. Neo-intestinal regeneration was observed and the intestinal mucosa covered the luminal side of the regenerated intestine across the anastomosis. Thus, the small intestine was regenerated by tissue engineering technology using an acellular collagen sponge scaffold. PMID- 11266044 TI - Novel evidence on hepatitis B virus infection in dialysis. PMID- 11266045 TI - Establishing the causes of childhood mortality in Ghana: the 'spirit child'. AB - Communities in remote regions of northeast Ghana record some of the highest rates of under five mortality in West Africa (23.9 per 1000 children/year (Binka, Maude et al. (1995). International Journal of Epidemiology, 24(1), 127-135). The communities, isolated geographically and culturally from the main tribal groups in Ghana continue to adhere very strongly to traditional beliefs and practices. A qualitative study of both traditional and modern maternal and child health care systems in the area, demonstrated that almost 15% of deaths of infants under 3 months of age were due to a belief in chichuru or spirit children, resulting in infanticide. It is therefore a significant public health problem, which has to be addressed in programs for the control of child mortality. A modification of the verbal autopsy method is proposed to assist in the identification of non biomedical causes of death. PMID- 11266046 TI - The medicalization of female "circumcision": harm reduction or promotion of a dangerous practice? AB - In recent decades the practice of female "circumcision" has come under intense international scrutiny, often conceptualized as a violation of women's basic right to health. Although the adverse health consequences of female "circumcision" form the basis of opposition to the practice, anti-circumcision activists, as well as many international medical associations, largely oppose measures to improve its safety. The debate over medicalization of female "circumcision" has, up until now, been cast as a moral dilemma: to protect women's health at the expense of legitimating a destructive practice? Or to hasten the elimination of a dangerous practice while allowing women to die from preventable conditions? This paper seeks to re-examine this debate by conceptualizing medicalization of female "circumcision" as a harm-reduction strategy. Harm reduction is a new paradigm in the field of public health that aims to minimize the health hazards associated with risky behaviors, such as intravenous drug use and high-risk sexual behavior, by encouraging safer alternatives, including, but not limited to abstinence. Harm reduction considers a wide range of alternatives, and promotes the alternative that is culturally acceptable and bears the least amount of harm. This paper evaluates the applicability of harm reduction principles to medical interventions for female "circumcision," and draws parallels to other harm reduction programs. In this light, arguments for opposing medicalization of female "circumcision", including the assertion that it counteracts efforts to eliminate the practice, are critically evaluated, revealing that there is not sufficient evidence to support staunch opposition to medicalization. Rather, it appears that medicalization, if implemented as a harm-reduction strategy, may be a sound and compassionate approach to improving women's health in settings where abandonment of the practice of "circumcision" is not immediately attainable. PMID- 11266048 TI - Disease and dislocation: the impact of refugee movements on the geography of malaria in NWFP, Pakistan. AB - Studies of the health implications of refugee movements have generally focused on the effects of dislocation on the health of refugees and the impacts on health care provision at the destination. A somewhat more neglected aspect of the refugee-health research has been the impact of refugee flows on the geography of disease, i.e., how the spatial patterns of disease prevalence are modified through the influx and settlement of refugee populations. We examine this issue by examining the changing geography of malaria in Pakistan's North West Frontier Province (NWFP) between 1972 and 1997. Until the late 1970s, the highest incidence of malaria in the region was seen in the southern and eastern parts. During the 1980s, however, two and a half million Afghan refugees entered the NWFP and were housed in tented villages along the border and in some interior areas. As the decade progressed, there was a significant shift in the spatial pattern of malaria, with the regions of highest incidence shifting to the west and north, coinciding strongly with refugee concentrations. Our study draws attention to the manner in which refugee influx and settlement can alter the ecology of the disease system, leading to long-term changes in the geography of malaria. PMID- 11266047 TI - Stressors, resources, and distress among homeless persons: a longitudinal analysis. AB - Relations among stressors, resources, and psychological distress were examined using two waves of data obtained from a probability sample of homeless persons (N = 430) residing in a large, demographically diverse county in North California. The focus of research was to examine whether and how social resources and housing resources directly affect distress and mediate the impact of stress factors on depressive symptoms. Path analysis results revealed that levels of psychological distress were responsive to change in objective housing circumstances, with the attainment of domicile status being associated with fewer distress symptoms. Our findings, however, indicated only modest effects of social resources on psychological distress through direct effects and mediating effects of life stressors on distress. Overall, the study suggests that the relationships among stressors, resources, and distress for homeless persons may be understood within the same analytical framework for the general population. PMID- 11266049 TI - Determinants of condom use intentions of university students in Ghana: an application of the theory of reasoned action. AB - The study examined the applicability of the Theory of Reasoned Action to the study of condom use intentions of students at a university in southern Ghana. The data supported the model, explaining 33% of the variance in students' condom use intentions. Subjective norms and the perceived disadvantages of condom use were significant determinants of intention, with the former being more important. Respondents who intended to use condoms consistently ("intenders") and those with no such intentions ("non-intenders") were equally motivated to comply with the wishes of their significant referents (sexual partners, close friends, parents and medical doctors). The critical difference was that "intenders" consistently held a stronger belief than "non-intenders" that their significant referents approved of condom use. Significantly, whereas "intenders" believed that their sexual partners would approve of condom use, the "non-intenders" held the contrary belief that their partners would disapprove of such behavior. This suggests that AIDS education interventions targeting a similar audience like the university students in this study should shift their foci away from individuals alone and instead, focus simultaneously on individuals, their sexual partners and their broader social networks in order to enhance perceptions of peer acceptance of condom use. PMID- 11266050 TI - The impact of AIDS, immigration and housing overcrowding on tuberculosis deaths in Sao Paulo, Brazil, 1994-1998. AB - The objective of this paper was to describe the distribution of tuberculosis (TB) mortality by area in the municipality of Sao Paulo, Brazil, from 1994 to 1998, and to evaluate its statistical association with several population characteristics. We surveyed TB deaths grouped by residential area, at the district level, and we calculated the rates for these areas standardized by gender and age groups. We applied simultaneous autoregressive--SAR regression analysis (autocorrelated errors model) in order to fit a "stepwise" model correlating TB deaths with the variables of interest. Significant associations were found between TB mortality rates and AIDS mortality rates, overcrowding at the household level, social development (expressed by a socioeconomic index), and rates of foreign immigration and immigration from other Brazilian States. Regression analysis allowed us to estimate the frequency of TB deaths virtually attributable to co-infection with HIV at 22.37% (95% confidence interval: 12.15 41.17%). TB death rates and utilization of public health services were not statistically associated, suggesting a reduced effectiveness of programs directed at control of the disease. The correlation between TB death rates and deprivation, measured by the socioeconomic index, indicates higher mortality in underprivileged areas. The significance of the association between housing overcrowding and TB deaths, in contrast to the absence of association with district-level overcrowding, indicates that prolonged contact is needed for disease transmission. Although the influx of foreigners and national migrants to the city diminished after the 1980s, immigration rates have been significantly correlated with TB mortality, suggesting greater vulnerability of these population segments to the disease. PMID- 11266051 TI - Injury in children of low-income Mexican, Mexican American, and non-Hispanic white mothers in the USA: a focused ethnography. AB - Several studies indicate that rates of serious pediatric injury are higher among Hispanics than among non-Hispanic whites in the USA. To investigate possible contributory factors, we interviewed 50 Mexican, 30 Mexican American, and 30 non Hispanic white mothers in their own homes in the same low-income neighborhoods of Southern California. Mothers were identified via door-to-door canvassing in areas with high rates of pediatric injury. We observed household conditions and behaviors and obtained a detailed family history, including accounts of any occurrence of serious injury in a child under 5 years old, the highest-risk age group for pediatric injury. Results show that Mexican families were poorer, less educated, and lived in more hazardous and crowded conditions than did families in the other two groups. Nevertheless, they benefited from strong family bonds and a cultural tradition in which responsible older children typically supervise younger siblings. In contrast, a number of Mexican American and white mothers had been abused as children and were estranged from their own mothers; hence they lacked support and models of good parenting. There was much less self-reported smoking, drug use, and mental dysfunction among the Mexican mothers and their male partners as well as much less excessively active and/or aggressive behavior among their children. The nature of the injuries reported by the various groups seemed to reflect these differences. Appropriate interventions for each group are discussed. The study illustrates the importance of using ethnographic methods to examine the context of pediatric injury at the household level. PMID- 11266052 TI - Looking means listening: coordinating displays of engagement in doctor-patient interaction. AB - This article studies the coordination of patients' production of their primary complaint and doctors' orientation to the patient on the one hand and to medical records on the other. In specific environments the doctor's shift of focus from interaction with the patient towards reading or writing the medical records is received by the patients as problematic. It is suggested that disengaging from interaction and engaging in studying the medical records may leave the patient puzzled about whether the doctor is listening or not. Thus, paying attention to the coordination and timing of these shifts in orientation may help the doctors to develop more patient-centered ways of interacting with the patient. Furthermore, studying the coordination of verbal and non-verbal aspects in medical interaction may complement the current ideas on the constituents of the ideal model of 'patient-centeredness' in medical interaction. PMID- 11266053 TI - Negotiating control and meaning: home birth as a self-constructed choice in Finland. AB - Each society has its own consensual understanding of birth and its determinants: caregivers, location, participants and loci of decision-making, which in the Western world are based on biomedical knowledge. However, two competing cultural models of childbirth, the biomedical/technocratic model and natural/holistic model, mediate women's choices and preferences for the place and caregiver in childbirth. This article explores the way in which these cultural models of birth and the existing practical possibilities for choices shape women's and men's understanding of home birth. Based on interviews with 21 Finnish women and 12 Finnish men, the reasons for and experiences of planning and building toward a home birth are examined through an analysis of birth narratives. The analysis focuses especially on the women's definitions of what is 'natural' and their relationship with health services where biomedical practices and knowledge are the norm. The analysis shows that the notion of 'natural birth' holds various meanings in Finnish women's narratives namely self-determination, control, and trust in one's intuition. I seek to demonstrate that just as the biomedical management of childbirth exhibits distinct cross-cultural variation, so also does resistance to biomedical hegemony, as such resistance is strongly embedded in the local socio-cultural situation. PMID- 11266054 TI - Clinical, provider and sociodemographic determinants of the number of antenatal visits in England and Wales. AB - The objective of this study was to measure the independent effects of clinical factors and non-clinical factors, such as provider and sociodemographic characteristics, on the number of antenatal visits made by women in England and Wales. The study was based on a survey of the secondary case records of 20,771 women with singleton pregnancies who were delivered between 1 August 1994 and 31 July 1995. The women in the survey attended one of nine maternity units in Northern England and North Wales selected within those areas to reflect geographical variations, as well as variations in the size and teaching status of the institution. A multivariate Poisson regression model was developed to examine differences in the number of antenatal visits made by women with different clinical and non-clinical characteristics. After controlling for non-clinical factors, primiparous women identified as high risk at booking made 1.0% more visits than primiparous women identified as low risk at booking (p = 0.196). Multiparous women identified as high risk at booking made 3.5% more visits than their low risk counterparts (p<0.001). High risk-defining criteria during antenatal care led to a 0.3% weekly increase in the number of antenatal visits amongst primiparous women (p <0.001) and a 0.4% weekly increase in the number of antenatal visits amongst multiparous women (p < 0.001). Several notable results, not reported elsewhere in the literature, were revealed by the regression analyses. After all independent variables were controlled for, women who were booked into urban teaching hospitals made 10% fewer antenatal visits than the women who were booked into the urban non-teaching hospitals. Women of Pakistani origin made 9.1% fewer antenatal visits than women of white British origin. Similar results were revealed for women of Indian origin and women from other ethnic groups. Non-smokers made 6.0% more antenatal visits than smokers. The planned pattern of antenatal care, number of carers seen, gestation at first presentation and maternal age also had significant independent impacts on the number of antenatal visits. The study highlights the sizeable impact of non clinical factors on the antenatal care delivery process and indicates ways in which variations in antenatal care might be reduced. PMID- 11266055 TI - Social patterns of birth weight in Hong Kong, 1984-1997. AB - Fetal growth is an indicator of social inequalities in health that may have a long-term impact persisting into later life. Little is known about the social patterns of birth weight in Hong Kong. This is a study of live-born singletons from 1984 to 1997 in a Hong Kong birth registry. Ordinary least-squares regression and logistic regression are used to analyse birth weight and low birth weight (< 2500 g), respectively. A gradient of birth weight and prevalence of low birth weight is demonstrated according to mothers' educational attainment. In relation to babies of the most educated mothers, babies of the least educated mothers had a mean deficit of 46g in birth weight and an odds ratio of 1.56 of low birth weight (each P<0.05). This social gradient was hidden unless parity was adjusted for. Unexpectedly, migrants from mainland China delivered heavier rather than lighter babies (each P<0.05). Type of living quarters and parental relation were also related to birth weight and low birth weight (each P<0.05). Continuous monitoring of the social patterns of birth weight is recommended. PMID- 11266056 TI - What do patients expect of psychiatric services? A systematic and critical review of empirical studies. AB - The article reviews literature concerning patients' expectations of psychiatric care. Early research (pre-1980) is outlined, followed by a systematic review of 21 studies fulfilling specific inclusion criteria from 1980 onwards. Overall, patients expected to improve as a result of psychiatric treatment, and had higher expectations of the helpfulness of psychological and combined treatments than other interventions. Few studies considered expectations of the process of psychiatric care or determinants of expectations. The majority of studies focused on examining the relation between expectations and outcomes. There were indications that expectations of improvement were linked to clinical outcomes, although the relationship appeared to be complex. There was also some evidence that when expectations of the process of care were incongruent with the service provided, outcomes were poorer. The findings of studies in the systematic review were generally congruent with earlier work (pre-1980), although expectations of improvement appeared to be higher in the later studies. Interventions to prepare patients for what to expect were found to have beneficial effects on attendance, satisfaction and the accuracy of expectations about the process of psychiatric care. Future research should focus on developing valid and reliable measures for use in different settings, and on determining the mechanism by which expectations may relate to outcomes, including clinical outcomes, attendance and satisfaction. PMID- 11266057 TI - Morbidity and Irish Catholic descent in Britain. Relating health disadvantage to socio-economic position. AB - In common with some other ethnic and religious minorities whose forebears migrated from their country of origin, Irish Catholics in Britain are less well off than the host population in terms of socio-economic position and health. Results are presented from a Scottish study, where Catholic religion of origin mainly indicates Irish ancestry, and it is estimated that about one-third of the population is of significant Irish descent. In this study, excess of physical and mental health problems and disability have previously been reported for those of Catholic background, particularly in the eldest cohort (aged 56 in 1988), and have not been fully explained by health-related behaviour. In this paper, we examine a number of key health measures, namely self-assessed health, number of symptoms in the month prior to interview, sadness or depression, disability and lung function, and various indicators of socio-economic position (head of household social class, main source of income, car ownership, housing tenure and school-leaving age), which all show Catholic disadvantage. Using longitudinal results from the 723 respondents who completed interviews both at sweeps one (1988) and three (1995), it is estimated that about half of the morbidity excess amongst middle-aged Catholics in Glasgow can be explained by socio-economic disadvantage. The health and socio-economic position of white minorities and disadvantaged religious minorities like Catholics in Scotland should be monitored by a co-ordinated information strategy. PMID- 11266058 TI - Molecules and mediators of inflammation in equine heaves: mechanisms and markers of disease. PMID- 11266059 TI - Neosporosis: an emerging protozoal disease of horses. PMID- 11266060 TI - The role of subchondral bone in joint disease: a review. AB - Subchondral bone plays a role in the pathogenesis of osteochondral damage and osteoarthritis in horses and humans. Osteochondral fragmentation and fracture, subchondral bone necrosis and osteoarthritis are common diseases in athletic horses, and subchondral bone is now thought to play an integral role in the pathogenesis of these diseases. There have been numerous research efforts focused on articular cartilage damage and its pathogenesis, yet comparatively little effort focused on subchondral bone pathology or the coordinated disease states of the osteochondral tissues. The purpose of this report is to review the current understanding of osteochondral disease in all species and its application to equine research and practice. It can be concluded from this review that our current understanding of osteochondral disease is based on clinical and pathological sources; and that the lack of information about joint tissue adaptation and disease has hampered objective studies of osteochondral tissues. PMID- 11266061 TI - MMP-9 as a marker of inflammation in tracheal epithelial lining fluid (TELF) and in bronchoalveolar fluid (BALF) of COPD horses. AB - Gelatinolytic activity was analysed to study whether elevated activity previously found at the tracheal level of the respiratory tract of horses with chronic obstructive pulmonary disease (COPD) could also be found at the lower part of the respiratory tract. Furthermore, presence and significance of the gelatinolytic matrix metalloproteinases (MMPs) MMP-2 and MMP-9 in respiratory secretions of healthy and COPD horses were determined. Elevated gelatinolytic matrix metalloproteinases were detected in bronchoalveolar and tracheobronchial secretions from COPD horses. The main pathologically elevated MMP was characterised to be MMP-9. Significantly increased MMP-9 activities as measured by gelatin zymography and Western blotting were found in all the respiratory samples from COPD horses compared to healthy horses. Elevation of active MMP-9 paralleled with increased gelatinase-associated lipocalin levels. Bronchoalveolar lavage fluid (BALF) epithelial cells, macrophages, neutrophils and lymphocytes expressed MMP-9 immunoreactivity demonstrated by immunocytochemistry and MMP-9 mRNA was expressed by bronchial epithelial cells of lung tissue section shown by in situ hybridisation. MMP-2 seems not to play a major role in chronic lung inflammation. No clear differences in MMP-2 or MMP-14 (a potent MMP-2 activator) levels were found when comparing the samples from COPD or healthy horses. These results suggests that MMP-9 could serve as a potential diagnostic marker for the active ongoing tissue remodelling in the acute phase of equine COPD. Increased gelatinolytic activity could be found at both tested respiratory tract levels. Therefore, tracheal epithelial lining fluid (TELF) samples can usefully serve as diagnostic material for detection of increased levels of the main gelatinolytic MMP, MMP-9, representing the entire diseased lung. PMID- 11266062 TI - Infection of endothelial cells with equine herpesvirus-1 (EHV-1) occurs where there is activation of putative adhesion molecules: a mechanism for transfer of virus. AB - Evidence is presented to show that activation of endothelial and leucoyte adhesion molecules is a key step in transferring virus from infected leucocytes; and determines the restricted tissue tropism. A range of tissues from 2 experimentally infected mares in late pregnancy at 4 and 8 days after infection with EHV-1 were compared with those from normal pregnant and nonpregnant mares. Rabbit antisera to equine activated endothelial cell molecules were used to identify which tissues expressed these molecules in normal nongravid and gravid mares, and to investigate whether the range of tissues was altered in pregnant mares as a consequence of infection. The results indicated that the endothelium of the pregnant reproductive tract did express these molecules. In the 2 pregnant mares infected with EHV-1, the endothelial cells in the nasal mucosa also expressed these activated endothelial cell molecules. Therefore, the sites expressing these molecules closely correlated with those where virus infection of endothelial cells has been described and is consistent with experimental in vitro data, indicating that expression of these molecules is an essential stage in the transference of virus from leucocytes to endothelial cells. PMID- 11266063 TI - Upregulation of mRNA of interleukin-1 and -6 in subchondral cystic lesions of four horses. AB - This study investigated the potential association of interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) in subchondral cystic lesions (SCL) in horses. With the technique of in situ hybridisation in paraffin sections of fibrous tissue of SCL and quantitative real-time PCR in fresh frozen fibrous tissue and undecalcified bone sections of SCL embedded in acrylic resin, upregulation of mRNA of both cytokines could be demonstrated. mRNA of IL-1beta was upregulated at the periphery of the cystic lesion adjacent to normal bone, whereas IL-6 mRNA was upregulated within the fibrous tissue found within the centre of the SCL. It was concluded that both cytokines are associated in pathological bone resorption observed in SCL and, in combination with increased production of prostaglandin E2, may be responsible for the slow healing, maintenance or further expansion of the cystic lesions. PMID- 11266064 TI - Morphology of the nerve endings in laryngeal mucosa of the horse. AB - To discuss the significance of laryngeal sensation on various disorders of the horse, we studied the morphological and topographical characteristics of sensory structures in the laryngeal mucosa using immunohistochemistry and immunoelectron microscopy. Various sensory structures, i.e. glomerular endings, taste buds and intraepithelial free nerve endings, were found in the laryngeal mucosa by immunohistochemistry for protein gene product 9.5 (PGP 9.5) and neurofilament 200kD (NF200). Glomerular nerve endings were distributed mainly in the epiglottic mucosa; some endings were also found in the arytenoid region arising from thick nerve fibres running through the subepithelial connective tissue. Some terminals directly contacted the epithelial cells. Taste buds were distributed in the epithelium of the epiglottis and aryepiglottic fold. In the whole mount preparation, the taste buds were supplied by the terminal branching of the thick nerve fibres. In some cases, the taste buds were arranged around the opening of the duct of the epiglottic glands. The intraepithelial free nerve endings were found to be immunoreactive for substance P (SP) and calcitonin gene-related peptide (CGRP). These nerve endings were surrounded by the polygonal stratified epithelial cells in the supraglottic region, and by the ciliated cells in the subglottic region. The density of the intraepithelial free nerve endings was highest in the corniculate process of the arytenoid region and lowest in the vocal cord mucosa. The densities of CGRP- and SP-immunoreactive nerve endings in the arytenoid region were (mean +/- s.d.) 30.6+/-12.0 and 10.0+/-4.9 per unit epithelial length (1 mm), respectively and in the vocal fold mucosa, 1.1+/-0.9 and 0.8+/-0.7, respectively. Approximately one half of the CGRP immunoreactive nerve endings were immunoreactive for SP, and most SP-immunoreactive nerve endings were also immunoreactive for CGRP. Well-developed subepithelial plexus with numerous intraepithelial fibres were observed in flat or round mucosal projections that existed on the corniculate process of the arytenoid region. In conclusion, the laryngeal mucosa of the horse seems to have morphology- and/or location-dependent sensory mechanisms against various endo-and exogenious stimuli. PMID- 11266065 TI - The force and contact stress on the navicular bone during trot locomotion in sound horses and horses with navicular disease. AB - Mechanical overload due to poor conformation or shoeing has been suggested to contribute to the development of navicular disease. While studies have determined the compressive force exerted on the navicular bone in normal horses, this has not been reported for horses with navicular disease. Also, the force has not been converted to stress by correction for contact area. In this study we developed a technique for the determination of the contact area between the deep digital flexor tendon and the navicular bone in vivo, and used a forceplate system to determine the force and stress on the bone at trot in 6 normal and eight diseased horses. The mean +/- s.d. peak force and peak stress were 5.62+/-1.45 N/kg and 2.74+/-0.76 MPa for the normal group and 6.97+/-1.50 N/kg and 3.07+/-0.55 MPa for the navicular disease group. The peak force and peak stress were similar for both groups but the force and stress in the horses with navicular disease were approximately double control group values early in the stance phase. This was due to a higher force in the deep digital flexor tendon, which was attributed to a contraction of the deep digital flexor muscle in early stance in an attempt to unload the heels. PMID- 11266066 TI - The effect of bilateral palmar digital nerve analgesia on the compressive force experienced by the navicular bone in horses with navicular disease. AB - Horses with navicular disease have an increased load on the navicular bone in early stance. This has been suggested to be a response to pain in the heel region. Seven horses with clinical, radiographic and scintigraphic signs of navicular disease underwent forceplate and kinematic analysis before and after desensitisation of the heel region with a bilateral palmar digital nerve block. The compressive force exerted on the navicular bone during stance, and stride kinematics, were determined in each state. After regional analgesia of the palmar digital nerves (PDNB) the compressive force on the navicular bone was lower throughout stance. The mean +/- s.d. peak force at the beginning of stance was 7.05+/-1.10 N/kg before, and 6.46+/-1.15 N/kg after PDNB (P = 0.01) and at the end of stance the mean peak values were 5.00+/-2.05 N/kg before, and 4.39+/-1.65 N/kg after PDNB (P = 0.05). We explained this finding as indicating that the horse responds to heel pain (including pain in the navicular region) by contracting the deep digital flexor muscle to unload the heels. This increases the compressive load on the navicular bone, which may cause remodelling and, in some horses, damage to the overlying flexor cartilage, which is then painful and identified as navicular disease. This mechanism identifies navicular disease as a possible end point for a variety of heel related conditions. PMID- 11266067 TI - Relationships of age and shape of the navicular bone to the development of navicular disease: a radiological study. AB - Estimating the shape of the proximal articular border of the navicular bone and grading the radiological navicular bone condition (grades 3 and 4 representing the most severe changes), the aim of this study was to assess potential age related implications of the previously reported shape predisposition to navicular disease in 746 normal and 174 clinically affected Dutch Warmbloods age 3-19 years. A significant, age-independent, shape-grade association found in normal and affected horses emphasises the fundamental character of the shape predisposition to navicular disease. A significant age-related increase of the least susceptible shape prevalence was found in elderly normal horses. A shape independent low grades 3 and 4 prevalence (mean 15%) was found in normal horses, vs. a high grades 3 and 4 prevalence (mean 85%) in the affected horses. Therefore, the clinical manifestation of the disease is grade-rather than shape dependent. A significant age-related appearance of inverted flask-shaped channels and enthesophytes was found in the clinically affected horses. However, considering the significant shape-radiological features association previously reported in 3-year-old normal horses, this association may be shape- rather than age-dependent. PMID- 11266068 TI - External skeletal fixation in the management of equine mandibular fractures: 16 cases (1988-1998). AB - Fifty-three cases of equine mandibular fractures were managed surgically from 1988-1998, of which 16 (30%) were repaired by external skeletal fixation (ESF). Three surgical methods were utilised: transmandibular 4.76 or 6.35 mm Steinmann pins incorporated into fibreglass casting material or nonsterile dental acrylic (methyl methacrylate - MMA) bars reinforced with steel; transmandibular 9.6 mm self-tapping threaded pins +/- 4.76 or 6.35 mm Steinmann pins incorporated into MMA bars reinforced with steel; and 4.5 mm or 5.5 mm ASIF cortical bone screws incorporated into MMA bars reinforced with steel or a ventral MMA splint. Fourteen horses were presented to the hospital for fixator removal at an average of 56.2 days. At removal, fractures were stable and occlusion of incisor and cheek teeth was considered adequate. Complications of the procedure occurred in 3 horses. Two horses with persistent drainage and ring sequestra from pin tracts required curettage 4 or 5 months after ESF removal. A third horse required replacement of the original fibreglass ESF with MMA bars to regain access to open, infected wounds. Another horse required removal of the second premolar at the time of fixator removal because the tooth root had been damaged in the original injury. ESF for the surgical management of mandibular fractures in horses has produced good results, with incisive and cheek tooth alignment reestablished in all horses. Horses that were managed via ESF had a rapid return to full feed and did not require any supplementation via nasogastric tube or oesophagostomy to maintain bodyweight or hydration status. PMID- 11266069 TI - Intra-articular pressure profiles of the cadaveric equine fetlock joint in motion. AB - The study of the influence of motion and initial intra-articular pressure (IAP) on intra-articular pressure profiles in equine cadaver metatarsophalangeal (MTP) joints was undertaken as a prelude to in vivo studies. Eleven equine cadaver MTP joints were submitted to 2 motion frequencies of 5 and 10 cycles/min of flexion and extension, simulating the condition of lower and higher (double) rates of passive motion. These frequencies were applied and pressure profiles generated with initial normal intra-articular pressure (-5 mmHg) and subsequently 30 mmHg intra-articular pressure obtained by injection of previously harvested synovial fluid. The 4 trials performed were 1) normal IAP; 5 cyles/min; 2) normal IAP; 10 cycles/min; 3) IAP at 30 mmHg; 5 cycles/min and 4) IAP at 30 mmHg; 10 cycles/min. The range of joint motion applied (mean +/- s.e.) was 67.6+/-1.61 degrees with an excursion from 12.2+/-1.2 degrees in extension to 56.2+/-2.6 degrees in flexion. Mean pressure recorded in mmHg for the first and last min of each trial, respectively, were 1) -5.7+/-0.9 and -6.3+/-1.1; 2) -5.3+/-1.1 and -6.2+/-1.1; 3) 58.8+/-8.0 and 42.3+/-7.2; 4) 56.6+/-3.7 and 40.3+/-4.6. Statistical analyses showed a trend for difference between the values for the first and last minute in trial 3 (0.05>P<0.1) with P = 0.1 and significant difference (P = 0.02) between the mean IAP of the first and last min in trial 4. The loss of intra-articular pressure associated with time and motion was 10.5, 16.9, 28.1 and 28.9% for trials 1-4, respectively. As initial intraarticular pressure and motion increased, the percent loss of intra-articular pressure increased. The angle of lowest pressure was 12.2+/-1.2 degrees (mean +/- s.e.) in extension in trials 1 and 2. In trials 3 and 4, the lowest pressures were obtained in flexion with the joints at 18.5+/-2.0 degrees (mean +/- s.e.). This demonstrated that the joint angle of least pressure changed as the initial intra-articular pressure changed and there would not be a single angle of least pressure for a given joint. The volume of synovial fluid recovered from the MTP joints in trial 3 compared to 4 (trials in which fluid was injected to attain IAP of 30 mmHg) was not significantly different, supporting a soft tissue compliance change as a cause for the significant loss of intra-articular pressure during the 15 min of trial 4. The pressure profiles generated correlate well with in vivo values and demonstrated consistent pressure profiles. Our conclusions are summarised as follows: 1. Clinically normal equine MTP joints which were frozen and then later thawed were found to have mostly negative baseline intra-articular pressures, as would be expected in living subjects. 2. Alternate pressure profiles of the dorsal and plantar pouch at baseline intra-articular pressure document the presence of pressure forces that would support 'back and forth' fluid movement between joint compartments. This should result in movement of joint fluid during motion, assisting in lubrication and nutrition of articular cartilage. 3. If joint pressure was initially greater than normal (30 mmHg), as occurs in diseased equine MTP joints, joint motion further increased joint capsule relaxation (compliance) and, therefore, reduced intra-articular pressure. 4. Peak intra articular pressures reached extremely high values (often >100 mmHg) in flexion when initial pressure was 30 mmHg. Joint effusion pressures recorded for clinical MCP joints are frequently 30 mmHg. These IAP values are expected to produce intermittent synovial ischaemia in clinical cases during joint flexion. 5. Additional in vivo studies are necessary to confirm our conclusions from this study and to identify the contributions of fluid absorption and the presence of ischaemia in a vascularised joint. PMID- 11266070 TI - Neurological disease associated with EHV-1-infection in a riding school: clinical and virological characteristics. AB - An outbreak of neurological disease caused by EHV-1 infection is described with emphasis on diagnosis and prognosis for recumbent horses. In April 1995, an outbreak of the neurological form of Equine herpesvirus type 1 (EHV-1) occurred in a well-managed riding school with 41 horses: 34 horses showed a temperature spike and 20 some degree of neurological signs, of which 10 were nursed intensively in the indoor arena of the riding school for 3 to 20 days, 8 having to be maintained in slings for 2-18 days, while 9 needed bladder catheterisation b.i.d. for 2-16 days. Within the first 3 days, one horse was subjected to euthanasia and another horse died. Postmortem examination revealed a mild vasculitis with perivascular mononuclear cuffing and axonal degeneration in the central nervous system. Clinical diagnosis was confirmed by serology and virology: 28 horses seroconverted in one or more tests during the outbreak, whereas 12 had already high CF and SN titres in the first sample, suggestive of recent infection. Virus was isolated from nasal swabs of 4 horses, and identified as EHV-1 with type-specific monoclonal antibodies. Restriction enzyme analysis revealed that the EHV-1 strains from this outbreak belonged to genome type EHV 1.IP. The electropherotypes were identical to those from another, epidemiologically unrelated, outbreak of neurological disease 2 months earlier. The timing of the temperature spikes and seroconversions indicated that the infection was probably introduced by a horse purchased 3 weeks before neurological signs occurred. At follow-up one year later, the 10 horses that showed mild neurological signs had recovered completely. Of the 8 horses that survived intensive care, 3 had returned to around their former performance level (2 of which had been in slings), while the other 5 had become pasture-sound. At follow-up 4 years later, all pasture-sound horses had been subjected to euthanasia because of persistent mild ataxia and incontinence. In conclusion, the prognosis for recumbent horses due to EHV-1 infection is grave. For virological diagnosis, extensive and strategic sampling of febrile in-contact horses is required, and the EHV-1-specific glycoprotein G (gG) ELISA is a valuable tool for specific serological diagnosis of EHV-1 infection causing neurological disease. PMID- 11266071 TI - Assessment of the rate of solid-phase gastric emptying in ponies by means of the 13C-octanoic acid breath test: a preliminary study. AB - The aim of this study was to assess the feasibility of applying the 13C-octanoic acid breath test for assessment of gastric emptying in ponies by investigating the pattern of 13C enrichment in breath following the administration of a test meal +/- 13C-octanoic acid. After a 14 h fast, the ponies received either no meal (Test I) or a standardised test meal labelled with 0 mg (Test II), 125 mg (Test III), 250 mg (Test IV) or 500 mg (Test V) 13C-octanoic acid. For each test (I-V), exhaled breath samples were collected in duplicate at 1 h and immediately before ingestion of the test meal and at frequent intervals thereafter for 12 h. Breath samples were analysed by continuous flow isotope ratio mass spectrometry. Three indices of breath 13C-enrichment were computed; half dose recovery time (t1/2), gastric emptying coefficient (GEC) and time to peak breath 13C-enrichment t(max). For Tests I and II, the ratio of 13CO2:12CO2 remained stable for the duration of the sampling period. For Tests III, IV and V, an increase in the ratio of 13CO2:12CO2 was detected. The test was reproducible within individuals, and intersubject variation was low. Further validation studies of this noninvasive technique are justified. PMID- 11266072 TI - Prevalence of antibodies to Neospora caninum in horses in France. PMID- 11266073 TI - Prevalence of placentophagia in horses. PMID- 11266074 TI - Salivary and plasma concentration of cortisol in normal horses and horses with Cushing's disease. PMID- 11266075 TI - Hepatoblastoma in a foal. PMID- 11266076 TI - Genomic organization of the human CYP3A locus: identification of a new, inducible CYP3A gene. AB - Proteins encoded by the human CYP3A genes metabolize every second drug currently in use. The activity of CYP3A gene products in the general population is highly variable and may affect the efficacy and safety of drugs metabolized by these enzymes. The mechanisms underlying this variability are poorly understood, but they include gene induction, protein inhibition and unknown genetic polymorphisms. To better understand the regulation of CYP3A expression and to provide a basis for a screen of genetic polymorphisms, we determined and analysed the sequence of the human CYP3A locus. The 231 kb locus sequence contains the three CYP3A genes described previously (CYP3A4, CYP3A5 and CYP3A7), three pseudogenes as well as a novel CYP3A gene termed CYP3A43. The gene encodes a putative protein with between 71.5% and 75.8% identity to the other CYP3A proteins. The highest expression level of CYP3A43 mRNA is observed in the prostate, an organ with extensive steroid metabolism. CYP3A43 is also expressed in several other tissues including liver, where it can be induced by rifampicin. CYP3A43 transcripts undergo extensive splicing. The identification of a new member of the CYP3A family and the characterization of the full CYP3A locus will aid efforts to identify the genetic variants underlying its variable expression. This, in turn, will lead to a better optimization of therapies involving the numerous substrates of CYP3A proteins. PMID- 11266077 TI - Rabbits possess a serum paraoxonase polymorphism similar to the human Q192R. AB - Serum paraoxonase (PON1) is a high-density lipoprotein (HDL)-associated enzyme that hydrolyses aromatic esters, organophosphates and lactones and can protect low-density lipoprotein (LDL) against oxidation. These properties are influenced by a well-characterized polymorphism (Q192R) in human PON1. We now report the identification and characterization of a phenotypically similar, but genetically distinct polymorphism in rabbit PON1. This polymorphism in rabbits was detected by phenotyping sera obtained from 16 inbred rabbit strains and 20 outbred New Zealand White rabbits by paraoxonase/arylesterase activity. The genetic basis of the rabbit polymorphism was determined by DNA sequencing and found to reside in a region distinct from the human Q192R and M55L polymorphisms. Three variant nucleotides within exon 4 (corresponding to P82S, K93E and S1O1G) were found to segregate with the observed rabbit PON1 phenotypes (rPON1A and rPON1B). The rPON1A and rPON1B proteins were purified and compared to the two human isoforms (192Q and 192R). The human and rabbit PON1s displayed similar characteristics with respect to physical properties and substrate specificity. However, rPON1A and rPON1B hydrolysed a variety of substrates at different rates. The rPON1A was also at least three-fold more efficient at protecting LDL from oxidation than rPON1B. Our characterization of a rabbit PON1 polymorphism provides useful insights into important functional residues in PON1. In addition, due to the observed similarities between the rabbit and human polymorphisms, the rabbit may serve as a good model to examine the effect of human PON1 polymorphisms in disease development. PMID- 11266078 TI - Further evidence of human leukocyte antigen-encoded susceptibility to clozapine induced agranulocytosis independent of ancestry. AB - To further examine the human leukocyte antigen (HLA)-encoded genetic susceptibility to clozapine-induced agranulocytosis (CA) we performed HLA genotyping in a sample of German schizophrenic patients, who suffered from this haematotoxic side-effect. Thirty-one schizophrenic patients with CA (17 women and 14 men) and 77 schizophrenic comparison subjects (40 women and 37 men) were included in the study. HLA-genotyping included identification of major histocompatibility complex (MHC) class I (HLA-A, B, Cw) and class II (HLA-DR, DQ) antigens. CA was significantly associated with HLA-Cw*7 (P<0.02), DQB*0502 (P<0.04), DRB1*0101 (P<0.03) and DRB3*0202 (P<0.02). These HLA-haplotypes are also partly linked to other diseases with a strong genetic background. All other antigens revealed no association to this haematotoxic reaction. In addition, we did not find gender-related effects, whereas age seemed to be a further major risk factor of CA (P<0.0003). Thus, HLA loci may serve as genetic marker to identify subjects of different ethnic subgroups prone to this severe idiosyncratic drug reaction of clozapine. Further studies are needed to investigate whether these associations with CA are due to causal involvement or linkage disequilibrium. PMID- 11266079 TI - Characterization of (+/-)-bufuralol hydroxylation activities in liver microsomes of Japanese and Caucasian subjects genotyped for CYP2D6. AB - Twenty-four genetic polymorphisms in the CYP2D6 gene were analysed in liver DNA samples of 39 Japanese and 44 Caucasians and compared with CYP2D6 protein levels and bufuralol 1'- and 6-hydroxylation activities in liver microsomes of these human samples. We detected 13 types of CYP2D6 genetic polymorphisms and classified these into 20 genotypes; nine types were found in Japanese and 14 types in Caucasian samples. CYP2D6*10B, but not CYP2D6*10A, was the most frequent (34.6%) in Japanese. In Caucasians, several CYP2D6 polymorphisms including CYP2D6*4, *4D, *4E, *4L, *3, *9, *5 and *2E (frequencies of 6.8, 3.4, 4.5, 9.1, 1.1, 2.3, 2.3 and 4.5%, respectively) were detected. A Caucasian having CYP2D6*3/*5 had a protein with slower gel mobility (immunoblotting with anti CYP2D6 antibody) and very low activity for bufuralol 1'-hydroxylation. Five Caucasian samples (CYP2D6*4/*4, *4/*4L, or *4D/*4L) had no measurable CYP2D6 protein and very low bufuralol 1'-hydroxylation activities. Seven Japanese subjects with CYP2D6*10B/*10B had CYP2D6 protein at levels of approximately 20% of those present in humans with CYP2D6*1 and *2 and catalysed bufuralol 1' hydroxylation at low rates. Kinetic analysis of bufuralol 1'- and 6-hydroxylation indicates that (i) the Km values for 1'-hydroxylation were lower in individuals with CYP2D6*1/*1, *1/*2, *1/*2X2, and *2/*2 than those with CYP2D6*4/*4, *4/*4L, *4D/*4L, or *10B/*10B and Vmax values tended to be higher in the former groups (*1, *2), and (ii) individuals with heterozygous CYP2D6*1/*4D, *1/*4L, and *1/*5 had relatively high Vmax/Km ratios, whereas individuals with heterozygous CYP2D6*1/*9, *2/4D, *2/*5, *2/*10B, *2E/*4E, *3/*5, *4L/*9, and *10B/*39 had lower Vmax/Km ratios for bufuralol 1'-hydroxylation. Quinidine inhibited bufuralol 1'-hydroxylation in liver microsomes, particularly at low substrate concentrations, in individuals with CYP2D6*1/*1, and 1/1*2, but not those with CYP2D6*4/*4 and very slightly in individuals with CYP2D6*10B/*10B. The latter two groups were found to be more sensitive to alpha-naphthoflavone than the former groups, indicative of the contribution of CYP1A2. These results support the view that CYP2D6*3, *4, *4D, and *4L are major genotypes producing poor metabolizer phenotypes in CYP2D6 in Caucasians, whereas CYP2D6*10B is a major factor in decreased CYP2D6 protein expression and catalytic activities in Japanese. PMID- 11266080 TI - Relevance of N-acetyltransferase 1 and 2 (NAT1, NAT2) genetic polymorphisms in non-small cell lung cancer susceptibility. AB - The highly polymorphic N-acetyltransferases (NAT1 and NAT2) are involved in both activation and inactivation reactions of numerous carcinogens, such as tobacco derived aromatic amines. The potential effect of the NAT genotypes in individual susceptibility to lung cancer was examined in a hospital based case-control study consisting of 392 Caucasian lung cancer patients [152 adenocarcinomas, 173 squamous cell carcinomas (SCC) and 67 other primary lung tumours] and 351 controls. In addition to the wild-type allele NAT1*4, seven variant NAT1 alleles (NAT1*3, *10, *11, *14, *15, *17 and *22) were analysed. A new method based on the LightCycler (Roche Diagnostics Inc.) technology was applied for the detection of the polymorphic NAT1 sites at nt 1088 and nt 1095. The NAT2 polymorphic sites at nt 481, 590, 803 and 857 were detected by polymerase chain reaction restriction fragment length polymorphism or LightCycler. Multivariate logistic regression analyses were performed taking into account levels of smoking, age, gender and occupational exposure. An increased risk for adenocarcinoma among the NAT1 putative fast acetylators [odds ratio (OR) 1.92 (1.16-3.16)] was found but could not be detected for SCC or the total case group. NAT2 genotypes alone appeared not to modify individual lung cancer risk, however, individuals with combined NAT1 fast and NAT2 slow genotype had significantly elevated adenocarcinoma risk [OR 2.22 (1.03-4.81)] compared to persons with other genotype combinations. These data clearly show the importance of separating different histological lung tumour subtypes in studies on genetic susceptibility factors and implicate the NAT1*10 allele as a risk factor for adenocarcinoma. PMID- 11266081 TI - A novel deletion in the flavin-containing monooxygenase gene (FMO3) in a Greek patient with trimethylaminuria. AB - Mutations of the flavin-containing monooxygenase type 3 gene (FMO3) that encode the major functional form present in adult human liver, have been shown to cause trimethylaminuria. We now report a novel homozygous deletion of exons 1 and 2 in an Australian of Greek ancestry with TMAuria, the first report of a deletion causative of trimethylaminuria. The deletion occurs 328 bp upstream from exon 1. The 3'-end of the deletion occurs in intron 2, 10013 base pairs downstream from the end of exon 2. The deletion is 12226 bp long. For the proband homozygous for the human FMO3 gene deletion, it is predicted that in addition to loss of monooxygenase function for human FMO3 substrates, such as TMA and other amines, the proband will exhibit decreased tolerance of biogenic amines, both medicinal and those found in foods. PMID- 11266082 TI - Polymorphism of the ABC transporter genes, MDR1, MRP1 and MRP2/cMOAT, in healthy Japanese subjects. PMID- 11266083 TI - Hypertension in black people: study of specific genotypes and phenotypes will provide a greater understanding of interindividual and interethnic variability in blood pressure regulation than studies based on race. AB - Hypertension is more frequent and more severe in some Black populations. Although many studies have focused on hypertension in black people in an attempt to understand the genetic and environmental factors that regulate blood pressure, this approach has not been productive. Study of the relationship between specific phenotypes and genotypes, both within and across ethnic groups, is more likely to advance our understanding of the regulation of blood pressure than studies focused on race and blood pressure. PMID- 11266084 TI - DNA tumor vaccines. AB - A new generation of vaccines are being developed to induce immune responses that fight off infectious agents, or erradicate cancerous cells. These new vaccines are based on a plasmid vector, which in transfected mammalian cells cause constitutive high-level expression of the target antigen. Expression of the target antigen, in turn, can induce a full-range of immunologic responses, including cell-mediated killing, cell-mediated cytokine release and the production of antigen-specific antibodies. Through molecular techniques, these nucleic acid vaccines can be enhanced to increase target antigen expression and facilitate antigen presentation. Additionally, genetic adjuvants expressed simultaneously with the target antigens can induce the immune responses to disease-associated antigens. The ease with which these genetic vaccines can be generated and the potency of their ability to generate immune-mediated responses make them highly effective, which creates hope for developing effective treatment and prevention of various diseases, most notably cancer. PMID- 11266085 TI - CD8+ T cell suppressor factors and the control of infection, replication and transcription of human immunodeficiency virus. AB - CD8+ T cells have been shown to produce factors which modulate HIV-1 replication in both T cells and monocytic cells. Examination of the literature reveals that this modulation may occur by the production of beta-chemokines which block viral entry. However, another CD8+ T cell-derived factor(s) targets the replication of HIV-1 at the level of transcription. CD8+ T cell factors strongly suppress replication at the level of transcription in T cells and T cell lines, the factors enhance both replication and transcription in cells of the monocyte/macrophage lineage. The enhancement of transcription and replication, which is pertussis toxin sensitive is induced by increased production of TNF alpha by the target cells. Thus, CD8+ T cells produce factors which mediate effects on transcription and replication of HIV-1 in a cell type-dependent manner. In this review a summary of the effects of chemokines and CD8-derived factors on HIV-1 transcription and replication is presented focusing on the cellular pathways which may mediate their effects on HIV transcription and replication in different cell types. The virus-host cell interactions that participate in the persistent replication of HIV in macrophages and the suppression of these functions in T cells require definition. The identification of CD8+ T cell factors which exert these controls on HIV-1 may lead to promising new therapies for HIV infection. PMID- 11266086 TI - Phage display as a tool for rapid cloning of allergenic proteins. AB - Allergic diseases represent an immune disorder associated with the production of immunoglobulin E (IgE) against normally innocuous antigens (allergens). Almost 20% of the population in industrialized countries suffer from type I allergic symptoms such as allergic rhinitis, conjunctivitis, urticaria or asthma. Although the mechanisms responsible for these allergic reactions are quite well understood, knowledge about the repertoire of molecules able to elicit type I symptoms is still limited. To clone and characterize entire allergen repertoires from complex allergenic sources in a fast and efficient way, new technologies are required. The phage surface display of cDNA libraries described here has proven to be a versatile cloning system to selectively isolate allergens physically linked to their genetic information. The screening of cDNA libraries displayed on phage surfaces with immobilised serum IgE from allergic patients reduces the time required for the selection of candidate clones to a few weeks. Robot-assisted high-throughput screening of the enriched library provides a fast and cost effective way to isolate complete allergen repertoires. The biotechnological production of recombinant allergens derived from these sequences bears a high potential for the improvement of the diagnosis of allergic diseases. PMID- 11266087 TI - Inhibition of angiogenesis in the treatment of tumors. AB - Angiogenesis is essential for tumor progression, growth and metastases. Many substances present in a normal organism can inhibit or stimulate the process of new vessel formation in tumors. The use of natural or synthetic angiogenesis inhibitors as anticancer drugs is currently under intense investigation. Such agents can have lower toxicity and are less likely to generate drug resistance than conventional cytotoxic drugs. Clinical trials are now underway to develop optimum treatment strategies for antiangiogenic drugs. This paper reviews the present achievements in preclinical and clinical studies with antitumor drugs based on inhibitors of angiogenesis. PMID- 11266088 TI - Immunomodulatory role of the corticotropin-releasing factor. AB - Corticotropin-releasing factor (CRF) was originally identified as a hypothalamic peptide which stimulates secretion of the hypophyseal adrenocorticotropin hormone. CRF exhibits its actions through G protein-dependent seven membrane domain receptors. Two subtypes of CRF receptors (CRF-R1 and CRF-R2) have been characterized thus far. CRF and its receptors were found in a number of brain regions, where they function by neuromodulation and also in several peripheral organs. Besides CRF, another naturally occurring CRF-like peptide, urocortin, has been characterized. In the immune system, CRF and CRF-R1 were so far detected at both mRNA and protein levels in several lymphoid organs and at sites of inflammation. Locally injected CRF was shown to modulate the severity of inflammation. This effect was not only a result of hemodynamic changes known to be induced by CRF or by activation of the hypothalamo-pituitary-adrenal axis, as CRF-binding sites were also found on immune cells. CRF was shown to directly modulate secretion of cytokines and neuropeptides, proliferation, chemotaxis and degranulation of purified macrophage and lymphocyte populations in vitro. Functional CRF-R was more recently demonstrated also on polymorphonuclear cells and significant amounts of CRF were shown to be produced in lymphoid organs, or delivered to lymphoid organs by peripheral nerves. Taken together, the experimental results obtained so far strongly point to the importance of CRF as a signaling molecule in lymphoid tissues and at the sites of inflammation. PMID- 11266089 TI - Expression and functional significance of CTLA-4, a negative regulator of T cell activation. AB - The generation of an effective immune response involves antigen-specific T cell expansion and differentiation of effector function. T cell activation requires at least two distinct signals, including signaling via the antigen-specific T cell receptor (TCR) and a costimulatory pathway. Antigen stimulation of T cells can lead either to a productive immune response, characterized by proliferation, differentiation, clonal expansion and effector function, or, in the absence of an appropriate costimulation, to a state of long-lasting unresponsiveness, termed anergy. Anergic T cells fail to proliferate and secrete cytokines in response to secondary stimulation. The interaction between the costimulatory molecule CD28 on T cells and members of the B7 family on antigen-presenting cell results in upregulation of T cell proliferation and cytokine production and induces the expression of the anti-apoptotic protein Bcl-xl. Based of these findings, the two signal requirement model for T cell activation is generally accepted today. The negative regulatory mechanisms during T cell activation are not well understood, but they are crucial for the maintainance of lymphocyte homeostasis. For several years the functional role of the enigmatic CD28 homologue cytotoxic T lymphocyte antigen-4 (CTLA-4) in T cell activation has been both obscure and controversial. CTLA-4 was initially supposed to provide a costimulatory signal in conjunction with TCR/CD3 signaling. Today we know that CD28 and CTLA-4 molecules may have diametrically opposed functions: signaling via CD28, in conjunctive with TCR, is required for T cell activation, while signaling via CTLA-4 is a negative signal that inhibits T cell proliferation. How the T cell integrates signals through the TCR/CD3 complex, CD28 and CTLA-4 to initiate, maintain and terminate antigen specific immune response is in fact not fully clarified. In this review, we will focus on the emerging role of CTLA-4 as a negative regulator of T lymphocyte activation and its role in the dynamic interplay of activatory and inhibitory signals. PMID- 11266090 TI - The murine Ly49 family: form and function. AB - The activity of natural killer (NK) cells is regulated by surface receptors that recognize class I MHC. Murine NK cells express a large family of lectin-related receptors (Ly49s) to perform this function, while human NK cells utilize a separate group of proteins containing Ig-related domains (KIRs). Although these receptor families are not structurally related, the Ly49 family appears to be the functional equivalent of human KIRs, since it uses similar signal transduction pathways for either activation or inhibition of NK cell function. Therefore, lessons learned from the study of the murine MHC class I receptor system may be relevant to human NK function. This review summarizes the current state of knowledge of the Ly49 family. PMID- 11266091 TI - A method for directly determining the number of dendritic cells and for evaluation of their function in small amounts of human peripheral blood. AB - Bone marrow-derived dendritic cells (DC) are highly potent antigen-presenting cells (APC) capable of initiating primary responses of naive T lymphocytes to antigen. Studies on DC in disease have been impeded by the lack of a defined method for accurate DC counting and for evaluation of their function in a small amount of blood. In order to detect and enumerate DC in whole peripheral blood preparations, we applied a direct two-color immunofluorescence method. Blood from healthy donors was stained with a mixture of fluorescein isothiocyanate (FITC) conjugated monoclonal antibodies (mAbs) recognizing lineage-associated molecules (CD3, CD14, CD16, CD20, CD57) and phycoerythrin (PE)-conjugated anti-HLA-DR mAb. DC were identified as lineage marker negative (lin-), HLA-DR highly positive cells. The mean percentage of these cells present in peripheral blood leukocytes (PBL) was 0.54%, and the mean absolute DC count was 31.4 x 10(6)/l of blood. DC stained directly in whole blood were heterogeneous with regard to their expression of CD2 and CD4 molecules, and did not express CD80 and CD83 molecules. Expression of CD80 and CD83 on DC was induced following a multistep isolation procedure, including overnight culture. We demonstrated a significant primary proliferative response to keyhole limpet hemocyanin (KLH) in cultures of peripheral blood mononuclear cells (PBMNC). Since primary proliferative response to neoantigens is entirely dependent on DC as APC, the cultures of unseparated PBMNC stimulated with KLH can be used to evaluate DC function in a relatively simple test. This test does not require previous isolation of DC and T lymphocytes and, therefore, can be performed on a small amount of blood. The elaborated flow cytometric method of DC counting in blood and the proliferative test of DC-dependent primary response to neoantigen are currently being applied in an ongoing study on the effect of chemotherapy on DC number and function in cancer patients. PMID- 11266092 TI - Malformations of angiogenesis in the low differentiated human carcinomas. immunohistochemical study. AB - Our previous observations showed that the perivascular mesenchyma of the thin walled vessels (capillaries) in cancers may be the source of organ-specific stem cells. We suggested that the cells forming vascular channels in altered stroma participate in the tumor development. This study was designed to examine the distribution of the vessels and their appearance in the breast, lung and colon cancers. Using immunohistochemical methods, we have shown that in the low differentiated tumors both CD31 and factor VIII antigens may be expressed in capillaries chiefly on the periphery of neoplastic foci. Many of these vessels were discontinuous, with interruptions or unformed tubules. Sporadically, CD31 protein and factor VIII antigens were not expressed in capillaries inside the very low differentiated cancer cases. It is difficult to assess by immunohistochemical means whether the vascular malformations are the primary or secondary phenomena in the malignancy and why these abnormalities were especially visible in some low differentiated cancers. PMID- 11266093 TI - TNF-alpha, IL-6 and their soluble receptor serum levels and secretion by neutrophils in cancer patients. AB - Simultaneous evaluation of cytokines and their soluble receptor production and the serum levels can be helpful in understanding the local and systemic immune response of a tumor-bearing host. In the present study we examined serum levels of TNF-alpha, IL-6 and their soluble receptors: sTNFRp55, sTNFRp75 and sIL-6R confronted with their production by the polymorphonuclear neutrophils (PMN) from cancer patients. Examinations were carried out in patients with adenocarcinoma breast cancer and squamous cell carcinoma of the oral cavity and related to the clinical course and to different phases of therapy. Secretion of IL-6, sTNFRp55 and sTNFRp75 by PMN appeared to be dependent on tumor type, clinical progression of disease as well as on therapy, suggesting a significant role of these cells at different phases of the immune response to cancer associated with these mediators. Changes in values of TNF-alpha, IL-6 and their soluble receptors in sera of both cancer groups, dependent on tumor type, clinical progression and cancer therapy, could have a diagnostic and prognostic role in cancer disease. PMID- 11266094 TI - Cytokine production in whole blood cell cultures of patients with B-lineage acute lymphoblastic leukemia. The influence of granulocyte-macrophage colony stimulating factor. AB - We investigated the levels of 6 different cytokines in the sera of 10 newly diagnosed patients with B cell lineage acute lymphoblastic leukemia (ALL) and detected a significant increase in IL-6 and IFN-alpha serum levels in comparison to that of healthy controls. Whole blood cell cultures of 10 ALL patients and 20 control individuals were induced with classical cytokine inducers, such as virus, PHA and LPS, and their ability to produce 9 different cytokines was compared. Blood cells of ALL patients produced significantly less IL-1alpha, IL-1beta, IL 10 and TNF-alpha than control cells and not significanly lower levels of IL-6, but comparable with control levels of IL-2, IL-4. rHuGM-CSF added to cell cultures 24 h before induction significantly enhanced the production of IL 1alpha, IL-1beta and TNF-alpha in controls, but only IL-1alpha and IL-1beta in the blood cell cultures of patients with ALL. GM-CSF did not significantly influence the production of IFN-alpha, IFN-gamma, IL-2, IL-4 and IL-10 in the control cells and the cells of ALL patients. The patients examined differed not only in the expression of CD10 and CD34 antigens on blast cells, but also in the reaction to GM-CSF treatment, which was found as very high standard deviation values. We suppose that these differences can partially explain the different effects of GM-CSF when used to ameliorate neutropenia of ALL patients after chemotherapy and to reduce the incidence of microbial infections. PMID- 11266095 TI - Estimating separately personal exposure to ambient and nonambient particulate matter for epidemiology and risk assessment: why and how. PMID- 11266096 TI - Estimating separately personal exposure to ambient and nonambient particulate matter for epidemiology and risk assessment: why and how. PMID- 11266097 TI - Estimating separately personal exposure to ambient and nonambient particulate matter for epidemiology and risk assessment: why and how. PMID- 11266098 TI - Movement and deposition of two organophosphorus pesticides within a residence after interior and exterior applications. AB - Post-application temporal and spatial distributions of two organophosphorus pesticides, diazinon (O,O-diethyl O-[6-methyl-2-(1-methylethyl)-4-pyrimidinyl] phosphorothioate, CAS No. 333-41-5) and chlorpyrifos [O,O-diethyl-O-(2-isopropyl 6-methyl-4-pyrimidinyl) phosphorothioate, CAS No. 2921-88-2], were monitored after homeowner applications for indoor and outdoor insect control. Samples of indoor air, vacuumable carpet dust, carpet dislodgeable residues, deposits on bare floors, table tops and dinnerware, surrogate food, and residues on children's hands and toys were taken before and up to 12 days after treatments in the family room, kitchen, and child's bedroom. Results from the study demonstrate the nature and magnitude of translocation of pesticides from the areas of application to surfaces accessible for human contact and permit comparisons of potential exposures via respiration and dermal contact/oral ingestion. Potential indoor inhalation exposures were estimated to be as high as 0.5 microg/kg/day for diazinon applied indoors and 0.05 microg/kg/day for chlorpyrifos applied to the outside perimeter of the house. While ingestion of carpet dust at the rate of 100 microg/day would have added a maximum of only approximately 0.01 microg/kg/day to the daily dose, residues found on the children's hands suggest that repeated mouthing could have contributed as much as 1-1.5 microg/kg/day. These estimates are below the U.S. Environmental Protection Agency (EPA) reference dose for chlorpyrifos, but exceed those for diazinon. PMID- 11266099 TI - Evaluation of health and safety risks in municipal solid waste recycling plants. AB - Designers and managers of recycling processing plants need to be informed about the inherent occupational health and safety risks; however, there are few studies in the literature describing these risks. Therefore, the objectives of this project were to document the biological, chemical, physical, and ergonomic risks in three household-waste recycling plants. Bioaerosols (molds and bacteria) were measured using the methodology recommended by the American Society for Testing and Materials (ASTM). Chemical contaminants and physical agents suspected of being present in this type of environment were measured using the standard methods of the Quebec Occupational Health and Safety Research Institute (IRSST). The ergonomic part of the study identified the work requirements and risk factors causing the workers' physical symptoms. In summer, the average concentrations of total bacteria were greater than the Scandinavian guideline of 10,000 colony forming units per cubic meter of air (CFU/m3) in the receiving areas in plants 1 and 3, in the sorting areas of the three plants, and in shipping in plants 1 and 3. When the average concentrations of gram-negative bacteria were compared to the Scandinavian guideline of 1000 CFU/m3 of air, only the sorting department in plant 2 in summer exceeded this value. Average indoor concentrations of molds that were statistically significantly greater than those measured in the upwind outdoor air were measured in all departments in plants 1 and 3, regardless of the season, and only in sorting in plant 2 during the summer. The only chemical contaminant measured at average concentrations greater than 50% of the Threshold Limit Values (TLVs) proposed by the American Conference of Governmental Industrial Hygienists (ACGIH) was CO in the sorting departments in plants 1 and 2 during the winter. Noise exceeded the ACGIH TLV in plant 2. The workers' physical symptoms seem to be caused by the posture and effort required while remaining in a stationary position. Action must be focused on improving aspects such as work organization and personal protection and on informing citizens of the need for cleanliness of the material to be recycled. PMID- 11266100 TI - Air emission flux from contaminated dredged materials stored in a pilot-scale confined disposal facility. AB - A pilot-scale field simulation was conducted to estimate the air emissions from contaminated dredged material stored in a confined disposal facility (CDF). Contaminated dredged material with a variety of organic chemicals, obtained from Indiana Harbor Canal, was used in the study. It was placed in an outdoor CDF simulator (i.e., a lysimeter of dimensions 4 ft x 4 ft x 2 ft). A portable, dynamic flux chamber was used to periodically measure emissions of various polynuclear aromatic hydrocarbons (PAHs). A weather station was set up to monitor and record the meteorological conditions during the experiment. The fluxes of several PAHs were monitored over time for 6 1/2 months. Initial 6-hr average fluxes varied from 2 to 20 ng/cm2/hr for six different PAHs. The flux values declined rapidly for all compounds soon after placement of the dredged material in the CDE Chemical concentrations derived from flux values were generally of low magnitude compared with ambient standards. Data obtained from the experiment were compared against those predicted using models for air emissions. Model simulations showed that initially the flux was largely from exposed pore water from saturated (wet) sediment, whereas the long-term flux was controlled by diffusion through the pore air of the unsaturated sediment. Model predictions generally overestimated the measured emissions. A rainfall event was simulated, and the dredged material was reworked to simulate that typical of a CDF operation. Increased flux was observed upon reworking the dredged material. PMID- 11266101 TI - A comparison of measured and simulated ozone concentrations in the rural areas of the eastern United States during summer 1995. AB - The recent regulatory actions toward a longer-term (i.e., 8-hr) average ozone standard have brought forth the potential for many rural areas in the eastern United States to be in noncompliance. However, since a majority of these rural areas have generally few sources of anthropogenic emissions, the measured ozone levels primarily reflect the effects of the transport of ozone and its precursor pollutants and natural emissions. While photochemical grid models have been applied to urban areas to develop ozone mitigation measures, these efforts have been limited to high ozone episode events only and do not adequately cover rural regions. In this study, we applied a photochemical modeling system, RAMS/UAM-V, to the eastern United States from June 1-August 31, 1995. The purpose of the study is to examine the predictive ability of the modeling system at rural monitoring stations that are part of the Clean Air Status Trends Network (CASTNet) and the Gaseous Pollutant Monitoring Program (GPMP). The results show that the measured daily 1-hr ozone maxima and the seasonal average of the daily 1 hr ozone maxima are in better agreement with the predictions of the modeling system than those for the daily 8-hr ozone maxima. Also, the response of the modeling system in reproducing the measured range of ozone levels over the diurnal cycle is poor, suggesting the need for improvement in the treatment of the physical and chemical processes of the modeling system during the nighttime and morning hours if it is to be used to address the 8-hr ozone standard. PMID- 11266102 TI - Carbon disulfide and hydrogen sulfide removal with a peat biofilter. AB - Simultaneous removal of H2S and CS2 was studied with a peat biofilter inoculated with a Thiobacillus strain that oxidizes both compounds in an acidic environment. Both sulfurous gases at concentrations below 600 mg S/m3 were efficiently removed, and the removal efficiencies were similar, 99%, with an empty bed retention time (EBRT) of more than 60 sec. Concentrations greater than 1300-5000 mg S/m3 caused overloading of the filter material, resulting in high H2SO4 production, accumulation of elemental sulfur, and reduced removal efficiency. The highest sulfur removal rate achieved was 4500 g-S/day/m3 filter material. These results indicate that peat is suitable as a biofilter material for the removal of a mixture of H2S and CS2 when concentrations of gases to be purified are low (less than 600 mg/m3), but it is still odorous and toxic to the environment and humans. PMID- 11266103 TI - Chemical composition and source apportionment of PM2.5 particles in the Sihwa area, Korea. AB - To investigate the chemical characteristics of fine particles in the Sihwa area, Korea, atmospheric aerosol samples were collected using a dichotomous PM10 sampler and two URG PM2.5 cyclone samplers during five intensive sampling periods between February 1998 and February 1999. The Inductively Coupled Plasma (ICP) Atomic Emission Spectrometry (AES)/ICP-Mass Spectrometry (MS), ion chromatograph (IC), and thermal manganese dioxide oxidation (TMO) methods were used to analyze the trace elements, ionic species, and carbonaceous species, respectively. Backward trajectory analysis, factor analysis, and a chemical mass balance (CMB) model were used to estimate quantitatively source contributions to PM2.5 particles collected in the Sihwa area. The results of PM2.5 source apportionment using the CMB7 receptor model showed that (NH4)2SO4 was, on average, the major contributor to PM2.5 particles, followed by nontraffic organic carbon (OC) emission, NH4NO3, agricultural waste burning, motor vehicle emission, road dust, waste incineration, marine aerosol, and others. Here, the nontraffic OC sources include primary anthropogenic OC emitted from the industrial complex zone, secondary OC, and organic species from distant sources. The source impact of waste incineration emission became significant when the dominant wind directions were from southwest and west sectors during the sampling periods. It was found that PM2.5 particles in the Sihwa area were influenced mainly by both anthropogenic local sources and long-range transport and transformation of air pollutants. PMID- 11266104 TI - Residential proximity to industrial sources of air pollution: interrelationships among race, poverty, and age. AB - This study builds on earlier work investigating statistical relationships between sociodemographic characteristics of populations and their residential proximity to industrial sources of air pollution. The analysis uses demographic data from the 1990 U.S. Census and industrial site data from the U.S. Environmental Protection Agency (EPA)'s 1990 Toxics Release Inventory (TRI). The focus is on examining interactions among race (African Americans and Whites), poverty (above and below household poverty threshold), and age (children from birth to 5 years of age and elderly people 65 years old or older). Results from three different study areas (Kanawha Valley in West Virginia, the Baton Rouge-New Orleans Corridor in Louisiana, and the greater Baltimore metropolitan area in Maryland) suggest there are important interactions among race, poverty, and age that are likely to have consequential ramifications for efforts aimed at investigating issues related to environmental justice. Our results indicate that a substantial proportion of all demographic groups studied live within a mile of the nearest facility, with values ranging from 22% of Whites above poverty in the Baton Rouge New Orleans Corridor to 60% of African Americans below poverty in Baltimore. Likewise, a substantial proportion of all demographic groups also live within 2 miles of four or more industrial facilities, with values ranging from 16% for Whites above poverty in the Corridor to 70% for African Americans below poverty in Baltimore. In all three study areas, African Americans were more likely than Whites to (1) live in households with incomes below the household poverty line, (2) have children 5 years of age or younger, (3) live closer to the nearest industrial emissions source, and (4) live within 2 miles of multiple industrial emission sources. Findings indicate that, compared with White children, a substantially higher proportion of African-American children 5 years of age or younger lived in poor households that were located in relatively close proximity to one or more industrial sources of air pollution. PMID- 11266105 TI - Urea as a PCDD/F inhibitor in municipal waste incineration. AB - Emissions of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) from municipal waste incineration have been widely studied because of their extensive toxicity, and many efforts have been made to restrict their emissions. Although a number of chemical compounds have been shown in laboratory-scale tests to inhibit the formation of PCDD/Fs, few have been tested in pilot- or full-scale plants. This work evaluates the effect of urea as a PCDD/F inhibitor in a pilot-scale incinerator that uses refuse-derived fuel (RDF). The decomposition of urea under the test conditions was also studied using detailed kinetic modeling. An aqueous solution of urea was injected into the flue gas stream after the furnace at approximately 730 degrees C, with varied urea concentrations and flue gas residence times used between the furnace and the sampling point. The results demonstrate that urea can successfully inhibit PCDD/F formation in waste incineration if concentrations and injection points are properly adjusted. The kinetic model showed that urea can be rapidly decomposed under appropriate flue gas conditions, indicating that in addition to the urea molecule itself, decomposition products of urea can also be responsible for the reduction of PCDD/F production during incineration. PMID- 11266106 TI - A chassis test procedure to mimic the heavy-duty engine transient emissions certification test. AB - In-use emissions from vehicles using heavy-duty diesel engines can be significantly higher than the levels obtained during engine certification. These higher levels may be caused by a combination of degradation of engine components, poor engine maintenance, degradation or failure of emissions after-treatment devices, and engine and emissions system tampering. A direct comparison of in-use vehicle emissions with engine certification levels, however, is not possible without removing an engine from the vehicle in order to perform engine dynamometer emissions testing. The goal of this research was to develop a chassis test procedure that mimics the engine performance, and as such the expected emissions levels, from the engine certification emissions test prescribed in the U.S. Code of Federal Regulations. Emissions measurements were taken from two engines during testing on an engine dynamometer using the transient heavy-duty Federal Test Procedure (FTP). Additionally, each engine was installed in an appropriate vehicle, and emissions measurements were taken using a chassis dynamometer while employing a vehicle driving schedule intended to match closely the instantaneous torque and speed schedule of the engine FTP. Engine and chassis testing was performed with the engines in stock (unmodified) condition as well as in several modes to simulate either tampered or poorly maintained conditions. The use of a chassis test as a predictive tool for determining whether an engine in a vehicle would pass the engine certification test has proven to be worthwhile. Analysis of the data shows that identification of chassis-mounted engines with NOx emissions above certification levels is possible by employing engine-specific correction factors. In the case of PM emissions, significant data scatter allowed only the identification of gross PM emitters. Engine tampering and poor maintenance can raise PM and NOx emissions, and these increases can be correctly identified by a chassis test. Analysis of chassis and engine CO and HC emissions did not reveal a strong enough correlation to warrant the use of the chassis test for emissions screening of these two pollutants. PMID- 11266107 TI - Smoke impacts from agricultural burning in a rural Brazilian town. AB - Agricultural and silvicultural biomass burning is practiced in many undeveloped portions of the Amazon basin. In Rond nia, Brazil, such burning is restricted to a brief period in the dry season of August and September to minimize the duration of air quality impacts and to attempt to control escaped fires. During this period, much of the region and the communities within it experience significant exposure to smoke from agricultural and forest fires. In cooperation with Brazilian scientists of the University of Brasilia, the Brazilian Organization for Agricultural Research (EMBRAPA), and the Alternative to Slash and Burn Program coordinated by the International Center for Research in Agroforestry (ICRAF), ambient air quality was measured in Theobroma, a small town in Rond nia, during one week of the open burning period of 1995 to supplement available air quality data and to foster public awareness of the impacts of widespread fires. Personal sampling equipment was used to measure ambient levels of formaldehyde (HCHO), acrolein, CO, benzene, and respirable PM in outdoor air. The data obtained were compared with established Brazilian and U.S. ambient air quality guidelines. Ambient levels of respirable PM averaged 191 microg/m3, HCHO averaged 12.8 ppb, CO averaged 4.2 ppm, and benzene averaged 3.2 ppb. Almost all acrolein samples were less than the detection limit of 1 ppb. The results showed that the public can be exposed to relatively high levels of pollutants under the prescribed burning smoke management strategy of a two- to three-week prescription burning period, although this is an improvement over past years when burning was unregulated and continued through most of the dry season. The results also demonstrate the feasibility of using personal exposure monitoring equipment for low-cost surveys of ambient air quality in polluted regions. PMID- 11266108 TI - Source apportionment of PM10 and PM2.5 in five Chilean cities using factor analysis. AB - Chile is a fast-growing country with important industrial activities near urban areas. In this study, the mass and elemental concentrations of PM10 and PM2.5 were measured in five major Chilean urban areas. Samples of particles with diameter less than 10 microm (PM10) and 2.5 microm (PM2.5) were collected in 1998 in Iquique (northern Chile), Valparaiso, Vina del Mar, Rancagua (central Chile), and Temuco (southern Chile). Both PM10 and PM2.5 annual mean concentrations (PM10: 56.9-77.6 microg/m3; PM2.5: 22.4-42.6 microg/m3) were significantly higher than the corresponding European Union (EU) and U.S. Environmental Protection Agency (EPA) air quality standards. Moreover, the 24-hr PM10 and PM2.5 U.S. standards were exceeded infrequently for some of the cities (Rancagua and Valparaiso). Elements ranging from Mg to Pb were detected in the aerosol samples using X-ray fluorescence (XRF). For each of the five cities, factor analysis (FA) was applied to identify and quantify the sources of PM10 and PM2.5. The agreement between calculated and measured mass and elemental concentrations was excellent in most of the cities. Both natural and anthropogenic sources were resolved for all five cities. Soil and sea were the most important contributors to coarse particles (PM10-PM2.5), whereas their contributions to PM2.5 were negligible. Emissions from Cu smelters and oil refineries (and/or diesel combustion) were identified as important sources of PM2.5, particularly in the industrial cities of Rancagua, Valparaiso, and Vina del Mar. Finally, motor vehicles and wood burning were significant sources of both PM2.5 and PM10 in most of the cities (wood burning was not identified in Iquique). PMID- 11266109 TI - Molecular cloning and characterization of a tobacco leaf cDNA encoding a phosphate transporter. AB - Phosphate acquired by roots is translocated to and utilized by the upper part of the plant, where the phosphate transport in the cell is also important in the phosphate metabolism. In order to study the role of the phosphate transporter in the regulation of the phosphate movement across the membranes in leaf cells, we isolated and characterized a 2,059 bp tobacco leaf cDNA clone, NtPT1. The 537 amino acid sequences, deduced from NtPT1, exhibited 93 and 91% identites to one of the high affinity phosphate transporters constitutively expressed in potato and tomato roots, respectively. The NtPT1 contains 12 membrane-spanning domain with a central hydrophilic region. The expression of NtPT1 in the yeast high affinity phosphate transporter mutant strain, NS219, complemented the mutant and promoted cell growth significantly. These results strongly suggest that NtPT1 encodes a functional phosphate transporter and that one of the high affinity phosphate transporters expressed in roots is also expressed in leaves. Southern analysis indicated that tobacco phosphate transporter genes are low copy number genes and members of a small multi-gene family. PMID- 11266110 TI - Cloning of cDNA for a novel fibrinogen/angiopoietin-related protein, FARP. AB - Using a low abundant gene screening strategy in the human dermal papilla cell cDNA library, we isolated a novel cDNA, which was 1,872 bp of nucleotides in length and contained an open reading frame encoding 405 amino acids. We designated it 'fibrinogen/angiopoietin-related protein' (FARP) as it contained the characteristic coiled-coil domain and fibrinogen-like domain in the NH2- and COOH-terminal, which are conserved in angiopoietins. FARP has a highly hydrophobic region at the N-terminus that is typical of a secretory signal sequence. Recently, a very similar gene, HFARP, was cloned and they have a difference of only 18 amino acids in N-terminus. While HFARP was expressed only in the liver, northern blot analysis showed that FARP mRNA is abundantly expressed in the liver, placenta, prostate, and ovary in human adult tissues. It was also expressed in the fetal liver and lung carcinoma cell line. Further study will be needed to clarify the function of the FARP gene. PMID- 11266111 TI - Molecular cloning of the nahG gene encoding salicylate hydroxylase from Pseudomonas fluorescens. AB - A gene encoding the salicylate hydroxylase was cloned from the genomic DNA of Pseudomonas fluorescens SME11. The DNA fragment containing the nahG gene for the salicylate hydroxylase was mapped with restriction endonucleases and sequenced. The DNA fragment contained an ORF of 1,305 bp encoding a polypeptide of 434 amino acid residues. The nucleotide and amino acid sequences of the salicylate hydroxylase revealed several conserved regions with those of the enzyme encoded in P. putida PpG7: The homology of the nucleotide sequence is 83% and that of amino acid sequence is 72%. We found large conserved regions of the amino acid sequence at FAD and NADH binding regions. The FAD binding site is located at the amino terminal region and a lysine residue functions as a NADH-binding site. PMID- 11266112 TI - Molecular characterization of the NeIF2Bbeta gene encoding a putative eIF2B beta subunit in Nicotiana tabacum. AB - The NeIF2Bbeta cDNA encoding beta-subunit of the translation initiation factor 2B (eIF2B-beta) was identified from Nicotiana tabacum through protein interaction with PRK1, a reproductive-organ-specific receptor-like kinase (Park et al., 2000). The eIF2B is a guanine nucleotide-exchange protein that consists of five subunits, which function in the regulation of translation in eukaryotic cells. The NeIF2Bbeta that shows a high homology in the amino acid sequence with other beta-subunits also exhibits sequence similarity to a and delta subunits of eIF2B from yeast and animals. The NeIF2Bbeta gene was expressed in all of the tissues examined, but the mRNA level was higher in reproductive tissues than in vegetative tissues. During anther development, the NeIF2Bbeta mRNA was detected in all stages with a slightly higher level in the earliest stage. The NeIF2Bbeta GFP fusion protein was mainly localized in the cytosol. PMID- 11266113 TI - Molecular characterization of the cDNA encoding an acidic isoform of PR-1 protein in rice. AB - Rice cDNA encoding an acidic type of pathogenesis-related protein-1 (PR-1a) was cloned and characterized. The deduced PR-1a protein consisted of 168 amino acid residues, including 24 hydrophobic signal sequences at the N-terminus. The predicted molecular mass of the PR-1a was 15,728 Da with a theoretical pI of 4.5, an indication of an acidic protein. The PR-la showed high homology to an acidic PR-1 of Zea mays (74%) and a previously identified basic type PR-1 of rice (64%). Both rice PR-1 and PR-1a genes were found to exist as small gene families through Southern blot hybridization analyses. The PR-1 mRNA was accumulated only in leaves, while the PR-1a transcript was accumulated throughout the plant at a low level. Expression of both PR-1 genes was induced by infections of the rice blast fungus, Magnaporthe grisea, or the bacterial leaf blight pathogen, Xanthomonas oryzae pv. oryzae, and the treatment of benzo (1, 2, 3) thiadiazole-7-carbothioic acid S-ethyl ester, H2O2, or CuSO4. The expression of both PR-1 genes was higher and more rapidly induced in an incompatible interaction than in a compatible interaction in the rice M. grisea interactions. PMID- 11266114 TI - A hot pepper cDNA encoding a pathogenesis-related protein 4 is induced during the resistance response to tobacco mosaic virus. AB - Hot pepper (Capsicum annuum) plants exhibit a hypersensitive response (HR) against infection by many tobamoviruses. A clone (CaPR-4) encoding a putative pathogenesis-related protein 4 was isolated by differential screening of a cDNA library prepared from resistant pepper plant leaves inoculated with tobacco mosaic virus (TMV) pathotype P0. The predicted amino acid sequence of CaPR-4 is very similar to those of other plant PR-4s. Southern blot analysis showed that small gene families of PR-4-related sequences were present in the pepper genome. Hot pepper cultivar Bugang, resistant to TMV-P0 and susceptible to TMV-P1.2, induced CaPR-4 expression by pathotype P0 inoculation in inoculated and systemic leaves, but not by pathotype P1.2. Effects of exogenously applied abiotic elicitors upon the CaPR-4 expression were also examined. The expression of the CaPR-4 gene was stimulated by methyl jasmonate (MeJA), ethephon and wounding treatment. However, application of salicylic acid (SA) did not trigger the expression. Evidence is emerging that jasmonic acid and ethylene play key roles in the SA-independent pathways of plant-pathogen interaction. Taken together, these results suggest that the CaPR-4 gene is one of the defense-related genes conferring resistance on pepper plants by the SA-independent pathway and the cross-talk between signaling compounds, jasmonic acid and ethylene could have a great regulatory potential in a plant's defense against TMV. PMID- 11266115 TI - Fibrin stimulates microfilament reorganization and IL-1beta production in human monocytic THP-1 cells. AB - Fibrin plays important roles in the wound healing processes, including blood clotting and platelet aggregation. Additional activities of fibrin were found in this study, which utilizes human THP-1 cells treated 1,25-(OH)2 vitamin D3 and plasminolytic fragments derived from fibrin. Coated fibrin fragment E on culture plates induced cell adhesions and morphological changes of the THP-1 cells, being resembled to tissue macrophages. Morphological changes of the THP-1 cells were caused by microfilament reorganization. IL-1beta production was increased in the THP-1 cells by adherent fibrin fragment E, but not by fibrin fragment D or by fibrinogen fragment E. The elevation of IL-1beta production is caused by transcriptional activation. Incubation with cytochalacin D, an actin polymerization inhibitor, prevents both microfilament reorganization and morphological changes, but has no effect on the IL-1beta production stimulated by fibrin fragment E. This data suggests that the IL-1beta production in the THP-1 cells do not require microfilament reorganization and integrin aggregation. Taken together, these results indicate that fibrin matrix plays an additional role in the stimulation of monocytes for production of IL-1beta, morphological changes and cell adhesion, resulting in the facilitation of the wound healing processes. PMID- 11266117 TI - Interleukin-4 induces two distinct GAS-binding complexes containing STAT6: evidence for DNA binding of STAT6 monomer. AB - Treatment of tonsillar mononuclear cells with interleukin-4 (IL-4) rapidly induced the formation of two distinct complexes upon reaction of the cell extracts with the IL-4-responsive element (IL-4RE) of the CD23b promoter in the electrophoretic mobility sift assays (EMSA). The two complexes were detected with a similar activation kinetics upon IL-4 stimulation. They were both immunoreactive with antibodies to signal transducer and activator of transcription (STAT) 6. The upper complex, however, appeared more stable in the competitor oligomer binding assays and more resistant in the anti-phosphotyrosine antibody inhibition assays than the lower complex. Western blot analysis revealed a single peptide of 105 kDa reacting with anti-STAT6 antibodies, which ruled out the possibility for multiple isoforms of STAT6 in these cells. Subsequently, Southwestern analysis demonstrated that monomeric STAT6 in the IL-4-treated nuclear extract can bind the labeled IL-4RE in an activation-dependent manner. Our data strongly suggests that in addition to dimeric STAT6, monomeric STAT6, albeit with a lower affinity, can bind the IL-4RE/gamma activation site (GAS) upon IL-4-induced activation. Also, the upper and lower bands observed in EMSA are likely to represent dimeric and monomeric STAT6 bound to the IL-4RE oligomer, respectively. The functional implication of the STAT6 monomer binding to GAS in the IL-4-induced gene activation is discussed. PMID- 11266116 TI - Correlation between genotype and phenotype in Korean patients with spinal muscular atrophy. AB - The goal of this study was to define the correlation between genotype and phenotype in Korean patients with spinal muscular atrophy (SMA). The SMA can be classified into three groups based on the age of onset and the clinical course. The candidate genes, survival motor neuron (SMN) gene, neuronal apoptosis inhibitory protein (NAIP) gene, and p44 gene were mapped and duplicated with telomeric and centromeric. The loss of the telomeric SMN occurs by a different mechanism. That is the deletion or conversion of telomeric SMN to centromeric SMN, in which case the conversion could produce a mild phenotype and deletion could produce a severe one. It has been known that there may be a balance between the numbers of copies expressed by the centromeric and telomeric SMN genes. In our study, ten patients with type I SMA and two type II patients were identified by their clinical findings and DNA studies. The major deletion of SMA candidate genes, deletion of the SMN gene, NAIP gene, and p44 gene were identified in six patients with type I SMA, while the rest of type I and all the type II patients showed the deletion of the SMN gene only. Allele numbers of the C212 marker were compared in patients and normal controls in order to find the correlation between the copy numbers and the clinical severity. The result was that type I patients had 2-5 alleles and the normal controls had 4-6. This suggests that the deletion is a major determining factor in the clinical phenotype. However, two type I patients with telomeric NAIP gene deletion notably had 4-5 alleles, as in the normal controls. This result implies that the correlation between the copy numbers and the severity is uncertain as opposed to the previous hypothesis. One type I patient showed the conversion of the centromeric SMN gene to the telomeric, which supports the conclusion that gene conversion is an important molecular mechanism for SMA. In the study of one hundred normal newborns, two physically normal newborns showed deletion of the centromeric SMN gene, suggesting frequent rearrangement in the locus. PMID- 11266118 TI - Cloning of androgen-inducible gene 1 (AIG1) from human dermal papilla cells. AB - Cultured human dermal papilla cells are useful for studying the androgen dependent growth of hair follicles. We cloned the human homolog of FAR-17a, a gene identified from the hamster flank organ as one of the androgen inducible genes, by degenerative PCR and human dermal papilla cDNA library screening. We isolated a novel cDNA clone, designated as AIG1 (Androgen-inducible Gene 1), whose expression was found to be inducible by androgen. AIG1 cDNA consists of 1,398 nucleotides in length, which encodes a protein of 238 amino acids (27 kDa). The deduced protein sequence showed 35% overall homology with FAR-17a. RT-PCR of human dermal papilla cDNA revealed two mRNA transcripts, which differed by 156 nucleotides. This results in an in-frame deletion of 52 amino acids. A computer analysis of hydropathy indicated five hydrophobic domains are present in the large protein sequence, while four hydrophobic portions are in the smaller protein sequence. In a Northern blot analysis, the major 1.5 kb and minor 1.2 kb bands of AIG1 mRNA were detected. AIG1 mRNA was expressed at a relatively high level in the heart, ovary, testis, liver, and kidney. However, they were expressed at a low level in the spleen, prostate, brain, skeletal muscle, pancreas, small intestine, and colon. When dermal sheath cells were stimulated with DHT, the level of AIG1 mRNA expression was increased at 30 ng/ml. The level of expression was higher in males than females. In this study, we cloned and initially characterized AIG1. Further study will be needed to understand the functions of AIG1 in the androgen-regulated hair cycle. PMID- 11266119 TI - Excision repair of adozelesin-N3 adenine adduct by 3-methyladenine-DNA glycosylases and UvrABC nuclease. AB - Adozelesin is a synthetic analog of the antitumor antibiotic CC-1065, which alkylates the N3 of adenine in the minor groove in a sequence-selective manner. Since the cytotoxic potency of a DNA alkylating agent can be modulated by DNA excision repair system, we investigated whether nucleotide excision repair (NER) and base excision repair (BER) enzymes are able to excise the bulky DNA adduct induced by adozelesin. The UvrABC nuclease and 3-methyladenine-DNA glycosylase, that exhibit a broad spectrum of substrate specificity, were selected as typical NER and BER enzymes, respectively. The adozelesin-DNA adduct was first formed in the radiolabeled restriction DNA fragment and its excision by purified repair enzymes was monitored on a DNA sequencing gel. The treatment of the DNA adduct with a purified UvrABC nuclease and sequencing gel analysis of cleaved DNA showed that UvrABC nuclease was able to incise the adozelesin adduct. The incision site corresponded to the general nuclease incision site. Excision of this adduct by 3 methyladenine-DNA glycosylases was determined following the treatment of the DNA adduct with a homogeneous recombinant bacterial, rat and human 3-methyladenine DNA glycosylases. Abasic sites generated by DNA glycosyalses were cleaved by the associated lyase activity of the E. coli formamidopyrimidine-DNA glycosylase (Fpg). Resolution of cleaved DNA on a sequencing gel showed that the DNA glycosylase from different sources could not release the N3-adenine adducts. A cytotoxicity assay using E. coli repair mutant strains showed that E. coli mutant strains defective in the uvrA gene were more sensitive to cell killing by adozelesin than E. coli mutant strain defective in the alkA gene or the wild type. These results suggest that the NER pathway seems to be the major excision repair system in protecting cells from the cytotoxicity of adozelesin. PMID- 11266120 TI - Molecular cloning and cultivar specific expression of MAP kinases from Capsicum annuum. AB - Two MAP kinases, MK1 and MK2, were cloned from Capsicum annuum (pepper) cv. Subicho using a parsley MAP kinase gene as a heterologous probe. MK1 and MK2 encode stress-inducible protein kinases that can contribute to the response to wounding, UV-C, and cold. MK1 has a 92% amino acid identity with WIPK of tobacco. It was transcriptionally induced in response to wounding. In contrast, no detectable MK1 transcript was found in unwounded leaves of pepper. MK2 has a high level of amino acid homology to MAP kinases, such as NTF4 and SIPK and was constitutively expressed in all tissues. Both MK transcripts were downregulated by UV-C treatment. Each MK protein activation was independently wound-inducible in a cultivar dependent manner. MK1 is phosphorylated in cv. Pungchon but not cv. Subicho; whereas, the MK2 protein activation by wounding is restricted to cv. Subicho. In addition, de novo synthesis of the MK1 protein and tyrosine phosphorylation was rapidly and transiently induced in cv. Pungchon by wounding. In contrast, it is highly unlikely that the MK1 protein is produced in cv. Subicho, even though there is an abundant expression of MK1 mRNA after wounding in this cultivar. In Escherichia coli, which overexpresses MK1, autophosphorylation is observed at conserved threonine and tyrosine phosphorylation sites. PMID- 11266121 TI - Primary structures and chain dominance of anti-DNA antibodies. AB - Using several anti-DNA autoantibodies, we analyzed the relative involvement of heavy and light chains in their interactions with DNA. We previously obtained eight hybridomas producing monoclonal anti-DNA autoantibodies by fusing spleen cells from an MRL-lpr/lpr mouse with myeloma cells. The chain dominance was analyzed by UV cross-linking experiments, in which the antibodies were covalently cross-linked with radioisotope-labeled oligonucleotides by short-wavelength UV light, and the cross-linked H and L chains were analyzed by SDS-PAGE and densitometric scanning. Among these, three were found to be heavy chain dominant antibodies in which heavy chains are dominantly involved in DNA binding. The other five were co-dominant antibodies in which both heavy and light chains are involved in DNA binding. To determine the factor(s) that can explain the chain dominance in DNA binding, we determined the amino acid sequences of the variable regions of both heavy (VH) and light (VL) chains of all eight monoclonal antibodies. By analyzing the data, we were able to draw the following conclusions: (1) The arginine residues are found in the CDR3 regions of both VH and VL of the co-dominant antibodies; whereas, the same residues are found only in the CDR3s of VH, but not in VL, of the heavy chain dominant antibodies. (2) The net charges of the V regions affect the chain dominance. From the results of this study it is suggested that the presence of arginine residue in CDR3 is a critical factor in determining chain-dominance, as well as DNA binding of anti DNA antibodies in general. PMID- 11266122 TI - Quantitative trait loci mapping associated with plant regeneration ability from seed derived calli in rice (Oryza sativa L.). AB - Quantitative trait loci (QTLs), which are associated with the ability of plant regeneration from seed derived calli, were detected using a recombinant inbred (RI) population from a cross between 'Milyang 23 (toingil)' and 'Gihobyeo (japonica)' in rice (Oryza sativa L.). A tongil type cultivar, 'Milyang 23', has a lower frequency of callus induction and plant regeneration than those of japonica 'Gihobyeo'. Transgressive segregations were observed for the callus induction rate and plant regeneration ability from seed derived calli of the RI population. An interval mapping analysis was used to identify the QTL controlling the plant regeneration ability. Two QTLs for the callus induction rate were detected on chromosomes 1 and 2, explaining the 10.9% total phenotypic variation. Four QTLs that are associated with the plant regeneration ability were located on chromosomes 2, 3, and 11, accounting for 25.7% of the total phenotypic variation. PMID- 11266123 TI - Molecular organization of the ribosomal RNA transcription unit and the phylogenetic study of Zymomonas mobilis ZM4. AB - Previously we reported that Zymomonas mobilis ZM4 contains three ribosomal transcription units (rrnA to C operons) which are clustered around the 50 min region, PacI fragments 13 and 6, on the physical map of Z. mobilis ZM4 [Kang, H. L. and Kang, H. S. (1998) Gene 206, 223-228]. The physical map reveals that the rrnA gene set is located on the 76 kb PacI fragment 13. The complete nucleotide sequence of the rrnA gene set has been determined. The total number of nucleotides of the rrnA gene set is about 6,250 bp. The structural genes of the 16S, 23S and 5S rRNA code for the 1,478 nt, 2,786 nt and 121 nt RNA chains, respectively. The length of the spacer regions between the 16S and 23S rRNA genes, and between the 23S and 5S rRNA genes, are 606 bp and 101 bp, respectively. Two tRNA genes, Ile-tRNA and Ala-tRNA, are found between the 16S and 23S rRNA genes and a fMet-tRNA gene is identified downstream of the 5S rRNA gene. Thus, the molecular organization of this rrnA gene set is the order of 5' 16S rRNA-tRNAIle-tRNAAla-23S rRNA-5S rRNA-tRNAfMet-3'. The secondary structure models of 16S, 23S and 5S rRNA are proposed. The phylogenetic tree, based on the 16S rRNA sequence, was constructed by neighbor-joining and maximum-parsimony methods. Zymomonas belongs to a group in which Aqorbacterium, Rhodobacter and Sphingomonas are included. PMID- 11266124 TI - Generation of a murine single chain Fv (scFv) antibody specific for cucumber mosaic virus (CMV) using a phage display library. AB - With the long-term goal of generating CMV-resistant transgenic plants using antibody genes, a single-chain variable fragment (scFv) antibody that binds to the cucumber mosaic virus was isolated from a scFv phage display library by four rounds of affinity selection with CMV-Mf as an antigen. The scFv has the identical binding specificity to CMV as a monoclonal antibody that is generated by the hybridoma fusion technique, and recognized purified preparations of CMV isolates belonging to either subgroup I or II in immunoblotting. The nucleotide sequences of the recombinant antibody showed that a heavy chain variable region (V(H)) gene belonged to the VH3 subgroup and the kappa light chain variable region (V kappa) came from the Vkappa4 subgroup. Our results demonstrate that the scFv phage display library, an alternative approach to the traditional hybridoma fusion technique, has a potential applicability in the study of plant virus and plant pathology. PMID- 11266125 TI - Identification of a nuclear protein ArpN as a component of human SWI/SNF complex and its selective association with a subset of active genes. AB - The hSWI/SNF complex remodels the chromatin structure to modulate gene expression. The hSWI/SNF complex is a multiprotein complex with at least 10 different proteins in mammals. In this study, we identified the 45 kDa subunit of the hSWI/SNF complex as ArpN, an actin-related protein. ArpN has a 36% identity and 50% similarity with the human beta-actin, but cannot be classified into any known class of actin-related proteins. ArpN is exclusively localized within the nucleus and appears as the unbound, chromatin-associated, or nuclear matrix associated forms in the nucleus. In the chromatin immunoprecipitation (ChIP) assay, we found the associations of ArpN with the Ets-2 and c-mycP2 promoter regions in HeLa cells. The promoter regions of the hsp70, cyclophilin, beta globin, TdT, and cd4 genes, however, were not associated with ArpN. The Ets-2 and c-mycP2 genes are expressed actively in HeLa cells, but beta-globin, TdT, and cd4 genes are inactive. The hsp70 and cyclophilin genes have a feature of stress inducibility. These selective associations of ArpN with a subset of active genes support the proposition that the requirement of hSWI/SNF complex in gene activation is gene specific. PMID- 11266126 TI - Stimulation of human DNA topoisomerase II activity by its direct association with the beta subunit of protein kinase CKII. AB - DNA topoisomerase II copurifies with and is phosphorylated by protein kinase CKII. In this study, a yeast two-hybrid system was used to investigate the interaction between human topoisomerase II isozymes and CKII subunits. The two hybrid test clearly showed that both topoisomerase IIalpha and IIbeta interact with the CKIIbeta, but not the CKIIalpha subunit. The two-hybrid test also demonstrated that topoisomerase IIbeta residues 1099-1263 and topoisomerase IIalpha residues 1078-1182 mediate the interaction with the CKIIbeta subunit, providing evidence that the leucine zipper motif and the major CKII-dependent phosphorylation sites of topoisomerase II are unnecessary for its physical binding to CKIIbeta. Furthermore, a DNA relaxation assay demonstrated that the CKII subunit enhances topoisomerase II activity by physical interaction with topoisomerase II. PMID- 11266127 TI - Molecular cloning and characterization of a novel mouse betaPix isoform. AB - BetaPix, a Pak-interacting guanine nucleotide exchange factor is known to be involved in the regulation of Cdc42/Rac GTPases and Pak kinase activity. Currently, three 1Pix isoforms, betaPix-a, -b, and -c have been reported. In this study, the cDNA of a novel Pix splice variant was isolated from a mouse brain cDNA library. The cloned betaPix isoform, named betaPix-d, lacks leucine zipper domain that is present in other Pix isoforms, and has a 11 amino acid addition at carboxyl terminus and distinct 3'-UTR Analysis of the tissue distribution of betaPix-d using RT-PCR revealed that its message was present mainly in brain and testis but in lower levels in heart, spleen, lung, liver, skeletal muscle and kidney. In situ hybridization studies with the 13Pix-d specific probes in the rat embryo show that betaPix-d isoform is expressed mainly in the central nervous system. Moreover, temporal expression pattern of the isoform is correlated with the active neurogenesis period in the cerebral cortex and cerebellum during rat brain development. These findings suggest that betaPix-d isoform may be developmentally regulated. PMID- 11266128 TI - Structure and expression of a CTP: phosphocholine cytidylyltransferase gene from Arabidopsis thaliana. AB - A genomic clone, which includes the CTP: phosphocholine cytidylyltransferase (CCT) open reading frame and its 5'- and 3'- flanking non-coding regions, has been isolated from Arabidopsis thaliana and sequenced. The CCT gene is approximately 3.0 kb in length and contains 8 exons interrupted by 7 introns, which range from 74 to 626 nucleotides. All nucleotide sequences for the intron 3' splice sites are consistent with the consensus AG sequence of plant pre-mRNA processing, while the major GT consensus sequence for the 5' splice site is conserved in 5 of 7 introns. Introns 5 and 6 have a minor GC consensus sequence instead. In the 5'-flanking region there are two sequences related to a cold responsive element found in the cold-inducible promoter of the A. thaliana cor15a gene, plus one gibberellin-response element. The results from reverse transcriptase-PCR indicate that the expression of A. thaliana CCT is regulated by temperature. The expression level of CCT increased after a 30-min treatment at 5 degrees C. When the plants were returned to 22 degrees C, the expression of CCT also decreased to the original level. PMID- 11266129 TI - A description of two new species of coccidia (Apicomplexa: Eimeriidae) from African reptiles with nomenclatural corrections for two Caryospora and one Eimeria species from snakes. AB - Two new species of coccidian parasites are described from African reptiles. Oocysts of Eimeria foulshami sp. n. from the plated lizard Gerrhosaurus major bottegoi Del Prato of Sudan are ellipsoidal, 24.1 x 14.9 (23-26.5 x 14-17.8) microm with a bilayered, colourless oocyst wall and lack polar granules. The ellipsoidal sporocysts average 8.6 x 4.6 (7-10.6 x 4.4-7) microm and possess a prominent, globular, sporocyst residuum. Oocysts of Caryospora regentensis sp. n. from the Eastern green mamba Dendroaspis augusticeps Smith, 1849 [corrected] of Kenya are spherical to subspherical, 16.8 x 16.4 (16-17.6 x 15-17.2) microm with a bilayered oocyst wall and a single polar granule. The ellipsoidal sporocysts average 13.0 x 10.3 (10.2-14 x 9.2-11) microm and possess a Stieda and substieda body and a prominent globular sporocyst residuum. Oocysts of Caryospora legeri Hoare, 1933 are reported from a hissing sand snake, Psammophis sibilans sibilans L. from Nigeria, representing a new geographical record. The oocysts are slightly larger than the type, but otherwise identical. Caryospora psammophi Bray, 1960 and C. hermae Bray, 1960 from Psammophis sibilans phillipsi, oocysts of which are morphologically similar to and overlap in dimensions with C. legeri Hoare, 1933, are synonymised with the latter species. Eimeria samiae Iskander et Tadros, 1979 is emended to E. samyadeli to reflect the gender of the person the species was named after and because E. sani is preoccupied. In addition to these findings, Eimeria bohemi Modry, Slapeta et Koudela, 2000 and oocysts of an unidentified spherical Eimieria sp. are reported from Chamaeleo dilepis dilepis Leach from Cameroon. PMID- 11266130 TI - Experimental transmission of Caryospora kutzeri (Apicomplexa: Eimeriidae) by rodent hosts. AB - Four laboratory-hatched European kestrels Falco tinnunculus L. were fed on laboratory mice and common voles Microtus arvalis Pallas previously inoculated with different doses of sporulated oocysts of Caryospora kutzeri Boer, 1982. Two kestrels that were fed infected mice shed C. kurtzeri oocysts 6 days after ingesting murine tissues. To compare direct and indirect transmissions, two of the kestrels were subsequently directly inoculated with 10(5) sporulated C. kutzeri oocysts and became patent on days 8 and 9 and shed caryosporan oocysts up to day 25 post inoculation. Additionally, four mice were inoculated with 10(6) oocysts in order to examine mouse tissues for the presence of developmental stages of C. kutzeri. No coccidian stages were found in the tissues of inoculated mice. The experiment showed that developmental stages of C. kutzeri are able to survive in mouse tissues and cause infection of suitable host after their ingestion. PMID- 11266132 TI - Grillotia borealis sp. n. (Cestoda: Trypanorhyncha) from five species of Bathyraja (Rajiformes: Arhynchobatidae) in the North Pacific Ocean with comments on parasite enteric distribution. AB - A new trypanorhynch cestode, Grillotia borealis sp. n., is described from the spiral intestines of softnose skates of the genus Bathyraja collected from subarctic waters of the North Pacific Ocean: B. parmifera (Bean) (type host), B. aleutica (Gilbert) and B. interrupta (Gill et Townsend) from the Bering Sea and B. minispinosa Ishiyama et Ishihara and B. sminovi (Soldatov et Pavlenko) from the Sea of Okhotsk off Japan. The new species is distinguished from other species of Grillotia by possession of the following combination of characters: four hooks per principal row, hooks 4(4') distinctly separated from hooks 3(3') of principal row, principal rows separated by 13-15 intercalary hooks in 2-3 rows, hooks 2(2') and 3(3') change in form along their respective files, hooks 1(1') do not change in form along the file, a broad band of microhooks on the external tentacular face, intermediary hooks are lacking, absence of a special basal armature, origin of the retractor muscle near middle of the bulb, average scolex ratio of 1:3:2:0.1, and a hermaphroditic sac. Grillotia borealis consistently favoured the most anterior regions of the spiral intestine. Seventy-one per cent of 21 attached worms occupied the most anterior chamber of the spiral valve and 52 per cent were embedded in the anterior surface of the spiral valve whorls. Factors which may limit the distribution of G. borealis within the spiral intestine of its host are discussed. Statistically significant differences occur in the mucosal morphology of B. aleutica and B. parmifera for villus length, diameter, spatial arrangement and number per unit area along the antero-posterior axis of the spiral intestine. PMID- 11266131 TI - Dendromonocotyle colorni sp. n. (Monogenea: Monocotylidae) from the skin of Himantura uarnak (Dasyatididae) from Israel and a new host record for D. octodiscus from the Bahamas. AB - Dendromonocotyle colorni sp. n. (Monogenea: Monocotylidae) is described from the dorsal skin surface of two specimens of Himantura uarnak (Forsskal) kept at the Eilat Underwater Observatory in Israel. Dendromonocotyle colorni is distinguished from the other eight species in the genus by the morphology of the terminal papillar sclerite on the haptor, the distal portion of the male copulatory organ and the morphology of the vagina. The development of the male copulatory organ is detailed for D. colorni and the adaptations of species of Dendromonocotyle to life on the dorsal skin surface of rays are discussed. Dendromonocotyle octodiscus Hargis, 1955 was identified from the dorsal skin surface of the southern stingray Dasyatis americana Hildebrand et Schroeder off Bimini, Bahamas and represents a new host record. PMID- 11266133 TI - Gill parasites of Cephalopholis argus (Teleostei: Serranidae) from Moorea (French Polynesia): site selection and coexistence. AB - The distribution and coexistence of gill ectoparasites of 121 specimens of Cephalopholis argus Bloch et Schneider, caught between October 1994 and October 1995, were investigated. Adults of the monogenean Benedenia sp. and copepod Hatschekia sp., the larval caligid copepod Caligus sp. (copepodite and chalimus stages), and praniza larvae of the isopod Gnathia sp. were found. All species were aggregated within the host population. Infracommunities were poor, with only 40.5% of fish infected by two parasite species. Only two individual fish harboured all the parasite species observed at the component community level. Prevalences were less than 50% and mean intensities were low (less than 6 parasites/host). No dominant parasite species were observed in the host population. The spatial distribution of each parasite species was studied on different partitions of the gill arches. Adult parasite stages that are mobile showed much overlap in their distribution, whereas temporarily attached larvae of Caligidae were more site specific. Copepodite and chalimus larvae showed niche restriction that is probably due to gregarious behaviour. Positive associations between caligid larvae reflected intraspecific interaction for site and/or resources. Each of the Caligus sp. larval stages prefers specific sites, as do the adults, which occur exclusively in the buccal cavity of the host. Infracommunities were too poor and too few to induce processes of interspecific competition. PMID- 11266134 TI - Colonisation and extinction in relation to competition and resource partitioning in acanthocephalans of freshwater fishes of the British Isles. AB - This paper challenges two paradigms long held in relation to the ecology of parasites in freshwater systems: (1) autogenic species are poorer colonisers than allogenic ones; and (2) parasites with direct life cycles are more successful colonisers than those with complex life cycles. Using new and existing data for Acanthocephala in freshwater fish from the British Isles, it is suggested that all six species present have been able to colonise and persist successfully, in spite of the supposed limitations of their autogenic life-style. It is proposed that these parasites have overcome these limitations by a variety of means, which apply equally to all species considered. Foremost among these is the utilisation of a migratory fish host as either a preferred or a suitable host in their life cycle, allowing colonisation of new areas and rescue effects in established areas, whilst equally important is the use of a common and widespread crustacean as the intermediate host. In addition, all six species appear to exhibit resource partitioning by host at either or both the larval and adult stages, thus reducing the potential for competition and further facilitating colonisation and survival. This hypothesis is supported by data from previous studies both on acanthocephalans from Europe and North America and on other autogenic parasites. It also provides an explanation for the apparently atypical host utilisation patterns of some acanthocephalan species in areas on the edge of their distributions, notably in Ireland. PMID- 11266135 TI - Some helminth parasites from Morelet's crocodile, Crocodylus moreletii, from Yucatan, Mexico. AB - An examination of three specimens of the Morelet's crocodile, Crocodylus moreletii Dumeril et Bibron, from the Lagoon of Celestun, Yucatan, Mexico revealed the presence of the following eight helminth species: Acanthostomum americanum (Perez Vigueras, 1956), Pelaezia loossi (Perez Vigueras, 1956), Telorchis sp. juv., Pseudoneodiplostomum groschafti sp. n. (all trematodes), Dujardinascaris helicina (Molin, 1860), Contracaecum sp. Type 2 larvae, Micropleura sp. and Paratrichosoma recurvum (Solger, 1877) (all nematodes). Pseudoneodiplostomum groschafti sp. n. is established by indication based on the description of specimens from Crocodylus rhombifer from Cuba, given by Groschaft and Barus (1970). Acanthostomum acuti Caballero et Brennes, 1959 is considered a synonym of A. americanum. A. americanum and D. helicina are recorded for the first time from Mexico and Micropleura sp. is the first American representative of the genus recorded outside South America. Findings of A. americanum, Telorchis sp., P. groschafti, D. helicina and Micropleura sp. in C. moreletii represent new host records. Some observations on the early development of D. helicina are provided. All species, except for P. recurvum, are briefly described and illustrated and some problems concerning their morphology, taxonomy and geographical distribution are discussed. PMID- 11266136 TI - The prevalence of Borrelia burgdorferi sensu lato in Ixodes persulcatus and I. ricinus ticks in the zone of their sympatry. AB - A total of 7210 unfed adult Ixodes persulcatus Schulze, 1930 and I. ricinus (L., 1758) ticks were collected from the vegetation by flagging in 35 study sites located in the zone of their sympatry (mainly in Leningrad region, Russia). Borrelia infection in ticks was estimated by the dark-field microscopic analysis of gut contents in standard vital preparations at a magnification of x600. No correlation was revealed between the series of parameters characterising the abundance of each tick species (tau = -0.13) and between the series of these parameters and the prevalence of Borrelia in each vector. It is concluded that in the broad zone of I. persulcatus and I. ricinus sympatry, the presence and proportion of one vector in the ecosystem does not have any significant effect on the extensity of infection and on the epizootic and epidemic significance of the other vector. Each tick species has its independent (of the other species) and relatively original functional role in the focal ecosystem. PMID- 11266137 TI - Polyplax guatemalensis sp. n. (Phthiraptera: Anoplura), a new sucking louse from Peromyscus grandis, a montane cloud forest rodent from Guatemala. AB - The adult male and female of Polyplax guatemalensis sp. n. are described from the sigmodontine murid rodent Peromyscus grandis Goodwin collected in the Reserva de Biosfera, Sierra de las Minas, Guatemala, at an elevation of 2,200 m. The new species extends the number of known native species of Polyplax in the New World to four with none of them recorded south of Panama. Polyplax guatemalensis is morphologically most closely related to Polyplax auricularis which parasitises a cluster of closely related New World sigmodontine rodents from Canada to Panama. These two species can be distinguished from all other known species of Polyplax by the presence of partially overlapping, subtriangular, anterior abdominal plates in both sexes. Polyplax guatemalensis can be separated from P. auricularis by the abundant tergal abdominal setae and longer pseudopenis in males, and by the presence of one fewer anterior abdominal, subtriangular tergite and sternite in females. PMID- 11266138 TI - Caryospora varaniornati sp. n. (Apicomplexa: Eimeriidae) in the Nile monitor, Varanus (Polydaedalus) niloticus species complex. AB - Parasitological examination of two ornate Nile monitors Varanus ornatus (Daudin, 1803) imported from Benin revealed the presence of a new species of Caryospora. Oocysts of Caryospora varaniornati sp. n. are spherical to slightly subspherical, 12.0 (11-12.5) x 11.5 (11-12) microm, without amicropyle and oocyst residuum, and occasionally possessing one small polar granule. Sporocysts are broadly ellipsoidal, 8.8 (8.5-9.5) x 6.7 (6.5-7) microm; a lentil-like Stieda body is present, ca. 0.5 x 1 microm; substieda body not visible. Experimental infection of a closely related host, Varanus niloticus (L.), did not lead to the oocyst excretion despite the fact that one of the experimentally inoculated monitors was immunosuppressed by dexamethasone. Histological examination did not reveal stages of coccidian development. Therefore, it is possible that C. varaniornati is strictly host specific. PMID- 11266139 TI - A simple staining method for the visualisation of metacercariae in small fish and tadpoles. PMID- 11266140 TI - Cadmium and lead concentrations in Contracaecum rudolphii (Nematoda) and its host, the cormorant Phalacrocorax carbo (Aves). PMID- 11266141 TI - How confident can we be that acrylonitrile is not a human carcinogen? PMID- 11266143 TI - Physical exposure assessment in monotonous repetitive work--the PRIM study. AB - OBJECTIVES: A program called the Project on Research and Intervention in Monotonous Work (PRIM) was initiated in 1994 as a prospective cohort study of work-related musculoskeletal disorders. The group-based exposure assessment strategy, focusing on task-related exposure and used to obtain baseline measures of physical exposures, is reported in this paper. METHODS: Monotonous, repetitive worktasks were evaluated at 19 factories. Tasks with an estimated similarity in physical exposure were aggregated before 103 exposure groups were formed. Subjects from the exposure groups were randomly sampled for measurements, and task-related exposure levels were quantified by 43 single exposure items using a real-time video-based observation method that allowed computerized estimates of repetitiveness, body postures, force, and velocity. In combination with questionnaire-based data on task distribution, the duration of exposure was calculated at the individual level. RESULTS: The video-based observational method and the large number of exposure variables enabled the establishment of detailed quantitative exposure profiles in 103 task-based exposure groups. However, methodological problems associated with the use of grouped exposure assessment were revealed. Despite efforts to optimize group homogeneity, the within-group variance was larger than the between-group variance for several shoulder postural variables. CONCLUSIONS: A task-based exposure-assessment strategy can be successful in solving some of the main problems associated with the assessment of physical workplace exposures. The large within-group variance in exposure to nonneutral shoulder postures may eventually require individual assessment or the inclusion of groups with maximal contrast in exposure or both. PMID- 11266142 TI - Reconciling animal and human data in a cancer risk assessment of acrylonitrile. AB - OBJECTIVES: Bioassays of rats exposed to acrylonitrile have consistently detected an elevated incidence of central nervous system (CNS) cancer. In contrast, epidemiologic studies have not found a statistically stable increase in CNS cancer mortality. The purpose of this paper is to examine whether or not CNS cancers predicted from the most appropriate inhalation bioassay in rats are consistent with CNS cancers observed in 3 recent, large epidemiologic studies. METHODS: A linearized multistage model was fit to dose-response data from a rat inhalation bioassay to estimate carcinogenic potency. This potency was applied to epidemiologic studies of acrylonitrile-exposed workers. After adjustment for less than complete lifetime follow-up in the epidemiologic studies, consistency was examined between CNS cancers predicted by the model fit to the animal data for the exposure levels and sample sizes of the epidemiologicy studies and the CNS cancers observed in the epidemiologic studies. RESULTS: The model predicted totals of 17.7, 3.6, and 7.6 CNS cancer deaths for the studies. These predictions were not far from the observed CNS cancer deaths (12, 6, and 6) and were well within their 95% confidence intervals of 6.9-22.3, 2.2-13.1, and 2.2-13.1, respectively. CONCLUSIONS: The CNS cancer potency estimated from the best available inhalation bioassay was consistent with the observed deaths in the epidemiologic studies as long as continuous lifetime exposure was chosen as the exposure metric. The lack of observed excess in CNS cancer among the studied workers may have been due to low exposures, insufficient follow-up times, or both. PMID- 11266144 TI - Questionnaire versus direct technical measurements in assessing postures and movements of the head, upper back, arms and hands. AB - OBJECTIVES: This study compares questionnaire-assessed exposure data on work postures and movements with direct technical measurements. METHODS: Inclinometers and goniometers were used to make full workday measurements of 41 office workers and 41 cleaners, stratified for such factors as musculoskeletal complaints. The subjects answered a questionnaire on work postures of the head, back, and upper arms and repeated movements of the arms and hands (3-point scales). The questionnaire had been developed on the basis of a previously validated one. For assessing worktasks and their durations, the subjects kept a 2-week worktask diary. Job exposure was individually calculated by time-weighting the task exposure measurements according to the diary. RESULTS: The agreement between the self-assessed and measured postures and movements was low (kappa = 0.06 for the mean within the occupational groups and kappa = 0.27 for the whole group). Cleaners had a higher measured workload than office workers giving the same questionnaire response. Moreover, the subjects with neck-shoulder complaints rated their exposure to movements as higher than those without complaints but with the same measured mechanical exposure. In addition, these subjects also showed a general tendency to rate their postural exposure as higher. The women rated their exposure higher than the men did. CONCLUSIONS: The questionnaire assessed exposure data had low validity. For the various response categories the measured exposure depended on occupation. Furthermore, there was a differential misclassification due to musculoskeletal complaints and gender. Thus it seems difficult to construct valid questionnaires on mechanical exposure for establishing generic exposure-response relations in epidemiologic studies, especially cross-sectional ones. Direct technical measurements may be preferable. PMID- 11266145 TI - Questionnaire-based mechanical exposure indices for large population studies- reliability, internal consistency and predictive validity. AB - OBJECTIVES: This study attempts to construct valid indices for mechanical exposure of the shoulder-neck region with relation to the development of shoulder neck pain in a 1-year perspective study of a general population. METHODS: A comprehensive questionnaire was presented to 14 556 subjects aged 45 or 65 years and repeated after 12 months. Twenty-four questions concerning positions, movements, and manual materials handling were registered on a 3-point impact scale. Musculoskeletal problems were reported on a slightly modified version of the Standardized Nordic Questionnaire for the Analysis of Musculoskeletal Symptoms. Test-retest stability after 2 weeks was calculated for 232 consecutive participants. Based on mechanistic theories, 4 exposure indices were formed. Another 5 constructs were obtained by factor analysis. RESULTS: All the indices showed good test-retest stability, and 5 of them had very good internal consistency. Due to overlaps between the indices, 2 indices stood out as having unique properties. One of them concerned mainly postures and the other dealt primarily with measured lifting. However, the latter was not related to the shoulder-neck pain outcome when adjusted for the posture index. The posture index showed an exposure-effect relationship with the outcome. The job titles implied a large degree of exposure misclassification. CONCLUSIONS: The posture index is recommended as a mechanical exposure index for analyses of interaction with other possible determinants of shoulder-neck pain (ie, psychosocial factors). The use of such an index instead of job titles in large population studies will reduce the risk of misclassification. PMID- 11266146 TI - Prevalence and occupational associations of neck pain in the British population. AB - OBJECTIVES: This study determined the prevalence of neck pain and its relation to occupation and occupational activities in the general population. METHODS: A questionnaire was mailed to 21 201 subjects aged 16-64 years, randomly selected from the patient registers of general practices in England, Scotland, and Wales, and to 993 subjects randomly selected from pay records of the armed services. Information was collected on occupation, workplace physical activities, neck pain in the past week and year, headaches, and feelings of tiredness or stress. Associations were explored by logistic regression, the resultant odds ratios being converted to prevalence ratios (PR). RESULTS: Among 12907 respondents, 4348 and 2528 reported neck pain in past year (1421 with pain interfering with normal activities) and week, respectively. Symptoms were the most prevalent among male construction workers [past week and year 24% and 38% (pain interfering with activities 11%), respectively], followed by nurses, armed services members, and the unemployed. Generally the age-standardized prevalence of neck pain varied little by occupation. Work with arms above the shoulders for >1 hours/day was associated with a significant excess of symptoms [PR 1.3-1.7 (women) and 1.2-1.4 (men)], but no associations existed for typing, lifting, vibratory tool use, or professional driving. Stronger neck-pain associations were found with frequent headaches (PR 2.3-2.8) and frequent tiredness or stress (PR 2.2-2.5) than with occupational activities. CONCLUSIONS: The data provide evidence against a strong association between neck pain and the examined occupational physical activities. They suggest that psychosocial factors may be more important. PMID- 11266147 TI - Reevaluation of lung cancer risk in the acrylonitrile cohort study of the National Cancer Institute and the National Institute for Occupational Safety and Health. AB - OBJECTIVES: The present study provides additional analyses of data obtained earlier on lung cancer risk among workers with acrylonitrile exposure. METHODS: The original authors provided the data. For total mortality and the cancer sites of a priori interest (lung, stomach, brain, breast, prostate, and the lymphatic and hematopoietic systems), standardized mortality ratios (SMR) and 95% confidence intervals (95% CI) were computed, the total United States and surrounding counties being used as standard populations. Regional rate-based SMR values were also computed between lung cancer and cumulative acrylonitrile exposure. RESULTS: Except for lung cancer, the external comparisons corroborated the earlier internal comparisons (no increased cancer mortality risk). For lung cancer, the external comparisons revealed death deficits for the unexposed workers (SMR 0.68, 95% CI 0.5-0.9) and all categories of acrylonitrile-exposed workers. The SMR obtained using external rates and the most exposed group (SMR 0.92. 95% CI 0.6-1.4) differed from the corresponding relative risk (RR) of the internal rates (RR 1.5, 95% CI 0.9-2.4). CONCLUSIONS: The analysis of the present study provides little evidence that acrylonitrile exposure increases the mortality risk of cancers of a priori interest, including lung cancer. The lung cancer findings of the external comparison differed from the earlier findings of the internal comparisons. Selection bias (as the healthy worker effect) was probably not responsible. Additional follow-up and analyses, especially of the unexposed workers with low lung cancer rates, may help elucidate the internal and external comparison differences. Results from both comparisons should be presented when the relative risks differ markedly, as both have advantages and disadvantages. PMID- 11266148 TI - Life-style intervention at the worksite--reduction of cardiovascular risk factors in a randomized study. AB - OBJECTIVES: This study tested a feasible method for screening for cardiovascular risk at the worksite and investigated the effects of a long-term comprehensive program of life-style intervention to prevent cardiovascular disease. METHODS: Employees in the public sector filled out a self-administered questionnaire with questions on social, medical, and work-related factors. The respondents numbered 454 (80%). A score sum for cardiovascular risk was calculated (range 1-20, median 7.0), and the 128 subjects with a sum above 8 were invited to a health examination including blood sampling. Thereafter the subjects were invited to participate, following randomization, in a comprehensive, 18-month, life-style intervention program to improve cardiovascular risk or in a control group. RESULTS: The intervention group significantly decreased body mass index, diastolic blood pressure, heart rate, low-density lipoprotein (LDL) cholesterol, and smoking habits during the intervention. The initially elevated serum cortisol, as a marker of stress reaction, normalized in the intervention group. In the control group LDL cholesterol also decreased, but the glucose and triglyceride levels increased, and smoking habits were unchanged. Sick days for a given period decreased after 1 year in the intervention group but not in the control group. CONCLUSIONS: Several cardiovascular risk factors can be improved and morning serum cortisol normalized during a long-term life-style intervention program with a randomized design using a worksite population of middle-aged subjects. The use of a 2-step screening program, with an initial questionnaire followed by a health check of subjects with elevated risk, is feasible for worksite settings. PMID- 11266149 TI - Consequences of workplace bullying with respect to the well-being of its targets and the observers of bullying. AB - OBJECTIVES: This study investigated the effects of workplace bullying and the psychological work environment on the well-being and subjective stress of the targets and observers of bullying. METHODS: In a questionnaire study, stress and psychological ill-health were measured, and the causes of reported stress were analyzed for municipal employees (N=949, 85% women, 15% men, mean age 41 years for the men and 40 years for the women). RESULTS: Both the targets of bullying and the observers reported more general stress and mental stress reactions than did respondents from the workplaces with no bullying. The targets also expressed feelings of low self-confidence more often than did those who had not been subjected to bullying. Being bullied, but also features of one's work, especially haste, excessively difficult tasks and poor goal clarity, predicted the stress reactions reported. Of the single forms of bullying, judging a person's work unjustly or in an offending manner, restricting a person's possibilities to express his or her opinions, and assaulting one's private life were the most clearly connected with all the stress reactions measured. Victim history was associated with feelings of low self-confidence. The targets of bullying used sleep-inducing drugs and sedatives more often than did the respondents who were not bullied. CONCLUSIONS: The study shows that not only the targets of bullying, but also bystanders, suffer when someone is bullied in the workplace. Bullying must therefore be regarded as a problem for the entire work unit and not merely as a problem of the target. PMID- 11266150 TI - Comparison of expert-rater methods for assessing psychosocial job strain. AB - OBJECTIVES: This study tested the reliability and validity of industry- and mill level expert methods for measuring psychosocial work conditions in British Columbia sawmills using the demand-control model. METHODS: In the industry-level method 4 sawmill job evaluators estimated psychosocial work conditions at a generic sawmill. In the mill-level method panels of experienced sawmill workers estimated psychosocial work conditions at 3 sawmills. Scores for psychosocial work conditions were developed using both expert methods and applied to job titles in a sawmill worker database containing self-reported health status and heart disease. The interrater reliability and the concurrent and predictive validity of the expert rater methods were assessed. RESULTS: The interrater reliability and concurrent reliability were higher for the mill-level method than for the industry-level method. For all the psychosocial variables the reliability for the mill-level method was greater than 0.90. The predictive validity results were inconclusive. CONCLUSIONS: The greater reliability and concurrent validity of the mill-level method indicates that panels of experienced workers should be considered as potential experts in future studies measuring psychosocial work conditions. PMID- 11266152 TI - Study on postural stress and shoulder disorders. PMID- 11266151 TI - Asthma risk, cleaning activities and use of specific cleaning products among Spanish indoor cleaners. AB - OBJECTIVES: Recent studies have shown an excess risk of asthma for cleaners, but it is not clear which cleaning-related exposures induce or aggravate asthma. METHODS: Risk factors for asthma were studied among indoor cleaners participating in the Spanish part of the European Community Respiratory Health Survey in 1992. In 1998, 78 of the 91 subjects reporting cleaning-related jobs in 1992 were identified. Of these, 67 indoor cleaners were interviewed by telephone about their cleaning activities and their use of cleaning products in 1992. These data were related to asthma prevalence in 1992, and the cleaners were compared with a reference group of office workers. RESULTS: Asthma prevalence was 1.7 times higher [95% confidence interval (95% CI) 1.1-2.6] among the cleaners than among the referents, being highest among private home cleaners (3.3, 95% CI 1.9-5.8). The prevalence of housedust mite sensitization amounted to 28% for the home cleaners and was significantly (P<0.01) higher than for other indoor cleaners (3%), but similar to the corresponding prevalence of office workers (22%). More than half of the cleaners reported work-related respiratory symptoms. The asthma risk of the home cleaners was mainly associated with kitchen cleaning and furniture polishing, with the use of oven sprays and polishes. CONCLUSIONS: The asthma risk of Spanish cleaners is primarily related to the cleaning of private homes. This relationship may be explained by the use of sprays and other products in kitchen cleaning and furniture polishing. PMID- 11266153 TI - Syntheses and evaluation of pyrido[2,3-dlpyrimidine-2,4-diones as PDE 4 inhibitors. AB - The syntheses and in vitro evaluation of a new series of pyrido[2,3-d]pyrimidine 2,4-diones bearing substituents at C-3 and/or C-4 positions on the pyridine ring are described. Some of these compounds, especially 51 and 6f, were found to be potent phosphodiesterase 4 (PDE 4) inhibitors exhibiting improved ratio of PDE 4 inhibitory activity:rolipram binding assay (RBA). PMID- 11266154 TI - Synthesis and separation of diastereomers of uridine 2',3'-cyclic boranophosphate. AB - The first boron-containing 2',3'-cyclic phosphate-modified analogue, uridine 2',3'-cyclic boranophosphate (2',3'-cyclic-UMPB), was synthesized. 5'-O-Protected uridine was cyclophosphorylated by diphenyl H-phosphonate to yield uridine 2',3' cyclic H-phosphonate, which upon silylation followed by boronation and subsequent acid treatment gave 2',3'-cyclic-UMPB in high yield. The two diastereomers of 2',3'-cyclic-UMPB were separated by HPLC. An alternative method for synthesis of uridine 2',3'-cyclic phosphorothioate (2',3'-cyclic-UMPS) via H-phosphonate was also described. PMID- 11266156 TI - 8-Carboxamidocyclazocine analogues: redefining the structure-activity relationships of 2,6-methano-3-benzazocines. AB - Unexpectedly high affinity for opioid receptors has been observed for a novel series of cyclazocine analogues where the prototypic 8-OH was replaced by a carboxamido group. For mu and kappa opioid receptors, the primary carboxamido derivative of cyclazocine ((+/-)-15) displayed high affinity (Ki=0.41 and 0.53 nM, respectively) nearly comparable to cyclazocine. A high enantiopreference ((2R,6R,11R)-) for binding was also observed. Compound (+/-)-15 also displayed potent antinociception activity in mice when administered icv. PMID- 11266155 TI - Synthesis and biological activities of potential metabolites of the non nucleoside reverse transcriptase inhibitor efavirenz. AB - Studies on the biotransformation of the clinically important non-nucleoside reverse transcriptase inhibitor efavirenz have shown that oxidation and secondary conjugation are important components of the processing of this molecule in vivo. We have synthesized metabolites of efavirenz to confirm their structure and to evaluate their activity as antivirals. PMID- 11266157 TI - Chemoenzymatic synthesis of functionalized cyclohexylglycines and alpha methylcyclohexylglycines via Kazmaier-Claisen rearrangement. AB - The synthesis of homochiral functionalized cyclohexylglycines and alpha methylcyclohexylglycines via chelated Kazmaier-Claisen rearrangement is described. These were shown to be potent scaffolds for the development of MMP inhibitors. PMID- 11266158 TI - Synthesis and structure-activity relationships of 5-substituted pyridine analogues of 3. AB - In an effort to probe the steric influence of C5 substitution of the pyridine ring on CNS binding affinity, analogues of 1 substituted with a bulky moiety- such as phenyl, substituted phenyl, or heteroaryl-were synthesized and tested in vitro for neuronal nicotinic acetylcholine receptor binding affinity. The substituted analogues exhibited Ki values ranging from 0.055 to 0.69 nM compared to a Ki value of 0.15 nM for compound 1. Assessment of functional activity at subtypes of neuronal nicotinic acetylcholine receptors led to identify several agonists and antagonists. PMID- 11266159 TI - 3-Anilino-4-arylmaleimides: potent and selective inhibitors of glycogen synthase kinase-3 (GSK-3). AB - Potent 3-anilino-4-arylmaleimide glycogen synthase kinase-3 (GSK-3) inhibitors have been prepared using automated array methodology. A number of these are highly selective, having little inhibitory potency against more than 20 other protein kinases. PMID- 11266160 TI - Synthesis and SAR of benzamidine factor Xa inhibitors containing a vicinally substituted heterocyclic core. AB - The selective inhibition of coagulation factor Xa has emerged as an attractive strategy for the discovery of novel antithrombotic agents. Here we describe highly potent benzamidine factor Xa inhibitors based on a vicinally-substituted heterocyclic core. PMID- 11266161 TI - Importance of the C28-C38 hydrophobic domain of okadaic acid for potent inhibition of protein serine-threonine phosphatases 1 and 2A. AB - Okadaic acid is a potent inhibitor of select serine/threonine protein phosphatases. The importance of the C28-C38 hydrophobic domain of okadaic acid for inhibition of PP1 and PP2A was investigated. The hydrophobic domain is required but not sufficient for potent inhibition, and it also contributes to differential inhibition between PP1 and PP2A. PMID- 11266162 TI - Anion recognition by thiostrepton. AB - A bicyclic polypeptide antibiotic thiostrepton forms both 1:1 and 1:2 complexes with anions (as tetrabutylammonium salts) in organic solvents with K2 >K1 for F- and K2<>Cl-, Br-, HSO4-, H2PO4-, but in CHCl3 they follow a different order: Cl- approximatelyHSO4- >F- approximately AcO- > Br > H2PO4-. The binding mode of anions to thiostrepton is discussed on the basis of solvent effects on the complexation selectivity. PMID- 11266164 TI - Synthesis, modelling and NK1 antagonist evaluation of a non-rigid cyclopropane containing analogue of CP-99,994. AB - A non-rigid cyclopropane-containing diamine analogue of CP-99,994 was synthesised and was found to have only moderate NK1 receptor binding affinity. Molecular dynamics calculations of the conformational space of the former compound gave good correlation between observed activity and a recently published pharmacophore model, lending predictive value to the latter. PMID- 11266163 TI - Influence of chain length and N-alkylation on the selective serotonin receptor ligand 9-(aminomethyl)-9,10-dihydroanthracene. AB - Comparison of the serotonin 5-HT2A receptor affinities of chain lengthened and N alkylated analogues of the novel ligand 9-aminomethyl-9,10-dihydroanthracene (AMDA) and a structurally similar prototypical tricyclic amine imipramine suggests that the two agents bind to the receptor in different fashions. The demonstration that AMDA is highly selective for serotonin receptors (5-HT2A, K = 20nM; 5-HT2C, Ki=43nM) versus the dopamine D2 receptor (Ki>10,000nM), as well as the serotonin and norepinephrine transporters (Ki>10,000nM) further suggests that AMDA and the nonselective ligand imipramine interact with these target macromolecules in different ways. PMID- 11266165 TI - Addressing the stability of C-capped dipeptide efflux pump inhibitors that potentiate the activity of levofloxacin in Pseudomonas aeruginosa. AB - Synthetic optimization of a biologically labile class of dipeptides that function as efflux pump inhibitors to potentiate the antibacterial agent levofloxacin in Pseudomonas aeruginosa has led to the discovery of a related series of compounds that are completely stable in a variety of biological matrices. Other than the stability profile, the in vitro profile of the new series is essentially identical to that observed with the original one. A prototypical compound from the new series demonstrates potentiation in an in vivo model of infection. PMID- 11266167 TI - Customized versus universal scoring functions: application to class I MHC-peptide binding free energy predictions. AB - A tailor-made free energy scoring method (Fresno) has been compared to six universal scoring functions (Chemscore, Dock, FlexX, Gold, Pmf, Score) for predicting the binding affinity of 26 peptides to the class I human major histocompatibility protein HLA-B*2705. Fresno clearly outperforms all six universal scoring functions. PMID- 11266166 TI - Sialidase inhibitors related to zanamivir. Further SAR studies of 4-amino-4H pyran-2-carboxylic acid-6-propylamides. AB - SAR investigations of the 4- and 5-positions of a series of 4-amino-4H-pyran-2 carboxylic acid 6-carboxamides are reported. Potent inhibitors of influenza A sialidase with marked selectivity over the influenza B enzyme were obtained when the basic 4-amino substituent was replaced by hydroxyl or even deleted. Modifications at the 5-position exhibited a tight steric requirement, with trifluoroacetamide being optimal. PMID- 11266169 TI - Design and synthesis of novel 2,3-dihydro-1H-isoindoles with high affinity and selectivity for the dopamine D3 receptor. AB - Starting from the tetrahydroisoquinoline SB-277011 1, a novel series of 5 substituted-2,3-dihydro-1H-isoindoles has been designed. Subsequent optimisation resulted in identification of 19, which has high affinity for the dopamine D3 receptor (pKi 8.3) and > or = 100-fold selectivity over other aminergic receptors. In rat studies 19 was brain penetrant with an excellent pharmacokinetic profile (oral bioavailability 77%, t1/2 5.2h). PMID- 11266168 TI - Gamma-lactone-Functionalized antitumoral acetogenins are the most potent inhibitors of mitochondrial complex I. AB - To study the relevance of the terminal alpha,beta-unsaturated gamma-methyl-gamma lactone moiety of the antitumoral acetogenins of Annonaceae for potent mitochondrial complex I inhibition, we have prepared a series of semisynthetic acetogenins with modifications only in this part of the molecule, from the natural rolliniastatin-1 (1) and cherimolin-1 (2). Some of the hydroxylated derivatives (1b, 1d and 1e) in addition to two infrequent natural beta-hydroxy gamma-methyl gamma-lactone acetogenins, laherradurin (3) and itrabin (4), are more potent complex I inhibitors than any other known compounds. PMID- 11266170 TI - The synthesis of beta-peptides containing guanidino groups. AB - The synthesis of the beta-peptide 1 by the postsynthetic modification of the corresponding amino-containing peptide 3 is described. The potential of 1 to act as a template for the ligation of complementary negatively-charged peptides is discussed. PMID- 11266171 TI - RPR203494 a pyrimidine analogue of the p38 inhibitor RPR200765A with an improved in vitro potency. AB - Following the discovery of RPR200765, a series of pyrimidine analogues have been prepared as backups. Amongst them, RPR203494 was identified with a better in vitro profile than RPR200765A. PMID- 11266172 TI - Carboproteins: a 4-alpha-helix bundle protein model assembled on a D galactopyranoside template. AB - We have recently introduced the concept of monosaccharides as templates for de novo design of protein models and described the synthesis of a model 'carbopeptide'. Here, we report the synthesis of a 64 amino acid (AA) 'carboprotein' by chemoselective ligation of a C-terminal hexadecapeptide aldehyde to a tetra-aminooxy functionalized methyl alpha-D-galactopyranoside (D Galp) template. Biophysical characterizations by CD spectroscopy and NMR amide H D exchange experiments indicated that the four-stranded carboprotein forms a 4 alpha-helix bundle structure. PMID- 11266173 TI - The identification of a potent, water soluble inhibitor of lipoprotein-associated phospholipase A2. AB - Modification of the pyrimidone 5-substituent in a series of 1-((amidolinked) alkyl)-pyrimidones, lipophilic inhibitors of lipoprotein-associated phospholipase A2, has given inhibitors of nanomolar potency and improved physicochemical properties. Compound 23 was identified as a potent, highly water soluble. CNS penetrant inhibitor suitable for intravenous administration. PMID- 11266174 TI - Design of non-peptide CCK2 and NK1 peptidomimetics using 1-(2 nitrophenyl)thiosemicarbazide as a novel common scaffold. AB - A beta-turn overlay hypothesis has been used to transform the core scaffold of a selective non-peptide bradykinin B2 receptor antagonist into ligands specifically recognized by the CCK2 or NK1 receptors. PMID- 11266175 TI - Synthetic analogues of SB-219383. Novel C-glycosyl peptides as inhibitors of tyrosyl tRNA synthetase. AB - Novel inhibitors of bacterial tyrosyl tRNA synthetase have been synthesised in which the cyclic hydroxylamine moiety of SB-219383 is replaced by C-pyranosyl derivatives. Potent and selective inhibition of bacterial tyrosyl tRNA synthetase was obtained. PMID- 11266177 TI - The C4 hydroxyl group of phorbol esters is not necessary for protein kinase C binding. AB - To investigate the role of the hydroxyl group at position 4 of the phorbol esters in protein kinase C (PKC) binding and function, 4beta-deoxy-phorbol-12,13 dibutyrate (4beta-deoxy-PDBu, 5a) and 4beta-deoxy-phorbol-13-acetate (6a) were synthesized from phorbol (1). The binding affinities of these 4beta-deoxy compounds (5a, 6a) to the 13 PKC isozyme C1 domains were quite similar to those of the corresponding 4beta-hydroxy compounds (4a, 4b), suggesting that the C4 hydroxyl group of phorbol esters is not necessary for PKC binding. Moreover, functional assays showed that 4beta-deoxy-PDBu (5a) exhibited biological activities (Epstein-Barr virus induction and superoxide generation) equally potent to those of PDBu (4a). These solution phase results differ from expectations based on the previously reported solid-phase structure of the complex of PKCdelta-C1B and phorbol-13-acetate (4b). PMID- 11266176 TI - Potent synthetic inhibitors of tyrosyl tRNA synthetase derived from C-pyranosyl analogues of SB-219383. AB - Novel pyranosyl analogues of SB-219383 have been synthesised to elucidate the structure-activity relationships around the pyran ring. Analogues with highly potent stereoselective and bacterioselective inhibition of bacterial tyrosyl tRNA synthetase have been identified. A major reduction in the overall polarity of the molecule can be tolerated without loss of the nanomolar level of inhibition. PMID- 11266178 TI - The amide hydrogen of (-)-indolactam-V and benzolactam-V8's plays a critical role in protein kinase C binding and tumor-promoting activities. AB - To investigate the role of the amide hydrogen of (-)-indolactam-V (1) and benzolactam-V8's on protein kinase C (PKC) binding and tumor promotion, 8 decylbenzolactone-V8 (6), a new lactone analogue of 8-decylbenzolactam-V8 (4), was synthesized from 2-nitrophenylpyruvic acid (7) in 11 steps. The PKC binding ability and tumor-promoting activities in vitro of 6 were much lower than those of 1 and 4, suggesting that the amide hydrogen of 1 and benzolactam-V8's plays a critical role in tumor promotion. However, it is noteworthy that 6 showed significant selectivity in the PKC isozyme surrogate binding. PMID- 11266179 TI - Formaldehyde-mediated modification of natural deoxyguanosine with amines: one-pot cyclization as a molecular model for genotoxicity. AB - Stable cross-linked adducts, 3-(2-deoxy-beta-D-ribofuranosyl)-7-phenyl-5,6,7,8 tetrahydro[1,3,5]triazino[1,2-a]purin-10(3H)-one and 7-butyl-3-(2-deoxy-beta-D ribofuranosyl)-5,6,7,8-tetrahydro[1,3,5]triazino[1,2-a]purin-10(3H)-one, that formed chemically from natural deoxyguanosine and aniline or buthyl amine in the presence of formaldehyde were identified. This reaction appears to be a general reaction of deoxyguanosine and primary amines, and it may be a model of DNA modification with carcinogenic aromatic amines without metabolic activation, if formaldehyde is present. PMID- 11266180 TI - The design of phenylglycine containing benzamidine carboxamides as potent and selective inhibitors of factor Xa. AB - Factor Xa, a critical serine protease in the blood coagulation cascade, has become a target for inhibition as a strategy for the invention of novel anti thrombotic agents. Here we describe the development of phenylglycine containing benzamidine carboxamides as novel, potent and selective inhibitors of factor Xa. PMID- 11266182 TI - Synthesis of substituted imidazopyrazines as ligands for the human somatostatin receptor subtype 5. AB - A new preparation of trisubstituted imidazopyrazines and dihydroimidazopyrazines via parallel synthesis using aminoacids and bromoketones resulted in the discovery of non-peptidic sst5 selective agonists. PMID- 11266181 TI - Structure-activity analysis of truncated orexin-A analogues at the orexin-1 receptor. AB - Truncated peptide analogues of orexin-A were prepared and their biological activity assesed at the orexin-1 receptor. Progressive N-terminal deletions identified the minimum C-terminal sequence required for maintaining a significant agonist effect, whilst an alanine scan and other pertinent substitutions identified key side-chain and stereochemical requirements for receptor activation. PMID- 11266183 TI - Structure-activity relationship on the human EP3 prostanoid receptor by use of solid-support chemistry. AB - Potent and selective EP3 receptor ligands were found by making a library using solid-support chemistry. These compounds can be obtained by a Suzuki coupling reaction of a solid-supported benzyl bromide using various boronic acids. The yields obtained for this reaction were in the range of 24-95% of arylmethyl cinnamic acid 1 after cleavage from the Wang resin. PMID- 11266184 TI - Diagnosis of carotid artery atheroma by magnetic resonance imaging. AB - Atheroma appears as a very low signal intensity area on 2-dimensional time-of flight (TOF) magnetic resonance (MR) images, and its components have various signal intensities on spin-echo (SE) images. The present study investigated atheroma of the carotid arteries in 37 subjects with risk factors (63+/-10 years of age; 19 men) by magnetic resonance imaging (MRI). On 2-dimensional (2D) TOF images, the carotid arteries were clearly demonstrated in all cases and atheroma was detected in 23 patients. The most common location of atheroma was at the origin of the internal carotid artery. There was vascular remodeling in all patients with atheroma. 2D-TOF images showed 97% agreement with ultrasonography. SE images clearly demonstrated atheroma in all 23 patients with atheroma. All patients with atheroma showing high signal intensity on T1-weighted images had hyperlipidemia. These findings indicate that the 2D-TOF imaging method is useful for detecting atheroma and SE-images are useful for its characterization. PMID- 11266185 TI - Appropriate indications for the use of a percutaneous cardiopulmonary support system in cases with cardiogenic shock complicating acute myocardial infarction. AB - Percutaneous cardiopulmonary support (PCPS) is now available for hemodynamic support in patients with cardiogenic shock, but there are no guidelines for its use. The present study determined the appropriate indications for the use of the PCPS in patients with cardiogenic shock complicating acute myocardial infarction (AMI). Sixty-four consecutive patients with cardiogenic shock complicating AMI had hemodynamic support with an intraaortic balloon pump (IABP; n=38) and/or PCPS (n=26). The shock score (0-15) was calculated immediately before starting these support systems to quantify the severity of shock. Multivariate logistic regression analysis determined the clinical factors affecting in-hospital mortality. The relationship between in-hospital prognosis and the shock score was also examined in the 2 groups. The most significant factor related to the in hospital prognosis was the shock score (p=0.0007; OR 2.16, 95% CI: 1.37-3.39). Another related factor was revascularization; however, this relationship did not reach statistical significance (p=0.069; OR 0.06). Among the 13 cases whose shock score was 4-8 (moderate shock), 5 survived in the PCPS group, but only 1 of 19 patients survived in the IABP group (p<0.05). None of the patients in either group whose shock score was more than 9 survived. The severity of shock is the most reliable independent predictor of in-hospital mortality in patients with cardiogenic shock complicating AMI. Using PCPS in patients with moderate cardiogenic shock may improve their in-hospital survival, but it must be used before the shock becomes severe. PMID- 11266186 TI - Electrocardiographic ST-segment elevation and changes in the regional work of the left ventricle during coronary angioplasty. AB - This study evaluated the reduction in regional work of the left ventricle caused by acute myocardial ischemia during coronary angioplasty, and correlated it with ST-segment elevation. Regional work of the left ventricular myocardium, which is derived from a stress-strain loop, is a useful index of the function of a diseased heart. However, the effects of transient ischemia on the regional work of the myocardium have not been fully elucidated. The subjects consisted of 25 patients who had proximal left anterior descending artery stenosis with normal wall motion and without collateral circulation. The patients were classified as showing ST-segment elevation > or = 0.2 mV (group A, 10 patients), or ST-segment elevation < 0.2mV (group B, 15 patients) during coronary angioplasty. Group A showed a greater reduction in the regional work of the interventricular septum than group B. Regional work recovered to the baseline level 30 s after balloon deflation in group B, but took 40 s in group A. A greater ST elevation during balloon inflation was associated with a greater, prolonged reduction of work performance in the ischemic region and a greater concomitant increase in the opposite nonischemic region. PMID- 11266187 TI - Prognostic value of iodine-123-metaiodobenzylguanidine imaging and cardiac natriuretic peptide levels in patients with left ventricular dysfunction resulting from cardiomyopathy. AB - This study assessed the prognostic value of Iodine-123-metaiodobenzylguanidine (MIBG) imaging and of the plasma level of cardiac natriuretic peptides in patients with left ventricular dysfunction resulting from cardiomyopathy. Predictors of cardiac death or hospitalization related to progressive heart failure were examined in 171 patients with chronic heart failure (96 patients with idiopathic cardiomyopathy and 75 patients with ischemic cardiomyopathy). All patients underwent MIBG imaging at rest and other hemodynamic studies. During a mean (+/-SD) follow-up period of 27+/-11 months, 11 patients died from heart failure and 16 required hospitalization. High MIBG washout was an independent predictor of cardiac death (relative risk [RR] = 1.158, p<0.0001) whereas the plasma level of brain natriuretic peptide (BNP: relative risk [RR] = 1.005, p<0.0001) and high MIBG washout (relative risk [RR] = 1.094, p<0.0001) were predictors of progressive heart failure (ie, combined cardiac death and hospitalization). Accelerated myocardial adrenergic nerve activity as assessed by MIBG imaging and the plasma levels of BNP are powerful predictors of the patient's prognosis. PMID- 11266188 TI - Atheromatous plaque in the distal aortic arch creating the potential for cerebral embolism during cardiopulmonary bypass. AB - The present study evaluated the risk in cardiac patients of rupture of a plaque by a jet stream from the arch cannula. The entire thoracic aorta and cardiac function were routinely monitored by transesophageal echocardiography (TEE) in 88 adult patients who underwent coronary artery bypass surgery. The changes in the atheromatous plaque in the distal aortic arch were observed before and after cardiopulmonary bypass. Of the 88 patients, 13 were found to have preoperative atheromatous plaque at the distal aortic arch and 8 (61.5%) of them suffered plaque rupture caused by jet stream from the arch cannula. Only 1 patient experienced apparent embolic episodes manifesting as cerebral and left leg embolisms; the remaining 7 had no clinical embolic symptoms. In order to prevent atheroembolic events, attention should be paid not only to the ascending aorta, but also to the distal arch and in this regard TEE is useful for detecting atheromatous changes of the aorta. PMID- 11266189 TI - Level of plasma tissue factor pathway inhibitor is inversely correlated with intraarterial diastolic pressure in subjects who underwent coronary angiography. AB - There are only a few studies of the relationship between hemostatic abnormalities and intraarterial pressure, so the present study investigated the association of various newer lipid and hemostatic variables with intraarterial pressure levels. Levels of total cholesterol, high-density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol, triglyceride, lipoprotein-(a), remnant-like particle cholesterol, cholesteryl ester transfer protein, uric acid, blood glucose, fibrinogen, free form of tissue factor pathway inhibitor (TFPI), C reactive protein, serum amyloid A protein, anti-Chlamydia pneumoniae immunoglobulin G and immunoglobulin A, and apolipoproteins (apo) A-I, B, and E were measured in 176 patients who underwent diagnostic coronary angiography. Intraarterial blood pressure was determined from central aortic pressure using a standard fluid-filled catheter-external transducer system. Multivariate regression analyses showed that TFPI level was the only independent factor associated with aortic diastolic pressure. The linear regression equation demonstrated a significant negative correlation of TFPI level with aortic diastolic pressure (r=-0.395, p=0.0011). With respect to the association with other parameters, the TFPI level showed significant correlations between the HDL C level and the apo A-I level, both in the overall patients and in the patients with coronary artery stenosis. This is the first evidence that the level of the plasma free form of TFPI is inversely correlated to aortic diastolic pressure. PMID- 11266190 TI - Diagnostic value of pulsed Doppler echocardiography in acute pulmonary thromboembolism--comparison with pulmonary angiography and pulmonary artery pressure. AB - The ratio of acceleration time to right ventricular ejection time (AcT/RVET) can be derived from the blood flow patterns recorded by pulsed wave Doppler echocardiography. In chronic cor pulmonale, AcT/RVET negatively correlates with pulmonary artery pressure (PAP). The present study evaluated the diagnostic value of AcT/RVET by comparing this variable with indices derived from pulmonary angiography (PAG) and PAP in 16 patients with acute pulmonary thromboembolism (APTE). AcT/RVET, PAP, and PAG severity indices (Miller index (MI) and UPET objective angiographic index (UI)) were measured during the acute phase on admission and the chronic phase after treatment. In the acute phase, AcT/RVET correlated with mean PAP (mPAP) (r=-0.68, p<0.05) and total pulmonary resistance (TPR) (r=-0.66, p<0.05), but not with MI or UI. During the chronic phase, AcT/RVET did not correlate with mPAP or TPR, but with both PAG indices (MI: r=0.76, p<0.05, UI: r=0.65, p<0.05). Before treatment of the APTE, AcT/RVET remained at low levels and could be used as an index of pulmonary hypertension. After treatment, however, following improvement of PAP, AcT/RVET was not useful for evaluating PAP, but might serve as an index for evaluating the volume of residual thrombi. PMID- 11266191 TI - Prediction of functional recovery in patients with myocardial infarction after revascularization--comparison of low-dose dobutamine stress echocardiography with fluorine-18 fluorodeoxyglucose positron emission tomography. AB - The present study investigated the agreement between low-dose dobutamine stress echocardiography (LDDSE) and fluorine-18 fluorodeoxyglucose positron emission tomography (FDG-PET) and compared each technique's ability to detect myocardial viability and predict functional recovery in 30 patients. All patients underwent revascularization, followed by echocardiography 5+/-3 months. Of the 390 segments analyzed by echocardiography before revascularization, 110 (28%) had abnormal wall motion. LDDSE showed viability in 66 sites of the 110 dyssynergic segments and 58 of these viable segments recovered their wall motion. With FDG-PET, 78 of the 110 dyssynergic segments were diagnosed as viable and 62 of these showed improvement of the wall motion. The sensitivities for LDDSE and FDG-PET to assess functional recovery were 84% and 90%, respectively; specificities were 80% and 64%, respectively. Positive predictive values for LDDSE and FDG-PET were 88% and 79%; negative predictive values were 75% and 78%, respectively. Both methods had good sensitivity for detecting improvement in regional function after revascularization, but LDDSE had a higher specificity for detecting viability and a better positive predictive value for left ventricular functional recovery. PMID- 11266192 TI - Association of seropositivity for antibody to Chlamydia-specific lipopolysaccharide and coronary artery disease in Japanese men. AB - Recent studies suggest an association between Chlamydia pneumoniae infection and coronary artery disease (CAD). To examine this relationship in Japanese men, serum IgA and IgG antibodies to Chlamydia-specific lipopolysaccharide were measured by enzyme-linked immunosorbent assay in 507 patients with CAD and 200 age-matched controls. CAD patients were divided into (1) 269 patients with myocardial infarction (MI) and (2) 238 patients with chronic coronary heart disease (CCHD). Compared with the control group, the CAD group did not differ in the prevalences of both antibodies (IgA: 23.7 vs 18.0%, p=0.10; IgG: 52.7 vs 51.0%, p=0.6). The index of IgG antibody was not significantly different between CAD and control groups (median 1.19 vs 1.18, p=0.3), whereas the index of IgA antibody was significantly higher in CAD than control group (median 0.60 vs 0.46, p<0.0001). Compared with the control group, the MI group had a significantly higher prevalence of IgA antibody (28.6 vs 18.0%, p=0.007); however, there was no difference in the prevalence of IgG antibody (58.0 vs 51.0%, p=0.13). The CCHD group did not differ in the prevalences of both antibodies (IgA: 18.1 vs 18.0%, p=0.9; IgG: 45.6 vs 51.0%, p=0.2). After the adjustment for coronary risk factors, odds ratios (ORs) of seropositive antibodies for CAD were 1.59 [95% confidence interval (CI): 0.88-2.87, p=0.12] for IgA seropositivity and 0.92 (95%CI: 0.58-1.47, p=0.7) for IgG seropositivity in all cases. In the MI and control groups, ORs of seropositive antibodies for MI were 2.67 (95%CI: 1.32 5.38, p=0.007) for IgA seropositivity, and 1.36 (95%CI: 0.79-2.36, p=0.2) for IgG seropositivity. This study discovered that IgA antibody to Chlamydia was significantly associated with CAD, especially with MI, in Japanese Men and the findings suggest that chronic infection of Chlamydia may be linked to the pathogenesis of MI. PMID- 11266193 TI - Helical computer assisted tomography in pulmonary hypertension complicating left to-right shunts--correlation with pulmonary hemodynamics. AB - The present study analyzed the helical computer-assisted tomography (CAT) findings in 30 patients with pulmonary hypertension (PH) associated with left-to right shunts; specifically, ventricular septal defect, 23; atrioventricular septal defect, 6; patent ductus arteriosus, 1. Eight patients had 21 trisomy. Age ranged from 1 to 18 (mean, 4.1) months, and body weight ranged from 2.6 to 10.7 (mean, 4.9) kg. In all patients, the chest CAT revealed patchy areas of high and low attenuation (mosaic pattern) and regional atelectasis in the lung fields. The volume of low attenuated lesions and of atelectasis, and the total lung volume were derived from integration of areas measured on the CAT image. The ratios of low attenuated lesion/total lung volume (Lo), volume of atelectasis/ total lung volume (Ate) and low attenuated lesion and volume of atelectasis/total lung volume (Lo&Ate) were compared with hemodynamic parameters measured at cardiac catheterization. The pulmonary to systemic resistance ratio correlated with Lo (r=0.61, p<0.01) and Lo&Ate (r=0.69, p<0.01), whereas the pulmonary vascular resistance correlated with Ate (r=0.53, p<0.01). Lo, Ate and Lo&Ate in the chest CAT are reliable parameters that can be used to estimate pulmonary vascular resistance in patients with PH associated with left-to-right shunts. PMID- 11266194 TI - Visceral fat accumulation contributes to insulin resistance, small-sized low density lipoprotein, and progression of coronary artery disease in middle-aged non-obese Japanese men. AB - Visceral fat accumulation plays an important role in the occurrence of coronary artery disease (CAD) associated with a cluster of multiple risk factors, such as glucose intolerance, insulin resistance and hyperlipoproteinemia. To clarify the detailed features of these factors, based on visceral fat accumulation, the present study examined the relationship between fat distribution and the characteristics of glucose metabolism and serum lipoproteins in middle-aged non obese Japanese men. First, the influence of visceral fat accumulation on glucose metabolism, insulin sensitivity, and the extent and severity of coronary artery lesions was investigated in 50 subjects with CAD and compared with 15 control subjects without CAD (Study 1) and with the lipoprotein characteristics in 44 subjects without CAD who were not treated with lipid-lowering drugs (Study 2). Body fat distribution was determined by abdominal computed tomography. In Study 1, the visceral fat area (VFA), blood pressure, fasting immunoreactive insulin (FIRI), and the plasma insulin area (PIA) obtained by oral glucose tolerance test in the subjects with CAD were all significantly higher than in the control subjects. The VFA was significantly correlated with FIRI, the homeostasis model of insulin resistance, PIA and steady state plasma glucose (SSPG) concentration as an index for insulin resistance (r=0.57, p<0.001, r=0.49, p<0.01, r=0.36, p<0.01, and r=0.50, p<0.05, respectively). Although the SSPG concentration did not correlate with the coronary atherosclerosis index as a score of the extent and severity of coronary lesions, the VFA was significantly correlated with this index (r=0.43, p<0.01). In Study 2, the VFA had significant positive correlations with serum total cholesterol, triglyceride, and apolipoprotein B and E levels and the cholesterol and triglyceride concentrations of very-low-density lipoprotein, intermediate-density lipoprotein, and low-density lipoprotein (LDL) fractions. There was a negative correlation between the VFA and LDL particle size (r=-0.34, p<0.05). In conclusion, visceral fat accumulation may contribute to the development of CAD through the progression of insulin resistance and the increase of apo B-containing lipoproteins and small-sized LDLs in middle-aged non-obese Japanese men. PMID- 11266195 TI - Case-control study of nonfatal myocardial infarction in relation to selected foods in Japanese men and women. AB - Most studies of diet and coronary heart disease (CHD) have focused on constituents rather than on whole foods. The present study examined the relationship of selected foods to nonfatal acute myocardial infarction (AMI) in Japan, with special reference to vegetables, fruits, fish, and tofu. A total of 660 cases with their first episode of AMI aged 40-79 years living in Fukuoka City or adjacent areas and 1,277 controls matched for age, sex, and residence were surveyed on lifestyle, including dietary factors. Participation rates were 87% of cases and 52% of controls. Consumption frequencies of 19 food/beverages items and daily amounts of 4 items were ascertained by interview. The final analysis was done with 632 cases and 1,214 controls. Although consumption of vegetables showed no clear association with the risk of AMI, fruit consumption appeared to reduce the risk of AMI in both men and women. The results also suggested that fish consumption was related to a decreased risk of AMI in men, although the trend was not statistically significant. In women only, tofu consumption was inversely related to the risk of AMI; relative risks for eating tofu <2, 2-3, and 4+ times per week were 1.0, 0.8, and 0.5, respectively, after adjustment for non-dietary factors (p for trend = 0.01). Further adjustment for consumption of fruit, fish and tofu did not alter the findings generally. The findings suggest that, in women at least, tofu consumption may be protective against the risk of AMI. Further studies are needed to corroborate the relationship of consumption of fish and fruit to AMI risk in Japanese men and women. PMID- 11266196 TI - Continuous observation of superoxide generation in an in-situ ischemia reperfusion rat lung model. AB - To investigate the time course of superoxide generation in ischemia-reperfusion in the in-vivo rat lung, the present study used an enhanced chemiluminescence method with 2-methyl-6-[p-methoxyphenyl]-3,7-dihydroimidazo[1, 2-alpha]pyrazin-3 one (MCLA) as a specific probe. The right pulmonary artery was occluded for 120 min, followed by 90-min reperfusion. Chemiluminescence induced by MCLA was continuously monitored by a photomultiplier exposed to the right lung. Chemiluminescence increased gradually in 30 min of reperfusion and remained elevated throughout reperfusion. The ratio of the luminescence count during reperfusion to the preischemic value increased to 2.20+/-0.31 (mean+/-SEM) (p<0.02 vs preischemic level), 2.50+/-0.39 (p<0.005), and 2.69+/-0.44 (p<0.005), at 30, 60, and 90 min of reperfusion, respectively. Bolus administration of superoxide dismutase during the reperfusion period significantly attenuated the chemiluminescence by 45.0+/-6.7% (p<0.01). The present results suggest that increasing oxygen radical formation leading to ischemia-reperfusion lung injury may occur even after a short period of occlusion of the pulmonary artery alone in vivo. PMID- 11266197 TI - Postischemic reperfusion injury can be attenuated by oxygen tension control. AB - Oxygen-derived free radicals cause cytotoxic damage during reperfusion after a period of ischemia and the production of these free radicals may be proportionate to oxygen tension (PO2). The present study tested the hypothesis that oxidative damage may be limited by maintaining a more physiologic PO2 following ischemia. An experimental study in Wistar rats were mounted on a Langendorff apparatus was conducted to estimate baseline aortic flow (AF), coronary flow (CF), cardiac output (CO), systolic pressure (SP), heart rate (HR), and the rate-pressure product (RPP: HRxSP). The hearts were divided into 3 groups (n=7, hearts/group): group 1, hypoxic (PO2=300+/-50 mmHg) reperfusion; group 2, middleoxic (PO2=500+/ 50 mmHg) reperfusion; and group 3, hyperoxic (PO2=700+/-50 mmHg) reperfusion. Following 30 min of warm ischemia, hearts in all groups were reperfused at each oxygen pressure. The recovery of cardiac function of each heart was measured at the end of reperfusion. Concentrations of lactate (LAC), lactate dehydrogenase (LDH), and creatine kinase (CK) in the coronary perfusate during reperfusion were measured. The recovery rate of CO, SP, and RPP in group 2 were all significantly better than in the other 2 groups. CK leakage in group 2 was significantly lower than in group 3. A clinical study was also conducted during elective coronary artery bypass grafts in 16 consecutive patients who underwent either hyperoxic (n=8, PO2=450-550 mmHg) or more physiologic (n=8, PO2=200-250 mmHg) cardiopulmonary bypass after aortic unclamping. The clinical study assessed CK MB, LDH, LAC, and malondialdehyde (MDA) in patient blood prior to starting the surgical procedure and at 30 min and 3, 9, and 21 h after unclamping. Cardiac index (CI), central venous pressure, pulmonary capillary wedge pressure, systolic arterial pressure, and the dose of cathecholamines were also measured. Although no significant differences were present in the dose of cathecholamines, the CI in the more physiologic oxygen tension group was significantly higher than in the hyperoxic group at 3 and 6 h after unclamping. The levels of MDA in the more physiologic PO2 group was significantly lower at 30 min after aortic unclamping than in the hyperoxic group. The present results suggest that in the experimental as well as in the clinical study, high PO2 leads to myocardial reperfusion damage; however, maintaining a more physiologic PO2 during reperfusion following ischemia may attenuate reperfusion injury. PMID- 11266198 TI - Leakage of energy to the body surface during defibrillation shock by an implantable cardioverter-defibrillator (ICD) system--experimental evaluation during defibrillation shocks through the right ventricular lead and the subcutaneous active-can in canines. AB - The leakage of electrical current to the body surface during defibrillation shock delivery by an implantable cardioverter-defibrillator (ICD) device (the Medtronic Jewel Plus PCD system) was evaluated in 5 dogs. The defibrillation shocks were delivered between the active-can implanted in the left subclavicular region and the endocardial lead placed in the right ventricle at the energy levels of 1, 2, 8, 12, 24 and 34 J. During each delivery, the electrical current leakage from the body surface was measured by electrodes connected to a circuit at 4 recording positions: (A) parallel-subcutaneous (the electrodes were fixed in the subcutaneous tissue of the left shoulder and the right lower chest, and the direction of the electrode vector was parallel to the direction of the defibrillation energy flow); (B) cross-subcutaneous (the electrodes were fixed in the subcutaneous tissue of the right shoulder and the left lower chest, and the vector of the electrodes was roughly perpendicular to the direction of the energy flow); (C) parallel-surface (the electrodes were fixed with ECG paste on the shaved skin surface at the left shoulder and the right lower chest); and (D) surface grounded (the electrodes were fixed on the shaved skin surface at the left shoulder and the left foot, which was grounded). The circuit resistance was set at a variable level (100-5,000 ohms) in accordance with the resistance measured through each canine body. Leakage energies were measured in 750 defibrillation shocks with each circuit resistance in 5 dogs. The leakage energy increased in accordance with the increase of the delivered energy and the decrease of the circuit resistance in all 4 recording positions. When the circuit resistance was set at 1,000 ohms, the leakage energy during shock delivery at 34 J was 32+/-17 mJ at position A, 5+/-9 mJ at B, 10+/-9 mJ at C, and 4+/-3 mJ at D (p=0.042). The peak current was highest at position A and was 87+/-22 mA with a circuit resistance of 1,000 ohms. The power of the leakage energy depended on the delivered energy and the impedance between the electrodes. The angle between the alignment of the recording electrodes and the direction of the energy flow was another important factor in determining the leakage energy. Although the peak current of the leakage energy reached the level of macro shock, the highest leakage energy from the body surface was considerably less because of the short duration of the shock delivery. PMID- 11266199 TI - Effects of in vivo gene transfer of fibroblast growth factor-2 on cardiac function and collateral vessel formation in the microembolized rabbit heart. AB - The effects of gene transfer of the secreted form of fibroblast growth factor-2 (FGF-2) were tested using an adenovirus vector in the microembolized rabbit heart. Japanese white rabbits underwent an intracoronary injection of 25-microm microspheres followed by recombinant adenovirus vectors encoding a secreted form of FGF-2 (FGF group), LacZ (LacZ group), or saline (saline group). Left ventricular (LV) systolic function was serially assessed by echocardiography. Vascular density was measured at 14 days with Azan and CD31 staining. The development of collateral vessels was assessed by measuring myocardial blood flow before and after the occlusion of the left anterior descending coronary artery. Percent fractional shortening (%FS) decreased after the microembolization, and improved gradually for 14 days in the FGF group only (41+/-1% (FGF) vs 32+/-1% (LacZ), 31+/-1% (saline), p<0.01). The vascular density and myocardial collateral blood flow were significantly higher in the FGF group in comparison with other groups. Transcoronary arterial gene transfer of the secreted form of FGF-2 was beneficial for the recovery of LV systolic function and development of collateral vessels in the microembolized rabbit heart. PMID- 11266200 TI - Usefulness of intracoronary angioscopy for elucidating the cause of subacute thrombosis after stenting. AB - Stent thrombosis is rare with anti-platelet therapy, which consists of aspirin and ticlopidine as a post-stenting administration. A 77-year-old man had repeated stent thrombosis, which was not predicted by coronary angiography, despite using contemporary periprocedural anti-platelet therapy. Only intravascular fiberscopy was able to detect the cause of the stent thrombosis. PMID- 11266201 TI - Torsade de pointes induced by intravenous and long-term oral amiodarone therapy in a patient with dilated cardiomyopathy. AB - A 70-year-old woman with dilated cardiomyopathy and ventricular tachyarrhythmia was initially treated in 1990 with intravenous amiodarone (240 mg). She developed a junctional escape rhythm (48 beats/min) with QT prolongation (QT: 0.68 s) and 8 h later developed torsade de pointes (TdP). Because other antiarrhythmic drugs did not suppress the arrhythmia, oral amiodarone (100 mg/day) was started in 1995, 7 weeks before she presented with congestive heart failure. The QT prolongation (QTc: 0.64) increased after administration of dopamine, and TdP again developed. This case suggests that amiodarone induces proarrhythmias by different mechanisms when administered intravenously or orally. PMID- 11266202 TI - Huge calcified aneurysm of the sinus of Valsalva. AB - Aneurysms of the sinus of Valsalva often remain undiagnosed until they rupture. A 61-year-old man had a huge, heavily calcified unruptured aneurysm, originating from the right sinus of Valsalva, detected incidentally on a chest radiograph taken for the diagnosis of cardiomegaly. Two-dimensional echocardiography revealed pericardial effusion with a huge calcified mass compressing the right ventricular outflow tract. The Doppler color-flow echocardiogram showed blood flow from the aortic root into the aneurysm. A chest computed tomographic scan revealed a large thrombosed aneurysm originating from the aortic root and measuring 10x10 cm. After pericardiocentesis, cardiac catheterization was performed, which showed that the right ventricular systolic pressure had elevated to 80 mmHg. Aortic root aortography demonstrated a huge unruptured calcified aneurysm in the sinus of Valsalva arising from the right coronary sinus. The patient underwent surgical correction to prevent aneurysmal rupture and to relieve the right ventricular outflow obstruction. PMID- 11266203 TI - Atrial pacing during radiofrequency ablation of junctional ectopic tachycardia--a useful technique for avoiding atrioventricular bloc. AB - Radiofrequency catheter ablation (RFCA) was performed on a 5-year-old boy with congenital junctional ectopic tachycardia (JET) that was refractory to medical management. Because of the lack of retrograde atrial depolarization during tachycardia, radiofrequency energy was delivered during atrial overdrive pacing to confirm the presence of preserved atrioventricular (AV) conduction. Although the procedure was complicated by complete right bundle branch block after ablation of the para-Hissian region, the patient regained sinus rhythm accompanied by normal AV conduction. Rapid atrial pacing during RFCA of JET may be safely used to avoid AV block. PMID- 11266204 TI - Rubella virus glycoprotein interaction with the endoplasmic reticulum calreticulin and calnexin. AB - Very little is known about the cellular factors that are required for the maturation of rubella virus glycoproteins (E2 and E1) in the endoplasmic reticulum of the infected cell. In the present study, we established the interaction of the ER chaperone proteins, calreticulin and calnexin, with the RV E1 and E2 proteins in cells stably expressing the viral proteins. The interaction between E2 and calnexin was significantly higher than with calreticulin. In pulse chase experiments, the half-life of the E2-calnexin was >45 min, whereas the half life of the calreticulin-E2 interaction was approximately 10 min. Tunicamycin and castanospermine treatments altered the mobilities of intracellular E1 and E2, due to either lack of oligosaccharide ligand addition or trimming of terminal glucose residues, respectively. Further, the drug treatments resulted in a loss of E1 and E2 interaction with calreticulin or calnexin, thereby demonstrating that the interaction is through monoglucosylated forms of RV proteins. These studies suggest that the interaction of RV glycoproteins with the ER chaperone proteins is essential for their maturation in the endoplasmic reticulum. PMID- 11266205 TI - Egg yolk antibodies against the E7 oncogenic protein of human papillomavirus type 16. AB - The E7 oncoprotein is the major transforming protein of Human Papillomavirus type 16 (HPV16) and the most abundant in cervical neoplasia. In this study we report the production of polyclonal antibodies to HPV16 E7 in rabbits and hens. The produced antibodies recognised the denatured and native form of HPV16 E7 protein by Western Blot, and immunoprecipitation. Epitope mapping demonstrated that hen antibodies reacted with a greater number of antigen determinants than the rabbit antibodies. In immunocytochemistry only hen antibodies were able to localize the E7 protein in a HPV positive cell line and in the high-grade squamous intraepithelial lesions, suggesting their possible usefulness in the screening of clinical samples. PMID- 11266206 TI - Complete nucleotide sequence of RNA-2 of Olive latent ringspot virus. AB - The complete nucleotide sequence of Olive latent ringspot virus (OLRSV) RNA-2 was determined. This RNA is 3969 nucleotides in length and contains a single open reading frame of 3448 nt, that encodes a polypeptide of 1146 amino acids, with a calculated Mr of 126,044. OLRSV RNA-2 has a structural organization typical of nepoviruses, with the coat protein (CP) cistron located in the C-terminal and the putative movement protein (MP) in the N-terminal regions of the polyprotein. Computer-assisted comparison of coat proteins of OLRSV and other nepoviruses disclosed relationships that tally with subgrouping based on physicochemical properties. PMID- 11266207 TI - Complete sequences and phylogenetic analyses of a Singapore isolate of broad bean wilt fabavirus. AB - The complete nucleotide (nt) sequence of a Singapore isolate of broad bean wilt fabavirus from Megakepasma erythrochlamys L., designated BBWV-ME, was determined. Its bipartite genome consisted of two positive-sense single-stranded ribonucleic acids (RNA). RNA1 (5951 nt in length) encoded a putative protease cofactor, nucleotide triphosphate (NTP)-binding domain (helicase), viral genome-linked protein (VPg), protease and RNA-dependent RNA polymerase (RdRp). RNA2 (3607 nt in length) encoded a putative movement protein (MP) and coat proteins (CP). Genome organization of BBWV-ME was similar to other viruses in the Comoviridae family. Phylogenetic analyses showed that fabaviruses were more closely related to the comoviruses than the nepoviruses. PMID- 11266208 TI - Hemagglutinin genotype profiles of canine distemper virus from domestic dogs in Japan. AB - Two distinct hemagglutinin genotypes were identified in canine distemper viruses circulating among dogs since 1991 in Japan. One is the native and numerically predominant KDK-1 genotype, while the other is the 98-002 genotype that may have evolved in the Far East beyond the border between Japan and the Republic of Korea. PMID- 11266209 TI - Nucleotide sequence and infectious cDNA clone of the L1 isolate of Pea seed-borne mosaic potyvirus. AB - The complete nucleotide sequence of Pea seed-borne mosaic potyvirus isolate L1 has been determined from cloned virus cDNA. The PSbMV L1 genome is 9895 nucleotides in length excluding the poly(A) tail. Computer analysis of the sequence revealed a single long open reading frame (ORF) of 9594 nucleotides. The ORF potentially encodes a polyprotein of 3198 amino acids with a deduced Mr of 363537. Nine putative proteolytic cleavage sites were identified by analogy to consensus sequences and genome arrangement in other potyviruses. Two full-length cDNA clones, p35S-L1-4 and p35S-L1-5, were assembled under control of an enhanced 35S promoter and nopaline synthase terminator. Clone p35S-L1-4 was constructed with four introns and p35S-L1-5 with five introns inserted in the cDNA. Clone p35S-L1-4 was unstable in Escherichia coli often resulting in amplification of plasmids with deletions. Clone p35S-L1-5 was stable and apparently less toxic to Escherichia coli resulting in larger bacterial colonies and higher plasmid yield. Both clones were infectious upon mechanical inoculation of plasmid DNA on susceptible pea cultivars Fjord, Scout, and Brutus. Eight pea genotypes resistant to L1 virus were also resistant to the cDNA derived L1 virus. Both native PSbMV L1 and the cDNA derived virus infected Chenopodium quinoa systemically giving rise to characteristic necrotic lesions on uninoculated leaves. PMID- 11266210 TI - Different effects of a single amino acid substitution on three adjacent epitopes in the gp41 C-terminal tail of a neutralizing antibody escape mutant of human immunodeficiency virus type 1. AB - The envelope protein of human immunodeficiency virus type 1 (HIV-1) comprises the outer gp 120 SU domain and the anchoring gp41 TM domain, and the conventional view is that it has a single transmembrane region with the following C-terminal sequence situated entirely within the virion. However, we have recently proposed that the gp41 C-terminal region comprises three transmembrane regions and an external loop structure. Part of this loop is the peptide 731PRGPDRPEGIEEEGGERDRDRS752 that carries three antibody epitopes, 734PDRPEG739, 740IEEE743, and 746ERDRD750. PDRPEG is not detected in virions but reacts with its cognate MAb (C8) in Western blots, IEEE is a linear and non-neutralizing epitope, and ERDRD is a conformational and neutralizing epitope. Here we show that escape mutants selected with neutralizing ERDRD-specific antibody had a single 732R-->G substitution, 14 residues upstream of the cognate epitope, and no longer bound the selecting antibody. The same amino acid substitution altered epitope PDRPEG in the virion so that it now reacted with MAb C8, but left epitope IEEE unaffected. Introduction of 732R-->G by site-specific mutagenesis into the gp41 of cloned HIV-1 NL4-3 virions allowed them to escape neutralization by ERDRD specific IgG, and confirms that 732R makes a major contribution to the neutralizing conformation of the 731-752 region of the C-terminal tail of gp41. PMID- 11266211 TI - Identification of rice black-streaked dwarf fijivirus in maize with rough dwarf disease in China. AB - Three virus isolates from maize with rough dwarf in different provinces in China were analyzed at the molecular level. When compared to an isolate from diseased rice plants in Hubei Province, all four isolates had identical genomic RNA electrophoretic profiles, which were composed of ten double-stranded (ds) RNAs. Full-length cDNAs of segment 10 (S10) from each of the four isolates were cloned by RT-PCR and the complete sequences were determined. Analysis of the sequences revealed that each consisted of 1801 nucleotides and contained a single open reading frame (ORF) which potentially encoded a protein with 558 amino acids. Further, the sequences showed more than 97.0% and 98.0% identity at nucleotide and amino acid levels, respectively. In addition, their identities to rice black streaked dwarf virus S10 were significantly higher than those to maize rough dwarf virus S10. Based on these results, it is suggested that the virus which causes this maize disease in China is rice black-streaked dwarf virus. PMID- 11266212 TI - MMTV-like env gene sequences in human breast cancer. AB - We have previously detected an MMTV env gene-like 660 bp sequence in 38% of human breast cancers, but not in normal tissues or other tumors. In this communication we report the sequences from eleven tumors and three breast cancer cell lines, and compare them to four strains of MMTV and to the known endogenous retroviral sequences. The breast cancer sequences were highly homogenous to the MMTV's, but not to the endogenous sequences suggesting an exogenous origin. PMID- 11266213 TI - Nucleotide sequence of genome segment 5 from Bombyx mori cypovirus 1. AB - The complete nucleotide sequences of the double-stranded RNA genome segments 5 (S5) from Bombyx mori cypovirus 1 (BmCPV-1) strains I and H were determined. The segments consisted of 2,852 nucleotides encoding putative proteins of 881 amino acids with molecular masses of approximately 101 kDa (p101). A homology search showed that p101 has high similarity (93%) to foot-and-mouth disease virus (FMDV) 2A protease (2Apro) at amino acid position 219 to 235. These findings suggest the possibility that p101 encoded by BmCPV-1 S5 might be cleaved into two non structural proteins by post-translational autocleavage involving a 2Apro-like protease. PMID- 11266214 TI - The 7th ICTV report. PMID- 11266215 TI - Polymorphism of the 5' terminal region of Citrus tristeza virus (CTV) RNA: incidence of three sequence types in isolates of different origin and pathogenicity. AB - Sequences of the 5' terminal region of the genomic RNA from eight isolates of Citrus tristeza virus (CTV) were previously classified into three types (I, II and III), with intragroup sequence identity higher than 88% and intergroup sequence identity as low as 44%. Sequencing of an additional 58 cDNA clones from 15 virus isolates showed that all sequences could be unequivocally assigned to one of the three types previously established. The relative frequency of each sequence type was assessed in 57 CTV isolates of different geographic origin and pathogenic characteristics by RT-PCR with sets of type-specific primers using CTV dsRNA as template. None of the isolates yielded amplification of the type I or II sequences alone, but in 19 of them type III sequences were the only amplification product detected. Within isolates containing more than one sequence type, eight had type II and III sequences, 11 had type I and III sequences, and 19 had sequences of the three types. Isolates containing only type III sequences caused only mild to moderate symptoms in Mexican lime, an indicator species for most CTV isolates, whereas isolates causing stem pitting in sweet orange an/or grapefruit, generally contained sequences type II. None of the sequence types could be traced to a precise geographic area, as all types were detected in isolates from at least nine of the 12 countries from which samples were taken. PMID- 11266216 TI - Mouse Mx2 protein inhibits hantavirus but not influenza virus replication. AB - The antiviral potential of Mx2 protein remains unknown, because the Mx2 gene in commonly used strains of laboratory mice is nonfunctional. Our previous study showed that functional Mx2 protein in some feral-origin strains was induced upon interferon treatment, was localized in the cytoplasm, and inhibited vesicular stomatitis virus replication. In the present study, we have demonstrated that the embryonic fibroblastic cells from a feral-origin strain (SPR) expressed 74 kDa Mx2 protein, which prevented the accumulation of viral transcripts and proteins of hantaviruses when the Mx2 gene was constitutively expressed in transfected Vero cells. Furthermore, the cells showed significantly lower titers of the virus than control cells. In contrast, influenza virus replication was not affected by the expression of Mx2 protein in the Vero cells. PMID- 11266217 TI - Characterization of a Paramyxovirus from a Fer de Lance viper (Bothrops jararaca): partial nucleotide sequence of the putative fusion protein. AB - During the generation of Expressed Sequence Tags (ESTs) from the Fer de Lance viper (Bothrops jararaca) venom glands, a partial cDNA (clone H8) coding for a protein with all the features of a paramyxovirus fusion protein was characterized. It has 920 bp and codes for a partial protein of 279 amino acids. Two potential N-glycosylation sites are present in the sequence which also possesses a typical membrane anchoring domain made of a stretch of hydrophobic amino acids. The polyadenylation signal sequence was identified. When compared to other fusion proteins, it showed the highest sequence similarity (37-39%) with those of human parainfluenza 3 and Sendai virus. PMID- 11266218 TI - Characterisation of the RNA-dependent RNA polymerase from Rabbit hemorrhagic disease virus produced in Escherichia coli. AB - All positive-strand RNA viruses encode a RNA-dependent RNA polymerase which in most cases has been only identified on the basis of its sequence conservation. Catalytic activity has been experimentally demonstrated in only a handful of these viral proteins, including that from Rabbit hemorrhagic disease virus. Studies from our laboratory have reported that RHDV RNA polymerase produced in Escherichia coli was enzymatically active showing poly(A)-dependent poly(U) polymerase as well as RNA polymerase activity on heteropolymeric substrates. In this work, we have investigated the in vitro activity of the recombinant 3Dpol from RHDV, including ion requirements, resistance to inhibitors, substrate specificity as well as data on the initiation mechanism of the template-linked products derived from heteropolymeric RNA substrates. Our study demonstrates that in an in vitro reaction recombinant RHDV RNA polymerase generated the minus strand of the heteropolymeric RNA substrates by a "copy-back" mechanism that initiated at the template 3'-terminal OH. PMID- 11266219 TI - Additional rep-encoding DNAs associated with banana bunchy top virus. AB - Banana bunchy top nanovirus (BBTV) has a multicomponent circular single-stranded DNA (cssDNA) genome consisting of at least six components. We have cloned, sequenced and analysed two additional cssDNA components, designated BBTV-S1 and S2, associated with a Taiwanese BBTV isolate. The sequences of BBTV-S1 and S2 comprised 1109 and 1095 nucleotides (nt), respectively, and like BBTV DNA-1, potentially encoded replication initiation proteins (Reps). However, the genome organisation of BBTV-S1 and S2 differed from that of BBTV DNA-1 in that (i) the stem sequence of the CR-SL was not conserved, (ii) the internal gene was absent and (iii) the probable TATA boxes were located 5' of the stem-loop. Further, sequence and phylogenetic analysis of the Rep genes indicated that BBTV DNA-S1 and S2 were distinct from BBTV DNA-1. When different geographical isolates of BBTV were tested for the presence of BBTV-S1/S2, these components were detected in various isolates from Vietnam, Taiwan, the Philippines, Tonga and Samoa but were not detected in isolates from Australia, Egypt, Fiji, and India. Based on these results, BBTV-S1 and S2 do not appear to be integral components of the BBTV genome and represent additional Rep-encoding DNAs associated with BBTV. PMID- 11266220 TI - Detection and subtyping (H5 and H7) of avian type A influenza virus by reverse transcription-PCR and PCR-ELISA. AB - Avian influenza virus infections are a major cause of morbidity and rapid identification of the virus has important clinical, economical and epidemiological implications. We have developed a one-tube Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) for the rapid diagnosis of avian influenza A. A panel of reference influenza strains from various hosts including avian species, human, swine and horse were evaluated in a one tube RT-PCR using primers designed for the amplification of a 218 bp fragment of the NP gene. The PCR products were detected by PCR-ELISA by use of an internal catching probe confirming the NP influenza A origin. The PCR-ELISA was about 100 times more sensitive than detection of PCR products by agarose gel electrophoresis. RT-PCR and detection by PCR-ELISA is comparable in sensitivity to virus propagation in eggs. We also designed primers for the detection of the influenza. A subtypes H5 and H7 shown to have pathogenic potential in poultry. The H5 primers cover the cleavage site of the HA gene and specifically amplify influenza A subtype H5. The H7 primers also cover the HA cleavage site and detected all H7 reference strains investigated. In addition, the H7 primers also amplified very weak and/or additional bands on an agarose gel from other subtypes. However, the H7 origin and the pathogenic potential defined by the presence or absence of basic amino acids at the cleavage site can be determined by sequencing of the PCR product. As far as we know this is the first demonstration of RT-PCR detection on a panel of H7 strains using only one primer set. PMID- 11266222 TI - Optimizing drug delivery to the lung: design of a CFC-free corticosteroid metered dose aerosol system. AB - The mandatory replacement of chlorofluorocarbons (CFCs) with ozone-friendly propellants, such as hydrofluoroalkanes (HFAs), has provided an opportunity to optimize aerosol design and improve drug delivery to pulmonary tissue. Asthma is an inflammatory disorder of the lungs that appears to affect both small and large airways, so ideally, inhaled corticosteroid should reach both central and peripheral sites. This review considers the development of an aerosol system containing beclomethasone dipropionate in hydrofluoroalkane-134a (HFA) propellant (Qvar, 3M Health Care, Loughborough, UK) designed to target medication delivery throughout the bronchopulmonary tree and to improve the therapeutic ratio (topical efficacy: systemic safety), thereby offering potential clinical benefits to asthma patients. PMID- 11266221 TI - Bovine viral diarrhoea virus genotype 1 can be separated into at least eleven genetic groups. AB - Seventy-eight bovine viral diarrhoea viruses (BVDV) recently collected in Austria, France, Hungary, Italy, Slovakia, Spain and UK were genetically typed in the 5'-untranslated (5'UTR) and autoprotease (Npro) regions of the pestivirus genome. Seventy-six of the isolates were BVDV-1 and two French isolates were of the BVDV-2 genotype. Phylogenetic analysis of the 5'UTR (245 nt), including additional BVDV-1 sequences from USA, Canada, Germany, New Zealand, Mozambique and Sweden, taken from GenBank and from our previous works, indicated that these viruses were clustered not only into the two generally accepted groups (BVDV-1a "NADL like" and BVDV-1b-"Osloss like"), but altogether into 11 phylogenetic groups. Similar clustering was observed with Npro region sequences (385 nt) and the highest bootstrap values (over 95%) were obtained by phylogeny combining 5'UTR and Npro sequences. Some associations between the genetic grouping and the origin of the isolates were apparent, probably reflecting historical trade contacts. Considering the variability of isolates it is recommended that diagnostic PCR primers should be re-examined to ensure coverage of all BVDV-1 groups. The genogroups were less clearly differentiated by monoclonal antibody typing, suggesting significant antigenic similarities within the BVDV-1 genotype. PMID- 11266223 TI - Effects of grinding with microcrystalline cellulose and cyclodextrins on the ketoprofen physicochemical properties. AB - Ground mixtures of ketoprofen (KETO) with native crystalline beta-cyclodextrin, amorphous statistically substituted methyl-beta-cyclodextrin, and microcrystalline cellulose were investigated for both solid phase characterization (differential scanning calorimetry (DSC) powder X-ray diffractometry, and infrared (IR) spectrometry) and dissolution properties (dispersed amount and rotating disk methods) to evaluate the role of the carrier on the performance of the final product. The effects of different grinding conditions, partial sample dehydration, and 1 year storage at room temperature were also investigated. The results pointed out the importance of the carrier nature on the efficiency of the cogrinding process. Both cyclodextrins were much more effective than was microcrystalline cellulose, even though no true inclusion complex formation occurred by mechanochemical activation. The best results were obtained from ground mixtures with methyl-beta-cyclodextrin, which showed the best amorphizing and solubilizing power toward the drug and permitted an increase of approximately 100 times its intrinsic dissolution rate constant, in comparison with the approximate 10 times increase obtained from ground mixtures with beta cyclodextrin. PMID- 11266224 TI - Oral sustained-release bioadhesive tablet formulation of didanosine. AB - The objective of this study was to formulate a hydrogel-forming bioadhesive drug delivery system for oral administration of didanosine (ddI). The aim of this tablet dosage form is to improve the oral absorption of ddI by delivering it in small doses over an extended period and localizing it in the intestine by bioadhesion. Compressed tablets of ddI using Polyox WSRN-303, Carbopol 974P-NF, and Methocel K4M as the bioadhesive release rate-controlling polymers were prepared. The effect of polymer concentration on the release profile and in vitro bioadhesion of the matrix tablets was studied. Tablet formulations with Polyox WSRN-303 (10%) and Methocel K4M (30%) showed 93 and 90% drug release, respectively, after 12 h. The drug release was found to be linear when fitted in the Higuchi equation (square-root time equation), suggesting zero-order release. Carbopol 974-P-NF was found to inhibit the complete release of ddI because of drug-polymer interaction; hence, is not suitable for formulation of ddI. Drug diffusion and swelling of the polymer (anomalous Fickian release) was found dominant in ddI release. In general, in vitro bioadhesion increased with an increase in polymer concentration. Tablets containing a single polymer can be designed to form hydrogels serving the dual purpose of bioadhesion and sustained release. PMID- 11266225 TI - Intraocular pressure-lowering activity and in vivo disposition of dipivalyl terbutalone in rabbits. AB - The ocular hypotensive activity and in vivo disposition of dipivalyl terbutalone, a chemical delivery system for terbutaline, were investigated in the albino rabbit model. The dose-response relationship was assessed at drug concentrations ranging from 0.5 to 4% in normal saline. Intraocular pressure (IOP)-lowering activity of dipivalyl terbutalone was compared with epinephrine in equal concentrations. The in vivo disposition was investigated after topical administration of dipivalyl terbutalone at 4% dose level for which two metabolic products, terbutaline, and terbutalone, were monitored in different anterior segment tissues/fluid of albino rabbits, including corneal and iris-ciliary body homogenates and aqueous humor. The instillation of 0.5, 1, 2, and 4% solutions (1 drop of 50 microL) significantly decreased the IOP of normotensive rabbits in a dose-dependant manner. At the highest dose, the maximum reduction (5.6 +/- 0.65 mm Hg) was observed at 3 h. In a comparative efficacy study, dipivalyl terbutalone was found to be more effective than epinephrine. In the in vivo distribution after the topical administration of dipivalyl terbutalone, terbutaline was found only in the iris-ciliary body, whereas terbutalone was found in all parts of the eye studied. This work suggests the potential use of dipivalyl terbutalone as an antiglaucoma agent, representing a new chemical delivery system for terbutaline. PMID- 11266226 TI - Physical properties and molecular behavior of chitosan films. AB - Chitosan films, varying in molecular weight and degree of deacetylation, were prepared by a casting technique using acetic acid as a dissolving vehicle. The physicochemical properties of the films were characterized. Both molecular weight and degree of deaceylation affected the film properties. Powder X-ray diffraction patterns and differential scanning calorimetry thermograms of all chitosan films indicated their amorphous state to partially crystalline state with thermal degradation temperature lower than 280-300 degrees C. The increase in molecular weight of chitosan would increase the tensile strength and elongation as well as moisture absorption of the films, whereas the increase in degree of deacetylation of chitosan would either increase or decrease the tensile strength of the films depending on its molecular weight. Moreover, the higher the degree of deacetylation of chitosan the more brittle and the less moisture absorption the films became. All chitosan films were soluble in HCl-KCl buffer (pH 1.2), normal saline, and distilled water. They swelled in phosphate buffer (pH 7.4), and cross linking between chitosan and phosphate anions might occur Finally, transmission infrared and 13C-NMR spectra supported that chitosan films prepared by using acetic acid as a dissolving were chitosonium acetate films. PMID- 11266227 TI - Influence of pH on the permeability of p-toluidine and aminopyrine through shed snake skin as a model membrane. AB - The influence of pH on the permeability of p-toluidine (pKa, 5.3) and aminopyrine (pKa, 5.0) through shed snake skin as a model membrane was studied. The pH was adjusted to several values, and the solubility of the drugs in each donor was measured. Flux rates and permeability coefficients were calculated from the steady-state penetration portions. The flux rates of p-toluidine decreased as the pH value in the donor solution increased. On the other hand, the flux rates of aminopyrine were constant at any pH value. The permeability coefficients of each drug increased as the pH value in the donor solution increased. The partition coefficients (octanol/buffer) of each drug were dependent on the molecular fraction of un-ionized species. From these results, it is suggested that ionized species of p-toluidine transports through shed snake skin, but the ionized species of aminopyrine does not. PMID- 11266228 TI - Effect of gelling conditions and mechanical treatment on drug availability from a lipogel. AB - This study has been conducted to determine whether the rheological differences depending on gelling and treatment conditions could have an influence on drug availability. Lipogels with constant composition were obtained by gelling olive oil with monodiglycerides at rest, under stirring, and milled after gelling. The considerable differences in rheological characteristics produced significant differences on in vitro drug release tests, whereas a lesser influence was observed on in vitro simulated absorption test. The rheological differences appeared not to influence in vivo drug availability. Also, rheological differences owing to the concentration of the gelling agent showed no significant influence on in vivo availability. PMID- 11266229 TI - Preparation and some physicochemical properties of cross-linked poloxamer hydrogel spheres. AB - The principal purpose of this paper is to report the preparation of cross-linked poloxamer hydrogel spheres in an aqueous two-phase system without the use of organic solvent and additional emulsifier and physicochemical properties related to drug release. Poloxamer 188 was modified with methacryloyl chloride to obtain the polymerizable derivative (macromer). The aqueous solution of the macromer was mixed with dextran/magnesium sulfate aqueous solution to form a water-in-water emulsion system. After polymerizing the macromer in the dispersion phase, nonporous particles with a mean diameter of micron level were prepared. Both the mean diameter and swelling ratio of spheres can be tailored by varying the starting composition of the preparations. The drug release experiments indicate that the release of vitamin B12 entrapped in the spheres follows first-order kinetics. PMID- 11266230 TI - Normal-phase TLC separation of enantiomers of 1.4-dihydropyridine derivatives. AB - A new TLC-based method was proposed for the separation of enantiomers and mixtures of racemic DHP derivatives differing in the kind of substituent in the phenyl ring. The conditions for the effective determination of the substances involved and the mechanism of their sorption were also studied. For the separation of felodipine, nilvadipine, and isradipine enantiomers, thin-layer chromatography was used, with a chiral stationary phase of the ligand exchange type, and developing phases of a different concentration of methanol (phi) as an organic modifier. The retention coefficient values k' were used to make the plots log k' = f(log phi) and log k' = f(phi). The processes taking place in the chromatographic systems were shown to be described by the Snyder-Soczewinski equation. PMID- 11266231 TI - Eosinophils and respiratory disease. PMID- 11266232 TI - RSV disease--is there a role for prevention? PMID- 11266234 TI - Compositional and functional changes of pulmonary surfactant in a guinea-pig model of chronic asthma. AB - Recent studies have found that severe surfactant dysfunction occurs during an asthma attack, but the changes in surfactant in a guinea-pig model of chronic asthma have not been studied. We therefore analysed the surfactant recovered from guinea-pigs after repeated inhalation of ovalbumin to see if the surfactant recovered from chronic asthmatic lungs would be intrinsically altered. Guinea pigs immunized through repeated inhalation of aerosolized ovalbumin (OA) were exposed to the antigen once a week for a month. Twenty-four hours after the last challenge the alveolar wash was recovered. We calculated saturated phosphatidylcholine (Sat-PC) and total protein (TP) pool sizes in alveolar spaces. Surfactant subtype conversion of large aggregate surfactant (LA) to small aggregate surfactant was studied in vitro by means of the surface area cycling technique. The phospholipid composition of LA was analysed by thin layer chromatography and the surface activity of LA was also determined. We found decreased surfactant pool sizes, decreased ratio of Sat-PC to TP in alveolar lavages in asthma groups, and surface activity of the surfactant recovered from asthmatic lungs to be inferior to that of the controls. Accelerated surfactant subtype conversion in vitro was also noted in the lungs of asthmatic animal models. In addition, the changes in phospholipid compositions which were similar to the pattern of acute lung injury suggested that alveolar inflammation might be involved in the pathogenesis of chronic asthma. These results indicate that surfactant is intrinsically abnormal in chronically asthmatic lungs. PMID- 11266233 TI - Reflexology and bronchial asthma. AB - Many asthma patients seek alternative or adjunctive therapies. One such modality is reflexology, whereby finger pressure is applied to certain parts of the body. The aim of the study was to examine the popular claim that reflexology treatment benefits bronchial asthma. Ten weeks of active or simulated (placebo) reflexology given by an experienced reflexologist, were compared in an otherwise blind, controlled trial of 20+20 outpatients with asthma. Objective lung function tests (peak flow morning and evening, and weekly spirometry at the clinic) did not change. Subjective scores (describing symptoms, beta2-inhalations and quality of life) and also bronchial sensitivity to histamine improved on both regimens, but no differences were found between groups receiving active or placebo reflexology. However, a trend in favour of reflexology became significant when a supplementary analysis of symptom diaries was carried out. It was accompanied by a significant pattern compatible with subconscious unblinding, in that patients tended to guess which treatment they had been receiving. No evidence was found that reflexology has a specific effect on asthma beyond placebo influence. PMID- 11266235 TI - Hypercalcaemia in Greek patients with tuberculosis before the initiation of anti tuberculosis treatment. AB - Hypercalcaemia has been known to occur in association with granulomatous diseases. The aim of this study was to ascertain the incidence of hypercalcaemia and determine the prevalence of symptoms associated with it in Greek patients with newly-diagnosed tuberculosis (TB), before the initiation of anti tuberculosis treatment. We prospectively evaluated all patients with newly diagnosed TB presenting, either as inpatients or as outpatients, to our hospital, during a 3-year period. We evaluated 88 patients with TB (50 males and 38 females), aged between 23 and 89 years (mean age+/-SD: 46.4+/-19 years), and 65 age- and sex-matched controls with chronic obstructive pulmonary disease (36 males and 29 females), aged between 28 and 88 years (mean age+/-SD: 47.2+/-18 years). Among TB patients, 56 had pulmonary TB, 20 had pleural TB without evidence of pulmonary parenchyma involvement, eight had pulmonary and pleural TB, and four had disseminated disease. The mean (+/-SD) albumin-adjusted serum calcium concentration and the mean ionized calcium concentration were significantly higher in the TB group (2.49+/-0.21 mmol l(-1) and 1.27+/-0.02 mmol l(-1) respectively) than in the control group (2.36+/-0.11 mmol l(-1) and 1.19+/ 0.02 mmol l(-1), P<0.05). In the TB group no correlation between type of disease and albumin-adjusted or ionized calcium concentration was seen. Hypercalcaemia was detected in 22 patients with TB (25%) but only three showed symptoms associated with it. We conclude that, although hypercalcaemia is a common laboratory finding among Greek patients with TB before anti-TB chemotherapy, it is usually asymtomatic. PMID- 11266236 TI - Respiratory disorders in common variable immunodeficiency. AB - Common variable immunodeficiency (CVID) is a heterogeneous immunodeficiency syndrome characterized by hypogammaglobulinemia, recurrent bacterial infections, and various immunologic abnormalities. The clinical presentation is generally that of recurrent pyogenic sinopulmonary infections. Our objectives were to study the prevalence of lung involvement and the response to intravenous immunoglobulin replacement therapy in 19 patients with CVID. Nineteen patients (12 men) with a mean age (SD) of 33.1 (17.1) years had a previous diagnosis of CVID and were treated with intravenous immunoglobulin replacement. All patients underwent complete pulmonary function tests and high-resolution computed tomography (HRCT) examination. Bronchiectasis was diagnosed in 11 (58%) patients and eight (42%) were multi-lobar bronchiectasis. Chronic airflow limitation (CAL) was present in 10 (53%) patients and a restrictive pattern was seen in one case. Eleven patients (58%) presented a decrease in single-breath carbon monoxide diffusing capacity of the lung (DL(CO)). Before intravenous immunoglobulin replacement therapy (INIRT), 84% of patients had suffered from at least one episode of pneumonia. Episodes of lower respiratory tract infection decreased significantly from 0.28 per patient and year before replacement therapy to 0.16 per patient and year after treatment. The mean duration of replacement therapy was 7.5 years. In conclusion lung involvement was frequent in patients with CVID. Long-term administration of intravenous gammaglobulin resulted in a substantial reduction of pneumonic episodes. PMID- 11266237 TI - Living with severe COPD. A qualitative exploration of the experience of patients in Leeds. AB - A sample of 37 patients with severe chronic obstructive pulmonary disease (COPD) drawn from a larger group of 64 patients being studied in a randomized trial of nebulizer versus high-dose inhaler therapy, and similar in age and gender mix, was interviewed in depth, with a view to illuminating standardized outcome measures and improving the understanding of these patients' needs. Three patients are described in detail. The interview material was analysed using non computerized methods. Quality of life was seen as depending mainly on family relationships, opportunities afforded locally for neighbourliness and freedom from fear, mobility and independence in the activities of daily living, and the absence or successful mitigation of symptoms of concomitant disease. Disease specific, symptom-oriented outcome measures may miss these. Additionally, the near and repeatedly threatening approach of death recommended extension of a palliative approach to endstage COPD. PMID- 11266238 TI - Comparing the adrenaline-Pirquet test with international PPD tuberculin tests. AB - Several tuberculins, strengths and setting methods are in everyday use. We wanted to compare the Norwegian reference adrenaline Pirquet test with the internationally recommended Mantoux PPD test and Rhoditest. In responders of a random sample of young adults, with randomization of test subjects, we intra individually compared the adrenaline-Pirquet (aP) test with Norwegian-produced synthetic medium tuberculin (SMT) with either the Mantoux test with PPD 2 tuberculin units (TU) (M2), the Mantoux-PPD 5 TU (M5) or the PPD 2 TU-Rhoditest (Rh). The criteria for a positive reaction were > or = 4mm for the aP test, > or = 10 mm for the M2 test, > or = 6 mm for the M5 test and > or = 2 mm for the Rh test. Strongly positive reactions were defined as aP test > or = 10 mm and M2/M5 test > or = 15 mm. One of the tuberculin tests was positive while the comparison test was negative in 14% of the M2 test group (n = 236), 15% in the M5 test group (n = 162) and 20% in the Rh test group (n = 187). The three PPD tests had positive reactions 3-10 times as often, with a simultaneous negative aP test, than vice versa. Strongly positive reactions occurred in 7% of the aP tests (> or = 10 mm), 32% of the M2 tests (> or = 15 mm) and in 41% of the M5 tests (> or = 15 mm). Receiver operating characteristic (ROC) curves gave the best agreement, with aP test >3mm compared with the M2 and the Rh tests. Regression equations are presented for transformations of the Norwegian reference method of adrenaline Pirquet results to internationally recommended PPD tests. The international recommendations, globalization in general and the skill of Norwegian public health nurses in performing intra-dermal BCG suggest a future shift to the PPD 2 TU Mantoux test in Norway. Due to the lack of sensitivity and specificity of all tuberculin tests they might be used in targeted tuberculin testing and not as a general screening procedure in a low-incidence tuberculosis area with BCG vaccinated inhabitants. PMID- 11266239 TI - Efficacy of HFA-beclomethasone dipropionate extra-fine aerosol (800 microg day( 1)) versus HFA-fluticasone propionate (1000 microg day(-1)) in patients with asthma. AB - Hydrofluoroalkane-134a beclomethasone dipropionate (HFA-BDP) extra-fine aerosol and HFA-fluticasone propionate (HFA-FP) are chlorofluorocarbon-free inhalers. We conducted an 8-week, open study to demonstrate the equivalence of HFA-BDP (800 microg day(-1)) and HFA-FP (1000 microg day(-1)) in moderate to severe asthma. Symptomatic patients on 500-1000 microg day(-1) CFC-BDP (or equivalent) and short acting beta-agonist, were randomized to HFA-BDP (n = 101) or HFA-FP (n = 97) after 7-14 (+/-2) day run-in. In the intent-to-treat (ITT) population (n = 198), both treatments provided clinically and statistically significant improvements in asthma control, with increases in peak expiratory flow in the morning (AM PEF) and asthma symptoms (within treatment analysis P<0.05). Mean (SE) change in AM PEF from baseline at week 8 was equivalent (defined as 90% CI for the mean difference between treatments within +/-25 l min(-1)) in the two groups: 29.59 (5.19) l min(-1) for HFA-BDP vs. 17.3 (5.45) l min(-1) for HFA-FP (90% CI-0.02, 24.91). For the perprotocol population (n = 121), the mean (SE) change in AM PEF from baseline was not equivalent; AM PEF improved to a significantly greater extent in the HFA-BDP group than HFA-FP group [34.84 (7.08) vs. 20.63 (7.32) l min(-1) P<0.01; 90% CI; 2.66, 31.10]. At week 8 in the ITT population, there were no statistically significant differences in FEV1, beta-agonist use, asthma symptom/sleep disturbance scores, or percentage of days without asthma symptoms/sleep disturbance. There was a significantly greater reduction from baseline in mean eosinophil count for HFA-BDP compared with HFA-FP at weeks 3 and 8 (P<0.01), and eosinophil cationic protein value at week 8 (P<0.01). Both treatments were well tolerated and there were no statistically significant differences in urinary cortisol creatinine parameters. In conclusion, this study showed that, in patients with moderate-to-severe symptomatic asthma, HFA-BDP extra-fine aerosol 800 microg(-1) was at least as effective and equally well tolerated as 1000 microg day(-1) HFA-FP. PMID- 11266240 TI - Expression of integrins in human cultured mesothelial cells: the roles in cell-to extracellular matrix adhesion and inhibition by RGD-containing peptide. AB - Integrins play key roles in cell-to-cell and cell-to-extracellular matrix (ECM) adhesion. We investigated integrin expression on pleural mesothelial cells (PMCs) and the inhibitory effect of arginine-glycine-asparate (RGD)-containing peptide on the adhesion of PMCs to fibronectin and collagen. Using flow cytometry and immunostaining, PMCs freshly isolated from pleural effusions and one mesothelial cell line were screened for different integrins. Intact pleural tissue was evaluated by immunohistochemistry. The adhesion of Met-5A cells to fibronectin and collagen types I, III and IV was assayed with prior treatment of various concentrations of glycine-arginine-glycine aspartate-serine (GRGDS). On primary PMCs, alpha2, alpha3, alpha5, beta1, beta3 and alphavbeta3 were highly expressed (>70%); alpha1 expression was intermediate (30-70%); and alpha4 and alpha6 expressions were low (< 30%). On Met-SA cells, alpha3, alpha5, alpha6 and beta1 were highly expressed (>70%); alpha1 was intermediate (30-70%); and alpha2, alpha4, beta3 and alphavbeta3 were low (<30%). The patterns of immunostaining on pleural tissues were similar to the results of flow cytometry for primary PMCs except for beta3. There was no statistically different expression in various disease states (transudate vs. exudate, benign vs. malignant). The inhibitory effect of GRGDS peptide on Met-5A cell adhesion to all four matrix proteins was dose-dependent. PMID- 11266241 TI - Comparison of asthma costs in patients starting fluticasone propionate compared to patients starting montelukast. AB - An observational study using pharmacy and medical claims was used to determine whether there are differences in asthma care cost between patients that are newly started on montelukast and low-dose fluticasone propionate. Patients were identified who had at least one ICD-9 (493.XX) claim for asthma and were newly prescribed inhaled fluticasone propionate 44 microg (FP) or montelukast 5 or 10 mg (MON). Subjects could not have had a claim for any inhaled corticosteroid or oral leukotriene modifier in 9 months prior to the first prescription claim for either FP or MON. They were subsequently followed for 9 months. Multi-variate regression analysis was used to determine the influence of these single controller therapies on post-index asthma related costs. Positively skewed cost variables were log-transformed prior to their inclusion into the multi-variate model. Asthma-related costs were adjusted for age, gender, health plan, co morbidities, pre-index asthma medication use and pre-index asthma care costs. Multivariate regression analysis, adjusting for baseline covariates, indicated that compared to treatment with montelukast, treatment with FP had significantly (P<0.001) lower post-index total asthma related costs. Adjusted least squares mean total asthma care costs for the 9-month post-index period were $US649 for FP 44 microg compared to $US1028 for montelukast. PMID- 11266242 TI - Prophylactic cranial irradiation in limited disease small-cell lung cancer in complete remission: a retrospective analysis. AB - Recently a meta-analysis showed an improved survival probability of prophylactic cranial irradiation (PCI) in limited disease small-cell lung cancer (LD SCLC) in complete remission after chemotherapy. We evaluated treatment results of PCI+ and PCI- in these patients. Whether PCI (n = 65) or no PCI (n = 37) was administered did not depend either on patients or on tumour characteristics. After 2 years the incidence of brain metastases was 11% in PCI+ patients and 51% in PCI- patients. Both disease-free survival and overall survival were significantly longer after PCI. PCI reduces the incidence of brain metastases, prolongs brain metastases free period, and overall survival in LD SCLC patients in complete remission after chemotherapy. PMID- 11266243 TI - Pitfalls in the diagnosis of complicated pulmonary hydatid disease. AB - The aim of this study was to present the pitfalls in the diagnosis of complicated pulmonary hydatid disease and to discuss the unusual radiological presentations of this endemic disorder in Turkey. We retrospectively evaluated 34 patients (12 females) aged between 8 and 64 years, who were operated on at our centre between 1991 and 1998 and diagnosed with complicated pulmonary hydatid cyst histopathologically. Computerized tomography (CT) scans of these patients were reviewed double-blind by two radiologists. The patients were then divided into two groups: group 1: initial radiological impression is pulmonary hydatid cyst and group 2: initial radiological impression is not pulmonary hydatid disease. These two groups were evaluated in terms of symptoms, radiographical presentation and laboratory tests. The presence of cystic appearance, water-lily sign, ring enhancement concomitant with intact cysts unanimously led the radiologists to the diagnosis of complicated pulmonary hydatid cyst, whereas solid appearance and presence of bronchial obliteration made the diagnosis unlikely. In such circumstances patient history, laboratory findings and bronchoscopic evaluation helped the diagnosis. In conclusion, in endemic regions like Turkey, atypical radiological presentation of complicated pulmonary hydatid disease should be considered in the differential diagnosis of solid pulmonary lesions. PMID- 11266244 TI - Staphylococcus aureus with reduced susceptibility to vancomycin--Illinois, 1999. AB - Staphylococcus aureus is one of the most common causes of hospital- and community acquired infections. Nosocomial methicillin-resistant S. aureus (MRSA) infections have become common, and cases of community-acquired MRSA infections also have occurred. Since 1996, vancomycin-intermediate S. aureus (VISA; vancomycin minimum inhibitory concentration [MIC = 8-16 microg/mL) has been identified in Europe, Asia, and the United States. The emergence of reduced vancomycin susceptibility in S. aureus increases the possibility that some strains will become fully resistant and that available antimicrobial agents will become ineffective for treating infections caused by such strains. This report describes the fourth case of confirmed VISA from a patient in the United States. PMID- 11266245 TI - Laboratory capacity to detect antimicrobial resistance, 1998. AB - Emerging mechanisms of antimicrobial resistance have clinical, microbiologic, and infection-control implications for health-care providers. Antimicrobial resistant organisms include Staphylococcus aureus with reduced susceptibility to vancomycin (minimum inhibitory concentration [MIC] > or = 4 microg/mL), including vancomycin intermediate S. aureus (VISA; vancomycin MIC = 8-16 microg/mL) and Enterobacteriaceae that produce extended spectrum beta-lactamases (ESBLS), which result in resistance to a broad range of beta-lactam antibiotics. Detecting VISA and ESBLs-producing gram-negative pathogens can be difficult for clinical microbiology laboratories. Although CDC and the National Committee for Clinical Laboratory Standards (NCCLS) have published screening and confirmatory methods for these pathogens (Tables 1 and 2), the extent of use of these methods is unknown. This report summarizes results from a survey of microbiology laboratories that participate in the Active Bacterial Core Surveillance (ABCs)/Emerging Infections Program (EIP) Network to assess the capacity of clinical microbiology laboratories to detect VISA and ESBL-producing pathogens; findings indicate that despite adequate capacity for proper testing, many laboratories do not have appropriate methodology to detect these resistant pathogens. PMID- 11266246 TI - Abortion surveillance: preliminary analysis--United States, 1997. AB - For 1997, CDC compiled data about legal induced abortions from the 50 states, New York City, and the District of Columbia. The total number of legal induced abortions was available from all reporting areas; however, not all these areas collected information about the characteristics of women who obtained abortions. This report presents preliminary data for 1997; final 1997 abortion data will be published during summer 2000. PMID- 11266247 TI - The rate of tumour growth does not distinguish between malignant and benign thyroid nodules. AB - OBJECTIVE: To clarify whether or not the rate of tumour growth is a useful measure for distinguishing papillary thyroid cancer from benign nodules. DESIGN: Retrospective study. SETTING: Teaching hospital, Japan. SUBJECTS: Fourteen patients with 27 nodules (<20 mm) followed up for 20 months or more. INTERVENTIONS: High resolution ultrasonography (US). MAIN OUTCOME MEASURE: Growth rate. RESULTS: There were 15 papillary thyroid cancers and 12 benign nodules. The median tumour diameters at the first and the last US examination were 9.0 mm (range 6-19) and 9.0 mm (range 5-21) in papillary thyroid cancers followed up for 37 months (range 21-85), and 11.5 mm (range 4-19) and 11.5 mm (range 5-23) in benign nodules followed up for 38 months (range 20-84), respectively. CONCLUSION: The rate of growth of thyroid nodules as documented by ultrasound did not prove to be useful in distinguishing between malignancy and benign disease of small thyroid nodules (<20 mm). PMID- 11266248 TI - Mesenteric injury after blunt abdominal trauma. AB - OBJECTIVE: To present our experience of mesenteric injuries after blunt abdominal trauma. DESIGN: Retrospective study. SETTING: University hospital, Greece. SUBJECTS: 31 patients with mesenteric injuries out of 333 who required operations for blunt abdominal trauma between March 1978 and March 1998. 21 were diagnosed within 6 hours (median 160 min, early group) and in 10 the diagnosis was delayed (median 21 hours, range 15 hours-7 days, delayed group). INTERVENTIONS: Emergency laparotomy. MAIN OUTCOME MEASURES: Mortality, morbidity, and hospital stay. RESULTS: There were no deaths. The diagnosis was confirmed by diagnostic peritoneal lavage in 17/21 patients in the early group whereas 7/10 in the delayed group were diagnosed by clinical examination alone. Most of the injuries (n = 23) were caused by road traffic accidents. 30 patients had injured the small bowel mesentery and 4 the large bowel mesentery. 25 of the 31 patients had associated injuries. There were no complications in the early group, compared with 6 wound infections and 1 case of small bowel obstruction in the delayed group (p < 0.0001). Median hospital stay in the early group was 11 days (range 3 24) compared with 23 days (range 10-61) in the delayed group (p = 0.004). CONCLUSION: Because delay in diagnosis is significantly associated with morbidity and duration of hospital stay we recommend that all patients admitted with blunt abdominal trauma should have a diagnostic peritoneal lavage as soon as possible PMID- 11266249 TI - Impact of fundus rotation gastroplasty on anastomotic complications after cervical and thoracic oesophagogastrostomies: a prospective non-randomised study. AB - OBJECTIVE: To find out the leak rate after cervical or thoracic anastomoses of oesophagus to fundus rotation gastric tubes after oesophagectomy. DESIGN: Prospective non-randomised study. SETTING: University hospital. Switzerland. SUBJECTS: 95 patients, of whom 62 had cervical and 33 thoracic anastomoses. INTERVENTIONS: Anastomoses were hand sewn in two layers between oesophagus and a gastric tube, that was elongated by 30% by a stapled fundus rotation gastroplasty. Anastomotic patency was studied clinically and radiographically between the 5th and 7th postoperative days. RESULTS: Five of the 62 patients had a clinical or radiological anastomotic leak (8%) in the neck and 2 of the 33 patients in the thorax (6%). Six patients died, one death being the result of a leak. CONCLUSION: Length and blood supply of fundus rotation gastroplasty tubes allows for safe anastomoses at thoracic and cervical levels. PMID- 11266250 TI - Rare indications for a Kausch-Whipple procedure. AB - OBJECTIVES: To find out whether the Kausch-Whipple operation is adequate for the cure of rare tumours of the pancreatic head. DESIGN: Retrospective study. SETTING: University hospital, Germany. PATIENTS: Of 640 patients who had Kausch Whipple procedures between 1972 and 1998 we found 42 (6.6%) who were operated on for rare tumours of the pancreatic head. RESULTS: Among these 42 patients 12 had functioning and non-functioning endocrine tumours, 11 had adenomas that were not locally resectable, 6 had leiomyosarcomas or oncocytomas, 4 had cystadenocarcinomas, 3 had acinar cell carcinomas, 2 had primary lymphomas, and 3 had metastases to the pancreatic head. Operative treatment (such as extended resection), postoperative course, and survival time after operation varied. Patients with adenomas had the most favourable mean survival time of 106.5 months. Among patients with cancer, those with endocrine malignancies had the best outcome with a mean survival duration of 58.3 months. PMID- 11266251 TI - Contrast radiography in small intestinal obstruction, a valuable diagnostic tool? AB - OBJECTIVE: To investigate the diagnostic and therapeutic potential of plain abdominal radiographs and contrast radiography in patients with suspected small intestinal obstruction. DESIGN: Retrospective study. SETTING: General hospital, Sweden. MATERIAL: 2357 sets of plain abdominal radiographic casenotes. MAIN OUTCOME MEASURES: Analysis of plain abdominal radiographs for small intestinal obstruction. Establishment of the time that subsequent contrast radiography medium took to reach the caecum, and its success rate. RESULTS: Of the 2357 plain abdominal films 1599 (68%) did not show small intestinal obstruction, 425 (18%) showed intermediate obstruction, and 333 (14%) showed small intestinal obstruction. The water-soluble contrast medium reached the colon in 394/591 (67%) of the cases with intermediate or complete small intestinal obstruction. Although the contrast medium passed to the colon there was remaining abnormality with dilated small intestine in 71/212 (33%) of the cases with intermediate obstruction and in 95/143 (66%) of the small intestinal obstruction group. The time for the contrast medium to reach the colon was 3.4 hours in the normal group, 5.5 hours in the intermediate group and 8.9 hours in the obstruction group. CONCLUSION: The plain abdominal radiographs seem to predict the success of follow-through examinations. Contrast radiography is safe and may have a therapeutic potential in small intestinal obstruction. PMID- 11266252 TI - Long-term follow-up of 1059 consecutive primary and recurrent inguinal hernias in a teaching hospital. AB - OBJECTIVE: To study the early and late outcome of various methods of inguinal hernia repair. DESIGN: Retrospective study. SETTING: Teaching hospital, Norway. SUBJECTS: 1059 repairs of inguinal hernias in men and women by 43 surgeons. INTERVENTIONS: Analysis of patients charts, results of questionnaires concerning 712 hernias (67%) and follow-up consultations when needed. MAIN OUTCOME MEASURES: Freedom from recurrence and postoperative groin symptoms after repairs of primary and recurrent hernias. RESULTS: After a median follow-up of 5.5 years, range 3-8, the recurrence rate was 8% for primary repairs and 29% after recurrent hernias. The incidence of permanent pain or discomfort was unexpectedly high, being 11% after primary repairs and 15% after recurrent hernia repairs. CONCLUSIONS: The number of recurrences at long-term follow-up after repairs of primary and recurrent inguinal hernias was unsatisfactory. The extent of postoperative pain was surprising as this was not given enough attention during the learning period. We have introduced a uniform treatment policy with a prospective surveillance programme with the aim of improving results in our teaching programme. PMID- 11266253 TI - Collagen tampons as aminoglycoside carriers to reduce postoperative infection rate in prosthetic repair of groin hernias. AB - OBJECTIVE: To find out whether collagen tampons treated with gentamicin would prevent postoperative infections in patients operated on for groin hernias by insertion of prostheses. SETTING: University hospital, Italy. DESIGN: Prospective randomised trial. PATIENTS: 595 patients who required prosthetic repair of a groin hernia. INTERVENTIONS: All repairs were by our standard surgical technique including prophylactic ceftriaxone, local anaesthesia, and insertion of a polypropylene mesh. 301 patients also had a gentamicin laced collagen tampon placed in front of the prosthetic mesh before the aponeurosis of external oblique muscle was sutured. RESULTS: 1/301 patients in the gentamicin group (0.3%) developed a postoperative wound infection compared with 6/294 in the control group (2.0%), (p = 0.04 Fisher exact test). CONCLUSIONS: Gentamicin-laced collagen tampons are effective in reducing the postoperative infection rate in patients operated on for groin hernia by insertion of a prosthesis. PMID- 11266254 TI - Excision and simple primary closure of chronic pilonidal sinus. AB - OBJECTIVE: To evaluate the outcome of asymmetrical complete excision of pilonidal sinus with simple primary closure without using drains or tension sutures. DESIGN: Prospective study. SETTING: Teaching hospital, Jordan. SUBJECTS: 46 patients with chronic pilonidal sinus treated between November 1994 and October 1998 by excision of the sinus down to the sacrococcygeal fascia and simple primary closure of the wound without tension sutures or drains. RESULTS: Patients stayed in hospital for 48 hours and postoperative pain was minimal. Complete healing was achieved in 41 patients (89%) after a mean follow up period of 36 months (range 12-60). Two patients (4%) developed recurrent sinuses and in three (7%) the wound broke down. All the patients who healed completely were back to work within three weeks of the operation. CONCLUSION: Excision and simple primary closure is a cost-effective way of treating chronic pilonidal sinus. PMID- 11266255 TI - Prevention of adhesions by surfactants and cellulose derivatives in mice. AB - OBJECTIVE: To evaluate the efficacy of hydrophobically modified ethyl (hydroxyethyl) cellulose (cellulose), sodium hyaluronate (hyaluronate) and phosphatidylglycerol, in the reduction of adhesion formation. DESIGN: Controlled study. SETTING: Experimental academic unit, Sweden. MATERIAL: NMRI mice. Solutions: (1) cellulose, (2) hyaluronate, (3) phosphatidylglycerol, (4) phosphatidylglycerol and cellulose, and (5) phosphatidylglycerol, cellulose and hyaluronate. INTERVENTIONS: A standard lesion was created in the parietal peritoneum in mice. One of the viscous solutions to be tested, or saline, was given intraperitoneally. MAIN OUTCOME MEASURES: Amount of adhesions found one week postoperatively. RESULTS: Cellulose; phosphatidylglycerol and cellulose; and phosphatidylglycerol, cellulose and hyaluronate all significantly reduced the amount of adhesions (p=0.0002, p=0.002, p < 0.0001), as did the hyaluronate alone (p < 0.05). Phosphatidylglycerol alone did not reduce the amount of adhesions. Combining cellulose with phosphatidylglycerol, or with hyaluronate, did not improve efficacy. CONCLUSION: Cellulose and hyaluronate were effective in reducing the formation of adhesions. Combining cellulose with hyaluronate or phosphatidylglycerol or both did not improve efficacy. PMID- 11266257 TI - A way to facilitate dissection in laparoscopic cholecystectomy. PMID- 11266256 TI - Suture materials and local recurrence in colorectal cancer: an experimental study. AB - OBJECTIVE: To investigate the carcinogenic role of suture materials in an experimental model in rats. DESIGN: Laboratory, experimental study. SETTING: University hospital, Spain. SUBJECTS: 125 Sprague-Dawley rats in 5 groups of 25 each. INTERVENTIONS: 3 Different suture materials (silk, polydioxanone, and titanium staples) were implanted in a region of the colon in 75 rats divided into three groups of 25. Another 25 animals were injured by puncture only in the same region, and a further 25 rats were not operated on (controls). Injections of 1 2,dimethylhydrazine were then given for 26 weeks. MAIN OUTCOME MEASURES: Number of rats in each group who had tumours in the region of interest, and level of infiltration of the tumours in each group. RESULTS: There were no differences between the injured and control rats in the number of tumours in the study zone. All groups in which suture material had been inserted had tumours in this zone. Titanium was more carcinogenic than either silk or polydioxanone. The tumours associated with titanium were the most infiltrating. CONCLUSIONS: We recommend the use of absorbable sutures, with a careful follow-up of patients in whom titanium staples have been used. PMID- 11266258 TI - Retroperitoneal fibrosis of the pancreatic head mimicking carcinoma of the pancreas. PMID- 11266259 TI - Pancreaticobiliary fistula: an unusual complication of necrotising pancreatitis. PMID- 11266260 TI - Exocrine pancreatic insufficiency caused by a somatostatinoma of the minor and major duodenal papilla in a patient with neurofibromatosis. PMID- 11266261 TI - A new technique for tying the surgeon's knot. PMID- 11266262 TI - Pain and convalescence after laparoscopic cholecystectomy. AB - Pain and speed of convalescence are the two main problems after uncomplicated laparoscopic cholecystectomy. We therefore identified interventional and descriptive studies in electronic databases and supplemented them with manual searches. Pain and interventional analgesic studies were analysed with reference to the effects on specific pain components after laparoscopic cholecystectomy. Convalescence and factors associated with early recovery were also analysed. Pain after cholecystectomy is most intense for the first 2-3 days with a high interindividual variability and dominated by incisional pain rather than other pain components. Early pain after cholecystectomy is reduced by minimising residual pneumoperitoneum and by giving incisional local anaesthetics, epidural analgesia, and non-steroidal anti-inflammatory drugs. There are inconclusive data about type and pressure of pneumoperitoneum, the use of intraperitoneal local anaesthetics, and the type of general anaesthesia. Pain and medico-cultural traditions are the main factors responsible for prolonged convalescence after laparoscopic cholecystectomy. To minimise pain and the duration of convalescence, we recommend multi-modal analgesic treatment in combination with short, standardised instructions to resume work and normal activity. PMID- 11266265 TI - Leadership and strategic alignment--getting people on board and engaged. PMID- 11266263 TI - Serum concentration of hepatocyte growth factor predicts perioperative surgical stress in children. AB - OBJECTIVE: To find out whether preoperative serum concentrations of hepatocyte growth factor (HGF) can indicate the general condition of sick children and can predict their postoperative inflammatory response. DESIGN: Non-randomised study. SETTING: University hospital, Japan. SUBJECTS: 41 children who required operation and 41 healthy controls. INTERVENTIONS: Samples of peripheral venous blood were obtained during the operation. MAIN OUTCOME MEASURES: Circulating concentrations of HGF, interleukin-6 (IL-6), and C-reactive protein (CRP); neutrophil counts; and nutritional variables including serum cholinesterase, albumin, and body weight: ideal body weight ratio. RESULTS: The mean serum concentration of HGF in the patients was significantly higher than in the normal controls. Preoperative HGF was related to the preoperative nutritional state, the postoperative IL-6 response, and the development of infective complications. CONCLUSIONS: The serum HGF concentration may be a useful variable for evaluating general condition and predicting perioperative surgical stress in sick children. PMID- 11266266 TI - Finance modeling in the delivery of medical care in tertiary-care hospitals in the Department of Veterans Affairs. AB - BACKGROUND: In the mid-1990s, the Department of Veterans Affairs (DVA) implemented the Veterans Equitable Resource Allocation (VERA), a new financial model developed to attempt to better distribute the approximately $18 billion annual budget among roughly 170 Veterans Administration Medical Centers (VAMCs). VERA is based on a Health Maintenance Organization (HMO) model. VERA provides reimbursement to each of the 22 regional Veterans Integrated Service Networks (VISNs), and subsequent VISN distribution to individual VAMCs is based on an individual medical center's enrollment of unique social security numbers (uniques). In HMO vocabulary these are individual "covered lives." METHODS: Currently available demographic and staffing information regarding the DVA's 23 tertiary hospital systems (Category 7 hospitals) on the KLF database (DVA Austin Data Base) and published information on the DVA website were reviewed. The following was obtained: (1) staffing information-physician and nurse full-time employment equivalent (FTEE) staffing; (2) patient demographics and hospital workload-facility uniques (u), outpatient facility uniques, average daily census (ADC), discharges, and outpatient clinic visits. The following staffing ratios were calculated for both physician and nursing: FTEE/(u/1000), FTEE/(discharges/1000), FTEE/(clinic visits/1000), FTEE/ADC. For all categories the means +/- SD were calculated and correlation coefficients were calculated on pertinent pairings. RESULTS: Although categorized as similar tertiary care facilities, the 23 "Group 7" VA hospitals are anything but equivalent when reviewed using the VERA financing model with respect to physician staffing, nurse staffing, and facility uniques. Using VERA methodology, average physician FTEE and total nursing FTEE staffing/(u/1000) are 3.67 +/- 0.89 and 15.53 +/- 3.77, respectively. Correlation statistics of staffing versus unique SSNs demonstrated correlation coefficients of 0.46 and 0.59 with respect to physician and nurse staffing, respectively. On the other hand, when physician FTEE and nursing FTEE staffing were compared with VAMC workload parameters (total ADC, discharges, and outpatient visits), correlation coefficients were more consistent, ranging from 0.62 to 0.86. CONCLUSIONS: In the VERA model, the reward of a larger annual budget for an individual VAMC or the regional VISN is realized when staffing of VAMCs is minimized, overall provided medical services (especially costly tertiary services) are limited, and the number of covered lives is maximized. A VAMC staffing system that equates medical services delivered in a tertiary VAMC setting based on an HMO model like VERA (where the user population is skewed toward the sicker, older patient) shows decreased correlation when compared with VAMC workload model parameters. PMID- 11266268 TI - An experimental study evaluating the effect of Mitomycin C on the prevention of postoperative intraabdominal adhesions. AB - BACKGROUND: Fibroblast proliferation is one of the well-known mechanisms for postoperative intraabdominal adhesion formation. Inhibition of fibroblast proliferation is an attractive field of investigation in the prevention of adhesions. Mitomycin C (MMC) is a cytotoxic agent that alkylates and crosslinks DNA and also inhibits fibroblast proliferation up to a few weeks. We aimed to determine the effect of MMC on the prevention of adhesions. MATERIALS AND METHODS: Generation of adhesions in rats by brushing a 1-cm(2) area of the cecum and the peritoneum on the right side of the abdominal wall was followed by intraperitoneal administration of saline, 1 mg/kg MMC, and 0.5 mg/kg MMC in saline. After 45 days, formation of adhesions was graded. RESULTS: The average adhesion scores of the control, and MMC (1 mg/kg), MMC (0.5 mg/kg) groups were 3.2 +/- 0.7, 0.8 +/- 0.6, and 0.7 +/- 0.8, respectively. Adhesion scores of the two MMC-treated groups were significantly lower than that of the control group (P < 0.001). There was no difference between the two MMC groups (P > 0.05). No side effect of MMC was observed. CONCLUSION: MMC was found to be very effective in the prevention of postoperative intraabdominal adhesions. PMID- 11266267 TI - Effect of 5-lipoxygenase inhibition on Kupffer cell clearance capacity in obstructive jaundiced rats. AB - BACKGROUND: Obstructive jaundice is a common surgical problem. It may cause hepatic and Kupffer cell dysfunction. Previous studies demonstrated that 5 lipoxygenase inhibition prevents hepatic injury. However, its effect on Kupffer cell clearance capacity has not been determined yet. MATERIALS AND METHODS: Rats were divided into four groups. In group 1 (sham control group), only bile duct dissection was performed. In other groups bile ducts were ligated and divided. In groups 1 and 2 saline, in group 3 ethanol, and in group 4 a 5-lipoxygenase inhibitor AA-861 was given intraperitoneally to the animals. Rats were sacrificed 14 days after the operations. Serum alkaline phosphatase, total bilirubin, and alanine aminotransferase levels were determined. Kupffer cell clearance capacity was measured using an in situ isolated hepatic perfusion technique. Hematoxylin eosin-stained liver samples were evaluated under light microscope for histopathologic scoring. RESULTS: Rats in the sham control group had significantly lower serum ALP and bilirubin values than those in the experimental groups with biliary obstruction. AA-861 administration significantly decreased serum ALT levels and histopathologic scores. There was no significant difference in ALT levels and histopathologic scores between the sham control and AA-861 groups. Kupffer cell clearance capacity was found to be significantly increased in the AA-861 group compared to other experimental groups with obstructive jaundice. CONCLUSIONS: This study shows that leukotriene synthesis inhibition using AA-861 prevents hepatic damage and improves Kupffer cell clearance capacity in obstructive jaundiced rats. This may have significant implications for the management of patients with obstructive jaundice. PMID- 11266269 TI - FAK induction in keratinocytes in an in vitro model of reepithelialization. AB - During reepithelialization, keratinocytes must become activated in order to migrate over the provisional extracellular matrix of the wound. Previously we have shown that focal adhesion kinase (FAK) is induced in activated keratinocytes. The mechanisms responsible for keratinocyte activation are unknown. Here we use an organ culture system to investigate FAK up-regulation and regulation of keratinocyte activation. Normal human skin was cultured on type I collagen. Keratinocytes migrated out of the explant onto the supporting collagen. Immunostaining for FAK showed induction in the migrating epithelium and also in the center of the explant some distance from the cut edge. Cells from the center of the explant expressed FAK and showed the activated phenotype as defined by their ability to spread on collagen. Since FAK is a tyrosine kinase, the tyrosine kinase inhibitors genistein or herbimycin A were added to the explant medium for 24 h. Inhibition of tyrosine kinase activity delayed epithelial migration, but keratinocytes were able to begin migrating after removal of the inhibitors. We conclude that FAK is up-regulated in keratinocytes in this whole skin explant model. Furthermore FAK up-regulation and keratinocyte activation are not confined to the migrating cells but are found in cells some distance from the skin margin. These data suggest that (1) cell migration, contact with wound matrix molecules, loss of cell-cell contact, or loss of basement membrane contact is not necessary for FAK up-regulation or keratinocyte activation; and (2) tyrosine kinase signaling pathways are important for reepithelialization. PMID- 11266270 TI - Antibody neutralization of vascular endothelial growth factor inhibits wound granulation tissue formation. AB - OBJECTIVE: The goal of this work was to test the functional role of vascular endothelial growth factor (VEGF) in promoting the vigorous granulation tissue formation, wound fluid accumulation, and angiogenic responses characteristic of this wound model. BACKGROUND: Formation of vessel-rich granulation tissue is central to wound repair and is thought to be regulated by locally liberated angiogenic factors. Despite the clinical importance of granulation tissue formation in the early stage of wound healing, surprisingly little is known about the molecular identity of signals leading to granulation tissue invasion of a wound space. Methods. A ventral hernia, surgically created in the abdominal wall of 15 swine, was repaired using silicone sheeting and skin closure. An osmotic minipump, inserted in a remote subcutaneous pocket, delivered saline (n = 5), an irrelevant control antibody (n = 5), or neutralizing anti-VEGF antibody (n = 5) into the wound environment. Serial ultrasonography on Days 2, 4, 7, 9, 11, and 14 was used to determine the dimensions of the subcutaneous granulation tissue and wound fluid compartment. VEGF and transforming growth factor beta1 (TGF-beta1) levels in serial wound fluid samples were quantitated by ELISA. On Day 14, animals were sacrificed and the abdominal wall was harvested for histologic, biochemical, and molecular analyses. RESULTS: In animals receiving saline or an irrelevant antibody, a nearly linear 4-fold increase in granulation tissue thickness and 7-fold increase in wound fluid volume were measured over the 14-day study interval. In contrast, in animals receiving anti-VEGF neutralizing antibody, Day 14 granulation tissue thickness and wound fluid volume measurements were essentially unchanged from Day 2 values. Moreover, in the anti-VEGF animals, ultrasonography was unable to resolve the "angiogenic zone" typical of both controls, and correspondingly, wound vessel count and vascular surface area estimates derived from image analysis of histological sections were 3-fold lower in the anti-VEGF animals compared with the saline and antibody controls. Finally, VEGF levels in wound fluid detectable by ELISA analysis were strikingly (10-fold) reduced in anti-VEGF animals on Postsurgery Days 7-14. In contrast, TGF-beta1 levels were unaffected by the anti-VEGF treatment. CONCLUSION: Functional VEGF is a key mediator in wound angiogenesis, fluid accumulation, and granulation tissue formation. PMID- 11266271 TI - Antioxidants reduce oxidative stress in claudicants. AB - BACKGROUND: Low-grade ischemia-reperfusion in claudicants leads to damage of local tissues and remote organs. Since this damage is partly caused by oxygen derived free radicals (ODFR), scavenging these ODFR could reduce the local and remote injury. METHODS: Using a new method by which a free radical reaction product (ortho-APOH) of the exogenous marker antipyrine is measured to quantify the oxidative stress, 16 stable claudicants performed a standard walking test before and after administration of vitamin E (200 mg) and vitamin C (500 mg) daily for 4 weeks. FINDINGS: Ortho-APOH was significantly increased during the reperfusion period (P = 0.026) before administration of the vitamins. After 4 weeks of vitamin supplementation no rise was found in the reperfusion period. Malondialdehyde showed no changes in either group. INTERPRETATION: These findings indicate that administering extra antioxidants to claudicants reduces oxidative stress in these patients. This may also have an effect on the remote ischemia reperfusion damage and reduce cardiovascular morbidity in this group. PMID- 11266272 TI - An experimental study of selective intrahepatic biliary ablation with ethanol. AB - BACKGROUND: Few attempts have been made to investigate the feasibility of selective intrahepatic biliary ablation with absolute ethanol. METHODS: Through a catheter cannulated into the bile duct of the left lateral and median lobes (70% of total liver weight), 0.2 mL of absolute ethanol was injected into rats. RESULTS: The weight of the infused lobes decreased to less than 50% of the entire liver weight 14 days after treatment, while the weight of the noninfused lobes increased to 1.6-fold of the original value. This increase was associated with a markedly elevated Ki-67 labeling index. Both bile flow and bile acid excretion in the noninfused lobes significantly increased to more than twice the original values on Day 14. Histologically, the interlobular bile ducts of the infused lobes were destroyed. Ethanol also soaked through Glisson's capsule and destroyed hepatocytes, which were replaced by fibrous tissue and proliferating bile ductules without necrosis by Day 14. The portal veins and hepatic arteries supplying the infused lobes were structurally well preserved throughout the study period. CONCLUSION: Selective biliary infusion of ethanol can be performed safely without serious complications, achieving lobar ablation with contralateral hypertrophy of the liver. PMID- 11266273 TI - Induction of E-selectin after partial hepatectomy promotes metastases to liver in mice. AB - BACKGROUND: The liver is the most frequent site of tumor metastasis. It has been suggested that partial hepatectomy promotes liver metastasis of malignant disease and that expression of E-selectin, a cell adhesion molecule, plays roles in tumor metastasis. However, no reports are available concerning the expression of E selectin after hepatectomy. METHODS: In the present study, we used BALB/c mice subjected to 30% partial hepatectomy after injection of 1 x 10(4) colon 26 cells to determine the effects of partial hepatectomy on tumor metastasis to liver. E Selectin expression within the liver after partial hepatectomy was evaluated using reverse transcription polymerase chain reaction and Western blotting. In addition, we injected polyclonal antibody to E-selectin into mice in which partial hepatectomy had augmented liver metastasis. RESULTS: Mice subjected to partial hepatectomy had significantly increased numbers of liver metastases (sham operation, 1.5 +/- 2.0, vs partial hepatectomy, 35.5 +/- 19.3; P < 0.001). Expression of E-selectin mRNA within the liver was markedly increased 4 h after partial hepatectomy, but subsequently decreased at 24 h. E-Selectin protein was detected 8 h after hepatectomy, but subsequently decreased at 24 h as measured by Western blotting. Mice subjected to intraperitoneal injection of neutralizing antibody after operation had significantly decreased numbers of liver metastases (phosphate-buffered saline, 20.6 +/- 9.2, P < 0.05, and normal IgG, 18.0 +/- 8.0, P < 0.05, compared with polyclonal antibody to E-selectin, 5.6 +/- 4.8). CONCLUSION: Induction of E-selectin by partial hepatectomy promotes hematogenous liver metastasis. Our findings can be applied to surgical treatment of liver tumor to reduce the recurrence of liver metastasis after hepatectomy by inhibiting E-selectin-mediated adhesion using reagents to E-selectin. PMID- 11266274 TI - Gadolinium chloride prevents mortality in hepatectomized rats given endotoxin. AB - The purpose of this study was to determine if inhibition of Kupffer cells by gadolinium chloride (GdCl(3)) affects the arterial ketone body ratio (AKBR), liver injury, and mortality in hepatectomized rats administered lipopolysaccharide (LPS). Rats treated with or without GdCl(3) received a 70% hepatectomy. Either LPS (5 mg/kg) or vehicle (saline) was administered 48 h after hepatectomy. Further, hepatectomized rats were administered superoxide dismutase (CuZnSOD, 9 x 10(4) U/kg) before and every 3 h after LPS injection up to 9 h to assess involvement of superoxide in liver injury in this model. All hepatectomized rats with saline died within 24 h after LPS administration. In contrast, GdCl(3) prevented this mortality completely. Serum AST levels were about 160 IU/L in hepatectomized rats with vehicle; however, values were increased approximately 25-fold by LPS administration. In contrast, these increases were blunted significantly by about 90% by GdCl(3). Further, GdCl(3) also prevented decreases in AKBR caused by LPS. LPS caused severe liver injury, which was stopped almost completely by GdCl(3). LPS-induced increases in superoxide production by isolated Kupffer cells were stopped by about 90% by GdCl(3). Importantly, SOD administration prevented decreases in AKBR, liver injury, and mortality significantly as well as GdCl(3). These results indicated that GdCl(3) prevented liver injury and mortality caused by LPS most likely by inhibiting superoxide production by Kupffer cells. Thus, inhibition of activation of Kupffer cells could be useful for preventing liver dysfunction in postoperative endotoxemia. PMID- 11266275 TI - A mechanism of interleukin-12 unresponsiveness associated with thermal injury. AB - An unresponsive state for the production of interleukin-12 (IL-12) is commonly observed in animals and patients with severe thermal injuries. In the present study, the participation of corticosteroids, prostaglandin E(2) (PGE(2)), and type 2 cytokines, which appeared in association with thermal injury, on the burn associated IL-12 unresponsiveness was studied. These substances have been described as inhibitors of IL-12 production. Less than 20 pg/ml serum IL-12 was produced in thermally injured mice (TI-mice) after stimulation with lipopolysaccharide (LPS), while 1037 pg/ml IL-12 was detected in sera of unburned mice equally stimulated with LPS. Almost complete restoration of the impaired IL 12 production was witnessed in TI-mice after treatment with soluble IL-4 receptor (50 ng/mouse, 2 h and 2 days after thermal injury). However, IL-12 was not induced by LPS stimulation in TI-mice treated with an inhibitor of PGE(2) (indomethacin, 0.1-5 mg/kg) or an inhibitor of corticosteroid production (ketoconazole, 10 mg/kg). LPS-stimulated IL-12 production was also impaired in normal mice inoculated with burn-associated type 2 T cells. In addition, in the presence of 1 microg/ml LPS, naive macrophages cocultured with burn-associated type 2 T cells did not produce IL-12 in their culture fluids, while IL-12 was produced by LPS-stimulated naive macrophages that were cocultured with naive splenic CD8(+) T cells. These results suggest that the IL-12-unresponsive state demonstrated in TI-mice is associated mainly with type 2 cytokines released from burn-associated type 2 T cells. PMID- 11266276 TI - The effect of nitric oxide inhibition on blood pressure depends on rat strain. AB - BACKGROUND: Nitric oxide is a continuously released endothelium-derived vasodilator and plays an important role in the maintenance of blood pressure (BP). Rat strains appear to differ in their resting BP and their response to the intravenous administration of N(omega)-nitro-l-arginine methyl ester (l-NAME), a nitric oxide synthase inhibitor. The presence of diabetes and hypertension also leads to differences in BP responses to l-NAME. We postulated that the contribution of NO to resting BP varies between rat strains and certain strains may be more sensitive to the effects of NO blockade. METHODS: Blood pressure was continuously measured using a carotid arterial catheter and the responses to l NAME were compared in anesthetized Lewis and Sprague-Dawley rats during a 2-h control period and a 2-h experimental period. l-NAME was given by a 50 mg/kg bolus followed by a 10 mg/kg/h infusion via a mesenteric vein. RESULTS: During the control period, the Lewis animals had lower systolic and diastolic BPs of 103 +/- 1 and 80 +/- 1 mm Hg compared with 127 +/- 1 and 105 +/- 1 mm Hg measured in Sprague-Dawley rats (P < 0.01). Although l-NAME infusion increased systolic BP in both strains compared with control values (P < 0.00005), the magnitude was significantly greater in Sprague-Dawley than Lewis animals (P = 0.0142); additionally, the BP was unstable in the Lewis animals. Furthermore, pulse pressure decreased during l-NAME in Lewis animals but increased in Sprague-Dawley animals (P < 0.00005). There were no significant changes in serum concentrations of aspartate transaminase nor of nitrite plus nitrate after l-NAME in either group. CONCLUSION: These results indicate that the effect of l-NAME on systemic BP differs markedly in Sprague-Dawley and Lewis rats, suggesting that the role of nitric oxide in regulation of resting vascular resistance may differ significantly between these rat strains. Rat strain is an important consideration for valid comparisons between studies. PMID- 11266277 TI - Role of endothelin in the circulatory changes associated with small bowel strangulation obstruction in pigs: effects of the endothelin receptor antagonist bosentan. AB - BACKGROUND: We have previously shown that experimental strangulation obstruction leads to increased release and concentration of endothelin-1 (ET-1) in venous blood from the strangulated bowel loop. The present study focuses on the microcirculatory effects of the released ET-1 in strangulation obstruction. METHODS: In anesthetized pigs strangulation obstruction was induced by increasing pressure in a baby pressure gasket placed around a loop of ileum until venous pressure reached 45 mm Hg. The pigs were randomly allocated into two groups. The nonselective ET(A)/ET(B) antagonist bosentan was administered intravenously (5 mg kg(-1)) to eight pigs (bosentan group) 30 min before strangulation, which was maintained for 90 min. Another eight pigs were treated in same manner except for the bosentan injection (control group). RESULTS: The concentration of ET in arterial and intestinal venous blood increased markedly after intravenous administration of bosentan. Intravenous infusion of bosentan was followed by a reduction in systemic arterial blood pressure. Bosentan reduced vascular resistance and increased blood flow in the normal intestinal mucosa. It also reduced muscularis blood flow in the beginning of the experiment. In strangulated small bowel bosentan inhibited the increase in vascular resistance usually caused by strangulation obstruction. Muscularis blood flow in strangulated small bowel was not affected by bosentan. CONCLUSION: Endothelin is involved in the normal regulation of arterial blood pressure. The increase in vascular resistance associated with strangulation obstruction is caused mainly by locally released endothelin. PMID- 11266278 TI - Role of platelet-activating factor in hepatectomy with Pringle's maneuver. AB - BACKGROUND: Interruption of hepatic inflow is commonly used to reduce blood loss during extensive liver resection, but may cause liver dysfunction. The present study investigated the effects of platelet-activating factor (PAF) antagonist E5880 on total liver warm ischemia and 70% hepatectomy. METHODS: Rabbits were used in this study and were divided into four groups: group 1, those treated with only 70% hepatectomy; group 2, those treated with only 20 min Pringle's maneuver; group 3, those treated with both Pringle's maneuver and 70% hepatectomy without pretreatment; and group 4, those pretreated with PAF antagonist E5880 (0.3 mg/kg) followed by Pringle's maneuver and 70% hepatectomy. The remnant liver function was then evaluated after reperfusion. RESULTS: Seven-day survival rates in both groups 1 and 2 were 100%. E5880 treatment significantly increased 7-day survival rate (group 4: 38% vs group 3: 0%, P < 0.05) after a combination of Pringle's maneuver and 70% hepatectomy. The elevations of serum liver enzymes (GOT, GPT, mGOT, and LDH) were significantly inhibited in group 4 at 1 and 4 h after reperfusion. Portal venous pressure and the energy charge of liver were also significantly improved in group 4, compared with those in group 3. Endothelin-1 levels of arterial and portal venous blood progressively increased after reperfusion; however, there were no significant differences between the two groups. Leukocyte infiltration into the liver was significantly inhibited in group 4. CONCLUSION: E5880 pretreatment has protective effects on liver function after 70% hepatectomy with Pringle's maneuver in rabbits. PMID- 11266279 TI - Collagen fibers in linea alba and rectus sheaths. AB - BACKGROUND: After the description of a general scheme of the architecture of collagen fibers in linea alba and rectus sheaths, variability and differences of fiber architectures were analyzed to describe their functional role. MATERIALS AND METHODS: Using confocal laser scanning microscopy the diameter of each layer of fibril bundles was measured in linea alba and rectus sheaths of 12 human cadavers, and each fibril bundle was classified according to its orientation (oblique I and II, transverse). RESULTS: The mean diameter of fibril bundles in the supraumbilical region of the linea alba was smaller than in the infraumbilical region, and in the supraumbilical region the thickness of the linea alba was smaller than in the infraumbilical region. Analyzing sex-dependent differences in the fiber architecture of the linea alba, a larger amount of transverse fibers relative to oblique fibers were found in females in infraumbilical regions. The thickness of the infraumbilical linea alba was smaller in females than in males, while its width was larger. CONCLUSIONS: There exist gender differences in the architecture of the linea alba. However, whether these morphological differences demonstrate the adaptability of this fiber architecture to biomechanical stress in raised intraabdominal pressure in pregnancy remains to be proven. The transverse fibers act as a counterpart to the intraabdominal pressure whereas the oblique fibers are involved mainly in movements of the trunk. PMID- 11266280 TI - Glutamine utilization in activated lymphocytes from rats receiving endotoxin. AB - BACKGROUND: A beneficial effect of supplemental glutamine for lymphocyte function in patients under metabolic stress has been suggested. Nevertheless, it is not clear how glutamine is used by lymphocytes when under stress. This time course study investigated the effect of endotoxin-induced stress on in vitro glutamine utilization and glutamine-dependent proliferation of activated lymphocytes. METHODS: Metabolic stress was modeled by intraperitoneal (ip) administration of endotoxin (5 mg/kg body wt) to rats. Control animals were injected with sterile saline. Cervical lymph node lymphocytes collected from animals 6, 12, 24, and 48 h following injection were activated with concanavalin A. Proliferation of these activated lymphocytes in the presence of 0.1-2 mM glutamine was determined. The glutamine utilization rate and glutaminase activity in the activated lymphocytes were also determined. RESULTS: The proliferation rate of lymphocytes was not affected by ip administration of endotoxin 6 h following the insult, however, 12, 24, and 48 h following the insult, the maximal response was suppressed (P < 0.05). In addition, at 12, 24, and 48 h, the concentration of glutamine for the maximal response of lymphocytes was lower than that for the control group (P < 0.05). Throughout the investigation period, both the glutamine utilization rate and glutaminase activity in the activated lymphocytes were decreased time dependently. CONCLUSION: The present study demonstrates that glutamine utilization by lymphocytes under a mitogenic challenge in vitro is significantly decreased in the late period after endotoxin injection. This is at least partly due to decreased glutaminase activity and is associated with decreased proliferation rate of mitogen-activated lymphocytes. PMID- 11266281 TI - Comparison of intradermal and subcutaneous injections in lymphatic mapping. AB - BACKGROUND: Sentinel node biopsy (SNB) for melanoma, with its intradermal (ID) injection, has a higher success rate than SNB for breast cancer, which is typically performed with a subcutaneous (SC) or peritumor injection. It is hypothesized that this is in part due to a slower transit time of lymphatic mapping agents through the parenchymal lymphatics of the breast. No study has investigated differences in transit time between different tissues to account for this clinical observation. The goal of the study was to compare transit time between ID and SC injections with common agents used in lymphatic mapping. METHODS: Four injection sites on five domestic pigs were used. Sites were bilateral and included cervical, forelimb, hindlimb, and flank areas. Agents included technetium sulfur colloid (Tc99, filtered and unfiltered), isosulfan blue (IB) dye, and fluorescein (FL) dye. At each site both ID and SC injections were made and the transit time to reach the sentinel node was recorded. The transit time differences were calculated per centimeter distance from the draining lymph node basin. RESULTS: Sentinel nodes were identified draining all sites and found to be hot, blue, or fluorescent (using a Wood's lamp for identification). The cervical and forelimb injection sites drained to the same cervical lymph node basin and both SC and ID injection sites drained to the same sentinel node. Similarly, the hindlimb and flank injection sites both drained to inguinal lymph node basins. The slowest transit time occurred with Tc99 injected SC and the fastest occurred with Tc99 injected ID, whereas both FL dye and IB traveled rapidly to the sentinel node whether injected SC or ID. Large differences were found using unfiltered Tc99 depending on its injection ID (2.7 s/cm +/- 0.5) vs SC (249 s/cm +/- 14.7, P = 0.008). CONCLUSIONS: Tc99 ID injections were significantly faster than SC injection. The slowest and fastest SC injection agents were unfiltered Tc99 and IB, respectively. Dermal injections provide faster transit of lymphatic agents and may improve the identification rate when applied to patients with breast cancer. PMID- 11266282 TI - Cardiac transplantation following a 24-h preservation using a perfusion apparatus. AB - BACKGROUND: We developed a new apparatus for heart preservation and have already reported successful transplantation following 12 h of preservation using this apparatus. The efficacy of coronary perfusion with an oxygenated Celsior solution was investigated through transplantation following 24 h of preservation using the apparatus. MATERIALS AND METHODS: After being harvested, grafts were preserved with a combination of immersion in a 4 degrees C Celsior solution and perfusion with an oxygenated Celsior solution using the apparatus in the coronary perfusion (CP) group and simply immersed in a 4 degrees C Celsior solution in the simple immersion(SI) group. beta-Adenosine triphosphate (beta-ATP), phosphocreatine (Pcr), and inorganic phosphate (P(i)) levels and myocardial pH (pH(i)) were measured immediately after the heart was excised and at 12 and 24 h after preservation. Following preservation, orthotopic transplantation was performed. Cardiac function was measured 2 h after weaning from cardiopulmonary bypass (CPB). RESULTS: beta-ATP/P(i), Pcr/P(i), and pH(i) levels were significantly higher in the CP group than in the SI group at 12 and 24 h after preservation. Four of six animals in the CP group and two of six in the SI group were successfully weaned from CPB. The recovery rates of cardiac function were better in the CP group than in the SI group. CONCLUSION: Twenty-four hours of heart preservation may be possible with a combination of immersion in a 4 degrees C Celsior solution and perfusion with an oxygenated Celsior solution using the perfusion apparatus. PMID- 11266283 TI - Mitochondrial respiration as an early marker of viability in cardiac-arrested rat lungs. AB - BACKGROUND: To evaluate the possibility of using pulmonary mitochondrial respiratory functions as early markers of ischemic lung viability in non-heart beating donors, we investigated the roles of the mitochondria in ischemia reperfusion injury of cardiac-arrested rat lungs. MATERIALS AND METHODS: Male Lewis rats were exposed to various periods of postmortem warm ischemia (0, 1, and 2 h at 21 degrees C). After a pulmonary artery flush and cold preservation (1 h at 4 degrees C), the rat lungs were reperfused using an isolated rat lung model. Each experimental group consisted of three subgroups (n = 7 in each subgroup) to examine pulmonary functions and biochemical measurements. RESULTS: The pulmonary functions after reperfusion were exacerbated after a 1-h postmortem warm ischemia and worsened after a 2-h warm ischemia following cardiac arrest. The mitochondrial respiratory control ratios already significantly decreased after a 1-h warm ischemia compared with nonischemic rat lungs, at which time the value was almost equivalent to that after a 2-h ischemia. There were no significant changes in the state 3 and 4 respiration of the mitochondria, the pulmonary lactate levels, or the lipid peroxide levels in the lung tissues and mitochondria during the first 1-h period of warm ischemia. The adenine nucleotide levels significantly decreased with the prolongation of the period of warm ischemia, but did not seem to be practical, because their determination required a much longer time than that of the mitochondrial respiratory control ratio. CONCLUSION: These results suggested that the mitochondrial respiratory control ratio may be a useful early marker for lung viability after cardiac arrest. PMID- 11266284 TI - Autologous bone marrow implantation induced angiogenesis and improved deteriorated exercise capacity in a rat ischemic hindlimb model. AB - BACKGROUND: Bone marrow possesses endothelial progenitor cells that secrete several growth factors and can contribute to the formation of new capillaries. In the present study, we investigated the extent of angiogenesis induced by implantation of autologous bone marrow cells (BMCs) in a rat ischemic hindlimb model and studied whether the increased collateral vessels can improve deteriorated physical function. MATERIALS AND METHODS: Ischemic hindlimb was created by ligation of the femoral artery and its branches in Dark Agouti (DA) rats. BMCs (1 x 10(7)) were injected percutaneously at six points into the gastrocnemius muscle. To assess angiogenesis, histologic evaluation and microangiography were performed at 2 weeks postligation. Severity of the ischemic insult was evaluated by measuring blood flow in the adductor and gastrocnemius muscles using nonradioactive colored microspheres and by determining the femoral arteriovenous oxygen difference (AVDO(2)) at 2 weeks postligation. Running time on a motor-driven treadmill was used to represent exercise capacity. RESULTS: The histologic evaluation and microangiogram showed that the implanted BMCs induce angiogenesis. Blood flow to the adductor muscle on the treated side in the bone marrow cell implantation (BMI) group was significantly restored to 77.3 +/- 19.3% of that of the normally perfused limb in comparison to that in control groups (P < 0.05). AVDO(2) in the BMI group significantly decreased when compared with AVDO(2) in control groups. Rats in the BMI group ran approximately 1.5 times longer than rats in control groups at 2 and 4 weeks postligation (P < 0.01). CONCLUSIONS: Implantation of autologous BMCs induced angiogenesis and improved deteriorated exercise capacity in our rat ischemic hindlimb model. PMID- 11266285 TI - Anomalous overexpression of p27(Kip1) in sporadic pancreatic endocrine tumors. AB - BACKGROUND: Little is known about the cellular defects and molecular mechanisms leading to pancreatic endocrine tumors (PETs). p27(Kip1) is a universal cyclin dependent kinase inhibitor (CDKI), which acts as a tumor suppressor and a negative regulator of cell cycle. From previous reports, quiescent cells show high levels of p27(Kip1) expression while neoplastic and proliferating cells show no detectable p27(Kip1) expression. We hypothesize that in malignant sporadic PETs, p27(Kip1) expression would be decreased compared with benign PETs and normal pancreatic tissue. METHODS: Western analysis was performed on 28 PETs (7 malignant, 21 benign), 2 nonendocrine cell lines, and 5 endocrine cell lines. Signal intensities were quantitated using densitometry and standardized to normal pancreas. RESULTS: Unexpectedly, increased p27(Kip1) expression as compared with control was seen in both benign and malignant tumors, as well as in all four pancreatic islet tumor cell lines, but not fibroblast or pituitary cell lines, evaluated. There was no difference in p27(Kip1) level between benign and malignant tumors. CONCLUSION: This represents the first report of anomalous p27(Kip1) overexpression in sporadic PETs, and is part of a growing literature describing the paradoxical overexpression of p27(Kip1) in human tumors that includes other endocrine tumors. These studies suggest a unique molecular pathway leading to endocrine tumorigenesis. PMID- 11266287 TI - A serine protease inhibitor, N-alpha-tosyl-l-lysine chloromethyl ketone, prolongs rat hepatic allograft survival. AB - BACKGROUND: Serine protease inhibitors have profound suppressive effects on cellular and humoral immune responses. We investigated the effect of a serine protease inhibitor, N-alpha-tosyl-l-lysine chloromethyl ketone (TLCK), on hepatic allograft survival in rats. Methods. Orthotopic hepatic transplantation was performed in an ACI (RT1(a))-to-LEW (RT1(1)) rat combination. TLCK was administered continuously at a dose of 4.4 mg/kg/day using an osmotic subcutaneous infusion minipump. RESULTS: TLCK prolonged hepatic allograft survival. Histologic staging of acute rejection based on Banff criteria in TLCK treated hepatic allografts was significantly lower than in untreated allografts. TLCK significantly reduced serum concentrations of interferon (IFN)-gamma and tumor necrosis factor (TNF) alpha in allograft recipients. TNF-alpha mRNA levels in TLCK-treated allografts were significantly lower than in untreated allografts. TLCK also decreased perforin mRNA levels in hepatic allografts. Hepatic infiltrates eluted from TLCK-treated allografts showed significantly lower cell mediated lympholytic activity against donor Con A blast cervical lymph node cells than those from untreated allografts. In vitro, TLCK suppressed interleukin-2 production and [(3)H]thymidine incorporation into an allogeneic mixed lymphocyte reaction. CONCLUSION: TLCK suppressed acute allograft rejection, suggesting a novel immunosuppressive strategy for therapy of acute organ rejection. PMID- 11266286 TI - Activation of interleukin-6/STAT3 and liver regeneration following transplantation. AB - BACKGROUND: Every liver that is procured, stored, and transplanted experiences injury from cold ischemia and reperfusion. Most recover quickly, but some grafts sustain enough injury to result in prolonged organ dysfunction or require retransplantation. The molecular mechanisms involved in early graft function and recovery following cold ischemia and reperfusion (I/R) after liver transplantation have not been well defined. Interleukin (IL)-6 is a critical factor in the mitogenic response within the liver, and is important for cell cycle progression and protection from injury. Activation of the latent transcription factor, STAT3, is dependent on IL-6 release. The role of the IL 6/STAT3 pathway and hepatocellular regeneration in graft recovery and cell cycle progression following cold ischemia and reperfusion was studied in a rat liver transplant orthotopic (OLT) model. Methods. Rat OLT was performed in a syngeneic model. The presence, time course, and magnitude of expression of IL-6, STAT3 activation, and upregulation of target immediate early genes were determined in liver grafts with minimal (<1 h) and prolonged (12 h) cold preservation times followed by transplantation. Progression of the cell cycle and replication was confirmed by BrdU uptake. RESULTS: Prolonged cold ischemia resulted in increased IL-6 expression and STAT3 activation. This correlated with upregulation of junB, c-fos, c-myc, and c-jun, immediate early genes associated with hepatic regeneration. Extensive DNA replication was present in livers with 12-h ischemia, demonstrating successful completion of the cell cycle. CONCLUSIONS: The participation of the IL-6/STAT3 pathway leading to cell cycle progression and regeneration is an important component in the recovery of organs immediately following cold preservation and transplantation. PMID- 11266288 TI - Emerging infectious diseases in Russia, 1990-1999. PMID- 11266289 TI - Bovine spongiform encephalopathy and variant Creutzfeldt-Jakob disease: background, evolution, and current concerns. AB - The epidemic of bovine spongiform encephalopathy (BSE) in the United Kingdom, which began in 1986 and has affected nearly 200,000 cattle, is waning to a conclusion, but leaves in its wake an outbreak of human Creutzfeldt-Jakob disease, most probably resulting from the consumption of beef products contaminated by central nervous system tissue. Although averaging only 10-15 cases a year since its first appearance in 1994, its future magnitude and geographic distribution (in countries that have imported infected British cattle or cattle products, or have endogenous BSE) cannot yet be predicted. The possibility that large numbers of apparently healthy persons might be incubating the disease raises concerns about iatrogenic transmissions through instrumentation (surgery and medical diagnostic procedures) and blood and organ donations. Government agencies in many countries continue to implement new measures to minimize this risk. PMID- 11266290 TI - Pesticides and public health: integrated methods of mosquito management. AB - Pesticides have a role in public health as part of sustainable integrated mosquito management. Other components of such management include surveillance, source reduction or prevention, biological control, repellents, traps, and pesticide-resistance management. We assess the future use of mosquito control pesticides in view of niche markets, incentives for new product development, Environmental Protection Agency registration, the Food Quality Protection Act, and improved pest management strategies for mosquito control. PMID- 11266292 TI - Geographic subdivision of the range of the malaria parasite Plasmodium vivax. AB - We examined geographically distinct isolates of Plasmodium vivax and categorized them according to developmental success in Anopheles albimanus. We found that parasites from Central America and Colombia form a group distinct from those of Asia. New World isolates have a distinct chromosomal translocation and an episomal variation in the open reading frame (ORF) 470 DNA sequence that distinguishes them from the other isolates tested. Old World types of P. vivax were introduced into the Americas, and a remnant of this lineage remains in P. simium. It is indistinguishable from Old World P. vivax to the extent determinable by using our encoded markers and the examination of its developmental pattern in mosquitoes. The cohesive characteristics that separate types of P. vivax are predictors of range and potential for transmission and hence require taxonomic distinction. PMID- 11266293 TI - Transferable plasmid-mediated resistance to streptomycin in a clinical isolate of Yersinia pestis. AB - Plasmid-mediated high-level resistance to multiple antibiotics was reported in a clinical isolate of Yersinia pestis in Madagascar in 1997. We describe a second Y. pestis strain with high-level resistance to streptomycin, isolated from a human case of bubonic plague in Madagascar. The resistance determinants were carried by a self-transferable plasmid that could conjugate at high frequencies to other Y. pestis isolates. The plasmid and the host bacterium were different from those previously associated with multiple-drug resistance, indicating that acquisition of resistance plasmids is occurring in this bacterial species. Emergence of resistance to streptomycin in Y. pestis represents a critical public health problem since this antibiotic is used as the first-line treatment against plague in many countries. PMID- 11266291 TI - Quinolone and macrolide resistance in Campylobacter jejuni and C. coli: resistance mechanisms and trends in human isolates. AB - The incidence of human Campylobacter jejuni and C. coli infections has increased markedly in many parts of the world in the last decade as has the number of quinolone-resistant and, to a lesser extent, macrolide-resistant Campylobacter strains causing infections. We review macrolide and quinolone resistance in Campylobacter and track resistance trends in human clinical isolates in relation to use of these agents in food animals. Susceptibility data suggest that erythromycin and other macrolides should remain the drugs of choice in most regions, with systematic surveillance and control measures maintained, but fluoroquinolones may now be of limited use in the empiric treatment of Campylobacter infections in many regions. PMID- 11266294 TI - Nested polymerase chain reaction for amplification of the Cryptosporidium oocyst wall protein gene. AB - We developed a sensitive nested polymerase chain reaction procedure for the Cryptosporidium oocyst wall protein (COWP) gene. Amplification and genotyping were successful in 95.2% of 1,680 fecal samples, 77.6% by the unnested and 17.6% by the nested COWP procedure. The COWP gene was amplified from 2,128 fecal samples: 71 from livestock animals and 2,057 from humans. This series included 706 cases from seven drinking water-associated outbreaks and 51 cases from five swimming pool-associated outbreaks, as well as 1,300 sporadic cases. PMID- 11266295 TI - Preoperative drug dispensing as predictor of surgical site infection. AB - The system used by the National Nosocomial Infection Surveillance (NNIS) program to measure risk of surgical site infection uses a score of 3 on the American Society of Anesthesiologists (ASA)-physical status scale as a measure of underlying illness. The chronic disease score measures health status as a function of age, sex, and 29 chronic diseases, inferred from dispensing of prescription drugs. We studied the relationship between the chronic disease score and surgical site infection and whether the score can supplement the NNIS risk index. In a retrospective comparison of 191 patients with surgical site infection and 378 uninfected controls, the chronic disease score and ASA score were highly correlated. The chronic disease score improved prediction of infection by the NNIS risk index and augmented the ASA score for risk adjustment. PMID- 11266296 TI - Lack of evidence of endogenous avian leukosis virus and endogenous avian retrovirus transmission to measles, mumps, and rubella vaccine recipients. AB - The identification of endogenous avian leukosis virus (ALV) and endogenous avian retrovirus (EAV) in chick cell-derived measles and mumps vaccines in current use has raised concern about transmission of these retroviruses to vaccine recipients. We used serologic and molecular methods to analyze specimens from 206 recipients of measles, mumps, and rubella (MMR) vaccine for evidence of infection with ALV and EAV. A Western blot assay for detecting antibodies to endogenous ALV was developed and validated. All serum samples were negative for antibodies to endogenous ALV by Western blot analysis. Peripheral blood lymphocyte samples from 100 vaccinees were further tested by polymerase chain reaction for both ALV and EAV proviral sequences; all were negative. Matching serum samples were tested by reverse transcriptase polymerase chain reaction for ALV and EAV RNA, and all 100 samples were negative, providing no evidence of viremia. These findings do not indicate the presence of either ALV or EAV infection in MMR vaccine recipients and provide support for current immunization policies. PMID- 11266297 TI - A flea-associated Rickettsia pathogenic for humans. AB - A rickettsia named the ELB agent, or "Rickettsia felis," was identified by molecular biology techniques in American fleas in 1990 and later in four patients from Texas and Mexico. We attempted to isolate this rickettsia from infected fleas at various temperatures and conditions. A representative isolate of the ELB agent, the Marseille strain, was characterized and used to develop a microimmunofluorescence test that detected reactive antibodies in human sera. The ELB agent was isolated from 19 of 20 groups of polymerase chain reaction-proven infected fleas. The microimmunofluorescence results provided serologic evidence of infection by the ELB agent in four patients with fever and rash in France (2) and Brazil (2), supporting the pathogenic role of this rickettsia. Our successful isolation of this rickettsia makes it available for use in serologic tests to determine its clinical spectrum, prevalence, and distribution. PMID- 11266298 TI - Gastroenteritis in sentinel general practices,The Netherlands. AB - From 1996 to 1999, the incidence of gastroenteritis in general practices and the role of a broad range of pathogens in the Netherlands were studied. All patients with gastroenteritis who had visited a general practitioner were reported. All patients who had visited a general practitioner for gastroenteritis (cases) and an equal number of patients visiting for nongastrointestinal symptoms (controls) were invited to participate in a case-control study. The incidence of gastroenteritis was 79.7 per 10,000 person years. Campylobacter was detected most frequently (10% of cases), followed by Giardia lamblia (5%), rotavirus (5%), Norwalk-like viruses (5%) and Salmonella (4%). Our study found that in the Netherlands (population 15.6 million), an estimated 128,000 persons each year consult their general practitioner for gastroenteritis, slightly less than in a comparable study in 1992 to 1993. A pathogen could be detected in almost 40% of patients (bacteria 16%, viruses 15%, parasites 8%). PMID- 11266299 TI - Active bacterial core surveillance of the emerging infections program network. AB - Active Bacterial Core surveillance (ABCs) is a collaboration between the Centers for Disease Control and Prevention and several state health departments and universities participating in the Emerging Infections Program Network. ABCs conducts population-based active surveillance, collects isolates, and performs studies of invasive disease caused by Streptococcus pneumoniae, group A and group B Streptococcus, Neisseria meningitidis, and Haemophilus influenzae for a population of 17 to 30 million. These pathogens caused an estimated 97,000 invasive cases, resulting in 10,000 deaths in the United States in 1998. Incidence rates of these pathogens are described. During 1998, 25% of invasive pneumococcal infections in ABCs areas were not susceptible to penicillin, and 13.3% were not susceptible to three classes of antibiotics. In 1998, early-onset group B streptococcal disease had declined by 65% over the previous 6 years. More information on ABCs is available at www.cdc.gov/ncidod/dbmd/abcs. ABCs specimens will soon be available to researchers through an archive. PMID- 11266300 TI - Emerging Chagas disease: trophic network and cycle of transmission of Trypanosoma cruzi from palm trees in the Amazon. AB - A trophic network involving molds, invertebrates, and vertebrates, ancestrally adapted to the palm tree (Attalaea phalerata) microhabitat, maintains enzootic Trypanosoma cruzi infections in the Amazonian county Paco do Lumiar, state of Maranhao, Brazil. We assessed seropositivity for T. cruzi infections in the human population of the county, searched in palm trees for the triatomines that harbor these infections, and gathered demographic, environmental, and socioeconomic data. Rhodnius pictipes and R. neglectus in palm-tree frond clefts or in houses were infected with T. cruzi (57% and 41%, respectively). Human blood was found in 6.8% of R. pictipes in houses, and 9 of 10 wild Didelphis marsupialis had virulent T. cruzi infections. Increasing human population density, rain forest deforestation, and human predation of local fauna are risk factors for human T. cruzi infections. PMID- 11266302 TI - Hospital control and multidrug-resistant pulmonary tuberculosis in female patients, Lima, Peru. AB - We examined the prevalence of tuberculosis (TB), rate of multidrug-resistant (MDR) TB, and characteristics of TB on a female general medicine ward in Peru. Of 250 patients, 40 (16%) were positive by sputum culture and 27 (11%) by smear, and 8 (3%) had MDRTB. Thirteen (33%) of 40 culture-positive patients had not been suspected of having TB on admission. Six (46%) of 13 patients whose TB was unsuspected on admission had MDRTB, compared with 2 (7%) of 27 suspected cases (p = 0.009). Five (63%) of 8 MDRTB patients were smear positive and therefore highly infective. In developing countries, hospital control, a simple method of reducing the spread of MDRTB, is neglected. PMID- 11266301 TI - Persistence and variability of Stenotrophomonas maltophilia in cystic fibrosis patients, Madrid, 1991-1998. AB - During 1991 to 1998 at least one Stenotrophomonas maltophilia pulmonary infection was observed in 25 (24%) of 104 cystic fibrosis patients at the same unit of our hospital in Spain. Ribotyping and pulse-field gel electrophoresis (PFGE) characterization of 76 S. maltophilia isolates from these patients indicated an overall clonal incidence of 47.1%, reflecting new strains in 44% of patients with repeated positive cultures for S. maltophilia. Six patients with repeated episodes were persistently colonized (> or = 6 months) with the same strain. S. maltophilia bacterial counts were higher (geometric mean, 2.9 x 10(8) cfu/mL) in patients with repeated episodes than in those with a single episode (8.4 x 10(4) cfu/mL, p < 0.01). Single episodes of S. maltophilia occurred in patients < 10 years of age (43% [6/14]), whereas chronic colonization occurred more frequently in older patients (> 16 years of age). PMID- 11266303 TI - Outbreak of West Nile virus infection, Volgograd Region, Russia, 1999. AB - From July 25 to October 1, 1999, 826 patients were admitted to Volgograd Region, Russia, hospitals with acute aseptic meningoencephalitis, meningitis, or fever consistent with arboviral infection. Of 84 cases of meningoencephalitis, 40 were fatal. Fourteen brain specimens were positive in reverse transcriptase-polymerase chain reaction assays, confirming the presence of West Nile/Kunjin virus. PMID- 11266304 TI - Rapid identification of Corynebacterium diphtheriae clonal group associated with diphtheria epidemic, Russian Federation. AB - We used 199 Corynebacterium diphtheriae isolated from 1995 to 1997 in Russia to evaluate the ability of random amplified polymorphic DNA (RAPD) to identify the unique clonal group that emerged there in 1990. Our data show that RAPD can reliably, reproducibly, and rapidly screen a large number of strains to identify the epidemic clonal group. PMID- 11266305 TI - Shigella spp. surveillance in Indonesia: the emergence or reemergence of S. dysenteriae. AB - From June 1998 through November 1999, Shigella spp. were isolated in 5% of samples from 3,848 children and adults with severe diarrheal illness in hospitals throughout Indonesia. S. dysenteriae has reemerged in Bali, Kalimantan, and Batam and was detected in Jakarta after a hiatus of 15 years. PMID- 11266306 TI - Tracking Cryptosporidium parvum by sequence analysis of small double-stranded RNA. AB - We sequenced a 173-nucleotide fragment of the small double-stranded viruslike RNA of Cryptosporidium parvum isolates from 23 calves and 38 humans. Sequence diversity was detected at 17 sites. Isolates from the same outbreak had identical double-stranded RNA sequences, suggesting that this technique may be useful for tracking Cryptosporidium infection sources. PMID- 11266308 TI - Disseminated Neocosmospora vasinfecta infection in a patient with acute nonlymphocytic leukemia. AB - We report Neocosmospora vasinfecta infection following chemotherapy for acute nonlymphocytic leukemia. N. vasinfecta, a plant pathogen, was identified by culture and genetic sequencing. Susceptibility testing revealed in vitro resistance for common antifungals. PMID- 11266307 TI - Pathologic studies of fatal cases in outbreak of hand, foot, and mouth disease, Taiwan. AB - In 1998, an outbreak of enterovirus 71-associated hand, foot, and mouth disease occurred in Taiwan. Pathologic studies of two fatal cases with similar clinical features revealed two different causative agents, emphasizing the need for postmortem examinations and modern pathologic techniques in an outbreak investigation. PMID- 11266309 TI - Adventitious viral genomes in vaccines but not in vaccinees. PMID- 11266310 TI - Strengthening national preparedness for smallpox: an update. PMID- 11266311 TI - High-level ciprofloxacin resistance in Neisseria gonorrhoeae: first report from Israel. PMID- 11266312 TI - An unusual bacterium causing a brain abscess. PMID- 11266313 TI - First glycopeptide-resistant Enterococcus faecium isolate from blood culture in Ankara, Turkey. PMID- 11266314 TI - Antimicrobial-drug use and methicillin-resistant Staphylococcus aureus. PMID- 11266315 TI - Lack of evidence for chloramphenicol resistance in Neisseria meningitidis, Africa. PMID- 11266316 TI - Iron loading and disease surveillance. PMID- 11266317 TI - Surprising findings following a Belgian food contamination with polychlorobiphenyls and dioxins. AB - We found that 12.1% of Belgian export meat samples from chicken or pork, unrelated to the PCB/dioxin crisis from 1999, contained more than 50 ng polychlorinated biphenyls (PCBs)/g fat and that 6.5% of samples contain more than 20 ng/g fat for the sum of 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT) and its metabolites. Part of this background contamination stems from imported animal feed ingredients (fish flour and grains), sometimes contaminated by recent use of DDT, as can be deduced from the ratio between DDT and its main metabolite, 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE). However, after comparing PCB concentrations in fish flour and grains with those found in meat, we suggest that the high concentrations stem from recycled fat. This is the first paper describing background concentrations of PCBs in animal meat from Belgium. PMID- 11266318 TI - Irreversible binding and adrenocorticolytic activity of the DDT metabolite 3 methylsulfonyl-DDE examined in tissue-slice culture. AB - The persistent adrenocorticolytic DDT metabolite 3-methylsulfonyl-DDE (MeSO(2) DDE) was originally identified in Baltic grey seals, a population suffering from adrenocortical hyperplasia. In mice, MeSO(2)-DDE induces mitochondrial degeneration and cellular necrosis in the adrenal zona fasciculata. In this study, we used precision-cut tissue slice culture to examine local CYP11B1 catalyzed irreversible binding of MeSO(2)-DDE in the murine adrenal cortex. We also examined effects on steroid hormone secretion, histology, and ultrastructure. As determined by microautoradiography, selective binding occurred in zona fasciculata of slices exposed to MeSO(2)-[(14)C]-DDE. Quantification of binding by phosphorautoradiography revealed a 3-fold reduction of binding in slices co-exposed to the CYP11B1 inhibitor metyrapone. As measured by HPLC, corticosterone and 11-deoxycorticosterone secretion to the medium increased linearly for at least 24 hr. Addition of the ACTH analog tetracosactide caused an 8-fold increase in corticosterone secretion. Addition of metyrapone reduced corticosterone secretion 4-fold. Exposure of slices to MeSO(2)-DDE (50 microM) reduced the rate of corticosterone secretion by 90% after 24 hr of incubation. As determined by electron microscopy, vacuolated mitochondria were present in zona fasciculata of slices exposed to MeSO(2)-DDE (50 microM) for 24 hr. Our findings show that all effects of MeSO(2)-DDE previously reported in vivo could be reproduced in adrenal slice culture ex vivo. This test system allows analysis of zone-specific irreversible binding and effects on steroid hormone secretion and target cell ultrastructure. We propose adrenal slice culture as a simple ex vivo test system with which to examine the adrenocorticolytic activity of xenobiotics in human and wild animal tissue. PMID- 11266319 TI - The masculinization of the fetus during pregnancy due to inhalation of diesel exhaust. AB - This study was conducted to determine the impact of diesel exhaust inhalation on the fetus. Seventy-two pregnant rats and 18 nonpregnant rats were divided into three groups: a group exposed to total diesel engine exhaust containing 5.63 mg/m(3) particulate matter, 4.10 ppm nitrogen dioxide, and 8.10 ppm nitrogen oxide; a group exposed to filtered exhaust without particulate matter; and a group exposed to clean air. The exposure period was from day 7 until day 20 of pregnancy. In addition, 15 pregnant rats were treated with aromatase inhibitors or testosterone to clarify the process by which diesel exhaust exerts its toxicity. The anogenital distance was significantly longer in male and female fetuses from both exhaust-exposed groups than in those of the control. Differentiation of the testis, ovary, and thymus was delayed and disturbed. Maternal testosterone and progesterone levels, which increased due to pregnancy whether or not the rats were exposed, were significantly higher and lower, respectively, in the pregnant rats exposed to total exhaust and filtered exhaust. The serum adrenocorticotropic hormone (ACTH) level and urinary excretion of 17 hydroxycorticosteroids (OHCS) did not differ among the pregnant groups. These results indicate that elevated testosterone did not result from elevated maternal adrenal function. The feto-placental-ovarian unit and inhibition of aromatase activity and synthesis caused by diesel exhaust inhalation might have played an essential role in the accumulation of testosterone. Since both exhaust-exposed groups showed almost the same reactions toward the inhalation, the gaseous phase must have included the relevant toxicants. PMID- 11266320 TI - Analysis of dietary intake of selected metals in the NHEXAS-Maryland investigation. AB - As part of a large pilot investigation of multimedia exposure to several classes of environmental contaminants, the National Human Exposure Assessment Survey (NHEXAS)-Maryland study, we collected 388 semiquantitative food checklists and duplicate diet solid food samples, analyzed for arsenic, cadmium, chromium, and lead concentrations, from 80 individuals in Maryland in 1995-1996 in a repeated measures design. Here we explore several methods to infer foods most strongly associated with concentrations of these metals observed in the duplicate diet in our data set. We employed two techniques in which logarithmically transformed metal concentrations in the duplicate diet were regressed on individual food item consumption using algorithms designed to identify the foods most associated with the observed duplicate diet concentrations. We also employed an alternative strategy in which foods to be used as independent variables in regression were selected using data collected in national food consumption and residue surveys, with regression procedures proceeding with the selected foods in a similar manner. The concordance of foods selected as major predictors among these three techniques is noteworthy and is discussed. Finally, the Dietary Exposure Potential Model (DEPM) was used with the Dietary Checklist data to predict duplicate diet concentrations within our sample. A comparison between the predicted values and those observed gave R(2) values of 0.180, 0.206, and 0.076 for As, Cd, and Pb, respectively (p < 0.0001 in all cases). We discuss the significance of these observations and the implications for dietary-exposure based risk analysis and dietary intake epidemiology. PMID- 11266322 TI - Estrogenic activity of phenolic additives determined by an in vitro yeast bioassay. AB - We used a recombinant yeast estrogen assay to assess the activity of 73 phenolic additives that are used as sunscreens, preservatives, disinfectants, antioxidants, flavorings, or for perfumery. Thirty-two of these compounds displayed activity: 22 with potencies relative to 17beta-estradiol, ranging from 1/3,000 to < 1/3,000,000, and 10 compounds with an impaired response that could not be directly compared with 17beta-estradiol. Forty-one compounds were inactive. The major criteria for activity appear to be the presence of an unhindered phenolic OH group in a para position and a molecular weight of 140-250 Da. PMID- 11266321 TI - Dermal in vitro penetration of methiocarb, paclobutrazol, and pirimicarb: effect of nonylphenolethoxylate and protective gloves. AB - Dermal exposure has become the major route of human occupational exposure to pesticides. Detergents are used as part of formulated pesticide products and are known to change the barrier properties of human skin in vitro. However, studies on the influence of detergents as well as protective glove materials on dermal penetration of pesticides are scarce. In an experiment using in vitro static diffusion cells mounted with human skin, we evaluated the effect of nonylphenol ethoxylate on dermal penetration of three extensively used pesticides- methiocarb, paclobutrazol, and pirimicarb--and the protection against dermal penetration offered by protective gloves made of latex or nitrile. There was a general tendency, though not statistically significant for all pesticides, for nonylphenolethoxylate to decrease the percutaneous penetration of the three pesticides. The nitrile generally offered better protection against percutaneous penetration of pesticides than did latex, but the degree of protection decreased over time and depended on the pesticides used. PMID- 11266323 TI - Beta(1-->3)-glucan in house dust of German homes: housing characteristics, occupant behavior, and relations with endotoxins, allergens, and molds. AB - beta(1-->3)-Glucans are potent proinflammatory agents that have been suggested to play a role in indoor-related respiratory health effects. The aim of this study was to assess whether beta(1-->3)-glucan concentrations in house dust are correlated with levels of endotoxins, allergens, and culturable mold spore counts in house dust. Further, the associations of beta(1-->3)-glucan with housing characteristics and occupant behavior were assessed. beta(1-->3)-Glucan was measured in settled house dust from living room floors of 395 homes of two German cities, Erfurt and Hamburg, with a specific enzyme immunoassay. Concentrations ranged from below the limit of detection to 19,013 microg/m(2) (22,588 microg/g dust). Concentrations per square meter were found to be correlated with endotoxins, mite and cat allergens, and culturable mold spores. Correlations were weaker when concentrations were expressed per gram of dust, indicating that variance in concentrations of all factors is largely determined by the amount of dust sampled. Associations between beta(1-->3)-glucan, housing characteristics, and occupant behavior were found for concentrations per square meter but not for concentrations per gram of dust. The following characteristics were associated with a significant increase in beta(1-->3)-glucan levels: carpets in the living room [means ratio (MR) = 1.9-2.1], keeping a dog inside (MR = 1.4), use of the home by four or more persons (MR = 1.4), use of the living room for > 180 hr/week (MR = 2.1), lower frequency of vacuum cleaning (MR = 1.6-3.0) and dust cleaning (MR = 1.2 and 1.4, respectively), and presence of mold spots during the past 12 months (MR = 1.4). We conclude that that the amount of dust sampled can be used as a proxy for hygiene and that beta(1-->3)-glucan concentrations per square meter are related to the amount of dust sampled. PMID- 11266324 TI - Longitudinal investigation of dietary exposure to selected pesticides. AB - Between September 1995 and September 1996, 4-day composite duplicate plate samples (379 solid food samples and 303 beverage samples) were obtained from a stratified random sample of 75 individuals in Maryland and analyzed for the presence of 10 pesticides. Samples were collected in each of six approximately equally spaced cycles as part of a larger pilot investigation of longitudinal exposure to pesticides and other elements. Chlorpyrifos was detected in 38.3% of the solid food samples, malathion in 75.2%, and p,p'-DDE in 21.4%. Other pesticides were detected in less than 10% of the solid food samples. Pesticide residues were not detected in duplicate beverage samples. In solid food samples, the mean concentration of chlorpyrifos was 0.7 (SD 1.7) microg/kg, 1.8 (2.1) for malathion, and 0.2 (0.6) for p,p'-DDE. The detection rate and mean concentration of chlorpyrifos, malathion, and p,p'-DDE varied by a factor of 2-3 among sampling cycles and significantly according to results from several statistical analyses. Co-occurrence of chlorpyrifos and malathion in solid food samples was found relatively frequently and also varied with time. Pesticides were detected in food samples with greatest frequency in spring and summer months and with lowest frequency in winter months. These results support the hypothesis that 4-day average exposure to chlorpyrifos and malathion varies over time for this population mean and for individual members of the population and that correlation between exposures to these two organophosphate pesticides can occur. The measurements of pesticide levels in duplicate plate samples presented here can be used to evaluate and set parameters for dietary exposure models. PMID- 11266326 TI - The influence of climate variation and change on diarrheal disease in the Pacific Islands. AB - Freshwater resources are a high-priority issue in the Pacific region. Water shortage is a serious problem in many small island states, and many depend heavily on rainwater as the source of their water. Lack of safe water supplies is an important factor in diarrheal illness. There have been no previous studies looking specifically at the relationship between climate variability and diarrhea in the Pacific region. We carried out two related studies to explore the potential relationship between climate variability and the incidence of diarrhea in the Pacific Islands. In the first study, we examined the average annual rates of diarrhea in adults, as well as temperature and water availability from 1986 to 1994 for 18 Pacific Island countries. There was a positive association between annual average temperature and the rate of diarrhea reports, and a negative association between water availability and diarrhea rates. In the second study, we examined diarrhea notifications in Fiji in relation to estimates of temperature and rainfall, using Poisson regression analysis of monthly data for 1978-1998. There were positive associations between diarrhea reports and temperature and between diarrhea reports and extremes of rainfall. These results are consistent with previous research and suggest that global climate change is likely to exacerbate diarrheal illness in many Pacific Island countries. PMID- 11266325 TI - Differences among black smoke, PM(10), and PM(1.0) levels at Urban Measurement Sites. AB - n Amsterdam, the Netherlands, we measured airborne particulate matter (PM) during winter 1998-1999, taking daily average measurements at an urban background site, at a busy street, and at a motorway. Comparison of black smoke, PM(10), and PM(1.0) levels showed that daily averages were highly correlated over time. Median daily concentrations were elevated at sites affected by traffic. The highest increase relative to the background in median daily concentration was noted for black smoke at the motorway (300%), whereas for PM(10) and PM(1.0) the increase was only 37% and 30%. These results indicate that mass measurements of ambient particulate matter underestimate the exposure to particles generated by traffic. PMID- 11266327 TI - Environmental sensitivities: prevalence of major symptoms in a referral center: the Nova Scotia Environmental Sensitivities Research Center Study. AB - Although the phenomenon of environmental sensitivities (ES) has no clear etiology nor well-accepted pathophysiology, affected individuals experience symptoms that cause varying levels of dysfunction. Through a dedicated, government-funded research and treatment center, a detailed questionnaire covering 217 symptoms in 13 systems was mailed in 1997-1998 to 812 individuals referred to the center by physicians. A total of 385 (47%) questionnaires were returned, and data were analyzed on 351 individuals. Participants tended to be women (80%), middle-aged individuals (37% age 40-49 years), and those in higher educational groups (28% completed university), but there was wide variation in demographic variables. General symptoms such as difficulty concentrating, fatigue, forgetfulness, and irritability dominated the overall prevalence of symptoms since the start of their illness. Those related to irritation such as sneezing, itchy or burning eyes, and hoarseness or loss of voice were more common after exposure to environmental irritants. Ranking of symptoms using severity scores was consistent between men and women. Overall scores were higher in women, in participants who were separated or divorced, and in low-income groups. The type and consistency of symptoms experienced after exposure to triggering substances may not fit a purely psychogenic theory. PMID- 11266328 TI - Diagnostic chelation challenge with DMSA: a biomarker of long-term mercury exposure? AB - Chelation challenge testing has been used to assess the body burden of various metals. The best-known example is EDTA challenge in lead-exposed individuals. This study assessed diagnostic chelation challenge with dimercaptosuccinic acid (DMSA) as a measure of mercury body burden among mercury-exposed workers. Former employees at a chloralkali plant, for whom detailed exposure histories were available (n = 119), and unexposed controls (n = 101) completed 24-hr urine collections before and after the administration of two doses of DMSA, 10 mg/kg. The urinary response to DMSA was measured as both the absolute change and the relative change in mercury excretion. The average 24-hr mercury excretion was 4.3 microg/24 hr before chelation, and 7.8 microg/24 hr after chelation. There was no association between past occupational mercury exposure and the urinary excretion of mercury either before or after DMSA administration. There was also no association between urinary mercury excretion and the number of dental amalgam surfaces, in contrast to recent published results. We believe the most likely reason that DMSA chelation challenge failed to reflect past mercury exposure was the elapsed time (several years) since the exposure had ended. These results provide normative values for urinary mercury excretion both before and after DMSA challenge, and suggest that DMSA chelation challenge is not useful as a biomarker of past mercury exposure. PMID- 11266329 TI - Associations of blood levels of PCB, HCHS, and HCB with numbers of lymphocyte subpopulations, in vitro lymphocyte response, plasma cytokine levels, and immunoglobulin autoantibodies. AB - Pentachlorophenol (PCP), hexachlorocyclohexane-[alpha], -beta, and -[gamma] (HCH [alpha], -beta, and -[gamma]), polychlorinated biphenyls (PCBs), and hexachlorobenzene (HCB) are widely distributed industrial chemicals. They are suspected to induce immunologic impairments in exposed individuals. We examined dose-response relationships of blood levels of these chemicals with cellular (numbers of lymphocyte subpopulations, in vitro lymphocyte response) or humoral (plasma cytokine levels, immunoglobulin autoantibodies) immunologic dysfunctions. We studied 146 patients who had been occupationally exposed primarily to PCBs for more than 6 months. Lymphocyte subpopulations, in vitro responses to mitogens and allogeneic stimulator cells, plasma neopterin, cytokines, soluble cytokine receptors, soluble adhesion molecules, anti-Ig autoantibodies, and liver transaminases were determined. Blood levels of the different compounds were strongly correlated with one another. There were only weak dose-response relationships between blood levels of PCBs with cellular immune parameters, and of HCHs and HCB with humoral immune parameters. An exception was the statistically significant negative association of HCB with interferon-[gamma] (IFN-[gamma]), indicating that HCB has a significant impact on Th1 lymphocytes. Patients with HCB blood levels above the mean of 1,109 ng/L more often had undetectable IFN-[gamma] blood levels than patients below the mean. Patients with increased PCB 138 (> 710 ng/L) had more frequently undetectable interleukin-4 blood levels than patients with PCB 138 below the mean, and patients with increased PCB 101 (> 31 ng/L) more often had low DR+ cell counts in the blood (< 190/microL) than patients with PCB 101 below the mean. To assess possible cumulative effects, we compared patients who had blood levels of all compounds below background with patients who had blood levels of all compounds above background. Patients with low or absent blood levels of the compounds studied had higher IFN-[gamma] plasma levels, providing some evidence for a cumulative effect of several weakly active compounds. In conclusion, exposure to PCBs, HCB, or HCHs is associated with weak immunologic abnormalities. These results contrast with those obtained in earlier studies of blood levels of PCP, which showed a strong dose-dependent relationship with immunologic impairments. Our data suggest that long-term exposure of patients to HCB suppresses IFN-[gamma] production. PMID- 11266330 TI - Chernobyl fallout and outcome of pregnancy in Finland. AB - Possible effects of Chernobyl fallout on outcome of pregnancy in Finland were evaluated in a nationwide follow-up study. The outcomes were the rate of live births and stillbirths, pregnancy loss, and induced abortions by municipality. Exposure was assessed based on nationwide surveys of radiation dose rate from the Chernobyl fallout, from both external and internal exposures. Using these measurements, we estimated the monthly dose rate for each of the 455 Finnish municipalities. On average, the dose rate from Chernobyl fallout reached 50 microSv per month in May 1986--a doubling of the natural background radiation. In the most heavily affected area, 4 times the normal background dose rates were recorded. Given the underlying regional differences in live birth, stillbirth, and abortion rates, we used longitudinal analysis comparing changes over time within municipalities. A temporary decline in the live birth rate had already begun before 1986, with no clear relationship to the level of fallout. A statistically significant increase in spontaneous abortions with dose of radiation was observed. No marked changes in induced abortions or stillbirths were observed. The decrease in the live birth rate is probably not a biological effect of radiation, but more likely related to public concerns of the fallout. The effect on spontaneous abortions should be interpreted with caution, because of potential bias or confounding. Further, there is little support in the epidemiologic literature on effects of very low doses of radiation on pregnancy outcome. PMID- 11266331 TI - Children's health, susceptibility, and regulatory approaches to reducing risks from chemical carcinogens. AB - Risk-based regulation of chemical exposures from the environment generally relies on assumptions about the extent of people's susceptibility to chemically induced diseases. Those assumptions are intended to be health-protective; that is, they err on the side of overstating susceptibility. Recent concern about children's special susceptibilities has led to proposals that would make risk-based regulations one-tenth more stringent, unless data are available to refute the assumption that children are more susceptible than adults. In this paper we highlight some of the questions that should be addressed in the context of risk assessment to determine whether such increased stringency would accomplish the desired result of improving children's health. In particular, characterizing benefits of greater stringency requires more information about dose-response relationships than is currently available. Lowering regulatory levels has attendant costs but may not achieve benefits, for example, if the previous level were already below an actual or practical threshold. Without an ability to understand the potential benefit (or lack thereof) of the additional stringency, an appropriate consideration of benefits and costs is not possible. PMID- 11266333 TI - Workshop Egyptox-2000: for better environmental education by the birth of the new millennium. PMID- 11266332 TI - Paternal occupational exposures and childhood cancer. AB - The objective of the study described here was to test the hypothesis that paternal occupational exposure near conception increases the risk of cancer in the offspring. We conducted a cohort study based on a population of 235,635 children born shortly after two different censuses in Sweden. The children were followed from birth to 14 years, and cases of cancer were identified in the Swedish Cancer Registry. Occupational hygienists assessed the probability of exposure to different agents in each combination of the father's industry and occupation as reported in the censuses. We also analyzed individual job titles. We compared the cancer incidence among children of exposed fathers to that among children of unexposed fathers using Cox proportional hazards modeling. The main findings were an increased risk of nervous system tumors related to paternal occupational exposure to pesticides [relative risk (RR) = 2.36; 95% confidence interval (CI), 1.27-4.39] and work as a painter (RR = 3.65; 95% CI, 1.71-7.80), and an increased risk of leukemia related to wood work by fathers (RR = 2.18; 95% CI, 1.26-3.78). We found no associations between childhood leukemia and paternal exposure to pesticides or paint. Our results support previous findings of an increased risk of childhood brain tumors and leukemia associated with certain paternal occupational exposures. Some findings in previous studies were not confirmed in this study. PMID- 11266335 TI - Opportunity and challenge in China. PMID- 11266334 TI - An unusual case of mixed-dust exposure involving a "noncommercial" asbestos. AB - Our health center evaluated an individual for suspected pneumoconiosis, which had resulted from exposures in a foundry/metal reclamation facility. Appropriate consent forms were obtained for the procedures. Historically, individuals who work in foundries have been exposed to various types of dusts. The clinical findings in this case were consistent with silicosis with a suspicion of asbestos induced changes as well. A sample from this individual, analyzed by electron microscopy, showed both classical and atypical ferruginous bodies. The uncoated fiber burden in this individual indicated an appreciable number of anthophyllite asbestos fibers. This finding, coupled with analysis of cores from ferruginous bodies and the presence of ferruginous bodies in areas of interstitial fibrosis, pathologically supported the diagnosis of asbestos-related disease. The unique factor associated with this case is that unlike in some settings in Finland where anthophyllite was mined and used commercially, this mineral fiber is not commonly found in commercially used asbestos products in the United States. Although the actual source of the asbestos exposure in this case is still being sought, it should be recognized that anthophyllite is a contaminant of many other minerals used in workplace environments, including foundries. The fiber burden indicates a unique type of exposure, differing from that usually construed as typical in occupational settings in the United States. PMID- 11266336 TI - Arsenic contamination in West Bengal and Bangladesh: statistical errors. PMID- 11266338 TI - Scientific theory versus legal theory. PMID- 11266340 TI - Gaps in pesticide reporting lead to underestimates of risk. PMID- 11266341 TI - Inaccurate models for mixtures. PMID- 11266342 TI - Wasting away in the South Pacific. PMID- 11266343 TI - Finding Pfiesteria fast. PMID- 11266344 TI - Prescription for mutant prevention. PMID- 11266345 TI - Leading to drug abuse. PMID- 11266346 TI - NIEHS trainees: hail fellows, well met. PMID- 11266347 TI - Toxic beryllium: new solutions for a chronic problem. PMID- 11266348 TI - Dam-building decisions: a new flood of fairness. PMID- 11266349 TI - Boycott! PMID- 11266350 TI - Comment on Lee M. Silver's article 'Reprogenetics: third millenium speculation' in EMBO reports, November 2000. PMID- 11266351 TI - Stem cell research--why is it regarded as a threat? An investigation of the economic and ethical arguments made against research with human embryonic stem cells. PMID- 11266352 TI - Myth, menace or medical blessing? The clinical potential and the problems of genetic vaccines. PMID- 11266353 TI - Changing the rules? The agreement between Celera and Science magazine concerning Celera's publication of its human genome sequence is upsetting many researchers in bioinformatics. PMID- 11266355 TI - Bacteria rule! The start of a new era in bacterial microbiology? PMID- 11266354 TI - Talking apples and oranges: The EU and the USA continue to struggle over exports of US hormone-treated beef to Europe. PMID- 11266356 TI - Can we afford to pay? The bioVision 2001 summit in France examined the impact of biotechnology in the developed and the Third World, and the cost of inaction when addressing some of humankind's most pressing problems. PMID- 11266357 TI - Making connections. Meeting: axon guidance and neural plasticity. PMID- 11266358 TI - New routes to membrane protein structures. Practical course: current methods in membrane protein research. PMID- 11266359 TI - Hens, cocks and avian sex determination. A quest for genes on Z or W? AB - The sex of an individual is generally determined genetically by genes on one of the two sex chromosomes. In mammals, for instance, the presence of the male specific Y chromosome confers maleness, whereas in Drosophila melanogaster and CAENORHABDITIS: elegans it is the number of X chromosomes that matters. For birds (males ZZ, females ZW), however, the situation remains unclear. The recent discovery that the Z-linked DMRT1 gene, which is conserved across phyla as a gene involved in sexual differentiation, is expressed early in male development suggests that it might be the number of Z chromosomes that regulate sex in birds. On the other hand, the recent identification of the first protein unique to female birds, encoded by the W-linked PKCIW gene, and the observation that it is expressed early in female gonads, suggests that the W chromosome plays a role in avian sexual differentiation. Clearly defining the roles of the DMRT1 and PKC1W genes in gonadal development, and ultimately determining whether avian sex is dependent on Z or W, will require transgenic experiments. PMID- 11266360 TI - Telomerase subunit overexpression suppresses telomere-specific checkpoint activation in the yeast yku80 mutant. AB - Ku is a conserved heterodimeric DNA-binding protein that plays critical roles in DNA repair and telomere homeostasis. In Saccharomyces cerevisiae, deletion of YKU70 or YKU80 results in an inability to grow at 37 degrees C. This is suppressed by overexpression of several components of telomerase (EST1, EST2 and TLC1). We show that overexpression of EST2 or TLC1 in yku80 mutants does not restore efficient DNA repair, or restore normal telomere function, as measured by telomere length, single-stranded G-rich strand or transcriptional silencing. Instead, yku80 mutants activate a Rad53p-dependent DNA-damage checkpoint at 37 degrees C and this is suppressed by overexpression of EST2 or TLC1. Indeed, deletion of genes required for Rad53p activation also suppresses the yku80 temperature sensitivity. These results suggest that activation of the DNA-damage checkpoint in yku mutants at 37 degrees C does not result from reduced telomere length per se, but reflects an alteration of the telomere structure that is recognized as damaged DNA. PMID- 11266361 TI - Ku-deficient yeast strains exhibit alternative states of silencing competence. AB - In Saccharomyces cerevisiae, efficient silencer function requires telomere proximity, i.e. compartments of the nucleoplasm enriched in silencing factors. Accordingly, silencers located far from telomeres function inefficiently. We show here that cells lacking yKu balance between two mitotically stable states of silencing competence. In one, a partial delocalization of telomeres and silencing factors throughout the nucleoplasm correlates with enhanced silencing at a non telomeric locus, while in the other, telomeres retain their focal pattern of distribution and there is no repression at the non-telomeric locus, as observed in wild-type cells. The two states also differ in their level of residual telomeric silencing. These findings indicate the existence of a yKu-independent pathway of telomere clustering and Sir localization. Interestingly, this pathway appears to be under epigenetic control. PMID- 11266362 TI - The Drosophila homolog of the human AF10 is an HP1-interacting suppressor of position effect variegation. AB - In chromosomal rearrangements of acute myeloid leukaemia patients the mixed lineage leukaemia (MLL) gene, a human homolog of the Drosophila gene trithorax, is frequently fused to AF10. Here we describe the identification and a functional characterization of the Drosophila homolog dAF10. We show that dAF10 functions in heterochromatin-dependent genomic silencing of position effect variegation, a phenomenon associated with chromosomal rearrangements that cause mosaic expression of euchromatic genes when relocated next to heterochromatin. We also demonstrate that dAF10 can associate with the heterochromatin protein 1 (HP1) in vitro and in vivo. The results indicate that dAF10 is an HP1-interacting component of the heterochromatin-dependent gene silencing pathway, which either contributes to the stability of the heterochromatin complex or to its function. PMID- 11266363 TI - Cleavage of non-tRNA substrates by eukaryal tRNA splicing endonucleases. AB - Eukaryal tRNA splicing endonucleases use the mature domains of pre-tRNAs as their primary recognition elements. However, they can also cleave in a mode that is independent of the mature domain, when substrates are able to form the bulge helix-bulge structure (BHB), which is cleaved by archaeal tRNA endonucleases. We present evidence that the eukaryal enzymes cleave their substrates after forming a structure that resembles the BHB. Consequently, these enzymes can cleave substrates that lack the mature domain altogether. That raises the possibility that these enzymes could also cleave non-tRNA substrates that already have a BHB. As predicted, they can do so, both in vitro and in vivo. PMID- 11266364 TI - ASK1 is required for sustained activations of JNK/p38 MAP kinases and apoptosis. AB - Apoptosis signal-regulating kinase (ASK) 1 is activated in response to various cytotoxic stresses including TNF, Fas and reactive oxygen species (ROS) such as H(2)O(2), and activates c-Jun NH(2)-terminal kinase (JNK) and p38. However, the roles of JNK and p38 signaling pathways during apoptosis have been controversial. Here we show that by deleting ASK1 in mice, TNF- and H(2)O(2)-induced sustained activations of JNK and p38 are lost in ASK1(-/-) embryonic fibroblasts, and that ASK1(-/-) cells are resistant to TNF- and H(2)O(2)-induced apoptosis. TNF- but not Fas-induced apoptosis requires ROS-dependent activation of ASK1-JNK/p38 pathways. Thus, ASK1 is selectively required for TNF- and oxidative stress induced sustained activations of JNK/p38 and apoptosis. PMID- 11266365 TI - Bacterial Na(+)-ATP synthase has an undecameric rotor. AB - Synthesis of adenosine triphosphate (ATP) by the F(1)F(0) ATP synthase involves a membrane-embedded rotary engine, the F(0) domain, which drives the extra membranous catalytic F(1) domain. The F(0) domain consists of subunits a(1)b(2) and a cylindrical rotor assembled from 9-14 alpha-helical hairpin-shaped c subunits. According to structural analyses, rotors contain 10 c-subunits in yeast and 14 in chloroplast ATP synthases. We determined the rotor stoichiometry of Ilyobacter tartaricus ATP synthase by atomic force microscopy and cryo-electron microscopy, and show the cylindrical sodium-driven rotor to comprise 11 c subunits. PMID- 11266366 TI - The structural GDP/GTP cycle of human Arf6. AB - The small GTP-binding protein Arf6 coordinates membrane traffic at the plasma membrane with aspects of cytoskeleton organization. This function does not overlap with that of other members of the ADP-ribosylation factor (Arf) family, although their switch regions, which are their major sites of interaction with regulators and effectors, have virtually identical sequences. Here we report the crystal structure of full-length, non-myristoylated human Arf6 bound to GTPgammaS. Unlike their GDP-bound forms, the active forms of Arf6 and Arf1 are very similar. Thus, the switch regions are discriminatory elements between Arf isoforms in their inactive but not in their active forms, a property that may generalize to other families of small G proteins. This suggests that GTP-bound Arfs may establish specific interactions outside the switch regions and/or be recognized in their cellular context rather than as isolated proteins. The structure also allows further insight into the lack of spontaneous GTPase activity of Arf proteins. PMID- 11266367 TI - The imd gene is required for local Cecropin expression in Drosophila barrier epithelia. AB - Surfaces of higher eukaryotes are normally covered with microorganisms but are usually not infected by them. Innate immunity and the expression of gene-encoded antimicrobial peptides play important roles in the first line of defence in higher animals. The immune response in Drosophila promotes systemic expression of antimicrobial peptides in response to microbial infection. We now demonstrate that the epidermal cells underlying the cuticle of larvae respond to infected wounds by local expression of the genes for the antimicrobial peptide cecropin A. Thus, the Drosophila epidermis plays an active role in the innate defence against microorganisms. The immune deficiency (imd) gene was found to be a crucial component of the signal-induced epidermal expression in both embryos and larvae. In contrast, melanization, which is part of the wound healing process, is not dependent on the imd gene, indicating that the signalling pathways promoting melanization and antimicrobial peptide gene expression can be uncoupled. PMID- 11266368 TI - An inner membrane platform in the type II secretion machinery of Gram-negative bacteria. AB - The type II secretion machinery allows most Gram-negative bacteria to deliver virulence factors into their surroundings. We report that in Erwinia chrysanthemi, GspE (the putative NTPase), GspF, GspL and GspM constitute a complex in the inner membrane that is presumably used as a platform for assembling other parts of the secretion machinery. The GspE-GspF-GspL-GspM complex was demonstrated by two methods: (i) co-immunoprecipitation of GspE-GspF GspL with antibodies raised against either GspE or GspF; (ii) interactions in the yeast two-hybrid system between GspF and GspE, GspF and GspL, GspL and GspM. GspL was found to have an essential role in complex formation. We propose a model in which the GspE-GspF-GspL-GspM proteins constitute a building block within the secretion machinery on top of which another building block, referred to as a pseudopilus, assembles. By analogy, we predict that a similar platform is required for the biogenesis of the type IV pilus. PMID- 11266370 TI - Pump up the volume: a sound policy? PMID- 11266369 TI - Non-specific activation of the epithelial sodium channel by the CFTR chloride channel. AB - The genetic disease cystic fibrosis is caused by mutation of the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR). Controversial studies reported regulation of the epithelial sodium channel (ENaC) by CFTR. We found that uptake of (22)Na(+) through ENaC is modulated by activation of CFTR in oocytes, coexpressing CFTR and ENaC, depending on extracellular chloride concentration. Furthermore we found that the effect of CFTR activation could be mimicked by other chloride channels. Voltage- and patch-clamp measurements, however, showed neither stimulation nor inhibition of ENaC-mediated conductance by activated CFTR. We conclude that the observed modulation of (22)Na(+) uptake by activated CFTR is due to the effect of CFTR-mediated chloride conductance on the membrane potential. These findings argue against the notion of a specific influence of CFTR on ENaC and emphasize the chloride channel function of CFTR. PMID- 11266371 TI - Adenoma characteristics as risk factors for recurrence of advanced adenomas. AB - BACKGROUND AND AIMS: The link between adenoma characteristics at baseline colonoscopy and adenoma recurrence is poorly understood. We assessed whether the number, size, location, or histology of resected adenomas was related to the probability of recurrence of advanced lesions. METHODS: Analyses were based on 1287 men and women in the wheat bran fiber (WBF) study, a randomized, double blind trial of WBF as a means of decreasing the probability of adenoma recurrence over a period of 3 years. Multiple logistic regression was used to calculate odds ratios (ORs) and 95% confidence intervals (CIs). RESULTS: Recurrence of advanced adenomas (>1 cm or tubulovillous/villous histology) was higher among individuals with adenomas >1 cm compared with those with adenomas <0.5 cm (OR, 2.69; 95% CI, 1.34-5.42) and among those with proximal than those with distal adenomas (OR, 1.65; 95% CI, 1.02-2.67). No association was observed for adenoma number or histology. A shift in location from the distal colon and rectum at baseline (54.6%) to more proximal recurrent adenomas (45.2%), including advanced lesions (42.8%), was observed. CONCLUSIONS: Large or proximally located adenomas are important indicators of recurrence of advanced lesions. Because most recurrences were detected in the proximal colon, careful surveillance of this area is warranted. PMID- 11266372 TI - Lack of prognostic influence of circulating tumor cells in peripheral blood of patients with colorectal cancer. AB - BACKGROUND AND AIMS: Circulating tumor cells in peripheral blood may be detected using high-sensitivity molecular techniques in several types of solid neoplasms, but their significance in colorectal cancer is controversial. The aim of this study was to assess the prognostic value of carcinoembryonic antigen (CEA) messenger RNA (mRNA) detection in peripheral blood samples from patients with colorectal cancer. METHODS: Peripheral vein blood samples from 95 consecutive patients with histologically confirmed colorectal carcinoma were obtained immediately before surgery to determine the presence of circulating tumor cells by use of a reverse-transcription polymerase chain reaction targeting CEA mRNA. Endpoints of the study were disease-free and overall survival. Results are referred to the whole series and, more importantly, to the 68 patients who underwent surgery for cure. RESULTS: After a median follow-up of 42 months, 19 of 68 patients (28%) operated on for cure had tumor relapse. In addition, 50 of 68 patients (73%) were alive. The probability of disease-free and overall survival was dependent on lymph node metastases and degree of differentiation, but not on the presence of circulating tumor cells (disease-free survival: relative risk, 1.00; 95% confidence interval [CI], 0.39-2.22, P = 0.99; overall survival: relative risk, 0.91, 95% CI, 0.34-2.43; P = 0.84). Similar results were obtained when all 95 patients with colorectal cancer were analyzed (disease-free survival: relative risk, 1.11; 95% CI, 0.63-1.95; P = 0.71; overall survival: relative risk, 1.21; 95% CI, 0.63-2.30, P = 0.55). CONCLUSIONS: Preoperative detection of blood circulating tumor cells by means of reverse-transcription polymerase chain reaction of CEA does not have prognostic significance in patients with colorectal cancer. PMID- 11266373 TI - Smoking cessation and the course of Crohn's disease: an intervention study. AB - BACKGROUND AND AIMS: To evaluate the benefit of smoking cessation in individuals with Crohn's disease, we performed an intervention study in a large cohort of smokers with the disease. METHODS: Repeated counseling to stop smoking, with easy access to a smoking cessation program, was given to 474 consecutive smokers with Crohn's disease. Patients who stopped smoking for more than 1 year (quitters) were included in a prospective follow-up study, which compared disease course and therapeutic needs with 2 control groups, continuing smokers and nonsmokers, paired for age, gender, disease location, and activity. RESULTS: There were 59 quitters (12%). Predictors of quitting were the physician, previous intestinal surgery, high socioeconomic status, and in women, oral contraceptive use. During a median follow-up of 29 months (1-54 months), the risk of flare-up in quitters did not differ from that in nonsmokers and was less than in continuing smokers (P < 0.001). Need for steroids and for introduction or reinforcement of immunosuppressive therapy, respectively, were similar in quitters and nonsmokers and increased in continuing smokers. The risk of surgery was not significantly different in the 3 groups. CONCLUSIONS: Patients with Crohn's disease who stop smoking for more than 1 year have a more benign disease course than if they had never smoked. PMID- 11266374 TI - Fat digestion modulates gastrointestinal sensations induced by gastric distention and duodenal lipid in humans. AB - BACKGROUND AND AIMS: It is unclear whether fat digestion is required for the induction of gastrointestinal sensations and whether different fats have different effects. We investigated the effect of fat digestion and of medium chain triglycerides (MCTs; C < 12) and long-chain triglycerides (LCTs; C > 16) on gastrointestinal sensations. METHODS: In a double-blind study, 15 healthy subjects were studied on 5 occasions during which LCT or MCT emulsions (2 kcal/min), with or without 120 mg tetrahydrolipstatin (THL, lipase inhibitor), or sucrose polyester (SPE, nondigestible fat) were infused intraduodenally in randomized order. After 30 minutes, the proximal stomach was distended in 1 mm Hg steps/min. Intensity of gastrointestinal sensations (on a 0-10 visual analog scale), plasma cholecystokinin (CCK) levels, and gastric volumes were assessed throughout. RESULTS: LCT and MCT increased gastric volume at baseline pressure compared with SPE, and LCT more than MCT. THL entirely abolished this effect (volumes [mL]: LCT, 213 +/- 19; LCT-THL, 39 +/- 3; MCT, 155 +/- 12; MCT-THL, 43 +/- 5; SPE, 44 +/- 5). Only LCT increased plasma CCK levels (pmol/L per 30 minutes: LCT, 21 +/- 2; LCT-THL, 9 +/- 1; MCT, 9 +/- 1; MCT-THL, 11 +/- 1; SPE, 9 +/- 1). During distentions, intragastric volumes were greater during infusion of LCT and MCT than during the respective THL conditions or SPE, but plasma CCK levels did not change. The intensity of sensations increased (hunger decreased) more with LCT than with MCT. During infusion of THL or SPE, the effects were smaller than during LCT or MCT. CONCLUSIONS: Fat digestion is required for the modulation of gastrointestinal sensations during gastric distention. The effects of fat depend on the fatty acid chain length and are not entirely explained by release of CCK. PMID- 11266375 TI - Complement activation directly induced by Helicobacter pylori. AB - BACKGROUND AND AIMS: Helicobacter pylori is a frequent gram-negative colonizer of the human stomach. Its interaction with complement may be involved in the pathogenesis of chronic gastritis, and was mechanistically studied in vitro. METHODS: Four H. pylori strains, 2 cytotoxin-associated genes (cag)A+ and 2 cagA , were isolated from infected patients. Bacteria or purified H. pylori lipopolysaccharides (LPSs) were incubated with nonimmune serum at 37 degrees C; the activation products C3b/iC3b/C3c (C3bc) and terminal complement complex (TCC) were then quantified by immunoassays. The serum sensitivity of 1 strain (L01, cagA+) was tested by counting the numbers of colony-forming units. RESULTS: All strains and LPSs generated large amounts of C3bc and TCC. Blocking of the classic complement pathway by the calcium chelator ethylene glycol tetraacetic acid (EGTA) markedly reduced the complement products, suggesting that H. pylori and its LPSs directly engage the classic activation pathway. H. pylori was shown to be serum sensitive, but 30% or more nonimmune serum was necessary to induce marked killing. After 5 minutes, swelled bacteria coated with C3bc and TCC were shown. CONCLUSIONS: H. pylori is complement sensitive and activates the classic pathway even in the absence of specific antibodies. Released cell wall constituents such as LPSs can activate complement and may explain why this bacterium induces gastric pathology without invading the mucosa. PMID- 11266376 TI - Regulatory role of phosphatidylinositol 3-kinase on TNF-alpha-induced cyclooxygenase 2 expression in colonic epithelial cells. AB - BACKGROUND AND AIMS: Cyclooxygenase (COX)-2 is up-regulated in most colonic cancers and in inflammatory bowel disease in which tumor necrosis factor (TNF) alpha is believed to play a central role. There has been recent speculation on the activation of phosphatidylinositol 3-kinase (PI 3-kinase) by TNF-alpha and its role in the regulation of genes controlled by NF-kappaB. We investigated the regulatory role of PI 3-kinase on COX-2 expression in colonic epithelial cells. METHODS: In HT-29 and Caco-2 colonic epithelial cells, COX-2 expression was induced by either TNF-alpha or interleukin (IL)-1alpha as observed by Northern and Western analyses. COX-2 activity was assessed by measuring prostaglandin E(2) (PGE2) production by enzyme-linked immunosorbent assay. NF-kappaB binding activity was assessed by electrophoretic mobility shift assay. PI 3-kinase activity was measured by quantifying the accumulation of PI 3-kinase-dependent D 3 lipid products by high-performance liquid chromatography. RESULTS: The PI 3 kinase inhibitor wortmannin up-regulated induced COX-2 expression in a concentration-dependent manner in both HT-29 and Caco-2 cells. An alternative PI 3-kinase inhibitor, LY294002, caused up-regulation of induced COX-2 messenger RNA (mRNA) in HT-29 cells at concentrations of < or =1 micromol/L. IL-4 and IL-13, which are known to activate PI 3-kinase, down-regulated HT-29 COX-2 mRNA, protein, and PGE2 production. NF-kappaB binding activity was unaltered by PI 3 kinase inhibition in HT-29 cells, in which TNF-alpha was shown to activate PI 3 kinase directly. CONCLUSIONS: COX-2 is negatively regulated by PI 3-kinase; we propose that the inhibitory effect of IL-4 and IL-13 is mediated via a PI 3 kinase-dependent pathway. This mechanism does not appear to involve NF-kappaB because PI 3-kinase inhibition did not alter NF-kappaB binding activity. TNF alpha can activate PI 3-kinase directly in addition to inducing COX-2. PMID- 11266377 TI - Mechanism of action of the calcium-sensing receptor in human antral gastrin cells. AB - BACKGROUND AND AIMS: Human G cells express the calcium-sensing receptor and respond to extracellular calcium by releasing gastrin. However, the receptor on G cells is insensitive to serum calcium levels. We investigated whether this is a result of differential regulation of signaling pathways compared with parathyroid or calcitonin cells. METHODS: Gastrin release from primary cultures of human antral epithelial cells enriched for G cells (35%) was measured by radioimmunoassay. G cells were stimulated by increasing extracellular calcium concentration for 1 hour in the presence or absence of antagonists of specific intracellular signaling pathways. Intracellular calcium levels were monitored to evaluate the effect of the antagonists on calcium influx. RESULTS: Inhibition of phospholipase C decreased calcium-stimulated gastrin release, but blockers of adenylate cyclase, phospholipase A(2), or mitogen-activated protein kinase had no effect. Inhibition of protein kinase C, nonselective cation channels, and phosphodiesterase increased basal and calcium-stimulated gastrin release while decreasing calcium influx. These data were consistent with basally active phosphodiesterase. CONCLUSIONS: The calcium-sensing receptor on the G cell activates phospholipase C and opens nonselective cation channels, resulting in an influx of extracellular calcium. Protein kinase C isozymes expressed by the G cells play multiple roles regulating both gastrin secretion and phosphodiesterase activity. PMID- 11266378 TI - Immunohistochemical demonstration of the NK(1) tachykinin receptor on muscle and epithelia in guinea pig intestine. AB - BACKGROUND AND AIMS: Previous immunohistochemical studies failed to reveal neurokinin (NK)(1) tachykinin receptors on intestinal muscle, despite convincing pharmacologic data indicating their presence. This study aimed to apply optimal immunohistochemical methods to reveal the receptors. METHODS: NK(1)-receptor immunoreactivity was examined by confocal microscopy in tissue incubated with or without 10(-7) mol/L substance P (SP), 10(-7) mol/L SP plus 10(-6) mol/L NK(1) receptor antagonist (CP99994), or with fluorescent cyanine 3.18 (Cy3) SP. RESULTS: Without incubation, NK(1)-receptor immunoreactivity was strong on muscle of the rectum and distal colon and weak in proximal colon and small intestine. NK(1) receptor was located on the surface of muscle cells in all gut regions. Exposure to SP increased the intensity of immunoreactivity, and the receptor moved into the cytoplasm. Mobilization of the receptor by SP was blocked by the NK(1)-receptor antagonist CP99994. Cy3-SP was internalized by muscle cells and colocalized with the receptor. NK(1)-receptor immunoreactivity occurred on crypt epithelial cells in the small intestine and the base of glands in the proximal colon. CONCLUSIONS: The NK(1) receptor occurs on the external muscle throughout the small and large intestines. SP binds and triggers NK(1)-receptor aggregation and internalization in the muscle. PMID- 11266379 TI - In vivo absorption of medium-chain fatty acids by the rat colon exceeds that of short-chain fatty acids. AB - BACKGROUND AND AIMS: Short-chain fatty acids (SCFAs) are main fuels of the colonic epithelium, and are avidly absorbed by the colon of animal and man. The current knowledge on colonic metabolism and absorption of medium-chain fatty acids (MCFAs) is limited. In some clinical situations, colonic absorption of high energy substances could compensate for reduced absorptive capacity because of a shortened or malfunctioning small bowel. We evaluated and compared colonic absorption and metabolism of MCFAs (octanoate, decanoate, and dodecanoate), SCFAs (acetate and butyrate), and long-chain fatty acids (LCFAs) (oleate). METHODS: Rats were surgically operated on to cannulate a 7-cm segment of proximal colon, isolate the vasculature, and cannulate the right colic vein draining this segment. The lumen was perfused with (14)C-labeled substrates for 100 minutes. Right colic vein blood was analyzed for total (14)C, (14)CO(2), and metabolites by scintillation counting and high-performance liquid chromatography. RESULTS: The transport from the colonic lumen to mesenteric blood of substrate carbon from MCFAs exceeded by 2-13-fold that of SCFAs and LCFAs. The CO(2) production from the oxidation of MCFAs was as high as or higher than that from SCFAs. CO(2) produced from the LCFA, oleate, was lower than from SCFAs or MCFAs. In addition to CO(2), ketone bodies were major metabolites of SCFAs and MCFAs. Ketogenesis from butyrate and the MCFAs was significantly higher than from acetate and oleate. A substantial proportion (50%-90%) of all substrates was absorbed without being metabolized. CONCLUSIONS: The colonic epithelium serves to absorb and partially metabolize MCFAs. For patients with a compromised small-bowel function, colonic absorption of MCFAs could represent an important way of receiving calories. PMID- 11266380 TI - Differential role of selectins in experimental colitis. AB - BACKGROUND AND AIMS: The role of selectins in experimental colitis remains unknown. The aims of this study were to characterize the time-course expression of selectins in trinitrobenzene sulfonic acid (TNBS)-induced colitis, the functional role of selectins in colonic leukocyte-endothelial cell interactions, and the therapeutic usefulness of selectin blockade in this model. METHODS: Control and TNBS-induced colitic rats were studied. Expression of P- and E selectin was assessed by the radiolabeled antibody technique, and L-selectin by flow cytometry. Leukocyte-endothelial cell interactions were studied in colonic venules by using intravital microscopy under basal conditions and after P-, E-, or L-selectin immunoblockade. Additional groups of animals were treated with anti P-selectin antibody, a nonbinding antibody, or dexamethasone, for 7 days. RESULTS: P-selectin and E-selectin expression were markedly up-regulated in colitic rats. Increased leukocyte rolling was abrogated by anti-P-selectin, but only attenuated by anti-E- or anti-L-selectin antibodies. Only pretreatment with anti-P- selectin decreased leukocyte adhesion. Animals chronically treated with dexamethasone, but not with anti- P-selectin, had significantly lower macroscopic and histologic damage scores, colon weight, and myeloperoxidase (MPO) activity than those treated with nonbinding antibody. CONCLUSIONS: P-selectin plays a key role on leukocyte rolling and its blockade attenuates leukocyte adhesion in TNBS induced colitis. However, treatment with an anti-P-selectin antibody does not significantly improve colitis. PMID- 11266381 TI - Effect of mature lymphocytes and lymphotoxin on the development of the follicle associated epithelium and M cells in mouse Peyer's patches. AB - BACKGROUND AND AIMS: Mechanisms regulating M-cell formation are still poorly understood. In vitro studies showed that lymphocytes trigger the conversion of enterocyte cell lines into M cell-like cells on coculture, whereas in vivo their role in M cell differentiation is still elusive. Our aim was first to examine Rag 1-/- mice, lacking B and T lymphocytes, for the presence of intestinal M cells. Second, we investigated the role of lymphotoxin alphabeta signaling on M-cell formation, given its pivotal role in the development of mouse Peyer's patches. METHODS: Small intestines of Rag-1-/- mice, injected or not with soluble lymphotoxin beta receptor-immunoglobulin fusion protein, were analyzed morphologically using whole mount cytochemical staining, immunohistochemistry, and electron microscopy. RESULTS: Small Peyer's patch-like aggregates were found in Rag-1-/- mice in normal number and location. The overlying epithelium of such aggregates was reduced in size but still harbored M cells. In vivo neutralization of lymphotoxin beta-receptor signaling partially reduced the percentage of M cells. CONCLUSIONS: The absence of mature lymphocytes does not prevent the formation of M cells, indicating that the signaling molecules that support M-cell differentiation, such as lymphotoxin alphabeta, may also be supplied by non-B and non-T cells. Mature B lymphocytes, however, are required for the formation of a full-sized follicle-associated epithelium. PMID- 11266383 TI - Compensatory phospholipid digestion is required for cholesterol absorption in pancreatic phospholipase A(2)-deficient mice. AB - BACKGROUND AND AIMS: Numerous studies have suggested phospholipid inhibition of dietary cholesterol absorption through the gastrointestinal tract. This study addressed the importance of luminal phospholipid hydrolysis in this process. METHODS: The effect of phospholipase inhibition on cholesterol transport from intestinal lumen to the lymphatics was evaluated in lymph fistula rats. Cholesterol and phospholipid absorption efficiency in intact animals was evaluated in control and phospholipase A(2) (PLA2) gene-targeted mice. RESULTS: The PLA2 inhibitor FPL 67047XX retarded cholesterol absorption in a lymph fistula rat model. Under basal chow-fed dietary conditions, cholesterol absorption efficiency from a single bolus meal, and plasma lipid levels, were similar among PLA2+/+, PLA2+/-, and PLA2-/- mice. Interestingly, the nonhydrolyzable phospholipid dioleoyl ether phosphatidylcholine suppressed cholesterol absorption by 10% to 18% in mice without regard to their PLA2 genotype. When 1-palmitoyl-2 [(14)C]oleoyl-phosphatidylcholine was used as the substrate, the radiolabeled phospholipid was found to be hydrolyzed and absorbed with equal efficiency in PLA2+/+, PLA2+/-, and PLA2-/- mice. CONCLUSIONS: These results suggested that although phospholipid digestion in the intestinal lumen is a prerequisite for efficient cholesterol absorption, additional enzyme(s) can compensate for pancreatic PLA2 in catalyzing phospholipid digestion and facilitating cholesterol absorption in PLA2 knockout mice. PMID- 11266382 TI - Nonalcoholic steatohepatitis: association of insulin resistance and mitochondrial abnormalities. AB - BACKGROUND AND AIMS: The pathogenesis of nonalcoholic steatohepatitis (NASH) is unknown. We tested the hypothesis that NASH is associated with 2 defects: (1) peripheral insulin resistance, which increases lipolysis, delivery of free fatty acids (FFA) to the liver, and hepatic fatty acid beta oxidation, thereby creating oxidative stress; and (2) an abnormality within the hepatocytes that might render them more susceptible to injury from oxidative stress. METHODS: The hypothesis was tested by evaluation of (1) insulin resistance by a 2-step hyperinsulinemic (10 and 40 mU. m(-2). min(-1)) euglycemic clamp; (2) insulin effects on lipolysis by enrichment of [U-(13)C]glycerol; (3) frequency and severity of structural defects in hepatocyte mitochondria in vivo; (4) fatty acid beta oxidation from serum [beta-OH butyrate], release of water-soluble radioactivity from (3)H palmitate by cultured fibroblasts and urinary dicarboxylic acid excretion; and (5) hepatic lipid peroxidation by immunohistochemical staining for 3 nitrotyrosine (3-NT). Subjects with NASH (n = 6-10 for different studies) were compared with those with fatty liver (n = 6) or normal controls (n = 6). RESULTS: NASH and fatty liver were both associated with insulin resistance, with mean glucose infusion rates (normal/fatty liver/NASH) of step 1, 4.5/1.6/0.9; step 2, 9.5/7.7/4.5 (P < 0.03 for both steps). Although baseline rates of glycerol appearance were higher in those with NASH than in those with fatty liver (means, 14.6 vs. 21.6 micromol. kg(-1). min(-1); P < 0.05), neither group significantly suppressed glycerol appearance at insulin infusion rates of 10 mU. m(-2). min( 1). NASH was associated with loss of mitochondrial cristae and paracrystalline inclusions in 9 of 10 subjects, compared with 0 of 6 subjects with fatty liver. However, no evidence of a generalized defect in fatty acid beta oxidation was noted in any group. Also, mean [beta-OH butyrate] was highest in those with NASH (means, 90 vs. 110 vs. 160 micromol/L; P < 0.04). Increased staining for 3-NT was present in fatty liver, and even greater staining was seen in NASH. CONCLUSIONS: These data indicate that peripheral insulin resistance, increased fatty acid beta oxidation, and hepatic oxidative stress are present in both fatty liver and NASH, but NASH alone is associated with mitochondrial structural defects. PMID- 11266384 TI - ATP-binding cassette transporter A1 (ABCA1) affects total body sterol metabolism. AB - BACKGROUND AND AIMS: Members of the family of ABC transporters are involved in different processes of sterol metabolism, and ABCA1 was recently identified as a key regulator of high-density lipoprotein (HDL) metabolism. Our aim was to further analyze the role of ABCA1 in cholesterol metabolism. METHODS: ABCA1 deficient mice (ABCA1-/-) and wild-type mice were compared for different aspects of sterol metabolism. Intestinal cholesterol absorption was determined by a dual stable isotope technique, and analysis of fecal, plasma, and tissue sterols was performed by gas chromatography/mass spectrometry. Key regulators of sterol metabolism were investigated by Northern and Western blot analyses or enzyme activity assays. RESULTS: ABCA1-disrupted sv129/C57BL/6 hybrid mice showed a significant reduction in intestinal cholesterol absorption. The decrease in cholesterol absorption was followed by an enhanced fecal loss of neutral sterols, whereas fecal bile acid excretion was not affected. Total body cholesterol synthesis was significantly increased, with enhanced 3-hydroxy-3-methyglutaryl coenzyme A (HMG-CoA) reductase observed in adrenals and spleen. In addition, ABCA1-/- mice showed markedly increased concentrations of cholesterol precursors in the plasma, lung, intestine, and feces. Reduced HMG-CoA reductase messenger RNA and enzyme activity in the liver suggest that enhanced cholesterol synthesis in ABCA1-/- mice occurs in peripheral tissues rather than the liver. CONCLUSIONS: The metabolism of cholesterol and cholesterol precursors is markedly affected by a lack of ABCA1 function. PMID- 11266385 TI - Expression of the peripheral-type benzodiazepine receptor and apoptosis induction in hepatic stellate cells. AB - BACKGROUND AND AIMS: Hepatic stellate cell (HSC) transformation and proliferation play an important role in liver fibrogenesis, and HSC apoptosis may be involved in the termination of this response. METHODS: Expression of the peripheral benzodiazepine receptor (PBR) and PBR-ligand-induced apoptosis were studied in cultured rat liver HSC. RESULTS: Transformation of HSC led to a transient expression of PBR at the messenger RNA and protein level, which was maximal after about 3 and 7 days of culture, respectively, and declined thereafter. Immunoreactive PBR showed a punctate staining and colocalized with mitochondrial manganese-dependent superoxide dismutase and adenine nucleotide translocator 1. The selective PBR ligands 1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3 isoquinolinecarboxamide (PK11195) and 4' chlorodiazepam (Ro5-4864), but not the centrally acting benzodiazepine ligand clonazepam, induced dose-dependent apoptosis in HSC. The apoptotic potency of PK11195 paralleled the level of PBR expression. PK11195 induced dephosphorylation of protein kinase B/Akt and Bad and a downregulation of Bcl-2. Collapse of the mitochondrial membrane potential preceeded PBR-ligand-induced apoptosis. No apoptosis was induced by PK11195 in parenchymal cells, despite the presence of PBR, and PK11195 had no effect in these cells on Bad phosphorylation and Bcl-2 expression. CONCLUSIONS: Transformation of HSC leads to a transient expression of PBR and renders the cells sensitive to PBR-ligand-induced apoptosis, involving protein kinase B/Akt and Bad-dependent mechanisms. PMID- 11266386 TI - Carbon monoxide from heme catabolism protects against hepatobiliary dysfunction in endotoxin-treated rat liver. AB - BACKGROUND AND AIMS: Liver is a major organ for heme detoxification under disease conditions, but its self-protective mechanisms against the toxicity are unknown. This study aimed to examine roles of carbon monoxide (CO), the gaseous product of heme oxygenase (HO), in ameliorating hepatobiliary dysfunction during catabolism of heme molecules in endotoxemic livers. METHODS: Vascular resistance and biliary flux of bilirubin-IXalpha, an index of HO-derived CO generation, were monitored in perfused livers of endotoxemic rats. Livers were perfused with HbO(2), which captures nitric oxide (NO) and CO, or metHb, a reagent trapping NO but not CO. RESULTS: In endotoxin-pretreated livers where inducible NO synthase and HO-1 overproduced NO and CO, HbO(2) caused marked vasoconstriction and cholestasis. These changes were not reproduced by the NO synthase inhibitor aminoguanidine alone, but by coadministration of zinc protoporphyrin-IX, an HO inhibitor. CO supplementation attenuated the events caused by aminoguanidine plus zinc protoporphyrin-IX, suggesting that simultaneous elimination of these vasorelaxing gases accounts for a mechanism for HbO(2)-induced changes. This concept was supported by observation that metHb did not cause any cholestasis; the reagent captures NO but triggers CO overproduction through rapid degradation of the heme by HO-1. CONCLUSIONS: These results suggest protective roles of CO against hepatobiliary dysfunction caused by heme overloading under stress conditions. PMID- 11266387 TI - Delivery of the Cu/Zn-superoxide dismutase gene with adenovirus reduces early alcohol-induced liver injury in rats. AB - BACKGROUND AND AIMS: Alcohol-induced liver injury is associated with an increase in oxidants from a variety of possible sources. Therefore, it was hypothesized that increased and stable expression of the antioxidant enzyme Cu/Zn-superoxide dismutase (SOD1) would diminish oxygen free radicals and reduce alcohol-induced liver injury. METHODS: To test this hypothesis, rats were given recombinant adenovirus containing Cu/Zn-superoxide dismutase (Ad.SOD1) or beta-galactosidase (Ad.lacZ) and fed ethanol enterally for 3 weeks. RESULTS: SOD was increased significantly 3-5-fold over endogenous levels in both hepatocytes as well as Kupffer cells 3 weeks after infection. Serum transaminase levels and pathology were elevated significantly in Ad.lacZ-treated animals by using an intragastric feeding model. This effect was blunted significantly in Ad.SOD1-infected animals. Importantly, electron spin resonance-detectable free-radical adducts caused by ethanol were also decreased by SOD1 overexpression. Moreover, the increase in nuclear factor kappaB (NFkappaB), tumor necrosis factor alpha (TNF-alpha), and interleukin 1 messenger RNA (mRNA) caused by ethanol was blunted in animals treated with Ad.SOD1. CONCLUSIONS: These data support the hypothesis that oxidant production is critical in early alcohol-induced liver injury and that gene delivery of antioxidant enzymes may be useful in prevention and treatment. PMID- 11266389 TI - Prevention of pancreatic cancer induction in hamsters by metformin. AB - BACKGROUND AND AIMS: Our previous study suggested that the known promotional effect of a high fat diet, which in hamsters induces peripheral insulin resistance, is related to a compensatory proliferation of islet cells. The present study was to examine whether the prevention of islet cell proliferation can inhibit the promotional effect of a high-fat diet in pancreatic carcinogenesis. METHODS: Two groups of high fat-fed hamsters were used. One group received Metformin in drinking water for life (HF+Met group), and the other group served as a control (HF group). At the time when the normalization of the plasma insulin level was expected, all hamsters were treated with the pancreatic carcinogen, N-nitrosobis-(2-oxopropyl)amine, and the experiment was terminated 42 weeks later. RESULTS: Although 50% of the hamsters in the high-fat group developed malignant lesions, none was found in the HF+Met group (P < 0.05). Also, significantly more hyperplastic and premalignant lesions, most of which were found within the islets, were detected in the high-fat group (8.6 lesions/hamster) than in the HF+Met group (1.8 lesions/hamster). CONCLUSIONS: The results lend further support on the significant role of islet cells in pancreatic carcinogenesis and may explain the association between pancreatic cancer and obesity, which is usually associated with peripheral insulin resistance. PMID- 11266388 TI - NF-kappaB stimulates inducible nitric oxide synthase to protect mouse hepatocytes from TNF-alpha- and Fas-mediated apoptosis. AB - BACKGROUND AND AIMS: Hepatocyte apoptosis is induced by tumor necrosis factor alpha (TNF-alpha) and Fas ligand. Although nuclear factor-kappaB (NF-kappaB) activation protects hepatocytes from TNF-alpha-mediated apoptosis, the NF-kappaB responsive genes that protect hepatocytes are unknown. Our aim was to study the role of NF-kappaB activation and inducible nitric oxide synthases (iNOSs) in TNF alpha- and Fas-mediated apoptosis in hepatocytes. METHODS: Primary cultures of hepatocytes from wild-type and iNOS knockout mice were treated with TNF-alpha, the Fas agonistic antibody Jo2, a nitric oxide (NO) donor (S-nitroso-N acetylpenicillamine), an NO inhibitor (N(G)-methyl-L-arginine acetate), and/or adenovirus-expressing NF-kappaB inhibitors. RESULTS: The IkappaB superrepressor and a dominant-negative form of IkappaB kinase beta (IKKbeta) inhibited NF-kappaB binding activity by TNF-alpha or Jo2 and sensitized hepatocytes to TNF-alpha- and Jo2-mediated apoptosis. TNF-alpha and Jo2 induced iNOS messenger RNA and protein levels through the induction of NF-kappaB. S-nitroso-N-acetylpenicillamine inhibited Bid cleavage, the mitochondrial permeability transition, cytochrome c release, and caspase-8 and -3 activity, and reduced TNF-alpha- and Fas-mediated death in hepatocytes expressing IkappaB superrepressor. N(G)-methyl-L-arginine acetate partially sensitized hepatocytes to TNF-alpha- and Fas-mediated cell killing. TNF-alpha alone or Jo2 alone induced moderate cell death in hepatocytes from iNOS(-)/(-) mice. CONCLUSIONS: NO protects hepatocytes from TNF-alpha- and Fas-mediated apoptosis. Endogenous iNOS, which is activated by NF-kappaB via IKKbeta, provides partial protection from apoptosis. PMID- 11266391 TI - Transepithelial signaling: making sense of stomach contents. PMID- 11266390 TI - A new cause of Zollinger-Ellison syndrome: non-small cell lung cancer. AB - Numerous epidemiologic studies suggest a relationship between lung cancer and peptic ulcer disease. Furthermore, various lung cancers synthesize and release a number of peptides such as gastrin and gastrin-releasing peptide that could cause acid hypersecretion; however, Zollinger-Ellison syndrome (ZES), because of a lung tumor, has never been described. We report such a patient for the first time. A 60-year-old man with a non-small cell lung carcinoma (large cell type) presented with diarrhea, heartburn, abdominal pain, and duodenal ulcers. Evaluation showed ZES was present (fasting hypergastrinemia, hyperchlorhydria) and control of all symptoms by omeprazole. No abdominal or cardiac tumor, the other known locations of gastrinomas causing ZES, was found on detailed tumor imaging studies. Resection of the lung tumor resulted in a decrease in gastrin levels to normal values. Plasma radioimmunoassays showed elevated gastrin, chromogranin A and normal levels of gastrin-releasing peptide, and 9 other hormones. The tumor showed similar immunocytochemical results. The characteristics of this case are compared with 100 cases of sporadic abdominal gastrinomas, and the evidence reviewed suggests why ZES should be considered in patients with lung cancer with peptic symptoms. PMID- 11266392 TI - Insulin resistance and mitochondrial abnormalities in NASH: a cool look into a burning issue. PMID- 11266393 TI - The up-and-down of hepatic stellate cells in tissue injury: apoptosis restores cellular homeostasis. PMID- 11266394 TI - Carbon monoxide and sepsis: is a toxic gas good for your liver? PMID- 11266395 TI - Sugar pills for pancreatic cancer: the benefits of becoming (insulin) sensitive. PMID- 11266396 TI - Turbo probiotics for IBD. PMID- 11266398 TI - Single-stage operations for Hirschsprung's disease: pushing the envelope. PMID- 11266397 TI - Helicobacter eradication versus prompt endoscopy for dyspepsia. PMID- 11266399 TI - The prevention of hepatocellular carcinoma by interferon: the story continues. PMID- 11266402 TI - Schizophrenia in late life: findings challenge traditional concepts. AB - Two notions about schizophrenia have persisted: (1) it is characterized by onset in adolescence or early adulthood, and (2) it has a progressively deteriorating course. Recent studies focusing on early-adulthood- and middle-age-onset schizophrenia challenge these views. Patients with early-onset schizophrenia and middle-age-onset schizophrenia (MAOS) are similar in terms of family history of schizophrenia, presence of minor physical anomalies, early childhood maladjustment, severity of positive symptoms, presence of gross structural abnormalities on cerebral magnetic resonance imaging, overall pattern of neuropsychologic deficits, and qualitative response to neuroleptic medications. Differences include a higher proportion of women among MAOS patients, and the tendency for MAOS patients to have less severe negative symptoms, better neuropsychologic performance (particularly in learning and abstraction/cognitive flexibility), and possibly larger thalamic volume and to respond to lower doses of neuroleptic medications. While onset of schizophrenia-like symptoms in very late life may reflect an acquired condition that is not "true schizophrenia," and that may be labeled "very-late-onset schizophrenia-like psychosis," findings suggest that true schizophrenia can arise after early adulthood. Middle-age-onset schizophrenia is predominantly neurodevelopmental, but it is also a distinct neurobiological subtype of schizophrenia. Our studies also demonstrate that neuropsychologic functioning remains stable in chronic schizophrenia outpatients, even when observed over several years and in the presence of significant fluctuations in the severity of clinical symptoms. PMID- 11266403 TI - Cognitive and functional impairments in elderly patients with schizophrenia: a review of the recent literature. AB - Although cognitive and functional impairments are very common in patients with schizophrenia, little is known about the characteristics of these impairments in older patients with the illness. Even less is known about the longitudinal course of these impairments. This article reviews the current knowledge with regard to cognitive and functional aspects of schizophrenia in late life, with a focus on the course of cognitive and functional deficits. Although the common belief has been that schizophrenia is static over the lifespan, recent evidence suggests that some patients may experience declines in their functioning over time. The predictors and course of these declines, as well as their potential underlying causes, are described in detail. Evidence regarding schizophrenia as a lifelong, dynamic brain disease is also reviewed. PMID- 11266405 TI - Neuroleptic treatment of late-life schizophrenia. AB - There is a paucity of studies on the use of neuroleptic medication for treatment of the core symptoms of schizophrenia in elderly patients. The studies that are available have significant methodologic problems, including the mixing of early- and late-onset patients, inadequate outcome criteria, and the lack of control groups. Studies of conventional neuroleptics suggest that older patients have a moderate therapeutic response but are likely to develop side effects. The few studies of atypical antipsychotics now available suggest efficacy for treatment of behavioral disturbance in the elderly and a more favorable side-effect profile. The usefulness of all neuroleptics for the treatment of the core symptoms of late-life schizophrenia may depend on the duration and severity of the symptoms, with a poor response associated with greater severity and duration. It appears that patients with later-onset symptoms may respond better to all neuroleptics. PMID- 11266404 TI - Contributions of neuropathology to understanding schizophrenia in late life. AB - The neurobiological basis of cognitive and functional deterioration commonly observed in elderly persons with schizophrenia is unclear. Despite superficial similarities in the clinical and neuropsychological profiles of schizophrenia in late life with neurodegenerative dementias, extensive neuropathological investigations have failed to find any evidence of neurodegeneration or neural injury beyond what is typically observed in brains of individuals without neuropsychiatric illness. In contrast, growing neuropathological data indicate aberrant brain development and connectivity in schizophrenia (including abnormalities in cytoarchitecture, innervation, and synaptic integrity) and abnormal molecular signaling pathways important in the formation of the nervous system and ongoing plasticity in maturity. These developmental abnormalities may represent a state of decreased cerebral reserve that causes persons with schizophrenia to be more vulnerable to the toxic effects of even "normal" accumulations of age-related neurodegenerative lesions. PMID- 11266406 TI - Conclusions: late-life schizophrenia. PMID- 11266407 TI - Treatment comparisons in HIV infection: the benefits and limitations of observational cohort studies. PMID- 11266408 TI - Studies of the operator region of the Staphylococcus aureus beta-lactamase operon. AB - The repressor proteins BlaI and MecI bind similarly to the bla operator implicated in the regulation of beta-lactamase synthesis in Staphylococcus aureus. BlaI binds to two separate dyads but neither copper-phenanthroline footprinting nor dimethyl sulphate (DMS) methylation protection assays produced any evidence of a change in the geometry of the DNA between the two dyads. It is concluded that BlaI molecules bound at the dyads probably do not cause bending or looping of the intervening DNA. DMS protection assays of BlaI binding to the bla operator in vitro and in vivo gave similar results so that it is tentatively concluded that the in vitro results are an accurate reflection of the in vivo situation. Deletion of the dyad nearest to the blaZ gene resulted in decreased synthesis of the chloramphenicol acetyltransferase reporter protein synthesized from the blaZ promoter/translation initiator. Explanations for this are considered. PMID- 11266409 TI - Application of a spectrophotometric method for the determination of post antibiotic effect and comparison with viable counts in agar. AB - The post-antibiotic effects of gentamicin and ciprofloxacin at 1x, 2x and 4x MIC on Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853 and Staphylococcus aureus ATCC 29213 were studied using a spectrophotometric method and the classic method of viable counts on agar as a reference. Monitoring of the growth kinetics was carried out by viability counting on the plate every hour and by means of the optical density of the cultures measured by spectrophotometry at a wavelength of 450 nm. No statistically significant differences were found between the results obtained with the spectrophotometric method and the reference method. The former method was much quicker, much easier to use and to replicate. PMID- 11266410 TI - A modified population analysis profile (PAP) method to detect hetero-resistance to vancomycin in Staphylococcus aureus in a UK hospital. AB - One hundred methicillin-resistant Staphylococcus aureus (MRSA) strains, isolated between 1983 and 1999, were tested alongside the vancomycin hetero-resistant S. aureus (hVRSA) strain Mu 3, and vancomycin-resistant S. aureus (VRSA) strain Mu 50, for their resistance to vancomycin. This was achieved using the screening method described by Hiramatsu, gradient plates, agar incorporation, standard Etest, macrodilution Etest and a modified population analysis. Using Hiramatsu's screening method, 5% of the 100 MRSA were identified as VRSA and 5% identified as hVRSA, the gradient plates identified 7% hVRSA, and the standard and macrodilution Etests identified no hVRSA. Mu 3 appeared to be vancomycin susceptible using both the agar incorporation and standard Etest methods, but was classified as hVRSA using the macrodilution Etest. The modified population analysis reliably detected vancomycin hetero-resistance in Mu 3 and identified no hVRSAs within the 100 MRSA sample. PMID- 11266411 TI - Effects of lansoprazole, clarithromycin and pH gradient on uptake of [14C]amoxycillin into rat gastric tissue. AB - The effect of lansoprazole and clarithromycin on the uptake of [(14)C]amoxycillin into rat gastric tissue was investigated. After oral administration of [(14)C]amoxycillin, the levels of radioactivity in gastrointestinal tissue were two to 15 times higher than those in plasma. The level of radioactivity in glandular stomach was significantly higher when lansoprazole and [(14)C]amoxycillin were administered together. After intravenous administration of [(14)C]amoxycillin, there was less radioactivity in gastric tissue than after oral administration, and co-administration of lansoprazole and clarithromycin had no obvious effect. The gastric emptying rate of [(14)C]amoxycillin was not apparently affected by the co-administration of lansoprazole and clarithromycin. In vitro uptake of [(14)C]amoxycillin into gastric tissue depended on the pH, with uptake at pH 7.4 being four times greater than that at pH 4.0. The apparent synergic effects of lansoprazole are due to enhanced penetration of amoxycillin in gastric mucus and tissue by increasing intragastric pH and play an important role in the eradication of H. pylori. PMID- 11266412 TI - Activity of moxifloxacin against clinical isolates of Streptococcus pneumoniae from England and Wales. AB - The activity of moxifloxacin was assessed against 1269 isolates of Streptococcus pneumoniae, comprising 462 isolates referred from UK hospitals, primarily for confirmation of resistance to first line agents, and 807 isolates from an enhanced surveillance of invasive infections. Resistance rates to penicillin, erythromycin, tetracycline and chloramphenicol were 88.7% (32.6% intermediate and 56.1% fully resistant), 50, 48 and 22.7%, respectively, for the former, and 8.0 (4.5% intermediate and 3.5% fully resistant), 14.7, 9.0 and 0.6% for the latter. Ninety-four per cent of the referred isolates and 99% of the surveillance isolates were susceptible to moxifloxacin at 83 times weaker than those against the wild-type enzyme. These results suggest that mutations in the corresponding genes confer quinolone resistance. PMID- 11266419 TI - Antimycobacterial activity of 1-deaza-7,8-dihydropteridine derivatives against Mycobacterium tuberculosis and Mycobacterium avium complex in vitro. AB - Twenty-five 1-deaza-7,8-dihydropteridine derivatives were screened for antimycobacterial activity against Mycobacterium tuberculosis strain H37Ra and three Mycobacterium avium clinical isolates (serovar 1, 4 or 6). Antibacterial activity was determined with a colorimetric microdilution broth assay. Seventeen of the compounds inhibited growth in the range >1.28 to MIC for sulbactam (1.84 h) was similar to that for imipenem (2.01 h). In the endocarditis model, imipenem (t > MIC, 2.12 h) was more efficacious than sulbactam (t > MIC, 1.17 h) in bacterial clearance from vegetations. These results show the efficacy of sulbactam in infections caused by susceptible strains of A. baumannii, with an MIC up to 4 mg/L, provided that doses reach a t > MIC similar to that of imipenem. The activity of sulbactam was time dependent. PMID- 11266428 TI - Intracellular concentration of the HIV protease inhibitors indinavir and saquinavir in human endothelial cells. AB - Human vascular endothelial cells may serve as targets and a reservoir for human immunodeficiency virus type 1 (HIV-1). The antiviral activity of HIV protease inhibitors is reported to be related directly to the intracellular amount of the drug. To assess intracellular concentrations of two HIV protease inhibitors, human umbilical venous endothelial cells (HUVECs) were exposed for 3 h and 24 h to 100, 10 and 1 mg/L indinavir and saquinavir. Intracellular drug concentrations and the total drug amount in the supernatant were measured by means of high performance liquid chromatography (HPLC). Exposure of HUVECs to 10 and 1 mg/L indinavir and saquinavir resulted in undetectable intracellular drug levels in 6 x 10(5) cells/well. Incubation of cells with solutions of 100 mg/L indinavir and saquinavir led to mean intracellular concentrations of indinavir (132 +/- 56 mg/L after 3 h and 150 +/- 29 mg/L after 24 h, respectively) and of saquinavir (96 +/- 10 mg/L after 3 h and 100 +/- 5 mg/L after 24 h) that were comparable to the levels determined for the substances in the supernatant over time (P > 0.001). These data indicate that intracellular concentrations of indinavir and saquinavir correlate well with the extracellular levels. Consequently, measurements of drug concentrations in patient's plasma by HPLC are assumed to be a good means of monitoring the intracellular drug concentration. PMID- 11266429 TI - Inhibition of H(+)-ATPase-mediated proton pumping in Cryptococcus neoformans by a novel conjugated styryl ketone. AB - We investigated the in vitro susceptibility of clinical isolates of Cryptococcus neoformans to the novel conjugated styryl ketone NC1175 by broth microdilution. The MIC(90) and the MFC of NC1175 for C. neoformans were 1 and 2 mg/L, respectively. NC1175 at low concentrations (1-4 mg/L) completely inhibited the glucose-induced acidification of the external medium caused by the extrusion of intracellular protons mediated by the plasma membrane located H(+)-ATPase. These data suggest that NC1175 is a fungicidal agent for C. neoformans and its possible cellular target(s) include the H(+)-ATPase. PMID- 11266430 TI - Current status of the aadA and dfr gene cassette families. PMID- 11266431 TI - Gentamicin diffusion in Mueller-Hinton agar plates from different manufacturers. PMID- 11266433 TI - Use of the t > MIC to choose between different dosing regimens of beta-lactam antibiotics. PMID- 11266432 TI - Dietary habits and gastrointestinal colonization by antibiotic resistant microorganisms. PMID- 11266434 TI - Correlation between increased consumption of fluoroquinolones in outpatients and resistance of Escherichia coli from urinary tract infections. PMID- 11266435 TI - The TOR kinases link nutrient sensing to cell growth. AB - Rapamycin is an immunosuppressive natural product that inhibits the proliferation of T-cells in response to nutrients and growth factors. Rapamycin binds to the peptidyl-prolyl isomerase FKBP12 and forms protein-drug complexes that inhibit signal transduction by the TOR kinases. The FKBP12 and TOR proteins are conserved from fungi to humans, and in both organisms the TOR signaling pathway plays a role in nutrient sensing. In response to nitrogen sources or amino acids, TOR regulates both transcription and translation, enabling cells to appropriately respond to growth-promoting signals. Rapamycin is having a profound impact on clinical medicine and was approved as an immunosuppressant for transplant recipients in 1999. Ongoing clinical studies address new clinical applications for rapamycin as an antiproliferative drug for chemotherapy and invasive cardiology. PMID- 11266436 TI - Activation of class III ribonucleotide reductase by thioredoxin. AB - Anaerobic ribonucleotide reductase provides facultative and obligate anaerobic microorganisms with the deoxyribonucleoside triphosphates used for DNA chain elongation and repair. In Escherichia coli, the dimeric alpha2 enzyme contains, in its active form, a glycyl radical essential for the reduction of the substrate. The introduction of the glycyl radical results from the reductive cleavage of S-adenosylmethionine catalyzed by the reduced (4Fe-4S) center of a small activating protein called beta. This activation reaction has long been known to have an absolute requirement for dithiothreitol. Here, we report that thioredoxin, along with NADPH and NADPH:thioredoxin oxidoreductase, efficiently replaces dithiothreitol and reduces an unsuspected critical disulfide bond probably located on the C terminus of the alpha protein. Activation of reduced alpha protein does not require dithiothreitol or thioredoxin anymore, and activation rates are much faster than previously reported. Thus, in E. coli, thioredoxin has very different roles for class I ribonucleotide reductase where it is required for the substrate turnover and class III ribonucleotide reductase where it acts only for the activation of the enzyme. PMID- 11266437 TI - Androgen receptor interacts with the positive elongation factor P-TEFb and enhances the efficiency of transcriptional elongation. AB - Androgen receptor (AR) may communicate with the general transcription machinery on the core promoter to exert its function as a transcriptional modulator. Our previous report demonstrated that the AR interacted with transcription factor IIH (TFIIH) under physiological conditions and that overexpression of Cdk-activating kinase, the kinase moiety of TFIIH, enhanced AR-mediated transcription in prostate cancer cells. In an effort to further dissect the mechanisms implicated in AR transactivation, we report here that AR interacts with PITALRE, a kinase subunit of positive elongation factor b (P-TEFb). Cotransfection of the plasmid encoding the mutant PITALRE (mtPITALRE), defective in its RNA polymerase II COOH terminal domain (CTD)-kinase activity, resulted in preferential inhibition of AR mediated transactivation. Indeed, AR transactivation in PC-3 cells was preferentially inhibited at the low concentration of 5,6-dichloro-1-beta-d ribofuranosylbenzimidazole (DRB), a CTD kinase inhibitor. These results suggest that CTD phosphorylation may play an important role in AR-mediated transcription. Furthermore, a nuclear run-on transcription assay of the prostate-specific antigen gene, an androgen-inducible gene, showed that transcription efficiency of the distal region of the gene was enhanced upon androgen induction. Taken together, our reports suggest that AR interacts with TFIIH and P-TEFb and enhances the elongation stage of transcription. PMID- 11266438 TI - Rescue of embryonic lethality in reduced folate carrier-deficient mice by maternal folic acid supplementation reveals early neonatal failure of hematopoietic organs. AB - The reduced folate carrier (RFC1) is an important route by which the major blood folate, 5-methyltetrahydrofolate, is transported into mammalian cells. In this study we determined the consequences of inactivation of RFC1 in mice by homologous recombination. While RFC1-null embryos died in utero before embryonic day 9.5 (E9.5), near-normal development could be sustained in RFC1(-)/- embryos examined at E18.5 by supplementation of pregnant RFC1(+/-) dams with 1-mg daily subcutaneous doses of folic acid. About 10% of these animals went on to live birth but died within 12 days. These RFC1(-)/- mice showed a marked absence of erythropoiesis in bone marrow, spleen, and liver along with lymphoid depletion in the splenic white pulp and thymus. In addition, there was some impairment of renal and seminiferous tubule development. These data indicate that in the absence of RFC1 function, neonatal animals die due to failure of hematopoietic organs. PMID- 11266439 TI - The pro domain of beta-secretase does not confer strict zymogen-like properties but does assist proper folding of the protease domain. AB - beta-Secretase (BACE) is a membrane-bound aspartyl protease that cleaves the amyloid precursor protein to generate the N terminus of the amyloid beta peptide. BACE is expressed as a precursor protein containing Pre, Pro, protease, transmembrane, and cytosolic domains. A soluble BACE derivative (PreProBACE460) that is truncated between the protease and transmembrane domains was produced by baculovirus-mediated expression. ProBACE460 was purified from conditioned media of infected insect cells using immobilized concanavalin A and immobilized BACE inhibitor, P10-P4' Stat(Val). Furin cleaves ProBACE460 between the Pro and protease regions to generate mature BACE460. The k(cat)/K(m) of ProBACE460 when assayed with a polypeptide substrate is only 2.3-fold less than that of BACE460. This finding and the similar inhibitory potency of P10-P4' Stat(Val) for ProBACE460 and BACE460 suggest that the Pro domain has little effect on the BACE active site. Exposure of ProBACE460 to guanidine denaturation/renaturation results in a 7-fold higher recovery of BACE activity than when BACE460 is similarly treated. The presence of free BACE Pro peptide during renaturation of BACE460 but not ProBACE460 increases recovery of activity. These findings show that the Pro domain in ProBACE460 does not suppress activity as in a strict zymogen but does appear to facilitate proper folding of an active protease domain. PMID- 11266441 TI - Nuclear position leaves its mark on replication timing. PMID- 11266443 TI - Cortical Num1p interacts with the dynein intermediate chain Pac11p and cytoplasmic microtubules in budding yeast. AB - Num1p, a cortical 313-kD protein, controls cytoplasmic microtubule (cMT) functions and nuclear migration through the bud neck in anaphase cells. A green fluorescent protein (GFP)-Num1p fusion protein localizes at the bud tip and the distal mother pole of living cells, apparently forming cMT capture sites at late anaphase. In addition, galactose-induced GFP-Num1p is seen at the bud neck and in lateral regions of the mother cortex. The bud tip location of Num1p depends on Bni1p but does not require Kar9p, Dyn1p, or cMTs, whereas cMT contacts with polar Num1p dots are reduced in cells lacking Dyn1p. Num1p associates with the dynein intermediate chain Pac11p in the presence of Dyn1p, and with the alpha-tubulin Tub3p, as shown by coimmune precipitation of tagged proteins. Num1p also forms a complex with Bni1p and Kar9p, although Num1p is not required for Bni1p- and Kar9p dependent nuclear migration to the bud neck in preanaphase cells. Our data suggest that Num1p controls nuclear migration during late anaphase by forming dynein-interacting cortical cMT capture sites at both cellular poles. In addition, Num1p may transiently cooperate with an associated Bni1p-Kar9p complex at the bud tip of early anaphase cells. PMID- 11266442 TI - Essential role of voltage-dependent anion channel in various forms of apoptosis in mammalian cells. AB - Through direct interaction with the voltage-dependent anion channel (VDAC), proapoptotic members of the Bcl-2 family such as Bax and Bak induce apoptogenic cytochrome c release in isolated mitochondria, whereas BH3-only proteins such as Bid and Bik do not directly target the VDAC to induce cytochrome c release. To investigate the biological significance of the VDAC for apoptosis in mammalian cells, we produced two kinds of anti-VDAC antibodies that inhibited VDAC activity. In isolated mitochondria, these antibodies prevented Bax-induced cytochrome c release and loss of the mitochondrial membrane potential (Deltapsi), but not Bid-induced cytochrome c release. When microinjected into cells, these anti-VDAC antibodies, but not control antibodies, also prevented Bax-induced cytochrome c release and apoptosis, whereas the antibodies did not prevent Bid induced apoptosis, indicating that the VDAC is essential for Bax-induced, but not Bid-induced, apoptogenic mitochondrial changes and apoptotic cell death. In addition, microinjection of these anti-VDAC antibodies significantly inhibited etoposide-, paclitaxel-, and staurosporine-induced apoptosis. Furthermore, we used these antibodies to show that Bax- and Bak-induced lysis of red blood cells was also mediated by the VDAC on plasma membrane. Taken together, our data provide evidence that the VDAC plays an essential role in apoptogenic cytochrome c release and apoptosis in mammalian cells. PMID- 11266445 TI - Downregulation of an AIM-1 kinase couples with megakaryocytic polyploidization of human hematopoietic cells. AB - During the late phase of megakaryopoiesis, megakaryocytes undergo polyploidization, which is characterized by DNA duplication without concomitant cell division. However, it remains unknown by which mechanisms this process occurs. AIM-1 and STK15 belong to the Aurora/increase-in-ploidy (Ipl)1 serine/threonine kinase family and play key roles in mitosis. In a human interleukin-3-dependent cell line, F-36P, the expressions of AIM-1 and STK15 mRNA were specifically observed at G2/M phase of the cell cycle during proliferation. In contrast, the expressions of AIM-1 and STK15 were continuously repressed during megakaryocytic polyploidization of human erythro/megakaryocytic cell lines (F-36P, K562, and CMK) treated with thrombopoietin, activated ras (H-ras(G12V)), or phorbol ester. Furthermore, their expressions were suppressed during thrombopoietin-induced polyploidization of normal human megakaryocytes. Activation of AIM-1 by the induced expression of AIM-1(wild-type) canceled TPA induced polyploidization of K562 cells significantly, whereas that of STK15 did not. Moreover, suppression of AIM-1 by the induced expression of AIM-1 (K/R, dominant-negative type) led to polyploidization in 25% of K562 cells, whereas STK15(K/R) showed no effect. Also, the induced expression of AIM-1(K/R) in CMK cells provoked polyploidization up to 32N. These results suggested that downregulation of AIM-1 at M phase may be involved in abortive mitosis and polyploid formation of megakaryocytes. PMID- 11266444 TI - Activation of endogenous thrombin receptors causes clustering and sensitization of epidermal growth factor receptors of swiss 3T3 cells without transactivation. AB - The G protein-coupled thrombin receptor can induce cellular responses in some systems by transactivating the epidermal growth factor (EGF) receptor. This is in part due to the stimulation of ectoproteases that generate EGF receptor ligands. We show here that this cannot account for the stimulation of proliferation or migration by thrombin of Swiss 3T3 cells. Thrombin has no direct effect on the activation state of the EGF receptor or of its downstream effectors. However, thrombin induces the subcellular clustering of the EGF receptor at filamentous actin-containing structures at the leading edge and actin arcs of migrating cells in association with other signaling molecules, including Shc and phospholipase Cgamma1. In these thrombin-primed cells, the subsequent migratory response to EGF is potentiated. Thrombin did not potentiate the EGF-stimulated EGF receptor phosphorylation. Thus, in Swiss 3T3 cells the G protein-coupled thrombin receptor can potentiate the EGF tyrosine kinase receptor response when activated by EGF, and this appears to be due to the subcellular concentration of the receptor with downstream effectors and not to the overall ability of EGF to induce receptor transphosphorylation. Thus, the EGF receptor subcellular localization which is altered by thrombin appears to be an important determinant of the efficacy of downstream EGF receptor signaling in cell migration. PMID- 11266446 TI - Biogenesis of porin of the outer mitochondrial membrane involves an import pathway via receptors and the general import pore of the TOM complex. AB - Porin, also termed the voltage-dependent anion channel, is the most abundant protein of the mitochondrial outer membrane. The process of import and assembly of the protein is known to be dependent on the surface receptor Tom20, but the requirement for other mitochondrial proteins remains controversial. We have used mitochondria from Neurospora crassa and Saccharomyces cerevisiae to analyze the import pathway of porin. Import of porin into isolated mitochondria in which the outer membrane has been opened is inhibited despite similar levels of Tom20 as in intact mitochondria. A matrix-destined precursor and the porin precursor compete for the same translocation sites in both normal mitochondria and mitochondria whose surface receptors have been removed, suggesting that both precursors utilize the general import pore. Using an assay established to monitor the assembly of in vitro-imported porin into preexisting porin complexes we have shown that besides Tom20, the biogenesis of porin depends on the central receptor Tom22, as well as Tom5 and Tom7 of the general import pore complex (translocase of the outer mitochondrial membrane [TOM] core complex). The characterization of two new mutant alleles of the essential pore protein Tom40 demonstrates that the import of porin also requires a functional Tom40. Moreover, the porin precursor can be cross-linked to Tom20, Tom22, and Tom40 on its import pathway. We conclude that import of porin does not proceed through the action of Tom20 alone, but requires an intact outer membrane and involves at least four more subunits of the TOM machinery, including the general import pore. PMID- 11266447 TI - The role of macrophages in demyelinating peripheral nervous system of mice heterozygously deficient in p0. AB - Mice heterozygously deficient in the p0 gene (P0(+/-)) are animal models for some forms of inherited neuropathies. They display a progressive demyelinating phenotype in motor nerves, accompanied by mild infiltration of lymphocytes and increase in macrophages. We have shown previously that the T lymphocytes are instrumental in the demyelination process. This study addresses the functional role of the macrophage in this monogenic myelin disorder. In motor nerves of P0(+/)- mice, the number of macrophages in demyelinated peripheral nerves was increased by a factor of five when compared with motor nerves of wild-type mice. Immunoelectron microscopy, using a specific marker for mouse macrophages, displayed macrophages not only in the endoneurium of the myelin mutants, but also within endoneurial tubes, suggesting an active role in demyelination. To elucidate the roles of the macrophages, we crossbred the myelin mutants with a spontaneous mouse mutant deficient in macrophage colony-stimulating factor (M CSF), hence displaying impaired macrophage activation. In the P0-deficient double mutants also deficient in M-CSF, the numbers of macrophages were not elevated in the demyelinating motor nerves and demyelination was less severe. These findings demonstrate an active role of macrophages during pathogenesis of inherited demyelination with putative impact on future treatment strategies. PMID- 11266448 TI - The role of dynamin and its binding partners in coated pit invagination and scission. AB - Plasma membrane clathrin-coated vesicles form after the directed assembly of clathrin and the adaptor complex, AP2, from the cytosol onto the membrane. In addition to these structural components, several other proteins have been implicated in clathrin-coated vesicle formation. These include the large molecular weight GTPase, dynamin, and several Src homology 3 (SH3) domain containing proteins which bind to dynamin via interactions with its COOH-terminal proline/arginine-rich domain (PRD). To understand the mechanism of coated vesicle formation, it is essential to determine the hierarchy by which individual components are targeted to and act in coated pit assembly, invagination, and scission. To address the role of dynamin and its binding partners in the early stages of endocytosis, we have used well-established in vitro assays for the late stages of coated pit invagination and coated vesicle scission. Dynamin has previously been shown to have a role in scission of coated vesicles. We show that dynamin is also required for the late stages of invagination of clathrin-coated pits. Furthermore, dynamin must bind and hydrolyze GTP for its role in sequestering ligand into deeply invaginated coated pits. We also demonstrate that the SH3 domain of endophilin, which binds both synaptojanin and dynamin, inhibits both late stages of invagination and also scission in vitro. This inhibition results from a reduction in phosphoinositide 4,5-bisphosphate levels which causes dissociation of AP2, clathrin, and dynamin from the plasma membrane. The dramatic effects of the SH3 domain of endophilin led us to propose a model for the temporal order of addition of endophilin and its binding partner synaptojanin in the coated vesicle cycle. PMID- 11266449 TI - Negative regulation of Ros receptor tyrosine kinase signaling. An epithelial function of the SH2 domain protein tyrosine phosphatase SHP-1. AB - Male "viable motheaten" (me(v)) mice, with a naturally occurring mutation in the gene of the SH2 domain protein tyrosine phosphatase SHP-1, are sterile. Known defects in sperm maturation in these mice correlate with an impaired differentiation of the epididymis, which has similarities to the phenotype of mice with a targeted inactivation of the Ros receptor tyrosine kinase. Ros and SHP-1 are coexpressed in epididymal epithelium, and elevated phosphorylation of Ros in the epididymis of me(v) mice suggests that Ros signaling is under control of SHP-1 in vivo. Phosphorylated Ros strongly and directly associates with SHP-1 in yeast two-hybrid, glutathione S-transferase pull-down, and coimmunoprecipitation experiments. Strong binding of SHP-1 to Ros is selective compared to six other receptor tyrosine kinases. The interaction is mediated by the SHP-1 NH(2)-terminal SH2 domain and Ros phosphotyrosine 2267. Overexpression of SHP-1 results in Ros dephosphorylation and effectively downregulates Ros dependent proliferation and transformation. We propose that SHP-1 is an important downstream regulator of Ros signaling. PMID- 11266450 TI - Intraarterial injection of muscle-derived CD34(+)Sca-1(+) stem cells restores dystrophin in mdx mice. AB - Duchenne muscular dystrophy is a lethal recessive disease characterized by widespread muscle damage throughout the body. This increases the difficulty of cell or gene therapy based on direct injections into muscles. One way to circumvent this obstacle would be to use circulating cells capable of homing to the sites of lesions. Here, we showed that stem cell antigen 1 (Sca-1), CD34 double-positive cells purified from the muscle tissues of newborn mice are multipotent in vitro and can undergo both myogenic and multimyeloid differentiation. These muscle-derived stem cells were isolated from newborn mice expressing the LacZ gene under the control of the muscle-specific desmin or troponin I promoter and injected into arterial circulation of the hindlimb of mdx mice. The ability of these cells to interact and firmly adhere to endothelium in mdx muscles microcirculation was demonstrated by intravital microscopy after an intraarterial injection. Donor Sca-1, CD34 muscle-derived stem cells were able to migrate from the circulation into host muscle tissues. Histochemical analysis showed colocalization of LacZ and dystrophin expression in all muscles of the injected hindlimb in all of five out of five 8-wk-old treated mdx mice. Their participation in the formation of muscle fibers was significantly increased by muscle damage done 48 h after their intraarterial injection, as indicated by the presence of 12% beta-galactosidase-positive fibers in muscle cross sections. Normal dystrophin transcripts detected enzymes in the muscles of the hind limb injected intraarterially by the mdx reverse transcription polymerase chain reaction method, which differentiates between normal and mdx message. Our results showed that the muscle-derived stem cells first attach to the capillaries of the muscles and then participate in regeneration after muscle damage. PMID- 11266451 TI - The Ndc80p complex from Saccharomyces cerevisiae contains conserved centromere components and has a function in chromosome segregation. AB - We have purified a complex from Saccharomyces cerevisiae containing the spindle components Ndc80p, Nuf2p, Spc25p, and Spc24p. Temperature-sensitive mutants in NDC80, SPC25, and SPC24 show defects in chromosome segregation. In spc24-1 cells, green fluorescence protein (GFP)-labeled centromeres fail to split during spindle elongation, and in addition some centromeres may detach from the spindle. Chromatin immunoprecipitation assays show an association of all four components of the complex with the yeast centromere. Homologues of Ndc80p, Nuf2p, and Spc24p were found in Schizosaccharomyces pombe and GFP tagging showed they were located at the centromere. A human homologue of Nuf2p was identified in the expressed sequence tag database. Immunofluorescent staining with anti-human Nuf2p and with anti-HEC, the human homologue of Ndc80p, showed that both proteins are at the centromeres of mitotic HeLa cells. Thus the Ndc80p complex contains centromere associated components conserved between yeasts and vertebrates. PMID- 11266452 TI - Convergence of alpha(v)beta(3) integrin- and macrophage colony stimulating factor mediated signals on phospholipase Cgamma in prefusion osteoclasts. AB - The macrophage colony stimulating factor (M-CSF) and alpha(v)beta(3) integrins play critical roles in osteoclast function. This study examines M-CSF- and adhesion-induced signaling in prefusion osteoclasts (pOCs) derived from Src deficient and wild-type mice. Src-deficient cells attach to but do not spread on vitronectin (Vn)-coated surfaces and, contrary to wild-type cells, their adhesion does not lead to tyrosine phosphorylation of molecules activated by adhesion, including PYK2, p130(Cas), paxillin, and PLC-gamma. However, in response to M CSF, Src(-/-) pOCs spread and migrate on Vn in an alpha(v)beta(3)-dependent manner. Involvement of PLC-gamma activation is suggested by using a PLC inhibitor, U73122, which blocks both adhesion- and M-CSF-mediated cell spreading. Furthermore, in Src(-/-) pOCs M-CSF, together with filamentous actin, causes recruitment of beta(3) integrin and PLC-gamma to adhesion contacts and induces stable association of beta(3) integrin with PLC-gamma, phosphatidylinositol 3 kinase, and PYK2. Moreover, direct interaction of PYK2 and PLC-gamma can be induced by either adhesion or M-CSF, suggesting that this interaction may enable the formation of integrin-associated complexes. Furthermore, this study suggests that in pOCs PLC-gamma is a common downstream mediator for adhesion and growth factor signals. M-CSF-initiated signaling modulates the alpha(v)beta(3) integrin mediated cytoskeletal reorganization in prefusion osteoclasts in the absence of c Src, possibly via PLC-gamma. PMID- 11266453 TI - A novel chromatin protein, distantly related to histone H2A, is largely excluded from the inactive X chromosome. AB - Chromatin on the mammalian inactive X chromosome differs in a number of ways from that on the active X. One protein, macroH2A, whose amino terminus is closely related to histone H2A, is enriched on the heterochromatic inactive X chromosome in female cells. Here, we report the identification and localization of a novel and more distant histone variant, designated H2A-Bbd, that is only 48% identical to histone H2A. In both interphase and metaphase female cells, using either a myc epitope-tagged or green fluorescent protein-tagged H2A-Bbd construct, the inactive X chromosome is markedly deficient in H2A-Bbd staining, while the active X and the autosomes stain throughout. In double-labeling experiments, antibodies to acetylated histone H4 show a pattern of staining indistinguishable from H2A Bbd in interphase nuclei and on metaphase chromosomes. Chromatin fractionation demonstrates association of H2A-Bbd with the histone proteins. Separation of micrococcal nuclease-digested chromatin by sucrose gradient ultracentrifugation shows cofractionation of H2A-Bbd with nucleosomes, supporting the idea that H2A Bbd is incorporated into nucleosomes as a substitute for the core histone H2A. This finding, in combination with the overlap with acetylated forms of H4, raises the possibility that H2A-Bbd is enriched in nucleosomes associated with transcriptionally active regions of the genome. The distribution of H2A-Bbd thus distinguishes chromatin on the active and inactive X chromosomes. PMID- 11266454 TI - The positioning and dynamics of origins of replication in the budding yeast nucleus. AB - We have analyzed the subnuclear position of early- and late-firing origins of DNA replication in intact yeast cells using fluorescence in situ hybridization and green fluorescent protein (GFP)-tagged chromosomal domains. In both cases, origin position was determined with respect to the nuclear envelope, as identified by nuclear pore staining or a NUP49-GFP fusion protein. We find that in G1 phase nontelomeric late-firing origins are enriched in a zone immediately adjacent to the nuclear envelope, although this localization does not necessarily persist in S phase. In contrast, early firing origins are randomly localized within the nucleus throughout the cell cycle. If a late-firing telomere-proximal origin is excised from its chromosomal context in G1 phase, it remains late-firing but moves rapidly away from the telomere with which it was associated, suggesting that the positioning of yeast chromosomal domains is highly dynamic. This is confirmed by time-lapse microscopy of GFP-tagged origins in vivo. We propose that sequences flanking late-firing origins help target them to the periphery of the G1-phase nucleus, where a modified chromatin structure can be established. The modified chromatin structure, which would in turn retard origin firing, is both autonomous and mobile within the nucleus. PMID- 11266455 TI - Mmm1p, a mitochondrial outer membrane protein, is connected to mitochondrial DNA (mtDNA) nucleoids and required for mtDNA stability. AB - In the yeast Saccharomyces cerevisiae, mitochondria form a branched, tubular reticulum in the periphery of the cell. Mmm1p is required to maintain normal mitochondrial shape and in mmm1 mutants mitochondria form large, spherical organelles. To further explore Mmm1p function, we examined the localization of a Mmm1p-green fluorescent protein (GFP) fusion in living cells. We found that Mmm1p GFP is located in small, punctate structures on the mitochondrial outer membrane, adjacent to a subset of matrix-localized mitochondrial DNA nucleoids. We also found that the temperature-sensitive mmm1-1 mutant was defective in transmission of mitochondrial DNA to daughter cells immediately after the shift to restrictive temperature. Normal mitochondrial nucleoid structure also collapsed at the nonpermissive temperature with similar kinetics. Moreover, we found that mitochondrial inner membrane structure is dramatically disorganized in mmm1 disruption strains. We propose that Mmm1p is part of a connection between the mitochondrial outer and inner membranes, anchoring mitochondrial DNA nucleoids in the matrix. PMID- 11266456 TI - Gradient of increasing affinity of importin beta for nucleoporins along the pathway of nuclear import. AB - Nuclear import and export signals on macromolecules mediate directional, receptor driven transport through the nuclear pore complex (NPC) by a process that is suggested to involve the sequential binding of transport complexes to different nucleoporins. The directionality of transport appears to be partly determined by the nucleocytoplasmic compartmentalization of components of the Ran GTPase system. We have analyzed whether the asymmetric localization of discrete nucleoporins can also contribute to transport directionality. To this end, we have used quantitative solid phase binding analysis to determine the affinity of an importin beta cargo complex for Nup358, the Nup62 complex, and Nup153, which are in the cytoplasmic, central, and nucleoplasmic regions of the NPC, respectively. These nucleoporins are proposed to provide progressively more distal binding sites for importin beta during import. Our results indicate that the importin beta transport complex binds to nucleoporins with progressively increasing affinity as the complex moves from Nup358 to the Nup62 complex and to Nup153. Antibody inhibition studies support the possibility that importin beta moves from Nup358 to Nup153 via the Nup62 complex during import. These results indicate that nucleoporins themselves, as well as the nucleocytoplasmic compartmentalization of the Ran system, are likely to play an important role in conferring directionality to nuclear protein import. PMID- 11266458 TI - Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. AB - In macroautophagy, cytoplasmic components are delivered to lysosomes for degradation via autophagosomes that are formed by closure of cup-shaped isolation membranes. However, how the isolation membranes are formed is poorly understood. We recently found in yeast that a novel ubiquitin-like system, the Apg12-Apg5 conjugation system, is essential for autophagy. Here we show that mouse Apg12 Apg5 conjugate localizes to the isolation membranes in mouse embryonic stem cells. Using green fluorescent protein-tagged Apg5, we revealed that the cup shaped isolation membrane is developed from a small crescent-shaped compartment. Apg5 localizes on the isolation membrane throughout its elongation process. To examine the role of Apg5, we generated Apg5-deficient embryonic stem cells, which showed defects in autophagosome formation. The covalent modification of Apg5 with Apg12 is not required for its membrane targeting, but is essential for involvement of Apg5 in elongation of the isolation membranes. We also show that Apg12-Apg5 is required for targeting of a mammalian Aut7/Apg8 homologue, LC3, to the isolation membranes. These results suggest that the Apg12-Apg5 conjugate plays essential roles in isolation membrane development. PMID- 11266459 TI - Drosophila aurora B kinase is required for histone H3 phosphorylation and condensin recruitment during chromosome condensation and to organize the central spindle during cytokinesis. AB - Aurora/Ipl1-related kinases are a conserved family of enzymes that have multiple functions during mitotic progression. Although it has been possible to use conventional genetic analysis to dissect the function of aurora, the founding family member in Drosophila (Glover, D.M., M.H. Leibowitz, D.A. McLean, and H. Parry. 1995. Cell. 81:95-105), the lack of mutations in a second aurora-like kinase gene, aurora B, precluded this approach. We now show that depleting Aurora B kinase using double-stranded RNA interference in cultured Drosophila cells results in polyploidy. aurora B encodes a passenger protein that associates first with condensing chromatin, concentrates at centromeres, and then relocates onto the central spindle at anaphase. Cells depleted of the Aurora B kinase show only partial chromosome condensation at mitosis. This is associated with a reduction in levels of the serine 10 phosphorylated form of histone H3 and a failure to recruit the Barren condensin protein onto chromosomes. These defects are associated with abnormal segregation resulting from lagging chromatids and extensive chromatin bridging at anaphase, similar to the phenotype of barren mutants (Bhat, M.A., A.V. Philp, D.M. Glover, and H.J. Bellen. 1996. Cell. 87:1103-1114.). The majority of treated cells also fail to undertake cytokinesis and show a reduced density of microtubules in the central region of the spindle. This is accompanied by a failure to correctly localize the Pavarotti kinesin-like protein, essential for this process. We discuss these conserved functions of Aurora B kinase in chromosome transmission and cytokinesis. PMID- 11266460 TI - Connection of the mitochondrial outer and inner membranes by Fzo1 is critical for organellar fusion. AB - Mitochondrial membrane fusion is a process essential for the maintenance of the structural integrity of the organelle. Since mitochondria are bounded by a double membrane, they face the challenge of fusing four membranes in a coordinated manner. We provide evidence that this is achieved by coupling of the mitochondrial outer and inner membranes by the mitochondrial fusion machinery. Fzo1, the first known mediator of mitochondrial fusion, spans the outer membrane twice, exposing a short loop to the intermembrane space. The presence of the intermembrane space segment is required for the localization of Fzo1 in sites of tight contact between the mitochondrial outer and inner membranes. Mutations in the intermembrane space domain of yeast Fzo1 relieve the association with the inner membrane. This results in a loss of function of the protein in vivo. We propose that the mitochondrial fusion machinery forms membrane contact sites that mediate mitochondrial fusion. A fusion machinery that is in contact with both mitochondrial membranes appears to be functionally important for coordinated fusion of four mitochondrial membranes. PMID- 11266462 TI - Hsp70 and antifibrillogenic peptides promote degradation and inhibit intracellular aggregation of amyloidogenic light chains. AB - In light chain (LC) amyloidosis an immunoglobulin LC assembles into fibrils that are deposited in various tissues. Little is known about how these fibrils form in vivo. We previously showed that a known amyloidogenic LC, SMA, can give rise to amyloid fibrils in vitro when a segment of one of its beta sheets undergoes a conformational change, exposing an Hsp70 binding site. To examine SMA aggregation in vivo, we expressed it and its wild-type counterpart, LEN, in COS cells. While LEN is rapidly oxidized and subsequently secreted, newly synthesized SMA remains in the reduced state. Most SMA molecules are dislocated out of the ER into the cytosol, where they are ubiquitinylated and degraded by proteasomes. A parallel pathway for molecules that are not degraded is condensation into perinuclear aggresomes that are surrounded by vimentin-containing intermediate filaments and are dependent upon intact microtubules. Inhibition of proteasome activity shifts the balance toward aggresome formation. Intracellular aggregation is decreased and targeting to proteasomes improved by overexpression of the cytosolic chaperone Hsp70. Importantly, transduction into the cell of an Hsp70 target peptide, derived from the LC sequence, also reduces aggresome formation and increases SMA degradation. These results demonstrate that an amyloidogenic LC can aggregate intracellularly despite the common presentation of extracellular aggregates, and that a similar molecular surface mediates both in vitro fibril formation and in vivo aggregation. Furthermore, rationally designed peptides can be used to suppress this aggregation and may provide a feasible therapeutic approach. PMID- 11266461 TI - Stromelysin-1 regulates adipogenesis during mammary gland involution. AB - The matrix metalloproteinase MMP-3/stromelysin-1 (Str1) is highly expressed during mammary gland involution induced by weaning. During involution, programmed cell death of the secretory epithelium takes place concomitant with the repopulation of the mammary fat pad with adipocytes. In this study, we have used a genetic approach to determine the role of Str1 during mammary involution. Although Str1 has been shown to induce unscheduled apoptosis when expressed ectopically during late pregnancy (Alexander, C.M., E.W. Howard, M.J. Bissell, and Z. Werb. 1996. J. Cell Biol. 135:1669-1677), we found that during post lactational involution, mammary glands from transgenic mice that overexpress the tissue inhibitor of metalloproteinases, TIMP-1 (TO), or mice carrying a targeted mutation in Str1 showed accelerated differentiation and hypertrophy of adipocytes, while epithelial apoptosis was unaffected. These data suggest that matrix metalloproteinases (MMPs) do not induce unscheduled epithelial cell death after weaning, but instead alter the stromal microenvironment. We used adipogenic 3T3-L1 cells as a cell culture model to test the function of MMPs during adipocyte differentiation. Fibroblastic 3T3-L1 progenitor cells expressed very low levels of MMPs or TIMPs. The transcription of a number of MMP and TIMP mRNAs [Str1, MT1-MMP, (MMP-14) collagenase-3 (MMP-13), gelatinase A (MMP-2), and TIMP 1, -2 and -3] was induced in committed preadipocytes, but only differentiated adipocytes expressed an activated MMP, gelatinase A. The addition of MMP inhibitors (GM 6001 and TIMP-1) dramatically accelerated the accumulation of lipid during differentiation. We conclude that MMPs, especially Str1, determine the rate of adipocyte differentiation during involutive mammary gland remodeling. PMID- 11266463 TI - A specific role of phosphatidylinositol 3-kinase gamma. A regulation of autonomic Ca(2)+ oscillations in cardiac cells. AB - Purinergic stimulation of cardiomyocytes turns on a Src family tyrosine kinase dependent pathway that stimulates PLCgamma and generates IP(3), a breakdown product of phosphatidylinositol 4,5-bisphosphate (PIP2). This signaling pathway closely regulates cardiac cell autonomic activity (i.e., spontaneous cell Ca(2+) spiking). PIP2 is phosphorylated on 3' by phosphoinositide 3-kinases (PI3Ks) that belong to a broad family of kinase isoforms. The product of PI3K, phosphatidylinositol 3,4,5-trisphosphate, regulates activity of PLCgamma. PI3Ks have emerged as crucial regulators of many cell functions including cell division, cell migration, cell secretion, and, via PLCgamma, Ca(2+) homeostasis. However, although PI3Kalpha and -beta have been shown to mediate specific cell functions in nonhematopoietic cells, such a role has not been found yet for PI3Kgamma. We report that neonatal rat cardiac cells in culture express PI3Kalpha, -beta, and -gamma. The purinergic agonist predominantly activates PI3Kgamma. Both wortmannin and LY294002 prevent tyrosine phosphorylation, and membrane translocation of PLCgamma as well as IP(3) generation in ATP-stimulated cells. Furthermore, an anti-PI3Kgamma, but not an anti-PI3Kbeta, injected in the cells prevents the effect of ATP on cell Ca(2+) spiking. A dominant negative mutant of PI3Kgamma transfected in the cells also exerts the same action. The effect of ATP was observed on spontaneous Ca(2+) spiking of wild-type but not of PI3Kgamma(2/2) embryonic stem cell-derived cardiomyocytes. ATP activates the Btk tyrosine kinase, Tec, and induces its association with PLCgamma. A dominant negative mutant of Tec blocks the purinergic effect on cell Ca(2+) spiking. Tec is translocated to the T-tubes upon ATP stimulation of cardiac cells. Both an anti-PI3Kgamma antibody and a dominant negative mutant of PI3Kgamma injected or transfected into cells prevent the latter event. We conclude that PI3Kgamma activation is a crucial step in the purinergic regulation of cardiac cell spontaneous Ca(2+) spiking. Our data further suggest that Tec works in concert with a Src family kinase and PI3Kgamma to fully activate PLCgamma in ATP stimulated cardiac cells. This cluster of kinases provides the cardiomyocyte with a tight regulation of IP(3) generation and thus cardiac autonomic activity. PMID- 11266464 TI - The karyopherin Kap142p/Msn5p mediates nuclear import and nuclear export of different cargo proteins. AB - We have identified a novel pathway for protein import into the nucleus. Although the product of Saccharomyces cerevisiae gene MSN5 was previously shown to function as a karyopherin (Kap) for nuclear export of various proteins, we discovered a nuclear import pathway mediated by Msn5p (also referred to as Kap142p). We have purified from yeast cytosol a complex containing Kap142p and the trimeric replication protein A (RPA), which is required for multiple aspects of DNA metabolism, including DNA replication, DNA repair, and recombination. In wild-type cells, RPA was localized primarily to the nucleus but, in a KAP142 deletion strain, RPA was mislocalized to the cytoplasm and the strain was highly sensitive to bleomycin (BLM). BLM causes DNA double-strand breaks and, in S. cerevisiae, the DNA damage is repaired predominantly by RPA-dependent homologous recombination. Therefore, our results indicate that in wild-type cells a critical portion of RPA was imported into the nucleus by Kap142p. Like several other import-related Kap-substrate complexes, the endogenous RPA-Kap142p complex was dissociated by RanGTP, but not by RanGDP. All three RPA genes are essential for viability, whereas KAP142 is not. Perhaps explaining this disparity, we observed an interaction between RPA and Kap95p in a strain lacking Kap142p. This interaction could provide a mechanism for import of RPA into the nucleus and cell viability in the absence of Kap142p. Together with published results (Kaffman, A., N.M. Rank, E.M. O'Neill, L.S. Huang, and E.K. O'Shea. 1998. Nature. 396:482 486; Blondel, M., P.M. Alepuz, L.S. Huang, S. Shaham, G. Ammerer, and M. Peter. 1999. Genes Dev. 13:2284-2300; DeVit, M.J., and M. Johnston. 1999. Curr. Biol. 9:1231-1241; Mahanty, S.K., Y. Wang, F.W. Farley, and E.A. Elion. 1999. Cell. 98:501-512) our data indicate that the karyopherin Kap142p is able to mediate nuclear import of one set of proteins and nuclear export of a different set of proteins. PMID- 11266465 TI - Plasminogen activator inhibitor 1 functions as a urokinase response modifier at the level of cell signaling and thereby promotes MCF-7 cell growth. AB - Plasminogen activator inhibitor 1 (PAI-1) is a major inhibitor of urokinase-type plasminogen activator (uPA). In this study, we explored the role of PAI-1 in cell signaling. In MCF-7 cells, PAI-1 did not directly activate the mitogen-activated protein (MAP) kinases, extracellular signal-regulated kinase (ERK) 1 and ERK2, but instead altered the response to uPA so that ERK phosphorylation was sustained. This effect required the cooperative function of uPAR and the very low density lipoprotein receptor (VLDLr). When MCF-7 cells were treated with uPA-PAI 1 complex in the presence of the VLDLr antagonist, receptor-associated protein, or with uPA-PAI-1(R76E) complex, which binds to the VLDLr with greatly decreased affinity, transient ERK phosphorylation (<5 min) was observed, mimicking the uPA response. ERK phosphorylation was not induced by tissue-type plasminogen activator-PAI-1 complex or by uPA-PAI-1 complex in the presence of antibodies that block uPA binding to uPAR. uPA-PAI-1 complex induced tyrosine phosphorylation of focal adhesion kinase and Shc and sustained association of Sos with Shc, whereas uPA caused transient association of Sos with Shc. By sustaining ERK phosphorylation, PAI-1 converted uPA into an MCF-7 cell mitogen. This activity was blocked by receptor-associated protein and not observed with uPA-PAI 1(R76E) complex, demonstrating the importance of the VLDLr. uPA promoted the growth of other cells in which ERK phosphorylation was sustained, including beta3 integrin overexpressing MCF-7 cells and HT 1080 cells. The MEK inhibitor, PD098059, blocked the growth-promoting activity of uPA and uPA-PAI-1 complex in these cells. Our results demonstrate that PAI-1 may regulate uPA-initiated cell signaling by a mechanism that requires VLDLr recruitment. The kinetics of ERK phosphorylation in response to uPAR ligation determine the function of uPA and uPA-PAI-1 complex as growth promoters. PMID- 11266466 TI - Activation of nuclear factor kappaB and Bcl-x survival gene expression by nerve growth factor requires tyrosine phosphorylation of IkappaBalpha. AB - NGF has been shown to support neuron survival by activating the transcription factor nuclear factor-kappaB (NFkappaB). We investigated the effect of NGF on the expression of Bcl-xL, an anti-apoptotic Bcl-2 family protein. Treatment of rat pheochromocytoma PC12 cells, human neuroblastoma SH-SY5Y cells, or primary rat hippocampal neurons with NGF (0.1-10 ng/ml) increased the expression of bcl-xL mRNA and protein. Reporter gene analysis revealed a significant increase in NFkappaB activity after treatment with NGF that was associated with increased nuclear translocation of the active NFkappaB p65 subunit. NGF-induced NFkappaB activity and Bcl-xL expression were inhibited in cells overexpressing the NFkappaB inhibitor, IkappaBalpha. Unlike tumor necrosis factor-alpha (TNF-alpha), however, NGF-induced NFkappaB activation occurred without significant degradation of IkappaBs determined by Western blot analysis and time-lapse imaging of neurons expressing green fluorescent protein-tagged IkappaBalpha. Moreover, in contrast to TNF-alpha, NGF failed to phosphorylate IkappaBalpha at serine residue 32, but instead caused significant tyrosine phosphorylation. Overexpression of a Y42F mutant of IkappaBalpha potently suppressed NFG-, but not TNF-alpha-induced NFkappaB activation. Conversely, overexpression of a dominant negative mutant of TNF receptor-associated factor-6 blocked TNF-alpha-, but not NGF-induced NFkappaB activation. We conclude that NGF and TNF-alpha induce different signaling pathways in neurons to activate NFkappaB and bcl-x gene expression. PMID- 11266467 TI - A novel 14-kilodalton protein interacts with the mitogen-activated protein kinase scaffold mp1 on a late endosomal/lysosomal compartment. AB - We have identified a novel, highly conserved protein of 14 kD copurifying with late endosomes/lysosomes on density gradients. The protein, now termed p14, is peripherally associated with the cytoplasmic face of late endosomes/lysosomes in a variety of different cell types. In a two-hybrid screen with p14 as a bait, we identified the mitogen-activated protein kinase (MAPK) scaffolding protein MAPK/extracellular signal-regulated kinase (ERK) kinase (MEK) partner 1 (MP1) as an interacting protein. We confirmed the specificity of this interaction in vitro by glutathione S-transferase pull-down assays and by coimmunoprecipitation, cosedimentation on glycerol gradients, and colocalization. Moreover, expression of a plasma membrane-targeted p14 causes mislocalization of coexpressed MP1. In addition, we could reconstitute protein complexes containing the p14-MP1 complex associated with ERK and MEK in vitro.The interaction between p14 and MP1 suggests a MAPK scaffolding activity localized to the cytoplasmic surface of late endosomes/lysosomes, thereby combining catalytic scaffolding and subcellular compartmentalization as means to modulate MAPK signaling within a cell. PMID- 11266468 TI - The plasminogen activator inhibitor PAI-1 controls in vivo tumor vascularization by interaction with proteases, not vitronectin. Implications for antiangiogenic strategies. AB - The plasminogen (Plg)/plasminogen activator (PA) system plays a key role in cancer progression, presumably via mediating extracellular matrix degradation and tumor cell migration. Consequently, urokinase-type PA (uPA)/plasmin antagonists are currently being developed for suppression of tumor growth and angiogenesis. Paradoxically, however, high levels of PA inhibitor 1 (PAI-1) are predictive of a poor prognosis for survival of patients with cancer. We demonstrated previously that PAI-1 promoted tumor angiogenesis, but by an unresolved mechanism. We anticipated that PAI-1 facilitated endothelial cell migration via its known interaction with vitronectin (VN) and integrins. However, using adenoviral gene transfer of PAI-1 mutants, we observed that PAI-1 promoted tumor angiogenesis, not by interacting with VN, but rather by inhibiting proteolytic activity, suggesting that excessive plasmin proteolysis prevents assembly of tumor vessels. Single deficiency of uPA, tissue-type PA (tPA), uPA receptor, or VN, as well as combined deficiencies of uPA and tPA did not impair tumor angiogenesis, whereas lack of Plg reduced it. Overall, these data indicate that plasmin proteolysis, even though essential, must be tightly controlled during tumor angiogenesis, probably to allow vessel stabilization and maturation. These data provide insights into the clinical paradox whereby PAI-1 promotes tumor progression and warrant against the uncontrolled use of uPA/plasmin antagonists as tumor angiogenesis inhibitors. PMID- 11266469 TI - Presenilin 1 negatively regulates beta-catenin/T cell factor/lymphoid enhancer factor-1 signaling independently of beta-amyloid precursor protein and notch processing. AB - In addition to its documented role in the proteolytic processing of Notch-1 and the beta-amyloid precursor protein, presenilin 1 (PS1) associates with beta catenin. In this study, we show that this interaction plays a critical role in regulating beta-catenin/T Cell Factor/Lymphoid Enhancer Factor-1 (LEF) signaling. PS1 deficiency results in accumulation of cytosolic beta-catenin, leading to a beta-catenin/LEF-dependent increase in cyclin D1 transcription and accelerated entry into the S phase of the cell cycle. Conversely, PS1 specifically represses LEF-dependent transcription in a dose-dependent manner. The hyperproliferative response can be reversed by reintroducing PS1 expression or overexpressing axin, but not a PS1 mutant that does not bind beta-catenin (PS1Deltacat) or by two different familial Alzheimer's disease mutants. In contrast, PS1Deltacat restores Notch-1 proteolytic cleavage and Abeta generation in PS1-deficient cells, indicating that PS1 function in modulating beta-catenin levels can be separated from its roles in facilitating gamma-secretase cleavage of beta-amyloid precursor protein and in Notch-1 signaling. Finally, we show an altered response to Wnt signaling and impaired ubiquitination of beta-catenin in the absence of PS1, a phenotype that may account for the increased stability in PS1-deficient cells. Thus, PS1 adds to the molecules that are known to regulate the rapid turnover of beta-catenin. PMID- 11266470 TI - Rab27a regulates the peripheral distribution of melanosomes in melanocytes. AB - Rab GTPases are regulators of intracellular membrane traffic. We report a possible function of Rab27a, a protein implicated in several diseases, including Griscelli syndrome, choroideremia, and the Hermansky-Pudlak syndrome mouse model, gunmetal. We studied endogenous Rab27a and overexpressed enhanced GFP-Rab27a fusion protein in several cultured melanocyte and melanoma-derived cell lines. In pigmented cells, we observed that Rab27a decorates melanosomes, whereas in nonpigmented cells Rab27a colocalizes with melanosome-resident proteins. When dominant interfering Rab27a mutants were expressed in pigmented cells, we observed a redistribution of pigment granules with perinuclear clustering. This phenotype is similar to that observed by others in melanocytes derived from the ashen and dilute mutant mice, which bear mutations in the Rab27a and MyoVa loci, respectively. We also found that myosinVa coimmunoprecipitates with Rab27a in extracts from melanocytes and that both Rab27a and myosinVa colocalize on the cytoplasmic face of peripheral melanosomes in wild-type melanocytes. However, the amount of myosinVa in melanosomes from Rab27a-deficient ashen melanocytes is greatly reduced. These results, together with recent data implicating myosinVa in the peripheral capture of melanosomes, suggest that Rab27a is necessary for the recruitment of myosinVa, so allowing the peripheral retention of melanosomes in melanocytes. PMID- 11266471 TI - Distinct protein sorting and localization to premelanosomes, melanosomes, and lysosomes in pigmented melanocytic cells. AB - Melanosomes and premelanosomes are lysosome-related organelles with a unique structure and cohort of resident proteins. We have positioned these organelles relative to endosomes and lysosomes in pigmented melanoma cells and melanocytes. Melanosome resident proteins Pmel17 and TRP1 localized to separate vesicular structures that were distinct from those enriched in lysosomal proteins. In immunogold-labeled ultrathin cryosections, Pmel17 was most enriched along the intralumenal striations of premelanosomes. Increased pigmentation was accompanied by a decrease in Pmel17 and by an increase in TRP1 in the limiting membrane. Both proteins were largely excluded from lysosomal compartments enriched in LAMP1 and cathepsin D. By kinetic analysis of fluid phase uptake and immunogold labeling, premelanosomal proteins segregated from endocytic markers within an unusual endosomal compartment. This compartment contained Pmel17, was accessed by BSA gold after 15 min, was acidic, and displayed a cytoplasmic planar coat that contained clathrin. Our results indicate that premelanosomes and melanosomes represent a distinct lineage of organelles, separable from conventional endosomes and lysosomes within pigmented cells. Furthermore, they implicate an unusual clathrin-coated endosomal compartment as a site from which proteins destined for premelanosomes and lysosomes are sorted. PMID- 11266472 TI - Rab27a is required for regulated secretion in cytotoxic T lymphocytes. AB - Rab27a activity is affected in several mouse models of human disease including Griscelli (ashen mice) and Hermansky-Pudlak (gunmetal mice) syndromes. A loss of function mutation occurs in the Rab27a gene in ashen (ash), whereas in gunmetal (gm) Rab27a dysfunction is secondary to a mutation in the alpha subunit of Rab geranylgeranyl transferase, an enzyme required for prenylation and activation of Rabs. We show here that Rab27a is normally expressed in cytotoxic T lymphocytes (CTLs), but absent in ashen homozygotes (ash/ash). Cytotoxicity and secretion assays show that ash/ash CTLs are unable to kill target cells or to secrete granzyme A and hexosaminidase. By immunofluorescence and electron microscopy, we show polarization but no membrane docking of ash/ash lytic granules at the immunological synapse. In gunmetal CTLs, we show underprenylation and redistribution of Rab27a to the cytosol, implying reduced activity. Gunmetal CTLs show a reduced ability to kill target cells but retain the ability to secrete hexosaminidase and granzyme A. However, only some of the granules polarize to the immunological synapse, and many remain dispersed around the periphery of the CTLs. These results demonstrate that Rab27a is required in a final secretory step and that other Rab proteins also affected in gunmetal are likely to be involved in polarization of the granules to the immunological synapse. PMID- 11266473 TI - Defective granule exocytosis in Rab27a-deficient lymphocytes from Ashen mice. AB - Because mutations in Rab27a have been linked to immune defects in humans, we have examined cytotoxic lymphocyte function in ashen mice, which contain a splicing mutation in Rab27a. Ashen cytotoxic T lymphocytes (CTLs) showed a >90% reduction in lytic activity on Fas-negative target cells compared with control C3H CTLs, and ashen natural killer cell activity was likewise diminished. Although their granule-mediated cytotoxicity pathway is profoundly defective, ashen CTLs displayed a normal FasL-Fas cytotoxicity pathway. The CD4/8 phenotype of ashen T cells and their proliferative responses were similar to controls. Ashen CTLs had normal levels of perforin and granzymes A and B and normal-appearing perforin positive granules, which polarized upon interaction of the CTLs with anti-CD3 coated beads. However, rapid anti-CD3-induced granule secretion was drastically defective in both CD8(+) and CD4(+) T cells from ashen mice. This defect in exocytosis was not observed in the constitutive pathway, as T cell receptor stimulated interferon-gamma secretion was normal. Based on these results and our demonstration that Rab27a colocalizes with granzyme B-positive granules and is undetectable in ashen CTLs, we conclude that Rab27a is required for a late step in granule exocytosis, compatible with current models of Rab protein function in vesicle docking and fusion. PMID- 11266475 TI - An essential role for the substrate-binding region of Hsp40s in Saccharomyces cerevisiae. AB - In addition to regulating the ATPase cycle of Hsp70, a second critical role of Hsp40s has been proposed based on in vitro studies: binding to denatured protein substrates, followed by their presentation to Hsp70 for folding. However, the biological importance of this model is challenged by the fact that deletion of the substrate-binding domain of either of the two major Hsp40s of the yeast cytosol, Ydj1 and Sis1, leads to no severe defects, as long as regions necessary for Hsp70 interaction are retained. As an in vivo test of this model, requirements for viability were examined in a strain having deletions of both Hsp40 genes. Despite limited sequence similarity, the substrate-binding domain of either Sis1 or Ydj1 allowed cell growth, indicating they share overlapping essential functions. Furthermore, the substrate-binding domain must function in cis with a functional Hsp70-interacting domain. We conclude that the ability of cytosolic Hsp40s to bind unfolded protein substrates is an essential function in vivo. PMID- 11266474 TI - Rab27a: A key to melanosome transport in human melanocytes. AB - Normal pigmentation depends on the uniform distribution of melanin-containing vesicles, the melanosomes, in the epidermis. Griscelli syndrome (GS) is a rare autosomal recessive disease, characterized by an immune deficiency and a partial albinism that has been ascribed to an abnormal melanosome distribution. GS maps to 15q21 and was first associated with mutations in the myosin-V gene. However, it was demonstrated recently that GS can also be caused by a mutation in the Rab27a gene. These observations prompted us to investigate the role of Rab27a in melanosome transport. Using immunofluorescence and immunoelectron microscopy studies, we show that in normal melanocytes Rab27a colocalizes with melanosomes. In melanocytes isolated from a patient with GS, we show an abnormal melanosome distribution and a lack of Rab27a expression. Finally, reexpression of Rab27a in GS melanocytes restored melanosome transport to dendrite tips, leading to a phenotypic reversion of the diseased cells. These results identify Rab27a as a key component of vesicle transport machinery in melanocytes. PMID- 11266476 TI - Where Notch and Wnt signaling meet. The presenilin hub. PMID- 11266477 TI - Of yeast, mice, and men. Rab proteins and organelle transport. PMID- 11266478 TI - Mechanical treatment of plantar fasciitis. A prospective study. AB - A randomized, prospective study was conducted to compare the effectiveness of three individual mechanical modalities in the treatment of plantar fasciitis. Two hundred fifty-five subjects were randomly assigned to one of three treatment groups: custom-made orthoses, over-the-counter arch supports, or tension night splints. Subjects were treated for 3 months, with follow-up visits at 2, 6, and 12 weeks. No statistically significant difference was noted among treatment groups with respect to final outcomes based on first-step pain or pain felt during the day. However, there was a statistically significant difference among the three groups with respect to early patient withdrawal from the study due to continued severe pain, noncompliance, or inability to tolerate the device. Patient compliance was greatest with the use of custom-made orthoses. PMID- 11266479 TI - Peg-in-hole, end-to-end, and V arthrodesis. A comparison of digital stabilization in fresh cadaveric specimens. AB - The proximal interphalangeal joint arthrodesis is frequently performed to correct hammer toe deformities. This study was conducted to compare the inherent stability of the three proximal interphalangeal joint arthrodeses--peg-in-hole, end-to-end, and V constructs--in the sagittal plane by means of load-to-failure testing of 30 fresh-frozen cadaveric specimens fixated with a 0.045 Kirschner wire. The peg-in-hole construct was associated with significantly higher peak loads at failure compared with the other two procedures. Furthermore, the peg-in hole construct had significantly higher stiffness values as compared with the V procedure. This study thus provides evidence that the peg-in-hole procedure is the most biomechanically stable surgical construct for proximal interphalangeal joint fusions under sagittal plane loading. PMID- 11266480 TI - Scientific approach to the axis of rotation at the midtarsal joint. AB - Current biomechanical models of the midtarsal joint describe it as having two axes of rotation, the oblique and the longitudinal. The considerable freedom of movement available at the midtarsal joint means that kinematic assessment of its function and determination of its axis of rotation must be conducted under conditions that enable the joint to function as normally as possible. The assessments on which the concepts of the longitudinal and oblique axes are based do not meet this criterion. Understanding of the motions at the midtarsal joint will improve as techniques of kinematic assessment improve. Future descriptions of the midtarsal joint should adopt the standard terms applied to the other joints in the lower limb, which will facilitate the study of the midtarsal joint in relation to the function of the rest of the lower limb. PMID- 11266481 TI - The grading of hallux valgus. The Manchester Scale. AB - This article describes a new, noninvasive method of assessing the severity of hallux valgus deformity by means of a set of standardized photographs. Six podiatrists were independently asked to grade the level of deformity of 13 subjects (26 feet) on a scale of 1 (no deformity) to 4 (severe deformity). The reliability of the four-point scale for the severity of hallux valgus was investigated by means of kappa-type statistics for more than two raters. The results showed that the grading method had excellent interobserver repeatability with a combined kappa-type statistic of 0.86, making it a suitable instrument for clinical and research purposes. PMID- 11266483 TI - Isolated cuboid fracture. A rare occurrence. AB - Solitary fractures of the cuboid are rare. Proper diagnosis and treatment of subtle cuboid fractures are essential to minimize long-term disability and must begin with the appropriate clinical suspicion. Treatment of overt cuboid fractures has changed over time and now includes aggressive repair of the fracture. The authors provide an overview of cuboid fractures and report on a patient with a minimally displaced fracture who was treated conservatively. PMID- 11266482 TI - Plantar verrucae in patients with human immunodeficiency virus. Clinical presentation and treatment response. AB - Several previous studies have yielded data showing that plantar and other cutaneous verrucae follow a more aggressive course in patients infected with human immunodeficiency virus (HIV) than in uninfected individuals. A pilot study was undertaken to identify trends in a sample population that would support this characterization of plantar verrucae in HIV+ patients and to determine whether there are differences in treatment response between HIV+ and HIV- patients. The results show that the HIV+ patients in the study presented with a significantly greater number and total area of lesions than did the HIV- patients. Furthermore, the HIV+ patients experienced a greater frequency of recurrence of their lesions following treatment with surgical curettage. These findings should provide the foundation for other extensive, multicenter studies to further characterize the treatment response of these lesions in HIV+ patients and to develop effective guidelines for their management. PMID- 11266485 TI - Venous aneurysm of the dorsal venous arch. AB - Clinically, a venous aneurysm is seen as a nonpulsatile mass with shrinkage upon elevation and enlargement with dependency. Confirmation of the diagnosis is best made by venography. In the case presented here of a rare aneurysm of the dorsal venous arch, the diagnosis of venous aneurysm was based on the histopathologic and intraoperative findings. If a venous aneurysm is symptomatic, it should be excised, with ligation of all feeder veins. In all cases, accurate preoperative diagnosis and evaluation allows for appropriate surgical planning. PMID- 11266484 TI - Atypical presentation of plantar fasciitis secondary to soft-tissue mass infiltration. AB - This article describes a patient with plantar fascial pain who presented to the office of one of the authors. Physical examination and the patient's description of the history of symptoms revealed classic signs and symptoms of plantar fasciitis. The patient was treated with numerous conservative modalities, including ultrasound, nonsteroidal anti-inflammatory medications, trigger-point injections, over-the-counter orthoses, and stretching exercises. When the pain was not relieved by these conservative measures, magnetic resonance imaging of the area was performed. Visualization of the insertional area of the plantar fascia revealed a mass inferior to, as well as infiltrated into, the plantar fascia. Surgical excision of the lesion resulted in complete elimination of the patient's pain. PMID- 11266486 TI - Hemangioma of the foot. PMID- 11266487 TI - Stress fracture of the foot secondary to osteoporosis: an atypical presentation. PMID- 11266488 TI - The case for multiple database searching in podiatric medicine. PMID- 11266489 TI - Manipulation method for the treatment of ankle equinus. PMID- 11266492 TI - Molecular quantification of human beta-glucuronidase levels in a patient with Vohwinkel's syndrome. AB - The skin must undergo the process of keratinization in order to perform its functions. During the process of differentiation, certain genes are activated while others are repressed, leading to changes in structural proteins and enzymes and in the synthesis of various lipids. An error in any of these steps can ultimately impair the process of keratinization. Vohwinkel's syndrome is the direct result of a defect in keratinization. Patients who have this epidermolytic palmoplantar keratoderma present clinically with hyperkeratosis of the stratum corneum. Hyperkeratosis has been linked to an increase in beta-glucuronidase levels. The authors studied the absolute concentration of human beta glucuronidase in a patient with Vohwinkel's syndrome as determined through a double-antibody sandwich enzyme-linked immunosorbent assay and a Western blot assay of the blood, urine, and skin of the patient. PMID- 11266491 TI - Pain at the site of tarsal tunnel incision due to neuroma of the posterior branch of the saphenous nerve. AB - The authors conducted a retrospective review of 16 patients who presented with the complaint of pain at the incision site after tarsal tunnel decompression. Specifically, the pain was located at the proximal aspect of the tarsal tunnel decompression scar. The mean duration of pain was 21 months (range, 6 to 34 months). The pain was eliminated by a block of the distal saphenous nerve, demonstrating that the pain was due to a neuroma of this nerve. The pain was treated by resection of the distal saphenous nerve in the distal leg and implantation of the proximal end of this nerve into the soleus muscle. At a mean of 18.5 months after surgery (range, 6 to 33 months), excellent relief of pain was achieved in 76% of cases and good relief of pain in 24% of cases. PMID- 11266493 TI - The reliability of clinical and caliper-based calcaneal bisection measurements. AB - The measurements of subtalar joint neutral position and hindfoot range of motion have been shown to be unreliable. The first step in making these measurements is to determine the calcaneal bisection. This study examines the reliability of bisecting the calcaneus with digital linear calipers. Five trials on each of six cadavers resulted in a mean absolute angular difference of 0.60 degree (SD +/- 1.17 degrees). These results were then compared with results from the typical visual method used clinically. Three raters each performed five trials on six cadavers. Visual bisection was more variable, with a mean absolute error of 3.61 degrees (+/- 3.13 degrees). A mean error of 6 degrees (+/- 1 degree) is certainly possible when the heel is visually bisected. It was determined that the caliper bisection was a valid technique for bisection of the heel, but that clinical visual bisection was not. PMID- 11266494 TI - Efficacy of terbinafine for toenail onychomycosis. A multicenter trial of various treatment durations. AB - The efficacy of terbinafine (250 mg/day) in the treatment of toenail onychomycosis was evaluated in a large open-label, multicenter trial of 12, 18, and 24 weeks of therapy. All 1,534 patients had onychomycosis, confirmed by either positive potassium hydroxide (KOH) wet mount, positive fungal culture, or both, and all received at least 12 weeks of treatment. Treatment was continued for an additional 6 or 12 weeks, depending on the extent of the disease at follow up. Mycologic cure rates (negative culture plus negative KOH) at week 72 were 72.1% in the 12-week treatment group, 72.5% in the 18-week group, and 77.0% in the 24-week group. In all groups, clinical cure rates were higher at week 72 than at week 48: 49.5% of the 12-week group, 49.2% of the 18-week group, and 44.6% of the 24-week group experienced clinical cure by the end of the study. Both mycologic and clinical recurrence rates were low in all treatment groups at the 72-week assessment. The results of this study confirm the efficacy of terbinafine in the treatment of toenail onychomycosis as demonstrated in previous registration and large-scale clinical trials. PMID- 11266495 TI - Effect of biofeedback-assisted relaxation training on foot ulcer healing. AB - A prospective, randomized study was conducted to determine the effect of biofeedback-assisted relaxation training on foot ulcer healing. For patients with chronic nonhealing foot ulcers, medical care was combined with a standardized biofeedback-assisted relaxation training program in the experimental group. The intervention was designed to increase peripheral perfusion, thereby promoting healing. A total of 32 patients with chronic nonhealing ulcers participated in the study. In the experimental group, 14 out of 16 ulcers (87.5%) healed, as compared with 7 out of 16 ulcers (43.8%) in the control group. PMID- 11266497 TI - Bilateral forefoot gangrene secondary to Lemierre's disease. PMID- 11266496 TI - Resection of a plantar calcaneal spur using the holmium:yttrium-aluminum-garnet (Ho:YAG) laser. AB - Many procedures have been described for the resection of plantar calcaneal spurs as treatment of heel spur syndrome and chronic plantar fasciitis. Most of these techniques involve a medial incision of between 2 and 6 cm for adequate exposure of the calcaneal spur. This article describes a new technique for resecting a calcaneal spur with a smaller medial incision using the holmium:yttrium-aluminum garnet (Ho:YAG) laser. This laser permits adequate resection of a plantar calcaneal spur as well as coagulation of the bone and surrounding tissues. This minimally invasive procedure has been used with good results over the past year by the senior author (W.K.S.) for the resection of calcaneal spurs. PMID- 11266498 TI - Smith-Lemli-Opitz syndrome: a genetic disorder with pedal manifestations. PMID- 11266499 TI - Activity monitors: should we begin dosing activity as we dose a drug? PMID- 11266500 TI - Council on Podiatric Medical Education Eighty-First Annual Report, 2000. PMID- 11266503 TI - Regulation of glucocorticoid receptor activity by 14--3-3-dependent intracellular relocalization of the corepressor RIP140. AB - Proteins belonging to the 14--3-3 family interact with various regulatory proteins involved in cellular signaling, cell cycle regulation, or apoptosis. 14- 3-3 proteins have been suggested to act by regulating the cytoplasmic/nuclear localization of their target proteins or by acting as molecular scaffolds or chaperones. We have previously shown that overexpression of 14--3-3 enhances the transcriptional activity of the glucocorticoid receptor (GR), which is a member of the nuclear receptor family. In this study, we show that 14--3-3 interacts with the nuclear receptor corepressor RIP140. In transfection assays, RIP140 antagonizes 14--3-3- enhanced GR transactivation. Using colocalization studies we demonstrate that 14--3-3 can export RIP140 out of the nucleus and, interestingly, can also change its intranuclear localization. Moreover, we also observed that 14 -3-3 can bind various other nuclear receptors and cofactors. In summary, our findings suggest that 14--3-3-mediated intracellular relocalization of the GR corepressor RIP140 might be a novel mechanism to enhance glucocorticoid responsiveness of target genes. They furthermore indicate a more general role for 14--3-3 protein by influencing the nuclear availability of nuclear receptor associated cofactors. PMID- 11266502 TI - The glucocorticoid receptor interacting protein 1 (GRIP1) localizes in discrete nuclear foci that associate with ND10 bodies and are enriched in components of the 26S proteasome. AB - The glucocorticoid receptor interacting protein-1 (GRIP1) is a member of the steroid receptor coactivator (SRC) family of transcriptional regulators. Green fluorescent protein (GFP) fusions were made to full-length GRIP1, and a series of GRIP1 mutants lacking the defined regulatory regions and the intracellular distribution of these proteins was studied in HeLa cells. The distribution of GRIP1 was complex, ranging from diffuse nucleoplasmic to discrete intranuclear foci. Formation of these foci was dependent on the C-terminal region of GRIP1, which contains the two characterized transcriptional activation domains, AD1 and AD2. A subpopulation of GRIP1 foci associate with ND10s, small nuclear bodies that contain several proteins including PML, SP100, DAXX, and CREB-binding protein (CBP). Association with the ND10s is dependent on the AD1 of GRIP1, a region of the protein previously described as a CBP-interacting domain. The GRIP1 foci are enriched in components of the 26S proteasome, including the core 20S proteasome, PA28alpha, and ubiquitin. In addition, the irreversible proteasome inhibitor lactacystin induced an increase in the total fluorescence intensity of the GFP-GRIP1 expressing cells, demonstrating that GRIP1 is degraded by the proteasome. These findings suggest the intriguing possibility that degradation of GRIP1 by the 26S proteasome may be a key component of its regulation. PMID- 11266504 TI - Nucleocytoplasmic shuttling of the thyroid hormone receptor alpha. AB - The thyroid hormone receptor alpha (TR alpha) exhibits a dual role as an activator or repressor of gene transcription in response to thyroid hormone (T(3)). Our studies show that TR alpha, formerly thought to reside solely in the nucleus tightly bound to DNA, actually shuttles rapidly between the nucleus and cytoplasm. The finding that TR alpha shuttles reveals an additional checkpoint in receptor control of gene expression. Using Xenopus oocyte microinjection assays, we show that there are two coexisting mechanisms for nuclear entry of TR alpha. First, nuclear import of TR alpha (molecular mass 46 kDa) was not sensitive to general inhibitors of signal-mediated transport, indicating that TR alpha can enter the oocyte nucleus by passive diffusion. Second, when TR alpha was tagged with glutathione-S:-transferase, import of the fusion protein (molecular mass 73 kDa) was completely blocked by these inhibitors, demonstrating that an alternative, signal-mediated import pathway exists for TR alpha. Nuclear retention of TR alpha in oocytes is enhanced in the presence of T(3), suggesting that more intranuclear binding sites are available for the ligand-bound receptor. Using mammalian cells, we show that shuttling of green fluorescent protein (GFP) tagged and untagged TR alpha is inhibited in both chilled and energy-depleted cells, suggesting that there is an energy-requiring step in the nuclear retention/export process. Nuclear export of TR alpha is not blocked by leptomycin B, a specific inhibitor of the export receptor CRM1, indicating that TR alpha does not require the CRM1 pathway to exit the nucleus. Dominant negative mutants of TR with defects in DNA binding and transactivation accumulate in the cytoplasm at steady state, illustrating that even single amino acid changes in functional domains may alter the subcellular distribution of TR. In contrast to TR alpha, nuclear export of its oncogenic homolog v-ErbA is sensitive to leptomycin B, suggesting that the oncoprotein follows a CRM1-mediated export pathway. Acquisition of altered nuclear export capabilities may contribute to the oncogenic properties of v-ErbA. PMID- 11266505 TI - Luteinizing hormone receptors are self-associated in slowly diffusing complexes during receptor desensitization. AB - We have previously shown that rat LH receptors (LHRs) occupied by human CG (hCG) exhibit slow receptor lateral diffusion and are self-associated. Here we have examined whether LHRs become self-associated and enter slowly diffusing structures in response to hormone binding and whether these receptors retain this organization while in the desensitized state. Before hormone exposure, wild-type rat LHRs coupled at the C terminus to enhanced green fluorescent protein (GFP-LHR wt) exhibited fast lateral diffusion, as assessed by fluorescent photobleaching recovery (FPR) methods, and most receptors were laterally mobile. After 30 min exposure to hCG and subsequent removal of hormone by low pH wash, hormone challenge at any time within the next 4 h produced no increase in cellular cAMP levels. During this time, LHRs were either laterally immobile or exhibited slower lateral diffusion. When LHRs were again responsive to binding of hormone, the rate of receptor lateral diffusion had become significantly faster and the fraction of mobile receptors was again large. Desensitized LHRs were also self associated and present in microscopically visible clusters on the plasma membrane. Fluorescence energy transfer (FET) methods were used to measure the extent of interaction between receptors coupled to either GFP or to yellow fluorescent protein (YFP). Before hormone treatment, there was essentially no energy transfer between LHRs. After desensitization of the receptors by 30 min exposure to hCG, energy transfer efficiency increased to 18%. Values for FET efficiency between desensitized receptors decreased over time, and receptors were responsive to hormone only after measurable energy transfer had completely disappeared. Together these results suggest that desensitized LHRs exist in large, slowly diffusing structures containing self-associated receptors and that these structures must dissipate before the receptor can again respond to hormone. PMID- 11266506 TI - Synergistic activation of the serotonin-1A receptor by nuclear factor-kappa B and estrogen. AB - Estrogen exerts profound effects on mood and mental state. The ability of estrogen to modulate serotonergic function raises the possibility that it may play a role in the mechanism associated with depression and its treatment. A cellular mechanism for estrogen to influence mood might be through the regulation of genes involved at various levels of the serotonin system. Here we report that estrogen can up-regulate the expression of the serotonin-1A receptor via a new mechanism involving synergistic activation by nuclear factor-kappa B (NF-kappa B) with estrogen receptor alpha. Interestingly, we observed that only estrogen receptor-alpha, and not -beta, was able to mediate this effect of estrogens. The partial antiestrogen, 4-hydroxytamoxifen, had the same effect as estrogen. In addition, mutation analysis showed that both the transactivation function of p65 and activation function 1 of estrogen receptor-alpha were essential for this synergistic regulation. Therefore, we propose that NF-kappa B complexes cooperate with estrogen receptor-alpha to recruit cofactors into the complex and thereby synergistically activate the serotonin-1A receptor promoter through nonclassical estrogen response elements by a mechanism that does not involve direct receptor binding to DNA. PMID- 11266507 TI - Beta(2)-adrenergic receptors potentiate glucocorticoid receptor transactivation via G protein beta gamma-subunits and the phosphoinositide 3-kinase pathway. AB - Glucocorticoid hormones influence manifold neuronal processes including learning, memory, and emotion via the glucocorticoid receptor (GR). Catecholamines further modulate these functions, although the underlying molecular mechanisms are poorly understood. Here, we show that epinephrine and norepinephrine potentiate ligand dependent GR transactivation in a hippocampal cell line (HT22) via beta(2) adrenergic receptors. This enhancement was strongest at low concentrations of glucocorticoids and was accompanied by increased GR binding to a glucocorticoid responsive element (GRE). beta(2)-Adrenergic receptor-mediated GR enhancement was relayed via G protein beta gamma-subunits, insensitive to pertussis toxin and independent of protein kinase A (PKA). In contrast, the catecholamine-evoked GR enhancement was strongly reduced by wortmannin, suggesting a critical role for phosphoinositide 3-kinase (PI3-K). In agreement, epinephrine directly activated PI3-K in vivo. Similarly, stimulation of tyrosine kinase receptors coupled to PI3 K activation, e.g. receptors for insulin-like growth factor I (IGF-I) or fibroblast growth factor (FGF), increased GR transactivation. Further analysis indicated that G protein-coupled receptor (GPCR) and tyrosine kinase receptor signals converge on PI3-K through separate mechanisms. Blockade of GR enhancement by wortmannin was partially overcome by expression of the downstream-acting protein kinase B (PKB/Akt). Collectively, our findings demonstrate that GPCRs can regulate GR transactivation by stimulating PI3-K. This novel cross-talk may provide new insights into the molecular processes of learning and memory and the treatment of stress-related disorders. PMID- 11266508 TI - Insulin induces specific interaction between insulin receptor and protein kinase C delta in primary cultured skeletal muscle. AB - Certain protein kinase C (PKC) isoforms, in particular PKCs beta II, delta, and zeta, are activated by insulin stimulation. In primary cultures of skeletal muscle, PKCs beta II and zeta, but not PKC delta, are activated via a phosphatidylinositol 3-kinase (PI3K)-dependent pathway. The purpose of this study was to investigate the possibility that PKC delta may be activated upstream of PI3K by direct interaction with insulin receptor (IR). Experiments were done on primary cultures of newborn rat skeletal muscle, age 5--6 days in vitro. The time course of insulin-induced activation of PKC delta closely paralleled that of IR. Insulin stimulation caused a selective coprecipitation of PKC delta with IR, and these IR immunoprecipitates from insulin-stimulated cells displayed a striking induction of PKC activity due specifically to PKC delta. To examine the involvement of PKC delta in the IR signaling cascade, we used recombinant adenovirus constructs of wild-type (W.T.) or dominant negative (D.N.) PKC delta. Overexpression of W.T.PKC delta induced PKC delta activity and coassociation of PKC delta and IR without addition of insulin. Overexpression of D.N.PKC delta abrogated insulin- induced coassociation of PKC delta and IR. Insulin-induced tyrosine phosphorylation of IR was greatly attenuated in cells overexpressing W.T.PKC delta, whereas in myotubes overexpressing D.N.PKC delta, tyrosine phosphorylation occurred without addition of insulin and was sustained longer than that in control myotubes. In control myotubes IR displayed a low level of serine phosphorylation, which was increased by insulin stimulation. In cells overexpressing W.T.PKC delta, serine phosphorylation was strikingly high under basal conditions and did not increase after insulin stimulation. In contrast, in cells overexpressing D.N.PKC delta, the level of serine phosphorylation was lower than that in nonoverexpressing cells and did not change notably after addition of insulin. Overexpression of W.T.PKC delta caused IR to localize mainly in the internal membrane fractions, and blockade of PKC delta abrogated insulin-induced IR internalization. We conclude that PKC delta is involved in regulation of IR activity and routing, and this regulation may be important in subsequent steps in the IR signaling cascade. PMID- 11266509 TI - The alpha-subunit of the epithelial sodium channel is an aldosterone-induced transcript in mammalian collecting ducts, and this transcriptional response is mediated via distinct cis-elements in the 5'-flanking region of the gene. AB - Aldosterone stimulates Na(+) reabsorption in the collecting ducts by increasing the activity of the epithelial sodium channel, ENaC. Systemic administration of aldosterone increases alpha ENaC mRNA expression in mammalian kidney, suggesting that the alpha ENaC gene is a target for aldosterone action in the distal nephron. To determine whether aldosterone increases alpha ENaC gene transcription, a portion of the alpha ENaC 5'- flanking region coupled to luciferase was transfected into MDCK-C7 cells, a collecting duct cell line with aldosterone-stimulated Na(+) transport. Both dexamethasone and aldosterone stimulated alpha ENaC-coupled reporter gene activity via the glucocorticoid receptor (GR), and this response correlated with the effect of these hormones on endogenous alpha ENaC expression. The aldosterone-stimulated alpha ENaC expression was blocked by actinomycin D, and aldosterone had no effect on alpha ENaC mRNA decay, confirming a transcriptional effect. In HT-29 cells, a GR/mineralocorticoid receptor (MR)-deficient colonic cell line with constitutive alpha ENaC expression, cotransfection with GR or MR restored aldosterone stimulated alpha ENaC gene transcription, although aldosterone had a functional preference for MR. Analysis of deletion constructs confirmed that a single imperfect glucocorticoid response element (GRE) is necessary and sufficient to confer the aldosterone responsiveness to the alpha ENaC gene promoter in MDCK-C7 and HT-29 cells. These results confirm that alpha ENaC is an aldosterone-induced transcript in the collecting duct and delineates the molecular mechanism for this effect. PMID- 11266510 TI - The endogenous fibroblast growth factor-2 antisense gene product regulates pituitary cell growth and hormone production. AB - Basic fibroblast growth factor (bFGF; FGF-2) is one of 19 related members of a growth factor family with mitogenic and hormone-regulatory functions. In Xenopus laevis oocytes, a 1.5-kb FGF-2 antisense (GFG) RNA complementary to the third exon and 3'-untranslated region (UTR) of FGF-2 mRNA has been implicated in FGF-2 mRNA editing and stability. The human homolog has been cloned, and we localized this gene by yeast artificial chromosome (YAC), somatic cell, and radiation hybrid panels to the same chromosomal site as FGF-2 (chromosome 4, JO4513 adjacent to D4S430), confirming this as a human endogenous antisense gene. The full-length GFG antisense RNA encodes a 35-kDa protein, which is highly homologous with the MutT family of antimutator nucleosidetriphosphatases (NTPases). We show that human pituitary tumors express FGF-2 and its endogenous antisense partner GFG. While normal pituitary expresses GFG but not FGF-2, pituitary adenomas express FGF-2 and have reduced levels of GFG; aggressive and recurrent adenomas expressed more FGF than GFG mRNA. To examine the effects of this antisense gene in the pituitary, we transfected the pituitary-derived GH4 mammosomatotroph cell line with constructs encoding the full-length human GFG cDNA. Transiently and stably transfected cells expressed the 35-kDa GFG protein that was localized to the cytoplasm. These cells exhibited enhanced PRL expression as documented by transiently transfected PRL-luciferase reporter assay and by endogenous PRL protein. GFG expression in these cells did not alter endogenous FGF-2 expression but increased the proportion of the higher molecular mass 22-kDa form of GH. Moreover, GFG expression inhibited cell proliferation as shown by [(3)H]thymidine incorporation, proliferating cell nuclear antigen (PCNA) nuclear staining, and cell cycle analysis. We conclude that the GFG-encoded protein has divergent hormone-regulatory and antiproliferative actions in the pituitary that are independent of FGF-2 expression. GFG represents a novel mechanism involved in restraining pituitary tumor cell growth while promoting hormonal activity. PMID- 11266511 TI - Stage-sensitive blockade of pituitary somatomammotrope development by targeted expression of a dominant negative epidermal growth factor receptor in transgenic mice. AB - The epidermal growth factor receptor (EGFR) and its ligands EGF and transforming growth factor-alpha (TGF alpha) are expressed in the anterior pituitary, and overexpression of TGF alpha in the lactotrope cells of the pituitary gland in transgenic mice results in lactotrope hyperplasia and adenomata, suggesting a role for EGFR signaling in pituitary cell proliferation. To address the role of EGFR signaling in pituitary development in vivo, we blocked EGFR signaling in transgenic mice using the dominant negative properties of a mutant EGFR lacking an intracellular protein kinase domain (EGFR-tr). We directed EGFR-tr expression to GH- and PRL- producing cells using GH and PRL promoters, and a tetracycline inducible gene expression system, to allow temporal control of gene expression. EGFR-tr overexpression in GH-producing cells during embryogenesis resulted in dwarf mice with pituitary hypoplasia. Both somatotrope and lactotrope development were blocked. However, when EGFR-tr overexpression was delayed to the postnatal period either by directing its expression with the PRL promoter or by delaying the onset of induction with tetracycline in the GH cells, no specific phenotype was observed. Lactotrope hyperplasia during pregnancy also occurred normally in the PRL-EGFR-tr mice. These data suggest that EGFR signaling is required for the differentiation and/or maintenance of somatomammotropes early in pituitary organogenesis but not later in life. (Molecular Endocrinology 15: 600-613, 2001) PMID- 11266512 TI - Analysis of the role of the mitogen-activated protein kinase in mediating cyclic adenosine 3',5'-monophosphate effects on prolactin promoter activity. AB - The mechanisms mediating cAMP effects to stimulate transcription of the PRL gene have been examined. Treatments that elevate intracellular cAMP concentrations were found to stimulate the mitogen-activated protein kinase (MAPK) in GH(3) cells. Elevated cAMP was also found to stimulate activation of the GTP-binding protein, Rap1. Rap1GAP1 reduced cAMP-induced phosphorylation of MAPK, offering evidence that Rap1 may play a role in mediating activation of MAPK. Treatment of GH(3) cells with PD98059, an inhibitor of the MAPK pathway, reduced the ability of forskolin to activate a PRL reporter gene, providing evidence that MAPK contributes to cAMP-mediated effects on the PRL promoter. As previous studies have implicated Ets factor binding sites within the PRL promoter in mediating responses to MAPK, we expected that the Ets sites would also play a role in cAMP responsiveness. Surprisingly, mutation of all of the consensus Ets factor binding sites in the proximal PRL promoter greatly reduced responsiveness to epidermal growth factor (EGF) and TRH but did not reduce cAMP responsiveness. Experiments using an expression vector for adenovirus 12S E1a provided evidence that the coactivators, CREB binding protein and/or p300, probably play a role in cAMP responsiveness of the PRL promoter. Interestingly, the ability of a GAL4-p300 fusion protein to enhance reporter gene activity was stimulated by cAMP in a MAPK dependent manner. These findings provide evidence for a model for cAMP-induced PRL transcription involving Rap1-induced MAPK activity leading to stimulation of the transcriptional coactivators, CBP and p300. PMID- 11266513 TI - Inhibition of protein phosphatase PP1 in GH3B6, but not in GH3 cells, activates the MEK/ERK/c-fos pathway and the human prolactin promoter, involving the coactivator CPB/p300. AB - The human (hPRL) PRL gene proximal promoter (-164/+15) is the target for numerous signal transduction pathways involving protein kinases. The inhibitor of Ser/Thr protein phosphatases okadaic acid (OA) was shown to induce this promoter in rat pituitary GH3B6 through a synergism between increased amounts of the ubiquitous factor AP-1 and the pituitary-specific factor Pit-1. Here we show that this activation results mainly from transcriptional stimulation of the c-fos promoter leading to increased AP-1 activity. We report the surprising absence of the hPRL and c-fos promoter stimulation by OA in GH3 cells, closely related to GH3B6 cells, and we use this discrepancy to dissect the precise mechanism of action. c fos gene activation involves the mitogen-activated kinase (MAPK)-ternary complex factor (TCF) pathway and can be obtained by expressing active V12ras in both cell lines. We show that OA acts by inhibiting protein phosphatase PP1, thereby protecting MAPK kinase (MEK)1/2 and/or a MEK1/2-kinase from dephosphorylation. PP1 inhibition of MEK activation by V12ras does not occur in GH3 cells, indicating that a distinct, PP1-sensitive phosphorylation site is used in GH3B6 cells to activate the TCF pathway in GH3B6 cells. Finally, we show that the synergistic OA activation of the hPRL promoter by Pit-1 and AP-1 is independent of the Pit-1 transactivation domain and is mediated by the general coactivator (CRE-binding protein)-binding protein (CBP)/p300. PMID- 11266514 TI - Identification of a decidua-specific enhancer on the human prolactin gene with two critical activator protein 1 (AP-1) binding sites. AB - Deletion analysis of the human PRL promoter in endometrial stromal cells decidualized in vitro revealed a 536-bp enhancer located between nucleotide (nt) 2,040 to -1,505 in the 5'-flanking region. The 536-bp enhancer fragment ligated into a thymidine kinase (TK) promoter-luciferase reporter plasmid conferred enhancer activity in decidual-type cells but not nondecidual cells. DNase I footprint analysis of decidualized endometrial stromal cells revealed three protected regions, FP1-FP3. Transfection of overlapping 100-bp fragments of the 536-bp enhancer indicated that FP1 and FP3 each conferred enhancer activity. Gel shift assays indicated that both FP1 and FP3 bind activator protein 1 (AP-1), and JunD and Fra-2 are components of the AP-1 complex in decidual fibroblasts. Mutation of the AP-1 binding site in either FP1 or FP3 decreased enhancer activity by approximately 50%, while mutation of both sites almost completely abolished activity. Coexpression of the 536-bp enhancer and A-fos, a dominant negative to AP-1, decreased enhancer activity by approximately 70%. Conversely, coexpression of Fra-2 in combination with JunD or c-Jun and p300 increased enhancer activity 6- to 10-fold. Introduction of JunD and Fra-2 into nondecidual cells is sufficient to confer enhancer activity. JunD and Fra-2 protein expression was markedly increased in secretory phase endometrium and decidua of early pregnancy (high PRL content) compared with proliferative phase endometrium (no PRL). These investigations indicate that the 5'-flanking region of the human PRL gene contains a decidua-specific enhancer between nt -2,040/-1,505 and AP-1 binding sites within this enhancer region are critical for activity. PMID- 11266515 TI - Inhibin binding protein in rats: alternative transcripts and regulation in the pituitary across the estrous cycle. AB - Inhibin binding protein (InhBP) and the transforming growth factor-beta (TGF beta) type III receptor, beta glycan, have been identified as putative inhibin coreceptors. Here we cloned the InhBP cDNA in rats and predict that it encodes a large membrane-spanning protein that is part of the Ig superfamily, as has been described for humans. Two abundant InhBP transcripts (4.4 and 1.8 kb) were detected in the adult rat pituitary. The larger transcript encodes the full length protein while the 1.8-kb transcript (InhBP-short or InhBP-S) corresponds to a splice variant of the receptor. This truncated isoform contains only the N terminal signal peptide and first two (of 12) Ig-like domains observed in the full-length InhBP (InhBP-long or InhBP-L). InhBP-S does not contain a transmembrane domain and is predicted to be a soluble protein. Beta glycan was also detected in the pituitary; however, it was most abundant within the intermediate lobe. Although we also observed beta glycan immunopositive cells in the anterior pituitary, they rarely colocalized with FSH beta-producing cells. We next examined physiological regulation of the coreceptors across the rat estrous cycle. Like circulating inhibin A and inhibin B levels, pituitary InhBP-L and InhBP-S mRNA levels were dynamically regulated across the cycle and were negatively correlated with serum FSH levels. Expression of both forms of InhBP was also positively correlated with serum inhibin B, but not inhibin A, levels. These data are particularly interesting in light of our in vitro observations that InhBP may function as an inhibin B-specific coreceptor. Pituitary beta glycan mRNA levels did not fluctuate across the cycle nor did they correlate with serum FSH. These observations, coupled with its pattern of expression within the pituitary, indicate that beta glycan likely functions as more than merely an inhibin coreceptor within the pituitary. A direct role for InhBP or beta glycan in regulation of pituitary FSH by inhibin in vivo has yet to be determined, but the demonstration of dynamic regulation of pituitary InhBP and its negative relation to serum FSH across the estrous cycle is an important step in this direction. PMID- 11266516 TI - Modulation of activin signal transduction by inhibin B and inhibin-binding protein (INhBP). AB - An antagonistic relationship between inhibin and activin is essential to the control of pituitary FSH release and to normal gonadal function. Two inhibin ligands, inhibin A and inhibin B, are made by the ovary in females, and each regulate pituitary FSH at different times during the reproductive cycle. Inhibin B, but not inhibin A, is produced by the testes and is therefore responsible for all inhibin-dependent FSH regulation in males. Although the activin signal transduction pathway has been well characterized, little is known about the mechanism of inhibin signaling and its relationship to activin antagonism. A recently cloned inhibin-binding protein, InhBP (p120), associates strongly with the type IB activin receptor (Alk4) in a ligand-responsive manner and interacts to a lesser extent with other activin and bone morphogenetic protein (BMP) type I and activin type II receptors. Activin stimulates the association of InhBP and Alk4, and inhibin B, but not inhibin A, interferes with InhBP-Alk4 complex formation. InhBP is necessary to mediate a specific antagonistic effect of inhibin B on activin-stimulated transcription. Appropriate stoichiometry between InhBP and the activin type I receptor is crucial to InhBP function. These findings suggest that InhBP is an inhibin B-specific receptor that mediates antagonism of activin signal transduction through the modulation of activin heteromeric receptor complex assembly. PMID- 11266517 TI - Neoplasms of the ampulla of vater with concurrent pancreatic intraductal neoplasia: a histological and molecular study. AB - Adenoma and adenocarcinoma of the ampulla of Vater are uncommon neoplasms of the gastrointestinal tract. Only one report has analyzed the relationship between ampullary adenocarcinoma and pancreatic intraductal neoplasia (PanIN), the precursor lesion of pancreatic adenocarcinoma. An association between PanIN and ampullary adenoma has not been reported previously. Case reports have documented the progression of PanIN to invasive pancreatic adenocarcinoma. We reviewed five resected ampullary adenoma and 17 ampullary adenocarcinoma cases and evaluated the pancreas for PanIN. Pancreatic sections from 35 autopsies were reviewed as a control group. Immunohistochemistry for overexpression of p53 and COX-2 proteins was performed in selected cases, as was PCR analysis for K-ras mutations. Follow up clinical data were obtained. All 22 ampullary neoplasms were associated with PanIN, which was high grade in two (40%) adenoma cases and seven (41%) adenocarcinoma cases. In 16 (73%) evaluable cases, PanIN extended to the pancreatic resection margin; two of which had high grade PanIN. Among the autopsy controls eight (23%) had low-grade PanIN. Seven of the 22 ampullary cases but none of the autopsy controls had coexistent pancreatitis. A smoking history was present in two of four autopsy cases in which this history was available. Overexpression of the p53 and COX-2 proteins was present in only one case of high grade PanIN. K-ras mutations were present in four of four of the PanIN lesions evaluated, including one autopsy case. Clinical follow-up revealed no progression of PanIN to invasive carcinoma in the remnant pancreas, although the follow-up period was too short to adequately assess that risk (an average of 3.8 y for adenoma cases and 2.5 y for adenocarcinoma cases). We conclude that adenomas and carcinomas of the ampulla are associated with PanIN, and often high-grade PanIN. Although its malignant potential has not been fully established, PanIN is underreported and often unrecognized. PanIN may be analogous to colorectal adenoma in that both are prevalent in the older adult population, but few progress to carcinoma. PMID- 11266518 TI - HPV DNA testing of the residual sample of liquid-based Pap test: utility as a quality assurance monitor. AB - HPV DNA testing of the residual sample volume of liquid-based Pap tests has been recommended as a way to determine the appropriate follow-up for women who have equivocal results in routine clinical screening. A major aspect of quality assurance in the cytopathology laboratory consists of correlation of smear interpretation with biopsy or conization results as mandated by CLIA '88. However, the use of histology as the gold standard suffers from similar problems of subjectivity and sampling as the Pap smear. In this study we explore the potential use of HPV DNA testing of the residual volume from the ThinPrep Pap Test (Cytyc Corporation, Boxborough, Massachusetts) as a substitute gold standard in quality assurance monitoring of a cervical cytology screening program. The residual samples from 397 ThinPrep Pap cases were retrospectively analyzed for high-risk HPV DNA using the Hybrid Capture II technique. Sensitivity (71.8%), specificity (86.5%), predictive value of positive (77.1%) and negative (82.9%) ThinPrep Pap interpretations were calculated on the basis of HPV DNA results for 266 cases classed as either squamous intraepithelial lesion (SIL) or negative. Overall, there was agreement between the two tests in 80.8% of cases (Cohen's kappa =.59). The percentage of HPV DNA-positive cases interpreted as atypical squamous cells of uncertain significance (ASCUS) was 43.7%, and the percentage of negative cases was 17.1%. We believe that this approach is an objective adjunct to the traditional quality assurance protocol, with the added benefit that it includes cases interpreted as negative, as well as abnormal cases that do not come to biopsy. PMID- 11266519 TI - Identification of herpes simplex virus genital infection: comparison of a multiplex PCR assay and traditional viral isolation techniques. AB - Genital herpes simplex virus (HSV) is of major public health importance, as indicated by the marked increase in the prevalence of genital herpes over the past two decades. Viral culture has traditionally been regarded as the gold standard for diagnosis. In this study, we compared viral culture and the amplification of HSV DNA by the polymerase chain reaction (PCR) with respect to sensitivity, cost, clinical utility, and turnaround time. Patient sample swabs from 100 individuals were inoculated onto MRC-5 cells for isolation. Positive results were confirmed via a direct fluorescent antibody technique, and serotyping, when requested, was performed using HSV-1 and -2-type-specific sera. PCR techniques employed an extraction step of the same initial swab specimen, followed by PCR amplification, using a multiplex assay for HSV-1, 2 DNA. HSV positive results were found in 32/100 samples via culture and in 36/100 samples via PCR. PCR-positive results yielded 16 (44%) patients infected with HSV-1 and 20 (56%) patients infected with HSV-2. Turnaround time for viral culture averaged 108 hours for positive results and 154 hours for negative results; PCR turnaround time averaged 24--48 hours. Laboratory cost using viral culture was $3.22 for a negative result and $6.49 for a positive result (including direct fluorescent antibody). Serotyping added $10.88 to each culture-positive test. Although laboratory costs for PCR were higher at $8.20/sample, reimbursement levels were also higher. We propose a multiplex PCR assay for diagnosis of HSV-1 and HSV-2 from patient swabs for use in a routine clinical laboratory setting. This assay offers increased sensitivity, typing, and improved turnaround time when compared with traditional viral culture techniques. Although it appears that PCR testing in a routine clinical laboratory setting is cost prohibitive compared with the case of nonserotyped viral culture, it may be very useful when clinical utility warrants distinguishing between HSV 1 and 2 and may be cost effective when reimbursement issues are examined. PMID- 11266520 TI - Concurrent angiomyolipoma and renal cell neoplasia: a study of 36 cases. AB - Little is known about the association of angiomyolipoma and adult renal-cell neoplasia. We studied the clinicopathologic features of 36 patients with concurrent angiomyolipoma and renal-cell neoplasia from the consultation and surgical pathology files of nine institutions. HMB-45 immunoreactivity was analyzed in both neoplasms. Twenty-five sporadic cases of patients with angiomyolipoma and renal-cell neoplasia and 11 cases of patients with tuberous sclerosis, as defined by Gomez' criteria, had mean ages of 59 and 53 years, respectively, and female-male ratios of 2:1 and 5:1, respectively. The mean size of the angiomyolipomas was 1 cm in the sporadic cases and 3 cm in those patients with tuberous sclerosis (medians: 0.5 and 3 cm, respectively, P =.002). The mean sizes of the renal-cell neoplasms were 5 cm in sporadic cases and 6 cm in patients with tuberous sclerosis (medians: 4 and 5 cm, respectively; P =.88). In both clinical settings, angiomyolipoma was more commonly the incidental tumor. Clear-cell (conventional) renal-cell carcinoma was the most common renal-cell neoplasm in both groups of patients, accounting for approximately two thirds of the tumors. In patients with tuberous sclerosis, 27% of renal-cell neoplasms were oncocytomas, compared with 8% in sporadic cases (P =.15). Papillary neoplasia, chromophobe, and collecting-duct renal-cell carcinoma were found only in sporadic cases. All of the 22 renal-cell neoplasms studied were negative for HMB-45, whereas all 25 angiomyolipomas studied were positive. PMID- 11266521 TI - Histopathologic analysis in 46 patients with pseudomyxoma peritonei syndrome: failure versus success with a second-look operation. AB - Pseudomyxoma peritonei syndrome is a disease characterized by mucinous ascites and mucinous tumor disseminated on peritoneal surfaces; the disease almost always originates from a perforated appendiceal epithelial tumor. Histopathologic assessment of aggressive versus noninvasive character of the mucinous tumor has been shown to have an impact on survival in patients treated with cytoreductive surgery and intraperitoneal chemotherapy. Out of a database of 312 patients having a complete cytoreduction for pseudomyxoma peritonei syndrome, 46 patients (24 male and 22 female) had at least one second-look surgery. Before this review, all 46 of these patients were clinically uniformly categorized with a diagnosis of pseudomyxoma peritonei. Using the criteria described by Ronnett and colleagues, all specimens from the multiple surgical procedures performed on these patients were reviewed and reclassified as disseminated peritoneal adenomucinosis (adenomucinosis), adenomucinosis/mucinous adenocarcinoma (hybrid), or mucinous adenocarcinoma. The review was performed in a blinded fashion by a single pathologist (HY). To facilitate a critical evaluation of these histopathologic assessments, the patients were separated into two groups: (1) 19 patients who had a second-look surgery that was unsuccessful in that they went on to die of their disease or in that they currently have disease progression and a limited survival and (2) 27 patients who had a successful second look and currently continue disease free with a minimum 3-year follow-up period. As a result of this review, 11 of 19 patients with an unsuccessful second look and originally designated pseudomyxoma peritonei were reclassified as hybrid-type malignancy (four patients) or mucinous adenocarcinoma (seven patients). Only two patients were reclassified in the successful second-look group (P =.0005). Transitions from a less aggressive to a more invasive histology from one cytoreduction to the next occurred on 13 occasions in patients whose second-look surgery failed and in one patient with a successful second-look surgery (P <.0001). Seven patients retained a histologic classification of disseminated peritoneal adenomucinosis but went on to die of an aggressive disease process. Clinical assessments suggested that failure of second-look surgery for pseudomyxoma peritonei was associated with a biologically more aggressive disease. Unsuccessful second-look surgery for patients with a clinical diagnosis of pseudomyxoma peritonei tumor was often related to an inaccurate initial histologic classification of appendiceal mucinous tumor. Also, a transition from less to more aggressive histology was frequently seen in patients dying of this disease. Assessment of tumor histology can predict the outcome if a uniform surgical treatment is used in patients with peritoneal dissemination of mucinous epithelial tumors of the appendix. PMID- 11266522 TI - Rosai-Dorfman disease isolated to the central nervous system: a report of 11 cases. AB - Sinus histocytosis with massive lymphadenopathy, also known as Rosai-Dorfman Disease (RDD), is an idiopathic histiocytic proliferation affecting lymph nodes. Although extranodal involvement has been reported in diverse sites, central nervous system (CNS) manifestation, particularly in the absence of nodal disease is uncommon. We report 11 cases of RDD primary to the CNS without evidence of other sites of involvement. The cases included 7 males and 4 females ranging in age from 22 to 63 years (mean: 41 y). The patients presented with headaches, seizures, numbness, or paraplegia. Eight cases involved the cranial cavity and three cases, the spinal canal. Lesions were most often extra-axial and dura based. Only one presented in the CNS parenchyma. Histologically, the lesions consisted of variable numbers of pale-staining histocytes with emperipolesis often overshadowed by extensive lymphoplasmacytic infiltrates and fibrosis in the background. Special stains for organisms were negative. By immunohistochemical analysis, the characteristic histiocytes were positive for S100 protein and CD68 and negative for CD1a. Treatment consisted of surgical biopsy or excision. Follow up, available for 10 cases with intervals ranging from 5 days to 42 months (mean: 15 mo), disclosed one patient dying of operative complications 5 days after biopsy and nine patients with no evidence of disease progression RDD should be considered in the differential diagnosis of inflammatory lesions of the CNS. Our study suggests that this entity may have been misdiagnosed in the past as plasma cell granuloma or inflammatory pseudotumor. PMID- 11266523 TI - Pleomorphic liposarcoma: a clinicopathologic analysis of 19 cases. AB - Pleomorphic liposarcoma is a variant of liposarcoma defined morphologically by the presence of pleomorphic lipoblasts. Because of its rarity, there are limited studies with long-term follow-up information. Nineteen pleomorphic liposarcomas were studied. Unequivocal pleomorphic lipoblasts were required for inclusion. In each case, the following features were noted: tumor site; tumor size; tumor depth; predominant histologic pattern (epithelioid or malignant fibrous histiocytoma [MFH]-like); extent of necrosis (absent, less than 15%, or at least 15%); mitotic counts; treatment and clinical follow-up. Patients were 11 females and 8 males, aged 33-87 years (mean, 64.5 y; median, 70 y). Tumors involved the extremities (13 patients: intramuscular in 10, subcutaneous in 2, depth unknown in 1), retroperitoneum (4 patients), mediastinum (1 patient), and paratesticular region (1 patient). Size ranged from 4.5--31 cm (mean, 11.9 cm; median, 12.0 cm). Predominant pattern was epithelioid in 7 and MFH-like in 12. Necrosis was present in 15 (79%) and was extensive (36 15%) in 14 patients. Mitotic counts ranged from 0.2--3.4/10 high-power fields (mean, 1.4; median, 1.4) by the average-count method and from 1--6/10 high power fields by the highest count method (mean, 2.9; median, 3.0). All patients were treated surgically; 10 patients received adjuvant chemotherapy and/or radiation therapy. On follow-up of 18 patients (range, 2--129 mo; mean, 35.4 mo; median, 23 mo) nine (50%) were dead of disease (range, 2--48 mo; mean, 20.1 mo; median, 12 mo), one died of other causes 2 months after diagnosis, two were alive with disease, five were disease free, and one was alive at 129 months (tumor status unknown). Five had recurrences (range, 3--28 mo; mean, 14.4 mo; median, 8 mo), and four of five (80%) with recurrences were dead of disease. Metastases developed in eight patients (range, 4--48 mo; mean, 19.5 mo; median, 11.5 mo), most commonly to the lungs. In conclusion, pleomorphic liposarcoma is a rare tumor of adulthood that occurs most commonly in the deep, soft tissues of the extremities. It behaves as a high-grade sarcoma that frequently metastasizes, most commonly to the lungs. Although this tumor has a wide range of histologic appearances, no clinical or pathologic feature is predictive of a more aggressive clinical course. PMID- 11266524 TI - Pathologic, immunohistochemical, and molecular features of benign and malignant phyllodes tumors of the breast. AB - The histologic distinction between benign and malignant Phyllodes tumors (PT) is often difficult and arbitrary. We analyzed a group of benign and malignant PT to determine whether specific histologic features and expression of Ki-67 and p53 could be useful in distinguishing benign PT from malignant tumors. We also determined whether deletions in Chromosome 3p at the FHIT and hMLH1 loci are common abnormalities in PT. Twenty PT were histologically classified as benign (7) or malignant (13). Seven of the malignant PT were low grade, and six were high grade. Ki-67 and p53 immunohistochemistry was performed on all tumors and analyzed for the stromal and for the epithelial component. PCR-based loss of heterozygosity analyses were performed with the following markers on Chromosome 3p: D3S1478 (3p21.2--21.3), D3S1289 (3p21.1--21.2), and D3S1295 (3p14.3--21.1). The distribution of immunoreactivity for Ki-67 was analyzed by quantifying the percentage of positive nuclei and expressed as the labeling index (LI). Patients' ages ranged from 13 to 71 years (median: 51 y). After a mean follow-up period of 8 years, none of the PT metastasized, whereas three recurred locally. Although malignant PT were larger than benign PT (means, 7.1 versus 4.3 cm), this difference was not statistically significant. Five tumors had infiltrating margins, and 14 were circumscribed. The Ki-67 LI in low-grade malignant PT (16 +/ 25.5) was significantly higher than that in benign PT (3.6 +/- 4.8), whereas the LI in the high-grade malignant PT group (50 +/- 21.9) was significantly higher than that in low-grade malignant tumors (P =.012). The Ki-67 LI in the three tumors that recurred was less than 10%. Two of seven (29%) benign PT were focally positive for p53, whereas four of seven (57%) low-grade malignant and three of six (50%) high-grade malignant PT were diffusely positive for p53. The three tumors that recurred initially were histologically benign, as were two of the recurrences. One recurrent tumor evolved to a high-grade malignant PT. Margins were greater than 1 cm in all tumors except four, three of which recurred locally. No allelic loss of 3p was found. In summary, Ki-67 expression may assist in distinguishing benign from malignant PT in diagnostically difficult cases. 3p deletions do not play a significant role in the development of these tumors. Neither Ki-67 nor p53 can reliably predict recurrence. Histologically high-grade malignant PT have a favorable prognosis if widely excised. We emphasize the importance of adequate margins in the treatment of benign and malignant PT. PMID- 11266525 TI - Follicular lymphoma with marginal zone differentiation: microdissection demonstrates the t(14;18) in both the follicular and marginal zone components. AB - On occasion, follicle center lymphomas (FCL) may contain a marginal-zone (MZ) component in which the interfollicular lymphoid cells take on an MZ cell morphology. In the past, these have been termed composite lymphomas. However, recent studies suggest that the two components are clonally related. It is unknown whether the bcl-2 translocation present in most FCLs is present in the cells that demonstrate MZ cell morphology. We have identified three cases of low grade FCL with a MZ component suitable for laser capture microdissection (LCM) of the two components. Cases were immunophenotyped in paraffin section with antibodies to CD10, CD20, bcl-2, and bcl-6. LCM was done to isolate cells from each component. Polymerase chain reaction for t(14;18) using primers to the major breakpoint region was performed on DNA extracts. The sensitivity of the PCR assay was decreased to 5%--10% follicle center cells in a background of reactive tonsil cells. All three cases showed different phenotypes in each component. The FCL component was positive for all four of the above markers, whereas the MZ component expressed only CD20 and bcl-2. Both components showed t(14;18) amplicons of identical size, with the MZ component signal being stronger than the 5%--10% sensitivity control, suggesting that the signal was not from rare, contaminating FCL cells. These results confirm that both components are clonally related and support the theory that these are indeed FCLs with MZ differentiation (that retain the t(14;18)) rather than the reverse, MZ lymphoma with follicle center differentiation. PMID- 11266526 TI - Prostaglandin D synthase (beta-trace) in meningeal hemangiopericytoma. AB - The level of prostaglandin D synthase (PGDS), a major protein constituent of cerebrospinal fluid (CSF), is altered in various brain diseases, including meningitis. However, its role in the brain remains unclear. PGDS is mainly synthesized in the arachnoid cells, the choroid plexus and oligodendrocytes in the central nervous system. Among brain tumors, meningiomas showed intense immunoreactivity to PGDS in the perinuclear region. Thus, PGDS has been considered a specific cell marker of meningioma. In this study, we examined 25 meningeal hemangiopericytomas (HPCs) and found that 16 of the tumors (64%) showed immunoreactivity for PGDS in the perinuclear region. For comparison, 15 meningiomas, 14 soft-tissue HPCs, 1 mesenchymal chondrosarcoma, 3 choroid plexus papillomas, and 7 oligodendrogliomas were also examined. Meningiomas showed positive immunoreactivity for PGDS in 13 cases (80%). Except for one case located at the sacrum, none of the other soft-tissue HPCs showed immunostaining for PGDS. Mesenchymal chondrosarcoma arises in the bones of the skull, and its histological pattern resembles that of HPC; however, it showed no immunoreactivity for PGDS. Neither choroid plexus papillomas nor oligodendrogliomas were immunopositive for PGDS. These findings suggest that meningeal HPCs may have a unique molecular phenotype that is distinct from that of the soft-tissue HPCs. The origin of meningeal HPCs may be more closely related to the arachnoid cells. PMID- 11266527 TI - Hamartin and tuberin expression in human tissues. AB - Tuberous sclerosis (TSC) is a bigenic autosomal dominant disease caused by mutations in one of two tumor-suppressor genes, TSC1 and TSC2, resulting in benign hamartomas and low grade neoplasms in multiple organs including brain, heart, kidney, and skin. We report the results of an immunohistochemical study of the expression of the TSC gene products, tuberin and hamartin, in multiple tissues obtained at autopsy from 12 non-TSC affected patients ranging in age from 20 weeks gestation to 8 years, and surgical specimens from some organs. Tuberin and hamartin are expressed and are colocalized in most tissues. Contrary to a previous report, immunostaining with our antisera detected hamartin in liver, small and large intestine, prostate, and testes. We did not detect significant developmental differences in tuberin or hamartin expression in comparable tissues from patients of different ages. Although tuberin and hamartin colocalize in most tissues and cell types, we provide data that hamartin is more abundantly expressed than tuberin in cells within some tissues including the distal nephron and a population of cells of the endocrine pancreas. These data support the hypothesis that hamartin and tuberin interact and may function together in many tissues where they are co-expressed, but also suggest that hamartin has a discrete and specialized function in certain cell types. PMID- 11266528 TI - The 2000 Long Course. Introduction. PMID- 11266529 TI - Breast cancer in the 21st century: neu opportunities and neu challenges. AB - Recent advances in the understanding of the molecular and genetic alterations underlying breast cancer development and progression have provided the opportunity to develop novel therapeutic strategies for this disease. None of these developments has had a greater recent impact on clinicians and pathologists than the recognition of the importance of the HER-2/neu (c-erbB-2) oncogene. Located on chromosome 17, this gene encodes a 185 kD transmembrane glycoprotein with tyrosine kinase activity that functions as a growth factor receptor. Amplification or overexpression of HER-2/neu is seen in approximately 20 to 30% of invasive breast cancers and this has been considered to be an adverse prognostic factor in many studies. However, recent interest in HER-2/neu has largely been focused on its role as a potential target for breast cancer treatment. In particular, recognition of the role of HER-2/neu in breast cancer growth led to the development of a humanized monoclonal antibody directed against the HER-2/neu protein as a therapeutic agent (Herceptin). Clinical studies have further suggested that HER-2/neu status can provide important information regarding sensitivity to certain forms of conventional systemic therapy, particularly anthracyclines. As a result of these developments, there has been increasing demand for pathologists to perform assays for HER-2/neu on current and archived breast cancer specimens. Immunohistochemistry and fluorescence in situ hybridization have emerged as the most viable assays for evaluation of HER-2/neu in routine clinical practice. However, each of these methods has its advantages and disadvantages. Determining the relative merits of these assays and developing clinically meaningful and reproducible systems to report the results are challenges pathologists must now address. The development of a therapeutic agent that directly targets a protein involved in a growth-signaling pathway represents a new paradigm in breast cancer treatment. Therapeutic strategies that target other molecules involved in breast cancer development and progression are on the horizon. It is crucial that pathologists become aware of these advances and assume a pivotal role in the development and application of assays to evaluate these new molecular targets. PMID- 11266531 TI - Practicing pediatric pathology without a microscope. AB - This article highlights changes in the field of pediatric pathology that have resulted from technical advances in prenatal diagnostics, immunohistochemistry, cytogenetics, and molecular genetics. The relatively new and growing need for specialized training in fetal pathology is used as an example. Comprehensive evaluation of human fetuses has become a requisite skill for many diagnostic pathologists, in part because contemporary prenatal diagnostic techniques have shifted the demographics of many congenital conditions from spontaneous term delivery to mid-gestation termination of pregnancy. The information provided by the pathologist has a tremendous impact for families and clinicians as they consider recurrence risks in future pregnancies. As most specimens from therapeutic terminations have gross dysmorphology, which may or may not constitute a recognizable pattern of human malformation, their analysis requires additional skills and methods that were traditionally the domain other specialists (e.g., medical geneticists). The pathologist must learn to identify syndromes, to be aware of their underlying etiology and pathogenesis, and to utilize advanced cytogenetic methods (e.g., fluorescence in situ hybridization), flow cytometry, or specific mutational analysis when appropriate. At a minimum, important anatomic details must be well documented and appropriate tissue samples should be obtained and stored to facilitate more specific diagnostic testing in the future. PMID- 11266530 TI - Anaplastic large cell lymphoma: the shifting sands of diagnostic hematopathology. AB - Anaplastic large cell lymphoma (ALCL) is a paradigm for the process used to define new disease entities, and provides a model that is applicable to all areas of pathology. ALCL was first recognized based on characteristic histologic features (sinusoidal invasion) and a distinctive immunophenotype (CD30+). However, neither sinusoidal invasion nor CD30-positivity proved to be entirely specific. Subsequently, a characteristic cytogenetic abnormality was identified, the t(2;5), that led to identification of the genes involved in the translocation (NPM/ALK) and insights into the pathogenesis. Generation of monoclonal antibodies to the aberrantly expressed anaplastic large cell lymphoma kinase (ALK) such as ALK-1 can be used diagnostically, and have led to improved definition of the diagnostic entity with important clinical and prognostic implications. These studies also have clarified the relationship of ALCL to Hodgkin's disease, another lymphoid malignancy associated with CD30 expression. We have learned that the ultimate histologic spectrum of ALCL is both narrower and broader than originally believed. The small cell and lymphohistiocytic variants of ALCL are ALK-positive, and are an accepted part of the disease entity, although the neoplastic cells may appear neither large nor anaplastic. Conversely, most cases of Hodgkin's-like ALCL have proved to be more closely related to true Hodgkin's disease, and are unrelated to ALCL. PMID- 11266532 TI - Carcinogenesis in the GI tract: from morphology to genetics and back again. AB - The genetic alterations in colorectal cancer progression are determined by one of two separate and distinct underlying pathways of genomic instability. The first pathway, chromosomal instability, is characterized by allelic losses and aneuploidy. The second pathway, microsatellite instability, is characterized by an abundance of subtle DNA mutations and diploidy. Although the genes causing chromosomal instability remain unknown, microsatellite instability is caused by inactivation of a DNA mismatch repair gene (predominantly MLH1 or MSH2). Microsatellite instability is present in 15% of colorectal cancers, and is diagnosed by analysis of tumor DNA from paraffin blocks and by demonstration of loss of mismatch repair protein expression in cancers. In addition to the unique profile of genetic alterations, colorectal cancers with microsatellite instability have distinct pathologic features and improved survival. Finally, cancers from most patients with hereditary non-polyposis colorectal cancer (or Lynch syndrome) have microsatellite instability due to germline mutations in the DNA mismatch repair genes. Identification of the microsatellite instability pathway has enormous implications for the clinical investigation and management of colorectal cancer patients. PMID- 11266533 TI - How familial cancer genes and environmentally induced oncogenes have changed the endocrine landscape. AB - The gene responsible for MEN-2, the ret proto-ocogene, has elucidated mechanisms of endocrine tumorigenesis. Activating mutations of this transmembrane tyrosine kinase receptor represent the first known example of an inherited oncogene. This knowledge has altered our approach to affected patients by allowing in utero screening and prophylactic thyroidectomy rather than provocative testing and morphologic analysis of C cell hyperplasia--will it result in eradication of medullary carcinoma of thyroid? The lessons from Chernobyl taught us how radiation can induce chromosomal rearrangements that involve the same gene. This has led to a better understanding of papillary thyroid carcinoma and provides a novel immunohistochemical marker that widens our diagnostic armamentarium. PMID- 11266534 TI - Applications of molecular diagnostics: solid tumor genetics can determine clinical treatment protocols. AB - Solid tumor diagnosis is now entering an era in which molecular genetics plays an important role. Clinical trials have shown different responses to various therapies that correlate with molecular alterations. Biological determinants related to treatment response markers aimed at individualized therapies are being defined and implemented. Patients are now being treated based on the profile of molecular genetic alterations in individual tumors. Protocols based on molecular markers will increase the chances for cure by opting for the right management approach. They also will improve, in most situations, the quality of life of patients with cancer, for example by facilitating organ preservation strategies. The molecular characterization therefore has an important prognostic and practical role in diagnosis. PMID- 11266535 TI - The continuing role of morphology in the molecular age. PMID- 11266536 TI - The 2000 Long Course. Conclusion. PMID- 11266537 TI - Correspondence re: Viswanatha DS, Foucar K, Berry BR, Gascoyne RD, Evans HL, Leith CP. Blastic mantle cell leukemia: an unusual presentation of blastic mantle cell lymphoma. Mod Pathol 2000;13:825-33. PMID- 11266538 TI - Correspondence re: Jimenez RE, Wallis T, Tabasczka P, Visscher DW. Determination of Her-2/Neu status in breast carcinoma: comparative analysis of immunohistochemistry and fluorescent in situ hybridization. Mod Pathol 2000;13:37 45. PMID- 11266540 TI - All Tcf HMG box transcription factors interact with Groucho-related co repressors. AB - Tcf/Lef family transcription factors are the downstream effectors of the Wingless/Wnt signal transduction pathway. Upon Wingless/Wnt signalling, beta catenin translocates to the nucleus, interacts with Tcf (1-3) and thus activates transcription of target genes (4,5). Tcf factors also interact with members of the Groucho (Grg/TLE) family of transcriptional co-repressors (6). We have now tested all known mammalian Groucho family members for their ability to interact specifically with individual Tcf/Lef family members. Transcriptional activation by any Tcf could be repressed by Grg-1, Grg-2/TLE-2, Grg-3 and Grg-4 in a reporter assay. Specific interactions between Tcf and Grg proteins may be achieved in vivo by tissue- or cell type-limited expression. To address this, we determined the expression of all Tcf and Grg/TLE family members in a panel of cell lines. Within any cell line, several Tcfs and TLEs are co-expressed. Thus, redundancy in Tcf/Grg interactions appears to be the rule. The 'long' Groucho family members containing five domains are repressors of Tcf-mediated transactivation, whereas Grg-5, which only contains the first two domains, acts as a de-repressor. As previously shown for Drosophila Groucho, we show that long Grg proteins interact with histone deacetylase-1. Although Grg-5 contains the GP homology domain that mediates HDAC binding in long Grg proteins, Grg-5 fails to bind this co-repressor, explaining how it can de-repress transcription. PMID- 11266541 TI - Enhancement of translation by the epsilon element is independent of the sequence of the 460 region of 16S rRNA. AB - The epsilon enhancer element is a pyrimidine-rich sequence that increases expression of T7 gene 10 and a number of Escherichia coli mRNAs during initiation of translation and inhibits expression of the recF mRNA during elongation. Based on its complementarity to the 460 region of 16S rRNA, it has been proposed that epsilon exerts its enhancer activity by base pairing to this complementary rRNA sequence. We have tested this model of enhancer action by constructing mutations in the 460 region of 16S rRNA and examining expression of epsilon-containing CAT reporter genes and recF-lacZ fusions in strains expressing the mutant rRNAs. Replacement of the 460 E.coli stem-loop with that of Salmonella enterica serovar Typhimurium or a stem-loop containing a reversal of all 8 bp in the helical region produced fully functional rRNAs with no apparent effect on cell growth or expression of any epsilon-containing mRNA. Our experiments confirm the reported effects of the epsilon elements on gene expression but show that these effects are independent of the sequence of the 460 region of 16S rRNA, indicating that epsilon-rRNA base pairing does not occur. PMID- 11266542 TI - Monitoring the structure of Escherichia coli RNase P RNA in the presence of various divalent metal ions. AB - Lead(II)-induced cleavage can be used as a tool to probe conformational changes in RNA. In this report, we have investigated the conformation of M1 RNA, the catalytic subunit of Escherichia coli RNase P, by studying the lead(II)-induced cleavage pattern in the presence of various divalent metal ions. Our data suggest that the overall conformation of M1 RNA is very similar in the presence of Mg(2+), Mn(2+), Ca(2+), Sr(2+) and Ba(2+), while it is changed compared to the Mg(2+)-induced conformation in the presence of other divalent metal ions, Cd(2+) for example. We also observed that correct folding of some M1 RNA domains is promoted by Pb(2+), while folding of other domain(s) requires the additional presence of other divalent metal ions, cobalt(III) hexamine or spermidine. Based on the suppression of Pb(2+) cleavage at increasing concentrations of various divalent metal ions, our findings suggest that different divalent metal ions bind with different affinities to M1 RNA as well as to an RNase P hairpin-loop substrate and yeast tRNA(Phe). We suggest that this approach can be used to obtain information about the relative binding strength for different divalent metal ions to RNA in general, as well as to specific RNA divalent metal ion binding sites. Of those studied in this report, Mn(2+) is generally among the strongest RNA binders. PMID- 11266543 TI - Monitoring S phase progression globally and locally using BrdU incorporation in TK(+) yeast strains. AB - Eukaryotic chromosome replication is initiated from numerous origins and its activation is temporally controlled by cell cycle and checkpoint mechanisms. Yeast has been very useful in defining the genetic elements required for initiation of DNA replication, but simple and precise tools to monitor S phase progression are lacking in this model organism. Here we describe a TK(+) yeast strain and conditions that allow incorporation of exogenous BrdU into genomic DNA, along with protocols to detect the sites of DNA synthesis in yeast nuclei or on combed DNA molecules. S phase progression is monitored by quantification of BrdU in total yeast DNA or on individual chromosomes. Using these tools we show that yeast chromosomes replicate synchronously and that DNA synthesis occurs at discrete subnuclear foci. Analysis of BrdU signals along single DNA molecules from hydroxyurea-arrested cells reveals that replication forks stall 8-9 kb from origins that are placed 46 kb apart on average. Quantification of total BrdU incorporation suggests that 190 'early' origins have fired in these cells and that late replicating territories might represent up to 40% of the yeast genome. More generally, the methods outlined here will help understand the kinetics of DNA replication in wild-type yeast and refine the phenotypes of several mutants. PMID- 11266539 TI - The RAG proteins in V(D)J recombination: more than just a nuclease. AB - V(D)J recombination is the process that generates the diversity among T cell receptors and is one of three mechanisms that contribute to the diversity of antibodies in the vertebrate immune system. The mechanism requires precise cutting of the DNA at segment boundaries followed by rejoining of particular pairs of the resulting termini. The imprecision of aspects of the joining reaction contributes significantly to increasing the variability of the resulting functional genes. Signal sequences target DNA recombination and must participate in a highly ordered protein-DNA complex in order to limit recombination to appropriate partners. Two proteins, RAG1 and RAG2, together form the nuclease that cleaves the DNA at the border of the signal sequences. Additional roles of these proteins in organizing the reaction complex for subsequent steps are explored. PMID- 11266544 TI - The human XPG gene: gene architecture, alternative splicing and single nucleotide polymorphisms. AB - Defects in the XPG DNA repair endonuclease gene can result in the cancer-prone disorders xeroderma pigmentosum (XP) or the XP-Cockayne syndrome complex. While the XPG cDNA sequence was known, determination of the genomic sequence was required to understand its different functions. In cells from normal donors, we found that the genomic sequence of the human XPG gene spans 30 kb, contains 15 exons that range from 61 to 1074 bp and 14 introns that range from 250 to 5763 bp. Analysis of the splice donor and acceptor sites using an information theory based approach revealed three splice sites with low information content, which are components of the minor (U12) spliceosome. We identified six alternatively spliced XPG mRNA isoforms in cells from normal donors and from XPG patients: partial deletion of exon 8, partial retention of intron 8, two with alternative exons (in introns 1 and 6) and two that retained complete introns (introns 3 and 9). The amount of alternatively spliced XPG mRNA isoforms varied in different tissues. Most alternative splice donor and acceptor sites had a relatively high information content, but one has the U12 spliceosome sequence. A single nucleotide polymorphism has allele frequencies of 0.74 for 3507G and 0.26 for 3507C in 91 donors. The human XPG gene contains multiple splice sites with low information content in association with multiple alternatively spliced isoforms of XPG mRNA. PMID- 11266545 TI - Evidence for regulation of protein synthesis at the elongation step by CDK1/cyclin B phosphorylation. AB - Eukaryotic elongation factor 1 (eEF-1) contains the guanine nucleotide exchange factor eEF-1B that loads the G protein eEF-1A with GTP after each cycle of elongation during protein synthesis. Two features of eEF-1B have not yet been elucidated: (i) the presence of the unique valyl-tRNA synthetase; (ii) the significance of target sites for the cell cycle protein kinase CDK1/cyclin B. The roles of these two features were addressed by elongation measurements in vitro using cell-free extracts. A poly(GUA) template RNA was generated to support both poly(valine) and poly(serine) synthesis and poly(phenylalanine) synthesis was driven by a poly(uridylic acid) template. Elongation rates were in the order phenylalanine > valine > serine. Addition of CDK1/cyclin B decreased the elongation rate for valine whereas the rate for serine and phenylalanine elongation was increased. This effect was correlated with phosphorylation of the eEF-1delta and eEF-1gamma subunits of eEF-1B. Our results demonstrate specific regulation of elongation by CDK1/cyclin B phosphorylation. PMID- 11266546 TI - Site-specifically located 8-amino-2'-deoxyguanosine: thermodynamic stability and mutagenic properties in Escherichia coli. AB - 2-Nitropropane (2-NP), an important industrial solvent and a component of cigarette smoke, is mutagenic in bacteria and carcinogenic in rats. 8-Amino-2' deoxyguanosine (8-amino-dG) is one of the types of DNA damage found in liver, the target organ in 2-NP-treated rats. To investigate the thermodynamic properties of 8-amino-dG opposite each of the four DNA bases, we have synthesized an 11mer, d(CCATCG*CTACC), in which G* represents the modified base. By annealing a complementary DNA strand to this modified 11mer, four sets of duplexes were generated each containing one of the four DNA bases opposite the lesion. Circular dichroism studies indicated that 8-amino-dG did not alter the global helical properties of natural right-handed B-DNA. The thermal stability of each duplex was examined by UV melting measurements and compared with its unmodified counterpart. For the unmodified 11mer, the relative stability of the complementary DNA bases opposite G was in the order C > T > G > A, as determined from their -DeltaG degrees values. The free energy change of each modified duplex was lower than its unmodified counterpart, except for the G*:G pair that exhibited a higher melting transition and a larger -DeltaG degrees than the G:G duplex. Nevertheless, the stability of the modified 11mer duplex also followed the order C > T > G > A when placed opposite 8-amino-dG. To explore if 8-amino-dG opposite another 8-amino-dG has any advantage in base pairing, a G*:G* duplex was evaluated, which showed that the stability of this duplex was similar to the G*:G duplex. Mutagenesis of 8-amino-dG in this sequence context was studied in Escherichia coli, which showed that the lesion is weakly mutagenic (mutation frequency approximately 10(-3)) but still can induce a variety of targeted and semi-targeted mutations. PMID- 11266547 TI - Comparison of intron-containing and intron-lacking human genes elucidates putative exonic splicing enhancers. AB - Of the rules used by the splicing machinery to precisely determine intron-exon boundaries only a fraction is known. Recent evidence suggests that specific short sequences within exons help in defining these boundaries. Such sequences are known as exonic splicing enhancers (ESE). A possible bioinformatical approach to studying ESE sequences is to compare genes that harbor introns with genes that do not. For this purpose two non-redundant samples of 719 intron-containing and 63 intron-lacking human genes were created. We performed a statistical analysis on these datasets of intron-containing and intron-lacking human coding sequences and found a statistically significant difference (P = 0.01) between these samples in terms of 5-6mer oligonucleotide distributions. The difference is not created by a few strong signals present in the majority of exons, but rather by the accumulation of multiple weak signals through small variations in codon frequencies, codon biases and context-dependent codon biases between the samples. A list of putative novel human splicing regulation sequences has been elucidated by our analysis. PMID- 11266548 TI - Hybrid characteristics of oligonucleotides consisting of isonucleoside 2',5' anhydro-3'-deoxy-3'-(thymin-1-yl)-D-mannitol with different linkage modes. AB - Oligonucleotides consisting of the isonucleoside repeating unit 2',5'-anhydro-3' deoxy-3'-(thymin-1-yl)-D-mannitol (4) were synthesized with the monomeric unit 4 incorporated into oligonucleotides as 1'-->4' linkage 4a (oligomer I) or 6'-->4' linkage 4b (oligomer II). The hybrid properties of the two oligonucleotides I and II with their complementary strands were investigated by thermal denaturation and CD spectra. Oligonucleotide I (4a) formed a stable duplex with d(A)(14) with a slightly reduced T(m) value of 36.6 degrees C, relative to 38.2 degrees C for the control duplex d(T)(14)/d(A)(14), but oligomer II (4b) failed to hybridize with a DNA complementary single strand. The spectrum of the duplex oligomer I/d(A)(14) showed a positive CD band at 217 nm and a negative CD band at 248 nm attributable to a B-like conformation. Molecular modeling showed that in the case of oligomer I: the C6' hydroxy group of each unit could be located in the groove area when hybridized to the DNA single strand, which might contribute additional hydrogen bonding to the stability of duplex formation. PMID- 11266549 TI - SfiI endonuclease activity is strongly influenced by the non-specific sequence in the middle of its recognition site. AB - The SfiI endonuclease cleaves DNA at the sequence GGCCNNNN NGGCC, where N is any base and downward arrow is the point of cleavage. Proteins that recognise discontinuous sequences in DNA can be affected by the unspecified sequence between the specified base pairs of the target site. To examine whether this applies to SFII, a series of DNA duplexes were made with identical sequences apart from discrete variations in the 5 bp spacer. The rates at which SFII cleaved each duplex were measured under steady-state conditions: the steady-state rates were determined by the DNA cleavage step in the reaction pathway. SFII cleaved some of these substrates at faster rates than other substrates. For example, the change in spacer sequence from AACAA to AAACA caused a 70-fold increase in reaction rate. In general, the extrapolated values for k(cat) and K(m) were both higher on substrates with inflexible spacers than those with flexible structures. The dinucleotide at the site of cleavage was largely immaterial. SFII activity is thus highly dependent on conformational variations in the spacer DNA. PMID- 11266550 TI - Methylation by a mutant T2 DNA [N(6)-adenine] methyltransferase expands the usage of RecA-assisted endonuclease (RARE) cleavage. AB - Properties of a mutant bacteriophage T2 DNA [N:(6)-adenine] methyltransferase (T2 Dam MTase) have been investigated for its potential utilization in RecA-assisted restriction endonuclease (RARE) cleavage. Steady-state kinetic analyses with oligonucleotide duplexes revealed that, compared to wild-type T4 Dam, both wild type T2 Dam and mutant T2 Dam P126S had a 1.5-fold higher k(cat) in methylating canonical GATC sites. Additionally, T2 Dam P126S showed increased efficiencies in methylation of non-canonical GAY sites relative to the wild-type enzymes. In agreement with these steady-state kinetic data, when bacteriophage lambda DNA was used as a substrate, maximal protection from restriction nuclease cleavage in vitro was achieved on the sequences GATC, GATN and GACY, while protection of GACR sequences was less efficient. Collectively, our data suggest that T2 Dam P126S can modify 28 recognition sequences. The feasibility of using the mutant enzyme in RARE cleavage with BCL:I and ECO:RV endonucleases has been shown on phage lambda DNA and with BCL:I and DPN:II endonucleases on yeast chromosomal DNA embedded in agarose. PMID- 11266551 TI - DNA methyltransferases of the cyanobacterium Anabaena PCC 7120. AB - From the characterization of enzyme activities and the analysis of genomic sequences, the complement of DNA methyltransferases (MTases) possessed by the cyanobacterium ANABAENA PCC 7120 has been deduced. ANABAENA has nine DNA MTases. Four are associated with Type II restriction enzymes (AVAI, AVAII, AVAIII and the newly recognized inactive AVAIV), and five are not. Of the latter, four may be classified as solitary MTases, those whose function lies outside of a restriction/modification system. The group is defined here based on biochemical and genetic characteristics. The four solitary MTases, DmtA/M.AVAVI, DmtB/M.AVAVII, DmtC/M. AVAVIII and DmtD/M.AVAIX, methylate at GATC, GGCC, CGATCG and rCCGGy, respectively. DmtB methylates cytosines at the N4 position, but its sequence is more similar to N6-adenine MTases than to cytosine-specific enzymes, indicating that it may have evolved from the former. The solitary MTases, appear to be of ancient origin within cyanobacteria, while the restriction MTases appear to have arrived by recent horizontal transfer as did five now inactive Type I restriction systems. One Mtase, M.AVAV, cannot reliably be classified as either a solitary or restriction MTase. It is structurally unusual and along with a few proteins of prokaryotic and eukaryotic origin defines a structural class of MTases distinct from all previously described. PMID- 11266552 TI - A single alteration 20 nt 5' to an editing target inhibits chloroplast RNA editing in vivo. AB - Transcripts of typical dicot plant plastid genes undergo C-->U RNA editing at approximately 30 locations, but there is no consensus sequence surrounding the C targets of editing. The cis-acting elements required for editing of the C located at tobacco rpoB editing site II were investigated by introducing translatable chimeric minigenes containing sequence -20 to +6 surrounding the C target of editing. When the -20 to +6 sequence specified by the homologous region present in the black pine chloroplast genome was incorporated, virtually no editing of the transcripts occurred in transgenic tobacco plastids. Nucleotides that differ between the black pine and tobacco sequence were tested for their role in C-->U editing by designing chimeric genes containing one or more of these divergent nucleotides. Surprisingly, the divergent nucleotide that had the strongest negative effect on editing of the minigene transcript was located -20 nt 5' to the C target of editing. Expression of transgene transcripts carrying the 27 nt sequence did not affect the editing extent of the endogenous rpoB transcripts, even though the chimeric transcripts were much more abundant than those of the endogenous gene. In plants carrying a 93 nt rpoB editing site sequence, transgene transcripts accumulated to a level three times greater than transgene transcripts in the plants carrying the 27 nt rpoB editing sites and resulted in editing of the endogenous transcripts from 100 to 50%. Both a lower affinity of the 27 nt site for a trans-acting factor and lower abundance of the transcript could explain why expression of minigene transcripts containing the 27 nt sequence did not affect endogenous editing. PMID- 11266553 TI - Target gene search for the metal-responsive transcription factor MTF-1. AB - Activation of genes by heavy metals, notably zinc, cadmium and copper, depends on MTF-1, a unique zinc finger transcription factor conserved from insects to human. Knockout of MTF-1 in the mouse results in embryonic lethality due to liver decay, while knockout of its best characterized target genes, the stress-inducible metallothionein genes I and II, is viable, suggesting additional target genes of MTF-1. Here we report on a multi-pronged search for potential target genes of MTF 1, including microarray screening, SABRE selective amplification, a computer search for MREs (DNA-binding sites of MTF-1) and transfection of reporter genes driven by candidate gene promoters. Some new candidate target genes emerged, including those encoding alpha-fetoprotein, the liver-enriched transcription factor C/EBPalpha and tear lipocalin/von Ebner's gland protein, all of which have a role in toxicity/the cell stress response. In contrast, expression of other cell stress-associated genes, such as those for superoxide dismutases, thioredoxin and heat shock proteins, do not appear to be affected by loss of MTF 1. Our experiments have also exposed some problems with target gene searches. First, finding the optimal time window for detecting MTF-1 target genes in a lethal phenotype of rapid liver decay proved problematical: 12.5-day-old mouse embryos (stage E12.5) yielded hardly any differentially expressed genes, whereas at stage 13.0 reduced expression of secretory liver proteins probably reflected the onset of liver decay, i.e. a secondary effect. Likewise, up-regulation of some proliferation-associated genes may also just reflect responses to the concomitant loss of hepatocytes. Another sobering finding concerns gamma glutamylcysteine synthetase(hc) (gamma-GCS(hc)), which controls synthesis of the antioxidant glutathione and which was previously suggested to be a target gene contributing to the lethal phenotype in MTF-1 knockout mice. gamma-GCS(hc) mRNA is reduced at the onset of liver decay but MTF-1 null mutant embryos manage to maintain a very high glutathione level until shortly before that stage, perhaps in an attempt to compensate for low expression of metallothioneins, which also have a role as antioxidants. PMID- 11266554 TI - Transcriptional adaptor and histone acetyltransferase proteins in Arabidopsis and their interactions with CBF1, a transcriptional activator involved in cold regulated gene expression. AB - The ARABIDOPSIS CBF transcriptional activators bind to the CRT/DRE regulatory element present in the promoters of many cold-regulated genes and stimulate their transcription. Expression of the CBF1 proteins in yeast activates reporter genes carrying a minimal promoter with the CRT/DRE as an upstream regulatory element. Here we report that this ability of CBF1 is dependent upon the activities of three key components of the yeast Ada and SAGA complexes, namely the histone acetyltransferase (HAT) Gcn5 and the transcriptional adaptor proteins Ada2 and Ada3. This result suggested that CBF1 might function through the action of similar complexes in ARABIDOPSIS In support of this hypothesis we found that ARABIDOPSIS has a homolog of the GCN5 gene and two homologs of ADA2, the first report of multiple ADA2 genes in an organism. The ARABIDOPSIS GCN5 protein has intrinsic HAT activity and can physically interact in vitro with both the ARABIDOPSIS ADA2a and ADA2b proteins. In addition, the CBF1 transcriptional activator can interact with the ARABIDOPSIS GCN5 and ADA2 proteins. We conclude that ARABIDOPSIS encodes HAT-containing adaptor complexes that are related to the Ada and SAGA complexes of yeast and propose that the CBF1 transcriptional activator functions through the action of one or more of these complexes. PMID- 11266555 TI - Ribozyme minigene-mediated RAD51 down-regulation increases radiosensitivity of human prostate cancer cells. AB - The strand transferase RAD51 is a component of the homologous recombination repair pathway. To examine the contribution of RAD51 to the genotoxic effects of ionising radiation, we have used a novel ribozyme strategy. A reporter gene vector was constructed so that expression of an inserted synthetic double stranded ribozyme-encoding oligonucleotide would be under the control of the cytomegalovirus immediate-early gene enhancer/promoter system. The prostate tumour cell line LNCaP was transfected with this vector or a control vector, and a neomycin resistance gene on the vector was used to create geneticin-resistant stable cell lines. Three stable cell lines were shown by western blot analysis to have significant down-regulation of RAD51 to 20-50% of the levels expressed in control cell lines. All three cell lines had a similar increased sensitivity to gamma-irradiation by 70 and 40%, respectively, compared to normal and empty vector-transfected cells, corresponding to dose-modifying factors of approximately 2.0 and 1.5 in the mid-range of the dose-response curves. The amount of RAD51 protein in transfected cell lines was shown to strongly correlate with the alpha parameter obtained from fitted survival curves. These results highlight the importance of RAD51 in cellular responses to radiation and are the first to indicate the potential use of RAD51-targeted ribozyme minigenes in tumour radiosensitisation. PMID- 11266556 TI - Trends in lipoplex physical properties dependent on cationic lipid structure, vehicle and complexation procedure do not correlate with biological activity. AB - Using a group of structurally related cytofectins, the effects of different vehicle constituents and mixing techniques on the physical properties and biological activity of lipoplexes were systematically examined. Physical properties were examined using a combination of dye accessibility assays, centrifugation, gel electrophoresis and dynamic light scattering. Biological activity was examined using in vitro transfection. Lipoplexes were formulated using two injection vehicles commonly used for in vivo delivery (PBS pH 7.2 and 0.9% saline), and a sodium phosphate vehicle previously shown to enhance the biological activity of naked pDNA and lipoplex formulations. Phosphate was found to be unique in its effect on lipoplexes. Specifically, the accessible pDNA in lipoplexes formulated with cytofectins containing a gamma-amine substitution in the headgroup was dependent on alkyl side chain length and sodium phosphate concentration, but the same effects were not observed when using cytofectins containing a beta-OH headgroup substitution. The physicochemical features of the phosphate anion, which give rise to this effect in gamma-amine cytofectins, were deduced using a series of phosphate analogs. The effects of the formulation vehicle on transfection were found to be cell type-dependent; however, of the formulation variables examined, the liposome/pDNA mixing method had the greatest effect on transgene expression in vitro. Thus, though predictive physical structure relationships involving the vehicle and cytofectin components of the lipoplex were uncovered, they did not extrapolate to trends in biological activity. PMID- 11266557 TI - Characterization of uracil-DNA glycosylase activity from Trypanosoma cruzi and its stimulation by AP endonuclease. AB - The intracellular pathogen Trypanosoma cruzi is the etiological agent of Chagas' disease. We have isolated a full-length cDNA encoding uracil-DNA glycosylase (UDGase), a key enzyme involved in DNA repair, from this organism. The deduced protein sequence is highly conserved at the C-terminus of the molecule and shares key residues involved in binding or catalysis with most of the UDGases described so far, while the N-terminal part is highly variable. The gene is single copy and is located on a chromosome of approximately 1.9 Mb. A His-tagged recombinant protein was overexpressed, purified and used to raise polyclonal antibodies. Western blot analysis revealed the existence of a single UDGase species in parasite extracts. Using a specific ethidium bromide fluorescence assay, recombinant T.cruzi UDGase was shown to specifically excise uracil from DNA. The addition of both Leishmania major AP endonuclease and exonuclease III, the major AP endonuclease from Escherichia coli, produces stimulation of UDGase activity. This activation is specific for AP endonuclease and suggests functional communication between the two enzymes. PMID- 11266559 TI - Enhancing the catalytic repertoire of nucleic acids: a systematic study of linker length and rigidity. AB - The incorporation of potentially catalytic groups in DNA is of interest for the in vitro selection of novel deoxyribozymes. A series of 10 C5-modified analogues of 2'-deoxyuridine triphosphate have been synthesised that possess side chains of differing flexibility and bearing a primary amino or imidazole functionality. For each series of nucleotide analogues differing degrees of flexibility of the C5 side chain was achieved through the use of alkynyl, alkenyl and alkyl moieties. The imidazole function was conjugated to these C5-amino-modified nucleotides using either imidazole 4-acetic acid or imidazole 4-acrylic acid (urocanic acid). The substrate properties of the nucleotides (fully replacing dTTP) with TAQ polymerase during PCR have been investigated in order to evaluate their potential applications for in vitro selection experiments. 5-(3-Aminopropynyl)dUTP and 5-(E 3-aminopropenyl)dUTP and their imidazole 4-acetic acid- and urocanic acid modified conjugates were found to be substrates. In contrast, C5-amino-modified dUTPs with alkane or Z-alkene linkers and their corresponding conjugates were not substrates. The incorporation of these analogues during PCR has been confirmed by inhibition of restriction enzyme digestion using XBAI and by mass spectrometry of the PCR products. PMID- 11266558 TI - Trypanosome spliced leader RNA genes contain the first identified RNA polymerase II gene promoter in these organisms. AB - Typical general transcription factors, such as TATA binding protein and TFII B, have not yet been identified in any member of the Trypanosomatidae family of parasitic protozoa. Interestingly, mRNA coding genes do not appear to have discrete transcriptional start sites, although in most cases they require an RNA polymerase that has the biochemical properties of eukaryotic RNA polymerase II. A discrete transcription initiation site may not be necessary for mRNA synthesis since the sequences upstream of each transcribed coding region are trimmed from the nascent transcript when a short m(7)G-capped RNA is added during mRNA maturation. This short 39 nt m(7)G-capped RNA, the spliced leader (SL) sequence, is expressed as an approximately 100 nt long RNA from a set of reiterated, though independently transcribed, genes in the trypanosome genome. Punctuation of the 5' end of mRNAs by a m(7)G cap-containing spliced leader is a developing theme in the lower eukaryotic world; organisms as diverse as EUGLENA: and nematode worms, including Caenorhabditis elegans, utilize SL RNA in their mRNA maturation programs. Towards understanding the coordination of SL RNA and mRNA expression in trypanosomes, we have begun by characterizing SL RNA gene expression in the model trypanosome Leptomonas seymouri. Using a homologous in vitro transcription system, we demonstrate in this study that the SL RNA is transcribed by RNA polymerase II. During SL RNA transcription, accurate initiation is determined by an initiator element with a loose consensus of CYAC/AYR(+1). This element, as well as two additional basal promoter elements, is divergent in sequence from the basal transcription elements seen in other eukaryotic gene promoters. We show here that the in vitro transcription extract contains a binding activity that is specific for the initiator element and thus may participate in recruiting RNA polymerase II to the SL RNA gene promoter. PMID- 11266560 TI - Different dynamics in nuclear entry of subunits of the repair/transcription factor TFIIH. AB - We report here the different ways in which four subunits of the basal transcription/repair factor TFIIH (XPB, XPD, p62 and p44) and the damage recognition XPC repair protein can enter the nucleus. We examined their nuclear localization by transiently expressing the gene products tagged with the enhanced green fluorescent protein (EGFP) in transfected 3T3 cells. In agreement with the identification of more than one putative nuclear localization signal (NLS) in their protein sequences, XPB, XPC, p62 and p44 chimeras were rapidly sorted to the nucleus. In contrast, the XPD-EGFP chimeras appeared mainly localized in the cytoplasm, with a minor fraction of transfectants showing the EGFP-based fluorescence also in the nucleus. The ability of the XPD chimeras to enter the nucleus was confirmed by western blotting on fractionated cell extracts and by functional complementation of the repair defect in the UV5 rodent cells, mutated in the XPD homologous gene. By deletion mutagenesis, we were unable to identify any sequence specific for nuclear localization. In particular, deletion of the putative NLS failed to affect subcellular localization and, conversely, the C terminal part of XPD containing the putative NLS showed no specific nuclear accumulation. These findings suggest that the nuclear entry of XPD depends on its complexation with other proteins in the cytoplasm, possibly other components of the TFIIH complex. PMID- 11266561 TI - Mitochondrial DNA ligase function in Saccharomyces cerevisiae. AB - The Saccharomyces cerevisiae CDC9 gene encodes a DNA ligase protein that is targeted to both the nucleus and the mitochondria. While nuclear Cdc9p is known to play an essential role in nuclear DNA replication and repair, its role in mitochondrial DNA dynamics has not been defined. It is also unclear whether additional DNA ligase proteins are present in yeast mitochondria. To address these issues, mitochondrial DNA ligase function in S.cerevisiae was analyzed. Biochemical analysis of mitochondrial protein extracts supported the conclusion that Cdc9p was the sole DNA ligase protein present in this organelle. Inactivation of mitochondrial Cdc9p function led to a rapid decline in cellular mitochondrial DNA content in both dividing and stationary yeast cultures. In contrast, there was no apparent defect in mitochondrial DNA dynamics in a yeast strain deficient in Dnl4p (Deltadnl4). The Escherichia coli ECO:RI endonuclease was targeted to yeast mitochondria. Transient expression of this recombinant ECO:RI endonuclease led to the formation of mitochondrial DNA double-strand breaks. While wild-type and Deltadnl4 yeast were able to rapidly recover from this mitochondrial DNA damage, clones deficient in mitochondrial Cdc9p were not. These results support the conclusion that yeast rely upon a single DNA ligase, Cdc9p, to carry out mitochondrial DNA replication and recovery from both spontaneous and induced mitochondrial DNA damage. PMID- 11266562 TI - Highly expressed and alien genes of the Synechocystis genome. AB - Comparisons of codon frequencies of genes to several gene classes are used to characterize highly expressed and alien genes on the SYNECHOCYSTIS: PCC6803 genome. The primary gene classes include the ensemble of all genes (average gene), ribosomal protein (RP) genes, translation processing factors (TF) and genes encoding chaperone/degradation proteins (CH). A gene is predicted highly expressed (PHX) if its codon usage is close to that of the RP/TF/CH standards but strongly deviant from the average gene. Putative alien (PA) genes are those for which codon usage is significantly different from all four classes of gene standards. In SYNECHOCYSTIS:, 380 genes were identified as PHX. The genes with the highest predicted expression levels include many that encode proteins vital for photosynthesis. Nearly all of the genes of the RP/TF/CH gene classes are PHX. The principal glycolysis enzymes, which may also function in CO(2) fixation, are PHX, while none of the genes encoding TCA cycle enzymes are PHX. The PA genes are mostly of unknown function or encode transposases. Several PA genes encode polypeptides that function in lipopolysaccharide biosynthesis. Both PHX and PA genes often form significant clusters (operons). The proteins encoded by PHX and PA genes are described with respect to functional classifications, their organization in the genome and their stoichiometry in multi-subunit complexes. PMID- 11266563 TI - Phylogenetic analysis of tmRNA genes within a bacterial subgroup reveals a specific structural signature. AB - Bacterial tmRNA mediates a trans-translation reaction, which permits the recycling of stalled ribosomes and probably also contributes to the regulated expression of a subset of genes. Its action results in the addition of a small number of C-terminal amino acids to protein whose synthesis had stalled and these constitute a proteolytic recognition tag for the degradation of these incompletely synthesized proteins. Previous work has identified pseudoknots and stem-loops that are widely conserved in divergent bacteria. In the present work an alignment of tmRNA gene sequences within 13 beta-proteobacteria reveals an additional sub-structure specific for this bacterial group. This sub-structure is in pseudoknot Pk2, and consists of one to two additional stem-loop(s) capped by stable GNRA tetraloop(s). Three-dimensional models of tmRNA pseudoknot 2 (Pk2) containing various topological versions of the additional sub-structure suggest that the sub-structures likely point away from the core of the RNA, containing both the tRNA and the mRNA domains. A putative tertiary interaction has also been identified. PMID- 11266564 TI - Identification of thermophilic species by the amino acid compositions deduced from their genomes. AB - The global amino acid compositions as deduced from the complete genomic sequences of six thermophilic archaea, two thermophilic bacteria, 17 mesophilic bacteria and two eukaryotic species were analysed by hierarchical clustering and principal components analysis. Both methods showed an influence of several factors on amino acid composition. Although GC content has a dominant effect, thermophilic species can be identified by their global amino acid compositions alone. This study presents a careful statistical analysis of factors that affect amino acid composition and also yielded specific features of the average amino acid composition of thermophilic species. Moreover, we introduce the first example of a 'compositional tree' of species that takes into account not only homologous proteins, but also proteins unique to particular species. We expect this simple yet novel approach to be a useful additional tool for the study of phylogeny at the genome level. PMID- 11266565 TI - GenEST, a powerful bidirectional link between cDNA sequence data and gene expression profiles generated by cDNA-AFLP. AB - The release of vast quantities of DNA sequence data by large-scale genome and expressed sequence tag (EST) projects underlines the necessity for the development of efficient and inexpensive ways to link sequence databases with temporal and spatial expression profiles. Here we demonstrate the power of linking cDNA sequence data (including EST sequences) with transcript profiles revealed by cDNA-AFLP, a highly reproducible differential display method based on restriction enzyme digests and selective amplification under high stringency conditions. We have developed a computer program (GenEST) that predicts the sizes of virtual transcript-derived fragments (TDFs) of in silico-digested cDNA sequences retrieved from databases. The vast majority of the resulting virtual TDFs could be traced back among the thousands of TDFs displayed on cDNA-AFLP gels. Sequencing of the corresponding bands excised from cDNA-AFLP gels revealed no inconsistencies. As a consequence, cDNA sequence databases can be screened very efficiently to identify genes with relevant expression profiles. The other way round, it is possible to switch from cDNA-AFLP gels to sequences in the databases. Using the restriction enzyme recognition sites, the primer extensions and the estimated TDF size as identifiers, the DNA sequence(s) corresponding to a TDF with an interesting expression pattern can be identified. In this paper we show examples in both directions by analyzing the plant parasitic nematode Globodera rostochiensis. Various novel pathogenicity factors were identified by combining ESTs from the infective stage juveniles with expression profiles of approximately 4000 genes in five developmental stages produced by cDNA-AFLP. PMID- 11266566 TI - Identification of 10 novel snoRNA gene clusters from Arabidopsis thaliana. AB - Ten novel small nucleolar RNA (snoRNA) gene clusters, consisting of two or three snoRNA genes, respectively, were identified from Arabidopsis thaliana. Twelve of the 25 snoRNA genes in these clusters are homologous to those of yeast and mammals according to the conserved antisense sequences that guide 2'-O-ribose methylation of rRNA. The remaining 13 snoRNA genes, including two 5.8S rRNA methylation guides, are new genes identified from A.thaliana. Interestingly, seven methylated nucleotides, predicted by novel snoRNAs Z41a-Z46, are methylated neither in yeast nor in vertebrates. Using primer extension at low dNTP concentration the six methylation sites were determined as expected. These snoRNAs were recognized as specific guides for 2'-O:-ribose methylation of plant rRNAs. Z42, however, did not guide the expected methylation of 25S rRNA in our assay. Thus, its function remains to be elucidated. The intergenic spacers of the gene clusters are rich in uridine (up to 40%) and most of them range in size from 35 to 100 nt. Lack of a conserved promoter element in each spacer and the determination of polycistronic transcription from a cluster by RT-PCR assay suggest that the snoRNAs encoded in the clusters are transcribed as a polycistron under an upstream promoter, and individual snoRNAs are released after processing of the precursor. Numerous snoRNA gene clusters identified from A.thaliana and other organisms suggest that the snoRNA gene cluster is an ancient gene organization existing abundantly in plants. PMID- 11266567 TI - Cooperative binding of effectors by an allosteric ribozyme. AB - An allosteric ribozyme that requires two different effectors to induce catalysis was created using modular rational design. This ribozyme construct comprises five conjoined RNA modules that operate in concert as an obligate FMN- and theophylline-dependent molecular switch. When both effectors are present, this 'binary' RNA switch self-cleaves with a rate enhancement of approximately 300 fold over the rate observed in the absence of effectors. Kinetic and structural studies implicate a switching mechanism wherein FMN binding induces formation of the active ribozyme conformation. However, the binding site for FMN is rendered inactive unless theophylline first binds to its corresponding site and reorganizes the RNA structure. This example of cooperative binding between allosteric effectors reveals a level of structural and functional complexity for RNA that is similar to that observed with allosteric proteins. PMID- 11266569 TI - ParAlign: a parallel sequence alignment algorithm for rapid and sensitive database searches. AB - There is a need for faster and more sensitive algorithms for sequence similarity searching in view of the rapidly increasing amounts of genomic sequence data available. Parallel processing capabilities in the form of the single instruction, multiple data (SIMD) technology are now available in common microprocessors and enable a single microprocessor to perform many operations in parallel. The ParAlign algorithm has been specifically designed to take advantage of this technology. The new algorithm initially exploits parallelism to perform a very rapid computation of the exact optimal ungapped alignment score for all diagonals in the alignment matrix. Then, a novel heuristic is employed to compute an approximate score of a gapped alignment by combining the scores of several diagonals. This approximate score is used to select the most interesting database sequences for a subsequent Smith-Waterman alignment, which is also parallelised. The resulting method represents a substantial improvement compared to existing heuristics. The sensitivity and specificity of ParAlign was found to be as good as Smith-Waterman implementations when the same method for computing the statistical significance of the matches was used. In terms of speed, only the significantly less sensitive NCBI BLAST 2 program was found to outperform the new approach. Online searches are available at http://dna.uio.no/search/ PMID- 11266568 TI - Ku80 is required for addition of N nucleotides to V(D)J recombination junctions by terminal deoxynucleotidyl transferase. AB - V(D)J recombination generates a remarkably diverse repertoire of antigen receptors through the rearrangement of germline DNA. Terminal deoxynucleotidyl transferase (TdT), a polymerase that adds random nucleotides (N regions) to recombination junctions, is a key enzyme contributing to this diversity. The current model is that TdT adds N regions during V(D)J recombination by random collision with the DNA ends, without a dependence on other cellular factors. We previously demonstrated, however, that V(D)J junctions from Ku80-deficient mice unexpectedly lack N regions, although the mechanism responsible for this effect remains undefined in the mouse system. One possibility is that junctions are formed in these mice during a stage in development when TdT is not expressed. Alternatively, Ku80 may be required for the expression, nuclear localization or enzymatic activity of TdT. Here we show that V(D)J junctions isolated from Ku80 deficient fibroblasts are devoid of N regions, as were junctions in Ku80 deficient mice. In these cells TdT protein is abundant at the time of recombination, localizes properly to the nucleus and is enzymatically active. Based on these data, we propose that TdT does not add to recombination junctions through random collision but is actively recruited to the V(D)J recombinase complex by Ku80. PMID- 11266570 TI - The length of telomeric G-rich strand 3'-overhang measured by oligonucleotide ligation assay. AB - A typical G-rich telomeric DNA strand, which runs 5'-->3' toward the chromosome ends, protrudes by several nucleotides in lower eukaryotes. In human chromosomes long G-rich 3'-overhangs have been found. Apart from the standard G-rich tail, several non-canonical terminal structures have been proposed. However, the mechanism of long-tail formation, the presence and the role of these structures in telomere maintenance or shortening are not completely understood. In a search for a simple method to accurately measure the 3'-overhang we have established a protocol based on the ligation of telomeric oligonucleotide hybridized to non denatured DNA under stringent conditions (oligonucleotide ligation assay with telomeric repeat oligonucleotide). This method enabled us to detect a large proportion of G-rich single-stranded telomeric DNA that was as short as 24 nt. Nevertheless, we showed G-tails longer than 400 nt. In all tested cells the lengths ranging from 108 to 270 nt represented only 37% of the whole molecule population, while 56-62% were <90 nt. Our protocol provides a simple and sensitive method for measuring the length of naturally occurring unpaired repeated DNA. PMID- 11266571 TI - Detection of mitochondrial single nucleotide polymorphisms using a primer elongation reaction on oligonucleotide microarrays. AB - We have developed a novel allele-specific primer elongation protocol using a DNA polymerase on oligonucleotide chips. Oligonucleotide primers carrying polymorphic sites at their free 3'end were covalently bound to glass slides. The generation of single-stranded targets of genomic DNA containing single nuclotide polymorphisms (SNPs) to be typed was achieved by an asymmetric PCR reaction or exonuclease treatment of phosphothioate (PTO)-modified PCR products. In the presence of DNA polymerase and all four dNTPs, with Cy3-dUTP replacing dTTP, allele-specific extension of the immobilized primers took place along a stretch of target DNA sequence. The yield of elongated products was increased by repeated reaction cycles. We performed multiplexed assays with many small DNA targets, or used single targets of up to 4.4 kb mitochondrial DNA (mtDNA) sequence to detect multiple SNPs in one reaction. The latter approach greatly simplifies preamplification of SNP-containing regions, thereby providing a framework for typing hundreds of mtDNA polymorphisms. PMID- 11266572 TI - Assembly of large genomic segments in artificial chromosomes by homologous recombination in Escherichia coli. AB - We developed a method for the reconstruction of a 100 kb DNA fragment into a bacterial artificial chromosome (BAC). The procedure makes use of iterative rounds of homologous recombination in Escherichia coli. Smaller, overlapping fragments of cloned DNA, such as cosmid clones, are required. They are transferred first into a temperature-sensitive replicon and then into the BAC of choice. We demonstrated the usefulness of this procedure by assembling a 90 kb genomic segment into an E.coli-STREPTOMYCES: artificial chromosome (ESAC). Using this procedure, ESACs are easy to handle and remarkably more stable than the starting cosmids. PMID- 11266573 TI - Manufacturing DNA microarrays of high spot homogeneity and reduced background signal. AB - Analyses on DNA microarrays depend considerably on spot quality and a low background signal of the glass support. By using betaine as an additive to a spotting solution made of saline sodium citrate, both the binding efficiency of spotted PCR products and the homogeneity of the DNA spots is improved significantly on aminated surfaces such as glass slides coated with the widely used poly-L-lysine or aminosilane. In addition, non-specific background signal is markedly diminished. Concomitantly, during the arraying procedure, the betaine reduces evaporation from the microtitre dish wells, which hold the PCR products. Subsequent blocking of the chip surface with succinic anhydride was improved considerably in the presence of the non-polar, non-aqueous solvent 1,2 dichloroethane and the acylating catalyst N:-methylimidazole. This procedure prevents the overall background signal that occurs with the frequently applied aqueous solvent 1-methyl-2-pyrrolidone in borate buffer because of DNA that re dissolves from spots during the blocking process, only to bind again across the entire glass surface. PMID- 11266574 TI - A GFP-equipped bidirectional expression module well suited for monitoring tetracycline-regulated gene expression in mouse. AB - Doxycycline (Dox)-sensitive co-regulation of two transcriptionally coupled transgenes was investigated in the mouse. For this, we generated four independent mouse lines carrying coding regions for green fluorescent protein (GFP) and beta galactosidase in a bicistronic, bidirectional module. In all four lines the expression module was silent but was activated when transcription factor tTA was provided by the alpha-CaMKII-tTA transgene. In vivo analysis of GFP fluorescence, beta-galactosidase and immunochemical stainings revealed differences in GFP and beta-galactosidase levels between the lines, but comparable patterns of expression. Strong signals were found in neurons of the olfactory system, neocortical, limbic lobe and basal ganglia structures. Weaker expression was limited to thalamic, pontine and medullary structures, the spinal cord, the eye and to some Purkinje cells in the cerebellum. Strong GFP signals were always accompanied by intense beta-galactosidase activity, both of which could be co regulated by Dox. We conclude that the tTA-sensitive bidirectional expression module is well suited to express genes of interest in a regulated manner and that GFP can be used to track transcriptional activity of the module in the living mouse. PMID- 11266576 TI - Role of the putative membrane-bound endo-1,4-beta-glucanase KORRIGAN in cell elongation and cellulose synthesis in Arabidopsis thaliana. AB - A temperature-sensitive, elongation-deficient mutant of Arabidopsis thaliana was isolated. At the non-permissive temperature of 31 degrees C, the mutation impaired tissue elongation; otherwise, tissue development was normal. Hypocotyl cells that had established cell walls at 21 degrees C under light-dark cycles ceased elongation and swelled when the mutant was shifted to 31 degrees C and darkness, indicating that the affected gene is essential for cell elongation. Analysis of the cell walls of mutant plants grown at 31 degrees C revealed that the cellulose content was reduced to 40% and the pectin content was increased to 162% of the corresponding values for the wild type grown at the same temperature. The increased amounts of pectin in the mutant were bound tightly to cellulose microfibrils. No change in the content of hemicellulose was apparent in the 31 degrees C-adapted mutant. Field emission-scanning electron microscopy suggested that the structure of cellulose bundles was affected by the mutation; X-ray diffraction, however, revealed no change in the crystallite size of cellulose microfibrils. The regeneration of cellulose microfibrils from naked mutant protoplasts was substantially delayed at 31 degrees C. The recessive mutation was mapped to chromosome V, and map-based cloning identified it as a single G-->A transition (resulting in a Gly(429)-->Arg substitution) in KORRIGAN, which encodes a putative membrane-bound endo-1,4-beta-glucanase. These results demonstrate that the product of this gene is required for cellulose synthesis. PMID- 11266575 TI - Efficient gene activation in cultured mammalian cells mediated by FLP recombinase expressing recombinant adenovirus. AB - A recombinant adenovirus (rAd) expressing Cre recombinase derived from bacteriophage P1 has already been extensively used for the conditional gene activation and inactivation strategies in mammalian systems. In this study, we generated AxCAFLP, a rAd expressing FLP recombinase derived from Saccharomyces cerevisiae and carried out quantitative comparisons with Cre-expressing rAd in both in vitro and in cultured cells to provide another efficient gene regulation system in mammalian cells. In the in vitro experiments, the relative recombination efficiency of FLP expressed in 293 cells infected with FLP expressing rAd was approximately one-thirtieth that of Cre even at 30 degrees C, the optimum temperature for FLP activity, and was approximately one-ninetieth at 37 degrees C. Co-infection experiments in HeLa cells using a target rAd conditionally expressing LacZ under the control of FLP showed that an FLP expressing rAd, infected at a multiplicity of infection (MOI) of 5, was able to activate the transgene in almost 100% of HeLa cells whereas the Cre-expressing rAd was sufficient at an MOI of 0.2. Since an MOI of 5 is ordinarily used in rAd experiments, these results showed that the FLP-expressing rAd is useful for gene activation strategies and is probably applicable to a sequential gene regulation system in combination with Cre-expressing rAd in mammalian cells. PMID- 11266577 TI - The chloroplast clpP gene, encoding a proteolytic subunit of ATP-dependent protease, is indispensable for chloroplast development in tobacco. AB - ClpP is a proteolytic subunit of the ATP-dependent Clp protease, which is found in chloroplasts in higher plants. Proteolytic subunits are encoded both by the chloroplast gene, clpP, and a nuclear multi gene family. We insertionally disrupted clpP by chloroplast transformation in tobacco. However, complete segregation was impossible, indicating that the chloroplast-encoded clpP gene has an indispensable function for cell survival. In the heteroplasmic clpP disruptant, the leaf surface was rough by clumping, and the lateral leaf expansion was irregularly arrested, which led to an asymmetric, slender leaf shape. Chloroplasts consisted of two populations: chloroplasts that were similar to the wild type, and small chloroplasts that emitted high chl fluorescence. Ultrastructural analysis of chloroplast development suggested that clpP disruption also induced swelling of the thylakoid lumen in the meristem plastids and inhibition of etioplast development in the dark. In mature leaves, thylakoid membranes of the smaller chloroplast population consisted exclusively of large stacks of tightly appressed membranes. These results indicate that chloroplast encoded ClpP is involved in multiple processes of chloroplast development, including a housekeeping function that is indispensable for cell survival. PMID- 11266578 TI - Blue light-induced branching in Vaucheria. Requirement of nuclear accumulation in the irradiated region. AB - When a narrow region of the fresh water coenocytic alga, Vaucheria terrestris sensu Gotz is irradiated with moderately intense blue light, a branch is induced from the center of the irradiated region after 4-5 h. Movement of organelles and microtubule bundles during the photocytomorphogenetic response were investigated. Chloroplasts in the cortical layer immediately started to accumulate in the blue light-irradiated region and their accumulation almost completely finished 30-40 min after the onset of light when the nuclei residing in endoplasm started to accumulate. Accumulation of nuclei was synchronized with disorientation and shortening of microtubule bundles, which originally run parallel to the cell axis. Not only amiprophos-methyl, a potent microtubule-decomposing reagent, but also cytochalasin A strongly inhibited the branch induction. Amiprophos-methyl completely and cytochalasin A mostly destroyed microtubules and completely inhibited nuclear accumulation, but both drugs allowed the accumulation of chloroplasts in the cortical layer of irradiated region. These indicate that the accumulation of nuclei is indispensable for branch induction while the chloroplast accumulation is insufficient by itself for branch induction. The ineffectiveness of cytochalasin A on chloroplast movement brings the conventional view of sliding movement of chloroplast on a long actin cable into question. The morphological and functional relationship between a nucleus and a microtubular bundle are discussed. PMID- 11266579 TI - A mechanism for promoting the germination of Zinnia elegans seeds by hydrogen peroxide. AB - H(2)O(2) promotes seed germination of cereal plants such as barley, wheat and rice, and several mechanisms have been proposed for its action [Naredo et al. (1998) Seed Sci. Technol. 26: 675-689]. We investigated the role of H(2)O(2) in the germination of Zinnia elegans seeds. H(2)O(2) promoted seed germination in a dose-dependent manner as did respiratory inhibitors, indicating that H(2)O(2) itself possibly promotes seed germination rather than O(2). Seed germination was promoted by removal of pericarp from seeds or by removal of ethanol-soluble compounds from the seeds with pericarp. The ethanol-soluble compounds suppressed the germination of seeds having no pericarp, and this effect was reversed by H(2)O(2). These findings indicate that oxidation of the germination inhibitor(s) present in the pericarp by H(2)O(2) promotes seed germination. Antioxidants which are derivatives of well-known germination inhibitors suppressed seed germination in a dose-dependent manner, suggesting that, to initiate seed germination, a germination inhibitor(s) should be decomposed by an oxidant such as H(2)O(2). PMID- 11266580 TI - Endoxyloglucan transferase is localized both in the cell plate and in the secretory pathway destined for the apoplast in tobacco cells. AB - Intracellular trafficking of enzymes responsible for constructing and modifying the cell wall architecture in plants is mostly unknown. To examine their translocation pathways, we employed an endoxyloglucan transferase (EXGT), a key enzyme responsible for forming and rearranging the cellulose/xyloglucan network of the cell wall. We traced its intracellular localization in suspension-cultured cells of tobacco bright yellow-2 by means of green fluorescent protein-fusion gene procedures as well as by indirect immunofluorescence. During interphase the protein was extensively secreted into the apoplast via the endoplasmic reticulum Golgi apparatus network, whereas during cytokinesis, the protein was exclusively located in the phragmoplast and eventually transported to the cell plate. These results clearly indicate commitment of EXGT protein to the construction of both the cell plate and the cell wall. This study also visualized the process of phragmoplast development at a level of vesicle translocation in the living cell. PMID- 11266581 TI - Auxin is a positive regulator for ethylene-mediated response in the growth of Arabidopsis roots. AB - The requirement of auxin for the ethylene-mediated growth response in the root of Arabidopsis thaliana seedlings was investigated using two ethylene-resistant mutants, aux1-7 and eir1-1, whose roots have been shown to have a defect in the auxin influx and efflux carriers, respectively. A 50% inhibition of growth (I(50)) was achieved with 0.84 microl liter(-1) ethylene in wild-type roots, but 71.3 microl liter( -1) ethylene was required to induce I(50) in eir1-1 roots. In aux1-7 roots, I(50) was not obtained even at 1,000 microl liter(-1) ethylene. By contrast, in the presence of 10 nM 1-naphthaleneacetic acid (NAA), the concentrations of ethylene required to induce I(50) in eir1-1 and aux1-7 roots were greatly reduced nearly to the level required in wild-type roots. Since the action of NAA to restore the ethylene response in aux1-7 roots was not replaced by IAA, an increase in the intracellular level of auxin is likely to be the cause for the restoration of ethylene response. NAA at 10 nM did not inhibit root growth when applied solely, but it was the optimum concentration to recover the ethylene response in the mutant roots. These results suggest that auxin is a positive regulator for ethylene-induced inhibition in root elongation. PMID- 11266582 TI - Molecular cloning and characterization of a cDNA for a novel ethylene receptor, NT-ERS1, of tobacco (Nicotiana tabacum L.). AB - The cDNA encoding a novel member (NT-ERS1) of ethylene receptor family of tobacco (Nicotiana tabacum L.) was obtained by a combination of RT-PCR and 5'-/3'-RACE cloning. The cDNA was 2,092 nucleotides long and had an open reading frame of 1,911 bp encoding 637 amino acids. The deduced polypeptide lacked a response regulator domain, indicating that the ethylene receptor belongs to an ERS-group. The amino acid sequence was similar to respective members of the tobacco ethylene receptor family: 67.8% to NT-ETR1, 39.1% to NTHK1 and 31.1% to NTHK2. Comparison of amino acid sequence suggested that NT-ERS1 is the counterpart of Nr in the ethylene receptor family of tomato, which belongs to Solanaceae as does tobacco. Northern blot analysis showed that mRNA of NT-ERS1 was present in leaf, shoot and root tissues, and accumulated in leaves treated with exogenous ethylene. A mutated NT-ERS1 cDNA transgene, obtained by introducing one nucleotide substitution into NT-ETR1 cDNA, conferred ethylene insensitivity in tobacco plants, indicating that the translation product of the cDNA actually functioned in the plants. PMID- 11266584 TI - Characterization of oxalate oxidase and cell death in Al-sensitive and tolerant wheat roots. AB - Several genes including oxalate oxidase (Oxo) are up-regulated in Triticum aestivum L. root tips exposed to Al. To better understand the function of Oxo during Al exposure, the protein level and enzyme activity were measured. The data indicate that both Oxo protein and activity are increased proportionally to the level of root growth inhibition (RGI). A high level of Oxo expression may result in excess H(2)O(2) production which could become toxic and induce cell death. However, the timing of H(2)O(2) production (observed after 24 h) indicates that it cannot be the primary cause of cell death first observed after 8 h. Moreover, at Al concentrations resulting in 50% RGI, we did not observe any cell death in the sensitive cultivar while a punctated pattern of death involving small groups of cells was found in the tolerant cultivar. This pattern was maintained for several days in the tolerant cultivar, suggesting the involvement of a cell death mechanism aimed at replacing epidermal cells intoxicated with Al while root growth is maintained. The accelerated epidermal cell turnover may represent a new detoxification mechanism helping to protect deeper cell layers of the meristematic and elongation zone essential for root growth. PMID- 11266583 TI - The regulation of ornithine decarboxylase gene expression by sucrose and small upstream open reading frame in tomato (Lycopersicon esculentum Mill). AB - We identified a near-full-length cDNA clone encoding ornithine decarboxylase (ODC) from tomato (Lycopersicon esculentum Mill). It contained a small upstream open reading frame (uORF) within its 5' untranslated region. An in vitro translation assay demonstrated that the uORF repressed expression of downstream ORF. Neither nucleotide nor predicted peptide sequence of the uORF was responsible for the repression. The presence of upstream AUG codon was shown to be responsible. ODC expression appeared to be organ specific. The ODC gene was expressed in roots, hypocotyls and sink leaves but not in source leaves. ODC transcripts were observed in apical meristem of primary roots, and were distributed in cells of cortex layer preferentially. ODC expression responded immediately to sucrose availability via the sucrose-specific pathway independent of hexokinase. Sucrose induction of ODC gene was seen in roots, hypocotyls and flowers but not in mature leaves. Moreover, only the root apical meristem responded to sucrose availability. These observations indicate that the spatial pattern of ODC expression is closely associated with cell proliferation and that sucrose sensing plays a major role in the spatial pattern of ODC expression. Also, the differential regulation of ODC and arginine decarboxylase gene expression by factors modulating plant growth suggests that they would have different physiological roles in plant development. PMID- 11266585 TI - Light response of the circadian waves of the APRR1/TOC1 quintet: when does the quintet start singing rhythmically in Arabidopsis? AB - We previously identified a novel class of proteins, named A:rabidopsis pseudo response regulators (APRRs), each of which (APRR1/TOC1, APRR3, APRR5, APRR7, APRR9) has an intriguing structural design containing an N-terminal pseudo receiver domain and a C-terminal CONSTANS motif. Expression of these APRR1/TOC1 family members is under the control of a coordinate circadian rhythm at the level of transcription such that the APRR-mRNAs start accumulating sequentially after dawn with 2 to 3 h intervals in the order of APRR9-->APRR7-->APRR5-->APRR3- >APRR1/TOC1 in a given 24 h photo-period. Based on these data, we previously proposed that these sequential and rhythmic events of transcription, termed 'circadian waves of APRR1/TOC1 quintet', may be a basis of a presumed Arabidopsis biological clock (named 'bar code clock') [Matsushika et al. (2000) Plant and Cell Physiol. 41: 1002]. Here we further characterized the event of circadian waves, by demonstrating that certain light stimuli are crucial determinants to induce the robust circadian waves, and accordingly, the first-boosted and light induced APRR9 appears to be primarily responsible for this light response of the circadian waves. Also, as such a light stimulus, a red light pulse that is presumably perceived by phytochromes appears to be sufficient to induce (or synchronize) the APRR1/TOC1 circadian waves. PMID- 11266586 TI - Mapping the virus and host genes involved in the resistance response in cucumber mosaic virus-Infected Arabidopsis thaliana. AB - A yellow strain of cucumber mosaic virus (CMV) [CMV(Y)] induces a resistance response characterized by inhibition of virus systemic movement with development of necrotic local lesions in the virus-inoculated leaves of Arabidopsis thaliana ecotype C24. In this report, the avirulence determinant in the virus genome was defined and the resistance gene (RCY1) of C24 was genetically mapped. The response of C24 to CMV containing the chimeric RNA3 between CMV(Y) and a virulent strain of CMV indicated that the coat protein gene of CMV(Y) determined the localization of the virus in the inoculated leaves of C24. The RCY1 locus was mapped between two CAPS markers, DFR and T43968, which were located in the region containing genetically defined disease resistance genes and their homologues. These results indicate that the resistance response to CMV(Y) in C24 is determined by the combination of the coat protein gene and RCY1 on chromosome 5. PMID- 11266588 TI - The Hans Neurath Award lecture of The Protein Society: proteins-- a testament to physics, chemistry, and evolution. PMID- 11266589 TI - Second virial coefficients as a measure of protein--osmolyte interactions. AB - The cytoplasm contains high concentrations of cosolutes. These cosolutes include macromolecules and small organic molecules called osmolytes. However, most biophysical studies of proteins are conducted in dilute solutions. Two broad classes of models have been used to describe the interaction between osmolytes and proteins. One class focuses on excluded volume effects, while the other focuses on binding between the protein and the osmolyte. To better understand protein--smolyte interactions, we have conducted sedimentation equilibrium analytical ultracentrifugation experiments using ferricytochrome c as a model protein. From these experiments, we determined the second virial coefficients for a series of osmolytes. We have interpreted the second virial coefficient as a measure of both excluded volume and protein--osmolyte binding. We conclude that simple models are not sufficient to understand the interactions between osmolytes and proteins. PMID- 11266590 TI - Crystal structure of an archaeal DNA sliding clamp: proliferating cell nuclear antigen from Pyrococcus furiosus. AB - The proliferating cell nuclear antigen (PCNA) is now recognized as one of the key proteins in DNA metabolic events because of its direct interactions with many proteins involved in important cellular processes. We have determined the crystal structure of PCNA from a hyperthermophilic archaeon, Pyrococcus furiosus (pfuPCNA), at 2.1 A resolution. pfuPCNA forms a toroidal, ring-shaped structure consisting of homotrimeric molecules, which is also observed in the PCNA crystals from human and yeast. The overall structure of pfuPCNA is highly conserved with other PCNA proteins, as well as with the bacterial ss clamp and the bacteriophage gp45. This result shows that the three-dimensional structure of the sliding clamp is conserved in the three domains of life. pfuPCNA has two remarkable features compared with the human and yeast PCNA molecules: it has more ion pairs and fewer intermolecular main chain hydrogen bonds. The former may contribute to the thermal stability of pfuPCNA, and the latter may be the cause of the stimulatory effect of pfuPCNA on the DNA synthesizing activity of P. furiosus DNA polymerases in the absence of the clamp loader replication factor C in vitro. PMID- 11266592 TI - Differentiation of the slow-binding mechanism for magnesium ion activation and zinc ion inhibition of human placental alkaline phosphatase. AB - The binding mechanism of Mg(2+) at the M3 site of human placental alkaline phosphatase was found to be a slow-binding process with a low binding affinity (K(Mg(app.)) = 3.32 mM). Quenching of the intrinsic fluorescence of the Mg(2+) free and Mg(2+)-containing enzymes by acrylamide showed almost identical dynamic quenching constant (K(sv) = 4.44 +/- 0.09 M(-1)), indicating that there is no gross conformational difference between the M3-free and the M3-Mg(2+) enzymes. However, Zn(2+) was found to have a high affinity with the M3 site (K(Zn(app.)) = 0.11 mM) and was observed as a time-dependent inhibitor of the enzyme. The dependence of the observed transition rate from higher activity to lower activity (k(obs)) at different zinc concentrations resulted in a hyperbolic curve suggesting that zinc ion induces a slow conformational change of the enzyme, which locks the enzyme in a conformation (M3'-Zn) having an extremely high affinity for the Zn(2+) (K*(Zn(app.)) = 0.33 microM). The conformation of the M3' Zn enzyme, however, is unfavorable for the catalysis by the enzyme. Both Mg(2+) activation and Zn(2+) inhibition of the enzyme are reversible processes. Structural information indicates that the M3 site, which is octahedrally coordinated to Mg(2+), has been converted to a distorted tetrahedral coordination when zinc ion substitutes for magnesium ion at the M3 site. This conformation of the enzyme has a small dynamic quenching constant for acrylamide (K(sv) = 3.86 +/ 0.04 M(-1)), suggesting a conformational change. Both Mg(2+) and phosphate prevent the enzyme from reaching this inactive structure. GTP plays an important role in reactivating the Zn-inhibited enzyme activity. We propose that, under physiological conditions, magnesium ion may play an important modulatory role in the cell for protecting the enzyme by retaining a favorable geometry of the active site needed for catalysis. PMID- 11266591 TI - An engineered leucine zipper a position mutant with an unusual three-state unfolding pathway. AB - The leucine zipper is a dimeric coiled-coil structural motif consisting of four to six heptad repeats, designated (abcdefg)(n). In the GCN4 leucine zipper, a position 16 in the third heptad is occupied by an Asn residue whereas the other a positions are Val residues. Recently, we have constructed variants of the GCN4 leucine zipper in which the a position Val residues were replaced by Ile. The folding and unfolding of the wild-type GCN4 leucine zipper and the Val to Ile variant both adhere to a simple two-state mechanism. In this study, another variant of the GCN4 leucine zipper was constructed by moving the single Asn residue from a position 16 to a position 9. This switch causes the thermal unfolding of the GCN4 leucine zipper to become three state. The unfolding pathway of this variant was determined by thermal denaturation, limited proteinase K digestion, and sedimentation equilibrium analysis. Our data are consistent with a model in which the variant first unfolds from its N terminus and changes the oligomerization specificity from a native dimer to a partially unfolded intermediate containing a mixture of dimers and trimers and then completely unfolds to unstructured monomers. PMID- 11266593 TI - Stabilizing interactions in the dimer interface of alpha-subunit in Escherichia coli RNA polymerase: a graph spectral and point mutation study. AB - The formation of alpha(2) dimer in Escherichia coli core RNA polymerase (RNAP) is thought to be the first step toward the assembly of the functional enzyme. A large number of evidences indicate that the alpha-subunit dimerizes through its N terminal domain (NTD). The crystal structures of the alpha-subunit NTD and that of a homologous Thermus aquaticus core RNAP are known. To identify the stabilizing interactions in the dimer interface of the alpha-NTD of E. coli RNAP, we identified side-chain clusters by using the crystal structure coordinates of E. coli alpha-NTD. A graph spectral algorithm was used to identify side-chain clusters. This algorithm considers the global nonbonded side-chain interactions of the residues for the clustering procedure and is unique in identifying residues that make the largest number of interactions among the residues that form clusters in a very quantitative way. By using this algorithm, a nine-residue cluster consisting of polar and hydrophobic residues was identified in the subunit interface adjacent to the hydrophobic core. The residues forming the cluster are relatively rigid regions of the interface, as measured by the thermal factors of the residues. Most of the cluster residues in the E. coli enzyme were topologically and sequentially conserved in the T. aquaticus RNAP crystal structure. Residues 35F and 46I were predicted to be important in the stability of the alpha-dimer interface, with 35F forming the center of the cluster. The predictions were tested by isolating single-point mutants alpha-F35A and alpha I46S on the dimer interface, which were found to disrupt dimerization. Thus, the identified cluster at the edge of the dimer interface seems to be a vital component in stabilizing the alpha-NTD. PMID- 11266594 TI - A model of dynamic side-chain--side-chain interactions in the alpha-lactalbumin molten globule. AB - Proteins in the molten globule state contain high levels of secondary structure, as well as a rudimentary, nativelike tertiary topology. Thus, the structural similarity between the molten globule and native proteins may have a significant bearing in understanding the protein-folding problem. To explore the nature of side-chain--side-chain interactions in the alpha-lactalbumin (alpha-LA) molten globule, we determined the effective concentration for formation of the 28--111 disulfide bond in 14 double-mutant proteins, each containing two hydrophobic core residues replaced by alanine. We compared our results with those of single alanine substitutions using the framework of double-mutant cycle analysis and found that, in the majority of cases, the effects of two alanine substitutions are additive. Based on these results, we propose a model of side-chain-side-chain interactions in the alpha-LA molten globule, which takes into consideration the dynamic nature of this partially folded species. PMID- 11266595 TI - The effect of substrate binding on the conformation and structural stability of Herpes simplex virus type 1 thymidine kinase. AB - The structure of Herpes simplex virus type 1 thymidine kinase (TK(HSV1)) is known at high resolution in complex with a series of ligands and exhibits important structural similarities to the nucleoside monophosphate (NMP) kinase family, which are known to show large conformational changes upon binding of substrates. The effect of substrate binding on the conformation and structural stability of TK(HSV1), measured by thermal denaturation experiments, far-UV circular dichroism (CD) and fluorescence is described, and the results indicate that the conformation of the ligand-free TK(HSV1) is less ordered and less stable compared to the ligated enzyme. Furthermore, two crystal structures of TK(HSV1) in complex with two new ligands, HPT and HMTT, refined to 2.2 A are presented. Although TK(HSV1):HPT does not exhibit any significant deviations from the model of TK(HSV1):dT, the TK(HSV1):HMTT complex displays a unique conformationally altered active site resulting in a lowered thermal stability of this complex. Moreover, we show that binding affinity and binding mode of the ligand correlate with thermal stability of the complex. We use this correlation to propose a method to estimate binding constants for new TK(HSV1)substrates using thermal denaturation measurements monitored by CD spectroscopy. The kinetic and structural results of both test substrates HPT and HMTT show that the CD thermal denaturation system is very sensitive to conformational changes caused by unusual binding of a substrate analog. PMID- 11266596 TI - Characterization of apo and partially saturated states of calerythrin, an EF-hand protein from S. erythraea: a molten globule when deprived of Ca(2+). AB - Calerythrin, a four-EF-hand calcium-binding protein from Saccharopolyspora erythraea, exists in an equilibrium between ordered and less ordered states with slow exchange kinetics when deprived of Ca(2+) and at low temperatures, as observed by NMR. As the temperature is raised, signal dispersion in NMR spectra reduces, and intensity of near-UV CD bands decreases. Yet far-UV CD spectra indicate only a small decrease in the amount of secondary structure, and SAXS data show that no significant change occurs in the overall size and shape of the protein. Thus, at elevated temperatures, the equilibrium is shifted toward a state with characteristics of a molten globule. The fully structured state is reached by Ca(2+)-titration. Calcium first binds cooperatively to the C-terminal sites 3 and 4 and then to the N-terminal site 1, which is paired with an atypical, nonbinding site 2. EF-hand 2 still folds together with the C-terminal half of the protein, as deduced from the order of appearance of backbone amide cross peaks in the NMR spectra of partially Ca(2+)-saturated states. PMID- 11266597 TI - Solution structure and backbone dynamics of the DNA-binding domain of mouse Sox 5. AB - The fold of the murine Sox-5 (mSox-5) HMG box in free solution has been determined by multidimensional NMR using (15)N-labeled protein and has been found to adopt the characteristic twisted L-shape made up of two wings: the major wing comprising helix 1 (F10--F25) and helix 2 (N32--A43), the minor wing comprising helix 3 (P51--Y67) in weak antiparallel association with the N-terminal extended segment. (15)N relaxation measurements show considerable mobility (reduced order parameter, S(2)) in the minor wing that increases toward the amino and carboxy termini of the chain. The mobility of residues C-terminal to Q62 is significantly greater than the equivalent residues of non-sequence-specific boxes, and these residues show a weaker association with the extended N-terminal segment than in non-sequence boxes. Comparison with previously determined structures of HMG boxes both in free solution and complexed with DNA shows close similarity in the packing of the hydrophobic cores and the relative disposition of the three helices. Only in hSRY/DNA does the arrangement of aromatic sidechains differ significantly from that of mSox-5, and only in rHMG1 box 1 bound to cisplatinated DNA does helix 1 have no kink. Helix 3 in mSox-5 is terminated by P68, a conserved residue in DNA sequence-specific HMG boxes, which results in the chain turning through approximately 90 degrees. PMID- 11266598 TI - Differential chemical labeling of the AlcR DNA-binding domain from Aspergillas nidulans versus its complex with a 16-mer DNA target: identification of an essential tryptophan involved in the recognition and the interaction with the nucleic acid. AB - DNA binding of the ethanol regulon transcription factor AlcR from Aspergillus nidulans was shown to involve a consensus basic region as in the other zinc cluster proteins. However, additional interactions between some residues and DNA were suspected, among which were a hypothetic hydrophobic interaction between Trp45 and the T residue of the consensus TGCGG sequence. In the present study, the differential chemical labeling of both the free protein and the protein/DNA complex showed significantly different behaviors of the three tryptophan residues comprised in the AlcR sequence toward the Koshland reagent. The spectacular decreased reaction rate for Trp45 within the complex confirmed the location of this residue at the protein/DNA interface. A similar result obtained with Trp53, an amino acid present at the C-terminal side of AlcR, also indicated its involvement in the DNA recognition. In contrast, the formation of the complex accompanied by an allosteric rearrangement allowed the Trp36 to be much more exposed to the solvent than in the free protein. These data provide additional evidence that the unique specificity of AlcR among the zinc binuclear cluster family results in new types of interactions between AlcR and its cognate targets. From a methodological point of view, the approach of differential chemical labeling combined with mass spectrometric analyses proved to be an interesting tool for the recognition of hydrophobic interactions between the tryptophan residues of a protein and its macromolecular target. PMID- 11266599 TI - Structure of soybean seed coat peroxidase: a plant peroxidase with unusual stability and haem-apoprotein interactions. AB - Soybean seed coat peroxidase (SBP) is a peroxidase with extraordinary stability and catalytic properties. It belongs to the family of class III plant peroxidases that can oxidize a wide variety of organic and inorganic substrates using hydrogen peroxide. Because the plant enzyme is a heterogeneous glycoprotein, SBP was produced recombinant in Escherichia coli for the present crystallographic study. The three-dimensional structure of SBP shows a bound tris(hydroxymethyl)aminomethane molecule (TRIS). This TRIS molecule has hydrogen bonds to active site residues corresponding to the residues that interact with the small phenolic substrate ferulic acid in the horseradish peroxidase C (HRPC):ferulic acid complex. TRIS is positioned in what has been described as a secondary substrate-binding site in HRPC, and the structure of the SBP:TRIS complex indicates that this secondary substrate-binding site could be of functional importance. SBP has one of the most solvent accessible delta-meso haem edge (the site of electron transfer from reducing substrates to the enzymatic intermediates compound I and II) so far described for a plant peroxidase and structural alignment suggests that the volume of Ile74 is a factor that influences the solvent accessibility of this important site. A contact between haem C8 vinyl and the sulphur atom of Met37 is observed in the SBP structure. This interaction might affect the stability of the haem group by stabilisation/delocalisation of the porphyrin pi-cation of compound I. PMID- 11266600 TI - Testing the role of chain connectivity on the stability and structure of dihydrofolate reductase from E. coli: fragment complementation and circular permutation reveal stable, alternatively folded forms. AB - The effects of chain cleavage and circular permutation on the structure, stability, and activity of dihydrofolate reductase (DHFR) from Escherichia coli were investigated by various spectroscopic and biochemical methods. Cleavage of the backbone after position 86 resulted in two fragments, (1--86) and (87--159) each of which are poorly structured and enzymatically inactive. When combined in a 1 : 1 molar ratio, however, the fragments formed a high-affinity (K(a) = 2.6 x 10(7) M(-1)) complex that displays a weakly cooperative urea-induced unfolding transition at micromolar concentrations. The retention of about 15% of the enzymatic activity of full-length DHFR is surprising, considering that the secondary structure in the complex is substantially reduced from its wild-type counterpart. In contrast, a circularly permuted form with its N-terminus at position 86 has similar overall stability to full-length DHFR, about 50% of its activity, substantial secondary structure, altered side-chain packing in the adenosine binding domain, and unfolds via an equilibrium intermediate not observed in the wild-type protein. After addition of ligand or the tight-binding inhibitor methotrexate, both the fragment complex and the circular permutant adopt more native-like secondary and tertiary structures. These results show that changes in the backbone connectivity can produce alternatively folded forms and highlight the importance of protein-ligand interactions in stabilizing the active site architecture of DHFR. PMID- 11266601 TI - A "loop entropy reduction" phage-display selection for folded amino acid sequences. AB - As a step toward selecting folded proteins from libraries of randomized sequences, we have designed a 'loop entropy reduction'-based phage-display method. The basic premise is that insertion of a long disordered sequence into a loop of a host protein will substantially destabilize the host because of the entropic cost of closing a loop in a disordered chain. If the inserted sequence spontaneously folds into a stable structure with the N and C termini close in space, however, this entropic cost is diminished. The host protein function can, therefore, be used to select folded inserted sequences without relying on specific properties of the inserted sequence. This principle is tested using the IgG binding domain of protein L and the lck SH2 domain as host proteins. The results indicate that the loop entropy reduction screen is capable of discriminating folded from unfolded sequences when the proper host protein and insertion point are chosen. PMID- 11266602 TI - Molecular dynamics simulation of Escherichia coli dihydrofolate reductase and its protein fragments: relative stabilities in experiment and simulations. AB - We have carried out molecular dynamics simulations of the native dihydrofolate reductase from Escherichia coli and several of its folded protein fragments at standard temperature. The simulations have shown fragments 1--36, 37--88, and 89- 159 to be unstable, with a C(alpha)RMSD (C(alpha) root mean squared deviation) >5 A after 3.0 nsec of simulation. The unfolding of fragment 1--36 was immediate, whereas fragments 37--88 and 89--159 gradually unfolded because of the presence of the beta-sheet core structure. In the absence of residues 1--36, the two distinct domains comprising fragment 39--159 associated with each other, resulting in a stable conformation. This conformation retained most of its native structural elements. We have further simulated fragments derived from computational protein cutting. These were also found to be unstable, with the exception of fragment 104--159. In the absence of alpha(4), the loose loop region of residues 120--127 exhibited a beta-strand-like behavior, associating itself with the beta-sheet core of the protein fragment. The current study suggests that the folding of dihydrofolate reductase involves cooperative folding of distinct domains which otherwise would have been unstable as independent folded units in solution. Finally, the critical role of residues 1--36 in allowing the two distinct domains of fragment 104--159 to fold into the final native conformation is discussed. PMID- 11266604 TI - Understanding thermostability in cytochrome P450 by combinatorial mutagenesis. AB - The cytochromes P450 are an important class of mono-oxygenases involved in xenobiotic metabolism and steroid biosynthesis in a diverse set of life forms. Discovery of CYP-119, a P450 from the archea Sulfolobus solfataricus has provided a means for understanding nature's method of stabilizing this important protein superfamily. To identify classes of stabilizing interactions used by CYP-119, we have generated a randomized library of point mutants and screened for mutants that are less thermostable than the wild type by monitoring the characteristic Soret band in the visible region of the cell lysis. The selected mutants were characterized by differential scanning calorimetry to compare the temperatures of the melting transitions of the various mutants. The identified mutations suggested that electrostatic interactions involving salt links and charge-charge interactions, as well as contributions from other interactions such as aromatic stacking, and side chain volume of hydrophobic residues contribute to enhanced thermostability in this cytochrome P450. PMID- 11266603 TI - Disulfide bond effects on protein stability: designed variants of Cucurbita maxima trypsin inhibitor-V. AB - Attempts to increase protein stability by insertion of novel disulfide bonds have not always been successful. According to the two current models, cross-links enhance stability mainly through denatured state effects. We have investigated the effects of removal and addition of disulfide cross-links, protein flexibility in the vicinity of a cross-link, and disulfide loop size on the stability of Cucurbita maxima trypsin inhibitor-V (CMTI-V; 7 kD) by differential scanning calorimetry. CMTI-V offers the advantage of a large, flexible, and solvent exposed loop not involved in extensive intra-molecular interactions. We have uncovered a negative correlation between retention time in hydrophobic column chromatography, a measure of protein hydrophobicity, and melting temperature (T(m)), an indicator of native state stabilization, for CMTI-V and its variants. In conjunction with the complete set of thermodynamic parameters of denaturation, this has led to the following deductions: (1) In the less stable, disulfide removed C3S/C48S (Delta Delta G(d)(50 degrees C) = -4 kcal/mole; Delta T(m) = -22 degrees C), the native state is destabilized more than the denatured state; this also applies to the less-stable CMTI-V* (Delta Delta G(d)(50 degrees C) = -3 kcal/mole; Delta T(m) = -11 degrees C), in which the disulfide-containing loop is opened by specific hydrolysis of the Lys(44)-Asp(45) peptide bond; (2) In the less stable, disulfide-inserted E38C/W54C (Delta Delta G(d)(50 degrees C) = -1 kcal/mole; Delta T(m) = +2 degrees C), the denatured state is more stabilized than the native state; and (3) In the more stable, disulfide-engineered V42C/R52C (Delta Delta G(d)(50 degrees C) = +1 kcal/mole; Delta T(m) = +17 degrees C), the native state is more stabilized than the denatured state. These results show that a cross-link stabilizes both native and denatured states, and differential stabilization of the two states causes either loss or gain in protein stability. Removal of hydrogen bonds in the same flexible region of CMTI-V resulted in less destabilization despite larger changes in the enthalpy and entropy of denaturation. The effect of a cross-link on the denatured state of CMTI-V was estimated directly by means of a four-state thermodynamic cycle consisting of native and denatured states of CMTI-V and CMTI-V*. Overall, the results show that an enthalpy-entropy compensation accompanies disulfide bond effects and protein stabilization is profoundly modulated by altered hydrophobicity of both native and denatured states, altered flexibility near the cross-link, and residual structure in the denatured state. PMID- 11266605 TI - A novel target recognition revealed by calmodulin in complex with the basic helix -loop--helix transcription factor SEF2-1/E2-2. AB - Calmodulin is the predominant intracellular receptor for Ca(2+) signals, mediating the regulation of numerous cellular processes. It can inhibit the DNA binding of basic helix--loop--helix transcription factors by a direct interaction of a novel type. To structurally characterize this novel calmodulin-target interaction, we decided to study the complex of calmodulin with a dimeric peptide corresponding to the DNA-binding domains of the dimeric basic helix-loop-helix transcription factor SEF2-1 (SEF2-1mp) using NMR. Here, we report that the stoichiometry of the calmodulin:SEF2-1mp complex is one dimeric peptide binding two calmodulin molecules. We also report the 1H, 13C, and 15N resonance assignments and the secondary structure of calmodulin in this for NMR large (approximately 38 kD) complex, as well as the 1H assignments and secondary structure of SEF2-1mp. In addition, we determined the amide proton exchange rates of calmodulin and measured intermolecular calmodulin:SEF2-1mp and calmodulin:calmodulin NOE contacts. The isotope-filtered experiments show a large number of SEF2-1mp to calmodulin NOE contacts indicating that a tight complex is formed, which is confirmed by an intermolecular calmodulin:calmodulin NOE contact. The secondary structure and amide proton exchange data show that the binding does not occur via the classical wraparound binding mode. Instead, the data indicate that calmodulin interacts with SEF2-1mp in a more open conformation, although the hydrophobic surfaces of the N- and C-terminal domains still form the main interaction sites. Interactions involving charged residues are also identified in agreement with the known relatively high sensitivity of the binding to ionic strength. Finally, the peptide does not form an alpha-helix as in the classical wraparound binding mode. PMID- 11266606 TI - Amyloid fibrils derived from the apolipoprotein A1 Leu174Ser variant contain elements of ordered helical structure. AB - We recently described a new apolipoprotein A1 variant presenting a Leu174Ser replacement mutation that is associated with a familial form of systemic amyloidosis displaying predominant heart involvement. We have now identified a second unrelated patient with very similar clinical presentation and carrying the identical apolipoprotein A1 mutation. In this new patient the main protein constituent of the amyloid fibrils is the polypeptide derived from the first 93 residues of the protein, the identical fragment to that found in the patient previously described to carry this mutation. The X-ray fiber diffraction pattern obtained from preparations of partially aligned fibrils displays the cross-beta reflections characteristic of all amyloid fibrils. In addition to these cross beta reflections, other reflections suggest the presence of well-defined coiled coil helical structure arranged with a defined orientation within the fibrils. In both cases the fibrils contain a trace amount of full-length apolipoprotein A1 with an apparent prevalence of the wild-type species over the variant protein. We have found a ratio of full-length wild-type to mutant protein in plasma HDL of three to one. The polypeptide 1--93 purified from natural fibrils can be solubilized in aqueous solutions containing denaturants, and after removal of denaturants it acquires a monomeric state that, based on CD and NMR studies, has a predominantly random coil structure. The addition of phospholipids to the monomeric form induces the formation of some helical structure, thought most likely to occur at the C-terminal end of the polypeptide. PMID- 11266608 TI - TM Finder: a prediction program for transmembrane protein segments using a combination of hydrophobicity and nonpolar phase helicity scales. AB - Based on the principle of dual prediction by segment hydrophobicity and nonpolar phase helicity, in concert with imposed threshold values of these two parameters, we developed the automated prediction program TM Finder that can successfully locate most transmembrane (TM) segments in proteins. The program uses the results of experiments on a series of host-guest TM segment mimic peptides of prototypic sequence KK AAAXAAAAAXAAWAAXAAAKKKK-amide (where X = each of the 20 commonly occurring amino acids) through which an HPLC-derived hydropathy scale, a hydrophobicity threshold for spontaneous membrane insertion, and a nonpolar phase helical propensity scale were determined. Using these scales, the optimized prediction algorithm of TM Finder defines TM segments by first searching for competent core segments using the combination of hydrophobicity and helicity scales, and then performs a gap-joining operation, which minimizes prediction bias caused by local hydrophilic residues and/or the choice of window size. In addition, the hydrophobicity threshold requirement enables TM Finder to distinguish reliably between membrane proteins and globular proteins, thereby adding an important dimension to the program. A full web version of the TM Finder program can be accessed at http://www.bioinformatics-canada.org/TM/. PMID- 11266607 TI - Application of photoaffinity labeling with [(3)H] all trans- and 9-cis-retinoic acids for characterization of cellular retinoic acid--binding proteins I and II. AB - Cellular retinoic acid-binding proteins (CRABPs) are carrier proteins thought to play a crucial role in the transport and metabolism of all-trans-retinoic acid (atRA) and its derivatives within the cell. This report describes a novel photoaffinity-based binding assay involving competition between potential ligands of CRABP and [(3)H]atRA or [(3)H]-9-cis-RA for binding to the atRA-binding sites of CRABP I and II. Photoaffinity labeling of purified CRABPs with [(3)H]atRA was light- and concentration-dependent, saturable, and protected by several retinoids in a concentration-dependent manner, indicating that binding occurred in the CRABP atRA-binding site. Structure-function relationship studies demonstrated that oxidative changes to the atRA beta-ionone ring did not affect ligand potency. However, derivatives lacking a terminal carboxyl group and some cis isomers did not bind to CRABPs. These studies also identified two novel ligands for CRABPs: 5,6-epoxy-RA and retinoyl-beta-D-glucuronide (RAG). The labeling of both CRABPs with 9-cis-RA occurred with much lower affinity. Experimental evidence excluded nonspecific binding of RAG to CRABPs and UDP glucuronosyltransferases, the enzymes responsible for RAG synthesis. These results established that RAG is an effective ligand of CRABPs. Therefore, photoaffinity labeling with [(3)H]atRA can be used to identify new ligands for CRABP and retinoid nuclear receptors and also provide information concerning the identity of amino acid(s) localized in the atRA-binding site of these proteins. PMID- 11266609 TI - Threading of a glycosylated protein loop through a protein hole: implications for combination of human chorionic gonadotropin subunits. AB - Chorionic gonadotropin (hCG) is a heterodimeric placental glycoprotein hormone essential for human reproduction. Twenty hCG beta-subunit residues, termed the seatbelt, are wrapped around alpha-subunit loop 2 (alpha 2) and their positions "latched" by a disulfide formed by cysteines at the end of the seatbelt (Cys 110) and in the beta-subunit core (Cys 26). This unique arrangement explains the stability of the heterodimer but raises questions as to how the two subunits combine. The seatbelt is latched in the free beta-subunit. If the seatbelt remained latched during the process of subunit combination, formation of the heterodimer would require alpha 2 and its attached oligosaccharide to be threaded through a small beta-subunit hole. The subunits are known to combine during oxidizing conditions in vitro, and studies described here tested the idea that this requires transient disruption of the latch disulfide, possibly as a consequence of the thioredoxin activity reported in hCG. We observed that alkylating agents did not modify either cysteine in the latch disulfide (Cys 26 or Cys 110) during heterodimer formation in several oxidizing conditions and had minimal influence on these cysteines during combination in the presence of mild reductants (1--3 mM beta-mercaptoethanol). Reducing agents appeared to accelerate subunit combination by disrupting a disulfide (Cys 93--Cys 100) that forms a loop within the seatbelt, thereby increasing the size of the beta-subunit hole. We propose a mechanism for hCG assembly in vitro that depends on movements of alpha 2 and the seatbelt and suggest that the process of glycoprotein hormone subunit combination may be useful for studying the movements of loops during protein folding. PMID- 11266610 TI - Design, synthesis, and characterization of a novel hemoprotein. AB - Here we describe a synthetic protein (6H7H) designed to bind four heme groups via bis-histidine axial ligation. The hemes are designed to bind perpendicular to another in an orientation that mimics the relative geometry of the two heme a groups in the active site of cytochrome c oxidase. Our newly developed protein design program, called CORE, was implemented in the design of this novel hemoprotein. Heme titration studies resolved four distinct K(D) values (K(D1) = 80 nM, K(D2) = 18 nM, K(D3) > or = 3 mM, K(D4) < or = 570 nM, with K(D3) x K(D4) = 1700); positive cooperativity in binding between the first and second heme, as well as substantial positive cooperativity between the third and forth heme, was observed. Chemical and thermal denaturation studies reveal a stable protein with native-like properties. Visible circular dichroism spectroscopy of holo-6H7H indicates excitonic coupling between heme groups. Further electrochemical and spectroscopic characterization of the holo-protein support a structure that is consistent with the predefined target structure. PMID- 11266611 TI - Molecular confinement influences protein structure and enhances thermal protein stability. AB - The sol-gel method of encapsulating proteins in a silica matrix was investigated as a potential experimental system for testing the effects of molecular confinement on the structure and stability of proteins. We demonstrate that silica entrapment (1) is fully compatible with structure analysis by circular dichroism, (2) allows conformational studies in contact with solvents that would otherwise promote aggregation in solution, and (3) generally enhances thermal protein stability. Lysozyme, alpha-lactalbumin, and metmyoglobin retained native like solution structures following sol-gel encapsulation, but apomyoglobin was found to be largely unfolded within the silica matrix under control buffer conditions. The secondary structure of encapsulated apomyoglobin was unaltered by changes in pH and ionic strength of KCl. Intriguingly, the addition of other neutral salts resulted in an increase in the alpha-helical content of encapsulated apomyoglobin in accordance with the Hofmeister ion series. We hypothesize that protein conformation is influenced directly by the properties of confined water in the pores of the silica. Further work is needed to differentiate the steric effects of the silica matrix from the solvent effects of confined water on protein structure and to determine the extent to which this experimental system mimics the effects of crowding and confinement on the function of macromolecules in vivo. PMID- 11266612 TI - Plasticity of quaternary structure: twenty-two ways to form a LacI dimer. AB - The repressor proteins of the LacI/GalR family exhibit significant similarity in their secondary and tertiary structures despite less than 35% identity in their primary sequences. Furthermore, the core domains of these oligomeric repressors, which mediate dimerization, are homologous with the monomeric periplasmic binding proteins, extending the issue of plasticity to quaternary structure. To elucidate the determinants of assembly, a structure-based alignment has been created for three repressors and four periplasmic binding proteins. Contact maps have also been constructed for the three repressor interfaces to distinguish any conserved interactions. These analyses show few strict requirements for assembly of the core N-subdomain interface. The interfaces of repressor core C-subdomains are well conserved at the structural level, and their primary sequences differ significantly from the monomeric periplasmic binding proteins at positions equivalent to LacI 281 and 282. However, previous biochemical and phenotypic analyses indicate that LacI tolerates many mutations at 281. Mutations at LacI 282 were shown to abrogate assembly, but for Y282D this could be compensated by a second-site mutation in the core N-subdomain at K84 to L or A. Using the link between LacI assembly and function, we have further identified 22 second-site mutations that compensate the Y282D dimerization defect in vivo. The sites of these mutations fall into several structural regions, each of which may influence assembly by a different mechanism. Thus, the 360-amino acid scaffold of LacI allows plasticity of its quaternary structure. The periplasmic binding proteins may require only minimal changes to facilitate oligomerization similar to the repressor proteins. PMID- 11266613 TI - Ulex europaeus agglutinin II (UEA-II) is a novel, potent inhibitor of complement activation. AB - Complement is an important mediator of vascular injury following oxidative stress. We recently demonstrated that complement activation following endothelial oxidative stress is mediated by mannose-binding lectin (MBL) and activation of the lectin complement pathway. Here, we investigated whether nine plant lectins which have a binding profile similar to that of MBL competitively inhibit MBL deposition and subsequent complement activation following human umbilical vein endothelial cell (HUVEC) oxidative stress. HUVEC oxidative stress (1% O(2), 24 hr) significantly increased Ulex europaeus agglutinin II (UEA-II) binding by 72 +/- 9% compared to normoxic cells. UEA-II inhibited MBL binding to HUVEC in a concentration-dependent manner following oxidative stress. Further, MBL inhibited UEA-II binding to HUVEC in a concentration-dependent manner following oxidative stress, suggesting a common ligand. UEA-II (< or = 100 micromol/L) did not attenuate the hemolytic activity, nor did it inhibit C3a des Arg formation from alternative or classical complement pathway-specific hemolytic assays. C3 deposition (measured by ELISA) following HUVEC oxidative stress was inhibited by UEA-II in a concentration-dependent manner (IC(50) = 10 pmol/L). UEA-II inhibited C3 and MBL co-localization (confocal microscopy) in a concentration-dependent manner on HUVEC following oxidative stress (IC(50) approximately 1 pmol/L). Finally, UEA-II significantly inhibited complement-dependent neutrophil chemotaxis, but failed to inhibit fMLP-mediated chemotaxis, following endothelial oxidative stress. These data demonstrate that UEA-II is a novel, potent inhibitor of human MBL deposition and complement activation following human endothelial oxidative stress. PMID- 11266614 TI - Sialidase-like Asp-boxes: sequence-similar structures within different protein folds. AB - Sequence similarity is the most common measure currently used to infer homology between proteins. Typically, homologous protein domains show sequence similarity over their entire lengths. Here we identify Asp box motifs, initially found as repeats in sialidases and neuraminidases, in new structural and sequence contexts. These motifs represent significantly similar sequences, localized to beta hairpins within proteins that are otherwise different in sequence and three dimensional structure. By performing a combined sequence- and structure-based analysis we detect Asp boxes in more than nine protein families, including bacterial ribonucleases, sulfite oxidases, reelin, netrins, some lipoprotein receptors, and a variety of glycosyl hydrolases. Although the function common to each of these proteins, if any, remains unclear, we discuss possible functions of Asp boxes on the basis of previously determined experimental results and discuss different evolutionary scenarios for the origin of Asp-box containing proteins. PMID- 11266615 TI - A noncanonical WD-repeat protein from the cyanobacterium Synechocystis PCC6803: structural and functional study. AB - SYNECHOCYSTIS: PCC6803 possesses several open reading frames encoding putative WD repeat proteins. One, the Hat protein, is involved in the control of a high affinity transport system for inorganic carbon that is active when the cells are grown under a limiting concentration of this carbon substrate. The protein is composed of two major domains separated by a hydrophobic linker region of 20 amino acid residues. The N-terminal domain of Hat has no homolog in standard databases and does not display any particular structural features. Eleven WD repeats have been identified in the C-terminal moiety. The region encompassing the four terminal WD repeats is essential for growth under a limiting inorganic carbon regime. The region encompassing the two most terminal WD repeats is required for the activity of the high-affinity transport system. However, because the Hat protein is located in the thylakoids, it should not be itself an element of the transport system. The structural organization of the WD-containing domain of Hat was modeled from the crystal structure of the G protein beta subunit (with seven WD repeats) and of hemopexin (a structural analog with four blades). Functional and structural data argue in favor of an organization of the Hat WD moiety in two subdomains of seven and four WD repeats. The C-terminal 4-mer subdomain might interact with another, yet unknown, protein/peptide. This interaction could be essential in modulating the stability of the 4-mer structure and, thus, the accessibility of this subdomain, or at least of the region encompassing the last two WD repeats. PMID- 11266616 TI - Solvation energetics and conformational change in EF-hand proteins. AB - Calmodulin and other members of the EF-hand protein family are known to undergo major changes in conformation upon binding Ca(2+). However, some EF-hand proteins, such as calbindin D9k, bind Ca(2+) without a significant change in conformation. Here, we show the importance of a precise balance of solvation energetics to conformational change, using mutational analysis of partially buried polar groups in the N-terminal domain of calmodulin (N-cam). Several variants were characterized using fluorescence, circular dichroism, and NMR spectroscopy. Strikingly, the replacement of polar side chains glutamine and lysine at positions 41 and 75 with nonpolar side chains leads to dramatic enhancement of the stability of the Ca(2+)-free state, a corresponding decrease in Ca(2+)-binding affinity, and an apparent loss of ability to change conformation to the open form. The results suggest a paradigm for conformational change in which energetic strain is accumulated in one state in order to modulate the energetics of change to the alternative state. PMID- 11266618 TI - Two-phase unfolding pathway of ribonuclease A during denaturation induced by dithiothreitol. AB - The dynamics of the unfolding process of bovine pancreatic ribonuclease A (RNase A) unfolded by dithiothreitol (DTT) at a low concentration of 1:30 were investigated in alkaline phosphate-buffered saline solutions at 303K and 313K by using proton nuclear magnetic resonance ((1)H NMR) spectra. Three NMR spectral parameters including Shannon entropy, mutual information, and correlation coefficient were introduced into the analysis. The results show that the unfolding process of RNase A was slowed to the order of many hours, and the kinetics of the unfolding pathway described by the three parameters is best fit by a model of two consecutive first-order reactions. Temperature greatly influences the rate constants of the unfolding kinetics with different temperature effects observed for the fast and the slow processes. The consistencies and the differences between the three sets of parameters show that they reflect the same relative denaturation pathway but different spectra windows of the unfolding process of RNase A. The results suggest that the unfolding process of RNase A induced by low concentrations of DTT is a two-phase pathway containing fast and slow first-order reactions. PMID- 11266617 TI - Stabilization of hen egg white lysozyme by a cavity-filling mutation. AB - Stabilization of a protein using cavity-filling strategy has hardly been successful because of unfavorable van der Waals contacts. We succeeded in stabilizing lysozymes by cavity-filling mutations. The mutations were checked by a simple energy minimization in advance. It was shown clearly that the sum of free energy change caused by the hydrophobicity and the cavity size was correlated very well with protein stability. We also considered the aromatic aromatic interaction. It is reconfirmed that the cavity-filling mutation in a hydrophobic core is a very useful method to stabilize a protein when the mutation candidate is selected carefully. PMID- 11266619 TI - Enantioselectivity in Candida antarctica lipase B: a molecular dynamics study. AB - A major problem in predicting the enantioselectivity of an enzyme toward substrate molecules is that even high selectivity toward one substrate enantiomer over the other corresponds to a very small difference in free energy. However, total free energies in enzyme-substrate systems are very large and fluctuate significantly because of general protein motion. Candida antarctica lipase B (CALB), a serine hydrolase, displays enantioselectivity toward secondary alcohols. Here, we present a modeling study where the aim has been to develop a molecular dynamics-based methodology for the prediction of enantioselectivity in CALB. The substrates modeled (seven in total) were 3-methyl-2-butanol with various aliphatic carboxylic acids and also 2-butanol, as well as 3,3-dimethyl-2 butanol with octanoic acid. The tetrahedral reaction intermediate was used as a model of the transition state. Investigative analyses were performed on ensembles of nonminimized structures and focused on the potential energies of a number of subsets within the modeled systems to determine which specific regions are important for the prediction of enantioselectivity. One category of subset was based on atoms that make up the core structural elements of the transition state. We considered that a more favorable energetic conformation of such a subset should relate to a greater likelihood for catalysis to occur, thus reflecting higher selectivity. The results of this study conveyed that the use of this type of subset was viable for the analysis of structural ensembles and yielded good predictions of enantioselectivity. PMID- 11266620 TI - Effects of i-propanol on the structural dynamics of Thermomyces lanuginosa lipase revealed by tryptophan fluorescence. AB - Influence of isopropanol (iPrOH) on the structural dynamics of Thermomyces lanuginosa lipase (TLL) was studied by steady-state, time-resolved, and stopped flow fluorescence spectroscopy, monitoring the intrinsic emission of Trp residues. The fluorescence of the four Trps of the wild-type enzyme report on the global changes of the whole lipase molecule. To monitor the conformational changes in the so-called "lid," an alpha-helical surface loop, the single Trp mutant W89m (W117F, W221H, W260H) was employed. Circular dichroism (CD) spectra revealed that iPrOH does not cause major alterations in the secondary structures of the wild-type TLL and W89m. With increasing [iPrOH], judged by the ratio of emission intensities at 350 nm and 330 nm, the average microenvironment of the Trps in the wild-type TLL became more hydrophobic, whereas Trp89 of W89m moved into a more hydrophilic microenvironment. Time-resolved fluorescence measurements revealed no major changes to be induced by iPrOH neither in the shorter fluorescence lifetime component (tau(1) = 0.5--1.2 ns) for the wild-type TLL nor in the longer fluorescence lifetime component (tau(2) = 4.8--6.0 ns) in the wild type TLL and the W89m mutant. Instead, for W89m on increasing iPrOH from 25% to 50% the value for tau(1) increased significantly, from 0.43 to 1.5 ns. The shorter correlation time phi(1) of W89m had a minimum of 0.08 ns in 25% iPrOH. Judged from the residual anisotropy r(infinity) the amplitude of the local motion of Trp89 increased upon increasing [iPrOH] 10%. Stopped-flow fluorescence spectroscopy measurements suggested the lid to open within approximately 2 ms upon transfer of W89m into 25% iPrOH. Steady-state anisotropies and longer correlation times revealed increasing concentrations of iPrOH to result also in the formation of dimers as well as possibly also higher oligomers by TLL. PMID- 11266621 TI - LiveBench-1: continuous benchmarking of protein structure prediction servers. AB - We present a novel, continuous approach aimed at the large-scale assessment of the performance of available fold-recognition servers. Six popular servers were investigated: PDB-Blast, FFAS, T98-lib, GenTHREADER, 3D-PSSM, and INBGU. The assessment was conducted using as prediction targets a large number of selected protein structures released from October 1999 to April 2000. A target was selected if its sequence showed no significant similarity to any of the proteins previously available in the structural database. Overall, the servers were able to produce structurally similar models for one-half of the targets, but significantly accurate sequence-structure alignments were produced for only one third of the targets. We further classified the targets into two sets: easy and hard. We found that all servers were able to find the correct answer for the vast majority of the easy targets if a structurally similar fold was present in the server's fold libraries. However, among the hard targets--where standard methods such as PSI-BLAST fail--the most sensitive fold-recognition servers were able to produce similar models for only 40% of the cases, half of which had a significantly accurate sequence-structure alignment. Among the hard targets, the presence of updated libraries appeared to be less critical for the ranking. An "ideally combined consensus" prediction, where the results of all servers are considered, would increase the percentage of correct assignments by 50%. Each server had a number of cases with a correct assignment, where the assignments of all the other servers were wrong. This emphasizes the benefits of considering more than one server in difficult prediction tasks. The LiveBench program (http://BioInfo.PL/LiveBench) is being continued, and all interested developers are cordially invited to join. PMID- 11266623 TI - Inactivation mechanism of the membrane protein diacylglycerol kinase in detergent solution. AB - We have examined the irreversible inactivation mechanism of the membrane protein diacylglycerol kinase in the detergents n-octyl-beta-D-glucopyranoside (OG) at 55 degrees C and n-decyl-maltopyranoside (DM) at 80 degrees C. Under no inactivation conditions did we find any direct evidence for the chemical modifications that are commonly found in soluble proteins. Moreover, protein inactivated at 55 degrees C in OG could be reactivated by an unfolding and refolding protocol, suggesting that the protein is inactivated by a stable conformational change, not a covalent modification. We also found that the inactivation rate decreased with both increasing protein concentration and increasing thermodynamic stability, consistent with an inactivation pathway involving transient dissociation and/or unfolding of the protein. Our results suggest that the primary cause of diacylglycerol kinase inactivation is not low solubility, but poor intrinsic stability in the detergent environment. PMID- 11266622 TI - Optimization of binding electrostatics: charge complementarity in the barnase barstar protein complex. AB - Theoretical and experimental studies have shown that the large desolvation penalty required for polar and charged groups frequently precludes their involvement in electrostatic interactions that contribute strongly to net stability in the folding or binding of proteins in aqueous solution near room temperature. We have previously developed a theoretical framework for computing optimized electrostatic interactions and illustrated use of the algorithm with simplified geometries. Given a receptor and model assumptions, the method computes the ligand-charge distribution that provides the most favorable balance of desolvation and interaction effects on binding. In this paper the method has been extended to treat complexes using actual molecular shapes. The barnase barstar protein complex was investigated with barnase treated as a target receptor. The atomic point charges of barstar were varied to optimize the electrostatic binding free energy. Barnase and natural barstar form a tight complex (K(d) approximately 10(-14) M) with many charged and polar groups near the interface that make this a particularly relevant system for investigating the role of electrostatic effects on binding. The results show that sets of barstar charges (resulting from optimization with different constraints) can be found that give rise to relatively large predicted improvements in electrostatic binding free energy. Principles for enhancing the effect of electrostatic interactions in molecular binding in aqueous environments are discussed in light of the optima. Our findings suggest that, in general, the enhancements in electrostatic binding free energy resulting from modification of polar and charged groups can be substantial. Moreover, a recently proposed definition of electrostatic complementarity is shown to be a useful tool for examining binding interfaces. Finally, calculational results suggest that wild-type barstar is closer to being affinity optimized than is barnase for their mutual binding, consistent with the known roles of these proteins. PMID- 11266624 TI - Solution nuclear magnetic resonance structure of a protein disulfide oxidoreductase from Methanococcus jannaschii. AB - The solution structure of the protein disulfide oxidoreductase Mj0307 in the reduced form has been solved by nuclear magnetic resonance. The secondary and tertiary structure of this protein from the archaebacterium Methanococcus jannaschii is similar to the structures that have been solved for the glutaredoxin proteins from Escherichia coli, although Mj0307 also shows features that are characteristic of thioredoxin proteins. Some aspects of Mj0307's unique behavior can be explained by comparing structure-based sequence alignments with mesophilic bacterial and eukaryotic glutaredoxin and thioredoxin proteins. It is proposed that Mj0307, and similar archaebacterial proteins, may be most closely related to the mesophilic bacterial NrdH proteins. Together these proteins may form a unique subgroup within the family of protein disulfide oxidoreductases. PMID- 11266626 TI - Effect of polymorphisms on ligand binding by mouse major urinary proteins. AB - Mouse urine contains an abundance of major urinary proteins, lipocalins, whose roles include slow release of semiochemicals. These proteins are highly polymorphic, with small sequence differences between individual members. In this study, we purified to homogeneity four of these proteins from two strains of inbred mice and characterized them by mass spectrometry. This analysis has led to the discovery of another variant in this group of proteins. Three of the polymorphic variants that map to the surface have no effect on the binding of a fluorescent probe in the binding cavity, but the fourth, characterized by a Phe to Val substitution in the cavity, shows a substantially lower affinity and fluorescence yield for the probe. These results are interpreted in light of the known crystal structure of the protein and molecular modeling calculations, which rationalize the experimental findings. This work raises the possibility that the calyx-binding site can show specificity for different ligands, the implications of which on pheromone binding and chemical communication are discussed. PMID- 11266625 TI - Role for cysteine residues in the in vivo folding and assembly of the phage P22 tailspike. AB - The predominantly beta-sheet phage P22 tailspike adhesin contains eight reduced cysteines per 666 residue chain, which are buried and unreactive in the native trimer. In the pathway to the native trimer, both in vivo and in vitro transient interchain disulfide bonds are formed and reduced. This occurs in the protrimer, an intermediate in the formation of the interdigitated beta-sheets of the trimeric tailspike. Each of the eight cysteines was replaced with serine by site specific mutagenesis of the cloned P22 tailspike gene and the mutant genes expressed in Escherichia coli. Although the yields of native-like Cys>Ser proteins varied, sufficient soluble trimeric forms of each of the eight mutants accumulated to permit purification. All eight single Cys>Ser mature proteins maintained the high thermostability of the wild type, as well as the wild-type biological activity in forming infectious virions. Thus, these cysteine thiols are not required for the stability or activity of the native state. When their in vivo folding and assembly kinetics were examined, six of the mutant substitutions -C267S, C287S, C458S, C613S, and C635S--were significantly impaired at higher temperatures. Four--C290S, C496, C613S, and C635--showed significantly impaired kinetics even at lower temperatures. The in vivo folding of the C613S/C635S double mutant was severely defective independent of temperature. Since the trimeric states of the single Cys>Ser substituted chains were as stable and active as wild type, the impairment of tailspike maturation presumably reflects problems in the in vivo folding or assembly pathways. The formation or reduction of the transient interchain disulfide bonds in the protrimer may be the locus of these kinetic functions. PMID- 11266627 TI - A comparison of salts for the crystallization of macromolecules. AB - Thirty-one proteins and viruses that we knew from our own experience could be crystallized, or had been reported to have been crystallized by others, were investigated. In this experiment, each protein or virus was subjected to a crystallization screen of 12 different salts, each titrated to pH 7.2 beforehand, at concentrations ranging from 20% saturation to 90% saturation. Eight macromolecules failed to crystallize at all from any salt and were omitted from consideration. From the remaining 23 proteins, each salt was scored according to how many proteins and viruses it successfully crystallized. Among several results, one was particularly striking. Sodium malonate clearly was much more successful than any other salt, resulting in the crystallization of 19 of the 23 macromolecules, almost twice as effective as the next most successful salt, which was a draw between sodium acetate, sodium tartrate, sodium formate, and ammonium sulfate (11 of 22). The high success rate of sodium malonate in producing crystals was even more impressive when an overall unique success rate with individual macromolecules was considered. PMID- 11266628 TI - Detection of early gene expression changes during activation of human primary lymphocytes by in vitro synthesis of proteins from polysome-associated mRNAs. AB - The rapid increase in protein synthesis during the mitogenic stimulation of human peripheral blood lymphocyte is the result of global and specific translational control mechanisms. To study some of these mechanisms, we examined the in vitro translatability of mRNAs associated with the polyribosome fraction. Polyribosome fractions were isolated from lymphocytes after activation with ionomycin and the phorbol ester PMA. The associated PAmRNAs were translated in the presence of mRNA depleted rabbit reticulocyte lysate and [(35)S]Met, and the protein products were analyzed by SDS--PAGE and autoradiography. There was little synthesis of protein from the PAmRNAs isolated from unactivated T cells, but the PAmRNAs isolated from activated T cells showed a rapid increase in translatability. Translation of the PAmRNAs was sensitive to edeine and m7GTP, suggesting their cap-dependent translation. With activation, the majority of proteins showed increasing in vitro translation, but two proteins, p72 and p33, were found to have increased synthesis within 30 min, which decreased in 1 h. Transcription inhibitors were used to ascertain if regulation of their expression was transcriptional or translational. To identify these proteins, we used biotinylated lysine during the in vitro translation reaction, and we extracted the biotinylated protein by using streptavidin magnetic beads. The protein product was analyzed by mass spectrometry. p33 was identified as a prohibitin-like protein (BAP37), but the identification of p72 was not found in the databases. The distinct up-regulation and down-regulation of their protein expression suggest their tightly controlled regulation during early T cell activation. PMID- 11266629 TI - Coupling of antibodies via protein Z on modified polyoma virus-like particles. AB - Therapeutic application of virus-based delivery systems often implies a change of the tropism of these vectors. This can be achieved by insertion of polypeptides (e.g., antibody fragments) in viral coat proteins. Such fusion proteins have only been used in viral vectors so far and, as part of a virus, they have not been available for a detailed biophysical characterization. We analyzed a fusion protein called VP1-Z, which is based on the polyoma virus coat protein VP1 and protein Z. Protein Z is an engineered antibody-binding domain derived from protein A from Staphylococcus aureus. The fusion VP1-Z was constructed by insertion of protein Z in the HI-loop of VP1. As wild-type VP1, VP1-Z formed pentameric capsomers and assembled to VLPs in vitro. The stability of these particles was very similar compared to that of VLPs of wild-type VP1. Protein Z was fully structured in the fusion protein and was still capable of binding antibodies on the surface of VLPs of VP1-Z. Using this fusion protein, we could change the tropism of polyoma VLPs toward cells presenting on their surface the antigen of the coupled antibody. PMID- 11266630 TI - Nested allosteric interactions in the cytoplasmic chaperonin containing TCP-1. AB - Initial rates of ATP hydrolysis by the chaperonin containing TCP-1 (CCT) from bovine testis were measured as a function of ATP concentration. Two allosteric transitions are observed: one at relatively low concentrations of ATP (<100 microM) and the second at higher concentrations of ATP. The data suggest that CCT has positive intra-ring cooperativity and negative inter-ring cooperativity in ATP hydrolysis, with respect to ATP, as previously observed in the case of GroEL. It is shown that the relatively weak positive intra-ring cooperativity found in the case of CCT may be due to heterogeneity in its subunit composition. Our results suggest that nested allosteric behavior may be common to chaperone double ring systems. PMID- 11266631 TI - Designed protein G core variants fold to native-like structures: sequence selection by ORBIT tolerates variation in backbone specification. AB - The solution structures of two computationally designed core variants of the beta 1 domain of streptococcal protein G (G beta 1) were solved by (1)H NMR methods to assess the robustness of amino acid sequence selection by the ORBIT protein design package under changes in protein backbone specification. One variant has mutations at three of 10 core positions and corresponds to minimal perturbations of the native G beta 1 backbone. The other, with mutations at six of 10 positions, was calculated for a backbone in which the separation between G beta 1's alpha-helix and beta-sheet was increased by 15% relative to native G beta 1. Exchange broadening of some resonances and the complete absence of others in spectra of the sixfold mutant bespeak conformational heterogeneity in this protein. The NMR data were sufficiently abundant, however, to generate structures of similar, moderately high quality for both variants. Both proteins adopt backbone structures similar to their target folds. Moreover, the sequence selection algorithm successfully predicted all core chi(1) angles in both variants, five of six chi(2) angles in the threefold mutant and four of seven chi(2) angles in the sixfold mutant. We conclude that ORBIT calculates sequences that fold specifically to a geometry close to the template, even when the template is moderately perturbed relative to a naturally occurring structure. There are apparently limits to the size of acceptable perturbations: In this study, the larger perturbation led to undesired dynamic behavior. PMID- 11266633 TI - Meeting report. Methods in Protein Structural Analysis Conference (MPSA2000), Charlottesville, Virginia, September 16-20, 2000. PMID- 11266632 TI - MPtopo: A database of membrane protein topology. AB - The reliability of the transmembrane (TM) sequence assignments for membrane proteins (MPs) in standard sequence databases is uncertain because the vast majority are based on hydropathy plots. A database of MPs with dependable assignments is necessary for developing new computational tools for the prediction of MP structure. We have therefore created MPtopo, a database of MPs whose topologies have been verified experimentally by means of crystallography, gene fusion, and other methods. Tests using MPtopo strongly validated four existing MP topology-prediction algorithms. MPtopo is freely available over the internet and can be queried by means of an SQL-based search engine. PMID- 11266635 TI - Why ethical standards? An introduction to the perianesthesia standards for ethical practice. PMID- 11266636 TI - Perianesthesia standards for ethical practice. PMID- 11266637 TI - Ethical decision making. AB - Members of the health care profession have a responsibility to identify and regulate their practice to protect consumers and assure the delivery of quality service. In this case study in which a patient wakes up during surgery, the nurse's code of ethics, standards of care, and ethical decision-making models provide guidance toward the resolution of this clinical dilemma. Such resolution requires sound nursing knowledge, knowledge of the facts of the situation, knowledge of the law, and application of critical thinking skills to facilitate positive outcomes for all parties involved. PMID- 11266638 TI - Evaluation of orthostatic blood pressure testing as a discharge criterion from PACU after spinal anesthesia. AB - Discharge readiness from a Phase I PACU after spinal anesthesia is frequently determined by recovery of sensory/motor function. However, no data exist indicating that recovery of sensory/motor function adequately predicts hemodynamic stability after spinal anesthesia. The conservative practice of waiting until the sensory/motor effects of spinal anesthesia have completely worn off often requires patients to remain in PACU for prolonged periods of time. The purpose of this study was to determine the safety and efficacy of using orthostatic blood pressure (BP) testing as a discharge criterion from PACU after spinal anesthesia. This study used a prospective, descriptive design to measure changes in mean arterial pressure (MAP) during orthostatic BP testing at 30 minute intervals after admission to the PACU following spinal anesthesia. A convenience sample of 121 patients admitted through the Same Day Surgery (SDS) unit was used. Results show that orthostatic BP criterion was safe and effective as an alternative to sensory/motor criteria in assessing hemodynamic stability and reducing the amount of time patients spend in the PACU after spinal anesthesia. This is a U.S. government work. There are no restrictions on its use. PMID- 11266639 TI - Preoperative assessment: safety considerations for patients taking herbal products. AB - The increased use of herbal therapy and other dietary supplements in the past decade has increased the risk of complications for patients undergoing surgery. The use of herbal products is projected to rise in the United States. Therefore, it is important for the perianesthesia nurse to be aware of the herbal products that patients are taking and the possible anesthesia and/or surgical complications related to herbal products. Current resources are reviewed in this article to increase awareness for the perianesthesia nurse. Recommendations for establishing local protocols are given, and patient education on the use of herbal products is also considered. PMID- 11266642 TI - Preparing for surgery: providing the details. PMID- 11266641 TI - Family visitation in PACU. PMID- 11266640 TI - Public speaking 101. PMID- 11266643 TI - Product review: refrigerated malignant hyperthermia cart. PMID- 11266644 TI - Evidence-based practice part 2: reliability and validity of selected acute pain instruments. AB - Pain management is an important aspect of perianesthesia patient care. PACU nurses need to be familiar with pain measurement to judge effectiveness of pain management. In fact, the 1999-2000 Joint Commission on Accreditation of Healthcare Organizations' (JCAHO) guidelines have included the measurement of pain before and after pain treatment in their standards of practice. This article reviews selected pain instruments that could be used to measure pain in perianesthesia patients and the available reliability and validity of the instruments. PMID- 11266646 TI - Accepting the challenge. PMID- 11266647 TI - Where is the evidence that cyclooxygenase inhibition is the primary cause of nonsteroidal anti-inflammatory drug (NSAID)-induced gastrointestinal injury? Topical injury revisited. AB - In this commentary, we take a critical look at the concept that the gastrointestinal (GI) side-effects of nonsteroidal anti-inflammatory drugs (NSAIDs) are due to the ability of these drugs to inhibit cyclooxygenase-1 (COX 1) that is constitutively expressed in the GI mucosa. Indeed, development of the new "super aspirins," such as Celebrex and Vioxx, that selectively inhibit the inducible COX-2, expressed in areas of inflammation, is a direct outgrowth of this concept. We discuss evidence from both the laboratory and the clinic that appears to be inconsistent with the above concept, and cite a number of examples where the depletion of mucosal prostaglandin levels and the development of GI injury can be dissociated. Instead, we revisit the possibility that NSAID-induced GI side-effects are mostly due to the ability of these drugs to topically injure the GI mucosa. We devote the remainder of the commentary to presenting evidence from our and other laboratories that NSAIDs can directly attenuate the surface hydrophobic barrier of the GI mucosa due to their ability to bind to zwitterionic phospholipids, and that even systemically administered NSAIDs that are secreted into the bile may induce GI ulceration and/or bleeding due to phospholipid interactions and the development of topical mucosal injury. PMID- 11266648 TI - Growth reduction in glioma cells after treatment with tetradecylthioacetic acid: changes in fatty acid metabolism and oxidative status. AB - During aerobic metabolism, a small amount of partially reduced oxygen is produced, yielding reactive oxygen species (ROS). Peroxisomes and mitochondria are major contributors to cellular ROS production, which is normally balanced by consumption by antioxidants. The fatty acid analogue tetradecylthioacetic acid (TTA) promotes mitochondrial and peroxisomal proliferation, and may induce oxidative stress and change the growth potential of cancer cells. In the present study, we found that TTA reduced [(3)H]thymidine incorporation in the glioma cell lines BT4Cn (rat), D54Mg (human), and GaMg (human) in a dose- and time-dependent manner. The 50% inhibitory TTA doses were approximately 125 microM for BT4Cn and D54Mg cells and 40 microM for GaMg cells after 4 days. alpha-Tochopherol counteracted this inhibition in GaMg cells. TTA enhanced the oxidation of [1 (14)C]palmitic acid, which could be explained by stimulation of enzymes involved in peroxisomal (fatty acyl-CoA oxidase) and/or mitochondrial (carnitine palmitoyltransferase) fatty acid oxidation. The glutathione content and the activities of glutathione peroxidase, glutathione reductase, and glutathione S transferase were differentially affected. Increased malondialdehyde (MDA) production was seen in TTA-treated GaMg and D54Mg cells, but not in BT4Cn cells, in vitro. In BT4Cn tumor tissue from TTA-treated rats, MDA was increased while the alpha-tocopherol content tended to decrease. TTA increased the level of cytosolic cytochrome c in BT4Cn cells, which suggests induction of apoptotic cascades. Although several mechanisms are likely to be involved in the TTA mediated effects on growth, we propose that modulation of cellular redox conditions caused by changes in fatty acid metabolism may be of vital importance. PMID- 11266649 TI - Expression of rat liver long-chain acyl-CoA synthetase and characterization of its role in the metabolism of R-ibuprofen and other fatty acid-like xenobiotics. AB - Our investigations of fatty acid metabolism and epimerization of the 2 arylpropionic acid derivative, R-ibuprofen, resulted in the successful purification of an acyl-CoA synthetase from rat liver microsomes that catalyzes the formation of both palmitoyl-CoA and R-ibuprofenoyl-CoA. To investigate whether R-ibuprofenoyl-CoA synthetase and long-chain acyl-CoA synthetase (LACS) are identical enzymes, we cloned the cDNA from LACS into the pQE30 expression vector and transformed the construct into Escherichia coli M15[pREP4]. Induction of the bacterial protein synthesis with 0.2 mM isopropyl-beta-D-galactoside resulted in a strong, time-dependent increase in LACS protein as determined by Western blot analysis using a polyclonal rabbit anti-LACS antibody. Incubations of the recombinantly expressed protein with palmitic acid as physiological LACS substrate or R-ibuprofen in the presence of Mg2+, ATP, and CoA resulted in a 5 fold increase in the thioesterification of both substrates. Western blot analysis using tissue homogenates of rat liver, heart, kidney, lung, brain, and ileum showed that LACS was found in every tissue investigated, with the greatest expression in the liver. Similar results were obtained with activity measurements using R-ibuprofen and palmitic acid as substrates. Northern blot analysis revealed a hybridization with a 3.8-kb mRNA transcript in rat liver, heart, and kidney, but no signal was observed in lung, brain and ileum, suggesting the expression of different LACS isoform(s) in these organs. In summary, our results further show that R-ibuprofenoyl-CoA synthetase and long-chain acyl-CoA synthetase are identical enzymes that are involved in the metabolism of various xenobiotics. PMID- 11266650 TI - [3H]MRS 1754, a selective antagonist radioligand for A(2B) adenosine receptors. AB - MRS 1754 [N-(4-cyanophenyl)-2-[4-(2,3,6,7-tetrahydro-2,6-dioxo-1,3-dipropyl-1H purin-8-yl)-phenoxy]acetamide] is a selective antagonist ligand of A(2B) adenosine receptors. This is the least well-defined adenosine receptor subtype, and A(2B) antagonists have potential as antiasthmatic drugs. For use as a radioligand, MRS 1754, a p-cyanoanilide xanthine derivative, was tritiated on the propyl groups in a two-step reaction using a p-carboxamido precursor, which was dehydrated to the cyano species using trifluoroacetic anhydride. [3H]MRS 1754 (150 Ci/mmol) bound to recombinant human A(2B) adenosine receptors in membranes of stably transfected HEK-293 cells. Specific binding was saturable, competitive, and followed a one-site model, with a K(D) value of 1.13 +/- 0.12 nM and a B(max) value of 10.9 +/- 0.6 pmol/mg protein. Specific binding utilizing 0.7 nM [3H]MRS 1754 was > 70% of total binding. The affinity calculated from association and dissociation binding constants was 1.22 nM (N = 4). Binding to membranes expressing rat and human A(1) and A(3) adenosine receptors was not significant, and binding in membranes of HEK-293 cells expressing human A(2A) receptors was of low affinity (K(D) > 50 nM). The effects of cations and chelators were explored. Specific binding was constant over a pH range of 4.5 to 6.5, with reduced binding at higher pH. The pharmacological profile in competition experiments with [3H]MRS 1754 was consistent with the structure-activity relationship for agonists and antagonists at A(2B) receptors. The K(i) values of XAC (xanthine amine congener) and CPX (8-cyclopentyl-1,3-dipropylxanthine) were 16 and 55 nM, respectively. NECA (5'-N-ethylcarboxamidoadenosine) competed for [3H]MRS 1754 binding with a K(i) of 570 nM, similar to its potency in functional assays. Thus, [3H]MRS 1754 is suitable as a selective, high-affinity radioligand for A(2B) receptors. PMID- 11266651 TI - Association of deletions and translocation of the reduced folate carrier gene with profound loss of gene expression in methotrexate-resistant K562 human erythroleukemia cells. AB - Severe impairment of methotrexate membrane transport in methotrexate-resistant K562 (K500E) cells was characterized by a nearly complete loss of reduced folate carrier (RFC) transcripts and RFC protein. As determined by 5'-rapid amplification of cDNA ends (5'-RACE), approximately 93% of the RFC transcripts in wild-type cells contained the KS43 5'-untranslated region transcribed from the RFC-B promoter. KS43 transcripts decreased > 90% in K500E cells. The basal and full-length RFC-B promoters were more active (3- and 2-fold, respectively) in directing transcription of a luciferase reporter gene in K500E than in wild-type cells. Treatment with a demethylating agent, 5-aza-2'-deoxycytidine, or with a histone deacetylase inhibitor, trichostatin A, did not increase the levels of RFC transcripts in K500E cells. No differences in RFC gene structure were detected between the lines on Southern blots; however, the RFC signals were decreased approximately 60% in K500E cells. DNA sequences were identical between the lines for the RFC coding region and the two 5'-non-coding exons and their respective promoters. Spectral karyotype analysis and fluorescence in situ hybridization in wild-type cells showed two normal chromosome 21 copies and one or two marker chromosomes, each with an RFC signal. In K500E cells, the RFC gene locus was no longer localized to a normal chromosome 21 (at 21q22.2), and a single RFC signal was associated with a small metacentric chromosome, characterized by a 21/22 translocation. Our results suggest that loss of RFC transcripts in K500E cells is unrelated to changes in the levels of critical transcription factors, or to differences in the extent of RFC promoter methylation or core histone deacetylation. Rather, this phenotype is due to the loss of one or more RFC alleles, and to a translocation of the remaining RFC allele with the formation of a 21/22 fusion chromosome. PMID- 11266653 TI - Enzymatic activation of a new antitumour drug, 5-diethylaminoethylamino-8 hydroxyimidazoacridinone, C-1311, observed after its intercalation into DNA. AB - The imidazoacridinone derivative, C-1311, is a new antitumour agent that exhibits strong antitumour activity against experimental colorectal cancer and has been selected for entry into clinical trial. The compound has previously been shown to have DNA non-covalent binding properties in vitro and to bind irreversibly to DNA of tumour cells. The latter effect has also been observed in a cell-free system, but only in the presence of activated enzymes. The present studies were aimed at finding out whether and in what way the enzymatic activation of C-1311 and its non-covalent binding to DNA influence or depend on each other. Enzymatic activation was performed with a model system containing horseradish peroxidase (HRP) and hydrogen peroxide (H2O2) and was followed by UV-VIS spectroscopy and by HPLC with UV-VIS and electrospray ionisation mass spectrometry detection. DNA non covalent binding was studied in the cell-free system by means of an unwinding assay and UV-VIS spectroscopy. It was shown that C-1311 was oxidised by the HRP/H2O2 system in a manner dependent on the drug:H2O2 ratio. In the case of ratios of 1:3 and 1:5, the reaction gave highly reactive species that were quickly transformed into the further products p2 and p3 that were unable to intercalate into DNA. In the presence of DNA, C-1311 first intercalated into DNA and the intercalated compound was then oxidised. This oxidation was directed to only one product. Therefore, DNA seems to play the role of a "scavenger" of the reactive oxidation product(s) yielded from the intercalated drug and prevents its further deactivation. We conclude that, under the conditions studied, intercalation of C-1311 into DNA is followed by its HRP-mediated activation, giving rise to the intercalated species that might irreversibly bind to DNA. Since peroxidase-type enzymes are present in the cell nucleus, the proposed sequence of events may also be expected to take place in the cellular environment in vivo. PMID- 11266652 TI - Enhancement of antioxidant and anti-inflammatory activities of bioflavonoid rutin by complexation with transition metals. AB - The antioxidant and anti-inflammatory activities of two transition metal complexes of bioflavonoid rutin, Fe(rut)Cl(3) and Cu(rut)Cl(2), were studied. It was found that Cu(rut)Cl(2) was a highly efficient in vitro and ex vivo free radical scavenger that sharply decreased (by 2-30 times compared to the parent rutin): oxygen radical production by xanthine oxidase, rat liver microsomes, and rat peritoneal macrophages; the formation of thiobarbituric acid-reactive products in microsomal lipid peroxidation; and the generation of oxygen radicals by broncho-alveolar cells from bleomycin-treated rats. The copper-rutin complex was also a superior inhibitor of inflammatory and fibrotic processes (characterized by such parameters as macrophage/neutrophil ratio, wet lung weight, total protein content, and hydroxyproline concentration) in the bleomycin treated rats. The antioxidant activity of Fe(rut)Cl(3) was much lower and in some cases approached that of rutin. Fe(rut)Cl(3) also stimulated to some degree spontaneous oxygen radical production by macrophages. We suggested that the superior antioxidant and anti-inflammatory activity of the copper-rutin complex is a consequence of its acquiring the additional superoxide-dismuting copper center. The inhibitory activity of Fe(rut)Cl(3) was lower, probably due to the partial reduction into Fe(rut)Cl(2) in the presence of biological reductants; however, similarly to the copper-rutin complex, this complex efficiently suppressed lung edema. PMID- 11266654 TI - Peripheral-type benzodiazepine receptor ligands: mitochondrial permeability transition induction in rat cardiac tissue. AB - Strong evidence is emerging that mitochondrial permeability transition (MPT) may be important in certain physiological conditions and, above all, in the processes of cell damage and death. Reversible MPT, triggered by inducing agents in the presence of calcium ions, has resulted in the opening of a dynamic multiprotein complex formed in the inner mitochondrial membrane and has caused large-amplitude mitochondrial swelling. In the present work, the exposure of de-energized rat cardiac mitochondria to peripheral benzodiazepine receptor (PBR) ligands (1-(2 chlorophenyl-N-methyl-1-methylpropyl)-3-isoquinolinecarboxamide (PK 11195), 7 chloro-5-(4-chlorophenyl)-1,3-dihydro-1-methyl-2H-1,4-benzodiazepin-2-one (Ro5 4864), and diazepam) produced a dose-dependent and cyclosporin A (CSP)-sensitive loss of absorbance, which was indicative of mitochondrial swelling. By contrast, the addition of a high-affinity central benzodiazepine receptor ligand (clonazepam) was ineffective, even at the highest concentration tested. The ultrastructural changes associated with swelling were similar in mitochondria exposed either to PK 11195 or to calcium. Supporting the apoptotic role of PK 11195-induced swelling, supernatants from mitochondria that had undergone permeability transition caused apoptotic changes in isolated cardiac nuclei. In addition, ultrastructural abnormalities were observed in rat cardiac tissue following in vivo PK 11195 administration, with these abnormalities being prevented by CSP co-administration. These data indicate that PBR ligands induce mitochondrial permeability transition and ultrastructural alterations in isolated cardiac mitochondria as well as in myocardiocytes, suggesting a novel strategy for studying the implication of PBR ligands as apoptosis inducers, through a probable effect on the MPT pore. PMID- 11266655 TI - Effect of the glutathione/glutathione disulfide redox couple on thiopurine methyltransferase. AB - The susceptibility of recombinant human thiopurine methyltransferase (hTPMT) to thiol-disulfide exchange was investigated. The enzyme was incubated in buffers of the redox couple GSH and GSSG. The values of the chosen concentrations and concentration ratios of the redox couple equaled those expected to occur in vivo. Activity measurements of the enzyme over time in these buffers at 30 degrees C indicated that thiol-disulfide exchange may be a part of the posttranslational modulation of hTPMT activity. Activity varied between 5% and 100%, with the lowest activities in buffers of low [GSH]/[GSSG] concentration ratios and of low total concentration of the redox couple. A thiol-disulfide exchange mechanism involving a mixed disulfide was proposed. Titration of the protein thiol groups with Ellmann's reagent (5,5'-dithiobis[2-nitrobenzoic acid]) revealed that at least two protein thiols were readily accessible for conjugation with the reagent, while others were conjugated more slowly. The previous model of hTPMT constructed by our group was in accordance with the experimental results. Inspection of the model indicated that one of the protein thiols subject to slow thiol-disulfide exchange may be situated at the binding site of the co-substrate of the enzyme and thus be responsible for the glutathione/glutathione disulfide modulation of the activity of hTPMT. PMID- 11266656 TI - No reduction of alpha-tocopherol quinone by glutathione in rat liver microsomes. AB - The cell membrane is protected against lipid peroxidation by endogenous antioxidants such as vitamin E (alpha-tocopherol). The oxidised form of alpha tocopherol (alpha-tocopherol quinone) does not have this antioxidant function. However, the literature indicates that alpha-tocopherol quinone can be reduced to alpha-tocopherol in vivo and thereby will add to the total antioxidant potential (Moore AN, Ingold KU. Free Radic Biol Med 1997;22:931-4). We found that GSH (reduced glutathione) did not mediate the reduction of alpha-tocopherol quinone, either directly in solution or in rat liver microsomes fortified with alpha tocopherol quinone. This renders GSH a less likely candidate for alpha-tocopherol quinone reduction in vivo. In addition, alpha-tocopherol quinone did not enhance GSH-dependent protection against lipid peroxidation, either in control microsomes, or in vitamin E-extracted microsomes. Indeed, alpha-tocopherol quinone blocked GSH-dependent protection against lipid peroxidation in vitamin E extracted microsomes. This indicates that alpha-tocopherol quinone can act as a pro-oxidant. PMID- 11266657 TI - Debenzylation of O(6)-benzyl-8-oxoguanine in human liver: implications for O(6) benzylguanine metabolism. AB - O(6)-Benzylguanine (BG) effectively inactivates the DNA repair protein O(6) alkylguanine-DNA alkyltransferase, and enhances the effectiveness of 1,3-bis(2 chloroethyl)-1-nitrosourea in cells in culture and tumor-bearing animals. BG is presently in phase II clinical trials. In humans, BG is converted to O(6)-benzyl 8-oxoguanine (8-oxoBG), a longer-lived, yet equally potent inactivator. We have isolated and identified the debenzylated product, 8-oxoguanine, in plasma and urine of patients following administration of BG. The purpose of this work was to determine the human liver enzymes responsible for the debenzylation of 8-oxoBG. Therefore, 8-oxoBG was incubated with human liver microsomes and cytosol, and the concentration of 8-oxoguanine was determined. No appreciable product was formed in the cytosol; however, increasing amounts of 8-oxoguanine were formed with increasing concentrations of pooled human liver microsomes. The amount of 8 oxoguanine formed increased with time and substrate concentration. Co-incubation of human liver microsomes with 8-oxoBG and various cytochrome P450 isoform selective inhibitors suggested the possible involvement of CYP1A2, 2E1, and/or 2A6 in this reaction. Incubation of 8-oxoBG with baculovirus cDNA-overexpressed CYP1A2, 2E1, 2A6, and 3A4 demonstrated that formation of 8-oxoguanine was due mainly to CYP1A2. Debenzylation of 8-oxoBG complied with Michaelis-Menten kinetics with K(m) and V(max) values of 35.9 microM and 0.59 pmol/min/pmol of CYP1A2, respectively. CYP1A2 appears to be mainly responsible for the debenzylation of 8-oxoBG in human liver. PMID- 11266658 TI - 2'-O-Acyl/alkyl-substituted arabinosyl nucleosides as inhibitors of human mitochondrial thymidine kinase. AB - Introduction of a bulky lipophilic acyl entity at the 2'-OH position of both 1 beta-D-arabinofuranosylthymine (araT) and (E)-5-(2-bromovinyl)-1-beta-D arabinofuranosyluracil (BVaraU), consistently resulted in a marked ( approximately 10-fold) increase in the inhibitory activity of these new arabinosyl nucleoside analogues for the mitochondrial thymidine kinase (TK-2) catalysed conversion of 2 microM [methyl-(3)H]dThd to [methyl-(3)H]dTMP. The most potent derivatives were inhibitory to [methyl-(3)H]dThd phosphorylation by TK-2 within the lower micromolar concentration range. Substitution of the arabinosyl nucleoside derivatives with the acyl groups also dramatically increased the selectivity of these compounds. The inhibitory activity of araT and BVaraU to dThd phosphorylation by other related nucleoside kinases, including herpes simplex virus type 1 TK, varicella-zoster virus TK, and cytosolic TK-1, was completely annihilated upon 2'-O-acyl substitution (IC(50) > or = 1000 microM). Kinetic analysis revealed purely competitive inhibition of 2'-O-acyl-BVaraU against TK-2-catalysed thymidine phosphorylation (K(i)/K(m): 2.3). However, 2'-O acyl-BVaraU was extremely poorly converted to the corresponding arabinosyl nucleoside 5'-monophosphate by TK-2 as revealed by [gamma-(32)P]phosphate transfer studies from [gamma-(32)P]ATP. Thus, the 2'-O-acyl derivatives of BVaraU did not behave as substrates, but rather as potent and highly selective inhibitors of TK-2. This is the first report on such a highly selective arabinosyl nucleoside inhibitor of mitochondrial TK-2, and opens perspectives for the rational design of selective mitochondrial TK-2 inhibitors. PMID- 11266659 TI - Investigation of nicotine binding to THP-1 cells: evidence for a non-cholinergic binding site. AB - Nicotine is known to modulate immune function, but reports have produced conflicting evidence as to whether nicotinic acetylcholine (nACh) receptors are responsible for these effects. This study was designed to examine the identity of nicotine-binding sites on immune cells using a human leukaemic monocytic cell line, THP-1, that is known to have functions that are modulated by nicotine. Binding studies were performed on THP-1 whole cells using [3H]nicotine as a probe to analyse any possible nicotine-binding sites on these cells. Saturation analysis of THP-1 cells revealed the presence of 2 distinct binding sites; one with a K(d1) of 3.5 +/- 2.1 x 10(-9) M and a B(max1) of 4100 +/- 560 sites/cell (designated the high-affinity site) and the other with a K(d2) of 27 +/- 9.2 x 10(-9) M and a B(max2) of 11,600 +/- 630 sites/cell (low-affinity site). Competition analysis revealed that one site had an affinity to a range of cholinergic ligands including epibatidine and cytisine. When saturation analysis of [3H](-)-nicotine to THP-1 cells was performed in the presence of 1 x 10(-6) M epibatidine, only one binding site was detected. Comparisons of K(d) and B(max) values showed that the high-affinity site was not occluded by epibatidine. No drugs tested displayed any affinity for the high-affinity site except the two enantiomers of nicotine. The high-affinity site was shown to be stereoselective for the (+)-enantiomer of nicotine as shown by K(i) values produced by competition analysis in the presence of 1 x 10(-6) M epibatidine. These values were 5.7 +/- 0.32 x 10(-11) M and 1.9 +/- 4.9 x 10(-9) M for (+)-nicotine and (-) nicotine, respectively. This study presents evidence for a possible non cholinergic binding site that may play a role in the mechanism of immunomodulation by nicotine. PMID- 11266660 TI - 2-Bromoethylamine as a potent selective suicide inhibitor for semicarbazide sensitive amine oxidase. AB - Semicarbazide-sensitive amine oxidase (SSAO) catalyzes the deamination of methylamine and aminoacetone to produce toxic aldehydes, i.e. formaldehyde and methylglyoxal, as well as hydrogen peroxide and ammonia. An increase of SSAO activity was detected by different laboratories in patients suffering from vascular disorders, i.e. diabetes and myocardial infarction. The enzyme has been suggested to play a role in vascular endothelial damage and atherogenesis. To date, there are no selective SSAO inhibitors. In the present study, 2 bromoethylamine (2-BrEA) was found to be a highly effective and selective inhibitor of SSAO obtained from different sources. The inhibition was irreversible and time dependent. It was competitive when the enzyme was not preincubated with the inhibitor, but became noncompetitive after incubation of the enzyme with 2-BrEA. The aldehyde trapping agent o-phenylenediamine was capable of preventing 2-BrEA-induced inhibition of SSAO activity. An aldehyde product was detected to be an initial product of 2-BrEA after it was incubated with SSAO. The inhibition, therefore, is mechanism-based. The SSAO inhibitory effects of eight structural analogues of 2-BrEA were assessed. It was concluded that a bromine atom at the beta position is quite important for exerting high potency of SSAO inhibition. The inhibition of SSAO activity by 2-BrEA was also demonstrated in vivo. It increased the urinary excretion of methylamine, an endogenous substrate for SSAO, in mice. 2-BrEA can be employed as a very useful tool in the investigation of SSAO. PMID- 11266662 TI - Blocking action of cytochalasin D on protein kinase A stimulation of a stretch activated cation channel in renal epithelial A6 cells. AB - We have shown that the apical membrane of renal epithelial A6 cells has a 29-pS stretch-activated nonselective cation (NSC) channel, which is activated by cytosolic cyclic AMP (cAMP) (J Gen Physiol 1997;110:327-36). In general, downstream signalings of cAMP are mediated through a cAMP-activated protein kinase (protein kinase A, PKA)-dependent pathway. Therefore, to study if the channel is activated by a PKA-dependent pathway, we applied a PKA catalytic subunit directly to the channel from the cytosolic surface in cytosol-free excised inside-out patches, using the single channel recording (patch clamp) technique. Application of PKA catalytic subunit with 2 mM ATP increased the open probability (P(o)) of the channel from 0.11 +/- 0.04 to 0.58 +/- 0.10 (mean +/- SD, N = 11, P < 0.001). The channel has a gating kinetics "C(L) <--> C(S) <--> O, " where C(L,) C(S,) and O are the long closed state, the short closed state, and the open state, respectively. PKA influenced the communication of the channel between C(L) and C(S) without affecting the communication between C(S) and O, leading the channel to only stay in C(S) and O. The PKA-induced increase in P(o) was attributable to the interruption of communication between C(L) and C(S) or to the reduction of time the channel stays in C(L.) Pretreatment with cytochalasin D (Cyt-D), an inhibitor of the polymerization of actin filaments, blocked the stimulatory effect of PKA on the channel. These observations suggest that phosphorylation of polymerized actin filaments regulates the gating kinetics of a stretch-activated channel in renal epithelium. PMID- 11266661 TI - Stimulation of stress-activated but not mitogen-activated protein kinases by tumour necrosis factor receptor subtypes in airway smooth muscle. AB - The multifunctional cytokine tumour necrosis factor-alpha (TNF) displays many physiological effects in a variety of tissues, especially proliferative and cytotoxic actions in immunological cells. Recently, we uncovered an important new mechanism by which TNF can sensitise airway smooth muscle (ASM) to a fixed intracellular Ca2+ concentration which in vivo would produce a marked hypercontractility of the airways. Here, we report that both 50-60 kDa type I TNFR (TNFR1) and 70-80 kDa type II TNFR (TNFR2) receptor subtypes were expressed in ASM cells and selectively activated the stress kinases, c-Jun N-terminal kinase and p38 mitogen-activated protein kinase (p38 MAPK). However, TNF caused no activation of p42/p44 MAPK or cytosolic phospholipase A(2) activity. In contrast, TNF stimulation of the TNFR1, but not the TNFR2, elicited nuclear factor-kappaB transcription factor function, a species known to be important in mediation of certain inflammatory cellular responses. This is the first report of TNF receptor subtypes in ASM cells causing selective kinase activation, which may prove important in therapeutic strategies for treating immune airway disorders such as chronic obstructive pulmonary disease and asthma. PMID- 11266664 TI - Frictional heating of total hip implants. Part 1: measurements in patients. AB - Hip implants heat up due to friction during long lasting, high loading activities like walking. Thermal damage in the surrounding soft and hard tissues and deteriorated lubrication of synovial fluid could contribute to implant loosening. The goal of this study was to determine the implant temperatures in vivo under varying conditions. Temperatures and contact forces in the joints were measured in seven joints of five patients using instrumented prostheses with alumina ceramic heads and telemetry data transmission. The peak temperature in implants with polyethylene cups rose up to 43.1 degrees C after an hour of walking but varied considerably individually. Even higher temperatures at the joints are probable for patients with higher body weight or while jogging. The peak temperature was lower with a ceramic cup, showing the influence of friction in the joint. During cycling the peak temperatures were lower than during walking, proving the effect of force magnitudes on the produced heat. However, no positive correlation was found between force magnitude and maximum temperature during walking. Other individual parameters than just the joint force influence the implant temperatures. Based on the obtained data and the available literature about thermal damage of biological tissues a detrimental effect of friction induced heat on the stability of hip implants cannot be excluded. Because the potential risk for an individual patient cannot be foreseen, the use and improvement of low friction implant materials is important. PMID- 11266665 TI - Frictional heating of total hip implants. Part 2: finite element study. AB - Due to higher friction artificial hip joints warm up more than natural joints during walking and other continuous activities. This could lead to thermal damage in the surrounding tissues and be a reason for long-term implant loosening, an effect which has not yet been investigated. In vivo measurements with instrumented implants showed temperatures inside the prosthetic head up to 43.1 degrees C (Part 1 of this work). Based on the experimental data a finite element model was developed to calculate the temperatures in the tissues surrounding the hip implant to determine whether these tissues can heat up to critical levels. Various parameters were investigated which could account for the variations in the measured temperatures in the patients, including the perfusion rate in tissues, the volume of synovial fluid, and different implant materials. We found that the synovial fluid is most endangered by thermal damage and consequent deterioration of lubricating properties. Implants with a cobalt-chromium head and a polyethylene cup are unfavourable as they can elevate the temperature in the synovia to more than 46 degrees C. With regard to thermal properties stems made from cobalt-chromium alloys are superior to titanium stems, by better conducting heat to the femur and minimizing the synovial fluid temperature. Factors determining the temperatures during walking are insufficiently known or cannot be determined in the individual patient. Therefore, the risk of a thermally induced implant loosening cannot currently be estimated. Under unfavourable conditions such a risk exists, however. General improvements of implant materials and clinical studies on the possibility of implant loosening due to high temperatures are therefore required. PMID- 11266667 TI - Dependence of release variables in the shot put. AB - When the shot is released above a horizontal plane, range from this point depends on release height, speed and angle. Measured distance is the sum of this range and horizontal distance of the release point from the throwing circle edge. Optimal release conditions can be calculated only if the dependence of release velocity on other variables, due to thrower limitations, is known. Experiments on two shot-putters investigated the hypothesis that there are constraint relationships among these four release parameters. A variable scaling scheme, using measurement of impact point and the known magnitude of g, corrected 2D data from one camera for out-of-plane motion and yielded accurate estimates of release parameters. Multivariate regression analyses determined approximate constraint surfaces limiting performance. Achievable release speed decreases with increasing release angle at about 1.7(m/s)/rad and decreases with increasing release height at about 0.8(m/s)/m, with only small differences in sensitivities between the throwers. Horizontal release distance also decreases with increasing release angle at about 1.7m/rad and increases with increasing release height at about 1.3m/m, again with only small differences between the two throwers. Optimal release conditions producing maximum range for a particular athlete can be determined using similar constraints for that athlete. PMID- 11266666 TI - Rotational moment arms of the medial hamstrings and adductors vary with femoral geometry and limb position: implications for the treatment of internally rotated gait. AB - Persons with cerebral palsy frequently walk with a crouched, internally rotated gait. Spastic medial hamstrings or adductors are presumed to contribute to excessive hip internal rotation in some patients; however, the capacity of these muscles to produce internal rotation has not been adequately investigated. The purpose of this study was to determine the hip rotation moment arms of the medial hamstrings and adductors in persons with femoral anteversion deformities who walk with a crouched, internally rotated gait. A musculoskeletal model with a "deformable" femur was developed. This model was used, in conjunction with kinematic data obtained from gait analysis, to calculate the muscle moment arms for combinations of joint angles and anteversion deformities exhibited by 21 subjects with cerebral palsy and excessive hip internal rotation. We found that the semimembranosus, semitendinosus, and gracilis muscles in our model had negligible or external rotation moment arms when the hip was internally rotated or the knee was flexed -- the body positions assumed by the subjects during walking. When the femur was excessively anteverted, the rotational moment arms of the adductor brevis, adductor longus, pectineus, and proximal compartments of the adductor magnus in our model shifted toward external rotation. These results suggest that neither the medial hamstrings nor the adductors are likely to contribute substantially to excessive internal rotation of the hip and that other causes of internal rotation should be considered when planning treatments for these patients. PMID- 11266668 TI - Trunk stiffness increases with steady-state effort. AB - Trunk stiffness was measured in healthy human subjects as a function of steady state preload efforts in different horizontal loading directions. Since muscle stiffness increases with increased muscle activation associated with increasing effort, it is believed that coactivation of muscles helps to stiffen and stabilize the trunk. This paper tested whether increased steady-state preload effort increases trunk stiffness. Fourteen young healthy subjects each stood in an apparatus with the pelvis immobilized. They were loaded horizontally at directions of 0, 45, 90, 135 and 180 degrees to the forward direction via a thoracic harness. Subjects first equilibrated with a steady-state load of 20 or 40% of their maximum extension effort. Then a sine-wave force perturbation of nominal amplitude of 7.5 or 15% of maximum effort and nominal period of 250ms was applied. Both the applied force and subsequent motion were recorded. Effective trunk mass and trunk-driving point stiffness were estimated by fitting the experimental data to a second-order differential equation of the trunk dynamic behavior. The mean effective trunk mass was 14.1kg (s.d.=4.7). The trunk-driving point stiffness increased on average 36.8% (from 14.5 to 19.8N/mm) with an increase in the nominal steady-state preload effort from 20 to 40% (F(1,13)=204.96, p<0.001). There was a smaller, but significant variation in trunk stiffness with loading direction. The measured increase in trunk stiffness probably results from increased muscle stiffness with increased muscle activation at higher steady-state efforts. PMID- 11266669 TI - Take-off aerodynamics in ski jumping. AB - The effect of aerodynamic forces on the force-time characteristics of the simulated ski jumping take-off was examined in a wind tunnel. Vertical and horizontal ground reaction forces were recorded with a force plate installed under the wind tunnel floor. The jumpers performed take-offs in non-wind conditions and in various wind conditions (21-33 m s(-1)). EMGs of the important take-off muscles were recorded from one jumper. The dramatic decrease in take-off time found in all jumpers can be considered as the result of the influence of aerodynamic lift. The loss in impulse due to the shorter force production time with the same take-off force is compensated with the increase in lift force, resulting in a higher vertical velocity (V(v)) than is expected from the conventional calculation of V(v) from the force impulse. The wind conditions emphasized the explosiveness of the ski jumping take-off. The aerodynamic lift and drag forces which characterize the aerodynamic quality of the initial take off position (static in-run position) varied widely even between the examined elite ski jumpers. According to the computer simulation these differences can decisively affect jumping distance. The proper utilization of the prevailing aerodynamic forces before and during take-off is a very important prerequisite for achieving a good flight position. PMID- 11266670 TI - A theoretical model of the effect of continuum damage on a bone adaptation model. AB - Throughout life, bone is continuously turning over by the well-regulated processes of bone formation and resorption. Everyday activities damage bone, and this damage is normally repaired in a continuous remodelling process. When an imbalance in this remodelling process occurs, bones may become more susceptible to fracture. This paper is devoted to a theoretical modelling of the competition between damage and internal remodelling in bones. The general theory of adaptive damaged-elastic materials proposed here as a model for the physiological process of damaged-bone remodelling follows the general framework of continuum thermodynamics where new damaged-bone remodelling law and associated thermodynamical restrictions are stated, and specialized to the case of small strain in isothermal processes. An attempt is also made to derive: (a) the damage force (adaptive damage energy release rate ) which controls the microcracks propagation and arrest, and (b) the damage rule by introducing damage thresholds and loading/unloading conditions. PMID- 11266671 TI - Mechanical energy analysis identifies compensatory strategies in disabled elders' gait. AB - Current concepts in disablement emphasize the importance of identifying mobility impairments in aging humans to enable timely intervention and, ultimately, prevent disability. Because mobility impairments are likely to result in compensatory movement strategies, recognizing and understanding those strategies may be critical in designing effective interventions for preventing disability. We sought to determine if mechanical energy methods are useful for identifying and understanding lower extremity compensatory movement strategies due to disabilities. Aleshinski's method was used to compute mechanical energy expenditure (MEE) and mechanical energy compensation (MEC) for the sagittal plane stance leg and low-back joints of healthy elders (HE) and disabled elders (DE) during preferred speed and paced (120 steps min(-1)) gait. DE subjects expended less ankle energy in late-stance and more low-back energy in mid-stance than did the HE subjects. When controlling for walking speed, the difference in ankle MEE disappeared, but mid-stance hip MEE was significantly higher for the DE subjects. Despite increased hip and low-back MEE, the DE subjects compensated hip and low back muscles greater then HE subjects by increasing energy transferred into the pelvis, particularly when walking faster than their self-selected speed. Increased energy transfers into the pelvis during mid-stance may be a strategy used to assist in advancing and controlling the contralateral limb's swing phase. Increased trunk energy, however, may compromise dynamic stability and increase the risk of falling. We conclude that mechanical energy methods are useful for identifying and understanding compensatory movement strategies in elders with disabilities. PMID- 11266672 TI - Analyses of myo-electrical silence of erectors spinae. AB - Electromyographic activity of the erector spinae was studied in 25 healthy, young individuals during forward bending and then coming back to erect posture. Sudden onset of electrical silence called the flexion-relaxation phenomenon was seen to occur in all at 57% of the maximum hip flexion and at 84% of the maximum vertebral flexion. Abrupt re-commencement of the activity was seen at almost similar flexion angle while coming back to erect position. The experiment was repeated with the buttocks held against the wall so as to prevent the posterior migration of the pelvis and also the hip flexion to some extent. The effect was to produce inhibition of the electrical activity earlier at 75% of maximum vertebral flexion (p<0.001) while reactivation of erector spinae occurred soon after the extension started from the maximum trunk flexion. Eleven male subjects repeated the experimental task holding 22 lb weight in front and then on their back tied around the iliac crest. In both the instances the myo-electrical silence was found to occur at greater vertebral flexion. It is concluded that the passive equilibrium between gravity induced tensile torque and the extension torque of stretched posterior vertebral ligaments is responsible for the flexion relaxation phenomenon than the stretch receptors. PMID- 11266673 TI - Age, sex, and grip strength determine architectural bone parameters assessed by peripheral quantitative computed tomography (pQCT) at the human radius. AB - The purpose of this study was to estimate the relation of some noninvasively derived mechanical characteristics of radial bone including architectural parameters for bone strength to grip strength and muscle cross-section. Sixty three males between 21 and 78yr of age and 101 females between 18 and 80yr of age were measured at the nondominant forearm using peripheral quantitative computed tomography (pQCT). We assessed the integral bone mineral density (BMD(I)) and content (BMC(I)) by pQCT at the distal and at the mid-shaft radius. Integral bone area (Area(I)), cortical thickness (C-th), and a newly proposed index for bone strength; the stress-strain index (SSI) were also calculated. The dynamometrically measured maximum grip strength was taken as a mechanical loading parameter and muscle cross-section as a substitute for it. Sex, grip strength, BMC(I) and BMD(I) (distal radius) were identified in a multiple regression analysis to significantly predict bone strength as expressed by SSI, after adjusting for all other independent variables, including age and sex (p<0.0001). Grip strength was closest related to age, sex, BMD(I) and SSI(p) of the distal radius. The cross-sectional area of muscle was not significantly determining the grip strength within the analysis model. In conclusion, our results suggested that architectural parameters at the distal radius were better related to grip strength than to cross-sectional muscle area in both males and females. Maximum muscle strength as estimated by grip strength might be a stronger determinant of mechanical characteristics of bones as compared with cross-sectional muscle area. PMID- 11266674 TI - Mechanical energetic processes during the giant swing before the Tkatchev exercise. AB - The goal of this study was to examine the possibility of the utilization of high bar and uneven parallel bar elasticity by the gymnasts through muscular work during the giant swing before the Tkatchev exercise on the high bar or uneven parallel bars. The performances were gathered during the Gymnastic World Championship in 1994. The set up consisted of two video cameras (50Hz) and two force measuring bars (500Hz). Twenty giant swings before the Tkatchev exercise, nine giant swings before the Tkatchev exercise after Tkatchev on the high bar and 15 giant swings before the Tkatchev exercise on the uneven parallel bars were analyzed. The giant swings were performed by 20 male and 15 female gymnasts. There are three phases during the giant swing exercise before the Tkatchev in which the systems (high bar-human body) total energy can be changed. During the first phase, energy is transferred from the gymnast's body into the bar. A clearly effective use of the bar's elasticity during the first phase could not be found. During the second phase, energy is transferred from the bar's back into the gymnast's body whose total energy increases. An increase in the energy of the system can only be achieved through muscular work. During the second phase of the various giant swing techniques no significant (p<0.05) difference in the energy increase through muscular work could be found. During the third phase, energy is once again produced by the gymnast through extension at the hip and shoulder joints. PMID- 11266675 TI - Introduction and evaluation of a gray-value voxel conversion technique. AB - In micro finite element analyses (microFEA) of cancellous bone, the 3D-imaging data that the FEA-models are based on, contain a range of gray-values. In the construction of the eventual FEA-model, these gray-values are commonly thresholded. Although thresholding is successful at small voxel sizes, at larger voxel sizes there is substantial loss of trabecular connectivity. We propose a new method: the gray-value method, where the microFEA-models use the information within the 3D-imaging data directly, without prior thresholding. Our question was twofold. First, how does the gray-value method compare to both plain and mass compensated thresholding? Second, what is the effect of element size on the results obtained with the gray-value method? We used nine microCT-scans of human vertebral cancellous bone. These were degraded to represent different resolutions, and converted into microFEA-models using plain thresholding, mass compensated thresholding, and the gray-value method. The apparent elastic moduli of the specimens were determined using microFEA. The different methods were compared on the basis of the apparent elastic moduli, compared to those calculated for a 28 microm reference model. The results showed that the gray value method greatly improves the results relative to other methods. The gray value method gives accurate predictions of the apparent elastic moduli, for voxel sizes up to one trabecular thickness (Tb.Th.). For voxel sizes greater than one Tb.Th. the accuracy, although still better than for both thresholding methods, becomes increasingly worse. PMID- 11266676 TI - A cross-validation of the biphasic poroviscoelastic model of articular cartilage in unconfined compression, indentation, and confined compression. AB - The biphasic poroviscoelastic (BPVE) model was curve fit to the simultaneous relaxation of reaction force and lateral displacement exhibited by articular cartilage in unconfined compression (n=18). Model predictions were also made for the relaxation observed in reaction force during indentation with a porous plane ended metal indenter (n=4), indentation with a nonporous plane ended metal indenter (n=4), and during confined compression (n=4). Each prediction was made using material parameters resulting from curve fits of the unconfined compression response of the same tissue. The BPVE model was able to account for both the reaction force and the lateral displacement during unconfined compression very well. Furthermore, model predictions for both indentation and confined compression also followed the experimental data well. These results provide substantial evidence for the efficacy of the biphasic poroviscoelastic model for articular cartilage, as no successful cross-validation of a model simulation has been demonstrated using other mathematical models. PMID- 11266677 TI - Biomechanical simulation of manual lifting using spacetime optimization. AB - Previous optimization techniques for the prediction of lifting motion patterns often require a change in either the number of variables or the order of the mathematical functions used to express the angular displacement of selected joints in response to change in variant conditions. The resolution of predicted results can also be seriously constrained by the number of variables used. These restrictions may often limit the applicability of these methodologies. In this paper, we proposed a new methodology for generating the optimum motion patterns for para-sagittal lifting tasks. A detailed description of this methodology is introduced. An example of an analysis using this methodology is presented. The computer program generated lifting motion patterns with a reduction of the overall objective function values. The actual versus predicted lifting motion patterns are compared. Using this method, constraints can be added anywhere within the lifting cycle without the need of rewriting the whole program. These features provide for a more flexible and efficient prediction of the lifting motion. PMID- 11266678 TI - A comparison of biomechanical properties between human and porcine cornea. AB - Due to the difficulty in obtaining human corneas, pig corneas are often substituted as models for cornea research. The purpose of this study is to find the similarities and differences in the biomechanical properties between human and porcine corneas. Uniaxial tests were conducted using an Instron apparatus to determine their tensile strength, stress-strain relationship, and stress relaxation properties. The tensile strength and stress-strain relation were very similar but significant differences between the two tissues were observed in the stress-relaxation relationship. Under the same stretch ratio lambda=1.5, porcine cornea relaxed much more than human cornea. If tensile strength and the stress strain relation are the only mechanical factors to be investigated, porcine cornea can be used as a substitute model for human cornea research. However, when stress relaxation is a factor, porcine corneas cannot be used as an appropriate model for human corneas in mechanical property studies. It is very difficult to get enough specimens of human cornea, so we did the experiments for stress-strain relationship at a specific value of strain rate (corresponding to the velocity of loading 10mm/min), and for stress relaxation at a specific stretch ratio lambda=1.5. PMID- 11266679 TI - Soft-tissue vibrations in the quadriceps measured with skin mounted transducers. AB - The purpose of this study was to develop a method to characterize the frequency and damping of vibrations in the soft tissues of the leg. Vibrations were measured from a surface-mounted accelerometer attached to the skin overlying the quadriceps muscles. The free vibrations in this soft tissue were recorded after impact whilst the muscle was performing isometric contractions at 0, 50, and 100% maximum voluntary force and with the knee held at 20, 40, and 60 degrees angles of flexion. The acceleration signals indicated that the soft tissue oscillated as under-damped vibrations. The frequency and damping coefficients for these vibrations were estimated from a model of sinusoidal oscillations with an exponential decay. This technique resolved the vibration coefficients to 2 and 7% of the mean values for frequency and damping, respectively. PMID- 11266680 TI - A minimally disruptive technique for measuring intervertebral disc pressure in vitro: application to the cervical spine. AB - A novel technique to measure in vitro disc pressures in human cervical spine specimens was developed. A miniature pressure transducer was used and an insertion technique was designed to minimise artefacts due to insertion. The technique was used to measure the intradiscal pressure in cervical spines loaded in pure axial compression. The resulting pressure varied linearly with the applied compressive force with coefficients of determination (r(2)) greater than 0.99 for each of the four specimens. Peak pressures between 2.4 and 3.5MPa were recorded under 800N of compression. PMID- 11266681 TI - A closed-loop cadaveric foot and ankle loading model. AB - Investigations of human foot and ankle biomechanics rely chiefly on cadaver experiments. The application of proper force magnitudes to the cadaver foot and ankle is essential to obtain valid biomechanical data. Data for external ground reaction forces are readily available from human motion analysis. However, determining appropriate forces for extrinsic foot and ankle muscles is more problematic. A common approach is the estimation of forces from muscle physiological cross-sectional areas and electromyographic data. We have developed a novel approach for loading the Achilles and posterior tibialis tendons that does not prescribe predetermined muscle forces. For our loading model, these muscle forces are determined experimentally using independent plantarflexion and inversion angle feedback control. The independent (input) parameters -- calcaneus plantarflexion, calcaneus inversion, ground reaction forces, and peroneus forces - are specified. The dependent (output) parameters -- Achilles force, posterior tibialis force, joint motion, and spring ligament strain -- are functions of the independent parameters and the kinematics of the foot and ankle. We have investigated the performance of our model for a single, clinically relevant event during the gait cycle. The instantaneous external forces and foot orientation determined from human subjects in a motion analysis laboratory were simulated in vitro using closed-loop feedback control. Compared to muscle force estimates based on physiological cross-sectional area data and EMG activity at 40% of the gait cycle, the posterior tibialis force and Achilles force required when using position feedback control were greater. PMID- 11266682 TI - Fine specificity of anti-GQ1b antibodies and clinical features. PMID- 11266683 TI - Multiple sclerosis: a white matter disease with associated gray matter damage. PMID- 11266684 TI - Fine specificity of anti-GQ1b IgG and clinical features. AB - Anti-GQ1b IgG frequently is present in sera of patients with Miller Fisher syndrome (MFS), Guillain-Barre syndrome (GBS) with ophthalmoplegia, Bickerstaff's brainstem encephalitis (BBE), and acute ophthalmoparesis (AO) in the acute phase. Why various clinical signs develop under these conditions, however, has yet to be clarified. We investigated the fine specificity of anti-GQ1b IgG and its clinical correlation in sera from 82 patients: 56 with MFS, 11 with GBS, 13 with BBE, and 2 with AO. Anti-GQ1b IgG antibodies were absorbed by GT1a in 80 (98%) of the 82 sera, by GD1b in 11 (13%), and by the other b-series gangliosides GD3, GD2, or GT1b in 24 (29%). The most frequent pattern of fine specificity was the cross reaction with GT1a alone, seen in 56 (68%) samples. Of the 11 patients with anti GQ1b IgG, cross-reacting with GD1b, 6 (55%) had impaired deep sense, and the association was significant (p=0.02). This is the first study to show that the fine specificity of anti-GQ1b IgG is heterogeneous and that the difference is correlated with the presence of a particular clinical sign. PMID- 11266686 TI - Gray matter T2 hypointensity is related to plaques and atrophy in the brains of multiple sclerosis patients. AB - Cortical and subcortical gray matter hypointensities on T2-weighted MR images (T2WI) occur commonly in MS brains and have been related to disease duration, clinical course, and the level of neurologic disability. These hypointensities have been reported to occur in the thalamus, basal ganglia, and rolandic cortex. We assessed whether T2 hypointensity is associated with the severity of white matter plaques and atrophy of MS brains. In 114 MS patients, hypointensity of the thalamus, putamen, caudate, and sensorimotor cortex was ordinally rated against age- and gender-matched normal controls on 1.5-T MRI fast spin-echo axial T2WI. Regional and global T2 hyperintense and T1 hypointense parenchymal lesion loads were ordinally rated. Enlargement of subarachnoid and ventricular spaces (atrophy) was ordinally rated vs. age- and gender-matched normal controls. T2 hypointensity was highly, positively correlated with many other MRI variables. Regression modeling showed that T2 hypointensity was related to total atrophy, total T2 lesion load, third ventricular enlargement, parietal atrophy, and to a lesser extent, frontal T1 lesions and cerebellar T2 lesions, but not related to gadolinium enhancement. Ordinal ratings of T2 lesions and central atrophy showed high correlations with quantitative computerized assessments. We conclude that gray matter hypointensity on T2WI may reflect pathologic iron deposition and brain degeneration in MS. This T2 hypointensity is associated with brain atrophy and other MR markers of tissue damage. Further study is warranted to determine if T2 hypointensity is predictive of disease course in MS and is a useful surrogate outcome measure in therapeutic trials. PMID- 11266685 TI - Magnetisation transfer ratio histogram analysis of primary progressive and other multiple sclerosis subgroups. AB - INTRODUCTION: Global magnetisation transfer ration (MTR) histogram analysis in the brain offers a method for evaluating pathological change both as a result of lesions and microscopic changes in normal appearing tissues. METHODS: 39 controls and 83 MS patients (46 primary progressive, 11 benign, 10 relapsing-remitting, 16 secondary progressive) were studied to explore the relationship of six conventional MTR histogram parameters with MS clinical subgroups and disability. Principal component (PC) analysis, which makes use of all the histogram data, was also used to examine the relationship between the MTR histogram and disability. RESULTS: When primary progressive patients were compared to controls, there were abnormalities of average MTR, and MTR at the 25th, 50th and 75th percentile. Disabled relapsing onset patients exhibited abnormalities in the same four parameters. Benign and nondisabled relapsing onset patients exhibited no significant abnormalities. Modest correlations were observed between disability and individual MTR parameters in relapse onset but not primary progressive patients--PC analysis revealed stronger and significant associations with disability in both subgroups. (r=0.40 for primary progressive and r=0.51 for relapsing onset). CONCLUSION: A number of MTR parameters are abnormal in primary progressive MS. MTR abnormalities are seen in disabled patients, whether of relapsing or primary progressive onset. The improved correlation with disability obtained by PC analysis suggests a useful role of this method for following clinically relevant pathological changes depicted in the MTR histogram. PMID- 11266687 TI - Cerebral venous thrombosis associated with tentorial subdural hematoma during oxymetholone therapy. AB - Androgen was reported to cause cerebral venous thrombosis (CVT) during replacement therapy for aplastic anemia. Oxymetholone, a synthetic androgen analogue, has been widely used in the treatment of aplastic anemia. A 40-year-old woman with aplastic anemia visited our hospital because of severe headache, nausea, vomiting, blurred vision and diplopia for a period of 1 month. She had taken oxymetholone for 2 years. Neurological examination revealed bilateral papilledema and bilateral sixth nerve palsies. Brain magnetic resonance imaging (MRI), performed at the time of admission, demonstrated left-sided tentorial SDH, and focal cerebral thrombosis of the left superficial sylvian vein and sigmoid sinus. MR venography revealed multiple irregularities in the superior sagittal sinus and left transverse sinus. CVT with tentorial subdural hematoma (SDH) caused by oxymetholone was strongly suggested. Oxymetholone was immediately discontinued, and her symptoms and signs disappeared. Because of the thrombocytopenia, anticoagulation was not started. She was discharged and visited the outpatient clinic without neurological symptoms for 6 months. This report supports the cautions given about the risk of CVT with oxymetholone supplementation in aplastic anemia. To the best of our knowledge, this is the first report of CVT associated with tentorial SDH that was probably caused by oxymetholone. PMID- 11266688 TI - Clinical phenotype in X-linked Charcot-Marie-Tooth disease with an entire deletion of the connexin 32 coding sequence. AB - To clarify the clinical phenotype and molecular mechanism in X-linked Charcot Marie-Tooth disease (CMTX) patients with a deletion of the whole connexin 32 (Cx32) coding sequence, we studied a family with this deletion by electrophysiology, Southern blotting and quantitative PCR analyses. Two brothers with no copy of Cx32, 27 and 25 years old, showed steppage gait, moderate muscle atrophy and weakness, and mild sensory disturbance in the distal parts of the legs. The clinical phenotypes in these brothers were not different from those in patients with other types of severe Cx32 mutations. Their mother, with one copy of Cx32, showed very mild muscle weakness and sensory disturbance. An electrophysiological study showed a nonuniform demyelinating neuropathy with some aspects of an axonal-loss neuropathy. Sural nerve biopsy showed loss of myelinated fibers, many relatively thin myelin sheaths, clusters of small myelinated fibers, and some onion bulb formations. The present findings suggest that both a demyelinating process and an axonal involvement were present in the patients with total defect of Cx32 probably due to loss of the function mechanism of Cx32 as the underlying molecular mechanism, because a dominant negative effect theory is not applicable in these patients. PMID- 11266690 TI - Spinocerebellar ataxia type 6: founder effect in Western Japan. AB - An accumulation of SCA6 cases has been observed in the Chugoku area of Western Japan. In the Tottori prefecture, located in the northeastern part of the Chugoku district, we observed a cluster of SCA6 families within the eastern area, suggesting that there may be a founder in the Japanese SCA6 population. Genotyping with DNA microsatellite markers linked to the CACNL1A4 gene on chromosome 19p13 demonstrated shared allelic characteristics and revealed a common haplotype in the majority of Japanese families. The common haplotype of the shared (CAG)(22) repeat found in this study may indicate the meiotic stability of CAG repeats in SCA6 patients. PMID- 11266689 TI - Clinical and physiological significance of abnormally prolonged central motor conduction time in HAM/TSP. AB - We measured the central motor conduction time (CMCT), central sensory conduction time (CSCT), F wave and mean F wave/M wave amplitude ratio in patients with HTLV I associated myelopathy/tropical spastic paraparesis (HAM/TSP) and controls. CMCTs in upper (U) and lower (L) limbs were significantly prolonged in HAM/TSP. L CSCT was significantly prolonged in HAM/TSP, but U-CSCT in HAM/TSP and controls were not significantly different. CMCT and CSCT were significantly correlated in HAM/TSP. U-CMCT, but not L-CMCT, correlated with the clinical severity of HAM/TSP. Although F wave conduction velocity and its occurrence were normal in HAM/TSP, U- and L-mean F wave/M wave amplitude ratio tended to be higher in HAM/TSP, and the L-mean F wave/M wave amplitude ratio was significantly correlated with the L- and thoracic CMCT. These findings demonstrate that the prolongation of CMCT sensitively reflects the extension of the lesions and the disinhibition to the anterior horn cells in HAM/TSP. PMID- 11266691 TI - Influence of physiologic oscillation of estrogens on cerebral hemodynamics. AB - The interactions between estrogens and the cerebrovascular system are complex and not fully understood. There are evidences suggesting that the hormones confer protection against cerebral ischemia. Our aim in this study was to investigate the effects of physiological variations of estradiol plasmatic concentration on cerebral hemodynamics. We investigated cerebrovascular reactivity to hypercapnia with transcranial Doppler ultrasonography and the breath-holding index method in the right middle cerebral artery of 20 young women during the menstrual and the ovulatory phase. Data were compared with those of 20 men matched for age. The mean value of the breath-holding index was significantly higher (p<0.001) in females during the ovulatory phase than in the menstrual phase. In men, values were similar to those of women during the menstrual phase. These results suggest that estrogens influence the adaptation capacity of the cerebrovascular system. The possible pathophysiological implications of the relationships between sex hormones and cerebral hemodynamics deserve further investigation. PMID- 11266692 TI - Damage to intracranial optic pathways in fatal closed head injury in man. AB - Head injury is a leading cause of visual impairment. This is partly due to direct trauma to the eye and optic nerve but much of the damage involves the intracranial optic pathways. We have studied the frequency, distribution and nature of the intracranial lesions of the optic pathways at autopsy in 45 cases of severe closed head injury, and examined the correlation between these post mortem lesions and the ante-mortem clinical findings. Twenty-four of the patients had been involved in road traffic accidents. The ages ranged from 9 to 88 years (mean 46.4), the Glasgow Coma Score (GCS) on admission ranged from 3 to 15 (mean 5), and the survival time after injury from 2.5 h to15 days (mean 3.3 days). Skull fractures were present in 75.6% of the cases. Histological assessment included the use of immunohistochemistry for beta-amyloid precursor protein (beta APP) and the microglial marker CD68. Axonal injury of varying severity was demonstrable in all cases, and in 39 (87%) the optic chiasm, tracts or radiations were involved, usually in more than one region. The severity of axonal injury was mild in 11 (24%), moderate in 9 (20%) and severe in 19 (42%) cases. The optic radiation at the level of the trigone of the lateral ventricle was particularly frequently and severely affected. The least affected parts of the intracranial optic pathways were the optic chiasm and the posterior segment of the optic nerve. The severity of injury to the optic pathways did not always reflect severity of axonal injury elsewhere in the brain and correlated poorly with the type of trauma (high- or low-velocity), presence of skull fractures or evidence of raised intracranial pressure (ICP). Of the 39 patients who survived more than 6 h, histological evidence of ischaemic injury to the primary optic cortex was present in 26 (67%) and was severe in 12. We conclude that the visual pathways are affected in a high proportion of patients with fatal closed head injury, nerve fibres in the optic radiations being particularly vulnerable. The findings suggest that damage to the posterior parts of the optic pathways may be under diagnosed among patients with head injury. PMID- 11266699 TI - Special address by His Holiness John Paul II, Rome, August 29, 2000. PMID- 11266700 TI - Presidential address. PMID- 11266693 TI - A large Japanese SPG4 family with a novel insertion mutation of the SPG4 gene: a clinical and genetic study. AB - We studied a large Japanese family with autosomal dominant pure hereditary spastic paraplegia (ADPHSP) clinically and genetically. To date, seven loci causing ADPHSP have been mapped to chromosomes 14q, 2p, 15q, 8q, 12q, 2q, and 19q. Among these loci, the SPG4 locus on chromosome 2p21--p22 has been shown to account for approximately 40% of all autosomal dominant hereditary spastic paraplegia (ADHSP) families. Very recently, Hazan et al. identified the SPG4 gene encoding a new member of the AAA (ATPases associated with diverse cellular activities) protein family, named spastin. We found a novel insertion mutation (nt1272--1273insA) in exon 8 of the SPG4 gene in the present family. Our study is the first to confirm the causative mutation of the SPG4 gene in Japanese. Clinically, it is noteworthy that the disease progression in the patients of this family was slow in spite of the late onset, and more than half of the patients showed severe constipation in addition to pure spastic paraplegia. PMID- 11266702 TI - Transplantation in the new millenium. PMID- 11266703 TI - Strategies for induction of clinical tolerance. PMID- 11266705 TI - Induction of transplantation tolerance by 1,25-dihydroxyvitamin D(3). PMID- 11266704 TI - Recombinant antigens to establish a model of autoimmunity in mice. PMID- 11266706 TI - Immunobiology of human NK cells. PMID- 11266707 TI - Hepatocyte transplantation: an experimental study to treat acute liver failure in pigs. PMID- 11266708 TI - Engineered vessels: importance of the extracellular matrix. PMID- 11266709 TI - Megadose approach to DBMC infusion-induced allograft hyporesponsiveness in living related renal allograft recipients. PMID- 11266710 TI - Induction of kidney allograft tolerance through mixed chimerism in miniature swine. PMID- 11266711 TI - Organ allograft recipients develop HLA class I-specific T suppressor cells. PMID- 11266712 TI - Intrathymic immune regulation with donor class I allopeptides leads to the development of immunoregulatory cells that maintain tolerance to cardiac allografts. PMID- 11266713 TI - Oral antigen induces allograft survival by linked suppression via the indirect pathway. PMID- 11266714 TI - CD8(+)CD28(-) T cells suppress alloresponse of CD4(+) T cells both in primates and rodents. PMID- 11266715 TI - Suppression of alloimmune responses in vitro and in vivo by CD3(+)CD8(-)CD4( )alphabeta(+) regulatory T cells. PMID- 11266716 TI - Intratracheal delivery of alloantigen induces donor-specific hyporesponsiveness: dendritic cells play a pivotal role as regulatory cells. PMID- 11266717 TI - Specific suppression of proliferative responses by primed lymphocytes of tolerant miniature swine. PMID- 11266718 TI - CTLA4Ig-induced linked regulation of allogeneic T cell responses. PMID- 11266719 TI - TCR transgenic regulatory T cells prevent lethal graft-versus-host disease. PMID- 11266720 TI - Mechanisms of acquired tolerance induced by adoptive transfer of MHC-specific alloreactive T cells: effector T cells migrate to the thymus. PMID- 11266721 TI - Autoimmune responses regulate alloimmunity for prolonged cardiac allograft survival. PMID- 11266722 TI - CD8alpha(+) (lymphoid-related) and CD8alpha(-) (myeloid) dendritic cells differentially regulate vascularized organ allograft survival. PMID- 11266723 TI - Tyrphostin AG490 selectively inhibits activation of the JAK3/STAT5/MAPK pathway and rejection of rat heart allografts. PMID- 11266724 TI - Thymic recognition of AlloMHC peptide-primed alloreactive T cells induces specific unresponsiveness to islets. PMID- 11266725 TI - Induction of acquired tolerance to cardiac allografts by adoptive transfer of in vivo allopeptide activated T cells. PMID- 11266726 TI - In utero induction of transplantation tolerance. PMID- 11266727 TI - Tolerance across a two-haplotype, fully MHC-mismatched barrier induced in miniature swine renal allografts treated with a 12-day course of tacrolimus. PMID- 11266728 TI - Liver transplants induce deletion of liver-specific T cells. PMID- 11266729 TI - CD8(+)CD28(-) T suppressor cells represent a distinct subset in a heterogeneous population. PMID- 11266730 TI - Donor bone marrow cells used for the induction of allograft acceptance in clinical transplantation as regulators of autologous immune responses. PMID- 11266731 TI - Intrathymic donor stem cell fractions increase chimerism but do not check alloantibody or alloreactivity responses in nonhuman primates. PMID- 11266732 TI - Essential role of MHC class II antigens in tolerance induction in allogeneic radiation chimera. PMID- 11266733 TI - Inhibition of in vitro donor-specific proliferative and cytotoxic T cell responses in chimeric CD40 ligand-deficient bone marrow transplant recipients treated perioperatively with CTLA4-Ig. PMID- 11266735 TI - Comparison of horse antithymocyte globulin with other T cell-depleting antibodies for induction of chimerism and renal allograft tolerance in nonhuman primates. PMID- 11266734 TI - Blockade of the CD28/B7 pathway is required for the establishment of donor cell chimerism in CD40 ligand-deficient recipients. PMID- 11266737 TI - Mixed bone marrow chimerism as a treatment for autoimmune diabetes. PMID- 11266736 TI - Mixed chimerism using a nonmyelosuppressive regimen in miniature swine. PMID- 11266738 TI - Heart allograft tolerance without development of posttransplant cardiac allograft vasculopathy in cyclophosphamide-induced tolerance. PMID- 11266739 TI - Cells of donor phenotype present in renal transplant recipients infused with donor marrow: potent regulators of antidonor immune responses of the recipient. PMID- 11266740 TI - Blockade of tryptophan catabolism prevents spontaneous tolerogenicity of liver allografts. PMID- 11266741 TI - Tolerance induction by alteration of T cell receptor (TCR) signaling using an allochimeric donor/recipient class I MHC protein. PMID- 11266742 TI - Modulation of primary T cell responses and tolerance induction by tyrphostin AG490. PMID- 11266743 TI - Mechanism of adenoviral-mediated CTLA4-IG gene-induced pancreatic allograft tolerance in rats. PMID- 11266744 TI - Donor cell migration and graft-infiltrating cell apoptosis in heart acceptance in a multiorgan transplant model. PMID- 11266745 TI - Mechanisms underlying the immunosuppressive activity of antitransferrin receptor monoclonal antibody. PMID- 11266746 TI - Graft acceptance and tolerance induction in mouse liver transplantation using wild mice. PMID- 11266747 TI - The role of CD30 in skin and heart allograft rejection in the mouse. PMID- 11266748 TI - Features of tolerance achieved by antigen and a single injection of an anti-CD45 monoclonal antibody in rats. PMID- 11266749 TI - Portal venous tolerance in the anti-class I L(d) alloantigen 2C transgenic mouse. PMID- 11266750 TI - Nihility: discovery of the immunologic zero corner where self-tolerance and clonal deletion are challenged. PMID- 11266751 TI - Expression of MHC class II antigen is essential in tolerance induction by donor bone marrow cell in antilymphocyte serum-treated and rapamycin-treated mice. PMID- 11266752 TI - Regulatory role of the thymic dendritic cells in acquired thymic tolerance: induction of tolerance to cardiac allografts by adoptive transfer of allopeptide pulsed host thymic dendritic cells. PMID- 11266753 TI - Donor-specific antigen transfusion-mediated cardiac allograft tolerance is prevented by prior treatment with anti-CD8, but not anti-CD4, antibody. PMID- 11266755 TI - The role of T-cell anergy in the maintenance of donor-specific hyporesponsiveness in renal transplant recipients. PMID- 11266754 TI - Mechanisms of donor-specific tolerance in a skin transplantation model. PMID- 11266756 TI - Proteome analysis in liver transplantation. PMID- 11266757 TI - Induction of long-term islet allograft survival in two rat strains by a single allochimeric class I MHC molecule. PMID- 11266758 TI - Intrathymic alloantigen-mediated tolerant, completely MHC-mismatched mouse hearts are specifically rejected by adoptively transferred in vitro-sensitized anti class I L(d+)-specific 2C cells. PMID- 11266759 TI - Putative regulatory function of alloreactive Th2 cells in tolerant recipients after small bowel transplantation. PMID- 11266760 TI - CD4(+), CD25(+) T cells as regulators of alloimmune responses. PMID- 11266761 TI - Specific T-cell deletion by transfected human monocytes expressing Fas ligand and antigen. PMID- 11266762 TI - The role of noninherited HLA haplotypes in inducing donor-specific hyporesponsiveness. PMID- 11266764 TI - Third-party passenger leukocytes prolong liver allograft survival. PMID- 11266763 TI - Tolerance induction in indirect alloresponses by analogs of HLA-derived peptides. PMID- 11266765 TI - Different role of cyclophosphamide-induced tolerance in heart and skin allograft tolerance. PMID- 11266766 TI - MHC class II compatibility between donor bone marrow and secondary organ grafts is required for tolerance based on mixed chimerism. PMID- 11266767 TI - The study of peripheral blood microchimerism in kidney transplantation. PMID- 11266768 TI - Allochimeric class I MHC inhibits chronic rejection in cardiac allografts. PMID- 11266769 TI - Dendritic cell TH2 cytokine gene therapy in sheep. PMID- 11266770 TI - A role for donor-derived B cells but not T cells in the maintenance of mixed hematopoietic chimerism. PMID- 11266771 TI - Adenovirus-mediated CTLA4-IgG gene therapy in orthotopic small intestinal transplantation in rats. PMID- 11266772 TI - Cellular basis of long-term rat liver allograft acceptance: role of donor and recipient leukocyte persistence in rat liver grafts. PMID- 11266773 TI - Induction of peripheral tolerance by posttransplant infusion of donor splenocytes. PMID- 11266774 TI - Novel CD40-IgG adenovirus-mediated gene therapy as a potent immunosuppressive treatment for liver transplantation in rats. PMID- 11266775 TI - Suppression of T-cell alloreactivity by gene-therapeutic modulation of human dendritic stimulator cells with TGF-beta adenoviral vectors. PMID- 11266776 TI - Conditioned dendritic cells as a temporal bridge between T helper and cytotoxic cells. PMID- 11266777 TI - Selective inhibition of NF-AT DNA binding by the short-chain fatty acid n butyrate. PMID- 11266778 TI - Targeting additional costimulatory pathways: a subtle role for CD2. PMID- 11266779 TI - Inhibition of Jak3 tyrosine kinase by PNU156804 blocks rat heart allograft rejection. PMID- 11266780 TI - The role of CD154 (CD40-ligand) in costimulation. PMID- 11266781 TI - The role of ezrin in T-cell receptor-dependent signaling. PMID- 11266782 TI - Transplantation and the CD28/CTLA4/B7 pathway. PMID- 11266783 TI - Intracellular signaling consequences of anti-IL-2Ralpha blockade by daclizumab. PMID- 11266784 TI - Role of STAT6 signaling in the induction and long-term maintenance of tolerance mediated by CTLA4-Ig. PMID- 11266786 TI - Transplantation tolerance by 1,25-dihydroxyvitamin D(3)-induced costimulation blockade. PMID- 11266785 TI - The effect of OX40/OX40L and CD27/CD70 pathways on allogeneic islet graft rejection. PMID- 11266788 TI - Effects of dose and duration of anti-CD154 antibody therapy in preventing renal allograft rejection in a nonhuman primate model. PMID- 11266787 TI - Costimulatory blockade for induction of mixed chimerism and renal allograft tolerance in nonhuman primates. PMID- 11266789 TI - Overexpression of A20 in endothelial cells of vascularized grafts creates a protective barrier against TNF- and FAS-mediated apoptosis. PMID- 11266790 TI - Hepatocyte-induced apoptosis of activated T cells, a mechanism of liver transplant tolerance, is related to the expression of ICAM-1 and hepatic lectin. PMID- 11266791 TI - Increased Fas ligand expression and activated T-lymphocyte apoptosis in islet graft tolerance induced by liver transplantation. PMID- 11266792 TI - A novel subset of dendritic cells propagated from the liver promotes differentiation of T regulatory cells and enhances allograft survival. PMID- 11266793 TI - Immaturity of donor dendritic cells regulates recipient T-cells toward Th2 deviation. PMID- 11266794 TI - Diltiazem affects human dendritic cell maturation. PMID- 11266795 TI - Fas ligand-transfected dendritic cells induce apoptosis of antigen-specific T cells. PMID- 11266796 TI - Phenotype and allostimulatory function of dendritic cells treated with antisense oligodeoxyribonucleotides targeting CD80 or CD86 mRNA. PMID- 11266797 TI - Diltiazem downregulates IL-12 production by human dendritic cells. PMID- 11266798 TI - Immature dendritic cells induce alternative CD4(+) T-cell responses in vivo. PMID- 11266799 TI - Effect of blockade of the costimulation pathway by anti-B7 antibodies in renal allotransplantation in baboons. PMID- 11266800 TI - Nonviral gene gun-mediated CTLA4-Ig gene transfer for modification of donor organs. PMID- 11266802 TI - Prolonged survival of allogeneic islet grafts in NOD mice treated with a combination of anti-CD45RB and anti-CD154 antibodies. PMID- 11266801 TI - CTLA4-Ig inhibits rejection of mouse liver allografts from Flt3-ligand-treated donors and is associated with increased lymphocyte apoptosis. PMID- 11266803 TI - Differential regulation of T-cell proliferation and apoptosis by hepatic CD8alpha(+) (lymphoid-related) and CD8alpha(-) (myeloid) dendritic cells. PMID- 11266804 TI - Altered expression of Bcl-x and Bax in cardiac allograft arteries of primates. PMID- 11266805 TI - CMV accelerates tubular apoptosis in a model of chronic renal allograft rejection. PMID- 11266806 TI - Survival- and apoptosis-inducing genes of the BCL-2 gene family expressed in urine lymphocytes to monitor renal transplant function. PMID- 11266808 TI - Expression of FasL as a stable cell-surface molecule is important to its apoptotic function. PMID- 11266807 TI - Early T-cell inactivation and apoptosis-critical events for tolerance induction after allogeneic liver transplantation. PMID- 11266809 TI - JNK(1)/SAPK(1) involvement in hypoxia-reoxygenation-induced apoptosis in rat hepatocytes. PMID- 11266810 TI - Target cells expressing CD95L are protected from lysis mediated by alloactivated T cells but are killed by resting NK cells. PMID- 11266811 TI - Early assessment of apoptosis in isolated islets of Langerhans. PMID- 11266812 TI - HO-1 upregulation protects the pancreatic cell line betaTC3 from cytokines and Fas-induced apoptosis. PMID- 11266813 TI - Transfection, but not retroviral transduction, upregulates apoptotic pathways in murine fibroblasts. PMID- 11266814 TI - Migration of donor MHC class II(+) cells and increase in apoptosis: correlate to graft outcome after heart and liver transplantation. PMID- 11266815 TI - Donor-derived costimulatory molecule-deficient dendritic cells impair the allostimulatory function of recipients' antigen-presenting cells. PMID- 11266816 TI - Prolonged allograft survival following ex vivo blockade of costimulatory signals B7-1/2 and CD40. PMID- 11266817 TI - Cyclosporin induces apoptosis of renal cells by enhancing nitric oxide synthesis: modulating effect of angiotensin II inhibitors. PMID- 11266819 TI - Role of Fas/Fasl in Kupffer cell-dependent deletion of alloantigen activated T cells following liver transplantation. PMID- 11266818 TI - Pretreatment of HL60 cells with Deoxyspergualin enhances trail-induced apoptosis independent of caspase 3 activation. PMID- 11266820 TI - Role of Fas-Fas ligand interaction in donor-specific transfusion-induced tolerance to H-Y antigen. PMID- 11266821 TI - HIV-1 Nef protein prolonged recipient survival in rat liver allografting. PMID- 11266822 TI - Anti-CTLA-4 human scFv antibodies prevent T-cell activation in transplantation. PMID- 11266823 TI - Prediction of rejection in lung transplantation. PMID- 11266825 TI - Early biomarkers for late graft loss. PMID- 11266826 TI - Genomic analysis of renal allograft dysfunction using cDNA microarrays. PMID- 11266824 TI - Molecular biology of transplant arteriosclerosis and sites of therapeutic intervention. PMID- 11266827 TI - Pathophysiology of the progression of renal graft dysfunction. PMID- 11266828 TI - Immunologic monitoring of T helper cell reactivity. PMID- 11266829 TI - CD40 ligand-CD40 T cell costimulation pathway is required for host sensitization in the immune cascade leading to accelerated allograft rejection. PMID- 11266830 TI - Specific suppression of interleukin-2 biosynthesis by synthetic antisense oligodeoxynucleotides does not influence allograft rejection. PMID- 11266831 TI - Presentation of MHC-disparate donor antigens predominantly by the indirect pathway results in the development of posttransplant vasculopathy: salutary effects of perioperative costimulatory blockade. PMID- 11266833 TI - Anti-HLA class I antibodies transduce signals in endothelial cells resulting in FGF receptor translocation, down-regulation of ICAM-1 and cell proliferation. PMID- 11266832 TI - Indirect allorecognition promotes the development of cardiac allograft vasculopathy. PMID- 11266834 TI - T cell receptor engagement can influence T cell motility. PMID- 11266835 TI - The BH4 domain of A1, an anti-apoptotic bcl family gene, is necessary and sufficient for its antiinflammatory function in endothelial cells. PMID- 11266836 TI - Antigen presentation by murine endothelial cells. PMID- 11266837 TI - Alloantibodies restore cardiac allograft rejection to IgKO mice. PMID- 11266839 TI - Role of MHC class I and CD8(+) T cells in the pathogenesis of chronic rejection. PMID- 11266838 TI - PDGF receptor inhibition prevents cardiac allograft arteriosclerosis in cholesterol-fed rabbits. PMID- 11266840 TI - Comparison of the effects of exposure to a single or multiple donor alloantigens in the development of transplant arteriosclerosis. PMID- 11266841 TI - Donor hypertension and recipient immune responsiveness in chronic rat cardiac allograft rejection. PMID- 11266842 TI - The persistence of transplant arteriosclerosis despite CD154 blockade. PMID- 11266843 TI - Development of transplant arteriosclerosis after allogeneic aorta transplantation in the rat: influence of recipient genotype. PMID- 11266844 TI - CFSE dye dilution mixed lymphocyte reactions quantify donor-specific alloreactive precursors in non-human primate cardiac graft rejection. PMID- 11266845 TI - Selective inhibition of IL-2 mRNA blocks allograft rejection by limiting T cell clonal expansion. PMID- 11266847 TI - FcgammaRllc 13Q/STP polymorphism influences the antibody-dependent cytotoxicity levels triggered by natural killer cells against pig aortic endothelial cells. PMID- 11266846 TI - Cyclosporin resistant allo-activated natural killer cells: possible evidence of functional natural killer memory. PMID- 11266848 TI - Proliferative responses to allogeneic HLA class I peptides. PMID- 11266849 TI - The graft-versus-leukemia effect in allogeneic irradiation bone marrow chimeras: possible suppressive role of irradiation-induced TGF-beta. PMID- 11266850 TI - Delayed type hypersensitivity in the CD8+ dominant anti-class I L(d) alloantigen 2C transgenic mouse is mediated by small but significant CD4 population. PMID- 11266852 TI - TGF-beta(1) gene expression in protocol biopsies from patients with stable renal allograft function. PMID- 11266851 TI - Changes of allo-intestinal graft survival and intragraft cytokine expression by bone marrow augmentation with tacrolimus treatment in rats. PMID- 11266854 TI - Inhibition of tumor necrosis factor-alpha does not prevent cardiac allograft arteriosclerosis in the rat. PMID- 11266853 TI - CD80/86 and Th1 cytokine expression in intestinal graft following reperfusion and endotoxemia. PMID- 11266855 TI - TNF-alpha IFN-gamma IL-6, IL-10, and TGF-beta1 gene polymorphisms in renal allografts. PMID- 11266856 TI - TNF-alpha and IL-6 gene polymorphism and rejection in kidney transplantation recipients. PMID- 11266857 TI - Bronchial epithelial cell-derived cytokine IL-10 and lung fibroblast proliferation. PMID- 11266858 TI - IL-10 transduction of liver allografts induces antiinflammatory monocytes in long term-surviving hosts. PMID- 11266859 TI - Absence of M-CSF-dependent tissue macrophages does not improve delayed function of islet of Langerhans grafts. PMID- 11266860 TI - Obliterative bronchiolitis develops in miniature swine transplanted across a minor histocompatibility barrier. PMID- 11266861 TI - Antisense NF-kappaB p65 prolongs experimental allo- and xenograft survival. PMID- 11266862 TI - Prolonged ischemia enhances acute rejection in rat kidney grafts. PMID- 11266863 TI - Effect of treatment with low-molecular-weight heparin on chronic renal allograft rejection in rats. PMID- 11266864 TI - Altered alloimmune response toward grafts from brain-dead donors in chronic rat cardiac allograft rejection. PMID- 11266867 TI - A model of vascular thymus transplantation in athymic rats. PMID- 11266865 TI - Leflunomide analog, MNA-715, plus cyclosporine reduces renal allograft rejection in mismatched dogs. PMID- 11266866 TI - Terminal complement proteins in mediating processes of accelerated graft arteriosclerosis in cardiac transplants. PMID- 11266868 TI - Addition of steroids blocks the tolerogenic potential of donor-specific blood transfusion. PMID- 11266869 TI - Induction of platelet-derived growth factor and its receptors in acute renal allograft rejection. PMID- 11266870 TI - Cytomegalovirus infection enhances connective tissue growth factor mRNA expression in a rat model of chronic renal allograft rejection. PMID- 11266871 TI - Diagnosis of acute rejection by analysis of urinary DNA of donor origin in renal transplant recipients. PMID- 11266872 TI - Blockade of the CD28/B7 and CD40/CD40L costimulatory pathways does not ameliorate chronic rejection in a mouse aortic allograft model of direct antigen presentation. PMID- 11266873 TI - Alternatively spliced variants of IL-2 mRNA in sequential transplant kidney core needle biopsies. PMID- 11266874 TI - Development of an oral formulation for ICAM-1 antisense oligonucleotides. PMID- 11266875 TI - Microgravity culture conditions decrease immunogenicity but maintain excellent morphology of pancreatic islets. PMID- 11266876 TI - MHC-independent allograft vascular disease: mRNA profile in the MHC congenic rat heterotopic cardiac transplant model. PMID- 11266877 TI - Improving screening strategies for HLA-specific antibody detection in renal transplant recipients. PMID- 11266878 TI - Sensitization to HLA by allografts is minimized by matching for HLA-A,B crossreactive groups. PMID- 11266879 TI - Donor specific antibody suppression in ABO incompatible kidney transplantation. PMID- 11266881 TI - Addition of rapamycin to cyclosporine-prednisone based immunosuppression reduces the clinical risk of pretransplant anti-HLA antibody. PMID- 11266880 TI - Influence of CsA treatment on adoptive transfer of immunity after allogeneic kidney transplantation in rats. PMID- 11266882 TI - Posttransplant immune monitoring of anti-HLA antibody. PMID- 11266883 TI - Solid-phase HLA antibody detection methods and risk of renal allograft rejection in children. PMID- 11266884 TI - Murine model of accelerated graft arteriosclerosis in primed recipients. PMID- 11266885 TI - Endothelin-1 is biologically active in experimental rat obliterative bronchiolitis. PMID- 11266886 TI - Function of CD40/CD40L in peritubular capillary of renal allograft rejection. PMID- 11266887 TI - An association between fibrinolytic activity and graft function in kidney transplantation recipients. PMID- 11266888 TI - Endothelial cell activation by tumor necrosis factor elicits human antiporcine cell-mediated rejection responses. PMID- 11266889 TI - New aspects of RANTES in context of liver transplantation. PMID- 11266890 TI - Interleukin 12 delays allograft rejection: effect mediated via nitric oxide. PMID- 11266891 TI - Blockade of the CD40 pathway fails to prevent CD8 T cell-mediated intestinal allograft rejection. PMID- 11266892 TI - Participation of IL-18 in human cholestatic cirrhosis and acute rejection: analysis in living donor liver transplantation. PMID- 11266893 TI - Detection of allospecific T cells by intracellular cytokine staining and flowcytometry. PMID- 11266894 TI - T cell proliferation is blocked by indoleamine 2,3-dioxygenase. PMID- 11266896 TI - Signaling via the CD28 molecule plays a particular role in the activation of human CD4(+)CD45R0(+) T cells. PMID- 11266895 TI - Acceptance of small bowel allografts by indirect allorecognition of donor class II MHC allopeptides. PMID- 11266897 TI - Indirect alloreactivity and cytokine production to HLA-DR peptides in human renal transplantation. PMID- 11266898 TI - The effect of alpha-galactosylceramide upon allogenic rejection. PMID- 11266899 TI - Human NK cells lyse porcine endothelial cells via the perforin/granzyme B pathway. PMID- 11266900 TI - Use of autologous dendritic cells loaded with apoptotic LCL for ex vivo generation of specific CTL from the PBMC of EBV(-) individuals. PMID- 11266901 TI - Detection of dendritic cells with down-regulated CD80/CD86, but normal MHC class II expression after rat liver transplantation. PMID- 11266902 TI - Cutaneous CsA-resistant veiled (dendritic) cells are responsible for uncontrolled skin allograft rejection. PMID- 11266903 TI - Natural killer cell regulatory receptor changes following tolerance induction. PMID- 11266904 TI - Comparison of the direct and indirect pathways of allorecognition in chronic allograft failure. PMID- 11266905 TI - A20 and BCL proteins exert a broad and complementary cytoprotective effect in endothelial cells via blockade of NF-kappaB and NFAT. PMID- 11266906 TI - Downregulation of nuclear factor kappa B expression in primate cardiac allograft arteries after E2F decoy transfection. PMID- 11266908 TI - Gamma-chain T-cell receptor transcripts are clonally expanded in the coronary arteries of cardiac allografts from patients with chronic rejection. PMID- 11266907 TI - Coronary arteries with chronic rejection contain oligoclonal T cells: persistence of clonally expanded T cell receptor transcripts from the early posttransplantation period through chronic rejection. PMID- 11266909 TI - Alloantigen-specific CD8(+) T cells stimulate endothelial cells to produce the T cell chemoattractants IP-10 and Mig. PMID- 11266910 TI - Immune reactivity toward HLA class II determinants in renal transplantation. PMID- 11266911 TI - New immunosuppressants and HLA matching. PMID- 11266912 TI - Low levels of HLA-specific antibody: relevance, detection, and treatment. PMID- 11266913 TI - Pregnancy: a two-edged sword. PMID- 11266914 TI - Pronase improves detection of HLA antibodies in flow crossmatches. PMID- 11266915 TI - An association between posttransplant antibody production and renal transplant rejection. PMID- 11266916 TI - Flow PRA to detect clinically relevant HLA antibodies. PMID- 11266917 TI - Relevance of posttransplant HLA class I and class II antibodies on renal graft outcome. PMID- 11266918 TI - Increased risk for cardiac allograft rejection in recipients with HLA-B8, DR3 type. PMID- 11266920 TI - Association between TGF-beta expression and severe GVHD in allogeneic bone marrow transplantation. PMID- 11266921 TI - Polymorphism in the Y box regulates cytokine-mediated expression of HLA-DR genes. PMID- 11266919 TI - Significance of cytokine gene polymorphism in renal transplantation. PMID- 11266922 TI - Cytokine genotypes in kidney, heart, and lung recipients: consequences for acute and chronic rejection. PMID- 11266923 TI - Microsatellites in the HLA region: HLA prediction and strategies for bone marrow donor registries. PMID- 11266924 TI - HLAMMATCHMAKER: a molecularly based donor selection algorithm for highly alloimmunized patients. PMID- 11266925 TI - Determination of a real-time fluorotyping strategy for the HLA-DR locus. PMID- 11266926 TI - Low-resolution HLA class II typing for solid organ transplantation is inadequate. PMID- 11266927 TI - Cytokine polymorphism and kidney graft survival at a single center. PMID- 11266928 TI - Cytokine gene polymorphism in patients with graft-versus-host disease. PMID- 11266929 TI - Blockade of costimulatory molecules on dendritic cells: implications for tolerance induction. PMID- 11266930 TI - Temporary treatment with sirolimus and low-trough cyclosporine prevents acute islet allograft rejection, and combination with starch-conjugated deferoxamine promotes islet engraftment in the preclinical pig model. PMID- 11266931 TI - IL-10: a tacrolimus-specific cytotoxic mediator in ongoing allograft rejection. PMID- 11266932 TI - Comparison of the in vitro metabolism of the macrolide immunosuppressants sirolimus and RAD. PMID- 11266933 TI - Prolonged survival of murine cardiac allografts by treatment with the 4-amino analog of tetrahydrobiopterin. PMID- 11266934 TI - Prolongation of pancreatic islet graft survival by blocking transferrin receptor (CD71). PMID- 11266935 TI - Immunosuppressive effects of Tripterygium wilfordii hook F in a rat liver transplant model. PMID- 11266936 TI - Castanospermine, an oligosaccharide processing inhibitor, reduces both lymphocyte endothelial cell binding and LFA-1alpha membrane expression. PMID- 11266937 TI - Prevention of intraportal islet failure by a highly selective iNOS inhibitor in the pig-to-nude rat model. PMID- 11266938 TI - Blockade of CD154 at the time of donor-specific blood transfusion does not improve kidney graft survival in rhesus monkeys. PMID- 11266939 TI - Neutralization of IL-12 reverses rejection of mouse liver allografts from Flt3 ligand-treated donors and is associated with suppression of both cellular and humoral responses. PMID- 11266940 TI - Chronic allograft rejection versus tolerance: a critical role for EIIIA(+) fibronectin. PMID- 11266941 TI - Generation of partial agonist ligands of the T-cell receptor by engineering of antibodies against CD3. PMID- 11266942 TI - FTY720 alters lymphocyte homing and protects allografts without inducing general immunosuppression. PMID- 11266943 TI - The technical aspects of combined intestinal and heart transplantation in rats. PMID- 11266944 TI - CD28-B7-2 but not CD28-B7-1 interaction is required for T cell costimulation in experimental obliterative bronchiolitis in the rat. PMID- 11266945 TI - Nitric oxide donor FK409 attenuates the development of neointimal hyperplasia in a rat aortic allograft model. PMID- 11266946 TI - Amelioration of hepatic ischemia/reperfusion injury with intercellular adhesion molecule-1 antisense oligodeoxynucleotides. PMID- 11266947 TI - Boswellic acid, a potent antiinflammatory drug, inhibits rejection to the same extent as high dose steroids. PMID- 11266948 TI - FTY720 prevents chronic rejection of rat heterotopic cardiac allografts. PMID- 11266950 TI - Methyl hydrogen fumarate inhibits acute and chronic rejection in rat kidney transplantation models. PMID- 11266949 TI - Differential expression of CD45 isoforms in tolerant and rejecting rat liver transplant recipients. PMID- 11266951 TI - Exquisitely small amounts of nonglucocorticoid natural steroids suppress the human allogeneic T-cell response. PMID- 11266952 TI - Prolonged class II MHC disparate skin allograft survival by treatment with antibodies to the chemokine Mig. PMID- 11266953 TI - Administration of antisense oligodeoxyribonucleotides against mRNA of CD80 or CD86 prolongs survival of cardiac allografts by inhibition of CTL activity. PMID- 11266954 TI - Effects of locally expressed CTLA4-Ig in a pancreatic beta cell line on the accelerated graft rejection response induced by DST. PMID- 11266955 TI - Cyclolinopeptide: a novel immunosuppressive agent with potential anti-lipemic activity. PMID- 11266956 TI - FK506 and SMS 201-995: effect on heterotopic heart transplantation in rats. PMID- 11266957 TI - Rapamycin reduces expression of fibrosis-associated genes in an experimental model of renal ischaemia reperfusion injury. PMID- 11266958 TI - Experimental study of tacrolimus immunosuppression on the mode of administration: efficacy of constant intravenous infusion avoiding C(max). PMID- 11266960 TI - Gene expression profiles of rat heart allografts with immunosuppression. PMID- 11266959 TI - Long-term survival following canine intrathymic islet allograft transplantation with short-course immunosuppressive therapy. PMID- 11266961 TI - Prolongation of survival and preservation of allograft structure and function by a signaling CD28 mAB in a rat model of chronic kidney rejection. PMID- 11266962 TI - Combination of antilymphocyte globulin and leflunomide leads to superior grafts. PMID- 11266963 TI - Tolerance induction by a single donor pretreatment with the adenovirus vector encoding CTLA4Ig gene in rat orthotopic liver transplantation. PMID- 11266964 TI - How can adenovirus-mediated catalase and superoxide dismutase gene transfer improve the outcome of pancreatic cells for transplantation? PMID- 11266965 TI - Adenovirus-mediated gene transfer of A20 in murine islets inhibits Fas-induced apoptosis. PMID- 11266966 TI - Soluble donor MHC class I gene therapy prevents accelerated heart allograft rejection in actively sensitized rat recipients. PMID- 11266967 TI - Heme oxygenase-1 gene therapy: a novel immunomodulatory approach in liver allograft recipients? PMID- 11266968 TI - Soluble allogeneic MHC class I molecule gene transfer promotes CTL apoptosis in vivo. PMID- 11266969 TI - Hepatic stem/progenitor cells with high proliferative potential in liver organ formation. PMID- 11266970 TI - cMet activation allows persistent engraftment of ectopically transplanted xenogenic human hepatocytes in mice. PMID- 11266971 TI - Efficient gene delivery to vascular smooth muscle cells using a nontoxic, synthetic peptide vector system targeted to membrane integrins: a first step toward the gene therapy of chronic rejection. PMID- 11266972 TI - Retrovirally transduced myeloid dendritic cells expressing mammalian or viral IL 10 differentially affect cell-mediated immunity and growth of transplantable tumors. PMID- 11266973 TI - Tolerance induction using transduction of Sca1+ cells with IL-10 gene. PMID- 11266974 TI - Tissue-engineered spinal cord. PMID- 11266975 TI - Generation of FASL-expressing antigen-presenting cells for suppression of alloreactive T cells in the rat. PMID- 11266976 TI - Protective role of NF-kappaB in liver cold ischemia/reperfusion injury: effects of IkappaB gene therapy. PMID- 11266977 TI - Long-term localized transgene expression in the pancreas achieved by intra arterial adenoassociated virus-mediated gene transfer. PMID- 11266978 TI - Transfection with genes encoding CTLA4Ig mediated by adenoassociated virus vectors prolongs survival of heart allografts. PMID- 11266979 TI - Prolonged survival of rat liver allografts with CrmA gene transfer. PMID- 11266980 TI - Nuclear factor kappa B expression in primate cardiac allograft arteries. PMID- 11266981 TI - Visualization of alloantigen-specific, EGFP-engineered T lymphocytes in a rat kidney transplantation model. PMID- 11266982 TI - Mechanical and histological characteristics of human trachea before and after cryopreservation: an opportunity for tracheal tissue banking. PMID- 11266983 TI - Tissue engineered alternatives to nerve transplantation for repair of peripheral nervous system injuries. PMID- 11266984 TI - Tolerance to vascularized musculoskeletal allografts. PMID- 11266985 TI - Investigation on reconstruction of skin for autologous transplantation. PMID- 11266986 TI - Lack of human endothelial cell activation by adult porcine islets. PMID- 11266987 TI - Psychological investigation protocol of candidates for hand transplantation. PMID- 11266988 TI - Viability of vascularized bone grafts; perfusion studies by dynamic enhanced MRI and bone scan. PMID- 11266989 TI - Assessment of cartilage viability in long-term cryopreserved tracheal allografts. PMID- 11266990 TI - Skin allografts-host cutaneous veiled cells initiate rejection reaction by indirect pathway of allorecognition. PMID- 11266991 TI - The fate of epidural autologous free fat grafts; longitudinal MRI surveillance. PMID- 11266992 TI - Microbiologic indicators as quality control in a valvular homografts bank. PMID- 11266993 TI - Low-dose, short-term treatment with anti-CD4 monoclonal antibody prolongs corneal allograft survival. PMID- 11266994 TI - The Derma Project: present and future possibilities of skin procurement for the treatment of large burns in Argentina, Tissue Engineering and the Cadaver Skin Bank. PMID- 11266996 TI - Transplantation of cultured autologous keratinocytes in fibrin sealant biomatrix to resurface chronic wounds. PMID- 11266995 TI - Helper T cell frequencies after implantation of aortic valve allografts in rats. PMID- 11266998 TI - Functional evaluation of the AMC-BAL to be employed in a multicentric clinical trial for acute liver failure. PMID- 11266997 TI - Role of immortalized hepatocyte transplantation in acute liver failure. PMID- 11266999 TI - FLIM58, a blocking peptide for FAS ligand, reduces apoptotic cell death of allogenic-transplanted hepatocytes in spleen. PMID- 11267000 TI - Early loss of transplanted autologous hepatocytes-lysis by leukocytes in vivo and in vitro. PMID- 11267001 TI - Hepatocyte isolation using human livers discarded from transplantation: analysis of cell yield and function. PMID- 11267002 TI - In vitro evaluation of growth and anabolism for C3A/HepG2 hepatoma cells with logistic equation and linear regression expression. PMID- 11267003 TI - Revascularization and remodeling of transplanted pancreatic islets. PMID- 11267004 TI - Multiaggregate hepatocyte (HP) spheroids in the hepato-cellular transplant: structural, functional and metabolic characterization. PMID- 11267006 TI - Steatotic versus cirrhotic livers as a source for human hepatocyte isolation. PMID- 11267005 TI - Islet autotransplantation combined with total pancreatectomy for treatment of pancreatic adenocarcinoma. PMID- 11267007 TI - Cryopreserved human parathyroid tissue: cell cultures for in vitro testing of function. PMID- 11267008 TI - Isolated pancreatic islets: structural and ultrastructural analysis. PMID- 11267009 TI - Purification of pancreatic islets using hydroxyethyl starch-Collins solution. PMID- 11267010 TI - FTY720, a novel immunosuppressive agent with insulinotropic activity, prolongs graft survival in a mouse islet transplantation model. PMID- 11267011 TI - Differentiation of endoderm derivatives, pancreas and intestine, from rhesus embryonic stem cells. PMID- 11267012 TI - Primate skin allotransplantation with anti-CD154 monotherapy. PMID- 11267013 TI - Comparison of Unisol with Euro-Collins solution as a vehicle solution for cryoprotectants. PMID- 11267014 TI - Temporal relationship of insulin, intact proinsulin and split proinsulin after islet autotransplantation. PMID- 11267015 TI - Mitogenic effects of rat Sertoli cells on adult homologous islet beta-cells: in vitro and in vivo studies. PMID- 11267016 TI - Surgical planning of human hand transplantation. PMID- 11267017 TI - Modulation of endothelial cell function in transgenic animals. PMID- 11267018 TI - The nature of elicited xenoantibodies. PMID- 11267019 TI - Altered infectivity of porcine endogenous retrovirus by "protective" avian antibodies: implications for pig-to-human xenotransplantation. PMID- 11267020 TI - Porcine cells are susceptible to infection by human alpha herpesviruses. PMID- 11267021 TI - Long-term monitoring of xenotransplanted baboons: no evidence for pig endogenous retrovirus transmission. PMID- 11267022 TI - The influence of donor brain death on long-term function of renal allotransplants in rats. PMID- 11267023 TI - Unsensitized human T cells do not infiltrate or injure quiescent pig coronary artery grafts in immunodeficient mouse hosts. PMID- 11267025 TI - Preventing anti-Gal response in xenograft recipients by an alpha-Gal toxin. PMID- 11267026 TI - Early induction of TIMP-1 in endothelial cells in response to human xenoreactive IgG. PMID- 11267027 TI - IL-5 prolongs allograft survival by downregulating IL-2 and IFN-gamma cytokines. PMID- 11267024 TI - Immunosuppression of direct T-cell-mediated xenorecognition in vitro. PMID- 11267028 TI - Induction of long-term chimerism in a pig-to-primate model of peripheral tolerance induction. PMID- 11267029 TI - Defect of severe combined immunodeficiency (Scid) MPhi/DC in acquired tolerance induction following rat-into-scid xenogeneic bone marrow cell chimeras. PMID- 11267030 TI - Inhibition of NF-kB activation in pig APC by xenospecific human T-suppressor cells. PMID- 11267031 TI - Comparison of the regulatory regions of the alpha1,3galactosyltransferase gene between murine and porcine species. PMID- 11267032 TI - Xenogeneic islets transplanted into primates are subject to a form of hyperacute rejection independent of anti-GAL-alpha 1-->3 gal antibody binding. PMID- 11267033 TI - Effects of virus expressing CTLA4-IG and IL1-RA on islet xenografts. PMID- 11267035 TI - Miniature swine and hDAF pig kidney transplantation in baboons treated with a nonmyeloablative regimen and CD154 blockade. PMID- 11267036 TI - Antibody responses in early graft rejection in pig-to-primate renal xenotransplantation. PMID- 11267034 TI - Acute vascular rejection of h-DAF transgenic porcine kidneys in immunosuppressed cynomolgus monkeys is associated with systemic and intragraft complement activation. PMID- 11267037 TI - Human thrombomodulin improves the microcirculation of the xeno-perfused porcine liver. PMID- 11267038 TI - The effect of anti-alphaGal antibody removal with immunoabsorption and splenectomy on CD46 transgenic kidney xenograft survival. PMID- 11267039 TI - Detection of elicited antiporcine antibodies by FACS and Galalpha1-3Gal-specific ELISA in cynomolgus recipients after porcine kidney transplantation. PMID- 11267040 TI - Soluble complement receptor 1 (TP10) preserves adult porcine islet morphology after intraportal transplantation into cynomolgus monkeys. PMID- 11267041 TI - T-cell-directed immunosuppression allows prolonged survival of xenogeneic pig islets in monkeys. PMID- 11267042 TI - Renal physiology in pig-to-baboon xenografts. PMID- 11267043 TI - Porcine heart xenograft function and hyperacute rejection: the role of thrombin inhibition. PMID- 11267045 TI - Analysis of hematopoesis in cynomolgus recipients of h-CD55 transgenic or unmodified porcine kidneys. PMID- 11267044 TI - Differential detection of alpha-gal and human CD59 molecules on pig-to-primate cardiac xenotransplantation: a marker of delayed xenograft rejection. PMID- 11267046 TI - Discordant lung xenotransplantation using alpha-GAL columns, pig-kidney adsorption, and complement depletion in baboons. PMID- 11267047 TI - Influence of cold ischemia time on hyperacute rejection and delayed graft function of porcine kidneys following discordant xenotransplantation from pig to cynomolgus monkey. PMID- 11267048 TI - Transgenic pigs with human N-acetylglucosaminyltransferase III. PMID- 11267049 TI - A codon exchange DAF is a powerful material for xenografting. PMID- 11267050 TI - The role of indirect recognition and TH2 response in cardiac xenograft rejection. PMID- 11267051 TI - Long-term survival of hamster hearts in presensitized rats. PMID- 11267052 TI - Long-term survival in hamster-to-rat cardiac xenotransplantation using cyclosporine A alone. PMID- 11267053 TI - Effects of gene transfection of human BCL-2 on graft coronary arteriosclerosis in hamster-to-rat cardiac xenografts. PMID- 11267054 TI - Complement inactivated, preformed antibodies do not play a pivotal role in delayed rejection of a guinea pig-to-rat cardiac xenograft. PMID- 11267056 TI - Activation of porcine hepatic microvascular sinusoidal endothelial cells in pig to-human liver xenotransplantation. PMID- 11267055 TI - Neonatal porcine pancreas as a source of islet transplantation. PMID- 11267057 TI - Apoptosis of pig endothelial cells following hyperacute rejection. PMID- 11267059 TI - Differential engraftment of human peripheral blood lymphocytes in various strains of immunodeficient mice. PMID- 11267058 TI - Removal of alphaGal antigens by ex vivo perfusion of pig kidneys with endo-beta galactosidase C. PMID- 11267060 TI - Effects of xenogeneic thymic transplantation in baboons. PMID- 11267061 TI - Simultaneous blockade of B7-CD28 and CD40-CD40L costimulation eliminates the direct xenorestricted human anti-porcine T-cell response. PMID- 11267062 TI - Transplantation of MCP cardiac xenografts into baboon recipients. PMID- 11267063 TI - Improved function of transgenic pig lungs in ex vivo lung perfusion with human blood. PMID- 11267064 TI - Overexpression of superoxide dismutase in a concordant xenograft does not alleviate acute rejection. PMID- 11267065 TI - TH2 cytokines regulate gene expression and proinflammatory responses in xenografts. PMID- 11267066 TI - Immune complex disease in immunosuppressed rat recipients of hamster liver xenografts. PMID- 11267067 TI - Adenovirus-mediated gene transfer of triple human complement regulating proteins in xenogeneic rat liver perfusion. PMID- 11267068 TI - Cellular immunity overrules the protective effect of human DAF as demonstrated in an ex vivo heart perfusion model. PMID- 11267069 TI - Sequential immunohistological analysis of porcine grafts in cynomolgus recipients after discordant kidney xenotransplantation. PMID- 11267070 TI - The role of indirect antigen recognition in islet xenograft rejection. PMID- 11267071 TI - Long-term survival of pig islets in the rat under cyclosporine monotherapy. PMID- 11267072 TI - Expression of tissue factor and tissue factor pathway inhibitor in porcine endothelium in response to natural antibody and complement. PMID- 11267073 TI - C1-inhibitor attenuates human serum induced E-selectin expression of pig aortic endothelial cells. PMID- 11267074 TI - Xeno-tolerance induced by blood transfusion in athymic nude rats. PMID- 11267075 TI - In vivo differentiation of human lymphocytes in a porcine microenvironment. PMID- 11267076 TI - For a rational approach to the critical points of the cadaveric donation process. PMID- 11267077 TI - The organ shortage and allocation issues. PMID- 11267078 TI - Crossover renal transplantation: hurdles to be cleared! PMID- 11267079 TI - Living kidney donation. PMID- 11267080 TI - Pancreas transplants from living donors: short- and long-term outcome. PMID- 11267081 TI - Three-year experience with the new Eurotransplant kidney allocation system 1996 1999. PMID- 11267082 TI - Role of local renin-angiotensin system in warm ischemia and reperfusion injury of the liver. PMID- 11267083 TI - Long-term results of renal transplantation using organs from non-heart-beating donors. PMID- 11267085 TI - Influence of donor characteristics on kidney sharing and discard: analysis of UNOS data. PMID- 11267084 TI - Donor cytokine gene polymorphisms are associated with increased graft loss and dysfunction after transplant. PMID- 11267086 TI - Comparison of CT and MR angiography for evaluation of living renal donors. PMID- 11267087 TI - Arginine-specific proteolytic activity predicts graft survival. PMID- 11267088 TI - Efficiency of special programs for highly sensitized patients within eurotransplant. PMID- 11267089 TI - Improvement of graft function without donor pretreatment in liver transplantation from non-heart-beating donors. PMID- 11267090 TI - Elimination of Kupffer cells suppresses activation of nuclear factor kappa B and production of cytokines and eicosanoids in non-heart-beating donors. PMID- 11267093 TI - Pathophysiology of brain death: effects on allograft function. PMID- 11267091 TI - Hearts from non-heart-beating donors: sodium-hydrogen-inhibitor cariporide improves functional recovery. PMID- 11267092 TI - Ischemia-reperfusion injury. PMID- 11267094 TI - Preservation of non-heart-beating donor kidneys: a clinical prospective randomised case-control study of machine perfusion versus cold storage. PMID- 11267095 TI - Attenuation of hepatic ischemia and reperfusion injury by serine protease inhibitor, FUT-175, in dogs. PMID- 11267096 TI - Effect of preconditioning in the liver against ischemia/reperfusion injury, protection of sinusoidal cells and alterations of gene transcription. PMID- 11267097 TI - Impact of prolonged cold ischemia and reperfusion on apoptosis, activation of caspase 3, and expression of bax after liver transplantation in the rat. PMID- 11267098 TI - Pyruvate prevents intestinal functional changes following ischemia-reperfusion injury. PMID- 11267099 TI - The effect of sodium nitroprusside on the chronic allograft damage in a rat model of renal transplantation. PMID- 11267100 TI - Methoxyethyl modification of phosphorothioate ICAM-1 antisense oligonucleotides improves prevention of ischemic/reperfusion injury. PMID- 11267101 TI - Loss of No-induced relaxation in abdominal aorta preserved in a Co-culture system. PMID- 11267102 TI - FTY 720 prevents ischemic reperfusion damage in rat kidneys. PMID- 11267103 TI - Inhibition of type II phospholipase A(2) by LY329722 attenuates ischemia and reperfusion injury of canine livers. PMID- 11267104 TI - Selective cyclooxygenase-2 inhibitor ameliorates warm ischemia-reperfusion injury of the canine liver. PMID- 11267105 TI - Targeted inhibition of the small GTPase protects against ischemia/reperfusion liver injury in mice. PMID- 11267106 TI - FR167653 ameliorates ischemia-reperfusion injury of the rat liver through P38 mitogen-activated protein kinase pathway. PMID- 11267107 TI - Endothelin converting enzyme protects postischemic liver. PMID- 11267109 TI - Histomorphometric analysis of the liver in brain-dead donors. PMID- 11267108 TI - Early and late inflammatory changes occurring in rat renal isografts from brain dead donors. PMID- 11267110 TI - Randomized clinical study comparing UW and Celsior solution in liver preservation for transplantation: preliminary results. PMID- 11267112 TI - Evaluation of a new preservation solution containing fosfomycin for 20-hour canine lung preservation. PMID- 11267111 TI - Pancreas preservation with Celsior solution in a pig autotransplantation model: comparative study with University of Wisconsin solution. PMID- 11267113 TI - The effect of FK409 on small bowel ischemia-reperfusion injury in dogs. PMID- 11267114 TI - Comparison of the protective effects of phosphate-buffered sucrose and University of Wisconsin solution in a non-heart-beating liver donor experiment. PMID- 11267115 TI - Comparison of two preservation solutions in the protection of the pH regulation mechanism of perfused rat livers after 24 hours of cold storage. PMID- 11267116 TI - Effect of sodium lactobionate sucrose solution on the function of liver grafts from non-heart-beating pig donors. PMID- 11267118 TI - Cold ischaemia further increases intrarenal resistance when non-heart-beating kidneys are pulsatile machine perfused. PMID- 11267117 TI - Optimal temperature in pancreas preservation by the two-layer cold storage method before islet isolation. PMID- 11267119 TI - Development of a human proximal tubule cell culture acidotic/WIT model. PMID- 11267120 TI - Involvement of calcium influx in hypoxia-induced bleb formation in human umbilical vein endothelial cells. PMID- 11267121 TI - Doppler ultrasound in prediction of the early mortality risk factors on the waiting list for pediatric liver transplantation recipients. PMID- 11267122 TI - Influence of a younger environment on organ transplantation from elderly donors. PMID- 11267123 TI - Intraoperative assessment of liver transplantation with marginal liver grafts. PMID- 11267124 TI - Renal transplantation from older living donors. PMID- 11267125 TI - Accelerated graft dysfunction in renal isografts from non-heart-beating donors. PMID- 11267126 TI - Newly developed cathether set for in situ protection of hearts. PMID- 11267128 TI - Preservation of rat livers by supercooling under high pressure. PMID- 11267129 TI - Sharing 0-antigen mismatched cadaveric kidneys does not decrease the graft survival of payback kidneys. PMID- 11267127 TI - Six-year experience in continuous hypothermic pulsatile perfusion kidney preservation. PMID- 11267131 TI - Enhanced endogenous adenosine protects from reperfusion injury after heart transplantation. PMID- 11267132 TI - In vivo measurement of radical scavenger efficacy of vitamins C and E in a pig model of pulmonary reperfusion injury. PMID- 11267130 TI - Celsior is superior to UW for graft preservation from non-heart-beating donors in a canine liver transplantation model. PMID- 11267133 TI - JNK and p38MAPK are activated during graft reperfusion and not during cold storage in rat liver transplantation. PMID- 11267134 TI - Effect of dipyridamole on ischemia and reperfusion injury of canine liver. PMID- 11267135 TI - Evaluation of the use of graft livers procured from old donors (70 to 87 years) for hepatic transplantation. PMID- 11267136 TI - Comparison between nonpulsatile and pulsatile machine perfusion preservation in liver transplantation from non-heart-beating donors. PMID- 11267137 TI - C1 inhibitor potentiates the protective effect of organ preservation solution on endothelial cells during cold storage. PMID- 11267139 TI - TP20 is superior to TP10 in reducing ischemia/reperfusion injury in rat lung grafts. PMID- 11267138 TI - International certificate DIN EN ISO 9001 for organ donation and procurement organization-a high-level guarantee of quality and safety. PMID- 11267140 TI - Protective effects of lactobionate in modified phosphate-buffered sucrose. PMID- 11267141 TI - Successful pancreas preservation before islet isolation by the simplified two layer cold storage method. PMID- 11267142 TI - Mitochondrial injury limits salvaging marginal livers by machine perfusion. PMID- 11267143 TI - Ischemic preconditioning enhances regenerative capacity of hepatocytes after prolonged ischemia. PMID- 11267145 TI - Hepatic artery resistance as a marker for preservation/reperfusion injury. PMID- 11267144 TI - High-pressure perfusion versus gravity perfusion in liver harvesting: results from a prospective randomized study. PMID- 11267146 TI - Prediction of chronic allograft failure using computerized image analysis of postperfusion biopsy specimen: study of cadaver kidney transplants. PMID- 11267147 TI - Application of exsanguineous metabolic support to human kidneys. PMID- 11267148 TI - A novel volunteer intervention to increase organ donation. PMID- 11267149 TI - Attitudes of high school students regarding organ donation. PMID- 11267150 TI - Donor pretreatment of grafts from marginal donors improves long-term graft outcome. PMID- 11267151 TI - Inhibitory effect of lidocaine on lipid peroxidation during cold preservation of pancreas. PMID- 11267152 TI - Influence of selenium therapy on liver microcirculation after warm ischemia/reperfusion: an intravital microscopy study. PMID- 11267153 TI - Phosphodiesterase type III inhibitor enhances the protective effects of adenosine against ischemic hepatic injury in rats. PMID- 11267154 TI - High dose of geranylgeranylacetone accumulate HSP72 mRNA in rat liver. PMID- 11267155 TI - Ginkgo biloba (EGB 761) improves microcirculation after warm ischemia of the rat liver. PMID- 11267156 TI - Elevation of cyclic nucleotides attenuates ischemia and reperfusion injury of liver. PMID- 11267158 TI - Steroid withdrawal in renal transplant recipients. PMID- 11267157 TI - Oxidant stress and antioxidant capacity in urine of renal transplant recipients predict early graft function. PMID- 11267159 TI - The practical problems of prednisone in pediatric renal transplantation. PMID- 11267160 TI - Immunosuppression and skeletal disorders. PMID- 11267161 TI - Tacrolimus as cornerstone immunosuppressant in kidney transplantation. PMID- 11267162 TI - Place of mycophenolate mofetil in renal transplantation. PMID- 11267163 TI - The role of newer monoclonal antibodies in renal transplantation. PMID- 11267164 TI - Polyclonal antibodies in immunosuppression. PMID- 11267165 TI - Mycophenolate mofetil decreases the risk for chronic renal allograft failure. PMID- 11267166 TI - Treatment with the chimeric anti-IL-2Ralpha basiliximab affects both the IL-2 and IL-15 signalling pathways after clinical kidney transplantation. PMID- 11267167 TI - Basiliximab significantly reduces acute rejection in renal transplant patients given triple therapy with azathioprine. PMID- 11267168 TI - Rapid steroid withdrawal versus standard steroid therapy in patients treated with basiliximab, cyclosporine, and mycophenolate mofetil for the prevention of acute rejection in renal transplantation. PMID- 11267169 TI - Daclizumab induction for primary kidney transplant recipients using tacrolimus, mycophenolate mofetil, and steroids as maintenance immunosuppression. PMID- 11267170 TI - Induction therapy with daclizumab as part of the immunosuppressive regimen in human small bowel and multiorgan transplants. PMID- 11267171 TI - Influence of donor-recipient HLA-DR matching on efficacy of anti-CD25 mAb in cardiac transplantation. PMID- 11267172 TI - Tacrolimus (FK 506) in kidney transplantation: five-year survival results of the U.S. multicenter, randomized, comparative trial. PMID- 11267173 TI - One-year follow-up of a large European trial comparing dual versus triple tacrolimus-based immunosuppressive regimens following renal transplantation. PMID- 11267174 TI - Conversion from cyclosporin to tacrolimus in chronic allograft nephropathy. PMID- 11267176 TI - Drug regimens affect cadaveric kidney graft survival. PMID- 11267177 TI - Sirolimus is effective for prevention of acute rejection in primary mismatched renal allograft recipients. PMID- 11267175 TI - Conversion to neoral provides effective rescue therapy for liver and kidney transplant patients intolerant of Prograf. PMID- 11267178 TI - Sirolimus rescue therapy for refractory rejection in renal transplant recipients. PMID- 11267180 TI - Effect of rapamycin and FK506 on mesangial cell proliferation. PMID- 11267179 TI - Conversion at first rejection: a prospective trial comparing cyclosporine microemulsion with tacrolimus in renal transplant recipients. PMID- 11267181 TI - Sirolimus-tacrolimus combination for combined kidney-pancreas transplantation: effect on renal function. PMID- 11267182 TI - Monitoring of mycophenolic acid in pediatric renal transplant recipients. PMID- 11267183 TI - Tacrolimus, MMF, steroid, and ALG immunotherapy for high immunological risk renal transplant recipients. PMID- 11267184 TI - FK506 and CsA differ in their effect on intracellular cytokine expression following kidney transplantation. PMID- 11267185 TI - ISA(TX)247: a novel calcineurin inhibitor. PMID- 11267186 TI - Determination of mycophenolate area under the curve by limited sampling strategy. PMID- 11267187 TI - The effect of mycophenolate mofetil on liver regeneration. PMID- 11267188 TI - Loss of cyclosporin-resistant allospecific T cells with age. PMID- 11267189 TI - Induction strategy using basiliximab combined with mycophenolate MMF and immediate low-dose cyclosporin is steroid sparing and more effective than OKT3. PMID- 11267191 TI - Sirolimus does not potentiate indices of chronic cyclosporin nephrotoxicity. PMID- 11267190 TI - Sparse-sampling algorithms and C2 monitoring are beneficial to optimize clinical outcomes for neoral. PMID- 11267192 TI - Lipid profiles in liver transplantation in patients receiving tacrolimus or cyclosporin. PMID- 11267193 TI - Pharmacokinetics of tacrolimus in pediatric renal transplant recipients. PMID- 11267195 TI - Acute rejection after liver transplantation despite intragraft coating by interleukin-2 receptor-alpha with basiliximab. PMID- 11267194 TI - Mycophenolate mofetil is superior in combination with tacrolimus compared to cyclosporine for immunosuppressive therapy after liver transplantation. PMID- 11267196 TI - A single-center open label randomized trial of the safety and efficacy of the use of sirolimus versus azathioprine in one-haplotype living related kidney transplant recipients-preliminary results. PMID- 11267197 TI - Prediction of the inhibition of IL-2 production by calcineurin inhibitors. PMID- 11267198 TI - Cyclosporine and tacrolimus alter renin-angiotesin system in mouse medullary thick ascending limb cultured cells. PMID- 11267199 TI - The acyl glucuronide metabolite of mycophenolic acid inhibits the proliferation of human mononuclear leukocytes. PMID- 11267200 TI - Mofetil mycophenolate in renal transplantation. PMID- 11267201 TI - Rescue therapy: a role for sirolimus in lung and heart transplant recipients. PMID- 11267202 TI - Pharmacokinetic interactions between sirolimus and cyclosporine exacerbate renal dysfunction. PMID- 11267203 TI - Treatment of recurrent hepatitis C after liver transplantation with IL-2r Ab. PMID- 11267204 TI - The impact of antilymphocyte therapy on sensitized patients in renal transplantation. PMID- 11267205 TI - Fast quantification method for sirolimus and its major metabolites. PMID- 11267206 TI - IVIG rescue therapy in renal transplantation. PMID- 11267208 TI - The dual kidney transplant registry. PMID- 11267207 TI - Choosing the right dose of new immunossuppressive drugs for new populations: importance of pharmacokinetic studies. PMID- 11267209 TI - Non-heart-beating donors: estimated actual potential. PMID- 11267210 TI - Retroperitoneoscopy-assisted living donor nephrectomy: 109 cases. PMID- 11267211 TI - Hand-assisted laparoscopic donor nephrectomy: comparable donor/recipient outcomes, costs, and decreased convalescence as compared to open donor nephrectomy. PMID- 11267212 TI - Improved recipient results after 5 years of performing laparoscopic donor nephrectomy. PMID- 11267213 TI - Laparoscopic donor nephrectomy: analysis of donor and recipient outcomes. PMID- 11267214 TI - Risk factors for kidney transplant acute rejection: a multivariate analysis. PMID- 11267215 TI - Recipient age as an independent risk factor for chronic renal allograft failure. PMID- 11267216 TI - Elderly transplant recipients may require less immunosuppression. PMID- 11267217 TI - Targeting cardiovascular risk in renal transplantation. PMID- 11267218 TI - Machine perfusion and viability assessment of non-heart-beating donor kidneys-a single-centre result. PMID- 11267219 TI - Prediction of outcome in non-heart-beating kidney transplantation. PMID- 11267220 TI - Multivariate analyses of factors contributing to early graft function in 759 kidney transplants from non-heart-beating donors. PMID- 11267222 TI - Allospecific human T suppressor cells in kidney transplantation. PMID- 11267221 TI - Results of transplantation with kidneys from non-heart-beating donors. PMID- 11267223 TI - Emerging profiles in 105 recipients of renal allografts functioning for 20 to 35 years: the "watershed" effect. PMID- 11267224 TI - Kidney transplantation with bone marrow augmentation: five-year outcomes. PMID- 11267226 TI - Renal allograft survival in patients who had simultaneous liver and kidney transplantation compared with those who had renal transplantation alone. PMID- 11267225 TI - Effect of HLA compatibility, pregnancies, blood transfusions, and taboo mismatches in living unrelated kidney transplantation. PMID- 11267227 TI - Renal transplantation using kidneys from elderly donors. PMID- 11267228 TI - Prevention of bone loss in kidney graft recipients. PMID- 11267229 TI - Influence of donor/recipient size in living donor kidney transplantation. PMID- 11267230 TI - Excellent outcome despite adverse histology after cadaveric renal transplantation. PMID- 11267231 TI - Histological injury and renal transplant outcome: the cumulative damage hypothesis. PMID- 11267232 TI - C4d deposits in renal allografts are associated with inferior graft outcome. PMID- 11267233 TI - Chronic allograft nephropathy categorised in chronic interstitial and vascular rejection. PMID- 11267234 TI - The new Banff classification of renal transplant biopsies: a major impact on the adequacy of the cores taken. PMID- 11267236 TI - Relationship between TGFbeta(1), intratubular CD103 positive T cells and acute renal allograft rejection. PMID- 11267237 TI - Improvement of long-term function in renal allografts from 'marginal donors' following the induction of heme-oxygenase-1. PMID- 11267235 TI - Plasma homocysteine levels and C677T MTHFR gene polymorphism in stable renal graft recipients. PMID- 11267239 TI - Reliability of clinical parameters for the selection of elderly cadaveric donors. PMID- 11267238 TI - Matching does not improve outcome from aged marginal kidney donors. PMID- 11267240 TI - Double kidney transplant (dual) with kidneys from older donors and suboptimal nephronal mass. PMID- 11267241 TI - Five and 10 year follow-up of En Bloc small pediatric kidneys in adult recipients. PMID- 11267242 TI - Proteinuria and death risk in the renal transplant population. PMID- 11267243 TI - Enhanced expression of angiotensin II type 1 receptor in the neointima of transplant renal arteriosclerosis in human renal allografts. PMID- 11267245 TI - Histological features of donor grafts for orthotopic liver transplantation. PMID- 11267244 TI - Effects of angiotensin-converting-enzyme inhibitors on progression to end-stage renal failure in chronic vascular rejection. PMID- 11267246 TI - Mortality and long term outcome of combined liver and kidney transplantations. PMID- 11267247 TI - The expression of endothelin and inducible nitric oxide synthase in human renal allografts and their role in chronic renal allograft nephropathy. PMID- 11267248 TI - Significant effect of HLA-DRB1 matching on acute rejection of kidney transplants within 3 months. PMID- 11267249 TI - Renal transplantation and HCV hepatitis: a longitudinal study. PMID- 11267250 TI - Nine year experience with kidney transplantation in patients with positive hepatitis C virus antibody. PMID- 11267251 TI - Time to first graft loss as a risk factor for second renal allograft loss. PMID- 11267252 TI - Need for individualized immunosuppression in elderly renal transplant recipients. PMID- 11267253 TI - Clinical evaluation of early predictors of renal transplantation results. PMID- 11267254 TI - Safety and efficacy of simvastatin for hyperlipidemia in renal transplant recipients: a double-blind, randomized, placebo-controlled study. PMID- 11267257 TI - Uricosuric effect of losartan in renal transplanted patients. PMID- 11267256 TI - Impairment of erythrocyte membrane fluidity in cyclosporine-treated renal transplant patients. PMID- 11267255 TI - CMV in kidney transplants in the tacrolimus-mycophenolate era. PMID- 11267259 TI - Deleterious effect of waiting time on renal transplant outcome. PMID- 11267258 TI - Long-term results of posttransplantation blood transfusion on kidney allograft survival. PMID- 11267260 TI - Collagen IV is upregulated in chronic transplant nephropathy. PMID- 11267262 TI - Initial bone loss in kidney transplant recipients: a prospective study. PMID- 11267261 TI - Differential homocysteine levels in renal transplant patients receiving neoral versus tacrolimus. PMID- 11267263 TI - Mortality's role in kidney transplant failures. PMID- 11267264 TI - Injury of peritubular capillaries correlates with graft function in chronic renal allograft nephropathy. PMID- 11267265 TI - Interleukin-6 in chronic renal allograft rejection: influence of nonimmunologic risk factors. PMID- 11267266 TI - Immunolocalisation of endothelin-1 and its receptors during acute renal allograft rejection. PMID- 11267267 TI - Long-term follow-up of living donor renal transplant recipients sensitized after donor specific blood transfusion. PMID- 11267268 TI - Treatment of calcineurin inhibitor toxicity by dose reduction plus introduction of mycophenolate mofetil. PMID- 11267269 TI - Impact of sirolimus on renal transplant outcomes in African Americans. PMID- 11267270 TI - Use of advanced age donors in living renal transplantation--is it justified? PMID- 11267271 TI - Treatment of acute kidney allograft rejection with the nonmitogenic CD3 mAb CLB T3/4.A (IgA-CD3): absence of a correlation of clinical responsiveness with inhibition of lymphocyte reactivity in vitro. PMID- 11267272 TI - Successful withdrawal of steroid after renal transplantation. PMID- 11267273 TI - Transplantation without a final crossmatch-it can be done. PMID- 11267275 TI - Clinical manifestations and diagnosis of cytomegalovirus infection in renal allograft recipients. PMID- 11267274 TI - Outcome after steroid withdrawal in adult renal transplant patients receiving tacrolimus-based immunosuppression. PMID- 11267276 TI - New method of kidney allograft pretreatment (six-year follow-up). PMID- 11267277 TI - Long-term outcome in hepatitis B sero-positive oriental renal transplant recipients. PMID- 11267278 TI - Intra-arterial stenting for recurrent transplant renal artery stenosis. PMID- 11267279 TI - Efficacy of a National Kidney Transplantation Service: need for, supply and outcome of transplantation. PMID- 11267280 TI - Clinicopathologic evaluation of IgA nephropathy in renal transplant recipients. PMID- 11267281 TI - Increased prevalence of carotid and femoral atherosclerosis in renal transplant recipients. PMID- 11267282 TI - Therapy in ESRD patients with antiphospholipid antibody syndrome. PMID- 11267283 TI - Analysis of primary and recurrent rejection following renal transplantation in a large, comparative, multicentre trial. PMID- 11267284 TI - Calcium oxalate microdeposition in failing kidney grafts. PMID- 11267286 TI - Simulect induction significantly decreases CMV infection in kidney recipients compared to OKT3. PMID- 11267285 TI - Canadian multicentre trial of tacrolimus/azathioprine/steroids versus tacrolimus/mycophenolate mofetil/steroids versus neoral/mycophenolate mofetil/steroids in renal transplantation. PMID- 11267287 TI - Sirolimus permits steroid withdrawal from a cyclosporine regimen. PMID- 11267288 TI - A calcineurin antagonist-free induction immunosuppression strategy for delayed graft function in renal transplantation. PMID- 11267289 TI - Positive effect of steroid withdrawal on bone mineral density in renal allograft recipients. PMID- 11267290 TI - Therapeutic drug monitoring of sirolimus for optimal renal transplant outcomes. PMID- 11267292 TI - Donor employment of vasopressors and its impact on allograft survival after transplantation. PMID- 11267291 TI - A large, multicentre trial to compare the efficacy and safety of tacrolimus with cyclosporine microemulsion following renal transplantation. PMID- 11267293 TI - Influence of brain death on cytokine release in organ donors and renal transplants. PMID- 11267295 TI - Differences in etiology for graft loss in female renal transplant recipients. PMID- 11267294 TI - Induction of heme-oxygenase-1 prevents ischemia/reperfusion injury and improves long-term graft outcome in rat renal allografts. PMID- 11267296 TI - Delayed graft function influences renal function but not survival. PMID- 11267297 TI - Impact of HLA-ABDR match on chronic rejection in kidney transplants. PMID- 11267298 TI - Arterial intimal thickening in stable renal allografts during the first year of follow-up. PMID- 11267299 TI - Donor bone marrow infusions are tolerogenic in human renal transplantation. PMID- 11267300 TI - Dyslipidemia and dietary modification in Chilean renal pediatric transplantation. PMID- 11267301 TI - Annual incidence and predicted risk of nonmelanoma skin cancer in renal transplant recipients. PMID- 11267302 TI - Liver transplantation: expanding the donor pool. PMID- 11267304 TI - Advances in pre- and posttransplant management of hepatitis B virus. PMID- 11267303 TI - Solid organ transplantation in HIV-infected individuals: obstacles and opportunities. PMID- 11267305 TI - CMV and EBV-PTLD after liver transplantation. PMID- 11267306 TI - Duct-to-duct biliary reconstruction in living donor liver transplantation. PMID- 11267307 TI - What is the limit of graft size for successful living donor liver transplantation in adults? PMID- 11267308 TI - 157 adult-to-adult living donor liver transplantation. PMID- 11267309 TI - A new approach to portal vein reconstruction in liver transplantation in patients with distal splenorenal shunts. PMID- 11267310 TI - Liver transplantation using cavoportal hemitransposition- a life-saving procedure in the presence of extensive portal vein thrombosis. PMID- 11267311 TI - Liver transplant with portocaval hemitransposition: experience at the University of Miami. PMID- 11267312 TI - Excessive portal venous inflow as a cause of allograft dysfunction in small-for size living donor liver transplantation. PMID- 11267313 TI - Epoprostenol and nitric oxide therapy for severe pulmonary hypertension in liver transplantation. PMID- 11267314 TI - Split liver transplantation: need for arterial reconstruction in adult patients. PMID- 11267315 TI - Alternative split liver technique: the equal size split. PMID- 11267316 TI - Split liver transplantation in adult patients: hepatic and portal vein division and reconstruction. PMID- 11267317 TI - Tacrolimus versus cyclosporin microemulsion in liver transplantation: results of a 3-month study. PMID- 11267318 TI - De novo use of low-dose tacrolimus and sirolimus in liver transplantation. PMID- 11267319 TI - Prospective randomized trial of tacrolimus and prednisone versus tacrolimus, prednisone, and mycophenolate mofetil: complete report on 350 primary adult liver transplantations. PMID- 11267320 TI - Improved early graft survival in patients receiving aprotinin during orthotopic liver transplantation. PMID- 11267321 TI - Influence of immunosuppression on patient survival after liver transplantation for hepatitis C. PMID- 11267322 TI - Psychiatric disorders in living-related liver transplantation. PMID- 11267323 TI - Donor bone marrow infusion in liver recipients: effect on the occurrence of acute cellular rejection. PMID- 11267324 TI - Early ribavirin treatment and avoidance of corticosteroids in hepatitis C virus (HCV)-positive liver transplant recipients: interim report of a prospective randomized trial. PMID- 11267326 TI - Factors determining outcome of liver transplantation for hepatocellular carcinoma associated with hepatitis C cirrhosis. PMID- 11267325 TI - Prevention of recurrent hepatitis C after liver transplantation with early interferon and ribavirin treatment. PMID- 11267327 TI - Management of hepatoma using cisplatin/epinephrine gel in patients awaiting orthotopic liver transplantation. PMID- 11267328 TI - Pediatric split liver transplantation using elderly donors. PMID- 11267329 TI - Surgical strategy in liver transplantation for polycystic liver disease. PMID- 11267330 TI - When is liver transplant indicated in cirrhosis with bleeding varices? PMID- 11267331 TI - FAP World Transplant Register and domino/sequential register update. PMID- 11267332 TI - A retrospective analysis of HLA matching and other factors on liver graft outcome. PMID- 11267333 TI - Prediction of primary graft failure by intraoperative quantification of liver perfusion. PMID- 11267334 TI - 3D CT angiography in patients before and after liver transplantation. PMID- 11267335 TI - Immunoregulation in liver transplant recipients: possible evidence of tolerance by DTH assay. PMID- 11267336 TI - TGF-beta expression in protocol transplant liver biopsies: a comparative study between cyclosporine-A (CyA) and tacrolimus (FK 506) immunosuppression. PMID- 11267337 TI - Allospecific T-suppressor cells in liver transplantation. PMID- 11267339 TI - Analysis of donor complications in living donor liver transplantation. PMID- 11267338 TI - Could expression of co-stimulatory molecules on B-PBL condition the acceptance or rejection of human liver grafts? PMID- 11267340 TI - Infrahepatic cavo-cavostomy as a rescue technique for complicated piggyback liver transplantation. PMID- 11267341 TI - GH/GHBP changes in the perioperative course of liver transplantation: pathophysiologic and clinical implications. PMID- 11267342 TI - The impact of liver transplantation on diabetes mellitus. PMID- 11267343 TI - Hemodynamic-volumetric versus pulmonary artery catheter monitoring during anesthesia for liver transplantation. PMID- 11267344 TI - Improved outcome for HCV recipients with normal serum transaminases after liver transplantation. PMID- 11267345 TI - The importance of early prevention of renal dysfunction in liver transplant recipients. PMID- 11267346 TI - Outcome of adult living donor liver transplantation using small volume of left liver graft less than 1% of body weight. PMID- 11267347 TI - Auxiliary orthotopic liver transplantation: new technique and results in toxic liver injury. PMID- 11267348 TI - Intravascular blood volume in cirrhotic patients. PMID- 11267349 TI - Hepatic artery thrombosis after orthotopic liver transplantation. PMID- 11267350 TI - Various types of donor hepatectomy according to the required graft volume in adult living donor liver transplantation. PMID- 11267351 TI - Liver retransplantation: indications and results over a 15-year experience. PMID- 11267352 TI - Indication, technique, and outcome of portal vein arterialization in orthotopic liver transplantation. PMID- 11267353 TI - The influence of donor age to graft volume increase rate in living donor liver transplantation. PMID- 11267355 TI - One liver for two adults: in situ split liver transplantation for two adult recipients. PMID- 11267354 TI - Daclizumab induction therapy in combination with tacrolimus. PMID- 11267356 TI - Phenotypic marker and cytotoxic potential of hepatic lymphocytes isolated from rejected livers. PMID- 11267357 TI - Successful orthotopic liver transplantation for treatment of intrahepatic Osler's disease. PMID- 11267358 TI - Preliver transplant viremic status and the choice of anti-HBV regimens. PMID- 11267359 TI - Sequential liver transplantation: 27 cases in 25 patients. PMID- 11267361 TI - Surgical treatment of Budd-Chiari syndrome--when is liver transplant indicated? PMID- 11267362 TI - Direct costs for one year of liver transplant care are directly associated with disease severity at transplant. PMID- 11267360 TI - IL-2 receptor antibody induction increases the risk for chronic rejection after liver transplantation. PMID- 11267363 TI - Celiac axis compression syndrome in liver transplantation. PMID- 11267364 TI - Long-term outcome of 27 patients after combined liver-kidney transplantation. PMID- 11267365 TI - Liver transplantation for small hepatocellular carcinoma in cirrhosis: analysis of our experience. PMID- 11267366 TI - Modulation of liver graft hemodynamics by partial ablation of the splenic circuit: a way to increase hepatic artery flow? PMID- 11267367 TI - Myocardial perfusion scintigraphy in patients with liver cirrhosis evaluated for orthotopic liver transplantation. PMID- 11267368 TI - Single-dose induction with daclizumab immediately after liver transplantation in pediatric patients. PMID- 11267369 TI - Parenthood following liver transplantation. PMID- 11267370 TI - Treatment of HCC: the role of liver resection in the era of transplantation. PMID- 11267371 TI - Natural alpha-IFN in HCV recurrence after liver transplantation. PMID- 11267372 TI - Split liver and living donation liver transplantation: the Berlin experience. PMID- 11267374 TI - Liver transplantation in recipients over 60. PMID- 11267373 TI - Tacrolimus or cyclosporine for immunosuppression after cardiac transplantation: which treatment reveals more side effects during long-term follow-up? PMID- 11267375 TI - Recurrent HCV infection reduces the requirement for tacrolimus after liver transplantation. PMID- 11267376 TI - Natural history of recurrent hepatitis C after liver transplantation. PMID- 11267377 TI - Use of microwave coagulation therapy in liver transplant candidates with hepatocellular carcinoma: a preliminary report. PMID- 11267378 TI - Steroid-free immunosuppression through thymoglobulin induction in liver transplantation. PMID- 11267379 TI - Transdermal estrogen therapy improves lipid profile and osteoporosis in postmenopausal liver transplant patients. PMID- 11267380 TI - Single centre experience with in situ right split liver transplantation in cirrhotic adults. PMID- 11267381 TI - Marginal liver: case report of a successful OLT from an 84-year-old donor. PMID- 11267382 TI - Circulating blood volume monitoring during liver transplantation for fulminant hepatic failure. PMID- 11267383 TI - Causes of death after liver transplantation in 4000 consecutive patients: 2 to 19 year follow-up. PMID- 11267384 TI - Significance of portal venous flow in graft regeneration after living related liver transplantation. PMID- 11267385 TI - Causes of retransplantation after primary liver transplantation in 4000 consecutive patients: 2 to 19 years follow-up. PMID- 11267386 TI - Liver transplant with cavoportal hemitransposition for portal and mesenteric thrombosis: case report. PMID- 11267387 TI - Center experience in liver transplantation for hepatocellular carcinoma associated with cirrhosis. PMID- 11267388 TI - Evaluation of non-heart-beating donors as a potential source for pancreatic islet transplantation. PMID- 11267389 TI - Correlation between intraoperative flows and hepatic artery strictures in liver transplantation. PMID- 11267390 TI - Sirolimus (rapamycin)-based rescue treatment following chronic rejection after liver transplantation. PMID- 11267391 TI - Comparison of quality of life and family stress in families of children with living-related liver transplants versus families of children who received a cadaveric liver. PMID- 11267392 TI - Safety limit of small partial liver allografts for orthotopic transplantation. PMID- 11267393 TI - Splenectomy and liver transplantation. PMID- 11267394 TI - Split liver transplantation in Asia. PMID- 11267395 TI - Donor age in liver transplantation: is there a limit? PMID- 11267396 TI - Standard cyclosporine A-based versus completely steroid-free FK506-based immunosuppression after liver transplantation. PMID- 11267397 TI - Necessity and risk of right lobe donor in living donor liver transplantation. PMID- 11267398 TI - Outcome of patients transplanted with liver from hepatitis C positive donors. PMID- 11267399 TI - Use of percutaneous liver biopsies in marginal liver donors. PMID- 11267400 TI - Mortality rate correlated with the number of pediatric liver transplants performed at a center. PMID- 11267402 TI - Experience with radiofrequency ablation of small hepatocellular carcinomas before liver transplantation. PMID- 11267401 TI - Decreasing viral load pretransplant and passive immunoprophylaxis with hepatitis B immunoglobulin posttransplant prevents hepatitis B virus recurrence after liver transplantation: an 8-year single-center experience. PMID- 11267403 TI - Liver transplantation in neurologic Wilson's disease. PMID- 11267405 TI - Are liver grafts from hepatitis B surface antigen negative/anti-hepatitis B core antibody positive donors suitable for transplantation? PMID- 11267406 TI - Does race-matched liver transplantation offer any graft survival benefit? PMID- 11267404 TI - A prospective, randomized trial with quadruple versus dual tacrolimus-based induction after liver transplantation. PMID- 11267407 TI - Delay in diagnosis: a factor in the poor outcome of late acute rejection of liver allografts. PMID- 11267408 TI - Daclizumab induction in liver transplant recipients. PMID- 11267409 TI - Cytokine-producing T lymphocytes as a marker of prognosis and rejection episodes in orthotopic liver transplantation. PMID- 11267410 TI - Leading causes of death in the liver transplant population compared to the corresponding age-matched general US population. PMID- 11267411 TI - Pharmacoeconomic study of tacrolimus-based versus cyclosporine-based immunosuppressive therapy following liver transplantation. PMID- 11267412 TI - Revisiting the use of hepatitis B core antibody-positive donor kidneys. PMID- 11267413 TI - Resection versus transplantation for liver metastases from neuroendocrine tumors. PMID- 11267414 TI - Technical approaches to multivisceral transplantation. PMID- 11267415 TI - Evolution of gastrointestinal transplantation at the University of Miami. PMID- 11267416 TI - Intestinal allografts delay rejection and prolong survival of combined donor specific and third party solid organ transplants. PMID- 11267417 TI - Segmental small bowel transplant from adult living-related donors. PMID- 11267418 TI - Infectious complications following living-related small bowel transplantation in adults. PMID- 11267419 TI - Late severe rejection of intestinal allografts: risks and survival outcome. PMID- 11267420 TI - Multivisceral transplantation for abdominal malignancy: indication, technique, and results in three patients. PMID- 11267421 TI - Genetic modulators of interleukin 1 activity influence the development of chronic rejection in human thoracic allografts. PMID- 11267422 TI - Dendritic cells expressing TGFbeta/IL-10, and CHO cells with OX-2, increase graft survival. PMID- 11267423 TI - TGF-beta1 gene polymorphism is a risk factor for renal dysfunction in heart transplant recipients. PMID- 11267424 TI - Influence of preoperative cyclosporine administration on HLA expression: a comparison of two treatment patterns. PMID- 11267425 TI - Neoral C-2 monitoring in cardiac transplant patients. PMID- 11267426 TI - Lamivudine treatment for HBV infection following thoracic organ transplantation. PMID- 11267427 TI - Cardiac allograft vasculopathy: adventitial immunoreactivity for PDGF-B and PDGFr beta in extra- versus intramural coronary arteries. PMID- 11267428 TI - Ethnic disparities in the pharmacologic characteristics of tacrolimus in heart transplantation. PMID- 11267429 TI - Monoclonal gammopathy in heart transplantation: risk factor analysis and relevance of immunosuppressive load. PMID- 11267430 TI - Expression of murine MD-1 regulates T-Cell activation/cytokine production. PMID- 11267431 TI - Correlation of angiographic and immunohistochemical findings in graft vessel disease after heart transplantation. PMID- 11267432 TI - Photopheresis immunomodulation after heart transplantation. PMID- 11267433 TI - Pediatric heart transplantation: changing indications and improved results. PMID- 11267435 TI - Coadministration of itraconazole and tacrolimus after thoracic organ transplantation. PMID- 11267434 TI - Heart transplantation in patients with inherited myopathies associated with end stage cardiomyopathy: molecular and biochemical defects on cardiac and skeletal muscle. PMID- 11267437 TI - Expression of the vasculoprotective estrogen receptor subtype beta in rat and human cardiac allograft vasculopathy. PMID- 11267436 TI - Mechanical circulatory support of heart transplant patients. PMID- 11267438 TI - A randomized comparison of an immunosuppressive strategy using tacrolimus and cyclosporine in black heart transplant recipients. PMID- 11267439 TI - High rejection score in the first year and risk of skin cancer in heart transplantation. PMID- 11267440 TI - Cytokine and TIRC7 mRNA expression during acute rejection in cardiac allograft recipients. PMID- 11267441 TI - Indirect allorecognition in heart allograft rejection. PMID- 11267442 TI - Racial differences in clinical outcome using tacrolimus and mycophenolate mofetil immunosuppression in heart transplantation. PMID- 11267443 TI - Obliterative bronchiolitis: prevention. PMID- 11267445 TI - Bilateral lung transplant: the procedure of choice for end-stage septic lung disease. PMID- 11267444 TI - Incidence and spectrum of infections in lung transplanted patients: comparison of four different immunosuppressive protocols. PMID- 11267446 TI - Long-term results of lung transplantation for cystic fibrosis. PMID- 11267447 TI - Pulmonary function after living donor lung transplantation. PMID- 11267448 TI - Usefulness of 2D echo Doppler in the preoperative assessment of cystic fibrosis patients who are candidates for lung transplantation. PMID- 11267449 TI - Predicting mortality after lung transplantation. PMID- 11267451 TI - Inhaled areosolized prostacyclin and pulmonary hypertension during anesthesia for lung transplantation. PMID- 11267450 TI - Improved results with lung transplantation for cystic fibrosis. PMID- 11267452 TI - Volumetric monitoring in multiorgan donor and related lung transplant recipients. PMID- 11267453 TI - Report for the international pancreas transplant registry-2000. PMID- 11267455 TI - Long-term outcome of pancreas transplantation. PMID- 11267454 TI - Comparison of enteric versus bladder drainage in pancreas transplantation. PMID- 11267457 TI - Pretransplant immunosuppression for pancreas transplants alone in nonuremic diabetic recipients. PMID- 11267456 TI - Steroid withdrawal for pancreas transplants under tacrolimus immunosuppression. PMID- 11267458 TI - Long-term survival following simultaneous kidney-pancreas transplantation versus kidney transplantation alone in patients with type 1 diabetes mellitus and renal failure. PMID- 11267459 TI - Simultaneous kidney-pancreas transplant with systemic-enteric versus portal enteric drainage. PMID- 11267460 TI - A prospective, randomized, open-label study of steroid withdrawal in pancreas transplantation-a preliminary report with 6-month follow-up. PMID- 11267461 TI - Matching in pancreas transplantation--a registry analysis. PMID- 11267462 TI - National transplantation pregnancy registry: postpregnancy graft loss among female pancreas-kidney recipients. PMID- 11267463 TI - Long-term benefit of kidney-pancreas transplants in type 1 diabetics. PMID- 11267464 TI - Role of surveillance biopsies in monitoring recipients of pancreas alone transplants. PMID- 11267465 TI - Outcome of simultaneous kidney-pancreas transplantation in African-American recipients: a case control study. PMID- 11267466 TI - Enteric versus bladder drainage for solitary pancreas transplants- a registry report. PMID- 11267467 TI - Ten-year survival after simultaneous pancreas/kidney transplantation with bladder drainage and tacrolimus-based immunosuppression. PMID- 11267468 TI - Long-term outcomes in simultaneous kidney-pancreas transplant recipients with portal-enteric versus systemic-bladder drainage. PMID- 11267469 TI - Nine-year experience with 126 pancreas transplants with portal-enteric drainage. PMID- 11267470 TI - Successful simultaneous pancreas-kidney transplantation from well-matched living related donors. PMID- 11267471 TI - Impact of the functioning pancreas on long-term renal function in pancreas-kidney transplantation. PMID- 11267472 TI - A multicenter trial of two daclizumab dosing strategies versus no antibody induction in simultaneous kidney-pancreas transplantation: interim analysis. PMID- 11267473 TI - Long-term prednisolone withdrawal after pancreas and kidney transplantation in patients treated with ATG, tacrolimus, and mycophenolate mofetil. PMID- 11267474 TI - Fate of the pancreas after asynchronous kidney loss in patients undergoing simultaneous kidney/pancreas transplantation. PMID- 11267475 TI - Long-term lipid metabolism in combined kidney-pancreas transplant recipients under tacrolimus immunosuppression. PMID- 11267476 TI - Results of solitary pancreas transplantation with enteric drainage: is there a benefit from monitoring urinary amylase levels? PMID- 11267478 TI - Current limitations of islet transplantation. PMID- 11267479 TI - Challenges toward standardization of islet isolation technology. PMID- 11267477 TI - Limited benefits of induction with monoclonal antibody to interleukin-2 receptor in combination with tacrolimus, mycophenolate mofetil, and steroids in simultaneous kidney-pancreas transplantation. PMID- 11267480 TI - Small bowel subserosal space as a site for islet transplantation and local drug delivery. PMID- 11267481 TI - Microcapsules and their ability to protect islets against cytokine-mediated dysfunction. PMID- 11267483 TI - Basic science foundation for clinical composite tissue transplantation. PMID- 11267482 TI - Islets of Langerhans encapsulated with a tissue-engineered membrane of rat chondrocytes maintain insulin secretion and glucose-insulin feedback for at least 30 days in culture. PMID- 11267484 TI - Peripheral nerve regeneration in human hand transplantation. PMID- 11267485 TI - Immunohistologic studies of the skin of human hand allografts: our experience with two patients. PMID- 11267486 TI - Advances in pediatric transplantation: heart and lung. PMID- 11267488 TI - Depressed oxidative metabolism of polymorphonuclear neutrophils after pediatric liver transplantation. PMID- 11267487 TI - Incidence, management, and outcome of posttransplant lymphoproliferative disease in pediatric liver transplant recipients. PMID- 11267489 TI - Predictive power of ELISA-detected anti-HLA class I antibodies in pediatric kidney transplantation. PMID- 11267490 TI - Risk factors for acute rejection after pediatric heart transplantation. PMID- 11267491 TI - Growth after pediatric heart transplantation. PMID- 11267492 TI - Pediatric liver transplantation for cystic fibrosis. PMID- 11267493 TI - Extrahilar mesenterico-left portal shunt for portal vein thrombosis after liver transplantation. PMID- 11267494 TI - Why is it more difficult to transplant children? A perspective in developing countries. PMID- 11267495 TI - A cause of late graft dysfunction after liver transplantation in children: de novo autoimmune hepatitis. PMID- 11267496 TI - Liver transplantation for hepatoblastoma in the pediatric population. PMID- 11267497 TI - Stem cell transplantation in utero for genetic diseases. PMID- 11267498 TI - Role of interleukin-12 in acute graft-versus-host disease(1). PMID- 11267499 TI - Autoreactive T-Cell subsets in acute and chronic syngeneic graft-versus-host disease. PMID- 11267500 TI - Engraftment of donor-derived stromal cells stimulates fast hematopoietic repopulation of vascularized bone marrow transplant recipients. PMID- 11267501 TI - Clinical relevance of the minor histocompatibility antigen HA-1 in allogeneic bone marrow transplantation between HLA identical siblings. PMID- 11267502 TI - CD4(-)CD8(-) regulatory T cells implicated in preventing graft-versus-host and promoting graft-versus-leukemia responses. PMID- 11267503 TI - Flow cytometric characterization of ex vivo expanded umbilical cord blood CD34(+) cells. PMID- 11267504 TI - Increased content of CD34(+)CD38(-) hematopoietic stem cells in the last collected umbilical cord blood. PMID- 11267505 TI - An ethnic role for chronic, but not acute, graft-versus-host disease after HLA identical sibling stem cell transplantation. PMID- 11267506 TI - The bidirectional relationship between cytomegalovirus and allograft injury. PMID- 11267507 TI - Use of kidneys from anti-HCV positive donors. PMID- 11267508 TI - Differential diagnosis between infection and rejection in renal allografts. PMID- 11267509 TI - Porcine endogenous retrovirus does not infect human cells using a bioartificial liver model system. PMID- 11267510 TI - Prevalence of mycobacterial infection in solid organ transplant recipients. PMID- 11267511 TI - Influenza vaccination among heart transplant recipients. PMID- 11267512 TI - Developing world perspective of posttransplant tuberculosis: morbidity, mortality, and cost implications. PMID- 11267513 TI - Transplantation of kidneys from Anti-HCV positive donors. PMID- 11267515 TI - TNF-alpha and sCD14 as early markers of CMV susceptibility after liver transplantation. PMID- 11267514 TI - Hepatitis C virus infection after renal transplantation: viral load and outcome. PMID- 11267517 TI - Antiviral drugs can inhibit lymphocyte apoptosis induced by cytomegalovirus antigens. PMID- 11267516 TI - Ganciclovir prophylaxis for cytomegalovirus infection in simultaneous kidney pancreas transplant recipients receiving tacrolimus, mycophenolate mofetil, and prednisone. PMID- 11267518 TI - Cytomegalovirus infection-enhanced chronic rejection in the rat is prevented by antiviral prophylaxis. PMID- 11267519 TI - CMV-specific CD8(pos) T lymphocyte differentiation in latent CMV infection. PMID- 11267520 TI - Is preemptive therapy for CMV infection following liver transplantation superior to symptom-triggered treatment? PMID- 11267521 TI - Polyomavirus PCR monitoring in renal transplant recipients: detection in blood is associated with higher creatinine values. PMID- 11267522 TI - Comparison between procalcitonin, serum amyloid A, and C-reactive protein as markers of serious bacterial and fungal infections after solid organ transplantation. PMID- 11267523 TI - Prophylactic antiviral therapy in CMV high-risk liver transplant recipients. PMID- 11267524 TI - Cellular and humoral parameters for vascular damage in blood during cytomegalovirus infections. PMID- 11267525 TI - Preemptive ganciclovir for CMV viremia in liver transplantation. PMID- 11267526 TI - Major infectious complications after kidney transplantation. PMID- 11267527 TI - A single center analysis of malignancies in first kidney graft recipients. PMID- 11267528 TI - Angiotensin II receptor blockade: a novel strategy to prevent immunosuppressant associated cancer progression. PMID- 11267529 TI - Kinetics and development of CMV-accelerated transplant vascular sclerosis in rat cardiac allografts is linked to early increase in chemokine expression and presence of virus. PMID- 11267530 TI - Donor-derived malignancy in organ transplant recipients. PMID- 11267531 TI - Recurrence of cancer after renal transplantation. PMID- 11267532 TI - Posttransplant lymphoproliferative diseases: report from a single center. PMID- 11267533 TI - A gene polymorphism associated with posttransplant lymphoproliferative disorder. PMID- 11267534 TI - Circulating EBV-DNA in the monitoring of EBV infection in pediatric liver transplant recipients. PMID- 11267535 TI - Posttransplant lymphoproliferative disorders: single center experience. PMID- 11267536 TI - Tumor incidence in heart transplant patients: report of the North Italy Transplant Program Registry. PMID- 11267537 TI - UNOS Transplant Tumor Registry: donors with a history of cancer. PMID- 11267538 TI - Predialysis immunosuppression is an independent risk factor for some cancers in renal transplantation. PMID- 11267539 TI - Skin diseases following organ transplantation--risk factors and new therapeutic approaches. PMID- 11267540 TI - Efficacy of computed tomography in detecting hepatocellular cancer in adult liver transplant candidates. PMID- 11267541 TI - A national survey of current practices for diagnosing and treating prostate cancer in transplant candidates. PMID- 11267542 TI - De novo malignancies after organ transplantation. PMID- 11267543 TI - Prospective follow-up of Epstein-Barr virus load in kidney transplant recipients. PMID- 11267544 TI - An allogeneic anti-cancer effect after hematopoietic stem cell transplantation. PMID- 11267545 TI - Effect of mycophenolate mofetil on the Anti-CMV serologic response after renal transplantation. PMID- 11267546 TI - CD8(POS) lymphocyte dynamics in primary CMV infection. PMID- 11267547 TI - Meta-analysis of prophylaxis of CMV disease in solid organ transplantation: is Ganciclovir a superior agent to Acyclovir? PMID- 11267549 TI - Increasing the availability of human organs for transplantation-some ethical issues. PMID- 11267548 TI - Lamivudine in recurrent hepatitis B after renal transplantation. PMID- 11267550 TI - Quality of life issues in transplantation: thoracic organ transplantation. PMID- 11267551 TI - Longitudinal prospective measurement of the quality of life before and after liver transplantation among adults. PMID- 11267553 TI - Medical and ethical issues in xenotransplantation: the opinion of the public, patients, and transplant candidates in Italy. PMID- 11267552 TI - Should culture-based gender bias in living donor renal transplantation be discouraged on ethical and graft survival grounds? PMID- 11267554 TI - Quality of life as a predictor of morbidity, mortality, and resource utilization after solid-organ transplant. PMID- 11267555 TI - Quality-of-life change after kidney transplantation. PMID- 11267556 TI - The quality of life of pancreas recipients with type-1 diabetes. PMID- 11267557 TI - Economic study of renal transplantation: a single-center analysis in Japan. PMID- 11267558 TI - Community promoters for organ donation: scheduling of the programme and assessment. PMID- 11267559 TI - Compliance with hemodialysis and kidney transplantation: a psychotherapeutic perspective. PMID- 11267560 TI - Relationship between donors and pediatric recipients of kidney transplants: a psychosocial study. PMID- 11267561 TI - Health utility scores following renal transplantation. PMID- 11267562 TI - Cost recovery from organ retrieval and implant: state of the art in the Province of Buenos Aires. PMID- 11267563 TI - Cost analysis of operative procedure for transplant patients. PMID- 11267564 TI - Psychodynamic evaluation of kidney transplant patients enrolled in a new immunosuppressive drug trial. PMID- 11267565 TI - Quality of life following solid organ transplantation in Medicaid beneficiaries. PMID- 11267566 TI - Peripheral administration of thymoglobulin for induction therapy in pancreas transplantation. PMID- 11267567 TI - Resuming life after transplantation: development of a support group for patients and their families. PMID- 11267568 TI - Ethics and quality of life of kidney transplant patient. PMID- 11267569 TI - Sexual disorders after heart transplantation. PMID- 11267570 TI - The cost of transplant graft maintenance following solid organ transplantation. PMID- 11267571 TI - Quality of life as a predictor of morbidity, mortality, and resource utilization after solid organ transplant. PMID- 11267572 TI - Economic outcome of simultaneous pancreas kidney transplantation compared with kidney transplantation alone. PMID- 11267573 TI - Quality of life after kidney transplantation--the impact of tacrolimus. PMID- 11267574 TI - Quality of life assessed by SF-36 health survey in renal transplant patients. PMID- 11267575 TI - Transplantation with living donor as ethical dilemma. PMID- 11267576 TI - Biochemical characterization of bioreactors for hybrid liver support: serum-free liver cell coculture of nonparenchymal and parenchymal cells. PMID- 11267577 TI - Improvement of the neurological status of pigs with acute liver failure by hepatocytes immobilized in alginate gel beads inoculated in an extracorporeal bioartificial liver. PMID- 11267579 TI - ECMO support for single lung transplantation. PMID- 11267578 TI - Bioartificial liver support: report of the longest continuous treatment with human hepatocytes. PMID- 11267580 TI - Use of MARS in the treatment of acute liver failure: preliminar monocentric experience. PMID- 11267581 TI - Prolongation of survival of pigs with ischemic liver failure by treatment with a bioartificial liver using glutamine synthetase transfected recombinant HepG2. PMID- 11267582 TI - A tightly regulated immortalized human fetal hepatocyte cell line to develop a bioartificial liver. PMID- 11267583 TI - Cryopreserved porcine hepatocytes in a liver biodialysis system. PMID- 11267584 TI - Improved diffusion properties of a new polysulfone membrane for the development of a bioartificial pancreas. PMID- 11267586 TI - Immune response in the early postoperative period after implantation of a left ventricular assist device system. PMID- 11267585 TI - Safety of continuous human liver support. PMID- 11267587 TI - Induction of T-cell apoptosis by polyurethane biomaterials used in left ventricular assist devices is dependent on calcineurin activation. PMID- 11267588 TI - Induction of CD40 ligand expression in human T cells by biomaterials derived from left ventricular assist device surface. PMID- 11267589 TI - Successful ex vivo liver perfusion system for hepatic failure pending liver regeneration or liver transplantation. PMID- 11267590 TI - Immunologic effects of implantation of left ventricular assist devices. PMID- 11267591 TI - Infectious complications in patients with the Novacor left ventricular assist system. PMID- 11267592 TI - Liberase HI enzyme versus collagenase type P for porcine hepatocyte isolation. PMID- 11267593 TI - Serum cytokines in left ventricular assist device candidates. PMID- 11267594 TI - Valuation of transmission of porcine endogenous retrovirus into patients subjected to hemoperfusion using an extracorporeal bioartificial liver support system. PMID- 11267595 TI - Pancreatic islets microencapsulation with polylactide-co-glycolide. PMID- 11267596 TI - Transplantation and humanism in the new millenium. PMID- 11267597 TI - The 11th report of the Latin American Transplant Registry: 62,000 transplants. PMID- 11267599 TI - South Mediterranean, Middle East, and subcontinent organ transplantation activity. PMID- 11267598 TI - Asian transplant registry (1999). PMID- 11267600 TI - The success of laparoscopic donor nephrectomy: is the open approach justified? PMID- 11267602 TI - Mini-invasive treatment of arterial and biliary complications after orthotopic liver transplantation. PMID- 11267601 TI - Morphologic changes in renal procurement biopsy and onset of graft function. PMID- 11267603 TI - The results of live related renal transplantation in black patients. PMID- 11267604 TI - Induction of anti-tumor activity following autologous stem cell transplantation: immunotherapeutic implications. PMID- 11267605 TI - Dyslipidemia and dietary modification in Chilean renal pediatric transplantation. PMID- 11267606 TI - The effect of losartan on posttransplant erythrocytosis. PMID- 11267607 TI - Urologic complications in 1275 consecutive renal transplantations. PMID- 11267608 TI - Serum troponin T in the early posttransplant period and long-term graft function in heart recipients. PMID- 11267609 TI - Management of acute necrotizing pancreatitis after renal transplantation. PMID- 11267610 TI - Effect of cyclosporine withdrawal on IL-10 production in kidney transplant recipients. PMID- 11267611 TI - Comparative study of helical CT scan angiography, conventional arteriography, and intraoperative findings for the evaluation of living renal transplant donors. PMID- 11267612 TI - Silver clip ligation technique for liver resection. PMID- 11267613 TI - Preliminary results of a prospective randomized study of basiliximab in kidney transplantation. PMID- 11267614 TI - En bloc and single kidney transplantation from donors weighing less than 15 kg into pediatric recipients. PMID- 11267615 TI - Angiotensin-converting enzyme inhibitors reduce hemoglobin concentrations, hematocrit, and serum erythropoietin levels in renal transplant recipients without posttransplant erythrocytosis. PMID- 11267616 TI - The health of living kidney donors 20 years after donation. PMID- 11267617 TI - Long-term survival of a lung cancer patient after kidney transplantation. PMID- 11267618 TI - Ureteral stenosis after kidney transplantation: interventional radiology or surgery? PMID- 11267619 TI - Organizacion Nacional de Trasplante de Venezuela: a new initiative to approach donation and transplantation in our country. PMID- 11267620 TI - Folic acid supplementation and homocyst(e)ine level in renal transplant recipients. PMID- 11267621 TI - MK801 increases feeding and decreases drinking in nondeprived, freely feeding rats. AB - The noncompetitive NMDA receptor antagonist MK801 has been reported to increase food intake in rats during scheduled test meals of palatable foods or after food deprivation, but not in nondeprived rats given rodent chow. To determine if MK801 has an effect on spontaneous meals, MK801 (100 microg/kg) was administered 15 min prior to dark onset to nondeprived rats maintained on powdered rodent chow, and spontaneous food and water access was measured. MK801 increased the length of the first meal and the amount of time spent feeding within the meal. Conversely, MK801 decreased the length and size of the first drinking bout. MK801 did not alter the latency to the first meal or drinking bout, nor the intervals between successive meals or bouts. The effects of MK801 on feeding and drinking bouts were partially confirmed by measuring total chow and water intake over the first 2 h of the dark period. Thus, acute MK801 can significantly alter spontaneous chow feeding and drinking in nondeprived rats when administered prior to dark onset. PMID- 11267623 TI - Anticonvulsant properties of N-salicyloyltryptamine in mice. AB - A new tryptamine analogue, N-salicyloyltryptamine (STP), a potential central nervous system (CNS) depressant, was tested in the pentylenetetrazol (PTZ) and maximal electroshock (MES) models of epilepsy in mice. When administered concurrently, STP (100 mg/kg ip) significantly reduced the number of animals that exhibited PTZ-induced seizures and eliminated the extensor reflex of maximal electric-induced seizures test in 50% of the experimental animals. In addition, it showed protection in the PTZ test by diminishing the death rate. PMID- 11267622 TI - Acute effects of nicotine and mecamylamine on tobacco withdrawal symptoms, cigarette reward and ad lib smoking. AB - Separate and combined effects of nicotine and the nicotinic antagonist mecamylamine were studied in 32 healthy volunteer smokers after overnight abstinence from smoking. Subjects participated in three sessions (3 h each), during which they wore skin patches delivering either 0 mg/24 h, 21 mg/24 h or 42 mg/24 h nicotine. Thirty-two subjects were randomly assigned to two groups receiving oral mecamylamine hydrochloride (10 mg) vs. placebo capsules. Two and one-half hours after drug administration, subjects were allowed to smoke ad lib, rating the cigarettes for rewarding and aversive effects. Transdermal nicotine produced a dose-related reduction in the subjective rewarding qualities of smoking. Nicotine also reduced craving for cigarettes and this effect was attenuated, but not eliminated, by mecamylamine. Mecamylamine blocked the discriminability of high vs. low nicotine puffs of smoke, and increased nicotine intake substantially during the ad lib smoking period. Some of the psychophysiological effects of each drug (elevation in blood pressure from nicotine, sedation and decreased blood pressure from mecamylamine) were offset by the other drug. The results supported the hypothesis that nicotine replacement can alleviate tobacco withdrawal symptoms even in the presence of an antagonist such as mecamylamine. Mecamylamine did not precipitate withdrawal beyond the level associated with overnight cigarette deprivation, suggesting its effects were primarily due to offsetting the action of concurrently administered nicotine as opposed to blocking endogenous cholinergic transmission. PMID- 11267625 TI - Chronic sucrose intake enhances nicotine-induced antinociception in female but not male Long-Evans rats. AB - Previous work has demonstrated that intake of palatable foods can alter the behavioral actions of opioid drugs. To investigate whether intake of palatable fare only affects opioid-induced behaviors or more generally influences drug induced responses, this study examined the effects of chronic intake of a palatable sucrose solution on nicotine-induced antinociception. Eight male and eight female Long-Evans rats were provided with ground chow and water (control group), while eight males and eight females were provided with chow, water and a 32% sucrose solution (sucrose group). After 3 weeks of exposure to the dietary conditions, all rats were tested for nicotine-induced antinociception using the tail flick test. Nicotine, administered using a cumulative dose regime (0.03, 0.1, 0.3 and 1.0 mg/kg sc), led to dose-dependent increases in tail flick latencies in male and female rats. Females in the sucrose group displayed significantly greater antinociceptive responses to nicotine than those in the control group. Similar results were obtained when females were retested after an additional 2 weeks. Comparison of males and females, revealed that sucrose enhanced nicotine's antinociceptive action in female but not in male rats. While previous research suggested that sweet tasting substances might affect drug action by acting on the endogenous opioid system, the present results indicate that sucrose intake could also alter the cholinergic system and possibly other systems involved in nicotine antinociception. PMID- 11267624 TI - Exogenous cortisol exerts effects on the startle reflex independent of emotional modulation. AB - Exogenous cortisol's modulation of the acoustic startle reflex (ASR) was tested alone and during exposure to affectively valenced photographs in healthy men and women. During nonmodulated startle, oral hydrocortisone had a biphasic dose effect, with 5 mg increasing and 20 mg decreasing, eyeblink reflex magnitude compared to placebo. During emotion modulation, 20 mg of hydrocortisone reduced reflex magnitude without affecting the usual pattern of modulation across positive, neutral, and negatively affective slides. Gender differences were not found in either relationship. These findings illustrate dose-dependent effects of cortisol on the startle pathway independent of emotional state and consistent across genders. PMID- 11267626 TI - Estrogen modulates 5-HT(1A) agonist inhibition of lordosis behavior but not binding of [(3)H]-8-OH-DPAT. AB - Previous studies showed that repeated estrogen treatment reduces the ability of the 5-HT(1A) receptor agonist, 8-hydroxy-2(di-n-propylamino) tetralin (8-OH DPAT), to inhibit lordosis behavior of female rats. The present study evaluated the effects of repeated estrogen treatment on lordosis behavior and 5-HT(1A) receptor binding and coupling to G protein in the hypothalamus-preoptic area using the agonist ligand [3H]-8-OH-DPAT, which binds selectively to G-protein coupled 5-HT(1A) receptors. Rats were injected twice with 25 or 50 microg of estradiol benzoate (EB) 7 days apart followed by 500 microg of progesterone (P) 48 h after the second EB injection. Controls received a single injection of 25 or 50 microg EB followed 48 h later by 500 microg of P. Four hours after P, 0.15 mg/kg 8-OH-DPAT was injected, and lordosis behavior examined for 30 min. Rats treated twice with EB showed significantly less 8-OH-DPAT inhibition of lordosis behavior than rats receiving a single EB injection. For receptor binding, rats received EB without P treatment. None of the estrogen treatments reduced [3H]-8 OH-DPAT binding density or affinity in the hypothalamus-preoptic area or hippocampus. These studies suggest that estrogen modulates 5-HT(1A) agonist potency without a measurable change in 5-HT(1A) receptor density or coupling to G protein. PMID- 11267627 TI - Bromocriptine-induced dissociation of hyperglycemia and prolactin response to restraint. AB - The present study investigated the effects of immobilization (restraint stress) on rat chronically treated with a D(2) receptor agonist (bromocriptine, 0.4 mg/100 g body weight, injected daily intraperitoneally (ip) for 2 weeks) on plasma glucose, prolactin, and insulin levels. During restraint, the plasma prolactin of vehicle-treated (VEH) rats increased rapidly, reaching a peak at 10 min (57.9 +/- 8.1 ng/ml, P < .01). In contrast, restraint failed to induce any significant change in the plasma prolactin levels of bromocriptine-treated (BR) rats. The hyperglycemic response to immobilization was 97% higher (P < .05) in BR rats than in VEH rats. Our data demonstrate that prolactin secretion and hyperglycemia in response to restraint can be dissociated by chronic treatment with BR, which also increased the hyperglycemic response to immobilization probably due to central D(2) dopaminergic activity. PMID- 11267628 TI - Effects of diazepam and beta-CCM on working memory in mice: relationships with emotional reactivity. AB - This study was aimed at determining the effects of systemic administration of diazepam and methyl beta-carboline-3-carboxylate (beta-CCM) both on spatial working memory and on emotional reactivity in mice. Results showed that diazepam and beta-CCM induced opposite effects in both memory and emotional reactivity tests. Indeed, as a function of dose, diazepam reduced anxiogenic-like reactions but increased vulnerability to interference in the memory task at a 30-s but not at a 5-s delay interval. As a function of dose, beta-CCM reduced vulnerability to interference and increased emotional reactivity, these effects being antagonised by concurrent administration of flumazenil (RO 15-1788). Thus, our study showed the bidirectional effects of these two drugs on a spatial working memory task involving a spontaneous processing of information and suggested a direct link between the emotional effects of the drugs and memory performance. PMID- 11267629 TI - Chronic inositol increases striatal D(2) receptors but does not modify dexamphetamine-induced motor behavior. Relevance to obsessive-compulsive disorder. AB - A large body of evidence suggests that the neuropathology of obsessive-compulsive disorder (OCD) lies in the complex neurotransmitter network of the cortico striatal-thalamo-cortical (CSTC) circuit, where dopamine (DA), serotonin (5HT), glutamate (Glu), and gamma-amino butyric acid (GABA) dysfunction have been implicated in the disorder. Chronic inositol has been found to be effective in specific disorders that respond to selective serotonin reuptake inhibitors (SSRIs), including OCD, panic, and depression. This selective mechanism of action is obscure. Since nigro-striatal DA tracts are subject to 5HT(2) heteroreceptor regulation, one possible mechanism of inositol in OCD may involve its effects on inositol-dependent receptors, especially the 5HT(2) receptor, and a resulting effect on DA pathways in the striatum. In order to investigate this possible interaction, we exposed guinea pigs to oral inositol (1.2 g/kg) for 12 weeks. Subsequently, effects on locomotor behavior (LB) and stereotype behavior (SB), together with possible changes to striatal 5HT(2) and D(2) receptor function, were determined. In addition, the effects of chronic inositol on dexamphetamine (DEX)-induced motor behavior were evaluated. Acute DEX (3 mg/kg, ip) induced a significant increase in both SB and LB, while chronic inositol alone did not modify LA or SB. The behavioral response to DEX was also not modified by chronic inositol pretreatment. However, chronic inositol induced a significant increase in striatal D(2) receptor density (B(max)) with a slight, albeit insignificant, increase in 5HT(2) receptor density. This suggests that D(2) receptor upregulation may play an important role in the behavioral effects of inositol although the role of the 5HT(2) receptor in this response is questionable. PMID- 11267631 TI - Behavioral analysis of stress controllability effects in a new swim stress paradigm. AB - Previous animal stress studies have illustrated the marked impact of coping on subsequent behavior and physiology by using shock as the stressor. The current study evaluates the generality of shock stress controllability effects in a new swim stress paradigm on several dependent measures: behavioral despair, analgesia, shuttlebox escape, and alcohol reactivity. In this new paradigm, rats in the escape group are able to learn the behavioral response as evidenced by significant reduction in the acquisition of a lever press response. Both escape and yoked subjects showed "behavioral despair" in comparison to both restrained and home cage controls when tested 24 h later. In the standard shuttlebox escape task 24-h post-stress, no group differences emerged, although a trend for poorer performance in the yoked subjects was evident. No group differences were observed in pain sensitivity after the first or second forced swim exposure. Finally, stress controllability effects were observed in behavioral reactivity to alcohol 2-h post-stress as measured by rotarod performance. This effect is opposite to the previous observations with the tailshock stress controllability paradigm. These results suggest that (1) there are certain similarities, but some fundamental differences between the behavioral endpoints measured following intermittent swim stress in comparison to the well-established effects of the intermittent tailshock stress model and (2) the qualitative nature of a stressor may markedly influence the behavioral and physiological consequences of stress and coping. PMID- 11267630 TI - Behavioral effects of buspirone in the marmoset employing a predator confrontation test of fear and anxiety. AB - In order to further validate the recently developed marmoset (Callithrix penicillata) predator confrontation model of fear and anxiety, we investigated the behavioral effects of buspirone with this method. The apparatus consisted of three parallel arms connected at each end to a perpendicular arm, forming a figure-eight continuous maze. A taxidermized wild oncilla cat (Felis tigrina) was positioned facing a corner of the parallel arms, alternating between the left or right side of the maze among animals tested. All subjects were first submitted to seven 30-min maze habituation trials (HTs) in the absence of the predator, and then to five randomly assigned treatment trials (TTs) in the presence of the predator: three buspirone sessions (0.1, 0.5 and 1.0 mg/kg), saline and sham injection controls. Twenty minutes after treatment administration, the animal was released into the maze and had free access to the apparatus for 30 min. All trials were taped for later behavioral analysis. Buspirone significantly decreased the frequency of scent marking, while increasing the time spent in proximity to the 'predator' stimulus, indicating an anxiolytic effect. Neither locomotor activity, exposure to a novel environment, stimulus location and habituation, nor gender influenced the effects of the drug treatments. These results further validate this method and demonstrate the potential usefulness of this ethologically based paradigm to test anxiety and fear-induced avoidance in nonhuman primates and its susceptibility to anxiolytic pharmacological manipulations. PMID- 11267632 TI - Separation-induced body weight loss, impairment in alternation behavior, and autonomic tone: effects of tyrosine. AB - We have investigated the effects of tyrosine on alternation behavior and hippocampal adrenergic and cholinergic tone in a model of self-induced weight loss caused by separation stress. Separation decreased body weight in mice (P < .001) and spontaneous alternations in the T-maze (P < .001). This impairment was associated with depletion of both norepinephrine (NE, P < .001) and dopamine (P < .01) while increasing MHPG (P < .05) and the ratio of MHPG/NE (P < .05). Increasing tyrosine availability restored performance to control levels (P < .001) and repleted dopamine (P < .05) and presumably also NE (indicated by increases in both MHPG, P < .001, and MHPG/NE, P < .05). Stress increased adrenergic alpha(2)-receptor density (P < .001) without changing its K(d) and the B(max) and K(d) of beta-receptors, suggesting that it decreased NE transmission through action on alpha(2)-receptors. The balance between beta- and alpha(2) receptors appeared to be related to alternation behavior as shown by the decrease (P < .01) and increase (P < .05) in their ratios induced by stress and tyrosine, respectively. With regard to cholinergic tone, separation stress increased M1 receptor density (P < .05) and its mRNA signal (P < .001). Tyrosine further increased M1 receptor density of stressed mice (P < .05). Tyrosine might be a potential therapy for cognitive and mood problems associated with the maintenance of a reduced body weight in the treatment of obesity and in the extreme case of anorexia nervosa. PMID- 11267633 TI - The possible cross-tolerance between morphine- and nicotine-induced hypothermia in mice. AB - In the present study, cross-tolerance between hypothermia induced by morphine and nicotine in mice has been investigated. Different doses of morphine or nicotine induced dose-dependent hypothermia. The sub-maximal doses of both drugs were used for interaction studies. Administration of mecamylamine either intracerebroventricularly (2-6 microg/animal icv) or intraperitoneally (0.5 and 1 mg/kg ip) decreased both morphine- or nicotine-induced hypothermia. Naloxone either intracerebroventricularly (2-6 microg/animal) or intraperitoneally (1 and 2 mg/kg) reduced the response to morphine, but not nicotine's response. Hexamethonium (5 and 10 mg/kg ip) caused a slight decrease in morphine's hypothermia, but not that of nicotine. Nicotine's response was decreased in the animals which were made tolerant to hypothermic effect of morphine. Pre-treatment of the animals with low doses of morphine (12.5 or 25 mg/kg), once daily for 3 days, did not cause significant tolerance to the hypothermic response to morphine or nicotine. However, the administration of low doses of morphine (12.5 or 25 mg/kg) plus nicotine (2 mg/kg), once daily for 3 days, increased levels of tolerance to hypothermia induced by either drug. It is concluded that nicotinic receptor mechanism may play a role in morphine-induced hypothermia and there is cross-tolerance between the two drugs. PMID- 11267634 TI - The effect of gamma-vinyl-GABA on the consumption of concurrently available oral cocaine and ethanol in the rat. AB - It has frequently been reported that a high percentage of individuals, identified as either alcohol- or cocaine-dependent, concurrently abuse both drugs. The experiments reported here represent a continuing effort to develop an animal model to predict the effects of a potential pharmacotherapeutic agent on concurrently available oral ethanol and cocaine. These experiments utilized drinkometer circuitry to assess the effects of gamma-vinyl-GABA (GVG), a gamma aminobutyric acid (GABA) transaminase inhibitor, on the consumption and temporal pattern of responses for orally self-administered ethanol and cocaine. The results of these experiments showed that GVG, at doses of 100, 200 and 300 mg/kg, reduced both ethanol and cocaine consumption in a dose-related manner. When compared to vehicle, GVG at all doses significantly reduced ethanol consumption while consumption of cocaine was significantly reduced only at 300 mg/kg. This is consistent with data showing that GVG reduces consumption of these drugs when administered alone and data showing that GVG is more potent in reducing ethanol induced compared to cocaine-induced extracellular dopamine in the nucleus accumbens. Analysis of the temporal pattern of drinking across the session suggests that GVG's effects are due to a disruption of the reinforcing properties of ethanol and cocaine rather than a more general reduction in motor behavior. These data suggest that GVG has potential for clinical use in populations that abuse either alcohol or cocaine alone or in combination. PMID- 11267635 TI - Modulation of cocaine's discriminative stimulus effects by dopamine D(1) agonists in rhesus monkeys. AB - Dopamine (DA) D(1) agonists are classified as high- or low-efficacy on the basis of in vitro functional measures as compared to DA. In monkeys self-administering cocaine, high-efficacy D(1) agonists have been shown to have reinforcing effects, while low-efficacy agonists do not. However, the relationship between D(1) agonist efficacy and cocaine-like discriminative stimulus effects, particularly in rhesus monkeys, is not clear. The present study investigated the discriminative stimulus effects of a high- (SKF 81297) and a low-efficacy (SKF 38393) D(1) agonist in rhesus monkeys (n=4) trained to discriminate cocaine from saline using a two-lever drug discrimination procedure. In a second experiment, the effects of agonist pretreatments, as well as pretreatment with a D(1) antagonist, on cocaine's discriminative stimulus effects were evaluated. SKF 81297 (0.01-1.7 mg/kg) fully substituted for cocaine in three of four animals (> 80% cocaine-appropriate responding), while SKF 38393 (0.3-10 mg/kg) occasioned < 50% cocaine-appropriate responding in all subjects. When given as a pretreatment, neither agonist altered cocaine's discriminative stimulus effects at the doses tested. In contrast, the D(1) antagonist SCH 23390 attenuated cocaine's discriminative stimulus effects. These results indicate that D(1) agonists have cocaine-like discriminative stimulus effects in rhesus monkeys that are consistent with their in vitro efficacies. However, when given in combination with cocaine, D(1) agonist efficacy does not appear to be a major factor in modifying cocaine's discriminative stimulus effects. PMID- 11267637 TI - Tolerance to nicotine's effects in the elevated plus-maze and increased anxiety during withdrawal. AB - In the elevated plus-maze test of anxiety, nicotine (0.1 mg/kg sc; 30 min after injection) had a significant anxiogenic effect, shown by specific decreases in the percentage of time spent on the open arms and in the percentage of open-arm entries. Tolerance developed to this anxiogenic effect after 7 days of nicotine treatment (0.1 mg/kg/day). Five minutes after an acute injection, nicotine (0.1 mg/kg) was ineffective, but after 7 days of treatment a significant anxiolytic effect, shown by specific increases in the percentage of time spent on the open arms and in the percentage of open-arm entries, emerged. After 14 days of nicotine treatment, tolerance developed to this anxiolytic effect. There was a complete dissociation between the effects of nicotine on the measures of anxiety, and on the locomotor activity as measured by closed-arm entries. No changes in closed-arm entries were found after acute administration of nicotine, but rats tested 30 min after their 7th injection made significantly fewer, and those tested 5 min after their 14th injection made significantly more, entries than their respective controls. Rats that were tested after 24 h withdrawal from six daily nicotine injections showed a significant anxiogenic effect. A low dose of nicotine (5 ng) injected into the dorsal hippocampus was without effect in vehicle pretreated rats, but it was able to reverse the anxiogenic effect found after 24 h of withdrawal from 6 days of nicotine treatment. PMID- 11267636 TI - Cannabinoids of diverse structure inhibit two DOI-induced 5-HT(2A) receptor mediated behaviors in mice. AB - We have recently shown that the selective cannabinoid CB(1) receptor antagonist SR 141716A produces robust frequencies of head-twitch response (HTR) and ear scratch response (ESR) in drug-naive mice. Both behaviors were potently blocked by the selective 5-HT(2A/C) receptor antagonist SR 46349B. Selective 5-HT(2A/C) agonists such as DOI also produce these behaviors in mice. The purpose of the present study was to: (1) investigate whether Delta(9)-tetrahydrocannabinol (Delta(9)-THC) and its analogs [Delta(8)-tetrahydrocannabinol (Delta(8)-THC), HU 210, CP 55,940, and WIN 55,212-2] can prevent the DOI-induced behaviors and (2) to see whether any correlation exists in the ID(50) potency order of these cannabinoids in inhibiting the DOI-induced HTR and ESR relative to their published ED(50) potency profiles in producing the tetrad of behaviors in mice. Thus, at 0 min, different groups of mice were injected intraperitoneally with either vehicle or varying doses of the following cannabinoids: Delta(9)-THC (0.25 20 mg/kg), Delta(8)-THC (2.5-20 mg/kg), HU-210 (0.02-0.5 mg/kg), CP 55,940 (0.004 0.5 mg/kg), and WIN 55,212-2 (0.5-10 mg/kg). Twenty minutes later, each mouse received an intraperitoneal injection of DOI (1 mg/kg) and the frequencies of DOI induced behaviors (mean +/- S.E.M.) were recorded for the next 20 min. The tested cannabinoids reduced the frequencies of both DOI-induced HTR and ESR in a dose dependent fashion. HU-210 was the most potent inhibitor of HTR, whereas CP 55,940 was most effective against ESR. The ID(50) potency order of cannabinoids in blocking the HTR is: HU-210 > CP 55,940 > WIN 55,212-2 > Delta(9)-THC > Delta(8) THC, which is identical to their published order of potency in producing the tetrad of behaviors in mice. On the other hand, they had the following ID(50) potency order against the ESR: CP 55,940 > HU-210 > WIN 55,212-2 > Delta(9)-THC > Delta(8)-THC. The tested cannabinoids were 3-30 times more potent in preventing the ESR than the HTR. The data show that cannabinoids inhibit 5-HT(2A) receptor mediated functions in a potent but differential manner. PMID- 11267638 TI - Flavor preferences conditioned by intragastric infusion of ethanol in rats. AB - Sprague-Dawley rats were trained 22 h/day to associate a flavored solution [conditioned stimulus (CS+)] with intragastric infusions of 6% ethanol and another flavored solution (CS-) with water infusions. The infusions were matched to the CS intakes so that the animals determined their timing and size. In Phase 1, chow and water were available ad libitum, and both CS flavors were initially sweetened with saccharin that was then faded out. The rats displayed a preference for the CS+ over the CS- under both reinforced and extinction conditions. When food-restricted in Phase 2, the rats displayed an increased preference for the CS+. In Phase 3, the rats were fed ad libitum chow and given preference tests with the CS+ paired with ethanol infusions of increasing concentration (6%, 12%, 18%, and 24%). Their preference for the CS+ over the CS- persisted, and self administered ethanol dose increased with concentration to 5 g/kg/day. The ethanol based conditioned flavor preference resembled those conditioned by carbohydrate and fat infusions, suggesting that at least some of reinforcing ability of ethanol may be related to its postingestive nutritive effects. PMID- 11267640 TI - Increased m-CPP-induced oral dyskinesia after lesion of serotonergic neurons. AB - Peripheral administration of the 5-hydroxytryptamine (5-HT)(2C/1B) agonist 1-(m chlorophenyl)piperazine (m-CPP) produces abnormal orofacial movements in rats. We have previously shown that this behavior is mediated by 5-HT(2C) receptors in the subthalamic nucleus [Neuroscience 72 (1996) 117]. The present studies examined this effect after serotonin depletion to determine whether removal of endogenous serotonin affected this behavioral response and/or subthalamic 5-HT(2C) receptors. Rats received an intraventricular infusion of the neurotoxin 5,7 dihydroxytryptamine (5,7-DHT, 100 mg/10 ml) or vehicle after desipramine pretreatment (25 mg/kg ip). The efficacy of serotonin depletion was confirmed by a decrease in serotonin uptake sites measured by autoradiography. Oral dyskinesia induced by peripheral administration of m-CPP (1.0 mg/kg ip) was markedly increased in lesioned rats compared to sham-operated controls 4 and 8 but not 12 days after the lesion. A subset of lesioned rats that displayed transient seizures after m-CPP injection did not prevent the measurement of oral dyskinesia during the observation period. No differences in 5-HT(2C) receptor levels were found with ligand-binding autoradiography in the subthalamic nucleus, or in other brain regions that express this receptor, in rats sacrificed 5 days following 5,7 DHT lesions. The data indicate that lesion of serotonergic neurons in adult rats induces a transient increase in motor responses mediated by subthalamic 5-HT(2C) receptors. These data suggest that functional alterations in serotonergic transmission in the subthalamic nucleus may be involved in the pathophysiology of hyperkinetic movement disorders. PMID- 11267639 TI - Diazepam during prior ethanol withdrawals does not alter seizure susceptibility during a subsequent withdrawal. AB - The number of cycles of alcohol detoxification is suggested to be an important variable in the predisposition to severe withdrawal seizures in alcohol-dependent individuals. Several clinical studies have suggested that exposure to repeated alcohol withdrawals may lead to increased severity of subsequent withdrawal episodes. Consistent with these observations, exposure to multiple cycles of ethanol withdrawal in our previous study significantly increased sensitivity to the convulsive effects of the GABA(A) receptor inverse agonist, Ro15-4513, in comparison to continuous ethanol exposure with no intermittent withdrawals. There was also a selective increase in the occurrence of spontaneous spike and sharp wave (SSW) activity in the EEG recorded from hippocampal area CA(3) in proportion to the number of withdrawal episodes experienced. It is hypothesized that during such repeated episodes of ethanol intoxication and withdrawal, changes in neuronal excitation during prior withdrawals could serve as initially subconvulsive kindling stimuli that might eventually result in the increased severity of the withdrawal syndrome. There is some evidence of the successful suppression of such neuronal excitation during acute ethanol withdrawal by positive modulators of the GABA(A) receptor. In the present study, the benzodiazepine agonist, diazepam, at a dose (4.0 mg/kg) that suppresses acute withdrawal symptoms, when administered during intermittent withdrawals, did not alter seizure sensitivity during a subsequent nonmedicated withdrawal. Diazepam treatment during prior withdrawals also did not have any effect on the multiple withdrawal-associated increase in SSW activity in hippocampal area CA(3) during an untreated withdrawal. This finding suggests that suppression of acute withdrawal symptoms by diazepam does not prevent long-lasting changes in CNS function resulting from repeated exposures to ethanol withdrawal. PMID- 11267641 TI - A role for serotonin in lipopolysaccharide-induced anorexia in rats. AB - Rats consistently reduce their food intake following injection of bacterial lipopolysaccharides (LPS). Because LPS increases CNS serotonin (5-HT) turnover, and because increases in CNS 5-HT turnover are associated with a decrease in food intake, we conducted a series of studies to examine 5-HT's potential role in LPS induced anorexia. Chronic CNS 5-HT depletion by cisterna magna (CM) administration of 5,7-dihydroxytryptamine (5,7-DHT) failed to attenuate LPS induced (100 microg/kg, ip) anorexia. In subsequent experiments, LPS was injected at lights out (hour 0) and [8-hydroxy-2-(di-n-propylamino)tetraline (8-OH-DPAT)] or N-CBZ-[(8beta)-1,6-dimethylergolin-8-yl]methylamine (metergoline) was injected at hour 5 - the time when LPS-treated rats become anorectic. Food intake was measured during the subsequent 2 h. In LPS-treated rats, 8-OH-DPAT (62.5, 125, or 250 microg/kg, sc) injection increased food intake. In a 2 x 2 factorial arrangement of LPS and 8-OH-DPAT, 125 microg/kg 8-OH-DPAT increased food intake significantly more in LPS-treated rats than in non-LPS-treated rats (significant LPS x 8-OH-DPAT interaction). In LPS-treated rats, 1 and 5 mg/kg metergoline significantly enhanced food intake. However, in a 2 x 2 arrangement of LPS and metergoline, 1 mg/kg metergoline failed to increase food intake in LPS and non LPS-treated rats in two separate trials. The ability of the 5-HT(1A) receptor agonist 8-OH-DPAT to attenuate LPS-induced anorexia in rats supports a role of 5 HT in LPS-induced anorexia. PMID- 11267643 TI - Purification and characterization of a Bacillus cereus exochitinase. AB - Five extracellular chitinases of Bacillus cereus 6E1 were detected by a novel in gel chitinase assay using carboxymethyl-chitin-remazol brilliant violet 5R (CM chitin-RBV) as a substrate. The major chitinase activity was associated with a 36 kDa (Chi36) gel band. Chi36 was purified by a one-step, native gel purification procedure derived from the new in-gel chitinase assay. The purified Chi36 has optimal activity at pH 5.8 and retains some enzymatic activity between pH 2.5-8. The temperature optimum for Chi36 was 35 degrees C, but the enzyme was active between 4-70 degrees C. Based on its ability to hydrolyze mainly p-nitrophenyl-(N acetyl-beta-D-glucosaminide)(2), Chi36 is characterized as a chitobiosidase, a type of exochitinase. The N-terminal amino acid sequence of mature Chi36 was determined (25 amino acids). Alanine is the first N-terminal amino acid residue indicating the cleavage of a signal peptide from a Chi36 precursor to form the mature extracellular Chi36. The N-terminal sequence of Chi36 demonstrated highest similarity with Bacillus circulans WL-12 chitinase D and significant similarity with several other bacterial chitinases. PMID- 11267642 TI - Molecular design, expression, and affinity immobilization of a trypsin streptavidin fusion protein*(1). AB - A trypsin-streptavidin (TRYPSA) fusion protein was designed and its expression in Escherichia coli was evaluated. The streptavidin gene was PCR modified and cloned into the pET expression vector. The trypsin gene was subsequently inserted into this plasmid, thus generating a colinear fusion of trypsin and streptavidin genes (pTRYPSA). This engineering strategy was verified, and TRYPSA was expressed after IPTG induction using the E. coli strains, BL21(DE3) and BL21(DE3)pLysS. Standard protein fractions of the cell lysate were prepared and trypsin activity was primarily detected in the periplasmic and inclusion body fractions. Immunoblotting showed a single Western-positive band exhibiting a molecular weight of 39,000 Da. A biotinylated porous glass affinity matrix was prepared and selective adsorption resulted in a one-step purification and immobilization of TRYPSA from crude cell lysate. Trypsin activity was verified using a synthetic substrate. This enzyme bioreactor should serve as an excellent prototype for future studies that will examine the effect of limited proteolysis on functional characteristics of milk proteins, including gelling, emulsifying and foaming properties. PMID- 11267644 TI - Combined effect of operational variables and enzyme activity on aqueous enzymatic extraction of oil and protein from soybean. AB - The individual effect of two different enzymes-protease and cellulase-on oil and protein extraction yields combined with other process parameters-enzyme concentration, time of hydrolysis, particle size and solid-to-liquid ratio-was evaluated by Response Surface Methodology. The selection of the enzymes for the study was based on preliminary experiments that showed higher increments in the extraction yield with the use of the two enzymes when compared to hemicellulase and pectinase. The levels of the quantitative parameters studied were: i) enzyme concentration: 0.1, 0.45, 2 w/w %; ii) liquid-to-solid ratio: 0.05, 0.125, 0.2; iii) mean particle size: 212.5, 449.5, 855 um; iv) time of hydrolysis: 30; 60; 120 min. Experimental data for both oil and protein extraction yields obtained with and without enzymes correlated very well with process parameters (P < 0.0001), resulting in models with high coefficient of determination for oil and protein extraction yields (r(2) = 0.9570 and r(2) = 0.9807, respectively). The use of protease resulted in significantly higher yields over the control (protein yield increased from 27.8 to 66.2%, oil yield increased from 41.8 to 58.7%) only when heat treated flour was used, or when non-heat treated flour with large particle sizes was used in the extraction. The yields of protein and oil from non heat treated material in general decreased slightly with the use of enzymes. PMID- 11267645 TI - Diffusion of (de)acylated antibiotic A40926 in alginate and carrageenan beads with or without cells and/or soybean meal. AB - Effective diffusion coefficients (D(e)) of antibiotic A40926 and its deacylated derivative were determined in Ca-alginate (2% wt/wt) and kappa-carrageenan (2.6% wt/wt) gel beads with or without immobilized Actinoplanes teichomyceticus cells and/or soybean meal (SBM). The method used was based on transient concentration changes in a well-stirred antibiotic solution in which gel beads, initially free of solute, were suspended. Unsteady-state diffusion in a sphere was applied and D(e) determined from the best fit of experimental data. A40926 showed markedly different diffusion characteristics than its deacylated derivative. Diffusivity of deacyl-A40926 in alginate or carrageenan gel beads was six to seven times that of A40926. Large differences in partition coefficients (Kp) were also found. In case of beads without additions, A40926, in contrast to deacyl-A40926, strongly partitioned to the liquid phase. Introduction of SBM and/or mycelium in the gel beads decreased the effective diffusivity of deacyl-A40926, but increased its partitioning to the solid phase. Our findings indicate that a relatively moderate structural change of a lipoglycopeptide molecule could lead to a major change in its diffusion/partition characteristics. PMID- 11267647 TI - Evaluating the basidiomycetes Poria medula-panis and Wolfiporia cocos for xylanase production. AB - Xylanase, oxidative enzymes and iron-binding compounds were detected in the filtrates of Wolfiporia cocos and Poria medula-panis grown in wheat bran liquid medium. Xylanase and iron-binding compounds were produced at high levels by the brown-rot fungus (BR) W. cocos and at low levels by the white-rot fungus (WR) P. medula-panis. Phenoloxidase was produced only by P. medula-panis. Polyacrylamide gel electrophoresis (PAGE) (SDS-PAGE) showed a wide variety of bands for extracellular proteins produced by W.cocos, with low molecular weight (<30 kDa) and minor bands with molecular weight above 45 kDa. Two bands with xylanase activity derived from W. cocos extracts were detected in the gels, whereas many different bands with xylanase activity were found in the extracts from P. medula panis. P. medula-panis is a selective lignin degrader, whereas W. cocos preferentially removes cellulose from wood. PMID- 11267646 TI - Immobilization of cycloisomaltooligosaccharide glucanotransferase for the production of cycloisomaltooligosaccharides from dextran. AB - Immobilization of cycloisomaltooligosaccharide glucanotransferase (CITase) and its application in the production of cycloisomaltooligosaccharides (CIs) from dextran were studied. Among various carrier materials examined, the enzyme adsorbed physically on Chitopearl BCW-3505 showed the highest activity (1.75 U/ml carrier). The activity remaining was 35%. The maximum CI yield in batch reactions at 0.2, 2 and 10% dextran was 28, 24 and 12%, respectively. The maximum CI yield at 2% dextran (24%) was slightly less than that with the free enzyme under the same conditions (26%). The concentration of linear oligosaccharides, the byproducts in the reaction mixture, was greater with the immobilized CITase than the free enzyme. The immobilized CITase was less thermostable than the free enzyme by about 10 degrees C. The pattern of influence of Ca(2+) concentration on the thermostability differed between the free and immobilized CITase. A Ca(2+) concentration of 50-100 mM was optimum for the thermostability of the immobilized CITase, 10-50 mM for the free enzyme. CIs were produced continuously by a column system packed with the immobilized enzyme at 40 degrees C with a space velocity (SV) of 6 h(-1). The three quarters life time was 4 weeks. We think that relatively long life time at fast SV was accomplished and CI production cost by this method should be lower than the batch reaction. This is the first report on immobilization of CITase. PMID- 11267648 TI - Enzymatic synthesis of amide surfactants from ethanolamine. AB - The condensation of a primary amine with fatty acids has been studied to determine optimum conditions for selective formation of amide surfactants via enzymatic amidification. Monoacylated ethanolamide and the diacylated amide-ester can be isolated from the reaction mixture, but the monoacylated ester cannot be isolated. The selectivity of the reaction depends on the solubility of the intermediate amide. Continuous precipitation of this product decreases the amount of amide-ester produced. Solubility values of the desired product (amide) are reported for different conditions.In acetonitrile, the ethyl ester of the corresponding fatty acid has been used successfully to avoid formation/precipitation of the ion-pair of the precursor reagents. In this medium, use of the transacylation reaction permits one to accelerate the reaction without producing a significant change in the selectivity toward the intermediate amide. This strategy is not successful in n-hexane where the solubilities of both ethanolamine and its ion-pair with lauric acid are similar.Results obtained for high loadings of substrates have been analyzed. In n-hexane and acetonitrile, the kinetics of the direct acylation reactions are controlled by the limited solubility of the ion pair formed by the two precursor reagents For the transacylation reaction in acetonitrile, at a sustrate loading of 2 mol l(-1,) selective production of as much as 92 mole percent N-acyl ethanolamine was observed in only 1.5 h. PMID- 11267649 TI - Purification and sequence analysis of the atypical maltohexaose-forming alpha amylase of the B. stearothermophilus US100. AB - The maltohexaose-forming alpha-amylase, of B. stearothermophilus US100, was purified to homogeneity by a combination of osmotic shock, starch adsorption and anion exchange chromatography. This enzyme has a relative molecular mass of 59 kDa. The analysis of the nucleotide sequence, of the corresponding gene, allowed the identification of a single open reading frame encoding a 549 amino acid protein, exhibiting a large homology to the other B. stearothermophilus alpha amylases. This homology reaches a maximum with those of DY-5 and DN1792 strains with respectively 3 and 4 aa different over 549. The relatively small differences, between Amy US100 and that of DN1792 strain, take in more importance since we have demonstrated that these enzymes differ essentially by their starch hydrolysis pattern. PMID- 11267650 TI - Chemical modification of lysine residues in Bacillus alpha-amylases: effect on activity and stability. AB - Chemical modification of lysine residues in two bacterial alpha-amylases, a mesophilic enzyme from Bacillus amyloliquefaciens (BAA) and a thermophilic enzyme from Bacillus licheniformis (BLA) was carried out using citraconic anhydride. 13 +/- 1 residues in BAA and 10 +/- 1 residues in BLA were found modified under defined experimental conditions. Modification brought about dramatic enhancement of thermal stability of BAA and catalytic activity of BLA. Such alterations were found dependent on the temperature and pH. Results obtained on Tm, the extent of deamidation, changes in the circular dichroism (CD) spectra and kinetic parameters before and after modification are discussed in terms of their contributions to the mechanism of irreversible thermoinactivation and activity enhancement. PMID- 11267651 TI - Infrared spectroscopy analysis of hemp (Cannabis sativa) after selective delignification by Bjerkandera sp. at different nitrogen levels. AB - Fourier-transform infrared (FT-IR) spectroscopy has been used to monitor changes in C/N-modified lignocellulosic substrates from Cannabis sativa L. in a 7-week solid-state fermentation with the white-rot fungus Bjerkandera sp. strain BOS55. The microbial transformation of hemp was considered as a pretreatment to pulping processes in paper industries. Special emphasis was paid on the N-content of the substrate, which was modified by: (i) external ammonium inputs, (ii) water extraction, and (iii) protease treatment.Selective delignification in the N limited media was observed. The most diagnostic FT-IR spectral bands in relation to changes in the lignocellulosic substrate were those corresponding to alkyl structures (2920, 1460 cm(-1)), carboxyl groups (1720 cm(-1)), amides (1650, 1540 cm(-1)) and carbohydrate (mainly 1030 cm(-1)). Simple and multiple regression functions revealed the potential of FT-IR in accurately reflecting substrate composition features previously determined by wet chemical methods. Correspondence analysis suggests C/N-dependent degradation patterns, and discriminant analysis confirmed that the differences between N-limited, N enriched and the original substrate were significant (P < 0.05) in terms of the intensities of five FT-IR diagnostic bands (1030, 1130, 1270, 1540 and 1650 cm( 1)).The results suggest that, in the system studied, the FT-IR spectroscopy is a reliable alternative to wet chemical analyses in the routine monitoring of the success of the biologic process since it reflects both qualitative and quantitative changes and it is very sensitive to lignin alteration and to carbohydrate and protein concentration. PMID- 11267652 TI - Estimation of oxygen mass transfer coefficient in stirred tank reactors using artificial neural networks. AB - The estimation of volumetric mass transfer coefficient, k(L)a, in stirred tank reactors using artificial neural networks has been studied. Several operational conditions (N and V(s)), properties of fluid (u(a)) and geometrical parameters (D and T) have been taken into account. Learning sets of input-output patterns were obtained by k(L)a experimental data in stirred tank reactors of different volumes. The inclusion of prior knowledge as an approach which improves the neural network prediction has been considered. The hybrid model combining a neural network together with an empirical equation provides a better representation of the estimated parameter values. The outputs predicted by the hybrid neural network are compared with experimental data and some correlations previously proposed in the literature for tanks of different sizes. PMID- 11267654 TI - Cloning and functional characterization of chicken p160 coactivator family members. AB - The factors SRC-1, TIF2 and ACTR were identified as interacting with nuclear receptors in a highly ligand-dependent manner. Because the molecular mass of each of these factors is approximately 160 kDa, they are collectively termed p160 coactivators. So far, p160 coactivators have been cloned from human, mouse and Xenopus. We report here the cloning of the chicken homologues of p160 coactivator members. As in human and mouse, chicken has three p160 coactivators. Each gene encodes an approximately 160 kDa protein which exhibits 70-80% amino acid sequence identity to human and mouse p160 coactivators. Chicken p160 coactivators also have the property of interacting with several liganded nuclear receptors. Moreover, we describe an imperfect LXXLL sequence, termed NR box 4, which is located downstream of NR box 3 and conserved between evolutionarily diverse species. The loss of NR box 4 results in a decrease of interaction with the nuclear receptor, which indicates that NR box 4 is required for efficient interaction. PMID- 11267653 TI - Rb and E2F-1 regulate telomerase activity in human cancer cells. AB - The ends of human chromosomes (telomeres) lose up to 200 bp of DNA per cell division. Chromosomal shortening ultimately leads to senescence and death in normal cells. Many human carcinoma lines are immortal in vitro, suggesting that these cells have a mechanism for maintaining the ends of their chromosomes. Telomerase is a ribonucleoprotein complex that synthesizes telomeric DNA onto chromosomes using its RNA component as a template. Recent studies have shown that inactivation of the retinoblastoma gene product pRb and the cyclin dependent kinase inhibitor p16(INK4A) is required for telomerase activity in epithelial cells. We have demonstrated previously that restoration of functional retinoblastoma (Rb) expression is sufficient to downregulate telomerase activity in carcinoma cells. To determine mechanisms by which Rb regulates telomerase expression, we examined the effects of cyclin dependent kinase (cdk) mediated Rb inactivation and the release of E2F-1 on telomerase activity in human carcinoma cells. Overexpression of cdk2 and cdk4 but not a dominant negative cdk2 rescued Rb mediated downregulation of telomerase activity. Overexpression of the cdk regulatory subunit cyclin D1 also rescued telomerase downregulation and p16 expression alone was sufficient to ablate activity. E2F-1 overexpression was sufficient to rescue Rb mediated reduction of telomerase activity, but an E2F-1 mutant defective in DNA and Rb binding activities failed to produce this effect. Tumor tissue from E2F-1 -/- mice was negative for telomerase activity, indicating a key regulatory role for this transcription factor. PMID- 11267655 TI - Cloning of the rat Slc14a2 gene and genomic organization of the UT-A urea transporter. AB - We cloned the Slc14a2 gene and determined the genomic organization of the rat urea transporter UT-A. Slc14a2, the gene encoding the rat UT-A transporter, extends for more that 300 kb. The four known rat mRNA isoforms: UT-A1, UT-A2, UT A3, and UT-A4 are transcribed from 24 exons. The Slc14a2 genomic map also accounts for 3'-untranslated sequences expressed alternatively in UT-A1, UT-A2, and UT-A3. We previously identified a TATA-less, tonicity-responsive promoter controlling the transcription of UT-A1, UT-A3, and UT-A4 from a single initiation site in the 5'-flanking region of the gene. Here, we describe a second, internal promoter in intron 12, which controls the transcription of UT-A2 starting from exon 13. This region contains a TATA motif upstream from the UT-A2 transcription start site, and shows consensus sequences for the cAMP response element (CRE) and for the tonicity enhancer (TonE) motif. Stimulation by cAMP induces UT-A2 mRNA expression in mIMCD3 cells, and luciferase activity in mIMCD3 cells transfected with those pGL3 constructs including the CRE sequences. Although long-term exposure to hypertonicity induces UT-A2 expression in mIMCD3 cells, hypertonicity does not induce significantly the activity of the promoter in intron 12. In summary, we describe the genomic structure of the rat UT-A urea transporter, encoded by the Slc14a2 gene. Our findings suggest that two promoters regulate transcription of the four UT-A isoforms, and that stimulation of transcription by vasopressin, mediated by cAMP and CRE sequences, and controlled by an intronic promoter, may contribute to the increase in UT-A2 expression during water deprivation. PMID- 11267656 TI - Diazepam and rolipram differentially inhibit cyclic AMP-specific phosphodiesterases PDE4A1 and PDE4B3 in the mouse. AB - Cyclic AMP is hydrolyzed by members of at least eight classes of cyclic nucleotide phosphodiesterases (PDEs). Although it has been reported that cyclic AMP PDE activity in mammalian tissues can be inhibited by benzodiazepines, it has not been conclusively demonstrated that members of the class of cyclic AMP specific, rolipram-inhibitable PDEs (PDE4s) are targets for these drugs. Moreover, no PDE4s expressed in mice have been characterized. To address these issues, we isolated two cDNAs representing homologues of PDE4A1 and PDE4B3 from a mouse brain library. After transient transfection in human embryonic kidney (HEK) 293 cells, the mouse PDEs hydrolyzed cyclic AMP with a low K(m) and were inhibited by rolipram; both are properties typical of other mammalian PDE4 enzymes. In addition, we found that diazepam inhibited cyclic AMP hydrolysis by the mouse PDE4 subtypes. Interestingly, PDE4B was significantly more sensitive to inhibition by both rolipram and diazepam than the PDE4A subtype. This is the first demonstration that recombinantly expressed PDE4s are inhibited by diazepam, and should facilitate future studies with mouse models of depression and anxiety. PMID- 11267657 TI - Structure and complex transcription pattern of the mouse SK1 K(Ca) channel gene, KCNN1. AB - Small conductance calcium-gated K(+) channels (SK channels) are encoded by the three SK genes, SK1, SK2, and SK3. These channels likely contribute to slow synaptic afterhyperpolarizations of apamin-sensitive and apamin-insensitive types. SK channels are also widely expressed outside the nervous system. The mouse SK1 gene comprises at least 12 exons extending across 19.8 kb of genomic DNA. This gene encodes a complex pattern of alternatively spliced SK1 transcripts widely expressed among mouse tissues. These transcripts exhibit at least four distinct 5'-nucleotide sequence variants encoding at least two N-terminal amino acid sequences. Optional inclusion of exons 7 and 9, together with two alternate splice donor sites in exon 8, yields transcripts encoding eight variant C terminal amino acid sequences for SK1. These include an altered putative S6 transmembrane span, modification of the C-terminal cytoplasmic domain binding site for calmodulin, and generation of two alternate predicted binding sites for PDZ domain-containing proteins. 20 of the 32 predicted mouse SK1 transcripts are expressed in brain at levels sufficient to allow consistent detection, and encode 16 SK1 polypeptide variants. Only four of these 16 polypeptides preserve the ability to bind calmodulin in a Ca(2+)-independent manner. Mouse SK1 also exhibits novel, strain-specific, length polymorphism of a polyglutamate repeat in the N-terminal cytoplasmic domain. The evolutionary conservation of this complex transcription pattern suggests a possible role in the tuning of SK1 channel function. PMID- 11267658 TI - Effects of mercuric chloride on the regulation of expression of the acute phase response components alpha(1)-acid glycoprotein and C/EBP transcription factors. AB - We have previously shown that in response to treatment with HgCl(2), the adult mouse liver exhibits both transcriptional and translational regulation of the acute phase response genes. In this study we asked whether the heavy metal treatment affects the regulation of the C/EBP transcription factors which play a key role in regulation of the acute phase response gene. Our studies have shown that the AGP gene is transcriptionally activated while transcription of the CCAAT/enhancer-binding trans-activating protein (C/EBP)alpha gene is slightly down-regulated and that of the C/EBPbeta gene does not respond. Both the C/EBPalpha and C/EBPbeta mRNAs produce multiple isoforms possibly by alternative translation initiation (ATI) of multiple internal AUG initiation sites. The C/EBPbeta mRNA appears to be stabilized. Although similar regulatory processes occur in response HgCl(2) vs. LPS, our data suggest that the translational processes (ATI) are differentially affected. In addition, a major difference lies in the fact that the C/EBPbeta gene is not transcriptionally activated by HgCl(2). Our data show decreased binding activity and pool levels of the C/EBPalpha isoform (p42(C/EBPalpha)) and increased binding activity and pool levels of C/EBPbeta isoform (p35(C/EBPbeta)) in response to HgCl(2). We propose that this isoform may be involved in the regulation of AGP gene expression in response to heavy metals and that there is a significant difference between the HgCl(2)-mediated and LPS-mediated inflammatory response. PMID- 11267659 TI - Nucleotide sequence of the mouse VEGF 3'UTR and quantitative analysis of sites of polyadenylation. AB - Sequencing of rat and human vascular endothelial growth factor (VEGF) cDNA clones has previously identified a 3' untranslated region of approximately 1.9 kb, although the apparent site of polyadenylation differed in the two species, despite a high degree of sequence conservation in the region. Neither site is preceded by a canonical AAUAAA polyadenylation signal, a situation frequently found in genes that are subject to alternative polyadenylation. We have sequenced 2.25 kb of the 3' region of the mouse VEGF gene and mapped the usage of potential polyadenylation sites in fibroblasts cultured under both normoxic and hypoxic conditions. We find that two sites for polyadenylation are present in the mouse VEGF gene but the majority of transcripts contain the longer form of the 3'UTR and that their usage is not effected by environmental oxygen tension. PMID- 11267660 TI - Molecular cloning and characterization of Kremen, a novel kringle-containing transmembrane protein. AB - Kringle domain, a triple-disulfide-linked domain, is conserved in diverse proteins which play important roles in various biological processes. We cloned Kremen, a novel member of kringle-containing proteins, using a newly developed unique strategy, 'Kringle-SAGE (serial analysis of gene expression)', which enables comprehensive analysis of kringle-containing proteins. Kremen is likely to be a type-I transmembrane protein composed of 473 amino acid residues. Kremen has a kringle domain, a WSC domain, and CUB domains in the extracellular region, while the intracellular region has no conserved motif involved in signal transduction. In the mouse embryo, the Kremen mRNA level, which was increased during embryonic development, was localized in the apical ectodermal ridge of limb buds, myotome, and sensory organs (e.g. optic vesicle, otic vesicle, nasal pit). In the adult mouse, Kremen mRNA was expressed in a variety of tissues with a relatively strong expression in the lung, heart, and skeletal muscle. Kremen mRNA expression in C2C12 and NIE-115 cells increased during respective differentiation into muscular and neural cells. These results suggest a potential role for Kremen in the regulation of cellular responses upon extracellular stimulus or cell-cell interaction in neuronal and/or muscle cells. Kringle-SAGE is expected to facilitate further elucidation of structure and functions of kringle proteins. PMID- 11267662 TI - Cloning of a cDNA for a constitutive NRT1 transporter from soybean and comparison of gene expression of soybean NRT1 transporters. AB - We have isolated a cDNA for a putative transporter, named GmNRT1-3, in the NRT1 family from soybean. It was predicted to have a similar topological structure not only to both GmNRT1-1 and GmNRT1-2 reported previously, but also to other members of the family. Two other cDNAs isolated have parts of the sequence for putative NRT1 transporters, GmNRT1-4 and GmNRT1-5, suggesting that at least five NRT1 transporters occur in soybean. These GmNRT1 genes and the GmNRT2 gene, encoding a soybean NRT2 nitrate transporter, showed different expression patterns to each other under various nitrogen conditions. Specifically, GmNRT1-3 was constitutively expressed in both roots and leaves, while GmNRT1-2 was gradually expressed as the roots developed in the presence of ammonium as a nitrogen source, but not in the presence of both ammonium and nitrate. Based on these results, we discussed the possible regulation in the expression and role of these transporters in nitrate uptake. PMID- 11267661 TI - Functional analysis and androgen-regulated expression of mouse organic anion transporting polypeptide 1 (Oatp1) in the kidney. AB - Mouse Oatp1 was recently identified as a new murine member of the organic anion transporting polypeptide (Oatp) family and suggested to represent the counterpart of rat Oatp1. Northern blot analysis detected expression of several mouse Oatp transcripts predominantly in liver and kidney. In the present study we describe the strict androgen-dependent expression of mouse Oatp1 mRNA in kidney and obtained further information about its substrate specificity using Xenopus oocytes. In addition to the previously reported estrone-3-sulfate, we demonstrate that mouse Oatp1 mediates sodium-independent uptake of the anionic steroid conjugates dehydroepiandrosterone sulfate (K(m) approximately 8 microM) and estradiol-17-glucuronide (K(m) approximately 5 microM) and also of the prostaglandin PGE(2). PMID- 11267664 TI - Alternative splicing and tissue-specific transcription of human and rodent ubiquitous 5-aminolevulinate synthase (ALAS1) genes. AB - The rate of haem synthesis in non-erythroid mammalian tissues is controlled by the ubiquitous isoform of 5-aminolevulinate synthase (ALAS1). In order to explore the regulation of mammalian ALAS1 genes, we have investigated the transcription of the human and rat genes. The 17 kb human gene differs from the rat gene in containing an additional untranslated exon that is alternatively spliced to produce a longer, minor mRNA transcript. Relative amounts of the two transcripts were similar in all tissues examined. Analysis of mRNA transcripts in human and rat tissues revealed tissue-specific differences in the use of transcription start sites by closely similar core promoters. In brain, initiation was from sites within and upstream from the TATA box, including an initiator-like element. In liver, initiation was TATA-driven from a single downstream site that appeared to be used exclusively for induction by drugs. Intermediate patterns were observed in other tissues and cell lines. Mutation of the TATA box did not impair transcription in transfected HeLa cells but activated upstream start sites, recapitulating the brain pattern. Our findings indicate that the conformation of the core ALAS1 promoter that directs assembly of the transcription pre-initiation complex may vary between tissues and have implications for understanding the tissue-specific regulated expression of this gene. PMID- 11267663 TI - Tn10 insertional mutations of Bacillus subtilis that block the biosynthesis of bacilysin. AB - Transposon mutagenesis was employed to isolate the gene(s) related with the biosynthesis of dipeptide antibiotic in Bacillus subtilis PY79 (a prototrophic derivative of the standard 168 strain). The blocked mutants were phenotypically selected from the transposon library by bioassay and the complete loss of biosynthetic ability was verified through ESI-mass spectrometry analysis. Four different bacilysin nonproducer mutants (Bac(-)::Tn10(ori-spc)) were isolated from the transposon library. The genes involved in bacilysin biosynthesis were identified as thyA (thymidilate synthetase), ybgG (unknown; similar to homocysteine methyl transferase) and oppA (oligopeptide permease), respectively. The other blocked gene was yvgW (unknown; similar to heavy metal-transporting ATPase); however, backcross studies did not verify its involvement in bacilysin biosynthesis. This gene, on the other hand, appeared to be necessary for efficient sporulation and transformation. Opp involvement was significant as it suggested that bacilysin biosynthesis is under or a component of the quorum sensing pathway which has been shown to be responsible for the establishment of sporulation, competence development and onset of surfactin biosynthesis. For verification, it was necessary to check the involvement of peptide pheromones (PhrA or PhrC) internalized by the Opp system and response regulator ComA as the essential components of this global control. phrA, phrC and comA deleted mutants of PY79 were thus constructed and the latter two genes were shown to be essential for bacilysin biosynthesis. PMID- 11267665 TI - Developmental expression of meprin metalloprotease subunits in ICR and C3H/He mouse kidney and intestine in the embryo, postnatally and after weaning. AB - Meprins are secreted and membrane-bound metalloendopeptidases highly expressed in kidney and intestinal epithelial cells. They are oligomeric glycoproteins composed of evolutionarily related alpha and/or beta subunits. The present work revealed that the messages for both meprin subunits were expressed in intestine and kidney in ICR and C3H/He mouse embryos (as early as day 11), indicating developmental functions for both subunits. During the first 2 weeks after birth, the mRNA levels for both subunits increased in ICR mice, but between 10 days and 3 weeks (time of weaning) the alpha subunit level in the intestine fell markedly. In adult ICR mice, meprin beta mRNA was consistently expressed in both kidney and intestine, whereas meprin alpha mRNA was highly expressed in kidney but only present at low levels in intestine. In C3H/He mice, the pattern of meprin alpha and beta subunit mRNA expression was similar to that of ICR mice, except that meprin alpha was barely detectable in kidney after birth. The results of postnatal studies indicate that the meprin alpha subunit has a role in the intestine during suckling but is not essential after weaning, and that the beta homooligomer is the major meprin form after weaning in both kidney and intestine. PMID- 11267666 TI - Expression and localization of transcripts of MT5-MMP and its related MMP in the ovary of the medaka fish Oryzias latipes. AB - cDNA clones of MT5-matrix metalloproteinase (MT5-MMP) and a related protein (designated MT5-MMP-del) were isolated by screening the cDNA library and by 3' rapid amplification of cDNA ends using an ovary RNA of the medaka fish Oryzias latipes. The MT5-MMP clone encodes a protein of 546 amino acids while the MT5-MMP del clone encodes a protein of 431 amino acids. Compared with mammalian counterparts, the fish MT5-MMP and MT5-MMP-del both lack the signal peptide and a part of the prodomain. The fish MT5-MMP and MT5-MMP-del were different in that the latter did not have the stem/transmembrane/cytoplasmic domain. The two fish MMPs were expressed in the ovary, testis, brain, and intestine. In the ovary, MT5 MMP mRNA was expressed in the oocytes of small growing follicles. In contrast, MT5-MMP-del mRNA was found in the stromal interstitial cells. These results strongly suggest that a MT5-MMP/gelatinase A cascade may possibly operate in the process of spawning and/or other events associated with ovulated oocytes or fertilized eggs of the medaka fish. PMID- 11267667 TI - Molecular cloning of human squamous cell carcinoma antigen 1 gene and characterization of its promoter. AB - The squamous cell carcinoma antigen (SCCA) serves as a serological marker for squamous cell carcinomas. Molecular cloning of the SCCA genomic region has revealed the presence of two tandemly arrayed genes, SCCA1 and SCCA2, which are 95% identical in nucleotide sequence. SCCA1 is a papain-like cysteine proteinase inhibitor, while SCCA2 is a chymotrypsin-like serine proteinase inhibitor. We analyzed here the sequence and the promoter activity of the 5'-flanking region of the SCCA1 gene. Deletion analysis of SCCA1 and SCCA2 promoter identified a 471-bp core promoter region upstream of the transcription start site. The transcriptional activity of SCCA1 promoter was up-regulated in squamous cell carcinoma cells, compared with keratinocyte and adenocarcinoma cells. The ratios of SCCA1 to SCCA2 promoter activity in squamous cell carcinoma, keratinocyte and adenocarcinoma cells were respectively 1.6, 5.3 and 2.8. Position -50 of SCCA1 and SCCA2 promoters played an important role in determining the promoter activities of SCCA1 and SCCA2. These findings suggest that the transcriptional regulation of SCCA1 and SCCA2 might differ among squamous cell carcinoma, keratinocyte and adenocarcinoma cells, and that SCCA1 promoter might be a potential target of gene therapy for squamous cell carcinoma. PMID- 11267668 TI - Isolation and characterization of the promoter region of the rat kidney-type glutaminase gene. AB - A lambdaEMBL3 rat genomic library was screened to clone a phage that contained the promoter region of the kidney-type mitochondrial glutaminase gene. The resulting lambdaGA1 phage contained 13.7 kb of genomic DNA that was mapped by Southern blotting and restriction analysis. The 2.22 kb and 0.83 kb SacI fragments of lambdaGA1 were sequenced and the transcription initiation site was identified by RNase mapping. The reported sequence contains 2287 bp of the promoter, the entire exon 1 (542 bp), and 223 bp of the initial intron of the glutaminase gene. The initial exon contains 141 bp of 5'-nontranslated sequence and 401 bp of coding sequence that encodes the 72-amino acid mitochondrial targeting presequence and 61 amino acids from the N-terminus of the mature 66 kDa glutaminase subunit. Various segments of the GA promoter were cloned into a chloramphenicol acetyltransferase (CAT) expression vector. The resulting GA-CAT constructs were transfected into LLC-PK(1)-F(+) kidney cells to assess the promoter function of the isolated genomic DNA. The GA(-402)CAT construct produced a 10-fold greater CAT activity than the promoter-less pCAT vector. Analysis of various deletion constructs indicated that elements located between -402 and -63 bp must act in synergy with more proximal elements to create a functional promoter. The initial 402 bp segment lacks a TATA sequence but is GC-rich and contains two CCAAT boxes and two Sp1 sites. PMID- 11267669 TI - Epigenetic mark sequence of the H19 gene in human sperm. AB - We have investigated the epigenetic mark in the human H19 gene. The H19 promoter is methylation-free in human sperm, but it is methylated in the paternally derived allele of most adult tissues. Consequently, the H19 gene is exclusively transcribed from the maternal allele. It was demonstrated that the differentially methylated region (DMR) located 2 kb upstream from mouse H19 is essential for the imprinting of H19. A 39 bp sequence in DMR has a high degree of similarity between humans, mice and rats. The highly conserved 15 bp core region of the consensus sequence contains four methylatable sites, and thus has been proposed as a potential imprinting mark region. In this study, fine epigenetic sequencing analysis was performed on the sperm DNA in comparison with other adult organs. Interestingly, the conserved sequence of the potential mark region was methylated in almost all the sperm genomes analyzed. Furthermore, the single dinucleotide CpG, whose methylation affects the accessibility of the element to CTCF, was methylated in the conserved core in the human sperm. These results suggest that the human core sequences may act as an imprinting center in the reciprocal monoallelic expression of H19. PMID- 11267670 TI - Structure and characterization of the human insulin-like growth factor binding protein (IGFBP)-6 promoter: identification of a functional retinoid response element. AB - The 1.7 kb human insulin-like growth factor binding protein (IGFBP)-6 gene 5' flanking region was sequenced and found to have promoter activity in human osteoblasts. The sequence contains four clustered transcription start sites and three retinoic acid response elements (RAREs) with widely spaced half-sites. Only the proximal DR15 RARE was functional. Retinoids increased IGFBP-6 promoter activity up to 3-fold. PMID- 11267671 TI - A novel fibroblast growth factor receptor-5 preferentially expressed in the pancreas(1). AB - Using the polymerase chain reaction on human embryonic cDNAs, we isolated a cDNA encoding a novel 504 amino acid protein, termed fibroblast growth factor receptor (FGFR)-5, which is highly homologous to known FGFRs. The NH(2)-terminal portion of FGFR5 contains a putative secretory signal sequence, three typical immunoglobulin-like domains, six cysteines, and an acidic box, but no HAV motif. The COOH-terminal portion of FGFR5 contains one transmembrane domain but no intracellular kinase domain. Recombinant FGFR5 expressed in COS-7 cells is not secreted, but recombinant truncated FGFR5 lacking the predicted transmembrane domain is secreted. Acidic fibroblast growth factor (aFGF) and basic fibroblast growth factor (bFGF) do not bind to FGFR5. Among 23 adult human tissues, FGFR5 mRNA is preferentially expressed in the pancreas. These results suggest that FGFR5 may provide a binding site for some other fibroblast growth factors and may regulate some pancreatic function. PMID- 11267673 TI - Molecular cloning, mapping and characterization of the human neurocalcin delta gene (NCALD). AB - We identified a new human gene that encodes a cognate of the bovine neurocalcin delta from a human fetal brain cDNA library; hence we named it human neurocalcin delta (NCALD) gene. The deduced polypeptide product of the cDNA is 22 kDa in size, and its amino acid sequence is 100% and 99% identical to that of the bovine and chicken neurocalcin, respectively. Northern blots showed that the NCALD gene is more abundantly expressed in brain, testis, ovary and small intestine. Tissue in situ hybridization confirmed the existence of the NCALD mRNA in the adult human testis. Radiation hybrid panel mapping localized the gene to chromosome 8 between molecular markers D8S270 and D8S257. PMID- 11267672 TI - Identification of hTLR10: a novel human Toll-like receptor preferentially expressed in immune cells. AB - Herein we describe the isolation of a cDNA encoding a novel human Toll-like receptor (hTLR) that we term hTLR10. Human TLR10 contains 811 amino acid residues. Deduced amino acid sequence analysis reveals that like the other known hTLRs (hTLR1-9) it is characterized by a signal peptide followed by multiple leucine-rich repeats (LRRs), a cysteine-rich domain, a transmembrane sequence and a cytoplasmic domain homologous to that of the human interleukin-1 receptor. Phylogenetic analysis indicates that among all the hTLRs, hTLR10 is most closely related to hTLR1 and hTLR6; the overall amino acid identity is 50% and 49%, respectively. hTLR10 mRNA is most highly expressed in lymphoid tissues such as spleen, lymph node, thymus, and tonsil. Expression analysis of cell lines indicates a predominance in a variety of immune cell types. Thus, hTLR10 is preferentially expressed in tissues and cells involved in immune responses. PMID- 11267674 TI - Isolation and characterization of cDNAs for the protein kinase HIPK2. AB - HIPK2 (homeodomain-interacting protein kinase 2) is a CD95 binding partner in yeast. Its primary amino acid sequence is highly conserved between human and mouse. The highest HIPK2 mRNA expression is found in neuronal tissue. The HIPK2 gene is located on human chromosome 7q33-35 and the protein is mainly localized in the nucleus. HIPK2 has been described to play a role as a co-repressor for homeodomain transcription factors. PMID- 11267675 TI - Molecular cloning of rat transmembrane domain protein of 40 kDa regulated in adipocytes and its expression in H9c2 cells exposed to ischemic hypoxia and reoxygenation. AB - We clone a 1230 bp complementary DNA encoding rat transmembrane domain protein of 40 kDa regulated in adipocytes (TPRA40), an orphan receptor, by reverse transcription-polymerase chain reaction using H9c2 cells derived from embryonic rat heart. The deduced amino acid sequence of rat TPRA40 consists of 369 amino acids and has a longer carboxyl terminus than that of the mouse protein. The level of TPRA40 mRNA decreases significantly throughout ischemic hypoxia and reoxygenation. PMID- 11267676 TI - Cloning of cDNA encoding a soybean allergen, Gly m Bd 28K. AB - A cDNA clone encoding a soybean allergen, Gly m Bd 28K, has been isolated. The clone has a 1567-bp cDNA insert with a 1419-bp open reading frame and a 148-bp 3' untranslated region, followed by a polyadenylation tail. The open reading frame was shown to encode a polypeptide composed of 473 amino acids. The chemically determined amino acid sequences of the peptides obtained from the allergen, including its N-terminal peptide, were shown to be contained in the N-terminal region of the amino acid sequence deduced from the cDNA, showing that the first half of the cDNA encodes the allergen with a preceding segment of 21 amino acids. The peptide fragment including the allergen was expressed as a fusion protein with glutathione S-transferase in Escherichia coli and immunoblotted with the sera of soybean-sensitive patients and the monoclonal antibody against the allergen. Furthermore, homology analyses demonstrate that the polypeptide for the cDNA exhibits high homology with the MP27/MP32 proteins in pumpkin seeds and the carrot globulin-like protein. This finding suggests that the polypeptide may consist of a 21-amino acid segment as a part of the signal peptide and the proprotein, which may be converted to two mature proteins, Gly m Bd 28K and a 23 kDa protein, during the development of soybean cotyledons. PMID- 11267677 TI - Molecular cloning of a cDNA for rat TM4SF4, a homolog of human il-TMP (TM4SF4), and enhanced expression of the corresponding gene in regenerating rat liver(1). AB - il-TMP (also known as TM4SF4) is a human tetraspanin that is expressed in human intestine and liver. We have cloned a novel cDNA for a rat gene with sequence similar to that of a cDNA for human il-TMP. The cDNA encoded a protein of 202 amino acids, designated rat TM4SF4. The corresponding transcript was detected in rat liver and testis. The expression of rat TM4SF4 was enhanced in regenerating liver after two-thirds partial hepatectomy. It was supposed that rat TM4SF4 might play a role in cell proliferation and in liver regeneration. PMID- 11267678 TI - Isolation, characterization, and mapping of a novel human KRAB zinc finger protein encoding gene ZNF463. AB - A novel human KRAB (Kruppel associated box) type zinc finger protein encoding gene, ZNF463, was obtained by mRNA differential display and RACE. It consists of 1904 nucleotides and encodes a protein of 463 amino acids with an amino-terminal KRAB domain and 12 carboxy-terminal C2H2 zinc finger units. The gene is mapped to chromosome 19q13.3 approximately 4 by FISH. As from Northern blot analysis ZNF463 is only expressed in testis, RT-PCR indicates that ZNF463 is expressed more highly in normal fertile adults than in fetus and azoospermic patients suggesting that it may play a role in human spermatogenesis. PMID- 11267680 TI - Characterization of human FSD1, a novel brain specific gene on chromosome 19 with paralogy to 9q31. AB - We have characterized a novel human gene, FSD1, on chromosome 19. FSD1 has a BBC, FN3 and SPRY domain, it is distantly related to the midline 1 gene and is expressed only in the brain. We have established its exon-intron structure and we have identified the corresponding orthologous genes in other species. In addition, the identification of FSD1 has led us to identify a homologous counterpart sequence on chromosome 9. PMID- 11267679 TI - Characterization and regulation of Schizosaccharomyces pombe gene encoding thioredoxin. AB - A cDNA coding thioredoxin (TRX) was isolated from a cDNA library of Schizosaccharomyces pombe by colony hybridization. The 438 bp EcoRI fragment, which was detected by Southern hybridization, reveals an open reading frame which encodes a protein of 103 amino acids. The genomic DNA encoding TRX was also isolated from S. pombe chromosomal DNA using PCR. The cloned sequence contains 1795 bp and encodes a protein of 103 amino acids. However, the C-terminal region obtained from the cDNA clone is -Val-Arg-Leu-Asn-Arg-Ser-Leu, whereas the C terminal region deduced from the genomic DNA appears to contain -Ala-Ser-Ile-Lys Ala-Asn-Leu. This indicates that S. pombe cells contain two kinds of TRX genes which have dissimilar amino acid sequences only at the C-terminal regions. The heterologous TRX 1C produced from the cDNA clone could be used as a subunit of T7 DNA polymerase, while the TRX 1G from the genomic DNA could not. The upstream sequence and the region encoding the N-terminal 18 amino acids of the genomic DNA were fused into the promoterless beta-galactosidase gene of the shuttle vector YEp357 to generate the fusion plasmid pYKT24. Synthesis of beta-galactosidase from the fusion plasmid was found to be enhanced by hydrogen peroxide, menadione and aluminum chloride. It indicates that the expression of the cloned TRX gene is induced by oxidative stress. PMID- 11267681 TI - Cloning and expression of novel mosaic serine proteases with and without a transmembrane domain from human lung. AB - Two cDNAs encoding novel mosaic proteins with a serine protease domain and potential regulatory modules, consisting of a protein kinase substrate and a low density lipoprotein receptor, were cloned from a human lung cDNA library by PCR. One with a transmembrane domain (MSPL) and the other without one (MSPS) comprise 581 and 537 amino acids, respectively. Except for the C-terminal ends, the two isoforms had an identical serine protease domain exhibiting 42, 39 and 43% identity with those of plasma kallikrein, hepsin and transmembrane protease serine 2, respectively. Both genes were predominantly expressed in human lung, placenta, pancreas and prostate. PMID- 11267682 TI - Identification and characterisation of the Drosophila homologue of the yeast Uba2 gene. AB - We have identified the Drosophila uba2 protein (dUba2). Analysis of the amino acid composition reveals similarity with both the mammalian (47% identity) and yeast (31% identity) homologues. dUba2 is present throughout the Drosophila life cycle but is most abundant during stages of proliferation. The protein is nucleoplasmic throughout much of the cell cycle, however it is lost from the nucleus during mitosis. The DUba2 localisation in the nucleoplasm is not uniform but is observed as concentrated patches reminiscent of the staining patterns seen for other proteins from this group. The nature of these sites is not clear, however the failure of dUba2 to localise to the sites of chorion amplification in ovaries suggests that they are not sites of ongoing DNA replication. PMID- 11267683 TI - Analysis of PBX1 as a candidate gene for type 2 diabetes mellitus in Pima Indians. AB - The human proto-oncogene PBX1 codes for a homeodomain containing protein that modulates expression of several genes, including those contributing to regulation of insulin action and glucose metabolism. PBX1 is located on chromosome 1q22, a region linked with type 2 diabetes in Pima Indians, Caucasians, and an Old Order Amish population. We have investigated the PBX1 genomic sequence to identify polymorphisms that may contribute to diabetes susceptibility in the Pimas. PBX1 is composed of nine exons spanning approx. 117 kb and is located within 300 kb of microsatellite D1S1677, which marks the peak of linkage to diabetes susceptibility in the Pima Indians. We detected 16 single nucleotide polymorphisms in PBX1 including one causing a glycine to serine substitution at residue 21. Comparison of the frequencies of the polymorphisms between affected and unaffected Pima Indians did not detect any significant differences, indicating that mutations in PBX1 do not explain the linkage of 1q with type 2 diabetes in this population. The genomic structure of PBX1 provides a basis for similar systematic examinations of this candidate locus in other populations in relation to both type 2 diabetes and other metabolic disorders. PMID- 11267684 TI - Stochastic modelling as a tool for planning animal-health surveys and interpreting screening-test results. AB - Traditionally, the planning of surveys (in particular, sample-size calculations) has relied on assumptions including the assumption of perfect screening tests. This paper presents a novel approach that can be used for planning animal-health surveys and interpreting screening-test results in the context of these surveys. A stochastic simulation model developed to assess the properties of herd-level sampling schemes and surveys has been adapted for large surveys aimed at substantiating freedom from infection at a national or regional level. We use a Bayesian approach to derive the post-survey probability of freedom from infection from the pre-survey probability of freedom and the likelihood ratio that is associated with screening-test results. We applied the model to two consecutive surveys conducted in 1998 and 1999 in Switzerland to substantiate freedom from infectious bovine rhinotracheitis (IBR) in the cattle population of about 56000 herds (median herd size of 15 cattle > 2 yr of age in 1999). In 1998, serum samples were taken from five cattle > 2 yr in 4672 herds, and in 1999 from all cattle > 2 yr old in 648 herds; samples were analysed by ELISA. The survey of 1999 provided less evidence than that of 1998 to support a status of freedom from infection; also, the characteristics of both herd-level sampling schemes were similar. We argue that the rationale for survey planning depends on the pre survey probability of freedom from infection (i.e. our level of confidence that the infection does not occur in the targeted animal population). In consequence, surveys should be tailored to individual populations in the respective countries or regions. The model has been developed in an Excel spreadsheet to allow flexibility of use, and adaptation to many other animal-health issues. PMID- 11267685 TI - No overall relationship between average daily weight gain and the serological response to Mycoplasma hyopneumoniae and Actinobacillus pleuropneumoniae in eight chronically infected Danish swine herds. AB - The association between the average daily weight gain (from approximately 4 to 20 weeks of age) and the serological responses to respiratory infections was examined in a longitudinal study including 825 pigs from eight chronically infected herds. Pigs were bled every 4th week (starting from approximately 4 weeks of age), and sera were analyzed for antibodies to Mycoplasma hyopneumoniae and Actinobacillus pleuropneumoniae serotypes 2, 5-7 and 12.Mixed analysis of covariance analyzed the relationship between the average daily weight gain and a categorical variable defining seroconversion as none, early or late as compared to the median time (estimated across herds) of seroconversion for the particular pathogen. The variables "gender", "weight at an approximate age of 4 weeks" and "time" (defining the exact length of the follow-up period), were included as explanatory variables, and "litter" and "herd" were included as explanatory random variables. The individual pig was the unit of concern. The variable defining time at seroconversion was not significantly associated with the average daily weight gain, when evaluating models across all eight herds. The apparent lack of effect could be because most pigs included in the study were subclinically infected, or because a temporary negative influence of the infections is hidden due to an increased growth in the period following infection. In conclusion, at least in these eight herds, seroresponses to M. hyopneumoniae and A. pleuropneumoniae could not be used to predict the effect of the pathogens on the daily weight gain. PMID- 11267686 TI - Comparison of economically optimized culling recommendations and actual culling decisions of Finnish Ayrshire cows. AB - Our purpose was to compare culling recommendations obtained from an economic optimization model with actual culling of Finnish Ayrshire cows. The dynamic programming (DP) model we used optimizes replacement and breeding decisions to maximize the net revenues from cows currently in a herd and their potential replacements over a 5-year decision horizon. Cows were described in the model by five state variables: parity, stage of lactation, month of calving, milk production level, and days open (pregnancy status). We performed survival analysis to study the effects of those five factors on culling and to compare the actual culling of cows in December 1993 and June 1994 with the optimized replacement recommendations for the same months and for cows in the same herds. The risk of culling increased as a cow grew older, both in the actual herds and in the DP recommendations for December. In the optimized replacements for June, however, the age of a cow did not play a significant role. A cow that had been in milk > 270 days had a lower risk of culling than cows in earlier stages of lactation. When 305-day milk production increased by 100 kg, the risk of culling decreased by 4% in the actual herds and by 6 and 12% in the DP recommendations for June and December, respectively. When the days open lengthened by a month, the risk of culling was 2.0- and 1.6-times higher in the actual herds and 1.7- and 2.0-times higher in the DP recommendations for June and December, respectively. Month of calving had a different effect in the optimized recommendations compared with the real-life situation: cows calving from January to August had a lower risk of culling than cows calving in the fall in the actual herds, but the optimization model recommended heavier culling for cows calving between January and August. The DP did not account for diseases and did not allow replacements during the first 2 months of lactation and some of the observed differences could be due to this. However, the results suggested that Finnish farmers might not be taking full advantage of the seasonality in milk pricing and production to maximize the profits of their herds--even though their culling decisions are rational and in quite close agreement with the optimized recommendations. PMID- 11267687 TI - The effects of health classification and housing and management of feeder pigs on performance and meat inspection findings of all-in-all-out swine-finishing herds. AB - The effects of health classification and of housing and management on performance and meat inspection findings were studied in 166 all-in-all-out finishing herds in Finland in 1995 and 1998. Producers could buy either health-classified (certified free of certain diseases) or standard-class pigs for their farms. Herds that had attained a certain level of housing and management were eligible to join the LSO 2000 management system for finishing herds. In 1995, none of the study herds were LSO 2000 units. In 1998, 76 of them had become LSO 2000 units, while 90 had remained as non-LSO 2000 units. Continuous outcome variables (daily gain and time in the finishing unit) were analysed with a mixed-model procedure with repeated measurements from the same farms. Discrete variables were analysed either with Poisson regression (mortality, whole- and partial-carcass condemnations, organ condemnations, arthritis, abscesses) or logistic regression (liver condemnations, pneumonia, pleuritis). The models were adjusted for the year, size of the herd and the average slaughter weight of the batch. The benefits obtained by the health classification of the feeder pigs included a substantial increase in daily gain (37g, about 5%) and decrease in time in the finishing unit (4 days, about 4%), decrease in mortality (odds ratio, OR 0.68), a substantial decrease in pneumonia (OR 0.37) and pleuritis (OR 0.60), and a substantial decrease in liver (OR 0.45) and organ (OR 0.72) condemnations. Classification of the pigs did not have an effect on the prevalence of whole carcass condemnations, arthritis or abscesses. The health-classified pigs had, unexpectedly, more partial-carcass condemnations (OR 1.15) than the standard class pigs. The obtained benefits of the LSO 2000 units were an increase in daily gain (14g, about 2%), decrease in time in the finishing unit (2 days, about 2%), a decrease in mortality (OR 0.69) and a decrease in whole-carcass (OR 0.69) and partial-carcass (OR 0.81) condemnations. The LSO 2000 units did not have any advantage over the non-LSO 2000 units in their prevalence of liver or organ condemnations, arthritis, abscesses, pneumonia or pleuritis. PMID- 11267688 TI - An epidemic of East Coast fever on a dairy farm in eastern Tanzania. AB - During the period August-September 1995, an epidemic of East Coast fever occurred at a dairy farm in Morogoro region of eastern Tanzania. Due to an intensive dipping scheme since 1970, a very unstable endemic status had been established in the animals. A breakdown in the dipping scheme caused a major disease outbreak; the dip wash was not changed for 18 months prior to the outbreak and dipping continued in a dip wash of unknown strength. There was also a delay in detecting the disease at an early stage. In total, 180 out of 432 (42%) of the cattle at the farm died--resulting in a loss of Tshs. 26,330,000 (US$ 42,879). The attack risk was nearly 77%. The outbreak points to the importance of adopting integrated strategies for the control of ticks and tick-borne diseases. PMID- 11267689 TI - Blowfly strike in sheep flocks as an example of the use of a time-space scan statistic to control confounding. AB - The use of a time-space scan statistic--defined by a cylindrical window with a circular geographic base and height corresponding to time--was investigated as a method of detecting clustering in veterinary epidemiology whilst controlling confounding. The example data set consisted of farmer-recorded occurrence of body strike and breech strike between August 1998 and May 1999 in 26 sheep flocks located in two local government areas of southeastern Queensland, Australia. This information was derived from a questionnaire survey mailed to farmers. Potentially confounding factors included in the investigation were flock size (< or = median, > median), flock structure (proportion of lambs, wethers, ewes and rams), pesticide application for flystrike control (yes, no) and rainfall (< or = median, > median). The total sheep population within selected flocks was 92,660; 1012 (1.09%) and 518 (0.56%) cases of body strike and breech strike were reported in 16 and 10 flocks, respectively.Clustering analyses of body strike and breech strike were undertaken separately, because different predisposing factors are associated with these diseases. Significant clustering of body strike (28.76 degrees S, 151.82 degrees E) during March 1999 and breech strike (28.73 degrees S, 151.16 degrees E) between February and May 1999 was detected. Adjusting for flock structure, flock size, pesticide use and rainfall did not alter the most likely cluster of body strike identified--although the relative risk changed (> 10%) after adjusting for flock structure. Adjustment for flock structure and rainfall resulted in different clusters of breech strike being identified. PMID- 11267690 TI - Detection model for mastitis in cows milked in an automatic milking system. AB - Automated detection of diseases (such as mastitis) in dairy cows might be an alternative for detection by observation during milking - especially when using an automatic milking system (AMS). An outline of a detection model is given. This detection model includes time-series models for two variables (milk yield and electrical conductivity of milk), with interpolation on previous values. The model is flexible in the number of variables actually used. Parameter values and the residual variances are updated by linear regression after each milking. Alerts for mastitis are given when the residuals fall outside given confidence intervals. A data set with 111 cows for 16 months (on average, 58 lactating cows per day) was used to test the model. Depending on the chosen confidence interval, 42-44 out of 48 cases of clinical mastitis were detected; the remaining cases were not detected because not all data needed were available. These results were better than the results obtained with the model usually used on the farm. The number of false-positive alerts depended on the chosen confidence interval and was higher than the number found with the model usually used. PMID- 11267691 TI - Persistent bovine viral diarrhoea virus infection in US beef herds. AB - In the summer of 1996, we screened 18,931 calves in 128 beef herds located in five US states for persistent bovine viral diarrhea virus (BVDV) infection. Of these, 76 herds were randomly selected from the client database of collaborating veterinary practices, and 52 herds were suspected by the collaborating veterinarians to have BVDV infection based on history or clinical signs. Serum was obtained from each calf in the cooperating herds prior to 4 months of age and tested for the presence of BVDV by microtiter virus isolation. Information about each of the herds (including management practices, vaccination history, and breeding- and calving-season production measures) were collected by the collaborating veterinarians using standardized questionnaires. A total of 56 BVDV positive calves in 13 herds were identified on initial screening. Ten (19%) of the BVDV-suspect herds and three (4%) of the randomly selected herds had > or = 1 BVDV-positive calf at initial screening. Multiple BVDV-positive calves were identified in 10 of those 13 herds. Follow-up information was obtained for 54 of the 56 positive calves. Ten out of 54 (18%) died prior to weaning, and 1 (2%) was sold because of unusually poor growth. Thirty-three out of 54 (61%) of the initially positive calves remained BVDV positive at 6 months of age - confirming persistent-infection (PI) status. Dams of 45 of the 56 positive calves were tested, with 3 (7%) identified as positive - indicating most PI calves were products of acute dam infection during gestation. The proportion of cows that were pregnant at the fall 1995 pregnancy examination was 5% lower in herds with PI calves born during the 1996 calving season than in randomly selected herds without PI calves. Most of the calves we identified with persistent BVDV infections survived to weaning, and could provide a constant source of virus to the herd throughout the breeding season and early gestation. PMID- 11267692 TI - Community participation and the delivery of veterinary services in Africa. AB - Community participation is now widely promoted as an important feature of aid projects in less-developed countries. However, definitions, uses and expectations of community participation vary considerably among professionals (including veterinarians). A lack of common understanding of community participation hinders the comparison of experiences between projects and can lead to false hopes regarding how community participation should be used and what it might deliver. This paper provides an overview of experiences with community participation in animal-health service development and research in Africa. By examining two types of community-based animal-health intervention, the paper also describes how community participation can vary in veterinary projects and relates this variation to project impact and sustainability. Projects that encourage types of community participation such as interactive participation and self-mobilisation are most likely to result in sustained benefits for livestock keepers. PMID- 11267693 TI - Use of interpolated climatic parameters to predict risk of blowfly-strike in Queensland sheep flocks. AB - The risk of blowfly-strike was investigated in 160 sheep flocks in Queensland, Australia. The association between blowfly-strike--measured by the use of pesticides in response to flystrike reported by flock managers during the period 1994-1997--and interpolated rainfall and temperature data was examined using survival analysis and stepwise regression. The statistical model explaining the most variation in risk of blowfly-strike included average maximum temperature and its second-order polynomial 4 months preceding flystrike. The model was validated using reports of blowfly-strike from 87 flocks during the period 1997-1999 (collected using the same method as for model construction) and interpolated maximum temperature data. Although there was overlap of 95% confidence intervals derived for model coefficients estimated using both construction and validation data sets, false-negative and false-positive misclassification percentages were 33.3 and 53.3%, respectively. False-negative misclassification was greater for flocks located in southern Queensland, and false-positive misclassification was greater for northern Queensland flocks. PMID- 11267694 TI - Distemper vaccination of farmed fur animals in Finland. AB - The most important farmed fur animal species in Finland are the American mink (Mustela vison), blue fox (Alopex lagopus), silver fox (Vulpes vulpes) and raccoon dog (Nyctereutes procyonoides); all are susceptible to canine distemper. The only distemper vaccines currently available are for mink, although they also have been used for fox and raccoon dogs in emergency situations. The efficacy in eliciting neutralizing antibodies and the safety of three mink-distemper vaccines were studied under field conditions with mink and silver fox. Two of the vaccines were also studied with raccoon dogs and blue fox. All three vaccines elicited a satisfactory antibody response in mink, whereas the response varied in the other species. No side effects were observed in any species tested. One of the vaccines was safe and immunogenic in all four species. PMID- 11267695 TI - Green fluorescent protein as a secretory reporter and a tool for process optimization in transgenic plant cell cultures. AB - Green fluorescent protein (GFP) is an attractive reporter for bioprocess monitoring. Although expression of GFP in plants has been widely reported, research on the use of GFP in plant cell cultures for bioprocess applications has been limited. In this study, the suitability of GFP as a secretory reporter and a useful tool in plant cell bioprocess optimization was demonstrated. GFP was produced and secreted from suspension cells derived from tobacco that was transformed with a binary vector containing mgfp5-ER cDNA, a modified GFP for efficient sorting to the endoplasmic reticulum, under control of the CaMV 35S promoter. For cell line gfp-13, extracellular and intracellular GFP accumulated to 15.4 and 29.4 mg x 1(-1), respectively. Extracellular GFP accounted for 30.9% of the total extracellular protein. The molecular mass of extracellular GFP was nearly identical to that of a recombinant GFP standard, indicating cleavage of the signal sequence. Neomycin phosphotransferase II, a cytosolic selection marker, was found almost exclusively in cellular extracts with less than 2% in the extracellular medium. These results suggest that extracellular GFP is most likely the result of secretion rather than nonspecific leakage from cells. Furthermore, medium fluorescence intensity correlated nicely with extracellular GFP concentration supporting the use of GFP as a quantitative secretory reporter. During the batch cultivation, culture GFP fluorescence also followed closely with cell growth. A medium feeding strategy was then developed based on culture GFP fluorescence that resulted in improved biomass as well as GFP production in a fed batch culture. PMID- 11267696 TI - Ethanol production from wheat straw hemicellulose hydrolysate by Pichia stipitis. AB - Ethanol production was evaluated from wheat straw (WS) hemicellulose acid hydrolysate using an adapted and parent strain of Pichia stipitis. NRRL Y-7124. The treatment by boiling and overliming with Ca(OH)(2) significantly improved the fermentability of the hydrolysate. Ethanol yield (Yp/s) and productivity (Qp av) were increased 2.4+/-0.10 and 5.7+/-0.24 folds, respectively, compared to neutralized hydrolysate. Adaptation of the yeast to the hydrolysate resulted further improvement in yield and productivity. The maximum yield was 0.41+/-0.01 g(p) g(s)(-1), equivalent to 80.4+/-0.55% theoretical conversion efficiency. Acetic acid, furfurals and lignins present in the hydrolysate were inhibitory to microbial growth and ethanol production. The addition of these inhibitory components individually or in various combinations at a concentrations similar to that found in hydrolysate to simulated medium resulted a reduction in ethanol yield (Yp/s) and productivity (Qp av). The hydrolysate used had the following composition (expressed in g x l(-1)): xylose 12.8+/-0.25; glucose 1.7+/-0.3; arabinose 2.6+/-0.21 and acetic acid 2.7+/-0.33. PMID- 11267697 TI - Development of stable cell lines for production or regulated expression using matrix attachment regions. AB - One of the major hurdles of isolating stable, inducible or constitutive high level producer cell lines is the time-consuming selection procedure. Given the variation in the expression levels of the same construct in individual clones, hundreds of clones must be isolated and tested to identify one or more with the desired characteristics. Various boundary elements (BEs), matrix attachment regions, and locus control regions (LCRs) were screened for their ability to augment the expression of heterologous genes in Chinese hamster ovary (CHO) cells. Of the chromatin elements assayed, the chicken lysozyme matrix-attachment region (MAR) was the only element to significantly increase stable reporter expression. We found that the use of the MAR increases the proportion of high producing clones, thus reducing the number of clones that need to be screened. These benefits are observed both for constructs with MARs flanking the transgene expression cassette, as well as when constructs are co-transfected with the MAR on a separate plasmid. Moreover, the MAR was co-transfected with a multicomponent regulatable beta-galactosidase expression system in C2C12 cells and several clones exhibiting regulated expression were identified. Hence, MARs are useful in the development of stable cell lines for production or regulated expression. PMID- 11267698 TI - Mechanisms of wood degradation by brown-rot fungi: chelator-mediated cellulose degradation and binding of iron by cellulose. AB - Iron, hydrogen peroxide, biochelators and oxalate are believed to play important roles in cellulose degradation by brown-rot fungi. The effect of these compounds in an 'enhanced' Fenton system on alpha-cellulose degradation was investigated specifically in regard to molecular weight distribution and cellulose-iron affinity. This study shows that the degradative ability of an ultrafiltered low molecular weight preparation of chelating compounds isolated from the brown-rot fungus Gloeophyllum trabeum (termed 'Gt chelator') increased with increasing Gt chelator concentration when the FeIII to Gt chelator ratio was greater than about 30:1. When this ratio was less than 30:1, increasing Gt chelator concentration did not accelerate cellulose degradation. In excess hydrogen peroxide, cellulose degradation increased and then decreased with increasing iron concentration when FeIII was present in excess of the Gt chelator. The critical ratio of FeIII to Gt chelator varied depending on the concentration of hydrogen peroxide in the system. Increasing iron concentration above a critical iron:chelator ratio inhibited cellulose degradation. The optimum pH for cellulose degradation mediated by Gt chelator was around 4.0. A comparison of the effects of 2,3-DHBA (a chelator that reduces iron similarly to Gt chelator) and Gt chelator with respect to cellulose degradation demonstrated the same pattern of cellulose degradation. Cellulose-iron affinity studies were conducted at three pH levels (3.6, 3.8, 4.1), and the binding constants for cellulose-FeIII, cellulose-FeII and cellulose-FeIII in the presence of Gt chelator were calculated. The binding constants for cellulose-FeIII at all three pH levels were much higher than those for cellulose-FeII, and the binding constants for cellulose-FeIII in the presence of Gt chelator were very close to those for cellulose-FeII. This is probably the result of FeIII reduction to FeII by Gt chelator and suggests that chelators from the fungus may be able to sequester iron from cellulose and reduce it in near proximity to the cellulose and thereby better promote depolymerization. The free radical generating system described has potential for use in a variety of industrial processing and pollution control applications. PMID- 11267699 TI - Analysis of alterations in gene expression after amplification of recombinant genes in CHO cells. AB - Dihydrofolate reductase (DHFR) based amplification of recombinant genes using increasing concentrations of methotrexate (MTX) is a common method to establish CHO cell lines producing high amounts of the desired protein. Once, cell lines with highly amplified target genes and good expression rates are isolated, further characterization of their transcriptional pattern is intended to clarify the question what other factors are elevated, as a prerequisite or consequence of recombinant protein production. In order to define genes which are upregulated in a cell line that shows high production rates, we have investigated alterations in gene expression which occur beside amplification of the recombinant genes. For this purpose, the suppression subtractive hybridization method was used to create a cDNA library enriched for differentially expressed sequences in the recombinant antibody producing CHO cell line versus the original counterpart. Differential expression was confirmed by Northern blotting and Northern ELISA. In addition to the expected recombinant genes, we have identified 5 transcripts which are upregulated in the recombinant cell line. One sequence has not been found in existing data bases, the others revealed to be genes involved in protein synthesis and regulation of transcription. Furthermore, an alternatively spliced, non-functional form of the DHFR mRNA was detected, suggesting a dramatic increase of the selection pressure exerted by MTX. PMID- 11267701 TI - Study of protein adsorption on indigo particles confirms the existence of enzyme- indigo interaction sites in cellulase molecules. AB - Adsorption of several crude and purified cellulases (from Trichoderma reesei, Penicillium verruculosum and Chrysosporium lucknowense) on indigo particles and Avicel cellulose was studied. Much higher amounts of protein were bound to indigo than to cellulose under similar conditions. For different purified enzymes, the quantity of bound protein per mg of adsorbent (indigo or cellulose) varied in the range of 57-111 and 0-62 microg x mg(-1), respectively. However, in general, the enzyme adsorption on indigo was less specific than the adsorption on cellulose. Three endoglucanases, having the highest indigo-binding ability, demonstrated the best washing performance in the process of enzymatic denim treatment. These data confirmed our previous findings that certain cellulases, which have indigo binding sites (clusters of closely located aromatic and other non-polar residues) on the surface of their molecules, may remove indigo from the denim fabric better than cellulases with lower content of hydrophobic residues exposed to solvent. PMID- 11267700 TI - Engineering zucchini yellow mosaic potyvirus as a non-pathogenic vector for expression of heterologous proteins in cucurbits. AB - Plant virus vectors provide an attractive biotechnological tool for the transient expression of foreign genes in whole plants. As yet there has been no use of recombinant viruses for the improvement of commercial crops. This is mainly because the viruses used to create vectors usually cause significant yield loss and can be transmitted in the field. A novel attenuated zucchini yellow mosaic potyvirus (AG) was used for the development of an environmentally safe non pathogenic virus vector. The suitability of AG as an expression vector in plants was tested by analysis of two infectious viral constructs, each containing a distinct gene insertion site. Introduction of a foreign viral coat protein gene into AG genome between the P1 and HC-Pro genes, resulted in no expression in planta. In contrast, the same gene was stably expressed when inserted between NIb and CP genes, suggesting that this site is more suitable for a gene vector. Virus mediated expression of reporter genes was observed in squash and cucumber leaves, stems, roots and edible fruit. Furthermore, AG stably expressed human interferon alpha 2, an important human anti-viral drug, without affecting plant development and yield. Interferon biological activity was measured in cucumber and squash fruit. Together, these data corroborate a biotechnological utility of AG as a non pathogenic vector for the expression of a foreign gene, as a benefit trait, in cucurbits and their edible fruit. PMID- 11267702 TI - Toxicological profile of diethyl phthalate: a vehicle for fragrance and cosmetic ingredients. AB - Diethyl phthalate (DEP; CAS No. 84-66-2) has many industrial uses, as a solvent and vehicle for fragrance and cosmetic ingredients and subsequent skin contact. This review focuses on its safety in use as a solvent and vehicle for fragrance and cosmetic ingredients. Available data are reviewed for acute toxicity, eye irritation, dermal irritation, dermal sensitization, phototoxicity, photoallergenicity, percutaneous absorption, kinetics, metabolism, subchronic toxicity, teratogenicity, reproductive toxicity, estrogenic potential, genetic toxicity, chronic toxicity, carcinogenicity, in vitro toxicity, ecotoxicity, environmental fate and potential human exposure. No toxicological endpoints of concern have been identified. Comparison of estimated exposure (0.73 mg/kg/day) from dermal applications of fragrances and cosmetic products with other accepted industrial (5 mg/m(3) in air) and consumer exposures (350 mg/l in water; 0.75 mg/kg/day oral exposure) indicates no significant toxic liability for the use of DEP in fragrances and cosmetic products. PMID- 11267703 TI - Effects of a water-soluble extract of rosemary and its purified component rosmarinic acid on xenobiotic-metabolizing enzymes in rat liver. AB - The effects of a water-soluble extract (WSE) of rosemary and its purified antioxidant rosmarinic acid (RA) on xenobiotic metabolizing enzymes (XME) were studied in rat liver after dietary administration. The modulation of phase I enzymes such as cytochrome P450 (CYP) 1A, 2B, 2E1, 3A, and phase II enzymes such as glutathione S-transferase (GST), quinone reductase (QR) and UDP glucuronosyltransferase (UGT) was evaluated by measuring enzyme activities with specific substrates. Protein levels of CYPs and rGST A1/A2, A3/A5, M1, M2 and P1 were measured using antibodies in Western blots. Caffeic acid was also studied because it results from RA biotransformation in rat after oral administration. Male SPF Wistar rats received the different compounds at 0.5% (w/w) incorporated into their diet for 2 weeks. WSE, containing RA, flavones and monoterpenes enhanced CYP 1A1, 2B1/2, 2E1 and GST (especially rGST A3/A5, M1 and M2), QR and UGT. On the contrary, no modification of XME was observed in response to RA or CA (except for a slight increase of UGT activity after CA treatment). The induction of XME by WSE could be attributed to flavones, monoterpenes or an additive effect of all components. PMID- 11267704 TI - The effect of sodium nitrite on some parameters of the immune system. AB - The effect of orally administered sublethal doses of 25, 50 and 100 mg/kg of sodium nitrite in drinking water ad lib. for 21 days on the immune response of Balb/c mice was investigated. The immunological parameters were examined at three phases: 1 day (phase A), 1 week (phase B) and 3 weeks (phase C) after the end of exposure to sodium nitrite. A significant decrease in dose-dependent manner was obtained in the following tests: lymphocyte percentages, concanavalin A (Con A)- and lipopolysaccharide (LPS)-induced lymphocyte proliferation assessed by the colorimetric MTT method, natural killer (NK) cell activity against WEHI-164 target cells, as well as IgM and IgG titers against injected sheep erythrocytes. Maximum suppressions were obtained in phase A after treatment with sodium nitrite at 100 mg/kg including lymphocyte count (17.5%), Con A-induced lymphocyte proliferation (40.1%), LPS-induced lymphocyte proliferation (31.4%), IL-2 stimulated NK cell activity (59.2%), unstimulated NK cell activity (59.6%), IgM titer (57.5%) and IgG titer (61.1%). On the other hand, a significant dose dependent increase in neutrophil count (71.3%) in phase A and phagocytic activation (133%) in the first two phases was obtained using the nitroblue tetrazolium (NBT) assay in the presence of phorbol myristate acetate (PMA). It was found that the immunosuppressive effect of sodium nitrite is reversible after cessation of exposure. PMID- 11267705 TI - Lymphocyte cytochrome P450-CYP2E1 expression in human IDDM subjects. AB - Cytochrome P4502E1, a phase I enzyme, has been shown to be induced in liver samples from diabetic and obese rats. One study demonstrated elevated levels of CYP2E1 in children with IDDM, using Western blot analysis. The aim of this investigation was to determine CYP2E1 expression in peripheral blood lymphocytes from eight well-controlled IDDM and eight sex- and age-matched control subjects using Western blot analysis and Phoretix image analysis. Levels of CYP2E1 were low to undetectable in human lymphocytes from healthy control subjects. However, levels of CYP2E1 were elevated in lymphocytes from IDDM subjects (mean 3.1-fold higher). The elevated levels of CYP2E1 in the IDDM subjects showed no correlation with HbA(1c) nor duration of IDDM; however, there were marked inter-individual differences in levels of induction of CYP2E1 between all subjects. The results of this study suggest that in human IDDM subjects, even with good metabolic control, expression of CYP2E1 is elevated when compared to controls. CYP2E1 is known to generate ROS in vivo, the modulation of this isoform in lymphocytes from IDDM subjects, could well add to the oxidative stress associated with IDDM and the development of associated complications. PMID- 11267706 TI - Topical permethrin exposure inhibits antibody production and macrophage function in C57Bl/6N mice. AB - Permethrin was applied to the shaved dorsal interscapular region of C57Bl/6N mice at doses of 0.5, 1.5 or 5.0 microl/day. These doses corresponded to approximately 22-220 mg/kg/day topical insecticide. Mice were exposed to permethrin in this manner daily for 10 or 30 consecutive days, or every other day for 7 or 14 exposures. The splenic macrophage chemiluminescent response was depressed in a dose-dependent manner at 2 and 10 days post-exposure to permethrin. Phagocytic ability of macrophages was not inhibited. Antibody production as shown by plaque forming cell (PFC) assay decreased significantly after 10 consecutive days of exposure to permethrin. These data indicate that topical permethrin exposure may produce systemic immune effects. PMID- 11267707 TI - Urinary thiocyanate levels as a biomarker for the generation of inorganic cyanide from benzyl cyanide in the rat. AB - A colorimetric procedure was developed and validated for the determination of thiocyanate in rat urine over the concentration range of 7-7000 microg/ml. It was applied to the determination of thiocyanate following its oral administration to male and female rats. The mean percentage urinary recoveries of sodium thiocyanate given by oral gavage at 10 and 100 mg/kg were 60 and 39%, respectively, for male rats and 89 and 73% for females over a period of 3 days. Most of the elimination occurred in the 0-48-h period post-dosing but significant amounts were still being excreted in the 48-72-h period. It was concluded from these results that the recoveries of urinary thiocyanate were such that this anion was suitable for use as a biomarker for the release of cyanide from organonitriles such as benzyl cyanide. Benzyl cyanide (150 mg/kg) administered orally to rats led to markedly increased urinary thiocyanate levels; for male rats this was equivalent to 54% of the dose and for females this was 65% over a period of 3 days. When adjusted for incomplete recoveries of the marker, thiocyanate, these values equated to 61 and 89%, respectively. It was concluded that this validated assay could be used to assess cyanide release from topically applied fragrance organonitriles (Potter, J., Smith, R.L., Api, A.M., 2000. An assessment of the release of inorganic cyanide from the fragrance materials, benzyl cyanide, geranyl nitrile and citronellyl nitrile applied dermally to the rat. Food and Chemical Toxicology 39, 147-151). PMID- 11267708 TI - An assessment of the release of inorganic cyanide from the fragrance materials benzyl cyanide, geranyl nitrile and citronellyl nitrile applied dermally to the rat. AB - Organonitriles are widely used as components of fragrances that are incorporated into consumer products, many of which are for human topical use. Some organontriles are readily broken down metabolically to potentially toxic inorganic cyanide. Studies were therefore undertaken to assess whether this occurs with three representative fragrance nitriles, namely, benzyl cyanide, geranyl nitrile and citronellyl nitrile when applied dermally to the rat. The nitriles (benzyl cyanide, 150 mg/kg; geranyl and citronellyl nitriles, 400 mg/kg) were applied to the shaved backs of rats and maintained under occlusion for 24 h. Urine samples were collected for 0-24 h, 24-48 h and 48-72 h from the time of first application. These samples were analysed for thiocyanate, a biomarker for cyanide formation in vivo, as described previously (Potter, J., Smith, R.L., Api, A.M., 2000. Urinary thiocyanate levels as a biomarker for the generation of inorganic cyanide from benzyl cyanide in the rat. Food and Chemical Toxicology 39, 141-146). In the case of benzyl cyanide, there was a marked increase in urinary thiocyanate levels attributable to the release of cyanide in vivo. The amount of thiocyanate recovered was equivalent to 37% of the dose for males and 32% for females. For geranyl nitrile there was no significant increase in urinary thiocyanate excretion and there was only a marginal increase in the case of citronellyl nitrile that was equivalent to 0.40% of the applied dose for males and 0.29% for females. PMID- 11267709 TI - The in vivo dermal absorption and metabolism of [4-14C] coumarin by rats and by human volunteers under simulated conditions of use in fragrances. AB - The disposition and metabolic fate of [4-14C]coumarin in a 70% aqueous ethanol solution was studied in male Lister Hooded rats after occluded dermal application and in three male volunteers after an exposure designed to simulate that which may be encountered when using an alcohol-based perfumed product. In both cases, the 6-h exposure was 0.02 mg/cm(2) (rats 0.023 mg/kg and humans 0.77 mg/kg). In both, coumarin was quickly absorbed, distributed and excreted in urine and feces, although fecal excretion of coumarin in humans was only 1% of the applied dose as opposed to 21% in rats. Total absorption was 72% of the applied dose with rats and 60% with humans. Peak plasma radioactivity in both was at 1 h. The mean plasma half-life of coumarin and metabolites was approximately 1.7 h for humans and 5 h for rats. In humans, coumarin was primarily metabolized to and excreted in urine as 7-hydroxycoumarin glucuronide and 7-hydroxycoumarin sulfate. Small amounts of unconjugated 7-hydroxycoumarin and o-hydroxyphenylacetic acid (o-HPAA) were also excreted. In rats, about twenty metabolites were present, but only o HPAA was identified. These studies show the rat is a very poor model for humans and toxicity in the rat cannot be extrapolated to humans. PMID- 11267710 TI - A preliminary study of the dermal absorption of aluminium from antiperspirants using aluminium-26. AB - Aluminium chlorohydrate (ACH), the active ingredient in many antiperspirants, was labeled with the radioisotope 26Al. The labeled ACH was then fractionated into about 100 samples using gel filtration chromatography. Each fraction was analyzed for 26Al and total aluminium content. Aluminium-26 was only detected in the fractions that also contained aluminium, which verified that the ACH was uniformly labeled. 84 mg of the labeled ACH was then applied to a single underarm of two adult subjects with blood and urine samples being collected over 7 weeks. Tape-stripping and mild washings of the skin were also collected for the first 6 days. Results indicate that only 0.012% of the applied aluminium was absorbed through the skin. At this rate, about 4 microg of aluminium is absorbed from a single use of ACH on both underarms. This is about 2.5% of the aluminium typically absorbed by the gut from food over the same time period. Therefore, a one-time use of ACH applied to the skin is not a significant contribution to the body burden of aluminium. PMID- 11267711 TI - Fate of ethanol topically applied to skin. AB - Ethanol is a major component of many aerosol sprays and consumer products that are designed to contact the skin. It is theoretically possible that small amounts of ethanol from alcohol-based sprays can be absorbed across the skin or inhaled during spraying. In order to assess the potential systemic dose, three parameters were measured: the evaporation of [14C]ethanol from the skin surface, the in vitro penetration of [14C]ethanol through excised pig skin and the ethanol concentration in the blood of human volunteers following simulated use of an alcohol based deodorant spray. The rate of evaporation from Benchkote and whole pig skin was similar (t(1/2)=13.6 sec and 11.7 sec, respectively) while that from glass was longer (t(1/2)=24.8 sec). Ethanol penetration through pig skin in vitro was greater in occluded cells than in non-occluded cells (2.19 mg/cm(2) and 0.10 mg/cm(2) in 24 hours, respectively). At the maximum flux seen in this experiment under occlusion, the amount of ethanol penetrating from a 1 m(2) area of skin would give a blood alcohol level of about 4 mg% in a 70-kg man. In the human use study, none of the blood samples taken from 16 human volunteers exhibited a detectable level of alcohol. These studies provide evidence that a systemic dose of ethanol is likely to be very low after the use of formulations delivering ethanol to the skin. PMID- 11267712 TI - "IARC Group 2B carcinogens" reported in cigarette mainstream smoke. AB - In the third and final part of a series surveying the international literature on the "IARC carcinogens" in cigarette mainstream smoke, the "IARC Group 2B carcinogens" are reviewed. A search of the published literature shows that of 227 chemical components classified as Group 2B, that is, "possible carcinogens," by the International Agency for Research on Cancer (IARC), 48 have previously been reported in cigarette mainstream smoke. Owing to its highly interactive molecular nature, removal from or inhibition of a given mutagenic or carcinogenic chemical within the complex aerosol mixture cannot reliably be predicted to reduce either the overall mutagenicity or carcinogenicity. However, in the absence of experimental data demonstrating an increase in adverse biological activity resulting from removal or inhibition of a potentially carcinogenic constituent, negation of the activity of the potential carcinogen may be considered as a desirable circumstance. PMID- 11267713 TI - Perinatal mortality rises both with prematurity and with the degree to which the baby's birthweight is below that expected for gestational age. PMID- 11267714 TI - Systematic reviews to evaluate diagnostic tests. AB - Diagnostic testing and screening is a critical part of the clinical process because inappropriate diagnostic strategies put patients at risk and entail a serious waste of resources. It is being increasingly recognised that absence of clear summaries of individual research studies on the repeatability, accuracy and impact of tests, which are often scattered across many different journals, is a major impediment. Just as the need to develop means to systematically review research assessing the effectiveness of treatments has been pursued over the last decade, so more recently attention has focused on how research on diagnostic tests might also be systematically reviewed. These reviews present a huge methodological challenge. This paper describes the use of a systematic approach to collation, appraisal and synthesis of information contained in the primary literature about accuracy of diagnostic strategies. PMID- 11267715 TI - Pathophysiology of preeclampsia and the role of serotonin. AB - Hypertensive disorders constitute the most common medical complications of pregnancy. In normal pregnancy, impressive physiological changes take place in the maternal cardiovascular system. Morphological changes are the result of invasion of migratory trophoblast cells into the walls of the spiral arteries. After destruction of elastic, muscular and neural tissue in the media, the trophoblast cells get incorporated into the vessel wall and the endothelial lining of the spiral arteries is restored. The physiological changes create a low resistance, low-pressure, high-flow system with the absence of maternal vasomotor control. Biochemical adaptations in maternal vasculature include changes in the prostaglandin system, the renin-angiotensin-aldosteron system and the kallikrein kinin system. In preeclampsia, physiological changes in the spiral arteries are confined to the decidual portion of the arteries. Myometrial segments remain anatomically intact and fail to dilate. In addition, the adrenergic nerve supply is left intact. The cause of this impaired endovascular trophoblast invasion is not yet elucidated. But in combination with the imbalance between vasodilator and vasoconstrictor eicosanoids, it gives rise to reduced perfusion of the intervillous space. In the absence of an adequate production of antiaggregatory prostacyclin (PGI(2)), nitric oxide, or both, surface-mediated platelet activation is supposed to occur on the surface of the spiral arteries. Because platelets are the principal source of circulating serotonin, the increased platelet aggregation in preeclampsia causes an increase in serotonin levels. Interaction of serotonin with serotonin(1)- or serotonin(2)-receptors depends on the state of the endovascular trophoblast or endothelium in the spiral arteries and has opposite effects with regard to vasodilating and vasoconstrictive influences. PMID- 11267716 TI - Pharmacological treatment of severe hypertension in pregnancy and the role of serotonin(2)-receptor blockers. AB - Hypertensive disorders of pregnancy are the leading cause of maternal and perinatal mortality and morbidity in developing and developed countries. The etiology of preeclampsia is still unknown. Delivering the baby is the only definite treatment. The benefits of acute pharmacological control of severe hypertension prior to and/or post-delivery are generally accepted. Most drugs commonly used in the management of severe hypertension in pregnancy have significant maternal and/or neonatal adverse side effects. Furthermore, some are not effective to acutely lower the blood pressure in patients with a hypertensive crisis. Until recently not one of the commonly used antihypertensive drugs has been tailored to the pathophysiology of severe preeclampsia, being a clinical syndrome characterized by endothelial cell dysfunction, vasospasm and platelet aggregation. Ketanserin, a serotonin(2)-receptor blocker, is a drug that appears to be tailored for treating this pregnancy-associated enthothelial cell dysfunction. The results of several prospective trials show that there is a definite place for serotonin(2)-receptor blockers in the treatment of pregnancy induced hypertensive disorders. This review provides a summary on the more established drugs as well as on some of the newer antihypertensive drugs used in pregnancy with emphasis on the existing experience with ketanserin. PMID- 11267717 TI - Non-invasive early prenatal diagnosis using fluorescent in situ hybridization on transcervical cells: comparison of two different methods for retrieval. AB - OBJECTIVE: We compared the efficiencies of uterine and endocervical lavage to retrieve fetal cells from first trimester pregnancies for further analysis with fluorescent in situ hybridization (FISH). STUDY DESIGN: Transcervical cell (TCC) samples were collected at 7-10 weeks of gestations by uterine lavage (13 women) and by endocervical lavage (12 women) who were scheduled for volunteer termination of pregnancy. A sample of placenta was also obtained for cytogenetic analysis to confirm the sex or genotype in the end of the procedure. FISH was performed using probes for the chromosomes 18, X and Y in a three color hybridization protocol. The statistical analysis included chi(2)-analysis, and t test. RESULTS: Sufficient cells were obtained in 12 of the 13 (92.3%) in uterine lavage and 10 of the 12 (83.3%) in endocervical lavage group for FISH procedures for fetal sex prediction. The mean success rate of signal detection for FISH procedure was 91.7% (range 83-97%). Fetal sex was correctly predicted in 11 of 12 (91.6%) with uterine lavage and 8 of 10 (80.0%) in endocervical lavage and the difference was statistically insignificant. CONCLUSION: This study demonstrated that there are available cells of fetal origin in the lower part of the uterus and these cells may be collected successfully as early as 7 weeks of the gestation. In addition, our results show that endocervical lavage method is as effective as uterine lavage. FISH has been successfully used to detect status of aneuploidy and sex of the fetus from TCC. PMID- 11267718 TI - Late normalisation of uterine artery velocimetry in high risk pregnancy. AB - OBJECTIVE: To test whether late normalisation of abnormal uterine velocimetry is a favourable prognostic factor in high risk pregnancies. STUDY DESIGN: Uterine artery colour Doppler velocimetry was performed at 24, 28-30 and 32-34 weeks in 282 high risk pregnancies treated with low dose aspirin. RESULTS: 88 patients had abnormal waveforms at 24 weeks and 77 delivered after the second assessment at 28 weeks. Of these, 38 (49%) had a normalisation of Doppler indices by 34 weeks. Compared with the persistently abnormal Doppler group, these patients delivered fewer small for gestational age babies (5/38 versus 26/39; p=0.0001) and had less gestational hypertension without proteinuria (3/38 versus 15/39; p=0.004). No patients with preeclampsia or other severe complications of pregnancy were observed in the normalised group. CONCLUSIONS: Although abnormal uterine artery velocimetry at 24 weeks is predictive of adverse pregnancy outcome, nearly half have late normalisation of the Doppler indices and a better perinatal outcome. Persistently abnormal waveforms are related to the worst pregnancy outcome. PMID- 11267719 TI - The effect of 17beta-estradiol on isolated omental arteries from preeclamptic women. AB - OBJECTIVE: To study the effect of 17beta-estradiol on isolated omental arteries from preeclamptic women. STUDY DESIGN: Rings of omental artery with intact endothelium were mounted in organ chambers for isometric tension recording. We studied the effect of pharmacological concentrations of 17beta-estradiol on potassium chloride-induced tension and the concentration-contraction relationships for norepinephrine and calcium. RESULTS: Cumulative application of 17beta-estradiol, in a concentration-dependent manner, relaxed potassium chloride contracted rings. Sixty minutes of incubation with 17beta-estradiol (10(-5)mol/l) attenuated the tension developed in response to potassium chloride, norepinephrine and calcium. Tamoxifen (10(-6)mol/l) did not antagonize the inhibitory actions of 17beta-estradiol. CONCLUSIONS: Pharmacological concentrations of 17beta-estradiol retain the capability for relaxing omental artery rings from preeclamptic women. The loss of refractoriness to norepinephrine, increased responsiveness to calcium ions and decreased ability of 17beta-estradiol to inhibit calcium-induced tension may be responsible for increased vascular reactivity in preeclampsia. PMID- 11267720 TI - Second trimester termination of pregnancy for fetal anomaly or death: comparing mifepristone/misoprostol to gemeprost. AB - OBJECTIVE: To assess the effect of changing the regimen for second trimester induction of labour from gemeprost to mifepristone/misoprostol. DESIGN AND SETTING: A retrospective study at a university teaching hospital over the 5-year period 1993-1997. SUBJECTS, METHODS and REGIMENS: 68 patients, 34 in the gemeprost group and 34 in the mifepristone/misoprostol group. The gemeprost group received 1mg vaginally every 3h to a maximum of five doses. The mifepristone/misoprostol group were pre-treated with 600 mg mifepristone orally followed by 800 microg misoprostol vaginally and then 400 microg orally every 3h to a maximum of four oral doses. MAIN OUTCOME MEASURES: Induction to abortion interval; delivery within 24h. RESULTS: The mifepristone/misoprostol group had a lower induction to abortion interval compared to the gemeprost group (median 8.9h versus 19.8h, respectively, p<0.01). The mifepristone/misoprostol regimen was more successful than the gemeprost regimen; 94% versus 68%, respectively, aborted without extra medical or surgical intervention, p=0.02. There were no significant differences in side effects, analgesia requirements or complications between the two groups. Three patients with previous Caesarean sections had a ruptured uterus; two from the gemeprost group and one from the mifepristone/misoprostol group. CONCLUSIONS: The new mifepristone/misoprostol regimen was more effective in second trimester induction of labour. Induction of labour with misoprostol or gemeprost should be used with care in patients with a previous Caesarean section. PMID- 11267721 TI - HELLP syndrome and factor V Leiden. AB - The association of thrombophilia and obstetrical complications is documented and well consistent with the hypothesis of an insufficient placental perfusion due to fibrin deposition as a major underlying pathophysiological mechanism. Factor V Leiden is one of the most frequent thrombophilic mutations. A high prevalence of this mutation has recently been reported in a group of 21 German women with haemolysis, elevated liver enzymes, low platelets (HELLP) syndrome. In this respect, we studied the prevalence of factor V Leiden in 18 women who were consecutively diagnosed at our Department of Obstetrics and Gynaecology as having HELLP syndrome, between 1995 and 1999. Women were tested either at the time of diagnosis or months or years after delivery for coagulation parameters, protein C (PC), protein S (PS), antithrombin III, lupus-like anticoagulant, anticardiolipin antibodies (ACA), activated protein C (APC) resistance and detection of the G1691A mutation (factor V Leiden). In all women, the parameters studied were normal and in none of the investigated cases was the G1691A mutation found. HELLP being a severe form of preeclampsia, we think that the reported association between factor V Leiden and HELLP may reflect the well-known association with preeclampsia. PMID- 11267722 TI - Correlations between serum assays of human chorionic gonadotrophin (hCG) and human placental lactogen (hPL) and pre-eclampsia or intrauterine growth restriction (IUGR) among nulliparas younger than 38 years. AB - OBJECTIVE: To study the relation between serum human chorionic gonadotrophin (hCG) levels measured at 15-18 weeks and gestational disorders, assess their correlation with the artery uteroplacental Doppler (AUD) at 24 weeks among nulliparas, and assess the predictivity of the hCG/hPL (human placental lactogen) ratio for pre-eclampsia. STUDY DESIGN: Retrospective study of two groups of women younger than 38 years old: one with an elevated serum hCG level (2 MoM (multiples of the median) or more) and a normal fetal karyotype (group A), and the other with a lower hCG level (group B). Within each group, we studied the nulliparas separately (respectively groups AO and BO). We analyzed the double screening, elevated hCG levels with abnormal AUD, for the predicting of hypertensive disorders. RESULTS: Elevated hCG levels were significantly (p<0.05) more prevalent among women who developed gestational diabetes (groups A and AO) and among nulliparas with pregnancy-induced hypertension and pre-eclampsia (AO). Among nulliparas, the combination of the hCG assay and a subsequent Doppler increased the positive predictive value (PPV) of the assay from 19 to 75%, without reducing its negative predictive value (NPV) for gestational vascular disorders. The hCG/hPL ratio did not improve the predictivity of the hCG assay alone for pre-eclampsia. CONCLUSIONS: An hCG level of 2 MoM or more at 15-18 weeks identifies a group of women at risk of gestational vascular disorders; it therefore ought to lead to an AUD at 24 weeks. This double screening should be able to define a population of women at risk of developing a hypertensive disorder, who could thus benefit from a preventive treatment, as aspirin. PMID- 11267723 TI - Neonatal outcome of spontaneous and assisted twin pregnancies. AB - OBJECTIVES: Over the last 10 years, diffusion of assisted reproduction techniques (ovarian stimulation, IVF, GIFT) has led to an increased incidence of multiple pregnancies and consequently, of the related obstetric-neonatal problems. In this study, multiple births have been studied, with particular reference to the twin births occurring in the Gemelli hospital, Rome. The hospital is also a reference centre for obstetric pathologies and infertility treatment. In particular, attention has been focused on neonatal outcome, comparing twins born from spontaneous and assisted pregnancies. STUDY DESIGN: 228 neonates from spontaneous twin pregnancies and 32 from assisted twin pregnancies were taken into consideration with regard to: premature birth, low birth-weight, intrauterine growth retardation, weight discordance, Apgar score, major neonatal diseases, and mortality. RESULTS: Results showed a significant higher incidence of prematurity and low birth-weight, as well as a significant lower gestational age, occurring more frequently in twins resulting from assisted pregnancies than in twins from spontaneous pregnancies. Furthermore, the incidence of severe depression at birth and respiratory disease was significantly higher in twins from assisted pregnancies than in those from spontaneous pregnancies, despite similar gestational age and birth-weight. PMID- 11267724 TI - The vaginal patch plastron for vaginal cure of cystocele. Preliminary results for 47 patients. AB - OBJECTIVE: We describe a new surgical technique (the vaginal plastron) for the treatment of cystocele by the vaginal route. The technique is based on bladder support by a vaginal strip (6-8cm in length and 4cm in width), isolated from the anterior colpocele, left attached to the bladder, associated with a suspension of this strip by its fixation to the tendinous arch of the pelvic fascia by six lateral sutures (three on each side of the plastron). The vaginal plastron is then covered by tucking it under the anterior colporraphy. STUDY DESIGN: We evaluated the short-term functional and anatomical results of the first 47 patients to have undergone this treatment between October 1997 and June 1998. The average age of the patients was 69 years. Cystoceles were associated with urinary stress incontinence in 38.3% of cases, with hysterocele or prolapse of the vaginal dome in 87.2% of cases, with an elytrocele in 19.1% of cases and a rectocele in 70.2% of cases. Of the 45 patients having had a hysterectomy combined with the vaginal plastron or in their past history, 44 (99.77%) had a Richter sacro-spino-fixation and 17 (38%) had a Campbell procedure combined with the vaginal plastron. All patients underwent a posterior perineorraphy with myorraphy of the elevators. RESULTS: Average follow-up was 16.4 months with extremes of 6-26 months and concerned 46 patients (one patient was unavailable). Ninety-three percent of the cystoceles were considered treated. One case of imperfect anatomical outcome was noted (persistence of stage 1 cystocele in one patient) together with two other cases of failure of the treatment of cystocele (relapse to stage 2 cystocele). CONCLUSION: Proposed as a curative treatment of cystocele and combined with the Richter fixation, the plastron provides a surgical solution to the problem of cystocele relapse arising after vaginal treatment of prolapse by sacro-spino-fixation alone (10-20% according to Richter). Short-term results are encouraging, however, medium- to long-term results (36-60 months) are necessary in order confirm the usefulness of this surgical technique. PMID- 11267725 TI - Non-invasive management of patients with suspected ectopic pregnancy: a survey among Dutch gynaecologists. AB - OBJECTIVE: To investigate if implementation of a non-invasive diagnostic work up of patients with suspected ectopic pregnancy, involving transvaginal sonography and serum human chorionic gonadotrophin (hCG) measurement decreases the likelihood of performing diagnostic laparoscopies. STUDY DESIGN: We interviewed 27 Dutch gynaecologists using 16 structured case summaries in a fractional factorial design. Each case summary concerned a hypothetical patient with suspected ectopic pregnancy. For each case presentation, the gynaecologists were asked for their inclination to perform laparoscopy. RESULTS: There were substantial differences in the degree to which data from a non-invasive work-up influenced the decision to perform a laparoscopy. Some gynaecologists would perform laparoscopy in all 16 patients at the first visit, whereas others would initially admit none of them. CONCLUSION: Dutch gynaecologists seem to be familiar with a non-invasive diagnostic approach in women with suspected ectopic pregnancy. However, there are considerable differences in management approach probably due to individual variability in weighing the risks and benefits of expectant management. PMID- 11267726 TI - The role of estrogen replacement therapy in Alzheimer's disease. AB - Multiple factors appear to contribute to the expression of Alzheimer's disease (AD). About 30% of cases of dementia of the Alzheimer's type (DAT) can be attributed to genetic factors. These observations raise the possibility of identifying multiple interventions that may modify the disease process and, therefore, the clinical expression of the dementia. Prominent among factors that may contribute to dementia and, specifically, to dementia of the Alzheimer's type is cerebral vascular disease. Estrogen is a potent factor that not only prevents vascular disease but also improves blood flow in diseased vessels, including blood flow in regions of the brain affected by AD. Estrogen also has direct effects on neuronal function that may play an important role not only in the preservation of neurons but in the repair of neurons damaged by the disease process. These effects of estrogen on the CNS suggest that the hormone may be effective not only in the prevention of dementia but also in its treatment. Given the distressingly high prevalence of AD among older women and the exorbitant social and economic costs associated with this disorder, a true risk reduction on the order of one-third to one-half, as suggested by several recent analytical studies, would be of tremendous public health importance. PMID- 11267727 TI - Low-dose mercury induces testicular damage protected by zinc in mice. AB - OBJECTIVES: This investigation was set out to determine whether mercury at a very low dose (4ppm) induces testicular damage on murine testis, and if so whether the toxic effects of mercury could be prevented by zinc. STUDY DESIGN: One of the following solutions was administered in the drinking water of CD-1 male mice: (1) 4ppm HgCl(2); (2) 800ppm ZnCl(2); (3) 4ppm HgCl(2)+800ppm ZnCl(2); or (4) deionised water; for 12 weeks. At the expiration of the treatment period, animals were sacrificed, testes excised and weighed, and epididymal sperm number taken. The testes were processed for histological examination. RESULTS: Both zinc and mercury significantly (p<0.05) decreased the absolute and relative testicular weights, with mercury producing the highest reduction in weight. Mercury reduced significantly (p<0.05) the epididymal sperm number, while zinc and mercury/zinc produced statistically same effect with control on the sperm number. Histological study showed that mercury at the concentration employed produced remarkable degenerative lesions on the testes, as the zinc-treated group showed a normal morphology. Majority of the animals in the mercury/zinc-treated group exhibited complete or partial protection as evidenced by the morphology of the seminiferous tubules. CONCLUSION: Zinc prevents mercury-induced testicular damage in mouse. These findings highlight the risks exposure to inorganic mercury might pose to male reproduction of mice, and suggests possible therapy with zinc. Study in humans is therefore advocated. PMID- 11267728 TI - The efficacy of bladder distention therapy in the treatment of frequency and urgency. AB - BACKGROUND: The aim of this retrospective study was to evaluate whether bladder distention therapy is effective in relieving the symptoms of patients experiencing urinary frequency and urgency. MATERIALS AND METHODS: Twenty-six female patients with a history of frequency and urgency who underwent multi channel urodynamic assessment, were treated with bladder distention therapy for symptomatic relief of their symptoms. Their mean age was 51 years of age (range from 28 to 75 years old). No patient had detrusor-urethral sphincter dyssynergia. Bladder distention therapy was performed in the operating theatre under epidural anesthesia. Statistical analysis was performed with the Student's t-test. RESULTS: The treatment was successful in 9 out of 26 patients after the first attempt (34.6%) and in 10 out of 26 patients after the second attempt (38%) at 1 month follow-up (FLU) and in 4 out of 28 patients at 9 months FLU (15%). Only one out of four patients who had this treatment twice showed improvement of her symptoms. CONCLUSION: Bladder distention therapy in patients with symptoms of overactive bladder dysfunction has a very small success rate (15%) and therefore, has no place in the modern treatment of bladder dysfunction. PMID- 11267729 TI - Prognostic significance of factors affecting disease free interval and overall survival for Stage II breast cancer in Greece. A multivariate cohort study. AB - BACKGROUND: Univariate analysis evaluates the impact of a prognostic factor on survival rates, either disease free (DFI) or overall (OS). Since many of the factors are interrelated, it is difficult to predict the prognosis of an individual patient. Multivariate analysis is therefore required in order to allow factors act together thus ending in the best possible combined predicting result. METHODS: A step-up procedure (Cox's Proportional Hazards Regression model) was used to include various prognostic parameters, relating to patients themselves, to the pathology of their tumours and to the treatment schedule followed. Two hundred and sixty-nine Stage II breast cancer Greek patients, treated from 1981 until 1991 and with a median 12-year of follow-up are studied. RESULTS: Five factors were found to be significant for patients DFI. In order of relevant importance, these were the number of infiltrated nodes, tumour size, postoperative radiotherapy, adjuvant chemotherapy and patients age. Regarding patients OS, tumour size, number of positive nodes, patients' age at entry and ER/PR status were the most important ones. CONCLUSION: Our long-term (12-year), single institution, single area results, suggest that, the prognostic factors for patients DFI and OS are the same with those of series from Europe and USA. Additionally, they remain unchanged after long-term follow-up, compared to a previously reported short-term national-wide study from this country. PMID- 11267730 TI - Endometrial and other primary cancers after tamoxifen treatment of breast cancer - results of retrospective cohort study. AB - OBJECTIVES: The aim of the retrospective cohort study was to evaluate the relationship between the influence of tamoxifen on the development of endometrial and other second primary cancers in the patients with invasive breast cancer. STUDY DESIGN: A cohort of 630 women diagnosed with breast cancer from 1987 to 1994 was selected from a population-based registry; 440 patients were treated with tamoxifen and 190 patients without it. The observation period was 8.5 years (range 5-12 years). The data were analysed by the relative risk (RR) calculation at a confidence interval (CI) of 95%, using a Mantel-Haenszel chi(2)-test and Fisher's p-test to evaluate statistical significance. RESULTS: There were no statistically significant differences between the group of breast cancer patients treated with tamoxifen and without it as regards the age at the breast cancer diagnosis, family medical histories, body mass, age at menopause, fertility, diabetes, hormone replacement therapy and oestrogen-hormone replacement therapy. In 41/440 (9.3%) tamoxifen-treated patients and in 8/190 (4.2%) non-users of tamoxifen, diagnostic curettage was performed due to benign endometrial changes and endometrial cancer (EC). The difference in the proportions of patients with diagnostic curettage in both group was statistically significant (chi(2)=4.45, p=0.03). In the group of patients treated with tamoxifen, with the median treatment duration of 40 months (range 1-97 months) and in the group of patients without tamoxifen, EC was diagnosed in 11 and in two patients, respectively. The evaluated RR was 2.38 (0.53-10.61, 95% CI). The second primary cancer, excluding contralateral breast cancer and EC, was diagnosed in the group of breast cancer patients treated with tamoxifen and without it in almost the same percentage, i.e. in 12 patients (3%) in the group of patients who were treated with tamoxifen and in 10 patients (5%) in the group of patients without tamoxifen treatment. CONCLUSION: Despite the fact that the calculated RR of EC in our study (2.4) was not statistically significant, due to a small number of patients, our results support the IARC evaluation that tamoxifen is carcinogenic to humans. Our data also suggest that tamoxifen does not increase the risk of other second primary cancers. However, the risk of individual second primary cancers cannot be reliably assessed due to a limited number of patients. PMID- 11267731 TI - Primary adenocarcinoma of the rectovaginal septum: a case report and literature review. AB - The authors report the third case of primary adenocarcinoma of the rectovaginal septum without associated endometriosis and discuss the pathogenesis of this tumour. Some of the tumour cells were stained with OC 125 antibody which recognises epithelium of coelomic origin; adenocarcinoma of the rectovaginal septum may arise directly from embryological Mullerian remnants. PMID- 11267732 TI - The risk of neonatal death in relation to birth weight and maternal hypertensive disease in infants born at 24-32 weeks. AB - OBJECTIVE: To determine the risk of neonatal death (NND) in relation to birth weight for gestational age and the presence or absence of maternal hypertensive disease in preterm neonates. DESIGN: Record linkage of maternity data and neonatal mortality data. SETTING: Scotland, UK. POPULATION: A group of 6946 live singleton preterm neonates without lethal congenital abnormalities born at 24-32 weeks between 1986 and 1992 inclusive. This group included 1448 cases of maternal hypertensive disease and 850 neonatal deaths. MAIN OUTCOME MEASURE: Neonatal death. RESULTS: The median birth weight for each gestational week was estimated from a fitted curve and each birth weight was recalculated as a multiple of the relevant median. The frequency of NND was much higher at lower gestations (73% at 24 weeks down to 2% at 32 weeks). Though the overall frequency of NND was lower in cases with hypertensive disease (8.6% versus 13.2%) this can be attributed to the fact that there were relatively fewer hypertensive cases in the high risk group at 24-27 weeks. In the 5498 cases not associated with maternal hypertensive disease, there were 726 NNDs. The mean MoM of birthweight for these NNDs was 0.982 (95% CI 0.967-0.996); this was only marginally different from the population mean (0.998; 95% CI 0.993-1.004). In the 1448 cases with maternal hypertensive disease, there were 124 NNDs. The overall birthweight for gestational age in the hypertensive group was substantially less than that of the whole population (mean MoM 0.84; 95% CI 0.83-0.85) and that of the 124 NNDs was still lower (mean MoM 0.75; 95% CI 0.724-0.782). For both hypertensive and non hypertensive cases, inspection of the data categorised into deciles showed that there was a continuous increase in the frequency of NND throughout the weight range, being lowest for the heaviest babies and highest for those in the lower centiles. CONCLUSION: (1) There is a relationship between birthweight for gestational age and risk of NND in infants born at 24-32 weeks; (2) this relationship is a continuum throughout the whole range of birthweight, not focused exclusively on a group defined as SGA; (3) provided appropriate birthweight standards are used, there is no extra effect on mortality from maternal hypertensive disease. PMID- 11267733 TI - Nitrofurantoin and congenital abnormalities. AB - OBJECTIVE: To study human teratogenic potential of oral nitrofurantoin treatment during pregnancy. MATERIALS AND METHODS: Pair analysis of cases with congenital abnormalities and matched population controls in the population-based dataset of the Hungarian Case-Control Surveillance of Congenital Abnormalities, 1980-1996. RESULTS: Of 38,151 pregnant women who had newborn infants without any congenital abnormalities (population control group), 774 (3.4%); of 22,865 case pregnant women who had newborns or fetuses with congenital abnormalities, 1079 (2.8%) and of 812 pregnant women who had newborns or fetuses with Down's syndrome (patient controls), 23 (2.8%) pregnant women were treated with nitrofurantoin. The above differences between population controls and cases may be connected with recall bias, because the case-control pair analysis did not indicate a teratogenic potential of nitrofurantoin use during the second and the third months of gestation, i.e. in the critical period for major congenital abnormalities. CONCLUSION: Treatment with nitrofurantoin during pregnancy does not present detectable teratogenic risk to the fetus. PMID- 11267734 TI - Perception of teratogenic risk of common medicines. AB - OBJECTIVE: To assess the perception of the teratogenic risk of common medication by professionals and lay people. STUDY DESIGN: A visual-analogue scale was used to measure the perceived percentage of mothers who will deliver a child with a malformation, including those exposed to a list of drugs. Fifteen general practitioners, 10 gynaecologists, 106 pre-clinical students, 150 students in their clinical training, 81 pregnant women and 63 non-pregnant women were interviewed. RESULTS: The perception of the teratogenic risk related to medication used in pregnancy was higher than the recognised risk in all groups, and for all drugs. The risk associated with safe medications was perceived to be higher by non-pregnant women as compared with the pregnant women. Pregnant women perceived the medication associated risk to be higher than physicians did for all drugs included in the questionnaire. CONCLUSIONS: The high and unrealistic perception of teratogenic risk amongst women and health professionals may lead to abortions of otherwise wanted and healthy children. PMID- 11267735 TI - Dietary factors and risk of spontaneous abortion. AB - OBJECTIVE: This study examines the association between dietary habits and risk of spontaneous abortion. DESIGN: Hospital-based case-control study. SETTING: Obstetric hospitals in Milan, Italy. SUBJECTS: Cases were: 912 women admitted for spontaneous abortion (within the 12th week of gestation). Controls were: women who gave birth at term to healthy infants on randomly selected days at the same hospitals where cases had been identified. RESULTS: The risk of spontaneous abortion was inversely and significantly related to green vegetables, fruit, milk, cheese, eggs and fish consumption. The multivariate odds ratios (OR), for highest versus lowest levels of intake, were 0.3 for fruit, 0.5 for cheese, 0.6 for green vegetables and milk and 0.7 for fish and eggs. The major type of seasoning fats have showed a direct association with risk of miscarriage. Comparing the highest with the lowest intake, the ORs were 2.0 (95% confidence interval, CI 1.1-3.6) and 1.6 (95% CI 1.1-2.3) for butter and oil, respectively. No consistent association emerged between meat, liver, ham and carrots intake and the risk of spontaneous abortion. CONCLUSIONS: This result suggests that a diet poor in several aspects, including vegetables and fruit, milk and dairy products, but rich in fats, may be a determinant or a correlate of increased risk of spontaneous abortion. PMID- 11267736 TI - Specialist life - William McBride. PMID- 11267737 TI - Knowledge representation and qualitative simulation of salmon redd functioning. Part I: qualitative modeling and simulation. AB - This work aims at representing empirical knowledge of freshwater ecologists on the functioning of salmon redds (spawning areas of salmon) and its impact on mortality of early stages. For this, we use Qsim, a qualitative simulator. In this first part, we provide unfamiliar readers with the underlying qualitative differential equation (QDE) ontology of Qsim: representing quantities, qualitative variables, qualitative constraints, QDE structure. Based on a very simple example taken of the salmon redd application, we show how informal biological knowledge may be represented and simulated using an approach that was first intended to analyze qualitatively ordinary differential equations systems. A companion paper (Part II) gives the full description and simulation of the salmon redd qualitative model. This work was part of a project aimed at assessing the impact of the environment on salmon populations dynamics by the use of models of processes acting at different levels: catchment, river, and redds. Only the latter level is dealt with in this paper. PMID- 11267738 TI - Knowledge representation and qualitative simulation of salmon redd functioning. Part II: qualitative model of redds. AB - This paper describes a qualitative model of the functioning of salmon redds (spawning areas of salmon) and its impact on mortality rates of early stages. For this, we use Qsim, a qualitative simulator, which appeared adequate for representing available qualitative knowledge of freshwater ecology experts (see Part I of this paper). Since the number of relevant variables was relatively large, it appeared necessary to decompose the model into two parts, corresponding to processes occurring at separate time-scales. A qualitative clock allows us to submit the simulation of salmon developmental stages to the calculation of accumulated daily temperatures (degree-days), according to the clock ticks and a water temperature regime set by the user. Therefore, this introduces some way of real-time dating and duration in a purely qualitative model. Simulating both sub models, either separately or by means of alternate transitions, allows us to generate the evolutions of variables of interest, such as the mortality rates according to two factors (flow of oxygenated water and plugging of gravel interstices near the bed surface), under various scenarios. PMID- 11267739 TI - Biocomplexity: adaptive behavior in complex stochastic dynamical systems. AB - Existing methods of complexity research are capable of describing certain specifics of bio systems over a given narrow range of parameters but often they cannot account for the initial emergence of complex biological systems, their evolution, state changes and sometimes-abrupt state transitions. Chaos tools have the potential of reaching to the essential driving mechanisms that organize matter into living substances. Our basic thesis is that while established chaos tools are useful in describing complexity in physical systems, they lack the power of grasping the essence of the complexity of life. This thesis illustrates sensory perception of vertebrates and the operation of the vertebrate brain. The study of complexity, at the level of biological systems, cannot be completed by the analytical tools, which have been developed for non-living systems. We propose a new approach to chaos research that has the potential of characterizing biological complexity. Our study is biologically motivated and solidly based in the biodynamics of higher brain function. Our biocomplexity model has the following features, (1) it is high-dimensional, but the dimensionality is not rigid, rather it changes dynamically; (2) it is not autonomous and continuously interacts and communicates with individual environments that are selected by the model from the infinitely complex world; (3) as a result, it is adaptive and modifies its internal organization in response to environmental factors by changing them to meet its own goals; (4) it is a distributed object that evolves both in space and time towards goals that is continually re-shaping in the light of cumulative experience stored in memory; (5) it is driven and stabilized by noise of internal origin through self-organizing dynamics. The resulting theory of stochastic dynamical systems is a mathematical field at the interface of dynamical system theory and stochastic differential equations. This paper outlines several possible avenues to analyze these systems. Of special interest are input-induced and noise-generated, or spontaneous state-transitions and related stability issues. PMID- 11267740 TI - Optically programming DNA computing in microflow reactors. AB - The programmability and the integration of biochemical processing protocols are addressed for DNA computing using photochemical and microsystem techniques. A magnetically switchable selective transfer module (STM) is presented which implements the basic sequence-specific DNA filtering operation under constant flow. Secondly, a single steady flow system of STMs is presented which solves an arbitrary instance of the maximal clique problem of given maximum size N. Values of N up to about 100 should be achievable with current lithographic techniques. The specific problem is encoded in an initial labeling pattern of each module with one of 2N DNA oligonucleotides, identical for all instances of maximal clique. Thirdly, a method for optically programming the DNA labeling process via photochemical lithography is proposed, allowing different problem instances to be specified. No hydrodynamic switching of flows is required during operation -- the STMs are synchronously clocked by an external magnet. An experimental implementation of this architecture is under construction and will be reported elsewhere. PMID- 11267741 TI - Comparison of field and laboratory methods for monitoring metallic mercury vapor in indoor air. AB - Real-time metallic mercury vapor levels of the indoor air were monitored at several mercury spill sites around the US in order to evaluate the effectiveness of site cleanup operations. Mercury readings taken in the field with a Jerome 431 Mercury Vapor Analyzer were compared with laboratory analysis using a modified National Institute for Occupational Safety and Health (NIOSH) 6009 method. Statistical evaluation showed that the data were highly comparable except at low concentrations, due to the large degree of uncertainty associated with measuring low levels of mercury with the Jerome analyzer. Regression analysis indicated that Jerome measurements of 10 microg/m(3) or greater are comparable for field analysis of mercury vapor in air. PMID- 11267742 TI - Application of a fiber-optic NIR-EFA sensor system for in situ monitoring of aromatic hydrocarbons in contaminated groundwater. AB - Interaction of analyte molecules with the evanescent wave of light guided in optical fibers is among the most promising novel sensing schemes that can be applied for environmental monitoring and on-line process analysis. By combining this measuring principle with the solid-phase extraction of analyte molecules into the polymer cladding of a fiber, it is possible to perform direct absorption measurements in the cladding, if the fiber is adapted to a conventional spectrometer/photometer. A big advantage of this arrangement is that the measurement is scarcely disturbed by matrix effects (background absorption of water in IR measurements, stray light due to turbidity in the sample). By using near-infrared (NIR) evanescent field absorption (EFA) measurements in quartz glass fibers coated with a hydrophobic silicone membrane it is possible to design and construct sensors for monitoring apolar hydrocarbons (HCs) in aqueous matrices.The paper presents a fiber-optic sensor system for the determination of aromatic HCs in groundwater or industrial wastewater. Generally, this instrument is suitable for quantitative in situ monitoring of pollutants such as aromatic solvents, fuels, mineral oils or chlorinated HCs with relatively low water saturation solubility (typically between 0.01 and 10 g l(-1)). The sensor probe is connected via all-silica fibers to a filter photometer developed at the IFIA, thus, allowing even remote analysis in a monitoring well. This portable instrument provides a total concentration signal of the organic compounds extracted into the fiber cladding by measuring the integral absorption at the 1st C--H overtone bands in the NIR spectral range. In situ measurements with the sensor system were performed in a groundwater circulation well at the VEGAS research facility of the University of Stuttgart (Germany). The NIR-EFA sensor system was tested within the frame of an experiment that was carried through in a tank containing sandy gravel with a groundwater-saturated aquifer, where soil and groundwater were contaminated with technical grade xylene. The goal of this experiment was to model and optimize the groundwater circulation well used for the remediation of the aquifer and soil surrounding the well. The sensor proved to trace reliably the total hydrocarbon concentration in the process water pumped from the well to a stripper column. Measurements were performed continuously over 4 months with C8 HC sum concentrations in the process water between 80 mg l(-1) down to the limit of detection, which is around 200 microg l(-1). It could be demonstrated that the fiber-optic sensor system is a valuable tool for near-real time control of a remedial action technique and verification and documentation of its success. PMID- 11267743 TI - Field screening test methods: performance criteria and performance characteristics. AB - Field-portable test methods may be quantitative, semi-quantitative, or qualitative and screening methods are often used in the field to determine if the concentration of a toxic substance exceeds regulatory or recommended standards or action levels. For on-site analysis, accurate quantitative tests for field measurements may not be available, depending on the analyte(s) or specific field situation. Thus, in lieu of more definitive test methods, screening tests which are based on qualitative or semi-quantitative methods are often used for making immediate decisions in the field, e.g. for compliance or risk assessment. Also, quantitative methods may be used for screening purposes in many instances. To ensure the quality of these screening tests and the decisions that are made based upon their results, screening methods need to be evaluated with sufficient data and should meet basic performance criteria prior to their being employed for decision-making purposes. Although quantitative, semi-quantitative and qualitative methods demonstrate different characteristics, it is desired that the performance criteria for all three method categories be consistent. If there is consistency, then one can have a sound basis for selecting the most appropriate test(s) for a given application. In order to unify the performance criteria for the different types of methods, a performance function is used to characterise both qualitative and semi-quantitative methods; in turn, this performance function is related to that for quantitative methods. False negative rates, false positive rates, sensitivity and specificity are key characteristics of screening methods that can be determined from the pertinent performance curves. The performance characteristics of each method are related to the uncertainty region that is associated with each method and the applicable uncertainty regions can be gleaned from the performance curves. Also, various options for using multiple test results to improve decisions based on test results are provided. PMID- 11267744 TI - Ultrasonic extraction and field-portable anodic stripping voltammetric measurement of lead in dust wipe samples. AB - Dust wipe samples were subjected to ultrasonic extraction (UE) in diluted nitric acid, and then analyzed for lead content using field-portable anodic stripping voltammetry (ASV). Recoveries of lead were determined from wipe materials which were spiked with certified reference materials (CRMs) containing known quantities of lead. Four different wipe materials and four different CRMs were tested, with and without filtration of aliquots of sample extract through 0.45 microm hydrophilic polytetrafluoroethylene filters. The CRMs consisted of paint, soil, particulate, and dust matrices. Wipe materials were chosen from those which have been found to meet the performance aspects of an ASTM standard specification. UE/ASV experiments were carried out in accordance with newly published ASTM procedures for on-site extraction and electroanalysis. Recoveries were found to vary for different wipe materials and CRMs. For several CRMs, quantitative (80- 120%) recoveries for UE/ASV were observed for one wipe material whether filtration was used or not, while other wipe materials required filtration for quantitative recovery. In the case of one wipe material which contained detergents, quantitative recoveries could not be achieved whether filtration was used or not. The total analysis time for a sample set of 6--12 samples was 60--90 min, including extraction time and sample manipulation. The results of this work have provided information on the choice of wipe materials that can be used for quantitative lead measurements by UE/ASV in materials that are representative of sources of lead in surface dust. PMID- 11267745 TI - Explosives detection in soil using a field-portable continuous flow immunosensor. AB - A field method for quantitative analysis of explosives in contaminated soil samples is described. The method is based on a displacement immunoassay performed in a commercial instrument, the FAST 2000, engineered by Research International Inc. The method can be used on-site to measure 2,4,6-trinitrotoluene (TNT) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) within 5min. For this study, replicate analyses were performed on soil extracts prepared from each field sample as well as appropriate controls, blanks, and laboratory standards. Statistical analyses were done to assess accuracy, bias, and predictability of the method. The results demonstrated that the immunosensor could be used effectively to screen environmental samples for the presence or absence of explosives. In most samples, the method also provided quantitative values that were in good agreement with standard laboratory analyses using HPLC. A limited number of sample matrices interfered with the immunoassay and produced results that varied significantly from the laboratory data. In each case, the compounds causing the problem have been identified and efforts are being made to minimize these matrix interferences in future field evaluations. PMID- 11267746 TI - An evaluation of field total petroleum hydrocarbon (TPH) systems. AB - An evaluation of several field kits and petroleum hydrocarbon measuring systems was conducted. The field kits were the immunoassay based EnviroGard petroleum fuels in soil test kit (EnviroGard, Millipore Canada, Mississauga, Ont., Canada), the turbidimetric based PetroFlag hydrocarbon test kit for soil (Dexsil, Hamden, CT, USA), a DR/2000 field kit (Hach Company, Loveland CO, USA) employing colorimetric test procedures and a total organic carbon (TOC) analysis instrument (Dohrmann Division, Rosemount Analytical Inc., Santa Clara, CA, USA) using oxidation principles. These procedures were compared to the traditional technique of extraction of the petroleum hydrocarbons using trichlorotrifluoroethane (Freon 113) as the solvent and subsequent infrared (IR) analysis using a portable fixed wavelength analyzer (Buck Scientific, East Norwalk, CT, USA). The EnviroGard kit was affected by the sample matrix. The soil type and the presence or lack thereof specific chemical components affected the capability to detect the petroleum hydrocarbon concentration. The PetroFlag soil test kit tended to generate results higher than the accepted concentration. The IR method was better capable of producing results similar to the expected concentration values of the prepared samples. Results indicate that the total organic carbon analysis technique evaluated is best suited for samples containing dissolved hydrocarbons in water and is not a preferred procedure for water samples containing dispersed or floating oil. At low concentrations of 10ppm and less, the TOC method and IR method have concentration values within a few parts-per-million (ppm) of each other, however, an examination of the trends in the results for all samples shows no similarity. This would indicate that the traditional extraction and infrared method and the total organic carbon method are not measuring the same parameter.Finally, the colorimetric field kit was capable of quantifying the concentration of oil in water samples within limits. The results from the oil-in water method built into the unit at the factory were not comparable with analysis carried out by the infrared technique. With specific methods for each oil incorporated into the spectrophotometer, the comparability of data increased significantly. Results generated by the kit are dependent upon the color and amount of the oil in the sample. The kit is best suited for dark colored oils and the water samples with concentrations in the range of 10 to 85ppm by weight. PMID- 11267747 TI - Solvent vapour monitoring in work space by solid phase micro extraction. AB - Solid phase micro extraction (SPME) is a fast, solvent-less alternative to conventional charcoal tube sampling/carbon disulfide extraction for volatile organic compounds (VOC). In this work, SPME was compared to the active sampling technique in a typical lab atmosphere. Two different types of fibre coatings were evaluated for solvent vapour at ambient concentration. A general purpose 100 microm film polydimethylsiloxane (PDMS) fibre was found to be unsuitable for VOC work, despite the thick coating. The mixed-phase carboxen/PDMS fibre was found to be suitable. Sensitivity of the SPME was far greater than charcoal sorbent tube method. Calibration studies using typical solvent such as dichloromethane (DCM), benzene (B) and toluene (T) showed an optimal exposure time of 5 min, with a repeatability of less than 20% for a broad spectrum of organic vapour. Minimum detectable amount for DCM is in the range of 0.01 microg/l (0.003 ppmv). Variation among different fibres was generally within 30% at a vapour concentration of 1 microg DCM/l, which was more than adequate for field monitoring purpose. Adsorption characteristics and calibration procedures were studied. An actual application of SPME was carried out to measure background level of solvent vapour at a bench where DCM was used extensively. Agreement between the SPME and the charcoal sampling method was generally within a factor of two. No DCM concentration was found to be above the regulatory limit of 50 ppmv. PMID- 11267748 TI - Field portable XRF analysis of environmental samples. AB - One of the critical factors for successfully conducting contamination characterization, removal, and remedial operations at hazardous waste sites is rapid and appropriate response to analyze samples in a timely fashion. Turnaround time associated with off-site analysis is often too slow to support efficient utilization of the data. Field portable X-ray fluorescence (FPXRF) techniques provide viable and effective analytical approaches to meet on-site analysis needs for many types of environmental samples. Applications include the in situ analysis of metals in soils and sediments, thin films/particulates, and lead in paint. PMID- 11267749 TI - On site PCB analysis in support of a transformer rebuilding project. AB - In December 1997, Emergencies Science Division (ESD) was contracted by Natural Resources Canada (NRCAN) to perform on-site analyses in support of a transformer rebuilding project at Sault Ste-Marie, Ont. Using a gas chromatograph with electron capture detector (GC/ECD) mounted in a mobile laboratory, PCB analyses were conducted on the original transformer oil, surface wipes, Varsol rinsing of the transformer tank interior and cooling fins. To assess the efficiency and validity of the decontamination process, PCB contamination was monitored closely on the rinse solvent. Surface wipe samples after wash down showed surface concentration of several hundred microg Aroclor 1254/100 cm(2), well below the acceptable limit of 8000 microg/100 cm(2). Because of the relatively large percentage of the internal surface area, the fin banks had to be rinsed exhaustively to meet the decontamination criteria. Final rinses of each of the seven fin banks of transformer 1 still showed presence of PCB, ranging from 80 to 590 ppm (microg/ml) with a mean value of 280 ppm. Upon completion of rebuilding, analysis of the R-Temp retro fill fluid showed 5 ppm at the initial power-up, increasing slightly to 16 ppm after 1 year of operation, which was far below the regulatory limit 50 ppm. The second transformer, by comparison, had a lower mean concentration of 54 ppm in the final fin rinse during decontamination. However, the backfill R-Temp showed an initial concentration of 38 ppm and remained essentially unchanged at 32 ppm after approximately 10 months of operation. Extensive comparison of GC and the quick test Clor-N-Oil kit were also carried out and showed generally good agreement. The use of an on-site GC was crucial in providing rapid and accurate analysis on-site, thus, enabling quick modifications to the decontamination strategies in order to meet the target PCB level. For projects of this nature, a GC/ECD was far superior to quick test kits by providing the selectivity and sensitivity for the diverse nature of the sample media. PMID- 11267750 TI - Use of a portable infra-red analyzer for low-level hydrocarbon emissions. AB - During some on-site tests, a portable infra-red (IR) analyzer was used successfully to monitor for hydrocarbon vapors. The detection limit of the IR analyzer is much lower than that of most other hydrocarbon vapor monitors and can be used in situations where, as in most ambient air monitoring situations, the levels are often less than a milligram per cubic metre (mg/m(3)). Traditional procedures used to measure hydrocarbon concentrations at lower levels involves the collection of samples on-site, which are then transported to a laboratory for analysis. The advantage of providing continuous sampling data is that it may indicate trends in the hydrocarbon vapor emissions that may not be apparent using a grab-type sample. The initial tests were designed to determine if the IR analyzer was capable of monitoring the low-level hydrocarbons in a field situation. The findings from that initial work was followed by modification of the test procedure to include an upwind IR analyzer, shortened sampling cycles to produce more data, and additional canister samples collected outside the burn period. The metered grab samples, using Summa canisters, were collected over a 1h period and any results would therefore, reflect an average value over the hour. The IR analyzer, with a sampling cycle of approximately 1min, was able to produce a near real-time distribution of the hydrocarbon vapors in the test site emissions. Because the testing parameters and methods are quite different, it is difficult to compare these two methods, but indications suggest strongly that the use of this portable IR instrument could help to describe the hydrocarbon emissions downwind from a source, as well as to monitor for these hydrocarbons continuously, including situations where the levels are below detection limits of most portable detectors. PMID- 11267751 TI - Preface. PMID- 11267752 TI - Screening of the anticoccidial effects of herb extracts against Eimeria tenella. AB - Ionophorous antibiotics have been popularly used in the treatment of avian coccidiosis. Tissue residue of these antibiotics may be found in poultry, we have sought safe alternative anticoccidial herbal materials for the control of avian coccidiosis. Efficacy of extracts from 15 different herbs, including Bupleurum chinese DC, Sophora flavescens Aiton, and Artemisia annua Linne was tested against Eimeria tenella. One-day-old broiler chicks were infected with a USDA reference book of E. tenella, and administered various herbal extracts. Survival rates, lesion scores, body weight gains, bloody diarrhea, and oocysts excretions were investigated at the first and the second week after infection. Bloody diarrhea in the S. flavescens and Sinomenium acutum treated groups was milder than that of the other infected groups. Survival rates in the groups treated with Ulmus macrocarpa (100%), Pulsatilla koreana, Torilis japonica, Artemisia asiatica and S. flavescens (90%) were higher than that of the infected control group (70%). Lesion scores in the groups treated with U. macrocarpa (1.40+/-1.14) or Pulsatilla koreana (1.60+/-1.82) were significantly lower than those of the infected control group (3.00+/-1.10). During the first week after infection, the weight gains in the groups treated with Quisqualis indica (232.9+/-43.5 g), S. flavescens (214.4+/-46.1 g) and S. acutum (211.3+/-29.4 g) were significantly higher than the infected control group (172.4+/-17.6 g). In conclusion, the data of the survival rates, bloody diarrhea symptoms, lesion scores, body weight gains and oocyst excretions indicate that the extract of S. flavescens was the most effective. P. koreana, S. acutum, U. macrocarpa and Q. indica were also effective. Further research on the above herbal materials will be carried out by the authors by chemical analysis of the extracts. PMID- 11267754 TI - Mitosis and differentiation in T-cells under cytotoxic action of Echinococcus granulosus hydatid fluid. AB - In the T-cell line, D10, thymidine uptake was used to measure the proportion of cells in S-phase, and the MTT assay to measure the number of viable cells. The effect of Echinococcus granulosus hydatid fluid (HF) on the lymphocytes was assayed in 3-day cultures of the T-cell line, D10, in increasing concentrations of HF. Apparent cytotoxic effects of HF were recorded as a log-linear decline in S-phase activity, which was reduced by the presence of IL-1, IL-2, or a combination of the two. In the presence of IL-2, however, mitogenic treatment with concanavalin A increased the cytotoxic effect in 3-day cultures, while in day-2 cultures, HF itself showed mitogenic effect. HF-induced decline in S-phase activity was not matched by a parallel decline in viable cells, suggesting that the apparent cytotoxicity of HF could result from cell-cycle arrest. Depending on its origin, HF enhanced membrane expression of CD25 and CD38 on human peripheral blood lymphoblasts, and diminished that of CD28. Taken together, these changes suggest that HF can induce T-cell mitosis and reduce co-stimulation with subsequent T-cell anergy or apoptosis. PMID- 11267755 TI - Survival of infective Ostertagia ostertagi larvae on pasture plots under different simulated grazing conditions. AB - This study was carried out to examine the survival of infective Ostertagia ostertagi larvae (L(3)) on pasture under different simulated conditions of grazing, i.e. mixed grazing of cattle and nose-ringed sows, or grazing by cattle alone. Standardised pats of cattle faeces containing O. ostertagi eggs were deposited on three types of herbage plots, which were divided into zone 1: faecal pat; zone 2: a circle extending 25cm from the edge of the faecal pat; zone 3: a circle extending 25cm from the edge of zone 2. For "tall herbage" (TH) plots, the herbage in zone 2 was allowed to grow naturally, while the herbage in zone 3 was cut down to 5-7cm fortnightly, imitating a cattle-only pasture. For "short herbage" (SH) plots, the herbage in both zones 2 and 3 were cut down to 5-7cm fortnightly, imitating mixed grazing of cattle and sows. The grass in the "short herbage and scattered faeces" (SH/SF) plots were cut as for SH plots, and the faeces were broken down 3 weeks after deposition and scattered within zone 2, imitating the rooting behaviour of co-grazing sows. Five faecal pats from each plot group were collected on monthly basis, along with the herbage from zones 2 and 3 cut down to the ground. Infective larvae were then recovered from both faeces and herbage. The numbers of L(3) recovered from zone 1 were higher in the TH plots than in the other two groups and, furthermore, the larval counts from SH plots were always higher than from SH/SF plots. The three groups followed a similar pattern during the season regarding numbers of L(3) in zone 2, and no clear patterns between plot types were obtained. The presence of L(3) in zone 3 was almost negligible. Important differences were seen throughout the study from the biological point of view; more L(3) were able to survive in faeces on the TH plots, presumably reflecting a better protection from heat and desiccation compared to those in the other plots. The overall results support the idea that mixed grazing of cattle and pigs favour the reduction of O. ostertagi larval levels in pasture. This reduction is mainly due to the grazing behaviour of pigs, which by grazing up to the very edge of the cattle faeces, will either expose the larvae in faeces to adverse environmental summer conditions or ingest cattle parasite larvae, or both. PMID- 11267756 TI - Effect and repeatability of Ascaridia galli egg output in cockerels following a single low dose infection. AB - Three groups of caged 20 Tetra-SL cockerels aged 1 day were orally infected with 30, 60 or 125 embryonated Ascaridia galli eggs. After 11, 12 and 13 weeks, faecal egg counts (FECs) were determined. All birds were slaughtered after the last sampling. A group of 25 control birds was sampled and slaughtered in parallel. The gastrointestinal tracts were examined for the presence of adult stages of A. galli. A random sample of 10% was also examined for the presence of immature stages of A. galli. The group with an infection dose of 125 eggs showed the highest average worm burden (p<0.05) and number of females (p<0.05), but the mean establishment rate was the lowest in this group. There was no significant difference in the mean logFEC between the groups. The logFEC per female worm was the highest in the low infection group (p<0.05). The average worm length and weight and the birds body weight were not significantly different among the groups. The estimated repeatabilities for mean logFEC of the different samples were reasonably high (0.55-0.87). This may open a way of genetic selection for A. galli resistance in chickens, which will be of importance for birds kept in alternative and organic farming systems. PMID- 11267753 TI - Leishmania infantum-specific IgG, IgG1 and IgG2 antibody responses in healthy and ill dogs from endemic areas. Evolution in the course of infection and after treatment. AB - The expression of IgG, IgG1 and IgG2 specific antibodies for Leishmania infantum was studied in five groups of dogs in Catalonia (Spain): I, 99 asymptomatic dogs (infected and uninfected) from a highly endemic area for leishmaniosis; II, 139 untreated dogs with clinically patent leishmaniosis; III, 11 naturally infected asymptomatic dogs monitored for up to 5 years since they were found seropositive to Leishmania antigen and without treatment; IV, 25 naturally infected dogs with clinically patent leishmaniosis and treated with either meglumine antimoniate and allopurinol or allopurinol alone and V, six experimentally infected dogs, treated with meglumine antimoniate and controlled for 5 years. The levels (ELISA units) of IgG, IgG1 and IgG2 in asymptomatic dogs (group I) were very variable (24+/-33, 32+/-31 and 26+/-31, respectively), and, as expected, lower than in ill dogs (group II) (168+/-34, 84+/-71 and 172+/-31, respectively). In both groups, the correlation between IgG and IgG2 levels (r=0.95, P<0.001 in group I and r=0.63, P<0.001 in group II) was higher than between IgG and IgG1 levels (r=0.01, P>0.05 in group I and r=0.31, P<0.001 in group II). In group III, IgG and IgG2 expression increased during infection, while IgG1 expression remained the same. In dogs of group IV, IgG levels after 1 year of treatment decreased more in responsive (mean values, 163+/-42 before treatment (b.t.) and 100+/-36 after treatment (a.t.)) than in unresponsive dogs (158+/-29 b.t. and 124+/-51 a.t.), especially for IgG1 (94+/-89 b.t. and 20+/-21 a.t. in responsive dogs and 35+/-25 b.t. and 22+/-13 a.t. in unresponsive dogs) rather than for IgG2 (156+/-16 b.t. and 114+/-45 a.t. in responsive and 151+/-11 b.t. and 125+/-36 a.t. in unresponsive dogs). Similar results were observed in the evolution of experimentally infected animals after consecutive and specific treatments. Overall results show the great variation in Leishmania-specific IgG1 expression in asymptomatic and symptomatic dogs, their lack of correlation with that of IgG2 and chemotherapy is more effective in dogs with initially high expression of IgG1. PMID- 11267757 TI - Factors affecting the frequency of ear canal and face infestation by Otodectes cynotis in the cat. AB - Otodectes cynotis is responsible for at least 50% of canker cases diagnosed in cats world-wide. The role of Demodex cati in the pathogenesis of otitis and acne is still obscure. The aims of this study were to estimate the prevalence of O. cynoyis and D. cati infestations in clinically normal cats in northern Greece, to determine the factors that are associated with the probability and severity of infestation in the cat, and to examine the importance of these mites in the pathogenesis of feline acne. Samples from 161 cats were examined by flushing the ear canals and by taking skin scrapings of the chin and lip area. The results were combined with various factors (sex, age, living style, hair coat type and presence of pruritus, of ear discharge, of acne-like lesions) in order to carry out a risk analysis. Two separate logistic regression analyses were performed. One, on the infestation/non-infestation potential with O. cynotis and the other, on the degree of such infestation as mild-to-moderate (< or =5 mites/field) or severe (>5 mites/field). D. cati was not detected in any of the 161 cats. The prevalence of O. cynotis was estimated at 25.5% (95% confidence interval (CI) 19 32). The rate of mite infestation was higher with the presence of ear discharge (odds ratio 9, 95% CI 3.3-24.5), periaural pruritus (odds ratio 3.6, 95% CI 1.8 8) and acne-like lesions (odds ratio 3.3, 95% CI 1.2-9). Cats with mild-to moderate degree of infestation had 18 times higher chance of exhibiting an ear discharge than those with a severe infestation. The log-odds of mild-to-moderate parasitism were linearly related to the age. PMID- 11267758 TI - Comparison of heartworm antigen test kit performance in dogs having low heartworm burdens. AB - Sensitivity and specificity of four in-clinic heartworm antigen test kits, AbboScreen (Abbott Laboratories), Snap PF (IDEXX Laboratories), Solo Step (HESKA Corporation), Witness (Synbiotics Corporation) and two heartworm antigen microwell plate assays, DiroCHEK (Synbiotics) and PetChek PF (IDEXX) were compared in a blinded study using serum or plasma drawn from 237 random source dogs, including 140 with necropsy-confirmed, low worm burden infections (minimum 1 worm, maximum 10, mean 2.3, median 3) and 97 confirmed heartworm-free at necropsy. In general, microwell format tests were more sensitive than membrane format tests and tests using ELISA technology were more sensitive than tests using lateral flow immunochromatographic technology. Percent sensitivity and specificity, respectively, were PetChek PF 76 and 97, DiroCHEK 71 and 94, SNAP PF 67 and 98, Solo Step 60 and 98, and AbboScreen 52 and 96. The Witness test protocol was changed by the manufacturer midway through the study, and the newer version of this test kit arrived containing a package insert alerting the user to a change in procedure, which purportedly resulted in improved sensitivity. PetChek was significantly more sensitive than all other tests except DiroCHEK and the new version of Witness. DiroCHEK was significantly more sensitive than all tests except PetCheck, SNAP and the new version of Witness. Snap was more sensitive than AbboScreen and the old version of Witness. Differences in specificity were not significant (P>0.05). PMID- 11267759 TI - Persistent efficacy of doramectin and moxidectin against Cooperia oncophora infections in cattle. AB - Duplicate studies in France and Northern Ireland were carried out to determine the persistent efficacy of topical moxidectin and doramectin against natural infections of Cooperia oncophora. In each study, groups of 15 nematode-naive calves were treated either with topical doramectin or moxidectin, and put out to graze on pasture contaminated with C. oncophora infective larvae. The persistent efficacy for preventing establishment of infection was assessed by the time to faecal egg excretion of C. oncophora eggs. It was found that the moxidectin treatment prevented infection for less than 10 days and the effect of doramectin lasted for 24 days. PMID- 11267761 TI - Isolation and phenotypic characterization of Pseudomonas aeruginosa pseudorevertants containing suppressors of the catabolite repression control defective crc-10 allele. AB - The amiE gene encodes an aliphatic amidase capable of converting fluoroacetamide to the toxic compound fluoroacetate and is one of many genes whose expression is subject to catabolite repression control in Pseudomonas aeruginosa. The protein product of the crc gene, Crc, is required for repression of amiE and most other genes subject to catabolite repression control in this bacterium. When grown in a carbon source such as succinate, wild-type P. aeruginosa is insensitive to fluoroacetamide (due to repression of amiE expression). In contrast, mutants harboring the crc-10 null allele cannot grow in the presence of fluoroacetamide (due to lack of repression of amiE). Selection for succinate-dependent, fluoroacetamide-resistant derivatives of the crc-10 mutant yielded three independent pseudorevertants containing suppressors that restored a degree of catabolite repression control. Synthesis of Crc protein was not reestablished in these pseudorevertants. All three suppressors of crc-10 were extragenic, and all three also suppressed a Delta crc::tetA allele. In each of the three pseudorevertants, catabolite repression control of amidase expression was restored. Catabolite repression control of mannitol dehydrogenase production was also restored in two of the three isolates. None of the suppressors restored repression of glucose-6-phosphate dehydrogenase or pyocyanin production. PMID- 11267762 TI - Characterization of the alanine racemases from two mycobacteria. AB - D-Alanine is a necessary precursor in the biosynthesis of the bacterial peptidoglycan. The naturally occurring L-alanine isomer is racemized to its D form through the action of a class of enzymes called alanine racemases. These enzymes are ubiquitous among prokaryotes, and with very few exceptions are absent in eukaryotes, making them a logical target for the development of novel antibiotics. The alanine racemase gene from both Mycobacterium tuberculosis and M. avium was amplified by PCR and cloned in Escherichia coli. Overexpression of the proteins in the E. coli BL21 system, both as native and as His-tagged recombinant products, has been achieved. The proteins have been purified to electrophoretic homogeneity and analyzed biochemically. A D-alanine requiring double knock-out mutant of E. coli (alr, dadX) was constructed and the cloned genes were able to complement its deficiencies. PMID- 11267763 TI - Monodansylcadaverine inhibits cytotoxicity of Vibrio parahaemolyticus thermostable direct hemolysin on cultured rat embryonic fibroblast cells. AB - The mechanism of action of Vibrio parahaemolyticus thermostable direct hemolysin (TDH) on cultured cells still remains unclear. We show that addition of osmotic stabilizers, such as polyethylene glycol and dextran, could not protect cultured rat embryonic fibroblast cells (Rat-1) against cytotoxicity induced by TDH, unlike their protection against the hemolytic activity of TDH. By contrast, 100 microM monodansylcadaverine, as well as the presence of 1 mM ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) in medium, protected the cells against cytotoxicity of TDH. Binding of TDH to Rat-1 cells and intracellular localization of TDH were affected by monodansylcadaverine and EGTA as analyzed by flow cytometry and confocal microscopy. On the hemolytic activity of TDH, monodansylcadaverine and EGTA had no effect. These results suggest that the mechanism of cytotoxicity of TDH on Rat-1 cells was different from that of hemolytic activity of TDH on red blood cells. PMID- 11267764 TI - Pleiotropic mutants hypersensitive to heavy metals and to oxidative stress in Chlamydomonas reinhardtii. AB - Insertional mutagenesis was used in Chlamydomonas reinhardtii to isolate original mutants hypersensitive to multiple drugs and physical agents. Out of 5200 transformants analyzed, 13 mutants belonging to seven phenotypic classes were isolated. Five were exclusively sensitive to cadmium and represented two loci. The other mutants were pleiotropic and presented a cross sensitivity to several (2--6) of the following agents: cadmium, copper, lead, paraquat, hydrogen peroxide, UVC and light. In all mutants analyzed, the hypersensitive phenotype was most probably due to a single mutational event. The sensitivity of several pleiotropic mutants to a broad range of physical and chemical agents suggests that the disrupted genes are involved in multiple stress responses. PMID- 11267765 TI - Purification and characterisation of a possible assimilatory nitrite reductase from the halophile archaeon Haloferax mediterranei. AB - The nitrite reductase from the extreme halophilic archaeon, Haloferax mediterranei, has been purified and characterised. H. mediterranei is capable of growing in a minimal medium (inorganic salts and glucose as a carbon source) with nitrate as the only nitrogen source. The overall purification was 46-fold with about 4% recovery of activity. The enzyme is a monomeric protein of approximately 66 kDa. A pH of 7.5 and high temperatures up to 60 degrees C are necessary for optimum activity. Reduced methyl viologen has been found to be an electron donor as effective as ferredoxin. NADPH and NADH, which are electron donors in nitrite reductases from different non-photosynthetic bacteria, were not effective with nitrite reductase from H. mediterranei. PMID- 11267766 TI - Identification and molecular characterization of a novel Bacillus strain capable of degrading Tween-80. AB - A Tween-80-degrading novel marine Bacillus strain, N10, has recently been isolated in Alexandria University, Egypt. The taxonomic position of this endospore forming bacterium was investigated on the basis of fatty acid analysis and 16S rRNA gene sequencing. Comparative computer database analyses revealed that the bacterium is a Bacillus subtilis strain. The gene encoding the small acid-soluble protein gamma-type (SASP-B), sspE, was successfully utilized in this study as a tool for discrimination between the two B. subtilis subspecies W23 and 168. Based on the alignment of 16S rRNA sequences and analysis of SASP-B relatedness, it has been demonstrated that the novel marine B. subtilis strain N10 is more closely related to the B. subtilis reference strain W23 than to 168. The strain, N10, has been deposited in the Bacillus Genetic Stock Center (BGSC) and assigned the accession number 3A17. PMID- 11267767 TI - Identification of conditionally expressed genes in Streptococcus pyogenes using RNA fingerprinting. AB - RNA fingerprinting using arbitrarily primed reverse transcription-polymerase chain reaction was employed on isolated RNA from Streptococcus pyogenes bacteria in order to identify genes that were regulated in response to environmental changes. When S. pyogenes was cultured under glucose-rich growth conditions a number of transcriptionally up-regulated products were identified, cloned and sequenced. Using the Streptococcal Genome Sequencing Project database and similarity searches against the GenBank database the corresponding genes encoding enzyme IIB and IIC component of a putative phosphotransferase system were identified. Thus, we show that RNA fingerprinting could be a useful tool to identify unknown genes in S. pyogenes that are expressed under certain environmental conditions. PMID- 11267769 TI - Inhibition of bacterial RNA polymerase by the cyanobacterial metabolites 12-epi hapalindole E isonitrile and calothrixin A. AB - The alkaloid 12-epi-hapalindole E isonitrile, from a cyanobacterial Fischerella species, and the indolophenanthridine calothrixin A, from Calothrix, inhibited Escherichia coli RNA polymerase competitively with respect to ATP, and non competitively with respect to UTP. The inhibition was dependent on the order of addition of the inhibitors. The K(I) values, with ATP as the variable substrate, were 1.3+/-0.2 mM and 0.23+/-0.11 mM, respectively. Based on comparisons with the sensitivity of whole cells to these inhibitors, it is concluded that other targets in addition to RNA polymerase may also be implicated in their action. PMID- 11267768 TI - Identification of archaeal rDNA from subgingival dental plaque by PCR amplification and sequence analysis. AB - A PCR assay for the amplification of small subunit ribosomal DNA (SSU rDNA) of Euryarchaea was developed and used to detect archaeal rDNA in 37 (77%) out of 48 pooled subgingival plaque samples from 48 patients suffering from periodontal disease. One major group of cloned periodontal sequences was identical to Methanobrevibacter oralis and a second minor group to Methanobrevibacter smithii. These two groups and a third novel group were found to be more than 98% similar to each other over an 0.65-kb segment of the 16S rRNA gene sequenced. M. oralis was found to be the predominant archaeon in the subgingival dental plaque. Phylogenetic analysis of partial SSU rDNA sequences revealed evidence for a distinct cluster for human and animal Methanobrevibacter sp. within the Methanobacteriaceae family. PMID- 11267770 TI - A simple (14)C-respirometric method for assessing microbial catabolic potential and contaminant bioavailability. AB - This paper describes the validation and application of a simple flask-based (14)C respirometer system designed to assess mineralisation of (14)C-labelled substrates under defined conditions. Validation of this respirometer system indicated stoichiometric CO(2) trapping up to a maximum of 400 micromol of CO(2) (in a single trap). Polycyclic aromatic hydrocarbon (PAH)-degrading bacteria were used to measure growth-linked biodegradation of [(14)C]naphthalene to (14)CO(2). A (14)C activity balance of 101.7+/-8.9% (n=6), after 74 h incubation time and 10 respirometer-opening events, indicated the suitability of the system for monitoring substrate mineralisation. This respirometric apparatus was then successfully applied to assess: (i) the PAH catabolism of microbes in a field contaminated soil, where naphthalene and phenanthrene were rapidly mineralised and (ii) soil-associated organic contaminant bioavailability, where increased soil-phenanthrene contact time resulted in a reduction in phenanthrene mineralisation in the soil. The described respirometer system differs from existing respirometer systems in that the CO(2) trap can be removed and replaced quickly and easily. The system is efficient, reproducible, adaptable to many situations, easy to construct and simple to use, it therefore affords advantages over existing systems. PMID- 11267771 TI - Enterobactin: the characteristic catecholate siderophore of Enterobacteriaceae is produced by Streptomyces species.(1). AB - Enterobactin is described in the literature as the typical iron-chelating compound (siderophore) produced by bacteria of the family Enterobacteriaceae. In the course of a HPLC with diode array detection screening programme for detection of novel secondary metabolites, enterobactin, its biosynthetic precursor 2,3 dihydroxy-N-benzoylserine and its linear dimer and trimer condensation products were found to be produced by two Streptomyces strains besides the trihydroxamate type siderophores desferri-ferrioxamine B and E. PMID- 11267773 TI - Identification of the intracellular polyhydroxyalkanoate depolymerase gene of Paracoccus denitrificans and some properties of the gene product. AB - Paracoccus denitrificans degraded poly(3-hydroxybutyrate) (PHB) in the cells under carbon source starvation. Intracellular poly(3-hydroxyalkanoate) (PHA) depolymerase gene (phaZ) was identified near the PHA synthase gene (phaC) of P. denitrificans. Cell extract of Escherichia coli carrying lacZ--phaZ fusion gene degraded protease-treated PHB granules. Reaction products were thought to be mainly D(--)-3-hydroxybutyrate (3HB) dimer and 3HB oligomer. Diisopropylfluorophosphonate and Triton X-100 exhibited an inhibitory effect on the degradation of PHB granules. When cell extract of the recombinant E. coli was used, Mg(2+) ion inhibited PHB degradation. However, the inhibitory effect by Mg(2+) ion was not observed using the cell extract of P. denitrificans. PMID- 11267772 TI - The Yersinia high-pathogenicity island is highly predominant in virulence associated phylogenetic groups of Escherichia coli. AB - The high-pathogenicity island (HPI) present in pathogenic Yersinia and encoding the siderophore yersiniabactin, has recently been identified in the asnT tRNA region of various Escherichia coli pathotypes, especially those responsible for bacteremia and urosepsis. Most E. coli strains causing such extra-intestinal infections belong to phylogenetic groups B2 and D. In this study we investigated (i) the distribution and localization of HPI among the different E. coli phylogenetic groups, using the ECOR reference collection; and (ii) the prevalence of HPI among a set of 124 phylogenetically characterized E. coli strains responsible for neonatal meningitis. Ninety-three percent of the ECOR strains belonging to groups B2 and D harbored HPI. In contrast, the island was present in 32% and 25% of strains belonging to groups A and B1, respectively, which are considered to be non-pathogenic. HPI was found in 100% of the neonatal meningitis strains, 13 of which belonged to groups A and B1, suggesting that HPI might contain virulent factors required for the development of neonatal meningitis. Moreover, we observed for the first time that HPI can be inserted in a site different from the asnT tRNA region. PMID- 11267774 TI - Taxon-specific oligonucleotide primers for detection of two ancient endomycorrhizal fungi, Glomus occultum and Glomus brasilianum. AB - A unique oligonucleotide pair, GOCC56:GOCC427, was designed that correctly primed specific amplification of a approximately 370-bp sequence spanning the ITS and 5.8S rDNA regions of Glomus occultum and Glomus brasilianum. In addition, this primer pair successfully detected G. occultum and G. brasilianum DNA in nested PCR using a primary PCR product amplified from highly diluted extracts of colonized corn (Zea mays) roots using modified ITS1:ITS4 primers. A second primer pair, GBRAS86:GBRAS388, primed specific amplification of a approximately 200-bp sequence spanning the ITS and 5.8S rDNA regions present only in G. brasilianum and Glomus strain GR582. Combined use of both primer pairs provides the means to detect and differentiate two ancient endomycorrhizal species, G. occultum and G. brasilianum, undetectable by standard root staining procedures. Sequence analysis showed that the purported G. occultum strain GR582 is likely a strain of G. brasilianum. PMID- 11267776 TI - Dependence of Staphylococcus epidermidis on non-transferrin-bound iron for growth. AB - The ability of Staphylococcus epidermidis strains to grow in the presence of human transferrin and varying amounts of ferric iron was studied. At initial bacterial densities up to 10(4) cfu ml(-1), none of the three strains grew when transferrin iron saturation was below the full saturation point, whereas the bacteria grew consistently when transferrin was fully iron-saturated and there was non-transferrin-bound iron in the medium. Precultivation of the bacteria under iron-restricted conditions to induce siderophore production did not abolish the growth dependence on non-transferrin-bound iron. At initial bacterial densities of 10(6) cfu ml(-1), the bacteria proliferated consistently also in the presence of partially saturated transferrin. The results indicate that at low bacterial densities, S. epidermidis cannot utilise transferrin-bound iron for growth and that its proliferation is dependent on non-transferrin-bound iron. PMID- 11267775 TI - Isolation and characterization of type IIS restriction endonuclease from Neisseria cuniculi ATCC 14688. AB - Neisseria cuniculi produces the restriction enzyme NcuI which is an isoschizomer of MboII. We have demonstrated that NcuI recognizes a pentanucleotide sequence (5'-GAAGA-3'/3'-CTTCT-5'), and cleaves the DNA 8 and 7 nucleotides downstream from the recognition site leaving a single 3'-protruding nucleotide. We have purified this enzyme to electrophoretic homogeneity using a four-step chromatographic procedure. NcuI endonuclease is a monomeric protein with a M(r)=48,000+/-1000 under denaturing conditions. The properties of NcuI are consistent with those for MboII, the position of the cleavage site being identical and the pH profile and divalent cation requirements being similar. Moreover, NcuI cross-reacts strongly with anti-MboII serum suggesting the presence of similar antigenic determinants. We have determined the sequence of 20 N-terminal amino acids for NcuI and concluded that this sequence is identical to the N-terminal portion of the MboII enzyme. PMID- 11267778 TI - The iron-induced ferredoxin FdxA of Campylobacter jejuni is involved in aerotolerance. AB - A gene encoding a putative 2[4Fe--4S] ferredoxin (FdxA) was identified upstream of, and divergent to the peroxide stress defense gene ahpC of the microaerophilic pathogen Campylobacter jejuni. The transcription start site of fdxA was located 27 and 28 bp upstream of the fdxA start codon. Transcriptional fusions of the fdxA promoter to a lacZ reporter gene demonstrated that expression of fdxA is iron-induced, and thus oppositely regulated to the iron-repressed ahpC gene. Insertional mutagenesis of the fdxA gene did not affect microaerobic growth of C. jejuni, but significantly reduced aerotolerance of C. jejuni. The fdxA gene is the first reported iron-induced gene of C. jejuni, and encodes a novel component of its oxidative stress defense. PMID- 11267777 TI - Cloning of the CYP51 gene from the eyespot pathogen Tapesia yallundae indicates that resistance to the DMI fungicide prochloraz is not related to sequence changes in the gene encoding the target site enzyme. AB - Resistance to sterol 14 alpha-demethylase inhibitor (DMI) fungicides has been correlated with mutations in the CYP51 gene encoding the target enzyme eburicol 14 alpha-demethylase. CYP51 was isolated from the eyespot pathogen Tapesia yallundae revealing a predicted 526-amino acid product exhibiting homology to other fungal CYP51s. CYP51 was sequenced from four field isolates sensitive or resistant to the DMI fungicide prochloraz and partially sequenced from two further isolates and eight progeny from a cross between prochloraz-sensitive and resistant parents. Two alleles of the gene were detected termed CYP51-1 and CYP51 2. No correlation was found between sequence change and fungicide sensitivity. Therefore prochloraz resistance involved a mechanism other than mutation in the target site gene. PMID- 11267779 TI - Identification and characterization of a gene encoding a 35-kDa protein from Mycobacterium avium subspecies paratuberculosis. AB - Mycobacterium avium subspecies paratuberculosis is the causative agent of Johne's disease, a chronic enteritis in ruminants. A gene homologous to that of 35-kDa antigen of Mycobacterium leprae was cloned and sequenced from Mycobacterium paratuberculosis. The database searches revealed 82.79% and 95.67% similarities of its nucleotide sequence, with those of immunodominant 35-kDa protein of M. leprae and M. avium, respectively. PMID- 11267780 TI - Initial degradation of dimethylphthalate by esterases from Bacillus species. AB - Dimethylphthalate (DMP), one of the phthalate esters, is used in the manufacture of plasticizers, insect repellents, and synthetic fibers, and contributes to environmental pollution. In the present study, we report a novel bacterium belonging to the Bacillus sp., which has the ability to utilize DMP as the sole source of carbon. The esterases from the cell-free extract of the Bacillus de esterified DMP. Native polyacrylamide gel electrophoresis showed the presence of four isoesterases designated Et1--4. The isoesterases Et-4 and Et-1 showed a higher preference towards DMP hydrolysis as compared with Et-2 and 3. A megaplasmid of about 60 kb was detected in this bacterium. The ability of this bacterium to utilize DMP as the sole source of carbon was lost upon plasmid curing. The isoesterases Et-1--4 were absent in the cell-free extracts of the cured bacterium. The results from our studies clearly demonstrate that de esterification is the initial step in the degradation of DMP and the genes for these esterases seem to be harbored on the plasmid in this bacterium. PMID- 11267781 TI - On the presence of peptide synthetase and polyketide synthase genes in the cyanobacterial genus Nodularia. AB - Nodularin is a hepatotoxin produced by the bloom-forming cyanobacterial species Nodularia spumigena. Putative peptide synthetase and polyketide synthase genes were detected in toxic strains of Nodularia by degenerate PCR. Using specific primer sets, peptide synthetase and polyketide synthase gene homologues were detected in nodularin-producing strains indicating a possible role of peptide synthetase and polyketide synthase enzyme complexes in the biosynthesis of nodularin. Strains of Nodularia isolated from around the world were also analyzed for the production of nodularin by the protein phosphatase 2A inhibition assay. The protein phosphatase inhibition assay and the molecular detection of peptide synthetase and polyketide synthase genes in Nodularia may be useful techniques for the assessment of nodularin-producing cyanobacteria in the environment. PMID- 11267782 TI - Monoclonal antibodies of IgA isotype specific for lipopolysaccharide of Salmonella enteritidis: production, purification, characterization and application as serotyping reagents. AB - Hybridomas secreting immunoglobulin A (IgA) monoclonal antibodies (MAbs) against Salmonella enteritidis lipopolysaccharide (LPS) were generated after mucosal immunization of BALB/c mice with heat killed bacteria. Antigen binding properties and specificity of the produced MAbs were studied in ELISA and immunoblotting with purified LPS. Two IgA MAbs agglutinated all Salmonella OD1 strains and all S. enteritidis clinical isolates. MAb 178H11 recognized O:9 antigen of subserogroup OD1 LPS. MAb 177E6/A9 reacted also with OD3 LPS antigen and agglutinated OD3 strains. These data suggest the existence of different O:9 antigen subspecificities, one presented in subgroup OD1 and the other common for OD1 and OD3. Thus the produced IgA MAbs prove to be useful reagents, which could differentiate OD1 and OD3 from OD2 strains. PMID- 11267783 TI - Expression and rapid one-step purification of biologically active His-tagged factor C by Ni(2+) affinity column chromatography. AB - Factor C is an unusual extracellular protein capable of inducing cytodifferentiation in certain Streptomyces strains. The protein is produced by Streptomyces griseus 45H at such a low amount that the study of its mode of action was hindered by the shortage of purified protein. We report here the expression of C-terminally hexa-His-tagged factor C in Streptomyces lividans and Escherichia coli. Expression in S. lividans is low while in E. coli it is relatively high, yielding about 5--10 mg of biologically fully active protein per liter culture. PMID- 11267784 TI - Modified xylE and xylTE reporter genes for use in Streptomyces: analysis of the effect of xylT. AB - The reporter gene xylE (encoding catechol 2,3-dioxygenase) has been modified for a more rational use in Streptomyces. Two reporter fragments, one containing xylE, and the other containing also the upstream gene xylT (which encodes a soluble ferredoxin), have been constructed to allow precise fusion of regulatory regions to the reporter genes. Identical fusions of these xylE and xylTE reporter fragments to the Streptomyces dagA and tipA promoters, in low and high copy number plasmids, show that the levels of xylE mRNA and catechol 2,3-dioxygenase activities are significantly higher when xylT is present. PMID- 11267785 TI - The role of Helicobacter pylori virulence factors in interleukin production by monocytic cells. AB - Helicobacter pylori infection results in chronic gastritis, which is initiated by the release of cytokines like interleukin (IL)-12 and IL-8 from mononuclear cells, and IL-8 from gastric epithelial cells. The severity of gastritis is influenced both by host factors and by bacterial factors such as the Cag proteins and the vacuolating cytotoxin VacA. Amounts of IL-12 and IL-8 produced by monocytic THP-1 cells differed considerably between the eight H. pylori isolates tested, but in contrast to H. pylori-induced IL-8 production by gastric epithelial cells, did not correlate to the Cag and VacA types of the strains. Apparently, in addition to Cag and VacA, other bacterial factors determine the extent in which H. pylori induced IL production in monocytes. PMID- 11267786 TI - Genetic differences between Escherichia coli O26 strains isolated in Brazil and in other countries. AB - Genomic diversity among 34 strains of Escherichia coli belonging to different serotypes of the O26 serogroup -- encompassing strains from different geographical origins and Shiga toxin-negative Brazilian strains -- was evaluated through random amplified polymorphic DNA (RAPD) analysis. Our results indicate that Brazilian and non-Brazilian O26 strains fall under distinct but closely related differentiation clusters. RFLP-PCR analysis of the fliC gene sequence was done in order to identify the H(-) serotypes and served to confirm the clustering pattern obtained in the dendrogram generated from RAPD data. The epidemiological significance of these data is discussed. PMID- 11267787 TI - Identification of genes regulated by prolonged acid exposure in Helicobacter pylori. AB - To investigate the influence of prolonged acid exposure on the gene expression, transcripts of Helicobacter pylori, grown under pH 5.5 and pH 7.4 for five successive passages, were analysed by differential display PCR. Eight genes were regulated by prolonged acid exposure. These genes included topA, tufB, ureB, flaA, atoE in the H. pylori genome and a cDNA fragment with 54% identity of the predicted amino acid sequence to a Bacillus cereus YkoW protein. The remaining two cDNA fragments had no significant homology to known sequences. Our data suggest that most of these genes might be required for the resistance of H. pylori to prolonged acid exposure. PMID- 11267788 TI - Cloning and DNA sequencing of the surface protein antigen I/II (PAa) of Streptococcus cricetus. AB - We have cloned and sequenced the gene encoding the surface protein antigen PAa (antigen I/II family) from Streptococcus cricetus E49 (serotype a) using degenerate PCR. The deduced amino acid sequence of PAa reveals two repeating regions (A region; alanine-rich region, P region; proline-rich region). Two additional tandem repeats were found in the A region and part of the P region was deleted compared to antigen I/II. Homology and phylogenetic analyses reveal that PAa is homologous to Streptococcus sobrinus PAg rather than Streptococcus mutans PAc. Using degenerate PCR a gene homologous to PAc was identified in Streptococcus intermedius, but not found in Streptococcus rattus or Streptococcus anginosus. PMID- 11267790 TI - Levels of ovine herpesvirus 2 DNA in nasal secretions and blood of sheep: implications for transmission. AB - A recently developed competitive PCR for ovine herpesvirus 2 (OvHV-2) was used to examine the levels of viral DNA in nasal secretions and peripheral blood leukocytes (PBL) of lambs and adult sheep. Viral DNA first appeared in the PBL of most lambs after about 3 months of age and the levels remained relatively constant thereafter. In most of the lambs (83%, n=12), viral DNA was undetectable by PCR in nasal secretions prior to 5 months of age. A dramatic rise of OvHV-2 DNA levels in the nasal secretions occurred starting at 5-6 months of age, which peaked at approximately 7 months. The highest level recorded in lamb nasal secretions was 7.5x10(8)copies/2microg DNA which were 75,000-100,000-fold higher than the levels in PBL of the same lambs. In adult sheep (n=10), the viral DNA levels in both PBL and nasal secretions were relatively stable over the 13-month period of the study, which included a lambing season. The data strongly suggest that neonatal lambs are not an important source for the transmission of OvHV-2 to clinically susceptible species, and that the nasal cavity is an important portal for shedding of infectious OvHV-2 in sheep. Furthermore, this study failed to identify a seasonal pattern in levels of viral DNA in nasal secretions or PBL of adult sheep that would provide a basis for the traditionally held belief that clinical cases of malignant catarrhal fever are significantly associated with lambing ewes. PMID- 11267789 TI - Experimental infection of pregnant ewes with bovine viral diarrhea virus type-2 (BVDV-2): effects on the pregnancy and fetus. AB - The reproduction effects of bovine viral diarrhea virus type-2 (BVDV-2) infection were investigated in ewes inoculated with a non-cytopathic BVDV-2 isolate at three stages of gestation. Virus inoculation was followed by a transient viremia, accompanied by a transient and mild hyperthermia and nasal discharge in a few animals. Some ewes were sacrificed at different time-points after virus inoculation to study the kinetics of fetal infection. Infectivity and viral antigens were detected in placentomes from day 7 to 36 post-inoculation (pi) and in fetal fluids and tissues between days 10 and 28 pi. Cardiac petechial hemorrhages and hemoperitoneum accompanied by a severe fibrinous ulcerative placentitis were observed in fetuses examined at days 21, 28 and 36 pi. Inoculation of ewes at days 55-60 of gestation resulted in a prolonged virus replication in placentomes and fetal tissues; ewes that were allowed to proceed with pregnancy had 77% of abortions or fetal and perinatal deaths. Seven stillbirths, unviable and viable lambs born to these ewes were virus-positive at birth. Infectious virus was repeatedly isolated from leukocytes of two lambs up to 2 and 6 months of age, indicating they were persistently infected. Ewes inoculated at days 65-70 of gestation had 66.6% of fetal and perinatal losses. Three viable lambs born to these ewes were healthy, BVDV antibody-positive and virus-negative. A transient viral replication in placentomes and in a few fetal tissues, followed by the rise of fetal neutralizing antibodies and virus clearance was the result of inoculating ewes at days 120-125 of gestation. Lambs born to these ewes were healthy, antibody-positive and virus-negative. These results demonstrate that the biology of BVDV-2 infection in pregnant sheep is essentially similar to that of BVDV-1 in pregnant cattle and sheep. These features make this species an attractive animal model for studying the pathogenesis of congenital BVDV-2 infection. PMID- 11267791 TI - Molecular epidemiological confirmation and circumstances of occurrence of sheep (S) strains of Mycobacterium avium subsp. paratuberculosis in cases of paratuberculosis in cattle in Australia and sheep and cattle in Iceland. AB - Distinct strains of Mycobacterium avium subsp. paratuberculosis with a tendency to segregate in either sheep, or cattle and other ruminants, have been described and are known as S and C strains, respectively. These strains can be distinguished by a polymorphism in the IS1311 element and other DNA-based methods. C strains are relatively easy to culture from tissues and faeces of animals with paratuberculosis but S strains are difficult to culture. A retrospective survey of archival formalin-fixed paraffin-embedded tissue samples from culture negative Australian paratuberculous cattle was undertaken to determine whether infection in these cases was due to S strains. Polymerase chain reaction and restriction endonuclease analysis of the amplified product was used to identify the polymorphism in IS1311. Three cases of bovine paratuberculosis due to S strain were confirmed from three different farms. A serological survey led to the identification of a further two cases on one of these farms. S strains were also identified in archival tissues from paratuberculous sheep and cattle from Iceland, confirming epidemiological and microbiological evidence that paratuberculosis in Iceland was due to S strain following importation of infected sheep from Europe. In each bovine case in both Iceland and Australia there had been direct or indirect contact of calves with paratuberculous sheep. We were unable to determine whether S strains had established endemic infection in cattle or whether repeated infection from sheep had occurred. Limited epidemiological evidence suggests that transmission of S strains to cattle in Australia has been uncommon under extensive grazing conditions. In Iceland, different husbandry practices appear to have favoured transmission of S strains to cattle. PMID- 11267792 TI - Characterisation and clonal relationships of Shiga-toxigenic Escherichia coli (STEC) isolated from Australian dairy cattle. AB - A total of 136 Shiga toxin-producing Escherichia coli (STEC) isolated during a longitudinal survey of three Australian dairy farms were examined to determine their virulence factors, serotype and genomic relationships. This study aimed to assess the potential of these STEC to cause disease in humans and to analyse the on-farm ecology of STEC. Virulence factors (stx, eae, ehxA) were used as determinants of potential to be enterohaemorrhagic E. coli (EHEC) and were examined using polymerase chain reaction (PCR). Among the cattle groups tested, calves, both before and during weaning, shed the most putative EHEC and were the main source of serotypes commonly associated with human disease. E. coli O157:H7 and E. coli O26:H11 represented 9.4 and 7.8% of cattle STEC isolates respectively, with other putative EHEC serotypes reported for the first time from cattle. Based on serotype and virulence factors, 20% of STEC were putative EHEC. Pulsed-field gel electrophoresis (PFGE) was used to compare the genomic profiles of STEC from dairy farms. Isolates common to cattle and the farm environment were identified. Multiple strains of STEC with high clonal turnover were detected in the faeces of cattle, and isolates appeared to be specific to individual farms. To fully assess the pre-slaughter EHEC risk factors on-farm, examination of STEC virulence is as important as determination of STEC prevalence. PMID- 11267793 TI - Genomic diversity of Bartonella henselae isolates from domestic cats from Japan, the USA and France by pulsed-field gel electrophoresis. AB - The genomic DNA diversity of 27 Bartonella henselae and three B. clarridgeiae isolates from 18 domestic cats from Japan, the USA and France was investigated by pulsed-field gel electrophoresis (PFGE) with NotI, AscI and SmaI restriction enzymes. A great diversity of genomic patterns was found for all B. henselae, but none for B. clarridgeiae isolates. The DNA size of B. henselae and B. clarridgeiae isolates were 1.7-2.9 and 1.7Mbp, respectively. All 13 Japanese cat isolates were identified as B. henselae type I. Furthermore, three of the four Japanese cats harbored genetically different B. henselae type I isolates, suggesting for the first time co-infection with various type I isolates. One French cat and one American cat were co-infected with B. henselae and B. clarridgeiae. B. henselae type I and type II were mainly grouped in two different clusters by PFGE using SmaI endonuclease in the dendrogram. PMID- 11267794 TI - Influence of Escherichia coli and Arcanobacterium pyogenes isolated from bovine puerperal uteri on phenotypic and functional properties of neutrophils. AB - When cows develop endometritis after birth, Escherichia coli and Arcanobacterium pyogenes are usually the most prominent bacteria present in bovine uterine lochial secretions. A. pyogenes alone is rarely found in the course of a disturbed puerperium. This was confirmed in this study, since average and high grade uterine contaminations were always associated with the presence of both bacteria. The contamination grade was positively correlated with uterine polymorphonuclear granulocyte (PMN) numbers and negatively correlated with blood PMN numbers. Whether E. coli and A. pyogenes affect the phenotype and function of bovine PMN in a similar or differential way was subject to in vitro studies. PMN were tested in the presence of washed bacterial fragments or culture supernatants taken as a source for soluble and/or secreted bacterial products. Fragments and soluble products differed only quantitatively in their effects on PMN. Usually, long-time exposure (24h) of PMN to fragments induced the strongest effects. Accelerated death of granulocytes was only moderately induced by both E. coli and A. pyogenes products. Both E. coli and A. pyogenes products induced the enhanced expression of a membrane molecule detected by mAb IL-A110 and of CD11b. Expression of other surface structures remained largely unchanged (MHC class I, CD11c). Functional parameters of PMN (phagocytosis; generation of reactive oxygen species, ROS; antibody-independent cellular cytotoxicity, AICC) generally declined after pre-incubation for 24h with products of E. coli or A. pyogenes. Interestingly, soluble products of A. pyogenes stimulated the phagocytosis of PMN. However, co-incubation with E. coli products abrogated this stimulatory effect. The results supply evidence for similar modes of action of the gram negative E. coli and the gram-positive A. pyogenes on bovine PMN. Alterations in PMN function and phenotype are mainly triggered by direct contact between bacterial fragments and PMN. Inhibition experiments with polymyxin B demonstrated that E. coli-mediated effects were not solely due to the action of lipopolysaccharide. The dominant functional depression of neutrophils by E. coli products strengthens the suggestion that the earlier appearance of E. coli in the uterus may support the co-infection of this organ by A. pyogenes at later times. PMID- 11267795 TI - Dichelobacter nodosus serotype M fimbrial subunit gene: implications for serological classification. AB - Dichelobacter nodosus fimbrial subunit gene (fimA) from a serotype M strain (M SPAHL) was investigated in this study. A primer set targeting the relatively conserved fimA regions and based on the published sequence from Nepalese serogroup M isolates (Nepalese M), failed to amplify the fimA of M-SPAHL. However, when the downstream primer was substituted with a primer that is specific for other serogroups of D. nodosus, the fimA was successfully amplified. Cloning followed by DNA sequencing, revealed that the M-SPAHL fimA was different to the Nepalese M fimA. The predicted amino acid sequence of the M-SPAHL fimA did not show homology to any known serogroups or serotypes. The most similar sequence was from serotype F1, and not Nepalese M. The consequences of serological relatedness and sequence dissimilarity are discussed. PMID- 11267796 TI - The role of pheromones and biostimulation in animal reproduction. AB - It is now known that pheromonal communication plays an important role in mammalian behaviour and reproductive processes. Chemical communication with pheromones is one means of transmitting such information. In mammals, signalling and priming pheromones are thought to act either singly or in combination through olfaction, auditory, visual (sight) or tactile stimuli. Pheromones are air-borne chemical substances ("signals") released in the urine or feces of animals or secreted from cutaneous glands that are perceived by the olfactory system and that elicit both behavioural and endocrine responses in conspecifics. Extensive studies in insects, rodents, swine, sheep, goats and cattle have established the importance of pheromones in the strong influence exerted by the male on reproductive activity in the female. There is a pheromone produced by the queen honey bee, which has two functions: inhibition of queen rearing and suppression of oogenesis in workers and in addition attracts drones during nuptial flight. It has also been demonstrated that the urine of male mice, rats, feral species and other wild rodents contains a priming pheromone that is responsible for hastening puberty in the females. Pheromones in the wool, wax and urine of a ram are sufficient to stimulate ewes to ovulate, while the buck has a strong characteristic seasonal odor and a buck jar containing the odor of the buck can be used as an aid in the detection of oestrus in does. The mere presence of the boar at the time of insemination of the sow improves sperm transport and ovulation, while the presence of the vasectomised bull has been reported to hasten the onset of puberty in heifers and also early resumption of ovarian activity in cattle following parturition. The role of pheromones in bovine reproduction is not as clearly defined as in sheep, goats and swine. Pheromones and other allelomimetic cues can exert profound effects on reproductive activity via the hypothalamic system that generates pulses of gonadotropin-releasing hormone. Manipulations of these factors and other pathways linking environmental inputs to reproductive output can lead to developing the concept of "control systems technologies", aimed at controlling reproductive performance. The knowledge acquired on the effectiveness of biostimulation; the factor which conditions it and the biological mechanism which produces it in livestock species, allows its use as a breeding management tool. The understanding of the role of pheromones could be of potential economic importance in addressing some of the problems associated with livestock production in the tropics. The biostimulation technique offers a potentially useful and practical way to improve reproductive efficiency in livestock species in the tropics. The exact nature of the cues and the role of biostimulation in livestock species especially swine, sheep, goats and cattle in developing countries require more attention. PMID- 11267797 TI - Developmental capacity of Bos indicus oocytes after inhibition of meiotic resuption by 6-dimethylaminopurine. AB - Several reports have suggested that a treatment before in vitro maturation might improve oocyte competence and increase its developmental potential. Therefore, the objectives of the present study were to establish the kinetics of IVM in Zebu oocytes, to assess the effect of 6-dimethylaminopurine (6-DMAP), a phosphorylation inhibitor, on meiotic resumption, and to verify the developmental potential of the blocked oocytes after removal of the inhibitory conditions. To establish the kinetics of in vitro maturation 1422 oocytes were obtained from Nellore cows ovaries and matured in presence and absence of gonadotropins. Samples of oocytes were taken from culture at 0, 6, 9, 12, 15, 18, 21 and 24h, and the oocytes were fixed, stained and evaluated for nuclear morphology. Germinal vesicle break down (GVBD) occurred between 6 and 12h of culture in both groups. By 21h the majority of the oocytes had reached metaphase II in presence (71%) and absence (62%) of gonadotropins. In order to examine the inhibitory effect of 6-DMAP, 585 oocytes were cultured for 12, 18 and 24h in the presence or absence of 2mM of 6-DMAP. At each time point the oocytes were evaluated for nuclear morphology. To test the reversibility of meiotic inhibition 366 oocytes were incubated for 0, 12, 18 and 24h in the presence of 6-DMAP and then were transferred to the maturation medium and cultured for further 24h. A total of 429 oocytes were used to evaluate the developmental potential after meiotic inhibition. The oocytes were cultured in the presence of 6-DMAP for 0, 12, 18 and 24h, and then were matured, fertilized and cultured in vitro. Culture of bovine oocytes in the presence of 6-DMAP up to 24h completely blocked GVBD with more than 90% of the oocytes at GV stage. The inhibitory effect of 6-DMAP was fully reversible since maturation rates were similar (P>0.05) among all treatment groups. The evaluation of embryo development after various periods of meiotic blockage showed that inhibition, regardless the time period, had no effect (P>0.05) on penetration and cleavage rates. However, the proportion of embryos at blastocyst stage was reduced after inhibition for 12 (20.2%), 18 (20.1%) and 24h (19.0%) compared with the control group (35.6%). 6-DMAP has a reversible effect on maintenance of meiotic arrest, but reduced further embryo development. PMID- 11267798 TI - The use of in vitro fertilization techniques to investigate the fertilizing ability of bovine sperm with proximal cytoplasmic droplets. AB - The objective of this experiment was to determine the effect of proximal droplets on sperm-oocyte binding, zona penetration, fertilization, and the developmental competence of oocytes fertilized by sperm with proximal droplets (PD) in an in vitro fertilization (IVF) and culture system. Frozen semen from three bulls (PD1, PD2 and PD3) with varying proportions of normal appearing sperm with proximal droplets and semen from a normal control bull (C) were used in this experiment. The mean number of sperm bound to the zona pellucida (26.8 +/- 2.0, n=100; 15.2 +/- 1.1, n=100; 16.2 +/- 1.0, n=100) for bulls PD1, PD2, and PD3, respectively, were significantly lower (P<0.05) than that of the control bull C (47.4+/- 1.9; n=114). No spermatozoa with PD were found bound to the zona pellucida and this finding was consistent among the three bulls. The percentage penetration of zonae for the bulls PD1, PD2 and PD3 (74%, 74/100; 71%, 71/100 and 69%, 69/100, respectively) were not different than that of bull C (72%, 179/245). Similarly, the mean number of sperm penetrating the zona pellucida (1.43+/- 1.2, 1.24 +/- 1.1 and 1.20 +/- 1.1, for bulls PD1, PD2 and PD3, respectively) were not different than that of bull C (1.45 +/- 1.1). However, fertilization rates (8.8%, 8/90; 16.8%, 16/95; and 10.6%, 11/103, for bulls PD1, PD2 and PD3, respectively) were lower (P<0.001) than that of bull C (68.7%; 77/112). Similarly, cleavage rates (5%, 10/200; 8%, 8/100 and 14%, 15/111) for the bulls PD1, PD2 and PD3, respectively, were lower than that of the control bull, C (60.7%; 79/130). Cleaved zygotes resulting from the fertilization of oocytes by apparently normal sperm from bulls with PD did not develop beyond cleavage, whereas, 43.8% (57/130) morulae and 20% (26/130) blastocysts were produced by oocytes fertilized by sperm from bull C. In summary, normal appearing sperm with PD did not bind to the zona pellucida. Apparently normal sperm with out proximal droplets co-existing in the semen along with sperm containing PD were also functionally deficient, resulting in reduced zonae binding and zygotes resulting from insemination with semen with a high percentage of PD did not develop beyond cleavage. PMID- 11267799 TI - Contraceptive efficacy of an intra-uterine device in Brahman cattle. AB - The contraceptive efficacy of an intra-uterine device was evaluated using 218 heifers and 212 cows on three north Australian cattle stations. The heifers were aged approximately 2 years and weighed 250-378 kg; the cows were aged 3-16 years and weighed 256-540 kg. All cattle were non-pregnant, non-lactating Brahmans. At the end of the monsoon (wet) season (April-June 1997), the cattle were allocated by stratified randomisation to the three treatments which were untreated controls (n=59), surgical ovariectomy (n=105), or implantation with a bovine intra-uterine device (BIUD; n=266). All cattle grazed and were managed as one group within each station. They were exposed to bulls (4 per 100 females) from soon after treatment until slaughter approximately 12 months later. The BIUD could not be implanted in 25% of heifers and 8% of cows due to narrow or twisted cervices. Correct placement of the BIUDs appeared to be achieved in 57% of heifers and 72% of cows. At slaughter, the devices were incorrectly positioned in 73% of heifers and 49% of cows into which BIUDs had been inserted and that remained non-pregnant. Uterine perforations by the BIUD were observed in 35 and 45% of these heifers and cows, respectively; most perforations appeared to occur during implantation. Low grade endometritis was observed at slaughter in most BIUD-implanted animals; 2% had pyometra.BIUD animals did not have significantly different growth to that of control or ovariectomised animals, other than when ovariectomy suppressed growth following surgery. Most animals implanted with BIUDs appeared to have normal ovarian function and animals were observed mating. All ovariectomised animals remained non-pregnant. Over 80% of controls were pregnant within 8 months of exposure to bulls, except heifers at one station where pregnancy rate was restricted to 25% as a result of severe nutritional conditions. Pregnancy was diagnosed in 21% of heifers and 33% of cows with implanted BIUDs. The device remained correctly positioned and with no pregnancy diagnosed in the year following implantation in only 2% of heifers and 14% of cows originally allocated. Because of the difficulties of implanting BIUDs, the high frequency of associated uterine injury, the high pregnancy rate in implanted animals, and that growth was unaffected by the presence of a BIUD, it was concluded that the device had poor contraception efficacy and no growth-promotant effect in Brahman cattle. PMID- 11267800 TI - Reduction of infectious epizootic hemorrhagic disease virus associated with in vitro produced bovine embryos by non-specific protease. AB - Infectious viruses bind more tenaciously to the zonae pellucidae of in vitro produced bovine embryos than to zonae of in vivo derived embryos. Currently, the International Embryo Transfer Society recommends that all in vivo derived embryos be subjected to a rigorous washing procedure in combination with exposure to trypsin to remove viruses adherent to the zonae. In contrast to in vivo derived embryos, this method is not effective for disinfecting in vitro produced embryos. Our hypothesis was that a more potent, non-specific protease from Streptomyces griseus (S. griseus) would provide a more effective treatment for virus removal from in vitro produced bovine embryos. Bovine oocytes were matured, fertilized, and cultured in completely defined in vitro conditions. Zygotes were washed according to the procedure outlined by the International Embryo Transfer Society, replacing trypsin with the experimental protease. Experimental incubations were with 0.1% (4 units/ml) protease for 0, 30, 45, 60 and 75s intervals. Embryos were able to withstand exposure to this enzymatic treatment for only 45s before their developmental potential was significantly reduced; 60s exposure was detrimental (P<0.05). Oocytes were exposed to epizootic hemorrhagic disease virus serotype 2 (EHDV-2, 10(6) TCID(50)/ml) during in vitro maturation. Resulting zygotes were washed according to the International Embryo Transfer Society procedure and either exposed to trypsin or protease. Exposure to EHDV-2 prevented cumulus expansion and markedly reduced embryonic development (P<0.05). There were no differences in development among virus exposed groups receiving no treatment or treatment with trypsin or protease. However, proportions of infected embryos were reduced after protease treatment versus positive controls and trypsin treated embryos. PMID- 11267801 TI - Influence of oocyte quality, culture media and gonadotropins on cleavage rate and development of in vitro fertilized buffalo embryos. AB - The present study was designed to examine the influence of oocyte quality, culture media and gonadotropins on cleavage rate and development of in vitro fertilized buffalo embryos. Three experiments were conducted. In experiment 1, oocytes were classified by number of cumulus cell layers and morphology of the ooplasm as good, fair or poor. Oocytes were cultured for IVM, IVF and IVC in CR1aa medium. In experiment 2, good quality oocytes were cultured for maturation in: (1) CR1aa; (2) CR2aa; (3) TCM-199; (4) MEM or (5) RPMI-1640, and then fertilized using frozen thawed buffalo spermatozoa in CR1aa. The oocytes were cultured in the same medium used for maturation after fertilization. In experiment 3, oocytes were classified into three groups: group (1) was without gonadotropin and serve as a control; group (2) in which IVM medium was supplemented with 10microg/ml FSH and group (3) in which IVM medium was supplemented with 10IUml(-1) eCG. In all experiments, oocytes were kept at 38.5 degrees C under 5% CO(2) for IVM, IVF, IVC and examined for cleavage and embryo development rates on days 3 and 8, respectively. Good and fair quality oocytes produced a higher cleavage rate (P<0.01) than poor quality oocytes. Morula production rate was also higher (P<0.01) for good as compared with fair quality oocytes. Embryo development with poor quality oocytes was arrested at the two to sixteen cell stage. In experiment 2, the cleavage rate was higher (P<0.05) in CR1aa than CR2aa, and higher (P<0.01) than TCM-199, MEM and RPMI-1640. The numbers of morulae and blastocysts were higher (P<0.01) for oocytes cultured in CR1aa and CR2aa media than TCM-199 or MEM. In experiment 3, the addition of FSH or eCG to the maturation medium increased (P<0.01) cleavage and developmental rates of buffalo embryo compared with control medium. In conclusion, the IVM of good quality buffalo oocytes in CR1aa or CR2aa medium and the addition of FSH or eCG in maturation medium produced higher cleavage and developmental rates of IVF buffalo embryos. PMID- 11267802 TI - The effect of low doses of naloxone on the preovulatory surge of LH and on the onset and duration of oestrus in the ewe with induced oestrus during the non breeding season. AB - The objective of this work was to study the effect of the endogenous opiate peptide (EOP) antagonist, naloxone, on the preovulatory LH surge and on the time of onset and duration of oestrus in the ewe with induced oestrus during the non breeding season. Forty Suffolk X Hampshire ewes 2-3-years-old and 50+/- 4kg were studied, ewes were divided at random in two groups of 20, housed in open paddocks under natural photoperiod (19 degrees latitude N); were fed with hay and commercial pellets, and provided water ad libitum. Group one received an intravaginal sponge with 45mg of medroxiprogesterone acetate for 14 days, and upon sponge withdrawal 250IU of eCG was administered i.m. Group two received the same treatment as group 1 but in addition they received two i.m. injections of 0.5mg of naloxone, one given on sponge withdrawal and the second 24h later (total dose 1.0mg). Oestrus in naloxone-treated ewes was present 32+/- 2h and in control ewes in 35+/- 3h after sponge withdrawal. Duration of oestrus in control ewes was shorter (27+/- 2.5h), than naloxone-treated ewes (39+/- 6h); (P<0.0001). The LH surge in naloxone-treated ewes was initiated 5h after onset of oestrus, and 8h after onset of oestrus in control ewes, and the difference was significative (P<0.0006). It was concluded that EOP are important modulators of reproductive function in the ewe. PMID- 11267803 TI - Ultrasonographic study of the effects of the corpus luteum on antral follicular development in unilaterally ovulating western white-faced ewes. AB - The objective of this study was to examine the local effects of the corpus luteum (CL) on ovarian antral follicle development by looking at follicle populations and dynamics in ovaries with or without CL, in unilaterally ovulating ewes, using a retrospective analysis of daily ultrasonographic records. The present report summarises the data from the first luteal phase of the breeding season (August October; n = 4), a luteal phase in the mid-breeding season (November-December; n = 5), the last luteal phase of the breeding season (January-March; n = 5), and the luteal phase after GnRH-induced ovulations in mid-anoestrus (May-June; n = 4) of western white-faced ewes. Mean daily numbers of 3mm follicles that did not grow any larger were significantly reduced in the CL-containing ovaries of ewes at all periods of study except for the transition to anoestrus. With all scanning periods combined, daily numbers of 3mm follicles not growing further increased (P<0.05) between day 6 and 15 after ovulation in the CL-containing ovaries. Based on mean data for the whole periods of observation, the non-CL-bearing ovaries of ewes in the transition to anoestrus had fewer (P<0.05) follicles growing from 3 to > or =5mm in size before regression compared with the mid-breeding season and mid-anoestrus. The lifespan of follicles reaching > or =5mm in diameter was shorter (P < 0.05) in the CL- compared with non-CL-containing ovaries of anoestrous ewes induced to ovulate with GnRH ((6.5+/- 1.3) and (9.0+/- 1.0) days, respectively). Circulating concentrations of progesterone were lower during both transitional periods (into and out of anoestrus) and mid-anoestrus than during the mid-breeding season (P < 0.001), and were less during anoestrus than during both transitional periods (P < 0.05). It was concluded that CL/luteal structures locally suppressed the growth of ovarian antral follicles to the 3mm size-range except during the transition to anoestrus, but that there was no inhibitory effect of the CL on the growth of ovarian follicles to larger diameters. The presence of CL/luteal structures did not affect the length of the lifespan of follicles reaching > or =5mm in diameter nor the number of ovulations per ovary in cyclic ewes, but shortened large follicle lifespan in anoestrous ewes. Variations in peripheral concentrations of progesterone across the breeding season and between the breeding season and anoestrus did not alter the lifespan of large antral follicles. In the transition to anoestrus and during mid anoestrus, the presence of the CL in an ovary appeared to maintain follicle development to ovulatory sizes and to increase the rate of turnover of large antral follicles, respectively. PMID- 11267804 TI - Freezability of spermatozoa from Finn and Dorset rams in multiple semen extenders. AB - Ten semen extenders were tested in two experiments for cryopreservation of semen collected from four Finn and four Dorset rams. Two ejaculates of semen were combined from each ram for testing each extender treatment. The extenders consisted of a series of commonly used egg yolk-TRIS media with and without sodium and triethanolamine lauryl sulfate (STLS), a similar extender with 3-N morpholino propane sulfonic acid (MOPS), and milk and whey extenders. In Experiment 1, extender treatments were replicated with three sets of collections from the eight rams, and in Experiment 2 with two sets. The egg yolk-TRIS glycerol-STLS (EY(1)TSTLS) extender was significantly superior to other extenders except whole milk in protecting the sperm during freezing and thawing. In Experiment 1, a 20% egg yolk-TRIS-glycerol-STLS extender preserved 71% of the progressively motile Finn sperm (post-thaw divided by pre-freeze percentage of motile sperm), and 76% of the Dorset sperm. In Experiment 2, the corresponding values for the same EY(1)TSTLS extender used with Finn and Dorset sperm were 86 and 64%, respectively. Without STLS the egg yolk extenders were significantly less effective in protecting cryopreserved ram sperm. This egg yolk-TRIS extender, containing STLS and glycerol, may hold promise for freezing ram sperm that could be used successfully for intracervical insemination. PMID- 11267806 TI - Origin of the preovulatory follicle in Mouflon sheep (Ovis gmelini musimon) and effect on growth of remaining follicles during the follicular phase of oestrous cycle. AB - Daily transrectal ultrasonographies were conducted to study development of all follicles with antral diameters > or = 2mm during the follicular phase of oestrous cycle in Mouflon, a strictly monovular wild-sheep. A total of 14 follicular phases was studied after oestrus synchronization with two cloprostenol doses, 9 days apart, in five cyclic Mouflon ewes. In 13 cycles (92.8%), the ovulatory follicle arose from those antral follicles present in both ovaries when luteolysis was induced, being the largest one with a mean size of 4.4+/- 0.3mm at that moment in 10 cycles (76.9%). The remaining cycles had a larger follicle, but it was decreasing in size. Appearance of new follicles > or =2mm in size remained unaffected during the follicular phase (3.7+/- 0.2), but there was found a linear decrease in the number of those growing to > or =3mm (2.5+/- 0.4 to 1.1+/- 0.2, P < 0.05) and > or = 4mm (0.6+/- 0.2 to 0.1+/- 0.1, P < 0.005), detection of new follicles growing to > or = 5mm was negligible. Then, number of medium (4-5mm) growing follicles present in both ovaries decreased from 1.5+/- 0.3 at 0 h to 0.3+/- 0.1 at 72 h (P<0.005). In conclusion, the single ovulatory follicle is the largest growing follicle present in both ovaries at the moment of luteolysis. This follicle is selected to grow and ovulate while development of other follicles is inhibited. PMID- 11267805 TI - Superovulation in ewes by a single injection of pFSH dissolved in polyvinylpyrrolidone (PVP): effects of PVP molecular weight, concentration and schedule of treatment. AB - Three experiments were carried out to evaluate induction in ewes of superovulation and embryo production by a single injection of a porcine pituitary extract (pFSH) dissolved in polyvinylpyrrolidone (PVP), investigating the effects of PVP molecular weight and its concentration (Experiment I), time and method of treatment (Experiments II and III). All ewes were synchronized for estrus by vaginal sponges impregnated with fluorogestone acetate (FGA; 30 mg, 9 days) plus PGF(2alpha) (Cloprostenol, 50 microg, 48h before sponge removal - s.r.), and superovulated by 250 IU pFSH. In Experiment I, 60 Gentile di Puglia ewes were subdivided into five experimental groups (n = 12): Group A, the control, received six decreasing intramuscular (i.m.) doses of pFSH, 12 h apart, beginning 48h before s.r.; Groups B and C were given 48 h before s.r. a single i.m. injection of pFSH dissolved in PVP with MW = 10,000, respectively, at concentrations of 15 and 30% w/v; Groups D and E received the same treatments as for B and C using PVP with MW = 40,000. None of the pFSH-PVP treatments were effective in inducing superovulation. In Experiment II, 22 Leccese ewes were subdivided into two groups (n = 11): Group A, control received i.m. four decreasing doses of pFSH, beginning 24 h before s.r., 12h apart; Group B was given a single i.m. injection of pFSH dissolved in PVP (MW = 40,000 at 30% w/v), 24 h before s.r. The pFSH-PVP treatment provided an ovulation rate similar to the control and tended to enhance embryo yield (4.4 versus 2.4, P>0.05). In Experiment III, 60 Leccese ewes were subdivided into six treatment groups (n = 10). Groups A and D served as controls and received i.m. 12 h apart, six doses (from 48 h before s.r.) and four doses (from 24h before s.r.) of pFSH, respectively. Groups B and C were treated by a single injection of pFSH in PVP (MW = 10,000; 30% w/v) 48 h before s.r., respectively by i.m. or subcutaneous (s.c.) administration. Groups E and F received the same treatments as for B and C 24 h before s.r. Intramuscular pFSH PVP administration 24 h before s.r. provided an ovulation rate (8.1), mean numbers of ova recovered (5.6) and fertilized (4.2) comparable to the six or four dose treatments and significantly higher (P <0.01) compared to the pFSH-PVP treatment carried out i.m. 48 h before s.r. These results show that a single injection of pFSH dissolved in PVP at 30% w/v, performed i.m. 24 h before s.r., is able to induce a superovulatory response comparable to that following multiple injection treatment, regardless of PVP molecular weight. PMID- 11267807 TI - Body weight loss during lactation and its influence on weaning-to-service interval and ovulation rate in Landrace and Yorkshire sows in the tropical environment of Thailand. AB - The aim of this study was to investigate the ovulation rate and the weaning-to service interval (WSI) of sows in relation to their body weight loss during lactation in tropical climatic conditions. Effect of lactation length (LL), number of total born piglets, number of live born piglets, litter birth weight, average piglet birth weight, number of pigs weaned, litter weaning weight and average pig weaned weight on sow weight loss during lactation were also studied. This study was conducted in two commercial purebred sow herds (A, B) in the central part of Thailand from August to December 1997. The herds had both Landrace (L) and Yorkshire (Y) sows. The 123 sows (55 L and 68 Y) in herd A and 153 sows (95 L and 58 Y) in herd B, parity 1-4, were weighed within 4 days after farrowing and at weaning. Lactation length, litter size at birth and at weaning, litter weight at birth and at weaning, and WSI were recorded for each of these sows. In herd A, 52 sows (20 L and 32 Y) were examined once by laparoscopy between days 8 and 14 after AI-service. These sows had farrowed at least seven piglets in the previous parturition. The numbers of corpora lutea (CL) in both ovaries were counted, and were assumed to equal the ovulation rate. L-sows had significantly (P < 0.05) higher relative weight loss during lactation (RWL) than Y-sows. The RWL increased by 0.7% for each extra pig weaned. When LL increased by 1 day, within the interval of 17-34 days, RWL decreased by 0.6%. Sows with a high weight loss had significantly (P < 0.05) longer WSI than sows with medium or low weight loss. Weight loss had a significant (P < 0.05) effect on WSI in parity 1 and 2 sows. Y-sows had more CL than L-sows (15.7 versus 14.0) (P < 0.05). RWL, parity and regression on lactation length had no significant effect on number of CL. In conclusion, sows with higher number of pigs weaned lose more weight. Under the restricted feeding regime applied, high weight loss during lactation prolongs WSI in parity 1 and 2 sows, but has no influence on the ovulation rate at first oestrus after weaning. The ovulation rate is higher in Yorkshire than in Landrace sows. The ovulation rate is independent of parity. PMID- 11267809 TI - Capnography alone is imperfect for endotracheal tube placement confirmation during emergency intubation. AB - This analysis primarily sought to determine the effectiveness of end-tidal capnography for tube placement confirmation during emergency airway management. Secondary objectives were validation of the rate of unanticipated esophageal placement during emergency intubation and quantification of the portion of intubations performed in patients with cardiac arrest where capnography is not recommended. The study was performed in two phases. For the primary objective, a meta-analysis was performed on all experimental capnography trials enrolling emergency populations. For the secondary objectives, inadvertent esophageal intubation and cardiac arrest rates were calculated from a large prospective multicenter observational study of emergency intubation cases. Data analysis included calculation of descriptive statistics, sensitivity, specificity, and confidence intervals (CI). Based on 2,192 intubations, a meta-analysis of previous capnography trials resulted in an aggregate sensitivity of 93% (95% CI 92-94%) and an aggregate specificity of 97% (CI 93-99%) for emergency tube placement confirmation. Thus, for emergency capnography use, the false-negative failure rate (tube in trachea but capnography reports esophagus) was 7% and the false-positive rate (tube in esophagus but capnography reports trachea) was 3%. This translates to potential harm for one patient in every 10 treated with capnographic confirmation alone (number needed to harm: 14 for false-negative, 33 for false-positive, and 10 for both). A further literature review demonstrated no sole method of tube placement confirmation is completely foolproof. Of 4,602 consecutive intubations reported to the National Emergency Airway Registry, 4% of emergency intubation attempts resulted in accidental esophageal intubation, and 10% occurred in nontraumatic cardiac arrest patients. During tracheal intubation of critically ill patients, it is concluded that the rate of accidental esophageal tube placement warrants continued improvement in emergency airway techniques. Misidentification of esophageal placement in the emergency setting may occur with capnography. Multiple methods of tube placement confirmation are superior to any single method because no single method has perfect accuracy. PMID- 11267808 TI - The photophase light intensity does not affect the scotophase melatonin response in the domestic pig. AB - This study investigated the effects of the photophase light intensity on the scotophase melatonin response. Twelve, 8-month-old crossbred gilts were allocated to three groups of four and housed in temperature- and lighting-controlled climate rooms. The rooms had a light intensity of 40, 200 or 10,000 lx and a light-dark cycle of 12 L:12 D. The gilts were allowed to acclimatize to a new lighting regimen for 1 week before being sampled at 2h intervals for 24h. Following the sampling, pigs were transferred under a different light intensity, allowed to adjust for 1 week and sampled again. The procedure was repeated three times so that all the groups went through all three lighting regimens (light intensities). All the gilts exhibited a clear circadian serum melatonin rhythm under each lighting regimen with high melatonin concentrations occurring during the scotophase. There was no difference in the scotophase melatonin response in terms of mean concentrations or duration of increased melatonin levels within or between the groups under different lighting regimens. There was considerable inter-individual variation in the dark phase melatonin response but the individual profiles were consistent under the different lighting regimens. It is concluded that when a certain threshold light intensity (<40lx) is exceeded, the photophase light intensity has no effect on the scotophase melatonin response. These results imply that extremely high light intensities during the photophase would provide no additional benefits compared with normal comfortable light intensity, if artificial lighting programs were introduced to commercial piggeries in order to reduce seasonal effects on reproduction. PMID- 11267810 TI - A prospective multicenter trial testing the SCOTI device for confirmation of endotracheal tube placement. AB - We sought to characterize the Sonomatic Confirmation of Tracheal Intubation (SCOTI) device's ability to confirm endotracheal tube location during real-time intubation in emergency and elective settings. Data were prospectively collected during a multicenter convenience-sample observational trial of emergency and elective intubation cases. In addition to tracheal and inadvertent esophageal intubations in emergency patients, intentional esophageal intubations were also performed to improve specificity calculations in consenting elective surgical patients. Data analysis included descriptive statistics as well as calculations of sensitivity, specificity, and 95% confidence intervals (CI). Data were obtained from 220 tracheal and 103 esophageal intubations, 137 (42%) performed in emergency patients. Fifteen tracheal intubations were incorrectly identified by SCOTI as esophageal and two esophageal intubations incorrectly as tracheal. Sensitivity and specificity were thus 93% (CI 90-97%) and 98% (CI 94-99%), respectively. The two false-positive cases were attributed to gaseous distension of the stomach and esophagus from prolonged bag-valve-mask ventilation. In addition to use in postprocedure tube placement confirmation, SCOTI aided the intubation procedure itself in 45 difficult emergency attempts (33%), 26 of which necessitated blind tube passage. We conclude that the SCOTI device has high sensitivity and specificity for tube placement confirmation during tracheal intubation attempts in both emergency and elective settings. It also facilitates tube placement itself during difficult intubations. As such it may be considered an adjunctive device to minimize the potentially fatal complication of esophageal intubation. PMID- 11267811 TI - Left lower quadrant pain of unusual cause. AB - The differential diagnosis of left lower quadrant abdominal pain in an adult man includes, among others, sigmoid diverticulitis; leaking abdominal aortic aneurysm; renal colic; epididymitis; incarcerated hernia; bowel obstruction; regional enteritis; psoas abscess; and in this rare instance, situs inversus with acute appendicitis. We report a case of situs inversus totalis with left-sided appendicitis and a brief review of the literature. There were several subtle indicators of total situs inversus present that were missed by the physicians and surgeons who initially evaluated the patient prior to surgery. Computed tomography scan with contrast, however, revealed the diagnosis immediately, and treatment was successfully initiated. PMID- 11267812 TI - Brain abscess in patients with hereditary hemorrhagic telangiectasia: case report and literature review. AB - Hereditary hemorrhagic telangiectasia (HHT), or Osler-Weber-Rendu disease, affects multiple organ systems. Brain abscess is a potential complication, and this disease carries a high mortality. In the setting of HHT the abscess most likely results from paradoxical septic emboli or bacterial seeding of an ischemic portion of the brain after paradoxical sterile emboli. Brain abscess is the diagnosis that must be ruled out in patients with HHT presenting with new onset neurologic symptoms. The clinician can be misled by seemingly benign and nonspecific symptoms, signs, and laboratory test results. Appropriate diagnostic imaging with computed tomography or magnetic resonance imaging of the head is mandatory. We present a case of brain abscess in a patient with HHT presenting to the Emergency Department. The review of the literature deals with the pathophysiology and manifestations of HHT with particular focus on the pathologic and clinical features, and management of cerebral abscess in this setting. Differences between patients with brain abscess with or without HHT are highlighted. PMID- 11267813 TI - Tourniquet syndrome: a review of constricting band removal. AB - A constricting band causing tourniquet syndrome is a common problem that can cause much frustration and pain for both the patient and practitioner. The following review classifies and describes the different aspects of the treatment of this condition. We also describe a policy and procedure for the motorized removal of large bands by using a high revolutions per minute cutting device. PMID- 11267814 TI - The transport of ciguatoxin: a case report. AB - Ciguatera fish poisoning has been responsible for as many as half of all food poisonings in the United States due to fish. Because the initial symptoms often include gastrointestinal symptoms, such as nausea, vomiting, diarrhea, and abdominal pain, patients may be discharged from the Emergency Department with a diagnosis of "acute gastroenteritis," only to return soon thereafter. This is a case report of such a patient who was evaluated and discharged only to subsequently return because of worsening of symptoms. PMID- 11267815 TI - Metformin-associated lactic acidosis. AB - In 1995, the oral antihyperglycemic agent, metformin, was introduced in the United States for treating diabetes mellitus. Rare cases of metformin-associated lactic acidosis caused by the accumulation of the drug in patients with renal dysfunction have been described, although a detailed time course of the resulting metabolic derangements has not been reported. A case of metformin-associated lactic acidosis is presented along with key serial laboratory abnormalities observed during the treatment phase. The patient made a complete recovery following therapy with hemodialysis and supportive care. PMID- 11267816 TI - Esophageal laceration and charcoal mediastinum complicating gastric lavage. AB - A 19-year-old woman underwent multiple attempts at orogastric lavage before success 5 h after ingesting approximately 24 grams of ibuprofen in a suicide attempt. Activated charcoal was administered via the lavage tube. She vomited charcoal shortly after administration and began experiencing difficulty breathing and an increase in the pitch of her voice. A chest X-ray study showed a widened mediastinum, pneumopericardium, and subcutaneous emphysema consistent with esophageal perforation that was confirmed by computed tomography scan. Surgical exploration revealed a tear in the proximal posterior esophagus with charcoal in the posterior mediastinum. She remained intubated for 7 days and was discharged 14 days after admission. This is a report of esophageal perforation with activated charcoal contamination of the mediastinum after gastric lavage. The risks and benefits of this procedure should be carefully considered in each patient prior to its use. Awake patients should be cooperative with the procedure to minimize any risk of trauma to the oropharynx or esophagus. PMID- 11267817 TI - Preoperative diagnosis of obturator hernia by computed tomography in six patients. AB - Obturator hernia is a rare condition, and the prognosis of patients with this condition is poor. A retrospective study was performed on six patients with obturator hernia between 1993 and 1998. They had been diagnosed preoperatively by computed tomography (CT). The initial CT scan of the abdomen, including the pelvic area, revealed an incarcerated bowel in the obturator foramen of all six patients. All patients underwent laparotomy on the day of admission. Resection of the small bowel was performed in three patients, and release of the small bowel was performed in the remaining three patients. There were no perioperative deaths. In elderly women who have evidence by abdominal plain X-ray studies of small bowel obstruction, we recommend performing CT scan of the abdomen, including CT scan of the pelvic area, for detection of obturator hernia. PMID- 11267818 TI - Efficacy of follow-up evaluation in penetrating thoracic injuries: 3- vs. 6-hour radiographs of the chest. AB - Pneumothorax (PTX) in patients with penetrating thoracic trauma is routinely ruled out with serial chest radiographs (CXRs). This study examined the efficacy of a shortened time period between initial and follow-up radiographs. Patients with penetrating torso injuries treated at a Level-1 trauma center received a CXR during their initial evaluation. If no pneumothorax or hemothorax was noted, and the patient did not require immediate admission to the Intensive Care Unit or operating room, a repeat chest film was taken at 3 and 6 h. Findings were treated as clinically indicated, and patients were discharged home if the last radiograph revealed no evidence of pathology. Over a 15-month period, 116 patients were evaluated for penetrating thoracic injuries (93 stabbings, 23 gunshot wounds) and had no pneumothorax detected on initial CXR. Two patients had pneumothorax detectable only by computed tomography. One patient had a normal initial CXR, but developed a PTX on the 3-h film, requiring tube thoracostomy. No patients developed a PTX on the 6-h study that was not present on the initial or 3-h CXR. In conclusion, extending the time between initial and final CXRs to 6 h in patients with penetrating thoracic trauma provided no additional information that was not available on the 3-h film. PMID- 11267819 TI - Hyperglycemic hyperosmolar nonketotic coma. PMID- 11267820 TI - Pancreatic pseudocysts. PMID- 11267821 TI - Abdominal wall hematoma as a complication of warfarinization. PMID- 11267822 TI - Total metacarpal dislocation. PMID- 11267825 TI - E-mail amplification of a mock code teaching round. AB - To determine whether e-mail could be used to supplement a teaching round, we implemented the following educational intervention: each Monday, a mock code was presented. Two e-mails were then sent the same day to all residents. One summarized the main teaching points whereas the second solicited discussion. Each Friday, a third e-mail was sent that summarized the discussion. We collected all e-mails and surveyed the residents. Fifteen of 18 residents completed the questionnaire; two were not participants in the e-round. Forty percent (7/15) of the residents attended fewer than half of the mock codes but most participants (10/13) reported reading >95% of the e-mails. A majority reported storing (11/13), printing (7/13), and reading e-mails a second time (12/13). Seven of 13 reported learning as much or more from the e-mails as from the mock code itself. We conclude that e-mail can increase learning from a traditional mock code teaching round. PMID- 11267823 TI - Viable pregnancy in the presence of an intrauterine contraceptive device. PMID- 11267824 TI - EM moonlighting: the focus should be on patient safety. PMID- 11267828 TI - Saving lives through community education. PMID- 11267826 TI - Follow-up of discrepancies in X-ray and electrocardiogram interpretations, and positive laboratory results. AB - This article describes the 1-year follow-up program implemented at Baystate Medical Center Emergency Department during 1999. Our previous system used staff who worked clinically, which led to prolonged delays in follow-up. Before initiating the program, 57% [95% confidence interval (CI): 55-59%] of all follow up cases were done within 3 days. After program implementation, 69% (95% CI: 67 72%) of all follow-up cases were completed in the same time frame. We reduced our "delayed" follow-up cases from 20% (95% CI: 18-22%) to 4% (95% CI: 3-5%) of all cases. Critical to the new system is the assignment of nurse-physician pairing to do follow-up when they are not doing direct patient care. PMID- 11267827 TI - AAEM, CORD, and SAEM reach a landmark position: consensus recommendations to the Federation of State Medical Boards (FSMB). American Academy of Emergency Medicine. Council of Emergency Medicine Residency Directors. Society of Academic Medicine. PMID- 11267829 TI - Complementation of chromosomal aberrations in AT/NBS hybrids: inadequacy of RDS as an endpoint in complementation studies with immortal NBS cells. AB - Nijmegen breakage syndrome (NBS) and ataxia telangiectasia (AT) are rare autosomal recessive hereditary disorders characterized by radiosensitivity, chromosomal instability, immunodeficiency and proneness to cancer. Although the clinical features of both syndromes are quite distinct, the cellular characteristics are very similar. Cells from both NBS and AT patients are hypersensitive to ionizing radiation (IR), show elevated levels of chromosomal aberrations and display radioresistant DNA synthesis (RDS). The proteins defective in NBS and AT, NBS1 and ATM, respectively, are involved in the same pathway, but their exact relationship is not yet fully understood. Stumm et al. (Am. J. Hum. Genet. 60 (1997) 1246) have reported that hybrids of AT and NBS lymphoblasts were not complemented for chromosomal aberrations. In contrast, we found that X-ray-induced cell killing as well as chromosomal aberrations were complemented in proliferating NBS-1LBI/AT5BIVA hybrids, comparable to that in NBS 1LBI cells after transfer of a single human chromosome 8 providing the NBS1 gene. RDS observed in AT5BIVA cells was reduced in these hybrids to the level of that seen in immortal NBS-1LBI cells. However, the level of DNA synthesis, following ionizing radiation, in SV40 transformed wild-type cell lines was the same as in NBS-1LBI cells. Only primary wild-type cells showed stronger inhibition of DNA synthesis. In summary, these results clearly indicate that RDS cannot be used as an endpoint in functional complementation studies with immortal NBS-1LBI cells, whereas the cytogenetic assay is suitable for complementation studies with immortal AT and NBS cells. PMID- 11267831 TI - Enhanced ribonucleotide incorporation by an O-helix mutant of Thermus aquaticus DNA polymerase I. AB - The O-helix of DNA polymerases has been implicated in substrate discrimination and replication fidelity. In this study, wild-type Thermus aquaticus DNA polymerase I (Taq pol I) and an O-helix mutant A661E was examined for their ability to discriminate between ribonucleotides and deoxyribonucleotides. Steady state nucleotide extension kinetics were carried out using a template cytidine and each nucleotide dNTP and rGTP. Wild-type Taq pol I and A661E demonstrated similar Vmax and Km values for the correct nucleotide dGTP. However, A661E discriminated between incorrect and correct nucleotide less well than wild-type; discrimination was reduced by factors of 9.5-, 5.6- and 15-fold for dATP, dTTP and rGTP, respectively. These data suggest that A661E is efficient polymerases in the presence of the correct deoxynucleotide, dGTP, but it is impaired in ability to discriminate between correct and incorrect deoxyribonucleotides or between ribo- and deoxyribonucleotides. A structural model of Taq pol I is described in which the mutation A661E alters the interactions between the O-helix and the terminal two phosphate groups in the primer strand. PMID- 11267830 TI - Characterization of the full length uracil-DNA glycosylase in the extreme thermophile Thermotoga maritima. AB - A full length (192 amino acids) uracil-DNA glycosylase (TMUDG) has been expressed and purified from the extreme thermophile Thermotoga maritima. This protein is active up to 85 degrees C. The enzyme is product inhibited by abasic sites in DNA and weakly inhibited by uracil. TMUDG was originally cloned from an ORF which encoded a protein of 185 amino acids. This shorter protein was stable up to 70-75 degrees C and it seemed unusual that this enzyme had an optimal activity temperature below the growth temperature of the organism (80-90 degrees C). Following the publication of the complete genomic sequence of T. maritima, it was shown that the gene contains an additional seven amino acids (LYTREEL) at the N terminal end of the protein. It is suggested that these seven residues are important in maintaining proper protein folding that results in increased temperature stability. We have also demonstrated that TMUDG can substitute for the Escherichia coli uracil-DNA glycosylase and initiate base excision repair using a closed circular DNA substrate containing a unique U:G base pair. PMID- 11267832 TI - The relative contribution of adduct blockage and DNA repair on template utilization during replication of 1,N2-propanodeoxyguanosine and pyrimido. AB - The role of replication blockage by the exocyclic DNA adducts propanodeoxyguanosine (PdG) and pyrimido[1,2-alpha]purin-10(3H)-one (M1G) was determined through the use of site-specifically adducted M13MB102 genomes containing a C:C-mismatch approximately 3000 base-pairs from the site of adduct incorporation. Genomes containing either dG, PdG, or M1G positioned at site 6256 of the (-)-strand were transformed into repair-proficient and repair-deficient Escherichia coli strains and the percent template utilization was determined by hybridization analysis. Unmodified genomes containing a C:C-mismatch resulted in a percent template utilization of approximately 60 and 40% for the (-)- and (+) strands, respectively. Transformation of PdG- or M(1)G-adducted genomes resulted in approximately a 60-40% and 50-50% (-)-strand to (+)-strand ratio, respectively, indicating that PdG and M(1)G are negligible blocks to replication in repair-proficient E. coli. This is in contrast to previous studies using (PdG:T)- and (M1G:T)-mismatched M13MB102 genomes, which resulted in a majority of the replication events using the unadducted (+)-strand and suggested that both adducts were significant blocks to replication [J. Biol. Chem. 272 (1997) 11434; Proc. Natl. Acad. Sci. U.S.A. 94 (1997) 8652]. The C:C-mismatch results, though, indicate that the large strand bias detected in the earlier studies is due to repair of the adducts and resynthesis of the (-)-strand using the (+)-strand as a template for repair synthesis. Transformation of adducted C:C-mismatched genomes into E. coli strains deficient in nucleotide excision repair did result in an increased strand bias with only approximately 20 and 34% of the replication events using the (-)-strand for PdG- and M1G-adducted genomes, respectively. The increased strand bias indicates the importance of nucleotide excision repair in the removal of PdG and M1G. PMID- 11267833 TI - Diversity of the damage recognition step in the global genomic nucleotide excision repair in vitro. AB - The XPC-HR23B complex, a mammalian factor specifically involved in global genomic nucleotide excision repair (NER) has been shown to bind various forms of damaged DNA and initiate DNA repair in cell-free reactions. To characterize the binding specificity of this factor in more detail, a method based on immunoprecipitation was developed to assess the relative affinity of XPC-HR23B for defined lesions on DNA. Here we show that XPC-HR23B preferentially binds to UV-induced (6-4) photoproducts (6-4PPs) as well as to cholesterol, but not to the cyclobutane pyrimidine dimer (CPD), 8-oxoguanine (8-oxo-G), O6-methylguanine (O6-Me-G), or a single mismatch. Human whole cell extracts could efficiently excise 6-4PPs and cholesterol in an XPC-HR23B-dependent manner, but not 8-oxo-G, O6-Me-G or mismatches. Thus, there was good correlation between the binding specificity of XPC-HR23B for certain types of lesion and the ability of human cell extracts to excise these lesions, supporting the model that XPC-HR23B initiates global genomic NER. Although, XPC-HR23B does not preferentially bind to CPDs, the excision of CPDs in human whole cell extracts was found to be absolutely dependent on XPC-HR23B, in agreement with the in vivo observation that CPDs are not removed from the global genome in XP-C mutant cells. These results suggest that, in addition to the excision repair pathway initiated by XPC-HR23B, there exists another sub-pathway for the global genomic NER that still requires XPC HR23B but is not initiated by XPC-HR23B. Possible mechanisms will be discussed. PMID- 11267834 TI - RAD9, RAD24, RAD16 and RAD26 are required for the inducible nucleotide excision repair of UV-induced cyclobutane pyrimidine dimers from the transcribed and non transcribed regions of the Saccharomyces cerevisiae MFA2 gene. AB - In this study, the effect of a prior UV irradiation on the removal of cyclobutane pyrimidine dimers (CPDs) from the transcribed strand (TS) and non-transcribed strand (NTS) of the MFA2 gene in haploid Saccharomyces cerevisiae (S. cerevisiae) cells was investigated. In NER competent cells, the pre-irradiation with a dose of 20J/m2 enhances the removal of CPDs induced by a second UV dose of 100J/m2 in the TS and the NTS of MFA2 gene except for the CPDs in the region +258 to +298 in the NTS, where the enhanced repair was absent. No inducible repair was observed in rad9, rad24, rad16 and rad26 cells, indicating two checkpoint genes RAD9 and RAD24, the global repair gene RAD16 and the transcription coupled repair gene RAD26 are essential for inducible NER. PMID- 11267835 TI - Isolation and genetic characterisation of the Drosophila homologue of (SCE)REV3, encoding the catalytic subunit of DNA polymerase zeta. AB - In Drosophila, about 30 mutants are known that show hypersensitivity to the methylating agent methyl methane sulfonate (MMS). Addition of this agent to the medium results in an increased larval mortality of the mutants. Using a P insertion mutagenesis screen, three MMS-sensitive mutants on chromosome II were isolated. One of these is allelic to the known EMS-induced mus205 (mutagen sensitive) mutant. In the newly isolated mutant, a P-element is detected in region 43E by in situ hybridisation. The localisation of mus205 to this region was confirmed by deficiency mapping. The gene was cloned and shows strong homology to the Saccharomyces cerevisiae REV3 gene. The REV3 gene encodes the catalytic subunit of DNA polymerase zeta, involved in translesion synthesis. The P-element is inserted in the first exon of the mus205 gene resulting in an aberrant mRNA, encoding a putative truncated protein containing only the first 13 of the 2130 aa native Drosophila protein. The mus205 mutant is hypersensitive to alkylating agents and UV, but not to ionising radiation. In contrast to reported data, in germ cells, the mutant has no effect on mutability by X-rays, NQO and alkylating agents. In somatic cells, the mutant shows no effect on MMS-induced mutations and recombinations. This phenotype of the Drosophila mus205 mutant is strikingly different from the phenotype of the yeast rev3 mutant, which is hypomutable after UV, X-rays, NQO and alkylating agents. PMID- 11267836 TI - Repair bias of large loop mismatches during recombination in mammalian cells depends on loop length and structure. AB - Repair of loop mismatches was investigated in wild-type and mismatch binding defective Chinese hamster ovary (CHO) cells. Loop mismatches were formed in vivo during extrachromosomal recombination between heteroallelic plasmid substrates. Recombination was expected to occur primarily by single-strand annealing (SSA), yielding 12- or 26-base nonpalindromic loop mismatches, and 12-, 26-, or 40-base palindromic loop mismatches. Nonpalindromic loops were repaired efficiently and with bias toward loop loss. In contrast, the 12-base palindromic loop was repaired with bias toward loop retention, indicating that repair bias depends on loop structure. Among the palindromic loops, repair bias was dependent on loop length, with bias shifting from loop retention to loop loss with increasing loop size. For both palindromic and nonpalindromic loops, repair efficiencies and biases were independent of the general (MSH/MLH) mismatch repair pathway. These results are discussed with respect to the maintenance of large nonpalindromic insertions, and of small and large palindromes, in eukaryotic genomes. PMID- 11267839 TI - Helicobacter pylori mutagenesis by mariner in vitro transposition. AB - We have developed a method for generating transposon insertion mutants using mariner in vitro mutagenesis. The gene of interest was PCR-amplified and cloned. A kanamycin-marked mariner transposon was randomly inserted into the purified plasmid in an in vitro transposition reaction. After repair and propagation in Escherichia coli, purified mutagenized plasmid was introduced into Helicobacter pylori by natural transformation. Transformants were selected by plating on kanamycin. Mutants were predominantly the result of double homologous recombination, and multiple mutants (with insertions in distinct positions) were often obtained. The site of insertion was determined by PCR or sequencing. We have made mutations in known or potential virulence genes, including ureA, hopZ, and vacA, using kanamycin- and kanamycin/lacZ-marked transposons. Colonies carrying a kanamycin/lacZ transposon appeared blue on medium containing the chromogenic agent X-gal, allowing discrimination of mutant and wild-type H. pylori in mixed competition experiments. PMID- 11267840 TI - Genetic complementation of the urease-negative Helicobacter pylori mutant N6ureB::TnKm. AB - Helicobacter pylori produces urease composed of the structural subunits UreA and UreB. Isogenic mutants produced by shuttle mutagenesis from the wild-type strain N6 are widely used in the literature. We describe the genetic complementation of the mutant N6ureB::TnKm by stable transformation with the vector pHel2 containing the cloned genes ureA and ureB and their specific promoter sequence. The orientation of the cloned insert was found to be crucial for urease expression. The majority of complemented clones functionally expressed urease at higher levels than did N6. Homologous recombination between chromosomal and cloned genes occurred at a frequency of 5%. PMID- 11267841 TI - Reduced intracellular survival of Helicobacter pylori vacA mutants in comparison with their wild-types indicates the role of VacA in pathogenesis. AB - The vacuolating cytotoxin VacA of Helicobacter pylori plays an important but yet unknown role in pathogenesis. We studied the impact of the vacuolating cytotoxin on H. pylori invasion of and survival within AGS cells (human gastric cell line derived from an antral adenocarcinoma). Isogenic vacA and cagA mutants were constructed in a wild-type clinical isolate H. pylori, AF4. An H. pylori VacA deficient mutant, AF4(vacA::kan), was cultured in significantly lower numbers from AGS cells after 24 h incubation with gentamicin added to the culture medium than were the type I wild-type strain AF4 (P<0.03) and an isogenic cagA mutant (P<0.01). Complementation of the AF4 vacA mutant with broth culture supernatant from wild-type AF4 improved the intracellular survival of the vacA mutant. We conclude that H. pylori's vacuolating cytotoxin improves the intracellular survival of H. pylori within AGS cells, suggesting the role of the vacuolating cytotoxin in H. pylori pathogenesis. PMID- 11267842 TI - Helicobacter pylori vacuolating cytotoxin binding to a putative cell surface receptor, heparan sulfate, studied by surface plasmon resonance. AB - The Helicobacter pylori vacuolating cytotoxin or VacA toxin is a major virulence factor in H. pylori infection and type B gastritis. We predicted heparin/heparan sulfate (H/HS) binding properties of the 58-kDa subunit of VacA cytotoxin using bioinformatics tools and showed this by surface plasmon resonance (SPR)-based biosensor studies. Putative H/HS binding peptides were synthesized and binding to HS was shown by SPR in the absence or presence of trifluoroethanol. We found that a recombinant cytotoxin VacA polypeptide binds to surface-immobilized HS and propose that HS might be a receptor/co-receptor for H. pylori VacA cytotoxin. PMID- 11267844 TI - A potential double role of anti-Lewis X antibodies in Helicobacter pylori associated gastroduodenal diseases. AB - In this study, we found Lewis X (Le(x)) determinants on 68% of Helicobacter pylori isolates from patients with chronic gastroduodenal diseases. Anti-Le(x) IgG were detected more frequently in the sera from dyspeptic children and adults (45 and 46%), with or without proved (culture) H. pylori infection, than in the sera from healthy individuals (14% and 25%). In contrast, the prevalence of anti Le(x) IgM was higher in the groups of healthy individuals than in the groups of dyspeptic patients. Moreover, anti-Le(x) monoclonal antibody of IgM class enhanced the uptake of Le(x)(+) but not Le(x)(-) H. pylori isolates by phagocytes. In the sera from some dyspeptic patients, we detected Le(x)-anti Le(x) IgG immune complexes (Le(x) ICs). There was a great difference between children and adults as regards the presence of Le(x) ICs. The immune complexes were found in the sera from nine out of 29 (27%) H. pylori-infected and three out of eight (37%) uninfected adult dyspeptic patients. In comparison, Le(x)-anti Le(x) IgG ICs were detected only for two out of 18 (11%) H. pylori-infected children. Le(x) ICs were not found in the sera from healthy individuals. Our results suggest that anti-Le(x) IgM may play a protective role in H. pylori infections. In contrast, anti-Le(x) IgG and particularly Le(x)-anti-Le(x) IgG ICs might contribute to the pathogenesis of chronic H. pylori infections. PMID- 11267843 TI - Infection with cagA- and vacA-positive and -negative strains of Helicobacter pylori in a mouse model. AB - To study the role of cytotoxin-associated protein (cagA) and vacuolating cytotoxin (vacA) in Helicobacter pylori infection in an experimental murine model, mice were infected with seven strains with different cagA and vacA status. Groups of 10 NMRI mice were challenged and were killed 5 weeks later. In a second study, 20 mice were challenged with a mixture of the same seven strains and killed 1, 3, 15 and 17 weeks post-inoculation. All seven strains were found to colonize the mice for the 5-week experimental period. Animals infected with vacA positive strains, regardless of cagA status, showed an elevation of antibody titers. Two cagA-negative and vacA-positive strains and one cagA- and vacA positive strain were found to 'take over' in the mixed infection as analyzed by the randomly amplified polymorphic DNA-polymerase chain reaction technique and in one mouse stomach we found coexistence of two of the strains. We found no evidence of the different strains colonizing different parts of the stomach. PMID- 11267845 TI - Inducible nitric oxide synthase expression before and after eradication of Helicobacter pylori in different forms of gastritis. AB - An increased expression of inducible nitric oxide synthase (iNOS) has been observed in the inflamed human gastric mucosa as well as in some tumors. This observation suggests a pathobiological role of elevated NO production. The purpose of this study was to compare the immunohistochemical iNOS expression in the different kinds of gastritis before and after the eradication of Helicobacter pylori. We performed iNOS and H. pylori immunohistochemical staining and counted iNOS positive cells. We detected elevated expression of iNOS around sites infected with H. pylori. iNOS expression in chemical gastritis was strongly elevated in mucosal glands. After treatment, we found a significant difference in iNOS expression in patients with classical H. pylori-induced antrum predominant gastritis and in patients with active autoimmune gastritis. In the special case of progressed gastritis with intestinal metaplasia we found persistence of intestinal metaplasia, and we could not find a significant difference in the number of positive iNOS cells before and after treatment. The persistence of IM as a possibly precancerous lesion is probably at least in the antrum a source of continuous iNOS induction even after H. pylori eradication. PMID- 11267847 TI - Helicobacter pylori in familial clusters based on antibody profile. AB - Studies have shown a high prevalence of Helicobacter pylori infection in close communities and that intrafamilial spread during early childhood may be a route of transmission. A total of 72 household members from 21 families were enrolled in this study. Sera from individuals showed 50/72 (69.4%) seropositive for IgG against H. pylori by ELISA. Western blots showed diversity in the protein profiles with molecular masses ranging from approximately 8 to 130 kDa. Cohen's kappa statistical analysis of the blot patterns showed that nine families demonstrated similar profiles (100%), while 4 other families showed varying similarities (17-50%). The results support the hypothesis of intrafamilial transmission of H. pylori. Furthermore, serological studies can be used as an effective approach to determine the familial status in relation to H. pylori infection. PMID- 11267846 TI - Role of virulence factors, cell components and adhesion in Helicobacter pylori mediated iNOS induction in murine macrophages. AB - To investigate the mechanisms involved in Helicobacter pylori-mediated inducible nitric oxide synthase (iNOS) upregulation in mononuclear cells we cocultivated human THP-1 acute monocytic leukemia cells and murine J774A.1 professional macrophages with different H. pylori wild-type strains and mutants. We have shown that H. pylori-mediated iNOS induction in J774A.1 is independent of established virulence factors but dependent on direct interaction between bacteria and cells. In J774A.1, iNOS was equally upregulated by the wild-type strains J99, 26695, P12, and P1 as well as by mutants lacking the cag pathogenicity island, vacA, katA, alpAB genes and the hp0043 gene taking part in lipopolysaccharide biosynthesis when direct cell contact was allowed but not when bacteria and cells were separated by protein-permeable filter membranes. In contrast, iNOS was not induced in THP-1. This indicates that H. pylori-mediated iNOS induction in J774A.1 is independent of important virulence factors whereas cell contact is crucial which suggests a role of adhesion or phagocytosis. PMID- 11267848 TI - Seropositivity to Helicobacter pylori heat shock protein 60 is strongly associated with intensity of chronic inflammation, particularly in antrum mucosa: an extension of an 18-year follow-up study of chronic gastritis in Saaremaa, Estonia. AB - Helicobacter pylori is a cause of chronic gastritis and leads to development of atrophy in some cases. There is evidence that the heat shock protein 60 (HSP60) of H. pylori is involved in induction of chronic inflammation. Seroprevalence of IgG antibodies to H. pylori HSP60 in an adult cohort from Saaremaa, Estonia (68 persons, median age 57 years), with a high prevalence of antibodies to cell surface proteins of H. pylori (92%) and a well characterized dynamics of chronic gastritis in an 18-year follow-up study, was tested using purified H. pylori HSP60 at a concentration of 1 microg ml(-1) with ELISA. The state of the gastric mucosa and the presence of H. pylori in histological sections in the samples of 1979 and 1997 were assessed in accordance with the Sydney system. Seropositivity for H. pylori HSP60 was 65%. Immunological response to H. pylori HSP60 is associated with the morphological presence of H. pylori in the antrum and corpus (P=0.01) and is strongly correlated with the grade of chronic inflammation, particularly in the antrum mucosa (r=0.34; P=0.003; OR=5.97 (95% CI 1.21-29.3)), but is not associated with development of atrophy during 18 years of follow-up, or with the activity of gastritis. This finding supports the evidence that immunological response to H. pylori HSP60 may play a role in triggering of the inflammatory process in the gastric mucosa. PMID- 11267849 TI - Analysis of host responses of guinea pigs during Helicobacter pylori infection. AB - Host responses of guinea pigs infected with Helicobacter pylori were investigated. Passaged H. pylori colonised the stomach for up to 13 weeks after infection, but after 1 month the number of bacteria fell sharply. Specific antibodies, predominantly of the IgG2 subtype, were present from week 3 onwards. Antibodies to urease A and flagella were abundant. Severe inflammation of the gastric mucosa and damage to the stomach epithelium was seen. Infiltrates of mononuclear cells and eosinophils were found near the parietal glands. As infection progressed, inflammation and tissue damage became more localised and more variable between individual animals. These parameters can be used as markers for colonisation of the stomach by H. pylori. PMID- 11267850 TI - Formulations of single or multiple H. pylori antigens with DC Chol adjuvant induce protection by the systemic route in mice. Optimal prophylactic combinations are different from therapeutic ones. AB - The ability to induce a protective response against Helicobacter pylori infection has been investigated by systemic immunization of mice with urease formulated with the cationic lipid DC Chol. This compound acts both as a formulating agent and as an adjuvant and induces a balanced Th1/Th2 response shown to be more effective for protection in our previous studies. Urease-DC Chol induced a significant protection in prophylaxis but not in therapeutic immunization. The protection level was between 1.5 and 2 log reduction of bacterial density measured by quantitative culture compared to unimmunized-infected mice. In parallel, the protective efficacy of other H. pylori antigens formulated in a similar way and administered with DC Chol was tested. These antigens were tested alone or in combination in prophylactic and therapeutic regimens. Some combinations of antigens induced a better prophylactic or therapeutic activity than urease alone (0.5-1.5 log further reduction in prophylaxis and therapy respectively, P<0.05). The combinations that induced the best protection were different in prophylaxis and therapy. In conclusion, DC Chol provides a convenient and efficient method to formulate different antigens even when they are present in non-compatible buffers initially. Moreover, the results obtained in protection against H. pylori with such formulations should lead the way to future clinical trials. PMID- 11267851 TI - Comparative study of Helicobacter pylori infection in guinea pigs and mice - elevation of acute-phase protein C3 in infected guinea pigs. AB - Eighteen Dunkin-Hartley guinea pigs and 50 NMRI mice were inoculated with Helicobacter pylori and the infection followed by culture, histopathology, antibody response, and plasma levels of the acute-phase proteins albumin, C3, and transferrin for up to 7 weeks. The immune response to H. pylori surface proteins was studied by an enzyme immunoassay (EIA) and Western immunoblot and the plasma levels of albumin, C3, and transferrin were analyzed by single radial immunodiffusion. Guinea pigs had a more severe gastritis and a higher EIA immune response than NMRI mice. Serum C3 levels were elevated in infected guinea pigs after 3 and 7 weeks indicating a systemic inflammatory response and a possible link between H. pylori infection and extragastric manifestations such as vasculitis associated with atherosclerosis. Serum cholesterol levels were analyzed in guinea pigs at 7 weeks and indicated a higher level in H. pylori infected than in control animals, but this difference was not statistically significant. PMID- 11267852 TI - Quality control in the locoregional treatment of breast cancer. PMID- 11267853 TI - Sentinel lymph node biopsy as an indicator for axillary dissection in early breast cancer. AB - Sentinel node biopsy (SNB) is a new component of the surgical treatment of breast cancer that accurately predicts axillary status. Although the procedure is still mainly investigational, many patients are requesting SNB to avoid axillary dissection if the sentinel node (SN) is negative. From March 1996 to December 1999, 373 patients with breast carcinoma and clinically negative axillary nodes underwent breast surgery, mainly conservative, and SNB. If the SN was histologically uninvolved no further surgical treatment was given. All patients were informed in detail and signed a consent form. SNB involved injection of labelled albumin particles close to the primary tumour, lymphoscintigraphy and location of the sentinel node with a gamma probe during surgery. 379 SNBs were performed on 373 patients (6 were bilateral). In 94, the SN was positive and complete axillary dissection was performed. In 285 cases (280 patients) the SN was negative and no dissection was performed: these were carefully followed with quarterly clinical examination of the axilla. A total of 343 years at risk were available for evaluation from which seven cases of axillary metastases were expected. No cases of clinically evident axillary node metastasis have occurred. These findings provide further confirmation of the validity of SNB and prompt us to suggest that it should become the method of choice for axillary staging in small-sized breast cancer. PMID- 11267855 TI - Short- and long-term anxiety and depression in women recalled after breast cancer screening. AB - The aim was to investigate the psychological consequences of further investigation after breast cancer screening. Study participants include 509 women (61%) recalled due to suspicious findings on screening mammograms, and a matched control group of 285 women (68%) with normal mammograms. Psychological distress was prospectively assessed with the Hospital Anxiety and Depression Scale (HADS). 46% of the women reported borderline or clinically significant anxiety prior to the recall visit. A few days after the visit, anxiety and depression had decreased significantly (P<0.01) in women informed about normal or benign results at the recall clinic, while reported distress remained at relatively high levels in women referred to surgical biopsy. The results demonstrate the adverse short term effect of a delay in receiving false-positive results, but do not indicate that the recall experience results in long-term anxiety or depression for a majority of women. PMID- 11267854 TI - Mammographic size of ductal carcinoma in situ does not predict the presence of an invasive focus. AB - A proportion of women thought to have ductal carcinoma in situ (DCIS) on mammography and a core biopsy showing DCIS only, in fact have an invasive focus on surgical excision. This study aims to identify the percentage of such patients who harbour an invasive focus and to ascertain features which can predict the presence of invasion. 140 patients had a core biopsy diagnosis of DCIS without invasion. All patients had their core biopsy graded and mammography was performed on 128 patients. Mammographic findings were classified by a radiologist blinded to the surgical findings into normal, mass/distortion or microcalcification. The extent of the microcalcifications was measured. The core biopsies were graded into high, intermediate or low grade DCIS groups. The core biopsy and radiological findings were compared to see if they could predict the presence of invasive disease at surgical excision. Of the 140 patients, 61 (44%) had an invasive focus. 8 (47%) of 17 patients with normal mammography had an invasive focus. 4 (36%) of 11 patients with a mammographic mass had evidence of invasion. Of the 100 patients with mammographic microcalcifications 48 (48%) had an invasive focus. In the 10 patients with low grade DCIS on core biopsy, 3 (30%) had an invasive focus. Comparative studies in patients with intermediate and high grade DCIS, were 7 of 18 (39%) and 51 of 112 (46%), respectively. Thus, 44% of women thought to have DCIS only on preoperative investigation had an invasive focus. In contrast to previous expressed opinions, neither mammography or grade were predictive. We have not identified any factor capable of predicting a higher likelihood of an invasive focus. PMID- 11267856 TI - Prospective randomised study of split-course radiotherapy versus cisplatin plus split-course radiotherapy in inoperable squamous cell carcinoma of the oesophagus. AB - Between 1983 and 1989, 211 patients with inoperable squamous cell carcinoma of the oesophagus were randomised in a study comparing split-course irradiation (two courses of 20 Gy in five fractions of 4 Gy, separated by a rest of 2 weeks) (arm A) and the same split-course irradiation in combination with cisplatin (CDDP) (3 4 days before each of the two courses of radiotherapy, repeated every 3-4 weeks, for a total of six cycles) (arm B). The Cox's regression model with retrospective stratification was used to compare the two arms to correct for the imbalance at randomisation of the T classification. The median overall survival was 7.9 (95% confidence interval (CI) 7.3-9.4) months in arm A and 9.6 (95% CI 8-13.5) months in arm B. The difference in overall survival was only borderline significant (P=0.048) with a reduction of the instantaneous rate of death of 24%. The 1 and 2 year overall survival rate were respectively 29% (95% CI 21-37%) and 15% (95% CI 8-22%) in arm A and 45% (95% CI 36-54%) and 20% (95% CI 13-27%) in arm B; thereafter, the survival curves became similar. The median progression free survival (PFS) was 5.0 (95% CI 4.6-5.7) versus 6.9 (95% CI 5.3-8.7) months (P=0.028) and the median time to local progression was 6.2 (95% CI 5.1-7.6) months versus 10.9 (95% CI 8.1-15.5) months (P=0.018), respectively, in arms A and B. Haematological toxicities were slightly more commonly observed in the combined group (1% versus 6%). This study shows that split-course irradiation in combination with CDDP is very well tolerated and should be preferred to radiotherapy alone. PMID- 11267857 TI - IGFBP-3 in epithelial ovarian carcinoma and its association with clinico pathological features and patient survival. AB - Insulin-like growth factor binding protein-3 (IGFBP-3) regulates the mitogenic and anti-apoptotic actions of insulin-like growth factors (IGFs). To study the role of IGFBP-3 in ovarian cancer progression, we measured IGFBP-3 concentrations in tumour tissues from 147 patients with epithelial ovarian carcinoma and examined its associations with clinicopathological features of disease and patient survival. The average age of the patients was 54.6 years (range 25-88 years) and the median follow-up time was 37 months. IGFBP-3 levels were measured with a commercial immunoassay kit. Low IGFBP-3 levels were significantly associated with unfavourable prognostic features of the disease, including advanced stage (P=0.048), large size of residual tumour (P=0.007), and suboptimal debulking outcome (P=0.007). Low IGFBP-3 levels were also associated with a significantly increased risk for disease progression (RR=1.92; 95% confidence interval (CI) 1.05-3.45; P=0.034), but the association was not sustained when other clinical and pathological variables were adjusted for in the analysis. No significant associations were observed between the IGFBP-3 level and patients' overall survival and response to chemotherapy. Findings of the study indicate that IGFBP-3 may play a role in the progression of epithelial ovarian cancer, but that it has no independent value in predicting either disease prognosis or the response of patients to chemotherapy. PMID- 11267858 TI - Prognostic importance of the soluble plasminogen activator receptor, suPAR, in plasma from rectal cancer patients. AB - Colorectal cancer is one of the most common tumour types with approximately one third of the tumours located within the rectum. Rectal cancer differs somewhat from colon cancer, e.g. regarding the method of operation and the use of preoperative radiotherapy due to a tendency for local tumour recurrence. Proteolytic enzymes have been identified as key molecules in tumour invasion and metastasis, and factors within the urokinase-plasminogen activation (uPA) system have been associated with prognosis in several tumour types, including colorectal cancer. Recently, methods have been developed to analyse the soluble fraction of the plasminogen activator receptor (suPAR) in blood samples. An association between elevated suPAR levels and poor prognosis has recently been demonstrated in colorectal cancer. We have measured suPAR levels in pretreatment plasma samples from 173 rectal cancer patients in order to confirm its prognostic strength in this clinical entity. suPAR levels were determined in ethylenediamine tetraacetic acid (EDTA) plasma by a kinetic enzyme-linked immunosorbent assay (ELISA) and analysed with respect to sex, age, Dukes' stage, tumour differentiation grade and survival. In a univariate analysis, continuous suPAR plasma levels were associated with survival (P<0.001) with shorter survival among patients with high suPAR values. Patients with suPAR values within the upper quartile had significantly shorter survival (hazard ratio (HR) 2.2, 95% confidence interval (CI) 1.3-43.7, P=0.002). In a multivariate Cox analysis, increasing suPAR values predicted shorter survival independent from Dukes' stage and tumour differentiation grade with an adjusted HR of 2.2 per ng/ml suPAR (95% CI 1.2-4.0, P=0.01). This study thus confirms that measurement of suPAR in preoperative plasma samples gives independent prognostic information in rectal cancer patients, higher values being associated with shorter survival. PMID- 11267859 TI - A possible role for methotrexate in the treatment of childhood acute myeloid leukaemia, in particular for acute monocytic leukaemia. AB - Acute myeloid leukaemia (AML) is thought to be methotrexate (MTX)-resistant. However, a small study suggested that acute monocytic leukemia (AML-M5) is sensitive to MTX. We measured MTX accumulation/polyglutamylation in 20 AML-nonM5, 37 AML-M5 and 83 common/preB-acute lymphoblastic leukaemia (c/preB-ALL) samples. Membrane transport was determined in 11 childhood AMLs (including 3 AML-M5) and in 25 c/preB-ALL samples. MTX sensitivity was determined in 23 AML-nonM5, 15 AML M5 and 63 common/preB-ALL samples using the thymidylate synthase (TS) inhibition assay. MTX transport was higher in AML samples compared with c/preB-ALL precluding a transport defect in AML. Accumulation of long-chain polyglutamates MTX-Glu(4-6) was 3-fold lower for AML-nonM5 compared with c/preB-ALL cells (median 268 versus 889 pmol MTX-Glu(4-6)/10(9) cells; P < or = 0.001); for AML-M5 samples, median accumulation of MTX-Glu(4-6) was 0 pmol/10(9) cells (P < or = 0.001). After short-term MTX exposure, AML-nonM5 was 6-fold more resistant to MTX compared with c/preB-ALL cells (2.16 versus 0.39 microM; P < 0.001), while AML-M5 was 2-fold more resistant (P = 0.02). In both AML-nonM5 and AML-M5 cells, MTX resistance was circumvented by continuous MTX exposure (median TSI(50) values: 0.052 and 0.041 microM, respectively) compared with a c/preB-ALL value of 0.066 microM. In conclusion, AML-M5 is relatively sensitive to MTX compared with other AML-subtypes even though polyglutamylation of MTX is poor. Using continuous exposure, AML-nonM5 and AML-M5 cells were at least as sensitive to MTX as c/preB ALL cells. This report suggests that MTX might be an overlooked drug in the treatment of childhood AML. PMID- 11267860 TI - APC I1307K and E1317Q variants are rare or do not occur in Swedish colorectal cancer patients. AB - Recently, a germ line mutation of the APC gene, I1307K, was discovered in a subset of Ashkenazi jews. The mutation involves an amino acid exchange and creates a tract consisting of eight contiguous adenosine residues believed to cause hypermutability in this region. Another germ line missense variant, E1317Q, not restricted to a certain ethnic population, could functionally alter the protein. These APC variants have been linked with increased colorectal cancer risk in several studies. However, they have not yet been investigated in Swedish colorectal cancer patients. Thus, our aim was to investigate the prevalence of I1307K and E1317Q in Swedish colorectal cancer patients in order to determine if these genetic variants are important predisposing factors to colorectal cancer in this population. To this end, sequence analysis was carried out of the APC gene in order to identify any I1307K and E1317Q variants in 106 unselected cases and 88 hereditary/familial colorectal cancer cases including 22 cases of hereditary non-polyposis colorectal cancer (HNPCC) fulfilling the Amsterdam criteria. Out of a total of 194 cases examined, we did not find any variants. It seems that these alterations are rare or absent in the Swedish population. PMID- 11267861 TI - Effects of hexadecylphosphocholine on thrombocytopoiesis. AB - Hexadecylphosphocholine (HePC) is the first representative of the alkylphosphocholines, a novel group derived from the cytotoxic etherlysophospholipids. HePC shows a broad spectrum of antiproliferative effects in neoplastic cells in vitro and in vivo. HePC has been tested successfully in several clinical studies. One of the remarkable features of this compound has been the induction of a leucocytosis and a thrombocytosis in most of the patients receiving HePC systemically. In this paper, we have investigated the biological and molecular mechanisms by which HePC exerts this interesting effect. We found that HePC acts as an unspecific costimulator on human megakaryocytic proliferation in a soft agar assay system predominantly together with thrombopoietin (TPO). Furthermore, HePC leads to the synthesis and secretion of several haematopoietic growth factors in monocytes and bone marrow fibroblasts, determined by the direct measurement of growth factors in cellular supernatants and by the measurement of growth factor mRNA in cell extracts. Thus, HePC seems to produce the increase of blood platelets in tumour patients by two different mechanisms. PMID- 11267862 TI - Downregulation of wild-type beta-catenin expression by interleukin 6 in human hepatocarcinoma HepG2 cells: a possible role in the growth-regulatory effects of the cytokine? AB - We investigated the antitumour effects of interleukin 6 (IL-6) on hepatocarcinoma HepG2 cells, endowed with high levels of a mutated, non-degradable, beta-catenin. IL-6 produced minimal growth-inhibitory effects and no apoptosis or gross changes in cell adhesion. Interestingly, however, it caused a consistent decrease in the cytoplasmic levels of wild-type, but not of mutated, beta-catenin protein. There was no effect on E-cadherin or gamma-catenin and a reduction in alpha-catenin occurred only at high concentrations. IL-4, a non-related cytokine, did not modify the content of beta-catenin. IL-6 did not influence beta-catenin mRNA levels. LiCl, a potent inhibitor of Glycogen Synthase Kinase 3beta (GSK3beta) activity, abrogated the IL-6-induced inhibition of wild-type beta-catenin. This indicates that IL-6 can affect wild-type beta-catenin through a post transcriptional mechanism, probably involving degradation of the protein. This effect might be related to the growth-regulatory activities of IL-6 in other situations, but can not counteract the oncogenic expression of mutated beta catenin in HepG2 cells or possibly in other tumour cells with similar gene mutations. PMID- 11267863 TI - Effects of a novel synthetic retinoid on malignant glioma in vitro: inhibition of cell proliferation, induction of apoptosis and differentiation. AB - Among six synthetic retinoids tested, the retinoid 6-[3-(1-adamantyl)-4 hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) was highly efficient in inducing growth inhibition of 8MG-BA and GL-15 human glioblastoma cell lines, with growth arrest at the S phase of the cell cycle. CD 437 also induced apoptosis in these cells, with 8MG-BA being the most sensitive. In these cells, induction of apoptosis by CD437 has been related to the downregulation of Bcl-2 expression and to CPP32 activation, but not to p53 expression. The remaining non apoptotic cells presented a morphological pattern of astroglial differentiation with overexpression of glial fibrillary acidic protein (GFAP) and glutamine synthetase (GS). The mechanism of action of CD437, originally developed as a RARgamma agonist, is not yet elucidated. However, our results suggest that it acts through an increase of the expression of retinoid-inducible genes, such as RARbeta2 and/or RARalpha2. PMID- 11267864 TI - Combination effect of adenovirus-mediated pro-apoptotic bax gene transfer with cisplatin or paclitaxel treatment in ovarian cancer cell lines. AB - To develop a novel therapeutic strategy for ovarian cancer, we constructed a recombinant adenovirus which highly expresses pro-apoptotic Bax protein and examined its therapeutic effect on a series of ovarian cancer cell lines: A2780, A2780/cDDP, OVCAR-3 and SK-OV-3. A recombinant adenovirus carrying the Bax-alpha gene (AxCALNKYbax) induced high expression of the Bax-alpha protein in all the cell lines. The cytotoxic effect of Bax was observed in three ovarian cancer cell lines: the per cent reduction in the number of cells was 40.0% for cisplatin sensitive A2780, 50.0% for cisplatin-resistant A2780/cDDP, and 64.8% for marginally cisplatin-resistant OVCAR-3. In contrast, it was only 12.3% for cisplatin-resistant SK-OV-3. Cisplatin-resistant A2780/cDDP had a p53 mutation and exhibited attenuated Bax induction after cisplatin treatment, which may explain why supplementation of Bax was effective in this chemoresistant ovarian cancer. Combination with cisplatin or paclitaxel enhanced the cytotoxic effect of Bax induction in all but one cell line including cisplatin-resistant A2780/cDDP. It appears that adenovirus-mediated Bax induction, with or without combination with conventional chemotherapy, useful strategy for the treatment of ovarian cancer. PMID- 11267865 TI - Identification of colon tumour-associated antigens by phage antibody selections on primary colorectal carcinoma. AB - Immunotargeting of solid tumours using antibodies has become a valuable tool for the detection of cancer metastases and the treatment of minimal residual disease. However, only few tumour antigens useful for targeting have been described to date. To identify cell-surface targets on colorectal carcinoma (CRC), we selected a large, human phage antibody repertoire on freshly isolated colon tumour cells. Two antibodies were identified that reacted with epithelial cell-restricted cell surface antigens, whereas one clone preferentially reacted with stromal cells. These antigens are tumour-associated antigens, as shown by their uniform expression in tumours of different patients and of different differentiation stages and by their limited expression on normal tissues. The pattern of reactivity in immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA) suggests that these antigens are different from previously identified tumour-associated antigens (e.g. Ep-CAM or c-ERB-2). This phage antibody-based method may lead to the cloning of novel tumour antigens that are useful for the immunotargeting of solid tumours. PMID- 11267866 TI - A role for Drosophila SMC4 in the resolution of sister chromatids in mitosis. AB - BACKGROUND: Faithful segregation of the genome during mitosis requires interphase chromatin to be condensed into well-defined chromosomes. Chromosome condensation involves a multiprotein complex known as condensin that associates with chromatin early in prophase. Until now, genetic analysis of SMC subunits of the condensin complex in higher eukaryotic cells has not been performed, and consequently the detailed contribution of different subunits to the formation of mitotic chromosome morphology is poorly understood. RESULTS: We show that the SMC4 subunit of condensin is encoded by the essential gluon locus in Drosophila. DmSMC4 contains all the conserved domains present in other members of the structural-maintenance-of-chromosomes protein family. DmSMC4 is both nuclear and cytoplasmic during interphase, concentrates on chromatin during prophase, and localizes to the axial chromosome core at metaphase and anaphase. During decondensation in telophase, most of the DmSMC4 leaves the chromosomes. An examination of gluon mutations indicates that SMC4 is required for chromosome condensation and segregation during different developmental stages. A detailed analysis of mitotic chromosome structure in mutant cells indicates that although the longitudinal axis can be shortened normally, sister chromatid resolution is strikingly disrupted. This phenotype then leads to severe chromosome segregation defects, chromosome breakage, and apoptosis. CONCLUSIONS: Our results demonstrate that SMC4 is critically important for the resolution of sister chromatids during mitosis prior to anaphase onset. PMID- 11267867 TI - Kinetics and regulation of de novo centriole assembly. Implications for the mechanism of centriole duplication. AB - BACKGROUND: Centriole duplication is a key step in the cell cycle whose mechanism is completely unknown. Why new centrioles always form next to preexisting ones is a fundamental question. The simplest model is that preexisting centrioles nucleate the assembly of new centrioles, and that although centrioles can in some cases form de novo without this nucleation, the de novo assembly mechanism should be too slow to compete with normal duplication in order to maintain fidelity of centriole duplication. RESULTS: We have measured the rate of de novo centriole assembly in vegetatively dividing cells that normally always contain centrioles. By using mutants of Chlamydomonas that are defective in centriole segregation, we obtained viable centrioleless cells that continue to divide, and find that within a single generation, 50% of these cells reacquire new centrioles by de novo assembly. This suggests that the rate of de novo assembly is approximately half the rate of templated duplication. A mutation in the VFL3 gene causes a complete loss of the templated assembly pathway without eliminating de novo assembly. A mutation in the centrin gene also reduced the rate of templated assembly. CONCLUSIONS: These results suggest that there are two pathways for centriole assembly, namely a templated pathway that requires preexisting centrioles to nucleate new centriole assembly, and a de novo assembly pathway that is normally turned off when centrioles are present. PMID- 11267868 TI - A role for myosin VII in dynamic cell adhesion. AB - BACKGROUND: The initial stages of phagocytosis and cell motility resemble each other. The extension of a pseudopod at the leading edge of a migratory cell and the formation of a phagocytic cup are actin dependent, and each rely on the plasma membrane adhering to a surface during dynamic extension. RESULTS: A myosin VII null mutant exhibited a drastic loss of adhesion to particles, consistent with the extent of an observed decrease in particle uptake. Additionally, cell cell adhesion and the adhesion of the leading edge to the substratum during cell migration were defective in the myosin VII null cells. GFP-myosin VII rescued the phagocytosis defect of the null mutant and was distributed in the cytosol and recruited to the cortical cytoskeleton, where it appeared to be enriched at the tips of filopods. It was also localized to phagocytic cups, but only during the initial stages of particle engulfment. During migration, GFP-myosin VII is found at the leading edge of the cell. CONCLUSIONS: Myosin VII plays an important role in mediating the initial binding of cells to substrata, a novel role for an unconventional myosin. PMID- 11267869 TI - Proneural enhancement by Notch overcomes Suppressor-of-Hairless repressor function in the developing Drosophila eye. AB - BACKGROUND: The receptor protein Notch plays a conserved role in restricting neural-fate specification during lateral inhibition. Lateral inhibition requires the Notch intracellular domain to coactivate Su(H)-mediated transcription of the Enhancer-of-split Complex. During Drosophila eye development, Notch plays an additional role in promoting neural fate independently of Su(H) and E(spl)-C, and this finding suggests an alternative mechanism of Notch signal transduction. RESULTS: We used genetic mosaics to analyze the proneural enhancement pathway. As in lateral inhibition, the metalloprotease Kuzbanian, the EGF repeat 12 region of the Notch extracellular domain, Presenilin, and the Notch intracellular domain were required. By contrast, proneural enhancement became constitutive in the absence of Su(H), and this led to premature differentiation and upregulation of the Atonal and Senseless proteins. Ectopic Notch signaling by Delta expression ahead of the morphogenetic furrow also caused premature differentiation. CONCLUSIONS: Proneural enhancement and lateral inhibition use similar ligand binding and receptor processing but differ in the nuclear role of Su(H). Prior to Notch signaling, Su(H) represses neural development directly, not indirectly through E(spl)-C. During proneural enhancement, the Notch intracellular domain overcomes the repression of neural differentiation. Later, lateral inhibition restores the repression of neural development by a different mechanism, requiring E(spl)-C transcription. Thus, Notch restricts neurogenesis temporally to a narrow time interval between two modes of repression. PMID- 11267870 TI - Plexin-B semaphorin receptors interact directly with active Rac and regulate the actin cytoskeleton by activating Rho. AB - Semaphorins and their receptors, plexins, are widely expressed in embryonic and adult tissues. In general, their functions are poorly characterized, but in neurons they provide essential attractive and repulsive cues that are necessary for axon guidance [1-3]. The Rho family GTPases Rho, Rac, and Cdc42 control signal transduction pathways that link plasma membrane receptors to the actin cytoskeleton and thus regulate many actin-driven processes, including cell migration and axon guidance [4-7]. Using yeast two-hybrid screening and in vitro interaction assays, we show that Rac in its active, GTP bound state interacts directly with the cytoplasmic domain of mammalian and Drosophila B plexins. Plexin-B1 clustering in fibroblasts does not cause the formation of lamellipodia, which suggests that Rac is not activated. Instead, it results in the assembly of actin:myosin filaments and cell contraction, which indicates Rho activation. Surprisingly, these cytoskeletal changes are both Rac and Rho dependent. Clustering of a mutant plexin, lacking the Rac binding region, induced similar cytoskeletal changes, and this finding indicates that the physical interaction of plexin-B1 with Rac is not required for Rho activation. Our findings that plexin-B signaling to the cytoskeleton is both Rac and Rho dependent form a starting point for unraveling the mechanism by which semaphorins and plexins control axon guidance and cell migration. PMID- 11267871 TI - Asymmetric spindle pole localization of yeast Cdc15 kinase links mitotic exit and cytokinesis. AB - The inactivation of mitotic cyclin-dependent kinases (CDKs) during anaphase is a prerequisite for the completion of nuclear division and the onset of cytokinesis [1, 2]. In the budding yeast Saccharomyces cerevisiae, the essential protein kinase Cdc15 [3] together with other proteins of the mitotic exit network (Tem1, Lte1, Cdc5, and Dbf2/Dbf20 [4-7]) activates Cdc14 phosphatase, which triggers cyclin degradation and the accumulation of the CDK inhibitor Sic1 [8]. However, it is still unclear how CDK inactivation promotes cytokinesis. Here, we analyze the properties of Cdc15 kinase during mitotic exit. We found that Cdc15 localized to the spindle pole body (SPB) in a unique pattern. Cdc15 was present at the SPB of the mother cell until late mitosis, when it also associated with the daughter pole. High CDK activity inhibited this association, while dephosphorylation of Cdc15 by Cdc14 phosphatase enabled it. The analysis of Cdc15 derivatives indicated that SPB localization was specifically required for cytokinesis but not for mitotic exit. These results show that Cdc15 has two separate functions during the cell cycle. First, it is required for the activation of Cdc14. CD14, in turn, promotes CDK inactivation and also dephosphorylates of Cdc15. As a consequence, Cdc15 binds to the daughter pole and triggers cytokinesis. Thus, Cdc15 helps to coordinate mitotic exit and cytokinesis. PMID- 11267872 TI - Evidence for a satellite secretory pathway in neuronal dendritic spines. AB - Long-term information storage within the brain requires the synthesis of new proteins and their use in synapse-specific modifications [1]. Recently, we demonstrated that translation sites for the local synthesis of integral membrane and secretory proteins occur within distal dendritic spines [2]. It remains unresolved, however, whether a complete secretory pathway, including Golgi and trans Golgi network-like membranes, exists near synapses for the local transport and processing of newly synthesized proteins. Here, we report evidence of a satellite secretory pathway in distal dendritic spines and distal dendrites of the mammalian brain. Membranes analogous to early (RER and ERGIC), middle (Golgi cisternae), and late (TGN) secretory pathway compartments are present within dendritic spines and in distal dendrites. Local synthesis, processing, and transport of newly translated integral membrane and secretory proteins may thus provide the molecular basis for synapse-specific modifications during long-term information storage in the brain. PMID- 11267873 TI - Serum-activated assembly and membrane translocation of an endogenous Rac1:effector complex. AB - Rho family GTPases (Cdc42, Rac1, and RhoA) function downstream of Ras [1], and in a variety of cellular processes [2]. Studies to examine these functions have not directly linked endogenous protein interactions with specific in vivo functions of Rho GTPases. Here, we show that endogenous Rac1 and two known binding partners, Rho GDP dissociation inhibitor (RhoGDI) and p21-activated kinase (PAK), fractionate as distinct cytosolic complexes. A Rac1:PAK complex is translocated from the cytosol to ruffling membranes upon cell activation by serum. Overexpression of dominant-negative (T17N) Rac1 does not affect the assembly or distribution of this Rac1:PAK complex. This is the first direct evidence of how a specific function of Rac1 is selected by the assembly and membrane translocation of a distinct Rac1:effector complex. PMID- 11267874 TI - Mitochondria are selectively eliminated from eukaryotic cells after blockade of caspases during apoptosis. AB - Pan caspase inhibitors are potentially powerful cell-protective agents that block apoptosis in response to a wide variety of insults that cause tissue degeneration. In many conditions, however, the blockade of apoptosis by caspase inhibitors does not permit long-term cell survival, but the reasons are not entirely clear. Here we show that the blockade of apoptosis by Boc.Aspartyl(O methyl)CH2F can result in the highly selective elimination of the entire cohort of mitochondria, including mitochondrial DNA, from both neurons and HeLa cells, irrespective of the stimulus used to trigger apoptosis. In cells that lose their mitochondria, the nuclear DNA, Golgi apparatus, endoplasmic reticulum, centrioles, and plasma membrane remain undamaged. The capacity to remove mitochondria is both specific and regulated since mitochondrial loss in neurons is completely prevented by the expression of the antiapoptotic protein Bcl-2 and partially suppressed by the autolysosomal inhibitor bafilomycin. Cells without mitochondria are more tolerant to an anaerobic environment but are essentially irreversibly committed to death. Prevention of mitochondrial loss may be crucial for the long-term regeneration of tissues emerging from an apoptotic episode in which death was prevented by caspase blockade. PMID- 11267875 TI - Bidirectional amyloid fiber growth for a yeast prion determinant. AB - The polymerization of many amyloids is a two-stage process initiated by the formation of a seeding nucleus or protofibril. Soluble protein then assembles with these nuclei to form amyloid fibers. Whether fiber growth is bidirectional or unidirectional has been determined for two amyloids. In these cases, bidirectional growth was established by time lapse atomic-force microscopy. Here, we investigated the growth of amyloid fibers formed by NM, the prion-determining region of the yeast protein Sup35p. The conformational changes in NM that lead to amyloid formation in vitro serve as a model for the self-perpetuating conformational changes in Sup35p that allow this protein to serve as an epigenetic element of inheritance in vivo. To assess the directionality of fiber growth, we genetically engineered a mutant of NM so that it contained an accessible cysteine residue that was easily labeled after fiber formation. The mutant protein assembled in vitro with kinetics indistinguishable from those of the wild-type protein and propagated the heritable genetic trait [PSI(+)] with the same fidelity. In reactions nucleated with prelabeled fibers, unlabeled protein assembled at both ends. Thus, NM fiber growth is bidirectional. PMID- 11267877 TI - Checks and balances needed for organ retention. PMID- 11267878 TI - Raising the priorities of disease prevention. PMID- 11267876 TI - Cortactin promotes and stabilizes Arp2/3-induced actin filament network formation. AB - Cortactin is a c-src substrate associated with sites of dynamic actin assembly at the leading edge of migrating cells. We previously showed that cortactin binds to Arp2/3 complex, the essential molecular machine for nucleating actin filament assembly. In this study, we demonstrate that cortactin activates Arp2/3 complex based on direct visualization of filament networks and pyrene actin assays. Strikingly, cortactin potently inhibited the debranching of filament networks. When cortactin was added in combination with the active VCA fragment of N-WASp, they synergistically enhanced Arp2/3-induced actin filament branching. The N terminal acidic and F-actin binding domains of cortactin were both necessary to activate Arp2/3 complex. These results support a model in which cortactin modulates actin filament dendritic nucleation by two mechanisms, (1) direct activation of Arp2/3 complex and (2) stabilization of newly generated filament branch points. By these mechanisms, cortactin may promote the formation and stabilization of the actin network that drives protrusion at the leading edge of migrating cells. PMID- 11267879 TI - How apples may lose their pips. PMID- 11267880 TI - Computational genomics. AB - We now know how to read the sequences of nucleotide letters that comprise the genome at a rather frightening speed--a several-million-base bacterial genome in several days is not a problem for one of the sequencing centers, and a billion base eukaryotic genome can be done in less than a year. But reading a text and understanding it are two different things. So how well can we understand the genome sequences? The answer to this question is central to the whole enterprise of genomics, and this is where computational analysis of genomes takes the driver's seat. Here I will try to briefly outline some major goals, problems, challenges and approaches of computational genomics. Such a young field is already quite diverse, and in this short article I will concentrate on several issues that seen to be critical for deciphering biology from genome sequences, rather than mathematical and computer-science aspects that are well covered in several excellent books. PMID- 11267881 TI - Centrosomes: Central no more? AB - It has recently been found that the zygotic development of a morphologically normal fly can occur without properly functioning mitotic centrosomes. Does this mean that centrosomes are not required for cell division in animals at all? PMID- 11267882 TI - Oogenesis: Setting one sister above the rest. AB - Recent studies have revealed that the Par-1 protein, in addition to its established role in anterior-posterior patterning of the Drosophila oocyte, has both microtubule-dependent and microtubule-independent roles very early in oogenesis. PMID- 11267883 TI - RNA localization: SHEdding light on the RNA-motor linkage. AB - Specific mRNAs achieve an asymmetric distribution in the cell by linking to molecular motors that walk along the cytoskeleton. Studies in S. cerevisiae have begun to define the nature of the RNA-motor linkage and identify She3p as an adaptor protein that links a type V myosin motor to specific ribonucleoproteins. PMID- 11267884 TI - Plant physiology: The importance of sucrose transporters. AB - Sucrose transport is essential for the distribution of carbohydrates in plants. Recent studies have shown that a specific transporter protein plays an essential role in loading sucrose into the phloem component of the plant vasculature. PMID- 11267885 TI - Inositol phosphates: Does IP(4) run a protection racket? AB - The phosphorylation of IP(3) by IP(3) 3-kinase leads to a number of physiological events, most of which are poorly understood. Recent findings about a hitherto unsuspected action of the IP(3) 3-kinase product, IP(4), suggest that the evolution of IP(3) 3-kinase may have even more far-reaching consequences than we thought. PMID- 11267886 TI - Cytokines: IL-21 joins the gamma(c)-dependent network? AB - The discovery of the cytokine IL-21 adds another member to the ever-growing list of small secreted molecules that have potent effects on lymphoid cells. Initial characterization of the IL-21 receptor complex suggests that IL-21 may belong to the cytokine family whose receptors share the common gamma chain, gamma(c). PMID- 11267887 TI - Cell cycle: Flies teach an old dogma new tricks. AB - E2F transcription factors are thought to influence the G1-S cell-cycle transition by controlling expression of genes required for growth and DNA synthesis. But emerging evidence suggests E2F complexes can control the cell cycle independently of transcription by directly regulating DNA replication origin usage during S phase. PMID- 11267888 TI - Genome analysis: More Drosophila Y chromosome genes. AB - The Drosophila melanogaster Y chromosome has long been known to contain few functional genes other than several required for male fertility. The D. melanogaster genome sequence has now allowed characterization of two more male fertility genes, shedding light on the function and evolution of Y chromosomes. PMID- 11267889 TI - Chromatin remodeling enzymes: who's on first? AB - A central problem in the regulation of eukaryotic gene expression is understanding how gene-specific transcriptional activators orchestrate the recruitment of the myriad proteins that are required for transcription initiation. An emerging view indicates that activators must first target two types of chromatin remodeling enzyme to the promoter region: an ATP-dependent SWI/SNF-like complex and a histone acetyltransferase. These two enzymes appear to act synergistically to establish a local chromatin structure that is permissive for subsequent events. Furthermore, several recent studies indicate that the recruitment of chromatin remodeling enzymes must follow an obligatory, sequential order of events that is determined by either promoter context or cell-cycle position. Here we review recent developments concerning the role of chromatin remodeling enzymes in gene regulation, and propose several models to explain how different chromatin remodeling activities can be functionally coupled. PMID- 11267890 TI - Identification of a juvenile hormone esterase gene by matching its peptide mass fingerprint with a sequence from the Drosophila genome project. AB - Juvenile hormone esterase (JHE, EC 3.1.1.1) from whole Drosophila melanogaster prepupae has previously been purified by selective precipitations, isoelectric focussing and two column chromatography steps. JHE bands from dried silver stained SDS-PAGE gels of that material were digested with trypsin. The masses of the tryptic digest peptides were determined by MALDI-TOF mass spectrometry. Only one predicted gene product (CG8425) from the D. melanogaster genome matches the JHE tryptic fingerprint with high confidence. This predicted JHE sequence includes features that are conserved among all active members of the serine carboxylesterase multigene family as well as features peculiar to JHEs from other species. Also we show that this JHE can be purified by an alternative method using anion exchange chromotography followed by trifluoromethylketone affinity chromatography. A cDNA encoding this JHE was isolated using 3' and 5' RACE. This sequence is in agreement with the Drosophila genome project's prediction except that the sixth predicted intron is not removed; instead there is a stop codon followed by a polyadenylation signal and a polyA tail. PMID- 11267891 TI - Cloning and functional expression of a chitinase cDNA from the common cutworm, Spodoptera litura, using a recombinant baculovirus lacking the virus-encoded chitinase gene. AB - A Chitinase cDNA named Slchi was cloned from the epidermis of the common cutworm, Spodoptera litura, and the enzymatic properties of its recombinant proteins were characterized. The Slchi cDNA encodes 552 amino-acid residues (aa) including a 19 aa putative signal peptide, with the calculated molecular mass of the putative mature protein 60,152 Da. A major transcript of Slchi about 2.8 kb was detected in the epidermis only during molting in the last instar larvae, suggesting its involvement in the digestive system for old cuticle. The E. coli-produced recombinant Slchi exhibited weak chitinolytic activity against 4MU (GlcNAc)(3)>4MU-(GlcNAc)(2)>4MU-(GlcNAc)(4), in this order, but not against 4MU (GlcNAc)(1). A recombinant Slchi with higher specific activity was obtained using recombinant Hyphantria cunea NPV (HycuNPV), which expresses Slchi under polyhedrin promoter. To discriminate chitinase activity of recombinant Slchi from an active chitinase encoded in HycuNPV genome (chiA), we further knocked out the chiA gene from the recombinant virus. The recombinant Slchi expressed in insect cell culture showed a similar substrate specificity against 4MU-(GlcNAc)(n) (n=1 4) to that produced in E. coli, while the viral chitinase showed the highest activity against 4MU-(GlcNAc)(2). The recombinant Slchi was secreted rapidly into the culture medium from the infected cells, whereas the viral chitinase retained predominantly in the cells. PMID- 11267892 TI - [3H]-Methyllycaconitine: a high affinity radioligand that labels invertebrate nicotinic acetylcholine receptors. AB - Nicotinic acetylcholine receptors (nAChR) of insect and other invertebrates are heterogeneous and new tools are needed to dissect their multiplicity. [(3)H] Methyllycaconitine ([(3)H]-MLA) is a novel radioligand which is a potent antagonist at vertebrate alpha7-type nAChR. Putative invertebrate nAChR of the aphid Myzus persicae, the moths Heliothis virescens and Manduca sexta, the fly Lucilia sericata, and the squid Loligo vulgaris were investigated in radioligand binding studies with [(3)H]-MLA. Saturable binding was consistent with a single class of high affinity binding sites for each of these invertebrates, characterised by a dissociation constant, K(d), of approximately 1 nM and maximal binding capacities, B(max), between 749 and 1689 fmol/mg protein for the insects and 14,111 fmol/mg protein for squid. [(3)H]-MLA binding to M. persicae membranes was characterised in more detail. Kinetic analysis demonstrated rapid association in a biphasic manner and slow, monophasic dissociation. Displacement studies demonstrate the nicotinic character of [(3)H]-MLA binding sites. Data for all nicotinic ligands, except MLA itself, are consistent with displacement from a high and a low affinity site, indicating that displacement is occurring from two or more classes of nicotinic binding site that are not distinguished by MLA itself. Autoradiographic analysis of the distribution of [(3)H]-MLA binding sites in Manduca sexta shows discrete labelling of neuropil areas of the optic and antennal lobes. PMID- 11267893 TI - Identification and characterization of the major Drosophila melanogaster mating plug protein. AB - In many insects, semen coagulates into a mating plug at the distal part of the female's genital tract. Mating plugs have been proposed to facilitate sperm movement or to prevent subsequent matings or sperm loss. The molecular constituents of insect mating plugs have not previously been characterized. Here we report that an abundant autofluorescent protein made by the Drosophila melanogaster male's ejaculatory bulb is a major constituent of the posterior region of the mating plug. Identities in size, chromosomal location and expression pattern indicate that the autofluorescent protein is PEB-me, an abundant ejaculatory bulb protein reported by Ludwig et al. [Biochem. Genet. 29 (1991) 215]. We cloned and sequenced the RNA encoding this protein. The transcript, which is male-specific and expressed only in the ejaculatory bulb, encodes a 377 a.a. predicted secreted protein with PGG repeats similar to those in homopolymer-forming proteins found in spider silk. PMID- 11267894 TI - Cloning and sequence of the gene encoding the muscle fatty acid binding protein from the desert locust, Schistocerca gregaria. AB - Muscle fatty acid binding protein (FABP) is a major cytosolic protein in flight muscle of the desert locust, Schistocerca gregaria. FABP expression varies greatly during development and periods of increased fatty acid utilization, but the molecular mechanisms that regulate its expression are not known. In this study, the gene coding for locust muscle FABP was amplified by PCR and cloned, together with 1.2 kb of upstream sequence. The sequence coding for the 607 bp cDNA is interrupted by two introns of 12.7 and 2.9 kb, inserted in analogous positions as the first and third intron of the mammalian homologues. Both introns contain repetitive sequences also found in other locust genes, and the second intron contains a GT-microsatellite. The promoter sequence includes a canonical TATA box 24 bp upstream of the transcription start site. The upstream sequence contains various potential myocyte enhancer sequences and a 160 bp segment that is repeated three times. In database searches in the genome database of Drosophila melanogaster, a gene with the same gene organization and promoter structure was identified, likely the dipteran homologue of muscle FABP. Upstream of both insect genes, a conserved 19 bp inverted repeat sequence was detected. A similar but reverse palindrome is also present upstream of all mammalian heart FABP genes, possibly representing a novel element involved in muscle FABP expression. PMID- 11267895 TI - Purification and characterization of a lipid transfer particle in Rhodnius prolixus: phospholipid transfer. AB - In this study we report the purification and characterization of a lipid transfer particle (LTP) from Rhodnius prolixus hemolymph, and its participation in phospholipid and diacylglycerol transfer processes. (3)H-diacylglycerol labeled low density lipophorin from Manduca sexta ((3)H-LDLp) was incubated with R. prolixus lipophorin (Lp) in the presence of Rhodnius hemolymph. Following incubation and isolation, both lipoproteins showed equivalent amounts of (3)H labeled lipids. Hemolymph was subjected to KBr gradient ultracentrifugation. SDS PAGE analysis of gradient fractions showed the enrichment of bands with molecular masses similar to the M. sexta LTP standard. LTP containing fractions were assayed and lipid transfer activity was observed. Purification of LTP was accomplished by (i) KBr density gradient ultracentrifugation, (ii) size exclusion, (iii) Cu(++) affinity and (iv) ion exchange chromatographies. LTP molecular mass was estimated approximately 770 kDa, comprising three apoproteins, apoLTP-I (315 kDa), apoLTP-II (85 kDa) and apoLTP-III (58 kDa). Phospolipid content of (32)P-LTP was determined after two-dimensional TLC. (32)P-phospholipid labeled and unlabeled lipophorins, purified from R. prolixus were incubated in the presence of LTP resulting in the time-dependent transfer of phospholipids. LTP-mediated phospholipid transfer was not a selective process. PMID- 11267896 TI - Purification and characterization of a plasmin-like protease from Tenodera sinensis (Chinese mantis). AB - A novel type of protease (mantis egg fibrinolytic enzyme, MEF-2) was isolated from the egg cases of Tenodera sinensis. The protease was homogeneous by SDS-PAGE and its apparent molecular mass was 32,900 Da. The amino acids in the N-terminal region were Ile-Val-Gly-Gly-Glu-Glu-Ala-Val-Ala-Gly-Asp-Phe-Pro-Ile-Val-Ser-Leu Gln-Glu. The enzyme was inhibited by PMSF, TLCK, aprotinin, benzamidine, soybean trypsin inhibitor and also slightly by elastatinal, EDTA, EGTA, cysteine and beta mercaptoethanol, but TPCK, iodoacetate and E-64 did not affect the activity. MEF 2 was not sensitive to alpha(1)-antitrypsin but antithrombin III and alpha(2) antiplasmin inhibited the enzyme. MEF-2 preferentially cleaved the oxidized B chain of insulin between Arg(22) and Gly(23). Among chromogenic protease substrates, the most susceptible to MEF-2 hydrolysis was benzoyl-Phe-Val-Arg-p nitroanilide with maximal activity at 30 degrees C and pH 5.0. These results indicate that MEF-2 belongs to the trypsin family. Upon incubation of crosslinked fibrin with MEF-2, a steady increase of D-dimer suggests that the enzyme has a strong fibrinolytic activity. In conclusion, MEF-2 is a new type of proteolytic enzyme and has some potential for practical application in fibrinolysis. PMID- 11267897 TI - Iridoid biosynthesis in staphylinid rove beetles (Coleoptera: Staphylinidae, Philonthinae). AB - The biosynthesis of chrysomelidial and plagiodial was studied in the rove beetle subtribe Philonthina (Staphylinidae). Glandular homogenates were found to convert synthetic (2E,6E)-[trideuteromethyl-5,5-(2)H(5)]octa-2,6-diene-1,8-diol (10) into nor-chrysomelidial (14) and nor-plagiodial (13). The overall transformation requires; i) oxidation of the substrate at C(1) and C(8), ii) cyclization of the resulting dialdehyde to nor-plagiodial followed by iii) isomerization to give nor chrysomelidial. The oxidase requires molecular oxygen as a cofactor and operates with removal of the pro-R hydrogen from C(1) and C(8) of synthetic (1R,8R,2E,6E) [1,8-(2)H(2)]-2,6-dimethyl-octa-2,6-diene-1,8-diol (15), producing a dialdehyde along with H(2)O(2). Unlike enzymes from iridoid-producing leaf beetle larvae, the Philonthus enzyme is able to oxidize saturated substrates such as citronellol. Crude protein extracts prepared from Philonthus glands by ammonium sulfate precipitation, were found to produce hydrogen peroxide at a rate of 0.085+/-0.003 ng H(2)O(2) (ng protein)(-1) hr(-1) with nerol as an oxidase substrate. The cyclase operates with opposite stereochemistry to the enzyme(s) from Phaedon cochleariae and other herbivorous leaf beetles, specifically removing the C(5)-H(R) hydrogen atom from (4R,5S,2E,6E)-[4,5-(2)H(2)]-2-methyl octa-2,6-diene-1,8-diol (17). These findings have enabled us to construct a detailed account of iridoid biosynthesis in rove beetles, which resembles the biosynthetic route in leaf beetle larvae, but exhibits distinct stereochemical differences. PMID- 11267898 TI - Effects of a mustard trypsin inhibitor expressed in different plants on three lepidopteran pests. AB - The effects of mustard trypsin inhibitor MTI-2 expressed at different levels in transgenic tobacco, arabidopsis and oilseed rape lines have been evaluated against three different lepidopteran insect pests. 1. Plutella xylostella (L.) larvae were the most sensitive to the ingestion of MTI-2. The inhibitor expressed at high levels in arabidopsis plants caused rapid and complete mortality. High mortality and significantly delayed larval development were also detectable in oilseed rape expressing MTI-2 at lower levels. 2. Mamestra brassicae (L.) larvae were sensitive only at high MTI-2 expression level, as obtained in transgenic tobacco and arabidopsis, whereas no effects were observed for larvae fed on plants showing relatively low expression levels such as those of oilseed rape lines. 3. Feeding bioassays with Spodoptera littoralis (Boisduval) larvae were carried out using the same oilseed rape lines, showing that at these low expression levels no mortality was observed although a delay in larval development did occur. The levels of insect gut proteolytic activities of the larvae still alive at the end of a 7 day feeding bioassay were usually higher than in the controls, but no new proteinases were expressed in any case. The combined results described in this paper demonstrate altogether the relevance of a case-by-case analysis [target insects and proteinase inhibitor (PI) level of expression in planta] in a PI-based strategy for plant protection. PMID- 11267899 TI - Characterization of acyl-CoA-binding protein (ACBP) in the pheromone gland of the silkworm, Bombyx mori. AB - Various fatty acyl-CoAs are involved as intermediates or precursors of sex pheromone components in the biosynthetic pathway of the pheromones in many lepidopteran insects. We have purified a 10-kDa protein from the cytosolic fraction of Bombyx mori pheromone glands by using affinity chromatography with a palmitoyl-CoA-agarose column and reversed-phase HPLC. Amino acid sequence analysis of the fragment peptides obtained from the purified protein, and a homology search, revealed that this protein was a member of acyl-CoA-binding proteins (ACBPs). MALDI-TOF mass spectral analysis of the purified protein and cloning of the gene from a pheromone gland cDNA library confirmed B. mori ACBP to be a 90 amino acid protein with 78.9% identity to that of Manduca sexta ACBP. The secondary structure of the recombinant B. mori ACBP was determined by NMR spectroscopy. Northern blot analysis demonstrated that B. mori ACBP was predominantly expressed in the pheromone gland and the corresponding transcript was expressed from the day before adult eclosion. Present results suggest that ACBP plays a significant role in the production of sex pheromones regulated by the neurohormone, pheromone biosynthesis activating neuropeptide (PBAN). PMID- 11267900 TI - Structural analysis of gene encoding cuticle protein BMCP18, and characterization of its putative transcription factor in the silkworm, Bombyx mori. AB - BMCP18(2) is one of the major cuticle proteins identified in the larval cuticle of the silkworm, Bombyx mori. A genomic clone coding for BMCP18 was isolated from a B. mori genomic library, and its structure was analyzed. The BMCP18 gene consists of three exons interspersed by two introns. Bm1 element-like sequences were identified around this gene, suggesting possible involvement of this retroposon in the duplication of B. mori cuticle protein genes during evolution. A structural comparison of the BMCP18 gene and related cuticle protein genes of other lepidopteran species (MSCP14.6 and HCCP12) showed that the 5' upstream region of the BMCP18, MSCP14.6, and HCCP12 genes has a 12-bp identical sequence matching the recognition sequence for transcription factors COUP-TF and HNF-4. This implies that molecular mechanisms regulating expression of these cuticle protein genes are also conserved. mRNAs coding for Bmsvp, the B. mori homolog of Drosophila Seven-up, which is known as a homolog of vertebrate COUP-TF, and BmHNF 4, a homolog of vertebrate HNF-4, were detected in the larval epidermis. Bmsvp bound to the 12-bp sequence in vitro, suggesting that Bmsvp regulates the BMCP18 gene expression. PMID- 11267901 TI - Multiple transport pathways for dibasic amino acids in the larval midgut of the silkworm Bombyx mori. AB - The transport pathways for dibasic amino acids were investigated in brush border membrane vesicles (BBMV) from the anterior-middle (AM) and posterior (P) regions of Bombyx mori midgut. In the absence of K(+), a low-affinity saturable transport of arginine in both AM- and P-BBMV (K(m) 1.01 mM, V(max) 4.07 nmol/7s/mg protein and K(m) 1.38 mM, V(max) 2.26 nmol/7s/mg protein, respectively) was detected. Arginine influx was dependent on the membrane electrical potential (Deltapsi) and increased raising the alkalinity of the external medium from pH 7.2 to 10.6. Competition experiments indicated the following order of substrate affinity: arginine, homoarginine, N(G)-monomethylarginine, N(G) nitroarginine>lysine>>ornithine>cysteine>methionine. Leucine, valine and BCH (2 amino-2-norbornanecarboxylic acid) did not inhibit arginine influx. In the presence of external K(+), the influx of arginine as a function of arginine concentration fitted to a complex saturation kinetics compatible with both a low affinity and a high-affinity component. The latter (K(m) 0.035 mM, V(max) 2.54 nmol/7s/mg protein) was fully characterized. The influx rate had an optimum at pH 8.8, was strongly affected by Deltapsi and was homogeneous along the midgut. The substrate affinity rank was: homoarginine>arginine, N(G) monomethylarginine>>cysteine, lysine>>N(G)-nitroarginine>ornithine>methionine. Leucine and amino acids with a hydrophobic side chain were not accepted. This system is also operative in the absence of potassium, with the same order of specificity but a very low activity. Lysine influx is mediated by two more transport systems, the leucine uniport and the K(+)/leucine symport specific for amino acids with a hydrophobic side chain that recognizes lysine at extravesicular pH values (pH(out)) exceeding 9. Both the uniport and the symport differ from the cationic transport systems so far identified in mammals because they are unaffected by N-ethylmaleimide, have no significant affinity for neutral amino acids in the presence of the cation and show a striking difference in their optimum pH. PMID- 11267902 TI - Identification of six chymotrypsin cDNAs from larval midguts of Helicoverpa zea and Agrotis ipsilon feeding on the soybean (Kunitz) trypsin inhibitor. AB - Lepidopteran insects like Helicoverpa zea and Agrotis ipsilon produce STI insensitive trypsins in the midgut following ingestion of dietary plant proteinase inhibitors like STI [Broadway, R. M., J. Insect Physiol. 43(9) (1997) 855-874]. In this paper, the effects of dietary STI on a related family of midgut serine proteinases, the chymotrypsins, were investigated. STI-insensitive midgut chymotrypsins were detected in larvae of H. zea and A. ipsilon feeding on diets containing 1% STI while STI-sensitive chymotrypsins were present in larvae feeding on diets containing 0% STI. These chymotrypsins were unaffected by TPCK, a diagnostic inhibitor of mammalian chymotrypsins but were fully inhibited by chymostatin. Four midgut cDNA libraries were constructed from larvae of each species fed either 0% STI or 1% STI diets. Six full-length cDNAs(1) encoding diverse preprochymotrypsins were isolated (three from H. zea and three from A. ipsilon) with certain sequence motifs that set them apart from their mammalian counterparts. Northern blots showed that some chymotrypsin mRNA were detected at higher levels while others were down-regulated when comparing insects reared on 0% STI and 1% STI diets. Southern hybridizations suggested that (like mammals) both species contained several chymotrypsin genes. A full-length chymotrypsin gene(1) from H. zea was sequenced for the first time and the presence of four introns was deduced. A first time comparison of 5' upstream regions(1) from three chymotrypsin genes and two trypsin genes of A. ipsilon indicated the presence of putative TATA boxes and regulatory elements. However a lack of consensus motifs in these upstream regions suggested the likelihood of multiple trans factors for regulation of genes encoding digestive proteinases and a complex response mechanism linked to ingestion of proteinase inhibitors. PMID- 11267903 TI - Transcriptional induction of diverse midgut trypsins in larval Agrotis ipsilon and Helicoverpa zea feeding on the soybean trypsin inhibitor. AB - Midgut trypsins insensitive to inhibition by the soybean trypsin inhibitor (STI) were found to be transcriptionally regulated in A. ipsilon and H. zea larvae feeding on STI, as demonstrated by injections with actinomycin, a transcriptional inhibitor, which abolished the production of these STI-insensitive trypsins. The induced, STI-insensitive trypsins differed from the constitutive, STI-sensitive trypsins in their susceptibility to inhibitors based on sizes, suggesting that the induced enzymes limited access to their active site by blocking bulky inhibitors. Twenty midgut cDNA fragments(1) were amplified using trypsin-specific PCR primers and at least twelve were shown to encode structurally diverse trypsins. High sequence diversity was observed for both the enzymes encoded by STI-induced mRNAs and those from larvae that had not been exposed to STI. Northern blots showed that midgut mRNAs hybridizing to various trypsin cDNA probes were either transcribed de novo or up-regulated following ingestion of STI. Southern hybridizations indicated the presence of multiple trypsin gene families in the insect genomes. The complete sequence of a trypsin gene(1) from A. ipsilon (AiT9) revealed the presence of three introns. Comparison of 5' upstream sequences(1) from AiT9 and AiT6 genes from A. ipsilon revealed putative TATA box and disparate regulatory motifs, within 500 bp of each translational start site. PMID- 11267904 TI - Adenylate cyclase in prothoracic glands during the last larval instar of the silkworm, Bombyx mori. AB - We have previously reported that the absence of prothoracicotropic hormone (PTTH) signal transduction during the early last larval instar of Bombyx mori plays a role in leading to very low ecdysteroid levels in the hemolymph, inactivation of the corpora allata, as well as larval-pupal transformation. In the present study, adenylate cyclase was characterized in crude preparations of prothoracic gland cell membranes in an effort to localize the cause of refractoriness to PTTH. It was found that cyclase activity of the prothoracic glands from the day 6 last instar showed activation responses to fluoride, a guanine nucleotide analogue, as well as calmodulin (CaM) in dose-dependent fashions. The additive effects of day 5 prothoracic gland adenylate cyclase stimulation by fluoride and CaM imply that there may exist Gs protein-dependent and CaM-dependent forms of adenylate cyclase. For day 1 last instar prothoracic glands, which showed no response to stimulation by PTTH in either cAMP generation or ecdysteroidogenesis, adenylate cyclase activity exhibited far less responsiveness to Ca(2+)/CaM than did that from day 5 glands. These findings suggest that day 1 prothoracic glands may possess some lesions in the receptor-Ca(2+) influx-adenylate cyclase signal transduction pathway and these impairments in PTTH signal transduction may be, at least in part, responsible for decreased ecdysteroidogenesis. PMID- 11267905 TI - Homologues of fibroin L-chain and P25 of Bombyx mori are present in Dendrolimus spectabilis and Papilio xuthus but not detectable in Antheraea yamamai. AB - Low molecular mass protein components of fibroin, whose electrophoretic patterns before and after the reductive cleavage of disulfide bonds were similar to those of L-chain and P25 of Bombyx mori, were identified in fibroin samples of Bombyx mandarina, Dendrolimus spectabilis and Papilio xuthus but not of Antheraea yamamai. Fibroin of A. yamamai is suggested to form a dimer of H-chain. Full length cDNA sequences were cloned for the homologues of L-chain and P25 from B. mandarina, D. spectabilis and P. xuthus. The deduced sequences of L-chain and P25 of B. mandarina are almost identical to those of B. mori, each containing a single amino acid change. Homologues of L-chain and P25 of D. spectabilis and P. xuthus show about 50% overall identity, respectively, with those of B. mori, but essential structural features; i.e. the three Cys residues in an L-chain and the eight Cys residues and one of the potential N-glycosylation sites in P25, are conserved in both species. These results, together with the published results for Galleria mellonella, suggest that the three-components (H-chain, L-chain and P25) complex of fibroin is rather common among Lepidopteran silk-producing insects, in contrast to the H-H dimer type found in the saturnid silkworm. PMID- 11267906 TI - Expression of CYP6B1 and CYP6B3 cytochrome P450 monooxygenases and furanocoumarin metabolism in different tissues of Papilio polyxenes (Lepidoptera: Papilionidae). AB - The CYP6B1 and CYP6B3 cytochrome P450 monooxygenases in the midgut of the black swallowtail participate in the metabolism of toxic furanocoumarins present in its host plants. In this study, biochemical analyses indicate that the fat body metabolizes significant amounts of the linear furanocoumarins bergapten and xanthotoxin after larvae feed on xanthotoxin. Northern analyses of the combined CYP6B1/3 transcript expression patterns indicate that transcripts in this P450 subfamily are induced in the midgut and fat body by xanthotoxin. Semi quantitative RT-PCR analyses of individual CYP6B1/CYP6B3 mRNAs indicate that CYP6B1 transcripts are induced by xanthotoxin in all tissues examined and that CYP6B3 transcripts are induced in the fat body only. These results indicate that the fat body participates in the P450-mediated metabolism of excess furanocoumarins unmetabolized by the midgut. Although transcripts of both genes were detected and CYP6B1 transcripts were induced by xanthotoxin in the integument, furanocoumarin metabolism was not detected. Comparison of these P450 promoters with the promoters of alcohol dehydrogenase genes expressed in the fat bodies of several Drosophila species suggest that the xanthotoxin inducibilities of these P450 genes in fat bodies are regulated by elements other than those modulating expression of Adh genes. PMID- 11267907 TI - Characterization and cloning of a Tenebrio molitor hemolymph protein with sequence similarity to insect odorant-binding proteins. AB - The yellow mealworm beetle, Tenebrio molitor, produces a number of moderately abundant low molecular weight hemolymph proteins ( approximately 12 kDa) which behave in a similar manner during purification and share antigenic epitopes. The cDNA sequence of the major component (THP12) was determined and the deduced protein sequence was found to be similar to those of insect odorant-binding proteins. Southern blot analysis suggests that at least some of the diversity in this family of proteins is encoded at the gene level. Both northern and western blot analysis indicate that THP12 is present in a variety of developmental stages and both sexes. THP12 was originally classified as an antifreeze protein, but the lack of antifreeze activity in the recombinant protein, as well as the clear separation of the antifreeze activity from THP12 following HPLC purification, has ruled out this function. The abundance of THP12, the similarity of THP12 to insect odorant-binding proteins, and the presence of hydrophobic cavities inside the protein (Rothemund et al., A new class of hexahelical insect proteins revealed as putative carriers of small hydrophobic ligands. Structure, 7 (1999) 1325-1332.) suggest that THP12 may function to carry non-water soluble compounds in the hemolymph. THP12 is also similar, particularly in structurally important regions, to other insect proteins from non-sensory tissues, suggesting the existence of a large family of carrier proteins which may perform diverse functions throughout the insect. PMID- 11267908 TI - Alternative splicing of the BSC1 gene generates tissue-specific isoforms in the German cockroach. AB - Voltage-gated sodium channels are integral transmembrane proteins responsible for the rapidly-rising phase of action potentials in most excitable cells. In mammals, the functional diversity and wide distribution of sodium channel proteins in various tissues and cell types are achieved mainly by selective expression of many distinct sodium channel genes. In the model insect, Drosophila melanogaster, however, only one confirmed sodium channel gene, para, and one putative sodium channel gene, DSC1, are known. We cloned and sequenced a DSC1 ortholog, BSC1, from the German cockroach, Blattella germanica. We found that the BSC1 transcript was present in a wide range of tissues, including nerve cord, muscle, gut, fat body and ovary, whereas the para transcript was detected only in nerve cord and muscle. Moreover, different tissues contained distinct alternatively spliced variants of BSC1, and two muscle-specific spliced variants are predicted to encode truncated proteins with only the first two of the four homologous domains. Therefore, alternative splicing and expression of distinct splicing variants in functionally different tissues may be a major mechanism by which insects increase BSC1 channel diversity in neuronal and non-neuronal tissues. PMID- 11267909 TI - Purification and characterization of trans-permethrin metabolizing microsomal esterases from workers of the eastern subterranean termite, Reticulitermes flavipes (Kollar). AB - Three alpha-naphthyl acetate hydrolyzing esterase isozymes were purified from microsomes prepared from Reticulitermes flavipes workers. The two step process involved sequential preparative IEF followed by continuous elution preparative electrophoresis on a 5% non-denaturing polyacrylamide gel. The first IEF run resulted in 5.4-fold purification with a yield of 46.1%. Subsequent IEF further purified the esterases 14.3-fold and 12% yield. Preparative electrophoresis of the pooled IEF fractions produced three major peaks of alpha-naphthyl acetate hydrolyzing activity. The esterases were correspondingly designated microsomal esterase (ME) 1, ME 2, and ME 3 based on increasing molecular retention on a native PAGE gel. ME 1, ME 2, and ME 3 were acidic proteins with pI values of 4.61, 4.70, and 4.77, respectively. Molecular mass as determined by gel filtration chromatography of ME 1, ME 2, and ME 3 was 69, 64, and 62 kDa, respectively. SDS-PAGE gels produced a single band for each of the isozymes with a molecular mass of 63 kDa indicating that the esterases were monomers. Specific activities of ME 1, ME 2, and ME 3 increased with increasing pH and the enzymes were active over a broad temperature range (25-55 degrees C). The three purified isozymes were inhibited at low concentration by paraoxon (10(-10) M), chlorpyrifos (10(-6) M), DEF (10(-6) M), and PMSF (10(-6) M) indicating that they were "B" type serine esterases. Conversely, inhibition was not observed at 10(-4) M eserine, PHMB, or CaCl(2), further supporting the conclusion that the microsomal esterases were of the "B" type. None of the isozymes was inhibited by 10(-4) M imidacloprid, fipronil, or PBO. Quantitatively, ME 1, ME 2 and ME 3 metabolized t-permethrin at 21.8, 21.0, and 38.8 nmol/h/mg protein, representing a purification factor of 333-, 318-, and 591-fold over microsomes, respectively. The three isozymes produced the same type and number of t-permethrin metabolites. PMID- 11267910 TI - Infection density of Wolbachia and incompatibility level in two planthopper species, Laodelphax striatellus and Sogatella furcifera. AB - Wolbachia, a bacterial endosymbiote of arthropods, causes cytoplasmic incompatibility (CI) in many insect species. CI traits were studied in two planthopper species, Laodelphax striatellus and Sogatella furcifera, and Wolbachia densities in these planthopper species were calculated by quantitative PCR methods. The CI level of L. striatellus was quite high and even aged males strongly caused CI. In contrast, S. furcifera showed partial CI, and males lost their ability to cause CI with age. Wolbachia infecting these two planthopper species were the same with respect to the nucleotide sequences of Wolbachia genes, 16S rDNA, ftsZ gene, groE genes, and wsp gene. Two methods for quantitative PCR, one using a DNA sequencer and the other a real-time sequence detection system, were established to calculate the amount of Wolbachia in the planthoppers. The density of Wolbachia in S. furcifera males was quite low. The difference in CI levels between the two planthopper species seems to be due to different amounts of Wolbachia infecting males. PMID- 11267912 TI - Dendritic vaccine promising for gliomas. PMID- 11267911 TI - Specific biochemical marker-based techniques for the identification of damage to Douglas-fir seed resulting from feeding by the western conifer seed bug, Leptoglossus occidentalis Heidemann (Hemiptera: Coreidae). AB - Specific biochemical marker-based techniques were tested for their ability to distinguish between seeds of Douglas-fir, Pseudotsuga menziesii (Mirbel) Franco, that were filled or unfilled (aborted) at maturity and those that were damaged or emptied by the western conifer seed bug, Leptoglossus occidentalis Heidemann. A polyclonal antibody raised against salivary gland extracts from L. occidentalis successfully identified residual salivary proteins on Western blots containing proteins from Douglas-fir seeds that had sustained various degrees of seed bug feeding damage. In a single blind experiment, the polyclonal antibody correctly identified 100% of undamaged control, 97% of unfilled control (aborted), and 98% of seed bug damaged seeds. Polyclonal antibodies raised against insoluble alfalfa crystalloid storage protein (11S globulin) detected the depletion of 11S globulin and the subsequent appearance of its hydrolyzed fragments in the soluble protein fraction of Douglas-fir seeds that were fed-upon by the seed bug. Feeding by L. occidentalis nymphs caused ca. 98% depletion of insoluble protein, but only ca. 53% reduction in the amount of soluble protein in seeds that appeared empty on radiographs. By comparison, unfilled (aborted) seeds contained significantly less insoluble and soluble protein than empty seeds that were fed-upon by L. occidentalis; moreover, no crystalloid (11S globulin) breakdown products were generated. The biochemical markers described in this study are reliable tools that can be used to identify conifer seeds that have sustained light to severe damage from L. occidentalis feeding. PMID- 11267914 TI - A future vaccine for diabetes? PMID- 11267913 TI - Trojan antibiotics. PMID- 11267917 TI - Jurgen Drews discusses the future of the industry. Interviewed by Rebecca N. Lawrence. PMID- 11267918 TI - Are drug targets missed owing to lack of physical activity? PMID- 11267919 TI - Gene therapy: will it deliver for RA? PMID- 11267920 TI - Wither solid-phase chemistry? - Reply. PMID- 11267921 TI - Multi-modal combination gene therapy for malignant glioma using replication defective HSV vectors. AB - Herpes simplex virus (HSV) may be modified to produce a non-pathogenic vector that is capable of delivering multiple transgenes simultaneously to cells, both safely and efficiently. We have exploited this property to develop viruses that target glioblastoma, a malignancy that is currently associated with a poor prognosis. Using rationally selected combinations of therapeutic transgenes coupled with gamma-knife radiotherapy, the ablation of experimental tumours in animal models has been demonstrated. Combination gene therapy using replication defective HSV vectors represents a promising and exciting approach to tackling malignancy in the CNS. PMID- 11267922 TI - Screening for human ADME/Tox drug properties in drug discovery. AB - There is no doubt that ADME/Tox drug properties, absorption, distribution, metabolism, elimination and toxicity, are properties crucial to the final clinical success of a drug candidate. It has been estimated that nearly 50% of drugs fail because of unacceptable efficacy, which includes poor bioavailability as a result of ineffective intestinal absorption and undesirable metabolic stability(1). It has also been estimated that up to 40% of drug candidates have failed in the past because of safety issues(2). In this review, the methodologies that are available for use in drug development as in vitro human-based screens for ADME/Tox drug properties are discussed. PMID- 11267923 TI - Methods used to assess pulmonary deposition and absorption of drugs. AB - The assessment of pulmonary drug absorption and deposition is becoming increasingly important in drug development. Absorption information can be used to maximize pulmonary selectivity, to screen drug candidates and to help evaluate the bioequivalence of generic inhalation products. Several methods are available to investigate pulmonary drug absorption and deposition, ranging from in vitro experiments to in vivo pharmacokinetic and pharmacodynamic analyses. In combination, these methods can indicate the fate of an inhaled drug. PMID- 11267924 TI - Monitor: molecules and profiles. AB - Monitor provides an insight into the latest developments in drug discovery through brief synopses of recent presentations and publications together with expert commentaries on the latest technologies. There are two sections: Molecules summarizes the chemistry and the pharmacological significance and biological relevance of new molecules reported in the literature and on the conference scene; Profiles offers commentary on promising lines of research, emerging molecular targets, novel technology, advances in synthetic and separation techniques and legislative issues. PMID- 11267925 TI - Monitor: molecules and profiles. AB - Monitor provides an insight into the latest developments in drug discovery through brief synopses of recent presentations and publications together with expert commentaries on the latest technologies. There are two sections: Molecules summarizes the chemistry and the pharmacological significance and biological relevance of new molecules reported in the literature and on the conference scene; Profiles offers commentary on promising lines of research, emerging molecular targets, novel technology, advances in synthetic and separation techniques and legislative issues. PMID- 11267926 TI - Relationship between partial inhibition of glycolysis and hemolysis after induction of gametocytogenesis in synchronous cultures of Plasmodium falciparum. AB - In the present study, the low molecular-weight fraction of the culture supernatant of anti-Plasmodium falciparum antibody-producing hybridoma cells (HybSL) was used in synchronous culture with P. falciparum FVO strain. When synchronous cultures were treated with HybSL solution on day 5, gametocytogenesis was also induced. Gametocytes were consistently found from the third day after treatment and reached a peak on the fourth day. An increase in pH and hemoglobin concentrations and decrease in lactate concentrations were observed on the first day after treatment. These phenomena suggested that HybSL solution partially inhibited glycolysis of erythrocytes parasitized with schizonts and resulted in hemolysis of infected erythrocytes. On the other hand, the production of gametocytes did not increase in cultures treated with HybSL solution on day 4 of synchronous cultures in which ring forms were plentiful. Most ring forms were not killed by HybSL solution and quickly developed to trophozoites and schizonts rather than gametocytes. Consequently, it is assumed that ring forms on day 4 of synchronous cultures have finished differentiation into the asexual stage. The conversion of asexual parasites to gametocytes may be triggered only when late stage trophozoites or early-stage schizonts are treated with HybSL solution. PMID- 11267927 TI - Variation of incubation time in an in vitro drug susceptibility test of Plasmodium falciparum isolates studied in the Solomon Islands. AB - A study on chloroquine resistance of falciparum malaria was conducted in the Solomon Islands. Both in vitro and clinical tests were performed. In our regular studies of in vitro chloroquine susceptibility tests on Plasmodium falciparum from non-immuners in Japan, the threshold point to differentiate resistant and susceptible isolates was set at a 0. 114 microM chloroquine in the semi-micro culture system, and this point was also applicable in the study of the malaria parasite taken in the highly endemic malarious area with good coincidence with clinical observation. Variation in the incubation time (24-63) to reach the schizont stage of the isolated parasites were noted. It appeared that chloroquine resistant P. falciparum showed traits to reach the schizont stage within a shorter incubation period. PMID- 11267929 TI - Eimeria auratae n. sp. (Apicomplexa: Eimeriidae) infecting the lizard Mabuya aurata in Saudi Arabia. AB - Eimeria auratae n. sp. was described from the gall bladder of the lizard Mabuya aurata collected at Al-Hofuf village, eastern region, Saudi Arabia. Morphology of sporulated as well as non-sporulated oocysts were studied. Sporulated oocysts were ellipsoidal 22-31.5x13.5-21.8 (27.7x18.5) microm with smooth brownish-yellow bilayered wall, 1.1 (0.9-1.3) microm. Micropyle, polar granule and oocyst residuum were absent. Sporocysts were ellipsoidal 10.5-12.8x7.5-9 (11.8x8.5) microm. Sporocyst residuum was present but Stieda body was absent. Sporozoites were crescent-shaped, blunt at one end and slightly tapered at the other. Eimeria species from Scincidae were compared. PMID- 11267928 TI - Cytokines and the acute phase response in post-treatment reactive encephalopathy of Trypanosoma brucei brucei infected mice. AB - Stimulation of the acute phase response during infection of mice with Trypanosoma brucei brucei (T. b. brucei) was investigated in an experimental model of the post-treatment reactive encephalopathy (PTRE), a common side-effect of anti trypanosome therapy. Plasma levels of the acute phase proteins (APP), haptoglobin (Hp) and serum amyloid P (SAP) increased by day 7 post-infection, but by day 20 had fallen to an intermediate level. This was accompanied by induction of the cytokines, interleukin (IL)-6 and tumour necrosis factor-alpha (TNFalpha) in both liver and brain. Treatment of mice on day 21 with a subcurative dose of diminazene aceturate (Berenil), a procedure known to induce a mild PTRE, cleared the parasite from the circulation with plasma APP and liver expression of mRNA for IL-6 and TNFalpha returning to the levels in the controls. Cytokine mRNA for both IL-6 and TNFalpha was detected in the brains of animals with developing PTRE although TNFalpha was not significantly greater than in the control group. A further subcurative dose of Berenil, leading to a more severe PTRE, was associated with elevated serum concentrations of Hp and SAP, increased TNFalpha mRNA in the liver and detectable IL-6 and TNFalpha mRNA in the brain. mRNA for IL 1alpha was expressed in brain and liver samples from all animals. A severe PTRE caused a systemic acute phase response which was not apparent with a mild PTRE. The pattern of cytokine mRNA induction was similar following both drug treatments. However, the difference in APP production could be caused by a breakdown in the blood-brain barrier during severe PTRE allowing cytokine synthesised in the brain to enter the circulation and maintain a systemic response. PMID- 11267931 TI - Characterisation of stage-specific proteins of Oesophagostomum dentatum by preparative isoelectric focusing and lectin blotting. AB - The nodular worm of pigs, Oesophagostomum dentatum, has previously been shown to undergo distinct biochemical changes during its life cycle. This phenomenon was studied in more detail for the early parasitic stages. Differences between infective third-stage larvae (L3), parasitic fourth-stage larvae cultivated in vitro (L4c), and pre-adult larvae recovered from the intestinal contents of pigs (L4p) were compared with respect to their protein and glycoprotein patterns by solubility-based protein fractionation and preparative isoelectric focusing followed by SDS-PAGE or by Western blotting with various lectins. While differences between the L4 were only minor (only three bands were specific for either L4c or L4p), L3 displayed distinctly different protein patterns with four L3-specific and nine L4-specific bands. Concanavalin A bound to a variety of glycoproteins, partly in a stage-specific manner, while Ricinus communis Agglutinin 120, Wheat Germ Agglutinin, Peanut Agglutinin and Soybean Agglutinin bound to fewer, partly stage-specific, molecules. PMID- 11267930 TI - A potent antimalarial activity of Hydrangea macrophylla var. Otaksa leaf extract against Plasmodium yoelii 17XL in mice. AB - The antimalarial activity of the hot-water extract of Hydrangea macrophylla var. Otaksa leaves was evaluated against Plasmodium yoelii 17XL in mice. Non-treated control mice died from 6 to 7 days after infection, but mice treated with the leaf extract survived during the experiment. Mice given the extract orally showed low parasitemia levels during administration. Following a transient recrudescence of malaria parasites in the bloodstream of treated mice, no parasites could be detected by a microscopic examination. Furthermore, the 30% MeOH aq. eluate and 50% MeOH aq. eluate from dried leaves of H. macrophylla var. Otaksa showed an antimalarial activity in vivo. Sulfamonomethoxine was orally given to infected mice to compare with the antimalarial activity of the hot-water extract of leaves. Sulfamonomethoxine given orally reduced parasitemia, but no complete cure of mice was observed. PMID- 11267932 TI - New tools for genetic analysis of Entamoeba histolytica: blasticidin S deaminase and green fluorescence protein. PMID- 11267933 TI - Effect of smoking on serum pepsinogen I level depends on serological status of Helicobacter pylori. AB - Serum pepsinogen (sPG) levels are used in gastric cancer screening programs. However, modification of sPG levels by smoking habit, according to the status of Helicobacter pylori (H. pylori) infection has been little investigated. This study investigated the effects of smoking on serum levels of pepsinogen I (PG I), pepsinogen II (PG II), and gastrin by IgG titer of antibody against H. pylori (Hp IgG titer) using the data from 356 current-smokers and 262 non-smokers (133 never smokers and 129 ex-smokers) in a cross-sectional study of 618 men aged 40 to 49 years. PG I, PG II, PG I / PG II ratio and gastrin were significantly associated with Hp-IgG titer in never-smokers [Spearman's correlation coefficient (95% confidence interval): 0.23 (0.07, 0.39), 0.52 (0.41, 0.63), -0.40 (-0.54,-0.27), and 0.25 (0.10, 0.41), respectively]. However, the correlation coefficients of PG I and PG II decreased in current-smokers, 0.02 (-0.1, 0.13) and 0.32 (0.22, 0.42), respectively. In H. pylori seronegative and low titer cases, the mean PG I level was significantly (P < 0.01) higher in current-smokers, compared with non smokers. However, in high titer cases, the mean PG I level was lower in current smokers. Mean PG II and gastrin levels, and PG I / PG II ratio did not differ according to smoking habits by Hp-IgG titer. The gastrin level was significantly correlated with PG II, but not PG I. These data indicate that current smoking influences the serum PG I level depending on Hp-IgG titer and the associations between sPGs and Hp-IgG titer. Gastrin is not involved in the modification of PG I levels by smoking. PMID- 11267934 TI - Sex hormone dependency of diethylnitrosamine-induced liver tumors in mice and chemoprevention by leuprorelin. AB - The prevalence of liver tumors throughout the world makes it imperative to seek chemopreventive agents. This tumor appears to be hormone-responsive and hormonal manipulations may therefore be beneficial. On this basis, both sexes of 12-day old B6C3F(1) mice were injected i.p. with diethylnitrosamine (DEN) at the dose of 2.5 mg / g body weight and observed for 32 weeks (males) or 36 weeks (females). In 100% of male mice, liver tumors were observed with an average diameter of 2.72 mm and multiplicity of 60.8. Orchidectomy at 6 weeks of age in these mice inhibited the incidence, multiplicity and size to 63%, 5.6 and 1.54 mm, respectively. By further implantation with an E(2) pellet at monthly intervals, these parameters were reduced to 26%, 0.6 and 0.61 mm, respectively. Administration of a gonadotropin-blocking chemical, leuprorelin, to DEN-treated male mice significantly reduced the multiplicity and size of tumors to 18.3 and 2.54 mm (P < 0.01 compared to those of DEN only). In female mice, the incidence of liver tumor was significantly smaller than that of males. However, ovariectomy and / or testosterone supplement significantly increased the occurrence of liver tumor. An anti-estrogen, toremifene, caused a marked further decrease of liver tumors. Mitotic indices with bromodeoxyuridine in tumor tissues paralleled the occurrence of liver tumors. Serum testosterone levels were significantly reduced by orchidectomy or by leuprorelin administration. These results further confirm that liver tumor is testosterone-responsive and hormonal manipulation by surgical orchidectomy or by chemical orchidectomy i.e. by leuprorelin, could substantially prevent the appearance of liver tumors. PMID- 11267935 TI - Targets of transcriptional regulation by transforming growth factor-beta: expression profile analysis using oligonucleotide arrays. AB - Transforming growth factor-betas (TGF-betas) are potent inhibitors of cell proliferation, and disruption of components of the TGF-beta signaling pathway leads to tumorigenesis. Mutations of transmembrane receptors and Smads mediating intracellular signaling have been reported in various cancers. To identify transcriptional targets of TGF-beta, we conducted an expression profile analysis. HaCaT cells derived from human keratinocytes and highly sensitive to TGF-beta were treated with TGF-beta in the absence or presence of cycloheximide (CHX). mRNAs extracted from the HaCaT cells were used for hybridization of oligonucleotide arrays representing approximately 5600 human genes. TGF-beta increased the expression of PAI-1, junB, p21 cdk inhibitor, Smad7, betaIG-H3, and involucrin that have been reported to be up-regulated by TGF-beta, validating the usefulness of this approach. The induction of betaIG-H3 by TGF-beta was completely abolished by CHX, suggesting that the transcription of betaIG-H3 is not directly regulated by TGF-beta. Unexpectedly, we identified more genes down regulated by TGF-beta than up-regulated ones. TGF-beta repressed the expression of epithelial specific Ets that may be involved in breast and lung tumorigenesis, which could contribute to tumor suppression by TGF-beta. Among a panel of cell cycle regulators, TGF-beta induced the expression of p21 cdk inhibitor; however, the induction of other cdk inhibitors was not significant in the present study. Taken together, the results suggest that TGF-beta may suppress tumorigenesis through positive and negative regulation of transcription. PMID- 11267937 TI - Glutathione S-transferases in small intestinal mucosa of patients with coeliac disease. AB - Patients with villous atrophy due to coeliac disease have an increased risk of developing small intestinal malignancies. Intestinal glutathione (GSH) and glutathione S-transferases (GST) are involved in the protection against carcinogenesis. The aim of this study was to evaluate GSH content and GST enzyme activity in small intestinal mucosa of untreated coeliacs compared to controls. We evaluated GSH content and GST enzyme activity, including the levels of GST classes alpha, mu, pi and theta, in small intestinal biopsies of untreated coeliacs (flat mucosa, Marsh IIIC, n = 12) compared to normal subjects (n = 23). Next, we evaluated GSH and GST's in coeliacs in remission (Marsh 0 - I, n = 11), coeliacs with persisting villous atrophy while on a gluten-free diet (partial villous atrophy, Marsh IIIA (n = 5); subtotal villous atrophy, Marsh IIIB (n = 6)) and patients with infiltrative / crypt-hyperplastic Marsh II lesions (n = 4). Total GST enzyme activity and content of GSTalpha are markedly suppressed in Marsh IIIC lesions compared to controls (resp. 220 +/- 79 vs. 464 +/- 189 nmol / mg protein*min (P < 0.001) and 2.79 +/- 2.46 vs. 6.47 +/- 2.29 mg / mg protein (P < 0.001)). In coeliacs in remission these levels normalized. Total GST enzyme activity and GSTalpha levels are proportionately lowered according to the degree of mucosal pathology in Marsh II, IIIA and IIIB. (Spearman's sigma correlation coefficient for total GST, -0.596, P < 0.001; GSTalpha, -0.620, P < 0.001). GSTmu, pi and theta and GSH levels are not significantly different in the selected study groups of mucosal pathology compared to controls. Total GST enzyme activity and content of GSTalpha in small intestinal mucosa are significantly lower in untreated coeliac disease compared to controls. In Marsh II, IIIA and IIIB, GST enzyme activity and GSTalpha content are proportionally lower according to the degree of mucosal pathology. Normal values are seen in coeliacs in remission. This correlation between coeliac disease and a suppressed GSH / GST detoxification system may explain in part the carcinogenic risk in untreated coeliac disease. PMID- 11267936 TI - The autocrine loop of epidermal growth factor receptor-epidermal growth factor / transforming growth factor-alpha in malignant rhabdoid tumor cell lines: heterogeneity of autocrine mechanism in TTC549. AB - To investigate the effects of the autocrine loop of epidermal growth factor receptor (EGFR)-epidermal growth factor (EGF) / transforming growth factor-alpha (TGF-alpha) on the proliferation and differentiation of malignant rhabdoid tumor (MRT), we used five MRT cell lines, TM87-16, STM91-01, TTC549, TTC642, and YAM RTK1. RT-PCR analyses revealed expression of EGFR mRNA in all MRT cell lines. In contrast, the expression of either EGF or TGF-alpha mRNA was detected in all MRT cell lines. Expression of EGF, TGF-alpha, and EGFR as determined by immunocytochemical staining and in situ hybridization, correlated with the results of RT-PCR. Upon differentiation-induction with 12-O-tetradecanoylphorbol 13-acetate (TPA), in TTC549, showing an expression of TGF-alpha but not EGF initially, de novo expression of EGF mRNA appeared abruptly on day 2 of TPA treatment. To confirm the EGFR-EGF / TGF-alpha autocrine loop, we used TGF-alpha, EGF, and their antibodies in the cultures. Monoclonal antibody (mAb) to EGFR alone significantly inhibited the growth of cell line TTC549. However, mAb to EGF or TGF-alpha could inhibit proliferation of this cell line only when administrated together. Our findings would suggest that growth of the TTC549 cell line is constitutionally regulated by TGF-alpha / EGFR, but that inhibition of this autocrine mechanism results in transient activation of an autocrine loop involving EGF / EGFR. Our results may indicate the presence of two different autocrine loops of EGFR-EGF and / or EGFR-TGF-alpha in MRT cell lines. The heterogeneity of autocrine mechanisms found in MRT cell lines would be consistent with the multiphenotypic diversity and aggressive characteristics of this enigmatic tumor. PMID- 11267938 TI - Aberrant expression of pRb, p16, p14ARF, MDM2, p21 and p53 in squamous cell carcinomas of lung. AB - Expression of cell cycle regulatory proteins in both the RB and p53 pathways was investigated in 50 cases of squamous cell carcinoma (SCC) of the lung using immunohistochemical techniques. Abnormality of pRb and p16 expression was seen at the frequencies of 16% and 78%, respectively, and appeared to be in a reciprocal relationship. On the other hand, strong and diffuse p53 immunoreactivity was seen in 60% of SCCs. MDM2 and p14ARF expressions were each observed in about half of the cases of SCC and were not significantly associated with strong p53 immunoreactivity. Statistical analysis revealed that p14ARF expression was significantly correlated with both p16 and MDM2 expression. Moreover, strong p53 expression was not correlated with the expression of p21. In comparing clinicopathological status with the immunohistochemical results, lack of p16 immunoreactivity was observed in the elderly group (over 65 years) as compared with the younger group (less than 65 years). Strong p53 expression was frequently observed in higher stages of SCC, with the developing tumor located in the central field of the lung. Similarly, the frequency of p14ARF expression was high in centrally developed SCC, but low in SCC developed in the periphery. These results suggest that disruption of the RB and p53 pathways is a frequent event in SCC, and that abnormal expression of p16 and p53 plays a more critical role than that of pRB, p14ARF and MDM2 in the development of SCC of the lung. PMID- 11267939 TI - Reduced expression of laminin alpha 3 and alpha 5 chains in non-small cell lung cancers. AB - The basement membrane is considered to act as a barrier which hinders cancer cells from invading the surrounding stroma. In order to assess changes in essential components during neoplasia in the lung, we immunohistochemically studied distribution patterns of laminins alpha 3 and alpha 5 in 40 adenocarcinomas and 8 squamous cell carcinomas. The a 5 chain was generally preserved at the periphery, frequently disrupted in foci with alveolar collapse and absent in foci of fibroblastic proliferation within adenocarcinomas. Fragmentation and absence of laminin alpha 3 chain were more prominent than for alpha 5 chain. Laminin alpha 3 chain was partially fragmented or absent in peripheral areas of adenocarcinomas, being significantly different from alpha 5 chain. Non-small cell lung cancers with reduced alpha 5 chain showed a tendency for greater lymph node metastasis. In cultured normal air way epithelial cells, both laminin alpha 3 and alpha 5 chains were found to be expressed by northern analysis. Eleven of the twelve cultured lung cancer cell lines did not express alpha 3 chain and expression of alpha 5 chain was reduced in three. Quantitative RT-PCR analysis also demonstrated expression of laminin alpha 3 chain in adenocarcinoma tissues to be significantly lower than in normal lung tissues. These results suggest that expression of laminin alpha chains is often reduced in lung cancer cells and this might contribute to basement membrane fragmentation and subsequent proliferation of stromal elements, as well as play some role in the process of cancer cell invasion. PMID- 11267940 TI - Progesterone receptor A and B isoforms in the human breast and its disorders. AB - Two different isoforms of progesterone receptor (PR), PRA and PRB, are expressed in target tissues at comparable levels. In this study, we first examined PRA and PRB immunoreactivity in human breast cancer and various intraductal proliferative epithelial lesions, and correlated these findings with clinicopathologic parameters. We then examined mRNA expression of PRA and PRB in six cases of invasive ductal carcinoma using RT-PCR. Immunoreactivity for both PRA and PRB was positive in the great majority of proliferative disease without atypia (PDWA) (85% for PRA and 96% for PRB) and atypical ductal hyperplasia (ADH) (100% for PRA and 100% for PRB), but the ratio of immunopositive cases and immunohistochemical (IHC) scores was significantly smaller in ductal carcinoma in situ (DCIS) (65% for PRA and 75% for PRB) and invasive ductal carcinoma (IDC) (66% for PRA and 55% for PRB) than in PDWA and ADH. There was a significant positive correlation between IHC scores for PRA and estrogen receptor alpha (ERalpha) in IDC, DCIS and ADH but not between PRB and ERalpha. In IDC, both PRA and PRB IHC scores were significantly associated with histological grade, but there was no association between PRA or PRB status and lymph node involvement, tumor size, or prognosis of the patients. The expression of mRNAs for both PRA and PRB was detected in all six cases of IDC examined. These results suggest that both PRA and PRB are strongly associated with ERalpha in human breast and this relation may be disturbed in breast cancer. PMID- 11267941 TI - Successful induction of tumor-specific cytotoxic T lymphocytes from patients with non-small cell lung cancer using CD80-transfected autologous tumor cells. AB - Cytotoxic T lymphocytes (CTL) against human lung cancer cells are difficult to induce by a conventional method using tumor cell stimulation probably due to an insufficiency of tumor antigens (TA) or costimulatory molecules such as CD80. We, therefore, investigated the potential of CD80-transfected tumor cells as stimulators of the in vitro induction of autologous tumor-specific CTL from regional lymph node lymphocytes in patients with lung cancer. Five non-small cell lung cancer cell lines (two adenocarcinomas, 1 squamous cell carcinoma, 1 large cell carcinoma and 1 adenosquamous cell carcinoma) were established from surgical specimens and were successfully transduced with a plasmid constructed with expression vector pBj and human CD80 cDNA, using a lipofection method. CD80 transfected tumor cells (CD80-AT) significantly augmented the proliferation of autologous lymphocytes from all cases as compared with non-transfected tumor cells (AT). AT-stimulated lymphocytes from 4 out of 5 cases did not show any cytotoxicity against AT; however, lymphocytes stimulated with CD80-AT exhibited substantial cytotoxicity against parental AT in all 5 cases tested. AT-stimulated lymphocytes derived from only one out of 5 cases showed major histocompatibility complex (MHC)-class I-restricted cytokine production in response to AT, while the MHC-class I-restricted responses were found in CD80-AT-stimulated lymphocytes from 4 out of 5 cases. These results indicate that CD80 on tumor cells could be a beneficial costimulatory molecule to elicit CTL against lung cancer, and also show that TA recognized by CTL was frequently expressed on lung cancer cells. PMID- 11267942 TI - Autoantibody to heat shock protein Hsp40 in sera of lung cancer patients. AB - Heat shock protein Hsp40 is a stress protein with chaperone activity and has a cooperative function with Hsp70 in mammalian cells. We examined the possible expression of Hsp40 in lung tumor tissues using immunoblotting and immunohistochemistry, and established an enzyme-linked immunosorbent assay (ELISA) method to detect IgG antibody to Hsp40 in the serum using purified human Hsp40. Sera were obtained from 130 normal subjects and 50 patients with lung cancer. Lung tumor tissues and cells specifically overexpressed Hsp40, and no such expression was detected in normal lung tissues. Compared with normal sera, significantly higher levels of autoantibody to Hsp40 were present in patients with lung cancer. The present study is the first to demonstrate overexpression of Hsp40 in human tumor tissue and the associated presence of autoantibody to Hsp40 in the serum. These results suggest that overexpression of Hsp40 in tumor cells may be recognized as a self-antigen. PMID- 11267943 TI - Efficient gene transduction by RGD-fiber modified recombinant adenovirus into dendritic cells. AB - Dendritic cells (DC) are important antigen-presenting cells in the development of an anti-tumor T cell response. To extend the range of current immuno / gene therapies, we tested luciferase-expressing RGD-adenovirus (Ad) (Ad5lucRGD) mediated transduction into DC. Phenotypically characterized DC were generated from peripheral blood CD14(+) cells by incubation with granulocyte-macrophage colony-stimulating factor, interleukin-4 and tumor necrosis factor alpha. On the 7th day of culture, the cells became mature DC with a CD1a(+), CD11c(+), CD80(+), CD83(+), CD86(+), human leukocyte antigen (HLA)-DR(+), CD14- phenotype. The expression of alpha( v)beta(3) integrin was enhanced on day 3 and returned to the basal level on day 7. We then compared the transduction efficiency of an Ad5lucRGD system to that using conventional Ad, in cells harvested on days 1, 3 and 7 of culture. Luciferase activity was negligible in AdCMVLuc, but remarkable in cells processed with Ad5lucRGD. Activity was maximal in cells that had been cultured for 3 days. Recombinant Ad5 fiber knob protein blocked AdCMVLuc- and Ad5lucRGD-mediated gene transduction by 90% and 20%, respectively. Surface markers and cytokine production were not affected by Ad5lucRGD-mediated transduction. PMID- 11267944 TI - Cisplatin-incorporated polymeric micelles eliminate nephrotoxicity, while maintaining antitumor activity. AB - cis-Diamminedichloroplatinum (II) (cisplatin, CDDP), a potent anticancer agent, was bound to the aspartic acid residues of poly(ethylene glycol)-poly(aspartic acid) (PEG-P(ASP)) block copolymer by ligand substitution reaction at the platinum atom of CDDP. The polymeric drug thus obtained was observed to form a micelle structure in aqueous medium, showing excellent water solubility. In the present study, in vitro and in vivo antitumor activity against several human tumor cell lines, toxicity and pharmacokinetic characteristics in rodents of CDDP incorporated polymeric micelles (CDDP / m) were evaluated in comparison with those of CDDP. In vitro, CDDP / m exhibited 10 - 17% of the cytotoxicity of CDDP against human tumor cell lines. CDDP / m given by intravenous (i.v.) injection yielded higher and more sustained serum levels than CDDP. In vivo CDDP / m treatment resulted in higher and more sustained levels in tumor tissue than CDDP, and showed similar antitumor activity to CDDP against MKN 45 human gastric cancer xenograft. CDDP / m treatment caused much less renal damage than CDDP. These results indicate that CDDP / m treatment can reduce CDDP-induced nephrotoxicity without compromising the anticancer cytotoxicity of CDDP. PMID- 11267945 TI - Germline mutation of dihydropyrimidine dehydrogenese gene among a Japanese population in relation to toxicity to 5-Fluorouracil. AB - 5-Fluorouracil (5FU) is most commonly used in chemotherapy for human malignancy. Over 80% of administered 5FU is metabolically degraded by dihydropyrimidine dehydrogenase (DPD), a primary and rate-limiting enzyme in the 5FU metabolic pathway. A DPD-deficient phenotype among cancer patients, which has posed a serious problem in 5FU-based chemotherapy, was reported to be in part ascribed to germline mutations in dihydropyrimidine dehydrogenase (DPYD) gene. Therefore, we for the first time examined the frequencies and types of germline mutations in the DPYD gene among a total of 107 Japanese cancer patients and healthy volunteers. Of 214 alleles examined among them, 181 alleles were of the same type, which was assigned as wild type; 21 alleles revealed a nucleotide substitution resulting in silent mutation; and the remaining 12 alleles showed five types of nucleotide deletion or substitutions resulting in one frameshift and four missense mutations. Three of them, A74G, 812delT and L572V, were novel mutations. None of the study subjects showed homozygous frameshift or missense mutated alleles. We also studied the association between toxic response to 5FU and heterozygous frame shift or missense mutation of the DPYD gene among eight cancer patients who had received 5FU-based chemotherapy. These patients did not show any adverse effects higher than grade 3, suggesting that heterozygotes are not associated with increased toxicity to 5FU. Our results indicate that a very small percentage, about 0.2%, of the Japanese population seems to carry homozygous mutations in DPYD gene, mutations which possibly indicate genetically increased toxicity of 5FU-based chemotherapy. PMID- 11267946 TI - Antitumor activity and pharmacokinetics of TAS-106, 1-(3-C-ethynyl-beta-D-ribo pentofuranosyl)cytosine. AB - We examined the effects of dosage schedule on antitumor activity in vitro and in vivo to determine the optimal administration schedule for a new nucleoside antimetabolite 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)cytosine (ECyd, TAS 106). The cytotoxicity of TAS-106 in vitro against human tumors was evaluated at three drug exposure periods. TAS-106 exhibited fairly potent cytotoxicity even with 4 h exposure, and nearly equivalent and sufficiently potent cytotoxicity with 24 and 72 h exposures. These results suggest that long-term exposure to TAS 106 will not be required to achieve maximal cytotoxicity. The antitumor activity of TAS-106 in vivo was compared in nude rat models bearing human tumors on three administration schedules, once weekly, 3 times weekly, and 5 times weekly for 2 or 4 consecutive weeks. TAS-106 showed strong antitumor activity without serious toxicity on all three schedules, but the antitumor activity showed no obvious schedule-dependency in these models. When tumor-bearing nude rats were given a single i.v. dose of [(3)H]TAS-106, tumor tissue radioactivity tended to remain high for longer periods of time as compared to the radioactivity in various normal tissues. Furthermore, when the metabolism of TAS-106 in the tumor was examined, it was found that TAS-106 nucleotides (including the active metabolite, the triphosphate of TAS-106) were retained at high concentrations for prolonged periods. These pharmacodynamic features of TAS-106 may explain the strong antitumor activity without serious toxicity, observed on intermittent administration schedules, in nude rat models with human tumors. We therefore consider TAS-106 to be a promising compound which merits further investigation in patients with solid tumors. PMID- 11267947 TI - Overexpression of CDK2 is a prognostic indicator of oral cancer progression. AB - Cyclins and cyclin-dependent kinases (CDKs) play key roles in cell cycle regulation, a process of which dysregulation can lead to uncontrolled cell growth and hence to cancer. We have already reported the alteration of CDK4 and cyclin D1 expression in oral cancer. In this study, we examined by immunohistochemistry the expression of CDK2, and cyclins A and E in 20 normal oral mucosa, 42 dysplastic epithelia, and 103 oral squamous cell carcinomas (SCCs). The expressions of CDK2, and cyclins A and E were not detected in the normal epithelium and significantly altered from epithelial dysplasia to SCC. While there were no significant correlations between the expression of cyclins A, E and the patients' survival, CDK2 expression was significantly correlated with lymph node involvement (P = 0.025), tumor differentiation (P = 0.032), mode of tumor invasion (P = 0.017), and shorter survival period (P = 0.0173). These results suggest that the elevated expression of CDK2 is a critical factor in oral cancer progression and can be used as a negative predictive marker of the patients' prognosis. PMID- 11267950 TI - Inhibitors against factor VIII in patients with cancer. Analysis of 41 patients. AB - BACKGROUND: The spontaneous formation of neutralizing antibodies (inhibitors) to factor VIII (FVIII) in patients with cancer is a well known phenomenon. However, to the authors' knowledge there is lack of information in the literature with respect to the clinical course of these patients and the nature of the association between malignant tumors and acquired hemophilia. METHODS: A retrospective study of 41 patients with cancer and acquired hemophilia was conducted. The patients were identified through a MEDLINE search between 1974 2000. All patients had detailed clinical and laboratory information available and descriptions of the course of the inhibitor in relation to cancer treatment. The patients were divided into two groups: responders and nonresponders. The stage of the tumor, inhibitor titer, FVIII level, and survival data were examined and compared between the two groups. RESULTS: A total of 63 hemorrhagic episodes were reported in 25 patients with solid tumors and 16 patients with hematologic malignancies. The median inhibitor titer at the time of the diagnosis of acquired hemophilia was 14 Bethesda units (BU) (range, 1-435 BU). The complete response (CR) rate to treatment of the inhibitor was 70% and patients who achieved a CR were more likely to have early stage tumors (P = 0.0007) and a lower median inhibitor titer at the time of presentation compared with nonresponders (12 BU vs. 78 BU; P = 0.007). The overall survival was significantly higher in patients who achieved a CR compared with patients with a persistent inhibitor (75% vs. 17%; P = 0.0006). CONCLUSIONS: Although it remains an uncommon occurrence, the development of inhibitors to FVIII should be considered as a cause of bleeding in some patients with malignant diseases. Because of the high response rate and the impact of this type of hemorrhage on cancer patients, efforts should be directed toward immunosuppression of the inhibitor in a fashion similar to that used in other patients with acquired hemophilia. To our knowledge the link between malignancy and the formation of antibodies to FVIII is unclear; however, it appears that treatment of cancer with chemotherapy or surgery may accelerate the eradication of the inhibitor in some patients. Long term prospective studies are needed to better assess the association between cancer and acquired hemophilia. PMID- 11267948 TI - The predictive value of vascular endothelial growth factor and nm23 for the diagnosis of occult metastasis in non-small cell lung cancer. AB - We assessed the association of vascular endothelial growth factor (VEGF) and nm23 expression with occult micrometastasis in lung cancer. As destination sites for micrometastasis, we scrutinized lymph node (LN) and bone marrow (BM) specimens. For LN, 122 stage I patients who had received curative operations were studied. As regards BM, 203 patients in stage I - IV who underwent operations were registered. Immunohistochemical anti-cytokeratin staining was used to detect microdissemination of cancer cells. The VEGF and the nm23 expression at the primary sites were immunohistochemically studied in 285 cases in total. The percentages of the patients with microdissemination were 28.7% for LN and 42.4% for BM. The outcome for the patients with LN or BM microdissemination was significantly worse than that for patients without it. The increased VEGF and the decreased nm23 expression within primary tumors were significantly associated with LN and BM microdissemination. The results indicate possible value of using these biological markers to predict the risk of systemic micrometastasis in non small cell lung cancer. PMID- 11267951 TI - Yield of mammography in selected patients age < or = 30 years. AB - BACKGROUND: An outcomes analysis study was performed to quantify the benefit of directed diagnostic imaging of selected very young women (defined as < or = 30 years of age) in our population. Summary results are presented. PATIENTS AND METHODS: Women's Imaging Services were queried for studies performed between April 1, 1997 and December 31, 1998 on women < or = 30 years of age. The authors' referral pathway mandates breast examination by a general surgeon or by the head of Women's Imaging before mammography in all such patients. Studies were excluded if there were reviews of scans performed at other sites. The resulting 142 mammograms were evaluated. RESULTS: Ninety percent of the 142 studies were within normal limits. Only 11 mammograms indicated any required action (7.8%), and only 5 of these merited biopsy. All biopsies revealed benign disease. No carcinomas were detected by biopsy or on clinical follow-up in this cohort of women. These values are congruent with the scarce literature on mammography in this population. CONCLUSIONS: The yield of mammography in the age < or = 30 years population is low. PMID- 11267949 TI - Psychosocial factors as a potential trigger of oxidative DNA damage in human leukocytes. AB - Although numerous studies have been carried out on the stress-cancer linkage, the results are still inconclusive. One of the useful, but rarely applied, methods to assess this linkage is to examine the relationship between psychosocial stress and cancer-predisposing genetic alterations simultaneously. We investigated whether various psychosocial factors can be associated with the levels of 8 hydroxydeoxyguanosine (8-OH-dG), a biomarker of cancer-related oxidative DNA damage, in peripheral blood leukocytes in 362 healthy workers (276 males and 86 females). After adjustments for age, body mass index, cigarette smoking, and alcohol use, female subjects showed positive relationships between the amount of 8-OH-dG and the Tension-Anxiety, Depression-Rejection, Anger-Hostility, Fatigue, and Confusion scores of the Profile of Mood States, respectively. The levels of 8 OH-dG also increased reliably in the female subjects who had poor stress-coping behaviors, particularly wishful thinking strategy, in the NIOSH general job stress instrument. There were positive relationships of the 8-OH-dG levels to average working hours, a self-blame coping strategy, and recent loss of a close family member in male subjects. These findings in a nonclinical sample of healthy adults not only provide evidence of a stress-cancer linkage, but also suggest possible sex differences in the mechanisms of stress-related cancer initiation. PMID- 11267952 TI - Effect of adjuvant chemotherapy on outcome in patients with metastatic breast carcinoma treated with first-line doxorubicin-containing chemotherapy. AB - BACKGROUND: The objective of the current study was to analyze the impact of adjuvant chemotherapy in comparison with other prognostic parameters on the outcome of a series of patients with breast carcinoma at time of metastatic recurrence. METHODS: Data from 1430 patients accrued in 8 prospective trials of anthracycline-based first-line chemotherapy conducted at the Institut Curie between 1977 and 1992 were reviewed. RESULTS: Patients who had not received adjuvant chemotherapy had better response rates (66%) than pretreated patients (56%; P < 0.0001). Median overall survival rates after metastatic recurrence were 26 months compared with 19 months, respectively (P < 0.0001). Local and regional recurrences as well as the number of organ sites involved with metastatic disease were reduced in patients who had received adjuvant chemotherapy. In a multivariate analysis, the following parameters if present at the initiation of treatment were associated with poor outcome: elevated lactico dehydrogenase (LDH), low Karnofsky index, short disease free interval, more than two involved sites, liver involvement, and prior adjuvant chemotherapy. This adverse prognostic effect of prior adjuvant chemotherapy was independent of the type of drugs and of the duration of the treatment and was present even in the subgroup patients with prolonged disease free intervals longer than 48 months. CONCLUSIONS: Adjuvant chemotherapy adversely affects overall response rates and overall survival rates in patients with metastatic breast carcinoma treated with first-line anthracycline based chemotherapy. PMID- 11267953 TI - Salvage treatment for local recurrence after breast-conserving surgery and radiation as initial treatment for mammographically detected ductal carcinoma in situ of the breast. AB - BACKGROUND: The purpose of the current study is to evaluate the outcome of salvage treatment for local recurrence after breast-conserving surgery and radiation as initial treatment for mammographically detected ductal carcinoma in situ (DCIS; intraductal carcinoma) of the breast. METHODS: An analysis was performed of 42 patients with local only first failure (n = 41) or local-regional only first failure (n = 1) after breast-conserving surgery and radiation treatment had been given for DCIS of the breast. Surgical treatment at the time of local recurrence included mastectomy (n = 37; 88%) or excision (n = 5; 12%). Adjuvant systemic therapy at the time of local recurrence was chemotherapy (n = 3; 7%), tamoxifen (n = 8; 19%), both (n = 1; 2%), none (n = 29; 69%), or unknown (n = 1; 2%). The median interval from the time of initial treatment to local recurrence was 4.8 years (range = 1.0-15.2 yrs). The median follow-up after salvage treatment was 4.5 years (range = 0.2-12.8 yrs). RESULTS: At the time of the local recurrence, 22 patients (52%) had invasive ductal carcinoma, 18 patients (43%) had DCIS, 1 patient (2%) had invasive lobular carcinoma, and 1 patient (2%) had angiosarcoma. After salvage treatment, the rate of overall survival and the rate of cause specific survival for all 42 patients were 92% at both 5- and 8-years after treatment. The rate of freedom from distant metastases was 89% at 5 and 8 years. Favorable prognostic factors after salvage treatment were DCIS as the histology of the local recurrence and mammography only as the method of detection of the local recurrence. CONCLUSIONS: The results of salvage treatment in the current study demonstrated that local recurrences were salvaged with high rates of survival and freedom from distant metastases. These results support the use of breast-conserving surgery and radiation for initial management of DCIS of the breast. PMID- 11267954 TI - Exclusive radical surgery for esophageal adenocarcinoma. AB - BACKGROUND: Because very poor survival rates were reported after exclusive nonradical surgery, the current opinion in the medical community is that very few esophageal adenocarcinoma patients can anticipate long-term survival after esophagectomy. In the current study the ability of exclusive radical surgery including very extended lymph node dissection to provide a substantial percentage of patients with long-term survival was examined. METHODS: Radical esophagectomy (including removal of the esophageal tube, excision of the potentially involved locoregional lymph nodes, and skeletization of the nonresectable vital organs in the mediastinum and upper abdomen) was attempted in 183 consecutive patients with either Barrett (n = 77) or non-Barrett (n = 106) adenocarcinoma of the esophagus or cardia. Esophagectomy was subtotal (neck anastomosis) or distal (chest anastomosis) in 103 patients and 80 patients, respectively. RESULTS: Radical esophagectomy (Ro resection) was feasible in 137 patients (75%) whereas 46 patients (25%) in whom a part of the neoplastic process was not resectable (R1 or R2 resection) underwent a palliative esophagectomy. The 5-year survival, including in-hospital deaths (4.3%), was 35.3% for the whole series, 48% after Ro resection, and 0% after R1 or R2 resection. The 5-year survival rate after any R resection was 57.2% in patients with Barrett adenocarcinoma compared with 20% in patients with non-Barrett adenocarcinoma (P < 0.0001) because of a higher prevalence of nontransmural tumors (Tis through T2, N0) in the former group (56.5%) compared with the latter group (6.6%) (P < 0.0001). The 5-year survival was related closely to the magnitude of both wall penetration and extraesophageal neoplastic spread (Ro, Tis-T1-T2, N0 = 83.5% vs. Ro, T3, N0 = 44.4% vs. Ro, any T, N1 < 5 metastatic lymph nodes = 37% vs. Ro, any T, N1 > or = 5 metastastic lymph nodes = 6.8% vs. R1, R2 = 0%; P < 0.0001). CONCLUSIONS: Exclusive radical esophagectomy provides a chance of long-term survival in 35% of esophageal adenocarcinoma patients in whom it is attempted and nearly 50% of those patients in whom it is feasible. The presence of a small number of metastatic lymph nodes does not appear to preclude a long-term favorable outcome. PMID- 11267955 TI - Long term results of radioactive gold grain implantation for the treatment of persistent and recurrent nasopharyngeal carcinoma. AB - BACKGROUND: Brachytherapy is useful for the reirradiation of nasopharyngeal carcinoma. In the current study, the long term treatment results of permanent radioactive gold(198) grain interstitial implantation in patients with persistent and recurrent nasopharyngeal carcinoma were reviewed. METHODS: Gold grain implantation was performed under direct vision with a split palate approach to provide 60 grays (Gy) 0.5 cm away from the plane of implantation. Between August 1986 and May 1999, 106 patients were treated with gold grain implantation (45 patients for persistent disease, 53 patients for first recurrence, and 8 patients for second recurrence in the nasopharynx). All patients had histologically proven disease by biopsy before undergoing implantation. RESULTS: Patients with persistent disease and those with first recurrence did well with the gold grain implantation. The 5-year local control rates for patients with persistent disease, first recurrence, and second recurrence in the nasopharynx were 87.2%, 62.7%, and 23.4%, respectively (P = 0.0004). The 5-year metastasis free survival rates were 68.1%, 60.3%, and 40%, respectively, for the 3 groups (P = 0.048). The overall survival rates at 5 years for the 3 groups were 79.1%, 53.6%, and 42.9%, respectively (P = 0.0047). Patients with computed tomography evidence of disease extension outside the nasopharynx had a lower local control rate compared with patients whose disease was confined to the nasopharynx (5-year local control rate of 52% vs. 72.3%; P = 0.031). The size of the lesion was not found to be an independent prognostic factor for local control after implantation. Multivariate analysis showed only an indication for implantation (persistent disease, first recurrence, and second recurrence) to be a significant prognostic factor for local control. Complications attributed to gold grain implantation included headache, palatal fistula, and mucosal radiation necrosis at the site of implantation, and were reported to occur in 28.3%, 18.9%, and 16%, respectively, of patients. CONCLUSIONS: For selected patients with disease confined to the nasopharynx, gold grain implantation is an effective salvage treatment for persistent and recurrent nasopharyngeal carcinoma. PMID- 11267956 TI - Clinicopathological significance of tumor nest configuration in patients with esophageal squamous cell carcinoma. AB - BACKGROUND: Cancer-stromal interactions are an important mediator of cancer invasion and metastasis. METHODS: The authors investigated the clinicopathological significance of tumor nest configuration and the surrounding stroma in 159 patients with advanced esophageal squamous cell carcinoma (ESCC). The tumors were classified microscopically into two types. Type A tumors had oval shaped or sheet-like tumor nests (with > 80% of the tumor area showing these features). Type B tumors had asteroid-shaped or scattered small tumor nests (with > 20% of the tumor area showing these features). RESULTS: Of the 159 tumors examined, 38 (24%) were type A and 121 (76%) were type B. Type B tumors had a significantly deeper invasion depth, more frequent lymphatic permeation and lymph node metastasis, more prominent active fibroblastic stroma, and less frequent inflammatory cell infiltration (P < 0.05). Both univariate (P < 0.05) and multivariate (P < 0.05) analysis of the patients' survival showed that the prognosis for patients with type B tumors was significantly worse than for patients with type A tumors. CONCLUSIONS: This study showed that tumor nest configuration, which corresponded to the behavior of tumor cells against stromal cells, correlated well with the aggressiveness of the tumor. PMID- 11267957 TI - A clinical scoring system predicts the yield of diagnostic laparoscopy in patients with potentially resectable hepatic colorectal metastases. AB - BACKGROUND: Laparoscopy may identify occult metastatic disease and prevent unnecessary laparotomy in some patients with potentially resectable colorectal liver metastases but is unnecessary in the majority of individuals who undergo resection. The objectives of the current study were to assess the impact of laparoscopy after extensive preoperative imaging and to determine whether a preoperative clinical risk score can identify those patients most likely to benefit from the procedure. METHODS: Between December 1997 and July 1999, 103 consecutive patients with potentially resectable colorectal liver metastases underwent laparoscopy prior to planned laparotomy and partial hepatectomy. Surgical findings, length of hospital stay, and hospital charges were analyzed. Patients were assigned a clinical risk score (CRS) based on five factors related to the primary tumor and the hepatic disease. The likelihood of finding occult unresectable disease and the yield of laparoscopy were analyzed with respect to the CRS. RESULTS: Seventy-seven patients (75%) underwent resection. Laparoscopy identified 14 of 26 patients with unresectable disease, 10 of whom were spared an unnecessary laparotomy. In patients who underwent biopsy only, the laparoscopic identification of unresectable disease shortened the hospital stay (1.2 +/- 0.6 days vs. 5.8 +/- 2.3 days; p = 0.0001) and reduced the total hospital charges by 55% (P = 0.0001). The CRS predicted the likelihood of occult unresectable disease, which was 12% in those with a score < or = 2 but increased to 42% in those with a score > 2 (P = 0.001). If laparoscopy were used only in high risk patients (CRS > 2), 57 laparoscopies would have been avoided and the net savings doubled. CONCLUSIONS: With extensive preoperative imaging, the vast majority of patients with potentially resectable hepatic colorectal metastases do not benefit from laparoscopy. However, in the minority of patients with occult unresectable disease, laparoscopy prevents unnecessary laparotomy and reduces hospital stay and the total hospital charges. The CRS, previously shown to predict survival after hepatic resection, identifies those high risk patients most likely to benefit from laparoscopy and may improve resource utilization. PMID- 11267958 TI - Increased expression of laminin-5 and its prognostic significance in lung adenocarcinomas of small size. An immunohistochemical analysis of 102 cases. AB - BACKGROUND: Laminin-5 plays an important role in cell migration during tissue remodeling and tumor invasion. METHODS: The authors studied the expression of laminin-5 immunohistochemically in 102 cases of small-sized lung adenocarcinoma (maximum dimension < or = 2 cm) using a monoclonal antibody against the laminin gamma2 chain, and they also investigated the associations of laminin-5 with clinicopathologic characteristics. Prognostic significance of increased laminin-5 expression was evaluated using the Kaplan-Meier method and the Cox proportional hazard model. RESULTS: Overall, laminin-5 expression was observed in 82 cases (80.4%): 7 of 18 (38.9%) bronchioloalveolar carcinomas and 75 of 84 (89.3%) invasive adenocarcinomas. Laminin-5 was preferentially localized in the cytoplasm of tumor cells at the tumor-stromal interface, where budding or dissociation of cancer cells was frequently observed. Overexpression of laminin-5 (24 cases, 23.5%) was associated with vascular invasion (P = 0.021) and stromal fibroblastic reaction (P = 0.005) but not with nodal involvement, lymphatic invasion, or pleural invasion. Survival analysis revealed that overexpression of laminin-5 was associated with shorter patient survival (P = 0.0027 by log rank test). On multivariate analysis, overexpression of laminin-5 was an independent prognostic factor (P = 0.030), as were nodal involvement (P < 0.0001), vascular invasion (P = 0.047), and lymphatic invasion (P = 0.0047) in a whole cohort of patients. Moreover, when patients with Stage I (International Union Against Cancer [UICC] staging system) disease were considered in multivariate analysis, overexpression of laminin-5 was the only significant prognostic factor (P = 0.022), whereas vascular invasion had a marginally significant impact (P = 0.07) on patient survival. CONCLUSIONS: The authors' results showed that laminin-5 is frequently expressed by cancer cells at the invasive front of lung adenocarcinoma. The study concluded that overexpression of laminin-5 may be a useful prognostic factor in patients with small-sized lung adenocarcinoma, especially in Stage I cases. PMID- 11267959 TI - Endoscopic evaluation of centrally located early squamous cell carcinoma of the lung. AB - BACKGROUND: Lung-sparing treatment recently has become a choice in the treatment of patients with early hilar lung carcinoma. To select the method of treatment, it is important to evaluate the histologic extent of the tumor using endoscopy. METHODS: A total of 46 patients who underwent surgery for an endoscopically evaluated early lung carcinoma of the tracheobronchial tree were analyzed. Initial surgery was performed in 16 patients and in 30 patients surgery was performed after preoperative laser therapy. The endoscopic findings were classified into three types: superficial, nodular, and polypoid. In the resected lung, the greatest tumor dimension, the depth of mural invasion, the presence of microscopic blood and lymph vessel invasion, and metastases to the dissected lymph nodes were examined. RESULTS: The mean greatest tumor dimensions were 14.6 +/- 7.3 mm (mean +/- standard deviation) in the nodular group, 23.3 +/- 12.8 mm in the superficial group, and 19.0 +/- 9.2 mm in the polypoid group. The greatest tumor dimension in the superficial group was significantly larger than that in the nodular group (P < 0.01). Preoperative endoscopic diagnosis of early hilar lung carcinoma was correct histologically in 34 of 46 cases (74.0%). Hilar lymph node metastases (N1), extrabronchial invasion, and extension to the peripheral bronchus were recognized in 12 cases. Complete disappearance of the tumor due to preoperative laser therapy was confirmed in eight patients. Lymph node metastasis was not found when the greatest tumor dimension measured < 8 mm. The overall absolute 5-year survival rate was 76.0% for all patients, 87.1% for the surgery alone group, and 70.0% for the surgery after preoperative laser therapy group. CONCLUSIONS: Curative treatment of early hilar lung carcinoma is possible using photodynamic therapy alone when the tumor size is < or = 8 mm and the lesion does not extend to the peripheral bronchus. PMID- 11267960 TI - Gender and other survival predictors in patients with metastatic melanoma on Southwest Oncology Group trials. AB - BACKGROUND: Some studies have suggested that women with metastatic malignant melanoma have a better survival rate than men. However, little is known about the effect of gender on survival in combination with other clinical variables and treatment variables. Thus, an analysis of 813 eligible patients from 15 consecutive Southwest Oncology Group (SWOG) Phase II or III trials evaluating chemotherapy or chemoimmunotherapy for metastatic melanoma was performed. METHODS: A multivariate Cox regression model was used. RESULTS: Poor performance status (P < 0.001), more organ sites with metastases (OSM) (P < 0.001), liver involvement (P < 0.001), and nonliver visceral involvement (P = 0.01) were highly significant predictors of worse survival, whereas the disease free interval (P = 0.08) had borderline significance. After adjustment for all factors, there was no difference in overall survival between men and women (P = 0.19). Women had a longer disease free interval (P = 0.003) and fewer OSM (P = 0.004) at study registration than men. CONCLUSIONS: The current study found that performance status, OSM, and type of visceral involvement were independent predictors of survival in patients with metastic malignant melanoma and should be used as stratification factors in future Phase III trials. However, the current study also found that gender did not appear to be a significant independent predictor of survival for this stage of disease. A longer disease free interval from initial diagnosis and fewer OSMs may partly explain the improved outcome reported for women in selected trials. The study concluded that further investigation of the biologic differences at early stage diagnosis should be undertaken to determine whether women truly have a different pace of disease progression and a different metastatic pattern. PMID- 11267961 TI - Critical role of extracellular signal-regulated kinase phosphorylation on menadione (vitamin K3) induced growth inhibition. AB - BACKGROUND: Although it is widely known that the extracellular signal-regulated kinase (ERK) pathway stimulates cell growth and protects cells from death, recent findings have proposed a proapoptotic action of ERK phosphorylation. Because the authors found that vitamin K3 (VK3) was a potent growth inhibitor and an inducer for ERK phosphorylation through a specific pathway in the stomach cancer cell line, the critical role of ERK phosphorylation in VK(3)-mediated growth inhibitory effect was examined. METHODS: The fluorochrome Hoechst 33258 assay (Hoechst AG [now Aventis] Frankfort, Germany) was used for counting cells (excitation at 360 nm; emission at 460 nm). For two-dimensional electrophoresis, cells were dissolved in urea standard buffer and applied first to isoelectronic focusing gels. Cell lysates were subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) using 10% polyacrylamide gels. To examine the phosphorylation of receptors, cell lysates were immunoprecipitated with receptor antibody. RESULTS: VK3 induced phosphorylation of hepatocyte growth factor receptor (c-met), epidermal growth factor receptor (EGFR), or external signal-regulated kinase (ERK), which increased progressively to a maximum level at 30 minutes, in a dose-dependent manner and occurred at growth inhibitory concentrations. VK(3)-mediated growth inhibition and protein tyrosine phosphorylation were nullified completely by glutathione or L-cysteine but not by nonthiol antioxidants, thus suggesting that sulfhydryl arylation might have been involved in VK(3)-mediated action. The phosphorylation of EGFR and c-met by VK(3) appeared to be functional, because these were coimmunoprecipitated with growth factor receptor-bound protein 2 (Grb2) and SOS1 antibody. Epidermal growth factor (EGF) or hepatocyte growth factor (HGF) stimulated increase of cyclin D1 protein after 12 hours and increased DNA content after 3 days in culture. In addition, U0126, which is a potent inhibitor for ERK phosphorylation, antagonized increase of cyclin D1, thus suggesting that EGF- or HGF-mediated ERK phosphorylation might have played an essential role for cell growth. By contrast, ERK phosphorylation by VK3 was more prolonged and intense than the signal induced by the growth factors. U0126 reduced ERK phosphorylation and prevented growth inhibition by VK3. Two-dimensional gels showed VK(3)-induced additional phospho-ERK spots, compared with those obtained from growth factors. This extra spot was completely antagonized by U0126. CONCLUSIONS: VK(3)-induced growth inhibition and protein tyrosine phosphorylation were mediated by the sulfhydryl arylation system. The tyrosine phosphorylation of EGFR or c-met by VK3 activated the Ras signaling pathway. The overexpressed ERK phosphorylation by VK3 seemed to originate from additional spots on two-dimensional gels, which played a critical role in VK(3) induced growth inhibitory action despite the fact that ERK phosphorylation by growth factors had had an essential association with cell growth. PMID- 11267962 TI - Phase I study of CI-958 in children and adolescents with recurrent solid tumors. AB - BACKGROUND: CI-958 is a synthetic intercalating agent of a new chemical class, the benzopyranoindazoles, with promising preclinical activity. Its mechanism of action is thought to be stabilization of the cleavable complex of DNA with topoisomerase II, as well as DNA helicase blockade. It is thought to have less cardiotoxicity than the anthracyclines. Early Phase I studies in adults showed the drug to be well tolerated, making it an attractive agent to pursue in Phase I clinical trials in children. METHODS: Children and adolescents with recurrent solid tumors received CI-958 at an initial dose of 450 mg/m(2) over 2 hours. Dose escalation was performed in a standard fashion in cohorts of three patients until dose limiting toxicity and the maximum tolerated dose were determined. RESULTS: Twenty-one patients were entered on the study. The maximum tolerated dose was found to be 650 mg/m(2). Dose limiting toxicities were Grade 4 neutropenia and Grade 4 hypotension at the dose level of 700 mg/m(2). CONCLUSIONS: The maximum tolerated dose of CI-958 in children and adolescents is 650 mg/m(2). No antitumor activity has been observed. PMID- 11267964 TI - Influence of Hispanic ethnicity on outcome after resection of carcinoma of the head of the pancreas. AB - BACKGROUND: Poor outcomes in Hispanic patients have been reported for tumors at a number of sites. The authors sought to determine whether a similar phenomenon occurs in Hispanics after the resection of solid epithelial tumors of the head of the pancreas. METHODS: Between 1983-1995, 273 patients with noncystic epithelial carcinoma of the head of the pancreas were evaluated. Resection was accomplished in 104 patients (38%); these patients were the focus of the current retrospective review. Of the patients who underwent resection, 26 (25%) were Hispanic and 78 (75%) were non-Hispanic. RESULTS: Although Hispanic patients tended to present at a significantly younger age and their serum bilirubin level was significantly higher, no other differences in clinical characteristics were observed. After resection, Hispanic patients had a median survival of only 11.4 months, whereas the non-Hispanic group had a median survival of 21.7 months (P = 0.009). Hispanic ethnicity, as well as age > 74 years and jaundice at the time of presentation also were found to be significant prognostic factors on multivariate analysis. Hispanic patients did not present with more advanced disease and no delays in assessment by a physician or in proceeding to surgery were observed. Furthermore, the rate of resection was the same in Hispanic patients and non-Hispanic patients. Long-term survival after palliative bypass was similarly worse in the Hispanic subgroup. CONCLUSIONS: Hispanic patients treated at the study center appeared to have a diminished survival after resection of a tumor of the head of the pancreas. No treatment-related factors were identified that could explain this discrepancy in outcome. PMID- 11267963 TI - Prognostic factors and imaging patterns of recurrent pulmonary nodules after thoracotomy in children with osteosarcoma. AB - BACKGROUND: In children with osteosarcoma who have undergone thoracotomy, it often is difficult to distinguish metastatic from benign recurrent pulmonary nodules. The authors of this study sought to identify any computed tomography (CT) imaging pattern of recurrent pulmonary metastases in this patient population. The authors also sought to identify associated prognostic factors. METHODS: CT scans obtained after thoracotomy were available for 35 patients with osteosarcoma who had undergone resection of presumed pulmonary metastases at St. Jude Children's Research Hospital (Memphis, TN). CT scans obtained before the initial thoracotomy were available for 33 of the 35. The authors recorded location, histologic diagnosis, and time of development of the original pulmonary nodules, time of recurrence of pulmonary disease; the location of recurrent nodules, and the presence of calcification, adenopathy, or progressive pleural disease, as well as patient demographic data, survival data, and location of the primary tumor site. RESULTS: Pulmonary nodules recurred in 32 of the 35 patients after thoracotomy. Nineteen of these patients underwent resection of the recurrent lesions and 1 who died underwent an autopsy; 18 of the 20 patients had metastatic disease. The only CT finding consistently associated with recurrent metastatic disease was progressive pleural thickening, which predicted a poor outcome. The occurrence of a solitary pulmonary nodule in the lung contralateral to the previous surgery was associated almost always with a benign process. CONCLUSIONS: CT imaging cannot distinguish reliably between benign and metastatic recurrent pulmonary disease after thoracotomy in patients with osteosarcoma. Recurrent pulmonary disease in this set of patients is likely to be metastatic, and aggressive surgical intervention is probably warranted. In this study, patients who had progressive pleural disease after thoracotomy consistently experienced pulmonary metastatic recurrence and had a poor prognosis. PMID- 11267965 TI - Brachytherapy. Results of two different therapy strategies for patients with primary glioblastoma multiforme. PMID- 11267967 TI - Enhancement of anti-tumor immunity by tumor cells transfected with the secondary lymphoid tissue chemokine EBI-1-ligand chemokine and stromal cell-derived factor 1alpha chemokine genes. AB - Several new lymphocyte-specific chemokines, which attract naive and memory T cells, B cells, dendritic cells and natural killer cells, have been isolated. We have found evidence of the anti-tumor effects of 3 major lymphocyte-specific chemokines, secondary lymphoid tissue chemokine (SLC), EBI-1-ligand chemokine (ELC) and stromal cell-derived factor (SDF)-1alpha, in murine models (Meth A fibrosarcoma and HM-1 ovarian tumor). In both naive and immunized mice, tumors expressing SLC, ELC or SDF-1alpha showed delayed progression compared with control tumors. In mice immunized with tumor cells expressing 1 of these 3 chemokine genes, challenge with parental tumor cells resulted in slightly slower progression than in control mice, while in mice immunized with tumor cells transfected to co-express IL-2 or granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as these chemokines, all tumors regressed. Furthermore, spleen cells from mice immunized with these "double-transfected" tumor cells exhibited higher proliferative responses and greater cytotoxic activity against parental tumor cells. These anti-tumor effects were associated with profound alterations in the leukocyte populations within the tumors and regional lymph nodes, and this was due to activation of type I T cell-dependent responses that produced high levels of IFN-gamma. These findings show that SLC, ELC and SDF 1alpha enhance anti-tumor immunity both systemically and locally and that these chemokines may be clinically useful, especially when combined with IL-2 and GM CSF. PMID- 11267968 TI - Genetic disruption of host nitric oxide synthase II gene impairs melanoma-induced angiogenesis and suppresses pleural effusion. AB - Our previous study showed that genetic disruption of nitric oxide (NO) synthase II (NOS II) expression inhibits the metastatic ability of non-immunogenic B16 melanoma cells in syngeneic mice. In the present study, the mechanisms for this metastasis suppression were determined. B16-BL6 and B16-F10 murine melanoma cells were injected i.v. into syngeneic wild-type (NOS II(+/+)) and NOS II-null (NOS II(-/-)) C57BL/6 mice. Both melanoma cells produced less and smaller experimental pulmonary metastases in NOS II(-/-) mice than in NOS II(+/+) mice. Moreover, less metastatic pleural effusion was observed in NOS II(-/-) mice than in NOS II(+/+) mice. Immunohistochemical analyses indicated that absence of NOS II expression was correlated with decreased vascular endothelial growth factor expression and tumor-associated vascular formation. After activation with lipopolysaccharide and IFN-gamma, neither melanoma cell line produced detectable levels of NO. Our data demonstrate that tumor-induced expression of host NOS II enhances melanoma metastasis and pleural effusion, at least in part, through regulation of vascular formation and vascular permeability. PMID- 11267969 TI - Natural T-helper immunity against human papillomavirus type 16 (HPV16) E7-derived peptide epitopes in patients with HPV16-positive cervical lesions: identification of 3 human leukocyte antigen class II-restricted epitopes. AB - Tumor-specific T-helper (Th) immunity was found to play a pivotal role in the natural and vaccine-induced immune defense against tumors. Since the majority of cervical cancers express human papillomavirus type 16 (HPV16) E7 oncoprotein, it is important to investigate the Th response against this target antigen in detail. By means of PBMC cultures from HLA-typed healthy donors, we identified the central part of HPV16 E7 (E7(41-72)) as the major immunogenic region within this antigen. Furthermore, we mapped 3 distinct Th epitopes within this region (DR15/E7(50-62), DR3/E7(43-77), DQ2/E7(35-50)). In a parallel approach, employing IFN-gamma ELISPOT analysis, we detected Th immunity against HPV16 E7 in subjects with HPV16+ lesions. Several of these responses matched with the 3 Th epitopes defined in our study. A number of other HPV16+ subjects did not display any E7 specific type 1 cytokine-producing T-cell immunity, indicating failure of the immune response. Our combined data argue for more extensive as well as longitudinal analysis of HPV16-specific T-cell immunity using the ELISPOT assay described, as well as for HPV-specific vaccination of individuals with HPV+ lesions. PMID- 11267970 TI - Microsatellite instability in synchronous gastric carcinomas. AB - Synchronous gastric carcinomas are found in 4% to 10% of all gastric carcinomas, and the tumor multiplicity is believed to be related to genetic predisposition. To investigate the role of mismatch repair error in synchronous gastric carcinomas, we analyzed the microsatellite instability (MSI) status of 101 cancers from 48 gastrectomy specimens and compared them with 149 solitary gastric carcinomas. Multiple synchronous gastric carcinomas are characterized by slightly older age, predominance in males, early stage and lower lymph node metastasis. Among the 48 cases, 8 (18 lesions) were associated with a gastric adenoma (type I) and 40 (83 lesions) were not associated with a gastric adenoma (type II). The MSI+ rate was 50% in the type I and 8.4% in the type II synchronous gastric carcinomas (p < 0.001), while that of solitary gastric carcinomas was 9.4%. In addition, the frameshift mutation rates of the TGF-betaRII, BAX and hMSH3 genes in the type I synchronous carcinomas were higher than those in the type II synchronous carcinomas. These findings indicate that a defect in the mismatch repair system might play a role in the carcinogenesis of a minor subset of multiple gastric carcinomas associated with adenomas. PMID- 11267971 TI - Overexpression of JunB in undifferentiated malignant rat oral keratinocytes enhances the malignant phenotype in vitro without altering cellular differentiation. AB - Our study examined the expression of AP-1 family members in keratinocytes derived from the rat-4NQO model of oral carcinogenesis in which extremes of epithelial differentiation and tumour cell aggressiveness are evident. The constitutive expression of JunB was diminished in the undifferentiated, more aggressive tumour phenotype compared with the well-differentiated, less aggressive keratinocytes, whereas the expression of other AP-1 family members (c-jun, junD, c-fos, fra1, fra2 and fosB) was either very weak or variable. After transfection of the undifferentiated keratinocytes with junB cDNA, clonal populations were isolated that expressed similar levels of JunB protein as the well-differentiated cells. Both untransfected and transfected cell lines were keratin negative and vimentin positive. Increased expression of JunB in the transfected cells resulted in up regulation of c-Jun and Fra1 and an enhanced AP-1 activity as demonstrated by transcriptional activation of the prototypic AP-1 dependent promoter, MMP-1. JunB transfected cells grew more quickly than vector-only controls and were refractory to the growth inhibitory effects of TGF-beta1. Over-expression of JunB resulted in the elevated expression of the AP-1 dependent proteinase, MMP-9, whereas the expression of the AP-1 independent enzyme, MMP-2, was unaffected. JunB transfected keratinocytes were highly invasive in an in vitro assay of tumour cell invasion compared with vector controls. The results indicate that increased expression of JunB above baseline levels in undifferentiated rat keratinocytes does not alter epithelial differentiation but enhances the malignant phenotype in vitro, possibly by altering the dynamics of the AP-1 complex. PMID- 11267972 TI - Elevated expression of UDP-N-acetylglucosamine: alphamannoside beta1,6 N acetylglucosaminyltransferase is an early event in hepatocarcinogenesis. AB - Previous reports have suggested that changes in oligosaccharide structures, especially beta1-6 branching in N-glycans, which are biosynthesized by UDP-N acetylglucosamine:alpha mannoside beta1,6 N-acetylglucosaminyltransferase (GnT V), are linked to tumor metastasis and invasion. In the present study, we investigated GnT-V expression in human hepatocellular carcinoma (HCC) tissues. High expression of GnT-V mRNA was observed in both HCC and the surrounding tissues but not in normal liver. Immunohistochemical study using a newly established monoclonal antibody against GnT-V revealed that positive staining of GnT-V was observed in 75% of HCC tissues and 60% of surrounding tissues and that liver cirrhosis showed much stronger staining of GnT-V than chronic hepatitis without liver cirrhosis (p = 0.0035). In contrast, all of 12 cases of atypical adenomatous hyperplasia diffusely expressed GnT-V. beta1-6 branching in N glycans, products of GnT-V, was increased in HCC tissues with high expression of GnT-V, as judged by lectin blotting. Levels of GnT-V expression in HCC tissues were positively correlated with a low Ki-67 labeling index (p = 0.0009), small size (p < 0.0001), poor differentiation (p < 0.0001) and absence of portal invasion (p = 0.018). Furthermore, HCC cases with low or no expression of GnT-V were more likely to show recurrence than cases with high expression (p = 0.0373). These findings strongly suggest that GnT-V expression is concerned mainly with an early phase of hepatocarcinogenesis. PMID- 11267973 TI - Expression and activity of matrix metalloproteases in human malignant mesothelioma cell lines. AB - The extracellular matrix metalloproteases (MMPs) secreted by various human tumor cells play a crucial role in tumor cell invasion and metastasis, but their expression in malignant mesothelioma (MM) cells has not been examined. In this study, we have investigated the spectrum of MMPs and tissue inhibitors of metalloproteases (TIMPs) produced by 8 MM cell lines. Using RT-PCR, we found that all investigated MM cell lines expressed genes encoding mRNA for MMP-1 (interstitial collagenase), MMP-2 (gelatinase A), MMP-3 (stromelysin-1), MMP-9 (gelatinase B) and TIMPs 1, 2 and 3. We also found that 6/8 MM cell lines expressed MMP-7 (matrilysin) and 3/8 MM cell lines expressed MMP-10 (stromelysin 2). MMP-11 (stromelysin-3) was not detected in any of the MM cell lines. Production of MMP-2 and MMP-9 was confirmed using gelatin zymography. In addition, all MM cell lines secreted a 66 kDa metalloprotease, while 3/8 MM cell lines secreted 46, 48, 51 and 63 kDa metalloproteases which specifically degraded the extracellular matrix components fibronectin, vitronectin and laminin. The 66 kDa protease was identified as MMP-3 by Western blot. Our results reveal a broad spectrum of MMPs and TIMPs produced by MM cells and indicate that different substrate specificities of MMPs may play a role in MM cell invasion. PMID- 11267974 TI - Tissue-specific alternate splicing of human telomerase reverse transcriptase (hTERT) influences telomere lengths during human development. AB - Direct genetic manipulations have shown that telomerase-mediated telomere elongation plays a key role in determining cellular replicative capacity and senescence. The mechanisms regulating the production of an active telomerase enzyme are still predominantly unknown, although roles for transcriptional control of hTERT, alternative-splicing of hTERT transcripts, and post translational phosphorylation of hTERT protein have been advocated. Here we show that hTERT is alternatively spliced in specific patterns by different tissue types during human development. Alternative splicing often prohibits the expression of hTERT protein containing functional reverse transcriptase domains. In these instances, telomerase activity is absent, leading to shortening of telomeres. The specific pattern of hTERT mRNA variants in human development provides evidence that alternative splicing is non-random and participates in the regulation of telomerase activity. PMID- 11267975 TI - Human breast-cancer cells stimulate the fusion, migration and resorptive activity of osteoclasts in bone explants. AB - A central event in bone resorption is the recruitment of osteoclasts to future resorption sites. Breast-cancer cells invariably metastasise to the skeleton and induce extensive bone destruction by osteoclasts. However, our understanding of the mechanisms by which cancer cells interact with osteoclasts remains unclear. Consequently, we compared the effects of conditioned medium (CM) from 2 human breast-cancer cell lines, MB-MDA-231 and MCF-7, with those of a normal human breast epithelial cell line, HME, on osteoclastic fusion, resorptive activity and migration from the periosteum to the developing marrow cavity of fetal mouse metatarsals in culture. Osteoclastic resorptive activity was assessed by pre labelling 17-day-old fetal metatarsal explants with 45Ca, whilst fusion and migration were monitored by histomorphometry and osteoclasts were identified by their tartrate-resistant acid phosphatase activity. CM from TPA-stimulated breast cancer cell lines produced a significant increase in osteoclastic resorptive activity, whilst the normal breast cell line produced a minimal increase. The breast-cancer cell lines also stimulated osteoclastic fusion and migration in the metatarsal explants, but the normal breast cell line was without effect. The stimulatory effect of CM from MDA-MB-231 cells on osteoclastic fusion, but not migration, was partially inhibited by preventing prostaglandin and leukotriene synthesis by cells within the bone explants. In contrast, a synthetic matrix metalloproteinase (MMP) inhibitor, but not a cysteine proteinase inhibitor, prevented the migration of osteoclasts to the calcified centre of the metatarsal explants in response to CM from MDA-MB-231 cells. MDA-MB-231 cells also induced an increase in the expression of MMP-9 by migrating osteoclasts. Fractionation of the TPA-stimulated breast cancer cell CM established that the resorptive activity was associated with factors of m.w. >3 kDa. We determined by immuno-assay that human breast-cancer cells secrete parathyroid hormone-related protein (PTH-rP), tumour necrosis factor-alpha (TNF-alpha) and interleukins (ILs) 6 and 11. Neutralizing experiments with human antibodies to these cytokines established that PTH-rP and TNF-alpha production by MDA-MB-231 cells were responsible for mediating their effects on osteoclastic migration and ultimately bone resorption in the metatarsal explants. PMID- 11267976 TI - Modulation of cell cycle-related protein expression by sodium butyrate in human non-small cell lung cancer cell lines. AB - To elucidate the mechanism of action of sodium butyrate (NaB), we examined its effect on the expression of some cell cycle-related proteins (cyclins D1 and E, p16(ink4), p21(waf1), p27(kip1)) in 2 human non-small cell lung cancer cell lines (NCI-460 and NCI-H23) characterized by wild- type and mutant TP53, respectively. The growth of both cell lines was inhibited in a dose-dependent manner and this process was accompanied by a modulation of cell cycle-related proteins. In NCI H460, the p27(kip1) and p16(ink4) protein levels were markedly increased following NaB treatment, whereas p21(waf1) was only slightly elevated, with a peak at 2 mM NaB, and p53 was unaffected by any concentration. By contrast, in NCI-H23, a marked increase in p21(waf1) protein was paralleled by decreased p53 levels, whereas all the other investigated proteins remained stable. The results suggest that NaB blocks the growth of both cell lines by induction of cyclin dependent kinase inhibitors (in particular, p21(waf1) in NCI-H23 and p27(kip1) and p16(ink4) in NCI-H460) through a p53-dependent or p53-independent mechanism, and open up interesting perspectives for the use of NaB as an alternative or additional strategy in the treatment of non-small cell lung carcinoma. PMID- 11267978 TI - Suppression of proline-directed protein kinase F(A) systemically inhibits the growth of human acute leukemia cells. AB - Initial studies revealed that proline-directed protein kinase F(A) (PDPK F(A)) was overexpressed in various cancerous tissues relative to normal controls. However, the functional role of overexpressed PDPK F(A) in cancer remains to be established. In this report, we explore the potential role of PDPK F(A) in leukemia cell growth by investigating the effects of partial inhibition of this kinase on human acute promyelocytic leukemia (HL-60) and acute lymphoblastic leukemia (Jurkat) cells. Cloning of PDPK F(A) cDNA and its recombinant antisense expression vector and antibody were successfully developed. Several stable antisense clones of HL-60 and Jurkat cells were subcloned, which expressed a low level of PDPK F(A) when compared with the control-transfected clone in immunoblot analysis. Moreover, these antisense clones potently inhibited cell growth, clonogenic growth in soft agar and serum-independent growth. The results taken together demonstrate that suppression of PDPK F(A) is able to interfere with the growth of HL-60 and Jurkat cells, suggesting an essential role of this PDPK in human acute leukemia cell growth. PMID- 11267977 TI - Telomerase in relation to clinicopathologic prognostic factors and survival in cervical cancer. AB - We investigated, in cervical cancer, the relation between telomerase activity, telomerase RNA (hTR) and mRNA of the catalytic subunit of telomerase, hTERT, with "classic" clinicopathological factors as well as survival. Frozen specimens were obtained from 107 consecutive patients with cervical cancer, treated with surgery or radiotherapy with or without chemotherapy. Telomerase activity was determined with fluorescence-based TRAP and hTR and hTERT with semi-quantitative RT-PCR. Eight normal cervical specimens served as controls. Analysis of prognostic factors and survival was limited to early-stage patients, treated primarily with radical hysterectomy. Telomerase activity was not detected in normal cervices and was present in 85 of 107 (79%) cervical cancers (p < 0.001). hTR was detected in all normal cervices and cervical cancers, while hTERT mRNA was detected in 1 of 8 (13%) normal cervices and in 83 of 104 (80%) cervical cancers (p < 0.001). In contrast to semi-quantitative hTR expression levels, semi-quantitative hTERT mRNA levels were related to telomerase activity levels (p < 0.01). In all patients, telomerase activity levels were related to differentiation grade (p < 0.05) but not to stage and histotype. In early-stage patients, telomerase activity, hTR and hTERT were not related to tumor volume, vascular invasion or presence of metastatic lymph nodes. Tumor volume, vascular invasion and presence of metastatic lymph nodes were related to (progression-free) survival, while telomerase activity and its subunits were not. Frequent up-regulation of telomerase activity and hTERT mRNA is especially observed in cervical cancers, while hTR is also detected in normal cervices. Telomerase is not applicable as a prognostic factor in early-stage cervical cancer patients. PMID- 11267979 TI - Silencing of the caspase-1 gene occurs in murine and human renal cancer cells and causes solid tumor growth in vivo. AB - Renal cell cancer is a unique solid tumor that occasionally shows spontaneous regression even at an advanced stage, of which the underlying mechanism is not well understood. To investigate a potential role of the pro-apoptotic molecule caspase-1 in the growth regulation of renal cell cancer, we created transfectants expressing exogenous caspase-1 from a murine renal cancer cell line, Renca. Overexpression of caspase-1 did not affect the growth of Renca cells in vitro at the exponential phase but induced apoptotic cell death at 50% to 75% confluence, whereas control cells underwent apoptosis only after reaching 100% confluence. When implanted into the flank of a syngeneic BALB/c mouse, caspase-1 overexpressing Renca cells did not effectively establish growth as a solid tumor, forming a measurable tumor in only 7 of 11 (64%) animals, whereas control cells formed a tumor in 6 of 6 (100%) animals. The growth of tumors from caspase-1 overexpressing cells slowed down markedly after the tumors reached 5 to 10 mm in diameter, and histological examination of such tumors revealed numerous apoptotic cells positively stained by TUNEL assay. Interestingly, endogenous caspase-1 was not detected in the tumors from control cells, which re-expressed caspase-1 when they were re-cultured and exposed to a demethylation reagent, 5-aza-2' deoxycytidine. Furthermore, treatment of a human renal cancer cell line, ACHN, with 5-aza-2'-deoxycytidine also caused recovery of caspase-1 expression, which was not detected before treatment. These data suggest that silencing of caspase-1 through DNA methylation may be involved in the oncogenesis of some renal cell cancers growing as a solid tumor. PMID- 11267980 TI - Comparative genomic hybridization (CGH) analysis of stage 4 neuroblastoma reveals high frequency of 11q deletion in tumors lacking MYCN amplification. AB - We have studied the occurrence and association of 11q deletions with other chromosomal imbalances in Stage 4 neuroblastomas. To this purpose we have performed comparative genomic hybridization (CGH) analysis on 50 Stage 4 neuroblastomas and these data were analyzed together with those from 33 previously published cases. We observed a high incidence of 11q deletion in Stage 4 neuroblastoma without MYCN amplification (59%) whereas 11q loss was only observed in 15% of neuroblastomas with MYCN-amplification (p = 0.0002) or 11% of cases with 1p deletion detected by CGH (p = 0.0001). In addition, 11q loss showed significant positive correlation with 3p loss (p = 0.0002). Event-free survival was poor and not significantly different for patients with or without 11q deletion. Our study provides further evidence that Stage 4 neuroblastomas with 11q deletions represent a distinct genetic subgroup that typically shows no MYCN amplification nor 1p deletion. Moreover, it shows that neuroblastomas with 11q deletion also often present 3p deletion. This genetic subgroup shows a similar poor prognosis as MYCN amplified 4 neuroblastomas. PMID- 11267981 TI - Mismatch repair and microsatellite instability in esophageal cancer cells. AB - Using in vitro mismatch repair (MMR) assay, we have identified 3 of 22 esophageal cancer cell lines exhibiting reduced MMR activity. By means of gel-shift assay, decreased binding ability to GT mismatch and CA loop was observed in these 3 cell lines. However, we could not find any mutations in the hMSH2, hMSH3 and hMSH6 genes, the protein products of which exhibit mismatch binding activity in human cells. In addition, when using antibodies against 5 MMR-related proteins (hMSH2, hMSH3, hMSH6, hPMS2 and hMLH1), no aberrant expression was detected in any of them. When we examined 9 microsatellite loci in endogenous genomic DNA, these 3 esophageal cancer cell lines, deficient in MMR, did not exhibit microsatellite instability. However, when we examined the repetitious sequence on exogenous plasmid DNA which was introduced into these 3 esophageal cancer cells, the results suggested that MMR deficiency in esophageal cancer cells could result in moderate instability of the exogenous sequence. PMID- 11267982 TI - Antibodies to Epstein-Barr virus thymidine kinase: a characteristic marker for the serological detection of nasopharyngeal carcinoma. AB - Patients suffering from nasopharyngeal carcinoma (NPC) generally exhibit elevated serum IgA antibody titres to Epstein-Barr virus (EBV) early antigen (EA) and virus capsid antigen (VCA). This property is frequently used as a diagnostic aid. Preliminary experiments suggested that an ELISA for IgA antibodies against the EBV-encoded thymidine kinase (TK) could form the basis of a more reliable diagnostic test. Here, we describe the construction of a recombinant baculovirus that expresses the EBV TK and present a full analysis of its use in serological surveys of NPC patients. Baculovirus-derived TK was used to develop a simple ELISA for serum IgA against this antigen. ELISA reactivity was strongly associated with NPC compared with an EBV-positive, normal control population. Comparison with the existing IgA-VCA and EA assays showed that the TK ELISA had higher sensitivity whilst the specificity was similar or higher. We conclude that the TK ELISA presents a strong predictor of NPC and, in its refined form, has improved pickup rates. In addition, results from patients with chronic nasopharyngitis (CNP) suggest that individuals with both symptoms of CNP and an elevated TK ELISA value may be at increased risk for the development of head-and neck cancer. PMID- 11267983 TI - In nasopharyngeal carcinoma-bearing patients, tumors and lymphocytes are infected by different Epstein-Barr virus strains. AB - Despite the fact that most adult humans worldwide are latently infected by the Epstein-Barr virus (EBV), only a very small percentage of them will develop an EBV-associated malignancy. We do not know whether this situation reflects the existence of more sensitive individuals or of particularly tumorigenic EBV strains. We postulated that if highly tumorigenic EBV strains did exist, they would be preferentially found in consistently EBV-associated tumors, such as nasopharyngeal carcinoma (NPC), and differ significantly from the strains present in other, non-pathological sites of the same patients. To test this hypothesis, we compared the BNLF1 gene of the EBV strains present in tumors and in "reservoir lymphocytes" of 6 NPC-bearing patients from Tunisia. Our results show that all of these patients were infected by more than 1 (and up to 7) EBV strains. Moreover, lymphocytes and tumor cells from the same individual were systematically infected by different viral strains. The origin and biological significance of these multistrain infections are discussed. PMID- 11267984 TI - Prostaglandins, but not tumor-derived IL-10, shut down concomitant tumor-specific CTL responses during murine plasmacytoma progression. AB - IL-10 is assumed to be a major immunosuppressive factor produced by most B-cell tumors. The immunosuppressive role of tumor-derived IL-10 was analyzed using the MHC class II-negative BALB/c plasmacytoma ADJ-PC-5 as a model tumor. Immune monitoring of tumor-bearing mice was based on the measurement of tumor burden, tumor-specific CTL cytotoxicity and intracellular cytokine staining using FACS. ADJ-PC-5 tumor progression in syngeneic recipients is associated with strong, concomitant, tumor-specific CTL responses during early stages of tumor progression which are sufficient to cause rejection of small s.c. autologous test tumors. These initial CTL responses gradually decline during later tumor stages. Blocking of IL-10 in vivo did not abolish CTL suppression or retard tumor growth. More strikingly, application of anti-IL-10 antibodies during early tumor stages abrogated CTL induction and markedly accelerated tumor growth. In contrast to anti-IL-10 treatment, application of cyclo-oxygenase inhibitors to ADJ-PC-5 tumor bearing mice led to enhanced tumor-specific CTL responses throughout all stages of tumor progression, paralleled by retarded tumor growth and a significantly delayed onset of suppression. Both findings contradict a dominant immunosuppressive role of IL-10 during B-cell tumor progression. Tumor-derived IL 10 must therefore be considered an immunostimulating factor, which accounts for the high immunogenicity of B-cell tumors, whereas prostaglandins, which are not produced by the tumor cells themselves, are the dominant immunosuppressors in this system. PMID- 11267985 TI - Cytokine secretion associated with the clearance of apoptotic bodies in renal cell carcinoma patients. AB - The factors determining the outcome of immunotherapy in metastatic renal cell carcinoma (RCC) patients remain elusive. Macrophages from normal donors that phagocytose apoptotic cells secrete the immunosuppressive cytokine IL-10 in vitro. Conversely, IL-10 genetic deletion enhances the immunogenicity of apoptotic tumor cells in vivo. Elevated pre-treatment levels of IL-10 are associated with an unfavorable outcome of RCC. We examined whether the ability to release IL-10 by macrophages from RCC patients that phagocytosed apoptotic cells correlated with the outcome of immunotherapy. To this aim, we derived macrophages from 30 patients with metastatic RCC and from 21 healthy subjects (11 sex- and age-matched healthy controls and 10 younger donors). Patients either had a clinical response after immunotherapy, with a median survival after treatment of more than 18 months (n = 16), or were beginning immunotherapy after diagnosis of metastatic disease (n = 14). Macrophages from responding patients challenged with apoptotic cells released significantly less IL-10 than controls (p = 0.0075) and recently diagnosed patients (p = 0.0198), as ascertained by a 2-sided Student's t test. This was not because macrophages from responding patients lost the ability to secrete IL-10, because antibody opsonization of apoptotic cells rescued IL-10 secretion. In contrast, macrophages from all groups of donors released similar amounts of TNF-alpha. The failure in IL-10 secretion by engulfing macrophages of responding subjects may exalt the immunogenicity of dying tumor cells, contributing to the success of immunotherapy. PMID- 11267986 TI - Inhibition of human tumor cell growth in vivo by an orally bioavailable inhibitor of human farnesyltransferase, BIM-46228. AB - Oncogenic mutations of the ras gene leading to constitutive activation of downstream effectors have been detected in a wide spectrum of human cancers (pancreas, thyroid, colon, non-small-cell lung cancer). Membrane anchorage of Ras, required for functional activity in signal transduction, is facilitated by post-translational modifications resulting in covalent attachment of a farnesyl group to the cysteine in the C-terminal CAAX motif. This attachment is mediated by farnesyltransferase (FTase). Here, we report a novel FTase inhibitor, BIM 46228, which showed (i) specific inhibition of purified human FTase enzyme, (ii) inhibition of proliferation in vitro in a large spectrum of human tumor cell lines, (iii) inhibition of growth of human tumor xenografts in athymic nude mice treated by per os administration and (iv) the benefits of in vitro combination of its activity with chemotherapy or radiotherapy. PMID- 11267987 TI - Expression of herpes simplex virus-thymidine kinase gene controlled by a promoter region of the midkine gene confers selective cytotoxicity to ganciclovir in human carcinoma cells. AB - A selective expression of suicide gene(s) in tumor cells should produce a preferential cytotoxic effect on tumors. Promoter region(s) of a gene that is expressed in tumors but not in normal tissues can be useful for tumor-specific transcription of a suicide gene. Midkine (MK), a growth/differentiation factor, is expressed predominantly in various types of human tumors, whereas its expression in adult normal tissues is highly restricted. In our study, we showed that a 2.3-kb fragment of genomic DNA in the 5' upstream region of the MK gene could activate transcription of a fused reporter gene in MK-positive cells but not in MK-negative cells. Efficiency of the cis-acting sequence to permit expression of an exogenous gene in tumor cells was comparable with that of the SV40 promoter. Regulated expression of the herpes simplex virus-thymidine kinase (HSV-TK) gene under the control of the MK promoter conferred increased sensitivity to ganciclovir (GCV) on MK-positive tumor cells. Administration of GCV into nude mice that were implanted with MK-positive tumor cells that expressed the HSV-TK gene under the control of the MK promoter could suppress the subsequent tumor growth. Expression of therapeutic genes restricted to tumors can be achieved by the use of the putative cis-acting MK promoter. PMID- 11267988 TI - Residential magnetic fields as a risk factor for childhood acute leukaemia: results from a German population-based case-control study. AB - Our objective was to investigate whether exposure to residential power-frequency (50 Hz) magnetic fields above 0.2 microT increases a child's risk of leukaemia and to confirm or reject a finding from a previous German study on this topic, which reported increased leukaemia risk with exposure to stronger magnetic fields during the night. A population-based case-control study was used, covering the whole of the former West Germany. Residential magnetic fields were measured over 24 hr for 514 children with acute leukaemia identified by the German Childhood Cancer Registry and 1,301 control children taken from population registration files. Magnetic fields above 0.2 microT were relatively rare in Germany (only 1.5% of the study population). Childhood leukaemia and 24 hr median magnetic fields were only weakly related (OR = 1.55, 95% CI 0.65-3.67). A significant association was seen between childhood leukaemia and magnetic field exposure during the night (OR = 3.21, 95% CI 1.33-7.80). A dose-response-relationship was observed after combining the data of all German studies on magnetic fields and childhood leukaemia. The evidence for an association between childhood leukaemia and magnetic field exposure in our study comes from a measure of exposure during the night. Despite the large size of our study, the results are based on small numbers of exposed children. If the observed association stands, the effect on a population level in Germany would be small. PMID- 11267990 TI - Human herpes virus-8 infection among pregnant women and their children: results from the Sardinia-IDDM Study 2. PMID- 11267989 TI - Free insulin-like growth factor-I and breast cancer risk. AB - Insulin-like growth factor-I (IGF-I) has mitogenic and anti-apoptotic effects on breast cancer cells. Epidemiologic studies have shown that high plasma levels of IGF-I and low levels of IGF binding protein (BP)-3 are associated with increased risk of breast cancer in premenopausal women. The actions of IGF-I are mediated through the IGF-I receptor (IGF-IR) and are regulated by IGFBPs. In circulation, most of the IGF-I binds to IGFBP-3, and binding of IGF-I to IGFBP-3 inhibits the actions of IGF-I. Since free IGF-I, which does not bind to IGFBPs, can readily cross the endothelial barrier to interact with IGF-IR, circulating free IGF-I is thought to be more relevant to the biologic activity of IGF-I. To examine the association of free IGF-I with breast cancer, we compared free IGF-I levels between 40 newly diagnosed breast cancer patients and 40 age- and race-matched healthy controls. Plasma levels of free IGF-I, total IGF-I and IGF-II, as well as total, intact and fragment IGFBP-3, were measured using commercial immunoassay kits. The association between IGF-I and breast cancer was examined using the conditional logistic regression analysis. Analysis of correlation (Spearman) showed that free IGF-I was correlated with total IGF-I and IGFBP-3 but not with IGF-II. The odds ratios for breast cancer patients having high plasma IGF-I (> or = median) after adjusting for menopausal status and IGFBP-3 were 2.00 (p < o r = 0.376) for total IGF-I and 6.31 (p < or = 0.047) for free IGF-I. A high ratio of IGF-I to IGFBP-3 was also associated with breast cancer (p < 0.05). No association was found for IGF-II, nor for total, intact and fragment IGFBP-3. The findings of this study suggest that measuring free IGF-I in circulation is more useful than measuring total IGF-I with respect to evaluation of an association between IGF-I and breast cancer risk. PMID- 11267991 TI - A BRCA2 germ-line mutation in familial pancreatic carcinoma. PMID- 11267992 TI - Risk of lung cancer from tobacco smoking among young women from Europe. PMID- 11267993 TI - Role of MED1 (MBD4) Gene in DNA repair and human cancer. AB - The human protein MED1, also known as MBD4, was isolated in a yeast two-hybrid screening as an interactor of the mismatch repair protein MLH1. MED1 contains an N-terminal 5-methylcytosine binding domain (MBD), which allows binding to methylated DNA, and a C-terminal catalytic domain with homology to bacterial DNA damage-specific glycosylases/lyases. This suggests that DNA methylation may play a role in human DNA repair. MED1 acts as a mismatch-specific DNA N-glycosylase active on thymine, uracil, 5-fluorouracil and, weakly, 3,N(4)-ethenocytosine paired with guanine. The glycosylase activity of MED1 prefers substrates in which the G:T mismatch is present in the context of methylated or unmethylated CpG sites. Since G:T mismatches can originate via spontaneous deamination of 5 methylcytosine to thymine, MED1 appears to act as a caretaker of genomic fidelity at CpG sites. Mutagenesis caused by these deamination events is a frequent mechanism of genetic instability in cancer; thus, based on the biochemical activity of its gene product, MED1 is a candidate tumor suppressor gene. Indeed, frameshift mutations of the MED1 gene have been reported in human colorectal, gastric, endometrial, and pancreatic cancer. In the future, efforts should be directed toward investigations of the functional role of the MED1 gene in the pathogenesis, prevention, and treatment of human cancer. PMID- 11267994 TI - Mismatch repair in correction of replication errors and processing of DNA damage. AB - The primary role of mismatch repair (MMR) is to maintain genomic stability by removing replication errors from DNA. This repair pathway was originally implicated in human cancer through an association between microsatellite instability in colorectal tumors in hereditary nonpolyposis colon cancer (HNPCC) kindreds. Microsatellites are short repetitive sequences which are often copied incorrectly by DNA polymerases because the template and daughter strands in these regions are particularly prone to misalignment. These replication-dependent events create loops of extrahelical bases which would produce frameshift mutations unless reversed by MMR. One consequence of MMR loss is a widespread expansion and contraction of these repeated sequences that affects the whole genome. Defective MMR is therefore associated with a mutator phenotype. Since the same pathway is also responsible for repairing base:base mismatches, defective cells also experience large increases in the frequency of spontaneous transition and transversion mutations. Three different approaches have been used to investigate the function of individual components of the MMR pathway. The first is based on the biochemical characterization of the purified protein complexes using synthetic DNA substrates containing loops or single mismatches. In the second, the biological consequences of MMR loss are inferred from the phenotype of cell lines established from repair-deficient human tumors, from tolerant cells or from mice defective in single MMR genes. In particular, molecular analysis of the mutations in endogenous or reporter genes helped to identify the DNA substrates for MMR. Finally, mice bearing single inactive MMR genes have helped to define the involvement of MMR in cancer prevention. PMID- 11267995 TI - The gene PC3(TIS21/BTG2), prototype member of the PC3/BTG/TOB family: regulator in control of cell growth, differentiation, and DNA repair? AB - PC3(TIS21/BTG2) is the founding member of a family of genes endowed with antiproliferative properties, namely BTG1, ANA/BTG3, PC3B, TOB, and TOB2. PC3 was originally isolated as a gene induced by nerve growth factor during neuronal differentiation of rat PC12 cells, or by TPA in NIH3T3 cells (named TIS21), and is a marker for neuronal birth in vivo. This and other findings suggested its implication in the process of neurogenesis as mediator of the growth arrest before differentiation. Remarkably, its human homolog, named BTG2, was shown to be p53-inducible, in conditions of genotoxic damage. PC3(TIS21/BTG2) impairs G(1) S progression, either by a Rb-dependent pathway through inhibition of cyclin D1 transcription, or in a Rb-independent fashion by cyclin E downregulation. PC3(TIS21/BTG2) might also control the G(2) checkpoint. Furthermore, PC3(TIS21/BTG2) interacts with carbon catabolite repressor protein-associated factor 1 (CAF-1), a molecule that associates to the yeast transcriptional complex CCR4 and might influence cell cycle, with the transcription factor Hoxb9, and with the protein-arginine methyltransferase 1, that might control transcription through histone methylation. Current evidence suggests a physiological role of PC3(TIS21/BTG2) in the control of cell cycle arrest following DNA damage and other types of cellular stress, or before differentiation of the neuron and other cell types. The molecular function of PC3(TIS21/BTG2) is still unknown, but its ability to modulate cyclin D1 transcription, or to synergize with the transcription factor Hoxb9, suggests that it behaves as a transcriptional co regulator. PMID- 11267996 TI - Mechanisms of P2 receptor-evoked DNA synthesis in thyroid FRTL-5 cells. AB - The expression of the P2 receptors and their functional responses were studied in rat thyroid FRTL-5 cells. RT-PCR analysis revealed transcripts for the G protein coupled P2Y(2), P2Y(4) and P2Y(6) receptors, and for the transmitter-gated ion channel P2X(3), P2X(4) and P2X(5) subunits. In Fura-2-loaded cells, UTP, ATP, ATPgammaS or UDP increased [Ca(2+)](i), and behaved as potent full agonists, while 2-Methylthio-ATP (2-MeSATP), alpha,beta-methylene-ATP (alpha,beta-meATP) and pure ADP were weak agonists. The agonist-mediated [Ca(2+) ](i) increases were diminished in Ca(2+) -free buffer, and by pertussis toxin (PTX) or suramin treatments. ATP, UTP, UDP and ATPgammaS increased (3)H-thymidine incorporation into DNA and expression of the protooncogenes c-Fos and c-Jun, while 2-MeSATP was ineffective, and alpha,beta-meATP gave a response only at 100-microM dose. The ATP-stimulated expression of c-Fos and c-Jun was dependent on Ca(2+), and protein kinase C, but not on calmodulin or Ca(2+)/calmodulin-dependent protein kinase II. Extracellular signal-regulated kinases (ERK1 and ERK2) are also involved as the MEK inhibitor, PD98059, reduced both ATP-evoked (3)H-thymidine incorporation and c-Fos and c-Jun expression. These results indicate that multiple P2Y receptor subtypes and at least the P2X(5) subtype are functionally expressed in FRTL-5 cells, and that nucleotides acting via P2 receptors are involved in the regulation of DNA-synthesis. PMID- 11267997 TI - Tpl-2 induces apoptosis by promoting the assembly of protein complexes that contain caspase-9, the adapter protein Tvl-1, and procaspase-3. AB - The Tpl-2 proto-oncoprotein promotes cellular proliferation when overexpressed in a variety of tumor cell lines. Here, we present evidence that when overexpressed in immortalized non-transformed cells, Tpl-2 induces apoptosis by promoting the activation of caspase-3 via a caspase-9-dependent mechanism, and that apoptosis is enhanced when Tpl-2 is co-expressed with the newly identified ankyrin repeat protein Tvl-1. The activation of caspase-3 by caspase-9 is known to depend on the assembly of a multimolecular complex that includes Apaf-1 and caspase-9. Data presented here show that co-expression of Tpl-2 with Tvl-1 promotes the assembly of a complex that involves several proteins that bind Apaf-1 including Tvl-1, itself, Tpl-2 and phosphorylated procaspase-9. More important, procaspase-3, which under normal growth conditions is not associated with the complex, binds Tvl-1 conditionally in response to Tpl-2-generated apoptotic signals. The conditional association of procaspase-3 with Tvl-1 promotes the in vivo proteolytic maturation of procaspase-3 by caspase-9, a process casually linked to apoptosis. PMID- 11267998 TI - Requirement of protein kinase Calpha, extracellular matrix remodeling, and cell matrix interaction for transforming growth factorbeta-regulated expression of E cadherin and catenins. AB - A hallmark of transforming growth factorbeta (TGFbeta) action is the induction of the synthesis and secretion of extracellular-matrix adhesion molecules and induction of the cell-surface expression of integrin receptors for these molecules (termed extracellular-matrix remodeling). The signal pathways leading to extracellular-matrix remodeling and the significance of extracellular-matrix remodeling in TGFbeta function is not well-understood. In the epithelium-derived human colon cancer cell line Moser, TGFbeta induces extracellular-matrix remodeling in a protein kinase Calpha-dependent manner. In this study we showed that TGFbeta was a potent inducer of the homotypic cell-cell adhesion molecule E cadherin and its undercoat-associated proteins, the catenins and dramatically increased the amount of E-cadherin/gamma-catenin complex formation. We found that the induction of E-cadherin and alpha- and beta-catenin by TGFbeta was also dependent on protein kinase Calpha, whereas the induction of gamma-catenin was independent of protein kinase Calpha but dependent on other protein kinase C isoforms. We also found that protein kinase Calpha-dependent induction of extracellular-matrix remodeling and subsequent cell-matrix interaction requiring both fibronectin and laminin were a prerequisite for the induction of E-cadherin (and alpha- and beta-catenin but not gamma-catenin) by TGFbeta. We therefore concluded that two signal pathways exist in TGFbeta-regulated expression of E cadherin and the catenins. We also concluded that a functional significance of TGFbeta-induced extracellular matrix remodeling is the activation of signal transduction mechanisms through increased interaction between extracellular matrix fibronectin and laminin and their cell-surface integrin receptors, which lead to the induction of E-cadherin (and alpha- and beta-catenin). PMID- 11267999 TI - P2Y(2) nucleotide receptor signaling in human monocytic cells: activation, desensitization and coupling to mitogen-activated protein kinases. AB - Activation of P2Y(2) receptors by extracellular nucleotides has been shown to induce phenotypic differentiation of human promonocytic U937 cells that is associated with the inflammatory response. The P2Y(2) receptor agonist, UTP, induced the phosphorylation of the MAP kinases MEK1/2 and ERK1/2 in a sequential manner, since ERK1/2 phosphorylation was abolished by the MEK1/2 inhibitor PD 098059. Other results indicated that P2Y(2) receptors can couple to MAP kinases via phosphatidylinositol 3-kinase (PI3K) and c-src. Accordingly, ERK1/2 phosphorylation induced by UTP was inhibited by the PI3K inhibitors, wortmannin and LY294002, and the c-src inhibitors, radicicol and PP2, but not by inhibitors of protein kinase C (PKC). The phosphorylation of ERK1/2 was independent of the ability of P2Y(2) receptors to increase the concentration of intracellular free calcium, since chelation of intracellular calcium by BAPTA did not diminish the phosphorylation of ERK1/2 induced by UTP. A 5-minute treatment with UTP reduced U937 cell responsiveness to a subsequent UTP challenge. UTP-induced desensitization was characterized by an increase in the EC(50) for receptor activation (from 0.44 to 9.3 microM) and a dramatic ( approximately 75%) decrease in the maximal calcium mobilization induced by a supramaximal dose of UTP. Phorbol ester treatment also caused P2Y(2) receptor desensitization (EC(50) = 12.3 microM UTP and maximal calcium mobilization reduced by approximately 33%). The protein kinase C inhibitor GF 109203X failed to significantly inhibit the UTP induced desensitization of the P2Y(2) receptor, whereas the protein phosphatase inhibitor okadaic acid blocked receptor resensitization. Recovery of receptor activity after UTP-induced desensitization was evident in cells treated with agonist for 5 or 30 min. However, P2Y(2) receptor activity remained partially desensitized 30 min after pretreatment of cells with UTP for 1 h or longer. This sustained desensitized state correlated with a decrease in P2Y(2) receptor mRNA levels. Desensitization of ERK1/2 phosphorylation was induced by a 5-minute pretreatment with UTP, and cell responsiveness did not return even after a 30 minute incubation of cells in the absence of an agonist. Results suggest that desensitization of the P2Y(2) receptor may involve covalent modifications (i.e., receptor phosphorylation) that functionally uncouple the receptor from the calcium signaling pathway, and that transcriptional regulation may play a role in long-term desensitization. Our results indicate that calcium mobilization and ERK1/2 phosphorylation induced by P2Y(2) receptor activation are independent events in U937 monocytes. PMID- 11268000 TI - Ebp1, an ErbB-3 binding protein, interacts with Rb and affects Rb transcriptional regulation. AB - Ebp1, an ErbB-3 binding protein, inhibits the proliferation and induces the differentiation of human breast cancer cells. The mechanisms of these effects are unknown. Rb, the product of the retinoblastoma gene, is an important modulator of cell cycle progression and cellular differentiation. We report that Rb is a binding target for Ebp1. Ebp1 was localized to both the nucleus and the cytoplasm of logarithmically growing AU565 breast cancer cells and HeLa cells as determined by confocal immunofluorescent microscopy. Ebp1 was present in Rb immunoprecipitates derived from AU565 breast cancer cells. GST-Rb also bound endogenous Ebp1. Using GST-Ebp1 constructs, we determined that the 72 C-terminal amino acids of Ebp1 were sufficient to bind Rb. Dephosphorylation of Ebp1 enhanced the interaction of Ebp1 with Rb. The overexpression of Ebp1 in MCF-7 and AU565 (Rb(+)) cells inhibited the activity of the E2F1 regulated cyclin-E promoter. Ebp1 bound E2F1 indirectly via Rb in lysates of MCF-7 cells. The interaction of Ebp1 with Rb may prove to be an important mechanism of Ebp1 induced changes in cell proliferation and differentiation. PMID- 11268001 TI - Endothelin-1 expression in long-term cultures of fetal rat calvarial osteoblasts: regulation by BMP-7. AB - Endothelin-1 (ET-1) is a vasoactive peptide that modulates bone metabolism via regulatory effects on osteoblasts, chondrocytes, and osteoclasts. While ET-1 may circulate in the blood stream, tissue-specific expression of this peptide is more physiologically relevant. In the present study we measured ET-1 synthesis in sections of fetal rat calvaria (FRC) and in cultured FRC osteoblasts. Regulation of ET-1 synthesis in FRC osteoblasts by bone morphogenetic protein-7 (BMP-7) and transforming growth factor-beta1 (TGF-beta1) also was examined. Immunohistochemical analysis revealed ET-1 staining in calvarial osteoblasts, endothelial cells, and osteocytes. ET-1 mRNA expression was detected in cultured FRC cells and ET-1 peptide was present in conditioned media. During long-term culture of FRC cells (26 days) ET-1 peptide production rose sharply and peaked during the time of cellular proliferation (Days 0-3) then returned to baseline levels by Day 18, when mineralized nodules were forming. Treatment of FRC cells with BMP-7 enhanced ET-1 levels by three-fold on Day 3 and enhanced nodule formation by 15-fold on Day 26. To determine whether ET-1 was involved in an autocrine manner in BMP-7-induced nodule formation, cells were cultured in the presence of BMP-7 and BQ-123, an ET(A) receptor antagonist. BQ-123 had no effect on nodule formation in control or BMP-7-treated cells, indicating that osteoblast derived ET-1 regulates other cell types in vivo during the bone formation process. PMID- 11268002 TI - Loss of functional caveolae during senescence of human fibroblasts. AB - Primary human fibroblasts have a finite replicative lifespan in culture that culminates in a unique state of growth arrest, termed senescence that is accompanied by distinct morphological and biochemical alterations. Senescent cell responses to extracellular stimuli are believed to be altered at a point after receptors are bound by ligand, leading to improper integration of the signals which initiate DNA replication. In this study we demonstrate that one of the key organizing membrane microdomains for receptor signaling, caveolae, are absent in senescent cells. A comparison of young and senescent cells indicated that senescent cells contained a higher total amount of caveolins 1 and 2 but had significantly less of both proteins in the caveolar fraction. Additionally, caveolar fractions from senescent cells completely lacked the tyrosine-kinase activity associated with functional caveolae. Furthermore, old cells had little caveolar protein exposed to the outer plasma membrane as estimated by using an in vivo biotinylation assay and no detectable caveolin 1 on the cell surface when processed for immunofluoresence and confocal microscopy. Together, these data suggest that a fundamental loss of signal integration at the plasma membrane of senescent cells is due to the loss of signaling competent caveolae. PMID- 11268003 TI - Delaying S-phase progression rescues cells from heat-induced S-phase hypertoxicity. AB - The mechanism by which a cell protects itself from the lethal effects of heat shock and other stress-inducing agents is the subject of much research. We have investigated the relationship between heat-induced damage to DNA replication machinery and the lethal effects of heat shock, in S-phase cells, which are more sensitive to heat shock than either G1 or G2. We found that maintaining cells in aphidicolin, which prevents the passage of cells through S-phase, can rescue S phase HeLa cells from the lethal effects of heat shock. When S-phase, HeLa cells were held for 5-6 h in 3 microM aphidicolin the measured clonogenic survival was similar to that for exponentially growing cells. It is known, that heat shock induces denaturation or unfolding of proteins, rendering them less soluble and more likely to co-isolate with the nuclear matrix. Here, we show that enhanced binding of proteins involved in DNA replication (PCNA, RPA, and cyclin A), with the nuclear matrix, correlates with lethality of S-phase cells following heat shock under four different experimental conditions. Specifically, the amounts of RPA, PCNA, and cyclin A associated with the nuclear matrix when cells resumed progression through S-phase correlated with cell killing. Heat-induced enhanced binding of nuclear proteins involved with other aspects of DNA metabolism, (Mrell, PDI), do not show this correlation. These results support the hypothesis that heat-induced changes in the binding of proteins associated with DNA replication factories are the potentially lethal lesions, which become fixed to lethal lesions by S-phase progression but are repairable if S-phase progression is delayed. PMID- 11268004 TI - Functional receptor-channel coupling compared in contractile and proliferative human vascular smooth muscle. AB - We have previously identified a human vascular smooth muscle clone that can reversibly convert between proliferative and contractile phenotypes. Here we compared receptor-channel coupling in these cells using fura-2 to monitor [Ca(2+)](i) and patch-clamp to record currents. Histamine elevated [Ca(2+)](i) in all cells and caused contraction of cells exhibiting the contractile phenotype. The rise of [Ca(2+)](i) persisted in Ca(2+)-free solution and was abolished by thapsigargin, indicating involvement of stores. Whole cell electrophysiological recording revealed that histamine evoked transient outward K(+) current, indicating functional receptor-channel coupling. The time-course and amplitude of the histamine-activated current were similar in cells of the proliferative and contractile phenotypes. Moreover, a large conductance K(+) channel was recorded in cell-attached patches and was activated by histamine as well as the Ca(2+) ionophore A-23187, identifying it as the large conductance Ca(2+)-dependent K(+) channel. This K(+) channel showed similar characteristics and activation in both proliferative and contractile phenotypes, indicating that expression was independent of phenotype. In contrast, histamine also elicited an inward Cl(-) current in some contractile cells, suggesting differential regulation of this current depending on phenotype. These studies demonstrate the usefulness of this human vascular cell clone for studying functional plasticity of smooth muscle, while avoiding complications arising from extended times in culture. PMID- 11268005 TI - Febrile and acute hyperthermia enhance TNF-induced necrosis of murine L929 fibrosarcoma cells via caspase-regulated production of reactive oxygen intermediates. AB - Previous studies have demonstrated the essential role of TNF-induced reactive oxygen intermediates (ROI) in the necrosis of L929 cells. We investigated the molecular basis for the interaction of hyperthermia and TNF in these cells. Hyperthermia, both febrile (40.0-40.5 degrees C) and acute (41.5-41.8 degrees C), strongly potentiated TNF killing, and sensistization was significantly quenched by the antioxidant, BHA. The broad-spectrum caspase inhibitor, Z-VAD, has been shown to markedly increase the TNF sensitivity of L929 cells at 37 degrees C; we observed that hyperthermia would also enhance the sensitivity of L929 cells to TNF + Z- VAD and that BHA could significantly quench the response, as well. The basis for hyperthermic potentiation was unlikely thermally-increased sensitivity to ROI, as treatment with hydrogen peroxide for 24 h killed L929 cells essentially equivalently, whether incubated continuously at 37 degrees C or at 40.0-40.5 degrees C, or for 2 h at 41.5-41.8 degrees C. However, febrile and acute hyperthermia markedly increased TNF-induced production of ROI, with or without Z-VAD. Hyperthermia dramatically accelerated the onset of this production, as well as the onset of necrotic death, as determined by oxidation of dihydro-rhodamine and propidium iodide staining, respectively, both of which were significantly quenchable with BHA. We conclude that hyperthermia potentiates TNF mediated killing in this cell model primarily by increasing the afferent, and not the efferent, phase of TNF-induced necrosis. PMID- 11268006 TI - Colocalization of GABA and glycine in the ventral nucleus of the lateral lemniscus in rat: an in situ hybridization and semiquantitative immunocytochemical study. AB - We have studied by in situ hybridization for GAD65 mRNA in thick sections and by semiquantitative postembedding immunocytochemistry in consecutive semithin sections, the expression of gamma-aminobutyric acid (GABA) and glycine in cell bodies and axosomatic puncta of the rat ventral nucleus of the lateral lemniscus (VNLL), a prominent monaural brainstem auditory structure. The in situ hybridization and the densitometric analysis of the immunostaining suggest that the rat VNLL contains two main populations of neurons. Approximately one-third of neurons are unstained with either technique and are presumably excitatory; their cell bodies are enveloped by a large number of glycine-immunoreactive puncta. Most if not all of the remaining two-thirds colocalize GABA and glycine and are assumed to be inhibitory. These two populations show a complementary distribution within the VNLL, with inhibitory neurons located mainly ventrally and excitatory neurons dorsally. In scatterplots of gray values measured from cell bodies, the double-labeled cells appear to form a single cluster in terms of their staining intensities for the two transmitter candidates. However, this cluster may have to be further subdivided because cells with extreme GABA/glycine ratios differ from those with average ratios with respect to location or size. The VNLL seems unique among auditory structures by its large number of neurons that colocalize GABA and glycine. Although the functional significance of this colocalization remains unknown, our results suggest that the VNLL exerts convergent excitatory and inhibitory influences over the inferior colliculus, which may underlie the timing processing in the auditory midbrain. PMID- 11268008 TI - Projections from the amygdalo-piriform transition area to the amygdaloid complex: a PHA-l study in rat. AB - The amygdalo-piriform transition area is a poorly defined region in the temporal lobe that is heavily connected with the olfactory system. As part of an ongoing project aimed at understanding the neuronal pathways that provide sensory information to the amygdala, we investigated the cytoarchitectonic and chemoarchitectonic features of the amygdalo-piriform transition area and its connections to the amygdaloid complex in 13 rats by using the anterograde tracer, Phaseolus vulgaris-leucoagglutinin. Our analysis indicates that the amygdalo piriform transition area has medial (rostral and caudal portions) and lateral parts. The rostromedial part projects heavily to the intermediate and lateral divisions of the central nucleus, whereas the caudomedial part projects mainly to the medial division. The lateral part of the amygdalo-piriform transition area projects heavily to the capsular and lateral divisions of the central nucleus. Electron microscopic analysis revealed that the projection to the lateral division of the central nucleus forms asymmetric contacts with the spines and shafts of postsynaptic neurons and, therefore, is assumed to be excitatory. The amygdalo-piriform transition area also projects moderately to other amygdaloid nuclei, including the parvicellular division of the basal nucleus, the anterior cortical nucleus, and the nucleus of the lateral olfactory tract. The lateral and medial parts of the amygdalo-piriform transition area also project to the distal temporal CA1 and distal temporal subiculum, respectively. Unlike the adjacent entorhinal cortex, the amygdalo-piriform transition area does not project to the dentate gyrus. These data suggest that the amygdalo-piriform transition area is a region that influences both emotional and memory processing in parallel by means of pathways to the amygdala and the hippocampus, respectively. PMID- 11268007 TI - Odorants as cell-type specific activators of a heat shock response in the rat olfactory mucosa. AB - Heat shock, or stress, proteins (HSPs) are induced in response to conditions that cause protein denaturation. Activation of cellular stress responses as a protective and survival mechanism is often associated with chemical exposure. One interface between the body and the external environment and chemical or biological agents therein is the olfactory epithelium (OE). To determine whether environmental odorants affect OE HSP expression, rats were exposed to a variety of odorants added to the cage bedding. Odorant exposure led to transient, selective induction of HSP70, HSC70, HSP25, and ubiquitin immunoreactivities (IRs) in supporting cells and subepithelial Bowman's gland acinar cells, two OE non-neuronal cell populations involved with inhalant biotransformation, detoxification, and maintenance of overall OE integrity. Responses exhibited odor specificity and dose dependency. HSP70 and HSC70 IRs occurred throughout the apical region of supporting cells; ubiquitin IR was confined to a supranuclear cone-shaped region. Electron microscopic examination confirmed these observations and, additionally, revealed odor-induced formation of dense vesicular arrays in the cone-like regions. HSP25 IR occurred throughout the entire supporting cell cytoplasm. In contrast to classical stress responses, in which the entire array of stress proteins is induced, no increases in HSP40 and HSP90 IRs were observed. Extended exposure to higher odorant doses caused prolonged activation of the same HSP subset in the non-neuronal cells and severe morphological damage in both supporting cells and olfactory receptor neurons (ORNs), suggesting that non neuronal cytoprotective stress response mechanisms had been overwhelmed and could no longer adequately maintain OE integrity. Significantly, ORNs showed no stress responses in any of our studies. These findings suggest a novel role for these HSPs in olfaction and, in turn, possible involvement in other normal neurophysiological processes. PMID- 11268009 TI - Light and electron microscopic study of the distribution of substance P immunoreactive fibers and neurokinin-1 receptors in the skin of the rat lower lip. AB - Cutaneous antidromic vasodilatation and plasma extravasation, two phenomena that occur in neurogenic inflammation, are partially blocked by substance P (SP) receptor antagonists and are known to be mediated in part by mast cell-released substances, such as histamine, serotonin, and nitric oxide. In an attempt to provide a morphological substrate for the above phenomena, we applied light and electron microscopic immunocytochemistry to investigate the pattern of SP innervation of blood vessels and its relationship to mast cells in the skin of the rat lower lip. Furthermore, we examined the distribution of SP (neurokinin-1) receptors and their relationship to SP-immunoreactive (IR) fibers. Our results confirmed that SP-IR fibers are found in cutaneous nerves and that terminal branches are observed around blood vessels and penetrating the epidermis. SP-IR fibers also innervated hair follicles and sebaceous glands. At the ultrastructural level, SP-IR varicosities were observed adjacent to arterioles, capillaries, venules, and mast cells. The varicosities possessed both dense core vesicles and agranular synaptic vesicles. We quantified the distance between SP IR varicosities and blood vessel endothelial cells. SP-IR terminals were located within 0.23-5.99 microm from the endothelial cell layer in 82.7% of arterioles, in 90.2% of capillaries, and in 86.9% of venules. Although there was a trend for SP-IR fibers to be located closer to the endothelium of venules, this difference was not significant. Neurokinin-1 receptor (NK-1r) immunoreactivity was most abundant in the upper dermis and was associated with the wall of blood vessels. NK-1r were located in equal amounts on the walls of arterioles, capillaries, and venules that were innervated by SP-IR fibers. The present results favor the concept of a participation of SP in cutaneous neurogenic vasodilatation and plasma extravasation both by an action on blood vessels after binding to the NK 1r and by causing the release of substances from mast cells after diffusion through the connective tissue. PMID- 11268010 TI - Projections of the oval nucleus of the hyperstriatum ventrale in the budgerigar: relationships with the auditory system. AB - The afferent and efferent projections of a vocal control nucleus, the oval nucleus of the hyperstriatum ventrale (HVo), were mapped out in a parrot, the budgerigar (Melopsittacus undulatus) to determine the relationships of this nucleus to the auditory system. In budgerigars, HVo is connected to both the anterior forebrain pathway as well as to nuclei forming the descending projection system to the brainstem (Durand et al. [1997] J. Comp. Neurol. 377:179-206). Previous studies (Brauth et al. [1997] Proc. N. Y. Acad. Sci. 807:368-385; Durand and Brauth [1998] Neurosci Abstr 24:78.9) indicate that HVo lesions disrupt vocal performance and that HVo neurons show long latency electrophysiologic auditory responses. HVo has also been shown to receive input from neurons in the immediately adjacent HV (Durand et al. [1997] J. Comp. Neurol. 377:179-206). Thus, the focus of the present study was to elucidate relationships between HVo, its immediately adjacent surround and telencephalic auditory nuclei. The results show that, although the lateral and medial portions of HVo are interconnected with one another, inputs to these areas and their surrounds are distinctively different. The most substantial auditory system inputs are derived from the frontal lateral neostriatum (NFl) and supracentral nucleus of the lateral neostriatum (NLs); these project primarily to the lateral HVo and lateral HVo surround. The medial HVo and surround receive only sparse or modest input from auditory nuclei, including the caudomedial neostriatum (NCM), neostriatum intermedium pars lateralis (NIL), Fields L1 and L3, and the neostriatum intermedium pars ventrolateralis (NIVL). Other sources of input to the HVo surround include the hyperstriatum accessorium (HA), the supralaminar area of the frontal neostriatum (NAs), the ventral anterior archistriatum (AAv), the medial archistriatum (Am) and the medial HV. Neurons in the HV immediately medial to HVo project to a shell region around the entire nucleus. Both the ventral paleostriatum (VP) and ventral part of the central nucleus of the lateral neostriatum (NLc) project to HVo but not to the surround. Previously described projections (Durand et al., 1997) from HVo to the NAom, NLc, and the magnicellular nucleus of the lobus parolfactorius (LPOm) were confirmed. PMID- 11268011 TI - The number of morphological synapses between neurons does not predict the strength of their physiological synaptic interactions: a study of dendrites in the nematode Ascaris suum. AB - Previous electrophysiological and anatomical studies of Ascaris suum motor neurons demonstrated a strong correlation between functional interactions and the presence of anatomically defined synapses. However, one example of a physiologically robust synaptic connection was encountered for which no anatomical evidence of direct chemical synapses was found. This involved synaptic transmission from an identified excitatory motor neuron to its inhibitory partner. In this study, pressure injection of horseradish peroxidase or nickel lysine into inhibitory motor neurons revealed numerous spines projecting from the main neuronal process toward the neuromuscular surface that then branched and extended fine, longitudinal processes up to 130 microm in length. Subsequent examination of nickel lysine-injected spines by electron microscopy revealed numerous chemical synapses, including for the first time direct inputs from the excitatory neuron. However, the numbers of synapses from this motor neuron were very small relative to inputs from other identified cells. Thus, direct synapses are unlikely to explain the robust nature of this physiological interaction. PMID- 11268012 TI - Neonatally elevated serotonin levels alter terminal arbors of individual retinal ganglion cells in superior colliculus of hamsters. AB - Previous studies from this laboratory showed that sprouting of serotoninergic (5 HT) axons in the hamster's superior colliculus (SC), induced by a single subcutaneous injection of 5,7-dihydroxytryptamine (5,7-DHT) at birth (postnatal day 0 [P-0]), resulted in an increased terminal distribution of the uncrossed retinocollicular projection that was not associated with any changes in the number or distribution of ipsilaterally projecting retinal ganglion cells. The present study was undertaken to determine what effect this manipulation had on the terminal arbors of such axons. Retinocollicular axons of normal and 5,7-DHT treated animals were anterogradely labeled with small intraretinal injections of the lipophilic dye 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) on P-16. After tissue processing on P-19, single retinocollicular axon arbors were reconstructed by using confocal microscopy. Quantitative analysis indicated that arbors from 5,7-DHT-treated hamsters had significantly greater total fiber lengths, areas, and volumes than those from normal animals. There were no differences between axons from the two groups in number of branch points, distribution of relative branch lengths, and numbers of bouton-like swellings. These results support the hypothesis that increased SC concentrations of 5-HT alter development of the uncrossed retinocollicular pathway such that a greater territory is covered by individual terminal arbors but that the number of synaptic contacts per arbor remains constant. This may explain, at least in part, the abnormally widespread distribution of the aggregate ipsilateral projection. PMID- 11268013 TI - Mono- and oligo-vesicular synapses and their connectivity in a Cnidarian sensory epithelium (Coryne tubulosa). AB - The spherical end-knobs of the tentacles of capitate hydropolyps are an evolutionarily early paradigm of a chemo- and mechanosensory epithelium composed of four types of sensory cells and one type of chemo-mechanosensitive nematocytes (stinging cells), all separated by supporting cells. The epithelium discriminates sites and compositions of stimuli and induces various kinds of behavior. Recent electrophysiological studies demonstrated rapid chemo-synaptic signal transmission of nematocytes and mechanosensory hair cells, graded in amplitude and duration. The present electron microscopic work, applying serial sectioning, analyses the ultrastructural basis of signal transmission and efference control in the tentacular spheres of Coryne tubulosa, a species also used in preceding studies. Neurites of sensory cells and of proximal ganglion cells constitute a nerve plexus at the base of the ectodermal cells. No ganglion cells are located within the spheres. Chemical synapses of the usual configuration connect neurites or are efferent to nematocytes and hair cells. Each of these synapses contains only 3-10 clear and/or dense-core vesicles of 70-150 nm diameter (oligo-vesicular synapses). For the graded afferent signal transmission of nematocytes and hair cells, the only candidates are regularly occurring zones of neurite contacts at the base of these cells. At their presynaptic side, mostly one (more seldom two to four) large vesicles (160-1100-nm-diameter magno-vesicles) are attached to a surface membrane density. In order to reconcile structural and functional data, a transient fusion and partial depletion of stationary vesicles is considered for the release of transmitter in mono-vesicular synapses, similar to recent findings for vertebrate endocrine secretion. The same principle is discussed for the usual oligo-vesicular synapses of Cnidaria. PMID- 11268014 TI - Variations of concentric hair cells in a Cnidarian sensory epithelium (Coryne tubulosa). AB - In capitate hydropolyps, the spherical end-knobs of the short tentacles present an exceptional concentration of sensory functions in one of the evolutionarily oldest nervous systems. The tentacular spheres are the basis of sensation and discrimination of objects and of capturing of prey-objects by the discharge of nematocytes (stinging cells). Recent electrophysiological studies of the spheres revealed combined chemo/mechanosensory functioning of the nematocytes and mechanosensitivity of further types of cells. The present electron microscopical study made use of the small size of the spheres of Coryne tubulosa to characterize all cells of some spheres. Five types of ectodermal cells were found to have sensory structural features and to be separated by or enclosed in supporting cells: 1) nematocytes of the stenotele type; 2) short and 3) long ciliated concentric hair cells, which carry a cilium-stereovilli bundle, similar to the cnidocil apparatus of nematocytes; 4) cells having a recessed cilium microvilli complex equipped with a thick cell-traversing rootlet (rootlet cells); and 5) cells having a recessed short cilium with no microvilli and only a short rootlet and containing, apically as well as basally, aggregations of dense-core vesicles (vesicle-rich cells). Types 1-4 vary the configuration of a concentric cilium-microvilli complex (variations of a concentric hair bundle) and were demonstrated or inferred to be mechanosensitive. Apical exocytotic activity, which is well known for the nematocytes (discharge of their cnidocyst), is indicated by ultrastructure for the nematocyte-resembling concentric hair cells and for the vesicle-rich cells. The tentacular spheres are considered an early paradigm of a sensory epithelium. Its synaptic structures and extensive connectivity are the subject of a subsequent paper. PMID- 11268015 TI - Sleep-disordered breathing in neuromuscular disorders: a condition in search of recognition. PMID- 11268016 TI - The effect of myelinating Schwann cells on axons. AB - Myelinating Schwann cells control the number of neurofilaments and elevate the phosphorylation state of neurofilaments in the axon, eventually leading to the typical large axon caliber. Conversely, absence of myelin leads to lower amounts of neurofilaments, reduced phosphorylation levels, and smaller axon diameters. In addition, myelinating Schwann cells mediate the spacing of Na(+) channel clusters during development of the node of Ranvier. When axons are associated with mutant Schwann cells in inherited neuropathies, their calibers are reduced and their neurofilaments are less phosphorylated and more closely spaced. Also, axonal transport is reduced and axons degenerate at the distal ends of long nerves. Myelin-associated glycoprotein may mediate some aspects of Schwann cell-axon communication, but much remains to be learned about the molecular bases of Schwann cell-axon communication. PMID- 11268017 TI - Reproducibility of motor unit number estimation in individual subjects. AB - Although the reproducibility of motor unit number estimation (MUNE) for groups of subjects has been studied, there is little such data for individuals. Prediction intervals represent a tool to study individual MUNE reproducibility and represent the range of values expected for a future MUNE if the true number of motor units remains unchanged. MUNE was performed using the statistical method on 48 normal individuals. The prediction interval was found to be a function of the intrasubject coefficient of variation. Using a commercial manufacturer's recommended technique and software, prediction intervals were found to be so broad as to be of uncertain value. We found that by averaging two MUNE observations for each determination, and using the method of weighted averages for calculating the size of an average single motor unit potential, the intrasubject coefficient of variation was reduced from 16.48% to 8.77%, and the 90% prediction interval became sufficiently narrow to be clinically useful. False negative rates were also lowered substantially using these techniques. Thus, simple modifications of an existing MUNE program improved the clinical utility of this program for the longitudinal study of patients in whom changes in motor unit number over time are of importance, such as those with motor neuron diseases. PMID- 11268018 TI - Role of brain-derived neurotrophic factor in wobbler mouse motor neuron disease. AB - Brain-derived neurotrophic factor (BDNF) is neuroprotective for motoneurons undergoing degeneration, including those in natural motor neuron disease (MND) in wobbler mice. To assess the role of BDNF in this model of MND, endogenous BDNF immunoreactivity was analyzed by semiquantitative video-image analysis. Affected cervical spinal cord motoneurons had significantly greater BDNF immunoreactivity compared to motoneurons of healthy littermates (P = 0.01) and affected lumbar spinal cord motoneurons (P = 0.008 at age 4 weeks; P = 0.005 at age 8 weeks). Neuronal nitric oxide synthase (n-NOS) immunocytochemistry revealed increased immunoreactivity in the affected cervical spinal cord motoneurons. Exogenous BDNF treatment partially inhibited the increased NOS activity, as quantitatively measured by nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry. The mean number of NADPH-d(+) motoneurons in the cervical anterior horn decreased from 3.5 +/- 1.2 to 1.5 +/- 1.2 (P = 0.002). The increase in endogenous BDNF immunoreactivity in the affected spinal cord may be compensatory in diseased motoneurons, yet it appears to still be inadequate because exogenous BDNF treatment is required to suppress increased NOS activity in degenerating motoneurons. Our study indicates that BDNF is important in halting nitric oxide (NO)-mediated motor neuron degeneration, which has potential implications for the treatment of neurodegenerative disorders. PMID- 11268019 TI - Lower body negative pressure: a test of cardiovascular autonomic function. AB - Lower body negative pressure (LBNP) may provide an alternative test of cardiovascular autonomic function for patients unable to perform the Valsalva maneuver (VM). LBNP at -40 mmHg for 30 s was compared to the VM at 40 mmHg for 15 s with heart rate and blood pressure measured continuously in three age groups: 10-25 years; 26-40 years; and 41-55 years. Heart rate and blood pressure responses were comparable, with moderately diminished changes in blood pressure and heart rate in the LBNP test. When heart response to LBNP was converted to a ratio similar to that calculated for the VM, a high degree of correlation was found (R(2) = 0.5711). The LBNP test shows promise as an alternative test of cardiovascular autonomic function based on studies in normal subjects. The less marked changes may relate to the more passive nature of the applied stress. Future work should improve the device's accessibility and establish values for patients with autonomic disorders. PMID- 11268020 TI - The spread of transgene expression at the site of gene construct injection. AB - Seven 2-day-old golden retriever pups were given focal intramuscular injections of a first generation adenovirus-dystrophin minigene construct and adenovirus beta-galactosidase construct as a 2:1 mixture into the left anterior tibial muscle. The spread of transgene expression within the anterior tibial muscle was compared with the spread of methylene blue dye after identical injection into the contralateral muscle. Transgene expression 5-7 days after intramuscular injection was shown to extend between 5.8 and 11.6 mm along the biopsied muscle length (range of biopsy lengths 11.1-12.2 mm). The level of transgene expression at 2 2.5-mm intervals from the site of injection was significantly related to the distance from the site of injection (dystrophin, P = 0.009; beta-galactosidase, P = 0.015). The spread of methylene blue dye within the anterior tibial muscle < or =24 h after identical intramuscular injection demonstrated a similar pattern to the transgene expression, with dye staining measured between 5.5 and 8.5 mm along the muscle sample length (range of biopsy lengths 5.6-15.6 mm). The greatest transgene expression and dye staining was measured 2-2.5 mm proximal to the site of injection with a maximum of 23% of muscle fibers expressing the dystrophin transgene, 95.2% expressing the beta-galactosidase transgene, and 98% of the tissue section stained with methylene blue dye. These results suggest transgene expression after focal intramuscular injection is relatively localized around the site of injection. Further research is required to develop techniques that will provide transgene expression throughout the length and breadth of a muscle. PMID- 11268021 TI - Laser evoked potentials using the Nd:YAG laser. AB - Pain-related cortical potentials were evoked by skin stimulation of the face and the limbs with 5-ns-duration laser pulses delivered by a Q-switched Nd:YAG laser. Such laser pulses, in the nanosecond range, were able to induce pinprick pain sensations and to evoke reproducible laser evoked potentials (LEPs) without visible skin lesions for an energy density of less than 18 mJ/mm(2). Low energy densities, around 10 mJ/mm(2), were sufficient to reach the pain threshold and to induce LEP. The mean conduction velocity of the stimulated afferent fibers was close to 20 m/s, consistent with the stimulation of Adelta fibers. The amplitude of LEP correlated with pain perception rather than with energy density. The differences, such as wavelength and stimulus duration, between the Q-switched Nd:YAG laser we used and the lasers that are currently used in LEP studies (i.e., CO(2), argon, or Tm:YAG lasers in the millisecond range) are discussed. Our study opens novel perspectives in the LEP field of research by using a new type of laser with a very short pulse duration. PMID- 11268022 TI - The susceptibility of muscle cells to oxidative stress is independent of nitric oxide synthase expression. AB - The free radical, nitric oxide (NO.), has been implicated in the pathogenesis of muscular dystrophies because the enzyme, nitric oxide synthase (NOS), which produces NO., binds to the dystrophin-glycoprotein complex (DGC). In various studies of tissue samples from human and animal muscular dystrophies due to DGC defects, correlations between reductions of NOS activity and disease severity have been reported. To test for any direct effect of NOS expression on muscle cell susceptibility, we examined muscle cells in vitro under conditions of experimentally altered NOS activity. There were no differences in susceptibility to oxidative stress between differentiated myotube cultures from wild-type and from neuronal NOS (nNOS)-deficient mice. Likewise, pharmacological inhibition of NOS did not alter cellular susceptibility to oxidative challenges. Overexpression of NOS neither enhanced nor diminished cellular susceptibility to oxidative stress. Finally, we assessed the effect of NOS overexpression on myotube cultures from dystrophin-deficient (mdx) mice. NOS protein was localized to both membrane and cytosolic compartments in the transduced cells. Still, no difference in susceptibility to oxidative stress was found between the NOS-overexpressing cells and control cells. These data suggest that muscle cell susceptibility to oxidative challenges is independent of the level of NOS expression. Therefore, any role NO. may play in the pathogenesis of muscular dystrophies is likely to be independent of its effect on the redox state of the cell. PMID- 11268023 TI - Quality of life and well-being of patients with myasthenia gravis. AB - The cardinal symptom of myasthenia gravis (MG) is weakness of voluntary muscles, a feature that may restrict full participation in life activities. In turn, such limitations may negatively affect quality of life (QOL) and well-being among individuals with the disease. In the present study, we administered a measure of QOL to 27 patients with generalized MG. Results revealed that functional status was negatively impacted in the domains of physical functioning, energy, and general health. However, a clinically meaningful difference was evident only on perceived ability to accomplish physical tasks. The results suggest that although MG requires accommodations in physical activities, general QOL and well-being does not differ markedly from the general population. PMID- 11268024 TI - Temporal effects of inactivty on myosin heavy chain gene expression in rat slow muscle. AB - Myosin heavy chain (MHC) mRNA and protein profiles in adult rat soleus and adductor longus were determined after 4, 8, 15, 30, 60, and 90 days of spinal cord isolation (SI). SI results in complete neuromuscular inactivity while leaving the motoneuron-muscle fiber connections intact. From 15 to 90 days, type I MHC mRNA was significantly decreased, whereas type I MHC protein did not significantly decrease until 30 and 60 days in the soleus and adductor longus, respectively. However, in both muscles, slow MHC downregulation was offset by significant upregulation of the faster MHC isoforms, primarily IIx. From 60 to 90 days, type I MHC was almost completely replaced with faster isoforms at the mRNA and protein levels. Thus, chronic inactivity and unloading of slow rat hindlimb muscles shifted the MHC profile from predominately type I to type IIx MHC mRNA and protein. PMID- 11268025 TI - Length dependence of variables associated with temporal dispersion in human motor nerves. AB - Temporal dispersion in motor nerves is associated with changes of amplitude, area, duration, and Fourier spectra of compound muscle action potentials (CMAPs) when comparing responses to proximal and distal stimulation. These changes depend on the length of the nerve segment. To quantitatively assess this dependence, motor conduction studies of nerve segments of various lengths were performed in the median, ulnar, and tibial nerves of 86 test subjects, aged 4 to 73 years. Amplitude, area, duration, and spectral energy above 49 Hz of CMAPs were measured. Values after distal and proximal stimulation of each nerve segment were compared to determine amplitude decay, area decay, protraction, and high frequency attenuation. A significant length dependence of amplitude decay was found in the tibial and ulnar nerves, of area decay in the median and ulnar nerves, and of CMAP duration in the ulnar and tibial nerves. The length dependence of the high-frequency attenuation was significant in all nerves studied. This report provides normative data for variables associated with temporal dispersion. PMID- 11268027 TI - Adaptations in maximal motor unit discharge rate to strength training in young and older adults. AB - Six young (mean = 23 years) and 6 older (mean = 76 years) adults participated in isometric resistance training 5 days/week for 6 weeks. The task involved isometric fifth finger abduction. Maximal motor unit discharge rates (MUDRs) were obtained from the abductor digiti minimi of each hand at 0, 2, 14, and 42 days of training using a quadrifilar needle electrode and automatic spike recognition software. In agreement with previous findings, maximal MUDR at baseline was significantly lower in older adults (P < 0.001), averaging 51.5 (+/-17.13) HZ in young and 43.3 (+/-14.88) HZ in older adults. In response to resistance training, maximal voluntary force increased 25% in young and 33% in older subjects (P < 0.001). Maximal MUDR increased significantly (11% young, 23% older) on day 2 [F(3,36) = 2.58, P < 0.05], but in older subjects returned to baseline levels thereafter. These adaptations in abductor digiti minimi MUDR suggest a two-part response to strengthening fifth finger abduction: early disinhibition followed by altered MU activation. PMID- 11268026 TI - Inhibition of the initial wave of NF-kappaB activity in rat muscle reduces ischemia/reperfusion injury. AB - Nuclear factor kappaB (NF-kappaB) is thought to play an important role in the expression of genes expressed in response to ischemia/reperfusion (I/R) injury. In this report, the activation of NF-kappaB in rat skeletal muscle during reperfusion following a 4-h ischemic period was studied. NF-kappaB activation displayed a biphasic pattern, showing peak activities from 30 min to 3 h postperfusion and 6 h to 16 h postperfusion, with a decline to baseline binding activity levels between 3 h and 6 h. Inhibition of NF-kappaB activation was investigated using proline dithiocarbamate (Pro-DTC). NF-kappaB binding activity during reperfusion was significantly reduced by intravenous administration of Pro DTC. Additionally, Pro-DTC resulted in decreased muscle edema and neutrophil activity, with an increased percentage of muscle survival compared with vehicle controls. These results demonstrate that NF-kappaB is activated during reperfusion in a biphasic manner and that the regulation of the initial phase of NF-kappaB activation affords physiological protection against a severe ischemic stress. Selective inhibition of NF-kappaB during early reperfusion may therefore be a therapeutic intervention for I/R injury. PMID- 11268028 TI - Gender, body mass and age as risk factors for ulnar mononeuropathy at the elbow. AB - Factors that predispose patients to ulnar mononeuropathy at the elbow (UME) are poorly defined. We compared 112 electrodiagnostic reports which met criteria for definite or probable UME to 104 reports which excluded UME. Male gender was strongly associated with definite (OR = 6.9, 95% CI = 2.4-20.4; P < 0.001) and all UME (OR = 2.2, 95% CI = 1.2-4.1; P = 0.010) after controlling for age and body mass index (BMI). Among men, UME was associated with increasing age (P = 0.008) but not a decreased BMI. Women, however, demonstrated an association between decreased BMI and UME (OR = 2.3, 95% CI = 1.3-4.2 for BMI < or =22.0 versus >22.0). These findings, in conjunction with gender differences in ulnar motor nerve parameters among UME subjects and controls, suggest that the pathophysiology of UME differs with gender. PMID- 11268029 TI - Correlation between nerve conduction studies and clinical scores in diabetic neuropathy. AB - Polyneuropathy, a frequent complication of diabetes, can be assessed clinically and electrophysiologically. Neurological examination can be quantified by validated scores, e.g., the neuropathy symptom score (NSS) or the neuropathy disability score (NDS). Such scores exclude electrophysiological aspects of the neuropathy. A software tool was designed to convert electrophysiological data into one single index of polyneuropathy (IPN). This index was calibrated to grade the severity of a polyneuropathy from 0.00 to 1.00. In a series of 38 diabetic patients, we have calculated NSS, NDS, and IPN. We found correlations between these variables, NDS and IPN exhibiting the more significant association. The use of IPN allowed us to demonstrate that nerve conduction values correlated with clinical scores in diabetic polyneuropathy. Such a software tool, by providing a single electrophysiological index, may facilitate clinico-electrophysiological assessment in large descriptive studies or therapeutic trials of diabetic polyneuropathy. PMID- 11268030 TI - Conduction block in neuralgic amyotrophy. AB - We describe two cases of neuralgic amyotrophy with electrophysiological evidence of conduction block across the lower trunk of the brachial plexus. Low-output impedance stimulation of the cervical spinal roots in combination with collision was used to accurately demonstrate the conduction block. Complete electrophysiological recovery of the conduction block occurred within 3 months. Early clinical and electrophysiological recovery in both patients suggests that, in some cases, demyelination may predominate early in the course of neuralgic amyotrophy. PMID- 11268033 TI - O'Brave new world that has such technologies in it! PMID- 11268031 TI - The motor cortex and amyotrophic lateral sclerosis. AB - On theoretical grounds, abnormalities of the motor cortex in patients with amyotrophic lateral sclerosis (ALS) could lead to anterograde ("dying-forward") transneuronal degeneration of the anterior horn cells as suggested by Charcot. Conversely, retrograde ("dying-back") degeneration of the corticospinal tracts could affect the motor cortex. Evidence derived from clinical, neuropathological, static, and functional imaging, and physiological studies, favors the occurrence of anterograde degeneration. It is hypothesized that transneuronal degeneration in ALS is an active excitotoxic process in which live but dysfunctional corticomotoneurons, originating in the primary motor cortex, drive the anterior horn cell into metabolic deficit. When this is marked, it will result in more rapid and widespread loss of lower motor neurons. In contrast, slow loss of corticomotoneurons, as occurs in primary lateral sclerosis (PLS), precludes excitotoxic drive and is incompatible with anterograde degeneration. Preservation of slow-conducting non-M1 direct pathways in PLS is not associated with excitotoxicity, and anterior horn cells survive for long periods of time. PMID- 11268034 TI - Merging sex and position. AB - The choice between male and female development arises from a simple binary decision made in early development. Studies in a few model organisms led to a detailed understanding of the regulatory mechanisms that convey male or female identity at the cellular level. We have learned little, however, of how this information translates into the actual sexual phenotype with regionally dimorphic characters. Where does positional information come from and how does it integrate with the sex-determining pathway? A recent report sheds light onto this enigma and reveals possible intersections between sex-determining and homeotic pathways in Drosophila. Such intersections may also play an important part in evolution, providing a basis for phenotypic diversity among related species. BioEssays 23:304-306, 2001. PMID- 11268035 TI - The Ras pathway and spindle assembly collide? AB - Although alterations in Ras signalling are found in about 30% of human cancers, the transforming activity of oncogenic Ras is not fully understood. In a recent paper, a putative Ras1 effector in S. pombe, named Scd1, was reported to localize to mitotic spindles. Scd1 physically associates with Moe1, a factor that may contribute to the inherent instability of microtubules (MTs) and appears to be needed for proper spindle function. Altered MT dynamics within the spindle are likely to affect spindle assembly and chromosome capture, processes that need to be delicately controlled if cells are to guard against genome instability and transformation. BioEssays 23:307-310, 2001. PMID- 11268036 TI - Wnt signalling: a theme with nuclear variations. AB - Wnt proteins are involved in a large number of events during development and disease. The crucial element in the transduction of the signal elicited by Wnt is the state and activity of beta-catenin. There are two pools of beta-catenin, one associated with cadherins at the cell surface and a soluble one in the cytolasm, whose state and concentration are critical for Wnt signalling. In the absence of Wnt, the cytoplasmic pool is low due to targetted degradation of beta-catenin. Upon Wnt signalling, beta-catenin is stabilized. As a consequence, it can access the nucleus where it interacts with members of the Tcf family of transcription factors to modulate the expression of defined targets. Recent reports indicate that, in addition to Tcfs, beta-catenin can interact with other nuclear proteins raising the possibility that Wnt signalling has a wider modulatory effect on transcription than is mediated by its interactions with Tcfs. BioEssays 23:311 318, 2001. PMID- 11268037 TI - Control of retinal growth and axon divergence at the chiasm: lessons from Xenopus. AB - Metamorphosis in frogs is a critical developmental process through which a tadpole changes into an adult froglet. Metamorphic changes include external morphological transformations as well as important changes in the wiring of sensory organs and central nervous system. This review aims to provide an overview on the events that occur in the visual system of metamorphosing amphibians and to discuss recent studies that provide new insight into the molecular mechanisms that control changes in the retinal growth pattern as well as the formation of new axonal pathways in the central nervous system. BioEssays 23:319-326, 2001. PMID- 11268038 TI - Sound silencing: the Sir2 protein and cellular senescence. AB - The model organism Saccharomyces cerevisiae is providing new insights into the molecular and cellular changes that are related to aging. The yeast protein Sir2p (Silent Information Regulator 2) is a histone deacetylase involved in transcriptional silencing and the control of genomic stability. Recent results have led to the identification of Sir2p as a crucial determinant of yeast life span. Dosage, intracellular localization, and activity of Sir2p all have important effects on yeast longevity. For instance, calorie restriction apparently increases yeast life span by increasing Sir2p activity. Since Sir2p related proteins have been identified in many prokaryotic and eukaryotic organisms, the fundamental principles derived from the studies in yeast may prove valuable in directing our future research toward an understanding of the mechanisms of aging in higher eukaryotes. BioEssays 23:327-332, 2001. PMID- 11268040 TI - Innate immunity and its evasion and suppression by hymenopteran endoparasitoids. AB - Recent studies suggest that insects use pattern recognition molecules to distinguish prokaryotic pathogens and fungi from "self" structures. Less understood is how the innate immune system of insects recognizes endoparasitic Hymenoptera and other eukaryotic invaders as foreign. Here we discuss candidate recognition factors and the strategies used by parasitoids to overcome host defense responses. We suggest that host-parasitoid systems are important experimental models for studying how the innate immune system of insects recognizes foreign invaders that are phylogenetically more closely related to their hosts. The strategies used by parasitoids suggest that insects may employ "hidden-self" recognition molecules for attacking foreign objects intruding the open circulatory system. BioEssays 23:344-351, 2001. PMID- 11268039 TI - The molecular machinery for lysosome biogenesis. AB - The lysosome serves as a site for delivery of materials targeted for removal from the eukaryotic cell. The mechanisms underlying the biogenesis of this organelle are currently the subject of renewed interest due to advances in our understanding of the protein sorting machinery. Genetic model systems such as yeast and Drosophila have been instrumental in identifying both protein and lipid components of this machinery. Importantly, many of these components, as well as the processes in which they are involved, are proving conserved in mammals. Other recently identified components, however, appear to be unique to higher eukaryotes. BioEssays 23:333-343, 2001. Published 2001 John Wiley & Sons, Inc. PMID- 11268041 TI - The DAP kinase family of pro-apoptotic proteins: novel players in the apoptotic game. AB - The DAP (Death Associated Protein) kinase family is a novel subfamily of pro apoptotic serine/threonine kinases. All five DAP kinase family members identified to date are ubiquitously expressed in various tissues and are capable of inducing apoptosis. The sequence homology of the five kinases is largely restricted to the N-terminal kinase domain. In contrast, the adjacent C-terminal regions are very diverse and link individual family members to specific signal transduction pathways. There is increasing evidence that DAP kinase family members are involved in both extrinsic and intrinsic pathways of apoptosis and may play a role in tumor progression. This review will focus on structural composition and subcellular localization of DAP kinase family members and on signal transduction pathways leading to their activation. Potential mechanisms of DAP kinase family mediated apoptosis will be discussed. BioEssays 23:352-358, 2001. PMID- 11268042 TI - Human infertility, reproductive cloning and nuclear transfer: a confusion of meanings. AB - The Chief Medical Officer of Health of the United Kingdom has recommended that the 1990 Human Fertilisation and Embryology Act should be amended to allow cloning in humans for research purposes only. He also recommended that: "The transfer of an embryo created by cell nuclear replacement into the uterus of a woman (so called 'reproductive cloning') should remain a criminal offence" (recommendation 7, Ref. 1). This recommendation implies that nuclear replacement and cloning are the same. They are not. Nuclear transfer constitutes reproductive cloning only when the individual created is genetically identical to the nuclear donor. In this paper, we describe a possible future use of nuclear transfer for the treatment of infertile individuals. The treatment yields an individual that receives approximately equal genetic contributions from each parent. We use this example to illustrate how semantic confusion might lead to plausibly moral and justifiable treatments being legally banned. In doing so, we hope to encourage a more accurate and informed use of language in science, law and politics, so that legislation is properly informed by science and achieves what it intends. BioEssays 23:359-364, 2001. PMID- 11268043 TI - Nadine Dobrovolskaia-Zavadskaia and the dawn of developmental genetics. AB - In one of the first genetic screens aimed at identifying induced developmental mutants, Nadine Dobrovolskaia-Zavadskaia, working at the Pasteur Laboratory in the 1920s, isolated and characterized a mutation affecting Brachyury, a gene that regulates tail and axial development in the mouse. Dobrovolskaia-Zavadskaia's analysis of Brachyury and other mutations affecting tail development were among the earliest attempts to link gene action with a tissue-specific developmental process in a vertebrate. Her analyses of genes that interacted with Brachyury led to the discovery of the t-haplotype chromosome of mouse. After 70 years, Brachyury and the multiple genes with which it interacts continue to occupy a prominent focus in developmental biology research. A goal of this review is to identify the contributions that Dobrovolskaia-Zavadskaia made to our current thinking about Brachyury and how she helped to shape the dawn of the field of developmental genetics. BioEssays 23:365-371, 2001. PMID- 11268044 TI - Calcium in development: from ion transients to gene expression. PMID- 11268046 TI - Measurement and pharmacokinetic analysis of unbound cephaloridine in rat blood by on-line microdialysis and microbore liquid chromatography. AB - A technique involving rapid sampling of cephaloridine in rat blood was achieved using a combination of microdialysis and sensitive microbore liquid chromatography. A microdialysis probe was inserted into the jugular vein/right atrium of a Sprague-Dawley rat. Then after a real-time collection of the analyte by microdialysis, the dialysate was automatically injected into a liquid chromatographic system via an on-line injector. Following a 2 h stabilization period after the surgical procedure, cephaloridine (20 mg/kg, i.v.) was then administered via the femoral vein. Isocratic elution of cephaloridine was carried out with a mobile phase containing methanol-20 mM monosodium phosphate (25:75, v/v, pH 5.5), and the flow rate of the mobile phase was 0.05 mL/min within 10 min. Intra- and inter-assay accuracy and precision of the assay were each less than 10%. The in vivo recovery of the cephaloridine from the microdialysate was 49.7 +/- 8.0% and 42.4 +/- 8.4% for 0.5 and 1 microg/mL standards (n = 6), respectively. Based on the pharmacokinetic analysis, the elimination half-life was 32.2 +/- 8.6 min by cephaloridine administration (20 mg/kg, i.v., n = 6). PMID- 11268045 TI - Determination and pharmacokinetic study of 1-p-(3.3-dimethyl-1-triazeno) benzoic acid in cancer patients by capillary gas chromatography. AB - A capillary gas chromatographic method was developed for determining 1-p-(3.3 dimethyl-1-triazeno) benzoic acid in the plasma and urine of cancer patients under pharmacokinetic study. The drug was extracted with ethyl acetate and methylated with diazomethane. Octadelane (10 microg/ml) was added as internal standard. The separation was carried out on an OV-1 quartz capillary column, 15 m x 0.32 mm (0.52 microm), with high-purity nitrogen as carrier gas and flame ionization detector (FID) as detector. The column temperature was held at 130 degrees C for 9 min and then programmed to 240 degrees C, at a rate of 35 degrees C/min. The temperature of both injector and detector was 260 degrees C. The standard curve was linear from 0.4 to 40 microg/mL in plasma, and from 0.8 to 20 microg/mL in urine, with correlation coefficients of 0.9979 and 0.9932. The relative standard deviations (RSD) were less than 9.7%. The minimum recovery of this method was 81.8%. This method was applied to the pharmacokinetic studies of 1-p-(3.3-dimethyl-1-triazeno) benzoic acid in cancer patients after a single dose (i.v.) of 160, 420 or 760 mg/m(2) was administered. They all conformed to the two compartment open model and showed linear pharmacokinetics. The excretion of this drug in the urine was minimal. PMID- 11268047 TI - Capillary electrophoresis of catecholamines with laser-induced fluorescence intensified charge-coupled device detection. AB - A capillary electrophoresis-laser-induced fluorescence-intensified charge-coupled device system was used for the separation and determination of catecholamines. Optimization of derivatization and separation conditions was investigated in order to reach a high separation efficiency and sensitivity. All fluorecein isothiocyanate derivatives of catecholamines were satisfactorily separated within 12 min. The detection limits were in attomole ranges. This method allows determination of catecholamines with high separation efficiencies and sensitivity. PMID- 11268048 TI - Simultaneous determination of flucytosine and fluorouracil in human plasma by high-performance liquid chromatography. AB - A validated, sensitive and precise reversed-phase high-performance liquid chromatographic method for the simultaneous determination of 5-flucytosine (5-FC) and 5-fluorouracil (5-FU) in human plasma is described. Two compounds, 5 methylcytosine (5-MC) and 5-chlorouracil (5-CU), were used as internal standards for the determination of 5-FC and 5-FU, respectively. Plasma samples were deproteinized with trichloroacetic acid and chromatographed on an octylsilica column, maintained at 30 degrees C during elution, using a 0.04 M phosphate buffer, pH 7.0, as eleunt. Spectrophotometric diode array detection was used at 266 nm. 5-FC, 5-FU, 5-MC and 5-CU were found to have retention times of 4.8, 5.8, 7.7 and 11.0 min respectively. Recoveries of 91-120% with reproducibility and repeatability coefficients of variation of 0.8-6% were obtained. Mean correlation coefficients of 0.99989 and 0.9995 were found for the linear calibration curves (n = 2) of 5-FC (4.816-192.6 mg/l) and 5-FU (0.05368-5.368 mg/l), respectively. The limits of quantitation were 0.3 mg/l for 5-FC and 0.05 mg/l for 5-FU. PMID- 11268049 TI - A fluorimetric, column-switching HPLC and its application to an elimination study of LLU-alpha enantiomers in rat plasma. AB - A method for the enantiomeric determination of 2,7,8-trimethyl-2-(beta carboxyethyl)-6-hydroxy chroman (LLU-alpha, gamma-CEHC) in rat plasma was developed using high-performance liquid chromatography (HPLC) with a fluorimetric derivatization with 4-N, N-dimethylaminosulfonyl-7-piperazino-2,1,3 benzoxadiazole (DBD-PZ) followed by O-acetylation with acetyl chloride. The proposed HPLC system used two non-chiral columns (phenyl and octadecylsilica) and a chiral column (a modified cellulose type), which were connected via two column switching valves. A derivatized sample prepared from rat plasma was first separated on the phenyl column, and the fraction including LLU-alpha derivative was introduced to the octadecylsilica column to quantify the concentration of the mixture of S- and R-LLU-alpha. Finally, the LLU-alpha derivative was directly injected into the chiral column to obtain the ratio of the enantiomers. The proposed HPLC system was applied to the enantiomeric determination of LLU-alpha in plasma after intravenous administration of racemic LLU-alpha. S-LLU-alpha was eliminated faster than R-LLU-alpha, and its concentration in plasma decreased to one-third at 2 min after dosing. PMID- 11268050 TI - Determination of amphetamine in dog plasma by gas chromatography with mass selective detection. AB - This paper describes the validation of an analytical method for the determination of amphetamine in beagle dog plasma by gas chromatography coupled to mass spectrometry (GC-MS). d-Amphetamine-d(6) was used as the internal standard. The method consisted of a rapid single-step liquid-liquid extraction and derivatization of amphetamine with 2,2,2-trichloroethyl chloroformate, followed by sensitive GC-MS detection. This is the first report utilizing the combination of trichloroethyl chloroformate as a derivatization reagent and a deuterated amphetamine analog as an IS for the quantification of amphetamine in plasma. The method was validated in terms of specificity, curve fit, precision, accuracy, recovery and stability, and was acceptable according to FDA draft guidelines for validation of bioanalytical methods. The limit of detection was 0.65 ng/mL. The calibration range was 5-150 ng/mL. The validated method was successfully employed for the quantitation of amphetamine in dog plasma samples for pharmacokinetic profiling. PMID- 11268051 TI - Validation study of assay method for DX-8951 and its metabolite in human plasma and urine by high-performance liquid chromatography/atmospheric pressure chemical ionization tandem mass spectrometry. AB - A new liquid chromatographic/mass spectrometric assay has been developed for the determination of DX-8951, a new anti-tumor drug, and its 4-hydroxymethyl metabolite (UM-1) in human plasma and urine. Solid-phase extractions were used for sample preparation. A gradient reverse-phase HPLC separation was developed with mobile phases consisting of trifluoroacetic acid and methanol. The detection was conducted using atmospheric pressure chemical ionization tandem mass spectrometry in the selected reaction monitoring mode. A structural analog, camptothecin (CPT), was used as the internal standard. The assay was validated for the determination of DX-8951 and UM-1 in human plasma and urine. The lower limits of quantitation of DX-8951 and UM-1 were 0.1 ng/mL in plasma and 1 ng/mL in urine. The method showed a satisfactory sensitivity, precision, accuracy, recovery and selectivity. PMID- 11268052 TI - Chiral derivatization reagents for drug enantioseparation by high-performance liquid chromatography based upon pre-column derivatization and formation of diastereomers: enantioselectivity and related structure. AB - Structures and related enantioselectivities of the respective chiral derivatization reagents (CDRs) for drug enantioseparation by high-performance chromatography based upon pre-column derivatization and diastereomeric formation are reviewed. The elution order of diastereomers caused reaction of some CDRs with enantiomeric amino acids and carboxylic acids. The development of new CDRs available for indirect HPLC methods is also discussed. (c) 2001 John Wiley & Sons, Ltd. PMID- 11268053 TI - Determination of 24,25-dihydroxyvitamin D(3) in human plasma using liquid chromatography-mass spectrometry after derivatization with a Cookson-type reagent. AB - A practical LC-MS method for determination of (24R)-24,25-dihydroxyvitamin D(3) [24,25(OH)(2)D(3)] in human plasma has been developed using derivatization with a Cookson-type reagent, 4-[4-(6-methoxy-2-benzoxazolyl)phenyl]-1,2,4-triazoline-3,5 dione (MBOTAD). The derivatization with MBOTAD significantly improved the ionization efficiency of the analyte with a detection limit of 18 fmol [equivalent to 7.5 pg of 24,25(OH)(2)D(3)] per injection. The method employed protein precipitation with acetonitrile, purification with OASIS HLB cartridge and silica gel column, derivatization with MBOTAD and atmospheric pressure chemical ionization MS detection. The mass spectrometer was operated in the positive-ion mode of mass chromatography and [26,26,26,27,27,27-(2)H(6)] 24,25(OH)(2)D(3) was used as an internal standard. The intra- and inter-assay coefficients of variation were below 3.4 and 2.5%, respectively, and the analytical recovery of 24,25(OH)(2)D(3) was quantitative. Assay linearity was obtained in the range of 0.05-1.2 ng per tube and the limit of quantitation was 0.23 ng/mL for a 0.3 mL plasma aliquot. The developed method was applied to plasma samples obtained from volunteers and gave satisfactory results. PMID- 11268054 TI - Stereoselective determination of p-hydroxyphenyl-phenylhydantoin enantiomers in rat liver microsomal incubates by reversed-phase high-performance liquid chromatography using beta-cyclodextrin as chiral mobile phase additives. AB - An analytical method was developed for determination of p hydroxyphenylphenylhydantoin enantiomers in rat liver microsome by using reversed phase high-performance liquid chromatography. A 50 mm C(8) column was used as the analytical column. The mobile phase was made up of 8.8 mmol/L beta-cyclodextrin, 0.25 mol/L urea and 0.05 mol/L ammonium acetate in water. The assay was linear from 2.05 to 410.0 micromol/L for each enantiomer. The limits of detection and of quantitation for the method were 0.90 and 2.05 micromol/L for each enantiomer, respectively. The analytical method afforded average recoveries of 93.59 +/- 2.75% and 94.72 +/- 1.78% for S- and R-p-hydroxyphenylphenylhydantoin, respectively. The method allowed study of the in vitro glucuronidation of p hydroxyphenylphenylhydantoin in rat liver microsomal incubates. The stereoselectivity of p-hydroxyphenylphenylhydantoin phase II metabolism was observed. PMID- 11268055 TI - Sperm storage in the oviduct of the internal fertilizing frog Ascaphus truei. AB - This study provides the first descriptions of sperm storage at the tissue and cellular levels in a female frog or toad. Oviducal anatomy was studied by light and electron microscopy in Ascaphus truei from north coastal California. Ascaphus truei is one of the few species of anurans in which fertilization is internal. Unlike other anurans with internal fertilization, however, mating in A. truei consists of a unique combination of amplectic and copulatory mechanisms that we term "copulexus." Posterior to a short, aglandular infundibular region, the oviduct possesses: 1) a proximal, convoluted ampullary region where intrinsic tubular glands secrete gelatinous envelopes around eggs; 2) a middle ovisac region where fertilization occurs; and 3) a distal oviducal sinus formed by medial junction of the ovisacs. Sperm storage tubules (SSTs) occur in the anterior portions of the ovisacs and consist of simple tubular glands. SSTs and the rest of the oviducal lining stain positively with the periodic acid-Schiff's procedure for neutral carbohydrates and this reaction is especially intense in reproductively active females. Sperm were found in the SSTs of gravid females as well as some nonvitellogenic females. The sperm are in orderly bundles in the SSTs, and although occasionally sperm nuclei were embedded in the epithelium, no evidence for spermiophagy was found. Oviducal sperm storage in A. truei is homoplastic, with closest structural similarities to squamate reptiles. Oviduct/sperm design constraints appear to limit the options for expression of features associated with oviducal sperm storage. PMID- 11268056 TI - Digestive, salivary, and reproductive organs of Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) B type. AB - A microscopic analysis of the morphology and ultrastructure of the digestive, salivary, and reproductive systems of adult Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) B type was conducted using light, scanning, and transmission electron microscopy. The internal anatomy of B. tabaci was found to be similar to that reported for Trialeurodes vaporariorum. In a microscopic analysis of the salivary glands, we have shown that each primary salivary gland is composed of at least 13 cells varying in morphology and staining differentially, while the accessory salivary glands are composed of four morphologically similar cells. We analyzed the course of the alimentary canal in B. tabaci, demonstrated the internal morphology of the organs, and clarified the location of the filter chamber relative to other organs in the whitefly. Our observations confirm that the pair of structures extending from the connecting chamber are caeca that may aid in fluid movement through the midgut and are not Malpighian tubules, as previously suggested. We confirm an earlier finding that the whitefly lacks Malpighian tubules, having instead specialized Malpighian-like cells within the filter chamber at the juncture with the internal ileum. Finally, we provide the first scanning electron microscopic analysis showing the reproductive organs of B. tabaci. Our investigation provides clarified terminology for several components of the digestive and excretory system. We also provide drawings and micrographs that will aid future researchers in localizing the internal organs of B. tabaci. We expect our analysis to provide a valuable tool for studying B. tabaci / plant virus interactions and physiological and biological aspects of this insect. PMID- 11268057 TI - Fine structure and development of the retina of the grenadier anchovy Coilia nasus (Engraulididae, Clupeiformes). AB - A study of the morphogenesis of the grenadier anchovy retina was undertaken using light and electron microscopy. Five developmental stages from prelarvae 3 days after fertilization to adult fish were studied. In addition to the general morphology of the eye and retina, special emphasis was given to the development of the photoreceptors and pigment epithelium (PE). The earliest retinae showing structural features indicative of a functioning eye are pure cone retinae composed of rows of alternating long and short cones forming a transient, tesselated pattern. At this stage there is a conventional PE containing melanin. In older stages cone rows are separated by the newly formed rods and by PE wedges filled with diffusely reflecting guanine crystallites. The findings are compared with the retinae of other engraulidids and with the development of teleost retinae in general. Moreover, the observed structural changes are discussed with respect to the photic habitat conditions of these anadromous fish that move between coastal waters, estuary, and river. PMID- 11268058 TI - Mineralized dermal layer of the Brazilian tree-frog Corythomantis greeningi. AB - Some species of anuran amphibians possess a calcified dermal layer (the Eberth Kastschenko layer) located between the "stratum spongiosum" and the "stratum compactum." This layer consists of calcium phosphate deposits, proteoglycans, and glycosaminoglycans. Although regarded as a protective layer against desiccation, a calcium reservoir, or possibly a remnant of a dermal skeleton present in anuran ancestors, very little is known about its origin, structure, and function. Thus, we studied the structure and composition of the mineralized dermal layer of Corythomantis greeningi, a peculiar hylid from the Brazilian semiarid region (caatinga), using conventional and cryosubstitution methods combined with transmission, scanning, and analytical electron microscopy. Results show that the dermal layer consists of dense, closely juxtaposed, globular structures. Although the electron opacity of the globules was variable, depending on the type of preparation, crystal-like inclusions were present in all of them, as confirmed by dark field microscopy. Electron probe X-ray microanalysis showed calcium, phosphorus, and oxygen, and electron diffraction revealed a crystalline structure comparable to that of a hydroxyapatite. PMID- 11268059 TI - Keratohyalin-like granules in lizard epidermis: evidence from cytochemical, autoradiographic, and microanalytic studies. AB - Epidermal sloughing in lizards is determined by the formation of an intraepithelial shedding complex in which keratohyalin-like granules are formed. The chemical nature of these granules is unknown, as is their role in keratinization. The goal of this study was to test whether they contain some amino acids similar to those found in mammalian keratohyalin. The embryonic and regenerating epidermis of lizards are useful systems to study the formation of these granules. Histochemically keratohyalin-like granules react to histidine and contain some sulfhydryl groups (cysteine). X-ray microanalysis shows that these granules contain sulfur and often phosphorus, two elements also present in the mature clear, oberhautchen, and beta layer. Instead the mesos, alpha, and lacunar layers contain only sulfur. Most sulfur is probably in a disulfide-bonded form, particularly in mature cells of the shedding complex, in large keratohyalin-like granules, and in the beta-keratin layer. Early differentiating beta-keratin cells have the maximal incorporation of tritiated proline, whereas tritiated arginine is slightly more concentrated in the basal layer of the epidermis. A high uptake of tritiated histidine is observed mainly in keratohyalin-like granules of the clear layer, but also in the oberhautchen layer and forming the alpha-lacunar layer. Immunogold electron microscopy shows that keratohyalin-like granules do not localize keratin but are embedded within a keratin network. These results suggest that keratohyalin-like granules of lizards, like mammalian keratohyalin, contain some sulfur-rich and histidine-rich proteins. These granules participate in the process of hardening of the clear layer that molds the spinulae of the deeper oberhautchen to form the superficial microornamentation. PMID- 11268062 TI - Self-Assembling Organic Nanotubes. AB - Hollow tubular structures of molecular dimensions perform diverse biological functions in nature. Examples include scaffolding and packaging roles played by cytoskeletal microtubules and viral coat proteins, respectively, as well as the chemical transport and screening activities of membrane channels. In the preparation of such tubular assemblies, biological systems make extensive use of self-assembling and self-organizing strategies. Owing to numerous potential applications in areas such as chemistry, biology, and materials science considerable effort has recently been devoted to preparation of artificial nanotubular structures. This article reviews design principles and the preparation of synthetic organic nanotubes, with special emphasis on noncovalent processes such as self-assembly and self-organization. PMID- 11268060 TI - The dorsal fin engine of the seahorse (Hippocampus sp.). AB - The muscles, fin ray joints, and supporting structures underlying the dorsal fin are described for two seahorse species: Hippocampus zosterae and Hippocampus erectus. A fan-shaped array of cartilaginous bones, the pterigiophores, form the internal supporting structure of the dorsal fin. Each pterigiophore is composed of a proximal radial that extends from a vertebra to the dorsal side of the animal, where it fuses to a middle radial. The middle radials fuse with each other to form a dorsal ridge upon which sit the spheroidal distal radials. Each distal radial articulates with a fin ray on its dorsal side and is attached to the dorsal ridge on its ventral side by a material that has been histologically identified as elastic cartilage. Together these connections form a two-axis joint that permits elevation, depression, and inclination of the ray. Each fin ray is actuated by two bilateral pairs of muscles, an anterior pair of inclinators, and a posterior pair of depressors. The anteriormost fin ray is actuated by three bilateral pair of muscles, the inclinators, the depressors, and a pair of elevator muscles that are positioned anterior to the inclinators. Preliminary examinations of the ray joints of the pectoral and anal fins of adult H. zostera and the pectoral fins of newborn H. erectus revealed structures similar to that seen in the dorsal fins. To further explore the structure and function of the dorsal fin gross dissections and simple functional tests were performed on H. erectus and H. barbouri and behavioral observations were made of all three species plus Hippocampus kuda. PMID- 11268063 TI - Tracelessness Unmasked: A General Linker Nomenclature. AB - Nowadays it is rare to find an issue of a major chemistry journal without at least one article on solid-phase synthesis. This is hardly surprising: the technique promises an end to arduous work-up procedures and the ability to facilitate the creation of vast libraries of compounds using combinatorial techniques. No longer is the technique only of interest to those involved in peptide synthesis: an enormous variety of product classes have now been prepared on and isolated from the solid phase. It is the "linker" which is the focus of this article. The linker's ultimate function is to release a product from the support into solution: it does this, without exception, with a chemical change to the product at the former linkage site. Some linkers, apparently, are "traceless". But what, in fact, is "tracelessness"? Twenty years ago, in a climate where cleavage of a linker resulted in formation of a polar carboxylic acid as the vestige of the support, the concept was attractive. Today the chemist is faced with a myriad of novel linkers which have the ability to release products bearing most major functionalities at the former linkage site and we will argue here that the term "traceless", although currently in widespread use, is meaningless. Instead, we propose a new categorization of linkers based on the functionality they release upon cleavage, and suggest a nomenclature to underpin this categorization. We anticipate that the article will also serve to highlight areas of linker technology in need of further research. PMID- 11268064 TI - Protean. PMID- 11268065 TI - Plastic Transistors Reach Maturity for Mass Applications in Microelectronics I wish to thank the Fonds der Chemischen Industrie and the Deutsche Forschungsgemeinschaft for a Liebig and a Habilitanden fellowship, and Prof. Peter Bauerle for numerous helpful discussions especially about the topic reviewed in this article. PMID- 11268066 TI - Labeling of Biomolecules for Medicinal Applications-Bioorganometallic Chemistry at Its Best The author thanks A. P. Schubiger and D. Grotjahn for reprints, and K. Severin for insightful discussions. The data in Figure 1 are used with the help and kind permission of M. Salmain and G. Jaouen. I am also grateful to Drs. H. & H. Weiss for their hospitality when writing this article. PMID- 11268067 TI - Highly Efficient and Ultrafast Phototriggers for cAMP and cGMP by Using Long Wavelength UV/Vis-Activation This work was supported by the Deutsche Forschungsgemeinschaft, the European Union, and the Fonds der Chemischen Industrie. We thank B. Dekowski and J. Lobetamann for technical assistance and S. Hecht for proof reading. PMID- 11268068 TI - Analysis of the Topology of the Chromophore Binding Pocket of Phytochromes by Variation of the Chromophore Substitution Pattern. PMID- 11268069 TI - Isolated Hexagonal Channels Built up by Three-Connected Ge(-) Ions in LiGe at High Pressure This work was supported by the Deutsche Forschungsgemeinschaft and the Fonds der Chemischen Industrie. PMID- 11268070 TI - A New Phase-Switch Method for Application in Organic Synthesis Programs Employing Immobilization through Metal-Chelated Tagging We gratefully acknowledge financial support from Pfizer Global Research and Development, the University of Ferrara (Italy, Postdoctoral Fellowship to A.M.), the EU (Marie Curie Fellowship to F.R.), and a Novartis Research Fellowship (to S.V.L.). PMID- 11268071 TI - 1H High-Resolution Magic Angle Spinning NMR Spectroscopy for the Investigation of a Ras Lipopeptide in a Lipid Membrane This work was supported by the Deutsche Forschungsgemeinschaft (SFB 197 and Innovationskolleg "Chemisches Signal und biologische Antwort"). D.H. is grateful for a BASF fellowship through the Studienstiftung des Deutschen Volkes. The authors thank Dr. A. Pampel for technical support with the DRX600. PMID- 11268072 TI - Enantioselective Synthesis of the Ricciocarpins A and B This work was supported by the Deutsche Forschungsgemeinschaft (Graduierten-Kolleg "Struktur-Eigenschafts Beziehungen bei Heterocyclen") and the Fonds der Chemischen Industrie. We thank Prof. Dr. A. Furstner, MPI Mulheim, for a sample of catalyst 9. PMID- 11268073 TI - The Ammonia-Synthesis Catalyst of the Next Generation: Barium-Promoted Oxide Supported Ruthenium. PMID- 11268074 TI - A New Photochemical Route to Cyclopropanes We gratefully thank the Deutsche Forschungsgemeinschaft for financial support (grant no.: We 1850/3-1). PMID- 11268075 TI - Formation of Super Wires of Clusters by Self-Assembly of Transition Metal Cluster Anions with Metal Cations. PMID- 11268076 TI - Heteroepitaxial Growth of a Zeolite T.W. and T.O. are grateful to H. Tsunakawa of the High Voltage Electron Microscope Laboratory, University of Tokyo (UT), and Prof. Y. Ikuhara, Engineering Research Institute, UT, for the 400-kV SAED experiments and their analyses, respectively. H. Shiga, T. Hayashi and T. Shiraki are acknowledged for preliminary experiments. This work was supported by a Grant in-Aid for Scientific Research from the Ministry of Education, Science, Sports and Culture and the TEPCO Foundation. PMID- 11268077 TI - Rotaxane-Encapsulation Enhances the Stability of an Azo Dye, in Solution and when Bonded to Cellulose This work was supported by the Engineering and Physical Sciences Research Council (UK) and by BASF plc. PMID- 11268078 TI - 2-Pyridyldimethylsilyl as a Removable Hydrophilic Group in Aqueous Diels-Alder Reactions This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science, Sports, and Culture, Japan, and in part by the Mitsubishi Foundation. We thank Professor Kazunari Akiyoshi (Kyoto University) for assistance with dynamic light-scattering experiments and fruitful discussions. Professor Susumu Yoshikawa and Professor Masahiro Kinoshita (Kyoto University) are gratefully acknowledged for critical discussions. PMID- 11268079 TI - The Combination of Spontaneous Resolution and Asymmetric Catalysis: A Model for the Generation of Optical Activity from a Fully Racemic System We thank the Centre National de la Recherche Scientifique and Rhodia for financial support, Prof. H. B. Kagan for helpful discussions, and H. Ait-Haddou and O. Hoarau for assistance in catalytic tests. PMID- 11268080 TI - Assembly of Nano-Scale Circular Supramolecular Arrays through pi-pi Aggregation of Arc-Shaped Helicate Units This work was supported by the University of Warwick (L. J. C.). We thank the EPSRC and Siemens Analytical Instruments for grants in support of the diffractometer and the EPSRC National Mass Spectrometry Centre, Swansea, for recording the mass spectra. PMID- 11268081 TI - A Dendritic Structure Containing a Designed Cleft which Controls Ligand Coordination Behavior in an Analogous Way to Proteins This work was supported by the Leverhulme Trust. We thank the University of Warwick National 9.4T FT-ICR Mass Spectrometry Facility (EPSRC/BBSRC) for recording the FT-ICR mass spectra and the EPSRC National Mass Spectrometry Centre, Swansea, for recording other mass spectra. PMID- 11268082 TI - [30]Metallacrown-10 Compounds: [Mn] PMID- 11268083 TI - Coordination Chemistry in the Solid: Study of the Incorporation of Cu(II) into Cyclam-Containing Hybrid Materials. PMID- 11268084 TI - Synthetic Studies on Ciguatoxin: A Highly Convergent Synthesis of the GHIJKLM Ring System Based on B-Alkyl Suzuki Coupling. PMID- 11268085 TI - Turning On Cell Migration with Electroactive Substrates We are grateful for support of this work by DARPA and the National Institute of Health (GM 54621). This work used facilities of the MRSEC supported by the National Science Foundation (DMR-9808595). M. M. is a Searle Scholar and an A. P. Sloan Fellow. B. T. Houseman is supported by MD/PhD Training Grant HD-09007. PMID- 11268086 TI - Asymmetric Induction by Helical Hydrocarbons: PMID- 11268087 TI - Interprotein Electron Transfer Reaction Regulated by an Artificial Interface We thank Prof. Dr. I. Morishima and his group for the arrangement of laser flash photolysis equipment. This work was supported by Nagase Science and Technology Foundation, and a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture, Japan. Y. H. was supported by Research Fellowships of the Japanese Society for the Promotion of Science for Young Scientists. PMID- 11268088 TI - Three-Dimensional Transmission Electron Microscopic Observations of Mesopores in Dealuminated Zeolite Y Supported by NWO under grant 98037. The research of A.J.K. has been made possible by a fellowship of the Royal Netherlands Academy of Arts and Sciences (KNAW). The authors thank J. E. M. J. Raaymakers for the nitrogen physisorption measurements, A. J. M. Mens for the XPS measurements, J. A. R. van Veen and E. J. Creyghton for physical data and useful discussions, and Shell International Chemicals and Zeolyst for the samples. PMID- 11268089 TI - Fluorescence Detection of Specific RNA Sequences Using 2'-Pyrene-Modified Oligoribonucleotides We are very grateful to Professor Hiroshi Sugiyama, Dr. Tetsuji Yamaoka, and Dr. Takashi Morii for ion-spray mass spectrometric measurements and helpful comments on this research. PMID- 11268090 TI - Versatile Approaches to the Polymer-Supported Synthesis of Bidentate Phosphorus Containing Ligands. PMID- 11268091 TI - Enantioselective Reduction of Ketones Catalyzed by Polymer-Supported Sulfonamide Using NaBH(4)/Me(3)SiCl (or BF(3) small middle dotOEt(2)) as Reducing Agent This work was supported by the Ministry of Sciences and Technology, the State Key Project of Basic Research (Project 973, No. G 2000048007). PMID- 11268092 TI - An Asymmetric Enzyme-Catalyzed Retro-Claisen Reaction for the Desymmetrization of Cyclic beta-Diketones This work was supported by the BBSRC (grants to N.J.T. and S.L.F.) and the European Commission (grant to J.G.). We also thank Mr. J. Millar and Dr. D. Uhrin for assistance with NMR spectroscopy. PMID- 11268093 TI - Synthesis of the First (1-3:6,7-eta-Cyclodecadienyl)ruthenium Complex by the Intramolecular Reaction of an Alkene and a Vinylcyclopropane We thank the National Science Foundation and the National Institutes of Health (NIH), General Medical Sciences, for their generous support of our programs. We are grateful to A. Cole for solving the X-ray structure of 3. Mass spectra were provided by the Mass Spectrometry Facility of the University of California, San Francisco, supported by the NIH Division of Research Resources. PMID- 11268094 TI - Stereoselective Cross Pinacol-Type Coupling between alpha,beta-Unsaturated Ketones and Aldehydes Mediated by Chromium(II) and R(3)SiCl Financial support by a Grant-in-Aid for Scientific Research on Priority Area No. 706 from the Ministry of Education, Culture, Sports, Science and Technology of Japan is gratefully acknowledged. PMID- 11268095 TI - Ligand-Controlled Chemoselectivity in the Classical Oxidative Addition Reactions of MeI and Aldehydes to Rhodium(I) Complexes This work was supported by the Israel Science Foundation, Jerusalem, Israel, by the MINERVA foundation, Munich, Germany, and by the Tashtiyot program of the Israeli Ministry of Science. D.M. is the holder of the Israel Matz Professorial Chair of Organic Chemistry. We thank Dr. L. Shimon and Dr. H. Rozenberg, Weizmann Institute of Science, for performing the X-ray structural analysis. PMID- 11268096 TI - Quasi One-Dimensional Organic Superconductor MDT-TSF small middle dotAuI(2) with T(c)=4.5 K at Ambient Pressure This work was supported by Grants-in-Aid for Scientific Research from the Ministry of Education, Science, Sports, and Culture of Japan. We thank the Cryogenic Center, Hiroshima University, for supplying liquid helium. PMID- 11268097 TI - Synthesis of PMID- 11268098 TI - Total Synthesis of (+)-Pyrenolide D This research was supported by the National Institutes of Health, Glaxo Wellcome Inc., the Alfred P. Sloan Foundation, and the Arnold and Mabel Beckman Foundation. PMID- 11268100 TI - arachno- PMID- 11268099 TI - Crown-Ether-Directed Assembly of Discrete and One-Dimensional Silver Aggregates Containing Embedded Acetylenediide Financial support from the Hong Kong Research Grants Council Earmarked Grant (CUHK 4268/00P) is gratefully acknowledged. PMID- 11268101 TI - Mass Spectrometry of an Intact Virus The authors gratefully acknowledge Jennifer Boydston for her helpful comments and suggestions. G.S. is grateful for support from the NIH (GM55775). The work at LBL was supported by the Director, Office of Energy Research, Office of Health and Environmental Research, Human Genome Program, U.S. Department of Energy under contract number DE-AC03-76SF00098. PMID- 11268102 TI - No Title. PMID- 11268103 TI - Biologically active compounds from marine organisms. PMID- 11268104 TI - Stimulation of protein biosynthesis in rat hepatocytes by extracts of Momordica charantia. AB - The in vivo effect of the administration of extracts of Momordica charantia on certain biochemical parameters of Sprague-Dawley rats was investigated. It was observed that there was an increase in muscle and liver protein levels, while there was a reduction in the levels of brain protein, muscle and liver glycogen. The activities of plasma L-alanine transaminase and alkaline phosphatase were reduced. The L-aspartate transaminase and adenosine triphosphatase activities were slightly elevated in whole plant extract treated rats while the L-aspartate transaminase was unaffected by the ethanol extract but reduced the adenosine triphosphatase activity. PMID- 11268105 TI - Effect of Mucuna urens (horse eye bean) on the gonads of male guinea-pigs. AB - The effect of Mucuna urens (seeds) on the gonads and sex accessory glands of male guinea-pigs was investigated. Sexually mature guinea-pigs of proven fertility were administered orally with 70 mg/kg and 140 mg/kg body weight of crude extract daily for 8 weeks respectively. Phytochemical screening of the seeds revealed the presence of alkaloids. No death or weight loss were observed during the duration of treatment. No pregnancy occurred in females mated with the treated males. Histological observations at high dose (140 mg/kg) showed complete degeneration of sperm in the testicular tubules. In some tubules, the acrosomal cap of the sperm cells was separated from the nuclei which underwent colour changes. In some tubules only the tails were left in the lumen. The spermatids, primary and secondary spermatocytes showed pycnosis while the morphology of spermatogonia and germinal epithelium appeared normal. Some epididymides were devoid of sperm while others contained degenerated spermatozoa and cell debris. In the prostate gland there was collapse of the villi and reduction of secretion in both the prostate and seminal vesicles. At low doses (70 mg/kg), there was spermatogenic arrest at spermatid stage. These observations have shown that M. urens is a potential male antifertility agent. PMID- 11268106 TI - The antioxidant activity of the essential oils of Artemisia afra, Artemisia abyssinica and Juniperus procera. AB - The essential oils of Artemisia afra Jacq., Artemisia abyssinica Schultz-Bip. and Juniperus procera Hoechst ex Endl. were examined for their potential radical scavenging activities. First a rapid evaluation of antioxidants was made using a TLC screening method. The abilities of the volatile oils to act as nonspecific donors for hydrogen atoms or electrons were checked in the diphenylpicrylhydrazyl assay. Oils from all three species showed positive results and were examined further. The oils of A. afra and J. procera were also effective hydroxyl radical scavenging agents when assessed in the deoxyribose degradation assay, whilst oils from A. abyssinica exhibited a paradoxical effect. In the in vitro assay for non enzymatic lipid peroxidation in liposomes, the oils of A. afra and J. procera also displayed antioxidant potential. It was not possible to measure the effect of A. abyssinica oil in this system because certain components, e.g. alk-2-enals, interfered with the assay. The compounds that contribute to the radical scavenging activities of A. afra and J. procera were identified and then assessed for their effects in the various test systems. Finally, the qualitative and quantitative compositions of the essential oils were studied by GC-MS. PMID- 11268107 TI - Augmentation of natural killer cell activity in vivo against tumour cells by some wild plants from Jordan. AB - Twelve aqueous extracts prepared from Jordanian plants that are currently used in traditional medicine to treat various types of cancer were tested in mice for their augmentation of natural killer cells in vivo in generating cytotoxicity against YAC tumour targets. After 1 week of oral administration of aqueous extracts of fresh Nigella sativum (N.s.) seeds and Allium sativum (A.s.) bulbs significant augmentation of splenic natural killer (NK) cells (62.3% +/- 6.4% and 52.6% +/-5.4% cytotoxicity, respectively), was obtained in comparison with spontaneous activity (24.5% +/- 1.6%) at 200:1 effector:target ratio. Onopordum acanthium (O.a.) stem and leaves and Allium cepa (A.c.) bulbs showed intermediate augmentation (38.6% +/- 3.8% and 30.6% +/- 3.4% cytotoxicity, respectively) while the rest showed insignificant augmentation activity. The fresh aqueous extract of a mixture of the plants with high and intermediate activity showed little insignificant augmentation activity (72.7% +/- 6.7% cytotoxicity) of NK cells compared with that obtained with each plant alone. PMID- 11268108 TI - Chemoprotective potentials of homoisoflavonoids and chalcones of Dracaena cinnabari: modulations of drug-metabolizing enzymes and antioxidant activity. AB - A series of homoisoflavonoids and chalcones, isolated from the endemic tropical plant Dracaena cinnabari Balf. (Agavaceae), were tested for their potential to inhibit cytochrome P4501A (CYP1A) enzymes and Fe-enhanced in vitro peroxidation of microsomal lipids in C57B1/6 mouse liver. The effects of the polyphenolic compounds were compared with those of prototypal flavonoid modulators of CYP1A and the well-known antioxidant, butylated hydroxytoluene. 2-Hydroxychalcone and partly 4,6-dihydroxychalcone were found to be strong inhibitors of CYP1A dependent 7-ethoxyresorufin O-deethylase (EROD) activity in vitro comparable to the effects of quercetin and chrysin. The first screening of flavonoids and chalcones of Dracaena cinnabari for antioxidant activity was done in an in vitro microsomal peroxidation assay. While chalcones were shown to be poor antioxidants, 7,8-methylenedioxy-3(4-hydroxybenzyl) chromane, as one of the tested homoisoflavonoids, exhibited a strong antioxidant activity comparable to that of the strongest flavonol antioxidant, quercetin. PMID- 11268109 TI - Antimicrobial activity of Streblus asper leaf extract. AB - Bactericidal activity was found in the 50% ethanol (v/v) extract of Streblus asper leaves. The extract possessed a selective bactericidal activity towards Streptococcus, especially to Streptococcus mutans which has been shown to be strongly associated with dental caries. The extract had no effect on cultures of Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa, Staphylococcus coagulase positive, Staphylococcus coagulase negative, Serratia marcescens, Klebsiella pneumoniae, Enterobacter, Pseudomonas aeruginosa, Burkholderia pseudomeallei and Candida albicans. The minimum growth inhibitory concentration and the minimum bactericidal concentration of S. asper extract against 10(8) CFU/mL of S. mutans was 2 mg/mL. The active compound is partially polar, partially heat labile, precipitated by 80% ammonium sulphate, and possesses a molecular weight larger than 10 000 Da. The potential for using S. asper extract as a natural product for controlling dental caries is discussed. PMID- 11268110 TI - Immunomodulatory principles of Pelargonium sidoides. AB - Extracts and isolated constituents (coumarins and phenols) of Pelargonium sidoides DC, a plant species used in folk medicine by the Southern African native population, were evaluated for their effects on nonspecific immune functions. Although this herbal medicine is also successfully employed in modern phytotherapy in Europe to cure infectious diseases of the respiratory tract, the scientific basis of its remedial effects is still unclear. Thus, functional bioassays including an in vitro model for intracellular infection with Leishmania parasites, an extracellular Leishmania growth assay, a fibroblast-virus protection assay (IFN activity), a fibroblast-lysis assay (TNF activity) and a biochemical assay for inorganic nitric oxides (iNO) were employed. None of the test samples revealed significant activity against extracellular, promastigote Leishmania donovani, the causative agent of human visceral leishmaniasis. In contrast, apart from the coumarin samples, all the Pelargonium extracts (EC(50) <0.1-3.3 microg/mL), gallic acid (EC(50) 4.4 microg/mL) and its methyl ester (EC(50) 12.5 microg/mL) significantly reduced the intracellular survival of L. donovani amastigotes within murine macrophages. These data indicate that the samples acted indirectly on Leishmania parasites, possibly by activating leishmanicidal macrophage functions. Macrophage activation was confirmed by detection of tumour necrosis factor (TNF-alpha) and inorganic nitric oxides (iNO) in supernatants of sample-treated macrophage cultures. Synthesis of iNO is a well known effector mechanism of macrophages against microorganisms such as Leishmania. Interestingly, blocking iNO-synthase with L-NMMA had no substantial effect on sample-induced intracellular Leishmania kill. From bioassay-guided fractionation, gallic acid and its methyl ester present in large amounts in P. sidoides and in its active extracts, were identified as the prominent immunomodulatory principle for this herbal medicine. The results, when taken together with recent reported antibacterial activity, provide a rational basis for both the traditional and the present utilization of P. sidoides in the claimed conditions. PMID- 11268111 TI - Screening of Brazilian plant extracts for antioxidant activity by the use of DPPH free radical method. AB - Brazilian plant extracts belonging to 16 species of 5 different families (71 extracts) were tested against the stable DPPH (2,2-diphenyl-1-picryl-hydrazyl hydrate) free-radical. The ability to scavenge DPPH radical was measured in these experiments by the discoloration of the solution. Ginkgo biloba and rutin, commonly used as antioxidants for medical purposes, were used as standards. Based on our results, we can say that as a general rule the ethanol extracts of plants belonging to the Verbenaceae family showed lower EC(50) values than the other plant extracts. Among the partitions, the more polar ones (ethyl acetate and n butanol) are those that generally have higher antioxidant activity (AA). PMID- 11268112 TI - Effects of antiinflammatory triterpenes isolated from Leptadenia hastata latex on keratinocyte proliferation. AB - Several triterpenes isolated from Leptadenia hastata latex were tested for their anti-inflammatory activity. Lupeol (1), its acetate (2) and palmitate (3) esters were found to be the main antiinflammatory constituents in the croton oil-induced ear oedema test. Furthermore, lupeol hemisuccinate (4), synthesized from lupeol, exhibited a higher activity than lupeol in the test. These results prove that the triterpenes play a pivotal role in the topical antiinflammatory effect of this latex. In addition, an in vitro model of human skin keratinocytes (epidermal explants) cultured at an air-liquid interface on a de-epidermized human dermis (DED) was used to investigate the effects of lupeol esters on skin repair in vitro. Compared with the control, compounds 2 and 3 improved keratinocyte proliferation at a concentration of 5 microM in the culture medium; however, they remained less active than compounds 1 and 4. In contrast to compound 1, all the lupeol esters (2-4), and particularly compound 4, induced a good differentiation of keratinocytes with a well-formed stratum corneum without parakeratosis. These results substantiate the topical use of Leptadenia hastata latex in traditional medicine and showed that both antiinflammatory activity and the effect on keratinocyte proliferation of compound 1 could be improved by its hemisuccinylation; on the contrary, esterification by acetylation or palmitoylation decreased these activities. PMID- 11268113 TI - Qualitative and quantitative olfactometric evaluation of different concentrations of ethanol peppermint oil solutions. AB - Selection of an adequate placebo is a major problem in clinical trials of Euminz(R) (10% peppermint oil/ethanol) which is used topically for the treatment of tension-type headache. This randomized, controlled, double-blind, cross-over study was performed to investigate whether there are qualitative differences between 10%, 1%, 0.5%, 0.1%, and 0% peppermint oil. Forty-one healthy subjects participated (age range 21-28 years); they rated both intensity, and hedonic tone of the stimuli. Verbal descriptions were combined to multiple response sets (MRS). In addition, the trigeminal impact of odorants was determined. Intensity ratings and MRS "menthol like" and "alcohol/solvent" changed with stimulus concentration. However, intensity had no significant effect on hedonics, trigeminal impact, or the number of descriptive items used. When MRS "menthol like" and "alcohol/solvent" were analysed after being weighted with intensity ratings, changes in relation to stimulus concentration were lost. Thus, the differences between the five concentrations of peppermint oil were--to their largest part--due to changes in stimulus intensity. Considering the large day-to day variability of olfactory sensitivity the present data support the hypothesis that the odour quality of 10% peppermint oil cannot be discriminated from the odour of 0.1%, 0.5%, or 1% peppermint oil when tested on separate days. PMID- 11268114 TI - Antimicrobial activity of honokiol and magnolol isolated from Magnolia officinalis. AB - The antimicrobial activity of honokiol and magnolol, the main constituents of Magnolia officinalis was investigated. The antimicrobial activity was assayed by the agar dilution method using brain heart infusion medium and the minimum inhibitory concentration (MIC) were determined for each compound using a twofold serial dilution assay. The results showed that honokiol and magnolol have a marked antimicrobial effect (MIC = 25 microg/mL) against Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Micrococcus luteus and Bacillus subtilis, but did not show antimicrobial activity (MIC > or = 100 microg/mL) for Shigella flexneii, Staphylococcus epidermidis, Enterobacter aerogenes, Proteus vulgaris, Escherichia coli and Pseudomonas aeruginosa. Our results indicate that honokiol and magnolol, although less potent than tetracycline, show a significant antimicrobial activity for periodontal pathogens. Hence we suggest that honokiol and magnolol might have the potential to be an adjunct in the treatment of periodontitis. PMID- 11268115 TI - The anxiolytic effect of Sho-ju-sen, a Japanese herbal medicine, assessed by an elevated plus-maze test in mice. AB - Sho-ju-sen (SK), a Japanese herbal medicine with a nourishing tonic action, is composed of a water extract of Kumazasa leaves (Sasa kurinensis Makino et Sibata) (SS), and ethanol extracts of Japanese red pine needles (Pinus densiflora Sieb. et Zucc) (PN) and Ginseng roots (Panax ginseng C. A. Meyer) (PX) in the ratio 8:1:1. In this study, an elevated plus-maze test in mice was carried out to assess whether SK had an anxiolytic effect. No significant change was observed in either the plus-maze or activity test following a single administration of SK (10 and 20 mL/kg p.o.). However, mice allowed a free intake of SK (10% solution) for 5 days and longer showed a significant prolongation of the time spent in the open arms (an anxiolytic effect), as long as that caused by the benzodiazepine anxiolytic diazepam (1 mg/kg p.o.). SK (1%, 3% and 30% solutions for 7 days) tended to develop the anxiolytic effect. Of the constituents of SK, SS (8% solution), but not PN (1% solution) or PX (1% solution), resulted in the anxiolytic effect. Except for a slight acceleration in the motor activity by PN (1% solution), no significant change in the motor activity was produced by any treatment with SK, SS or PX. The combined treatment of SK (10% solution) or SS (8% solution) with 1 mg/kg diazepam enhanced the anxiolytic effect. Flumazenil (0.1 mg/kg s.c.), a benzodiazepine receptor antagonist, alone did not change the time spent in the open arms. However, it completely reversed the anxiolytic effect of SK, SS and diazepam. The present results suggest that: (1) long-term treatment with SK develops an anxiolytic effect, (2) SS is the main constituent for the anxiolytic effect of SK, and (3) benzodiazepine receptors are involved in the anxiolytic effect of SK and SS. PMID- 11268116 TI - In vitro leishmanicidal activity of naturally occurring chalcones. AB - A variety of chalcones have been shown to exhibit activity against Leishmania parasites. In contrast to synthetic or semisynthetic chalcones, only a few plant derived compounds have been investigated. To provide a scientific rational for the antiprotozoal potency of plants used in ethnomedicine and containing chalcones, and in the search for new antiprotozoal drugs, we have carried out a primary screening for in vitro leishmanicidal activity of 20 chalcones isolated from plants. The compounds were tested against extracellular promastigotes of Leishmania donovani, L. infantum, L. enrietii and L. major, and against intracellular amastigote L. donovani residing within murine macrophages. Against the extracellular Leishmania (L. donovani), most compounds were active with EC(50) values between 0.07 and 2.01 microg/mL. Some of these chalcones, 2',4' dihydroxy-4-methoxychalcone, 2'-hydroxy-3,4-dimethoxychalcone and 2-hydroxy-4,4' dimethoxychalcone also significantly inhibited the intracellular survival of L. donovani parasites with EC(50) values between 0.39 and 0.41 microg/mL. When tested against murine bone marrow-derived macrophages as a mammalian host cell control, all compounds with antileishmanial activities also proved to be cytotoxic to varying extents (EC(50) 0.19-2.06 microg/mL). Correlations between molecular structures and antileishmanial activity are discussed in detail. Specific compounds are illustrated with emphasis on their mode of action and potential for the development of selective antiprotozoal agents. PMID- 11268117 TI - Central nervous system stimulatory action from the root extract of Plumbago zeylanica in rats. AB - The effects of a 50% ethanol extract of the root of Plumbago zeylanica (P. zeylanica) were investigated on locomotor behaviour and central dopaminergic activity in rats. The effects on the ambulatory behaviour were assessed along with the levels of dopamine (DA) and its metabolite homovanillic acid (HVA) in the striatum after a single oral dose (100, 200 and 300 mg/kg body weight) of the extract. The extract significantly increased the spontaneous motility in animals. The ambulatory and rotatory behaviour in the treated groups were higher than in the control group (p < 0.05). There were marked differences in the ambulatory behaviour between 100 and 300 mg/kg, indicating that the responses were stimulatory and dose-dependent. The stereotypic behaviour which is characteristic of a dopamine agonist showed biphasic effects. However, there was no significant difference between the groups (p > 0.05). The results showed that the extract of the root of P. zeylanica specifically enhanced the spontaneous ambulatory activity without inducing stereotypic behaviour. The neurochemical estimations revealed elevated levels of DA and HVA in striatum compared with the control rats (p < 0.01). The levels were higher for the 100 mg/kg treated group than the other groups. The levels declined by increasing the dosage of the extract to 200 mg/kg and 300 mg/kg, however, these levels remained higher than the control group. The relationship between motor activity and levels of dopamine are not parallel. These behavioural and biochemical results indicated stimulatory properties of the extract of the root of P. zeylanica, which may be mediated by dopaminergic mechanisms in the rat brain. PMID- 11268118 TI - Effect of Aloe vera leaves on blood glucose level in type I and type II diabetic rat models. AB - Aloe vera (L.) Burm. fil. (= A. barbadensis Miller) (Liliaceae) is native to North Africa and also cultivated in Turkey. Aloes have long been used all over the world for their various medicinal properties. In the past 15 years, there have been controversial reports on the hypoglycaemic activity of Aloe species, probably due to differences in the parts of the plant used or to the model of diabetes chosen. In this study, separate experiments on three main groups of rats, namely, non-diabetic (ND), type I (IDDM) and type II (NIDDM) diabetic rats were carried out. A. vera leaf pulp and gel extracts were ineffective on lowering the blood sugar level of ND rats. A. vera leaf pulp extract showed hypoglycaemic activity on IDDM and NIDDM rats, the effectiveness being enhanced for type II diabetes in comparison with glibenclamide. On the contrary, A. vera leaf gel extract showed hyperglycaemic activity on NIDDM rats. It may therefore be concluded that the pulps of Aloe vera leaves devoid of the gel could be useful in the treatment of non-insulin dependent diabetes mellitus PMID- 11268119 TI - Comparative pharmacological study of hydroethanol extracts of Passiflora alata and Passiflora edulis leaves. AB - Several species of the genus Passiflora, known in Brazil as "maracuja", have widespread use in folk medicine as sedatives and tranquillizers. The anxiolytic activity of hydroethanol extracts of P. alata and P. edulis leaves was evaluated using the elevated plus-maze test. The extracts presented anxiolytic activity in dosages around 50, 100 and 150 mg/kg. PMID- 11268120 TI - Antibacterial activity of Ailanthus excelsa (Roxb). AB - The antibacterial activity of different fractions of a methanol extract obtained from the dried stem bark of Ailanthus excelsa (Roxb) was studied using different bacterial strains. The ethyl acetate fraction inhibited the growth of all test bacteria. The MIC of the EA fraction was found to be 6 mg/disc. PMID- 11268121 TI - Inhibitory effects of sanguinarine on monoamine oxidase activity in mouse brain. AB - The effects of benzophenanthridine alkaloids, such as sanguinarine and chelidonine, on monoamine -oxidase (MAO) activity in mouse brain were investigated. Sanguinarine showed an inhibitory effect on MAO activity in a concentration dependent manner (53.4% inhibition at 25 microM). However, chelidonine did not inhibit MAO activity. The IC(50) value of sanguinarine was 24.5 microM. Sanguinarine inhibited non-competitively MAO activity using kynuramine as a substrate. The K(i) value for sanguinarine was 22.1 microM. These results suggest that sanguinarine partially contributes to the regulation of catecholamine content. PMID- 11268122 TI - Insect repellent activity of diterpenoid alkaloids. AB - Diterpenoid and norditerpenoid alkaloids were tested against Tribolium casteneum (Herbst.) in order to assess their repellent activity. Of 29 tested alkaloids, 21 compounds showed promising insect repellent activity, while eight of them were not found to be active. The alkaloids were obtained from Delphinium, Consolida and Aconitum species. The highest activity was found in hetisine, a diterpene alkaloid (59.37%) and the lowest activity in another diterpene alkaloid venulol (31.25%). PMID- 11268123 TI - Garcinia extract inhibits lipid droplet accumulation without affecting adipose conversion in 3T3-L1 cells. AB - Garcinia extract was isolated from the fruit of the Garcinia cambogia and was used as a potential antiobesity agent. In week 3 of culture with insulin, the fat cells exhibited more numerous and larger intracytoplasmic lipid droplets (i.e. 30 40 microm(2)). When Garcinia extract and insulin were added simultaneously, the accumulation of lipid droplets was inhibited and the peak droplet area shifted to become smaller (10-20 microm(2)). The activities of glycerophosphate dehydrogenase, a marker of adipose differentiation, were not significantly inhibited by the Garcinia extract. These findings suggest that the Garcinia extract inhibits lipid droplet accumulation in fat cells without affecting adipose conversion. PMID- 11268124 TI - Antiinflammatory and antioxidant property of saponins of tea [Camellia sinensis (L) O. Kuntze] root extract. AB - Two groups of saponins, TS-1 and TS-2, isolated from tea root extract (TRE) were tested for antiinflammatory and in vitro antioxidant activity. Both TS-1 and TS-2 inhibited carrageenan-induced paw oedema in rats. The antioxidant activity of these compounds was evaluated using the xanthine-xanthine oxidase system. The study indicated that the previously observed antitumour activity of TRE might be mediated through scavenging of free radicals by saponins and their antiinflammatory activity. PMID- 11268125 TI - Effects of Shilajit on the development of tolerance to morphine in mice. AB - Effects of concomitant administration of Processed Shilajit (PS, 0.1 and 1 mg/kg, i.p.), in Swiss mice were evaluated on the development of tolerance to morphine induced analgesia in the hot plate test. Chronic administration of morphine (10 mg/kg, i.p., b.i.d.) to mice over a duration of 10 days resulted in the development of tolerance to the analgesic effect of morphine. Concomitant administration of PS with morphine, from day 6 to day 10, resulted in a significant inhibition of the development of tolerance to morphine (10 mg/kg, i.p.) induced analgesia. Processed Shilajit per se, in the doses used, did not elicit any significant analgesia in mice; nor did the chronic concomitant administration of Processed Shilajit alter the morphine-induced analgesia. These findings with Processed Shilajit indicate its potential as a prospective modifier of analgesic tolerance to morphine. PMID- 11268128 TI - Structural elucidation of disinfection by-products in treated drinking water. AB - Two unusual disinfection by-products have been detected periodically in the gas chromatography/mass spectrometry (GC/MS) characterization analyses of semi volatile organics in treated drinking water. The electron ionization (EI) mass spectra contained mono-chlorinated and mono-brominated ions at m/z 107/109 and 151/153, respectively. Library searching techniques suggested mono-halogenated butanol structures but no matches could be found. Positive ion chemical ionization (CI) spectra contained mono-chlorinated and mono-brominated ions at m/z 105/107 and 149/151, respectively. No [M + H]+ ions were initially observed. Accurate mass measurements confirmed the empirical formulae for the significant ions in the EI spectra and the mono-halogenated butanol structures. Further CI experiments with other reagent gases and instruments revealed possible molecular weights of 139 and 183 Da, respectively. Tandem mass spectrometry (MS/MS) experiments in EI and CI were used to elucidate the fragmentation schemes. The two compounds have been tentatively identified as 1-aminoxy-1-chlorobutan-2-ol and 1-aminoxy-1-bromobutan-2-ol, respectively. PMID- 11268129 TI - Electron ionisation induced fragmentation of fused norbornene analogues containing SiMe2 or GeMe2 and oxygen bridges. Migration of SiMe2 and GeMe2 groups. AB - The main electron ionisation induced fragmentation processes of fused norbornene analogues containing SiMe2 or GeMe2 and oxygen bridges, as well as their dependence on substitution, were investigated using mass (MS) and tandem mass (MS/MS) spectrometric analysis. Formation of the rearrangement ions of m/z 176 in the mass spectra of fused norbornene analogues containing SiMe2 and m/z 222 in the mass spectrum of norbornene analogues containing GeMe2 provides firm evidence for the migration of a SiMe2 and GeMe2 bridge, respectively. PMID- 11268130 TI - A highly automated 96-well solid phase extraction and liquid chromatography/tandem mass spectrometry method for the determination of fentanyl in human plasma. AB - A high-throughput bioanalytical method based on automated sample transfer, automated solid phase extraction, and fast liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis, has been developed for the determination of the analgesic fentanyl in human plasma. Samples were transferred into 96-well plates using an automated sample handling system. Automated solid phase extraction (SPE) was carried out using a 96-channel programmable liquid-handling workstation using a mixed-mode sorbent. The extracted samples were then dried down, reconstituted and injected onto a silica column using an aqueous/organic mobile phase with tandem mass spectrometric detection. The method has been validated over the concentration range 0.05-100 ng/mL fentanyl in human plasma, based on a 0.25-mL sample size. The assay is sensitive, specific and robust. More than 2000 samples have been analyzed using this method. The automation of the sample preparation steps not only increased the analysis throughput, but also facilitated the transfer of the method between different bioanalytical laboratories of the same organization. PMID- 11268131 TI - Electrospray ionisation mass spectrometric study of degradation products of quercetin, quercetin-3-glucoside and quercetin-3-rhamnoglucoside, produced by in vitro fermentation with human faecal flora. AB - Eight phenolic compounds, obtained by in vitro fermentation of quercetin, quercetin-3-glucoside and quercetin-3-rhamnoglucoside were analysed by electrospray ionisation mass spectrometry (ESI-MS). Low-energy collision-induced dissociation tandem mass spectrometry (CID-MS/MS) was performed on the [M - H]- precursor ions to obtain specific fragmentation. Typical fragmentation of the phenolic acids was loss of 44 (CO2) and 18 (H2O) u. Production of m/z 108 by loss of neutral radicals, e.g. HCO2, CH3 or HCO, was also favoured. Structures of the compounds, numbered 1-8, were suggested based on the fragmentation patterns. PMID- 11268132 TI - Metal-assisted esterification: glutaric acid-iron(II) complexes in the gas phase. AB - Transition metal ions are routinely used to assist organic reactions; however, direct detection of the intermediates in such reactions is uncommon. Here, we demonstrate a transition metal ion-assisted reaction between glutaric acid (L) and methanol, using electrospray ionization mass spectrometry (ESI-MS). Esterification of glutaric acid does not occur in aqueous methanol solution under ESI conditions, but the FeII-bound acid cluster, [FeII L2 - H]+, adds methanol and dehydrates to give rise to an abundant product ion with a 14 Da increased mass. The occurrence of methyl esterification is supported by collision-induced dissociation and isotopic labeling data, which indicate that the sequence by which the product ion is generated is loss of water, followed by the addition of methanol. Electrospray ionization conditions, specifically the tube lens offset voltage, strongly affect the reaction efficiency, presumably through control of the dehydration process. Other transition metal ions, such as NiII, ZnII, CoII and CuII, also show distinctive metal-assisted reactions. PMID- 11268133 TI - The solvent shells of cluster ions produced by direct electric field extraction from glycerol/water mixtures. AB - Electric field extraction of gaseous negative ions directly from water/glycerol solutions by use of a track membrane technique was investigated. The distributions of numbers of solvent molecules in the extracted cluster ions for different compounds were obtained. It is shown that the extraction mechanism is a direct field-stimulated evaporation of cluster ions from liquid, with a subsequent loss of several solvent molecules in the vacuum. For relatively simple ions a good correspondence of results was obtained with a continuous medium model. It was found that the number of solvent molecules in a cluster shell, for more complicated ions such as amino acids, is significantly greater than that for halide ions or ions of simple organic acids. An increase in the number of solvent molecules in the case of amino acid negative ions is rationalized in terms of the existence of several charged groups, each of which gives an additional contribution to the cluster shell. PMID- 11268134 TI - Gas chromatography/combustion/isotope-ratio-monitoring mass spectrometric analysis of methylboronic derivatives of monosaccharides: a new method for determining natural 13C abundances of carbohydrates. AB - Monosaccharides were derivatized using methylboronic acid and N,O bis(trimethylsilyl)trifluoroacetamide (BSTFA), and the delta13C values of these derivatives measured by gas chromatography/combustion/isotope-ratio-monitoring mass spectrometry to determine the original 13C-content of the monosaccharides. Comparison with the measured off-line delta13 values of the monosaccharides shows that no fractionation in 13C takes place during derivatization. The methylboronic derivatization method has proven to be a new method for natural abundance isotopic analysis of intact monosaccharides (arabinose, xylose, fucose, fructose and glucose). The method is rapid, does not involve isotopic fractionation during derivatization, and gives more precise delta13C values than other methods reported. The method was successfully applied to determine the delta13C value of glucose of the freshwater alga Scenedesmus communis. PMID- 11268135 TI - Referencing strategies and techniques in stable isotope ratio analysis. AB - Stable isotope ratios are reported in the literature in terms of a deviation from an international standard (delta-values). The referencing procedures, however, differ from instrument to instrument and are not consistent between measurement facilities. This paper reviews an attempt to unify the strategy for referencing isotopic measurements. In particular, emphasis is given to the importance of identical treatment of sample and reference material ('IT principle'), which should guide all isotope ratio determinations and evaluations. The implementation of the principle in our laboratory, the monitoring of our measurement quality, the status of the international scales and reference materials and necessary correction procedures are discussed. PMID- 11268136 TI - Shared decision-making in question. AB - Over recent years, communication within the physician-patient relationship has been profoundly changing. New modes of conveying diagnostic and therapeutic information influence the way in which decisions regarding treatment are made. We propose a critical review of the various theoretical models as presented in the literature, from the paternalistic to the shared decision model, in order to reveal conceptual ambiguities and their related methodological problems. This analysis leads to a project for clarifying these problems through a research protocol based on shared decision-making. PMID- 11268137 TI - Awareness and barriers to use of cancer support and information resources by HMO patients with breast, prostate, or colon cancer: patient and provider perspectives. AB - This study assessed patient awareness and use--as well as obstacles to use--of HMO- and community-based psychosocial support services designed for cancer patients. Participants were a randomly selected group of patients from a large Northwest HMO, with breast (N=145), prostate (N=151), or colon cancer (N=72), and their oncology and urology providers (N=29). Patient awareness was highest for HMO-based services (68-90%) and lower for community- (33%) or Internet-based (10 14%) services, and use rates were low across all services (range 2-8%). Providers reported referring 70% of their patients to HMO cancer support services, but their estimates of actual patient use of these services (40%) were inflated. Providers reported few barriers to referring patients to support services. The most commonly reported patient barriers to using such services were already having adequate support, lack of awareness of the service, and lack of provider referral. Results of regression analyses suggest that education, physician referral, social support, and spirituality may be important influences on use of cancer support services. This study takes a first step toward understanding patient use of existing cancer support services and suggests ways to increase participation in these services. PMID- 11268138 TI - Art therapy with adult bone marrow transplant patients in isolation: a pilot study. AB - Psycho-social interventions for cancer patients in isolation for bone marrow transplant (BMT) have been advocated in the recent literature. It is not clear what type of interventions would be most appropriate. This study was conducted at Memorial Sloan-Kettering Cancer Center (MSKCC), with three aims. (1) To test the feasibility of introducing art therapy as a supportive intervention for adult BMT patients in isolation. Nine patients were seen in art therapy sessions twice a week while in isolation, and were helped to develop free personal images. The three art therapists used the same art therapy program as a model. (2) To assess how patients would use the program. Forty-two images were made by the nine patients during the art therapy sessions. A thematic analysis of the images showed that the patients used art therapy effectively in three ways: (a) to strengthen their positive feelings, (b) to alleviate their distress, and (c) to clarify their existential/spiritual issues. (3) The third aim was to identify which patients would most benefit from art therapy. Our results suggest that the non-verbal metaphorical modality of art therapy may be especially beneficial for patients who need to deal with emotional conflicts, and with feelings about life and death, in a safe setting. PMID- 11268139 TI - 'Dividing the desolation': clients views on the benefits of a cancer counselling service. AB - This paper describes clients' accounts of the benefits they derived from a short course of cancer counselling provided within a humanist framework. Three hundred and two clients who had attended at least one session of a short course of cancer counselling received an evaluation form, which incorporated both fixed-choice and open-ended questions. One hundred and forty two (47%) clients returned evaluation forms; those who had attended more sessions were significantly more likely to do so. Quantitative data were analysed using SPSS (Statistical Package for the Social Sciences) for Windows and qualitative data using a thematic approach. Almost all clients indicated that they felt they had benefited from counselling. Analysis of the open-ended questions identified nine main benefits of counselling and four key avenues or processes through which clients derived these benefits. Overall, counselling was seen as helping them to work through powerful thoughts and feelings and so to come to terms with cancer and to regain a sense of control in their lives. The benefits of a short course of counselling which clients identified reflect the aims of humanistic counselling which are not well captured by psychiatric assessments or most standard research instruments. In evaluating cancer counselling services, assessments which include these client-defined outcomes may provide a more sensitive way of gauging the value of counselling to a non-clinic population. PMID- 11268140 TI - Low-income women with early-stage breast cancer: physician and patient decision making styles. AB - BACKGROUND: Poor women have low rates of breast conservation therapy not explained by differences in insurance status or treatment preferences. The purpose of this study was to explore how low-income women make decisions about breast cancer treatment. METHODS: Twenty-five women diagnosed with early-stage breast cancer through the Nebraska Every Woman Matters program were interviewed about their experiences selecting treatment options. These interviews were transcribed and then analysed using established qualitative techniques. RESULTS: More than half of the women (n=16) described playing a passive role in decision making. Choice was determined by medical factors or not offered by their physicians. Intense emotional distress affected some women's ability to compare options. The women who did engage in a rational decision-making process (n=9) based their choices on concerns about body image and fear of recurrence. CONCLUSIONS: When presented with a choice, and when able to objectively weigh treatment options, low-income women base their treatment decisions on the same issues as those of higher income. Whether differences in income strata alter the doctor-patient power dynamic in favor of physician control over decision making, or whether low-income women are less prepared to engage in a rational deliberative process warrants further study. PMID- 11268141 TI - Married couples' perspectives on prostate cancer diagnosis and treatment decision making. AB - Prostate cancer is a disease of men. But for married men, it is commonly and quite reasonably expected that their wives will be significantly involved in dealing with the disease and its treatment. Yet, there are few data on how wives perceive the diagnosis and their role in responding to it. This study explores men's and their wives' perceptions of the prostate cancer diagnosis and their role in treatment decision-making. We conducted separate focus group interviews with seven married men with metastatic prostate cancer and their wives. The accounts of receiving the diagnosis and deciding on treatment that were told by the men differed in significant ways from the stories told in the wives' groups. We found that many men do not share their prostate-related health problems with their wives and some men choose their treatment without much spousal consideration. Our findings suggest that physicians may be in a position to influence how men and their wives interact in reaching a treatment decision. PMID- 11268142 TI - Assessment of depression among cancer patients: the role of pain, cancer type and treatment. AB - One hundred consecutive cancer patients were assessed using two structured methods for assessing major depressive disorder-Structured Clinical Interview for DSM III-R (SCID) and Endicott criteria-and using a depression rating scale Hamilton Depression Rating Scale (HAMD). Forty-nine percent of patients were depressed using SCID (DSM III-R criteria), whereas 29% of patients were depressed using Endicott criteria. Twenty-eight percent of patients were depressed using both criteria. Age and sex did not have any influence on the assessment of major depression. Both the structured interview and the rating scale were able to identify suicide ideation. Depressed patients were not shown to have more lifetime depression than non-depressed patients using both structured methods. Patients who were depressed using both assessments of depression had more metastasis and pain than non-depressed patients. PMID- 11268143 TI - Considerations of psychosocial illness phase in cancer survival. AB - Rarely are biomedical, clinical and psychosocial data considered simultaneously as influences on cancer patient outcomes. This study utilized medical record and interview data from 152 adult cancer patients with various tumor types in a model of survival estimation. Predictors included disease stage of the neoplasm (TNM stage), clinical functioning of the patient (Karnofsky performance status), and psychosocial demands of the disease course (psychosocial illness phase). Psychosocial illness phase captures developmental time phases of illness (i.e. 'crisis', 'early chronic', 'late chronic' and 'terminal'), essentially locating patients along the disease course relative to treatment and treatment response. The analysis utilized the Kaplan-Meier (product-limit) method to estimate stratum specific survival functions. Model comparisons employed the differences in the likelihood ratio chi-squares between nested models, and Cox proportional hazard models assisted in explaining the effects of the predictors on survival times. Results indicate that psychosocial illness phase makes an independent contribution to survival time estimation (p<0.05) when all three dimensions are considered simultaneously. PMID- 11268144 TI - Attributions of cause and recurrence in long-term breast cancer survivors. AB - Women are bombarded with information about the purported causes and the prevention of breast cancer. This survey sought to determine to what women survivors of breast cancer attributed the cause and lack of recurrence of their breast cancer, and whether these views were associated with specific health behaviors. METHODS: Women who had survived breast cancer without recurrence for at least 2 years were surveyed by mail about their views on the cause and lack of recurrence of their breast cancer. They were also asked to estimate their personal risk of cancer recurrence, report on their health behaviors, describe what advice they would give to women newly diagnosed with breast cancer, and what they would change if they had to relive their breast cancer experience. RESULTS: 378 (75.6%) women breast cancer survivors responded who had been recurrence free for a mean of 8.6+/-11.8 years. Women (n=322) who responded to the question about the cause of breast cancer attributed it to stress (42.2%), genetics (26.7%), environment (25.5%), hormones (23.9%), don't know (16.5%), diet (15.5%), and breast trauma (2.8%). Women (n=330) who responded to the question about what prevented cancer recurrence attributed it to positive attitude (60.0%), diet (50.0%), healthy lifestyle (40.3%), exercise (39.4%), stress reduction (27.9%), prayer (26.4%), complementary therapies (11.2%), don't know (5.1 %), luck (3.9%), and tamoxifen (3.9%). Most women felt their recurrence risk was average (44.8%), or below average (35.8%). Some attributions of breast cancer cause or lack of recurrence were associated with specific health behaviors. The majority of women survivors would advise other women with breast cancer to be positive, and if they had to relive their cancer experience they would take more control of their treatment. DISCUSSION: Despite lack of evidence substantiating stress as a cause of breast cancer, many breast cancer survivors believed stress caused their cancer. An even higher percentage of survivors believed their positive attitude had prevented breast cancer recurrence and they would advise other women with breast cancer accordingly. Attribution beliefs clearly affected survivors' health behaviors. CONCLUSION: Healthcare providers should consider the personal beliefs of patients about cancer cause and recurrence, which may be at variance with scientific evidence. This may assist in framing the management of patients in personally meaningful ways, which may increase health behaviors, adherence, satisfaction and quality of life. Whether it will increase survival remains unknown. PMID- 11268145 TI - Planting seeds of knowledge about inflammatory bowel disease: half a century of science, prescience, and prophecy in the pages of Mount Sinai s Journal. AB - This is a review of those clinical observations, innovative concepts, and predictions concerning inflammatory bowel disease that were first published in The Journal of the Mount Sinai Hospital, renamed in 1970 The Mount Sinai Journal of Medicine. The review was based on a hand search of every volume of The Journal from its inception in 1934 to the present day. PMID- 11268146 TI - Hepatology at Mount Sinai: the present and the future. AB - The Division of Liver Disease at Mount Sinai, now into its fifth decade, has evolved through two remarkable periods in its development and is on the cusp of a third exciting era. The first, extending from the division's creation in 1957 to the retirement in 1988 of its first division chief, Fenton Schaffner, was characterized by brilliant clinical and pathophysiologic insights derived from the unique collaboration of Schaffner, a master clinician, with Hans Popper, a world renowned pathologist widely acknowledged as the father of the modern discipline of hepatology. The second, extending from the appointment in 1988 of Paul D. Berk as Schaffner's successor to the present day, has witnessed enormous growth in the clinical and scientific activities of the division, together with the emergence of a world-class liver transplant program at Mount Sinai. During this recent period, an extensive program of formal clinical research was established; the basic research program then expanded into the areas of hepatic transport, molecular virology, and the cellular and molecular pathogenesis of hepatic fibrosis; and both the clinical and research productivity of the division increased dramatically. A major undertaking, now in its second year, has been the creation of the Center for the Study of Primary Biliary Cirrhosis; Mount Sinai has contributed important advances toward the understanding of this disease. Funding for the Center, from the Artzt Family Foundation Trust, supports a series of interrelated basic studies on the immunology and pathobiology of the disease, as well as creation of a unique clinical database, a serum and tissue bank, and a program of clinical studies. This integration of basic and clinical research in pursuit of new methods of diagnosis and treatment serves as a model for the division's continued leadership role. PMID- 11268147 TI - The Mount Sinai division of gastroenterology at the beginning of the 21st century. AB - The Mount Sinai Division of Gastroenterology has an international reputation for outstanding contributions to the study of digestive diseases, especially inflammatory bowel disease. A discussion of the current structure of the gastroenterology (GI) fellowship training program is provided, along with an overview of the GI Division at the turn of the 21st century. PMID- 11268148 TI - Pediatric gastroenterology at the Mount Sinai hospital. AB - The history of pediatric gastroenterology at Mount Sinai begins in 1960. Early publications by Drs. Korelitz and Gribetz on the management of inflammatory bowel disease in children served as the preface to forty years of progress in this important area. The history of pediatric gastroenterology includes important work by many individuals, including Horace Hodes, Lotte Strauss and Frederick Kopel. Early observations on the nature of inflammatory bowel disease (IBD), and its course, preceded work on nutritional therapies for IBD, mechanisms of gene nutrient interactions, regulation of gene transcription, and molecular processes involved in bile transport in the liver and small intestine. Over the last twenty years, the division has grown in size and reputation. Today there are fourteen full-time faculty - 9 M.D.'s and 5 Ph.D.'s - who work in three funded research laboratories. There are also five advanced practice nurses (including three nurse practitioners), two social workers and two nutritionists, as well as several administrators and assistants. In addition to being recognized as a premier center for the treatment of children with general pediatric gastroenterological problems, especially inflammatory bowel disease, the division is also known as one of the nation's largest pediatric liver and liver transplant centers, and it is rapidly becoming one of the largest pediatric short gut syndrome and small bowel transplant centers. PMID- 11268149 TI - The evolution of gastrointestinal endoscopy at the Mount Sinai Hospital. AB - Gastrointestinal endoscopy came to The Mount Sinai Hospital in the 1950s, along with the Wolf-Schindler gastroscope. In 1961, it was supplemented by the Eder Hufford semi-flexible esophagoscope and later by the Olympus gastrocamera and then the Hirschowitz fiberoptic instruments from ACMI and Olympus. A formal training program was started by Jerome Waye in 1966 for flexible gastroscopy and esophagoscopy. In 1969, endoscopic retrograde cholangiopancreatography (ERCP) was introduced. Colonoscopy was at first performed under x-ray control, and subsequently replaced by the nonfluoroscopic method of colonoscopic topography, which was developed by Dr. Waye. A full-time nurse who was in charge of the endoscopy unit founded the Society for Gastrointestinal Nurses and Assistants while working at The Mount Sinai Hospital. PMID- 11268150 TI - Ileostomy and colostomy support groups. AB - The first organized ostomy support group in the world was formed at The Mount Sinai Hospital, in 1950, through the efforts of a surgeon and the patients themselves. Later, similar groups were set up in other locations and some even took the name of Mount Sinai's first such society (QT). These groups had two major functions: Psychological: reassurance and understanding from other ostomates before and after the operation; advice on how to deal with oneself and others. Educational: instruction on the details of stoma management; information for surgeons on the proper location and other details of fashioning a stoma; information to the public on the existence and needs of ostomates. In order to extend services to more people and with more individual attention, a special clinic was established at The Mount Sinai Hospital devoted entirely to patients with ileostomy and colostomy. It too was the first of its kind anywhere. Years later, two surgeons arranged an all-day convention of the ostomy groups in the greater New York City, New Jersey, and Connecticut areas, at the New York Academy of Medicine. At the convention, the idea of a national society was conceived. In the following years - in Detroit, then Cleveland, and finally Los Angeles - the United Ostomy Association was developed, structured, and incorporated. The educational uses of the ostomy groups were later supplemented and partly replaced by enterostomal therapists who originally were trained and practiced in Cleveland. Now the therapists, the societies, and the ostomates themselves are available to patients with various types of stomas, whether recent or longstanding, before and after surgery. PMID- 11268151 TI - To make a difference: the founding of the Crohn's and Colitis Foundation of America. AB - In 1965, with the help of Dr. Henry D. Janowitz, Irwin M. Rosenthal established the Foundation for Research in Ileitis, Inc., now known as the Crohn's and Colitis Foundation of America, Inc. He was joined shortly thereafter by William D. Modell. At that time, the entire annual NIH budget for research on inflammatory bowel disease was only $25,000. Successful fund raising made it possible to recruit a research fellow at The Mount Sinai Hospital in the Division of Gastroenterology to study ileitis. Thereafter, the efforts of the foundation expanded nationwide. It supported a nationally coordinated research program and sponsored education for physicians, patients and the public. In addition, it established support groups to help patients and their families cope with Crohn's disease and ulcerative colitis. With the energy and philanthropy of the foundation's many lay leaders, tens of millions of dollars have been raised for research and education in inflammatory bowel disease. PMID- 11268152 TI - Impairments: always linked to meaningful disability? PMID- 11268153 TI - Relationship between duration of therapy services in a comprehensive rehabilitation program and mobility at discharge in patients with orthopedic problems. AB - BACKGROUND AND PURPOSE: The purpose of this study was to examine the relationship between duration of physical therapy and occupational therapy and mobility at the time of discharge from a comprehensive rehabilitation program in a group of patients with orthopedic diagnoses. SUBJECTS: Subjects were 116 consecutive patients with orthopedic diagnoses (mean age=72.6 years, SD=12.0, range=21-99) who were admitted to a comprehensive inpatient rehabilitation program. METHODS: This retrospective cohort study utilized the Uniform Data Set, social service records, and quality assurance records to provide demographic and medical information. The Functional Independence Measure (FIM) provided information regarding mobility at admission and discharge. The duration of physical therapy and occupational therapy was measured in hours. RESULTS: Subjects received an average of 40.8 hours of therapy and showed an average change in FIM mobility subscale scores of 24.5. Multiple linear regression was used to demonstrate that duration of therapy was a predictor of FIM score at the time of discharge (partial correlation=.069) after controlling for length of stay, number of diagnoses, FIM cognitive subscale score at admission, and FIM mobility subscale score at admission. Duration of therapy accounted for 6.9% of the variance in the model. CONCLUSION AND DISCUSSION: This study indicates that the amount of physical therapy and occupational therapy that patients with orthopedic diagnoses receive during enrollment in an inpatient comprehensive rehabilitation program is related to the FIM mobility subscale score at the time of discharge. The authors suggest that increasing the hours of therapeutic intervention that a patient receives in inpatient rehabilitation could improve functional outcomes at discharge. PMID- 11268154 TI - Student performance and perceptions of a lecture-based course compared with the same course utilizing group discussion. AB - BACKGROUND AND PURPOSE: Self-directed learning is believed to be an important aspect of the reflective clinical practitioner. This study was a comparison of student learning and student perceptions of course and instructor effectiveness, course difficulty, and amount learned between the active learning and lecture sections of a course. SUBJECTS: Participants in this study were 170 physical therapist students in 3 sections of a physiology course in the first year of their professional program. METHODS: Course grades and the results of teacher course evaluations were compared between a lecture section and an active learning section. The students in the original active learning section were reassessed 1 year later to determine their perceptions of the course. The differences were analyzed using Kruskal-Wallis and Mann-Whitney U tests. RESULTS: Course grades were higher in both active learning sections than in the lecture section. However, the students in both active learning sections perceived that they had learned less than students in the lecture section. Students' perceptions of course and instructor effectiveness were lower in the active learning sections than in the lecture section. There were no differences between the lecture and active learning sections on the students' perceptions of course difficulty. CONCLUSION AND DISCUSSION: Although they did better in the active learning environment, physical therapist students in a basic sciences course (physiology) in the first year of their professional program perceived that they had learned less in active learning courses. They also had lower perceptions of course and instructor quality. PMID- 11268155 TI - Relationships among selected measures of impairment, functional limitation, and disability in patients with cervical spine disorders. AB - BACKGROUND AND PURPOSE: Little is known about the relationship among impairments, functional limitations, and disability in people with cervical spine disorders (CSD) despite the fact that these concepts are routinely used in clinical practice. The primary purpose of this study was to investigate the relationships among commonly assessed impairment, functional limitation, and disability measures in patients with CSD. A secondary purpose was to determine the influence of payment source and time since onset of symptoms on these same measures. SUBJECTS: Eighty patients (mean age=45.7 years, SD=15.9, range=20-88) with CSD who were referred for physical therapy participated in the study. METHODS: Data were obtained for 3 measures of impairment, 2 measures of functional limitation, and 3 measures of self-reported disability during the initial visit. RESULTS: All 3 sets of variables (ie, impairment, functional limitation, disability) correlated with each other, with the highest correlation occurring between the impairment measures and the functional limitation measures (r=.82). Other correlations were noted between individual variables. There was no effect of payment type or time since onset of symptoms on the variables. CONCLUSION AND DISCUSSION: Positive correlations were noted among the 3 sets of measures, which supports the assumption that impairments, functional limitations, and disability are related in patients with CSD. PMID- 11268156 TI - Patients in general practice in Denmark referred to physiotherapists: a description of patient characteristics based on general health status, diagnoses, and sociodemographic characteristics. AB - BACKGROUND AND PURPOSE: Both musculoskeletal illness and mental illness characterized by somatic symptoms are common in primary care, and it is hypothesized that many patients with musculoskeletal illness have relatively poor mental health. The purpose of this study was to describe the characteristics of patients in general practice in Denmark who are referred to physiotherapists with signs and symptoms of musculoskeletal illness. SUBJECTS AND METHODS: One hundred ninety-four general practitioners, representing 124 practices, participated in a survey of 2,042 consecutive patients with musculoskeletal illness. RESULTS: The diagnoses were generally poorly defined. Compared with the general population, patients with musculoskeletal illness had markedly poorer physical health and poorer mental health. Patients with poorly defined diagnoses did not differ from patients with well-defined diagnoses in terms of physical health, but they scored lower on the mental health component summary scale of the Danish version of the Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36). CONCLUSION AND DISCUSSION: Of patients referred to physiotherapists by general practitioners in Denmark, the subgroup with poorly defined diagnoses had lower mental health scores than those with well-defined diagnoses, suggesting that a biopsychosocial approach to care may be appropriate for this group of patients. PMID- 11268157 TI - Gait re-education based on the Bobath concept in two patients with hemiplegia following stroke. AB - BACKGROUND AND PURPOSE: This case report describes the use of gait re-education based on the Bobath concept to measure the changes that occurred in the gait of 2 patients with hemiplegia who were undergoing outpatient physical therapy. CASE DESCRIPTION: One patient ("NM"), a 65-year-old woman, was referred for physical therapy 6 weeks following a right cerebrovascular accident. She attended 30 therapy sessions over a 15-week period. The other patient ("SA"), a 71-year-old woman, was referred for physical therapy 7 weeks following a left cerebrovascular accident. She attended 28 therapy sessions over a 19-week period. Clinical indexes of impairment and disability and 3-dimensional gait data were obtained at the start of treatment and at discharge. Therapy was based on the Bobath concept. OUTCOMES: At discharge, NM demonstrated improvements in her hip and knee movements, reduced tone, and improved mobility. At discharge, SA demonstrated improved mobility. During gait, both patients demonstrated more normal movement patterns at the level of the pelvis, the knee, and the ankle in the sagittal plane. SA also demonstrated an improvement in hip extension. DISCUSSION: These cases demonstrate that recovery of more normal movement patterns and functional ability can be achieved following a cardiovascular accident and provide insight into the clinical decision making of experienced practitioners using Bobath's concept. PMID- 11268158 TI - Physical therapy for spinal accessory nerve injury complicated by adhesive capsulitis. AB - BACKGROUND AND PURPOSE: The authors found no literature describing adhesive capsulitis as a consequence of spinal accessory nerve injury and no exercise program or protocol for patients with spinal accessory nerve injury. The purpose of this case report is to describe the management of a patient with adhesive capsulitis and spinal accessory nerve injury following a carotid endarterectomy. CASE DESCRIPTION: The patient was a 67-year-old woman referred for physical therapy following manipulation of the left shoulder and a diagnosis of adhesive capsulitis by her orthopedist. Spinal accessory nerve injury was identified during the initial physical therapy examination, and a program of neuromuscular electrical stimulation was initiated. OUTCOMES: The patient had almost full restoration of the involved muscle function after 5 months of physical therapy. DISCUSSION: This case report illustrates the importance of accurate diagnosis and suggests physical therapy intervention to manage adhesive capsulitis as a consequence of spinal accessory nerve injury. PMID- 11268159 TI - Malignant hyperthermia. PMID- 11268161 TI - A danger to society. PMID- 11268162 TI - Free access to cDNA provides impetus for gene function work. PMID- 11268163 TI - Anti-AIDS drugs available 'at cost'. PMID- 11268164 TI - Network aims to link species data from global collections. PMID- 11268165 TI - London museum puts its animals on public display. PMID- 11268166 TI - Theorists reject challenge to standard model. PMID- 11268168 TI - Mine mooted as underground home for physicists. PMID- 11268167 TI - Congress accused of slighting sound science. PMID- 11268169 TI - Trepidation greets plan for cloning humans. PMID- 11268170 TI - Singapore invests in bioinformatics. PMID- 11268172 TI - Into the mind of a killer. PMID- 11268173 TI - Fellowship fund would help eastern Europe to retain its young talent. PMID- 11268181 TI - Questions of direction. PMID- 11268174 TI - Arabidopsis could shed light on human genome. PMID- 11268175 TI - Climate panel looked at all the evidence. PMID- 11268182 TI - Genesis by definition. PMID- 11268183 TI - Palaeontology. Turning over a new leaf. PMID- 11268184 TI - Brown dwarf is a radio star. PMID- 11268185 TI - Molecular clocks. A vivid loop of light. PMID- 11268186 TI - Sizing up the Sun. PMID- 11268188 TI - Biogeochemistry: Oxygenating the atmosphere. PMID- 11268187 TI - Neurobiology: New memories from new neurons. PMID- 11268190 TI - Animal behaviour: Greensleaves. PMID- 11268191 TI - Cognitive neuroscience: Repression revisited. PMID- 11268192 TI - Superconductivity: Magnetic glue exposed. PMID- 11268193 TI - Structural biology: Protein crystal mimics reality. PMID- 11268195 TI - Frequency-dependent Batesian mimicry. PMID- 11268196 TI - Pattern formation: Instabilities in sand ripples. PMID- 11268197 TI - Palaeoanthropology: Did our ancestors knuckle-walk? PMID- 11268198 TI - Palaeoanthropology: Did our ancestors knuckle-walk? PMID- 11268201 TI - Structure of the bacterial flagellar protofilament and implications for a switch for supercoiling. AB - The bacterial flagellar filament is a helical propeller constructed from 11 protofilaments of a single protein, flagellin. The filament switches between left and right-handed supercoiled forms when bacteria switch their swimming mode between running and tumbling. Supercoiling is produced by two different packing interactions of flagellin called L and R. In switching from L to R, the intersubunit distance ( approximately 52 A) along the protofilament decreases by 0.8 A. Changes in the number of L and R protofilaments govern supercoiling of the filament. Here we report the 2.0 A resolution crystal structure of a Salmonella flagellin fragment of relative molecular mass 41,300. The crystal contains pairs of antiparallel straight protofilaments with the R-type repeat. By simulated extension of the protofilament model, we have identified possible switch regions responsible for the bi-stable mechanical switch that generates the 0.8 A difference in repeat distance. PMID- 11268200 TI - Guard cell abscisic acid signalling and engineering drought hardiness in plants. AB - Guard cells are located in the epidermis of plant leaves, and in pairs surround stomatal pores. These control both the influx of CO2 as a raw material for photosynthesis and water loss from plants through transpiration to the atmosphere. Guard cells have become a highly developed system for dissecting early signal transduction mechanisms in plants. In response to drought, plants synthesize the hormone abscisic acid, which triggers closing of stomata, thus reducing water loss. Recently, central regulators of guard cell abscisic acid signalling have been discovered. The molecular understanding of the guard cell signal transduction network opens possibilities for engineering stomatal responses to control CO2 intake and plant water loss. PMID- 11268202 TI - Discovery of radio emission from the brown dwarf LP944-20. AB - Brown dwarfs are not massive enough to sustain thermonuclear fusion of hydrogen at their centres, but are distinguished from gas-giant planets by their ability to burn deuterium. Brown dwarfs older than approximately 10 Myr are expected to possess short-lived magnetic fields and to emit radio and X-rays only very weakly from their coronae. An X-ray flare was recently detected on the brown dwarf LP944 20, whereas previous searches for optical activity (and one X-ray search) yielded negative results. Here we report the discovery of quiescent and flaring radio emission from LP944-20, with luminosities several orders of magnitude larger than predicted by the empirical relation between the X-ray and radio luminosities that has been found for many types of stars. Interpreting the radio data within the context of synchrotron emission, we show that LP944-20 has an unusually weak magnetic field in comparison to active M-dwarf stars, which might explain the previous null optical and X-ray results, as well as the strength of the radio emissions compared to those at X-ray wavelengths. PMID- 11268203 TI - Strong coupling between local moments and superconducting 'heavy' electrons in UPd2Al3. AB - The electronic structure of heavy-fermion compounds arises from the interaction of nearly localized 4f- or 5f-shell electrons (with atomic magnetic moments) with the free-electron-like itinerant conduction-band electrons. In actinide or rare earth heavy-fermion materials, this interaction yields itinerant electrons having an effective mass about 100 times (or more) the bare electron mass. Moreover, the itinerant electrons in UPd2Al3 are found to be superconducting well below the magnetic ordering temperature of this compound, whereas magnetism generally suppresses superconductivity in conventional metals. Here we report the detection of a dispersive excitation of the ordered f-electron moments, which shows a strong interaction with the heavy superconducting electrons. This 'magnetic exciton' is a localized excitation which moves through the lattice as a result of exchange forces between the magnetic moments. By combining this observation with previous tunnelling measurements on this material, we argue that these magnetic excitons may produce effective interactions between the itinerant electrons, and so be responsible for superconductivity in a manner analogous to the role played by phonons in conventional superconductors. PMID- 11268204 TI - Loss of superconductivity with the addition of Al to MgB2 and a structural transition in Mg1-x AlxB2. AB - The basic magnetic and electronic properties of most binary compounds have been well known for decades. The recent discovery of superconductivity at 39 K in the simple binary ceramic compound magnesium diboride, MgB2, was therefore surprising. Indeed, this material has been known and structurally characterized since the mid 1950s (ref. 2), and is readily available from chemical suppliers (it is commonly used as a starting material for chemical metathesis reactions). Here we show that the addition of electrons to MgB2, through partial substitution of Al for Mg, results in the loss of superconductivity. Associated with the Al substitution is a subtle but distinct structural transition, reflected in the partial collapse of the spacing between boron layers near an Al content of 10 per cent. This indicates that superconducting MgB2 is poised very near a structural instability at slightly higher electron concentrations. PMID- 11268205 TI - Electrical spin injection and accumulation at room temperature in an all-metal mesoscopic spin valve. AB - Finding a means to generate, control and use spin-polarized currents represents an important challenge for spin-based electronics, or 'spintronics'. Spin currents and the associated phenomenon of spin accumulation can be realized by driving a current from a ferromagnetic electrode into a non-magnetic metal or semiconductor. This was first demonstrated over 15 years ago in a spin injection experiment on a single crystal aluminium bar at temperatures below 77 K. Recent experiments have demonstrated successful optical detection of spin injection in semiconductors, using either optical injection by circularly polarized light or electrical injection from a magnetic semiconductor. However, it has not been possible to achieve fully electrical spin injection and detection at room temperature. Here we report room-temperature electrical injection and detection of spin currents and observe spin accumulation in an all-metal lateral mesoscopic spin valve, where ferromagnetic electrodes are used to drive a spin-polarized current into crossed copper strips. We anticipate that larger signals should be obtainable by optimizing the choice of materials and device geometry. PMID- 11268206 TI - Shear-induced molecular precession in a hexatic Langmuir monolayer. AB - Liquid crystalline behaviour is generally limited to a select group of specially designed bulk substances. By contrast, it is a common feature of simple molecular monolayers and other quasi-two-dimensional systems, which often possess a type of in-plane ordering that results from unbinding of dislocations-a 'hexatic' liquid crystalline phase. The flow of monolayers is closely related to molecular transport in biological membranes, affects foam and emulsion stability and is relevant to microfluidics research. For liquid crystalline phases, it is important to understand the coupling of the molecular orientation to the flow. Orientationally ordered (nematic) phases in bulk liquid crystals exhibit 'shear aligning' or 'tumbling' behaviour under shear, and are described quantitatively by Leslie-Ericksen theory. For hexatic monolayers, the effects of flow have been inferred from textures of Langmuir-Blodgett films and directly observed at the macroscopic level. However, there is no accepted model of hexatic flow at the molecular level. Here we report observations of a hexatic Langmuir monolayer that reveal continuous, shear-induced molecular precession, interrupted by occasional jump discontinuities. Although superficially similar to tumbling in a bulk nematic phase, the kinematic details are quite different and provide a possible mechanism for domain coarsening and eventual molecular alignment in monolayers. We explain the precession and jumps within a quantitative framework that involves coupling of molecular orientation to the local molecular hexatic 'lattice', which is continuously deformed by shear. PMID- 11268208 TI - Increases in greenhouse forcing inferred from the outgoing longwave radiation spectra of the Earth in 1970 and 1997. AB - The evolution of the Earth's climate has been extensively studied, and a strong link between increases in surface temperatures and greenhouse gases has been established. But this relationship is complicated by several feedback processes most importantly the hydrological cycle-that are not well understood. Changes in the Earth's greenhouse effect can be detected from variations in the spectrum of outgoing longwave radiation, which is a measure of how the Earth cools to space and carries the imprint of the gases that are responsible for the greenhouse effect. Here we analyse the difference between the spectra of the outgoing longwave radiation of the Earth as measured by orbiting spacecraft in 1970 and 1997. We find differences in the spectra that point to long-term changes in atmospheric CH4, CO2 and O3 as well as CFC-11 and CFC-12. Our results provide direct experimental evidence for a significant increase in the Earth's greenhouse effect that is consistent with concerns over radiative forcing of climate. PMID- 11268207 TI - Evolution of leaf-form in land plants linked to atmospheric CO2 decline in the Late Palaeozoic era. AB - The widespread appearance of megaphyll leaves, with their branched veins and planate form, did not occur until the close of the Devonian period at about 360 Myr ago. This happened about 40 Myr after simple leafless vascular plants first colonized the land in the Late Silurian/Early Devonian, but the reason for the slow emergence of this common feature of present-day plants is presently unresolved. Here we show, in a series of quantitative analyses using fossil leaf characters and biophysical principles, that the delay was causally linked with a 90% drop in atmospheric pCO2 during the Late Palaeozoic era. In contrast to simulations for a typical Early Devonian land plant, possessing few stomata on leafless stems, those for a planate leaf with the same stomatal characteristics indicate that it would have suffered lethal overheating, because of greater interception of solar energy and low transpiration. When planate leaves first appeared in the Late Devonian and subsequently diversified in the Carboniferous period, they possessed substantially higher stomatal densities. This observation is consistent with the effects of the pCO2 on stomatal development and suggests that the evolution of planate leaves could only have occurred after an increase in stomatal density, allowing higher transpiration rates that were sufficient to maintain cool and viable leaf temperatures. PMID- 11268209 TI - Fossil evidence of water lilies (Nymphaeales) in the Early Cretaceous. AB - Phylogenetic analyses have identified the water lilies (Nymphaeales: Cabombaceae and Nymphaeaceae), together with four other small groups of flowering plants (the 'ANITA clades': Amborellaceae, Illiciales, Trimeniaceae, Austrobaileyaceae), as the first diverging lineages from the main branch of the angiosperm phylogenetic tree, but evidence of these groups in the earliest phases of the angiosperm fossil record has remained elusive. Here we report the earliest unequivocal evidence, based on fossil floral structures and associated pollen, of fossil plants related to members of the ANITA clades. This extends the history of the water lilies (Nymphaeales) back to the Early Cretaceous (125-115 million years) and into the oldest fossil assemblages that contain unequivocal angiosperm stamens and carpels. This discovery adds to the growing congruence between results from molecular-based analyses of relationships among angiosperms and the palaeobotanical record. It is also consistent with previous observations that the flowers of early angiosperms were generally very small compared with those of their living relatives. PMID- 11268210 TI - Mesoscale vertical motion and the size structure of phytoplankton in the ocean. AB - Phytoplankton size structure is acknowledged as a fundamental property determining energy flow through 'microbial' or 'herbivore' pathways. The balance between these two pathways determines the ability of the ecosystem to recycle carbon within the upper layer or to export it to the ocean interior. Small cells are usually characteristic of oligotrophic, stratified ocean waters, in which regenerated ammonium is the only available form of inorganic nitrogen and recycling dominates. Large cells seem to characterize phytoplankton in which inputs of nitrate enter the euphotic layer and exported production is higher. But the size structure of phytoplankton may depend more directly on hydrodynamical forces than on the source of available nitrogen. Here we present an empirical model that relates the magnitude of mesoscale vertical motion to the slope of the size-abundance spectrum of phytoplankton in a frontal ecosystem. Our model indicates that the relative proportion of large cells increases with the magnitude of the upward velocity. This suggests that mesoscale vertical motion-a ubiquitous feature of eddies and unstable fronts-controls directly the size structure of phytoplankton in the ocean. PMID- 11268211 TI - Ecological importance of trichromatic vision to primates. AB - Trichromatic colour vision, characterized by three retinal photopigments tuned to peak wavelengths of approximately 430 nm, approximately 535 nm and approximately 562 nm (refs 1, 2), has evolved convergently in catarrhine primates and one genus of New World monkey, the howlers (genus Alouatta). This uniform capacity to discriminate red-green colours, which is not found in other mammals, has been proposed as advantageous for the long-range detection of either ripe fruits or young leaves (which frequently flush red in the tropics) against a background of mature foliage. Here we show that four trichromatic primate species in Kibale Forest, Uganda, eat leaves that are colour discriminated only by red-greenness, a colour axis correlated with high protein levels and low toughness. Despite their divergent digestive systems, these primates have no significant interspecific differences in leaf colour selection. In contrast, eaten fruits were generally discriminated from mature leaves on both red-green and yellow-blue channels and also by their luminance, with a significant difference between chimpanzees and monkeys in fruit colour choice. Our results implicate leaf consumption, a critical food resource when fruit is scarce, as having unique value in maintaining trichromacy in catarrhines. PMID- 11268212 TI - Suppressing unwanted memories by executive control. AB - Freud proposed that unwanted memories can be forgotten by pushing them into the unconscious, a process called repression. The existence of repression has remained controversial for more than a century, in part because of its strong coupling with trauma, and the ethical and practical difficulties of studying such processes in controlled experiments. However, behavioural and neurobiological research on memory and attention shows that people have executive control processes directed at minimizing perceptual distraction, overcoming interference during short and long-term memory tasks and stopping strong habitual responses to stimuli. Here we show that these mechanisms can be recruited to prevent unwanted declarative memories from entering awareness, and that this cognitive act has enduring consequences for the rejected memories. When people encounter cues that remind them of an unwanted memory and they consistently try to prevent awareness of it, the later recall of the rejected memory becomes more difficult. The forgetting increases with the number of times the memory is avoided, resists incentives for accurate recall and is caused by processes that suppress the memory itself. These results show that executive control processes not uniquely tied to trauma may provide a viable model for repression. PMID- 11268213 TI - Masking unveils pre-amodal completion representation in visual search. AB - When one object is partly occluded by another, its occluded parts are perceptually 'filled in', that is, the occluded object appears to continue behind its occluder. This process is known as amodal completion. The completion of a partially occluded object takes about 200 ms, and pre-completion information (that is, information from before amodal completion has occurred) exists in the visual system for that duration. It has been suggested, however, that observers cannot make use of this information, even when it is beneficial to do so: visual search for a target that appears to be partly occluded, for example, is slower than for a target that does not undergo occlusion, despite both targets being physically identical. Here we show that visual search does have access to pre completion representations, but only for a limited time that depends on the size of the occluded region. PMID- 11268214 TI - Neurogenesis in the adult is involved in the formation of trace memories. AB - The vertebrate brain continues to produce new neurons throughout life. In the rat hippocampus, several thousand are produced each day, many of which die within weeks. Associative learning can enhance their survival; however, until now it was unknown whether new neurons are involved in memory formation. Here we show that a substantial reduction in the number of newly generated neurons in the adult rat impairs hippocampal-dependent trace conditioning, a task in which an animal must associate stimuli that are separated in time. A similar reduction did not affect learning when the same stimuli are not separated in time, a task that is hippocampal-independent. The reduction in neurogenesis did not induce death of mature hippocampal neurons or permanently alter neurophysiological properties of the CA1 region, such as long-term potentiation. Moreover, recovery of cell production was associated with the ability to acquire trace memories. These results indicate that newly generated neurons in the adult are not only affected by the formation of a hippocampal-dependent memory, but also participate in it. PMID- 11268215 TI - Effects of chronic exposure to cocaine are regulated by the neuronal protein Cdk5. AB - Cocaine enhances dopamine-mediated neurotransmission by blocking dopamine re uptake at axon terminals. Most dopamine-containing nerve terminals innervate medium spiny neurons in the striatum of the brain. Cocaine addiction is thought to stem, in part, from neural adaptations that act to maintain equilibrium by countering the effects of repeated drug administration. Chronic exposure to cocaine upregulates several transcription factors that alter gene expression and which could mediate such compensatory neural and behavioural changes. One such transcription factor is DeltaFosB, a protein that persists in striatum long after the end of cocaine exposure. Here we identify cyclin-dependent kinase 5 (Cdk5) as a downstream target gene of DeltaFosB by use of DNA array analysis of striatal material from inducible transgenic mice. Overexpression of DeltaFosB, or chronic cocaine administration, raised levels of Cdk5 messenger RNA, protein, and activity in the striatum. Moreover, injection of Cdk5 inhibitors into the striatum potentiated behavioural effects of repeated cocaine administration. Our results suggest that changes in Cdk5 levels mediated by DeltaFosB, and resulting alterations in signalling involving D1 dopamine receptors, contribute to adaptive changes in the brain related to cocaine addiction. PMID- 11268216 TI - BRI1 is a critical component of a plasma-membrane receptor for plant steroids. AB - Most multicellular organisms use steroids as signalling molecules for physiological and developmental regulation. Two different modes of steroid action have been described in animal systems: the well-studied gene regulation response mediated by nuclear receptors, and the rapid non-genomic responses mediated by proposed membrane-bound receptors. Plant genomes do not seem to encode members of the nuclear receptor superfamily. However, a transmembrane receptor kinase, brassinosteroid-insensitive1 (BRI1), has been implicated in brassinosteroid responses. Here we show that BRI1 functions as a receptor of brassinolide, the most active brassinosteroid. The number of brassinolide-binding sites and the degree of response to brassinolide depend on the level of BRI1 protein. The brassinolide-binding activity co-immunoprecipitates with BRI1, and requires a functional BRI1 extracellular domain. Moreover, treatment of Arabidopsis seedlings with brassinolide induces autophosphorylation of BRI1, which, together with our binding studies, shows that BRI1 is a receptor kinase that transduces steroid signals across the plasma membrane. PMID- 11268217 TI - Regulation of CD40 and CD40 ligand by the AT-hook transcription factor AKNA. AB - Proteins containing AT hooks bind A/T-rich DNA through a nine-amino-acid motif and are thought to co-regulate transcription by modifying the architecture of DNA, thereby enhancing the accessibility of promoters to transcription factors. Here we describe AKNA, a human AT-hook protein that directly binds the A/T-rich regulatory elements of the promoters of CD40 and CD40 ligand (CD40L) and coordinately regulates their expression. Consistent with its function, AKNA is a nuclear protein that contains multiple PEST protein-cleavage motifs, which are common in regulatory proteins with high turnover rates. AKNA is mainly expressed by B and T lymphocytes, natural killer cells and dendritic cells. During B lymphocyte differentiation, AKNA is mainly expressed by germinal centre B lymphocytes, a stage in which receptor and ligand interactions are crucial for B lymphocyte maturation. Our findings show that an AT-hook molecule can coordinately regulate the expression of a key receptor and its ligand, and point towards a molecular mechanism that explains homotypic cell interactions. PMID- 11268220 TI - Pharmacogenetics initiative galvanizes public and private sectors. PMID- 11268218 TI - Transcription coactivator p300 binds PCNA and may have a role in DNA repair synthesis. AB - The transcriptional coactivator p300 interacts with many transcription factors that participate in a broad spectrum of biological activities, such as cellular differentiation, homeostasis and growth control. Mouse embryos lacking both p300 alleles die around mid-gestation, with pleiotropic defects in morphogenesis, in cell differentiation and, unexpectedly, in cell proliferation because of reduced DNA synthesis. Here we show that p300 may have a role in DNA repair synthesis through its interaction with proliferating cell nuclear antigen (PCNA). We show that in vitro and in vivo p300 forms a complex with PCNA that does not depend on the S phase of cell cycle. A large fraction of both p300 and PCNA colocalize to speckled structures in the nucleus. Furthermore, the endogenous p300-PCNA complex stimulates DNA synthesis in vitro. Chromatin immunoprecipitation experiments indicate that p300 is associated with freshly synthesized DNA after ultraviolet irradiation. Our results suggest that p300 may participate in chromatin remodelling at DNA lesion sites to facilitate PCNA function in DNA repair synthesis. PMID- 11268221 TI - Physicians vs Physicians. PMID- 11268222 TI - Americans' views on the use and regulation of dietary supplements. AB - This article presents the views of Americans on what the government's future role should be in regulating or overseeing the growing sales of dietary supplements for health purposes. Based on results of multiple national opinion surveys, including the views of both users and nonusers of supplements, we found that a substantial percentage of Americans surveyed reported that they regularly take dietary supplements as a part of their routine health regimen. However, they reported that they do not discuss the use of dietary supplements with their physicians because they believe that the physicians know little or nothing about these products and may be biased against them. Many users felt so strongly about the potential health benefits of some of these products that they reported that they would continue to take them even if they were shown to be ineffective in scientifically conducted clinical studies. However, there also was broad public support for increased government regulation of these products. We found that a majority of Americans surveyed supported the following: to require that the Food and Drug Administration review the safety of new dietary supplements prior to their sale; to provide increased authority to remove from sale those products shown to be unsafe; and to increase government regulation to ensure that advertising claims about the health benefits of dietary supplements are true. PMID- 11268224 TI - Cardiovascular fitness as a predictor of mortality in men. AB - OBJECTIVE: To examine the relations of cardiorespiratory fitness, as measured by maximal oxygen uptake and exercise test duration at the initiation of the study, with overall, cardiovascular disease (CVD)-related, and non-CVD-related mortality. METHODS: A population-based cohort study of 1294 men with no CVD, pulmonary disease, or cancer at baseline in Kuopio and surrounding communities in eastern Finland. During an average follow-up of 10.7 years, there were 124 overall, 42 CVD-related, and 82 non-CVD-related deaths. RESULTS: The relative risk of overall death in unfit men (maximal oxygen uptake <27.6 mL/kg per minute) was 2.76 (95% confidence interval, 1.43-5.33) (P =.002), and the relative risk of CVD-related death was 3.09 (95% confidence interval, 1.10-9.56) (P =.05), compared with fit men (maximal oxygen uptake >37.1 mL/kg per minute) after adjusting for age, examination years, smoking, and alcohol consumption. The relative risk of non-CVD-related death in unfit men was almost the same magnitude as for overall death. Furthermore, adjustment for serum lipid levels, blood pressure, plasma fibrinogen level, diabetes, and fasting serum insulin level did not weaken these associations significantly. Exercise test duration also had a strong inverse relation to overall, CVD-related, and non-CVD-related mortality. Poor cardiorespiratory fitness was comparable with elevated systolic blood pressure, smoking, obesity, and diabetes in importance as a risk factor for mortality. CONCLUSIONS: Cardiorespiratory fitness had a strong, graded, inverse association with overall, CVD-related, and non-CVD-related mortality. Maximal oxygen uptake and exercise test duration represent the strongest predictors of mortality. PMID- 11268223 TI - Garlic shows promise for improving some cardiovascular risk factors. AB - OBJECTIVES: To summarize the effects of garlic on several cardiovascular-related factors and to note its adverse effects. METHODS: English and non-English citations were identified from 11 electronic databases, references, manufacturers, and experts from January 1966 through February 2000 (depending on the database searched). Reports of cardiovascular-related effects were limited to randomized controlled trials lasting at least 4 weeks. Reports of adverse effects were not limited by study design. From 1798 pertinent records, 45 randomized trials and 73 additional studies reporting adverse events were identified. Two physicians abstracted outcomes and assessed adequacy of randomization, blinding, and handling of dropouts. Standardized mean differences of lipid outcomes from placebo-controlled trials were adjusted for baseline differences and pooled using random effects methods. RESULTS: Compared with placebo, garlic preparations may lead to small reductions in the total cholesterol level at 1 month (range of average pooled reductions, 0.03-0.45 mmol/L [1.2-17.3 mg/dL]) and at 3 months (range of average pooled reductions 0.32-0.66 mmol/L [12.4-25.4 mg/dL]), but not at 6 months. Changes in low-density lipoprotein levels and triglyceride levels paralleled total cholesterol level results; no statistically significant changes in high-density lipoprotein levels were observed. Trials also reported significant reductions in platelet aggregation and mixed effects on blood pressure outcomes. No effects on glycemic-related outcomes were found. Proven adverse effects included malodorous breath and body odor. Other unproven effects included flatulence, esophageal and abdominal pain, allergic reactions, and bleeding. CONCLUSIONS: Trials suggest possible small short-term benefits of garlic on some lipid and antiplatelet factors, insignificant effects on blood pressure, and no effect on glucose levels. Conclusions regarding clinical significance are limited by the marginal quality and short duration of many trials and by the unpredictable release and inadequate definition of active constituents in study preparations. PMID- 11268225 TI - Electron-beam computed tomography in the diagnosis of coronary artery disease: a meta-analysis. AB - BACKGROUND: Electron-beam computed tomography (EBCT) is a new, noninvasive method of detecting coronary artery calcification that is being increasingly advocated as a diagnostic test for coronary artery disease (CAD). Before its clinical use is justified, however, the overall accuracy of EBCT must be better defined. OBJECTIVE: To estimate the accuracy of EBCT in diagnosing obstructive CAD. DATA SOURCES: English-language studies from January 1, 1979, through February 29, 2000, were retrieved using MEDLINE and Current Contents databases, bibliographies, and expert consultation. STUDY SELECTION: We included a study if it (1) used EBCT as a diagnostic test; (2) reported cases in absolute numbers of true-positive, false-positive, true-negative, and false-negative results; and (3) used coronary angiography as the reference standard for diagnosing obstructive CAD (defined as > or = 50% diameter stenosis). DATA EXTRACTION: Data were extracted from the included articles by 2 independent reviewers. DATA SYNTHESIS: Weighted pooled analysis and summary receiver operating characteristic (ROC) curve analysis were used to determine sensitivity and specificity rates. Results from 9 studies with 1662 subjects were included. Pooled sensitivity for EBCT was 92.3% (95% confidence interval [CI], 90.7%-94.0%) and pooled specificity was 51.2% (95% CI, 47.5%-54.9%). Maximum joint sensitivity and specificity for EBCT from its summary ROC curve was 75%. As the threshold for defining an abnormal test varied, sensitivity and specificity changed. For a threshold that resulted in a sensitivity of 90%, specificity was 54%; when sensitivity was 80%, specificity rose to 71%. CONCLUSION: The performance of EBCT as a diagnostic test for obstructive CAD is reasonable based on sensitivity and specificity rates from its summary ROC curve. PMID- 11268226 TI - Improving lipid evaluation and management in medicare patients hospitalized for acute myocardial infarction. AB - BACKGROUND: The control of low-density lipoprotein cholesterol (LDL-C) levels in patients with known coronary artery disease, particularly in those with acute myocardial infarction, has been shown to reduce the rates of disease progression, recurrent events, and mortality. OBJECTIVES: To evaluate and improve hospital based processes for measuring and treating, when necessary, LDL-C levels above 3.36 mmol/L (>130 mg/dL) in patients with an acute myocardial infarction. DESIGN: A nonrandomized retrospective baseline study followed by a collaborative educational intervention with participating hospitals and a second nonrandomized postintervention study. PATIENTS: Four hundred six preintervention patients discharged from the hospital alive after a confirmed acute myocardial infarction in 1996, and 498 postintervention patients discharged from the hospital in 1999. INTERVENTIONS: Performance of lipid profiles on admission to the hospital and during hospitalization and drug and dietary interventions. RESULTS: The measurement of LDL-C level on admission to the hospital increased from 8% preintervention in 1996 to 32% postintervention in 1999. The measurement during hospitalization increased from 14% preintervention to 48% postintervention. Hospitals that initiated programs to ensure early lipid evaluations through preprinted orders and policy changes achieved an average patient LDL-C measurement rate of 70% in 1999. Hospitals lacking standard policies averaged only 23% at the same time. Of the patients with a measured LDL-C level greater than 3.36 mmol/L (>130 mg/dL) who were not undergoing drug therapy on admission to the hospital, 46% were given lipid-lowering agents by discharge from the hospital during the postintervention period. During this same period, only 11% of the patients were prescribed this therapy if they had either a lower measured level or no LDL-C measurement at all. CONCLUSION: Active hospital-based programs to ensure routine LDL-C measurements in patients admitted for acute myocardial infarction increased the use of appropriate lipid-lowering therapy in these high risk individuals and could contribute to reducing the incidence of recurrent coronary artery disease. PMID- 11268227 TI - Early switch from intravenous to oral antibiotics in hospitalized patients with bacteremic community-acquired Streptococcus pneumoniae pneumonia. AB - BACKGROUND: The identification of Streptococcus pneumoniae bacteremia in hospitalized patients with community-acquired pneumonia is considered by some investigators to be an exclusion criterion for early switch from intravenous to oral therapy. OBJECTIVE: To determine whether the switch from intravenous to oral therapy in such patients, once the bx;1patient reaches clinical stability, is associated with poor clinical outcome. METHODS: The medical records of 400 patients with community-acquired pneumonia hospitalized at the Veterans Affairs Medical Center of Louisville (Louisville, Ky) were reviewed to identify patients with bacteremic S pneumoniae. Four criteria were used to define when a patient reached clinical stability and should be considered a candidate for switch therapy: (1) cough and shortness of breath are improving, (2) patient is afebrile for at least 8 hours, (3) white blood cell count is normalizing, and (4) oral intake and gastrointestinal tract absorption are adequate. RESULTS: A total of 36 bacteremic patients were identified. No clinical failures occurred in 18 patients who reached clinical stability and were switched to oral therapy or in 7 patients who reached clinical stability and continued intravenous therapy. Clinical failures (5 deaths) occurred in the group of 11 patients who did not reach clinical stability. CONCLUSION: Once a hospitalized patient with community acquired pneumonia reaches clinical stability, it is safe to switch from intravenous to oral antibiotics even if bacteremia caused by S pneumoniae was initially documented. PMID- 11268228 TI - Characteristics and work experiences of hospitalists in the United States. AB - BACKGROUND: Little is known about the personal characteristics, work-related attitudes, or professional experiences of hospitalists. In considering the future of hospital medicine as a viable career choice for physicians (primarily, internists), these issues should be examined in a systematic fashion. Learning more about hospitalists and their work can enhance dialogue about the advantages and shortcomings of such a career from the perspective of the individual physician. METHODS: A self-administered mail survey was sent to 820 hospitalists who are dues-paying members of the National Association of Inpatient Physicians and who spend 50% or more of their time doing clinical work, teaching, or research related to hospital medicine. Attitudes about topics such as job-related burnout and job satisfaction were tapped, as well as information about different professional and social experiences. The analyses were performed using descriptive statistics and analysis of variance techniques. RESULTS: Analysis was based on 393 responses (48% response rate). Results show hospitalists to be a group of younger, mostly male, early-career individuals with high levels of job satisfaction and autonomy, low levels of burnout, and a long-term commitment to remaining in the role. Hospital medicine is a source of positive social and professional work experiences related to interactions with physician peers, patients and their families, and nonphysician hospital coworkers. Key components of hospitalists' jobs, practices, and workload are coalescing. However, certain developments, such as changing patterns of compensation and the enlisting of more general internists and women as hospitalists, merit further examination. CONCLUSIONS: The results offer insight into the physicians who are becoming hospitalists, the jobs and settings in which they work, and how hospitalists experience their everyday work lives. Valuable baseline data are provided for assessment of attitudes, such as burnout, that should be examined regularly in this fledgling group. This study complements research looking at the performance related outcomes of hospitalists, and it can be used by various stakeholders to better understand and assess the long-term potential of what is being proposed as a new career path. PMID- 11268229 TI - Eradication of methicillin-resistant Staphylococcus aureus from a health center ward and associated nursing home. AB - BACKGROUND: Long-term health care facilities have been recognized as reservoirs of multiresistant bacterial strains, especially methicillin-resistant Staphylococcus aureus (MRSA). Efforts to control MRSA in this setting usually have been only partially effective. We describe herein the eradication of epidemic MRSA from a Finnish health care center ward and affiliated nursing home. METHODS: The methods to control MRSA included (1) contact isolation precautions, (2) screening for asymptomatic carriage, (3) eradication of carriage, and (4) education of staff on hygienic measures. The first 6 patients with MRSA-positive findings were referred without delay to the Infectious Diseases Unit of the adjacent university hospital for eradication treatment. Later, an isolation unit of 6 rooms was founded in the health care center, where the MRSA-colonized patients were nursed as a separate cohort until they, in succession, were referred to the Infectious Diseases Unit for decolonization. RESULTS: From May 20 through August 17, 1993, the epidemic MRSA strain was isolated from 8 long-term patients on the 40-bed ward of the health care center, 4 of the 59 residents of the nursing home, and 1 member of the staff. Eradication of carriage was successful in all except 1 patient with dementia, who was nursed in contact isolation in the health care center until his death 21 months later. CONCLUSIONS: It is possible to eradicate MRSA from a long-term health care facility even after 13 cases by applying strict control measures. Our experience may be valuable in the future decision-making process for control of new and more challenging multiresistant bacteria, eg, vancomycin-resistant strains of MRSA. PMID- 11268230 TI - Monoclonal gammopathy of undetermined significance and exposure to breast implants. AB - BACKGROUND: Animal studies and uncontrolled case series in humans have suggested a possible association between breast implant exposure and monoclonal gammopathy. OBJECTIVE: To assess whether there is an increased risk of monoclonal gammopathy in women with silicone breast implants, we conducted a retrospective study of women exposed to breast implants and matched nonexposed women nested within a prospective cohort study (the Nurses' Health Study). METHODS: We used serum protein electrophoresis and immunoglobulin subtype by immunofixation to test 288 women exposed to breast implants and 288 age-matched, nonexposed women who previously had provided a blood sample (1989-1990) for monoclonal proteins. RESULTS: Among the women exposed to breast implants, 5 had monoclonal gammopathy of undetermined significance (MGUS) compared with 4 women among those not exposed (odds ratio, 1.25; 95% confidence interval, 0.27-6.39). The distribution of isotypes was similar across exposure groups. The exposed women with MGUS tended to be older than the nonexposed women (mean age, 60.4 years vs 52.5 years, respectively; P =.03). None of the 9 women with MGUS had reported multiple myeloma or other hematologic malignancies up through 1996. CONCLUSIONS: We find little evidence to support a substantial increased risk of MGUS in women exposed to breast implants. Larger studies are needed to determine if a more modest relationship exists. PMID- 11268231 TI - Communicating with dying patients within the spectrum of medical care from terminal diagnosis to death. AB - BACKGROUND: Efforts to improve communication between physicians and dying patients have been unsuccessful, and guidelines for improving patient-physician communication about end-of-life care are based primarily on expert opinion. This study assessed which aspects of communication between patients and physicians are important in end-of-life care. METHODS: Twenty focus groups were held with 137 individuals, including patients with chronic and terminal illnesses, family members, health care professionals from hospice or acute care settings, and physicians with expertise in end-of-life care. Focus group analyses determined domains of physician skill at end-of-life care. Communication with patients was identified as one of the most important domains. Analyses of components important in communicating with dying patients and their families were performed. RESULTS: The following 6 areas were of central importance in communicating with dying patients: talking with patients in an honest and straightforward way, being willing to talk about dying, giving bad news in a sensitive way, listening to patients, encouraging questions from patients, and being sensitive to when patients are ready to talk about death. Within these components, subthemes emerged that provide guidelines for physicians and educators. Dying patients also identified the need to achieve a balance between being honest and straightforward and not discouraging hope. CONCLUSIONS: Several areas emerged for physicians to focus their attention on when communicating with dying patients. These findings provide guidance in how to improve this communication. They also highlight the need to approach communication about end-of-life care as a spectrum that requires attention from the time of a terminal diagnosis through death. PMID- 11268232 TI - Clinical predictors of mental disorders among medical outpatients. AB - BACKGROUND: Mental disorders are common among primary care patients and often not detected by primary care physicians. We report on clinical cues that may allow physicians to target patients for psychiatric screening. METHODS: Two hundred fifty consecutive adults presenting to a walk-in clinic completed previsit surveys assessing demographics, symptom characteristics, recent stress, functional status (Medical Outcomes Study Short Form-6), and mental disorders (Primary Care Evaluation of Mental Disorders [PRIME-MD]). Patients with positive findings for a mental disorder on the PRIME-MD underwent a semistructured interview. Immediately after the visit, physicians completed the Difficult Doctor Patient Relationship Questionnaire. RESULTS: Patients averaged 50.5 years of age (range, 18-92 years). Little more than half were women (53%); 43%, white; 44%, African American; 8%, Hispanic; and 6%, other. Twenty-six percent had an underlying mental disorder; 11% had more than 1 mental disorder. Sixteen percent had a depressive disorder; 6%, major depression; 11%, an anxiety disorder; 2%, panic disorder; and 9%, a somatoform disorder. Independent correlates of a mental disorder included reporting recent stress (odds ratio [OR], 6.7; 95% confidence interval [CI], 3.3-13.6), having 5 or more physical symptoms (OR, 4.0; 95% CI, 2.1-7.9), or reporting health to be less than very good (OR, 2.2; 95% CI, 1.1 4.3). There was a stepwise increase in the likelihood of having a mental disorder and number of correlates present. Among patients with no predictors, only 2% had an underlying mental disorder, compared with 72% among patients with all 3 clinical predictors. CONCLUSIONS: Patients who report recent stress, 5 or more physical symptoms, or poor health are more likely to have an underlying mental disorder. These clinical cues may allow clinicians to select patients in whom formal screening for mental disorders would be particularly fruitful. PMID- 11268233 TI - Adverse drug effects, compliance, and initial doses of antihypertensive drugs recommended by the Joint National Committee vs the Physicians' Desk Reference. AB - BACKGROUND: Compliance problems are common causes of the inadequate treatment of hypertension, with 16% to 50% of patients quitting treatment within 1 year. Dose related adverse drug events (ADEs) frequently cause compliance problems, and many ADEs occur with the initial doses of antihypertensive drugs. Thus, it is an established tenet to initiate antihypertensive therapy at low doses to avoid ADEs that diminish patients' quality of life and reduce compliance. However, what are the lowest effective doses of antihypertensive drugs? OBJECTIVE: To compare the initial doses recommended in the Physicians' Desk Reference (PDR) with those recommended by the Sixth Report of the Joint National Committee on the Detection, Evaluation, and Treatment of High Blood Pressure (JNC VI). METHODS: Review of the latest JNC VI report (1997) and the 1999 and 2000 editions of the PDR and the medical literature. RESULTS: The JNC VI recommends substantially lower initial doses for 23 (58%) of 40 drugs, compared with the PDR. In addition, for 37 (82%) of 45 drugs, PDR guidelines do not suggest lower initial doses for old or frail patients than for younger adults. CONCLUSIONS: Although the PDR is the drug reference most used by physicians, it does not reflect the lowest initial doses that are recommended by the JNC VI for many of the most prescribed antihypertensive drugs. Because avoidance of ADEs is essential to maintaining compliance with antihypertensive therapy, and because many antihypertensive ADEs are dose related, physicians must know the very lowest, effective, least ADE prone doses. Patients and physicians would benefit by establishing mechanisms to make this information readily available to all practicing physicians. PMID- 11268234 TI - High-normal serum creatinine concentration is a predictor of cardiovascular risk in essential hypertension. AB - BACKGROUND: Determination of serum creatinine concentration is recommended in all patients with hypertension as a marker of target organ damage. However, the possibility that creatinine values within the reference range might contribute to stratification of cardiovascular risk in essential hypertension has never been tested. PATIENTS AND METHODS: In the setting of the Progetto Ipertensione Umbria Monitoraggio Ambulatoriale Study, for up to 11 years (mean, 4 years) we followed up 1829 white patients with hypertension (mean +/- SD age, 51 +/- 12 years; 53% men; office blood pressure, 157/98 mm Hg) free of cardiovascular events and with normal pretreatment creatinine levels (men, <136 micromol/L [<1.5 mg/dL]; women, <120 micromol/L [<1.4 mg/dL]) who also underwent 24-hour blood pressure monitoring and electrocardiography before therapy. RESULTS: During follow-up, there were 175 fatal or nonfatal major cardiovascular morbid events (2.4 per 100 patient-years). Event rate increased progressively from the first to the fourth sex-specific quartiles of creatinine distribution (1.5, 2.3, 2.3, and 3.5 per 100 patient-years; P =.003 by log-rank test). After adjustment (in a multivariate Cox model) for age, sex, diabetes, cholesterol, smoking, left ventricular hypertrophy, and 24-hour pulse and mean blood pressures (P<.05 for all), creatinine concentration was an independent adverse predictor of cardiovascular morbid events (P =.01). The observed excess risk was 1.30 (95% confidence interval, 1.07-1.59) for a 20-micromol/L (0.23-mg/dL) increase in creatinine concentration. CONCLUSIONS: A serum creatinine value within the reference range is a predictor of cardiovascular morbidity in white patients with essential hypertension. Its prognostic value persists after adjustment for several powerful confounders, including average 24-hour blood pressure and left ventricular hypertrophy. PMID- 11268235 TI - Preclinical Cushing disease. PMID- 11268236 TI - Acute rhabdomyolysis associated with cerivastatin therapy. PMID- 11268237 TI - New sham method in auricular acupuncture. PMID- 11268238 TI - A randomized controlled trial of auricular acupuncture for cocaine dependence: treatments vs outcomes. PMID- 11268240 TI - An automated screening strategy to identify patients with alcohol problems in a primary care setting. PMID- 11268244 TI - A piece of my mind. A simple song of gratitude. PMID- 11268245 TI - Easing the elderly online in search of health information. PMID- 11268246 TI - Awareness of "silent strokes" stressed. PMID- 11268247 TI - New vaccine decreases rate of nosocomial infections. PMID- 11268253 TI - Public health risks of not vaccinating children. PMID- 11268254 TI - Public health risks of not vaccinating children. PMID- 11268258 TI - A woman experiencing difficulty with breastfeeding. PMID- 11268256 TI - Postexposure rabies prophylaxis in immunosuppressed patients. PMID- 11268259 TI - A woman experiencing difficulty with breastfeeding. PMID- 11268261 TI - Accuracy of hair mineral analysis. PMID- 11268262 TI - Accuracy of hair mineral analysis. PMID- 11268264 TI - Immunogenicity of bivalent AC polysaccharide meningococcal vaccine in children aged 6 through 24 months. PMID- 11268265 TI - Physical abuse of women before, during, and after pregnancy. AB - CONTEXT: Clinicians who care for new mothers and infants need information concerning postpartum physical abuse of women as a foundation on which to develop appropriate clinical screening and intervention procedures. However, no previous population-based studies have been conducted of postpartum physical abuse. OBJECTIVES: To examine patterns of physical abuse before, during, and after pregnancy in a representative statewide sample of North Carolina women. DESIGN, SETTING, AND PARTICIPANTS: Survey of participants in the North Carolina Pregnancy Risk Assessment Monitoring System (NC PRAMS). Of the 3542 women invited to participate in NC PRAMS between July 1, 1997, and December 31, 1998, 75% (n = 2648) responded. MAIN OUTCOME MEASURES: Prevalence of physical abuse during the 12 months before pregnancy, during pregnancy, and after infant delivery; injuries and medical interventions resulting from postpartum abuse; and patterns of abuse over time in relation to sociodemographic characteristics and use of well-baby care. RESULTS: The prevalence of abuse before pregnancy was 6.9% (95% confidence interval [CI], 5.6%-8.2%) compared with 6.1% (95% CI, 4.8%-7.4%) during pregnancy and 3.2% (95% CI, 2.3%-4.1%) during a mean postpartum period of 3.6 months. Abuse during a previous period was strongly predictive of later abuse. Most women who were abused after pregnancy (77%) were injured, but only 23% received medical treatment for their injuries. Virtually all abused and nonabused women used well baby care; private physicians were the most common source of care. The mean number of well-baby care visits did not differ significantly by maternal patterns of abuse. CONCLUSION: Since well-baby care use is similar for abused and nonabused mothers, pediatric practices may be important settings for screening women for violence. PMID- 11268266 TI - Relation of gemfibrozil treatment and lipid levels with major coronary events: VA HIT: a randomized controlled trial. AB - CONTEXT: A low plasma level of high-density lipoprotein cholesterol (HDL-C) is a major risk factor for coronary heart disease (CHD). A secondary prevention study, the Veterans Affairs High-Density Lipoprotein Intervention Trial (VA-HIT), demonstrated that CHD events were significantly reduced during a median follow-up of 5.1 years by treating patients with the fibric acid derivative gemfibrozil when the predominant lipid abnormality was low HDL-C. OBJECTIVE: To determine if the reduction in major CHD events with gemfibrozil in VA-HIT could be attributed to changes in major plasma lipid levels. DESIGN: Multicenter, randomized, double blind, placebo-controlled trial conducted from September 1991 to August 1998. SETTING: The Department of Veterans Affairs Cooperative Studies Program, in which 20 VA medical centers were participating sites. PARTICIPANTS: A total of 2531 men with a history of CHD who had low HDL-C levels (mean, 32 mg/dL [0.83 mmol/L] ) and low low-density lipoprotein cholesterol (LDL-C) levels (mean, 111 mg/dL [2.88 mmol/L]). INTERVENTION: Participants were randomly assigned to receive gemfibrozil, 1200 mg/d (n = 1264), or matching placebo (n = 1267). MAIN OUTCOME MEASURE: Relation of lipid levels at baseline and averaged during the first 18 months of gemfibrozil treatment with the combined incidence of nonfatal myocardial infarction and CHD death. RESULTS: Concentrations of HDL-C were inversely related to CHD events. Multivariable Cox proportional hazards analysis showed that CHD events were reduced by 11% with gemfibrozil for every 5-mg/dL (0.13-mmol/L) increase in HDL-C (P =.02). Events were reduced even further with gemfibrozil beyond that explained by increases in HDL-C values, particularly in the second through fourth quintiles of HDL-C values during treatment. During gemfibrozil treatment, only the increase in HDL-C significantly predicted a lower risk of CHD events; by multivariable analysis, neither triglyceride nor LDL-C levels at baseline or during the trial predicted CHD events. CONCLUSIONS: Concentrations of HDL-C achieved with gemfibrozil treatment predicted a significant reduction in CHD events in patients with low HDL-C levels. However, the change in HDL-C levels only partially explained the beneficial effect of gemfibrozil. PMID- 11268268 TI - Validation of a clinical decision aid to discontinue in-hospital cardiac arrest resuscitations. AB - CONTEXT: Most patients undergoing in-hospital cardiac resuscitation do not survive to hospital discharge. In a previous study, we developed a clinical decision aid for identifying all patients undergoing resuscitation who survived to hospital discharge. OBJECTIVE: To validate our previously derived clinical decision aid. DESIGN, SETTING, AND PARTICIPANTS: Data from a large registry of in hospital resuscitations at a community teaching hospital in Georgia were analyzed to determine whether patients would be predicted to survive to hospital discharge (ie, whether their arrest was witnessed or their initial cardiac rhythm was either ventricular tachycardia or ventricular fibrillation or they regained a pulse during the first 10 minutes of chest compressions). Data from 2181 in hospital cardiac resuscitation attempts in 1987-1996 involving 1884 pulseless patients were analyzed. MAIN OUTCOME MEASURE: Comparison of predictions based on the decision aid with whether patients were actually discharged alive from the hospital. RESULTS: For 327 resuscitations (15.0%), the patient survived to hospital discharge. For 324 of these resuscitations, the patients were predicted to survive to hospital discharge (sensitivity = 99.1%, 95% confidence interval, 97.1%-99.8%). In 269 resuscitations, patients did not satisfy the decision aid and were predicted to have no chance of being discharged from the hospital. Only 3 of these patients (1.1%) were discharged from the hospital (negative predictive value = 98.9%), none of whom were able to live independently following discharge from the hospital. CONCLUSION: This decision aid can be used to help physicians identify patients who are extremely unlikely to benefit from continued resuscitative efforts. PMID- 11268267 TI - Leukocyte-reduced red blood cell transfusions in patients with anemia and human immunodeficiency virus infection: the Viral Activation Transfusion Study: a randomized controlled trial. AB - CONTEXT: Allogeneic blood transfusions have immunomodulatory effects and have been associated with activation of human immunodeficiency virus (HIV) and cytomegalovirus (CMV) in vitro and of HIV in small pilot studies. Retrospective studies suggest that transfusions adversely affect the clinical course of HIV. Data in selected non-HIV-infected patients requiring blood transfusion have suggested clinical benefit with leukocyte-reduced red blood cells (RBCs). OBJECTIVE: To compare the effects of leukoreduced and unmodified RBC transfusions on survival, complications of acquired immunodeficiency syndrome, and relevant laboratory markers in HIV-infected patients. DESIGN AND SETTING: Double-blind randomized controlled trial conducted in 11 US academic medical centers from July 1995 through June 1999, with a median follow-up of 12 months (24 months in survivors). PATIENTS: A total of 531 persons infected with HIV and CMV, aged 14 years or older, who required transfusions for anemia; 259 received leukoreduced transfusions and 262 received unmodified transfusions (10 did not receive the planned transfusion). MAIN OUTCOME MEASURES: Survival and change in plasma HIV RNA level 7 days after transfusion, compared by type of transfusion. RESULTS: At entry, the groups were similar in demographic, clinical, and relevant laboratory characteristics. A total of 3864 RBC units were transfused. Two hundred eighty nine deaths occurred (151 with leukoreduced transfusion; 138 with unmodified transfusion); median survival was 13.0 and 20.5 months, respectively (relative hazard [RH], 1.20; 95% confidence interval [CI], 0.95-1.51; log-rank P =.12). Analyses adjusted for prognostic factors suggested possible worse survival with leukoreduction (RH, 1.35; 95% CI, 1.06-1.72). There was no difference in time to new opportunistic event/death or frequency of transfusion reactions. No changes in plasma HIV RNA level were seen in either group at days 7, 14, 21, or 28, even in patients not taking antiretroviral drugs. There were no differences in trends between groups in CMV DNA, CD4 cell counts, activated (CD38% or human leukocyte antigen-DR) CD8 cell counts, or plasma cytokine levels. CONCLUSIONS: We found no evidence of HIV, CMV, or cytokine activation following blood transfusion in patients with advanced HIV infection. Leukoreduction provided no clinical benefit in these patients. These data demonstrate the importance of conducting controlled studies of effects of leukoreduction in different patient populations, since smaller studies in other patient populations have suggested leukoreduction may be beneficial. PMID- 11268269 TI - Association of maternal endothelial dysfunction with preeclampsia. AB - CONTEXT: Preeclampsia is believed to result from release of placental factors that damage maternal vascular endothelium. However, because most studies have been conducted during pregnancy, it has not been possible to separate maternal from placental mechanisms underlying endothelial dysfunction in preeclampsia. OBJECTIVE: To determine whether endothelial function is impaired in nonpregnant women with previous preeclampsia and whether endothelial dysfunction is mediated by oxidative stress. DESIGN AND SETTING: Case-control study conducted at 3 hospital maternity units in London, England, between July 1997 and June 2000. PARTICIPANTS: A total of 113 women with previous preeclampsia (n = 35 with recurrent episodes; n = 78 with a single episode) and 48 women with previous uncomplicated pregnancies, all of whom were at least 3 months (median, 3 years) postpartum. MAIN OUTCOME MEASURES: Brachial artery flow-mediated (endothelium dependent) and glyceryl trinitrate-induced (endothelium-independent) dilatation were compared between previously preeclamptic women and controls. To investigate oxidative stress, these measurements were repeated after administration of ascorbic acid, 1 g intravenously, in 15 cases and 15 controls. RESULTS: Mean (SD) flow-mediated dilatation was lower in women with previous preeclampsia compared with controls (recurrent group, 0.9% [4.1%]; single-episode group, 2.7% [3.5%]; and control group, 4.7% [4.3%]; P<.001). In contrast, glyceryl trinitrate-induced dilatation was similar in the 3 groups (recurrent, 19.5% [5.9%]; single-episode, 21.0% [8.0%]; and control, 21.0% [8.3%]; P =.65). Impaired flow-mediated dilatation in previously preeclamptic women was not accounted for by recognized vascular risk factors. Ascorbic acid administration increased flow-mediated dilatation in previously preeclamptic women (baseline, 2.6% [3.3%]; after administration, 5.6% [3.0%]; P =.001) but not in controls (baseline, 6.2% [3.3%]; after administration, 6.7% [5.0%]; P =.72). CONCLUSIONS: Our results indicate that endothelial function is impaired in women with previous preeclampsia and is not explained by established maternal risk factors but is reversed by antioxidant ascorbic acid administration. PMID- 11268271 TI - Cocaine and pregnancy--time to look at the evidence. PMID- 11268270 TI - Growth, development, and behavior in early childhood following prenatal cocaine exposure: a systematic review. AB - CONTEXT: Despite recent studies that failed to show catastrophic effects of prenatal cocaine exposure, popular attitudes and public policies still reflect the belief that cocaine is a uniquely dangerous teratogen. OBJECTIVE: To critically review outcomes in early childhood after prenatal cocaine exposure in 5 domains: physical growth; cognition; language skills; motor skills; and behavior, attention, affect, and neurophysiology. DATA SOURCES: Search of MEDLINE and Psychological Abstracts from 1984 to October 2000. STUDY SELECTION: Studies selected for detailed review (1) were published in a peer-reviewed English language journal; (2) included a comparison group; (3) recruited samples prospectively in the perinatal period; (4) used masked assessment; and (5) did not include a substantial proportion of subjects exposed in utero to opiates, amphetamines, phencyclidine, or maternal human immunodeficiency virus infection. DATA EXTRACTION: Thirty-six of 74 articles met criteria and were reviewed by 3 authors. Disagreements were resolved by consensus. DATA SYNTHESIS: After controlling for confounders, there was no consistent negative association between prenatal cocaine exposure and physical growth, developmental test scores, or receptive or expressive language. Less optimal motor scores have been found up to age 7 months but not thereafter, and may reflect heavy tobacco exposure. No independent cocaine effects have been shown on standardized parent and teacher reports of child behavior scored by accepted criteria. Experimental paradigms and novel statistical manipulations of standard instruments suggest an association between prenatal cocaine exposure and decreased attentiveness and emotional expressivity, as well as differences on neurophysiologic and attentional/affective findings. CONCLUSIONS: Among children aged 6 years or younger, there is no convincing evidence that prenatal cocaine exposure is associated with developmental toxic effects that are different in severity, scope, or kind from the sequelae of multiple other risk factors. Many findings once thought to be specific effects of in utero cocaine exposure are correlated with other factors, including prenatal exposure to tobacco, marijuana, or alcohol, and the quality of the child's environment. Further replication is required of preliminary neurologic findings. PMID- 11268272 TI - Screening mothers for intimate partner abuse at well-baby care visits: the right thing to do. PMID- 11268273 TI - Medical malpractice and legal resolution systems in Japan. AB - Medical malpractice claims and dispute resolution systems have been examined in Western societies for their impact on the quality of care and efficient compensation for injured patients. However, little is known about the Japanese malpractice environment because claim information has been closely guarded. Based on data from the Japanese Supreme Court, the Ministry of Health, Labor, and Welfare (formerly the Ministry of Health and Welfare), and the Japan Medical Association (JMA), which provides malpractice insurance to 43.5% of Japan's 250 000 physicians, we review Japanese malpractice liability systems and the frequency of claims during the last 30 years. Annual premiums for physician professional liability insurance are relatively low (454 dollars-491 dollars). Although the frequency of claims in Japan is lower than that reported in the United States, England, and Germany, the number of claims is increasing in Japan. According to publicly available data from the Japanese Supreme Court, the annual number of medical malpractice suits filed in district courts has increased from 102 in 1970 to 629 in 1998 (from 0.09 to 0.25 per 100 physicians). The proportion of awards greater than 89 dollars 300 increased from 13.6% in 1976 to 65.4% in 1987. Among JMA members, claims increased 31% from 1987 to 1999, but the frequency of claims has remained at approximately 0.3 per 100 JMA members. The JMA's professional liability program offers a nonbinding out-of-court review of claims that is faster and less expensive than judicial resolution (a few months with no attorney required vs 35 months and attorney fees), but is a poor means of deterrence or discipline. Since JMA data represent only 43.5% of Japanese physicians, generalizations cannot be made about all Japanese physicians and institutions. The lack of data on all claims hinders adequate evaluation of dispute resolution methods, development of appropriate risk management activities, and proactive education for Japanese physicians. PMID- 11268280 TI - EGTA enhancement of adenovirus-mediated gene transfer to mouse tracheal epithelium in vivo. AB - Administration of recombinant adenoviral (AdV) vectors to animals can lead to inflammatory and immune responses. For therapeutic indications in which repeated treatment is necessary, such as cystic fibrosis (CF), these responses can limit the therapeutic usefulness of the vector. In principle, the utility of the vector can be improved by increasing its therapeutic index, that is, by either increasing its efficacy or decreasing its toxicity. A strategy that would enhance the efficacy of an adenoviral approach would allow the use of fewer virus particles to achieve a given level of transgene expression, and thereby also reduce unwanted effects such as immune responses. Following up on our observation that treating polarized normal human bronchial epithelial cells with calcium (Ca(2+))-free medium or the calcium chelator ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) significantly enhanced the subsequent transfection of these cells with cationic lipid:pDNA complexes, we have now asked whether such a treatment protocol might also improve the ability of AdV to infect these cells. Treating polarized airway epithelial cells with EGTA led to a dramatic increase in AdV-mediated transduction, as demonstrated by an approximately 50-fold increase in transgene expression. This strategy was also tested in vivo and resulted in substantial increases (up to 50-fold) in the ability of AdV vectors to infect mouse tracheal epithelium. Transfection of mouse trachea with an AdV aerosol was also significantly increased by pretreatment with EGTA. The enhancing effects of EGTA could not be duplicated with hypo- or hyperosmotic treatments. Light microscopy of mouse trachea that had been EGTA treated and then infected with AdV demonstrated an EGTA-mediated AdV infection of airway epithelial cells. The apparent enhanced potency of AdV for airway cells resulting from this strategy provides a significant increase in the therapeutic index of this gene delivery vector, and may increase the likelihood that it can be used for clinical indications requiring chronic administration of the vector. PMID- 11268281 TI - Safety evaluation of hemagglutinating virus of Japan--artificial viral envelope liposomes in nonhuman primates. AB - We tested, in cynomolgus monkeys, the safety and effectiveness of a hybrid liposome vector, hemagglutinating virus of Japan (HVJ)--artificial viral envelope (AVE) liposomes, for human therapeutic gene transfer in a series of experiments. In a repetitive intramuscular administration study, vehicle control macaques (n = 2), which were treated with HVJ--AVE liposome suspension, received repetitive intramuscular injections of 2 ml of test substance. Human hepatocyte growth factor (HGF) cDNA-inserted expression vector (pUC-SR alpha/HGF) injection animals (n = 2), which were treated with HVJ--AVE liposome suspension containing pUC-SR alpha/HGF, received repetitive intramuscular injection of 2 ml of test substance. General body condition, hematology, blood chemistry, and serum HGF were determined sequentially before treatment and 7, 21, 28, and 29 days after treatment. Elevations in HGF were detected in monkeys injected with pUC-SR alpha/HGF. After this observation period, macaques were killed for autopsy and histological examination. pUC-SR alpha/HGF was detected by polymerase chain reaction (PCR) analysis in the liver, spleen, and at the injection site. In single intravenous administration study, control macaques (n = 4) received a single intravenous injection of 10 ml of physiological saline. Vehicle control animals (n = 5) received a single intravenous injection of 10 ml of HVJ--AVE liposome suspension. DNA-treated animals (n = 7) received a single intravenous injection of 10 ml of HVJ--AVE liposome suspension containing plasmid DNA [pcDNA 3.1(+)]. General body condition, body weight, hematology, blood chemistry, and urine composition were determined sequentially before treatment and 1, 14, 21, and 28 days after treatment. After this observation period, macaques were killed for autopsy and histological examination. pcDNA 3.1(+) was detected by PCR analysis on day 1 in lung, liver, and spleen of all monkeys, in kidney of one of two monkeys, and in heart of one of two monkeys. However, no DNA was detected in any of the tissues examined on days 14, 21, and 28. No virus genomic RNA was detected by reverse transcription (RT)-PCR analysis with HVJ-specific primers. In this series of safety evaluations, the animals tolerated the safety study with no change in body weight or general condition. No hematological changes or alterations in blood chemistry or urine composition was detected. Moreover, no histological changes were observed. This safety evaluation study demonstrates the safety, feasibility, and therapeutic potential of the novel transfection vehicle, HVJ--AVE liposomes, in humans. PMID- 11268283 TI - Adenovirus-mediated atrial natriuretic protein expression in the lung protects rats from hypoxia-induced pulmonary hypertension. AB - Endogenous as well as exogenous atrial natriuretic peptide (ANP) attenuates the development of chronic hypoxic pulmonary hypertension (CHPH) in rats. We built a recombinant adenovirus type 5 containing ANP cDNA under the control of the Rous sarcoma virus long terminal repeat (Ad.ANP). The efficiency of this vector in delivering the ANP gene was first examined in rat primary cultures of pulmonary vessel smooth muscle cells (SMCs) in comparison with Ad.beta GAL. Conditioned medium collected from Ad.ANP-infected cells (1000 TCID(50)/cell) contained 5 x 10(9) M immunoreactive ANP and elicited relaxation of isolated rat pulmonary arteries preconstricted with phenylepinephrine. To examine the effects of adenovirus-mediated ANP expression in the CHPH rat lung, Ad.ANP or Ad.beta GAL was administered via the tracheal route. Immunoreactive ANP was detected in bronchoalveolar fluid as early as 4 days and until 10-17 days after Ad.ANP administration (5 x 10(8) TCID(50)). Lung ANP immunostaining was mainly localized in bronchial and alveolar epithelial cells. As compared with Ad.beta GAL-treated controls, rats given Ad.ANP (5 x 10(8) TCID(50)) on the day before a 2-week exposure to hypoxia (10% O(2)) had lower values for pulmonary artery pressure (32.1 +/- 1.93 vs. 35.5 +/- 2 mmHg, p < 0.01) and Fulton's index (0.52 +/- 0.089 vs. 0.67 +/- 0.12, p < 0.001) and less severe right ventricular hypertrophy and distal vessel muscularization. These results suggest that induction of ANP expression in the lung may hold promise in the treatment of pulmonary hypertension. PMID- 11268282 TI - Adenovirus-mediated interleukin 3 beta gene transfer by isolated limb perfusion inhibits growth of limb sarcoma in rats. AB - Cytokine gene transfer using (multiple) intratumoral injections can induce tumor regression in several animal models, but this administration technique limits the use for human gene therapy. In the present studies we describe tumor growth inhibition of established limb sarcomas after a single isolated limb perfusion (ILP) with recombinant adenoviral vectors harboring the rat IL-3 beta gene (IG.Ad.CMV.rIL-3 beta). In contrast, a single intratumoral injection or intravenous administration did not affect tumor growth. Dose-finding studies demonstrated a dose-dependent response with a loss of antitumor effect below 1 x 10(9) IU of IG.Ad.CMV.rIL-3 beta. Perfusions with adenoviral vectors bearing a weaker promoter (MLP promoter) driving the rIL-3 beta gene did not result in antitumor responses, suggesting that the rIL-3 beta-mediated antitumor effect depends on the amount of rIL-3 beta protein expressed by the infected cells. Furthermore, it was shown by direct comparison that ILP with IG.Ad.CMV.rIL-3 beta in the ROS-1 osteosarcoma model is at least as efficient as the established therapy with the combination of TNF-alpha and melphalan. Treatment with IG.Ad.CMV.rIL-3 beta induced a transient dose-dependent leukocytosis accompanied by an increase in peripheral blood levels of histamine. Leukocyte infiltrations were also histopathologically demonstrated in tumors after perfusion. These results demonstrate that ILP with recombinant adenoviral vectors carrying the IL 3 beta transgene inhibits tumor growth in rats and suggest that cytokine gene therapy using this administration technique might be beneficial for clinical cancer treatment. PMID- 11268284 TI - AdTIMP-2 inhibits tumor growth, angiogenesis, and metastasis, and prolongs survival in mice. AB - TIMP-2 is a natural matrix metalloproteinase (MMP) inhibitor that prevents the degradation of extracellular matrix proteins. It abolishes the hydrolytic activity of all activated members of the metalloproteinase family and in particular that of MT1-MMP, MMP-2, and MMP-9, which are selective for type IV collagenolysis. Since MMPs have been implicated in both cancer progression and angiogenesis, we generated a recombinant adenovirus to deliver human TIMP-2 (AdTIMP-2) and evaluated its anticancer efficiency in three murine models. Our results demonstrated that overexpression in vitro of TIMP-2 inhibited the invasion of both tumor and endothelial cells without affecting cell proliferation. Its in vivo efficiency has been evaluated in murine lung cancer LLC, and colon cancer C51 in syngeneic mice as well as in human breast cancer MDA MB231 in athymic mice. Preinfection of tumor cells by AdTIMP-2 resulted in an inhibition of tumor establishment in more than 50% of mice in LLC and C51 models and in 100% mice in the MDA-MB231 model. A single local injection of AdTIMP-2 into preestablished tumors of these three types significantly reduced tumor growth rates by 60--80% and tumor-associated angiogenesis index by 25--75%. Lung metastasis of LLC tumor was inhibited by >90%. In addition, AdTIMP-2-treated mice showed a significantly prolonged survival in all the cancer models tested. These data demonstrate the potential of adenovirus-mediated TIMP-2 therapy in cancer treatment. PMID- 11268285 TI - Intercellular transfer of the virally derived precursor form of acid alpha glucosidase corrects the enzyme deficiency in inherited cardioskeletal myopathy Pompe disease. AB - Pompe disease is a lethal cardioskeletal myopathy in infants and results from genetic deficiency of the lysosomal enzyme acid alpha-glucosidase (GAA). Genetic replacement of the cDNA for human GAA (hGAA) is one potential therapeutic approach. Three months after a single intramuscular injection of 10(8) plaque forming units (PFU) of E1-deleted adenovirus encoding human GAA (Ad-hGAA), the activity in whole muscle lysates of immunodeficient mice is increased to 20 times the native level. Direct transduction of a target muscle, however, may not correct all deficient cells. Therefore, the amount of enzyme that can be transferred to deficient cells from virally transduced cells was studied. Fibroblasts from an affected patient were transduced with AdhGAA, washed, and plated on transwell culture dishes to serve as donors of recombinant enzyme. Deficient fibroblasts were plated as acceptor cells, and were separated from the donor monolayer by a 22-microm pore size filter. Enzymatic and Western analyses demonstrate secretion of the 110-kDa precursor form of hGAA from the donor cells into the culture medium. This recombinant, 110-kDa species reaches the acceptor cells, where it can be taken up by mannose 6-phosphate receptor-mediated endocytosis. It then trafficks to lysosomes, where Western analysis shows proteolytic processing to the 76- and 70-kDa lysosomal forms of the enzyme. Patient fibroblasts receiving recombinant hGAA by this transfer mechanism reach levels of enzyme activity that are comparable to normal human fibroblasts. Skeletal muscle cell cultures from an affected patient were also transduced with Ad-hGAA. Recombinant hGAA is identified in a lysosomal location in these muscle cells by immunocytochemistry, and enzyme activity is transferred to deficient skeletal muscle cells grown in coculture. Transfer of the precursor protein between muscle cells again occurs via mannose 6-phosphate receptors, as evidenced by competitive inhibition with 5 mM mannose 6-phosphate. In vivo studies in GAA knockout mice demonstrate that hepatic transduction with adenovirus encoding either murine or human GAA can provide a depot of recombinant enzyme that is available to heart and skeletal muscle through this mechanism. Taken together, these data show that the mannose 6-phosphate receptor pathway provides a useful strategy for cell-to-cell distribution of virally derived recombinant GAA. PMID- 11268286 TI - Cochlear gene delivery through an intact round window membrane in mouse. AB - Cochlear gene transfer studies in animal models have utilized mainly two delivery methods: direct injection through the round window membrane (RWM) or intracochlear infusion through a cochleostomy. However, the surgical trauma, inflammation, and hearing loss associated with these methods lead us to investigate a less invasive delivery method. Herein, we studied the feasibility of a vector transgene-soaked gelatin sponge, Gelfoam, for transgene delivery into the mouse cochlea through an intact RWM. The Gelfoam absorbed with liposomes and adenovirus, but not with adeno-associated virus (AAV), was successful in mediating transgene expression across an intact RWM in a variety of cochlear tissues. The Gelfoam technique proved to be an easy, atraumatic, and effective, but vector-dependent, method of delivering transgenes through an intact RWM. Compared with the more invasive gene delivery methods, this technique represents a safer and a more clinically viable route of cochlear gene delivery in humans. PMID- 11268287 TI - Robust prostate-specific expression for targeted gene therapy based on the human kallikrein 2 promoter. AB - Tissue-specific transcriptional regulatory elements can increase the safety of gene therapy vectors. Unlike prostate-specific antigen (PSA/hK3), whose expression displays an inverse correlation with prostate cancer grade and stage, human glandular kallikrein 2 (hK2) is upregulated in higher grade and stage disease. Therefore, our goal was to develop a strong and prostate-specific hK2 based promoter for targeted gene therapy. We identified the minimum "full strength" hK2 enhancer and built transcriptional regulatory elements composed of multiple tandem copies of this 1.2-kb enhancer, fused to the hK2 minimal promoter. Relative to the weak induction of the minimal hK2 promoter by androgen analog (R1881) in androgen receptor (AR)-positive LNCaP cells, transcriptional activity was increased by 25-, 44-, 81-, and 114-fold when one to four enhancers were spliced to the hK2 promoter, respectively. In contrast, the enhancer/promoter elements were inactive in the AR(-) prostate cancer line PC-3 and in a panel of nonprostate lines, including 293, U87, MCF-7, HuH-7, and HeLa cells. Furthermore, we generated a recombinant adenovirus, ADV.hK2-E3/P-EGFP, expressing enhanced green fluorescent protein (EGFP) under the control of the hK2 triplicate enhancer/promoter, and compared its properties with ADV.CMV-EGFP expressing EGFP under the control of the cytomegalovirus (CMV) enhancer/promoter. Unlike the CMV promoter, the hK2-E3/P promoter was at least 100-fold inducible by R1881 in the adenoviral backbone. Compared with in situ injection of subcutaneous LNCaP tumors with ADV.CMV-EGFP, which led to detectable EGFP expression in tumor, liver, and brain tissue, ADV.hK2-E3/P-EGFP injection led to robust but tumor restricted EGFP expression. These results suggest that the hk2 multienhancer/promoter should be a powerful novel reagent for safer targeted gene therapy of prostate cancer. PMID- 11268289 TI - Gene therapy of malignant gliomas: a pilot study of vaccination with irradiated autologous glioma and dendritic cells admixed with IL-4 transduced fibroblasts to elicit an immune response. PMID- 11268292 TI - Anxiety and uncertainty in informed decision making. PMID- 11268288 TI - CMV-beta-actin promoter directs higher expression from an adeno-associated viral vector in the liver than the cytomegalovirus or elongation factor 1 alpha promoter and results in therapeutic levels of human factor X in mice. AB - Although AAV vectors show promise for hepatic gene therapy, the optimal transcriptional regulatory elements have not yet been identified. In this study, we show that an AAV vector with the CMV enhancer/chicken beta-actin promoter results in 9.5-fold higher expression after portal vein injection than an AAV vector with the EF1 alpha promoter, and 137-fold higher expression than an AAV vector with the CMV promoter/enhancer. Although induction of the acute-phase response with the administration of lipopolysaccharide (LPS) activated the CMV promoter/enhancer from the context of an adenoviral vector in a previous study, LPS resulted in only a modest induction of this promoter from an AAV vector in vivo. An AAV vector with the CMV-beta-actin promoter upstream of the coagulation protein human factor X (hFX) was injected intravenously into neonatal mice. This resulted in expression of hFX at 548 ng/ml (6.8% of normal) for up to 1.2 years, and 0.6 copies of AAV vector per diploid genome in the liver at the time of sacrifice. Neonatal intramuscular injection resulted in expression of hFX at 248 ng/ml (3.1% of normal), which derived from both liver and muscle. We conclude that neonatal gene therapy with an AAV vector with the CMV-beta-actin promoter might correct hemophilia due to hFX deficiency. PMID- 11268294 TI - News from the Society for Women's Health Research: seeking effective topical microbicides. PMID- 11268295 TI - Toward optimal health: the experts discuss weight control drugs. Interview by Jodi Godfrey Meisler. PMID- 11268296 TI - Can variability in the hormonal status of elderly women assist in the decision to administer estrogens? AB - Hormone replacement therapy (HRT) has been proposed for the prevention and treatment of many chronic conditions, ranging from osteoporosis, heart disease, urinary incontinence, and Alzheimer's disease. With the exception of osteoporosis, however, many of the suggested benefits remain controversial. Part of the controversy stems from the relative absence of randomized controlled trials, particularly those enrolling sufficient numbers of elderly women. We propose that another factor may also contribute, one that has been overlooked - failure to consider the variable endogenous estrogen status of elderly women. Highly variable levels of estrogens are present in nearly all postmenopausal women, even at advanced ages. Similar to other endocrine systems, estrogen deficiency and the need for its replacement are, therefore, likely to be relative rather than absolute. Recent studies indicate that elderly women who are less able to compensate for declining ovarian 17beta-estradiol production by adipose synthesis of estrone (E1) may be at greater risk for certain chronic conditions associated with relative estrogen deficiency. Because many markers of estrogen deficiency exhibit overlap between risk groups, their clinical usefulness as predictors of frailty, disability, and response to HRT has been limited. Future studies will need to focus not only on the use of highly variable circulating serum estrogen levels but also on markers of overall estrogenic effects at the level of individual target tissues (i.e., markers of bone turnover, karyopyknotic index on a vaginal wall smear). We propose that a clinical approach that takes into consideration the remarkable heterogeneity (physiological as well as psychological) of elderly women will enable us to approach the decision about HRT in a more individualized and possibly better targeted fashion. PMID- 11268297 TI - Cardiovascular health interventions in women: What works? AB - Women's Cardiovascular Health Network members representing 10 Prevention Research Centers completed a literature review of approximately 65 population-based studies focused on improving women's cardiovascular health through behavior change for tobacco use, physical inactivity, or diet. A framework was developed for conducting the search. Databases (Medline, Psychlit, Smoking and Health, Cumulative Index to Nursing and Allied Health Literature) of studies published from 1980 to 1998 were searched. The review was presented at a meeting of experts held in Atlanta, Georgia. Output from the meeting included identification of what has worked to improve cardiovascular health in women and recommendations for future behavioral research. Additional information is available at www.hsc.wvu.edu/womens-cvh. Cardiovascular health interventions geared toward women are scant. Based on the available studies, program components that emerged as effective included personalized advice on diet and physical activity behaviors and tobacco cessation, multiple staff contacts with skill building, daily self monitoring, and combinations of strategies. Recommendations for community-based tobacco, physical activity, and diet interventions are discussed. A few overarching recommendations were to (1) conduct qualitative research to determine the kinds of interventions women want, (2) examine relapse prevention, motivation, and maintenance of behavior change, (3) tailor programs to the stage of the life cycle, a woman's readiness to change, and subgroups, that is, minority, low socioeconomic, and obese women, and (4) evaluate policy and environmental interventions. The effects of cardiovascular interventions in women have been inappropriately understudied in women. Our review found that few studies on cardiovascular risk factor modification have actually targeted women. Hence, adoption and maintenance of behavior change in women are elusive. Intervention research to improve women's cardiovascular health is sorely needed. PMID- 11268299 TI - Acceptability research on female-controlled barrier methods to prevent heterosexual transmission of HIV: Where have we been? Where are we going? AB - Acceptability research is an important component of any product development process. As researchers move into a new, accelerated phase of vaginal microbicide development, it is important to take stock of the acceptability research conducted to date and determine future research priorities. In this paper, we review findings from acceptability research conducted to date in four categories: hypothetical product acceptability research, existing product research (spermicide acceptability studies), acceptability research within the context of clinical trials, and postmarketing acceptability research conducted around the female condom. Finally, we highlight areas where additional research is needed in light of recent progress in microbicide development and discuss a possible framework for the introduction and acceptability of new sexually transmitted disease (STD) prevention technologies. PMID- 11268298 TI - Medical and surgical therapies for pain associated with endometriosis. AB - Endometriosis is a common condition for which a number of treatments have been proposed. Medical treatments are based on the hormonal responsiveness of endometriosis implants. These therapies include progestins (with or without estrogens), androgens, and gonadotropin-releasing hormone (GnRH) analogs. Surgical treatments may include hysterectomy with oophorectomy or organ-sparing surgery involving ablation or resection of visible lesions of endometriosis and restoration of pelvic anatomy. There are no studies that directly compare the effectiveness or adverse effects of medical therapy and surgical therapy. Studies on medical therapy compare different treatments with placebo or with other active treatments. Hormone-based therapies for endometriosis show 80%-100% effectiveness in relief of pelvic pain over a 6-month course of therapy. Serious adverse outcomes after medical therapy are unusual. Studies on surgical therapy are largely anecdotal, with noncomparative reports on a variety of surgical methods. A few comparative surgical studies have been reported. Because of the noncomparative nature of many of the surgical studies, the use of combinations of surgical procedures and techniques in the reported studies, and the reporting of results from surgeons with an unusually high level of technical skill, the gynecological practitioner has little basis in the literature for assessing the optimum surgical approach. Surgical complications are believed to be underreported and may be related to how aggressive a surgical procedure is undertaken. PMID- 11268300 TI - We've come a long way, maybe: recruitment of women and analysis of results by sex in clinical research. AB - During the last decade, North American policymakers have started to demand more representative research populations. Several papers have suggested that there has been improvement, over the last decade, in the number of studies that include women as subjects, yet these same papers have expressed concern that many investigators omit analysis of data by sex from their research reports. Our study examined all clinical research ethics applications from July 1, 1995, to June 30, 2000, at a tertiary care Canadian university teaching hospital to determine whether the investigator planned to recruit both men and women and whether he or she intended to perform analysis of data by sex. For research studying nonsex specific conditions, 97.6% of researchers intended to recruit both men and women, yet only 20.2% planned to perform analysis of data by sex. This proportion decreased from 29.9% in 1995-1996 to 16.9% in 1999-2000. Seventy-seven percent of the applications submitted were for studies involving drugs, and only 17% of these nonsex-specific studies planned an analysis of data by sex. The results of this study indicate that although researchers in Canada are aware of the importance of planning to recruit women into clinical trials, more needs to be done to ensure that they plan and perform analyses of data by sex. PMID- 11268301 TI - Preliminary observations on differing psychological effects of conjugated and esterified estrogen treatments. AB - During a double-blind comparison of menopausal replacement therapy with estrogen alone compared with estrogen plus methyltestosterone (meT), subjects who had been on conjugated equine estrogen (CEE) said they felt better when placed on esterified estrogen (EE). We, therefore, tested whether these estrogen treatments differed in their neuropsychological effects. Subjects were 34 healthy menopausal respondents to advertisements younger than age 66 who were on CEE at baseline. Each was randomized into the EE condition, either immediately after baseline or after they first took EE plus added meT for 8 weeks. We compared neuropsychological measures between these two conditions. Data included cognitive performance test results and symptom self-ratings. Multivariate techniques were used to adjust for the effects of treatment order. Compared with prior CEE treatment, EE treatment was associated with significantly improved scores on the Zung Self-Rated Depression Scale and on Switching Attention Test performance. Further investigation is warranted to determine if different forms of estrogen replacement induce different neuropsychological effects. PMID- 11268305 TI - Prostate cancer screening. PMID- 11268303 TI - Rural physicians' perspectives on cervical and breast cancer screening: a gender based analysis. AB - Several studies highlight the role of physicians in determining cervical and breast cancer screening rates, and some urban studies report higher screening rates by female physicians. Rural women in North America remain underscreened for breast and cervical cancers. This survey was conducted to determine if there were significant gender differences in practices and perceptions of barriers to breast and cervical cancer screening among rural family physicians in Ontario, Canada. One hundred ninety-one family physicians (response rate 53.1%) who practiced in rural areas, small towns, or small cities completed a mail questionnaire. The physicians' mean age was 44.4 years (SD 9.9), and mean number of years in practice was 16.6 years (SD 10.3). Over 90% of physicians reported that they were very likely to conduct a Pap test and clinical breast examination (CBE) during a periodic health examination, and they had high levels of confidence and comfort in performing these procedures. Male (68%) and female (32%) physicians were similar in their likelihood to conduct screening, levels of confidence and comfort, and knowledge of breast and cervical cancer screening guidelines. However, the self-reported screening rates for Pap tests and CBE performed during last year were higher for female than male physicians (p < 0.01). Male physicians reported they were asked more frequently by patients for a referral to another physician to perform Pap tests and CBE (p < 0.001). Also, male physicians perceived patients' embarrassment as a stronger barrier to performing Pap tests (p < 0.05) and CBE (p < 0.01) than female physicians. No gender differences were observed in screening rates or related barriers to mammography referrals. These findings suggest that physicians' gender plays a role in sex-sensitive examination, such as Pap tests and CBE. There is a need to facilitate physician patient interactions for sex-sensitive cancer screening examinations by health education initiatives targeting male physicians and women themselves. The feasibility of providing sex-sensitive cancer screening examinations by a same sex health provider should also be explored. PMID- 11268302 TI - Anxiety/uncertainty reduction as a motivation for interest in prophylactic oophorectomy in women with a family history of ovarian cancer. AB - Most women at familial risk for ovarian cancer must decide about prophylactic oophorectomy without conclusive genotypic information about their risk level. Some women with relatively low-risk profiles seek prophylactic oophorectomy or are recommended the procedure by their physicians, if they appear "cancerphobic." This study investigated the desire to reduce anxiety in relation to other factors associated with interest in prophylactic oophorectomy in a group of women with varying degrees of familial risk for ovarian cancer. Ninety-four women enrolled in an ongoing program for women with a family history of ovarian cancer received personalized risk counseling and were classified as having a sporadic, familial, or putative hereditary pedigree by a genetics counselor. Eligible enrollees were interviewed by telephone about current and future interest in prophylactic oophorectomy, perceived risk of ovarian cancer, severity of cancer anxiety, stress-related ideation, and reasons for and against surgery. Reduction of anxiety/uncertainty was the factor most strongly associated with current interest in prophylactic oophorectomy, independent of objective risk classification, perceived risk, severity of cancer anxiety, intrusive ideation, or other variables. Future interest in prophylactic oophorectomy was predicted by other perceived benefits of surgery. Current, but not future, interest in prophylactic oophorectomy appears motivated in part by seeking immediate relief from anxiety. Interest in prophylactic oophorectomy may fluctuate based on varying exposure to cues that trigger anxiety. Women seeking prophylactic oophorectomy, particularly those with lower-risk family pedigrees, should be offered options for anxiety management as part of informed consent for prophylactic oophorectomy. PMID- 11268307 TI - The application of appendiceal Mitrofanoff principle to Stanford pouch. AB - OBJECTIVE: The present paper reports the functional aspects of a novel continent cutaneous reservoir. PATIENTS AND METHODS: A continent cutaneous reservoir was constructed by the application of appendiceal Mitrofanoff principle to Stanford pouch in four male and three female patients between 1995-1998 in our clinic. The mean age of the patients was 45.6 years (7-67 years) and the etiological factor was carcinoma of the bladder in four, interstitial cystitis in one and extrophy epispadias complex in two cases. Patients were followed with arterial blood gas determination, serum biochemistry, urinalysis and urine culture at postoperative 3 weeks and by 3-month intervals thereafter. Additionally, pouch graphy, abdominal ultrasonography and urodynamic tests were performed every 6 months. RESULTS: After the operation all the patients were continent. Stoma was transferred from the umbilicus to the right lower quadrant in one case (14.3%) because of difficulty in catheterisation. Pouch graphy at postoperative 6 months revealed low-grade vesicoureteral reflux in two (28.6%) patients and one (14.3%) of them required suppressive antibiotic therapy because of pyelonephritis episodes. Another patient developed hyperchloremic metabolic acidosis and needed oral alkaline supplementation. The mean pouch capacity measured at postoperative 6 months was 423 (350-550) ml and the mean end-filling pressure was found as 21 (18-25) cmH2O. After a mean follow-up period of 37 (18-45) months all the patients remained continent and stable. CONCLUSION: The continent cutaneous reservoir presented herein is our alternative to orthotopic neobladder in female patients undergoing radical cystectomy. It also provided continence as well as good quality of life in patients with extrophy-epispadias complex and male patients after radical cystectomy and urethrectomy. PMID- 11268306 TI - Surveillance programs for early stage non-seminomatous testicular cancer: a practice guideline. AB - BACKGROUND AND PURPOSE: To identify an appropriate surveillance program for men with clinical stage I non-seminomatous germ cell tumors of the testis (NSGCT). MATERIALS AND METHODS: A systematic review of the published literature was combined with a consensus process, around the interpretation of the evidence in the context of conventional practice, to develop an evidence-based practice guideline. RESULTS: No randomized controlled trials (RCTs) comparing surveillance schedules were found, but data from 12 case series and one RCT which compared radiotherapy with surveillance were reviewed. Variations in the schedules were not associated with observed variations in relapse, salvage, or survival rates. CONCLUSIONS: Men with clinical stage I testicular cancer, as defined by a normal physical examination, normal radiological scans (computed tomography [CT]) and serum markers (alpha-fetoprotein [AFP] and beta-subunit of human chorionic gonadotropin (betaHCG) which are normal or fall within normal limits during their expected half-lives, are eligible for surveillance. A recommended surveillance schedule is as follows: 1) Physical examination, blood serum marker tests (AFP and HCG), and chest x-rays should be conducted every month in the first year, every 2 months in the second year, every 3 months in the third year, and every 6 months in the fourth and fifth years; and 2) CT scans of the abdomen and pelvis should be conducted every 3 months in the first year, every 4 to 6 months in the second year and every 6 months in the third year, and once a year in the fourth and fifth year. PMID- 11268308 TI - Surgical confirmation of ProstaScint abnormalities in two patients with high risk prostate cancer. AB - We describe our initial experience using ProstaScint scanning in addition to conventional imaging modalities for staging of high risk prostate cancer. Using our protocol, ProstaScint images detected abnormalities in pelvic lymph nodes not seen on CT scan or magnetic resonance imaging (MRI) in two patients. Subsequent surgical pelvic lymphadenectomy confirmed these abnormalities. Further patients will be accrued on this study to estimate the sensitivity and specificity of ProstaScint scanning. PMID- 11268309 TI - Testicular seminoma and Down's syndrome. AB - OBJECTIVE: To review cases of testicular seminoma in Down's syndrome (DS) patients. METHODS: The case of a patient diagnosed and treated at our institution is reported. The literature is reviewed for previous cases, with attention paid to age, stage, treatment and outcome. RESULTS: Nineteen cases were found. The mean age at diagnosis is 32. Seventy three percent are Stage I. At 47, our patient is the oldest reported case. A body of evidence exists for an association between DS and testicular seminoma. An increased rate of cryptorchidism, increased gonadotropin levels and genetic instability are possible pathways. CONCLUSION: The age distribution of seminoma in DS is skewed by the decreased life expectancy of patients with DS. More cases should be seen as this life expectancy increases. The stage at presentation in DS patients is comparable to that seen in the general population. Standard therapy has been successfully delivered in these patients. PMID- 11268310 TI - Small cell carcinoma of the prostate: an underrecognized entity. AB - Adenocarcinoma is by far the most commonly diagnosed histologic subtype among prostate malignancies. Historically, there has been little awareness of the rare but lethal small cell carcinoma (SCC) in association with prostate cancer. Within the last decade, however, several reports have documented the existence of a neuroendocrine-like tumor arising from cells in the prostate. There is evidence that the development of poorly-differentiated neuroendocrine cells (similar to those found in oat cell carcinomas of the lung) can be seen in the progression of an initially pure adenocarcinoma, possibly due to the totipotential nature of the basal or reserve cells normally present in the prostatic acini. Although pure SCC is rare, admixtures of adenocarcinoma and small cell components may be more prevalent than previously believed. Since effective treatment of a prostatic tumor, or part of a tumor, with an SCC etiology differs from that of pure adenocarcinoma, early recognition of any histologic or clinical changes in the patient with prostate cancer may alter the course of the disease. PMID- 11268311 TI - Hypothesis testing for the genetic background of quantitative traits. AB - The testing of Bayesian point null hypotheses on variance component models have resulted in a tough assigmment for which no clear and generally accepted method exists. In this work we present what we believe is a succeeding approach to such a task. It is based on a simple reparameterization of the model in terms of the total variance and the proportion of the additive genetic variance with respect to it, as well as on the explicit inclusion on the prior probability of a discrete component at origin. The reparameterization was used to bypass an arbitrariness related to the impropriety of uninformative priors onto unbounded variables while the discrete component was necessary to overcome the zero probability assigned to sets of null measure by the usual continuous variable models. The method was tested against computer simulations with appealing results. PMID- 11268312 TI - Marker assisted selection for the improvement of two antagonistic traits under mixed inheritance. AB - A Monte Carlo simulation was used to investigate the potential of Marker Assisted Selection (MAS) in a multiple-trait situation. Only additive effects were considered. The base population was assumed to be in linkage equilibrium and, next, the population was managed over 15 discrete generations, 10 males and 50 females were chosen out of the 100 candidates of each sex. Performance for two traits was simulated with an overall heritability of a given trait equal to 0.25 or 0.10 and the overall genetic correlation between traits was generally equal to -0.4 except in one case where it was equal to 0. The model involved one biallelic QTL, accounting for 10 or 20% of the genetic variance of a given trait, plus polygenes. Initial allelic frequencies at the QTL, were generally equal to 0.5 but in one case were equal to 0.1 and 0.9. A marker with 120 different alleles in the 60 founder parents was simulated in the vicinity of the QTL. Two values of the recombination rate between these two loci were considered, 0.10 and 0.02. The genetic evaluation was based on a multiple-trait BLUP animal model, accounting (MAS) or not (conventional BLUP) for marker information. Two sets of simulations were run: (1) a "missing data" case, with males having no record for one of the traits, and (2) a "secondary trait" case, with one trait having a weight in the aggregate genotype 4 times less than the other trait and the QTL acting only on this secondary trait. In the first set, evaluation methods were found to mainly affect the accuracy of overall genetic values prediction for the trait with missing data. In comparison with BLUP, MAS led to an extra overall genetic response for the trait with missing data, which was strongly penalised under the conventional BLUP, and to a deficit in response for the other trait. This more balanced evolution of the two traits was obtained, however, at the expense of the long-term overall cumulated response for the aggregate genotype, which was 1 to 2.5% lower than the one obtained under the conventional BLUP. In the second set of simulation, in the case of low initial frequency (0.1) of the QTL allele favourable to the secondary trait, MAS was found to be substantially more efficient to avoid losing this allele than BLUP only when the QTL had a large effect and the marker was close. More benefits should be expected from MAS with more specific applications,such as early selection of animals, or by applying dynamic procedures i.e. letting the respective weights to QTL and polygenic values in the selection criterion vary across generation. PMID- 11268313 TI - Relationships between type and longevity in the Holstein breed. AB - The relationship between type traits and longevity was studied in the French Holstein breed using a survival analysis model. In this model, the phenotypic value adjusted for systematic fixed effects, the estimated breeding value, or the residual value (defined as the difference between the adjusted phenotypic value and the estimated breeding value) of the cow for each type trait was included as a risk factor. This was done separately for two subpopulations (registered and nonregistered herds) and with or without adjustment for production traits, i.e., considering true or functional longevity. For both types of herds, udder traits (and above all, udder depth) clearly influenced the length of productive life. There seemed to be a more pronounced voluntary culling on type traits in registered herds. The correction for the within herd-year class of production traits, as a way to approximate functional longevity, increased the importance of udder traits and decreased the weight of capacity traits. The same results were obtained when the phenotypic value of the cow for type was replaced by her estimated breeding value, whereas residuals had little impact. The relationship between longevity and type traits was most often nonlinear, in particular for udder traits, but in this study, no trait with a clear intermediate optimum was found. PMID- 11268314 TI - A possible dominant white gene in Jersey cattle. AB - A white heifer ("Snow") was born in 1991 from coloured registered Jersey parents. She produced six calves sired by coloured Jersey bulls: three white bull calves, two white heifer calves, and one coloured bull calf. One of the white bull calves was mated with 40 Hereford x Friesian yearling heifers (white face, predominantly black body with some white patches). The 38 resulting calves included 16 white and 22 coloured calves. Twelve of the 16 white calves were heifers and four were bulls. Red or black spotting was recorded on some white calves. The results are consistent with an autosomal dominant mutant causing the white phenotype. The mutation appears to have arisen spontaneously in Snow, then passing to her white progeny and white grand-progeny. The white individuals varied from entirely white in a few cases, to most having some residual small areas of red or black pigmentation in patterns not typical of other reported white spotting patterns of cattle. PMID- 11268315 TI - Functional study and regional mapping of 44 hormono-regulated genes isolated from a porcine granulosa cell library. AB - cDNA clones from a pig granulosa cell cDNA library were isolated by (differential hybridisation for follicle stimulating hormone (FSH) regulation in granulosa cells in a previous study. The clones that did not match any known sequence were studied for their expression in granulosa cells (treated or not by FSH) and in fresh isolated ovarian follicles mainly by comparative RT-PCR analysis. These results give functional data on genes that may be implicated in follicular growing. These ESTs have been localised on the porcine genome, using a somatic cell hybrid panel, providing new type I markers on the porcine map and information on the comparative map between humans and pigs. PMID- 11268316 TI - Specific cytogenetic labeling of bovine spermatozoa bearing X or Y chromosomes using fluorescent in situ hybridization (FISH). AB - X and Y specific probes were identified in order to apply the fluorescent in situ hybridization (FISH) technique to bovine spermatozoa. For Y chromosome detection, the BRY4a repetitive probe, covering three quarters of the chromosome, was used. For X chromosome detection, a goat Bacterial Artificial Chromosome (BAC) specific to the X chromosome of bovine and goats and giving a strong FISH signal was used. Each probe labeled roughly 45% of sperm cells. The hybridization method will be useful for evaluating the ratio of X- and Y- bearing spermatozoa in a sperm sample and consequently can be used to evaluate the efficiency of sperm sorting by different techniques such as flow cytometry. PMID- 11268317 TI - Non-myeloablative allogeneic stem cell transplantation in children. PMID- 11268318 TI - Cord blood-derived hematopoietic progenitor cells: in vitro response to hematopoietic growth factors and their recruitment into the S-phase of the cell cycle. AB - BACKGROUND AND OBJECTIVES: In the recent years many studies on the expansion of cord blood (CB)-derived progenitor cells have been performed, whereas less information is available on their cycling status. The objective of this study was to evaluate the cycling status of CB-derived colony-forming cells (CFC) and long term culture-initiating cells (LTC-IC), and their recruitment into the S-phase of the cell cycle in response to a combination of cytokines. DESIGN AND METHODS: CB derived CFC and LTC-IC were first quantified by standard clonogenic assay and long-term culture, respectively. In a second set of experiments, CB-derived progenitor cells were incubated with interleukin(IL)-3, stem cell factor (SCF) and granulocyte colony-stimulating factor (G-CSF) and their cell cycle status assessed both by the cytosine arabinoside (Ara-C) suicide approach and by flow cytometric DNA analysis. RESULTS: We found that only small proportions of both CFC and LTC-IC were in the S-phase of the cell cycle. These estimates were confirmed by flow cytometric DNA analysis, which showed that 96% +/- 2% of CB derived CD34+ cells were in G0/G1 and only 1.6% +/- 0.4% in the S-phase. Staining of CD34+ cells with an anti-statin monoclonal antibody, a marker of the G0 phase, indicated that among CD34+ cells with a flow cytometric DNA content typical of the G0/G1 phase, 68% +/- 7% of cells were in the G0 phase of the cell cycle. Twenty-four hour incubation with IL-3, SCF and G-CSF significantly increased the proportion of cells in the S-phase for both CFC and LTC-IC without inducing any loss in their number. Flow cytometric DNA analysis also showed an increase of CD34+ cells in the S-phase upon continuous exposure to these cytokines. INTERPRETATIONS AND CONCLUSIONS: Our findings indicate that: i) a small number of CB-derived CFC and LTC-IC are in the S-phase of the cell cycle; ii) a substantial number of CD34+ cells with a flow cytometric DNA content typical of the G0/G1 fraction are cycling, as they are found in the G1 phase of the cell cycle; iii) 24-hour incubation with IL-3, SCF and G-CSF can drive a proportion of progenitor cells into the S-phase without reducing their number. PMID- 11268319 TI - Allogeneic bone marrow transplantation in hematologic disorders of childhood: new trends and controversies. AB - Bone marrow transplantation (BMT) represents an important therapeutic choice for several kinds of disorders: hematologic, metabolic and neoplastic pathologies can be treated with this strategy. The aim of this article is to describe the main indications for allogeneic BMT in haematologic disorders of childhood and possible problems related to this procedure. We consider only hematologic aspects, paying particular attention to unusual disorders of infancy as myelodysplastic syndromes and aplastic anemia. We also consider quality of life after a BMT in patients with sickle cell anemia and thalassemia major and compare this with quality of life of patients receiving chronic periodic blood transfusions. PMID- 11268320 TI - Cord blood transplantation in childhood. PMID- 11268322 TI - Haploidentical bone marrow transplantation in leukemia and genetic diseases. AB - From 1986 to June 2000, sixty children suffering from acute and chronic leukemia (n = 42, 33 of which in resistant relapse), genetic diseases (n = 11), aplastic anemia (n = 2, one of which with platelet refractoriness and bleeding), myelodysplasia (n = 5) received an haploidentical bone marrow, mismatched for 2-3 HLA loci. The donor's marrow was treated in vitro with vincristine and methylprednisolone to obtain a functional T depletion (MLC and CTL inhibition, functional blockade of Th1 and Th2). The prevalence of infectious complications and GVHD was similar to that recorded in matched unrelated donor (MUD) transplants. In situations of high risk of rejection (chronic leukemia, genetic diseases) we infused immediately one half of the harvest and then frozen aliquots from the second week. Of the 25 ALL and 8 AML in resistant relapse, 3 survived, disease-free at 14, 8 and 1 years respectively. Of the 3 ALL, transplanted during remission, 1 is surviving at 18 months. Of the 6 CML, 1 had fractionated bone marrow and is surviving at 3 years, and 5 had standard single dose infusion and died of progression of their disease after rejection of the graft (4) or blast crisis after complete engraftment (1). The 2 patients with aplastic anemia, those with myelodysplasia, and 6 of the 10 with genetic disorders died of transplant related complications or disease progression. 4 patients with osteopetrosis (n = 2), MLD (n = 1), Wiskott Aldrich dis. (n = 1) survive at 8, 2, 5 and 1.5 years respectively. In patients transplanted with fractionated marrow GVHD > 2nd grade occurred in 15%. Only one patient rejected the graft. Compared with MUD transplantation, mismatched BMT whenever performed in patients in good conditions provides similar outcome and widens the donor availability. PMID- 11268321 TI - Unrelated donor marrow transplantation: an update of the experience of the Italian Bone Marrow Transplant Group (GITMO). AB - Unrelated donor bone marrow transplant (UD-BMT) has become an attractive alternative source of hematopoietic cells for patients lacking a matched sibling. The aim of this paper was to report on results of the 696 UD BMTs performed in 31 Italian institutions during the first 10 years of activity of the Italian Bone Marrow Donor Registry (IBMDR). In 1989 the Italian Bone Marrow Transplant Group (GITMO) established the IBMDR to facilitate donor search and marrow procurement for patients lacking an HLA identical sibling. By end of December 1999, 260,000 HLA-A, B typed volunteer donors had been cumulatively registered and 2,620 searches had been activated for Italian patients. At least one HLA-A, B, DRB1 matched donor was found for 54% of the patients and 696 UD BMTs were performed. In 50% of cases the donor was found in the IBMDR and in 50% in 15 other Registries. The average time from search activation to transplant was 6 months for disease other than CML. For CML it was 14 months. Actuarial 12-month transplant-related mortality (TRM) was 68% in patients grafted between 1979 and 1992 and 44% for patients grafted after 1993. Twenty-eight per cent of patients developed grade III or IV acute GvHD and 24% developed extensive chronic GvHD. The rate of disease free survival at three years was 57% for patients with 1st chronic phase CML, 37% for patients with 1st or 2nd CR ALL, 31% for AML or MDS patients < or = 18 years of age and 54% for patients with inborn errors. We conclude that the IBMDR has benefited a substantial number of patients lacking a matched sibling and has facilitated the recruitment of UDs into the international donor pool. The long time required for the search is the major obstacle to the success of this programme. This suggests that early transplant and a decrease in TRM could further improve these encouraging results. PMID- 11268323 TI - Haploidentical peripheral blood and marrow stem cell transplantation in nine cases of primary immunodeficiency. AB - Bone marrow transplantation (BMT) is the treatment of choice in children affected by primary immunodeficiency (PID). Because only 10-15% of affected children have a familial HLA-identical donor alternative therapeutic options are BMT from a matched unrelated donor or an haploidentical BMT. In our experience only 40% of these children find a donor within the International Registry. Therefore, the remaining 50% children affected by PID are candidates for haploidentical BMT. Unfortunately, in PID other than sever-combined immunodeficiency (SCID), low engraftment rates have been reported because of minimal residual immunity. In order to enhance engraftment rate in haploidentical BMT in PID we suggest a protocol with addition of donor peripheral stem cells after mobilization with granulocyte colony-stimulating factor (G-CSF) (16 micrograms/kg for 5 days) and bone marrow cells. This procedure increases the cell load, which allows intensification of the conditioning regimen for induction of faster engraftment. The separation of CD34+ cells from leukapheresis products was achieved in the first 6 patients by the Isolex 300 system (Baxter) with a CD34+ cell purity range of 80-95% and in another three patients by the Clinimacs System (Miltenyi). The peripheral blood stem cells were cryopreserved until BMT, 15 days after G-CSF stimulation when the bone marrow was harvested, processed and T-cell depleted with Campath 1-M in the first 6 cases while the Clinimacs System was used in the remaining cases and no T-cell depletion was required. We included 9 patients in the study protocol: SCID (4), Omenn's syndrome (3), LAD (1) and CID (1). The mean value of peripheral CD34+ cells infused was 13.42 x 10(6)/kg and the mean CD3+ cells number was 0.385 x 10(5)/kg; the mean value of BM CD34+ cells infused was 10.62 x 10(6)/kg and the mean CD3+ cell number was 2.39 x 10(5)/kg. The mean number of infused CFU was 8.1 x 10(5)/kg for PBSC and 3.59 x 10(5)/kg for BM. The 9 patients achieved more than 0.5 x 10(9) peripheral blood neutrophils/L at a mean of 14.6 days (range: 6-22 days). One patient affected by SCID showed complete chimerism, but he died after BMT of systemic CMV infection; the other 8 patients are alive and well and 4 of them show complete chimerism in all cell lines. Split chimerism was documented in 2 SCID cases (CD3+ lymphocytes were of donor origin, monocytes were autologous and granulocytes were mainly autologous); 1 patient affected by Omenn's syndrome received 3 transplants (1 from the mother and 2 from the father, T-cells alone and bone marrow) and achieved engraftment with complete chimerism after the third transplant; the patient affected by LAD also received 3 transplants (2 bone marrow infusions and 1 PBSC infusion) achieving complete chimerism after the third one. In conclusion, the engraftment achieved in all treated patients, and the acceptable conditioning-related toxicity suggest that this approach could be successfully applied to children affected by PID and candidates for haploidentical BMT. PMID- 11268325 TI - Surveillance of cytomegalovirus infections in bone marrow transplant in Trieste: seven years' experience. AB - Forty-five consecutive patients submitted to a bone marrow transplant (BMT) were followed up weekly in order to evaluate the incidence of cytomegalovirus (CMV) infections on the basis of CMV antigenemia and polymerase chain reaction. All but one transplanted patients engrafted; fourteen patients out of these were CMV antigenemia positive after 16-184 days (median 32.5, mean 43.4) with an 31.8% incidence. CMV infections were associated with graft-versus-host disease and immunogenetic relationship between the donor and the recipient. No CMV infection was detectable in autologous transplants while antigenemia was demonstrated in 3/11 and 6/7 patients with BMT from respectively mismatched related and matched unrelated donors. PMID- 11268324 TI - Treatment of childhood acute lymphoblastic leukemia after the first relapse: curative strategies. AB - BACKGROUND AND OBJECTIVES: Treatment of recurrent childhood acute lymphoblastic leukemia (ALL) has been controversial in the last decade. Conventional intensive chemotherapy (CHEMO) can cure up to 30% of children who have relapsed after ALL: similar results have been obtained with autologous bone marrow transplantation (ABMT), but allogeneic bone marrow transplantation (AlloBMT) seems to be the best therapeutic option. In this review the authors point out the contribution of current strategy in the setting of children with ALL who experience a first relapse and should be offered optimal treatment in order to obtain the best disease-free survival (DFS). The principal objective of this issue is to reach a possible consensus on the more controversial points regarding factors considered strong predictors of the outcome of the relapsed patients, second-line chemotherapy, optimal timing and type of transplantation. EVIDENCE AND INFORMATION SOURCE: Data published in the literature over the last decade concerning early and late relapse in childhood ALL suggest that improvements in cure rates may be achieved by intensification of the relapse treatment both with chemotherapy and with different types of transplantation. An accurate search for Medline data has been performed in order to understand the risk-benefit ratio of aggressive therapy adopted in this setting. STATE OF ART: Modern first-line treatment protocols for childhood ALL have contributed to curing an ever larger number of patients but this strategy could be responsible for creating a more resistant leukemic clone at the time of systemic or extramedullary relapse. This hypothesis emerges from a number of single or multicenter experiences; thus clinical and biological features in relapsed patients are being studied carefully in order to understand which risk-directed second-line therapy should be best adopted. The BFM group classified ALL relapses as "very early", "early", or "late" according to the time from diagnosis to first relapse (i.e. < 18, > 18 and < 30 or more than 30 months) and has shown that about 2/3 of the small fraction of children with late extramedullary relapses or late non T-marrow relapses or early combined non T-relapses can be rescued by chemotherapy; in contrast ALL early relapses or T-immunophenotype ALL relapses can be rescue only by AlloBMT. Since 1990 the AIEOP group adopted BFM-like first-line treatment and experienced similar situations for relapsed patients so that, even in absence of a real common relapse protocol, they went in the same direction as the BFM group as far as hematopoietic stem cell transplantation (HSCT) procedures and decision were concerned. A recent AIEOP study on the destiny of 192 consecutive patients with ALL in 2nd complete remission and not having an HLA suitable sibling donor is presented in this issue. The value of different HSCT procedures is addressed and the protection against a new relapse seems to be real, although, of course, the risk-benefit ratio should always be evaluated. PERSPECTIVES: Very few prospective studies on the treatment of childhood ALL relapse have been set up in the last decade because of many difficulties regarding common second-line therapies, some reluctance versus HSCT in consideration of the transplant-related mortality and the so-called late effects. Additional modifications of allogeneic family and unrelated donor HSCT strategies and the promising results both of cord HSCT and auto-grafting methods including in vitro purging or post-transplant immunotherapy, are making transplantation procedures for ALL relapsed patients more appropriate and increasing confidence in their use. The possibility of performing common prospective international studies should be encouraged over the next years in order to elucidate an area of great research as is that of the treatment of childhood ALL relapse. PMID- 11268327 TI - The central venous catheter in a bone marrow transplant unit: an unresolved problem. AB - Bone marrow transplantation (BMT) is feasible with a bearable risk and discomfort for patients only if good venous access is provided. Therefore a major task for nurses of a BMT unit is management of a patient's central venous catheter. There is not general agreement about the procedure of handling a CVC and infection prophylaxis. We collected data from some Italian BMT and hematology units by means of a questionnaire. The responses to this questionnaire were not comparable except for some particulars. Each center has its own ritual procedure; even the use of sterile gloves while handling the most dangerous connections of the catheter is not the rule everywhere. It is noteworthy that only a minority of physicians are able to handle a catheter correctly. PMID- 11268326 TI - Artificial nutrition and bone marrow transplantation. AB - Parenteral nutrition has a central role in the supportive therapy of patients submitted to a BMT. A central catheter is mandatory for transfusions, antibiotic therapy and a proper nutrition. A good nutritional support contributes to maintain hydration, reduce lean body mass loss, increase patient comfort and improve survival in patients who can not eat or absorb for a prolonged period of time. After a BMT metabolic complications are frequent and require careful monitoring; in critical care patients, the major risks are electrolyte and glucose disturbances. Liver disease is a main metabolic complication of PN, but it can occur in any cancer patient due to therapy or to graft-versus-host disease. Its best prevention requires the avoidance of prolonged enteral fasting. PMID- 11268329 TI - Immunoablation followed or not by hematopoietic stem cells as an intense therapy for severe autoimmune diseases (SADS). New perspectives, new problems. PMID- 11268328 TI - High dose therapy and autologous hematopoietic stem cell transplantation in poor risk solid tumors of childhood. AB - In the last two decades autologous hematopoietic stem cell transplantation (HSCT) has been increasingly used in the treatment of several poor risk solid tumors of childhood. Examples are recurrent or resistant cancers, metastatic presentation at diagnosis, incomplete surgical resection, unfavorable histologic and biological features. Results from the Children's Cancer Group randomized trial confirm the data from retrospective studies which reported the superiority of HSCT over standard chemotherapy for neuroblastoma. Several retrospective analyses support the use of HSCT in Ewing's sarcoma and in some brain tumors. No evidence of utility has been reported for rhabdomyosarcoma. The most widely utilized source of stem cells is peripheral blood, while there are conflicting data regarding the use of total body irradiation and purging of stem cells. PMID- 11268330 TI - Immunoablation followed by autologous hematopoietic stem cell infusion for the treatment of severe autoimmune disease. AB - BACKGROUND AND OBJECTIVES: The aim of this study was to evaluate the tolerability and effectiveness of a non-myeloablative conditioning regimen followed by autologous hematopoietic stem cell infusion for the treatment of severe autoimmune diseases. DESIGN AND METHODS: From 1996 patients with severe autoimmune disease not responsive to conventional immunosuppressive treatment were selected. The patients' blood or marrow cells were harvested after incubation with vincristine and methylprednisolone. Two different immunoablative conditioning regimens were employed. The first used cyclophosphamide (2500 mg/m2 in one day) and antilymphocyte globulin (ALG) (15 vials/m2 in three days) and the second used fludarabine (300 mg/m2 in two courses of 5 days) plus ALG (25 vials/m2 in 5 days). RESULTS: Nineteen patients (14 female, 5 male) with severe autoimmune diseases were treated. Nine had a rheumatologic disorder (5 juvenile chronic arthritis, 1 rheumatoid arthritis, 1 systemic vasculitis, 1 Sjogren's syndrome, 1 Behct's disease), 4 a neurologic disorder (3 multiple sclerosis, 1 myasthenia), 3 a haematologic disease (2 pure red cell aplasia, 1 autoimmune thrombocytopenia), 2 had a gastrointestinal disease (1 Crohn's disease, 1 autoimmune enteropathy) and 1 had a multiple autoimmune disorder. There was no regimen-related toxicity and no opportunistic infections occurred. Ninety percent of the patients improved and/or had a complete remission after the procedure. Fifty percent of the subjects went into complete or partial remission after a median follow-up of 15 months (range 3-25) while 50% relapsed after a median follow-up of 11 months, (range 6-16). The incidence of relapse in the group treated with fludarabine was lower (30%). INTERPRETATION AND CONCLUSIONS: A non myeloablative conditioning regimen was able to induce persistent remission in some patients with severe autoimmune diseases. There was no mortality or morbidity related to the procedure. The extent of remission does, however, remain to be established. PMID- 11268332 TI - Autologous bone marrow transplantation versus alternative drugs in pediatric rheumatic diseases. AB - A minority of children suffering from severe rheumatic diseases are unresponsive to conventional treatments. These patients can now be managed with a variety of immunosuppressive therapies. Methotrexate is considered the first choice disease modifying agent for adult and juvenile rheumatoid arthritis. In patients unresponsive to low doses of methotrexate, medium or high-doses can be useful. Instead of methotrexate, a recently developed immunosuppressive drug, mycophenolate-mofetil, which inhibits T- and B-lymphocyte proliferation, can be used. Another possibility for refractory rheumatic diseases, with no increase in toxicity, is combination therapy, for example methotrexate plus cyclosporine, or methotrexate plus salazopyrine or intravenous pulses of cyclophosphamide and methylprednisone. More recently two distinct inhibitors of tumor necrosis factor (etanercept and infliximab have been used successfully for intractable rheumatic diseases (juvenile idiopathic arthritis, psoriatic arthritis, spondyloarthropathies) but the follow-up is still too short to establish their long-term effectiveness. If all these treatments are unsuccessful, an autologous bone marrow transplantation can be proposed to selected patients. Interesting results have been obtained in pediatric rheumatic diseases such as juvenile idiopathic arthritis, systemic lupus erythematosus and systemic sclerosis. Further studies are required to assess the best procedures able to induce remission with a minimal risk of fatal events. PMID- 11268331 TI - In vitro incubation of bone marrow and peripheral stem cells with vincristine and methylprednisolone: functional T-cell depletion for haploidentical and autologous transplants. AB - A mismatched bone marrow transplantation is feasible only if the donor's marrow lymphocytes are eliminated from the graft. This can be achieved by several methods, but all have the disadvantage of inducing a long-lasting immune deficiency while the risk of graft rejection and leukemic relapse increase. We use a sort of functional T-cell depletion by treating the cells with vincristine and methylprednisolone. This method is surely the cheapest and has allowed us to perform 60 transplants with a tolerable risk of GVHD. The treatment of the donor's lymphocytes has already been demonstrated to be able to block the mixed lymphocyte culture reaction in vitro. In this experiment Th1 and Th2 activities were almost completely blocked without reduction of lymphocyte viability and apoptosis induction. PMID- 11268333 TI - Indications and role of allogeneic bone marrow transplantation in childhood very high risk acute lymphoblastic leukemia in first complete remission. AB - BACKGROUND AND OBJECTIVES: Acute lymphoblastic leukemia (ALL) accounts for approximately one third of all cancers in children and its outcome depends on risk factors at the time of diagnosis. While uniform chemotherapy adopted in multicenter studies provided a constant improvement in cure rates for standard risk patients, the results reached in very high risk patients have been disappointing. The objective of this review is to point out the role of allogenic bone marrow transplantation (alloBMT) in very high risk childhood ALL on the basis of results from the current clinical trials. EVIDENCE AND INFORMATION SOURCE: Data covered by Medline and produced by the authors involved in ongoing international studies cover a vast "scenario" of children with very high risk ALL who underwent allogeneic BMT. STATE OF ART: The author outlines the crucial point of very high risk factors in childhood ALL in order to identify those children who are at risk of early relapse. The main reasons for pursuing alloBMT in this particular category of patients concern poor prognostic factors such as molecular biology markers, structural chromosomal abnormalities and biological factors (poor prednisone response) including resistance to initial induction chemotherapy. AlloBMT in childhood ALL in first complete remission seemed to lead to a promising disease-free survival in this patient population when compared with chemotherapy. The principal biases of the retrospective studies were the variable very high risk eligibility criteria, the different first-line therapies adopted before alloBMT and above all the waiting time to transplant which could have accounted for some advantage to alloBMT patients versus chemotherapy patients. PERSPECTIVES: The author touches upon the preliminary results of an ongoing international prospective study as an example of reaching a consensus in the controversial treatment of childhood very high risk ALL. This attempt should provide more information regarding the role of alloBMT in this setting and should cover an area of particular interest. PMID- 11268334 TI - Does stress make you sick and belief make you well? The science connecting body and mind. PMID- 11268335 TI - Hypothalamic mechanisms of pain modulatory actions of cytokines and prostaglandin E2. AB - A decrease and subsequent increase in nociceptive threshold in the whole body are clinical symptoms frequently observed during the course of acute systemic infection. These biphasic changes in nociceptive reactivity are brought about by central signal substances induced by peripheral inflammatory messages. Systemic administration of lipopolysaccharide (LPS) or interleukin-1 beta (IL-1 beta), an experimental model of acute infection, may mimic the biphasic changes in nociception, hyperalgesia at small doses of LPS, and IL-1 beta and analgesia at larger doses. Our behavioral and electrophysiological studies have revealed that IL-1 beta in the brain induces hyperalgesia through the actions of prostaglandin E2 (PGE2) on EP3 receptors in the preoptic area and its neighboring basal forebrain, whereas the IL-1 beta-induced analgesia is produced by the actions of PGE2 on EP1 receptors in the ventromedial hypothalamus. An intravenous injection of LPS (10-100 micrograms/kg) produced hyperalgesia only during the period before fever develops and was abolished by microinjection of NS-398 (an inhibitor of cyclooxygenase 2) into the preoptic area, but not into the other areas in the hypothalamus. The hyperalgesia induced by the cytokines PGE2 and LPS may explain the systemic hyperalgesia clinically observed in the early phase of infectious diseases, which probably warns the organisms of infection before the full development of sickness symptoms. The switching of nociception from hyperalgesia to analgesia accompanied by sickness symptoms may reflect changes in the host's strategy for fighting microbial invasion as the disease progresses. PMID- 11268336 TI - Molecular mechanisms of fever and endogenous antipyresis. AB - This review summarizes recent studies on endogenous antipyretic mechanisms. Fever is the result of a balance between pyrogenic and cryogenic cytokines and hormones. Although there is considerable evidence that fever evolved as a host defense response, it is important that the rise in body temperature not be too high. Many endogenous cryogens or antipyretics that limit the rise in body temperature have been identified during the last 25 years. These include alpha MSH, arginine vasopressin, glucocorticoids, TNF (under certain circumstances), and IL-10. Most recently, evidence has accumulated that cytochrome P-450 (P-450), part of the alternative pathway for arachidonic acid metabolism, plays an important role in reduction of fever and inflammation. Supporting a role for P 450 in endogenous antipyresis and antiinflammation includes evidence that (1) inducers of P-450 reduce fever, (2) inhibitors of P-450 cause a larger fever, (3) and P-450 arachidonic acid metabolites reduce fever. PMID- 11268337 TI - CXC chemokine receptors expression during chronic relapsing experimental autoimmune encephalomyelitis. AB - Chemokines are small proinflammatory cytokines that possess the ability to stimulate migration of inflammatory cells towards the tissue site of inflammation. Previous reports showed that several chemokines may be involved in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), an animal model of autoimmune central nervous system (CNS) inflammation. Inflammatory cells respond to chemotactic chemokine gradient through the chemokine receptors (ChRs). The goal of this study was to analyze expression of ChRs belonging to CXC subfamily during different stages of chronic relapsing EAE. We found significantly increased expression of CXCR2 and CXCR4 in the spinal cord during the first and second disease attacks. The kinetics of this expression in CNS and blood suggests that CXCR2 is expressed by leukocytes migrating from the blood, but CXCR4 is expressed mainly by CNS parenchymal cells. Those results support the interpretation that chemokine-chemokine receptor interactions may play an important role in the development of CNS autoimmune inflammation. PMID- 11268338 TI - Contribution of differently localized alpha 2- and beta-adrenoceptors in the modulation of TNF-alpha and IL-10 production in endotoxemic mice. AB - Evidence is presented that the immune response to endotoxemia is under tonic control of the sympathetic nervous system. Adrenergic agents may influence the immune response both directly through alpha- and beta-adrenergic receptors expressed by immunologically competent cells and indirectly via alteration of the endogenous NA level by influencing the activity of release-regulating presynaptic alpha 2-adrenoceptors located on the sympathetic nerve terminals. In the immunomodulatory effect of NA/adrenergic drugs, their action on beta adrenoceptors was dominant, but the considerable role of alpha-adrenoceptors on macrophages was also demonstrated. According to our findings, regulation of the ascending wing of the inflammatory response, that is, TNF-alpha production, is more sensitive to the adrenoceptor effect, whereas modulation of its deregulation by IL-10 production also involves some other determining factors. PMID- 11268339 TI - Regulation of cytokine secretion and amyloid precursor protein processing by proinflammatory amyloid beta (A beta). AB - Neurodegenerative processes in Alzheimer's disease (AD) are thought to be driven, in part, by the deposition of amyloid beta (A beta), a 39-43-aminoacid peptide product resulting from an alternative cleavage of amyloid precursor protein (APP). In addition to its neurotoxic properties, A beta may influence neuropathology by stimulating glial cell cytokine and acute phase protein secretion in affected areas of the brain (e.g., cortex, hippocampus). Using an in vitro human astrocyte model (U-373 MG astrocytoma cells), the effects of A beta treatment on acute phase protein (APP and alpha-1-antichymotrypsin [alpha 1-ACT]) and interleukin-8 (IL-8) were examined. U-373 MG cells secreted increased levels of alpha 1-ACT and neurotrophic/neuroprotective alpha-cleaved APP (alpha APP) after exposure to interleukin-1 beta (IL-1 beta) for 24 hours. A beta treatment resulted in a similar, but modest increase in alpha 1-ACT secretion, a two- to threefold stimulation of IL-8 production, and, conversely, a profound reduction in the levels of secreted alpha APPs. A beta inhibited alpha APP secretion by U 373 MG cells in a concentration- and conformation-dependent manner. Moreover, the reduction in alpha APP secretion was accompanied by an increase in cell associated APP. Another proinflammatory amyloidogenic peptide, human amylin, similarly affected APP processing in U-373 astrocytoma cells. These data suggest that A beta may contribute to Alzheimer's-associated neuropathology by lowering the production of neuroprotective/neurotrophic alpha APPs. Moreover, the concomitant increase in cell-associated APP may provide increased substrate for the generation of amyloidogenic peptides within astrocytes. PMID- 11268340 TI - Increased sensitivity of the baroreceptor reflex after bacterial endotoxin. AB - Lipopolysaccharide (LPS), an endotoxin that elicits the production of several cytokines, induces cardiovascular changes characterized by increased perfusion of immune organs and compensatory sympathetic vasoconstriction in other tissues. We therefore hypothesized that to adapt to altered blood flow distribution following LPS administration, changes in the sensitivity of reflexes that control blood pressure would occur. Our data show that the sensitivity of the baroreceptor reflex increases significantly two and three hours after the intravenous administration of a subpyrogenic dose of the endotoxin. This change in sensitivity that could occur at peripheral or central levels may underlie necessary adjustments of cardiovascular mechanisms during the course of certain immune responses. PMID- 11268342 TI - Mechanisms of behavioral and neuroendocrine effects of interleukin-1 in mice. AB - Interleukin-1 beta is a key molecule in brain-immune interactions that, apart from its immune effects, stimulates the hypothalamo-pituitary-adrenal (HPA) axis and induces behavioral alterations. However, its physiological role during stress responses remain to be elucidated. The possible mechanisms involved in IL-1 mediated stimulation of the HPA axis during stress were assessed by using different approaches. They were first studied in mice deficient for the IL-1 beta converting enzyme (ICE) gene. Mature IL-1 beta derives from a precursor, the pro IL-1 beta, devoid of any conventional signal sequence that is mainly processed by ICE. After immune or stress stimulation, ICE-deficient mice were shown to have a hyperactive HPA axis and to able to produce immunoreactive IL-1 beta. This indicates that the greater reactivity of the HPA axis could result from a higher sensitivity to non-ICE-matured IL-1 beta, as suggested by a higher basal transcription of hypothalamic IL-1 receptor type I (IL-1 RI) in ICE-deficient mice. The biological effects of IL-1 beta are mediated by IL-1 RI associated with the IL-1 receptor accessory protein (IL-1RAcP). IL-1RAcP is an essential component for IL-1 action at the periphery, but its role in the brain is not well known. Therefore, the effects of i.c.v. IL-1 beta were studied in IL-1RAcP deficient mice. In normal mice, i.c.v. IL-1 beta depresses peripheral immune responses, induces the production of plasma IL-6, and stimulates the HPA axis. None of these effects were observed in IL-1RAcP-deficient mice, indicating that IL-1RAcP is necessary for the induction of the main neuroendocrine and immune effects of central IL-1 beta. In normal mice, the role of IL-1 beta was assessed by pretreating the animal with the IL-1 receptor antagonist (IL-1Ra). IL-1Ra did modify the activation of the HPA axis observed during stress, except when the animals were previously sympathectomized. This suggests that the sympathetic nervous system can downregulate the IL-1 beta-induced stimulation of the HPA axis. Finally, the modulation of the production and physiological activities of IL-1 were studied in normal mice, taking advantage of interindividual differences in brain-immune interactions linked to cerebral lateralization. Behavioral/brain lateralization was shown to be related to behavioral response to peripheral administration of IL-1, and to the production of IL-1 and IL-6 in response to LPS. This suggests that cytokines, and especially IL-1 beta, may represent one of the factors responsible for interindividual differences in brain-immune interactions. PMID- 11268341 TI - Not all peripheral immune stimuli that activate the HPA axis induce proinflammatory cytokine gene expression in the hypothalamus. AB - Administration of low doses of lipopolysaccharide (LPS) that do not disrupt the blood-brain barrier (BBB) results in the expression of interleukin-1 beta (IL-1 beta), IL-6, and tumor necrosis factor-alpha (TNF alpha) in the hypothalamus in parallel to stimulation of the hypothalamus-pituitary-adrenal (HPA) axis. This endocrine response is triggered by peripheral cytokines, and we recently obtained evidence that brain-borne IL-1 contributes to its maintenance. LPS preferentially stimulates cells of the macrophage lineage and B lymphocytes. The possibility that primarily stimulation of other types of peripheral immune cells also results in the expression of proinflammatory cytokines in the brain and in the activation of the HPA axis was investigated. Our results showed that, in contrast to LPS, administration of the superantigen staphylococcal enterotoxin B (SEB), which stimulates T cells by binding to appropriate V beta domains of the T-cell receptor, did not result in induction of IL-1 beta, IL-6, and TNF alpha expression in the hypothalamus. Furthermore, although IL-2 transcripts in the spleen were highly increased, expression of this gene was not detected in the brain. However, as with LPS, SEB administration also results in elevated levels of glucocorticoids in blood. Therefore, our data suggest that increased expression of proinflammatory cytokines in the brain is not a necessary step in the stimulation of the HPA axis by SEB. PMID- 11268343 TI - Genetic regulation of nerve avulsion-induced spinal cord inflammation. AB - In the animal model for multiple sclerosis (MS), experimental autoimmune encephalitis (EAE), genetic loci correlating with incidence or severity of disease are located both within and outside of the major histocompatibility complex (MHC). Whereas polymorphisms within MHC class I and II molecules are likely to be a major determinant of MHC gene influence in rat EAE, it is still unclear how non-MHC gene regions influence disease. Genetic control of inflammation can hypothetically be either general or specific for a particular target tissue. For the latter, gene regulation of pathomechanisms in the CNS could affect reactivity of microglia or astrocytes, local cytokine/chemokine production, or even neuronal vulnerability. We have obtained strong support for this notion by observations of rat strain-dependent variation in the inflammatory response after ventral root avulsion, a model in which mainly non-antigen specific elements of the immune system promote inflammation. A comparison of strains with similar MHC haplotypes on different backgrounds and strains with different MHC haplotypes on the same background, respectively, demonstrates that the inflammatory phenotype is regulated mainly by non-MHC genes. Interestingly, different features of the inflammatory response, such as induction of MHC class II expression, glial activation, cytokine expression, and neuronal vulnerability, varied between rat strains and were largely independent of each other. The genetic control of several basic features of inflammation in the CNS is of great relevance not only for MS/EAE, but also for several other neurological conditions with inflammatory components such as cerebrovascular and neurogenerative dieases and trauma. PMID- 11268344 TI - Bidirectional heterologous desensitization of opioid and chemokine receptors. AB - Opioids are known to suppress a number of elements of the immune response, including antimicrobial resistance, antibody production, and delayed-type hypersensitivity. Phagocytic cells may be particularly susceptible to opioid administration, since reduced production of the cytokines IL-1, IL-6 and TNF alpha, monocyte-mediated phagocytosis, and both neutrophil and monocyte chemotaxis have all been well established. Earlier studies have shown that both mu- and delta-opioid agonists induce a chemotactic response in monocytes and neutrophils. In addition, mu- and delta-opioid administration inhibited the chemotactic response of these cell populations to a number of chemokines through a process of heterologous desensitization. We report here that mu-, delta-, and kappa-opioid agonists also induce a chemotactic response in T lymphocytes. Using the human T-cell line Jurkat, we have confirmed previous observations that pre incubation with met-enkephalin (MetEnk), an endogenous opioid agonist, prevents the subsequent chemotactic response to the chemokine RANTES. On the other hand, treatment with MetEnk does not alter the response to the chemokine SDF-1 alpha. Moreover, we found that pre-treatment with RANTES prevented a subsequent response of monocytes to the mu-opioid agonist DAMGO. These results suggest that activation of members of the opioid and chemokine receptor families leads to downregulation of each other's leukocyte migratory activities. PMID- 11268345 TI - C-fos and IL-2 gene expression in rat brain cells and splenic lymphocytes after nonantigenic and antigenic stimuli. AB - Immunostimulatory or immunosuppressive stress models were used: (1) rotation stress (RS) and (2) immobilization (restraint) stress (IS). Intravenous injection of tetanus toxoid (anatoxin) (TT) was chosen as the antigenic stimulus (500 micrograms/kg weight), and intravenous injection of saline solution was used as the control. Splenic lymphocytes (CBA mice) or different brain structures (Wistar and Sprague-Dawley rats) were analyzed. The c-fos and interleukin-2 (IL-2) mRNA expression was measured using a digoxigenin (Dig)-labeled cDNA probe by spot or in situ hybridization. Rotation stress stimulated IL-2 mRNA synthesis in lymphocytes in the presence of ConA and rIL-2 by 40%. IL-2 mRNA synthesis in lymphoid cells obtained from animals after IS and after IS in combination with the administration in vitro of the cytotoxic drug CsA to the splenic lymphocytes was inhibited (30% and 99%), accordingly, as compared with control rats. Induction of c-fos mRNA synthesis in rat brain cells was noted 30 minutes after RS in the hypothalamus (lateralis hypothalamic area, LHA), thalamus, corpus collosum, and sensorimotor zone of the brain cortex. IL-2 mRNA synthesis was shown two hours after RS in the same structures. The increased number of c-fos mRNA-positive cells two hours after TT injection was shown in the posterior hypothalamus area (PHA), LHA, dorsomedial nucleus (DMH), ventromedial nucleus (VMH), and anterior hypothalamus area (AHA) as compared to the effect of i.v. saline injection. Moreover, IL-2 mRNA-positive cell induction was noted in the PHA, DMH, and VMH. Six hours after TT injection, c-fos mRNA expression was decreased in the PHA, LHA, and AHA. Activation of c-fos and IL-2 mRNA was detected in the paraventricularis nucleus 6 hours after TT i.v. injection. Thus, inhibition or stimulation of IL-2 gene expression in lymphoid cells depends on the nature of the stressors. RS or antigenic stimuli induce c-fos and IL-2 gene expression in definite structures of the brain. The dynamics of this process are time dependent. The partial correlation between c-fos and IL-2 mRNA expression in localization in brain structures and time dependence was shown. PMID- 11268346 TI - Tumor necrosis factor-alpha induces neuronal death by silencing survival signals generated by the type I insulin-like growth factor receptor. AB - Within the central nervous system, the proinflammatory cytokine tumor necrosis factor (TNF)-alpha is best characterized by its ability to directly foment signals of death. However, recent evidence suggests that TNF-alpha also promotes neurodegeneration through inhibition of a vital survival signal, insulin-like growth factor-I (IGF-I). By inhibiting essential components of the IGF-I survival response, such as phosphatidylinositol 3'-kinase (PI 3-kinase), low nontoxic concentrations of TNF-alpha indirectly trigger the death of neurons. We suggest that this inhibition of survival signaling is a pathophysiologically relevant action of TNF-alpha in the brain. This type of cross-talk by which vastly different receptors utilize shared intracellular substrates is potentially applicable to a broad number of receptors that are coexpressed on the same cell. The use of neuronal growth factors in the treatment of neurodegenerative diseases, such as cerebral ischemia and the AIDS dementia complex, may prove much more effective if the elevated expression of TNF-alpha in these disorders is neutralized. PMID- 11268347 TI - The neuroimmunomodulatory peptide alpha-MSH. AB - Alpha-melanocyte-stimulating hormone (alpha-MSH), a neuroimmunomodulatory peptide of ancient origin, is known to be involved in the control of host responses. In inflammatory cells, in the periphery and within the central nervous system, alpha MSH modulates the production and action of proinflammatory cytokines. This broad influence occurs via endogenous alpha-MSH (melanocortin) receptors. The key to this anti-inflammatory influence is inhibition of NF-kappa B. Indeed alpha-MSH inhibits activation of this nuclear factor through preservation of I kappa B alpha, which binds to NF-kappa B and prevents its migration to the nucleus. Cells transfected with alpha-MSH plasmid vector are resistant to challenge with bacterial lipopolysaccharide. The peptide also act on central melanocortin receptors to modulate inflammation in the periphery. In brief, alpha-MSH and certain of its fragments such as alpha-MSH [11-13] KPV modulate inflammation via three general actions: direct actions on peripheral host cells; actions on inflammatory cells within the brain to modulate local reactions; and descending neural anti-inflammatory pathways that control inflammation in peripheral tissues. PMID- 11268348 TI - The neuropeptide alpha-MSH in host defense. AB - The presence of the ancient peptide alpha-melanocyte-stimulating hormone (alpha MSH) in barrier organs such as gut and skin suggests that this potent anti inflammatory molecule may be a component of the innate host defense. In tests of antimicrobial activities, alpha-MSH and its fragment KPV showed inhibitory influences against the gram-positive bacterium Staphylococcus aureus and the yeast Candida albicans. Anti-tumor necrosis factor and antimicrobial effects of alpha-MSH suggest that the peptide might likewise reduce replication of human immunodeficiency virus (HIV). Treatment with alpha-MSH reduced HIV replication in chronically and acutely infected human monocytes. At the molecular level, alpha MSH inhibited activation of the transcription factor NF-kappa B known to enhance HIV expression. alpha-MSH that combines antipyretic, anti-inflammatory, and antimicrobial effects could be useful in the treatment of disorders in which infection and inflammation coexist. PMID- 11268349 TI - The role of alpha-MSH as a modulator of cutaneous inflammation. AB - Among various neuropeptides such as substance P, calcitonin gene-related peptide and others, alpha-melanocyte-stimulating hormone (alpha-MSH) was found to be produced in the skin. Moreover, melanocortin receptor 1 (MC-1R), which is specific for alpha-MSH and ACTH, is expressed in the skin on keratinocytes, dendritic cells, macrophages and endothelial cells. In monocytes, macrophages and dendritic cells alpha-MSH inhibits the production and activity of immunoregulatory and proinflammatory cytokines such as IL-2, IFN-gamma, TNF-alpha and IL-1. It downregulates the expression of costimulatory molecules such as CD86 and CD40 and induces the production of suppressor factors such as the cytokine synthesis inhibitory factor IL-10. On endothelial cells alpha-MSH is capable of downregulating the LPS-induced expression of adhesion molecules such as vascular cell adhesion molecule (VCAM) and E-selectin. Moreover, the LPS-induced activation of transcription factors such as NF kappa B is downregulated by alpha MSH. In a mouse model i.v. or topical application of alpha-MSH was found to inhibit the induction phase as well as the effector phase of contact hypersensitivity (CHS) reactions and to induce hapten-specific tolerance. These findings indicate that the production of immunosuppressing neuropeptides such as alpha-MSH by epidermal cells may play an essential role during the pathogenesis of immune and inflammatory reactions in the skin. PMID- 11268350 TI - Neuropeptide regulation of immunity. The immunosuppressive activity of alpha melanocyte-stimulating hormone (alpha-MSH). AB - The ocular microenvironment is an extreme example of regional immunity. Within its microenvironment, expression of delayed type hypersensitivity (DTH) is suppressed. This immunosuppression is mediated in part by the constitutive expression of alpha-MSH. Previously we have found that alpha-MSH suppresses the production of IFN-gamma by activated effector T cells. Recently we have found that alpha-MSH can mediate induction of TGF-beta-producing T cells that act as regulatory T cells. This has encouraged us to further examine the potential for alpha-MSH to suppress T cell-mediated inflammation (autoimmune disease) and to regulate lymphokine production by effector T cells. When alpha-MSH was injected i.v. into mice at the time of peak retinal inflammation, the severity of experimental autoimmune uveitis (EAU) was significantly suppressed. Effector T cells activated in vitro in the presence of alpha-MSH proliferated and produced IL-4 and enhanced levels of TGF-beta while their IFN-gamma and IL-10 production was suppressed. The alpha-MSH-treated T cells functioned as regulatory T cells by suppressing in vitro IFN-gamma production by other inflammatory T cells. This regulatory activity was the function of alpha-MSH-treated CD4+ CD25+ T cells. Therefore, alpha-MSH mediates immunosuppression by inducing a differential expression of lymphokine production and by inducing activation of regulatory functions in T cells. This implies that alpha-MSH may take part in regional mechanisms of immunosuppression and possibly peripheral tolerance. Thus, alpha MSH can be used to suppress autoimmune disease and possibly reestablish tolerance to autoantigens. PMID- 11268351 TI - Natural immunity and neuroimmune host defense. AB - Innate resistance is mediated by non-immune defense and by natural immunity. Non immune defense includes diverse mechanisms (e.g., physico-chemical defense by bile acids). Natural killer (NK) cells, gamma delta T lymphocytes and CD5+ B lymphocytes are key mediators of natural immunity. These cells utilize germ-line coded receptors that recognize highly conserved, homologous epitopes (homotopes). Typically, it is not the antigen, but cytokines and hormones that regulate the level of NK-mediated cytotoxicity. These include interleukin-2, interferons, prolactin and growth hormone. Less is known about gamma delta T lymphocytes. CD5+ B lymphocytes produce germ-line coded antibodies (predominantly IgM) that are polyspecific, and able to recognize a great variety of microorganisms, cancer cells and self-components. Antigen is not an effective stimulus for natural antibody (NAb), but bacterial lipopolysaccharide (LPS) is. During the acute phase response (febrile illness) the T-cell-regulated adaptive immune response is switched off and natural immune mechanisms are amplified several hundred to a thousand times within 24-48 hours (immunoconversion). This immunoconversion is initiated by immune-derived cytokines, and involves profound neuroendocrine and metabolic changes, all in the interest of host defense. Immune recognition is assured by natural antibodies and by some liver-derived acute phase proteins, such as C-reactive protein or endotoxin-binding protein, the level of which is elevated in the serum. Thus, natural immunity is essential for a first and last line of defense and the neuroendocrine system is an important promoter of this activity. PMID- 11268352 TI - Control of salivary secretion by nitric oxide and its role in neuroimmunomodulation. AB - In many in vivo systems exposure to endotoxins (LPS) leads to the co-induction of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), which is important to the regulation of the function of different systems during infection. In submandibular glands (SMG) neural (n)NOS is localized in neural terminals and in striated, granular convoluted and excretory ducts, endothelial (e)NOS in vascular endothelium and ducts, and iNOS in macrophages and in tubules and ducts. In normal adult male rats, injection of an inhibitor of NOS decreased the stimulated salivary secretion and a donor of NO potentiated it, indicating that NO exerts a stimulatory role. A single high dose of LPS (5 mg/kg, i.p.) induced an increase in NOS activity measured by the 14C-citrulline method, increased PGE content almost 100% as measured by RIA, and blocked stimulated salivary secretion. The administration of a specific iNOS inhibitor, aminoguanidine (AG), with LPS not only decreased NOS activity but significantly decreased PGE content, indicating that NO triggered the activation of COX-2. LPS increased conversion of labeled arachidonate to prostaglandins (PGs) showing that COX was induced. Since a PGE1 analogue blocked stimulated salivation, the LPS induced inhibition of salivation is probably due to release of PGs. Therefore, the use of inhibitors of iNOS and COX-2 could be very useful to increase salivation during infection since saliva has antimicrobial actions. PMID- 11268353 TI - Nerve growth factor and neuroimmune interactions in inflammatory diseases. AB - Discovered almost 50 years ago, nerve growth factor (NGF) has been extensively studied in various biological systems. NGF has recently been suggested to play an important role in mediating and/or regulating immune response, in addition to its trophic and tropic effects on nerve growth and regeneration It is clear that in complex interactions between immune cells and nervous system NGF plays a central role. We have only just begun to identify and understand the direct mechanisms by which NGF activates target cells, the precise identity of the target cells, and the particular factors released from target cells. Nerve growth factor together with possibly other neurotrophins such as BDNF (brain-derived nerve growth factor), GDNF (glial-derived nerve growth factor) or NT3 are important modulators of immunity. More detailed studies are needed at the receptor, mediator and cellular levels to better understand the neuroimmunomodulatory properties of neurothrophins and NGF. The nature of the involvement of NGF in inflammation and inflammatory diseases remains a particularly interesting question. By blocking NGF or mediators released upon NGF activation, we are able to control the progress of inflammation, thereby opening many therapeutic opportunities for the future. PMID- 11268354 TI - Nuclear factor-kappa B is involved in the catecholaminergic suppression of immunocompetent cells. AB - Catecholamines are known to exert a powerful impact on the immune system by downregulation of proliferation and differentiation, and induction of apoptosis. However, the mechanism for this regulatory route is still unclear. Therefore well established human monocytic cell-lines and nontransformed human monocytes, obtained from peripheral blood, were incubated with an optimal concentration of LPS and varying concentrations of the catecholamine dopamine. The proliferative response to LPS was determined by [3H]thymidine incorporation, and a significant suppressive effect by dopamine was obtained. LPS-induced binding of NF-kappa B to DNA, determined by electrophoretic mobility shift assay, was inhibited by extrinsic dopamine, leading to a decreased proliferation and cytokine expression. In contrast, the intracellular ceramide concentration was not affected by incubation of peripheral blood lymphocytes with dopamine. Our findings suggest that the NF-kappa B-I-kappa B transcription machinery may well be involved in the catecholaminergic regulation of the immune system, while the ceramide-SAPK/JNK cascade appears not to play a significant role in this suppression. PMID- 11268355 TI - Neurohormones and catecholamines as functional components of the bone marrow microenvironment. AB - A variety of cytokines and growth factors exert a finely tuned control on the complex series of proliferative and differentiative events called hematopoiesis. Recent studies have shown that neuroendocrine and neural factors may also regulate hematopoiesis. In particular, besides its important immunoenhancing properties, the pineal neurohormone melatonin can also rescue hematopoiesis from the toxic effect of anti-cancer drugs via the action of T-helper cell novel opioid cytokines. In turn, these substances bind kappa-opioid receptors expressed in GM-CSF-activated macrophage-like stromal cells and seem to stimulate IL-1. Adrenergic agents can also affect hematopoiesis. We demonstrated that pre-B cells express alpha 1B-adrenoceptors (alpha 1B-AR) and that their activation by catecholamines results in suppressed myelopoiesis in vitro or protection in vivo against supralethal doses of carboplatin. Most recently, we found that alpha 1B AR gene knockout mice show a deranged hematopoietic recovery after sublethal irradiation. Regeneration of pre-B cells (the cell type expressing alpha 1B-AR) and of erythrocytes was much faster in knockout than in wild-type mice. Most interesting, bone marrow cells can synthesize both melatonin and catecholamines. As far as melatonin is concerned, human and murine bone marrow cells contain and synthesize melatonin at a concentration that is three orders of magnitude higher than that normally found in serum. Catecholamines are also present in substantial amounts and originate both from nerve endings and bone marrow cells. These findings open interesting new perspectives and include hematology among the disciplines that would benefit from the integrative NIM approach. PMID- 11268356 TI - The peripheral CRH/urocortin system. AB - The hypothalmus-pituitary-adrenal (HPA) axis and the immune system communicate at multiple levels: On the one hand, immune system-derived substances, such as interleukin-1, interleukin-6, tumor necrosis factor alpha, and leukemia inhibitory factor can stimulate the HPA axis. On the other hand, HPA axis-derived substances, most importantly glucocorticoids, can modulate the immune response. Furthermore, factors that were originally thought to be restricted to the HPA axis have been found to be expressed by immune cells. Proteins belonging to the CRH (corticotropin-releasing hormone) family represent important examples of such hormones. In the early 1990s, it was shown that immunoreactive CRH was present at sites of chemically induced inflammation. Administration of anti-CRH antibodies reduced the degree of inflammation, pointing to a pro-inflammatory role of "peripheral" CRH. We and others could show that lymphocytes are one source of immunoreactive CRH; however, the antiserum used in our study as well as in previous reports crossreacted with urocortin, a newly discovered member of the CRH family. Using RT-PCR, we could clearly demonstrate that human lymphocytes expressed urocortin but not CRH mRNA. These results were confirmed by immunocytochemistry, employing urocortin- and CRH-specific antibodies, respectively. The possible functional roles of urocortin expression in the immune system are discussed. PMID- 11268357 TI - Neural control of ocular immune privilege. AB - Ocular immune privilege arises from interactions between the immune apparatus and the eye itself, thereby providing immune protection for the eye that is devoid of sight-threatening inflammation. On the one hand, antigens injected intraocularly elicit deviant systemic immune responses that are devoid of immunogenic inflammation (Anterior Chamber-Associated Immune Deviation, ACAID). On the other hand, the ocular microenvironment (aqueous humor, secreted by cells that surround this chamber) suppresses intraocular expression of immunogenic inflammation. Several lines of evidence indicate that ocular immune privilege is under neural control. First, aqueous humor contains neuropeptides (alpha-MSH, VIP, CGRP) that inhibit and alter the functional properties of T lymphocytes and macrophages. Second, when corneal nerves are severed, the tissues surrounding the anterior chamber cease secreting immunosuppressive factors and ACAID fails--until the nerves regrow. Third, light deprivation abolishes the capacity of the anterior chamber to support ACAID induction, a process that is sensitive to neuropeptides and melatonin. The photoreceptor(s) responsible for ACAID are connected to the nervous system and may reside in the anterior segment and/or the retina. Thus, neural elements from the central nervous system and within the eye help to shape both the induction and the expression of ocular immunity, thereby promoting immune privilege. PMID- 11268358 TI - Nerve-driven immunity. The direct effects of neurotransmitters on T-cell function. AB - We carried out studies to explore whether neurotransmitters can directly interact with their T-cell-expressed receptors, leading to either activation or suppression of various T-cell functions. Human and mouse T cells were thus exposed directly to neurotransmitters in the absence of any additional molecule, and various functions were studied, among them cytokine secretion, proliferation, and integrin-mediated adhesion and migration. In this review, I describe the effects of four neuropeptides: somatostatin (SOM), calcitonin-gene-related peptide (CGRP), neuropeptide Y (NPY), and substance P (Sub P), and one non peptidergic neurotransmitter--dopamine. We found that SOM, NPY, CGRP, and dopamine interact directly with T cells, leading to the activation of beta 1 integrins and to the subsequent integrin-mediated T-cell adhesion to a component of the extracellular matrix. In contrast, Sub P had a reverse effect--full blockage of integrin-mediated T-cell adhesion triggered by a variety of signals. Each of these neurotransmitters exerted its effect through direct interaction with its specific receptor on the T-cell surface, since the effect was fully blocked by the respective receptor-antagonist. Taken together, this set of findings indicates that neurotransmitters can directly interact with T cells and provide them with either positive (integrin-activating, pro-adhesive) or negative (integrin-inhibiting, anti-adhesive) signals. We further found that the above neurotransmitters, by direct interaction with their specific receptors, drove T cells (of the Th0, Th1, and Th2 phenotypes) into the secretion of both typical and atypical ("forbidden") cytokines. These results suggested that neurotransmitters can substantially affect various cytokine-dependent T-cell activities. As a whole, our studies suggest an important and yet unrecognized role for neurotransmitters in directly dictating or modulating numerous T-cell functions under physiological and pathological conditions. PMID- 11268359 TI - Cytokines and neurotrophins interact in normal and diseased states. AB - Neurotrophins (NTs) such as nerve growth factor (NGF) as well as cytokines, for example, interleukin-6 (IL-6), are communicators between the nervous and immune systems. There is evidence for mutual interactions between NTs and cytokines. Strategies are being developed to elucidate the molecular mechanism/s of interactions and to understand how cytokines are involved in health and disease. Analysis of underlying signaling pathways in glial cells indicates that different transcription factors, such as NF-kappa B, cAMP-responsive-element binding protein (CREB), and activator protein 1 (AP-1), are involved in NT induction. IL 6 and NTs of the NGF family are coexpressed at sites of nerve injury. Interactions of these factors could modulate both neuronal de- and regeneration: IL-6 in conjunction with its soluble IL-6 receptor induces a specific pattern of NTs in astrocytes in defined brain regions. This indicates that the IL-6 system mediates a local supply of NTs that participate in diverse CNS functions, such as protection of neurons from insults, neuronal survival, and neuroimmune responses. PMID- 11268360 TI - Overproduction of IFN-gamma and TNF-alpha from natural killer (NK) cells is associated with abnormal NK reactivity and cognitive derangement in Alzheimer's disease. AB - Alterations of natural killer (NK) function can be involved in the neuroimmune mechanism of neurodegeneration in dementia of the Alzheimer's type (DAT). NK cell cytotoxicity (NKCC) and the generation and release of IFN-gamma and TNF-alpha (spontaneous and modulated by IL-2) from pure NK cells (CD 16+, CD 56+, CD 3-) were studied together with circulating IFN-gamma and TNF-alpha levels and cognitive function in 22 old patients with DAT and 15 healthy old subjects. Higher (p < 0.001) IL-2 modulated NKCC (with IL-2 50 U/mL and 100 U/mL) was demonstrated in DAT patients (+35% and +99% from baseline) than in healthy subjects (+6% and +76% from baseline). Increased spontaneous and IL-2-induced release of IFN-gamma and TNF-alpha from NK cells were found in DAT patients compared to healthy subjects (p < 0.001), whereas no difference of serum IFN gamma and TNF-alpha was demonstrated between DAT and control groups. Significant negative correlations among the spontaneous release of IFN-gamma and TNF-alpha from NK and the decrease of the score of cognitive function (MMSE) were found in patients with DAT. In conclusion, alterations of NKCC control and NK-derived cytokine release in DAT could be involved in the neuroinflammatory mechanism related to the progression of neurodegeneration and dementia. PMID- 11268361 TI - Beneficial autoimmune T cells and posttraumatic neuroprotection. AB - Injuries of the central nervous system (CNS) lead to an inevitable and irreversible loss of function because of the lack of neurogenesis, poor regeneration, and the spread of degeneration. In most tissues, protection and repair are the function of the immune system. It has long been thought that this does not apply to the CNS, where--because of its immune-privileged character--any immune activity was assumed to be detrimental. We have recently proposed, however, that provided care is taken to avoid the attendant risks, both repair and protection of injured CNS neurons can benefit from immune intervention. In the following I will summarize the data that led to this concept and describe the evidence supporting it. PMID- 11268362 TI - Feedsidewards: intermodulation (strictly) among time structures, chronomes, in and around us, and cosmo-vasculo-neuroimmunity. About ten-yearly changes: what Galileo missed and Schwabe found. AB - The spectrum of biological rhythms is extended far beyond circadians, circannuals, and ultradians, such as 1.5-hourly melatonin and 8-hourly endothelin 1 (ET-1) rhythms by statistics of natality, growth, morbidity, and mortality, some covering decades or centuries on millions of individuals. These reveal infradian cycles to be aligned with half-weekly rhythms in ET-1, weekly and half yearly ones in melatonin, and even longer--about 50-, about 20-, and about 10 year cycles found in birth statistics. About daily, weekly, yearly, and ten yearly patterns are also found in mortality from myocardial infarctions; the 10 yearly ones are also in heart rate and its variability; in steroid excretion, an aspect of resistance, for example, to bacteria; and in the genetic changes of the bacteria themselves. Automatic physiological measurements cover years and, in one case, cover a decade; the latter reveal an about 10-year (circadecennial) cycle. ECGs, covering months beat-to-beat, reveal circaseptans, gaining prominence in response to magnetic storms or after coronary artery bypass grafting. A spectrum including cycles from fractions of 1 Hz to circasemicentennians is just one element in biological time structures, chronomes. Chaos, trends, and any unresolved variability are the second to fourth elements of chronomes. Intermodulations, feedsidewards, account for rhythmically and thus predictably recurring quantitive differences and even for opposite treatment effects of the same total dose(s) of (1) immunomodulators inhibiting or stimulating DNA labeling of bone in health or speeding up versus slowing down a malignant growth and thus shortening or lengthening survival time, or (2) raising or lowering blood pressure or heart rate in the vascular aspect of the body's defense. Latitude dependent competing photic and nonphotic solar effects upon the pineal are gauged by alternating yearly (by daylight) and half-yearly (by night) signatures of circulating melatonin at middle latitudes and by half-yearly signatures at noon near the pole. These many (including novel near 10-yearly) changes, for example, in 17-ketosteroid excretion, heart rate, heart rate variability, and myocardial infarction in us and those galactic, solar, and geophysical ones around us have their own special signatures and contribute to a cosmo-vasculo-immunity and, if that fails, to a cosmo(immuno?) pathology. PMID- 11268363 TI - Melatonin and its relation to the immune system and inflammation. AB - Melatonin (N-acetyl-5-methoxytryptamine) was initially thought to be produced exclusively in the pineal gland. Subsequently its synthesis was demonstrated in other organs, for example, the retinas, and very high concentrations of melatonin are found at other sites, for example, bone marrow cells and bile. The origin of the high level of melatonin in these locations has not been definitively established, but it is likely not exclusively of pineal origin. Melatonin has been shown to possess anti-inflammatory effects, among a number of actions. Melatonin reduces tissue destruction during inflammatory reactions by a number of means. Thus melatonin, by virtue of its ability to directly scavenge toxic free radicals, reduces macromolecular damage in all organs. The free radicals and reactive oxygen and nitrogen species known to be scavenged by melatonin include the highly toxic hydroxyl radical (.OH), peroxynitrite anion (ONOO-), and hypochlorous acid (HOCl), among others. These agents all contribute to the inflammatory response and associated tissue destruction. Additionally, melatonin has other means to lower the damage resulting from inflammation. Thus, it prevents the translocation of nuclear factor-kappa B (NF-kappa B) to the nucleus and its binding to DNA, thereby reducing the upregulation of a variety of proinflammatory cytokines, for example, interleukins and tumor neurosis factor alpha. Finally, there is indirect evidence that melatonin inhibits the production of adhesion molecules that promote the sticking of leukocytes to endothelial cells. By this means melatonin attenuates transendothelial cell migration and edema, which contribute to tissue damage. PMID- 11268364 TI - The stress response and immune function: clinical implications. The 1999 Novera H. Spector Lecture. PMID- 11268365 TI - Gene regulation by melatonin. AB - The physiological and neuroendocrine functions of the pineal gland hormone, melatonin, and its therapeutic potential critically depend on the understanding of its target sites and its mechanisms of action. This has progressed considerably in the last few years through the cloning of G protein-coupled seven transmembrane melatonin receptors (Mel1a and Mel1b) as well as of nuclear receptors (RZR/ROR alpha and RZR beta) that are associated with melatonin signaling. The transcription factor RZR/ROR alpha appears to mediate a direct gene regulatory action of the hormone, and specific binding sites have been identified in promoter regions of a variety of genes, such as 5-lipoxygenase (5 LO), p21WAF1/CIP1, and bone sialoprotein (BSP). The membrane signaling pathway clearly shows higher ligand sensitivity than the nuclear signaling pathway, but details of its signal transduction cascade, and target genes are presently unknown. Membrane melatonin receptors are expressed mainly in the central nervous system, whereas RZR/ROR alpha is prominently expressed both in the periphery and the brain. The action of membrane melatonin receptors and their specific agonists have been associated with circadian rhythmicity, whereas direct effects of melatonin in the periphery, such as immunomodulation, cellular growth, and bone differentiation, mainly appear to be mediated by RZR/ROR alpha. It is hypothesized in this review that, in some cases, RZR/ROR alpha may be a primary target of membrane melatonin receptors. PMID- 11268366 TI - Involvement of nuclear receptors in the enhanced IL-2 production by melatonin in Jurkat cells. AB - This report shows that melatonin enhances IL-2 production by Jurkat cells via a nuclear receptor-mediated mechanism. Jurkat cells express nuclear (RZR alpha, ROR alpha 1, and ROR alpha 2) and membrane (mt1) melatonin receptors, and melatonin binds to Jurkat nuclei and membranes with the same affinity described for human peripheral blood mononuclear cells (PBMCs). Melatonin enhances IL-2 production by Jurkat cells activated by either phytohemagglutinin (PHA) or phorbol myristate acetate (PMA). PHA activation of Jurkat cells does not change the profile of melatonin receptor expression; on the contrary, PMA activation negatively regulates the mt1 receptor. In the absence of the membrane receptor, melatonin still activates the IL-2 production. These results show that the expression of the nuclear melatonin receptor is sufficient for melatonin to activate IL-2 production by Jurkat cells. PMID- 11268367 TI - The mechanism of action of cytokines to control the release of hypothalamic and pituitary hormones in infection. AB - During infection, bacterial and viral products, such as bacterial lipopolysaccharide (LPS), cause the release of cytokines from immune cells. These cytokines can reach the brain by several routes. Furthermore, cytokines, such as interleukin-1 (IL-1), are induced in neurons within the brain by systemic injection of LPS. These cytokines determine the pattern of hypothalamic-pituitary secretion that characterizes infection. IL-2, by stimulation of cholinergic neurons, activates neural nitric oxide synthase (nNOS). The nitric oxide (NO) released diffuses into corticotropin-releasing hormone (CRH)-secreting neurons and releases CRH. IL-2 also acts in the pituitary to stimulate adrenocorticotropic hormone (ACTH) secretion. On the other hand, IL-1 alpha blocks the NO-induced release of luteinizing hormone-releasing hormone (LHRH) from LHRH neurons, thereby blocking pulsatile LH but not follicle-stimulating hormone (FSH) release and also inhibiting sex behavior that is induced by LHRH. IL-1 alpha and granulocyte macrophage colony-stimulating factor (GMCSF) block the response of the LHRH terminals to NO. The mechanism of action of GMCSF to inhibit LHRH release is as follows. It acts on its receptors on gamma-aminobutyric acid (GABA)ergic neurons to stimulate GABA release. GABA acts on GABAa receptors on the LHRH neuronal terminal to block NOergic stimulation of LHRH release. IL-1 alpha inhibits growth hormone (GH) release by inhibiting GH-releasing hormone (GHRH) release, which is mediated by NO, and stimulating somatostatin release, also mediated by NO. IL-1 alpha-induced stimulation of PRL release is also mediated by intrahypothlamic action of NO, which inhibits release of the PRL inhibiting hormone dopamine. The actions of NO are brought about by its combined activation of guanylate cyclase-liberating cyclic guanosine monophosphate (cGMP) and activation of cyclooxygenase (COX) and lipoxygenase (LOX) with liberation of prostaglandin E2 and leukotrienes, respectively. Thus, NO plays a key role in inducing the changes in release of hypothalamic peptides induced in infection by cytokines. Cytokines, such as IL-1 beta, also act in the anterior pituitary gland, at least in part via induction of inducible NOS. The NO produced inhibits release of ACTH. The adipocyte hormone leptin, a member of the cytokine family, has largely opposite actions to those of the proinflammatory cytokines, stimulating the release of FSHRF and LHRH from the hypothalamus and FSH and LH from the pituitary directly by NO. PMID- 11268368 TI - Melatonin mediates seasonal changes in immune function. AB - Field studies indicate that immune function is compromised and the prevalence of many diseases are elevated during winter when energetic stressors are extensive. Presumably, individuals would enjoy a survival advantage if seasonally recurring stressors could be anticipated and countered by shunting energy reserves to bolster immune function. The primary environmental cue that permits physiological anticipation of season is daily photoperiod, a cue that is mediated by melatonin. However, other environmental factors, including low food availability and ambient temperatures, may interact with photoperiod to affect immune function and disease processes. This paper will review laboratory studies that consistently report enhanced immune function in short day lengths. Prolonged melatonin treatment mimics short days, and both in vitro and in vivo melatonin treatment enhances various aspects of immune function, especially cell-mediated immune function, in nontropical rodents. Reproductive responsiveness to melatonin appears to affect immune function. In sum, melatonin may be part of an integrative system to coordinate reproductive, immunologic, and other physiological processes to cope successfully with energetic stressors during winter. PMID- 11268370 TI - Neuropeptide control of bone marrow neutrophil production. A key axis for neuroimmunomodulation. AB - Nerve fibers project into the bone marrow and terminate in association with stromal cells. Nerve terminals are also associated with antigen-processing and presenting cells throughout the body and have been shown to be important in leukocyte trafficking and wound healing as well as hemopoiesis. Here we show that neuropeptide input to the bone marrow is vital to normal granulopoiesis and that deletion of the neuropeptides, substance P, and calcitonin gene-related peptide (CGRP), with the neurotoxin, capsaicin, abrogates normal blood cell production. Norepinephrine, neurokinins a and 2, and vasoactive intestinal peptide all have inhibitory effects on in vitro CFU-GM colony formation. Substance P, neurokinin 1, nerve growth factor, and CGRP have stimulatory effects on CFU-GM. Furthermore, in vitro experiments show that, apart from CGRP, all the neuroactive substances we tested operate through effects on accessory cells, stimulating the release of regulatory molecules that have a direct effect on purified CFU-GM. PMID- 11268369 TI - Effects of preprotachykinin-I peptides on hematopoietic homeostasis. A role for bone marrow endopeptidases. AB - Hematopoiesis is maintained by "fine-tuned" regulation among cytokines, neuropeptides, neurotransmitters, and neurotrophic factors. Neurotransmitters, derived from PPT-I exert immune and hematopoietic regulation. PPT-I is also expressed locally in bone marrow (BM) stromal cells. PPT-I peptides induce the production of cytokines in BM cells, resulting in regulation of both committed progenitors (CFU-GM) and primitive hematopoietic progenitors (CAFC). Both types of progenitors are regulated differently by the two major PPT-I peptides, SP and NK-A. Endopeptidases, present in BM cells, can digest SP to produce SP(1-4) and SP(4-11). In this study, we investigated the hematopoietic effects of these fragments on CFU-GM and CAFC. Similar to the two major intact PPT-I peptides (SP and NK-A), we observed different hematopoietic effects by SP(1-4) and SP(4-11). Whereas SP(1-4) exerted inhibitory effects on CFU-GM and CAFC, SP(4-11) mediated stimulatory effects. Similar to NK-A, the inhibitory effects of SP(1-4) can be partly explained by the induction of suppressive cytokines (TGF-beta, TNF-alpha, and INF-gamma). Use of antagonists and screening of a dodecapeptide expression library determined that the effects of SP(1-4) were mediated by NK-1. These results show that PPT-I peptides and their endopeptidase-derived fragments may add to the fine-tuned regulation on hematopoiesis. Furthermore, PPT-I may be exerting autoregulation to protect hematopoietic stem cells. These studies have relevance to stem cell protection and BM transplant. PMID- 11268371 TI - Light and immunomodulation. AB - The immune system is susceptible to a variety of stresses. Recent work in neuroimmunology has begun to define how mood alteration, stress, the seasons, and daily rhythms can have a profound effect on immune response through hormonal modifications. Central to these factors may be light through an eye-brain hormonal modulation. In adult primates, only visible light (400-700 nm) is received by the retina. This photic energy is then transduced and delivered to the visual cortex and, by an alternative pathway, to the suprachiasmatic nucleus (SCN), the hypothalamic region that directs circadian rhythm. Visible light exposure also modulates the pituitary and pineal glands, leading to neuroendocrine changes. Melatonin, norepinephrine, and acetylcholine decrease with light activation, whereas cortisol, serotonin, GABA, and dopamine levels increase. The synthesis of vasoactive intestinal polypeptide (VIP), gastrin releasing peptide (GRP), and neuropeptide Y (NPY) in rat SCN has been shown to be modified by light. These induced neuroendocrine changes can lead to alterations in mood and circadian rhythm as well as immune modulation. An alternative pathway for immune modulation by light is through the skin. Visible light (400-700 nm) can penetrate epidermal and dermal layers of the skin and may directly interact with circulating lymphocytes to modulate immune function. In contrast to visible light, in vivo exposure to UV-B (280-320 nm) and UV-A (320-400 nm) radiation can alter normal human immune function only by a skin-mediated response. It is therefore important, when reporting neuroendocrine immune findings, to control the intensity, timing and wavelength of ambient light. PMID- 11268372 TI - Complex coping patterns and their role in adaptation and neuroimmunomodulation. Theory, methodology, and research. AB - This paper describes the evolution of a model of adaptative coping, as well as an example of the converse, a maladaptive coping pattern, type C. It was hypothesized that the more closely a coping process resembles the inverted U shaped function that characterizes homeostasis for most biological systems, the more likely it is to be adaptive, and to be associated with more positive health outcomes. Maladaptive learned coping patterns, such as type C coping, represent deviations from homeostasis in that they fail to recognize, respond appropriately to, and/or resolve stressors, thus keeping the physiological stress response chronically engaged, with subsequent long-term damage to implicated biological systems. This interpretation of how maladaptive coping patterns such as type C can influence health outcomes is consistent with findings from the author's 20 year program of research on the type C pattern, its assessment, and its association with poorer health indicators and outcomes in cancer (malignant melanoma) and HIV/AIDS. PMID- 11268373 TI - The immune system and schizophrenia. An integrative view. AB - Immune alterations in schizophrenia have been described for decades. Modern immunological methods and new insights into the highly developed and functionally differentiated immune system allow an integrative view of both the older and the recent findings of immunological abnormalities in schizophrenia. Both the unspecific and the specific arms of the immune system seem to be involved in the dysfunction of the immune system in schizophrenia. The unspecific, "innate" immune system shows signs of overactivation in unmedicated schizophrenic patients, as indicated by increased monocytes and gamma delta-cells. Increased levels of interleukin-6 (IL-6) and the activation of the IL-6 system in schizophrenia might be the result of the activation of monocytes/macrophages, too. On the other hand, several parameters of the specific cellular immune system are blunted, such as, for example, the decreased T helper-1 (TH-1)-related immune parameters in schizophrenic patients both in vitro and in vivo. It seems that a TH-1-TH-2 imbalance with a shift to the TH-2 system is associated with schizophrenia. During antipsychotic therapy with neuroleptics, the specific TH-1 related immune answer becomes activated, but in addition the B cell system and antibody production increase. PMID- 11268374 TI - Prenatal influences on neuroimmune set points in infancy. AB - Many factors during fetal life and early infancy have been found to affect the development of immune responses in animals. This study investigated whether acute exposure of the fetal monkey to high levels of corticosteroids would also have a lingering effect on the expression of immune responses still manifest postpartum in yearling juveniles. One month prior to parturition, pregnant rhesus monkeys were administered dexamethasone for two days. Lymphocyte proliferative responses to mitogen were then examined in their offspring when they were between 1.0-1.5 years of age. In addition, cell sensitivity to corticosteroid feedback was assessed by testing the ability of a gradation of cortisol doses to inhibit proliferation. Monkeys generated from dexamethasone-treated pregnancies tended to have lower responses to concanavalin A. Further, their cells were less sensitive to in vitro incubation with cortisol, suggesting that elevated adrenal activity in vivo had downregulated hormone receptors on their cells. These findings concur with the view that steroidal hormones in utero can influence the fetal immune system, resulting in prolonged effects on immune responses after birth. The similarity of the dexamethasone condition to the clinical treatment used in obstetrical practice raises a potential concern about the widespread antenatal exposure of premature infants to steroidal drugs. PMID- 11268375 TI - Illness, cytokines, and depression. AB - Various medical conditions that involve activation of the immune system are associated with psychological and neuroendocrine changes that resemble the characteristics of depression. In this review we present our recent studies, designed to investigate the relationship between the behavioral effects of immune activation and depressive symptomatology. In the first set of experiments, we used a double-blind prospective design to investigate the psychological consequences of illness in two models: (1) vaccination of teenage girls with live attenuated rubella virus, and (2) lipopolysaccharide (LPS) administration in healthy male volunteers. In the rubella study, we demonstrated that, compared to control group subjects and to their own baseline, a subgroup of vulnerable individuals (girls from low socioeconomic status) showed a significant virus induced increase in depressed mood up to 10 weeks after vaccination. In an ongoing study on the effects of LPS, we demonstrated significant LPS-induced elevation in the levels of depression and anxiety as well as memory deficits. These psychological effects were highly correlated with the levels of LPS-induced cytokine secretion. In parallel experiments, we demonstrated in rodents that immune activation with various acute and chronic immune challenges induces a depressive-like syndrome, characterized by anhedonia, anorexia, body weight loss, and reduced locomotor, exploratory, and social behavior. Chronic treatment with antidepressants (imipramine or fluoxetine) attenuated many of the behavioral effects of LPS, as well as LPS-induced changes in body temperature, adrenocortical activation, hypothalamic serotonin release, and the expression of splenic TNF-alpha mRNA. Taken together, these findings suggest that cytokines are involved in the etiology and symptomatology of illness-associated depression. PMID- 11268376 TI - Experimental immunomodulation, sleep, and sleepiness in humans. AB - Infection, inflammation, and autoimmune processes are accompanied by serious disturbances of well-being, psychosocial functioning, cognitive performance, and behavior. Here we review those studies that have investigated the effects of experimental immunomodulation on sleep and sleepiness in humans. In most of these studies bacterial endotoxin was injected intravenously to model numerous aspects of infection including the release of inflammatory cytokines. These studies show that human sleep-wake behavior is very sensitive to host defense activation. Small amounts of endotoxin, which affect neither body temperature nor neuroendocrine systems but slightly stimulate the secretion of inflammatory cytokines, promote non-rapid-eye-movement sleep amount and intensity. Febrile host responses, in contrast, go along with prominent sleep disturbances. According to present knowledge tumor necrosis factor-alpha (TNF-alpha) is most probably a key mediator of these effects, although it is likely that disturbed sleep during febrile host responses involves endocrine systems as well. There is preliminary evidence from human studies suggesting that inflammatory cytokines such as TNF-alpha not only mediate altered sleep-wake behavior during infections, but in addition are involved in physiological sleep regulation and in hypnotic effects of established sedating drugs. PMID- 11268377 TI - Psychoneuroimmunology and HIV/AIDS. PMID- 11268378 TI - Cooperation of pituitary hormone prolactin with interleukin-2 and interleukin-12 on production of interferon-gamma by natural killer and T cells. AB - The pituitary hormone prolactin (PRL) is also produced by cells of the immune system and participates in early and late T cell activating events. We have previously shown a modulatory role of PRL during maturation of dendritic cells (DC). Production of IL-12 by T cell receptor (TCR)-activated DC is necessary for T cells to acquire the Th1 cytokine (i.e., IFN-gamma secreting) profile, which is associated with activation of cellular response. In a separate work, PRL has been shown to increase IFN-gamma synthesis by natural killer (NK) cells. We have extended that study by exploring the ability of PRL to induce IFN-gamma production by T and NK cells in the presence of the specific stimuli IL-12 and IL 2. The individual effect of PRL, IL-12, and IL-2 was specific for NK cells, and IL-2 and IL-12 were much more efficient than PRL. Cooperation of IL-2 and PRL was observed on NK cells. IL-2-induced synthesis of IFN-gamma was increased by physiological concentrations of PRL but was unaffected or inhibited by high concentrations. By contrast, optimal enhancement of IL-12-induced IFN-gamma release was observed with T cells but not with NK cells. Unexpectedly, interaction between PRL and IL-12 occurred only at high concentrations of PRL. These data indicate a complex role of PRL in the cytokine network and point to a revaluation of the proposed immunosuppression by stress-related hyperprolactinemia. PMID- 11268379 TI - Role of cyclophilins in somatolactogenic action. AB - Prolactin (PRL) and growth hormone (GH) are members of the somatolactogenic hormone family, the pleiotropic actions of which are necessary for vertebrate growth and mammary differentiation. The basis for the specific function of these hormones has remained uncertain; however, their action is associated with internalization and translocation into the nucleus. A yeast two-hybrid screen identified an interaction between PRL and cyclophilin B (CypB), a peptidyl prolyl isomerase (PPI) found in the endoplasmic reticulum (ER), extracellular space, and nucleus. The interaction between CypB and PRL/GH was confirmed in vitro and in vivo through the use of recombinant proteins and coimmunoprecipitation studies. The exogenous addition of CypB potentiated the proliferation of PRL- and GH dependent cell lines 18- and 40-fold, respectively. The potentiation of PRL action by CypB was accompanied by a dramatic increase in the nuclear retrotranslocation of PRL. Immunogold electron microscopy has revealed this retrotransport to occur via a vesicular pathway. A CypB mutant, termed CypB-NT, was generated that lacked the putative wild-type N-terminal nuclear localization sequence. Although CypB-NT demonstrated levels of PRL binding and PPI activity equivalent to wild-type CypB, it was incapable of mediating the nuclear retrotranslocation of PRL or enhancing PRL-driven proliferation. These studies reveal CypB as an important chaperone facilitating the nuclear retrotransport and action of the somatolactogenic hormone family. PMID- 11268380 TI - Prolactin regulation of apoptosis-associated gene expression in T cells. AB - Evidence accumulated over the last two decades indicates important actions for prolactin (PRL) in regulation of several functions of the immune system. That PRL can serve to facilitate immune cell proliferation is well established. In addition, PRL appears to play a salient role in the genesis and/or potentiation of certain autoimmune diseases. Recent evidence from several laboratories has extended the spectrum of PRL actions in immunological systems to include regulation of lymphocyte pool size through the process of apoptosis. Experimental results obtained using lactogen-dependent rat pre-T cell lines, the Nb2 lymphoma, have demonstrated that PRL suppresses cell death mechanisms activated by cytokine/hormone deprivation and cytotoxic drugs such as glucocorticoids. In this paper, we review results from studies conducted to investigate the mechanism(s) underlying PRL-regulated apoptosis suppression. Effects of the hormone on expression of apoptosis-associated genes of the Bcl-2 family as well as the protooncogene pim-1 in proliferating Nb2 sublines and in cells exposed to apoptotic stimuli are presented. It is concluded that PRL-mediated apoptosis suppression in immune cells reflects a complex interaction among several gene products. PMID- 11268381 TI - Growth hormone and prolactin expression in the immune system. AB - Prolactin (PRL) and growth hormone (GH) are pituitary hormones that play pivotal roles in lactation and body growth, respectively. In addition, both hormones have been implicated as modulators of immune responses. Since the expression of GH and PRL by leukocytes points to autocrine or paracrine roles during immune responses, our study is aimed at PRL- and GH-production in leukocytes. We show that human peripheral blood granulocytes, which express GH and PRL mRNA, contain high molecular-weight immunoreactive variants of GH and PRL (37 and 43 kDa, respectively), but not the pituitary-sized hormones. Secretion of these variants, or biologically active material as assessed by the Nb2 bioassay, was not detected. On the other hand, certain leukemic myeloid cells secrete 23-kDa, pituitary-sized, PRL, which is biologically active. PMID- 11268382 TI - The role of bim, a proapoptotic BH3-only member of the Bcl-2 family in cell-death control. AB - Apoptosis is an evolutionarily conserved process for killing unwanted cells. Genetic and biochemical experiments have indicated that three groups of proteins are necessary for activation of the cell-death effector machinery: cysteine proteases, their adaptors, and proapoptotic Bcl-2 family members. Antiapoptotic Bcl-2 family members are needed for cell survival. We have cloned Bim, a proapoptotic Bcl-2 family member that shares with the family only a 9-16 aa region of homology [Bcl-3 homology region(BH3)], but is otherwise unique. Bim requires its BH3 region for binding to Bcl-2 and activation of apoptosis. Analysis of Bim-deficient mice has shown that Bim is essential for the execution of some but not all apoptotic stimuli that can be antagonized by Bcl-2. Bim deficient mice have increased numbers of lymphocytes, plasma cells, and myeloid cells, and most develop fatal autoimmune glomerulonephritis. In healthy cells, Bim is bound to the microtubule-associated dynein motor complex, and is thereby sequestered from Bcl-2. Certain apoptotic signals unleash Bim and allow it to translocate to intracellular membranes, where it interacts with Bcl-2 or its homologues. These results indicate that BH3-only proteins are essential inducers of apoptosis that can be unleashed by certain death signals. Unleashed BH3-only proteins neutralize the prosurvival function of Bcl-2-like molecules, and this is thought to liberate Apaf-l-like adapters to activate caspase zymogens, which then initiate cell degradation. PMID- 11268383 TI - Stress and chemotherapy. Combined effects on tumor progression and immunity in animal models. AB - In mice bearing Lewis lung carcinoma, rotational and restraint stress specifically increases the formation of lung metastasis, and restraint stress markedly attenuates the antitumor effects of cyclophosphamide. The aim of this investigation was therefore to examine the effects of restraint stress on tumor metastasis in mice bearing MCa mammary carcinoma, and on the effectiveness of CCNU and DTIC. Restraint stress increases MCa mammary carcinoma metastasis, causes a marked reduction in cyclophosphamide activity, and a minor attenuation of the effects of CCNU and DTIC. The possible occurrence of seasonal factors, observed for the increase by rotational stress of Lewis lung carcinoma metastasis, was also determined for cyclophosphamide effectiveness. The survival time of control mice is longer in February than in June, and is not appreciably modified by rotational stress. The effects of cyclophosphamide are similar in both seasonal periods, and are similarly attenuated by rotational stress. The seasonal effects of rotational stress, and the reduction of the effects of cyclophosphamide caused by rotational stress, are accompanied by corresponding variations in the number of CD3+ and CD4+ splenic T-lymphocyte subsets and in the CD4+/CD8+ ratio, respectively. The reported effects of stress on tumor progression and on the effectiveness of cyclophosphamide thus appear to occur via modulation of immune responses of the host directed against the tumor. These data appear of interest for their experimental implications, and suggest the opportunity to consider the role that the stress during treatment may play in determining the effectiveness of clinical antitumor chemotherapy. PMID- 11268384 TI - Modulation of anticancer cytokines IL-2 and IL-12 by melatonin and the other pineal indoles 5-methoxytryptamine and 5-methoxytryptophol in the treatment of human neoplasms. AB - Lymphocyte number still remains one of the most important immune parameters predicting the prognosis of advanced cancer patients. IL-2 and IL-12 are the main antitumor cytokines in humans, and their effect is modulated by the neuroendocrine system, mainly by the pineal gland through the circadian release of melatonin (MLT) and perhaps that of other indole hormones, such as 5 methoxytryptamine (5-MTT), and 5-methoxytryptophol (5-MTP). MLT has been proven to exert important antitumor immunomodulating effects, whereas the possible immunomodulatory properties of the other pineal indoles are still controversial. In an attempt to better define the pineal neuroendocrine regulation of the anticancer cytokine network, we have evaluated in metastatic solid-tumor patients the effects on lymphocyte number induced by different neuroimmune regimens, consisting of MLT alone (20 mg/day orally in the evening), subcutaneous (s.c.) low-dose IL-2 alone (3 MIU/day in the evening for 6 days/week), s.c. low-dose IL 12 alone (0.5 mcg/kg once/week in the morning), IL-12 plus MLT, IL-2 plus MLT, and IL-2 plus MLT plus 5-MTT (10 mg/day orally in the afternoon) plus 5-MTP (5 mg/day orally at noon). The results showed the following evidence: (1) MLT alone is unable to induce lymphocytosis; (2) MLT significantly enhances IL-2-induced lymphocytosis; (3) IL-12 alone determines lymphocytopenia, which can be reversed by MLT; (4) IL-2 plus IL-12 induces a very pronounced lymphocytosis, which can be further amplified by MLT; (5) a total pineal endocrine replacement therapy with MLT, 5-MTT, and 5-MTP further increases IL-2-induced lymphocytosis with respect to MLT plus IL-2 alone. Therefore, this study confirms that IL-2- and IL-12 dependent anticancer immunity is under a pineal modulation. PMID- 11268385 TI - What's in a name? Neuroimmunomodulation or psychoneuroimmunology? AB - Compelling evidence is presented to support the hypothesis that psychological processes affect immune function. Consequently, it is argued that psychological processes should be included in human immunological studies and that neuroimmunomodulation could accurately by called psychoneuroimmunology. PMID- 11268386 TI - Sympathetic innervation affects superantigen-induced decrease in CD4V beta 8 cells in the spleen. AB - The stimulation by superantigens of T cells expressing an appropriate V beta chain results in a strong proliferative response that is followed by a state of energy specific for the antigen used. This model was used to continue our studies on immunoregulatory host neuroendocrine responses. We have recently found that four days after administration of the superantigen staphylococcal enterotoxin B (SEB) into mice, that is, at an early stage of the anergic phase, the decrease in the percentage of splenic CD4V beta 8 was accompanied by a decrease in the splenic concentration of the sympathetic neurotransmitter noradrenaline (NA) as compared to vehicle-injected mice. No comparable changes were detected in the kidney. At this point, blood levels of NA, adrenaline, and corticosterone were comparable in SEB- and vehicle-injected mice. We have also found that the decrease in splenic CD4V beta 8 cells was not observed in animals that had been chemically sympathectomized prior to the administration of the superantigen. These results indicate that the sympathetic response induced by SEB may have immunoregulatory implications. PMID- 11268387 TI - Variability of interactions between neuroendocrine and immunological functions in physiological aging and dementia of the Alzheimer's type. AB - A link between neuroendocrine and immunological changes has been suggested in the pathophysiology of dementia of the Alzheimer's type (DAT). Healthy young and old subjects and patients with DAT were recruited to evaluate the chrononeuroendocrine organization of cortisol, GH, and melatonin (MLT) secretions. The study was carried out together with the evaluation of natural killer (NK) cell function: cytotoxic activity (NKCC) and TNF-alpha and IFN-gamma release after exposure to IL-2 (100 U/mL). Moreover, a cerebral morphometric analysis of hippocampus and temporal lobe (MRI) was performed. The activation of hypothalamo-pituitary-adrenal (HPA) axis and the decrease of GH, and MLT nocturnal peaks were associated with normal NKCC and TNF-alpha/IFN-gamma in healthy elderly subjects, whereas in DAT patients the same neuroendocrine changes occurred together with abnormal NKCC (spontaneous and IL-2/IFN-beta-modulated) and with alterations of TNF-alpha/INF-gamma generation from NK. Moreover significant correlations among the increase of NKCC and TNF-alpha and the decrease of cognitive function were found in the DAT group. These correlations were associated with the impairment of nocturnal GH and MLT levels and with the relatively higher serum cortisol concentrations. Moreover, the impairment of cortisol suppression after dexamethasone (1 mg orally at 23:00) was significantly correlated with the increase of spontaneous release of TNF-alpha and with IL-2 modulated NKCC. Finally the imunoneuroendocrine alterations found in DAT were associated with the reduction of cerebral volume in hippocampus and temporal lobes. Taken together these data indicate that the immunoneuroendocrine balance is maintained in physiological aging, whereas NK immune dysregulation in DAT could contribute to altering the neuroendocrine functions and to extend the progression of neurodegeneration and dementia. PMID- 11268388 TI - Interleukin-1 beta and thymic peptide regulation of pituitary and glial cell cytokine expression and cellular proliferation. AB - Interleukin-6 (IL-6) is a B-cell differentiating and T-cell activating cytokine that is expressed in T cells, neutrophils, monocytes, macrophages, and mast cells. Because IL-6 is also synthesized and released by anterior pituitary cells and IL-6 stimulates pituitary hormone release, this cytokine may serve a paracrine or autocrine role within the pituitary. Interleukin-1 beta (IL-1 beta) stimulates IL-6 release from anterior pituitary cells through a mechanism that involves lysophosphatidylcholine (LPC 18:0) generation and protein kinase C activation. In the rat C6 glioma cell line, IL-1 beta synergistically stimulates IL-6 release in the presence of increased intracellular cAMP concentrations. The catecholamines and serotonin also synergistically stimulate IL-6 release in the presence of IL-1 beta. LPC 18:0 synergistically increases IL-6 release in the presence of norepinephrine, and IL-1 beta transiently increases LPC 18:0 formation in C6 cells. Therefore, IL-1 beta induction of LPC 18:0 may lead to increases in IL-6 production via activation of a kinase cascade. The bovine thymic preparation, thymosin fraction 5 (TF5), also stimulates IL-6 release from C6 glioma cells in a protein kinase C-dependent manner. Of interest, TF5 inhibits the proliferation of C6 cells, pituitary adenoma MMQ cells, and promyelocytic HL 60 cells. We suggest that a thymic hormone immune surveillance mechanism may suppress neuroendocrine and hematopoietic tumor formation. Thus, IL-1 beta and certain thymic peptides act to increase IL-6 expression in neuroendocrine cells. The enhanced production of neuroendocrine cytokines may affect hormone secretion, neurotransmission, and the development of certain neurodegenerative disorders (e.g., Alzheimer's disease). The isolation of the active component of TF5 that inhibits neuroendocrine and hematopoietic tumor cell proliferation will provide a potential therapeutic strategy for the treatment of these tumors. PMID- 11268389 TI - Cytokine activation of the HPA axis. AB - The observation that administration of interleukin-1 (IL-1) to animals activates the hypothalamo-pituitary-adrenocortical (HPA) axis stimulated great interest in the significance and mechanism of this response, and in whether other cytokines have similar activities. Interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF alpha) share HPA-activating activity, although they are less potent and effective than IL-1, whereas IL-2 and interferon alpha(IFN alpha) lack activity. Small increases in body temperature occur in response to IL-1, IL-6 and TNF alpha, but these changes are prevented by inhibitors of cyclooxygenase (COX) and do not appear to be related to the HPA-activation. The rapid HPA-activating effects of IL-1 are impaired by COX inhibitors, but the more prolonged HPA activation associated with intraperitoneal injections is not affected, indicating multiple mechanisms for IL-1-induced HPA activation. The HPA response to IL-6 is not sensitive to COX inhibitors, but that to TNF alpha appears to be. The HPA activating activity of IL-1 is associated with increases in the apparent release of brain noradrenaline (NA) and serotonin (5-HT), but not dopamine, as well as with increased brain tryptophan. The NA changes, but not these in serotonin metabolism and tryptophan, are prevented by COX inhibitors. IL-6 has effects on serotonin and tryptophan like those of IL-1, but no detected effect on NA. TNF alpha has some effect on NA and tryptophan, but only at relatively high doses. IFN alpha lacks activity on these neurochemicals. Manipulation of noradrenergic, but not serotonergic systems alters the IL-1-induced HPA activation, suggesting the involvement of NA. However, brain NA does not appear to be essential for HPA activation in mice. PMID- 11268390 TI - In vivo immunomodulation by peripheral adrenergic and cholinergic agonists/antagonists in rat and mouse models. AB - Our work is devoted to defining relationships between the immune system and the adrenergic and cholinergic systems in vivo. In the rat model, we have shown that the cells of different immune compartments express the genes of a defined set of adrenergic/cholinergic receptors, and it was shown that lymphocytes are a site of non-neuronal production of norepinephrine and acetylcholine. Furthermore, using implantable slow-release tablets containing adrenergic or cholinergic agonists/antagonists, distinct and partly opposite effects were observed on peripheral immune functions. Concerning sympathetic immunoregulation, our data- in contrast to those of other studies--suggest that an enhanced adrenergic tonus leads to immunosuppression primarily via alpha 2-receptor-mediated mechanisms. Beta-blockade strongly enhances this effect, most likely by inhibition of pineal melatonin synthesis. In recent experiments on the kinetics it was found that the continuous alpha-adrenergic treatment entails a strong suppression of cellular responsiveness during the first few hours, which is increasingly followed by a general loss of lymphocytes in blood and lymphoid organs most likely due to enhanced apoptosis. More recently, we have extended our studies to the mouse model. First data obtained with RNAse protection assays suggest a biphasic effect on the gene expression of several cytokines in spleen cells due to adrenergic in vivo treatment. PMID- 11268391 TI - Possible function of IL-6 and TNF as intraadrenal factors in the regulation of adrenal steroid secretion. AB - Interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF alpha) and their mRNAs are present in the human, rat, and bovine adrenal cortex. The release of these cytokines from adrenal cells is regulated by factors that alter adrenal function (e.g., ACTH, angiotensin II, interleukin-1). IL-6 and TNF type 1 receptors are also present on adrenocortical cells. Exposure to IL-6 increases cortisol or corticosterone release from human, bovine, and rat adrenal cells. IL-6 increases basal and ACTH-stimulated aldosterone release, but inhibits angiotensin II stimulated aldosterone secretion from bovine adrenal cells. IL-6 increases dehydroepiandrosterone (DHEA) release from human cells, but decreases DHEA secretion from bovine cells. TNF alpha inhibits corticosterone release from normal rat adrenal cells or fragments, but increases corticosterone release from cholestatic rat adrenal slices. TNF alpha decreases cortisol release from bovine and fetal human adrenal cells, but increases cortisol release from adult human adrenal cells. TNF alpha inhibits aldosterone secretion from rat and bovine adrenocortical cells. TNF alpha does not affect DHEA secretion from fetal human adrenocortical cells, but inhibits basal and ACTH-stimulated DHEA release from bovine adrenal cell. Because IL-6 and TNF alpha are produced in the adrenal gland and modify adrenal steroid secretion, these cytokines may function as intraadrenal factors in the regulation of adrenal steroid secretion. PMID- 11268392 TI - Gaseous neuromodulators in the control of neuroendocrine stress axis. AB - The gaseous neuromodulator carbon monoxide has been shown to reduce the stimulated release of stress neuropeptides, such as vasopressin and oxytocin, from the rat hypothalamus in vitro, while evidence concerning corticotropin releasing hormone is controversial. In vivo studies have been conducted in the rat, inhibiting heme oxygenase activity--and hence carbon monoxide biosynthesis- in the central nervous system by means of specific heme oxygenase blockers; these studies showed that basal heme oxygenase activity tends to oppose exaggerated increases in vasopressin secretion following immune-inflammatory challenges, whereas it favors the normal rise in circulating ACTH which follows footshock. Another gas normally produced in mammalian brains under basal conditions, hydrogen sulfide, also appears to play a role in the control of the hypothalamo pituitary-adrenal axis. Indeed, increases in hydrogen sulfide levels within the hypothalamus, either obtained with hydrogen sulfide-enriched media or by the addition of the hydrogen sulfide precursor S-adenosyl-methionine, are associated with the inhibition of the stimulated release of corticotropin-releasing hormone from rat hypothalamic explants. Parellel in vivo experiments in the rat under resting conditions and after stress-induced adrenocortical activation show that S adenosyl-methionine significantly reduces the rise in serum corticosterone levels caused by 1-h exposure to cold. These results demonstrate the pathophysiological importance of both carbon monoxide and hydrogen sulfide in the regulation of neuroendocrine function. PMID- 11268393 TI - Cytokines, leptin, and the hypothalamo-pituitary-adrenal axis. AB - The endocrine and immune systems are linked via an elaborated communication system constituted by an array of cytokines and neuropeptides which interact to modulate the integrated response of an organism to infection. Weight loss and anorexia, probably secondary to cytokine release, frequently accompany infection, but leptin could also play a role. Like cytokines, leptin serves as a peripheral messenger to convey signals to the brain. Expression of leptin is stimulated by glucocorticoids, endotoxins, and cytokines; on the other hand, leptin seems to inhibit the activation of the hypothalmo-pituitary-adrenal (HPA) axis. Indeed leptin exerts a direct, dose-dependent inhibition of stimulated cortisol secretion by normal human and rat adrenal cells in vitro. These effects are mediated by the long isoform of the leptin receptor, because its transcript is expressed in the adrenal tissue. In addition we investigated the role played by the glucocorticoids in the development of tolerance of the hypothalamo corticotropic, immune and adipose system responses to repeated endotoxin administration. Unlike that of the corticotropic axis, tolerance of the immune and adipose systems is at least partially glucocorticoid-independent. This crosstalk between the endocrine, immune, and adipose systems may be of prime importance to homeostasis in pathophysiological events occurring during infection. PMID- 11268394 TI - SOCS proteins: modulators of neuroimmunoendocrine functions. Impact on corticotroph LIF signaling. AB - Several members of the newly characterized family of suppressor of cytokine signaling (SOCS) proteins-such as SOCS-1, SOCS-3, and CIS-act as negative regulators of the cytokine-induced Jak-STAT signaling cascade. The expression of SOCS proteins is stimulated by a variety of cytokines and hormones in a tissue specific manner. This article reviews our current understanding of SOCS proteins and their role as modulators of neuroimmunoendocrine functions, for example, in signaling of leptin, growth hormone, and prolactin, specially focusing on the impact of SOCS proteins on corticotroph leukemia inhibitory factor (LIF) signaling. LIF, a member of the gp130 sharing cytokine family, modulates pituitary development, POMC gene expression, and ACTH secretion. Current data on the negative autoregulatory function of the suppressor of cytokine signaling, SOCS-3, in LIF-induced POMC gene expression and ACTH secretion are extensively discussed. PMID- 11268395 TI - Macrophage migration inhibitory factor (MIF). A critical neurohumoral mediator. PMID- 11268396 TI - Functional cross-talk among cytokines, T-cell receptor, and glucocorticoid receptor transcriptional activity and action. AB - The main communicators between the neuroendocrine and immune systems are cytokines and hormones. We studied the molecular interaction between immune activators (cytokines and T-cell receptors [TCRs]) and the glucocorticoid receptor (GR) in cells in which glucocorticoids play a key regulatory function: (1) cellular targets of TNF-induced cytotoxicity; (2) the pituitary gland; and (3) thymic cells. Cytokines (TNF-alpha and IL-1) increase glucocorticoid-induced transcriptional activity of the GR via the DNA-glucocorticoid response elements (GREs) in cells transfected with a glucocorticoid-inducible reporter plasmid. As a functional physiological correlate, priming of fibroblastic cells with a low dose of TNF significantly increases the sensitivity to glucocorticoid inhibition of TNF-induced apoptosis (without involving NF-kappa B). Priming of AtT-20 mouse corticotrophs and Cushing pituitary cells with IL-1 increases the sensitivity to glucocorticoid inhibition of CRH-induced ACTH/POMC expression. In thymocytes, activation of the T-cell receptor counteracts the glucocorticoid-induced thymic apoptosis by downregulating the glucocorticoid action on GRE-driven apoptotic genes. Thus, cytokines and immune mediators prevent their own deleterious effects not only by stimulating glucocorticoid production, but also by modifying the sensitivity of the target cells for the glucocorticoid counter-regulatory action. The functional cross-talk at the molecular level between immune signals and glucocorticoids is essential to determine the biological response to both mediators and constitutes the ultimate level of interaction between the immune and neuroendocrine mediators. PMID- 11268397 TI - Gender, neuroendocrine-immune interactions and neuron-glial plasticity. Role of luteinizing hormone-releasing hormone (LHRH). AB - Signals generated by the hypothalamic-pitutary-gonadal (HPG) axis powerfully modulate immune system function. This article summarizes some aspects of the impact of gender in neuroendocrine immunomodulation. Emphasis is given to the astroglial cell compartment, defined as a key actor in neuroendocrine immune communications. In the brain, the principal hormones of the HPG axis directly interact with astroglial cells. Thus, luteinizing hormone releasing hormone, LHRH, influences hypothalamic astrocyte development and growth, and hypothalamic astrocytes direct LHRH neuron differentiation. Hormonally induced changes in neuron-glial plasticity may dictate major changes in CNS output, and thus actively participate in sex dimorphic immune responses. The impact of gender in neuroimmunomodulation is further underlined by the sex dimorphism in the expression of genes encoding for neuroendocrine hormones and their receptors within the thymus, and by the potent modulation exerted by circulating sex steroids during development and immunization. The central role of glucocorticoids in the interactive communication between neuroendocrine and immune systems, and the impact of gender on hypothalamic-pituitary-adrenocortical (HPA) axis modulation is underscored in transgenic mice expressing a glucocorticoid receptor antisense RNA. PMID- 11268398 TI - Oncology in neuroimmunomodulation. What progress has been made? AB - In 1987 in Dubrovnik, Yugoslavia, N.H. Spector named a new discipline: Neuroimmunomodulation. R. Ader called this new discipline psychoneuroimmunomodulation when the major emphysis was on its behavioral aspects. Neuroimmunomodulation (NIM) is devoted to the study of the interactions at different morphologic and functional levels among the immune, nervous, and endocrine systems. In fact, this science is the modern manifestation of an old science: in the words of B.D. Jankovic (1987), "Neuroimmunomodulation is a modern reflection in neurosciences and immunosciences of the ideas and experience of philosophers and ingenious observers of ancient Egypt, Greece, China, India, and other civilizations that the mind is involved in the defense against diseases." Twelve years ago NIM was regarded by many conventional scientists almost as a form of witchcraft. Today it may be the fastest growing area of biomedical science research in the world. Important clinical applications will not be far behind. NIM research has also progressed in the field of oncology research. Topics such as treatment of hormone-dependent cancer with analogues of hypothalamic hormones, the role of opioids and T cells in cancer, stress-cancer immune connections, the anticancer role of melatonin and cytokines, immunotherapy of cancer, and the role of psychotherapy in cancer patients represent some lines of research that have been or are being investigated by scientists. Some areas remain to be thoroughly investigated such as the influence of physical exercise (sports), music (classical or modern), and/or relaxation techniques (e.g. yoga) on the development of human cancer. This paper reviews the role of NIM in oncology and provides some perspectives for further research and development of clinical applications. PMID- 11268399 TI - Thymic neuroendocrine self-antigens. Role in T-cell development and central T cell self-tolerance. AB - The repertoire of thymic neuroendocrine precursors plays a dual role in T-cell differentiation as the source of either cryptocrine accessory signals in T-cell development or neuroendocrine self-antigens presented by the thymic major histocompatibility complex (MHC) machinery. Thymic neuroendocrine self-antigens usually correspond to peptide sequences highly conserved during the evolution of one family. The thymic presentation of some neuroendocrine self-antigens is not restricted by MHC alleles. Oxytocin (OT) is the dominant peptide of the neurohypophysial family. It is expressed by thymic epithelial and nurse cells (TEC/TNCs) of different species. Ontogenetic studies have shown that the thymic expression of the OT gene precedes the hypothalamic one. Both OT and VP stimulate the phosphorylation of p125FAK and other focal adhesion-related proteins in murine immature T cells. These early cell activation events could play a role in the promotion of close interactions between thymic stromal cells and developing T cells. It is established that such interactions are fundamental for the progression of thymic T-cell differentiation. Insulin-like growth factor 2 (IGF 2) is the dominant thymic polypeptide of the insulin family. Using fetal thymic organ cultures (FTOCs), the inhibition of thymic IGF-2-mediated signaling was shown to block the early stages of T-cell differentiation. The treatment of FTOCs with an mAb anti-(pro)insulin had no effect on T-cell development. In an animal model of autoimmune type 1 diabetes (BB rat), thymic levels of (pro)insulin and IGF-1 mRNAs were normal both in diabetes-resistant and diabetes-prone BB rats. IGF-2 transcripts were clearly identified in all thymuses from diabetes-resistant adult (5-week) and young (2- and 5-days) BB rats. In marked contrast, the IGF-2 transcripts were absent and the IGF-2 protein was almost undetectable in +/- 80% of the thymuses from diabetes-prone adult and young BB rats. These data show that a defect of the thymic IGF-2-mediated tolerogenic function might play an important role in the pathophysiology of autoimmune Type 1 diabetes. PMID- 11268400 TI - Human T lymphopoiesis. In vitro and in vivo study models. AB - Successive steps in T lymphocyte differentiation and T potential of human stem cells (HSC) can be tested in the following models: (a) the infusion of cells in NOD-SCID mice, (b) the injection of cells in renconstituted SCID/hu mice, (c) the differentiation of cells in fetal thymus organ culture (FTOC), and (d) on thymic stromal layers. Using mixed human-murine FTOC, we showed (a) TCR alpha beta, TCR gamma delta lymphocytes, NK cells, and dendritic cells complete their differentiation, (b) IL-7R alpha signaling and IL-7 are essential, (c) a detailed phenotypic and functional analysis of discrete successive steps of positively selected thymocytes, (d) an efficient transduction of genes in HSC with persistent gene expression throughout the T-lymphocyte differentiation, and (e) adaptation to submerging high oxygen culture increases the test sensitivity to a clonal assay. Other approaches are the in vivo SCID/hu reconstitution model. With this method small fragments of human fetal liver and thymus are implanted under the kidney capsule of an adult SCID mouse with result in an impressive human thymus organ, six months after transplantation. We use this model to study thymus T-cell developmental kinetics, development of gene-marked precursor cells and thymic homing of precursor cells. PMID- 11268401 TI - Role of glucocorticoids in early T-cell differentiation. AB - The results of the T-cell differentiation in the progeny of adrenalectomized pregnant rats (Adx fetuses), an experimental model that ensures the absence of glucocorticoids (GCs) during the first stages of development, are summarized. In Adx thymuses there is an accelerated maturation of thymocytes that is reversed by in vivo GC replacement. In addition, Adx thymuses show decreased cell content, which correlates with both the increased numbers of apoptotic cells and an early migration of DP (CD4+CD8+) and SP (both CD4+CD8- and CD4-CD8+) thymocytes to the spleen. As shown by in vitro recolonization assays, accelerated T-cell differentiation is a consequence of changes in the biology of lymphoid precursors occurring in the fetal liver of Adx fetuses. They arrive at the thymic primordium earlier and mature faster than the fetal liver lymphoid progenitors from Sham control fetuses. After the establishment of a fetal hypothalamus-pituitary-gland adrenal-gland (HPA) axis, there is a gradual normalization of the T-cell development Adx fetuses. PMID- 11268402 TI - The thymus at the crossroad of neuroimmune interactions. AB - The numerous relationships existing between the nervous and the immune systems suggest that the neural networks present in the intrathymic microenvironment may influence T-cell development. We previously reported that thymic neural-crest derived stromal cells are involved in a neural differentiation pathway and are able to produce neurotrophic factors and neurokines that are in turn able to increase and/or modulate thymic-stromal cell neuronal phenotype. We also showed that EGF promotes a neural phenotype in thymic epithelial cells by enhancing the expression of neuronal-specific markers, neurotransmitters, and neuropoietic cytokines, such as IL-6 and CNTF. More recently we showed that the effect of EGF in directing thymic epithelial cells toward a neural-oriented cell fate is mediated by modulating the expression of genes directly involved in neurotypic differentiation (i.e., thrombospondin-1). EGF-induced regulation of stromal cells may also affect T-cell differentiation, as we observed that an EGF-pretreatment reduces the ability of thymic epithelial cells to sustain thymocyte differentiation in vitro. Finally, we demonstrated that a complex network involving the neurotrophin BDNF and its specific receptors may have a role in sustaining thymocyte precursor survival and supporting the thymocyte differentiation process. All together, our results suggest that the thymus may be the site of integration of different neuroimmune networks that are potentially involved in the regulation of thymocyte survival and/or differentiation. PMID- 11268403 TI - Is there a role for growth hormone upon intrathymic T-cell migration? AB - Intrathymic T-cell differentiation is essentially driven by the thymic microenvironment, a tridimensional network formed by thymic epithelial cells and to a lesser extent, dendritic cells, macrophages, fibroblasts, and extracellular matrix components. Thymocyte migration throughout the thymus is partially dependent on extracellular-matrix (ECM)-mediated interactions. Herein we investigated the putative role of growth hormone (GH) upon events related to intrathymic T-cell migration. We demonstrated that GH upregulates the expression of ECM ligands and receptors in distinct preparations of cultured thymic epithelial cells TECs). We also showed that adhesion of thymocytes to thymic epithelial cells was significantly increased by GH treatment, an effect that could be consistently abrogated when TECs were treated to antifibronectin, anti VLA5, antilaminin, or anti-VLA6 antibodies before addition of thymocytes to the cultures. We also studied thymic nurse cells (TNCs), lymphoepithelial complexes that can be isolated ex vivo from the thymus. In this system, we had previously demonstrated that ECM ligands and receptors control both inward and outward thymocyte traffic. We then showed that GH enhances thymocyte release from TNCs, as well as the reconstitution of these lymphoepithelial complexes. Lastly, we evaluated the in vivo influence of GH on thymocyte exit. This was done by means of intrathymic injection of GH plus fluorescein isothiocyanate (FITC), and further analysis of recent thymic emigrants (FITC+ cells) in peripheral lymphoid organs, as defined by CD4/CD8-based cytofluorometric phenotyping. The proportions of FITC+ T cells appeared augmented in lymph nodes in GH-treated mice, as compared to controls. Taken together, these data indicate that GH stimulates intrathymic T-cell traffic, an effect that is at least partially mediated by extracellular matrix-mediated interactions. PMID- 11268404 TI - The opioid antagonist naloxone induces a shift from type 2 to type 1 cytokine pattern in normal and skin-grafted mice. AB - Opioid peptides affect different immune functions. We present evidence that these effects could be mediated by the modulation of Th1/Th2 cytokine production. The acute and chronic treatment with the opioid receptor antagonist naloxone decreased the production of IL-4 by splenocytes of C57BL/6 and Balb/cJ mice, that present a Th1/Th2 dominance, respectively, immunized with the protein antigen KLH. In contrast, IL-2 and IFN-gamma levels were increased after naloxone treatment. These results indicate that naloxone increases Th1 and decreases Th2 cytokine production. Moreover in C57BL/6 mice, naloxone treatment was able to accelerate skin-graft rejection, a Th1-mediated phenomenon, by increasing Th1 cytokine production. The effect of naloxone could be ascribed to the removal of the regulatory effects exerted by endogenous opioid peptides, which could activate Th2 and suppress Th1 cytokines. PMID- 11268405 TI - Expression of delta opioid receptors and transcripts by splenic T cells. AB - Delta opioid receptors (DORs) and preproenkephalin-A-derived opiate peptides are expressed by mononuclear cells in various lymphoid organs. DOR ligands modulate a variety of immune functions, such as T-cell proliferation, calcium mobilization, and cytokine production. Recently, quiescent T cells were found to express low levels of DOR transcripts, which increased due to the following: cell culture of unstimulated murine splenocytes (depending on cell density); cross-linking the T cell receptor (TCR) with anti-CD3-epsilon; and a single in vivo exposure to staphylococcal enterotoxin B (SEB). Enhanced expression of DOR mRNA was mediated transcriptionally. Moreover, PMA + ionomycin, which mimic the proliferative signal of anti-CD3, inhibited the expression of DOR mRNA. Using semiquantitative immunofluorescence to detect DORs, SEB was found to increase the fraction of T cells that expressed DOR and to enhance the relative level of DOR expression per T cell. Previous studies have shown that DOR agonists inhibited the anti-CD3 stimulated production of interleukin-2 and T-cell proliferation. Therefore, the enhanced expression of DORs by activated T cells may be capable of downregulating the T-cell activation program. PMID- 11268406 TI - Acute effects of morphine on blood lymphocyte proliferation and plasma IL-6 levels. AB - Administration of morphine (10 mg/kg) to rats was found to decrease the proliferative potential of blood lymphocytes by 60-80% and concurrently elevate circulating levels of the cytokine, interleukin-6 (IL-6), 2- to 4-fold. Both parameters were similarly altered upon the central administration of morphine and were blocked upon pretreatment of animals with the opioid receptor antagonist, naltrexone. These results suggest that the activation of central opioid receptors is involved in morphine-induced inhibition of lymphocyte proliferation as well as increases in circulating levels of IL-6. Studies addressing the potential peripheral mechanisms demonstrated that intact ganglionic transmission was required for both effects of morphine. Although the suppression by morphine of lymphocyte proliferation appeared to be largely independent of stimulation of the hypothalamic-pituitary-adrenal axis, the elevation of IL-6 was completely abolished in adrenalectomized animals. Collectively, these results suggest that central opioid receptor activation results in changes in different immune parameters that can be mediated through distinct peripheral mechanisms. PMID- 11268407 TI - Expression of preproenkephalin mRNA and production and secretion of enkephalins by human thymocytes. AB - Human thymocytes were tested for the capacity to express the preproenkephalin (PPE) gene and for production of the end product metenkephalin (MENK). It is shown here for the first time that the cytokines IL-2, IL-4, IL-6, and TGF-beta are capable of inducing PPE mRNA expression. Moreover, a culture of thymocytes with the cytokines results in intracellular expression of MENK as determined by immunohistochemistry. Thymocytes do not secrete detectable amounts of MENK, however, but only the larger MENK-containing peptides or proteins. Cytokines IL-1 beta and IL-10 increase the expression of PPE mRNA in 50% of the thymuses tested, whereas IFN-gamma does not induce changes in PPE mRNA expression. PMID- 11268408 TI - Susceptibility to autoimmune disease and drug addiction in inbred rats. Are there mechanistic factors in common related to abnormalities in hypothalamic-pituitary adrenal axis and stress response function? AB - DA and LEW inbred rats are extraordinarily susceptible to a wide range of experimental autoimmune diseases. These diseases include rheumatoid arthritis models such as collagen-induced arthritis (CIA) and adjuvant-induced arthritis (AIA), multiple sclerosis models such as myelin-basic-protein (MBP)-induced experimental autoimmune encephalomyelitis (MBP-EAE), and autoimmune uveitis models such as retinal S antigen (SAG) and interphotoreceptor-retinoid-binding protein (IRBP)-induced experimental autoimmune uveitis (SAG-EAU and IRBP-EAU, respectively). DA and LEW rats are also addiction-prone to various drugs of abuse, such as cocaine. Moreover, they exhibit a variety of behavioral and biochemical characteristics that appear to be related to their susceptibility to addiction. By contrast, F344 and BN rats show quite different phenotypes. They are relatively resistant to CIA, AIA, MBP-EAE, SAG-EAU, and IRBP-EAU, and they are relatively resistant to addiction. Interestingly, both DA and LEW rats, in contrast to F344 and BN rats, have abnormalities in hypothalamic-pituitary adrenal (HPA) axis function. For example, circadian production of corticosteroids is very abnormal in DA and LEW rats; that is, they exhibit minimal circadian variation in corticosterone levels. Since corticosteroids potentially have significant influences on immune function and autoimmune disease susceptibility and may also influence sensitivity to drugs of abuse, we have begun to dissect genetic control of these various phenotypic differences, focusing initially on the regulation of autoimmune disease expression. Using genomewide scanning techniques involving F2 crosses of DA x F344 (CIA and AIA), DA x BN (CIA), and LEW x F344 [IRBP-EAU and streptococcal-cell-wall arthritis (SCWA)], we have identified, to date, 14 genomic regions [quantitative trait loci (QTL)] that regulate disease expression in these crosses. Development and analysis of QTL congenic rats involving these loci are in progress and should permit us to address the relationships among autoimmune disease susceptibility, drug addiction, and HPA axis and stress response function. These initial data, however, indicate that the genetic control of the autoimmune disease traits is highly complex. PMID- 11268409 TI - Neuroendocrine manifestations in Sjogren's syndrome. Relation to the neurobiology of stress. AB - Evidence suggests that autoimmune rheumatic diseases are associated with neuroendocrine dysfunction. Sjogren's syndrome (SS) is proposed as an ideal model to study perturbations in the neuroimmune axis, since patients tend to be medication free and studies are not confounded by the effects of chronic immunosuppressive therapy. The functional integrity of the adrenal, gonadal, and thyroid axes was assessed in SS. Pituitary function of the HPA axis was evaluated directly by determining the ACTH released during oCRH stimulation, while adrenal function was assessed indirectly by endogenous ACTH released during oCRH stimulation. Low basal activity of the HPA axis was associated with pituitary hyporesponsiveness to exogenous CRH, as well as hyporesponsiveness of the adrenal glands to endogenous ACTH. These findings are compatible with a central deficiency of the adrenal axis. An overall attenuated and delayed LH and FSH response to LHRH stimulation was also indicative of central dysfunction of the gonadal axis in SS. SS patients demonstrated elevated basal TSH levels and evidence of mild hypothyroidism. Basal prolactin concentrations were also elevated in SS, and both TSH and PRL showed relatively increased responses to TRH stimulation. The data suggest a central deficiency in all three neuroendocrine axes: adrenal, gonadal, and thyroid. It is not clear if any one system plays a primary role in the expression of the disease. Rather, it is likely that the net effect involves the synergistic and antagonistic effects of multiple hormones. Taken together, adrenal and gondadal steroid hormone deficiency, plus elevated PRL levels, probably greatly affect immune function in SS patients. PMID- 11268410 TI - Gonadotropin-releasing hormone and G proteins: potential roles in autoimmunity. AB - The hypothalamic homone gonadotropin-releasing hormone (GnRH) displays gender specific actions. Pituitary responsiveness to GnRH is generally increased by estrogens and decreased by androgens. GnRH is now known to be produced by the immune system and to exert potent immunologic actions. Our central hypothesis is that gender differences in responsiveness to GnRH in the immune system play a pivotal role in the gender differences in immunity and autoimmunity. Studies in lupus-prone mice demonstrate that GnRH exacerbates murine lupus in a gender specific fashion. Subsequent studies from our laboratory suggest that the gender differences in immunologic responsiveness to GnRH may relate to differences in the expression of the signal transducers through which GnRH acts, namely, the G proteins, Gs, and Gq/11. We have further demonstrated gender differences in second messengers for GnRH, IP3, and cAMF in immune cells. We have also demonstrated that GnRH agonist increases the quantities and/or activity of G proteins in immune cells in a gender-specific fashion. We speculate that gender differences in GnRH production and action, and in G protein expression play a role in a variety of autoimmune diseases that affect females predominantly. PMID- 11268411 TI - Neuroendocrine and other factors in the regulation of inflammation. Animal models. AB - A variety of animal models have been used to study the role of neuroendocrine responses in various aspects of autoimmune/inflammatory disease. Complex models of autoimmune disease, such as inflammatory arthritis in rats and thyroiditis in chickens, indicate a role for blunted HPA axis and dysregulated sympathoneuronal responses in susceptibility to autoimmune disease. A variety of approaches including pharmacological, surgical (ablation, transplantation), genetic linkage and segregation studies have been used to identify factors contributing to the phenotypes of susceptibility or resistance to inflammatory/autoimmune disease. Innate inflammation, or the earliest nonspecific form of the inflammatory response, which is characterized by fluid exudation and migration of immune cells to inflammatory sites, is a subtrait of these forms of inflammatory disease. Genetic linkage and segregation studies in inflammatory susceptible and resistant rat strains indicate that this subtrait is multigenic and polygenic; that is, that multiple loci on multiple chromosomes, each with a weak effect, control this trait, and that there is a large environmental component to the variability of this trait. Such information derived from animal studies can be used to target candidate genes for further study and to inform the design of human studies. PMID- 11268412 TI - Perturbations of arginine vasopressin secretion during inflammatory stress. Pathophysiologic implications. AB - Pro-inflammatory cytokines, such as interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF alpha), released from inflammatory foci, can activate the hypothalamus to produce corticotrophin-releasing hormone (CRH) and arginine vasopressin (AVP). These hypothalamic peptides in synergy increase ACTH production by the pituitary gland and hence corticosteroid (CS) secretion by the adrenal cortices. CS dampens inflammation. The pituitary also produces prolactin (PRL), which is pro-inflammatory, and macrophage inhibitory factor (MIF), which by counteracting the anti-inflammatory and immunosuppressive effects of CS, is pro-inflammatory. Lewis rats develop a variety of induced autoimmune inflammatory conditions, such as streptococcal cell wall arthritis, whereas the histocompatible F344 Fisher rats are resistant to this condition. Lewis rats have a defective hypothalamic-pituitary adrenal (HPA) response to a variety of hypothalamic stimuli, but have augmented systemic secretion of AVP. Patients with rheumatoid arthritis (RA) have deficient CS with exaggerated PRL responses to inflammatory stimuli. Within inflammatory foci, CRH is pro inflammatory. AVP, which augments autologous mixed lymphocyte reactions, can replace the IL-2 requirement for gamma IFN production by T cells via V1a receptors, and potentiates primary antibody responses, is also pro-inflammatory. Lewis rats have significantly high plasma levels, hypothalamic content, and in vitro release of AVP in comparison to the inflammatory disease-resistant Fischer rats. Immunoneutralization of AVP attenuates inflammatory responses. In Sprague Dawley rats, AVP potentiates PRL secretion. Preliminary studies in patients with RA have shown that the circulating levels of AVP are significantly increased, which might be a compensatory response to low CS levels or a result of elevated levels of IL-6 in these patients but could nevertheless contribute to rheumatoid inflammation. A similar observation has been made in patients with ankylosing spondylitis. PMID- 11268413 TI - The hypothalamic-pituitary-adrenal and gonadal axes in rheumatoid arthritis. AB - The hypothalamic-pituitary-adrenal (HPA) and the hypothalamic-pituitary-gonadal (HPG) axes involvement or response to immune activation seems crucial for the control of excessive inflammatory and immune conditions such as autoimmune rheumatic diseases, including rheumatoid arthritis (RA). However, female patients seem to depend more on the HPA axis, whereas male patients seem to depend more on the HPG axis. In particular, hypoandrogenism may play a pathogenetic role in male RA patients because adrenal and gonadal androgens, both products of the HPA and HPG axes, are considered natural immunosuppressors. A significantly altered steroidogenesis of adrenal androgens (i.e., dehydroepiandrosterone sulfate, DHEAS and DHEA) in nonglucocorticoid-treated premenopausal RA patients has been described. The menopausal peak of RA suggests that estrogens and/or progesterone deficiency also play a role in the disease, and many data indicate that estrogens suppress cellular immunity, but stimulate humoral immunity (i.e., deficiency promotes cellular Th1-type immunity). A range of physical and psychosocial stressors are also implicated in the activation of the HPA axis and related HPG changes. Chronic and acute stressors appear to have different actions on immune mechanisms with experimental and human studies indicating that acute severe stressors may be even immunosuppressive, while chronic stress may enhance immune responses. The interactions between the immunological and neuroendocrine circuits is the subject of active and extensive ongoing research and might in the near future offer highly promising strategies for hormone-replacement therapies in RA. PMID- 11268414 TI - The role of pro- and antiinflammatory cytokines in neurodegeneration. AB - Experimental and clinical damage to the brain leads to rapid upregulation of an array of cytokines predominantly by glia. These cytokines may exert neurotoxic or neuroprotective actions. This paper will focus on the pro-inflammatory cytokine interleukin-1 (IL-1), which participates in diverse forms of brain damage including ischemia, brain trauma, and excitotoxic injury. Administration of low doses of IL-1 markedly exacerbates these forms of brain damage, whereas blocking IL-1 release or actions reduces neuronal death. IL-1 receptor antagonist (IL-1ra) is also upregulated by brain damage (mainly by neurons) and acts as an endogenous inhibitor of neurodegeneration, presumably by blocking IL-1 actions on its receptor. We have studied the actions of both IL-1 and IL-1ra in experimental models of ischemic and neurotoxic injury in rats, and have found site-specific effects within the striatum. On the basis of this and further work, we propose that IL-1 can exacerbate cell death in these conditions by modifying polysynaptic anterograde pathways leading from the striatum to the cortex. The precise nature of these pathways remains undetermined, as do the underlying mechanisms by which IL-1 can exert its effects, but appear to involve induction of IL-1 in specific brain regions and activation of cortical glutamatergic pathways. PMID- 11268415 TI - Environmental factors favoring the allergen-specific Th2 response in allergic subjects. AB - Allergen-reactive type 2 helper T cells (Th2) play a triggering role in the activation and/or recruitment of IgE antibody-producing B cells, mast cells, and eosinophils, i.e., the cellular triad involved in the allergic inflammation. Interleukin (IL)-4 production at the time of antigen presentation to the Th cell is critical for the development of Th2 cells. Other cytokines, such as IL-1 and IL-10, and hormones, such as calcitriol and progesterone, also play a positive role. In contrast, cytokines such as interferon (IFN)-alpha, IFN-gamma, IL-12, and transforming growth factor (TGF)-beta, and relaxin play a negative regulatory role on the development of Th2 cells. However, the mechanisms underlying the preferential activation by environmental allergens of Th2 cells in atopic individuals still remain obscure. Some gene products selectively expressed in Th2 cells or selectively controlling the expression of IL-4 have recently been described. Moreover, cytokines and other gene products that dampen the production of IL-4, as well as the development and/or the function of Th2 cells, have been identified. These findings allow us to suggest that the upregulation of genes controlling IL-4 expression and/or abnormalities of regulatory mechanisms of Th2 development and/or function may be responsible for Th2 responses against common environmental allergens in atopic subjects. PMID- 11268416 TI - Conditioning rhinitis in allergic humans. AB - A classical Pavlovian paradigm pairing an olfactory cue with allergen challenge for a single training trial was used to produce conditioned histamine release and conditioned nasal airflow decrease in seasonal allergic rhinitis sufferers. There was no conditioned increase in subjective symptoms. Histamine release and airflow decrease showed evidence of extinction by a second test trial. A second study comparing the effects of the number of training trials showed that three training trials produced greater histamine release and airflow decrease than a single training trial, suggesting stronger effects with additional training. PMID- 11268417 TI - Androstenetriol and androstenediol. Protection against lethal radiation and restoration of immunity after radiation injury. AB - Androstenetriol (AET) and Androstenediol (AED) upregulate host immunity, leading to increased resistance against infections. AET augments IL-2, IL-3, IFN gamma levels, and counteracts hydrocortisone immune suppression. AET and AED at a dose of 0.75 mg/- and 8.0 mg/25-g mouse, protected 60 and 70%, respectively, of C57/BL/6J mice irradiated with a lethal dose. These hormones also protected mice irradiated with 6 Gy and infected with a coxsackievirus B4 LD50. AET significantly increased spleen lymphocyte numbers at 7, 14, and 21 days after a 6 Gy exposure. Fluorescent activated cell-sorter analysis of irradiated mice, spleen, and bone marrow showed that AET significantly augmented the myeloid precursor markers, CD11b/Mac-1, and B220 (pan B), as well as the absolute numbers of CD4+/CD8+ cells over the 21 days of testing. Overall, the data are consistent with AET/AED inducing a more rapid recovery of all hematopoietic precursors from the small number of surviving stem cells. PMID- 11268418 TI - Altered glucocorticoid regulation of the immune response in the chronic fatigue syndrome. AB - It is increasingly recognized that glucocortiocoids (GCs) can have subtle modulatory effects in immunoregulation rather than having generalized immunosuppressive effects. GCs suppress Th1 cells and cellular immunity, but may favor Th2 responses and humoral immunity. The chronic fatigue syndrome (CFS) appears to be associated with a disturbed HPA-axis. Moreover, CFS patients show several immunological changes suggestive of decreased cellular immunity. It is postulated herein that in CFS patients a decreased Th1/Th2 balance may be the result of selective effects of GC on the IL-10/IL-12 regulatory circuit. PMID- 11268419 TI - Acute stress enhances while chronic stress suppresses skin immunity. The role of stress hormones and leukocyte trafficking. AB - Delayed-type hypersensitivity (DTH) reactions are antigen-specific, cell-mediated immune responses that, depending on the antigen, mediate beneficial (resistance to viruses, bacteria, fungi) or harmful (allergic dermatitis, autoimmunity) aspects of immunity. Contrary to the widely held notion that stress is immunosuppressive, we have shown that under certain conditions, stress can enhance immune function. DTH reactions can be studied in rats or mice by challenging the pinnae of previously sensitized animals with antigen. Studies have shown that acute stress administered immediately before antigen exposure significantly enhances skin DTH. In contrast, chronic stress significantly suppresses skin DTH. Stress-induced changes in leukocyte distribution may contribute to these bidirectional effects of stress, since acute stress induces a significant mobilization of leukocytes from the blood to the skin, whereas chronic stress suppresses leukocyte mobilization. In order to identify the hormonal mediators of the observed effects of stress, we first showed that adrenalectomy (ADX) eliminates the stress-induced enhancement of DTH. Acute administration (to ADX animals) of low doses of corticosterone and/or epinephrine significantly enhances skin DTH. In contrast, acute administration of high doses of corticosterone, low doses of dexamethasone, or chronic administration of moderate doses of corticosterone, suppress skin DTH. Thus, the timing and duration of stress may significantly affect the nature (enhancing versus suppressive) of the effects of stress on skin immune function. These results suggest that during acute stress, stress hormones may help enhance immune function by informing the immune system about impending challenges (e.g., wounding or infection) that may be imposed by a stressor (e.g., an aggressor). Thus, during acute stress, the brain may send a warning signal to the immune system, just as it does to other fight/flight systems in the body. PMID- 11268420 TI - The role of oxidative stress in viral infections. AB - Oxidative stress is implicated as a pathogenic factor in a number of viral infections. Our work has shown that nutritionally induced oxidative stress exacerbates the pathogenesis of coxsackievirus B3 (CVB3) infection in mice. Of particular note, mice fed on a diet deficient in antioxidants developed myocarditis when infected with a normally benign strain of CVB3. This change in virulence was found to be due to changes in the viral genome. Immune functions of the oxidatively stressed mice were also altered. Another example of the effect of oxidative stress on a viral pathogen took place in Cuba in the 1990s. An epidemic of optic and peripheral neuropathy in the population occurred that was associated with a lack of dietary antioxidants and with smoking (a pro-oxidant). A coxsackie like virus was isolated from the cerebrospinal fluid from 84% of patients cultured. Thus, oxidative stress can have profound effects, not only on the host, but on the pathogen as well. PMID- 11268422 TI - Chemical sympathectomy alters cytotoxic T lymphocyte responses to herpes simplex virus infection. AB - Numerous studies have sought to delineate the impact of neuroendocrine function on overall immune responsiveness. Using various murine models, we and others have previously shown that both adrenal-dependent and adrenal-independent mechanisms regulate components of the primary and memory cellular immune responses to herpes simplex virus type 1 (HSV-1) infection. We have extended these studies by determining the impact of 6-hydroxydopamine (6-OHDA)-induced peripheral sympathetic denervation on these responses. C57BL/6 mice treated with 6-OHDA (200 mg/kg) were inhibited in their ability to generate primary, HSV-specific cytotoxic T lymphocytes (CTL) in response to HSV infection. Sympathectomy also suppressed the activation and function of HSV-specific memory CTL (CTLm). In addition, administration of 6-OHDA resulted in a transient but substantial increase in levels of circulating corticosterone and hypothalamic Fos expression. Together, these findings suggest that peripheral sympathetic denervation may modulate immune function via activation of the hypothalamic-pituitary-adrenal (HPA) axis. PMID- 11268421 TI - Local regulation of glucocorticoid activity in sites of inflammation. Insights from the study of tuberculosis. AB - In sites of inflammation there is a change in the equilibrium between the enzymes that inactivate cortisol by conversion to cortisone and those that reactivate cortisone by conversion to cortisol. Current evidence suggests that during an immune response with a Type 1 cytokine profile such as tuberculosis, there is locally enhanced reductase activity with locally increased cortisol concentrations due to recruitment of cortisone. This results in enhanced cortisol mediated feedback on the inflammatory process, and deviation of the response towards Type 2. Preliminary data suggest that eventually, in the presence of Type 2 cytokine polarization, the enzyme equilibrium may reverse again and cortisol is then locally inactivated to cortisone. Together with changes in glucocorticoid receptor expression and function this may result in local cortisol resistance and susceptibility to tissue damage mediated by proinflammatory cytokines. These observations help to explain the sequence of events in several infectious, inflammatory and autoimmune diseases. PMID- 11268423 TI - Steroid hormone regulation of antiviral immunity. AB - Recent observations in both humans and animals have demonstrated that stress is immunomodulatory and can alter the pathogenesis of microbial infections to the extent that it may be adverse to health. Stress disrupts homeostasis, and the body responds through endocrine and nervous system interactions in an effort to re-establish the health of the host. However, the resulting physiologic changes associated with stress, such as the rise in serum glucocorticoids (GCs), are implicated in suppression of antiviral immunity. Therefore, it would be of significance to counterregulate stress-mediated immunosuppression during viral infection to improve immune responses and limit virus-mediated damage. The data in this study focus upon the antiglucocorticoid influence of a native steroid hormone that has been shown to augment immune function and protect animals against lethal viral infections. Androstenediol (5-androstene-3 beta,17 beta diol, AED), a metabolite of dehydroepiandrosterone (DHEA), confers protection against lethal infection with influenza A virus. The protective activity appears to counterbalance the function of the regulatory GCs because AED prevents GC mediated suppression of IL-1, TNF-alpha, and IL-2 secretion. Furthermore, AED inhibits GC-induced transcription of a GC-sensitive reporter gene. PMID- 11268424 TI - Neuroendocrine regulation of IL-12 and TNF-alpha/IL-10 balance. Clinical implications. AB - Interleukin-12 and tumor necrosis factor (TNF)-alpha promote T-helper (Th) 1 responses and cellular immunity, whereas IL-10 suppresses Th1 activities and stimulates Th2 and humoral immune responses. Recent evidence indicates that glucocorticoids, norepinephrine, epinephrine, histamine, and adenosine inhibit the production of human IL-12 and TNF-alpha, whereas they do not affect or even stimulate the production of IL-10. Through this mechanism these neuroendocrine mediators may cause a selective suppression of Th1 responses and a Th2 shift rather than generalized Th suppression. The substantial Th2-driving force of endogenous stress mediators, as well as histamine and adenosine, can be amplified to a great extent during certain conditions and may play a role in increased susceptibility of the organism to various infections that are normally cleared by Th1 responses. In addition, conditions that contribute to a substantial increase or decrease of local or systemic concentrations of these mediators via modulation of IL-12, TNF alpha/IL-10 balance may also play a role in induction, expression, and progression of certain autoimmune diseases, allergic/atopic reactions, and tumor growth. These conditions include: acute or chronic stress; cessation of chronic stress or chronic hypoactivity of the stress system; severe exercise; serious surgical procedures or traumatic injuries; major burns; severe ischemia or hypoxia; pregnancy and the postpartum period. Thus, better understanding of the neuroendocrine regulation of IL-12, TNF-alpha/IL-10 balance might help the development of new therapeutic strategies for the treatment of Th1- and Th2 mediated human diseases. PMID- 11268425 TI - CNS penetration by noninvasive viruses following inhalational anesthetics. AB - The effects of inhalational anesthetics on brain penetration by the neurovirulent noninvasive West Nile virus (WN-25) were studied in mice. WN-25 injected intracerebrally causes encephalitis and kills adult mice, but when injected intraperitoneally (i.p.) it is unable to invade the brain and kill. Under stress conditions, this strain causes encephalitis and death even after i.p. inoculation. In the study described in this paper, we used two inhalational anesthetics, a single short-term exposure to 2% halothane for 10 min in oxygen, or 70% nitrous oxide (N2O) for 30 min in air. Both inhalational anesthetics induced WN-25 encephalitis and death in 33% and 20% of the tested mice, respectively. Exposure of inoculated mice to halothane for prolonged periods or for repeated exposures (two or three times) markedly increased the mortality rate (up to 75%). Exposure to 30% CO2, a known modulator of blood-brain barrier (BBB) activity, was used as a positive control (80% mortality). No death was observed in the control non-exposed injected mice. Virus levels were found to be more than 10(7) plaque-forming units (PFU)/brain in all moribund mice. Additional parameter demonstrating the "stressor-like" nature of inhalation anesthetics was the induction of a significant decrease in weight of the lymphoid organs of inoculated mice. We suggest that inhalational anesthetics induces BBB breaching with subsequent entrance of the noninvasive WN-25 virus into the brain, causing encephalitis and death. PMID- 11268426 TI - Fat redistribution in HIV-infected patients. A new hormonal-immune disorder? AB - Multidrug antiretroviral regimes in HIV-infected patients may have side effects. The most frequent side effects are changes in fat metabolism and distribution. We describe a particular pattern of fat redistribution (FR), characterized by a progressive enlargement of breast and abdominal girth and fat loss in the lower limbs, which occurs in approximately 10% of HIV-infected women treated with combined antiretroviral therapy. To elucidate the metabolic, endocrine, and immunologic consequences of the observed disturbance, we measured serum lipids, glucose, C-peptide, ACTH, plasma, urinary cortisol, and cytokines IL-2, IFN gamma, Il-4, IL-10, Il-12, and TNF alpha in 36 patients with FR and in a control group without FR. There were no significant differences in hormonal and metabolic laboratory testing between the two groups. Immunology studies showed that in vitro production of TNF alpha and IL-10 was lower and IL-12 production higher in SR patients. Whether or not such immune alterations may be reponsible or be caused by fat redistribution remains to be explained. One year after the follow up, 50% of the patients treated with triple therapy developed lipodystrophy, characterized by weight loss, face-wasting, and hyperglycemia; the remaining 50% remained unchanged. In 13 patients the 3TC withdrawal was followed by improvements of the syndrome in 50% and of lipodystrophy in about 25%. These data suggest that the FR syndrome is frequent in patients treated with 3TC and that it is associated with characteristic changes in the cytokine production. PMID- 11268427 TI - Immunoendocrinologic abnormalities in human immunodeficiency virus infection. AB - Alterations in the production of adrenal steroids and a complex pattern of dysregulation in cytokine profiles accompany the progression of HIV infection. Cortisol levels increase in HIV infection, while those of dehydroepiandrosterone (DHEA), a physiologic antagonist of the immunoregulatory activities of cortisol, decrease. A shift from type-1 to type-2 cytokine production is also detected in most patients during disease progression. This shift is summarized as a defective production of interferon gamma (IFN gamma), interleukin-2 (IL), and IL-12 accompained by increased production of IL-4, IL-5, IL-6, and IL-10. IFN gamma and IL-2 are suppressed, while the generation of IL-4 is stimulated by cortisol and pharmacological doses of glucocorticoids (GC). GC and IL-4 stimulate the differentiation of B lymphocytes into IgE-producing plasma cells, the concentration of which augments in HIV infection. Finally, GC induces programmed cell death (PCD) in a variety of different cells, including mature T lymphocytes. Because (1) TH1 but not TH2 undergo rapid Fas-mediated PCD upon antigen stimulation, and (2) TH2 clones preferentially survive in vitro cell cultures, the progressive shift from type-1 to type-2 cytokine production observed in HIV infection could be at least partially provoked by the increase in the production of cortisol and the reduction of DHEA. Progression of HIV infection to AIDS can be controlled by highly active antiretroviral therapy (HAART); HAART drastically reduces HIV plasma viremia, but is less effective in immune reconstitution. Additionally HAART is associated in a sizable portion of patients by complex lypodistropyc phenomena that often involve the endocrine system. PMID- 11268428 TI - Changes in cortisol/DHEA ratio in HIV-infected men are related to immunological and metabolic perturbations leading to malnutrition and lipodystrophy. AB - HIV-1 infection is associated with immune deficiency and metabolic perturbations leading to malnutrition and lipodystrophy. Because immune response and metabolic perturbations (protein and lipid metabolism) are partly regulated by glucocorticoids and DHEA, we determined serum cortisol and DHEA concentrations, and the cortisol/DHEA ratio in HIV-positive men, either untreated or receiving various antiretroviral treatments (ART), including highly active antiretroviral therapy (HAART). Cortisol levels were found increased in all patients, whatever the stage of the disease and independently of the ART treatment. In contrast, serum DHEA was elevated in the asymptomatic stage, and it was below normal values in AIDS patients, either untreated or mono-ART-treated. The DHEA level was low in HAART-treated patients with lipodystrophy (LD+) and highly increased in HAART treated patients without lipodystrophy (LD-). Consequently, the cortisol/DHEA ratio was similar to controls in asymptomatic untreated or mono-ART-treated patients, but increased in AIDS patients. Interestingly, this ratio was increased in LD+ HAART-treated men, but normalized in LD- HAART-treated patients. Changes in the cortisol/DHEA ratio were negatively correlated with the in vivo CD4 T-cell counts, with the malnutrition markers, such as body-cell mass and fat mass, and with the increased circulating lipids (cholesterol, triglycerides, and apolipoprotein B) associated to the lipodystrophy syndrome. Our observations show that the cortisol/DHEA ratio is dramatically altered in HIV-infected men, particularly during the syndromes of malnutrition and lipodystrophy, and this ratio remains elevated whatever the antiretroviral treatment, including HAART. These findings have practical clinical implications, since manipulation of this ratio could prevent metabolic (protein and lipid) perturbations. PMID- 11268429 TI - 7 alpha-hydroxy-dehydroepiandrosterone and immune response. AB - In human and murine lymphoid organs, circulating 3 beta-hydroxysteroids, including pregnenolone (PREG), dehydroepiandrosterone (DHEA), and epiandrosterone (EPIA), are 7 alpha-hydroxylated by a cytochrome P450 identified in the hippocampus as P4507B1. Mouse and human lymphoid organs produced different patterns of 3 beta-hydroxysteroid 7 alpha-hydroxylation with the absence of pregnenolone and epiandrosterone hydroxylation in human and mouse, respectively. Both 7 alpha-hydroxy-DHEA and 7 alpha-hydroxy-EPIA triggered a significant increase of antitetanus toxoid and anti-Bordetella pertussis toxins IgGs production in cultures of activated B + T cells derived from human tonsils, whereas both 7 alpha-hydroxy-PREG and 7 alpha-hydroxy-DHEA increased the immune response in mouse. Paracrine action of 7 alpha-hydroxysteroids resulted from their production in cells of the lymphoid organs. Comparison of P4507B1 sequences in rat, human, and two mouse species showed that one amino acid change might explain important differences in KM for 7 alpha-hydroxylation, and suggested that such differences might contribute to the extent of immune response. PMID- 11268430 TI - Antiemetic effect of ramosetron during hyperthermo-chemo-radiotherapy for esophageal cancer. AB - BACKGROUND: The antiemetic effects of serotonin receptor antagonists during chemoradiotherapy for solid tumors have never been reported. We have developed hyperthermo-chemo-radiotherapy (HCR) for esophageal cancer. However, with this treatment, the more potent the chemotherapy was, the more frequently emesis was experienced. MATERIALS AND METHODS: Fifteen patients with esophageal cancer underwent HCR (6 courses of hyperthermia, cisplatin 20 mg/m2 x 6, 5-FU 300 mg/m2 x 15 and radiation 1.5 Gy x 30). Ramosetron was administered intravenously (0.3 mg x 15). The emesis inhibition rate was defined as the rate of patients having neither vomiting nor severe nausea. RESULTS: The incidence of patients without nausea gradually decreased to 60% at the end of chemotherapy. However, vomiting was completely avoided except in one patient for two days. The emesis inhibition rates of weeks 1, 2, 3 and 4 were 100.0, 93.3, 89.5 and 95.2%, respectively. The overall inhibition rate was 94.5% and the rate of "well inhibited" was 79.0%. There were no ramosetron-related adverse reactions. CONCLUSIONS: These findings suggest that ramosetron is a useful antiemetic agent for nausea and vomiting induced by chemoradiotherapy for solid tumors. PMID- 11268431 TI - Correlations between serum progesterone and smoking, and the growth fraction of cervical squamous cell carcinoma. AB - BACKGROUND: Possible correlations between growth fraction of squamous cervical carcinomas and serum progesterone (se-P) concentrations, smoking habits and DNA ploidy were studied. MATERIALS AND METHODS: The DNA S-phase fraction (SPF), measured by flow cytometry was used as a marker of tumour growth in 103 cases of squamous cervical cancer stage IB-IV. DNA-ploidy (peridiploidy vs. aneuploidy), Se-P, se-Estradiol, smoking habits, parity, menopausal status, clinical stage and histopathological grading were compared to SPF < 14% vs. SPF > or = 14%. RESULTS: Aneuploidy, (odds ratio (OR) 10.0), se-P > or = 2.6 nmol/l (OR 7.5) and smoking (OR 3.0) were significantly associated with SPF > or = 14%, after adjustments for all factors included in the study. The association with se-P and smoking was attributed to an increased risk for the premenopausal women in the study. DISCUSSION: In this study an increased tumour growth was associated with increased leves of se-P, smoking and aneuploidy in women with invasive squamous cervical carcinoma. This study seems to experimentally confirm epidemiological studies, where smoking and long-term use of oral contraceptives have been linked to cervical neoplasms. PMID- 11268432 TI - Detection of esophageal carcinoma using single photon emission computed tomography with technetium-99m tetrofosmin. AB - Technetium-99m tetrofosmin (Tc-TF) a thallium-201 (Tl-201) and technetium-99m methoxy-isobutyl-isonitrile (Tc-MIBI) competitor, being used as a new radiopharmaceuticalfor myocardial imaging was assessed to understand its value in detecting esophageal carcinoma (Eso-Ca). Forty patients with Eso-Ca underwent Tc TF single photon emission computed tomography (SPECT) of the chest. Meanwhile 15 controls also accepted Tc-TF SPECT of the chest for comparison. Among the 40 patients Tc-TF chest SPFCT detected Eso-Ca in 33 (82.5%) but not in 7 (17.5%). In contrast all 15 normal controls (100.0%) had negative Tc-TF chest SPECT Detection sensitivities were higher for tumors located in the middle portion of the esophagus for epidermoid ca and for tumors with poor differentiation when compared with tumors in the upper or lower portion for adenocarcinoma and tumors with moderate differentiation. However, the differences were not significant (p values < 0.05). Significantly higher detection sensitivity was found for tumors of larger size than for tumors of smaller size (p value < 0.05). Our results suggest that Tc-TF chest SPECT is a helpful test for detecting Eso-Ca especially larger tumors. PMID- 11268433 TI - A phase I study of 90Y-2IT-BAD-Lym-1 in patients with non-Hodgkin's lymphoma. AB - BACKGROUND: Prior clinical trials proved that all histologic grades of chemotherapy-resistant B-cell non-Hodgkin's lymphoma (NHL) could respond to radio immunotherapy (RIT) with 131I-Lym-1 and 67Cu-2IT-BAT-Lym-1. This Phase I study was conducted to determine the safety and maximum tolerated dose (MTD) of 90Y-2IT BAD-Lym-1. METHODS: Lym-1 is a mouse monoclonal antibody that preferentially targets malignant B-lymphocytes. 90Y has beta emissions suitable for therapy but no gamma emissions, therefore, 111In-2IT-BAD-Lym-1 is used for imaging. The macrocyclic chelator, DOTA, bound 90Y tightly to form a stable drug. Patients with chemotherapy-resistant NHL received 90Y-2IT-BAD-Lym-1 at administered doses of: 0.185, 0.278, or 0.370 GBq/m2. RESULTS: Myelotoxicity, especially thrombocytopenia, was dose-limiting. No significant non-hematologic toxicity occurred. Human anti-mouse antibody (HAMA) developed in 3/8 patients. The mean radiation dose to the 33 imaged tumors was 7.0 Gy/GBq. Lung, kidney and liver received mean radiation doses of 1.3, 2.4, and 6.4 Gy/GBq, respectively from 90Y 2IT-BAD-Lym-1. The tumor: body and tumor:bone marrow (by imaging) ratios were 16.4:1 and 5.8:1, respectively. In this Phase I study, 5/8 patients that failed prior chemotherapy had a partial response or stabilization of NHL after RIT. CONCLUSION: The safety and toxicity of 90Y-2IT-BAD-Lym-1 were determined and the MTD was 0.370 GBq/m2, a dose used in 4 patients. 90Y-2IT-BAD-Lym-1 may be useful for future clinical trials because 90Y is readily available and can deliver potent beta emissions to NHL. Bone marrow support however, will be required for further dose escalation. PMID- 11268434 TI - Chemosensitivity of breast cancer lymph node metastasis compared to the primary tumor from individual patients tested in the histoculture drug response assay. AB - Lymph node metastasis is often the first indication of the aggressiveness of breast cancer. Effective chemotherapy in breast cancer depends on targeting the metastatic component of the disease. In order to optimize chemotherapy in the metastatic target of breast cancer, the histoculture drug response assay (HDRA) was performed on surgical specimens of primary tumor and axillary lymph node metastasis from 30 breast cancer patients. The surgical specimens were cut into approximately 10 mg pieces, and placed onto the collagen gel sponges in the medium containing previously-determined cutoff concentrations of doxorubicin (DXR), 5-fluorouracil (5-FU), cisplatin (DDP), and mitomycin C (MMC). After incubation for 7 days, the chemosensitivity of the tumor fragments was evaluated with the 3-(4,5-dimethythiazol2yl)-2,5-diphenyl-2H tetrazolium bromide (MTT) endpoint. The lymph node metastases were more resistant than the primary tumor for DXR, 5-FU, and MMC (p < 0.05) but not for CDDP. The data suggest that both primary tumor and metastases from individual patients should be tested in the HDRA to enhance clinical efficacy of chemotherapy. PMID- 11268435 TI - Toremifene as a substitute for adjuvant tamoxifen in breast cancer patients. AB - BACKGROUND: Toremifene is a new antiestrogen, which in nonclinical studies appears less carcinogenic than tamoxifen. Clinical trials of adjuvant toremifene vs. tamoxifen in breast cancer patients are ongoing. This study aimed to evaluate the short-term effects of changing from adjuvant tamoxifen to toremifene. PATIENTS AND METHODS: Twenty postmenopausal breast cancer patients receiving adjuvant tamoxifen, 20 mg/day, were switched to toremifene 60 mg/day. The effects on the uterus were evaluated prospectively by transvaginal ultrasound; tolerability was assessed clinically. RESULTS: In 14 patients who had uterine abnormalities (endometrial thickening or polyps) under tamoxifen, no significant changes occurred during a median of 18 months (range 7-24) of toremifene treatment. Out of six patients who had entered the study due to intolerance to tamoxifen, however, 3 tolerated toremifene well. CONCLUSION: Toremifene does not modify previous uterine changes induced by tamoxifen. For some patients who do not tolerate tamoxifen, however, switching to toremifene may allow the continuation of adjuvant antiestrogenic therapy. PMID- 11268436 TI - Docetaxel, a promising novel chemotherapeutic agent in advanced breast cancer. AB - Clinical practice in chemotherapy of breast cancer is undergoing changes. This study investigated the efficacy and toxicity of docetaxel, a novel chemotherapeutic agent in metastatic breast cancer. Focus was on the effect of the cumulative dose of previous anthracycline treatment on response rate, toxicity and survival in our own patients; published data were reviewed. PATIENTS AND METHODS: Thirty-one women, (median age 52 years, range 40-65) treated for metastatic breast cancer with docetaxel were included. RESULTS: The overall response rate was 48%, with 3 complete and 11 partial responses (95% CI 29-66). The duration of response was 7 months (range 2 to 16 months), the median overall survival after docetaxel 13.7 months for responding patients, 14.3 months in no change patients and 6.5 months in patients with progressive disease. The mean cumulative anthracycline dose prior to docetaxel was 860 mg (range 200-1760 mg); in the case of responders, the previous cumulative total epirubicin doses were 200-1575 mg (median 766 mg.). Total dose or schedule of previous epirubicin treatment had no impact on docetaxel response rate, toxicity or survival. The response seen in this study is within the published range (24 to 60%) observed for docetaxel in anthracycline-treated patients. CONCLUSION: We conclude that docetaxel is active in metastatic breast cancer even as third- line treatment. Previous treatment with, or response to, epirubicin does not influence the response to docetaxel and this promising new drug is currently being tested for adjuvant use in breast cancer. PMID- 11268437 TI - Tumor volumetry: proposal of a new concept to predict lymph node metastasis in early gastric cancer. AB - BACKGROUND: The present study was conducted to evaluate the significance of tumor volumetry in early gastric cancer for predicting lymph node metastasis. MATERIALS AND METHODS: [corrected] Computer-generated three-dimensional images of tumors from 147 patients with early gastric cancer, who underwent curative gastrectomy with lymphadenectomy, were reconstructed from continuous tissue sections and the volume (mm3) of each tumor was measured by the surface rendering method. Logarithmic tumor volume and conventional pathological factors were then studied with respect to the prevalence of metastasis to regional lymph nodes. RESULTS: The results of univariate analysis showed that lymph node metastasis was associated with a larger tumor diameter, larger logarithmic tumor volume, a higher rate of blood and lymphatic vessel invasion and a higher incidence of submucosal invasion. However, the results of logistic regression analysis showed that only two factors, logarithmic tumor volume and blood vessel invasion, were independent variables that correlated with lymph node metastasis. Moreover, logarithmic tumor volume was the most important variable correlated with lymph node metastasis in early gastric cancer (p = 0.004, odds ratio: 23.831). CONCLUSIONS: Based on the present results, tumor volumetry appears to represent a novel method of predicting the likelihood of lymph node metastasis in early gastric cancer. PMID- 11268438 TI - Gemcitabine as single agent in the treatment of elderly patients with advanced non small cell lung cancer. AB - BACKGROUND: In phase II trials gemicitabine proved to be an active agent in NSCLC, producing a clinical benefit, often higher than response rate. PATIENTS AND METHODS: We assessed the impact of gemcitabine treatment on response rate and quality of life in 21 untreated elderly patients (aged > 70 years) with NSCLC. Gemcitabine (1250 mg/sm) was administered days 1-8 every 21 days. Response and toxicity were analyzed according to WHO criteria. The assessment of quality of life was performed by analysing a disease symptom related questionnaire completed by the patient. RESULTS: All the patients were evaluable: we found 7 PR (33%), 5 SD (24%) and 9 PD; the median duration of response was 24 weeks; the median overall survival 32 weeks; WHO grade 2 leukopenia (in 4 patients) and thrombocytopenia (grade 3 in 1 patient and grade 2 in two patients) were the main toxic effects. A clinical benefit was demonstrated in all 12 patients with PR or SD and in 3 patients with PD. CONCLUSIONS: These data confirm that gemcitabine is a well tolerated and active therapeutic approach in elderly non small cell lung cancer patients. PMID- 11268439 TI - Microalbuminuria during cisplatin therapy: relation with pharmacokinetics and implications for nephroprotection. AB - BACKGROUND: To assess the relation of cisplatin-induced nephrotoxicity to its pharmacology. PATIENTS AND METHODS: In 22 chemonaive patients (median age, 32 years) receiving 100-150 mg/m2 cisplatin for a total of 54 courses of therapy pharmacokinetics of ultra-filtrable platin were analyzed. Nephrotoxicity was sensitively assessed by nephelometric analyses of urinary marker-proteins. RESULTS: The parameters calculated for ultrafiltrable platin were (two compartment-model): terminal half-life, 36 hours (coefficient of variation [CV], 22%); AUC, 12852 ng h/ml (33%); volume of distribution, 3531 (44%); total clearance, 285 ml/min (30%); renal clearance, 149 ml/min (23%); maximum concentration, 1720 ng/ml (66%); renal elimination, 57% of applied dose (26%). A pathological urinary excretion of albumin > 20 mg/l and alpha-1-microglobulin > 10 mg/l was detected in 39 out of 54 and 42 out of 54 cycles, respectively. The degree of albuminuria was related with urinary monoaquoplatin concentrations (p = 0.003). CONCLUSION: Nephrotoxicity of cisplatin appears to depend on the urinary monoaquoplatin concentrations which may be modulated by application of saline. PMID- 11268440 TI - Breast cancer screening by mammography in women aged under 50 years in Japan. AB - BACKGROUND: The effectiveness of mammographic screening in women aged over 50 years has been confirmed in the United States and Europe, but its effectiveness in women aged from 40 to 49 years remains controversial. The optimum age for effective screening of subjects for breast cancer by mammography in Japan was studied based on the results of mammographic screening. METHOD: The benefit of breast cancer screening in women was examined by stratifying the results of mammographic screening in a Tokushima trial on the basis of age: under 50 years and 50 years or older. The results of conventional screening by physical examination alone, which we performed in a Zentsuji trial, were used as the control. RESULTS: The examinees numbered 13,982 and 18,619 in mammographic screening and screening by physical examination, respectively. Breast cancer was detected in 43 and 22 patients, respectively. The detection rate of breast cancer was 0.31% by mammographic screening, which is about 3 times higher than that (0.12%) by screening using physical examination. Mammographic screening thus showed significantly higher sensitivity (93.5% vs 73.3%, p = 0.015). The proportion of stage I cancer and the absence of nodal involvement were 67.4% and 79.1% by mammographic screening, compared with 31.8% and 59% by physical examination. Our results obtained with mammographic screening were equal to or higher than the results obtained in the United States and Europe. The clinical stage of the breast cancers detected by mammographic screening in the subjects aged under 50 years was stage 0 (DCIS) in one case and stage I in 10 cases, while the group aged 50 years or older showed stage 0 in 11 cases and stage I in 19 cases. There were three cases of false-negative; two false-negative cases were aged under 50 year, while one case was aged 50 years or older. The detection rates of cancer in the group under 50 years and that of 50 years or more were 0.19% and 0.39% by mammograpic screening and 0.09% and 0.15% by the physical examination. The sensitivities in the group under 50 years and that of 50 years or more were 84.6% and 97.0% by mammographic screening and 72.7% and 73.7% by physical examination, showing no significant difference. In the results of mammograms by Wolfe's classification with respect to the age groups, the proportion of DY (dense breast) pattern decreased significantly from 3.5% to 0.2% in women of 50 years or more and from 16.6% to 2.4% in those under 50 years when the values were compared between the period from 1992 to 1995 and the period from 1998 to 1999, respectively (p < 0.0001). CONCLUSION: The above findings suggested the possible effectiveness of mammographic screening not only in women aged 50 years or more but also in those aged under 50 years, in Japan. Therefore, introduction of mammography should be considered at an early date, even for women aged from 40 to 49 years. PMID- 11268441 TI - Surgical outcome of node-positive early gastric cancer with particular reference to nodal status. AB - BACKGROUND: The risk of recurrence according to nodal status in patients with node-positive early gastric cancer (EGC) remains unclear and no appropriate treatment approaches have yet been established for such patients. MATERIALS AND METHODS: The surgical outcome of gastrectomy in combination with lymphadenectomy was examined in a total of 100 patients (54 males and 46 females, ranging in age from 25 to 84 years; average 56.6 years) with EGC and metastasis to lymph nodes. The outcome was assessed with particular reference to the extent of lymph node metastasis. RESULTS: The 5 and 10-year overall survival rates were 93.5 and 89.8%, respectively. Significant differences in survival were detected when anatomical distribution of lymph node metastasis (p < 0.0001), number of positive nodes (p = 0.0004) and tumor size (p = 0.0085) were examined. In particular, in 73 patients for whom the metastasis was limited to a perigastric node, prognosis was excellent and no recurrence was observed during the follow-up period. On the other hand, 27 patients with metastasis to a lymph node beyond the perigastric region were defined as comprising a high risk group for recurrence among node positive EGC patients due to their poor prognosis (10-year survival rate, 58.5%). CONCLUSION: The results of the present study have suggested that radical gastrectomy combined with lymphadenectomy is essential to achieve complete remission in patients with lymph node metastasis restricted to perigastric nodes. For patients with a high risk of recurrence in EGC, whose condition is complicated by lymph node metastasis beyond the perigastric region, care should be taken to prevent recurrence by conducting long-term follow-up even after radical surgery. In order to improve survival, an appropriate protocol for post operative adjuvant therapy may be needed for patients such as those with advanced gastric cancer. PMID- 11268442 TI - Atypical squamous and glandular cells of undetermined significance (ASCUS and AGUS) of the uterine cervix. AB - ASCUS (Atypical Squamous Cells of Undetermined Significance) and AGUS (Atypical Glandular Cells of Undetermined Significance), or AGCUS, are two acronyms introduced in 1988 by The Bethesda System (TBS) for reporting borderline cytological changes in cervical cytology. ASCUS and AGUS categories should be subclassified. Five ASCUS subgroups were proposed: 1) ASCUS due to processing defects, 2) with "mature" cytoplasm, 3) in post-menopausal women (a--in the setting of atrophy and b--with estrogen stimulation), 4) atypical metaplasia, and 5) ASCUS with keratinized cytoplasm. AGUS subgroups may be subcategorized in endometrial or endocervical on the basis of origin. Endocervical AGUS should be further qualified, but the analysis of atypical glandular cells may be really difficult and the conclusive diagnosis is frequently "AGUS not otherwise specified". The subclassification of ASCUS and AGUS is useful for an appropriate clinical management, but pertinent patient information (such as age, date of last menstrual period, mechanical therapies, tamoxifen therapy, and others) is needed to avoid an overdiagnosis and consequently an overtreatment. In fact various subgroups require different clinical management. Therefore, an effective communication between cytopathologists and referring physicians is essential in the analysis of squamous and glandular atypias. PMID- 11268443 TI - Histological features of cirrhosis with hepatitis C virus for prediction of hepatocellular carcinoma development; a prospective study. AB - The histological features of pre-neoplastic lesions in HCV-associated cirrhosis remain uncertain. The aim of this prospective study was to elucidate histological features for predicting the development of hepatocellular carcinoma (HCC). A cohort of 72 consecutive patients with hepatitis C-associated cirrhosis, which was diagnosed by histology investigated for development of HCC. Seven histological features including small cell dysplasia (SCD) and large cell dysplasia (LCD) of liver cirrhosis were evaluated with regard to the development of HCC. In addition, proliferation and apoptosis were investigated using immunohistochemistry by proliferating cell nuclear antigen and TUNEL method, respectively. At enrollment, SCD was observed in the biopsy specimens of 18 out of 72 (25.0%) patients and LCD was observed in 20 out of 72 (27.8%). Twenty eight out of 72 patients (38.9%) developed HCC during a mean follow-up period of 72.4 months. Among the histological parameters, SCD, active inflammation and complete nodule were statistically significant factors for the cumulative probability of developing HCC. However, LCD did not appear to be important for HCC development. In multivariate analysis, SCD was the highest independent risk factor for HCC. Samples with SCD demonstrated a higher proliferative rate and a lower apoptotic rate than normal hepatocytes or samples with LCD. These results indicate that SCD is a major risk factor for HCC. Careful assessment of liver histology may be important in order to predict HCC development in patients with HCV-related cirrhosis. PMID- 11268444 TI - Clinicopathological studies of esophageal carcinoma in achalasia: analyses of carcinogenesis using histological and immunohistochemical procedures. AB - Achalasia of the esophagus is a benign disease caused by dyskinesia of the lower esophagus and cardia and is presumed to be a premalignant lesion leading to an increased risk of squamous cell carcinoma. We analyzed six surgically or endoscopically resected carcinomas among 54 cases of esophageal achalasia using histological and immunohistochemical procedures. The mean interval between the diagnosis of achalasia and carcinoma was 21.5 years. Four of the six cases were superficial early-stage cancers whilst the other two were advanced cancers invading the adventitia. Histological mapping of the resected esophageal specimens demonstrated marked hyperplastic changes of stratified squamous epithelium and multiple foci of dysplastic changes. The squamous cell carcinomas showed well-differentiated type with low-grade atypia, closely associated with dysplastic foci. Immunohistochemical staining for p53, p21, p16 and epidermal growth factor receptor suggested that the dysplastic epithelium was a borderline lesion between hyperplasia and in situ carcinoma. Our observations suggested that esophageal food stasis induces chronic hyperplastic esophagitis and eventually malignant transformation of esophageal epithelial cells, associated with dysplasia-carcinoma sequence. PMID- 11268445 TI - The primary mediastinal growing teratoma syndrome. AB - We encountered a case of mediastinal immature teratoma which revealed the feature of the so-called growing teratoma syndrome. A 20-year-old male with a cough was discovered to have an abnormal shadow in the mediastinum. The serum AFP was elevated to 3600 ng/ml. The specimen with percutaneous needle biopsy revealed mature teratoma. The tumor was suspected to be mature teratoma with a malignant component because of the high level of serum AFP and he underwent chemotherapy. The serum AFP declined markedly but the tumor further enlarged. The resected tumor was diagnosed as immature teratoma, although most of the tumor tissue was mature component. PMID- 11268447 TI - Sarcoma of the maxillofacial region: follow-up of 25 cases. AB - BACKGROUND: Maxillofacial sarcomas are rare tumors, varying according to their type, grade, site of occurrence, response to treatment and prognosis. Approximately 10% of all cases of osteosarcoma, a subtype with a particularly poor prognosis, occur in the head and neck. PATIENTS AND METHODS: During the 35 years from 1963 until 1997, the 25 patients treated for sarcomas at our center were identified and evaluated according to a variety of parameters. RESULTS: Overall 2- and 5-year survival rates were 72% and 60%, respectively. Sarcomas were found most commonly in the mandible, maxilla and tongue. Survival was not significantly correlated with gender, ethnic origin or histopathological staging, but significant correlation was found between type of sarcoma and survival whilst younger patients had better survival rates. CONCLUSION: Advanced facial reconstruction methods may enable the performance of more radical ablative surgery, thereby improving survival outcome for head and neck osteosarcoma patients. Osteosarcoma appears to be as lethal in the maxillofacial region as in the extremities. PMID- 11268446 TI - Randomized controlled trial of 5-fluorouracil (5-FU) infusion combined with 1 hexylcarbamoyl-5-fluorouracil (HCFU) oral administration and HCFU alone as postoperative adjuvant chemotherapy for colorectal cancer. AB - BACKGROUND: Although surgical resectability is an important prognostic factor, recurrences are commonly noted in advanced colorectal cancer patients, even after apparently curative surgery. Because such recurrences cannot be cured, better adjuvant chemotherapies are urgently required. PATIENTS AND METHODS: We studied the effect of postoperative chemotherapy using 1-hexylcarbamoyl-5-fluorouracil (HCFU) oral administration with or without 5-fluorouracil (5-FU) infusion for curatively resected Stage II and III colorectal cancer. This study was prospectively randomized and controlled and 303 (95.6%) of 316 patients were determined to be candidates for statistical assessment. Group A received oral HCFU, 300 mg daily for 52 weeks beginning 2 weeks after surgery. Group B also received 5-FU intravenous injection, 333 mg/m2 body surface area/24 hours continuously for 72 hours beginning on postoperative day 0 and 6. RESULTS: There were no differences in overall 5-year survival or disease-free survival between Groups A and B. Group B had better 5-year disease-free survival (47.6%) than Group A (42.9%) (p = 0.062) and significantly prolonged interval from surgery to recurrence (p = 0.003) for patients with lymph node metastasis. In contrast, group B had significantly shortened 5-year disease-free survival (p = 0.010) and increased recurrence rate in patients without lymph node metastasis. CONCLUSION: Inductive therapy with 5-FU in combination with oral HCFU is beneficial as adjuvant chemotherapy for advanced colorectal cancer with lymph node metastasis. PMID- 11268448 TI - Results of fast neutron therapy of adenoid cystic carcinoma of the salivary glands. AB - BACKGROUND: Adenoid cystic carcinomas (ACC) seem to have a better response to fast neutron irradiation than to photon beam therapy because of the higher relative biological effectiveness of neutron radiation. METHODS: Between 1986 and 1995, 72 patients with ACC of the salivary glands were treated in Munster with fast neutrons. The median age was 54 years. All the patients had either recurrent or macroscopic rest tumor prior to neutron therapy. The median total dose was 15.03 Gy. Median follow-up was 50 months. RESULTS: 39.1% of the patients achieved a complete remission and 48.6% a partial remission. The survival probability was 86% after one year, 73% after two years and 53% after five years. The recurrence free survival was 83% after one year, 71% after two years and 45% after five years. CONCLUSION: Neutron beam therapy seems to have been an effective treatment in these selected patients. PMID- 11268449 TI - Technetium-99m methoxy-isobutyl-isonitrile chest single photon emission computed tomography to detect mediastinal lymph node metastasis in patients with non-small cell lung cancer: comparison with computed tomography. AB - This study evaluated the clinical role of Tc-99m-methoxyisobtylisonitrile (Tc MIBI) single photon emission computed tomography (SPECT) of the chest in the detection of mediastinal lymph node (MLN) metastasis in patients with non-small cell lung cancer (NSCLC). Twenty-five patients with proven NSCLC were enrolled in this study. Each of the patients received computed tomography (CT) of the chest and Tc-MIBI SPECT of the chest for presurgical staging. A postsurgical pathologic diagnosis was made and these patients were evaluated for the study of mediastinal lymph nde (MLN) involvement. Meanwhile, 10 volunteers also accepted Tc-MIBI SPECT of the chest for comparison. The results showed that the diagnostic sensitivity, specificity and accuracy of Tc-MIBI chest SPECT were 81.8%, 85.7% and 84% and for chest CT they are 36.3%, 85.7% and 64%, respectively. Our results indicated that Tc-MIBI chest SPECT was more sensitive and accurate than chest CT in the evaluation and detection of MLN involvement in the NSCLC patients. PMID- 11268450 TI - The role of technetium-99m methoxyisobutylisonitrile scintimammography in diagnosis of breast cancer in patients with mammographically dense breasts. AB - The purpose of this study was to assess the contribution of techentium-99m methoxyisobutylisonitrile (Tc-99m sestamibi) scintimammography in the diagnosis of breast cancer in 32 female patients with indeterminate mammographic probability of malignancy because of mammographically dense breasts. All the breast masses were removed and histopathological diagnoses were obtained in all cases. The results showed that scintimammography with Tc-99m sestamibi was positive in 21 patients (20 true-positive, 4 false-positive) and negative in 11 patients (7 true-negative, 4 false-negative). The diagnostic sensitivity, specificity and accuracy rates were 83%, 88% and 84%, respectively, in the differentiation of malignant from benign breast masses in the patients with mammographically dense breasts. The detection of malignant breast tumors by Tc 99m sestamibi scintimammography was independent of the density of the breast tissue. In conclusion, Tc-99m sestamibi scintimammography appears to be a useful diagnostic method for the detection of breast cancer in patients with non diagnostic mammograms because of mammographically dense breasts. PMID- 11268451 TI - Expression of apoptosis, cell proliferation, and drug resistance genes in pediatric Wilms' tumors. AB - The expression of genes associated with apoptosis, cell proliferation and drug resistance in tumor cells was investigated in two pediatric Wilms' tumor patients (MCH-WT-1 and MCH-WT-3) for their association with cell cycle, daunorubicin accumulation and clinical data. DNA content, cell cycle and drug accumulation were analyzed immediately after surgery by flow cytometry and mRNA expression by reverse transcriptase-polymerase chain reaction (RT-PCR) assay. Primary cell cultures were established from tumor specimens and tumor cells in both cases showed epithelial morphology. Although cell proliferation markers (Ki67 and PCNA) were expressed in both cases, MCH-WT-3 showed higher levels of mRNA expression, which corresponded, with metastatic behavior of the tumor in the patient. While p53 and p21 mRNAs were expressed at low levels in MCH-WT1, MCH-WT-3 showed high levels of p21 mRNA only. The increased expression of cyclin kinase inhibitor (p21) in MCH-WT-3 compared to MCH-WT-1 correlated with a higher percentage of G0/G1 cell population in the tumor specimen. Despite the expression of multidrug resistance markers (MDR1 and LRP) in MCH-WT-1, flow cytometric analysis showed tumor cell populations with very low and high daunorubicin accumulation and with the absence of any effect for verapamil and dipyridamole on daunorubicin accumulation of tumor cells. PMID- 11268452 TI - Comparative study of the acute nephrotoxicity from standard dose cisplatin +/- ifosfamide and high-dose chemotherapy with carboplatin and ifosfamide. AB - The nephrotoxic effects of different platinum compounds based combination chemotherapies were compared. Chemotherapy consisted of either cisplatin fractionated over 5 days (5 x 20 mg/m2) or given as a single-day infusion (1 x 50 mg/m2) plus ifosfamide (4 g/m2) or high-dose chemotherapy was applied including carboplatin (3 x 500 mg/m2) and ifosfamide (3 x 4 g/m2) fractionated over three consecutive days. Conventional parameters such as serum creatinine and glomerular filtration rate (GFR), as well as urinary protein excretion of N-acetyl-beta-D glucosaminidase (NAG)) and alpha 1-micro-globulin were assessed in 52 patients. Fractionation over 5 days without adding other nephrotoxic agents, i.e. ifosfamide, prevented decreases in GFR following cisplatin, whereas the combination of conventional dose cisplatin and ifosfamide, given as a single-day infusion, and high-dose carboplatin/ifosfamide yielded a pronounced fall of GFR. All groups showed increases in the urinary excretion levels of serum derived proteins and NAG, but with significant differences; about 2 to 3-fold for 5-days cisplatin, 3 to 5-fold for single-day cisplatin/ifosfamide, and 20 to 35-fold for high-dose chemotherapy. Thus, conventional approaches can reduce but not prevent the nephrotoxicity of cisplatin-based chemotherapy. In particular, high-dose chemotherapy regimens including carboplatin and ifosfamide are associated with comparable or even higher nephrotoxicity to single-day cisplatin/ifosfamide. In the light of the long-term consequences of persistent renal damage prevention of nephrotoxicity should be further improved. PMID- 11268453 TI - Expression of the neutral amino acids transporter ASCT1 in esophageal carcinomas. AB - Cancers cells utilize more glucose and amino acids than their benign counterparts. Overexpression of the facilitative glucose transporter Glut1 in human cancers was found to correlate with aggressive biologic behavior. The aim of this work was to determine whether the neutral amino acid transporter ASCT1 is expressed in human esophageal carcinomas, and to correlate the findings with Glut1 expression. Sections of formalin-fixed and paraffin-embedded tissue from 42 cases of esophageal carcinomas were entered in the study. Immunohistochemical staining was performed using a rabbit anti-ASCT1 IgG developed in our laboratory, and anti-Glut1 antibody, using standard avidin-streptavidine immunoperoxidase method. Sections of formalin-fixed and paraffin-embedded HepG2 cells were used as positive controls. The percent of ASCT1-positive cells was scored. Statistical analysis was performed using Fisher's exact test. ASCT1 immunoreactivity was cytoplasmic, whereas Glut1 was membranous. Significantly more adenocarcinomas expressed ASCT1 than squamous cell carcinomas (p < 0.0001), whereas Glut1 expression was similar in both tumor types. There was no association between the expression of either transporter and lymph node metastasis. Glut1 was expressed more often in the better differentiated than the poorly differentiated squamous carcinomas (p = 0.003). These results suggest that unlike in squamous cell carcinoma, ASCT1 plays a significant role in the recruitment of amino acids in adenocarcinoma of the esophagus, and suggest that the metabolic needs, and uptake of nutrients, are regulated differently in these two tumor types. Additional studies with larger number of patients are needed to determine the biological significance of ASCT1 expression in esophageal carcinomas. PMID- 11268454 TI - Paclitaxel and carboplatin for recurrent salivary gland malignancies. AB - BACKGROUND: The use of chemotherapy for recurrent salivary gland carcinomas is under investigation. PATIENTS AND METHODS: Fourteen patients (10 males, 4 females; median age 55 years, range 20-70) with recurrent carcinomas of major (9 patients) and minor (5 patients) salivary gland origin (histology: 1 adenocarcinoma, 10 adenoid cystic carcinoma, 2 undifferentiated carcinoma, 1 mucoepidermoid carcinoma) were treated with carboplatin AUC 5.5 + paclitaxel 175 mg/m2 (3-hour infusion) on day 1 (interval = 3 weeks). All patients had been previously treated with surgery + radiotherapy and 8 with a cisplatin combination. One patient had a local lesion, 7 locoregional recurrence and metastases and 6 patients had metastases only. RESULTS: Overall 65 courses were given (median 5; range 2-6). Responses were: PR in 2 patients (14%) lasting 5 and 12 months; 7 NC (50%) with a median duration of 8.5 months (5-12); and 5 PD (36%). The median survival time was 13.5 months for PR/NC patients, 6 months for non responders; median overall survival was 12.5 months (3-17+). CONCLUSION: This combination had a moderate activity; the treatment was well tolerated and toxicity was manageable. PMID- 11268455 TI - Cryosurgery as a means to improve surgical treatment of patients with multiple unresectable liver metastases. AB - BACKGROUND: The aim of the study was to evaluate the results of cryosurgery in patients with multiple (five or more), heavily pretreated, unresectable liver metastases. MATERIALS AND METHODS: Nineteen patients with multiple unresectable liver metastases were entered into a prospective nonrandomized trial. The liver tumours were treated during surgery under ultrasound guidance. All the patients were followed-up to assess complications, treatment response and sites of recurrence. RESULTS: 140 metastases were identified in 19 patients (mean, 7; range, 5-25) and 13 patients had a synchronous liver resection. Cryosurgery was used to treat 90 metastases (mean diameter, 30 mm; range, 10-135). There were no treatment-related deaths and the overall rate of complications was 21%. During a mean follow-up of 28 months (range, 5-60), tumours recurred at the site of cryosurgery in two patients (10%), in the remaining liver in nine patients (47%) and elsewhere in five patients (26%). Three patients had no evidence of disease 48, 50 and 60 months after liver cryosurgery, respectively. CONCLUSION: Cryosurgery may be effective in the treatment of patients with multiple unresectable liver metastases and should be investigated in multimodality treatment programmes. PMID- 11268456 TI - Independent prognostic importance of microvessel density in clinically localized prostate cancer. AB - BACKGROUND: Previous studies have reported a possible prognostic importance of microvessel density (MVD) in prostate cancer, although the significance after radical prostatectomy is not clear. The purpose of this study was to assess the prognostic value of MVD in clinically localized prostatic adenocarcinomas, focusing on moderately-differentiated tumours. MATERIALS AND METHODS: We examined a series of 104 patients treated for presumed organ-confined cancer in the period 1988-94. The area of highest tumour grade was selected from the prostatectomy specimens and vessels were high-lighted by staining for factor-VIII-related antigen. MVD was quantitated in the "hot spot" area and related to biochemical failure and clinical recurrence. RESULTS: In moderately differentiated tumours (WHO grade) (n = 66), MVD was associated with preoperative s-PSA and positive surgical margins. In univariate 5-year analysis, microvessel density (MVDmean > 122 mm-2, median) (p = 0.0074), s-PSA, tumour dimension, capsular penetration, seminal vesicle invasion and positive surgical margins were all significant predictors of biochemical failure, while MVDmean (p = 0.0084) was the only statistically significant predictor of clinical recurrence. In multivariate Cox' analysis, MVDmean (p = 0.0003), capsular penetration and tumour dimension remained as independent predictors of biochemical failure. CONCLUSION: Assessment of MVD in moderately differentiated prostatic adenocarcinomas may improve the prognostic stratification of patients after radical prostatectomy. PMID- 11268457 TI - Evaluation of 8-hydroxydeoxyguanosine in human oral cells: the importance of tobacco smoke and urban environment. AB - The DNA adduct 8-hydroxydeoxyguanosine (8-OHdG) has been widely used as a sensitive biomarker for oxidative damage. To investigate the role of environmental factors on oxidative DNA damage formation, the level of 8-OHdG was determined in oral cells from 109 healthy volunteers by an immunohistochemical method. A statistically significantly higher content of 8-OHdG was detected in oral cells from smokers (111 +/- 55, n = 38) compared with non smokers (78 +/- 48, n = 71), (p < 0.01). Moreover, subjects living in an urban area showed a higher level of oxidative damage with respect to those living in a countryside suburban area (99 +/- 53, n = 58 vs. 78 +/- 51, n = 51), (p = 0.03). No significant association was detected between 8-OHdG in oral cells and other variables such as passive smoke, oral infections, alcohol or vitamin intake and grilled food consumption. This work suggests that tobacco smoke and environmental exposure to pollutants lead to a measurable increase of oxidative damage in oral cells and confirms that the immunoperoxidase method is an appropriate approach for epidemiological analyses. PMID- 11268458 TI - Nerve contact with muscle component in neuromuscular hamartoma. AB - Neuromuscular hamartoma is a very rare soft tissue tumor, of which only 20 cases have been reported previously. None of these reports has described the relation between hamartomatous skeletal muscle and nerve fibers in the tumor. We experienced a patient with neuromuscular hamartoma arising at the brachial plexus. In this tumor, the localization of synaptophysin (SYP), S-100 protein (SP), neuron-specific enolase (NSE) neurofilament protein (NFP) and myoglobin (MG) was immunohistochemically detected. The results showed that SYP and MG were diffusely localized in the hamartomatous muscle fibers, SP in the schwann cells, and NSE and NFP in the axons of the hamartomatous nerve. Therefore, it is suggested that in the neuromuscular hamartoma, the structure of the neuromuscular junction may be similar to that in the motor end-plate of the normal muscle, but it may not be functional, because the hamartomatous muscles could not contract by nerve stimulation. PMID- 11268459 TI - Relationship between doxorubicin binding ability and tumor volume decrease after chemotherapy in adult malignant soft tissue tumors in the extremities. AB - We have previously reported that the doxorubicin (DOX) binding ability detected by the DOX (or adriamycin) binding assay closely correlated with the chemosensitivity of human osteosarcomas (1). We performed the present study to clarify the relationship between the DOX binding ability (%DB) and the histologic response, rate of decrease in tumor volume of malignant soft tissue tumors after preoperative chemotherapy and prognosis. Nine malignant soft tissue tumors (4 liposarcomas, 3 synovial sarcomas, one malignant fibrous histiocytoma (MFH) and one extraskeletal osteosarcoma (EOS)) which arose at the extremities of adult patients were analyzed by the DOX binding assay using freshly biopsied specimens. After preoperative chemotherapy including DOX or pirarubicin (THP), the rate of decrease in tumor volume was measured using magnetic resonance imaging, and the histologic response expressed as tumor necrosis to chemotherapy was also investigated. All the patients, apart for one, were continuously disease-free after treatment. One patient with EOS died of metastatic disease before surgery. The histologic response in 8 tumors without EOS was poor. The %DB of 5 tumors was greater than 80% (average: 95.90%), whereas that of 4 tumors was less than 80% (average: 38.33%). Although there was no correlation between the %DB and the histologic response, or prognosis, a significantly positive correlation was found between the %DB and the rate of decrease in tumor volume (r = 0.7455, p < 0.05). These results suggest that in malignant soft tissue tumors, the rate of decrease in tumor volume after chemotherapy might be a better indicator for chemosensitivity than the histologic response and also that the DOX binding ability might be a good predictor for chemosensitivity before chemotherapy. PMID- 11268460 TI - Case report of secondary chondrosarcoma showing spontaneous regression after frequent recurrences. AB - We report a case of secondary chondrosarcoma arising in the ilium showing spontaneous regression after frequent local tumor recurrences followed by multiple surgeries of marginal or intralesional excision. The patient was a 16 year-old boy who had been diagnosed as having multiple exostosis from 9 years of age. He experienced an increasing abdominal tumor mass that formed a huge tumor. Although marginal resection of the tumor was attempted, intraperitoneal dissemination was caused by rupture of the tumor capsule and the peritoneum, as a result of severe tumor adhesion to the peritoneum. During the 5 years after the initial operation, local recurrences occurred seven times in various areas of the intra- or retro-peritoneum and marginal or intralesional excision was performed every time for a total of 14 tumors. However, since the seventh operation, the patient has had no evidence of recurrence or metastasis of the disease for more than 10 years. Therefore, we considered that the cancer might have spontaneously regressed. PMID- 11268462 TI - Altered expression of hMSH2 in sporadic colorectal cancer, surrounding mucosa and at distant colonic mucosa. AB - BACKGROUND: Mismatch repair gene hMSH2 is involved in correction of mispairing during replication and its mutation is associated both with microsatellite instability and with hereditary colorectal cancer. We evaluated its involvement in sporadic colorectal cancer tumorigenesis too. MATERIALS AND METHODS: The protein expression pattern of hMSH2 was evaluated on 29 cases of resected sporadic adenocarcinoma using an immunohistochemical approach. RESULTS: In 14 cases, lack of hMSH2 protein expression was observed in adenocarcinoma and in peritumoral mucosa. In 12 patients, hMSH2 resulted in strong expression in the tumour as well as in the surrounding mucosa and at distant mucosa. In three cases, hMSH2 protein expression in tumoral, adjacent and at distance normal mucosa resulted negative. CONCLUSION: Repair genes could play an important role in tumour progression and in sporadic colorectal cancer. Detection of protein expression by immunohistochemistry may be a method to select tumours for successive genetic investigations. PMID- 11268461 TI - Immunohistochemical expression of transforming growth factor beta 1 and nm-23 H1 antioncogene in prostate cancer: divergent correlation with clinicopathological parameters. AB - BACKGROUND: Prostate cancer is one of the main causes of morbidity and mortality among men. Several oncogenes and growth factors have been studied in an attempt to explain the molecular basis of carcinogenesis and progress of this carcinoma. In this study we correlated the immunohistochemical expression of antioncogene nm 23 H1 and transforming growth factor beta 1 (TGF-beta 1) with the clinical stage, PSA values, Gleason score and survival in prostate cancer. MATERIALS AND METHODS: Fifty nine patients with prostate cancer were evaluated. PSA measurement, Gleason score determination and clinical staging were recorded for all the patients by the time of initial diagnosis and prior to any treatment. Follow-up ranged from 12 to 40 months. Tissue sections from representative areas of the tumors were immunohistochemically stained for nm-23 H1 and TGF-beta 1. The expression of these markers was correlated with stage, PSA values, Gleason score and survival. RESULTS: There was a negative correlation between nm-23 H1 staining and tumor stage and grade. High grade (Gleason score 8-10) and stage D tumors showed weaker staining than low stage and grade tumors. There was a positive correlation between TGF-beta 1 staining, tumor stage and serum PSA levels. Additionally, TGF beta 1 proved to be a negative predicting factor for patient survival. In tumors expressing both markers, TGF-beta 1 was the one to determine the aggressiveness of the carcinoma. CONCLUSIONS: nm-23 H1 appears to be a tumor suppressor gene in prostate cancer, while TGF-beta 1 may act as a stimulating agent provoking aggressive behavior and metastasis. Their immunohistochemical staining may constitute complementary information in the evaluation of prostate cancer patients. PMID- 11268463 TI - Telomerase activity and p53 gene mutation in familial polyposis coli. AB - BACKGROUND: Familial adenomatous polyposis (FAP) is a dominantly inherited disorder and it is difficult to know when polyps change to carcinoma. In this study, we retrospectively evaluated whether telomerase activity and p53 gene mutations are useful parameters for detecting malignant changes in polyps in patients with FAP. MATERIALS AND METHODS: Twenty-one tissue samples (carcinoma: 5, large polyp: 8, and small polyp: 8) were obtained from 8 patients with FAP. Colorectal carcinomas were detected in 5 patients. As a control, 68 colorectal carcinomas and 11 adenomatous polyps with synchronous colorectal carcinoma from 68 patients and 8 polyps from 8 patients without colorectal carcinoma were obtained. The telomeric repeat amplification protocol (TRAP) assay was used for the detection of telomerase activity. In 21 samples from FAP patients, mutations of exon 5 to 8 in the p53 gene were analyzed by the polymerase chain reaction single strand conformation polymorphism (PCR-SSCP) method. RESULTS: In 5 patients with FAP who had carcinomas in their colon and rectum, all the samples (carcinomas, large polyps and small polyps) showed telomerase activity. In contrast, in the 3 patients with FAP who had no carcinomas, telomerase activity was not detected in either the large or small polyps. Telomerase activity was detected in 51 out of 68 control colorectal carcinomas (75%) and in 6 out of 19 adenomas (32%). The p53 gene mutation was detected in only 2 carcinomas from 2 patients with FAP and was not detected in any large and small polyps. CONCLUSION: These findings suggest that telomerase is activated before the occurrence of the p53 gene mutation. In FAP, genetic changes associated with the development of colonic adenoma into carcinoma might activate telomerase and such genetic changes might occur in whole polyps of the colon and rectum. PMID- 11268464 TI - Prognostic value of S-phase fraction in T3N0M0 gastric cancer. Implications for adjuvant chemotherapy. AB - INTRODUCTION: The prognosis of patients with T3N0M0 gastric cancer is still unfavourable and the role of adjuvant chemotherapy is unclear. We addressed this study to evaluate whether the analysis of the S-Phase Fraction (SPF) might have prognostic implications in serosa-positive, node-negative gastric cancer. MATERIALS AND METHODS: Specimens of resected gastric cancer were studied by flow cytometry for SPF analysis. Consecutive patients with stage pT3N0M0, adequate follow-up information and sufficient archival tumor tissue were considered eligible for the study. The tumor SPF indices were related to the timing of recurrences, the relapse rate and the disease-free survival of patients. RESULTS: The analysis was carried out on samples of 137 patients with surgically-resected, stage pT3N0M0 gastric cancer. SPF resulted high and low in 39% and 61% of cases, respectively. Fifty-seven patients relapsed (42%) and early recurrences (within 18 months after surgery) occurred more frequently among cases with high SPF (p < .03). Patients with high SPF tumors showed a worse relapse rate and disease-free survival than patients with low SPF tumors. (p < .005). CONCLUSION: The SPF analysis showed prognostic differences among patients with stage pT3N0M0 gastric cancer. These data may be of value in the planning of future adjuvant chemotherapy trials in gastric cancer. PMID- 11268465 TI - Fine needle aspiration and core needle biopsy techniques in the diagnosis of nodular thyroid pathologies. AB - It is widly accepted that Fine Needle Aspiration Biopsy (FNAB) is the main test to distinguish benign from malignant thyroid lesions. Nevertheless, this technique presents some limits such as the possibility of false-negative or inadequate samples and it is unable to cytologically discriminate among adenomathosus goiter, follicular adenoma and well-differentiated follicular carcinoma. The aim of this study was to evaluate the possibility of restricting these limitations using Core Needle biopsy (CN) technique. Therefore we selected for CN, 40 out of 136 patients who underwent FNAB during a one year period; among these patients only 32 agreed a to this technique. Forty-two out of 136 patients underwent surgery; 29 of them were subjected to both biopsies. Sixteen of the diagnostic microbiopsies have been histologically confirmed. We had no discordant cases between cytological and microhistological diagnosis, except for one case which appeared cytologically colloid goiter, microhistologically follicular neoplasm and histologically follicular adenoma. In this case it was not possible to microhistologically discriminate benign from malign follicular lesion. In our experience not all patients accepted CN biopsy as well as FNAB and, moreover, this technique showed no advantage over FNAB diagnosis. On this base we think that actually FNAB should be the main procedure in the diagnosis of the thyroid lesions. It is easily performed, accepted by the patients and has a low cost benefit ratio. If the sample is not diagnostic it can be easily repeated and false-negative cases could be discovered thanks to an adequate clinical and ultrasonographical follow-up of the patients. PMID- 11268466 TI - Adjuvant tamoxifen versus tamoxifen plus CMF in the treatment of early breast cancer in Greece. Fifteen-year results of a randomised prospective trial and the potential risks of the antioestrogen. AB - BACKGROUND: CMF and Tamoxifen are the most commonly administered drugs for the adjuvant treatment of early-stage breast cancer. We present the 15-year follow-up of our 250-patient series and evaluate the oestrogenic side-effect of Tamoxifen on the endometrium. PATIENTS AND METHODS: 250 women entered this prospective study from 1981-1986. They had all undergone modified radical mastectomyl and were randomly assigned to receive either Tamoxifen only for 4 years or combination of Tamoxifen with 6 cycles of standard CMF. Abdominal sonogram was used to determine endometrial thickness, with 6 mm as cut-off limit for endometrial biopsy. RESULTS: After 15.6 years of follow-up DFS and OS rates were better for the CMF + Tamoxifen, group (52.8% vs 39.2%--p = 0.043 and 57.6% vs 40.8%--p = 0.006 respectively). Only patients with more than 4 infiltrated nodes did not significantly benefit from adjuvant CMF. Postmenopausal women suffered more proliferative endometria compared to premenopausal ones (40.3% vs 15.6%), while life-threatening lesions (cancer and atypias) were found in 3.3% of the postmenopausal patients only. CONCLUSION: CMF + Tamoxifen combination offers better long-term results for early-stage breast cancer patients. Dose reduction must be avoided if maximum results are to be achieved. More than 4 positive nodes seem to require additional chemotherapeutic manipulation. Tamoxifen's oestrogenic side-effect on the endometrium is quite common, but life-threatening lesions are rare, thus proving the drug's safety. PMID- 11268467 TI - Quantitative image analysis of oesophageal squamous cell carcinoma from the high incidence area of China, with special reference to tumour progression and papillomavirus (HPV) involvement. AB - BACKGROUND: Despite much research effort, the major prognostic factor of oesophageal squamous cell cancer (ESCC) remains the pathological stage of the disease as defined by the TNM classification, whereas tumour grading is of limited value in this respect, mainly due to its low reproducibility. A better means for disease prognostication based on improved understanding of the pathogenetic mechanisms is urgently required. MATERIALS AND METHODS: Among the cohort of 700 ESCC patients from the high-incidence area of China, previously subjected to extensive testing for Human papillomavirus (HPV) involvement by in situ hybridization (ISH) and PCR, a group of 273 patients was randomly selected for analysis of the primary tumour, adjacent mucosa and regional lymph nodes, by histopathology and quantitative image analysis. All these and the HPV data were subjected to extensive univariate and multivariate analysis to disclose independent predictors of progressive disease. RESULTS: For the analyses, the tumours were graded into two categories: well-moderately and poorly differentiated. HPV DNA was detected in 116 (18.9%) of the carcinomas by ISH and in 15.2% by PCR. In univariate analysis, lymph node status (considered as the surrogate marker of progressive disease) was significantly (p < 0.01) predicted by the following nuclear parameters: nuclear area, G0/G1 ratio, HPV DNA status, integrated optical density (IOD), mean optical density (MOD) and S-Phase. In multivariate (stepwise backward LR) analysis, 6 variables remained as independent predictors of disease progression (at p < 0.05 level), the three most significant ones being nuclear perimeter, nuclear roundness and equivalent diameter (p < 0.01). CONCLUSION: A series of quantitatively measured nuclear parameters seem to bear a close correlation with ESCC differentiation and progression in univariate analysis and some of these variables proved to be significant independent predictors of disease progression in multivariate modelling as well. These data clearly advocate the use of quantitative image analysis in searching for additional prognostic factors of ESCC. PMID- 11268468 TI - Prognostic value of circulating sialyl Tn antigen in colorectal cancer patients. AB - To examine the prognostic value of assessing the concentration of circulating sialyl Tn antigen (STN) after surgery, we determined serum STN levels in peripheral venous blood (designated "p-") in 308 colorectal cancer patients and what we have termed the "d-p gradient" (obtained by subtracting the serum concentration in peripheral venous blood from that in the tumor's drainage venous blood) in 144 patients. The prognostic value of STN and carcinoembryonic antigen (CEA) was compared. Patients were divided into low or high p-antigen groups and low, intermediate, or high d-p gradient groups. Univariate and multivariate analyses revealed that high STN d-p gradient, high p-CEA, or high CEA d-p gradient were each independent variables for poor patient outcome after surgery, separate from stage. In conclusion, a high STN d-p gradient was a predictor of poor outcome after resection for colorectal cancer, while p-STN was not independent of stage. PMID- 11268469 TI - Clinical pharmacokinetics and metabolism of paclitaxel after polychemotherapy with the cytoprotective agent amifostine. AB - BACKGROUND: Cytoprotection of healthy cells represents a new approach in cancer chemotherapy, but a pharmacokinetic drug interaction between the cytostatic and the cytoprotectant is undesired. METHODS: The purpose was to evaluate the clinical pharmacokinetics (PHK) of paclitaxel (PACLI) and its metabolites under cytoprotection in patients suffering from breast cancer. PACLI was administered alone and in a second cycle in combination with amifostine (AMI) as a paired cross over. RESULTS: In both treatment schedules the steady state of PACLI occurred after 3 hours, the tmax of metabolites between 3 and 4 hours. The mean steady-state concentration was cmax = 5432 +/- 1238 ng/ml in the control group and cmax = 5140 +/- 2407 ng/ml in the AMI group. For the serum metabolites, the findings were very similar: 6-OH-PACLI: cmax 413 +/- 153 ng/ml versus 432 +/- 304 ng/ml, 3"-OH-PACLI: cmax 99 +/- 103 ng/ml versus 123 +/- 98 ng/ml, 3",6-DiOH PACLI: cmax 43 +/- 55 ng/ml versus 75 +/- 85 ng/ml. AUC values of metabolites were slightly higher in the AMI group, but PHK seemed equal. CONCLUSION: The results gave evidence, that cytoprotection with AMI has no clinical consequences on PACLI pharmacokinetics and biotransformation. PMID- 11268471 TI - PAPNET-assisted primary screening of conventional cervical smears. AB - The PAPNET System is the only device with a neural-network-based-artificial intelligence to detect and show the images of abnormal cells on the monitor to be evaluated in an interactive way. We effectively used the PAPNET in rescreening of conventional cervical smears and we detected its advantages and its disadvantages. In this paper, we report our results from PAPNET-assisted primary screening performed on 20,154 conventional smears. The smears were classified as Negative or as Review. The Negative cases were rapidly rescreened mainly near the coverslip edges, which are the slide areas not analyzed by automated devices because of focusing problems. The Review cases were fully reanalyzed by the optic microscope. In summary, 140 positive smears were detected: 57 cases showed changes due to HPV, 63 LSIL, 15 HSIL, and 5 carcinomas. Therefore, the PAPNET System was confirmed as useful in primary screening of conventional cervical samples as well as rescreening. PMID- 11268470 TI - Influence of hormonal status of patients with cystic disease on the composition of cyst fluid and breast cancer risk. AB - The relationship between the composition of breast cyst fluid (BCF), the menstrual status and in addition some endocrine events in the history of patients (n = 131) with gross cystic breast disease was investigated. The dehydroepiandrosterone (DHA) levels in type II (K+/Na+ < 1) cysts of the follicular group were significantly higher compared to the type II cysts of the luteal or postmenopausal groups. For testosterone a significant difference existed between the type I (K+/Na+ > or = 1) follicular and type I postmenopausal groups. Estrone levels were significantly higher in type I BCF of patients in the luteal phase compared to both the follicular and postmenopausal type I cysts. Progesterone levels were lowest in the postmenopausal subgroups (both in type I and II cyst). Significant correlations were found between the number of pregnancies and the levels of DHA-sulfate and also progesterone in BCF. DHA levels were correlated with the period of lactation. The K+/Na+ ratios were the lowest in women who lactated for the longest period. The estrone was lowest in BCF of current oral contraceptive (o.c.) users while the estradiol was lowest in patients who had never used o.c. A history of previous o.c. use was associated with a significantly high mean DHA level. A significantly higher DHA and lower testosterone level were demonstrated in BCF of patients who had some previous gynecological interventions. The composition of BCF and the "life of cysts" and thus the rate of breast cancer risk may depend on hormonal status during the menstrual cycles or postmenopause and also on endocrine history of patients. PMID- 11268472 TI - Expression and clinical implications of bcl-2 in extrahepatic bile duct carcinoma: its relationship with biological features. AB - bcl-2 expression in 38 extrahepatic bile duct carcinomas was immunohistochemically investigated in order to clarify its clinical significance in this carcinoma. bcl-2 was expressed in 17 cases (44.7%), in which more than 10% of the carcinoma cells were positive for bcl-2. bcl-2 expression was more frequently absent in cases with advanced stages (p = 0.0642), lymph node metastasis (p = 0.0009), perineural invasion (p = 0.0647) and aberrant p53 expression (p = 0.0181). Furthermore, bcl-2 expression was inversely related to the incidence of apoptosis (p = 0.0002) and proliferating activity of the cells (p < 0.0001). These results suggest that bcl-2 plays a role in negatively regulating apoptosis and its loss can be significantly linked to the biological aggressiveness of this carcinoma. PMID- 11268473 TI - Immunohistochemical study of the receptors for retinoic acid in prostatic adenocarcinoma. AB - BACKGROUND: Retinoids play an important role in regulating cellular proliferation and differentiation. Although their effects are mediated by retinoic acid receptors (RARs) and retinoid X receptors (RXRs), limited information is available about the expression of RARs and RXRs in prostatic adenocarcinoma. We intended in this study to elucidate further the participation of those receptors in tumor progression of human prostate. MATERIALS AND METHODS: The immunohistochemical expression of RARs and RXRs proteins was studied using paraffin-embedded archival tissues obtained from patients with and without neoadjuvant hormonal therapies for prostatic adenocarcinoma. RESULTS: The immunoreactivity against RAR-alpha, RAR-gamma, RXR-alpha and RXR-gamma were detected in many, if not all, prostatic adenocarcinoma cells of the cases without and with neoadjuvant hormonal therapy, whereas less expression of both RAR-beta and RXR-beta were identified. Positively stained adenocarcinoma cells with RAR beta and RXR-beta were found to be decreased in the cases with neoadjuvant therapy. In addition, the specimens showing positive immunoreaction of RAR-beta were limited to the cases of moderately- and poorly-differentiated but not well differentiated, adenocarcinomas. CONCLUSION: We first demonstrated the expression of RARs and RXRs proteins in prostatic adenocarcinoma using subtype-specific antibodies. Immunohistochemistry using those antibodies might give an indication of susceptibility of the patients to retinoid therapy as a possible treatment of prostatic adenocarcinoma. PMID- 11268474 TI - Serum tetranectin and CA125 in endometrial adenocarcinoma. AB - BACKGROUND: CA125 and tetranectin (TN) are prognostic markers in ovarian cancer. This study examines the values of these markers in endometrial cancer. MATERIALS AND METHODS: TN and CA125 were determined preoperatively in 99 patients with primary endometrioid adenocarcinoma and evaluated in relation to tumor grade, stage and cancer survival. RESULTS: The CA125 levels correlated significantly with tumor stage. Dichotomized according to a cut-off level of 35 U/ml, CA125 significantly correlated with cancer death. Multivariate regression analysis of cancer survival time showed that CA125 > 35 U/ml was not an independent factor when stage was introduced. TN levels were within the normal range in all patients and did not show any association with tumor grade, stage or survival. CONCLUSIONS: The study confirmed the role of CA125 as a prognostic factor in endometrial cancer and may be of aid in pointing out patients at high risk, whereas tetranectin did not show any prognostic effect. PMID- 11268475 TI - Influence of tumor localization on the prognostic value of P53 protein in colorectal adenocarcinomas. AB - BACKGROUND: The prevalence of genetic alterations is different in primary carcinomas from the proximal colon when compared with carcinomas from the distal colorectum. The objective of this work was to explore the existence of possible differences in the informative weight of the risk of tumor recurrence provided by p53 immunostaining depending on the localization of the neoplasm. PATIENTS AND METHODS: Nuclear immunohistochemical expression of p53 protein was determined in formalin-fixed paraffin-embedded archival tumor tissue samples from 190 primary colorectal adenocarcinomas. The relative prognostic importance on the risk of recurrence of each variable was assessed in a Cox's proportional hazard regression analysis. Multiplicative interaction terms between p53 and tumor site were included in the multivariate models in order to test their joint effect on survival. RESULTS: One hundred and one patients (53.1%) manifested nuclear accumulation of the protein. P53 overexpression was more frequent in distal than in proximal tumors (58.5% ve s 41.7%) (p = 0.03). Disease-free survival was lower in p53-positive cases (75% versus 38%) (p = 0.006), but significance of the association varied according to the localization of the tumor (p = 0.004 in proximal carcinomas and p = 0.049 in distal carcinomas). Multivariate analysis identified p53 positivity and distal tumor localization as the factors significantly associated with a high risk of recurrence Interaction between p53 expression and localization was present. P53 exhibited different prognostic value in distal and proximal colon. While adjusted hazard ratio for positive p53 was 1.99 in distal cancers, it was 8.04 for proximal tumors. CONCLUSION: The prognostic with value of tumor recurrence associated overexpression of p53 protein is influenced by the location of the tumor. The negative predictive weight is significantly higher in proximal than in distal cancers. PMID- 11268476 TI - Distribution and prognostic value of the fibroblast growth factor-2 low-affinity binding sites in human breast cancer. AB - We performed a competitive binding study with 125I-labelled FGF (fibroblast growth factor)-2 and unlabelled FGF-2 in an unselected series of two hundred and thirty human primary breast cancers. One hundred and ninety-two breast cancer biopsies possessed FGF-2 low-affinity binding sites (FGF-2 LABS). The median dissociation constant was 2.4 nM (range, 1.03-18) and the median concentration of membrane protein was 6187.5 fmol/mg (range, 831-90,000). FGF-2 LABS concentrations were positively correlated to the progesterone receptor level. Cox univariate analyses showed that the FGF-2 LABS (> or = upper quartile) was associated to a longer overall survival (p = 0.05; RR = 0.042); node involvement, estrogen receptor progesterone receptor and histoprognostic grading were also prognostic. In Cox multivariate analyses, only the progesterone receptor, estrogen receptor, node involvement and FGF-2 LABS were prognostic factors; the FGF-2 LABS were associated with a longer overall survival (p = 0.033; RR = 0.068). The present study showed that FGF-2 LABS have only a limited role as a prognostic factor in breast cancer. PMID- 11268477 TI - Intrapulmonary teratoma associated with thymic tissue. AB - A case of benign, cystic intrapulmonary teratoma occurring in the right lobe of a 22-year old female is described with grossly and microscopically findings. The connection between the tumor and the segmental bronchus, together with the absence of germ cell neoplasms in other locations, clearly established the true intrapulmonary nature of the lesion. The unusual finding of thymic tissue within the wall supports the possible origin from the third pharyngeal pouch. PMID- 11268478 TI - Detection of circulating malignant cells by RT-PCR in long-term clinically disease-free I stage melanoma patients. AB - Recently, reverse transcriptase-polymerase chain reaction (RT-PCR) for the detection of circulating tumor cells has been suggested as a potential technique for staging cancer. In this report, 43 melanoma patients (including 4 in situ melanoma patients) were tested for tyrosinase mRNA in blood by RT-PCR. All patients had melanoma thinner than 1.5 mm (stage I). Circulating melanoma cells were detected in 8 (18.6%) out of 43 MM patients tested: 5 (16.1%) of 31 patients with melanoma thinner than 0.76 mm and 3 (42.8%) out of 7 patients with melanoma thicker than 0.76 mm. Moreover, in the tyrosinase-negative group we found only 4/31 patients (13%) with histologic signs of regression, but in the tyrosinase positive group, 3 out of 8 patients (37.5%) showed, at histologic examination, signs of regression. At the time of this analysis all the patients enrolled (tyrosinase-negative and tyrosinase-positive ones) were free of disease, probably due to the short median time of follow-up after the inclusion in the study. The presence of regression is an important cause of melanoma understaging and the tyrosinase test could represent an effective tool in order to achieve a realistic staging in this subgroup of melanoma patients. Probably, maximum sensitivity of the diagnostic RT-PCR approach to monitor MM patients with either localized or advanced disease could be achieved by using additional markers expressed with high frequencies in melanoma. We propose that one such marker could be the sign of regression. PMID- 11268479 TI - P53 protein expression in gastric adenocarcinoma. Negative predictor of survival after postoperative adjuvant chemotherapy. AB - BACKGROUND: The aim of the present study was to evaluate the influence of p53 protein on the survival of patients undergoing radical gastrectomy and postoperative adjuvant chemotherapy for gastric cancer. PATIENTS AND METHODS: It was a retrospective study of 46 patients with gastric adenocarcinoma (Stage II and III of the Japanese staging system). Alypatients were treated by curative radical gastrectomy with regional lymphadenectomy plus adjuvant chemotherapy. This regime included Mitomycin (20 mg one hour before surgery, followed by 10 mg the day after) and Fluorinated Pyrimidine (UFT) (400 mg/m2/day orally) (started four weeks after operation, and continued for one year). Immunohistochemical expression of p53 protein was determined on tumor samples from the removed specimens. The influence of p53 on survival was assessed in a Cox's proportional hazard regression analysis. RESULTS: Sixteen tumors (34.7%) manifested nuclear overexpression of p53 protein. Patients with p53-negative tumors showed higher cumulative survival at 4 years follow-up than patients with p53-positive tumors (82% versus 45%) (p < 0.01). Multivariate analysis identified p53 overexpression as a negative independent predictive factor (hazard ratio: 11.15) (95% CI: 1.93 64.42). Multivariate analysis performed on patients with Stage III tumors, separately, confirmed the predictive effect of p53 overexpression. CONCLUSION: The results suggest that postoperative adjuvant chemotherapy acted differently in p53-positive than in p53-negative gastric tumors. Absence of p53 overexpression is associated to longer survival when adjuvant therapy is administered. PMID- 11268480 TI - Evaluation of HPV, CMV, HSV and EBV in esophageal squamous cell carcinomas from a high-incidence area of China. AB - BACKGROUND: Certain viruses, notably human papillomavirus (HPV), cytomegalovirus (CMV), herpes simplex virus (HSV) and Epstein-Barr virus (EBV), are known to produce tumors in animals and cell transformation in vitro and they have been implicated in the pathogenesis of human cancers. All these viruses are also known to infect the esophagus. This study was aimed to determine whether these viruses play any causal role in the etiology of esophageal squamous cell carcinoma. MATERIALS AND METHODS: A series of 103 esophageal squamous cell carcinomas derived from patients in the high-incidence area of northern China were analyzed by DNA in situ hybridization and polymerase chain reaction (PCR) for the presence of HPV DNA sequences and, using immunohistochemistry, for the demonstration of CMV, HSV and EBV infections. RESULTS: Six (5.8%) of the 103 tumors were found to contain HPV 16, 18 or 30 DNA sequences. HPV types 6, 11 and 53 were not detected in any of the cases. Amplified HPV DNA sequences were found in 17 out of 101 (16.8%) carcinoma specimens by PCR with L1 consensus primers. None of the 103 carcinomas tested was immunohistochemically positive for CMV, HSV or EBV. CONCLUSION: Our results confirmed the HPV involvement in esophageal carcinomas and provided further evidence to support a causal association of HPV infection with esophageal squamous cell carcinoma. However, the three herpesviruses, CMV, HSV and EBV, are highly unlikely to be involved in the pathogenesis of this malignancy in the high-incidence area of China. PMID- 11268481 TI - Expression of thymidine phosphorylase and vascular endothelial growth factor in epithelial ovarian cancer: correlation with angiogenesis and progression of the tumor. AB - OBJECTIVE: The aim was first, to determine whether the expression of thymidine phosphorylase (TP) and vascular endothelial growth factor (VEGF) by epithelial ovarian cancer cells is correlated with the density of microvessels within the tumor and with ultrasonographic findings (B-mode classification and pulsed Doppler blood flow) and second, to speculate how these two angiogenesis factors participate in the tumorigenesis and tumor progression of epithelial ovarian cancer. METHODS: B-mode ultrasonography and color Doppler imaging and pulsed Doppler spectral analysis were used to scan patients with an overt ovarian mass immediately before laparotomy. Sections of malignant tumors were analyzed for the cellular expression of TP and VEGF and the intratumoral density of microvessels by immunohistochemistry using antibodies to TP, VEGF and Factor VIII related antigen, respectively. The main outcome measures were the histological classification of the tumor, the stage of the disease, whether or not the tumor cells were TP and VEGF positive or negative, the microvessel count and B-mode classification and the peak systolic velocity (PSV). RESULTS: Forty-four epithelial ovarian cancers were studied (6 of low malignant potential, 15 serous cystadenocarcinoma, 9 mucinous cystadenocarcinoma, 8 endometrioid adenocarcinoma, 4 clear cell carcinoma and 2 malignant Brenner tumor); 19 were Stage I, 6 Stage II, 15 Stage III and 4 Stage IV. Fourteen tumors (32%) were classified as TP positive and 21 (48%) as VEGF positive. The proportion of Stage I tumors that were TP positive (16%) was significantly lower (p = 0.022) than the corresponding value for Stages II-IV (44%), but the proportion with VEGF positive, the values for microvessel count and PSV were similar. Microvessel count did not significantly related to TP nor VEGF expression. The PSV was significantly higher in TP-positive tumors (p = 0.02) and VEGF-positive tumors (p = 0.006), respectively. There was a significant correlation between the microvessel count and the PSV (r = 0.34, p = 0.024). Moreover, specific B-mode classification significantly associated with disease stage and with TP expression, but not with VEGF expression. CONCLUSIONS: Angiogenesis is an early, critical step in the tumorigenesis of epithelial ovarian cancer, however, it does not provide significant information about tumor aggressiveness. When TP expression is superimposed upon the VEGF expression, the tumor might acquire the aggressive tumor phenotype. PMID- 11268482 TI - Telomerase activity in long-standing ulcerative colitis. AB - Telomerase activity is frequently associated with neoplasia. It is a ribonucleoprotein capable of replacing telomeric DNA sequences that are lost at each cell division. Neoplastic progression in chronic ulcerative colitis is characterized by the development of epithelial dysplasia which is accompanied by genetic alterations. Therefore we tested telomerase activity in 128 biopsy samples of four colectomy specimens with long-standing ulcerative pancolitis by using the Telomerase PCR ELISA System. In three patients with multiple dysplastic or carcinomatous lesions, telomerase activity was detected in 22 samples with a regional association to dysplastic or carcinomatous areas. 15 of the samples with telomerase activity (68%) were found in dysplastic/carcinomatous samples or in the direct vicinity of dysplastic areas, 4 (18%), 2 positions (about 4 cm) and the remaining three (14%) not more than 3 positions away from such areas. In the fourth patient, resected because of clinical deterioration despite medical treatment and who had no dysplastic lesions, no telomerase activity was detected. These results show that telomerase activity might be used as a complementary marker to histology for the identification of patients with ulcerative colitis who are at an increased risk for neoplastic progression. PMID- 11268483 TI - Sequential high dose-density chemotherapy in advanced ovarian cancer. AB - Introduction of paclitaxel or anthracyclines can improve the results of chemotherapy in advanced ovarian cancer. Dose intensification (by shortening of intervals between cycles) and sequential administration of active regimens at least theoretically may improve chemotherapy effectiveness. 18 patients entered into a pilot trial of combination chemotherapy. Treatment consisted of cisplatin 50 mg/m2, epidoxorubicin 60 mg/m2 and cyclophosphamide 500 mg/m2 every 14 days for six cycles, followed by paclitaxel 175 mg/m2 (3-hour infusion) every 14 days for four cycles. Granulocyte colony stimulating factor at 300 mcg was employed between cycles on days 5-10. 16 out of 18 patients who entered the study received a full dose chemotherapy with a ratio between actually received and planned dose intensity of 0.8 or more. No life-threatening side effect was observed and toxicity was acceptable. This new approach based on sequential administration of active regimens at high dose intensity proved feasible, active and devoid of unacceptable toxicity. The administration the booth of paclitaxel and epidoxorubicin with cisplatin and cyclophopshamide has been rendered possible. Further studies are warranted. PMID- 11268484 TI - Extraabdominal desmoid tumor with dissemination detected by thallium-201 scintigraphy. AB - Extraabdominal desmoid tumor is a locally aggressive tumor despite being histologically benign. To avoid local recurrence, it is important to preoperatively detect the exact localization and extension of the infiltrating or disseminating lesion in this tumor. We report a case of recurrent extraabdominal desmoid tumor, which arose in the posterior thigh region, detected with Tl-201 (Tl) scintigraphy. In this case, Tl accumulated in the small disseminating lesion and to the recurrent tumor. This lesion was not identified by palpation because of its small size, deep localization and absence of symptoms, although MR imaging, which was performed after the Tl scintigraphy, clearly showed the lesion. After tumor resection, Tl did not accumulate in any region. These results suggest that Tl scintigraphy may be useful, not only for the diagnosis of extraabdominal desmoid tumor, but also for the detection of the exact localization or extension of small infiltrating or disseminating lesions before treatment. PMID- 11268485 TI - Assessment of chemosensitivity in patients with malignant bone and soft tissue tumors using thallium-201 scintigraphy and doxorubicin binding assay. AB - This study was performed to compare the accumulation of thallium (Tl)-201 which is correlated with malignancy and the doxorubicin binding ability, which is correlated with chemosensitivity, in nine patients who received preoperative chemotherapy with doxorubicin and cisplatin. Tl-201 scintigraphy was performed at 15 minutes (early image) and 3 hours (delayed image) after injection of 111 MBq of Tl-201. The change of degree of the radionuclide uptake between the early and delayed images was evaluated before and after preoperative chemotherapy. The doxorubicin binding ability (%DB) to nuclear DNA in living tumor cells isolated from biopsy materials was assessed by doxorubicin binding assay. The histologic response to preoperative chemotherapy was evaluated by the percentage of tumor necrosis. Before preoperative chemotherapy no changes of Tl-201 uptake between the early and delayed images was detected in any tumors. Five patients, who had no change of Tl-201 uptake after preoperative chemotherapy, showed a poor histologic response and had a %DB ranging from 10% to 70% (mean: 36.0%). The other four patients, who had a %DB greater than 90%, showed a good histologic response. All of these four patients had decreased Tl-201 uptake after preoperative chemotherapy. This study demonstrated that doxorubicin binding assay and midcourse Tl-201 scintigraphy are useful methods to assess the response to chemotherapy early in malignant bone and soft tissue tumors. PMID- 11268486 TI - The loss of NM23 protein in malignant melanoma predicts lymphatic spread without affecting survival. AB - NM23 is considered to be a metastasis suppressor gene the role of which as prognosticator in the case of malignant skin melanoma (MM) is highly controversial due to different results on gene, and protein expressions. Accordingly, we analyzed the protein expression of NM23 in 32 primary skin melanomas with a follow-up period of 5 years minimum. We found that NM23 expression was the lowest in the thickest primary tumors (based on the % of the positive cells and the incidence of low expressor tumors) but the difference was not significant statistically due to the extreme heterogeneity of the tumors. When primary tumors were grouped according to their biological behavior (non metastatic; lymph-node (LN) metastatic; organ and LN metastatic tumors) we observed that the lowest NM23 protein expression (based on the % of positive tumor cells as well as on the incidence of low expressor tumors) was found in the LN metastatic tumors compared to other groups (p < 0.05). The NM23 phenotype of the primary tumors remained stable in the corresponding LN metastases in the case of the 5 analyzed tumors. There was no difference in the 5-year survival between patients with low (< 50% positive cells) or high NM23 protein expressing primary tumors. Collectively, these data suggest that the NM23 protein expression in the primary tumors of MM predicted lymphatic spread but did not affect 5-year survival because it did not correlate with organ metastasis. PMID- 11268487 TI - Urokinase-type plasminogen activator, plasminogen activator inhibitor type 1 and cathepsin D: analysis of their prognostic significance in squamous cell carcinoma of the head and neck. AB - BACKGROUND: The aim of the study was to evaluate the prognostic significance of tumour and serum concentrations of urokinase-type plasminogen activator (uPA), its type 1 inhibitor (PAI-1) and cathepsin D (Cath D) in patients with squamous cell carcinoma of the head and neck (SCCHN). PATIENTS AND METHODS: Determinations of uPA and PAI-1 were made using enzyme-linked immunosorbent assays in tumour and serum samples of 47 and 32/47 patients, respectively. For the determination of tumour (94 patients) and serum (34/94 patients) Cath D concentrations, an immunoradiometric assay was used. RESULTS: In an univariate survival analysis, the risk of disease recurrence and SCCHN-related death was significantly higher in the patients with high uPA (P = 0.046, P = 0.010) tumours, compared to those with low uPA tumours. In addition, the high serum levels of uPA correlated positively with the rate of relapse (P = 0.007), but not with the mortality rate (P = 0.200). There was no statistically significant difference between low and high PAI-1 groups, regarding either tumour or serum concentration of the inhibitor, and between low and high Cath D tumours. Low Cath D serum levels appeared to be related to longer disease-free interval (P = 0.055), but not to disease-specific survival (P = 0.120). CONCLUSIONS: The tumour levels of uPA, as well as serum levels of uPA and Cath D could potentially predict the survival probability of patients with SCCHN. However, the strength of this association remains to be investigated on a larger and more homogeneous group of patients. PMID- 11268488 TI - Preoperative thrombocytosis is an independent prognostic factor in stage III and IV endometrial cancer. AB - BACKGROUND: To identify prognostic factors in stage III and IV endometrial cancer with special attention to pretreatment platelet count. MATERIALS AND METHODS: 59 patients with FIGO stage III or IV disease operated on between 1983 and 1993 were analyzed. Patients with preoperative thrombocytosis were compared with those without thrombocytosis. Prognostic factors were analyzed with a Cox proportional hazard model. RESULTS: With the exception of age, there were no significant differences between patients with or without thrombocytosis. At multivariate analysis, five-year disease-free survival was influenced significantly by FIGO stage (stage III vs stage IV; p = 0.009), thrombocytosis (p = 0.02) and cervical involvement (p = 0.024). Similarly, overall five-year survival was significantly influenced by stage (p < 0.001), cervical involvement (p = 0.005) and thrombocytosis (p = 0.01). Age, histology, grade, myometrial invasion, lymph vascular space involvement or spread to adnexae were not significantly associated with survival. CONCLUSION: Thrombocytosis is an independent prognostic factor in stage III and IV endometrial cancer. PMID- 11268489 TI - Pilot study of immunotherapy with interleukin-2 after autologous stem cell transplantation in advanced breast cancers. AB - Median survival for advanced breast cancer does not exceed 2 years. Immunotherapy following Hihg Dose Chemotherapy (HDC) and autologous stem cell transplantation (ASCT) is a procedure that could hypothetically decrease relapse rate. The mechanism implicated is induction of immune modulation and a possible Graft Versus Tumor effect (GVHT). Tolerance and feasibility of rIL-2 administered after HDC with ASCT was analyzed in twenty one advanced breast cancer patients. The patients were treated either with intra-venous high-dose rIL-2 (9 patients) or subcutaneous low dose (12 patients). With intra-venous high-dose rIL-2, 50% of the scheduled dose was administered and 100% of the scheduled dose was administered at a lower dose in the subcutaneous route. rIL-2 was administered safely after HDC and ASCT, particularly in the subcutaneous low dose arm. However no clinical beneficial effect was documented for these advanced heavily pretreated breast cancers. Immune modulation with rIL-2 earlier requires further investigation. PMID- 11268490 TI - Malignant melanoma of soft tissues (clear cell sarcoma) of the foot. Is MRI able to perform a specific diagnosis? Report of one case and review of the radiological literature. AB - The magnetic resonance (MR) findings in malignant melanoma of soft tissues, also called clear cell sarcoma of tendons and aponeuroses, have been described as a focal abnormality with a specific MR pattern of increased signal intensity (relative to normal muscle) on T1 weighted sequences and variably decreased signal intensity on T2 weighted sequences (relative to surrounding fat). We have reported here a case of malignant melanoma of soft tissues of the foot, studied with ultrasonography (US) and MR, in which MR showed T1-hypointensity, T2 hyperintensity and marked gadolinium uptake. We have described the relationship between the intracytoplasmic melanin amount of and these atypical MR findings. PMID- 11268492 TI - Cytological diagnosis of malignant mixed mullerian tumor of the uterus in ascitic fluid. AB - The malignant mixed Mullerian tumour (MMMT) is a rare and aggressive neoplasm of the uterus, seen in postmenopausal women. In this case, an uncommon neoplasm was diagnosed cytologically in the ascitic fluid of a woman 58 years old and was confirmed histologically after hysterectomy and bilateral adnexectomy. PMID- 11268491 TI - New triplet chemotherapy combination with carboplatin, paclitaxel and gemcitabine plus amifostine support in advanced non small cell lung cancer: a phase II study. AB - New triplet chemotherapy combinations are under investigation in advanced non small cell lung cancer (NSCLC). Carboplatin, plus paclitaxel, plus gemcitabine is among the most active and promising regimens. The use of more aggressive chemotherapy in order to improve results can increase toxicity. Amifostine (WR 2721) reduces toxicity of radiotherapy and chemotherapy and protects selectively a number of normal, but not neoplastic, tissue. Based on this background, we performed a phase II study on carboplatin, plus paclitaxel, plus gemcitabine with amifostine support in advanced NSCLC. Patients received chemotherapy at the following dosage: carboplatin AUC 5, i.v., at day 1; paclitaxel 175 mg/m2, i.v. by 3-hour infusion, at day 1; gemcitabine 1000 mg/m2, i.v. by 3-hour infusion, at days 1 and 8; every 3 weeks for a maximum of 6 cycles. Amifostine was administered at the dose of 740 mg/m2, i.v., at day 1 of each cycle. Seventeen patients entered the study. They were prevalently male, median age was 62 years, PS (ECOG) was 0 in 10 cases (58.8%), 1 in 6 (35.3%) and 2 in 1 (5.9%). Histology was epidermoid in 8 cases (47%) and adenocarcinoma in 9 (53%). We observed 8 (47.5%) objective responses with 2 (11.7%) complete responses. Median time to progression and median survival were 24 and 36 weeks, respectively. Treatment was well tolerated. The main toxicity was as follows: grade 3 neutropenia, grade 2 thrombocytopenia and grade 3 anemia in one (5.8%) case; grade 2 peripheral neurologic toxicity in 3 (17.6%) patients; grade 2 cardiac toxicity (atrial fibrillation) in one case; and grade 3 respiratory toxicity (dispnoea) in one patient. These data indicate that this combination has promising activity and tolerability. A randomized trial comparing carboplatin plus paclitaxel, plus gemcitabine versus carboplatin, plus paclitaxel, plus gemcitabine, plus amifostine in advanced NSCLC is warranted. PMID- 11268493 TI - Prognostic significance of high microsatellite instability in a Spanish series of gastric adenocarcinomas. AB - In colorectal cancer different levels of microsatellite instability (MSI) have been described. MSI-H (high) characterizes a unique clinical and pathological phenotype known as hereditary non-polyposis colorectal cancer syndrome, whereas MSI-L (low) and MSS (stable) are considered similar phenotypes without pathological implications. MSI has been also described as a frequent genetic alteration in a subset of gastric adenocarcinomas. However, the clinicopathological and prognosis significance of this abnormality in these tumors remains unclear. To investigate the role of genetic instability in gastric carcinogenesis we examined 10 microsatellite loci in 37 patients. MSI-H was found in 37.8% patients. We observed a trend of MSI-H tumors to be associated with elderly patients, intestinal histological type, advanced clinical stages and less aggressiveness with better survival. In conclusion, MSI-H can be considered as a good prognosis factor in a subset of gastric tumors. PMID- 11268494 TI - Sequential high dose-density chemotherapy with two active regimens for advanced small cell lung cancer. AB - 16 patients with advanced small cell lung cancer were treated with a combination of cyclophosphamide (1000 mg/m2 day 1), epidoxorubicin (60 mg/m2 day 1) and vincristine (1.4 mg/m2 day 1) every 14 days for six cycles followed by a combination of cisplatin (40 mg/m2 days 1 & 2) and etoposide (100 mg/m2 days 1-3) every 14 days for four cycles. Shortening of intervals was obtained with the prophylactic employment of granulocyte colony-stimulating factor (filgrastim, 300 mcg subcutaneously from day 5 to dsy 10). In 11 patients ratio between actually delivered dose intensity and planned dose intensity of > 80% was obtained. Toxicity was acceptable and no life-threatening toxicities were observed. An objective response (partial or complete) was observed in 11 patients. The new regimen, incorporating the concepts of dose-intensification and sequential administration of regimens, is feasible and may be considered for further studies. PMID- 11268495 TI - MRP expression of testicular cancers and its clinical relevance. AB - BACKGROUND: The expression of a multidrug resistance associated protein (MRP) has been investigated in a variety of human tumors. However, there is a lack of data regarding its expression in germ cell testicular tumors (GCTTs). PATIENTS AND METHODS: MRP expression was examined by immunohistochemistry (IHC) using mouse monoclonal antibody (MRPm6) against human MRP in 56 testis cancer specimens. This antigen was also correlated with the histology, metastatic behavior, clinical stage and tumor suppressor protein p53 immunostaining of GCTTs. RESULTS: All testis tumors, regardless of their histology, metastatic status and clinical stage gave positive signals. MRP was positive not only in the cytoplasm but, very interestingly, in the nuclei. CONCLUSION: Our results suggested that ala GCTTs express high levels of MRP protein with no relation to any of clinicopathological variables investigated here. Since germ cell tumors are very sensitive to chemotherapy, the role of MRP as mediator of drug resistance seems unconvincing in this malignancy. MRP is located in the cytoplasm and the nuclei of tumor cells and may be involved in transportation and/or redistribution certain substrates from the nucleus to the cytoplasm. PMID- 11268496 TI - Carboplatin, cisplatin and paclitaxel in the treatment of patients with epithelial ovarian cancer. AB - BACKGROUND: Prognosis of advanced ovarian cancer is unsatisfactory. Chemotherapy can be intensified combining active drugs at their highest possible doses. PATIENTS AND METHODS: In this phase I/II trial, 77 untreated patients received escalating doses of paclitaxel (135, 155, 175, 195 and 215 mg/m2, infused over 3 hours) with carboplatin (AUC 3.6) and cisplatin (60 mg/m2). Nine, 16, 13, 8 and 3 patients were treated at the five levels, respectively. A further 28 patients were treated at the maximum tolerable dose (MTD). RESULTS: Dose-limiting toxicities (one WHO grade 3 constipation, one grade 2 prolonged peripheral neurotoxicity and one grade 3 cardiac toxicity) occurred at 215 mg/m2 in 3 out of 3 patients. MTD was reached at level 4 paclitaxel dose (195 mg/m2). Response was evaluated in 62 patients. A complete response was achieved in 23 patients (37.1% 95% CI 25.2-50.3), including 16 (25.8%) pathological and partial response in 28 (45.2%), for an overall response rate of 82.3% (95% exact CL: 70.5%-90.8%). The probability of response was affected by the degree of initial debulking (p = 0.002) and not by the paclitaxel dose. In patients with stage III-IV disease, median progression-free survival was 17 months (95% CI 14-25). After a median follow-up of 28 months, median survival had not been reached; 2-year estimated survival was 67%. CONCLUSION: Paclitaxel can be safely given at the dose of 195 mg/m2 in combination with carboplatin (AUC 3.6) and cisplatin (60 mg/m2). This combination is active and safe and could be considered in clinical settings requiring intensive short treatment. PMID- 11268497 TI - Prognostic significance of P53 histochemistry and DNA histogram parameters in head and neck malignancies. AB - BACKGROUND: The aim of the study was to determine the role of quantitative pathological parameters in prognosis of head and neck malignancies. MATERIALS AND METHODS: 51 head and neck squamous cell carcinoma patients were examined for mutant p53 gene expression (45 out of 51 patients) by immunohistochemistry and for cellular DNA-content (44 out of 51 patients) using digital picture analyzer. Statistical analysis was performed using BMDP package. RESULTS: No correlation with prognosis was found for age, sex, localization, T-classification and therapy. There was significant relationship between N-status and overall survival (p = 0.0008). No correlation was found with overall and disease-free survival for either histologic type or grading. P53: No significant correlation was detected with overall survival. A relationship was found between mutant p53 and metastasis free time (p = 0.06). Ploidy: There were no significant differences between aneuploid and euploid tumors for either disease-free or overall survival. Synthetic (S)-phase fraction: A correlation was found for both survival rates (p = 0.029) and metastasis-free time (p = 0.05). Polyploid fraction (PF): correlation was shown for both overall survival (p = 0.0128) and metastasis-free time (p = 0.0038). CONCLUSION: There is correlation between p53 overexpression and metastatic potential and there is a significant relationship between SPF and PF value and prognosis (metastasis-free and overall survival) of head and neck cancer. PMID- 11268499 TI - Paclitaxel/carboplatin as first-line chemotherapy in advanced ovarian cancer: efficacy and adverse effects with special consideration of peripheral neurotoxicity. AB - BACKGROUND: The aim of this study was to investigate the response and survival probability of patients with advanced ovarian cancer treated with a combination of paclitaxel and carboplatin as first-line chemotherapy. Additionally we investigated the extent of adverse effects due to chemotherapy with special consideration for peripheral neurotoxicity. MATERIALS AND METHODS: Thirty-seven women with epithelial ovarian cancer, treated with a combination chemotherapy consisting of paclitaxel and carboplatin, were included in the analysis. A total of 234 courses of paclitaxel/carboplatin were applied. Paclitaxel was administered at a dose of 175 mg/m2, infused over 3 hours, every 21 days. Carboplatin was administered at an area under the concentration-time curve (AUC) of 6. RESULTS: Thirty of the 37 patients responded to the chemotherapy, demonstrating an overall response of 81%. Seven patients died of the disease (19%). The mean overall survival was 20 months (25% quartile: 19, median and 75% quartile not reached). Thirteen patients (34%) developed peripheral neurotoxicity. In 10 cases (76%) neurotoxicity occurred after the fifth and sixth chemotherapy cycle. In all cases of severe neurotoxicity pathologic sensory nerve conduction-measurements were observed. In one patient a weakness of the left leg was observed. Apart from alopecia, other adverse effects were rare. CONCLUSION: This study confirmed the therapeutic benefit of the combination of paclitaxel and carboplatin as first-line chemotherapy in patients with ovarian cancer. Neurologic toxicity, increasing with every cycle of the chemotherapy, was a clinically significant adverse effect in our study. However, peripheral neuropathy mainly affected sensory fibers, without involving motor nerves. PMID- 11268498 TI - Continuous subcutaneous octreotide in gastrointestinal cancer patients: pain control and beta-endorphin levels. AB - BACKGROUND: Somatostatin is a naturally occurring hormone widely identified in a number of human tissues, with a broad spectrum of physiological actions. Octreotide is a synthetic analogue of somatostatin, which seems to be promising in clinical use. AIMS: a. to evaluate the efficacy of octreotide in pain control of patients with advanced gastrointestinal cancer, as well as octreotide's outcome in the hepatic function; b. to investigate the relationship between pain intensity and beta-endorphin blood levels in the patients. PATIENTS: The study group consisted of 25 patients (age range: 48-89 years, 14 males, 11 females) with far advanced gastrointestinal cancer. METHODS: All the patients were under s.c. morphine administration using a continuous infusion pump. When pain intensity increased, 0.6 mg/day of octreotide was added to the therapeutic regimen in the same syringe of the continuous infusion pump. Pain intensity and beta-endorphin blood levels were measured five times: Once before octreotide administration and the other four 12, 24, 48 hours and 7 days after. A complete blood count and a biochemical screening profile were taken before the administration of octreotide as well as on the 7th and the 14th day. RESULTS: 24 out of 25 cases showed a reduction in pain intensity (pretreatment x = 5.3, post treatment x = 0.6). beta-endorphin blood levels increased significantly during the study (an increase of 184.78% was observed on the 7th treatment day). In one patient pain control was achieved by increasing morphine dosage. Statistically significant changes were observed in hepatic function indices (p < 0.02). Significant side-effects were not observed. CONCLUSION: Octreotide can be used as an adjuvant analgesic in the management of gastrointestinal cancer pain which is managed by continuous s.c. administration. Although fuither research needs to be done, octreotide's administration seemed to improve hepatic function of these patients, therefore, it could potentially have a positive effect in the patient's quality of life. PMID- 11268500 TI - A paclitaxel-containing chemotherapy does not cause central nervous adverse effects: a prospective study in patients with ovarian cancer. AB - BACKGROUND: The objective of this study was to evaluate the possible effects of a paclitaxel containing chemotherapy on different neuropsychological parameters in women with ovarian cancer. MATERIALS AND METHODS: Twenty-eight women with histologically documented epithelial ovarian carcinoma and treated with a combination chemotherapy consisting of paclitaxel and carboplatin entered the study. The patients were tested with a battery of different neuropsychological tests before, after 3 cycles and at the end of the chemotherapy. RESULTS: Twenty of the 28 patients responded to the chemotherapy (71%). Eleven patients (39%) developed peripheral neurotoxicity. The median values of 6 tests performed before the first chemotherapy cycle scored out of the normal range. These patients with deviant test results at the beginning of the paclitaxel/carboplatin infusions did not deteriorate during chemotherapy. We found a statistically significant improvement of the alphabetical cross out test from the first to the third measurement (mean increase = 4.07; 95% confidence interval = [0.99; 7.15]) (p < 0.05), indicating an improvement of the short-term attention, the concentration and the constancy of working during chemotherapy. The other tests failed to show statistically significant changes during chemotherapy (p > 0.05). CONCLUSION: According to our results, a chemotherapy consisting of paclitaxel/carboplatin caused no signs of acute central nervous toxicity or neuropsychological deterioration. PMID- 11268501 TI - Human resources. Thoroughly modern matron. PMID- 11268502 TI - Human resources. Lean on me. AB - Appointing a full-time staff support co-ordinator in a community trust has reduced absence and been welcomed by staff. The pilot, which has been operating for two years, offers staff confidential interviews. Most of those using the service do not need more than two sessions. The co-ordinator has also acted as mediator where there is conflict between staff. PMID- 11268503 TI - Quick quick, slow. PMID- 11268504 TI - On the evidence. Patient information. PMID- 11268505 TI - Consultants. The seven-year hitch. PMID- 11268506 TI - General practice. Catch a falling star. AB - A panel set up to identify and support poorly performing GPs in one health authority relies on patients and health professionals to raise their concerns. Doctors from small and single-handed practices account for many of those investigated. The average length in practice of those investigated was 28 years. Panel members believe the system is working reasonably well but does not provide quick solutions to problems of poor performance. PMID- 11268507 TI - US healthcare. Taking to the Bush. PMID- 11268508 TI - Equal opportunities. Fair do's. PMID- 11268509 TI - Schistosomiasis and soil-transmitted helminth infections. PMID- 11268510 TI - Remarks on the discovery of carbonic anhydrase. PMID- 11268511 TI - Carbonic anhydrase (CA)-related proteins (CA-RPs), and transmembrane proteins with CA or CA-RP domains. PMID- 11268512 TI - Regulation of the CA1, CA2 and CA3 genes. PMID- 11268513 TI - Introduction to the carbonic anhydrases. PMID- 11268514 TI - Use of carbonic anhydrase II-deficient mice in uncovering the cellular location of membrane-associated isoforms. PMID- 11268515 TI - X-ray crystallographic studies of mammalian carbonic anhydrase isozymes. PMID- 11268516 TI - The catalytic mechanism of mammalian carbonic anhydrases. PMID- 11268517 TI - Activation of carbonic anhydrase isozymes. PMID- 11268518 TI - Active-site engineering of carbonic anhydrase and its application to biosensors. PMID- 11268519 TI - Folding and stability of human carbonic anhydrase II. PMID- 11268520 TI - Membrane transport and provision of substrates for carbonic anhydrase: in vertebrates. PMID- 11268521 TI - Respiratory and renal roles of carbonic anhydrase in gas exchange and acid-base regulation. PMID- 11268522 TI - Evolution and distribution of the carbonic anhydrase gene families. PMID- 11268523 TI - The roles of carbonic anhydrase in metabolism, cell growth and cancer in animals. PMID- 11268524 TI - The roles of carbonic anhydrase in gustation, olfaction and chemical irritation. PMID- 11268525 TI - Carbonic anhydrases in striated muscle. PMID- 11268526 TI - Inherited deficiencies and activity variants of the mammalian carbonic anhydrases. PMID- 11268527 TI - Carbonic anhydrase inhibition in ophthalmology: carbonic anhydrases in cornea, lens, retina and lacrimal gland. PMID- 11268528 TI - Carbonic anhydrase inhibition in ophthalmology: aqueous humor secretion and the development of sulfonamide inhibitors. PMID- 11268529 TI - The design of new carbonic anhydrase inhibitors. PMID- 11268530 TI - Roles of carbonic anhydrases in the alimentary tract. PMID- 11268531 TI - Carbonic anhydrase in the nervous system. PMID- 11268532 TI - Carbonic anhydrases in calcified tissues. PMID- 11268533 TI - Carbonic anhydrases of higher plants: an overview. PMID- 11268534 TI - Plant carbonic anhydrases: structure and mechanism. PMID- 11268535 TI - Algal carbonic anhydrase. PMID- 11268536 TI - Bacterial carbonic anhydrases. PMID- 11268537 TI - Keeping pace with a fast enzyme: steps and missteps. PMID- 11268538 TI - Carbonic anhydrase research: a clinical perspective, past and future. PMID- 11268539 TI - An overview of the distribution and function of carbonic anhydrase in mammals. PMID- 11268540 TI - The membrane carbonic anhydrases: from CO2 transport to tumor markers. PMID- 11268541 TI - [The challenge of informatics in primary care]. PMID- 11268542 TI - [Research activity on pediatric vaccines in Spain: descriptive analysis of prospective studies published between 1990 and 1998]. AB - OBJECTIVE: To describe the overall characteristics of prospective studies on vaccines in children, performed by Spanish investigators and published between 1990 and 1998. METHODS: Through a bibliographic research on MEDLINE and EMBASE, 24 prospective studies, performed in Spain, published as original papers, and with objectives directly related to the administration of vaccines to children have been identified. These studies were grouped as: clinical trials (group A), studies performed on established vaccination programmes (group B), and those that could not be included in the above mentioned groups (group C). RESULTS: 5, 9 and 10 studies belonged to groups A, B and C, respectively. More than 12,800 subjects participated in these studies, belonging to both normal population or specific risk groups. In 11 studies, the study population comprised newborns and infants. The vaccines under investigation were: hepatitis B (10 studies), DTPe/Pa (6), MMR (3), flu (2), Hib (1), hepatitis A (1), and meningococcus A and C (1) to address different objectives (in most of them, immunogenicity and/or reactogenicity). Nine had external financial support; 21 were performed by hospital and/or primary care investigators, and 18 in the Vasque Coutry, Madrid or Valencia. 13 publications reported obtaining informed consent, and 8 on the approval of the study protocol by an independent committee. Ten studies were published by international journals. CONCLUSIONS: This study shows that most of the studies are conducted by clinicians, with vaccines targetted to newborns and infants, with no external financial support, in a small number of autonomous communities, and usually published in Spanish Journals. The submission of this type of studies to a research ethics committee is desirable, something done to a lesser extent than obtaining informed consent. PMID- 11268543 TI - [Anxiety about death in primary care: relationship with frequency of consultation and psychomorbidity of patients]. AB - OBJECTIVES: To calculate the prevalence of psychological morbidity and anxiety about death among patients. To observe the association of these variables with the presence of chronic illness, consumption of psychiatric drugs and frequency of attendance. DESIGN: Cross-sectional, descriptive study. SETTING: Semi-urban primary care centre. PATIENTS: The study was proposed to 281 patients and accepted by 226 over 18 years old, selected systematically from among 1829 people who attended for consultation between May and July 1999. We calculated 30% prevalence of psychological morbidity. MEASUREMENTS AND MAIN RESULTS: Face-to face Interviews and questionnaires using the Goldberg Anxiety and Depression scales and Templer's scale of Anxiety about death. 63.7% women and 36.3% men responded, with an average age of 52.2 (SD 16.2). Positive Anxiety and Depression scale (prevalence of psychological morbidity) 55.3%; 61.8% in women and 43.9% in men (p = 0.009). Mean score on scale of anxiety about death 6.08 (SD 3.15); 6.66 in women (SD 2.67) and 5.05 in men (SD 2.67) (p < 0.0005). In over-attenders, anxiety about death was 6.61 (SD 3.23); and in those who are not, 5.81 (SD 3.10) (p = 0.074). The group with psychiatric morbidity had 38.4 mean visits (SD 27.28) against 35.5 (SD 22.59) for those without psychiatric morbidity (NS). In patients with psychiatric morbidity, the mean on the scale of anxiety about death was 7.51 (SD 3.33); and 5.33 (2.65) in patients without it (p < 0.0005). CONCLUSIONS: High prevalence of psychiatric morbidity. The majority are women, who have a mean value of anxiety about death which is greater than men's. We found no association between anxiety about death and over-attendance. The patients with psychiatric morbidity are more anxious about death. Chronic illness, age and consumption of psychiatric drugs are linked to a greater use of the clinic. We only found a relationship between psychiatric morbidity and a greater demand for health-care among the over-70s. PMID- 11268544 TI - [Family dysfunction as predisposing factor of mental disease. Does that relationship exist?]. AB - OBJECTIVES: The purpose is to describe the relationship between family disfunction and mental disorder. The secondary objective is to know the prevalence and distribution of mental disorders in primary care attended population. DESIGN: A cross-sectional study was conducted in a primary care setting. PATIENTS AND METHODS: Random sample was selected over 280 subjects from consultant population. The variables (family function, family structure, social and economic conditions and mental disorders) were collected through interview. APGAR test and Mini International Neuropsychiatric Interview test were performed. RESULTS: 264 patients were finally included (64% women). Mean age was 45.6 years (SD 16.7). Mental disorders were detected in 87 patients (33%). The most prevalent disorders were generalized anxiety disorder, dysthymia and major depression, family disfunction was found in 32 patients (12.3%). Prevalence of mental disorders wasn't statistically different in the group with family disfunction. CONCLUSION: Mental disorders are a common problem between primary care attended population. There wasn't any association between family disfunction and mental disorders, because of the limitations in the APGAR test in detecting family disfunction. PMID- 11268545 TI - [Do urocultures change our therapy approach?]. AB - OBJECTIVE: To describe to what extent the results of urocultures modify approaches to therapy and the factors linked to this change. DESIGN: Cross sectional, descriptive study by means of review of records. SETTING: Primary care. PARTICIPANTS: 222 adult urocultures requested at 8 health centres between March and May 1999. INTERVENTIONS: We extracted from the records age, sex, symptoms, risk factors, and approach to therapy before and after the uroculture. We discarded 358 urocultures because of not finding the clinical record or because the episode or data on the change in approach to therapy was lacking in the record. RESULTS: The urocultures belonged to patients with a mean age of 54.2, 73.1% of whom were women. 34.7% presented no risk factor for UTI. 44% had no symptoms of UTI. 21.2% of urocultures were positive, with E. coli the most frequently isolated bacteria (69.4%). Empirical antibiotic treatment was called for in 44.6% (70.1% quinolones, 9.3% fosfomycin). After receipt of the result, there was a change of approach in 25 cases (11.4%, SE 2.1%), of whom 15 did not receive empirical treatment (6 with symptoms and 9 without). The antibiotic was changed in 9 of the 99 cases treated empirically (always because of resistance). Among those with change of approach, there was a higher percentage of risk factors (84% against 62%, p < 0.05, chi 2 = 4.47). There were no differences for age, sex, symptoms or bacteria between the two groups. CONCLUSIONS: There is a quite considerable percentage of urocultures that lead to a change in approach to therapy, although most of the patients had linked risk factors and/or did not receive empirical antibiotic treatment. PMID- 11268546 TI - [Role of primary care teams in hospitalization of children under 2 years of age?]. AB - OBJECTIVE: To determine whether the structure of primary care teams on carrying out the healthy child health programme leads to a drop in the risk of admission to hospital of children under two, in comparison with the traditional clinic or out-clinic health system. DESIGN: Case-reference epidemiological study. CASES: 40% of the children under 24 months admitted to paediatric or neonate floors of the Marques de Valdecilla University Hospital. Reference: 15% of the recently born children alive in this hospital. Information was gathered through face-to face interview and by examining health cards. The study ran from April 1995 to May 1996. RESULTS: Children under two monitored habitually by a doctor belonging to a primary care team showed a drop in risk of hospital admission for all clinical diagnoses of 0.57 (95% CI, 0.35-0.93), after adjustment due to various confusion factors such as maternal education, social class, ethnic background, mother's age, mother's tobacco consumption, natural breast-feeding at birth, admission at birth. There was a drop of risk of hospital admission for high temperature without apparent cause in those children monitored habitually by a team doctor (adjusted RR = 0.41; 95% CI, 0.19-0.90). CONCLUSIONS: The advantages of the paediatric health care reform with the structuring of the primary care teams and the accompanying activities performed lead to a drop in the risk of hospital admission of those children under two years old who are habitually monitored by a doctor belonging to a primary care team. PMID- 11268547 TI - [Seasonal change in blood concentration of uric acid and its potential clinical implications]. AB - OBJECTIVE: To determine whether uric acid in plasma of patients registered at a PCC varies with the season. DESIGN: Descriptive study, with two transversal cuts, one in winter and one in summer. SETTING: Health district on the outskirts of Valencia. PATIENTS: Selected at random from all patients over 18 with medical records, with appointments in January-February and July-August 1999. Sample size was calculated for paired data with an alpha error of 0.05 and beta error of 20%. The pertinent level of uraemia was set at 0.4 mg/dl. Variability was deduced from a mini-sample of 17 cases in a sample of 72 patients. The following were recorded: sex, age, BMI, blood count, glucaemia, total cholesterol, HDL-c, LDL-c, uric acid and triglycerides. Meteorological data were supplied by the Valencia Weather Centre. MEASUREMENTS AND MAIN RESULTS: Temperature (12.8 degrees C versus 25.8 degrees C), sun (9.9 hours a day versus 5.9) and relative humidity (67.1% versus 61.4%) were greater in summer than in winter. There were no differences in mean atmospheric pressure (760.1 mmHg versus 760.7). 75 patients with a mean age of 63 finished the study. Mean uric acid was higher in summer at 5.64 mg/dl (95% CI, 5.29-5.99) than in winter at 5.23 mg/dl (CI, 4.92-5.56). We found no significant differences in the BMI, red corpuscles, haematocrits, glucaemia, total or divided cholesterol, or triglycerides. CONCLUSION: Seasonal variation in the plasma concentration of uric acid was found in the sample studied. PMID- 11268548 TI - [Test on knowledge about climacteric: development and validation process]. AB - OBJECTIVE: To construct and validate a test of knowledge about the menopause. SETTING: The context for use of this test is the Autonomous Community of Andalusia. METHOD: 1. Construction of the theoretical universe through the Delphi Technique. 2. Weighting of each of the dimensions of the theoretical universe. 3. Formulation and selection of the conceptual elements to introduce into the questionnaire and formulation of the corresponding items. 4. Selection of items and analysis of their internal consistency, homogeneity, stability and equivalence after a pilot study. PARTICIPANTS: The questionnaire's pilot study was run with 150 Andalusian women, chosen through sampling by quotas of age, habitat and educational background. MEASUREMENTS AND RESULTS: A questionnaire with 56 true and false (T/F) items grouped in four dimensions was obtained: biological aspects (14 items), psychological and social aspects (14), health risks linked to the menopause (6), care and preventive and health-improvement activities at the menopause (22). Its validity and reliability were determined by the following results: internal consistency and overall homogeneity (Cronbach's alpha = 0.86); stability or absence of significant differences at the moment the questionnaire is run (Student's t = 1.09; p = 0.28); intra-class correlation coefficient (ICC) = 0.753; equivalence or absence of bias of the questioners (Student's t = 1.09; p = 0.28), ICC = 0.761. CONCLUSIONS: The results lead us to conclude that the instrument is stable, allowing for the independence of the time and questioner variables, with high internal consistency and representing well the conceptual elements that shape the theoretical universe. PMID- 11268549 TI - [Evaluation of knowledge about climacteric in Andalusian women]. AB - OBJECTIVES: To evaluate the level of knowledge about the menopause of Andalusian women between 30 and 60 years old; and to determine their knowledge's relationship with social and demographic variables, health service use and their position as regards the climacteric. DESIGN: Cross-sectional study. PARTICIPANTS: A sample of 770 Andalusian women between 30 and 60 was chosen, for a sample error of +/- 5% and 95% confidence interval, chosen through multi-stage sampling: at random with proportionality for province and size of habitat, and sampling by age quotas and educational background. MEASUREMENTS: The principal study variable was women's level of knowledge about the menopause, evaluated through a validated test of 56 dichotomous questions. In the descriptive analysis, the mean, standard deviation and 95% CIs were obtained; in the analysis of relationships, the test of homogeneity of means, and variance and regression analysis were used. RESULTS: On a scale of 0-56, the mean on the sample was 18.17 with a standard deviation of 14.37 (95% CI: 17.15-19.17). 57.9% of the sample polled had values of low or very low level of knowledge. In the analysis of relationships, and according to the variance analysis data, we found that the level of understanding is related (p < 0.001) to age (F = 64.21), educational background (F = 131.19), type of menopause (F = 8.94), and having received information on the menopause (F = 7.57). Of these four variables it is educational background which most explains the variability in knowledge (r = 0.52, p < 0.001). However, experience of the menopause or use of the health services for the menopause do not seem to affect knowledge about the climacteric period and its treatment. CONCLUSIONS: The lack of knowledge of the menopause shown by the population studied demonstrates the need for Health Education on this stage of life. The relationships analysis leads us to conclude that the profile of women studied who knew most was someone with middle/high educational background, aged 30-40, who had had a surgical menopause, and had received information about the menopause. PMID- 11268550 TI - [Expectancy in urinary incontinence treatment from the perspective of an elderly population]. AB - AIMS: To determine the attitudes of community-dwelling elderly patients about the possibilities for cure of urinary incontinence. DESIGN: Cross-sectional study. INTERVENTIONS: A specific questionnaire on urinary incontinence and the Barthel Index Modified by Shah. LOCATION: Basic Health Zone of Cabra (Cordoba, Spain). PATIENTS: A random age-stratified sample of 793 patients selected from a total of 5139 persons > or = 65 years of age. MEASUREMENTS: Home interview on urinary incontinence (presence, characteristics, patient attitude regarding possibilities for cure). PRINCIPAL RESULTS: Forty-eight percent of elderly persons felt that incontinence could be cured, 21% thought that it could be improved, 9% stated that nothing could be done about it and the remaining 22% had no opinion. A significant association was found between the type of response and age, sex, state of health as perceived by the subject and level of self-care. The presence of incontinence did not influence the nature of the responses given by patients about the possibilities for successful treatment of incontinence. CONCLUSIONS: Our elderly persons were not aware of the possibilities for treatment of incontinence, a fact that may have a negative impact on sufferers seeking help for this remediable problem. PMID- 11268551 TI - [Impact of a community network of psychosocial support in perinatal health]. AB - OBJECTIVE: Determine the impact of a community intervention in perinatal health in, cities of the State of Chihuahua, Mexico. PATIENTS, MATERIAL AND METHODOLOGY: A community intervention was designed including pregnant women (n = 1233), and finally 1148 (6.74% of loss) group intervention (n = 261) control group (n = 887). Intervention consisted of five visits prior to delivery during the 22nd, 26th, 30th, 34th and 38th weeks, as well as a visit during the first 8 days in puerperium, providing social and educational support. In regards to the control group, they were visited in two different occasions, week 22 and purperium, indicating them to continue with their regular care. The outcome variables were perinatal mortality, premature birth (< 37 weeks), low weight at birth (< 2500 grams) and perinatal morbidity. RESULTS: Perinatal mortality rate was smaller in the intervention group 3.8 per 1000 born alive; perinatal mortality in the control group was 13.1. No statistically considerable difference was found in premature birth, in low weight at birth and in perinatal morbidity. CONCLUSION: Community intervention provides a support to pregnant women under psychosocial risk factors; nevertheless, it does not contribute to considerably decrease premature birth, low weight at birth or perinatal complications. PMID- 11268552 TI - [Informatics in primary care (I)]. PMID- 11268553 TI - [Do we prescribe omeprazol properly?]. PMID- 11268554 TI - [From evidence based medicine to medicine based on what is evident]. PMID- 11268555 TI - [Prevalence of female ejaculation. A failed study]. PMID- 11268556 TI - [Hyperprolactinemia secondary to hypotensive treatment with verapamil]. PMID- 11268557 TI - [Ischemia-reperfusion tissue injuries and possibilities of pharmacologic intervention]. AB - Tissue can be paradoxically more severely damaged by reperfusion than by ischaemia itself (ischaemic-reperfusion injury--IRI). The mechanism of IRI has been intensively studied. Both the excessive formations of reactive oxygen radicals and calcium overload participate in the development of IRI. However, the significance of interaction between neutrophils and endothelial cells has been revealed recently. Imbalance of factors formed by endothelial cells and those formed by blood elements as well as the increased tendency to adhesion of neutrophils to the vascular endothelia during IRP has been proved. IRP effects on tissues can be pharmacologically influenced by: 1. anti-oxidative therapy, 2. substitution of cytoprotective factors formed in endothelial cells and blood elements, 3. inhibition of cytotoxic factors of endothelial cells and blood elements, 4. influencing the adhesion of neutrophils to the vascular endothelia, 5. administration of drugs acting by several of the above mentioned mechanisms. However, beside the application of the anti-oxidative therapy during the organ transplant operations, only few of those approaches has been used in the clinical practice. PMID- 11268558 TI - [Portal hypertension and the endothelium]. AB - Portal hypertension is defined as a rise in the pressure gradient between the portal vein and the inferior vena cava by more than 10 mmHg. The basic factors in the development of portal hypertension include increased resistance in intrahepatic circulation, increased portal blood flow, and development of the hyperdynamic syndrome. Apart from structural changes in the hepatic tissue, the major role in the pathogenesis of portal hypertension is played by endothelial dysfunction. In patients with portal hypertension, endothelial dysfunction reduced the activity of vasodilator, anti-aggregation, and anti-adhesion substances (NO/EDRF, prostacyclin, CO) and to increased production of vasoconstrictor and proliferative substances (endothelins in particular) in intrahepatic the circulation. On the other hand, vasodilator agents become predominant in splanchnic and systemic circulation. As a result, the intrahepatic vascular resistance and portal blood flow rise. It was followed by a subsequent increase in portal pressure and the development of hyperdynamic circulation. Identification of the pathophysiological mechanisms of portal hypertension has paved the way to new trends in pharmacological therapy. The aim of effective treatment of portal hypertension is the permanent reduction of the portal pressure, which markedly decreases the risk for complications, and the risk for esophageal variceal bleeding in particular. The currently most preferred agents are those with vasodilator and vasoconstrictor effects. PMID- 11268559 TI - [New findings on the role of leptins in regulation of the reproductive system in humans]. AB - The reproductive activity is closely related to the nutritional status of the organism. A new hormone--leptin has been discovered a few years ago, which informs the brain about fat supplies in the body. Leptin participates in the regulation of the reproductive system at both, peripheral and central, levels. There are discussions about leptin as a possible trigger of puberty. The prepubertal leptin levels are low in both sexes; they increase with the onset of puberty. It has been suggested that leptin resistance might exist in the childhood, allowing the onset of the puberty only at the moment, when the adipose stores are adequate for the strongly demanding period of the adolescence. The role of leptin in pathogenesis of polycystic ovary syndrome (PCOS) is studied. The significance of leptin during pregnancy and after birth remains still unclear. PMID- 11268560 TI - [Effect of ultrasound on sex differentiation and embryonic mortality]. AB - Effects of ultrasound on the sex differentiation and embryonic mortality was tested on the set of 772 eggs of the Rhode Island Red and Hampshire strain chicken. In control groups, embryos were not exposed to ultrasound during incubation. In experimental groups, chicken embryos were exposed to ultrasound generated by the transducer oscillating at the frequency of 30 kHz with the power of 60 W. Embryonic mortality was significantly higher in the experimental groups exposed to ultrasound. Pathogenic effects of ultrasound were more pronounced in embryos with already developed allantochorionic blood circulation. It can be assumed that the impairment of allantoid vessels reduced in the intensity of oxidative processes in embryonic tissues. The decrease of blood pH, resulting from the carbon dioxide accumulation, set the sex differentiation in flavour of males. PMID- 11268561 TI - [Cholinesterases and their importance in the etiology, diagnosis and therapy of Alzheimer's disease]. AB - Cholinesterases belong to esterases and represent important animal enzymes with multiple biological function. Acetylcholinesterase (AChE) plays the key role in cholinergic neurotransmission, whereas the function of butyrylcholinesterase (BuChE) is still unrevealed. Both enzymes seem to act as neurogenic factors during the early embryogenesis and later on they may participate in some pathological alteration. In humans, these degenerative changes are related to the deposition of pathologic proteins in the brain and with the progress of the Alzheimer's dementia (AD). Both AChE) and BuChE become recently the target for the most frequently used therapy of AD--cholinesterase inhibitors. PMID- 11268562 TI - [Competence and quality in medical practice. Or the necessity for organising continuing medical education in Lebanon]. PMID- 11268563 TI - Karyotype of amniotic fluid cells at the AUB-MC results on 2000 cases. AB - We report results on 2000 cases of amniotic fluid referred for karyotype analysis. Referrals were advanced maternal age in 64% of cases and abnormal ultrasound in 12% of cases. The frequency of chromosome aneuploidy was 2.4% in the first category and that of chromosome abnormalities 8% in the second. The incidence of marker chromosomes was 0.25%, that of mosaicism 0.3%, and maternal cell contamination was observed in 0.6% of cases. The overall culture failure rate was 0.9%. Our results are mostly in accordance with figures from larger surveys, published in the literature and differences might be due to the smaller number of samples in this series and variation in referral and/or sampling protocols. PMID- 11268565 TI - [Otosclerosis: experience at the Hotel-Dieu in France. 71 surgical cases]. AB - Otosclerosis is one of the common causes of hearing loss. The incidence varies between 0.1% and 2%. In Lebanon otosclerosis is a common entity that has not been well evaluated. To the best of our knowledge there is no epidemiologic analysis of the incidence or outcomes of otosclerosis in Lebanon. We collected the number of stapedectomies performed for otosclerosis in different hospitals between Jan. 1994 and Dec. 1995. We also retrospectively reviewed the charts of 71 cases who underwent stapedectomy at Hotel-Dieu de France-St Joseph University Hospital Medical Center, Beirut, Lebanon. Between 1992 and 1996 the incidence of otosclerosis in Lebanon as revealed through stapedectomy is 5/100,000. We report also on the pathology, technique, complications and outcomes of stapedectomy surgery for otosclerosis in Hotel-Dieu Hospital. Further epidemiologic studies and screening is required to reveal the exact incidence of this common entity that could be underdiagnosed or untreated in our country. PMID- 11268564 TI - Atypical squamous cells of undetermined significance (ASCUS), low-grade squamous intraepithelial lesion (LSIL), high-grade squamous intraepithelial lesion (HSIL) and histology. AB - OBJECTIVE: A new terminology for cytologic diagnosis of cervical lesions has been introduced by Bethesda System. This includes: 1) Atypical squamous cells of undetermined significance (ASCUS), 2) Low-grade squamous intraepithelial lesion (LSIL), 3) High-grade squamous intraepithelial lesion (HSIL), 4) squamo-cellular carcinoma. The aim of this study was to assess the correlation between the colpocytologic test (Pap psmear), the histologic response and colposcopy. METHODS: We re-examined the cytologic results of 447 patients who underwent routine cytologic tests, with a diagnosis of various grades of atypia, from ASCUS to HSIL. A histologic test was carried out on a colposcopic basis in 210 cases and cytologic results were correlated with the histologic and colposcopic pictures. RESULTS: For ASCUS the histology was positive in 19.1% (31/163) of cases and negative in 10.4% of the colposcopically positive cases (17/163), while in the remaining 70.5% (115/163) the colposcopy resulted negative. In LSIL with the presence of human papillomavirus (HPV) the histologic findings, confirmed the cytologic result in 35.8% (48/134) while in the group of cervical intraepithelial neoplasia (CIN I) results corresponded in 40.6% (41/101). In high squamous intraepithelial lesions there was an histologic confirmation in 59.2% (29/49). CONCLUSIONS: The data obtained indicate an increase in positive histology results from the types of low grade (ASCUS-LSIL) to those of a high grade (HSIL). Our results indicate that the presence of ASCUS should be assessed colposcopically and histologically, where indicated. PMID- 11268566 TI - Febrile neutropenia in cancer patients in a tertiary care medical center in Lebanon: microbial spectrum and outcome. AB - We prospectively analyzed the episodes of febrile neutropenia at the American University of Beirut Medical Center. One hundred and four episodes were studied in 64 patients over a period of 15 months: 81 (78%) with leukemia, 11 (10.5%) with lymphoma, 3 (2.8%) with multiple myeloma, and 9 (8.6%) with solid tumors. Bacteremia was confirmed in 30 episodes (29%), of which 18 (60%) were caused by gram-negative bacilli and 12 (40%) by gram-positive cocci. The predominant organisms were: E. coli (9), coagulase negative staphylococci (CNS) (6), Pseudomonas aeruginosa (5), and S. aureus (4). In seven episodes (6.7%) urinary tract infections were diagnosed, 6 with E. coli. Sputum cultures were positive in eight episodes (7%), 2 with P. aeruginosa, and 2 with methicillin resistant S. aureus. All patients were started empirically on antibacterial agents. In twenty one episodes, a single antibiotic was started, ceftazidime being the most commonly used agent. In most cases, however, 2 or 3 antibacterial agents were started empirically. Antifungal therapy with amphotericin B (11) or fluconazole (20) was added because of persistent fever despite broad antibacterial coverage. Thirteen patients died (20%), 6 of them had bacteremia; 2 with gram-negative bacilli, and 4 with gram-positive cocci. Except for one, all patients had been started, at the onset of the fever, on antimicrobial agents to which the isolated microorganisms turned out to be susceptible. Our results show that infections with gram-negative bacteria continue to predominate unlike what has been reported recently from European and North American trials. A trend toward a higher mortality of infections caused by gram-positive cocci was noted. PMID- 11268567 TI - Bacterial etiology of otitis media with effusion in a group of Lebanese children. AB - Identification of the main bacteria causing otitis media with effusion (OME) in a given population is essential. It indicates the degree of involvement of a given bacterium in a particular disease of that population. Knowledge of the most prevalent bacteria would initiate the search for the mode of acquisition of such bacteria and may aid in establishing appropriate control and prevention programs, which may decrease the incidence of OME. The rapid response of most OME to a variety of broad-spectrum antimicrobials deprives the clinician from knowing the particular bacteriologic agent prevailing in a community. With the emergence of resistant strains and the change over time of the relative distribution of bacteriologic agents known to cause OME, the identification of the bacterial etiology of OME in Lebanese children was initiated. PMID- 11268568 TI - [Radical surgical treatment of vulvar epidermoid cancer]. AB - During an eighteen-year period, forty-nine patients with invasive epidermoid carcinoma of the vulva were treated by a radical vulvectomy at the Hotel-Dieu de France hospital in Beirut. The age, the clinical presentation, and the lymph nodes involvement were close to what was published in the literature. The overall five-year survival rate was 72%. Those with stage I disease lesions had an excellent survival rate. A review of the literature would indicate that a conservative approach in the treatment of early invasive lesions can give satisfactory results, and that adjuvant radiotherapy is more beneficial than pelvic lymph nodes dissection when there is a high risk of pelvic involvement. PMID- 11268569 TI - [Chronic suppurative otitis media. Experience with 140 cases at the Hotel-Dieu of France]. AB - OBJECTIVES: Chronic suppurative otitis media (CSOM) is a common disease especially in developing countries. In Lebanon, CSOM is a frequent complaint. This retrospective study is an objective evaluation of the cases we operated in HDF. METHODS: One hundred and forty cases of CSOM were collected between 1993 and 1997: 88/140 cases were simple CSOM and the others 52/140 cases had cholesteatomas. RESULTS: There were no immediate postoperative complications reported (< 6 weeks). Beyond 6 weeks, with a mean follow-up of 20 months, 15% of the patients were lost for follow-up and we had 93.4% of success rate in CSOM and 88% in cholesteatomas. The recurrence rate of cholesteatoma was 14%. Children's cholesteatoma (below 15 years of age) were more aggressive than adult's cholesteatomas. In CSOM without cholesteatoma, the hearing gain was 5 dB. In CSOM with cholesteatoma the mean hearing gain was 4 dB. In conclusion the study revealed the importance of this pathology in our country with the relative high incidence of cholesteatoma compared to simple CSOM, this could be explained by the chronicity of the disease and the lack of preventive measures. PMID- 11268570 TI - Low-dose aprotinin in cardiac operations. AB - BACKGROUND: Patients with heart disease are frequently maintained on a regimen of aspirin. During cardiac surgery aspirin-induced platelet dysfunction increases the risk of bleeding. METHODS: The files of 82 patients who had undergone open heart surgery were selected to study the efficacy of the low-dose (2 million KIU) aprotinin regimen in decreasing blood loss and transfusion requirements for patients receiving aspirin. Patients were divided into three groups: Group 1, includes those who received neither aspirin nor aprotinin (n = 41), Group 2, received both aspirin and aprotinin (n = 27) and Group 3, did not receive aprotinin despite aspirin intake (n = 14). Primary outcome measures in this study were total volume of blood loss (postoperative chest tube drainage) and volume of transfusions during hospitalization. RESULTS: Patients treated with aprotinin (Group 2) had significantly lower total blood loss (359 ml versus 527 ml and 628 ml in Group 1 and Group 3, p < 0.05), rates of bleeding (17.2 ml/hr versus 25.7 and 30.4 ml/hr in groups 1 and 3 respectively). A significant difference was also found when comparing the volume of blood transfusions (224.4 ml versus 262.4 and 537.5 ml) and prevalence of transfusion (33.3% versus 36.6% and 64.3%). CONCLUSION: Low-dose aprotinin significantly reduces blood loss and blood transfusions in patients receiving aspirin who undergo cardiac operations. PMID- 11268571 TI - The government health center role redefined: cervical cancer screening in the Shouf area. PMID- 11268572 TI - [Pseudo-Takayasu in Behcet's disease]. AB - Behcet's disease is a chronic multisystem vasculitis that is frequent in Lebanon. The great arteries involvement is rare. We report here an unusual case of subclavian artery occlusion (pseudo-Takayasu) with a literature review. PMID- 11268573 TI - [An unusual cause of acute myelopathy: a dural arteriovenous fistula at the craniocervical junction]. AB - BACKGROUND: Dural arteriovenous fistulas (DAVF) account for 10% to 15% of all intracranial arteriovenous malformations. Since the first case published by Woimant et al. in 1982, many type V DAVF, i.e. with spinal venous drainage, have been reported. Fistulas located at the craniocervical junction (CCJ) however, are exceptional and only 10 cases of CCJ fistulas associated with myelopathy have been described. CASE REPORT: The authors present a 36-year-old male patient without previous medical history, suffering from acute myelopathy. Cervical MRI showed multiple serpiginous flow-voids along the cord surface and cerebral angiography disclosed a dural fistula of the CCJ fed by the right posterior meningeal and occipital arteries. The venous drainage was directed caudally towards the perimedullary veins. Embolization through the occipital artery, using cyanoacrylate, was performed and resulted in complete cure of the malformation with rapid clinical recovery. DISCUSSION: The authors discuss the pathophysiology and clinical consequences of intracranial DAVF with myelopathy (named V, m+), that are usually identical to those of spinal dural fistulas and related to intramedullary venous hypertension. Early treatment is essential to reverse the patient's myelopathy. Embolization, if technically possible, is the preferred treatment and cyanoacrylate remains the best embolic agent. Following glue deposition, systemic high-dose steroids should be administered to prevent edema. CONCLUSION: In conclusion, this is the first case of DAVF of the foramen magnum causing myelopathy to be detected early and cured by glue embolization alone, with rapid and total clinical recovery. PMID- 11268574 TI - [Mapping of brain function with positron emission tomography for pathophysiological analysis of neurological disorders]. PMID- 11268575 TI - [Functional brain mapping using synthetic aperture magnetometry(SAM)]. PMID- 11268576 TI - [Study of human speech function by magnetic stimulation]. PMID- 11268577 TI - [High-field MR and brain functional analysis]. PMID- 11268578 TI - [Optical imaging of brain function]. PMID- 11268579 TI - [Definition of individual language related area by awake surgery]. PMID- 11268581 TI - [Microcirculatory changes in the development of the cerebrovascular adaptation]. AB - Recently, long-term cerebrovascular adaptations after unilateral carotid ligation that increase the tolerance of the brain to subsequent episode of ischemia was reported(J Cereb Blood Flow Metab, 1998). However, the pathophysiological mechanisms underlying the phenomenon was unknown. We examined regional cerebral blood flow(rCBF) before, during and after subsequent ischemia in the development of the adaptation. Male Wistar rats(n = 18) were used. Unilateral(right) carotid artery ligation was performed 3 hours(group A: n = 8), 3 days(group B: n = 10) before forebrain ischemia. With bilateral carotid arteries occlusion, mean arterial blood pressure(MABP) was reduced by hypobaric hypotension down to 50 mmHg and maintained constant for 30 min. After hypotension, the experiment continued for 90 min. Local CBF were registered at 25(5 x 5) identical locations by laser Doppler scanning in the cortex during the experiment in bilateral hemispheres(at control phase, after the left carotid artery ligation, during ischemia and after hypotension). The physiological variable such as blood pressure and gas analysis were within normal range in all groups. There were no differences of rCBF between the right and left hemispheres in group A during the experiment. In group A, rCBF of both hemispheres significantly decreased after the ligation of the left carotid artery, during the induced ischemia (P < 0.05), and recovered to the value of before ischemia. Whereas, in group B rCBF of the left side(non-occluded side) decreased after the ligation of the left carotid artery, during the induced ischemia(P < 0.05), and recovered to the value of before ischemia, but rCBF of the right side(occluded side) decreased only during the induced ischemia(P < 0.05) and rCBF of the right side was significantly higher than the left brain after ligation of the carotid artery, during ischemia and after hypotension(P < 0.05). On the basis of these data, we concluded that cerebrovascular adaptations can be acquired by 3 days after unilateral carotid artery occlusion and the tolerance phenomenon is certainly attributed to microcirculatory improvement. PMID- 11268580 TI - [Prevalence of Martin-Gruber anastomosis on motor nerve conduction studies]. AB - Prevalence of median to ulnar anastomosis in the forearm(Martin-Gruber anastomosis; MGA) to the first dorsal interosseous(FDI), abductor digiti quinti (ADQ) and adductor pollicis(AP) was investigated. Subjects contained 106 patients with normal nerve conduction or patients with various neuropathies. Recording electrodes were placed on the motor point of FDI, ADQ and AP. Supramaximal stimulations were given to the median and ulnar nerves at the wrist or above the elbow. The diagnosis of MGA was made by the following criteria; amplitude of compound muscle action potential(CMAP) increased after elbow stimulation as compared with the wrist stimulation in median nerve conduction studies. The corresponding decrease in CMAP amplitude was found after above elbow stimulation as compared with the wrist stimulation in ulnar nerve conduction studies. No MGA was found in 80(75%) out of 106 patients. MGA to FDI was found in all 26 patients who had MGA. MGA to ADQ and AP was found in 11% and 10% of the patients, respectively. Only 8 out of 26 patients had MGA to all 3 muscles. In the presence of MGA median motor nerve conduction studies demonstrate larger CMAP, with a small initial positivity, after elbow stimulation than after wrist stimulation. And moreover, ulnar motor nerve conduction studies reveal a conduction block-like finding in the forearm. In this study MGA was found in 25% of the patient to FDI, in 11% to ADQ and in 10% to AP. Although a very small MGA might be overlooked in our method, such a small MGA doesn't mislead us into erroneous interpretation of motor nerve conduction studies. PMID- 11268582 TI - [Cerebral infarction presenting pure motor monoparesis: diagnosis by diffusion weighted MR imaging]. AB - We studied 10 patients with acute ischemic cerebrovascular disorders presenting paralysis confined to one limb, unaccompanied by sensory signs(pure motor monoparesis, PMM) on diffusion-weighted MR imaging(DWI). DWI revealed fresh ischemic lesions in all patients, except for 2 cases of transient ischemic attack. On DWI, acute infarction in multiple lesions was identified, and small superficial lesions were clearly described. Superficial lesions were seen in 4 patients, and deep lesions were also seen in 4 patients. DWI is useful for lesion analysis in cerebral infarction with PMM. PMID- 11268584 TI - [Acute cerebellitis presenting as acute hydrocephalus: case report]. PMID- 11268583 TI - [A case of central nervous system anomalies (agenesis of corpus callosum, colpocephaly, hydrocephalus, congenital dermal sinus) associated with congenital heart disease(double-outlet right ventricle, complete endocardial cushion defect, atrial septal defect, pulmonary arterial stenosis, patent ductus arteriosus)]. AB - A case of central nervous system anomalies(agenesis of corpus callosum, colpocephaly, hydrocephalus, congenital dermal sinus) associated with congenital heart disease(double-outlet right ventricle, complete endocardial cushion defect, atrial septal defect, pulmonary arterial stenosis, patent ductus arteriosus) is reported. Female patient had been already diagnosed as hydrocephalus during pregnancy and ventricular drainage was performed soon after the delivery. Prostaglandin E 1 was also applied for heart disease, but saturation of O2 decreased to 80% on arterial blood gas analysis. Blalock-Taussig operation and ligation of ductus arteriosus was done 41 days after the delivery and ventricle peritoneal shunt was also made for the progressive hydrocephalus on the same day. Chromosome analysis showed no abnormality. The genesis of this complicated brain and heart anomaly is discussed from the viewpoint of neural crest cell abnormality. PMID- 11268585 TI - [Physiology of appetite and feeding behavior: introduction]. AB - Food intake is regulated by the central nervous system depending on macronutrients and environmental changes. The hypothalamus is the target of hunger and satiety signals arising from the peripheral organs and the brain. Noradrenaline-neuropeptide Y and opioid-galanine are involved in carbohydrate and fat intake, respectively, while serotonin-CCK-insulin and dopamine-cyclic dipeptides systems inhibit them. Histamine and proinflammatory cytokines are involved in stress- and sickness-induced anorexia. Leptin accelerated intrahypothalamic anorexic mechanisms executed by POMC/CART and CRH but suppresses orexigenic mechanisms promoted by NPY and orexin. Although these mechanisms elegantly regulate appetite and feeding behavior, disruption of weight control has been accelerated and the incidence of obesity and eating disorder are dramatically increasing recent years in our modern society. New approach may be necessary to solve the problems of weight control. PMID- 11268586 TI - [Functional relationship between circadian rhythm and control of food intake]. AB - Living things on the earth including bacteria, plants and animals show circadian rhythms in their behaviors and physiological phenomena, and these circadian rhythms are usually synchronized with environmental changes having the period of 24 h on the earth. In mammals including human beings, the hypothalamic suprachiasmatic nucleus (SCN) functions as a master circadian oscillator, and generates a circadian rhythm of food intake. Sometimes the circadian oscillation of the SCN is disturbed with physical and psychological stressors. This review describes the functional relationship in respect to connections between the circadian oscillator in the SCN and food regulatory centers and neurons in the brain focusing on its mechanism in human beings, and a possible involvement of the circadian oscillator of the SCN in the abnormality of the appetite control. PMID- 11268587 TI - [Role of leptin and its receptor in the regulation of appetite and body fat]. AB - The role of leptin and its receptor on the regulation of appetite and body fat was summarized. Leptin directly exerts its anorexigenic effects on arcuate nucleus via proopiomelanocortin and neuropeptide Y neurons. The anorexia and sympathetic nerve activation result in the reduction of body fat. But physiological concentrations of leptin could not reduce body fat in obese people, while genetic loss of central leptin effects induces obesity in children. Melanin concentrating hormone, orexin, and corticotropin-releasing hormone may be directly regulated by leptin. Serotonergic neurons may be separate from leptin effects. Phosphorylation of 985- and 1138-tyrosine of long-form leptin receptor activates SHP-2 and STAT3, respectively. Soluble leptin receptor concentrations in serum are negatively correlated with BMI. Clinical usefulness of leptin is now in progress. PMID- 11268588 TI - [A role for orexins and melanin-concentrating hormone in the central regulation of feeding behavior]. AB - The hypothalamus is the most important region in the control of food intake and body weight. The ventromedial 'satiety center' and lateral hypothalamic 'feeding center' have been implicated in the regulation of feeding and energy homeostasis by various studies of brain lesions. Orexins(orexin A and orexin B) and melanin concentrating hormone (MCH), whose intracerebroventricular injections increase food intake, are localized in the lateral hypothalamus and provide diffuse projections throughout the brain. Orexins and MCH neurons have a coextensive distribution, but are not colocalized. Orexins and MCH may affect feeding behavior through distinct neuronal pathways. We here describe the effect of orexins and MCH on feeding behavior from the physiological, neuroanatomical and molecular studies. PMID- 11268589 TI - [Regulation of appetite by melanocortin and its receptors]. AB - alpha, beta, gamma-MSH and ACTH are derived from the same precursor, POMC(proopiomelanocortin), and are classified as melanocortin. alpha-MSH plays an important role in the regulation of appetite and energy expenditure via central melanocortin receptor, melanocortin 4 receptor(MC4R), which is expressed mainly in hypothalamus. alpha-MSH or its analogue shows inhibitory effect on appetite and inversely MC4R antagonist stimulates appetite. MC4R knock-out mice has adult onset obesity and decreased energy expenditure. POMC gene expression in hypothalamus is partially regulated by leptin. Agouti-related peptide(AgRP), a homologue of agouti peptide and antagonist of MC3R and MC4R, is expressed in human brain and may act as a inhibitor of alpha-MSH. From the genetical aspect, the region near POMC gene, 2p23, is one of the susceptibility loci of human obesity. POMC gene mutations are found in two families, where mutations in both alleles cause human obesity, red hair, adrenal dysfunction, due to alpha-MSH and ACTH deficiencies. In morbidity obese patients, heterozygous MC4R gene mutations are found among 4% of them. These results suggest the importance of melanocortin and its receptors on appetite regulation in human. PMID- 11268590 TI - [The relationship between beta 3-adrenoceptor and regulation of body fat mass, and food intake]. AB - beta 3-adrenoceptor(beta 3-AR) plays important roles in thermogenesis of brown adipose tissue(BAT) and lypolysis of white adipose tissue(WAT). Anti-obesity effect of beta 3-agonists is reported, and the Trp64Arg point mutation of the human beta 3-AR gene is associated with abdominal obesity. beta 3-agonist decreases food intake in rat and mice, and its effect is confirmed by both direct infusions to the brain and peripheral injections. Stimulated thermogenesis of BAT increases glucose utilization, then 'glucose dip' signals meal initiation. Risen core temperature leads to meal termination. But, because of decreased ability for thermogenesis, meal size increases in many obese animal models. Further investigations are being carried out to make these problems clear. PMID- 11268591 TI - [The role of anorectic and orexigenic peptides(CART, NPY etc)]. AB - Leptin inhibits food intake and increase energy expenditure via an interaction with specific leptin receptors located in the hypothalamus. Leptin receptors have been identified in cocaine and amphetamine-regulated transcript(CART), neuropeptide Y(NPY), galanin-containing neurons of the arcuate nucleus, respectively, and in corticotropin-releasing hormone(CRH)-containing neurons of the paraventricular nucleus, suggesting that these peptides are mediators of leptin's action in the hypothalamus. Anabolic pathways such as those including NPY and galanin promote feeding and are inhibited by leptin, whereas catabolic pathways which include CART and CRH have the opposite effect and are stimulated by leptin. In the current review we examine the significant role of the CART, CRH, NPY and galanin in the regulation of energy balance. PMID- 11268592 TI - [Genetic abnormalities of regulatory mechanism of appetite]. AB - Recently, leptin was cloned and characterized as a sateity factor which acts through the hypothalamus. alpha-melanocyte-stimulating hormone derived from pro opiomelanocortin(POMC) and melanocortin receptor-4(MC4-R) have been reported to be involved in the downstream of the effect of leptin. In this paper, we summarized the clinical characteristics and the mechanisms of obesity caused by genetic abnormalities involved in the regulatory mechanism of appetite such as leptin, leptin receptor, POMC, MC4-R and prohormone convertase 1. PMID- 11268593 TI - [Distorted control of feeding behavior induced by abnormally regulated peripheral energy metabolism]. AB - To regulate feeding behavior and energy metabolism, a variety of neuronal and hormonal messages are integrated by glucose sensing system located in peripheral organ and central nervous system. Among them, a hepatoportal glucose sensor plays a major role in perception of peripheral humoral information such as brain-gut peptides and cytokines. These signals are transmitted to the hypothalamus through the hepatic afferent vagus nerve and the nucleus of the solitary tract. Recent molecular approaches using knock out or transgenic mice have indicated important effects of peripheral energy metabolism on feeding control. Particularly, fatty acid metabolism in the liver and functions of uncoupling protein family in the brown adipose tissue and the muscle play an essential role as detector for energy storage, and in turn may be contributable to regulation of feeding behavior. PMID- 11268594 TI - [Genetic abnormality and the mechanism of ingestive disorder in obese animal model]. AB - The objective of this review article is to present the genetic abnormalities in obese animal models up to present times and to suggest the mechanism of ingestive disorder. Leptin is an anorectic ob gene product and activates the anorexigenic POMC and CART neurons in the ARC and suppresses orexigenic NPY and AGRP neurons. TUB gene product also activates the anorexigenic POMC neurons. These anorexigenic neurons project to the second-order hypothalamic neuron, CRH, TRH and so on in the PVN and suppression of orexigenic neurons project to the orexin in the LHA. PMID- 11268595 TI - [Pathology and significance of leptin resistance in obesity]. AB - Leptin, the protein product of the ob gene, is predominantly secreted from white adipose tissue, and acts on the brain to regulate food intake, energy expenditure, and neuroendocrine function. Obese rodent and humans are mostly associated with high circulating leptin levels. These findings have led to the conclusion that obese individuals are relatively insensitive to endogenous leptin termed 'leptin resistance'. The potential sites for leptin resistance include the blood-brain-barrier transport system and the leptin signaling mechanism in leptin responsive neurons in the hypothalamus. In this review, we describe leptin, leptin receptor, and potential hypothesis of leptin resistance. PMID- 11268596 TI - [Molecular mechanism in the development of the complications associated with obesity--the physiological and pathological role of adipocytokines]. AB - Visceral fat accumulation often accompanies various complications, such as insulin resistance, hypertension, dyslipidemia and atherosclerosis. Adipose tissue has been found to secrete various biologically active adipocytokines including free fatty acids. Accumulation of visceral fat increases the portal free fatty acids concentration to cause insulin resistance and dyslipidemia. Tumor necrosis alpha (TNF alpha) deteriorates insulin resistance in obesity. The levels of plasminogen activator inhibitor(PAI)-1 increase and plasma adiponectin concentration decreases in obesity leading to the development of vascular disease. The finding of genes specifically expressed in visceral fat and new adipocytokines should facilitate clarification of the mechanism for the development and complications of visceral fat accumulation. PMID- 11268597 TI - [Abnormality in feeding and body weight regulation in obesity]. AB - Obesity is, with rare exceptions, a complex phenotype resulting from interactions between environmental and genetic risk factor. Obesity is associated with diabetes, hypertension, and hyperlipidemia, leading to arteriosclerosis. Dysregulation of food intake and energy expenditure, and thus energy homeostasis, is now recognized as playing a major role in development of obesity. A detailed understanding of the physiology and genetics of the regulatory systems is critical for the development of new approaches for treating obesity and its sequelae. PMID- 11268598 TI - [Eating disorders concurrent with type 1 diabetes: pathology and management]. AB - Development of disordered eating is not unusual in young females with type 1 diabetes. Although the debate continues over whether or not young diabetic females are at increased risk for developing eating disorders, no one questions the devastating effect of eating disorders on the clinical course of diabetes, successful intervention is exceedingly difficult. This manuscript introduces the most current research on the epidemiology, pathology, possible mechanism of development, and management of eating disorders in type 1 diabetes. It also discusses ongoing study at Kyushu University related to the clinical characteristics of and therapy for females with type 1 diabetes and recurrent binge eating. PMID- 11268599 TI - [The characteristics and mechanism of onset of abnormal eating behavior due to stress or psychosomatic diseases]. AB - Abnormal eating behavior is characterized by two major categories. One, stress induced abnormal eating behavior, is characterized by abnormality of degree and frequency of diet, which stands for appetite loss or appetite gain. They are connected with psychosocial or somatic stress and show temporary disorder for almost all the people. The other, abnormal eating behavior accompanied with psychosomatic diseases, is characterized by chronically devastated cognition and behavior of diet, which represents Anorexia Nervosa(AN) or Bulimia Nervosa(BN). The characteristics and mechanism of onset of these disorders are generally investigated. Abnormal eating behavior in AN or BN are associated with distorted cognition and adjustment difficulty against stress and psychological problems. PMID- 11268600 TI - [Appetite and regulation of food intake in uremia]. AB - Anorexia, nausea and vomiting are common symptoms in patients with severe renal failure. Abnormalities in body fluids volume, serum electrolytes concentrations and acid-base balance and accumulations of uremic toxic substances might be primarily contributing to the suppression of appetite. However, the detailed mechanisms that cause appetite suppression in uremia are poorly understood. Impaired gastric emptying, constituents of peritoneal dialysate, plasma high leptin, high cholecystokinin and low neuropeptide Y are also considered to be contributing to appetite suppression in uremia. PMID- 11268601 TI - [Mechanisms of cancer-induced anorexia]. AB - Possible mediators of anorexia-cachesia syndrome are hormones, cytokines, and leptin at the peripheral tissues, and neuropeptides, cytokines, and hormones in the hypothalamus. Insulin resistance and relative hyper-glucagonemia might stimulate anorexia. Interleukin(IL)-6, and tumor necrosis factor(TNF)-alpha as well as IL-1 and leukemia inhibitory factor(LIF) are some of those cytokines. Although leptin in the adipose tissue is an attractive addition, changes in the tumor-bearing state does not support its active role in the induction of anorexia. Newer neuropeptides related with anorexia and oxygenia have been found in the hypothalamus. Hypothalamic neuropeptide Y(NPY) and ciliary neurotropic factor(CNTF) are some of those promising mediators. Although current trend has been progestational drugs, newer drugs such as anticytokines, anabolic agents, and neuropeptide agonists/antagonists are now under investigations. PMID- 11268602 TI - [Mechanism and management of drug-induced eating disorder]. AB - Eating disorders induced by drugs, especially anorexia, nausea and vomiting are frequently reported as side effects of many available drugs. Since such disorders can affect patients' quality of life, suitable treatment plans based on the mechanisms of their induction are warranted. Eating disorders induced by drugs can be classified according to the mechanism of their induction into disorders resulting from: 1) gastrointestinal mucosal injury; nonsteroidal antiinflammatory drugs, antibiotics, etc. 2) chemical stimulation of the central nervous system; anticancer drugs, morphine, digitalis, etc. 3) non-specific symptoms following other systemic side effects; drug-induced depression, liver derangement, etc. This review addresses the aforementioned points and the best countermeasures that should be tried for successful control of these disorders. PMID- 11268603 TI - [Definition and classification of eating disorders]. AB - This paper describes definition and classification of Eating Disorders which centered on the atypical cases. Eating disorders in DSM-IV were further classified into 3 groups. Three groups were Anorexia Nervosa, Bulimia Nervosa and Eating Disorders Not Otherwise Specified. Binge Eating Disorders frequently transfer to obese patient. This disease entity become independent of Anorexia Nervosa and Bulimia Nervosa. Body weight changing trend evaluated not only cross section but also longitudinal observation. There are some experience cases which Anorexia Nervosa cause by diet therapy of obesity patient. A lot of Eating Disorder patients revealed atypical courses during clinical treatment. The symptom of disturbances in the way in which their body weight and sharp could not easy to confirm routine history taking. One type of eating disorders eat throughout the day with no planned mealtimes. PMID- 11268604 TI - [Anorexia nervosa (AN)--epidemiology, cause, therapy, outcome]. AB - The number of patients suffering from anorexia nervosa is rising steadily. More than 90 percent of patients are female in preadolescents or adolescents. AN is characterized by abnormal eating behavior and excessive loss of weight. The self esteem is highly dependent on their body shape and weight. The purposes of behavior therapy for AN are to remove various types of avoidance behavior, to reestablish desirable eating behavior and social skill behavior. The course and outcome of AN are variable. Of patients admitted to our hospital to receive behavior therapy, 60 percent are mostly recovered, 30 percent exhibit a fluctuating pattern of weight gain followed by relapse, and 10 percent are chronically deteriorating course of the illness over many years. The long-term mortality from AN is about 6 percent. PMID- 11268605 TI - [Bulimia nervosa (BN)--epidemiology, etiology, clinical features, treatment, prognosis]. AB - Bulimia nervosa is a distinctive disorder that was identified in the late 1970s. At first bulimia nervosa was closely associated with anorexia nervosa, but gradually the two disorders became partly separate. The characteristic of bulimia nervosa is the frequent occurrence of binge-eating episodes and sense of loss of control during eating episodes. Bulimia nervosa is complex disorder that are caused and then maintained by various social, psychological, and biological factors. The result of research over the 20 years have contributed significantly to cognitive-behavioural and pharmacological treatment approach for people with bulimia nervosa. PMID- 11268606 TI - [Eating disorders and immunodeficiency]. PMID- 11268607 TI - [Endocrine and reproductive disturbances in anorexia nervosa and bulimia nervosa]. AB - Numerous endocrine abnormalities are associated with anorexia nervosa and bulimia nervosa. The principal complication is amenorrhoea. Hypothyroidism and hypercortisolism have been described as a protective mechanism to conserve energy. Growth hormone concentrations are often increased as a result of starvation. Insulin and blood sugar concentrations are decreased, but prolactin concentrations are remain normal. Considerable evidence exists of hypothalamic dysfunction in patients with eating disorders. This dysfunction is reflected in disturbances of endocrine function. Endocrine disturbances may be not solely related to the low body weight. Hypothalamic monoamines, neuropeptides and leptin are involved in the regulation of human appetite, and in several ways they are changed in eating disorders. However, it remains to be clarified whether the altered appetite regulation is secondary or etiologic. PMID- 11268608 TI - [Bone mineral density of eating disorder]. AB - Osteoporosis recently has been added to growing list of medical complications assisted with eating disorder in particular Anorexia nervosa. These often occur early in the course of anorexia in adolescent girls, presumably because this disorder not only interrupts the normal rapid bone accretion characteristic of adolescences, but also accelerate bone loss. The pathogenesis of osteoporosis in AN has not been completely characterized. While low body mass and amenorrhea are clearly important variables, other focters also may be involved. Some possible contributing factors in patients with AN include low Ca intake, increased glucocorticoids, insulin growth factor 1 deficiency. Simple weight gain needs to be part of the treatment, although it and exercise remain of unproven benefit for osteoporosis in patients with AN. But some studies have found that the clinical course of osteoporosis is not reversed simply with weight restoration. Long term studies are needed to answer the question of whether osteoporosis assisted with eating disorder is reversible. PMID- 11268609 TI - [Pica: pathogenesis and therapeutic approach]. AB - Pica is one of eating disorders characterized by continuous or repeated ingestion of non-food materials or sometimes of a tremendous amount of specified food. Known risk factors for developing pica include iron deficiency anemia, malnutrition, mental disorders, pregnancy, psychiatric conflict, and social and ethnic habituation, although the mild pica tendency associated with mental disorders is excluded from the definition of pica according to the fourth edition of Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) by American Psychiatric Association. Pica is regarded as the common disorder than expected but commonly missed, and therefore is important to suspect of this disorder for its diagnosis when people with known risk factors show unexplained signs and symptoms. PMID- 11268610 TI - [Night-eating syndrome]. AB - Morning anorexia, evening hyperphagia and insomnia characterized night-eating syndrome. This syndrome is described in 1955 by Stunkard, et al. It occurred during periods of stress and was associated with a poor outcome of efforts at weight reduction. The prevalence of this syndrome was about 26% of severely obese population in US. In Japan, there is few clinical study of this syndrome. It is thought that this syndrome increases in prevalence with increasing adiposity. The behavior study showed that a coherent pattern of behavior was found in subjects with night-eating syndrome. And neuroendocrine study indicated that the leptin, which was produced from the adipocyts, related this syndrome and night eating behavior. PMID- 11268611 TI - [Human obesity and point mutations of leptin and leptin receptor]. AB - Obesity-related genes were isolated and identified in these several years. Development of transgenic mice has clarified the molecular mechanisms of obesity. There have been reported point mutations of the genes correspond to leptin(ob gene) and leptin receptor(db-gene). Thereby leptin is considered to be one of the most important regulators of energy metabolism also in human. PMID- 11268612 TI - [Evaluation of obesity and diagnostic criteria of obesity as a disease for Japanese]. AB - Obesity causes many undesirable health disorders such as diabetes mellitus, hyperlipidemia, hypertension and so on. Recently, those life style-affecting diseases is increasing, especially the increment of diabetes mellitus is prominent. In 2000, Japan obesity society issued the new standard of the evaluation of obesity and new diagnostic criteria of obesity as a disease for Japanese. According to this issue, obesity was evaluated by body mass index(BMI). And, 18.5 < BMI < 25 is normal, 25 < BMI < 30 is obese 1, 30 < BMI < 35 is obese 2, 35 < BMI < 40 is obese 3, and 40 < is obese 4. Obesity as a disease is defined by two cases. The first category is composed of two items; one is BMI > 25, and the other is having one disease worsen by obesity, such as diabetes mellitus, hyperlipidemia, hypertension, hyperuricemia, coronary heart disease, cerebral infarction, sleep apnea syndrome, fatty liver, deformative arthritis. The second category is the visceral type of obesity with BMI > 25, which was diagnosed by west size, over 85 cm for men, and over 90 cm for women, and by visceral fat area over 100 cm2 in abdominal CT. PMID- 11268613 TI - [Binge-eating in simple obesity]. AB - Binge eating in simple obesity has recently been recognized as a serious clinical problem. In the obese population with binge eating, distinct characteristics have been found by many researchers, such as an early onset of obesity and diet, frequent body weight fluctuation, the amount of time spent dieting, extreme restriction of food intake and an unrealistic ideal of diet. Obese binge eaters also exhibit more psychiatric symptomatology such as distortion of body image, low self-esteem, low self-efficacy, a high level of depression, strong perfectionism, high impulsivity and comorbidity of personality disorders, especially a borderline personality disorder. Cognitive-behavioral therapy and interpersonal psychotherapy are promising for treatment of obese binge eaters. Supportive psychotherapy will add some help by ameliorating psychological distress. PMID- 11268614 TI - [Classification and pathophysiology of symptomatic obesity]. AB - Symptomatic obesity was classified by 1) endocrine obesity, 2) central nerve related obesity, 3) genetic syndrome with obesity and 4) drug-induced obesity. Pathogenesis and pathophysiology of the diseases which induce symptomatic obesity, were briefly described. The origins of hyperphagia and obesity in these diseases have not yet clarified. Gene abnormalties of the genetic syndrome with obesity will be elucidated in near future by the rapidly progress of molecular biological techniques. PMID- 11268615 TI - [Specific features of obesity in children and its management]. AB - Obesity during childhood is caused by both congenital and acquired causes. Obese children usually have family history, especially of their mothers. Identical twins have similar weight even if they are reared apart. In very rare cases of heritable obesity, genetic defects in leptin synthesis and its receptor, POMC, MC4 receptor, and prohormone converting enzyme have been reported. In addition, body weight of children and adolescents is related with their life styles, and the prevalence of obesity in recent years is higher than before probably due to changes in calorie intake and energy expenditure. Diagnosis of obesity is based on the assessment of overweight using BMI in most cases of adults. During childhood and adolescence, BMI can not be applied as in adults and its percentile values are useful for children. Percentage of overweight for the standard weight for height has been used as well to demonstrate over- or underweight in children and adolescents. Evaluation of fat volume and its distribution is essential for the precise diagnosis of obesity in children as well as adults. PMID- 11268617 TI - [Efficacy of new devices for behavior modification therapy for obesity]. AB - In the treatment of obesity, the behavior modification therapy has aimed at maintaining weight reduction by reforming wrong daily habitual eating behavior which leads obesity. However, a distorted cognitive pattern or a deviated physical lies behind the problematic eating behavior. In that aspect, we have developed new devices as belows. The ingestive behavioral questionnaire discloses problematic eating behavior and its related cognitive and sensational pattern at an early stage of the treatment. The charting of daily weight pattern is convenient for both therapist and patient himself to visually look out an actual ingestive pattern and reinforces maintenance of weight reduction by long term self-monitoring. The chewing chart recording is useful for the recovery of physical satiety sensation, and prevents over eating. PMID- 11268616 TI - [Ideal way of diet therapy to improve the eating behavior pattern]. AB - As for obesity, the intake energy increases compared with the energy consumption and the synthesis to the body fat develops at a long term. How the eating behavior pattern took part in the disorder of adjustment function of such an energy balance have been examined. In addition, I have clarified how to improve the eating behavior of the purchase, the cooking, the preservation of the food, and the speed, the frequency, and distribution and the sense at the meal for prevention and treatment of obesity. PMID- 11268618 TI - [Indications for surgery for morbid obesity and effectiveness of bariatric surgery]. AB - In this paper we describe a history and technical aspects of bariatric surgery. And surgical techniques of Divided Vertical Banded Gastroplasty and perioperative management are presented. Our indications for surgery for morbid obesity are almost equal to the guidelines for obesity surgery adopted by the American Society for Bariatric Surgery October 1986. From 1982 to 2000, 64 bariatric surgical procedures were performed at the author's institution. Vertical banded gastroplasty was performed in 45 patients, Horizontal gastric partitioning in 8 patients, Gastric bypass in 10 patients, and Divided vertical banded gastroplasty in 1 patient. The average weight loss one year after Vertical banded gastroplasty is 1/3 of the patient weight. Most of the preexisting comorbid conditions related to the obesity showed improvement or were completely resolved after surgery. No major complications were observed postoperatively. We concluded that surgical procedures for morbid obesity are very effective therapy. PMID- 11268619 TI - [Anti-obesity drugs--mechanism and medical indication of appetite suppressants]. AB - Obesity has become a big problem in Japan as in Europe and United States. Principle of the treatment of obesity is diet and exercise therapy, but these treatments are often failed. Pharmacotherapy is now second choice, but is expected great developments by recent advances of molecular biology of obesity. Appetite suppressants were previously applied for clinical use and showed a certain effect to decrease body weight. In Japan, only madindol is available and sibutramine is one the clinical trial. In the medical treatment of obesity including pharmacotherapy, we have to select the obese patients who should be under the medical control, and we should not only reduce the body weight, but medicate the complications of obesity. PMID- 11268620 TI - [Study of factors that influence the use of ambulances in Japanese prefectures]. AB - PURPOSE: The objective of this study was to clarify the factors that influence the rate of patient-carriage by ambulances in Japanese Prefectures. METHOD: The study was conducted using data on cases of patients with mild conditions carried by ambulances in Japanese prefectures in 1993, concentrating the factors that influence the rate of usage. The cases were analyzed focusing on three major types of medical emergencies (accounting for 93.4% of the total): sudden illnesses, traffic accidents and general injuries. SPSS for Windows was used for statistical analysis. RESULTS AND DISCUSSION: Data analysis by age group and type of medical emergency showed a positive correlation (r > 0.7) for the rate of cases with mild conditions carried by ambulances in all age groups (early childhood, adolescence, adult and senior) and the three major types of medical emergencies. Particularly, a strong correlation (r > 0.9) was observed with regard to patients suffering from sudden illnesses and general injuries. Multiple regression analysis showed that the rate for patients with mild cases carried by ambulances was higher in prefectures where; (1) there were more cases of administrative litigation related to individuals' rights, and (2) there were many nuclear families that tended to lack the capacity for family care. In addition, it also became evident that the rate for patients with mild injuries from traffic accidents was higher in prefectures where; (1) the rate for male-driver license holders for small- and medium-size cars was higher, and (2) the ratio of numbers of vehicles to total roadway area was higher. CONCLUSIONS: The findings suggest that factors unrelated to medical emergencies have a major influence on use of ambulance. Reevaluation of policy in order to promote appropriate utilization is necessary. PMID- 11268621 TI - [Inference of the means and the confidential intervals for epidemiological indices in enterohemorrhagic]. PMID- 11268622 TI - [Knowledge, behavior, and attitudes toward sex among adolescent students at a junior/high school in Cote d'Ivoire]. AB - OBJECTIVES: To be able to provide appropriate information about sex to adolescent students in the Cote D'Ivoire, we conducted a study to determine knowledge, behavior, and attitudes toward sex. METHODS: We conducted a cross-sectional study at a junior/high school using a self-administered questionnaire which contained students' characteristics (age, sex, grade, tribe, region), their knowledge about sex, sexual behavior (including experience of sex and contraception), and attitudes toward sex. Knowledge about sex and proportions employing contraception were compared between males and females and between the lower and the upper grades. We also examined associations between attitudes toward sex and contraception. RESULTS: A total of 695 (males: 278, females: 417) students filled in our questionnaire (response rate: 33.1%). The proportion of the students who had experienced sex was 84.2% for males, 46.5% for females, and the average age of the first sex was 14.3 and 15.6 years in males and females, respectively the proportions using contraception was 49.6% and 46.9%. In students of the lower grades, males had greater knowledge than females, but this difference was reduced with progression through the upper grades. The proportion using contraception in the upper grades was also higher and an association between attitude toward sex and contraception was clear. CONCLUSION: Adolescent students' knowledge, behavior, and attitudes toward sex could be relatively easily investigated at a junior/high school of Cote D'Ivoire. The majority of males and half of the females in this setting already had experience of sexual intercourse. The study showed that difference in sexual knowledge between males and females shrunk with increase in school grade. We also showed an association between attitude toward sex and contraception. PMID- 11268623 TI - [Several problems on infectious waters. The responsibility of medical institutions and the information and evaluation about contractors]. PMID- 11268624 TI - [Examination of validity of the subjective fatigue scale for young adults]. AB - OBJECTIVE: The purpose of this study was to examine the validity of a new type of Subjective Fatigue Scale for young adults (SFS-Y). METHODS: The SFS-Y consisted of 24 item questions representing 6 sub-scales of difficulty of concentrated thinking, languor, reduced activation, reduced motivation, drowsiness and feeling of physical disintegration. The SFS-Y, subjective symptoms index (SSI), chronic fatigue symptoms index (CFSI), multidimensional fatigue index (MFI) and the Chalder fatigue scale (CFS) were administered to 5435 students aged 15-20 yr. RESULTS: It was inferred that SFS-Y can evaluate individual differences from the viewpoints of score distribution and discrimination power. It was considered that the SFS-Y has high generality because it includes many question items with a similar content to existing fatigue scales and it can do multiple evaluations covering almost all aspects of existing scales. The relationships between sub scales in the SFS-Y and existing fatigue scales were high in sub-scales with similar names and was low in ones with different names. CONCLUSION: The Subjective Fatigue Scale developed to evaluate subjective fatigue for youths was considered to be excellent in validity and to be effective scale. PMID- 11268625 TI - [Maternal partiality in attachment with multiple birth children and the related factors]. AB - Multiple births are associated with an increased risk of child abuse and neglect. It is reported that only one child is abused in almost cases, and most abusers are the mothers. Maternal partiality regarding attachment has been suggested as the reason for this tendency. This study investigated the prevalence of this phenomenon in families with multiple birth children and identified factors associated with increased risk. The subjects were 231 mothers of multiple birth children. The following results were obtained. 1. Overall, 10.0% of mothers with multiple birth children reported that they didn't equally attach themselves to all their offspring. 2. Mothers who didn't equally attach themselves exhibited significantly poor health conditions and a higher frequency of upper respiratory infections, compared with mothers who demonstrated no partiality. Moreover, they were more likely to complain of severe fatigue (physical and mental) and poor sleeping conditions. 3. The mothers who didn't equally attach themselves to all their multiple birth children had a higher rate of handicapped children. CONCLUSION: Mothers who do not equally attach themselves to all their multiple birth children show poor health conditions and a higher frequency of upper respiratory infections, and complain of severe fatigue and poor sleeping conditions. They also have a higher rate of handicapped children. PMID- 11268627 TI - [Comprehensive view of bacteriocins and evolution of colicin structures]. PMID- 11268626 TI - [Prediction of mortality from findings of annual health checkups utility for health care programs]. AB - OBJECT: To clarify relationships between the findings of annual health checkups and mortality in men and women living in Ibaraki prefecture. METHOD: The subjects were 32,705 men and 63,959 women aged 40 to 79 years who participated in annual health checkups in 1993. They were followed up until November 30, 1998, with a systemic review of resident registration and death certificates. The Cox's proportional hazards model was used to estimate relative risk, after adjustment for age, smoking status, usual alcohol intake, hypertension category, serum total cholesterol, HDL cholesterol, blood glucose, serum creatinine, body mass index (BMI) and urinary protein. RESULTS: During the 5.2-year follow-up, there were 2,937 deaths (including 384 deaths from stroke, 242 from coronary heart disease and 1,305 from cancer). Significant predictors of mortality from all causes were smoking, usual alcohol intake, hypertension, low serum total cholesterol, low BMI, high blood glucose level, proteinuria for men and women, and low HDL cholesterol for men, and high serum creatinine for women. Significant predictors of mortality from all cardiovascular diseases were smoking, hypertension, low BMI, high serum creatinine, proteinuria for men and women, usual alcohol intake and low HDL cholesterol for men, and serum total cholesterol and high blood glucose level for women. Significant predictors of mortality from stroke were hypertension, low BMI, high serum creatinine for men and women, and proteinuria for women. Significant predictors of mortality from coronary heart disease were smoking, high serum total cholesterol, high blood glucose level, proteinuria for men and women, hypertension, low HDL cholesterol for men. Significant predictors of mortality from cancer were smoking, usual alcohol intake, BMI for men and women, low serum total cholesterol, low HDL cholesterol and proteinuria for men, and high blood glucose level for women. Smoking, usual alcohol intake, low HDL cholesterol and proteinuria were significant predictors of mortality from lung cancer for men. CONCLUSION: Smoking, usual alcohol intake, hypertension, BMI, serum level of total cholesterol, HDL cholesterol, blood glucose, creatinine, and urinary protein are significantly associated with mortality. We obtained the new finding that serum creatinine level is a significant predictor of mortality from all cardiovascular diseases in Japanese men and women, and that the multivariate relative risk in female moderate alcohol drinkers (46-68 g ethanol intake/day) vs non-drinkers is significantly elevated for death from all causes. The results of our study are useful for planning of health care education and services. PMID- 11268628 TI - [Group behaviour of gram-positive bacteria controlled by peptide pheromone]. PMID- 11268629 TI - [Structure and mode of action of bacteriocins produced by lactic acid bacteria]. PMID- 11268631 TI - [Diversity and uniqueness of pyocins produced by Pseudomonas aeruginosa]. PMID- 11268630 TI - [A cytotoxic ribonuclease targeting specific tRNAs]. PMID- 11268632 TI - [Yeast killer toxins: a study toward structure and function]. PMID- 11268633 TI - [Molecular mechanism of action of ribotoxins]. PMID- 11268634 TI - [Natural insecticides]. PMID- 11268635 TI - [Antimicrobial proteins of insects: a speculation of their origin and functions]. PMID- 11268636 TI - [Antitumor and antimicrobial proteins from marine mollusca Aplysiids]. PMID- 11268637 TI - [Horseshoe crab animicribial proteins with chitin-binding activity]. PMID- 11268638 TI - [Pierisin, an apoptosis-inducing protein from cabbage butterfly]. PMID- 11268639 TI - [Internal defense factors in terrestrial slug]. PMID- 11268640 TI - [Host defensive protein of planaria]. PMID- 11268641 TI - [Neurotoxins in arthropod venom]. PMID- 11268642 TI - [Mastoparan as a G protein activator]. PMID- 11268643 TI - [Spider and scorpion toxins as the ion channel blocker]. PMID- 11268644 TI - [Bioactive molecule supporting blood feeding: unique activities from the salivary glands in the kissing bug, Rhodnius prolixus]. PMID- 11268646 TI - [Biological activity of lysenin]. PMID- 11268645 TI - [Chemical structures and the mechanism of action of peptide toxins from cone shells]. PMID- 11268647 TI - [Dimerization of snake venom protein generates a new function]. PMID- 11268648 TI - [Relationship between structure and function of cholera toxin and Escherichia coli enterotoxin: structural and functional similarities to other toxins]. PMID- 11268649 TI - [Shiga toxins produced by enterohemorrhagic Escherichia coli]. PMID- 11268650 TI - [Tetanus and botulinum toxins]. PMID- 11268651 TI - [Bordetella dermonecrotizing toxin and Escherichia coli cytotoxic necrotizing factors: bacterial toxins activating Rho family GTPases]. PMID- 11268652 TI - [Structure and mode of action of staphylococcal bi-component pore-forming toxins]. PMID- 11268653 TI - [Diphtheria toxin receptor]. PMID- 11268654 TI - [Translocation of virulence factors into the host cell by bacterial delivery systems]. PMID- 11268655 TI - [Helicobacter pylori vacuolating cytotoxin (VacA) and its modulatory effects in host cells]. PMID- 11268656 TI - [Impairment of host defense via activation of host's proteases by bacterial proteases]. PMID- 11268657 TI - [Molecular mechanism of membrane pore formation with cholesterol binding cytolysin: streptolysin O and perfringolysin O]. PMID- 11268658 TI - [Recombinant immunotoxin for cancer therapy: current status and prospective]. PMID- 11268659 TI - [The gastrointestinal immune system: cholera toxin and E. coli heat-labile enterotoxin as immunomodulators]. PMID- 11268660 TI - [Interaction between membrane-damaging toxin of Listeria monocytogenes and host immune system]. PMID- 11268661 TI - [Toll-like receptors and innate immune system]. PMID- 11268662 TI - [Structures of bacterial superantigen on the molecular basis and their activities to the immune system]. PMID- 11268663 TI - [Anti microbial, LPS-neutralizing protein (CAP 18) and host defense]. PMID- 11268664 TI - [Early events in the development of the nervous system]. AB - In the recent years, real advances have been made in our understanding of some aspects of early neural development. A remarkable finding is the identification of neural inducers, the existence of which had been predicted by Mangold and Spemann almost seventy years ago. Another recent and important discovery consists in the identification of several developmental genes and growth factors. Indeed these two classes of molecules participate in the induction and in the physiological and morphological compartmentalization of the nervous system along its anteroposterior and dorsoventral axes. PMID- 11268666 TI - [Gene transfer]. AB - Mammalian cells in culture can be modified by gene transfer and these procedures are routinely used in experimental biology. Yet, efficient approaches to modify certain complex cell populations, stem cells or cells organized within a tissue are still lacking. The hurdles to gene transfer can be listed by describing the pathway of a nucleic acid molecule, from the external medium towards the cell nucleus where its encoded information will be expressed. The requirements include the necessity to compact the size of the macromolecule, to overcome electrostatic repulsion, to cross a series of membranes and to establish itself permanently. Viruses have evolved to achieve these goals and understanding their strategies for cell invasion allows to design vectors for gene transfer. Chemicals or biochemicals able to bind DNA, as well as physical methods inducing changes in the structure of membranes can also be useful for gene transfer. The outcome of gene transfer technologies and their improvement pave the way to inovative medical applications and provide powerful tools for exploring the living world. PMID- 11268665 TI - [Nuclear transfer and cell differentiation: contribution of cloning to the study of embryonal stem cells]. AB - The birth of physiologically normal and fertile animal of several mammalian species following the transfer of somatic nuclei into recipient enucleated oocytes has stimulated researches on the functional plasticity of cells available from living organisms. This experimental approach, termed cloning, has up to a low efficiency. It however allows the study of mechanisms compatible with the reacquisition of an undifferentiated nuclear status and contributes to researches aimed at controlling the fate of stem cells for therapeutically applications. PMID- 11268668 TI - [Medical use, current and future, of human stem cells]. AB - Since many years, the ability to harvest and then graft haematopoietic stem cells in man has allowed to treat with success several lethal diseases in haematology and oncology. Progress in cell biology recently made possible the obtention of human stem cells, particularly of embryonic origin which, theoretically, have all the potentialities of development. Therefore, in spite of other many obstacles still ahead, the perspective of using elements produced from such cells for therapeutic purpose seems closer. PMID- 11268667 TI - [Neurogenesis in the adult brain: hope for brain repair?]. AB - It has been known for many years now that the vocal centers in the hyperstriatum (cortex) of birds with seasonal song behaviour are the site of neuronal generation and, thus, that stem cells capable of giving rise to real neurons exist in the adult. Since then, the notion that neuronal progenitors exist in the adult has been extended to mammals. It seems that the number of brain regions in which neuronal generation is taking place is limited. In addition to the olfactory epithelium where olfactory receptors are under constant renewal, the sites of neuronal regeneration are the olfactory bulb, the dentate gyrus of the hippocampus, the associative cortex--in the macaque--and the cerebellum. Although this represents only a limited number of regions and neuronal types (primarily GABAergic interneurons), the observation that new neurons can be created throughout life raises the possibility to use them in replacement therapy in the human. PMID- 11268669 TI - [Hematopoietic stem cells]. AB - Hematopoietic stem cells, which share with other stem cells of adult tissues the ability to maintain constant the number and diversity of differentiated mature cells throughout adult life offer a fabulous system to analyze mechanisms controlling cell proliferation and differentiation. Cytokines controlling the differentiation of intermediate progenitors into mature cells of the various lineages have been characterized and have been widely used, in vitro as in vivo, to increase the output of differentiated cells. In contrast, despite significant technological advances, molecular events associated with the stem cell decisions first to either self-renew or differentiate, and then to irreversibly commit to one of the lymphoid or of the myeloid pathways are still very badly understood. This is partly explained by the lack of reliable assays, particularly in humans, to assess stem cell activity, and by the difficulty to dissect the composition of molecular complexes regulating gene expression in these very rare cells. Despite these limitations, recent evidence suggests that there is some flexibility in the initial decisions of stem cells, and that extracellular factors may influence stem cell fate. If this is confirmed, it may then become possible to propose new therapeutic strategies based on the manipulation of stem cell properties. PMID- 11268670 TI - [Therapeutic potential of fetal neuron grafts in neurodegenerative diseases]. AB - Neurodegenerative diseases are a major public health problem since they affect as a mean 1% of the people in industrialized countries, and will progress along with the ageing of the population. For ten years, a major research endeavor aims at developing new therapeutics against these diseases. These new approaches are based either on the substitution of affected neurons by homogous neuroblasts that are able to replace them anatomically and functionally (fetal neural grafts), or on the use of neuroprotective proteins (by gene therapy) that are able to promote natural defenses of neural cells. Clinical results have been obtained with fetal neural transplants over the past decade, in patients with Parkinson's and, more recently, Huntington's disease. The neuroprotective approach is under evaluation. PMID- 11268671 TI - [Ethical aspects of human cloning for therapeutic purposes and the use of embryonal stem cells]. AB - Embryo definition is without any ambiguity: it consists in a stage of development able to give rise to an autonomous organism. In this sense, it is obvious that human embryos could be produced by cloning. If the therapeutic prospects of therapeutic cloning are confirmed, there will be a tension between two ethical logics: to respect human embryos as possible persons ... and to improve the condition of severely affected patients. Whatever the definitive solution, its moral significance should be denied. In contrast, it seems possible to use spare embryons for selected research without considering them only as things. PMID- 11268672 TI - [The ethical question of the embryo]. AB - The possible use of embryonic stem cells for therapeutic purposes raises once more the ethical question concerning the position of the embryo in relation to medical needs. Since this technology treats the embryo as a raw material, the debate must incorporate a semantic clarification, in order to identify the conceptual ambiguities that exist between popular thought, science, anthropology and the religious and philosophical convictions of the individual. The problem is knowing whether the end always justifies the means and whether the future may be sacrificed to the present. PMID- 11268673 TI - [Pollicization, remains of the past or current operation]. AB - Pollicization is a long time honored operation but more recent introduction of microsurgical techniques allowing toe to thumb transfers has changed the indications in trauma and congenital malformations. We reviewed two series of patients to assess the long-term result in both traumatized and malformed hands. Twenty-seven pollicized mutilated fingers were reviewed with a mean follow up of 9.5 years; the longest follow up published in the literature. Despite the age quite advanced of the population and multidigital amputations (more than two fingers were amputated in 16 cases), 21 patients had a good or fair result on a global score including mobility, sensibility, daily use, cortical integration, cold intolerance or pain and appearance. In congenital malformations, hypoplasia or aplasia of the thumb in presence of long fingers remains one of the best indication of pollicization. Out of 35 operated children, 27 were reviewed with a sufficient follow up (mean 6 years) to assess the result. Sensibility and growth were excellent (3 cases need some secondary shortening) mobility was close to normal in 61% of cases but the main concern was the strength which reached only 42% of the standard. PMID- 11268674 TI - [Significance of paramyxovirinae protein F in physiopathology and immunity]. AB - The Paramyxovirinae sub-family contains viruses of major importance for humans (measles and mumps viruses) and animals (Newcastle disease virus, canine distemper virus, rinderpest virus) but also zoonotic viruses (Hendra and Nipah viruses). Newcastle disease virus, a Rubulavirus, which is specific of birds, can be used as a model for understanding the pathogenicity mechanisms of Paramyxovirinae and for the study of vaccination against the diseases they cause. The F protein of pathogenic strains of Newcastle disease virus have a polybasic cleavage site (K/R-Q-K/R-R) which is specific of furine, a cellular enzyme present in all cellular types, what allows viral multiplication in all cells of the host. Such cleavage site is also present in the F protein of measles and mumps viruses. Antibodies against the F protein are the most effective in the immune response against Newcastle disease virus. The major role played by the F protein in the immunity against Paramyxovirinae arises also from the research performed on other animal (rinderpest and canine distemper viruses) and human (measles virus) viruses. Recombinant and sub-unit vaccines as well as DNA vaccination based on the expression of viral glycoproteins allow effective vaccination of chickens against Newcastle disease. The use of a similar approach could lead to the development of new vaccines against measles or mumps and if necessary to produce vaccines against zoonotic Paramyxovirinae. PMID- 11268675 TI - [Should acute kidney failure be treated in people over 80 years of age at an intensive care unit?]. AB - This is the first retrospective study aimed to analyze the clinical symptoms, the etiology, the morbidity and the lethality of acute renal failure in patients over 80 years. The criterion of inclusion was the occurrence of acute renal failure defined on the basis of high plasma creatinine associated with normal kidney size in patients of this class of age who had been hospitalized in an intensive care unit between October 1971 and September 1996. In the case of a preexisting moderate nephropathy, acute renal failure was defined by an increase of plasma creatinine of at least 50% over its basal value. Three hundred and eighty-one patients over 80 years out of a total of 2,111 patients with acute renal failure were included in this study. The various etiologies and mechanisms of the disease are described. Twenty-nine% of the patients underwent dialysis. The global lethality reached 40% during the time of hospitalization. The factors significantly associated with a poor prognosis were identified as cancer essentially, but also sepsis and preexisting cardiovascular diseases. The mean survival after hospitalization was of 19 months. In summary, the frequency of admission for acute renal failure of patients over eighty in intensive care units is increasing but the rate of lethality observed is less than expected. A pattern of pathological associations leading to death in these patients cannot be defined with certitude. Therefore, the methods of renal replacement therapy available in modern intensive care units must be utilized in this class of age as it is the case in younger patients. PMID- 11268676 TI - [Mucoviscidosis: progress in the diagnosis and management]. AB - Cystic fibrosis (CF) is the most common severe autosomal recessive disorder amongst Caucasian populations. We have learned a great deal on CF during the past 20 years, mainly with the identification of the CFTR gene in 1989. In the same time, improvements in the therapeutic management dramatically changed its clinical outcome: while in 1946 the median survival was 4-5 years, in 2,000, it reaches 30 years in reference centers. Since the prognosis depends on respiratory functions, a large number of clinical trials were designed to improve them; medical, physiotherapic, and surgical interventions such as a bipulmonary grafts allowed to slow the natural decline of respiratory functions. Early diagnosis and maintaining optimal nutrition such as bipulmonary grafts allowed to slow the natural decline of respiratory functions. Early diagnosis and maintaining optimal nutrition are two other determining factors of prognosis. Nowadays the next step is to resolve the problem of the benefit of gene therapy but no significant progress was observed in that field for the past five years. PMID- 11268677 TI - [HIV infections, AIDS, and STDs in a setting under epidemiologic surveillance: course among army personnel]. PMID- 11268678 TI - Arthropods as predators of ticks (Ixodoidea). AB - The existing information on arthropods as predators of ticks is based mainly on sporadic observations and their role in reducing tick populations and in most cases is still not clear. Some reports suggest that in certain ecological habitats arthropods play an important role in the control of the tick population. This publication reports on some 100 relevant publications that appeared between 1906 and 1999. Ants, beetles, and spiders seem to be the major arthropods preying on ticks. In general, engorged ticks are more often preyed upon by arthropods than are unfed or feeding ticks. PMID- 11268679 TI - Temporal changes in prevalence of scrub typhus rickettsia (Orientia tsutsugamushi) infecting the eggs of Leptotrombidium imphalum (Acari: Trombiculidae). AB - Eggs from seven colony lines of the chigger mite Leptotrombidium imphalum (Vercammen-Grandjean & Langston) were examined for infection with Orientia tsutsugamushi (Hyashi), the etiologic agent of scrub typhus. The polymerase chain reaction (PCR) using primers OtP 56.809 and OtM 56.1221, which amplify a 291 bp region of the P56 gene of O. tsutsugamushi, was used to detect scrub typhus within single eggs. All seven chigger mite lines produced infected eggs with varying rates of infection (Li1 = 8.1%, n = 124; Li2 = 45.6%, n = 90; Li3 = 30.1%, n = 144; Li4 = 31.7%, n = 145; Li5 = 21.3%, n = 136; Li6 = 41.6% n = 77; Li7 = 22.5%, n = 110). The 3 wk with the highest infection rates for each line using Fourier analysis were as follows: Li1 = 2, 7, 14; Li2 = 4, 6, 12; Li3 = 3, 6, 12; Li4 = 4, 6, 12; Li5 = 5, 7, 14; Li7 = 4, 6, 12. Li6 only had nine measurements over time; therefore, Li6 was excluded from individual analysis. Infection rates of scrub typhus in eggs occurred in a 2-wk 2-d cycle, using Fourier analysis of combined data. Not only did infection rates vary among the progeny of females, but temporal variation also occurred. PMID- 11268680 TI - Seasonal occurrence of Ctenocephalides felis felis and Ctenocephalides canis (Siphonaptera: Pulicidae) infesting dogs and cats in an urban area in Cuernavaca, Mexico. AB - The seasonal occurrence of Ctenocephalides felis felis (Bouche and Ctenocephalides canis (Curtis) infestation on dogs and cats in Cuernavaca City in Mexico, was determined by examining 1,803 dogs and 517 cats at two veterinary clinics during 1995-1997. The overall flea infestation was 30.3 and 30.1% for dogs and cats, respectively. There were no significant differences (P > 0.05) in percentage of infestation among years for both hosts. The infestation was somewhat higher in spring, summer, and autumn than in winter, but no statistical differences was found among seasons (P > 0.05) for both pets. No relationship existed between percentage of flea infestation and temperature or rainfall among seasons. On dogs, 81.1% were infested with only C. felis felis, 16.8% with C. canis, and 2% had both flea species; whereas 92.3% of the cats were infested with C. felis felis and 7.7% with C. felis felis and C. canis. The cat flea was the most prevalent flea species found other than C. canis; no other species were found on the dogs and cats. It appeared that flea life cycle development continued throughout the year. PMID- 11268681 TI - Interdigital gland substances of white-tailed deer and the response of host seeking ticks (Acari: Ixodidae). AB - Host-seeking male and female blacklegged ticks, Ixodes scapularis Say, exhibited an arrestant response when contacting substances from front and rear interdigital glands of male and female white-tailed deer, Odocoileus virginianus (Zimmermann). Female I. scapularis responded positively to substances from interdigital glands on the fore legs and hind legs of female deer, whereas male I. scapularis responded only to samples from the fore legs of does. These results showed that previously reported responses of I. scapularis to residues from interdigital glands of hind legs of deer were independent of tarsal gland substances, which may contaminate interdigital gland substances on hind legs. Nymphs of I. scapularis and female lone star ticks, Amblyomma americanum (L.), did not show an arrestant response to substances from hind legs of does, but male A. americanum did. PMID- 11268682 TI - Preliminary survey of ticks (Acari: Ixodidae) parasitizing wild turkeys (Aves: Phasianidae) in eastern Kansas. AB - During the spring and fall turkey hunting seasons of 1999, hunters and Kansas Department of Wildlife and Parks field personnel examined wild turkeys, Meleagris gallopavo L., for ticks and submitted them to us for identification. From springtime hunting, we received 113 ticks from 12 turkeys killed in nine counties, all in the eastern one-third of Kansas. Collectors reported examining three additional wild turkeys on which no ticks were found. All ticks were nymphal lone star ticks, Amblyomma americanum (L.). Of 11 wild turkeys examined in seven counties during October, one was parasitized by 30 A. americanum larvae. Data from this study and accounts from the published literature suggest that parasitism of wild turkeys by immature lone star ticks is commonplace wherever this host and ectoparasite are sympatric. Our study suggests that M. gallopavo may be an important host that supports lone star tick populations. PMID- 11268683 TI - Gene flow between natural and domestic populations of Lutzomyia longipalpis (Diptera: Psychodidae) in a restricted focus of American visceral leishmaniasis in Venezuela. AB - The epidemiology of the visceral leishmaniasis in the Americas is associated with both a natural and a domestic cycle. The existence of reproductively isolated populations of Lutzomyia longipalpis (Lutz & Neiva), and the scarcity of records of this species from natural habitats in areas where it has been associated with domestic habitats indicated that natural populations could be genetically distinct from domestic ones. Therefore, we compared the genetic structure and estimated the gene flow between L. longipalpis from domestic and peridomestic habitat and from an adjacent undisturbed natural environment along a 1.2-km transect. The analyses were performed on electrophoretic data from eight isozyme loci. The absence of fixed differences in the diagnostic loci Ak and Hk indicated that all specimens belonged to one of the two cryptic species identified in Venezuela. The average number of alleles per locus ranged from 2.0 to 2.9 and the average heterozygosity ranged from 7.8 to 13.4%. No differences were detected in the genetic structure of this species from domestic or peridomestic habitats and those trapped as far as 1.2 km from human dwellings. Nm, estimated from Wright's Fst, indicated that at least 208 individuals per generation migrated between the peridomestic habitat and a 1.2-km distant point to maintain the observed similarities in allelic frequencies. This high rate of gene flow indicated that this species has high migration rates between domestic and natural environments, and has the potential to transport for Leishmania from natural to domestic environments. PMID- 11268684 TI - Dogs as a favored host choice of Anopheles gambiae sensu stricto (Diptera: Culicidae) of Sao Tome West Africa. AB - The host source and human blood index (HBI) of an exophilic population of the "forest" cytoform of Anopheles gambiae Giles sensu stricto, from a peri-urban area of the island of Sao Tome were assessed. Blood meals of 434 An. gambiae females from all-night indoor light-trap collections, 193 from indoor and 422 from outdoor resting collections, were determined by ELISA. Significant differences were found in the HBI estimates from insects collected indoors (0.93) and outdoors (0.27). Blood-fed insects collected resting outdoors provided the most representative sample for host determination. Dogs were the predominant hosts, followed by humans and pigs. Of all human feeds, it was estimated that 81.5% were taken inside houses. The low HBI of 0.27 for the An. gambiae population explains the low sporozoite rate and the meso-endemicity of malaria in the island. PMID- 11268685 TI - Vertical transmission of Orientia tsutsugamushi in two lines of naturally infected Leptotrombidium deliense (Acari: Trombiculidae). AB - Vertical transmission of Orientia tsutsugamushi (Hayashi), the etiologic agent for scrub typhus, was studied in two lines of naturally infected Leptotrombidium deliense Walch. In one line of mites originating from a single adult (V3M), the rate of filial transmission was 100% for the first two laboratory generations, but declined to 86.6% in the third laboratory generation. The vertical infection rate in this line of mites was 100% for the parental generation, but declined to 95.6% for the F1 generation and 88.6% for F2. The transmission of O. tsutsugamushi in another line of L. deliense (V3F) was less efficient than mites originating from V3M. In the initial laboratory generation of V3F a filial transmission rate of 100% was recorded. However, none of the F2 generation of this line transmitted rickettsiae to mice (Mus musculus L.), resulting in a filial transmission rate of 0%. Transmission of O. tsutsugamushi to mice by progeny from cohort larvae originally from the same adult (V3F) was also studied in the laboratory and these were found to be relatively poor transmitters of rickettsiae. The filial infection rate of F2 larvae was 60%, F3 was 88.8%, and F4 was 55.9%. The biology of infected L. deliense was studied and compared with uninfected mites reared under the same laboratory conditions. The results showed that infected female L. deliense laid approximately the same or more eggs as uninfected adults. The rate of development of the progeny of infected L. deliense was not significantly different from uninfected mites. PMID- 11268686 TI - Effects of available sugar on the reproductive fitness and vectorial capacity of the malaria vector Anopheles gambiae (Diptera: Culicidae). AB - Although females of most mosquito species are known to use sugar as a necessary source of energy, female Anopheles gambiae Giles sensu stricto are thought to use it facultatively or not at all. However, field evidence of sugar-free living is inconclusive, and the implications for reproductive fitness and vectorial capacity are unknown. To evaluate the role that sugar may play in the ecology of these mosquitoes, mated female An. gambiae in the laboratory were given access to either no food (water only), 10% sucrose, human blood, or human blood + 10% sucrose, and comparisons of daily mortality, fecundity, and biting frequency were made. The effect of sugar availability on vectorial capacity and the intrinsic rate of increase, a measure of fitness, then were determined. Females (pooled and individual) given blood + sugar lived significantly longer than did those on the other diets. Daily fecundity was higher for females given blood alone than for those fed blood + sugar (13 versus 9 eggs per female daily). However, total fecundity and intrinsic rate of increase were not affected by sugar availability. Biting frequency was significantly higher (0.41 versus 0.26 bites per female per day) for females given blood alone. Despite the reduced survivorship, exclusive blood-feeding led to a theoretically higher vectorial capacity for Plasmodium falciparum at 27 degrees C. These data indicate that female An. gambiae could replace sugar with increased blood feeding without suppressing reproductive fitness. Increased blood feeding could, in turn, increase the rate of malaria transmission and may explain the unusual efficiency of this vector. PMID- 11268687 TI - Meconial peritrophic membranes and the fate of midgut bacteria during mosquito (Diptera: Culicidae) metamorphosis. AB - The location of midgut bacteria relative to meconial peritrophic membranes (MPMs) and changes in bacterial numbers during midgut metamorphosis were studied in Anopheles punctipennis (Say), Culex pipiens (L.), and Aedes aegypti (L.) pupae and newly emerged adults. After adult emergence in Aedes, Anopheles, and most Culex, there were few to no bacteria in the midgut. In most newly emerged adult mosquitoes, few bacteria were found in either the lumen or within the MPMs/meconia. In a few Culex specimens, high numbers of bacteria were found in the MPMs/meconia and low numbers in the lumen. In all three species bacterial counts were high in fourth instars, decreased after final larval defecation, increased in young pupae, and increased further in old pupae. A very effective gut sterilization mechanism is operating during mosquito metamorphosis and adult emergence. This mechanism appears to involve the sequestration of remaining larval gut bacteria within the confines of the meconium and one or two MPMs and the possible bactericidal effect of the exuvial (molting) fluid, which is ingested during the process of adult emergence. PMID- 11268688 TI - Ixodes scapularis (Acari: Ixodidae): abundance and rate of infection with Borrelia burgdorferi in four state parks in Wisconsin. AB - Four state parks located in Lyme disease endemic regions of Wisconsin were surveyed for the presence of Ixodes scapularis Say during May and June of 1998 by drag sampling along hiking trails. Nymphal abundance varied between parks, with the average number of nymphs encountered in 1 h ranging from 6.2 +/- 3.8-47.1 +/- 36.3 (mean +/- SD). Questing nymphs were tested for the presence of Borrelia burgdorferi by culture in BSK medium and 7-12% was found to be infected. The average risk of encountering an infected nymph (entomologic risk index) ranged from 0.5 to 5.2 infected nymphs per hour. The highest entomological risk index was recorded from a small island park in northwestern Wisconsin during the last week in May (8.0 infected nymphs per hour). These results indicate a lower risk for human Lyme disease exposure in Wisconsin state parks in comparison with highly endemic areas of the northeastern United States. PMID- 11268689 TI - Green fluorescent protein-tagged Leishmania in phlebotomine sand flies. AB - In this work we have used for the first time green fluorescent protein (GFP) tagged cells of the human parasite Leishmania donovani to observe its development in the gut of phlebotomine sand flies. Low numbers of GFP-tagged L. donovani were more easily detected than nontagged Leishmania, suggesting that GFP-tagged Leishmania could be used to efficiently study the biology of Leishmania in their vectors, and open the possibility of using nonaxenic flies. Using this method, we found that GFP-tagged L. donovani, the ethiological agent of Old World Kala-azar, were able to establish an infection within the gut of Lutzomyia species, which are vectors of New World Leishmania. The GFP-tagged parasites divide successfully in the gut of colonized and in wild caught Lu. longipalpis (Lutz & Neiva, 1912), Lu. ovallesis (Ortiz, 1952), and Lu. youngi (Feliciangeli & Murillo, 1985). In the case of Lulongipalpis the labeled parasite exhibited a normal anterior development as the one observed in its natural vector. PMID- 11268690 TI - Use of an allele-specific polymerase chain reaction assay to genotype pyrethroid resistant strains of Boophilus microplus (Acari: Ixodidae). AB - A polymerase chain reaction-based assay was developed to detect the presence of a pyrethroid resistance-associated amino acid substitution in Boophilus microplus (Canestrini). The assay uses a simple method for the extraction of genomic DNA from individual larvae and genotypes individuals for the presence of a Phe-->Ile amino acid substitution in the S6 transmembrane segment of domain III of the para like sodium channel, clearly distinguishing heterozygotes from homozygotes. High frequencies for this amino acid substitution were found in the Corrales and San Felipe strains, which have target site insensitivity mechanisms for pyrethroid resistance. The Caporal resistant strain contained lower yet substantial numbers of amino acid-substituted alleles. Low amino acid substitution frequencies were found in the susceptible reference Gonzales strain and the Coatzacoalcos strain, which has metabolic esterase-mediated pyrethroid resistance. The amino acid substitution was not found in six other strains that were susceptible to pyrethroids. PMID- 11268691 TI - L-lactic acid as a mosquito (Diptera: Culicidae) repellent on human and mouse skin. AB - The attraction of Aedes albopictus (Skuse) to hands and forearms of human subjects treated with several concentrations of L-LA solution were studied in a test chamber containing proboscis-amputated mosquitoes. Fewer mosquitoes alighted on L-LA treated human skin than on water-treated control skin. Similar results were found using normal mosquitoes following L-LA and water treatment of mouse skin. The relative repellent effects of L-LA varied with concentration. The minimum repellent concentration was lower than previously reported for human skin. The number of alightments decreased at increasing concentrations of L-LA, demonstrating the absolute repellency of L-LA. Unlike previous reports suggesting that L-LA attracted mosquitoes, our studies using human and mouse skin showed that L-LA exhibited both relative and absolute repellency. PMID- 11268692 TI - Cryopreservation of embryos of Culicoides sonorensis (Diptera: Ceratopogonidae). AB - The successful long-term storage of insects that are used for research purposes can eliminate the need for ongoing colony maintenance on a large scale. In addition, rare and valuable genotypes of insects can be preserved. This study was conducted to determine whether cryopreservation is a suitable means of storing embryos of Culicoides sonorensis Wirth & Jones, an important vector of animal pathogens. We determined that eggs of C. sonorensis can withstand vigorous treatments of dechorionation, permeabilization, and loading with the cryoprotectant, elthylene glycol. Although their viability was reduced, an average of 80.3% of the embryos developed into larvae. Dehydration in vitrification solution caused a much greater reduction in egg viability (42.7% survival), and freezing in liquid propane further reduced the number of eggs that developed into larvae (40.1%), pupae (22.9%) and adults (18.8%). This work demonstrated that this procedure may prove useful for the cryopreservation of standard laboratory colonies and genetic lines of C. sonorensis. PMID- 11268693 TI - Interrupted blood-feeding by Culiseta melanura (Diptera: Culicidae) on European starlings. AB - To determine whether Culiseta melanura (Coquillett) mosquitoes tend to take multiple blood meals when birds of certain species serve as hosts, we compared the frequencies with which such mosquitoes fed upon caged starlings and robins and determined whether similar volumes of blood were imbibed from each. The blood of robins (Turdus migratorius) and European starlings (Sturnus vulgaris) was marked contrastingly by injecting birds with rubidium or cesium salts. Caged birds were placed together in a natural wetland setting overnight. Mosquitoes captured nearby on the following morning were analyzed for each of the elemental markers. Where marked robins and starlings were equally abundant, 43% of freshly engorged Cs. melanura fed on more than or equal to two hosts. More Cs. melanura fed on robins than on starlings. Individual mosquitoes tended to contain far more robin- than starling-associated marker, indicating that mosquitoes "feasted" on robins but only "nibbled" on starlings. Mosquitoes marked with both elements apparently fed meagerly on the starlings then abundantly on the robins. Our estimates of bloodmeal volume indicate that 85% of mosquitoes that fed on marked starlings obtained < 0.5 microliter of blood from them. We suggest that defensive behavior by starlings interrupts mosquito blood-feeding and that, in a communal roost of starlings, each mosquito will tend to feed on more than one bird, thereby promoting rapid transmission of such ornithonotic arboviruses as eastern equine encephalomyelitis virus and West Nile virus. PMID- 11268694 TI - Prevalence of infection in ticks submitted to the human tick test kit program of the U.S. Army Center for Health Promotion and Preventive Medicine. AB - In 1997, ticks removed from humans and received alive by the Tick-Borne Disease Laboratory of the U.S. Army Center for Health Promotion and Preventive Medicine (USACHPPM) were tested for pathogens by polymerase chain reaction (PCR). Thirty three of 222 (15%) Amblyomma americanum (L.) DNAs produced amplicons of the expected size of Ehrlichia chaffeensis Anderson, Dawson & Wilson and 26/222 (12%) produced amplicons indicating Borrelia burgdorferi Johnson, Schmid, Hyde, Steigalt & Brenner. Five (2%) appeared to be co-infected with both organisms. Thirteen of 308 (4%) Dermacentor variabilis (Say) were PCR-positive for spotted fever group rickettsiae. Restriction fragment-length polymorphism analysis indicated all were Rickettsia montana. One hundred twenty-seven D. variabilis from Monroe County, WI, were tested for B. burgdorferi and 14 (11%) were positive. Five of 24 (21%) Ixodes scapularis Say were positive for B. burgdorferi and one (2%) was positive for the agent of human granulocytic ehrlichiosis. Different species of ticks transmit different pathogens, and most tick-borne diseases have similar early symptoms, therefore knowing the species and infection status of the tick enhances the physician's ability to consider tick-borne agents as a potential cause of disease and recommend appropriate therapy. Ongoing surveillance of the vector species of human diseases provides an additional estimate of human encounters with infected ticks, and testing ticks removed from humans may increase our knowledge of the vector status of tick species for transmitting tick-borne pathogens. PMID- 11268695 TI - Field trial of systemically delivered arthropod development-inhibitor (fluazuron) used to control woodrat fleas (Siphonaptera: Ceratophyllidae) and ticks (Acari: Ixodidae). AB - An orally delivered arthropod development-inhibitory (fluazuron) was evaluated for its potential to reduce the number of flea and tick vectors found on the dusky-footed woodrat Neotoma fuscipes Baird, a reservoir host important in disease enzootiology in northern California. Pigmented bait cubes containing fluazuron were distributed monthly to woodrat nests in a chaparral habitat for 1 yr. When compared with control woodrats, the numbers of fleas [primarily Orchopeas sexdentatus (Baker)] on treated woodrats were significantly reduced 3-4 mo after initial application, and remained so for the duration of the application period. By contrast, tick numbers were not significantly reduced on treated woodrats. After the cessation of treatments, flea indices remained lower on treated animals for up to 2 mo after application. Approximately 93% of woodrats captured in the treatment area excreted pigmented feces and 93% of distributed bait cubes were removed by woodrats, which indicates that the bait cube formulation and delivery system were highly effective. Bait cubes also were attractive to small rodents and ground-frequenting birds. The results of this study suggest that a monthly application program of fluazuron delivered by bait cube is effective in reducing woodrat flea-burdens, but is not effective, at least in the short-term, in controlling ticks. PMID- 11268697 TI - Movement of Aedes aegypti (Diptera: Culicidae) released in a small isolated village on Hainan Island, China. AB - A mark-release-recapture experiment was conducted in a small isolated village on Hainan Island, China, to examine the dispersal and movement of adult Aedes aegypti (L.). Two cohorts of mosquitoes marked with uniquely colored fluorescent dye were released at two different sites and recaptured for 6 d at every house in the village using human bait collections. The distribution pattern of houses around release site affected dispersal. The recapture rate of females released at the center of the village was higher (3.49%) than females released at the edge of the village (2.47%). The average day of recapture differed significantly between sexes, but not cohorts. The average day of recapture of females and males released at the center was 2.5 and 1.54 d, respectively. The total number of mosquitoes recaptured was the greatest at premises near the release site, and decreased at a constant rate of 0.43-0.48 with increasing distance from the release site. The proportion of nulliparous females decreased during the first 4 d and proportion of females with developing or mature ovaries increased during the latter half of the experiment. The daily survival rate for females and males released at the center of the village was estimated by log-regression to be 0.763 and 0.52, respectively. PMID- 11268696 TI - Effects of juvenile hormone on eggs and adults of the cat flea (Siphonaptera: Pulicidae). AB - Juvenile hormone III plays a major role in regulating feeding and reproduction in the adult cat flea, Ctenocephalides felis (Bouche). Both blood consumption and egg production increased in a dose-dependent manner up to a maximum at 1,250 ppm when fleas were continuously exposed to concentrations up to 12,500 ppm juvenile hormone. Histological studies demonstrated that juvenile hormone III also stimulated cellular differentiation of salivary gland epithelia, midgut epithelia, and fat body cells, enhancing the ability of the adult flea to digest blood and synthesize vitellogenins for the maturing oocytes. In unfed fleas, exposure of adults to concentrations of > or = 1,000 ppm juvenile hormone III applied to filter paper resulted in membrane lysis and destruction of salivary gland and midgut epithelial cells, fat body cells, and ovarian tissue. Unlike juvenile hormone mimics, which have potent ovicidal effects in fleas, juvenile hormone had little effect in preventing egg hatch; 58% of the eggs laid by fleas treated with 12,500 ppm juvenile hormone III hatched, and a concentration of 30,000 ppm was required to reduce hatch to 2% in untreated eggs exposed to treated filter paper for 2 h. Compared with the juvenile hormone mimic pyriproxyfen, juvenile hormone III was less toxic to fed adult fleas. However, at a concentration of 12,500 ppm, juvenile hormone killed approximately 45% of the adults and caused autolysis and yolk resorption in the developing oocytes. Thus, at high concentrations, juvenile hormone appears to have a pharmacological effect on fleas, which is highly unusual in insects. PMID- 11268698 TI - Contrasts in tick innate immune responses to Borrelia burgdorferi challenge: immunotolerance in Ixodes scapularis versus immunocompetence in Dermacentor variabilis (Acari: Ixodidae). AB - The blacklegged tick, Ixodes scapularis Say, transmits the Lyme disease spirochete Borrelia burgdorferi, whereas the American dog tick, Dermacentor variabilis (Say), is unable to transmit the bacterium. We compared the innate immune response of these ticks against spirochetes directly inoculated into the hemocoel cavity of ticks. In I. scapularis, some Borrelia were found associated with hemocytes, while numerous other spiral-shaped, intact bacteria remained free in the hemolymph. In contrast, in D. variabilis only remnants of the bacteria were evident in the hemolymph, indicating lysis; intact spirochetes were rare. Spirochetes were observed bound to or within the organs of both tick species, although many more spirochetes were found associated with the I. scapularis organs. The few spirochetes observed with the D. variabilis organs appeared to be dead because D. variabilis tissues rarely contained culturable bacteria, unlike I. scapularis tissues. When spirochetes were incubated with I. scapularis hemolymph plasma in vitro, bacterial survival and motility were not reduced. In contrast, incubation of spirochetes with D. variabilis hemolymph plasma resulted in > 50% of the spirochetes becoming nonmotile by 45 min. The differences in the responses of the two different tick species indicate that I. scapularis is immunotolerant when challenged with B. burgdorferi and dependent on a slow phagocytic response to clear Borrelia from the hemolymph. In contrast, D. variabilis is highly immunocompetent (i.e., innate immunity), using plasma borreliacidal factors and a rapid increase in phagocytic cells to clear the infection and limit tissue invasion. PMID- 11268700 TI - [Pulmonary embolism in children and adolescents]. AB - We present a necropsy study of 25 children (age 0-15 years) and 7 adolescents (age 16-18 years) with the diagnosis of macroscopic pulmonary thromboembolism (PTE). PTE occurred along with a serious disease, e.g. tumour or general infection, and when the subjects had some of the following risk factors, e.g. immobilization, tumours, cardiovascular diseases, operations, obesity, septic state, peritonitis, cachexia and placement of central venous catheters. PMID- 11268699 TI - [Factors which affect cytodiagnosis in gynecology]. AB - The incidence of invasive carcinoma of the cervix in the Czech Republic is higher than in Western Europe. The only effective test for early discovery of the precancerous lesions is cytodiagnostics--"PAP" smears. The study points to preventing mistakes in the cytodiagnostic process and stresses quality control according to the International Academy of Cytology (I.A.C.) and the European Federation of Cytological Societies (E.F.C.S.). PMID- 11268701 TI - [Simultaneous occurrence of papillary carcinoma, oncocytic carcinoma and malignant lymphoma of the thyroid gland in a female patient with Hashimoto's disease]. AB - A 51-year-old woman was admitted for a painless enlargement of the thyroid lasting over 6 months. Hashimoto's thyreoiditis was diagnosed and three tumors were found: oncocytic carcinoma, malignant lymphoma and papillary carcinoma. In the right lobe, oncocytic carcinoma and high grade malignant lymphoma composed of cells with irregular, lobulated nuclei were found. The lymphoma was confined to the thyroid gland. The oncocytic carcinoma invaded the capsule and the surrounding tissues. In the left lobe, there was a papillary carcinoma. PMID- 11268702 TI - [A multimedia atlas of skin pathology]. AB - We prepared a digital atlas of non-tumourous skin pathology. Currently, the available digital atlases can go beyond the scope of written publications, especially with regards to ease of access, speed of preparation, updating information, fair price and unlimited capacity. This atlas can be used as a diagnostic aid, as well as for postgraduate and pregraduate teaching. PMID- 11268704 TI - [Consultation (second opinion on biopsies) as an instrument of quality control in histopathologic diagnosis]. AB - As the whole spectrum of morphological patterns encountered in surgical pathology cannot be, in its entirety, diagnosed correctly by a general pathologist alone, one has to seek consultation for difficult or unusual cases with specialised experts. In the Czech Republic, the consultation practice performed by specialists from different medical institutions is the only way to assess the quality of histopathological diagnostic performance, i.e. it represents an audit system. Audit systems functioning in other countries are described here. A proposal of a standard consultation process as well as problems concerning the financial costs and increased workload of consultants are discussed. PMID- 11268703 TI - [Herwig Hamperl in Prague]. AB - Prof. Herwig Hamperl, M.D. (1899-1976), Chairman of the German Institute of Pathology, Charles University, Prague in years 1940-1945. The account of his work in Prague is first of all based on his autobiography (A Pathologist's Life and Evolution), published in 1972 in a German edition only. It describes the history of the university and conditions of work in the German Institute of Pathology 60 years ago. Hamperl recalls here his contemporaries, his relationships with the Czech pathologists, Prof. Sikl and his Czech students. The autopsy and the cause of death of Reinhard Heydrich are reported as well as the situation at the university and in Prague during the war years. PMID- 11268705 TI - CD 68 positivity of the so-called meconium corpuscles in human foetal intestine. AB - CD 68 antibodies are most frequently used as a reliable paraffin marker of histiocytes. The CD 68 clusters of antibodies recognize a 110-kDa glycoprotein associated with lysosomes and therefore these antibodies show visible reactivity with cells of the monocyte/macrophage lineage. We observed distinct CD 68 positivity of granular intraepithelial inclusions in the small intestine of 12 week-old human foetuses. These PAS positive inclusions corresponded to the so called meconium particles, whose lysosomal origin was confirmed by electron microscopy. Our new observation shows that CD 68 antibodies are specific mainly for organelles and not for a cell type and therefore the staining of epithelial cells containing lysosomes is possible. PMID- 11268707 TI - Human dermal safety studies with eflornithine HCl 13.9% cream (Vaniqa), a novel treatment for excessive facial hair. AB - Eflornithine HCl 13.9% cream (Vaniqa) is a novel treatment for the management of unwanted facial hair in women. This paper reports the results of four modified open-label, within-subject vehicle-controlled studies evaluating the dermal safety of this topical treatment. In a repeated insult patch test (230 subjects), erythema with oedema occurred in 38.9% of subjects treated with eflornithine HCl 13.9% cream and 4.8% of subjects treated with vehicle cream. Challenge applications at previously untested sites following the three-week induction period produced noticeable erythema or greater on only four sites treated with eflornithine HCl 13.9% cream and one vehicle-treated site. The erythema at these sites subsided substantially within 24 hours. In a three-week cumulative irritation study (30 subjects), the mean irritation score for sites treated with eflornithine HCl 13.9% cream was 1.33, compared with 0.76 at vehicle-treated sites and 3.09 at positive-control (sodium lauryl sulphate-treated) sites (p < 0.001 between all three groups). In a phototoxicity study (25 subjects), irradiated sites showed either no reaction (40% of both sites treated with eflornithine HCl 13.9% cream and vehicle-treated sites), or mild erythema subsiding in all cases but one within 24 hours. No reaction was seen at non irradiated sites. In a photocontact allergy study (30 subjects), challenge with eflornithine HCl 13.9% cream or its vehicle alone produced either no reaction or mild erythema subsiding within 24 hours at both irradiated and non-irradiated sites. No serious adverse events were reported during the studies, and the only adverse events considered related to treatment were pruritus (three subjects) and dry skin at test site (one subject). These results demonstrate that eflornithine HCl 13.9% cream does not have contact sensitising, photocontact allergic or phototoxic properties. It can cause irritation under exaggerated conditions of use. Eflornithine HCl 13.9% cream, therefore, has a favourable dermal safety profile appropriate for a topical treatment to be applied routinely. PMID- 11268706 TI - A phase II trial of fractionated vinorelbine/doxorubicin as first-line therapy for advanced breast cancer. AB - In western countries breast cancer is still the leading cause of death of women. Very promising results have been obtained by combining vinorelbine and doxorubicin, two of the most active drugs in metastatic breast cancer. However, despite the activity reported, this combination has shown a 10% rate of grade 2-4 cardiac toxicity, mainly due to the total cumulative doses of anthracycline delivered. The aim of this study was to divide the total dose of doxorubicin into two administrations on days 1 and 8, in order to cut down its toxicity while maintaining the same activity. Fifty-two chemotherapy naive patients with metastatic breast cancer entered into the study and were treated with vinorelbine 25 mg/m2 plus doxorubicin 25 mg/m2 both on days 1 and 8 every three weeks. Fifty one patients were eligible and evaluable for toxicity while 47 of them were evaluable for activity. Haematological toxicity was predominantly related to neutropenia, with grade 3/4 in 16% of cycles. Non-haematological toxicity was represented by alopecia grade 3 (which affected 65% of the patients), local phlebitis and severe constipation. No clinically significant cases of neuropathy or cardiac dysfunction were seen. With regard to activity, 38 out of 47 patients (80%) responded to therapy, nine of them achieving complete responses (19%). Median response duration was 16 months and the median overall survival was 22.7 months. We conclude that the fractionated administration of vinorelbine and doxorubicin is associated with excellent haematological and non-haematological tolerability (especially as regards cardiac toxicity), coupled with high levels of activity comparable to those observed using regimens based on unfractionated administration of treatment. PMID- 11268708 TI - A randomised, single-blind, crossover comparison of the acceptability of the calcium and vitamin D3 supplements Calcichew D3 Forte and Ad Cal D3 in elderly patients. AB - BACKGROUND: Supplements of calcium and vitamin D are given to elderly patients in order to reduce the likelihood of osteoporotic fractures. The acceptability of the preparation is an important component of the compliance of such patients with their treatment. AIM: To compare the acceptability of Calcichew D3 Forte (CDF) and Ad Cal D3 (ACD). METHOD: This was a randomised, crossover, comparative study of two formulations of calcium and vitamin D. Patients took CDF for seven days followed by ACD for seven days, or vice versa, according to the randomisation schedule. At the end of each treatment period, patients used visual analogue scales (VAS) to indicate the grittiness, chalkiness, taste, ease of chewing, ease of swallowing and stickiness of each preparation. RESULTS: One hundred and two elderly patients taking calcium supplements were recruited. Of these, 94 were suitable for inclusion in the efficacy analysis. The VAS scores were significantly lower for CDF than for ACD for grittiness (p = 0.0051), chalkiness (p = 0.0005), ease of chewing (p = 0.0001), ease of swallowing (p = 0.0001) and stickiness (p = 0.0001). These findings indicate that patients found CDF more acceptable than ACD. There was no difference between the groups for the taste score (p = 0.64). Overall, 79.8% of patients stated a preference for CDF, 10.6% preferred ACD and 9.6% had no preference. CONCLUSION: Patients preferred CDF and found it more acceptable than ACD. PMID- 11268709 TI - The prevention of breast cancer: an overview. AB - The role of lifestyle modifications, antioestrogens, cyclo-oxygenase-2 inhibitors and prophylactic mastectomy in reducing breast cancer is reviewed. It is concluded that avoiding postmenopausal obesity and regular physical activity are simple measures that seem to reduce breast cancer risk. There is no conclusive evidence that dietary modification and vitamin supplementation significantly reduce the risk of breast cancer. The evidence suggests that tamoxifen significantly reduces the risk of breast cancer in women at increased risk, but whether it reduces breast cancer mortality remains unknown. Ongoing clinical trials may prove that raloxifene is superior to tamoxifen in breast cancer prevention due to its anti-oestrogenic effects on the endometrium. Bilateral prophylactic mastectomy reduces the risk of breast cancer by 90% in high risk women. PMID- 11268710 TI - Doxofylline: a new generation xanthine bronchodilator devoid of major cardiovascular adverse effects. AB - Doxofylline (7-(1,3-dioxalan-2-ylmethyl) theophylline) is a novel xanthine bronchodilator which differs from theophylline in that it contains a dioxalane group in position 7. Similarly to theophylline, its mechanism of action is related to the inhibition of phosphodiesterase activities, but in contrast it appears to have decreased affinities towards adenosine A1 and A2 receptors, which may account for its better safety profile. The bronchodilating activities of doxofylline have been demonstrated in clinical trials involving patients with either bronchial asthma or chronic obstructive pulmonary disease. In contrast to other bronchodilators, experimental and clinical studies have shown that the drug is devoid of direct stimulatory effects. This may be of importance because the arrhythmogenic actions of bronchodilators may have a negative impact on the survival of patients with respiratory diseases. PMID- 11268711 TI - The effect of atorvastatin on serum lipids, lipoprotein(a) and plasma fibrinogen levels in primary dyslipidaemia--a pilot study involving serial sampling. AB - We conducted an open-label study to test the effects of atorvastatin on serum lipids, lipoprotein(a) [Lp(a)] and plasma fibrinogen levels. A total of 90 dyslipidaemic, non-smoking patients (45 patients with primary hypercholesterolaemia and 45 patients with primary mixed hyperlipidaemia) aged 48 +/- 11 years were studied. The patients were treated with 20 mg of atorvastatin for 24 weeks, in a single nocturnal dose. At baseline and every eight weeks, the fasting lipid profile, together with serum Lp(a) and plasma fibrinogen levels (Clauss method), were measured. Atorvastatin was highly effective in normalising the serum lipid profile. No significant change in median serum Lp(a) levels was observed in the whole group of patients (0.14 g/l before, vs. 0.16 g/l after, treatment) as well as in patients with raised (> 0.30 g/l) baseline levels (n = 32). A small non-significant increase of plasma fibrinogen was found (3.04 g/l vs. 3.14 g/l) after 24 weeks of atorvastatin administration. The effects of atorvastatin on both these variables did not differ in patients with hypercholesterolaemia or mixed hyperlipidaemia. In conclusion, our findings suggest that the effect of atorvastatin on plasma fibrinogen levels in dyslipidaemic patients without evident vascular disease is not clinically relevant. Furthermore, any rise in fibrinogen levels that may occur is likely to be transient in nature. Further studies are necessary to clarify this issue. There was no evidence that atorvastatin influences serum Lp(a) levels. PMID- 11268713 TI - Achieving LDL-C target levels: the role of statins. AB - Epidemiological and clinical studies have clearly established the link between elevated low-density lipoprotein cholesterol (LDL-C) and coronary heart disease (CHD). Consequently, treatment guidelines have been developed that identify LDL-C as a causative factor for CHD and as a target for lipid-lowering therapy. The 3 hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) are considered the drugs of choice for lowering LDL-C in patients with hypercholesterolaemia. However, despite the proven efficacy of statins in reducing LDL-C, many patients do not achieve recommended LDL-C target levels. Surveys suggest that patients do not receive the appropriate drugs in adequate dosage to reach LDL-C target values. Success in achieving targets is lowest in patients at the highest risk, i.e. those with established CHD. The introduction of more potent HMG-CoA reductase inhibitors, or the use of higher doses of older statins, should allow more patients to achieve LDL-C target levels. PMID- 11268712 TI - The Twenty-third Annual San Antonio Breast Cancer Symposium. AB - This paper reviews the recent Twenty-third Annual San Antonio Breast Cancer Symposium. A total of 580 studies were presented either orally or as posters. Two phase III multi-centre clinical trials found that fulvestrant (Falsodex), given as a once-monthly intramuscular injection (250 mg), was well-tolerated and at least as good as anastrozole (1 mg) in postmenopausal women with advanced breast cancer that had progressed or recurred on prior endocrine therapy. Another phase III randomised trial found that letrozole (2.5 mg daily) was superior to tamoxifen as a neoadjuvant therapy in postmenopausal women with ER- and/or PgR positive breast cancer unsuitable for breast-conserving surgery. In a phase III study, capecitabine (Xeloda) was found to be well-tolerated and able to improve survival by three months when added to Taxotere. Cutting edge data on microarray gene profiling in breast cancer were presented. The potential role of this new technology in predicting outcome and selecting therapy was discussed. Furthermore, its limitations and the need for validation were highlighted. The role of new diagnostic tools, such as fibre-optic ductoscopy (FDS) and microcatheters to obtain ductal cells, was discussed. Finally, the worldwide overview was presented. PMID- 11268714 TI - Comparison of acarbose and gliclazide as first-line agents in patients with type 2 diabetes. AB - AIM: To compare the effect of acarbose and gliclazide on clinical findings, biochemical parameters and safety in type 2 diabetic patients insufficiently controlled with medical nutrition therapy (MNT). METHODS: Seventy-two patients (age 35-70 years, BMI < or = 35 kg/m2), who had not taken any oral antidiabetic drug previously, were randomised into two groups after a four-week placebo period, and treated for 24 weeks with acarbose (100 mg two to three times daily) and gliclazide (40-80 mg twice daily). The study was open and 57 patients (33 males and 24 females) completed it. MNT was provided for each patient based on personal requirements as defined by a dietitian. The effect of treatment was evaluated by fasting and postprandial (PP) metabolic parameters (blood glucose, insulin and C peptide levels), HbA1c and plasma lipid levels. In addition, side effects were recorded and clinical examinations performed. RESULTS: Both drugs were effective in reducing of HbA1c, fasting and PP blood glucose levels. However, PP serum insulin levels in the gliclazide group increased more than those in the group treated with acarbose (p = 0.007). Moreover, a small weight reduction was obtained with acarbose treatment but not with gliclazide. Lipid levels were favourably affected by both drugs. Total cholesterol levels decreased in both groups, the decrease only reaching significance in the acarbose group (p = 0.013). However, serum levels of LDL cholesterol decreased in both groups (acarbose and gliclazide, p = 0.033 and p = 0.023, respectively), but the ratio of HDL to LDL cholesterol increased in the acarbose group only (p = 0.045). Both treatments were generally well tolerated. Common complaints in the acarbose group were flatulence and meteorism (29.6%). However, 10.0% of the patients in the gliclazide group reported at least one mild hypoglycaemic episode. CONCLUSIONS: The results of the study demonstrate that acarbose and gliclazide were reasonably effective in improving metabolic control in patients insufficiently controlled with diet alone, and both treatments were well tolerated. Because of its effects on weight reduction and PP hyperinsulinaemia, acarbose may be preferred as a first-line drug, particularly in the treatment of overweight type 2 diabetic patients. PMID- 11268715 TI - Neuroprotection in the newborn infant: interactions between stress, glutamate, glucocorticoids and development. PMID- 11268716 TI - Pharmacology and neuroprotective actions of mGlu receptor ligands. PMID- 11268717 TI - Placental inflammation and brain injury in preterm infants. PMID- 11268718 TI - Possible strategies to protect the preterm brain against the fetal inflammatory response. AB - The accruing evidence that a fetal inflammatory response is the link between antenatal infection and white matter damage in the preterm newborn infant offers room for speculation how this harmful sequence could be interrupted. Enhancement of endogenous protection, response modification, and damage limitation downstream could be helpful strategies for intervention design. Appropriate observational and experimental studies are needed before clinical interventions can be initiated. PMID- 11268719 TI - ATP-sensitive K+ channels and cytoprotection. PMID- 11268720 TI - Is perinatal dexamethasone treatment safe in preterm infants? PMID- 11268721 TI - Transient hypothyroxinaemia in preterm infants. PMID- 11268722 TI - Catecholamines, hypoxic defence and the neonatal brain. PMID- 11268723 TI - Telencephalic angiogenesis: a review. PMID- 11268724 TI - Clinical experience with therapeutic hypothermia in asphyxiated infants. PMID- 11268725 TI - Systemic hypothermia in infants with hypoxic-ischaemic encephalopathy: a French pilot study. PMID- 11268726 TI - Hippocampal selective regional vulnerability and development. PMID- 11268727 TI - Breast-feeding, human milk, long-chain polyunsaturated fatty acids and development. PMID- 11268728 TI - [Value of estrogen and progesterone receptors in the management of intraepithelial squamous lesions of low grade]. AB - The objective of this study was to know the prognostic value of estrogen receptors and progesterone receptors positives (ER+, PR+) in the management of low grade squamous intraepithelial lesions [(LGSIL), human papillomavirus (HPV) and cervical intraepithelial neoplasia grade 1 (CIN 1) and menopausal status]. MATERIAL AND METHODS: From January 1995 to January of 1999, was studied in 144 women with abnormal citology to HPV and CIN grades 1, 2 or 3. Colposcopy was carried out and two biopsies were taken from the suspicious lesion, they were sent for histopathological study, and for ER/PR by the dextran-coated charcoal technique. In addition to the receptor status information, histological type of squamous intraepithelial lesions (SIL), and menopausal status were recorded and analyzed and were correlated later. RESULTS: In normal cervix, 89% and 60% of specimens were ER+, PR+ (+6 fmol/mg protein) There was significant difference in ER status between the normal cervix con HPV and CIN, (P < 0.003), but no with PR (P > 0.53). ER levels in premenopausal and postmenopausal patients there was no significant difference (P > 0.27) but PR levels there was significant difference (P < 0.04). ER/PR levels in HPV o CIN grade no correlated to the histologic grade and menopausal status (P > 0.35, > 0.97, respectively). CONCLUSION: The women with levels ER+ were significant higher in the normal cervix than SIL, ER+, PR+ levels no correlated to the prognostic value in the management of LGSIL and menopausal status. PMID- 11268729 TI - [Induction of nitric oxide synthesis in mononuclear cells in culture using peritoneal fluid from women with endometriosis, in relation to the percentage of T lymphocytes and NK cells identified in an such environment]. AB - Even though endometriosis represents a reproductive health problem of the greatest importance due to the fact that it is one of the most common benign gynecological conditions, its aetiology is still unknown. The most accepted hypothesis is the one proposed by John Sampson, suggesting that the endometrial cells and tissues derived from menstrual flow during uterine scaling reach the peritoneum through the tubes by reversed flow and, under the specific conditions of the peritoneal microenvironment, they are able to implant and proliferate in an ectopic manner. Some evidence shows that the number and activation of macrophages are increased in the peritoneal medium of women with endometriosis. It is known that the activation of this cell group leads to a greater synthesis of diverse molecules associated with this condition. OBJECTIVE: Evaluating the association between the nitric oxide (NO) synthesis induction capacity of the peritoneal fluid, the percentage of cooperative T lymphocytes and NK cells present in the peritoneal medium of women with different stages of endometriosis, as compared to fertile and healthy women. We also tried to find the correlation between the concentration of TNF-alpha identified in the peritoneal fluid of both groups with the NO synthesis induction that was carried out. Material and methods. The study group was formed by women with endometriosis (WEN) from the National Institute of Perinatology, and the control group was formed by patients attending the Family Planning Clinic of the Northeast Regional Unit (Culiacan, Sin.) (HFW). A NO synthesis induction was performed using lymphocytes stimulated with peritoneal fluid from WEN and HFW in order to measure the concentration of cooperative T lymphocytes and NK cells, the TNF-alpha of the peritoneal fluid was also measured. RESULTS: The NO synthesis induction capacity of peritoneal fluid observed with lymphocytes from a culture was greater than the one presented by healthy women. CONCLUSION: Nitric oxide was recently described as a potent inhibitor of effector cytotoxic activity associated to the immunological response of cooperative T lymphocytes of the TH-1 type promoting cytotoxic activity on different cell strains. Evidence suggests that NO inhibits INF-alpha synthesis, the later being a potent proliferation and cytotoxic activity inducer in NK cells, cytotoxic T lymphocytes, and cooperative T lymphocytes. A role of NO as a regulator of NK cell activity has also been described. PMID- 11268731 TI - [Treatment of cervix uteri cancer. 1946]. PMID- 11268730 TI - [Ovarian volume as a predictive factor of ovarian reserve in response to exogenous stimulation with gonadotropins and its correlation with ovum/embryo development in FIVTE/ICSI results]. AB - The evaluation of ovarian reserve in regard to the exogenous response elicited by gonadotropines is directly related to the interconnection of diverse intrafollicular factors involved in ovum components, and thus, to a better embryonic development, which will be reflected in the pregnancy ratios of assisted reproduction programs. A series of markers have been proposed in order to dynamically evaluate the ovarian response: hormone determinations (FSH, E2, FSH:LH index), biochemical factors (inhibine-B), intrafolicular (cytokines, leptin), neuromodulators (GNRH test), and so forth. However, none has shown a specific sensitivity in order to accurately determine the best treatment depending on the values they provide. OBJECTIVE: To evaluate the capacity of ovarian response on the basis of ovarian volume determination. Two groups of patients were studied: group 1 (n = 19) with a basal ovarian volume smaller than 3 cm3, and group 2 (n = 21), those with a volume greater than 3 cm3. Patients in group 2 showed a better response to ovarian stimulation, as well as the collection of better quality ovarian, increased fertilization, segmentation and pregnancy ratios and a lower cancellation index as compared to group 1. It can be concluded that patients with an ovarian volume lower than 3 cm3 on the day before the stimulation will present a poor response to exogenous gonadotropins, thus, this variable must be considered as a marker of ovarian follicular response. PMID- 11268732 TI - [Folic acid levels, homocysteine and polymorphism of methylenetetrahydrofolate reductase enzyme (MTHFR) in patients with pre-eclampsia and eclampsia]. AB - Preeclampsia and eclampsia are the primary causes of maternal mortality. In the state of Nuevo Leon, from 1990 to 1998, these conditions represented 44.1% of maternal deaths. The presence of thrombogenic substances (homocysteine, C protein, and anticardiolipin antibodies) in the mother's blood has been related to this problem. The C677T polymorphism of the enzyme methylene tetrahydrofolate reductase (MTHFR) favors the increase of homocysteine levels, while folic acid (FA) supplementation decreases its levels. OBJECTIVE: To establish the role of FA in the physiopathology of preeclampsia in our environment. KIND OF STUDY: Longitudinal, prospective and comparative. MATERIAL AND METHODS: CASES: Women with severe preeclampsia and/or eclampsia (n-13). CONTROLS: Women in the third trimester of a normal pregnancy (n + 15). 20 mL Blood samples were taken during the first 24 hours of puerperium, and their AF, homocysteine and MTHFR polymorphism were measured. The t Student test and the Exact Fisher test were used to compare between both groups. RESULTS: The values obtained for homocysteine were (x + SD): CASES: 9.85 micromoles/L + 2.88, and controls: 7.61 micromoles/L + 1.32 (p < 0.04). The frequency (%) of the genetic polymorphism for MTHFR was: positive homozygotes (T/T): 38.46 vs. 20, heterozygotes (C/T): 38.46 vs. 26.6, negative homozygotes (C/C): 23 vs 53, for cases and controls, respectively. CONCLUSIONS: According to our study, the frequency of the homozygote state (T/T) of MTHFR and increased blood levels of homocysteine is greater in women suffering from preeclampsia. PMID- 11268733 TI - [Clinical effects of meropenem on infectious diseases in obstetrics and gynecology]. AB - The efficacy and the safety of meropenem (MEPM) were examined in obstetric and gynecological infections and the results shown below were obtained. 1. The subjects were patients with infectious diseases during a non-pregnant period (n = 14), a pregnant period (n = 13) and a puerperal period (n = 11). After intravenous drip infusion of MEPM at 1-2 g/day, the response rate was 38/39 (97.4%) (Excellent: 13/39 (33.3%), Effective: 25/39 (64.1%)). 2. The response rates in a group receiving 1 g/day, a group receiving 2 g/day and a group receiving a combination of both were, respectively, 25/26 (96.2%), 9/9 (100%) and 4/4 (100%). 3. In cases that did not respond to previous treatments, the response rate to MEPM was 20/20 (100%). 4. In clinical efficacy classified by causative organisms, the rate of Excellent was 10/23 (43.5%) and the rate of Effective was 13/23 (56.5%), and the bacterial eradication rate was 21/23 (91.3%). 5. No subjective and objective adverse reactions or abnormalities in clinical laboratory tests due to administration of MEPM were observed. PMID- 11268734 TI - [Nutritional therapy in gastrointestinal diseases. Diets--necessary or superfluous?]. AB - Diets form a part of the treatment concept in numerous gastrointestinal diseases. Their effectiveness, however, varies considerably from one disease to another. Thus, for example, diet is of decisive importance in celiac disease and lactose intolerance. In contrast, dietary measures are ineffective in the treatment of gallstones, and uncertain as a prophylactic measure against biliary colic. While dietetic measures are an important temporary measure in acute pancreatitis, in chronic pancreatitis such an approach is often not complied with, since it includes abstinence from alcohol. In chronic inflammatory bowel disease, diet can ameliorate a number of complications, although it leaves the pathological process itself unaffected. High-fiber diet is, for the most part, ineffective in patients with irritable bowel syndrome. The present article discusses the benefits of dietary measures in a number of gastroenterological disorders. PMID- 11268735 TI - [Improving prognosis of chronic liver diseases with specific nutrition]. AB - A sensible diet is capable of reducing the complications of chronic liver disease and improving the patient's prognosis. In patients with compensated liver disease, adequate nutrition should be ensured, but specific therapeutic measures are not generally required. Patients with decompensated liver disease often have deficient nutrition or malnutrition. In these patients specific dietary measures make good sense. PMID- 11268736 TI - [Emergencies in general practice, 5. Acute vision loss]. PMID- 11268737 TI - [Minimally invasive procedures in heart surgery. How does it work and who profits?]. AB - The leading minimally invasive procedures employed in coronary surgery are minimally invasive direct coronary arterial bypass surgery (MIDCAB) and the Octopus system. These interventions are performed on the beating heart and require no extracorporeal circulation (ECC), thus avoiding the side effects, such as pulmonary or neurological complications, associated with ECC. In surgery on the mitral or aortic valve, the procedures are carried out via small incisions in the non-beating heart, and endovascular bypass systems (e.g. Port-Access) are sometimes needed for EEC. The advantages of small incisions are a reduction in the risk of infection, shorter hospital stay and, in particular, improved cosmesis. A disadvantage is the longer operating time. Only careful patients selection guarantees successful surgery. PMID- 11268738 TI - [Blood group record. Patient record series]. PMID- 11268740 TI - [Spastic syndrome. More mobile again after stroke]. PMID- 11268739 TI - [Genetic diagnosis for general practice. What the general practitioner can decipher from genes]. PMID- 11268741 TI - [Hepatitis C. Chances for treatment increase]. PMID- 11268742 TI - [16. Edema--consider the heart, liver and kidney!]. PMID- 11268743 TI - [Quality assurance. Make guidelines for patients!]. PMID- 11268744 TI - [Cell phones and radio devices again in the news. Eye melanoma caused by telephoning? (interview by Petra Eiden)]. PMID- 11268745 TI - [Patient with dyspnea and thoracic pain. Too infrequently cor pulmonale is suspected]. PMID- 11268746 TI - [Abdominal pain after eating. Caution with "food allergy" diagnosis]. PMID- 11268747 TI - [Enlarged lymph nodes as an incidental finding. How can carcinoma be excluded?]. AB - Swollen lymph nodes are a very common pathological sign. The majority of cases met with at the primary care level are of an unspecific reactive nature. The first aim of the diagnostic evaluation is to exclude a malignant condition. Of decisive importance for any assessment of enlarged lymph nodes is the overall clinical presentation. History-taking should include data on age, course over time, general symptoms and occupation (contact with animals). The physical examination should aim to identify the distribution, size, consistency, tenderness, and site of the swellings. The diagnostic work-up is completed by laboratory investigations and imaging procedures. A stepped approach to the diagnostic evaluation is recommended. PMID- 11268748 TI - [Leukocytosis as an incidental finding. Clues in the blood picture]. AB - An increase in the number of leukocytes is often an early pathological signal that may be seen during the course of numerous diseases. Leukocytosis on its own in the absence of clinical symptoms is highly unspecific. If exogenous influences have been excluded, and if laboratory investigations repeatedly indicate leukocytosis, a differentiation must be made between a reactive and a malignant cell proliferation. In addition to the patient's history and clinical findings, a differential blood count is mandatory. Of differential diagnostic significance, is the extent of the left shift. For the qualitative interpretation of the differential blood count, careful microscopic assessment of a blood smear is required. Should atypical cells be found, further diagnostic measures must be initiated without delay, in a center with appropriate facilities. PMID- 11268749 TI - [Leukocytopenia as an incidental finding. Finding the etiology]. AB - Any leukopenia of less than 1000/microliter poses an acute threat to life, and mandates an immediate search for the underlying cause. An extensive history taking (use of drugs? visits abroad? previous illnesses?) and physical examination (splenomegaly? exanthema? signs of hemorrhage?) are mandatory. In addition to a manual differential blood count, bone marrow aspiration for cytological and histological evaluation must be requested. In this overview, the major differential diagnoses, such as allergic agranulocytosis, leukemia, pernicious illnesses (e.g. malaria), hypersplenic syndrome and a number of infectious diseases are discussed. PMID- 11268750 TI - [Aging without becoming old. 2 anti-aging days per week]. PMID- 11268751 TI - [Treating hypertension only in case of high risk? Evidence-based therapy with cost control]. PMID- 11268752 TI - [Emergencies in general practice, 2. Acute tonsillitis. Differential diagnosis and complications]. PMID- 11268753 TI - [Diagnostic quiz. Abdominal pain with hemolysis. Alcoholic pancreatitis]. PMID- 11268754 TI - [Continuously updating the pain concept. Morphine is a reference, but not a gold standard]. PMID- 11268755 TI - [Youthful appearance from the surgeon. Tightening and tucking]. PMID- 11268757 TI - [One year moxifloxacin. Still effective in the respiratory tract]. PMID- 11268756 TI - [Diabetic patient with insulin analog treatment. More flexible meal regimen]. PMID- 11268758 TI - [Hereditary hair loss. A tincture against expanding alopecia]. PMID- 11268759 TI - [Berlin study examines life after 70. Far removed from wishful thinking]. PMID- 11268760 TI - [Subthreshold psychological disorders. The problem of specificity of unspecified diagnoses]. PMID- 11268762 TI - [Subthreshold psychological disorders]. AB - Subthreshold psychic disorders are mild, masked, atypical, or intense but brief psychopathological syndromes below the threshold of standardized diagnoses. They indicate beginning, intermittent, or residual states of well-known psychic disorders or ("comorbid") syndromes associated to other psychic or somatic disorders or possibly partially morbid states by themselves. At least the subthreshold depressions and anxiety disorders are more than twice as frequent as specified diagnosable psychic disorders and have serious consequences with regard to both individual suffering and cost. Primarily they are a problem for primary medical care. It is mainly up to the general practitioner to recognise beginning or residual psychic disorders underlying disturbed well-being or uneasiness in order to treat them preventively or curatively or to co-treat such comorbid states in somatic diseases. Open questions of research and possible consequences for health policy are discussed. PMID- 11268761 TI - [Recurrent brief depressive episodes. Epidemiology, clinical aspects, diagnosis and therapy]. AB - Recurrent brief depression (RBD), an affective disorder with a similarly high risk of suicidal behavior as major depression (MD), is characterized by depressive episodes occurring about once a month that last only a few days. The combination of RBD and MD, called combined depression (CD), increases the risk of suicidal behavior enormously. Whereas patients with CD are usually in the care of psychiatrists or neurologists, epidemiological data demonstrate that RBD patients usually see only general practitioners and are not recognized as suffering from an affective disorder. Diagnostic criteria for RBD can be found in the ICD-10 and are helpful in both research and clinical routine. Recurrent brief psychiatric syndromes should be taken into differential diagnostic consideration and are discussed in detail in this review. However, the possibility of prospective diagnostic confirmation of RBD and the way of evaluating drug responses in prophylactic intervention of RBD differ essentially from those in treatment of MD and are more related to clinical procedures used in treating migraine or epilepsy. Differences in the courses of RBD and MD are responsible for different requirements in the design of drug treatment studies. Denial of special methodological needs and highly selected patient samples have probably been responsible for false negative results in double-blind, placebo-controlled treatment studies. Although several authors reported successful treatment of RBD with different compounds in about 60 patients, no treatment algorithm can be given. Future treatment studies without the limitations of previous studies are clearly needed in the field of RBD. PMID- 11268763 TI - [Expectations of established neurologists and general practitioners regarding the psychiatric clinic]. AB - OBJECTIVE: Psychiatrists' and general practitioners' expectations regarding psychiatric hospitals were evaluated using qualitative methodology. DESIGN: A content analysis was carried out of interviews with 27 psychiatrists and 24 general practitioners. The statements were analysed quantitatively to assess their relative importance. RESULTS: Immediate delivery of discharge summaries and continuous patient-related communication were mentioned most frequently. In addition, psychiatrists assigned much importance to therapy referral after completion of inpatient care and to the prescription of less expensive drugs. Nearly half of all statements related to the communication process between local and clinic physicians, a quarter to aspects of referral, and a quarter to quality of inpatient care. Differences between psychiatrists and general practitioners mainly concerned aspects of medical education. CONCLUSION: Taking into account these findings, more attention should be paid to psychiatrists' and general practitioners' expectations to reduce problems of interaction between clinicians and their colleagues in private practice; this might be helpful to improve the continuity of psychiatric care. PMID- 11268764 TI - [Practical application of the CERAD test battery as a neuropsychological dementia screening test]. AB - The CERAD neuropsychological test battery is becoming the standard measure for screening cognitive deficits associated with dementia. The seven subtests of the CERAD battery (Mini-Mental State Examination or MMSE, verbal fluency, Modified Boston Naming Test or MBNT, construction ability, learning of word lists, recall, and recognition), a short test of crystallized intelligence (vocabulary test or WST), and a simple test of visuo-motor tracking (number relation test-G or ZVT-G) were applied to 30 healthy control subjects, 49 depressed patients, and 98 mildly to severely demented patients. All subtests of the CERAD battery separated controls from mildly demented patients. Overall, depressed patients scored between controls and mildly demented patients. Score differences between depressed patients and mildly demented patients were significant for MMSE, learning and recall of the word list, verbal fluency, and MBNT. This paper contains a profile sheet for the CERAD battery developed according to published norms that allows rapid transformation of test results into a scaled graphical representation. PMID- 11268765 TI - [HIV patients with psychiatric illnesses. Treatment strategies and drug interactions]. AB - Since the beginning of the endemic phase of the human immunodeficiency virus (HIV), about 60,000 humans have been infected in Germany by this retrovirus. Epidemiological data indicate approximately 2,500 new infections annually. Due to the multiagent antiviral therapy available, only about 600 cases progressed to the clinical picture of full-blown acquired immunodeficiency syndrome (AIDS) in 1999. [Robert-Koch-Institut (Berlin), http://www.rki.de] If an HIV infection or AIDS is current, a higher prevalence of psychiatric disorders in these persons can be anticipated. Psychoactive drugs and their dosages need to be chosen under several considerations and with regard to the HIV infection, since interactions between antiviral and psychotropic drugs occur frequently. The use of psychotropic drugs is often required and thus cannot be ignored. Therefore, psychiatric disorders, medical treatment strategies, interactions between psychotropic and antiretroviral drugs, and biotransformation mechanisms are discussed with regard to an underlying HIV infection. PMID- 11268767 TI - [A case of delusional jealousy in Parkinson disease ]. AB - Delusional jealousy has been reported in organic and "functional" psychotic disorders. In organic psychoses, delusional jealousy is usually associated with impaired orientation or other cognitive disturbances. Delusional jealousy in Parkinson's disease has rarely been reported in the scientific literature. A case of a 49-year-old man suffering from this pathology in the course of Parkinson's disease is presented as to clinical picture, therapy, course, and outcome. PMID- 11268766 TI - [Are there predictors for sexual risk behavior in HIV-infected patients?]. AB - The influence of four coping strategies ("rumination," "search for affiliation," "threat minimization," and "search for information"), four social network dimensions ("affectively positive," "affectively negative," "accepting confidants," and "liking confidants") and sociodemographics on the sexual risk behavior of HIV-infected persons were analyzed in sexual encounters with steady and casual partners. The analysis examines bi- and multivariately the predictors for sexual risk behavior. The study population consisted of 103 asymptomatic HIV infected persons (80 men, 23 women, mean age 34 years, range 21-69 years) who participated in our prospective study and reported their sexual contacts during the previous 6 months. In sexual encounters with steady partners, the risk of unprotected behavior increased with the frequency of sexual contact. In these encounters, coping strategies and social network had no influence on sexual risk behavior. In sexual contacts with casual partners, the number of contacts with these partners was also of importance. The coping strategy "rumination" correlates significantly with enhanced risk behavior. In sexual contacts with casual partners, there was no correlation between sexual risk behavior and the three other coping strategies or social network. In multivariate analysis, the number of sexual contacts was the only significant predictor for sexual risk behavior with steady partners as well as casual ones. As sexual activity with HIV infected persons is not absolutely safe, further prevention campaigns have to focus more on the motivation for safe sex, its situational aspects, and partners' responsibility. PMID- 11268768 TI - [Huntington chorea. A clinically relevant differential diagnosis in geriatric psychiatry patients?]. AB - We report on seven geropsychiatric patients, aged 62 to 86, who had received either in- or outpatient psychiatric treatment for different disorders before being diagnosed with Huntington's disease (HD) by molecular biological methods. At the time of diagnosis, these patients presented only with mild involuntary movements in addition to other, nonspecific psychiatric symptoms such as depressive, paranoid, or dementia symptoms. In six of the seven cases, the HD symptoms had previously been misdiagnosed as tardive dyskinesia because these patients had been treated with neuroleptics in the past. Family histories were nonspecific. Three of the seven patients had family members who were described as "mentally ill" and already deceased. Huntington's disease (HD) should be considered as a differential diagnosis in geropsychiatric patients presenting nonspecific psychiatric symptoms. PMID- 11268769 TI - [History and prospects of transcultural psychiatry and psychotherapy]. AB - Transcultural psychiatry and psychotherapy gained importance with the presence of immigrants and refugees in contemporary society. Transcultural psychiatry examines the interaction of culture with the symptoms and course of mental disorders. It focuses on communication and knowledge transfer between professionals and patients and reflects cultural limits in the perception and classification of psychiatric diseases. The past, present, and future of transcultural psychiatry were discussed during a symposium held in Berlin at a meeting of the Society for Philosophy and the Psychological Sciences and the German Society for Anthropological Medicine, Psychology, and Psychotherapy. PMID- 11268770 TI - [Psychiatry as a subject--graduate education as a requirement]. PMID- 11268771 TI - [Psychiatry as a subject--graduate education as a requirement. On the status of the discussion regarding graduate education in psychiatry and psychotherapy]. PMID- 11268772 TI - Synthesis of Mannich bases of some 2,5-disubstituted 4-thiazolidinones and evaluation of their antimicrobial activities. AB - 5-Phenyl/methyl-5-morpholinomethyl/pyrrolidinomethyl-2-(5- aryl-1,3,4-oxadiazol-2 yl)imino]-4-thiazolidinones (5a-m) were synthesized by the reaction of 5 phenyl/methyl-2-[(5-aryl-1,3,4-oxadiazol -2-yl)imino]-4-thiazolidinones (4a-j) with formaldehyde and morpholine or pyrrolidine. The structures of the compounds were determined by analytical and spectral (IR, 1H-NMR, EIMS) methods. The antibacterial activities of the novel compounds against Staphylococcus aureus ATCC 6538, Staphylococcus epidermidis ATCC 12228, Escherichia coli ATCC 8739, Klebsiella pneumoniae ATCC 4352, Pseudomonas aeruginosa ATCC 1539, Salmonella typhi, Shigella flexneri and Proteus mirabilis and antifungal activity against Candida albicans ATCC 10231 were tested using the disk diffusion method. Compounds 5a, 5b, 5c, 5e, 5g, and 5h were found to be active against S. aureus ATCC 6538 (MIC: 312.5; 39; 19.5; 39; 156; and 78 micrograms/mL respectively) and compounds 5c and 5h against S. flexneri (MIC: both 312.5 micrograms/mL). The minimal inhibitory concentrations of these compounds were determined using the micro dilution method. PMID- 11268773 TI - Non-thiol farnesyltransferase inhibitors: structure-activity relationships of aralkylsubstituted benzophenones. AB - We describe a novel class of benzophenone-based farnesyltransferase inhibitors exploiting a novel aryl binding region in the farnesyltransferase's active site. The present study was mainly focussed on structural modifications of the trimethylene spacer of the 4-phenyl butyroyl residue of our lead structure (IC50 = 530 nM). These modifications turned out to have little effect on activity as had the replacement of the terminal aryl by cyclohexyl (IC50 = 440 nM vs. IC50 = 530 nM). PMID- 11268774 TI - Benzophenone derivatives and related compounds as potent histamine H3-receptor antagonists and potential PET/SPECT ligands. AB - Para-substituted aromatic ethers with benzophenone or related structural elements and a 3-(1H-imidazol-4-yl)propyloxy moiety were prepared by Mitsunobu-type ether synthesis or SNAr reaction. Most of the title compounds possess high antagonist potency in histamine H3-receptor assays in vitro as well as in vivo in mouse CNS following oral administration. After defining 4-(3-(1H-imidazol-4 yl)propyloxy)phenyl phenyl methanone as a new lead, structure-activity relationships were investigated for this new class of compounds. Substitution of the meta'-position of the benzophenone moiety with halogen atoms (e.g., iodine, fluorine) led to compounds with high antagonist potency in vitro as well as in vivo (Ki = 9.3 and 4.3 nM, ED50 = 0.7 and 0.47 mg/kg p.o., 18 and 12, respectively). A receptor profile of several functional in vitro assays for several biogenic amine receptors for the meta'-iodinated derivative demonstrated high selectivity toward the histamine H3 receptor. PMID- 11268777 TI - [Herber Weiner: on his 80th birthday]. PMID- 11268775 TI - Synthesis and biological activity of the structural analogues of (-)-cabenegrin A I. AB - A series of phenylbutene and butanol derivatives (6a-j, 12, 13, 15, 17, 24b,c, 26, 27a,b) were prepared from the readily available resorcinol derivatives 2a-f and 7-hydroxy-chroman (18). The products were tested for inhibitory activity on the LPS-induced TNF-alpha production in the plasma in comparison with that of cabenegrin A-I (1a). PMID- 11268778 TI - [Prof. Dr. Dr. h.c. Herbert Weiner, an interview]. PMID- 11268776 TI - Structural determinants for AMPA agonist activity of aryl or heteroaryl substituted AMPA analogues. Synthesis and pharmacology. AB - We have previously reported the synthesis and pharmacological characterization of analogues of 2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA, 1a), in which the methyl group was replaced by a phenyl group (APPA, 1b) or heteroaryl groups. While 2b and its 3-pyridyl analogue 2-amino-3-[3-hydroxy-5-(3 pyridyl)-4-isoxazolyl]propionic acid (3-Py-AMPA, 3) show very low affinity for AMPA receptors, introduction of heteroaryl substituents containing heteroatom in the 2-position provides potent AMPA receptor agonists. We here report the synthesis and pharmacology of 2-amino-3-(3-hydroxy-5-pyrazinyl-4 isoxazolyl)propionic acid (7) (IC50 = 1.2 microM), which is weaker as an AMPA agonist than AMPA (IC50 = 0.040 microM; EC50 = 3.5 microM) but comparable in potency with 2-Py-AMPA (4) (IC50 = 0.57 microM; EC50 = 7.4 microM), as determined in radioligand binding and electrophysiological experiments, respectively. The AMPA analogues 8a-c, containing 2-, 3-, or 4-methoxyphenyl substituents, respectively, and the corresponding hydroxyphenyl analogues, 9a-c, were also synthesized and evaluated pharmacologically. With the exception of 2-amino-3-[3 hydroxy-5-(2-hydroxyphenyl)-4-isoxazolyl]propionic acid (9a), which is a very weak AMPA agonist (IC50 = 45 microM; EC50 = 324 microM), none of these compounds showed detectable effect at AMPA receptors. PMID- 11268779 TI - [Factoral structure and psychometric properties of the German version of the Toronto Alexithymia Scale (TAS-20) in psychosomatic patients]. AB - The present paper describes the attempt to cross-validate the three-factorial structure of the German Version of the 20-Item Toronto Alexithymia Scale in a population of 419 patients, suffering of psychogenic disorders. However, results neither revealed good internal consistencies nor was it possible to obtain an acceptable adaptation of the empirical data on the postulated model. Therefore an explorative factor analysis was performed in order to get a more consistent factor model. The hereby resulting four-factorial structure was congruent with reference to theoretical and empirical data of Alexithymia. However, additional cross-validation in other populations has to be investigated. PMID- 11268780 TI - [Patients' need for information before surgery]. AB - Frequently, prior to surgery, patients are provided with information in order to meet legal and hospital requirements, but without taking their psychological needs into consideration. In order to learn how to improve this situation, the present study was performed. By means of a semi-structured interview including a newly constructed card system, that avoids giving patients undemanded and possibly harmful implicit information, the need for information of 60 patients before hip- or knee-replacement surgery were investigated in great detail. The results show that 83.3% of the patients want to be prepared by the surgeon, preferably by means of an oral communication (75.0%). Most frequently the day when decision for surgery is made and the day of hospitalization were preferred as point of time for preparation (30% each). Patients were more interested in information about the operation and recovery (43.3% each) than about risks (33.3%). Only 11.7% wanted to get psychological preparation prior to surgery. We conclude that coming into contact and establishing a trustful relationship with the surgeon who does the operation is the most crucial need of surgical patients. He should give presurgical information according to the patient's needs, which often do not focus on the risks, as legal regulations do. By providing psychological preparation for the small number of patients in need, psychological specialists may contribute to improve satisfaction and outcome of surgical patients. PMID- 11268781 TI - [Adaptation of parents to the diagnosis of a chronic disease in their child]. AB - The course and the adaptivity of parental coping with a chronical disease in their child (leukemia, solid tumors, diabetes or epilepsy) was studied during the first three months after diagnosis. 66 parents answered questionnaires to their coping and their quality of life as well as to the perceived quality of life of their children 1-2 weeks and again 8-12 weeks after diagnosis. As coping was stable, the parents' and children's quality of life increased over time. Coping strategies using communication and social support improved the physical well being of the parents. Family orientation and optimism are helpful for parents and children, too, but rumination decreased emotional well-being of the parents. Psychosocial care for families with a chronically ill child should improve open communication and social orientation in an early stage of the disease and should try to change maladaptive cognitive reactions. PMID- 11268782 TI - [Sexual satisfaction of women--development and results of a questionnaire]. AB - In sexology the existing questionnaires do not sufficiently consider the experiencing of sexuality and the extent of sexual satisfaction. That is why a questionnaire was developed which includes, besides the frequency and duration of sexual activities, the satisfaction with frequency and duration of these activities and the desired sexual behaviour. A first study with this questionnaire was carried out on 112 women with heterosexual behaviour, aged 20 to 48 years. The frequent desire with regard to coital orgasm as one result of our investigation confirms the centering of orgasm in other studies. But half of the women do not describe orgasm as the favoured feeling during sexual intercourse. For 37% of the women the emotional and physical closeness to the partner is explicitly more important than experiencing an orgasm. According to the comparison of extreme groups sexual satisfaction particularly correlates with self-determination realized in partnership and with satisfaction of communicational desires and need for tenderness within the partnership. PMID- 11268783 TI - [Psychotherapy and religion: a survey of Northern Bavarian psychotherapists]. AB - By taking a survey of physicians and psychologists accredited as psychotherapists in Franconia (northern Bavaria), we attempted to draw conclusions about the importance of religion for the therapists themselves and for their therapeutic settings. 253 physicians and 78 psychologists returned usable questionnaires (a return rate of 70% for each group). 30% of each group were non-denominational, significantly more men than women were not religious or considered themselves "agnostic/atheistic." Catholic therapists appeared to be more loyal toward their church and more open-minded toward religion in general than protestants and non denominationals, respectively. Around 1/5 of the psychologists had prayed for their patients. Among the psychoanalysts there were no fewer non-denominationals or agnostics/atheists than among physician/psychotherapists or behavioural therapists. The physician/psychoanalysts classified the role of religion as less important than physician/psychotherapists; however, there were marked differences between the psychoanalytic schools. Many therapists could imagine consulting a spiritual counselor in appropriate cases. The results indicate a correlation between the subjective attitude of therapists toward religion and their handling of this topic in therapeutic practice. PMID- 11268785 TI - Thank you and goodbye. PMID- 11268784 TI - Evaluating fire safety in older persons through home visits. AB - The evaluate elders' risk factors for fire injury, we performed in-home assessments on our Geriatric Clinic clients. Nearly two-thirds of the subjects had physical impairments that could compromise escaping a fire. Fire safety equipment often was suboptimal. Nearly three-fourths of our subjects were not worried about fire injury, yet all had at least one fire injury risk factor. Fire safety knowledge was poor. Apathy was common, with fewer than one-third of our subjects complying with our recommendations. PMID- 11268786 TI - Stereotactic breast biopsy: a six-year surgical experience. AB - A retrospective review was done of all stereotactic breast biopsies performed at the Central Baptist Hospital Breast Center from February 1994 through December 1999. A total of 1,080 biopsies were performed in 1,026 patients, all by surgeons working independently. Masses were biopsied in 54% and calcifications in 40%. Eighteen percent of biopsies were malignant. The most common benign diagnosis was fibrocystic disease (72%), followed by fibroadenoma (19%), lymph node (2%), and papilloma (2%). The most common malignant diagnosis was invasive ductal carcinoma (40%) followed by ductal carcinoma in situ (32%) and mixed invasive and in situ ductal carcinoma (19%). A prebiopsy BI-RADS mammographic Category III was associated with a 2% incidence of malignancy; Category IV--17%; Category V--90%. Atypical ductal hyperplasia on stereotactic biopsy was upgraded to a malignant diagnosis after reexcision in 19% of the cases. The false-negative rate was 0.4% (sensitivity 99%) and the complication rate was 3%, mostly related to bleeding. Stereotactic biopsy is a safe and accurate technique for the minimally-invasive diagnosis of abnormal mammograms. PMID- 11268787 TI - Improvements for multipurpose bacteriological identification tables to suit the diagnosis of Vibrio cholerae. AB - The aim of the study was to reduce to key tests the 4 extensive polyvalent diagnostic biochemical tables most widely used in Croatia and to adapt them for the demonstration of Vibrio cholerae and its differentiation from the 3 Vibrios (V. alvinolyticus, V. mentschikovii, V. fluvialis) important in differential diagnosis. The fourth table has now been adapted to differentiate among all 12 Vibrio species known to be human pathogens (V. mimicus, V. cincinatiensis, V. holisae, V. damsela, V. furnisi, V. parahaemolyticus, V. vulnificus, V. carchariae). Using the inductive Learning by Logic Minimization Method (ILLM), we analyzed 2 tables (i.e. identification matrices) that were a part of bioMerieux's commercial packaged polyvalent identification systems widely used in Croatia (API 20E and ATB 32E), as well as 2 compilation tables by M. T. Kelly et al. The tables contained 27, 32, 59 and 8 tests, respectively. Cutting these solely to the key tests involved rationalizing them from 59 to the 5 necessary to differentiate Vibrio cholerae from 3 related Vibrios. Further rationalizations were from 32 to 2 and from 27 to the 3 necessary to differentiate Vibrio cholerae from 2 related Vibrios. By reducing the table of 8 tests to 7, and adding 4 new ones to these we achieved an optimization permitting mutual differentiation of all 12 known human Vibrio pathogens. Use of the selective TCBS plating medium was the only precondition for making these tables effective. PMID- 11268788 TI - Clinical value of biochemical markers in the diagnosis of acute myocardial infarction. AB - Current strategy for the use of biochemical markers in the diagnosis of acute myocardial infarction is not yet uniform. New markers of myocardial damage have significantly altered the former viewpoints. The study included 41 patients with confirmed acute myocardial infarction (25 males and 16 females, age range 42-85 years). Control group comprised of 25 patients with chronic renal failure without signs of acute coronary event (n = 11) and patients with confirmed coronary artery disease (n = 14). The level and activity of CKMB (microgram/L and U/L), and the level of myoglobin and cTnl were determined. The results showed the sensitivity of CKMB (microgram/L) in the first six hours from the onset of pain to be statistically significantly higher than the sensitivity of cTnl, while myoglobin was confirmed to be the earliest marker. Determination of CKMB (U/L) activity should be abandoned since it was found to have the lowest sensitivity and specificity. Also, a combination of myoglobin and CKMB (microgram/L) showed a statistically significantly higher sensitivity and diagnostic efficacy but lower diagnostic specificity compared to the combination of myoglobin and cTnl. PMID- 11268789 TI - Metastasis of renal adenocarcinoma to the pelvic region. AB - Adenocarcinoma is among the most common tumors with a 95% incidence. Renal tumor metastases can occur by the lymphatic, lymphohematogenous, or hematogenous route. A 59-year-old female with metastasis of renal adenocarcinoma, at an unusual localization is presented. Diagnosis was made by ultrasound and cytologic examination, computerized tomography, angiography, and tumor biopsy. Tumor biopsy was the sole successful technique to detect the metastasis of renal adenocarcinoma. PMID- 11268791 TI - Nutrition and dietetics--neglected subjects in medical education. AB - A brief historical review of the development of nutrition and dietetics in the world is followed by concise description of the history of the discipline in Croatia after the foundation of the School of Medicine in Zagreb in 1919. As differentiated from the Schools of Medicine in Zagreb, Rijeka, Osijek and Split, the third, nutritional curriculum at the School of Food Science and Biotechnology has allocated as many as 280 theoretical and 110 practical periods in the form of seminars (15 periods) and practical exercise (95 periods) to nutrition and dietetics. The curriculum includes special course of lectures in dietetics, delivered by a physician specialized in the field. This is followed by the observation that none of the four Croatian Schools of Medicine in Zagreb, Rijeka, Osijek and Split has introduced systematic education in the field in either undergraduate or postgraduate curricula. There are only some sporadic related topics included in undergraduate and postgraduate studies. Thus, the paper points to the serious neglect of this important subject in the curricula of our medical schools, and present some ideas how to solve the problem in the nearest future. The possible organization of a special postgraduate study in nutrition and dietetics for physicians willing to devote themselves professionally to this field of medicine is strongly advocated, because modern nutritional practices and dietetics are likely to soon impose the need of such experts in every large hospital and clinical department. PMID- 11268790 TI - Successful treatment of alloimmune thrombocytopenia using corticosteroid therapy in a woman with two consecutive neonatal deaths--case report. AB - Alloimmune thrombocytopenia is a serious fetal disorder resulting from platelet antigen incompatibility between the mother and the fetus. In mild cases, the diagnosis is usually made upon detection of neonatal thrombocytopenia, but serious consequences such as fetal intracranial hemorrhage and/or unexplained fetal death may complicate the disorder. Various treatment modalities are suggested in the management of alloimmune thrombocytopenia, however, none has yet been confirmed as obviously superior. We report on the successful use of corticosteroids during pregnancy in a woman with a history of two consecutive neonatal deaths due to severe thrombocytopenia and HPA 5b platelet-specific antigen incompatibility. PMID- 11268792 TI - The scientific challenges for the third millennium. PMID- 11268793 TI - Effect of opioid peptide methionine-enkephalin in long-term cultures of human bone marrow. AB - Methionine enkephalin, an opioid peptide belonging to the family of neuropeptides, has been shown to function as a neurotransmitter, hormone and growth factor. The present work explored its effects in long-term culture of bone marrow cells, harvested from a patient with acute lymphoid leukemia (ALL-L3) in the second complete remission. Nine cultivation flasks were established and maintained for five weeks, with medium renewal once a week. At each re-feeding, methionine-enkephalin was added to the cultures in final concentrations 10(-8), 10(-10) or 10(-12) M, and granulocyte-macrophage progenitor cells (GM-CFU) were determined among the harvested, nonadherent cell populations. The total number of nonadherent cells was 8% to 42% higher in the treated cultures than in the control, nontreated cultures, and the GM-CFU counts were three to four times higher. Those changes, although evident, did not reach statistical significance because of the small group sizes. In 1 of 9 cultures the adherent cell layer was atypical, the cell population consisted of small cells resembling the lymphoblasts, and the cell count was 2-8 times higher than in the controls. That aberrant culture has presumably arisen from residual leukemic cells remaining in the bone marrow after chemotherapy. The findings support the idea that opioid peptides, including methionine-enkephalin, participate in regulation of hematopoiesis. Two mechanisms may have accounted for the observed effects of enkephalin on cultured bone marrow cells: an indirect action, via interleukins secreted from the stromal cells upon stimulation of the opioid receptors, or a direct action on hematopoietic precursors. PMID- 11268795 TI - IOM committee calls for complete revamping of health care system to achieve better quality. AB - The health care system can no longer deliver quality care the way it is currently organized, according to a new report, Crossing the Quality Chasm, released March 1 by the Institute of Medicine's Committee on Quality of Health Care in America. Instead, the committee says that it is time to reinvent the system. PMID- 11268794 TI - Diagnosis and management of childhood food allergy. AB - The pediatrician plays a pivotal role in the initial diagnosis of food allergy. Alternative diagnoses are considered as a careful history, physical examination, and directed laboratory tests determine the type of adverse reaction and the responsible food. Through elimination diets in infants, appropriately selected tests for specific IgE, and, in some cases, supervised oral food challenges, a diagnosis is secured. Treatment consists of strict dietary elimination with provisions for emergency management of accidental ingestions. Referral to an allergist and dietitian is made as warranted by the severity and type of allergy and for follow-up for possible resolution of the allergy. The pediatrician also provides information to the family for the prevention of allergy in at-risk newborns. Future diagnostic tests and treatment modalities are likely to simplify the management of the food allergic child. PMID- 11268796 TI - Psychrometric chart aids evaluation of IAQ problems--Part III. PMID- 11268797 TI - The search for welding safety. Many good Web sites are out there, just waiting to be discovered. You have to dig down a bit to find those nuggets. PMID- 11268798 TI - Controlling the human element. PMID- 11268799 TI - Fall protection. Networking with fall prevention experts. PMID- 11268800 TI - Fall protection. Engineered fall protection systems. PMID- 11268801 TI - Focus on hearing, head & face protection. Ten steps to welding safety. PMID- 11268802 TI - Focus on hearing, head & face protection. Construction cowboys & head protection. PMID- 11268803 TI - Focus on hearing, head & face protection. A good start for the new millennium. PMID- 11268804 TI - Focus on hearing, head & face protection. The many faces of hearing loss prevention. PMID- 11268805 TI - [Alcohol and pregnancy]. PMID- 11268806 TI - [Child and adolescent psychiatry. Research perspectives]. PMID- 11268807 TI - [The secrecy aspect of donor insemination]. AB - A literature review among couples treated with donor insemination (DI) showed that almost all participants had planned not to inform the child that this treatment had taken place. Many of the couples had told other people about the treatment, and thus opened up the possibility of a traumatic disclosure. Though secrecy about DI treatment could be taxing for the couple, this strain did not negatively influence the assessment of the relationship, parenthood, or the parent-child relation. Psychosocial development of DI-children was not different from other children's development. It was not possible to show a deleterious effect of keeping the treatment a secret. There is a lack of qualitative studies about the consequences of secrecy for the relationship between DI-parents, for parenthood and for the child. Only one study included adult DI-children. PMID- 11268809 TI - [Audiometric surveillance of occupational environment]. AB - Noise levels in Danish workplaces at which hearing impairment will occur in more than 10% of the workers are still frequent. Annually 300 cases are compensated for noise induced hearing loss at a national level. This may indicate that the current national hearing protection strategy, which relies on noise reduction only, is insufficient. By audiometric screening early hearing loss will be detected and further progression may be halted. This paper reviews the methods of continuous audiometric screening of noise exposed worker populations. Reliability and validity are discussed, and an overview of quality assurance programmes is given. It is concluded that a national hearing conservation program should be implemented in Denmark. PMID- 11268808 TI - [Treatment of esophageal varices]. AB - Patients with oesophageal varices without previous bleeding have a risk of 25-30% for the development of a bleeding episode within two years. It is important to identify patients with a high risk of bleeding due to the high mortality of 30 40% within six weeks after a bleeding episode. Treatment with a non-selective betablocker possibly combined with isosorbidnitrate should be initiated if an upper endoscopy shows the presence of medium- or large-sized oesophageal varices. Sclerotherapy or ligation initiated within a short time after start of an acute bleeding episode reduces mortality and risk of bleeding. Pharmacological treatment may have effect on the acute episode, and most data advocate for Terlipressin to be the first choice. If bleeding continues transjugular intrahepatic portosystemic shunt (TIPS) should be considered. Fifty to seventy percent of patients will experience a rebleeding episode if otherwise untreated. Endoscopic ligation should be preferred possibly in combination with a non selective betablocker. If another bleed occurs the patient should be considered for TIPS or transplantation. PMID- 11268810 TI - [Did the changed guidelines on alcohol and pregnancy by the National Board of Health and Welfare change alcohol consumption of pregnant women?]. AB - BACKGROUND: In March 1999, the National Board of Health changed its recommendations about alcohol drinking in pregnancy. A "No drinking" policy was changed to three recommendations: 1. Avoid, as far as possible, alcohol in pregnancy; 2. Never take more than one drink a day; and 3. Do not drink alcohol every day. AIM: By means of data from the Danish National Birth Cohort, to monitor changes in pregnant women's reporting of alcohol drinking before and after the change in policy. METHODS: From October 1, 1997, to September 30, 1999, a total of 30,899 pregnant women were interviewed by the end of first trimester. Information on alcohol intake reported among women interviewed from July 1, to September 30, 1998 was compared with the same information obtained from interviews completed in the same months in 1999. RESULTS: Overall, there were no changes in mean alcohol intake in the two periods. Hence, the mean intake was 0.6 drinks per week in the period before and 0.7 drinks per week in the period after introduction of the recommendations. The proportion of women drinking more than two drinks per week was 6.4% before vs 7.4% after the new and less restrictive recommendations (p = 0.12). The proportion of women having one or more binge episode (e.g. drinking five or more drink at one occasion) was 26.7 vs 27.4 (p = 0.65). CONCLUSION: The study showed no significant changes in drinking habits among Danish pregnant women after relaxation of the guidelines for sensible drinking during pregnancy. PMID- 11268811 TI - [Legal abortion. An analysis of factors which can affect frequency of complications]. AB - INTRODUCTION: Due to a rather high complication rate, we decided to analyze data from women, who were readmitted to hospital with complications to legal abortion. MATERIAL: 760 legal abortions performed in 1993-1994 were retrospectively reviewed for complications. RESULTS: We found 66 complications (8.7% (95% confidence limits (6.7-10.7)), that resulted in readmission to hospital. The distribution of complications was as follows: retention 41 (62% (50-74)), infection 12 (18% (9-27)), bleeding and pain 11 (17% (8-26)) and perforation two (3% (0-7)). There was a slight correlation between a stronger midline echo on ultrasound measurement of the uterine cavity and finding placental tissue on histological examination of the material obtained from reevacuation. The time interval between legal abortion and reevacuation was nine days. Retroflexio uteri was not correlated with reevacuation. CONCLUSIONS: We found a statistically increased risk of complications following legal abortion relative to the figures reported by the national health authority, but not significantly higher than that found in other reports. Moreover, a conservative attitude to retention can reduce the number of reevacuatio uteri after legal abortion. PMID- 11268812 TI - [Surgeons' experience of complication frequency--registration validity for legal abortion]. AB - INTRODUCTION: The aim was to investigate the degree to which experience influenced the complication rate (cr) in legal abortion. We also compared our cr with that registered by the National Health Service for the same group of patients. MATERIAL: 760 legal abortions from 1993 to 1994 were reviewed retrospectively. RESULTS: We found an overall cr of 8.7% (95% confidence limit: 6.7-10.7). The cr was the same for all groups of doctors, from the youngest to the most senior and experienced surgeons. The experience in months of the younger surgeons was noted for each intervention, and we found that they did not improve their results over time or by practice. In the same study, we found that the National Health Service had received, registered, and published statistical data from the hospitals, which were not entirely accurate. CONCLUSION: The complication rate was not associated with the experience of the surgeon. PMID- 11268813 TI - [Picture of the month: perihepatic adherences in Fitz-Hugh-Curtis syndrome]. PMID- 11268814 TI - [Antibiotics to newborn infants with sepsis: effect and ecology]. PMID- 11268815 TI - [Do more women lose their breast following screening--and when are we supposed to discuss it?]. PMID- 11268816 TI - [Open letter to the editors of Rationel Farmakoterapi. A greeting from the target group]. PMID- 11268817 TI - Update on anticonvulsants for the treatment of alcohol withdrawal. AB - Some anticonvulsants have been shown to be as effective as some benzodiazepines for the treatment of alcohol withdrawal. Anticonvulsants may offer advantages over benzodiazepines in the outpatient treatment of alcohol withdrawal: they lack abuse potential, have minimal interactions with alcohol, and may be more effective in ameliorating psychiatric symptoms of alcohol withdrawal. Carbamazepine appears to be as effective as lorazepam and oxazepam in ameliorating the symptoms of alcohol withdrawal. In addition, a recent study indicates that carbamazepine may suppress post-withdrawal alcohol use. Divalproex may also reduce symptoms of alcohol withdrawal, based on several open-label studies. However, both carbamazepine and divalproex have limited usefulness in alcoholics with severe hepatic or hematologic complications. Newer anticonvulsants, such as gabapentin and vigabatrin, also appear to reduce alcohol withdrawal symptoms in preclinical and open-label clinical trials while lacking the toxicities of carbamazepine and divalproex. Controlled trials are underway exploring the efficacy and safety of newer anticonvulsants for the treatment of alcohol withdrawal. PMID- 11268818 TI - Carbohydrate-deficient transferrin: an aid to early recognition of alcohol relapse. AB - Although the primary use of biochemical markers of heavy drinking is to assist in screening for alcohol problems, laboratory tests may also aid in early identification of relapse. This report reviews research findings on a new marker, carbohydrate deficient transferrin (CDT), in alcoholics receiving treatment or in follow-up. It also offers recommendations on how CDT may be employed by clinicians monitoring drinking status. PMID- 11268819 TI - Liver diseases by alcohol and hepatitis C: early detection and new insights in pathogenesis lead to improved treatment. AB - Much progress has been made in the understanding of the pathogenesis of alcoholic liver disease, resulting in improvement of treatment. Therapy must include correction of nutritional deficiencies, while taking into account changes of nutritional requirements. Methionine is normally activated to S adenosylmethionine (SAMe). However, in liver disease, the corresponding enzyme is depressed. The resulting deficiencies can be attenuated by the administration of SAMe but not by methionine. Similarly, phosphatidylethanolamine methyltransferase activity is depressed, but the lacking phosphatidylcholine (PC) can be administrated as polyenylphosphatidylcholine (PPC). Chronic ethanol consumption increases CYP2E1, resulting in increased generation of toxic acetaldehyde and free radicals, tolerance to ethanol and other drugs, and multiple ethanol-drug interactions. Experimentally, PPC opposes CYP2E1 induction and fibrosis. Alcoholism and hepatitis C infection commonly co-exist, with acceleration of fibrosis, cirrhosis, and hepatocellular carcinoma. PPC is being tested clinically as a corresponding antifibrotic agent. Available antiviral agents are contraindicated in the alcoholic. Anti-inflammatory agents, such as steroids, may be selectively useful. Finally, anticraving agents, such as naltrexone or acamprosate, should be part of therapy. PMID- 11268820 TI - New developments in the pharmacotherapy of alcohol dependence. AB - Neuroscientific underpinnings and pharmacotherapeutic treatments of substance use disorders are rapidly developing areas of study. In particular, there have been exciting new developments in our understanding of the involvement of excitatory amino acid neurotransmitter systems and the opiate and serotonin systems in the pathophysiology of alcohol withdrawal, alcohol dependence, and in subtypes of individuals with alcoholism. In this article, new developments in the pharmacotherapy of alcohol dependence will be reviewed. In particular, the use of anticonvulsants in alcohol withdrawal and protracted abstinence syndromes will be discussed. New data on opiate antagonists and acamprosate, an agent that exerts actions through excitatory amino acid systems in relapse prevention, will be reviewed. Finally, there will be a review of new data concerning the use of serotonin reuptake inhibitors in subtypes of alcoholism and the use of combination pharmacotherapy. PMID- 11268821 TI - The community reinforcement approach to the treatment of substance use disorders. AB - Empirical support is presented for the Community Reinforcement Approach (CRA), a broad-spectrum cognitive-behavioral treatment for substance use disorders. At the core of CRA is the belief that an individual's environment can play a powerful role in encouraging or discouraging drinking and drug use. Consequently, it attempts to rearrange contingencies so that sober behavior is more rewarding than substance-abusing behavior. Originally tested in the early 1970s with a small sample of alcohol-dependent inpatients, it has repeatedly proven to be successful over the years with larger, diverse populations. Empirical backing is also presented for a new variant of CRA that works through family members to engage treatment-resistant individuals into substance abuse treatment. PMID- 11268822 TI - Intimate partner violence, dependence symptoms and social consequences from drinking among white, black and Hispanic couples in the United States. AB - This study estimates the association of intimate partner violence (IPV) and alcohol-dependence symptoms, social consequences from drinking, and drug use among white, black, and Hispanic couples. A probability sample of 555 white, 358 black, and 527 Hispanic couples in the U.S. household population was interviewed in 1995. The response rate was 85%. Bivariate analysis indicates that most problem status variables are associated with increased rates of male-to-female (MFPV) and female-to-male (FMPV) partner violence. Logistic regression analyses showed that predictors of MFPV and FMPV vary by ethnicity. PMID- 11268824 TI - Prevention of alcohol problems in the 21st century: challenges and opportunities. AB - The community is the "new frontier" for alcohol and other drug prevention. New prevention initiatives at the community level suggest that effective strategies will often be quite different from national or state policies and will require a different perspective. Alcohol and other drug use is part of routine community life and must be considered in the context of the community, which is itself a dynamic and self-adapting system. To develop effective community-level interventions, prevention planners and policy makers must understand how various aspects of the community influence alcohol and other drug use and even contribute to alcohol and other drug problems. This paper outlines the basis for a systems approach to community prevention and the policy options that this approach suggests. It also examines the new science of complexity, differentiates between catchment and a systems approach to prevention, describes a public health model within a systems approach, and describes using local policy as a means to produce system changes as well as recent findings from community-based prevention efforts that employed local alcohol policies. PMID- 11268823 TI - Alcohol introxication and violent crime: implications for public health policy. AB - Despite extensive public health campaigns, the consequences of alcohol intoxication continue to be a serious public health concern. Alcohol intoxication, for example, is a prevalent feature of crime, especially violent crime. Previous studies of alcohol intoxication and violent crime have used samples of police reports, correctional populations (arrestees, jail detainees, or convicted offenders), or community surveys. Studies using police reports and correctional populations are biased because few police-citizen encounters result in police reports or arrest. Community surveys avoid these biases but rely on the subject's assessment of both the victims' and the suspects' intoxication. We took a different approach and directly observed 2,365 police-citizen encounters. Observers used the Alcohol Symptom Checklist to determine the level of alcohol intoxication or impairment. We compared the prevalence of suspects' and victims' alcohol intoxication (equivalent to a blood alcohol level [BAL] of .05 or above) by type of encounter and computed odds ratios to assess the association between intoxication and type of encounter. We also controlled for demographic characteristics (race, gender, age, and socio-economic status) to assess the relationships among perpetration, victimization, and intoxication. Overall, suspects are far more likely than victims to be intoxicated; not surprisingly, suspects in public order/vandalism encounters are the most likely to be intoxicated. Alcohol intoxication appears to contribute substantially to violent victimization. The role of alcohol intoxication is largest among groups that, if not intoxicated, are generally less vulnerable to violence: whites, males, and persons of higher socio-economic status. We discuss the implications of these findings for services and public health policy. PMID- 11268825 TI - Maternal transmission of nicotine dependence: psychiatric, neurocognitive and prenatal factors. AB - This paper reviews the literature on maternal influences on smoking behaviors of offspring from the perspective of neuropsychiatric deficits that may be transmitted from mother to child. In particular, we review what is known regarding associations between: (1) in-utero exposure to smoking, (2) adolescent neurocognitive functioning and psychiatric comorbidity, and (3) the patterns of smoking and progression of nicotine dependence. Furthering our knowledge of these differences in susceptibility to nicotine dependence among youth will provide additional avenues for prevention and intervention efforts targeted toward those at high risk for dependence. PMID- 11268826 TI - Screening and diagnosis of anxiety and mood disorders in substance abuse patients. AB - The aims of this study are (1) to study the prevalence of anxiety and mood disorders in a clinical substance abuse population, (2) to asses the pre- and post-detoxification change in SCL-90 score in this population for subjects with psychopathology compared to subjects without psychopathology, and (3) to asses the value of the SCL-90 and the Addiction Severity Index-psychiatric problems scale as clinical diagnostic screening instrument for psychopathology. The design was a longitudinal prospective cohort study with pre-detoxification ASI and SCL 90 data and post-detoxification CIDI and SCL-90 data on a clinical sample of 116 substance abuse patients. The present results indicate that the ASI-Psychiatric problems score is a limited indicator of psychiatric comorbidity. When a pre detoxification screening for psychopathology is warranted, the present results show that the use of the SCL-90 is preferable above the ASI-PSY scale. Of the screening scores under study, the post-detoxification SCL-90 score is found to be the most valid screening instrument for diagnosis of anxiety and mood disorders in a clinical substance abuse population. Although the post-detoxification SCL-90 score holds moderate specificity combined with high sensitivity, applying this instrument in clinical substance abuse practice will result in a substantial reduction of patients unnecessarily referred for further psychiatric diagnostic evaluation. Further studies aimed at improvement of screening instruments in this population are needed. PMID- 11268827 TI - Modifying residents' professional attitudes about substance abuse treatment and training. AB - Some physicians have negative attitudes and beliefs towards patients with addiction. Moreover, few residents are inclined towards a subspecialty fellowship in addiction psychiatry. We aimed to determine if a one-day educational conference could facilitate attitudinal change among 52 general psychiatry residents. Significant changes (p < 0.05) in attitudes were reported following the conference, including enhanced beliefs that physicians can motivate their addicted patients to seek treatment and increased physician interest in pursuing advanced addiction training. A one-day educational intervention may be effective in improving professional attitudes toward addiction treatment by reinforcing previously acquired medical education. The duration of these changes remains to be determined. PMID- 11268828 TI - Assessing the risks and benefits of benzodiazepines for anxiety disorders in patients with a history of substance abuse or dependence. AB - In this article, the authors reevaluate the traditional position that benzodiazepines should be avoided in anxiety disorder patients with a history of substance abuse or dependence. The efficacy of benzodiazepines in each of the anxiety disorders is reviewed, as are their side effects and toxicity. The definitions of benzodiazepine abuse and dependence are discussed, and relevant animal, experimental, and clinical data are reviewed and analyzed. A manual and computerized (MEDLINE) search was performed from 1966 to the present to examine the English-language literature published on benzodiazepines, substance abuse, and each of the anxiety disorders listed in DSM-IV. The authors found that benzodiazepines have demonstrated efficacy in generalized anxiety disorder, panic disorder, and agoraphobia; they are promising agents in the treatment of social phobia and alcohol-induced anxiety disorders. They are generally well tolerated. There is much ambiguity over appropriate definitions for benzodiazepine abuse and dependence: although most benzodiazepine abusers concurrently abuse other substances, there is little evidence to indicate that a history of substance abuse is a major risk factor for future benzodiazepine abuse or dependence. Furthermore, benzodiazepines do not appear to induce relapse of substance abuse in these patients. The authors conclude that the position that benzodiazepines are contraindicated in former substance abusers appears to lack empirical justification. Benzodiazepines may be indicated in certain patients with anxiety disorders and a history of substance abuse or dependence. PMID- 11268829 TI - Predictors of nonadherence to HIV-related medication regimens during methadone stabilization. AB - Nonadherence to HIV-related medication regimens among drug-abusing patients decreases therapeutic effectiveness and may limit patient access to newer, highly active antiretroviral therapies (HAART). A number of factors have been associated with medication nonadherence; however, few studies have examined predictors of nonadherence specifically in HIV-positive drug abusers. In the current study, a comprehensive assessment battery was administered to 42 HIV-positive, injection drug users beginning methadone maintenance. HIV-related medication adherence was assessed weekly by self-report during the 4-week methadone stabilization phase. Thirty-six percent of patients reported less than 80% adherence to their medication regimen at entry into methadone. Medication adherence increased significantly during the 4-week stabilization phase. Significant zero-order correlations were found between nonadherence during stabilization and viral load, low educational attainment, depression, and neuropsychological tests of problem solving ability and cognitive flexibility. Independent predictors of nonadherence were low levels of education and poor emotional functioning. Implications for early intervention are discussed. PMID- 11268830 TI - Clinical review of inhalants. AB - While the rate of inhalant abuse continues to rise in this country, it remains one of the least studied or discussed groups of abused substances. This review focuses on the current knowledge of the epidemiology, pharmacology, and sequela of inhalant abuse. We will discuss three groups of inhalants: volatile solvents, nitrous oxide, and nitrites. We will then conclude by proposing means by which inhalant abuse may be prevented and treated. PMID- 11268831 TI - Confidence of compliance: a Bayesian approach for percentile standards. AB - Rules for assessing compliance with percentile standards commonly limit the number of exceedances permitted in a batch of samples taken over a defined assessment period. Such rules are commonly developed using classical statistical methods. Results from alternative Bayesian methods are presented (using beta distributed prior information and a binomial likelihood), resulting in "confidence of compliance" graphs. These allow simple reading of the consumer's risk and the supplier's risks for any proposed rule. The influence of the prior assumptions required by the Bayesian technique on the confidence results is demonstrated, using two reference priors (uniform and Jeffreys') and also using optimistic and pessimistic user-defined priors. All four give less pessimistic results than does the classical technique, because interpreting classical results as "confidence of compliance" actually invokes a Bayesian approach with an extreme prior distribution. Jeffreys' prior is shown to be the most generally appropriate choice of prior distribution. Cost savings can be expected using rules based on this approach. PMID- 11268833 TI - Determination of 1-aminopropan-2-one, a dissolved sewage component, in water samples. AB - A new method for the determination of 1-aminopropan-2-one (APR) in water samples was developed. APR was synthesised as its hydrochloride and derivatized with 2,4 dinitrophenylhydrazine (DNPH) for determination by high-pressure liquid chromatography with ultraviolet detection (UV-HPLC). APR was determined in water samples at pH 12 using a gas-stripping chamber, connected to a cartridge containing DNPH. Acidified water samples were injected into the gas-stripping chamber and a solution of NaOH added to bring the solution to pH 12. APR was volatilised and stripped from solution and passed onto the cartridge under a constant stream of nitrogen gas. Gas flow rates were carefully controlled to allow maximum contact of APR with DNPH on the cartridge. When the reaction time had elapsed, the cartridge was disconnected and the derivative eluted with a fixed volume of acetonitrile and injected onto the HPLC, where the APR hydrazone was resolved isocratically with a mobile phase consisting of acetonitrile and water (60:40). The HPLC was calibrated using standard solutions of the APR hydrazone. Recoveries of APR from standard samples were 90-100% at the 10 microM level and the detection limit for the method was calculated as 18 nM. Detection of APR in urine and primary-treated sewage samples (41 nM and 1.225 microM, respectively) confirmed the applicability of the technique to analysis of environmental samples. PMID- 11268832 TI - Process development for the removal of lead and chromium from aqueous solutions using red mud--an aluminium industry waste. AB - Red mud, an aluminium industry waste, has been converted into an inexpensive and efficient adsorbent and used for the removal of lead and chromium from aqueous solutions. Effect of various factors on the removal of these metal ions from water (e.g. pH, adsorbent dose, adsorbate concentration, temperature, particle size, etc.) has been studied and discussed. The effect of presence of other metal ions/surfactants on the removal of Pb2+ and Cr6+ has also been studied. The material exhibits good adsorption capacity and the data follow both Freundlich and Langmuir models. Thermodynamic parameters indicate the feasibility of the process. Kinetic studies have been performed to understand the mechanism of adsorption. Dynamic modelling of lead and chromium removal on red mud has been undertaken and found to follow first-order kinetics. The rate constant and mass transfer coefficient have also been evaluated under optimum conditions of removal in order to understand the mechanism. Column studies have been carried out to compare these with batch capacities. The recovery of Pb2+ and Cr6+ and chemical regeneration of the spent column have also been tried. PMID- 11268834 TI - Bioaccumulation of polychlorinated dibenzo-p-dioxins and dibenzofurans in the foodweb of Ya-Er Lake area, China. AB - Bioaccumulation of PCDD/F in the foodweb was investigated in the Ya-Er Lake area, which was heavily polluted by PCDD/F. The high concentrations of PCDD/F in sediment can be transferred and bioaccumulated by aquatic organisms and humans through various pathways. Benthonic invertebrate animals and aquatic plants with a lot of fibers in the root can accumulate PCDD/F from sediment and water. Snail (Bellamya aeruginosa), shrimp (Macrobranchium sp.) and freshwater mussel (Acuticosta chinensis (Lea)) took up PCDD/F from the water and maintained the emission patterns, whereas fish tended to selectively accumulate 2,3,7,8 substituted isomers. The tissues of fish-eating bird and duck (Anas platyrhynchos) were very highly contaminated by PCDD/F due to ingestion of fish and other aquatic organisms from sediment. The residual concentration in breast milk depended on the original concentration of PCDD/F in the food. A resident in Ya-Er Lake area showed a daily intake of PCDD/F of about 9.14 pg TEQ/kg body weight/day. This is higher than the tolerable daily intake (TDI) for PCDD/F (1 pg TEQ/kg body weight/day), which was recommended by the World Health Organization (WHO). PMID- 11268835 TI - Evaluation of biofilm image thresholding methods. AB - To evaluate biomass distribution in heterogeneous biofilms from their microscope images, it is often necessary to perform image thresholding by converting the gray-scale images to binary images consisting of a foreground of biomass material and a background of interstitial space. The selection of the gray-scale intensity used for thresholding is arbitrary but under the control of the operator, which may produce unacceptable levels of variability among operators. The quality of numerical information extracted from the images is diminished by such variability, and it is desirable to find a method that improves the reproducibility of thresholding operations. Automatic methods of thresholding provide this reproducibility, but often at the expense of accuracy, as they consistently set thresholds that differ significantly from what human operators would choose. The performance of five automatic image thresholding algorithms was tested in this study: (1) local entropy; (2) joint entropy; (3) relative entropy; (4) Renyi's entropy; and (5) iterative selection. Only the iterative selection method was satisfactory in that it was consistently setting the threshold level near that set manually. The extraction of feature information from biofilm images benefits from automatic thresholding and can be extended to other fields, such as medical imaging. PMID- 11268836 TI - Phosphorus removal by sands for use as media in subsurface flow constructed reed beds. AB - Sorption of P to the bed sand medium is a major removal mechanism for P in subsurface flow constructed reed beds. Selecting a sand medium with a high P sorption capacity is therefore important to obtain a sustained P-removal. The objective of this study was to evaluate the P-removal capacities of 13 Danish sands and to relate the removal to their physico-chemical characteristics. The P removal properties were evaluated in short-term isotherm batch-experiments as well as in 12-week percolation experiments mimicking the P-loading conditions in constructed reed bed systems. The P-removal properties of sands of different geographical origin varied considerably and the suitability of the sands for use as media in constructed reed beds thus differs. The P-removal capacity of some sands would be used up after only a few months in full-scale systems, whereas that of others would persist for a much longer time. The most important characteristic of the sands determining their P-removal capacity was their Ca content. A high Ca content favours precipitation with P as sparingly soluble calcium phosphates particularly at the slightly alkaline conditions typical of domestic sewage. In situations where the wastewater to be treated is more acid, the contents of Fe and Al may be more important as the precipitation reactions with these ions are favoured at lower pH levels. The maximum P-sorption capacities estimated using the Langmuir-isotherm plots did not correspond to or correlate with the actual amount of P removed in the percolation columns. Hence, the Langmuir-isotherm does not estimate the P-removal capacities of sands. It is suggested that a suitable quick method of screening a selection of potential media for P-removal potential is to perform simple removal isotherm studies using water with a similar chemical composition as the wastewater to be treated. This method will not provide a direct estimate of the P-removal capacity that can be obtained in full-scale systems, but it is a means of comparing the relative performance of potential media. PMID- 11268837 TI - Nitrification of high strength ammonia wastewaters: comparative study of immobilisation media. AB - Due to legislative pressures, sludge production and processing in the UK will increase substantially in the future resulting in a supernatant liquid high in ammonia (500-1000 mg l-1) and "hard" COD (approximately 500 mg l-1). A small footprint reactor is required to effectively nitrify this effluent, and the aim of this work was to compare a number of immobilisation media under a variety of conditions in order to determine which media held the most promise for future development. Laboratory-scale continuously stirred tank reactors containing freely suspended and immobilised biomass were operated with a high-strength synthetic ammonia wastewater (500 mg N l-1) to determine the nitrification rates at various temperatures, and ammonia and COD loadings. COD:NH3 ratios in sludge liquors vary widely depending on the treatment processes employed, and therefore ratios of 1:1 and 2:1 were tested as being fairly typical. The freely suspended nitrifiers were washed out of the reactors at a 1 d hydraulic retention time (HRT), whereas the reactors containing adsorption particles (Linpor and Kaldnes) and PVA-encapsulated nitrifiers continued partially nitrifying down to 12 h, and oxygen addition enhanced nitrification. A decrease in temperature from 25 to 16 degrees C only caused a small (10%) decrease in nitrification in the immobilised cell reactors, demonstrating that nitrification was mass transfer rather than kinetically controlled. A reduction in nitrification occurred when glucose (500 mg l-1) was added to the feed due to the growth of a heterotrophic population. The adsorbed biomass reactors lost 35% of nitrification compared to only 7% with PVA, and it appears that the colonisation of PVA by heterotrophs is more difficult than for Linpor and Kaldnes. Respiration rates for all particles increased with time in the reactors, and nitrifiers immobilised in PVA retained approximately 40% of their viability after immobilisation. Volumetric nitrification rates were generally higher for the PVA reactor than for Linpor and Kaldnes, and were: suspended biomass reactor: 0.36; Linpor: 0.57; Kaldnes: 0.53 and PVA: 0.70 kg N m-3-reactor d-1 for a 25% reactor fill. These equate to 2.28, 4.24 and 3.97 g N m-2-media d-1 for Linpor, Kaldnes and PVA respectively, hence other reactor fill rates for Kaldnes warrant further investigation. However, the PVA particles with the highest nitrification rates under all conditions showed promise as an immobilisation medium, and are amenable to further optimisation for the nitrification of high-strength ammonia wastewaters. PMID- 11268838 TI - Inhibiting effects of chloroform on anaerobic microbial consortia as monitored by the Rantox biosensor. AB - The Rantox biosensor was designed for anaerobic wastewater treatment process control, and detects modifications of the feed based on the response of the acetoclastic methanogens contained in the sensor to periodic pulses of a concentrated organic substrate. The biosensor was tested under various operating conditions at the laboratory scale, in parallel with a digester under control fed on the same substrate. The aim was to evaluate the response of the biosensor in the presence of an incoming organic toxic compound (CHCl3). The experimental set up, i.e. the biosensor and the digester, was connected to an automated control system developed under LabVIEW environment for data acquisition and operational sequence programming (the Rantox Virtual Instrument). Biomasses with different activities were used as inocula, and inhibition was induced by dosing chloroform according to two different procedures. The results showed good sensitivity and rapid response of the biosensor to feed intoxication. The presence of chloroform was detected by the Rantox with a rapid and visible response, and well in advance with respect to the digester. PMID- 11268839 TI - Effect of tapering on the performance of waste stabilization ponds. AB - A model for wastewater degradation in a tapered waste stabilization pond was derived as a modified Bessel function by materials balance approach. Based on hypothetical data, the tapered model gave lower faecal bacteria removal than the conventional (rectangular) model for various values of dispersion number, die-off rate coefficient, average width and shape factors. The above results were corroborated by data collected from two laboratory ponds operated in parallel; one having a tapered and the other a rectangular surface area. The latter gave slightly higher hydraulic efficiency and BOD5 removal. Besides, faecal bacteria removal was significantly lower in the tapered pond than in the rectangular pond at 0.10 level of significance. Calculated faecal bacteria reduction using the tapered model was in good agreement with measured data with coefficient of correlation and standard error of 0.904 and 0.014, respectively. Effects of tapering on ponds with respect to construction cost, operational and maintenance ease and accuracy of estimated design parameters are also discussed. PMID- 11268840 TI - Stability analysis of the biodegradation of mixed wastes in a continuous bioreactor with cell recycle. AB - The stability characteristics of a continuous bioreactor with cell recycle for biodegradation of mixed wastes are investigated. The system involves a pure culture of Pseudomonas putida and media containing phenol and glucose as carbon and energy sources. The model growth kinetics for the two substitutable substrates were experimentally validated in a previous study. The stability analysis carried out using elementary principles of bifurcation theory shows rich dynamics characteristics of the reactor model, including steady-state multiplicity and hysteresis. The effect of the bioreactor operating parameters on the stability behavior of the model is discussed. Practical criteria are also derived for the safe operation of the unit and to prevent the occurrence of wash out conditions. PMID- 11268841 TI - Environmental and nutritional factors affecting geosmin synthesis by Anabaena sp. AB - A cyanobacterium isolated from a source-water reservoir during a spring odor and taste episode and identified as Anabaena sp. consistently produced geosmin during laboratory culture on modified BG-11 liquid medium. Maximal geosmin/biomass occurred at 20 degrees C and a light intensity of 17 microE/m2/s; geosmin/chla values directly correlated with increasing light intensity (r2 = 0.95, P < 0.01). It was concluded that at 20 degrees C, increasing light intensity favors less chla synthesis and higher geosmin synthesis; at 17 microE/m2/s, increasing temperature stimulates chla production (to 25 degrees C) while repressing geosmin synthesis (above 20 degrees C). Nutritional factors promoting biomass, chla, and geosmin synthesis by Anabaena sp. were also investigated. For cultures grown at 17 microE/m2/s and 20 degrees C for 20 days, both ammonium-N and nitrate-N generally enhanced the growth of Anabaena sp. Nitrate-N promoted more chla production (r2 = 0.99) than ammonium-N. Geosmin synthesis was directly correlated with ammonium-N concentrations (r2 = 0.89), with low nitrate-N (123.5 micrograms/l) favoring maximal geosmin production (2.8 micrograms/l). Increasing nitrate-N concentrations promoted a three-fold increase in chla content with geosmin synthesis decreased by two-fold. Geosmin/mg biomass was directly related to ammonium-N concentration; high nitrate-N levels suppressed geosmin production. No geosmin was detected at or below 118 micrograms phosphate-phosphorus/l. Geosmin, dry weight biomass, and chla production were correlated with increasing phosphorus (P) concentration (r2 = 0.76, 0.96 and 0.98, respectively). No geosmin was detected when copper was present in growth media at or above 6.92 micrograms Cu2+/l (CuSO4.5H2O). Dry weight biomass and chla production were negatively correlated with Cu2+ ion concentrations. PMID- 11268842 TI - The utilization of aniline, chlorinated aniline, and aniline blue as the only source of nitrogen by fungi in water. AB - The ability of fungi to degrade aniline and its derivatives in water is reported. Several fungi are able to degrade aniline and its derivatives as a sole nitrogen, carbon and energy source. Some of these fungi were obtained from activated sludge by enrichment technique. Among the 10 studied fungi, Fusarium sp. and Rhizopus sp. utilize aniline as a sole nitrogen, carbon and energy source, with production of acetanilide and catechol. Fusarium sp. utilized 70% of 10 mmol aniline and produced 3.55 mM ammonia during 30 days. Rhizopus sp. utilized 65% of 10 mmol aniline during 30 days. Rhizopus sp. and Fusarium sp. utilized only 2 chloroaniline and 3-chloroaniline as nitrogen source in the presence of glucose, with production of catechol, ammonium and chloride. The utilization of 2 chloroaniline was better than 3-chloroaniline, by Fusarium sp. and Rhizopus sp. Cladosporium sp. was the best isolate which could use aniline blue as the only source of nitrogen. This fungus reduced 89% of aniline blue, and ammonia is produced as the result of aniline blue biodegradation by Cladosporium sp. PMID- 11268843 TI - A chemometric approach to understanding the bioelimination of anionic, water soluble dyes by a biomass using empirical and semi-empirical molecular descriptors. AB - Single correlation and multiple linear regression analyses have been applied to understand the bioelimination of 103 anionic, water-soluble dyes by a biomass at a wastewater treatment works. The chemometric approach highlighted that anionic, water-soluble dyes with larger molecular size/ionic charge ratios and containing more primary aromatic amines and unsulphonated naphthalene nuclei and fewer aliphatic alcohol groups had superior levels of bioelimination. PMID- 11268844 TI - Oestrogens and oestrogenic activity in raw and treated water in Severn Trent Water. AB - Sewage effluent discharged to surface water has been shown to contain human hormones, particularly oestrogens, and synthetic chemicals which may be able to disrupt the endocrine system. Since many surface waters which receive sewage effluent are subsequently used as drinking water sources, it is important to demonstrate that treated drinking water is not contaminated. Oestrogenic activity in rivers and drinking water in the region of Severn Trent Water was studied using a combination of bioassay, to integrate exposure over time, and advanced chemical analysis. There was little or no evidence of substances that were oestrogenic, even in waters receiving significant amounts of sewage effluent. Oestrogenic activity, as measured in the rainbow trout vitellogenin assay, was seen at the Tame/Trent confluence but this activity was relatively weak. There was no activity detected at raw water intakes and no hormones or substances that are oestrogenic were detected in the final drinking water. PMID- 11268845 TI - Photocatalytic degradation of the cyanotoxin cylindrospermopsin, using titanium dioxide and UV irradiation. AB - Cylindrospermopsis raciborskii produces the cyanotoxin cylindrospermopsin, which is commonly found in SouthEast Queensland water reservoirs, and has been responsible for the closure of these reservoirs as a source of drinking water in recent times. Thus, alternative more effective treatment methods need to be investigated for the removal of toxins such as cylindrospermopsin. This study examined the effectiveness of two brands of titanium dioxide under UV photolysis for the degradation of cylindrospermopsin. Results indicate that titanium dioxide is an efficient photocatalyst for cylindrospermopsin degradation. The titanium dioxide (TiO2), brand Degussa P-25 was found to be more efficient than the alternate brand Hombikat UV-100. There was an influence from solution pH (4, 7, and 9) with both brands of titanium dioxide, with high pH resulting in the best degradation rate. Importantly, there was no adsorption of cylindrospermopsin to titanium dioxide particles as seen with other cyanotoxins, which would adversely influence the degradation rate. Degradation rates were not influenced by temperature (19-34 degrees C) when P-25 was the source of TiO2, some temperature influence was observed with UV-100. Dissolved organic carbon concentration will reduce the efficiency of titanium dioxide for cylindrospermopsin degradation, however the presence of other inorganic matter in natural waters greatly assists the photocatalytic process. With minimal potentially toxic by-product formation expected with this treatment, and the effective degradation of cylindrospermopsin, titanium dioxide UV photolysis is a promising speculative alternative water treatment method. PMID- 11268846 TI - Separation of titanium dioxide from photocatalytically treated water by cross flow microfiltration. AB - This study focuses on the separation of titanium dioxide from water by cross-flow microfiltration (CMF) within wastewater treatment by photocatalysis using slurry reactor systems. The systematic studies have shown that the separation performance of TiO2 particles is strongly affected by cross-flow velocity, transmembrane pressure, feed concentration, pH of the suspension and ionic strength. An extreme sensitivity to pH and electrolyte concentration indicates the importance of interfacial effects in solid-liquid separation of TiO2 particles. Under optimal conditions, permeate fluxes of up to 1250 l m-2 h-1, approaching those of pure water, could be obtained with a polypropylene membrane which is not sensitive to abrasion. The obtained results makes TiO2 separation by cross-flow microfiltration attractive in solar-catalytic detoxification. PMID- 11268847 TI - Modeling and Monte Carlo simulation of TCDD transport in a river. AB - A one-dimensional water quality model to assess the long-term fate of 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) in three compartments (water, sediment, fish) of a river has been developed using the literature data on various model parameters. The transient deterministic model with constant or nonrandom parameters is solved numerically by the method of orthogonal collocation, while an analytical solution is developed for the steady-state model. The impact of uncertainty in several model parameters has been studied by means of Monte Carlo simulations assuming that the uncertain parameters are uncorrelated and can be modeled by three probability distributions (uniform, normal and lognormal). For the case of a high TCDD discharge into a small, shallow river, we find that the maximum TCDD contents of water and fish are well below the prescribed safe limits. We also find that the effects of uncertainty on water quality metrics are quite complex or nonintuitive and can be substantial. This is especially true for TCDD in fish, which can be higher by as much as 50-70% than the deterministic predictions, if the parameter uncertainties follow uniform distributions. PMID- 11268848 TI - Activated sludge monitoring with combined respirometric-titrimetric measurements. AB - A short review of different respirometric methods is presented, and advantages and disadvantages of different principles are discussed. In this study a combined respirometric-titrimetric set-up was applied to monitor the degradation processes during batch experiments with activated sludge. The respirometer consists of an open aerated vessel and a closed non-aerated respiration chamber. It is operated with two oxygen probes resulting in two sources of information on the oxygen uptake rate; both collected at a high frequency. The respirometer is combined with a titrimetric unit that keeps the pH of the activated sludge sample at a constant value through the addition of acid and/or base. The cumulative amount of added acid and base serves as a complementary information source on the degradation processes. Interpretation of respirometric data resulting from validation experiments (additions of acetate and urea as ammonium source) showed that the set-up provided reliable data. Data interpretation was approached in two ways: (1) via a basic calculation procedure, in which the oxygen uptake rates were obtained by an oxygen mass balance over the respiration chamber, and (2) via a model-based procedure in which substrate transport was included for a more accurate data interpretation. Simulation examples showed that the presence of substrate transport in the model may be crucial for a correct data interpretation, since experimental conditions (e.g. low flow rate) and/or the biodegradation kinetic parameters (e.g. high Ks) may otherwise lead to data interpretation errors. Earlier studies already pointed out that titrimetric data can be related to nitrification, and this was also confirmed in this study. However, in addition, it was shown here for experiments with acetate that the amount of acid dosed was clearly related to the amount of acetate degraded. This indicates that the titrimetric data can be used to study the carbon source degradation. For the titrimetric data in this study, a model-based analysis was however only applied for the nitrification process. For an experiment with ammonium, it was illustrated that the estimation of biodegradation kinetics on a combined respirometric-titrimetric data set significantly improves confidence intervals of the parameters compared to the parameter estimation based on respirometric or titrimetric data separately. PMID- 11268849 TI - Alternative analysis of BOD removal in subsurface flow constructed wetlands employing Monod kinetics. AB - A new, mechanistic approach for design and analysis of subsurface flow (SSF) constructed wetlands is presented. The model is based on the assumption that the biological processes in wetlands, like other biological systems, exhibit Monod kinetics. A Monod approach fits well with observed wetland performance. It predicts first-order behaviour at low concentrations, that is, pollutant removal rates which increase with increasing pollutant concentration; and zero-order or saturated behaviour at high pollutant concentrations, that is, a maximum pollutant removal rate. A kinetic analysis of subsurface flow constructed wetlands exhibiting Monod kinetics reveals that loading rate, as well as the zero order degradation rate constant, are essential parameters for efficient wetlands design for the removal of organic carbon. In particular, Monod kinetics enables the identification of an absolute maximum removal rate which is necessary to prevent undersizing in design. This is significant because it represents a theoretical upper bound on loading rate for wetlands design. The analysis is applied to wetlands data collected in North America by the US EPA in order to extract design criteria for BOD removal. It reveals that maximum loadings for SSF wetlands are at least 80 kg ha-1 d-1 for BOD. In addition, a new dimensionless performance efficiency parameter, omega, is presented as a more effective means of comparing wetland performance. PMID- 11268850 TI - Determination of N-chloramines in As-samra chlorinated wastewater and their effect on the disinfection process. AB - Chlorine is the most widely used disinfectant of wastewater due to its capacity to inactivate most pathogenic microorganisms quickly. However, chlorine reacts with natural organic compounds present in wastewater to produce some undesirable organic byproducts. One such class of compounds is the nitrogenous compounds. The reaction between chlorine and compounds containing a nitrogen atom with one or more hydrogen atoms attached to it will form chloramines which have lower disinfection efficiency. Eighty percent of the wastewater generated in Jordan is treated at the Khirbet As-Samra wastewater treatment plant for eventual use in agriculture. In this study efficiency of chlorination was studied by collecting samples from the effluent of the treatment plant. The yield concentration of N chloramines upon chlorination was determined. Efficiency of disinfection process as a function of contact time, concentration of chlorine dosage, concentration of nitrogenous compound and pH were studied. In this study, it has been found that at a chlorine dosage of 15 mg/L and contact time of 15 min, the percentage total coliform kill in As-samra wastewater sample was 100%. After addition of histidine, glycine and phenylalanine (15 mg/L in each case) to the wastewater sample, the percentage of total coliform kill dropped to 58, 78 and 79% respectively. When chlorine dosage was increased to 24 mg/L the percentage total coliform kill reached 96, 99 and 100% in wastewater samples treated with 5 mg/L histidine, glycine and phenylalanine, respectively. The percentage total coliform kill dropped to zero in wastewater samples treated with histidine, glycine and phenylalanine at a concentration of 30 mg/L each. This work supports the theory that amino acids and ammonia preferentially react with chlorine to form N chloramine which significantly reduces the disinfection efficiency of the chlorination process. PMID- 11268851 TI - Biodegradability and change of physical characteristics of particles during anaerobic digestion of domestic sewage. AB - At the high-rate anaerobic treatment of domestic sewage, both biological and physical processes play an important role. Therefore, the anaerobic biodegradability of raw, paper-filtered and membrane-filtered sewage and black water has been investigated in batch experiments. Additionally, the effect of anaerobic digestion on physical characteristics, like particle size, surface tension and zeta-potential, of the present particles is studied. The biodegradability of domestic sewage and black water at 30 degrees C is almost similar (71-74%). Moreover, a high methanogenesis of the colloidal fraction in domestic sewage (86 +/- 3%) is achieved, showing that the low removal of colloidal particles in continuous high-rate anaerobic reactors is due to low physical removal rather than biodegradability. The lowest biodegradability is demonstrated for the dissolved fraction (62%). The results show that after anaerobic digestion the average radius of particles with diameter < 4.4 and < 0.45 microns increased for domestic sewage, while it decreased for black water. Part of the surface-active components in domestic sewage is not biodegraded during anaerobic batch digestion, as indicated by the development of the surface tension. The negative zeta-potential of all particles hardly changes during digestion, showing that colloidal interactions were not affected by anaerobic digestion. PMID- 11268852 TI - Microorganisms' activity and energy fluxes in Lake Varese (Italy): a field method. AB - Microorganisms are fundamental components of energy fluxes in aquatic ecosystems. Interest in plankton production and respiration has recently stimulated exploration of the use of electron transfer system (ETS) activity in oceanography and limnology. If we consider microorganism production (MP) and microorganism respiration (MR), due to aerobic and anaerobic metabolism, microorganism growth efficiency can be defined as MGE = MP/(MP + MR). In order to calculate MGE, we measured independently the two components of ETS (photosynthesis electron transfer system activity (PETS) and respiration electron transfer system activity (RETS)) during an annual cycle using a portable biosensor microorganisms amperometric detector system (MiDAS) on the site. MGE was calculated in samples collected from the photic and aphotic zones and the superficial sediment and ranged between 0.60 and 0.45 and dropped to 0.15 at the end of the summer. This substantial decrease is probably due to the prevalence of the anaerobic heterotrophic metabolism after a pronounced state of anoxia during the summer algal bloom. PMID- 11268853 TI - The application of powdered activated carbon for MIB and geosmin removal: predicting PAC doses in four raw waters. AB - Blooms of blue-green algae in reservoirs often produce the musty-earthy taste and odour algal metabolites 2-methylisoborneol (MIB) and geosmin. MIB and geosmin are not removed by conventional water treatment and their presence in the distribution system, even at low ng L-1 levels, can result in consumer complaints. Powdered activated carbon (PAC) can effectively remove MIB and geosmin when the correct dose is applied. The homogeneous surface diffusion model (HSDM) was used to predict PAC doses required to reduce MIB and geosmin concentrations to below 10 ng L-1 at four water treatment plants in Adelaide, South Australia. In jar tests, undertaken under treatment plant conditions, the predicted doses were found to produce water of the desired quality in three of the four waters. The poor predictions found in the fourth water, which had a considerably higher turbidity, were attributed to the incorporation of PAC in a larger, denser floc, leading to a reduced effective contact time of the adsorbent. It was found that higher doses of PAC were required for both compounds to produce acceptable quality water when turbidities rose above 26 NTU. PMID- 11268854 TI - Regeneration of N, P and Si near the sediment/water interface of lakes from Southwestern China Plateau. AB - Lake water, pore water, and sediments were sampled in both polluted and unpolluted lakes from Southwestern (SW) China Plateau. The results show that although the mechanisms of nutrient regeneration were similar in the polluted and unpolluted lakes, the processes, however, were much stronger in the polluted lakes. Nitrogen regeneration was mainly of organic process. Phosphorus regeneration was essentially controlled by iron redox cycling near sediment/water interface. Nutrient upward fluxes were in the order SiO2 > NH4+ > PO4(3-). This study has significance for further investigating the response of nutrient biogeochemistry to the increasing nutrient levels in aquatic environments. PMID- 11268855 TI - The role of hydroxyl radicals for the decomposition of p-hydroxy phenylacetic acid in aqueous solutions. AB - The chemical decomposition of p-hydroxyphenylacetic acid, a priority phenolic pollutant present in wastewaters from some agro-industrial plants, is studied by means of a single photochemical process produced by a polychromatic UV radiation and by hydroxyl radicals generated by the combination of UV radiation plus hydrogen peroxide and by the Fenton's reagent (hydrogen peroxide plus ferrous salts). Batch experiments were conducted to establish the degradation levels obtained and the quantum yields in the single photodecomposition process. An improvement in the decomposition of the phenolic acid in the combined UV/H2O2 oxidation is observed, due to the generation of OH radicals, and the contribution of the radical reaction to the global process is determined. In the Fenton's reagent oxidation, the effects of the operating variables (H2O2 and Fe2+ initial concentrations, pH, type of buffer used) are established and the rate constant for the reaction of p-hydroxyphenylacetic acid with OH radicals is evaluated from a kinetic model, its value being 7.02 x 10(8) M-1 s-1 at 20 degrees C. PMID- 11268856 TI - A new design of equilibrator to monitor carbon dioxide in highly dynamic and turbid environments. AB - A new design of equilibrator for carbon dioxide monitoring in natural waters is described. It consists in a vertical tube filled with marbles through which water is flowing while equilibrating with a closed air circuit. It offers several advantages compared with classical equilibrators, among which is a fast response time (half-life constant approximately 30 s) and the potential to work in very turbid water. The proposed equilibrator is of particular interest to monitor carbon dioxide in coastal ecosystems, such as estuaries, which are known to be turbid and highly dynamic. Two performance tests and some field results are presented to illustrate the efficiency of the proposed system. PMID- 11268857 TI - The potential for phytoremediation of MTBE. AB - This paper examines the potential for phytoremediation of MTBE, a gasoline additive that has become a prevalent and persistent groundwater pollutant, due to its' non-sorbing and non-reactive nature in water. A novel experimental design is developed to measure plant uptake and transpiration of MTBE from hydroponic systems, separating these processes from passive volatilization of the chemical. Plant uptake experiments indicate 30% reduction in MTBE mass in water over a 1 week period by small poplar saplings, at both high (1600 ppb) and low (300 ppb) MTBE concentrations. Active plant uptake of MTBE was approximately double that achieved by passive volatilization through a balsa wood control. MTBE was detected in biomass at the 100-ppb level, confirming passage of MTBE through the plant. A mass balance indicated that MTBE was largely untransformed during transport through the small poplar saplings to air. The high degree of MTBE removal achieved by small plants over a short period of time indicates great potential for successful phytoremediation of subsurface MTBE plumes using poplar trees. The fraction of MTBE removed from the hydroponic systems correlated well with volume of water transpired by the plants; the correlation enabled computation of the MTBE transpiration stream concentration factor of approximately 1, an important parameter for the design of engineered MTBE phytoremediation systems. PMID- 11268858 TI - Cyanobacteria diversity and toxicity in a wastewater treatment plant (Portugal). AB - Cyanobacteria are common in eutrophic natural waters. Being favoured by warm, stable and nutrient-enriched waters they may constitute an important part of the phytoplankton community in Wastewater Treatment Plants (WWTP). The phytoplankton communities of two ponds (facultative and maturation) of the WWTP of Esmoriz (North Portugal) were studied, with particular importance given to cyanobacteria. Mouse bioassays were performed with cyanobacteria samples during some of the blooms and ELISA assays specific for hepatotoxic microcystins were carried out. During the study period (January-July 1999) cyanobacteria were frequently dominant in the ponds ranging from 15.2 to 99.8% of the total phytoplankton density. The main species were Planktothrix mougeotii, Microcystis aeruginosa and Pseudanabaena mucicola. Mouse bioassays were performed during Oscillatoria bloom period but the results were negative, in spite of the high cyanobacteria biomass. ELISA assays were performed for both ponds but only in the maturation pond positive values were found. Microcystin concentrations (as MCYST-LR equivalents) varied from 2.3 to 56.0 micrograms/l on the margin of the pond and between 1.7 and 4.6 micrograms/l in the outflow of this pond. These values indicate that WWTP may be a source of contamination of water bodies with cyanobacteria toxins. PMID- 11268859 TI - Novel cake characteristics of waste-activated sludge. AB - Breaking down the time limit constraints for conventional compression-permeation (C-P) cell test, this work has, for the first time, experimentally evaluated the cake characteristics of viable waste-activated sludge subject to polyelectrolyte flocculation and to freeze/thaw treatment under a pressure range of 25-200 kPa. There exists a threshold pressure exceeding which the cake structure would significantly deteriorate. Also, the present biological sludge is a "super compactible" sludge, whose compactibility is greater than most data ever reported in open literature. The information presented herein has implications to filter design/operation and can be used as a reference data set for examining the existing filtration theories. PMID- 11268860 TI - Comments on "Development of an improved synthetic sludge: a possible surrogate for studying activated sludge dewatering characteristics". Banu Ormeci and P. Aarne Vesilind (2000) Wat. Res. 34, 1069-1078. PMID- 11268861 TI - Comment on "Theories of cake filtration and consolidation and implications to sludge dewatering". PMID- 11268862 TI - Maternal worry about neonatal hearing screening. AB - OBJECTIVE: To identify and compare the prevalence and degree of maternal worry about neonatal hearing screening at the time of an initial neonatal hearing screen and rescreen in 1997 and 1999. STUDY DESIGN: We report on a prospective cross-sectional investigation of maternal worry about newborn hearing screening. Demographic data, maternal knowledge of hearing screening, and degree of maternal worry were collected on 307 mothers at the time of the neonatal screen and 40 mothers at the time of the rescreen. RESULTS: Degree of maternal worry was significantly greater at the rescreen compared to the screen. Mothers who reported greater worry at the time of the screen were more likely to be socioeconomically disadvantaged. Although maternal knowledge about hearing screening increased between the two time periods, degree of worry remained unchanged. CONCLUSION: Efforts to minimize the neonatal false-positive hearing screen rates and to educate mothers about hearing screening are indicated to minimize unnecessary worry. PMID- 11268863 TI - The effect of anemia on retinopathy of prematurity in extremely low birth weight infants. AB - OBJECTIVE: Numerous risk factors for development of retinopathy of prematurity (ROP) in very low birth weight infants have been identified in the literature. However, the role of anemia in the development of ROP has not been adequately addressed. STUDY DESIGN: We retrospectively examined the medical records of all infants weighing < or = 800 g who were admitted to a university hospital between July 1, 1992 and December 30, 1997. Highest and lowest hemoglobin and hematocrit values and the number of blood transfusions were recorded at each week of life during hospitalization. Gestational age at birth, birth weight, race, sex, oxygen status, history of bronchopulmonary dysplasia, length of hospital stay, and sepsis were also identified as potential risk factors. Data were analyzed using logistic regression to adjust for these confounding variables. RESULTS: Infants were grouped according to ROP status in the following manner: stage 0 to 1 ROP, stage 2 ROP, and stage 3 to threshold ROP. Sex, gestational age at birth, bronchopulmonary dysplasia, ventilator days, length of hospital stay, and number of blood transfusions were significantly associated with severity of ROP by univariate analysis. Using a logistic regression model, only gestational age (p = 0.007) and number of blood transfusions (p = 0.04) remained statistically significant. CONCLUSIONS: Anemia did not affect severity of ROP as an independent risk factor. However, the number of blood transfusions did affect the highest stage of ROP in this group of premature infants. Infants who remained severely anemic (Hgb < or = 8 g/dl or Hct < or = 25%) for longer periods of time developed milder ROP than less anemic infants. PMID- 11268864 TI - Delivery room resuscitation decisions for extremely low birthweight infants in California. AB - OBJECTIVE: To characterize physician-parent counseling and delivery room resuscitation of extremely low birthweight (ELBW) infants. STUDY DESIGN: Cross sectional survey of 473 California neonatologists detailing counseling patterns, resuscitation thresholds, and acceptance of parental decision making. RESULTS: The response rate was 61%. After 23 weeks' gestation, > 80% of neonatologists counseled parents expecting ELBW infants. All (> 99%) counseled parents about mortality; > 25% reported not discussing limiting resuscitation or death despite resuscitation. Decisions to limit resuscitation were affected by congenital anomalies, parents' wishes, or perceptions of pain, suffering, and quality of life. Nearly 70% of neonatologists supported parental decision making at 22 to 23 weeks, whereas 66% to 74% responded that parents should not be allowed to make nonresuscitation decisions after 26 weeks. Median resuscitation thresholds were 23 weeks (range 20-28) and 500 g (range 350-1000). CONCLUSIONS: Neonatologists' failure to discuss nonresuscitation options, variations in resuscitation thresholds, and unwillingness to accept nonresuscitation decisions for more mature ELBW infants may restrict parental decision making. PMID- 11268865 TI - The association of coagulase-negative staphylococci isolated from the chorioamnion at delivery and subsequent development of cerebral palsy. AB - OBJECTIVE: To find out whether there is an association between cultures positive for coagulase negative staphylococci (CONS) taken from babies in the Neonatal Intensive Care Unit (NICU) and a subsequent outcome of cerebral palsy. STUDY DESIGN: At delivery, we obtained cultures from the chorioamnion space and, when medically indicated, we obtained bacterial cultures from children in the NICU. Surviving neonates underwent final examination for cerebral palsy at age 18 months. RESULTS: Of six children in the Magnesium and Neurologic Endpoints Trial who had cerebral palsy, chorioamnion cultures had been obtained for five of six. Four of these five children (80%) had CONS-positive cultures, whereas 26 of 102 (25%) children without cerebral palsy were CONS positive (p = 0.02). In the NICU, of children with cerebral palsy, the prevalence of culture-proven CONS was 80% (4/5); for those without cerebral palsy, the prevalence was 17% (15/86) (p = 0.01). Using multivariable logistic regression to control for confounding, CONS in the chorioamnion remained significant (adjusted odds ratio [OR] 37.7, 95% confidence interval [CI] 3.0 to +infinity; p = 0.003). However, when controlled for extremely low birth weight, nonvertex presentation, and being on a ventilator > or = 20 days, the association between culture-proven CONS in the NICU and cerebral palsy became insignificant (adjusted OR 3.0, 95% CI 0.2 to +infinity; p = 0.42). CONCLUSION: CONS in the chorioamnion space are associated with cerebral palsy, but in these data, CONS in the NICU are not found to be associated with cerebral palsy. PMID- 11268866 TI - The changing face of medicine: health care on the Internet. PMID- 11268867 TI - Hyperbilirubinemia and early discharge. PMID- 11268868 TI - Neuroimaging and the timing of fetal and neonatal brain injury. AB - Current and advanced structural and functional neuroimaging techniques are presented along with guidelines for utilization and principles of imaging diagnosis in fetal and neonatal central nervous system abnormalities. Pattern of injury, timing issues, and differential diagnosis are addressed with emphasis on neurovascular disease. Ultrasonography and computed tomography provide relatively rapid and important screening information regarding gross macrostructural abnormalities. However, current and advanced MRI techniques often provide more definitive macrostructural, microstructural, and functional imaging information. PMID- 11268869 TI - One-sided high-frequency oscillatory ventilation in the management of an acquired neonatal lobar emphysema: a case report and review. AB - We describe a premature infant (gestational age 28 weeks and birth weight 1280 g) with a left-sided acquired lobar emphysema (ALPE). Left lateral decubitus positioning, right-sided conventional ventilation (CV), tracheal high-frequency oscillatory ventilation (HFOV), and dexamethasone administration were subsequently used in the treatment without success. The emphysema was resolved and the patient was extubated after selective intubation and HFOV of the right unaffected lung. We also review the reported cases of ALPE in neonates that were treated by one-sided high-frequency ventilation (HFV). PMID- 11268870 TI - Survival of a preterm twin following infarction of the right hepatic lobe. AB - Survival following hemorrhagic necrosis of a lobe of the liver in preterm infants is very uncommon. We present the clinical and radiologic findings of such a case, discuss the clinical management of the hemorrhage that resulted in the infant's survival, and present a basis for the evolution of the liver pathology. PMID- 11268872 TI - Placental pathology casebook. Complete hydatidiform mole with coexistent term twin pregnancy. AB - A case of twin pregnancy consisting of a complete hydatidiform mole with a coexistent, viable fetus is presented. The case is distinctive for its presentation on ultrasound, its unusually low levels of serum hCG, its remarkable histology, and its term delivery. PMID- 11268871 TI - Case report: total parenteral nutrition extravasation associated with spinal cord compression and necrosis. AB - A preterm infant, whose course was complicated by sepsis, necrotizing enterocolitis with jejunal perforation, intraventricular hemorrhage and cerebellar hemorrhage, suffered permanent and total paralysis below the neck from extravasation of parenteral nutrition fluids through a femoral venous catheter. MRI imaging revealed extravasation of fluid into the paraspinus musculature with extension into the spinal canal. This fluid was identified as hyperalimentation and intralipid. Postmortem examination found evidence of necrosis of the spinal cord as well as perforation of the right iliac vein. PMID- 11268873 TI - Fetal heart rate monitoring casebook. Nonreassuring fetal heart rate, attempted instrumental delivery, forceps rotation, cord prolapse during rotation maneuver, and rescue by extreme emergency abdominal delivery. AB - In the course of an attempted instrumental delivery, prolapse of a pulseless umbilical cord occurred, concomitant with total collapse of the fetal heart rate pattern. Rescue was by extreme emergency abdominal delivery. PMID- 11268874 TI - Early-onset neonatal group B streptococcal sepsis: intrapartum antibiotic prophylaxis in the clinical setting. AB - OBJECTIVE: To evaluate how guidelines for the use of intrapartum antibiotics for the prevention of early-onset Group B streptococcal infection are utilized in a clinical setting. STUDY DESIGN: Review of maternal/infant records for the year 1993 in a perinatal center. RESULTS: Intrapartum antibiotics were administered to 77.8% of 443 Group B streptococcus (GBS)-colonized women. There were 452 infants born to these mothers, of which four developed GBS infection. During the same period, an additional 11 infants with GBS infection were born to women with "negative" or "unknown" GBS status (the women did not receive intrapartum antibiotics). Infants of GBS-colonized women who had not receive antibiotics were more likely to develop infection than GBS negative or unknown status, odds ratio 9.0, 95% confidence interval (2.8-29.1). CONCLUSION: This study supports the use of intrapartum antibiotics as an important means of preventing early-onset neonatal GBS infection but demonstrates problems that may be encountered in the clinical application of guidelines for intrapartum antibiotic prophylaxis. PMID- 11268875 TI - [Rapid reviews for evidence-based decision support. (Restricting) requirements]. AB - Since evidence-based decision making is increasingly gaining acceptance among German health care authorities the need for evidence-based support for time critical decisions becomes obvious. The best method to identify and summarize the available evidence is the conduct of systematic reviews of the literature. However, generally systematic reviews are not applicable in matters of urgency, because of their time consuming preparation. Concepts for "rapid reviews" are urgently required which ensure that these reviews still produce reliable results. The article proposes strategies to accelerate the preparation of a systematic review and discusses their possible influences on the conclusions of the review. Research efforts are needed to clarify which of the accelerations actually yield valid results. PMID- 11268876 TI - [Evidence-based scientific analysis of fundamental Medical Services of Statutory Health Insurance internal principle assessment]. AB - In Germany, the increasing relevance of Evidence-based medicine (EbM) is not only a consequence of growing economic limitations in the health care system but of a changed jurisdiction, too: liability of the statutory health insurances (SHI) to pay for medical care depends on the proof of its effectiveness. This is of special importance for the Medical Services in their role as an advisor of the SHI. The article depicts the basic assignments of the Medical Services of the Statutory Health Insurance, their legal frame and the role of EbM in sociomedical expertising. The way of fundamental sociomedical expertising, its internal and external effects, the personnel and technical/logistic requirements are described as well as potential of improvement. PMID- 11268877 TI - [The importance of guidelines in the area of malpractice procedure]. AB - Generally available national guidelines are the only way to guarantee that the clinically active doctor will consider the same rules as the doctor as an expert within a malpractice procedure. Especially the following question has to be ruled out by national guidelines: From what time on is an innovation to be considered as "normal"--as "standard"? The same is true for continued expert discussions and for the "medical-economic balance". Without national guidelines recognized as well-known rules, the doctor accused in a malpractice procedure will find himself in an unacceptable position of legal uncertainity. PMID- 11268878 TI - [Quality project in invasive/interventional cardiology]. AB - With installing a new catheterization laboratory, a quality project with focus on indications for invasive/interventional procedures was implemented. Health insurance companies as budget holders were involved in the project, external control is accomplished by their medical service (MDK). The focus on indications is new, since most approaches in this area deal with structure and/or process quality. The actual concept of this quality project makes medical performance transparent with regard to adequate indication as the first and important step to excellent quality of results. Further, the concept contains a rational approach to the controversial discussion about the increasing frequency of catheter-based coronary interventions. PMID- 11268879 TI - [Patients' involvement in and and their demands on evidence-based medicine]. AB - Representatives of evidence-based medicine have repeatedly declared their goal of integrating patients and their demands on the health care system in discussions and networking forums. If one takes various indicators of consumer demands into consideration--like, for example, the Berliner Mangelliste (a list of inadequacies and deficiencies of the Public Health Care System compiled by members of the Berliner Selbsthilfe-Forum chronisch kranker und behinderter Menschen (a platform of discussion for chronically ill or handicapped participants in self-help groups) or, for example, the results of a study which researched consumer demands on information in decision-making situations, then a number of mutual fields of interest between patients and evidence-based medicine professionals can indeed be found. On the other hand there is a series of issues which patients repeatedly articulate, which are not among those most important to evidence-based medicine. Such issues include, for example, more personal attention, acceptance, holistic treatment methods, and the elimination of deficits in the quality of the Health Care System. To successfully involve patients in the networking process in a meaningful and permanent way and to make their involvement in the activities of evidence-based medicine more than a rhetorical aim, the discourse should not exclusively correspond to specialised interests of professionals, but rather should take up the issues given priority to by patients and should consider their working conditions, their stress-factors and their often limited resources. If it is seriously intended that patients and members of self-help groups take an active interest in promoting those parts of evidence based medicine helpful and efficient for them, then more effort must be made, on the one hand, to motivate them and to provide more comprehensive information and, on the other hand, patients and their initiatives must receive more support. PMID- 11268880 TI - [The LORAS Project and quality assurance. In four years from input- to outcome oriented financing in public health. 1: The LORAS Project]. AB - This series of three articles is a summary of the operations, findings and results of the hospital reform projects in the Canton of Zurich, termed LORAS. With the aid of the LORAS project within four years Zurich hospitals have been transformed. Whereas they used to adhere to input-oriented covering of deficits they now operate with outcome-oriented prospective financing of output. Part 1 describes the whole Project. Part 2 focuses on the development of outcome measurement. Part 3 finally describes the implementation of the outcome measurement in the canton of Zurich. PMID- 11268881 TI - [The DEGAM guidelines "Dysuria" by the German Society of General Practice and Family Medicine (DEGAM)--possible consequences of the implementation in general practice]. AB - INTRODUCTION: In Germany, there are hardly any reliable data on patient care in the primary care setting which warrant the development and implementation of clinical guidelines. In this paper, data generated by a prospective observational study of patients with urinary tract symptoms are compared to the recommendations of an evidence-based clinical guideline. PATIENTS AND METHODOLOGY: Over a period of 6 months all patients consulting one of 6 General Practitioners in southern Germany with symptoms of dysuria have been documented on a standardised patient record. Data were compared to the recommendations of the guideline "Dysuria" by the German Society of General Practice and Family Medicine (DEGAM) to assess the relevance and feasibility of the guideline. In a scenario, compliance with the guideline is extrapolated to the realm of primary care. RESULTS: Basic demographic and epidemiological data agree with basic assumptions of the guideline. As far as diagnostic and therapeutic strategies are concerned there are significant discrepancies between the recommendations and the realm of primary care. Microbiologic cultures are ordered far less then recommended, second line drugs are prescribed far more often then recommended, macroscopic urinoscopy is performed widely but not covered by the guideline at all. If GPs complied completely with the guideline, many more diagnostic procedures would be performed and a different palette of antimicrobial drugs would be prescribed. CONCLUSION AND OUTLOOK: The "Dysuria-Guideline" of DEGAM was developed for a prevalent and relevant topic in primary care in Germany. There are significant discrepancies between the recommendations and the realm of primary care. Post-hoc analysis is an informative and feasible tool to identify potential obstacles against implementation of guidelines. PMID- 11268882 TI - [The medical service medical quality assurance]. PMID- 11268883 TI - [Evidence-based medicine in the further development of health care systems. Comments on a current discussion]. AB - The concept of evidence-based medicine is applicable also in the field of guiding health care systems to overcome deficits in adequate patient care. EbM has already been used in this field and has passed first tests of its practical application. There are some obstacles against the future esteem of EbM. These include some misunderstanding concerning the concept itself, missing routine where external evidence has to be put into practice, and some general resistance. Details are presented and an adequate usage of EbM is argued for. PMID- 11268884 TI - [How much is "evidence-based"? An overview of the state of the art in research]. AB - The proportion of evidence-based health care gives rise to controversy. Estimates range from 4%-20%. Studies trying to ascertain the proportion of evidence-based healthcare show wide variations in the results depending on design, setting, subject, main outcome measures, methodology and definition of evidence from 11 82%. The results should not be generalized to community health care and should not be misused in public discussions. PMID- 11268885 TI - [Beta blockers in asthma and COPD--a therapeutic dilemma?]. PMID- 11268886 TI - [Risk of radiation exposure in X-ray examination of the thorax. German Central Committee for the Control of Tuberculosis (DZK)]. PMID- 11268887 TI - [Lung function reference data in school-age children]. AB - The practical interpretation of lung function data depends to a very great extent on the quality of reference values. METHODS: From a population of n = 2615 schoolchildren between 6 and 12 years of age 15,404 lung function measurements were taken in accordance with commonly accepted guidelines. In the present study the validity of reference equations from the literature is examined with regard to our measured data. RESULTS: Employing linear regression analysis, the natural logarithm (ln) of forced vital capacity in ml (FVC) and expiratory volume in one second in ml (FEV1) were explained on the basis of the equation ln y = a + b* ln x (y = FVC, FEV1 (ml); x = height (cm)). Analyses were performed for girls (lnFVC = -4.8789 + 2.5504* lnheight, lnFEV1 = -4.3078 + 2.4070* lnheight) and boys (lnFVC = -4.5241 + 2.4917* lnheight, lnFEV1 = -3.7338 + 2.2985* lnheight), separately. Our coefficients correspond best to the literature dealing with a population of the same age group. On the other hand, for example, if reference values are derived from equations from the literature for the age group 6 to 18 years, they result at 6 years in an underestimation of the volume measured by us (FVC -150 ml) and at 12 years to an overestimation (FVC + 120 ml). CONCLUSIONS: The extent of the proven systematic deviations of the reference values from the measured values is of clinical and epidemiological relevance. To avoid misinterpretations, special reference values should be applied for preadolescents, at least with regard to FVC and FEV1. PMID- 11268888 TI - [Congenital cystic adenomatoid malformation--clinical spectrum and management]. AB - Congenital cystic adenomatoid malformation (CCAM) is one of the most frequent dysplasias of the lung. Diagnosis is often suspected in utero and urges obstetricians, pediatricians, and pediatric surgeons to make appropriate management decisions as to an optimal management for the affected patients. We report on a preterm baby with a gestational age of 33 weeks, suffering from hydrops fetalis and postnatal respiratory distress syndrome, a two-year old boy with clinical signs of a foreign body aspiration, and a seven-year old boy with a funnel chest. In each case, surgical resection was performed, the histology revealing CCAM. Our case report describes the broad clinical spectrum of CCAM. An algorithm is presented, helping to make diagnostic and therapeutic decisions. PMID- 11268889 TI - [Evaluation of a structured education program for adult outpatient asthmatics]. AB - BACKGROUND AND METHODS: The efficacy of a structured education programme (AFAS) unter outpatient conditions was evaluated in a pilot study including 25 mild to severely ill adult asthmatics (age 41 +/- 2 yrs.) over a period of two and a half years. The main teaching items of the programme are: self-control of the disease with regular peak flow measurements, monitoring of symptoms with a patient diary, effects and side effects of the treatment, correct inhalation technique of asthma medication and the ability of self-management with regard to the actual degree of airflow limitation by the patients. RESULTS: After AFAS the knowledge of the patients regarding the disease as well as the medication increased significantly. There was an improvement of drug therapy: before AFAS only 52% of the patients used inhaled steroids on a regular basis with regard to 96% one and two years after participation of AFAS (p < 0.01). The self-control of the disease was improved: before AFAS no patient measured peak flow during acute dyspnoea, compared with 88% (p < 0.001) and 75% (p < 0.001) one and two years after AFAS, respectively. The number of severe asthma-attacks decreased significantly from 10.7 +/- 2.5 per patient and year before education to 1.3 +/- 0.2 (p < 0.001) after the first year and to 2.0 +/- 0.3 (p < 0.05) after the second year. The total number of hospital days due to asthma decreased from 219 days in the year before the participation in AFAS to zero (p < 0.001) in the first year after the education and to 17 days (p < 0.001) after the second year. CONCLUSIONS: The efficacy of patient education with AFAS is still evident two years after the course, but a reduction of self-control of the disease was observed during the follow-up period. In conclusion, structured education programmes for adult asthmatics can be effective even under outpatient conditions. PMID- 11268890 TI - [The significance of health related quality of life for the evaluation of interventional measures in patients with COPD]. AB - Health related quality of life (HRQOL) is an important criterion for the evaluation of rehabilitation measures in patients with chronic obstructive pulmonary disease (COPD). The present paper reviews the current literature about the effects of pulmonary rehabilitation on the HRQOL of patients with COPD. The aim is to summarize critically methods, results and unanswered issues of the present research on the effects of pulmonary rehabilitation on HRQOL. The rehabilitation of patients suffering of COPD is mainly based on six types of interventions: 1. long-term oxygen therapy (LTO), 2. pharmacological management, 3. surgical therapy (bilateral reduction of lung volume), 4. physical therapy, 5. nutritional therapy (special diets), and 6. psycho-social interventions (e.g. psychotherapy, training and education). Thirty-one studies could be included in which HRQOL served as an outcome criterion for the rehabilitation of COPD patients. In 14 (45%) studies exclusively a disease-specific measure for the assessment of HRQOL was employed, while in 12 (39%) studies a generic instrument was applied. In the remaining five (16%) studies two ore more measures were used, whereas four of them combined a generic and a disease-specific method. The St. Georges Respiratory Questionnaire (SGRQ) und the Chronic Respiratory Disease Questionnaire (CRDQ) belonged to the group of the specific instruments, while among the generic measures the Sickness Impact Profile (SIP), the Nottingham Health Profile (NHP), the SF-36 and the Quality of Well-Being Scale (QWB) were most frequently used in COPD patients. The surgical bilateral reduction of lung volume, pharmacological therapy, upper extremities muscle training and psychological measures as single interventions proved to have persistent positive effects on the HRQOL. Several rehabilitation programs, composed of a wide variety of different interventions were effective in terms of HRQOL. On the other hand, at follow-up, the short-term positive effects had decreased in two of the three studies, where the rehabilitation took place exclusively in an inpatient setting. However, in three of four programs implemented in an outpatient setting, a persistent positive effect on HRQOL could be demonstrated. In conclusion from the as of yet available findings, we suggest for future studies to use only such measures of HRQOL which have been tested psychometrically in patients with COPD and to combine disease-specific and generic measures. In order to achieve lasting positive effects of rehabilitation on HRQOL, outpatient settings or ambulatory refreshment sessions following rehabilitation on an inpatient basis should be preferred. PMID- 11268891 TI - [Superficial pyoderma requiring oral antibiotic therapy: fusidic acid versus pristinamycin]]. AB - OBJECTIVE: This study was aimed to compare the clinical and antibacterial efficacy of fusidic acid 500 mg twice a day, per os, over 7.5 days) to pristinamycin 1 g twice a day, per os, over 10 days). METHODS: Patients aged over 18, suffering from a superficial pyoderma requiring antibiotherapy and having given their informed consent were enrolled in a controlled, multicentre, double blind double dummy, parallel groups study. From day 0 to day 10, the patients received the randomised treatment. Those who were cured at day 11 had a visit at day 25 without any treatment between day 11 and day 25. A swab was performed on days 0, 11 and 25. The two treatment groups were compared in terms of efficacy, safety and global cost. RESULTS: 334 patients seen in dermatologic consultation were included in the study. 313 patients were analysed on an intent-to-treat basis. 158 received fusidic acid (FA) and 155 were treated with pristinamycin (P). At D11, 126 patients were cured in the FA group (79.7%) and 118 in the P group (76.1%) (p = 0.44). The bacteriological success rate was 85.2% in the FA group and 82.7 in the P group (p = 0.67). The recovery was confirmed in 92.6% of the FA patients and 90.4% of the P patients at D25 (p = 0.56). Digestive tolerance was better with fusidic acid than with pristinamycin. In economic terms, fusidic acid was cheaper than pristinamycin: 443 French francs in the FA group versus 545 FF in the P group. CONCLUSION: Therefore we conclude that an oral course of 7.5 days with fusidic acid is an efficient and cheaper alternative to a treatment with pristinamycin over 10 days. PMID- 11268892 TI - [De La Chapelle syndrome]. AB - OBJECTIVE: The De La Chapelle syndrome (XX male) is a peripheral hypogonadism concerning males with 46,XX karyotype. We conducted a retrospective study of 18 cases and report the main clinical biological and hormonal characteristics. PATIENTS AND METHODS: Clinical features (weight, height, aspect of the external genital organs, body hair, gynecomastia), hormone levels (testosterone, gonadotrophin, baseline and stimulated prolactin estradiol), and results of a Barr test and karyotype were recorded in all patients in addition to search for the SRY gene (in 8 of the 18 patients). Findings were compared with a matched male population and a Klinefelter syndrome population. RESULTS: Microrchidia was found in almost all the patients while the penis had a normal size. Signs of hypoandrogenism were frequent and gynecomastia was present in half the cases. De La Chapelle patients differed from Klinefelter patients by the absence of dysmorphism. DISCUSSION: Patients with De La Chapelle syndrome diagnosed around the age of 20 years do not have borderline disorders associating genitalia anomalies or sexual ambiguity. The majority of the patients bear the testis determining SRY gene on one of the X chromosomes, providing the rational explanation of the male phenotype, but 20% of the XX males doe not have this gene. The role of certain key genes that could be implicated in abnormal sexual differentiation is known, but the complexity and heterogeneous nature of this syndrome leaves many questions unanswered. Therapy is based on androgen replacement therapy given at an early stage. PMID- 11268894 TI - [Late reversal leprous reaction appearing 18 months after the termination of treatment]. PMID- 11268893 TI - [Fainting after drinking a glass of whiskey-soda...]. AB - BACKGROUND: Alcohol intolerance is a rare syndrome that usually involves pain, pruritus, or vasomotor phenomena consecutive to moderate alcohol intake. This syndrome may unmask various malignancies, especially Hodgkin's disease or non Hodgkin lymphoma, and carcinoma of the cervix. CASE REPORT: A patient who became unconscious after drinking a glass of whiskey-cola was explored for alcohol intolerance. High-level hypereosinophilia was discovered. The diagnosis of idiopathic hypereosinophilia was retained after extensive evaluation. DISCUSSION: Idiopathic hypereosinophilia may be a premalignancy expression of a lymphoproliferative disorder. In our patient, who also had alcohol intolerance known to be associated with later diagnosis of malignancy, lymphocyte phenotype studies revealed an abnormal population. However, rearrangement studies of T-cell and B-cell receptors, as well as bone marrow cytology and histology did not enable identification of a lymphoproliferative disorder. This patient will require close follow-up due to the risk of developing a lymphoproliferative disorder. PMID- 11268895 TI - [Heparin-induced priapism]. PMID- 11268896 TI - [Scurvy revealed by hemarthrosis]. PMID- 11268897 TI - [Parkinson disease: diagnostic and therapeutic criteria]. PMID- 11268899 TI - [Where's the obstacle?]. PMID- 11268898 TI - [Parkinson's disease]. PMID- 11268900 TI - [Neurosurgery and pituitary tumors: etio-pathogenic considerations]. AB - PITUITARY ADENOMA: Based on the experience of nearly 5000 cases of surgically treated pituitary tumors at the neurosurgery department of the Foch Hospital, the pituitary adenoma is the most frequent pituitary tumor. Secreting tumors lead to a clinical syndrome depending on the level of hormone overproduction. Gonadotrop or non-functioning pituitary adenomas are mainly macroadenomas presenting with visual symptoms, hypopituitarism or as an incidentaloma. Anatomical features dictate the surgical approach. OTHER TUMORS: The other types of hypophyseal tumors, such as craniopharyngioma, Rathke's cleft cyst or others are usually surgical tumors because medical treatment is ineffective. Malignant pituitary tumors are unusual. PMID- 11268901 TI - [Neurosurgery and pituitary tumors: from preoperative tests to postoperative follow-up]. AB - DIAGNOSIS: Accurate diagnosis of a pituitary tumor requires coordinated medical collaboration. Guidance learned from experience is essential for MRI exploration of pituitary tumors, especially microadenomas. Therapeutic strategy depends largely on the accurate examination of the radiological data that guides the surgical approach and determines, for the patient and family, the possibility of tumor and/or endocrinological cure. PITUITARY FUNCTION: Secreting tumors are diagnosed by assessment of hypophyseal function, predictive of medical treatment efficacy and the value of hormone replacement therapy in case of surgery. PMID- 11268903 TI - [HIV: a guide on management of seropositive patients]. PMID- 11268902 TI - [Neurosurgery and pituitary tumors: surgical indications and outcome]. AB - TWO MAIN FEATURE: Indication for surgery in patients with pituitary tumors depends first on the anatomical situation: the enclosed or invasive nature of the tumor. Total resection of an enclosed tumor, even if it is a huge one, can be expected to be successful. For invasive tumors, surgery will be subtotal unless the invasion is very limited. The second consideration is the efficacy and limitations of medical treatment. At present, only secreting pituitary adenomas are accessible to medical therapy. Other pituitary tumors, and non-functioning pituitary adenomas are not suitable for valid medical treatment and may warrant a surgical strategy. CHOICE OF THE OPERATIVE APPROACH: Anatomical and radiological considerations are determining. For secreting pituitary adenomas, first intention surgery via a transphenoidal access is advocated when surgery can be expected to achieve complete tumor resection without damaging the normal gland. For other cases, medical treatment has to be instituted prior to surgery which will be discussed in case of failure, intolerance or for tumor reduction. Transphenoidal surgery is strongly advocated in case of a pituitary incidentaloma with a visual danger, even in old patients. OUTCOME: Visual symptoms are often improved after neurosurgery for pituitary tumors. Hormone cure is frequent in microsecreting pituitary adenomas, rarely in invasive tumors. PMID- 11268904 TI - [Boerhaave's syndrome: also in the emergency room]. AB - Two men, aged 52 and 57 years, had vomited and then developed chest pain, dyspnoea and tachypnoea. After a myocardial infarction had been excluded in the cardiac emergency room, further examination revealed a rupture of the oesophagus. This was treated surgically with the ultimate creation of a tubular stomach. Both patients then recovered well. The Boerhaave's syndrome, a 'spontaneous' perforation of the oesophagus, is a rare and potentially lethal condition which should be diagnosed at an early stage. Pain in the chest, dyspnoea and vomiting are frequent symptoms. A cardiac cause is sometimes erroneously suspected. Subcutaneous emphysema is a major indication for a perforation of the oesophagus. The chest X-ray shows also mediastinal emphysema and infiltrative abnormalities; in case of doubt a second X-ray should be made some hours later. PMID- 11268905 TI - [Destruction of medical records in psychiatry; a disturbing development]. AB - The Netherlands have the 'Wet op de Geneeskundige Behandelingsovereenkomst', a law concerning the medical treatment agreement with the patient. Among other things it regulates the duties regarding the keeping and destruction of medical records. A critical evaluation is necessary, because the existing regulation is expected to have a negative effect on patient care and on scientific research. Development of better criteria for destruction of medical records is necessary. Until then, the destruction of medical records should be stopped. PMID- 11268906 TI - [Save or destroy? The importance of medical record for former patients and future patients]. AB - In 1995, a new privacy law was introduced in the Netherlands. According to this law, medical records should be saved for 10 years, and then destroyed, unless keeping the records for a longer period follows reasonably from the duties of the treating physician (as is the case, for example, when treating patients with a chronic disease). There are serious concerns with regard to the future availability of medical record data for clinical research and patient care after 2005. Evaluation of the late effects of many medical treatments will no longer be possible in the Netherlands. Patient care, particularly genetic counselling, will be also seriously compromised. As a possible solution the profession might name diagnoses and treatments regarding for which, from the point of view of good care, it is necessary for files to be kept for longer than 10 years. For a uniform nationwide policy it would be better if all files, perhaps after sorting by diagnosis and treatment, should be obligatorily kept for much longer than 10 years, preferably for the duration of the life expectancy. PMID- 11268907 TI - [Determination of cardiac troponins for diagnosis of 'acute myocardial infarct']. AB - The diagnosis of 'myocardial infarction' was recently redefined by a commission of European and American cardiologists. The main element of the new definition is a raised serum level of heart-specific troponin (T or I). In healthy adults virtually no cardiac troponin is demonstrable, so that every rise of the level of the heart-specific troponin in the blood means that there is myocardial damage. An infarction during a coronary bypass operation and myocardial damage during skeletomuscular injury can be diagnosed almost faultlessly by a troponin assay. A raised troponin level in acute coronary ischaemia is associated with a raised mortality risk. However, even when the level is normal, a risk of cardiac complications is present. For the diagnosis of a recurrent infarction during a developing infarction, determination of the peak level of the creatine kinase muscle brain mass (CK-MBmass) is most appropriate. Also, the value of including troponin in the existing rule-out protocols has not yet been proven. Now that most Dutch hospitals shortly will be capable of troponin assays, cardiologists and clinical chemists should consider the implementation of this assay in clinical practice. PMID- 11268908 TI - [Development and developmental disorders of the human brain. III. Neuronal migration disorders of the cerebrum]. AB - Neuronal migration disorders of the cerebral cortex form a heterogeneous group of abnormalities, characterised by mental retardation, epilepsy and hypotonia. They are prevalent in 1% of the population and in 20-40% of the untreatable forms of epilepsy. Disorders at the start of the migration result in nodular heterotopias. Bilateral periventricular nodular heterotopias are X-linked disorders, in which cortical neurons are unable to leave their position at the ventricular surface due to the absence of filamin 1. The large group of lissencephalies can be divided into a number of syndromes, each of which is characterised by a gene mutation (LIS1, DCX, RELN). These mutations result in agyria and pachygyria, which are characteristic for this group. A number of these abnormalities, especially the smaller nodular heterotopias and focal cortical dysplasia, may be treated by neurosurgical excision. PMID- 11268909 TI - [From gene to disease; fragile X-syndrome: hereditary mental retardation due to a developmental gene]. AB - The fragile X syndrome is the most common cause of familial intellectual disability. The identification of the 'fragile-X mental retardation' (FMR1) gene disclosed a novel genetic mechanism: an intergenerational instable cytosine guanine-guanine (CGG) repeat leading to the absence of FMR1 protein above a threshold of 200 repeats and, subsequently, leading to familial mental retardation. PMID- 11268910 TI - [Diagnostic image (28). Hydatidiform mole]. AB - A 23-year-old woman suffered from vaginal bleeding during pregnancy caused by a mola hydratidosa. PMID- 11268911 TI - [Magnetic resonance cholangiopancreatography: sensitive and specific diagnostic method for suspected choledolithiasis]. AB - OBJECTIVE: To evaluate the role of magnetic resonance pancreaticography (MRCP) in the diagnostic process of common bile duct stones. DESIGN: Retrospective. METHOD: All 27 MRCPs performed in the period December 1997-December 1998 in the Deventer Hospital, the Netherlands, were evaluated using chart examination. The group comprised 11 males and 16 females with an average age of 57 years (SD 3.2) with anamnestic or biochemical cholestasis. If at MRCP stones were diagnosed, endoscopic retrograde cholangiopancreaticography (ERCP) was performed. If MRCP was without abnormalities, no further diagnostic procedures were performed. The findings at MRCP were compared with those at ERCP and with the clinical course. The MRCP examinations were performed on a 1.5 Tesla MR unit. RESULTS: In 16 patients MRCP was performed before laparoscopic cholecystectomy and in 5 there after. In 5 MRCP was performed to rule out a biliary cause of acute pancreatitis and in 1 patient because of an elevated alkaline phophatase after laparotomy for an abdominal stab injury. There was one false-positive MRCP result and no false negative ones. Accordingly, the sensitivity of MRCP for choledocholithiasis was 100% and the specificity 95%. PMID- 11268912 TI - [Perinatal mortality assessed: results of a regional audit]. AB - OBJECTIVE: To assess the level of suboptimal care prior to cases of perinatal death and the extent to which perinatal mortality can be reduced by further improvements in care. DESIGN: Retrospective panel audit investigation. METHOD: Cases of perinatal death occurring in 1996 and 1997 among women living in the region Zuid-Holland-Noord, the Netherlands, were identified by approaching midwives, obstetricians/gynaecologists and paediatricians/neonatologists. The medical records of the cases were studied by an expert panel using a checklist of evidence-based criteria for standard care in order to determine circumstances and actions that did not comply with professional protocols, or that indicated either low compliance of the mother or an inadequate healthcare infrastructure (so called sub-standard factors). The panel also assessed whether the perinatal death could have been prevented. RESULTS: A total of 342 perinatal deaths were found. For 332 cases sufficient information was available for a panel assessment and for 318 cases the panel reached a consensus on the assessment. One or more sub standard care factors were identified in more than half of the cases. In 19% of the cases the panel agreed that the sub-standard factor had 'possibly' contributed to the death, and in 6% they agreed that the sub-standard factor had 'probably' contributed to the death. In the last group the main problems involved were antenatal care (particularly a failure to detect or inadequate management of intrauterine growth retardation) and intrapartum care (too much of a 'wait and see' approach). CONCLUSIONS: This regional audit revealed that further quality improvement of obstetric care is possible if clinical practice guidelines for effective and safe care are better implemented. It is expected that these improvements could reduce the perinatal mortality rate by between 6% and 25%. PMID- 11268913 TI - [Dyspnea after pneumonectomy due to atrial septum defect]. AB - A 55-year-old woman presented with complaints of recurrent dyspnoea one year after pneumonectomy carried out as treatment for a tumour of the left lung. During several months her symptoms progressed and eventually mechanical ventilation became necessary. On admission a patent foramen ovale was found with transoesophageal ultrasound but this was judged not to be the cause of her symptoms. The pulmonary angiogram showed a intracardiac shunt with no intrapulmonary shunts. After repeated transoesophageal ultrasound a second defect was found of a sinus venosus type. This large defect was proven to be clinically significant during catherisation of the heart, when occlusion with a balloon was performed. After surgical repair of these defects with an artificial patch, the patient recovered well. Since then she has been without complaints. PMID- 11268914 TI - [Treatment of severely delayed gastric emptying]. PMID- 11268915 TI - [Interferon for adjuvant therapy of melanoma: approved, not indicated]. PMID- 11268916 TI - [Screening for asymptomatic Chlamydia trachomatis infection: cost-effectiveness favorable at a minimum prevalence rate of 3% or more]. PMID- 11268917 TI - [Confusion surrounding bovine spongiform encephalopathy (BSE) and the risk of new variant Creutzfeldt-Jakob disease]. PMID- 11268918 TI - [Circumcision from a historical perspective]. PMID- 11268919 TI - [Circumcision from a historical perspective]. PMID- 11268920 TI - [What is your diagnosis? (Atypical) bronchial carcinoid, Cushing syndrome due to ectopic ACTH production (somatostatin receptor positive tumor). Incidentaloma of the hypophysis]. PMID- 11268921 TI - [Comparison and reproducibility of 3 indirect methods for measuring blood pressure]. AB - It is rare to find systematic comparisons of various methods for measuring blood pressure. This study aimed to compare blood pressure measurements performed by the doctor, by the patient and on a 24-hour basis with regard to their reliability and reproducibility. A total of 84 patients were included in the study. All patients had their blood pressure measured at the doctor's office twice on day 1 and 10, by themselves and with a 24-h blood pressure measuring unit. The doctors obtained the highest means. The mean systolic and diastolic value taken on the day 1 was higher by 13 mmHg and 8 mmHg than the ABPM value or the measurement obtained by the patient at home, respectively. This difference was still present at the end of the study (7 mmHg and 5 mmHg compared to ABPM and 9 mmHg and 6 mmHg compared to the patient's measurements, respectively). The values obtained by 24-h measurement and the patients themselves showed a better reproducibility than the doctor's measurements. Both methods are thus suitably good for use in diagnostics and therapy monitoring. Compared to the patient's own measurements, the 24-hour blood pressure measurement is less dependent on the patient's reliability. PMID- 11268922 TI - [The significance of postprandial glycemia]. AB - Both acute and chronic hyperglycemia cause tissue lesions which ultimately lead to micro- and macrovascular diabetic complications. The newly developed insulin secretagogues and quick-acting insulin analogues are pharmacological options to reduce the amplitude of glucose fluctuations, thereby reducing acute glucose toxicity. There is hope that new pathophysiologically based therapeutic options will improve glucose control of type 2 diabetes and that physicians can be convinced to change their frequently expressed opinion that rising glucose levels represent the natural course of the disease. The Diabetes Control and Complications Trial has clearly shown that intensive insulin therapy is effective in treating complete insulin deficiency in type 1 diabetes over many years, achieving a stable HbA1c and thereby preventing diabetic complications [1]. PMID- 11268923 TI - [Primary vesiculo-ureteral reflux and reflux nephropathy--new knowledge]. PMID- 11268924 TI - [Dramatic initial manifestation of celiac disease in an adult woman]. PMID- 11268925 TI - [49-year-old patient with cervical lymphadenopathy. Cervical lymph node tuberculosis]. PMID- 11268926 TI - Psychomotor and audiological assessment of infants born to mothers with subclinical thyroid dysfunction in early pregnancy. AB - BACKGROUND: To investigate the frequency and the effects of various degrees of maternal thyroid dysfunction in the first trimester of pregnancy, before the onset of fetal thyroid function, on psychomotor and audiological outcome of the offspring. METHODS: In a cohort of 691 pregnant women, undergoing thyroid screening between the 8th and 10th gestational week, eight were found to have a subclinical form of hypothyroidism and one was frankly hypothyroid. Treatment with L-thyroxine was started soon after diagnosis was made. Their nine offspring had a psychomotor and audiological assessment at the age of nine months. Psychomotor development was evaluated with the Brunet-Lezine test, while audiological function was assessed with auditory brainstem responses (ABR's). RESULTS: Psychomotor developmental quotients were not different in patients and controls (99 +/- 6 vs 101 +/- 4). Regarding ABR pattern, there were no significant differences between patients and controls. Moreover, no correlation was found between maternal fT4 and psychomotor as well as audiological outcome in the offspring. CONCLUSIONS: These findings are reassuring, since various degrees of maternal thyroid dysfunction in early pregnancy seem to have no adverse effects on the psychomotor and audiological outcome of the offspring up to nine months of age. A longer follow-up however is needed before definitive statements can be made. PMID- 11268927 TI - [Evaluation of the effects of a nucleotide-enriched formula on the incidence of diarrhea. Italian multicenter national study]. AB - BACKGROUND: Aim of this study is to evaluate the incidence of diarrhea in children fed with a nucleotide-supplemented formula (Similac FormulaPlus) in comparison with formula without nucleotide supplementation. METHODS: We investigated the effects of a nucleotide-supplemented formula on the incidence of diarrheal episodes in 3315 infants with a multicenter study conduced by 386 pediatricians since March 1998 until October 1999. The study population has been divided into 4 groups based on the type of feeding; group 1 (n = 958) = exclusively nucleotide-supplemented formula, group 2 (n = 824) = exclusively formula without nucleotide supplementation, group 3 (n = 920) = mixed breast feeding and nucleotide-supplemented formula, group 4 (n = 613) = mixed breastfeeding and formula without nucleotide supplementation. At the beginning of the study the 4 groups did not differ for body weight, length and mass index. The infants were enrolled since the first month up to the end of the third month of life and they were followed-up to the end of the sixth month of life. During the period of observation the growth of lenght, weight and mass index was similar among the 4 groups. RESULTS: Monthly incidence of diarrhea was computed and the comparison between group 1 and group 2 using the summary odds-ratio of Mantel Haenszel showed that in group 1 the incidence of diarrhea was significantly lower than un group 2 (RR = 0.567); CI 95% = 0.440-0.732); similar results were obtained comparing the incidence of diarrhea between group 3 and group 4, having the former a RR = 0.630 (CI 95% = 0.476-0.834). CONCLUSIONS: The conclusion drawn is that the supplementation with nucleotide of the formula milk decreases the risk of diarrheal episodes during the first six month of life in healthy infants. Such a positive effect is present both in exclusively nucleotide-supplemented formula and in mixed breast-feeding and nucleotide-supplemented formula fed infants. Interpretation of these results is that nucleotides, much more present in human milk than in formula milk, improve the immune defense of the infants stimulating particularly the cell-mediated immunity. PMID- 11268928 TI - [Mouth mucosa free-flap grafts in repeat operations of hypospadias]. AB - BACKGROUND AND AIMS: Buccal mucosal grafts are widely used in the treatment of primary hypospadias and urethral stenosis owing to their elasticity, optimal attachment, possibility of generous harvesting and easy preparation. The aim of this study was to check whether buccal mucosal flaps are also valuable in redo surgery for hypospadias complicated by large breaks in the urethra and with scarce residual genital tissue. METHODS: Fourteen patients aged between 3 and 11 years old (mean age 6.6) were selected and operated between December 1993 and June 1999. The patients presented extensive fistulous tracts, roughly ellipsoidal in shape and with a maximum diameter of between 7 and 42 mm (mean length 18 mm). The original technique was: Duplay (7 patients); Onlay buccal graft (1 patient); Snodgrass (1 patient); Tubulised preputial flap (2 patients); Onlay preputial graft (2 patients); Tubulised vesical mucosal flap (1 patient). The mucosal flap, taken from the lower lip, was used to cover the gap as an onlay patch and recovered with residual genital skin with the interposition, where possible, of a de-epithelised flap. RESULTS: An optimal cosmetic and functional result was achieved in 10/14 cases with flowmetry > 25 percentile according to Toguri nomograms. Three patients presented fistulas: one punctiform fistula resolved spontaneously. The other two cases resolved after corrective surgery. One patient showed meatal regression with slight stenosis that was resolved with MAGPI. CONCLUSIONS: These results appear to be encouraging. Buccal mucosal graft may represent a valid alternative also in the treatment of secondary hypospadia with large breaks in the urethra. No complication was reported in the harvesting area, even if this was carried out at a second stage in the labial area. PMID- 11268929 TI - Diabetes mellitus in the pre-school child. AB - Diabetes mellitus presents rarely in the pre-school child and presents specific problems because of the peculiarity of the young child physiology. Pathogenesis involves the classic immunological mechanisms, with a higher incidence of other autoimmunity and family history of diabetes. Because of the rarity of the condition in this age group, the delay of the recognition of the signs and symptoms, which are often subtle at onset, determines the increased incidence of ketoacidosis. The reasons for the lower glycaemic control in this age group include the persistence of endogenous insulin, but also a more detailed involvement by the parents in organising diabetes. For the same reason ketoacidosis is an unusual occurrence after diagnosis. As to insulin therapy, three or more injections a day should be recommended, as in the older child, while the modern devices for blood glucose monitoring have proved useful to improve glycaemic control and to decrease the frequency of nocturnal hypoglycaemia, which gives particular concern given the vulnerability of the nervous system in this age group. Management of diabetes in the pre-school child may result very difficult for both parents and health carers because of the erratic daily pattern of activity, sleep and feeding; however, with a cautious strategy which involves insulin therapy, diet and monitoring it is possible to achieve satisfactorily the following aims: physical well-being of the young child, normal growth, lack of hyperglycaemia or hypoglycaemia, acceptable value of glycosilated haemoglobin. PMID- 11268930 TI - [Congenital abnormalities of the temporal bone. Clinical picture and therapeutic indications]. AB - The embryology of the ear and the different congenital anomalies depending on the arrest in development at a particular embryologic date are reviewed. The historical and the actual attitude towards surgical correction is discussed. The classification system, the evaluation and patients selection, the timing of auricular reconstruction and atresiaplasty, the preoperative evaluation, the patient and parents counselling, the surgical technique, the complications and alternative techniques are discussed. PMID- 11268931 TI - [Obstructive syndrome caused by trichobezoars: historical disease or disease still current? Description of a case in adolescence]. AB - Trichobezoars are masses of entangled material, found in the stomach and intestines, composed of hair ingested by the patient. When the mass grows, symptoms of intestinal occlusion can appear. Trichobezoars in pediatrics are usually found in adolescent females presenting personality disorders and trichophagia. This work describes a case of trichobezoar diagnosed in a 13 year old girl, wearing a brace for serious scoliosis but absolutely normal from the emotional and psychical point of view, with normal scholastic and familiar situation. Already hospitalized three months before for pneumonia from mycoplasma, the girl comes to our observation for the appearance of vomiting and constipation. The clinical examination reveal an epigastric mass as big as an orange. General conditions and hematochemical tests are good. Lab tests are performed (abdomen echography and upper abdomen MNR) but is the oesophago gastroscopy which allows the diagnosis. A big trichobezoar is then surgically removed and the gastroenteric symptoms completely disappear. PMID- 11268932 TI - Spontaneous gastric perforation in premature twins. AB - Two pairs of identical and non-identical premature neonates proceeding from twin pregnancies were operated on for spontaneous gastric perforation. The newborns in our case, one girl and one boy two different pregnancies were delivered by emergency cesarean section. Their gestational ages were 30 and 32 weeks, and their birth weight 1400 and 2100 g, respectively. Both of the neonates were being treated in the Neonatal Intensive Care Unit when the perforations were diagnosed. They presented clinically abrupt symptoms of abdominal distension and pneumoperitoneum. The sites of the ruptures were located at the anterior gastric wall near the gastroesophageal junction. The sibling twins were consequently also observed very carefully and fortunately they did not develop any similar clinical symptoms. All four twins were finally discharged from the hospital in good condition. PMID- 11268933 TI - [Bacteremia in children. Evaluation of a caseload assisted in a period of 21 months at a pediatrics division]. PMID- 11268934 TI - [Equilibrium and orthognathodontic surgery: correlations in a group of patients undergoing treatment]. AB - BACKGROUND: The aim of this study was to evaluate the main parameters provided by the static stabilometric test (mean X, mean Y, mean velocity, length of tracing, standard deviation of velocity, ellipse area) in the follow-up of patients suffering from skeletal occlusive pathology undergoing orthognathodontic surgery to confirm the re-establishment of postural equilibrium. METHODS: Fifteen patients with skeletal dysgnathia were correlated with a group of 10 healthy subjects. The same parameters were analysed in the dysgnathic subjects at 6 and 12 months after surgical correction. The patients enrolled in this study underwent surgery at the Division of Maxillofacial surgery of Turin University. Student's "t"-test and multivariate statistical analysis (Cox regression) were used for the statistical analysis of results. RESULTS: A significant variability was noted in some of the main parameters analysed (mean X, mean Y, tracing length) between the two populations (healthy and dysgnathic) compared to visual signs (eyes opened-closed). The change in stabilometric values within the group of dysgnathic patients was highly significant 6 and 12 months after surgery, not only in terms of visual signs but also the cervical component (retroflexion of the head), above all the value of mean Y (p = 0.001). CONCLUSIONS: An analysis of these results shows that static stabilometry can be a valuable aid both during the preoperative evaluation and during the follow-up in patients undergoing jaw surgery since it can quantify the improvement of body balance. PMID- 11268935 TI - [Titanium joints welded with laser and infrared techniques: comparative analysis of its microstructure]. AB - BACKGROUND: Aim of the study is to compare microstructure of Titanium joints soldered with Laser and Infrared methods by performing SEM analysis, metallography and microhardness evaluation. METHODS: Wax specimens were separated in the middle area corresponding to the joint region, machined and then soldered. 40 Titanium samples were fused and divided in 2 groups of 20 samples each. First group was soldered by laser welding, second one by infrared brazing. SEM analysis and standard metallography were carried out at joined areas and unsoldered surfaces. Microhardness test was performed on longitudinally sectioned samples with 150 g load for 15 second up to 7 mm distance from the joined area. RESULTS: SEM photomicrographs revealed for group 1 a homogeneous metal-joint interface, without microporosities; for group 2 exhibited a distinct demarcation of metal joint interface. Metallography evaluation showed for laser joined samples only the presence of Titanium; infrared joined samples showed in soldered regions also Ni and Cu. Microhardness values detected at the joined surfaces seem to be higher for both considered groups. CONCLUSIONS: Laser joining method with exclusive presence of Titanium seems to be ideal for monometallism; both techniques exhibited microstructural changes in the heated surface layer. PMID- 11268936 TI - [Clinical evaluation, radiologic and histologic analysis in mandibular alveolar distraction procedures. Preliminary study]. AB - BACKGROUND: Alveolar distraction osteogenesis is a process to form new alveolar bone to correct alveolar deformities in ridge height and width. Aim of this work is to study the bone processes to optimize the implantoprosthetic rehabilitation. METHODS: Alveolar distraction osteogenesis was applied in 7 patients with ridge deformities to obtain the desired ridge augmentation. Clinical and radiological evaluations were performed during the following 12 weeks, before implant insertion. Biopsies at 40, 60 and 88 days were studied after general, specific and histochemical staining of slides; microradiographs were analyzed to evaluate the trabecular bone volume. RESULTS: Forty days after the end of distraction, soft callus shows the start of ossification. Sixty days after the end of distraction, soft callus was widely converted into a network of trabecular woven bone; osteogenic activities were low; trabecular bone volume was about 50%. Eighty-eight days after the end of distraction bone amount appeared reduced, with a more ordered structure, further reduction of bone formation activity, whereas osteoclast erosion was active. CONCLUSIONS: Results show an almost steady-state bone deposition processes 60 days after the end of distraction and a regress with longer time. The results suggest the possibility of an early implant insertion to avoid bone loss due to mechanical unloading. PMID- 11268938 TI - [Treatment of mandibular condylar hyperplasia in developmental age. Clinical case]. AB - A case of hyperplasia of the mandibular condyle in a growing-up subject, observed at the Department of Maxillo-Facial Surgery of the University of Rome "La Sapienza", is described. Hyperplasia of the mandibular condyle is a facial asymmetry due to the unilateral overdevelopment of the mandibular bone. In this study the authors underline how bone scintigraphy, 3D tomography and electrognatographic analysis, associated with standard radiography and cephalometry, are important methods of diagnosis in order to make an early diagnosis of hyperplasia of the mandibular condyle and differential diagnosis with other pathologies. In particular, bone scintigraphy is a useful screening procedure to detect if the pathology is in an active phase or not. The 3D tomography is used in pre-surgery to evaluate precisely morphological and structural alterations of the craniofacial bones on a tridimentional base. Finally, the electrognatographic test records the mandibular activity both in physiological and pathological conditions. All these instrumental techniques allow to make a diagnosis and lead to a possible therapeutical approach. PMID- 11268937 TI - [Bacterial virulence in the etiology of periodontal diseases]. AB - Strong relationships have been very often described between various form of periodontal disease (PD) and certain bacterial species, so that nowadays periodontal disease is recognized as an infectious disease. Destruction of periodontal supporting tissues happens as a response to very intricate host parasite interactions. When the clinician will be able to fully understand and identify such phenomena it would be possible to succeed in a properly diagnosis and control of the active phase of periodontal disease. The first step in such a direction would be to analyze the common characteristic of some bacterial species, the so called suspected periodontopathogens. Such species namely Gram negative, associated with the outbreak of periodontal disease have in common the capacity to disrupt the integrity of the host defences by means of the so called virulence factors. These factors may enhance the bacterial colonization or may interfere with the host response that ultimately results in periodontal support breakdown. The present review focuses on the virulence factors of the main suspected periodontopathogens evaluating the effects on the host immune response and directly on the periodontal tissues. PMID- 11268939 TI - Hepatic resection under selective portal vein occlusion using degradable starch microspheres. A simple procedure of an experimental study using the pig. AB - A simple technique is described to perform hepatectomy. The technique consists of portal vein blood flow occlusion using degradable starch microspheres injected through an ultrasonically-guided puncture at the point of resection root and hepatic artery occlusion. The method minimizes ischemic damage to the liver remnant and is easy to perform. PMID- 11268940 TI - Synchronous gastric adenocarcinoma and MALT lymphoma in a patient with H. pylori infection. Could the two neoplasms share a common pathogenesis? AB - Low-grade primary MALT (mucosa-associated lymphoid tissue) lymphoma of the stomach is a neoplasm with an indolent course and a good prognosis. Patients with this type of neoplasm seem to have a higher risk for other neoplasms. Of interest is the association of gastric MALT lymphoma with gastric adenocarcinoma of intestinal type. We report the case of a patient, with a history of H. pylori related gastritis, in whom a diagnosis of synchronous gastric adenocarcinoma of intestinal type and low-grade MALT lymphoma, occurring as collision tumors, was made. The stage procedures confirmed the presence of a locally advanced gastric tumor staged as T3 N1. The patient underwent two cycles of neoadjuvant EEP (etoposide, epirubicin, cisplatin) chemotherapy. After 2 months, a R0 total gastrectomy with D2-lymphoadenectomy was successfully performed. The development of simultaneous primary gastric lymphoma and carcinoma is a rare event. The possible coexistence of both tumors should be kept in mind, especially in patients infected with H. pylori, since a possible etiopathogenetic role of this bacterium has been differently postulated for both disease. PMID- 11268941 TI - Splenoadrenal shunt. An original portosystemic decompressive technique. AB - Management of gastrointestinal hemorrhage from rupture of esophageal and gastric varices due to portal hypertension remains a debated question. In patients with sclerotherapy-resistant esophagogastric varices, and preserved hepatic function, a surgical shunt is considered the treatment of choice. A 63-year-old male was admitted in our Department with a diagnosis of idiopathic fibrosis of the liver, portal hypertension, esophageal and gastric varices and previous history of variceal bleeding. A distal splenorenal shunt was planned. During the isolation, a large diameter left adrenal vein was identified. An end-to-end anastomosis utilizing the distal splenic vein and the proximal adrenal stump was performed. The procedure was uneventful. An ultrasound color-Doppler on the 3rd postoperative day, showed normal intrasplenic resistance index, demonstrating the efficacy of the shunt. A splenic angiography carried out on the 8th postoperative day showed the complete patency of the splenoadrenal shunt. At the 15th postoperative day, the patient was discharged. In patients with portal hypertension, sclerotherapy-resistant esophagogastric varices and preserved hepatic function, a surgical portosystemic shunt is mandatory. Splenoadrenal shunt, utilizing a left adrenal vein represent an excellent option in selected cases. PMID- 11268942 TI - Gastric rupture after Heimlich maneuver and cardiopulmonary resuscitation. AB - Choking is a common emergency problem. The Heimlich maneuver is unquestionably effective in relieving airway obstruction. Serious and life-threatening complications may arise, however, if the maneuver is applied incorrectly. Two cases of gastric rupture after Heimlich maneuver are reported. Lay public, paramedics and the medical professionals should be educated with the correct technique of Heimlich maneuver and its potential complications. All patients receiving Heimlich maneuver should be examined by an experienced physician. PMID- 11268943 TI - Gallbladder adenomyomatosis presenting as fever of unknown origin: a case report. AB - Gallbladder adenomyomatosis is a rare disorder, characterized by benign hyperplasia of the gallbladder mucosa creating invaginations through the muscular layer, known as Rokitansky-Aschoff sinuses. It is considered an acquired disease, with pathophysiology similar to that of the diverticular disease of the colon. Diagnosis is often achieved by ultrasound, but a significant percentage is misdiagnosed as chronic cholecystitis, whereas the diagnosis is finally achieved histologically. We describe a case of gallbladder adenomyomatosis presenting as fever of unknown origin. The patient was a 17-year-old girl with a history of sustained fever of 38.5 degrees C of two months' duration. There were no accompanying symptoms and the whole diagnostic workup, including abdominal ultrasound, was negative. Gallbladder inflammation was evident during an eventual investigatory laparoscopy, and cholecystectomy was performed. The histologic results were consistent with diffuse adenomyomatosis. The patient became afebrile immediately after cholecystectomy. To our knowledge, fever has never been associated with gallbladder adenomyomatosis before in bibliography, nor has adenomyomatosis been mentioned as a cause of fever of unknown origin. We therefore believe that gallbladder adenomyomatosis should be considered in the differential diagnosis of sustained fever with negative workup. PMID- 11268944 TI - Radical surgery of colon cancers directly invading the duodenum, pancreas and liver. AB - BACKGROUND/AIMS: Adjacent organ invasion is observed in 5-12% of the colorectal cancers and is rarely faced with invasion of the duodenum, pancreas or liver. METHODOLOGY: We reviewed 4 patients with invasion of the duodenum, pancreas or liver or more than one at the same time, to emphasize the importance of aggressive radical procedures. RESULTS: Three patients underwent en bloc pancreaticoduodenectomy with right hemicolectomy, and one patient underwent en bloc pancreaticoduodenectomy with right hemicolectomy and 5th and 6th hepatic segments resection. Perioperative need for blood transfusion was 3-5 units. There was no postoperative morbidity and mortality, except for the patient who had a bile leakage which got well with medical treatment. Although malignant invasion was proved by pathologic evaluation, only one patient had lymph node involvement. While one patient was lost at the 8th postoperative month, 3 patients are living disease free within a range of 14-41 months. CONCLUSIONS: The chances of longer survival can be given to the patient by en bloc radical resections of locally advanced right colon tumors accompanied by invasion of duodenum, pancreas and liver, with low morbidity and mortality rates. PMID- 11268945 TI - Differing effects of two nitric oxide synthase inhibitors on experimental colitis. AB - BACKGROUND/AIMS: The aim of this study was to investigate the effects of two inhibitors of nitric oxide synthase on a rat model of colitis. METHODOLOGY: Colitis was induced by administration of an enema containing trinitrobenzene sulfonic acid. This colitis was treated everyday for one week with NG-nitro-L arginine (10 mg/kg, i.v.), which is a non-selective inhibitor of both constitutive nitric oxide synthase and inducible nitric oxide synthase, or aminoguanidine (10 mg/kg, i.v.) which is an inhibitor of inducible nitric oxide synthase. Exposure to the trinitrobenzene sulfonic acid enema inhibited the increase in body weight of rats, and markedly increased the colonic damage scores, wet weight, thiobarbituric acid-reactive substances and myeloperoxidase activity. RESULTS: The inhibition of weight increase caused by trinitrobenzene sulfonic acid was significantly reduced by aminoguanidine treatment, whereas weight loss tended to be aggravated by NG-nitro-L-arginine treatment. The increases in the colonic damage scores, wet weight, thiobarbituric acid-reactive substances and myeloperoxidase activity in trinitrobenzene sulfonic acid-colitis were significantly inhibited by aminoguanidine treatment, although they tended to be aggravated by NG-nitro-L-arginine treatment. CONCLUSIONS: These results suggest that an inhibitor of inducible nitric oxide synthase, but not of constitutive nitric oxide synthase, was effective in treating experimental colitis in rats. PMID- 11268946 TI - Results of lymphadenectomy for obvious lateroaortic lymph node metastases from colorectal primaries. AB - BACKGROUND/AIMS: To analyze the results of surgery for macroscopically or radiologically obvious (i.e., easily detectable by computed tomography scan or by palpation) synchronous or metachronous lateroaortic lymph node metastases from colorectal primaries. METHODOLOGY: Thirty-one highly selected patients who underwent a lateroaortic lymphadenectomy for obvious lateroaortic lymph node metastases from January 1989 to January 1999 were analyzed retrospectively. An associated metastatic lesion was present in 68% of the cases before or concomitantly with the lateroaortic lymph node metastases. Ten lateroaortic lymph node metastases were synchronous with the primary, and 21 were metachronous. Decision for lymphadenectomy was taken after a multidisciplinary meeting and a period of observation. Median follow-up after lymphadenectomy was 24.2 months (range: 6-120). All the patients received at least two systemic lines of chemotherapy before or after the lateroaortic lymphadenectomy. RESULTS: There was no postoperative mortality. Resection was macroscopically complete (R0-1 of UICC) in 26 cases (84%). Twenty-six (83.8%) patients developed recurrent lesions or had progressive residual disease. The most frequent first site of recurrence was intrathoracic (54.8%) for the entire series, except for the subgroup of isolated lateroaortic lymph node metastases in which recurrent lesions were mainly lateroaortic. Three-year global and disease-free survival rates were, respectively, 39% and 9.6%. No significant difference was noted in survival between lateroaortic lymph node metastases that were synchronous or metachronous with the primary. However, the most important prognostic factor was the presence of associated metastases. Indeed 3-year survival attained 30% when lateroaortic lymph node metastases were isolated but 0% when lateroaortic lymph node metastases were associated with another metastatic site (P = 0.006). CONCLUSIONS: Obvious lateroaortic lymph node metastasis is rarely isolated. However, when it is isolated, in selected cases (objective response to systemic chemotherapy, good general status), resection can be beneficial whatever its synchronous or metachronous appearance. PMID- 11268947 TI - Postsurgical sequential methotrexate/5-FU and leucovorin on outpatient basis for advanced colorectal carcinoma. AB - BACKGROUND/AIMS: The present study compared the effects of sequential methotrexate and 5-fluorouracil followed by leucovorin rescue (MFL), as an adjuvant chemotherapy versus a combination of UFT and mitomycin C (MMC), on patient survival and recurrence after surgery for colorectal carcinoma. METHODOLOGY: Between January 1990 and December 1997, a total of 55 patients with advanced colorectal cancer were treated postsurgically by adjuvant chemotherapy using MFL or UFT-MMC. Surgical treatment was performed according to standardized procedures for radical resection of colorectal cancer. The patients were divided into 2 groups after surgery. The MFL regimen consisted of MTX (100 mg/m2) and 5 FU (600 mg/m2) at hour 24, followed by leucovorin rescue. The UFT-MMC regimen consisted of MMC (12 mg/m2) intraoperatively and MMC (6 mg/m2) every other week after surgery for 2 months, and oral UFT (375 mg/m2/day), a combination of tegafur and uracil in a molar ratio of 1:4, was continued for 3 years or longer depending on the patients tolerance. RESULTS: The overall survival rates after surgery were significantly (P < 0.05) higher in the MFL than the UFT-MMC group. Recurrence rates were significantly lower in the MFL than the UFT-MMC group, especially for liver recurrence. Disease-free survival was significantly (P < 0.05) higher in the MFL than the UFT-MMC group. CONCLUSIONS: These results demonstrated the superiority of MFL therapy for improving postsurgical survival in patients with advanced colorectal cancer, in particular those patients with a high risk of recurrence following potential curative resection. PMID- 11268948 TI - Interventional radiology and endoscopic therapy for recurrent esophageal varices. AB - BACKGROUND/AIMS: We investigated the impact long-term prognosis of combined interventional radiology and endoscopic therapy in patients with esophageal varices. METHODOLOGY: Patients with recurrent esophageal varices underwent treatment as follows: 54 were treated with endoscopic therapy alone and 32 underwent endoscopic therapy plus interventional radiologic procedures. Primary endpoints during 5-year follow-up included recurrent bleeding, second retreatment, and death. RESULTS: The bleeding rates were 11.1% in the endoscopy group, and 9.4% in the combined therapy group. Second retreatment rates at 1 year, 3 years, and 5 years in the endoscopy group and combined therapy group were 25.4% and 17.2%, 70.2% and 39.3%, and 85.0% and 69.6%, respectively. The second retreatment rates in the combined therapy group were significantly reduced compared to the endoscopy alone group (P = 0.05). Cumulative retreatment rates in Child's class C cases were significantly lower in the combined therapy group than in the endoscopy group (P = 0.01). Survival at 3 years was 97.1% in the endoscopy group and 92.0% in the combined therapy group, and 5-year survival was 79.1% and 83.6%, respectively. CONCLUSIONS: The combination of interventional radiologic and endoscopic therapy is highly effective and improves long-term prognosis in patients with recurrent esophageal varices. PMID- 11268949 TI - True carcinosarcoma of the esophagus with osteosarcoma. AB - We experienced a case of true carcinosarcoma of the esophagus with osteosarcoma. Computed tomography scan revealed calcification of the tumor. Immunohistochemically, vimentin staining was positive, and osteoid formation was found in the sarcomatous portion. It was clear that the lesion contained osteosarcoma of mesenchymal origin, and thus we diagnosed it as true carcinosarcoma. PMID- 11268950 TI - Unenhanced spiral CT for evaluating acute appendicitis in daily routine. A prospective study. AB - BACKGROUND/AIMS: The purpose of this study was to define in a routine setting the role of spiral computed tomography in patients with suspected acute appendicitis and to determine the effect of computed tomography on the treatment of such patients. METHODOLOGY: Appendiceal computed tomography was performed in 120 consecutive patients with acute appendicitis in the differential diagnosis, whose clinical findings were insufficient to perform surgery or to discharge from the hospital. Each scan was obtained in a single breath hold from the lower abdomen to the upper pelvis using a 5-mm collimation and a pitch of 1.6. Computed tomography results were correlated with surgical and pathologic findings at appendectomy or clinical follow-up. RESULTS: Eighty-eight of the 93 patients with acute appendicitis were correctly diagnosed by computed tomography, 24 of the 27 patients without acute appendicitis were correctly diagnosed by computed tomography (95% sensitivity, 89% specificity). Computed tomography signs of acute appendicitis included fat stranding (100%), enlarged appendix (> 6 mm) (97%), adenopathy (63%), appendicoliths (43%), abscess (10%), and phlegmon (5%). CONCLUSIONS: The use of spiral computed tomography in patients with equivocal clinical presentation suspected of having acute appendicitis led to a significant improvement in the preoperative diagnosis and a lower negative appendectomy rate. Appendiceal computed tomography is an accurate technique even if performed in the daily routine of scanning. PMID- 11268952 TI - Validation of MPI and PIA II in two different groups of patients with secondary peritonitis. AB - BACKGROUND/AIMS: There are several scoring systems designed to predict mortality in patients with peritonitis, which need validation in different patient populations. Our aim was to evaluate Mannheim Peritonitis Index (MPI) and Peritonitis Index of Altona (PIA II) in patients with postoperative peritonitis and other causes of secondary peritonitis. METHODOLOGY: The records of patients operated for intraabdominal infection between 1987-1996 in Hacettepe University Department of General Surgery, were reviewed retrospectively. A total of 473 patients were included in the study; 75 of them had postoperative peritonitis (POSTOP group) and the remaining 398 had secondary peritonitis due to other causes (OTHER group). Using multiple logistic regression, MPI and PIA II were combined in an equation and this new variable was called combined peritonitis score (CPS); CPS = -9 + (0.3 x MPI) + (-1.2 x PIA II). All patients were scored according to MPI, PIA II and CPS. Receiver-operator characteristic (ROC) curves and sharpness of scores were compared. Also mean scores in both groups, proportions of correct predictions of outcome according to scores and correlation of scores with mortality were compared. RESULTS: Overall mortality was 17.8% in OTHER group and 33.3% in POSTOP group (P = 0.0018). Higher MPI scores, lower PIA II scores and higher CPS scores were associated with higher mortality in both groups (P < 0.0001). Mean MPI values were higher, mean PIA II values were lower and mean CPS values were higher in POSTOP group (P < 0.001). The areas under ROC curves of CPS were bigger than MPI and PIA II in both groups. Sharpness of CPS was higher in both groups compared to MPI and PIA II (P < 0.05). Proportion of correct predictions of outcome was highest in CPS among the three scores (P = 0.0074). CPS had the best correlation with observed mortality. CONCLUSIONS: POSTOP group patients had higher MPI, lower PIA II and higher CPS values ending up with higher mortality. This may be because of the delay in diagnosis and treatment, resulting with higher organ failure rates. Generally the results of evaluations for MPI and PIA II are similar. When these two peritonitis scores are combined and used together in the form of CPS, all the parameters improve. PMID- 11268951 TI - What is the essential role in the normalized rat intestinal transit? Calcium or calcitonin. AB - BACKGROUND/AIMS: Both calcium and calcitonin have been involved in mediating gastrointestinal motility. Using a thyroidectomized rat model the present study tried to determine which one of them was essential for normal small intestinal transit. METHODOLOGY: Adult Sprague-Dawley male rats simultaneously received thyroidectomy or a sham operation plus a laparotomy for duodenal tubing. After two-week housing, a small intestinal transit study was conducted via geometric center computation of radioactivities of the intestinal segments counted from intraduodenal feeding of radiochromium. Before feeding, these rats were i.p. injected with saline or human calcitonin in the doses of 0.1, 1 and 10 microM/kg, respectively. Another group of Tx rats received i.v. infusion of saline or CaCl2 for 30 min before motility study. RESULTS: Thyroidectomization effectively delayed transit compared to the sham-operated controls (3.59 +/- 0.15 vs. 2.98 +/ 0.32, P < 0.05). High doses of calcitonin treatment in thyroidectomized rats further inhibited transit (P < 0.01), whereas calcitonin did not restore the suppressed calcium level. Delayed transit in thyroidectomized rats was restored after calcium infusion (2.95 +/- 0.22 vs. 3.78 +/- 0.42, P < 0.05) while their hypocalcemia was already corrected. CONCLUSIONS: Thyroidectomization disturbs small intestinal transit, while peripheral calcitonin replacement further inhibits transit. Since calcium replacement enables to restore small intestinal transit, it is apparent that calcium rather than calcitonin is essential for normal small intestinal transit. PMID- 11268953 TI - Therapy of HCC-radiofrequency ablation. AB - Radiofrequency interstitial hyperthermia has been used for percutaneous ablation of hepatocellular carcinoma, under ultrasound guidance in local anesthesia. Conventional needle electrodes require a mean number of 3 sessions to treat tumors of diameter < or = 3 cm. Tumors up to 3.5 cm in diameter can be treated in 1 or 2 sessions by expandable needle electrodes. With both methods in all treated cases, ablation of tumors was obtained. In a group of patients with long follow up, survival rate at 5 years was 40%. In a mean follow-up of 23 months 41% of patients had recurrences (local recurrences in 5%; new lesions in 36%), which often could be retreated by a new course of radiofrequency application. In recent experience large hepatocellular carcinomas (up to 6.8 cm in diameter) were treated by a combination of segmental transcatheter arterial embolization followed by radiofrequency application. In this way most tumors were ablated in one session of radiofrequency therapy. No fatal complications were observed. Major complications were: strong pain due to capsular necrosis in one patient; hemotorax in one case; a fluid collection in the site of ablated tumor in one patient treated by combination of transcatheter arterial embolization and radiofrequency application. PMID- 11268954 TI - The impact of clinical types of disease manifestation on the risk of early postoperative recurrence in Crohn's disease. AB - BACKGROUND/AIMS: Surgery for Crohn's disease is frequently followed by symptomatic recurrence, which in up to 40% requires reoperation within 6 years. Whilst there is evidence that postoperative medical prophylaxis can be efficient, the results of clinical trials are inconsistent regarding the achieved benefit for the patient. Several parameters have been claimed to indicate an increased intrinsic risk of early surgical recurrence. METHODOLOGY: Patient charts of 287 patients who had undergone abdominal surgery for Crohn's disease were reviewed. Mean follow-up was 4.4 years. Recurrence-free intervals were calculated by the Kaplan-Meier method. A uni- and multivariate analysis was conducted to assess the impact of possible indicators of the need of repeated surgery. RESULTS: Patients with fistulizing type of symptoms, extraintestinal manifestations, corticosteroid treatment or male gender experienced significantly earlier reoperation. Recurrent disease, histologic evidence of inflamed resection margins, patient's age at the time of primary diagnosis and operation and the presence of epitheloid granulomas did not show significant influence on recurrence-free intervals. CONCLUSIONS: We conclude that the natural course of disease after intestinal resection in patients with one or more of these risk factors tends towards earlier recurrence requiring surgical intervention. The risk factors identified in this trial may be useful for patient stratification for randomized trials on the efficacy of medical prophylaxis. PMID- 11268955 TI - The therapeutic strategies in performing emergency surgery for gastroduodenal ulcer perforation in 130 patients over 70 years of age. AB - BACKGROUND/AIMS: Gastroduodenal ulcer is a very common illness in Japan. As the number of elderly persons in Japan increases the same as in Europe and America, the number of such patients requiring a gastroduodenal emergency operation has also increased. Regarding the complications of peptic ulcer, a perforation remains the most important fatal complication. The aim of this study is to investigate the operative risk factors and the long-term recurrence rates and to define the optimal surgical procedures in emergency situations in elderly patients. METHODOLOGY: From April 1988 through March 1997, 130 patients over 70 years of age with a perforated gastroduodenal ulcer (a duodenal ulcer perforation in 50 patients and a gastric ulcer perforation in 80 patients) were operated on in an emergency situation in our clinic. We investigated the following items; medical illness, preoperative risk factor, optimal surgical procedure, postoperative organ failure and the cumulative recurrence-free rates after surgical treatment. RESULTS: A significant correlation with mortality was observed in patients with established comorbidity in the following organs: lung (P = 0.03), heart (P = 0.02), kidney (P = 0.04), and diabetes (P = 0.03). The highest postoperative mortality rate was recorded in patients who underwent a simple closure of a duodenal ulcer perforation (4 patients; 26.7%), while the lowest postoperative mortality rate was recorded in patients who underwent a simple closure and vagotomy of a duodenal ulcer perforation (3 patients; 12.5%). In gastric ulcers, the mortality rate in patients with a gastrectomy was significantly higher than in patients with a simple closure. The practical application of the three risk factors (preoperative shock, delay to surgery over 24 hours, and medical illness) was shown by the progressive rise in the mortality rate with the increasing number of risk factors. Based on the 5 postoperative years after treating a perforated duodenal ulcer, the cumulative recurrence rate after a simple closure (63.6%) was significantly higher than that after a simple closure and vagotomy (38.1%) (n = 0.02) or after gastrectomy (0%) (P < 0.001). At 5 years postoperatively, the cumulative recurrence rate after a simple closure (41.2%) was significantly higher than that after a gastrectomy (15.9%) (P < 0.01). CONCLUSIONS: In conclusion, in an emergency situation, elderly patients are in a highly unfavorable prognostic condition due to their advanced age, and comorbidity, which thus leads to poorer results, not only worldwide, but also in Japan. Based on our findings, in duodenal ulcer cases, a simple closure and vagotomy is recommended because of its low mortality and minimal stress, except for cases with a giant perforation measuring over 20 mm in diameter at the perforation hole or with severe duodenal stenosis. In stomach ulcer cases, a gastrectomy may be recommended because of its low recurrence rate. PMID- 11268956 TI - Percutaneous radiofrequency ablation therapy using a clustered electrode in the animal liver. AB - BACKGROUND/AIMS: The aim of the present study was to examine the safety and effectiveness of percutaneous radiofrequency ablation therapy using a needle with cluster radiofrequency electrodes in an animal model. METHODOLOGY: A total of 10 radiofrequency applications were performed in the normal liver of 5 domestic pigs with real-time ultrasonography until roll-off occurred two times. Aspartate aminotransferase, alanine aminotransferase, lactic dehydrogenase, and total bilirubin were evaluated before the procedure and 1 h, 24 h, and 7 days following percutaneous radiofrequency ablation therapy. The animals were euthanized 1 or 2 weeks after percutaneous radiofrequency ablation therapy, and the livers were removed for gross and histopathologic analysis for coagulation necrosis. RESULTS: There were no complications in any of the experimental animals. Aspartate aminotransferase, alanine aminotransferase, and lactic dehydrogenase levels peaked 24 h following percutaneous radiofrequency ablation therapy, and decreased with time thereafter. Total bilirubin was not elevated in any of the animals at any time. Macroscopic examination revealed that the area of coagulated necrosis was 28 x 21 mm when using a 2.0-cm needle, and 41 x 35 mm when using a 3.5-cm needle. Coagulation necrosis did not occur near large vessels. Microscopic examination of the fixed tissue revealed that coagulation necrosis occurred in preserving lobular structure. CONCLUSIONS: Percutaneous radiofrequency ablation therapy using a clustered electrode is a safe and effective treatment for liver tumor. Incomplete coagulation necrosis, however, can occur when percutaneous radiofrequency ablation therapy is performed for tumors located near large vessels. PMID- 11268957 TI - The value of serum-ascites albumin gradient for the determination of portal hypertension in the diagnosis of ascites. AB - BACKGROUND/AIMS: The aim of the present study was to evaluate the correlation between serum-ascites albumin gradient and portal pressure gradient in a population with ascites related to multiple conditions. METHODOLOGY: Thirty-seven patients were divided into two groups: group 1: 30 patients with cirrhosis as the cause of ascites, and group 2: 7 patients with ascites due to other causes. All patients were submitted to paracentesis and blood examination to determine the serum-ascites albumin gradient and the hepatic venous pressure gradient was measured. RESULTS: Mean serum-ascites albumin gradient was 2.0 g/dL in group 1 and 0.6 g/dL in group 2. Mean hepatic venous pressure gradient was 14.7 mm Hg in group 1 and 1.3 mm Hg in group 2. CONCLUSIONS: There was a significant correlation between the serum-ascites albumin gradient and the hepatic venous pressure gradient (r = 0.502), indicating the reliability of the serum-ascites albumin gradient in demonstrating the presence of portal hypertension and its relationship with the origin of ascites. PMID- 11268958 TI - Long-term outcome of kidney transplantation in patients with hepatitis C virus infection. AB - BACKGROUND/AIMS: The impact of HCV (hepatitis C virus) infection on the long-term outcome of kidney transplant patients is controversial. METHODOLOGY: Eighty-four renal allograft recipients who were seronegative for hepatitis B surface antigen and had been screened for antibody to hepatitis C virus (anti-HCV) were included. The outcome and survival were compared between anti-HCV-positive (n = 30, group 1) and anti-HCV-negative (n = 54, group 2) kidney transplant patients. Group 1 patients were further compared to 52 anti-HCV-positive end-stage renal disease patients (group 3) who were on chronic dialysis. RESULTS: Group 1 patients had a higher prevalence of chronic hepatitis than group 2 and group 3 patients did (67% vs. 2% and 31%). Liver-related complications and deaths between group 1 and group 2, and group 1 and group 3 patients were not significantly different. The comparisons of the long-term survival between these groups showed no significant differences, despite group 3 patients had a higher overall mortality rate. Cox regression analysis confirmed that age more than 45 years was the only independent factor that affected survival in anti-HCV-positive end-stage renal disease patients with or without kidney transplantation. CONCLUSIONS: HCV infection is not a contraindication to kidney transplantation. For anti-HCV positive end stage renal disease patients, survival is better in younger patients, and is not influenced by kidney transplantation or continuing dialysis. PMID- 11268959 TI - Effect of translocation of cirrhotic liver on hepatic venous hemoglobin oxygen saturation during hepatectomy. AB - BACKGROUND/AIMS: Comparison of conventional abdominal approach and right thoracoabdominal approach for the resection of hepatocellular carcinoma in cirrhotic liver located at right upper lobe was made in accordance with the effect of translocation of right lobe on hepatic venous hemoglobin oxygen saturation. METHODOLOGY: From 1990-1994, 92 initial hepatectomies were performed in patients with hepatocellular carcinoma at our department. Nine patients underwent resections of hepatocellular carcinoma located at the right upper lobe with the right thoracoabdominal approach. On the other hand, 10 patients underwent resections of hepatocellular carcinoma at the same location but with the abdominal approach. We evaluated the effect of translocation of liver by monitoring the hepatic venous hemoglobin oxygen saturation. RESULTS: The outcome was favorable for the right thoracoabdominal approach for those patients with liver cirrhosis considering less reduction in frequency and degree of hepatic venous hemoglobin oxygen saturation during operation. CONCLUSIONS: The right thoracoabdominal approach may cause less damage to the liver with severe cirrhosis and preferable to this type of operation compare to the abdominal approach. PMID- 11268960 TI - Treatment of vascular complications following liver transplantation: multidisciplinary approach. AB - BACKGROUND/AIMS: Complications affecting the vascularization of the graft following orthotopic liver transplantation still represent a significant cause of graft loss and patient mortality. Strategies have recently been developed for the early detection and treatment of these complications before irreversible graft failure takes place. METHODOLOGY: A series of 429 consecutive liver transplants performed on 384 patients between April 1986 and December 1998 was retrospectively reviewed to assess the incidence of all the vascular complications and the results of their treatment with either surgery or interventional radiology. RESULTS: The incidence of vascular complications was 6.06% for the hepatic artery, 2.56% for the inferior vena cava and 1.16% for the portal vein. As regards anastomotic stenosis and thrombosis, the requirement of retransplantation decreased progressively with the advent of systematic postoperative screening with duplex Doppler ultrasonography and the introduction of graft-salvage procedures, falling from 50% for those cases diagnosed before 1996 to 19% for those diagnosed from 1996 on. Mortality following 18 graft salvage procedures was 11.1% versus 41.6% following retransplantation. Graft salvage procedures were successful in 14 out of 18 cases. CONCLUSIONS: Close surveillance of the vascular anastomoses and multidisciplinary approach to the treatment of vascular complication after liver transplantation considerably reduces graft loss and patient mortality. PMID- 11268961 TI - Usefulness of redox tolerance test in evaluating fatty liver. AB - BACKGROUND/AIMS: In living-related liver transplantation, fatty liver should be exactly detected in the healthy donor with noninvasive measurement before the surgery. The study aimed to investigate the usefulness of redox tolerance test in diagnosing fatty liver. METHODOLOGY: The subjects were 32 patients who underwent an abdominal surgery. They did not show any abnormal values in biochemical evaluations, nor had they diabetes. Under informed consent, liver specimens were obtained intraoperatively, and the subjects were divided into three groups according to the degree of hepatic fatty deposit: group A has fatty deposits at less than 10% of hepatocytes (n = 12), group B showed the deposits at 10-30% (n = 10), group C has the deposits of more than 30% (n = 10). Before the surgery, redox tolerance test was performed as follows; arterial blood samples were obtained successively at 75 g oral glucose load over a 120-min period, and the arterial ketone body ratio and blood glucose level were determined. The ratio of increased arterial ketone body ratio (AKBR) to increased blood glucose (BG) level (100 x delta AKBR/delta BG) was calculated as redox tolerance index. RESULTS: After fasting state, arterial ketone body ratio and blood glucose level did not differ among the three groups. However, the values of redox tolerance index in groups B (0.73 +/- 0.08) and C (0.46 +/- 0.04) were significantly lower than those in group A (1.85 +/- 0.31). CONCLUSIONS: The redox tolerance test was exceedingly sensitive indicator for objectively diagnosing the fatty liver. PMID- 11268962 TI - Cytokine response to human liver ischemia-reperfusion injury during hepatectomy: marker of injury or surgical stress? AB - BACKGROUND/AIMS: The aim of this study was to evaluate the inflammatory or antiinflammatory cytokine response to ischemia-reperfusion during hepatectomy and to find a useful marker of injury or surgical stress during hepatic ischemia reperfusion. METHODOLOGY: In 9 patients with liver disease who underwent hepatectomy using the Pringle maneuver, serum cytokines, including alanine transaminase, aspartate transaminase, and hyaluronic acid, were measured just prior to vascular occlusion; 5, 10 and 15 min after initial clamping; and 3 min after initial declamping. RESULTS: The mean concentrations of aspartate transaminase and alanine transaminase did not significantly differ before and after ischemia-reperfusion during hepatectomy. However, mean concentrations of hyaluronic acid after ischemia-reperfusion were significantly (P < 0.03) higher than before clamping. Although there were no significant differences in the mean concentrations of IL-1 beta, IL-6, IL-8, IL-10 and TNF-alpha among, before and after ischemia-reperfusion, the mean concentrations of granulocyte colony stimulating factor after ischemia-reperfusion and macrophage colony-stimulating factor after reperfusion were significantly (P < 0.05) higher than before clamping. CONCLUSIONS: Although hepatic parenchymal cell function was maintained after ischemia-reperfusion during hepatectomy, sinusoidal endothelial cell dysfunction was found. Release of granulocyte colony-stimulating factor and macrophage colony-stimulating factor after ischemia-reperfusion were also found. These cytokines and hyaluronic acid may be useful indicators in the early phase of human ischemia-reperfusion injury during hepatectomy. PMID- 11268963 TI - Hepatitis C virus infection: other biological fluids than blood may be responsible for intrafamilial spread. AB - BACKGROUND/AIMS: Several epidemiological studies have shown the existence of other routes of transmission of the hepatitis C virus besides the parenteral one, but the mechanisms involved are not yet understood. The general aim of this study was to evaluate the prevalence of hepatitis C virus infection in family contacts of infected patients and to analyze the possible risk factors and alternative transmission routes. METHODOLOGY: One hundred and thirty-eight relatives of 45 patients (index cases) affected by C virus-related chronic hepatitis were studied. The relatives were 45 spouses, 89 children and 4 cohabitants who underwent detection of serum anti-HCV antibodies; the anti-HCV-positive subjects were tested for serum HCV-RNA. The index cases, all the spouses and only other infected relatives were tested for the presence of HCV-RNA in saliva RESULTS: Antibodies to hepatitis C virus were detected in 5.7% of the family members while 11.1% of the analyzed spouses were serum HCV-RNA-positive. HCV-RNA was found in 44% of the examined saliva and 39% of these were found serum HCV-RNA-negative. The prevalence of hepatitis C virus among household contacts, excluding cases with previous parenteral exposure, was 3.6%. CONCLUSIONS: The epidemiological data on the intrafamilial spread of hepatitis C virus may be underestimated owing to the existence of infected relatives serum-negative but saliva-positive for the presence of the virus. The whole of these observations suggests a possible role of biological fluids in intrafamilial spread of hepatitis C virus. PMID- 11268964 TI - FR128998 ameliorates liver injury in extended liver resection with ischemia in dogs. AB - BACKGROUND/AIMS: Platelet-activating factor is one of the most potent phospholipid mediators associated with inflammatory conditions such as ischemia and reperfusion injury. FR128998 (FR) is a novel platelet-activating factor receptor antagonist. In this study, we evaluated the effect of FR during an extended liver resection with ischemia in a canine model. METHODOLOGY: Animals were divided into two groups: the control group (n = 8), and the FR-treated group (n = 7) in which FR was administered via the portal vein. While the right portal pedicle was clamped for 60 min, the left portal branch remained patent to avoid splanchnic congestion. Following reperfusion, 75% of the liver (including the right central, quadrate, left central, left lateral, and papillary lobes) was resected. Hepatic venous blood was collected to measure GPT, GOT, and LDH levels. Hepatic tissue blood flow was measured by a laser Doppler flow meter. The liver specimen was harvested for histological study. RESULTS: GPT, GOT, and LDH levels after reperfusion were significantly (P < 0.05) lower in the FR-treated group than in the control group. Hepatic tissue blood flow was maintained significantly (P < 0.05) higher in the FR-treated group than in the control group. Histologically, accumulation of polymorphonuclear neutrophils significantly (P < 0.05) decreased in the FR-treated group compared with the control group. The 2 day survival rate was statistically (P < 0.05) better in the FR-treated group than in the control group. CONCLUSIONS: FR128998 provides a protective effect for liver parenchyma and sinusoidal endothelial cells during extended liver resection with ischemia. PMID- 11268965 TI - Percutaneous ethanol injection in the treatment of hepatocellular carcinoma in cirrhosis. AB - BACKGROUND/AIMS: In this study we discuss and evaluate the use of percutaneous ethanol injection for the treatment of hepatocellular carcinoma in cirrhosis. METHODOLOGY: Percutaneous ethanol injection was performed under ultrasound guidance, with multiple sessions at an outpatient department or with "single session" technique under general anesthesia, according to the size and number of the lesions. RESULTS: In our patients with Child A (293), B (149), or C (20) cirrhosis and single hepatocellular carcinoma 5 cm or smaller, the 1-, 3- and 5 year survival rates were 98, 79 and 47%, 93, 63 and 29%, and 64, 12 and 0%, respectively. In our 108 patients with larger hepatocellular carcinoma, 1- and 3 year survival rates were: 72 and 57% in single, encapsulated tumors, 73 and 42% in single infiltrating or multiple encapsulated tumors, and 46 and 0% in symptomatic or with advanced portal thrombosis tumors. CONCLUSIONS: Percutaneous ethanol injection proved to be a safe, effective, repeatable, easy and low-cost therapy for hepatocellular carcinoma. Survival after percutaneous ethanol injection was comparable to that after surgical resection, probably because of a balancing between greater complete ablation rate of surgery versus absence of early mortality and liver damage of percutaneous ethanol injection. On the basis of the percutaneous ethanol injection rationale, other ablation techniques were proposed using radiofrequency, laser or acetic acid. Their initial results are promising. PMID- 11268966 TI - Surgical experience of hydatid disease of the liver: omentoplasty or capitonnage versus tube drainage. AB - BACKGROUND/AIMS: The aim of this paper was to report our experience and comparison of surgical treatment methods for hepatic hydatid cyst disease. METHODOLOGY: Between January 1990 and December 1998, 66 patients with hepatic hydatid disease in two centers were operated on. Patients were assessed by clinical examination, laboratory methods and ultrasonography and computed tomography and magnetic resonance. We also compared omentoplasty or cappitonage with external drainage with or without cyctectomy. RESULTS: Common pathology was solitary cysts and most of them were placed in the right lobe. Omentoplasty was performed for 35 cysts and cappitonnage for 36 and external drainage for 31 cysts. No operative mortality was reported. Patients with omentoplasty developed fewer complications and had a significantly shorter hospitalization than those with external drainage. CONCLUSIONS: Although omentoplasty seems to be the best possible surgical alternative for the radical treatment of hepatic hydatid cysts, the management of hydatid cysts should be flexible, taking into consideration a number of factors and variables. PMID- 11268967 TI - Hepatic arterial infusion of 5-fluorouracil for liver metastases from pancreatic carcinoma: results from a pilot study. AB - BACKGROUND/AIMS: Liver metastasis is a common progression of pancreatic carcinoma, but an effective chemotherapy has not been established. The purpose of this study was to examine the efficacy and safety of a hepatic arterial infusion of 5-FU in patients with liver metastasis from pancreatic carcinoma. METHODOLOGY: Thirteen patients were enrolled in a pilot study of a hepatic arterial infusion of 5-FU therapy. They received 5-FU for 5 days at a dose of 500 mg/m2/day by continuous hepatic arterial infusion every 4 weeks. RESULTS: One patient showed a partial response, while 6 showed no change. Of these 6 patients, 2 showed a minor response. The overall response rate was 8% (95% confidence interval: 0-22%). Nausea and vomiting were the most common types of toxicity. Three patients (23%) had hepatic arterial occlusion. There were no life-threatening toxicities or complications. The overall median survival time was 15.9 weeks. CONCLUSIONS: Hepatic arterial infusion of 5-FU in patients with liver metastasis from pancreatic carcinoma is tolerable but is minimally effective at this dose and schedule. The schedule of administration should be modified. PMID- 11268968 TI - Hepatic alpha-smooth muscle actin expression in hepatitis C patients before and after interferon therapy. AB - BACKGROUND/AIMS: The mechanism of hepatic fibrogenesis with chronic viral hepatitis is not well understood. Persistent activation of hepatic stellate cells is felt to play a role in the development of fibrogenesis and progression to cirrhosis. METHODOLOGY: We determined the expression of hepatic alpha-smooth muscle actin, a marker of hepatic stellate cell activation, in 29 patients with chronic hepatitis C and varying degrees of liver injury and fibrosis. In addition to a baseline evaluation, we assessed the effect of interferon therapy on alpha smooth muscle actin expression in 11 patients, including 6 with a sustained response to therapy. Specimens were evaluated by light microscopy for grade of inflammation and stage of fibrosis. Expression of alpha-smooth muscle actin was assessed semiquantitatively by immunohistochemical staining. RESULTS: At baseline, all patients had alpha-smooth muscle actin expressed within the liver without an obvious correlation with the severity of liver injury. However, among sustained responders, a reduction in hepatic necroinflammatory activity was associated with a trend towards a decrease in alpha-smooth muscle actin expression. This however did not reach statistical significance. CONCLUSIONS: Hepatic alpha-smooth muscle actin expression, as a marker of hepatic stellate cell activation appears reversible and tends to correlate with necroinflammatory activity. PMID- 11268969 TI - Mechanism of cold ischemia-reperfusion-induced graft injury after orthotopic liver transplantation in rats. AB - BACKGROUND/AIMS: The purpose of this study was to clarify the mechanism of cold ischemia-reperfusion-induced graft injury after liver transplantation, especially with regard to the relationship between hepatocyte, sinusoidal endothelial cell injury, and hepatic hemodynamic alteration. METHODOLOGY: We evaluated changes in hepatocyte and sinusoidal endothelial cell function, and hepatic hemodynamics after reperfusion in an isogeneic rat liver-transplantation model. The livers of male LEW rats were stored in 4 degrees C lactated Ringer's solution for 1 hr, 3 hr (viable graft), and 6 hr (nonviable graft) before implantation. After reperfusion, hepatocyte function was assessed by serum alanine aminotransferase level and bile output; sinusoidal endothelial cell function was evaluated by serum hyaluronic acid level. Furthermore, we measured hepatic venous oxygen saturation, and portal venous blood flow using a transit time blood flow meter. RESULTS: At 2 hr after reperfusion, the hepatocyte function was similar in all groups. However, the sinusoidal endothelial cell function deteriorated severely in the nonviable graft group, and significantly decreased hepatic venous oxygen saturation levels were observed, suggesting poor hepatic circulation. At 4 hr after reperfusion, the hepatocyte injury was markedly increased in the nonviable graft group. Although systemic blood pressure remained stable, significantly decreased portal venous blood flow in the nonviable graft group was found compared with the viable graft groups. Histopathological studies showed that massive ischemic necrosis was seen in zone III (central) of hepatic lobule 8 hr after reperfusion in the nonviable graft group. CONCLUSIONS: These data suggest that the sinusoidal endothelial cell injury was predominant in the early phase of reperfusion, and might cause microcirculatory disturbances, resulting in decreased portal venous blood flow. This phenomenon may subsequently cause ischemic damage to the hepatocyte, with eventual graft failure. PMID- 11268970 TI - The effect of granulocyte colony stimulating factor on the immune parameters in experimental obstructive jaundice. AB - BACKGROUND/AIMS: Obstructive jaundice is an important clinical problem. It may cause complications such as renal insufficiency, cardiovascular sequels, coagulation defects, gastrointestinal bleeding, delayed wound healing, secondary biliary cirrhosis and sepsis. We investigated the effect of GM-CSF on immunological parameters in the experimental obstructive jaundice. METHODOLOGY: In our experimental study we studied four groups that consisted of 28 rats. The 1st group consisted of sham rats, the 2nd group consisted of sham and GM-CSF applied rats and the 3rd and 4th groups consisted of rats that had obstructive jaundice. In the 4th group we applied 4 micrograms/kg GM-CSF subcutaneously to the rats for 7 days. We measured the levels of neutrophils, lymphocytes, leukocytes, interferon-alpha, CD32, CD34 and CD64 in all groups. RESULTS: In the jaundice group neutrophil, lymphocyte and leukocyte counts were found to be significantly lower compared to the other groups (P < 0.005). interferon-alpha and CD levels were found to be lower in the jaundice group compared to the other groups. In the GM-CSF applied jaundice group neutrophils, lymphocytes, leukocytes and interferon-alpha levels were found to be higher. CD34- CD64 levels were insignificantly increased in the GM-CSF group whereas CD32 levels were significantly increased. CONCLUSIONS: We believe that in the prevention of serious septic complications which have a high mortality risk, GM-CSF application to restore the macrophage-neutrophil functions should be supported by advanced clinical studies. PMID- 11268971 TI - Angiogenesis in hepatocellular carcinoma as evaluated by alpha smooth muscle actin immunohistochemistry. AB - BACKGROUND/AIMS: Angiogenesis has been known to be associated with tumor development. In this study, neovascularization in small hepatocellular carcinoma was investigated by evaluation of intratumoral arteriole counts, using alpha smooth muscle actin antibody immunohistochemistry. METHODOLOGY: Surgical specimens from 38 patients with small hepatocellular carcinoma were immunostained for alpha smooth muscle actin and proliferating cell nuclear antigen. The correlation between intratumoral arteriole density and clinicopathological factors including angiographic findings, proliferative activity, and patient prognosis were analyzed. RESULTS: Significant difference in intratumoral arteriole density were observed between well-differentiated hepatocellular carcinoma and poorly differentiated hepatocellular carcinoma (P = 0.004) or moderately differentiated hepatocellular carcinoma and poorly differentiated hepatocellular carcinoma (P = 0.011). The mean intratumoral arteriole count in the tumors showing angiographic hypervascularity was significantly higher than that in the tumors without angiographic hypervascularity (P = 0.011). A significant and positive correlation was found between proliferating cell nuclear antigen labeling index and intratumoral arteriole density (r = 0.5232, P = 0.001). A high intratumoral arteriole density in tumor was significantly correlated with shorter patients survival (P = 0.018). Cox's multivariate regression analysis showed that the intratumoral arteriole density was independent prognostic factors (P = 0.0306). CONCLUSIONS: Intratumoral arteriole density was found to be significantly associated with histological grade, proliferative activity, and patient survival. It also reflected the angiographic findings. Alpha smooth muscle actin antibody immunohistochemistry would provide a simple and biologically significant method which is usable to screen neovascularization and arterial blood supply in hepatocellular carcinoma, and may have predicting utility for patients outcome. This technique is applicable to routine paraffin sections, and may be useful as an adjunct to surgical pathology of hepatocellular carcinoma. PMID- 11268972 TI - Prognostic value of the modified TNM (Izumi) classification of hepatocellular carcinoma in 53 cirrhotic patients undergoing resection. AB - BACKGROUND/AIMS: Few studies have assessed the significance of prognostic factors in cirrhotic patients undergoing resection for hepatocellular carcinoma. METHODOLOGY: Overall survival and disease-free survival were evaluated in 53 cirrhotic patients undergoing hepatic resection for supervening hepatocellular carcinoma. The value of the UICC TNM classification, and the Izumi modified staging system on prognosis were analyzed. RESULTS: In multivariate analysis lack of micro/macrovascular invasion were predictive for long-term outcome. The difference between stages 1 and 2 or stage 3 and 4A using the UICC TNM classification was not significant with respect to survival or disease-free survival. UICC TNM classification was modified as follows; stage 1, solitary tumor without vascular invasion; stage 2, solitary or multiple tumor(s) involving adjacent vessel branch; stage 3, tumor(s) involving major vessel branch or with regional lymph node metastases; stage 4, tumor(s) with distant metastases. TNM (modified in accordance with Izumi) showed a significant difference between each stage with respect to survival and disease-free survival. CONCLUSIONS: A uniform tumor classification of hepatocellular carcinoma is advocated. Our results show that the UICC TNM classification for hepatocellular carcinoma is inadequate and may even on occasion lead to unnecessary resection. The modified staging system of Izumi is superior in determining outcome for cirrhotic patients with supervening hepatocellular carcinoma undergoing resection. PMID- 11268973 TI - Results of 22 years of experience in radical surgical treatment of hepatic hydatid cysts. AB - BACKGROUND/AIMS: As there is still no effective parasiticide, treatment of hydatid cysts continues to be surgical. The possibility of treatment by PAIR. (puncture-aspiration-instillation-reaspiration) or laparoscopy has intensified the debate on the need for radical surgery. This study aims to show that radical surgical resection of the hepatic hydatid cyst is a safe and very effective technique, based on our results after 22 years of experience. METHODOLOGY: Between 1974 and 1996 in 2 large Madrid hospitals we operated on 459 patients with 630 hydatid cysts. As technical advances and experience may vary results, patients were divided into 2 groups according to the period when they had undergone surgery: group A between 1974 and 1984; and group B between 1985 and 1996. Results of radical surgical resection and changes over the course of evolution of this technique were analyzed. RESULTS: A progressive drop was observed in morbidity and mortality. There were no deaths related to technical complications amongst total cystopericystectomy cases. Between 1990 and 1996 mortality was 0%, 2% of patients presented biliary fistula and 4% infection of the residual cavity. Mean hospital stay was 15.2 days. Only 1 patient of the 459 presented recurrence. CONCLUSIONS: As regards morbidity and mortality, technical advances and accumulated experience permit safe treatment of hepatic hydatid cysts by radical resection, with an almost nil recurrence rate. This makes it the technique of choice over others such as partial resection, PAIR or laparoscopy. PMID- 11268974 TI - Hepatic resection for liver metastasis of sigmoid colon cancer after incomplete percutaneous microwave coagulation therapy. AB - We report a case of colon cancer with liver metastasis that had been treated previously by sigmoidectomy and partial hepatic segmentectomy. A 55-year-old woman presented with two asynchronous liver metastases, which were treated with percutaneous microwave coagulation therapy. However, evaluation by dynamic computed tomography one week later showed incomplete necrosis in at least one tumor. Surgical resection was subsequently performed and histopathological examination showed the presence of viable cancer cells in both tumors. We conclude that surgical resection is perhaps the best curative method of treatment of metastatic liver tumors of colorectal carcinomas and that dynamic computed tomography is not always accurate for evaluating the effect of microwave coagulation therapy. PMID- 11268976 TI - Nonpercutaneous therapies of hepatocellular carcinoma. AB - Treatment of hepatocellular carcinoma depends largely on local resources, the stage of the disease and the presence of cirrhosis, but is limited overall by the lack of efficient chemotherapy. Hepatic resection is the treatment of choice for the few patients with hepatocellular carcinoma and normal liver. Five-year survival without recurrence in patients with a tumor of mean diameter 8 cm was 33%. Liver transplantation is the best chance for cure in patients with cirrhosis and a single small tumor, but its widespread application is limited by a number of obstacles, including cost. Tumor size and number, and liver status were common guidelines for selecting patients. Five-year survival of transplant patients was > 50%, compared to 0% in historical untreated controls. Patients with well preserved liver function and a small tumor at the periphery could equally benefit from hepatic resection, although cirrhosis entails the risk of morbidity due to portal hypertension and development of de-novo tumors. Another major drawback of hepatic resection is the early spread of tumor cells, facilitating early tumor recurrence after operation. For patients with compensated cirrhosis and a small tumor who were hardly eligible for surgery, transcatheter arterial chemoembolization appeared to be a cost-saving and effective treatment modality. Transcatheter arterial chemoembolization has been largely employed also for the palliative treatment of patients with large tumors, but the benefits on survival are doubtful. Conventional radiotherapy with external irradiation was not effective against hepatocellular carcinoma. PMID- 11268975 TI - Liver cystadenocarcinoma originating in cystadenoma without mesenchymal stroma. Therapeutic strategy in case of atypical radiological criteria. A case report. AB - Optimal treatment of cystadenoma if diagnosed consists of complete resection of the tumor. In case of atypical radiological criteria, therapeutic strategy is not well defined. The attitude we adopt is to regularly monitor the lesion. Surgical removal of the tumor is of course indicated facing any significant change on sonography or tomodensiometry. PMID- 11268978 TI - Hepatic angiomyolipoma with concomitant hepatocellular carcinomas. AB - Angiomyolipoma is a rare lipomatous tumor in the liver. Definitive preoperative diagnosis is becoming easier by the use of ultrasonography, computed tomography, and magnetic resonance imaging techniques. Nonsurgical treatment has been advocated for its benign nature. However, recently we encountered one case of hepatic angiomyolipoma with two concomitant hepatocellular carcinomas on a hepatitis B carrier. Although his serum alpha-fetoprotein was normal, under the above impression these lesions were resected. The pathologic findings showed a typical angiomyolipoma and two well-differentiated hepatocellular carcinomas with marked fatty metamorphosis. This is the first report of angiomyolipoma with concomitant hepatocellular carcinomas in the literature. Nonsurgical treatment of angiomyolipoma in an endemic area for hepatocellular carcinoma should proceed with caution because cases of fat-rich minute hepatocellular carcinomas will make the diagnosis difficult. PMID- 11268977 TI - Clinical features and imaging diagnosis of biliary cystadenocarcinoma of the liver. AB - Biliary cystadenocarcinoma of the liver is a relatively rare disease. Herein, we reported a case of biliary cystadenocarcinoma with a review of the literature. A 71-year-old female was admitted with the chief complaint of epigastralgia. The imaging studies revealed a biliary cystadenocarcinoma in the left hepatic lobe with suspicion of direct invasion to the left and middle hepatic veins and inferior vena cava. However, there was no direct invasion of the tumor to these veins in operation findings, and an extended left hepatic resection was performed without resection of inferior vena cava. The tumor was histologically diagnosed as biliary cystadenocarcinoma of the liver. Diagnosis of biliary cystadenocarcinoma is usually difficult preoperatively, however, a diagnosis was possible with the use of imaging studies. It was suggested that this tumor originated from a benign cystadenoma because of the existence of a transitional zone between normal cells and atypical cells in the cystic wall. Systematic hepatectomy was recommended as the initial treatment in consideration of the features of cystadenocarcinoma. PMID- 11268980 TI - Liver transplantation using a right lateral sector graft from a living donor to her granddaughter. AB - One of the major concerns regarding living-related liver transplantation is graft size disparity. The left liver graft is too small while the right is too large in some recipients. To overcome this problem, the right lateral sector (Segments VI and VII) was transplanted from a living donor (55 kg) to her granddaughter (17 kg). The common hepatic trunk had to be anastomosed end-to-end to the graft hepatic vein without being compressed by the graft overriding the vena cava and without unfavorable tension of the anastomosis. The anterior wall of the hepatic vein of the donor was resected as much as possible. The superficial left, left, middle and right hepatic veins of the recipient were made confluent by incision of the intervening venous walls, and the nicks were sutured to form a wide and long common venous trunk. The recipient received a graft corresponding to 75% of her standard liver volume. She was complicated with gastric dilation and acute rejection, but recovered with no signs of anastomotic stricture. Right lateral sector graft obtained by this innovative procedure may be useful for overcoming borderline graft-recipient size and shape differences. PMID- 11268979 TI - Transfusion transmissible virus TTV and its putative role in the etiology of liver disease. AB - TTV, the transfusion transmissible hepatitis virus infects mainly patients at risk for parenteral exposure and hence, prone to develop chronic liver disease, as well as healthy populations worldwide. Most TTV infections appear to occur parenterally, with viremia detected frequently in blood donors and blood products. The substantial proportion of asymptomatic individuals never exposed to blood-borne agents, and its high prevalence among healthy subjects implicates the fecal-oral route as another potential for transmission. According to the TTV DNA levels detected in liver tissue, it apparently replicates in hepatocytes, and TTV DNA is present in sera of patients with posttransfusion hepatitis of unknown etiology closely correlated with ALT levels. However, TTV initiating the development of chronic liver disease or causing posttransfusion hepatitis could not be confirmed, as most patients positive for TTV DNA remain asymptomatic and those progressing towards chronic liver disease are invariably coinfected with either the hepatitis B or C virus. Also, TTV coinfection does not aggravate the symptoms associated with hepatitis B or C. Similarly, it does not cause posthepatitis aplastic anemia, and high-risk patients can immunologically clear the viral DNA. In conclusion, being widely distributed and apparently nonpathogenic, TTV might represent an opportunistic but innocent virus reminiscent of hepatitis G virus, with a negligible role in the etiology of chronic liver disease. PMID- 11268981 TI - Practical guidelines for the preservation of the pancreaticoduodenal arteries during duodenum-preserving resection of the head of the pancreas: clinical experience and a study using resected specimens from pancreaticoduodenectomy. AB - BACKGROUND/AIMS: The purpose of this study was to create a practical guideline for vascular preservation during duodenum-preserving resection of the head of the pancreas. METHODOLOGY: We examined the anatomy of pancreaticoduodenal arteries by specimen angiography and dissection using 12 pancreaticoduodenectomy specimens. We also reviewed our experiences with duodenum-preserving resection of the head of the pancreas. RESULTS: In the specimens, the posterior pancreaticoduodenal artery and its duodenal branches were easily separated from the posterior surface of the pancreas, and its papillary branch was identified in two-thirds of the cases. It was difficult to dissect the anterior superior pancreaticoduodenal arteries from the pancreas because they were partially buried in the pancreatic parenchyma. The anterior inferior pancreaticoduodenal artery located in the posterior and inferior surface of the pancreas could be safely dissected in two thirds of the cases. Duodenum-preserving resection of the head of the pancreas was performed in 7 patients. In every case, the anterior superior pancreaticoduodenal artery was sacrificed and the anterior inferior pancreaticoduodenal artery was preserved. In 3 cases, the entire posterior pancreaticoduodenal artery was preserved and in 4 cases a short segment of the posterior pancreaticoduodenal artery was removed accidentally. The pancreatic head was totally removed and the intrapancreatic common bile duct was preserved. There were 3 postoperative complications, pancreatic leakage, intraabdominal fluid collection and bile duct stricture. They improved with conservative management. CONCLUSIONS: To safely perform duodenum-preserving resection of the head of the pancreas, preservation of the whole posterior pancreaticoduodenal artery and anterior inferior pancreaticoduodenal artery is recommended because they can be safely dissected from the pancreas, and the posterior pancreaticoduodenal artery provides the major blood supply to the papilla and distal bile duct. However, removal of a short segment of posterior pancreaticoduodenal artery does not preclude a good blood supply to the duodenum because of bidirectional blood flow. PMID- 11268983 TI - A case report of cribriform carcinoma of the pancreas with long time survival. AB - A 62-year-old man had complained of left abdominal pain and tenderness and a body weight loss. Abdominal ultrasonography and computed tomography revealed homogeneous tumor with clear margin, and an irregular shape (3.5 x 2.0 cm) in the body of the pancreas. Endoscopic retrograde cholangiopancreatography showed a shadow defect in the main pancreatic duct adjacent to the tumor, which suggested intraductal tumor spread. Distal pancreatectomy with splenectomy and left paraaortic lymph node dissection was performed. Microscopically, the tumor showed microtubular carcinoma, which was characterized by a cribriform pattern, medullary growth, and little interstitium. The tumor was encapsulated by a relatively thick fibrous capsule. The patient was discharged uneventfully, and he is alive 33 months after operation without a distinct sign of recurrence. In conclusion, cribriform carcinoma of the pancreas has specific characteristics, such as good prognosis, expansive growth with little invasion, intraductal growth and spread without mucin production and histological marked cribriform pattern. This type of carcinoma should be classified as a new disease entity of carcinoma of the pancreas. PMID- 11268982 TI - Surgery for chronic obstructive pancreatitis: comparison of end-to-side pancreaticojejunostomy with pancreaticoduodenectomy. AB - BACKGROUND/AIMS: Chronic obstructive pancreatitis usually manifests with intractable pain and recurrent episodes of chronic pancreatic inflammation. The side-to-side pancreaticojejunostomy is used for those patients with a large pancreatic duct. But for the patients with small pancreatic duct, the optimal surgical procedure needs to be evaluated. A prospective study was designed to compare the different results between distal pancreatectomy plus end-to-side pancreaticojejunostomy and pancreaticoduodenectomy. METHODOLOGY: The patients were chosen prospectively and randomly to undergo either a distal pancreatectomy plus end-to-side pancreaticojejunostomy or pancreaticoduodenectomy in the last 3 years. Eighteen patients with chronic obstructive pancreatitis were randomly divided into two groups. Ten patients (group A) underwent distal pancreatectomy plus end-to-side and ductal to mucosal pancreaticojejunostomy, and the other 8 patients (group B) underwent pancreaticoduodenectomy were compared. RESULTS: The operative time was significantly shorter and operative blood loss was less in group A. The postoperative follow-up of patients in group A had better outcome with increased body weight and no steatorrhea or diabetes mellitus, and all of them had complete pain relief. CONCLUSIONS: We concluded that distal pancreatectomy with end-to-side pancreaticojejunostomy provided a better surgical treatment for the patients with chronic obstructive pancreatitis and small pancreatic duct. PMID- 11268984 TI - A new embryologic hypothesis of annular pancreas. AB - Annular pancreas is a developmental anomaly of the pancreas. There are two major hypotheses concerning development of the annular pancreas from the ventral pancreatic anlage; adhesion of the right ventral anlage to the duodenal wall (Lecco's theory), and persistence of the left ventral anlage (Baldwin's theory) reported in 1910, but each theory has some problems and can account for only a few types of annular pancreas. We report a new embryologic hypothesis of annular pancreas which can account for the developmental mechanism of three types of arrangement of annular ducts. The tip of the left ventral anlage adheres to the duodenum and stretches to form a ring. Whether the tip is proximal or distal to the bile duct creates several arrangements of the annular duct. PMID- 11268985 TI - Comparison of the UICC/AJCC 1992 and 1997 pN categories for gastric cancer patients after surgery. AB - BACKGROUND/AIMS: UICC/AJCC 1997 classification changes pN category. We evaluated its prognostic impact. METHODOLOGY: A total of 710 patients who underwent a > or = D2 gastrectomy were recruited. Among them, the data of 319 patients who had involved regional lymph nodes and no evidence of distant metastases were used for comparing the 1992 and 1997 pN categories. RESULTS: For 1997 category, 201 patients (64%) were pN1, 75 (23.5%) pN2, and 43 (13.5%) pN3. For 1992 category, 143 patients (44.8%) were pN1, and 147(46.1%) pN2. 29 patients (9.1%) with lymph node metastasis to the hepatoduodenal ligament were distant metastasis. The 1997 pN category was a more powerful prognostic discriminant (relative risk: 2.086) than the 1992 category. Compared to the 1992 stage classification, the 1997 one had a skewed distribution of patients with marked shift of patients of stage IIIA (105-126 patients), IIIB (116-58 patients), and IV (100-122 patients). The survival difference between stage IIIA and IIIB for the 1997 stage classification is narrower than for 1992. CONCLUSIONS: The 1997 pN category allows for estimation of prognosis superior to the 1992 category. PMID- 11268987 TI - Clinical advances in primary liver cancer in China. AB - Primary liver cancer is one of the most common cancers in China. According to the statistics of our country, primary liver cancer claims 20.40 lives per 100,000 people annually, with 19.98 per 100,000 in cities and 23.59 per 100,000 in rural areas, ranking as the 2nd and the 1st leading cause of cancer death respectively. Of all the newly enrolled cases in the world each year, 45% are found in the mainland of China. In the southeast areas of high incidence, the situation is even worse with tumors tending to occur in a younger age group. In this paper, we summarized our 40-year experience and progress in diagnosis and treatment of primary liver cancer. PMID- 11268986 TI - Palliative gastrectomy for advanced gastric cancer. AB - BACKGROUND/AIMS: Although palliative gastrectomy for advanced gastric cancer may be favorable in selected patients presenting with bleeding and obstruction, little has been reported about the clinical significance of palliative gastrectomy, including prognosis. METHODOLOGY: A retrospective comparison between 84 patients with palliative gastrectomy (PG group) and 100 patients with unresectable operation (UO group) for advanced gastric cancer was carried out. RESULTS: The incidence of serosal invasion, peritoneal dissemination, hepatic and lymph node metastases, and undifferentiated tissue type in the UO group were significantly higher than in the PG group. Median survival after operation in the PG group (20.6 months) was significantly longer than in the UO group (5.7 months). Also, in stage IVb patients, median survival time in the PG group (10.2 months) was significantly longer than in the UO group (5.0 months). However, median survival in the patients with synchronous liver metastasis between PG (8.4 months) and UO (4.6 months) groups was not significantly different. Survival rates after operation of 6 months, 1 year and 2 years in all patients between the palliative gastrectomy group versus UO group were 83.6% versus 38.3% (P < 0.01), 63.0% versus 9.3% (P < 0.01) and 35.2% versus 0% (P < 0.01), respectively. CONCLUSIONS: Palliative gastrectomy compared to unresectable operation may be effective for improvement of prognosis even if stage IVb patients with peritoneal dissemination and/or distant lymph node metastasis. However, it may be unfavorable on survival of patients with synchronous liver metastasis. PMID- 11268988 TI - Prediction of survival period for patients with postoperative recurrence after curative resection for advanced gastric carcinoma. AB - BACKGROUND/AIMS: Although many studies have attempted to clarify the prognostic indicators for gastric carcinoma, there have been few studies regarding the factors that correlate with the survival period of patients with postoperative recurrence. METHODOLOGY: Among 504 advanced gastric adenocarcinoma patients who had undergone curative gastrectomy, 188 patients who had died of recurrence were used in this study. RESULTS: Univariate analysis indicated that age, the presence of lymph node metastasis and blood vessel invasion, the number of positive lymph nodes, and gastrectomy significantly correlated with the survival period. Multivariate analysis indicated that the length of the survival period was independently influenced by the number of positive lymph nodes and blood vessel invasion. The survival time of patients with less than 3 positive lymph nodes and no accompanying blood vessel invasion was significantly longer than that of other patients. CONCLUSIONS: The number of positive lymph nodes and the presence of blood vessel invasion are the most important factors predicting the survival period of patients with postoperative recurrence after curative resection for advanced gastric carcinoma. PMID- 11268989 TI - Prognostic factors in early gastric cancer. AB - BACKGROUND/AIMS: Opportunities of observing patients with recurrent early gastric cancer are rare. Assessment of prognostic factors for the recurrence of early gastric cancer is important for determining adequate strategies for managing early gastric cancer. METHODOLOGY: Clinicopathologic variables were compared in 8 patients with and 453 without recurrence who were followed for over 5 years after curative resection. Expression of mutant p53 and Ki-67 was evaluated in 16 patients with n2 or above. RESULTS: Eight patients died of gastric cancer with recurrence. There were no inter-group differences in mean diameter, histologic classification, patterns of infiltrating growth, or cancer-stroma relationship; but macroscopic appearance, depth of invasion, lymphatic invasion, venous invasion and lymph node metastases were significantly more frequent on univariate analysis in those with recurrence. Lymph node metastases was an independent prognostic factor by the Cox proportional hazards regression model. In patients with n2 or above, mutant p53 expression was higher in recurrent than in nonrecurrent cases. CONCLUSIONS: Lymph node metastasis was the only independent prognostic factor for the recurrence of early gastric cancer. The expression of mutant p53 may be an indicator of recurrence in patients with n2 or above. PMID- 11268990 TI - Gastric emptying rate for solid and for liquid test meals in patients with dyspeptic symptoms after partial gastrectomy and after vagotomy followed by partial gastrectomy. AB - BACKGROUND/AIMS: Gastric emptying rate for solid and for liquid test meals was investigated retrospectively in patients with longstanding epigastric distress after partial gastrectomy, either as primary treatment or after failure of vagotomy for peptic ulcer in order to find an explanation for the postoperative symptoms. METHODOLOGY: Radionuclide-labeled liquid and solid test meals were used to evaluate gastric emptying rate, at least one year after surgery. RESULTS: The lag phase for liquid test meals disappeared in all operated patients. Partial gastrectomy usually lead to fast emptying but this resective procedure, if performed after vagotomy, lead to stasis in a significant number of patients. Gastric emptying rate for solids increased in only a few of these symptomatic patients. In most of them however, there was a normal to decreased emptying rate. If a vagotomy had preceded the resective procedure, gastric emptying rate decreased significantly. CONCLUSIONS: In all these symptomatic patients, gastric emptying had been disturbed for at least one type of test meal. This makes investigation for both meals necessary, especially since there is a lack of correlation. Furthermore, if vagotomy fails to prevent ulcer recurrence, one should carefully consider all options before performing partial gastrectomy since gastric emptying rate after these consecutive procedures worsens considerably. PMID- 11268992 TI - Endoscopic ultrasonography features of calcified gastric cancer. AB - Calcifications are a rare finding described in benign and malignant tumors located in any site of the body. Their presence in stomach and colon carcinomas is very rare. Most of the cases described are mucinous adenocarcinomas. We present the case of one patient with this disease studied with endoscopic ultrasonography. There were punctate calcifications in the submucosa layer that tended to take on a crown-like shape in the outer-most area, producing an acoustic shadow. The pathological study of the surgical specimen showed amorphic calcifications inside some mucin lakes. More cases need to be studied with this technique in order to define their endosonographic characteristics. PMID- 11268991 TI - Therapy of HCC--TACE for liver tumor. AB - Nowadays, TACE is routinely performed in a subgroup of patients with hepatocellular carcinoma who are not eligible for surgical resection or percutaneous tumor ablation and who do not preset contraindications to locoregional treatment. When comparing randomized to nonrandomized studies, TACE efficacy upon survival is still controversial in this subgroup despite a 40-50% response rate. Nevertheless, in a pragmatic approach, and as long as other therapies still fail in this subgroup, TACE can be recommended in Okuda I or II patients, the sessions being prolonged after the second one only for the responders who do not present severe side effects. PMID- 11268993 TI - Resection of hepatocellular carcinoma. AB - The majority of hepatocellular carcinoma occurs in patients with liver cirrhosis. Although the tumors are often discovered at an early stage during surveillance of these patients, the underlying cirrhosis renders the surgery more difficult and exposes the patients to higher rates of postoperative morbidity and mortality than occurs in other types of liver surgery. Over the past 20 years surgeons have developed new surgical procedures and techniques to firstly reduce the unnecessary resection of liver parenchyma and to decrease intraoperative blood loss. Better patient selection and understanding of prognostic factors will hopefully result in a further decrease in operative risk and postoperative recurrence. Adjuvant chemotherapy may prove effective in reducing the postoperative recurrence but at this stage surgery still remains as the best treatment for patients with recurrent tumor which is accessible to resection. PMID- 11268994 TI - Minimally invasive approaches to hepatic surgery. AB - During the last 10 years we have seen dramatic changes in minimally invasive surgery. Important advancements in videoscopy, instrumentation, and surgical skills have allowed us to perform surgical procedures recently viewed as well outside the domain of minimally invasive surgeon. Solid organ surgery has become common place; in particular, hepatic surgery has been shown to be amenable to minimally invasive techniques. Benign cyst excision, abscess drainage, local tumor ablation, and liver resection are all hepatic procedures described in recent literature. Though it is clear that the trend toward less invasive alternatives towards open liver surgery will continue, proper indications and utilization of such procedures requires careful scrutiny using outcome oriented research. This paper gives a brief overview of the technology available and its application in minimally invasive approaches to liver surgery. PMID- 11268995 TI - Laparoscopic radiofrequency of hepatocellular carcinoma (HCC) in liver cirrhosis. AB - BACKGROUND/AIMS: In this report, the feasibility and efficacy of laparoscopic radiofrequency interstitial thermal ablation of hepatocellular carcinoma has been evaluated in 20 patients. METHODOLOGY: The laparoscopic approach with the use of intraoperative ultrasonography allowed us to obtain additional information regarding liver nodules and a complete treatment of the liver lesions. RESULTS: The complication rate was low and there was no operative mortality. A complete necrosis has been obtained in 90% of the cases at 1 month dynamic computed tomography following the treatment. CONCLUSIONS: Laparoscopic radiofrequency thermal ablation of hepatocellular carcinoma proved to be a safe and effective technique; its use may be proposed in selected patients. Larger series are needed to accurately assess its role among the other ablative therapies of hepatocellular carcinoma. PMID- 11268996 TI - Liver resection in advanced hepatocellular carcinoma. AB - BACKGROUND/AIMS: The aim of this study was to assess the results of major liver resection in patients with advanced hepatocellular carcinoma in terms of safety and survival. METHODOLOGY: The subjects of this study are 19 patients that underwent 24 resections for advanced (stage IV) hepatocellular carcinoma. Eighteen of these resections were performed for primary tumor and 6 were repeat resections. Nine patients presented without cirrhosis, 5 with cirrhosis, and 5 patients had the fibrolamellar variant of hepatocellular carcinoma. RESULTS: Hospital mortality was recorded in 1 case (5%). Morbidity was noted in 7(37%) cases. All patients with fibrolamellar variant of hepatocellular carcinoma are alive at 78, 41, 24, 12 and 9 months (P = 0.008), compared with a median survival of 18 and 9 months for the noncirrhotic hepatocellular carcinoma and cirrhotic hepatocellular carcinoma groups, respectively (P = 0.24). CONCLUSIONS: We conclude that an aggressive policy of major liver resection with vascular reconstruction was justifiable in patients with advanced fibrolamellar variant of hepatocellular carcinoma and in selected patients with noncirrhotic hepatocellular carcinoma, and of doubtful value in patients with cirrhosis. PMID- 11268997 TI - Cholangiocarcinoma and the role of radiation and chemotherapy. AB - Cholangiocarcinoma is a rare tumor. Many cases are localized while metastatic disease within the liver and abdomen do occur. There is as yet no standard therapy for advanced bile duct tumors. Radiotherapy and chemotherapy are not curative modalities in this condition but are being assessed adjuvantly following surgery, and as palliative treatment in an attempt to either extend progression free and overall survival or to palliate symptoms. Advances may be made by: (i) The combined use of radiation and chemotherapy, (ii) High dose conformal radiotherapy, (iii) Novel chemotherapeutic agents. Patients should be given the opportunity to participate in clinical trials. PMID- 11268998 TI - Surgery for cholangiocarcinoma. AB - Cholangiocarcinoma is a cancer arising from bile duct epithelium and commonly occurs in the main bile duct or at the bile duct confluence. The patients present obstructive jaundice and often have advanced disease. Treatment in the past has frequently consisted of palliative measures aimed at relieving jaundice either by surgical bypass or by endoscopic or percutaneous drainage usually in combination with stenting. A better understanding of the ways of invasion of the pathology of cholangiocarcinoma together with improvements in surgical techniques and perioperative management have lead to an increase in the number of patients in whom resection may be contemplated. Resection offers the only chance of cure and the best chance of long-term survival. Current problems associated with resection of hilar cholangiocarcinoma are discussed in this review article. PMID- 11268999 TI - Endoscopic stenting for definitive treatment of irretrievable common bile duct calculi. A long-term follow-up study of 49 patients. AB - BACKGROUND/AIMS: The value of endoprostheses for long-term management of bile duct stones has not been formally established. A prospective evaluation of results and complications of the insertion of biliary endoprostheses was performed in patients with endoscopic irretrievable bile duct stones. METHODOLOGY: From January 1990 to September 1998, 49 patients (18 men and 31 women; average age 75.5 years), underwent endoscopic biliary stenting for endoscopically irretrievable bile duct stones. RESULTS: Successful biliary drainage was achieved in all patients. Early complications occurred in 12.2% of cases. Over the long-term follow-up (median follow = 39 months) late complications occurred in 40.8% of cases, with 3 cases of biliary-related death. CONCLUSIONS: For immediate bile duct drainage, endoprostheses proved a safe and effective alternative for treatment of patients with endoscopically irretrievable bile duct stones. Because of the risk of subsequent complications, its use as a definitive treatment should be confirmed to highly selected cases. PMID- 11269000 TI - Diagnostic workup before laparoscopic cholecystectomy--which diagnostic tools should be used? AB - BACKGROUND/AIMS: A prerequisite for successful laparoscopic cholecystectomy is the exclusion of potential risks such as cholangiolithiasis, anatomical malformations or diseases of the stomach. As there is no general agreement regarding the appropriate preoperative diagnostic workup, we compared different diagnostic methods as to their value in detecting unknown accompanying diseases and complications. METHODOLOGY: Between 9/90 and 8/93, we performed 850 laparoscopic cholecystectomies. The first 700 were included in this study. A prospective comparison was carried out of the diagnostic accuracy of history, physical examination, laboratory tests, upper gastrointestinal endoscopy or barium meal, i.v. cholangiography and abdominal ultrasound. RESULTS: Measurement of the diameter of the common bile duct was found to be a good noninvasive method for diagnosing common bile duct stones (sensitivity 80%, specificity 99%). In combination with the history and the laboratory tests the sensitivity could be improved to 99%. The sensitivity of i.v. cholangiography in detecting common bile duct stones was 80%, the specificity 99.3%. 646/700 patients underwent preoperative endoscopy/barium meal. In 53 (8.2%) patients pathological findings were found, but only in 4 cases (0.6%) they influenced the indication for laparoscopic cholecystectomy. In 1 patient an advanced gastric cancer was diagnosed 6 months after laparoscopic cholecystectomy, the preoperative barium meal did not show any pathological findings. CONCLUSIONS: The results show that routine ultrasonography in combination with history and laboratory tests prior to laparoscopic cholecystectomy can be recommended for detecting common bile duct stones. In patients with 1 or more pathologic finding endoscopic retrograde cholangiopancreatography should be performed preoperatively. A gastroscopy should be done in patients with nonspecific upper abdominal pain, history of peptic ulcer disease and persisting pain after laparoscopic cholecystectomy. PMID- 11269001 TI - The immune response to alkaline phosphatase and other immunogens in patients with primary biliary cirrhosis. AB - BACKGROUND/AIMS: Primary biliary cirrhosis is a potentially lethal hepatobiliary disorder in which 90% of the patients are women. Histologically, the disease is characterized by a progressive destruction of intrahepatic bile ducts by autoreactive T lymphocytes. Although the underlying etiology remains unknown, potential hypotheses must take into account; a) the predilection of the disease for women of childbearing age, b) the frequent coexistence of bone and intestinal involvement, and c) the high prevalence of autoantibodies directed towards intracellular enzymes. With these considerations in mind, we hypothesized that exposure to P-ALP (placental alkaline phosphatase) during pregnancy results in autoreactivity directed towards all human tissues harboring the ALP enzyme (liver, bone and intestine) in genetically predisposed individuals. METHODOLOGY: To test this hypothesis, we stimulated peripheral blood mononuclear cells of primary biliary cirrhosis patients (n = 17) cholestatic liver disease controls (n = 6) and healthy controls (n = 14) with P-ALP, polyclonal activators (phytohemagglutinin [PHA], anti-CD3) and recall antigens (tetanus toxoid, streptokinase). We then determined their proliferative and cytokine responses by 3H-thymidine incorporation and ELISA assays for Il-10, IL-6, tumor necrosis factor-alpha and interferon-gamma, respectively. RESULTS: The results of the study revealed that the proliferative response to P-ALP was similar in peripheral blood mononuclear cells from primary biliary cirrhosis patients, cholestatic and healthy controls. Although the proliferative responses to PHA (P < 0.001) and anti-CD3 (P < 0.001) were decreased in peripheral blood mononuclear cells from primary biliary cirrhosis patients when compared to both control groups, responses to the recall antigens; tetanus toxoid and streptokinase were similar in the three groups. Cytokine production following exposure to P-ALP, polyclonal activators or recall antigens in peripheral blood mononuclear cells from primary biliary cirrhosis patients was similar to that of cholestatic and healthy controls. CONCLUSIONS: The results of the above experiments suggest that P-ALP is unlikely to be the target autoantigen in primary biliary cirrhosis. The results also support the findings of other investigators that primary biliary cirrhosis patients have suppressed proliferative responses to polyclonal stimulation. PMID- 11269002 TI - Human neutrophil functions in obstructive jaundice. AB - BACKGROUND/AIMS: The effect of obstructive jaundice on neutrophil function has not been extensively studied. Therefore, the present study aimed at evaluating the effect of obstructive jaundice on human neutrophils. METHODOLOGY: Twelve patients with obstructive jaundice due to common bile duct obstruction underwent endoscopic biliary drainage. Neutrophil functions (chemotaxis and superoxide anion generation) were evaluated before and 7 days after drainage. RESULTS: Neutrophil chemotaxis in response to FMLP (formyl-methionyl-leucyl-phenylalanine) or interleukin-8 was abnormally increased before drainage, and was normalized after drainage. Similarly, enhanced superoxide anion generation in response to FMLP or phorbol myristate acetate before drainage was alleviated after drainage. CONCLUSIONS: The results suggest neutrophil overactivity in patients with obstructive jaundice. The ameliorating effect of biliary drainage on neutrophil overactivity might play a role in the prevention of postoperative complications. PMID- 11269003 TI - Laparoscopic cholecystectomy: what are the factors determining difficulty? AB - BACKGROUND/AIMS: The experience in laparoscopic cholecystectomy is increasing, more difficult cases are performed even by the junior surgeons. This policy has led the investigators to look for methods to identify potentially difficult laparoscopic cholecystectomy. METHODOLOGY: A prospective study was performed to find out the factors that determine technical difficulty in laparoscopic cholecystectomy. Two hundred and twenty-seven patients (170 females and 57 males) with symptomatic gallbladder stones were recruited for this study from June 1995 to September 1997. The overall difficulty score as a dependent variable was based on the following operative parameters: duration of surgery, bleeding, dissection of gallbladder wall, adhesions, spillage of bile, spillage of stones and difficulty of gallbladder extraction. Multiple regression analysis was used to assess the significance of the following potential difficulty variables (Independent) in predicting the variation in the overall difficulty score: age, sex, body mass index, gallbladder size, common bile duct diameter, gallbladder wall thickness, liver size and the presence of adhesions. RESULTS: Gallbladder wall thickness, presence of adhesions, liver size and gallbladder size were found to be significant predictors of the variation in overall difficulty score (Adjusted R2 = 0.48). CONCLUSIONS: We believe that the above-mentioned four difficulty factors are important, however, experience of the surgeon plays a major role in reducing difficult laparoscopic cholecystectomy and consequently decreasing it's complications. PMID- 11269004 TI - Portal vein thrombosis associated with hilar bile duct carcinoma and liver abscess. AB - As most portal vein occlusion in hilar bile duct carcinoma is caused by tumor invasion to the portal vein, other mechanisms of its occlusion are very rare. We report the case of a 69-year-old man who underwent surgical resection for an advanced hilar bile duct carcinoma associated with unusual portal vein occlusion. Preoperative diagnosis was advanced hilar bile duct carcinoma with liver abscess and right portal vein occlusion due to tumor invasion. Extended right hepatectomy combined with resection of caudate lobe was performed. Intraoperatively, tumor invasion to the portal vein was not evident and resected margin of the right portal vein showed thrombosis and no evidence of malignancy histologically. To our knowledge, this is the first reported case of a patient with a combination of portal vein thrombosis and liver abscess in hilar bile duct carcinoma. Although portal vein occlusion due to thrombosis is an unusual complication in hilar bile duct carcinoma, the presence of liver abscess may be a useful diagnostic implication of this occlusion. PMID- 11269006 TI - Benign biliary stricture associated with atherosclerosis. AB - We report a case of benign bile duct stricture that could not be differentiated from intrahepatic bile duct carcinoma preoperatively. The patient was a 79-year old man. Computed tomography showed dilatation of the intrahepatic bile duct in the left lobe. Direct cholangiography showed segmental stricture of the left bile duct. Angiography showed narrowing of the left hepatic artery. Although bile cytology did not show malignant cells, we suspected intrahepatic bile duct carcinoma preoperatively. We performed extended left hepatic lobectomy. Histopathologic examination of the resected duct also showed no malignant cells; fibrosis with infiltration by lymphocytes was seen at the bile duct stricture. In addition, the resected liver specimen showed sclerotic change in the intrahepatic arteries. The postoperative course was uneventful for more than 26 months, without recurrence or cholangitis. We encountered a very rare case of benign segmental bile duct stricture, which was difficult to differentiate from bile duct carcinoma. We think the biliary stricture was secondary to atherosclerosis which may have caused bile duct ischemia. PMID- 11269005 TI - Combination of interventional therapies in hepatocellular carcinoma. AB - Many interventional techniques aimed at achieving nonsurgical ablation of hepatocellular carcinoma have been developed and clinically tested over the last decade. Percutaneous image-guided therapies such as ethanol injection and radiofrequency thermal ablation provide an effective means for treating hepatocellular carcinoma lesions smaller than 3 cm, but do not ensure successful ablation of larger tumors. In view of the limitations of available interventional therapies, there is currently a focus on a multimodality strategy for the treatment of large hepatocellular carcinomas. Combination of transcatheter arterial chemoembolization and ethanol injection overcomes the weakness of each of the two procedures, enhancing local therapeutic effect and long-term survival. More recently, a new technique for single-session ablation of large hepatocellular carcinoma lesions has been devised by combining transcatheter hepatic arterial balloon occlusion/embolization and radiofrequency treatment. This combined approach substantially increases the thermal necrosis volume that can be created with respect to the conventional radiofrequency technique, as a result of the reduction of heat loss caused by convection. In a pilot multicentric clinical trial performed in 62 patients, successful ablation of hepatocellular carcinoma lesions ranging 3.5-8.5 cm in diameter was achieved in 82% of cases in the absence of major complications. This new technique seems to have the potential to replace other interventional methods for the treatment of large hepatocellular carcinoma. PMID- 11269007 TI - Hepatic resection for metastatic tumors from noncolorectal carcinoma. AB - BACKGROUND/AIMS: The role of liver resection for hepatic metastases from noncolorectal carcinomas has yet to be clarified. The present study examines a single institutional experience of hepatic resection for noncolorectal metastases. METHODOLOGY: From January 1987 to March 1999, 14 patients underwent curative resection for liver metastases from noncolorectal carcinomas. Records of these patients were reviewed. RESULTS: Resections were performed for liver metastases from gastric cancers (n = 8), pancreatic cancers (n = 2), and cancers of bile duct, the papilla of Vater, kidney, and breast (n = 1, each). Six patients (5 with gastric cancers and 1 with pancreas cancer) presented with synchronous disease and 8 with metachronous disease. In the gastric cancer patients, there are 2 disease-free survivors (26 and 53 months) in the metachronous group, though all of the 5 patients with synchronous disease died within 29 months. All of the 4 patients with pancreatobiliary carcinomas died within 2 years. One case of breast cancer and another of renal cell cancer are alive without disease at 49 and 9 months, respectively. CONCLUSIONS: For metastases from gastric cancers, better survival after hepatic resection is expected in metachronous cases than in synchronous cases. Hepatic resection may afford little benefit for patients with liver metastases from pancretobiliary cancers. PMID- 11269009 TI - Electrochemical therapy--comparison with other local treatment methods on rat model. AB - BACKGROUND/AIMS: Electrochemical therapy is an alternative to treat hepatoma. We compare this method with the other local injection methods on rat liver. METHODOLOGY: Five groups of Wister rats (24 in each) were anaesthetized. Electrochemical therapy was set under direct current, 6 volts, electrodes were 0.5 cm apart, 0.5 cm deep into exposed parenchyma for 10 min. Local injection was done with 50 microL of 95% alcohol, 30 microL of 20% acetic acid, 30 microL of 35% hydrochloric acid, and 30 microL of 20% sodium hydroxide via a 27-gauge needle under direct vision into each rat. Rats and their livers were examined postmortem on day 1, 3, 7 and 14. RESULTS: In electrochemical therapy, the treated area showed coagulation necrosis and without blood cells on day 1; then the margin became blurred. Mononuclear cell infiltration, reperfusion and fibrous band formation were prominent from day 3 through day 14. In local injection groups, the necrosis is quite irregular and unpredictable. The regeneration went under similar process. CONCLUSIONS: To destroy tissue locally, electrochemical therapy is unique for its predictability in destructive area and the recovery process and is as effective as the other injection methods. Therefore, it has great potential for hepatoma treatment. PMID- 11269008 TI - Hepatic stellate cell activation occurs in nonalcoholic steatohepatitis. AB - BACKGROUND/AIMS: Hepatic stellate cell activation has a major role in the pathogenesis of hepatic fibrosis, considered to constitute part of the healing response to a necroinflammatory stimulus. However, steatosis per se, has also been shown to induce this activation. This study evaluates if hepatic stellate cell activation is present, and how it correlates with steatosis, in nonalcoholic steatohepatitis, whose hallmark is steatosis. METHODOLOGY: Steatosis, hepatocyte damage, inflammation and fibrosis were graded from 0 to 3+, in liver biopsies from 15 well documented nonalcoholic steatohepatitis and 5 normal controls. Activated hepatic stellate cell activation were identified immunohistochemically using a monoclonal antibody raised against cytoplasmic alpha-smooth muscle actin, and semiquantitatively graded using a scoring method. RESULTS: Nonalcoholic steatohepatitis patients showed significantly greater numbers of alpha-smooth muscle actin-reactive hepatic stellate cell than controls: hepatic stellate cell index of 3.6 +/- 1.9 versus 1.5 +/- 0.5, P < 0.05. The distribution of alpha smooth muscle actin-reactive hepatic stellate cell was higher in the perivenular areas, than in the intermediate zone and portal area, with no significant association between steatosis and alpha-smooth muscle actin-expressing hepatic stellate cell. However, a significant association was found between portal and lobular inflammation and hepatic stellate cell index, r = 0.72, P = 0.0005 and r = 0.75, P = 0.0002, respectively. CONCLUSIONS: This study demonstrates that hepatic stellate cell activation occurs in nonalcoholic steatohepatitis, clearly correlating with portal and lobular inflammation, but not with steatosis, suggesting that the mechanisms implicated in fibrosis in nonalcoholic steatohepatitis are probably related with inflammation. PMID- 11269010 TI - Long-term results following resectional surgery for Klatskin tumors. A twenty year personal experience. AB - BACKGROUND/AIMS: The purpose of this study was to assess whether long-term survival in patients suffering from cholangiocarcinomas of the porta hepatis is significantly different when comparing results between local and extended procedures in order to justify increased mortality and morbidity following extensive resections. METHODOLOGY: From November 1991 to May 2000, 46 patients with Klatskin tumor were assigned to two groups. Group A patients (n = 25) had local resection and group B patients (n = 21) had local resection plus hemihepatectomy. On admission, all patients were drained via percutaneous transhepatic biliary drainage. In all patients we proceeded with an internal biliary drainage in order to anticipate jaundice and decompensated liver function. Internal biliary drainage was carried out 35-40 days before surgery. At the end of the operation an arterial catheter was introduced into the common hepatic artery for adjuvant locoregional targeting immunochemotherapy, which was initiated 20 days following surgery in all patients. RESULTS: Overall survival for group A patients ranged from 14 months to 76 months (mean: 29). Disease-free survival ranged from 10-52 months (mean: 25). Five-year survival rate was 10%. Five-year disease-free survival was 0%. Overall survival for group B patients ranged from 28 months to 79 months (mean: 39). Disease-free survival ranged from 17-72 months (mean: 32). Five-year survival rate was 20%. Five-year disease-free survival rate was 10%. CONCLUSIONS: Combined tumor and liver resection is associated with significantly better results when compared with those following tumor resection alone. PMID- 11269011 TI - [Epileptic seizures from etomidate? The human Kv1.1 potassium channel in humans]. AB - OBJECTIVE: It is a matter of dispute whether etomidate exhibits an epileptogenic action. It is also disputed whether frequently observed myocloni induced by etomidate are related to seizure-like activity in the EEG. The clinical effects of anaesthetic agents reflect their molecular action. A possible epileptogenic action of etomidate should, therefore, result from action on a molecular target that has yet to be identified. Suppression of Kv1.1 channels may be associated with epilepsy in men. Inhibition of Kv1.1 channels by etomidate at clinically relevant concentrations would, thus, argue in favour of an epileptogenic action of etomidate. METHODS: Patch-clamp recordings of the pharmacological action of etomidate on cloned and functionally expressed human Kv1.1 channels. RESULTS: Etomidate inhibits human Kv1.1 channels in a concentration-dependent and reversible manner. The IC50-value is 400 microM, the Hill-coefficient is 2. The ratio of free clinical plasma concentrations and the IC50-value is 0.006. At plasma concentrations determined in a clinical setting Kv1.1 channels would be suppressed by less than 0.01%. CONCLUSION: The small effect of etomidate on human Kv1.1 channels at these concentrations questions an epileptogenic action of etomidate caused by inhibition of Kv1.1 channels. PMID- 11269012 TI - [Evaluation of a training program "Psychiatry for EmergencyPhysicians"]. AB - OBJECTIVE: Psychiatric education was neglected in emergency medicine until lately. Although measures for assuring the quality of care are established, there are up to now no investigations about the effectiveness of the psychiatric programs which are used in the German Emergency Physicians' Basic Training Program (EPTP-B, "Fachkundenachweis Rettungsdienst"). Herewith a first evaluation of a training program for psychiatric emergencies is presented. METHODS: Participants of the EPTP-B in Hamburg were imparted defined contents to achieve the educational objective. Before and afterwards two semi-structured questionnaires had to be answered asking for previous knowledge, attitudes, educational success and necessity for further training. RESULTS: At the beginning of the course 42 medical doctors (m/f: 22/20, mean age: 32.2 +/- 4.6 years) estimated their knowledge as poor and the necessity for training as high. At the end, the majority rated the program as educative, informative (69%) and practical (81%). However, the training was judged to be too short (81%) and the participants still felt too distant from the educational objective. CONCLUSIONS: There are necessity, demand and interest for an intensified psychiatric education of emergency physicians who are confronted with a high number and variety of psychic disturbances. On-the-job training of the theoretical matters is essential. PMID- 11269013 TI - [Osteochondroplastic tracheobronchopathy]. AB - The words tracheobronchopathia osteochondroplastica (TO) were coined by Aschoff in 1910, defining a rare disease of the tracheobronchial system which most often is only "accidentally" discovered in the course of examinations, a bronchoscopy or computerized tomography, which are undertaken for other reasons. A characteristic sign for this disease are multiple cobblestone-like protrusions in the trachea and bronchus which are caused by calcifications, chondrifications and ossifications of the upper layer of the mucous membrane. We report on a 81-year old patient who was diagnosed with the disease while undergoing a fiberoptic bronchoscopy during a prolonged weaning phase. In the beginning we discussed the possibility of a neoplasia or a chronic inflammatory process as differential diagnosis and based on that we considered a dilatative tracheotomy for the weaning period. After histological confirmation of a TO we refrained from the measure, the reason being that in our opinion the patient would not profit from it. To reach a correct diagnosis a good interdisciplinarian cooperation is essential. The patient was successfully extubated on the 11th post-operative day after a patient conventional weaning which was supported by accompanying symptomatic measures. Apart from the case report we present a summary of the literature on TO concerning epidemiology, pathology, diagnosis and therapeutical measures. PMID- 11269014 TI - [Morphine for pain treatment in a patient wih acute intermittent porphyria]. PMID- 11269015 TI - [Robotic surgery--surgical and anesthesiologic implications]. AB - Because of the restricted freedom of movement inherent in endoscopic standard instruments wholly endoscopic heart surgery has not been possible up to now. The development of such techniques was especially hindered by the rib cage. Now the precision which is imperative for bypass surgery or valve reconstruction is made possible by telemanipulator systems. Preconditions for this method are cardiopulmonary bypass-techniques which allow a cardioplegic cardiac arrest with closed chest and extensive hemodynamic monitoring which enables the anesthesiologist to make exact diagnoses without having a direct view on the heart. After an extensive experimental phase a telemanipulation system is successfully in use since May 1998 in the Cardiac Center in Leipzig. PMID- 11269016 TI - [Comments on "Severity in the prognosis following head-brain trauma." Krier C., Kienzle F. Anaesthesiol Intensivmed Notfallmed Schmerzther 2000;35:63-66 and "Outcome factors in severe head-brain trauma." Thomas A et al. Anasthesiol Intensivmed Notfallmed Schmerzther 2000;35:91-8]. PMID- 11269017 TI - [Bovine spongiform encephalopathy (BSE)]. PMID- 11269018 TI - [Transmissible spongiform encephalopathies--anesthesiologic and intensive care management]. AB - The transmissible spongiform encephalopathies (TSE) are known to affect humans and various animals. The bovine spongiform encephalopathy (BSE) and the human Creutzfeldt-Jacob disease (CJD) are among the most notable degenerative disorders caused by prions. Considering the BSE epidemic and the description of a new variant of Creutzfeldt-Jacob disease (nvCJD), which is probably related to bovine spongiform encephalopathy, TSE have recently gained a lot of public attention. Although the causative factors (prions, viruses) are still under discussion, none of the present concepts are explanatory for all aspects of the human CJD. CJD may present as a sporadic, genetic, or infectious illness and there is now considerable concern that bovine prions may have been passed to humans. To exclude transmission of CJD via medical products and instruments, the effectiveness of cleaning, disinfection and sterilization procedures must be firmly established. This manuscript presents an overview to anaesthesiology and intensive care medicine of recommended inactivation procedures and assessed these procedures in the light of the inactivation of prions. PMID- 11269019 TI - [Coupling of cardiac output or systemic O2 transport and metabolism during catecholamine therapy]. AB - Catecholamines increase cardiac output (CO) and thus systemic oxygen delivery, but they also increase the tissue's oxygen demand (thermogenic or calorigenic effect). Therefore, it is of particular interest for the choice of a catecholamine as to what extent CO is increased in relation to oxygen demand (VO2), because the tissue's oxygen balance is improved only if CO and thus oxygen delivery increases more than oxygen demand. For this purpose we reviewed the literature and analysed the relation between CO and VO2 during physiological as well as during pathological conditions. In particular, we compared the slopes of the regression lines of the individual CO/VO2-relation for each catecholamine. Dependent on study conditions, the extent of the increases in CO and VO2 varies substantially already for one and the same agent. During physiological conditions, the synthetic agents dobutamine and dopexamine primarily increase CO, whereas the endogenous ones epinephrine and particularly norepinephrine increase both CO and VO2 to about the same extent. During pathological conditions the literature is inconsistent, but it appears that the CO/VO2-relations do not differ substantially from those observed under physiological conditions. With due caution the current information implies that the synthetic catecholamines dobutamine and in particular dopexamine might be preferred in the treatment of low cardiac output states because they increase CO and thus oxygen delivery above the increase in metabolic demand. PMID- 11269020 TI - [Oral lichen planus. 2. Therapy possibilities and current treatment concepts]. PMID- 11269021 TI - [Possibilities of early diagnosis of familial adenomatous polyposis of the gastrointestinal system based on dentomaxillary findings]. PMID- 11269022 TI - [Role of forensic medicine and forensic psychiatry in solving the problem of organized crime]. AB - Term--organized crime is not satisfactorily defined todate neither in forensic sciences, nor in lexical formulations. It is necessary to come to grips with the criminalistic and social pathological meaning of three terms--individual crime, grouped and organized crime as well as participation of forensic sciences including forensic medicine on solving problems of organized crime. Forensic medicine and forensic psychology can help to solve this acute problem of present development of social life. It can help in criminalistic expertize and insider activity in profiliation of perpetrators and victims. Mainly it will be about searching and improving of methodical approaches in solving of incriminated cases and its analysis for investigative organs and courts. Important asset in this problem must be also preventive portion in preclusion of criminality. PMID- 11269023 TI - [Age determination using peptide mapping of non-collagenous proteins in human dentin]. AB - The submitted paper deals with possibilities of assessment of human age based on findings of stereospecific breakdown of proteins containing D-forms of aspartic acid. The stereospecificity of enzymatic breakdown assumes that after enzymatic hydrolysis peptide breakdown products with different molecular weights--the so called peptide map--will be formed, depending how many D-aspartyl residues the protein contains. The authors proved in the submitted preliminary study in subjects of different age the formation of breakdown products of different size in non-collagenous proteins of human dentin which was hydrolyzed by protease V8 from Staphylococcus aureus. PMID- 11269024 TI - [Quantification of cerebral contusions using the contusion index]. AB - The submitted results provide evidence that calculation of the contusion index (CI) for objective expression of the extent and depth of cerebral contusion is very suitable for various comparative studies. The authors assessed, using the CI the influence of direction, site of violent action, and age on the extent and depth of cerebral contusion in different types of head injuries. Consistent with the literature, the greatest contusion developed in areas of the frontal and temporal lobes. In case of violence from the back and from the side, in subjects above 50 years of age the CI of contusion par contrecoup was markedly greater than at the site of violence. The highest CI of the whole brain was in case of violence from the side. During falls from major heights or walking during alcohol intoxication the contusion was worse than in casualties during traffic accidents where pedestrians predominated. Fractures of the skull in junior subjects occurred only after an intensity which at the same time led also to the development of more extensive and deeper contusion as compared with older subjects where CI in fractures of the skull and intracranial haemorrhage were markedly smaller. PMID- 11269026 TI - [Hypothyroidism due to blocking type anti-TSH receptor antibodies]. PMID- 11269027 TI - [Painless thyroiditis]. PMID- 11269028 TI - [Autoimmune hypoparathyroidism]. PMID- 11269029 TI - [Autoimmune orchitis and male infertility]. PMID- 11269030 TI - [Autoimmune oophoritis]. PMID- 11269031 TI - [Idiopathic Addison disease, autoimmune adrenalitis]. PMID- 11269032 TI - [Insulitis of autoimmune diabetes]. PMID- 11269033 TI - [Insulin autoimmune syndrome]. PMID- 11269034 TI - [Type B syndrome of insulin resistance]. PMID- 11269035 TI - [Isaacs' syndrome]. PMID- 11269036 TI - [Polyglandular autoimmune syndrome]. PMID- 11269037 TI - [Evans syndrome]. PMID- 11269038 TI - [White cell aplasia]. PMID- 11269039 TI - [Pernicious anemia]. PMID- 11269040 TI - [Immune megakaryocytopenia]. PMID- 11269041 TI - [Thrombotic thrombocytopenic purpura]. PMID- 11269042 TI - [Autoimmune myelodysplasias]. PMID- 11269043 TI - [Autoimmune hemolytic anemia]. PMID- 11269044 TI - [Tolosa-Hunt syndrome]. PMID- 11269045 TI - [Autoimmune neutropenia]. PMID- 11269046 TI - [Autoimmune lymphocytopenia]. PMID- 11269047 TI - [Autoimmune pancytopenia, aplastic anemia]. PMID- 11269048 TI - [Autoimmune lymphoproliferative syndrome(ALPS)]. PMID- 11269049 TI - [Idiopathic thrombocytopenic purpura in childhood]. PMID- 11269050 TI - [Pure red cell aplasia]. PMID- 11269051 TI - [Idiopathic thrombocytopenic purpura (ITP)]. PMID- 11269052 TI - [Autoimmune basophilopenia]. PMID- 11269053 TI - [Chorea-acanthocytosis]. PMID- 11269054 TI - [Drug-induced autoimmune neutropenia(drug-induced AIN)]. PMID- 11269056 TI - [Lupoid thrombocytopenia]. PMID- 11269055 TI - [Drug-induced autoimmune hemolytic anemia]. PMID- 11269057 TI - [Myasthenia gravis]. PMID- 11269058 TI - [Autoimmune atrophic gastritis]. PMID- 11269059 TI - [Ulcerative colitis]. PMID- 11269060 TI - [Crohn's disease]. PMID- 11269061 TI - [Autoimmune hepatitis(AIH)]. PMID- 11269063 TI - [Autoimmune cholangitis]. PMID- 11269062 TI - [Primary biliary cirrhosis]. PMID- 11269064 TI - [Primary sclerosing cholangitis (PSC)]. PMID- 11269065 TI - [Autoimmune-related pancreatitis, autoimmune pancreatitis]. PMID- 11269066 TI - [Immunologic pleuritis]. PMID- 11269067 TI - [Idiopathic interstitial pneumonia, idiopathic pulmonary fibrosis]. PMID- 11269068 TI - [Juvenile myasthenia gravis]. PMID- 11269069 TI - [Primary autoimmune glomerular disease]. PMID- 11269070 TI - [Primary autoimmune tubulointerstitial nephritis]. PMID- 11269071 TI - [Pemphigus vulgaris]. PMID- 11269073 TI - [Herpetiform pemphigus]. PMID- 11269072 TI - [Pemphigus foliaceus]. PMID- 11269074 TI - [Paraneoplastic pemphigus (PNP)]. PMID- 11269075 TI - [IgA pemphigus]. PMID- 11269076 TI - [Concentric sclerosis (Balo)]. PMID- 11269077 TI - [Dermatitis herpetiformis Duhring]. PMID- 11269078 TI - [Bullous pemphigoid]. PMID- 11269079 TI - [Herpes gestationis]. PMID- 11269080 TI - [Epidermolysis bullosa acquisita]. PMID- 11269081 TI - [Cicatricial pemphigoid]. PMID- 11269082 TI - [Linear IgA bullous dermatosis]. PMID- 11269083 TI - [Erythema nodosum syndrome]. PMID- 11269084 TI - [Psoriasis vulgaris]. PMID- 11269085 TI - [Erythema exsudativum multiforme (EEM) syndrome, Stevens-Johnson syndrome (SJS)]. PMID- 11269086 TI - [Pustulosis palmaris et plantaris (PPP)]. PMID- 11269087 TI - [Vogt-Koyanagi-Harada disease]. PMID- 11269088 TI - [Sympathetic ophthalmia]. PMID- 11269089 TI - [Lens-induced uveitis]. PMID- 11269090 TI - [Fisher's syndrome]. PMID- 11269091 TI - [Goodpasture's syndrome]. PMID- 11269093 TI - [Churg-Strauss syndrome]. PMID- 11269092 TI - [Endomyocardial fibrosis]. PMID- 11269094 TI - [Henoch-Schonlein purpura]. PMID- 11269095 TI - [Wegener's granulomatosis (WG)]. PMID- 11269096 TI - [Microscopic polyangiitis]. PMID- 11269097 TI - [Classical polyarteritis nodosa]. PMID- 11269098 TI - [Temporal arteritis (giant cell arteritis)]. PMID- 11269099 TI - [Takayasu's aortitis, aortitis syndrome]. PMID- 11269100 TI - [Polyangiitis overlap syndrome]. PMID- 11269101 TI - [Guillain-Barre syndrome]. PMID- 11269102 TI - [Cutaneous vasculitis, cutaneous leukoclastic angiitis, hypersensitivity angiitis]. PMID- 11269103 TI - [Primary angiitis of the central nervous system]. PMID- 11269104 TI - [Thromboangiitis obliterans]. PMID- 11269105 TI - [Behcet's disease]. PMID- 11269106 TI - [Cogan's syndrome]. PMID- 11269107 TI - [Mouth and genital ulcers with inflamed cartilage]. PMID- 11269108 TI - [Shulman syndrome, diffuse eosinophilic fasciitis]. PMID- 11269109 TI - [Sjogren's syndrome]. PMID- 11269110 TI - [Weber-Christian disease]. PMID- 11269111 TI - [Malignant rheumatoid arthritis]. PMID- 11269112 TI - [Palindromic rheumatism]. PMID- 11269113 TI - [Scleroderma]. PMID- 11269114 TI - [Hypergammaglobulinemic purpura]. PMID- 11269115 TI - [Lewis-Sumner syndrome]. PMID- 11269116 TI - [Antiphospholipid syndrome]. PMID- 11269117 TI - [Mixed connective tissue disease]. PMID- 11269118 TI - [Relapsing polychondritis]. PMID- 11269119 TI - [Sarcoidosis]. PMID- 11269120 TI - [Jaccoud arthritis (chronic post rheumatic fever arthropathy)]. PMID- 11269121 TI - [Adult Still's disease]. PMID- 11269122 TI - [Fibromyalgia syndrome]. PMID- 11269124 TI - [Polymyositis and dermatomyositis]. PMID- 11269123 TI - [Systemic lupus erythematosus]. PMID- 11269125 TI - [Overlap syndrome]. PMID- 11269126 TI - [Essential cryoglobulinemia]. PMID- 11269127 TI - [Cryofibrinogenemia]. PMID- 11269128 TI - [Rheumatoid arthritis]. PMID- 11269129 TI - [Acute disseminated encephalomyelitis (ADEM)]. PMID- 11269130 TI - [Undifferentiated connective tissue syndrome]. PMID- 11269131 TI - [Arthritis mutilans]. PMID- 11269132 TI - [Polymyalgia rheumatica]. PMID- 11269133 TI - [Lymphomatoid granulomatosis]. PMID- 11269134 TI - [Rheumatic fever]. PMID- 11269135 TI - [Juvenile rheumatoid arthritis]. PMID- 11269136 TI - [Systemic lupus erythematosus in childhood]. PMID- 11269137 TI - [Scleroderma]. PMID- 11269138 TI - [Multiple sclerosis]. PMID- 11269139 TI - [Mixed connective tissue disease in childhood]. PMID- 11269140 TI - [Aortitis syndrome (Takayasu's arteritis) in childhood]. PMID- 11269141 TI - [Juvenile dermatomyositis, juvenile polymyositis]. PMID- 11269142 TI - [Behcet disease]. PMID- 11269143 TI - [Sjogren's syndrome]. PMID- 11269144 TI - [Drug-induced lupus erythematosus]. PMID- 11269145 TI - [Scleroderma, scleroderma related diseases]. PMID- 11269147 TI - [Other autoimmune diseases]. PMID- 11269146 TI - [Primary autoimmune hemolytic anemia (warm antibody)]. PMID- 11269148 TI - [Neonatal transient myasthenia gravis]. PMID- 11269151 TI - [Neonatal alloimmune thrombocytopenia]. PMID- 11269149 TI - [Neonatal hyperthyroidism]. PMID- 11269150 TI - [Neonatal hypothyroidism]. PMID- 11269152 TI - [Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP)]. PMID- 11269153 TI - [Neonatal hypoglycemia]. PMID- 11269155 TI - [Multiple chemical sensitivity]. PMID- 11269154 TI - [Neonatal lupus erythematosus]. PMID- 11269157 TI - [Allergic conjunctivitis]. PMID- 11269156 TI - [Allergic rhinitis]. PMID- 11269158 TI - [Urticaria and angioedema]. PMID- 11269159 TI - [Atopic dermatitis]. PMID- 11269160 TI - [Allergic contact dermatitis]. PMID- 11269161 TI - [Bronchial asthma]. PMID- 11269162 TI - [Hypereosinophilic syndrome]. PMID- 11269163 TI - [Lambert-Eaton myasthenic syndrome]. PMID- 11269164 TI - [Allergic broncho pulmonary aspergillosis (ABPA)]. PMID- 11269166 TI - [Food allergy]. PMID- 11269165 TI - [Hypersensitivity pneumonitis]. PMID- 11269167 TI - [Serum sickness]. PMID- 11269168 TI - [Heiner's syndrome]. PMID- 11269169 TI - [Hopkins syndrome (acute postasthmatic amyotrophy)]. PMID- 11269170 TI - [Aspirin-induced asthma]. PMID- 11269171 TI - [Allergic tension-fatigue syndrome (ATFS)]. PMID- 11269172 TI - [Stiff-man syndrome]. PMID- 11269174 TI - [Exercise induced asthma (EIA)]. PMID- 11269173 TI - [Exercise-induced anaphylaxis]. PMID- 11269175 TI - [Insect allergy]. PMID- 11269176 TI - [Cough variant asthma]. PMID- 11269177 TI - [Puberal asthma]. PMID- 11269178 TI - [Sick house syndrome, sick building syndrome, indoor harmful substance sensitivity]. PMID- 11269179 TI - [Metallic salts asthma]. PMID- 11269180 TI - [Latex allergy]. PMID- 11269181 TI - [Baker's asthma]. PMID- 11269182 TI - [Paraneoplastic opsoclonus myoclonus syndrome (POMS)]. PMID- 11269184 TI - [Hoya (sea-squirt) asthma]. PMID- 11269183 TI - [Red soft coral induced allergic symptoms]. PMID- 11269185 TI - [Mushroom allergy]. PMID- 11269186 TI - [Allergy due to air pollutants (SPM, SO2, NO2, etc)]. PMID- 11269187 TI - [Allergy due to occupational vanadium exposure]. PMID- 11269188 TI - [Allergy of external medicine]. PMID- 11269189 TI - [Drug allergy due to systemic use]. PMID- 11269190 TI - [Paraneoplastic cerebellar degeneration]. PMID- 11269191 TI - [Paraneoplastic encephalomyelitis/sensory neuropathy]. PMID- 11269192 TI - [Carcinoma-associated retinopathy]. PMID- 11269193 TI - [Chronic hypertrophic cranial pachymeningitis]. PMID- 11269194 TI - [Motor neuropathy associated with autoantibodies]. PMID- 11269195 TI - [Motor neuron disease with autoantibodies]. PMID- 11269196 TI - [Autoimmune hypophysitis (lymphocytic hypophysitis)]. PMID- 11269197 TI - [HTLV-I--associated myelopathy/tropical spastic paraparesis (HAM/TSP)]. PMID- 11269198 TI - [Graves' disease]. PMID- 11269199 TI - [Primary hypothyroidism]. PMID- 11269200 TI - [Hashimoto disease]. PMID- 11269201 TI - Conflict of interest: organizational vs. executive ethics in health care. AB - This survey of 207 senior- and mid-level administrators from across the U.S. details the surprising level and type of ethical conflicts and unethical practices they face on the job and how their organizations create and respond to these challenges. The findings suggest widespread ethical conflict caused by situational and environmental factors, financial incentives, boards of directors, and stakeholder pressure. Particularly poignant is the apparent inability of executives to alter the ethical environment. Substantial differences were found between nonprofit and proprietary sector administrators as well as gender and age groups. PMID- 11269202 TI - Gaining strategic advantages through partnerships with public health departments. AB - Public health officials have advocated in public health and public policy journals for collaboration with private sector health care organizations for nearly a decade. There has been little written in the management literature on this topic, however. There are several important areas in which public health departments have expertise that could be valuable to private sector health care organizations, including health maintenance organizations (HMOs). These include the delivery of services in some geographic areas and to some special populations, provision of preventive and health promotion services to HMO members, performance of epidemiology services, assistance in accreditation, and repair of the damaged image of HMOs. HMOs and local health departments in many parts of the country are already entering into contracts for these purposes. Such partnerships between HMOs and local health departments can improve the health of the members of HMO plans and contribute to improving the health of the community. PMID- 11269203 TI - Hospital competition in major U.S. metropolitan areas: empirical evidence. AB - In response to dramatic rises in health care costs, policy-makers have been debating the relative merits of competitive strategies as a means of containing costs. This article represents a study of the 29 largest MSAs for 1991. Controlling for environmental conditions in each market, the impact of competition on hospital costs was examined. Competition was found to have had a significant positive impact on overall hospital costs. PMID- 11269204 TI - Physician unionization in the United States: fad or phenomenon? AB - This article explores the current trends and issues surrounding physician unionization in the United States, using data from secondary sources and nine interviews with leaders of organizations at the forefront of physician unionizing efforts. Several key points are supported by these data and prior unionization research. First, unions should become a viable organizing alternative for the almost 50% of doctors who are salaried employees because of fewer legal barriers to collective representation, the involvement of national labor unions with resources to spend on organizing, more physicians belonging to demographic groups less hostile to organized labor. and work-related pressures faced by physician employee under managed care. A second key point is that unions will find it difficult to represent self-employed physicians because of the influence of organized medicine and legal barriers to gaining collective bargaining rights for this group. This discussion is intended to raise awareness of the physician union issue among health care policy-makers and researchers. PMID- 11269205 TI - New patterns of community care: coordinated services for dually diagnosed adults in North Carolina. AB - North Carolina has developed coordinated care systems for people dually diagnosed with both a mental retardation and a different major mental illness. In response to a class action lawsuit, the state has become a leader in treatment of this form of dual diagnosis. Systems of care for this "Thomas S class" operate in each of 41 area programs for mental health, developmental disabilities, and substance abuse. Networks of care among leaders in mental health and developmental disability promote the coordination of health, housing, social, and vocational services. A survey of 100 area program leaders finds extensive cooperation and a variety of services provided and contracted for, both within and beyond each area program, particularly among developmental disability specialists. Cooperation among leaders is associated with service variety and inter-organizational linkages. The extent of relationships among provider organizations is associated with better access to care. Best practice includes a single portal of entry and inter-agency councils. PMID- 11269206 TI - Medical students' and doctors' perceptions about Medicaid: a content analysis. AB - This research investigates the similarities and differences in medical students' and doctors' perceptions about Medicaid. One hundred, ninety-seven medical students and doctors were asked to answer 4 three open-ended questions about Medicaid. Their responses were content analyzed using standard rules of manifest and content analysis. Results indicate important differences between the groups. Implications are provided. PMID- 11269207 TI - International health and emerging infectious diseases. AB - This article explored the role of international health in reducing the impact of infectious diseases by espousing the monumental application of global electronic communication and socioeconomic development initiatives. The interaction between the society and environmental changes have dramatic effects on the frequency of infectious diseases worldwide. Development of dams, human population expansion, migration patterns, urbanization and the invasion of hitherto virgin forests, and global warming enhance the proliferation of vectors of infectious diseases. Sustainable development and emphasis on primary prevention initiatives, coupled with the application of technology to improve farming, provision of safe water and electrification of rural communities, are significant steps in infectious disease control. PMID- 11269208 TI - Vestibular testing and the big E. PMID- 11269209 TI - Bilateral laryngoceles in a young trumpet player: case report. PMID- 11269210 TI - Large nonobstructing exostoses of the external auditory canal. PMID- 11269211 TI - Unusual cartilaginous lesion of the torus tubarius as a cause of otalgia. PMID- 11269212 TI - Laryngeal candidiasis. PMID- 11269213 TI - 'Don't touch me' lesions of the petrous apex. PMID- 11269214 TI - Severe, disabling dizziness after intratympanic aminoglycoside treatment for dizziness. PMID- 11269215 TI - The microbiology of chronic rhinosinusitis: results of a community surveillance study. AB - In view of the rapidly changing patterns of antibiotic resistance, community surveillance studies are providing important information to help guide practitioners in making their choice of antibiotics. For this community surveillance study, we performed a retrospective chart review of nasal and sinus culture data obtained from 83 patients with typical symptoms of chronic rhinosinusitis who visited a community otolaryngologist in Rochester, New York. Pathogens were isolated in 59 of these patients (71%). The most common were coagulase-negative staphylococci (31% of isolates). Among the other isolated pathogens were Hemophilus influenzae (25%), Streptococcus pneumoniae (12%), Moraxella catarrhalis (10%), Pseudomonas aeruginosa (7%), alpha-hemolytic streptococci (5%), and Staphylococcus aureus (3%). Approximately 39% of the coagulase-negative staphylococci isolates were resistant to penicillin. Some 20% of the H influenzae isolates were beta-lactamase-positive, and 14% of all isolates were resistant to multiple antibiotics. Approximately 12% of the 83 patients cultured positive for multiple organisms. Our findings provide important surveillance information about the resistance patterns of pathogens in our area. Although the prevalence of beta-lactamase-positive H influenzae that we observed was consistent with those of other reports, we found a lower prevalence of polymicrobial flora. Our findings suggest that culture- and sensitivity-directed therapy should be effective in the treatment of chronic rhinosinusitis. PMID- 11269216 TI - Chondrolipoma of the oropharynx. AB - We describe the startling case of a man who was able to extend and retract a smooth round mass in and out of his oropharynx at will. On examination under anesthesia, the mass was found to be attached to the posterior tonsillar pillar by a stalk. The lesion was excised, and histopathology determined that it was a chondrolipoma. We believe that this is the first report of a chondrolipoma at this anatomic site. PMID- 11269217 TI - Lateralized carotid artery: an unusual cause of pulsatile tinnitus. AB - We describe the case of a patient who had a pulsatile tinnitus that was caused by a laterally displaced internal carotid artery. Her condition was treated with the use of a hearing did, which suppressed the tinnitus. We also review the literature on laterally displaced internal carotid arteries, and discuss their differentiation from a congenitally aberrant artery. PMID- 11269218 TI - Myoepithelioma of the parotid gland: a report of two cases. AB - We describe the clinical and pathologic features of two benign myoepitheliomas of the parotid gland. Through 1985, only 42 other cases had been reported in the literature--39 benign and three malignant. Fewer than 100 cases had been reported through 1993. Since then, two other reports have been published. But are these tumors really rare, or are they simply not well recognized? It is our opinion that they are not as rare as is generally believed because the number of case reports is increasing as pathologists have become more aware of their existence. PMID- 11269220 TI - Lymphoma: an update on evolving trends in staging and management. AB - Because lymphoma frequently manifests as a neck mass, otolaryngologists are often the first to evaluate and diagnose it. Excisional biopsy of lymph nodes generally provides the definitive diagnosis. After diagnosis, however, the otolaryngologist's involvement generally wanes as subsequent treatment of the patient is provided by an oncology team. Nevertheless, it is important for the otolaryngologist to be familiar with current concepts in the comprehensive evaluation and treatment of lymphoma patients. This knowledge allows us to participate in and facilitate timely testing, which helps avoid undue delays between the documentation of physical findings and the initiation of treatment. Otolaryngologists also need to be up to date on recent developments in the treatment of lymphomas in order to be able to answer questions regarding staging and prognosis, to make the proper referrals, and to help our patients understand current controversies and treatment practices. PMID- 11269219 TI - Frontal recess surgery for diving-related frontal pain: case report. AB - We report the case of a professional scuba diver who was unable to dive because he began experiencing severe frontal pain on descent. Following endoscopic surgery to open the frontal recess, the man was able to resume diving unrestricted by pain. We discuss the causes and treatment of this complaint, and we suggest that this might be considered a new indication for surgery in a limited number of cases. PMID- 11269221 TI - Securing cochlear implants to the skull: two alternate methods. AB - In view of the various problems encountered with the traditional methods of securing cochlear implants--including dural tear and suture dissolution following infection--we devised two alternate methods of performing this procedure. We use a titanium mesh or a Gore-Tex patch secured with two 4-mm screws to fix the receiver to the skull. No patient who has undergone either of these procedures at our institution has experienced any of the complications that are associated with the older silk, nylon, and Dacron sutures. Moreover, our two alternate methods are less technically difficult and can be performed in a shorter period of time. PMID- 11269222 TI - The autoimmune lymphoproliferative syndrome. A disorder of human lymphocyte apoptosis. PMID- 11269223 TI - Immune deficiency presenting as mycobacterial infection. AB - All the discrete genetic defects identified to date that seem to specifically predispose to infection with NTM or BCG have occurred in the pathways involving the generation of or response to IFN-gamma (Fig. 3). This natural genetic survey therefore suggests that one of the most critical cytokines in the control of NTM and BCG is IFN-gamma. Unfortunately, this recognition does not give us a clear sense of the critical mechanism(s) involved and still leaves us at a phenomenological level of understanding. Therefore, even though IFN-gamma appears to be the most critical cytokine in the control of mycobacteria by the "experiments of nature" cited earlier, it is likely that other cytokines are involved in the more proximal events of killing of intracellular parasites and viral control. These cytokines or chemokines may perform better therapeutically than [figure: see text] does IFN-gamma if they are better able to evoke the critical antimycobacterial mechanism(s). Dissection of these pathways, identification of the most proximal factors, and exploitation of these findings for the treatment of mycobacterial and other intracellular infections is the critical charge for the future. PMID- 11269224 TI - The hyper-IgE syndrome. PMID- 11269225 TI - The Lambert-Eaton myasthenic syndrome. PMID- 11269226 TI - Signal-transduction defects in T cells. AB - In conclusion, multiple receptors and signal transduction cascades influence T cell function and fate. During the past few years many of these important aspects of T-cell biology were identified. The complexity of the various signaling pathways has made appreciation of their clinical significance difficult. One way of studying the function of these molecules is to create mice deficient of these components. However, frequently the murine phenotype is far from reflecting the homologous human deficiency. It is therefore beneficial to define the human immunodeficiencies in order to understand the role of a certain signaling molecule in humans. Further, mutations that result in partial deficiencies may result in a different phenotype from null mutations. This information may aid in improving structure/function analysis of these signaling components. PMID- 11269227 TI - Primary T-lymphocyte immunodeficiencies. PMID- 11269228 TI - The DiGeorge anomaly. AB - The DiGeorge anomaly, originally considered a clinical paradigm for isolated thymus deficiency, has now been redefined as a member of a group of disorders that share in common a chromosome deletion resulting in monosomy 22q11 (CATCH-22 or DGA/VCFS). In addition to the thymus defect, conotruncal heart anomalies, dysmorphism, hypoparathyroidism, and cleft palate are prominent features. Despite the emphasis on thymus involvement in DGA, a clinically significant thymus defect is found only in a small percentage of these patients probably occurring in less than 5% of the cases. Maldescent of the thymus, however, is extremely common, leading to an absence of mediastinal thymic tissue in nearly all cases. The basic embryological fault in these disorders is an inadequate development of the facial neural-crest tissues, resulting in defective organogenesis of pharyngeal pouch derivatives that receive cephalic neural-crest contribution to the mesenchmyme. The causes for this maldevelopment are both genetic and extragenetic in origin and the genetic lesions act in concert with random environmental events to produce the ultimate clinical picture. The modern research approaches now available have cleared away most of the confusion clouding the clinical and theoretical aspects of DGA and related disorders, providing the clinician with useful landmarks to assess and treat these intriguing clinical challenges. PMID- 11269231 TI - Model.it: building three dimensional DNA models from sequence data. AB - A WWW server is described for creating 3D models of canonical or bent DNA starting from sequence data. Predicted DNA trajectory is first computed based on a choice of di- and tri-nucleotide models (M.G. Munteanu et al., Trends Biochem. Sci. 23, 341-347, 1998); an atomic model is then constructed and optionally energy-minimized with constrained molecular dynamics. The data are presented as a standard PDB file, directly viewable on the user's PC using any molecule manipulation program. AVAILABILITY: The model.it server is freely available at http://www.icgeb.trieste.it/dna/ CONTACT: kristian@icgeb.trieste.it; pongor@icgeb.trieste.it SUPPLEMENTARY INFORMATION: a series of help files is available at the above address. PMID- 11269232 TI - Characterization of the mupA gene in strains of methicillin-resistant Staphylococcus aureus with a low level of resistance to mupirocin. PMID- 11269229 TI - The Wiskott-Aldrich syndrome. PMID- 11269230 TI - The pathogenesis of ataxia-telangiectasia. Learning from a Rosetta Stone. PMID- 11269233 TI - In vitro activity of rifaximin against enteropathogens producing traveler's diarrhea. PMID- 11269234 TI - Identification of vat(E-3), a novel gene encoding resistance to quinupristin dalfopristin in a strain of Enterococcus faecium from a hospital patient in the United Kingdom. PMID- 11269235 TI - [Which allergy tests do you order for chronic urticaria?]. PMID- 11269236 TI - [Anal fissures: treatment with botulinum toxin]. PMID- 11269237 TI - Two classes of tRNA synthetases suggested by sterically compatible dockings on tRNA acceptor stem. PMID- 11269238 TI - Sixth international workshop on scleroderma research, Oxford, UK, 30 July--22 August 2000. PMID- 11269239 TI - Role of semen in the female-to-male transmission of HIV. PMID- 11269241 TI - Ask the doctor. I'm 78 years old. I've known for years that I have a narrowing of the aortic valve of my heart. My doctors are always asking me whether I have chest pain, fainting spells, or any other special symptoms. Until recently, the answer has been no. A few days ago, however, I was washing dishes in my kitchen, and I suddenly felt lightheaded and fell to the ground. I didn't black out, but I almost did. Now my doctor is saying that I should have my aortic valve replaced. Isn't this rather an extreme response to just one spell of lightheadedness? PMID- 11269242 TI - [Mite infestations (scabies). Diagnosis, treatment and prevention]. PMID- 11269243 TI - The correlation between presenting ST-segment depression and the final size of acute myocardial infarcts in patients with acute coronary syndromes. AB - The use of reperfusion therapy in patients with ST elevation acute coronary syndromes had been established. However, reperfusion therapy is usually considered contra-indicated in those with ST depression, despite the knowledge that regional posterior infarction is typically indicated by ST depression maximal in leads V1 to V3 and nonregional subendocardial infarction is typically indicated by marked ST depression maximal in other leads. This study of patients with non-ST-elevation acute coronary syndromes investigates the quantitative relationship between presenting ST depression and final QRS changes in both of these subgroups. The final QRS score was significantly higher (2.44 points) than that of a control group with not ST depression, (1.55 points) in the group with maximal ST depression in V1 to V3 (P = 0.04). However, in the entire population, there was a highly significant correlation (P = .003) between the sum of the presenting ST depression and the final QRS score. Trials of reperfusion therapy will be required to determine if such evolution to electrocardiogram documented acute myocardial infarction can be prevented in patient with marked ST depression acute coronary syndromes. PMID- 11269244 TI - A 12-year-old boy with a limp. PMID- 11269246 TI - Artificial intelligence applications in the intensive care unit. AB - OBJECTIVE: To review the history and current applications of artificial intelligence in the intensive care unit. DATA SOURCES: The MEDLINE database, bibliographies of selected articles, and current texts on the subject. STUDY SELECTION: The studies that were selected for review used artificial intelligence tools for a variety of intensive care applications, including direct patient care and retrospective database analysis. DATA EXTRACTION: All literature relevant to the topic was reviewed. DATA SYNTHESIS: Although some of the earliest artificial intelligence (AI) applications were medically oriented, AI has not been widely accepted in medicine. Despite this, patient demographic, clinical, and billing data are increasingly available in an electronic format and therefore susceptible to analysis by intelligent software. Individual AI tools are specifically suited to different tasks, such as waveform analysis or device control. CONCLUSIONS: The intensive care environment is particularly suited to the implementation of AI tools because of the wealth of available data and the inherent opportunities for increased efficiency in inpatient care. A variety of new AI tools have become available in recent years that can function as intelligent assistants to clinicians, constantly monitoring electronic data streams for important trends, or adjusting the settings of bedside devices. The integration of these tools into the intensive care unit can be expected to reduce costs and improve patient outcomes. PMID- 11269245 TI - Two-stage response to endotoxin infusion into normal human subjects: Correlation of blood phagocyte luminescence with clinical and laboratory markers of the inflammatory, hemostatic response. PMID- 11269247 TI - Commentary: Early discontinuation violates Helsinki principles. PMID- 11269248 TI - Rectal bleeding and colorectal cancer. Inclusion criteria of study need clarification. PMID- 11269249 TI - Rectal bleeding and colorectal cancer. Results of study were incorrectly interpreted. PMID- 11269250 TI - All epidemiological evidence is important in colorectal cancer. PMID- 11269252 TI - Impact of NHS Direct on demand for immediate care. Meaningful review is still outstanding. PMID- 11269251 TI - Impact of NHS Direct on demand for immediate care. NHS Direct must be better marketed and deal with problems more effectively. PMID- 11269253 TI - Impact of NHS Direct on demand for immediate care. NHS Direct can help accident and emergency departments. PMID- 11269254 TI - Should NICE's advice be handled centrally or locally? PMID- 11269255 TI - Catheter ablation for cardiac arrhythmias. Ablation should not be denied to elderly patients on basis of age. PMID- 11269256 TI - Surveillance of Haemophilus influenzae infection. Surveillance data for assessing impact of vaccination are valid. PMID- 11269257 TI - Outcome of pregnancy in diabetic women. Authors did not define criterion for case selection. PMID- 11269258 TI - Outcome of pregnancy in diabetic women. More investigation is needed into whether control of diabetes is really poorer in England than Norway. PMID- 11269260 TI - Ultrasonography in diagnosis of acute appendicitis. Active observation is often sufficient to make diagnosis. PMID- 11269259 TI - Ultrasonography in diagnosis of acute appendicitis. Diagnostic laparoscopy is often more useful than ultrasonography. PMID- 11269261 TI - Interrupting antiretroviral treatment needs particular care. PMID- 11269262 TI - Smoking and use of mobile phones. Data have been wrongly interpreted. PMID- 11269263 TI - Smoking and use of mobile phones. Italian data don't show the same pattern. PMID- 11269264 TI - Smoking and use of mobile phones. No correlation in Switzerland either. PMID- 11269265 TI - Hospital mergers may be painful but have positive aspects too. PMID- 11269266 TI - The fragile male. Male zygotes are often formed at suboptimal times in fertile cycle. PMID- 11269267 TI - The fragile male. Men should follow example of women's health movement. PMID- 11269268 TI - Title of Dr should be sufficient for all doctors. PMID- 11269269 TI - Mesh compared with non-mesh methods of open groin hernia repair: systematic review of randomized controlled trials and laparoscopic compared with open methods of groin hernia repair: systematic review of randomized controlled trials. PMID- 11269270 TI - Mesh compared with non-mesh methods of open groin hernia repair: systematic review of randomized controlled trials and laparoscopic compared with open methods of groin hernia repair: systematic review of randomized controlled trials. PMID- 11269271 TI - Functional outcome after restorative panproctocolectomy for ulcerative colitis decreases an otherwise enhance quality of life. PMID- 11269272 TI - Vaccination with irradiated cercariae of Schistosoma mansoni preferentially induced the accumulation of interferon-gamma producing T cells in the skin and skin draining lymph nodes of mice. AB - Cytokine response to schistosomula of Schistosoma mansoni was evaluated in the skin of mice during the initial 72 h following infection. These studies showed a significant increase in the levels of IL-4 and IL-10 message in the skin in areas of cercarial penetration. The IL-4 message was detectable in the skin as early as 8 h after infection and the message for IL-10 appeared from 16 h after infection. However, mRNA for IFN-gamma was undetectable in the skin samples for up to 72 h after infection with normal cercariae. In sharp contrast, vaccination with irradiated cercariae induced IFN-gamma and IL-2 responses in the skin within 24 h. Analysis of the cytokine profile of cells isolated from the skin during these early time points showed that T cells are probably not a source of IL-4 or IL-10 in the skin of mice infected with normal cercariae. However, in vaccinated animals, the majority of the IFN-gamma is derived from skin-residing T cells. In vaccinated animals, responses in the skin were mirrored in the skin-draining lymph nodes as well. Analysis of the CD4/CD8 ratio showed a significant decrease in the skin following vaccination suggesting an increase in CD8+ cells. Interestingly however, when vaccinated animals were challenged with normal cercariae, there was a significant reduction in IFN-gamma response in the skin and its draining lymph nodes. These results show that vaccination with irradiated cercariae of S. mansoni, preferentially induce the accumulation of IFN-gamma producing T cells in the skin and skin-draining lymph nodes of mice. PMID- 11269273 TI - Coproantigen survey for Echinococcus multilocularis prevalence of red foxes in Hokkaido, Japan. AB - An epidemiological survey was conducted on the seasonal variation of Echinococcus multilocularis prevalence in red foxes from 1997 to 1998, using a monoclonal antibody-based detection of the tapeworm coproantigen. Thirty-six breeding dens of reproductive fox families were identified in the endemic area of Koshimizu, eastern Hokkaido, Japan. Fecal samples from each site were examined by coproantigen detection assay and fecal egg examination. Whereas the prevalence of coproantigen positive feces showed no seasonal fluctuation (51.6-66.7%), variation was found in the prevalence of egg positive feces in which a higher prevalence was observed in the summer and winter (31.1 and 38.7%) than spring and autumn (13.3 and 13.5%). Significant differences were observed between juveniles and adult foxes in both examinations. Samples from juvenile foxes gave higher coproantigen positive results and taeniid egg intensity. Those results suggest more juveniles infected with the cestode than adults in the same period. The practical use of coproantigen assay as a survey tool and factors which affect the prevalence and host age-related difference are discussed. PMID- 11269274 TI - Sequence analysis of genes encoding ribosomal proteins of amitochondriate protists: L1 of Trichomonas vaginalis and L29 of Giardia lamblia. AB - Two genes encoding the ribosomal proteins were cloned and sequenced from amitochondriate protists, L1 (L10a in mammalian nomenclature) from Trichomonas vaginalis and L29 (L35 in mammalian nomenclature) from Giardia lamblia. The deduced amino acid sequences were analyzed by sequence alignments and phylogenetic reconstructions. Both the T. vaginalis L1 and the G. lamblia L29 displayed eukaryotic sequence features, when compared with all the homologs from the three primary kingdoms. PMID- 11269275 TI - Improved detection of Dirofilaria repens DNA by direct polymerase chain reaction. AB - Diagnosis of human infection by Dirofilaria repens, depends mainly on microscopic evaluation of tissue cross-sections and the macroscopic characteristics of the worm. Tissue degeneration and/or poor specimen preparation practices however, often render many cases of subcutaneous dirofilariasis elusive to such morphological diagnostic approaches. The early PCR protocols, developed to satisfy these complex diagnostic needs, failed to amplify dirofilariae DNA from formalin preserved material. To overcome these difficulties, we developed an improved PCR protocol using a set of primers designed to amplify a rather stable, highly repetitive D. repens-specific genomic DNA target. We report the performance of this protocol with a large variety of dirofilariae infected DNA specimens, including those extracted from formalin preserved biological material for up to 20 days. Our findings support its potential application to routine clinical diagnosis. PMID- 11269276 TI - A case of Diphyllobothrium nihonkaiense infection successfully treated by oral administration of Gastrografin. AB - A diphyllobothriid cestode infection found in a 54-year-old male residing in Oita, Japan, was successfully treated by oral administration of Gastrografin in combination with a intramuscular injection of Vagostigmin. The strobila expelled was 6.14 m long with a scolex, and morphologically identical with Diphyllobothrium nihonkaiense except unusual ovaries of which posterior horns were confluent in each proglottid. This is the first case of treatment of cestode infection by oral administration of Gastrografin. PMID- 11269277 TI - Suppressed expression of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) in an irradiation-attenuated Plasmodium berghei XAT strain. AB - Plasmodium berghei XAT (XAT) is a non-reversible, non-lethal type malaria parasite strain derived from the highly virulent lethal P. berghei NK65 (NK65) by X-irradiation. The difference in polypeptide expression between NK65 and XAT was examined in this study. Western blot patterns of the parasite polypeptides showed that a 30-kDa polypeptide was not detected in XAT. In the present paper, we focused the study on the difference in the expression of the 30-kDa polypeptide between XAT and NK65. Although several other significant differences were noted in the spots shown by two-dimensional gel electrophoresis, the 30-kDa polypeptide was isolated by means of preparative 2D-gel electrophoresis followed by HPLC, and N-terminal amino acid sequence of the polypeptide was eventually determined. Complementary DNA clones encoding the 30-kDa polypeptide were isolated and characterized. Full-length cDNA clones from XAT encoded a protein of 231 amino acid residues with a 693-bp open reading frame. The deduced amino acid sequence exhibited 67% identity with that for P. falciparum hypoxanthine-guanine phosphoribosyltransferase (HGPRT; EC 2.4.2.8), suggesting that this protein is P. berghei HGPRT. Northern blot analysis revealed that expression of HGPRT in XAT was only one-eighth of that in NK65. This finding indicates that HGPRT gene expression is markedly suppressed in XAT. The amino acid sequence of HGPRT from NK65 was identical to that from XAT. This finding showed that the amino acid sequence of XAT-HGPRT was not mutated and had not undergone deletion. PMID- 11269279 TI - A new avian cestode Malika turdi n. spp. Dilepididae: Dipylidiinae) from the golden mountain thrush, Turdus dauma aureus Holandre in Japan. AB - Malika turdi n. spp. (Dilepididae: Dipyliidinae) was described from the small intestine of a golden mountain thrush, Turdus dauma aureus Holandre, at Ogori City, Fukuoka Prefecture, Japan. By having two rows of rostellar hooks, it resembles M. daviesi, M. himantopodis, M. kalawewaensis, and M. zeylanica, but is readily distinguished from the first three species by having non-spiny cirrus, and from the latter species by having a smaller scolex and more testes. This is the first report of Malika in Japan. PMID- 11269278 TI - Production of interleukin-10 by peripheral blood mononuclear cells from residents of a marshland area in China endemic for Schistosoma japonicum. AB - Interleukin-10 (IL-10) cytokine production was assessed using peripheral blood mononuclear cells (PBMC) from 67 individuals living in an area endemic for schistosomiasis japonica in China (Dongting Lake, Hunan Province), and 11 control subjects from a non-endemic part of the same Province. Production of IL-10 was measured following in vitro stimulation of PBMC using whole parasite extract (SWAP) or a panel of recombinant Schistosoma japonicum antigens (22-kDa tegumental membrane-associated antigen, glyceraldehyde-3-phosphate dehydrogenase, paramyosin, 14-kDa fatty acid-binding protein and 28-kDa glutathione S transferase) which are of recognized interest in the development of protective immunity to schistosomiasis. Significantly, PBMC isolated from the exposed population compared with the non-exposed population produced higher levels of IL 10. There was a trend towards higher mean levels of IL-10 release in putatively resistant (insusceptible) (consistently egg negative but highly exposed) individuals compared with susceptible (egg-positive) subjects from the exposed population. Analysis of individual exposure (the duration of water contact and the percent body surface area in contact with water, expressed as m2 h/day) vs. IL-10 production indicated a weak but consistent and statistically significant inverse correlation, with lower levels of exposure being associated with higher levels of IL-10. These results suggest an association between IL-10 production and resistance to S. japonicum in subjects from this Chinese population exposed to infection. PMID- 11269280 TI - Primary infections with Babesia microti are not prolonged by concurrent Heligmosomoides polygyrus. AB - Seven experiments were carried out to test the hypothesis that concurrent infection with the chronic and immunomodulatory intestinal nematode parasites, Heligmosomoides polygyrus, and the piroplasm, Babesia microti, would result in more intense and long-lasting infections with the hemoprotozoan. However, despite variations in the experimental protocols (different mouse strains, varying levels of infection and different intervals between infection with the two species) a significantly higher B. microti parasitaemia was detected on only one occasion, and a significantly lower parasitaemia on two occasions, relative to control mice. In none of our experiments was the duration of infection prolonged. We conclude that the presence of H. polygyrus does not interfere markedly with the host's ability to mount a protective response against B. microti and hence season dependent peaks of abundance of H. polygyrus in wild rodents are unlikely to present a particular threat to human communities by providing a greater reservoir of infection with B. microti in wild rodents than at other times of the year. PMID- 11269282 TI - Uncertainty on bioweapons treaty. PMID- 11269281 TI - A budget out of balance. PMID- 11269283 TI - Emerging diseases. Russia, NIH float big plan for former Soviet bioweapons lab. PMID- 11269284 TI - Paleoanthropology. Fossil tangles roots of human family tree. PMID- 11269285 TI - Space biology. New cuts in station could spark walkout. PMID- 11269287 TI - Dark matter. Astronomers glimpse galaxy's heavy halo. PMID- 11269286 TI - Paleontology. New fossil may change idea of first mollusk. PMID- 11269288 TI - Ecology. U.N. report suggests slowed forest losses. PMID- 11269289 TI - Superconductivity. Physicists scramble to recapture the magic. PMID- 11269290 TI - Pakistan. Shake-up in nuclear weapons program. PMID- 11269291 TI - Genomics. Chimp sequencing crawls forward. PMID- 11269292 TI - Foot-and-mouth disease. Barricading U.S. borders against a devastating disease. PMID- 11269293 TI - Science policy. Can a king of catalysis spur U.K. science to new heights? PMID- 11269294 TI - Condensed-matter physics. Doing the Bose nova with your main squeeze. PMID- 11269295 TI - Taxonomy. Linnaeus's last stand? PMID- 11269296 TI - Rescuing Venice from a watery grave. PMID- 11269297 TI - Ethical behavior as a stakes game. PMID- 11269298 TI - In a map for human life, count the microbes, too. PMID- 11269299 TI - Defining distress. PMID- 11269300 TI - Information access. Building a "GenBank" of the published literature. PMID- 11269301 TI - Science's response. Is a government archive the best option? PMID- 11269303 TI - Astronomy. The Birth of asteroseismology. PMID- 11269302 TI - Immunology. Memory T cells--local heroes in the struggle for immunity. PMID- 11269305 TI - Structural biology. Flipping a switch. PMID- 11269306 TI - Materials science. Watching grains deform. PMID- 11269304 TI - Development. The path to the heart and the road not taken. PMID- 11269307 TI - Dynamic combinatorial chemistry. PMID- 11269309 TI - Saving lives with sugar. PMID- 11269308 TI - Searching for medicine's sweet spot. PMID- 11269310 TI - Sugar separates humans from apes. PMID- 11269311 TI - After the fall. PMID- 11269312 TI - The best of both worlds? PMID- 11269313 TI - Bent out of shape. PMID- 11269314 TI - Toward automated synthesis of oligosaccharides and glycoproteins. AB - The discovery of previously unknown functions associated with carbohydrates and the study of their structure-function relations are of current interest in carbohydrate chemistry and biology. Progress in this area is, however, hampered by the lack of convenient and effective tools for the synthesis and analysis of oligosaccharides and glycoconjugates. Development of automated synthesis of such materials is necessary to facilitate research in this field. This review describes recent advances in carbohydrate synthesis, with particular focus on developments that have potential application to the automated synthesis of oligosaccharides, glycopeptides, and glycoproteins. PMID- 11269315 TI - Glycoprotein structure determination by mass spectrometry. AB - The human genome encodes 30,000 to 40,000 proteins, and a major challenge is to understand how posttranslational events, such as glycosylation, affect the activities and functions of these proteins in health and disease. Glycosylated proteins are ubiquitous components of extracellular matrices and cellular surfaces where their oligosaccharide moieties are implicated in a wide range of cell-cell and cell-matrix recognition events. The power of ultrahigh-sensitivity mass spectrometric strategies for defining the primary structures of highly complex mixtures of glycoprotein glycoforms is set to revolutionize structural glycobiology in the coming postgenomic era. PMID- 11269316 TI - Chemical glycobiology. AB - Chemical tools have proven indispensable for studies in glycobiology. Synthetic oligosaccharides and glycoconjugates provide materials for correlating structure with function. Synthetic mimics of the complex assemblies found on cell surfaces can modulate cellular interactions and are under development as therapeutic agents. Small molecule inhibitors of carbohydrate biosynthetic and processing enzymes can block the assembly of specific oligosaccharide structures. Inhibitors of carbohydrate recognition and biosynthesis can reveal the biological functions of the carbohydrate epitope and its cognate receptors. Carbohydrate biosynthetic pathways are often amenable to interception with synthetic unnatural substrates. Such metabolic interference can block the expression of oligosaccharides or alter the structures of the sugars presented on cells. Collectively, these chemical approaches are contributing great insight into the myriad biological functions of oligosaccharides. PMID- 11269317 TI - Intracellular functions of N-linked glycans. AB - N-linked oligosaccharides arise when blocks of 14 sugars are added cotranslationally to newly synthesized polypeptides in the endoplasmic reticulum (ER). These glycans are then subjected to extensive modification as the glycoproteins mature and move through the ER via the Golgi complex to their final destinations inside and outside the cell. In the ER and in the early secretory pathway, where the repertoire of oligosaccharide structures is still rather small, the glycans play a pivotal role in protein folding, oligomerization, quality control, sorting, and transport. They are used as universal "tags" that allow specific lectins and modifying enzymes to establish order among the diversity of maturing glycoproteins. In the Golgi complex, the glycans acquire more complex structures and a new set of functions. The division of synthesis and processing between the ER and the Golgi complex represents an evolutionary adaptation that allows efficient exploitation of the potential of oligosaccharides. PMID- 11269318 TI - Glycosylation and the immune system. AB - Almost all of the key molecules involved in the innate and adaptive immune response are glycoproteins. In the cellular immune system, specific glycoforms are involved in the folding, quality control, and assembly of peptide-loaded major histocompatibility complex (MHC) antigens and the T cell receptor complex. Although some glycopeptide antigens are presented by the MHC, the generation of peptide antigens from glycoproteins may require enzymatic removal of sugars before the protein can be cleaved. Oligosaccharides attached to glycoproteins in the junction between T cells and antigen-presenting cells help to orient binding faces, provide protease protection, and restrict nonspecific lateral protein protein interactions. In the humoral immune system, all of the immunoglobulins and most of the complement components are glycosylated. Although a major function for sugars is to contribute to the stability of the proteins to which they are attached, specific glycoforms are involved in recognition events. For example, in rheumatoid arthritis, an autoimmune disease, agalactosylated glycoforms of aggregated immunoglobulin G may induce association with the mannose-binding lectin and contribute to the pathology. PMID- 11269319 TI - Glycosylation of nucleocytoplasmic proteins: signal transduction and O-GlcNAc. AB - The dynamic glycosylation of serine or threonine residues on nuclear and cytosolic proteins by O-linked beta-N-acetylglucosamine (O-GlcNAc) is abundant in all multicellular eukaryotes. On several proteins, O-GlcNAc and O-phosphate alternatively occupy the same or adjacent sites, leading to the hypothesis that one function of this saccharide is to transiently block phosphorylation. The diversity of proteins modified by O-GlcNAc implies its importance in many basic cellular and disease processes. Here we systematically examine the current data implicating O-GlcNAc as a regulatory modification important to signal transduction cascades. PMID- 11269320 TI - Armed and dangerous: Toxoplasma gondii uses an arsenal of secretory proteins to infect host cells. AB - Toxoplasma gondii is a protozoan parasite that infects a wide variety of warm blooded animals and humans, in which it causes opportunistic disease. As an obligate intracellular parasite, T. gondii must invade a host cell to survive and replicate during infection. Recent studies suggest that T. gondii secretes a variety of proteins that appear to function during invasion or intracellular replication. These proteins originate from three distinct regulated secretory organelles called micronemes, rhoptries and dense granules. By discharging the contents of its secretory organelles at precise steps in invasion, T. gondii appears to timely deploy secretory proteins to their correct target destinations. Based on the timing of secretion and the characteristics of secretory proteins, an emerging theme is that T. gondii compartmentalizes its secretory proteins according to general function. Thus, it appears that micronemal proteins may function during parasite attachment to host cells, rhoptry proteins may facilitate parasite vacuole formation and host organellar association, and dense granule proteins likely promote intracellular replication, possibly by transporting and processing nutrients from the host cell. However, as more T. gondii secretory proteins are identified and characterized, it is likely that additional functions will be ascribed to each class of proteins secreted- by this fascinating invasive parasite. PMID- 11269321 TI - Schistosomiasis control in China. AB - This review on the control of schistosomiasis in China consists of the disease in the past, epidemiology, control programme, control approaches, achievements, problems existed and an appendix: the criteria for control and elimination of schistosomiasis in China. PMID- 11269322 TI - Preliminary study on genetic variability of Dicrocoelium dendriticum determined by random amplified polymorphic DNA. AB - Genetic variability of adult specimens of Dicrocoelium dendriticum has been studied using random amplified polymorphic DNA (RAPD). The worms were collected from the infected livers of different sheep from several localities in Leon province (NW Spain). DNA fragments were amplified by means of decamer primer oligonucleotides of arbitrary sequence. Some primers produce complex and highly variable patterns of amplified DNA in D. dendriticum. Intra- and inter-population genetic variability of adult parasites were analyzed, scoring polymorphic and monomorphic reproducible bands by means of the Jaccard similarity, and dendrograms showing genetic relationships between individuals were obtained using the FITCH method. Genetic variability seems to be high in this parasite and genetic similarity within a population (worms infecting a single animal) is similar to the average similarity between worms from different sheep. These results suggest that each sheep is infected by numerous genetically different parasites from one or more populations of infected ants. PMID- 11269323 TI - DNA immunization with Plasmodium falciparum serine repeat antigen: regulation of humoral immune response by coinoculation of cytokine expression plasmid. AB - We immunized mice with plasmid expressing the 47-kDa amino-terminal domain of the Plasmodium falciparum serine repeat antigen (SERA) using gene gun and investigated humoral immune response to SERA antigen. Significant SERA-specific IgG was observed in BALB/c mice after immunization three times with SERA expression plasmid. Furthermore, these levels were increased by the coinoculation of cytokine (IFN-gamma, IL-4, GM-CSF, or IL-12) expression plasmid. In respect to the SERA-specific Ig subclasses, coinoculation of IFN-gamma, GM-CSF, or IL-12 expression plasmid increased the levels of SERA-specific IgG2a, and these were much higher than that in mice immunized with SERA expression plasmid alone. In contrast to the SERA-specific IgG2a, coinoculation of any cytokine expression plasmid did not change the levels of SERA-specific IgG1. These results indicate that cytokine expression plasmid enhances and regulates humoral immune response elicited by SERA DNA immunization. PMID- 11269324 TI - Molecular cloning and functional expression of a cDNA encoding the major endoplasmic reticulum-associated calcium-binding protein, calreticulin, from Philippine strain Schistosoma japonicum. AB - We describe the cloning of a full length calreticulin (CR)-encoding cDNA clone isolated by immunoscreening of a cDNA library prepared with mRNA from adult worms of the Philippine strain of Schistosoma japonicum, the cause of Asian schistosomiasis. The sequence of the cDNA is presented, and its molecular characterisation and functional expression as a Ca2+-binding protein described. The potential role of CR in inducing protective immunity in the schistosomes is discussed. PMID- 11269326 TI - Differences in nucleosome organization over episomally located plasmids coincides with aberrant promoter activity in P. falciparum. AB - Here we investigated whether the Plasmodium falciparum GBP130 promoter maintains its developmental activity during the intraerythrocytic cycle when located on an episomal plasmid introduced using transient transfection. Comparing its activity with that of the endogenous chromosomally located GBP130 promoter indicates that the episomally located GBP130 promoter looses its developmental restriction, being rendered constitutively active. Loss of developmental restriction coincides with the absence of phased nucleosomal arrays over the episome. These data suggest that epigenetic factors may play a role in developmentally regulated gene expression in P. falciparum. PMID- 11269325 TI - Non-ulcerative cutaneous lesion in immunodeficient mice with Leishmania amazonensis infection. AB - Cutaneous leishmaniasis begins as papules or nodules at the site of promastigote inoculation. The next key pathogenic event in this disease is the formation of an ulcer at this site. Leishmania infection in immunodeficient mice, however, showed non-ulcerative cutaneous lesions suggesting the involvement of the immune system in ulcer formation. Severe combined immunodeficient (SCID), recombination activating gene 2 knockout (RAG-2-/-), and immunocompetent mice were inoculated subcutaneously with cultured L. amazonensis promastigotes. Macroscopic nodules appeared at the inoculation site within 2 weeks of infection in all the mice and gradually extended to the surrounding skin tissue. Although nodules of immunocompetent mice ulcerated within 6 weeks, immunodeficient mice did not form ulcers even after 25 weeks of inoculation. These results strongly suggest the importance of functional T and B cells in ulcer formation of cutaneous leishmaniasis and are consistent with clinical features of non-ulcerative cutaneous leishmaniasis in some AIDS patients. The present study also indicates that the L. amazonensis-infected immunodeficient mouse model might be suitable for studying the mechanisms of ulcer formation in cutaneous leishmaniasis. PMID- 11269327 TI - Differentiation of two Oesophagostomum spp. from pigs, O. dentatum and O. quadrispinulatum, by computer-assisted image analysis of fourth-stage larvae. AB - A method was developed to differentiate between fourth-stage larvae (L4) of two species of porcine nodular worms, Oesophagostomum dentatum and O. quadrispinulatum, by computer-assisted analysis of digitised microphotographs of L4 grown in vitro for various time periods and of L4 ex vivo. The overall lengths of the larvae and the lengths of the oesophagus as well as parameters describing the shape of the oesophagus and buccal capsule were measured and a formula based on these parameters was developed that could differentiate between the two species on the basis of the morphometric data. It was demonstrated that morphometry can produce unbiased data which can be employed for the calculation of indices suitable for the differentiation of morphologically different specimens. Computer-based techniques facilitate the processing of the complex data and offer the option for automation of measurements for routine applications. PMID- 11269328 TI - Immune response against protozoal and nematodal infection in mice with underlying Schistosoma mansoni infection. AB - Schistosoma mansoni infection induces T helper (Th) 2-dominant immune response in mice not only to S. mansoni itself but also to other coexisting antigens. In the present study, we challenged S. mansoni-infected mice with the intestinal nematode, Strongyloides venezuelensis, and the intracellular protozoa, Leishmania major to see whether such Th2-dominant immune responses alter susceptibility of the host to other concomitant parasitic infections. The recovery of S. venezuelensis adult worms from the small intestine was significantly decreased by S. mansoni infection, and the protection to S. venezuelensis appeared to act on migrating larvae. Antibodies elicited by S. mansoni infection showed cross binding to third-stage larvae antigen of S. venezuelensis. On the other hand, S. mansoni infection did not affect the outcome of L. major infection in both susceptible BALB/c and resistant C57BL/6 mice. Popliteal lymph node cells of BALB/c mice expressed mRNA for interleukin (IL)-10 rather than IL-4, regardless of S. mansoni infection, and those of C57BL/6 mice expressed IFN-gamma mRNA upon L. major antigen stimulation, even in S. mansoni-infected mice. Our findings suggest that Th2-dominant immune response induced by S. mansoni protects mice from intestinal helminthic infections, whereas they do not always modulate protozoal infections. PMID- 11269329 TI - Diagnosis of human neurocysticerocosis in endemic countries: a clinical study in Honduras. AB - With the purpose of evaluating the available methodology for neurocysticercosis (NCC) diagnosis, 60 neurological patients were studied during a 4-year period in Honduras. Neurological evaluation, Computed Tomography (CT), cysticercosis Enzyme Linked Immunoelectrotransfer blot (EITB) assay, electroencephalographic studies, and collection of epidemiological information were performed to assess a final diagnosis. The presenting clinical manifestations were: epileptic seizures (52%), headache without intracranial pressure (27%) and intracranial hypertension (10%). A protocol for the diagnosis of NCC is suggested. According to this protocol, patients with active (live) cysticercus and/or antibodies in Cerebrospinal fluid (CSF) were diagnosed as definitive cases of NCC, whereas those with only brain calcifications were diagnosed as probable cases. NCC diagnosis was definitive in 14 (23%) patients, probable in 32 (54%) and ruled out in 14 (23%). Of the patients with epileptic seizures, six (19%) had definitive and 20 (65%) had probable NCC. Overall seropositivity was 28%. EITB positivity varied from 14 to 100%, and from 20 to 35% in definitive and probable cases of NCC, respectively. When compared to CT, EITB overall sensitivity for definitive, active cases, was 50% in serum and 63% in CSF. These results suggest that brain images combined with neurological evaluation remains the best approach for neurocysticercosis diagnosis, and that EITB, even though its variable sensitivity, offers valuable information, especially if performed in CSF. PMID- 11269330 TI - Prevalence of Sarcocystis spp. cysts in Japanese and imported beef (Loin: Musculus longissimus). AB - A survey was carried out to investigate the occurrence of Sarcocystis infection in the loin (Musculus longissimus) of Japanese and imported beef. In all, the muscle tissue of 482 samples were examined by histological method. The prevalence of Sarcocystis unspecified species cysts was lower in Japanese beef (total 6.31%: 0% in Holstein castrated, 12.96% in Holstein milk cow, 3.33% in Japanese shorthorn and 11.58% in Japanese black cattle) than in beef imported from America (36.78%) or Australia (29.49%). The infection density of imported beef, especially in American, was higher than in Japanese beef. All detected cysts except one were identified as Sarcocystis cruzi. One thick walled cyst was found in Australian beef but it could not be distinguished as either Sarcocystis hirsuta or Sarcocystis hominis. PMID- 11269331 TI - Neurocysticercosis case with a single cyst in the brain showing dramatic drop in specific antibody titers within 1 year after curative surgical resection. AB - We report on one Japanese case subject with neurocysticercosis (NCC) who had a single cyst in the brain and had undergone curative surgical resection. Pathological examination revealed that it was a cysticercus of Taenia solium. Serological examination of the pair serum samples just before and 1 year after surgery revealed that antibody responses against the glycoproteins, highly specific to NCC and detected in the serum sample just before surgery by both immunoblot and ELISA became negative within 1 year after the surgery. It is, therefore, strongly suggested that this case had a single cysticercus in his whole body and the presence of a single cysticercus was sufficient to evoke antibody responses against it. PMID- 11269332 TI - Cutaneous gene transfer and therapy: the present and the future. AB - The easy accessibility of the skin as a therapeutic target provides an exciting potential for this organ for the development of gene therapy protocols for cutaneous diseases and a variety of metabolic disorders. Thus far, full phenotypic reversion of a diseased phenotype has been achieved in vivo for junctional epidermolysis bullosa and X-linked or lamellar ichthyosis and in vitro for xeroderma pigmentosum. These recessive skin diseases are characterized by skin blistering, abnormalities in epidermal differentiation and increased development of skin cancers, respectively. Corrective gene delivery at both molecular and functional levels was achieved by transduction of cultured skin cells using retroviral vectors carrying the specific curative cDNA. These positive results should prompt clinical trials based on transplantation of artificial epithelia reconstructed ex vivo using genetically modified keratinocytes. Promising results have also been obtained in phenotypic reversion of cells isolated from patients suffering from a number of metabolic diseases such as gyrate atrophy, familial hypercholesterolemia or phenylketonuria. In these diseases transplantation of autologous artificial epithelia expressing the transgenes of interest or direct transfer of the DNA to the skin represents a potential therapeutic approach for the systemic delivery of active molecules. Successful cutaneous gene therapy trials, however, require development of protocols for efficient gene transfer to epidermal stem cells, and information about the host immune response to the recombinant polypeptides produced by the implanted keratinocytes. The availability of spontaneous animal models for genodermatoses will validate the gene therapy approach in preclinical trials. PMID- 11269333 TI - Gene therapy for hemophilia. AB - Hemophilia A and B are X-chromosome linked recessive bleeding disorders that result from a deficiency in factor VIII (FVIII) and factor IX (FIX) respectively. Though factor substitution therapy has greatly improved the lives of hemophiliac patients, there are still limitations to the current treatment that have triggered interest in alternative treatments by gene therapy. Significant progress has recently been made in the development of gene therapy for the treatment of hemophilia A and B. These advances parallel the technical improvements of existing vector systems including MoMLV-based retroviral, adenoviral and AAV vectors, and the development of new delivery methods such as lentiviral vectors, helper-dependent adenoviral vectors and improved non-viral gene delivery methods. Therapeutic and physiologic levels of FVIII and FIX could be achieved in FVIII- and FIX-deficient mice and hemophilia dogs by different gene therapy approaches. Long-term correction of the bleeding disorders and in some cases a permanent cure has been realized in these preclinical studies. However, the induction of neutralizing antibodies often precludes stable phenotypic correction. Another complication is that certain promoters are prone to transcriptional inactivation in vivo, precluding long-term FVIII or FIX expression. Several gene therapy phase I clinical trials are currently ongoing in patients suffering from severe hemophilia A or B. No significant adverse side effects were reported, and semen samples were negative for vector sequences by sensitive PCR assays. Most importantly, some subjects report fewer bleeding episodes and occasionally have very low levels of clotting factor activity detected. The results from the extensive preclinical studies in normal and hemophilic animal models and encouraging preliminary clinical data indicate that the simultaneous development of different strategies is likely to bring a permanent cure for hemophilia one step closer to reality. PMID- 11269334 TI - Systematic analysis of clinically applicable conditions leading to a high efficiency of transduction and transgene expression in human T cells. AB - BACKGROUND: The transduction of human peripheral blood T cells with retroviral vectors constitutes an attractive approach for the correction of a number of genetic diseases. In this study we have conducted a systematic analysis of the relevance of a large number of parameters currently considered to affect the transduction of, and transgene expression in, human T cells. METHODS: Retroviral vectors encoding the human nerve growth factor receptor (NGFR) were used for transducing human T cells from normal volunteers. The proportion of T cells that expressed the marker transgene was determined by flow cytometry using anti-NGFR antibodies. RESULTS: Spinoculation and static fibronectin (FN)-assisted infections improved to a similar extent the transduction efficiency of PHA/IL-2 stimulated T cells, when compared with samples subjected to standard static infections. When immobilized anti-CD3 (anti-CD3i) or anti-CD3i/28i-stimulated T cells were considered, static infections in FN-coated plates were reproducibly more efficient than spinoculation infections performed on FN-uncoated plates. Under optimized manipulation conditions (three infection cycles of anti-CD3i/28i stimulated T cells in FN-coated plates) the total number of NGFR+ T cells harvested after 7 days of incubation represented, on average, twice the total number of T cells seeded at Day 0, and up to 95% of the human T cells efficiently expressed the marker transgene. Similar results were obtained regardless of whether samples were manipulated in medium supplemented with fetal bovine serum or with heat-inactivated autologous serum. CONCLUSIONS: Our study offers new experimental conditions for the transduction of human T cells, with obvious implications for the development of gene therapy protocols. PMID- 11269335 TI - Increasing endothelial cell permeability improves the efficiency of myocyte adenoviral vector infection. AB - BACKGROUND: Gene delivery to the myocardium using blood-borne adenoviral vectors is hindered by the endothelium, which represents a barrier limiting the infection rate of underlying myocytes. However, endothelial permeability may be modulated by pharmacological agents. METHODS: In the present study, we modeled the endothelial barrier in vitro using a human umbilical vein endothelial cell (HUVEC) monolayer seeded on a Transwell membrane as a support and diffusion of fluorescent dextrans as a permeability index. We used alpha-thrombin (100 nM) as a pharmacological agent known to increase endothelial permeability and tested the barrier function of the endothelial cell monolayer on adenovector-mediated luciferase gene transfer to underlying isolated cardiac myocytes. RESULTS: A confluent HUVEC monolayer represented a considerable physical barrier to dextran diffusion; it reduced the permeability of the micropore membrane alone to fluorescein isothiocyanate (FITC)-labeled dextrans of molecular weights 4, 70, 150 and 2000 kDa by approximately 54, 78, 88 and 98%, respectively. Alpha thrombin (100 nM) increased the permeability coefficients (P(EC)) by 276, 264, 562 and 4166% for the same dextrans, respectively. A confluent HUVEC monolayer represented a major impediment to adenovector-mediated luciferase gene transfer to cardiac myocytes, largely reducing gene transfer efficiency. However thrombin induced a nine-fold increase in myocyte infection. CONCLUSION: In our model, the endothelial cell monolayer represents a major impediment to myocyte adenovector mediated gene transfer which can be partially improved by pharmacologically increasing endothelial permeability. The Transwell model is therefore particularly useful for testing the efficiency of pharmacological agents in modulating adenovector passage through the endothelial barrier. PMID- 11269336 TI - Stabilization of transgenes delivered by recombinant adenovirus vectors through extrachromosomal replication. AB - BACKGROUND: A major limitation of adenovirus-mediated gene therapy for metabolic and inherited diseases is the instability of transgene expression in vivo. This instability results, at least in part, from the inability of the vector genome to maintain the transgene through replication or integration. In this study we evaluated the possibility of stabilization of an adenovirus-delivered transgene by non-adenovirus replicative elements. METHODS: We have developed a novel system for the maintenance of transgenes delivered by adenovirus vectors through extrachromosomal replication. In its initial configuration, this system combines the Epstein-Barr virus (EBV) replicative elements, a tetracycline (Tc)-inducible expression system, and the Cre-lox recombination system in the context of a single E1/E3/E4-deleted adenovirus vector. Induction of Cre expression initiates a Cre-mediated recombination, resulting in the excision of a fragment of the vector genome and its circularization into an EBV-based episome. RESULTS: In vitro studies have demonstrated that excision of the circular episome can occur in a cell-free system as well as in cultured cells transfected with plasmid DNA or transduced by a virus vector carrying the episome-excising cassette. PCR studies have shown that in proliferating, non-permissive, cultured primate cells the episome generated from the adenovirus vector is maintained much more stably than the genome of the parent vector. This episome was also able to replicate in mammalian cells. CONCLUSION: Together these studies demonstrate the feasibility of this approach for the stabilization of transgenes delivered to dividing cells by adenovirus vectors. PMID- 11269337 TI - Optimised helper virus-free production of high-quality adeno-associated virus vectors. AB - BACKGROUND: Clinical development of adeno-associated virus (AAV) requires standardised, safe, efficient and scalable procedures for the manufacture of the rAAV vector, including production, purification and testing. Several strategies have been reported for the approach to the manufacturing problem. We report a helper virus-free process that produces high quality rAAV stocks. METHODS: rAAV were produced by triple transfection, a helper virus-free process. After lysis of the cells in the presence of nuclease, the rAAV produced were purified by HPLC through two ion-exchange columns in tandem followed by dialysis. rAAV stocks were thoroughly characterised for biological activity and for the presence of residual contaminants. The titer of infectious particles and of rep + particles was determined by dRA assay. Contaminating DNA and RNA were determined by fluorescent dye binding and real-time PCR. The protein content of the rAAV stocks was characterised by SDS-PAGE, ELISA test, Western blot and specific enzymatic assays for putative residual contaminating protein. The in vivo biological activity of the stocks was evaluated in mouse muscle. RESULTS: rAAV stocks obtained following this procedure elicit: 2-5 x 10(12) pp/ml; 3-6 x 10(10) ip/ml; < 10(3) rep + particles/ml; <0.3 mUeq/ml of residual benzonase activity; non-detectable Ad or beta-galactosidase proteins; <35 pg/ml of cellular genomic DNA; in vivo expression in mouse muscle without any immune reaction detected. CONCLUSIONS: This work demonstrates the possibility of producing purified high-quality rAAV free of helper virus. The procedure described in this paper is easily adaptable for large-scale production of clinical rAAV vectors. PMID- 11269338 TI - Atomic force microscopy imaging of DNA-cationic liposome complexes optimised for gene transfection into neuronal cells. AB - BACKGROUND: Cationic liposomes represent an important gene delivery system due to their low immunogenicity, but are relatively inefficient, with optimisation of DNA-liposome complexes (lipoplexes) for transfection necessary for each cell type of interest. There have been few studies examining optimisation in neuronal cell types or determining how the structure of lipoplexes affects transfection efficiency. METHODS: Four commercially available cationic liposome formulations were used to optimise transfection efficiency in neuronal cells. The DNA to liposome ratio and the amount of DNA used in transfections were varied. Transfection efficiency was determined by the percentage of cells positive for the micro-galactosidase reporter gene product. The structure of lipoplexes was studied using atomic force microscopy. Lipoplexes were characterised further using dynamic light scattering to determine size and fluorescence techniques to show DNA compaction. RESULTS: Optimal transfection conditions were found to differ between immortalised cell lines and primary cells. High transfection efficiencies in immortalised cell lines were achieved predominantly with multivalent cationic liposomes while primary neuronal cells showed optimal transfection efficiency with monovalent cationic liposomes. The structure of lipoplexes was observed with atomic force microscopy and showed globular complexes for multivalent cationic liposomes, while monovalent liposomes gave less compact structures. In support of this finding, high levels of DNA compaction with multivalent liposomes were observed using fluorescence quenching measurements for all DNA to liposome ratios tested. One monovalent liposome showed increasing levels of compaction with increasing liposome amount. Dynamic light scattering showed little change in complex size when the different lipoplexes were studied. CONCLUSIONS: Optimisation of transfection efficiency was different for cell lines and primary neurons. Immortalised cells showed optimal transfection with multivalent liposomes while primary neurons showed optimal transfection with monovalent liposomes. The charge ratio of the monovalent liposome was below one, suggesting a different mechanism of lipoplex binding and uptake in primary neurons. The structure of lipoplexes, as PMID- 11269341 TI - The Journal of Gene Medicine 2000 Young Investigator Award. Identification of a new cis-acting replication element (CARE) in the AAV-2 genome involved in viral DNA replication and increase in vector titers. PMID- 11269339 TI - Cationic liposome-mediated gene transfer to rat salivary epithelial cells in vitro and in vivo. AB - BACKGROUND: Previously we have shown that gene transfer to salivary gland epithelial cells readily occurs via recombinant adenoviruses, although the response is short-lived and results in a potent host immune response. The aim of the present study was to assess the feasibility of using cationic liposomes to mediate gene transfer to rat salivary cells in vitro and in vivo. METHODS: Initially, for transfection in vitro, we used two cationic liposome formulations (GAP-DLRIE/DOPE and DOSPA/DOPE) complexed with plasmid encoding human growth hormone (hGH) as a reporter gene. Thereafter, using GAP-DLRIE/DOPE, plasmids were transferred to rat salivary glands in vivo, and hGH levels measured in saliva, serum and gland extracts. RESULTS: Under optimal conditions, transfection of rat submandibular glands (SMGs) was consistently observed. Approximately 95% of the cells transfected with a plasmid encoding beta-galactosidase were acinar cells. Maximal hGH expression was obtained during the first 48 h post-transfection using a plasmid encoding the hGH cDNA and complexed with GAP-DLRIE/DOPE. hGH was detected in gland extracts and saliva, and occasionally in serum. No systemic or local gland pathology was consistently or significantly observed. CONCLUSIONS: The levels of the reporter gene product, hGH, obtained after GAP-DLRIE/DOPE mediated gene transfer are considerably lower (<0.5%) than those achieved with adenoviral vectors (10(8) PFU). Nonetheless, cationic liposome-mediated gene transfer to salivary glands may be useful for potential therapeutic applications. PMID- 11269342 TI - Identification of YAC clones containing the mutable slender glume locus slg in rice (Oryza sativa L.). AB - A mutable slender glume gene slg, which often reverts to the wild-type state, was induced by gamma-ray irradiation of seeds of the japonica rice cultivar 'Gimbozu'. The final goal was to understand whether the slender glume mutation was associated with the insertion of a transposable element, utilizing map-based cloning techniques. The RFLP (restriction fragment length polymorphism) analysis revealed that the slg locus was located between two RFLP loci, XNpb33 and R1440, on chromosome 7 with recombination values of 3.1% and 1.0%, respectively. Using these two RFLP loci as probes, five YAC (yeast artificial chromosome) clones containing either of these two loci were selected from a YAC library. Subsequently, both end fragments of these YAC clones, amplified by the inverse PCR (IPCR) method, were used to select new YAC clones more closely located to the slg locus. After repeating such a procedure, we successfully constructed a 6-cM YAC contig, and identified four overlapping YAC clones, Y1774, Y3356, Y5124, and Y5762, covering the slg locus. The chromosomal location of the slg was narrowed down to the region with a physical distance of less than 280 kb between the right end fragments of Y1774 and Y3356. PMID- 11269343 TI - An efficient method for the physical mapping of transgenes in barley using in situ hybridization. AB - The genetic transformation of crops by particle bombardment and Agrobacterium tumefaciens systems have the potential to complement conventional plant breeding programmes. However, before deployment, transgenic plants need to be characterized in detail, and physical mapping is an integral part of this process. Therefore, it is important to have a highly efficient method for transgene detection by fluorescence in situ hybridization (FISH). This study describes a new approach, which provides efficient control of probe length and labelling, both of which play an important role in in situ hybridization of transgenes. The approach is based on reducing the size of the plasmid prior to labelling by nick translation, rather than using the whole or linearized plasmid, or varying the amounts of DNaseI in the nick translation mixture. This provided much more efficient labelling of the probe, which yielded optimal hybridization. minimal fluorescent background, and accurate physical location of the transgene. PMID- 11269344 TI - Towards an expanded and integrated linkage map of cucumber (Cucumis sativus L.). AB - Linkage maps in cucumber (Cucumis sativus var. sativus L.) have been constructed using morphological traits, isozymes, restriction fragment length polymorphisms (RFLPs), and random amplified polymorphic DNAs (RAPDs). The lack of polymorphism in cucumber has led to the construction of relatively unsaturated maps (13- to 80 point). We have added amplified fragment length polymorphism (AFLP) markers to existing narrow-based (within C. sativus) and wide-based (C. sativus x C. sativus var. hardwickii) maps. JOINMAP v. 2.0 was used to construct maps and to join these with historical maps from several previous studies. Our narrow- and wide based merged maps contain 255 and 197 markers, respectively, including morphological traits, disease resistance loci, isozymes, RFLPs, RAPDs, and AFLPs. Condensation of total map distance occurred in merged maps compared to historic maps using many of the same markers. This phenomenon is most likely due to differences in map construction algorithms. The merged maps represent the best fit of the data used and are an important first step towards the construction of a comprehensive linkage map for cucumber. Identification of additional anchor markers between the narrow- and wide-based maps presented here may allow their future integration into a unified model. PMID- 11269345 TI - Sau3A in situ digestion of human chromosome 3 pericentrometric heterochromatin. I. Differential digestion of alpha-satellite and satellite 1 DNA sequences. AB - In situ digestion with the restriction endonuclease (RE) Sau3A (Sau3A REISD) uncovers a polymorphism for the pericentromeric heterochromatin of human chromosome 3, which can be positively stained (3+) or not (3-), and has proven useful to differentiate donor and recipient cells after sex-matched bone marrow transplantation and to analyze the so-called hemopoietic chimerism. The aim of the present investigation was to obtain insight into the molecular basis of such polymorphism to optimize its use for chimerism quantification using methodological approaches other than REISD. To this end, fluorescence in situ hybridization (FISH) assays using probes for the satellite DNA sequences that mainly constitute chromosome 3 pericentromeric heterochromatin (alpha-satellite and satellite 1 DNA) were performed on control and Sau3A-digested chromosomes. The results obtained suggest that chromosome 3 alpha-satellite DNA is digested in all individuals studied, irrespective of the karyotype obtained by Sau3A REISD (3++, 3+-, 3--), and thus it does not seem to be involved in the polymorphism uncovered by Sau3A on this chromosome. Satellite 1 DNA is not digested in any case, and shows a polymorphism for its domain size, which correlates with the polymorphism uncovered by Sau3A in such a way that 3+ chromosomes show a large domain (3L) and 3- chromosomes show a small domain (3S). It seems, therefore, that the cause of the polymorphism uncovered by Sau3A on the pericentromeric region of chromosome 3 is a difference in the size of the satellite 1 DNA domain. Small satellite 1 DNA domains fall under the resolution level of REISD technique and are identified as 3-. PMID- 11269346 TI - Different chromosomal distribution patterns of radiation-induced interchange breakpoints in barley: first post-treatment mitosis versus viable offspring. AB - Translocation breakpoints (TBs) induced by ionizing radiation are nonrandomly distributed along barley chromosomes. When first post-treatment mitoses were evaluated, centromeres and the heterochromatin-containing proximal segments tended to be more than randomly involved, and terminal segments to be less than randomly involved in translocations. Contrary to this, small chromosomal regions in median and distal arm positions, characterized by high recombination rates and high gene density, were identified as preferred sites for the origination of viable translocations, probably due to deviations in chromatin organization. Apparently, the position of a TB has an influence on the rate of viability versus elimination of the carrier cells. Surprisingly, TBs within centromeres and heterochromatin-containing segments seem to be more harmful for survival than those induced in gene-rich regions. PMID- 11269347 TI - Random amplified polymorphic DNA diversity of marginal and central populations in Pinus contorta subsp. latifolia. AB - Fifteen populations of lodgepole pine (Pinus contorta subsp. latifolia) were surveyed for diversity across 52 random amplified polymorphic DNAs (RAPDs). The objective was to compare single-locus and multilocus structures in four marginal, three intermediate, and eight central populations. Single-locus estimates indicated average observed and expected heterozygosity to be 0.19 and 0.17, respectively. When these estimates were split into population categories, a clear trend of increasing diversity was detected in the direction of marginal to central populations. F-statistics indicated an excess of heterozygotes, with F(IS) ranging from -0.08 for marginal populations to -0.15 for central populations and averaging -0.12 over 15 populations. The estimates of F(ST) decreased towards the margins of the species range, indicating increased population differentiation. Forty-nine of 52 RAPDs tested neutral in the Ewens Watterson analysis. Multilocus analysis showed significant two-locus and high order gametic disequilibria in all 15 populations. The most prominent components of the two-locus analysis were the variance of disequilibrium (VD, 46.2%) and the multilocus Wahlund effect (31.9%). This high value for VD indicated that founder effects could explain much of the observed multilocus associations. When analyzed by population categories, the VD showed a decreasing trend indicating that variation due to founder effects was more prominent in marginal populations. The two-locus Wahlund effect (WC) that is characteristic of strong population subdivision was highest in the central populations. This indicated significant levels of gene flow between populations with different allelic combinations. PMID- 11269348 TI - Molecular cytogenetic analysis of DNA sequences with flanking telomeric repeats in Triticum aestivum cv. Begra. AB - A cloned genomic DNA fragment (pTa241) formerly derived from a DNA fraction obtained from isolated nuclei of embryos of a Polish cultivar of wheat (Triticum aestivum cv. Begra) comprises a tandem repeat of the telomeric array CCCTAAA, and hybridizes in situ exclusively to the telomeres of all chromosome arms of the somatic chromosome complement of wheat. A second cloned fragment (pTa637) derived from the same fraction is 637 bp long, flanked by 28 bp of the same telomeric repeat unit, and hybridizes in situ to the entire lengths of all the chromosomes of the complement. The same pattern of hybridization was observed when the flanking telomeric sequences were removed. A third DNA fragment (pTa1439), derived from unfractionated genomic DNA and flanked with 62 bp of the same telomeric unit, showed the same patterns of distribution. Together with additional evidence from Southern analysis, these observations were interpreted to mean that these sequences are associated with mobile DNA elements and are distributed widely throughout the genome. The chromosomal distribution of the non telomeric parts of the clones is consistent with the dispersed genomic distribution characteristic of transposons and retroelements. PMID- 11269349 TI - Production of durum wheat substitution haploids from durum x maize crosses and their cytological characterization. AB - The objective of this study was to investigate the effect of individual durum wheat (Triticum turgidum L.) chromosomes on crossability with maize (Zea mays L.) and to cytologically characterize the haploids recovered. Fourteen 'Langdon' (LDN) D-genome disomic substitution lines, a LDN Ph mutant (Ph1b ph1b), and normal 'Langdon' were pollinated with maize pollen. After pollination, hormonal treatment was given daily for up to 14 days. Haploid embryos were obtained from all lines and were aseptically cultured. From a total of 55,358 pollinated florets, 895 embryos were obtained. Only 14 of the embryos germinated and developed into healthy plants. Different substitution lines showed varying degrees of success. The most successful was the substitution 5D(5B) for both embryo formation and haploid plantlet production. These results indicate that the substitution of 5D for 5B confers on durum wheat a greater ability to produce haploids. Fluorescent genomic in situ hybridization (GISH) showed that the substitution haploids consisted of 7 A-genome chromosomes, 6 B-genome chromosomes, and 1 D-genome chromosome. Triticum urartu Turn. genomic DNA was efficient in probing the 7 A-genome chromosomes, although the D-genome chromosome also showed intermediate hybridization. This shows a close affinity between the A genome and D genome. We also elucidated the evolutionary translocation involving the chromosomes 4A and 7B that occurred at the time of evolution of durum wheat. We found that the distal segment translocated from chromosome 7B constitutes about 24% of the long arm of 4A. PMID- 11269350 TI - Organization of 5S ribosomal RNA genes in tea (Camellia sinensis). AB - The 5S rRNA genes in the Camellia sinensis (L.) O. Kuntze (tea) genome are arranged as tandem repeat units of 300 and 325 bps. The 2 classes of tandem repeats were discovered by Southern hybridisation of tea genomic DNA with a 5S rRNA gene PCR product. PMID- 11269351 TI - Development of a STS marker assay for detecting loss of heterozygosity in rice hybrids. AB - The RAPD (random amplified polymorphic DNA) markers OPE15750 and OPE15300 were affected by loss of heterozygosity (LOH) in rice hybrids AMR x 'M202' and AMR x 'L202'. The markers were mapped to the same locus at or near the centromere of rice chromosome 2. The two RAPD products were cloned, sequenced, and found to have lengths of 734 bp and 297 bp, respectively. The 297-bp sequence shares a 98% homology with one end of the 734-bp sequence, accounting for the cross hybridization previously observed between the two clones. Based on the DNA sequence of the 734-bp fragment, a pair of STS (sequence-tagged site) primers was designed and tested. Rice plants homozygous for either OPE15734 or OPE15297 all produced PCR fragments of the same length, 482 bp. However, the two PCR products were discernible by digestion with the restriction enzyme XbaI prior to gel electrophoresis. The STS product from plants homozygous for OPE15734 was cut into two fragments of 239 and 240 bp, which appeared as one single band in an agarose gel; whereas the STS product from plants homozygous for OPE15297 was not cut by XbaI and was characterized by a 482-bp band in the agarose gel. These STS primers were tested in rice hybrids and F2 progenies derived from the hybrids of AMR x 'M202' and AMR x 'L202'. Homozygosity for OPE15297 was confirmed for all F2 panicle rows derived from AMR x 'M202'. In contrast, F2 panicle rows of AMR x 'L202' exhibited two different segregation patterns (genotypes), i.e., either uniformly homozygous for the 240-bp marker (OPE15734/OPE15734) or segregating for the 482- and 240-bp markers (OPE15734/OPE15297). This STS-marker system provides a robust assay for detecting the occurrence of LOH in rice hybrid progenies. PMID- 11269352 TI - Pattern of X-Y chromosome pairing in the Taiwan vole, Microtus kikuchii. AB - Pairing of X and Y chromosomes at meiotic prophase and the G- and C-banding patterns and nucleolar organizer region (NOR) distribution were analyzed in Microtus kikuchii. M. kikuchii is closely related to M. oeconomus and M. montebelli, karyologically and systematically. The formation of a synaptonemal complex between the X and Y chromosomes at pachytene and end-to-end association at diakinesis--metaphase I are only observed in three species in the genus Microtus; M. kikuchii, M. oeconomus, and M. montebelli. All the other species that have been studied so far have had asynaptic X-Y chromosomes. These data confirm that M. kikuchii, M. oeconomus, and M. montebelli are very closely related, and support the separation of asynaptic and synaptic groups on the phylogenetic tree. PMID- 11269353 TI - A physical map with yeast artificial chromosome (YAC) clones covering 63% of the 12 rice chromosomes. AB - A new YAC (yeast artificial chromosome) physical map of the 12 rice chromosomes was constructed utilizing the latest molecular linkage map. The 1439 DNA markers on the rice genetic map selected a total of 1892 YACs from a YAC library. A total of 675 distinct YACs were assigned to specific chromosomal locations. In all chromosomes, 297 YAC contigs and 142 YAC islands were formed. The total physical length of these contigs and islands was estimated to 270 Mb which corresponds to approximately 63% of the entire rice genome (430 Mb). Because the physical length of each YAC contig has been measured, we could then estimate the physical distance between genetic markers more precisely than previously. In the course of constructing the new physical map, the DNA markers mapped at 0.0-cM intervals were ordered accurately and the presence of potentially duplicated regions among the chromosomes was detected. The physical map combined with the genetic map will form the basis for elucidation of the rice genome structure, map-based cloning of agronomically important genes, and genome sequencing. PMID- 11269354 TI - Identification of barley genome segments introgressed into wheat using PCR markers. AB - Barley has several important traits that might be used in the genetic improvement of wheat. For this report, we have produced wheat-barley recombinants involving barley chromosomes 4 (4H) and 7 (5H). Wheat-barley disomic addition lines were crossed with 'Chinese Spring' wheat carrying the phlb mutation to promote homoeologous pairing. Selection was performed using polymerase chain reaction (PCR) markers to identify lines with the barley chromosome in the ph1b background. These lines were self pollinated, and recombinants were identified using sequence-tagged-site (STS) primer sets that allowed differentiation between barley and wheat chromosomes. Several recombinant lines were isolated that involved different STS-PCR markers. Recombination was confirmed by allowing the lines to self pollinate and rescreening the progeny via STS-PCR. Progeny testing confirmed 9 recombinants involving barley chromosome 4 (4H) and 11 recombinants involving barley chromosome 7 (5H). Some recombinants were observed cytologically to eliminate the possibility of broken chromosomes. Since transmission of the recombinant chromosomes was lower than expected and since seed set was reduced in recombinant lines, the utility of producing recombinants with this method is uncertain. PMID- 11269355 TI - Large-scale mutagenesis directed at specific chromosomes in wheat. AB - A novel approach has been developed to allow for the efficient selection of loss of-function wheat mutants in the M1 generation, following either physical or chemical mutagenesis. This has generated an order of magnitude increase in the efficiency of identification of mutants, and also greatly increases the likelihood that selected individuals reflect mutation events at the target locus, rather than at genes acting elsewhere in the disease resistance pathway. The approach relies only on prior knowledge of the chromosomal location of the target gene, and uses the polyploidy of wheat to construct populations for mutagenesis in which large numbers of individuals are hemizygous for the target gene. The idea is illustrated with the mass identification of mutants at three independent genes for race-specific resistance to yellow rust, and one gene for resistance to powdery mildew. PMID- 11269357 TI - Development of sequence-characterized amplified regions (SCARs) from amplified fragment length polymorphism (AFLP) markers tightly linked to the Vf gene in apple. AB - Amplified fragment length polymorphism (AFLP) markers have become widely used in saturating the region of a gene of interest for the ultimate goal of map-based cloning of the gene or for marker-assisted selection. However, conversion of AFLP markers into restriction fragment length polymorphism (RFLP) or polymerase chain reaction (PCR)-based markers will greatly expand their usefulness in genetic applications. Previously, we have identified 15 AFLP markers tightly linked to the Vf gene conferring scab resistance in apple. In this study, we have successfully converted 11 of these AFLPs into sequence-characterized amplified region (SCAR) markers. Of the remaining four nonconverted AFLP markers, one, ET2MC8-1, has been found to be very short (83 base pairs) and is an A/T rich (90%) marker; a second, EA2MG11-1, has shown identical sequences between Malus floribunda 821 (the original source of the Vf gene) and scab-susceptible apple cultivars; while the other two, EA12MG16-1 and ET8MG1-1, have not been cloned. Using the 11 converted SCAR markers along with 5 previously identified SCAR markers, a high-resolution linkage map around the Vf gene has been constructed, and found to be consistent with its corresponding AFLP map. Three converted SCAR markers (ACS-3, -7, and -9) are inseparable from the Vf gene; whereas one (ACS-6) is located left of, and the remaining seven (ACS-1, -2, -4, -5, -8, -10, and -11) are located right of the Vf gene at genetic distances of 0.4 and 0.2 cM, respectively. A reliable and robust procedure for development of SCAR markers from AFLP markers is presented. PMID- 11269356 TI - Development of microsatellite markers in potato and their use in phylogenetic and fingerprinting analyses. AB - Three genomic libraries were constructed using a mixture of DNA from Solanum phureja Juz. & Buk., and S. chacoense Bitt. Two of the libraries were enriched for ATT and GT repeats (a 27-fold enrichment was achieved). In total, 3500 clones of the conventional library, 1,000 of the library enriched for ATT, and 12,000 of the one enriched for GT were screened with five different repeat motifs, and a total of 18 primer pairs was obtained. Another group of 12 primer pairs was obtained from the SSR-containing sequences in the public databases (18 SSR containing sequences were utilized). From among 30 newly developed primer pairs, 12 previously published ones, and 12 pairs developed for tomato, 7 were used to identify 12 different potato cultivars and introductions, and 12 were used to study phylogenetic distance among seven wild and cultivated potato species. Two SSR markers were sufficient to discriminate the 12 cultivars. The mean number of alleles per polymorphic locus was 5 for the 12 cultivars and 4.5 for the seven species. The results obtained in this study confirm those achieved in similar studies in other plant species regarding the abundance and use of SSR markers in identifying species and cultivars. PMID- 11269358 TI - The porcine gonadotropin-releasing hormone receptor gene (GNRHR): genomic organization, polymorphisms, and association with the number of corpora lutea. AB - The interaction of gonadotropin-releasing hormone (GNRH) and its receptor (GNRHR) is critical in the endocrine regulation of reproduction. The gene (GNRHR) encoding the receptor has been mapped to porcine chromosome 8. There is evidence for three quantitative trait loci (QTL) influencing ovulation rate on this chromosome. We obtained an almost complete sequence (3993 bp, excluding intron 1) of the porcine GNRHR gene using PCR-based comparative genomic walking and inverse genomic walking approaches. Twelve polymorphisms were detected by sequencing of pooled DNA of Chinese Taihu and European Large White pigs, including 7 base substitutions and 5 insertions-deletions (indels). A F2 population of Meishan x European Large White pigs was genotyped for a TG indel in the promoter region, and a C/G substitution in the 3' UTR (untranslated region). A significant association of the C/G substitution with number of corpora lutea at first parity was observed. PMID- 11269359 TI - Autoradiographic study of transcription and dosage compensation in the sex and neo-sex chromosome of Drosophila nasuta nasuta and Drosophila nasuta albomicans. AB - Cellular autoradiography is used to study the transcription patterns of the polytene X chromosomes in Drosophila nasuta nasuta and D. n. albomicans. D. n. nasuta, with 2n = 8, includes a pair of complete heteromorphic sex chromosomes, whereas D. n. albomicans, with 2n = 6, has a pair of metacentric neo-sex chromosomes representing incomplete heteromorphic sex chromosomes. The neo-X chromosome has two euchromatic arms, one representing the ancestral X while the other represents the ancestral autosome 3 chromosomes. The metacentric neo-Y chromosome has one arm with a complete heterochromatic ancestral Y and the other arm with a euchromatic ancestral autosome 3. The transcription study has revealed that the X chromosome in D. n. nasuta is hyperactive, suggesting complete dosage compensation, while in the neo-X chromosome of D. n. albomicans the ancestral X chromosome is hyperactive and the ancestral autosome 3, which is part of the neo sex chromosome, is similar to any other autosomes. This finding shows dosage compensation on one arm (XLx/-) of the neo-X chromosome, while the other arm (XR3/YR3) is not dosage compensated and has yet to acquire the dosage compensatory mechanism. PMID- 11269360 TI - Genetic mapping of resistance factors to Phytophthora palmivora in cocoa. AB - Phytophthora palmivora causes pod rot, a serious disease on cocoa widespread throughout the producing regions. In order to ascertain the genetic determination of cocoa resistance to P. palmivora, a study was carried out on two progenies derived from crosses between a heterozygous, moderately resistant Forastero clone, T60/887, and two closely related and highly susceptible Forastero clones, one completely homozygous, IFC2, and one partially heterozygous, IFC5. The cumulative size of both progenies was 112 individuals. Plants were subjected to natural and artificial inoculation of P. palmivora in C te d'Ivoire. The genetic maps of T60/887 and of IFC5 were constructed using amplified fragment length polymorphism (AFLP) markers and microsatellites. The map of T60/887 comprised 198 markers assembled in 11 linkage groups and representing a total length of 793 cM. The map of IFC5 comprised 55 AFLP markers that were assembled into six linkage groups for a total length of 244 cM. Ratio of rotten over total number of fruit under natural infection was measured for each tree over two harvests. Artificial inoculations were performed on leaves and pods. These tests were weakly correlated with the pod rot rate in the field. Five quantitative trait loci (QTLs) of resistance were detected for T60/887 but none were common between the three traits measured. Stability and reliability of the experimental procedures are discussed and revealed the difficult use of these artificial tests on adult trees for a good prediction of field resistance. PMID- 11269361 TI - Characterization of euploid backcross progenies derived from interspecific hybrids between Oryza sativa and O. eichingeri by restriction fragment length polymorphism (RFLP) analysis and genomic in situ hybridization (GISH). AB - Restriction fragment length polymorphism (RFLP) analysis and GISH (genomic in situ hybridization) were performed on euploid plants derived from crosses between Oryza sativa (2n = 24, AA) and two brown planthopper-resistant accessions of O. eichingeri (2n = 24, CC). After screening with 164 RFLP markers, 60 of the 67 euploid plants were identified as introgression lines, each carrying 1-6 small O. eichingeri segments integrated on chromosomes 1, 2, 6, or 10. In the somatic chromosome preparations of F1 hybrid, O. eichingeri chromosomes, fluorescing greenish-yellow in the sequential GISH, appeared to be longer and to contain more heterochromatin than O. sativa ones, and this karyotypic polymorphism can be used to detect some introgressed O. eichingeri segments in euploid plants. In addition, GISH identification presented direct evidence for the transfer of small segments from O. eichingeri to O. sativa chromosome(s) which were subsequently recognized according to their condensation pattern, arm ratio, and chromosome length. The present results would contribute to the molecular mapping and selection of O. eichingeri--derived brown planthopper-resistant gene and positive yield QTLs. PMID- 11269362 TI - Centromeric tandem repeat from the chaffinch genome: isolation and molecular characterization. AB - A new family of avian centromeric satellites is described. The highly repeated sequence, designated FCP (Fringilla coelebs PstI element), was cloned from the 500-bp PstI digest fraction of the chaffinch (Fringilla coelebs L.) genomic DNA, sequenced, and characterized. The FCP repeat was found to have 505-506 bp length of monomer, 57% content of GC, to compose about 0.9% of the chaffinch genome, and to be highly methylated. Results of Southern-blot hybridization of cloned FCP element onto genomic DNA digested with different restriction enzymes, and sequencing directly from total genomic DNA using FCP-specific primers and ThermoFidelase enzyme (Fidelity Systems Inc.) were in agreement with a tandem arrangement of this repeat in the chaffinch genome. Five positions of single nucleotide polymorphism (SNP) were found in the FCP monomers using direct genomic sequencing. Fluorescence in situ hybridization (FISH) with FCP probe and primed in situ labelling (PRINS) with FCP specific primers showed that the FCP elements occupy pericentric regions of all chaffinch chromosomes. On chromosome spreads, the fluorescent signals were also observed in the intercentromeric connectives between nonhomologous chromosomes. The results suggest that the centromeric FCP repeat is responsible for chromosome ordering during mitosis in chaffinch. PMID- 11269363 TI - Enterovirus 71: the virus, its infections and outbreaks. AB - Enterovirus 71 (EV71) was first recognized in 1974. Since then it has been implicated in 13 small and large outbreaks world-wide. Large outbreaks of hand, foot and mouth disease (HFMD), mostly benign, occurred in Japan in 1973 and 1978. Four outbreaks with brain stem encephalitis and significant numbers of deaths occurred in Bulgaria and Hungary in the late 1970's and in Malaysia and Taiwan in 1997 and 1998 respectively. During the latter two epidemics, pulmonary edema and hemorrhage often leading to quick deaths in children aged from 0.5 to 3 years old was first recognized. In Taiwan 78 deaths and over 100,000 cases of HFMD occurred. Coxsackie A16 cocirculated with EV 71, without however, causing any severe illnesses. The transmission of EV 71 was related to number of siblings in a household, rural residence and contact with cases of HFMD. Genotype analyses show that genotypes have changed with time in the United States and Japan. Recent isolates from Japan are similar to the isolates from Malaysia and Taiwan in 1997 and 1998, respectively. Even though genotype analysis has not identified specific sequences responsible for neurovirulence, the strains causing brain stem encephalitis and pulmonary edema in the Far East are similar and have arisen since 1997. Seroepidemiological studies in Taiwan suggest that children aged from 0.5 to 4 years old are most susceptible while the rest of the population are over 50% immune. Theoretically there is a pool of such susceptible subjects every few years. In prevention for another major outbreak, a simple, inactivated Salk type vaccine should be immediately prepared and made available. PMID- 11269364 TI - Using buffy coat for reverse transcriptase-polymerase chain reaction in the diagnosis of dengue virus infection: preliminary study. AB - Using reverse transcriptase-polymerase chain reaction (RT-PCR) to detect and type from viremic human serum samples for dengue virus infection is widely used today. However, a few false-negative results were reported due to very low titers of the virus particle in serum samples. As mononuclear cells, macrophages or monocytes are target cells for dengue virus infection, and the replication of virions can be observed in peripheral leukocytes frequently, the amount of virus particle in buffy coat should be higher than those in serum samples. Here, we describe a procedure in which RNA extraction from the buffy coat of a patient with a false negative serum sample yielded specific viral RNA amplifiable by RT-PCR, thereby providing an alternative choice for the accurate diagnosis of dengue infection. PMID- 11269365 TI - High protease activity of Chryseobacterium indologenes isolates associated with invasive infection. AB - In order to understand virulence factors of Chryseobacterium indologenes isolates associated with invasive infection, enzymatic activities and cellular fatty acid profiles of 42 isolates recovered at National Taiwan University Hospital from January 1994 to December 1996 were studied. Among them, 12 blood isolates were considered as invasive and 30 (recovered from urine, sputa, infected burn wounds, and catheter tips) were noninvasive. All isolates showed strong activities of alkaline phosphatase, acid phosphatase, naphthol-AS-BI-phosphohydrolase, and N acetyl-beta-glucosaminidase, and had no activities for alpha-galactosidase, beta galactosidase, beta-glucuronidase, alpha-mannosidase, and alpha-fucosidase. The activities of other enzymes were variable. Thirty-two isolates (76%) had varying degrees of protease activity. Two profiles (profiles I and II) of cellular fatty acids of the isolates were found and profile I predominated. There was no significant difference of distribution of cellular fatty acid profiles and activities of enzymes between invasive and noninvasive isolates, except protease activity which was significantly higher in invasive isolates than that in noninvasive isolates. Protease activity may play an important role in virulence on invasive infections caused by C. indologenes. PMID- 11269366 TI - The evaluation of allergens and allergic diseases in children. AB - Knowing the incidence of allergic diseases and their relationship with allergens is a crucial requirement for therapeutic judgment. We present our experience on the incidence, clinical features and allergens of the allergic diseases detected by multiple allergosorbent chemiluminescent assay (MAST-CLA) in children from 1997 to 1999 at the Taipei Veterans General Hospital. The incidence of bronchial asthma, allergic rhinitis and atopic dermatitis are significantly different when stratified by age groups. Among the enrolled 2008 patients, 980 (48.8%) patients have positive MAST-CLA results. Of these, 562 (57.3%) are male and 418 (42.7%) are female. A significant increase among patients with positive allergens is also found when stratified by age group. Inhalant allergen is the major allergen detected in our patients. House dust mites Dermatophagoides pteronyssinus (Dp) and Dermatophagoides farinae (Df), cockroaches, feathers, and dog dander show the highest incidence in the 7- to 12-year-old group. In the fungal group, Aspergillus and Penicillium also show a significant difference in the incidence among different age groups. Pollen allergens, on a whole, show significant difference in incidence among different age groups. The food allergen group shows variable significant difference in incidence. Crab, milk, and egg white show the highest significant incidence in the 2- to 6-year-old group. These results suggest that the incidence of allergens detected in allergic diseases varies among different age groups. PMID- 11269367 TI - Household distribution of house dust mite in central Taiwan. AB - House dust mite (HDM) is a common inhalant allergen which can precipitate atopic disease episodes including asthma and allergic rhinitis. However, the relationship between HDM and asthma in subtropical regions of Asia, which may be affected by differences in climate and environmental variables, has not been widely studied. To assess this relation in the subtropical region of central Taiwan, we collected HDM samples from the houses of eight asthmatic patients as well as four normal subjects over a 1-year period. HDMs were collected by vacuum from the following four areas: living room floor, sofa, the top surface of child's mattress and bedroom floor. The mite concentrations, site distribution, seasonal variation, individual species and correlation with asthmatic attacks were studied. The HDM concentration had a seasonal variation, with the highest concentrations noted from July to November with gradually decrease from December to June. Among the four areas of collection, the highest concentration of mites was found on the child's mattress (p < 0.05). Dermatophagoides pteronyssinus was the dominant species (77%) and Dermatophagoides farinae was the second (13%). Our data showed that: 1. The highest concentrations of HDM occurred during the period from July to November. 2. The child's mattress was the household region with the highest percentage of HDM and thus should be considered of great concern as a likely source of the exacerbation of asthma. 3. D. pteronyssinus was the dominant species. PMID- 11269368 TI - Mastoiditis: a disease often overlooked by pediatricians. AB - Although mastoiditis can be a life threatening disease, clinicians often overlook it because it is uncommon. We reviewed the presentation and management of all children younger than 15 years of age with the discharge diagnosis of mastoiditis in our hospital from January 1994 through December 1999. Nineteen patients that fulfilled the case definition were included. The most common clinical presentation in this series was fever. More specific findings, such as otorrhea, postauricular pain, swelling, and redness of mastoid could be found in less than half of these patients. Only two patients had characteristic physical findings, and mastoiditis was diagnosed in only three patients upon admission. Plain radiographic evidence of mastoiditis was usually not apparent early in the course. In this series, the majority of patients were diagnosed by computed tomography (CT) scans. The present study demonstrates that mastoiditis most commonly presents without a clearly diagnostic set of physical examination and laboratory findings. Mastoiditis should be considered in patients with otitis media or with fever of unknown origin (FUO). The empirical antibiotic treatment should cover organisms commonly found in acute otitis media (AOM), including Haemophilus influenzae, Streptococcus pneumoniae, and Moraxella catarrhalis. PMID- 11269369 TI - Outcomes of Aeromonas bacteremia in patients with different types of underlying disease. AB - Over a 6-year period, 42 patients with different underlying diseases developed Aeromonas bacteremia in our hospital. The male to female ratio was 2:1. The vast majority of these patients had underlying diseases, including various types of neoplasm (n = 14), liver cirrhosis (n = 11), biliary tract disorder (n = 10) and other illnesses (n = 7). Community-acquired bacteremia was predominant (33 cases, 79%). Aeromonas hydrophila was the most common species isolated (88%). Monomicrobial bacteremia was more common than polymicrobial bacteremia (64% vs 36%). Monomicrobial bacteremia was associated with neoplasm or liver cirrhosis in 80% of patients. Polymicrobial bacteremia was more common in patients with biliary tract disorder than in patients from other groups (60% vs 40%). Escherichia coli (60%) was the predominant concomitant organism isolated. The major clinical manifestations were fever (74%), jaundice (57%), and abdominal pain (45%). Recognized infection sites included biliary tract, soft tissue involvement, peritoneal involvement, while 50% of patients had no recognized infection site. Eight patients (80%) received cholecystectomy due to gall stone with acute cholecystitis. However, none of the cirrhotic patients with necrotizing fasciitis received surgical treatment. The mortality attributed to Aeromonas bacteremia was 70%. Patients with liver cirrhosis or malignancy had a higher acute mortality (death within 7 days after admission) than the other patients (89% vs 11%). We conclude that Aeromonas bacteremia can cause a rapidly fatal outcome and should be considered an important pathogen for septicemia in patients with liver cirrhosis or neoplasm. PMID- 11269370 TI - Complications of varicella in children: emphasis on skin and central nervous system disorders. AB - A review of medical records at a tertiary hospital in southern Taiwan from June 1988 through May 1998 identified 136 children who had been hospitalized for varicella-related complications. Of the children, 83% (113/136) were healthy before the onset of varicella and 17% (23/136) had underlying illnesses. The mean age was 4.7 years (ranged from 1 day to 18 years) with a male predominance (1.7:1). The mean hospital stay was 5.5 days (ranged from 1 to 22 days). Secondary bacterial skin or soft tissue infections were the most common complications (44%), followed by central nervous system (CNS) involvement (23%), pneumonia (18%), thrombocytopenia (12%), and liver function impairment (10%). Among the 60 patients with secondary bacterial cutaneous infection, 16 (27%) had positive isolates, including 12 isolates of Staphylococcus aureus and four Streptococcus pyogenes. Age above 8 years was significantly associated with the development of varicella-associated CNS complications (p = 0.019). Of the 23 immunocompromised hosts, the most common underlying conditions were hematological diseases (11 patients, 48%), followed by neonatal varicella (7 patients, 30%) and chronic illness with steroid treatment (5 patients, 22%). All of the subjects in this study had a favorable outcome except for three lethal cases, resulting in a case-fatality rate of 2.2%. The cause of death was S. aureus septicemia in one patient, streptococcal toxic shock syndrome in one patient, and encephalitis with brain herniation in one patient. Our results demonstrate that varicella continues to be a serious disease that occasionally results in life-threatening complications in healthy and immunocompromised children. Routine immunization of all healthy children against varicella is recommended. PMID- 11269371 TI - Antimicrobial susceptibility and species identification for clinical isolates of enterococci. AB - In order to describe the susceptibility patterns and determine the identities of Enterococcus spp. isolated at a regional teaching hospital in Taichung during the period from June through November 1998, 96 clinical isolates of enterococci were collected for further analysis. Major sources of these isolates included urine, wound, and pus. Species identification was performed using the API 20 Strep system and supplemental tests. The minimum inhibitory concentrations (MICs) of six antimicrobial agents were determined by E-test for each isolate. Disk diffusion tests were also performed and the results were compared with those reported by the clinical laboratory. Because gentamicin susceptibility tests showed inconsistent results in many isolates, MIC determinations by the microbroth dilution method were also performed for these isolates. All isolates were tested for beta-lactamase production using the chromogenic method. The results showed that Enterococcus faecalis was the most frequently encountered species (86.5%), followed by Enterococcus faecium (7.3%), Enterococcus avium (5.2%) and Enterococcus casseliflavus (1.0%). The MIC90 of ampicillin, penicillin, vancomycin, teicoplanin, ciprofloxacin and gentamicin for total isolates were 1, 3, 2, 0.25, 1, and more than 1024 microg/mL, respectively. All isolates were susceptible to vancomycin and teicoplanin. The same rate of resistance (3.1%) was found to penicillin, ampicillin and ciprofloxacin in all isolates. There were 50 (52%) and 48 (50%) isolates with high level streptomycin and gentamicin resistance, respectively. The MIC90 of ampicillin and penicillin for E. faecium were significantly higher than those for other species (96 and > 256 vs. < or = 1 and < or = 3 microg/mL, respectively); in fact, ampicillin or penicillin-resistance was only found in E. faecium. No organism was found to produce beta-lactamase. There were 29 isolates showing discrepant results between the study findings and clinical laboratory report for gentamicin susceptibility. Most isolates (27/ 29) reported as susceptible to gentamicin by the clinical laboratory showed a high level gentamicin resistance by MIC determinations. The inconsistent results may be due to discrepancy in interpretations for heterogeneous resistance, bias in selecting colonies for the disk diffusion test, or variation in the quality of Mueller-Hinton agar used. The results suggest that any pure isolates growing some colonies within the inhibition zone should be considered as gentamicin resistant even if the zone diameter is equal or greater than the susceptible breakpoint. In order to obtain accurate results, the gentamicin MIC should be determined by the dilution method for enterococcal isolates that yield intermediate inhibition zones or zones just above the susceptible limit. PMID- 11269372 TI - Comparison of antimicrobial susceptibility of Citrobacter freundii isolates in two different time periods. AB - Citrobacter freundii was first identified in 1932, since then it has been reported to cause a variety of infections in aged, immunocompromised, and debilitated patients. With the use of broad-spectrum antibiotics, C. freundii has become increasingly resistant to antimicrobial agents. In order to determine the chronological changes in susceptibility and current susceptibility status of C. freundii, we compared the antimicrobial susceptibility of C. freundii in two different time periods, from 1987 to 1988 and from 1997 to 1998. In both time periods, 61 isolates of C. freundii were randomly selected for study from all clinical isolates at National Taiwan University Hospital. The minimum inhibitory concentrations and susceptible rates of 15 antimicrobial agents were compared, and it was found that most C. freundii isolates were resistant to anti pseudomonal penicillins, first, second, and third generation cephalosporins, gentamicin, tobramycin, and aztreonam. The results indicate that the susceptible rates of C. freundii to aminoglycosides and ciprofloxacin decreased markedly during the period from 1987 to 1998. Cefepime, cefpirome, imipenem, and meropenem remained the most active agents against C. freundii. PMID- 11269373 TI - Childhood Churg-Strauss syndrome: report of a case. AB - Churg-Strauss syndrome (CSS) (allergic granulomatosis and angitis) is an uncommon form of systemic vasculitis, which is rare in children. It is characterized by peripheral blood hypereosinophilia, systemic necrotizing vasculitis, and a preceding history of bronchial asthma. We described a boy with initial presentation of poorly controlled bronchial asthma, allergic rhinitis, recurrent sinusitis and several episodes of hemoptysis since the age of 9. He then developed purpuric skin lesions, generalized soreness, and symptoms of mononeuritis multiplex at age 11. On admission to our hospital at the age of 12, he developed marked pericardial effusion. After a series of studies including chest computed tomography (CT), skin biopsy, nerve conduction study, and serological tests for autoantibodies, CSS was diagnosed. Thereafter, he received regular corticosteroid therapy, and his symptoms were generally well-controlled with occasional acute exacerbation. The clinical characteristics, diagnosis and management of CSS in children are also reviewed. PMID- 11269374 TI - Neonatal fungemia caused by Hansenula anomala: a case report. AB - Hansenula anomala, an ascosporogenous yeast of the class Ascomycetes, is a free living organism isolated from the environment. It is also a part of the normal or transient flora of the human throat and alimentary tract. It has been recognized as an opportunistic pathogen and its infection is very rare. A premature infant, a victim of right femoral osteomyelitis and right hip arthritis caused by oxacillin-resistant Staphylococcus aureus, was found to have developed H. anomala fungemia just before the initiation of the antimicrobial therapy with teicoplanin. Antifungal agents (fluconazole and amphotericin B) were prescribed for 10 days despite the absence of clinical sign of systemic fungal infection. His general condition remained good, with a subsequent sterile blood culture. The patient was discharged after completing 5 weeks of antimicrobial therapy, and he remained well during follow-up at our outpatient clinics. Here, we also review the risk factors, the clinical presentations, and the therapeutic strategies of H. anomala infection in the literature. PMID- 11269375 TI - Guidelines for antimicrobial therapy of urinary tract infections in Taiwan. PMID- 11269376 TI - Design of a unique PCR primer for detection of oral HPV infection. AB - Human papillomavirus (HPV) has been known as a pathogen of oral dysplasia. However, suitable PCR primers for the detection of oral HPV infection have not been reported. The aim of this study was to design unique consensus primers. The consensus primers were designed by homologues analyses between subtypes 2, 6, 11, 13, 16, 18, 30, 32 and 58, which frequently infect the oral membrane. PCR, with our designed primer, detected HPV DNA subtypes 2, 6, 11, 16, 18 and 58, and also showed PCR product from a clinical papilloma sample. These results indicate that our designed consensus primer can be used for the study of the relationships between oral disease and HPV infection. PMID- 11269377 TI - The effects of functional treatment on the sagittal position of the mandibular condyle. AB - The aim of the current study was to evaluate the effects of functional treatment appliances, a U-Bugel type I activator and a conventional activator, on the mandibular condyle considering the sagittal direction. Alterations in the growth direction and quantity of the condyle effects the ramal inclination and, consequently, the whole mandible. This situation is important when considering treatment prognosis and stability. The material consisted of pre- and post treatment lateral cephalograms and hand-wrist films of 49 individuals having skeletal and dental Class II/div.1 malocclusions. Although the elimination of overjet and correction of Class II/div.1 anomalies was achieved, statistically significant changes were not observed in the sagittal direction of the mandibular condyle. PMID- 11269378 TI - Oral biopsy turnaround time: 20-year experience of the Department of Oral Pathology, Oral Medicine and Periodontology, Faculty of Dentistry, University of Malaya. AB - The turnaround time (TAT) for oral biopsies received for histological examination by the Department of Oral Pathology, Oral Medicine and Periodontology, Faculty of Dentistry, University of Malaya, for the years 1978, 1988 and 1998 was evaluated. For the three years studied, TATs for 61, 233 and 463 specimens were retrospectively analysed. Testing intervals, that is, from the dates the surgeons procured the specimens, the laboratories accessioned them and until the pathologists signed off the diagnoses, were used to calculate TAT. The performance level of the respective pathologists, the growth of tissue diagnostic services and the possible variables that influence TAT were also evaluated. As prompt diagnosis means prompt treatment, which in turn has a bearing on prognosis, the TAT pertinent to oral malignant tumors was emphasized. The mean TAT, its mode and median fell significantly in 1998 compared with the previous 2 years; it was lower for soft tissue than for hard tissue specimens, and lower for malignant, than for non-malignant specimens. The progression of tissue diagnostic services is up to a satisfactory level, as 88.89 % of biopsies could render diagnoses within a fair period of time in 1998. PMID- 11269379 TI - Diagnostic efficacy of three-dimensional images by helical CT for lesions in the maxillofacial region. AB - The purpose of this study was to evaluate the efficacy of three-dimensional (3-D) images produced with a helical CT for the diagnosis of lesions occurring in the maxillofacial region. Thirty-four patients, who had lesions in the maxillofacial region, were examined by plain radiography (intra and extraoral) and the helical CT. Further, 3-D images were reconstructed from the data provided by the helical CT using the volume rendering method. These images were compared with plain radiographic images and conventional two-dimensional (2-D) CT images in terms of the information they provided for diagnosis. Using the 3-D images for tumors, bone destruction, inner components, extent of the lesion,the relationship between the lesion and surrounding anatomical landmarks, and the roots of the adjacent teeth were observed in overall views. We conclude that 3-D images produced by helical CT may provide useful information for the diagnosis of lesions. PMID- 11269380 TI - Detection of mitochondrial DNA mutations in human gingival tissues. AB - The accumulation of mitochondrial DNA mutation is an important contributor to the aging process. Polymerase chain reaction (PCR) amplification was carried out on total DNA from gingival tissues of human subjects with ages of 19 to 64 years, and 5.0-kb and 7.4-kb deletions were found in the mitochondrial genomes of these subjects. This is the first report of mitochondrial DNA deletion detectable in human gingival tissues. PMID- 11269381 TI - Helicobacter pylori in oral ulcerations. AB - Helicobacter pylori is an important pathogen involved in the development of gastrointestinal ulcers, but its involvement in oral ulcerous lesions is unclear. As culture is generally recognized as the gold standard for diagnosis of H. pylori infection, we employed this approach to assess the association of H. pylori with oral mucosal ulcerations. Samples were collected from patients with oral mucosal ulcerative disorders: 12 cases of recurrent aphthous stomatitis (RAS), 7 cases of herpes simplex virus (HSV) stomatitis, and 3 cases of erosive lichen planus (LP). Serum IgG antibodies against H. pylori were examined in all cases. All of the RAS and erosive LP cases were culture-negative for H. pylori, while two cases of HSV stomatitis were positive. The two culture-positive cases were also seropositve for the H. pylori antigen. It is suggested that H. pylori might not have a direct association with oral ulcerations. However, H. pylori in the oral cavity might exist in a non-culturable coccoid state without productive infection, and might form colonies only under special conditions such as HSV infection. PMID- 11269382 TI - Relationship between the quantity of gingival crevicular fluid and clinical periodontal status. AB - In order to analyze the possible relationship between the quantity of gingival crevicular fluid (GCF) and clinical periodontal status, the severity of gingival inflammation (gingival index (GI) scores) and probing depth (PD) were recorded and GCF samples were obtained from 1,111 sites. These sites were further analyzed on the basis of distinct tooth groups to evaluate the significance of particular anatomical sampling locations. Statistical analysis of cumulative data showed significant increases in GCF volume with greater GI scores and PD. Correlations between GCF volume and both of the clinical measures were also strongly positive and significant for all sites. However, significant differences in GCF volume were observed between the anterior and posterior sampling sites. Increases in volume with increasing GI and PD were more marked for incisor and canine teeth. Similarly, the relationship between the quantity of GCF and clinical periodontal status was more clear and absolute in the anterior region than in the premolar and molar areas. These findings suggest that the quantity of GCF is not constant throughout the entire dentition, and that the relationship between GCF measurements and clinical periodontal status is site-based. This unique feature of GCF seems to be an essential factor in the design of GCF-related studies. PMID- 11269383 TI - Profiles of cytokine expression in radicular cyst-lining epithelium examined by RT-PCR. AB - Levels of messenger RNA (mRNA) extracted from five samples of radicular cyst lining epithelium were analyzed for cytokines, growth factors and epithelial cell growth-related receptors by RT-PCR. All five samples expressed IL-1alpha, -1beta, IL-6, IL-8, IL-11, TGF-beta1, PDGF-A and aFGF, and receptors for EGF (c-erbB), KGF, HGF (c-met) and IL-6. Some of the specimens expressed MIP-1alpha, RANTES, GM CSF, M-CSF, TNF-alpha, PDGF-B and bFGF, but no expression of IL-2, IL-4, IFN gamma, IGF-I, EGF and KGF was detected. These results indicate that radicular cyst-lining epithelium, which is considered to be identical to the cell rests of Malassez, may play a role in periodontal pocket formation or apical cyst formation by interaction with surrounding connective tissue or hematopoietic cells through the expression of various cytokines. PMID- 11269384 TI - Oral myiasis caused by hypoderma bovis larvae in a child: a case report. AB - Myiasis is the invasion of living tissue of humans and other mammals by the eggs or larvae of flies of the order of Diptera. It occurs mainly in the tropic, and is associated with inadequate public and personal hygiene. Oral myiasis in humans appears to be rare. This article records a case of oral myiasis caused by larvae of Hypoderma bovis. Two different pathologic soft tissue sockets were observed in the vestibular sulcus at the level of the both deciduous laterals along the deep upper lip tissues. PMID- 11269385 TI - Parental autonomy granting during adolescence: exploring gender differences in context. AB - This study investigated the ways in which 2 indicators of parental autonomy granting, adolescents' decision-making input and parental knowledge of adolescents' daily experiences, differed as a function of contextual factors (i.e., parents' gender role attitudes or sibling dyad sex composition) and boys' and girls' personal qualities (i.e., gender, pubertal status, developmental status, or birth order) in a sample of 194 families with firstborn (M = 15.0 years) and second-born (M = 12.5 years) adolescents. Firstborns were granted more autonomy than second borns, especially in families with firstborn girls and second-born boys. Girls in families marked by traditional maternal gender role attitudes were granted fewer autonomy opportunities. Postmenarcheal second-born girls were granted more opportunities for autonomy than were premenarcheal second born girls, but only in families with less traditional maternal gender role attitudes. PMID- 11269386 TI - Maternal intrusive support in the academic context: transactional socialization processes. AB - Although transactional models of socialization have received support, there has been little investigation of the processes involved. The goal of this research was to move in this direction in the context of the socialization of achievement. Mothers and their elementary school children (N = 166) took part in an 18-month longitudinal study including a 2-week daily checklist. The results suggested that children's low achievement elicits intrusive support from mothers through 2 mechanisms. Mothers worried over their children's performance, and this was associated with heightened intrusive support. Children's low achievement manifested itself in uncertainty, which was linked to heightened intrusive support. The achievement of children whose mothers frequently used intrusive support improved over time but did not exceed that of children whose mothers infrequently used intrusive support. Day-to-day analyses suggested that although intrusive support promotes success, it also fosters failure for low-achieving children. PMID- 11269388 TI - Viewing imitation as child responsiveness: a link between teaching and discipline domains of socialization. AB - The authors observed 106 children's imitation and responses to maternal control at 14 and 22 months. Imitation was observed in a teaching task in which mothers modeled 3 standard pretend-play sequences. Responses to control were observed in typical discipline contexts. Girls imitated more than boys. Responsive imitation measures were coherent and longitudinally stable and correlated significantly with responsiveness to maternal control. The authors propose that a young child's willingness to imitate his or her parent in a teaching context and to comply in a control context both reflect a responsive or receptive stance toward parental socialization. The consistency of children's responsiveness across contexts has implications for both sociomoral and cognitive development. PMID- 11269387 TI - The consolidation/transition model in moral reasoning development. AB - The consolidation/transition model conceptualizes development as entailing a cyclical pattern of alternating consolidation and transition phases and posits that stage advance is predicted by a specific distribution of reasoning across stages indicative of disequilibrium (more reasoning above than below the mode, with a high degree of mixture). The validity of this model was examined in the context of moral reasoning development with the use of standard statistical techniques as well as Bayesian techniques that can better account for classification error. In this longitudinal study. 64 children and adolescents participated in 5 annual administrations of the Moral Judgment Interview. The distribution of their reasoning across stages was used to predict subsequent development. The results support the hypotheses regarding cyclical patterns of change and predictors of stage transition and demonstrate the utility of Bayesian techniques for evaluating developmental change. PMID- 11269389 TI - Do adolescent symptomatology and family environment vary over time with fluctuations in paternal alcohol impairment? AB - This study tested whether adolescent internalizing problems, externalizing problems, heavy alcohol use, fathers' parenting, and family conflict varied over time with fluctuations in fathers' alcohol impairment and also whether children of recovered alcoholic fathers differed from children of nonalcoholic fathers. Fathers and adolescent children (N = 267 families) were interviewed in 3 annual assessments. Results showed that adolescent symptomatology and the family environment did not vary over time as a function of different trajectories of paternal alcohol impairment. However, children of recovered alcoholic fathers exhibited more symptomatology than did children of nonalcoholic fathers. Even though paternal alcoholism has remitted in these families, children of recovered alcoholic fathers might remain on a general higher risk trajectory relative to children of nonalcoholic fathers. PMID- 11269390 TI - Japanese children's numerical competencies: age- and schooling-related influences on the development of number concepts and addition skills. AB - Using a cutoff design (J. Bisanz, F. J. Morrison, & M. Dunn, 1995) to separate school-related influences from those that are age related, the study investigated the development of number concepts and addition skills in Japanese children. Three groups of kindergarten and 1st grade children who differed in age and/or school experiences completed tasks on their numerical competencies 1 and 6 months after school entrance. Children's use of addition strategies, rather than their solution accuracy, changed primarily as a function of schooling, not age. Children's Base 10 number concepts improved with the amount of schooling, as well as with other social and age-related factors. Results suggest that schooling is an important determinant in developing Japanese-speaking children's numerical competencies, which were not explained solely by their language characteristics or by age-related factors. PMID- 11269391 TI - The development of cognitive and academic abilities: growth curves from an early childhood educational experiment. AB - In the Abecedarian Project, a prospective randomized trial, the effects of early educational intervention on patterns of cognitive and academic development among poor, minority children were examined. Participants in the follow-up were 104 of the original 111 participants in the study (98% African American). Early treatment was full-time, high-quality, educational child care from infancy to age 5. Cognitive test scores collected between the ages of 3 and 21 years and academic test scores from 8 to 21 years were analyzed. Treated children, on average, attained higher scores on both cognitive and academic tests, with moderate to large treatment effect sizes observed through age 21. Preschool cognitive gains accounted for a substantial portion of treatment differences in the development of reading and math skills. Intensive early childhood education can have long-lasting effects on cognitive and academic development. PMID- 11269392 TI - Body image across the life span in adult women: the role of self-objectification. AB - This study aimed to investigate women's body image across the entire life span from within the theoretical perspective provided by objectification theory (B. L. Fredrickson & T.-A. Roberts, 1997). In a cross-sectional study, a sample of 322 women ranging in age from 20 to 84 years completed a questionnaire measuring body dissatisfaction, self-objectification, and its proposed consequences. Although body dissatisfaction remained stable across the age range, self-objectification, habitual body monitoring, appearance anxiety, and disordered eating symptomatology all significantly decreased with age. Self-objectification was found to mediate the relationship between age and disordered eating symptomatology. It was concluded that objectification theory helps clarify the processes involved in the changes in body image that occur with age. PMID- 11269394 TI - Lexical input as related to children's vocabulary acquisition: effects of sophisticated exposure and support for meaning. AB - A corpus of nearly 150,000 maternal word-tokens used by 53 low-income mothers in 263 mother-child conversations in 5 settings (e.g., play, mealtime, and book readings) was studied. Ninety-nine percent of maternal lexical input consisted of the 3,000 most frequent words. Children's vocabulary performance in kindergarten and later in 2nd grade related more to the occurrence of sophisticated lexical items than to quantity of lexical input overall. Density of sophisticated words heard and the density with which such words were embedded in helpful or instructive interactions, at age 5 at home, independently predicted over a third of the variance in children's vocabulary performance in both kindergarten and 2nd grade. These two variables, with controls for maternal education, child nonverbal IQ, and amount of child's talk produced during the interactive settings, at age 5, predicted 50% of the variance in children's 2nd-grade vocabulary. PMID- 11269393 TI - The younger siblings of teenage mothers: a follow-up of their pregnancy risk. AB - This study followed 243 younger brothers and younger sisters of parenting teens and nonparenting teens across a 1.5-year period. The average age of siblings was 13.6 years at Time 1 and 15 years at Time 2. Relative to other youths, the sisters of parenting teens exhibited a sharp increase in drug and alcohol use and partying behavior across time and had the highest pregnancy rate at Time 2 (15%). The siblings of parenting teens spent 10 hr a week caring for their sisters' children, and, for girls, many hours of child care was associated with negative outcomes including permissive sexual behavior. Findings suggest that the younger sisters of parenting teens are at very high risk of early pregnancy and that this risk becomes increasingly pronounced across time. PMID- 11269395 TI - Large-scale production of the carbohydrate portion of the sialyl-Tn epitope, alpha-Neup5Ac-(2-->6)-D-GalpNAc, through bacterial coupling. AB - Alpha-Neup5Ac-(2-->6)-D-GalpNAc, the carbohydrate portion of sialyl-Tn epitope of the tumor-associated carbohydrate antigen, was prepared by a whole-cell reaction through the combination of recombinant Escherichia coli strains and Corynebacterium ammoniagenes. Two recombinant E. coli strains overexpressed the CMP-Neup5Ac biosynthetic genes and the alpha-(2-->6)-sialyltransferase gene of Photobacterium damsela. C. ammoniagenes contributed to the production of UTP from orotic acid. Alpha-Neup5Ac-(2-->6)-D-GalpNAc was accumulated at 87 mM (45 g/L) after a 25-h reaction starting from orotic acid, N-acetylneuraminic acid, and 2 acetamide-2-deoxy-D-galactose. PMID- 11269396 TI - Synthesis of Neu5Ac- and Neu5Gc-alpha-(2-->6')-lactosamine 3-aminopropyl glycosides. AB - In order to prepare 3-aminopropyl glycosides of Neu5Ac-alpha-(2-->6')-lactosamine trisaccharide 1, and its N-glycolyl containing analogue Neu5Gc-alpha-(2-->6') lactosamine 2, a series of lactosamine acceptors with two, three, and four free OH groups in the galactose residue was studied in glycosylations with a conventional sialyl donor phenyl [methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5 dideoxy-2-thio-D-glycero-alpha- and beta-D-galacto-2-nonulopyranosid]onates (3) and a new donor phenyl [methyl 4,7,8,9-tetra-O-acetyl-5-(N-tert butoxycarbonylacetamido)-3,5-dideoxy-2-thio-D-glycero-alpha- and beta-D-galacto-2 nonulopyranosid]onates (4), respectively. The lactosamine 4',6'-diol acceptor was found to be the most efficient in glycosylation with both 3 and 4, while imide type donor 4 gave slightly higher yields with all acceptors, and isolation of the reaction products was more convenient. In the trisaccharides, obtained by glycosylation with donor 4, the 5-(N-tert-butoxycarbonylacetamido) moiety in the neuraminic acid could be efficiently transformed into the desired N-glycolyl fragment, indicating that such protected oligosaccharide derivatives are valuable precursors of sialo-oligosaccharides containing N-modified analogues of Neu5Ac. PMID- 11269397 TI - Synthesis and antiperoxidant activity of new phenolic O-glycosides. AB - We describe the synthesis of some 3-tert-butyl-4-hydroxyphenyl D-glycopyranosides by reaction of tert-butylhydroquinone with beta-D-pentaacetyl-glucose, beta-D pentaacetyl-galactose, 2-acetamido- and 3,4,6-tri-O-acetyl-2-butanamido-2-deoxy beta-D-glucopyranosyl chlorides as well as the formation of anomeric 3-tert-butyl 4-hydroxyphenyl 4,6-di-O-acetyl-2,3-dideoxy-D-erythro-hex-2-eno-pyranosides by reaction between tert-butylhydroquinone and 3,4,6-tri-O-acetyl-D-glucal. All compounds, except 3-tert-butyl-4-hydroxyphenyl alpha- and beta-D glucopyranosides, inhibited lipid peroxidation with a degree of potency comparable to that of tert-butyl hydroxyanisole. PMID- 11269398 TI - The formation of 2-hydroxy-4-hydroxymethyl-3-(indol-3-yl)-cyclopent-2-enone derivatives from ascorbigens. AB - A facile preparation is described of 3-(indol-3-yl)-2-hydroxy-4 hydroxymethylcyclopent-2-enone and its N-derivatives in 15-40% yields by the degradation of ascorbigen or its N-derivatives in a warm solution of L-ascorbic acid through a sequential domino reaction. The same cyclopentenone derivatives were obtained in 30-40% yields by the condensation of (N-alkylindol-3-yl)glycolic acids with ascorbic acid. 2,6-Dihydroxy-1-(indol-3-yl)hexa-1,4-diene-3-one and 2 hydroxy-4-hydroxymethyl-5-(indol-3-yl)cyclopent-2-enone were identified as intermediates in this reaction. PMID- 11269399 TI - Conjugating oligosaccharides to proteins by squaric acid diester chemistry: rapid monitoring of the progress of conjugation, and recovery of the unused ligand. AB - Samples that are periodically withdrawn from the mixture of a conjugation reaction can be analyzed on a picomolar scale without any work-up or pre purification using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) in combination with the ProteinChip System. The technique provides rapid information about the increasing molecular mass of the glycoconjugate formed, thereby allowing termination of the process when the desired incorporation of the ligand onto the carrier protein is achieved. The excess oligosaccharide used at the onset of conjugation can be recovered and used in preparation of a similar neoglycoconjugate. The overall economy of conjugations, which often involve labor-intensive linker-equipped oligosaccharides, can be markedly increased in this way. PMID- 11269401 TI - Structural studies of the antigen III cell wall polysaccharide of Trichosporon domesticum. AB - Cell wall and soluble polysaccharides that reacted with Trichosporon domesticum factor III serum were isolated from the type strain of T. domesticum. The fractions contained O-acetyl groups, which contributed to the serological reactivity. The antigenic structure was characterized by chromatographic and spectroscopic methods. The polysaccharide has an alpha-(1-->3)-D-mannan backbone with hetero-oligosaccharide side chains consisting of a 2-O-substituted beta-D glucuronic acid residue bound to O-2 of the mannose residue, beta-D-xylopyranosyl residues located in the middle of the side chain, and a nonreducing terminal alpha-L-arabinopyranosyl residue bound to 0-4 of xylose. The mannan backbone is O acetylated at O-6 of the mannose residues. PMID- 11269400 TI - Enzymatic syntheses of T antigen-containing glycolipid mimicry using the transglycosylation activity of endo-alpha-N-acetylgalactosaminidase. AB - Thomsen-Friedenreich antigen (T antigen) disaccharide, beta-D-galactose-(1-->3) alpha-N-acetyl-D-galactosamine (beta-D-Gal-(1-->3)-alpha-D-GalNAc), containing glycolipid mimicry was synthesized using the transglycosylation activity of endo alpha-N-acetylgalactosaminidase from Bacillus sp. This enzyme could transfer the disaccharide from a p-nitrophenyl substrate to water-soluble 1-alkanols and other alcohols at a transfer ratio of 70% or more. Although the transfer ratios were lower for water-insoluble than water-soluble alcohols, they were shown to increase by adding sodium cholate to the reaction mixtures. The enzyme also transferred the disaccharide directly from asialofetuin to 1-alkanols. The anomeric bond between the disaccharide and 1-alkanols of the transglycosylation product is in the alpha configuration as determined by sequential digestion of jack bean beta-galactosidase and Acremonium alpha-N-acetylgalactosaminidase. Since the transglycosylation product, beta-D-Gal-(1-->3)-alpha-D-GalNAc-(1-->O) hexyl, efficiently inhibits the binding of anti-T antigen monoclonal antibody to asialofetuin, it has potential as an agent for blocking T antigen-mediated cancer metastasis. PMID- 11269402 TI - Location of the O-methyl groups in the O polysaccharide of Pseudomonas syringae pv. phaseolicola. AB - The O-methylation pattern of the O polysaccharide (OPS) of the lipopolysaccharide of Pseudomonas syringae pv. phaseolicola GSPB 1552 was revealed by methylation (CD3I) analysis, Smith degradation, and NMR spectroscopy. Together with the major O repeats consisting of D-rhamnopyranose (D-Rhap) and D-fucofuranose (D-Fucf), there are minor repeats (approximately 30%) containing 3-O-methyl-D-rhamnose (D acofriose), which is 2-substituted in the interior repeats and occupies the terminal non-reducing end of the OPS. It was suggested that the methylated O repeats are linked to each other nearby the non-reducing end of the OPS and that the 'biological' O repeat of the OPS has the following structure: [molecular structure: see text]. PMID- 11269404 TI - Synthesis of an L-quinovose-containing disaccharide. AB - The synthesis of a versatile L-rhamnose monosaccharide synthon is described. This synthon is used in the synthesis of a disaccharide containing the rare sugar, 6 deoxy-L-glucose, linked to the 3-C-hydroxymethyl group of methyl 2,3-O isopropylidene 3-C-(hydroxymethyl)-beta-D-erythrofuranoside. PMID- 11269403 TI - Syntheses of daidzein-7-yl beta-D-glucopyranosiduronic acid and daidzein-4',7-yl di-beta-D-glucopyranosiduronic acid. AB - Syntheses of the title compounds--commonly known as 'daidzein 7-glucuronide' and 'daidzein 4',7-diglucuronide'--are described. Selective 7-deacetylation of 4',7 di-O-acetyldaidzein is employed. PMID- 11269405 TI - Structure of the O-specific polysaccharide of Hafnia alvei PCM 1196. AB - An acidic O-specific polysaccharide was isolated from Hafnia alvei PCM 1196 lipopolysaccharide and studied by sugar and methylation analyses along with one- and two-dimensional 1H and 13C NMR spectroscopy, including NOESY and HMBC experiments. The following structure of the pentasaccharide repeating unit was established: -->4)-alpha-D-GalpA-(1-->3)-beta-D-GlcpNAc-(1-->2)-beta-D-Galp-(1- >6)-alpha-D-Glcp-(1-->6)-alpha-D-GlcpNAc-(1-->. PMID- 11269407 TI - The structure of the carbohydrate backbone of the core-lipid A region of the lipopolysaccharide from Proteus penneri strain 40: new Proteus strains containing open-chain acetal-linked N-acetylgalactosamine in the core part of the LPS. AB - Analysis of the core part of the LPS from several strains of Proteus revealed that P. penneri strains 2, 11, 19, 107, and P. vulgaris serotypes 04 and 08 have the same structure with a new type of linkage between monosaccharidesan open chain acetal--that was previously determined for P. vulgaris OX2 and P. penneri 17. The LPS from P. penneri strain 40 contains the same structure substituted with one additional monosaccharide: [molecular structure: see text] where (1S) GalaNAc1 is a residue of N-acetyl-D-galactosamine in the open-chain form. It is connected as a cyclic acetal to positions 4 and 6 of the galactosamine residue having a free amino group. All other sugars are in the pyranose form. PMID- 11269406 TI - A disaccharide repeat unit is the major structure in fucoidans from two species of brown algae. AB - The predominant repeating structure of a fraction of the fucoidan from Ascophyllum nodosum prepared by acid hydrolysis and centrifugal partition chromatography (LMWF) was established as: [-->3)-alpha-L-Fuc(2SO3-)-(1-->4)-alpha L-Fuc(2,3diSO3-)-(1]n by NMR spectroscopy and methylation analysis. The proton and carbon NMR spectra of this unit have been assigned and found to correspond with features in the spectra of the whole purified fucan from A. nodosum which account for most of the integrated intensity. The same structure has also been recognised in the fucoidan of Fucus vesiculosus. The fraction LMWF has in vitro anticoagulant activity, indicating that the above structure may be partly responsible for biological activity in the native fucoidan. PMID- 11269408 TI - Eighth International Symposium on Microbial Ecology (ISME-8)--microbial ecology on the doorstep of the 21st century. PMID- 11269409 TI - Bridge to transplantation with a Thoratec left ventricular assist device in a 17 kg child. AB - A Thoratec left ventricular assist device (LVAD) was used to support a 7-year-old 17-kg boy with viral cardiomyopathy for 23 days before heart transplantation. The boy is still living more than 1 year posttransplant, and functional except for some spastic paresis of the left hand, a residual from a stroke during device support. He is the smallest person to be supported with this device. We discuss techniques for using the Thoratec in children. PMID- 11269410 TI - Aerosolized iloprost for severe pulmonary hypertension as a bridge to heart transplantation. AB - Preexisting pulmonary hypertension in pediatric patients is associated with poor outcome after cardiac transplantation because of donor right ventricular dysfunction. To avoid a combined heart-lung transplantation in a 17-year-old patient, we used an intensified pretreatment with intravenous prostacyclin and dobutamine combined with an inhalative therapy with the aerosolized prostacyclin analog Iloprost. With this regimen, the patient was hemodynamically stabilized for the waiting period of 21 days after which an uneventful cardiac transplantation was performed. PMID- 11269411 TI - Eleven-year follow-up after descending thoracic aorta replacement in a small child. AB - Eleven years after the prosthetic replacement of a hypoplastic thoracic aorta in a 3.5-year-old child, there was normal growth of the aortic arch and abdominal aorta without signs of restenosis. This case illustrates that growth of the aorta can be normal after replacement of its thoracic part in a growing child. PMID- 11269412 TI - Origin of the left pulmonary artery from the aorta: embryologic considerations. AB - We observed a case of anomalous origin of the left pulmonary artery from the aorta in which the media of the abnormal vessel and the main pulmonary artery were fused, but without communication. This is the fifth isolated case of repair without the use of cardiopulmonary bypass reported in the literature. This pathology should be included in the aortic arch anomalies as a partial or complete failure of development of the left sixth arch. PMID- 11269413 TI - Management of pseudoaneurysm of the ascending aorta performed under circulatory arrest by port-access. AB - Pseudoaneurysms of the ascending aorta following previous thoracic surgery pose a difficult surgical management problem. In this report, we present a case of a patient with aortic insufficiency and a pseudoaneurysm of the ascending aorta at the site of a previous anastomosis. The particularity of this case is in the atypical use of Port-Access technology (Heartport, Redwood City, CA) to overcome surgical concerns [1]. PMID- 11269414 TI - Stapling of an aortic arch aneurysm. AB - Saccular aneurysms of the aortic arch are usually managed with patch graft aortoplasty or with tube graft replacement. In either case, hypothermic circulatory arrest is necessary. The use of stapling devices has revolutionized pulmonary and gastrointestinal surgery; however, these instruments have rarely been used in aortic surgery except during thromboexclusion procedures. We present a simple and seemingly innovative stapling method that eliminates the need for circulatory arrest. PMID- 11269415 TI - The left gastric artery as an in-situ conduit in coronary artery bypass grafting. AB - Unsuitability of the in-situ right gastroepiploic artery in coronary bypass grafting occurs. Sometimes free-grafting can be performed, although this should not be considered in patients with a diseased ascending aorta. We describe the successful use of the left gastric artery as an alternative in-situ arterial conduit in a patient with a severely atherosclerotic ascending aorta. PMID- 11269416 TI - Giant left atrial intrapericardial aneurysm: noninvasive preoperative imaging. AB - Congenital giant intrapericardial aneurysms of the left atrium are rare. A 17 year-old boy presented with paroxysmal episodes of palpitations, chest pain, and dyspnea. A chest roentgenogram showed an enlarged left cardiac silhouette. Transthoracic echocardiography imaging showed an intrapericardial aneurysm of the left atrium. Cardiac magnetic resonance imaging confirmed the diagnosis and delineated adjacent structures to plan the surgical resection. We have found no previous reports of cases of diagnosis and preoperative assessment based solely on noninvasive imaging. PMID- 11269417 TI - An unusual cause of severe cyanosis in infancy. AB - An infant presented with cyanosis due to a diaphragmatic Morgagni hernia compromising right ventricular diastolic filling and resulting in right-to-left atrial-level shunting as demonstrated by contrast echocardiography. There was complete resolution of cyanosis after repair of the hernia. PMID- 11269418 TI - Surgical repair for double outlet right ventricle and intact ventricular septum. AB - We report the case of a neonate with a complex cardiac anomaly that consisted of a double outlet right ventricle, intact ventricular septum, small left ventricle, and pulmonary stenosis who underwent surgical repair with a successful outcome. We have not previously found a case report of a patient with double outlet right ventricle and intact ventricular septum who is still alive and in good health at an intermediate postoperative follow-up. PMID- 11269420 TI - Hemoptysis secondary to pulmonary pseudoaneurysm 30 years after a gunshot wound. AB - A 49-year-old man presented with intermittent hemoptysis from a traumatic pulmonary artery pseudoaneurysm 30 years following a thoracic gunshot wound. The patient was asymptomatic for 28.5 years, when he began experiencing recurrent hemoptysis, chest pain, and a cough. A left lower lobe mass on chest x-ray film was investigated with contrast-enhanced computed tomography and pulmonary angiogram confirming a 1.5-cm pseudoaneurysm. Intraluminal coil embolization was attempted, but a left lower lobectomy was ultimately necessary to treat persistent hemoptysis. PMID- 11269419 TI - Penetrating thoracic trauma in arrow injuries. AB - Arrow wounds are very rare. We present herein a case of hilar penetrating thoracic trauma caused by an arrow, and a review of the literature, to clarify the management of these cases and their indications for surgery. Depending on the type of arrowhead, the tissue elasticity can narrow the wound track around the shaft of the arrow, sometimes causing a tamponade effect. In the mediastinal or hilar area, an arrow should not be removed before an injury to the major blood vessels or the heart has been ruled out. PMID- 11269421 TI - Pulmonary artery aneurysm in a pregnant woman. AB - We present a patient with a pulmonary artery (PA) aneurysm who has none of the documented causes of PA aneurysm but who is pregnant. We believe that this patient represents a case of primary pregnancy-associated PA aneurysm. PMID- 11269422 TI - Postpartum coronary artery dissection complicated by heparin-induced thrombocytopenia and thrombosis. AB - A 32-year-old woman presented with an acute anterior-lateral myocardial infarction 7 days postpartum. Coronary angiography revealed an occlusive left main coronary artery dissection. After coronary artery bypass surgery the patient's cardiac function improved and stabilized. Thrombocytopenia and a femoral artery thrombosis after 9 days of heparin exposure marked the development of heparin-induced thrombocytopenia and thrombosis that was successfully managed with argatroban, a direct thrombin inhibitor anticoagulant. PMID- 11269423 TI - Innominate artery rupture after transcervical drainage for descending necrotizing mediastinitis. AB - We present a case of innominate artery rupture after descending necrotizing mediastinitis (DNM) on day 36 of cervicomediastinal drainage. The patient recovered after aortosubclavian arterial bypass grafting followed by resection of the eroded artery. Because mechanical pressure caused by drains in addition to the inflammatory process can cause major vessel erosion, prolonged transcervical tube drainage for treating descending necrotizing mediastinitis should be avoided even if the drains applied are soft and thin. PMID- 11269425 TI - Port site metastasis after laparoscopic staging of esophageal carcinoma. AB - The case history of a patient who underwent laparoscopic staging of an esophageal carcinoma is presented. After neoadjuvant chemoradiation therapy a port site metastasis was found at esophagectomy. Possible etiologies, implications for the continued use of minimally invasive staging for esophageal carcinoma, and prevention of port site metastasis are discussed. PMID- 11269424 TI - Postpneumonectomy aortic arch mycotic aneurysm. AB - A 31-year-old woman who had undergone left pneumonectomy for a tuberculosis destroyed left lung 3 years previously presented in respiratory distress after a pregnancy complicated by preeclampsia and aspiration pneumonia. Investigation revealed a large aortic arch aneurysm as well as a filling defect in the descending thoracic aortic lumen. Emergency aortic arch reconstruction was performed for a massive pseudoaneurysm or contained rupture filling the entire postpneumonectomy space. Pathologic and microbiological examination demonstrated Aspergillus fumigatus and active inflammation. PMID- 11269426 TI - Bronchial carcinoid of an accessory tracheal bronchus. AB - We present a rare case of bronchial carcinoid tumor arising in an accessory right tracheal bronchus and involving the associated tracheal lobe in a 48-year-old man, who presented with a history of recurrent respiratory infections and recent onset of hemoptysis. Diagnosis was established on preoperative bronchoscopy and biopsy. The tumor was completely removed by right upper lobectomy with the tracheal bronchus resected flush to its origin from the right lower tracheal wall. Final histology revealed a typical carcinoid tumor. PMID- 11269427 TI - Stenosis of the trachea caused by retrosternal ossification. AB - An abnormal anatomic structure of the thoracic inlet causing stenosis of the trachea is rare. This case report illustrates a man presenting tracheal stenosis caused by retrosternal ossification, and discusses its clinical significance and possible pathogenesis. PMID- 11269428 TI - Lung volume reduction surgery in a ventilated patient with severe pulmonary emphysema. AB - There are a limited number of reports in the literature cocerning lung volume reduction surgery in patients receiving mechanical ventilation. We present a case in which a ventilator-dependent patient with apparent endstage pulmonary emphysema underwent lung volume reduction with a successful outcome. Although the role of this procedure for selected nonventilated patients has been widely discussed its use in ventilated patients is still not clearly defined. We show that lung volume reduction surgery may facilitate ventilatory weaning in such cases and improve functional status. PMID- 11269429 TI - Subatmospheric pressure dressing for saphenous vein donor-site complications. AB - Newer endoscopic techniques have been successful at reducing saphenous vein donor site wound complications, but not entirely eliminating them. Tissue necrosis with superimposed infection is typically treated with antibiotics and surgical debridement. Typically, primary reclosure is not possible and the open leg wound is allowed to slowly granulate with dressing changes until a skin graft can be performed. This report describes an alternative treatment using subatmospheric pressure dressing to promote granulation tissue and wound closure in saphenous vein donor-site wounds. PMID- 11269430 TI - Emergency cardiopulmonary bypass in a bilaterally nephrectomized patient with a history of heparin-induced thrombocytopenia: successful reexposure to heparin. AB - We present a case of emergency coronary artery bypass surgery in a bilaterally nephrectomized patient with a history of heparin-induced thrombocytopenia. The patient was reexposed short term to heparin during cardiopulmonary bypass and did not develop any complications related to heparin-induced thrombocytopenia. Despite intraoperative neutralization of heparin severe bleeding complications occurred, probably resulting from preoperative therapeutic anticoagulation with rhirudin in conjunction with an increased half-life of more than 2 days. PMID- 11269431 TI - Vanishing pulsatile mass on the anterior chest wall. PMID- 11269432 TI - Left atrial reduction and mitral valve surgery: the "functional-anatomic unit" concept. AB - The desired outcome for patients undergoing mitral valve surgery includes both good function of the mitral valve, and preservation and restoration of sinus rhythm. To achieve such an outcome, we evolved the concept of the left atrium and mitral valve as a "functional anatomic unit." In this report, we describe a technique for reduction in left atrial size, isolation of the pulmonary veins, and amputation of the left atrial appendage in combination with mitral valve repair. We performed such a procedure in 4 patients, with rheumatic mitral valve disease and chronic atrial fibrillation, with restoration of good valve function and sinus rhythm at 16 to 20 months after surgery. PMID- 11269433 TI - Pulmonary venous drainage into the left atrial appendage facilitates transplantation of the left lung with difficult exposure. AB - Heterotopic implantation of the pulmonary venous confluence into the left atrial appendage during left lung transplantation is a reasonable alternative technique to reestablish venous drainage when exposure of the native left pulmonary veno atrial connection may be problematic. We used this approach in a 39-year-old woman with chronic bronchiectasis who underwent bilateral sequential lung transplantation through a clam-shell approach. Dense hilar scarring and a small left atrial size made exposure of the native left pulmonary veno-atrial connection difficult. PMID- 11269434 TI - Complex reconstruction of the aortic and mitral annuli: a simplification. AB - Simultaneous reconstruction of the aortic and mitral annuli is a useful but complex procedure. Implantation of a bovine pericardial gusset can be facilitated by ex vivo attachment to the sewing ring of the mitral valve prosthesis. PMID- 11269436 TI - Aortic root remodeling with the "cuff" technique for stentless valve implantation. AB - Aortic root and sinotubular junction dilatation and aneurysm of ascending aorta are considered relative contra-indications to implantation of a stentless valve prosthesis, because the modified aortic geometry leads to aortic incompetence and early failure of the prosthesis. Aortic root reconstruction can be performed according to various techniques. We present a surgical technique in which a tubular graft, replacing an ascending aortic aneurysm, allows sinotubular remodeling and satisfactory implantation of a stentless prosthesis. The native aorta is inserted into the vascular prosthesis at the level of the sinotubular junction which is wrapped in order to prevent commissure spreading. Sizing of the vascular and valve prosthesis is made according to annular diameter. Since October 1999, 6 patients have been operated using this technique with good results. PMID- 11269435 TI - Elephant trunk anastomosis between left carotid and subclavian arteries for aneurysmal distal aortic arch. AB - There is an increased risk of rupture with attempting a distal anastomosis when the distal aortic arch exceeds 5 to 6 cm. To circumvent this problem, we describe performing the anastomosis between the left common carotid and the left subclavian arteries and, at the second-stage operation, interposing a tube graft between the left subclavian artery and the descending aortic tube graft. PMID- 11269437 TI - Coronary revascularization without cardiopulmonary bypass in high-risk patients: a route to the future. AB - Previous reports have demonstrated that reoperative coronary revascularization, advanced age, female sex, and impaired left ventricular dysfunction are independent predictors of operative mortality after coronary artery bypass grafting (CABG). CABG without cardiopulmonary bypass (off-pump CABG) has been proposed as a potential therapeutic alternative in these high-risk patient groups. Despite the substantial learning curve associated with off-pump CABG, early outcomes of off-pump CABG in high-risk patients are better than those associated with the conventional on-pump CABG approach. These results suggest that off-pump CABG is a safe alternative to on-pump CABG in high-risk patients. Randomized prospective studies are needed to validate the results of these initial retrospective reports and to demonstrate the long-term benefits of this approach. PMID- 11269438 TI - As originally published in 1993: Brain protection via cerebral retrograde perfusion during aortic arch aneurysm repair. Updated in 2001. PMID- 11269439 TI - As originally published in 1993: Protection from postischemic spinal cord injury by perfusion cooling of the epidural space. Updated in 2001. PMID- 11269440 TI - Thrombogenicity of the St. Jude medical prosthesis with and without silzone coated sewing cuffs. PMID- 11269441 TI - Lower ministernotomy for the repair of atrial septal defects. PMID- 11269443 TI - Congenitally bicuspid aortic valve and associated aortic medical disease. PMID- 11269442 TI - Temporary tricuspid detachment in closure of ventricular septal defect. PMID- 11269444 TI - Complications on sternal reentry. PMID- 11269445 TI - Postoperative drug therapy and survival after coronary artery bypass grafting. PMID- 11269447 TI - Collaborative practice: a personal journey. PMID- 11269446 TI - Selective graft and coronary sinus perfusion in off-pump CABG: is it necessary? PMID- 11269448 TI - Malnutrition, outcome, and nutritional support: time to revisit the issues. PMID- 11269449 TI - Lowest hematocrit on bypass and adverse outcomes associated with coronary artery bypass grafting. Northern New England Cardiovascular Disease Study Group. AB - BACKGROUND: Cardiac surgery patients' hematocrits frequently fall to low levels during cardiopulmonary bypass. METHODS: We investigated the association between nadir hematocrit and in-hospital mortality and other adverse outcomes in a consecutive series of 6,980 patients undergoing isolated coronary artery bypass graft surgery. The lowest hematocrit during cardiopulmonary bypass was recorded for each patient. Patients were divided into categories based on their lowest hematocrit. Women had a lower hematocrit during bypass than men but both sexes are represented in each category. RESULTS: After adjustment for preoperative differences in patient and disease characteristics, the lowest hematocrit during cardiopulmonary bypass was significantly associated with increased risk of in hospital mortality, intra- or postoperative placement of an intraaortic balloon pump and return to cardiopulmonary bypass after attempted separation. Smaller patients and those with a lower preoperative hematocrit are at higher risk of having a low hematocrit during cardiopulmonary bypass. CONCLUSIONS: Female patients and patients with smaller body surface area may be more hemodiluted than larger patients. Minimizing intraoperative anemia may result in improved outcomes for this subgroup of patients. PMID- 11269450 TI - Coronary artery bypass combined with bilateral carotid endarterectomy. AB - BACKGROUND: Surgical management of patients presenting for coronary artery bypass grafting with significant bilateral carotid artery stenosis has not been well defined. In this study, our preliminary results of coronary artery bypass grafting with concomitant bilateral carotid endarterectomy have been reviewed. METHODS: A retrospective nonrandomized chart review was performed in 33 patients with unstable angina and bilateral carotid artery stenosis, more than 70%, undergoing simultaneous coronary artery bypass grafting and bilateral carotid endarterectomy using an eversion technique. RESULTS: Concomitant coronary artery bypass grafting with bilateral carotid endarterectomy was performed urgently in 24 (73%) and electively in 9 (27%) patients. The average carotid artery cross clamp and total perfusion times were 14.7 +/- 4.9 minutes and 123 +/- 29.2 minutes, respectively. The average length of stay in the cardiopulmonary intensive care unit was 4.2 +/- 14.2 days and total hospital stay was 16.2 +/- 20.5 days. Postoperative in-hospital stay was 14.9 +/- 20.3 days. There were no postoperative strokes. Twenty-one (64%) patients were discharged before the tenth postoperative day. Nonfatal postoperative complications occurred in 27% (9 of 33) of patients. The overall 30-day mortality was 6.1% (2 of 33) and that was unrelated to primary cardiac or cerebrovascular events. CONCLUSIONS: Favorable outcome supports the justification for performing concomitant coronary artery bypass grafting with bilateral carotid endarterectomies in selected patients. PMID- 11269451 TI - Bilateral internal thoracic artery use for dialysis patients: does it increase operative risk? AB - BACKGROUND: The efficacy and risk of using the bilateral internal thoracic artery (BITA) for coronary artery bypass grafting in dialysis patients is virtually unknown. METHODS: Twenty-five patients on dialysis who underwent coronary artery bypass grafting using the BITA were retrospectively studied (BITA group). For comparison purposes, 52 patients on dialysis who underwent coronary artery bypass grafting using the left ITA were selected (LITA group). RESULTS: No wound healing problems occurred in the BITA group. Mean postoperative bleeding volume was 1,427 +/- 808 mL and 800 +/- 508 mL in the BITA and LITA groups, respectively (p = 0.00009). Blood transfusions for the BITA and LITA groups required an average of 6.8 and 6.2 units of packed red blood cells, respectively, with no significant difference. Five patients in the BITA group (20%) showed severe atherosclerotic deterioration of the ascending aorta, precluding clamping. Hospital mortality was 4% (1 of 25 patients) in the BITA group and 7.7% (4 of 52 patients) in the LITA group, with no significant difference (p = 0.49). CONCLUSIONS: In patients on dialysis, especially those with severe atherosclerotic or calcified deterioration of the ascending aorta, coronary artery bypass grafting using BITA grafting (arterial in situ conduits) may offer the easiest and most suitable solution without increased operative risk. PMID- 11269452 TI - Flow wire measurements after complete arterial coronary revascularization with T grafts. AB - BACKGROUND: The T-graft procedure achieves complete arterial revascularization in coronary three-vessel disease. In this technique, all bypass anastomoses are supplied by the left internal mammary artery (IMA). This prospective study explores the question of whether the quantitative flow in such grafts is influenced by the pathology in the native coronary arteries. METHODS: Eighty-two patients with coronary three-vessel disease were studied after complete arterial coronary revascularization with T-grafts. Quantitative flow and coronary flow reserve were measured in the proximal IMA with a Doppler guide wire. Three groups were compared: group 1, all native coronary arteries were stenosed but patent (n = 31); group 2, one occluded native coronary vessel (n = 33); group 3, two or more occluded native coronary arteries (n = 18). RESULTS: Quantitative flow was significantly higher in group 3 than in group 2 at 1 week (93.9 +/- 39.5 vs 75.8 +/- 27.3 mL/min, p < 0.05) and 6 months postoperatively (86.0 +/- 40.1 vs. 69.1 +/- 35.5 mL/min, p < 0.05). Flow in group 2 was significantly (p < 0.05) higher than in group 1 (1 week: 58.0 +/- 28.4 mL/min, 6 months: 55.2 +/- 29.2 mL/min) in both examinations. There were no significant differences in coronary flow reserve between the three groups (1: 2.88 +/- 0.97, 2: 2.84 +/- 0.96, 3: 2.74 +/- 0.94). CONCLUSIONS: After complete arterial revascularization with T-grafts, the quantitative flow in the IMA is influenced by the status of the native coronary arteries. As a result of competitive flow phenomena, blood flow in the bypasses is significantly lower when the coronary arteries are affected only by stenosis. PMID- 11269453 TI - Single aortic cross-clamp technique reduces S-100 release after coronary artery surgery. AB - BACKGROUND: Neurologic impairment after coronary artery bypass grafting is associated with cerebral embolization. An important cause of embolism is aortic manipulation. Constructing both distal and proximal anastomoses during a single period of aortic cross-clamping avoids this source of embolism and may reduce neurologic injury after coronary artery bypass grafting. METHODS: Fifty consecutive patients undergoing coronary artery bypass grafting were prospectively randomized to group 1, in which a single aortic cross-clamping was used to construct distal and proximal anastomoses, or to group 2, in which the proximal anastomoses were each constructed with a partial occluding aortic clamp. Levels of S-100 and troponin-T release were measured preoperatively and postoperatively. RESULTS: Aortic cross-clamp time was significantly longer in group 1, but other preoperative and intraoperative variables were equally represented in both groups. Control group levels of S-100 and troponin-T were similar. Postoperative S-100 levels were significantly higher in group 2 than in group 1 (p < 0.015). No significant difference was found between the groups in postoperative troponin-T levels. CONCLUSIONS: The results of this trial suggest improved cerebral protection is associated with the single aortic cross-clamp technique for coronary artery bypass grafting with no increase in myocardial damage. The single aortic cross-clamp technique is simple and inexpensive. We recommend its wider use. PMID- 11269454 TI - High early patency of saphenous vein graft for coronary artery bypass harvested with surrounding tissue. AB - BACKGROUND: Surgical trauma to the saphenous vein, used as a conduit for coronary artery bypass grafting, affects their occlusion rate. This study evaluates the early patency of saphenous vein grafts harvested with a pedicle of surrounding tissue that protects the vein from spasm and trauma. METHODS: Fifty-two patients underwent coronary artery bypass grafting with saphenous veins harvested with surrounding tissue. Forty-five patients, who received a total of 124 vein grafts and 42 left internal mammary arteries, underwent angiographic follow-up at a mean of 18 months (9 to 24 months). RESULTS: Patency for saphenous vein grafts was 95.4% and for left internal mammary arteries, it was 93.3%. Twenty-nine of 30 (96.7%) vein grafts anastomosed to arteries 2.0 mm or more, 65 of 67 (97%) grafts to 1.5 mm, and 10 of 13 (77%) anastomosed to 1-mm arteries were patent. Nineteen of 22 (86.4%) vein grafts with flow rates 20 mL/min or less, 32 of 34 (94.1%) with flow between 20 and 40 mL/min, and 50 of 51 (98%) with flow more than 40 mL/min were patent. Other registered surgical and clinical factors did not contribute to vessel occlusion. CONCLUSIONS: Early patency rate of saphenous veins harvested with surrounding tissue is very high, even in saphenous vein grafts demonstrating low blood flow. Preservation of graft endothelium using our harvesting technique may be the explanation of this success. PMID- 11269455 TI - Proinflammatory cytokines in cerebrospinal fluid in repair of thoracoabdominal aorta. AB - BACKGROUND: Little is known about alterations of cytokine levels in cerebrospinal fluid (CSF) during thoracoabdominal aortic surgery. We measured perioperative CSF cytokine levels to determine their clinical significances. METHODS: Perioperative serum and CSF levels of cytokine were measured in 15 adult patients undergoing repair of the descending thoracic aorta (n = 4) or thoracoabdominal aorta (n = 11). All patients underwent prosthetic replacement and perioperative CSF drainage. Serum and CSF levels of tumor necrosis factor-alpha, Interleukin- (IL-) 1beta, IL-6, IL-8, IL-10, and IL-12 were measured before operation and at 0, 6, 12, 18, 24, 48, and 72 hours postoperatively using enzyme-linked immunosorbent assays. RESULTS: There were no hospital deaths, but 1 patient suffered paraplegia. Cerebrospinal fluid IL-8 levels peaked at immediately after operation (751.7 +/- 42.1 pg/mL versus preoperative levels, 54.9 +/- 24.6 pg/mL; p < 0.001), and the higher levels persisted for 72 hours. In contrast, serum IL-8 levels did not change and remained lower than CSF levels. The patient with paraplegia had the highest CSF IL-8 levels throughout the study period. Serum and CSF levels of tumor necrosis factor-alpha, IL-1beta, IL-6, and IL-12 did not significantly change. Serum and CSF levels of IL-10 were significantly elevated after operation compared with preoperative levels. In contrast to IL-8, serum IL 10 levels surpassed CSF levels. CONCLUSIONS: Cerebrospinal fluid IL-8 levels are significantly elevated in thoracoabdominal aortic operation, and may be the most sensitive to the inflammatory response in the ischemic spinal cord injury. Persistent elevation of CSF IL-8 levels may be predictive of further development of neurologic deficits, and a reduction of proinflammatory cytokine levels may be a beneficial effect of CSF drainage, but this requires further investigation. PMID- 11269456 TI - Impact of minimally invasive valvular heart surgery: a case-control study. AB - BACKGROUND: The port access (PA) approach for valvular heart surgery is widely used, but few studies evaluating outcomes compared with the sternotomy approach have been performed. METHODS: One hundred nine consecutive patients undergoing PA isolated valve surgery were compared with 88 matched patients who underwent sternotomy-isolated valve surgery before the institution of the PA program. Case matching was performed by age, surgeon, congestive heart failure, position of operated valve, and history of previous surgery. RESULTS: Analysis revealed that PA was associated with similar hospital mortality (p = 0.62), longer bypass times (p < 0.001), shorter length of stay (p = 0.02), fewer transfusions (p = 0.02), and fewer septic complications (p = 0.05). CONCLUSIONS: The PA approach for isolated valvular heart surgery provided patients with significantly improved clinical outcomes in their immediate perioperative course. Further studies are required to measure the impact of the PA approach on the patients' recovery after hospitalization. PMID- 11269457 TI - Fifteen-year clinical experience with the biocor porcine bioprostheses in the mitral position. AB - BACKGROUND: Bioprosthetic valve use represents a crucial improvement in surgical treatment of mitral valve disease. The aim of this study is to determine the long term durability of the Biocor porcine bioprosthetic mitral valve. METHODS: Between 1985 and 1989, a total of 158 Biocor porcine bioprosthetic valves were placed in the mitral position, and long-term results of these patients were investigated retrospectively in 1999. RESULTS: Thirty-day mortality was 4.4% (7 patients). Total follow-up was 1,499 patient-years. Actuarial survival was 83.66% +/- 3% at 5 years, 77.78% +/- 3.36% at 13 years (1.8% patient-year). Multivariate analysis demonstrated younger age, duration of implantation, congestive heart failure, and functional class to be significant predictors of late mortality. Actuarial freedom from valve-related mortality was 98.58% +/- 1% at 15 years (0.13% patient-year). Actuarial freedom from structural valve deterioration was 95.49% +/- 1.8% at 5 years, 70.2% +/- 4.12% at 10 years, and 64.82% +/- 5.34% at 13 years (2.6% patient-year). Actuarial freedom from structural valve deterioration-related reoperation was 98.43% +/- 1.1% at 5 years, 89.15% +/- 2.85% at 10 years, and 76.82% +/- 7.91% at 14 years. Multivariate analysis showed younger age and duration of implantation to be significant predictors of structural valve deterioration and its related reoperation. CONCLUSIONS: By studying a 15-year time period, it is seen that this new generation porcine bioprosthetic valve should be considered an alternative for mechanical valves in selected patients. PMID- 11269458 TI - Modifications of the Cox-Maze III procedure. AB - BACKGROUND: The extended operative time needed for surgery with complicated atrial incisions may preclude application of the Cox-Maze III procedure (CM-III) as a concomitant operation. And after the CM-III, left atrial (LA) contraction has been reported to recover in reduced magnitude compared with right atrial (RA) contraction. METHODS: To decrease operative time, we have modified the CM-III (modification I) by: obliterating the LA appendage instead of excising it; cryoablating the bridge between the LA appendage and margin of the pulmonary vein encircling incision; extending the lateral incision of RA onto the RA appendage without excising it, and extending the incision more inferiorly toward the inferior vena cava; and omitting the T-incision of RA. We compared the clinical results of the conventional CM-III (group 1, n = 18) with those of the modified CM-III group (group 2, n = 23) performed in patients with rheumatic mitral valve (MV) disease. To enlarge the contractile area of the LA, we modified the CM-III to encircle the right and left pulmonary veins separately (modification II), and compared the LA contractilities of the conventional CM-III (group A, n = 15) with those of the second modification (group B, n = 9). RESULTS: Modification I: Mean aortic cross-clamp (ACC) times (135 +/- 29 versus 104 +/- 18 minutes, p < 0.005) and cardiopulmonary bypass (CPB) times (240 +/- 33 versus 185 +/- 42 minutes, p < 0.001) were significantly decreased in group 2 compared with those in group 1. In group 1, sinus rhythm was restored in 16 patients (88.9%). RA contractility was demonstrated in 100% of patients with sinus rhythm (16 of 16) and LA contractility in 75% (12 of 16) in the latest follow-up echocardiography. In group 2, sinus rhythm was restored in 21 patients (91.3%). RA contractility was demonstrated in 100% of patients with sinus rhythm (21 of 22) and LA contractility in 76.2% (16 of 21). Modification II: Mean ACC times were increased in group B compared with group A (133 +/- 32 versus 172 +/- 39 minutes, p = 0.02). The A velocities at LA contraction and the ratio of atrial contraction to peak early diastolic filling velocity (A/E ratio) of the trans-mitral flow were 0.14 +/- 0.20 m/sec and 0.23 +/- 0.11 in group A, and 0.58 +/- 0.33 m/sec and 0.47 +/- 0.19 in group B, respectively, both showing a significant increase in group B compared with group A (p < 0.05). CONCLUSIONS: Our first modification of the CM-III showed comparable sinus conversion rates and incidence of atrial contractility restoration with significantly shorter ACC and CPB times than the conventional CM-III. The second modification of the CM-III significantly increased the LA contractility when compared with the conventional CM-III, although the second modification required a longer ACC time. PMID- 11269459 TI - The role of apolipoprotein E in cognitive decline after cardiac operation. AB - BACKGROUND: Recently, Tardiff and colleagues have suggested that the presence of the apolipoprotein E, epsilon4 allele was associated with increased likelihood of cognitive decline after coronary artery bypass grafting. The objective of the current study was to replicate this earlier work using an increased sample size. The increased sample also enabled an analysis by individual genotype in cognitive decline after coronary artery bypass grafting. METHODS: Apolipoprotein E genotyping was performed on 111 individuals undergoing coronary artery bypass grafting. Each participant underwent a battery of nine neuropsychological tests before operation and 4 to 7 weeks after operation. RESULTS: Cognitive decline, assessed by both continuous Z change scores and two categoric measures of cognitive deficit, was not significantly associated with either individual apolipoprotein E genotypes or categorization by the presence or absence of the epsilon4 allele. The examination of potential moderating factors did not alter this finding. CONCLUSIONS: This study suggests that the epsilon4 allele is not associated with cognitive decline in the weeks after coronary artery bypass grafting. PMID- 11269460 TI - Stimulation of neutrophil activation during coronary artery bypass grafting: comparison of crystalloid and blood cardioplegia. AB - BACKGROUND: During myocardial ischemia, activation of polymorphonuclear neutrophils (PMNs) results in the production of free oxygen radicals, which increase myocardial injury. It has been shown that PMNs also produce nitric oxide. It is not clear whether PMNs become activated as a result of their direct contact with ischemic/reperfused myocardium or if PMN activation and free oxygen radical production are effects of specific stimuli released during coronary artery bypass grafting (CABG). The aim of the current study was to evaluate plasma-mediated neutrophil stimulation and production of superoxide anion (O2) and nitric oxide in patients undergoing CABG, and to verify whether crystalloid and blood cardioplegia can modify such stimulation. METHODS: Coronary sinus, peripheral arterial, and venous plasma samples were collected from 50 patients who underwent CABG and were divided into 2 equal groups which received either crystalloid or blood cardioplegia: directly before myocardial ischemia and aortic cross-clamping; at the beginning of reperfusion after aortic clamp release; and 30 minutes after reperfusion. O2 and nitric oxide production by PMN was evaluated by standard methods. RESULTS: There was a significant (p < 0.05) increase in O2 production by PMN incubated with plasma obtained from the coronary sinus immediately after reperfusion in patients receiving crystalloid cardioplegia compared to blood cardioplegia. No difference was observed in plasma stimulation of nitric oxide production by PMN in the 2 groups of patients at different times during the procedure. CONCLUSIONS: Cardioplegia may affect release of neutrophil oriented stimuli from ischemic myocardium and modify neutrophil activation during coronary artery bypass grafting. PMID- 11269461 TI - Early and intensive continuous hemofiltration for severe renal failure after cardiac surgery. AB - BACKGROUND: The aim of this study was to test whether early and intensive use of continuous venovenous hemofiltration (CVVH) achieved a better than predicted outcome in patients with severe acute renal failure undergoing cardiac operations, and whether a simple and yet accurate model could be developed to predict their outcome before starting CVVH. METHODS: Medical record analysis with collection of demographic, clinical, and outcome information was used. RESULTS: Sixty-five consecutive patients were treated with early and intensive CVVH (mean operation to CVVH time, 2.38 days; pump-controlled ultrafiltration rate, 2 L/h) after coronary artery bypass grafting (56.9%), single valve procedure (16.9%), or combined operations (26.2%). In 32.3% of patients, intraaortic balloon counterpulsation was required and 20% of patients were emergencies. Sustained hypotension despite inotropic and vasopressor support occurred in 40% of patients and prolonged mechanical ventilation in 58.5%. Using an outcome prediction score specific for acute renal failure, the predicted risk of death was 66%. Actual mortality was 40% (p = 0.003). Using multivariate logistic regression analysis and neural network analysis, patient outcome could be predicted with good levels of accuracy (receiver operating characteristic 0.89 and 0.9, respectively). CONCLUSIONS: Early and aggressive CVVH is associated with better than predicted survival in severe acute renal failure after cardiac operations. Using readily available clinical data, the outcome of such patients can be predicted before the implementation of CVVH. PMID- 11269462 TI - Comparison of epsilon aminocaproic acid and low-dose aprotinin in cardiopulmonary bypass: efficiency, safety and cost. AB - BACKGROUND: In this study we compared the clinical efficiency, safety, and economic benefit of low-dose aprotinin with epsilon aminocaproic acid (EACA) in reducing bleeding after cardiopulmonary bypass operation. METHODS: In a double blind, randomized study, 100 patients received low-dose aprotinin (2 x 10(6) kallikrein inhibitor units) or EACA (20 g). The surgical procedure was single- or double-valve replacement with or without coronary artery bypass grafts. RESULTS: Mediastinal chest drainage and transfusion requirements with both therapies were similar. There were no urgent reoperations to secure hemostasis in either group. Similar levels of D-dimer with both therapies indicate a similar inhibition of fibrinolysis. Release of troponin I was less in the low-dose aprotinin group 1 and 4 hours after bypass, although electrocardiographic measurements did not reflect this difference. Levels of S-100beta and neuron-specific enolase were similar with both therapies, confirming that there was no difference in the occurrence of any adverse neurologic events in either group. CONCLUSIONS: Low dose aprotinin and EACA showed similar effects on the reduction of intraoperative and postoperative bleeding. The lower cost of EACA with no change in safety outcome suggests it is the preferred treatment. PMID- 11269463 TI - Factors affecting functional outcome after autologous skeletal myoblast transplantation. AB - BACKGROUND: This study assessed the extent to which the initial degree of functional impairment and the number of injected cells may influence the functional improvement provided by autologous skeletal myoblast transplantation into infarcted myocardium. METHODS: One week after left coronary artery ligation, 44 rats received into the infarcted scar, autologous skeletal myoblasts expanded in vitro for 7 days (mean, 3.5 x 10(6), n = 21), or culture medium alone (controls, n = 23). Left ventricular function was assessed by two-dimensional echocardiography. RESULTS: When transplanted hearts were stratified according to their baseline ejection fraction, a significant improvement occurred at 2 months in the less than 25% (from 21.4% to 37%), 25% to 35% (from 29% to 43.8%), and in the 35% to 40% (from 37.2% to 41.7%) groups, compared to controls (p = 0.048, 0.0057, and 0.034, respectively), but not in the more than 40% stratum. A significant linear relationship was found between the improvement in ejection fraction and the number of injected myoblasts, both at 1 and 2 months after transplantation (p < 0.0001). CONCLUSIONS: Autologous myoblast transplantation is functionally effective over a wide range of postinfarct ejection fractions, including in the sickest hearts provided that they are injected with a sufficiently high number of cells. PMID- 11269464 TI - Combination of preconditioning and delayed flap elevation: evidence for improved perfusion and oxygenation of the latissimus dorsi muscle for cardiomyoplasty. AB - BACKGROUND: Atrophy and fibrosis of the distal part of the latissimus dorsi muscle (LDM) wrap is a recognized complication of cardiomyoplasty that has been attributed to ischemia. Failure of the muscle wrap contributes to the late attrition seen in clinical cardiomyoplasty. In this study we examined the role of two-staged mobilization and of preconditioning by electrical stimulation on the regional perfusion and oxygenation of the LDM. METHODS: In a rabbit model (n = 36) the LDM was preconditioned as follows: group A muscles received preconditioning in situ; group B muscles were partially mobilized by dividing the intercostal perforators and then preconditioned; and group C muscles were completely mobilized and wrapped around a silicone-rubber mandrel before conditioning. Controls received no conditioning. The preconditioning regimen consisted of 2 weeks of continuous stimulation at 2.5 Hz. At completion of preconditioning the muscles were fully mobilized and mounted on a muscle-testing apparatus. Purpose-built microelectrodes measured regional PO2 and perfusion using a diffusible gas tracer technique. Muscles were weighed and processed for fiber typing and capillary counting. RESULTS: All preconditioned muscles demonstrated fiber transformation, with increased fatigue resistance. Perfusion of preconditioned muscles both at rest and during contraction was higher than control in the proximal part of the muscle. Distal regions of group B muscles had higher perfusion and capillary density than any other group (p < 0.05). Distal regions of group C had the lowest perfusion and capillary density, and showed muscle atrophy and histologic evidence of necrosis. During fatigue testing there was a decrease in the PO2 in the distal regions of the control and group C muscles (p < 0.05), whereas it was maintained at resting levels in both group A and B muscles. CONCLUSIONS: Conditioning in situ improves perfusion of the distal LDM and prevents a fall in tissue PO2 during contraction. Two-stage mobilization further improves distal perfusion and capillary density. In contrast, shortterm elevation followed by conditioning produces impaired distal perfusion, decrease in PO2, and fiber necrosis in the distal muscle. The present study suggests that partial mobilization of the LDM performed at the same time as placement of electrodes for preconditioning may prepare the LDM better for the demands of cardiomyoplasty. PMID- 11269465 TI - Effect of extracorporeal membrane oxygenation on left ventricular function of swine. AB - BACKGROUND: Previous clinical and experimental investigations have produced inconsistent data describing the effects of veno-arterial extracorporeal membrane oxygenation (VA ECMO) on intrinsic left ventricular (LV) function. We report an animal model that allows investigation of the effects of VA ECMO on the mechanics of the LV using two load-insensitive indices: end-systolic pressure-minor axis dimension relationship (ESPDR) and preload recruitable dimensional stroke work (PRDSW). METHODS: Eight piglets (5 to 11 kg) were anesthetized, instrumented, and placed on VA ECMO. Throughout the experiment, systemic and left atrial partial pressure of oxygen were maintained between 100 to 200 mm Hg. At ECMO flow rate of 50% of baseline cardiac output, data were collected prior to ECMO, at 4 and 6 hours during ECMO, and after weaning from ECMO. Data measured or calculated for each time point included heart rate, LV pressures and minor axis dimensions at different pre-loads, first derivative of LV pressure with respect to time, velocity of circumferential fiber length shortening (VCF), LV shortening fraction (LVSF), ESPDR, and PRDSW. RESULTS: A significant (p < 0.05) decrease in LVSF and VCF was seen at 4 and 6 hours during ECMO when compared to baseline, but the ESPDR and PRDSW did not change during ECMO. CONCLUSIONS: VA ECMO alone changes some of the load-dependent parameters of contractility, but intrinsic function of the heart is not significantly affected as measured by load-insensitive indices of LV performance. PMID- 11269466 TI - Left ventricular dysfunction during extracorporeal membrane oxygenation in a hypoxemic swine model. AB - BACKGROUND: Perfusion of the coronary circulation with hypoxemic blood from the left ventricle has been postulated to cause myocardial dysfunction during venoarterial extracorporeal membrane oxygenation for respiratory support. METHODS: We investigated this hypothesis in 10 anesthetized open-chest piglets (7 to 9 kg) undergoing venoarterial extracorporeal membrane oxygenation after placement of minor-axis sonomicrometry crystals and left ventricular micromanometer. The left atrial partial pressure of oxygen was made hypoxemic (25 to 40 mm Hg) after initiation of extracorporeal membrane oxygenation by ventilation with a hypoxic gas mixture. Left ventricular contractile function, including peak LV pressure, shortening fraction, maximum rate of increase of left ventricular pressure, velocity of circumferential fiber shortening, end-systolic pressure-minor axis dimension relationship, and preload recruitable dimensional stroke work, was measured or calculated on extracorporeal membrane oxygenation before (baseline) and at 4 and 6 hours after rendering the left atrial blood hypoxemic. RESULTS: Left ventricular shortening fraction and velocity of circumferential fiber shortening were significantly lower (p < 0.05) at 4 and 6 hours when compared with baseline. The slope of the end-systolic pressure-minor axis dimension relationship decreased but was not significantly different at 4 and 6 hours when compared with baseline owing to poor linear correlation (r = 0.30 to 0.93). The preload recruitable dimensional stroke work was more linear (r = 0.87 to 0.99), and the slope was significantly lower (p < 0.01) at 4 and 6 hours when compared with baseline. CONCLUSIONS: Hypoxemic cardiac output from the left ventricle during venoarterial extracorporeal membrane oxygenation is associated with depression of left ventricular systolic function in this animal model. Current use of venoarterial extracorporeal membrane oxygenation for respiratory support may not provide adequate oxygen supply to the myocardium. PMID- 11269467 TI - Proper timing of blood cardioplegia in infant lambs: superiority of a multiple dose regimen. AB - BACKGROUND: In the pediatric and infant age groups, it is unclear whether repeated infusions of blood cardioplegia solution during ischemic arrest are beneficial or detrimental when compared with a single-dose regimen. METHODS: Twenty lambs (aged 6 to 7 weeks) were placed on cardiopulmonary bypass. A miniature glass-tip electrode measured myocardial pH and hydrogen ion concentration, [H+], in the anterior wall. The aorta was clamped for 2 hours. Group S (n = 10) received a single dose of blood cardioplegia solution. Group M (n = 10) received multiple doses of blood cardioplegia solution at 20-minute intervals. RESULTS: The amount of [H+] generated during the cross-clamp period was greater in group S than in group M (39.2 +/- 10.1 nmol/L versus 0.4 +/- 1.4 nmol/L, p < 0.008). The percent increase in the time constant, tau, an index of diastolic relaxation, was more prolonged after cardiopulmonary bypass in group S when compared with group M (51.4% +/- 2.8% versus 6.4% +/- 3.0%, p < 0.0001). Similarly, the percent decrease in end systolic elastance, a measure of systolic contractility, was greater in group S after cardiopulmonary bypass when compared with group M (29.5% +/- 1.4% versus 7.3% +/- 1.3%, p < 0.0001). CONCLUSIONS: In this infant lamb model, multiple doses of blood cardioplegia solution provided superior metabolic preservation and hemodynamic support after 2 hours of aortic clamping when compared with a single-dose regimen. PMID- 11269468 TI - Time course of cranial ultrasound abnormalities after arterial switch operation in neonates. AB - BACKGROUND: The object of this study was to investigate the time course and fate of abnormal findings in cranial ultrasound after performing an arterial switch operation in neonates with transposition of the great arteries, and to analyze the relationship to cerebral cell damage. METHODS: Cranial ultrasound was performed prospectively in 35 neonates with transposition of the great arteries before the operation as well as 4 hours, 1, 2, and 3 days, and 1 and 2 weeks postoperatively. Blood levels of neuron-specific enolase, a marker of cerebral cell damage, were determined before, during, and 4 and 24 hours postoperatively. RESULTS: In 17 of 35 neonates (49%), early postoperative cranial ultrasound revealed abnormalities indistinguishable from intraventricular hemorrhage. In 11 neonates findings were transient and were normalized 2 weeks postoperatively, whereas in the remaining 6 neonates there was evidence of resolving hemorrhage. In all neonates there was a rise in neuron-specific enolase blood concentrations during and 4 hours after extracorporal circulation without correlation to sonographic findings. CONCLUSIONS: Enhanced echogenicity of the choroid plexus or dilatation of the cerebral ventricular system is a frequent early postoperative finding that may be caused by transient plexus edema rather than intraventricular hemorrhage and is not related to cerebral cell damage. PMID- 11269469 TI - Neurodevelopmental outcome related to cerebral risk factors in children after neonatal arterial switch operation. AB - BACKGROUND: Neurodevelopmental outcome after neonatal arterial switch operation for complete transposition of the great arteries is an important topic needing prospective assessment. METHODS: A group of 33 unselected children (3.0 to 4.6 years) operated on as neonates with combined deep hypothermic circulatory arrest and low flow cardiopulmonary bypass and a control group of 32 age-matched healthy children (3.0 to 4.8 years) underwent evaluation of socioeconomic and clinical neurological status and a standardized test comprising all areas of child development. Results of patients were related to those of the control group, to population norms, and to preoperative, perioperative, and postoperative cerebral risk factors. RESULTS: Clinical neurological status was normal in 26 patients (78.8%) and reduced in 7 (21.2%). Complete developmental score and the subscores for motor function, visual perception, learning and memory, cognitive function, language, and socioemotional functions were not different compared to population norms. Compared to the patients, the children of the control group scored higher on tests of complete development, cognition, and language, but also on socioeconomic status. Complete developmental score and the scores for motor, cognitive, and language functions were weakly inversely related to the duration of circulatory arrest, but not to the duration of bypass. Cerebral risk factors such as serum levels of the neuron-specific enolase, perinatal acidosis, perinatal asphyxia, peri- and postoperative cardiocirculatory insufficiency, or clinical seizures were not correlated to the test results. CONCLUSIONS: Neonatal arterial switch operation with combined circulatory arrest and low flow bypass is associated with neurological impairment, but not with reduced development as assessed by formal testing of motor, cognitive, language, and behavioral functions. Perioperative serum level of the neuron-specific enolase is not a valid marker for later developmental impairment. PMID- 11269470 TI - In vivo flow dynamics of the total cavopulmonary connection from three dimensional multislice magnetic resonance imaging. AB - BACKGROUND: The total cavopulmonary connection (TCPC) design continues to be refined on the basis of flow analysis at the connection site. These refinements are of importance for myocardial energy conservation in the univentricular supported circulation. In vivo magnetic resonance phase contrast imaging provides semiquantitative flow visualization information. The purpose of this study was to understand the in vivo TCPC flow characteristics obtained by magnetic resonance phase contrast imaging and compare the results with our previous in vitro TCPC flow experiments in an effort to further refine TCPC surgical design. METHODS: Twelve patients with TCPC underwent sedated three-dimensional, multislice magnetic resonance phase contrast imaging. Seven patients had intraatrial lateral tunnel TCPC and 5 had extracardiac TCPC. RESULTS: In all patients in both groups a disordered flow pattern was observed in the inferior caval portion of the TCPC. Flow at the TCPC site appeared to be determined by connection geometry, being streamlined at the superior vena cava-pulmonary junction when the superior vena cava was offset and flared toward the left pulmonary artery. Without caval offset, intense swirling and dominance of superior vena caval flow was observed. In TCPC with bilateral superior vena cavae, the flow patterns observed included secondary vortices, a central stagnation point, and influx of the superior vena cava flow into the inferior caval conduit. A comparative analysis of in vivo flow and our previous in vitro flow data from glass model prototypes of TCPC demonstrated significant similarities in flow disturbances. Three-dimensional magnetic resonance phase contrast imaging in multiple coronal planes enabled a comprehensive semiquantitative flow analysis. The data are presented in traditional instantaneous images and in animated format for interactive display of the flow dynamics. CONCLUSIONS: Flow in the inferior caval portion of the TCPC is disordered, and the TCPC geometry determines flow characteristics. PMID- 11269471 TI - Development of pulmonary arteries after central aortopulmonary shunt in newborns. AB - BACKGROUND: Central shunt (CS) is frequently used to treat diminished pulmonary blood flow in newborns. We analyzed the impact of CS on the growth of the pulmonary arteries (PAs). METHODS: Twenty-two consecutive newborns underwent a CS procedure. In 15 newborns the preoperative angiograms and angiograms taken before undergoing anatomic or hemodynamic correction procedures were analyzed. The patients were divided retrospectively into two groups by the size of the PA in the preoperative angiogram: group I, patients with PAs more than 4 mm (n = 10), group II, PAs 4 mm or less (n = 5). To compare the development of the PAs in the groups, the Nakata index, McGoon ratio, and lower lobe indices were calculated from angiograms. RESULTS: The indices were significantly higher in group I before CS, but no differences was found between the groups before anatomic or hemodynamic correction. The postoperative Nakata indices and the McGoon ratios in the groups were higher when compared with preoperative values (group I, p = 0.037 and p = 0.013; group II, p = 0.043 and p = 0.043, respectively). The significant increase of the lower lobe indices only in group II (p = 0.043) suggests faster growth of the PA in this group. CONCLUSIONS: Optimal diameters of the CS promote growth of the PAs, which was confirmed by the increased Nakata and McGoon indices. The benefit in smaller PAs is greater. PMID- 11269472 TI - Long-term results of relief of subaortic stenosis in univentricular atrioventricular connection with discordant ventriculoarterial connections. AB - BACKGROUND: We set out to examine the long-term results of relief of subaortic stenosis by enlargement of ventricular septal defect in patients with univentricular atrioventricular connection to a dominant left ventricle and discordant ventriculoarterial connections. METHODS: Twenty-four patients underwent enlargement of ventricular septal defect between 1985 and 1998 at a median age of 3.2 years (range, 3 weeks to 14 years). Ten patients were younger than 1 year of age. Eighteen had undergone previous banding of the pulmonary trunk, 9 of whom also required repair of coarctation of the aorta. The median subaortic gradient before enlargement was 46 mm Hg. Twenty-three patients had a patch to enlarge the rudimentary right ventricle. RESULTS: Five patients (21%) died in the early postoperative period. The overall survival at 1 and 3 years was 73%, and at 5 and 10 years was 68% and 60%, respectively. Complete heart block requiring insertion of a pacemaker occurred in 2 patients (8%). A Fontan operation was performed in 10 patients, 5 underwent a bidirectional Glenn procedure, and 2 required cardiac transplantation. Follow-up was complete in all survivors at a median time of 6.7 years (range, 8 months to 13 years). From the earlier part of the series, 3 patients experienced aortic insufficiency and 2 had recurrent obstruction. Factors adversely affecting survival were age younger than 1 year at operation and presence of obstruction within the aortic arch. CONCLUSIONS: Our experience shows that, in patients with univentricular atrioventricular connection to a dominant left ventricle and subaortic stenosis, enlargement of the ventricular septal defect provides satisfactory relief of obstruction except in those younger than 1 year of age, and those who have associated obstruction in the aortic arch. PMID- 11269473 TI - Right ventricular outflow tract reconstruction with an allograft conduit. AB - BACKGROUND: Allograft conduits are used for reconstruction of the right ventricular outflow tract in patients with congenital heart disease and in the pulmonary autograft procedure. A retrospective evaluation of our experience with the use of allograft conduits for reconstruction of the right ventricular outflow tract was conducted. METHODS: Between August 1986 and March 1999, 316 allografts (246 pulmonary, 70 aortic) were implanted in 297 patients for reconstruction of the right ventricular outflow tract. Main diagnostic groups were aortic valve pathology (n = 112, 35%), tetralogy of Fallot (n = 71, 22%), and pulmonary atresia with ventricular septal defect (n = 46, 14%). Kaplan-Meier analyses were done for survival, valve-related reoperation, and valve-related events. In addition, Cox regression analysis was used for evaluation of potential risk factors. RESULTS: Mean age at operation was 18 years (range, 7 days to 61 years). Mean follow-up was 4 years (range, 2 days to 12 years). Twelve patients (4%) died within 30 days after operation. Patient survival was 90% (95% confidence interval [CI], 86% to 94%) at 5 years and 88% (95% CI, 83% to 94%) at 8 years. Twenty-four reoperations were required for allograft dysfunction in 23 patients; 21 allografts were replaced. Freedom from valve-related reoperation was 91% (95% CI, 86% to 95) at 5 years and 87% (95% CI, 81% to 93%) at 8 years. Twenty-nine valve related events were reported (2 deaths, 24 reoperations, 2 balloon dilatations, and 1 endocarditis). Freedom from valve-related events was 90% (95% CI, 85% to 94%) at 5 years after implantation, and 84% (95% CI, 77% to 91%) at 8 years. Risk factors for accelerated allograft failure were extra-anatomic position of the allograft (p = 0.03; hazard ratio, 9.7) and the use of an aortic allograft (p = 0.02; hazard ratio, 2.4). CONCLUSIONS: Right ventricular outflow tract reconstruction with an allograft conduit has good medium-term results, although progression of allograft degeneration is noted. Aortic allografts should preferably not be used for reconstruction of the right ventricular outflow tract. PMID- 11269474 TI - Renewal of the Fontan circulation with concomitant surgical intervention for atrial arrhythmia. AB - BACKGROUND: Atrial arrhythmia remains one of the major complications in the longer term after the Fontan procedure. METHODS: Conversion to total cavopulmonary connection was carried out concomitantly with surgical intervention for atrial arrhythmia in 4 patients undergoing the Fontan procedure by atriopulmonary connection and having continual atrial fibrillation or flutter in the longer term after the initial procedure. RESULTS: The surgical intervention restored sinus rhythm. Transient atrial fibrillation occasionally occurred after the reoperation in 1 patient in whom duration of preoperative arrhythmic period had been 6 years, and defibrillation was needed twice. In the other 3 patients, no episodes of paroxysmal arrhythmia have been noted. Subsequent to renewal of the Fontan circulation, cardiac index increased, with systemic venous pressure decreasing. All 4 patients are currently doing well with their functional status of New York Heart Association functional class I. CONCLUSIONS: Combination of conversion to total cavopulmonary connection and concomitant surgical intervention for atrial arrhythmia is effective, when used appropriately and in a timely manner in patients with atrial arrhythmia in the longer term after the initial Fontan procedure by atriopulmonary connection. PMID- 11269475 TI - Individualized heparin and protamine management in infants and children undergoing cardiac operations. AB - BACKGROUND: Measurements of activated coagulation time do not correlate with plasma concentration of heparin. This study investigated the effects of a patient specific method to manage anticoagulation and its reversal in pediatric patients undergoing cardiopulmonary bypass. METHODS: Infants and children were randomly assigned to receive either a standard dose of heparin (300 IU/kg; group C, n = 13) or an individualized dose, calculated by an in vitro heparin dose-response test (group HC, n = 13). Protamine dose was based on a 1 mg/l mg ratio of total administered heparin for patients in group C and of the residual heparin concentration in group HC. RESULTS: Administered heparin was significantly higher and total protamine dose was significantly reduced in the HC group (both p < or = 0.001). There was less thrombin generation (p = 0.02) and fibrinolysis (p = 0.05) in group HC. Blood loss and requirement for transfusion of blood and fresh frozen plasma were also lower in group HC (all p < or =0.05). CONCLUSIONS: An individualized management of anticoagulation and its reversal results in less activation of the coagulation cascade, less fibrinolysis, and reduced blood loss and need for transfusions. Further studies are warranted to better define the clinical impact of these findings. PMID- 11269476 TI - Nutritional status of patients undergoing lung cancer operations. AB - BACKGROUND: Patients referred for lung cancer operations were reported to be nutritionally depleted. This may be relevant in determining patient outcome after surgical procedures. A study was undertaken to measure a range of nutritional variables including dietary intake of patients referred to a regional cardiothoracic center for curative lung cancer operations. METHODS: Anthropometric measurements, grip strength, fat-free mass (FFM), serum protein concentrations, lymphocyte count, creatinine-height index, subjective global assessment, and data on daily intakes of energy, protein, and vitamin C were collected prospectively. Anthropometric indices were also measured in a group of control patients with mild chronic obstructive pulmonary disease. RESULTS: Sixty patients and 22 control patients were recruited. Weight, skin-fold thickness, and grip strength were not significantly different between patients and control patients, and both groups were similar to the general population. However, 8 patients (13.3%) had a body mass index (BMI) less than 20, and 14 patients (24.1%) had a fat-free mass index less than 15. Serum albumin and transferrin concentrations and lymphocyte count were very rarely depressed but prealbumin and retinol-binding protein levels were below normal in 11.9% and 8.3% of patients, respectively. Thirty percent of patients reported low energy intake, 13% reported a low protein intake, and 61.7% had reduced vitamin C intake. CONCLUSIONS: Severe nutritional depletion was uncommon in patients referred for operations for lung cancer and its frequency may have been overestimated in some previous reports. A low intake of vitamin C was common in our patients but its clinical significance is unclear. PMID- 11269477 TI - The influence of nutritional status on complications after operations for lung cancer. AB - BACKGROUND: Nutritional status is known to play an important role in determining outcome after many types of operations but its importance relative to nonnutritional indices in patients undergoing an operation for lung cancer is unclear. METHODS: Detailed nutritional and nonnutritional assessment of 52 patients undergoing surgical resection of lung cancer was performed. The frequency of postoperative complications and length of intercostal drainage time were recorded, and the relation between preoperative indices and postoperative outcome was assessed. RESULTS: Patients who died or needed reventilation had poorer nutritional status, worse lung function, and lower maximum expiratory pressures than those who did not. Using multiple logistic regression, the best model (R2 = 0.39) to predict death combined operation type, preoperative carbon monoxide transfer factor (% predicted), and maximum expiratory pressure (% predicted). Operation type and the fat-free mass index (FFMI) alone were only slightly less informative (R2 = 0.35). For reventilation the best model (R2 = 0.80) combined operation type, body mass index (BMI), and maximum expiratory pressure (% predicted). Intercostal drainage time after lobectomy was significantly related only to preoperative lymphocyte count (p = 0.004) and subjective global assessment score (p = 0.02). CONCLUSIONS: Impaired nutrition is an important predictor of death and the need for reventilation after an operation for lung cancer, and the selection of patients for lung resection might be improved by measuring simple nutritional indices such as BMI and the FFMI. PMID- 11269478 TI - Autocrine motility factor receptor expression in patients with stage I non-small cell lung cancer. AB - BACKGROUND: Expression of autocrine motility factor receptor (AMFR) associates with increased cell migration and poor survival in certain types of human cancers. We assessed the possible correlation between AMFR, clinicopathologic features, and survival in stage I non-small cell lung cancer (NSCLC). METHODS: AMFR expression was analyzed immunohistochemically, using a monoclonal antibody (3F3A) in tumor specimens from 97 patients with curative resection. Vascular endothelial growth factor (VEGF) expression was also examined after accounting for AMFR expression. RESULTS: Out of 97 tumors, 38 (39.2%) were positively stained with AMFR. The AMFR expression was significantly associated with histologic type of tumor, mainly in adenocarcinoma. Overall survival of patients with AMFR-positive tumors was significantly worse than that of AMFR-negative tumors (p = 0.0050). The AMFR expression appears to be associated with VEGF expression. Patients who were AMFR positive and had high VEGF expression had a worse prognosis compared with the AMFR-negative and low VEGF-expression group (p < 0.0001). Multivariate analysis revealed an independent prognostic impact of AMFR on survival (p = 0.0039). CONCLUSIONS: These results indicate that evaluation of AMFR expression may provide useful guidance in follow-up of patients with NSCLC. PMID- 11269479 TI - E-cadherin expression associated with differentiation and prognosis in patients with non-small cell lung cancer. AB - BACKGROUND: E-Cadherin plays a major role in maintaining the intercellular junctions in epithelial tissues. The reduction of E-cadherin expression in cancer cells may be associated with tumor differentiation, metastasis, and a poor prognosis. METHODS: Immunohistochemistry for E-cadherin expression was performed on 109 tumors from patients with non-small cell lung cancer who underwent operations. RESULTS: With respect to membranous immunostaining, 57 carcinomas were E-cadherin-positive, 39 carcinomas E-cadherin-reduced, and 13 carcinomas E cadherin-negative. The percentage of poorly differentiated tumors in the impaired E-cadherin expression group was significantly higher than that in the E-cadherin positive group (p = 0.005). Furthermore, the frequency of lymph node metastases in tumors with impaired E-cadherin expression was significantly higher than that in the E-cadherin-positive tumors (p = 0.011). A Cox regression analysis revealed that E-cadherin expression was a significant factor in the prediction of survival for patients with non-small cell lung cancer (p = 0.002). CONCLUSIONS: E-Cadherin expression was associated with tumor differentiation, lymph node metastasis, and prognosis in patients with non-small cell lung cancer. PMID- 11269480 TI - Is segmentectomy with lymph node assessment an alternative to lobectomy for non small cell lung cancer of 2 cm or smaller? AB - BACKGROUND: Lesser resection than the standard lobectomy for small-sized cT1N0M0 non-small cell lung cancers continues to be debated. METHODS: We reviewed specimens of 139 patients after lobectomy for cT1N0M0 cancer of 2 cm or less. In addition, we prospectively enrolled 70 patients able to tolerate a lobectomy, in a trial of lesser resection for these lesions. The limited procedure consisted of segmentectomy in which the resection line was delivered beyond the burdened segment, plus exploration of lymph nodes by frozen sectioning. This procedure was modified if the result was positive; this modified procedure was called extended segmentectomy. RESULTS: The nodal status after lobectomy was pN0, 107 patients; pN1, 12 patients; and pN2, 20 patients. Of the pN1 patients, 2 had only intralobar nodal involvement within the same segment of the main tumor. In the remaining 30 patients with nodal involvement, we ascertained the nodal involvement during the operation. Regarding intrapulmonary metastasis, 1 of 8 patients having this metastasis had the lesion at the segment where the main tumor was not located and had N2 disease, which was detected intraoperatively. If extended segmentectomy had been performed instead of lobectomy, the lesion could have been removed completely. The 5-year survival of patients with cT1N0M0 cancer of 2 cm or less was 87.3% after extended segmentectomy. There were no local recurrences and three noncancer-related deaths. Among patients with pT1N0M0 cancer of 2 cm or less, the 5-year survival was 87.1% in the extended segmentectomy group and 87.7% in the lobectomy group (p = 0.8008). CONCLUSIONS: Extended segmentectomy should be considered as an alternative for patients with cT1N0M0 non-small cell lung cancer of 2 cm or smaller. PMID- 11269481 TI - Surgical resection of non-small cell carcinoma after treatment for small cell carcinoma. AB - BACKGROUND: Development of non-small cell lung carcinoma (NSCLC) in patients previously treated for small cell carcinoma (SCLC/NSCLC) is well described; however, little is known about clinical outcome. METHODS: A single-institution 20 year review was performed. Patient characteristics and survival for SCLC/ NSCLC patients were compared with those for control patients matched for stage, resection, and previous malignancy. RESULTS: One thousand four hundred four patients with small cell carcinoma were identified, and 29 underwent therapy for metachronous NSCLC: 11 of 29 patients underwent surgical resection, 10 of these 11 (90%) were stage I. Compared with surgically treated stage I NSCLC patients, SCLC/NSCLC patients were more likely to have squamous histology (70% versus 35%, p = 0.026); and subanatomic resection (90% versus 17.4%, p < 0.0005). The SCLC/NSCLC patients had significantly poorer survival when compared with stage I NSCLC patients undergoing any resection (24.53 versus 74.43 months, p = 0.003) and stage I NSCLC patients receiving wedge resection (24.53 versus 58.39 months, p = 0.006). Survival was similar to NSCLC patients with a history of previous treated extrathoracic solid malignancy. CONCLUSIONS: Surgical resection for SCLC/NSCLC patients is feasible, but poorer prognosis is noted when compared with stage-matched control patients. Surgical candidates should be carefully chosen, and alternative local control modalities considered. PMID- 11269482 TI - Postoperative fluorescence bronchoscopic surveillance in non-small cell lung cancer patients. AB - BACKGROUND: Second lung primaries occur at a rate of 1% to 3% per patient-year after complete resections for non-small cell lung carcinoma (NSCLC). Fluorescence bronchoscopy appears to be a sensitive tool for surveillance of the tracheobronchial tree for early neoplasias. METHODS: Patients who were disease free after complete resection of a NSCLC were entered into a fluorescence bronchoscopy surveillance program. All suspicious lesions were biopsied along with two areas of normal mucosa to serve as negative controls. RESULTS: A total of 73 fluorescence bronchoscopies were performed after conventional bronchoscopy in 51 patients at a median of 13 months postresection. The majority (46 of 51) of patients had stage I or II NSCLC, whereas 10% (5 of 51) had stage IIIA. Three intraepithelial neoplasias and one invasive carcinoma were identified in 3 of 51 patients (6%), all current or former smokers. Of the four lesions identified, three were in the 20 patients with prior squamous cell carcinomas. No intraepithelial neoplasias were identified by white-light bronchoscopy, whereas two of three were detected by fluorescence examination. The one invasive cancer detected was apparent on both white-light and fluorescence bronchoscopic examinations. CONCLUSIONS: Surveillance with fluorescence bronchoscopy identified lesions in 6% of postoperative NSCLC patients thought to be disease-free. Patients with prior squamous cell carcinomas appear to be a population that may warrant future prospective study of postoperative fluorescence bronchoscopic surveillance. PMID- 11269483 TI - Early results of a prospective study of limited resection for bronchioloalveolar adenocarcinoma of the lung. AB - BACKGROUND: We reported that bronchioloalveolar adenocarcinoma (BAC) without active fibroblastic proliferation of the lung had no lymph node and pulmonary metastasis and had a favorable prognosis. However, there has been no prospective trial regarding limited pulmonary resection for this type of BAC. The purpose of this study is to confirm the effectiveness of limited resection for histologically confirmed BAC without active fibroblastic proliferation. METHODS: From 1996 through 1999, 42 patients who had small peripheral lung tumors (< or = 20 mm), suspected of being BAC, were enrolled in this trial. The patient population consisted of 24 men and 18 women with a mean age of 58.4 years. Limited resection was completed when BAC, without both active fibroblastic proliferation and lymph node metastasis, was confirmed histologically by intraoperative pathologic examination. RESULTS: Limited resection was completed in 36 patients, wedge resection in 34, and segmentectomy in 2 patients. In 6 patients, the procedure was converted into lobectomy because of pathologic invasive sign in 3, active fibroblastic proliferation in 1, and for other reasons in 2 patients. All patients have been followed for a median follow-up period of 30 months and are alive without sign of recurrence. CONCLUSIONS: Our early results indicate that limited resection may be an acceptable alternative to lobectomy for histologically confirmed BAC without active fibroblastic proliferation. PMID- 11269484 TI - Surgical treatment of hepatic and pulmonary metastases from colon cancer. AB - BACKGROUND: Surgical resection of isolated hepatic or pulmonary metastases secondary to colorectal cancer has been shown to yield acceptable long-term survival. However, results are inconclusive for surgical resection of both hepatic and pulmonary metastases. METHODS: We reviewed the records of all patients who underwent surgical resection of both hepatic and pulmonary metastases from colorectal cancer between 1980 and 1998. RESULTS: A total of 58 patients underwent resection of both hepatic and pulmonary metastases secondary to colorectal cancer. All patients had local control of their primary cancer before metastasectomy. There were no operative deaths. Morbidity occurred in 12% of patients. Follow-up was complete in all patients, with a median duration of 62 months (range, 6 to 201 months). The 5- and 10-year survivals were 30% and 16%, respectively. A premetastasectomy carcinoembryonic antigen level greater than 5 ng/mL increased the risk of early death (p = 0.029). Neither the number of pulmonary lesions nor the time interval between the primary surgery and the metastasectomy had a significant impact on survival (p = 0.67). At 5 years, 55% of patients were free of disease. Four patients had lymph node involvement at the time of pulmonary resection and all 4 patients died within 22 months of their pulmonary metastasectomy. CONCLUSIONS: Resection of both hepatic and pulmonary metastases secondary to colorectal cancer in highly selected patients is safe and results in long-term survival. Thoracic lymph node involvement and elevated carcinoembryonic antigen levels before pulmonary metastasectomy are associated with reduced survival. PMID- 11269485 TI - Resection of adrenal metastases from non-small cell lung cancer: a multicenter study. AB - BACKGROUND: In recent case reports and limited series, adrenalectomy was recommended for an isolated adrenal metastasis from non-small cell lung cancer (NSCLC). METHODS: We retrospectively studied patients with a solitary adrenal metastasis from NSCLC who had undergone potentially curative resection in eight centers. RESULTS: Forty-three patients were included. Their adrenal gland metastasis was discovered synchronously with NSCLC in 32 patients, and metachronously in 11. It was homolateral to the NSCLC in 31 patients and contralateral in 12 (p < 0.01). Median survival was 11 months, and 3 patients survived more than 5 years. There was no difference between the synchronous and metachronous groups regarding recurrence rate or survival. Survival was not affected by the homolateral location of the metastasis, the histology of the NSCLC, TNM stage, any adjuvant and neoadjuvant treatment, or, in the metachronous group, a disease-free interval exceeding 6 months. CONCLUSIONS: We confirm the possibility of long-term survival after resection of isolated adrenal metastasis from NSCLC, but no clinical or pathologic criteria were detected to identify patients amenable to potential cure. PMID- 11269486 TI - Thoracoscopic resection of pulmonary nodules after computed tomographic-guided coil labeling. AB - BACKGROUND: A limiting factor in performing video-assisted thoracic surgery for resection of peripheral solitary pulmonary nodules has been the recognition of the lesion visually. This study reports our clinical experience of injecting a small metallic marker under computed tomographic scan guidance before the operation, allowing localization of the lesion. METHODS: A series of 14 patients underwent video-assisted thoracic surgery for removal of 15 pulmonary nodules situated in the outer third of the lung. Before operation, a radiopaque microcoil was injected just behind the lesion and then used to locate, under fluoroscopy, the area to be resected during thoracoscopy. The technique was evaluated for accuracy, reliability, and ease of use. RESULTS: Microcoil labeling of peripheral pulmonary nodules allowed in every case a complete resection and a histologic identification of the lesion. It is more stable and accurate than methylene blue dye marking, and it is as easy to perform as computed tomographic scan-guided biopsy. The incidence of complication was small in spite of our inexperience with the technique. CONCLUSIONS: Our experience with microcoil injection shows that it provides consistent and highly accurate marking of pulmonary nodules for video assisted thoracic surgery, allowing secure resection with a safe margin. PMID- 11269487 TI - Airway complications after lung transplantation: treatment and long-term outcome. AB - BACKGROUND: Airway complications are a significant cause of morbidity after lung transplantation. Effective treatment reduces the impact of these complications. METHODS: Data from 123 lung (99 single, 24 bilateral) transplants were reviewed. Potential risk factors for airway complications were analyzed. Stenoses were treated with expanding metal (Gianturco) stents. RESULTS: Mean follow-up was 749 days. Thirty-five complications developed in 28 recipients (complication rate: 23.8%/anastomosis). Mean time to diagnosis was 47 days. Only Aspergillus infection and airway necrosis were significantly associated with development of complications (p < 0.00001 and p < 0.03, respectively). Stenosis was diagnosed an average of 42 days posttransplant. Average decline in forced expiratory volume in 1 second (FEV1) was 39%. Eighteen patients (13 single and 5 bilateral) required stent insertion. Mean increase in FEV1 poststenting was 87%. Two stent patients died from infectious complications. Six patients required further intervention. Long-term survival and FEV1 did not differ from nonstented patients. CONCLUSIONS: Aspergillus and airway necrosis are associated with the development of airway complications. Expanding metal stents are an effective long-term treatment. PMID- 11269488 TI - Economic analysis of lung volume reduction surgery as part of the National Emphysema Treatment Trial. NETT Research Group. AB - BACKGROUND: In today's cost-conscious health care environment, obtaining timely and accurate economic information regarding new medical technologies has become extremely important. The National Emphysema Treatment Trial, a multicenter, randomized controlled trial of lung volume reduction surgery (LVRS) plus medical therapy, versus medical therapy for patients with severe emphysema, includes a parallel cost-effectiveness analysis. METHODS: The analysis is designed to determine the cost-effectiveness of LVRS versus medical therapy for those who are eligible for the procedure. After describing theoretical foundations of cost effectiveness analysis as they apply to this study, we describe the economic and quality of life data that are being collected alongside the clinical trial, methods of analysis, and approach to presenting the results. RESULTS: The cost effectiveness of LVRS relative to medical therapy will be presented as costs per quality-adjusted life years gained. CONCLUSIONS: This analysis will provide timely economic data that can be considered alongside the clinical results of the National Emphysema Treatment Trial. As one of the largest clinical trials to include a parallel, prospective cost-effectiveness analyses, this study will also provide valuable practical information about conducting an economic analysis alongside a multicenter clinical trial. PMID- 11269489 TI - Cautious advance. Gene therapy is more complex than anticipated. PMID- 11269490 TI - Research on human embryonic stem cells. NIH issues new guidelines a week after British recommendations. PMID- 11269491 TI - Toxins for terrorists. Do scientists act illegally when sending out potentially dangerous material? PMID- 11269492 TI - Fight for reputation. Judah Folkman counter-sued Abbott in the legal battle over kringle 5. PMID- 11269493 TI - Fusion proteins and fusion pores. Workshop: regulated exocytosis and the vesicle cycle. PMID- 11269494 TI - Organization of complex situations in the immune system. Conference: signal processing through protein complexes. PMID- 11269495 TI - Chunnel vision. Export and efflux through bacterial channel-tunnels. AB - The Escherichia coli TolC protein is central to toxin export and drug efflux across the inner and outer cell membranes and the intervening periplasmic space. The crystal structure has revealed that TolC assembles into a remarkable alpha helical trans-periplasmic cylinder (tunnel) embedded in the outer membrane by a contiguous beta-barrel (channel), so providing a large duct open to the outside environment. The channel-tunnel structure is conserved in TolC homologues throughout Gram-negative bacteria, and it is envisaged that they are recruited and opened, through a common mechanism, by substrate-specific inner-membrane complexes. PMID- 11269496 TI - The Cdc7/Dbf4 protein kinase: target of the S phase checkpoint? AB - Cdc7/Dbf4 is a protein kinase that is required for the initiation of DNA replication in eukaryotes. Recent work has provided new clues to the role that Cdc7/Dbf4 plays in this process. A range of other observations suggest that Cdc7/Dbf4 also plays another, less well characterized, role in checkpoint function and in the maintenance of genomic integrity. In this review we attempt to bring together new information to explain how Cdc7/Dbf4 may perform these two distinct functions. PMID- 11269497 TI - New genes with old modus operandi. The connection between supercoiling and partitioning of DNA in Escherichia coli. AB - The process of partitioning bacterial sister chromosomes into daughter cells seems to be distinct from chromatid segregation during eukaryotic mitosis. In Escherichia coli, partitioning starts soon after initiation of replication, when the two newly replicated oriCs move from the cell centre to quarter positions within the cell. As replication proceeds, domains of the compact, supercoiled chromosome are locally decondensed ahead of the replication fork. The nascent daughter chromosomes are recondensed and moved apart through the concerted activities of topoisomerases and the SeqA (sequestration) and MukB (chromosome condensation) proteins, all of which modulate nucleoid superhelicity. Thus, genes involved in chromosome topology, once set aside as 'red herrings' in the search for 'true' partition functions, are again recognized as being important for chromosome partitioning in E. coli. PMID- 11269498 TI - DNA conformation driven by AP-1 triggers cell-specific expression via a strong epithelial enhancer. AB - We report here the characterization of the regulatory region of the human LAMA3 gene, coding for the alpha3A chain of laminin-5. A 202 bp fragment is sufficient to confer epithelial-specific expression to a thymidine kinase promoter through the cooperative effect of three AP-1 binding sites. Remarkably, removal of the sequences located between the AP-1 sites does not modify the promoter activity in keratinocytes but allows strong expression in fibroblasts. Replacement of the deleted sequences by non-homologous ones fully restores the restricted enhancement in keratinocytes. Functional analysis and mutagenesis experiments demonstrate that a minimal distance between the AP-1 sites is required for the enhancer DNA fragment to adopt a particular conformation driven by the binding of Jun-Fos heterodimers. In non-permissive cells, this conformation leads to the anchorage of non-DNA-binding fibroblastic cofactors to form an inhibitory ternary complex. Therefore, our results describe for the first time an unusual conformation-dependent epithelial-specific enhancer. PMID- 11269499 TI - Uncoupling yeast intron recognition from transcription with recursive splicing. AB - Pre-mRNA splicing has to be coordinated with other processes occurring in the nucleus including transcription, mRNA 3' end formation and mRNA export. To analyze the relationship between transcription and splicing, we constructed a network of nested introns. Introns were inserted in the 5' splice site and/or branchpoint of a synthetic yeast intron interrupting a reporter gene. The inserted introns mask the recipient intron from the cellular machinery until they are removed by splicing. Production of functional mRNA from these constructs therefore requires recognition of a spliced RNA as a splicing substrate. We show that recurrent splicing occurs in a sequential and ordered fashion in vivo. Thus, in Saccharomyces cerevisiae, intron recognition and pre-spliceosome assembly is not tightly coupled to transcription. PMID- 11269500 TI - Efficient membrane assembly of the KcsA potassium channel in Escherichia coli requires the protonmotive force. AB - Very little is known about the biogenesis and assembly of oligomeric membrane proteins. In this study, the biogenesis of KcsA, a prokaryotic homotetrameric potassium channel, is investigated. Using in vivo pulse-chase experiments, both the monomeric and tetrameric form could be identified. The conversion of monomers into a tetramer is found to be a highly efficient process that occurs in the Escherichia coli inner membrane. KcsA does not require ATP hydrolysis by SecA for insertion or tetramerization. The presence of the proton-motive force (pmf) is not necessary for transmembrane insertion of KcsA; however, the pmf proved to be essential for the efficiency of oligomerization. From in vivo and in vitro experiments it is concluded that the electrical component, deltapsi, is the main determinant for this effect. These results demonstrate a new role of the pmf in membrane protein biogenesis. PMID- 11269501 TI - Activation of the Drosophila NF-kappaB factor Relish by rapid endoproteolytic cleavage. AB - The Rel/NF-kappaB transcription factor Relish plays a key role in the humoral immune response in Drosophila. We now find that activation of this innate immune response is preceded by rapid proteolytic cleavage of Relish into two parts. An N terminal fragment, containing the DNA-binding Rel homology domain, translocates to the nucleus where it binds to the promoter of the Cecropin A1 gene and probably to the promoters of other antimicrobial peptide genes. The C-terminal IkappaB-like fragment remains in the cytoplasm. This endoproteolytic cleavage does not involve the proteasome, requires the DREDD caspase, and is different from previously described mechanisms for Rel factor activation. PMID- 11269502 TI - The Drosophila caspase Dredd is required to resist gram-negative bacterial infection. AB - The Drosophila innate immune system discriminates between pathogens and responds by inducing the expression of specific antimicrobial peptide-encoding genes through distinct signaling cascades. Fungal infection activates NF-kappaB-like transcription factors via the Toll pathway, which also regulates innate immune responses in mammals. The pathways that mediate antibacterial defenses, however, are less defined. We have isolated loss-of-function mutations in the caspase encoding gene dredd, which block the expression of all genes that code for peptides with antibacterial activity. These mutations also render flies highly susceptible to infection by gram-negative bacteria. Our results demonstrate that Dredd regulates antibacterial peptide gene expression, and we propose that Dredd, Immune Deficiency and the P105-like rel protein Relish define a pathway that is required to resist gram-negative bacterial infections. PMID- 11269503 TI - HNF1alpha controls renal glucose reabsorption in mouse and man. AB - Recently it has been shown that dominant mutations in the human hepatocyte nuclear factor 1alpha (HNF1alpha) gene, encoding for a homeoprotein that is expressed in liver, kidney, pancreas and intestine, result in maturity onset diabetes of the young type 3 (MODY3). HNF1alpha-null mice are diabetic, but at the same time suffer from a renal Fanconi syndrome characterized by urinary glucose loss. Here we show that MODY3 patients are also characterized by a reduced tubular reabsorption of glucose. The renal murine defect is due to reduced expression of the low affinity/high capacity glucose cotransporter (SGLT2). Our results show that HNF1alpha directly controls SGLT2 gene expression. Together these data indicate that HNF1alpha plays a key role in glucose homeostasis in mammals. PMID- 11269506 TI - T cell mediated neuroprotection is a physiological response to central nervous system insults. AB - The physiological conditions under which beneficial autoimmunity is evoked have never been documented. We recently demonstrated that autoimmune T cells directed against myelin-associated self-proteins, when passively transferred into rats or mice, reduce the spread of damage after traumatic injury to central nervous system axons. This finding raised a fundamental question: does this beneficial effect represent a physiological neuroprotective response that normally is too weak to be effective and requires boosting, or is it simply the welcome result of an ex vivo manipulation? It appears from our studies that trauma, at least in the central nervous system, evokes a stress signal that activates a T cell dependent response directed against self antigens, and that this response is physiological in nature, beneficial in intent, and amenable to boosting by active or passive immunization. PMID- 11269504 TI - Tip cell-derived RTK signaling initiates cell movements in the Drosophila stomatogastric nervous system anlage. AB - The stomatogastric nervous system (SNS) of Drosophila is a simply organized neural circuitry that innervates the anterior enteric system. Unlike the central and the peripheral nervous systems, the SNS derives from a compact epithelial anlage in which three invagination centers, each giving rise to an invagination fold headed by a tip cell, are generated. Tip cell selection involves lateral inhibition, a process in which Wingless (Wg) activity adjusts the range of Notch signaling. Here we show that RTK signaling mediated by the Drosophila homolog of the epidermal growth factor receptor, DER, plays a key role in two consecutive steps during early SNS development. Like Wg, DER signaling participates in adjusting the range of Notch-dependent lateral inhibition during tip cell selection. Subsequently, tip cells secrete the DER ligand Spitz and trigger local RTK signaling, which initiates morphogenetic movements resulting in the tip cell directed invaginations within the SNS anlage. PMID- 11269505 TI - Basic renal physiology for molecular biologists. PMID- 11269507 TI - Suicide gene therapy for pediatric tumors. AB - Tumor gene therapy is potentially very specific and efficacious. Suicide genes are promising tools in the arsenal of tumor gene therapy. However, problems of tumor targeting, low in vivo efficacy of nucleic acid transfer, and recent reports of adverse effects hinder the translation of this approach into clinical practice. Therefore vector design, tumor targeting, mechanisms of cell kill and killing of untransfected tumor cells must be improved. Once these problems are solved in vitro and in animal models, gene therapy holds great promise for pediatric oncology given the abundance of specific targets in pediatric tumors. This review describes the current state of preclinical research in tumor suicide gene therapy, provides an outline of pediatric suicide gene therapy protocols, and identifies potential targets in pediatric malignancies. PMID- 11269508 TI - Control of the cell survival/death decision by cannabinoids. AB - Cannabinoids, the active components of Cannabis sativa (marijuana), and their derivatives produce a wide spectrum of central and peripheral effects, some of which may have clinical application. The discovery of specific cannabinoid receptors and a family of endogenous ligands of those receptors has attracted much attention to cannabinoids in recent years. One of the most exciting and promising areas of current cannabinoid research is the ability of these compounds to control the cell survival/death decision. Thus cannabinoids may induce proliferation, growth arrest, or apoptosis in a number of cells, including neurons, lymphocytes, and various transformed neural and nonneural cells. The variation in drug effects may depend on experimental factors such as drug concentration, timing of drug delivery, and type of cell examined. Regarding the central nervous system, most of the experimental evidence indicates that cannabinoids may protect neurons from toxic insults such as glutamaergic overstimulation, ischemia and oxidative damage. In contrast, cannabinoids induce apoptosis of glioma cells in culture and regression of malignant gliomas in vivo. Breast and prostate cancer cells are also sensitive to cannabinoid-induced antiproliferation. Regarding the immune system, low doses of cannabinoids may enhance cell proliferation, whereas high doses of cannabinoids usually induce growth arrest or apoptosis. The neuroprotective effect of cannabinoids may have potential clinical relevance for the treatment of neurodegenerative disorders such as multiple sclerosis, Parkinson's disease, and ischemia/stroke, whereas their growth-inhibiting action on transformed cells might be useful for the management of malignant brain tumors. Ongoing investigation is in search for cannabinoid-based therapeutic strategies devoid of nondesired psychotropic effects. PMID- 11269509 TI - The GAA repeat expansion in intron 1 of the frataxin gene is related to the severity of cardiac manifestation in patients with Friedreich's ataxia. AB - Friedreich's ataxia is an autosomal recessive disorder characterized by spinocerebellar degeneration. It is caused by an unstable GAA trinucleotide repeat expansion (>120 repeats) in the first intron of the frataxin gene on chromosome 9 (9q13) in both alleles. Concentric left ventricular hypertrophy has been recognized as the major cardiac manifestation of Friedreich's ataxia. Our aim was to investigate the influence of the frataxin repeat length on cardiac hypertrophy in patients with Friedreich's ataxia and in patients with hypertrophic and dilated cardiomyopathy. Thirty-one patients with Friedreich's ataxia, 86 patients with hypertrophic cardiomyopathy, 134 patients with dilated cardiomyopathy, and 32 healthy individuals without cardiac disease were analysed by electrocardiography and 2D-M-mode echocardiography. Then, the size of the frataxin repeat was determined by polymerase chain reaction (PCR) and agarose gel electrophoresis. The number of GAA repeats in patients with hypertrophic and dilated cardiomyopathy was not different from the length in patients without cardiac disease (hypertrophic cardiomyopathy, 8+/-2 repeats on GAA 1 allele and 11+/-5 repeats on GAA 2 allele; dilated cardiomyopathy, 7+/-2 repeats on GAA 1 allele and 11+/-5 repeats on GAA 2 allele; Control, 9+/-1 repeats on GAA 1 allele and 12+/-6 repeats on GAA 2 allele). The septal and posterior wall thickness of these patients was not related to the GAA repeat length. All patients with Friedreich's ataxia had two enlarged alleles with a mean GAA repeat length of 757+/-316 and 1012+/-231, respectively. The lengths of both alleles were significantly greater than the lengths in the controls (P<0.0001), patients with hypertrophic cardiomyopathy (P<0.0001) and dilated cardiomyopathy (P<0.0001). A significant correlation was revealed between interventricular septal hypertrophy and frataxin repeat length in the smaller allele. Furthermore, the ratio of septal to posterior wall thickness was significantly correlated to GAA repeat size on the smaller allele. In conclusion, the size of the GAA repeat on the smaller allele in the frataxin gene is associated with the degree of left ventricular hypertrophy in patients with Friedreich's ataxia but is not related to the severity of hypertrophic cardiomyopathy. PMID- 11269510 TI - Impaired sodium excretion, decreased glomerular filtration rate and elevated blood pressure in endothelin receptor type B deficient rats. AB - The renal endothelin (ET) system, particularly the ET type B receptor, has been implicated in the regulation of sodium excretion and glomerular filtration rate (GFR). We analyzed kidney morphology and function in a rat strain characterized by complete absence of a functional ETB receptor. Due to Hirschsprung's disease limiting lifetime in these rats, studies were performed in 23-day-old rats. Kidney size and morphology (glomerular and interstitial matrix content, glomerular size and cell density and intrarenal vascular morphology) were normal in ETB-deficient rats. There were also no evidence of altered kidney cell cycle regulation in these rats. GFR was significantly lower, by 72% (P<0.001), in homozygous ETB-deficient rats than in wild-type rats. Fractional sodium excretion was likewise markedly reduced by 84% in homozygous ETB-deficient rats (P<0.001 versus wild-type rats). Treatment with the specific epithelial sodium channel blocker amiloride led to a much higher increase in fractional sodium excretion in ETB-deficient rats (934.2+/-73% in ETB-deficient rats versus 297+/-20% in wild type rats, expressed as percentage of corresponding placebo treated control; P<0.001). Mean arterial blood pressure was elevated by 7.9 mmHg in homozygous ETB deficient rats (P<0.05 versus wild-type rats). Our study demonstrates that ETB deficiency causes early onset kidney dysfunction characterized by a markedly reduced sodium excretion, decreased GFR, and slightly elevated blood pressure. The complete absence of the ETB receptor causes in the kidney--in contrast to the colon--a functional rather than a developmental, neural crest cell dependent disease, since kidney morphology was normal in ETB-deficient rats. The much higher increase in the fractional sodium excretion in ETB-deficient rats after pharmacological blockade of the epithelial sodium channel indicates that the decreased fractional sodium excretion in ETB-deficient rats is most probably due to a lack of the inhibitory property of the ETB receptor on the epithelial sodium channel activity. PMID- 11269511 TI - Cytomegalovirus inhibits p53 nuclear localization signal function. AB - Endothelial cells (EC) infected with the VHL strain of cytomegalovirus (CMV) are resistant to p53-mediated apoptosis, which may be relevant to EC dysfunction and atherogenesis. This resistance to apoptosis may be mediated by cytoplasmic sequestration of p53, which functions only in the nucleus. We explored the hypothesis that CMV sequesters p53 in the cytoplasm by blocking p53 nuclear localization signal (NLS) function. We transfected VHL CMV infected EC with recombinant p53 NLSI conjugated with chicken muscle pyruvate kinase (PK) plasmid. NLSI is responsible for 90% of p53 nuclear localization, and PK is not normally translocated to the nucleus after cytoplasmic production. Thus it cannot be localized in the nucleus without the assistance of the artificial NLSI. A double labeling immunofluorescence staining method was used to identify the localization of p53 NLSI-conjugated PK in CMV-infected EC. We found that CMV infection sequesters PK and p53 in the cytoplasm by blocking NLSI function. This inactivation of NLSI function is dependent upon infection stage; it occurs only in the early and late phases and not the immediate early phase of infection. These findings may be relevant to endothelial dysfunction and initiation of atherogenesis. Our study also suggests a novel mechanism of the p53 inactivation by virus, which may be important for atherogenesis and tumorgenesis. PMID- 11269513 TI - Advances in sample preparation in electromigration, chromatographic and mass spectrometric separation methods. AB - The quality of sample preparation is a key factor in determining the success of analysis. While analysis of pharmaceutically important compounds in biological matrixes has driven forward the development of sample clean-up procedures in last 20 years, today's chemists face an additional challenge: sample preparation and analysis of complex biochemical samples for characterization of genotypic or phenotypic information contained in DNA and proteins. This review focuses on various sample pretreatment methods designed to meet the requirements for the analysis of biopolymers and small drugs in complex matrices. We discuss the advances in development of solid-phase extraction (SPE) sorbents, on-line SPE, membrane-based sample preparation, and sample clean-up of biopolymers prior to their analysis by mass spectrometry. PMID- 11269512 TI - Mutation analysis of the MECP2 gene in British and Italian Rett syndrome females. AB - Rett syndrome is an X-linked dominant neurological disorder, which appears to be the commonest genetic cause of profound combined intellectual and physical disability in Caucasian females. Recently, this syndrome has been associated with mutations of the MECP2 gene, a transcriptional repressor of still unknown target genes. Here we report a detailed mutational analysis of 62 patients from UK and Italian archives, representing the first comparative study among different populations and one of the largest number of cases so far analyzed. We review the literature on MECP2 mutations in Rett syndrome. This analysis has permitted us to produce a map of the recurrent mutations identified in this and previous studies. Bioinformatic analysis of the mutations, taking advantage of structural and evolutionary data, leads us to postulate the existence of a new functional domain in the MeCP2 protein, which is conserved among brain-specific regulatory factors. PMID- 11269515 TI - Chiroptical detection during liquid chromatography 7. The rotation angle/absorbance ratio of chiral molecules. Its possible use for on-line analysis during preparative separations of enantiomers. AB - The rotation angle/absorbance ratios C+ = alpha+/A+ and C- = a-/A-, determined via detection by a polarimeter and a photometer, were checked for the first time with reference to their use for on-line analysis during preparative separations. For this purpose, (+)-, (-)- and (+/-)-carvones were investigated by liquid chromatography (LC) on microcrystalline tribenzoylcellulose. It turned out that the ratios C+ and C- depend only slightly upon concentration (Table 1). Overlapped peaks of enantiomers were successfully submitted to computer deconvolution (e.g. Fig. 2, bottom). A procedure for on-line analysis during preparative LC is proposed. PMID- 11269514 TI - Shielding of protein-boronate interactions during boronate chromatography of neoglycoproteins. AB - A method for separating glycoproteins on a boronate column under conditions which suppress the interactions between the protein moiety and the boronic acid ligand has been developed. A model system consisting of non-glycosylated chymotrypsin and maltose-modified chymotrypsin (cht-mal) was utilised in the investigations. Chymotrypsin was chosen as the model protein because of its known interaction with boronate. By coupling maltose to chymotrypsin, a neoglycoprotein was created which has the property of binding to the affinity matrix both via the protein moiety and via the carbohydrate residues. The introduction of a so-called shielding reagent into the buffer solutions during chromatography resulted in the prevention of the protein-boronate interactions while the carbohydrate-boronate interaction was little influenced. Different types of, mainly low-molecular-mass, polyhydroxyl chemicals were screened in order to correlate the shielding efficiency to the chemical structure of the investigated compounds. Polyhydroxyl chemicals with a conformation that allows the formation of tridentate complexes with the boronate anion provided the highest shielding efficiencies. PMID- 11269516 TI - Characterization and identification of a "mystery" oil spill from Quebec (1999). AB - This paper describes a case study in which advanced chemical fingerprinting and data interpretation techniques were used to characterize the chemical compositions and determine the source of an unknown spilled oil from Quebec. On 28 February 1999, significant amounts of oil was reported on the river banks of St. Laurence River in front of a company named "Thermex" (in a town - Beauharnois, Quebec, about 50 km northwest of Montreal). The spilled oil was suspected to be released from a nearby factory. In response to this specific site investigation needs, a tiered analytical approach using GC-MS and GC-flame ionization detection was applied. A variety of diagnostic ratios of "source specific marker" compounds, in particular isomers of biomarkers and alkylated series of polycyclic aromatic hydrocarbons within the same alkylation groups, were determined and analyzed. The hydrocarbon analysis results reveal the following: (1) the spilled oil is very "specific", and is significantly different from most crude oils in chemical composition; (2) the oil in samples come from the same source, however, the spill sample 2569 was identified to contain a small amount (approximately 10%) of diesel; (3) the spilled oil was relatively "fresh", its chemical composition has not undergone significant alteration yet; (4) the spilled oil showed unusually high concentration of the US Environmental Protection Agency priority polycyclic aromatic hydrocarbons (PAHs). The "Pyrogenic Index" values were determined to be as high as 0.11-0.13, significantly higher than crude oils (<0.010) and heavy Bunker type fuels (0.015 0.060). This indicates significant contribution of PAH composition from pyrogenic components; (5) biomarkers were also detected, but their concentrations were unusually low in comparison to most crude oils. PMID- 11269517 TI - Determination of alcohols by high-performance liquid chromatography with fluorimetric detection after pre-column derivatisation with carbazole-9-N-acetic acid and chromatographic behaviour of alcoholic derivatives. AB - Alcohols were derivatised to their carbazole-9-N-acetic acid (CRA) esters with 1 ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC.HCl) as the dehydrating agent. Studies on derivatisation conditions indicated that the coupling reaction proceeded rapidly and smoothly in the presence of a base catalyst in acetonitrile to give the corresponding sensitively fluorescent derivatives. The retention behaviour of alcohol derivatives was investigated by varying mobile phase compositions (ACN-water and MeOH-water). The parameters from the equation log k'=A-BX were evaluated by retention data of derivatives using an isocratic elution with different mobile phases. The results indicated that the parameters derived allowed computation of retention factors in good agreement with experiments. At the same time, a general equation was derived that makes possible predictions of partition coefficient in binary mobile phases with different proportions of organic solvent to water based on some simple regression analysis. The LC separation for the derivatised alcohols containing higher carbon alcohols showed good reproducibility on a reversed-phase C18 column with gradient elution. The detection limits (excitation at 335 nm, emission at 360 nm) for derivatised alcohols (signal-to-noise ratio=3:1) were in the range of 0.1-0.4 pg per injection. PMID- 11269518 TI - Characterization of metal affinity of green fluorescent protein and its purification through salt promoted, immobilized metal affinity chromatography. AB - Immobilized metal affinity chromatography (IMAC) was investigated as a method of recovery for green fluorescent protein (GFPuv). It was found that in the absence of genetic modification to enhance metal affinity, GFPuv displayed strong metal affinity to Cu(II) and Ni(II), and weak or negligible affinity to Zn(II) and Co(II). Changes in the mobile phase NaCl concentration during Ni(II)-IMAC strongly affected purity and yield of GFPuv, with fine resolution under higher NaCl concentrations. Finally, IMAC via Cu(II) and Zn(II) with intervening diafiltration was used to recover GFPuv with high yield and purity. PMID- 11269520 TI - Determination of the major isosteroidal alkaloids in bulbs of Fritillaria by high performance liquid chromatography coupled with evaporative light scattering detection. AB - A new direct HPLC analytical method using evaporative light scattering detection coupled with a low-temperature adapter for the simultaneous determination of the major biologically active isosteroidal alkaloids in Bulbus Fritillariae, a commonly used antitussive traditional Chinese medicinal (TCM) herb, has been developed. The simultaneous separation of eight Fritillaria alkaloids was achieved on a reversed-phase C8 column with an isocratic mobile phase system consisting of acetonitrile-methanol-water (66.5:3.5:30, v/v) containing 0.006% triethylamine. This method provides good reproducibility and sensitivity for the quantification of six major isosteroidal alkaloids, namely peimissine, verticine, verticinone, imperialine, isoverticine and ebeiedine in different Fritillaria species with overall intra- and inter-day precision and accuracy of less than 11% and higher than 90%, respectively. The assay was successfully utilized to quantify the major biologically active alkaloids in five Fritillaria species. The results demonstrate that this method is simple, selective, and suitable for the quality control of this commonly used antitussive TCM herb, Bulbus Fritillariae. reserved. PMID- 11269519 TI - Effect of buffer concentration on gradient chromatofocusing performance separating protiens on a high-performance DEAE column. AB - Gradient chromatofocusing is a recently developed chromatographic technique that overcomes the limitations of conventional chromatofocusing. This technique employs a HPLC gradient system and simple low-molecular-mass buffer components to generate linear or other function pH gradients on ion-exchange columns. Results of the present work show a superior separation of beta-lactoglobulin A and B in gradient chromatofocusing compared to salt gradient chromatography using the same DEAE column, with an optimized resolution of 2.3 obtained with gradient chromatofocusing compared to 1.1 obtained with NaCl gradients at constant pH. A significant advantage of the gradient chromatofocusing technique over the conventional chromatofocusing technique is its ability to employ a relatively wide range of buffer concentrations in the mobile phase, the effect of which is studied in the present work. Five proteins (conalbumin, ovalbumin, bovine serum albumin, beta-lactoglobulin A and B) are chromatographed on a DEAE polymethacrylate HPLC anion-exchange column using the same approximately linear pH gradient profile but different mobile phase buffer concentrations. Results show a significant effect of buffer concentration on peak width, separation factor and resolution. For example, resolution increases from 1.5 to 2.3 in the separation of beta-lactoglobulin A and B when the concentration of each of the components in the 100% elution buffer is increased from 6.25 to 25.0 mM (with the same outlet pH gradient). This separation trend is also seen in the chromatography of ovalbumin from a commercial source, noting a progressive increase in resolution of two peaks in the sample (resolution increased from 0.7 to 2.4) when the concentration of each of the components in the 100% elution buffer is increased from 6.25 to 37.5 mM (same outlet pH gradient). The gains in the resolution are attributed to an increase in the separation factor, since the peak widths are generally noted to also increase with increased buffer concentration. These results point to a significant interplay between buffer concentration and pH, which is not effectively exploited in either conventional chromatofocusing or in conventional ion-exchange chromatographic procedures employing salt gradient elution at constant pH. Gradient chromatofocusing has the ability of optimizing both parameters, thus providing it with unique capabilities in protein separations. PMID- 11269521 TI - Determination of RS,E/Z-tocotrienols by HPLC. AB - Synthetic alpha-tocotrienol was separated into four geometrical E/Z side chain isomers by preparative HPLC (permethylated beta-cyclodextrin phase). The isolated isomers were resolved in ethylene glycol dimethyl ether, converted into the corresponding methyl ether using dimethyl sulfate, and the tocotrienol methyl ethers were extracted with n-hexane. A subsequent HPLC separation on a chiral phase (adsorbent cellulose derivated with 3,5-dimethyl phenyl carbamate) discriminates between the enantiomers of each E/Z side chain isomer, achieving the complete resolution of the eight occurring synthetic RS,E/Z-alpha tocotrienols. The method can be shortened by omitting the preparative separation of the E/Z tocotrienol isomers prior to the chromatography on the chiral dimethyl phenyl carbamate phase. The simplified method achieved the following separation: RS,E/Z-alpha-tocotrienol separated into five peaks, RS,E/Z-beta-tocotrienol into eight, RS,E/Z-gamma-tocotrienol into six and RS,E/Z-delta-tocotrienol into eight peaks. The naturally occurring R,E-E-tocotrienol isomer could be identified within the synthetic RS,E/Z-isomers by co-chromatography with tocotrienol methyl ethers derived from natural sources, respectively. PMID- 11269522 TI - High-performance liquid chromatographic separation of microcystins and nodularin, cyanobacterial peptide toxins, on C18 and amide C16 sorbents. AB - Four C18 columns and a novel amide C16 column were assessed in the HPLC separation of eight microcystins and nodularin-R. Gradient mobile phases of acetonitrile combined with trifluoroacetic acid, formic acid or ammonium acetate were compared. Special attention was paid to the resolution of four possible coeluting microcystin pairs. Generally speaking, the acidic mobile phases were superior to the ammonium acetate-based mobile phase in terms of resolution and selectivity. The amide C16 column had the best overall performance and unique selectivity properties. PMID- 11269523 TI - Determination of volatile components in peptic powder by gas chromatography-mass spectrometry and chemometric resolution. AB - Gas chromatography-mass spectrometry (GC-MS) coupled with chemometric resolution upon two-dimensional data was proposed as a method for the analysis of volatile components in a traditional Chinese medicinal preparation peptic powder which contains Rhizoma Atractylodis, Pericarpium Citri Reticulatae, Cortex Magnoliae Officinalis and Radix Glycyrrhizae. Ninety-three components were separated and 65 of them were qualitatively and quantitatively analyzed which represented about 90.28% of the total content. With the help of chemometric resolution, the data were resolved into a pure chromatogram and a mass spectrum of each chemical component. The accuracy of qualitative and quantitative results was greatly improved by using the two-dimensional comprehensive information of chromatograms and mass spectra. The example showed that chemometric resolution could greatly enhance separation ability. This makes it possible to analyze complicated practical systems like traditional Chinese medicinal preparations with the help of coupled instruments and chemometric resolution methods. PMID- 11269524 TI - Analysis of nonvolatile species in a complex matrix by headspace gas chromatography. AB - This study developed a phase reaction conversion (PRC) headspace gas chromatographic (HS-GC) technique for the measurements of nonvolatile species in liquid or solid samples. The technique is demonstrated by the measurements of carbonate in aqueous carbonate solutions and in kraft pulp mill liquor samples. A very small amount of sulfuric acid (volume of 0.5 ml, concentration of 2 mol/l) is used to acidify a sample of less than 300 microl in volume and convert the dissolved carbonate into carbon dioxide (gas) in a sample vial (reactor) that is analyzed by thermal conductivity detection through a headspace sampler. The carbonate concentrations measured by PRC-HS-GC in seven kraft liquor samples agree very well with those measured using a coulometric and a titrametric method. Simultaneous analysis of multiple species was also conducted to demonstrate the versatility of the method. The present method is very simple, rapid, reliable, accurate, and fully automated. It can be applied to analyze other nonvolatile species in various industrial and environmental samples. PMID- 11269525 TI - Multivariate calibration methods for quantification in strongly overlapping capillary electrophoretic peaks. AB - Capillary zone electrophoresis with diode-array detection was applied to the separation of ebrotidine and its metabolites. However, three of these, which are neutral in the conditions studied, co-migrated with the electroosmotic flow signal. Therefore, strongly overlapping peaks were observed. The main aim of this study was to show the potentiality of capillary electrophoresis in combination with chemometrics. Multivariate calibration methods were applied to quantify these analytes in synthetic mixtures. The results obtained using partial least squares (PLS) were in agreement with actual values, with an overall prediction error of 9.7%. PMID- 11269526 TI - Mixed triblock copolymers used as DNA separation medium in capillary electrophoresis. AB - A polymer solution, formed by mixing two polyoxybutylene-polyoxyethylene polyoxybutylene (BEB) triblock copolymers (B10E270B10 and B6E46B6), was tested as a new separation medium for double-stranded DNA separation in capillary electrophoresis. The mixture of B10E270B10 and B6E46B6 has a viscosity-adjustable property and a dynamic coating ability, which makes the medium very easy to handle. The performance of the mixture on the DNA separation is greatly affected by the mass ratio of the two constituents. There is a minimum amount of concentration for B10E270B10, below which the medium will lose its performance. The addition of B6E46B6 increases both the selectivity and the separation efficiency. The optimal concentration, with 3% (w/v) B10E270B10 and 5% (w/v) B6E46B6, is determined with the consideration of both speed and resolution. A resolution of 1.3 was achieved on the separation of 123/124 base pairs in the pBR322/HaeIII digest within 20 min by using a 10 cm column of 75 microm I.D., demonstrating the potential use of mixtures of amphiphilic block copolymers as an effective DNA separation medium. PMID- 11269527 TI - Restricted access chromatographic sample preparation of low mass proteins expressed in human fibroblast cells for proteomics analysis. AB - Two-dimensional electrophoresis and modern image analysis systems have made it possible to study protein expression and regulation of proteins in biological systems. Proteins in the molecular mass region of 20-120 kDa are well investigated and described. However, proteins with masses below 20 kDa are the least investigated as they are rarely seen on 2D-PAGE due to fast migrations in the electric field and lack of staining efficiency. This paper describes a technique that enriches proteins in the lower mass region using solid-phase extraction. The purification step is carried out using C18 functionalised "restricted access" affinity chromatography whereby simultaneous trace enrichment and sample clean up is achieved. In this study expression patterns of TGF-beta stimulated and non-stimulated fibroblasts were compared after the solid-phase fractionation procedure. An increased expression pattern was obtained whereby 400 protein spots could be detected by image analysis in the <20-kDa region. Out of these, specific regulations of 14 spots were found by quantitative image analysis and spots of interest were identified with MALDI TOF-MS. The regulated and identified proteins were triosephosphate isomerase, cofilin and heat shock 27-kDa protein. PMID- 11269528 TI - Ion-exchange-based eluent-free preconcentration of some anions. AB - Preconcentration procedures based on ion-exchange methods are often used to enhance the sensitivities of analytical techniques where the eluent used for eluting the preconcentrated ions does not influence the subsequent analytical step. Until recently, only a limited use of ion-exchange-based sample preconcentration procedures has been found in those analytical techniques where the eluent components strongly influence the separation procedure [e.g., capillary electrophoresis (CE)]. In this paper, we present a preconcentration procedure based on (i) the preconcentration of anions on an ion-exchange resin, (ii) the subsequent elution of analytes, and (iii) on-line removal of eluent components by chemical suppression using an appropriate suppressor device (either packed-bed suppressor column or micromembrane suppressor). The adjustment of the system parameters, combined with a computer-controlled, sensing/switching system, resulted in a minimal additional dilution of the eluted preconcentrated anions. The efficiency of the proposed enrichment/matrix removal procedure was tested by using off-line CE analysis of collected preconcentrated samples, reaching a LOD of 1 microg/l for a selected anion. PMID- 11269529 TI - Separation of kaempferols in Impatients balsamina flowers by capillary electrophoresis with electrochemical detection. AB - Capillary electrophoresis with wall-jet amperometric detection was used to detect kaempferol and its derivatives kaempferol-3-glucoside, kaempferol-3 glucosylrhamnoside and kaempferol-3-(p-coumaroyl)glucoside. The influence of buffer pH on separation was investigated and optimized. With a phosphate buffer at pH 7.5, nearly complete separation of the four kaempferols was achieved according to their different electrophoretic mobilities. The detection potential was also evaluated and optimized. At detection potential of +0.80 V vs. saturated calomel electrode, an amperometric response with high sensitivity and stability was obtained for these four compounds. Detection limit estimated for all the kaempferols examined was less than 1.4 fmol, based on S/N=3. The use of this method for the separation and detection of these compounds present in balsam flowers (Impatiens balsamina) is reported. PMID- 11269530 TI - Enantiomeric separations of primary amino compounds by capillary electrochromatography with monolithic chiral stationary phases of chiral crown ether-bonded negatively charged polyacrylamide gels. AB - A novel enantiomeric separation method by capillary electrochromatography with chiral crown ether-bonded negatively charged polyacrylamide gels is presented. Two kinds of chiral crown ether derivatives, (+)-tetraallyl 18-crown-6 carboxylate and (+)-18-crown-6 tetracarboxylic acid 2-allyl ester were synthesized and allowed to covalently bind to a negatively charged polyacrylamide gel, a so-called monolithic stationary phase, respectively. The gel was placed in fused-silica tubing, the walls of which had been activated with a bifunctional reagent to make the resulting gel bind covalently to the inner surface. Enantiomeric separations of 12 primary amino compounds were achieved using these columns and mobile phases of 200 mM triethanolamine-300 mM boric acid buffers with high efficiencies of up to 135000 plates m(-1). Both the within- and between run reproducibilities of retention time and separation factor were good. The reproducibilities of retention time and separation factor for three different columns prepared from a different batch of monomers were acceptable. The gel filled capillaries were stable for at least 13 months with intermittent use for 3 months followed by storage at room temperature for 10 months. The result of the optical purity test of alanine-2-naphthylamide is also described. PMID- 11269531 TI - Clinical significance of low titer anti-nuclear antibodies in early rheumatoid arthritis: implications on the presentation and long-term course of the disease. AB - The objective of this study was to evaluate the clinical significance of anti nuclear antibodies (ANA) detected in the early stages of rheumatoid arthritis (RA), by a retrospective comparison of the clinical, laboratory, and therapeutic characteristics of patients with or without ANA. The files of 99 longstanding seropositive RA patients were reviewed. Data relating to demographics, medical history, family history, physical findings, extra-articular complications, laboratory tests, drugs [dosage, duration. efficacy, combinations, adverse effects (AEs)], intra-articular injections, and surgery were recorded. Patients with or without ANA at presentation of their disease were compared using chi square and t-tests. Fifty-two ANA positive (group 1) and 47 ANA negative (group 2) patients were enrolled in the study. All were comparable in terms of their mean age, age at diagnosis, follow-up duration (approximately 10.5 years), and male:female (M:F) ratio. On admission, pain complaints were more pronounced in group 1 (P = 0.004 in the feet), but the physical findings did not differ. Deformities and nodules developed in similar numbers. Extra-articular complications were evenly distributed; vasculitis, however, was significantly more prevalent in ANA positive (10/52) than in ANA negative (2/47) patients. Thyroid disease was more common in group 2 (10/47 vs 3/52). Laboratory tests (presentation and maximal values) were similar, with the exception of higher anti DNA (but within normal ranges) and gamma-globulin% in group 1. Group 1 used more drugs prior to diagnosis. Corticosteroids and disease-modifying anti-rheumatic drugs (DMARDs) were evenly used. Combination therapy, joint injections, and surgery were more prevalent in group 2. AEs to various DMARDs were more common in group 1. Although similar in many aspects, RA patients with ANA tend to present with more pain complaints, a higher risk of vasculitis and AEs relating to use of DMARDs, while those without ANA needed more aggressive therapeutic modalities. PMID- 11269533 TI - The role of quantitative ultrasound in predicting osteoporosis defined by dual X ray absorptiometry. AB - The aim of this study was to establish whether quantitative ultrasound (QUS) parameters could identify patients classified as osteoporotic and osteopenic on the basis of dual energy X-ray absorptiometry (DEXA). One hundred and twenty three patients (39 male, 84 female) with osteoporosis and suspected of having osteoporosis were included in this study. Broadband ultrasound attenuation (BUA) and speed of sound (SOS) were measured and bone mineral densities (BMD) of the lumbar spine and left hip was measured by DEXA. Subjects were classified into three groups (normal, osteopenic and osteoporotic) on the basis of BMD T-scores measured by DEXA. QUS parameters of the osteoporotic group were significantly lower than those of osteopenic and normal groups; there was no difference in QUS parameters between the normal and osteopenic groups. Correlations of both right and left SOS and BUA with the spine and femoral neck BMD were moderate (r = 0.343 0.539, P < 0.001). There was also reasonable correlation between DEXA and QUS T scores (r = 0.364-0.510, P < 0.001). QUS had a sensitivity of 21% and a specificity of 95% for diagnosing osteoporosis. We concluded that, although DEXA and QUS parameters were significantly correlated, QUS parameters can not predict osteopenia as defined by DEXA, and sensitivities and specificities of QUS parameters were not sufficiently high for QUS to be used as an alternative to DEXA. PMID- 11269532 TI - Differential in vitro effects of IL-4, IL-10, and IL-13 on proinflammatory cytokine production and fibroblast proliferation in rheumatoid synovium. AB - The purpose of this study was to compare the potential of interleukin-4 (IL-4), IL-10, and IL-13 to interrupt two major inflammatory pathways in rheumatoid arthritis (RA), i.e., overexpression of proinflammatory cytokines and cytokine mediated fibroblast growth. IL-4, IL-10, and IL-13 were all able to significantly inhibit the production of IL-1beta, tumor necrosis factor-alpha (TNF-alpha), IL 6, and IL-8 by freshly isolated RA synovial tissue cells, IL-10 was most effective in terms of IL-1beta and TNF-alpha reduction. The IL-1 receptor antagonist was enhanced by IL-4 and IL-13, but only slightly enhanced by IL-10. Spontaneous interferon-gamma secretion was diminished by IL-4 and IL-10 but not by IL-13. Addition of anti-IL-10 neutralizing antibody to RA synovial tissue cells resulted in a substantial increase in IL-1beta and TNF-alpha levels, whereas neither anti-IL-4 nor anti-IL-13 antibody had a significant effect. IL 1beta-stimulated proliferation of RA synovial fibroblast cell lines was inhibited by IL-4 and IL-13, but not by IL-10; IL-4 was over tenfold more effective than IL 13. These results suggest that IL-4, IL-10, and IL-13 all have the therapeutic potential to regulate the disease activity mediated by proinflammatory cytokines in RA, but each cytokine may have different potencies. PMID- 11269534 TI - Hyperuricemia in Saudi Arabia. AB - The objective of this prospective study was to determine the prevalence of hyperuricemia and gout in a sample of Saudi individuals and their relationship to certain risk factors, namely, obesity, serum glucose, triglycerides, cholesterol, age, and sex. A total of 487 Saudis (250 males and 237 females) from 14 primary care clinics were interviewed, examined, and investigated. The mean age for the males was 46.89 +/- 17.01 years (range 14-83) and for the females 45.08 +/- 13.67 years (range 21-80). Serum uric acid (SUA) values above 420 micromol/l for males and 360 micromol/l for females were considered to be high. Of the 487 individuals, 41 (8.42%; 20 males and 21 females) had hyperuricemia. The mean SUA was 308.41 +/- 90.64 micromol/l for males and 254.59 +/- 85.79 micromol/l for females. In females, uric acid levels correlated significantly with age, body mass index (BMI), serum creatinine, and the erythrocyte sedimentation rate (ESR), but not with serum cholesterol or triglycerides. In males, uric acid levels only correlated significantly with BMI and serum creatinine. No case of gout was found. PMID- 11269535 TI - Alendronate in rheumatoid arthritis patients treated with methotrexate and glucocorticoids. AB - Rheumatoid arthritis (RA) is a systemic inflammatory disease. Along with synovial joint inflammation, extra-articular involvement is a common feature of RA. Periarticular and generalized osteoporosis are seen both as an extra-articular feature of the disease itself and due to various medications like glucocorticoids and methotrexate (MTX). In this study, we investigated the effects of oral alendronate in RA patients treated with MTX and prednisolone by comparing the effects of "alendronate+calcium" and "only calcium" on bone mineral density (BMD). Fifty RA patients classified according to American Rheumatism Association (ARA) criteria were included in the study. The control group consisted of 20 postmenopausal osteoporotic patients. The RA patients were divided randomly into two groups. All patients were started on MTX 7.5 mg/week, 2.5-mg daily folic acid, and 7.5-mg daily prednisolone. The first group, consisting of 25 female RA patients, was also given 10-mg daily alendronate and 1000-mg daily calcium. The second group also consisted of 25 female patients and was given only 1000-mg calcium per day. The postmenopausal control group was given daily 10-mg alendronate and 1000-mg calcium. Bone mineral densities were measured by dual energy x-ray absorptiometry (DEXA) and again at the end of the sixth month. At the end of the study, RA patients given only calcium had reduced mean BMD, and patients treated with alendronate and calcium showed increased mean BMD almost in all regions. This increase was significant in the L2 and L1-4 total regions. In postmenopausal osteoporotic patients, we saw statistically significant increases in BMD in all regions. The increase in BMD values in RA patients treated with alendronate was smaller than in those of the control group of postmenopausal osteoporosis patients. In conclusion, RA itself has a risk factor for osteoporosis in addition to the risks of the medications like corticosteroids and MTX. In the prevention and treatment of RA-associated osteoporosis, alendronate and calcium therapy is effective and well tolerated. PMID- 11269536 TI - Scintigraphic evaluation of synovial inflammation in rheumatoid arthritis with (99m)technetium-labelled human polyclonal immunoglobulin G. AB - The aim of this study was to investigate whether (99m)technetium-labelled polyclonal human immunoglobulin G (99mTc-IgG) scintigraphy reflects synovial inflammation in patients with rheumatoid arthritis (RA). We evaluated 29 patients with RA for this reason and found a highly significant correlation between total scintigraphic scores and total tenderness scores (r = 0.781, P < 0.001). A significant correlation was also found between 99mTc-IgG scintigraphic scores and tenderness in all joints other than the shoulders. The 99mTc-IgG scintigraphy had a sensitivity of 69% and specificity of 88% in cases with tenderness and 72% and 81%, respectively, in cases with swelling. Total scintigraphic scores were correlated with serum levels of C-reactive protein (r = 0.401, P < 0.05) but not with erythrocyte sedimentation rate (r = 0.149, P > 0.05). The correlation between disease activity scores and total scintigraphic scores was also found to be significant (r = 0.812, P < 0.001). We suggest that 99mTc-IgG scintigraphy is a reliable and objective method in detecting synovial activity and can be appropriate for observing disease prognosis in clinical trials with RA. PMID- 11269537 TI - Progression of primary APS (Hughes syndrome) into serological SLE: case report. AB - An Arab woman presented with a history of multiple foetal losses and spontaneous venous thromboembolism, which recurred on several occasions. The presence of antiphospholipid antibodies in the absence of other clinical and serological features of systemic lupus erythematosus (SLE), including negative antinuclear antibodies (ANA), confirmed the diagnosis of primary antiphospholipid syndrome (PAPS). More than 15 years after the beginning of clinical events and 10 years after diagnosis, she progressed into the immunological domain of SLE without concurrent clinical features. The patient exhibited weakly positive ANA of a speckled pattern, strongly positive anti (ds) DNA antibodies and false positive VDRL. Lymphopenia has not been observed at any stage of the follow-up. Although the evolution of PAPS into SLE has been infrequently reported, this seems to be another case suggesting that PAPS in some patients may be an early manifestation of lupus. PMID- 11269538 TI - Primary lung cancer associated with polymyositis/dermatomyositis, with a review of the literature. AB - It has been suggested that lung cancer is frequently associated with polymyositis/dermatomyositis (PM/DM). The purpose of this study was to describe the clinical features of primary lung cancer associated with PM/DM. We first describe the clinical features of two cases treated in our hospital, and then provide a review of the literature. Finally, 24 patients (five females and 19 males) with primary lung cancer associated with PM/DM are retrospectively evaluated. Histological types of lung cancer were as follows: small cell lung cancer (n = 7), squamous cell carcinoma (n = 5), adenocarcinoma (n = 2), others (n = 5), and unknown (4). The onset of PM/DM is frequently observed before the detection of lung cancer. This is the first report to describe the clinical features of lung cancer associated with PM/DM. PMID- 11269539 TI - Results of arthroscopic treatment of symptomatic loss of extension following anterior cruciate ligament reconstruction. AB - Symptomatic loss of knee extension is an important cause of postoperative morbidity following anterior cruciate ligament reconstruction. In a series of 342 consecutive reconstructions performed by the senior author, 17 knees in 16 patients had symptomatic extension deficits (>5 degrees) refractory to a minimum of 4 months of intensive physical therapy that required arthroscopic debridement. Thirteen knees in 12 patients were available for evaluation at a mean follow-up of 3.9+/-1.7 years and form the treatment group. Twenty-six knees in 26 patients who underwent reconstruction but did not develop arthrofibrosis were matched to the treatment group and served as controls. At a mean of 12+/-8 months following reconstruction, patients in the treatment group underwent examination under anesthesia, arthroscopic debridement, revision notchplasty as necessary, and controlled manipulation. Postoperatively, patients were assigned to a closely supervised rehabilitation protocol emphasizing restoration of knee extension. At final evaluation, knee extension deficits had improved from a preoperative mean of 10 degrees (SD 5 degrees) to 3 degrees (SD 4 degrees) (P<.001). Multiple functional rating scales also were used to evaluate the treatment and control groups. With the numbers available, there was no statistically significant difference in function at final evaluation between the treatment and control groups. The best treatment for loss of knee extension is preventive. Complications are avoided by careful patient selection, appropriate timing of surgery, attention to operative detail, and aggressive rehabilitation. However, patients reaching a plateau in rehabilitation with significant residual extension deficits, patellofemoral symptoms, or both predictably benefit from arthroscopic debridement. PMID- 11269540 TI - The effect of tubularization on the mechanical properties of patellar tendon grafts. AB - To determine the effect of tubularization on the prefailure mechanical properties of bone-patellar tendon-bone autografts used for anterior cruciate ligament repair, 10 bovine bone-patellar tendon-bone grafts were tested in tension before and after tubularization with running suture. The testing protocol involved a 5-N preload, 10 preconditioning cycles to 200 N, and a final test cycle to 950 N at 1000 N/sec. Five of the grafts were tested first as harvested (flat) and then again following tubularization. The remaining five grafts were tubularized prior to the initial testing, and final testing was done with the suture removed. Raw testing data were reduced to determine the amount of stretching associated with preconditioning, as well as laxity and stiffness of the preconditioned grafts. Tubularized grafts stretched significantly more than flat grafts during preconditioning: 3.5 times as much after the first preconditioning cycle (3.8+/ 1.9 mm versus 1.1+/-0.78 mm) and 3.1 times as much after 10 cycles (5.0+/-2.1 mm versus 1.6+/-0.9 mm). There was no statistically significant difference in the stiffnesses of the tubularized and flat grafts, nor did tubularization have an effect on graft laxity. Interestingly, there was a slight increase in laxity the second time each graft was tested, regardless of whether the graft was flat or tubularized when it was first tested. These results highlight the importance of preconditioning patellar tendon grafts before fixation, especially those that have been tubularized. PMID- 11269541 TI - Radiographic analysis of femoral tunnel position in anterior cruciate ligament reconstruction. AB - Successful reconstruction of the anterior cruciate ligament (ACL) depends on anatomic placement of a graft ligament substitute. This study examined the accuracy of a plain radiograph in determining femoral tunnel position during ACL reconstruction. Nine cadaveric distal femurs had six separate tunnels made in each specimen: 12:00 (high position), 1:30 (anatomic position), and 3:00 (low position) in the left femora and 12:00 (high), 10:30 (anatomic), and 9:00 (low) in the right femora. At each position on the clock face, two 9-mm tunnels were drilled, leaving 2 mm (correct) and 12 mm (incorrect) of posterior wall intact. With a radiopaque tunnel dilator in each tunnel, a true lateral radiograph, a 10 degree externally rotated lateral radiograph, and a 10 degree internally rotated lateral radiograph were obtained. All radiographs were analyzed for femoral tunnel placement in the anteroposterior plane with the four-quadrant method described by Harner et al and the ratio method described by Aglietti et al. Statistically significant differences could only be distinguished between anatomic (10:30), anterior (12-mm rim), and posterior (2-mm rim) positions. There were no statistically significant differences for any of the other positions when comparing true laterals to true laterals, true laterals to internal or external oblique views, or when comparing internal and external oblique views. A malpositioned anterior tunnel (12-mm rim posterior), which was "low" at 9:00 or "high" at 12:00 in the notch (malplaced), could not be distinguished reliably from an anatomically correct placed tunnel with a single-plane lateral radiograph. PMID- 11269542 TI - Outcome results after total knee arthroplasty: does the patient's physical and mental health improve? AB - An ongoing prospective study with continuing enrollment and follow-up of 279 patients analyzed the postoperative mental and physical outcomes on an interim basis using the SF-36 Physical and Mental Health Summary Scale, which allows for patient self-assessment of total knee arthroplasty (TKA) outcome. Results of this study support the hypothesis that patients can assess their own physical health after surgery and suggest that while physical health component scores increased dramatically subsequent to TKA, mental health component scores remained essentially unchanged. The lack of significant improvement of the mental health component score can be explained in part by noting that the mean mental health component score for the study patient cohort was already above average preoperatively. Thus, mental health component scores did not correlate with the dramatic improvement of physical health component scores. Physical health component score appears to be a more accurate indicator of the benefits of TKA than mental health component score. PMID- 11269544 TI - Articular cartilage damage, peripheral migration, and device failure as meniscal arrow complications: case report. PMID- 11269543 TI - Asymmetric patella resurfacing in total knee arthroplasty. AB - Three hundred consecutive primary, cemented, condylar total knee arthroplasties (TKAs) were reviewed for the presence of asymmetric patella resurfacing using a postoperative Merchant or sunrise patellar radiograph. Twenty-one knees in 14 patients were found to have the patella asymmetrically resurfaced. Asymmetric resurfacing typically involved the inadvertent preferential resurfacing of the lateral facet with underresection of bone from the medial patellar facet. All patients underwent follow-up for a minimum of 5 years, with a mean follow-up of 7.5 years. Of the 21 knees, 3 revisions were required for patellar complications. One patellar component was loose on radiographs and there was marked patellofemoral pain in 6 knees. Overall, 11 of 21 knees (52%) underwent revision or were recommended for revision for patellar complications or had anterior knee pain that limited activities. Inadvertent asymmetric patella resurfacing using the kinematic condylar implant adversely affects the outcome after TKA. PMID- 11269545 TI - Multifocal osteonecrosis of the knee. PMID- 11269546 TI - Building a successful practice-owned, office-based ambulatory surgery center. AB - If the orthopedic practice is able to avoid the pitfalls noted above, the practice-owned, office-based ASC can be an ideal venue for retrieving services back into the orthopedic practice. These services have been outsourced to the hospitals and multispecialty ASCs for many decades. These practice-owned, office based facilities provide significant enjoyment, convenience, efficiency, and in most cases profitability for many practices throughout the country. Few, if any, new orthopedic practice facilities are being designed today without strong consideration of a practice-owned, office-based ASC. PMID- 11269547 TI - Pros and cons of practice-owned and office-based ambulatory surgery centers. AB - A detailed feasibility analysis is imperative to ensure the success of a practice owned ASC. Analysis of the payer mix and the market relating to surgical volume that can be performed at the ASC is imperative. If overbuilding, overequipping, and overstaffing are avoided and the group has adequate volume that can be managed at the ASC, the facility should be a success. Building a practice-owned ASC without an accurate and detailed financial feasibility and payer study can place the endeavor at risk. A well-planned, economically constructed and properly managed ASC will result in an efficient and successful ancillary service for the orthopedic group practice. PMID- 11269548 TI - Laparoscopic surgery in hepatic hydatid cysts: a technical improvement. AB - Hepatic hydatid cysts are a common surgical problem that is encountered in many tropical countries, including India. Open surgical exploration and excision had been the mainstay of treatment until the advent of laparoscopy. In 1998, we successfully managed six cases of large hepatic hydatid cysts using the videoendoscope, with excellent postoperative follow-up results. Four men and two women participated in this study, with patient ages ranging from 28 to 42 years. The duration of the disease ranged from 1.3 to 2.8 years. All patients had undergone preoperative albendazole therapy for more than 2 months. Complete evacuation of the cyst contents, including all daughter cysts and laminated membrane, along with a subtotal excision of the extrahepatic part of the cyst wall, was accomplished without any spillage into the peritoneal cavity. The saucerized cavity was drained. The drains were removed 6 to 9.4 days after a check ultrasound. Postoperative follow-up ranged from 3 to 9 months and revealed no evidence of a recurrence in the abdomen. It is possible with carefully planned placement of trocars to completely eliminate the risk of spillage, and therefore not compromise the standard principles of hydatid surgery. PMID- 11269549 TI - Laparoscopic repair of colonic perforation associated with colonoscopy: use of passing sutures and endoscopic linear stapler. AB - Laparoscopic surgery recently has been conducted to repair colonic perforation that is associated with colonoscopy. The authors describe their laparoscopic repair of perforation using passing sutures and an endoscopic linear stapler. One 12-mm and several 5-mm trocars were inserted in the lower abdomen under general anesthesia. Observing with a laparoscope, passing sutures were threaded transversely through all layers of the margin of defect and pulled up with forceps to hold the margin straight, along which the defect was stapled with an endoscopic linear stapler. The authors applied this method for five patients, where the perforation occurred in the sigmoid colon or in the cecum (perforation size ranging from 10 mm to 50 mm). Perforation was successfully repaired in all patients, with no complications because of perforation or the procedures. The current method is beneficial because the perforated lesion is safely and easily closed and postoperative colonic stenosis is avoided. PMID- 11269550 TI - Surgical management of peptic ulcer disease in the Helicobacter era--management of bleeding peptic ulcer. AB - Bleeding continues to be a significant cause of morbidity and mortality for patients with peptic ulcer disease. Recent advances have changed the management of this disease. Upper endoscopy with or without endoscopic therapy is the preferred procedure during the initial evaluation of upper gastrointestinal bleeding. With its excellent success rates, many patients are being cured with endoscopic therapy followed by eradication of Helicobacter pylori. H. pylori is now thought to have an important role in the pathogenesis of a majority of gastric and duodenal ulcers. This finding has led to the recommendation that patients with peptic ulcer disease be treated with regimens effective against this organism. Currently, patients who are older and who have more severe underlying medical conditions present a challenge. This review will address the options for treatment of peptic ulcer bleeding. In addition, knowledge gained regarding H. pylori infection and use of nonsteroidal anti-inflammatory drugs will be discussed. PMID- 11269551 TI - Laparoscopic appendectomy in children: evaluation of different techniques. AB - Patients and surgeons frequently opt for laparoscopic appendectomy for treatment of acute appendicitis. Clinical studies have shown this approach to be a reasonable alternative to open appendectomy. The objective of the current study was to assess the outcome of laparoscopic appendectomy using three different techniques. The study sample consisted of 150 children with acute appendicitis who underwent surgery at Al-Azhar University Hospitals, Cairo, Egypt, and at Al Mishary Hospital in Riyadh, Saudi Arabia, between October 1997 and October 1999. The patients were allocated to undergo extracorporeal laparoscopic appendectomy, Endoloop laparoscopic appendectomy, or EndoGIA (Ethicon Endo-surgery, Inc., Cincinnati, OH, USA) laparoscopic appendectomy. All patients were assessed for the severity of the disease at baseline using clinical and hematologic indicators. The ages of the children ranged from 7 to 14 years, with a mean of 10 years (SD, 2.14 years). Of the children, 55.3% were female. The results showed that children who underwent laparoscopic appendectomy using the EndoGIA had statistically significant shorter operating times, did not have complications, and had the shortest duration of hospital stay (although duration of hospital stay did not reach the statistically significant level of P > 0.05). Therefore, the study showed that laparoscopic appendectomy using the EndoGIA is the procedure that is most recommended, except for the relatively high cost of the disposable materials. Endoloop laparoscopic technique was the second most preferable procedure, and the least preferred procedure was extracorporeal laparoscopically assisted appendectomy. The major drawback of the last technique is the high frequency of complications. Endoloop laparoscopic appendectomy with a purse-string suture can be performed safely if the EndoGIA is not available. PMID- 11269552 TI - Laparoscopic hernia repair enhances early return of physical work capacity. AB - Several researchers have documented less postoperative pain and a quicker return to daily activities after laparoscopic herniorrhaphy. However, little objective data that validates this hypothesis exists. This study compares the rate of postoperative physical work capacity with return to preoperative levels, which is measured by a standard treadmill test in patients who underwent laparoscopic and conventional open hernia repair. Patients completed a 6-minute walking test preoperatively and 1 week postoperatively using a nonmotorized treadmill. The distance walked was recorded. If the distance that a patient achieved at 1 week was not within 0.02 miles of the preoperative values of the patient, the patient was asked to return at 1 month for repeat testing. Patients were enrolled prospectively in this study from October 1997 to February 1999. Sixty-six patients participated in the study (27 laparoscopic herniorrhaphies and 39 open herniorrhaphies were performed). There was no significant difference in age, body mass index, or preoperative distance achieved among the two groups. At 1 week, patients who underwent laparoscopic repair demonstrated a mean increase of 18 meters from preoperative distance (P = 0.07). In the open group, patients demonstrated a mean decrease of 90 meters at 1 week (P = 0.001). The change in distance at 1 week between the laparoscopic and the open groups was statistically significant (P = 0.001). However, at 1 month, there was no significant difference among the two groups. Measured using treadmill walking, laparoscopic hernia repair seems to offer an early advantage to open repair in return-to-physical work capacity. PMID- 11269553 TI - Laparoscopic intraperitoneal onlay polytetrafluoroethylene mesh repair (IPOM) for inguinal hernia during spinal anesthesia in patients with severe medical conditions. AB - In patients with severe pulmonary disease, laparoscopic techniques are not advised. The authors report their preliminary experience with laparoscopic intraperitoneal onlay polytetrafluoroethylene mesh repair for inguinal hernia during spinal anesthesia in patients with chronic obstructive pulmonary disease. Spinal anesthesia was performed using hyperbaric bupivacaine (3-3.5 mL) injected at L2-L3. If necessary, additive opioid therapy was administered. Under low pressure pneumoperitoneum (10 mm Hg), polytetrafluoroethylene mesh was stapled securely on the posterior inguinal wall to spare epigastric and iliac vessels. Fifteen patients underwent surgery. Median age was 62 years. All patients were classified American Society of Anesthesiologists physical status III/IV. Mean forced expiratory volume in the first second was 1.1 L/s. Median operating time was 20 minutes. Postoperative recovery was uneventful for all patients. The average duration of hospital stay was 1.5 days. Seroma or hematoma was not noted. Six-month follow-up did not show recurrence or infection. This technique is an effective method of repair of inguinal hernia in patients with severe chronic obstructive pulmonary disease, and it provides a maximum of comfort. PMID- 11269554 TI - Endoscopic axillary lymphadenectomy without prior liposuction in 100 patients with invasive breast cancer. AB - The purpose of this study was to evaluate intra- and postoperative outcome after endoscopic axillary lymphadenectomy without liposuction. One hundred patients with early stage breast cancer were treated by breast conserving therapy and endoscopic technique. The median duration of operation was 75 minutes (30-130 minutes). Switching from endoscopy to an open technique was necessary for two patients. The median number of removed lymph nodes was 16. Postoperatively, seroma developed in four patients, temporal winged scapula developed in three patients, and a wound infection in one patient. On postoperative day 5, arm mobility was not restricted for 89 patients. After a median follow-up of 14 months, 14 patients reported persistent impairment of sensibility, and two patients did not have full shoulder mobility. Axillary recurrence has not developed in any patient. Endoscopic axillary lymphadenectomy can be performed safely without previous liposuction. PMID- 11269555 TI - Staple-line reinforcement with a new type of polyglycolic acid felt. AB - Although various materials have been used for reinforcement in lung-volume reduction surgery to buttress pulmonary staple-line, absorbable materials are not available for use in thoracoscopic surgery. Moreover, even nonabsorbable types of reinforcements have been used only for lung volume reduction surgery. However, elderly patients with spontaneous pneumothorax secondary to emphysematous lung are well treated with staple-line reinforcement. The authors developed a new type of polyglycolic acid felt to buttress staple-line. This felt is absorbable, easier to cut with a stapler knife than is the conventional polyglycolic acid felt, and inexpensive enough to use for various types of thoracic surgeries for emphysematous lungs in Japan, and it can be attached to staplers with a small amount of fibrin glue. These strips were used to reinforce pulmonary staple lines for resection of emphysematous lungs in 14 patients: pulmonary emphysema (n = 1), bilateral giant bullae (n = 1), ipsilateral giant bullae (n = 6), spontaneous pneumothorax with multiple bullae in an emphysematous lung (n = 5), and lung cancer in a patient with pulmonary emphysema (n = 1). There were no air leaks during surgery. Air leaks were noted in three patients after surgery. In two patients, the air leaks stopped within 2 weeks. In one patient, the air leak was found to originate from an untouched lobe during reoperation. No infection or allergic reaction developed in a patient during a mean follow-up of 12 months (range, 1 to 24 months). PMID- 11269556 TI - Preperitoneal Richter hernia after a laparoscopic gastric bypass. AB - Trocar-site incisional hernias are potentially dangerous because of their susceptibility to become Richter hernias. The authors describe a morbidly obese patient in whom developed an unusual type of Richter hernia after a laparoscopic isolated Roux-en-Y gastric bypass at a 10-mm trocar site. Although the fascial closure of the trocar hernia site was intact, a hernia developed through the peritoneum into the preperitoneal space. For morbidly obese patients, the thick preperitoneum is a potential space that allows for the development of a Richter hernia, despite adequate fascial closure. It is recommended that all 10-mm and 12 mm trocar sites be closed, incorporating the peritoneum into the fascial closure to obliterate the preperitoneal space, to prevent this postoperative complication. PMID- 11269557 TI - Gallstone in abdominal wall--a complication of laparoscopic cholecystectomy. AB - A 39-year-old woman presented with abdominal wall mass 9 years after she underwent laparoscopic cholecystectomy for symptomatic gallstones. After surgical resection, a pathologic examination identified an abscess cavity within the abdominal wall that was surrounded by a wide, diffuse, poorly defined wall of dense fibrous tissue. An examination did not show neoplastic tissue. The cavity was bile-stained and contained a 2.5-cm gallstone. This case shows a complication of laparoscopic cholecystectomy. Gallstones spilled during the extraction of the gallbladder through the abdominal wall incision may lead to a reactive process that clinically and microscopically may resemble a fibro-proliferative disorder, including a neoplastic process. This complication of laparoscopic cholecystectomy is rare. Pathologists must be aware of its occurrence because examination of the solid fibrous wall may lead to diagnoses of reactive or neoplastic fibro proliferative processes. PMID- 11269558 TI - Hand-assisted laparoscopic splenectomy for idiopathic thrombocytopenic purpura during pregnancy. AB - A successful case of a hand-assisted laparoscopic splenectomy with low-pressure pneumoperitoneum for autoimmune thrombocytopenic purpura in a patient at 23 weeks' gestation is reported. Preoperative splenic arterial embolization was performed on the same day as the operation using painless contour embolic material and super-absorbent polymer microspheres. The abdominal wall retraction method first was applied to avoid the effects of pneumoperitoneum on systemic hemodynamic alterations. However, a sufficient surgical view could not be obtained, as the intra-abdominal organs were elevated because of the enlarged uterus. A surgical view with 4 to 6-mm Hg pneumoperitoneum was available for the hand-assisted splenectomy. The postoperative course was uneventful, and the patient vaginally delivered a healthy infant. A hand-assisted laparoscopic splenectomy with low-pressure pneumoperitoneum after splenic arterial embolization would be feasible for patients with autoimmune thrombocytopenic purpura during a relatively advanced pregnancy. PMID- 11269559 TI - Laparoscopic surgery using a newly designed suction lifter. AB - The authors describe a simple and available abdominal wall-lift technique for endoscopic surgery that involves a new suction lifter that they designed for laparoscopic surgery. Since July 1998, the authors have used this technique successfully in eight patients with appendicitis, in one patient with a perforated duodenal ulcer, and in one patient with an adenoma of the ascending colon. PMID- 11269560 TI - Primary segmental infarction of the greater omentum: a rare cause of RLQ syndrome: laparoscopic resection. AB - The authors report a rare case of a patient with a primary segmental infarction of the greater omentum who reported acute abdominal pain. Despite preoperative clinical studies and imaging evaluation, an etiologic diagnosis could not be determined. The diagnosis of this uncommon disease was determined after initial laparoscopic exploration. A laparoscopic resection was performed. The patient had an uneventful recovery and was discharged within 12 hours. The differential diagnosis of the right lower quadrant syndrome includes several disorders, of which the primary segmental infarction of the greater omentum is not frequent. The authors emphasize the usefulness of routine laparoscopic exploration in patients with RLQ syndrome because it adds the possibility of mini-invasive treatment to the initial diagnosis. PMID- 11269561 TI - Laparoscopic ovarian cystectomy using a single umbilical puncture method. AB - To establish a minimally invasive technique to perform ovarian cystectomy, the authors applied a single umbilical puncture method. A 2-month old female infant was admitted to the hospital because of an ovarian cyst that showed no spontaneous shrinkage after her birth. An umbilical semicircular incision was made to insert a 10-mm trocar, into which a 3-mm laparoscope was inserted. The ovarian cyst was grasped using forceps inserted through an operating channel of the scope, and the cyst was removed through the incision. After aspiration of the cyst, the free cyst wall was resected, leaving the intact ovarian tissue. The operation was performed without difficulty or complication. The postoperative course was uneventful. The wound was inconspicuous, and the result was cosmetically excellent. The case demonstrated the feasibility of the minimally invasive technique using a single umbilical puncture for ovarian cystectomy in an infant. PMID- 11269562 TI - Laparoscopic segmental resection for infiltrating endometriosis of rectosigmoid colon: a preliminary report. PMID- 11269563 TI - Laparoscopic versus open vertical banded gastroplasty for the treatment of morbid obesity. AB - Vertical banded gastroplasty (VBG) is an effective treatment for morbid obesity. Recent advancement in laparoscopic surgery has made laparoscopic VBG possible. The authors compared retrospectively the outcomes of laparoscopic VBG versus open VBG in patients with morbid obesity. From June 1998 to April 1999, 100 patients (18 men, 82 women; average age, 32.6 years) underwent laparoscopic VBG, and 40 patients (7 men, 33 women; average age, 28.8 years) underwent conventional open VBG. The two groups were similar regarding sex, age, and body mass index distribution. Mean surgical time, blood loss estimate, duration of postoperative recovery, analgesic usage, complications, and weight reduction were compared among the two groups. Laparoscopic VBG was successful in 99 (99%) of the 100 patients. Mean surgical time was longer in duration for the laparoscopic VBG group than it was for the open VBG group (173 vs. 101 minutes, P < 0.01). The laparoscopic VBG group had earlier flatus passage (1.9 vs. 2.6 days; P < 0.01), less usage of analgesics (meperidine 50 mg/unit; 0.9 vs. 2.3 units; P < 0.01), and a shorter postoperative hospital stay (3.7 vs. 6.0 days; P < 0.01). Estimated blood loss, surgical complication rate, and weight reduction were similar among the two groups. Although laparoscopic VBG required a longer surgical time and was technically more demanding, it resulted in shorter recovery time, less analgesic use, and less severe physical discomfort. The authors' findings show that the two methods were approached safely equally. PMID- 11269564 TI - Comparative safety of bone remodeling agents with a focus on osteoporosis therapies. AB - This article reviews the different treatments currently available for osteoporosis and examines the benefits and adverse events that are associated with each. While emphasizing safety considerations, this review summarizes the following treatments for osteoporosis: calcium supplements, fluoride, hormone replacement therapy, raloxifene, bisphosphonates, salmon calcitonin, and calcitriol. Before prescribing any of these agents, the clinician should review the risk/benefit profile of each drug in the context of the individual patient's history, concomitant diseases, concurrent medications, and general physical condition. PMID- 11269565 TI - Pharmacodynamic modeling of lansoprazole using an indirect irreversible response model. AB - A mechanism-based pharmacokinetic/pharmacodynamic model was used to assess lansoprazole effects on gastric pH. The irreversible inactivation of the H+/K+ ATPase enzyme by lansoprazole controls the secretion rate of H+ ions and gastric pH values. The basal circadian rhythm of gastric acid production was taken into account as well as the effects of food intake. A model was applied to multiple dose data from a crossover study of four dosage regimens of lansoprazole in two groups of normal male subjects. Model parameters were estimated by nonlinear regression and were compared to historical values reported in the literature. The predicted mean gastric ion concentration was 23.2 mM (pH 1.6) with the peak time at 12.6 hours (8:30 p.m.), and the half-time for H+ removal from the stomach averaged 1.7 hours. The estimated half-life of gastric food removal was 0.8 hours. The rate constant for normal H+/K+-ATPase degradation was 0.045 h(-1). The pharmacodynamic parameter describing lansoprazole action on gastric acid secretion was the second-order enzyme inactivation constant, which averaged 0.16 microg(-1) x L x h(-1). The parameters obtained for both the baseline and drug treatment data were consistent with the literature and physiologically relevant with the exception of effective food volume, which was large presumably due to buffer effects. The model successfully incorporated the physiological regulation of gastric acid production, the effects of food on gastric acid, and the effects of multiple-dosing regimens of lansoprazole on gastric acid production to give reasonable profiles of gastric pH. PMID- 11269566 TI - Dichloroacetate: population pharmacokinetics with a pharmacodynamic sequential link model. AB - Dichloroacetate (DCA) is a small molecule that reduces ambient concentrations of lactate in man. It was the purpose of this study to develop pharmacokinetic and pharmacodynamic models for determination of a dose for a pivotal Phase III clinical trial of DCA in patients with traumatic brain injury (TBI). Population pharmacokinetic and pharmacodynamic models were developed for DCA using NONMEM software. The pharmacokinetic data were fit to a physiologic two-compartment model, and the pharmacodynamic data were fit to an indirect physiologic response model. Simulations were employed to evaluate various dosing strategies for consideration in a pivotal Phase III clinical trial of DCA. For the pharmacokinetic model, it was discovered that the clearance of DCA decreased on multiple dosing from 4.82 L/h to 1.07 L/h and that the pharmacokinetics and pharmacodynamics in TBI patients could not be predicted from normal volunteers. Population pharmacokinetic modeling and simulation of the expected effects of several dosing strategies were useful procedures for designing a Phase III trial. PMID- 11269567 TI - Pharmacokinetics of mycophenolic acid after mycophenolate mofetil administration in liver transplant patients treated with tacrolimus. AB - The pharmacokinetics of mycophenolic acid (MPA) was studied after oral administration of mycophenolate mofetil (MMF) in 8 liver transplant patients. The mean (+/- SD) maximum MPA plasma concentration of 10.6 (+/- 7.5) mg/ml was achieved within 0.5 to 5 hours. The mean (+/- SD) steady-state area under the plasma concentration versus time curve (AUC(0-12)) was 40 (+/- 30.9) mg/ml/h. The mean (+/- SD) half-life was 5.8 (+/- 3.8) hours. There was poor correlation between trough blood concentrations of tacrolimus (r = -0.004) or serum creatinine (r = 0.689) with MPA AUC, while the serum bilirubin concentrations correlated (r = 0.743) well with MPA AUC, suggesting impairment in MPA conjugation in patients with liver dysfunction. The mean (+/- SD) ratio of the AUC of mycophenolic acid glucuronide (MPAG) to MPA was 64 (+/- 84), which correlated significantly with serum creatinine (r = 0.72) but not with serum bilirubin concentrations (r = 0.309), indicating accumulation of MPAG in patients with renal dysfunction. In 7 primary liver transplant patients on the same dose of MMF, the trough plasma concentrations of MPA during the first week of therapy ranged from < 0.3 to 1.5 microg/ml. The MPA concentrations increased by several folds during the next few weeks, which correlates well with increases in serum albumin concentrations. Changes in albumin appear to partially contribute to the variations in the pharmacokinetics of MPA in liver transplant patients. PMID- 11269568 TI - Abacavir/lamivudine/zidovudine as a combined formulation tablet: bioequivalence compared with each component administered concurrently and the effect of food on absorption. AB - A single-center, open-label, three-way crossover study was conducted in 24 healthy subjects to assess (1) the bioequivalence of a combined abacavir 300 mg/lamivudine 150 mg/zidovudine 300 mg (A/L/Z) combination tablet relative to the separate brand-name components administered simultaneously and (2) the effect of food on the bioavailability of the drugs from the combination tablet. The subjects were randomly assigned to receive each of the following three treatments, separated by a 2-day washout period: one A/L/Z combination tablet after an overnight fast, one abacavir 300 mg tablet + one lamivudine 150 mg tablet + one zidovudine 300 mg tablet sequentially after an overnight fast, or one A/L/Z combination tablet 5 minutes after completing a standardized high-fat breakfast (67 g fat, 58 g carbohydrate, and 33 g protein). Serial blood samples were collected up to 24 hours postdose for determination of abacavir, lamivudine, and zidovudine serum concentrations. Standard pharmacokinetic parameters were estimated. Treatments were considered bioequivalent if 90% confidence intervals (CI) for geometric least squares (GLS) mean ratios for abacavir, lamivudine, and zidovudine area under the serum concentration-time curve (AUC(infinity)) and maximum observed serum concentration (Cmax) fell entirely within 0.80 to 1.25 for log-transformed parameters. The combined A/L/Z tablet was bioequivalent in the extent (AUC) and rate of absorption (Cmax and time of Cmax [tmax]) to the individual brand-name drug components administered concurrently under fasted conditions. GLS ratios and 90% CI for AUC(infinity) and Cmax were 0.99 (0.96, 1.03) and 1.00 (0.90, 1.11), respectively, for abacavir; 0.95 (0.91, 0.99) and 0.90 (0.84, 0.99), respectively, for lamivudine; and 0.95 (0.89, 1.02) and 0.96 (0.80, 1.15), respectively, for zidovudine. The extent of absorption of abacavir, lamivudine, and zidovudine from the combination tablet was not altered by administration with meals, indicating that this formulation may be administered with or without food. However, food slowed the rate of absorption, delayed the tmax, and reduced the Cmax of abacavir, lamivudine, and zidovudine. These changes, which were consistent with those observed with the individual reference formulations when administered with food, were not considered clinically important. All formulations were well tolerated underfasted and fed conditions. PMID- 11269569 TI - Effect of time of meal consumption on bioavailability of a single oral 5 mg tacrolimus dose. AB - Tacrolimus (FK506, Prograf) is marketed for the prophylaxis of organ rejection following allogenic liver or kidney transplantation. This study investigated the effect of timing of a standardized breakfast meal on both the rate and extent of tacrolimus absorption following a single 5 mg oral dose. The protocol used a randomized, open-label, four-period, four-treatment, four-sequence crossover design in 16 healthy, nonsmoking, drug-free male subjects between the ages of 22 and 45 years who were within 15% of their ideal body weight. The four treatments were the following: (A) fasting for 10 hours, (B) ingestion 1 hour before breakfast, (C) ingestion immediately following consumption of the breakfast, and (D) ingestion 1.5 hours after beginning consumption of the breakfast. The breakfast, which was consumed over 15 minutes, contained 848 kcal, with 30%, 16%, and 54% of calories derived from fat, protein, and carbohydrate, respectively. Tacrolimus absorption in the fasting state provided the greatest relative bioavailability (p < 0.05 compared with all other three treatments). AUC(0 infinity)) averaged 312, 276, 205, and 203 ng x h/mL for treatments A, B, C and D, respectively. In contradistinction to taking the drug 1 hour prior to a meal, which had a relatively minor impact on the relative extent of absorption (approximately 12%) compared to the fasting state, ingestion of tacrolimus immediately after a meal (treatment C) or 1.5 hours subsequent to a meal (treatment D) had a more pronounced influence. Mean AUC(0-infinity) ratios (fasting to either postmeal treatments) were approximately 1.5, indicating that absorption extent was considerably reduced by ingesting tacrolimus capsules immediately after eating or 1.5 hours thereafter. Absorption was also prolonged following drug ingestion after a meal, as indicated by a mean tmax value in the fasting state of 1.84 hours, relative to 3.41 hours (immediately aftermeal, p = 0.0035) and 3.22 hours (1.5 hours postmeal, p = 0.0094). The only discernable difference in parameters between treatments C and D was with Cmax, with values of 7.19 and 9.04 ng/mL, respectively, but was not statistically significantly different (p = 0.231). Based on these results and those from a prior study, it is recommended that under therapeutic conditions, oral tacrolimus be administered in a consistent manner, both with respect to the type of meal as well as timing of ingestion relative to consumption of the meal. PMID- 11269570 TI - Food increases the bioavailability of tolterodine but not effective exposure. AB - The objective of this study was to investigate the influence of food on the pharmacokinetics of tolterodine, its active 5-hydroxymethyl metabolite (5-HM), and exposure to the active moiety (sum of unbound tolterodine + 5-HM) in healthy volunteers. Serum concentrations of tolterodine and 5-HM were measured for up to 12 hours after a single oral dose (2 mg) of tolterodine L-tartrate, administered either on an empty stomach or with a standardized medium-fat breakfast. All 23 subjects completing the study were classified as extensive metabolizers (phenotyped with debrisoquine). Pharmacokinetic data on tolterodine and the active moiety were evaluable for 22 subjects; all completing subjects were evaluable for 5-HM pharmacokinetics. Based on Cmax and AUC(infinity) ratios, relative bioavailability of tolterodine in the presence of food was 1.49 (90% confidence interval [CI], 1.35-1.71) and 1.53 (1.35-1.72), respectively. The pharmacokinetics of 5-HM and the active moiety were unaffected by food, as were the rates of drug absorption and terminal half-lives of tolterodine and 5-HM. Given that bioequivalence was observed for the active moiety underfed and fasting conditions, the authors concluded that coadministration of tolterodine with food is not expected to have any clinically relevant effects. PMID- 11269572 TI - Effect of grapefruit juice on pharmacokinetics of microemulsion cyclosporine in African American subjects compared with Caucasian subjects: does ethnic difference matter? AB - This study aims to determine the effect of grapefruit juice (GJ) on microemulsion cyclosporine (CsA) in 11 African American subjects, and it was compared to those in 11 Caucasian subjects. Each subject received two oral doses of CsA with water (W) or GI as well as i.v. CsA. Regardless of race, GJ significantly increased the peak concentration (Cmax) and area under the time-curve (AUC) of CsA; however, the magnitude of GJ effects was different between African American subjects and Caucasian subjects (p = 0.0003). GJ increased peak concentration of CsA by 39% in African American subjects, while the difference in Caucasian subjects was only 8% (p > 0.05). GJ also increased AUC of CsA in African American subjects by 60%, while GJ increased that in Caucasian subjects by 44% (p = 0.0001). The absolute bioavailability of CsA was 21% lower in African American subjects compared with Caucasian subjects when it was given with water (p = 0.048), but these differences disappeared when it was given with GJ (p = 0.6). These findings suggest that concurrent administration of GJ increases the bioavailability of CsA in African American subjects in greater magnitude compared with Caucasian subjects. PMID- 11269571 TI - Lack of correlation between in vitro inhibition of CYP3A-mediated metabolism by a PPAR-gamma agonist and its effect on the clinical pharmacokinetics of midazolam, an in vivo probe of CYP3A activity. AB - RG 12525 (2-[[4-[[2-(1H-tetrazole-5-ylmethyl)phenyl]methoxy]phenoxy]methyl] quinolone) is a novel peroxisome proliferator-activated receptor gamma (PPAR gamma) agonist. In vitro microsomal inhibition assays indicated that RG 12525 is a potent inhibitor of CYP3A4, with a Ki value of 0.5 microM. With the conservative assumption that the total plasma concentration of drug was available to metabolic enzymes following RG 12525 oral administration, marked inhibition of CYP3A4 was expected to substantially reduce the systemic clearance of compounds metabolized by this enzyme. The possibility also existed for inhibition of intestinal and hepatic CYP3A4 by RG 12525 to reduce "first-pass" metabolism and increase absolute bioavailability of CYP3A4 substrates orally coadministered. Consequently, an in vivo drug-drug interaction study was performed to evaluate the effects of orally administered RG 12525 on in vivo CYP3A4 activity in healthy male subjects. The pharmacokinetics of oral midazolam, a probe for intestinal and hepatic CYP3A activity, was not influenced by either the low (100 mg qd for 4 days) or high (600 mg qd for4 days) RG 12525 dosing regimen despite the resulting total plasma concentrations of inhibitor that were well above in vitro Ki values. The point estimates and 90% confidence intervals for the ratios of mean midazolam AUC for subjects administered 100 mg RG 12525 (110.6; 98.7-124.1) and 600 mg RG 12525 (98.4; 84.4-114.7) versus midazolam alone were within 80% to 125%. To explain these results, factors that could limit the accuracy of in vitro models in predicting metabolic drug interactions, mainly the high degree of RG 12525 protein binding (> 99.9%), were considered. The lack of correlation between the in vitro inhibition of CYP3A4 by RG 12525 and the inconsequential effects of this compound on midazolam pharmacokinetics accentuate the need to recognize factors other than plasma drug concentrations and potency of in vitro enzyme inhibition when extrapolating in vitro data to predict in vivo drug-drug interactions. PMID- 11269573 TI - Race but not age affects erythromycin breath test results in older hypertensive men. AB - Erythromycin breath tests (ERBT) were performed to determine age and racial effects on CYP3A4-mediated hepatic clearance in hypertensive men (n = 43) in the clinical setting. Older hypertensive African American men (n = 19: 71 +/- 8 years, mean +/- SD) had faster ERBT clearance compared with Caucasian (n = 20: 72 +/- 6 years) hypertensive men (at 20 minutes after dosing: 0.042 +/- 0.01 percent dose/min exhaled vs. 0.033 +/- 0.013; at 60 minutes after dosing: 0.030 +/- 0.05 vs. 0.023 +/- 0.007 percent dose/min exhaled; ANOVA, p = 0.007), while age, smoking, and reported alcohol intake did not affect ERBT. The data suggest faster hepatic CYP3A-mediated clearance in African American men compared with Caucasian men, and that race may significantly affect CYP3A-mediated hepatic clearance in patients treated for hypertension. PMID- 11269574 TI - Pharmacokinetics, safety, and tolerability of BAY 12-9566 and nonsteroidal anti inflammatory agents (naproxen, ibuprofen) during coadministration in patients with osteoarthritis. AB - The pharmacokinetic interactions between BAY 12-9566 and two nonsteroidal anti inflammatory drugs (NSAIDs), naproxen and ibuprofen, were investigated in osteoarthritis (OA) patients. The study comprised six groups: two NSAID groups with three levels of treatment (BAY 12-9566 400 mg, BAY 12-9566 100 mg, and placebo). Plasma pharmacokinetic parameters (AUC(0-tau), Cmax, and tmax) were determined for each treatment group following 5 days of NSAID administration, 14 days of BAY 12-9566 administration, and 14 days of concurrent NSAID and BAY 12 9566 administration. For most conditions, the total plasma drug concentrations of both NSAID and BAY 12-9566 were diminished by coadministration; total plasma BAY 12-9566 was not affected by ibuprofen treatment. Importantly, the free drug concentrations were largely unaffected by coadministration. Most side effects were mild or moderate in intensity, and all events, with the exception of headache, were reported in both NSAID groups and in both placebo and BAY 12-9566 groups. PMID- 11269575 TI - Lack of citalopram effect on oral digoxin pharmacokinetics. AB - The effect of chronic administration of citalopram on the single oral dose pharmacokinetics of digoxin was evaluated in 11 healthy adult subjects in an open, one-way crossover study. Subjects received 1 mg digoxin on day 1. Serial blood samples and total urine were collected over 192 hours, followed by an 11 day washout period. On days 22 through 50, subjects received 40 mg citalopram once daily. On day 43, a single dose of 1 mg digoxin was coadministered; again, serial blood samples and total urine were collected over 192 hours after the digoxin dose. There were no statistically significant differences in any of the digoxin pharmacokinetic parameters (AUC(0-->24), AUC(0-->infinity), Cmax, tmax, t(1/2), CL/F, CLrenal, and Ae(0-->infinity)), and the 90% confidence intervals for treatment differences for the parameters (except for tmax) were all within 80% to 125%. Concomitant digoxin administration did not significantly affect citalopram pharmacokinetics. The treatment was well tolerated by all subjects; no serious adverse events and no clinically significant ECG changes were observed. These data suggest that it is unlikely that concomitantly administered citalopram would have any significant effect on serum digoxin concentrations in patients who are receiving chronic digoxin therapy. PMID- 11269576 TI - Prospective randomized studies. PMID- 11269577 TI - Postoperative bleeding following notchplasty in anterior cruciate ligament reconstruction: thermal radio frequency versus powered instrumentation. AB - This study compared postoperative bleeding during anterior cruciate ligament (ACL) reconstruction following notchplasty by power instrumentation with that following radiofrequency. Between January 1998 and April 1998 we prospectively divided 24 consecutive patients undergoing arthroscopic ACL reconstruction with bone-patellar tendon-bone autograft into two groups. Notchplasty was performed by powered instrumentation in group A (n=12) and by radiofrequency in group B (n=12). Two Redi-Vac suction drains were placed, one intra-articularly and the other subcutaneously at the harvest site and tibial tunnel. All drains were removed 48 h postoperatively. The first drainage measurement (end of surgery, 6 a.m. postoperative day 1) showed average total bleeding of 124.16 cc in group A and 65.41 cc in group B (P<0.001); per hour this was 10.21 cc in group A and 5.49 cc in group B (P<0.001). The second drainage measurement (6 a.m. postoperative day 2) showed average total bleeding of 44.55 cc in group A and 17.78 cc in group B (P<0.01); per hour this was 1.85 cc in group A and 0.74 cc in group B (P<0.001). Radiofrequency technology can be used when performing intercondylar notchplasty in ACL reconstruction. As a result of this technique postoperative intra-articular bleeding was significantly reduced. PMID- 11269578 TI - The natural history of the knee following arthroscopic medial meniscectomy. AB - We examined the natural history of arthroscopic medial meniscectomy in knees with an isolated meniscal injury by reviewing 317 of 894 cases following medial meniscectomy. At the time of the initial surgery none of the knees had been operated on, and there was no evidence of ligament injury. The patients were reviewed clinically and radiologically after a mean of 11.5 years (range 10-15). The knee was considered "normal" or "nearly normal" by 91% of patients. In 218 patients the contralateral knee was asymptomatic without history of operation or significant injury and could be used as control for comparison. Radiology showed 22.4% greater excess prevalence of joint space narrowing in the operated compared to the control knee. The factors predisposing to a poor radiological result were age above 35 years, the presence of medial compartment cartilage degeneration at the time of the first arthroscopy, resection of the posterior one-third of the meniscus, and meniscal rim resection. Preoperative participation in sport was a predictor of a better outcome. PMID- 11269579 TI - Delayed gastrocnemius muscle response to sudden perturbation in rehabilitated patients with anterior cruciate ligament reconstruction. AB - This nonrandomized, posttest-only comparison between two experimental groups and a control group compared the lower extremity muscle activation latencies of patients following rehabilitated unilateral anterior cruciate ligament (ACL) reconstruction (allograft or autograft bone-patellar tendon-bone tissue) and normal control subjects. Twenty-three subjects (seven allograft, eight autograft, eight normal control) of similar age, height, weight, isokinetic knee extensor, and flexor peak torque/bodyweight, functional capability (single leg broad jump and single leg vertical jump) and recreational activity level participated in this study. Experimental group subjects were 21.3+/-5 months (allograft) and 27.6+/-10 months (autograft) after surgery. Kinematic and electromyographic data were sampled during ten randomly timed unilateral perturbations. Experimental group gastrocnemius latencies were delayed (allograft 59.5+/-25 ms, autograft 69+/-20 ms) compared to the control group (31.8+/-11 ms). The allograft (r=0.80) and autograft (r=0.40) unilateral ACL reconstruction groups displayed moderate and weak positive relationships between anterior knee laxity and knee angular displacements following perturbation, respectively. Control group subjects did not display significant relationships between these variables (r=-0.07). In the allograft group there was also a moderate inverse relationship between gastrocnemius latency and knee angular displacement following perturbation (r= 72). The autograft (r=-0.06) and control (r=-0.21) groups did not show similar relationships between these variables. Delayed gastrocnemius latencies for the experimental groups suggested prolonged neuromuscular deficits during weight bearing dynamic knee stabilization. Knee angular displacement magnitude following sudden perturbation was related more strongly to knee laxity and gastrocnemius latency among subjects who had undergone ACL reconstruction using allograft bone patellar tendon-bone tissue. PMID- 11269580 TI - The anatomy of the patellar tendon. AB - The morphology of the attachment of the patellar tendon, its bundle orientation, the differential fascicles length and the position of the apex of the patella were assessed in 22 cadaveric human knees. The patellar apex was 39+/-6% of the width of the tendon from its medial edge. The bulk of tendon was attached to the distal two-thirds of the anterior aspect of the patella. In six cases tendon fibres originated from the posterior surface of the apex of the patella, forming a ridge on the back of tendon. This may represent an anatomical variant accounting for the increased tendon thickness noted on MRI, both incidentally and during assessment for patellar tendonitis. Fascicles were parallel in the sagittal plane but converged in the frontal plane toward their tibial attachment. When bone-patellar tendon-bone (B-PT-B) grafts were harvested, as for anterior cruciate ligament reconstruction, the grafts narrowed distally. When harvesting B PT-B, the oblique orientation of the fibres in the coronal plane must be borne in mind. PMID- 11269581 TI - A comparative study of 'isometric' points for anterior cruciate ligament graft attachment. AB - Anterior cruciate ligament (ACL) reconstruction depends critically on isometric graft placement. Unfortunately, different supposedly isometric points have been published, and no prior work has compared them to find out which are really isometric. The purpose of this study was to compare the isometry of previously published 'isometric' points for ACL reconstruction. The isometric points and knee loadings of previous studies were reproduced accurately in 12 fresh cadaveric knees. The length changes were measured through 140 degrees knee flexion, using an intra-articular suture attached to a displacement transducer. Six points had less than 1 mm length change and were located proximally in the natural ACL attachment at the posterior end of Blumensaat's line. The other seven points had length change patterns that would cause ACL graft tightening or slackening with knee flexion if they were used as the sites of bone tunnels for graft placement. This study confirms the existence of an isometric zone close to the posterior end of Blumensaat's line under several loading conditions. Other graft attachment points are less suitable for ACL reconstruction. PMID- 11269582 TI - Instrumented measurement of glenohumeral joint laxity: reliability and normative data. AB - This study assessed shoulder laxity using an instrumented arthrometer. We compared anterior and posterior translations at various force levels to determine the reliability of our measurement technique and to provide normative data in healthy shoulders. Fifty shoulders were assessed for glenohumeral joint laxity in two directions (anterior and posterior) and at four force levels (67, 89, 111, and 134 N). The dependent measure was joint displacement. Laxity values were widely, yet normally, distributed in our group of healthy shoulders. Intraclass correlation coefficients revealed excellent between-trial reliability (0.92) and fair between-session (0.73) and between-examiner (0.74) reliability. The average standard error of measurement between trials (0.56 mm), sessions (1.5 mm), and examiners (1.7 mm) demonstrated an unprecedentedly high degree of precision for quantifying glenohumeral joint laxity. Paired t tests revealed no significant laxity differences between sides (P>0.05), indicating bilateral symmetry. A 2 (direction) x 4 (force) analysis of variance revealed significant differences in laxity between directions (P<0.0001) and force levels (P<0.0001). Our results show that our instrumented technique for quantifying glenohumeral joint laxity is precise and reproducible. Posterior translation was significantly greater than anterior, and a significant increase in translation was observed between increasing levels of force. PMID- 11269583 TI - Superior short-term results with eccentric calf muscle training compared to concentric training in a randomized prospective multicenter study on patients with chronic Achilles tendinosis. AB - In a previous uncontrolled pilot study we demonstrated very good clinical results with eccentric calf muscle training on patients with painful chronic Achilles tendinosis located at the 2-6 cm level in the tendon. In the present prospective multicenter study (Sundsvall and Umea) patients with painful chronic Achilles tendinosis at the 2-6 cm level in the tendon were randomized to treatment with either an eccentric or a concentric training regimen for the calf muscles. The study included 44 patients, with 22 patients (12 men, 10 women; mean age 48 years) in each treatment group. The amount of pain during activity (jogging or walking) was recorded by the patients on a visual analogue scale, and patient satisfaction was assessed before and after treatment. The patients were instructed to perform their eccentric or concentric training regimen on a daily basis for 12 weeks. In both types of treatment regimen the patients were told to do their exercises despite experiencing pain or discomfort in the tendon during exercise. The results showed that after the eccentric training regimen 82% of the patients (18/22) were satisfied and had resumed their previous activity level (before injury), compared to 36% of the patients (8/22) who were treated with the concentric training regimen. The results after treatment with eccentric training was significantly better (P<0.002) than after concentric training. The good clinical results previously demonstrated in the pilot study with eccentric calf muscle training on patients with chronic Achilles tendinosis, were thus reproduced in this multicenter, showing superior results to treatment with concentric training. PMID- 11269584 TI - Injuries in mountain biking. AB - Despite still growing attraction mountain biking as a matter of sports traumatology still lacks relevant data based on large cross-sectional surveys. To obtain an overview of risk factors, types, and main body sites of injuries occurring in mountain biking we assessed the results of a questionnaire answered by 3873 athletes. A total of 8133 single lesions were reported by 3474 athletes, 36% of whom regularly participated in competitions. The incidence of injuries in mountain biking is comparable to that in other outdoor sports, the majority of injuries being minor. Mountain biking athletes were found to have an overall injury risk rate of 0.6% per year and 1 injury per 1000 h of biking. The main risk factors included slippery road surface, cyclist's poor judgement of the situation, and excessive speed, representing personal factors that could be altered by preventive measures. Of all injuries 14% were due to collision with some part of the bike, especially the pedals and the handlebar. While 75% of the injuries were minor, such as skin wounds and simple contusions, 10% were so severe that hospitalization was required. A breakdown of the injuries according to body site and frequency of occurrence is presented. PMID- 11269585 TI - Preconditioning patellar tendon autografts in arthroscopic anterior cruciate ligament reconstruction: a prospective randomized study. AB - This prospective randomized evaluated the effect of preconditioning patellar tendon autografts before implantation and fixation during anterior cruciate ligament (ACL) reconstruction. Fifty-three patients with a unilateral ACL rupture were included in the study. One group of patients had their patellar tendon autograft preconditioned by passive stretching at a constant load of 39 N for 10 min immediately prior to implantation (group P). The other group underwent no preconditioning before the implantation of the graft (group NP). The follow-up examination was performed by independent observers after 26 months (23-29) in group P and after 25 months (23-30) in group NP (n.s.). At follow-up the KT-1000 laxity test revealed a total side-to-side difference of 2.5 mm (-1.5 to +8.5) in group P and 3.0 mm (-7 to +6.5) in group NP (n.s.). The Lysholm score was 86 points (47-100) in group P and 94 points (44-100) in group NP (n.s.). The Tegner activity level was 6 (2-9) in group P and 7 (3-9) in group NP (n.s.). There was no significant difference between the study groups regarding IKDC classification. Patients who underwent ACL reconstruction using a preconditioned patellar tendon autograft had no advantages in terms of restoration of laxity or clinical outcome at 2-year follow-up. PMID- 11269587 TI - Gas chromatographic analysis of trace gas impurities in tungsten hexafluoride. AB - Highly reactive fluorinated gaseous matrices require special equipment and techniques for the gas chromatographic analysis of trace impurities in these gases. The impurities that were analysed at the low-microg/l levels included oxygen, nitrogen, carbon dioxide, carbon monoxide, sulfur hexafluoride and hydrogen. This paper describes the use of a system utilising backflush column switching to protect the columns and detectors in the analysis of trace gas impurities in tungsten hexafluoride. Two separate channels were used for the analysis of H2, O2, N2, CO, CO2 and SF6 impurities with pulsed discharge helium ionisation detection. PMID- 11269586 TI - Practical aspects of ultrahigh pressure capillary liquid chromatography. AB - A novel pressure-balanced injection valve was evaluated for use with ultrahigh pressure liquid chromatography (UHPLC) at pressures up to 120 MPa (1,200 bar). Fused-silica capillaries (30-33 cm x 100 microm I.D.) packed with nonporous 1.5 microm isohexylsilane-modified (C6) silica particles were employed to study maximum pressure, injection reproducibility, injection time, and sample amount consumed for an injection. The new valve was more reproducible, convenient, and required much less sample than previously used injection systems. The effect of column diameter on efficiency and sensitivity was studied. The 100 microm I.D. columns demonstrated approximately 40% lower efficiency but 10-fold higher sensitivity than the 29 microm I.D. columns. Columns packed with nonporous C6 particles produced higher efficiencies than columns packed with a 1.5 microm porous octadecylsilane-modified (C18) material. PMID- 11269588 TI - High-resolution analysis of polyprenols by supercritical fluid chromatography. AB - A high-resolution analysis of polyprenol mixtures was achieved by supercritical fluid chromatography (SFC). The separation of polyprenols was examined on an octadecylsilane-packed column with liquid carbon dioxide as the mobile phase and ethanol as modifier. Using this chromatography system, the resolution of separation (Rs) between octadecaprenol (prenol 18) and nonadecaprenol (prenol 19) was two times higher than that using conventional reversed-phase high-performance liquid chromatography. Our SFC technique allows the advantage of baseline separation of polyprenol samples containing hydrophobic components such as terpenes or fatty acids that are unfavorable for good separation. This method is very useful for the analysis of structurally close polyprenol analogues of rubber plant metabolites. PMID- 11269589 TI - Identification and quantification of base and nucleoside markers in extracts of Ganoderma lucidum, Ganoderma japonicum and Ganoderma capsules by micellar electrokinetic chromatography. AB - The present paper describes the development of a micellar electrokinetic chromatographic method for the determination of nucleoside (adenosine, uridine) and base (uracil) markers in aqueous extracts of Ganoderma medicinal preparations. The markers were successfully separated within 10 min using an 80 mM borate buffer, with 25 mM sodium dodecyl sulfate adjusted to pH 9.0, an operating voltage of 22 kV, temperature of 20 degrees C and a hydrodynamic injection time of 5 s. Separations were carried out in a fused-silica capillary with peak detection by direct UV at 254 nm. Following semi-validation of the method, with each analyte showing a good linear relationship over a 0.2 to 20 ppm concentration range (correlation coefficients from 0.9986 to 0.9998), the amounts of the three markers in the various forms of Ganoderma were easily determined using a relatively simple extraction procedure. PMID- 11269590 TI - Separation of uranium(VI) and transition metal ions with 4-(2 thiazolylazo)resorcinol by capillary electrophoresis. AB - A capillary electrophoresis method utilizing 4-(2-thiazolylazo)resorcinol (TAR) was developed to separate uranium, cobalt, cadmium, nickel, titanium and copper metal ions. TAR was chosen as the visible absorbing chelating ligand because of its ability to form stable complexes with a wide variety of metals. Several parameters that included pH, electrophoretic run buffer concentration, buffer type and the influence of chelating ligand in the electrophoretic run buffer were examined to determine the best separating conditions. Optimum separation of the six metal chelates was achieved in a 15 mM Na2B4O7-NaH2PO4, pH 8.3 buffer containing 0.1 mM TAR. Method validation included injection and method precision studies as well as detection limit and linear dynamic range determination. High ppb to low-ppm (w/w ratio) detection limits were achieved with linear dynamic ranges between 0.1 and 75 ppm. PMID- 11269592 TI - Fast determination of phenols in contaminated soils. AB - An extraction method for the determination of phenols in contaminated soils, based on the application of solid-phase microextraction (SPME) coupled with GC flame ionization detection analysis, was developed and tested. This method was developed using a natural soil spiked with phenol to a concentration level typical of an acute contamination event that can occur in an industrial site. The effects of the extraction parameters (pH, extraction time and salt concentration) on the extraction efficiency were studied and the method was then applied to determine the pollutant concentration at the beginning and during the biological treatment of a soil, contaminated with phenol and 3-chlorophenol, respectively. The SPME results were validated by comparison with those obtained with an US Environmental Protection Agency certified extraction method. The SPME method was also successfully applied to the determination of the adsorption behavior of 3 chlorophenol on a natural clay soil and was shown to be suitable for different matrices and phenolic compounds. Application of SPME technique results in a sharp reduction of the extraction times with negligible solvent consumption. PMID- 11269591 TI - Separation and characterization of oligomers by reversed-phase high-performance liquid chromatography: a study on well-defined oligothiphenes. AB - Reversed-phase high-performance liquid chromatography (RP-HPLC) was used for the separation of 3-hexylthiophene oligomers in the range of 3 to 30 monomeric units, while systematically varying stationary and mobile phases. A set of different columns was chosen, covering a broad range of silica types, pore sizes and bonding chemistry. Mobile phases of tetrahydrofuran (THF) combined with water, acetonitrile (ACN) or methanol (MeOH) were used. Although differences between columns were small, a higher selectivity correlated with a lower hydrophobicity parameter from the Galushko column test. The model of Jandera, based on the linear solvent strength model of Snyder, was used to describe the retention of the oligomers in gradient mode. This gave information about selectivities on different stationary phases similar to the hydrophobicity parameter. Contrary to the stationary phase, the mobile phase had a major influence on the selectivity. The THF-water combination gave much higher selectivities compared to THF combined with MeOH or ACN. Using the aqueous mobile phase even enabled separation of different isomers. Determination of thermodynamic parameters for the model compounds showed that retention of the different isomers was mainly determined by the orientation of the side chains at both ends of the chain. An additional repeating unit in the middle of the polymer backbone gave a similar contribution to retention, irrespective of the orientation of its side chain. Three model isomers were separated by preparative RP-HPLC and identified by proton nuclear magnetic resonance spectroscopy. The combination of subsequent preparative size exclusion chromatography, RP-HPLC and matrix-assisted laser desorption ionization time-of-flight mass spectrometry enabled the identification of the two major oligomeric series in the sample as the regioregular product with one bromine end group and, in smaller amounts, a regioirregular product with two bromine end groups. reserved. PMID- 11269593 TI - Quantitative interpretation of Fourier-transform infrared spectoscopic data from a size--exlusion chromatorgraphy solvent- evaporation interface. AB - Quantitative evaluation of polymer composition across the SEC chromatogram can provide more accurate characterization of heterogeneous polymer samples for problem solving and for material specification. To this end Fourier-transform infrared spectroscopy (FTIR) with solvent-evaporation interfaces has become a very powerful detector for size-exclusion chromatography (SEC). The solvent evaporation interface removes the mobile phase at the exit of the chromatograph and deposits the separated molecular sizes as polymer films on infrared transparent substrates. Quantitative interpretation of the FTIR spectra obtained from these films has recently been found to be best accomplished by using partial least squares. In this paper, polystyrene and poly(methylmethacrylate), alone, as blends, and a copolymer were analyzed in a SEC equipped with an evaporative interface. Molecular weight effects, wavelength selection, the effect of averaging spectra on results, and selection of the best data preprocessing method were investigated. General methods of evaluating these variables were developed to arrive at conditions for this particular "model" situation in order to provide a basis for the analysis of more complex polymers. PMID- 11269595 TI - Cyclodextrin biospecific-like displacement in dye-affinity chromatography. AB - Interactions between Cibacron Blue F3GA (CB F3GA), as a model of triazine dye, and 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CD), as a model of cyclodextrin, were investigated by monitoring the spectral shift that accompanies the binding phenomena. Matrix analysis of the difference spectral titration of CB F3GA with HP-beta-CD revealed only two absorbing species, indicating a host-guest ratio of 1:1. The dissociation constant for this HP-beta-CD-CB F3GA complex, Kd, was found to be 0.43 mM. The data for HP-beta-CD forming inclusion complexes with CB F3GA were used to develop the concept of competitive elution by inclusion complexes in dye-affinity chromatography. When this concept was applied to the elution of L lactate dehydrogenase from a CB F3GA affinity matrix, it was shown to be an effective elution strategy. It provided a 15-fold purification factor with 89% recovery and sharp elution profile (0.8 column volumes for 80% recovery), which is as good as that obtained by specific elution with NADH (16-fold, 78% recovery and 1.8 column volumes). In addition, the new elution strategy showed a better purification factor and sharper elution profile than traditional non-specific elution with KCl (4.5-fold, and 1.4 column volumes). Hence, competitive elution by inclusion complexes may be a promising strategy for eluting proteins with high recoveries and purification factors in dye-affinity chromatography. PMID- 11269594 TI - Identification and quantification of molecular species of diacyl glyceryl ether by reversed-phase high-performance liquid chromatography with refractive index detection and mass spectrometry. AB - We have developed a method to identify and quantify the molecular species of diacyl glyceryl ether (DAGE) using high-performance liquid chromatography (HPLC) equipped with a refractive index detector and an electrospray ionization and time of flight mass spectrometer (LC-RI-MS). An octadecyl silica column with a mixture of acetonitrile and dichloromethane (65:35, v/v) as an eluant was used for the HPLC. When the LC-RI-MS method was applied to a mixture of synthetic DAGEs; 1-O hexadecyl-2,3-dioleoylglycerol (O-16:0-18:1-18:18:1), 1-O-octadecyl-2,3 dioleoylglycerol (O-18:0-18:1-18:1), 1-O-octadecenyl-2,3-dioleoylglycerol (O-18:1 18:1-18:1), 1-O-octadecyl-2,3-didocodahexaenoylglycerol (O-18:0-22:6-22:6), and 1 O-octadecenyl-2,3-didocosahexaenoylglycerol (O-18:1-22:6-22:6), good separation and quantification were obtained on the refractive index chromatogram. A pseudo molecular ion [M+NH4]+ and a monoacyl glyceryl ether ion [M-RCO2] + were observed for all synthetic DAGEs on the mass spectrum. It was found that the fatty acids and glyceryl ether in DAGE could be easily identified by these mass spectra. When this LC-RI-MS method was applied to the DAGEs extracted from muscle of Stromateus stellatus, approximately 18 peaks were observed on LC-RI-MS chromatograms and the major molecular species of DAGEs were identified as O-16:0-18:1-18:1. PMID- 11269596 TI - Studies on factors influencing stability and recovery of paclitaxel from suspension media and cultures of Taxus cuspidata cv Densiformis by high performance liquid chromatography. AB - An HPLC method was developed for quick scanning of taxanes from large numbers of plant cell suspension samples. The method was optimized for analysis of a range of taxanes of differing polarity. Identification of a standard mixture of paclitaxel and 12 related taxanes was achieved in less than 15 min using a gradient mode and a Microsorb-MV C8 column. The method was used to investigate the influence of several factors on stability and recovery of paclitaxel from suspension media and cultures of Taxus cuspidata cv Densiformis. Incubation time had the most significant influence on stability of paclitaxel, contributing 88% to the total variation. Shaking contributed 6% to the total variation. Light contributed only 0.25% to the total variation. Analysis of test samples of suspension cultures of T. cuspidata cv Densiformis over a 4 week period show paclitaxel, 10-deacetylbaccatin III, and baccatin III levels ranging from 0 to 149 microg/L, from 0 to 1.9 mg/L, and from 0 to 583 microg/L, respectively. PMID- 11269597 TI - Studies on the response mechanism of thermionic detection to organophosphorus compounds. AB - The response of thermionic/nitrogen-phosphorus detection (TID) to a series of organophosphonate esters has been studied. The response of TID is found to decrease with the increase in the alkyl chain length of the molecules. An attempt has been made to propose the response mechanism of TID for these compounds. The charge carriers accountable for the response do not necessarily arise by combustion of the molecule but by a reaction involving alkali metal and the compound. This has been supported by thermodynamic parameters and molecular descriptors. The mechanism of the reaction that appears to be bimolecular has been explained by steric effects. The study successfully explains the observed change in the response of TID with minor changes in structures of the molecules. PMID- 11269598 TI - Gas chromatographic-mass spectrometric determination of hydrophilic compounds in environmental water by solid-phase extraction with activated carbon fiber felt. AB - Simple gas chromatographic-mass spectrometric determination of hydrophilic organic compounds in environmental water was developed. A cartridge containing activated carbon fiber felt was made by way of trial and was evaluated for solid phase extraction of the compounds in water. The hydrophilic compounds investigated were acrylamide, N,N-dimethylacetamide, N,N-dimethylformamide, 1,4 dioxane, furfural, furfuryl alcohol, N-nitrosodiethylamine and N nitrosodimethylamine. Overall recoveries were good (80-100%) from groundwater and river water. The relative standard deviations ranged from 4.5 to 16% for the target compounds. The minimum detectable concentrations were 0.02 to 0.03 microg/l. This method was successfully applied to several river water samples. PMID- 11269599 TI - Determination of thiodyglycol in groundwater using solid-phase extraction followed by gas chromatography with mass spectrometric detection in the selected ion mode. AB - A highly sensitive analytical procedure is described for determining thiodiglycol in groundwater. Samples are initially fortified with 3,3'-thiodipropanol (surrogate), then both species are extracted using sequential solid-phase extraction with both C18 and Ambersorb 572 columns. The C18 column, which removes extraneous groundwater components, is discarded; the Ambersorb 572 column is dried thoroughly before eluting polar components with a small volume of dichloromethane. The extract is taken to dryness using dry flowing nitrogen, and the resulting residue is derivatized using N-(tert.-butyldimethylsilyl)-N methyltrifluoroacetamide and pyridine. The derivatized products are diluted to a final volume with toluene, chromatographed using a fused-silica capillary column, and detected with a quadrupole mass spectrometric detector in its selected-ion mode. Two independent, statistically unbiased, procedures were used to evaluate the detection limits for thiodiglycol; the values ranged between 4 and 16 microg( 1) groundwater. PMID- 11269600 TI - Development of a screening method for cocaine and cocaine metabolites in urine using solvent microextraction in conjunction with gas chromatography. AB - A simple, quick and inexpensive screening method for cocaine and cocaine metabolites has been developed. Drug extraction was achieved using the relatively new technique of solvent microextraction (SME). Complete analysis is achieved in 13 min, using, a 6-min extraction with a 2-microl drop followed by separation on a gas chromatograph. The developed procedure was tested as a screening method for cocaine and cocaine metabolites in spiked urine samples. Using SME, concentrations as low as 0.125 microg ml(-1) of cocaine, ecgonine methyl ester, cocaethylene and anhydroecgonine methyl ester were measurable with relative standard deviation values averaging 9.0%. PMID- 11269601 TI - Food & fitness: build a healthy lifestyle. PMID- 11269602 TI - Dietitians can prevent listeriosis. PMID- 11269603 TI - Understanding dietary patterns essential to patient care. PMID- 11269604 TI - Genetically engineered "golden" rice unlikely to overcome vitamin A deficiency. PMID- 11269605 TI - Determining when obesity is a disease. PMID- 11269606 TI - Dietary reference intakes: vitamin A, vitamin K, arsenic, boron, chromium, copper, iodine, iron, manganese, molybdenum, nickel, silicon, vanadium, and zinc. PMID- 11269607 TI - Genetic variants, diet, and physical activity. PMID- 11269609 TI - Evaluating brief measures of fruit and vegetable consumption frequency and variety: cognition, interpretation, and other measurement issues. AB - To evaluate whether items from 3 brief measures of fruit and vegetable consumption were understood and interpreted as intended, cognitive testing was conducted in a purposive sample of 31 white, African-American and Hispanic persons. The measurement instruments tested were the fruit and vegetable module from the Behavioral Risk Factor Surveillance System (to measure frequency), and 1 fruit and 1 vegetable variety measurement instrument developed by the investigators. The cognitive testing interviews were analyzed qualitatively to identify interpretation difficulties and other measurement issues. The testing identified a number of measurement issues, including issues related to time frame, wording, interpretation, grouping of items, and serving size. Recommendations based on the findings were incorporated into revised versions of each instrument, which were further tested in a small sample. As revised and presented in this article, these instruments for assessing fruit and vegetable frequency and variety appear to be understood and interpreted as intended across different racial and ethnic groups, and may be useful in situations requiring brief dietary assessment, although further testing is needed. PMID- 11269608 TI - Dietary fiber intakes and insulin requirements in pregnant women with type 1 diabetes. AB - OBJECTIVE: To determine whether higher dietary fiber intake (water soluble and insoluble) is associated with lower insulin requirements and better glycemic control in pregnant women with type 1 diabetes consuming a self-selected diet. DESIGN: A longitudinal, observational study. SUBJECTS: Pregnant women (n=141) with type 1 diabetes participating in an interdisciplinary program examining the effects of glycemic control on pregnancy outcome (Diabetes and Pregnancy Program, University of Cincinnati Medical Center). MEASUREMENTS: We determined total, water soluble and insoluble fiber intakes from 3-day food records kept each trimester during pregnancy. Outcome measures were insulin dose, pre-meal blood glucose, and glycated hemoglobin concentrations. STATISTICAL ANALYSES: Correlation coefficients, multiple regression, mixed-model analysis of variance. RESULTS: Mean intakes (g/day) of total, water soluble fiber, and insoluble fiber were 14.0 (range, 1.8-33.1), 4.8 (range, 0.6-10.5) and 9.0 (range, 1.1-24.0), respectively. In the second and third trimesters of pregnancy, insulin requirements were inversely associated with total, water soluble, and insoluble fiber intakes; the correlation coefficients ranged from -0.22 to -0.17 (P=.02 to .08). Insulin requirements associated with a higher fiber intake (20.5 g/day) were 16% to 18% lower than for a lower fiber intake (8.1 g/day). These relations remained after adjustment for body weight, disease severity and duration, insulin type, and study year in the second (P=.03 to .10) but not in the third trimester. Pre-meal blood glucose and glycated hemoglobin concentrations were not associated with fiber intake. CONCLUSIONS: Among pregnant women with type 1 diabetes, higher fiber intake is associated with lower daily insulin requirements. Dietary fiber intake should be considered when counseling patients about the management of blood glucose concentrations. PMID- 11269610 TI - How do I help patients with pica? PMID- 11269611 TI - Energy and macronutrient intakes of elite figure skaters. AB - OBJECTIVES: Dietary guidelines for athletes emphasize complex carbohydrates. This study examined dietary intakes of elite figure skaters relative to current recommendations in sports nutrition. PARTICIPANTS: Subjects were male (n=80) and female (n=81) figure skaters taking part in a series of training camps held in Colorado between 1988 and 1995. Mean age was 18 years for men and 16 years for women. DESIGN: Measures of height, weight, and skinfold thickness were used to calculate body mass index and percent body fat. Blood samples were drawn for analysis of nutritional status. Energy and nutrient intakes were based on 3-day food records. STATISTICAL ANALYSES: Multivariate regression model and correlation analyses used the SPSS for Windows program. RESULTS: Values of body mass index and percent body fat were similar to those obtained for elite athletes in other studies. Plasma chemistries were in the normal range. Energy intakes (2,329 kcal/day for men and 1,545 kcal/day for women) were below recommended values for sex and age. The skaters derived approximately 50% of their daily energy from sugars and fat. Sugars alone accounted for 25% of daily energy intakes--the skaters consumed between 100 g (women) and 142 g of sugars per day. Sugar and fat intakes, when expressed as percent of daily energy, were inversely linked, providing evidence of a fat-sugar seesaw. Higher-energy diets were higher in fat but lower in carbohydrate and protein. APPLICATIONS: High consumption of sugars and fat by elite athletes was not associated with overweight or excess body fat. Although recommended diets are usually built around complex carbohydrates, dietetics professionals can address the increased energy needs of elite athletes by recommending energy-dense foods. Sugars and fats are efficient sources of energy per unit volume. PMID- 11269612 TI - Helping adolescent athletes achieve a gold-medal diet. PMID- 11269613 TI - Diets with either beef or plant proteins reduce risk of calcium oxalate precipitation in patients with a history of calcium kidney stones. AB - OBJECTIVE: To determine the effect of substituting equal amounts of dietary protein as animal protein (beef) for plant protein (legumes, seeds, nuts, and grains) on urinary components associated with calcium oxalate precipitability risk. DESIGN: Randomized crossover trial. SUBJECTS: Twenty-three normocalciuric patients with a history of calcium kidney stones (8 women and 15 men, mean age 50.7+/-14.6 years) with 24-hour urinary calcium < or =10.3 micromol, 24 hour urinary oxalate excretion between 228 and 963 micromol, and a urinary calcium increase of < or =1.0 micromol in 4 hours after a 25 micromol oral calcium load. SETTING: Four-day, free-living adaptation period, followed by 2-day metabolic unit study. INTERVENTION: The study compared consumption of 2 servings of beef (43 g protein for women and 50 g for men) daily with an equal amount of protein from plant foods including legumes, nuts, and grains. MAIN OUTCOME MEASURES: Tiselius risk index (TRI) for calcium oxalate precipitability calculated from urinary calcium, oxalate, magnesium, citrate, and volume. STATISTICAL ANALYSES: Paired t tests. RESULTS: Urinary calcium, oxalate, magnesium, citrate, phosphorus, volume, and TRI did not differ between diets. Urinary sodium and potassium were higher for patients on the plant protein diet. After correcting for variations in urinary sodium and potassium between diets, the difference in urinary calcium remained insignificant. TRI was lower on both beef- and plant protein diets compared with self-selected prestudy diets for all participants. CONCLUSION/APPLICATIONS: Balanced diets containing moderate amounts of either beef or plant protein are equally effective in reducing calcium oxalate kidney stone risk based on changes in urinary composition. PMID- 11269614 TI - Facilitating dietary change: the patient-centered counseling model. AB - Recent data indicate that the patient-centered counseling model enhances long term dietary adherence. This model facilitates change by assessing patient needs and subsequently tailoring the intervention to the patient's stage in the process of change, personal goals, and unique challenges. This article describes this model, including its theoretical foundations, a 4-step counseling process, and applications. This behavioral counseling model can help nutrition professionals enhance patient adherence to nutrition care plans and dietary guidelines. PMID- 11269615 TI - Inadequate folic acid intakes are prevalent among young women with neural tube defects. PMID- 11269616 TI - Liquid meal replacement vs traditional food: a potential model for women who cannot maintain eating habit change. PMID- 11269617 TI - Vitamin A, vitamin C, calcium, and iron content of federally funded preschool lunches in Virginia. PMID- 11269618 TI - Dietary intake and energy expenditure of female collegiate swimmers during decreased training prior to competition. PMID- 11269619 TI - Acceptability of low-fat, sugar-free cakes: effect of providing compositional information during taste-testing. PMID- 11269620 TI - A proposed model for effective nutrition care. PMID- 11269621 TI - The QT interval. AB - The effects of disease states and therapeutic drugs on the QT interval have been extensively studied in an attempt to understand the relationship between QT and the risk of torsade de pointes and sudden cardiac death. Differences in heart rate correction methods, electrocardiogram lead placement, and other internal (eg, genetic, physiologic) and external (eg, food, time of day) factors have confounded the interpretation of this relationship. A comprehensive review of the epidemiologic literature suggests that the corrected QT interval (QTc) is an important but imprecise marker of cardiovascular disease. The association between QTc prolongation and mortality has been identified in patients with cardiac disease but is unclear in patients without cardiac disease. Drug-related prolongation of QTc can clearly increase the risk of torsade de pointes, but this arrhythmia is rarely associated with a QTc of less than 500 ms. It also appears that noncardiac drugs that are associated with QTc prolongation are not identical in their proarrhythmic capacities and that increased exposure via clinically significant drug interactions is a major contributor to the liability of noncardiac drug-induced arrhythmia. Recognition of the aforementioned variables in conjunction with careful QTc measurements assists in establishing a more precise benefit-risk ratio for a specific drug therapy or for arrhythmia risk associated with various pathophysiologic or genetic states. PMID- 11269622 TI - Introduction to the special section: hypnosis and EMDR. Eye Movement Desensitization and Reprocessing. PMID- 11269623 TI - The challenges of treatment evolution and integration. PMID- 11269625 TI - Accessing the power in the patient with hypnosis and EMDR. Eye Movement Desensitization and Reprocessing. AB - Both Ernest Rossi's ideodynamic accessing model of hypnosis and EMDR are intended to access information stored in the mind-body system. In this paper the author is suggesting that treatment effectiveness can be enhanced by combining these particular models. The similarities and the uniqueness of each method, both theoretically and in terms of the different protocols, are compared to provide a rationale for combining them. Verbatim examples from clinical cases are presented to demonstrate exactly how these models can be usefully combined in clinical practice. PMID- 11269624 TI - ECEM (eye closure eye movements): integrating aspects of EMDR with hypnosis for treatment of trauma. AB - The paper addresses distinctions between hypnotic interventions and Eye Movement Desensitizing and Reprocessing (EMDR) and discusses their effect on persons who have symptoms of Posttraumatic Stress Disorder (PTSD). Eye movements in hypnosis and EMDR are considered in terms of the different ways they may affect responses in treatment. A treatment intervention within hypnosis called ECEM (Eye Closure, Eye Movements) is described. ECEM can be used for patients with histories of trauma who did not benefit adequately from either interventions in hypnosis or the EMDR treatment protocol used separately. In ECEM the eye movement variable of EMDR is integrated within a hypnosis protocol to enhance benefits of hypnosis and reduce certain risks of EMDR. PMID- 11269627 TI - Integrative psychotherapy: combining ego-state therapy, clinical hypnosis, and eye movement desensitization and reprocessing (EMDR) in a psychosocial developmental context. AB - The principles of this conceptual framework are: (1) personality organization is dissociative as well as associative, consisting of ego states, and progresses through stages of psychosocial development; (2) inappropriately activated ego states cause dysfunction, which is habitual or due to the intense affect of disrupted development or unresolved grief or trauma; (3) completely overcoming dysfunction requires therapy with both individual ego states and the personality system; (4) clinical hypnosis provides techniques to enhance accessing ego states; and (5) EMDR combines ego-state therapy with eye movements (EMs) to produce a powerful psychotherapy method. During assessment, ego states responsible for dysfunctional emotional reactions and behavior are identified together with those that could be appropriate instead. Included in the treatment protocol, EMs and clinical hypnosis promote: (1) corrective developmental experiences; (2) resolution of grief and trauma; (3) acquisition of skills and abilities; (4) co-consciousness; and (5) negotiation among ego states. The outcome is an integrated "family of self" that has effectively overcome developmental crises, grief, and trauma, is aware of essential inner resources, and can consciously activate appropriate ego states. PMID- 11269626 TI - Recommendations and illustrations for combining hypnosis and EMDR in the treatment of psychological trauma. AB - Three experienced therapists, trained in hypnosis and EMDR, distilled some tentative hypotheses about the use of hypnosis in EMDR from fifteen cases, two presented here. When a therapist uses hypnosis with EMDR, it seems that the client is having difficulty or the therapist anticipates that the client will have difficulty managing the experiences processed with EMDR. Hypnosis initiated either during the introduction to EMDR or within a therapy session prior to the initiation of EMDR seems to have served two functions. The first function is to activate inner work that prepares the client to use EMDR successfully, and the second function is to facilitate overtly the processing of the traumatic experience. Clients might have two kinds of difficulties in managing affect or distress: (1) they may have a long-standing, irrational and strongly held belief that interferes with managing affect or distress, and (2) they may never have developed the capacity to tolerate intense affect, distress or pain. Should a therapist use hypnosis during the closing down phase of a session without preparing the client with hypnosis during the introduction to EMDR, the therapist should seriously reconsider the pace and focus of EMDR and the client's resources to manage affect and distress. PMID- 11269628 TI - Potential contributions of hypnosis to ego-strengthening procedures in EMDR. Eye Movement Desensitization Reprocessing. AB - This paper explores how hypnotic principles can be systematically incorporated into the standard EMDR protocol to enhance various ego strength capacities during EMDR treatment. Expanding these skill areas can widen the therapeutic window of possibility for clients with a variety of complex clinical issues, including posttraumatic, dissociative or personality disorders, anxiety symptoms, and depressive difficulties. Clinical case examples are used to illustrate ways of integrating hypnotic principles within a proposed EMDR protocol to promote ego strengthening and facilitate therapeutic change. PMID- 11269629 TI - EMDR and hypnosis in the treatment of phobias. Eye Movement Desensitization and Reprocessing. AB - Clinical hypnosis and EMDR have both been employed in the treatment of phobias. EMDR has been a controversial treatment method with the research showing mixed results concerning its efficacy. Many studies have shown the effectiveness of hypnosis in the treatment of phobias, but no studies have directly compared hypnosis to EMDR. This paper discusses each approach to treatment, with special emphasis on EMDR. Relevant research and current theories are reviewed along with questions raised and recommendations for future research. PMID- 11269630 TI - The wreathing protocol: the imbrication of hypnosis and EMDR in the treatment of dissociative identity disorder and other dissociative responses. Eye Movement Desensitization Reprocessing. AB - Dissociative Identity Disorder (DID), a chronic childhood onset posttraumatic stress disorder, is currently recognized as a treatable condition. It is considered the paradigmatic dissociative condition and carries with it extreme posttraumatic symptomatology. Therapists skilled in the treatment of DID are typically fluent in the uses of hypnosis for stabilization, affect management, building a safe place and grounding to name of few. EMDR, which has come to the forefront of clinical awareness in the last ten years, seems aptly suited for the treatment of trauma, but can be destabilizing. This paper proposes a protocol, called Wreathing Protocol, for the imbricated use of EMDR and hypnosis in the treatment of not only DID (though this will be the primary focus of the paper), but also Dissociative Disorder Not Otherwise Specified (DDNOS) and chronic Posttraumatic Stress Disorder (PTSD). This protocol is useful to advanced clinicians skilled in both modalities independently. The sequential steps of the Wreathing Protocol will be described and illustrated by a clinical vignette on DID. The clinical implications of the use of the Wreathing Protocol will be discussed in DID as well as the chronic post traumatic spectrum. PMID- 11269631 TI - Hypnosis, the hidden observer, and not-so-hidden consent. PMID- 11269632 TI - Informed dissent regarding hypnosis and its not-so-hidden observers: comment on Lynn. PMID- 11269633 TI - Informed consent and the standard of care in the practice of clinical hypnosis. PMID- 11269635 TI - Different perspectives on informed consent and clinical hypnosis. PMID- 11269634 TI - Informed consent and uninformed clinical practice: dissociation, hypnosis and false memories. PMID- 11269636 TI - What suggestion is best for pain? PMID- 11269637 TI - Pain management for neonatal circumcision. AB - Circumcision is the most common surgical procedure performed in the neonatal period in North America. If untreated, the pain of circumcision causes both short and long term changes in infant behaviours. The most widely studied pharmacological intervention for pain management during circumcision is dorsal penile nerve block (DPNB) by injected lidocaine (lignocaine). Randomised controlled trials have demonstrated its efficacy; infants premedicated with lidocaine have significantly smaller changes in physiological and pain-related behaviours compared with infants who are not given analgesics. A meta-analysis of injection-related adverse effects (bruising/haematoma) yielded a risk of 6.7% (95% confidence interval, 0.5 to 12.9%). Systemic toxicity from injected local anaesthesia has not been reported. Less effective modalities include topical anaesthesia with lidocaine-prilocaine cream [Eutectic Mixture of Local Anaesthetics (EMLA)], lidocaine cream and oral administration of sucrose. The good tolerability of lidocaine-prilocaine cream has been demonstrated by a lack of clinically significant methaemoglobinaemia when used appropriately. Nonpharmacological interventions (pacifier, specially designed restraint chair) reduce distress during the procedure, and paracetamol (acetaminophen) may provide postoperative analgesia. No single agent has been demonstrated to ameliorate pain for all infants undergoing circumcision. A multimodal approach of pharmacotherapy is currently recommended. Studies evaluating the efficacy of combined analgesia have demonstrated significant benefits for combinations of 2 or more forms of treatment (such as DPNB and sucrose-dipped pacifier) compared with single interventions. The instrument used to perform the circumcision is also important. The Mogen clamp has been shown to be associated with a shorter procedure time and less pain compared with the Gomco clamp. If circumcision is to be performed on infants, it is, therefore, recommended that combined analgesia and the Mogen clamp technique are used, and nonpharmacological stress reducing interventions such as pacifiers and comfortable restraining chairs should also be employed. PMID- 11269638 TI - Antimalarial chemoprophylaxis in infants and children. AB - The evolving patterns of drug resistance in malaria parasites and changes in recommendations for malaria prevention present a challenge to physicians who advise travellers on chemoprophylaxis. Because compliance with personal protection measures is usually low, children should receive appropriate chemoprophylaxis, including breast-fed infants who are not protected through maternal chemoprophylaxis. For travel to areas where chloroquine resistance has not yet been reported (i.e. parts of Central America, the Caribbean and parts of the Middle East), chloroquine alone is sufficient for antimalarial prophylaxis. Mefloquine is the drug of choice for chemoprophylaxis in areas with chloroquine resistant Plasmodium falciparum, and can be given to infants and young children. The combination of chloroquine and proguanil is well tolerated in children but is much less effective against drug-resistant malaria. Further research is needed to determine the best dosage regimen for antimalarial drugs used for chemoprophylaxis in children. PMID- 11269639 TI - Treatment of hydatid disease. AB - Hydatid disease caused by Echinococcus granulosus presents medical, veterinary and economic problems worldwide. Hydatidosis can be treated by medical, surgical and percutaneous modalities. Benzoimidazole carbamates are effective against E. granulosus. Although mebendazole, the first benzoimidazole used, has some beneficial effects on the disease in selected patients, it has also been associated with treatment failure in some cases, perhaps because of its poor absorption. Albendazole, a more recently developed benzoimidazole, is more effective than mebendazole. Praziquantel, an isoquinoline derivative, has recently shown value in the treatment of human echinococcal disease and its use in combination with albendazole is recommended in some patients. Ultrasound guided cyst puncture is another choice of treatment which has been used successfully in selected patients, although anaphylactic and allergic reactions due to spillage of the cyst contents have occurred. Surgical therapy in echinococcal hydatid disease is indicated for large cysts with multiple daughter cysts, superficially located single liver cysts which have a risk of rupture, complicated cysts such as those accompanied by infection, compression or obstruction, and cysts located in vital organs or which are exerting pressure on adjacent vital organs. However, surgical therapy carries high risk of mortality, morbidity or recurrence. Therefore, medical therapy may be an alternative option in uncomplicated cysts and in patients at high risk from surgery. The adjuvant use of drugs with surgery and percutaneous treatment can also be recommended for some patients. PMID- 11269641 TI - Management of neonatal herpes simplex virus infection. AB - Herpes simplex viruses (HSV) are ubiquitous pathogens which can be transmitted vertically causing significant morbidity and mortality in neonates. Neonatal HSV infection is infrequent with an incidence ranging from 1 in 3,500 to 1 in 20,000, depending on the population. Neonatal HSV infection is much more frequent in infants born to mothers experiencing a primary HSV infection with an incidence approaching 50%, while infants born to mothers experiencing recurrent HSV infection have an incidence of less than 3%. Neonatal infections are clinically categorised according to the extent of the disease. They are: (i) skin, eye and mouth (SEM) infections; (ii) central nervous system infection (encephalitis)- neonatal encephalitis can include SEM infections; and (iii) disseminated infection involving several organs, including the liver, lung, skin and/or adrenals. The central nervous system may also be involved in disseminated infections. Caesarean section, where the amniotic membranes are intact or have been ruptured for less than 4 hours, is recommended for those women who have clinical evidence of active herpes lesions on the cervix or vulva at the time of labour. This procedure significantly decreases the risk of transmission to the infant. Diagnosis of neonatal infection requires a very high level of clinical awareness as only a minority of mothers will have a history of genital HSV infection even though they are infected. Careful physical examination and appropriate investigations of the infant should accurately identify the infection in the majority of cases. Treatment is recommended where diagnosis is confirmed or there is a high level of suspicion. The current recommendation for treatment is aciclovir 20 mg/kg 3 times daily by intravenous infusion. Careful monitoring of hydration and renal function as well as meticulous supportive care of a very sick infant is also required. The newer anti-herpes agents, valaciclovir and famciclovir, offer no advantage over aciclovir and are not recommended for neonatal HSV infection. Prognosis is dependent upon the extent of disease and the efficacy of treatment, with highest rates of morbidity and mortality in disseminated infections, followed by central nervous system infection and the least in SEM infection. However, SEM infection is associated with poor developmental outcome even in infants who do not have encephalitis. Studies to improve the outcome of SEM infection are in progress. Neonatal HSV infections, although being relatively uncommon, are associated with significant morbidity and mortality if unrecognised and specific treatment is delayed. Diagnosis relies on a high level of clinical suspicion and appropriate investigation. With early therapy, the prognosis for this infection is considerably improved. PMID- 11269642 TI - Eating disorders in children and adolescents: epidemiology, diagnosis and treatment. AB - Eating disorders such as anorexia nervosa (AN) and bulimia nervosa (BN) are increasingly prevalent among children and adolescents. Whereas AN has a peak age of onset in early to mid-adolescence, BN typically presents during or after late adolescence. There is a spectrum of eating disorders that can be categorised by the criteria in the fourth edition of the American Psychiatric Association's Diagnostic and Statistical Manual of Mental Disorders into AN, BN and 'eating disorder not otherwise specified (ED-NOS)'. The key clinical signs of AN are those of protein calorie malnutrition. In BN, signs of purging are also important. Despite marked physical changes, metabolic decompensation occurs late and when present is an indication for hospital admission. During refeeding, electrolyte disturbances, in particular hypophosphataemia, should be serially monitored. For females with AN, restoration of gonadotropins, oestradiol and resumption of menses is a cardinal indicator of nutritional recovery. Treatment should address the medical, nutritional and psychological needs of children and adolescents with eating disorders. No single professional can be proficient in all spheres. Children and adolescents with eating disorders are best managed by a 'team approach'. Treatment may occur in a variety of inpatient, daypatient or outpatient settings. The aims of medical treatment are to promote bodyweight gain and nutritional recovery. Psychiatric goals address the psychosocial precipitants, treat comorbid mood symptoms and assist the patient to develop alternative coping skills. The crude mortality of AN has decreased to around 6%. For children and adolescents, the morbidity from malnutrition is increased because of the biological changes that are interrupted. PMID- 11269640 TI - Cefpodoxime proxetil: a review of its use in the management of bacterial infections in paediatric patients. AB - Cefpodoxime proxetil is an oral third generation cephalosporin with a broad spectrum of antibacterial activity. The drug has in vitro activity against many common Gram-positive and Gram-negative pathogens associated with common paediatric infections, making the drug a useful option for empirical therapy. In randomised controlled trials conducted in children with acute otitis media, oral cefpodoxime proxetil 8 to 10 mg/kg/day (usually administered in 2 divided doses) for 5 to 10 days was at least as effective as standard regimens of amoxicillin/ clavulanic acid, cefixime, cefuroxime axetil or cefaclor as assessed by either clinical or bacteriological criteria. Cefpodoxime 8 to 10 mg/kg/day (administered in 2 divided doses) for 5 to 10 days was at least as effective as standard 10-day regimens of penicillin V in the treatment of children with pharyngitis and/or tonsillitis. Significant differences in favour of cefpodoxime proxetil were demonstrated in terms of clinical (1 study) and bacteriological (2 studies) criteria. The clinical efficacy of 5 days of treatment with cefpodoxime proxetil is similar to that of 10 days of treatment with penicillin V. In children with lower respiratory tract infections (primarily pneumonia), clinical and bacteriological efficacy rates achieved with cefpodoxime proxetil treatment were similar to those produced by cefuroxime axetil or amoxicillin/clavulanic acid in randomised controlled trials. Cefpodoxime proxetil also demonstrated clinical efficacy in paediatric patients with skin and soft tissue infections. In randomised studies that included both adults and children with a variety of infections (e.g. abscess, atheroma, furuncle and carbuncle, infected wounds, cellulitis), cefpodoxime proxetil showed efficacy similar to that of cefuroxime axetil or cefaclor. Cefpodoxime proxetil is well tolerated by paediatric patients, with adverse events (primarily gastrointestinal tract disturbances and skin rashes) that are consistent with those reported for other oral cephalosporins. CONCLUSION: Cefpodoxime proxetil is a third generation cephalosporin with a broad spectrum of antibacterial activity and a favourable pharmacokinetic profile which allows twice-daily administration. It is generally well tolerated and demonstrates good bacteriological and clinical efficacy in paediatric patients with various infectious diseases, including acute otitis media, tonsillitis and/or pharyngitis. Based on these characteristics, cefpodoxime proxetil is a suitable option for the treatment of paediatric patients with various common bacterial infections. PMID- 11269643 TI - cDNA arrays and the enigma of melanoma immune responsiveness. AB - Melanoma is a cancer characterized by predisposition to natural and induced immune rejection. The occurrence of this phenomenon, however, remains sporadic and capricious while there is no clear understanding of the chain of events responsible. With the progression of the understanding of the molecular immunology of melanoma during the last decade, several theories suggesting immune tolerance (inadequate immune response) and/or immune escape (adaptation of cancer cells to a vigorous immune pressure) as possible modulators of melanoma growth have emerged. As the number of these hypotheses, mostly based on molecular or pre clinical models, has increased exponentially, the relevance of each individual one has been conversely decreasing leaving the observer without confidence that a theory likely to explain this phenomenon can be soon formulated. In this review, we will discuss a direct approach whereby new theories might be identified by direct observation of human phenomena as they occur by depiction of molecular portraits of melanoma lesions using global transcript analysis. PMID- 11269644 TI - DNA microarrays in pediatric cancer. AB - Childhood cancer, like all cancer, is at heart a genetic disease. Consequently, fundamental understanding of the oncogenic process is likely to be beneficially addressed by genetic methodology. Current methods have largely focused on single gene defects, like chimeric genes, which are present in many sarcomas and leukemias. Real understanding is more likely to derive from a genome-wide analysis of these malignancies. Recent technologic advances have made it possible to simultaneously assess the entire expressed gene profile, or transcriptome, of a given cancer. Foremost among these methods is gene expression profiling using DNA microarrays. Two basic approaches predominate: spotted arrays and photolithography arrays. Regardless of the method, the resulting information can be used to create disease profiles, but only if appropriate bioinformatic solutions are employed. Common analytic approaches include two-way expression comparisons, or scatter analyses; outlier gene analysis, to identify significantly dysregulated genes; dendrogram analyses, as pioneered by Eisen; cluster analyses to identify diagnostic or biologic groups; and various forms of functional analyses to identify relevant genes and biologic pathways. Studies of both adult and pediatric cancer have demonstrated the feasibility of such analyses to identify both diagnostic and prognostic groups of tumors. Acute childhood leukemias have been grouped into myelogenous and lymphoid, and even B- and T-cell subsets. Breast cancer prognostic groups have been identified on the basis of a small subset of expressed genes. In addition, preliminary data on childhood sarcomas appear to identify both diagnostic and prognostic subsets. Specifically, embryonal rhabdomyosarcoma could be distinguished from alveolar rhabdomyosarcoma, and even morphologically mixed embryonal and alveolar rhabdomyosarcoma showed similar gene expression profiles in both histologies. Further, collaborative studies using clustering analyses appear to identify prognostic groups of diverse sarcomas. Larger institutional and cooperative group studies are currently underway to validate these preliminary findings. PMID- 11269645 TI - Tissue microarray: a new technology for amplification of tissue resources. AB - Tissue microarrays are a method of harvesting small disks of tissue from a range of standard histologic sections and placing them in an array on a recipient paraffin block such that hundreds of cases can be analyzed simultaneously. This technique allows maximization of tissue resources by analysis of small-core biop sies of blocks, rather than complete sections. Using this technology, a carefully planned array can be constructed with cases from pathology tissue block archives, such that a 20-year survival analysis can be performed on a cohort of 600 or more patients by use of only a few microliters of antibody in a single experiment. The reflex criticism of this technique is that the tissue analyzed is not representative, especially in antigens with heterogeneous staining patterns. This review addresses this issue, as well as the issue of antigen preservation or durability, which validates construction of arrays from archives. Strategies and methods of construction and analysis of the arrays are discussed, as well as some other unusual array applications. This technique can provide a highly efficient, high-throughput mechanism for evaluation of protein expression in large cohorts. It has the potential to allow validation of new genes at a speed comparable to the rapid rate of gene discovery afforded by DNA microarrays. PMID- 11269646 TI - The role of tissue microdissection in cancer research. AB - Tissue microdissection is a laboratory method that is used to procure specific cells or cell populations from a histology slide under direct microscopic visualization. The recovered cells can be studied with a variety of DNA, messenger RNA, and protein analysis methods, including new high-throughput gene expression and proteomics technologies. This approach is permitting investigators to comprehensivelyexamine the molecular anatomy of cells in tissue sections forthe first time. This article reviews the development and evolution of tissue microdissection techniques, summarizes examples of research studies, and discusses related challenges that the research community must address. Additional information and complete laboratory protocols are available on a website at http://cgap-mf.nih.gov/. PMID- 11269647 TI - Implications of oncogenomics for cancer research and clinical oncology. AB - The explosion of information generated by large-scale functional genomics technologies has resulted in an exponential increase in the number of potential genes and proteins available for pharmaceutical and diagnostic research development. In order to tap this potential, the primary challenge is to develop a strategy to effectively integrate and extract meaning from the human genomic sequence information that has been generated since the start of the Human Genome Project. This article deals with the strategies being applied by academics and by the biotechnology sector to sort and triage this information with the ultimate goal of identifying future therapeutic targets for cancer and other diseases. PMID- 11269648 TI - A new cancer genome anatomy project web resource for the community. AB - The National Cancer Institute's Cancer Genome Anatomy Project (CGAP) is developing publicly accessible information, technology, and material resources that provide a platform for the interface of cancer research and genomics. CGAP's efforts have focused toward (1) building and annotating catalogues of genes expressed during cancer development, (2) identifying polymorphisms in those genes, and (3) developing resources for the molecular characterization of cancer related chromosomal aberrations. To date, CGAP has produced more than 1,000,000 expressed sequence tags, approximately 3,300,000 serial analysis of gene expression tags, and identified more than 10,000 human gene-based single nucleotide polymorphisms. To enhance access to these datasets by the research community, a new Cancer Genome Project web site (http://cgap.nci.nih.gov/) is being introduced. The web site includes genomic data for humans and mice, including transcript sequence, gene expression patterns, single-nucleotide polymorphisms, clone resources, and cytogenetic information. Descriptions of the methods and reagents used in deriving the CGAP datasets are also provided. An extensive suite of informatics tools facilitates queries and analysis of the CGAP data by the community. One of the newest features of the CGAP web site is an electronic version of the Mitelman Database of Chromosome Aberrations in Cancer. PMID- 11269649 TI - The genealogic approach to human genetics of disease. AB - The goal of modern human genetics is to correlate genes with disease or, more specifically, relate genetic variation to phenotypic variation. Although this correlation is usually straightforward in the Mendelian disorders, it has proved to be much more difficult to find in the common diseases because they appear to be more complex, likely involving an interplay among multiple genes and between genes and the environment. Although the strategy of linkage mapping of families was very successful when it was applied to the rare monogenic diseases, few common diseases have been mapped to statistical significance. Many investigators are now abandoning linkage analysis altogether and are moving to a candidate gene case-control strategy. In this article, we describe a genealogic approach to mapping human disease genes and provide three examples of how we have used it to map common diseases to statistical significance. We focus on a simple population with little historic migration and use a computerized genealogy database to increase the number of patients who can be compared with other affected relatives through high-density microsatellite genotyping. The genealogy helps determine which phenotypic classification is inherited and therefore possible to map. It may represent a more efficient strategy than candidate gene case-control studies for determination of what alleles or haplotypes are shared by patients in a population. We suggest that the genetics community not give up on linkage analysis, nor should it assume that the common diseases are too complex to map. PMID- 11269650 TI - Cancer proteomics: from biomarker discovery to signal pathway profiling. AB - In the post-genomic era of science, the field of proteomics promises the discovery of new molecular targets for therapy, biomarkers for early detection, and new endpoints for therapeutic efficacy and toxicity. Patient-specific targeted therapeutics with reduced toxicity and increased efficacy, the ultimate goal for rational drug development, can only be achieved if direct analyses of the tissue cells in the actual human malignancy are analyzed. To that end, technologies such as Laser Capture Microdissection (LCM), is providing unparalleled access to the purified diseased human cells directly from tissue specimens. However, limited availability of patient material is a challenge towards the development of new highly sensitive proteomic methodologies. Two dimensional gel electrophoresis (2D-PAGE), still the mainstay of most proteomic analysis of disease, is being complemented, and in some instances replaced by new exciting approaches to multiparametric protein characterization. The use of rapid, high throughput mass spectrometric-based fingerprints of peptides and proteins may prove to be valuable for new molecular classification of human tumors and disease stages. Coupled with LCM, high-density protein arrays, antibody arrays, and small molecular arrays, could have a substantial impact on proteomic profiling of human malignancies. Finally, detailed real-time knowledge about the states of intracellular signaling circuitry and pathways in the normal and malignant cells before, during and after therapy will yield invaluable information about mechanism of action and efficacy of existing and novel therapeutics for the treatment of human cancer. PMID- 11269651 TI - Combinatorial chemistry in cancer drug development. AB - The field of combinatorial chemistry has grown at an enormous rate in recent years, both in response to high-throughput capabilities and the discovery of a plethora of novel therapeutic targets. This review attempts to outlinethe recent developments of combinatorial chemistry in the search for novel cancer-related therapeutic agents. PMID- 11269652 TI - Detection and quantification of protein phosphatase inhibitor-1 gene expression in total rat liver and isolated hepatocytes. AB - The mRNA expression of protein phosphatase inhibitor-1 (inhibitor-1) in rat liver was demonstrated using highly sensitive semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Quantification by real-time RT-PCR (LightCycler technology) yielded the same copy number of inhibitor-1 mRNA in total rat liver and isolated hepatocytes (12 copies per cell). This novel finding shows that rat liver expresses indeed inhibitor-1 mRNA, albeit in low amounts. The low copy number explains why the mRNA had not been detected by Northern blotting so far. For comparison, about 425 copies/cell were detected in brain and 2500 copies/cell in skeletal muscle from rat. The full-length coding sequence of rat liver inhibitor-1 was cloned and sequenced, 100% homology with the muscle cDNA was obtained, indicating the expression of the same gene in liver and muscle. In vitro transcription and translation yielded a protein (Mr approximately 30 kDa) which could be detected with a specific antibody by immunoblotting. This indicates an intact open reading frame of inhibitor-1 in rat liver. Immunoblotting of liver extract yielded a very weak band which comigrated with the inhibitor-1 proteins from muscle and brain. It is concluded that mRNA expression of inhibitor-1 may have implications for the regulation of protein phosphatase-1 (PP1) in rat liver. PMID- 11269653 TI - Levels of IL-8 and myeloperoxidase in the lungs of pneumonia patients. AB - Interleukin-8 (IL-8) is considered as the major polymorphonuclear neutrophils (PMNs) chemoattractant cytokine in lung diseases such as asthma and adult respiratory distress syndrome (ARDS). However, controversial results were obtained regarding the involvement of IL-8 in the pathogenesis of pneumonia. This study examines the role of IL-8 in the recruitment and activation of PMNs in the lung of pneumonia patients. The interesting aspect of this study is that it is a site- specific analysis of the infected and uninfected lungs of the same patient. The level of IL-8 mRNA, protein and myeloperoxidase present in the cells of the bronchioalveolar lavages (BALs) taken from the areas of known pneumonic consolidations on chest X-ray (infected lung) are compared with the BALs obtained from areas of no obvious infiltrate (non-infected lung). The results obtained from the infected and non-infected lungs of pneumonic patients were further compared with that of a control group of non-smoking patients. The level of IL-8 mRNA and protein were determined by RT-PCR and ELISA respectively. There was a significant increase in the level of IL-8 mRNA in the infected lung as compared to its level in the non-infected lung (p < 0.001). In correlation with the increase in mRNA, IL-8 protein concentrations in BAL fluids from the infected lung were 6 fold higher than those taken from the non-infected lung (p < 0.0001). This pattern was also consistent with MPO activity in the BALs (4.5 fold more MPO activity in the infected lung as compared to that of the non-infected lung), indicating that IL-8 is directly implicated in neutrophil accumulation that follows acute respiratory infection. The results of the present study, therefore, indicate the involvement of IL-8 in the pathogenesis of pneumonia. PMID- 11269654 TI - Involving of the cytoplasmic region of leukemia inhibitory factor receptor alpha subunit, IL-6 related signal transducer-gp130 or fas death domain for MAPK p42/44 activation in HL-60 cell with LIF or anti-Fas IgG. AB - The chimeric receptors were prepared by exchanging the cytoplasmic region between leukemia inhibitory factor (LIF) receptor alpha subunit (gp190) and the other subunit-gp130 (190/130,130/190) and separately transduced into leukemia line HL 60 (to have the wild type subunit). The purpose is to investigate which subunit for activating MAPK p42/44 in leukemia cell while the cytoplasmic region homodimerization (190cyt-190cyt, 130cyt-130cyt) was induced by LIF. The results showed that MAPK p42/44 expression level after LIF stimulation 5 h was lower in the transformants with pED 130/190 (190cyt- 190cyt) (p < 0.01) and higher in the transformants with pED 190/130 (130cyt- 130cyt) (p < 0.05) than those in the parent cells. Meanwhile, MAPK p42/44 phosphorylation (Thr202/Tyr204) was ascended and the highest at 10 min in the 190/130 and descended in the 130/190. It suggests that gp130 activate MAPK p42/44 and gp190 indirectly regulate its expression and function. In order to analyses the relation of the subunit oligomerization and MAPK p42/44 we also prepared the recombination of the extracellular and transmembrane region of Fas and the cytoplasmic region of each LIFR subunit (Fas/190, Fas/130). After transduction into HL-60 with lipofection and induction by anti-Fas IgG, we found that MAPK p42/44 expression levels were lower in the Fas/190 than in the Fas/130 and parent cells (p < 0.01) and no difference between the Fas/130 and the wild type receptor. However, phospho-MAPK p42/44 were increased in the Fas/130 than the parent cells. It suggests that the oligomerization of the cytoplasmic regions of gp130 be potential to normally initiate MAPK p42/44 for the signal of HL-60 proliferation. We also determine that the separated oligomerization FasDD (no dimerization) can initiate the corresponding signal molecules, then regulate MAPK p42/44 expression and phosphorylation in leukemia cells. PMID- 11269655 TI - In vivo effects of vanadium on GLUT4 translocation in cardiac tissue of STZ diabetic rats. AB - The effect of vanadium treatment on insulin-stimulated glucose transporter type 4 (GLUT4) translocation was studied in cardiac tissue of streptozotocin (STZ) induced diabetic rats by determining the subcellular distribution of GLUT4. Four groups of rats were examined: control and diabetic, with or without bis(maltolato)oxovanadium(IV) (BMOV, an organic form of vanadium) treatment for 8 weeks. The effect of vanadium on insulin-induced GLUT4 translocation was studied at 5 min as the early insulin response and at 15 min after insulin injection as the maximal insulin response. At 5 min after insulin injection, plasma membrane GLUT4 level in the diabetic-treated group was not different from the control groups and was significantly higher than that of the insulin-stimulated diabetic group, indicating an enhancement of insulin response on GLUT4 translocation brought about by vanadium treatment. In contrast to that at 5 min after insulin injection, no significant difference in the plasma membrane GLUT4 level was observed between the diabetic and the diabetic-treated groups at 15 min after insulin injection. GLUT4 mobilization from the intracellular pool in response to insulin was also investigated at 15 min after insulin injection. Basal intracellular GLUT4 content was significantly higher in the diabetic-treated group when compared to the diabetic group under the same condition. However, the increased basal intracellular GLUT4 in the diabetic-treated group did not result in more insulin-mediated GLUT4 translocation at 15 min after insulin injection. In conclusion, the finding that plasma membrane GLUT4 in the diabetic-treated group is significantly higher than that of the diabetic group at 5 min but not at 15 min post-insulin injection indicates that vanadium treatment enhances insulin mediated GLUT4 translocation in cardiac tissue by enhancing its early response. PMID- 11269656 TI - Modulation of MLC-2v gene expression by AP-1: complex regulatory role of Jun in cardiac myocytes. AB - Hypertrophic stimulation of cardiac myocytes results in rapid induction of a number of transcription factors, including members of the AP-1 family, which is followed by a programmed alteration in the pattern of gene expression. In the ventricular cardiocytes there is re-expression of the fetal atrial natriuretic factor (ANF) gene and upregulation of its myosin light chain-2 (MLC-2v). The mechanism(s) by which the induction ofAP-1 is coupled to the promoters of these target genes is largely unknown. In this report, we demonstrate that in transient co-transfection assay, c-Jun inhibited while Jun B stimulated the MLC-2v promoter activity. Mutant c-Jun recombinants, in which the activation domains were deleted, still remained inhibitory, but a specific mutation in the leucine zipper, which changes the alignment of Jun with its dimerization partner, caused a reversal of its effect on the target MLC-2v promoter. Based on these findings, we propose that in chicken cardiac myocytes, the regulation of MLC-2v promoter by Jun may occur via its interaction with other proteins, possibly of the leucine zipper family. PMID- 11269657 TI - Arsenite inhibits Ras-dependent activation of ERK but activates ERK in the presence of oncogenic Ras in baboon vascular smooth muscle cells. AB - Exposure to arsenical compounds enhances the risk of atherosclerosis. The reason is unknown but it might be because an effect of arsenite (As3+) on plaque smooth muscle cells (SMCs) activation of extracellular signal-regulated kinase (ERK), a crucial mediator of SMC function. We found that arsenite inhibits the activation of ERK by platelet-derived growth factor-BB (PDGF-BB). This inhibitory effect depends on the time of arsenite exposure, is reversible, and is attenuated by preincubation of SMCs with the antioxidant N-acetyl-cysteine. These observations are consistent with the assumption that oxidative stress is involved. The blockade of ERK by arsenite may be mediated by an inhibition of Ras as arsenite prevents GTP-loading of Ras in response to PDGF-BB. Moreover, the Ras blockade by arsenite is not specific for PDGF-BB because it was also observed following stimulation of SMCs with EGF. To address the role of Ras, we expressed constitutively active, GTP-bound Ha-Ras (V12Ras). Unexpectedly, in V12Ras expressing-SMCs, arsenite stimulates ERK, but still decreases ERK activity in the presence of PDGF-BB. Our data suggest that arsenite inhibits the Ras/ERK pathway in SMCs, and that arsenite may activate ERK in Ras-transformed cells by mechanisms different from those employed by growth factors. PMID- 11269658 TI - Valine 77 of heterocystous ferredoxin FdxH2 in Anabaena variabilis strain ATCC 29413 is critical for its oxygen sensitivity. AB - Ferredoxins are small iron sulfur proteins necessary for electron donation. FdxH1 and FdxH2 are associated with two different nif gene clusters where they transfer electrons for the reduction of nitrogenase complex. FdxH1 was observed to be stable towards oxygen, whereas, FdxH2 was relatively unstable. We had identified the amino acid involved in oxygen sensitivity of ferredoxin protein using protein modeling. The exchange of valine to leucine at position 77 was critical for ferredoxin proteins in relation to its oxygen sensitivity. This exchange leads to a longer side chain, which inhibits the accessibility of oxygen to the iron sulfur cluster. Site directed mutagenesis and in vitro experiments confirms that valine indeed is involved in the oxygen sensitivity. The exchange of leucine to valine in FdxH1 makes it oxygen unstable. Thus, from the above results we can conclude that the position of leucine at position 77 is critical for oxygen sensitivity of ferredoxin and protein modeling can be used to identify specific amino acids in other oxygen-sensitive proteins. PMID- 11269659 TI - Cardioprotective effects of adenosine A1 and A3 receptor activation during hypoxia in isolated rat cardiac myocytes. AB - Adenosine (ADO) is a well-known regulator of a variety of physiological functions in the heart. In stress conditions, like hypoxia or ischemia, the concentration of adenosine in the extracellular fluid rises dramatically, mainly through the breakdown of ATP. The degradation of adenosine in the ischemic myocytes induced damage in these cells, but it may simultaneously exert protective effects in the heart by activation of the adenosine receptors. The contribution of ADO to stimulation of protective effects was reported in human and animal hearts, but not in rat hearts. The aim of this study was to evaluate the role of adenosine A1 and A3 receptors (A1R and A3R), in protection of isolated cardiac myocytes of newborn rats from ischemic injury. The hypoxic conditions were simulated by exposure of cultured rat cardiomyocytes (4-5 days in vitro), to an atmosphere of a N2 (95%) and CO2 (5%) mixture, in glucose-free medium for 90 min. The cardiotoxic and cardioprotective effects of ADO ligands were measured by the release of lactate dehydrogenase (LDH) into the medium. Morphological investigation includes immunohistochemistry, image analysis of living and fixed cells and electron microscopy were executed. Pretreatment with the adenosine deaminase considerably increased the hypoxic damage in the cardiomyocytes indicating the importance of extracellular adenosine. Blocking adenosine receptors with selective A1 and A3 receptor antagonists abolished the protective effects of adenosine. A1R and A3R activation during the hypoxic insult delays onset of irreversible cell injury and collapse of mitochondrial membrane potential as assessed using DASPMI fluorochrom. Cardioprotection induced by the A1R agonist, CCPA, was abolished by an A1R antagonist, DPCPX, and was not affected by an A3R antagonist, MRS 1523. Cardioprotection caused by the A3R agonist, Cl-IB-MECA, was antagonized completely by MRS 1523 and only partially by DPCPX. Activation of both A1R and A3R together was more efficient in protection against hypoxia than by each one alone. Our study indicates that activation of either A1 or A3 adenosine receptors in the rat can attenuate myocyte injury during hypoxia. Highly selective A1R and A3R agonists may have potential as cardioprotective agents against ischemia or heart surgery. PMID- 11269660 TI - The importance of glycolytically-derived ATP for the Na+/H+ exchange activity in guinea pig ventricular myocytes. AB - The cardiac subtype of Na+/H+ exchanger (NHE-1) plays an important role in the regulation of intracellular pH (pHi) and also can be a major route for Na+ influx. Although intracellular ATP is required for the optimal function of NHE-1, the regulation of the exchanger by ATP is less well characterized. This study was designed to investigate which intracellular ATP generated by oxidative phosphorylation or by glycolysis is dominant for the activation of NHE-1 in intact cardiac myocytes. Isolated guinea pig ventricular myocytes were loaded with the pHi-sensitive fluorescent indicator, 2'-7'-bis(carboxyl)-5',6'-carboxy fluorescein (BCECF), and exposed to 20 mM 2-deoxyglucose (2-DG) or 2 mM sodium cyanide (CN) to inhibit glycolysis or oxidative phosphorylation, respectively. The activity of NHE-1 was estimated with pHi recovery following transient application of 15 mM NH4Cl (NH4Cl prepulse). After the NH4Cl prepulse, pHi decreased from 7.00 +/- 0.03 (mean +/- S.E.) to 6.60 +/- 0.06 and recovered to 6.94 +/- 0.13 at 10 min (n = 7). The pHi recovery was suppressed in the presence of 2-DG (6.67 +/- 0.05, p < 0.01, n = 7), but was not changed in the presence of CN (6.88 +/- 0.18, n = 6). Since there was no difference in the intrinsic H+ buffering power, the estimation of the net acid efflux demonstrated that the activity of NHE-1 was significantly depressed in 2-DG. The inhibitory effect of 2 DG was not due to more extensive depletion of global intracellular ATP or secondary to the change in either intracellular Na+ or Ca2+ concentration. We concluded that ATP generated by glycolysis rather than by oxidative phosphorylation is essential to activate NHE-1 in ventricular myocytes. PMID- 11269661 TI - Three dimensional atomic model and experimental validation for the ATP-Regulated Module (ARM) of the atrial natriuretic factor receptor guanylate cyclase. AB - Atrial natriuretic factor (ANF) receptor guanylate cyclase (ANF-RGC) is a single chain transmembrane-spanning protein, containing both ANF binding and catalytic activities. ANF binding to the extracellular receptor domain activates the cytosolic catalytic domain, generating the second messenger cyclic GMP. Obligatory in this activation process is an intervening transduction step, which is regulated by the binding of ATP to the cyclase. The partial structural motif of the ATP binding domain of the cyclase has been elucidated and has been termed ATP Regulatory Module (ARM). The crystal structures of the tyrosine kinase domains of the human insulin receptor and haematopoietic cell kinase were used to derive a homology-based model of the ARM domain of ANF-RGC. The model identifies the precise configuration of the ATP-binding pocket in the ARM domain, accurately represents its ATP-dependent features, and shows that the ATP-dependent transduction phenomenon is a two-step mechanism. In the first step, ATP binds to its pocket and changes its configuration; in the second step, via an unknown protein kinase, it phosphorylates the cyclase for its full activation. PMID- 11269663 TI - The effect of tyrosol, hydroxytyrosol and oleuropein on the non-enzymatic lipid peroxidation of rat liver microsomes. AB - The effects of the polyphenolic compounds from virgin olive oil: tyrosol, hydroxytyrosol and oleuropein on the non-enzymatic lipid peroxidation induced by ascorbate-Fe2+ of rat liver microsomes were examined. The inhibition of light emission (maximal induced chemiluminescence) by oleuropein was concentration dependent. Hydroxytyrosol showed a substantial degree of inhibition against ascorbate-Fe2+ induced lipid peroxidation in rat liver microsomes that was at least 6 times higher than that observed in the presence of oleuropein. Inhibition of lipid peroxidation by tyrosol was not observed. In rat liver microsomes incubated alone or in the presence of tyrosol, the fatty acid composition was profoundly modified when subjected to in vitro peroxidation mediated by ascorbate Fe2+, with a considerable decrease of 18:2n6 and 20:4n6; however, changes in fatty acid composition were not observed when microsomes were incubated with hydroxytyrosol. When oleuropein was used at low concentration (5, 15 microM) a considerable decrease of 20:4n6 was observed, but 18:2n6 was not modified; at higher concentration (30, 60 microM) changes in fatty acid composition were not observed. There was a very good correlation between the presence of oxidized phospholipids and the changes in polyunsaturated fatty acids previously observed. Thus, hydroxytyrosol showed the highest protection again oxidized phospholipid formation. The presence of oleuropein at low concentration (5, 15 microM) does not prevent the formation of oxidized phospholipids (8.02 +/- 1.22 and 1.22 +/- 1.22) but concentration higher than 30 microM avoids completely the formation of this molecules whereas tyrosol at any concentration assayed was found to be ineffective and allows the formation not only of oxidized phospholipids but also of oxidized cholesterol. PMID- 11269662 TI - Accelerated HER-2 degradation enhances ovarian tumor recognition by CTL. Implications for tumor immunogenicity. AB - We investigated the ubiquitination and degradation of a tumor antigen, the HER 2/neu (HER-2) protooncogene product which is overexpressed in epithelial cancers. HER-2 degradation was investigated in the ovarian tumor line, SKOV3.A2, that constitutively overexpressed long-life HER-2. We used as agonist geldanamycin (GA), which initiated downmodulation of HER-2 from the cell surface. HER-2 was polyubiquitinated and degraded faster in the presence than in the absence of GA. GA did not decrease HLA-A2 expression. Presentation of the immunodominant cytotoxic T lymphocyte (CTL) epitope, E75 (369-377) from SKOV.A2 was inhibited by proteasome inhibitors, such as LLnL but was enhanced by cysteine protease inhibitors such as E64, indicating that both the proteasome and cysteine proteases are involved in epitope formation but have different effects. Enhanced tumor recognition was not an immediate or early effect of GA treatment, but was evident after 20 h of GA treatment. In contrast, 20 h GA treatment did not increase tumor sensitivity to LAK cell lysis. Twenty hour GA-treated SKOV3.A2 cells expressed an unstable HER-2 protein synthesized in the presence of GA, of faster electrophoretic mobility than control HER-2. This suggested that the newly synthesized HER-2 in the presence of GA was the main source of epitopes recognized by CTL. Twenty hour GA-treated SKOV3.A2 cells were better inducers of CTL activity directed to a number of HER-2 CTL epitopes, in peripheral blood mononuclear cells compared with control untreated SKOV3.A2 cells. Thus, induction of HER-2 protein instability enhanced the sensitivity of tumor for CTL lysis. Increased HER-2 CTL epitopes presentation may have implications for overcoming the poor immuno-genicity of human tumors, and design of epitope precursors for cancer vaccination. PMID- 11269664 TI - Modulation of biosynthesis of phosphatidylcholine via CDP-choline in rat liver: influence of ethanol on the microsomal cholinephosphotransferase activity. AB - We have studied in vitro the effects of ethanol on the different enzymes involved in the biosynthesis of phosphatidylcholine (PC) via CDP-choline. Ethanol alters neither choline kinase (CK) nor CTP:phosphocholine cytidylyltransferase (CT) activities but, at levels higher than 50 mM, it does significantly inhibit microsomal cholinephosphotransferase (CPT) activity concomitantly with an increase in the ethanol concentration. A study of the kinetics of the reaction catalysed by CPT shows that ethanol decreases Vmax without altering Km, indicating a non-competitive inhibitory effect. An analysis of the thermodependence of CPT activity in the absence of ethanol reveals a break in the Arrhenius plot and thus a straight relationship between enzyme activity and the physico-chemical state of the microsomal membrane. Incubation of microsomes in the presence of ethanol increased the transition temperature from 25.8-28.2 degrees C. Microsomes were also incubated with n-alkanols with chain-lengths of fewer than five carbon atoms at concentrations which, according to their partition coefficients, produce equimolar levels in the membrane. Under these conditions all the alkanols caused the same inhibitory effect. All these results demonstrate that ethanol modulate the PC biosynthesis at the level of CPT activity and does not affect the CT enzyme. The inhibition found on CPT is clearly dependent on the alteration produced by ethanol on the hepatic microsomal membrane. PMID- 11269665 TI - Telomerase activation and incidence of HPV in human gastrointestinal tumors in North Indian population. AB - The present study was initiated with the objective of finding out the role of possible factors in the etiology of gastrointestinal tract cancers. HPV-DNA was detected in 62.5% (25/40) of the patients by PCR. Telomerase activity as shown by TRAP-ELISA assay was detected in 82.5% (33/40) of the tumor samples and absent in 85.7% (30/35) of the normal samples taken from the same patients. As many as 53.6% (15/28) of the invasive cases were positive both for telomerase activity and for HPV, while 39.3% (11/28) of them, although indicating telomerase expression, showed no signal for HPV. This suggests that activation of telomerase could be by a pathway independent of HPV activation, although both parameters could act as diagnostic and prognostic markers for gastrointestinal tract cancers. PMID- 11269666 TI - Differential expression of cytosolic and mitochondrial 3-hydroxy-3-methylglutaryl CoA synthases during adipocyte differentiation. AB - Mitochondrial and cytosolic 3-hydroxy-3-methylglutaryl CoA synthase (m-HMS and c HMS) genes show high identity at the nucleotide and amino acid level, but no homology has been found in the promoter area. The main regulator for c-HMS is SREBP. The best known transcription factor that regulates m-HMS is PPAR alpha. Three types of PPAR, alpha, gamma and delta have been described in vertebrates. Here we found that they display distinct ligand response profiles in the m-HMS promoter. In some conditions PPAR gamma is a significant activator of m-HMS. Thus, the m-HMS gene is transiently expressed during the clonal expansion phase of 3T3-L1 differentiation. We found that C/EBP delta and PPAR gamma activate the m-HMS promoter in 3T3-L1 cells synergistically. This synergistic effect was only observed in the whole promoter (-1148 to +28). A small construct (-116 to +28) which contains the PPRE did not respond to C/EBP delta and/or PPAR gamma. This suggests that a putative C/EBP site lie somewhere between -1148 and -116 bp. We also show that C/EBP delta was more efficient that C/EBP alpha and C/EBP beta to activate the m-HMS promoter. The time course of c-HMS mRNA expression during 3T3 L1 differentiation was different, with a significant increase at terminal adipogenesis. We found that the transcription factor C/EBP alpha did not activate the c-HMS promoter. The differential pattern of expression shown by these two genes, which have a common ancestor, exemplifies refinements of transcriptional control during evolution. PMID- 11269667 TI - A study on distribution of different hydroxyproline fractions in the bovine ocular tissues. AB - The purpose of this study was to determine the content of total, free, peptide bound, protein-bound, soluble- and insoluble collagen hydroxyproline (Hyp) in tissues of bovine eye. The results show that lens had the highest content of free Hyp. This was followed by cornea, retina, iris and aqueous humor. The difference between the Hyp content of lens and iris (p < 0.01) and aqueous humor (p < 0.001) was significant. The peptide-bound Hyp was highest in iris followed by cornea, ciliary body, sclera, lens, aqueous humor and retina. Significant differences (p < 0.001) was observed between the concentration of peptide-bound Hyp of iris and ciliary body, sclera, lens, aqueous humor and retina. Protein-bound Hyp was highest in iris, followed by ciliary body, sclera, cornea, lens, retina and aqueous humor. The difference between the protein-bound Hyp levels of iris and sclera, cornea, lens, retina and aqueous humor was significant (p < 0.001). No peptide-bound and protein-bound Hyp was detected in vitreous humor. Iris had the highest content of total Hyp. This was followed by cornea, ciliary body, sclera, lens, retina, vitreous humor and aqueous humor. The difference in the Hyp content of iris with ciliary body, sclera, lens, retina, vitreous humor and aqueous humor was significant (p < 0.001). Cornea had significantly (p < 0.001) higher content of soluble- and insoluble-collagen Hyp as compared to other tissues. This was followed by ciliary body, sclera, lens, iris and retina. Iris had the highest content of collagen. This was followed by cornea, ciliary body, sclera, lens, retina, vitreous humor and aqueous humor. The difference in the collagen content of iris with ciliary body, sclera, lens, retina, vitreous humor and aqueous humor was significant (p < 0.001). PMID- 11269668 TI - Inducible nitric oxide synthase in the myocard. AB - Recognition of significance of nitric oxide synthases (NOS) in cardiovascular regulations has led to intensive research and development of therapies focused on NOS as potential therapeutic targets. However, the NOS isoform profile of cardiac tissue and subcellular localization of NOS isoforms remain a matter of debate. The aim of this study was to investigate the localization of an inducible NOS isoform (NOS2) in cardiomyocytes. Employing a novel immunocytochemical technique of a catalyzed reporter deposition system with tyramide and electron microscopical immunocytochemistry complemented with Western blotting and RT-PCR, we detected NOS2 both in rat neonatal and adult cultured cardiomyocytes and in the normal myocard of adult rats as well as in the human myocard of patients with dilative cardiomyopathy. NOS2 was targeted predominantly to a particulate component of the cardiomyocyte--along contractile fibers, in the plasma membrane including T-tubules, as well as in the nuclear envelope, mitochondria and Golgi complex. Our results point to an involvement of NOS2 in maintaining cardiac homeostasis and contradict to the notion that NOS2 is expressed in cardiac tissue only in response to various physiological and pathogenic factors. NOS2 targeting to mitochondria and contractile fibers suggests a relationship of NO with contractile function and energy production in the cardiac muscle. PMID- 11269669 TI - L-carnitine reduces malondialdehyde concentrations in isolated rat hearts in dependence on perfusion conditions. AB - The study investigated the influence of L-carnitine on the formation of malondialdehyde, an indicator of lipid peroxidation, in isolated Langendorff rat hearts. Earlier investigations of hemodynamic parameters and the recovery of ATP and creatine phosphate, carried out by means of 31P-NMR spectroscopy, had demonstrated that, depending on the composition of the perfusates (content of glucose, fatty acids, and carnitine), quite strong differences may occur in the reperfusion period after ischemia. In order to determine a possible relationship between these differences and the addition of carnitine, the study investigated whether carnitine penetrated into the tissue during the experiments, and whether it was able to reduce the concentration of detrimental substances. The concentrations of free and total carnitine as well as the malondialdehyde content as an indicator of ischemia/reperfusion damage were determined in different parts of the cardiac tissue as follows: After the Langendorff-experiments the hearts were dissected, homogenized and reconditioned; then carnitine and malondialdehyde were determined. The study included 63 hearts, which were divided into 8 different perfusion groups. Carnitine concentrations in heart tissue perfused with L-carnitine were much higher than those of the controls. Since exogenous L carnitine and formed esters could be found in the tissue after the experiment, they must have permeated the cellular membrane rapidly. The concentrations of malondialdehyde behaved in an inverted way; as expected they were lower in carnitine-perfused hearts. The favourable effects of L-carnitine, expressed both by improved cardiac dynamics and ATP and CrP recovery in the reperfusion period, are obviously due to the fact that L-carnitine reduces ischemic damage. PMID- 11269670 TI - Expression of the mRNA encoding truncated PPAR alpha does not correlate with hepatic insensitivity to peroxisome proliferators. AB - Two alternatively spliced forms of human PPAR alpha mRNA, PPAR alpha1 and PPAR alpha2, have been identified. PPAR alpha1 mRNA gives rise to an active PPAR alpha protein while PPAR alpha2 mRNA gives rise to a form of PPAR which lacks the ligand-binding domain. PPAR alpha2 is unable to activate a peroxisome proliferator response element (PPRE) reporter gene construct in transient transfection assays. Both PPAR alpha1 and PPAR alpha2 mRNA are present in human liver, kidney, testes, heart, small intestine, and smooth muscle. In human liver, PPAR alpha2 mRNA abundance is approximately half that of PPAR alpha1 mRNA; a correlation analysis of PPAR alpha1 and PPAR alpha2 mRNA mass revealed an r-value of 0.75 (n = 18). Additional studies with intact liver from various species, showed that the PPAR alpha2/PPAR alpha1 mRNA ratios in rat, rabbit, and mouse liver were less than 0.10; significantly lower than the 0.3 and 0.5 ratios observed in monkey and human livers, respectively. To determine if a high PPAR alpha2/PPAR alpha1 mRNA ratio was associated with insensitivity to peroxisome proliferators, we treated human, rat, and rabbit hepatocytes with WY14643, a potent PPAR alpha activator, and measured acyl CoA oxidase (ACO) mRNA levels. Rat ACO mRNA levels increased markedly in response to WY14643 while human and rabbit hepatocytes were unresponsive. Thus, although the PPAR alpha2/PPAR alpha1 mRNA ratio is low in rabbits, this species is not responsive to peroxisome proliferators. Further studies with male and female rats, which vary significantly in their response to peroxisome proliferators, showed little difference in the ratio of PPAR alpha2/PPAR alpha1 mRNA. These data suggest that selective PPAR alpha2 mRNA expression is not the basis for differential species or gender responses to peroxisome proliferators. PMID- 11269671 TI - Alterations in the oxidative metabolic profile in vascular smooth muscle from hyperlipidemic and diabetic swine. AB - High cholesterol, especially LDL cholesterol, has been associated with the development of atherosclerotic plaques in arteries. To investigate the changes in cellular substrate metabolism early in the atherogenic process, Sinclair miniature swine were treated for 12 weeks with either a control diet, a high fat diet, or a high fat diet with the addition of alloxan to induce diabetes. The fractional entry into the TCA cycle of 1,2-(13)C-acetate (5 mM), 1-(13)C-glucose (5 mM), and unlabeled, endogenous lipids was determined in control, hyperlipidemic, and diabetic/hyperlipidemic pigs using 13C-isotopomer analysis of glutamate. The diabetic state of the pigs was validated by plasma glucose measurements made after 10 weeks of alloxan treatment for control (65 +/- 6 mg/dL), hyperlipidemic (63 +/- 5 mg/dL), and diabetic/hyperlipidemic (333 +/- 52 mg/dL) pigs. Plasma glucose values did not correlate with the percentage of glucose entry into the TCA cycle (R2 = 0.0819, n = 10). Alterations in the pattern of substrate oxidation were better correlated with changes in plasma lipids (cholesterol and triglycerides) than with changes in plasma glucose. Plasma total cholesterol and total triglyceride levels significantly correlated with changes in acetate metabolism (R2 = 0.7768 and R2 = 0.4787, respectively) and with changes in glucose metabolism (R2 = 0.6067 and R2 = 0.4506, respectively). We conclude that alterations in lipid profile, especially those that were observed in the diabetic milieu, are associated with early changes in vascular smooth muscle oxidative metabolism. These changes in oxidative metabolism may precede alterations in smooth muscle phenotype and, therefore, may play an important role in the early pathogenesis of atherosclerosis. PMID- 11269672 TI - An analysis of cardiac mortality in patients with new-onset end-stage renal disease in New York State. AB - End-stage renal disease (ESRD) is associated with an overall one-year mortality of 23.5% in the US, of which cardiac causes constitute 50% of all deaths. Data on incident ESRD patients were obtained from the Health Care Financing Administration's 2728 and 2746 forms by special request from the ESRD Network of New York. 4,948 ESRD patients, who started dialysis in New York State from April 1, 1995, through April 1, 1996, were assessed to identify risk factors present at the initiation of dialysis that predict cardiac death. 899 deaths were registered during the 19-month-follow-up period, 50% of which were from cardiac causes. Using the Cox-proportional hazards model, the increasing age category, white race, the presence of one or more vascular co-morbid conditions, and the presence of diabetes and one or more cardiac co-morbid conditions significantly predicted cardiac death (p < 0.05). Diabetes increased the risk for cardiac death by 48% for those patients without any cardiac co-morbidities (RR = 1.48, p < 0.0082). In contrast with results observed in the general population, gender, serum albumin and body mass index were not significant predictors of cardiac death. In identifying risk factors present at the initiation ofdialysis that predict cardiac death, this study highlights factors that may be modified prior to dialysis initiation in order to improve life expectancy and mortality rates and decrease health care costs for the ESRD population. PMID- 11269673 TI - Coronary artery disease as a definitive risk factor of short-term outcome after starting hemodialysis in diabetic renal failure patients. AB - Cardiovascular disease is a major cause of morbidity and mortality in patients with diabetes who are receiving dialysis. It is known that cardiac mortality is high in the first year of hemodialysis (HD). The purpose of this study was to clarify the prevalence of coronary artery disease (CAD) in patients with diabetes at the initiation of HD, and to investigate the relationship between short-term outcomes after starting hemodialysis and CAD. We performed coronary angiography (CAG), whether or not patient had angina, within one month of initiation of maintenance HD in 17 of 34 consecutive unselected diabetic patients. We studied the two-year survival rate of those with CAD. Eleven patients (65%) were classified CAD-positive. Nine (82%) of these 11 had multivessel disease. Clinical and hematologic factors did not differ significantly between the groups with and without CAD. During the two-year follow-up period, 28 (82%) of 34 patients survived. In patients with CAD the two-year survival rate was 60%. None of the patients without CAD died within 2 years after starting HD. These results suggest that the presence of CAD at the initiation of HD may play a critical role in short-term outcomes after starting HD. We believe that an evaluation of CAD should be performed at the initiation of HD. PMID- 11269674 TI - Effect of sympathetic and plasma renin activity on hemodialysis hypertension. AB - BACKGROUND: The reasons for poor control of blood pressure in hemodialysis (HD) patients are not clear, while patients have achieved their desirable dry weight and excess weight are not different between the hypertensive and normotensive patients. A link between sympathetic activity and HD hypertension could be an alternative explanation. PATIENTS AND METHODS: We studied the effect of sympathetic and plasma renin-aldosterone activity of 10 hypertensive patients, 5 men and 5 women, aged from 30 to 60 years. The results were compared to those of another 10 normotensive hemodialysis patients. Blood samples were taken before HD and at the end of a 4-hour HD session for plasma aldosterone (ALDO), renin activity (PRA), adrenaline and noradrenaline determination. One month dialysis records, which includes 13 dialysis sessions and 26 blood pressure readings for each patient were used, for pre-dialysis and post dialysis mean arterial blood pressure (MAP) measurement. RESULTS: Pre-dialysis plasma adrenaline was 124.12 +/ 12.93 pg/ml vs. 121.12 +/- 14.71 pg/ml and plasma noradrenaline was 260.88 +/- 140.86 pg/ml vs. 138.11 +/- 122 +/- 32 pg/ml for hypertensive and normotensive patients, respectively. Post-dialysis plasma adrenaline and noradrenaline levels were 119.37 +/- 8.81 pg/ml vs. 120.62 +/- 15.35 pg/ml and 210.44 +/- 126.71 pg/ml vs. 94.88 +/- 64.05 pg/ml for hypertensive and normotensive patients, respectively. Pre-dialysis PRA was 8.70 +/- 6.37 ng/ml/h vs. 2.77 +/- 1.8 ng/ml/h and plasma ALDO was 457.07 +/- 245.54 pg/ml vs. 197.74 +/- 87.46 pg/ml for hypertensive and normotensive patients, respectively. Pre-MAP was 109.76 +/- 5.21 mmHg vs. 99.28 +/- 7.13 mmHg and post-MAP was 107.22 +/- 6.74 mmHg, vs. 91.61 +/- 7.27 mmHg for hypertensive and normotensive patients, respectively. Plasma ALDO and fluid volume removed by ultrafiltration were found to be significantly correlated (p < 0.05). PRA and plasma adrenaline-noradrenaline levels were not correlated with MAP or body weight alterations. CONCLUSIONS: It is suggested that sympathetic activity, as it was expressed by plasma catecholamine alterations, is not associated with hemodialysis hypertension. PMID- 11269675 TI - Increase of peripheral natural killer T cells in hemodialysis patients. AB - AIMS: Patients with end-stage renal disease often present a state of immunodeficiency. To elucidate the involvement of natural killer T (NKT) cells in the development of pathologies associated with uremia, we examined the percentages and characteristics of NKT cells (CD56+ T cells and CD57+ T cells) in peripheral lymphocites from hemodialysis (HD) patients. MATERIALS AND METHODS: Thirty-three patients who had undergone HD therapy for 2 to 5 years (group A, n = 15) or for more than 10 years (group B, n = 18) and 17 normal controls participated in this study. Cell surface antigens of lymphocytes were analyzed using immunofluorescent flow cytometry. RESULTS: The percentage of peripheral CD56+ T cells was increased in group A and B compared with controls, and that of CD57+ T cells was increased in group B compared with controls and group A. The most abundant population in CD56+ T cells was DN T cells, and that in CD57+ T cells was CD8+ T cells. In CD57+ T cells, a higher proportion of CD8+ T cells and a lower proportion of DN T cells were found in group B compared with controls. CONCLUSIONS: We detected increases of CD56+ T cells and CD57+ T cells in HD patients. The elevation of NKT cells might affect some complications associated with HD. PMID- 11269676 TI - Prevalence of biliary lithiasis in a Sicilian population of hemodialysis patients. AB - Studies performed to date on the prevalence of biliary lithiasis (BL) in chronic renal failure patients on hemodialysis (HD) have given contradictory results. The aims of the present study were to evaluate the prevalence of BL and its main associated risk factors in a population of hemodialysis patients, and to compare the results with those we had obtained previously in an overt population of the same zone. The study included 171 patients (83 M, 88 F), mean age 62.5 years and mean duration of dialysis 66.7 months. The screening protocol also included body mass index (BMI), a number of biochemical parameters and an ultrasound scan of the gallbladder and biliary tract. The general prevalence of BL was 33.3% (30.1% in men and 36.4% in women), and this figure was significantly higher than that found in our previous study. Prevalence increased with age in both sexes (Mantel Haenszel Chi-squared = 5.4, p < 0.03), but not with duration of dialysis. The main risk factors, evaluated with multiple logisstic regression, were the presence of diabetes mellitus and high serum phosphorus levels. Specific symptoms were also significantly associated in BL patients. No association was found with parity, BMI or serum lipid alterations. In conclusion, the prevalence of BL in a Sicilian population of HD patients was higher than that found in an overt population of the same area and the associated main risk factors were not coincident. Further studies are needed to establish the role played by the phase of end-stage renal disease before HD and to correct the metabolic disturbances to limit a high percentage of morbidity in a disease already in itself sufficiently disabling. PMID- 11269677 TI - Re-evaluating criteria for peritoneal dialysis in "classical" (D+) hemolytic uremic syndrome. AB - BACKGROUND: Indications for peritoneal dialysis in children with post-dysenteric hemolytic uremic syndrome (D+ HUS) have not been thoroughly evaluated. Although early institution of dialysis may reduce mortality, the procedure has attendant complications. AIM: To determine whether the use of more stringent criteria for instituting dialysis had a better outcome to that of using conventional criteria. METHOD: Following an outbreak of Shigella dysenteriae type 1 D+ HUS in KwaZulu/Natal during June 1994 to October 1995, we compared the renal outcome and mortality between two periods: before May 1995 (69 children) when conventional criteria for dialysis were employed and after May 1995 (70 children) when more stringent criteria for dialysis were applied. RESULTS: The mean age of presentation was 35 months, 79 (56.8%) were males. Both groups were comparable except for gut perforation, which was more frequent before May 1995, and hypertension and severe disease, which were more frequent after May 1995. Seventy patients underwent dialysis, 36 (52.2%) before May 1995. There were no significant differences in renal outcome or death following discharge from hospital in both groups. Overall mortality was 20.1%, 15 (53.6%) of the 28 children that demised presented before May 1995. OUTCOME: Accordingly, although children with more severe disease and a higher frequency of hypertension presented after May 1995, there were no significant differences in morbidity or mortality in those using stringent criteria for dialysis, compared to those in whom conventional criteria were used. CONCLUSION: We showed that several children, who would previously have been dialyzed, may be managed conservatively, without an increase in mortality or morbidity. PMID- 11269679 TI - Risk factors of cyclosporine nephrotoxicity after conversion from Sandimmune to Neoral. AB - BACKGROUND: In 1995 - 1996, we switched from a once-daily Sandimmune dose to a twice-daily dose regimen of Neoral. Concurrent with the switch we changed our target trough level from 100 microg/l at 24 hours to the generally accepted 12 hour level of 150 microg/l. We performed a retrospective cohort study to assess cyclosporine toxicity following this switch and to identify risk factors for nephrotoxicity. PATIENTS AND METHODS: Of 212 patients with a stable graft function pre-conversion clinical parameters at 1 and 12 months post-conversion were compared with those at time of conversion. Cyclosporine nephrotoxicity was defined as a significant decline of the reciprocal of the serum creatinine concentration over time post-conversion in the absence of other obvious causes for declining graft function. Risk factors of cyclosporine nephrotoxicity were assessed using logistic regression analysis. RESULTS: The mean cyclosporine trough level rose from 87 microg/l at the time of conversion to 139 microg/l at 12 months post-conversion whereas the daily drug dose increased over the same period from 233 mg to 252 mg. Mean serum creatinine increased by 10% from 135 to 148 micromol/l (p < 0.001). Cyclosporine nephrotoxicity was present in 42 patients (20%). Cyclosporine dose and trough level did not predict nephrotoxicity but beta-blockers (OR 0.35, 95% CI 0.17-0.72) and calcium channel blockers (OR 0.35, 95% CI 0.19-0.82) reduced the risk of nephrotoxicity, independent from an effect on blood pressure. CONCLUSION: 20% of stable renal transplant patients experienced chronic cyclosporine nephrotoxicity after conversion from a once daily Sandimmune regimen to a twice-daily Neoral regimen with dose adjustments to a trough level of 150 microg/l. beta-blockers and calcium channel blockers reduced the risk of nephrotoxicity. PMID- 11269678 TI - Initial remission-inducing effect of very low-dose cyclosporin monotherapy for minimal-change nephrotic syndrome in Japanese adults. AB - AIM: Cyclosporin A (CsA) in combination with corticosteroids can be used effectively in steroid-sensitive nephrotic syndrome. However, reports documenting the effectiveness ofCsA monotherapy against such a condition have been scarce. In 11 adults with minimal-change nephrotic syndrome, we have tried very low-dose CsA in the hope of inducing remission without using either corticosteroid or any other immunosuppressive drugs. PATIENTS AND METHODS: Indications for treatment included steroid-sensitive relapsing nephrotic syndrome (7 patients) and first episode nephrotic syndrome (4 patients). In all patients, corticosteroid and cytotoxic agents had not been given before entry. CsA was administered orally at an initial dose of 2.4 (range 1.5 - 3.1) mg/kg per day. RESULTS: Analysis of the clinical course revealed that 8 of 11 patients entered complete remission after a mean duration of 44 +/- 31 days, whereas 3 patients failed to enter remission to CsA alone, resulting in complete remission combined with methylprednisolone pulse therapy without conventional oral prednisolone. CsA dosages and trough levels between responders and non-responders were similar. Non-responders had much higher levels of serum total cholesterol and higher daily urinary excretion of protein than those of responders, respectively. No patients had significant decrease in creatinine clearance, development of hypertension or suffered from other CsA associated serious side-effects. CONCLUSION: The present data suggest that CsA monotherapy at a very low dose could induce complete remission in adult patients with minimal-change nephrotic syndrome. Conversely, severe hypercholesterolemia would be likely to inhibit the action of CsA against nephrotic conditions. PMID- 11269680 TI - High serum apolipoprotein AIV levels in renal transplant recipients. AB - BACKGROUND: Human apolipoprotein (apo) AIV might play a role in post-transplant reverse cholesterol transport, which appears to be comparable to that seen in healthy subjects. However, there may be subtle differences between healthy individuals and renal transplant recipients, given the other abnormalities of lipoprotein metabolism in the latter. Therefore, the aim of the present study was to investigate possible changes of serum apo AIV levels in renal transplant recipients, and to evaluate potential factors influencing these levels. PATIENTS AND METHODS: Total and free serum apo AIV was determined in 36 clinically stable renal transplant recipients and in 20 sex- and age-matched healthy control subjects. RESULTS: Mean total serum apo IV concentrations (+/- SD) were significantly higher in renal transplant recipients than in control subjects (202 +/- 102 vs 79 +/- 45 mg/l, p < 0.01). The percentage of lipoprotein-free fractions of apo AIV was comparable in both groups. The elevated total serum concentrations of apo AIV were mainly related to creatinine clearance and partially to serum triglyceride levels in renal transplant recipients. CONCLUSION: Our data suggest that the observed elevation of serum apo AIV concentrations in renal transplant recipients is essentially related to the presence of impaired renal function. PMID- 11269681 TI - Outcome of renal transplantation in fibrillary glomerulonephritis. AB - Fibrillary glomerulonephritis (FGN) is a rare but progressive glomerular disease usually with end-stage renal disease (ESRD) developing within months or few years following the diagnosis. Little is known about the outcome of renal transplantation in patients with ESRD due to FGN. We report four patients with FGN who received renal allografts. Two patients developed recurrent FGN in their grafts. One patient was diagnosed to have recurrent FGN 9 years post-transplant, and lost his graft 4 years thereafter. Another patient had recurrent disease 2 years post-transplant but has stable graft function after 7 years. One patient died with normal renal allograft function 7 years following transplantation. The fourth patient has chronic transplant nephropathy 34 months post-transplant without evidence of recurrent FGN. A literature review revealed 10 additional patients who received 11 renal allografts due to ESRD caused by FGN. Four of these 10 patients had biopsy-proven recurrence (one patient in two subsequent grafts), but this caused graft loss only in 2 patients 56 months and 7 years post transplant, respectively. The earliest recurrence was diagnosed 2 years post transplant. We conclude that although the recurrence rate of FGN in renal transplants is high (around 50%), the recurrent disease has a relatively benign course and prolonged graft survival is possible. PMID- 11269682 TI - Hemolytic uremic syndrome after capnocytophaga canimorsus (DF-2) septicemia. AB - A 66-year-old man developed a hemolytic uremic syndrome (HUS) with acute renal failure, thrombocytopenia, fragmented red cells in the blood film and elevated serum LDH following a capnocytophaga canimorsus (DF-2) infection after a dog bite. He was treated with antibiotics, plasmapheresis and hemodialysis. Although hematologic values improved, the patient remained hemodialysis-dependent for six months. In the literature several cases of renal failure following capnocytophaga canimorsus septicemia have been described, caused by hypotension or disseminated intravascular coagulation (DIC). In our patient there were no signs of hypotension or extensive DIC. A few case reports described HUS and thrombotic thrombocytopenic purpura (TTP) following DF-2 sepsis. PMID- 11269683 TI - Tubulointerstitial nephritis associated with acute intermittent porphyria. AB - Progressive renal impairment associated with acute intermittent porphyria is not well recognized and the mechanism of renal damage remains unclear. We report a case of a 51-year-old female with acute intermittent porphyria and long-term follow-up who developed proteinuria and renal insufficiency. Her biopsy showed marked tubulointerstitial damage with mitochondrial abnormalities. Urinary excretion of lipid peroxidation was increased compared to healthy controls. The porphyrin precursors may increase lipid peroxidation products and damage mitochondria leading to tubulointerstitial nephritis. PMID- 11269684 TI - An incomplete form of the Fanconi syndrome in a patient with osteogenesis imperfecta. PMID- 11269685 TI - Takayasu arteritis and renovascular hypertension. PMID- 11269686 TI - Immunocomplex-mediated crescentic glomerulonephritis associated with anti-DNA antibody: a renal limited lupus? PMID- 11269687 TI - Pseudohyperkalemia, thrombocytosis and renal cancer. PMID- 11269688 TI - Acute fatal colchicine intoxication in a patient on continuous ambulatory peritoneal dialysis (CAPD). Possible role of clarithromycin administration. PMID- 11269689 TI - Oxidative stress and cardiovascular disease in hemodialysis. PMID- 11269690 TI - Measurement of human chylomicron triglyceride clearance with a labeled commercial lipid emulsion. AB - Human chylomicron triglyceride (TG) kinetics has been difficult to determine directly owing to technical limitations. This report describes a new method for studying chylomicron metabolism. Healthy volunteers (n = 10) sipped a drink providing 175 mg fat x kg(-1) h(-1) for 7.5 h to produce a steady-state chylomicronemia. A commercial 10% intravenous lipid emulsion was labeled with [3H]triolein, purified by high-performance liquid chromatography, and sterilized. A trace amount of labeled emulsion was injected intravenously 30 min before (i.e., in the fasting state) and 5, 6, and 7 h after sipping began (i.e., triplicate determinations in the fed state). Chylomicron half-lives were calculated from the monoexponential decay curves, and apparent distribution volumes were estimated by back-extrapolation to time zero. Plasma and estimated chylomicron TG concentrations increased from 89+/-13 and 0.8+/-0.3 to 263+/-43 and 91+/-39 mg/dL (mean +/- SEM), respectively, with feeding. Tracer-determined chylomicron TG half-lives were 5.34+/-0.58 and 6.51+/-0.58 min during the fasting and fed states, respectively (P < 0.01). The apparent distribution volume during the fasting state was 24% greater than plasma volume (4515+/-308 vs. 3630+/-78 mL, P < 0.02), consistent with significant margination of lipid emulsion particles to endothelial binding sites. Margination was reduced during the fed state, suggesting that native chylomicrons competed with lipid emulsion particles for endothelial lipoprotein lipase. The results indicate that a radiolabeled commercial lipid emulsion is metabolized in a fashion similar to native chylomicron TG, and thus can be used to study chylomicron TG kinetics. PMID- 11269692 TI - Low density lipoprotein receptor mRNA in rat liver is affected by resistant starch of beans. AB - The effects of resistant starches of beans on serum cholesterol and hepatic low density lipoprotein (LDL) receptor mRNA in rats were investigated. Rats were fed a cholesterol-free diet with 150 g/kg corn starch (CS), 150 g/kg adzuki (Vigna angularis) starch (AS), 150 g/kg kintoki (Phaseolus vulgaris, variety) starch (KS), or 150 g/kg tebou (P. vulgaris, variety) starch (TS) for 4 wk. There were no significant differences in body weight among groups through the experimental period. The liver weight in the CS group was 1.1-1.2 times higher than that in the AS, KS, and TS groups. The cecum weight in the TS was 1.4 times higher than that in the CS group, and the cecal pH in the CS group was significantly higher than in the other groups. The serum total cholesterol, very low density lipoprotein + intermediate density lipoprotein + LDL-cholesterol and high density lipoprotein (HDL)-cholesterol concentrations in the bean starch groups were significantly lower than those in the CS group through the feeding period. The total cholesterol/HDL-cholesterol ratio in the bean starch groups was also significantly lower than that in the CS group at the end of the 4-wk feeding period. The hepatic cholesterol concentration in the TS group was significantly higher than in the CS group at the end of the 4-wk feeding period. The relative quantity of hepatic apo B mRNA in the AS group was 1.2 times higher than that in the CS group, and the hepatic LDL receptor mRNA levels in the AS and TS groups were 1.8-2.0 times higher than that in the CS group. The results of this study demonstrate that AS, KS, and TS lowered the serum total cholesterol level by enhancing the hepatic LDL receptor mRNA level. PMID- 11269691 TI - Major clofibrate effects on liver and plasma lipids are independent of changes in polyunsaturated fatty acid composition induced by dietary fat. AB - The effects of clofibrate on the content and composition of liver and plasma lipids was studied in mice fed for 4 wk on diets enriched in n-6 or n-3 polyunsaturated fatty acids (PUFA) from sunflower oil (SO) or fish oil (FO), respectively; both oils were fed at 9% of the diet (dry weight basis). Only FO was hypolipidemic. Both oil regimes led to slightly increased concentrations of phospholipids (PL) and triacylglycerols (TG) in liver as compared with a standard chow diet containing 2% fat. Clofibrate promoted hypolipidemia only in animals fed SO. Its main effect was to enlarge the liver, such growth increasing the amounts of major glycerophospholipids while depleting the TG. SO and FO consumption changed the proportion of n-6 or n-3 PUFA in liver and plasma lipids in opposite ways. After clofibrate action, the PUFA of liver PL were preserved better than in the absence of oil supplementation. However, most of the drug induced changes (e.g., increased 18:1n-9 and 20:3n-6, decreased 22:6/20:5 ratios) occurred irrespective of lipids being rich in n-6 or n-3 PUFA. The concentration of sphingomyelin (SM), a minor liver lipid that virtually lacks PUFA, increased with the dietary oils, decreased with clofibrate, and changed its fatty acid composition in both situations. Thus, oil-increased SM had more 22:0 and 24:0 than clofibrate-decreased SM, which was significantly richer in 22:1 and 24:1. PMID- 11269693 TI - Dietary supplementation with conjugated linoleic acid does not alter the resistance of mice to Listeria monocytogenes infection. AB - Conjugated linoleic acid (CLA) has been used experimentally as a dietary supplement to increase lean body weight and to modulate inflammation in a variety of animal species. In addition, human use of dietary CLA as a supplement to regulate body fat has received both scientific and public attention. No reports have been published regarding the effects of dietary CLA on antimicrobial resistance. In this study, we provide evidence that feeding CLA for up to 4 wk does not alter host defense against Listeria monocytogenes in mice. These findings suggest that the anti-inflammatory effects of CLA do not impair cellular immunity to this intracellular pathogen. PMID- 11269694 TI - Effect of conjugated linoleic acid on fungal delta6-desaturase activity in a transformed yeast system. AB - Conjugated linoleic acid (CLA; 18:2), a group of positional and geometric isomers of linoleic acid (LA; 18:2n-6), has been shown to modulate immune function through its effect on eicosanoid synthesis. This effect has been attributed to a reduced production of n-6 polyunsaturated fatty acid (PUFA), the precursor of eicosanoids. Since delta6-desaturase is the rate-limiting enzyme of the n-6 PUFA production, it is our hypothesis that CLA, which has similar chemical structure to LA, interacts directly with delta6-desaturase. A unique and simple model, i.e., baker's yeast (Saccharomyces cerevisiae) transformed with fungal delta6 desaturase gene, previously established, was used to investigate the direct effect of CLA on delta6-desaturase. This model allows LA to be converted to y linolenic acid (GLA; 18:3n-6) but not GLA to its metabolite(s). No metabolites of CLA were found in the lipids of the yeast transformed with delta6-desaturase. The inability to convert CLA to conjugated GLA was not due to the failure of yeast cells to take up the CLA isomers. CLA mixture and individual isomers significantly inhibited the activity of delta6-desaturase of the transformed yeast in vivo. Even though its uptake by the yeast was low, CLA c9,t11 isomer was found to be the most potent inhibitor of the four isomers tested, owing to its high inhibitory effect on delta6-desaturase. Since CLA did not cause significant changes in the level of delta6-desaturase mRNA, the inhibition of GLA production could not be attributed to suppression of delta6-desaturase gene expression at the transcriptional level. PMID- 11269695 TI - Cultured fish cells metabolize octadecapentaenoic acid (all-cis delta3,6,9,12,15 18:5) to octadecatetraenoic acid (all-cis delta6,9,12,15-18:4) via its 2-trans intermediate (trans delta2, all-cis delta6,9,12,15-18:5). AB - Octadecapentaenoic acid (all-cis delta3,6,9,12,15-18:5; 18:5n-3) is an unusual fatty acid found in marine dinophytes, haptophytes, and prasinophytes. It is not present at higher trophic levels in the marine food web, but its metabolism by animals ingesting algae is unknown. Here we studied the metabolism of 18:5n-3 in cell lines derived from turbot (Scophthalmus maximus), gilthead sea bream (Sparus aurata), and Atlantic salmon (Salmo salar). Cells were incubated in the presence of approximately 1 microM [U-14C]18:5n-3 methyl ester or [U-14C]18:4n-3 (octadecatetraenoic acid; all-cis delta6,9,12,15-18:4) methyl ester, both derived from the alga Isochrysis galbana grown in H14CO3-, and also with 25 microM unlabeled 18:5n-3 or 18:4n-3. Cells were also incubated with 25 microM trans delta2, all-cis delta6,9,12,15-18:5 (2-trans 18:5n-3) produced by alkaline isomerization of 18:5n-3 chemically synthesized from docosahexaenoic acid (all cis delta4,7,10,13,16,19-22:6). Radioisotope and mass analyses of total fatty acids extracted from cells incubated with 18:5n-3 were consistent with this fatty acid being rapidly metabolized to 18:4n-3 which was then elongated and further desaturated to eicosatetraenoic acid (all-cis delta8,11,14,17,19-20:4) and eicosapentaenoic acid (all-cis delta5,8,11,14,17-20:5). Similar mass increases of 18:4n-3 and its elongation and further desaturation products occurred in cells incubated with 18:5n-3 or 2-trans 18:5n-3. We conclude that 18:5n-3 is readily converted biochemically to 18:4n-3 via a 2-trans 18:5n-3 intermediate generated by a delta3,delta2-enoyl-CoA-isomerase acting on 18:5n-3. Thus, 2-trans 18:5n-3 is implicated as a common intermediate in the beta-oxidation of both 18:5n-3 and 18:4n-3. PMID- 11269696 TI - Characterization of N-acylphosphatidylethanolamine and acylphosphatidylglycerol in oats. AB - Two polar lipid classes, both with three acyl groups, were isolated from an extract of oats and characterized by nuclear magnetic resonance spectroscopy, electrospray mass spectrometry (MS), and electron ionization MS (EIMS). Distortionless enhancement by polarization transfer (DEPT) and the two dimensional correlation experiments 1H-detected heteronuclear multiple quantum coherence spectroscopy, heteronuclear multiple bond correlation spectroscopy, double quantum filtered correlation spectroscopy, and total correlation spectroscopy provided sufficient information for determination of the structure of the two lipid classes. The polar lipid classes were found to be N acylphosphatidylethanolamine [1,2-diacyl-sn-glycero-3-phospho-(N-acyl)-1' ethanolamine; N-acyl-PE] and acylphosphatidylglycerol [1,2-diacyl-sn-glycero-3 phospho-(3'-acyl)-1'-sn-glycerol]. High-performance liquid chromatography with electrospray ionization MS (HPLC-ESMS) and with electrospray ionization tandem MS (HPLC-MS/MS) were utilized for the separation and subsequent determination of molecular species. With HPLC-ESMS, ions of deprotonated molecules were obtained and with HPLC-MS/MS carboxylate ions (representing acyl groups) were obtained as well as other structurally significant ions. Fifty molecular species of N acylphosphatidylethanolamine and 24 molecular species of acylphosphatidylglycerol were found, with a molecular mass range of 924-1032 Da and 959-1035 Da, respectively. Identification of the fatty acid isomers, as picolinyl ester derivatives, was done with gas chromatography with EIMS. Three isomers of 16:1 fatty acids were found in N-acyl-PE, and their double bond positions were determined to 6, 9, and 11 with a relative abundance of 4:10:1. PMID- 11269697 TI - Fatty acid composition of lipids present in selected lichenized fungi: a chemotyping study. AB - The total-lipid composition of 21 lichens of the ascomycetous genera Cladonia (11) and Cladina (1) of the family Cladoniacea, Cladia (1), Parmotrema (3), Ramalina (2), Leptogium (1), Cetraria (1), and the basidiomycetous genus Dictyonema (1) was determined. Analyses of those of Dictyonema glabratum were carried out with a total extract and those obtained after successive extractions with various solvents. Each extract was partitioned between n-heptane/isopropanol and 1 M sulfuric acid, giving triglycerides (TG) in the upper phase. Extracts were methanolyzed and the resulting methyl esters were analyzed by gas chromatography-mass spectrometry. Methanolyzates of TG unexpectedly contained esters of 9-oxodecanoic, 9-methyl-tetradecanoic, 6-methyl-tetradecanoic, 3 hydroxy-decanoic, nonanedioic, and decanedioic acids, as well as common fatty acids. Fatty acid methyl ester profiles from the lichens were submitted to cluster analysis, and the resulting dendogram showed a cluster consistent with Cladonia spp., suggesting an efficient aid to lichen taxonomy. The total carbohydrate content of each lipid extract was determined by a modified phenol sulfuric acid method, which compensated for the presence of pigments. PMID- 11269698 TI - A new ceramide from the basidiomycete Russula cyanoxantha. AB - A new phytosphingosine-type ceramide (1) was isolated along with nine other compounds-5alpha,8alpha-epidioxy(22E,24R)-ergosta-6,22-dien-3beta-ol, 5alpha,8alpha-epidioxy-(24S)-ergosta-6-en-3beta-ol, (24S)-ergosta-7-ene 3beta,5alpha,6beta-triol, (22E,24R)-ergosta-7,22-dien-3beta,5alpha,6beta-triol, inosine, adenine, L-pyroglutamic acid, fumaric acid, and D-allitol from the ethanol and chloroform/methanol extract of the fruiting bodies of the basidiomycete Russula cyanoxanotha (Schaeff.) Fr. The structure of (1) was established as (2S,3S,4R,2'R)-2-(2'-hydroxytetracosanoylamino) octadecane-1,3,4 triol by means of spectroscopic and chemical methods. PMID- 11269699 TI - Ab initio and density functional theory studies for the explanation of the antioxidant activity of certain phenolic acids. AB - Ab initio and density functional theory molecular orbital calculations were carried out at both the HF/6-31 +G(d) and B3LYP/6-31+G(d) levels for the four antioxidants, p-hydroxycinnamic acid derivatives, namely, the p-coumaric, caffeic, ferulic, and sinapinic acid and the corresponding radicals, in an attempt to explain the structural dependency of the antioxidant activity of these compounds. Optimized resulting geometries, vibrational frequencies, absolute infrared intensities, and electron-donating ability are discussed. Both the high degree of conjugation and the extended spin delocalization in the phenoxyl radicals offer explanation for the scavenging activity of the four acids. In structurally related compounds, the calculated heat of formation value in radical formation appears as a meaningful molecular descriptor of antioxidant activity in accordance with experimental data. This becomes more clear at the B3LYP level. PMID- 11269700 TI - Formation of cleavage products by autoxidation of lycopene. AB - The cleavage products formed by autoxidation of lycopene were evaluated in order to elucidate possible oxidation products of lycopene in biological tissues. Lycopene solubilized at 50 microM in toluene, aqueous Tween 40, or liposomal suspension was oxidized by incubating at 37 degrees C for 72 h. Among a number of oxidation products formed, eight products in the carbonyl compound fraction were identified as 3,7,11 -trimethyl-2,4,6,10-dodecatetraen-1-al, 6,10,14-trimethyl 3,5,7,9,13-pentadecapentaen-2-one, acycloretinal, apo-14'-lycopenal, apo-12' lycopenal, apo-10'-lycopenal, apo-8'-lycopenal, and apo-6'-lycopenal. These correspond to a series of products formed by cleavage in the respective 11 conjugated double bonds of lycopene. The maximal formation of acycloretinal was 135 nM in toluene, 49 nM in aqueous Tween 40, and 64 nM in liposomal suspension. Acycloretinoic acid was also formed by autoxidation of lycopene, although its formation was lower in the aqueous media than in toluene. The pig liver homogenate had the ability to convert acycloretinal to acycloretinoic acid, comparable to the conversion of all-transretinal to all-trans-retinoic acid. These results suggest that lycopene might be cleaved to a series of apolycopenals and short-chain carbonyl compounds under the oxidative conditions in biological tissues and that acycloretinal is further enzymatically converted to acycloretinoic acid. PMID- 11269701 TI - Cyclodehydration reactions of methyl 9,10-; 10,12-; and 9,1 2-dioxostearates with 1,2-diaminoethane under ultrasonic irradiation. AB - Reaction of methyl 9,10-dioxostearate (1) and 9,12-dioxostearate (2) with 1,2 diaminoethane under concomitant ultrasonic irradiation (10-15 min, 60 degrees C) in water furnished the corresponding 2,3-dihydropyrazine (4, 79%) and 1,2,3,4 tetrahydro-1,4-diazocine (5, 70%) derivatives, respectively. Reaction of methyl 10,12-dioxostearate (3) with 1,2-diaminoethane was successful only when glacial acetic acid was used instead of water under ultrasonic irradiation (4 x 10 min, 70 degrees C) to give a 2,3-dihydro-1H-1,4-diazepine (6, 95%) derivative. The structures of these novel six-membered (4), seven-membered (6), and eight membered (5) N-heterocyclic fatty ester derivatives were confirmed by a combination of infrared, nuclear magnetic resonance spectroscopic and mass spectral analyses. PMID- 11269702 TI - Dialysis and gel filtration of isolated low density lipoproteins do not cause a significant loss of low density lipoprotein tocopherol and carotenoid concentration. AB - The resistance of isolated low density lipoprotein (LDL) to copper-initiated oxidation is often used as a measure of effectiveness of an antioxidant intervention. Prior to oxidation, excess salt and EDTA are removed via dialysis or gel filtration of the LDL sample. However, there is concern over whether the antioxidant content of dialyzed or gel-filtered LDL is truly representative of native LDL extracted from a blood sample. Previously, the experiments done after the storage of native and dialyzed LDL at -80 degrees C showed that the dialysis step can cause a loss of up to 60% in the tocopherol and carotenoid content of LDL. In the present study, a comparison of the micronutrient concentration in freshly prepared dialyzed and native LDL from 35 subjects showed that after the correction for cholesterol, only lycopene (13%, P < 0.001) and to a lesser extent alpha-carotene (8%, P < 0.02) were significantly decreased, and the absolute fall in concentration was far smaller than previously reported. Other experiments done with smaller numbers of samples suggested that there were minimal micronutrient losses following gel filtration and that it was important to include 10 micromol/L EDTA in the dialysis and elution buffer; otherwise micronutrient losses did occur. In summary, immediate dialysis of freshly isolated LDL in the presence of 10 micromol/L EDTA does not cause any major loss in the concentration of tocopherol and most carotenoids. PMID- 11269703 TI - Alpha-linolenic acid and its delta5-desaturation product, coniferonic acid, in the seed lipids of Tsuga and Hesperopeuce as a taxonomic means to differentiate the two genera. PMID- 11269704 TI - Overestimates of oleic and linoleic acid contents in materials containing trans fatty acids and analyzed with short packed gas chromatographic columns. PMID- 11269705 TI - Production, isolation and structure determination of novel fluoroindolocarbazoles from Saccharothrix aerocolonigenes ATCC 39243. AB - Saccharothrix aerocolonigenes ATCC 39243 produces an indolocarbazole antitumor agent rebeccamycin under submerged fermentation conditions. Adding DL-6 fluorotryptophan to culture of S. aerocolonigenes ATCC 39243 induces the formation of two novel indolocarbazoles, fluoroindolocarbazoles A and B. Feeding DL-5-fluorotryptophan to culture of S. aerocolonigenes ATCC 39243 induces the production of a novel indolocarbazole, fluoroindolocarbazole C. These fluoroindolocarbazoles have been isolated from culture broth and purified to homogeneity by vacuum liquid chromatography and column chromatography. All three fluoroindolocarbazoles are more potent than rebeccamycin against P388 leukemia by ip route in murine model. PMID- 11269708 TI - Short and convergent synthesis of asterriquinone B1 and demethylasterriquinone B1. PMID- 11269706 TI - Isolation, and biological properties of a new cell cycle inhibitor, curvularol, isolated from Curvularia sp. RK97-F166. AB - A new cell growth inhibitor, curvularol, was isolated from the fermentation broth of Curvularia sp. RK97-F166. Curvularol showed no antibacterial activity, and very weak antifungal activity. However, curvularol inhibited the cell cycle progression of normal rat kidney (NRK) cells in G1 phase at 150 ng/ml. Curvularol induced the morphological reversion of srcts-transformed NRK cells at 100 ng/ml, and inhibited protein synthesis same as cycloheximide. PMID- 11269707 TI - AJI9561, a new cytotoxic benzoxazole derivative produced by Streptomyces sp. PMID- 11269709 TI - Biomimetric total synthesis of quinolactacin B, TNF production inhibitor, and its analogs. PMID- 11269710 TI - New cyclic depsipeptide antibiotics, clavariopsins A and B, produced by an aquatic hyphomycetes, Clavariopsis aquatica. 1. Taxonomy, fermentation, isolation, and biological properties. AB - Clavariopsins were isolated from the fermentation broth of Clavariopsis aquatica AJ 117363. Clavariopsins are cyclic depsipeptide antibiotics with the molecular weight of 1,153 and 1,139. Clavariopsins showed in vitro antifungal activity against not only Aspergillus fumigatus but also, although to a lesser extent, A. niger and Candida albicans. PMID- 11269711 TI - New cyclic depsipeptide antibiotics, clavariopsins A and B, produced by an aquatic hyphomycetes, Clavariopsis aquatica. 2. Structure analysis. AB - The structures of new cyclic decadepsipeptides, clavariopsins A and B, were determined to be cyclo[-(R)-2-hydroxyisovaleryl-L-pipecoyl-L-MeVal-L-Val-L-MeAsp L-MeIle-L-MeIle-Gly L-MeVal-L-Tyr(OMe)-] and cyclo[-(R)-2-hydroxyisovaleryl-L pipecolyl-L-Val-L-Val-L-MeAsp-L-Melle-L-MeIle-Gly-L-MeVal-L-Tyr(OMe)-], respectively, by spectroscopic analyses, especially using 2D NMR techniques. The absolute stereochemistry was elucidated by the advanced Marfey's method and chiral HPLC analysis. PMID- 11269712 TI - Bioxanthracenes from the insect pathogenic fungus. Cordyceps pseudomilitaris BCC 1620. I. Taxonomy, fermentation, isolation and antimalarial activity. AB - Eleven bioxanthracenes and two monomers, six novel in nature, were isolated from the insect pathogenic fungus Cordyceps pseudomilitaris BCC 1620. Growth optimization of the strain led to the improvement of bioxanthracenes production. The bioxanthracenes were evaluated for their antimalarial activity and cytotoxicity. PMID- 11269713 TI - Bioxanthracenes from the insect pathogenic fungus Cordyceps pseudomilitaris BCC 1620. II. Structure elucidation. AB - Structures of eleven bioxanthracenes (1 approximately 11) and two monomers (12 and 13), isolated from the insect pathogenic fungus Cordyceps pseudomilitaris BCC 1620, were elucidated. The structure, including the axial stereochemistry, of one of the major symmetrical dimers (1) was determined by X-ray crystallographic analysis, while the stereochemistries of the other isomers were deduced by chemical conversions and spectroscopic means. PMID- 11269714 TI - Design of a novel mammalian screening system for the detection of bioavailable, non-cytotoxic streptogramin antibiotics. AB - Screening and development of new antibiotic activities to counteract the increasing prevalence of multidrug-resistant (MDR) human pathogenic bacteria has once again become a priority in human chemotherapy. Here we describe a novel mammalian cell culture-based screening platform for the detection of streptogramin antibiotics. Quinupristin-dalfopristin (Synercid), a synthetically modified streptogramin, is presently the sole effective agent in the treatment of some MDR nosocomial infections. A Streptomyces coelicolor transcriptional regulator (Pip) has been adapted to modulate reporter gene expression (SEAP, secreted alkaline phosphatase) in Chinese hamster ovary cells (CHO) in response to streptogramin antibiotics. This CHO cell-based technology was more sensitive in detecting the production of the model streptogramin pristinamycin, from Streptomyces pristinaespiralis, than antibiogram tests using a variety of human pathogenic bacteria as indicator strains. The reporter system was able to detect pristinamycin compound produced by a single S. pristinaespiralis colony. The assay was rapid (17 hours) and could be carried out in a high-throughput 96-well plate assay format or a 24-well transwell set-up. This novel mammalian cell-based antibiotic screening concept enables detection of bioavailable and non-cytotoxic representatives of a particular class of antibiotics in a single assay and represents a promising alternative to traditional antibiogram-based screening programs. PMID- 11269715 TI - Extremophilic orgainisms as and unexplored source fo antifungal compounds. AB - Extracts of the biomasses and fermentation broths of 217 extremophilic microorganisms isolated from a number of locales were screened for antifungal activity using whole-cell and mechanism-based in vitro assays. Importantly, eleven broth extracts had activity against several Candida species and Aspergillus fumigatus in whole-cell in vitro assays. One broth specifically inhibited (1,3)beta-glucan synthase activity and four specifically inhibited ketol-isomerase activity, suggesting a mode of action of the antifungal compound(s) present in these extracts. The extract from one thermophile, a novel species of Pseudomonas, was fractionated, an active compound purified and its structure determined. The compound was identified as pyochelin, a previously identified iron-binding compound with heretofore undescribed antifungal activity. To our knowledge, this is the first report demonstrating that extremophiles synthesize compounds that have antifungal activity. PMID- 11269716 TI - Construction and physiological studies on a stable bioengineered strain of shengjimycin. AB - Shengjimycin is a group of 4"-acylated spiramycins with 4"-isovalerylspiramycin as the major component, produced by recombinant S. spiramyceticus F21 harboring a 4"-O-acyltransferase gene from S. mycarofaciens 1748. A stable bioengineered strain of Streptomyces spiramyceticus WSJ-1 was constructed by integrating the 4" O-acyltransferase gene (ist) by homologous recombination into the chromosome of the spiramycin-producing strain S. spiramyceticus F21. In this construction, a Streptomyces/E. coli shuttle plasmid pKC1139 (AmR) was used as the vector with the tsr gene used as selection marker for homologous recombination. The constructed strain, S. spiramyceticus WSJ-1,was genetically stable in production titer and proportion of components of shengjimycin as well as in maintaining the tsr selective marker when grown without selection. Southern hybridization confirmed the integrated status of the ist gene in the host genome. The production and the proportion of major component of 4"-isovalerylspiramycin of S. spiramyceticus WSJ-1 was also improved comparing with the strain harboring an autonomous plasmid -S. spiramyceticus F21/pIJ680(311) as shown by HPLC analysis. Physiological studies indicated that increase of the VDH ( valine dehydrogenase ) and LDH ( leucine dehydrogenase ) activities of WSJ-1 may be involved in this improvement. PMID- 11269718 TI - Borrelidin inhibits a cyclin-dependent kinase (CDK), Cdc28/Cln2, of Saccharomyces cerevisiae. AB - We identified borrrelidin, a member of macrolide antibiotic, as an inhibitor of a cyclin-dependent kinase of the budding yeast, Cdc28/Cln2. A 50% inhibition concentration (IC50) of borrelidin for Cdc28/Cln2 was 24 microM. In addition, borrelidin arrests both haploid and diploid cells in G1 phase at the point indistinguishable from that of alpha-mating pheromone, at concentrations not affecting the gross protein synthesis. Although the inhibition of CDK activity may not be a solo cause of the G1 arrest, our results indicate that borrelidin is a potential lead compound for developing novel CDK inhibitors of higher eukaryotes. PMID- 11269717 TI - Novel fungal metabolites as cell wall active antifungals: fermentation, isolation, physico-chemical properties, structure and biological activity. AB - Two novel antifungal compounds, 1 (SCH 466457), and 2 (SCH 466456), active in a "cell wall" assay, were isolated from the fermentation broth of an unidentified fungus. The active compounds were separated from the broth filtrate by adsorption on a macroreticular resin and were purified on reverse phase HPLC. Detailed mass spectrometric and NMR experiments and degradative studies helped in elucidating the structures of these compounds. The compounds were identified to be peptides containing amino acids such as alanine, aminoisobutyric acid, proline, leucine, valine, glycine and a previously identified beta-keto acid, 2-methyl 3 oxotetradecanoic acid. (5) Both compounds were active against Candida, dermatophytes and Aspergillus (Geometric Mean MIC's, 8.9, 20 and 16 microg/ml, and 64, 128 and 23 microg/ml, respectively for 1 and 2). PMID- 11269719 TI - Identification of four genes from the granaticin biosynthetic gene cluster of Streptomyces violaceoruber Tu22 involved in the biosynthesis of L-rhodinose. AB - Four genes, ORF 22 approximately 25, from the granaticin biosynthetic gene cluster of Streptomyces violaceoruber Tu22 were analyzed for their involvement in the biosynthesis of the two deoxysugar moieties of the granaticins. Each gene was individually inactivated on a cosmid carrying the entire gra gene cluster and the mutant cosmids were transformed into S. coelicolor CH999. Analysis of the pattern of pigment production by the transformants revealed that each of the four ORFs is required for the formation/attachment of the L-rhodinose moiety of granaticin B, but not that of the D-olivose moiety of granaticin. Based on these results and sequence homologies a pathway of dTDP-L-rhodinose formation is proposed which implicates ORF23, and possibly also ORF 24, in the 3-deoxygenation reaction, ORF 25 in the epimerization and ORF 22 in the final 4-ketoreduction. PMID- 11269720 TI - From histoalchemy to molecular marvels: a sojourn through periodontal connective tissue research. PMID- 11269721 TI - Analysis of ordinal dental data: evaluation of conflicting recommendations. AB - Although recommendations for the appropriate analysis of non-normal and ordinal scaled data have appeared in the dental research literature for many years, there is no consensus. When one is conducting statistical tests for differences between groups, the central concern is whether it is safe to use parametric tests (e.g., analysis of variance), or if only non-parametric ranking tests should be considered. Relevant statistical and scientific issues associated with non normality and measurement scale are reviewed, and three conclusions are reached regarding the analysis of dental data: (1) Parametric tests are sufficiently robust relative to typical violations of normality; (2) presumed statistical prohibitions against the application of parametric methods to ordinal data do not actually exist; and (3) 'ordinal' dental indices have sufficient quantitative meaning to be considered quasi-interval. For these reasons, parametric tests should not be avoided; they will be valid and usually more powerful and more easily applied to complex designs than non-parametric alternatives. PMID- 11269723 TI - Biological organization of hydroxyapatite crystallites into a fibrous continuum toughens and controls anisotropy in human enamel. AB - Enamel forms the outer surface of teeth, which are of complex shape and are loaded in a multitude of ways during function. Enamel has previously been assumed to be formed from discrete rods and to be markedly aniostropic, but marked anisotropy might be expected to lead to frequent fracture. Since frequent fracture is not observed, we measured enamel organization using histology, imaging, and fracture mechanics modalities, and compared enamel with crystalline hydroxyapatite (Hap), its major component. Enamel was approximately three times tougher than geologic Hap, demonstrating the critical importance of biological manufacturing. Only modest levels of enamel anisotropy were discerned; rather, our measurements suggest that enamel is a composite ceramic with the crystallites oriented in a complex three-dimensional continuum. Geologic apatite crystals are much harder than enamel, suggesting that inclusion of biological contaminants, such as protein, influences the properties of enamel. Based on our findings, we propose a new structural model. PMID- 11269722 TI - Incipient analysis of mesenchymal stem-cell-derived osteogenesis. AB - Tissue regeneration strategies invoke cell-based therapies for effective tissue formation. Current assessment of mesenchymal stem cell (MSC) directed bone regeneration during in vivo assays is dependent on histologic determination of bone formation. It was the aim of this study to determine the relationship between bone sialoprotein (BSP) expression and osteocalcin expression with subsequent osteogenesis occurring in MSC-based implants. RT-PCR assessment of human actin, collagen type I, BSP, and osteocalcin indicated that undifferentiated cells did not express BSP or osteocalcin. Three weeks following implantation, human BSP could be identified in RNAs isolated from the retrieved implants. For every implant from which human BSP cDNA was amplified, parallel implants harvested at 6 weeks demonstrated bone formation at the histologic level. This study confirms that, in the context of the severe combined immunodeficiency disease (SCID) mouse model, culture-expanded, cryopreserved human MSCs have osteogenic potential and demonstrates that implanted cell gene expression can reveal the early onset of bone formation. PMID- 11269724 TI - Mass properties of the pig mandible. AB - Specification of mass properties is an essential step in the modeling of jaw dynamics, but obtaining them can be difficult. Here, we used three-dimensional computed tomography (CT) to estimate jaw mass, mean bone density, anatomical locations of the mass and geometric centers, and moments of inertia in the pig jaw. High-resolution CT scans were performed at one-mm slice intervals on specimens submerged in water. The mean estimated jaw mass was 12% greater than the mean wet weight, and 33% more than the mean dry weight. Putative bone marrow accounted for an extra 13% of mass. There was a positive correlation between estimated mean bone density and age. The mass center was consistently in the midline, near the last molar. The mean distance between the mass center and geometric center was small, especially when bone marrow was taken into account (0.58 +/- 0.21 mm), suggesting that mass distribution in the pig jaw is almost symmetrical with respect to its geometric center. The largest moment of inertia occurred around each mandible's supero-inferior axis, and the smallest around its antero-posterior axis. Bone marrow contributed an extra 9% to the moments of inertia in all three axes. Linear relationships were found between the actual mass and a mass descriptor (product of the bounding volume and mean bone density), and between the moments of inertia and moments of inertia descriptors (products of the mass descriptor and two orthogonal dimensions forming the bounding box). The study suggests that imaging modalities revealing three dimensional jaw shape may be adequate for estimating the bone mass properties in pigs. PMID- 11269725 TI - Improvement of strength parameters of a leucite-reinforced glass ceramic by dual ion exchange. AB - An innovative dual ion-exchange process can improve the limited strength and the scatter-in-strength of technical glasses. The objective of this study was to prove whether such a two-stage ion-exchange process can also improve the limited strength and the problematic scatter-in-strength of dental ceramic materials. The first exchange was done in KNO3 on a leucite-reinforced glass-ceramic material, the second exchange in 70 mol% KNO3, 30 mol% Na NO3 at different treatment times and temperatures. The dual-exchange process approximately doubled the Weibull strength. Moreover, the Weibull modulus showed a four-fold increase, i.e., the coefficients of variation were reduced from 18.3 to 4.7%. We conclude that the dual-exchange process may help significantly to increase the clinical reliability of glass-ceramic dental restorations, because the strength and the scatter-in strength will be substantially improved by this treatment. PMID- 11269726 TI - Influence of dentition status on physical disability, mental impairment, and mortality in institutionalized elderly people. AB - The loss of teeth is known to influence the mastication of foods and nutritional status. Therefore, we hypothesize that poor dentition status can impair the systemic health of the aged. To clarify the influence of dentition status on deterioration in physical ability, mental impairment, and mortality, we conducted a six-year prospective cohort study of the institutionalized elderly living in 29 of the 30 institutions for the elderly in Kitakyushu, Japan. Bivariate analysis revealed that worse dentition status at baseline led to significantly worse physical and mental impairment, and higher mortality. In a multiple logistic regression analysis, the physical ability of edentulous subjects without dentures significantly deteriorated compared with that of dentate subjects with 20 or more teeth. The six-year mortality rate of the edentulous subjects without dentures was significantly higher than that of the subjects with 20 or more teeth. Poorer dentition status, especially edentulousness without dentures, may therefore be related to deterioration in the systemic health of the aged. PMID- 11269727 TI - Immunodominant region of Actinobacillus actinomycetemcomitans 40-kilodalton heat shock protein in patients with rheumatoid arthritis. AB - Bacterial heat shock proteins have been implicated in the pathogenesis of several diseases, and the immunological relationship between rheumatoid arthritis (RA) and Escherichia coli DnaJ has been reported. Since there are similarities in the tissue destruction process of RA and periodontitis, we examined the reactivities of antibodies in sera from RA patients to the DnaJ protein from Actinobacillus actinomycetemcomitans. An enzyme-linked immunosorbent assay showed that IgG titers to the N-terminal conservative region of the DnaJ are significantly higher in RA patients compared with the healthy controls (p < 0.05). Furthermore, we examined IgG titers of disease controls to determine the specificity of the immune responses to this region in RA patients. The difference between RA and infectious disease patients was also significant (p < 0.05). These results suggest that the N-terminal region of DnaJ from A. actinomycetemcomitans may contribute to the etiologic analysis of RA. PMID- 11269728 TI - Interferon-gamma down-regulates gene expression of cathepsin K in osteoclasts and inhibits osteoclast formation. AB - The cytokine, IFN-gamma, has been shown in vitro to inhibit bone resorption, but the mechanisms responsible for this inhibition have not been clearly defined. Cathepsin K is a major protease responsible for bone resorption. IFN-gamma may inhibit bone resorption through down-regulation of osteoclast genes, including cathepsin K. To test the hypothesis, we investigated the effect of IFN-gamma on cathepsin K expression in the MOCP-5 and wild-type mouse bone marrow co-culture systems by Northern blot as well as osteoclast formation at different stages of differentiation. The results show that IFN-gamma down-regulates mRNA levels of cathepsin K in a time- and dose-dependent manner. Consequently, cathepsin K protein production is also reduced by IFN-gamma. Moreover, our results indicate that IFN-gamma inhibits osteoclast formation only early in osteoclast differentiation. IL-6 and TNFalpha did not significantly affect cathepsin K gene expression in osteoclasts. However, IL-1alpha stimulated gene expression. In conclusion, our data suggest that the actions of IFN-gamma on osteoclastic bone resorption may be mediated by its effects on both osteoclast formation at an early stage and osteoclast gene expression in mature osteoclasts. PMID- 11269729 TI - Large-scale early in vitro response to actinobacillus actinomycetemcomitans suggests superantigenic activation of T-cells. AB - The mode of T-cell response to Actinobacillus actinomycetemcomitans is largely unknown. The present study sought to investigate the hypothesis that A. actinomycetemcomitans expresses superantigens, capable of antigen-non-specific T cell activation. To that end, peripheral blood mononuclear cells were stimulated with A. actinomycetemcomitans, and T-cell expression of the early activation marker, CD69, was determined by flow cytometry. Results showed that A. actinomycetemcomitans activated a large number of T-cells with magnitude similar to that of staphylococcal enterotoxin superantigens. A. actinomycetemcomitans sonicate preferentially activated T-cells expressing Vbeta5.1 and Vbeta8, while the extracellular preparation activated Vbeta5.1+, Vbeta8+, and Vbeta12+ T-cells. T-cell response to A. actinomycetemcomitans was observed in the presence of autologous, as well as heterologous, antigen-presenting cells, suggesting a MHC non-restricted response. Thus, the in vitro response to A. actinomycetemcomitans is characterized by large-scale T-cell activation in a Vbeta-specific and MHC-non restricted manner, consistent with the involvement of superantigens. PMID- 11269730 TI - Validation of an in vitro biofilm model of supragingival plaque. AB - The study of biofilm structure and function mandates the use of model systems for which a host of environmental variables can be rigorously controlled. We describe a model of supragingival plaque containing Actinomyces naeslundii, Veillonella dispar, Fusobacterium nucleatum, Streptococcus sobrinus, and Streptococcus oralis wherein cells are cultivated anaerobically in a saliva-based medium on hydroxyapatite discs coated with a salivary pellicle, with material and pieces of apparatus common to all microbiology laboratories. After 0.5 hr, 16.5 hrs, 40.5 hrs, and 64.5 hrs, the composition of adherent biofilms was analyzed by culture techniques, live/dead fluorescence staining, and confocal laser scanning microscopy. Repeated independent trials demonstrated the repeatability of biofilm formation after 40.5 hrs and 64.5 hrs. Brief exposures of biofilms to chlorhexidine or Triclosan produced losses in viability similar to those observed in vivo. This biofilm model should prove very useful for pre-clinical testing of prospective anti-plaque agents at clinically relevant concentrations. PMID- 11269731 TI - pH-regulated secretion of a glyceraldehyde-3-phosphate dehydrogenase from Streptococcus gordonii FSS2: purification, characterization, and cloning of the gene encoding this enzyme. AB - Streptococcus gordonii and other viridans streptococci (VS) are primary etiologic agents of infective endocarditis, despite being part of the normal oral microflora. Recently, a surface-bound glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has been found on the cells of all tested streptococcal species, where it has been implicated as a virulence factor. In contrast, we observed that a soluble extracellular GAPDH was the major secreted protein from S. gordonii FSS2, an endocarditis strain. The biochemical properties and gene sequence of S. gordonii GAPDH are almost identical to those of other streptococcal GAPDHs. Growth at defined pHs showed that secretion of GAPDH is regulated by environmental pH. GAPDH was primarily surface-associated at growth pH 6.5 and shifted to > 90% secreted at growth pH 7.5. Others have identified S. gordonii promoters that are up-regulated by a pH shift similar to that experienced by organisms entering the blood stream (neutral) from the oral cavity (slightly acid). Analysis of our results suggests that secretion of GAPDH may be a similar adaptation by S. gordonii. PMID- 11269732 TI - Assessment of acid production by various human oral micro-organisms when palatinose or leucrose is utilized. AB - One promising way of reducing caries is by using sucrose substitutes in food, e.g., palatinose or leucrose. Previous experiments addressing cariogenic potential of sucrose substitutes have focused mainly on Streptococcus mutans. However, given the many other micro-organisms in the oral cavity, this study compared the acid production of 100 bacterial strains representing 44 different species, by batch fermentation in a test tube containing, as a sole carbohydrate source, glucose, sucrose, palatinose, or leucrose. Selected strains were further analyzed in a fermenter. Additionally, 30 yeast strains were tested by an auxanographic sugar assimilation test. Only Lactobacillus spp., Stomatococcus mucilaginosus, Leuconostoc mesenteroides, and Weissella paramesenteroides, and some of the yeasts studied-i.e., Candida albicans, C. tropicalis, C. parapsilosis, and Saccharomyces cerevisiae-utilized leucrose and/or palatinose well. Strikingly, Stomatococcus mucilaginosus produced water-insoluble polysaccharides by fermentation of leucrose and palatinose. In the fermenter, the respective sucrose substitutes were not only cleaved but also utilized. Thus, extracellular cleavage by autochthonous micro-organisms may produce cariogenic cleavage products (glucose, fructose) that can be used by other well characterized cariogenic bacteria found in the oral flora. Therefore, the anticariogenic potential of sucrose substitutes in food might be limited. PMID- 11269734 TI - Innervation of human tooth pulp in relation to caries and dentition type. AB - The neural status of carious teeth, particularly those associated with a painful pulpitis, is largely unknown. This study sought to determine differences in the innervation density of human primary and permanent teeth and whether caries or painful pulpitis was associated with anatomical changes in pulpal innervation. Coronal pulps were removed from 120 primary and permanent molars with a known pain history. Teeth were categorized as intact, moderately carious, or grossly carious. Using indirect immunofluorescence, we labeled sections for the general neuronal marker, protein gene product 9.5. Using image analysis, we found permanent teeth to be significantly more densely innervated than primary teeth. While there was no significant correlation with reported pain experience, neural density in both dentitions increased significantly with caries. Analysis of these data suggests that caries-induced changes in neural density may be functionally more important in the regulation of pulpal inflammation and healing than in the processing and perception of dental pain. PMID- 11269733 TI - New in vitro model for the acquired enamel pellicle: pellicles formed from whole saliva show inter-subject consistency in protein composition and proteolytic fragmentation patterns. AB - The present investigation was undertaken to investigate the variability of proteins in whole saliva which adsorb to hydroxyapatite and are thus likely to be precursors of the acquired enamel pellicle. Whole-saliva proteins from 18 subjects were absorbed to hydroxyapatite, and the gel filtration patterns of released proteins revealed four major peaks and three minor peaks eluting between the major peaks. Amino acid analysis indicated that minor peaks contained fragments of proteins in major peaks, and this was confirmed by sequence analysis. Major peaks comprised 95% and minor peaks comprised 5% of protein absorbed to hydroxyapatite, suggesting a limited proteolytic capacity of whole saliva. HPLC elution patterns of components in minor peaks suggested that proteolysis is not totally random but is an orderly and consistent process. These studies suggest that whole saliva may be suitable as a model system for the investigation of post-secretory modifications of salivary proteins important for the formation of the acquired enamel pellicle. PMID- 11269735 TI - Pharmacokinetics, distribution, metabolism and excretion of. AB - PURPOSE: To evaluate the metabolic fate of UCN-01, a signal transduction inhibitor, blood and plasma concentrations, distribution, metabolism and excretion were investigated in rats and dogs after intravenous administration of [3H]UCN-01. METHODS: The radioactivity in plasma, blood and tissues was measured after intravenous administration of UCN-01. In addition, the radioactivity excreted in bile, urine and feces was also determined. RESULTS: The radioactivity in rat and dog plasma disappeared triphasically with terminal half-lives of 21.3 and 27.2 h, respectively. The ratios of the blood-to-plasma concentrations ranged from 0.82 to 1.13 in rats and 0.81 to 1.73 in dogs. From 0.5 to 4 h after giving [3H]UCN-01 to rats, the radioactivity in all tissues except the brain and testes was higher than in plasma. The highest concentration was observed in the lungs followed by the liver and kidneys. The radioactivity was mainly excreted in feces, reaching 96.0% of the radioactivity dose in rats and 78.4% in dogs up to 168 h after injection. Since the biliary excreted radioactivity was 67.2% over 48 h in bile duct-cannulated rats, most of the radioactivity excreted in feces was from biliary radioactivity. There were several metabolites in bile samples, but little UCN-01. CONCLUSIONS: UCN-01 is mainly eliminated by the liver, and there are high concentrations of radioactivity derived from [3H]UCN-01 in all tissues except the brain and testes. PMID- 11269736 TI - Phase I study of mitoxantrone, raltitrexed, levofolinic acid and 5-fluorouracil in advanced solid tumours. AB - PURPOSE: We have recently evaluated the combination of raltitrexed, levofolinic acid (LFA) and 5-fluorouracil (5-FU) in advanced head and neck and colorectal cancer, and we have shown that this combination is well tolerated and has clinical activity. Clinical combination studies have shown that raltitrexed and anthracyclines can be combined at full doses without unexpected toxicities. Based on these observations, we started a phase I study of mitoxantrone plus raltitrexed administered on day 1, followed by LFA and 5-FU on day 2 in patients with advanced solid tumors. PATIENTS AND METHODS: Mitoxantrone was given at a starting dose of 6 mg/m2, raltitrexed at a fixed dose of 3 mg/m2, LFA at a fixed dose of 250 mg/m2, and 5-FU at a starting dose of 750 mg/m2. Mitoxantrone and 5 FU doses were subsequently escalated alternately up to dose-limiting toxicity. Treatment was repeated every 14 days. RESULTS: Four dose levels were tested in 18 patients. All three patients treated at the fourth dose level had grade 4 neutropenia after the first cycle. Therefore, this level was defined as the maximum tolerated dose and the dose level immediately below (mitoxantrone 7 mg/m2 and 5-FU 900 mg/m2) was selected for further evaluation. Neutropenia was the main toxic effect. Nonhaematologic side effects were mild. One complete response and five partial responses (all but one in patients with head and neck cancer) were observed, for an overall response rate of 33% (95% confidence interval, 13% to 59%). CONCLUSIONS: Mitoxantrone, raltitrexed and 5-FU can be combined at doses which are close to those used in monotherapy. The observed activity is encouraging, especially in the subset of patients with head and neck cancer. PMID- 11269737 TI - Pharmacokinetics and pharmacodynamic effects of 5-fluorouracil given as a one hour intravenous infusion. AB - PURPOSE: Clinical toxicity associated with 5-fluorouracil (5-FU) is related to the area under the plasma concentration-time curve (AUC). Recently, short-term infusions of 5-FU given over 30 or 60 min have been substituted for conventional "bolus" 5-FU given over 3-5 min in randomized clinical trials, but there are only limited pharmacokinetic data for these altered infusion durations. We therefore wished to determine the pharmacokinetics and toxicity associated with 5-FU given as a 1-h intravenous (i.v.) infusion. METHODS: A group of 22 adults with advanced gastrointestinal tract cancers and no prior systemic chemotherapy for advanced disease received interferon alpha-2a (5 MU/m2 s.c., days 1-7), leucovorin (500 mg/m2 i.v. over 30 min, days 2-6) and 5-FU (370 mg/m2 i.v. over 1 h, days 2-6). The doses of 5-FU and interferon-alpha were adjusted according to individual tolerance. The pharmacokinetics and clinical toxicity were retrospectively compared with patients receiving the same regimen under the same treatment guidelines except that 5-FU was given over 5 min. RESULTS: The regimen was well tolerated, and 41% of the patients tolerated 5-FU dose escalations to 425-560 mg/m2 per day. Grade 3 or worse diarrhea and fatigue ultimately occurred in 14% of the patients each. Granulocytopenia, mucositis, and diarrhea appeared to be appreciably milder in the present trial compared with our prior phase II experience in colorectal cancer. The peak 5-FU plasma levels and AUC with 370 mg/m2 5-FU given over 1 h were 7.3-fold and 2.4-fold lower than previously measured in 31 patients who received 5-FU over 5 min. CONCLUSION: Increasing the length of 5-FU infusion to 1 h seemed to substantially reduce the clinical toxicity with this modulated 5-FU regimen, likely due to markedly lower peak 5-FU plasma levels and AUC. Changes in the duration of a short infusion of 5-FU clearly affects the clinical toxicity, but raises the concern of a potentially adverse impact on its antitumor activity. These results suggest the importance of including precise guidelines concerning the time over which 5-FU is given in clinical trials. Having a specified duration of 5-FU infusion is also important if 5-FU dose escalation is considered. PMID- 11269738 TI - Upregulation of gap junctional intercellular communication and connexin 43 expression by cyclic-AMP and all-trans-retinoic acid is associated with glutathione depletion and chemosensitivity in neuroblastoma cells. AB - PURPOSE: Downregulation of gap junctional intercellular communication (GJIC) has been implicated in carcinogenesis. This is a result of altered expression of connexins, the proteins that mediate GJIC, including connexin 43 (Cx43). Our aim was to evaluate the effect of known inducers of Cx43 on the chemosensitivity of the human neuroblastoma cell line IMR-32 to chemotherapeutic agents. METHODS: We examined the effect of dibutyryl-cyclic AMP (db-cAMP) and all-trans-retinoic acid (tRA) on Cx43 and GJIC, glutathione (GSH) and gamma-glutamyl-cysteine-synthetase (gamma-GCS) levels, and glutathione S-transferase (GST) activity. Finally, we performed cell survival assays to measure the response of IMR-32 cells to the chemotherapeutic drugs doxorubicin, melphalan and bis-chloronitrosourea (BCNU), after treatment with db-cAMP and/or tRA. RESULTS: Exposure to db-cAMP led to the upregulation of GJIC and Cx43 expression and phosphorylation. On the other hand, exposure to tRA led to the upregulation of GJIC but Cx43 expression and phosphorylation were not greatly affected. The combination of both agents was more potent in inducing GJIC in comparison to treatment with db-cAMP or tRA alone. Treatment with db-cAMP, but not with tRA, was associated with a significant increase in the cytotoxic effects of the anticancer drugs doxorubicin, melphalan and BCNU as shown by a decrease in their IC50 values. Concomitant exposure to db-cAMP and tRA, however, had a more pronounced effect on cell sensitization to chemotherapy drugs (particularly doxorubicin) than exposure to db-cAMP or tRA alone. Under the db-cAMP and tRA treatment conditions (which upregulate GJIC and modulate drug response), GSH levels were significantly reduced while the levels of GST and gamma-GCS activities remained unchanged. CONCLUSIONS: This study suggests that GJIC plays a role in cellular drug resistance, and highlights the potential use of GJIC modulators in combination with chemotherapy. Also, this is the first study exploring the ability of both db cAMP and tRA to enhance cell chemosensitivity. PMID- 11269739 TI - Differential sensitivity to etoposide (VP-16)-induced S phase delay in a panel of small-cell lung carcinoma cell lines with G1/S phase checkpoint dysfunction. AB - PURPOSE: The highly schedule-dependent cytotoxic agent etoposide (VP-16) is initially effective in the treatment of small-cell lung cancer (SCLC), particularly in multidrug combination chemotherapy. Heterogeneity in cellular sensitivity to cell cycle arrest may underpin the inadequacy of low-dose extended cycle single-agent regimes in tumours with partially dysfunctional apoptotic signalling pathways. We have studied the longevity and dose dependency of cell cycle and to a limited extent the apoptotic responses of a panel of seven unselected SCLC cell lines, screened for TP53 status. METHODS: Cells were analysed using flow cytometry for the cell cycle responses and field inversion gel electrophoresis for apoptotic patterns. The mitotic inhibitor nocodazole was used to assess and correct cell line response data for differences in cell cycle traverse per se. RESULTS: An overall lack of G1/S arrest and muted DNA fragmentation were consistent with the presence of TP53 gene abnormalities. Maximal G2 arrest but with clear recovery potential occurred at an exposure dose (ED, concentration of drug x time) value of approximately 24 microM h. Higher doses (ED values >48 microM h) revealed a wide variation in S phase delay that was independent of population doubling time and could not be compensated for by drug concentration changes alone. CONCLUSION: The results suggest that heterogeneity in the in vitro sensitivity of unselected SCLC cell lines to S phase arrest is demonstrable at ED values projected to be critical for clinical activity. Such variation in S phase responsiveness may reflect differences in checkpoint activation and offer a functional target for the design of more effective combination therapy. PMID- 11269740 TI - Phase II and pharmacokinetic/pharmacodynamic trial of sequential topoisomerase I and II inhibition with topotecan and etoposide in advanced non-small-cell lung cancer. AB - PURPOSE: In vitro and in vivo preclinical models have demonstrated synergistic activity when topoisomerase I and II inhibitors are administered sequentially. Topoisomerase I inhibitors increase topoisomerase II levels and increase cell kill induced by topoisomerase II poisons. We evaluated this hypothesis in a cohort of patients with advanced non-small-cell lung cancer (NSCLC). METHODS: A group of 19 patients with advanced NSCLC (70% adenocarcinoma) received topotecan at a dose of 0.85 mg/m2 per day as a continuous 72-h infusion from days 1 to 3. Etoposide was administered orally at a dose of 100 mg twice daily for 3 days on days 7-9 (schedule and dose derived from prior phase I trials). Total and lactone topotecan concentrations were measured at the end of the 72-h infusion. Blood samples were obtained immediately after each 72-h topotecan infusion in order to measure the mutational frequency at the hypoxanthine phosphoribosyl transferase (HPRT) locus in peripheral lymphocytes. RESULTS: A total of 55 cycles were administered. Toxicity was mainly hematologic with grade 4 neutropenia occurring in 7% of courses. Only one partial response and two stable diseases were observed. The 1-year survival rate was 33%. There was a statistically significant difference between steady-state lactone concentrations between cycle 1 and cycle 2 with decreasing concentrations with cycle 2 (P = 0.02). This was explained by a statistically significant increase in the clearance of topotecan lactone during cycle 2 (P = 0.03). Total but not lactone concentrations correlated with nadir WBC, ANC and platelet levels. Steady-state plasma lactone levels correlated with the mutational frequency at the HPRT locus (P = 0.06). In the one patient with a partial response a sixfold increase in HPRT mutational frequency was observed, which was not seen in patients with progressive disease. CONCLUSION: The combination of topotecan and etoposide in this schedule of administration has minimal activity in adenocarcinoma of the lung. This lack of activity may be due to the delay in administration of etoposide after the topotecan as studies have shown that the compensatory increase in topoisomerase II levels after treatment with topoisomerase I inhibitors is shortlived (<24 h). The HPRT mutational frequency results suggest that the lack of clinical response may be associated with failure to achieve sufficient cytotoxic dose as indicated by a lack of increase in mutational frequency in those patients with progressive disease. HPRT mutational frequency may correlate with plasma steady-state topotecan lactone levels. Future studies should be directed toward earlier administration of topoisomerase II inhibitors after topoisomerase I inhibition. PMID- 11269742 TI - Detection and cytotoxicity of cisplatin-induced superoxide anion in monolayer cultures of a human ovarian cancer cell line. AB - Superoxide anions (O2-) generated by cisplatin [cis-diamminedichloroplatinum (II), DDP] were determined by measuring the chemiluminescence from the luminescence probe, 2-methyl-6-[p-methoxyphenyl]-3,7-dihydroimidazo[1,2-a]pyrazin 3-one (methyl Cypridina luciferin analog, MCLA), in monolayer cultures of a human ovarian cancer cell line (A2780) in physiological saline at pH 7.0. In a time course study, chemiluminescence of MCLA (C-MCLA) showed a peak level at 10 min and a background level at 60 min after the addition of DDP. The intensity of C MCLA increased with increasing concentrations of DDP or MCLA in a limited concentration range, and was significantly correlated (r = 0.960) with the number of A2780 cells. DDP-induced C-MCLA was completely inhibited by the addition of the O2- scavenger, superoxide dismutase (SOD). However, SOD did not decrease DDP cytotoxicity in terms of clonogenic cell survival. These findings suggest that DDP generates extracellular O2-, probably by interaction with the cellular membrane in A2780 cells, and O2- does not lead to cellular damage. PMID- 11269741 TI - Treatment of VX2 carcinoma implanted in the liver with arterial and intraperitoneal administration of oily anticancer agents. AB - PURPOSE: Long-term survival and cure cannot be achieved in patients with unresectable, advanced abdominal cancer, because no chemotherapeutic treatment has definite antitumor activity for malignant solid tumor and its dissemination. In this study, arterial and intraperitoneal administration of oily anticancer agents, which have properties that permit targeted chemotherapy for VX2 carcinoma implanted in the liver, was attempted to achieve long-term survival. MATERIALS AND METHODS: Rabbits bearing VX2 tumors in the liver measuring 1-2 cm in diameter received an arterial injection of 0.2 ml of nitrogen mustard N-Oxide (HN2-O) dissolved in Lipiodol (7.5 mg/ml), a newly developed oily anticancer agent, for the tumor and an intraperitoneal injection of a cocktail of oily anticancer agents for the prevention of intraperitoneal dissemination. RESULTS: Twelve out of thirteen rabbits survived and VX2 cancer was not observed in these 12 rabbits. The controls received a sham operation, an intraperitoneal injection of the cocktail of oily anticancer agents alone, or an arterial injection of HN2 O/Lipiodol alone. In these control groups, 27 out of 29 rabbits died of cancer. To examine the dose form for arterial injection, 14 rabbits received an arterial injection of the simple mixture of HN2-O dissolved in physiological saline and Lipiodol, with an additional intraperitoneal injection of the cocktail. Eight of these 14 rabbits died of enlargement of the hepatic tumor and peritoneal dissemination. CONCLUSION: Long-term survival and cure was achieved in almost all rabbits bearing VX2 tumor in the liver by simultaneous arterial and intraperitoneal injection of oily anticancer agents. PMID- 11269744 TI - Cryptophycin-induced hyperphosphorylation of Bcl-2, cell cycle arrest and growth inhibition in human H460 NSCLC cells. AB - Bcl-2 has been described as a factor that can protect from apoptosis. The protective effect of Bcl-2 may be lost if the protein is phosphorylated. Bcl-2 phosphorylation can be induced by agents that affect microtubule depolymerization or prevent microtubule assembly. In 13 human tumor cell lines there was a high degree of heterogeneity in Bcl-2 protein expression. Human H460 non-small-cell lung carcinoma (NSCLC) cells expressed high levels of Bcl-2 and were selected for study. Western blot analysis for Bcl-2 phosphorylation was carried out after 4 h and 24 h of exposure to cryptophycin 52, cryptophycin 55, paclitaxel or vinblastine. Cryptophycin 52 and cryptophycin 55 were very potent inducers of Bcl 2 phosphorylation. After 4 h of exposure, Bcl-2 phosphorylation was evident with 0.05 nM cryptophycin 52, 0.25 nM cryptophycin 55, 5 nM vinblastine and 50 nM paclitaxel. The hyperphosphorylated form of Bcl-2 was evident after 24 h exposure of H460 cells to 0.25 nM cryptophycin 52 or cryptophycin 55 and 50 nM vinblastine or paclitaxel. The effects of the compounds on the cell cycle paralleled those on Bcl-2 phosphorylation. In H460 cells 90% cell killing was obtained with 0.13 nM cryptophycin 52, 0.2 nM cryptophycin 55, 20 nM paclitaxel and > 100 nM vinblastine after 24 h of exposure as determined by colony formation. In Bcl-2 negative Calu-6 NSCLC cells, 90% cell killing was obtained with 0.03 nM cryptophycin 52, 0.1 nM cryptophycin 55, 11 nM paclitaxel and 0.5 nM vinblastine using the same experimental design. Thus, cryptophycins are potent inducers of Bcl-2 phosphorylation. The cryptophycins were more potent cytotoxic agents in Bcl 2-negative Calu-6 cells than in Bcl-2-positive H460 cells indicating that pathways triggered by Bcl-2 phosphorylation are involved in cryptophycin-induced lethality. PMID- 11269743 TI - P53-independent downregulation of p73 in human cancer cells treated with Adriamycin. AB - P73, a new p53 homologue, has been recently identified as a candidate tumor suppressor gene. PURPOSE: We studied the alterations in p73 in a panel of human cancer cell lines treated with the chemotherapeutic agent, Adriamycin (ADR), in comparison with the changes in p53. METHODS: P73 and p53 mRNA and protein were determined using semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, respectively. ADR cytotoxicity was examined by a trypan blue dye exclusion assay. RESULTS: The cell lines bearing wild-type p53 were more susceptible to ADR than the cell lines bearing mutant p53. ADR treatment resulted in a significant accumulation of p53 protein and mRNA expression in the wild-type p53 cell lines and caused little (slight increase) or no influence on p53 expression in the cell lines with p53 mutation and deletion. However, in striking contrast to the alterations in p53, a decline in p73 at both the protein and mRNA levels was observed in all the cell lines examined following ADR treatment. Further studies indicated that this p53-independent downregulation of p73 was induced by ADR in a dose- and time-dependent manner. Moreover, the p73 protein decline was abrogated by the presence of proteasome inhibitors. CONCLUSIONS: Our findings revealed that although p73 shares a similar structural and functional composition with p53, there is a significant difference in the mechanisms that govern the responses of p53 and p73 to ADR-induced DNA damage. PMID- 11269745 TI - Effect of E7010 on liver metastasis and life span of syngeneic C57BL/6 mice bearing orthotopically transplanted murine Colon 38 tumor. AB - PURPOSE: E7010 is an orally active sulfonamide antitumor agent showing good activity against various subcutaneously inoculated rodent tumors and human tumor xenografts. The purpose of this study was to evaluate the effect of E7010 on liver metastasis and life span of mice bearing orthotopically transplanted murine Colon 38 tumor. METHODS: Orthotopic transplantation of murine Colon 38 tumor as intact tissue yielded hepatic metastasis with a high incidence in about 1 month in C57BL/6 mice, and the mice died in about 2 months with cachexia. In this model, the maximum tolerated dose of E7010 (100 mg/kg per day) was administered orally on various schedules, including for 14 days or daily until death, starting at 14 days after transplantation, or for 8 days from 21 days after transplantation. RESULTS: E7010 showed tumor growth inhibition (T/C=40%) at the orthotopic site similar to that at the subcutaneous site (T/C = 32%) when administered from 14 days after transplantation. When E7010 was started from 21 days after transplantation, it significantly decreased the number of hepatic metastases (control 17.1+/-20.8, E7010 2.6+/-5.3), although inhibition of tumor growth at the orthotopic site was only moderate (T/ C=60%). The administration of E7010 until death produced a significant increase in life span (control 49.8+/ 8.9 days, E7010 62.5+/-6.1 days). Although the tumor weight of the E7010-treated group on the day of death was similar to that of the untreated group (control 1.166+/-0.507 g, E7010 1.211+/-0.632 g), there were significantly fewer liver metastases in the E7010-treated group (control 41.3+/-31.1, E7010 2.0+/-2.0). CONCLUSION: E7010 suppressed tumor growth at both primary and metastatic sites and increased life span in an orthotopic transplantation model of murine Colon 38 tumor in syngeneic C57BL/6 mice. Hepatic metastasis was inhibited more effectively than the growth of the primary tumor. PMID- 11269746 TI - TNF-alpha further augments natural killer cells when co-administered with an interferon inducer to irradiated, leukemic, bone-marrow-transplanted mice. AB - PURPOSE: We have recently demonstrated that the interferon inducer Poly I:C significantly augments both natural killer (NK) cell numbers and the life span of leukemic, irradiated mice given syngeneic bone marrow transplants (SBMT). The cytokine tumor necrosis factor-alpha (TNF-alpha) also stimulates NK cells directly through receptor-ligand mechanisms. We have combined in the present study the NK-enhancing properties of IFN (Poly I:C-induced) and TNF-alpha by giving Poly I:C to leukemic mice for 8 days after irradiation and SBMT, concomitant with TNF-alpha during the first 4 days immediately after SBMT. All mice were sampled at day 9 following irradiation, transplant, and treatment. METHODS: NK cells were identified and quantified by immunoperosidase labeling methods combined with a hematologic staining technique. RESULTS: The data reveal that TNF-alpha, added to the Poly I:C administration protocol, significantly boosted NK cell numbers 2.4-fold over that achieved by Poly I:C alone. CONCLUSIONS: Since the role of NK cells in the immediate post-transplant period is (a) to destroy residual tumor cells, and (b) to produce hemopoiesis-driving cytokines, it appears that two NK cell stimulants are better than one, at least in the crucial, early post-transplant period. PMID- 11269747 TI - Preclinical antitumor activity of two novel taxanes. AB - PURPOSE: Two taxane analogs, BMS-184476 and -188797, were evaluated for their in vitro cytotoxicity and in vivo antitumor activity, and compared with paclitaxel and occasionally docetaxel (Taxotere). METHODS: Cytotoxicity was assessed in vitro using a panel of human tumor cell lines. Several different murine and human tumor models were used in vivo to evaluate the taxane analogs. RESULTS: Both compounds were found to have cytotoxic potency similar to paclitaxel and to partially overcome two different forms of paclitaxel resistance. BMS-184476 was found to be clearly superior to paclitaxel in three human xenograft tumor models: A2780 ovarian carcinoma; HCT/pk, a moderately paclitaxel-resistant colon carcinoma; and L2987 lung carcinoma. Additionally, in the clinically derived TAXOL-unresponsive ovarian carcinoma, HOC79, BMS-184476 performed slightly better than paclitaxel and Taxotere. BMS-184476 and paclitaxel were inactive in two murine model systems, M5076 sarcoma and the paclitaxel-resistant M109/txlr lung carcinoma. Against the parental M109 tumor, both BMS-184476 and paclitaxel performed comparably. BMS-184476 was never found to be inferior to paclitaxel. The other taxane analog, BMS-188797, displayed efficacy superior to paclitaxel in four in vivo tumor models: HOC79, HCT/ pk, M109, and L2987 carcinomas. Like paclitaxel and BMS-184476, BMS-188797 was inactive versus M5076 sarcoma. CONCLUSIONS: Two new taxane analogs were characterized as superior to paclitaxel or Taxotere in several in vivo tumor models. Both BMS-184476 and -188797 are currently in phase I or II clinical trials. PMID- 11269748 TI - Communicating with clinicians. PMID- 11269749 TI - Improved immunoturbidimetric assay for cystatin C. AB - An immunoturbidimetric assay for cystatin C was optimized with respect to assay imprecision. After investigating the optimum pH, polyethylene glycol concentration and specimen volume, two modifications were introduced: an increase in specimen volume to 25 microL; and an extension of the pre-incubation period to 240 s. These modifications produced an assay with between-batch imprecision (coefficient of variation, n = 10 or 11) ranging from 3-9% at 0.72 mg/L to 1.3% at 5.29 mg/L. The assay was susceptible to interference from lipaemia and haemolysis but not bilirubinaemia in both the original and modified protocol. Extending the pre-incubation to 240 s improved tolerance to common interferences and retained assay applicability in the routine clinical setting. PMID- 11269750 TI - Correction formula for carbamylated haemoglobin in diabetic uraemic patients. AB - The measurement of carbamylated haemoglobin is a useful indicator of uraemic state during the preceding few weeks in patients with renal failure. In diabetic uraemic patients with hyperglycaemia, glycation of haemoglobin may interfere with its carbamylation, as both reactions involve the free amino groups of the protein. The aim of this study was to investigate the carbamylation of haemoglobin in the presence of hyperglycaemia. The study included 29 patients with chronic renal failure on regular haemodialysis, 14 diabetic and 15 non diabetic patients, and 10 healthy controls. We found a significant correlation between the degree of haemoglobin carbamylation and mean blood urea concentration in both uraemic and control subjects. Carbamylation of haemoglobin was higher in both diabetic and non-diabetic chronic renal failure patients, but there were no significant differences between the groups regarding mean blood urea concentration or level of haemoglobin carbamylation. Carbamylated haemoglobin per unit of blood urea concentration was lower in the diabetic patients. Using a correction formula to account for the degree of haemoglobin glycation, there was no longer a significant difference in carbamylation per unit of blood urea concentration. In vitro incubation of red blood cells from six healthy and six diabetic non-uraemic patients in 70 mmol/L urea showed a significantly lower carbamylation in the diabetic patients, but there was no significant difference when using corrected carbamylated haemoglobin values. We conclude that glycation of haemoglobin affects its carbamylation and that monitoring of uraemia in a diabetic patient necessitates the use of carbamylated haemoglobin value corrected for the degree of glycation. PMID- 11269751 TI - Decreased longevity in Japanese men, associated with low-molecular-weight apolipoprotein(a) phenotypes. AB - Lipoprotein(a) [Lp(a)] is an established risk factor for atherosclerosis. High plasma Lp(a) concentrations are associated with low-molecular-weight (LMW) apolipoprotein(a) (apo a) phenotypes, which raises the question of whether LMW apo a phenotypes occur less frequently in the elderly. To assess this possibility, we studied apo A phenotype and allele frequencies in Japanese subjects > or = 80 years old (men:women = 40:34) in comparison with those <80 years old (men:women = 221:296). Significantly fewer LMW phenotypes and LMW alleles were observed in > or = 80 year old men compared with those < 80 years old: 2.5% versus 16.1% for the LMW phenotype frequency and 1.3% versus 9.7% for the LMW allele frequency (P<0.05, Fisher's exact probability test). Similar differences were not found in the women. Consistent with this, plasma Lp(a) concentrations were significantly lower in the men > or = 80 years old than in the younger group. These results indicate that LMW apo a phenotypes are associated with a shorter lifespan in men but not in women, possibly reflecting the higher susceptibility of men to atherosclerosis. PMID- 11269752 TI - Effect of lipoprotein lipase on binding of chylomicrons to LDL receptor-deficient Chinese hamster ovary cells. AB - The authors investigated the binding of human plasma 125I-labelled chylomicrons to Chinese hamster ovary (CHO) cells, i.e. native CHO cells are mutant ldl-A7 cells lacking the low-density lipoproteins receptor, in the absence and presence of exogenous bovine milk lipoprotein lipase (LPL) in the culture medium. Only a small amount of binding to either cell was observed in the absence of added LPL. Exogenously added LPL increased the specific binding of chylomicrons to ldl-A7 cells, as well as to native CHO cells. The enhanced binding of chylomicrons to ldl-A7 cells or native CHO cells by LPL was inhibited by heparinase and a monoclonal antibody against LPL (5D2) which recognizes the carboxyl terminal of LPL. However, the enhanced binding was not inhibited by 1 M NaCl, which abolishes the enzymatic activity of LPL in either ldl-A7 cell or native CHO cells. These results suggest that LPL enhances the binding of chylomicrons to heparan sulphate proteoglycans of CHO cells, and that it is the carboxyl terminal of LPL but not the enzymatic activity of LPL that is essential for LPL to mediate the binding of chylomicrons to CHO cells. PMID- 11269753 TI - Apolipoprotein E polymorphism and serum lipoprotein(a) concentrations in a Korean male population. AB - The purpose of this study was to investigate the effect of apolipoprotein E polymorphism on lipoprotein(a) metabolism by comparing serum lipoprotein(a) concentration with apolipoprotein E genotype in a Korean male population whose high molecular weight (HMW) lipoprotein(a) frequency was 95-98%. Serum lipoprotein(a), total cholesterol, triglyceride and high-density lipoprotein cholesterol concentrations were measured and the apolipoprotein E genotype determined in 1189 healthy Korean males. The medians of serum lipoprotein(a) concentration in the apo E 2/3 group (0.105 g/L) and the apo E 3/4 group (0-116 g/L) were significantly lower than that in the apo E 3/3 group (0.155 g/L; P < 0.001). The medians of serum triglyceride were 1.497 mmol/L in the apo E 2/3 group, 1.356 mmol/L in the apo E 3/4 group, and 1.452 mmol/L in the apo E 3/3 group (P<0.05). With the significant difference in the serum lipoprotein(a) concentration in Korean males according to apolipoprotein E genotype, and with the negative correlation between serum triglyceride concentration and serum lipoprotein(a) concentration, it is suggested that apolipoprotein E polymorphism and serum triglyceride participate in the metabolism of lipoprotein(a) with HMW. PMID- 11269754 TI - Troponin T: role in altering patient management and enabling earlier discharge from a district general hospital. AB - The use of troponin T to facilitate early patient discharge was investigated in a prospective study in a district general hospital. Troponin T was measured in 91 patients admitted over a period of 6 months with chest pain but without evidence of myocardial infarction. The main outcome measure was length of hospital stay. A negative troponin T was found in 70 patients. Fifty of these were discharged within 24 h of the troponin result being available and they had a significantly shorter hospital stay than a case control group and a historical control group from the previous 6 months. Troponin T measurement has a role in altering patient management by enabling early discharge, resulting in significant cost savings and increasing bed availability. PMID- 11269755 TI - Comparison of Abbott AxSYM, Behring Opus Plus, DPC Immulite and Ortho-Clinical Diagnostics Vitros ECi for measurement of cardiac troponin I. AB - Myocardial infarction is a common cause of morbidity and mortality in patients with chest pain. The presence of human cardiac troponin I (cTnI) in serum is considered to be a highly specific biochemical marker of acute myocardial infarction. In this study we compare the performances of the Abbott AxSYM, Behring Opus Plus, DPC Immulite and Ortho-Clinical Diagnostics Vitros ECi for the measurement of cTnI. The first two methods use a fluorogenic enzyme-linked immunoassay. whereas the last two use chemiluminescent immunometric assays. All procedures are completely automated. Total percentage coefficients of variation using pooled sera ranged from 5.9 to 6.5% for the AxSYM, 14.4 to 25.6% for the Opus, 6.9 to 9.8% for the Immulite and 4.5 to 5.2% for the Vitros ECi method. The closest correlation between methods, obtained from 120 fresh serum samples, was observed between the Vitros ECi and the Immulite methods, with r=0.99, and the regression line was Immulite cTnI 1.505 (95% confidence interval 1.474-1.536) x Vitros cTnI--0.154 (-0.702 to 0.394). Receiver operating characteristic curves were nearly identical for all assays, and the areas under the curves were 0.972, 0.927, 0.967 and 0.969 for the AxSYM, Opus, Immulite and Vitros ECi methods, respectively. There was a significant difference between the AxSYM and Opus methods (P= 0.036). PMID- 11269756 TI - Thyroid stimulating hormone secretion is a dynamic process. PMID- 11269757 TI - Diagnosis of latent acute intermittent porphyria by genetic analysis. PMID- 11269758 TI - Improved assay characteristics of an immunoturbidimetric assay for cystatin C. PMID- 11269759 TI - Interference by bilirubin in salicylate measurement. PMID- 11269760 TI - Measurement and importance of plasma brain natriuretic peptide and related peptides. PMID- 11269761 TI - Are natriuretic peptides clinically useful as markers of heart failure? PMID- 11269762 TI - Elective extracorporeal membrane oxygenation: an improved perioperative technique in the treatment of tracheal obstruction. AB - The surgical management of children with tracheal stenosis and obstruction is complicated by the perioperative needs of pressure ventilation and indwelling endotracheal tubes. These factors predispose to surgical failure and anastomotic breakdown, restenosis. and pneumomediastinum. The use of extracorporeal membrane oxygenation (ECMO) to manage ventilation during tracheal repair allows better visualization at the surgical site and obviates the need for indwelling endotracheal tubes and high-pressure ventilation. Six children were treated with elective ECMO at a tertiary care hospital. All 6 underwent successful surgical repair, and 4 of the 6 were ultimately extubated. There were no significant complications at the surgical site. There was 1 death from postoperative complications, and 2 patients required tracheotomy. One tracheotomy was performed for upper airway obstruction secondary to retrognathia, and this patient was subsequently decannulated. Medical complications were confined to 2 patients and included sepsis, hyperbilirubinemia, seizure disorder, renal failure, intracranial hemorrhage, and hydrocephalus. Elective ECMO provides a reliable perioperative technique for airway management of children with tracheal stenosis or obstruction. This technique offers the advantage of improved visibility at the operative site and eliminates the need for high-pressure ventilation, thereby likely reducing the risk of perioperative morbidity. PMID- 11269763 TI - Arytenoid prolapse as a consequence of cricotracheal resection in children. AB - Cricotracheal resection (CTR) is a technique introduced comparatively recently for treating severe laryngotracheal stenosis in children. The recognized complications of CTR include recurrent laryngeal nerve damage, anastomotic dehiscence, and restenosis. We describe a further complication of CTR, namely, prolapse of the arytenoid cartilage. The presentation may be late, with symptoms of shortness of breath on exertion and nocturnal stertor with a poor sleep pattern, or the prolapse may be an asymptomatic incidental finding. The diagnosis is performed with flexible nasopharyngoscopy with the patient unanesthetized, or with rigid endoscopy with the patient lightly anesthetized and spontaneously ventilating. The affected arytenoid cartilage is noted to prolapse anteriorly and medially with inspiration, partly obstructing the airway. If treatment is required, endoscopic laser partial arytenoidectomy is effective. In a series of 44 children who underwent CTR, 20 were noted to develop arytenoid prolapse after operation. Twelve were asymptomatic, and 8 required laser arytenoidectomy, 2 of whom now require continuous positive airway pressure for moderate supraglottic collapse. PMID- 11269764 TI - Long-term monitoring of respiration with a mediastinal pressure sensor in dogs. AB - The ability of an implanted mediastinal pressure sensor to produce a stable respiratory signal that could be used to trigger stimulation of upper airway muscles was examined. In 6 dogs, a pressure sensor was secured to the manubrium (4 by wires and 2 by transmanubrial placement). In 6 other dogs, the pressure sensor was placed in the upper anterior mediastinum. The animals were monitored for a minimum of 8 weeks (2 transmanubrial sensors for 12 months). Sensors that were able to maintain a midline position, high in the mediastinum, had the best signals. A caudal sensor position or abutment against an intrathoracic structure caused signal inversion (unusable signals). Transmanubrial placement resulted in a stable signal for 1 year. We conclude that long-term monitoring of respiration with a mediastinal pressure sensor can be successfully performed in dogs, providing an adequate signal for nerve-muscle stimulation. Separation from cardiovascular structures improves signal quality. PMID- 11269765 TI - Molecular markers in head and neck squamous cell carcinoma: their biological function and prognostic significance. AB - Head and neck squamous cell carcinoma affects more than 500,000 people worldwide each year. Despite optimal treatment with surgery, irradiation, and chemotherapy, disease recurrence and progression remains a common and challenging oncological problem. Recently, interest has developed in identifying novel molecular markers that allow identification of those patients at increased risk for locoregional recurrence and death. This article reviews several such molecular markers studied in head and neck cancer, including p53, angiogenesis-related markers, cyclin D1, and epidermal growth factor receptor. The biological function of these markers and the potential clinical implications are discussed. The purpose of this review is to update the otolaryngologist on a rapidly emerging segment of applied translational research in our field. PMID- 11269766 TI - Objective voice analysis after autologous fat injection for unilateral vocal fold paralysis. AB - This study was designed to objectively compare a patient's voice after onset of unilateral vocal fold paralysis (UVFP) to his or her own normal voice, and to compare the results after treatment by intrafold injection of autologous fat. Acoustic recordings were obtained for 2 male patients before thoracic surgery and after the onset of iatrogenic left UVFP. Vocal fold augmentation was performed 10 days after UVFP. The acoustic recordings were repeated within 3 days and at 1 month. The phonation quotient, pitch perturbation quotient, amplitude perturbation quotient, harmonics-to-noise ratio, cepstral peak prominence, and long-term average spectrum were analyzed. All parameters improved after treatment, with a return to preparalytic values for most. During the first month, some deterioration was noted. This is the first study comparing a subject's own normal voice to his or her voice after vocal fold augmentation. We recommend overinjection of fat if vocal fold atrophy is expected. PMID- 11269767 TI - Enhancement of submicroscopic damage of the nasal epithelium by topical allergen challenge in patients with perennial nasal allergy. AB - The purposes of this study were to clarify whether damage of the nasal epithelium exists in patients with nasal allergy, and how the morphology of the epithelium changes after topical allergen challenge. Electron microscopy revealed 2 characteristic features in the nasal epithelium of patients with perennial nasal allergy--an increase in the number of epithelial cells with cytoplasmic vacuoles, and markedly widened intercellular spaces--although these changes were unclear under light microscopy. The density of vacuolated cells significantly increased 24 hours after allergen challenge. Further, the number of eosinophils that were associated with vacuolated cells was significantly higher in patients with nasal allergy than in controls. These morphological changes, thus, were considered to be types of damage to the nasal epithelium associated with nasal allergy. Such changes may be among the causes of nasal hyperreactivity, which is an important feature of nasal allergy. PMID- 11269769 TI - Adenoid cystic carcinoma of the base of the tongue. AB - A retrospective review of 14 patients with adenoid cystic carcinoma of the tongue treated between 1955 and 1997 was performed. Treatment consisted of surgery (n = 2), radiotherapy (n = 2), chemotherapy (n = 1), or combination therapy (n = 9). The 2-, 5-, and 10-year absolute survival rates were 92%, 79%, and 63%, respectively. Seventy-five percent of the patients who died of cancer succumbed to distant metastases. However, long-term survival was common despite a high incidence of local and distant recurrence. The presence of positive surgical margins, the incidence of regional metastases, the incidence of perineural invasion, the initial stage of disease, and the eventual development of locoregional recurrence and distant metastases did not significantly alter the survival rate. Surgical extirpation combined with postoperative radiotherapy is advocated for the treatment of adenoid cystic carcinoma of the tongue. Given the indolent nature of this disease process, surgery should be directed toward conservation of speech and swallowing function. PMID- 11269768 TI - Mucosa-associated lymphoid tissue in middle ear and eustachian tube. AB - The presence of mucosa-associated lymphoid tissue (MALT) was investigated histopathologically in every 20th section from 99 vertically cut, celloidin embedded temporal bone-eustachian tube (ET) specimens. Among specimens from infants and children between 1 month and 7 years of age, MALT was found in 22 of 44 (50%). However, in 26 adults over 18 years of age, MALT was found in only 2 specimens (7.7%), a significantly lower incidence than that in infants and children. Moreover, MALT did not appear in any of the 21 neonates under the age of 1 month. All 99 specimens were classified into 2 groups: 41 specimens with otitis media (OM) and 58 specimens without OM. The presence of MALT was significantly higher in specimens with OM (43.9%) than in specimens without OM (13.8%). Mucosa-associated lymphoid tissue was found in the ET, middle ear, and mastoid process in 18 specimens (43.9%). 5 specimens (12.2%), and 1 specimen (2.4%) with OM, respectively, and in 8 (13.8%), 0, and 0 specimens without OM. In regard to the distribution of MALT, it occurred more frequently in the pharyngeal half of the cartilaginous portion of the ET than in the rest of the ET, middle ear, and mastoid; the presence was significantly greater in the inferior half of the cartilaginous portion of the ET than in the superior half. Inflammatory cell infiltration in the cartilaginous and bony portions of the ET was significantly greater in specimens with OM than in specimens without OM with no MALT. However, even in some specimens without OM, inflammatory cells were found in the ET, particularly in the pharyngeal half of the cartilaginous portion of the ET. These findings suggest that MALT has a close relationship to OM and that it may be a local response to repeated infection. PMID- 11269770 TI - Is pressure dressing necessary after ear surgery? AB - Compressive bandages carry intrinsic risks and complications. The purpose of this study was to assess whether compressive head bandages are necessary after ear surgery to prevent complications such as hematoma or wound infection. A prospective, randomized, controlled study was conducted from August 1, 1993, to June 1, 1999. We randomly assigned 420 consecutive middle ear or mastoid operations to either a pressure bandage group or to a no-pressure bandage group. A careful follow-up was planned, and complications were recorded. In the pressure bandage group, 3 patients had bruising over the pinna and 70 patients had minor skin erythema when the dressing was removed. No patient had bruising or erythema in the no-pressure bandage group. No patient had hematoma, wound infection, or any other wound complication in either group. As a result of this study, we have decided to abandon the use of compression bandages after uncomplicated ear surgery. PMID- 11269771 TI - Hypertrophic neuropathy of the facial nerve. AB - Hypertrophic neuropathy is a peripheral nerve lesion that is histologically characterized by onion bulb formations around axons. This histologic picture, which is usually seen in generalized hypertrophic neuropathies, can occasionally be observed in single nerves as localized hypertrophic neuropathy. Cranial involvement of such localized hypertrophic neuropathy represents a very rare entity; only a few cases have been reported in the literature. We report the history of a progressive facial paralysis with a tumorous enlargement of the seventh cranial nerve that was clinically suspected of being a schwannoma. Pathological examination permitted the diagnosis of hypertrophic neuropathy. PMID- 11269772 TI - Bilateral laryngeal pseudoparalysis in xanthoma disseminatum treated by endoscopic laser medial arytenoidectomy. AB - Xanthoma disseminatum is a rare non-Langerhans' cell histiocytosis, characterized by papular cutaneous eruption, possible mucosal involvement, and frequent association with vasopressin-sensitive diabetes insipidus. Herein we report a case of xanthoma disseminatum with pharyngolaryngeal involvement. In this patient, mucosal xanthomas involving the arytenoid cartilages and the interarytenoid area resulted in laryngeal stenosis and severe impairment of both cricoarytenoid joints' motility. Endoscopic CO2 laser medial arytenoidectomy, according to the technique described by Crumley (1993), and vaporization of interarytenoid xanthomas were successfully performed, thus reestablishing bilateral cordal motility and the laryngeal airway. Four years later, a CO2 laser revision was necessary because of recurrence of xanthomas in the posterior larynx. Two years after the latter operation, the patient has no signs of laryngeal obstruction and has a normal voice quality. This case report suggests that endoscopic medial arytenoidectomy may be successfully used in the treatment of bilateral laryngeal pseudoparalysis secondary to xanthoma disseminatum. PMID- 11269773 TI - Tracheal length of infants under three months old. AB - The measurement of tracheal length in infants is difficult to perform in vivo. In adults, tracheal length may be consistent with age, but in infants, tracheal length may vary much more with age and other factors. This study used video rigid ventilation bronchoscopy to evaluate the length of the airway, concentrating on the population younger than 3 months old. There were 34 infants in this study: 14 boys and 20 girls. The mean length from the superior border of the vocal fold to the carina was 5.04 cm, and the mean tracheal length (from the ridge of the first tracheal ring to the carina) was 4.12 cm. There was no significant difference between boys and girls in the length from the vocal fold to the carina or in the tracheal length. The length from the vocal fold to the carina is best correlated to body weight, followed by body height and age. PMID- 11269774 TI - Long-term changes in off-lesion endocochlear potential after induction of localized lesions in the lateral wall. AB - Localized lesions were produced in various turns of the guinea pig cochlea by means of a photochemical reaction between systemically administered rose bengal dye and green light illumination. The endocochlear potential (EP) was measured at various off-lesion sites, and a morphological examination was performed. In a previous study, this same investigation was done at 3 days, at which time all sites apical to the lesion showed significant EP depression, and damage to the stria vascularis at the lesion was ongoing. In the present 2-week study, the apical EP values were not different from the basal values, and all experimental values were essentially the same as the EP values found in control animals. Morphological examination revealed that the previously damaged structures were greatly repaired. Localized damage and early apical EP depression followed by damage repair and eventual EP recovery could account for the clinical course of certain cases of idiopathic sudden hearing loss involving low-tone deafness. PMID- 11269775 TI - Ligand-gated purinergic receptors are differentially expressed in the adult rat vestibular periphery. AB - To further characterize the pattern of expression of the ligand-gated purinergic P2X receptors in the peripheral vestibular system, we conducted reverse transcription-polymerase chain reaction amplification of P2X1 and P2X2 messenger RNA extracted from adult rat vestibular ganglia (Scarpa's ganglia) and vestibular end organs. Transcripts encoding P2X1 were found in both Scarpa's ganglia and the end organs, but transcripts encoding P2X2 were found only in the vestibular end organs. These results support previous electrophysiological data, and they provide a more complete understanding of the specific role of purinergic (adenosine-5'-triphosphate) transmission in the vestibular periphery. PMID- 11269776 TI - Effect of furosemide on basal lamina anionic sites in guinea pig labyrinth. AB - The authors studied the effects of acute furosemide administration on the basal lamina (BL) anionic sites in the stria vascularis, ampullar crista, and endolymphatic sac by using cationic polyethyleneimine (PEI). Furosemide was intravenously administered to albino guinea pigs with normal Preyer's reflexes. After 20 minutes, the bony labyrinth was removed and processed for histologic evaluation. Under a transmission electron microscope, a marked enlargement of the intercellular spaces was observed in the stria vascularis. The PEI distribution decreased significantly on the capillary BL in the stria vascularis and on the subepithelial BL in the sensory, transitional, and dark cell areas. However, no significant change was observed on the capillary BL or the subepithelial BL in the endolymphatic sac. These findings suggest that acute furosemide administration severely alters the distribution of the anionic sites in the strial capillary BL and in the subepithelial BL in the ampullar crista, but not in the capillary BL or the subepithelial BL of the endolymphatic sac. PMID- 11269777 TI - Rhinoscleroma mimicking nasal polyposis. AB - Rhinoscleroma is increasing in incidence in the United States. It should be considered in patients who are immigrants from endemic countries and present with nasal polyposis that exhibits significant adherence to the nasal septum and relative sparing of the sinuses. Fluoroquinolones are emerging as the adjunctive antibiotic treatment of choice to complement surgical extirpation of the disease. PMID- 11269778 TI - Comparative efficacy of primary angioplasty with stent implantation and thrombolysis in restoring basal coronary artery flow in acute ST segment elevation myocardial infarction: quantitative assessment using the corrected TIMI frame count. AB - Following thrombolysis and primary percutaneous transluminal coronary angioplasty (PTCA) for acute ST segment elevation myocardial infarction, basal flow in the culprit artery is known to influence prognosis. The purpose of this study was to determine if differences exist in basal flow in culprit and nonculprit coronary arteries in patients with acute ST segment elevation myocardial infarction who were treated with thrombolysis or primary PTCA with stent implantation. Twenty patients were randomized to thrombolysis (with recombinant tissue plasminogen activator) and 24 to primary PTCA with stent implantation within 3 hours of onset of acute ST segment elevation myocardial infarction. Coronary angiography was performed 90-120 minutes after thrombolysis or immediately after PTCA with stent implantation and again at 18-36 hours after intervention in both groups. Patients who failed to achieve thrombolysis in myocardial infarction (TIMI) grade 2 or 3 flow were excluded. The corrected TIMI frame count was used as the index of basal coronary artery flow. Early after intervention the mean corrected TIMI frame count in the culprit coronary artery was significantly lower in the primary PTCA with stent group (27.4 +/- 7.7 frames) than in the thrombolysis group (39.8 +/- 10 frames, p < 0.001). Eight thrombolysis patients (40%) and 20 primary PTCA patients (83%, p < 0.01) achieved TIMI grade 3 flow early after intervention. By 18-36 hours after intervention there were no significant differences in the mean correct TIMI frame count between the thrombolysis and primary PTCA with stent groups. There were no significant differences in the mean corrected TIMI frame count between these two groups in the nonculprit coronary artery, either early after intervention or at 18-36 hours. In successfully reperfused coronary arteries following acute ST segment elevation myocardial infarction, primary angioplasty with stent implantation reestablished TIMI grade 2 or 3 flow faster and more effectively than thrombolysis did. PMID- 11269780 TI - Microalbuminuria, pulse pressure, left ventricular hypertrophy, and myocardial ultrasonic tissue characterization in essential hypertension. AB - Microalbuminuria (UAE) may be considered a marker of systemic vascular dysfunction, while pulse pressure (PP) is an indicator of the stiffness of vascular conduits. Both these parameters, together with left ventricular hypertrophy (LVH), are linked to cardiovascular morbidity in hypertensive patients. The aim of this study was the analysis of the possible relationships among UAE, PP, and LVH with ultrasonic myocardial textural parameters, which are altered in hypertensives patients. A group of male (n = 70) essential hypertensive patients (mean age: 58 +/- 7 yr) was analyzed with a group of age comparable normotensive healthy subjects as controls (n = 32). Ambulatory blood pressure monitoring (ABPM) was performed with an oscillometric monitor; UAE was measured by nephelometry. A conventional 2D-Doppler echocardiography (to analyze left ventricular mass: LVM) and a quantitative analysis of the echocardiographic digitized imaging with the use of a calibrated digitization system (to calculate the septum and the posterior wall textural parameters) were performed on all subjects. The myocardial mean gray level was calculated to derive the cyclic variation index (CVI). The CVI was significantly lower in hypertensives both for the septum (- 16.3 +/- 22.8 vs 34.7 +/- 15.3%; p < 0.001) and for the posterior wall (- 15.2 +/- 23.6 vs 38.2 +/- 15.4%; p < 0.001). A significant negative correlation was found between logUAE and the CVI of the septum (r = -0.42; p < 0.001), between the PP and the CVI of the septum (r = -0.40; p < 0.002) and between the CVI and the LVM (r = -0.38; p < 0.001). Multiple regression analysis having as dependent variable the CVI at septum level showed as significantly related independent variables: PP (p < 0.01), logUAE (p < 0.001), and LVM (p < 0.05) (multiple R: 0.76, squared multiple R: 0.57; p < 0.001). It was found that LVM, logUAE, and PP are all correlated with textural parameters, and the CVI can be considered a sensitive parameter in the identification of an abnormal myocardial texture in hypertension. A high level of arterial stiffness and the presence of vascular dysfunction in essential hypertension could participate in the determination of myocardial alterations and permit the identification of patients with the worst prognosis in terms of morbidity or mortality due to cardiovascular events. PMID- 11269781 TI - Recurrent ischemia resulting from left internal mammary artery-to-pulmonary artery fistula. AB - This report describes a case series of recurrent ischemia after coronary artery bypass grafting resulting from left internal mammary artery-to-pulmonary artery fistula. An angiographic demonstration of this fistula is presented. PMID- 11269779 TI - Increased autoantibodies against oxidized low-density lipoprotein in coronary circulation in patients with coronary spastic angina. AB - Oxidized low-density lipoproteins are important in the progression of atherosclerosis. Autoantibodies against malondialdehyde-modified low-density lipoproteins have been reported to be predictive of the progression of atherosclerosis. This study sought to examine whether plasma levels of autoantibodies against oxidized low-density lipoprotein increase in the coronary circulation in patients with coronary spastic angina. The authors examined plasma antioxidized low-density lipoprotein antibody levels (activity unit values (AcU)/mL) simultaneously in the coronary sinus and the aortic root in 20 patients with coronary spastic angina, 23 patients with stable exertional angina, and 15 control subjects by measuring plasma levels of immunoglobulin G (IgG) autoantibodies against malondialdehyde-modified low-density lipoproteins by enzyme-linked immunosorbent assay. The plasma antioxidized low-density lipoprotein antibody levels (AcU/mL) in the coronary sinus increased in coronary spastic angina (38 +/- 16) compared with stable exertional angina (23 +/- 7) and control subjects (20 +/- 6) (p < or = 0.0001). The levels (AcU/mL) in the aortic root also increased in coronary spastic angina (33 +/- 12) compared with stable exertional angina (23 +/- 7) and control subjects (20 +/- 6) (p < 0.005). Furthermore, the coronary sinus-arterial differences of the levels (AcU/mL) were also higher in coronary spastic angina (5 +/- 9) than in stable exertional angina (0 +/- 6) and healthy subjects (-1 +/- 5) (p < 0.05). The generation of malondialdehyde-modified low-density lipoproteins is reported to be associated with atherothrombosis. These findings suggest that elevated levels of autoantibodies against malondialdehyde-modified oxidized low-density lipoproteins in coronary circulation are associated with the development of atherothrombosis from the progression of atherosclerosis rather than with the extent of coronary atherosclerosis in patients with coronary spastic angina. PMID- 11269782 TI - Prevalence of subclavian artery stenosis in patients with peripheral vascular disease. AB - Internal mammary arteries (IMA) as conduits in coronary artery bypass grafting are superior to saphenous vein grafts. If there is subclavian artery stenosis (SAS) proximal to the IMA graft, impairment of flow to the IMA may occur. If the stenosis is severe, retrograde flow from the grafted coronary artery to the brachial artery may lead to angina. Following the identification of 2 cases of angina secondary to subclavian artery stenosis at their institution, the authors prospectively performed arch angiography in a cohort of patients with manifestations of peripheral vascular disease undergoing diagnostic coronary angiography to assess the prevalence of subclavian stenosis. Fifty-two patients were enrolled in the protocol, with 48 patients having technically acceptable studies. Of these 48, 41.6% had measurable stenosis of at least one of the brachiocephalic arteries, with 35% of patients with at least a 30% stenosis of the left subclavian artery and 18.7% with more than 50% stenosis. They conclude that patients with significant peripheral vascular disease undergoing coronary angiography who are potential candidates for revascularization may benefit from arch angiography as part of their initial evaluation. PMID- 11269783 TI - The 1999 French Venous Disease Survey: epidemiology, management, and patient profiles. AB - A recent (1999) Sofres survey of representative samples of the adult French population aged 15 and over showed that almost half this population suffered from lower limb venous complaints and that 43% of them were untreated. Of those treated, 24.2% received venotropics, including 21.5% by prescription, while 6.0% practiced self-medication. Venous disease sufferers form a relatively underprivileged sector of the population in terms not only of age, income, work and living conditions, but also of general health and medical history. Despite its clinical efficacy and potential social utility, venotropic treatment is possible only if backed by adequate state health insurance coverage supplemented by mutual and private insurance schemes. Any restriction to such coverage will only decrease access to prescription venotropics. PMID- 11269784 TI - Association between mitral annular calcification and carotid atheroma. AB - Carotid artery atherosclerosis is a strong predictor of stroke and represents a potential source of cerebral emboli. The aim of this study was to investigate whether an association exists between mitral annular calcification and carotid atheroma. In addition, the characteristics of carotid atheromas were compared between patients with and without mitral annular calcification. The authors found that there was a significant association between the presence of mitral annular calcification and carotid atheroma. Mitral annular calcification in the elderly may be a form of atherosclerosis. PMID- 11269785 TI - Percutaneous balloon mitral commissurotomy during pregnancy. AB - Percutaneous balloon mitral commissurotomy was performed in 16 pregnant women aged 23 +/- 3 years (range, 16-39 years) who had severe mitral stenosis at pregnancies of mean gestational age 25 +/- 6 weeks. Ten patients were in New York Heart Association functional class III, and six patients were in functional class IV at the time of the procedure. All patients were symptomatic despite maximal medical therapy. The procedure was performed with the Inoue balloon. The mitral valve area increased from 0.9 +/- 0.3 to 1.8 +/- 0.3 cm2 (p < 0.05). The mitral valve pressure gradient decreased from 23 +/- 7 to 6 +/- 3 mm Hg (p < 0.05). The left atrial pressure decreased from 28 +/- 8 to 10 +/- 4 mm Hg (p < 0.05). The pulmonary artery pressure decreased from 59 +/- 18 to 33 +/- 12 mm Hg (p < 0.05). Fourteen patients continued their pregnancies to mean gestational age 37 +/- 2 weeks; all infants were healthy. Two patients had premature deliveries more than 1 month after the procedure due to obstetrical reasons. The two newborns died at day 2 of respiratory distress. Eleven women had vaginal deliveries and five had cesarean sections. All clinically improved to New York Heart Association functional class I or II. Excessive blood loss from the femoral puncture site that required transfusion occurred in one patient. Mitral regurgitation increased one degree in four patients, from 0 to 1+. Patients were observed until delivery. None had restenosis. The degree of mitral regurgitation remained unchanged. Percutaneous balloon mitral commissurotomy can be performed safely during pregnancy. It will effectively improve hemodynamics and symptoms in pregnant patients who have severe mitral stenosis and persistent congestive heart failure symptoms despite conventional medical treatment. There are no immediate detrimental effects of radiation on the fetus. Risks are lower than previously reported when surgical commissurotomy was performed. PMID- 11269786 TI - Isolated pulmonic valve endocarditis caused by group B streprococcus (Streptococcus agalactiae)--a case report and literature review. AB - The pulmonic valve is the least commonly involved valve in infective endocarditis. Pulmonic valve endocarditis is usually associated with tricuspid valve endocarditis, and isolated pulmonic valve endocarditis is exceedingly rare. The predisposing factors for developing pulmonic valve endocarditis include a congenitally anomalous pulmonic valve, intravenous drug abuse, and the presence of indwelling intravenous or flow-directed pulmonary artery catheters. More cases of group B streptococcus endocarditis are being reported. The risk factors for group B streptococcus endocarditis include diabetes mellitus, cancer, alcoholism, malnutrition, immunocompromised status, intravenous drug abuse, postpartum and postabortion states, and underlying valvular disease. The vegetations of this type of endocarditis are usually large and have a higher tendency to result in embolism. The presentation of group B streptococcus endocarditis is usually acute and may result in rapid valve destruction if not treated promptly. A case of isolated pulmonic valve endocarditis caused by group B streptococcus, Streptococcus agalactiae, is presented that was diagnosed with multiplane transesophageal echocardiography in a 40-year old, alcoholic, malnourished man, who was successfully treated with intravenous penicillin G. The literature on the isolated pulmonic valve endocarditis caused by group B streptococcus is reviewed. PMID- 11269787 TI - Ergotamine-induced acute vascular insufficiency of the lower limb--a case report. AB - Ergotamine-containing drugs are widely used in the treatment of acute migraine attacks. Spastic vasoconstriction is one of the possible side effects usually affecting the lower extremities and sometimes leading to gangrene. A 28-year-old woman was hospitalized for severe acute arterial insufficiency of the limbs. The initial surgical approach was not successful since the diagnosis was missed. Overuse of ergotamine derivative was acknowledged by the patient, who had a long history of migraine headaches. Therefore, the patient was treated conservatively with intravenous heparin and prostaglandin infusion and sympatheticolysis via epidural catheter. The vascular complications, angiographic findings, and different modalities of treatment of ergotamine-induced peripheral vascular insufficiency of the lower limb are reviewed. PMID- 11269789 TI - Regarding "myocardial infarction with acute insulin poisoning". PMID- 11269788 TI - Ehlers-Danlos syndrome type IV and multiple aortic aneurysms--a case report. AB - Beside atherosclerosis, aortic aneurysms can be part of the clinical spectrum of many systemic diseases, including infectious, inflammatory, genetic and, less often, congenital disorders. A 48-year-old white man presented with multiple large aneurysms of the aorta and its main branches. Medical history was unremarkable except for the presence of a softened abdominal mass since he was 28 years old. On the physical examination, an arterial murmur was heard over the left carotid artery and a palpable mass was noted in the whole right side of the abdomen. No skin or joint abnormalities were noted. Aortography, computed tomography, and magnetic resonance angiography showed multiple large aneurysms of the descending thoracic and abdominal aorta. Aneurysms of the innominate, left subclavian, and carotid arteries were also seen. This case resembles those previously reported, in which multiple aortic aneurysms were associated with abnormalities of the type III procollagen gene (COL3A1). Although the classic stigmas of the Ehlers-Danlos syndrome type IV were lacking, this genetic disease may be the cause of the multiple aneurysms in this patient. PMID- 11269790 TI - Role of impaired lower-limb venous innervation in the pathogenesis of the chronic fatigue syndrome. AB - BACKGROUND: In patients with acute orthostatic hypotension, there is excessive pooling of blood in the legs, which may result from the strikingly subnormal compliance that is demonstrable in the pedal veins during norepinephrine infusion. The common occurrence of delayed orthostatic hypotension and/or tachycardia in the chronic fatigue syndrome (CFS) led to the present studies of foot vein compliance in CFS patients with a linear variable differential transformer. METHODS: Seven patients with CFS were compared with 7 age- and gender matched healthy control subjects in their blood pressure, heart-rate, and plasma norepinephrine responses to prolonged standing and in measurements of their foot vein contractile responses to intravenous norepinephrine infusions with the linear variable differential transformer. RESULTS: Excessive, delayed (usually after 10 min) orthostatic reductions in systolic and diastolic blood pressure (P < 0.01) and inconsistently excessive increases in heart rate were found in the CFS patients, in whom venous compliance in response to infused norepinephrine was significantly reduced (P < 0.05). CONCLUSIONS: In these patients with CFS, delayed orthostatic hypotension was clearly demonstrable, and, as in previously reported patients with orthostatic hypotension of acute onset, this was associated with reduced pedal vein compliance during norepinephrine infusion, implying impaired sympathetic innervation of foot veins. The rapid symptomatic improvement demonstrated in previous studies of CFS patients during correction of orthostatic venous pooling by inflation of military antishock trousers (MAST) to 35 mm Hg may suggest that excessive lower body venous pooling, perhaps by reducing cerebral perfusion, is involved in the orthostatic component of fatigue in these patients. PMID- 11269791 TI - Increased excretion of N-acetyl-beta-D-glucosaminidase and beta2-microglobulin in gestational week 30. AB - BACKGROUND: Little is known about when the urinary excretion of a combination of N-acetyl-beta-D-glucosaminidase (NAG) and beta2-microglobulin (beta2MG) concentration [relative to creatinine (Cr)] reaches maximal values during uncomplicated normotensive pregnancy. This study was thus designed to analyze when urinary excretion of biochemical parameters was increased during normotensive pregnancy. METHODS: NAG, beta2MG, total protein, albumin, and Cr were simultaneously measured in random (untimed) midstream urine samples from 22 healthy nonpregnant women and from 82 normotensive pregnant women (22 in gestational week 20, 25 in week 30, and 35 in week 37). RESULTS: NAG/Cr and beta2MG/Cr ratios were significantly higher (P < 0.01-0.05) in the normotensive pregnant women in gestational week 30 than in the nonpregnant control subjects and normotensive pregnant women in gestational week 20. The NAG/Cr and beta2MG/Cr ratios showed maximal values in gestational week 30. The total protein/Cr ratio was significantly higher in gestational weeks 20, 30, and 37 than in the control subjects. The albumin/Cr ratio was significantly higher in women in gestational week 30 and 37 than in women in gestational week 20 and in the control subjects. CONCLUSIONS: The excretion of both NAG and beta2MG relative to Cr was increased and showed the maximal values in gestational week 30 during normotensive pregnancy. The increase in a tubular enzyme (NAG) might be caused by renal tubular damage, and that in a low molecular weight protein (beta2MG) might result from decreased renal tubular reabsorption. These findings suggest that renal tubular damage and reabsorption dysfunction were increased in gestational week 30. PMID- 11269792 TI - Poststreptococcal reactive arthritis in adults: long-term follow-up. AB - BACKGROUND: Poststreptococcal reactive arthritis (PSReA) is a recognized inflammatory articular syndrome that follows group A streptococcal infection in persons not fulfilling the Jones criteria for the diagnosis of acute rheumatic fever. Characteristic features include nonmigratory arthritis, lack of response to aspirin or nonsteroidal anti-inflammatory agents, and the presence of extra articular manifestations, including vasculitis and glomerulonephritis. Whether or not patients with PSReA develop carditis is a point of contention. METHODS: We analyzed the clinical features, laboratory findings, response to therapy, and outcome in patients diagnosed with PSReA between 1983 and 1998 and observed through April 2000. All patients were contacted, reexamined, and repeat antistreptolysin, rheumatoid factor, C3 and C4 complement components, and echocardiograms were performed. RESULTS: Seventeen patients (4 men and 13 women) were included. All were of low socioeconomic status. All patients had acute severe arthritis that began shortly after a sore throat episode. Extra-articular involvement including tenosynovitis, vasculitis, and glomerulonephritis was relatively common. More importantly, none exhibited clinical and/or echocardiographic evidence of cardiac involvement. Longterm antibiotic therapy was not given. CONCLUSION: Cardiac involvement did not occur in this group of patients with PSReA. Prolonged prophylactic antibiotic therapy may not be required for adult patients presenting with PSReA. PMID- 11269793 TI - School absenteeism, parental work loss, and acceptance of childhood influenza vaccination. AB - BACKGROUND: Influenza causes school absenteeism and may cause parents to miss work to care for sick children. However, it is not known whether these factors influence parental acceptance of childhood vaccination. METHODS: A survey was mailed to parents of 1,805 children attending 3 elementary schools. It included questions about school absenteeism and employment status for adults who stayed home to care for an ill child. Parents were asked if they would consider vaccinating their child against a common wintertime respiratory virus. RESULTS: Of the 972 surveys returned (54% return rate), 954 could be analyzed. Only 13% of respondents stated that they would not consider vaccination for their child. Sixty-nine percent of children had been absent from school because of a nonasthma respiratory illness, with an average of 2.54 days missed per child. Among respondents whose child had missed any school, 33% would definitely consider vaccination compared with 24% of respondents whose child had not missed school (P < 0.01). As children missed more school days, vaccine acceptance increased. In 53% of families, an adult had to miss work to care for the ill child. Vaccine acceptance was higher if an adult caretaker had to lose time from work because of a child's illness (35% versus 25% for non-working caretakers, P < 0.01). CONCLUSION: Vaccine acceptance was closely linked with the amount of absenteeism caused by respiratory illness in the previous year. Parents who had to miss work to care for an ill child were more accepting of the vaccine than were other parents. PMID- 11269794 TI - NSAIDs and the kidney revisited: are selective cyclooxygenase-2 inhibitors safe? AB - Selective cyclooxygenase-2 (COX-2) inhibitors have provided relief for patients suffering from chronic pain and other inflammatory conditions and have reduced adverse gastrointestinal effects. The documented reduction in gastric erosions, ulcerations, and perforations during the use of COX-2-selective inhibitors raises the question: would the kidney be similarly spared? Our understanding of these enzyme isoforms in the kidney is incomplete. However, kidney tissue seems to possess "constitutive" or homeostatic COX-2 enzyme, suggesting a role for prostaglandins produced by this isoform. In addition, studies evaluating the renal effects of the selective nonsteroidal anti-inflammatory drugs (NSAIDs) are inconclusive, and available data on the renal effects of COX-2-selective inhibitors are conflicting. Inadequate numbers, varied baseline patient characteristics, and different doses and lengths of drug treatment hampers comparison of the small number of clinical investigations available for review. Therefore, this article reviews the role of cyclooxygenase enzyme activity and associated prostaglandins in the kidney and the adverse renal effects of nonselective NSAIDs. We also touch on the COX-1/COX-2 selectivity of NSAIDs, the localization of COX enzymes in kidneys, and clinical studies examining the renal effects of selective COX-2 inhibitors. PMID- 11269796 TI - Hidradenitis suppurativa and acne conglobata associated with spondyloarthropathy. AB - Hidradenitis suppurativa and acne conglobata are well-described chronic dermatologic diseases. Although the exact incidence of these disorders is unknown, both are relatively uncommon conditions. The incidence of spondyloarthropathy is less than 1% in the general population. Therefore, a triad of hidradenitis suppurativa, acne conglobata and spondyloarthropathy is a rare syndrome described only in a few case reports in the literature. We report a case of hidradenitis suppurativa and acne conglobata associated with spondyloarthropathy. PMID- 11269795 TI - A 41-year-old, HIV-infected man with fever, pulmonary nodules, and pancytopenia. PMID- 11269797 TI - Unexplained systemic symptoms and Gallium-67--guided decisions. AB - Over 3 months, a healthy man developed prominent systemic symptoms that defied investigation. Physical examination was noncontributory, and extensive studies revealed only a marked acute-phase response associated with increased serum IL-6 levels. A whole body Gallium-67 scan was crucial in diagnosis, directing attention to high uptake in the left paraspinal and psoas muscles. Open surgical excision biopsy was performed, guided by intraoperative use of a gamma-probe. Removed tissue was diagnosed as diffuse, large B-cell non-Hodgkin lymphoma of muscle (stage IE), a rare extranodal lymphoma. Cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) therapy was given, and the patient became asymptomatic with normal blood tests and was thought to be in remission. However, a repeat Gallium-67 scan revealed recurrent multifocal disease and salvage chemotherapy was instituted. A 47,XXY karyotype (Klinefelter syndrome) was later identified, possibly associated with the lymphoma. PMID- 11269798 TI - Microscopic polyangiitis presenting as idiopathic pulmonary fibrosis: is anti neutrophilic cytoplasmic antibody testing indicated? AB - We report a 55-year old woman with microscopic polyangiitis who presented with idiopathic pulmonary fibrosis and 1 year later developed hematuria and proteinuria. She had a high serum level of perinuclear anti-neutrophilic cytoplasmic antibodies. Renal angiogram was normal. The diagnosis of microscopic polyangiitis was confirmed by renal biopsy, which showed pauci-immune crescentic glomerulonephritis. The patient received immunosuppressive therapy and improved markedly. Consideration of small vessel vasculitis is important in the differential diagnosis of idiopathic pulmonary fibrosis. PMID- 11269799 TI - Primary lymphoma of the esophagus in a chronically immunosuppressed patient with hepatitis C infection: case report and review of the literature. AB - Adenocarcinoma and squamous cell carcinoma account for the vast majority of esophageal malignancies. Other malignancies that can involve the esophagus include melanoma, sarcoma, and lymphoma. Gastrointestinal involvement with lymphoma has a variable incidence, as reported in the literature. However, primary involvement, as defined by Dawson, is extremely rare. Lymphoma has been linked to immunosuppressive conditions (such as AIDS), medications, and transplantation. We present what we believe to be the first case of primary esophageal lymphoma in a patient on long-term immunosuppression with azathioprine who was also infected with the hepatitis C virus (HCV). HCV has been postulated to have a relationship with B cell lymphomas. PMID- 11269800 TI - Lymphocytic hypophysitis in a man presenting with hypercalcemia. AB - A 59-year-old man with a 30-year history of type 2 diabetes mellitus presented with fatigue, confusion, and weight loss over a 3-month period. He was found to be hypercalcemic (11.8 mg/dL) and dehydrated, and his hypercalcemia improved with intravenous fluids. While in the hospital, he developed hyponatremia, hypoglycemia, and hypotension. He was found to have a subnormal cortisol level of 2.3 microg/dL at baseline, which increased to only 5.6 microg/dL 60 minutes after a 250-microg corticotropin intravenous stimulation test. The patient developed pneumonia and adult respiratory distress syndrome and died of an acute myocardial infarction. During the autopsy, he was found to have lymphocytic hypophysitis with a severe reduction in corticotropin-producing anterior pituitary cells. No malignancy was identified at autopsy. He is the first male patient to be described in the literature who presented with hypercalcemia caused by lymphocytic hypophysitis. PMID- 11269801 TI - Allergic bronchopulmonary disease caused by Bipolaris hawaiiensis presenting as a necrotizing pneumonia: case report and review of literature. AB - We report a case of allergic bronchopulmonary disease caused by Bipolaris hawaiisensis in an immunocompetent host, presenting with symptoms and radiographic findings suggestive of necrotizing pneumonia. Cultures of the plugs and bronchial washing yielded the pathogenic fungi. Laboratory tests revealed eosinophilia and elevation of serum IgE. This patient was successfully treated with steroids, amphotericin B lipid complex, and itraconazole. Review of 10 previously reported cases and their clinical manifestations and treatment are presented. PMID- 11269802 TI - By the way, doctor. Recently I saw an advertisement for a new product called Slenderstrip that's supposed to help you lose weight without dieting. That sounds too good to be true. Can you tell me anything about it? PMID- 11269803 TI - US healthcare system too geared to acute medicine. PMID- 11269804 TI - Spinal cord monitoring in neuromuscular scoliosis. AB - This article reviews the use of spinal cord monitoring in neuromuscular scoliosis, a condition having a higher incidence of true positive results than idiopathic scoliosis. While somatosensory cortical evoked potentials (SCEP) are unreliable, somatosensory spinal evoked potentials (SSEP) are possible to obtain in most cases and a method using an epidural electrode is described. The '50% rule' is satisfactory having good specificity and sensitivity with it rare for post-operative paralysis to have occurred undetected. The spinal cord in these cases appears to have increased susceptibility particularly during the passage of sublaminar wires with the incidence of complications reduced using modern instrumentation. PMID- 11269805 TI - Surgical management of paralytic scoliosis in myelomeningocele. AB - A retrospective analysis of 54 patients with paralytic scoliosis due to myelomeningocele, who underwent surgical treatment, was performed. The aim of this study was to compare different surgical techniques and to identify clinical parameters influencing primary and midterm results. Three surgical techniques were used: 1) group I, posterior fusion/instrumentation; 2) group II, anterior fusion/no instrumentation combined with posterior fusion/instrumentation; and 3) group III, anterior and posterior fusion/instrumentation. Average age at surgery was 13.1 years. A preoperative scoliosis angle of 90 degrees [interquartile range (25th-75th percentile) (IQR), 76-106 degrees] was primarily reduced to 38 degrees (IQR, 30-50 degrees). At final follow-up (mean, 3.3 years), correction deteriorated to 44 degrees (IQR, 38-65 degrees). The group III procedure resulted in a better midterm correction of scoliosis compared with group I (P = 0.02). The extension of anterior fusion correlated with primary and midterm correction of scoliosis (P < 0.03). Patients with a thoracic level of paralysis had a higher relative loss of correction compared with patients with a lumbar level (P < 0.06). This finding can be attributed mostly to group I patients (P = 0.011). Hardware complications occurred in 16 patients (30%). Relative loss of correction among these patients was high (P < 0.01) and relative midterm correction low (P = 0.001). We recommend anterior and posterior fusion, each with instrumentation for the treatment of paralytic scoliosis in myelomeningocele. In patients with a thoracic level of paralysis, the two-stage procedure is mandatory to reduce the risk of hardware complications and subsequent major loss of correction. PMID- 11269806 TI - Spinal stabilization in Duchenne muscular dystrophy: principles of treatment and record of 31 operative treated cases. AB - The aim of this study was to report results of prophylactic spinal stabilization in patients with Duchenne muscular dystrophy. There is still debate regarding the ideal instrumentation. A prospective study of a consecutive series of 31 patients stabilized with the ISOLA system from D2 to S1 will be presented. The mean follow up was 22 months (range, 1-60 months). The evaluation of the Cobb angle and pelvic obliquity revealed the following: 1) Cobb angle: preoperation, 48.6 degrees (range, 22-82 degrees); postoperation, 12.5 degrees (range, 0-30 degrees); follow-up, 12.5 degrees (range, 0-42 degrees); and 2) pelvic obliquity: preoperation, 18.2 degrees (range, 3-40 degrees); postoperation, 3.8 degrees (range, 0-13 degrees); follow-up, 5.1 degrees (range, 0-14 degrees). Spinal stabilization with the ISOLA system was found to be a suitable treatment for scoliosis owing to Duchenne muscular dystrophy. It should be carried out after loss of ambulation as soon as a progressive curve of more than 20 degrees is documented. The complication rate was found to be high. PMID- 11269807 TI - Epiphyseal changes after proximal femoral osteotomy. AB - This study was conducted to evaluate the risk factors for epiphyseal changes suggestive of osteonecrosis after proximal femoral osteotomy for hip subluxation associated with cerebral palsy. Forty-eight children with 94 hips were reviewed. Two observers rated the radiographs using a written protocol on two occasions each so that reproducibility of these observations could be assured. Concomitant pelvic osteotomy proved to have the greatest association with risk of epiphyseal changes. These findings, suggestive of osteonecrosis, were present in 7 of 68 (10%) hips that had isolated femoral osteotomy, and in 12 of 26 (46%) hips that had concomitant pelvic osteotomy. PMID- 11269808 TI - Salter osteotomy for treatment of acetabular dysplasia in developmental dysplasia of the hip in patients under 10 years. AB - Serial radiographs of 44 hips in 39 patients undergoing Salter innominate osteotomy for the treatment of dysplastic acetabulum owing to developmental dysplasia of the hip were reviewed. The hips were also evaluated clinically at the last follow-up examination, 7 years to 13 years postoperatively. At 7 years to 13 years postoperative, excellent or good clinical results were assessed in 43 hips (98%), and excellent or good radiographic results in 32 hips (73%). In patients with a postoperative center edge (CE) angle > 24 degrees, the CE angle remained significantly greater throughout the follow-up period compared with patients with a postoperative CE angle < or = 24 degrees. A positive correlation was found between the degree of operative CE angle correction and radiographic findings 7 years to 13 years postoperatively. PMID- 11269809 TI - Chiari osteotomy in children and young adults. AB - From 1972 to 1991, 50 Chiari procedures were performed in 41 patients. The follow up period ranged from 3 years to 24 years and 3 months, with an average of 7 years. Age of the patients at operation averaged 10 years and 6 months (3 years and 1 month to 28 years and 6 months). The best results were obtained in patients older than 10 years of age. There was no statistical relationship between the technical details of Chiari osteotomy (angle of Chiari, displacement and distance to the femoral head) and the results. PMID- 11269810 TI - Wagner multiple K-wire osteosynthesis to correct coxa vara in the young child: experience with a versatile 'tailor-made' high angle blade plate equivalent. AB - In 1978, Wagner described a technique using multiple Kirschner wires (K-wires) to stabilize an intertrochanteric osteotomy performed for the correction of coxa vara in small children. Multiple K-wires are used to create a custom high-angle blade plate for valgus osteotomy. The authors have evaluated a retrospective series of 17 Wagner intertrochanteric osteotomies that were performed in 10 children with coxa vara between the ages of 1 year and 8 years. The neck-shaft angle was corrected from 93.5 degrees to 129.5 degrees at long-term follow-up, and the Hilgenreiner epiphyseal angle was corrected from 71 degrees to 37.6 degrees at long-term follow-up. Revision surgery was performed on five hips with inadequate initial surgical correction. Complications included a single broken K wire, a femur fracture after hardware removal, and one hip developed avascular necrosis postoperatively. PMID- 11269811 TI - The value of early postoperative bone scan in slipped capital femoral epiphysis. AB - The purpose of this study was to evaluate the sensitivity and predictive value of early postoperative bone scan for detection of avascular necrosis (AVN) of the femoral head after surgical treatment of slipped capital femoral epiphysis. We reviewed records of 49 patients (64 hips) operated on between 1980 and 1997 with a mean follow-up of 3 years. Sixty-one out of 64 hips went through an early postoperative bone scan. The three hips that developed AVN showed significant loss of radionuclide uptake. There were neither false-positive or false-negative cases in this series. Early postoperative bone scan has an excellent sensitivity and predictive value for detection of AVN after surgical treatment of slipped capital femoral epiphysis. PMID- 11269812 TI - Femoral fractures in children from 4 years to 10 years: conservative treatment. AB - Some authors have widened the indications for surgical management of isolated femoral shaft fractures in children between 4 years and 10 years of age. We address this study to evaluate the results of such femoral fractures treated conservatively in 41 children. All fractures were closed, isolated, and diaphyseal. The mean age was 6.5 years (standard deviation, 1.7 years) and the mean follow-up was 2.3 years (standard deviation, 1.7 years). All fractures were managed conservatively with skin traction (mean hospitalization time, 20.7 days), alignment of the fragments was serially followed by X-ray, and a spica cast was applied (9.7 weeks), usually without a general anesthesia. Angular deformity was assessed by measurement of the fracture-site diaphyseal angle as well as by measurement of the interphyseal angle described by Wallace and Hoffman. No significant complications were recorded regarding leg-length discrepancy, deformity, refractures, etc. Mean cost is not a factor in determining method of treatment at our hospital. We feel that this type of fracture in the 4 years to 10 years age group can be safely treated with a conservative approach. PMID- 11269813 TI - Scoliosis in cerebral palsy: an overview and recent results. AB - The natural history of scoliosis in cerebral palsy as well as the treatments that include observation, bracing, and surgery are reviewed. If surgery is indicated, a careful preoperative evaluation is essential, focusing especially on the patient's nutritional, gastrointestinal, neurologic, and orthopedic systems. Intraoperative and perioperative issues such as blood loss control are reviewed. The Luque-Galveston method of instrumentation for scoliosis secondary to cerebral palsy is our preferred method of treatment. Our recent results compare favorably with the literature with fewer complications. PMID- 11269814 TI - Epidemiological features of supracondylar fractures of the humerus in Chinese children. AB - We studied 450 children with a supracondylar fracture of the humerus in the period 1991 to 1995, and were able to collect full management details in 403 of them (253 boys and 150 girls). The median age at presentation was 6 years (6.6 years in boys, and 5 years in girls), with the nondominant humerus 1.5 times more commonly injured. Fifteen percent of children presented more than 1 day after the injury. Garland type III fractures constituted 45% of cases, type I 30%, and type II 24%, with flexion type fractures present only in 1% of the children. A nerve injury was associated with the fracture in 19 cases. Although the radial pulse was not palpable at presentation in nine patients, only one child had diminished distal circulation requiring exploration. Concomitant fractures were present in 14 patients. Elbow hyperextension was greater than in a comparable group of noninjured children. Open reduction was necessary in 20% of these children, most being managed by manipulation under anaesthesia, at times associated with percutaneous Kirschner wiring. The hospital stay was 2 days or less in two-thirds of the patients, with more than 90% discharged home within 1 week of admission. In conclusion, many Chinese patients attend hospital later than their Western counterparts, and the rate of flexion-type injuries is low. PMID- 11269816 TI - Bilateral symmetric stress fractures in a toddler. AB - Stress fractures are a common injury among adolescent athletes and military recruits. The increase in child participation in organized sport activities has contributed to the inclusion of the skeletally immature age group among those who may suffer from this problem. Bilateral simultaneous symmetric tibial stress fractures that are infrequent in older children are even more rare in toddlers. This entity may cause a diagnostic problem as it must be differentiated from infectious disease, acute trauma or even from the result of a battered child. PMID- 11269815 TI - Fractures of the lateral process of the talus in children. AB - Fractures of the lateral process of the talus are an uncommon injury, which are often misdiagnosed as severe ankle sprain. This error may result in inappropriate treatment of an intraarticular fracture, with subsequent posttraumatic arthrosis. To date, only one fracture of a lateral talar process has been reported in a child, in whom delayed diagnosis and initial mismanagement led to a suboptimal result. The sport of 'snowboarding', which is gaining in popularity, has been significantly associated with fractures of the lateral talar process, leading some authors to dub this fracture 'Snowboarder's Fracture'. This and the ever increasing incidence of major trauma lead us to believe that this fracture will be encountered more frequently, even in the pediatric population, as the two factors mentioned do not pass over this group. We report lateral talar process fractures in two children: one in a 9-year-old girl and one in an 11-year-old boy, the latter associated with talar neck and body fractures. Timely diagnosis enabled prompt open reduction and internal fixation, preventing subtalar arthrosis. We discuss the pertinent anatomy and mechanism, and present the clinical picture, imaging studies and treatment. Two important points are exemplified by these cases. First, this fracture, although rare, does occur in children, and should be sought in appropriate settings. Second, despite the severe talar injury in the 11 year old, early diagnosis and intervention conserved foot function. PMID- 11269817 TI - Obstetric dislocation of the thoracic spine: case report and review of the literature. AB - A fracture dislocation of the upper thoracic spine with spinal cord injury is reported in a neonate. This rare injury is associated with attendant predisposing obstetric circumstances (breech transverse presentations, large baby size) that can alert clinicians of potential problems and aid in the diagnosis of neonatal hypotonia and paralysis. PMID- 11269818 TI - Large cell lymphoma of bone presented by limp. AB - We present a rare case of anaplastic large cell lymphoma of the bone in the leg of a child. The patient initially presented with suspected osteomyelitis of the fibula and was treated by antibiotics without apparent success. Thereafter, an open biopsy of the lesion was performed and the correct diagnosis was established. This rare case demonstrates the difficulties that a treating physician meets in establishing the correct diagnosis in a child presenting with limping. A review of the pertinent literature is introduced. PMID- 11269819 TI - Radiology administrators' opinions of education. AB - This article reports results of a survey assessing the opinions of California radiology administrators regarding the importance of the baccalaureate degree in radiologic technology. Results revealed no relationship between the degree and employment, retention, promotion, salary, career orientation or willingness to cross-train. PMID- 11269820 TI - Stress, burnout and hardiness in R.T.s. AB - This article reports results of a survey assessing the relationship among occupational stress, "personality hardiness" and burnout in 95 radiographers employed in Connecticut hospitals. Regression analyses indicated a positive correlation between burnout and occupational stress, and an inverse relationship between personality hardiness and burnout. Personality hardiness had a beneficial effect at all stress levels. PMID- 11269821 TI - Violence in the workplace. AB - Health care professionals are at high risk for on-the-job violence. This article discusses the scope and nature of violence in the health care workplace, how to prevent it and what to do if it occurs. PMID- 11269822 TI - Pediatric skeletal trauma. AB - Children with traumatic injuries present an emotionally and professionally demanding situation for radiologic technologists. A child's fear and anxiety, developmental stage and the psychological impact on parents and caregivers all affect the technologist's ability to perform a rapid but technically excellent exam. This article examines the specific causes and presentations of unintentional (as opposed to abuse-related) skeletal trauma in children, with emphasis on the role of imaging. The differences between children and adults in terms of injury patterns, healing and complications are discussed, along with efforts to reduce the incidence of childhood skeletal trauma. PMID- 11269823 TI - Are you sure you want to be a manager? AB - Management has a substantial effect on organizational outcome and is critical for the organization in terms of obtaining long-term goals. Therefore, the selection of successful managers is crucial to attaining organizational objectives. However, a staff technologist who functions very successfully in his or her present role may not have the characteristics needed for successful management. Unfortunately, success in one's current, nonsupervisory position is usually the very attribute that is considered when a management position is offered. Because of this, there are many who fail miserably in the management role. These managers are ill-suited for the duties they must perform, unhappy in the position, disliked and seen as ineffectual by their subordinates and colleagues. It is essential that any individual who is offered a management position critically evaluate his or her abilities and personality before accepting the offer. For a candidate who is unsure of his or her abilities, tests are available that can assist in measuring attributes such as leadership behavior, power management, power orientation, conflict management, perceptions of organizational climate and the individual's "organizational fit" with the organization offering the position. These instruments should be used with caution because their reliability and validity in predicting management success depend on a wide variety of factors. Ultimately, it is the prospective manager's personal judgment that must be the final determinant of whether a management position is the best choice. PMID- 11269824 TI - Reactions to iodinated contrast. PMID- 11269825 TI - Osteoporosis and bone mineral density. PMID- 11269827 TI - Change of heart. PMID- 11269826 TI - Trends in examinee age. PMID- 11269828 TI - Microcoils improve MR imaging. PMID- 11269829 TI - Frank Lautenberg. Long-time US Senator from New Jersey. PMID- 11269831 TI - New OIG compliance guidelines. PMID- 11269830 TI - Prevention of perinatal HIV transmission. AB - Preventing perinatal HIV transmission is a multistep, multidisciplinary process that includes ensuring women's access to early prenatal care, acquiring knowledge about the HIV status of pregnant women, educating them regarding HIV infection and its transmission, and prescribing antiretroviral agents to women with HIV infection and to the HIV-exposed neonate and ensuring their consistent use. It also includes mobilizing social and supportive services for these patients and their families. The collaborative nature of this care and treatment options available are discussed in this article. PMID- 11269832 TI - Recruiting and retention tips and techniques. AB - By incorporating today's technology and common sense into their recruiting and retention efforts, businesses will update age-old practices and reduce unnecessary costs. PMID- 11269833 TI - Aetna US Healthcare FAQs. PMID- 11269834 TI - Dr. Y.T. Johnson. One of a kind. PMID- 11269835 TI - Book end? Budget cuts in Minnesota's medical education. PMID- 11269837 TI - MMA legislative priorities advancing as session unfolds. PMID- 11269836 TI - May the (work) force be with us. PMID- 11269838 TI - Investing in the future of medicine. PMID- 11269839 TI - Mental health issues gaining attention in 2001. PMID- 11269840 TI - Assessing latex allergy among health care employees using workers' compensation data. AB - Latex allergies among health care workers have garnered considerable attention from medical researchers and practitioners. However, the majority of research on natural rubber latex allergy has focused on clinical methodologies and emphasized the quantification of employee sensitization rates as opposed to actual incidents of reactivity. Workers' compensation data provide information on the number and impact of reactions to latex use. This article presents an analysis of health care workers' compensation data from North Dakota to estimate the prevalence, costs, and nature of claims associated with latex allergic reaction. The results show an annual average claim rate of 1.52 per 10,000 health care workers employed in the state, and annual costs averaging about $.08 per health care worker. Skin disorders were the most commonly reported condition. These findings are compared with previous studies of Minnesota and Rhode Island and demonstrate similar results. PMID- 11269841 TI - Relationship of workers' compensation status and duration of preoperative carpal tunnel symptoms. AB - Carpal tunnel syndrome (CTS) is one of the most common work-related injuries and a leading cause of work disability. We studied patients from a single community who had carpal tunnel releases (CTRs), comparing duration of preoperative symptoms and severity of nerve dysfunction among workers compensation and nonworkers compensation recipients. Included in the study were 131 patients who had a total of 187 primary CTRs done by a single surgeon. Duration of preoperative symptoms was significantly longer for nonworkers compensation patients than for workers compensation patients (p < 0.01). A smaller proportion of workers compensation patients had severe electromyography (EMG) findings (p = 0.04), and a larger portion had borderline EMG findings. People with work-related CTS appear to receive surgical treatment for CTS earlier than people whose CTS is not related to work covered under workers compensation laws. PMID- 11269842 TI - Continuing medical education. What do Minnesota physicians want? AB - CONTEXT: Continuing medical education (CME) is necessary for ongoing licensure and is critical in maintaining professional expertise. However, educators may not always consider their students preferences when developing new programs. OBJECTIVE: To determine physician preference for the format of CME programs and to learn what factors contribute to selecting a CME activity. DESIGN: Survey with 12 multiple response items pertaining to educational objectives, past educational experiences, and demographic information. PARTICIPANTS: A total of 1,967 Minnesota physicians were sent the survey; 385 physicians returned surveys within 2 months of mailing date (20% return rate). RESULTS: The vast majority of respondents reported participating in traditional CME programs during the preceding two years, and most said they planned to attend a traditional program in the next two years. CONCLUSIONS: Minnesota physicians overwhelmingly prefer attending traditional CME programs to participating in more interactive, technology-based activities. Before new technology such as the Internet can be widely used in CME, it must be made attractive to the practicing physician. PMID- 11269843 TI - Keep the sample closet door open. PMID- 11269844 TI - A life of leadership. PMID- 11269845 TI - [System for recording heart intervals in man]. AB - The equipment for observation and recording of human cardiointervals which is based on arterial pulse wave registration is proposed. PMID- 11269847 TI - [Influence of cooling and warming of the arm on the reciprocal 1a inhibition of flexor muscles of the forearm of man]. AB - In electrophysiological investigation on the healthy people the cooling (24 degrees C) and warming (42 degrees C) of arm was studied for their effects on the reciprocal 1a inhibition of flexor muscle of forearm during condition stimulation of extensor. Was observed, that cool and warm temperature stimulation enhanced this inhibition in flexor muscle. The cooling was more effectively in this enhancing. The obtained data can be connected with segmental and supraspinal mechanisms in the control of movements under sensor flow. PMID- 11269846 TI - [Role of estrogens in the regulation of puberty in female rats]. AB - Various correlations of testosterone metabolites during puberty were modulated in female rats. It has been fixed that estrogenes stimulate female pubescence only under conditions of non-aromatized androgenes availability in organism. Role of estrogenes in first-stage puberty (vagina opening) consists in activation of fermentative (5a-reductive) system which realises testosterone biotransformation into 5 alpha-reduced androgenes. Being dependent on the dose estrogenes shorten the period between initial and final (establishing positive feed-back way in ovaries-hypophysis system) puberty stages, hasten the first ovulation. PMID- 11269848 TI - [The relationship between psychophysiological and some anthropometric indexes in men and women]. AB - The relationships between functional mobility of nervous processes (FMNP), brain efficiency, index of successive work, nervous system strength of human and his some anthropometric indexes: mass, height, index mass body and proper basal metabolism were investigated. We showed that men in comparison with women have reliable higher levels of FMNP and also reliable higher levels of mass, height, proper basal metabolism and both in the whole group and in the subgroups with different level of FMNP. It was shown that mass of women is nonlinearly depended on functional mobility of nervous processes. Besides, such psychophysiologycal properties as brain efficiency and index of successive work correlated with mass, index mass body and proper basal metabolism. PMID- 11269849 TI - [Influence of eosin y and orthovanadate on the steady-state of Ca(+)2 entry into secretory cells of exocrine glands and their spontaneous secretion]. AB - Ca(2+)-pump blocators in low concentrations (eozyn Y up to 5 mM and ortovanadate up to 40 mM) essentially increases of Ca2+ content in salivary gland of Chironomus plumosus larvae's and spontaneous protein secretion. It was shown that eozyn Y much more effectively suppresses of plasma membrane Ca(2+)-pump then ortovanadate. Eozyn Y and ortovanadate in higher concentrations essentially decrease of Ca2+ content in glands and spontaneous protein secretion. The former is evoked by suppression of endoplasm reticulum Ca(2+)-pump, decreasing of Ca2+ influx in cells following by diminishing of Ca2+ transmembrane gradient. Therefore, energydependent Ca2+ transporting systems of plasma membrane and endoplasm reticulum effectively regulate steady-state Ca2+ entry in secretory cells of Chironomus plumosus salivary glands and maintain relatively low level of spontaneous secretion. PMID- 11269850 TI - [Physical endurance in rats exposed prenatally to stress of exogenous glucocorticoids]. AB - Gender- and age-related peculiarities of the influence of prenatal stress or exogenous glucocorticoids on physical endurance in rats were found in the experiments using fatigue test. Normal adult females were of greater endurance than males. Prenatal stress resulted in increased physical fatigue mainly in females while sexual differences were reversed. Hydrocortisone acetate administration to pregnant rats prevented sexual dimorphism in physical fatigue development in adult animals due to increase of physical endurance in males and its decrease in females. In aging rats disappearance of both sexual differences and prenatal stress effects took place against a background of fast physical fatigue development. PMID- 11269851 TI - [Cerebrocrast correction of reparative osteogenesis on the background of chronic stress]. AB - Protective action of cerebrokrast--a derivative from 1,4-dihydropyridine--on the processes of reparative osteogenesis in rats in case of combined action of chronic emotional-pain stress and trauma of mandibular was determined. It is stipulated by a antioxidant properties of cerebrokrast, as well as its stabilizing influence on glycoproteins and calcium homeostasis. The redeived data extend the idea about general-metabolic action of the investigated nootrop at the level of the whole organism and testify worthwhile of its use for prophylaxis of extreme conditions. PMID- 11269852 TI - [Regulatory mechanisms of erythropoiesis by leukocytes in inflammation]. AB - On the model of carrageenan-induced acute aseptic peritonitis in rats with use of the inhibitors of some leukocytic products it is shown that lysosomal proteinases inhibit erythropoiesis activation in inflammation. Lipoxygenase metabolites stimulate erythropoiesis. Eicosanoids as a whole stimulate erytropoiesis still more. Reactive oxygen species promote the development of erythron hyperplasia. Dexamethasone administration increases the erythropoiesis intensification. PMID- 11269853 TI - [State of lipid peroxidation and antioxidant activity of blood in skin wounds induced by mechanical and radiation injury]. AB - The state of lipid peroxidation and antioxidant system in blood were been studied on the models of skin wound and radioinduced ulcer in rats. It was shown that the healing of skin wound by secondary strength is accompanied by activation of processes lipid peroxidation in plasma and erythrocytes during 15 days. The development of radioinduced ulcer is characterized by more high degree activation of lipid peroxidation beginning with latent period and till forming ulcer (30 days). Only in process of radioinduced ulcer development two phase of activation lipid peroxidation and decrease of antioxidant enzyme activity--catalase and superoxid dismutase--were established. Secondary phase of lipid peroxidation activation was coincided with period of clinical appearances and it was possibly explained by exhaustion of antioxidant system. PMID- 11269854 TI - [Central hemodynamics in different types of weather under orthostatic and clinical conditions]. AB - The results of research of weather influence on the state of the central haemodynamics in primary condition (after 10 minutes adaptation lying on the back) and on 1, 5, 10 minutes of ortostatic and clinostatic tests are presented in the work. It's established that reactivity of the cardiovascular system on ortostasis is decreased in unfavourable weather conditions. It was observed greater decreasing of striking and heart induces compare comparatively with ones in favorable weather. These facts prove the state of disadaptation in unfavourable weather conditions. PMID- 11269855 TI - [Production of superoxide anion radical and nitric oxide in renal tissues sutured with different surgical suture material]. AB - The generation of superoxide anion radicals (in mitochondria, microsomes and under respiratory burst of leucocytes) and nitric oxide (NO) in renal tissue has been studied in the experiment with white rats, which had been carried out nephrotomy with following usage for suture such absorbable surgical threads as plain and chromic catgut, biofil (of dura mater spinalis of the cattle), Dexon II (polyglycolic acid) and biofil modified with aethonium, succinate and mexidol. The research proves the use of plai and chromic catgut leads to the development longer oxidative stress with increasing of cytotoxic agents production (superoxide anion and NO). The risk of longitudinal oxidative stress decreases under the use of biofil suture modified with biological active compounds (aethonium, succinate and mexidol). In this case, the generation of superoxide anion radicals in mitochondria and microsomes is normalised earlier. The superoxide generation with respiratory burst of leucocytes and NO production decreases in 14 day of postoperative period under the use of biofil suture modified with succinate and mexidol. PMID- 11269856 TI - [Effect of sodium alpha-ketoglutarate injected after x-ray treatment on the respiration and oxidative phosphorylation of hepatic mitochondria]. AB - It have been found that total X-ray treatment (everyday expose to 1 Roentgen up to achievement of total doses 10, 20 30 Roentgen) inhibits the rate of ADP stimulated respiration of rat liver mitochondria, decreases its efficiency and makes phosphorylation less couple to respiration. All this effects are present from the first period after treatment till the end of treatment. Intraperitoneal alpha-ketoglutarate injections during treatment decreases the inhibition of respiration and oxidative phosphorylation, increases the efficiency of respiration during the period from 1-st injected till the end of experiment. PMID- 11269857 TI - [Effect of intraventricular infusion of Licvorin on general convulsive activity of rats]. AB - It was studiet anticonvulsive action of the biopreparation licvorin had been made from a cerebrospinal fluid (CSF) of a cattle on the epiliptic activity provoking by korasol. The preparation was infused intraventriculary. Research showed preparation had the antiepileptic effect. It was manifested by decreasing expressiveness of convulsions, averting of general epileptic fit and mortality. Anticonvulsive action of the biopreparation licvorin was registrated in 2 hours and was achieved maximum effect 24 hours after injection. Anticonvulsive properties of the biopreparation licvorin realized by influence his active factors on structures of a brain, specifically on the GABAdependent systems. PMID- 11269858 TI - [New ways of correcting metabolic acidosis in experimental periodontitis]. AB - Established prevention of atrophy of osseous cloth alveolar rats jaws sprouts and correction of metabolic acidosis of mineral concentrate "VITA" attached to modeling parodontit by dint of creation in them of fortune compensated metabolic acidosis. Mineral concentrate realizes normalization redoksfortune, thiol disulphid system, lowering of relations SH/SS-group, multiplies maintenance oxidized and lowers maintenance restored of nicotinamid copherments, normalize relation HA[symbol: see text]/HA[symbol: see text]H in osseous jaws cloth of experimental rats attached to modeling in them metabolic acidosis and parodontit in distinction from fortune of alkalosis. Mineral complex will normalize in rats maintenance of regulators and orientation of exchange processes in cloths attached to parodontit, whereat indicates lowering of activity key gluconeogenesis processes enzyme fructosodiphosphatasa of liver cloths, mucous gum and alveolar sprouts of jaws. PMID- 11269859 TI - [Role of apoptosis in the pathogenesis of atherosclerosis]. AB - The recent information about importance of programmed cell death in the development of atherosclerosis was reviewed. It was emphasized, that intensiveness of apoptosis was various for smooth muscle cells, macrophages, endothelial cells, lymphocytes, and changed during the development of such a multistage process as atherosclerosis. Is was proved, that exactly apoptosis gave specific features to the atherosclerotic process. The agents inducing atherosclerosis (modified lipoprotein, mechanical influences, viruses, bacterial toxins, etc.), have insufficient for initiation of necrosis. The damage to vascular wall in atherosclerosis is characterized not by intensive influence of pathogenic factors, but rather constant chronic influence of the agents of moderate power. Therefore, it is just apoptosis that makes up the critical mechanism of the reactive-regeneration answer of vascular wall structures to an injure. Evolutionally, the apoptic way of the development of answer to an injure appears to be more rational, as for as it results in minimal losses of cells due to both secondary alteration and maintenance of fast restoration of the primary cellular architecture of injured tissues at the expense of the natural mechanisms of regulation of apoptosis and proliferation. In the certain situation, these positive aspects of apoptosis make up a basis for the development of a pathology- failures in the program of apoptosis result in chronic proliferative-degenerative processes, that is in atherosclerosis. PMID- 11269861 TI - [Cardiac reflexes under conditions of heart injury]. AB - Analysis of the previous experimental literary data and own authors data about the pattern of cardiogenic reflectory effects during rise and development of the local injury is represented in this review. The role of double afferent innervation of the heart is discussed and author suggests its mayor functional role under some kind of heart diseases. PMID- 11269860 TI - [Role of opiate system in mechanisms of formation of alcohol-dependence]. AB - Effect of naloxon on the activity of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) in the blood serum and content of catecholamines (CA) in the brain structures (hypothalamus, midbrain, new cortex) and blood of rats under alcoholic dependence were studied. Participation of opiate structures of receptors in formation of alcoholic dependence is shown. PMID- 11269862 TI - A review of interproximal wear grooves on fossil hominin teeth with new evidence from Olduvai Gorge. AB - Interproximal (approximal) grooves at the cementum-enamel junction of premolar and molar teeth have been observed in a broad range of human ancestors and related extinct species from 1.84 million years ago to the present. Many hypotheses have been presented to explain the aetiology of these grooves, though their form and positioning are most consistent with tooth-picking behaviours. This paper reviews occurrences of interproximal grooves in the cheek teeth of modern and fossil humans, evaluates hypotheses on their cause, and reports on a previously undescribed groove found in OH 60, a molar tooth from Olduvai Gorge. This specimen is among the earliest to show such grooving, and is most likely attributable to Homo erectus. It is concluded that, because interproximal grooves have been observed only on Homo teeth, they probably reflect a behaviour or behaviours unique to that genus. PMID- 11269863 TI - Compositional analysis of human acquired enamel pellicle by mass spectrometry. AB - Relatively little is known about the formation of the acquired enamel pellicle other than that it involves the selective adsorption of specific proteins from oral fluids. Previous studies on the identification of pellicle components have relied largely on immunological or enzymatic detection and have been hampered by the fact that only minute quantities of pellicle can be removed from tooth surfaces. The present work describes an improved method of harvesting pellicle that combines mechanical and chemical removal; this approach was used to investigate systematically the desorption of in vitro pellicle components with different solutions. Eleven major in vitro pellicle proteins were identified by using a combination of electrophoretic separation and matrix-assisted laser desorption/ionization-reflectron time-of-flight mass spectrometry. A similar analysis of in vivo-formed pellicle revealed the presence of intact statherin, lysozyme, albumin and amylase. Further analysis of in vivo pellicle by liquid chromatography-electrospray ionization mass spectrometry suggested the presence of numerous low molecular-weight fragments of precursor proteins. The protein composition of in vitro whole-salivary pellicle adsorbed to hydroxyapatite and that of in vivo enamel pellicle differed for proline, the result of a reduction in the content of acidic proline-rich proteins in the in vivo samples. Unique features of the oral environment such as enzymatic activities or mineral surface properties may account for these differences between in vivo and in vitro pellicle formation. PMID- 11269864 TI - Ultrastructure of early mineral deposition during hyaline layer formation in rat molars. AB - There is no consensus on whether the first mineralized layer, the hyaline layer, that is juxtaposed to root dentine is a variety of dentine or cementum or even a tissue of epithelial origin. Some suggest that there is no intermediate tissue between the acellular extrinsic fibre cementum (AEFC) and the root dentine. Here, to study hyaline layer formation and mineralization we examined by transmission electron microscopy the early stages of root development in upper molars from 10 to 13 day old Wistar rats. In addition to conventionally processed material, undemineralized and unstained sections were examined, which showed the deposition of fine mineral crystals in contact with the mineralized surface of root dentine. Early mineralization of the hyaline layer occurred in the region of the inner basement membrane, which persisted between the inner cellular layer of Hertwig's epithelial root sheath and the outer mineralized root dentine. When the root sheath began its fragment, collagen fibrils from the developing periodontal ligament began to insert into the mineralising hyaline layer, which was 0.5-0.8 micron wide. As the fragmentation of the root sheath HERS increased, more collagen fibrils appeared intermingled with the mineralising hyaline layer. In more advanced stages, when the hyaline layer had become fully mineralized and the formation of the AEFC began, the hyaline layer could no longer be identified. Thus, the hyaline layer is clearly discernible at early stages of periodontal development. Subsequently, it is masked by intermingling of cementum and dentine and therefore it is not possible to detect it in the formed roots of rat molars. PMID- 11269865 TI - Sialogogic activity in the rat of peptides analogous to [Tyr8]-substance P in which substitutions have been made in the N-terminal amino acids. AB - In order to elucidate the regulatory roles for salivation of amino acids in positions 1-4 of the N-terminal region of [Tyr8]-substance P (SP), the structure sialogogic activity correlations of various synthetic octa- to undecapeptides replaced in positions 1-4 of [Tyr8]-SP with each of 19 common amino acids, one by one, and with the same sequence of the C-terminal hepatapeptide as that of [Tyr8] SP, were studied in the submandibular glands of rats after intraperitoneal injection. Each of 19 octa-, nona-, deca- and undecapeptides with replaced amino acids and a penta- to decapeptide with the progressive elimination of the N terminal portion were newly synthesized by the multipin peptide method. All octa- to undecapeptides replaced with each of 19 common amino acids in positions 1-4 had sialogogic activities. In 19 octa- and decapeptides in which P4 and P2 had been replaced, four and three replacements, respectively, had significantly increased secretory activities. In contrast, in 19 nonapeptides in which K3 had been replaced, none had significantly increased secretory activities. Furthermore, in 19 undecapeptides in which R1 had been replaced, most replacements had significantly increased or equipotent activities for fluid secretion. It is concluded that amino acids in the N-terminal region of various tachykinins may not need to be strictly conserved and that amino acid residues in the N-terminal portion, R1 in particular and P2, may strongly inhibit secretory activity. PMID- 11269866 TI - Oral colonization and cariogenicity of Streptococcus gordonii in specific pathogen-free TAN:SPFOM(OM)BR rats consuming starch or sucrose diets. AB - The significance of Streptococcus gordonii in dental caries is undefined, as is that of other alpha-amylase-binding bacteria (ABB) commonly found in the mouth. To clarify the ecological and cariological roles of S. gordonii our specific pathogen-free Osborne-Mendel rats, TAN:SPFOM(OM)BR, were fed either diet 2000 (containing 56% confectioner's sugar, most of which is sucrose) or diet 2000CS (containing 56% cornstarch, in lieu of confectioner's sugar) and inoculated with S. gordonii strains. Uninoculated rats were free of both indigenous mutans streptococci (MS) and ABB, including S. gordonii, as shown by culture on mitis salivarius and blood agars of swabs and sonicates of dentitions after weanlings had consumed these diets for 26 days. ABB were detected by radiochemical assay using [125I]-amylase reactive to alpha-amylase-binding protein characteristic of the surface of S. gordonii and other ABB. No ABB were detected (detection limit < 1 colony-forming units in 10(6) colony-forming units). Thus the TAN:SPFOM(OM)BR colony presents a 'clean animal model' for subsequent study. Consequently, S. gordonii strains Challis or G9B were used to inoculate weanling rat groups consuming either the high-sucrose diet 2000 or the cornstarch diet 2000CS. Two additional groups fed each of these diets remained unioculated. Recoveries of inoculants were tested 12 and 26 days later by oral swabs and sonication of the molars of one hemimandible of each animal, respectively. Uninoculated animals were reconfirmed to be free of ABB and mutans streptococci, but inoculated ones eating diet 2000CS had S. gordonii recoveries of 1-10% or, if eating diet 2000, 10-30% of total colony-farming units in sonicates. There were no statistically significant differences among the inoculated and uninoculated animal groups' caries scores when they ate the cornstarch diet. Lesion scores for sucrose-eating rats were, however, from 2.4-5.1-fold higher than for cornstarch-eating rats, P < 0.001, and were still higher if animals had been inoculated with either Challis (1.41-fold) or G9B (1.64-fold), than if uninoculated, both P < 0.001, so long as the rats ate the sucrose diet. Therefore, TAN:SPFOM(OM)BR rats do not harbour ABB or S. gordonii but can be colonized by S. gordonii. Colonization levels of S. gordonii on the teeth are higher in the presence of high sucrose than with high starch-containing diets. Caries scores are augmented by sucrose compared with starch, and are further augmented by S gordonii colonization. S. gordonii is thus cariologically significant in the presence of sucrose, at least in this rat. PMID- 11269867 TI - Identification and isolation of differentially expressed genes in osmotically stressed human oral keratinocytes. AB - Complementary DNA fragments which showed differential expression relative to unstressed controls were identified and isolated from human oral keratinocytes exposed to hyperosmotic stress. The up- or downregulation of the expression of nine of these cDNAs in response to osmotic stress was determined by Northern blotting. Sequence analysis showed that clones K-5 and K-46 contained identical sequences. Homology searches revealed that K-13 and K-33 were fragments of unknown genes. Among the upregulated cDNAs, K-16 and K-32 were 94 and 83% identical to chromosome 16 bacterial artificial chromosome (CIT987K-A-418G10) and a cDNA (ai49b01.sl) clone, respectively. Another clone, K-34, encoded a protein 73% identical to Bax epsilon. Among the downregulated genes, K-5/46 and K-45 were 99% identical to the og24d08.s1 cDNA clone and to mitochondrial genes for tRNAs and 12S and 16S ribosomal RNAs, respectively, while K-50 was 100% identical to KIAA0905 protein. The gene expression induced by osmotic stress occurred in parallel with the induction of apoptosis and a reduction in protein biosynthesis. This observation, together with the characteristics of the some of the differentially expressed genes, suggests that among the major events induced in oral keratinocytes by hyperosmotic stress are the induction of apoptosis and a decrease in protein biosynthesis, brought about by upregulation of pro-apoptotic genes and downregulation of genes involved in protein biosynthesis. PMID- 11269868 TI - Fluoride alters casein kinase II and alkaline phosphatase activity in vitro with potential implications for dentine mineralization. AB - Dentine phosphoprotein (DPP), a major non-collagenous acidic protein of dentine, undergoes altered phosphorylation in vivo in the presence of high fluoride concentrations. This has major implications for the altered mineralization patterns found during fluorosis. In dentine, casein kinase II is involved in phosphorylating DPP, and alkaline phosphatase (ALP) is ascribed roles in the dephosphorylation of DPP, increasing the inorganic phosphate at the mineralization front and the removal of pyrophosphate. Here the influence of fluoride in vitro on the activity of purified casein kinase II and ALP and its relation to altered patterns of mineralization were examined. Kinetic analysis showed that casein kinase II activity was completely inhibited at 0.04 M NaF. Vmax when compared to the control assay was significantly decreased (P < 0.0001) between concentrations 4 x 10(-4)-4 x 10(-8) M NaF. Significant changes to the Km (P < 0.0001) were also observed. ALP activity was inhibited by NaF (0.09-9 x 10( 8) M), with Vmax significantly decreased (P < 0.0001) at 0.09 M NaF. Alterations in the activity of these enzymes in the presence of fluoride may in part explain the decreased phosphorylation observed in DPP isolated from fluorotic dentine and may aid understanding of the altered matrix mediated mineralization patterns found during fluorosis. PMID- 11269869 TI - The effect of food consistency and dehydration on reflex parotid and submandibular salivary secretion in conscious rats. AB - Changes in salivary secretion with different consistency of diet and dehydration were studied in male Wistar rats under unrestricted conditions. To measure the salivary secretion, a stop-flow method was used. There was little unstimulated salivary secretion from the parotid and submandibular glands, but eating solid, powdered, and liquid diets induced parotid and submandibular saliva. There was no significant change in the volume and flow rate of saliva in bilateral parotid glands during the eating of solid diets. The solid and powdered diets induced significantly more salivary secretion from the parotid gland than did the liquid. The salivary flow rate with solid diets was significantly greater from the parotid gland than from the submandibular. On the other hand, the salivary flow rate with the liquid diet was significantly smaller from the parotid gland than from the submandibular. Appreciable amounts of submandibular saliva, but little parotid saliva were secreted during grooming. Clearly, parotid and submandibular saliva have different roles in the rat. When injected intraperitoneally with 1.5 M NaCl solution or water-deprived for 24 h, rats took longer to eat the solid diets, and had increased salivary volume and decreased flow rate from the parotid gland. These results indicate that the moisture content of the diet and the dryness of the mouth alters the volume of parotid saliva secreted in rats and show that parotid saliva plays an important part in mastication and swallowing. PMID- 11269870 TI - Reduction in the rate of inositol 1,4,5-trisphosphate synthesis in rat parotid acini by lithium. AB - Stimulation of muscarinic cholinergic receptors on rat parotid acinar cells causes a rapid production of inositol phosphates, with the key metabolic event being the breakdown of phosphatidylinositol 4,5-bisphosphate into inositol 1,4,5 trisphosphate (Ins(1,4,5)P3) and diacylglycerol. Here a high-performance liquid chromatographic technique was used to measure the effects of intracellular lithium ions on the amount of various inositol phosphates produced. When acini were stimulated maximally with acetylcholine (ACh), the sum of all inositol phosphates produced followed a monoexponential function with a production rate constant for Ins(1,4,5)P3 of 0.07 +/- 0.01 solidus/sec. The presence of 23 mM LiCl intracellularly reduced the production rate constant of Ins(1,4,5)P3 induced by ACh to 0.03 +/- 0.01 solidus/sec, resulting in a decrease in the Ins(1,4,5)P3 production as well as in the magnitude of the rise in the intracellular free Ca2+ concentration. The lithium ion (Li+) did not affect the rate of conversion of Ins(1,4,5)P3 to either inositol 1,4-bisphosphate or inositol 1,3,4,5 tetrakisphosphate. The rate of the inositol phosphate production after the addition of the Ca2+ ionophore ionomycin was unaffected by intracellular Li+ (23 mM), which implies that the action of Li+ was at the muscarinic cholinergic receptor, on G-protein or on the interactions between G-proteins and phospholipase C. Thus, in the early events after receptor stimulation with ACh, Li+ causes a reduction in the concentration of the cellular messengers Ins(1,4,5)P3 and Ca2+. PMID- 11269871 TI - A histometric study in rats of the effect of the calcium antagonist amlodipine on bone healing after tooth extraction. AB - The purpose was to investigate whether amlodipine, a second-generation calcium antagonist used for the treatment of hypertension and angina, interferes with healing of rat alveolar bone. A progressive increase in volume density of new bone filling the socket was quantified by a histometric differential point counting method 7-42 days after tooth extraction. The results showed a 20-30% decrease in bone volume fraction in the alveolus of amlodipine-treated animals from 7 days on, in addition to a higher (7-35%) volume fraction of connective tissue and a tendency toward an increase in the volume fraction of persisting coagulum. If confirmed in humans, the knowledge of a deleterious effect of Ca channel blockers in hindering alveolar bone healing would be important in planning oral operations involving bone tissue, including those for device implantation. PMID- 11269872 TI - Motor potentials evoked by transcranial magnetic stimulation during isometric and dynamic masseter muscle contraction in humans. AB - The facilitation of muscle motor potentials evoked by transcranial magnetic stimulation (TMS) has been demonstrated convincingly during both isometric and dynamic activity in the limbs but not in the jaw muscles. An experimental design involving TMS, surface electromyography and controlled muscle-activity was employed to investigate the motor response of the human masseter during voluntary isometric and dynamic voluntary conditions. During the isometric condition, an increase in muscle facilitation resulted in a progressive increase in motor evoked potential (MEP) amplitude that was consistently greater on the side contralateral to that subjected to TMS (P < 0.05). No difference in MEP amplitude or laterality of response was revealed for the two dynamic conditions. The sample size may have been too small to reveal any differences. The modulation of MEPs during isometric activity was probably due to cortical and brainstem mechanisms. Putative variation in masseter MEPs during dynamic conditions cannot be discounted. PMID- 11269873 TI - Stress response to surgical procedures in the submandibular region and its influence on salivary secretion in mice. AB - Saliva stimulated by pilocarpine was collected from the mouths of adult ICR male mice after either submandibular-sublingual gland (SM-SL) exposure and SM-SL removal under pentobarbital anaesthesia. Salivary flow rates decreased and salivary protein concentrations increased after both surgical procedures. Serum cortisol was elevated after the operations. Salivary protein separation patterns on sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) from surgically treated animals were very similar to those from mice given an alpha 1 adrenergic agonist (phenylephrine), but differed significantly from those given a beta-adrenergic agonist (isoproterenol). When the SM-SL was completely removed (i.e. parotid saliva), protein concentrations also dramatically increased. The SDS-PAGE protein separation pattern in the mice whose SM-SL had been removed was very similar to that in those whose SM-SL had been exposed. Hence, a surgical insult in the submandibular region in mice enhances the secretion of salivary protein, mainly in response to alpha 1-adrenergic receptor stimulation in the parotid gland. This study suggests the usefulness of alpha 1-adrenergic receptor stimulated parotid salivary protein as a marker of surgical insult. PMID- 11269874 TI - [Intracranial complications of otitis]. AB - Since the advent of antibiotics, endocranial complications due to infectious ear disease are actually unfrequent. They occur predominantly in young adults with a long term chronic otitis media and cholesteatoma. The mortality for endocraneal complications still remains significant. We report six cases of endocraneal complications due to infectious ear disease, that have been seen in our service over the last eight years; remarking that half of the studied patients had an acute ear disease. A review of the literature helps to understand the actual standard of diagnosis and treatment. PMID- 11269875 TI - [Molecular changes in epidermoid carcinoma of the oropharynx]. AB - In most of the studies about molecular alterations in squamous cell carcinomas of the head and neck there is not distinction between the different subsites of this area. The objective of this study is to describe the molecular alterations in squamous cell carcinomas of the oropharynx. Twenty-nine oropharyngeal carcinomas, with a minimum follow-up of 36 months, were studied. The molecular alterations analyzed were: the amplification of 11q13 region (in the 29 cases), and the MYC and ERBB1 oncogenes (in 22 cases); the integration of Human Papillomavirus (HPV) types 6b and 16 (in 22 cases); the loss of heterozygosity (LOH) of p53 and N acetyltransferase-2 (NAT2) gene (in 12 and 13 informative cases, respectively); and the cellular DNA content (in 13 cases). The most frequent alterations found were the LOH at p53 (67%), and NAT2 (54%) locus, followed by 11q13 amplification (49%). ERBB1 amplification was found in 14% of the cases, and MYC amplification only in one (5%). Integration of the HPV was found in 23% of the cases. Nine (69%) of the 13 analyzed cases were aneuploid. The only alteration with a prognostic significance was 11q13 amplification that showed a tendency to be associated with a higher frequency of nodal metastases and tumor recurrence. PMID- 11269876 TI - [Validity of hand-held systems for otoacoustic emissions in hearing screening]. AB - Hearing screening objective is the development of an universal screening. With that purpose, new hand-held system of Otoaccoustic Emissions have been designed, which are easier to perform and to interpret the results, to detect hearing loss disordersas soon as possible. To validate the hand-held systems of Otoaccoustic Emissions in the hearing screening, we have compared the results of the tests with Otoaccoustic Emissions clinic systems and tonal audiometry in two groups of population: new-borns and hearing-loss adults aged between 16 and 56. Our study showed that the hearing screening results in children with hand-held system were similar to classic Otoaccoustic Emissions system, but there were more than 40% of adult hearing losses not detected with this system. These results can invalidate the test. PMID- 11269877 TI - [Cancer of the tonsillar region. Retrospective study and review of the literature]. AB - It has been carried out a retrospective analysis of 34 patients suffering from carcinoma epidermoid of the tonsillar fossa being treated with two different therapies from 1989 to 1996. Twenty three of them were treated with surgery (transoral resection or bloc radical tonsillectomy with myocutaneous flap reconstruction and ipsilateral lymph node dissection). Eleven of them were treated with chemotherapy and radiotherapy. The rate of overall survival at 5 years were 41.2%. Actuarial survival rates were (71.4%) in stages I-II and 33.3% in stages III-IV (p = 0.08). On multivariate analysis, age (p = 0.28) and modality of treatment (p = 0.80) were not significant effect on survival. Advanced stages (III-IV) showed 3.4 times much more risk of death than early stages (I-II) (p = 0.11). PMID- 11269878 TI - [Reconstruction of oropharyngeal defects with lateral arm flap]. AB - The lateral arm flap, is a fasciocutaneous flap with great versatility, but underused in head and neck reconstruction. Its qualities include a intermediate thickness between the radial forearm flap and the pectoralis major, ideal to reconstruct oropharyngeal defect, a consistent vascular pedicle, a pliable soft tissue and a low donor site morbidity. Use of this flap does not require the sacrificing of a major feeding vessel to the arm. We have chosen this technique to reconstruct four cases with surgical defects in oral cavity and oropharynx. The anatomic and functional results have been satisfactory and the complications rate is comparable to other microvascular techniques. We think that the lateral arm free flap is a useful reconstructive technique in specific areas of head and neck. PMID- 11269880 TI - [Laryngeal tuberculosis: study of 11 cases]. AB - We report 11 patients with laryngeal tuberculosis seen in our hospital, January 1990 to July 2000. Eight were men and all cases presented with dysphonia and/or disphagia. In 8 pulmonary tuberculosis was associated. Mycobacterium tuberculosis was isolated from the sputum in 7 patients. Granulomatous laryngitis was demonstrated in the eight patients with laryngeal biopsy. The evolution with medical treatment was favourable in all patients. PMID- 11269879 TI - [Preoperative evaluation in thyroplasty: the laryngeal lateral compression]. AB - Thyroplasty type I has provided significant improvement to the treatment of patients with glottal incompetence. It is essential that patients be preoperatively evaluated using objective criteria. Laryngeal manual compression test are manipulations of the thyroid and cricoid cartilages that result in modifications of the position of the vocal folds. The most valuable laryngeal manual compression test for patients with glottal insufficiency is the lateral compression test. When this test results in a preoperative improvement in voice suggest that surgery will be successful. In this paper we present the objective evaluation of the effects of lateral compression test upon glottic incompetence by means of narrow band power spectrum analysis. PMID- 11269881 TI - [Usefulness of RET proto-oncogene in the diagnosis of hereditary-type medullary carcinoma of the thyroid. Correlation with surgical findings]. AB - In about 25% of cases medullary thyroid carcinoma (MTC) is hereditary. In this group is possible to detect germline point mutations of the RET proto-oncogene in about 95% of the studied cases. The purpose of the present paper is to confirm the value of the RET in the screening of the hereditary MTC. We studied 43 subjects at risk for a Multiple Endocrine Neoplasia Type 2A Syndrome. RET analysis was positive for MEN 2A in 22 subjects. Fifteen of the 22 have undergone a total thyroidectomy at our facility. Histopathological study of the surgical specimens confirmed the presence of a MTC in all the cases. According with our experience. RET analysis is a 100% sensitive and specific method of hereditary MTC screening, and we think it has obvious advantages over the calcitonin tests in technical, economic and ethic aspects. PMID- 11269882 TI - [Giant ethmoid neurinoma with intracranial extension]. AB - The schwannomas are tumors arising from nervous tissue. It appears very rarely in the nose and paranasal sinuses. Intra and extracraneal extension of these tumors are even more uncommon. In this paper we report a case of a esfeno-ethmoidal schwannoma with anterior skull fossa extension. We describe the CT scan and magnetic resonance imaging and the histological features. Surgical resection was carried through a craniofacial approach. Some data about diagnosis, treatment and outcome of these tumors are revised. PMID- 11269883 TI - [Leiomyoblastoma of the tongue]. AB - Leiomyoma is a benign tumor of smooth muscle origin that is rarely found in oral cavity. We report a case of leiomyoblastoma of the tongue. The interest of this lesion is supported by its location and by the difficulty to determine its biological behavior. About 25% of the oral smooth muscle tumors are malignant, and have been reported "leiomyoma" locally invasive. PMID- 11269884 TI - [Papillary carcinoma on thyroglossal duct cyst]. AB - Papillary carcinoma ansing on a thryroglosal duct cyst is a rare tumor. Since Ucherman described first case in 1915, only 150 cases have been reported. We present clinic evolution and treatment of a new case and review literature and discussion about tumor origin, adequate diagnosis test and treatment. This should include Sistrunk procedure but further surgery on thyroid gland is not accepted by all authors. PMID- 11269885 TI - [Laryngeal metastasis from colloid adenocarcinoma of the colon: report of a case]. AB - Metastatic involvement of the laryngeal is very rare, with around 150 cases reported to the literature. In eight of these cases, the primary tumor was a colon adenocarcinoma. We report the case of a 80 year-old woman treated of a colloid adenocarcinoma of 7 years earlier, referred to us for chronic and progressive dyspnea. Endoscopic examination showed a subglottic spherical mass, which caused an important compromise of the respiratory airway. Tomographic studies revealed also a thyroid mass. The patient was treated with a tracheostomy, resection of the subglottic mass (with intraoperative diagnosis of "mucin producing tumor"), and total thyroidectomy. The final pathologic diagnosis of the subglottic mass was a metastasis of colloid adenocarcinoma of the colon. In the literature reviewed there are no previous reports of metastatic involvement of the larynx with this type of colon adenocarcinoma. We discuss the clinical and radiological findings, and therapeutic options for metastasis to the larynx, as well as pathological differential diagnosis. PMID- 11269886 TI - [Results of tympanoplasty with antrum exclusion in the treatment of middle ear cholesteatoma]. AB - INTRODUCTION: Exclusion of the antrum was designed to avoid the problem of mastoid ventilation by means of insufficient eustachian tube. MATERIAL AND METHODS: We retrospectively reviewed 72 ears with cholesteatoma who underwent antrum exclusion tympanoplasty (57% in men, mean age 40.9 years). Mean follow-up was 31.2 months (range, 12-64 mo). RESULTS: Exclusively epitympanic-antral cholesteatoma location (28.4%), and the necrosis of the long branch of the incus (54%) were the most common. The most used material to reconstruct the ossicular chain was Torp prosthesis (73.6%), followed by incus autograft (11.1%). We reported the following complications: postoperatory otorrhea (5.5%), perforation of neotympanum (16.7%), prosthesis extrusion (23.6%, time average of 15.6 months), and recurrence of cholesteatoma (4.2%). DISCUSSION/CONCLUSION: The technique with antrum exclusion has reported acceptable results in the control cholesteatoma recidivism. Although the functional results are similar to those of the open techniques, the resulting cavity with the antroexclusion is more anatomic. PMID- 11269888 TI - The effects of dexamethasone on insulin release and biosynthesis are dependent on the dose and duration of treatment. AB - Complex results concerning the effect of glucocorticoids on insulin secretion have been reported. The aim of this study is to clarify the direct effects of glucocorticoids on pancreatic islets and to determine whether the effect of glucocorticoids on insulin biosynthesis or release is dependent on the dose and duration of treatment with glucocorticoid. Studies on insulin secretion and biosynthesis were performed with different concentrations (0, 1, 10, 100 nmol/l) and durations (1 and 6 h) of treatment with dexamethasone (dexa) in rat pancreatic islets. (1) One nmol/l dexa had no inhibitory effect on insulin secretion and biosynthesis. Ten and 100 nmol/l had an inhibitory effect on insulin secretion, which was mainly due to suppression of the first phase of insulin secretion. (2) Insulin content was significantly increased regardless of the concentration in 1-h treated islets. However, insulin content was markedly diminished with 100 nmol/l dexa in 6-h treated islets. (3) The preproinsulin mRNA expression of 6-h treated islets was suppressed in a dose-dependent manner. Our data revealed that, in the condition of short-term and low-dose glucocorticoid exposure, insulin secretion and biosynthesis are not affected. The secretory process of insulin seems to be the initial step of the inhibitory action of glucocorticoid. Both insulin release and biosynthesis are inhibited by chronic exposure to high dose dexamethasone. It can be concluded that glucocorticoid might be involved in the multisteps of insulin release and biosynthesis. PMID- 11269887 TI - Hypotriglyceridemic and hypocholesterolemic effects of anti-diabetic Momordica charantia (karela) fruit extract in streptozotocin-induced diabetic rats. AB - Momordica charantia (karela) is commonly used as an antidiabetic and antihyperglycemic agent in Asian, Oriental and Latin American countries. This study was undertaken to investigate the effects of long term feeding (10 weeks) of M. charantia fruit extract on blood plasma and tissue lipid profiles in normal and streptozotocin (STZ)-induced Type 1 diabetic rats. The results show that there was a significant (P < 0.05) increase in plasma non-esterified cholesterol, triglycerides and phospholipids in STZ-induced diabetic rats, accompanied by a decrease in high density lipoprotein (HDL)-cholesterol. A moderate increase in plasma (LPO) product, malonedialdehyde (MDA), and about two-fold increase in kidney LPO was also observed in STZ-induced diabetic rats. The treatment of diabetic rats with M. charantia fruit extract over a 10-week period returned these levels close to normal. In addition, karela juice also exhibited an inhibitory effect on membrane LPO under in vitro conditions. These results suggest that M. charantia fruit extract exhibits hypolipidemic as well as hypoglycemic effects in the STZ-induced diabetic rat. PMID- 11269889 TI - Peptide-specific cytotoxicity of T lymphocytes against glutamic acid decarboxylase and insulin in type 1 diabetes mellitus. AB - Cytotoxic T lymphocytes (CTL) against pancreatic beta-cells probably play a major role in the etiology of type 1 diabetes mellitus (DM). CTLs recognize a complex formed between MHC class I and antigenic peptides fragments derived from intracellular processing of proteins. However, the exogenous peptides, which show strong affinities to MHC class I, can be presented. In this study, we focused on the cytotoxic activity of peripheral lymphocytes in patients with type 1 DM against the peptides of glutamic acid decarboxylase (GAD) and insulin, which can bind MHC class 1 A24. Lymphocytes were isolated from peripheral blood of 12 type 1 DM patients and eight healthy control subjects. The effector cells were cultured with peptides, IL-2 and IL-7, restimulated weekly by autologous antigen presenting cells, which were cultured with IL-4 and GM-CSF. On day 21, CTL activities of cultured effector cells were tested against autologous EB-blast cells as target cells pulsed with the stimulating peptides using 51Cr release assay. The results showed that cytotoxicity against insulin peptide binding to MHC class I A24 was observed in lymphocytes of four out of ten patients with type 1 DM. The mean cytotoxicity was 46.0% of the maximum release. The antibody against HLA-class I inhibited this effect. Cytotoxicity against GAD peptide which bind MHC class I A24 was not observed in seven patients. None of healthy controls showed cytotoxicity against GAD or insulin peptides was observed. This is the first report describing the cytotoxic activity of CD8+ T lymphocytes against insulin in type 1 DM. PMID- 11269890 TI - Insulin resistance and coronary risk factors in Japanese type 2 diabetic patients with definite coronary artery disease. AB - Insulin resistance is known as an important risk factor for coronary artery disease (CAD). However, CAD-related mortality in Japanese type 2 diabetics is lower than in Caucasians. To investigate whether insulin resistance is related to CAD in Japanese type 2 diabetics, we measured insulin sensitivity and several coronary risk factors in Japanese patients with type 2 diabetes with and without CAD. Thirty-three patients with definite CAD and 33 age- and sex-matched patients without CAD (control) were studied. Insulin sensitivity was assessed by the K index of insulin tolerance test (KITT). Clinical characteristics, classical risk factors, lipoprotein (a), and insulin sensitivity were compared between the two groups. Patients with CAD had a significantly longer duration of diabetes (9.0 +/ 1.4 vs. 5.5 +/- 0.9 years, P < 0.05, respectively), were mostly hypertensive (69.7 vs. 39.4%, P < 0.05), and more likely to be treated with insulin (45.5 vs. 18.2%, P < 0.05) compared with the control. Concerning the metabolic parameters, patients with CAD had a significantly higher insulin resistance than control (2.40 +/- 0.15 vs. 3.23 +/- 0.17%/min, P < 0.01, respectively), higher triglyceride (1.39 +/- 0.10 vs. 1.05 +/- 0.05 mmol/l, P < 0.05), lower HDL cholesterol (1.05 +/- 0.05 vs. 1.28 +/- 0.06 mmol/l, P < 0.05), and higher lipoprotein (a) (27.5 +/- 4.3 vs. 17.4 +/- 2.0 mg/dl, P < 0.05). Multiple logistic regression analysis indicated that hypertension, insulin resistance, high lipoprotein (a) and triglyceride, and low HDL cholesterol were independently related to CAD. Our results suggest that insulin resistance per se is a significant risk factor for CAD in Japanese patients with type 2 diabetes. PMID- 11269891 TI - A comparison of insulin suppression tests performed with somatostatin and octreotide with particular reference to tolerability. AB - To evaluate the tolerability of insulin suppression test (IST) using octreotide instead of somatostatin, we compared the steady-state plasma glucose (SSPG) values and the safety during and after the test in 17 normal volunteers. The subject received IST twice (with somatostatin or with octreotide) in random order. During the test, all subjects were infused with regular insulin and glucose simultaneously for 180 min. In addition, either somatostatin or octreotide was infused intravenously over the same period of time. Plasma glucose, insulin and C-peptide were measured. The subject response to the test was recorded during and one day after the test by a structured questionnaire. The SSPG and the steady-state plasma insulin (SSPI) values reached during IST were similar, irrespective of the use of somatostatin or octreotide. There was a positive correlation between the SSPG values obtained from both methods (r = 0.67, P = 0.003). However, the mean intra-individual coefficient of variation is 17.9% for SSPG. The SSPG levels, no matter from which method, correlated positively with the 2-h insulin after oral glucose challenge. Most adverse events (especially gastrointestinal discomfort) occurred after the test, and increased much more after using octreotide than somatostatin (P = 0.002 by chi 2 test). In conclusion, the SSPG values measured by IST using octreotide or somatostatin are similar in normal healthy subjects. Yet, the octreotide method has more adverse events after the test. PMID- 11269892 TI - Risk factors for the development of diabetic retinopathy in Japanese type 2 diabetic patients. AB - This study investigated the risk factors for development of diabetic retinopathy (DR) in 787 type 2 diabetic patients with no retinopathy at the first visit. The subjects were followed up for at least 3 years (mean, 6.7 years). Among the baseline factors, significant correlations were observed between the development of DR and HbA1c (P < 0.0001), the method of therapy (P < 0.005), the duration of diabetes at the first visit (P < 0.005) and the past maximal body mass index (BMI) (P < 0.01). No significant correlation was found with the blood pressure, age, gender, TC or BMI. Among the follow-up variables, the mean HbA1c (P < 0.0001) and duration of diabetes (P < 0.001) correlated significantly with DR development, whereas the blood pressure and age did not. We found that a 1% decrease in HbA1c led to a 35% reduction in the risk of development of DR during the follow-up. The patients whose HbA1c at the first visit was higher than the median value of 8.2% showed a higher probability of development of DR during the next 3 years even when the same blood glucose control was maintained during the follow-up. In conclusion, our study demonstrated that the most important risk factor influencing the development of DR was the blood glucose control. Moreover, we found that the glycemic level at the first visit also influenced the development of DR. PMID- 11269893 TI - Improved glycaemic control with miglitol in inadequately-controlled type 2 diabetics. AB - OBJECTIVE: The study compared the long-term efficacy and safety of miglitol to placebo in Type 2 diabetic outpatients inadequately controlled on combination therapy of diet, glibenclamide and metformin. METHODS: Type 2 diabetic patients (n = 154) receiving glibenclamide 7-20 mg/day and at least one 500-850 mg tablet metformin per day were randomized to receive additional miglitol or placebo for 24 weeks, titrated up stepwise from 25 to 100 mg trice daily. RESULTS: Addition of miglitol to sulphonylureas and metformin (per protocol analysis) produced a statistically, significantly greater reduction in HbA1c (-0.55%, P = 0.04) and postprandial glucose (-2.6 mmol/l, P = 0.0009) from baseline to endpoint than placebo (-0.2% and -0.6 mol/l, respectively). Reduction in fasting blood glucose was greater with miglitol than placebo, and there was a possible difference in favor of miglitol for fasting and postprandial triglyceride levels, but these did not reach statistical significance. Flatulence and diarrhea were reported by statistically, significantly more patients receiving miglitol than placebo, but adverse events overall were reported by only 10% more patients in the miglitol group. No cases of hypoglycaemia were reported. CONCLUSIONS: Miglitol can safely and effectively be added to long-term combination therapy in people with Type 2 diabetes inadequately controlled with glibenclamide plus metformin. PMID- 11269894 TI - Is insulin resistance or diabetes mellitus associated with stroke? An 18-year follow-up study. AB - Insulin resistance and/or diabetes are risk factors for coronary artery disease. However, it is still controversial whether they are associated with the development of stroke. A total of 304 Japanese men and women, aged 20-69 years, were selected on the basis of casual high blood glucose concentrations from 2732 participants of a population-based health examination in 1980. They all underwent a 50 g oral glucose tolerance test in 1981. Homa IR (index of insulin resistance) and Homa beta-cells (index of beta-cell function) were calculated from their fasting insulin and glucose using the formulas for the homeostasis model. They were followed-up for 18 years. Incidence of stroke was investigated by computed tomography. During 18 years, 28 subjects had a stroke; 21 had ischemic and nine had hemorrhagic strokes (two had both). Baseline variables, which showed an independent association with the incidence of stroke in the Cox proportional hazard model, were blood pressure, use of anti-hypertensive medications, and Homa beta-cell index (inversely) after adjustments for age and sex. After further adjustment for blood pressure using a step-forward method, Homa beta-cell was significantly related to the incidence of stroke (Hazard ratio: 0.65, 95% confidence interval: 0.44-0.95). In addition to hypertension, diabetes but not insulin resistance, is a risk factor for stroke. PMID- 11269895 TI - New administration routes in immunotherapy. PMID- 11269896 TI - Local conjunctival immunotherapy: the effect of dermatophagoides pteronyssinus local conjunctival immunotherapy on conjunctival provocation test in patients with allergic conjunctivitis. AB - BACKGROUND: We evaluated the effect of local conjunctival immunotherapy (LCIT) with standardized dermatophagoides pteronyssinus (Dp) extracts on antigen specific conjunctival provocation test (CPT) in patients with allergic conjunctivitis in a double blind, placebo-controlled study. We use the CPT because in our experience is the more objective parameter to evaluate the sensitivity to allergens in this patients. METHODS: The patients were selected on the basis of symptoms, positive prick test, positive CPT and elevated serum and tears total and specific IgE. The CPT was assessed with increased dilution of Dp extracts instilled into the lower fornix. Conjunctival hyperemia, tearing, itching, burning and swelling of eyelids were scored according a 4-point rating scale. Patients were randomly assigned to 2 groups of 12. The first group was treated with Dp extracts and the second group with placebo during 6 months. A drop of diluted antigen was instilled in both eyes daily, in 2-fold increased concentrations, the first 10 AU/ml. The maintenance dose was 1,000 AU/ml or the maximal dose which did not provoke symptoms. The symptoms were controlled with oral and/or local antihistamines. We evaluated the CPT before and after the treatment. The patients did not receive antihistamines during the 15 previous days to carrying-out the CPT. RESULTS: Ten of the twelve patients of the active group complete the treatment. One of the patients dropped out of the study because experienced local reaction with a dose of 1,000 AU/ml and refused to continue with the treatment. Other patient was disqualified for failure to comply with the protocol. One patient, which experienced itching and tearing with a dose of 1,000 AU/ml, tolerate 100 AU/ml. We continue with this dose until the end of treatment. The remaining patients tolerate 1,000 AU/ml as maintenance dose. A significant difference was observed in the score of CPT between LCIT treated patients and placebo group after 6 month of LCIT. CONCLUSIONS: We propose LCIT as a useful alternative to traditional subcutaneous immunotherapy in patients with allergic conjunctivitis. PMID- 11269897 TI - The inverse association between serum anti-streptolysin-O titers and the frequency of exacerbations of asthma in childhood. AB - BACKGROUND: The decline in infections in childhood may contribute to the rising severity and prevalence of atopic disorders in developed countries. Support for this hypothesis has been obtained from findings of an inverse association between tuberculin responses and atopy and from findings of high prevalence of asthma in certain islands with low prevalence of respiratory infections. With this regard, we investigated the association between serum anti-streptolysin-O (ASO) titers and the frequency of exacerbations of asthma in childhood. METHODS: Thirty atopic asthmatic children who has no sign of upper respiratory tract infection at the time of presentation or during the previous two months were included in the study. Serum ASO titer was measured as an indicator of past streptococcal upper respiratory tract infections. ASO titer > or = 200 Todd units was accepted as positive. RESULTS: A statistically significant association is found between high anti-streptolysin-O titers and decreased number of exacerbations in those children. CONCLUSIONS: Our data suggests that streptococcal infections might be a factor attenuating asthma in childhood. PMID- 11269898 TI - Effects of sublingual immunotherapy in patients sensitised to Ambrosia. An open controlled study. AB - BACKGROUND: Allergy to Ambrosia is a disease of growing importance in Europe. Injective and non-injective immunotherapy have been recognised as safe and effective but no evidence is currently available for sublingual immunotherapy (SLIT) in patients sensitised to Ambrosia. This study was planned to assess the effects and the safety of SLIT in patients clinically sensitised to Ambrosia. METHODS: 19 patients clinically sensitised to Ambrosia and treated with SLIT were compared to 14 patients treated only with drugs. Diary cards with symptoms and drug consumption were filled-in by patients during the pollen season whereas specific nasal challenge and skin prick test were run two months before and after the pollen season. Patients and doctors were also asked to express their subjective assessment about symptoms and drug consumption during the season. RESULTS: SLIT-treated patients had less symptoms and a significantly minor drug intake (p = 0.04) as compared to untreated patients. Nasal challenge test improved significantly in the SLIT group (p = 0.0001) but not in the control group (p = 0.6875) with a significant difference between groups at the end (p = 0.0413) but not at the beginning of the trial (p = 0.213). The decrease in skin reactivity was significant in the control group (p = 0.0186) and highly significant in the SLIT group (p < 0.0001), with no difference between groups (p = 0.2987). Subjective assessment from both patients and doctors was favorable to SLIT (p = 0.0005 for symptoms; p = 0.0019 for drug consumption). Only one minor local side effect was registered during SLIT. CONCLUSIONS: According to our data, SLIT in patients allergic to Ambrosia is safe and able to improve both subjective and objective parameters. PMID- 11269899 TI - Safety of nimesulide, meloxicam and rofecoxib as alternative analgesics. AB - Paracetamole and codeine are safe alternative analgesics for analgesic intolerant patients. Recently marketed selective and specific COX2 inhibitors are also considered to be safe for this group of patients. In this survey we wanted to disclose the safety of nimesulide and meloxicam and rofecoxib where they have been marketed recently in Turkey. PMID- 11269900 TI - Common variable immunodeficiency, insulin-dependent diabetes mellitus and celiac disease. AB - Common variable immunodeficiency is a disorder characterised by hypogammaglobulinemia with B-lymphocytes in peripheral blood and repeated infections. We report a child with a diagnosis of diabetes mellitus and celiac disease during lactation, and in whom common variable immunodeficiency was diagnosed at the age of 5. During evolution of the disease he presented multiple respiratory infections in spite of substitution therapy with gamma globulins. He presented pulmonary fibrosis with a pulmonary volume reduced, and a spirometric restrictive patron. Immunologically, he presents reduction in CD4 lymphoid population. He expresses the alleles DQ2 A1 0501 and B1 which are strongly associated with susceptibility to insulin-dependent diabetes mellitus and celiac disease, but don't express antigens HLA class II DR3 and DR4 that are more frequent in these entities. The main disease and all the complications had affected his curve pondostatural. PMID- 11269901 TI - Repeated furunculosis in adult male with abnormal neutrophil activity. AB - A 21 years old male suffered from repeated furunculosis in different regions of the body over the last two years. This coincided with the start of professional activities in hospital surroundings. The purulent secretions all showed growth of Staphylococcus aureus. All laboratory tests were normal except for a decrease of the neutrophil phagocytic ingestion phase. Before the diagnosis of defective phagocytosis was made, antibiotic treatment was started about 4 to 5 days after the appearance of the infectious process and the furunculosis led to abscess formation with difficult healing and cellulitis. After the diagnosis of defective phagocytosis ingestion phase, personal hygiene was intensified during and after work shifts at the hospital and antibiotic treatment was started at the first signs of folliculitis, which showed healing. PMID- 11269902 TI - Anaphylactic shock after intra-articular injection of carboxymethylcellulose. AB - BACKGROUND: Sodium carboxymethylcellulose (SCMC) is the sodium salt of a polycarboxymethyl ether of cellulose. SCMC is widely used in pharmaceutical and food industries. We present the case of a 47-year-old man who suffered an anaphylactic shock after an intra-articular injection of Trigon depot. METHODS AND RESULTS: Prick and intradermal tests with Trigon depot and its components (triamcinolone acetonide, Tween 80, benzylalcohol, SCMC), mepivacaine 2% and latex were performed. Challenge test with mepivacaine 2% was also realized. RESULTS: Showed a positive intradermal test to Trigon depot and carboxymethylcellulose, with negative results to the rest (including challenge test to mepivcaine 2%). CONCLUSIONS: Our results support the diagnosis of anaphylactic shock after intra-articular injection of carboxymethylcellulose and we consider necessary to emphasize that excipients must be taken into account as a potential source of adverse reactions to drugs. PMID- 11269903 TI - Aspirin-induced asthma needs a classification. PMID- 11269904 TI - [Involuntary hospitalization in the first psychotic episodes: associated factors]. AB - INTRODUCTION: The great majority of involuntary admissions at acute hospital units are related to severe psychiatric diseases. The aim of this study is to determine which are the characteristics and associated factors in first psychotic episodes not due to medical illness. METHOD: It had been evaluated, 61 first psychotic episodes hospitalised in a General Hospital Psychiatric Unit. It was used a protocol with the SCID, SCID-PANSS, DSM-IV criteria, unified clinical history, and PANSS, and Philips scales. RESULTS: 67.2% of the patients were committed to the hospital. Only the PANNS positive subscale was significantly associated with the involuntary admission. CONCLUSIONS: There is a high admission rate of involuntary inpatients with first psychotic episode. The involuntary admission is highly related to psychotic positive symptoms. PMID- 11269905 TI - [Body image in patients with dentofacial deformities treated by maxillofacial surgery]. AB - OBJECTIVE: Considerable controversies exist about the psychological impact of dentofacial deformities and the effects of treatment on body image. This research, which is a part of a longitudinal prospective study about a number of psychological aspects of adult patients with severe dentofacial deformities treated by orthognatic surgery, assesses the body image of a sample of patients before and after treatment. METHOD: Two subscales, general and facial-oclusal, of the modified Secord and Jourard test were used to evaluate body image of a sample of orthognatic surgery patients (N = 58). The questionnaires were completed at four time points: one week before surgery (T0), and one (T1), six (T6) and twelve months (T12) after surgery. RESULTS: Despite the severity of dentofacial deformity, presurgical mean scores of both subscales of body image test were normal. T6 scores were significantly higher than normative levels in both subscales (p < 0.001). T6 scores were also significantly higher than T0 in both general (p < 0.05) and facial-oclusal subscales (p < 0.001). T12 measurements showed significant differences in both subscales with respect to normative levels but significant differences between T12 and T0 were only found in facial oclusal subscale (p < 0.001). No differences were found between T12 and T6 scores. CONCLUSION: The positive effect of orthognatic surgery on body image is encouraging, but longer follow up is needed to evaluate the persistence of results and the eventual occurrence of any late change. PMID- 11269906 TI - [Assessment of the functioning of the daily life in people with long term mental disorder. Adaptation and reliability of the Spanish version of the "Basic Everyday Living Skills" (BELS)]. AB - INTRODUCTION: There is a close relationship between mental illness and the decay of global functioning. The level of support and care needed by a person will also depend closely of the level of his or her functioning. The aim of this article is to present the Spanish version of the Basic Every Day Living Schedule (BELS) and its reliability that has been studied on a sample of Spanish population. METHODOLOGY: Once the first translation into Spanish was satisfactory back translated into English, 77 residents in different sheltered homes were assessed by two pair of researchers. For the statistical analysis the Kappa coefficient was used. RESULTS: Kappa mean values for the opportunity scale was 0.791 and 0.743 for the performance scale. CONCLUSION: BELS is an assessment instrument with adequate reliability properties for the purpose for which it was conceived: to assess the basic abilities for the every day living. PMID- 11269907 TI - [Teaching survey for consultation-liaison psychiatry]. AB - BACKGROUND: Consultation-Liaison Psychiatry plays a major role in the training and clinical activity during the General Psychiatry Residency. Nevertheless, our national residency program for Psychiatry does not define either goals or distinct activities for the C-L Psychiatry rotation. METHODS: In order to: a) assess its current situation within the official teaching centers in Spain and b) determine the perceived opinion of the residents; a 41 item survey was mailed to 253 third and fourth year residents in Psychiatry to evaluate organizational issues of the C-L services, main aspects of this rotation, perceived quality, level of satisfaction and training needs. RESULTS: At least one resident from 40 out of the 67 accredited training centers answered the survey. The information gathered shown, as it was foreseen, that C-L training within the general psychiatry residency is very heterogeneous; furthermore it was documented some unsatisfaction regarding both the level of supervision and the rotation itself. CONCLUSIONS: Future studies are needed to get a closer look at all these aspects in order to improve the quality of both the rotation in C-L psychiatry and the standards of the C-L service itself. This endeavour shall benefit the training in C-L Psychiatry of the future spanish general psychiatrists. PMID- 11269908 TI - [Indications of group therapy for alcoholism treatment]. AB - INTRODUCTION: It exist a general need of improving the efficiency of psychiatry and psychological treatment based on evidence and individualized treatments. Group psychotherapy (GP) for alcoholics are used extensively in most specialized centers of Spain, but in our knowledge they are mostly indicated by routine and in a indiscriminate way. Furthermore, this therapies are scarcely evaluated and they do not exist homogeneous and objective criteria about their indication. MATERIAL AND METHOD: A cohort of 329 alcoholic patients was followed up until they finish the group psychotherapy treatment or either they drop out. RESULTS: Women show smaller drop out rates of the GP (51% to 2 years) that men (61%) and abandoned significantly afterwards (98 vs. 34 average weeks). In the younger patients is given the otherwise: they are retained less and they abandon before in spite of counting with specific groups for them. Subjects with medium educational level abandon sooner than those of high or low educational levels. Of all the psychiatric, alcohological and demographic antecedents only the presence of jealousy or of alcoholic hallucinations worsens significantly the outcome of GP. DISCUSSION: It is outlined the convenience of avoiding indications of GP when exist one of these worsening indicators and of minimizing indications of GP in younger patients or male patients, or otherwise to improve the therapeutic cares given to them during the critical phase of the first two months. Other alternative are to identify the personal and psychological characteristic of women and adults that favor the GP adherence to elaborate therapeutic strategies to maximize them in the other patients. PMID- 11269909 TI - [New advances in neuroimaging in the diagnosis of obsessive-compulsive disorder]. AB - Brain-imaging research provides evidence to suggest that the underlying disfunction in Obsessive-Compulsive Disorder (OCD) is likely to be in the prefrontal cortex-basal ganglia thalamic circuit rather then in any one single brain region. Early computerized tomography and magnetic resonance imaging studies have shown morphological changes in the basal ganglia. Now more sophisticated techniques are enhancing the information available, specially with regard to the caudate nucleus. The serotonergic hypothesis remains a necessary but not sufficient explanation for the pathogenesis of OCD. Most evidence remains focussed on the basal ganglia and on a 5-HT/dopamine inter-relationship. Given the basal ganglia receive such rich innervation from both 5-Th and dopamine neurones, it has been postulated that OCD is subserved by a neuronal dysfunction in the basal ganglia and orbitofrontal cortex circuit. Combining behavioural challenge with brain imaging may be a better approach to capturing brain function while patients with OCD and control subjects are actually observed. Using this techniques has made it possible to identified changes in response to treatment, whether the treatment is pharmacological or behavioural. However, there are not data enough that allow us to understand the anatomical, physiological and chemical mechanisms implicated in OCD. PMID- 11269910 TI - [Non-specific differences in neurocognitive performance in psychotic disorders?]. AB - INTRODUCTION: In the last years, there has been a growing interest in the study of cognitive functioning, not only in schizophrenia but also in other psychoses and psychiatric disorders. Attention, memory and information processing deficits may have great relevance as predictors of social adjustment and quality of life. OBJECTIVE AND METHODS: [corrected] We revised literature concerning comparative studies of neuropsychological performance in schizophrenia and affective disorders, mainly bipolar disorders. RESULTS: Studies analyzed show contradictory results. Some authors report a more global dysfunction or a more limited and concrete dysfunction in schizophrenia. However, most of the studies, especially those with homogeneous samples of psychotic affective disorders do not find any difference in cognitive functioning between schizophrenic and bipolar patients. DISCUSSION: Although some factors as sample selection, clinical state, course of the disorder, make difficult the interpretation of results, it would not exist specific deficits in any of the diagnostic groups studied. PMID- 11269911 TI - [Efficiency of melatonin in the treatment of insomnia]. AB - The aim of this research consists of studying whether melatonin (MLT) may be considered as an alternative for the treatment of insomnia. METHODS: A 33-year period (1966-1998) of computerised search using Medline database was performed. Melatonin, pineal gland and insomnia were used as key words. RESULTS: 93 articles were elicited; 85 were excluded because they were reviews (44) or were not directly related to the research topic (41). 111 insomniac patients were treated with MLT in 8 articles. 60% of the patients reported an improvement of sleep quality. Objective sleep measures also improved; there was a decrease in the sleep latency time and the number of awakenings (62% and 50% of patients respectively) after MLT treatment. CONCLUSIONS: Melatonin may have a promising future for the treatment of insomnia. The lack of standardised criteria for diagnosing sleep disorders and the lack of structured psychiatric interviews for ruling out psychiatric pathology are clear obstacles in generalizing these results. PMID- 11269912 TI - [Electroconvulsive therapy and serotonergic system]. AB - Evidence from several studies supports the involvement of several neurotransmitter systems in the mechanism of action and the clinical efficacy of the electroconvulsive therapy (ECT). However, more recent studies have reported serotonin, through the activation of several receptors, to be the neurotransmitter most frequently altered in ECT. With regard to the serotonergic system, several levels of alteration can be targeted, that concerning serotonin and its metabolite concentrations, that concerning changes in the density of presynaptic and postsynaptic receptors located both in brain tissue or in platelets, and finally, alterations at the intracellular signalling system or second messenger level. PMID- 11269913 TI - [Criminal behavior after orbitofrontal lesion]. AB - INTRODUCTION: Orbitofrontal lesions may produce abnormal social conduct, making impossible to live independently and even producing antisocial behaviour. CLINICAL CASES: Two young people with orbitofrontal lesion due to TH1 showed as chronic symptoms (18 months and seven years after injury) comportmental changes similar to antisocial personality disease, including criminal actions. Neuropsychologically the first case showed memory, fluency and secuentiation impairment. The second patient' performance was normal. CONCLUSIONS: Prefrontal lesion may impair self-regulation, producing syndromes that prevent normal every day life but are no much relevant in neurological and neuropsychological assessments. Orbitofrontal cortex plays an important role in social cognition which is the function that allows complex social behaviour. Because of the complexity of this function, symptoms can worse with the passage of time, thus a long follow-up of these patients is required. PMID- 11269914 TI - Single-cell RT-PCR as a tool to study gene expression in central and peripheral autonomic neurones. AB - In studies of the central and peripheral autonomic nervous system, it has become increasingly important to be able to investigate mRNA expression patterns within specific neuronal populations. Traditionally, the identification of mRNA species in discrete populations of cells has relied upon in situ hybridization. An alternative, relatively simple procedure is 'multiplex' reverse transcription polymerase chain reaction (RT-PCR), conducted on single neurons after their in vitro isolation. Multiplex single-cell RT-PCR can be used to examine the expression of multiple genes within individual cells, and can be combined with electrophysiological, pharmacological and anatomical (retrograde labelling) studies. This review focuses on a number of key aspects of this approach, methodology, and both the advantages and the limitations of the technique. We also provide specific examples of work performed in our laboratory, examining the expression of alpha 2-adrenergic receptors in catecholaminergic cells of the rat brainstem and adrenal medulla. The application of single-cell RT-PCR to future studies of the autonomic nervous system will hopefully provide information on how physiological and pathological conditions affect gene expression in autonomic neurones. PMID- 11269915 TI - Splanchnic-mesenteric capacitance bed in the postural tachycardia syndrome (POTS). AB - BACKGROUND: Gastroenterologic symptoms are common in the postural tachycardia syndrome (POTS), and postprandial worsening of orthostatic symptoms often occurs. We, therefore, investigated splanchnic-mesenteric vasoregulation in POTS. SUBJECTS AND METHODS: Eleven patients with POTS (one man, 10 women, 29.4 +/- 7.7 (S.D.) years) and 10 controls (two men, eight women, 27.9 +/- 5.6 years) participated in this study. The protocol included 5 min of 70 degrees head-up tilt (HUT) before and after a liquid meal, as well as 1.5 min of hyperventilation. Blood pressure (BP), heart rate (HR), endtidal CO2, and cardiovascular indices derived from thoracic electrical bioimpedance were continuously monitored. Superior mesenteric artery (SMA) blood flow was measured by real time Doppler ultrasound and analyzed off-line. Cross-sectional area of SMA (SMA-area) and time-averaged velocity (SMA-TAV) were measured; SMA blood flow (SMA-BF) and vascular resistance (SMA-VR) were derived. RESULTS: The following significant results were found: at supine rest, the POTS group had higher HR, BP, SMA-TAV and SMA-BF and a lower SMA-VR than the control group. HUT resulted in a reduction of pulse pressure, CO2 level, SMA-area, SMA-TAV and SMA-VF and increment of HR and SMA-VR in both groups. The POTS group underwent greater increment of HR and greater reduction of CO2 than controls. Hyperventilation induced increment of HR and cardiac index (CI) and reduction of SMA-VR in controls; no significant change occurred in POTS. The test meal induced increments of HR, CI, SMA-area, SMA-TAV and SMA-VF and reduction of SMA-VR in patients and controls for both supine rest and HUT. CONCLUSION: The main novel observations of increased resting SMA-BF, SMA-TAV supine, and reduced SMA-VR when compared with controls support the notion that there is excessive splanchnic capacity (pooling) at rest in POTS. PMID- 11269916 TI - Linear and non-linear 24 h heart rate variability in chronic heart failure. AB - It has recently been demonstrated that SDNN of heart rate variability (HRV) is a useful independent prognostic tool in chronic heart failure (CHF). The purpose of the present study was to evaluate if spectral and non-linear analysis of 24-h HRV, considered markers of autonomic cardiac modulation, contain independent prognostic information in CHF patients. Twenty normal subjects and thirty consecutive outpatients with clinically stable CHF were studied for 2 years. Periods of 300 R-R intervals were analyzed from Holter recordings. The power spectral analysis, the slope of the linear relationship between log-power versus log-frequency (1/f), and the complexity content (using corrected conditional entropy; CCE) of the R-R series were calculated. The normalized power of the low frequency spectral component (LF) and the 1/f slope were significantly lower in patients compared to controls (respectively 30.1 +/- 3.0 vs. 48.6 +/- 3.4 and 1.27 +/- 0.04 vs. -1.08 +/- 0.05; P < 0.05). Moreover, the patients who died during the study presented a reduced LF (20.9 +/- 4.1 vs. 35.5 +/- 3.5 nu; P < 0.05) and a steeper 1/f slope (-1.40 +/- 0.09 vs. -1.21 +/- 0.04 nuts, P < 0.05) compared to survivors. These results remained significant in a logistic model including heart rate and SDNN. The information content present in spectral and non-linear analysis of HRV in CHF patients has prognostic relevance independently from the time domain measures of HRV. In particular, the reduction of LF power seems the best indicator among those considered. PMID- 11269917 TI - Topical anesthetic before microneurography decreases pain without affecting sympathetic traffic. AB - Pain associated with the microneurography procedure varies among human research volunteers, and may influence baseline sympathetic neural activity. The purpose of this study was to evaluate the efficacy and effects of applying a topical anesthetic prior to microneurography. Ten volunteers underwent microneurography twice, separated by a minimum of 4 weeks. Using a single-blind, randomized cross over design, EMLA cream (2.5% lidocaine and 2.5% prilocaine in oil emulsion) or an aqueous placebo cream was applied 2 h prior to each session. Subjects rated pain on a scale from 0 (no pain) to 4 (extreme pain). The electrocardiogram, and efferent sympathetic nerve traffic from peroneal nerve muscle fascicles at the popliteal fossa were recorded during a 10-min supine rest period. EMLA cream significantly reduced perception of pain (P < 0.05), but did not affect burst reflex latencies from preceding R-waves or total muscle sympathetic nerve traffic (P > 0.05). These data show that use of EMLA cream prior to microneurography is innocuous, and do not support the hypothesis that baseline sympathetic traffic is increased by pain or discomfort associated with microneurography. PMID- 11269918 TI - Peripheral sympathetic function as a predictor of complex regional pain syndrome type I (CRPS I) in patients with radial fracture. AB - Complex regional pain syndrome type I (CRPS I) is a frequent complication after injuries of the upper limbs. The pathophysiology of this disease remains unclear, although disturbances of the sympathetic nervous system have been detected in several clinical studies, and sympathetic blocks resolve the symptoms in many of the cases. To investigate the meaning of sympathetic dysfunction at the beginning of the disease, 27 patients with distal radial fracture were examined prospectively during the course of the disease with regard to their clinical symptoms and their peripheral sympathetic nervous function. Sympathetic nervous function was examined by testing the vasoconstrictor response to sympathetic stimuli--recorded with laser Doppler fluxmetry--of the fingertips of both hands. Four patients developed CRPS I during the 12-week observation time and two patients presented an incomplete clinical CRPS I picture ('borderline patients'). The complaints of all patients (normal fracture patients, CRPS I patients, borderline patients) were similar during the first week after trauma with focus on pain, motoric disturbances and autonomic symptoms. After 1 or 2 weeks, a larger clinical difference developed between normal fracture patients and CRPS I or 'borderline patients'. In CRPS I patients and 'borderline patients', the sympathetic vasoconstrictor response was diminished or absent from the first posttraumatic day throughout the observation time, whereas the normal fracture patients revealed slightly impaired sympathetic nervous function on the first posttraumatic day and normal results during the rest of the observation time. With regard to the unaffected contralateral hand, CRPS I patients also showed impaired sympathetic nervous function. The results of the present study suggest that the disturbances in the sympathetic nervous system in CRPS I patients are systemic and not limited to the affected limb. Their occurrence before the clinical breakout of the disease may serve as a marker that might be useful for early therapy and lead to further understanding of the pathophysiology of CRPS I. PMID- 11269919 TI - Substance P and calcitonin gene-related peptide immunoreactive sensory DRG neurons innervating the lumbar facet joints in rats. AB - The rat L5/6 facet joint is innervated from L1 to L5 dorsal root ganglions (DRGs) multisegmentally. Sensory fibers from L1 and L2 DRGs were reported to innervate nonsegmentally through the paravertebral sympathetic trunks, while those from L3 to L5 DRGs segmentally innervate the L5/6 facet joint. The presence of substance P (SP) and calcitonin gene-related peptide (CGRP) immunoreactive (ir) nerve fibers has been demonstrated in the lumber facet joints, but their ratios have not been determined. Fluoro-gold (F-G) labeled neurons innervating the L5/6 facet joint were distributed throughout the DRGs for levels L1 to L5. Of the F-G labeled neurons, the ratios of SP-ir L1, L2, L3, L4 and L5 DRG neurons were 13, 15, 29, 31 and 30%, respectively, and those of CGRP-ir neurons were 17, 24, 44, 56 and 50%, respectively. The ratios of SP and CGRP-ir neurons in L1 and L2 DRGs were significantly less than those in L3, L4 or L5 DRGs. In conclusion, the neurons of L3, L4 and L5 DRGs may have a more significant role in pain sensation of the facets than L1 and L2 DRG neurons. PMID- 11269920 TI - Dorsal horn administration of L-arginine accentuates the pressor response evoked by activation of muscle mechanoreceptors. AB - Recent work from our laboratory suggests that nitric oxide production in the dorsal horn has a modulatory influence on the pressor reflex evoked by static contraction of skeletal muscle. In this study, we tested the hypothesis that nitric oxide production in the dorsal horn is involved in producing the pressor reflex elicited by activation of skeletal muscle mechanoreceptors. Cats were anesthetized with alpha-chloralose (80 mg/kg) and urethane (100 mg/kg) and a laminectomy was performed. With the exception of the L7 dorsal root, the dorsal and ventral roots from L5 to S2 were sectioned on one side. Muscle mechanoreceptors were activated by manually stretching the ipsilateral triceps surae muscle 1.5 cm. To block nitric oxide synthase, a 50 mM solution of nt-nitro L-argenine methyl ester (L-NAME) (a dose that altered the pressor reflex to static contraction) was microdialyzed into the dorsal horn at L6 and S1. Dialysis of L-NAME failed to attenuate the peak change in mean arterial pressure evoked by muscle stretch (45 +/- 6 mmHg before and 44 +/- 9 mmHg after 2 h of L-NAME dialysis). On the other hand, 2 h of L-arginine dialysis (50 mM) increased the peak pressor response to muscle stretch from 43 +/- 3 to 57 +/- 5 mmHg. These data suggest that administration of L-arginine enhances the excitability of dorsal horn cells receiving input from muscle mechanoreceptors, thus increasing the pressor response evoked by activation of this type of muscle afferent neuron. PMID- 11269921 TI - Cotransmission from sympathetic vasoconstrictor neurons: differences in guinea pig mesenteric artery and vein. AB - Vasoconstrictor responses to electrical field stimulation (EFS, 0.2-32 Hz, 0.1 ms, 12 V, for 1 min) were measured in endothelium-denuded segments of guinea-pig mesenteric vein and compared to responses in mesenteric artery. The distribution of both tyrosine-hydroxylase-like immunoreactivity (TH-LI) and neuropeptide Y like immunoreactivity (NPY-LI) was also studied using anti-TH and anti-NPY antibodies. The effect of exogenous NPY (10 nM) on EFS (8 Hz, 0.3 ms, 12 V, for 1 min)-evoked overflow of noradrenaline (NA) was also studied using an HPLC technique with electrochemical detection. Veins responded with contractions at lower frequencies of stimulation than arteries. Prazosin (0.1 microM) abolished the EFS-evoked contractions in artery at 0.5-32 Hz and in vein at 0.2-1 Hz of stimulation. However, in vein, the contractile responses to EFS at 2-32 Hz of stimulation were only reduced by prazosin. Phentolamine (1 microM) abolished the responses to 0.5-4 Hz and reduced the responses to 8-32 Hz of EFS in artery. In vein, phentolamine (1 microM) abolished the responses to 0.2-1 Hz and facilitated the contractions elicited by 16-32 Hz. The NPY-receptor antagonist BIBP3226 (1 microM), in combination with phentolamine, abolished contractions in vein. Yohimbine (0.1 microM) abolished the responses to lower frequencies of stimulation in both artery (0.5-2 Hz) and vein (0.2-1 Hz). The responses to greater frequency stimulation were not affected by yohimbine in artery, and were facilitated in vein. Pre-treatment of animals for 24 h with reserpine abolished contractile responses to EFS in artery, whereas in vein, responses to 0.2-2 Hz were abolished while responses to 4-32 Hz were unchanged. Suramin (100 microM) or alpha,beta-methylene ATP (alpha,beta MeATP; 10-100 microM) treatment did not affect the contractile responses to EFS in either artery or vein. Pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid tetrasodium (PPADS; 30 microM), even potentiated the responses to 2-16 Hz in vein. However, following resperine treatment, both PPADS and suramin reduced the nerve-evoked contractions of vein. Either BIBP3226 (1 microM) alone or BIBP3226 in combination with PPADS or suramin abolished the contractile response to EFS in reserpine-treated veins. NPY (100 nM) produced significantly more contraction in vein than in artery (i.e., 93 +/- 2.5 versus 7 +/- 4% of the response to 70 mM KCl, respectively). NPY (10 nM) significantly reduced the NA overflow evoked by EFS at 8 Hz. Flat mount preparations and cryostat sections of both mesenteric artery and vein revealed that TH-LI and NPY-LI were co-localized in a dense network of fibers within the adventitial layer. In conclusion, NA exclusively mediates the contractile response to sympathetic nerve stimulation in guinea-pig mesenteric artery, whereas at least three neurotransmitters [i.e., NA, adenosine 5'-triphosphate (ATP) and NPY] are involved in the neural response of mesenteric vein. PMID- 11269922 TI - Vagal ganglionic and nonadrenergic noncholinergic neurotransmission to the ferret lower oesophageal sphincter. AB - In the present study we aimed to discretely characterise ganglionic and neuroeffector transmission to the ferret lower oesophageal sphincter (LOS) using a novel preparation of LOS muscle with intact vagal innervation in conjunction with isolated LOS muscle strips. In this way we could compare vagally mediated LOS relaxation with that of enteric inhibitory motorneurones which were directly stimulated. Preparations of LOS muscle, with or without attached vagus nerves, were dissected from adult ferrets and maintained under preload in organ baths, where LOS muscle developed spontaneous tone. LOS relaxations in response to vagal stimulation (0.5-5 Hz, 30 V) were recorded, alone and following pretreatment with tetrodotoxin (TTX), hexamethonium (Hex), Hex and atropine and NG-nitro-L-arginine (L-NNA). Direct activation of enteric inhibitory motorneurones was performed via electrical field stimulation (EFS). Vagal stimulation elicited frequency dependent relaxations of the LOS that were abolished by tetrodotoxin (1 microM) and markedly reduced following L-NNA pretreatment (100 microM), but unaltered following pretreatment with the selective VIP or PACAP receptor antagonists VIP (10-28) or PACAP (6-38), respectively (each at 5 microM). The potent NOS inhibitor S-methyl-L-thiocitrulline (100 microM) inhibited LOS relaxation to the same degree at 5 Hz. Hex alone (500 microM) reduced maximal relaxation by 50%; in combination with atropine (2 microM), relaxation was almost abolished. In isolated LOS muscle strips, neither VIP (10-28) nor PACAP (6-38) altered EFS induced relaxation. Taken together, these results suggest ganglionic neurotransmission to the ferret LOS occurs mainly through a combination of nicotinic and muscarinic receptors and utilises nitroxidergic enteric inhibitory motorneurones to relax the LOS. Moreover, LOS relaxation due to direct activation of inhibitory motorneurones also utilises primarily nitric oxide and other as yet undefined neurotransmitters. Neither VIP nor PACAP are involved in vagally mediated or direct enteric neuronally stimulated LOS relaxation in the ferret. PMID- 11269924 TI - Constitutive nitric oxide release modulates neurally-evoked chloride secretion in guinea pig colon. AB - The role of constitutive nitric oxide (NO) release in enteric neural pathways regulating ion transport was examined in guinea pig distal colon, in vitro and ex vivo. In in vitro studies, 43% of colonic preparations exhibited oscillations in baseline short-circuit current (Isc), which were reduced by tetrodotoxin (TTX). The NO chelator, hemoglobin (Hb), and neuronal NO synthase inhibitor, 7 nitroindazole (7-NI), significantly increased the baseline Isc in these tissues, which was reduced by TTX. In tissues without oscillations in baseline Isc, Hb reduced the Isc, while 7-NI had little effect. In all tissues, electrical field stimulation (EFS; 15 V/10 Hz) caused a biphasic increase in the Isc which was enhanced by both Hb and 7-NI. In the ex vivo studies, basal release of nitric oxide was significantly lower in colonic segments isolated from guinea pigs administered N omega-nitro-L-arginine methyl ester (L-NAME) i.p. compared to control tissues. Moreover, carbachol, caused a 10-fold increase in NO release in control tissues, but had no effect in tissues isolated from the L-NAME group. L NAME increased tissue conductance and EFS-induced changes in Isc, which were reversed by L-arginine. However, carbachol-induced ion secretion was unaltered in the L-NAME group compared to control animals. The results suggest that, in guinea pig colon, constitutive enteric NO release tonically suppresses submucous neural activity and it is involved in the maintenance of basal epithelial chloride secretion and mucosal permeability. Hence, constitutive NO promotes a delicate balance between pro-absorptive and pro-secretory processes in guinea pig colon. PMID- 11269923 TI - Peptide immunoreactivity and ultrastructure of rat urinary bladder nerve fibers after topical desensitization by capsaicin or resiniferatoxin. AB - In the present study the decrease of neuropeptide containing nerve fibers and the increase in the volume threshold to reflex micturition occurring in the rat bladder after intravesical application of capsaicin or resiniferatoxin were compared. The ultrastructure of bladder terminal axons was evaluated at the moment of maximal peptide depletion and compared to that of nerve fibers after systemic capsaicin application. Adult Wistar rats were treated intravesically for 30 min with 0.5 ml of 100 nM RTX, 1 mM capsaicin or 30% ethanol in saline, the vehicle solution. Twenty-four hours and 1, 2, 3, 4 and 8 weeks later the bladders were immunostained for CGRP, SP, VIP and NPY. Cystomanometric studies were performed 24 h and 1, 8, and 12 weeks after vanilloid instillation. Twenty-four hours after systemic capsaicin or intravesical capsaicin or RTX, bladders were prepared for electron microscopic (EM) observation. Intravesical capsaicin or RTX decreased, in a similar way, the number of CGRP and SP-IR (immunoreactive) fibers coursing in the muscular layer and the mucosa. IR fibers amounted to less than 20% of controls at 24 h and returned to normal levels in the eighth week. At the EM level, bladders treated with topical vanilloids did not show morphological changes in terminal axons coursing in the mucosa. In contrast, bladders from animals treated systemically with capsaicin contained numerous grossly degenerated nerve fibers. VIP and NPY-IR fibers were not affected by the treatment. Cystometrograms showed an increase of the volume threshold to reflex micturition that started at 24 h and disappeared at 12 weeks. We conclude that intravesical capsaicin or RTX were equally effective in terms of reducing the number of SP and CGRP-IR fibers and increasing the volume threshold for reflex micturition. Both changes were transient and were not associated with ultrastructural changes of the bladder nerve fibers, excluding terminal axon degeneration as the main mechanism of action of intravesical vanilloids. PMID- 11269925 TI - Effects of both the emotional behavior and feeding conditions on the circulating plasma volume and plasma glucose levels in cats. AB - Influence of hypothalamically induced emotional behavior on the circulating plasma volume, plasma levels of glucose, epinephrine (E), norepinephrine (NE), dopamine (DA) and cortisol were examined in awake cats under both fasted and fed conditions. Restlessness was evoked intermittently for 6 h by electrical stimulation of the anteromedial hypothalamus (AMH). Blood was sampled immediately before, 1 h after and 6 h after the start of stimulation. Changes in the plasma volume was calculated by changes of hemoglobin (Hb) and hematocrit (Ht). As the control group, another 7 cats with electrodes implanted but unstimulated were identically treated under both fasted and fed conditions. Both E and glucose levels in restlessness group once markedly increased after 1 h and then tended to decrease after 6 h, whereas NE levels in restlessness group increased after 1 h and further increased after 6 h, whether cats were fasted or fed. DA levels increased under the fasted condition of restlessness. The cortisol level markedly increased in both fasted and fed restlessness groups. The plasma volume in control group increased under the fed condition, while in restlessness group it decreased remarkably and tended to decrease more in a fasted state than in a fed state. These results indicated that AMH induced restlessness elicited marked sympatho-adrenal activation, hyperglycemia and hemoconcentration, whether cats were fasted or fed. Relationship among such responses, and the difference in responses between fasted and fed conditions were also discussed in the paper. PMID- 11269926 TI - Nociceptin inhibits capsaicin-sensitive contraction to mesenteric nerve stimulation in the guinea-pig isolated ileum. AB - Mesenteric nerve stimulation (MNS) in the presence of guanethidine and hexamethonium antidromically stimulated extrinsic sensory nerve fibers and cholinergic myenteric motor neurons, resulting in longitudinal muscle contraction in the isolated guinea-pig ileum. Nociceptin (NC) is a recently discovered neuropeptide that structurally resembles an opioid peptide. The aim of the current study was to examine how NC affects the contractile responses to MNS in the isolated guinea-pig ileum, in comparison with an opiate, methionine enkephalin. These contractions were auxotonically recorded and their amplitude was analyzed. NC (1-100 nM) and methionine-enkephalin (0.1-10 microM) concentration-dependently inhibited the response to MNS (20 Hz, 0.5 ms, supramaximal currents). Naloxone (10 microM) significantly diminished the inhibitory effect of methionine-enkephalin (0.1-10 microM), but did not antagonize the inhibitory effect of NC (1-100 nM). We conclude that NC receptors, distinct from opioid receptors, exist on the capsaicin-sensitive sensory nerve fibers and/or myenteric cholinergic motor neurons in the guinea-pig ileum and that specific antagonists for these NC receptors are not found yet. PMID- 11269927 TI - Effect of hypoxia on the activity of respiratory and non-respiratory modulated retrotrapezoid neurons of the cat. AB - The retrotrapezoid nucleus (RTN), a part of the rostral ventrolateral medulla, is involved in the control of breathing. A recent immunohistological study suggested a possible involvement of the RTN in hypoxic chemoreflex loop. The present electrophysiological study performed in the cat demonstrates that 23 out of 24 RTN neurons were stimulated during the biphasic respiratory response to hypoxia, which consists of a reinforcement followed by a depression of respiratory activity. This confirms the previous immunohistological study. While 15 RTN neurons might integrate either phase I (n = 7) or phase II (n = 8) O2 chemosensitive inputs, the remaining eight RTN neurons stimulated by hypoxia are susceptible to integrate both phase I and phase II O2-chemosensitive inputs. In conclusion, our results suggest that the different subsets of RTN neurons may influence respiratory output by conveying signals originating from peripheral and/or central chemoreceptors stimulated during hypoxia. PMID- 11269928 TI - Descending influences from the infralimbic cortex on vago-vagal reflex control of gastric motor activity in the rat. AB - In experiments on urethane anaesthetised rats the influence of electrical stimulation of ventral areas of the medial prefrontal cortex (mPFC) on spontaneous and vagally-mediated gastric motility were studied. Stimulation of the mPFC resulted in gastric relaxation manifested as a fall in intragastric pressure from a baseline value of 5.0 +/- 0.5 cm H2O. These were most prominent following a short latency when the infralimbic cortex (IL) was stimulated (27.4 +/- 2.5% fall in gastric pressure). Electrical stimulation of the central end of one cervical vagus nerve caused a comparable decrease in gastric pressure (27.1 +/- 2.9%). The cortical mediated relaxation was reduced by atropine and abolished by vagotomy. The cortically induced gastric relaxation followed a shorter latency (5.9 +/- 1.0 s), time to nadir (20.1 +/- 2.7 s) and the half recovery time (21.5 +/- 4.0 s) than vagally mediated-relaxations (9.9 +/- 2.3, 56.0 +/- 5.3 and 83.4 +/- 9.5 s, respectively). Vagally mediated relaxations were inhibited by simultaneous stimulation of the infralimbic cortex. In this case the decrease of gastric pressure, the time to nadir and the half recovery time were significantly decreased in comparison with the gastric relaxatory responses to vagal stimulation alone (P < 0.05). We conclude that one way in which the mPFC influences gastric motility is via corticofugal projections from the infralimbic cortex to the brain-stem which modulate transmission of vago-vagal reflexes. PMID- 11269929 TI - Evidence that nerve growth factor promotes the recovery of peripheral neuropathy induced in mice by cisplatin: behavioral, structural and biochemical analysis. AB - In this study we investigate the neurotoxic action of Cisplatin (6 micrograms/g body weight for 5 treatment cycles during 15 weeks with a total dose of 30 micrograms/g), an antitumor drug, and its effect on the level of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) in peripheral tissues. We found that Cisplatin in adult rodents impairs peripheral sensory function and both sympathetic and sensory peripheral innervation as shown by the hot-plate response, catecholamine distribution and substance P immunoreactivity respectively. These changes are associated with decreased NGF in intestine, paws, and bladder while NGF increased in the spinal cord. Also BDNF decreased in bladder and paws and increased in spinal cord and intestine. To further investigate the role of NGF in the pathogenesis of Cisplatin-induced peripheral neuropathies a group of animals was injected with NGF (1 microgram/g every 4 days for 4 times) following Cisplatin treatment and evaluated for sensory function, sympathetic and sensory innervation and BDNF levels. Data demonstrated that exogenous NGF administration is able to restore biochemical, structural and functional changes induced by Cisplatin. These findings suggest that the reduction of NGF availability could be a cause of Cisplatin-induced peripheral neuropathies and that NGF exogenous administration could prevent or reduce Cisplatin neurotoxicity also in cancer patients, reducing the side effects of chemotherapy. PMID- 11269930 TI - Severe cardiovascular autonomic dysfunction in a patient with Guillain-Barre syndrome: a case report. AB - Autonomic dysfunction is a frequent and severe complication of Guillain-Barre syndrome. It is often responsible for cardiovascular abnormalities, even cardiac arrest. We report a 49-year-old patient, who suffered from Guillain-Barre syndrome necessitating mechanical ventilation. He showed wide fluctuations of blood pressure and heart rate spontaneously or in relation with medical procedures. Though heart rate variability (HRV) and baroreflex sensitivity (BRS) values derived from different methods were extremely low, vigorous stimuli, like eyeball pressure test and carotid sinus massage, produced exaggerated responses, like severe bradycardias, hypotension and cardiac arrest. Despite the plasma exchange and supportive therapies, the patient finally developed adult respiratory distress syndrome (ARDS), sepsis and died due to septic shock. PMID- 11269931 TI - Patterns of palmar skin temperature alterations during transthoracic endoscopic T2 sympathectomy for palmar hyperhidrosis. AB - Transthoracic endoscopic T2 sympathectomy has been widely applied to the treatment of a variety of sympathetically mediated disorders. Palmar hyperhidrosis is probably the most common indication for thoracic sympathectomy, especially in certain subtropical areas. Which sympathetic ganglion is to be ablated and how extensive such ablation is enough to eliminate palm sweating are two important issues. Intraoperative monitoring of palmar skin temperature (PST) is the most frequently used method for assessing the accuracy as well as adequacy of ablation of the target sympathetic ganglia. With continuous monitoring of bilateral PST during the operative course of T2 sympathectomy, it was possible to depict the alterations of bilateral PST in response to specific surgical procedures in a real-time manner. For each case, a PST graph was obtained, which represented the graphical expression of intraoperatively recorded bilateral PST data plotted against time. The PST graphs of 93 consecutive cases were analysed. Three types of PST graphs existed, reflecting different responses of bilateral PST to different surgical procedures during the operation. In Type I PST graph pattern, found in 58 cases, skin incision and intercostal muscle dissection caused dramatic bilateral PST drop; and unilateral T2 sympathectomy induced synchronous bilateral PST elevation. Twenty-four cases demonstrated Type II PST graph pattern, in which unilateral T2 sympathectomy caused only ipsilateral PST elevation, although the PST-depressing effect of skin incision and muscle dissection was as significant as in Type I graph pattern. In the 11 cases who showed Type III PST graph pattern, neither skin incision nor T2 sympathectomy induced any apparent changes of PST on either side, giving rise to two rather flat PST curves on the PST graphs. These findings implicate that reciprocal interactions between bilateral sympathetic activities exist in the majority of cases, and that crossover sympathetic modulation may play a role in the neural control of the sudomotor and vasomotor activities of the palms. This study also provides information regarding how PST would possibly change following specific surgical procedures during transthoracic endoscopic T2 sympathectomy, which may be of importance to those who use intraoperative PST monitoring as a guide in determining whether or not the correct sympathetic ganglia are ablated for adequate sympathetic denervation of the palms. PMID- 11269932 TI - Effect of fatty acids on phase behavior of hydrated dipalmitoylphosphatidylcholine bilayer: saturated versus unsaturated fatty acids. AB - The effect of some fatty acids on the phase behavior of hydrated dipalmitoylphosphatidylcholine (DPPC) bilayer was investigated with special interest in possible difference between saturated and unsaturated fatty acids. The phase behavior of hydrated DPPC bilayer was followed by a differential scanning calorimetry and a Fourier transform infrared spectroscopy. The addition of palmitic acid (PA) increased the bilayer phase transition temperature with the increase of the PA content in the mixture. In addition, DPPC molecules in gel phase bilayer became more rigid in the presence of PA compared with those in the absence of PA. This effect of PA on the phase behavior of hydrated DPPC bilayer is common to other saturated fatty acids, stearic acid, myristic acid, and also to unsaturated fatty acid with trans double bond, elaidic acid. Contrary to these fatty acids, oleic acid (OA), the unsaturated fatty acid with cis double bond in the acyl chain, exhibited quite different behavior. The effect of OA on the bilayer phase transition temperature was rather small, although a slight decrease in the temperature was appreciable. Furthermore, the IR spectral results demonstrated that the perturbing effect of OA on the gel phase bilayer of DPPC was quite small. These results mean that OA does not disturb the hydrated DPPC bilayer significantly. PMID- 11269933 TI - The application of dimethyldioxirane for the selective oxidation of polyfunctional steroids. AB - Oxidation and epoxidation reactions of a series of structurally different steroids related to methyl 5 beta-cholanoates having hydroxyl groups and/or double bonds by treatment with dimethyldioxirane (DMDO) are described. Steroidal alcohols, olefines, and unsaturated alcohols and conjugated enones with DMDO were transformed into ketones, epoxides, and epoxy-ketones, respectively, in good isolated yields. The regio- and stereoselectivities for DMDO reaction differing from those observed for organic peracids, tert-butyl hydroperoxide and alkaline hydrogen peroxide are also discussed. PMID- 11269934 TI - A novel fluorescent coronenyl-phospholipid analogue for investigations of submicrosecond lipid fluctuations. AB - A fluorescent phospholipid derivative, the 2'-(4-coronenylbutyric) ester of lyso egg phosphatidylcholine, has been synthesized for use in studies of submicrosecond lipid dynamics. Synthesis of the phospholipid derivative involves Friedel-Crafts acylation of free coronene, followed by a Huang-Minlon reduction to yield the fatty-acyl derivative, 4-coronenylbutyric acid. Esterification of the carboxylic acid with lyso-phosphatidylcholine is achieved through a mixed anhydride intermediate. The resultant coronenyl-phospholipid adduct (Cor-PC) has been incorporated into sonicated unilamellar vesicles of dimyristoylphosphatidylcholine (DMPC) for dynamic lipid studies. Fluorescence quenching studies using potassium iodide, together with steady-state emission anisotropy (EA) measurements, confirm that the coronene moiety of the phospholipid adduct resides towards the head group interfacial region of the lipid bilayer. Unique properties of this new fluorescent phospholipid adduct are its long mean fluorescence lifetime (tau av approximately 112 ns at 14 degrees C), the planar symmetry of the fluorophore and its defined bilayer location. As a consequence, depolarizing motions of the coronene moiety target submicrosecond 'gel-fluid' lipid dynamics arising from a relatively narrow bilayer distribution. Our data suggest that the sensitivity of this new long-lived fluorescent phospholipid analogue to localized transverse submicrosecond lipid dynamics can provide important biological insights into varied processes including lipid peptide interactions, bilayer fluidity gradients and passive ion transport. PMID- 11269935 TI - Interfacial properties of phosphatidylcholine bilayers containing vitamin E derivatives. AB - alpha-Tocopherol and alpha-tocopheryl succinate are biologically active lipids. The activity of these lipids may be related to how they affect membrane physical chemical properties. Utilizing fluorescence methods, we have investigated the effect of alpha-tocopherol, alpha-tocopheryl succinate, and alpha-tocopheryl acetate on the properties of model membranes consisting of 1-palmitoyl-2-oleoyl sn-glycero-3-phosphocholine and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine. In liquid-crystalline phase phospholipid bilayers, alpha-tocopherol decreased acyl chain mobility and decreased the interfacial polarity, but had no effect on the interfacial surface charge. In contrast, alpha-tocopheryl succinate had little effect on acyl chain motion or interfacial hydration, but increased the interfacial surface charge. alpha-Tocopheryl acetate had very little effect on any of the measurements of these bilayer properties. In a gel phase bilayer, alpha-tocopherol decreased acyl chain order, whereas alpha-tocopheryl succinate and alpha-tocopheryl acetate did not. Each alpha-tocopheryl derivative had a different effect on interfacial polarity, however, only alpha-tocopheryl succinate increased the interfacial surface charge. The acylation of alpha tocopherol abolishes its antioxidant activity and generates molecules with different membrane physical properties. The non-polar acetate group of alpha tocopheryl acetate locates this compound in a region of the bilayer where it has little effect on bilayer interfacial properties. The free carboxyl group of alpha tocopheryl succinate is located in the interfacial region of the bilayer where it increases the membrane surface charge. PMID- 11269936 TI - Photophysical studies of 3,3' dioctadecyloxacarbocyanine dye in model biological membranes and different solvents. AB - The absorption and fluorescence spectra of 3,3'-dioctadecyloxacarbocyanine [DiOC18(3)], a cationic oxacarbocyanine dye have been studied in aqueous and nonaqueous media containing egg phosphatidylcholine (PC) as well as in different solvents of diverse nature. The results show the evidence of complex formation of the dye in the ground and in the excited states with PC. The excited state interaction of the dye with PC suggests the electron transfer from PC to dye and this is supported by photovoltage generation in a photoelectrochemical cell consisting of dye and PC in aqueous medium. An attempt has been made to determine the polarity of the microenvironment of the dye in PC liposome or PC reverse micelle from the spectral studies of the dye in different solvents of known polarity. PMID- 11269937 TI - Membrane perturbing properties of sucrose polyesters. AB - Sucrose polyester (SPE), in the form of sucrose octaesters and sucrose hexaesters of palmitic (16:0), stearic (18:0), oleic (18:1cis), and linoleic (18:2cis) acids, have many uses. Applications include: a non-caloric fat substitute, detoxification agent, and oral contrast agent for human abdominal (MRI) magnetic resonance imaging. However, it has been shown that the ingestion of SPE was shown to generate a depletion of physiologically important lipidic vitamins and other lipophilic molecules. In order to better understand, at the molecular level, the type of interaction between SPE and lipid membrane, we have, first synthesized different type of labelled and non-labelled SPEs. Secondly, we have studied the effect of SPEs on multilamellar dispersions of dielaidoylphosphatidylethanolamine (DEPE) and dipalmitoylphosphocholine (DPPC) as a function of temperature, SPE composition and concentration. The effects of SPEs were studied by differential scanning calorimetry (DSC), X-ray diffraction, 2H and 31P NMR spectroscopy. At low concentration (< 1 mol%) all of the SPEs lowered the bilayer to the inverted hexagonal phase transition temperature of DEPE and induced the formation of a cubic phase in a composition dependent manner. At the same low concentration, SPEs in DPPC induce the formation of a non-bilayer phase as seen by 31P NMR. Order parameter measurements of DPPC-d62/SPE mixtures show that the SPE effect on the DPPC monolayer thickness is dependent on the SPE, concentration, chains length and saturation level. At higher concentration (> or = 10 mol%) SPE are very potent DEPE bilayer to HII phase transition promoters, although at that concentration the SPE have lost the ability to form cubic phases. SPEs have profound effects on the phase behaviour of model membrane systems, and may be important to consider when developing current and potential industrial and medical applications. PMID- 11269938 TI - Efficient synthesis of the cholinephosphate phospholipid headgroup. AB - In search of an efficient method to prepare cholinephosphate headgroups in phospholipids under mild conditions (where the diacylglycerol moiety is not subject to oxidation), a method was developed for phosphorylation using a trialkyl phosphite and I2. The active intermediate is a phosphoryl iodide formed by oxidation of the phosphite with I2. 2-Bromoethanol, dimethyl chlorophosphite, and an alcohol (diglyceride) are converted to a phosphate triester in a one-pot reaction with high yield. In the second reaction, the phosphate triester is demethylated, and the ethyl bromide group is converted to choline by treatment with aqueous trimethylamine. This procedure is applied to the synthesis of hexadecylphosphocholine, and 1,2-didecanoyl-1-deoxy-1-thio-sn-glyceryo-3 phosphocholine. PMID- 11269939 TI - The lipid charge density at the bilayer surface modulates the effects of melittin on membranes. AB - The influence of melittin on two DMPA membrane systems at pH 4.2 and 8.2 has been investigated by solid-state 31P and 2H NMR, as a function of temperature and peptide concentration. Melittin promotes greater morphological changes for both systems in the fluid phase, the effect being larger at pH 4.2. Close inspection of fatty acyl chain dynamics suggests that some parallels can be drawn between the DMPA/melittin at pH 8.2 and PC/melittin systems. In addition, at pH 8.2 a direct neutralization at the interface of one of the lipid negative charges by a positive charge of the peptide occurs, as can be monitored by 31P NMR at the molecular level. For the system at pH 4.2 and at high temperature, a lipid-to peptide molar ratio of 30 is sufficient to transform the whole system into an isotropic phase, proposed to be inverted micelles. When the system is cooled down towards the gel phase one observes an intermediate hexagonal phase in a narrow range of temperature. PMID- 11269940 TI - Ceramide-epoxides. AB - Previously unknown 4,5-epoxy-N-acetyl-sphingosine (1) was synthesized by epoxidation of N-acetyl-sphingosine with 1,1-dimethyldioxirane. A by-product generated by HPLC purification is the tetrahydrofuryl derivative of acetamide (2). Mainly allylic oxidation was observed when natural ceramides were reacted with dimethyldioxirane. PMID- 11269941 TI - The human hair follicle contains two distinct K19 positive compartments in the outer root sheath: a unifying hypothesis for stem cell reservoir? AB - Up to now, the localization of stem cells in human anagen hair follicle relied on three complementary approaches; namely, detection of slow cycling cells, detection of high colony forming cells, and differential immunohistochemical staining. These techniques, however, gave conflicting results since stem cells were localized either as long label retaining cells in the so-called bulge area or as high colony forming cells in the lower third of the follicle. In the present study we investigated the expression of cytokeratin 19, a marker for putative stem cell-containing epithelial compartments, in order to characterize stem cell distribution in the human hair follicle throughout the hair cycle. We found that anagen human hair follicles contain two distinct reservoirs for stem cells located in the upper and lower thirds of the follicle. These two reservoirs fuse during the catagentelogen transition phase and individualize again in the newly forming anagen hair follicle. PMID- 11269942 TI - Identification of cadherin-11 down-regulation as a common response of astrocytoma cells to transforming growth factor-alpha. AB - Transforming growth factor-alpha (TGF-alpha) and its receptor are frequently co expressed in high-grade astrocytomas, suggesting a role for TGF-alpha autocrine/paracrine loops in the malignant progression of astrocytomas. To identify genes that may be critical in mediating TGF-alpha impact on the malignant progression of astrocytomas, we have used cDNA arrays to investigate TGF-alpha effects on the gene expression profile of U-373 MG glioblastoma cells. We found that in these cells approximately 50% of the TGF-alpha regulated genes code for cell motility/invasion-related proteins. TGF-alpha action on the expression of four of these proteins, alpha-catenin, IQGAP1, RhoA, and cadherin 11, was further investigated by immunoblotting in four astrocytoma cell lines and in normal astrocytes. The results demonstrate that the effects of TGF-alpha on IQGAP1, alpha-catenin, and RhoA expression are cell-line dependent. On the other hand, under TGF-alpha treatment, cadherin-11 expression is consistently decreased in all astrocytoma cell lines tested but is increased in normal astrocytes. In addition, we found that cadherin-11 is consistently down-regulated in astrocytomas versus normal brain tissues. Altogether, these results suggest that the down-regulation of cadherin-11 is a frequent molecular event in the neoplastic transformation of astrocytes and that this down-regulation may be initiated and/or amplified by TGF-alpha autocrine/paracrine loops during tumor progression. PMID- 11269943 TI - Maintenance of human islets in long-term culture. AB - The long-term maintenance of human islets in culture has remained a challenge. Despite advancements in culture techniques, human islets proved to have a short life span in vitro. For the first time, we have succeeded in maintaining human islets in a defined culture medium for more than 12 months. Freshly isolated islets from a 38-year-old donor were cultured in M3:5 medium and placed on a rocker for 14 days to remove contaminated exocrine and mesenchymal cells which attached to the bottom. The floating islets were purified by daily hand-picking and transfer into fresh medium. After 14 days, purified islets were allowed to attach to the bottom of the flasks and to expand. At various time points, islets were examined immunohistochemically and electron microscopically, and the secretion of islet hormones and their mRNA were determined by radioimmunoassay and reverse transcriptase polymerase chain reaction, respectively. Within seven days of culture, ductular and acinar cells developed within the initially normal islets. With time, exocrine cell types expanded while the number of the endocrine cells and their secretion decreased. At day 60, only a few endocrine cells were identifiable, whereas most of the cells appeared undifferentiated and expressed cytokeratin 7 and 19, neuron specific enolase, tomato lectin, phaseolus leucoagglutinin, laminin, and vimentin. After 60 days, the culture consisted entirely of undifferentiated cells which could be maintained in culture for 270 days before they became senescent. This is the first report on the long-term maintenance of human islet cells in culture and allows an insight into the complex process of endocrine cell differentiation. PMID- 11269944 TI - Upregulation of 9 genes, including that for thioredoxin, during epithelial differentiation of mesothelioma cells. AB - Malignant mesothelioma is a tumour originating from mesothelial cells, and it exhibits epithelial, fibrous, or biphasic differentiation. This tumour is highly resistant to therapy, and presence of a sarcomatous growth pattern has been associated with worse prognosis. A mesothelioma cell line with retained ability to differentiate into either epithelial or fibroblast-like phenotype was subjected to subtractive hybridisation in order to identify the genes coupled to tumour cell differentiation. Nine genes were found to be selectively overexpressed in the epithelial sub-line, compared to only two genes in the fibroblast-like phenotype. This may support the idea that the sarcomatous phenotype represents a less differentiated tumour. One of the genes that is differentially expressed by the epithelial cells was thioredoxin, a small redox active protein associated with cell-growth and differentiation. This overexpression was accompanied by increased protein levels both intracellularly and in the medium. Thioredoxin is reduced by the selenoprotein thioredoxin reductase and NADPH. The activity of this enzyme was high in both cell sub-lines but induced 2-fold in the epithelially-differentiated cells. Overexpression of thioredoxin might be a factor behind the poor prognosis and reduced responsiveness to therapy of mesotheliomas. Epithelial differentiation in this cell line has previously been linked to increased synthesis of heparan sulphate proteoglycans. The possible formation of complexes including thioredoxin, thioredoxin reductase, and heparan sulphate proteoglycans might play a role in the local control of cell growth and differentiation. PMID- 11269945 TI - Characterisation of a DNA sequence element that directs Dictyostelium stalk cell specific gene expression. AB - The ecmB gene of Dictyostelium is expressed at culmination both in the prestalk cells that enter the stalk tube and in ancillary stalk cell structures such as the basal disc. Stalk tube-specific expression is regulated by sequence elements within the cap-site proximal part of the promoter, the stalk tube (ST) promoter region. Dd-STATa, a member of the STAT transcription factor family, binds to elements present in the ST promoter-region and represses transcription prior to entry into the stalk tube. We have characterised an activatory DNA sequence element, that lies distal to the repressor elements and that is both necessary and sufficient for expression within the stalk tube. We have mapped this activator to a 28 nucleotide region (the 28-mer) within which we have identified a GA-containing sequence element that is required for efficient gene transcription. The Dd-STATa protein binds to the 28-mer in an in vitro binding assay, and binding is dependent upon the GA-containing sequence. However, the ecmB gene is expressed in a Dd-STATa null mutant, therefore Dd-STATa cannot be responsible for activating the 28-mer in vivo. Instead, we identified a distinct 28-mer binding activity in nuclear extracts from the Dd-STATa null mutant, the activity of this GA binding activity being largely masked in wild type extracts by the high affinity binding of the Dd-STATa protein. We suggest, that in addition to the long range repression exerted by binding to the two known repressor sites, Dd-STATa inhibits transcription by direct competition with this putative activator for binding to the GA sequence. PMID- 11269946 TI - Aberrant folate response and premature development in a mutant of Dictyostelium discoideum. AB - Growth and development are mutually exclusive in Dictyostelium discoideum. The transition between the two stages of the life cycle is regulated by the relative abundance of nutrients and proteins secreted by the cells which reflect population density. At the transition from growth to development, the discoidin genes--developmental markers--are induced by the "quorum" protein PSF. The effect of PSF is counteracted by food bacteria and by folate [8]. We show that folate treatment during growth delays morphologic development. Furthermore, we demonstrate that in a mutant of Dictyostelium discoideum (V188, renamed HBW3), which expresses discoidinI during growth and which develops rapidly [46], discoidinI expression is less sensitive to folate than in wild type cells. Finally, we present evidence that fragments of the discoidinI gamma promoter which are unresponsive to PSF and CM are sufficient for misregulation in the mutant. The only known regulator of these promoter elements is folate. Changes in the expression of other early developmental genes are also shown. Taken together, these data suggest that the reduced sensitivity to folate might be the cause for the "rapid development" phenotype of the mutant and that folate regulates developmental timing. PMID- 11269947 TI - Emerin expression at the early stages of myogenic differentiation. AB - Emerin is an ubiquitous protein localized at the nuclear membrane of most cell types including muscle cells. The protein is absent in most patients affected by the X-linked form of Emery-Dreifuss muscular dystrophy, a disease characterized by slowly progressive muscle wasting and weakness, early contractures of the elbows, Achilles tendons, and post-cervical muscles, and cardiomyopathy. Besides the nuclear localization, emerin cytoplasmic distribution has been suggested in several cell types. We studied the expression and the subcellular distribution of emerin in mouse cultured C2C12 myoblasts and in primary cultures of human myoblasts induced to differentiate or spontaneously differentiating in the culture medium. In differentiating myoblasts transiently transfected with a cDNA encoding the complete emerin sequence, the protein localized at the nuclear rim of all transfected cells and also in the cytoplasm of some myoblasts and myotubes. Cytoplasmic emerin was also observed in detergent-treated myotubes, as determined by electron microscopy observation. Both immunofluorescence and biochemical analysis showed, that upon differentiation of C2C12 cells, emerin expression was decreased in the resting myoblasts but the protein was highly represented in the developing myotubes at the early stage of cell fusion. Labeling with specific markers of myogenesis such as troponin-T and myogenin permitted the correlation of increased emerin expression with the onset of muscle differentiation. These data suggest a role for emerin during proliferation of activated satellite cells and at the early stages of differentiation. PMID- 11269948 TI - Embryonic, fetal, and neonatal tongue myoblasts exhibit molecular heterogeneity in vitro. AB - Variable gene expression patterns have been shown to exist between embryonic, fetal, and neonatal lineages of limb skeletal myoblasts in vitro and in vivo. In this study, we examined the molecular phenotype of embryonic, fetal, and neonatal tongue myoblasts in primary culture for comparison with in vivo developmental tongue myoblasts. Myogenic regulatory factor (MRF) and myosin heavy chain (MHC) gene expression were determined in culture during both growth and differentiation conditions by PCR, immunoblotting, and immunohistochemistry. Unlike their in vivo tongue myoblast equivalents, developmental tongue myoblast cultures featured the expression of MyoD when kept in growth conditions. Differentiation conditions in vitro induced myogenic tongue lineages to maintain characteristics of their in vivo morphologic and contractile gene phenotype. Both in vivo and in vitro, embryonic tongue lineages predominantly expressed MHC-embryonic isoforms, while fetal and neonatal tongue lineages predominantly expressed fast and perinatal isoforms of contractile genes. A notable difference from the in vivo condition that was observed in differentiated tongue myotubes in vitro was the presence of the MHC-slow protein. It was previously demonstrated that MHC-slow protein was undetectable during the in vivo development of the tongue musculature despite the abundance of slow isoform transcripts. The present characterization of primary tongue myogenic cultures indicates that murine myoblast heterogeneity exists primarily between developmental lineages at the level of contractile gene expression. Outside their native surroundings, developmental myogenic tongue populations are unable to recapitulate the determination and differentiation molecular profiles that occur in vivo. PMID- 11269949 TI - Larval-to-adult conversion of a myogenic system in the frog, Xenopus laevis, by larval-type myoblast-specific control of cell division, cell differentiation, and programmed cell death by triiodo-L-thyronine. AB - For the clarification of larval-to-adult muscle conversion, the authors established primary culture methods for adult- and larval-type myoblasts in the frog, Xenopus laevis, and examined the hormonal response in each case. The cell types were enzymatically dissociated from adult frog leg and tadpole tail muscles, respectively. The cells became attached to culture plates, proliferated, and fused with each other to form multinucleated myotubes within one week. Five significant differences between the two cell types were noted. (1) Adult cells showed greater proliferation activity than larval cells, the former increasing 5.5-fold over 6 days while the latter increase only 2.5-fold. (2) Differentiation (fusion) of larval type myoblasts started earlier. Cell fusion began on day 2 or 3 in larval cells and on day 4 in adult cells. (3) The metamorphic hormone, triiodo-L-thyronine (T3) decreased larval cell numbers to 56% of that of control cultures on day 7 but had no effect on adult cell number. DNA synthetic activity (3H-thymidine incorporation) in larval cells decreased under T3 (10(-8) M) to 45% of the control level on day 7. (4) Differentiation of adult myoblasts into myotubes was promoted by T3, whereas that of larval cells diminished by half. (5) Myotube death was induced by T3 specifically in larval but not in adult cultures. In addition to the myotube death, double staining with TUNEL (in situ DNA nick end labeling) and anti-desmin antibody indicated that T3 induces myoblast (desmin+ cell) death specifically in larval but not in adult cells. It is thus evident that the conversion of a larval-type myogenic system during metamorphosis becomes possible through nearly totally specific control of cell division, cell differentiation, and programmed cell death at a precursor cell level by T3. PMID- 11269950 TI - Generation of mononucleate cells from post-mitotic myotubes proceeds in the absence of cell cycle progression. AB - The remarkable regenerative ability of adult urodele amphibians depends in part of the plasticity of differentiated cells at the site of injury. Limb regeneration proceeds by formation of a mesenchymal growth zone or blastema under the wound epidermis at the end of the stump. Previous work has shown that when cultured post-mitotic newt myotubes are introduced into the blastema, they re enter the cell cycle and undergo conversion to mononucleate cells which divide and contribute to the regenerate [11, 13]. In order to investigate the interdependence of these two aspects of plasticity, we have blocked cell cycle progression of the myotubes either by X-irradiation or by transfection of the CDK4/6 inhibitor p16. In each case, the efficacy of the block was evaluated in culture after activation of S phase re-entry by serum stimulation. The experimental myotubes were implanted into limb blastemas along with a differentially labelled control population of myotubes containing an equivalent number of nuclei. X-irradiated myotubes gave rise to mononucleate cells in the limb blastema, and the progeny were blocked in respect of S phase entry. Comparable results were obtained with the p16-expressing myotubes. We conclude that progression through S or M phase is not required for generation of mononucleate cells and suggest that such cells may arise by budding from the muscle syncytium. PMID- 11269951 TI - Degradation of pentachlorophenol aqueous solutions using a continuous flow ultrasonic reactor: experimental performance and modelling. AB - The degradation of aqueous solutions of pentachlorophenol (PCP) in a three-stage sonochemical reactor operating in the continuous flow mode has been investigated. The experimental reactor may be considered as a series of three high-frequency ultrasonic units. The influence of several parameters such as ultrasonic power, reactor volume and volumetric feed flow rate on the reactor performance is reported. Application of classical basic chemical engineering principles leads to a model that enables us to predict the PCP concentration within the reactor. In steady state, experimental conversion rates are shown to be in good agreement with model predictions. PMID- 11269952 TI - A reaction kinetics model of water sonolysis in the presence of a spin-trap. AB - The time development of the concentration of a spin-trapped OH radical was studied by electron spin resonance at various sound intensities and various 5,5 dimethyl-1-pyrroline N-oxide (DMPO) concentrations in water sonolysis. The lifetime of the spin-trapped OH radical was also studied, and factors governing sonolysis are discussed. We found that the production of spin-trapped OH radical increases with increasing ultrasound intensity. The lifetime of a spin-trapped OH radical decreases linearly with increase in sonication time. This result suggests that an unknown scavenger is produced by ultrasound. Based on the above results, we suggested a model of the reaction kinetics and estimated the production rate of OH radical from this model. PMID- 11269953 TI - SINNMR studies of acoustically induced rotation of suspended particles. AB - Experiments and data analyses are reported on the 20 kHz acoustic manipulation of samples of trisodium phosphate dodecahydrate, of varying particle size, suspended in suitable support media. Data are obtained to expand the understanding and optimisation of the SINNMR (sonically induced narrowing of the nuclear magnetic resonance spectra of solids) technique, and use this to determine acoustically induced particle rotational correlation times. It is concluded that the average particle rotational correlation time decreases with increasing particle size and decreasing viscosity/density of the support media. Acoustic cavitation and accompanying interparticle collisions are shown to be important in the observation of SINNMR spectra. PMID- 11269955 TI - Influence of different factors on the output power transferred into medium by ultrasound. AB - The influence of several factors (amplitude of ultrasonic waves, external static pressure, temperature and viscosity of medium) acting, either individually or in combination, on the amount of power transferred to a liquid medium during ultrasonication (power output) was measured by calorimetry. At constant amplitude (150 microns) and pressure (200 kPa), the power output decreased as the temperature was raised. The effect of temperature could be compensated by increasing pressure. The magnitude of the increase in power output due to raising the pressure depended on the pressure range and the treatment temperature. At all temperatures and pressures studied, the power output increased exponentially when the amplitude was increased linearly. The magnitude of this power output did not depend on the temperature or pressure of treatment. At 40 degrees C the magnitude of the increase in power output due to increasing the pressure was not influenced by the amplitude of sonic waves. The power output increased as the viscosity of the medium was increased. The magnitude of this effect did not depend on the amplitude but on the static pressure. PMID- 11269954 TI - Homogeneous and heterogeneous asymmetric reactions: Part 11. Sonochemical enantioselective hydrogenation of trifluoromethyl ketones over platinum catalysts. AB - Enantioselective sonochemical hydrogenation of alpha,alpha,alpha-trifluoromethyl ketones, namely, 1,1,1-trifluoroacetophenone and 1,1,1-trifluoro-phenylacetone was carried out over various platinum catalysts modified by cinchonidine in different solvents. Both compounds yielded the (R)-alcohol as major product. The reaction rates and the enantiomeric excesses were determined over Pt/C, Pt/SiO2, Pt/K-10 and Pt/Al2O3 catalysts under conventional conditions. Since Pt/Al2O3 exhibited the best catalytic performance the effect of ultrasound on the catalytic activity and enantioselectivity was tested using this catalyst applying different sonochemical pretreatments. These methods included a pretreatment before the reaction in hydrogen and oxygen or both. The ultrasonic irradiation was found to be highly advantageous in the case of trifluoroacetophenone, whereas only moderate changes were observed using trifluoro-phenylacetone. After insonation of the catalyst, the enantioselectivity was considerably improved. Both the aerobic and anaerobic sonochemical pretreatments resulted in significant improvement in optical yield (up to 49% and 17% ee, at room temperature under 10 bar in 1,2-dichlorobenzene). In parallel, the hydrogenation rates increased to a small extent (1.1-1.2-fold increase) after hydrogenative ultrasonic pretreatment, whereas the rates decreased by a factor of 1.5-2 after aerobic insonation. To obtain more insight into the process, the effect of insonation time on the activity and enantioselectivity and actual changes of the catalyst were also studied. PMID- 11269956 TI - Comparison of classical and ultrasound-assisted extraction of polysaccharides from Salvia officinalis L. AB - After preparation of medicine tinctures from the herbal plant Salvia officinalis by classical and ultrasound-assisted extraction with aqueous ethanol, the insoluble plant residues were subsequently treated with hot water and dilute alkali to isolate polysaccharide cell wall components. The yields of the hot water extract as well as total extracted polysaccharides were higher in the case of the ultrasound-treated plant in both laboratory and pilot plant experiments. The water-extractable polysaccharide fractions, in all cases, contained glucose, galactose and arabinose as main sugar components, whereas the alkali-extractable fractions were rich in xylans. The fractions also contained considerable amounts of proteins. The water-soluble polysaccharides may contribute to the biological activity of the plant decoction. The results indicate that the addition of a subsequent extraction step during the preparation of the herbal tincture might contribute to a better exploitation of the raw material. PMID- 11269958 TI - Water stress responses of seedlings of four Mediterranean oak species. AB - Effects of water stress on phenology, growth, stomatal activity and water status were assessed from April to November 1996 in 2-year-old seedlings of Quercus frainetto Ten. (Quercus conferta Kit.), Quercus pubescens Willd., Quercus macrolepis Kotschy (Quercus aegilops auct.) and Quercus ilex L. growing in containers in northern Greece. All four species developed more than 50% of their total leaf area before the beginning of June--an adaptation to arid climates. Well-irrigated plants tended to develop greater individual leaf area, number of leaves per plant, total plant leaf area, height and root:shoot ratios than water stressed plants, but the difference between treatments was not significant for any parameter in any species. Quercus macrolepis appeared to be the most drought tolerant of the four species. It maintained the highest number of leaves of the smallest size and increased the proportion of fine roots during drought. In all species, drought caused significant decreases in stomatal conductance and predawn and midday water potentials from mid-July until the end of August, when the lowest soil water content and highest mean daily air temperatures and midday leaf temperatures occurred; however, the responses were species-specific. Among the four species, Quercus macrolepis sustained the highest stomatal conductance despite very low water potentials, thus overcoming drought by means of desiccation tolerance. Quercus ilex decreased stomatal conductance even before severe water stress occurred, thereby avoiding desication during drought. Quercus pubescens had the highest water potential despite a high stomatal conductance, indicating that its leaf water status was independent of stomatal activity. Quercus frainetto was the least drought-resistant of the four species. During drought it developed very low water potentials despite markedly reduced stomatal aperture. PMID- 11269959 TI - Dependence of winter water relations of mature high-elevation Picea engelmannii and Abies lasiocarpa on summer climate. AB - Water relations of Engelmann spruce (Picea engelmannii Parry) and subalpine fir (Abies lasiocarpa (Hook.) Nutt.) trees growing at an elevation of 3230 m on Mt. Evans, Colorado, USA, were studied during the winters of 1995-1996 and 1996-1997. During both winters, current-year and 1-year-old shoots were collected weekly and their relative water contents (RWC) determined. Measured meteorological parameters were used in a conifer winter water relations model, WINWAT, to simulate changes in shoot RWC of P. engelmannii and A. lasiocarpa during the winter. The model failed to predict shoot RWCs in 1996-1997 when calibrated with 1995-1996 data. The cold early summer of 1995 inhibited xylem formation, which appears to have caused lower rates of water recharge to the needles during the 1995-1996 winter than during the 1996-1997 winter. We conclude that summer climate strongly affects winter water relations in these subalpine species, and that changes in both summer and winter climate must be considered when predicting future ranges of these species. PMID- 11269957 TI - Effect of ultrasonic treatment on the molecular weight of carboxymethylated chitin-glucan complex from Aspergillus niger. AB - Different factors affecting the efficiency of the ultrasonic depolymerization of the high-molecular-weight carboxymethylated chitin-glucan prepared from the fungal mycelium of Aspergillus niger have been investigated. The influence of the following parameters was examined: concentration of the chitin-glucan complex, duration of ultrasonic irradiation, reaction temperature and volume of the ultrasonicated solution. The optimized conditions for the efficient ultrasonic depolymerization include: polysaccharide concentration--0.2 mg ml-1; volume of the sonicated solution--25 ml; duration of the sonication--10 min; and constant cooling of the sonicated sample in an ice-water bath. PMID- 11269960 TI - Changes in volatile terpene and diterpene resin acid composition of resistant and susceptible white spruce leaders exposed to simulated white pine weevil damage. AB - Induced (traumatic) resin in white spruce (Picea glauca (Moench) Voss) leaders resistant or susceptible to the white pine weevil (Pissodes strobi Peck) was analyzed for volatile terpenes and diterpene resin acids after simulated white pine weevil damage. Leaders from 331 trees were wounded just below the apical bud with a 1-mm diameter drill, coinciding with the natural time of weevil oviposition in the spring. Leaders were removed in the fall, and the bark and xylem from the upper and lower regions of the leader extracted and analyzed by gas chromatography. Unwounded trees had low amounts of resin in xylem compared with bark. In response to wounding, volatile terpenes and diterpene resin acids increased in the upper xylem (area of wounding), with resistant trees showing a greater increase than susceptible trees. Wounding caused monoterpenes in particular to decrease in the lower region of the leader (away from the drilled area) in greater amounts in susceptible trees than in resistant trees. In response to wounding, the proportion of monoterpene to resin acid increased in the upper and lower xylem of resistant trees, and slightly increased in the upper xylem of susceptible trees. Monoterpene-enriched resin is more fluid than constitutive resin, and probably flows more readily into oviposition cavities and larval mines, where it may kill immature weevils. Loss of resin components in the lower xylem suggested catabolism and transport of these materials to the site of wounding; however, energetic and regulatory data are necessary to confirm this hypothesis. This study provides a basis for measuring the ability of a tree to undergo traumatic resinosis that could be used to screen for resistance to white pine weevil. PMID- 11269961 TI - Relationship between carbohydrate concentration and root growth potential in coniferous seedlings from three climates during cold hardening and dehardening. AB - Greenhouse-cultured, container-grown seedlings of Aleppo pine (Pinus halepensis Mill.), radiata pine (Pinus radiata D. Don), and interior Douglas-fir (Pseudotsuga menziesii var. glauca (Beissn.) Franco) were cold acclimated and deacclimated in growth chambers over 24 weeks. Needle and root cold hardiness and root growth potential (RGP) were measured weekly. Root, needle and stem analyses for soluble sugars and starch were performed biweekly. In all tissues, there was a close correspondence between cold hardiness and the absolute concentration of soluble sugars, as well as between the increase and decrease in concentration of soluble sugars during cold hardening and dehardening, respectively, supporting the theory that soluble sugars function as cryoprotectants in plant tissues. The magnitude of starch concentration did not parallel the magnitude of the cold hardiness attained, and changes in starch concentration were related to production and consumption factors, rather than timing of changes in cold hardiness. The rise and fall of RGP paralleled the rise and fall of total carbohydrate concentration in roots. The behavior of the three species was surprisingly similar, considering the different climates to which they are adapted. PMID- 11269962 TI - Foliar nutrient retranslocation in Eucalyptus globulus. AB - We measured patterns of change in concentrations and contents of nitrogen, phosphorus, potassium, magnesium and calcium in fully expanded leaves of young Eucalyptus globulus (Labill.) trees growing in a plantation in southeastern Australia, over a 12-month period beginning at the onset of spring. There was significant net retranslocation of mobile nutrients on a seasonal basis from green leaves, coinciding with continued growth and production of foliage. There was a close positive relationship between initial nutrient content (N, P and K) of the leaf and amount retranslocated, and a tight coupling between N and P retranslocated from leaves. Net retranslocation was significantly correlated with basal area growth increments. Artificial shading of leaves resulted in senescence and reduction in leaf mass. It also induced retranslocation of N, P and K from leaves of different ages and from different position in the canopy. Although the mechanisms underlying the effects of shading intensity on these changes were not elucidated, shading provided an experimental tool for studying retranslocation. Comparison of the results with published data for Pinus radiata (D. Don) grown in the same environment indicated a similarity between the species in patterns of change in foliar nutrient contents and in factors governing foliar nutrient retranslocation, giving rise to unifying principles. PMID- 11269963 TI - Differentiation among effects of nitrogen fertilization treatments on conifer seedlings by foliar reflectance: a comparison of methods. AB - Analysis of reflectance can be used to estimate foliar concentrations of photosynthetic pigments, thus providing information on the physiological status of green plants. We compared several methods of reflectance analysis for the capacity to differentiate among effects of fertilization treatments across different irradiances on seedlings of Engelmann spruce (Picea engelmanii Parry ex Engelm.). Seedlings were grown in two light treatments (0 and 60% shade) and three nitrogen (N) treatments (10, 25 and 100 mg N l-1) for one growing season, after which foliar reflectance of the needles was measured. Five indices were tested: R550 (% reflectance at 550 nm); red edge position; the ratio R698:R760; the structure independent pigment index (SIPI); and the photochemical reflectance index (PRI). Both the light and nutrient treatments significantly affected foliar chlorophyll a and b and carotenoid concentrations. Among the indices tested, R550, red edge position and R698:R760 ratio were related to chlorophyll concentration, and were significantly affected by both light and N treatments. Both SIPI and PRI were related to chlorophyll and carotenoid concentrations. Among these relationships, PRI was affected by both treatments, whereas SIPI was sensitive to N treatment but not to light treatment. All five indices were weakly but significantly correlated with growth as measured by dry weight. PMID- 11269964 TI - Stable carbon isotope discrimination: an indicator of cumulative salinity and boron stress in Eucalyptus camaldulensis. AB - Saplings of Eucalyptus camaldulensis Dehn. Clone 4544, irrigated with water of differing salinities (2 to 28 dS m-1) and boron concentrations (1 to 30 mg l-1), integrated the history of these stresses through the discrimination of stable isotopes of carbon in leaf and woody tissues. Carbon isotope discrimination (delta) was reduced primarily by salinity. Decreases in discrimination in response to boron stress were detected in the absence of salinity stress, but the decreases were significant only in leaf tissues with visible boron injury. Sapwood core samples indicated that salinity- and boron-induced reductions in delta increased with increasing tree age. Absolute values of delta varied with location of leaf or wood tissue, but relative effects of salinity on the relationship between delta and transpiration efficiency (W) were similar. In response to increasing salinity stress, relative decreases in delta paralleled relative decreases in biomass and both indices yielded similar salt tolerance model parameters. The strong correlations between delta, tree fresh weight, leaf area and W suggest that delta is a useful parameter for evaluating salt tolerance of eucalyptus PMID- 11269966 TI - Day at the capitol. Arkansas physicians spend time lobbying local legislators. PMID- 11269965 TI - Effects of serpentine soil factors on Virginia pine (Pinus virginiana) seedlings. AB - Effects of simulated serpentine soil conditions (elevated Mg:Ca ratio and Ni concentration) on seedlings from populations of Virginia pine (Pinus virginiana Mill.) from serpentine and non-serpentine sites were evaluated in sand culture. We determined (1) how seedlings are affected by elevated Mg:Ca ratio and Ni concentrations, (2) if there are interactive effects between Mg:Ca ratio and Ni concentrations on seedling growth, needle pigment concentrations, and nutrition, and (3) if Virginia pine populations from serpentine areas are edaphic ecotypes. A Mg:Ca ratio of 5 and 50 microM Ni both reduced seedling growth compared with control seedlings grown in the presence of the standard Mg:Ca ratio of 0.5 and no Ni. Interactive effects between Mg:Ca ratio and Ni concentrations were highly significant for growth, foliar pigments, and needle and root elemental concentrations. Nickel-mediated reductions in growth and foliar pigment concentrations were less at the serpentine Mg:Ca ratio of 5 than at the standard (non-serpentine) Mg:Ca ratio of 0.5. Foliar N was reduced by Ni concentrations as low as 10 microM, and foliar and root K, Ca and P concentrations were significantly reduced by Ni concentrations above 25 microM, with greater reductions at a Mg:Ca ratio of 0.5 than at a Mg:Ca ratio of 5. There were no population x serpentine soil factor interactions for seedling growth, foliar pigment concentrations, or nutrition, suggesting that seedlings from trees growing on serpentine soils are not edaphic ecotypes. We conclude that serpentine conditions present at the site of seed collection have not resulted in the selection of edaphic ecotypes of Virginia pine with respect to Mg:Ca ratio and Ni concentration. PMID- 11269967 TI - Thoracic aortic aneurysm revisited. PMID- 11269968 TI - Use of diffusion-weighted images. PMID- 11269969 TI - Arkansas Patient Safety Initiative. PMID- 11269971 TI - Epidemiology in the post-genomic era. PMID- 11269970 TI - Pediatric injuries resulting from use of all-terrain vehicles. AB - Annually, 20,000 children are injured while operating all-terrain vehicles (ATVs). The purpose of this paper was to review child-ATV injuries in Arkansas and identify any areas in need of further investigation. An analysis of emergency medical-service transports was done for children 0-19 years who had ATV-related injuries in Arkansas from 1998 to 1999. Prehospital-reported child-ATV emergencies were identified, separated by county, and emergency encounter rates were calculated. Our results indicate that emergency medical services (EMS) transported 319 children in Arkansas from 1998 to 1999. ATV injury information is limited in Arkansas, but available data indicate high injury rates existed for many rural counties. PMID- 11269972 TI - Evaluation of technetium-99m ciprofloxacin (Infecton) in the imaging of infection. AB - INTRODUCTION: The aim of this study was to evaluate the usefulness of technetium 99m (Tc-99m) ciprofloxacin in imaging inflammation/infection. The ciprofloxacin for labelling, as a kit, was obtained from St Bartholomew's Hospital in London. MATERIALS AND METHODS: Patients were injected intravenously with Tc-99m ciprofloxacin and imaging was done at 10 minutes, 4 hours and 24 hours if necessary. Tomographic images (SPECT) were obtained in a few patients. Ninety-six patients were studied using Tc-99m ciprofloxacin. Forty-eight patients had bone scans and 22 had Tc-99m IgG scans. Eight patients were imaged using Tc-99m HMPAO labelled white blood cell, and bacteriological culture results were available in 24 patients. Organisms cultured included Acinetobacter baumanii, Streptococcus, Staphylococcus aureus, Pseudomonas, Klebsiella, Blastococidia, Methicillin resistant S. aureus, Salmonella and Candida. RESULTS: Findings were evaluated against microbiology, alternative imaging modalities and clinical outcome. There were 47 true positives, 33 true negatives, 5 false positives and 11 false negatives, giving a sensitivity of 81% and specificity of 87%. The positive and negative predictive values were 90% and 75%, respectively. There were no side effects and the scan was particularly useful in the evaluation of painful joint prosthesis to exclude infection. Repeat studies on 8 patients given antibiotics over a long period were very useful in deciding on termination of the antibiotic treatment. PMID- 11269974 TI - Correlation of baseline quantitative plasma human immunodeficiency (HIV) type 1 RNA viral load with clinical status and CD4+ T-cell counts in treatment-naive HIV positive patients in Singapore. AB - INTRODUCTION: Quantitative measurement of plasma HIV-1 RNA viral load has been available in Singapore since 30 November 1996. This study investigates the relationship, in antiretroviral-naive, HIV-positive Singapore residents, between the baseline HIV-1 RNA viral load and clinical status at the time of quantification. The association of HIV-1 RNA viral load with CD4+ T-cell counts was also studied. MATERIALS AND METHODS: HIV-1 RNA viral load was determined using Amplicor HIV-1 Monitor Test. One hundred and eighty subjects had baseline plasma HIV-1 RNA levels quantified during the period 30 November 1996 to 27 July 1998. They were classified into three clinical groups: A for asymptomatic infection (n = 110), B for symptomatic infection (n = 29) and C for AIDS (n = 41). RESULTS: The differences between mean HIV-1 RNA levels were statistically significant (P < 0.001) for groups A and B (mean difference = -0.61 log10), and for groups A and C (mean difference = -0.75 log10). However, there was no statistically significant difference (P = 0.68) between groups B and C (mean difference = -0.13 log10). Of those subjects with CD4+ T-cell counts measured within 30 days of viral load quantification, there were statistically significant negative correlations between HIV-1 viral load and CD4+ T-cell counts for groups A (n = 91, r = -0.536, P < 0.01) and C (n = 34, r = -0.446, P < 0.01) but not group B (n = 26, r = -0.297, P > 0.05). CONCLUSION: This suggest that the more advanced the phase of HIV infection, the higher the baseline plasma viral load and the lower the CD4+ T-lymphocyte counts. PMID- 11269975 TI - Granuloma annulare: a review of 41 cases at the National Skin Centre. AB - INTRODUCTION: Granuloma annulare (GA) is a benign, inflammatory skin disorder that has no proven aetiology or widely accepted theory of pathogenesis. We present the results of a local study examining the characteristics of this disorder. MATERIALS AND METHODS: This is a retrospective study of 41 patients with granuloma annulare seen at a tertiary dermatological skin hospital in Singapore. The disease was classified based on whether it was localised or generalised. Remission rates and associated diseases were documented. RESULTS: The majority (93.9%) of lesions were localised. GA tends to occur within the first three decades of life, and 39.4% of patients were 10 years old and below. There was a slight female preponderance, and no predilection for any particular racial group. Remissions were seen in 67.6% of patients who were followed up for at least 12 months, and remissions only occurred with the localised form of GA. There were 4 cases of disseminated GA and 1 case of perforating GA. There were 6 patients with associated diabetes mellitus, 2 with pulmonary tuberculosis, and 4 patients each with hypercholesterolaemia and a history of atopy. CONCLUSION: The characteristics of GA in our local population do not differ significantly from that of overseas studies. While localised forms of GA have a high chance of resolving, the generalised forms of the disease are difficult to treat and do not tend to resolve totally. PMID- 11269973 TI - Antineutrophil cytoplasmic antibodies (ANCAs) in patients with inflammatory bowel disease show no correlation with proteinase 3, lactoferrin, myeloperoxidase, elastase, cathepsin G and lysozyme: a Singapore study. AB - INTRODUCTION: The pathogenic importance of antineutrophil cytoplasmic antibodies (ANCAs) in inflammatory bowel disease (IBD) is unclear and target antigen localisation studies may lend insight to the specific pathogenic mechanisms of IBD. In this pilot study, we looked at occurrence of ANCA in Asian IBD patients. In ANCA-positive samples, we analysed for the presence of target antigens i.e. proteinase 3, lactoferrin, myeloperoxidase, elastase, cathepsin G and lysozyme. MATERIALS AND METHODS: This prospective study was carried out from July 1997 to February 1998. Sera were screened for ANCAs with indirect immunofluorescent test and tested with an enzyme immunoassay (ELISA) kit which provides a semi quantitative assay for human IgG autoantibodies against 6 antigens: proteinase 3, lactoferrin, myeloperoxidase, elastase, cathepsin G and lysozyme. RESULTS: A total of 75 patients were studied: 50 with IBD and 25 controls with functional bowel disease. Ten had Crohn's disease (CD) and 40 had ulcerative colitis (UC). There was no racial predilection among the Chinese, Malays or Indians. In CD, 1 was positive for cytoplasmic ANCA (cANCA) and 2 for perinuclear ANCA (pANCA). In UC, 4 were positive for pANCA, 15 for atypical perinuclear ANCA (apANCA) and 1 for cANCA. In the CD and UC population, the proportion positive for ANCA was 30% and 50%, respectively. There was no ANCA detected among the controls. Of those ANCA-positive IBD patients (n:23), only 1 demonstrated anti-myeloperoxidase antibodies. No antibodies were detected against the other 5 antigens tested. CONCLUSIONS: This pilot Singapore study concludes that there is no significant ANCA association with proteinase 3, lactoferrin, myeloperoxidase, elastase, cathepsin G and lysozyme. PMID- 11269976 TI - Transaminitis in Duchenne's muscular dystrophy. AB - BACKGROUND: Persistently raised transaminase levels often prompt the clinician to investigate for liver pathology. Previously, some of our patients with Duchenne's muscular dystrophy have had investigations to look for liver disorders when the alanine transaminases (ALTs) were found incidentally to be high. AIM: The objective of the study was to ascertain the levels of the transaminases in patients with Duchenne's muscular dystrophy and to see if the levels of transaminases correlated with muscle enzymes such as creatine kinase (CK). MATERIALS AND METHODS: This is a case series of 19 patients with Duchenne's muscular dystrophy. Alanine and aspartate transaminase (AST) levels and CK levels were measured in the serum of the patients. RESULTS: In this series, ALT and AST levels were all found to be raised significantly in patients with biopsy-proven Duchenne's muscular dystrophy and Becker's muscular dystrophy. Alanine transaminase, in particular, was raised to a mean of 356 mmol/L, 9 times above the mean for normal. There was also good correlation between ALT and CK levels with a correlation coefficient of r = 0.80 and P value 0.01. Similar to CK, the transaminase levels were inversely proportional to mobility and to age. CONCLUSION: The presence of hypertransaminasemia in patients with muscular dystrophy should be attributed to muscle breakdown rather than to liver pathology and such patients should not be over-investigated for liver disease. In the absence of liver pathology, raised transaminases may be an early sign of occult muscular dystrophy and such patients should have CK levels checked to look for evidence of muscular dystrophy. PMID- 11269977 TI - Extended resection of locally advanced (T4) stomach cancer. AB - INTRODUCTION: The surgical management of locally advanced (T4) stomach cancer remains controversial and many still question the benefits of an extended resection. The aims of this study were to examine the morbidity and mortality associated with extended resection and also to determine the survival benefit. MATERIALS AND METHODS: A retrospective review of all stomach cancer operations performed from 1989 to 1998 was carried out and the relevant case histories retrieved and analysed. RESULTS: Out of the 980 stomach cancer operations performed, 784 (80%) had tumour resection and 20 (2.5%) also had extended resection of adjacent involved organs. These 20 patients had a 10% postoperative morbidity and the operative mortality was 15%. The survival ranged from 2 to 35 months (median 17 months). CONCLUSIONS: Extended resection of T4 stomach cancer is feasible and carries an acceptable operative morbidity and mortality. However, there is poor survival benefit and it should only be performed in a selected group of patients. PMID- 11269978 TI - Transurethral alprostadil for the treatment of erectile dysfunction: results of a multicentre trial. AB - INTRODUCTION: The incidence of erectile dysfunction (ED) has been shown to increase significantly in Singaporean male population. Thus, this prospective study (no controls) assessed the clinical efficacy and safety of a medicated urethral system for erection (MUSE) in Singaporean male patients with a known history of ED. SUBJECTS AND METHOD: Eighty-six men with a mean age of 55.7 years with differential causes of ED were administered with MUSE in the clinic with a titration adjustment of 4 possible dose regimes (125, 250, 500 and 1000 mcg) till efficacy is achieved. Subsequently, patients were subjected to home treatment for a duration of 3 months. RESULTS: Sixty per cent of inclinic patients exhibited good responses and were given MUSE for home treatment. The efficacy rate for home treatment was 86%. Overall, the patients (n = 86) had a 51.2% success rate in achieving satisfactory sexual intercourse. Diabetic and psychogenic patients were noted to respond well to MUSE. No severe adverse events were found in this study. CONCLUSION: MUSE showed to be a safe, less invasive, well-tolerated and efficient alternative treatment for ED in Singaporean men. PMID- 11269979 TI - Initial experience with an autocapture pacemaker system. AB - INTRODUCTION: Autocapture management aims to extend pacemaker longevity without compromising on patient safety by automatically monitoring the pacing threshold and adjusting the pacemaker output for consistent capture. This paper describes our initial experience with the Pacesetter Regency pacemaker with autocapture management. MATERIALS AND METHODS: Nineteen patients were implanted with single chamber pacemakers with autocapture management. Autocapture was programmed "ON" the day after implantation if Evoked Response (ER) amplitude was at least 2.8 mV. The patients were followed up at 2 weeks, 2 months and 6 months. At each visit, pacing threshold and lead impedance were measured. Autocapture was turned "ON" during follow-up if it had not been done previously. RESULTS: In 16 out of 19 patients, autocapture could be turned "ON" the day after implantation. One patient had an ER signal that was less than 2.8 mV and 2 patients were in fast atrial fibrillation of more than 120 beats per minute which precluded ER signal testing. These patients could not have autocapture programmed "ON". CONCLUSION: The benefits of autocapture management can only be realised if an ER signal of at least 2.8 mV is obtained. This requires intraoperative testing of the ER signal. Since there is no commercially available pacing system analyser presently that can measure this, modification of the standard implantation procedure with some prolongation of procedure time is needed. PMID- 11269980 TI - Minimally invasive repair of atrial septal defects--a case series. AB - INTRODUCTION: Repair of atrial septal defects in adults has become so safe that emphasis is now placed on improving cosmesis without compromising safety. The results of our series of minimally invasive repair of atrial septal defects done through a lower partial sternotomy are reported. MATERIALS AND METHODS: Eight adult patients underwent minimally invasive repairs of atrial septal defects over a period of 1 year (Group 1). Conventional repairs of atrial septal defects were performed in 26 patients over the same period (Group 2). We describe the technique and compare our results on time taken for surgery, bypass time, cross clamp time, time to extubation and total hospital stay. Early complications and complications at 1 year are also presented. RESULTS: No difference in the demographic data was found between the two groups. The time taken for surgery was more, while cross clamp times and bypass times were shorter in Group 1. There was also no difference in the amount of blood loss, early postoperative complications and later complications like cardiac rhythm or wound pain. CONCLUSION: The improved cosmetic appearance, both in males and females, is an important factor to recommend minimally invasive repair of atrial septal defects in an adult population. Surgical closure of atrial septal defects can be performed through a lower partial sternotomy without compromising access or safety with good cosmetic outcome. PMID- 11269981 TI - Cross-sectional study of near-work and myopia in kindergarten children in Singapore. AB - INTRODUCTION: In view of the high and increasing myopia rates amongst young Singaporean children, we aimed to assess the relationship between near-work and myopia in 414 pre-school children aged 4 to 6 years. MATERIALS AND METHODS: We measured near-work indices such as tuition classes outside school and other possible risk factors via a questionnaire. We then measured myopia with a hand held autorefractor. RESULTS: Children who had 3 or more hours per week of near work classes outside school had a higher rate [odds ratio 1.39 (95% confidence interval 1.02, 2.53)] of myopia. CONCLUSIONS: This suggests that there may be an association between near-work and myopia, even at such a young age. Given the increasing emphasis on near-work in Singapore, it may be important to call for increased visual health awareness, although further studies will be needed to establish if near-work causes myopia. PMID- 11269982 TI - Diabetic retinopathy. AB - INTRODUCTION: To provide a review of the current standard of care in diabetic eye management. METHODS: A non-systematic evidence-based review utilising available data and consensus statements on the subject matter. RESULTS: Diabetes mellitus affects some 9.0% of Singaporeans, and more than 60% of patients with diabetes in this population remained undiagnosed. Diabetic retinopathy is an important complication among diabetic patients and adverse visual outcome associated with this condition can be reduced by more than 95% by taking measures including good glycaemic control, timely and appropriate laser therapy, and vitrectomy surgery when indicated. An important aspect of management is the accurate disease classification which is essential for prognostication, appropriate follow-up schedule, and timing of therapeutic intervention purposes. CONCLUSION: Diabetic retinopathy will remain a significant problem if the current trend in diabetes among Singapore residents prevails. As the severe visual impairment associated with diabetic retinopathy can be largely prevented with appropriate and timely intervention, the major challenge to the health care providers today is the identification and education of patients with diabetes, and the enrollment of these patients in a life-long comprehensive ophthalmic management programme in order to minimise visual morbidity. PMID- 11269984 TI - Squamous cell carcinoma arising in a cutaneous epidermal cyst--a case report. AB - INTRODUCTION: Cutaneous epidermal cysts are common lesions, but fortunately, malignant transformation of their epithelium is rare. There are only a few cases mentioned in recent literature, mostly occurring in epidermal cysts in the head and neck region. CLINICAL PICTURE: We present a case of malignant transformation in a long-standing buttock epidermal cyst, presenting a month after trauma to the cyst. TREATMENT: An excision biopsy and rotation flap was done. When the pathology was confirmed, a second resection was carried out and the defect covered with a free latissimus dorsi flap, a buttock rotation flap and a posterior thigh rotation flap. CONCLUSION: Although malignant transformation is rare, this case illustrates the importance of routine histology in excision of epidermal cysts. PMID- 11269983 TI - Otogenic lateral sinus thrombosis--a case report. AB - INTRODUCTION: We portray and discuss a case of lateral sinus thrombosis following acute otitis media and mastoiditis. CLINICAL PICTURE: The patient presented with otorrhoea, otalgia, neck pain, fever and chills. TREATMENT: Cortical mastoidectomy was performed. Intravenous antibiotics and heparin were administered. OUTCOME: The patient had a complete recovery with no sequelae. CONCLUSIONS: Neurotologic complications of suppurative otitis media like meningitis, cerebral abscess, extradural abscess and dural sinus thrombosis are rare in the antibiotic era. Hence, doctors today have to maintain extra vigilance and a high index of suspicion for such complications. PMID- 11269985 TI - Primitive neuroectodermal tumour of the chest wall--a report of two cases and review of literature. AB - INTRODUCTION: Primitive neuroectodermal tumours (PNETs) of the chest wall are rare entities and little is known regarding its biological activity and prognostic factors. Two cases are reported and the available literature reviewed to highlight the presentation and management of these tumours. CLINICAL FEATURES: We report 2 patients who were diagnosed with PNET of the chest wall in our centre. As there are no clinical features or basic diagnostic measures which are characteristic of these tumours, diagnosis is based on special tests. With the advent of newer immunohistochemical methods, it is now diagnosed more confidently. TREATMENT: Both patients received multidisciplinary modalities of treatment, comprising extensive surgical resection, chemotherapy and radiotherapy. OUTCOME: One patient succumbed to the disease one year after diagnosis and the other is currently disease-free, both clinically and radiologically at 24 months. CONCLUSION: Despite multidisciplinary modalities of treatment, the prognosis of PNET is still generally poor. Early diagnosis and treatment are important to improve the chances of survival. PMID- 11269986 TI - The use of the laryngeal mask airway in post-tonsillectomy haemorrhage--a case report. AB - INTRODUCTION: The use of the laryngeal mask airway in elective adenotonsillectomy has been well described. However, there is no literature to support its use in post-tonsillectomy haemorrhage. CLINICAL PICTURE: We report a case of a patient who presented with primary post-tonsillectomy haemorrhage, which required general anaesthesia for haemostasis after undergoing bilateral functional endoscopic sinus surgery, uvulopalatopharyngoplasty and tonsillectomy. TREATMENT AND OUTCOME: The laryngeal mask airway was used successfully after an initial attempt at endotracheal intubation had failed. There were no complications. CONCLUSIONS: The laryngeal mask airway can be used to secure the airway for haemostasis for post-tonsillectomy haemorrhage if intubation is not possible. PMID- 11269987 TI - Beneficial effect of combination therapy with ozagrel and pranlukast in exercise induced asthma demonstrated by krypton-81 m ventilation scintigraphy--a case report. AB - INTRODUCTION: We evaluated the effect of combination therapy with thromboxane A2 synthesis inhibitor and leukotriene receptor antagonist in a patient with exercise-induced asthma using krypton-81 m ventilation scintigraphy. CLINICAL PICTURE: In a patient with exercise-induced asthma, we found exercise-induced abnormalities of respiratory function test and ventilation scintigraphy, and increases in plasma concentrations of thromboxane B2 and leukotriene C4 with exercise. TREATMENT: A thromboxane A2 synthesis inhibitor (ozagrel) and a leukotriene receptor antagonist (pranlukast) were prescribed. OUTCOME: After treatment for 2 weeks, abnormalities of respiratory function test and ventilation scintigraphy improved. CONCLUSIONS: The combination therapy with ozagrel and pranlukast might be useful for the relief of symptoms in patients with exercise induced asthma, and krypton-81 m ventilation scintigraphy could be a useful tool for visible evaluation of treatment. PMID- 11269988 TI - A case report: persistent acantholytic dermatosis in chronic renal failure. AB - INTRODUCTION: This is the second case of persistent acantholytic dermatosis in a patient with chronic renal failure, seen at the same institution in 2 years. CLINICAL PICTURE: A 70-year-old Chinese man with end-stage renal failure on continuous ambulatory peritoneal dialysis for 6 months presented with pruritic rashes over the scalp and chest for 3 months. Histologically, the lesions resembled Darier's disease. Differential diagnoses include Darier's disease, Grover's disease and perforating dermatosis in chronic renal failure. TREATMENT: The patient was treated with anti-histamines, topical steroids and emollients. OUTCOME: Resolution of pruritus was documented. However, the extensive hyperkeratotic papules remained persistent. CONCLUSION: A chronic non-remitting course is to be expected for this dermatoses in which the aetiology is unknown. PMID- 11269989 TI - Retroperitoneal Castleman's disease in the perinephric space--imaging appearance: a case report and a review of the literature. AB - INTRODUCTION: Castleman's disease (CD) is a rare lymphoid tumour usually found in the mediastinum. Extrathoracic sites are uncommon. Its radiological findings may be similar to other retroperitoneal tumours, making diagnosis difficult. CLINICAL PICTURE: A 54-year-old female was found to have an incidental hypoechoic mass in the left posterior perinephric space on routine ultrasound. Abdominal computed tomography (CT) scan demonstrated an isodense mass which enhanced brightly with intravenous contrast. Angiogram confirmed a hypervascular mass. TREATMENT: The retroperitoneal mass was excised. OUTCOME: Histology revealed CD of hyaline vascular type. CONCLUSION: CD should be considered in the differential diagnosis of a retroperitoneal mass, which demonstrates homogeneous and intense enhancement. PMID- 11269990 TI - Perioperative wheezing--a report of three cases. AB - CLINICAL PICTURE: We describe 3 patients with perioperative wheezing, 2 of whom were treated with bronchodilators on the presumptive diagnosis of bronchospasm. TREATMENT AND OUTCOME: Subsequent clinical improvement occurred when it was recognised that the wheezing was due to upper airway obstruction and stridor, not bronchospasm and rhonchi, and appropriate treatment instituted. PMID- 11269991 TI - An unusual tumour metastasis to the cervix. AB - INTRODUCTION: The vast majority of tumours in the cervix are either primary carcinomas or direct extension of primary tumours from nearby sites such as the endometrium, myometrium, rectum and bladder. Patients usually present with abnormal bleeding, pain and dyspareunia. A smaller number of patients are asymptomatic and their tumours can be diagnosed early by PAP smears. CLINICAL PICTURE: We present an unusual case of an elderly lady with breast cancer that had metastasized to the cervix 4 years after primary treatment of the breast malignancy. Although the appearance of the cervix was "normal looking", it was firm and indurated on palpation. PATHOLOGY: A definite diagnosis of metastatic infiltration could only be made by colposcopic biopsy. Early PAP smears had shown some abnormal cells suggestive of metastatic lobular carcinoma but were not conclusive of metastasis from breast carcinoma. Subsequent bone scans and CT scans of the abdomen revealed metastatic lesions in the skull, spine, left femoral shaft and liver. PMID- 11269992 TI - Echocardiographic features and management of neonatal ductal aneurysm. AB - OBJECTIVES: To determine the case incidence of ductus arterious aneurysm among our neonates, and to report on our experience in the presentation, echocardiographic features, management and outcome of this condition. METHODS: Retrospective review of cases diagnosed within a 1-year period from 1 July 1998 to 30 June 1999. RESULTS: Eight cases of neonatal ductus arteriosus aneurysm (DAA) were diagnosed from 1 July 1998 to 30 June 1999. There were 998 new neonatal echocardiograms done during this period, giving us a case incidence of 0.8 per 100 echocardiograms. Only 1 patient was diagnosed antenatally, all others were detected incidentally on echocardiography done for other indications. None had symptoms related directly to the DAA and there was no suggestive mediastinal mass on chest X-ray. Majority were term infants (88%) and there was a predominance of male infants (75%). Three were associated with patent ductus arteriosus (PDA), while in the others, the ductus arteriosus were non-patent at first echocardiography. In the 3 infants whose ductus arteriosus was patent, the PDA was inserted into the pulmonary artery from an unusually superior direction. Close serial echocardiography on some of our patients suggested that resolution of the aneurysm is by thrombosis, manifesting as an echogenic area, followed by regression. In our series, 7 had total resolution, while 1 patient had a persistent echogenic area which became progressively smaller. CONCLUSIONS: We found ductus arteriosus aneurysms in 0.8% of our neonatal echocardiograms. An unusually superior insertion of PDA into the pulmonary artery is a marker for its presence. Asymptomatic aneurysms resolve spontaneously and should be managed expectantly. Thrombosis plays a part in the resolution of ductal aneurysm. PMID- 11269993 TI - Predisposing factors and gross examination findings in periodontal disease. AB - Periodontal disease is the most frequently encountered condition by small animal practitioners. A complete oral examination including visual examination, periodontal probing, and intraoral radiographs are necessary to determine the grades of disease present, and the predisposing factors contributing to it. PMID- 11269994 TI - Complicated periodontal disease. AB - Not all patients experience "normal" levels of periodontal disease. More complicated cases with deep periodontal pockets or excessive inflammation require specialized procedures and therapy. PMID- 11269995 TI - How to obtain and interpret periodontal radiographs in dogs. AB - Oral radiography plays an important role in the diagnosis of periodontal disease. The diagnostic yield of radiographs is high, and they should be obtained in all cases presenting for periodontal treatment and to assess the long-term success of therapy. Diagnostic-quality radiographs for evaluating the periodontium are best obtained with a dental x-ray unit and with the patient under general anesthesia. The standard full-mouth radiographic survey contains a minimum of 6 views, and with practice, can be obtained with minimal effort and time. Interpretation of dental radiographs, however, requires a keen understanding of the normal radiographic anatomy of a tooth and its supporting structures. The radiographic diagnosis of periodontal disease is characterized by rounding of the alveolar crest with loss in continuity of the lamina dura, widening of the periodontal ligament space, and loss of alveolar crest height. PMID- 11269996 TI - Treatment planning based on examination results. AB - Various methods of assessment, from visual to tactile to radiographic are used to accurately evaluate oral and dental problems. From this evaluation, the appropriate therapy can be determined. PMID- 11269998 TI - Home care: products and techniques. AB - Periodontal disease is common in dogs and cats. Prevention and treatment is important for general health and well-being of our pets. Both prevention and treatment of periodontal disease have two components, namely maintenance of oral hygiene and professional periodontal therapy. Maintenance of oral hygiene is performed by the owner and is, therefore, also called home care. The preventions and long-term control of periodontal disease requires adequate home care. This chapter details home care techniques and available products. PMID- 11269997 TI - Nonsurgical periodontal therapy. AB - The primary etiology of periodontal disease is bacterial infection. Bacteria exist as a biofilm (plaque) on the tooth and soft-tissue surfaces of the mouth. Biofilm is extremely resistant to antimicrobial activity. To effectively treat periodontal disease, the bacterial load must be reduced to allow healing of the inflamed tissues. Reduction of the bacterial load can be accomplished by surgical methods, nonsurgical methods, or a combination of the two. This article focuses on the nonsurgical treatment of periodontal disease. A thorough oral examination, which includes visual inspection and the use of a periodontal probe, is needed to determine the best therapy. Supragingival cleaning with power and hand scalers is the first step in the therapy process. The next step, subgingival scaling, is necessary to remove bacteria that are in direct contact with the periodontium. Effective subgingival plaque removal is time intensive and requires motivation, manual dexterity, and meticulous technique. Most veterinarians and veterinary technicians lack the training, instruments, and time to remove subgingival plaque effectively. To improve therapeutic results, adjunctive therapy in the form of oral systemic antibiotics or a locally applied doxycycline-containing polymer may be used. The success of periodontal therapy also is dependent on dental home care that takes place after professional treatment. The veterinarian and staff must be willing to educate and reinforce the dental home care efforts of the pet owner. PMID- 11269999 TI - Periodontal surgery equipment. AB - Periodontal surgery requires the operator to have specific knowledge and skills. The use of proper equipment will improve the operator's skill and will decrease surgical time. It will also result in less tissue trauma, thus increasing healing time and reducing postoperative complications and patient discomfort. Various surgical procedures can be enhanced with the usage of equipment that is designed for specific tasks of the procedure. Proper care of equipment is essential for extended instrument life. PMID- 11270001 TI - Oronasal fistula repair. AB - Oronasal fistula is a relatively common complication associated with maxillary canine tooth extraction, problematic healing of maxillectomy, and repair of secondary cleft palate in small animals. Regardless of the clinical scenario associated with oronasal fistula, therapy requires surgical treatment. Principles for surgical repair of oronasal fistula include development of mucosal flaps with excellent vascular supply to transpose over the defect to restore continuity of the nasal and oral cavities. The specific surgical technique may vary but includes either single or double mucosal flaps. Oronasal fistula refractory to multiple attempts at surgical repair may be obturated by using a prosthodontic device. PMID- 11270000 TI - Advanced periodontic techniques. AB - Periodontal disease is multifaceted in nature. Problems occur, such as mucogingival problems, osseous deformities, and gingival enlargement, and they require multiple types of treatment. There is no one way to approach each problem or procedure. Philosophy, talent, experience, training, and objectives ultimately determine treatment selection. PMID- 11270002 TI - Reconstruction of oral defects using mucogingival pedicle flaps. AB - There are many oral pathological conditions that greatly impact the health and the longevity of small animals and require surgical closure. Surgical treatment success is dependent on an accurate diagnosis of the problem. The pet's signalment, medical history, clinical signs, and adjunctive diagnostics (radiographs and histology) are all used concurrently to ascertain a working diagnosis. Once determined, the pathology's management and the subsequent reconstruction of normal anatomy become the surgeon's goals. The use of mucogingival flaps in this reconstruction process is integral to the therapy's success. The oral cavity's abundant blood supply and rapid healing allow these flaps to be well tolerated by the animal and quite resistant to breakdown. Flap designs and implementation are important to this healing process. This article discusses 4 diverse oral cases. Their presentations, diagnoses, and surgical corrections are discussed to show how mucogingival flaps may be used in closures of oral surgical defects. PMID- 11270003 TI - CONSORT, QUOROM, and structured abstracts--new rules for authors, new tools for readers. PMID- 11270004 TI - A comparison of remifentanil and fentanyl in patients undergoing carotid endarterectomy. AB - BACKGROUND AND AIM: We investigated the haemodynamic stability and emergence characteristics of isoflurane/nitrous oxide anaesthesia supplemented with remifentanil or fentanyl in patients undergoing carotid endarterectomy. METHODS: Anaesthesia was induced with propofol (1-2 mg kg-1) and either remifentanil (0.5 microgram kg-1) or fentanyl (1 microgram kg-1), followed by an infusion of remifentanil (0.2 microgram kg-1 min-1) or fentanyl (2 micrograms kg-1 h-1). RESULTS: There were no significant differences between the groups in haemodynamic variables, postoperative pain, nausea or vomiting. After induction there was a significant decrease in mean arterial pressure for both groups (P < 0.001) and a decrease in heart rate (P = 0.001) in the remifentanil group. In both groups these haemodynamic changes continued during maintenance of anaesthesia (P < 0.05). The time to eye opening after surgery was significantly shorter with remifentanil compared with fentanyl (6.62 +/- 3.89 vs. 18.0 +/- 15.18 min, P = 0.015). CONCLUSION: Remifentanil appears to be a comparable opioid to fentanyl when supplementing isoflurane/nitrous oxide anaesthesia for carotid endarterectomy. PMID- 11270005 TI - Comparative analysis of costs of total intravenous anaesthesia with propofol and remifentanil vs. balanced anaesthesia with isoflurane and fentanyl. AB - BACKGROUND AND AIM: We evaluated the costs and benefits of total intravenous anaesthesia compared with a balanced anaesthesia regimen. METHODS: One-hundred and twenty-four patients undergoing cataract surgery were randomized to either a propofol/remifentanil or an isoflurane/fentanyl group. In the propofol/remifentanil group, both drugs were used for induction and maintenance of anaesthesia; in the isoflurane/fentanyl group, anaesthesia was induced with etomidate and fentanyl and maintained with isoflurane and fentanyl. All patients received mivacurium for muscle relaxation and the lungs were ventilated mechanically. The use of propofol and remifentanil resulted in a faster emergence and an overall savings per case of [symbol: see text] 12.25 due to a reduction in personnel costs which outweighs the higher drug acquisition costs. RESULTS: In the propofol and remifentanil group, more patients were satisfied and would accept the same anaesthetic again. CONCLUSION: We conclude that propofol and remifentanil is more cost-effective than isoflurane/fentanyl due to its better recovery profile, reduced total direct costs and higher patient satisfaction. PMID- 11270006 TI - Ketamine-midazolam total intravenous anaesthesia for prolonged abdominal surgery. AB - BACKGROUND AND AIM: Infusion of ketamine and midazolam can maintain prolonged anaesthesia, but delayed recovery is a limitation. We aimed to develop an approximation regimen for the infusion of ketamine and midazolam to obtain steady state anaesthesia with acceptable recovery. METHODS: Thirty-one patients undergoing radical cystectomy were studied. The initial regimen was calculated from drug pharmacokinetic variables and tailored in a pilot study (15 patients) to develop the approximation regimen dosage. Anaesthesia was induced with midazolam (150 micrograms kg-1) and ketamine (2 mg kg-1). Tracheal intubation and ventilation with oxygen enriched air (FiO2 = 0.35) were facilitated by muscle relaxants. Anaesthesia was maintained by the approximation regimen doses. Routine monitoring was used for all patients, but pulmonary artery catheters were inserted in 11 patients, to obtain haemodynamic and oxygenation variables. RESULTS: Steady-state anaesthesia was obtained with minimal deviations in the regimen in some patients followed by reasonable recovery. CONCLUSION: It is concluded that infusion of ketamine and midazolam in the approximation regimen doses can be used to maintain anaesthesia for prolonged abdominal surgery. PMID- 11270007 TI - Management of difficult intubation. AB - Appropriate airway management is an essential part of the anaesthetist's role. Difficult intubation, which can now be quantified using the 'Intubation Difficulty Scale', should be anticipated whenever possible. A strategy needs to be developed in order to anticipate problems. The first part of this paper reviews the different factors that contribute to make intubation and/or ventilation difficult. Problems with intubation (or ventilation of the lungs) can be caused by abnormal laryngeal structures (e.g. tumour, stenosis), or by difficulty in seeing the glottis. The clinical history will usually help identify the former problem, while physical examination of the airway is required to reveal either disproportion between the various structures of the airway (e.g. tongue, larynx), and/or difficulties in aligning the oral, pharyngeal, and laryngeal axes. The different techniques used to diagnose these problems are described. The second part of this paper summarizes the algorithms used by the anaesthetist when management of the airway is found difficult. Three situations are considered: (a) anticipated difficult intubation, for which awake fibreoptic intubation would appear to be the technique of choice in the majority of cases, (b) unforeseen difficult intubation in a patient whose lungs can be ventilated; here, various techniques for control of the airway will be briefly described, and (c) both tracheal intubation and lung ventilation are impossible; this is a life threatening emergency, for which three solutions are proposed. These include use of the laryngeal mask airway, the Combitube, or transtracheal ventilation. These situations will be analysed with the aim of proposing management strategies that always guarantee the safety of the patient. PMID- 11270008 TI - The effects of two single doses of tramadol on sleep: a randomized, cross-over trial in healthy volunteers. AB - BACKGROUND AND AIM: The effects of analgesic drugs on sleep are poorly understood. We investigated short- and medium-term effects of tramadol on sleep structure. METHODS: Eight healthy volunteers received a placebo (predrug placebo night), then, in a randomized, double-blind, cross-over fashion a single oral dose of tramadol 50 mg or 100 mg (drug-night), and finally, again a placebo (postdrug placebo-night). Standardized polysomnography (electroencephalogram, electro-oculogram, submental electromyogram) was continuously recorded during placebo- and drug-nights. RESULTS: During drug-nights both doses of tramadol significantly increased the duration of stage 2 sleep, and significantly decreased the duration of slow-wave sleep (stage 4). Tramadol 100 mg but not 50 mg significantly decreased the duration of paradoxical (rapid eye movement) sleep. In the placebo-night after tramadol 100 mg (but not after 50 mg) duration of stage 2 sleep was significantly shorter, and duration of stage 4 sleep was significantly longer compared with the predrug placebo-night. CONCLUSION: In healthy volunteers, a single dose of tramadol 50 mg disturbs sleep in the night of drug application. With 100 mg, sleep is disturbed in both the night of drug application and in the subsequent night. PMID- 11270009 TI - Minimal flow sevoflurane and isoflurane anaesthesia and impact on renal function. AB - BACKGROUND AND AIM: Compound A generation and accumulation in sevoflurane anaesthesia is dependent on fresh gas flow. We investigated the extent of generation of compound A. METHODS: After Institutional Review Board approval and informed consent, patients with normal renal function were randomized to receive either sevoflurane (n = 33) or isoflurane (n = 43) minimal flow anaesthesia (0.5 L min-1) for at least 2 h under standardized conditions. Compound A concentrations were quantified and blood and urine samples were taken to assess renal involvement. Both groups were comparable. RESULTS: No significant differences concerning blood chemistry and urine measurements were found. The maximum mean compound A concentration was observed 90 min after flow reduction being 40 +/- 9 p.p.m. at a corresponding mean sevoflurane concentration of 2.1 +/ 0.5 vol%. Mean inspiratory compound A exposure was 102 +/- 33 p.p.m h-1. CONCLUSION: Compound A concentrations using 0.5 L min-1 fresh gas flow and a heated absorber were higher than previously published values using an inflow of 1 L min-1. Compound A exposure was similar to other clinical studies which did not show changes in renal and hepatic function. PMID- 11270010 TI - Gum elastic bougie, capnography and apnoeic oxygenation. AB - BACKGROUND AND AIM: This study assessed the accuracy of using capnography with a modified, hollow gum elastic bougie in predicting tracheal intubation, and its effectiveness as a method of apnoeic oxygenation. METHODS: Patients were randomly allocated to having the gum elastic bougie inserted, under anaesthesia, in the trachea or the oesophagus. End-tidal carbon dioxide measurements were made at 10 and 20 s. The position of the gum elastic bougie was correctly predicted in 89.2% of patients. We tested the apnoeic oxygenation on an anaesthetic simulator model, which is housed in the Scottish Anaesthesia Simulator Centre, Stirling, UK. RESULTS: The time taken for the oxygen saturation to fall to 90% was significantly prolonged when the gum elastic bougie was used for apnoeic oxygenation. CONCLUSION: The modification of the gum elastic bougie allows a more objective assessment of correct placement than the previous tactile method. The current design of bougie is unsuitable but can be modified. PMID- 11270011 TI - Interscalene brachial plexus anaesthesia with small volumes of ropivacaine 0.75%: effects of the injection technique on the onset time of nerve blockade. AB - BACKGROUND AND AIM: We evaluated the effect of the injection technique on the onset time and efficacy of interscalene brachial plexus anaesthesia. METHODS: With Ethical Committee approval and written consent, 30 patients undergoing elective shoulder acromioplasty or capsuloplasty were randomly allocated to receive interscalene brachial plexus block with 20 mL of ropivacaine 0.75% by using either a single injection (Single group, n = 15) or multiple injection (Multiple group, n = 15). Nerve blocks were placed with the aid of a nerve stimulator using short bevelled, Teflon coated needles. The stimulation frequency was set at 2 Hz and the intensity of stimulating current, initially set at 1 mA, was gradually decreased to < or = 0.5 mA after each muscular twitch was observed. In the Single group, the anaesthetic solution was slowly injected after the first muscular twitch had been observed. In the Multiple group, 8 mL were injected at shoulder abduction, 6 mL were injected at arm flexion, and 6 mL at the extension of the arm. RESULTS: Placing the block required 5 min (4-8 min) in the Multiple group and 3 min (1-10 min) in the Single group (P = 0.001); however, total preoperative time (from skin disinfection to complete loss of pinprick sensation from C4 to C7 with inability to elevate the limb from the operating table) was shorter in the Multiple group (15 min; range 10-28 min) than in the Single group (23 min; range 14-60 min) (P = 0.03). Additional intravenous fentanyl supplementation was required in two patients of the Multiple group (13%) and eight patients of the Single group (53%) (P = 0.05). CONCLUSION: We conclude that using a multiple injection technique shortened the preparation time and improved the quality of interscalene brachial plexus anaesthesia performed with small volumes of ropivacaine 0.75%. PMID- 11270013 TI - Use of remifentanil for major abdominal surgery. PMID- 11270012 TI - Pre-emptive efficacy of epidural fentanyl in elective abdominal surgery. AB - BACKGROUND AND AIM: This study determines whether epidural fentanyl given before incision decreases the requirements for opioid analgesia postoperatively, compared with the same dose of epidural fentanyl given after the surgery. METHODS: Forty patients scheduled to undergo elective abdominal surgery were randomly allocated between two groups according to the time of administered of fentanyl. None of the patients in either group received premedication. Prior to induction of general anaesthesia an epidural catheter was inserted at the L2-3 interspace and flushed with 0.9% NaCl. Patients then received 100 micrograms fentanyl in 10 mL 0.9% NaCl through this catheter either 15 min before awaking at the end of the operation (group I), or else the same dose given at an estimated time of 15 min before the start of surgery (group II). Postoperative analgesia consisted of patient-controlled intravenous fentanyl. The amount of fentanyl used by the patients was noted at 2, 4, 8, 12 and 24 h after surgery. Pain scores and sedation scores were assessed at 0, 2, 4, 8, 12 and 24 h postoperatively. RESULTS: The consumption of fentanyl was similar in both groups in all studied periods postoperatively. The mean pain score was lower for patients in group I than group II immediately after operation. There were no statistically significant differences between the mean pain scores of groups at 2, 4, 8, 12 and 24 h after operation. Mean sedation scores were similar in both groups at all times postoperatively. CONCLUSION: This study showed that the dose of fentanyl administered epidurally prior to surgical incision did not produce any clinically useful pre-emptive analgesic effect. PMID- 11270014 TI - [Depression among the elderly]. PMID- 11270015 TI - [Depression in the advanced age: therapeutic aspects]. PMID- 11270016 TI - [Depression among the elderly. Etiological and clinical aspects]. PMID- 11270017 TI - [Clinical, biological and social aspects of depression the aged 80 and over]. PMID- 11270018 TI - Hypertonic saline (7.5%) after coronary artery bypass grafting. AB - BACKGROUND AND OBJECTIVE: Patients undergoing coronary artery bypass grafting often require volume loading after operation. In this situation hypertonic saline may be beneficial in restoring the haemodynamic balance and removing excess extravascular fluid. METHODS: Forty coronary artery bypass grafting patients were randomly assigned to receive either hypertonic saline 7.5% (20 patients) or 0.9% saline (20 patients) as a single dose of 4 mL kg-1 over 30 min in the postoperative rewarming phase in the intensive care unit. RESULTS: Mean arterial pressure increased in the hypertonic saline group from 82 +/- 10 (SD) to 104 +/- 17 mmHg (P = 0.002) vs. the normal saline group), and the cardiac index rose from 2.3 +/- 0.5 to 3.4 +/- 0.8 L min-1 m2 (P = 0.002 vs. the normal group). The haemodynamic effect of hypertonic saline lasted only about 1 h. Diuresis was greater in the hypertonic saline group both at 1 h (hypertonic saline: 490 +/- 274 vs. normal saline: 204 +/- 130 mL; P = 0.001) and 10 h (hypertonic saline: 1952 +/- 554 vs. normal saline: 1421 +/- 514 mL; P = 0.003). CONCLUSIONS: No adverse effects were seen. The hypertonic saline had a strong diuretic effect and may be beneficial in coronary artery bypass graft patients after operations. This is because of its value as a short-term plasma expander and the diuresis eliminates excessive fluid from the body. A larger study is needed to determine whether the benefits outweigh the possible side-effects in these patients. PMID- 11270020 TI - Propofol as a continuous infusion during cardiopulmonary bypass does not affect changes in serum free fatty acids. AB - Perioperative myocardial infarction or ischaemia is a potential consequence of cardiac surgery and elevated free fatty acids can increase the severity of myocardial ischaemic damage. We investigated perioperative changes in serum free fatty acids, and other serum lipids, as a consequence of using propofol infusions for cardiac surgery during cardiopulmonary bypass. Twenty-five patients undergoing elective coronary artery bypass grafting were allocated to two groups. One group of 12 patients was given a continuous infusion of propofol and the other group of nine patients received intermittent boluses of midazolam as a hypnotic agent. Serum lipid concentrations were measured at four periods perioperatively. Changes in free fatty acid concentrations were similar between the two groups. Lipid concentrations related to triglyceride in the propofol group decreased on one occasion but subsequently returned to control value. On the other hand, such values in the midazolam group remained lower than control values. Propofol is not a contraindication as an anaesthetic for cardiac surgery in respect of concern regarding the effects of free fatty acids. PMID- 11270019 TI - Small doses of remifentanil or sufentanil for blunting cardiovascular changes induced by tracheal intubation: a double-blind comparison. AB - BACKGROUND AND OBJECTIVE: To compare the effects on cardiovascular changes induced by tracheal intubation when small doses of either remifentanil or sufentanil are used in the presence of midazolam. METHODS: Thirty normotensive, ASA physical status I-II patients, receiving general anaesthesia for major abdominal surgery, received an intravenous midazolam premedication (0.05 mg kg-1) 10 min before induction. They were randomly allocated to receive in a double blind fashion an intravenous bolus of either (a) remifentanil given as a bolus dose 1 microgram kg-1 (n = 15), or else (b) sufentanil 0.1 microgram kg-1 infused over 60 s (n = 15). In each instance this loading dose was followed by a continuous intravenous infusion (0.1 microgram kg-1 min-1 or 0.01 microgram kg-1 min-1 of remifentanil or sufentanil, respectively). General anaesthesia was induced with propofol (2 mg kg-1), followed by atracurium besilate (0.5 mg kg-1) to facilitate tracheal intubation. Following intubation, the lungs were mechanically ventilated with a 60% nitrous oxide in oxygen mixture and a 1% inspired sevoflurane. RESULTS: Arterial pressure and heart rate were recorded before induction of anaesthesia (baseline), immediately before intubation, immediately after tracheal intubation and every minute for the first five minutes thereafter. No differences in systolic and diastolic arterial pressures were observed between the two groups. At the end of the study period, systolic and diastolic pressures slightly decreased from preinduction values in both groups. Four patients in the remifentanil group (26%) and five patients in sufentanil group (33%) showed at least one systolic pressure value < 90 mmHg during the study period (P = not significant); however, the observed decreases in systolic pressure were transient and did not require treatment. Heart rate values were not affected by tracheal intubation in either group. CONCLUSIONS: In healthy normotensive patients without cardiovascular disease the use of a relatively small dose of either remifentanil or sufentanil after standard midazolam premedication results in a similar and clinically acceptable effectiveness in blunting the cardiovascular changes induced by tracheal intubation. PMID- 11270021 TI - Successful resuscitation after catastrophic carbon dioxide embolism during laparoscopic cholecystectomy. AB - A 92-year-old female was scheduled for laparoscopic cholecystectomy. Following intraperitoneal carbon dioxide insufflation and removal of her gallbladder, the patient developed serious haemodynamic deterioration associated with a decrease of both end-tidal carbon dioxide concentration (ETCO2) and chest compliance. Carbon dioxide embolism was suspected and the diagnosis was confirmed by aspiration of 20 mL of foamy blood from the central venous line. The patient was successfully resuscitated after discontinuation of carbon dioxide insufflation and ventilation of the lungs with 100% oxygen. Carbon dioxide embolization must always be suspected during laparoscopic surgery whenever sudden haemodynamic deterioration associated with a decrease in ETCO2 and chest compliance occur. PMID- 11270022 TI - Severe life threatening reaction to Haemaccel in a patient with bronchial asthma. PMID- 11270023 TI - Use of 10% lidocaine in the cuff of the endotracheal tube. PMID- 11270024 TI - Is PEEP, truly a PEEP? PMID- 11270025 TI - Use of a metal tracheostomy tube for laser surgery to a subglottic stenosis. PMID- 11270026 TI - Speciality status for intensive care medicine? PMID- 11270027 TI - Perineural injection to nerve root and radicular blood flow: a clinical study during spinal surgery. AB - BACKGROUND AND OBJECTIVE: To investigate the effects of the perineural injection of lidocaine or corticosteroids on radicular blood flow during spinal surgery. METHODS: After lumbar discectomy, a probe for laser Doppler flowmetry was placed directly on the 4th or 5th lumbar nerve root. Thirty patients undergoing lumbar discectomy were randomly assigned to one of three groups. Each group received one of three protocols for a perineural injection to the nerve root: 1.0 mL 0.9% saline in group A, 1.0 mL 1% lidocaine in group B or 1.0 mL dexamethasone (4 mg) in group C. Measurements included radicular blood flow, mean arterial pressure, haemoglobin concentration, percutaneous oxygen saturation and end-tidal carbon dioxide tension. Radicular blood flow was measured by laser Doppler flowmetry before the injection and 15 min after these injections. The three groups were similar with respect to mean arterial pressure, haemoglobin concentration, percutaneous oxygen saturation and end-tidal carbon dioxide tension. RESULTS: Radicular blood flow did not change after the injection in any of the groups. CONCLUSIONS: The results suggest that the perineural injection of 1% lidocaine or dexamethasone does not affect radicular blood flow during lumbar discectomy. PMID- 11270028 TI - Comparison of the effects of clonidine and hydroxyzine on haemodynamic and catecholamine reactions to microlaryngoscopy. AB - BACKGROUND AND OBJECTIVE: This study compares the effect of oral clonidine vs. hydroxyzine on the haemodynamic and catecholamine responses to microlaryngoscopy. METHODS: Thirty-five ASA II-III patients were included in this double-blind randomized trial. The patients received either hydroxyzine 1 mg kg-1 (n = 18) or clonidine 3 micrograms kg-1 (n = 17) for their oral premedication 100 min before an intravenous induction of anaesthesia using propofol (2-3 mg kg-1) and fentanyl (2 micrograms kg-1). Arterial pressure and heart rate were measured before premedication, and throughout the procedure and recovery. Plasma catecholamine levels were determined before premedication, after induction, and 1.5, 30 and 120 min after laryngoscopy. RESULTS: Mean arterial pressure was significantly lower after clonidine, whereas there was no difference in heart rate and plasma catecholamine levels between the two groups. CONCLUSIONS: Clonidine for premedication significantly decreased mean arterial pressure during microlaryngoscopy and the following recovery phase but did not modify the overall haemodynamic response to the suspension microlaryngoscopic nociceptive stimulus. PMID- 11270029 TI - The plasma elimination rate and urinary secretion of procalcitonin in patients with normal and impaired renal function. AB - BACKGROUND AND OBJECTIVE: The amount of procalcitonin eliminated in the urine and the plasma disappearance rate of procalcitonin were evaluated in patients with normal and impaired renal function, because patients with sepsis are a main target group for procalcitonin measurement, and these patients often develop renal dysfunction. METHODS: Elimination of procalcitonin in the urine (microgram 12 h-1) was measured in 76 patients. In another 67 patients, the 50% plasma disappearance rate (t1/2, h) was evaluated 48 h after peak concentrations (procalcitonin > 2 micrograms L-1). Renal function was assessed by creatinine clearance. RESULTS: Procalcitonin elimination in the urine was significantly reduced in patients with severe renal dysfunction. However, the plasma disappearance rate correlated only weakly with renal dysfunction (Spearman's rank correlation R = -0.36, P = 0.004, regression t1/2 = 49.87-0.15 creatinine clearance). The 25% quartile and median were 25.2 h and 30.0 h in patients with normal renal function, and 36.3 h and 44.7 h in patients with severely impaired renal function (creatinine clearance < 30 mL min-1). CONCLUSIONS: Renal elimination of procalcitonin is not a major mechanism for procalcitonin removal from the plasma. Although the plasma disappearance rate may be prolonged up to 30 50% in some patients with renal dysfunction, clinical diagnostic decisions may not be severely influenced by this moderate prolongation of procalcitonin elimination. We conclude that procalcitonin can be used diagnostically in patients with renal failure as well as in those with normal renal function. PMID- 11270030 TI - Intravenous ketamine attenuates arterial pressure changes during the induction of anaesthesia with propofol. AB - BACKGROUND AND OBJECTIVE: To investigate whether the administration of ketamine before induction with propofol produces a smaller decrease in arterial pressure. METHODS: Twenty-two patients were assigned to one of two groups to receive either propofol with ketamine (n = 11) or propofol alone (n = 11, control). Anaesthesia was induced with 2 mg kg-1 propofol and 0.5 mg kg-1 ketamine or 2 mg kg-1 propofol alone. Ketamine was administered 1 min prior to induction with propofol. Immediately after induction with propofol, vecuronium (0.15 mg kg-1) was administered. Four minutes after administration of vecuronium, tracheal intubation was performed. Anaesthesia was maintained using sevoflurane (0.5%) in 66% nitrous oxide until 3 min after intubation. Systolic, diastolic and mean arterial pressure and heart rate were recorded on arrival, directly before induction with propofol, prior to tracheal intubation, immediately after intubation and at 3 min after intubation. RESULTS AND CONCLUSIONS: Administration of ketamine before induction with propofol preserved haemodynamic stability compared with induction with propofol alone. PMID- 11270031 TI - Changes in jugular bulb oxygenation in patients undergoing warm coronary artery bypass surgery (34-37 degrees C). AB - BACKGROUND AND OBJECTIVE: Imbalance between cerebral oxygen supply and demand is thought to play an important role in the development of cerebral injury during cardiac surgery with cardiopulmonary bypass. METHODS: We studied jugular bulb oxygen saturation, jugular bulb oxygen tension, arterial-jugular bulb oxygen content difference and oxygen extraction ratio in 20 patients undergoing warm coronary artery bypass surgery (34-37 degrees C) with pH-stat blood gas management. RESULTS: Only two patients showed desaturation (jugular bulb oxygen saturation < 50%) at 5 min on bypass, and none from 20 min onwards. Multiple regression models were performed after using bypass temperature, mean arterial pressure, cerebral perfusion pressure, haemoglobin concentration and arterial carbon dioxide tension as independent variables, and arterial-jugular bulb oxygen content difference, jugular bulb oxygen saturation, oxygen extraction ratio and jugular bulb oxygen tension as individual dependent variables. CONCLUSIONS: We found that jugular bulb oxygen saturation, jugular bulb oxygen tension and oxygen extraction ratio are mainly dependent on arterial carbon dioxide tension, and arterial-jugular bulb oxygen content difference is dependent on arterial carbon dioxide tension and the bypass temperature. Our results suggest jugular bulb oxygenation is mainly dependent on arterial carbon dioxide tension during warm cardiopulmonary bypass. PMID- 11270032 TI - [Early cellular immune response in experimental labyrinthitis: immunohistochemical study]. AB - Labyrinthitis ossificans is a recently recognized entity with the extending cochlear implant surgery. Up to date there are not so many studies of the cellular response leading the organization of the inflammatory reaction within the cochlea. Immunemediated labyrinthitis is a valid experimental model that allow the knowledge of the cellular infiltration mechanisms within the cochlea. The inner ear communicates with systemic immunity via the circulation by the passage of cells through the spiral modiolar vein (SMV) and its collecting venules in the scala tympani. Inflammatory reaction within the cochlea lead to the formation of fibrotic tissue and bone (osteoneogenesis) inside it and injuring the neurosensory organs of hearing and balance. To test which cells are proliferating early in an inflammatory response, an animal model of Keyhole limpet haemocyanin (KLH) induced labyrinthitis was utilized, showing a granulomatous lesions not previously reported in this experimental model. PMID- 11270033 TI - [Morphofunctional changes in olfactory bulb after elimination of smell stimuli (I). Morphologic study]. AB - We study the olfactory bulb plasticity by olfactory deprivation in the jerbil (Meriones Unguiculatus). We analyze what effects has the odorous stimuli privation in the first days after the birth on the olfactory bulb. We have found that this stimuli absence reduces the size of the bulb in the discussed side and alters the volumes of some of the bulbars layers. This indicates that the structural development of the nervous system specifies of an integrity of their sensorial afferencies. PMID- 11270034 TI - [Cultural adaptation of 2 questionnaires for health measurement in patients with vertigo]. AB - OBJECTIVE: To perform a transcultural adaptation form English to Spanish of two common questionnaires of handicap assessment in vestibular disorders. STUDY DESIGN: Prospective study. PATIENTS: 337 patients seen for non-acute dizziness from peripheral or central origin in a tertiary referral setting. MAIN OUTCOME MEASURES: Dizziness Handicap Inventory test and UCLA-Dizziness Questionnaire after transcultural adaptation following the method of translation backtranslation, expert assessment and statistical validation. RESULTS: The results after cultural adaptation and reliability assessment provide a firm basis to demonstrate the close relation of the Spanish and English version in all the items and their meaning. CONCLUSION: This adapted questionnaires can be used to assess vestibular disability with no loss of metric values of the original version. PMID- 11270035 TI - [Clinical and nasometric study of velopharyngeal function in two-stage palatoplasty]. AB - Since 1976 children born with complete cleft lip and palate have been treated according to two stage cleft palate repair regime, soft palate at one year of age and hard palate between 3-12 years of age. The objective of this paper is to present the clinical and instrumental speech evaluation and the velopharyngeal function of 41 patients, mean age 8.2 years, 28 unilaterals and 13 bilateral clefts; 7 cases had palatal fistulae larger than 5 mm2. The results showed moderate-severe hypernasality in 20% of cases, persisting articulatory errors with backing in 43% of cases, glotal stops in 2%, the intelligibility was deficient in 7%. The evaluation of the oro-nasal resonance with Nasometry showed suspicious or VPI values in 14 and 20% of cases respectively. Nasendoscopy findings showed same degree of VPI in 35% of cases. Repalatoplasty or fistula closure was needed in 17% and pharyngeal flap in 12% of cases. The great incidence of articulatory errors have promoted us to discontinue this technique. PMID- 11270036 TI - [Correlation of clinical, radiologic, and histopathologic findings in laryngeal, hypopharyngeal, and oropharyngeal cancer]. AB - The correct staging of the tumors has important consequences in the planning of the treatment in the patients with cancer of larynx, hypopharynx and oropharynx. The objective of this paper is to study the correlation of the clinical, radiologic and pathologic staging with the purpose to evaluate if the computed tomography (CT) is effective in the diagnostic of the stage in the different tumoral findings. We did a retrospective study in 34 patients with pharynx and larynx cancer in the "Hospital Universitario Principe de Asturias" between the years 1994-1998. The method was: 1. Clinical history and ENT examination. 2. Head and neck CT. 3. Direct laryngoscopy and biopsy. 4. Surgical treatment and posterior pathologic staging of the removed specimen. The lesions were studied according the TNM-UICC classification with the T and N stage. The first thing to do was the clinical stage then the radiologic stage and at last compare it with the pathologic stage after surgical removal. The results are presented in percentages and confirm that the clinical and radiologic combined information improve the correlation between clinical and pathologic staging in the cancer of larynx, hypopharynx and oropharynx. PMID- 11270037 TI - [Our management protocol and surgical technique in cerebrospinal fluid rhinorrhea treated with an endonasal approach]. AB - Five patients with cerebrospinal fluid fistula (CFF) have been treated with intratecal fluoresceine, 2 cc at 2%, and endoscopic nasal surgery. In 3 patients CFF was postraumatic; one case spontaneous and another case iatrogenic. In all the cases CFF have been solved in the first time. Postoperatory follow-up vary from 8 to 14 months, and no recurrence was observed. Fluorescein must be managed adequately for prevent neural complications. PMID- 11270038 TI - [Sinonasal intestinal-type adenocarcinoma: report of 7 cases]. AB - The intestinal-type adenocarcinoma of the nasal cavity and paranasal sinuses are uncommon tumors, with less than 4% of the total of malignancies of this region. They have histological similitudes with the glandular estructure of the intestinal mucosa. In some aspects they are similar to others tumors of this area, symptoms, an etiological relation with the exposure to wood dust ... but they have differences in the local aggressivity, this is important for the tractament, evolution and survival. The authors present a revision about clinic characteristics, diagnostic and tractament of seven cases of nasosinusal intestinal type adenocarcinoma. PMID- 11270039 TI - [Non-invasive fungal sinusitis: allergic fungal sinusitis and sinusal mycetomas]. AB - We review non-invasive fungal sinusitis (FS) cases treated at our Unit following the recent SF classification, based on physiopathology, treatment and prognosis. We report 7 FS cases treated during 2 years and followed a minimum of 24 months. They are two allergic FS, one of them related to an allergic bronchopulmonary aspergillosis, and five mycetomas, two of them without a sinusal foreign body, sphenoidal and maxillary respectively, and three caused by a maxillary foreign body. All were treated by endoscopic sinus surgery (ESS). The two allergic cases were also treated with systemic corticoids in the postoperative period. We review the clinical presentation of the different types of non-invasive FS, the importance of endoscopy and imaging techniques in their management, the usefulness of ESS, the common association of FS to foreign bodies, and their prognosis. PMID- 11270040 TI - [Frontolateral laryngectomy. Our experience in La Rioja]. AB - It has been carried out a retrospective study on 16 patients who have suffered from glotic tumorus in T1b stage and have been treated by frontolateral laryngectomy from 1989 to 1998. Data related to age, sex, toxic habits, anatomopathological diagnosis, surgery, recurrence and survival were analysed. The disease affects more in males, in the 5th and 6th decade of life, with a medium to a high consumption of alcohol. The anatomopathological diagnosis in 96.75% of the cases was epidermoid carcinoma. The survival analysis showed that 87.5% of the patients survived free of recurrence for the first five years and only 6.25% had a local recurrence. PMID- 11270041 TI - [Variability in the digital voice analysis depending on the analyzed vocal, in normal patients and in patients with dysphonia]. AB - The aim of this study is to decide which of Spanish's 5 vowels is the most suitable for the analysis of the voice by means of the program Dr. Speech 3.0. We assessed 98 patients: 51 normal subjects and 47 hoarse patients (34 polyps, 7 nodules, 2 hyperfunctional dysphonia, 4 other pathologies) before and after following a surgical and/or speech therapy. The methodology of the study included: protocolized questionnaire, ENT examination, perceptual evaluation of the hoarse voices and physical analysis of the 5 sustained vowels. Jitter, shimmer, NNE and HNR in the voice varied depending on the analyzed vowel and from normal subjects to hoarse patients. NNE and HNR for vowels [o] and [u] was higher than for [a], such differences were statistically significant (p < 0.001). Shimmer for vowels [o] and [u] was lower than shimmer for [a] in hoarse patients (p < 0.05). Jitter for [a] was higher than jitter for the other vowels in normal patients. All algorithms improveded with surgical and/or speech therapy. Especially levels of noise for vowels [a] [e] and [o] presented statiscally significant improvements (p < 0.001), shimmer for [a] and [e] (p < 0.001) and jitter for [e] (p < 0.01). The hoarse patients presented values more pathological than the normal subjects, especially jitter for vowels [a] [e] and [o] (p < 0.001), shimmer for [a] [e] [i] and [u] (p < 0.001) and level of noise for [a] [e] [o] and [u] (p < 0.001). CONCLUSION: The analysis of vowel [e] presented higher r than [a] when correlated with perceptual evaluation of voice or with voice quality that gave us Dr. Speech 3.0. Also, the [e] is the best vowel to express the improvements of jitter, shimmer and level of noise of hoarse patients after following surgical and/or speech therapy. PMID- 11270042 TI - [Positron-emission tomography (PET) in the diagnosis of malignancy, recurrence, and staging in patients with head and neck tumors]. AB - We report a prospective study with 15 patients with the diagnosis of head and neck tumors. They underwent two types of studies, the radiological one with computed tomography or magnetic resonance image scan and the positron emission tomography (PET) with 18-fluorodeoxyglucose (FDG). Five patients did not receive any treatment before, and the PET was performed to evaluate the possibility of malignancy, to determine the stage of the disease and if a recurrence was suspected. The study with the PET has the advantage of detecting small lesions and it is not influenced by radiotherapy or surgery. PMID- 11270043 TI - [Allergic fungal sinusitis]. AB - Allergic fungal sinusitis is a recently described clinical entity that has gained increased attention as a cause of chronic sinusitis. Consist in a benign noninvasive sinus disease related to a hypersensitivity reaction to fungal antigens. It should be suspected in any atopic patient with refractory nasal polyps. Computed tomography (CT) findings are characteristics, but not diagnostic. Diagnosis requires show allergic mucin in the histopathologic examination and hiphae in special fungal stains. The suitable treatment includes the allergic mucin removal and sinus aeration accomplished endoscopically, perioperative systemic steroids and immunotherapy with fungal antigens. We present a case of this kind of chronic sinusitis describing the characteristic histopathologic and radiologic findings, the pathogenic theories and recent advances in immunotherapy. PMID- 11270044 TI - [Hemifacial microsomia or syndrome of the first and second branchial arch: clinical features and therapeutic considerations]. AB - The syndrome of hemifacial microsomia is formed by a constellation of congenital malformations of the originating face structures of the I and II branchial archs, in which the first pharingeal stock and the first soft palate take part to, along with the nucleus of the temporary bone. We present a new case of hemifacial microsomia, and reviewed its clinical characteristics and therapeutic possibilities. PMID- 11270045 TI - [Laryngeal rhabdomyoma: unusual case of dysphonia. Review of the literature]. AB - Rhabdomyomas of the larynx are extremely rare benign tumors. Only 25 have been well documented until now. These tumors display a low growing pattern. Diagnosis is based on immunocytochemical studies and electron microscopy. Three histological types are distinguished: adult, fetal and fetal myxoidal or genital. Surgery is the treatment of choice. A new case, adult type, is presented and the literature reviewed. PMID- 11270046 TI - [Head and neck lesions caused by systemic progressive ossifying fibrodysplasia (Munchmeyer's disease]. AB - Fibrodysplasia ossificans progressiva is a rare hereditary connective tissue disorder due to an unknown mutation conditioning the ossification of striated muscle and soft tissues frequently related to joints and stimulated by traumatisms or aggressive iatrogenic manouvres. It produces stiffness, pain, disability, swelling and fibro-osseus masses disseminated everywhere, with a very spectacular, invalidant and irreversible clinical picture. Affection of head and neck is unusual, excepting lesions in sternocleidomastoid or masticatiry muscles. We show a systemic case of fibrodysplasia osificans progressiva with enhanced clinical and radiological findings in cervical rhachis, pharynx and larynx, oral cavity, facial bones and ear. Association of biphophonates and steroids for a long period of time seems up to now stopping the evolution but not its regression. PMID- 11270047 TI - [Dysphagia caused by Zenker's diverticulum after total laryngectomy]. AB - Zenker's diverticulum is a mucosal lined outpouching of pharynx through Lainert's space that causes dysphagia of the upper digestive tract. Multiples theories try to explain the acquired etiology of this entity, attributing its origin to a disfunction of pharynx-esophageal sphincter. A case of total larynguectomy with hypopharyngeal diverticulum and progressive dysphagia to solid food is presented. We analyze the etiopathogenic mechanisms and the definitive characteristics of this entity. We review mundial literature, being exceptional the fact of finding clinical manifestations in diverticulum of larynguectomized patients. PMID- 11270049 TI - [Classic Kaposi's sarcoma in the ORL area]. AB - Kaposi's Sarcoma is a malignant neoplasty originated in vascular endothelial cells. Such medical case appears as its classical type located in the tongue and inferior limbs. The therapy consisted of intralesional injections of interferon alfa, leading into better clinic and macroscopic conditions of the injuries. PMID- 11270050 TI - Counseling for type A and recurrent heart disease: Friedman et al. (1986). PMID- 11270048 TI - [Non-Hodgkin's lymphoma of the parotid gland: 2 cases]. AB - We report two cases of bilateral parotid swelling at initial clinical assessment of non-Hodgkin lymphoma. The swelling was not associated with other symptoms. They both were stage IV and were treated by chemotherapy. We have reviewed the literature about first manifestations of the disease, clinical diagnosis and incidence. The importance of early biopsy of such lesions is emphasised. PMID- 11270051 TI - Psychological stress and wound healing: Kiecolt-Glaser et al. (1995). PMID- 11270052 TI - Placebos and their effectiveness: Roberts (1995). PMID- 11270053 TI - Support groups and metastatic breast cancer: Spiegel et al. (1989). PMID- 11270055 TI - A structured psychiatric intervention for patients with malignant melanoma: Fawzy et al. (1990b). PMID- 11270056 TI - Psychosocial influences and mortality: Ruberman et al. (1984). PMID- 11270058 TI - Mind-body common sense. PMID- 11270057 TI - Parental love and health: Russek & Schwartz (1997a). PMID- 11270059 TI - An evidence base for identifying patients' thoughts and feelings. PMID- 11270060 TI - Psychological stress and the common cold: Cohen et al. (1991). PMID- 11270061 TI - Supportive-expressive group therapy and life extension of breast cancer patients: Spiegel et al. (1989). PMID- 11270062 TI - Colds and the stress-illness connection: Cohen et al. (1991). PMID- 11270063 TI - Prayer and distant healing: Sicher et al. (1998). PMID- 11270065 TI - Social relationships and health: Berkman & Syme (1979). PMID- 11270064 TI - Positive and negative beliefs and the course of AIDS: Taylor et al. (2000). PMID- 11270066 TI - The sociopsychological ecology of illness: Stewart Wolf and Helen Goodell's revised second edition of Harold G. Wolff's Stress and Disease (1968). PMID- 11270067 TI - Hostility and heart disease: Williams et al. (1980). PMID- 11270069 TI - A critical view. PMID- 11270068 TI - Prospective studies offer promise in establishing the mind-body connection: Reed et al. (1994) and Bower et al. (1998). PMID- 11270070 TI - Ivan Pavlov and the conditioning of physiological responses. PMID- 11270071 TI - A positive assessment. PMID- 11270072 TI - Seeing what we can see: on the coming exchange between Jon Kabat-Zinn and Arnold S. Relman. PMID- 11270073 TI - Nonlocal mind: the seminal experiments of Bernard Grad. PMID- 11270074 TI - Titanium alloys for fracture fixation implants. AB - This paper is intended to provide an overview of the composition, mechanical properties, biocompatibility, and clinical applications for titanium alloys that are used for fracture fixation implants. A new class of titanium implant alloys has emerged in recent years that exhibits a beta microstructure and a unique combination of mechanical properties. Important information regarding notch sensitivity testing and clinical significance is also discussed. Attributes such as stress corrosion cracking resistance, fatigue strength, and wear characteristics are also essential for specific clinical applications, but are beyond the scope of this presentation. PMID- 11270075 TI - Cobalt-base alloys used in bone surgery. AB - Cobalt-base alloys may be generally described as non magnetic, wear, corrosion and heat-resistant (high strength even at elevated temperature). Many properties of the alloy originate from the crystallographic nature of cobalt, the solid solution-strengthening effect of chromium and molybdenum, the formation of extremely hard carbides and the corrosion resistance imparted by chromium. Cobalt base alloys are difficult to fabricate which is why their use has been limited, but continuous work led to the development of specialized casting methods. Due to its excellent resistance to degradation in the oral environment, the first medical use of cobalt-base alloys was in the cast of dental implants. Various in vitro and in vivo tests have shown that the alloys are biocompatible and suitable for use as surgical implants. Today, the use of Co alloys for surgical applications is mainly related to orthopaedic prostheses for the knee, shoulder and hip as well as to fracture fixation devices. Joint endoprostheses are typical long-term implants and the applied implant material must therefore meet extremely high requirements with regard to biocompatibility with the surrounding body tissue material and corrosion resistance to body fluids. PMID- 11270076 TI - Stainless steel in bone surgery. AB - Today, stainless steel is one of the most frequently used biomaterials for internal fixation devices because of a favorable combination of mechanical properties, corrosion resistance and cost effectiveness when compared to other metallic implant materials. The biocompatibility of implant quality stainless steel has been proven by successful human implantation for decades. Composition, microstructure and tensile properties of stainless steel used for internal fixation is standardized in ISO and ASTM material specifications. Metallurgical requirements are stringent to ensure sufficient corrosion resistance, nonmagnetic response, and satisfactory mechanical properties. Torsional properties of stainless steel screws are different from titanium screws. Stainless steel bone screws are easier to handle because the surgeon can feel the onset of plastic deformation and this provides adequate prewarning to avoid overtorquing the screw. New nickel-free stainless steels have been recently developed primarily to address the issue of nickel sensitivity. These stainless steels also have superior mechanical properties and better corrosion resistance. The Ni-free compositions appear to possess an extraordinary combination of attributes for potential implant applications in the future. PMID- 11270077 TI - Nonresorbable polymers in bone surgery. AB - Sufficient strength and stiffness, biocompatibility and long-term stability are important criteria that polymers have to fulfill for successful implant applications in bone surgery. An additional requirement is sterilization without degradation of the polymer properties. Ultra-high molecular weight polyethylene is used as a carrier material in joint replacement components because of good wear and damping properties. Nevertheless, reduction of wear is still a major challenge in polyethylene research as wear particles are responsible for early loosening of prosthetic devices. Acrylic bone cement is used for the fixation of artificial joints, providing an immediate and strong fixation of the implant. Possible problems with this bone cement are the residual (toxic) monomers and the high curing temperature that may cause thermal bone necrosis. Polyacetal resins are implantable high performance polymers. Due to their high chemical and mechanical resistance, they are used in joint replacement components and other long-term implants. Polyetheretherketone is a new and even more stable high performance polymer. Due to its stability at sterilization temperatures and under irradiation, PEEK is intended to replace POM in future. PEEK is available in a "medical grade" quality suitable for long-term implantation. PMID- 11270078 TI - Bioresorbable polymers in trauma and bone surgery. AB - During the last few decades interest in resorbable polymeric materials has been steadily increasing. As with other materials for implantable devices, they have to satisfy several biological and technical requirements. Implants should maintain adequate mechanical properties in vivo for the time required and degrade at an effective rate. The conditions of polymer synthesis, further processing into implants and the sterilization process determine the mechanical properties of resorbable implants. Degradation of implants is manifested by implant fragmentation, strength loss and the decrease of polymer molecular weight. The rate of degradation and the tissue reaction are strongly affected by the material chemical composition and to some extent also by the mechanical properties. Potentially, devices made from bioresorbable polymers can overcome problems associated with metal implants like stress protection, potential for corrosion, wear and debris formation, as well as the necessity of implant removal. Resorbable polymers have proven to be good materials for a range of devices in trauma surgery. However, modifications and optimizations are still required. Three-dimensional porous scaffolds in various geometrical forms offer a good potential for the manufacture of tissue-engineered implants in the future. PMID- 11270080 TI - Calcium orthophosphates in medicine: from ceramics to calcium phosphate cements. AB - Calcium phosphate (CaP) compounds are becoming of increasingly great importance in the field of biomaterials and, in particular, as bone substitutes. Recent discoveries have accelerated this process, but have simultaneously rendered the field more complicated for the everyday user. Subtle differences in composition and structure of CaP compounds may have a profound effect on their in vivo behaviour. Therefore, the main goal of this article is to provide a simple, but comprehensive presentation of CaP compounds. Reference is made to the most important commercial products. PMID- 11270079 TI - Inert bioceramics (Al2O3, ZrO2) for medical application. AB - Ceramics is defined as the art and science of making and using solid articles having as essential components inorganic non-metallic materials and which are composed largely of them (Kingery et al., 1976). The inherent brittleness of traditional ceramics has limited their ability to compete with ductile metals and polymers for technical applications. However, over the last 100 years innovative techniques in the fabrication of ceramics have led to their use as high-tech materials. Inert bioceramics, such as Al2O3 and ZrO2, have inherently low levels of reactivity compared with other materials such as polymers and metals as well as surface reactive or resorbable ceramics. In a human body, they are expected to be non-toxic, non-allergenic, and non-carcinogenic for a life-time, which leads to a corresponding range of engineering design philosophies for medical application. Due to their excellent frictional properties technical ceramics are nowadays mainly used in endoprosthetics. PMID- 11270081 TI - About composite materials and their use in bone surgery. AB - Composite materials consist of two or even more different material components or phases, which are combined with the aim to improve physical, mechanical and/or biological properties. Such structures are designed to fulfil very specific requirements with respect to a selected device application making full use of their higher weight-specific strength and/or stiffness. Furthermore, these materials offer an opportunity for constructing radiolucent devices. In medical technology, composite materials mainly consist of a polymer matrix and fibres as a reinforcement phase. Composites similar to those known from technical applications reveal a number of specific biological problems. This is due to the materials and manufacturing processes available for the construction of such composites preventing their unrestricted use in direct bone contact. Nevertheless, an application potential for these materials in bone surgery exists and justifies further research and development efforts. PMID- 11270083 TI - AO Scientific Supplement to Injury deals with the different implant materials used in the treatment of locomotor trauma and disease. PMID- 11270082 TI - Unalloyed titanium for implants in bone surgery. AB - Commercially pure (c.p.) titanium has proven its suitability as an implant material in bone surgery over many years in the fields of osteosynthesis, oral implantology, and in certain applications in joint prosthetics. Excellent biocompatibility and corrosion resistance are outstanding features. Furthermore, c.p. titanium is known for not causing allergic reactions. The different grades of c.p. titanium and their minimum mechanical properties are specified in ISO and ASTM standards for implant materials. Typical mechanical properties are given for AO ASIF implant applications. The properties and clinical performance of c.p. titanium are discussed and compared to those of implant stainless steel and titanium alloys. In brief some specific features relating to c.p. titanium implant material are treated, including biocompatibility and soft tissue and bone response and taking into account the effects of implant surface configurations at the same time. In addition, issues are addressed which arise from frequent inquiries from clinics. PMID- 11270084 TI - . . . and clinical immunology? PMID- 11270085 TI - Passive smoking and expired carbon monoxide concentrations in healthy and asthmatic children. AB - BACKGROUND: Carbon monoxide (CO) in expired air has been reported to be an indirect measurement for the quantity of passive smoking. Since endogen CO is produced in inflammatory processes and inflammation is the main pathogenetic mechanism of asthma, it was aimed to investigate the relationship between the intensity of passive smoking and CO concentration in expired air of healthy and asthmatic children. METHODS AND RESULTS: The study was performed in the outpatient pediatrics clinics and day care centers. Knowledge about indoor smoking habits were obtained from parents. The exhaled CO concentrations were measured by a portable device in 235 healthy (mean age, 4.4 +/- 2.3 years) and 54 asthmatic (mean age, 4.5 +/- 1.7 years) children. Children with no smoking parents had the lowest exhaled CO concentrations. Significant relationships were found between the number of smoking cigarettes in the house and exhaled CO concentrations in both healthy (p = 0.003) and asthmatic (p = 0.01) children. Carbon monoxide concentrations were higher in asthmatic children than healthy ones (mean +/- SD, 1.32 +/- 1.50 ppm and 0.86 +/- 1.35 ppm, respectively, p = 0.028) if their parental smoking habits were not taken into account. Asthmatic children of non-smoking parents had higher CO concentrations than healthy subjects of non-smoking parents (1.05 +/- 1.55 ppm vs 0.37 +/- 0.53 ppm, p = 0.01). On the other hand, asthmatic children who has no smoking parents and did not receive inhaled steroids had significantly higher CO concentrations (1.75 +/- 1.45 ppm) than those who received steroids (0.58 +/- 0.65 ppm, p = 0.024). CONCLUSIONS: Exhaled CO can be used as an indicator of passive smoking in children. Higher expired CO of asthmatic children may reflect inflammation of the lung in asthma. PMID- 11270086 TI - Antigens of Fusarium solani: types and criteria for immunochemical characterization. AB - BACKGROUND: There are no established methods for the preparation and standardization of Fusarium solani antigens. This lack of standardization makes it difficult to use these antigens in allergenic diagnostic tests. OBJECTIVE: To obtain an appropriate standardized method for the preparation of the different antigen types of F. solani. METHODS: Production of fungal extracts, followed by biochemical and immunological characterization. RESULTS: The somatic antigens presented the greatest protein content most of these proteins are common to the metabolic and hydrosoluble antigens, particularly those proteins at 35-39 kDa, 29 32 kDa and 15-16 kDa, as detected by electrophoresis and immunoblotting. The hydrosoluble antigens presented the highest protein diversity; these proteins were the most specific, showing minor determinants in common with the other antigens. CONCLUSIONS: It is recommended that a mixture of the different antigen sources be used in order to obtain extracts which would cover the maximum number of diagnostic possibilities. PMID- 11270087 TI - Hereditary angioedema. Long-term follow-up of 88 patients. Experience of the Argentine Allergy and Immunology Institute. AB - Since the detection of the first patient with hereditary angioedema (HA) in 1978, 88 new patients belonging to 16 families have been referred to our clinic. Eighty patients had Type I disease, 5 Type II, and 3 Type III (secondary). We describe the clinical onset, frequent complications, diagnostic tests of the complement system, and abnormalities of the coagulation pathway linked to complement activation. Particular attention was paid to family members who could present succedaneum symptoms. The results of danazole and other therapies and protective and preventive treatment for surgery also are discussed. PMID- 11270088 TI - [Data on the safety of antihistaminics obtained from published clinical trials]. AB - BACKGROUND: We studied H1 antihistaminic safety, with respect to CNS, and clinical trial validity to determine its adverse effects and generalize the results. METHODS AND RESULTS: We employed clinical trials published between 1990 94, selecting them according to several quality criterion to obtain reliable and extrapolable conclusions. Patients that use modern antihistaminics have less adverse effects (32.5%), than those patients that use classics ones (78.6%) and similar to placebo (27.7%). Loratadine (28.4%) and noberastine (22.3%) are the antihistaminics with less adverse effects. Sedation (13.2%) and fatigue (4.9%) are the adverse effects more frequently produced by antihistaminics. Headache (6.7%) that is a frequent adverse effect, has a doubtful causality relationship with the use of antihistaminic. CONCLUSIONS: Second generation antihistaminics have less adverse effects and similar efficacy than classics ones. Noberastine and loratadine are the antihistaminics with less adverse effects in this study. PMID- 11270089 TI - Is serum ECP level helpful in determining discontinuation of inhaled corticosteroid therapy in asthmatic children? AB - BACKGROUND: Serum eosinophil cationic protein (ECP) has been promoted as a direct marker of eosinophilic inflammation of the airways in patients with asthma. However, its role in monitoring disease activity and management of inhaled corticosteroid (ICS) therapy is not well defined. METHODS: We determined serum ECP (s-ECP) levels in 95 children (mean +/- SD age, 6.2 +/- 3.9 years) with asthma. At the time of measurements, 34 out of 95 children were symptomatic whereas 61 were in stable condition; and 56 of 95 patients were on maintenance ICS therapy. ICS prophylaxis was withdrawn in 16 of those 56 patients who remained asymptomatic with a dose of 100 micrograms/day of budesonide for 8 weeks. Eight out of these 16 children had to restart ICS therapy within the following 12 weeks, while the remaining 8 children continued to be asymptomatic within the same period. RESULTS: ECP values and number of patients with a high ECP level (> or = 15 micrograms/L) were significantly higher in the symptomatic group (p = 0.01 and p = 0.006, respectively). Also, ECP levels were significantly lower in the group who achieved clinical remission (n = 16) in which ICS therapy was withdrawn when compared with those who needed to continue ICS prophylaxis. On the other hand, no difference was observed in the comparison of the ECP levels of children who had to restart ICS therapy and those who did not. CONCLUSION: Our results suggest that, although the determination of s-ECP levels are in accordance with clinical evaluation of disease activity, it is not useful in determining discontinuation of ICS therapy. PMID- 11270090 TI - Childhood asthma in developing countries. Low income aspects and related matters. PMID- 11270091 TI - Anaphylactic reaction to the ingestion of raw onion. A case report. AB - A case of severe systemic reactions (intense itching, urticaria, confusion, blurred vision, transient loss of consciousness, sweating, tachycardia) after ingestion of raw or lightly-cooked onion is described. The patient, a 44-year-old woman, had no troubles with well-cooked onions. Differently from the cases of sensitivity to onion described in literature, this patient was monosensitized, being skin tests negative to pollens, inhalants and other foods. The patient had 3.7 kU/L of onion-specific segum IgE, as determined by REAST. The density of onion-specific IgE (calculated as percent ratio to total IgE) was 30.8%. The reactivity of patient's serum IgE towards thermolabile and thermostable components has been tested with unheated and heated (30' at 100 degrees C) onion extracts bound to polystyrene beads and tested in the RAST system. Unheated extract resulted positive in class 2, heated extract negative, demonstrating that this patients, differently from similar clinical cases described in literature, had IgE antibodies recognizing just thermolabile onion fraction. This is the first case described in literature of a monosensitization to the thermolabile component of onion, negative also to related foods (Liliacee) and characterized by severe systemic reactions. The importance of specific-IgE density (%) rather their absolute amount (kU/L) as parameter predictive for the clinical severity of allergic reactions is discussed. PMID- 11270092 TI - Status asthmaticus: is respiratory physiotherapy necessary? AB - We present the case of a four year-old girl diagnosed of moderate extrinsic asthma that in the course of an episode of asthmatic status, she presented after treatment with respiratory physiotherapy an abrupt worsening of its clinical state, with appearance of a pneumotorax that precised intensive care treatment. The use of respiratory physiotherapy is dissuaded as part of the treatment in the initial phase of acute asthma, being reserved this treatment later in the recovery phase, anytime when a component of hypersecretion exists and the intensity of the bonchoconstriction has diminished. PMID- 11270093 TI - Spontaneous reconstruction of the canine hypogastric nerve over a long period after removing half of its length. AB - Spontaneous reconstruction of the sympathetic pathway projecting to the seminal tract after serious injury has not been well understood. Multiple cross innervation mechanisms from the spinal cord via the hypogastric nerve to the seminal tract have been demonstrated currently. This study was undertaken to explore long-term spontaneous reconstruction of the canine hypogastric nerve (HGN), which controls ejaculation, after removing half of its length. Further, the study tried to identify the crossed-pathway(s) reconstructed. Responses of the vas deferens/epididymis and bladder neck to electrical stimulation of the lumbar splanchnic nerve (LSN) or the HGN were examined. In six dogs whose hypogastric nerve was injured bilaterally as described above 4 years before, corresponding to more than 20 years in human, nine (43%) and 13 (57%) of the 21 LSNs stimulated elicited elevation of vasal and bladder neck pressure, respectively. By combining re-transection of a HGN, the following pathways to the vas deferens/epididymis were identified to have been reconstructed: (1) to the ipsilateral was deferens/epididymis without crossing to the other side; (2) to the contralateral vas deferens/epididymis by crossing to the other side at the caudal mesenteric plexus (CMP); (3) to the contralateral vas deferens/epididymis by crossing to the other side from the ipsilateral HGN at the commissural branches between the right and left pelvic plexuses (CBPP); and (4) to the ipsilateral vas deferens/epididymis by crossing twice at the CMP to the other side and at the CBPP again from the contralateral HGN to the ipsilateral side. The similar patterns of reconstruction were also observed in the bladder neck. The above results indicate that the sympathetic pathways via the HGN to the canine seminal tract can be reconstructed spontaneously in a high rate over a long period after serious injuries and that their cross-innervation system can be repaired. PMID- 11270094 TI - Interaction of calcitonin gene-related peptide (CGRP), substance P (SP) and conventional autonomic agonists in rat submandibular salivary peroxidase release in vitro. AB - Our previous immunohistochemical studies reveal that several neuropeptides, such as substance P and calcitonin gene-related peptide, innervate the major salivary glands of the mouse, rat and human. The aim of the study was to clarify their interactions by measuring their effects alone or with conventional autonomic agonists (carbachol, phenylephrine and isoproterenol) on peroxidase secretion of incubated submandibular gland slices. Calcitonin gene-related peptide evoked significant increase in peroxidase activity of the secretion only when used at 10(-5) M concentration, while substance P evoked significant, dose-dependent increase at much lower concentrations (10(-10) M). Adrenergic and cholinergic agonists enhanced peroxidase activity. Interestingly, substance P inhibited both phenylephrine and isoproterenol induced increase in peroxidase activity. Calcitonin gene-related peptide did not affect the inhibition caused by substance P. Our results demonstrate that in the salivary gland tissue substance P alone or in conjunction with adrenergic agonists result in opposing secretory responses with the doses used in vitro. Conversely, the response mediated by adrenergic receptors may be critically affected by simultaneous occupation of substance P receptors. PMID- 11270095 TI - Role of the solitary tract nucleus in mediating nociceptive evoked cardiorespiratory responses. AB - We compared the cardiorespiratory reflex responses evoked by noxious stimulation of the forelimb and cornea. Due to the depressant effects of anaesthesia on visceral reflexes we compared data from an unanaesthetised decerebrate rat model- the working heart-brainstem preparation (WHBP), with the anaesthetised rat. In both experimental models stimulation of the forelimb (mechanical pinch) evoked a tachycardia (WHBP: 19 +/- 2 bpm) and a decrease in respiratory cycle length (WHBP: from 4.1 +/- 0.2 to 2.3 +/- 0.1 s). The magnitude of response in anaesthetised animals depended on anaesthetic depth. Mechanical stimulation of the cornea evoked a bradycardia (-49.2 +/- 4.8 bpm) and an increase in respiratory cycle length from 4 +/- 0.36 to 5.88 +/- 0.2 s which was only present in the WHBP. In the WHBP activation of forelimb and corneal nociceptors both elicited significant pressor effects; in anaesthetised rats there were inconsistent changes in arterial pressure. To determine a role for the nucleus of the solitary tract (NTS) in mediating nociceptive evoked responses in the WHBP, synaptic transmission was blocked reversibly following bilateral microinjections of cobalt chloride. The heart rate responses evoked from either forelimb or corneal nociceptors were attenuated by approximately 50% (P < 0.05). A similar effect was observed using isoguvacine, a GABAA receptor agonist, to hyperpolarise NTS neurones. In conclusion, activation of forelimb and corneal nociceptors evoked contrasting patterns of cardiorespiratory response in the WHBP while in the anaesthetised rat the magnitude of the cardiorespiratory response to forelimb stimulation was quantitatively dependent on anaesthetic dose. In the WHBP, NTS neurones appear important for mediating the cardiac component of the reflex response following stimulation of nociceptive reflex pathways. PMID- 11270096 TI - Effect of stimulating non-myelinated vagal axons on atrio-ventricular conduction and left ventricular function in anaesthetized rabbits. AB - It has previously been demonstrated in several species that stimulation of myelinated vagal efferent fibres evokes slowing of heart rate and atrio ventricular (A-V) conduction and a decreased ventricular contractility but recruitment of non-myelinated fibres did not further increase the response. Only in rabbits was a significant bradycardia evoked on recruiting non-myelinated fibres. However, if stimulating myelinated fibres produced a near maximal response, then effects of further activation of non-myelinated fibres may have been missed. Indeed, selective stimulation of non-myelinated fibres alone now has been shown to evoke a slowing of heart rate independent of the effects of myelinated fibres. In the present study we tested in rabbits whether selective stimuli are also capable of slowing A-V conduction and changing ventricular contractility. In rabbits pretreated with the beta 1-adrenoceptor antagonist atenolol, ECG, arterial blood pressure, left ventricular pressure and dP/dt were recorded before and during stimulation of non-myelinated vagal efferent fibres using an anodal block technique (J. Physiol. 273 (1977) 539). R-R interval and A V conduction times were computed off-line. Stimulation of non-myelinated vagal fibres increased R-R interval by 97.7 +/- 18.8 ms from a baseline of 315.3 +/- 7.7 ms, increased A-V conduction time by 9.9 +/- 1.1 ms from a baseline of 81.9 +/- 3.1 ms and decreased left ventricular dP/dtmax by 2486 +/- 362 mmHg s-1 from a baseline of 11,186 +/- 795 mmHg s-1. When hearts were paced at a rate about 10% higher than normal, A-V conduction time still increased by 13.3 +/- 1.9 ms from a baseline of 104.2 +/- 3.6 ms and dP/dtmax still fell by 2300 +/- 188 mmHg s-1 from a baseline of 11,200 +/- 777 mmHg s-1. Ganglionic blockade with hexamethonium (15-20 mg kg-1) always abolished the evoked increases in A-V conduction time, whilst there was still an increase in R-R interval in seven of the 12 animals tested. The data demonstrate that non-myelinated vagal efferent fibres can modulate chronotropic, dromotropic and inotropic actions on the heart. PMID- 11270097 TI - Effect of prolonged head-down bed rest on complex cardiovascular dynamics. AB - We postulated that a change in complex dynamics of the cardiovascular system could be involved in the orthostatic intolerance observed after simulated weightlessness. Supine recordings of 1024 consecutive pulse intervals and systolic blood pressures were obtained on 7 subjects adapted to a 42 day head down bed rest (day 22 and 42) but also before and 6 days after head-down bed rest (-6 degrees). Coarse graining spectral analysis was used to extract the non harmonic (fractal) component from each time series. The power spectral densities of this fractal component are inversely proportional to their frequency (1/f beta). We fitted an inverse power law estimate to the fractal component to determine the spectral exponent beta. The complex dynamics of blood pressure and heart rate variability were also analyzed by correlation dimension and non-linear prediction. Bed rest induced orthostatic intolerance in 4 subjects. There was a significant increase in the spectral exponent beta of RR-interval variability during and after head-down bed rest (before: 1.039 +/- 0.090; during: 1.552 +/- 0.080 and 1.547 +/- 0.100; after: 1.428 +/- 0.040). Analysis of the blood pressure dynamics indicated lower correlation dimensions during head-down bed rest and higher coefficients of predictability after head-down bed rest. Complexity alterations of RR-interval and blood pressure variability were not linked with one another during head-down bed rest. These alterations seemed to be correlated with the orthostatic intolerance observed after bed rest. These results suggest a change of the integration level of cardiovascular autonomic regulation. PMID- 11270098 TI - Autonomic consequences of brainstem infarction. AB - OBJECTIVE: It is well known that patients with brainstem infarctions sometimes experience dizziness, vertigo and falls, although the exact mechanism is not clear. Therefore, we designed a study to quantify autonomic function in patients with brainstem infarction. PATIENTS AND METHODS: We examined autonomic function in 15 patients with brainstem infarctions, who had a history of vertigo, nausea, floating sensation and/or general fatigue during standing, and 31 age-matched controls using the composite autonomic scoring scale (CASS), which was used to grade autonomic function. The patients underwent initial autonomic assessment and then were subjected to aniracetam therapy. The drug was given orally (dose of 600 mg/day) for a duration of 56 days. Upon completion of aniracetam administration, the CASS was again tested. RESULTS: Upon initial assessment, the patients had mild reductions in mean blood pressure (MBP) and lack of an increasing heart rate (HR) within 5 min of head up-tilt, an impairment in BP correction during late phase II and reduced phase IV beat-to-beat BP response to the Valsalva maneuver, and reduced heart rate response to deep breathing (HRdb). CASS indicated mild autonomic dysfunction. After 8 weeks of treatment with aniracetam, the patients' symptoms improved and the autonomic tests showed improvement in autonomic function. CONCLUSION: Part of the pathogenesis of recurrent vertigo or dizziness with brainstem infarction might be due to mild autonomic dysfunction. Aniracetam, which activates the cholinergic system in brain, might correct the cardiovagal system in these patients. The CASS may be a sensitive tool for assessing mild autonomic dysfunction in patients with brainstem infarction. PMID- 11270099 TI - Neurogenic failures of the external urethral sphincter closure and relaxation; a videourodynamic study. AB - Urinary urgency and voiding difficulty are common features in neurological diseases, which can be attributed to dysfunction of the urethral sphincter and the detrusor. However, little is known about dynamic sphincter behaviour in neurological diseases. The present study aimed at investigating neurogenic failures of the external urethral sphincter closure and relaxation by videourodynamic study. We recruited 44 neurological patients with urinary urgency and frequency, 27 men and 17 women, mean age 61 years, and 28 of them had voiding difficulty as well. None had abnormal finding of digital examination or ultrasound echography of the pelvic organs. Using triple-lumen 7F catheter under X-ray fluoroscope, we measured detrusor pressure, external urethral sphincter pressure (Pura) and external sphincter EMG in all patients. We also performed pressure-flow study and obtained the Abram-Griffiths (AG) number, a numerical grade of obstruction. During filling 30 had detrusor hyperreflexia. EMG cystometry showed uninhibited external sphincter relaxation (UESR) in eight patients, seven of whom had detrusor hyperreflexia as well. Patients with UESR showed an abnormal reduction of Pura, mean reduction 64 +/- 27 cmH2O (mean +/- standard deviation). During UESR the Pura and EMG activity fluctuated, and fluoroscopic image showed bladder neck opening in four with extreme urge sensation, including one without detrusor hyperreflexia. During an attempt of voiding three patients with voiding difficulty had detrusor-external sphincter dyssynergia (DESD) with detrusor contraction and eight had unrelaxing external sphincter without detrusor contraction. Fluoroscopic image showed an incomplete or absent urethral opening at the external sphincter. Four of them had severe straining on voiding together with intermittent increment of EMG activity without a normal funneling of the bladder neck. The mean reduction of Pura during voiding was 6.4 +/- 6.7 cmH2O and 5.0 +/- 9.5 cmH2O (in women and men, respectively) with DESD or unrelaxing external sphincter which was less than 39 +/- 25 cmH2O and 53 +/- 47 cmH2O in those without (P < 0.01). The mean AG number was 15 +/- 21 and 51 +/- 19 (for women and men, respectively) with DESD or unrelaxing external sphincter which was larger than 6.2 +/- 34 and 35 +/- 22 in those without (P < 0.05). In conclusion, UESR and DESD/unrelaxing external sphincter could be a factor for urinary urgency and voiding difficulty in neurological patients, evidence of central dysregulation affecting the Onuf's nucleus and its fibres to the external urethral sphincter. PMID- 11270100 TI - [Morphofunctional changes in the olfactory bulb after odor stimuli deprivation (II). Histochemical study]. AB - By histochemical and immunohistochemical methods we study the effects that has the odorous stimuli privation in the first day after the birth on the activity of the olfactory bulb of the gerbil (Meriones Unguiculatus). This sensory deprivation of stimuli causes drastic reductions in given activities (Tyrosine Hydroxylase) no altering other (DOPA-Decarboxylase). This indicates that the functional ripeness of given enzymatic systems specify of an integrity of their sensorial afferences, while it is not necessary for the normal activity of other enzymes even within the same cell. PMID- 11270101 TI - [Health measurement instruments in patients with vertigo]. AB - OBJECTIVE: Compare disability and handicap in patients with dizziness by means of two questionnaires. STUDY DESIGN: Prospective study. PATIENTS: 337 patients seen for non-acute dizziness from peripheral or central origin in a tertiary referral setting. MAIN OUTCOME MEASURES: Spanish version of the Dizziness Handicap Inventory test and UCLA-Dizziness Questionnaire after transcultural adaptation following the method of translation-back-translation. RESULTS: We obtained a good correlation between the frequency of dizzy spells and quality of health as perceived by the patient; also there was good correlation between the intensity of each spell and limitation for performance of daily activities. Quality of life is mainly related to handicap in these patients. CONCLUSION: Vertigo, as a non fatal health outcome, can be studied following the two main conceptual frameworks of Impairment, Disabilities and Handicap and Health-Related Quality of Life. PMID- 11270102 TI - [Identification of respiratory syncytial virus and adenovirus in children with chronic serous otitis]. AB - OBJECTIVE: To study the presence of respiratory syncytial virus (RSV) and adenovirus in the effusions of the middle ear and adenoid tissue from patients with chronic otitis media with effusion by use of the polymerase chain reaction. METHOD: The effusions and adenoid samples were collected from 32 children undergoing myringotomy and ventilation tube placement for chronic otitis media with effusion. The samples were separated for viral culture, immunofluorescence and PCR analysis. RESULTS: One (3%) effusion sample was positive for adenovirus by the PCR. None of samples (effusion and adenoid) were positives for RSV. The results of viral culture and immunofluorescence were all negative. CONCLUSIONS: Our results can not support that respiratory viruses play a role in the pathogenesis of chronic otitis media with effusion. PMID- 11270103 TI - [Experience in the treatment of 98 carcinomas of the nasopharynx. Long-term follow-up and analysis of prognostic factors]. AB - This was a retrospective study of 98 patients (pts.) with histologically confirmed nasopharyngeal carcinoma. The clinico-demographic characteristics were: median age of 53 years (11-83); 74 males and 24 females (ratio 3:1); histology subtype OMS 2-3 in 89 pts. (90.8%); cranial nerve deficits in 11 pts. (11.2%); 50 (51%) were stage T3T4; 68 pts. (69.4%) N2N3 and 77 pts. (78.6%) stage IV. The therapeutic modalities were: radical radiotherapy (RT) alone in 42 pts., chemotherapy (CT) alone in 4 pts., RT + adjuvant CT in 10 pts. and neoadjuvant CT + RT in 42 pts. RT was delivered in wide fields, doses between 50-75 Gy with conventional fractionation. CT consisted in cisplatinum-based schedules (PF in 34 pts., BEC in 9 and others in 13 pts.). Analyzed by treatment, more males and stages N2N3 and IV were accrued in neoCT + RT arm (p < or = 0.05). For the entire population, the overall complete response was achieved in 65 pts. (66.3%); in 27/35 pts. (77.1%) of the RT group and 30/51 pts. (58.8%) of CT + RT group (p 0.07) of pts. with III-IV stages. With a median follow-up of 74.5 months, 32 pts. (32.65%) are alive and free of disease. The projected OS for all pts. was 40 months (m), 51.4% at 3 years (y) and 45.5% at 5 y with a disease free survival of 37 m (0-236). No differences between treatment arms were found (p 0.4). In univariant analysis for OS in stage III-IV pts., age > 50 y, histology OMS1, cranial nerve deficits, stage T3T4 and N2N3, were considered adverse prognostic factors (p < or = 0.05). In multivariant analysis, only age > 50 y and stages T3 T4, N2-N3 were significant (p < or = 0.05). In conclusion, we demonstrated good long term survival without any differences among treatment modalities in pts. with advanced nasopharyngeal carcinomas. New therapeutic approaches are warranted in order to improve the outcome of this patients. PMID- 11270104 TI - [Epistaxis: prospective study on emergency care at the hospital level]. AB - We prospectively studied 279 patients with epistaxis referred to ENT specialist from emergency room in our hospital in one year. Masculine sex (62%), medium or old ages (median 56 year), and associated diseases (HBP 22.9%, anticoagulants or antiplatelet drugs 11.1%), were the most common. Predominant local etiologic factors were trauma (12.9%) and inflammation (14%). Epistaxis had been essential in 36.9%. The most frequent location in all ages was the anterior one, especially located in the Kiesselbach's area, although the incidence of the posterior epistaxis increases in characteristic way from fourth decade, affecting more to men. Most of epistaxis (> 99%) were treated successfully by conservative approach and only two patients required surgical or interventional therapy. Patients between 30-59 year and the posterior epistaxis was the most refractory to the treatment. Hospitalized patients were older than ambulatory ones and had more associated diseases. Their average hospital stay was 9.2 days. PMID- 11270105 TI - [Ambulatory septopyramidal surgery: methods, results and patient satisfaction]. AB - Outpatient surgery is an activity that increases constantly in Otolaryngology. There are studies about some nasal surgery techniques, such as septoplasty or endonasal endoscopic performed as outpatient surgery, but so far nobody has reported any comparative analysis on the results of outpatient septorhinoplasty, although this is a frequent practice. We report our experience with 40 cases of outpatient septoplasty and 40 outpatient septorhinoplasty, performed by the same surgeon, and we make a comparative study with the same number of operations performed on in-patients with traditional surgical techniques. Aesthetic and functional results and satisfaction level in postoperative interviews were compared in both cases. No serious complications were present in either group. The number of complications, the functional and aesthetic results and the degree of patients satisfaction were similar in both groups. Septopyramidal surgery (septoplasty and septorhinoplasty) is suitable to be performed on outpatient with the same quality as in-patients. PMID- 11270106 TI - [Nasal mucosa and systemic repercussion of the topical and prolonged use of budesonide in patients with perennial allergic rhinitis]. AB - In a prospective study, measurement of a 9.00 a.m. basal serum cortisol and biopsies of the inferior turbinate mucosa were taken from 40 patients using topical nasal corticosteroid (Budesonide) continuously for month or years. No systemic adverse effects and no histopathological changes of significance were found. These findings do no suggest that topically corticosteroids are harmful to the nasal mucosa and no produce systemic effects. PMID- 11270108 TI - [Diagnostic-therapeutic approach to internal jugular thrombosis]. AB - The internal jugular vein thrombosis is a rare and critical disease. The infection of the head and neck deep spaces it is not the main cause since the antibiotic era. The ENT neoplasms and iatrogenic etiology are more frequent. Anodyne symptoms and vital complications are important to obtain an early diagnose and therapy, to improve the prognosis. The objective of this review is to obtain a certainly therapeutic criteria in the approach to this entity. PMID- 11270107 TI - [Detection tonsillar pathology by superoxide dismutase levels. Comparative study of surgical indications for tonsillectomy]. AB - If oxygen-derived free radicals are considered the definitive cause of tonsillar damage after infection, it seems reasonable that scavenger antioxidants levels could be used as a detector of tissue impairment. So, superoxide dismutase (SOD) amounts were measured in palatine tonsils and peripheral blood on subjects bearing of hypertrophy without infection (H, n = 83), recurrent tonsillitis (RA, n = 75), and peritonsillar abscess (PA, n = 12). SOD levels in both tonsillar cultures supernatants and peripheral blood erythrocytes were detected progressively increased in groups with H, RA and PA, which were statistically significative (ANOVA-test; p < 0.001). A significative correlation between tissue and blood was observed for all the groups. We can conclude that SOD concentration in palatine tonsils and/or peripheral blood increases proportionally to infections incidence, which allows detecting patients with functional damage, and recommending objectively tonsillectomy or at least monitoring clinical response for a therapy. Practical use and results obtained from comparison to tonsil biopsies are discussed. PMID- 11270109 TI - [Cholesterol granuloma of the middle ear: cause of idiopathic hemotympanum]. AB - We report a case of a 49-year-old man with a cholesterol granuloma of the middle ear in an only hearing ear. As it mimicked a vascular middle ear tumor, a contrast cranial computed tomography and a gadolinium-enhanced-magnetic resonance imaging with vascular reconstructions were performed, confirming that both carotid artery and jugular vein were near the lesion but not involved. This patient could be managed with a conservative hearing preserving approach. Differential diagnosis of vascular middle ear anomalies is reviewed, specially in relation to cholesterol granulomas as a cause of idiopathic hemotympanum. PMID- 11270110 TI - [Mucoepidermoid carcinoma of the nasal vestibulum]. AB - Mucoepidermoid carcinoma is a malignant epithelial tumor, with a mucosecretor component that is exceptional in the upper airways. It seems to be a kind of immunity in this area to the development of this tumors. In the review of the literature we only have found three cases in the nasal cavity. We present the case of a 18th years old woman with a tumor in the right nostril for two months. Several biopsy were required to achieve its histopathological diagnosis as a high malignant mucoepidermoid carcinoma. Radical surgery over the tumor and metastasis was performed but the case has a bad prognosis. The prognosis of the mucoepidermoid carcinoma depends on the histological grade, the early diagnosis and the surgical treatment which has to be so radical as possible to obtain free tumors limits. PMID- 11270111 TI - [Pansinusitis secondary to inflammatory myofibroblastic tumor]. AB - Inflammatory myofibroblastic tumors are aetiologically enigmatic and biologically unpredictable lesions. The lungs are the organs of apparent predilection. These tumors have also been documented in a number of extrapulmonary sites including head and neck. Paranasal location is rare. We report a case of a maxillary location in a 10 years and 10 months old girl initiated as an acute pansinusitis. PMID- 11270113 TI - [Lingual involvement in progressive systemic sclerosis]. AB - A case of a 40 year old female patient who began to present alterations in the tongue and who was diagnosed of Progressive Sclerosis of that organ is presented. The manifestation and diagnostic tools used in this case, as well as the treatment use, is given. The rareness of this disease, which is possible in systemic diseases, stands out. PMID- 11270112 TI - [Peritonsillar abscess with parotid and peripharyngeal extension. Report of a case]. AB - Peritonsillar abscess is the most frequent complication of a tonsillar infection. The purulent material can spread from peritonsillar space to the fascial neck spaces. These deep neck infections may be a life threatening complication. The correct treatment of these infections is an appropriate antimicrobial therapy and abscess drainage, most of the times by surgery. We present a clinical case of a large perypharyngeal, and parotid abscess originated from a peritonsillar abscess. It was necessary surgical treatment to drain it. PMID- 11270115 TI - [Lingual hamartoma. Report of a case]. AB - The hamartomas that are located in the lingual region are very strange tumours. There are very few published in the world literature. A patient that presented a hamartoma was diagnosed and treated in our department. PMID- 11270114 TI - [Basosquamous carcinoma of the floor of the mouth]. AB - Basaloid-squamous cell carcinoma is a rare tumor characterized by Wain et al in 1986 as a separate pathological entity. In head and neck more than 100 cases have been reported, with a strong predilection for hypopharynx, base of tongue, and supraglotic larynx. This entity also has been described in nasal cavity, oesophagus, tonsils and oral cavity. Only 14 cases has been reported arising in the floor of the mouth, and none in our literature. We present a new case in floor of the mouth and clinicopathological features are emphasized. We have undertaken moreover a bibliographic review of this uncommon and aggressive neoplasm. PMID- 11270116 TI - [Localized laryngeal amyloidosis. Endoscopic surgery with CO2 laser]. AB - Laryngeal amyloidosis is a very rare disease. In the upper airways, the most common localization is the laryngeal organ, particularly the supraglotic region. Usually laryngeal amyloidosis is a localized primary form, although it has occasionally been reported in conjunction with system amyloidosis. We are reporting a new case of localized primary laryngeal amyloidosis in the right arytenoid mucosa in a 66 years old woman who underwent CO2 laser endoscopic surgery. PMID- 11270117 TI - [Branchial cysts with heterotopic salivary tissue in the upper third of the neck]. AB - The presence of heterotopic salivary gland tissue (HSGT) in rather uncommon in the neck. Usually it has been located in its lower third. HSGT in the upper neck is believed to be a rare entity. Two cases of HSGT in the upper neck are presented, with a review of their clinical and histopathological characteristics. The embryologic derivation of salivary tissue and close association with the branchial apparatus are discussed. The possibility of neoplastic transformation must always be considered in these uncommon lesions. PMID- 11270118 TI - Altered prestarvation response in a nystatin resistant Dictyostelium discoideum mutant. AB - Wild-type Dictyostelium amoebae secrete an autocrine, prestarvation factor (PSF) that allows them to measure the amount of food bacteria compared to their cell density. When the ratio of PSF to bacteria reaches a threshold, the cells are signaled to prepare for eventual starvation. This prestarvation response (PSR) usually starts three to four generations before the end of exponential growth, leading to the accumulation of several aggregation specific genes during growth. We characterize a nystatin-resistant mutant, HK19, that expresses the PSR genes three generations earlier than wild type but has an otherwise wild-type PSR. Although HK19 has a full PSR during growth, HK19 continues to grow at the wild type rate and reaches normal cell densities. Because HK19 temporally separates the PSR from starvation, it became possible to test whether starvation is required for development. Since HK19 growing at low density can be induced to clump with either cAMP or folate, it appears that the PSR and an external signal are sufficient for entry into development. These data suggest that the PSR is a complex genetic pathway that induces genes involved in the exit from growth and the entry into development. PMID- 11270119 TI - Biphasic expression of rnrB in Dictyostelium discoideum suggests a direct relationship between cell cycle control and cell differentiation. AB - Cell differentiation in Dictyostelium is strongly affected by the cell cycle. Cell cycle control is well-understood in other systems, but this has had almost no impact on the study of Dictyostelium cell differentiation, in part because the cell cycle in Dictyostelium is unusual, lacking a G1 phase. Here we describe the cell-cycle regulated expression of rnrB, which codes for the small subunit of ribonucleotide reductase and is a marker of late G1 in many systems. There appear to be two expression peaks, one in mid-G2 and the other near the G2/M transition. Using Xgal/anti-BrdU double staining, we show that cells in asynchronously growing cultures express in both phases, with a gap between them during which the gene is transcriptionally silent. Cold-synchronized cells show exclusively G2/M expression, while mid-G2 expression is seen in high-density synchronized cells and can also be inferred in cells undergoing synchronization by either method. rnrB expression occurs in other systems shortly after cells pass a point (the "restriction point" or "start") at which they commit to complete their current cell cycle. We demonstrate a similar commitment point in Dictyostelium and show that this occurs shortly before the mid-G2 rnrB expression peak. The Dictyostelium cell cycle thus appears to include a well-defined though inconspicuous event, between early and mid-G2, with some features which are normally associated with the G1/S transition. Others have described a switch from stalk to spore differentiation preference at about this time. Since Dictyostelium cells switch back from spore to stalk preference approximately at the G2/M rnrB expression maximum, cell differentiation as well as rnrB expression may be regulated directly by fundamental cell cycle control processes. PMID- 11270121 TI - Visualization of endogenous BMP signaling during Xenopus development. AB - The TGF-beta superfamily of growth factors is known to transmit signals to the nucleus mainly through the Smads, intracellular signaling components that are highly conserved from nematodes to humans. The signaling activity of the Smads is regulated by their ligand-stimulated phosphorylation through Ser/Thr kinase receptors. Here, to examine the in vivo role of BMP, we investigated the spatio temporal activation of BMP-regulated signals during Xenopus development, using a polyclonal antibody that specifically recognizes the phosphorylated form of BMP regulated Smads. BMP signaling was observed uniformly in embryos as early as stage 7, but was restricted to the ventral side of the embryo at the late blastula stage, supporting the proposed role of BMP4 as a ventralizing factor in Xenopus embryos. In addition, localized staining was detected in several developing organs, consistent with the predicted function of BMP family members in organogenesis. PMID- 11270122 TI - Lactation affects expression of intermediate filaments in human breast epithelium. AB - The human breast contains two epithelial lineages, luminal epithelial and myoepithelial. Specific patterns of expression of intermediate filaments have previously been demonstrated in the resting breast. To determine how terminal differentiation and lactation influenced expression of intermediate filaments in breast epithelial cells, we used Western blot analysis to measure the levels of vimentin, alpha-smooth muscle actin, keratin 14, and keratin 18 in the resting and lactating breast. Confocal immunofluorescence was used to determine the subcellular site of localization of the intermediate filaments. Vimentin was localised to myoepithelial cells in both the resting and lactating gland. There was a four-fold increase in vimentin protein levels in lactating tissue relative to resting tissue, and this may be related to increased cellular activity of the myoepithelial cells which surround secretory alveoli. Alpha-smooth muscle actin and keratin 14 were detected in myoepithelial cells, and similar levels of expression were found in lactating and resting tissue. In the resting breast, keratin 18 and keratin 8 were detected in luminal epithelial cells in a filamentous form, whereas in lactating tissue it was present in a punctate form in luminal cells and also seen as granules in the lumen of alveoli. Our results indicate that intermediate filament expression patterns are altered in the lactating human breast, and this may reflect their role in the fully functional gland. PMID- 11270120 TI - Interaction of gdt1 and protein kinase A (PKA) in the growth-differentiation transition in Dictyostelium. AB - The gdt1 gene is a negative regulator of the growth-differentiation-transition (GDT) in Dictyostelium. gdt1- cells express the GDT marker discoidin earlier and at higher levels and prematurely enter the differentiation pathway. Protein kinase A is a positive regulator of the GDT and is required for multicellular development. Disruption of the PKA catalytic subunit or overexpression of a constitutively active mutant of the regulatory subunit results in cells which do not form multicellular aggregates and which show strongly reduced levels of discoidin. We have created PKA-/gdt1- double mutants and show that these display high levels of discoidin expression but no aggregation, suggesting that gdt1 may be a downstream target of PKA in a branched signaling cascade initiating differentiation. Data obtained with the PKA inhibitor H89 support these result: in wild type cells H89 inhibits discoidin expression while in gdt1- mutants there is no obvious effect. However, since PKA-/gdt1- cells display less discoidin expression than the single gdt1 mutant, we propose that PKA and gdt1 are in two parallel interacting pathways. To get insight into the mechanism how PKA may block gdt1, we have tested two putative PKA phosphorylation sites in the protein and found that one of them is efficiently phosphorylated by PKA in vitro. A model for the interplay between PKA and gdt1 during the growth-differentiation transition is discussed. PMID- 11270123 TI - Phenotypic modulation of arterial smooth muscle cells is associated with prolonged activation of ERK1/2. AB - Arterial smooth muscle cells grown in primary culture on a substrate of fibronectin in serum-free medium are converted from a contractile to a synthetic phenotype. This process is dependent on integrin signaling and includes a major structural reorganization with loss of myofilaments and formation of a large secretory apparatus. Functionally, the cells lose their contractility and become competent to migrate, secrete extracellular matrix components, and proliferate in response to growth factor stimulation. Here, it is demonstrated that the mitogen activated protein kinases ERK1/2 play a vital role in the fibronectin-mediated modification of rat aortic smooth muscle cells. Immunoblotting showed that phosphorylated ERK1/2 (p44/p42) were expressed throughout the period when the change in phenotypic properties of the cells took place. Moreover, phosphorylated ERK1/2 accumulated in the nucleus as revealed by immunocytochemical staining. Additional support for an active role of ERK1/2 in the shift in smooth muscle phenotype was obtained by the finding that PD98059, an inhibitor of the upstream kinase MEK1, potently suppressed both the expression of phosphorylated ERK1/2 and the fine structural rebuilding of the cells. In conclusion, the observations point to an important and multifaceted role of ERK1/2 in the regulation of differentiated properties and growth of vascular smooth muscle cells. PMID- 11270125 TI - Pulmonary embolism in the elderly. AB - The elderly are at increased risk for pulmonary embolism because of both the conditions common to this age group, and the immobility that often accompanies them. Whether aging alone represents a hypercoagulable state is unclear. The incidence of pulmonary embolism rises with age, however, as does pulmonary embolism mortality. The diagnosis of pulmonary embolism is difficult and frequently missed because elderly patients and their physicians may attribute nonspecific symptoms to underlying cardiopulmonary disease or to age itself. Routine laboratory examinations are also nonspecific. Lower extremity studies to diagnose DVT should always be pursued because a positive study results in identical treatment, without the need for further testing. D-dimer concentrations are useful when low, but are commonly elevated in the elderly because of other comorbid conditions. Lung scanning remains the most common initial study to diagnose pulmonary embolism, although spiral CT is as sensitive and specific. Pulmonary angiography should always be considered when the initial studies are nondiagnostic and clinical suspicion is high, and this test is well tolerated in the elderly. The role of newer diagnostic techniques, such as MR imaging, cannot be determined until well-designed outcomes trials are completed. Prophylaxis with appropriate pharmacologic agents or mechanical measures should be administered not only to patients undergoing hip or knee reconstruction surgery, but to all bed-ridden elderly medical and general surgery patients. Treatment for pulmonary embolism with anticoagulation reduces the mortality rate and should be administered in all elderly patients without contraindications. In addition, thrombolysis should be considered for all hemodynamically unstable patients with pulmonary embolism, regardless of age. Vena caval filters are warranted when anticoagulation is contraindicated, although evidence of the long-term benefit of these devices is lacking. At present, pulmonary embolism is underdiagnosed and undertreated in the elderly. By heightening awareness of this diagnosis and its appropriate management in this age group, considerable morbidity and mortality may be avoided. PMID- 11270124 TI - Oral anticoagulants. Pharmacologic issues for use in the elderly. AB - There has been a marked expansion of the indications for oral anticoagulant therapy, particularly among the elderly. Despite the documented benefits, the use of warfarin remains strikingly low among patients 80 years of age and older. Elderly patients often exhibit an enhanced dose response to warfarin. On average, steady-state warfarin doses decrease by 11% per decade of age. Pharmacokinetic changes in the elderly are negligible. Pharmacodynamic differences have not been well characterized. Initiating warfarin dosing in the elderly should be done cautiously, with doses of 5 mg or less. Doses should be adjusted downward in the presence of congestive heart failure, advanced obstructive lung disease, liver disease, malignancy, protracted diarrhea, enteral feedings, or concurrent potentiating medications. Numerous medications interfere with the anticoagulant response of warfarin. The most powerful potentiating drugs are those that interfere with the metabolism of (S)-warfarin. Examples include amiodarone, trimethoprim-sulfamethoxazole, and metronidazole. These drugs should be prescribed with caution in the elderly and mandate frequent INR monitoring during the induction period. An extensive assessment of patient-specific factors that might increase the hazards related to warfarin therapy needs to be conducted and documented before initiating oral anticoagulant therapy. Patients and their caregivers need to understand the risks and benefits, and to recognize signs of abnormal bleeding and the need for frequent monitoring. Patients should be encouraged to maintain consistency in their vitamin K intake and should strive to meet the recommended dietary allowance for vitamin K. To improve anticoagulation control, physicians and other health care providers need to be aware of the many warfarin drug interactions and be cognizant of the increased dose response of warfarin seen in the elderly. Concurrent prescription of multiple drugs known to affect warfarin's anticoagulant response should be minimized and use of nonselective nonsteroidal anti-inflammatory drugs should be limited given their deleterious effects on the gastric mucosa. Transitions from inpatient care to subacute care and back to outpatient care are particularly vulnerable periods for patients' anticoagulation control. Enhanced provider communication and more seamless transitions help to ensure optimal INR follow-up and timely warfarin dose adjustment if indicated. PMID- 11270126 TI - Antithrombotic therapy in atrial fibrillation. AB - Atrial fibrillation is a common condition affecting elderly individuals; as many as 10% of people older than age 80 years have AF. AF is also a potent risk factor for ischemic stroke, raising the risk of stroke fivefold. A set of consistent randomized controlled trials has demonstrated that long-term anticoagulation can largely reverse the risk of stroke attributable to AF. In these trials, anticoagulation generally proved quite safe, raising the risk of intracranial hemorrhage by less than 0.5% per year. The anticoagulation target for AF is INR 2 to 3 with INR 2.5 as the specific goal. The trials were much less consistent about the efficacy of aspirin, although it seems that aspirin has a small stroke preventive effect. The recommended dose of aspirin is 325 mg per day. Because it raises the risk of hemorrhage and adds the burden of frequent monitoring of INR values, anticoagulation is recommended for those patients with AF at higher risk of stroke. Such higher risk is conferred by the following risk factors: (1) a history of a prior stroke, TIA, or other systemic embolic event; (2) a history of hypertension; (3) diabetes mellitus; (4) left ventricular dysfunction; (5) mitral stenosis; and (6) older age. The exact age threshold conferring sufficiently increased risk is uncertain, with some research indicating the threshold should be age 65 years, and other research indicating the threshold should be age 75 years. For lower-risk patients, aspirin is recommended. Future research should focus on the oldest patients with AF. These individuals face the highest risk of ischemic stroke without anticoagulation and the highest risk of major hemorrhage with anticoagulation. Only small numbers of such elderly patients were included in the randomized trials. Future research should also focus on improved risk stratification, allowing better targeting of anticoagulation. Discoveries of new antithrombotic agents and new drugs and devices for preservation of sinus rhythm could radically improve stroke-preventive strategies for AF. PMID- 11270127 TI - Antithrombotic and thrombolytic therapy for ischemic stroke. AB - Antithrombotic and thrombolytic agents form the cornerstone of stroke prevention and treatment. Large, randomized trials have also highlighted the effectiveness and safety of early and continuous antiplatelet therapy in reducing atherothrombotic stroke recurrence. Aspirin is the antiplatelet treatment standard against which several other antiplatelet agents (ticlopidine, clopidogrel, aspirin-dipyridamole) have been shown to be more effective. The prevention of cardioembolic stroke is best accomplished with oral anticoagulation, barring any contraindications. The thrombolytic agent, rt-PA, improves outcome in ischemic stroke patients treated within 3 hours of onset. The risk-benefit ratio is narrow because of an increased risk for bleeding but studies do not support a higher risk in the geriatric population. PMID- 11270128 TI - Direct and indirect antithrombins. Heparins, low molecular weight heparins, heparinoids, and hirudin. AB - With the eclipse of UH by newer anticoagulants, the field has opened up to search for new and better drugs. Hirulog or bivalirudin is another direct antithrombin that has been used in initial trials. It is smaller than hirudin, at 20 amino acids. Currently under investigation, it seems to have a short half-life, a narrow therapeutic window, and a reverse dose effect, with lower levels achieving better cardiac post-thrombolysis patency than higher doses. Other antithrombins being examined are the hirudisins, where four amino acids of hirudin have been replaced by the RGDS integrin-binding sequence and thrombin receptor antagonist peptides. In addition, many other inhibitors of activated clotting factors are being studied for future therapeutic value. Tick anticoagulant protein studies are underway, as are studies on a group of benzamidine isoxazoline derivates, which are direct Xa inhibitors. We are truly at an age of discovery with the newer anticoagulants and it may take many years until we can distinguish the advantages and disadvantages of all the newer therapies. It looks like an ever more real possibility that medicine may find an antithrombotic regimen that is highly effective, highly reversible, and nontoxic. PMID- 11270130 TI - Thrombolysis and antithrombotic therapy for coronary artery disease. AB - Aspirin in a dose of 160 to 325 mg should be administered on day 1 of an acute MI and continued indefinitely on a daily basis. Thrombolytic therapy should be administered within 6 to 12 hours after the onset of an acute MI with ST segment elevation or with left bundle branch block. Primary coronary angioplasty when available should be used rather than thrombolytic therapy in the treatment of older persons with acute MI who are poor candidates for thrombolytic therapy. Intravenous heparin should be given in persons with acute MI undergoing primary coronary angioplasty or surgical coronary revascularization and in persons with acute MI at high risk for systemic embolization. Long-term oral warfarin should be given after MI for the secondary prevention of MI in post-MI persons unable to tolerate daily aspirin, in post-MI persons with persistent atrial fibrillation, and in post-MI persons with left ventricular thrombus. Platelet GP IIb/IIIa inhibitors should be administered along with aspirin and enoxaparin in the acute phase of management of persons with unstable angina pectoris or non-Q wave MI. Aspirin should be administered daily indefinitely to persons after MI, to persons with unstable angina pectoris, to persons with stable angina pectoris, and to persons undergoing coronary revascularization. Aspirin plus ticlopidine or aspirin plus clopidogrel should be used in persons undergoing coronary artery stenting. Platelet GP IIb/IIIa inhibitors should be used at the time of coronary angioplasty, coronary atherectomy, or coronary stenting. Aspirin, 160 to 325 mg daily, is recommended in older men and postmenopausal women who are at high risk for developing coronary events in addition to treating their coronary risk factors. PMID- 11270129 TI - Antithrombotic therapy in valvular heart disease. AB - Whether antithrombotic therapy in elderly patients differs from that in younger populations depends on whether the risk for such bleeding differs in the elderly. Regarding long-term therapy with warfarin derivatives, evidence shows that there is a difference. The anticoagulation response to warfarin is exaggerated with advancing age. This article discusses antithrombotic therapies for valvular heart disease, including bioprosthetic and mechanical prosthetic heart valves, aspirin and dipyridamole in combination with oral anticoagulant therapy, antiplatelet agents alone or in combination with very low dose warfarin, tilting disk valves, valve position, first-generation valves compared with modern valves, interruption of anticoagulant therapy, and miscellaneous indications for antithrombotic therapy. PMID- 11270131 TI - Perioperative management and reversal of antithrombotic therapy. AB - Patients of advanced age commonly undergo invasive procedures and surgery. With the number of elderly individuals being treated with long-term anticoagulant therapy growing annually, it is not uncommon that surgery is contemplated for older adults on long-term anticoagulant therapy. This article focuses on the management of elderly patients who are on long-term anticoagulant therapy, principally with warfarin, who must undergo invasive procedures. Although no consensus has been reached regarding the perioperative management of patients on long-term anticoagulation therapy, this discussion presents the current status and some recommendations for practice. PMID- 11270132 TI - Antiplatelet therapy in the elderly. Aspirin, ticlopidine-clopidogrel, and GPIIb/GPIIIa antagonists. AB - Antiplatelet agents including aspirin, dipyridamole, the thienopyridines, and the GPIIb/IIIa antagonists have collectively demonstrated their ability to have a significant impact on the incidence of recurrent MIs, strokes, and other vascular ischemic events in the geriatric population. Low-dose aspirin also seems to be effective and safe for the primary prevention of ischemic heart disease in men considered at high risk. There is no evidence that the recommendations from these studies had increased relevance to younger adults, and the studies considering age as a variable found antiplatelet agents had either similar or increased benefit in older patients. In view of the relatively reduced adverse effects of these agents when compared with their potential therapeutic benefit, it is important that they be considered in all older patients for secondary prevention and in certain high-risk groups for primary prevention of cardiovascular morbidity and mortality. PMID- 11270133 TI - Hemorrhagic complications of oral anticoagulant therapy. AB - Hemorrhage is the major complication of anticoagulant therapy. The criteria for classifying the severity of bleeding has varied between studies, which has resulted in variability in the rate of bleeding reported in the literature. The major determinants of oral anticoagulant-related bleeding are the intensity of the anticoagulant effect, baseline patient characteristics, and the length of therapy. Older patients have characteristics that may place them at higher risk for anticoagulant-related bleeding, but they also have characteristics that make them more likely to benefit. The risk for anticoagulant-related bleeding cannot be considered in isolation and the potential benefits need to be weighed carefully in each individual patient, irregardless of age. PMID- 11270134 TI - Clinical evaluation of hypercoagulable states. AB - Although thromboembolic disease associated with primary thrombophilic conditions most often presents before age 50 years, older adults can be affected by such disorders. This article addresses congenital and acquired prothrombotic states, with specific attention to the likelihood of such disorders occurring in the geriatric patient population. Recommendations for testing and therapy are reviewed. PMID- 11270135 TI - Prevention of venous thromboembolism. AB - Recognition of the patient at risk and delivery of appropriate prophylaxis are imperative to reduce the incidence of VTE (including fatal PE) in hospitalized patients. This article provides estimates of risk and a summary of effective prophylaxis methods such that physicians can tailor an individual patient's prophylaxis regimen to maximize DVT risk reduction in the safest and most cost effective manner. PMID- 11270136 TI - Treatment of deep venous thrombosis and pulmonary embolism with low molecular weight heparin in the geriatric patient population. AB - Low-molecular-weight heparins have provided a new approach to treating DVT-PE. These anticoagulants have better bioavailability, a longer half-life, and a more predictable antithrombotic effect than UFH. In the studies reviewed, LMWHs were shown to be safe and effective in preventing recurrent thrombotic events when compared with the more precise UFH dosing schedules. There has been no evidence of an increased incidence of major bleeding or recurrent thromboembolic events based on age in these trials. With these findings LMWHs given subcutaneously, without laboratory monitoring, in a dose determined by actual body weight allows clinicians involved in the care of the elderly to manage these patients with DVT with or without PE in the nursing home setting, skilled nursing care facilities, rehabilitation units, acute care hospital setting, or as described in this article at home. These patients can continue their physical therapy programs because intravenous infusion lines and the necessity to be maintained at bedrest do not encumber them. PMID- 11270137 TI - Transduction capabilities of neonatal ventricular afferent neurons in vivo. AB - We sought to determine the capacity of ventricular sensory nerve endings (neurites) associated with neonatal nodose ganglion afferent neurons to transduce mechanical and chemical stimuli in situ. Spontaneous activity generated by 17 nodose ganglion cardiac afferent neurons was identified in 8 anesthetized neonatal pigs (10-21 days old) using extracellular recording recording techniques. The activity generated by afferent neurons was studied when their ventricular sensory neurites were exposed to local mechanical or chemical stimuli, following systemic administration of specific chemicals or during brief periods of apnea. Gentle mechanical distortion of their ventricular sensory fields enhanced the activity generated by 6 spontaneously active afferent neurons, while suppressing the activity generated by another 3 neurons. Afferent neuronal activity was either enhanced or suppressed when the following chemicals were applied to identified ventricular epicardial sensory fields: the sodium channel modifier veratridine (92% of tested neurons); the P1-purinoceptor agonist adenosine (92%); the neuropeptides angiotensin II (100%), bradykinin (90%) and substance P (90%); and the nitric oxide donor S-nitroso-N-acetylpenicillamine (100%). Epicardial application of isoproternol or nicotine induced modest neuronal responses. Cardiac afferent neurons were also affected when these chemicals were administered systemically. Apnea of 60-100 s duration modified (enhanced, n = 2; suppressed, n = 5) the activity generated by most identified afferent neurons. The estimated average conduction velocity of afferent axons associated with these neurons was 1.0 +/- 0.2 m/s. It is concluded that neonatal nodose ganglion cardiac afferent neurons respond to many of the chemicals known to modify adult cardiac afferent neurons. That cardiac afferent neurons are capable of sensing the mechanical and chemical milieu of the neonatal heart should be taken into account when considering altered neonatal cardiovascular status such as occurs during apnea. PMID- 11270138 TI - Novel dual 'small' vesicle model of ATP- and noradrenaline-mediated sympathetic neuromuscular transmission. AB - The main question asked was if sympathetic nerves in guinea-pig vas deferens release the co-transmitters ATP and noradrenaline from the same, or different vesicles, i.e. in fixed combinations or independently. The extracellularly recorded excitatory junction current (EJC) and the fractional increase in overflow of tritium (delta T) were used to monitor the per pulse secretion of ATP and [3H]NA, respectively, during electrical stimulation with 1-3000 pulses at 0.1 40 Hz. The frequency- and train length-dependence and alpha 2-adrenoceptor mediated autoinhibition of these parameters, and of the ATP-mediated twitch contraction, were compared first in the presence of cocaine (to block noradrenaline reuptake), then after brief exposure to phenoxybenzamine (PBA, to irreversibly 'destroy' alpha 2-autoreceptors). Parallel variations of EJC/p(ulse) and delta T/p(ulse) under all conditions would support, non-parallel variations argue against exocytosis of ATP and noradrenaline from the same vesicles. The main findings were that facilitation and alpha 2-autoinhibition of EJC/p and delta T/p were remarkably similar during stimulation at 2 Hz but increasingly dissimilar at higher frequencies. delta T/p remained strongly facilitated and tightly controlled by activation of alpha 2-autoreceptors at 10-40 Hz, but both the facilitation and the sensitivity to alpha 2-autoinhibition of EJC/p were inversely related to frequency. At 40 Hz EJCs were 'small', minimally facilitated and totally unaffected by cocaine or PBA, i.e. insensitive to alpha 2 autoinhibition. Nevertheless, activation of alpha 2-receptors during the 40 Hz train strongly restricted the 'post-tetanic augmentation' (PTA) of the first EJC 10 s after the tetanus. Comparison between the frequency dependence of EJCs and the twitch contraction in the presence of cocaine or after PBA treatment indicates that it is the 'summed EJC per second', i.e. the ATP-driven current injection per unit time into smooth muscle, which triggers the twitch. The working hypothesis is proposed that these nerves use two classes of 'small vesicles' (SVs) to store and release either 'big' or 'small' ATP and noradrenaline 'quanta', and that differences in properties (Ca2+ affinity, capacity) of Ca2+ receptors in the SV membranes enable the nerves to selectively secrete 'big quanta' at low frequency and 'small quanta' during trains at high frequency. PMID- 11270139 TI - Regulation of parasympathetic neurons by mast cells and histamine in the guinea pig heart. AB - The potential interaction between the immune system and the autonomic nervous system was examined in the cardiac ganglia of guinea pigs. Intracellular voltage recordings were used to determine the effects of mast cell degranulation on the membrane properties of parasympathetic neurons in animals actively sensitized to ovalbumin. Stimulation of mast cell degranulation by perfusion with ovalbumin (10 micrograms/ml) produced a depolarization and increase in the excitability of intracardiac neurons. These effects could be mimicked by histamine application, either by perfusion (10 microM) or by local pressure application (100 microM, 1-2 s application). In either case, histamine application resulted in a similar membrane depolarization and increase in excitability. Immunohistochemical experiments demonstrated that histamine-immunoreactive mast cells are located in close proximity to parasympathetic postganglionic neurons. The histamine response was not due to release of other neurotransmitters from adjacent nerve terminals and both the depolarization and increase in excitability were inhibited by the H1 antagonist, pyrilamine (300 nM), and were unaffected by the H2 antagonist cimetidine (5 microM). Incubation of cardiac ganglion preparations from sensitized animals with pyrilamine prior to ovalbumin perfusion resulted in the inhibition of both the depolarization and increase in excitability. These results demonstrate that mast cell degranulation, and the subsequent release of histamine, results in the stimulation of intracardiac neurons via the activation of H1 receptors. Thus, local inflammatory reactions in the cardiac tissue can lead to the rapid activation of parasympathetic neurons, thereby altering cardiac function. PMID- 11270141 TI - Characterization of adenosine A1 receptor in cultured myoblast C2C12 cells of mice. AB - In an attempt to investigate the presence of adenosine A1 receptor in cell line, we used N6-cyclopentyladenosine (CPA), an agonist of adenosine A1 receptor, to incubate with C2C12 cells in vitro. CPA increased the uptake of radioactive glucose into C2C12 cells in a concentration-dependent manner and this action was abolished by the antagonists, both 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) (1,3-dipropy1-8-cyclopentylxanthine) and 8-(p-sulfophenyl)theophylline (8-SPT), at concentrations sufficient to block adenosine A1 receptor. Northern blot analysis showed the expression of adenosine A1 receptor mRNA by C2C12 cells. Western blotting also indicated a positive correlation (r = 0.99) of antibody recognized adenosine A1 receptor with membrane protein. The presence of adenosine A1 receptor in C2C12 cells can thus be considered. In the presence of U73312 (1 [6[[(17 beta)-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H- pyrrole-2,5 dione), the specific inhibitor of phospholipase C, glucose uptake stimulated by CPA into C2C12 cells was reduced concentration-dependently while it was not modified by U73343 (1-[6[[(17 beta)-3-methoxyestra-1,3,5(10)-trien-17 yl]amino]hexyl]-2,5- pyrrolidinedione), the negative control of U73312. Moreover, chelerythrine and GF 109203X (3-[1-[3-(dimethylamino)propyl]-1H-indol-3-yl]-4-(1H indol-3- yl)-1H-pyrrole-2,5-dione) also diminished the CPA-stimulated glucose uptake at concentrations sufficient to inhibit protein kinase C. The obtained data suggest that activation of adenosine A1 receptor in C2C12 cells may increase the glucose uptake via phospholipase C-protein kinase C pathway. PMID- 11270140 TI - Effects of nitric oxide in the dorsal motor nucleus of the vagus on the extrahepatic biliary system in rabbits. AB - To investigate whether nitric oxide (NO) in the dorsal motor nucleus of the vagus (DMV) mediated an influence on the extrahepatic biliary system, we studied the effects of microinjection of NO-producing drugs into DMV on the motilities of the gallbladder (GB) and the sphincter of Oddi (SO) in anesthetized rabbits. Microinjection of the NO precursor L-arginine into the rostral DMV produced an increase in the GB and SO motilities, which can be counteracted by both NG-nitro L-arginine-methyl (L-NAME), an inhibitor of NOS, and reduced hemoglobin (rHb), a scavenger of NO, and were eliminated by bilateral cervical vagotomy. On the other hand, the NO donor sodium nitroprusside (SNP) was able to mimic the excitatory effect of L-arginine. This effect can be antagonized by rHb, but not by L-NAME, for SNP supplied exogenous NO without activating NOS. These results indicate that NO in the DMV mediates an excitatory effect on the extrahepatic biliary system. PMID- 11270143 TI - [Role of physiology in solving major problems of occupational medicine]. AB - The article deals with materials concerning solution of topical problems in occupational medicine. The authors present some theoretic principles and practical solutions resulting from those materials. Being natural and experimental basis of various medical branches, clinical physiology methods are proved prospective for solution of scientific and practical problems of occupational medicine. PMID- 11270142 TI - Local opiate receptors in the sinoatrial node moderate vagal bradycardia. AB - Met-enkephalin-arg-phe (MEAP) interrupts vagal bradycardia when infused into the systemic circulation. This study was designed to locate the opiate receptors functionally responsible for this inhibition. Previous observations suggested that the receptors were most likely located in either intracardiac parasympathetic ganglia or the pre-junctional nerve terminals innervating the sinoatrial node. In this study 10 dogs were instrumented with a microdialysis probe inserted into the sinoatrial node. The functional position of the probe was tested by briefly introducing norepinephrine into the probe producing an increase in heart rate of more than 30 beats/min. Vagal stimulations were conducted at 0.5, 1.2 and 4 Hz during vehicle infusion (saline ascorbate). Cardiovascular responses during vagal stimulation were recorded on-line. MEAP was infused directly into the sinoatrial node via the microdialysis probe. The evaluation of vagal bradycardia was repeated during the nodal application of MEAP, diprenorphine (opiate antagonist), and diprenorphine co-infused with MEAP. MEAP introduced into the sinoatrial node via the microdialysis probe reduced vagal bradycardia by more than half. Simultaneous local nodal blockade of these receptors with the opiate antagonist, diprenorphine, eliminated the effect of MEAP demonstrating the participation by opiate receptors. Systemic infusions of MEAP produced a reduction in vagal bradycardia nearly identical to that observed during nodal administration. When local nodal opiate receptors were blocked with diprenorphine, the systemic effect of MEAP was eliminated. These data lead us to suggest that the opiate receptors responsible for the inhibition of vagal bradycardia are located within the sinoatrial node with few, if any, participating extra-nodal or ganglionic receptors. PMID- 11270144 TI - [Characteristics of thermoregulation of hands in patients with vibration disease]. AB - Calorimetry proved disorders of peripheral thermoregulation in vibration disease patients. The authors suggested formulae to calculate specific rate of heat flow from hand surface. Calorimetry was proved appropriate for early diagnosis of vibration disease associated with acroangiospasm fits. PMID- 11270145 TI - [Effects of infrasound and noise on contractility of lymphatic vessels (an experimental study)]. AB - Infrasound and noise inhibit contractility of lymphatic vessels. 16 Hz (100 dB) infrasound considerably decreases parameters of contractile response. Elixir "Altaisky" stimulates pump function of lymphatic vessels. PMID- 11270146 TI - [Work and health status of workers of shoe manufacturing industries]. AB - According to work conditions, severity and intensity, the main shoe-making occupations are assigned to III class of I-II jeopardy grade. If new technology applied, the work is assigned to I-II jeopardy class, being optimal--allowable. Increased mortality with liver cancer and lympholeucosis was revealed among workers contacting chloroprene. PMID- 11270147 TI - [Evaluating relative risk of morbidity or mortality for persons who participated in the clean-up of the Chernobyl accident (based on the DALY index)]. AB - The article covers a method evaluating the relative morbidity or mortality risk for those who liquidated Chernobyl accident consequences. The method is based on number of years lost due to untimely death and on number of years spent in disability due to new diseases or traumas. The highest mortality risk was revealed in the group who received at least 10.0 cGy, in workers with traumas and poisonings, in office workers with cardiovascular diseases. PMID- 11270148 TI - [Value of bone densitometry in the determination of vertebral mineral density in participants of the clean-up after Chernobyl accident]. AB - Scanning osteodensitometry helped to study mineral saturation of lumbar vertebra in individuals who liquidated Chernobyl accident consequences. Bone mineral density appeared to depend on radiation dose obtained by the examinees. High risk of osteoporosis revealed in the examinees who participated in the liquidation in 1986. The authors stress that the examinees should be covered by the skeletal state monitoring possible only with osteodensitometry. The article justify that biphotonic scanning osteodensitometers are expedient in Health Care Centers for individuals who liquidated Chernobyl accident consequences. PMID- 11270149 TI - [Psychoemotional disorders in vibration disease]. PMID- 11270150 TI - [Cytotoxicity of high-clay refractory dust]. PMID- 11270151 TI - [Vibration disease in miners of the Far North]. AB - The article covers a specified concept model of vibration disease pathogenesis. The leading mechanisms underlying the disease are nervous and hormonal dysfunction presenting hypercortisolemia and low serum levels of thyroid hormones, associated tunnel compression of upper limb nerves and immune disorders resulting in autoimmune aggression. Informative diagnostic methods are suggested and diagnostic values are justified. The authors proved efficiency of therapeutic measures aimed to relieve compression and ischemic changes in peripheral nerves, to correct immune disorders. PMID- 11270152 TI - Providing the best for our patients: evidence-based practice. PMID- 11270153 TI - Right idea, wrong approach. PMID- 11270154 TI - Expertise is what matters. PMID- 11270155 TI - T-cell lymphoma products. PMID- 11270156 TI - Assessing and treating epididymitis. PMID- 11270157 TI - Charcot's foot in diabetes. PMID- 11270158 TI - Preventing and managing exercise-induced asthma. AB - Managing exercise-induced asthma is a challenge for the patient and the clinician. As the incidence of exercise-induced asthma increases, health care providers need effective protocols to diagnose and treat these patients in outpatient settings. This article discusses the incidence, risk factors, pathophysiology, diagnosis, and treatment of exercise-induced asthma. Both traditional and new methods of disease diagnosis and treatment in the outpatient setting are explored. PMID- 11270159 TI - Post myocardial infarction treatment in the older adult. AB - Treatments for acute myocardial infarctions (AMIs) have advanced over the past few decades. Although AMIs are considered medical emergencies, continuing research has provided protocols and guidelines that significantly decrease mortality and reinfarction rates. Beta-blockers and aspirin are considered standard treatment for post-AMI patients; however, studies involving the elderly reveal that this population is less likely to receive beta-blocker and aspirin therapy. This article discusses current recommendations and treatments for post AMI elderly patients. PMID- 11270160 TI - Diagnosing malnutrition in the elderly. AB - Numerous research studies have documented the inability of many health care providers to identify nutritional deficit vulnerability and early and advanced malnutrition states in the elderly. This article provides a clinical evaluation guide for identifying risks and diagnosing incipient and advanced malnutrition. Diagnosis and intervention can prevent loss of function and independence and decrease morbidity and mortality in the elderly. PMID- 11270161 TI - Acting with awareness and care. PMID- 11270162 TI - Provider and physician: title use in HMO provider panels. PMID- 11270163 TI - Diagnosing infant botulism. PMID- 11270164 TI - Falls in the elderly: identifying and managing peripheral neuropathy. PMID- 11270165 TI - [Health for all and health management]. PMID- 11270166 TI - [Employment precariousness: an emerging public health problem?]. PMID- 11270167 TI - [Shared surveillance: meningococcal disease vs influenza]. AB - OBJECTIVE: To analyse the association between the behavior of meningococcal disease and influenza, using for this purpose population statistics for Spain for the period of 1964 to 1997. METHODS: Ecological study of the incidence of meningococcal disease and influenza in Spain from 1964 to 1997, inclusive. The study used weekly statistical data for these diseases supplied by the Compulsory Disease Reporting System (Enfermedades de Declaracion Obligatoria, EDO). The deterministic component of the meningococcal disease and influenza series was studied by means of spectral analysis based on the Fast Fourier Transformation, and the non-deterministic component was studied using the ARIMA model. The Box Jenkins method was used for pre-bleaching the series, and a cross-correlation was subsequently established between the residuals in order to detect the presence of any significant correlations between the meningococcal disease and influenza series. RESULTS: During the period from 1964 to 1997, the week that showed, on average, the greatest number of cases for the season was week 7 in the case of meningococcal disease and week 6 in the case of influenza. Spectral analysis of the meningococcal disease and influenza series clearly demonstrated the annual periodicity of both series, and periodicity of nearly 11 years for meningococcal disease and slightly over 10 years for influenza. When cross-correlation is established after prebleaching the series, positive correlations are obtained in the results of lags 0, 1, 2, and 3. Introducing influenza as an exogenous variable in the multivariate model of meningococcal disease corroborates these results. There was a statistically significant relationship between the two processes during the same week and with a three-week lapse. CONCLUSIONS: By means of a methodology not previously applied to this subject, and by the use of prolonged time-span, country-comprehensive population statistics (which includes several epidemics waves), an association was shown to exist between meningococcal disease and influenza. This suggests the need for the surveillance of the two processes in an interrelated manner. PMID- 11270168 TI - [Social and clinical characteristics of a group of mothers infected with HIV in Valencia: influence of parenteral drug addiction]. AB - OBJECTIVE: In the first years after the finding of HIV-infection, the main was for its transmission among western women was the intravenous drug addiction. The objective of our work consisted in evaluating the main social and clinical characteristics of a group of seropositive mothers, and in analyzing their potential relationship with intravenous consumption of drugs. METHOD: We performed an observational prospective study in 220 HIV-infected women who had recently given birth to children with high risk for infection. We included every women at an HIV Unit of a hospital in Valencia who had been diagnosed of HIV infection at any moment between the first mother-child transmission reported in 1985 and 1993. The analysis was based on univariate analysis. RESULTS: The virus transmission was produced by heterosexual relations in a 27.7% of the study women and due to the parenteral drug addiction in 69.1%. We detected more women who had an abortion, with criminal antecedents and parental abandonment among those who were intravenous drug users (IVDU), with OR of 1.8 (p = 0.087), 8.95 (p = 0.012) and 15 (p = 0.000), when compared with those mothers non-IVDU. Besides, IVDU presented a higher probability for getting hepatitis B or C infection and for other toxic habits, as smoking (OR = 6.19, p = 0.000) or alcoholism (OR = 5.91, p = 0.017). CONCLUSION: Many of the analysed characteristics in these women were more related with the consumption of injected drug than with the HIV infection, such as the greater frequency of elective abortions, criminal antecedents, parental abandonment, multiple drug abuse and pathological precedents like hepatitis B or C. PMID- 11270169 TI - [Variation in medical practice. Apropos of the use of CAT and NMR in INSALUD]. AB - OBJECTIVE: In this study the variability on the utilization of nuclear magnetic resonance (NMR) and computerized tomography (CT) scan among hospitals and provinces in the INSALUD (Spanish National Health System) is evaluated as well as the role of the availability of resources in the variability. METHOD: Data on availability of resources, its use and the reference population for each hospital were obtained from the Specialized Care Information System (SIAE) for the years 1996-1997. The units of analysis were the hospitals and the provinces in the INSALUD territory. The independent variables were the ratio of technologies and professional per inhabitant. Also the waiting list and the economical level of the province were used. Data analysis included the extremal quotient and multiple linear regression. RESULTS: The ratio of the highest to lowest rate of CT and NMR use is 15 and 27 among hospitals and 3 and 4 among provinces, respectively. The number of neurosurgeons, number of CT apparatus, waiting list for CT and rate of NMR use, all standardized per population, explains 61% of CT variability among hospitals. Among provinces, the number of CT apparatus explains 31% of all variability. For NMR use among hospitals, the number of neurosurgeons, number of orthopedic surgeons and CT use, all variables standardized per population, explains 42% of variability. The amount of equipment is not associated with NMR rate among provinces. CONCLUSIONS: The variation found in the INSALUD territory for the two procedures is high and ecologically associated to the availability of resources. It would be convenient to perform an observational study to confirm the findings and evaluate the possible contribution of inappropriate use to the variation. PMID- 11270170 TI - [Cost-effectiveness analysis of self-monitoring of blood glucose in type 2 diabetics]. AB - OBJECTIVE: Compare the cost-effectiveness of self-monitoring of blood glucose (MBG) with your non-use. DESIGN: Descriptive and retrospective study covering the period 1995-97 in the 597 type-2 diabetes patients: 286 practicing MBG on a stable basis and 311 not doing so. All are registered in seven health districts in the territorial ambit of Tortosa Primary Care. Were quantified the direct costs in relation to consumption of reagent strips for the practice of MBG, outpatients visits in your primary care center, derivations to specialist of reference and complementary test according to recommendations of the European NIDDM Policy Group in the population user of MBG and no-user; the annual cost increment, the average annual cost and the total annual cost in the population user of MBG and in the application of a ideal model of quantitative and qualitative cover according to clinical recommendations of the Gedaps; and the cost-effectiveness. RESULTS: While the 78% of the total diabetic population satisfy some clinical indication for prescribing MBG, only the 42.5% practice the MBG. The consumption of reagent strips rising of 8% to 15% of the global cost of the diabetic population. In the application of the ideal model of cover, this cost increase up the 30% of global cost. The effectiveness obtained, an 27%, not are significantly different in the population user of MBG and no user. The cost effectiveness in the user of MBG increased of 210.789 ptes/year to 213.148 ptes/year; and no-user of 162.019 ptes/year to 162.051 ptes/year. The application of ideal model of cover and the gain of an effectiveness near to possible level of efficiency imply an descent average of cost-effectiveness of approximately 60%: 78.904 ptes/year in user MBG and 54.682 ptes/year in no-user. CONCLUSIONS: 1. We choose in the presents conditions the option of no-user MBG. 2. The average cost-effectiveness per diabetic patient will increase by the needs of accommodate the therapy to new standards of metabolic control. 3. Are clear opportunity for the improve the management and to motivate an efficient use of technology associate to defects of public sanitary market. 4. The model of ideal cover associated to greater effectiveness are necessary for to unify the economic and clinic efficiency. PMID- 11270171 TI - [Establishing maximum permissible values in soil for health protection with the Lur model]. AB - OBJECTIVE: Environmental pollution can be the origin of health problems and it is seen by populations as a hazard. Therefore, when environmental maximum levels of contaminants are set it is necessary to guarantee public health protection and to promote a healthy environment. In the Basque Autonomous Community (Spain) the establishment of maximum levels of contaminants in soil has followed the model named LUR, based on the risk assessment methodology. These values are levels of pollutant that are supposed not to represent an unacceptable risk for human beings. METHODS: Maximum levels of contaminants in soil arise from establishing a maximum tolerable risk and assessing toxicity information and human exposure in relation to land uses. The model has considered 5 land uses for exposure: children playground, residential with garden, residential, public park and industrial/commercial. The routes of exposure taken into account have been soil ingestion, consumption of home-grown vegetables, soil particle and vapour inhalation and dermal absorption. When inhalation and dermal absorption contribute largely to contaminant intake, standards are less robust because uncertainty in exposure assessment in these cases is high. RESULTS AND CONCLUSIONS: The results of applying this methodology to the case of two contaminants are presented: cadmium, as a non carcinogenic inorganic contaminant, and benzo(a)pyrene, as a carcinogenic organic contaminant. Limitations of the methodology are discussed. PMID- 11270172 TI - [Comparison of 2 methods for calculating uncertainty in laboratory analysis]. AB - OBJECTIVE: To compare two methods in the estimation of the uncertainty in laboratory quality control. METHODS: A computerized simulation is performed to compare the delta method (suggested by the International Organization for Standardization and the Entidad Nacional de Acreditacion) and a bootstrap-based method. The simulation includes several situations with different environmental conditions and different relationships between the analyzed variables. RESULTS: The mean in the coverage obtained by the estimated confidence intervals is higher and closer to the nominal using the bootstrap than using the delta method. The most important differences are observed in the coverage percent distribution: while using the bootstrap, a great number of simulations obtain coverage near the nominal of 95%; using the delta method the coverage are more dispersed, including coverage of 100% in some occasions and lesser than 80% in others. The bootstrap offers very similar results under different conditions, including in the presence of unknown and unmeasured variables or when the analyzed variables are correlated. The delta method shows poorer results in both situations. CONCLUSION: The uncertainty in the laboratory quality control can be estimated more accurately with bootstrapping than with the delta method. PMID- 11270173 TI - [Media impact of the SESPAS (Spanish Public Health Association) 2000 report]]. AB - OBJECTIVE: To measure the mediatic effect of different communication strategies used in public health advocacy. More specifically, to compare the effectiveness of the World Wide Web as a tool to attract the attention of journalists, with other more traditional formulas. METHODS: For the Spanish Public Health Association (SESPAS) Report 2000, two types of media strategies to communicate the report contents were programmed: a) traditional and passive strategies, centred in approaching journalists through press releases and press conferences around the SESPAS meeting (November 15-20 1999); b) interactive strategies, since August 15 to December 30, focused towards attracting health journalists to the non-embargoed, full text SESPAS report launched in a web site. To facilitate the web page use, we wrote a letter, in the first week of August, to all the members of the Spanish Health Communicators Association giving them the URL and the website map. In parallel, a monitoring system of the media impact was established from August to December 99, covering 250 magazines and 70 newspapers, in order to locate and recover all the stories about the SESPAS report for further analysis. RESULTS: Sixty-six stories were recovered; they were published in 32 press media from 24 provinces with an advertising value of 18,243,873 Ptas. As a whole, smaller circulation rate papers published more stories than larger ones. During five months, the SESPAS report was present in the press agenda, even though stories were not distributed homogeneously over time. Information concentrated around three moments: the first one, a week after our summer mailing; the second one, in the occasion of the publication of a story about the increase in traffic accidents in El Pais, and the third one during the SESPAS meeting. There were significant differences among those stories published from the traditional strategies of communication and those published from the interactive ones, the latter being more diverse, with more contributions of the journalists and tackling a wider range of issues. CONCLUSIONS: The combination of traditional and alternative communication strategies was a effective option. Unlike previous experiences in this occasion, with the network aid, the presence of SESPAS in media was not punctual around the Congress, but maintained during five months. The results and the obtained experience of this research can be useful for future public health advocacy interventions in Spain. PMID- 11270174 TI - [Declaring conflict of interest in scientific publications. Time for the spotlights and stenographers in the backroom of research financed by the industry?]. AB - The term conflict of interests is applied to those situations in which the research validity and integrity may be influenced by a secondary interest, typically an economic benefit, but also an ideological, personal or professional interest. In this work we describe some ways of conflict of interests- particularly those related with the publication of clinical and epidemiological research supported by the industry--and the regulation of this problem from medical journals, including references to the situation in Spain. The conflict of interest is not synonymous of scientific fraud neither malpraxis in research, but in the medical literature there exists enough evidence to consider it as an important source of biases. The usual form of facing the conflict of interests is to make it public, so that readers can judge its importance. The editorial policies of the Spanish journals are, in general, far from giving importance to this problem, an aspect which could favor an attitude of the investigators, to maintain funding or to obtain new contracts, unnecessarily subordinated to the interests of the companies. PMID- 11270175 TI - [Debate on frequentative vs Bayesian methods]. PMID- 11270176 TI - [Primary care networks in Catalonia. Structure and process analysis]. PMID- 11270177 TI - [The role of communications media in public health]. PMID- 11270178 TI - Newborns with ambiguous genitalia, impalpable gonads, and hyperpigmentation. PMID- 11270179 TI - Sialic acid in human milk: composition and functions. AB - Breast milk is the best nutrient source for infants. It contains all elements needed for a normal growth and development of infants. Human milk contains a large amount of sialic acid compared with bovine milk. Sialic acid contained in oligosaccharides, glycolipids and glycoproteins in milk is considered to play important roles in physiological functions in infancy. Thus, we have investigated the sialic acid composition and the functions of sialylated compounds in human milk. Sialic acids comprise a family of neuraminic acid derivatives present in secretions, fluids and tissues of mammals. In milk, sialic acid is present in different sialoglycoconjugate compounds such as oligosaccharides, glycolipids and glycoproteins, not in a free form. Human milk contains 0.3-1.5 mg/ml of sialic acid. Sialic acid bound to oligosaccharides accounts for about 75% of the total sialic acid contained in human milk. Most of the sialic acid contained in human milk is found in the form of sialyllactose, an oligosaccharide formed from lactose and sialic acid. In milk, gangliosides, sialic acid-containing glycolipid, occur mainly as monosialoganglioside 3 (GM3) and disialoganglioside 3 (GD3). The concentration of GM3 in human milk increases, while that of GD3 concentration decreases during lactation. Because the brain and central nervous system contain considerable level of sialic acid in infancy, it is considered to play important roles on the expression and development of their functions. Moreover, we found that some sialylated compounds had inhibited the adhesion of toxins, bacteria and viruses to the receptors on the surface of epithelial cells. Additionally, we found that some sialylated compounds had growth-promoting effects on bifidobacteria and lactobacilli, predominantly present in the intestinal flora of infants fed with human milk. The results suggested that sialylated compounds in human milk possibly behaved as a physiological component in the intestinal tract of infants to protect them against enteric infections. PMID- 11270180 TI - Pediatric asthma: promoting best practice (AAAAI & AAP) 1999. PMID- 11270181 TI - The predisposing factors of coagulase negative staphylococcal bacteraemia in neonatal intensive care unit. AB - Coagulase negative staphylococci are the commonest blood culture isolate from infants on neonatal intensive care units. The differentiation of contaminants from isolates representing true infection remains a significant clinical problem. Data from two neonatal intensive care units were collected prospectively in order to find those parameters, which best correlated with actual sepsis. Each case was assessed using clinical parameters to categorise infants into infection and contaminant groups. Logistic regression was then performed to find significant correlates. Three correlates were found, namely the presence of a long line (P = 0.001), abnormal white cell count (P = 0.037) and abnormal white cell morphology (P = 0.027). Abnormal white cell morphology was assessed by two experienced hematologists. More than half the isolates were probable contaminants and true infection may occur in the absence of a long line in this patient group. PMID- 11270182 TI - Neurologic involvement in an outbreak of enterovirus 71 infection: a hospital based study. AB - Enterovirus (EV) can cause varied clinical manifestations. Involvement of the central nervous system (CNS) with the nonpolio EVs are common and important causes of morbidity in children. To investigate the manifestations of nonpolio enteroviral infections with CNS involvement during the EV outbreak, from February 1998 to January 1999, we collected 153 hospitalized patients in our pediatric ward caused by nonpolio EV infections which were diagnosed by history, clinical features, or detected from viral cultures. Fourteen patients (9.2%) had CNS presentations, 13 males and one female. The ages ranged from one month to 10.3 years. The spectrum of CNS presentations included aseptic meningitis (4 cases, 28.6%), encephalitis (5 case, 35.7%), encephalomyelitis (3 cases, 21.4%), and poliomyelitis-like syndrome (2 cases, 14.3%). Among these patients, 8 cases (57.1%) were isolated with EV71 from at least one site of rectal or throat swab sampling. Two fatal cases were presented as encephalitis and complicated with pulmonary edema. Generally, enteroviral infections are considered as a benign infectious disease in children. However, pediatricians should keep in mind that EV71 has caused several endemic outbreaks and continues to be an occasional cause of severe CNS disease. Early evaluation and appropriate treatment of CNS enteroviral infections may minimize the neurologic sequelae. PMID- 11270183 TI - Final height of children with type 1 diabetes: the effects of age at diagnosis, metabolic control, and parental height. AB - Normal growth is one of the major goals in the treatment of children with type 1 diabetes. We prospectively monitored the linear growth and metabolic control of 44 children (13 boys) with type 1 diabetes from the time of diagnosis to the attainment of adult height and analyzed the relationship between the height and the age at diagnosis, metabolic control, and genetic target height. At diagnosis, girls at puberty were taller (height in standard deviation score: 0.60 +/- 0.94, p = 0.022), while boys (-0.03 +/- 0.67) and prepubertal girls (0.24 +/- 0.86) were similar to the age-controlled children. During the following years, they lost height compared to their height at diagnosis (p = 0.009), but they still attained an average final height (-0.13 +/- 0.66 in boys, -0.05 +/- 0.86 in girls) correlated with their height at diagnosis (r = 0.37, p = 0.014), as well as their genetic target height (r = 0.78, p < 0.005). The final height as well as the reduction in height was not linearly correlated with the age at diagnosis. The mean HbA1c level of the 44 children was 10.33 +/- 1.74%, boys had better control compared with girls (mean HbA1c 9.45 +/- 1.28 v.s. 10.71 +/- 1.78%, p = 0.013). The final height or the reduction in height was not linearly correlated with the mean HbA1c level. PMID- 11270184 TI - Thanatophoric dysplasia type I. AB - Thanatophoric dysplasia is a sporadic, nearly always lethal congenital skeletal dysplasia. It is characterized by shortening of the limbs, a severely small thorax, macrocephaly, and platyspondyly. There are two major subtypes: a short, curved femur characterizes type I, and a straighter femur with cloverleaf skull characterizes type II. Recently, mutations in the fibroblast growth factor receptor 3 (FGFR-3) gene have been identified in both subtypes, which suggest that thanatophoric dysplasia is a genetically homogenous skeletal disorder. Most affected neonates die of respiratory failure, due to narrow thorax with pulmonary hypoplasia. Antenatal sonographic diagnosis is feasible in the second trimester of pregnancy, but differentiating thanatophoric dysplasia from non-lethal skeletal disorders is very important. At the present time, however, prenatal genetic screening seems unpractical. PMID- 11270186 TI - Chylus-like urine as a complication of percutaneous hyperalimentation catheter in an infant: report of one case. AB - A very low-birth-weight neonate developed chylus-like urine after receiving parenteral nutrition (PN) via percutaneous central venous catheters (CVC) for 7 weeks. A perirenal fluid collection could be seen under sonography. This kind of complication has not been described in literature. After withdrawing the CVC for 5 cm, the urine cleared up. For patients under prolonged PN via CVC and repeated change dressing of the CVC, close monitoring and regular evaluation of the position of the catheter tip are warranted. PMID- 11270185 TI - Congenital complete heart block. AB - Congenital complete heart blocks (CCHB) are mostly related to the neonatal lupus syndrome. The purpose of this paper was to assess the clinical spectrum of CCHB in our hospital. Nine patients were retrospectively enrolled between 1994 and 1999. The birth history, electrocardiography, 24-hour Holter monitoring, pacemaker insertion and its complications, maternal disease, and maternal and infant autoantibody levels were studied. All nine cases were diagnosed prenatally. Hydrops fetalis was noted in five (55.6%). Six cases were live births and the other three were terminated. No anatomical heart defects were noted. Initial electrocardiography revealed the atrial rates ranged from 150 to 166 beats per minute. The minimal ventricular rates ranged from 46 to 80 beats per minute. VVI mode pacemakers were inserted through xyphoid approach in all live birth infants. Complications were noted in three of them (50%). Antinuclear antibody and anti-SSA/Ro antibody were positive in all 8 mothers (100%). The anti SSB/La antibody was positive in 6 of the eight mothers (75%). Five infants tested positive for anti-SSA/Ro antibody. None of the infants tested positive for anti SSB/La antibody. In conclusion, all CCHBs in our series were associated with maternal autoantibodies. Because of high complication rate of permanent pacemaker insertion during the neonatal period, it should be restricted in certain conditions. PMID- 11270187 TI - Food protein-induced enterocolitis syndrome: report of one case. AB - We report a 76-day old infant who got diarrhea within the first week of life. He was treated as acute gastroenterocolitis and kept on feeding with regular infant formula. Because the symptoms persisted, the feeding formula was shifted to soy based formula then to the highly-hydrolyzed formula and got improvement. But severe bloody diarrhea, vomiting, dehydration and fever developed after feeding with regular infant formula again. Based on the history and clinical presentations, cow's milk allergy was suspected. He received total parenteral nutrition for 5 days then fed with highly-hydrolyzed formula with slowly increasing amount. Thereafter tests for total eosinophil counts, total serum IgE, milk specific IgE antibodies and milk extract skin prick test were all unremarkable. Under the impression of food protein-induced enterocolitis syndrome (FPIES), a double-blind placebo-controlled food challenge (DBPCFC) with infant formula was performed. Regular infant formula induced severe vomiting, diarrhea, fever, acidosis and elevation of absolute neutrophil counts (ANC) of peripheral blood by 27,640/mm3. Based on the laboratory findings and challenge results, the patient fit the diagnostic criteria of food protein-induced enterocolitis syndrome. PMID- 11270188 TI - Neurologic complications of enterovirus 71 infection in children: lessons from this Taiwan epidemic. PMID- 11270189 TI - Zellweger syndrome: report of one case. AB - Zellweger syndrome is a fatal autosomal-recessive hereditary disease characterized by the absence of peroxisomes in liver and kidneys. The absence of peroxisomes results in impairment of many metabolic pathways, especially beta oxidation of very long chain fatty acids (VLCFAs). We report a case of a three month-old male infant with facial dysmorphism, hypotonia, psychomotor retardation, and hepatomegaly. He had an elder brother with the same facial features and hypotonia who died of hepatic failure at four months of age. Biochemical studies revealed elevation of blood pipecolic acid and VLCFAs, compatible with peroxisomal disorder. Electron microscopy of liver biopsy revealed absence of peroxisomes. Zellweger syndrome was diagnosed. Because this syndrome is usually fatal in early life, genetic counseling and prenatal diagnosis are crucial. PMID- 11270190 TI - Kawasaki disease: a new disease? AB - Because the etiology of Kawasaki disease (KD) is still unknown, KD cannot yet be called a disease entity but is still regarded as a clinical entity. The 1999 paper by Sarah S. Long and the 1999 letter to the editor by Stanford T. Shulman were both examined. We must await the discovery of the etiology before we can make a definite statement whether or not KD is a new disease. PMID- 11270191 TI - [Electrocochleography and phase audiometry in diagnosis of Meniere disease]. AB - The diagnosis of Meniere's disease is uncertain when the typical symptoms do not occur completely and definitely. A reliable finding of an endolymphatic hydrops (EH) is the base for a correct prognosis and therapy. Electrocochleography is a proven diagnostic procedure but requires a lot of time and of technical know-how. The mobility of cochlear partition can be tested by low-frequency masking (LFM) recording the phase dependent subjective masked threshold of a short test tone. We performed electrocochleography and LFM in 29 patients with suspected Meniere's disease at the same day. Both tests pointed at an EH in 62%, and in 24% the results showed correspondent negative results. In 14% the results were inconsistent. Both methods also showed in 59% of the contralateral ears without symptomatic signs the indication of an EH. Considering the good conformity of both tests the easier LFM can be recommended for detection of EH. PMID- 11270192 TI - [Oncocytic neoplasms of the parotid gland. Differential diagnosis, clinical course and review of the literature]. AB - BACKGROUND AND OBJECTIVE: Oncocytic neoplasms of the salivary glands are rare. PATIENTS/METHODS: We report on seven cases of oncocytic neoplasms of the parotid gland (one multifocal nodular oncocytic hyperplasia, five oncocytomas, and one oncocytic carcinoma). RESULTS: While the history, clinical presentation, and histology of all oncocytic neoplasms showed no characteristic differences, intraoperatively the well-differentiated oncocytic carcinoma displayed an infiltrative, locally aggressive growth pattern. A local carcinoma recurring 7 years later corresponded to the primary tumor. There were no metastases. Immunohistochemistry revealed that all oncocytic neoplasms were positive for markers of cytokeratins (KI-1) and negative for markers of carcinoembryonal antigen (CEA), S-100 protein, and smooth muscle actin (SMA). In contrast to the benign neoplasms, the oncocytic carcinoma showed an increased rate of proliferating cells (MIB-1) and a strongly positive reaction with the antibody MA 903 against high molecular weight cytokeratins. CONCLUSIONS: In a review of the literature, we could identify a group of locally aggressive, low-grade oncocytic carcinomas with a considerably better prognosis, similar to that of our case. The therapeutic significance of these findings is discussed. PMID- 11270193 TI - [Early detection of pediatric hearing loss. Visual and automated procedures compared]. AB - The recording of otoacoustic emissions is suitable for the detection of hearing loss in small children. The test meets the following requirements for hearing screening: it is carried out in a few minutes, specialized personnel is not necessary, the results do not depend on the vigilance of the child, and total costs are comparably low. However, the choice of a suitable device is quite essential. A test of the ILO88, ILO92, Echosensor (Otodynamics Ltd.), and Echoscreen (Madsen) devices was performed with 102 children (aged < 1 year). Additionally, a software package (Otoclass) for offline analysis of ILO88 data files was tested. The results indicate that all devices applying OAE as a screening measure was able to detect every ear (n = 25) with a hearing loss indicated by the outcome of a control BERA (stimulus level 35 dB nHL), thus reaching a sensitivity equal to 100%. The specificity of the different OAE devices depends on the underlying detection strategy. The best results were achieved with the automated Echoscreen device (95.9%) followed by the Otoclass analysis software (94.2%). The Echosensor device failed in our study to provide good specificity (77.3%). Reflex audiometry, which is favored by pediatricians in Germany, when used alone is completely inadequate as a screening method, even if conducted by a well-trained investigator. In our study, only 61.5% of the children with hearing loss were detected with reflex audiometry, and 42.7% of the children with normal hearing were misclassified. These results deviated from the results presented in ref. 13 to a large extent, as the Hanover group attested sensitivity and specificity of 100% for reflex audiometry (HNO 43, Reuter et al.). The deviating results are discussed in detail. PMID- 11270194 TI - [Metastasis in the frontal skull base from hepatocellular carcinoma]. AB - Metastatic tumours involving the nose and the paranasal sinuses are rare. Especially metastatic spread to the sphenoid sinus is an extremely rare occurrence. The most common metastatic tumour is the renal cell carcinoma. Only four cases of hepatocellular carcinoma presenting a sphenoid sinus metastasis could be found in a search of the literature. We report on the case of a 59-year old male who suffered from a sphenoid sinus mass. A biopsy showed the tumour to be a metastatic hepatocellular carcinoma. The suspected primary tumour was then found in the left liver lobe. The early diagnosis of paranasal sinus malignancies is difficult because of the varied and nonspecific symptoms and signs. In cases of late diagnosis, the treatment is usually palliative with a poor prognosis. The importance of endoscopic examination and CT or MRI scan for early detection must be emphasized. PMID- 11270195 TI - [Middle ear adenoma. Long-term course of a rare neoplasm]. AB - Adenomas of the middle ear are rare benign glandular neoplasms arising from the middle ear mucosa. After previous operations 25 and 15 years before, a 67-year old female complained about dizziness, tinnitus, and unilateral hearing loss on the left side. A tumor in the tympanum that was revealed by otoscopy could be removed completely. Histological examinations showed an adenoma of the middle ear with cholesteatoma. This was inconsistent with the histological result of the operation in 1983, which had described a hidradenoma. An exact analysis of the preparations confirmed that a middle ear adenoma had already been present in 1983. Hidradenoma is one of the most important differential diagnoses. The characteristic histological sign of middle ear adenomas in contrast to hidradenomas is the lack of myoepithelial cells. In addition, it is very difficult to differentiate middle ear adenoma and adenocarcinoma using histopathological and clinical methods. Therefore, thorough follow-up is mandatory for patients after surgical treatment of middle ear adenomas. PMID- 11270196 TI - [Cystic neck tumor. Extracranial schwannoma of the hypoglossal nerve]. PMID- 11270197 TI - [Malpractice issue in bilateral excision of exostoses]. PMID- 11270198 TI - [Reconstruction of large tongue defects]. PMID- 11270199 TI - [Statistics on allergology and environmental medicine in the ENT specialty in Germany]. AB - Environmental medicine and allergy were found to be important in the daily ENT practice after statistical analysis and an inquiry among all ENT doctors in Germany. 64% of ENT colleagues do practice allergy, 20% environmental medicine. With reference to all registered ENT doctors 24% are titled allergists, 4% specialists in environmental medicine. Focussing on the teaching there is a big difference between both groups. Whereas training facilities for allergists are reasonable, there are only a few chief instructors for environmental medicine in the ENT society. This fact has to be considered on educational plannings in future. PMID- 11270202 TI - Codman Award winners describe improvements. PMID- 11270200 TI - [Rupture of the round window--detection with fluorescence endoscopy]. AB - Rupture of the round window membrane as a special cause of inner ear deafness is widely accepted after changing pressure levels, e.g. in diving. However, even without a barotrauma before, the spontaneous rupture of the round window membrane is suspected occasionally in patients with sudden hearing loss and/or vertigo and tinnitus. To carry through the tympanotomy is decided by ENT surgeons often in cases of progressive hearing loss despite infusion therapy. Perilymph fistulas have been detected relatively seldom, compared to the number of reported operations by several authors. However, covering the round niche with connective tissue leads to the improvement of symptoms sometimes even in cases without microscopical evidence of fistula. Within the last 3 years 14 patients suffering sudden hearing loss of one ear underwent tympanotomy in our department. Of these patients 8 reached restitution of the hearing ability. Especially 2 patients with sudden deafness caused by spontaneous rupture of the round window membrane are reported in the following article. Perilymph fistulas were detected in these cases by IV-application of fluorescein and fluorescence endoscopy of the middle ear. Both patients obtained a normal hearing curve within 1 week after surgical intervention and obliteration of the round niche. PMID- 11270201 TI - [Rheologic infusion therapy, neurotransmitter administration and lidocaine injection in tinnitus. A staged therapeutic concept]. AB - BACKGROUND AND OBJECTIVE: Cochlear dysfunction and tinnitus are treated by means of hemorrheological infusions in order to increase the cochlear oxygen supply and restore function of hair cells, neurotransmission and central processing of auditory information. PATIENTS/METHODS: In a retrospective analysis of the charts of 123 patients treated between February 1993 and May 1994, we analyzed effectiveness and safety of a gradual therapeutic regimen, consisting of dextrane/procaine infusions, lidocaine i.v. injection and infusion therapy with the neurotransmitter glutamic acid. RESULTS: Tinnitus decreased in 83.7% of patients with acute tinnitus (AT) and 16.1% of patients with chronic tinnitus (CT) during dextrane/procaine infusion. The majority (89%) experienced their tinnitus relief during the first 5 days. Treatment with glutamic diethylester and glutamic acid resulted in a 26.5% overall improvement. Application of lidocaine intravenously over a period of 10 min diminished tinnitus loudness or frequency in 16.7% (AT) and 38.9% (CT) of cases respectively. The long-term effects of therapy were investigated by a follow-up mailing action: 66.7% of the AT and 15.6% of the CT sufferers stated a clear therapy effect over time. Nonserious side effects were noted in 4% of the treated patients. Therefore safety was excellent. CONCLUSIONS: For acute and chronic tinnitus a gradual therapeutic regimen is recommended: (1) infusions with dextrane and procaine over 5 days; (2) intravenous application of 100 mg lidocaine over 10 min; and if necessary (3) administration of glutamic diethylester and glutamic acid for 3 days. This resulted in overall tinnitus relief in 95.3% of the acute and 26.7% of the chronic tinnitus sufferers. PMID- 11270203 TI - Analyzing data in measures with small populations. PMID- 11270204 TI - Updating PPO accreditation. PMID- 11270206 TI - Simplifying compliance activities. PMID- 11270205 TI - Setting a cap on scoring for new patient safety requirements. PMID- 11270207 TI - Questions on the environment of care. PMID- 11270208 TI - Trending to reduce medication errors. PMID- 11270209 TI - Self-reported training needs and training issues of advisors to self-advocacy groups for people with mental retardation. AB - My purpose in this paper is to describe the self-reported training needs and training information of advisors to self-advocacy groups for people with mental retardation. A telephone survey was administered to 118 advisors randomly selected, resulting in 90 completed surveys and a response rate of 76%. Major findings are reported in training topics, preferred training formats, hours and frequency of training, how training needs change, and differences in training needs based on advisor demographic data. The results can be used by individuals and groups to help better support the self-advocacy movement by assisting self advocacy groups to better prepare and train individuals to be successful and useful advisors. PMID- 11270210 TI - Community services, issues, and service gaps for individuals with developmental disabilities who exhibit inappropriate sexual behaviors. AB - Inappropriate sexual behaviors represent the most challenging behaviors for community service providers. A national survey of 243 community agencies was conducted to describe services provided for individuals with developmental disabilities who exhibit high-risk sexual behaviors and to identify issues and service gaps. The most common types of offenses were sexual behavior (a) in public situations, (b) that inappropriately involved others, and (c) involved minors. Community agencies used multifaceted approaches to serve these individuals. The major issues and problems were systemic, specifically staff issues and service gaps, followed by funding. Implications of this study are that increased knowledge and skills related to sexuality and inappropriate sexual behavior and mental health resources are needed to build community capacity to serve this population. PMID- 11270211 TI - Teaching mathematics to students with mild-to-moderate mental retardation: a review of the literature. AB - A systematic search of the literature from 1989 through 1998 was conducted to identify and analyze mathematics interventions for students with mild-to-moderate mental retardation. We found that the focus of instruction has shifted from basic skills instruction to computation and problem-solving instruction. Techniques such as constant-time delay, peer tutoring, time trials, and direct instruction proved beneficial in improving mathematics skills. Further, students with mental retardation learned to employ cognitive strategies successfully when these techniques were included. Although this information is promising, we recommend that further studies be conducted in secondary schools and in inclusive settings. PMID- 11270213 TI - Naming, defining, and classifying in mental retardation. AB - Recent discussions about changing the term mental retardation to a different term may be considered in the broader framework of three distinct but related processes: naming (terminology), defining, and classifying. The three processes are analyzed according to their purposes and functions: In naming, a term is assigned; in defining, the term is explained; and in classifying, the group is divided into subgroups according to stated principles. The current status of each process is described, especially as represented in the 1992 AAMR manual, Mental Retardation: Definition, Classification, and Systems of Supports. We suggest three sets of guiding questions that may help evaluate proposed changes in naming, defining, or classifying. PMID- 11270212 TI - National estimates of vocational service utilization and job placement rates for adults with mental retardation. AB - Data from the 1994 and 1995 Disability Supplements of the National Health Interview Survey (NHIS) were used to estimate rates of utilization of vocational services and examine employment outcomes for adults with disabilities who have received vocational services. Those living outside the formal long-term care system, and who were self or proxy identified as having mental retardation, were compared with other adults with disabilities. Analyses suggest that compared to other working-age persons with disabilities, adults with mental retardation (a) have a different population profile, (b) receive different types of services, (c) experience similar levels of satisfaction, (d) have much lower rates of competitive employment, and (e) are much more likely to be employed in segregated work settings. Research and policy implications of findings are discussed. PMID- 11270214 TI - Children with mental retardation/developmental disabilities: do physicians ever consider needed dental care? PMID- 11270215 TI - Reflections on participatory action research with people who have developmental disabilities. PMID- 11270216 TI - Adapting the person-centered approach in Singapore: a situated perspective. PMID- 11270217 TI - Selectivity and specificity in analytical chemistry. General considerations and attempt of a definition and quantification. AB - Selectivity and specificity are performance characteristics of analytical methods which are frequently used in analytical literature. In general, the terms are applied verbally and a quantification of selectivity and specificity is given rarely. Excepted are methods like chromatography and ISE sensoring which use individual quantities such as selectivity coefficients, indices and other parameters to characterize analytical procedures and systems. Here a proposal is given to characterize selectivity and specificity quantitatively by relative values in a range of 0 to 1 expressing so a certain degree of selectivity and specificity. By examples it will be shown that the derived quantities characterize analytical methods and problems in a plausible way. PMID- 11270218 TI - On the effect of catalyst status in the quantitative determination of platinum in Pt-Sn/MgO materials. AB - The development of an analytical method for the determination of platinum in MgO based Pt/Sn catalysts is described. Electrothermal atomic absorption spectroscopy (ETAAS) was chosen because of its high sensitivity and robustness against spectral interferences. All the sources of chemical interferences were statistically analyzed and corrections were adopted for the presence of magnesium oxide support. The effectiveness of different mineralization procedures was critically evaluated as a function of the chemical of the solid catalyst. Microwave digestion allowed recovery of metal of 100% for all the catalysts analyzed and exhibited significant better precision values than other digestion methods, which could nevertheless be utilized under proper conditions in selected cases. PMID- 11270219 TI - Analysis of sintered products of iron ore fines by flame atomic absorption spectrometry using a matrix modifier. AB - A very precise and accurate method for chemical analysis of sintered products (from iron ore fines in "pot grate furnace") is discussed. A matrix modifier/buffer (a mixture of KCl, tartaric acid, HCl and H2SO4) is used to prevent interference of iron in the determination of calcium and magnesium by flame atomic absorption spectrometry. Also, an EDTA titration method is recommended for calcium and magnesium after separating iron in the form of mixed oxides by ammonia-ammonium chloride buffer. Statistical data for a series of experiments are presented and precision values are found to be comparable with those of conventional methods used for complex metallurgical products. For the majority of cases, the agreement between the two methods is extremely good. A slight deviation has been noted in a few samples which may be overcome by a more thorough sampling prior to analysis. PMID- 11270221 TI - Continuous preconcentration system for nitrate ions using anionic reverse osmosis tubes coupled to an ion chromatograph. AB - A continuous preconcentration system for nitrate ions was developed using cation exchange tubing made from Nafion perfluorosulfonic acid membrane. This method is based on ion exclusion effects and reverse osmosis phenomena. The system was evaluated by connecting it to an ion chromatograph. The concentration ratios could be increased by raising the pressures between the two sides of the cation exchange tubing. Twenty-fold concentration of nitrate ion was achieved when the pump pressure was 20 x 10(5) Pa. The relative standard deviations of the preconcentration ratio at four different pump pressures, 5, 10, 15 and 20 x 10(5) Pa were 1.2-2.8% (n = 5). PMID- 11270220 TI - A comparison of phosphated and sulfated beta-cyclodextrins as chiral selectors for capillary electrophoresis. AB - The enantioseparation capabilities of three different functionalized beta cyclodextrins, two sulfated beta-cyclodextrins with 4 and 15 nominal degrees of substitution and a phosphated beta-cyclodextrin with 8 degrees of substitution, were compared. While anodic detection was used with both sulfated cyclodextrins, the phosphated cyclodextrin required cathodic detection suggesting either lower ionization of the phosphated cyclodextrin or generally lower affinity of the analytes for the phosphated cyclodextrin. The effects of several experimental parameters were evaluated with respect to enantioseparation. The degrees of substitution of the cyclodextrin, pH of the background electrolyte as well as the concentration of the functionalized beta-cyclodextrin, each had a significant influence on the successful enantiomeric separation of the chiral drugs investigated. PMID- 11270222 TI - Synthesis of glycosylated porphyrins as neutral ionophores for a berberine sensitive electrode. AB - For preparing a berberine-sensitive electrode, 5,10,15,20-tetrakis[2-(2,3,4,6 tetraacetyl-beta-D-glucopyranosyl)-1-O- phenyl]porphyrin (T(o-glu)PPH2) was synthesized from the reaction of pyrrole with ortho-acetylglycosylated benzaldehyde by Lindsay's method. The electrode based on T(o-glu)PPH2 with an optimized membrane composition exhibits Nernstian response to berberine in the concentration range 2.4 x 10(-7)-5.0 x 10(-3) mol L-1, with a pH range from 3.9 to 10.2, and a fast response time of 30 s. The electrode shows fair selectivity towards berberine with respect to common co-existing species. T(o-glu)PPH2 shows better potentiometric response characteristics comparing to chloro[5,10,15,20 tetrakis[2-(2,3,4,6-tetraacetyl-beta-D- glucopyranosyl)-1-O-phenyl]-porphinato] manganese (MnT(o-glu)PPCl) and better selectivity towards berberine than tetraphenylporphyrin (TPPH2). The effect of the composition of the electrode membrane has been studied and the experimental conditions optimized. The contents of berberine in pharmaceutical tablets were determined by direct potentiometry and the results agreed with values obtained by the pharmacopoeia method. PMID- 11270223 TI - Characterization of single-stranded DNA on chitosan-modified electrode and its application to the sequence-specific DNA detection. AB - Single-strand DNA could bind with chitosan on a platinum electrode via forming a tight DNA-chitosan complex. The salt concentration of the ssDNA solution had an obvious effect on the surface coverage, the immobilization was remarkably reduced at high salt concentration. The sample ssDNA immobilized on the chitosan-modified electrode can hybridize efficiently with the complementary sequences and be successfully used for the sequence-specific DNA detection. The same results could be obtained using a gold or graphite electrode modified with chitosan. The stability of this electrode has been also discussed. PMID- 11270224 TI - Voltammetric behavior of L-cysteine in the presence of CPB at a silver electrode. AB - The voltammetric behavior of L-cysteine at a silver electrode is described. L Cysteine can be anodically accumulated at a silver electrode surface as a sparingly soluble silver salt; at more negative potentials, the insoluble compound is stripped cathodically yielding a small current peak at about -1.10 V (vs. SCE). In the presence of cetyl pyridine bromide (CPB), the stripping peak shifts slightly to a more negative potential, and the peak height increases significantly. Thus, the peak becomes more useful for the determination of L cysteine. In contrast to other surfactants, CPB can improve the accumulation and stripping of L-cysteine obviously. The voltammetric behavior of cysteamine, 3 mercaptopropionic acid and homocysteine is discussed as well. PMID- 11270225 TI - Continuous determination of chloroquine in plasma by laser-induced photochemical reaction and fluorescence. AB - A flow injection fluorimetric method is proposed for the determination of chloroquine based on the photochemical derivatisation in an alkaline medium of the analyte and fluorescence generation after irradiation with a pulsed Nd:YAG laser operated at 355 nm. Chemical, hydrodynamic and laser variables were studied in order to obtain the best conditions for quantification. A linear range from 25 to 600 micrograms/L was achieved, with a correlation coefficient of 0.997 (n = 8), an RSD of 4.3% (n = 11) and a detection limit of 8 micrograms/L (3 sigma). The sample throughput was 10 h-1. The method was successfully applied to the determination of chloroquine in human plasma. The increase of sensitivity with respect to the method based on monitoring the intrinsic fluorescence of chloroquine itself was 1.7 times. PMID- 11270226 TI - Validated method for the determination of the novel organo-ruthenium anticancer drug NAMI-A in human biological fluids by Zeeman atomic absorption spectrometry. AB - NAMI-A is a novel ruthenium-containing experimental anticancer agent. We have developed and validated a rapid and sensitive analytical method to determine NAMI A in human plasma, plasma ultrafiltrate and urine using atomic absorption spectrometry with Zeeman correction. The sample pretreatment procedure is straightforward, involving only dilution with an appropriate hydrochloric acid buffer-solution. Because the response signal of the spectrometer depended on the composition of the sample matrix, in particular on the amount of human plasma in the sample, all unknown samples were diluted to match the matrix composition in which the standard line was prepared (plasma-buffer 1:10 v/v). This procedure enabled the measurement of samples of different biological matrices in a single run. The validated range of determination was 1.1-220 microM NAMI-A for plasma and urine, and 0.22-44 microM for plasma ultrafiltrate. The lower limit of detection was 0.85 microM in plasma and urine and 0.17 microM in plasma ultrafiltrate. The lower limit of quantitation was 1.1 and 0.22 microM, respectively. The performance of the method, in terms of precision and accuracy was according to the generally accepted criteria for validation of analytical methodologies. The applicability of the method was demonstrated in a patient who was treated in a pharmacokinetic phase I trial with intravenous NAMI-A. PMID- 11270227 TI - Determination of tartaric acid in wines by FIA with tubular tartrate-selective electrodes. AB - A flow injection analysis (FIA) system comprising a tartrate-(TAT) selective electrode has been developed for determination of tartaric acid in wines. Several electrodes constructed for this purpose had a PVC membrane with a complex of quaternary ammonium and TAT as anion exchanger, a phenol derivative as additive, and a more or less polar mediator solvent. Characterization of the electrodes showed behavior was best for membranes with o-nitrophenyl octyl ether as solvent. On injection of 500 microL into a phosphate buffer carrier (pH = 3.1; ionic strength 10(-2) mol/L) flowing at 3 mL/min, the slope was 58.06 +/- 0.6 with a lower limit of linear range of 5.0 x 10(-4) mol/L TAT and R2 = 0.9989. The interference of several species, e.g. chloride, bromide, iodide, nitrate, gallic acid, tannin, sucrose, glucose, fructose, acetate, and citrate, was evaluated in terms of potentiometric selectivity coefficients. The Hofmeister series was followed for inorganic species and the most interfering organic ion was citrate. When red and white wines were analyzed and the results compared with those from an independent method they were found to be accurate, with relative standard deviations below 5.0%. PMID- 11270228 TI - Ultrasound-assisted solubilization of trace and minor metals from plant tissue using ethylenediaminetetraacetic acid in alkaline medium. AB - A simplified and fast sample pretreatment method based on ultrasound-assisted solubilization of metals from plant tissue with ethylenediaminetetraacetic acid in alkaline medium is described. Powdered unknown and certified plant samples (particle size < 50 microns) were slurried in the solubilization medium and subjected to high intensity ultrasonication by a probe ultrasonic processor (20 kHz, 100 W). Metal solubilization can be accomplished within 3 min using a 30% vibrational amplitude and 0.1 M EDTA at pH 10, the supernatant obtained upon centrifugation being used for analysis. The method is applied to several food plants with unknown metal contents and certified plant samples such as CRM GBW07605 tea leaves, BCR CRM 61 aquatic moss and BCR CRM 482 lichen, with good trueness and precision. Intensive treatments with concentrated acids involving total matrix decomposition can be avoided. Metal determination (Ca, Cd, Mg, Mn, Pb and Zn) in the alkaline extracts was carried out by flame and electrothermal atomic absorption spectrometry. PMID- 11270229 TI - Quantification of carbohydrate structures in size fractionated aquatic humic substances by two-dimensional nuclear magnetic resonance. AB - Two-dimensional phase sensitive C,H correlation spectra were successfully applied to the quantification of carbohydrate substructures in aquatic humic substance (HS) fractions obtained by tangential flow multistage ultrafiltration (TFMSTUF) of a selected bog water HS (HO13, German Research Program DFG-ROSIG) as well as a river HS (Suwannee River Fulvic Acid Reference of the International Humic Substances Society, IHSS). It turns out that after size fractionation the HS samples give very well resolved C,H-correlation spectra which offer a great potential for substructure quantification. Details of the combined substructure quantification technique, novel in HS characterization, are presented. The results of the combined procedure point out that carbohydrate moieties predominantly occur in higher molecular mass fractions (> 10 kDa) of isolated HS. PMID- 11270230 TI - Slurry sampling fluorination assisted electrothermal vaporization-inductively coupled plasma-atomic emission spectrometry for the direct determination of metal impurities in aluminium oxide ceramic powders. AB - A new analytical procedure for the direct determination of metal impurities (Cr, Cu, Fe and V) in aluminium oxide ceramic powders by slurry sampling fluorination assisted electrothermal vaporization-inductively coupled plasma-atomic emission spectrometry (ETV-ICP-AES) is reported. A polytetrafluoroethylene (PTFE) emulsion was used as a fluorinating reagent to promote the vaporization of impurity elements in aluminium oxide ceramic powders from the graphite tube. A vaporization stage with a long ramp time and a short hold time provided the possibility of temporal analyte-matrix separation. The experimental results indicated that a 10 microL 1% m/v slurry of aluminium oxide could be destroyed and vaporized completely with 600 micrograms PTFE under the selected conditions. Two aluminium oxide ceramic powder samples were used without any additional pretreatment. Analytical results obtained by using standard addition method with aqueous standard solution were checked by comparison of the results with pneumatic nebulization (PN)-ICP-AES based on the wet-chemical decomposition and analyte-matrix separation. The limits of detection (LODs) between 0.30 microgram g-1 (Fe) and 0.08 microgram g-1 (Cu) were achieved, and, the repeatability of measurements was mainly better than 10%. PMID- 11270231 TI - Simultaneous determination of Fe(II) and Fe(III) in waters by capillary isotachophoresis. AB - An ITP method for the simultaneous determination of Fe(II) and Fe(III) in waters, based on separation of their EDTA and fluoride complexes, respectively, was developed. The leading electrolyte used consists of chlorides, La(III) as co counter ion and is buffered with beta-alanine to pH = 3.5. The terminating electrolyte contains caproic acid and L-histidine (pH = 4.5). The method was validated and tested with samples of artificial, ground and treated water with good results, comparable to those obtained by other analytical techniques. Fe(II) and Fe(III) up to 20 mg/L were measured with an RSD = 1.4-1.5% and detection and determination limits of 0.8-0.9 and 3.0-3.5 mg/L, respectively. The ITP method can be recommended for routine utilization in hydroanalytical laboratories. PMID- 11270232 TI - [Diagnosis of status epilepticus by continuous EEG monitoring in a neurointensive care unit]. AB - Continuous EEG monitoring use has documented a surprisingly high incidence of convulsive and non-convulsive status epilepticus in patients with acute brain injuries. Seizures are the cause of secondary insult. Many problems may be encountered during EEG recording, such as electrical interferences and artefacts arising from the patient. To minimise these problems, we suggest the following: train the bedside nurse, make a library of the artefacts, have the EEG technologist check electrodes and establish low impedance, have the electroencephalographer examine the EEG, correlate the activity and movements of the patient. In the diagnosis and the management of convulsive and non-convulsive status epilepticus, the value of continuous EEG monitoring appears to be established. PMID- 11270233 TI - [Antiepileptic treatment in cerebral lesions: what are the indications and the main practical use?]. AB - The aim of this report is to review the indication and the practical use of the antiepileptic drugs in patients with cerebral lesions. The use of antiepileptic drugs to treat seizure or status epilepticus in an emergency is well catalogued and reported in this paper. Practical use of antiepileptic drugs, after a first seizure or to prevent a seizure, in patients with a cerebral lesion, is controversial. The question of antiepileptic drugs in seizures and in prophylaxis is discussed in different types of cerebral lesions: head injury, stroke, cerebral arteriovenous malformation and cerebral tumour. PMID- 11270234 TI - [Neurofunctional tests in presurgical strategies for partial epilepsies]. AB - The presurgical evaluation of drug-resistant partial epilepsies primarily relies on two major investigations, including a long term video-EEG monitoring which aimed at recording the patient's typical seizures, and a specifically designed high quality magnetic resonance imaging (MRI). The latter demonstrates an abnormality within the epileptogenic lobe in the majority of cases, which might not, however, necessarily match the epileptogenic zone. Numerous functional neuro imaging techniques have been progressively added to the pre-surgical evaluation of refractory partial epilepsies, such as the study of cerebral glucose metabolism, benzodiazepine receptor availability, and methionine incorporation using positron emission tomography (PET), the evaluation of ictal cerebral blood flow changes using single photon emission computerized tomography (SPECT), the measurement of N-acetyl-aspartate concentration with magnetic resonance spectroscopy, and the mapping of eloquent areas using functional MRI. These investigations can help to confirm the origin of seizure onset previously suggested by MRI and electro-clinical data, and provide independent prognostic information regarding the chance of a successful surgical treatment. Moreover, functional neuro-imaging data can have a critical diagnostic value when MRI is strictly normal or shows multifocal abnormalities. However, the variety and rapid evolution of functional neuro-imaging techniques makes it difficult to propose a standard protocol. Finally, it remains mandatory to proceed to an intracranial EEG investigation in a substantial number of patients, including the majority of those suffering from an extra-temporal epilepsy. PMID- 11270235 TI - [Functional neurosurgery for epilepsy]. AB - Introduced at the end of the last century, epilepsy surgery is indicated in patients with intractable partial seizures and based on the resection of the epileptogenic cerebral tissue from which ictal discharges originate. Palliative procedures include seizure spread pathways interruption (callosotomy, multiple subpial transections) and chronic stimulation of the vagus nerve. Complete preoperative investigations including seizure observation, clinical tests, video EEG, MRI and functional MRI, and PET-scan are performed in order to identify the epileptogenic zone. In difficult cases, invasive seizure monitoring through depth electrode implantation (SEEG) is performed. Resections for temporal lobe seizures are associated with favorable outcome: 60 to 90% of patients will be seizure-free after surgery. A less favorable outcome is observed after extra-temporal resections: 40 to 60% seizure-free patients. A better outcome is observed after surgery for epilepsy associated with an image-defined lesion, most often a tumor, rather than for cryptogenic epilepsy. Tumors associated with chronic partial epilepsy are indolent, most of them are dysembryoplastic neuroepithelial tumors (DNET). Outcome after palliative procedures are more variable, depending on the etiology of epilepsy. PMID- 11270236 TI - [Anesthesia for epilepsy surgery]. AB - Epilepsy is rather common, affecting 0.5 to 2% of the population. Numerous patients, particularly those resistant to the antiepileptic therapy, can be surgically treated after a thorough evaluation. Surgery for epilepsy can be carried out either under general or local anaesthesia with sedation. This second approach is reserved for the extirpation of foci localised in motor, sensory or language areas. During the preoperative anaesthetic evaluation, two specific points have to be taken into account: the psychological aspect and the antiepileptic medication. During the procedure, an electrocorticography with or without stimulation may be indicated, particularly when a perioperative stimulation is scheduled. Low doses of volatile agents are chosen, and no curare and large doses of benzodiazepines and barbiturates. Awakening takes place on the operation table for a rapid and reliable neurological evaluation. During procedures performed under local anaesthesia, the anaesthetist must be ready at any time to intubate the patient in order to secure the airway. PMID- 11270237 TI - [Status epilepticus: pharmacokinetic basis of anticonvulsant treatment in adults]. AB - In status epilepticus, the optimal efficacy of the antiepileptic drugs depends notably on effective, quickly reached and sufficiently lasting cerebral concentrations and the optimal tolerability notably on the lack of excessive storage in the brain and other tissues. So, the best efficacy-tolerability ratio of these drugs is largely determined by their pharmacokinetic properties. A linear kinetics, a not too short distribution half-life, a neither too brief nor too long elimination half-life, a fast and easy crossing of the blood-brain barrier and the lack of long-lasting accumulation in fat tissues are among the main ideal pharmacokinetic properties. Any of the antiepileptic drugs currently used in status epilepticus has all these properties together. An accurate knowledge of the pharmacokinetics is absolutely crucial to rationally decide the route of administration, the loading dose and the maintenance doses. However, pharmacokinetics must only complete, but cannot replace, the clinical experience and judgement, especially because some limitations: kinetic equations are mathematically exact but theoretical; individual kinetics in a given patient is exceptionally known in clinical practice; finally the pharmacokinetics may be significantly modified during a status epilepticus, especially of the generalized convulsive type, due to systemic consequences and complications of the seizures. In the emergency situation of status epilepticus, the correlation between the clinical efficacy and the so-called "therapeutic" plasma levels remains ill defined. The reported values are often very high and their range appears very large. Nevertheless plasma levels are useful, especially for the monitoring of the evolution; they are mandatory for nonlinear-kinetics drugs. PMID- 11270238 TI - [Epileptogenic drugs in anesthesia]. AB - Most anaesthetics and analgesics have both pro- and anticonvulsant activity. The data in the literature should be analysed with respect to the patient population, the recording of epileptic activity and the method of EEG analysis. Among inhaled anaesthetics, isoflurane has strong anticonvulsant properties. In some circumstances, sevoflurane may induce an epileptic activity. With the exception of ketamine and etomidate, all intravenous hypnotics may be used for anesthesia of the epileptic patient. Midazolam is a potent anticonvulsant. Among narcotics, fentanyl and alfentanil may induce clinical or electroencephalographic seizures. Considering the large number of patients treated with these agents without any neurological adverse effect, the clinical relevance of these data is unclear. Neuromuscular blocking agents do not possess pro- or anticonvulsant properties. PMID- 11270239 TI - [Practice in electroconvulsive therapy]. AB - The aim of this paper is to describe the practice of electroconvulsive therapy in CHU Dijon, according to the Anaes guidelines. Seven component procedures are listed, particularly from the standpoint of the psychiatrist. PMID- 11270240 TI - [Anesthesia for electroconvulsive therapy]. PMID- 11270241 TI - [Plasma osmolarity and cerebral volume]. AB - Under normal physiological conditions, the osmolarity of extracellular fluids (ECFs) and natremia are controlled by two regulatory mechanisms modulating the water balance and sodium outflow from information collected by the osmoreceptors and baroreceptors, respectively. As well, under normal physiological conditions, water and electrolytes of brain ECFs are secreted by the endothelial cells of brain capillaries. Furthermore, isotonicity is present on both sides of the blood brain barrier. In the event of systemic osmolarity disorders, water transport subject to osmosis laws occurs at the level of the blood-brain barrier. In the case of plasmatic hyperosmolarity cerebral dehydration is observed, while cerebral edema occurs in the contrary case. However, plasmatic osmolarity disorders have less effect on the cerebral volume when their introduction is slow. Experimentation in acute conditions shows that measured variations of the cerebral water content are lower than calculated variations, thus suggesting the existence of an adaptive mechanism, that is, the cerebral osmoregulation which limits the variation of the volume of brain cells by modulating their osmoactive molecule content. These osmoactive molecules are, on the one hand, the electrolytes, which are early and rapidly mobilized, and, on the other hand, the organic osmoles (amino acids, etc.), whose secretion is slower and delayed. This phenomenon should be taken into account in the treatment of osmolarity disorders. Thus, the related-risk of treatment for natremia disorders is therapeutic reversal of the osmotic gradient at the level of the blood-brain barrier. This reversal, which corresponds to a second osmotic stress, requires the implementation of a new procedure of cerebral osmoregulation in the opposite direction of the preceding one. As successive osmotic stresses decrease the effectiveness of brain osmoregulation, the risk for cerebral dehydration and pontine myelinolysis increases when the treatment of chronic hyponatremia is too aggressive. Finally, the choices for infusion solutions in neurosurgery and treatment for osmolarity disorders are also based on the relationships that exist between plasmatic osmolarity and the brain. PMID- 11270242 TI - [Hyponatremia in neurologic intensive care: cerebral salt wasting syndrome and inappropriate antidiuretic hormone secretion]. AB - Hyponatraemia is a frequent complication in neurologically injured patients; it is a secondary cerebral injury. Hyponatraemia leads to consciousness problems, convulsions, worsening of the neurological status and thus the neurological evaluation. Hyponatraemia is secondary to free water retention (inappropriate ADH secretion) or to renal salt loss. The cerebral salt wasting syndrome (CSWS) has been described with head injury, subarachnoid haemorrhage and after several sorts of brain insults. It is characterised by an increased natriuresis and diuresis. Diagnosis is based on hyponatraemia, hypernatriuresis, increased diuresis and hypovolaemia. However, inappropriate ADH secretion and CSWS share several diagnostic criteria. The atrial natriuretic factor and the C-type natriuretic factors play a role in the development of the CSWS. The diagnostic approach and monitoring are based on the assessment of sodium and water losses. Therapy is based on correction of the circulating volume and natraemia. Speed of correction is a matter of debate: slow correction presents the risk of further neurological injury whereas rapid correction presents the risk of central pontine myelinosis. PMID- 11270244 TI - [Status epilepticus--sodium and osmolarity]. PMID- 11270243 TI - [Hypernatremia in neurosurgical pathology]. AB - Hypernatraemia is defined as an increase in extracellular sodium concentration, associated with plasma hyperosmolality and cellular dehydration. It can result from excessive water loss, from an increase in the total sodium content or from both mechanisms. As far as neurosurgical pathology is concerned, hypernatraemia due to excessive water loss may be observed in patients who do not sense thirst or are unable to ingest water. Urinary water loss is seen in diabetes insipidus and osmotic diuresis. Extrarenal water losses from pulmonary origin may be observed in intubated or tracheotomized patients. Hypernatraemia with sodium and water retention may be encountered in patients suffering from Cushing diseases or syndromes, or more frequently in those who are given excessive amounts of sodium (hypertonic saline, sodium salts). Clinical manifestations of hypernatraemia consist of neurologic symptoms related to cellular dehydration; their severity is correlated with the rapidity of the onset of the electrolytic disorder. Depending on the pathophysiological mechanism, treatment of hypernatraemia involves stopping sodium intake, restoring normovolaemia and administering hypotonic fluids. Treatment of diabetes insipidus relies on the administration of the antidiuretic hormone and of drugs that increase its secretion rate or its responsiveness in the kidneys. PMID- 11270245 TI - [Pathophysiology of epileptic seizures and status epilepticus]. AB - Primary and secondary epileptogenesis involves multiple genetic and acquired factors. Epileptogenesis is a complex result of combined factors including membrane factors, neurotransmitter and environmental factors. Ion channel-related diseases, GABA and glutamate dysfunction, and glial reaction intervene in different epileptic conditions. The understanding of the mechanisms which emphasize initiation and maintenance of status epilepticus (SE) are in progress. Prognosis of SE is related to the duration of epileptic activity and to the acute cerebral and systemic consequences. Delayed cellular and molecular alterations after SE are responsible for secondary epileptogenesis. Glutamate receptor activation is the main key point leading to an excessive intraneuronal accumulation of ionic calcium by which a cascade of reactions is induced. Apoptotic neuronal death, glial reaction axonal sprouting and neurogenesis contribute to a state of hyperexcitability and hypersynchrony. A better understanding of underlying mechanisms of epileptogenesis may serve the development of new drugs with both anticonvulsant and antiepileptic (prevention or neuroprotection) actions. PMID- 11270247 TI - [Predicting the outcome of very premature infants: a medical and public health challenge]. PMID- 11270246 TI - [Traffic accidents in adolescents: a growing health risk]. PMID- 11270248 TI - [Outcome of 249 premature infants, less than 29 weeks gestational age]. AB - The aim of this paper was to report the vital and neurological outcome of 249 preterm infants of less than 29 weeks born between 1990 and 1996, and included in a prospective study until two years of age. RESULTS: The initial mortality rate was 19%. This was related to gestational age and severe transfontanellar ultrasonographically (TFU) detected abnormalities. The rate of follow-up at two years of age was 98%. Neurological sequelae amounted to 12.8%, including four cases of deafness. The possibility of survival without neurological sequelae increased from 52% at 24-25 weeks to 72% at 26-28 weeks of gestational age (p < 0.005). The presence of sequelae was significantly related to severe cranial ultrasonographically-detected abnormalities, to parental social level, and to early neonatal anemia. Normal TFU and/or isolated periventricular hyperechogenicity could not exclude the presence of neurological sequelae which, however, appeared to be less severe than at the onset. CONCLUSION: Gestational age, severe TFU abnormalities and neonatal anemia play a major role in the rate of mortality and in the neurological sequelae in preterm infants, and can influence the decisions concerning the treatment of this pediatric population. PMID- 11270249 TI - [Family socioeconomic deprivation and vulnerability in the pediatric emergency room: evaluation and management]. AB - OBJECTIVE: The aim of this study was to evaluate the importance of socioeconomic deprivation and other forms of vulnerability in families attending the pediatric emergency unit (PEU). It was based on a five-level classification of family types and an analysis of responses provided by the French healthcare system. METHOD: The prospective study involved 150 families admitted to the PEU on the basis of open-response interviews that analyzed demographic and socio-economic characteristics, motivations for consultation, the child's quality of life, family problems experienced by the parents, and their support network. RESULTS: Five categories of families were defined as follows: complete destitution necessitating immediate social measures (0.7%); acknowledged and well-managed economic deprivation (13.3%), unacknowledged and/or complex economic deprivation with a significant deterioration in the quality of life (22.2%), familial psychological vulnerability without economic deprivation (30.4%), and families without any apparent problems (33.7%). Consultation at the PEU appears to be a multifactorial phenomenon motivated by socioeconomic, psychological and cultural factors. This phenomenon is connected with the present-day imbalance in the healthcare system, which does not adequately respond to the needs of vulnerable families. CONCLUSION: This survey contributes to the current reflection on the integration of psychosocial factors in child healthcare at both the hospital and local community level. The method described herein has determined the factors of vulnerability and the risks of economic deprivation. It can contribute to the development of improved communication and cooperation between practitioners, the hospital and local social workers. PMID- 11270250 TI - [Anastomotic stenosis after surgical treatment of esophageal atresia: frequency, risk factors and effectiveness of esophageal dilatations]. AB - Anastomotic stricture is the most common complication following the surgical repair of esophageal atresia, and is usually treated by esophageal dilation. OBJECTIVES: The aims of this study were to assess in an infant population operated on at birth for type III or IV esophageal atresia: 1) the frequency of esophageal stenosis following the repair of esophageal atresia, and associated factors; 2) the efficacy of esophageal dilation by the Savary-Gaillard bougie technique. MATERIALS AND METHODS: The medical records of 52 children presenting with esophageal atresia over a 5-year period were retrospectively reviewed. Gestional age and birth weight, duration of mediastinal and transanastomotic drainage, and anastomotic complications including leakage, stricture, and the presence of gastroesophageal reflux were recorded and analysed. Patients presenting with anastomotic stricture were compared with a group of children without stricture. The number of esophageal dilations, their efficacy and the complication rate were analyzed. RESULTS: Anastomotic stricture developed in 20 (40%) of the 50 patients undergoing primary repair for esophageal atresia. The occurrence of anastomotic stricture was related to anastomotic tension during esophageal surgical repair (p < 0.03). Young children required esophageal dilation at a mean age of 142 days (24-930 days). Stricture resolution occurred after a mean of 3.2 dilations (1-15) over an average period of 7.9 months (range: 0-30 months). Dilation was successful in 90% of the 20 patients. Seven patients required only one dilation. Perforation of the esophagus occurred in one case, and this severe complication led to the death of the child. Esophageal dilation was unsuccessful in two patients, who presented prolonged severe dysphagia. CONCLUSION: Anastomotic stricture following repair of esophageal atresia is connected with the length of the gap that has to be repaired, and tension during suture. Esophageal dilation by the Savary-Gaillard bougie technique is an effective method for treating esophageal stricture. Several dilations are usually needed before the disappearance of dysphagia. PMID- 11270251 TI - [Evaluation of 47,213 infants in neonatal screening for cystic fibrosis, using pancreatitis-associated protein and immunoreactive trypsinogen assays]. AB - OBJECTIVES: The increasing evidence of the benefits of neonatal screening for cystic fibrosis (CF) indicates that this procedure could soon be implemented throughout France. The screening strategy currently used involves the detection of infants with elevated levels of immunoreactive trypsinogen (IRT) (approximately 1% of the population), followed by the detection of CFTR gene mutations. However, genetic analysis has certain drawbacks, the most important of which being the management of heterozygotes, and in France the requirement by law of previous informed consent. In cases of CF, pancreatic alterations are already present in utero. A previous study has demonstrated the value of pancreatitis associated protein (PAP) as a screening test for CF, and has indicated that a feasible two-stage strategy could involve the following: 1) selection of infants with elevated PAP levels; 2) in this group of infants, subsequent detection of those with elevated IRT levels for direct CF diagnosis by the sweat test thereby avoiding the use of genetic analysis. The study aim was to evaluate this strategy in a large number of neonates. METHODS AND RESULTS: The aforementioned strategy was evaluated in a prospective study involving 47,213 infants in the Provence region of France. In infants with a PAP > 7.5 ng/mL (1.28%), 176 had an elevated IRT level > 700 ng/mL (0.37%). In this limited population sample (0.37% of the total), the sweat test diagnosed five cases of CF. A sixth case involving the monozygous twin of an infant with diagnosed CF remained undetected, probably because of a registration error. Genetic analysis confirmed the diagnosis, and also detected another case in an infant with two CFTR mutations but with a normal phenotype at 20 months of age. As the observed incidence was similar to that which had previously been reported, and as no further case was subsequently detected two years after the end of the study, this indicated that the sensitivity of this screening strategy was satisfactory. Its specificity makes the direct diagnosis of CF cases by the sweat test feasible, without further selection by genetic analysis. CONCLUSION: The PAP/IRT technique for CF detection seems to be suitable for mass screening, without the drawbacks of genetic testing. PMID- 11270252 TI - [Acute myelitis of an unusual cause in a child: the lymphocytic choriomeningitis virus]. AB - Acute transverse myelitis is a rare disorder in childhood. It usually occurs as a post-infectious disease, but a precise infectious agent is identified in only 20% of cases. OBSERVATION: The diagnosis of acute transverse myelitis was made in a 5.5-year-old girl who initially presented with left Claude-Bernard-Horner syndrome and meningitis. A few days later, motor and sensory tetraparesia with bladder dysfunction was observed. Magnetic resonance imaging showed a diffuse lesion in the medulla, with a hypersignal in the T2 and a hyposignal in the T1 sequences. Serum analysis showed the presence of a viral infection due to the lymphocytic choriomeningitis (LCM) virus. The outcome was marked by complete recovery of the sensorimotor deficit, but a persistence of the left Claude Bernard-Horner syndrome. CONCLUSION: In rare cases, the LCM virus is responsible for myelitis. In the present case, the Claude-Bernard-Horner syndrome was secondary to the cervico-medullary lesion. Recent reports in the literature have been discussed, in particular as regards the use of immunomodulatory therapy, which clearly improves patient prognosis. PMID- 11270253 TI - [Type B botulism: a family outbreak]. AB - CASE REPORTS: Three cases of an outbreak of familial foodborne botulism are reported. The food incriminated could not be identified despite a careful investigation into the food history of the patients. The outcome was good following endotracheal ventilation and botulism antitoxin trivalent therapy. CONCLUSION: In France, foodborne botulism is an uncommon public health disease, and with a good prognosis when the diagnosis is promptly performed. The value of emergency electromyographic findings is emphasized, particularly when the repetitive stimulation of the motor nerve shows a presynaptic block of neuromuscular transmission. Management depends on the symptomatology, and trivalent antitoxin therapy is the only specific treatment. PMID- 11270254 TI - [Eccrine neutrophilic hidradenitis: idiopathic plantar form in children]. AB - In this study, two cases have been reported of idiopathic plantar hidradenitis, an uncommon dermatological pathology with a spontaneous favorable outcome. OBSERVATIONS: Two children aged 12 and 14 years presented with a painful papulo nodular plantar rash with major functional impairment. The diagnosis of idiopathic plantar hidradenitis was considered, and then confirmed in one case by plantar biopsy. Non-steroidal antiinflammatory drugs, associated with paracetamol in one case were administered. The symptoms disappeared spontaneously within a few days in both cases, without any recurrence. CONCLUSION: A knowledge of the symptoms connected with plantar hidradenitis in the child allows a rapid diagnosis to be made without hospitalization or further medical examination. Analgesic treatment and rest seem to be the only useful approaches. Biopsy to investigate eccrine gland infiltration by neutrophils can only be proposed in the case of an abnormally prolonged duration or an atypical presentation of this pathology. PMID- 11270256 TI - [Specifics in pediatric arterial cerebral infarctions]. AB - Despite the number of potential causes, most children with ischemic cerebrovascular disease are classified into a few major categories: 1) cardioembolic stroke; 2) moyamoya syndrome; 3) complication(s) connected with systemic disease, with a known risk of a cerebrovascular event; 4) cervicocephalic arterial dissection; and 5) transient cerebral arteriopathy of undefined origin (probable arteritis). The etiological diagnosis is rapidly established by complementary investigations based on the initial clinical and imaging findings: cardiac exploration, magnetic resonance imaging and angiography or echo-doppler of the cervical arteries if cervical dissection is suspected; intra-arterial catheter cerebral angiography and analysis of the cerebrospinal fluid to investigate an intracranial arteriopathy. The outcome and treatment depend on the type of stroke, on its accurate identification, and on prediction regarding the risk of recurrence. Although there is a constitutional predisposition to cerebrovascular accidents, environmental triggers such as trauma, infectious disease and cardiac surgery also play a major role. PMID- 11270257 TI - [Genetic deafness:the primary cause of sensorineural hearing loss in children]. AB - Genetically-transferred hearing impairments account for more than 50% of cases of pediatric sensorineural hearing defects. Multiple clinical aspects are involved in genetic hearing impairment, including the involvement of other organs, genetic inheritance, and the degree and age at onset of hearing loss. Diagnosis relies on family history, on the systematic investigation of the symptomatology including an associated syndrome, and audiometry testing in parents and siblings. Analysis of the connexin 26 gene is also indicated, as it is frequently involved in this disorder. Further genetic analysis in affected families will aid in detecting other as yet unidentified genes responsible for hearing impairment. PMID- 11270255 TI - [Intrapleural fibrinolytic treatment and infectious pleuresies: three pediatric cases]. AB - Intrapleural instillation of fibrinolytic agents has been proposed for the treatment of loculated pleural effusions, or whenever the biochemical characteristics of the pleural fluid (pH, glucose level, LDH) indicate the risk of a complicated outcome due to a pleural effusion with complications and the possible development of empyema. At present, there is no consensus regarding the use of intrapleural fibrinolytic agents in children. CASE REPORTS: In this study, the successful treatment by fibrinolytic agents and standard drainage are successfully performed in three children with a pleural effusion due to an infection. CONCLUSION: The clinical utility, in terms of the reduction of the duration of hospitalization and additional surgical treatment, should be assessed in prospective studies. PMID- 11270258 TI - [Radiological case of the month. Crohn's disease]. PMID- 11270259 TI - [Secondary effects of vaccinations]. AB - The adverse effects of vaccines include local reactions and systemic symptoms or illnesses. Local reactions are frequent, most often presenting as transient pain, redness, edema and/or nodule. Fever of short duration is the main systemic symptom, generally occurring within 24-48 hours following vaccination. Some vaccines have recognized specific adverse effects such as thrombocytopenic purpura for the measles-mumps-rubella vaccine, and febrile convulsions for the pertussis vaccine. Hepatitis B vaccine and Haemophilus influenzae type b vaccine have been respectively suspected to be responsible for neurological demyelinating disease and insulin-dependent diabetes mellitus, but large-scale epidemiological studies have failed to confirm these allegations. PMID- 11270260 TI - [Sciatic paralysis after a buttock intramuscular injection in children: an ongoing risk factor]. AB - Intramuscular injections are regularly recommended for the administration of certain drugs in children. This article underlines the fact that buttock intramuscular injection risks injury to the sciatic nerve, which may lead to lower limb palsy, most often presenting as paralytic drop foot. This condition rarely results from direct traumatic lesion of the sciatic nerve, but usually from the caustic effect of the injected drug. It may occur in older children and adolescents, as well as in infants and younger children. Therefore, the buttocks should not be used as an intramuscular injection site in children whatever their age. In the case of sciatic nerve injury following intramuscular injection, extrafascicular neurolysis may prevent the occurrence of paralysis. PMID- 11270261 TI - [Management of children with fever without localizing signs of an infection]. AB - The management of infants and young children with fever without source (FWS) is a difficult challenge for pediatricians. Of 100 children with FWS, ten to 20 will have a serious bacterial infection (SBI) and 4 to 5% bacteriemia. Because no single aspect of the medical history, physical examination and laboratory parameters can reliably identify a child at increased risk for SBI, most management strategies now focus on identifying infants that are less likely to have SBI. The negative predictive value for 'low-risk criteria' SBI is close to 100%. Therefore, if it is possible to carry out a daily clinical examination, antibiotic treatment can be withheld from these children. For children who do not fulfill the low-risk criteria, antibiotics must be administered until the results of blood and urine samples and possibly CSF cultures have been obtained. PMID- 11270262 TI - [Handicapped epidemiology in children]. PMID- 11270263 TI - [64th Congress of the American College of Rheumatology, Philadelphia, October 28 November 2, 2000]. PMID- 11270264 TI - [Specific inhibitors of Cox-2 and hemostasis: point of view of internal medicine]. PMID- 11270265 TI - [Chronic urticaria and Hashimoto-Hashimoto's thyroiditis: report of 6 cases]. AB - PURPOSE: Chronic urticaria is a common skin disorder. The cause is rarely determined. Autoimmune diseases, particularly autoimmune thyroiditis, have been implicated in the occurrence of chronic urticaria. METHODS: We reviewed clinical records of patients with Hashimoto's disease and chronic urticaria. RESULTS: In our department, six patients had presented chronic urticaria associated with Hashimoto's thyroiditis: four patients, of which three treated with L-thyroxine were euthyroid, the other two were hypothyroid. Hashimoto's thyroiditis had been diagnosed for three patients during the investigation of chronic urticaria. Three patients developed chronic urticaria though they were treated with thyroid suppression for Hashimoto's disease. Two of them had a dramatic improvement with opotherapy. One patient who was euthyroid without treatment improved with hormonal therapy. The fourth patient had a partial remission with thyroid hormones and was cured with corticotherapy. CONCLUSION: The mechanism by which thyroid autoimmunity is associated with urticaria is poorly understood. A cross linking of IgE receptors of mastocytes induced by antithyroid antibodies may be a cause of histamine release. Hormonal therapy may be a potent event for the clinical improvement by the suppression of chronic thyroid stimulation. Assay of thyroid hormone and antithyroid antibodies should be performed in patients with chronic urticaria. Discovery of Hashimoto's thyroiditis with chronic urticaria requires thyroid hormone replacement not only in hypothyroid but also euthyroid patients. PMID- 11270266 TI - [Nutritional status assessment in 57 hospitalized aged patients: impact of the causal disease]. AB - PURPOSE: To study the nutritional status in elderly patients hospitalized for rehabilitation and to compare it among patients with hip fracture and those with medical care. METHODS: Patients were nutritionally assessed upon admission (d0) to our unit by measurement of anthropometric, biological parameters and dietary intake. Thirty-seven patients were operated for hip fracture (group I) and 21 were hospitalised for medical disease (group II). Nutritional status was compared in the two groups on d0 and was evaluated after one month (d30) in the operated group. RESULTS: No significant difference could be observed for any anthropometric or biologic (albumin, transthyretin and transferrin) in the two groups. Daily food intake related to body weight was much the same in both groups (31 kcal/kg). C-reactive protein and orosomucoid levels were above the reference range in both groups. Hip fracture operated patients had higher orosomucoid than non-operated ones (1.50 +/- 0.4 versus 1.14 +/- 0.4 g/L; P = 0.002). One-month follow-up of nutritional status performed in 31 operated patients showed a significant decrease in TST and MAC (respectively p = 0.02 and p = 0.007) and in orosomucoid (p = 0.003) although daily food intake increased. CONCLUSION: Twenty eight percent of patients were undernourished upon admission in the unit. A moderate inflammatory state still remained in all our patients, particularly in those who had undergone surgery. This inflammatory state persisted two months after surgery. PMID- 11270267 TI - [From the first symptoms to terminal renal failure: need for a nephrologic follow up]. AB - PURPOSE: As patients with chronic renal failure are frequently referred late to nephrologists, we decided to quantify the magnitude of late referral and its consequences. METHODS: We studied retrospectively an inception cohort of 62 patients starting dialysis (either hemodialysis or continuous ambulatory peritoneal dialysis) during 1993 with a 4-year follow-up. RESULTS: The mean delay between either first symptoms of renal disease, or first evidence of renal failure and nephrologist referral was 10 years and 3 years 56 days, respectively. About 47% of the patients were referred less than 6 months before starting dialysis, and 27.5% less than 1 month. Blood pressure levels were higher in patients referred less than 6, 3 and 1 month (P < 0.05), as was creatinine concentration in patients referred less than 1 month (P < 0.05). In contrast, plasma calcium was lower for referral less than 6 months (P < 0.05) and 3 months (P < 0.005), as was bicarbonate concentration for referral less than 3 and 1 month (P < 0.05). Initial hospitalisation stay was prolonged (x1.5) for late referral less than 3 months (56.4 +/- 39 days vs 35.9 +/- 33.6 days, P < 0.05) as was 6 months hospitalisation length for referral less than 3 months (x1.6) (52.9 +/- 40.6 days vs 33.2 +/- 28.7 days, P < 0.05) and less than 1 month (x1.8) (61 +/- 45 days vs 33.9 +/- 28.7 days, P < 0.05) and < 1 month (x1.8) (61 +/- 45 days vs 33.9 +/- 28.7 days, P < 0.05). Only 44.1% of patients started hemodialysis with a functioning arteriovenous fistula, and patients requiring temporary access had a 4.4-fold longer initial (60.1 +/- 41.7 days vs 13.6 +/- 11.6 days, P < 0.005) and 6-month (59.6 +/- 39 days vs 13.6 (9.1, P < 0.005) hospitalisation stay. The four-year mortality rate was unaffected by the delayed referral but strongly and independently predicted by age, diabetes and hypoalbuminemia. CONCLUSION: Early nephrologic referral and timely initiated dialysis decrease morbidity at the start of dialysis and both hospitalisation length and costs. PMID- 11270268 TI - [Genetic hypokalemia]. AB - INTRODUCTION: Hypokalemia is the most frequent electrolytic disturbance in hospitalized patients. It is sometimes familial. Careful clinical and biological evaluation may guide further genetic analysis. CURRENT KNOWLEDGE AND KEY POINTS: Genetic hypokalemia is linked to disorders of mineralocorticoid hormone synthesis or action (glucocorticoid-remediable hyperaldosteronism, congenital adrenal hyperplasia, apparent excess of mineralocorticoids), to renal tubular disorders (Liddle's syndrome, Bartter's and Gitelmann's syndrome, tubular acidosis) or to disorders of cellular transfer of potassium (hypokalemic periodic paralysis). FUTURE PROSPECTS AND PROJECTS: Molecular mechanisms of adult Bartter's syndrome are probably different from pediatric syndromes. A better clinical and biological evaluation with longitudinal follow-up could allow significant progress in the knowledge of the natural history and prognosis of these syndromes. PMID- 11270269 TI - [Role and modalities of insulin treatment in type 2 diabetics]. AB - INTRODUCTION: The natural history of type 2 diabetes mellitus is characterized by an inescapable and gradual worsening of a decrease in insulin secretion. Thus after several years of progress, less than half of type 2 diabetic patients have good glycemic control. This explains the increase in insulin prescription to type 2 diabetic patients in France in recent years. This work's objective is to take into account recent publication data to clarify the status of and adjustments in insulin therapy. CURRENT KNOWLEDGE AND KEY POINTS: The benefit of insulin treatment-mediated glycemic control optimization on microvascular complications is now proven. However, there is still controversy concerning macrovascular complications. Hypoglycemic risk in type 2 diabetic patients is limited and the main problem with insulin treatment is weight gain. Following failure with treatment by tablets, the most suitable treatment in terms of metabolic improvement, weight gain limitation and treatment adhesion is to add an intermediate insulin injection at bedtime. The next step remains several injections a day, with metformine addition if possible. FUTURE PROSPECTS AND PROJECTS: Therapeutic treatment in type 2 diabetes mellitus may become an earlier start of insulin therapy to preserve the remaining pancreatic insulin reserve. The role of brief and long-lasting insulin analogues, as well as inhaled insulin, which will soon be available, should be specified. PMID- 11270270 TI - [Is alopecia areata a psychosomatic disease?]. AB - INTRODUCTION: Destruction of hair follicles by lymphocytes induces alopecia aerata. Hence, immunological mechanisms are involved in this disease, as for numerous dermatoses. Nonetheless, alopecia aerata appears to be a psychosomatic disease. Is there any contradiction? CURRENT KNOWLEDGE AND KEY POINTS: Alopecia aerata often occurs after stress, particularly during mourning. Psychiatric disorders are more frequent in patients with alopecia than in healthy subjects. But these disorders might be secondary to the visible hair disease. Psychopathological mechanisms need to be clarified but alexithymia seems to be the key for understanding how stress could induce hair loss. In the skin (and the scalp) all functions are narrowly controlled by nerve fibers. Among these functions are hair growth and immunity. Immune cells and hair follicle cells possess receptors for neurotransmitters, which are synthesized by neuronal endings. When activated, these receptors are able to modulate cell properties. The same phenomena are described with stress-induced hormones. In alopecia aerata, like in numerous other diseases, psychosomatics and immunology are not opposed because immune cells are controlled by the nervous system through neurotransmitters. FUTURE PROSPECTS AND PROJECTS: Research needs to be thorough in both the fields of psychology and neurobiology. Psychotherapies or psychotropes appear to be useful in the treatment of alopecia aerata. PMID- 11270271 TI - [Hemolytic anemia disclosing Wilson's disease. Report of 2 cases]. AB - INTRODUCTION: The liver and central nervous system are the usual targets of Wilson's disease, an inherited disorder of copper metabolism. Severe hemolytic anemia is an unusual complication of Wilson's disease. EXEGESIS: We report two cases of Wilson's disease revealed by acute intravascular hemolytic anemia associated with liver failure. Blood smear analysis showed stippled red cells in one case; hemolytic anemia improved within a few weeks in both patients but progressive liver failure required transplantation in the other. Hemolysis probably results from the toxic effect of free serum copper on erythrocyte membrane. CONCLUSION: Diagnosis of Wilson's disease must be considered in case of acute hemolytic anemia associated with liver failure in young adults. PMID- 11270273 TI - [Macro-CK disclosing disseminated epidermoid carcinoma of the pyriform sinus in a patient with dermatomyositis]. AB - INTRODUCTION: The association between dermatomyositis and cancer is clearly established, but its frequency remains difficult to define. EXEGESIS: We report the case of an association between a dermatomyositis and a cancer of the piriform antrum. Four months after surgical treatment and radiotherapy, the increased macro-CK level gave us reason to suspect a cancer relapse with pulmonary, hepatic, splenic and renal metastases without progressive clinical signs. CONCLUSION: Mitochondrial macro-CK detection must evoke the presence of neoplasia with or without metastasis, which would be a poor prognosis, as has been shown in our patient. PMID- 11270272 TI - [Pulmonary hyalinising granuloma: report of 2 original cases with cervicofacial and orbital involvement]. AB - INTRODUCTION: Pulmonary hyalinizing granuloma is a rare fibrosing nodular disease of the lung characterized by its histological appearance, which includes at the center of the lesion a dense network of concentric hyalinized collagen lamella surrounded by perivascular lymphoplasmacytic infiltrate that rarefies in the center of the nodule. EXEGESIS: We report two new cases: the first with laryngeal (endoluminal tumor-like), orbital (subeyelid nodule) and mesenteric (9 x 6 cm mass) location of hyalinizing granuloma; the second with cervical, facial (trismus), orbital (pseudotumor) and limb (ankylosing elbow) fibrosis. CONCLUSION: The extrapulmonary diffusion of the disease is extremely rare. In one of the cases, with corticosteroids and after a follow-up of 12 months, the pulmonary tumors vanished but the fibrosis resolved only partially. PMID- 11270274 TI - [Neurologic toxicity caused by zelitrex (valaciclovir) in 3 patients with renal failure. Is overdose associated with improvement of product bioavailability improvement?]. AB - INTRODUCTION: We report three cases of neurotoxicity in patients with renal failure, treated with Zelitrex (valacyclovir). EXEGESIS: The patients are two women and a man, aged 76 +/- 4.6 years, who presented acute mental confusion during a treatment with valacyclovir. In two cases, the patients previously had altered renal function and were under peritoneal dialysis. In the last case, the patient had simultaneous neurotoxicity and acute renal failure. After the discontinuation of the drug, the outcome was favourable in all cases. CONCLUSION: Our cases focus attention on the possible neurotoxicity of valacyclovir, which is an amino acid ester prodrug of acyclovir, rapidly and almost completely hydrolysed to acyclovir prior to systemic exposure. The bioavailability of valacyclovir is 54% compared to approximately 20% for oral acyclovir and may account for unexpected overdoses, which may lead to serious neurological toxicity. PMID- 11270275 TI - [ECG of a 100-year-old female patient]. PMID- 11270276 TI - [Value of methylprednisolone perfusions in the corticodependent forms of Horton's disease]. PMID- 11270278 TI - [Common variable immunodeficiency of late onset complicated with granulomatosis]. PMID- 11270277 TI - [Gynecomastia caused by diabetic mastopathy: 2 cases]. PMID- 11270279 TI - [Atypical focal nodular hyperplasia]. PMID- 11270280 TI - [Influence of the family history of alcohol consumption in men and women]. AB - OBJECTIVE: To assess the risk for alcohol abuse among individuals with a positive family history of alcohol abuse (FH+). MATERIAL AND METHODS: The study population was a sample (n = 8,890) drawn from a 1988 national survey on addictions in Mexico City's urban population. Data analysis consisted of frequency and association measures, using family history of alcohol abuse as the exposure factor. RESULTS: Prevalence of heavy drinking was 13.7% for males and 0.6% for females. Alcohol dependence syndrome was found in 9.9% of males and 0.6% of females. Men with HF+ were twice more likely to develop dependence syndrome than HF- males. The odds ratio for women was 1.27. CONCLUSIONS: Differential patterns by gender were found for familial transmission of alcohol abuse; parental alcohol intake is a main risk factor for developing alcohol dependence syndrome. PMID- 11270281 TI - [Prevalence of Moraxella catarrhalis colonization in asymptomatic carriers under 6 years of age]. AB - OBJECTIVE: To determine the prevalence of upper respiratory tract colonization by Moraxella catarrhalis in children under six years of age. MATERIAL AND METHODS: A survey was conducted between January and December 1998 in Mexico City, among children aged 2 months to 5 years, selected through cluster sampling. Pharyngeal samples were taken for M. catarrhalis identification. The minimal inhibitory concentration to different antibiotics was obtained and beta-lactamases were determined by the iodometric test. Statistical analysis consisted of frequency distributions, odds ratios, 95% confidence intervals, and Mantel-Haenszel chi 2. Statistical significance was set at p < 0.05. RESULTS: After excluding 37 children, the study population was 604 children from Mexico City; M. catarrhalis was present in 130 pharyngeal specimens (22.9%). Most of the strains were positive for beta-lactamase production (75.4%). Eighty percent of the strains was resistant to penicillin and 70% to ampicillin and amoxicillin. None were resistant to cefotaxime, imipenem, meropenem and erythromycin. CONCLUSIONS: Prevalence of M. catarrhalis upper respiratory tract colonization is similar to that of other respiratory pathogens. These findings warrant future research on the role of M. catarrhalis as an etiologic agent in acute and chronic respiratory infections in Mexico. PMID- 11270282 TI - [Activity-based cost model applied to tracer cardiovascular diseases]. AB - OBJECTIVE: To analyze the costs of outpatient care on tracer ischemic cardiovascular diseases events in public healthcare institutions. MATERIALS AND METHODS: The study was carried out from April to October 1998, on a sample of 2,000 (290 tracer diseases and 1,710 non-tracer diseases) first-time outpatient visits at the San Roque de Connet General Hospital, Buenos Aires, Argentina. Costs were evaluated using the Activity-Based Cost (ABC) method. RESULTS: Outpatient care activity improvements would result in significant savings in indirect costs of 7.11% on average for products defined as high blood pressure, dyslipemia and diabetes. Total savings in unit cost per product from elimination of activities would be 11.78% for high blood pressure, 13.96% for dyslipemia, 19.05% for diabetes, and 11.45% for non-tracer diseases. A total of 66.26% of the total indirect costs corresponding to dyslipemia and 61.80% of the total indirect costs corresponding to diabetes were inefficiently allocated or misspent. The total unit cost of medical care assessed by the traditional method is $22.98, a figure that in some cases is quite below the cost obtained by the ABC method used in this study. CONCLUSIONS: It is necessary to work on re-designing the patient healthcare process, to evaluate the activities which do not add any value, and that turn out to be a nuisance and delay for the patient. These activities make the system inefficient since resources are allocated to activities that hinder the process and that are therefore charged to the cost of medical visits. PMID- 11270283 TI - [Practice of traditional medicine in Latin America and the Caribbean: the dilemma between regulation and tolerance]. AB - OBJECTIVE: This paper characterizes the current stage of traditional medicine in nine countries of Latin America and the Caribbean. MATERIAL AND METHODS: This qualitative study was conducted between March and December 1998. Data were collected on the components of traditional health systems in countries of Latin America and the Caribbean, by means of a network of individuals and institutions from different countries that acted as expert informants from different specialty areas. RESULTS: Findings from the analysis of traditional medicine regulation are presented in three groups: a) Countries with some developments in the area of legislation; b) Countries where legislation is underway; and, c) Countries with no legislation or incipient regulation. CONCLUSIONS: Several stages of traditional medical practice legislation are found in the region. This heterogeneity shows the complexity involved in regulating the practice of providers with low levels of formal training, with different therapeutic practices, and with customs that are frequently difficult to include within the standards of the official health system. These findings are important for designing and implementing healthcare policies to adequate traditional medical practices to the needs of populations that commonly use them. PMID- 11270284 TI - [Comments on Psychosocial situation of adolescents and toxemia of pregnancy]. PMID- 11270286 TI - Toward better access to health insurance coverage for U.S. retirees in Mexico. AB - Many retirees from the United States of America have limited health insurance coverage while living in Mexico. Medicare and Medicaid benefits are not portable to other countries and Medigap (private insurance that supplements Medicare) is very limited. This causes economic and medical hardships and serves as a barrier to retirement to Mexico. Increasing numbers of U.S. retirees will be interested in moving to Mexico in the future because of the climate, the culture, and the lower cost of living. The numbers are increasing as a result of several factors such as aging "baby boomers" and the rapidly growing Mexican-origin population in the U.S.A. who are citizens or permanent residents but would like to return to their communities of origin after working in the U.S.A. There are several policy initiatives that could provide opportunities for improving health insurance coverage for these retirees that could be cost-effective. PMID- 11270285 TI - [Prevention of maternal RhD isoimmunization with anti-D gamma isoimmunization]. AB - OBJECTIVE: To report our experience in preventing RhD maternal isoimmunization by using anti-D gamma globulin among Rh-negative women. MATERIAL AND METHODS: Between 1982 and 1995, immunologic and hematologic data were collected from all Rh-negative women seen at Mexico's National Perinatology Institute. Women at risk of Rh isoimmunization were given a prophylactic dose of 150 micrograms of anti-D gamma globulin. RESULTS: A total of 4,857 Rh-negative women were seen during the study period (4.85% of the total population of women seen at the Institute), 629 (13.0%) of whom developed RhD isoimmunization; 542 (86.2%) of these women were already isoimmunized when first seen at our Institute. Twenty-two women (3.5%) developed isoimmunization even after receiving a proper dose of anti-D gamma globulin. Prophylaxis was given to 2,605 women (53.6%); 2,039 received a single dose, and 475 two doses. Prophylaxis failed in 22 cases; four were women with multiple pregnancy and 18 developed obstetric pathologic conditions. CONCLUSIONS: The use of anti-D gamma globulin resulted in a reduction of maternal Rh isoimmunization to less than one case per 1,000 women. Failures to prevent isoimmunization were associated to additional obstetric conditions and to lack of adherence to prevention guidelines. PMID- 11270287 TI - [Learn more about the counter-role of women physicians, proposes a reader of an article about the identity and experiences of these professionals]. PMID- 11270288 TI - [Eating disorders and personal behavioral control]. AB - OBJECTIVE: To explore the three components of personal behavioral control: Objective control, subjective control and control beliefs among normal women, women at risk of anorexia or bulimia and women with sub-clinical eating disorders. MATERIAL AND METHODS: In 1997, a cross-sectional study was conducted in Caracas, Venezuela. The study population consisted of 87 women: 21 with sub clinical eating disorders, 33 at risk of having an eating disorder, and 33 normal women. Measurement instruments used were: Eating Attitude Test, Composite International Diagnostic Interview, Self-control Inventory, Inventory for Perceived Self-efficacy for Self-control, and the Locus of Control Inventory. Data analysis consisted of analysis of variance, and post hoc comparisons were done with the Student-Neuman-Keuls test. RESULTS: Women with sub-clinical eating disorders were found to have difficulty for behavior self-control, lower self efficacy to regulate their behavior, and beliefs of control by powerful others over their behavior and its consequences. CONCLUSIONS: The results from this study are a first attempt to understand the role of the psychological variable "personal control of behavior", as a protective or risk factor for developing sub clinical anorexia or bulimia. PMID- 11270289 TI - Original and practical. PMID- 11270290 TI - Botanical briefs: garlic--Allium sativum. PMID- 11270291 TI - Photo quiz. Intradermal tophaceous gout. PMID- 11270292 TI - Cutaneous T-cell lymphoma at a young age. AB - A case of cutaneous T-cell lymphoma (CTCL) in a 22-month-old patient is discussed, emphasizing the importance of screening for CTCL even in very young patients with atypical symptoms of eczema, atopic dermatitis, or parapsoriasis. The clinical, histologic, and immunologic diagnostics can now be supported by molecular methods; therefore, patients at the earlier stages of CTCL can be diagnosed and treated with good results. PMID- 11270293 TI - Sneddon's syndrome: a case report. AB - We report a case of Sneddon's syndrome with the triad of livedo reticularis, hypertension, and neurologic symptoms. The procedures for diagnosis and the tests to delineate clotting abnormalities are examined. PMID- 11270294 TI - Acantholytic dermatosis of the vulvocrural area. AB - Acantholytic dermatosis of the vulvocrural area is a rare skin disorder characterized by solitary or multiple skin-colored to white, smooth papules or plaques. Histopathological features of both Hailey-Hailey disease and Darler's disease are present. There is acantholysis, which may involve the full thickness of the epidermis, and dyskeratosis with corps ronds and grains. There may be marked hyperkeratosis and focal parakeratosis. We report a case of this rare disease and discuss its differential diagnosis and treatment. PMID- 11270295 TI - Unilateral acquired nevus flammeus in women. AB - Congenital nevus flammeus is a benign vascular tumor characterized by pink to pale red patches that thicken as the patient ages, producing a dull red to reddish blue, cobblestone-textured plaque. We present the cases of 3 women with unilateral acquired nevus flammeus on the cheek whose lesions resolved after minimal treatment with a 585-nm pulsed dye laser. The etiology of acquired nevus flammeus is reviewed and tumor response rates to laser surgery are discussed. PMID- 11270296 TI - Lymphangioma circumscriptum of the vulva. AB - Lymphangioma circumscriptum is a benign disease of the lymph ducts and an unusual pathologic process that rarely affects the vulva. The etiology of this lesion is not clear, but obstruction of the lymph vessels has been suggested as a possible cause in some cases. We report the case of a 44-year-old woman with lesions similar to lymphangioma circumscriptum of the vulva and chronic idiopathic lymphedema of the lower right limb. Because there was no obvious cause, we propose that the lymphangioma was caused by the lymphedema. PMID- 11270297 TI - Vulvodynia: an indicator or even an early symptom of vulvar cancer. AB - Vulvodynia is a symptom of chronic, painful vulvar discomfort of multicausal origin. Vulvar cancer is an underestimated cause of vulvodynia. Even early stages of vulvar neoplasia can lead to aching lesions. Three cases of vulvar carcinoma eliciting persistent pain have been diagnosed within a 2-year period. In 2 of our case studies, women had antecedent periods of vulvar pruritus of long duration (5 and 20 years, respectively). We conclude that early histologic examination of all visible vulvar lesions is necessary to exclude the presence of malignant vulvar neoplasia. PMID- 11270298 TI - Acquired vulvar lymphangiomas: a sequela of radiation therapy. AB - Lymphangiomas of the vulva are rare clinical entities. Acquired or secondary lymphangiomas have characteristically been reported after radiation therapy for cervical carcinoma and appear on the vulva years after this treatment. Local surgery, scrofuloderma, and Crohn's disease may also damage vulvar lymphatic flow and lead to the development of vulvar lymphangiomas. We report a case of acquired vulvar lymphangiomas that occurred in a patient 15 years after she received radiation therapy for squamous cell carcinoma of the uterine cervix. PMID- 11270300 TI - Tufted hair folliculitis after scalp injury. AB - We describe the case of a 38-year-old epileptic man with tufted hair folliculitis. The condition started 5 years ago after a scalp laceration that had been sustained 3 months earlier during an epileptic crisis. There then appeared a circumscribed inflammatory bulging lesion (with exudation and crusts) that evolved to scarring alopecia with tufts of 20 to 30 apparently normal hair shafts. Results of bacteriologic examination of pus extruding from the dilated follicular ostia revealed Staphylococcus aureus. The cutaneous pathologic examination showed polymorphous inflammatory exudate in the upper and mid dermis, which was mostly perifollicular, and the presence of normal and independent follicles in the deep dermis, which, while ascending, converged to a common dilated follicular channel. The patient was treated successively with oral flucloxacillin, erythromycin, ciprofloxacin, and amoxicillin/clavulanic acid and with topical application of erythromycin, clindamycin, povidone iodine, and ketoconazole. Transient improvement was followed by recurrence and enlargement of the affected area. PMID- 11270299 TI - Atypical mycobacterial infection in a patient with hairy cell leukemia. AB - A case of a cutaneous tumor caused by atypical mycobacterial infection (Mycobacterium kansasii) in a patient with hairy cell leukemia is reported. Surgical removal of the lesion and subsequent combination antituberculotic treatment led to a cure of this infection. Remission of the leukemia was achieved with interferon alfa. PMID- 11270301 TI - Annular lichen sclerosus et atrophicus. AB - Lichen sclerosus et atrophicus (LSA) is an idiopathic skin condition characterized by ivory-colored, atrophic papules and plaques. Many variants of LSA have been described. Only rarely has an annular variant been noted. We present a case of annular LSA and discuss the other reported cases exhibiting an annular shape. PMID- 11270302 TI - Nonendemic pemphigus foliaceus presenting as fatal bullous exfoliative erythroderma. AB - Pemphigus foliaceus is a cutaneous autoimmune blistering disease that is characterized by lower morbidity and mortality than those observed in pemphigus vulgaris or paraneoplastic pemphigus. However, erythrodermic forms of the endemic variant of pemphigus foliaceus have been associated with a higher mortality. We report a case of nonendemic pemphigus foliaceus that presented as fatal bullous exfoliative erythroderma, and thus, we will emphasize the inclusion of this entity in the differential diagnosis and the use of skin direct immunofluorescence in the evaluation of patients with erythroderma. PMID- 11270303 TI - Suspected induction of a pyoderma gangrenosum-like eruption due to sulpiride treatment. AB - A case of a pyoderma gangrenosum (PG)-like eruption due to the antipsychotic drug sulpiride, a form of risperidone, is described. The contribution of sulpiride to the etiology of the PG-like lesion is based on the reduction and healing of the ulcer upon cessation of the drug and the formation of a bulla following the drug's re-administration. The literature on drug-induced PG or PG-like eruptions is discussed. The selectivity of sulpiride for dopamine receptors and its limited effect on other neuronal pathways differentiates sulpiride from other types of antispychotic drugs commonly used in Israel, including phenothiazine, butyrophenone, and thioxanthene. Adverse systemic and cutaneous reactions to sulpiride and to risperidone are described. To our knowledge, this is the first report of a PG-like eruption due to the former. PMID- 11270304 TI - Efficacy and safety of terbinafine 1% solution in the treatment of interdigital tinea pedis and tinea corporis or tinea cruris. AB - Two randomized, double-blind, vehicle-controlled, multicenter studies assessed the efficacy and safety of a new terbinafine 1% solution for the treatment of interdigital tinea pedis and tinea corporis or tinea cruris (tinea corporis/cruris). Patients with interdigital tinea pedis applied terbinafine 1% solution or vehicle twice daily for 1 week with 7 weeks of follow-up (N = 153), and patients with tinea corporis/cruris applied terbinafine 1% solution or vehicle once daily for 1 week with 3 weeks of follow-up (N = 66). Efficacy was assessed mycologically and clinically at the end of treatment and throughout follow-up. In the tinea pedis study, 66% of patients were effectively treated with terbinafine compared with 4% of the group treated by vehicle (P < .001; Mantel-Haenszel test). In the tinea corporis/cruris study, treatment was effective in 65% of the terbinafine group compared with 8% of the vehicle group (P < .001). There were no significant differences in the frequency of cutaneous adverse events between the 2 groups in either study. We conclude that one week of therapy with terbinafine 1% solution is highly effective, superior to vehicle, and safe for use in superficial fungal infections. PMID- 11270306 TI - Hospital interventions improve care of North Carolina Medicare patients with acute myocardial infarction. PMID- 11270305 TI - The original Siamese twins. We know why Chang died, but why did Eng? PMID- 11270307 TI - Urinary incontinence. No need to be wet and upset. AB - Over 17 million people in the United States suffer from urinary incontinence. UI limits functional and social activities and is a common cause of anxiety, social withdrawal, and depression. Primary care providers should take a proactive approach in searching for the presence of, and then investigating the reasons for, incontinence. Usually the diagnosis is apparent from the data obtained from a good history, physical examination, measurement of post-voiding residual urine, and urinalysis. A voiding diary can quantify the magnitude of the problem and response to treatment. Conservative treatment options include behavioral techniques such as timed or prompted voiding, changes in diet, pelvic floor exercises, and medications. If these measures fail, referral to a urological surgeon is appropriate so that further diagnostic studies can be offered as well as minimally invasive procedures or surgery. PMID- 11270308 TI - 'DOC.TOR n. [fr. Latin docere, to teach]. A person trained in the healing arts. PMID- 11270309 TI - A rare congenital condition discovered (happily) late in life. Tetralogy of Fallot with absent pulmonary valve. PMID- 11270310 TI - Hospice. Care when there is no cure. PMID- 11270311 TI - The charms of music. Step by step prescription for patients. PMID- 11270312 TI - Depression and vascular function in older adults. Evaluating the benefits of exercise in a new study at Duke University. PMID- 11270313 TI - Health and social problems of a primary care clinic population after a disaster. The Hurricane Floyd flood. PMID- 11270314 TI - Prescription and adherence to statins of patients with coronary artery disease and hypercholesterolemia. AB - OBJECTIVE: Statins have proved to be safe and effective in the secondary prevention of coronary artery disease, but the level of prescription and the reasons for nonadherence to treatment in many coronary diseases treatment centers has not been determined. The purpose of this study was to identify reasons for nonadherence to statin therapy. METHODS: We analyzed 207 consecutive patients with coronary artery disease and hypercholesterolemia (total cholesterol > or = 200 mg/dL or LDL-cholesterol > or = 130 mg/dL). Patients' average age was 61.7 +/ 10 year; 111 (53.6 %) male were and 94 (46.6 %) were female. We analyzed the level of prescription and adherence to treatment with statins. RESULTS: Statins were prescribed for 139 (67 %) patients, but only 85 (41 %) used the drug. In spite of being indicated, statins were not prescribed in 68 (33 %) patients. Of 54 (26 %) patients, nonadherent to statins, 67 % did not use the drug due to its high cost, 31 % due to the lack of instruction, and only 2 % due to side effects. Total cholesterol (260.3 +/- 42.2 vs 226.4 +/- 51.9; p < 0.0001) and LDL cholesterol (174.6 +/- 38.1 vs 149.6 +/- 36.1; p < 0.0001) were lower in patients on medication. HDL-cholesterol increased from 37.6 +/- 9.6 to 41.5 +/- 12.9 mg/dL (p = 0.02), and triglycerides were not modified in patients using statins. CONCLUSION: The prescription of statins in patients with coronary artery disease and dyslipidemia is high; however, its adherence is far from satisfactory, due to the high cost of the medication. Reduction in total cholesterol and LDL cholesterol levels did not reach the targets recommended by the Brazilian Consensus on Dyslipidemia. PMID- 11270315 TI - Clinical and laboratory evaluation of hyperlipemic and hypothyroid patients. AB - OBJECTIVE: To determine the frequency of hypothyroidism in a sample of hyperlipemic patients and evaluate clinical and laboratory factors indicative of thyropathy among them. METHODS: Fifty-one hyperlipemic patients, grouped according to an earlier or recent diagnosis of their thyroid function into euthyroid and hypothyroid, were evaluated with clinical and laboratory examinations of blood levels of free T4 and TSH (by radioimmunoassay). Patients were on average 46.8 +/- 11.7 years old, predominantly of the female sex (62.5 %); 31 % had a previous diagnosis of hypothyroidism and were under treatment with thyroxin. RESULTS: Fourteen three percent of patients analyzed had hypothyroidism, which had not been detected before. Differentiating attributes of the groups analyzed were: a predominance of females among the hypothyroid patients and a higher HDL serum concentration among those recently diagnosed. CONCLUSION: In the present study, new cases of hypothyroidism in hyperlipemic patients were a frequent occurrence, yet few clinical and laboratory data except tests evaluating free T4 and TSH in the blood indicated which patients had thyroid dysfunction. PMID- 11270316 TI - Congestive heart failure. Correlation between functional class and systolic and diastolic functions assessed by Doppler echocardiography. AB - OBJECTIVE: To evaluate the influence of systolic or diastolic dysfunction, or both on congestive heart failure functional class. METHODS: Thirty-six consecutive patients with a clinical diagnosis of congestive heart failure with sinus rhythm, who were seen between September and November of 1998 answered an adapted questionnaire about tolerance to physical activity for the determination of NYHA functional class. The patients were studied with transthoracic Doppler echocardiography. Two groups were compared: group 1 (19 patients in functional classes I and II) and group 2 (17 patients in functional classes III and IV). RESULTS: The average ejection fraction was significantly higher in group 1 (44.84 % +/- 8.04 % vs. 32.59 % +/- 11.48 % with p = 0.0007). The mean ratio of the initial/final maximum diastolic filling velocity (E/A) of the left ventricle was significantly smaller in group 1 (1.07 +/- 0.72 vs. 1.98 +/- 1.49 with p = 0.03). The average maximum systolic pulmonary venous velocity (S) was significantly higher in group 1 (53.53 cm/s +/- 12.02 cm/s vs. 43.41 cm/s +/- 13.55 cm/s with p = 0.02). The mean ratio of maximum systolic/diastolic pulmonary venous velocity was significantly higher in group 1 (1.52 +/- 0.48 vs. 1.08 +/- 0.48 with p = 0.01). A predominance of pseudo-normal and restrictive diastolic patterns existed in group 2 (58.83 % in group 2 vs. 21.06 % in group 1 with p = 0.03). CONCLUSION: Both the systolic dysfunction index and the patterns of diastolic dysfunction evaluated by Doppler echocardiography worsened with the evolution of congestive heart failure. PMID- 11270317 TI - Cardiac extension of primary mediastinal seminoma compressing the right ventricular outflow tract. AB - We report the case of a 33-year-old male with primary seminoma of the anterior mediastinum with initial clinical manifestations suggestive of heart disease. PMID- 11270318 TI - Alcohol and atherosclerosis. AB - Observational studies have attributed a protective effect to alcohol consumption on the development of atherosclerosis and cardiovascular morbidity and mortality. Alcohol intake in the amount of one to two drinks per day results in an estimated 20-40% reduction in cardiovascular events. An additional protective effect, according to major cohort studies, has been attributed to wine, probably due to antioxidant effects and platelet antiaggregation agents. On the other hand, the influence of different patterns of alcohol consumption and environmental factors may explain a great part of the additional effect of wine. Protection may be mediated by modulation of other risk factors, because alcohol increases HDL-C, produces a biphasic response on blood pressure, and modulates the endothelial function, while it neither increases body weight nor impairs glucose-insulin homeostasis. Alcohol may also have a direct effect on atherogenesis. Despite these favorable effects, the current evidence is not enough to justify prescribing alcohol to prevent cardiovascular disease. PMID- 11270319 TI - Myths about cardiovascular diseases. PMID- 11270320 TI - Dynorphin A analogs containing a conformationally constrained phenylalanine derivative in position 4: reversal of preferred stereochemistry for opioid receptor affinity and discrimination of kappa vs. delta receptors. AB - Analogs of the opioid peptide [D-Ala8]dynorphin A-(1-11)NH2 containing optically pure (R)- and (S)-2-aminotetralin-2-carboxylic acid (Atc) in position 4 were synthesized and evaluated for opioid receptor affinity. These peptides are the first reported dynorphin A analogs containing a conformationally constrained amino acid in place of the important aromatic residue Phe4. By incorporating resolved Atc isomers, the opioid receptor affinity and the stereochemistry of the constrained residue could be unambiguously correlated. Both Dyn A analogs containing Atc in position 4 retained nanomolar affinity for kappa and mu opioid receptors. Unexpectedly the peptide containing (R)-Atc, corresponding to a conformationally constrained D-Phe analog, displaying higher affinity for both kappa and mu receptors than the peptide containing (S)-Atc. In contrast [D-Phe4,D Ala8]Dyn A-(1-11)NH2 exhibited significantly lower affinity for kappa and mu receptors than the parent peptide, as expected. Conformational restriction of the Phe4 sidechain or incorporation of D-Phe in position 4 had the largest effect on delta receptor affinity, yielding compounds with negligible affinity for these receptors. Thus, there appear to be distinctly different structural requirements for this residue for kappa vs. delta receptors, and it is possible to completely distinguish between these two receptors by changing a single residue in Dyn A. PMID- 11270321 TI - Enantioselective pharmacokinetics of the enantiomers of clevidipine following intravenous infusion of the racemate in essential hypertensive patients. AB - The aim of the study was to characterize the individual pharmacokinetics of (-)-R and (+)-S-clevidipine following intravenous constant rate infusion of rac clevidipine to essential hypertensive patients. Twenty patients received three out of five randomized treatments with clevidipine. The pharmacokinetics of the separate enantiomers were evaluated by compartmental analysis of blood concentrations vs. time curves using the population approach. The derived pharmacokinetic parameters were used to simulate the time for 50 and 90% postinfusion decline following various infusion times of rac-clevidipine. A two compartment model was used to describe the dispositions of the enantiomers; there were only minor differences between the estimated pharmacokinetic parameters of the separate enantiomers. The mean blood clearance values of (-)-R- and (+)-S clevidipine were 0.103 and 0.096 l/min/kg, and the corresponding volumes of distribution at steady state were 0.39 and 0.54 l/kg, respectively. The context sensitive half-time was approximately 2 min regardless of stereochemical configuration, and a 90% decline in concentration was achieved approximately 8 min postinfusion for (-)-R-clevidipine and 11 min for (+)-S-clevidipine, following clinically relevant infusion times with clevidipine. In conclusion, both enantiomers are high-clearance compounds with similar blood clearance values. The volume of distribution for the enantiomers is slightly different, presumably due to differences in the protein binding. From a pharmacokinetic point of view, the use of a single enantiomer as an alternative to the racemic clevidipine will not offer any clinical advantages. PMID- 11270322 TI - Synthesis of the antistroke drug lubeluzole and its enantiomer. Lipase-catalyzed resolution of chiral building block. AB - 1-Chloro-3-(3,4-difluorophenoxy)-2-propanol was kinetically resolved by lipase catalyzed esterification with vinyl butanoate in organic medium to yield the (S) butanoate and the (R)-alcohol as the remaining substrate. In an enantioconvergent synthesis the mixture was subject to Mitsunobu esterification in one pot which converted the (R)-alcohol to the (S)-ester. The (S)-butanoate was hydrolyzed by lipase catalysis to give (S)-1-chloro-3-(3,4-difluorophenoxy)-2-propanol. The two enantiopure chiral building blocks were used for synthesis of Lubeluzole and its enantiomer respectively. PMID- 11270323 TI - High-performance liquid chromatographic analysis of the sulfation of 4 hydroxypropranolol enantiomers by monkey liver cytosol. AB - We developed a new high-performance liquid chromatographic method using an ODS column and a chiral column for the assay of racemic 4-OH-PL sulfate and enantiomeric 4-OH-PL sulfates, respectively. The method was successfully applied to measure phenolsulfotransferase (PST) activities for 4-OH-PL in cytosolic fractions from livers of Japanese monkeys (Macaca fuscata) and for comparison with its activity of cytosolic fractions from rat, rabbit, dog, and human livers and Hep G2 cells. The activity was ranked as Hep G2 cells > monkeys = humans = dogs = rats > rabbits. To evaluate the Japanese monkey as a nonhuman animal model in drug metabolism studies, we further characterized sulfation of 4-OH-PL as a further metabolic pathway in monkey livers to compare that with human livers. Inhibition studies in which cytosolic fractions were preincubated at 43 degrees C or 2,6-dichloro-4-nitrophenol (DCNP) used as a PST inhibitor indicated that two kinds of PSTs, thermolabile, low-Km and DCNP-resistant PST and thermostable, high Km and DCNP-sensitive PST were involved in 4-OH-PL sulfation by monkey liver cytosol, which is very similar to the reported profile of 4-OH-PL sulfation by human liver cytosol. Sulfation kinetics in a low concentration range of 4-OH-PL enantiomers demonstrated that apparent Km values were similar between human and monkey liver cytosolic fractions, but the Vmax values were different, so that intrinsic clearance values (Vmax/Km, Clint) were higher in monkeys than in humans. Furthermore, enantiomer selectivity of [R(+)-4-OH-PL > S(-)-4-OH-PL] was observed in the Vmax and CLint values of monkey liver cytosol. These results indicate that the profile of sulfation of 4-OH-PL by liver cytosolic fractions is similar in humans and Japanese monkeys. PMID- 11270325 TI - Composition and chirality of amino acids in aerosol/dust from laboratory and residential enclosures. AB - Initial results from the analyses of geological and anthropological samples for amino acids were difficult to accept because of the high enantiomeric purities of the analytes (i.e., predominantly L-amino acids). Consequently, sources of contamination had to be considered. All sources were eliminated except for direct atmospheric contamination. Essentially invisible, microscopic, aerosol/dust was found to rapidly contaminate the surface of samples and sample containers even after brief exposure times in clean laboratories. Contamination increased with exposure time. The aerosol/dust amino acids were contained predominantly in a proteinaceous material. Aerosol/dust from different locations can contain different percentages of proteinoid/amino acid material. However, the relative concentrations of the amino acids were similar for both laboratory and residential samples. The enantiomeric purity of the L-amino acids studied in aerosol/dust appears to be 99% or greater for the samples examined. Thus, even slight contamination of any sample with microscopic dust or aerosol particles can skew the results of trace amino acid analyses and amino acid e.e. determinations. PMID- 11270324 TI - Enantioselectivity of 3-amino-2-oxazolidinone derivatives with potential psychotropic activity on cellulose tris(4-methylbenzoate) chiral selector. AB - The behavior of a series of 3-amino-2-oxazolidinone derivatives with a potential hypnotic activity on achiral (octadecylsilane) and chiral (cellulose tris(4 methylbenzoate)) stationary phases was examined. The compounds differed in the composition of a substituted aromatic ring containing different substituents in different positions. It was possible to resolve all the compounds with selectivity 1.11 < or = alpha < or = 2.74. The enantiodifferentiating power of substituents was correlated to their electron donating ability and position in the aromatic ring. PMID- 11270326 TI - From 6-t-butylfulvene to highly functionalized five-membered rings. AB - 6-t-Butylfulvene was used as the starting material for asymmetric dihydroxylation. The obtained protected diol (2) was used for several stereoselective functionalizations such as epoxidations, hydroboration, and aminohydroxylation. However, these fulvene derivatives proved to be rather unreactive and gave no conversion under, e.g., palladium (0), catalyses or hydroboration conditions. PMID- 11270327 TI - Stereoselectivity of the folate transporter in rabbit small intestine: studies with amethopterin enantiomers. AB - Stereoselectivity of the folate transporter was examined using rabbit intestinal brush border membrane vesicles (BBMV). Methotrexate (MTX) and the antipode (D amethopterin) were used as model substrates of the transporter. Folic acid (FA) and MTX were actively taken up into BBMV in the presence of an H+ gradient. Initial uptake of FA and MTX was concentration-dependent with Km values of 1.5 and 1.6 microM for FA and MTX, respectively. FA and MTX mutually inhibited uptake in a competitive manner, with Ki values being similar to the corresponding Km values, demonstrating that FA and MTX share the folate transporter. D Amethopterin also inhibited FA uptake competitively, with a Ki value approximately 60-fold greater than that of MTX, showing that the affinity of the D-isomer (D-amethopterin) to the folate transporter is much less than that of the L-isomer (MTX). The extent of stereoselectivity observed in the present study is consistent with the previously reported differences in plasma concentration between amethopterin enantiomers following oral administration in humans. PMID- 11270328 TI - Topological diversity of artificial beta-barrels in water. AB - Rigid-rod beta-barrels are composed of interdigitating, short, amphiphilic peptide strands flanked by stabilizing rigid-rod "staves". We here report studies on the topological diversity of these recently devised artificial beta-barrels with regard to their length. For this purpose, homologous p-octiphenyl, p sexiphenyl, and p-quarterphenyl rods were equipped with complementary tripeptide strands based on the sequences Lys-Leu-Lys and Glu-Leu-Glu. The stability of rigid-rod beta-barrels of different length was determined by denaturation with guanidinium chloride. Free energies of delta GH2O = -5.2 kcalmol-1, delta GH2O = 2.9 kcalmol-1, and delta GH2O < -0.3 kcalmol-1 found for homologous p-octiphenyl, p-sexiphenyl, and p-quarterphenyl beta-barrels demonstrated strong dependence of beta-barrel stability on beta-barrel length. These results revealed a very qualitative minimal (approximately 23 A) and an "ideal" beta-barrel length (approximately 34 A), synergistic formation (alpha = 1.4) and remarkable stability for "ideal" p-octiphenyl beta-barrels exceeding that of several proteins and most synthetic models. Rigid-rod beta-barrels with p-oligophenyl "staves" longer than approximately 34 A will be very difficult to make and study because of rapidly decreasing rod solubilities. However, a strategy to bypass this apparent upper limitation of beta-barrel length is introduced: supramolecular matching of mismatched rods yielded elongated beta-barrels (61 A) of acceptable stability (delta GH2O = 2.2 - 3.1 kcalmol-1). PMID- 11270329 TI - Experimental study of the hydrodynamic behaviour of a high frequency ultrasonic reactor. AB - In relation to design and modeling of sonochemical reactors, the hydrodynamic behaviour of a high-frequency ultrasonic reactor has been investigated. Residence time distribution (RTD) measurements have been performed by means of a tracer method. The influence of ultrasound on the response to an inlet pulse was evidenced. It was shown that the reactor behaves like a completely stirred tank reactor (CSTR) as soon as ultrasonic irradiation operates. Preliminary observations on acoustic streaming occurring within the reactor will also be presented. PMID- 11270330 TI - Sonolysis of chlorobenzene in aqueous solution: organic intermediates. AB - The ultrasonic degradation of 1.72 mM chlorobenzene was investigated. The sonolysis of chlorobenzene followed first-order kinetics. The influence of the pH of the aqueous solution and the effect of the saturating gas, air or argon, was measured. No pH effect was noticed, and saturation with the monoatomic argon accelerated the degradation. Furthermore, the addition of the radical scavenger benzoate demonstrated that no significant degradation took place in the bulk solution. For air-saturated solutions, the following organic degradation products were identified: methane, acetylene, butenyne, butadiyne, benzene, chlorophenols, phenylacetylene and other chlorinated and non-chlorinated monocyclic and dicyclic hydrocarbons. For argon-saturated solutions, the same products were found, except for the chlorophenols. The presence of the chlorophenols in the case of air saturation only demonstrated the interaction between the radicals formed and oxygen, and no direct degradation by OH. radicals. The kinetics of several organic degradation products and chloride were determined for the sonolysis of air- and argon-saturated solutions. PMID- 11270331 TI - Influence of ultrasound power on the alkylation of phenylacetonitrile under solid liquid phase transfer catalysis conditions. AB - The influence of ultrasound power on the C-alkylation of phenylacetonitrile by ethyl bromide was studied under solid-liquid phase transfer catalysis in the presence of potassium hydroxide and tetrabutylammonium hydrogenosulfate. Experimental results are reported on the influence of the ultrasonic power on the yields. The optimum efficiency of ultrasounds is determined and the way in which ultrasound power may affect the yields is discussed. PMID- 11270332 TI - Sonochemical and thermal redox reactions of triphenylmethane and triphenylmethyl carbinol in nitrobenzene. AB - The reaction of triphenylmethane and triphenylcarbinol with nitrobenzene under thermal or ultrasonic activation was studied. It was shown beyond doubt that the thermal reaction of the aforementioned systems at 210 degrees C occurs through electron transfer. The sonochemical reactions occur at 40 degrees C, although slowly, while heating at the same temperature leaves the system unchanged. Electron transfers are also involved but an unexpected reductive process was evident. PMID- 11270333 TI - Acceleration of formaldehyde reactions with proteins due to ultrasound. AB - We have found that the reactions between caseine and formaldehyde or Urotropin are markedly increased under the influence of ultrasound. We also present a probable interpretation of the mechanism causing the acceleration of these reactions. PMID- 11270334 TI - Possible new route for the production of C60 by ultrasound. AB - The production of C60 by ultrasonic irradiation of liquid benzene has been studied. After irradiating 150 ml of liquid benzene for 1 h (600 W, 20 kHz), approximately 1 microgram of C60 is produced. PMID- 11270335 TI - Ultrasound assisted PTC catalyzed saponification of vegetable oils using aqueous alkali. AB - A few vegetable oils were saponified using aqueous KOH and different PTCs at room temperature in the presence of ultrasound. The extent of saponification was studied using the saponification value as a reference. Optimizations of various parameters such as time, selection of PTC, quantity of PTC, quantity of KOH and quantity of water were carried out using soyabean oil as a sample oil under sonication with stirring. To study the effect of ultrasound, the saponification was also carried out at 35 +/- 2 degrees C under different conditions, namely stirring, sonication, stirring and sonication, and heating at 100 degrees C. It was found that the heterogeneous liquid-liquid phase saponification of different vegetable oils using aq. KOH/CTAB was remarkably accelerated at 35 +/- 2 degrees C in the presence of ultrasound along with stirring. PMID- 11270336 TI - OH-radical formation by ultrasound in aqueous solution--Part II: Terephthalate and Fricke dosimetry and the influence of various conditions on the sonolytic yield. AB - Terephthalate and Fricke dosimetry have been carried out to determine the sonolytic energy yields of the OH free radical and of its recombination product H2O2 in aqueous solutions under various operating conditions (nature of operating gas, power, frequency, temperature). For example, in the sonolysis of Ar saturated terephthalate solutions at room temperature, a frequency of 321 kHz, and a power of 170 W kg-1, the total yield [G(.OH) + 2 G(H2O2)], equals 16 x 10( 10) mol J-1. This represents the total of .OH that reach the liquid phase from gas phase of the cavitating bubble. The higher the solute concentration, the lower the H2O2 production as more of the OH free radicals are scavenged, in competition with their recombination. Fricke dosimetry, in the absence and presence of Cu2+ ions, shows that the yield of H atom reaching the liquid phase is much lower, with G(H.) of the order of 3 x 10(-10) mol J-1. These sonolytic yields are smaller in solutions that are at the point of gas saturation, and increase to an optimum as the initial sonication-induced degassing and effervescence subsides. The probing of the sonic field has shown that the rate of sonolytic free-radical formation may vary across the sonicated volume depending on frequency and power input. PMID- 11270337 TI - Sonophotochemical destruction of aqueous solution of 2,4,6-trichlorophenol. AB - The combination of ultrasound and photochemistry have been used to degrade an aqueous solution of 2,4,6-trichlorophenol. An ultrasonic probe of 22 kHz frequency and a UV tube of 15 W have been used. Anatase grade TiO2 was used as the semiconductor catalyst. The effect of parameters such as ultrasonic intensity, operating conditions, type of ultrasonic equipment, and mode of UV transmission have been studied. The sonophotochemical degradation has been found to be dependent on the intensity of sonication, temperature of the reaction, and the type of ultrasonic equipment used, but was independent of the mode of UV transmission. Enhancement in the degradation rate has been observed at a higher sonication intensity and temperature of the solution. PMID- 11270338 TI - A comparative study of local sensors of power ultrasound effects: electrochemical, thermoelectrical and chemical probes. AB - In order to propose standard methods for the local measurement of the effects of power ultrasound inside a reactor, we compare three methods: a chemical dosimeter (Weissler reaction), a thermal sensor (embedded thermocouple) and an electrochemical probe (developed in our laboratory). The same emission device, i.e. the resonant tube (Sonitube-Sodeva), was used for all these methods. Similar trends were observed using various measurements: ultrasound effects vary significantly along the tube axis (due to standing waves in the resonant tubular emitter), but only slightly from the tube axis to the wall. More reliable and reproducible results were obtained with the thermal and electrochemical probes than with the chemical dosimeter. PMID- 11270339 TI - Sonochemical polymerization of benzene derivatives: the site of the reaction. AB - Sonochemical polymerization of benzene and halogen-substituted benzenes has been studied. The difference of absorption spectra of polymerization products can be explained qualitatively using bond energies of the primary products. The relative rate constant of the polymerization reaction is apparently proportional to the inverse of the vapour pressure of the liquids. Using this relation, we analysed the relative rate constant of the polymerization in benzene/chrolobenzene mixtures. From this, we conclude that sonochemical polymerization proceeds in the vapour phase of a bubble. PMID- 11270340 TI - Sonochemical reduction of carbon dioxide. AB - Sonolysis of carbon dioxide dissolved in water was performed from a standpoint of reducing this material in atmosphere. During one hour of sonication, the amount of CO2 decreased to about half at 5 degrees C under CO2-Ar atmosphere. The decreasing rate for CO2 followed the order Ar > He > H2 > N2 and it was down with increasing temperature in the range of 5-45 degrees C. The most favorable concentration for reducing CO2 was 0.03 (mole fraction of CO2 in gas phase). This concentration in gas phase means an equal mixture of CO2 and Ar in water, because CO2 is more soluble than Ar. Since carbon dioxide dissolved in water would be partly ionized, the roles of ions on the sonolysis were also examined. Gaseous reaction products were CO, H2 and a small amount of O2. Carbon monoxide and hydrogen might be obtained from CO2 and H2O by sonolysis, respectively. Both gases are fuel and react each other to C1 compounds such as methanol, and so on. Therefore, irradiation of ultrasonic waves should be an important technique for reducing CO2. PMID- 11270341 TI - Monolithic diphasic gels of mullite by sol-gel process under ultrasound stimulation. AB - Diphasic gel in the mullite composition was prepared from a colloidal sol of boehmite mixed with a hydrolyzed tetraethoxisilane (TEOS) solution. The boehmite sol was obtained by peptization of a poorly crystallized or very small mean crystallite size (approximately 34 A) precipitate, resulting from the reaction between solutions of aluminum sulfate and sodium hydroxide. Ultrasound was utilized in the processes of the TEOS hydrolysis and the boehmite peptization, and also for complete homogenization of the mixture to gel. The wet gel is almost clear and monolithic. The gel transparency is lost on drying, when syneresis has ended, so that the interlinked pore structure starts to empty and is recovered upon water re-absorption. Cracking closely accompanies this critical drying process. Differential thermal analysis (DTA) and X-ray diffraction (XRD) show that the solid structure of the gel is composed of an amorphous silica phase, as a matrix, and a colloidal sized crystalline phase of boehmite. Upon heat treatment, the boehmite phase within the gel closely follows the same transition sequence as in pure alumina shifted towards higher temperatures. Orthorhombic mullite formation was detected at 1300 degrees C. PMID- 11270342 TI - Ultrasound-promoted preparation of disteryl ethers catalyzed by montmorillonite K 10. AB - In the presence of montmorillonite K 10, 5(6)-unsaturated sterols (1) were heated under ultrasound at 45 degrees C in dichloromethane for 2-5.5 h to provide 3 beta,3'beta-5(6)/5'(6')-unsaturated disteryl ethers (2) in 35-79% yield. Meanwhile, 3 alpha,3'alpha-diandrost-5-en-17-one-3-yl ether (3a) and 3 alpha,3'alpha-dipregn-5-en-20-one-3-yl ether (3b) were also obtained as by products from 3 beta-hydroxyl-androst-5-en-17-one (1a) and 3 beta-hydroxyl pregnan-5-en-20-one (1b) respectively. PMID- 11270343 TI - Protonation of norharmane as a sonochemical dosimeter for organic media. The effect of temperature. AB - The sonolysis of different organic and aqueous media in the presence of norharmane produce its protonation. This simple and reversible reaction is particularly suitable as a dosimetric reaction in order to measure the relative amount of cavitation induced by the ultrasonic irradiation in non aqueous solutions. The protonation rate increases when small amounts of chloroform are added to the solution. The frequency (20 and 475 kHz) and temperature effects on the reaction rate are also studied. Our results show that sonication of aqueous solutions at high frequency leads to a strongly oxidant medium. PMID- 11270344 TI - Effect of liquid-phase properties on ultrasound intensity and cavitational activity. AB - The intensity of ultrasound is attenuated due to various properties of the liquid, such as viscosity, density, etc. In this paper, a simple method is proposed to measure the combined attenuation and cavitational activity of ultrasound intensity in various organic liquids using standard KI decomposition reaction. A modified experimental attenuation coefficient is proposed and its dependence on liquid viscosity reasonably matches the theoretical predictions made by Stokes [G.G. Stokes, Trans. Camb. Philos. Soc. 8 (1849) 287]. Exploratory work to determine the effect of other liquid properties on cavitational activity is carried out. Correlations are proposed to explain the dependence of the attenuated cavitational activity on various properties of a liquid. PMID- 11270345 TI - Comparison of ultrasound effects in different reactors at 20 kHz. AB - To compare the performances of three power ultrasonic devices at 20 kHz: a horn, a cup horn and a tube, the local intensity distributions of local effects of cavitation have been investigated. The sensor is an electrochemical probe, measuring the solid-liquid mass transfer rate, related to the cavitation intensity. The axial and radial profiles of mass transfer coefficients have been investigated in three devices, at various power inputs. In all these equipments very strong heterogeneities have been characterized, whether a standing wave appears or not. PMID- 11270346 TI - Sonochemistry of organic compounds in homogeneous aqueous oxidising systems. AB - The influence of ultrasound on the degradation of trichloroethylene, o chlorophenol and 1,3-dichloro-2-propanol in some Fenton type aqueous systems was investigated. Kinetic analysis of the results revealed that the ultrasonic waves do not really enhance the reactivity of the system, but rather add their own degradation mechanism to the chemical degradation. This effect is comparable to the well known sonochemical switching effect. As a consequence, the degradation rate of the combined systems is the sum of the pure sonochemical degradation rate and the silent chemical degradation rate. The only exception to this rule is the degradation of o-chlorophenol in the presence of hydrogen peroxide, in the absence of catalysts. Care had to be taken in order to avoid artifacts that may arise, e.g., when neglecting temperature effects. PMID- 11270347 TI - Dielectric polarization in the Planck theory of sonoluminescence. AB - Sonoluminescence observed in the cavitation of liquid H2O may be explained by the Planck theory of SL, which treats the bubbles as collapsing miniature masers having optical waves standing in resonance with the dimensions of the bubble cavity. Microwaves are shown to be created from the Planck energy of the standing waves, provided the bubble wall can be treated as a perfect blackbody surface. Liquid H2O is strongly absorbent in the ultraviolet and there the bubble approaches a Planck blackbody enclosure. The microwaves are created at frequencies proportional to the bubble collapse velocity only to be promptly absorbed by the rotation quantum states of the H2O and other bubble wall molecules. The microwaves are absorbed discretely at rotation line frequencies, or continuously by dipole rotation at frequencies from 1 to 30 GHz. In the liquid state, molecular rotation of the H2O molecule is hindered and the microwave energy is rapidly turned into bending energy by intermolecular collisions. Subsequently, the bubble wall molecules may thereby ionize and produce visible photons. The microwaves create intense electrical fields in the bubble wall by dielectric polarization. If the gases adjacent to the bubble wall undergo electrical breakdown, free electrons are created, thereby providing sonoluminescence with a magnetic field effect. PMID- 11270348 TI - [Controversy: systemic amniocentesis for women 38 years and more? Is it not premature to no longer accept performing amniocentesis on a patient age 38 years or more?]. PMID- 11270349 TI - Role of neurotransmitter autoantibodies in the pathogenesis of chagasic peripheral dysautonomia. AB - Chagas' disease is caused by a parasite, Trypanosoma cruzi, which is widely distributed in South and Central America. Dysautonomias, derangements of sympathetic and parasympathetic nervous system function, are seen fairly often during the chronic course of Chagas' disease. Many infected subjects developed, in the course of the disease, neurogenic cardiomyopathy or digestive damage. Our investigations show the existence of circulating antibodies in Chagas' disease that bind to beta-adrenergic and muscarinic cholinergic receptor (mAChR). The neurotransmitter receptor-autoantibody interaction triggers in the cells intracellular signal transductions that alter the physiological behavior of the target organs, leading to tissue damage. Moreover, the deposit of autoantibodies behaving as agonists induces desensitization and/or down regulation of the receptors. This in turn can lead to a progressive blockade of them with sympathetic and parasympathetic denervation. Using synthetic peptides for immunoblotting and enzyme immunoassay, we demonstrated that these autoantibodies reacted against the second extracellular loop of the human heart beta 1 adrenoceptor and M2 cholinoceptor. Also, the corresponding affinity-purified antipeptide antibodies displayed an agonist-like activity associated with specific receptor activation. A strong association between circulating antipeptide M2 mAChR autoantibodies and the presence of patients' low heart rate variability index, bradycardia and cardiac or esophageal autonomic dysfunction in chronic chagasic patients was verified. This fact make these antipeptide antibodies a proper marker of cardiac neuromyopathy and achalasia. PMID- 11270351 TI - 1999 Novera Herbert Spector Award for significant contributions in both leadership and research in neuroimmunomodulation. Recipient: George P. Chrousos, M.D., Sc.D. PMID- 11270350 TI - Social disruption, immunity, and susceptibility to viral infection. Role of glucocorticoid insensitivity and NGF. AB - Glucocorticoid (cort) responses have been shown to suppress inflammatory reactions by inhibiting the trafficking of immune cells. Recently, it was demonstrated that restraint stress (RST) and psychosocial stress (social reorganization; SRO) differentially affected the pathophysiology and survival in the mouse influenza viral infection model. While both stressors activated the HPA axis, only SRO affected survival. In RST, elevated cort diminished recruitment of inflammatory cells following intranasal challenge of C57BL/6 mice with A/PR8 virus. However, infected SRO mice developed hypercellularity in the lungs and were more likely to die from lung consolidation than controls. Since elevated cort failed to be anti-inflammatory in SRO mice, the hypothesis that psychosocial stress induced steroid insensitivity was tested. An in vitro cort suppression test was performed by stimulating splenocytes from SRO and control mice with mitogen in the presence or absence of cort. Proliferation of ConA-stimulated cells was inhibited by cort in a dose-dependent fashion in controls, but splenocytes from SRO mice stimulated with ConA were resistant to cort-induced suppression. Thus, psychosocial stress induced a state of steroid insensitivity. SRO also induced the release of nerve growth factor (NGF) from the salivary glands into circulation; plasma NGF correlated with development of steroid insensitivity. NGF has been reported to negatively regulate the expression of type II glucocorticoid receptors, and thus may be a key factor in the induction of steroid insensitivity. PMID- 11270352 TI - Low temperatures in the study of chemical reactions and molecular structures. Proceedings of the 3rd International Conference on Low Temperature Chemistry. Nagoya, Japan, 26-30 July 1999. PMID- 11270353 TI - [Physicians caught in the controversies. Proceedings of the 24th Symposium for Lawyers and Physicians. Berlin, Germany 18-19 February 2000]. PMID- 11270354 TI - [2nd Congress of the European Society for Agricultural and Food Ethics (EURSAFE)]. PMID- 11270355 TI - Re: Bacteriology of burn wounds in Enugu, Nigeria. PMID- 11270357 TI - Recent references. PMID- 11270356 TI - Skin substitutes in burns. PMID- 11270358 TI - Lp(a) concentrations in hyperapobetalipoproteinemia. PMID- 11270359 TI - Sensory, digestive and metabolic influences on preference and intake. AB - In the 1960s. scientific work on the mechanisms of appetite began to shift from the effects of deprivation on reinforcement, motivation and sensation (hunger and thirst) to the influences of ingestion itself on what and how much was chosen at a bout of eating and drinking (the sating of appetite). This personal overview starts with John D. Davis's involvement in this trend to studying sensory, digestive and metabolic controls of meal volume, as he and the author each joined the field. The main topic is the diverse series of innovative experiments that Davis carried out in the author's research group in 1970 1971. This work exemplified Davis's technical inventiveness and theoretical precision. It generated influential findings on behavioural roles of dietary flavours, gastric distension and hepatic metabolism. PMID- 11270360 TI - Microstructure of eating behavior in humans. PMID- 11270361 TI - Trichlorobacter thiogenes should be renamed as a Geobacter species. PMID- 11270362 TI - Mycobacterial 19-kDa lipoprotein mediates Mycobacterium tuberculosis-induced apoptosis in monocytes/macrophages at early stages of infection. PMID- 11270363 TI - The PYRIN domain: a novel motif found in apoptosis and inflammation proteins. PMID- 11270364 TI - Sequence as well as functional similarity for DIABLO/Smac and Grim, Reaper and Hid? PMID- 11270366 TI - Characterization and Diagnosis of Prion Diseases. Proceedings of a symposium. September 23-25, 1999, Tubingen, Germany. PMID- 11270365 TI - Symposium on endocrinology and aging: the 1st meeting of the European Interest Group hormones and aging. PMID- 11270367 TI - Heavy Metals in the Environment. Proceedings of the II National Conference. Sassari, Italy, 4-5 May 2000. PMID- 11270369 TI - Proceedings of the Northwestern University Medical School 2nd Annual Oncology Consensus Conference: Recent Advances and New Therapeutic Directions for Hematologic Malignancies. January 12-16, 2000. Kailua-Kona, Hawaii. PMID- 11270368 TI - Antidepressant augmentation with low-dose olanzapine in obsessive-compulsive disorder. PMID- 11270370 TI - Quinolone resistance-determining regions of gyrA and parC in Pasteurella multocida strains with different levels of nalidixic acid resistance. PMID- 11270372 TI - [Variola in Ostrava]. PMID- 11270373 TI - Proceedings of the Women in Thoracic Surgery Symposium. Fort Lauderdale, Florida, USA. January 31, 2000. PMID- 11270371 TI - Identification of a Tn1546-like (type 2) element in vancomycin-resistant Enterococcus faecium isolated from hospitalized patients in Japan. PMID- 11270374 TI - Successful Aging Through Oral Health. Proceedings of a conference. March 14-16, 1999. Vancouver Island, British Columbia, Canada. PMID- 11270375 TI - Sleep deprivation and host defense. PMID- 11270376 TI - A job for life? PMID- 11270377 TI - A new partnership. PMID- 11270378 TI - Piercing difficulties. PMID- 11270379 TI - Root resorption. PMID- 11270380 TI - Antibiotic prescribing for acute sinusitis. PMID- 11270381 TI - Highspeed not a winning tool. PMID- 11270382 TI - Dental strategy. PMID- 11270383 TI - A report of an elective to the Hospital for Sick Children and Bloorview Macmillan Centre, Toronto. AB - The Department of Dentistry based at the Hospital for Sick Children and Bloorview MacMillan Centre in Toronto is unique. It provides treatment for cleft lip and palate patients and all forms of specialist dental treatment to children and adolescents resident in Ontario. The quality of service provided by the department has been externally recognised and validated through an ISO 9002 award (February 1999). At that time, it was the first Dental Clinic in North America to receive the ISO 9002 standard. In addition, the American Cleft Palate Craniofacial Association has recognised the department for its organisation and contributions in the field of cleft lip and palate and craniofacial surgery. PMID- 11270384 TI - Dental attendance in 1998 and implications for the future. AB - The 1998 survey of Adult Dental Health in the UK was carried out under the auspices of the Office of National Statistics together with the Universities of Birmingham, Dundee, Newcastle-upon-Tyne and Wales. A key behavioural indicator in these decennial surveys is whether people say they go to a dentist for a regular dental check-up, an occasional dental check-up or only when they have trouble with their teeth. The proportion of dentate adults in the UK who report attending for regular dental check-ups has risen from 43% in 1978 to 59% in 1998. Older adults (over 55 years old) in 1998 were the most likely to say they attend for regular dental check-ups. Many younger adults (16-24) in 1998 said they went to a dentist less often than 5 years previously, they were also the least likely to say they attend for regular dental check-ups. Dental anxiety remains a problem for many dental patients but another factor of importance to many is their want to be involved in the treatment process and especially to be given an estimate of treatment costs. PMID- 11270385 TI - Connectors. AB - This article describes the types and functions of connectors for RPDS. It also considers the relative merits and limitations of these connectors. PMID- 11270387 TI - Capitation registration and social deprivation in England. An inverse 'dental' care law? AB - OBJECTIVE: To examine associations between NHS child dental registration data and area deprivation scores of English Health Authorities (N= 100) in 1996/97 and 1997/98. METHOD: The Department of the Environment index of local conditions and the Jarman Underpriviledge Area Score from the 1991 census were used to measure deprivation. Prior to September 1997, children got free dental treatment under a capitation scheme with an NHS dentist. If they did not attend within 24 months their registration lapsed on the last day of December of the second registration year and they were deleted from the capitation list. After September 1997 the registration period was reduced to 15 months. OUTCOME: Curve-linear regression of the Health Authority (HA) percentage of children registered, lapses in capitation registrations and deprivation scores. RESULTS: In England 68% of children were registered in December 1996. The percentage registered in each Health Authority was associated with deprivation (DoE, r2=0.33, Jarman, r2=0.27 p<0.01). In January 1997, 17.8% (1,345,142) of children registered lapsed (HA range 12.8% to 30.3%) and this was also significantly associated with deprivation (DoE r2=0.66, Jarman, r2=0.51 p<0.01). Similar results were found in 1997/98. CONCLUSIONS: Registration and lapse rates were significantly associated with social deprivation confirming that there is an inverse 'dental' care law for children in England. NHS capitation may widen dental health inequalities. PMID- 11270386 TI - Comparison of decisions regarding prophylactic removal of mandibular third molars in Sweden and Wales. AB - OBJECTIVE: To test the hypothesis that Swedish dentists schedule more mandibular third molars for prophylactic removal compared with UK dentists and oral surgeons. DESIGN: Clinical and radiographic information relating to a stratified sample of 36 disease-free mandibular third molars (equal distribution of males and females, patients' age, angular position and degree of impaction) was presented to 26 general dental practitioners (GDPs) and 10 oral surgeons in Sweden and 18 GDPs and 10 oral surgeons in Wales who were asked to decide whether or not the third molars should be removed. RESULTS: There was no evidence of any difference in mean number of molars scheduled for removal by the GDPs, but the Swedish oral surgeons scheduled significantly more third molars for removal than oral surgeons in Wales. CONCLUSION: The less interventionist approach among oral surgeons in the UK may reflect the development and application of authoritative guidelines in the UK and an extensive debate concerning appropriateness of prophylactic removal there. PMID- 11270388 TI - A national survey of dental hygienists: working patterns and job satisfaction. AB - OBJECTIVE: To describe the working practices and level of job satisfaction of dental hygienists in the United Kingdom. DESIGN: Postal questionnaire survey of 3,955 dental hygienists registered with the General Dental Council. Replies were received from 2,533 (64%). RESULTS: At the time of the survey only a small proportion of respondents (11%) were not working as dental hygienists, the most common reason for a current career break being child rearing. The majority of dental hygienists (78%) were employed in general dental practices, and most worked in more than one practice (64%). Approximately half worked part-time (fewer than 30 hours per week), and part-time working was more common amongst those respondents with childcare responsibilities. In the region of 60% of respondents had taken one or more career breaks during their working life, and the average total duration of career breaks was 11 months, the most common reason for all career breaks was child rearing. Additional qualifications had been gained by 35% of the sample, a high proportion (75%) had attended training courses in the previous year. The respondents expressed a high degree of job satisfaction, those who were older and who had childcare responsibilities expressed higher levels of job satisfaction. CONCLUSIONS: Dental hygienists express a high level of job satisfaction. A proportion take breaks in their career, most commonly for pregnancy and child rearing. The majority return to part-time employment after their career break. Planning of future requirements for the training of professionals complementary to dentistry should be informed by a consideration of the working patterns of dental hygienists. PMID- 11270389 TI - Long term periodontal problems--the chemotherapeutic aspect. PMID- 11270390 TI - Angiotensin II-induced hypertension in bradykinin B2 receptor knockout mice. AB - The present study was performed to examine the role of endogenous bradykinin (BK) in the development of angiotensin II (Ang II)-induced hypertension in mice. BK B2receptor knockout (B2R-/-) and wild-type (B2R+/+) mice (22to 26 g) were infused with either saline (SAL) or Ang II (40ng/min) via an osmotic minipump implanted intraperitoneally. On day 12after implantation, there was no difference in systolic blood pressure (SBP, tail-cuff plethysmography) between SAL/B2R+/+ and SAL/B2R-/- mice(128+/-5 versus 133+/-6 mm Hg, n=24/group). In contrast, SBP was higher on day 12 of infusion in Ang II/B2R-/- than in Ang II/B2R+/+ mice (173+/ 6versus 156+/-5 mm Hg; P<0.05, n=27 and 28). Mean arterial pressure (MAP)was also higher in anesthetized Ang II/B2R-/- mice than in Ang II/B2R+/+mice (139+/-3 versus 124+/-3 mm Hg; P<0.05, n=16 and 14). Unlike Ang II, long-term norepinephrine (NE) infusion via an osmotic minipump (45ng/min) caused equivalent increases in SBP in B2R+/+ and B2R-/- mice measured on day 12 after implantation (151+/-4 versus 149+/-5 mm Hg, n=9and 8). MAP also did not differ on day 13 after implantation between NE/B2R+/+ and NE/B2R-/- mice (120+/-6 versus 122+/-4 mm Hg, n=9 and 8). There were no differences in glomerular filtration rate and urinary sodium excretion among the groups. However, renal plasma flow (RPF) was lower in Ang II/B2R-/- mice than in Ang II/B2R+/+ mice (2.34+/-0.06 versus 4.33+/-0.19 mL x min-1 x g-1; P<0.05). Acute inhibition of NO synthase (NOS)with nitro-L arginine-methyl ester (0.5 microg x g-1 x min-1) in SAL/B2+/+ and SAL/B2-/- mice caused equal increases in MAP (142+/-1 versus 145+/-1 mmHg) and decreases in RPF (2.06+/-0.06 versus 2.12+/-0.15 mL x min-1 x g-1).However, short-term NOS inhibition caused a greater increase in MAP of Ang II/B2R+/+ mice than of Ang II/B2R-/- mice, such that MAP after NOS inhibition in Ang II/B2R+/+ approached that of Ang II/B2R-/- mice (156+/-2versus 159+/-2 mm Hg). These changes were associated with a decrease in RPF in Ang II/B2R+/+ mice to values similar to those of Ang II/B2R-/- mice before NOS inhibition (2.12+/-0.09 versus 2.34+/-0.06 mL x min-1 x g-1). These results demonstrate that the kallikrein-kinin system selectively buffers the vasoconstrictor activity of Ang II. Furthermore, the enhanced susceptibility of B2R-/- mice to Ang II-induced hypertension and renal vasoconstriction is likely due to an impaired ability to release NO by endogenous kinins. PMID- 11270391 TI - 80 Nobel laureates urge support for embryonic stem cell research. PMID- 11270392 TI - Three groups identify gene for DiGeorge syndrome. PMID- 11270393 TI - Jump in sudden cardiac deaths reported in younger people during past decade. PMID- 11270395 TI - Development and use of complex probes for DNA fingerprinting the infectious fungi. AB - A variety of methods have emerged for genetic fingerprinting the infectious fungi. One of the most versatile is Southern blot hybridization with species specific complex DNA probes that include sequences that identify hypervariable, moderately variable and invariant genomic sequences. These probes assess genetic relatedness at all the necessary levels including identical, highly related but non-identical, moderately related and unrelated. Methods are described for cloning complex probes, characterizing them and verifying their effectiveness at the different levels of resolution. The complex probes that have been developed for Candida albicans, C. glabrata, C. dubliniensis, C. tropicalis, C. parapsilosis and Aspergillus fumigatus are described and discussed. PMID- 11270394 TI - Coccidioides immitis isolated from armadillos (Dasypus novemcinctus) in the state of Piaui, northeast Brazil. AB - Natural infection of armadillos with Coccidioides immitis was studied in the state of Piaui, northeast of Brazil, endemic for coccidioidomycosis. In 1998, 26 nine-banded armadillos (Dasypus novemcinctus) were captured in 4 different counties. The animals were sacrificed under deep anesthesia with ether. At necropsy fragments of spleen, liver, lungs and heart were homogenized and seeded onto Sabouraud dextrose agar with and without cycloheximide (BBL, USA). Part of each organ was also processed for histological examination. Suspected colonies of filamentous fungi observed after the second week of incubation at room temperature, exhibiting barrel-shaped arthroconidia alternating with empty spaces, were inoculated intraperitoneally into mice. Three armadillos proved to be infected with C. immitis. Mice inoculated with suspected colonies obtained from homogenized spleen of three and liver of two armadillos developed disseminated coccidioidomycosis and immature and mature spherules of C. immitis were disclosed in several organs. For the first time armadillos (D. novemcinctus) were found naturally infected with C. immitis, adding new data on the ecology and on a possible role of these ancestral mammals in the evolutionary life cycle of this fungus. PMID- 11270396 TI - The use of flow cytometry as a tool for monitoring filament formation of fungi. AB - Flow cytometry (FC) has the ability to discriminate a variety of cell parameters including cell size and complexity, and fluorescence intensity. As yeast cells or fungal spores germinate they undergo a morphological transformation from round oval shaped cells to elongate filamentous forms. To date, monitoring these events has been performed using microscopic examination. Microscopic examination is a labor intensive process that examines a very small percentage of the total cell population. We have developed a method using FC that is rapid, simple to perform, and reproducible. The major advantages of FC include analysis of a larger number of cells, increased objectivity due to nonselective measurements of all cells in the population studied, and the computer related data analysis capability of the flow cytometer. PMID- 11270397 TI - Identification and expression of multidrug resistance-related ABC transporter genes in Candida krusei. AB - Infections with Candida krusei have increased in recent years as a consequence of its intrinsic resistance to fluconazole, an antifungal azole widely used in immunocompromised individuals to suppress infections due to azole-susceptible C. albicans. One established mechanism for azole resistance is drug efflux by ATP binding cassette (ABC) transporters. Since these transporters recognize structurally diverse drugs, their overexpression can lead to multidrug resistance (MDR). To identify C. krusei genes potentially involved in azole resistance, PCR was performed with primers corresponding to conserved sequences of MDR-related ABC transporters from other fungi. Two genes, ABC1 and ABC2, were identified; Southern blots suggested that both have one or two related gene copies in the C. krusei genome. ABC1 RNA was constitutively expressed at low levels in log phase cells while ABC2 RNA was undetectable. However, both genes were upregulated as cultures approached stationary phase, and this upregulation was correlated with decreased susceptibility to the lethal activity of the azole derivative miconazole. Furthermore, ABC1 was upregulated following brief treatment of C. krusei with miconazole and clotrimazole (but not other azoles), and the unrelated compounds albendazole and cycloheximide. The latter two compounds antagonized fluconazole activity versus C. krusei, supporting a role for the ABC1 transporter in azole efflux. Finally, miconazole-resistant mutants selected in vitro demonstrated increased constitutive expression of ABC1. Based on these expression data, genetic and functional characterization of the ABC1 transporter to directly test its role in C. krusei azole resistance would appear to be warranted. PMID- 11270398 TI - Genotypic identification of Candida dubliniensis isolated from HIV patients by MLEE. AB - Candida dubliniensis is a novel species only recently described. This emerging pathogen shares some of the phenotypic characteristics specific to C. albicans but is genetically different. In this study we typed four strains of atypical C. albicans isolated in our laboratory and compared them to 41 strains of C. albicans and 11 strains of C. dubliniensis by several phenotypic methods and by multilocus enzyme electrophoresis. Using factorial correspondence analysis, we distinguished C. dubliniensis and the atypical C. albicans strains from all strains of C. albicans. Atypical C. albicans strains were identified as C. dubliniensis. PMID- 11270399 TI - Adherence of clinical isolates of Saccharomyces cerevisiae to buccal epithelial cells. AB - A number of isolates of Saccharomyces cerevisiae have been associated with disease in immunocompromised individuals. Such isolates display a variety of characteristics that enable colonization and persistence in the host. The aim of the work presented here was to establish whether clinical isolates of S. cerevisiae were capable of adhering to epithelial tissue. Adherence to host tissue has been shown to be crucial to the virulence of the pathogenic yeast Candida albicans, and identification of this ability in S. cerevisiae might indicate a role for adherence in tissue colonization by this emerging pathogen. Clinical S. cerevisiae isolates were found to be capable of adhering to exfoliated buccal epithelial cells (BECs) but to a lesser degree than C. albicans. In contrast to the situation evident with C. albicans, the adherence of S. cerevisiae isolates to BECs was not influenced by the carbon source in which the yeast was grown. Treatment of S. cerevisiae with trypsin or proteinase K resulted in a significant reduction in adherence ability while adherence was unaffected by treatment of cells with mannosidase, thus indicating a possible role for proteins rather than mannoproteins in the adherence of S. cerevisiae to BECs. PMID- 11270400 TI - Effect of sub-MICs of antimycotics on expression of intracellular esterase of Trichophyton rubrum. AB - The electrophoretic pattern of the intracellular esterase of the dermatophyte Trichophyton rubrum was altered when this fungus was grown in the presence of subinhibitory concentrations of the antimycotics tioconazole or griseofulvin. All strains (original isolate and antimycotic resistant mutants) presented five clearly visible bands when cultivated on medium containing below-minimum inhibitory concentrations (sub-MICs) of tioconazole or griseofulvin, and only two clearly visible bands when cultivated in medium without antimycotics. No extra bands were detected in the electrophoretic patterns of the extracellular esterase of these fungi (mutants or the original isolate) when cultivated with or without tioconazole or griseofulvin (sub-MIC values). These results suggest that additional forms of esterase are produced inside the cell and may be a nonspecific response to cellular stress, or may participate in cellular detoxification processes in the presence of these antimycotics. PMID- 11270401 TI - Phaeohyphomycosis due to Cladosporium cladosporioides. AB - Phaeohyphomycosis is a clinical entity caused by dematiaceous fungi. We describe a clinical case of phaeohyphomycosis due to Cladosporium cladosporioides in a 45 year-old white male, apparently healthy, human immunodeficiency virus-negative. The patient was treated with terbinafine for 9 months, with regression of a skin lesion. Three months after discontinuation of the therapy, there was a clinical and mycological relapse. After progression of the disease with inadequate treatment, there was no response to amphotericin B and flucytosine. Finally, we obtained a clinical response with itraconazole oral solution at 600 mg day(-1) for a 6-month period. PMID- 11270402 TI - Rhinosporidiosis in a domestic cat. AB - Rhinosporidiosis was diagnosed in a domestic shorthair cat from a suburb of Washington DC, USA. The clinical presentation of protracted sneezing and epistaxis was associated with a polypoid lesion in the right nostril. Light microscopic examination revealed a polypoid lesion with numerous sporangia containing maturing endospores. Free endospores were present in the stroma of the polyp and lumen of the nasal cavity. Transmission electron microscopy revealed ultrastructural features typical of Rhinosporidium seeberi. The case was followed clinically for a total of 70 months and there were five attempts at surgical excision. This is the first reported case of rhinosporidiosis in a domestic cat. PMID- 11270403 TI - Keratomycosis caused by Cylindrocarpon lichenicola. AB - We present a case of keratomycosis caused by Cylindrocarpon lichenicola in a 30 year-old immunocompetent male patient living in a rural area of Formosa Province (north-eastern Argentina). No ocular trauma is reported in his case history. There are no previous reports of infections caused by this fungus in Argentina. PMID- 11270404 TI - Isolation of Sporothrix schenckii from the nails of domestic cats (Felis catus). AB - We report the first isolation of Sporothrix schenckii from the nail surfaces of cats. The fungus grew from nail clippings of three cats associated with three household outbreaks of sporotrichosis involving cats and human beings. The identification of the isolates was based on macroscopic and microscopic morphological characteristics at 25 degrees C and conversion of S. schenckii to the yeast-like form at 37 degrees C. PMID- 11270405 TI - A new dominant selectable marker for use in Cryptococcus neoformans. AB - Cryptococcus neoformans is an excellent model system for studies on the molecular pathogenesis of fungal infections. There is only one dominant selectable market that can be used in the transformation of this organism, and we wanted to develop another. We found that various strains of C. neoformans are very sensitive to the aminoglycoside antibiotic nourseothricin, and that spontaneous resistance to this drug must be an extremely rare event. Resistance to nourseothricin is conferred by the product of the nourseothricin acetyltransferase gene (nat1) from Streptomyces noursei. In order to express this gene in C. neoformans, we created a fusion construct by driving expression of natl with the promoter sequence from a C. neoformans actin gene. Biolistic transformation of the serotype A C. neoformans strain H99 and the serotype D strain JEC21 with this construct resulted in transformation efficiencies of approximately 1,000 transformants microg(-1) of DNA and 20 transformants microg(-1) of DNA, respectively. Southern blots were performed using DNA from some of the H99 transformants, and this confirmed that all of the resistant isolates had the construct integrated in a random fashion within the genome. There was no cross-resistance of the nourseothricin-resistant transformants to hygromycin B, which is the other antibiotic used as a dominant selection marker in C. neoformans. The development of nourseothricin resistance as a second dominant selectable market will be helpful in future molecular studies on this important pathogenic fungus. PMID- 11270406 TI - Reliability of clinical research on invasive fungal infections: a systematic review of the literature. AB - The aim of this study was to assess the comparability and reliability of clinical research on invasive fungal infections (IFIs) which is accumulating in the medical literature. A Medline search strategy was developed covering the years 1985-1997. The 7,086 articles identified this way were further limited to 173 by reading the abstracts to studies involving immunocompromised patients and deep tissue infections. Diagnostic criteria used for definitions of IFIs were evaluated according to the level of confidence in diagnosis. For analysing the reliability, we used the sample of 397 patients from the European Organization for Research and Treatment of Cancer (EORTC) database. Each patient in the database was evaluated according to the definitions employed in each article. The level of overall agreement among the articles was very low (Kappa = 0.253). The results of this analysis show the discrepancy in the medical literature in diagnosing IFIs and the necessity for the standardization of definitions. PMID- 11270407 TI - Role of chemokines in fungal infections. AB - The production of chemokines at the site of a fungal infection is critical for effective recruitment of leukocytes to that site. Over 40 chemokines and 20 chemokine receptors have been identified. The most intriguing biological property of chemokines is that they often play non-redundant roles in vivo even though they are highly related, have multiple activities and bind multiple chemokine receptors. Almost all of the chemokine studies to date have concentrated on responses to Cryptococcus, Candida, Aspergillus or Pneumocystis. The role of chemokines in infections caused by fungi such as Histoplasma, Blastomyces, Coccidioides and Paracoccidioides remains to be explored. In this review we have summarized what is currently known about the role of chemokines during fungal infection, including the influence of these signaling proteins on effector cell recruitment and development of cell-mediated immunity. PMID- 11270408 TI - Cryptococcosis in India: the awakening of a giant? AB - The menace of cryptococcosis has assumed global proportions over the years. The tropical climate of the Indian subcontinent offers a suitable environment for Cryptococcus neoformans, and the onslaught of the acquired immune deficiency syndrome (AIDS) pandemic since the early 1990s has substantially influenced the situation. Coupled with that are the advances in laboratory diagnostic techniques that have made accurate diagnosis increasingly available. These factors together have led to a sharp increase in the number of reported cases of cryptococcosis. This review attempts to present an overview of the status of cryptococcosis in India from its first description to the most recent times. The disease has been reported from almost all parts of the country. C. neoformans var. neoformans is predominantly found in clinical samples, while C. n. var. gattii infection has also been reported. An organ commonly involved is the central nervous system, among others. Both immunocompromised and apparently immunocompetent patients have been affected. Laboratory diagnosis is mostly by conventional methods, while effective therapeutic options are limited. Early diagnosis followed by institution of specific therapy, where possible, has effectively reduced mortality. Awareness of the disease and maintenance of a high index of clinical suspicion is required. An integrated approach to patient management with active interaction between the clinicians and the laboratory personnel would be highly beneficial. The wide variety of presentations of the disease seen in India suggests the possibility of occurrence of strain variation which needs to be investigated fully. Introduction of routine testing of antifungal susceptibility of clinical isolates is also important in order to obtain baseline data on susceptibility patterns and to predict in advance any shift in those patterns in the population. To maintain a high standard in all such endeavours, the establishment of an external quality control system is desirable. PMID- 11270410 TI - Candidal and bacterial bloodstream infections in premature neonates: a case control study. AB - Nosocomial bloodstream infections (BSI) in premature neonates are an important cause of morbidity and mortality. The early and efficient diagnosis of a neonatal BSI and the differentiation between bacterial and fungal BSI remains a challenging task. We compared the clinical features and blood test results in preterm infants with proven candidal or bacterial BSI in order to identify potential risk factors for developing a candidal BSI. Preterm infants with proven candidal BSI were significantly more prematurely born (mean age of gestation 27.7 vs. 29.8 weeks), had previously received significantly more antibiotics of multiple classes (mean 4.4 vs. 1.2) for significantly longer periods (mean 19.3 vs. 3.2 days), were ventilated more intensively, had a significantly longer stay at the neonatal intensive care unit before the onset of the BSI (mean 26.5 vs. 9.4 days), and had C-reactive protein values even higher than in preterm infants with a bacterial BSI (mean 90 vs. 71 mg l(-1)). The presence of thrombocytopenia ( < 150 x 10(9) cells l(-1)) in all the preterm infants with candidal BSI was a significant difference. No differences were seen with regard to birth-weight, use of central intravascular catheters, total parenteral nutrition, white blood cell count and differentiation. In conclusion, candidal BSI can be strongly expected after the third week of admittance in the most premature neonates on a respirator and treated with multiple classes of antibiotics for a prolonged period of time. The presence of these risk factors in a 'septic' premature infant on antibiotic treatment justifies the empiric use of antifungals. PMID- 11270409 TI - COS-l, a putative two-component histidine kinase of Candida albicans, is an in vivo virulence factor. AB - The human fungal pathogen, Candida albicans, has three putative histidine kinases showing homology to those of plants, bacteria and other fungi. We have constructed a homozygous deletion strain and a hemizygous reconstituted strain of one of these histidine-kinase-encoding genes, COS-1, in C. albicans. Neither strain showed any growth defect in a number of liquid media nor increased resistance or sensitivity to a number of antifungal drugs. Importantly, we show that the COS-1 homozygous disruption strain had significantly reduced virulence in a systemic murine model of candidosis. Thus, COS-1 appears to be an in vivo virulence factor and may represent a novel target for the development of antifungal drugs. PMID- 11270411 TI - Isolation and characterization of the Candida albicans MOT2 gene. AB - A putative Candida albicans homologue of Saccharomyces cerevisiae MOT2 (modulator of transcription) has been cloned and analyzed. A cDNA fragment corresponding to a portion of S. cerevisiae MOT2 was used to isolate a similar C. albicans gene (CaMOT2). CaMOT2 is comprised of two exons of 50 bp and 1,714 bp, respectively, with a single 82 bp intron located near the 5' end of the gene. The gene encodes a protein (CaMot2p) with an estimated mass of 67 kDa. The 5' region of the gene shows sequence homology with S. cerevisiae MOT2, whereas no significant similarity was observed in the 3' region. Similarly, the N-terminal portion of C. albicans Mot2p exhibits approximately 80% homology with S. cerevisiae Mot2p, while no significant homology to any known protein was observed in the carboxy terminal half of the C. albicans protein. The N-terminal portion of CaMot2p contains a cysteine-rich domain (amino acids 18-62). The distribution of the cysteine residues identifies CaMot2p as a zinc-finger protein. The data suggest two potential Zn-binding sites, similar to the arrangement found in S. cerevisiae. Reverse-transcriptase polymerase chain reaction was used to compare the level of CaMOT2 expression between C. albicans grown in vitro and growth during in vivo infection in the rat model of oral candidiasis. The results showed CaMOT2 is down-regulated during growth in the rat oral cavity compared to in vitro culture. Although the function of C. albicans MOT2 has not been determined, comparison to S. cerevisiae MOT2 suggests the gene product may act as a general negative regulator. PMID- 11270412 TI - Interferon-gamma production in peripheral lymphocytes of patients with tinea pedis: comparison of patients with and without tinea unguium. AB - The precise mechanism of the host defense that protects the nail from dermatophyte invasion is not known. Recent immunological findings in dermatophytosis suggest the hypothesis that the T helper 1 (Th1) response may play a role in protecting the nail from dermatophyte invasion. Our present study focused on interferon-gamma (IFN-gamma) release in patients with tinea pedis with or without tinea unguium, and pathogenesis of tinea unguium is discussed in relation to the association with a possible deficiency of Th1 response in the host defense mechanism. The production of IFN-gamma by peripheral blood mononuclear cells from the patients with tinea unguium in response to stimulation with trichophytin was not impaired in contrast to that from the patients without tinea unguium. Comparable lymphocyte proliferation to trichophytin was observed in both groups. Normal healthy persons with no clinical evidence of tinea could be divided into two groups based on lymphocyte proliferation and IFN-gamma production in response to trichophytin: high responder and low responder, with high responders being correlated with a clinical history of previous tinea pedis. In this study, a lack of a Th1 response to dermatophyte antigen was not shown in patients with tinea unguium by measuring the release of IFN-gamma, which plays a role in the effector phase of the delayed-type hypersensitivity reaction. A deficiency in the Th1 response to dermatophyte antigen, therefore, does not appear to play an important role in the establishment of tinea unguium. PMID- 11270413 TI - Yeast identification in the clinical microbiology laboratory: phenotypical methods. AB - Emerging yeast pathogens are favoured by increasing numbers of immunocompromised patients and by certain current medical practices. These yeasts differ in their antifungal drug susceptibilities, and rapid species identification is imperative. A large variety of methods have been developed with the aim of facilitating rapid, accurate yeast identification. Significant recent commercial introductions have included species-specific direct enzymatic colour tests, differential chromogenic isolation plates, direct immunological tests, and enhanced manual and automated biochemical and enzymatic panels. Chromogenic isolation media demonstrate better detection rates of yeasts in mixed cultures than traditional media, and allow the direct identification of Candida albicans by means of colony colour. Comparative evaluation of rapid methods for C. albicans identification, including the germ tube test, shows that chromogenic media may be economically advantageous. Accurate tests for single species include the Bichrolatex Albicans and Krusei Color tests, both immunologically based, as well as the Remel Rapid Trehalose Assimilation Broth for C. glabrata. Among broad-spectrum tests, the RapID Yeast Plus system gives same-day identification of clinical yeasts, but performance depends on inoculum density and geographic isolate source. The API 20 C AUX system is considered a reference method, but newer systems such as Auxacolor and Fungichrom are as accurate and are more convenient. Among automated systems, the ID 32 C strip, the Vitek Yeast Biochemical Card and the Vitek 2 ID YST system correctly identify >93% of common yeasts, but the ID-YST is the most accurate with uncommon yeasts, including C. dubliniensis. Spectroscopic methods such as Fourier transformed-infrared spectroscopy offer potential advantages for the future. Overall, the advantages of rapid yeast identification methods include relative simplicity and low cost. For all rapid methods, meticulous, standardized multicenter comparisons are needed before tests are fully accepted. PMID- 11270414 TI - Synergistic interaction of terbinafine with triazoles or amphotericin B against Aspergillus species. AB - The in vitro activity of terbinafine alone and in combination with other antifungal agents was tested against isolates of Aspergillus fumigatus, A. flavus and A. niger. Testing was performed in a modified National Committee for Clinical Laboratory Standards (NCCLS) macrodilution broth assay, and interactions were examined using a checkerboard design. Terbinafine was highly active against Aspergillus isolates (minimum inhibitory concentration [MIC] 0.01 to 2 microg ml( 1)) with a primary fungicidal action (minimum fungicidal concentration [MFC] 0.02 to 4 microg ml(-1)). Amphotericin B was also highly active and cidal as expected (MIC 1 microg ml(-1), MFC 1 to 4 microg ml(-1)). The triazoles itraconazole and voriconazole were highly active but showed a variable degree of cidal activity against the different strains, voriconazole having the more potent cidal activity. Fluconazole had no significant activity (MIC > 128 microg ml(-1)). Drug combinations were tested in the A. fumigatus and A. niger strains. Terbinafine and amphotericin showed an additive to synergistic interaction depending on the isolate. Combinations of terbinafine with itraconazole or voriconazole displayed a potent synergistic interaction and fungicidal activity against all isolates. Surprisingly, fluconazole also potentiated the activity of terbinafine in an additive to synergistic fashion, despite its lack of activity alone. The results suggest potential clinical application of terbinafine in aspergillosis, either alone or in combination with amphotericin or triazoles. PMID- 11270415 TI - Ramichloridium mackenziei brain abscess: report of two cases and review of the literature. AB - We report two cases of brain abscesses caused by Ramichloridium mackenziei, a neurotropic dematiaceous fungus that seems to be geographically restricted to the Middle East. One of the patients had chronic myelomonocytic leukemia but did not receive any chemotherapeutic agents. The other patient was a normal host. Both cases had a fatal outcome despite aggressive antifungal therapy and surgical intervention. Herein, we review all previously described cases in the literature, and discuss the epidemiology, mycology and histopathology of this life threatening organism. PMID- 11270416 TI - HER-2/neu oncogene expression in advanced breast cancer. AB - Tumor tissue from patients with advanced breast cancer was analyzed for HER-2/neu and p53 expression. The tissue samples from primary tumor and from axillary lymph nodes or distant metastases from 118 breast cancer patients were obtained. Sections from formalin-fixed, paraffin-embedded materials were immunostained for HER-2/neu and p53 oncoprotein expression. Staining results were correlated with survival times and disease-free survival times, flow cytometric synthesis phase fractions, and DNA ploidy. No correlation could be found between HER-2/neu and p53 or any other tested factor, but grade I primary cancers that were positive for HER-2/neu showed a tendency for better outcome. The HER-2/neu staining of the metastases was independent of the staining of the primary tumor. HER-2/neu can be used as a prognostic marker for advanced breast cancer, when the primary tumor is well differentiated. PMID- 11270417 TI - Biodegradation of the antineoplastics vindesine, vincristine, and vinblastine and their toxicity against bacteria in the aquatic environment. AB - Antineoplastics are excreted into sewage, because patients often poorly metabolize them after administration or they are metabolized into more biologically reactive metabolites. There is little information on their biodegradation and toxicity in aquatic environments. Therefore, the biodegradability of the vinca alkaloids, and their toxicity towards wastewater bacteria were investigated in this study. The biodegradability of vindesine, vincristine, and vinblastine was examined in the closed bottle test (CBT). Additionally, the biodegradability of vinblastine as a model compound of the vinca alkaloids was tested in the Zahn-Wellens test (ZWT). The growth inhibition test with Pseudomonas putida was conducted, and a toxicity control in the CBT and the ZWT was used. The colony-forming units were monitored in the CBT; the test results for the biodegradability after 28 days were: 30% for vincristine, 20% for vindesine, and 10% for vinblastine. Therefore, none of the test compounds met the criteria for being readily biodegradable (> or = 60%). Vinblastine was biodegraded up to 18% in the ZWT after 40 days, and therefore, not inherently. Toxicity towards wastewater bacteria was not found. PMID- 11270418 TI - Tumor biology and prognosis in black breast cancer patients: a review. AB - American black women have a risk of developing breast cancer lower than that of American white women. but they have a worse prognosis when they do develop the disease. A major factor responsible for this discrepancy is a relatively high poverty level in the black population, with the consequent likelihood of delayed diagnosis and presentation with more advanced disease. However, breast cancer in black women also occurs at a younger age, more often is estrogen receptor negative, and more frequently exhibits aggressive biological behavior as judged by histopathologic grade, high tumor cell proliferation rate, and altered p53 expression. Obesity, known to be associated with a poor prognosis primarily as a consequence of increased estrogen production and bioavailability, is more common in black than in white breast cancer patients. An additional factor may be an earlier age at first completed pregnancy for black women, which is associated with a reduced breast cancer risk but also a poorer prognosis. Both the epidemiologic features and the tumor biology of breast cancer in black women serve to stress the particular importance of developing effective, specifically tailored strategies for early diagnosis in this ethnic group. PMID- 11270419 TI - Sexually transmitted infections and cervicovaginal dysplasia in a cohort of human immunodeficiency virus-positive women in Turin. AB - The correlation between sexually transmitted infections and cervicovaginal dysplasia has been evaluated in a cohort of 135 women who tested positive for human immunodeficiency virus type I (HIV-1) and were admitted to Amedeo di Savoia Hospital of Turin during the years 1997 and 1998 (stages B2 and B3 or C2 and C3). Of these women. 31 presented with sexually transmitted diseases (STDs; mean age, 33.5 +/- 5.9 years). Among them, 14 were affected by cervicovaginal dysplasia of differing severity; human papillomavirus (HPV) infection was found in 13 subjects (10 with cervicovaginal dysplasia). Herpes simplex virus type 2 (HSV-2) infection was detected in six women. Finally, Trichomonas vaginalis and Candida albicans were found in 10 and in 6 patients, respectively. Immunologic and hematologic evaluations were performed in the patients affected by STDs; in 28 patients of our case report unaffected by STDs but of similar ages (34.1 +/- 5.6 years) and stage of infection; and in 20 HIV-negative women unaffected by STDs. A significant reduction among the patients affected by STDs, as compared to those unaffected, was found in the case of white cells, CD4+ T cells, and ratio values (CD4 +/ and CD8 + T cells). Moreover, red cell count and hemoglobin concentration were lower in those women in the STD group. A lack of correlation was found between HIV RNA loads and CD4 + T cell counts and between HIV RNA and hemoglobin concentration in the patients with cervicovaginal dysplasia and in those affected by HSV-2 infection, which differed from the findings in subjects affected only by trichomoniasis or candidiasis. This suggests that the two former pathologic conditions (cervico-vaginal dysplasia and HSV-2 infection), other than HIV- I infection, may contribute to the impairment of these values. Moreover in our case report, T vaginalis and HSV-2 infections, which are suspected to have an oncogenic potential, do not seem to be relevant in the induction or facilitation of genital neoplastic diseases. Noteworthy is that the patients affected by HSV-2 infection, such as those affected by genital neoplastic diseases, showed the most compromised values of total white cells, CD4+ T cells, ratio index, red cells, and hemoglobin concentration. PMID- 11270420 TI - Quality assessment of cervical screening: a population-based case-control study in the C te-D'Or region, France. AB - Our objectives were to evaluate the effectiveness of cervical cancer screening outside organized programs in the prevention of cervical carcinoma in situ (CIS) and to enhance the way in which case control studies avoid some common biases. In our case-control study, we assessed all incident, histologically verified cases of CIS registered from 1987 to 1997 in the population-based cancer registry of C te-d'Or, France (N = 104) and 208 controls randomly selected from the screened population and matched for age, date of last screening, residence, and pathology laboratory results. We considered as appropriate for controls screened women who had had at least one Papanicolaou smear in the 3 years preceding the diagnosis or similar period. Screening for controls was higher (67.8%) than for cases (41.4%; P < .001), with a relative protection against CIS of 3.09 (95% confidence interval, 1.83-5.22) and a prevented fraction in the screened population of 45% to 50%. These findings suggest a protective advantage for CIS even in the absence of organized screening. The methodologic approach has advantages as compared to previous types of case-control studies. Although further refinements still are warranted, learning about the protective effect of screening for CIS provides information that may be useful in assessing the impact of a screening policy on women actually at risk of invasive cervical cancer. PMID- 11270421 TI - Parameters of differentiation and proliferation in adult granulosa cell tumors of the ovary. AB - We evaluated parameters of cell differentiation and proliferation to improve prognostication of ovarian adult granulosa cell tumors. Recurrent tumors (n = 10, REC group) and nonrecurrent tumors (n = 30, NED group) were compared in terms of cellular atypia, nuclear area, p53 overexpression, ploidy, DNA index, mitosis count, S-phase fraction, and nucleolar organizer region number and area per cell. Cellular atypia was significantly more frequent in REC than NED tumors (50% versus 13%; P = .03). Mean nuclear area was significantly larger in the REC than in the NED group (44 microm2 versus 36 microm2; P = .006). Mitotic count was significantly higher in REC than NED tumors (mean of 4.8 versus 1.7; P = .004). S phase fraction and ploidy did not predict outcome: neither did nucleolar organizer region numbers and area per cell, or p53 overexpression. Cellular atypia and mitotic count may help in determining the prognosis of adult granulosa tumors of the ovary. The histochemical parameters evaluated did not provide additional information. PMID- 11270422 TI - Detection of APC and k-ras mutations in the serum of patients with colorectal cancer. AB - Detection of tumor DNA in peripheral blood of patients with colorectal cancer (CRC) may allow early diagnosis of tumor disease and be of prognostic value. However, a reliable tumor marker detectable in the serum of patients with this disease is missing. Because k-ras and APC mutations occur frequently and at an early stage in CRCs, these mutations might also be detected in the serum of CRC patients and serve as tumor markers. Hence, tumor tissues of CRC patients were examined for the presence of mutations in the k-ras and APC genes. If a mutation was detected in the tumor, the serum of the patient was screened subsequently for the presence of this mutation. K-ray mutations were detected in 22 of 30 colorectal tumor tissues, but only in six patients was the mutation identified in their serum samples. Mutations of the APC gene were identified in 25 of 65 tumors: 20 of these 25 patients showed the respective mutation in their serum. Given their higher detection rate, APC mutations could be a more informative serum marker than k-ras in CRC patients. PMID- 11270423 TI - Metallothionein isoform 1 and 2 gene expression in the human bladder: evidence for upregulation of MT-1X mRNA in bladder cancer. AB - The goals of this study were to determine the expression of metallothionein isoform 1 and 2 proteins (MT-1 and MT-2) in bladder cancer and then to determine which MT isoform-specific genes promoted the expression of these proteins. Immunohistochemical analysis disclosed no immunoreactivity for MT-1 and MT-2 (designated as MT-1/2 to reflect the nonspecificity of the antibody for the two isoforms) in cells comprising the normal bladder or in nonmalignant bladder disorders, such as cystitis and interstitial cystitis. Immunohistochemical analysis demonstrated that MT-1/2 was overexpressed in all samples of carcinoma in situ and in high-grade bladder cancer, with variable overexpression in low grade bladder cancer and dysplastic lesions. The intensity and frequency of MT 1/2 staining correlated with the grade of the tumor. The MT-1 and MT-2 proteins are encoded by a family of eight genes (MT-1A, MT-1B, MT-1E, MT-1F, MT-1G, MT-IH, MT-1X, and MT-2A), and reverse transcriptase-polymerase chain reaction was used to determine which genes were expressed in the normal bladder and in bladder cancer. This analysis demonstrated that both normal and cancerous bladder tissue expressed mRNA for the MT-2A and MT-1X genes. The expression of MT-1E mRNA was variable in both normal bladder and bladder cancer specimens. Comparison of expression relative to that of beta-actin demonstrated that the level of MT-1X mRNA was overexpressed greatly in bladder cancer as compared to the level in normal bladder tissue. In contrast, the level of MT-2A mRNA was similar in both the normal and the bladder cancer specimens. The level of MT-1X expression did not vary with tumor grade. These studies suggest that the overexpression of MT 1/2 protein in bladder cancer is a result of the overexpression of the MT-1X gene. PMID- 11270424 TI - Possible implications of arginase and diamine oxidase in prostatic carcinoma. AB - Polyamine metabolism in prostatic tissue, which may give rise to prostatic hyperplasia by inducing cell proliferation, is controlled primarily by two enzymes: arginase and diamine oxidase (DAO). Arginase catalyzes the synthesis of ornithine, the precursor for polyamines from arginine, whereas DAO catalyzes the oxidation of diamines. such as spermine and spermidine, to a much less active compound called putrescine. In this study, we evaluated arginase and DAO activities in prostate tissues of 50 patients with benign prostatic hyperplasia and in 23 patients with prostatic carcinoma. Both DAO and arginase activities were found to be elevated in cancer tissues as compared to benign prostatic hyperplasia (fivefold and twofold, respectively; P < .001). A strong inverse correlation between arginase and DAO activities was observed in those in the cancer group (r = -0.65; P < .001). Gleason grades of prostatic carcinomas were well correlated with both arginase and DAO activities. Results suggest that DAO and arginase might play an essential role in the mechanism of prostatic disease process by modulating polyamine levels. PMID- 11270425 TI - Thymidine kinase 1 immunoassay: a potential marker for breast cancer. AB - Previous research indicates that thymidine kinase I (TKI) possesses value as a tool for both prognosis and diagnosis in breast cancer. However, drawbacks to the existing radioassay for thymidine kinase have frustrated its clinical use. To overcome these drawbacks, we developed a monoclonal antibody to TK1. We have assessed this antibody for a linear antibody-antigen response and for reproducibility using ELISA techniques. We also have evaluated this antibody for TKI specificity as determined by Western blot. To test the accuracy of this monoclonal antibody further, we treated human MCF-7 breast cancer cells with tamoxifen and measured decreasing TKI activity and protein levels with the radioassay and with our monoclonal antibody in an ELISA, respectively. We then used the radioassay and our monoclonal antibody to measure TK1 activity and protein levels, respectively, in 218 serum samples of postoperative breast cancer patients and found a correlation between the two assays. Our results demonstrated that the TK1 immunoassay not only had a linear, reproducible, and specific response but accurately measured TK1 levels in both MCF-7 breast cancer cells and serum. Thus, our monoclonal antibody may demonstrate potential for practical use in a clinical setting for the management of breast cancer. PMID- 11270426 TI - Irradiation-induced effects on mast cells, neuropeptides, and atrial natriuretic peptide in the rat heart and lung: bases for further studies. AB - We examined the effects of irradiation over the thorax of the rat on the mast cells, the neuropeptide-containing nerve fibers, and the expression of atrial natriuretic peptide in the heart and lung. The total doses were 20 to 36 Gy delivered as single doses or fractionated irradiation. Immunohistochemical and radioimmunoassay methods were used. The number of mast cells was much reduced in both the lung and heart in response to irradiation. A trend for lowering the atrial natriuretic peptide levels in plasma was noted both 1 day and 9 days after irradiation. In contrast to the situation in other organs (salivary and laryngeal glands, the intestine), no changes occurred in the immunohistochemical expression of neuropeptides. With these observations and those made in previous studies about the effects of radiotherapy on other organs, the functional significance and basis for further research in the fields are discussed. PMID- 11270427 TI - Assessment of the cytotoxic and clastogenic activities of the sesquiterpene lactone lynchnopholide in mammalian cells in vitro and in vivo. AB - Lychnopholide (LNP), a sesquiterpene lactone with antitumor, trypanocidal, and antimicrobial activities, was isolated from Vanillosmopsis erythropappa. The present study was carried out to assess the cytotoxic and clastogenic potential of this new agent in human cultured lymphocytes and Swiss bone marrow cells before the agent was used in medicine. The mitotic index, chromosomal aberrations (CAs), sister chromatid exchanges (SCEs), and proliferation index were investigated. There was no alteration in the number of CAs and SCEs in the continuous in vitro treatment. However, the highest concentration (0.2 microg/ml) of LNP was cytotoxic. LNP (0.1, 0.2, and 0.4 microg/ml) induced a significant increase in CA frequency at the G2 phase in all treated cultures. Only the highest concentration (26.67 mg/kg) caused a significant increase in the total number of CAs in the in vivo investigation. On the basis of these results, LNP had a clastogenic effect on both test systems and a cytotoxic effect in vitro. PMID- 11270428 TI - Active auditory mechanics in mosquitoes. AB - In humans and other vertebrates, hearing is improved by active contractile properties of hair cells. Comparable active auditory mechanics is now demonstrated in insects. In mosquitoes, Johnston's organ transduces sound-induced vibrations of the antennal flagellum. A non-muscular 'motor' activity enhances the sensitivity and tuning of the flagellar mechanical response in physiologically intact animals. This motor is capable of driving the flagellum autonomously, amplifying sound-induced vibrations at specific frequencies and intensities. Motor-related electrical activity of Johnston's organ strongly suggests that mosquito hearing is improved by mechanoreceptor motility. PMID- 11270429 TI - Polyploidy alters advertisement call structure in gray treefrogs. AB - Whole-genome duplication is believed to have played a significant role in the early evolution and diversification of vertebrate animals. The establishment of newly arisen polyploid lineages of sexually reproducing animals requires assortative mating between polyploids. Here, we show that genome duplication can directly alter a phenotypic trait mediating mate choice in the absence of genotypic change. Our results suggest that the direct effect of polyploidy on behaviour is a consequence of increased cell size. PMID- 11270430 TI - Kin-selected conflict in the bumble-bee Bombus terrestris (Hymenoptera: Apidae). AB - Kin selection theory predicts conflict in social Hymenoptera between the queen and workers over male parentage because each party is more closely related to its own male offspring. Some aspects of the reproductive biology of the bumble-bee Bombus terrestris support kin selection theory but others arguably do not. We present a novel hypothesis for how conflict over male parentage should unfold in B. terrestris colonies. We propose that workers delay laying eggs until they possess information showing that egg laying suits their kin-selected interests. In colonies where queens start to lay haploid eggs early, we hypothesize that this occurs when workers detect the presence of queen-produced male brood in the brood's larval stage. In colonies where queens start to lay haploid eggs late, we hypothesize that it occurs when workers detect a signal from the queen to female larvae to commence development as queens. Our hypothesis accounts for previously unexplained aspects of the timing of reproductive events in B. terrestris, provides ultimate explanations for the results of a recent study of mechanisms underlying queen-worker conflict and helps explain this species' characteristic bimodal (split) sex ratios. Therefore, kin selection theory potentially provides a good explanation for reproductive patterns in B. terrestris. PMID- 11270431 TI - Predator experience on cryptic prey affects the survival of conspicuous aposematic prey. AB - Initially, aposematism, which is an unprofitable trait, e.g. noxiousness conspicuously advertised to predators, appears to be a paradox since conspicuousness should increase predation by naive predators. However, reluctance of predators for eating novel prey (e.g. neophobia) might balance the initial predation caused by inexperienced predators. We tested the novelty effects on initial predation and avoidance learning in two separate conspicuousness levels of aposematic prey by using a 'novel world' method. Half of the wild great tits (Parus major) were trained to eat cryptic prey prior to the introduction of an aposematic prey, which potentially creates a bias against the aposematic morph. Both prey types were equally novel for control birds and they should not have shown any biased reluctance for eating an aposematic prey. Knowledge of cryptic prey reduced the expected initial mortality of the conspicuous morph to a random level whereas control birds initially ate the conspicuous morph according to the visibility risk. Birds learned to avoid conspicuous prey in both treatments but knowledge of cryptic prey did not increase the rate of avoidance learning. Predators' knowledge of cryptic prey did not reduce the predation of the less conspicuous aposematic prey and additionally predators did not learn to avoid the less conspicuous prey. These results indicate that predator psychology, which was shown as reluctance for attacking novel conspicuous prey, might have been important in the evolution of aposematism. PMID- 11270433 TI - Depletion models can predict shorebird distribution at different spatial scales. AB - Predicting the impact of habitat change on populations requires an understanding of the number of animals that a given area can support. Depletion models enable predictions of the numbers of individuals an area can support from prey density and predator searching efficiency and handling time. Depletion models have been successfully employed to predict patterns of abundance over small spatial scales, but most environmental change occurs over large spatial scales. We test the ability of depletion models to predict abundance at a range of scales with black tailed godwits, Limosa limosa islandica. From the type II functional response of godwits to their prey, we calculated the handling time and searching efficiency associated with these prey. These were incorporated in a depletion model, together with the density of available prey determined from surveys, in order to predict godwit abundance. Tests of these predictions with Wetland Bird Survey data from the British Trust for Ornithology showed significant correlations between predicted and observed densities at three scales: within mudflats, within estuaries and between estuaries. Depletion models can thus be powerful tools for predicting the population size that can be supported on sites at a range of scales. This greatly enhances our confidence in predictions of the consequences of environmental change. PMID- 11270432 TI - Effects of demanding foraging conditions on cache retrival accuracy in food caching mountain chickadees (Poecile gambeli). AB - Birds rely, at least in part, on spatial memory for recovering previously hidden caches but accurate cache recovery may be more critical for birds that forage in harsh conditions where the food supply is limited and unpredictable. Failure to find caches in these conditions may potentially result in death from starvation. In order to test this hypothesis we compared the cache recovery behaviour of 24 wild-caught mountain chickadees (Poecile gambeli), half of which were maintained on a limited and unpredictable food supply while the rest were maintained on an ad libitum food supply for 60 days. We then tested their cache retrieval accuracy by allowing birds from both groups to cache seeds in the experimental room and recover them 5 hours later. Our results showed that birds maintained on a limited and unpredictable food supply made significantly fewer visits to non-cache sites when recovering their caches compared to birds maintained on ad libitum food. We found the same difference in performance in two versions of a one-trial associative learning task in which the birds had to rely on memory to find previously encountered hidden food. In a non-spatial memory version of the task, in which the baited feeder was clearly marked, there were no significant differences between the two groups. We therefore concluded that the two groups differed in their efficiency at cache retrieval. We suggest that this difference is more likely to be attributable to a difference in memory (encoding or recall) than to a difference in their motivation to search for hidden food, although the possibility of some motivational differences still exists. Overall, our results suggest that demanding foraging conditions favour more accurate cache retrieval in food-caching birds. PMID- 11270434 TI - Environmental response of upper trophic-level predators reveals a system change in an Antarctic marine ecosystem. AB - Long-term changes in the physical environment in the Antarctic Peninsula region have significant potential for affecting populations of Antarctic krill (Euphausia superba), a keystone food web species. In order to investigate this, we analysed data on krill-eating predators at South Georgia from 1980 to 2000. Indices of population size and reproductive performance showed declines in all species and an increase in the frequency of years of low reproductive output. Changes in the population structure of krill and its relationship with reproductive performance suggested that the biomass of krill within the largest size class was sufficient to support predator demand in the 1980s but not in the 1990s. We suggest that the effects of underlying changes in the system on the krill population structure have been amplified by predator-induced mortality, resulting in breeding predators now regularly operating close to the limit of krill availability. Understanding how krill demography is affected by changes in physical environmental factors and by predator consumption and how, in turn, this influences predator performance and survival, is one of the keys to predicting future change in Antarctic marine ecosystems. PMID- 11270435 TI - Evolution of reduced dispersal mortality and 'fat-tailed' dispersal kernels in autocorrelated landscapes. AB - Models describing the evolution of dispersal strategies have mostly focused on the evolution of dispersal rates. Taking trees as a model for organisms with undirected, passive dispersal, we have developed an individual-based, spatially explicit simulation tool to investigate the evolution of the dispersal kernel, P(r), and its resulting cumulative seed-density distribution, D(r). Simulations were run on a variety of fractal landscapes differing in the fraction of suitable habitat and the spatial autocorrelation. Starting from a uniform D(r), evolution led to an increase in the fraction of seeds staying in the home cell, a reduction of the dispersal mortality (arrival in unsuitable habitat), and the evolution of 'fat-tailed' D(r) in autocorrelated landscapes and approximately uniform D(r) in random landscapes. The evolutionary process was characterized by long periods of stasis with a few bouts of rapid change in the dispersal rate. PMID- 11270436 TI - Coevolution between a cockroach and its bacterial endosymbiont: a biogeographical perspective. AB - Cryptocercus are subsocial, xylophagous cockroaches that live in temperate forests. Like other cockroaches, Cryptocercus harbour endosymbiotic bacteria in their fat bodies. Two species of Cryptocercus occur in the palaearctic, one each in eastern Russia and south-central China. In the USA, there are five species: one in the north-west and four in the south-east. Little is known about the relationship between the Eurasian and North American Cryptocercus or the causes of the disjunct distribution. Here, a molecular phylogeny for six out of the seven Cryptocercus species and their endosymbionts is inferred in an attempt to understand the evolution and biogeography of the genus. Our analysis showed that the North American Cryptocercus are monophyletic, suggesting that a single colonization event was followed by vicariance. There was complete concordance between the host and endosymbiont phylogenetic trees. Divergence estimates based on endosymbiont DNA sequences suggested that the palaearctic and nearctic Cryptocercus diverged 70-115 million years (Myr) ago and the eastern- and western USA species diverged 53-88 Myr ago. These divergence estimates were correlated with biogeographical events, and a hypothesis is presented to explain the current distribution of Cryptocercus. Our findings suggest that Cryptocercus has had a long evolutionary history, dating back to the Jurassic. PMID- 11270437 TI - Genetic divergence of the seminal signal-receptor system in houseflies: the footprints of sexually antagonistic coevolution? AB - To understand fully the significance of cryptic female choice, we need to focus on each of those postmating processes in females which create variance in fitness among males. Earlier studies have focused almost exclusively on the proportion of a female's eggs fertilized by different males (sperm precedence). Yet, variance in male postmating reproductive success may also arise from differences in ability to stimulate female oviposition and to delay female remating. Here, we present a series of reciprocal mating experiments among genetically differentiated wild-type strains of the housefly Musca domestica. We compared the effects of male and female genotype on oviposition and remating by females. The genotype of each sex affected both female oviposition and remating rates, demonstrating that the signal-receptor system involved has indeed diverged among these strains. Further, there was a significant interaction between the effects of male and female genotype on oviposition rate. We discuss ways in which the pattern of such interactions provides insights into the coevolutionary mechanism involved. Females in our experiments generally exhibited the weakest, rather than the strongest, response to males with which they are coevolved. These results support the hypothesis that coevolution of male seminal signals and female receptors is sexually antagonistic. PMID- 11270439 TI - A phylogeny of the land snails (Gastropoda: Pulmonata). AB - We have undertaken the first large-scale molecular phylogenetic analysis of the Stylommatophora. Sequences of the ribosomal RNA gene-cluster were examined in 104 species of snails and slugs from 50 families, encompassing all the currently recognized major groups. It allows an independent test of the present classification based on morphology. At the level of families our molecular phylogeny closely supports the current taxonomy, but the deep branches within the tree do not. Surprisingly, a single assemblage including the families Achatinidae, Subulinidae and Streptaxidae lies near the base of the tree, forming a sister group to all remaining stylommatophorans. This primary division into 'achatinoid' and 'non-achatinoid' taxa is unexpected, and demands a radical reinterpretation of early stylommatophoran evolution. In particular, the Orthurethra appear to be relatively advanced within the 'non-achatinoid clade', and broadly equivalent to other super-familial clusters. This indicates that supposedly primitive features such as the orthurethran kidney are derived. The molecular tree also suggests that the origin of the Stylommatophora is much earlier than the main period of their diversification. PMID- 11270438 TI - Semelparity in a large marsupial. AB - Complete mortality of males after mating is known in several small dasyurid and didelphid species (up to 300g) and has previously been suggested to be a consequence of their small size and their inability to sequester sufficient fat reserves for an intense rut in the winter. Males of these species use increased corticosteroid levels to allow protein catabolism, enabling them to support their mating effort with other body reserves. However, increased corticosteroid levels have negative consequences such as anaemia, gastrointestinal ulceration, immune suppression and disease. The Australian dasyurid Dasyurus hallucatus shows complete male die off after mating in tropical savannah, yet males of this species may weigh as much as 1120 g and continue to eat during the rut. Die off in D. hallucatus shows many similarities to that in the smaller species including weight loss, fur loss, parasite infestation, increased testosterone levels and anaemia. However, in contrast to smaller species, there is no evidence of elevated corticosteroid levels or gastrointestinal ulceration. Consequently, the phenomenon of male die off after mating lacks a universal explanation. PMID- 11270440 TI - Experimental manipulation of female reproduction reveals an intraspecific egg size-clutch size trade-off. AB - A negative relationship, or trade-off, between egg size and clutch size is a central and long-standing component of life-history theory, yet there is little empirical evidence for such a trade-off, especially at the intraspecific level. Here, I show that female zebra finches (Taeniopygia guttata) treated chronically during egg formation with the anti-oestrogen tamoxifen lay smaller eggs (by 8%) but produce larger clutches (on average two eggs more) than controls. Decreased egg mass in tamoxifen-treated females was associated with a 50% decrease in plasma levels of the two yolk precursors, vitellogenin and very-low-density lipoprotein. Although tamoxifen-treated females laid more, smaller eggs (and had a higher total expenditure in their clutch), they did not differ from controls in the number of chicks fledged, the mass or size of these chicks at fledging, or the chicks' egg-production performance at three months of age. However, tamoxifen treated females had lower relative hatching success: they laid more eggs but hatched the same number of chicks. Among individual tamoxifen-treated females, birds that laid the smallest eggs early in their laying sequence laid the largest number of additional eggs, that is, there was a negative correlation, or trade off, between egg size and clutch size. PMID- 11270441 TI - Cytotoxic T-lymphocyte memory, virus clearance and antigenic heterogeneity. AB - Cytotoxic T-lymphocyte (CTL) memory to viruses has traditionally been studied in an isolated setting. However, recent experiments have indicated that the presence of antigenically heterologous challenges can result in the attrition of CTL memory. Here we use mathematical models in order to explore the consequence of these dynamics for the ability of the immune system in controlling multiple infections. Mathematical models suggest that antigen-independent persistence of CTL memory is required in order to resolve and clear an infection. This ensures strong immunological pressure at low loads when the virus population declines towards extinction. If the number of antigenic stimuli exposed to the immune system crosses a threshold, we find that immunological pressure is significantly reduced at low loads and this can prevent virus clearance and reduces overall control of viral replication. Hence, exposure to many heterologous challenges reduces the ability of CTL memory to contribute to virus control. The higher the number of infections present in the host, the higher the overall virus load and the higher the total number of memory CTLs. Beyond a given threshold, addition of new viruses to the system results in accelerated loss of virus control which eventually leads to a reduction in the overall memory CTL population. These dynamics might contribute to the progressively weaker immunity observed as a result of ageing. In this context, antigenically variable pathogens expose the immune system to many heterologous challenges within a short period of time and this could result in accelerated ageing of the immune system. These results have important implications for vaccination and treatment strategies directed against viral infections. PMID- 11270442 TI - Slow and fast visual motion channels have independent binocular-rivalry stages. AB - We have previously reported a transparent motion after-effect indicating that the human visual system comprises separate slow and fast motion channels. Here, we report that the presentation of a fast motion in one eye and a slow motion in the other eye does not result in binocular rivalry but in a clear percept of transparent motion. We call this new visual phenomenon 'dichoptic motion transparency' (DMT). So far only the DMT phenomenon and the two motion after effects (the 'classical' motion after-effect, seen after motion adaptation on a static test pattern, and the dynamic motion after-effect, seen on a dynamic-noise test pattern) appear to isolate the channels completely. The speed ranges of the slow and fast channels overlap strongly and are observer dependent. A model is presented that links after-effect durations of an observer to the probability of rivalry or DMT as a function of dichoptic velocity combinations. Model results support the assumption of two highly independent channels showing only within channel rivalry, and no rivalry or after-effect interactions between the channels. The finding of two independent motion vision channels, each with a separate rivalry stage and a private line to conscious perception, might be helpful in visualizing or analysing pathways to consciousness. PMID- 11270443 TI - The gender identity disorder in the DSM-IV classification: a critical evaluation. AB - OBJECTIVE: The DSM-IV classification in its definition and description of the gender identity disorder omits a number of diagnostically significant features. This paper attempts to correct the deficiencies. METHOD: The text under the headings: 'Diagnostic features', 'Specifiers', 'Associated disorders', 'Laboratory findings', 'Prevalence', 'Course' and 'Differential diagnosis' is subjected to a detailed scrutiny, using the author's experience as consultant psychiatrist to the Monash University Gender Dysphoria Clinic over a period of 25 years as source and background. Results of two studies of male-to-female and female-to-male transsexuals are given in the Table. RESULTS: DSM-IV criteria are augmented and the symptomatology focused. The existing gaps in the delineation of specifiers and associated features are closed by providing additional clinical material. The description of the course and the differential diagnosis are enriched. CONCLUSIONS: Although the critical analysis of the DSM-IV classification of the gender identity disorder has shown the manual to be adequate, it nevertheless has shortcomings which may impede exact diagnosis. PMID- 11270444 TI - The role of economic evaluation in mental health care. AB - OBJECTIVE: A consequence of the integration of psychiatry into acute and public health medicine is that psychiatrists are being asked to evaluate their services. There is pressure on mental health-care systems because it is recognized that funds should be directed where they can provide the best health outcomes, and also because there are resource constraints which limit our capacity to meet all demands for health care. This pressure can be responded to by evaluation which demonstrates the effectiveness and efficiency of psychiatric treatment. This paper seeks to remind psychiatrists of the fundamental principles of economic evaluation in the hope that these will enable psychiatrists to understand the methods used in evaluation and to work comfortably with evaluators. METHOD: The paper reviews the basic principles behind economic evaluation, illustrating these with reference to case studies. It describes: (i) the cost of the burden of illness and treatment, and how these costs are measured; (ii) the measurement of treatment outcomes, both as changes in health status and as resources saved; and (iii) the various types of economic evaluation, including cost-minimization, cost effectiveness, cost-utility and cost-benefit analysis. RESULTS: The advice in the paper provides psychiatrists with the necessary background to work closely with evaluators. A checklist of the critical questions to be addressed is provided as a guide for those undertaking economic evaluations. CONCLUSIONS: If psychiatrists are willing to learn the basic principles of economic evaluation and to apply these, they can respond to the challenges of evaluation. PMID- 11270445 TI - Improving services for people with a psychotic disorder and problematic substance use: a multifaceted approach to project evaluation. AB - OBJECTIVE: The objective of this study was to conduct a multifaceted formative evaluation of the Central Sydney Area Health Service (CSAHS) Psychosis and Substance Use Project. METHOD: Four evaluative methods were used: (i) description and interpretation of the Project's documented processes and outcomes; (ii) a benchmark comparison of the Project processes and outcomes against three of the 11 National Standards for Mental Health Services; (iii) a survey of the Project's key stakeholders; and (iv) interviews with 12 purposefully sampled key informants. RESULTS: The Project achieved its aim to develop a strategy to improve services for people with comorbid psychosis and problematic substance use. Three of the five Project objectives were fully achieved: examination of current clinical services, development of a clinical services plan, and development of a staff education programme. The Project partially achieved two objectives: development of an information system, and a research agenda. The Project and CSAHS performed well when measured against three of the National Mental Health Standards. Project participants perceived the Project to have been successful and worth continuing, identified some shortcomings and made recommendations for the second phase. CONCLUSIONS: The participatory approach to the Project and the evaluation was successful. With some improvements the Project is worth continuing into a second phase. A multifaceted approach and qualitative research methods are useful for formative evaluation of health service programmes. PMID- 11270446 TI - A clinical validation of the Dysmorphic Concern Questionnaire. AB - OBJECTIVE: The current study addressed the concept of dysmorphic concern as a symptom that may exist in a number of disorders. The aims of the study were to: (i) validate a recently developed questionnaire that measures dysmorphic concern, the Dysmorphic Concern Questionnaire (DCQ); and (ii) evaluate the relationship of dysmorphic concern to depressed mood, social phobia, and obsessive-compulsive symptomatology. METHOD: Sixty-five psychiatric inpatients were diagnosed using the computerized version of the Composite International Diagnostic Interview (CIDI-A). They then completed the DCQ, and questionnaires measuring body dysmorphic disorder (the Body Dysmorphic Disorder Examination, or BDDE), depression, social phobia, and obsessive-compulsive disorder (OCD). The factor structure and convergent validity of the DCQ were determined, and associations with mood and anxiety symptoms explored. RESULTS: The DCQ was found to be a reliable and valid instrument that is sensitive to dysmorphic concern. Furthermore, although dysmorphic concern was associated with body dysmorphic disorder (BDD), depression, social phobia and OCD, only the score from the BDDE predicted DCQ score in a multiple regression analysis. Finally, BDD symptomatology was best defined by the presence of negative body beliefs as measured by the DCQ. CONCLUSIONS: Negative body beliefs are the hallmark of BDD. However, the existence of dysmorphic concern does not necessarily imply a diagnosis of BDD. The DCQ is a quick and efficient means of identifying dysmorphic concern in those who present with depression, OCD, social phobia or BDD. PMID- 11270447 TI - Psychotic depression, subcortical arteriosclerotic encephalopathy and holocaust conditioned posttraumatic stress disorder. PMID- 11270448 TI - Committal of a patient due to media influence? A case report. PMID- 11270449 TI - Signal transmission, schizophrenia and the limits to biological psychiatry. PMID- 11270450 TI - Maintenance electroconvulsive therapy in schizophrenia. PMID- 11270451 TI - Uncommon self-mutilation in a borderline personality disorder patient. PMID- 11270453 TI - Publishing ethics in psychiatry. AB - OBJECTIVE: The ethics of publishing in psychiatry has gained only limited attention. We examine, in a historical context, pertinent ethical problems and offer a series of recommendations for dealing with them. METHOD: The study was conducted by exploration of medical databases and websites, and systematic discussion with ethicists, legal experts, publishers and researchers. RESULTS: Serious 'publishing misconduct' appears to have been rare in the psychiatric literature, but any occurrence of redundant publication, plagiarism and publication of fraudulent or inhumane research is disturbing and should be prevented. Difficulties around authorship, sensitive use of language, conflict of interest and bias in the publishing process are additional issues claiming our attention. CONCLUSIONS: A clearly articulated publishing ethos is desirable. Potential strategies to achieve this include devising guidelines on publishing ethics, teaching the subject to new writers, journals committing themselves to the ethical dimension of their operations, and penalising colleagues who violate agreed principles of good conduct. PMID- 11270452 TI - Evolutionary approaches to psychopathology: the role of natural defences. AB - OBJECTIVE: Psychoanalytic theories of the mind emerged in the immediate post Darwinian era of the 1880s and 1890s. Since that time much has changed in both psychoanalytic and evolutionary theorizing. This paper explores recent evolutionary thinking on psychopathology. METHOD: Relevant literature was reviewed. RESULTS: This paper outlines some of the common behavioural defence mechanisms and then explores ways in which they are represented in various disorders, with a focus on depression. This paper suggests that 'symptoms' can be related to the activation of evolved defence mechanisms to respond to losses and threats. Such will involve, for example, anxious arousal and heightened vigilance and attention to the threat, with the type of defence (e.g. fight, flight, submit, help seeking) being mirrored in particular symptom presentations. CONCLUSION: Defences can become pathological when they are too easily aroused or prolonged, are arrested (aroused but not expressed) and/or ineffective. PMID- 11270454 TI - Better mental health services for young people: responsibility, partnerships and projects. AB - OBJECTIVE: This paper argues that adolescent psychiatry is best linked with child psychiatry and opposes separate youth mental health programmes for 12-25-year olds. It reports on the current status of services and considers how adult mental health services (AMHS) can improve services for young adults (18-25-year-olds). METHOD: Factors in development, psychopathology, prevention, training and service systems are reviewed to suggest that current child and adolescent mental health service systems (CAMHS) are appropriate for 0-17-year-olds. Improvements in CAMHS are described from a Victorian perspective, including the model of specialist clinical programmes or teams for specific patient populations. Mechanisms are outlined for AMHS to better assist young adults from 18 to 25 years of age. RESULTS: The model of clinical projects or clinical programme teams, developed in partnership with primary health and others, is a suitable vehicle to help AMHS to improve clinical services to their young adult populations. These may be funded from a variety of sources, including re-engineering existing service resources. CONCLUSIONS: Such developments complement the work of specialist research units and build local competencies. More programme development and evaluation is needed, which will require the support of the College and State and Commonwealth Mental Health Branches. PMID- 11270455 TI - Implications for Australia of the multimodal treatment study of children with attention-deficit/hyperactivity disorder. AB - OBJECTIVES: This paper reviews the implications of the large-scale USA Multimodal Treatment Study of Children with Attention-Deficit/Hyperactivity Disorder (MTA) by the MTA Cooperative group for planning of clinical services for children with attention deficit hyperactivity disorder in Australia. METHOD: The MTA study findings of no significant difference between active medication treatment alone, and combined medication combined with behaviour therapy treatments, on core symptom outcomes, are examined. Service traditions, workforce issues, diagnostic and ethical and philosophical considerations are discussed in relation to their impact on service planning in Australia. Implications of pharmacogenomic research are examined. Ethical and philosophical questions are raised in relation to the use of stimulant medications for subthreshold symptoms and for socialization of children. RESULTS: A critical evaluation of results reveals that combined treatments allow the use of lower medication doses, and that multimodal treatments are effective for comorbid symptoms. Completion of the Human Genome Project promises increasing technological advances. Arguments for and against a technological approach to child rearing are posited. CONCLUSIONS: The MTA study raises not just service planning questions, but also important ethical and philosophical considerations about optimal degree of medicalization of services in an increasingly technological society. Child psychiatrists will be required to have an understanding of technological developments if they wish to contribute to future debates. PMID- 11270457 TI - Transcranial magnetic stimulation: experience, knowledge and attitudes of recipients. AB - OBJECTIVE: To examine the experience, knowledge and attitudes of recipients of transcranial magnetic stimulation (TMS) regarding the treatment. There have been no studies of patient views about TMS. METHOD: A 60-item survey was administered by telephone to persons with depressive illness who received TMS at Royal Hobart Hospital, Tasmania. RESULTS: Forty-eight patients were interviewed. About two thirds also had a history of treatment with electroconvulsive therapy (ECT). Experience and opinions about TMS were generally very positive. Almost three quarters of interviewees believed TMS had been helpful. The vast majority rated TMS as more acceptable than having, or the prospect of having, ECT. The majority would have TMS again and would recommend it to others. CONCLUSIONS: The mostly favourable experiences and attitudes reported by interviewees will be reassuring to patients, their families and treating health professionals when TMS is being considered. PMID- 11270456 TI - Relative misery and youth suicide. AB - OBJECTIVE: To test the 'absolute misery hypothesis' that suicide rates are a proxy measure of psychological maladjustment within the general population of young people. METHOD: Study I regressed World Health Organization statistics on youth suicide rates on measures of adolescent adjustment across seven countries. Study II analysed the results of a Canadian survey involving 2,111 children from 31 schools in grades seven to 12 (ages 11-20 years, mean = 15.5, SD = 1.7). The survey contained measures of suicidality, depressed affect and social comparison. RESULTS: Study I found that male suicide was much more likely in psychologically well-adjusted countries than in less well-adjusted countries. Although not statistically significant in a sample of this size (n = 7), correlation analysis suggested that the relationship between suicide and adjustment was in the opposite direction for females. Study II found that suicidality in boys was not associated with depressed affect on its own, or with social comparison on its own, but was associated with the combination of depressed affect and negative social comparison. By contrast, suicidality in girls was significantly associated both with absolute and comparative levels of unhappiness. CONCLUSIONS: A new, 'relative misery hypothesis' is proposed to account for these results. Under this hypothesis, the disposition of vulnerable young men towards suicide is influenced by their affective state relative to others. When those around them are perceived to be better off than they are, the predisposition of young men to suicide is increased. By contrast, female suicide is predicted to be less influenced by young women's relative state, and more by their absolute level of unhappiness. The primary implication of the relative misery hypothesis is that the prevention of young male suicide in particular is likely to require methods that discourage vulnerable individuals from making negative social comparisons. PMID- 11270458 TI - Cognitive-behavioural therapy via videoconferencing to a rural area. AB - OBJECTIVE: This case report describes the use of cognitive-behavioural therapy via two-way, interactive audiovisual videoconferencing and identifies issues involved in using this form of technology to provide therapy. CLINICAL PICTURE: A 38-year-old married woman living in rural South Australia presented with panic disorder with agoraphobia and major depression. The patient had refused antidepressant treatment. TREATMENT: The patient was treated with 12 sessions of cognitive-behavioural therapy delivered via videoconferencing. OUTCOME: Anxiety and depressive symptoms resolved with concomitant improvement in function. CONCLUSIONS: Providing this form of therapy via videoconferencing can be effective. PMID- 11270460 TI - Obstetric risk factors for postnatal depression in urban and rural community samples. AB - OBJECTIVE: The objective of this study was to examine obstetric risk factors for postnatal depression in an urban and rural community sample, with concurrent consideration of personality, psychiatric history and recent life events. METHODS: This was a prospective study with women planning to give birth in one of the four participating hospitals recruited antenatally. Obstetric information was obtained from the New South Wales Midwives Data Collection, completed shortly after delivery. Personality, psychiatric history and life-events information were obtained from a questionnaire, administered within 1 week postpartum. Depression status was assessed at 8 weeks postpartum using the Edinburgh Postnatal Depression Scale. RESULTS: Complete data were obtained from 490 women. Several non-obstetric risk factors for the development of postnatal depression at 8 weeks postpartum were reported including: sociodemographic (up to technical college level education, rented housing, receiving a pension/benefit), personality (those who described themselves as either nervy, shy/selfconscious, obsessional, angry or a worrier), psychiatric history (familial history of mental illness, personal history of depression or anxiety or a history of depression in the participant's mother) and recent life-events (major health problem, arguments with partner and friends/relatives). None of the obstetric variables were significantly associated with increased risk for postnatal depression, but several showed marginally significant increases (multiparous women, antepartum haemorrhage, forceps and caesarean section deliveries). CONCLUSIONS: The results emphasize the importance of psychosocial risk factors for postnatal depression and suggest that most obstetric factors during pregnancy and birth do not significantly increase risk for this depression. Early identification of potential risk for postnatal depression should include assessment of sociodemography, personality, psychiatric history and recent life events, as well as past and present obstetric factors. PMID- 11270459 TI - Treatment of psychogenic polydipsia: comparison of risperidone and olanzapine, and the effects of an adjunctive angiotensin-II receptor blocking drug (irbesartan). AB - OBJECTIVE: Our objective was to determine the outcome of novel strategies in managing a case of severe polydipsia. CLINICAL PICTURE: The patient was a 39-year old male with a 20-year history of paranoid schizophrenia who, despite only mild residual psychotic symptoms, had been hospitalized for the previous 10 years because of severe polydipsic behaviour complicated by water intoxication. TREATMENT: Novel antipsychotic agents, risperidone and olanzapine, as well as the specific angiotensin-II receptor blocking drug, irbesartan were employed at selected intervals in a study lasting nearly 3 years. A strict behavioural management programme was ongoing, in which diurnal weight change and the number of breaches of weight limits, requiring management in a low-stimulus environment, were documented on a daily basis. Summary measures of diurnal weight change and behavioural intervention were charted against changes in treatment. OUTCOME: Polydipsic behaviour improved on risperidone up to 4 mg daily, but was not sustained. Olanzapine was similarly successful in stabilizing polydipsia, and improvement was achieved with the addition of irbesartan. CONCLUSION: We suggest that the D2-sparing profiles of receptor binding achieved with low-dose risperidone and olanzapine may account for this beneficial effect. The benefit derived with irbesartan implicates the involvement of brain angiotensin systems centrally in helping to regulate drinking behaviour. PMID- 11270461 TI - The mental and physical health of female sex workers: a comparative study. AB - OBJECTIVES: The objective of this study was to compare the mental and physical health, adult abuse experiences and social networks of female sex workers with data previously collected from two large community samples of age-matched women. METHOD: A convenience sample of sex workers were interviewed and completed two well-established questionnaires, the General Health Questionnaire (GHQ-28) and the Intimate Bond Measure (IBM). Sex workers were invited to reflect on their experiences of their work. RESULTS: There were no differences in mental health on the GHQ-28 or in self-esteem (measured by an item on the Present State Examination) between the two groups. Neither were there any differences in their assessment of their physical health or the quality of their social networks. Sex workers were less likely to be married and had been exposed to more adult physical and sexual abuse than the comparison group. They were more likely to smoke and to drink heavily when they drank. One-third said that their general practitioner was not aware of their work. A subgroup not working with regular clients or in a massage parlour had higher GHQ-28 scores and may be an at-risk group. Narrative information about the work, particularly its intermittent nature, is presented. CONCLUSIONS: No evidence was found that sex work and increased adult psychiatric morbidity are inevitably associated, although there may be subgroups of workers with particular problems. The illegal and stigmatized nature of sex work are likely to make usual public health strategies more difficult to apply, considerations which should give concern from a preventive health standpoint. PMID- 11270462 TI - Assessing stressful life events in relation to liability and compensation. AB - OBJECTIVE: The aim of this paper is to outline limitations in the assessment of the relationship between stressful experiences and psychological disorder in the medico-legal setting. METHOD: A research-derived approach to more objectively assessing stressful life events and disorder is discussed in the light of limitations or biases which may arise in the evaluation of the clinical significance of stressors in psychological disorder particularly in a medico legal context. RESULTS: There may be considerable bias in the assessment of stressful experiences in a medico-legal setting. In addition to the purely subjective approach used in the appraisal of stressful life events it may be useful to (i) apply common sense and population-based appraisal as an initial basis for assessing the significance of a stressor; (ii) use evidence-based findings to support the link between stressors and disorder; and (iii) apply recognized criteria of causality where applicable. CONCLUSIONS: Conflicting psychiatric opinions in the medico-legal setting may arise both from psychiatrists' positional biases, from the often complex relationships that may exist between stressors and depression, and from a failure to use evidence-based findings to support psychological explanations. PMID- 11270464 TI - A conceptual review of functioning: implications for the development of consumer outcome measures. AB - OBJECTIVE: Australia's National Mental Health Strategy aims to achieve improved consumer outcomes. The development and refinement of consumer outcome measures is targeted within the Second National Mental Health Plan. The National Standards for Mental Health Services identify measures of functioning, quality of life and satisfaction with services as relevant to assessing and monitoring consumer outcome. Consumers have described gauging their own recovery through the achievement of functional goals in everyday life. This paper reviews how functioning is viewed within the mental health field, and implications for developing better functional outcome measures. METHOD: Literature describing the development of measures of functioning, principles of outcome measurement, and functional outcomes for people with severe mental illness was identified, using PsycLIT. A review yielded themes reflecting a number of assumptions about the concept of functioning. RESULTS: Functioning is inadequately defined, raising issues about what is focused on, and from whose viewpoint, each of which has implications for using measures of functioning to monitor consumer outcome. Conflation of dissimilar functional domains, and flawed assumptions about the importance of symptomatology in influencing functional outcome limit the sensitivity to meaningful change of functional measures. Consumer perspectives are relatively neglected in functional tool development. CONCLUSIONS: A conceptual framework that recognizes lived experience and the interaction between persons and their environment is much needed to guide the development of functional outcome measures. Qualitative and quantitative research methodologies should be used to advance understanding of functioning and to address limitations of current approaches to functional outcome measurement. PMID- 11270463 TI - Stalking: new constructions of human behaviour. AB - OBJECTIVE: In the last decade stalking has emerged as a significant social problem, which now constitutes a specific form of criminal offence in most English-speaking nations. This paper examines why stalking has become a major social problem and why it should be of particular concern to mental health professionals. METHOD: Using the extant literature, the history of the emergence of stalking as social, legal and behavioural science discourses is presented. An attempt is made to understand the social and cultural forces which shaped our current understanding of the phenomenon of stalking. RESULTS: Stalking flourishes in a variety of contexts; the social conditions conducive to such behaviour include greater instability in intimate relationships, a culture of blame and entitlement and a growing social anxiety that emphasizes vulnerability to crime and suspicion regarding the intentions of strangers. Stalking is now an established category whose utility is in directing social, legal and health energies to support victims and relieve stalkers of their burden of pursuit. CONCLUSIONS: Stalking is a curious construction born of a range of tensions in contemporary culture but has proved to be a useful label and a useful concept. In part due to the emergence of the concept of stalking, laws are now available to protect, and services increasingly geared to support, the victims of persistent harassment. PMID- 11270465 TI - An analysis of consumer perspectives following contact with an eating-disorders service. AB - OBJECTIVE: The views of consumers following contact with treatment for eating disorders represent an underresearched aspect of service provision. The aim of this paper is to examine patterns of consumer satisfaction following contact with a specialist eating-disorders service. METHOD: Using both a structured and an open-ended questionnaire format, consumer perspectives were sought routinely through postal survey 3 months after the point of first contact. Responses were analysed from 120 patients who returned their questionnaires during the 2-year period ending in December 1998. RESULTS: Although the structured response format indicated high rates of satisfaction, the open-ended format revealed five categories describing the perceived best and worst aspects following consultation with the service. The category of therapeutic alliance drew the majority of positive comments, while the most frequently cited worst aspect of consultation was the category of treatment type. CONCLUSIONS: People with eating disorders form a unique group of mental health consumers to survey for satisfaction. While approval ratings prompted by both structured and open-ended questions were high, and centred around the theme of therapeutic alliance, the most frequent source of negative commentary was activities and structures considered essential by traditional treatment modalities. This provides important insights into the predicaments of people with eating disorders presenting for treatment, and the importance of developing satisfaction surveys to accommodate such predicaments and concerns. PMID- 11270466 TI - A non-comparative trial of the efficacy and safety of fexofenadine for treatment of perennial allergic rhinitis. AB - An open-label, non-comparative study was performed in three Otolaryngology centers in Bangkok, Thailand, to assess the efficacy, safety and tolerability of fexofenadine in Thai patients with perennial allergic rhinitis. Altogether 101 perennial allergic rhinitis patients were included, 33 males and 68 females. Mean age was 33 years, average duration of symptoms was 6 years. All patients received fexofenadine hydrochloride 120 mg once daily (OD) in the morning for 2 weeks. Patients recorded their allergy symptoms daily using a 5 point rating scales in the diary card. At the end of 2 weeks, patients and investigators assessed the overall efficacy of treatment. Adverse events and onset of symptom relief were also recorded by every patient. Blood test and ECG were performed before and after treatment in one center (Siriraj Hospital). Total symptom scores and nasal scores decreased significantly from a baseline at 1 week and 2 weeks after treatment (p < 0.05). The mean onset of symptom relief was 2 hours and 12 minutes. The global assessment of the treatment by patients and investigators showed significant concordance. There was no significant change in either the vital signs, laboratory tests or ECG. The incidence of treatment related adverse events was 8% but all were mild and easily tolerated. Drowsiness was reported from only one patient. This study suggests that fexofenadine 120 mg once daily was an effective, safe and well tolerated treatment for perennial allergic rhinitis in Thai patients. PMID- 11270467 TI - Acceptance of the Accuhaler, a multi-dose powder inhaler, among asthmatic patients: a comparison with the pressurized metered-dose inhaler. AB - This study aimed to evaluate dry powder inhaler naive asthmatic patients' perception and preference of the Accuhaler, a multidose dry powder inhaler and the pressurized metered dose inhaler (pMDI). After the first instruction, 66.7% of 48 patients enrolled in the study could demonstrate the correct use of the Accuhaler. When the patients were asked to compare the pMDI and the Accuhaler after using the Accuhaler to administer salmeterol for 4 weeks, the Accuhaler scored significantly better than the pMDI for the following features: knowing how many doses are left, presence of an attached cover, taste, instruction for use, attractiveness, ease of use, ease of holding, shape, and comfortable mouthpiece. The pMDI scored better to the Accuhaler in terms of size. More patients preferred the Accuhaler than the pMDI; the presence of a dose counter and perceived ease of use were the main reasons cited for their preference for the Accuhaler. PMID- 11270468 TI - Clinical studies on bronchial asthma caused by contact with hamsters. AB - Bronchial asthma induced by contact with hamsters and other small rodents is receiving higher attention from the medical profession not only because of the problem of laboratory animal allergies (LAA), but also because of increasing household allergens for asthma, since keeping these pets has become more common in Japanese homes. The present report describes our studies on the backgrounds of nine patients with asthma who kept Dzungarian Dwarf hamsters as household pets. The following features were recognized among patients with bronchial asthma induced by contact with hamsters: 1) earlier onset of symptoms than for keeping other household pets, at an average of 14.7 months or within 12 months in 78% of the cases following the start of pet keeping; 2) adults ranging from their late 30s to 40s who have children of primary school age; 3) dwelling in apartments; 4) relatively high level of serum IgE and ECP; 5) positive for both immediate and late type asthmatic responses on an inhaling induction test; and 6) rapid remission after the cessation of pet keeping. PMID- 11270469 TI - Buckwheat allergy and reports on asthma and atopic disorders in Taiyuan City, Northern China. AB - Allergy to common buckwheat (Fagopyrum esculentum) has been reported from Europe and Japan, and a 24 kDa globulin protein has been identified as one of the major allergens. In China also another type, tartary buckwheat (Fagopyrum tartaricum) is grown and consumed. Three groups of individuals in Shanxi province, China, were investigated for buckwheat allergy using skin prick test. The groups were: agricultural researchers with occupational exposure to buckwheat (N = 16); workers in a food industry producing buckwheat noodles (N = 25), and patients with diabetes or cardiovascular disease consuming buckwheat as functional food (N = 20). Information on atopic disorders and adverse food reactions were collected by a doctors-administered questionnaire. One male industrial worker had a positive skin prick test to buckwheat, but no symptoms while eating or handling buckwheat products. In total, 34% consumed buckwheat food at least every week, and 23% had a weekly consumption of tartary buckwheat. The prevalence of doctor's diagnosed asthma was low (1.6%). Four subjects (6.6%) reported a history of allergic rhinitis, with allergy to cedar pollen, carnation and peach. PMID- 11270470 TI - Evaluation of three methods for using the Duotip-Test device for skin testing. AB - The sensitivity and precision of rotation, prick and puncture methods of using the Duotip-Test for epicutaneous allergy skin testing were evaluated. Forty-one volunteers who had not taken any antihistamines within the previous two weeks were recruited. The mean age was 21.6 years (range 18 to 25 years). Histamine hydrochloride 1 mg/ml and 50% glycerol saline were used as positive and negative controls, respectively. Each method of testing was performed in triplicate on the volar surface of both forearms. Wheal and flare were measured 15 minutes later. Rotation, prick and puncture methods produced histamine mean wheal diameter +/- standard deviation of 6.61 +/- 0.87 mm, 3.86 +/- 1.03 mm, and 3.00 +/- 0.65 mm, respectively (p < 0.01). The coefficient of variation of rotation method was 13.13%. It was the only method that gave coefficient of variation lower than 20%. False negative and false positive proportions of rotation method using a 4 mm criterion for positive reaction were 1.5% and 0.75%, respectively. Rotation method was well accepted by the volunteers although it was ranked highest in pain. We concluded that the rotation method of using Duotip-Test is a highly reliable technique for skin testing. PMID- 11270471 TI - Comparison of in vitro assay for specific IgE and skin prick test with intradermal test in patients with allergic rhinitis. AB - Among many diagnostic tests for allergic rhinitis, the intradermal (ID) test is practical and reliable. However, there are several factors affecting compliance, practicability and interpretation, and also problems on hypersensitivity of the ID. For these reasons, we evaluated other tests which have been thought to have high reliability as diagnostic and/or screening assays, namely, skin prick test and specific IgE detection in seventy-four perennial rhinitis patients (51 males and 23 females whose ages were between 15-60 years). In this study, Dermatophagoides pteronyssinus and D. farinae extracts, known to be the most common aeroallergens in Thailand, were used as the allergens/antigens. Compared to the standard ID test, sensitivities to D. pteronyssinus and D. farinae of the studied patients tested by skin prick test were 90.4% and 86.4%, and specificities were 99.5% and 93.1%, respectively. Sensitivities to D. pteronyssinus and D. farinae using specific IgE assay were 96.3% and 88.9%, and specificities were 96.2% and 88.9%, respectively. It was concluded that the skin prick test can be used as a screening method for patients with allergic rhinitis, while the specific IgE detection can be used as an alternative for diagnosis of patients who are susceptible to the ID test or for those who are severely susceptible to allergic rhinitis such that medication can not be withdrawn for the ID test. PMID- 11270472 TI - Parvovirus B19 antibodies in immunocompromized children in Thailand. AB - Parvovirus B19, a non-enveloped single stranded DNA virus is distributed worldwide. Sero-prevalence in adult populations amounts to approximately 50%. Clinical manifestations vary depending on the Immune status of the infected individuals and may include mild childhood Infection as well as hydrops fetalis due to intrauterine infection. To determine the prevalence of this infection among the immunocompromized individuals in Thailand, we determined, by indirect ELISA, levels of IgM and IgG antibodies to the parvovirus B19 in 106 immunocompromized children. These included 49 children who were on chemotherapy for treatment of malignancies, 18 who were receiving immunosuppressive drugs after organ transplantations, 14 who were under a regimen of corticosteroids and 25 who were positive for antibodies to HIV. The average prevalence of IgG antibodies in 106 children was 16.0%; the prevalence of antibodies was 33.3% in post-transplanted group, 16% in children positive for HIV, 12.2% in the group receiving chemotherapy for malignancies and 7.6% in the group treated with corticosteroids. All children were negative for IgM antibodies to parvovirus B19. PMID- 11270473 TI - X-linked hyper IgM syndrome: a report of the first case in Thailand with a confirmed mutation of CD40 ligand gene. AB - X-linked hyper IgM (XHIM) syndrome is a rare congenital immunodeficiency disease caused by failure of B cell to isotype switch from IgM to other classes of immunoglobulins in response to infections. Recently, a molecular cloning of the gene responsible for the syndrome, the CD40L gene has been accomplished and the gene was successfully mapped to the long arm of X chromosome at the position Xq26. We, herein, report the first case of molecular proven XHIM in a Thai boy with a classic presentation and with a confirmed mutation of the CD40L gene. CASE REPORT: A.S. was a 1 year 7 month old boy referred from Buriram Provincial Hospital for a work up and treatment for his recurrent infections consisted of chronic respiratory tract infections with otitis media (since 6 months of age), chronic diarrhea (since 9 months of age) and malnutrition (marasmus) secondary to his longstanding illnesses. He was a product of a consanguineous marriage but without history of similar illness observed in his pedigree. Abnormal laboratory works up included IgG of 300 mg/dl, IgA 10 mg/dl, IgM 1,635 mg/dl, positive stool examinations for Cryptosporidium, chronic colitis on radiographic gastrointestinal follow through study, a positive acid fast bacillus (AFB) stain of gastric aspirate and multiple positive bacterial cultures from various body sources. His anti-HIV serology was negative. His hospital course was significant for several bouts of infections of gastrointestinal, respiratory, and genitourinary systems. His treatment consisted of multiple courses of antibiotics, antituberculous drugs and IVIG administrations. His hospital course was complicated with feeding problem from an esophageal stricture requiring several esophageal dilatations. The analysis of CD40L gene revealed a point mutation of exon 5 (A619T) of the CD40L gene resulting in a stop codon confirming that indeed he had XHIM. He died with Pseudomonas septicemia during the waiting period for a bone marrow transplantation from a cord-blood stem cell. PMID- 11270474 TI - Histamine induced decrease of lecithin levels in broncho-alveolar lavage fluid of rats is mediated by H2 receptor. AB - Lecithin, a major surface active substance of the surfactant system of the lung, was estimated in broncho-alveolar lavage (BAL) fluid in four groups of healthy adult male albino rats. Rats from group I were not administered any drug and acted as controls. Group II were administered histamine diphosphate. Group III were given H1 blocker (pyrilamine maleate) followed by histamine diphosphate. Group IV received H2 blocker (ranitidine hydrochloride) followed by histamine diphosphate. Lecithin content of BAL fluid in the control group was compared with that in the other three groups. A significant decrease in lecithin content was observed in the rats that received either histamine diphosphate or H1 blocker followed by histamine diphosphate. However, compared to control rats no significant difference in lecithin content was seen in rats that received H2 blocker followed by histamine diphosphate. The results clearly indicate that the decrease in surface active lecithin content in BAL fluid following administration of histamine diphosphate was unaffected by prior administration of H1 blocker, but was blocked by prior administration of H2 blocker. It was concluded that histamine induced decrease in lecithin content of BAL fluid is mediated through H2 receptors. Since the predominant source of intra-alveolar lecithin are Type II cells of the alveolar epithelium, It is possible that Type II cells have H2 receptors, stimulation of which resulted in decreased intraalveolar lecithin. PMID- 11270475 TI - Prevalence and clinical relevance of serum anti-p53 antibodies in patients with cholangiocarcinoma. AB - Cholangiocarcinoma (CCA) constitutes carcinoma of the bile duct found at a high prevalence in northeastern Thailand. In the present study, we examined the sera of altogether 82 Thai CCA patients for the presence of anti-p53 antibodies in order to investigate a role of the tumor suppressor gene, p53 in the carcinogenesis. Our results revealed anti-p53 antibodies in 7.3% of the cases tested, which conforms to the prevalence rate of p53 gene mutation recently reported at 5% among Thai patients. With limited number of the patients, anti-p53 antibodies were rapidly detected more frequently among patients with peripheral tumors than those with central tumors. However, further studies is required to establish significance and prognostic value of the antibodies in the context of CCA. PMID- 11270476 TI - Evaluation of microalb immunoturbidimetric test for albuminuria screening. AB - Microalb, an immunoturbidimetric test for screening urinary albumin levels was evaluated for its potential as a screening test for microalbuminuria in diabetic patients. It was compared with the current standard, the radioimmunoassay (RIA). The results showed that the test lacks sensitivity while its specificity was acceptable. Therefore, it can not replace the RIA as the screening method. This study also showed that the first early morning urine could be used if a 24- hour collection was not possible, as its albumin content fairly correlated (r = 0.78) with the 24-hour urine collection of the diabetic patients. PMID- 11270477 TI - Evidence for a direct effect of melatonin on mitochondrial genome expression of Siberian hamster brown adipocytes. AB - Photoperiod variations are known to participate in the regulation of energy balance in different rodent species via melatonin, a neurosecretory product synthesized by the pineal gland during the night. A direct effect of melatonin on adipose tissue has been suggested since binding sites for the indole have been described on brown adipocytes. The aim of this study was to investigate a genetic effect of melatonin on isolated Siberian hamster brown adipocytes using differential display RT-PCR (DDRT-PCR). Brown adipose cells were isolated from brown adipose tissue and treated for 3 hr with 0.1 and 10 microM melatonin. Total RNA was extracted and DDRT-PCR experiments were performed. A differential band, which disappeared after melatonin treatment, was detected. After confirmation and cloning, the corresponding cDNA fragment B18 was sequenced. B18 had 85 and 81% similarity with a portion of rat and mouse cytochrome b mRNA, respectively, suggesting that B18 corresponds to hamster cytochrome b. This hypothesis was confirmed by the close parallel between the changes in mRNA content, detected by B18, and by cytochrome b mRNA content, detected by a rat probe. Cytochrome b mRNA is encoded by the mitochondrial genome, suggesting a similar effect of melatonin on the whole mitochondrial transcripts. Indeed, 3 hr of treatment with melatonin (10 nM and 0.1 microM) decreased by 44% mitochondrial transcript contents. This work constitutes the first evidence of a direct biological effect of melatonin on Siberian hamster brown adipocytes. PMID- 11270478 TI - Melatonin counteracts potentiation by lysophosphatidylcholine of serotonin induced vasoconstriction in human umbilical artery: relation to calcium influx. AB - We evaluated mechanisms underlying the antioxidant property of melatonin as related to vasospastic effects in the human umbilical artery from lysophosphatidylcholine (LPC), a component of oxidized lipoprotein. Helical sections of umbilical artery without endothelium were obtained at elective cesarean delivery between 37 and 39 wk of gestation. Changes in 5 hydroxytryptamine (5-HT)-induced vasoconstriction were measured. Arterial sections were treated with LPC (15 or 30 microM) alone or pretreated with a hydroxyl radical (.OH) scavenger, mannitol (20 mM), an H2O2 scavenger, catalase (1,200 U/mL), or melatonin (0.1 or 1.0 microM). The effect of LPC on the response of arterial sections to external calcium in the presence of KCl (20 mM) was determined. LPC potentiated 5-HT-induced vasoconstriction in a concentration dependent manner; pretreatment with mannitol or catalase significantly reduced this vasospastic effect. LPC (30 microM) significantly augmented the contractile response to external calcium. Melatonin (1.0 microM) pretreatment significantly suppressed the contractile response to external calcium. The results suggest that LPC potentiates 5-HT-induced umbilical artery vasoconstriction, in part by increasing the calcium influx into smooth muscle cells via activation of voltage dependent calcium channels. Given a previous finding, the vasospastic effect of LPC on the umbilical artery also appears to involve the suppression of endothelial nitric oxide production. Melatonin significantly suppresses the vasospastic effect of LPC, probably by scavenging .OH arising from LPC. PMID- 11270479 TI - Thrombopoietic properties of 5-methoxytryptamine plus melatonin versus melatonin alone in the treatment of cancer-related thrombocytopenia. AB - Recent studies have shown that the hematopoietic system is under neuroendocrine control. In particular, thrombopoiesis has been proven to be stimulated by melatonin, and the pineal indole has been shown to be effective in the treatment of thrombocytopenia resulting from different causes. At present, however, there are no data concerning the possible thrombopoietic activity of pineal indoles other than melatonin. The present study was carried out to evaluate the effect of a concomitant administration of the pineal indole 5-methoxytryptamine in patients with cancer-related thrombocytopenia who did not respond to melatonin alone. The present study included 30 patients, who were randomized to receive melatonin alone (20 mg/day orally in the evening) or melatonin plus 5-methoxytryptamine (1 mg/day orally in the early afternoon). A normalization of platelet count was achieved in 5/14 (36%) patients treated with melatonin plus 5-methoxytryptamine and in none of the patients treated with melatonin alone (P < 0.05). Moreover, mean platelet number significantly increased only in the patients treated with melatonin plus 5-methoxytryptamine. This preliminary clinical study would suggest that 5-methoxytryptamine, a pineal indole, may also exert thrombopoietic activity. Further studies, however, will be required to establish whether 5 methoxytryptamine may play a direct thrombopoietic activity, or whether it may act by improving melatonin's efficacy. PMID- 11270480 TI - Cholelithiasis, oxidative stress and melatonin. PMID- 11270481 TI - Melatonin, mitochondria, and cellular bioenergetics. AB - Aerobic cells use oxygen for the production of 90-95% of the total amount of ATP that they use. This amounts to about 40 kg ATP/day in an adult human. The synthesis of ATP via the mitochondrial respiratory chain is the result of electron transport across the electron transport chain coupled to oxidative phosphorylation. Although ideally all the oxygen should be reduced to water by a four-electron reduction reaction driven by the cytochrome oxidase, under normal conditions a small percentage of oxygen may be reduced by one, two, or three electrons only, yielding superoxide anion, hydrogen peroxide, and the hydroxyl radical, respectively. The main radical produced by mitochondria is superoxide anion and the intramitochondrial antioxidant systems should scavenge this radical to avoid oxidative damage, which leads to impaired ATP production. During aging and some neurodegenerative diseases, oxidatively damaged mitochondria are unable to maintain the energy demands of the cell leading to an increased production of free radicals. Both processes, i.e., defective ATP production and increased oxygen radicals, may induce mitochondrial-dependent apoptotic cell death. Melatonin has been reported to exert neuroprotective effects in several experimental and clinical situations involving neurotoxicity and/or excitotoxicity. Additionally, in a series of pathologies in which high production of free radicals is the primary cause of the disease, melatonin is also protective. A common feature in these diseases is the existence of mitochondrial damage due to oxidative stress. The discoveries of new actions of melatonin in mitochondria support a novel mechanism, which explains some of the protective effects of the indoleamine on cell survival. PMID- 11270482 TI - Effect of propranolol plus exercise on melatonin and growth hormone levels in children with growth delay. AB - The pineal gland in humans is under both alpha- and beta-adrenergic control, although it seems that beta1-adrenoceptors are mainly implicated in melatonin secretion. In the present study, we evaluated the role of beta-adrenergic innervation on melatonin production and its relation with the production of growth hormone (GH). Thirty-four children (15 males and 19 females, mean age 10.5 +/- 0.8 years) from the University of Granada Hospital were studied. The children were included in a protocol for the evaluation of growth delay using the propranolol + exercise test. This standardized test allowed us to study simultaneously the role of an unspecific beta-adrenergic blocker such as propranolol and of an adrenergic stimulus such as exercise on the pineal production of melatonin. Changes in plasma levels of melatonin and GH were determined at basal, 120 and 140 min after the test was applied. Hormonal determinations were carried out by commercial radioimmunoassay kits previously standardized in our laboratory. The results show a significant decrease in plasma melatonin levels at 120 and 140 min after the test (P < 0.05), whereas GH levels increased significantly at 140 min (P < 0.001). The decrease of melatonin levels was a consequence of the test, since in a control group, the circadian decay of melatonin was significantly less pronounced (P < 0.05). These data suggest an inverse relationship between melatonin and GH after the propranolol + exercise test, and the reduction in melatonin may be related to its depletion by exercise induced oxidative stress. PMID- 11270483 TI - Roles of nocturnal melatonin and the pineal gland in modulation of water immersion restraint stress-induced gastric mucosal lesions in rats. AB - The roles of melatonin and the pineal gland in the circadian variation of water immersion restraint stress-induced gastric mucosal lesions in rats were investigated. Fasted rats were subjected to water-immersion restraint stress during both the diurnal and nocturnal phases of a light:dark cycle. Pinealectomized and sham-operated rats were also subjected to water-immersion restraint stress at night. The lesion area after 4 hr of stress during the dark phase was significantly lower than in light-phase controls. Pinealectomy increased the lesion area in the dark phase, compared to the sham operation, but this effect was counteracted by intracisternal melatonin preadministration at a dose of 100 ng/rat. Melatonin concentrations in control rats during the light phase were significantly increased 4 hr after water-immersion restraint stress. In contrast, melatonin concentrations 4 hr after water-immersion restraint stress in the dark phase were significantly depressed compared with the control levels at the corresponding time. Melatonin levels after stress exposure were markedly decreased in pinealectomized rats as compared with sham-operated rats. These results suggest that circadian rhythm has an important role in the formation of stress-induced gastric mucosal lesions in rats and that melatonin responses to water-immersion restraint stress differ between day and night. The pineal gland modulates the stress response and melatonin contributes to gastric protection via a mechanism involving the central nervous system. PMID- 11270484 TI - The modulation of neuronal activity by melatonin: in vitro studies on mouse hippocampal slices. AB - The influence of melatonin on evoked potentials recorded from the CAI field of mouse hippocampal slices was investigated. Melatonin (0.1-2.0 mM) and its analog, 6-chloromelatonin (0.1-0.5 mM) depressed evoked potentials (EPSP and the population spike) in a concentration-dependent manner. The melatonin-induced depression was followed by a slow recovery phase. Since the fiber potential was not affected, it was concluded that melatonin influenced synaptic efficiency and/or cell excitability. Luzindole, an antagonist of MT2 melatonin receptors, although slightly depressing evoked potentials when applied by itself (100 microM), blocked any further inhibition by melatonin when added afterwards. We concluded that melatonin reduced synaptic efficiency and/or excitability of hippocampal neurons most likely through interaction with MT2 melatonin receptors, but other possible mechanisms of melatonin action are also considered. PMID- 11270485 TI - A test of the coincidence and duration models of melatonin action in Siberian hamsters: the effects of 1-hr melatonin infusions on testicular development in intact and pinealectomized prepubertal Phodopus sungorus. AB - The pineal hormone melatonin is known to play an important role in mediating photoperiodic messages to the reproductive system in seasonal breeding animals. Our goal was to test, in a single experimental paradigm, two hypotheses that have been forwarded to describe how the circadian rhythm of pineal melatonin transmits photoperiodic information to the reproductive system: 1) induction, i.e., a short day effect, occurs when secreted melatonin and a circadian rhythm of sensitivity to melatonin coincide in time; 2) induction occurs following exposure to elevated circulating melatonin levels for a prescribed duration. In order to determine the relative validity of these hypotheses, we investigated the testicular maturation response to 1-hr daily infusions of 10, 25, and 50 ng of melatonin in pinealectomized intact and prepubertal Siberian hamsters (Phodopus sungorus). Animals received, beginning on day 15 of life, programmed subcutaneous infusions of melatonin or vehicle at one of five time points (19:00-20:00, 20:00-21:00, 21:00-22:00, 24:00-01:00, and 03:00-04:00 hr) for 15 days. In animals gestated and raised in a long photoperiod (LD16:8 = 16L, where L is the duration of light in hours, and D that of dark), melatonin infusion right after lights off (20:00 21:00 hr) significantly retarded gonadal maturation; this dose was ineffective at other times tested. Doses of 10 and 25 ng melatonin were ineffective at all time points. Identical results were obtained in prepubertal hamsters gestated in a short photoperiod (LD10:14 = 10L) and raised in 16L; these results were independent of the presence or absence of the pineal gland. In animals gestated and raised in 10L, melatonin infusions failed to suppress testicular development beyond that induced by the photoperiod; testicular development was maximally suppressed in all groups. The results of these investigations are best explained under the experimental conditions employed here: 1) the photoperiodic gonadal response in juvenile Siberian hamsters is regulated by the coincidence in time of exogenously administered melatonin with an intrinsic rhythm of sensitivity to melatonin, which, under the constraints imposed by our experimental design, occurred at 20:00-21:00 hr; and 2) the duration of the melatonin signal alone, equal in all groups, cannot explain the results. PMID- 11270486 TI - Vesicular storage of adenosine triphosphate in the guinea-pig cochlear lateral wall and concentrations of ATP in the endolymph during sound exposure and hypoxia. AB - Previous studies have revealed putative vesicular stores of adenosine triphosphate (ATP) in the marginal cells of the cochlear stria vascularis which may serve as a source of ATP for purinergic signalling. This study aimed to provide further evidence of ATP storage in the cochlea and to see whether ATP levels in the endolymph are affected by noise and hypoxia. Tissues from the lateral wall and organ of Corti of the guinea-pig cochlea were fractionated to obtain vesicular (VF) and mitochondrial (MF) fractions. Free and total ATP were then measured by the luciferase-luciferin reaction from which membrane-bound vesicular ATP was calculated. In the lateral wall, the VF contained 2.02+/-0.04 nmol ATP/mg protein (n = 5), significantly greater (p < 0.001; paired Student's t test) than the concentration of ATP in the MF (0.36+/-0.05). In the organ of Corti, the VF contained 0.69+/-0.08 nmol ATP/mg protein (n = 4), significantly smaller than the amount in the VF of the lateral wall tissues (p < 0.001; non paired Student's t-test). Small amounts of fumarase. an enzyme of the mitochondrial matrix, in the VF, excluded the possibility of mitochondrial ATP contamination. To investigate the effect of hypoxia and noise on the ATP concentrations in the endolymph, fluid samples were collected from the first (basal) cochlear turn of anaesthetized guinea-pigs. As a result of hypoxia (15 min, 13% F1O2), ATP concentrations (nM, mean +/- SEM) increased from 6.2+/-2.3 to 9.3+/-4.5 (n = 4), but the difference was not statistically significant. As a result of noise (15 min, 10 kHz, 110 dB SPL. broad band), the ATP levels increased significantly from 7.4+/-1.2 to 16.0+/-1.8 (p = 0.01; Student's t-test: n = 4). This study has demonstrated the presence of a vesicular store of ATP in the stria vascularis of the cochlea and described an increase in the ATP levels in the endolymph during noise exposure. The findings suggest that ATP is actively secreted from the vesicular store under conditions of metabolic stress. The presence of ATP under basal conditions supports a role for ATP in the sound transduction process during normal function. PMID- 11270487 TI - 1,25(OH)2 vitamin D3 induces elevated expression of the cell cycle-regulating genes P21 and P27 in squamous carcinoma cell lines of the head and neck. AB - The biologically active form of vitamin D3, 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], inhibits proliferation and induces differentiation for various malignant cells, including squamous cell carcinoma cell lines of the head and neck (SCCHN). These effects are due to an arrest of cells in the G0/G1 phase of the cell cycle and are predominantly mediated by the vitamin D receptor. To further explore the molecular mechanisms of the antiproliferative activity in SCCHN we studied the influence of 1,25(OH)2D3 on the expression of the G1 phase regulating proteins cyclin D1, p21 and p27. Furthermore, as a direct target of G1 protein complexes, we investigated the phosphorylation status of the retinoblastoma protein (pRb). Synchronized cells of 2 SCCHN cell lines [JPPA (laryngeal carcinoma) and SCC 9 (tongue carcinoma)] and human immortalized keratinocytes (HaCaT) were cultured for 96 h in the presence or absence (ethanol as control) of 1,25(OH)2D3 (10(-7) M). At various time intervals the cell cycle status was detected by fluorescence-activated cell sorting (FACS) analysis and in parallel the expression of cell cycle-regulating proteins was determined at the protein and mRNA levels. In all cell lines tested 1,25(OH)2D3 caused an arrest of cells in the G0/G1 phase of the cell cycle and markedly induced the expression of the inhibitors p21 and p27. No influence was detectable on the expression of cyclin D1. Induction of p21 and p27 mRNA revealed transcriptional regulation by the vitamin D receptor. Simultaneously, hyperphosphorylated pRb was transformed to the hypophosphorylated form. Our results demonstrate that the biologically active form of vitamin D3 directly regulates the expression of p21 and p27, inducing a G0/G1 phase arrest: one mechanism by which 1,25(OH)2D3 controls cell proliferation inSCCHN. PMID- 11270488 TI - Pharmacological models for inner ear therapy with emphasis on nitric oxide. AB - Nitric oxide (NO)-mediated neurotoxicity may be an appropriate pathophysiological model with which to explain a variety of inner ear diseases characterized by acute or progressive hearing loss, tinnitus and vertigo. The localization of NO synthase (NOS) isoforms was examined in the inner ear of the pigmented guinea pig after intratympanic injection of 1 mg lipopolysaccharide (LPS) or 5 mg gentamicin (GM) using an immunohistochemical method, revealing the expression of NOS II in the inner ear. Production of NO in the isolated organ of Corti and utricle or in the isolated vestibular and cochlear hair cells after stimulation with L arginine, glutamate, GM and LPS was investigated using the fluorescence indicator 4,5-diaminofluorescein diacetate. The fluorescence intensity of the sensory cells was augmented by stimulation with L-arginine, glutamate, GM and LPS. A significant increase in NO production was also noted in the LPS-treated animals. These findings imply that NO from constitutive NOS may mediate ototoxicity in the early phase, whereas NO from NOS II may contribute to the late phase of tissue damage in the inner ear. Based on this hypothesis, reduction of glutamatergic excitotoxicity and inhibition of NOS, scavenging superoxide and scavenging peroxynitrite are thought to attenuate NO-mediated otoneurotoxicity. PMID- 11270489 TI - Short-term adaptation in the peripheral auditory system is related to the AMPA receptor. AB - The role of glutamate receptors was investigated by infusing N-methyl-D-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methyl-isoxazol-propionate (AMPA) into the guinea pig cochlea. Auditory brainstem response thresholds and forward masking were used to determine auditory sensitivity. In the presence of 330 microM NMDA, the auditory brainstem response (ABR) thresholds were elevated by 20-30 dB at 2 kHz and 8 kHz, and the slopes of the forward masking curves were not significantly different from controls. When a high concentration of NMDA (15 mM) was used, ABR thresholds were elevated by 40-50 dB at 2 kHz and 8 kHz and the slopes of the forward masking curves were significantly decreased. In contrast, when AMPA (150 microM) was infused, ABR thresholds were elevated by 20-35 dB at 2 and 8 kHz and the slopes of the forward masking curves were significantly decreased from the control group. When the concentration of AMPA was decreased (100 microM). ABR thresholds were not significantly altered but the slopes of the forward masking curves were significantly decreased from control values. The present study suggests that AMPA receptors play a significantly more important role in short-term adaptation than NMDA receptors. PMID- 11270490 TI - Inner ear autoantibodies and their targets in patients with autoimmune inner ear diseases. AB - Immunological mechanisms are thought to play an important role in the pathogenesis of some cochleo-vestibular diseases. This study attempts to present further evidence of autoantibodies reactive against guinea pig inner ear proteins found in patients with autoimmune inner ear diseases (AIED) and specifically identifies the main target antigens of these antibodies. Sera from 110 patients with a clinical diagnosis of either rapidly progressive sensorineural hearing loss (n = 32). Meniere's disease (n = 41), sudden deafness (n = 6) or other aetiologies of hearing loss (n = 11) were screened by the Western blot technique. Forty-four percent of the patients' sera had antibodies to several inner ear proteins, of which the 30, 42 and 68 kDa proteins were found to be the most reactive. These highly reactive proteins were identified by gas-phase micro sequencing after digestion with trypsin and separation of peptide fragments by high-performance liquid chromatography. A partial sequence of each protein was determined. These data, together with those obtained from 2-dimensional gel electrophoresis followed by Western blotting, demonstrated that the 30 and 42 kDa inner ear proteins are the major peripheral myelin protein P0 and the beta-actin protein, respectively, while sequence analysis indicated that the 68 kDa protein is novel. These findings further support the hypothesis that several populations of antibodies may contribute to the enhanced immunological activity of AIED patients. They also add a new dimension to our knowledge of AIED and may open new avenues in the development of simple serological assays, which are easier to perform and more rapid than Western blotting. PMID- 11270491 TI - Vascular variations of the inner ear. AB - Vascular anomalies of the inner ear have been documented in only a few isolated case reports. The goal of our study was to describe, qualify and quantify vascular variations of the inner ear in 122 temporal bones from 64 pediatric subjects aged between 0 and 10 years. The average age was 11.6 months. Horizontal sections of the temporal bone, examined by light microscopy, revealed vessels coursing freely through the perilymphatic space of the cochlea, especially in the apical turn. Other findings included abnormally wide vessels in the stria vascularis as well as a vascular malformation of the internal auditory canal. Our study demonstrated more atypical vessels in the cochlea than in the vestibular labyrinth. We found a statistically significant positive correlation between vascular variations of the inner ear and concomitant cardiac anomalies or endolymphatic hydrops. We also discuss the possible etiology and potential significance of these findings in terms of disturbances of the function of the inner ear. PMID- 11270492 TI - Unpredictable hearing loss after intratympanic gentamicin treatment for vertigo. A new theory. AB - A new hypothesis is advanced suggesting that unpredictable cases of profound hearing loss after intratympanic gentamicin treatment (IGT) may be caused by decreased patency of the communication routes between the inner ear and the cerebrospinal fluid, primarily of the cochlear aqueduct. A tympanic displacement analyzer, which can indirectly analyze inner ear and intracranial pressure changes and can also evaluate the efficiency of communication between these two compartments, was used. Two cases are presented: in the first, a patient who became deaf after IGT showed signs of decreased patency of the communication routes with the tympanic membrane displacement (TMD) test; in the second, a patient without hearing damage after IGT had efficient communication evaluated by the TMD test. These preliminary findings are in accordance with the proposed pathophysiology. If future clinical studies confirm the present theory and findings, it may prove possible to predict and prevent deafness after IGT and possibly also after systemic aminoglycoside treatment. PMID- 11270493 TI - Detection of mucin gene expression in normal rat middle ear mucosa by reverse transcriptase-polymerase chain reaction. AB - Hypersecretion of mucin is a common feature of chronic and mucoid otitis media which may play an important role in hearing loss. The mechanisms controlling mucin secretion in the middle ear are not completely understood. Our reverse transcriptase-polymerase chain reaction results demonstrate that mRNAs of MUC1, MUC2, MUC3, MUC4 and MUC5AC are expressed in normal rat middle ear mucosa. Moreover, the expression of mRNA of the secretory mucins MUC2, MUC3 and MUC5AC was threefold lower in normal middle ear mucosa than that in the intestine or trachea. In contrast, expression of the membrane-bound mucins MUC1 and MUC4 was approximately the same in both middle ear mucosa and the intestine or trachea. MUC5AC proteins were also identified immunohistochemically in normal rat middle car epithelium. The methodology used in this study provides useful baseline information for investigation of the mechanisms of regulation of mucin gene expression during otitis media. PMID- 11270494 TI - Evidence-based surgery in chronic rhinosinusitis. AB - Considerable interest has been focused in recent years on an evidence-based approach to clinical medicine. For obvious reasons, however, it has proved difficult to examine the benefit of surgical procedures in randomized, double blind. placebo-controlled trials. This review considers the evidence available in the literature for surgery in chronic rhinosinusitis and examines the validity of the studies in the context of evidence-based medicine. PMID- 11270495 TI - Vestibulopathy and age effects on head stability during chair rise. AB - It is unknown how vestibular dysfunction and age differentially affect balance control during functional activities. The objective of this study was to gain insight into the effects of age and vestibulopathy on head control when rising from a chair. Head relative to trunk (head-on-trunk) sagittal plane angular and linear control strategies were studied in patients with bilateral vestibular hypofunction (BVH) and in healthy subjects aged 30-80 years. A two-way analysis of variance was used to compare head-on-trunk kinematics by age (young vs elderly) and diagnosis (healthy vs BVH) at the time of liftoff from the seat. Angular control strategies differed with age but not diagnosis: young (healthy and BVH) subjects stabilized head rotations in space while elderly (healthy and BVH) subjects stabilized head rotations on the trunk. In contrast, linear control strategies differed by diagnosis but not age: BVH subjects (young and old) allowed a greater rate of head-on-trunk translation while healthy subjects (young and old) inhibited such translations. Young BVH subjects stabilized head-in-space rotations (as did young healthy subjects) without a functioning vestibular system, suggesting cervicocollic reflex and/or other sensory compensation for vestibular loss. Elderly BVH subjects stabilized head rotation with respect to the trunk, as did healthy elders, but did not stabilize head-on-trunk translations, suggesting a reliance on passive mechanical responses of the neck to sense head movements. We conclude that compensation strategies used by patients with vestibulopathy are age-dependent and appear to be more tractable in the younger BVH patient. PMID- 11270496 TI - Quality of life of vestibular schwannoma patients after surgery. AB - The quality of life of vestibular schwannoma (VS) patients after surgery was investigated. The subjects consisted of 236 unilateral VS patients who underwent tumor removal between 1990 and 1997. A questionnaire was sent to all patients regarding their hearing, tinnitus, dizziness and the changes in their daily life after surgery; 176 out of 204 patients (86%) who received the questionnaire completed and returned it. The answers were compared with recent data reported in other clinical studies. Ninety percent of the patients with postoperative class A hearing were satisfied with their hearing. However, only 30% of patients with postoperative class B hearing were satisfied. Tinnitus worsened after surgery more often in patients who underwent a labyrinthectomy than in those who did not. Dizziness improved after surgery in the majority of VS patients. However, 30% of patients had difficulty driving a car and 50% of patients could not enjoy activities such as playing sport after surgery. PMID- 11270497 TI - The vestibular evoked response to linear, alternating, acceleration pulses without acoustic masking as a parameter of vestibular function. AB - In this study, short latency vestibular evoked potentials (VsEPs) were recorded in five guinea pigs in response to alternating linear acceleration pulses with and without acoustic masking. A steel bolt was implanted in the skull and coupled to a shaker. Linear acceleration pulses (n = 400) in upward, downward or alternating directions were given, with a peak acceleration of 4g after 0.5 msec. Tests were repeated with acoustic masking, after modiolus destruction and after application of KCl in the vestibule. Stimuli of the vestibular nerve were recorded with a platinum electrode in the bony facial nerve canal in the bulla. Unilateral linear acceleration showed a shallow plateau at 0.5 msec, which disappeared with alternating acceleration impulses and after modiolus destruction. Therefore all further tests were done with alternating impulses. After a latency time of 0.8 msec a multiwave response was seen, with a first positive peak P1 at 1.16 ms. These were followed by other positive and negative peaks (N1, P2, N2, P3, N3). With the elimination of cochlear influences by using acoustic masking, P1 remained stable, while subsequent peaks were altered or eliminated. After modiolus destruction, the P1 peak remained, although with a smaller amplitude due to vestibular damage. After application of a saturated KCl solution in the vestibule all responses, including P1, disappeared, thus confirming the vestibular origin of these responses. We conclude that the onset latency of the VsEP and the peak latency and level of the first positive peak P1 in response to alternating linear acceleration pulses without acoustic masking, measured in the facial canal, are good and stable parameters of vestibular function in guinea pigs. PMID- 11270498 TI - Symptoms, findings and treatment in patients with dehiscence of the superior semicircular canal. AB - Recently Minor and co-workers described patients with sound- and pressure-induced vertigo due to dehiscence of the superior semicircular canal. Identifying patients with this 'new' vestibular entity is important, not only because the symptoms are sometimes very incapacitating, but also because they can be treated. We present symptoms and findings in eight such patients, all of whom reported pressure-induced vertigo that increased during periods of upper respiratory infections. Pulse-synchronous tinnitus and gaze instability during head movements were also common complaints. All patients lateralized Weber's test to the symptomatic ear. In some of the patients the audiogram also revealed a small conductive hearing loss. However, the stapedius reflexes were always normal. A vertical/torsional eye movement related to the superior semicircular canal was seen in most of the patients in response to pressure changes and/or sound stimulation. One patient also had superior canal-related positioning nystagmus. Testing vestibular evoked myogenic potentials revealed in all patients a vestibular hypersensitivity to sounds. In the coronal high-resolution 1-mm section CT scans the dehiscence was visible on 1 to 4 sections. Moreover, the skull base was rather thin in this area and cortical bone separating the middle ear and the antrum from the middle cranial fossa was absent in many of the patients. Two of the patients have undergone plugging of the superior semicircular canal using a transmastoid approach and both patients were relieved of the pressure-induced symptoms. PMID- 11270499 TI - Human respiratory cells from nasal polyps as a model for gene transfer by non viral cationic vectors. AB - The influence of cell differentiation and proliferation on cationic vector mediated gene transfer into the explant-outgrowth cell culture from nasal polyps was investigated. Respiratory cells were categorized into two groups based on the expression of cytokeratin filaments (CKs), which were used as differentiation markers. Outgrowths grown for 2 weeks expressed similar levels of CKs 14, 13 and 18 showing a de-differentiated phenotype, while outgrowths cultured for 4 weeks presented very high levels of CK 13, high CK 14 and low CK 18 expression and were squamous differentiated. De-differentiated cells presented higher proliferation indexes than squamous cells. Gene transfer levels, as evaluated using a quantitative reporter gene (firefly luciferase), were significantly higher in the 2- than in the 4-week-old outgrowths. Cationic vector transfected respiratory cells were located both proximally and distally to the explant, as shown by enzymatic staining of beta-galactosidase-positive cells. Respiratory cell outgrowths from nasal polyps can be considered a suitable model to study gene transfer protocols in vitro. PMID- 11270500 TI - The anti-inflammatory effect of erythromycin and its derivatives, with special reference to nasal polyposis and chronic sinusitis. AB - Macrolides have been used for decades as an important chemotherapeutic agent in the treatment of infectious diseases. In the last 10 years there has also been increasing interest in the interaction between macrolide antibiotics and the immune system. The aim of this review is to focus on the anti-inflammatory action of erythromycin and its derivatives in the treatment of chronic sinusitis and nasal polyps. Systematic clinical investigations have been few and to the author's knowledge there have been no placebo-controlled studies. However there have been, especially from Japan, a number of clinical reports stating that long term, low-dose macrolide antibiotics are effective in treating chronic sinusitis incurable by surgery or glucocorticosteroid treatment, with an improvement in symptoms varying between 60% and 80% in different studies. In animal studies macrolides have increased mucociliary transport, reduced goblet cell secretion and accelerated apoptosis of neutrophils, all factors that may reduce the symptoms of chronic inflammation. There is also increasing evidence in vitro of the anti-inflammatory effects of macrolides. Several studies have shown macrolides to inhibit interleukin gene expression for IL-6 and IL-8 and also to inhibit the expression of intercellular adhesion molecule essential for the recruitment of inflammatory cells. There is also evidence in vitro, as well as clinical experience, showing that macrolides reduce the virulence and tissue damage caused by chronic bacterial colonization without eradicating the bacteria. The benefit of long-term, low-dose macrolide treatment seems to be that it is, in selected cases, effective when steroids fail. The exact mechanism of action is not known, but it probably involves downregulation of the local host immune response as well as a downgrading of the virulence of the colonizing bacteria. In the future, placebo-controlled studies should be performed to establish the efficacy of macrolides if this treatment is to be accepted as evidence-based medicine. PMID- 11270501 TI - Localization of cystic fibrosis transmembrane conductance regulator in epithelial cells of nasal polyps and postoperative polypoid mucosae. AB - Cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride channel protein that plays an important role in electrolyte and water transport through the respiratory epithelial cells. In order to understand the possible role of CFTR in the pathogenesis of nasal polyps and postoperative polypoid mucosae, we aimed to characterize the localization of CFTR in the epithelia of nasal polyps and postoperative polypoid mucosae of subjects who did not manifest the phenotypic expression of cystic fibrosis. Immunohistochemical staining for CFTR, using monoclonal mouse anti-human CFTR, was performed on tissue sections of 4 normal turbinates and nasal polyps and postoperative polypoid mucosae from 10 patients who underwent endoscopic intranasal operations. CFTR showed a typical apical distribution in the normal turbinate mucosae whereas, in the nasal polyps, CFTR demonstrated a heterogenous pattern of localization comprising diffuse or scattered cytoplasmic labelling, very low to undetectable labelling, intense perinuclear staining and intermingled typical apical location. In postoperative polypoid mucosae, the pattern of CFTR localization was less heterogeneous than in the nasal polyp epithelial cells and showed a more prominent feature of diffuse cytoplasmic staining. These results suggest that an altered localization of the CFTR may have a role in the pathogenesis of nasal polyps and postoperative polypoid mucosa. PMID- 11270502 TI - Examination of nasopharyngeal epithelium with contact endoscopy. AB - Primarily to evaluate the potential of contact endoscopy, the nasopharyngeal mucosa were examined using contact microscopy. With contact endoscopy it has been possible to visualize (60 x, 150 x ) the superficial cell layers of the nasopharyngeal epithelium, previously stained with methylene blue in vivo and in situ. Normal nasopharyngeal epithelium (20 cases) and cases with pathology (18 cases of chronic nasopharyngitis, 5 cases of nasopharyngeal cyst, and 57 cases of poorly differentiated squamous carcinoma) were assessed with contact endoscopy. The results showed that contact endoscopy can offer clear morphology and arrangement of the epithelial cells of the superficial layers and microvascular networks, such as the size and shapes of the cells or nuclei, nucleus/cytoplasm ratio, karyokinesis, etc. Our study indicated that contact endoscopy can permit the mapping of cellular alteration over mucosa and constitutes a new method of monitoring a high-risk population or precancerous lesions of nasopharyngeal carcinoma. PMID- 11270503 TI - Alpha-tocopherol protects against monocyte Mac-1 (CD11b/CD18) expression and Mac 1-dependent adhesion to endothelial cells induced by oxidized low-density lipoprotein. AB - Alpha-tocopherol supplementation is reported to protect against cardiovascular disease and to influence cells involved in atherogenesis, such as monocytes. Interactions between monocytes and vascular endothelial cells occur early in atherogenesis, and adhesion is mediated by integrins. We evaluated the effects of alpha-tocopherol on expression of Mac-1 (CD11b/CD18) by monocytes after stimulation with oxidized low-density lipoprotein (LDL), which is implicated as a potent chemotactic agent in atherogenesis. Incubation of whole blood with oxidized LDL (100 microg/ml) increased Mac-1 expression on monocytes, and preincubation with alpha-tocopherol reduced this upregulation in a concentration dependent manner. In another experiment, whole blood was obtained from healthy adult volunteers after 10 days of alpha-tocopherol administration (600 mg/day) and was incubated with oxidized LDL (100 microg/ml). There was a decrease in the upregulation of Mac-1 compared with that measured before administration. Adherence of oxidized LDL-stimulated monocytes to human umbilical vein endothelial cells was reduced by pretreatment with alpha-tocopherol, and was also inhibited by an anti-CD18 monoclonal antibody. Experiments with protein kinase C inhibitors suggested that reduction of Mac-1 upregulation by alpha-tocopherol was secondary to a decrease of protein kinase C activity. In conclusion, alpha tocopherol suppressed the upregulation of Mac-1 expression on monocytes by oxidized LDL. PMID- 11270504 TI - Properties of fucoidan from Cladosiphon okamuranus tokida in gastric mucosal protection. AB - To elucidate the anti-ulcer potential of Cladosiphon fucoidan, anti-peptic activity, bFGF stabilizing activity and inflammatory properties of this and related substances were investigated. Anti-peptic activity was observed with this and other sulfated polysaccharides such as dextran sulfate, carrageenan, and Fucus fucoidan. However, non-sulfated polysaccharides such as mannan and dextran did not exert the anti-peptic activity. The loss of bFGF bioactivity was prevented by all sulfated polysaccharides tested except chondroitin sulfate, at pH 7.4 and at pH 4.0. At pH 2.0, only heparin protected the bFGF activity. The generation of superoxide by macrophages and PMNs was stimulated by dextran sulfate, carrageenan, and Fucus fucoidan, whereas Cladosiphon fucoidan, heparin and chondroitin did not. Dextran sulfate, carrageenan, and Fucus fucoidan also stimulated the secretion of TNFalpha from macrophages, while Cladosiphon fucoidan did not. Thus, Cladosiphon fucoidan is a sulfated polysaccharide without inflammatory action. These results suggest that Cladosiphon fucoidan is a safe substance with potential for gastric protection. PMID- 11270505 TI - Adenosine modulates cell growth in baby hamster kidney (BHK) cells. AB - Adenosine is known to modulate cell growth in a variety of mammalian cells either via the activation of receptors or through metabolism. We investigated the effect of adenosine on Baby Hamster Kidney (BHK) cell growth and attempted to determine its mechanism of modulation. In wild-type BHK cells, adenosine evoked a biphasic response in which a low concentration of adenosine (1-5 microM) produced an inhibition of colony formation but at higher concentrations (up to 50 microM) this inhibition was progressively reversed. However, no biphasic response was observed in an "adenosine kinase" deficient BHK mutant, "5a", which suggests that adenosine kinase plays an important role in the modulation of growth response to adenosine. Adenosine receptors did not appear to have a role in regulating cell growth of BHK cells. Specific A1 and A2 receptor antagonists were unable to reverse the effect of adenosine on cell growth. Even though a specific A3 adenosine receptor antagonist MRS-1220 partly reversed the inhibition in colony formation at 1 microM adenosine, it also affected the transport of adenosine. Thus adenosine transport and metabolism appears to play the major role in this modulation of cell growth as 5'-amino-5'-deoxyadenosine, an adenosine kinase inhibitor, reversed the inhibition of cell growth observed at 1 microM adenosine. These results, taken together, would suggest that adenosine modulates cell growth in BHK mainly through its transport and metabolism to adenine nucleotides. PMID- 11270506 TI - Inhibition of angiotensin-converting enzyme protects endothelial cell against hypoxia/reoxygenation injury. AB - Cardiovascular tissue injury in ischemia/reperfusion has been shown to be prevented by angiotensin-converting enzyme (ACE) inhibitors. However, the mechanism on endothelial cells has not been assessed in detail. Cultured human aortic endothelial cells (HAEC) were exposed to hypoxia with or without reoxygenation. Hypoxia enhanced apoptosis along with the activation of caspase-3. Reoxygenation increased lactate dehydrogenase release time-dependently, along with an increase of intracellular oxygen radicals. ACE inhibitor quinaprilat and bradykinin significantly lessened apoptosis and lactate dehydrogenase release with these effects being diminished by a kinin B2 receptor antagonist and a nitric oxide synthase inhibitor. In conclusion, hypoxia activated the suicide pathway leading to apoptosis of HAEC by enhancing caspase-3 activity, while subsequent reoxygenation induced necrosis by enhancing oxygen radical production. Quinaprilat could ameliorate both apoptosis and necrosis through the upregulation of constitutive endothelial nitric oxide synthase via an increase of bradykinin, with the resulting increase of nitric oxide. PMID- 11270508 TI - Acute withdrawal from repeated cocaine treatment enhances latent inhibition of a conditioned fear response. AB - Psychostimulant-induced locomotor sensitization and disrupted latent inhibition (LI) of a classically conditioned association are two paradigms that have been widely studied as animal behavioural models of psychosis. In this study we assessed the effects of withdrawal from the repeated intermittent administration of cocaine on LI of a conditioned fear response. Animals which were either preexposed (PE) to a tone conditioned stimulus (CS) or naive to the tone (i.e. non-preexposed: NPE) subsequently experienced 10 pairings of the tone CS with footshock. Afterwards, both groups received five daily injections of cocaine (20 mg/kg, i.p.) or saline. After 3 days of withdrawal from drug treatment, animals were tested for conditioned freezing to the context of the footshock chamber, and 1 day later, for conditioned freezing to the tone CS. Cocaine-sensitized animals exhibited markedly enhanced LI compared to saline-treated animals, due to the fact that NPE-cocaine animals spent more time freezing during the tone CS than NPE-saline animals, whereas PE-cocaine animals showed a tendency toward reduced freezing compared to the saline groups. While these results suggest the presence of increased anxiety in cocaine-withdrawn NPE animals, the absence of this effect in cocaine-withdrawn PE rats indicates that cocaine withdrawal also influences the retrieval of previously learned information. PMID- 11270507 TI - The D3R partial agonist, BP 897, attenuates the discriminative stimulus effects of cocaine and D-amphetamine and is not self-administered. AB - Growing attention has been directed towards the potential involvement of the dopamine D3 receptor (D3R) in modulating effects of psychomotor stimulants. BP 897 (N-[4-[4-(2-methoxyphenyl)-1-piperazinyl]butyl]-2-naphthylcarboxamide; aka BP 4.897 and DO897) is amongst the most selective partial agonists for the D3R receptor thus far reported. BP 897 was tested for its ability to support self administration in rhesus monkeys (0.3-30 microg/kg) and for its ability to produce cocaine- and D-amphetamine-like discriminative stimulus effects in mice (0.01-17 mg/kg i.p.). BP 897 was not self-administered above vehicle and saline levels in any of the four monkeys tested, and produced less than 30% generalization from either the cocaine or D-amphetamine stimulus. When BP 897 was administered before administrations of cocaine or D-amphetamine, percent drug lever selections were reduced. These results suggest that BP 897 has a profile of activity suitable for consideration as a potential treatment for cocaine dependency disorders. PMID- 11270509 TI - Intravenous self-administration of heroin/cocaine combinations (speedball) using nose-poke or lever-press operant responding in mice. AB - Acquisition and dose-related self-administration of heroin (H)/cocaine (C) combinations in C57BL/6 x SJL mice were studied in nose-poke or lever-press operant responding procedures. C57BL/6 x SJL mice readily acquired self administration in both operant procedures with a combination of doses known to be ineffective when each drug was used alone (H: 15 microg/kg and C: 150 microg/kg per injection). Similar numbers of infusions were obtained under conditions of fixed-ratio (FR) 3 versus 1 for the nose-poke and lever-press responses, respectively. Dose-effect curves for heroin:cocaine combinations revealed a pattern corresponding to a leftward shift of the dose-response for intravenous cocaine self-administration. Curves were similar whether generated with 1 or 3 days of availability per dose, or including subjects that did not respond preferentially (> 70% responses) to the hole or lever associated with drug delivery, along with those that did. Motor activity induced by a combination of low doses for each drug was examined (H: 0.375 mg/kg and C: 3.75 mg/kg, i.p.). Under these conditions, the combination of both drugs induced an initial cocaine like stimulation of horizontal activity, in contrast to the tendency of heroin to decrease activity. It is concluded that heroin:cocaine combinations used in the present study had reinforcing effects in C57BL/6 x SJL mice, and produced a cocaine-like effect in the early part of drug-induced activity sessions, followed by a locomotor profile corresponding to the average of both drugs. PMID- 11270510 TI - The discriminative stimulus and reinforcing effects of nicotine in humans following nicotine pretreatment. AB - Smokers often report that the first cigarette of the day is the most rewarding, and subsequent smoking is less rewarding. Reduction in smoking enjoyment later in the day may be related to acute tolerance to the discriminative stimulus effects of nicotine. We examined changes in nicotine discrimination behaviour in humans as a function of acute nicotine pretreatment. Male and female dependent smokers (n = 15) were initially trained to discriminate 20 microg/kg nicotine by nasal spray from placebo (0 microg/kg) without nicotine pretreatment. They then were tested on generalization of discrimination across a range of spray doses from 0 20 microg/kg following pretreatment with placebo, moderate dose (14-21 mg) or high dose (28-42 mg) transdermal nicotine. Generalization testing involved both two- and three-response ('novel' option) quantitative procedures. Subjects also engaged in a self-administration phase at the end of each session, involving choices between nicotine (20 microg/kg) and placebo spray. Nicotine pretreatment significantly attenuated nicotine-appropriate responding at higher nicotine spray doses, suggesting acute tolerance, but only in women. Similar results were seen for subjective 'head rush', suggesting this effect may be related to discrimination behaviour in women. However, nicotine pretreatment also increased novel-appropriate responding, especially in men, following intermediate generalization doses, suggesting qualitatively different stimulus effects. Although differences were not significant, nicotine self-administration tended to be inversely associated with nicotine pretreatment dose in men but not in women. These results only modestly support the notion of acute tolerance to the discriminative stimulus effects of nicotine, and even then only in women and not in men. PMID- 11270511 TI - Matching strategies for drug studies of prepulse inhibition in humans. AB - Prepulse inhibition (PPI), a measure of sensorimotor gating, is impaired in certain neuropsychiatric disorders. Animal studies have revealed drug effects on PPI that may be relevant to understanding the biology of gating deficits in human populations. Recent efforts have examined similarities and differences in drug effects on PPI between rodents and humans. Experimental designs are needed that most effectively translate these drug studies across species. In the course of a larger set of studies of drug effects on startle in normal human subjects, we examined the potential utility of one design element that is utilized in rodent PPI drug studies: pre-testing to diminish variability across dose groups. Startle was measured during a screening session; 7-10 days later, 20 subjects were retested after consuming a placebo pill. Acoustic and tactile startle, and unimodal and cross-modal PPI, were measured in five sessions over a period of 3 hours post-placebo. There were significant and robust correlations between levels of startle magnitude and PPI during pre-testing and testing, for both left and right eyeblink measures. Comparable correlations were evident for both unimodal and cross-modal testing. Pre-testing values were most predictive of test performance early in the 3-hour test session, and predictive strength diminished or disappeared towards the end of testing. The utility of a pre-testing design could be seen clearly by comparing groups 'matched', based on pre-test data, versus groups created by alternating or random group assignments. It is concluded that pre-test designs can effectively match groups with comparable levels of startle or PPI, and thereby diminish between-group variability in human PPI drug studies. For studies using repeated testing to assess drug time course, the predictive value of pre-testing is greatest in early test sessions. PMID- 11270512 TI - Effects of acute D-amphetamine and ketamine on the performance of rats in a serial negative patterning procedure. AB - This study assessed the effects of acute amphetamine and ketamine on the performance of rats in a serial negative patterning procedure. A 5 s auditory target stimulus and a 5 s visual feature cue were each followed by food, but the target stimulus was not followed by food if preceded by the feature. There was a 5 s empty gap between feature termination and target onset in the latter, non reinforced trials. Thus, the feature functioned as a cue signalling the non reinforcement of the target. The interval between the feature and the target was varied in the non-reinforced trials following pretreatment with subcutaneous saline, D-amphetamine (0.5 mg/kg) or ketamine (5 mg/kg). The main behaviour measured was visits to the place of food delivery during target presentation. Under saline, the response frequency during the target was lowest when the interval between the feature and the target exactly matched the interval used during training. Either shortening or lengthening the interval enhanced responding. Neither D-amphetamine nor ketamine disturbed this temporal pattern, although D-amphetamine and ketamine non-specifically increased and decreased response frequencies, respectively, in all the trial types. The results are discussed in the framework of the amphetamine and ketamine models of schizophrenia. PMID- 11270513 TI - Differences in locomotor response to an inescapable novel environment predict sensitivity to aversive effects of amphetamine. AB - Differences in locomotor response to an inescapable novel environment have previously been shown to predict sensitivity to amphetamine reward, where high responders (HR), compared to low responders (LR), showed greater initial sensitivity to amphetamine self-administration. The present experiments sought to extend these findings and assessed the relationship between locomotor response to an inescapable novel environment and conditioned taste aversion (CTA) with amphetamine and lithium chloride (LiCl). Male Sprague-Dawley rats were tested for their locomotor response to an inescapable novel environment and divided into high (HR) or low (LR) responders, based on whether their locomotor scores were above or below the median activity level of the subject sample. After several days, the animals were tested in a CTA procedure and conditioned with either amphetamine or lithium chloride. Compared to HR rats, LR rats showed greater sensitivity to amphetamine CTA at the doses tested. In contrast, the results with LiCl showed no relationship between locomotor response to an inescapable novel environment and CTA. Taken together, the present results suggest that LR, compared to HR, rats show less sensitivity to the rewarding effects of amphetamine because they are more sensitive to aversive effects of amphetamine, as reflected in CTA. In contrast, HR rats display less sensitivity to aversive effects of amphetamine, which may explain their greater propensity to self administer amphetamine. PMID- 11270514 TI - Reversal effect of sulpiride on rotational behaviour of rats with unilateral frontal cortex ablation: an alternative explanation for the pharmacological mechanism of its antidepressant effect. AB - The antidepressant effect of sulpiride has been generally explained as the result of its preferential blocking effect on self-inhibitory presynaptic dopamine autoreceptors at low doses. Low dose haloperidol has the same blocking effect. In rats with unilateral ablation of the frontal cortex, methamphetamine administration induced mild contralateral rotation 10 days after the operation. We examined whether low dose sulpiride and haloperidol would have the same effect on this rotational model. High dose sulpiride (100 mg/kg) or low dose haloperidol (0.05 mg/kg) prevented this methamphetamine-induced rotation. However, low dose (15 mg/kg) sulpiride clearly reversed the direction of rotation. This reversal effect of low dose sulpiride is not explained by the preferential blocking effect on dopamine autoreceptors. The results suggest that low dose sulpiride, unlike low dose haloperidol, has a prominent blocking effect on D2 receptors in the frontal cortex. This unique effect of sulpiride may be relevant to its clinical antidepressant and anxiolytic effects at low doses. PMID- 11270515 TI - Lack of sex-related differences in the prevention by baclofen of the morphine withdrawal syndrome in mice. AB - In previous studies we have demonstrated a possible interaction between the GABAergic and opioid systems involved in the antinociceptive effect of the GABA(B) agonist, baclofen (BAC). On the other hand, sex differences have been observed for the antinociceptive effect of morphine (MOR). In the present study, we analyzed sex-related differences in the MOR abstinence syndrome and its prevention with BAC. Prepubertal male and female Swiss-Webster albino mice (27-33 g) were rendered dependent by intraperitoneal (i.p.) injection of MOR (2, 4 and 8 mg/kg), twice daily for 9 days. On the tenth day the dependent animals were divided into two groups: one received naloxone (NAL) (6 mg/kg, i.p.) 60 min after the last dose of MOR, to precipitate the abstinence syndrome; the other group received BAC (2 mg/kg, i.p.) followed by NAL (6 mg/kg, i.p.), injected 30 and 60 min after the last dose of MOR, respectively. Behavioral signs were recorded in the open field for 30 min. Although there were sex differences in the MOR withdrawal syndrome, we found a lack of sex differences in the prevention of the MOR abstinence syndrome by BAC. PMID- 11270516 TI - Effects of the herbal medicine "Sai-rei-to" on in vitro interferon-gamma production of peripheral blood mononuclear cells. AB - The herbal medicine "Sai-rei-to" has been used in the treatment of swellings and edemas for about 3000 years in China. Recently, this drug has been prescribed as an adjuvant in the treatment of rheumatoid arthritis (RA) among Japan's western medicine doctors. It is thought to possess regulatory effects on the immune system, although its mode of action is not yet fully described. In the present in vitro study, we at first induced interferon-gamma (IFN-gamma) in the cultures of peripheral blood mononuclear cells of healthy volunteers by adding pokeweed mitogen (PWM), phytohemagglutinin (PHA), recombinant interleukin-2 (IL-2), or control medium. We then added "Sai-rei-to" to these cultures and investigated the effects of this drug on IFN-gamma production levels. The results demonstrated that "Sai-rei-to" had 2 different effects: (i) it inhibited the IFN-gamma production in cultures with PWM or PHA (which induced large volumes of IFN gamma), and (ii) it increased IFN-gamma production in the cultures with IL-2 (IL 2 induced only small volumes of IFN-gamma). These findings indicate that "Sai-rei to" possesses regulatory effects on the IFN-gamma production. IFN-gamma is an important cytokine in the immune system, and it has also attracted attention as a factor related to the pathogeneses of RA. Therefore, concomitant administration of such a medicine which can appropriately control IFN-gamma production in vivo could be beneficial for RA patients from the immunological viewpoint. Clinical experience in the past has shown that "Sai-rei-to" has a very low incidence of side effects, and can be administered orally for long periods. We expect that concomitant administration of "Sai-rei-to" with current therapy could be clinically useful in the management of RA patients. PMID- 11270517 TI - Circulating immune complex levels measured by new ELISA kits utilizing monoclonal anti-C1q and anti-C3d antibodies correlate with clinical activities of SLE but not with those of RA. AB - The authors studied sera from both patients with SLE and from those with RA to evaluate clinical usefulness and significance of circulating immune complexes (CIC) detected with new ELISA kits utilizing monoclonal anti-C1q and anti-C3d antibodies. CIC values of patients with SLE significantly correlated to severity of disease activities evaluated by clinical symptoms and laboratory tests, especially for serum complement levels. Since substances detected with the ELISA kits were closely related to serum complement components, it was determined that a direct relationship exists between clinical activities and CIC values appearing in SLE patients with hypocomplementemia. In RA patients, CIC values did not correlate to clinical activities evaluated by Lansbury's index, anatomical bone damage with X-ray or functional assessment of activities of daily living, but did significantly correlate to levels of IgM-RF, serum IgG concentrations and some markers of systemic inflammation. Detection of IC after fractionation of RA sera revealed a broad range of molecular sizes detectable with the ELISA kits, which indicated that CIC in vivo were heterogenic and complicated in formation, degradation and interaction with serum complements. PMID- 11270518 TI - Cerebrospinal fluid beta 2-microglobulin in AIDS related central nervous system involvement. AB - We measured cerebrospinal fluid (CSF) and serum beta 2-microglobulin (beta 2-M) in Acquired Immunodeficiency Syndrome (AIDS) patients with or without clinical evidence of central nervous system (CNS) involvement. The CSF beta 2-M level was significantly higher than the serum level in AIDS patients with neurological symptoms, but not in AIDS without neurological symptoms, suggesting an increased shedding of this protein in CSF, as a result of rapid cellular turnover within CNS. CSF beta 2-M level increases both in Human Immunodeficiency Virus type 1 (HIV-1) related and in opportunistic CNS syndromes, confirming that beta 2-M is a non specific marker of CNS involvement in AIDS. Nevertheless, the highest CSF beta 2-M values were observed in patients with severe dementia and autoptic diagnosis of multifocal giant cells encephalitis (MGCE) without other opportunistic diseases. This observation could have important implications for monitoring AIDS dementia complex in AIDS patients. Finally, 5 out of 7 (71%) AIDS patients with cryptococcal meningitis showed a decline in CSF beta 2-M level well related to the decrease of cryptococcal antigen (Crypto-Ag) titres and the clinical remission. This data suggests that CSF beta 2-M determination could be used as a useful test in monitoring efficacy of therapy of CNS pathologies in AIDS patients. PMID- 11270519 TI - Inhibition of PHA-stimulated proliferation of peripheral blood mononuclear cells by human seminal plasma inhibin (HSPI). AB - This report deals with the effect of purified HSPI (10.7 kDa single chain prostatic peptide of 94 amino acids) on PHA-stimulated proliferation of peripheral blood mononuclear cells (PBMNC). HSPI was found to inhibit the incorporation of 3H-thymidine by PHA-stimulated PBMNC in a dose-dependent manner, albeit with the requisite of pretreatment of cells with HSPI. A maximum inhibition was observed on pretreatment for 30 min. These studies thus indicate that HSPI may be interfering with the very early events of PHA-induced signal transduction process in PBMNC. PMID- 11270520 TI - Unilateral cerebral ventriculomegaly: is one better than two? PMID- 11270521 TI - Neonatal nucleated red blood cell counts: relationship to abnormal fetoplacental circulation detected by Doppler studies. AB - Increased neonatal nucleated red blood cell counts are thought to be related to intrauterine hypoxemia. We sought to determine the effect of increasing circulatory impairment in fetuses on the neonatal nucleated red blood cell count. One hundred thirty-four singleton pregnancies were included in the study and were allocated to 4 study groups according to Doppler findings. The systolic-to diastolic ratios of the umbilical artery, fetal aorta, middle cerebral artery, and uterine arteries were recorded. Fetuses were assigned to the following groups on the basis of the last Doppler examination before delivery: group 1, normal systolic-to-diastolic ratios in the examined vessels; group 2, a systolic-to diastolic ratio greater than 2 SD above the mean for gestational age in the umbilical artery or fetal aorta and no abnormal Doppler findings in the uterine arteries; group 3, systolic-to-diastolic ratios greater than 2 SD above the mean for gestational age in all examined vessels; and group 4, absence of end diastolic velocity in the umbilical artery or fetal aorta and systolic-to diastolic ratios greater than 2 SD above the mean for gestational age in the uterine arteries. A blood sample from the umbilical artery was obtained within 1 minute after birth, and nucleated red blood cells per 100 white blood cells were counted by light microscopy. Nucleated red blood cell counts were higher in fetuses in group 4 (median, 72.0; range, 9-720; P < .001) and group 3 (median, 38.4; range, 7-201; P < .001) than in fetuses in group 1 (median, 5.1; range, 0 20). Neonates in group 4 had significantly lower birth weights (P < .001), lower arterial and venous pH values (P < .05), and lower Apgar scores after 5 minutes (P < .01) as well as an increased likelihood of cesarean delivery because of fetal distress (P < .001). The number of fetuses in group 4 with a cord blood base deficit of less than -8 mmol/L was increased. Nucleated red blood cell counts were comparable in fetuses in group 2 (median, 5.4; range, 0-37) and group 1. In groups 1 to 3 no brain-sparing effect occurred, whereas in 15 of 21 cases in group 4 a brain-sparing effect was present. Multivariate analysis revealed that Doppler results of the umbilical artery, fetal aorta, and uterine arteries were independent determinants of neonatal nucleated red blood cell count. Increasing abnormalities seen on fetoplacental Doppler studies are associated with increasing numbers of nucleated red blood cells at birth. Given the known relationship between abnormal Doppler flow and intrauterine hypoxemia, the neonatal nucleated red blood cell count might become an additional valuable tool in the surveillance of growth-restricted fetuses. PMID- 11270522 TI - Subjective versus objective evaluation of amniotic fluid volume of pregnancies of less than 24 weeks' gestation: how can we be accurate? AB - This study was undertaken to compare subjective versus objective ultrasonic evaluation of amniotic fluid volume in pregnancies of less than 24 weeks' gestation. Amniotic fluid volume was subjectively (visualization without ultrasonic measurements) and objectively (visual interpretation with ultrasonic measurements) evaluated in 42 singleton pregnancies undergoing termination. The actual amniotic fluid volume was then determined using a dye-dilution technique. The women evaluated were in their mid-20s, primarily African American, and between 15 and 23 weeks' gestation. There was no significant difference in the total number of correct estimates of amniotic fluid volume when the data were stratified by level of operator experience (P = .34), ultrasonic technique (P = .33), or the combined correct subjective versus combined correct objective estimates (P = .68). We have concluded that the accuracy of amniotic fluid volume assessment in pregnancies of less than 24 weeks is not influenced by the level of operator experience or the type of ultrasonic measurement. PMID- 11270523 TI - Low-intensity pulsed ultrasound initiates bone healing in rat nonunion fracture model. AB - Low-intensity pulsed ultrasound exposure has been shown clinically to shorten the fracture repair process and to induce healing of nonunions in humans, but its mechanism of action remains unclear. In this study we investigated the effect and mechanism of low-intensity pulsed ultrasound on nonunion fracture healing in rat tibias. A consistently reproducible nonunion was produced in rat tibias by muscle interposition without osteotomy. This model was produced by creating a closed tibial fracture with only the distal end of the tibialis anterior muscle interposed into the fracture site. One limb was noninvasively exposed to low intensity pulsed ultrasound (a 200-millisecond burst of sine waves of 1.5 MHz, repeating at 1.0 kHz) for 20 minutes daily. The incident intensity was approximately 30 mW/cm2. Rats were killed at intervals between 2 and 6 weeks. The events were assessed by radiographs, microfocus X-ray computed tomograms, and histologic examination. After 6 weeks of exposure, 7 of 14 nonunion fractures showed healing on radiologic assessment. The results of three-dimensional microfocus X-ray computed tomographic reconstruction and histologic examination also supported this finding. On the other hand, all control tibias remained in a state of nonunion during the same period. These results indicate that low intensity pulsed ultrasound promotes healing in the rat nonunion fracture model. PMID- 11270524 TI - Duplex criteria for determination of 50% or greater carotid stenosis. AB - Recently the North American Symptomatic Carotid Endarterectomy Trial investigators reported a benefit of carotid endarterectomy compared with medical therapy for symptomatic patients with 50% or greater carotid stenosis. This has necessitated the development of screening parameters for diagnosis of 50% or greater carotid stenosis on the basis of the reference standards used in the study by the North American Symptomatic Carotid Endarterectomy Trial. The duplex scans and arteriograms of 110 patients (210 carotid arteries) were reviewed by blinded readers. Duplex measurements of peak systolic velocity and end diastolic velocity were recorded, and the ratio of these velocities in the internal and common carotid arteries was calculated. The criteria determined for detection of 50% or greater stenosis were as follows: peak systolic velocity of the internal carotid artery greater than 170 cm/s (sensitivity, 92%; specificity, 90%; positive predictive value, 92%; negative predictive value, 90%; and accuracy, 91 %); end diastolic velocity of the internal carotid artery greater than 60 cm/s (sensitivity, 92%; specificity, 86%; positive predictive value, 95%; negative predictive value, 79%; and accuracy, 91 %); ratio of peak systolic velocity of the internal carotid artery to peak systolic velocity of the common carotid artery greater than 2 (sensitivity, 93%; specificity, 75%; positive predictive value, 83%; negative predictive value, 89%; and accuracy, 85%); and ratio of end diastolic velocity of the internal carotid artery to end diastolic velocity of the common carotid artery greater than 2.4 (sensitivity, 96%; specificity, 79%; positive predictive value, 88%; negative predictive value, 92%; and accuracy, 89%). It is concluded that 50% or greater carotid artery stenosis can be reliably determined by duplex criteria. The use of receiver operating characteristic curves allows the individualization of duplex criteria to the clinical situation. PMID- 11270526 TI - Analysis of color Doppler signal intensity variation after levovist injection: a new approach to the diagnosis of thyroid nodules. AB - The objective of this study was to evaluate the usefulness of a galactose-based ultrasonographic contrast agent, Levovist (Schering AG, Berlin, Germany), in differentiating benign from malignant thyroid nodules by analysis of the time intensity curves correlating the variation of the intensity signal value during the contrast transit time. Fifty-four patients scheduled for surgical removal of a nodule or the thyroid gland or both after cytologic examination were enrolled in this study; all of the nodules underwent a baseline color and power Doppler evaluation and then to a color Doppler examination after an intravenous bolus injection of Levovist. The time-intensity curves were analyzed with respect to the histologic results. Carcinomas showed a significantly earlier arrival time of Levovist than nodular hyperplastic benign nodules and adenomas (8.1 +/- 1.41 versus 19.6 +/- 2.2 and 16.1 +/- 2.8 seconds; P < .0001), although no significant difference occurred between hyperplastic benign nodules and adenomas; carcinomas and adenomas showed an earlier time to peak than hyperplastic benign nodules (14.6 +/- 1.2 and 23.1 +/- 3.8 versus 33.0 +/- 3.0 seconds; P < .0001). No significant difference was found in baseline, peak, final intensity signal, and percent variation of intensity signal among hyperplastic benign nodules, adenomas, and carcinomas. Although cytologic examination still remains the standard of reference for the presurgical diagnosis of thyroid nodules, the preliminary data of this pilot study demonstrate that the analysis of time intensity curves after Levovist injection might provide useful, complementary, and quantitative information to differentiate benign from malignant thyroid nodules. PMID- 11270525 TI - Cephalic vein and hemodialysis fistula: surgeon's observation versus color Doppler ultrasonographic findings. AB - The aim of this study was to evaluate whether preoperative color Doppler ultrasonography improves immediate success rates of arteriovenous fistulas for dialysis. One hundred twenty-four patients with chronic renal failure underwent color Doppler ultrasonographic examination of both arms, including the cephalic vein, before arteriovenous fistula construction. Patients were randomly divided into 2 groups: A and B. In group A, there were 52 patients, and the surgeon planned to construct arteriovenous fistulas depending only on physical examination. In group B, which comprised 72 patients, surgeons performed arteriovenous fistula construction on sites labeled by color Doppler ultrasonography. In group A, of 52 patients who had surgery for arteriovenous fistula construction, 13 had fistulas that did not function. Among these 13 patients, 8 were found to have chronic thrombotic changes in the cephalic vein on color Doppler ultrasonography, and 5 had none of these changes. When we checked the color Doppler ultrasonographic findings, we noted that these 5 patients had decreased volume flow in the radial artery. On the whole, the arteriovenous fistulas worked in 39 patients (75%) and did not function in 13 patients (25%). In group B, surgeons followed the color Doppler ultrasonographic results. Of 72 patients who underwent the procedure, 68 patients (94.4%) had functioning fistulas, whereas 4 (5.6%) had fistulas that did not work. These 4 patients were found to have low volume flow in the radial artery. When both groups were compared by chi2 analysis, the difference was statistically significant (P = .002). Group B, in which patients were preoperatively evaluated by color Doppler ultrasonography, had a high success rate. We found that color Doppler ultrasonography is very helpful as a noninvasive procedure for this evaluation. Although many surgical clinics still perform arteriovenous fistula construction without the aid of color Doppler ultrasonographic findings, we think that the use of color Doppler ultrasonography should be emphasized before surgeons proceed with arteriovenous fistula construction. PMID- 11270527 TI - Overlaying ultrasonographic images on direct vision. AB - The objective of this technical advance is to permit in situ visualization of ultrasonographic images so that direct hand-eye coordination can be used during invasive procedures. A method is presented that merges the visual outer surface of a patient with a simultaneous ultrasonographic scan of the patient's interior. The method combines a flat-panel monitor with a half-silvered mirror such that the image on the monitor is reflected precisely at the proper location within the patient. The ultrasonographic image is superimposed in real time on the patient, merging with the operator's hands and any invasive tools in the field of view. Instead of looking away from the patient at an ultrasonographic monitor, the operator sees through skin and underlying tissue as if it were translucent. Two working prototypes have been constructed, demonstrating independence of viewer location and requiring no special apparatus to be worn by the operator. This method could enable needles and scalpels to be manipulated with direct hand-eye coordination under ultrasonographic guidance. Invasive tools would be visible up to where they enter the skin, permitting natural visual extrapolation into the ultrasonographic slice. Biopsy needles would no longer be restricted to lie in the plane of the ultrasonographic scan but could instead intersect it. These advances could lead to increased safety, ease, and reliability in certain invasive procedures. PMID- 11270528 TI - Sonographic depiction of ovarian vascularity and flow: current improvements and future applications. AB - The objective of this image presentation is to update the reader with the current clinical applications and future developments concerning the sonographic depiction of ovarian vascularity and flow. This topic is discussed and illustrated using numerous figures that show actual images and their histologic correlation as well as illustrative drawings of central concepts. Color Doppler sonography is an accurate means to depict ovarian vascularity and flow. This information can be used to detect ovarian cancer and adnexal torsion. Future developments include the use of sonographic contrast agents and three-dimensional imaging. This report describes the current state of the art regarding Color Doppler sonography of ovarian vascularity and flow. It also describes areas for future research and development. PMID- 11270529 TI - Intravascular ultrasonographic findings in May-Thurner syndrome (iliac vein compression syndrome). AB - The objective of this series was to describe the findings in 2 types of iliac vein compression syndrome on intravascular ultrasonography. We conducted a retrospective review of the intravascular ultrasonographic findings in 2 patients with iliac vein compression syndrome due to 2 different types of venous spur and correlated those findings with the contrast-enhanced venographic findings. Intravascular ultrasonography allowed differentiation of the 2 different types of iliac vein compression syndrome in analogy to the venographic classification. Both cases had compression of the left common iliac vein between the right common iliac artery and the vertebral bodies. In addition, hyperechogenic eccentric wall thickening, the presence of multiple vascular channels separated by hyperechogenic structures, and the presence of sequelae of deep venous thrombosis such as synechiae and organized thrombus can be seen on intravascular ultrasonography. The ability to perform exact measurements of the venous dimensions is an added benefit of intravascular ultrasonography. Intravascular ultrasonography can assist in the diagnosis and classification of iliac vein compression syndrome, allows assessment of its complications, and can be potentially helpful for planning endovascular treatment. PMID- 11270530 TI - Outcome of prenatally diagnosed mild unilateral cerebral ventriculomegaly. AB - The objective of this study was to determine the frequency of prenatally diagnosed unilateral cerebral ventriculomegaly and also to assess neonatal outcome in infants with this prenatal diagnosis. A computerized ultrasonography database identified fetuses with isolated and nonisolated unilateral cerebral ventriculomegaly from October 1994 to June 1999. The Denver II Developmental Screening Test was used to assess developmental skills. Unilateral cerebral ventriculomegaly was diagnosed in 15 of 21,172 (1 per 1,411) pregnancies. The width of the enlarged lateral ventricle ranged from 1.0 to 1.9 cm. In 10 (67%) of 15 cases unilateral cerebral ventriculomegaly was an isolated finding. Eight of the 14 infants who were born at 36 weeks' gestation or later had postnatal cranial imaging, and ventricular asymmetry was confirmed in 5 (63%). One infant with an arachnoid cyst and cerebral palsy died at 2 years of age. The remaining 11 infants in whom developmental milestones were assessed had age-appropriate skills. Unilateral fetal ventriculomegaly is usually an isolated finding and when isolated has little measurable effect on developmental outcome. PMID- 11270531 TI - Postnatal outcome of fetuses with the prenatal diagnosis of asymmetric hydrocephalus. AB - We sought to assess the sonographic findings and postnatal outcome in fetuses with the prenatal diagnosis of asymmetric hydrocephalus. The sonograms from cases of asymmetric hydrocephalus diagnosed prenatally at our institution were reviewed. Postnatal outcome was obtained from maternal, neonatal, and pediatric records. Fourteen fetuses at 17.3 to 38.9 weeks' gestational age on prenatal sonography had a maximum ventricular measurement of 10.2 to 48.8 mm, with the degree of asymmetry ranging from 2.2 to 27.3 mm. Thirteen of 14 had a normal sized contralateral ventricle. Other fetal anomalies identified at sonography included Dandy-Walker malformation, intraventricular hemorrhage, porencephalic cyst, hydronephrosis, pleural effusion, and mild dilatation of a renal pelvis. Eleven fetuses had follow-up prenatal sonography. Among these, ventricular dilatation resolved in 5, remained the same in 3, increased in 2, and decreased in 1. Postnatal outcome was normal in 6 cases (43%) and abnormal in 8 (57%), including 2 cases of in utero intracranial hemorrhage, 2 with congenital syndromes, 1 with an imperforate foramen of Monro, 1 with tuberous sclerosis, 1 with developmental delays, and 1 with cerebral palsy. Asymmetric unilateral hydrocephalus appears to represent an entity different from bilateral hydrocephalus in that there is less risk of perinatal death, there are fewer associated anomalies, and the overall prognosis is better. Outcome may be normal, but fetuses with increasing unilateral ventriculomegaly and cases associated with other brain abnormalities tend to have a poor neurologic outcome. PMID- 11270532 TI - Ultrasonographic appearance of Ascaris lumbricoides in the small bowel. AB - Roundworm infestation, one of the most common helminthic diseases worldwide, is caused by Ascaris lumbricoides, one of the largest parasites that infests the human bowel. A lumbricoides is virtually universal at some stage of childhood in semitropical and tropical regions. This study describes our experience with the ultrasonographic appearance of intestinal ascariasis in 84 patients, 2.5 to 42 years of age, examined over 2 years beginning October 1997. The patients' conditions ranged from acute intestinal obstruction to no clinical features pertaining to obstruction. Ultrasonographic examination was performed with an Echocee power Doppler real-time unit with a variable-frequency 3.7-MHz convex, 7.5-MHz linear probe. In longitudinal section the Ascaris worm presented as a linear intraluminal mass with 3 or 4 linear echogenic interfaces; in the cross section, it was round, sometimes appearing as a "target" sign. Some worms also showed serpentine movements. Sonographic examination of the patients in the left lateral decubitus position after ingestion of water improved detection and visualization of the worms in some cases. It is concluded that A lumbricoides in the small bowel has a sonographic appearance that can be recognized by the wary observer. PMID- 11270533 TI - Ultrasonographic appearance of colon taeniasis. AB - We present the case of a 50-year-old woman with abdominal pain, nausea, loss of appetite, and frequent stools in whom the routine ultrasonographic examination demonstrated a double-reflective, ribbon-like structure in the lumen of the initial segment of the ascending colon, which suggested colon taeniasis. Because the initial parasitologic analysis yielded negative results and application of albendazol did not have any therapeutic effect, the diagnosis was confirmed by barium enema and subsequently by parasitologic examination of proglottids passed in the stool after application of niclosamide. The double-reflective, ribbon-like structure in the lumen of the intestine seems to be specific to the ultrasonographic appearance of intestinal taeniasis. Transcutaneous ultrasonography of the gastrointestinal tract, performed as a screening method before conventional radiologic or endoscopic examination, can point to the ultimate diagnosis of colon taeniasis. PMID- 11270534 TI - Sonographic diagnosis of multiple unilateral ovarian teratomas. PMID- 11270535 TI - Treating complex nervous system vascular disorders through a "needle stick": origins, evolution, and future of neuroendovascular therapy. AB - In the past few decades, dramatic improvements have occurred in the field of neuroendovascular surgery. Endovascular therapy today is a well-established treatment modality for a variety of cerebrovascular and nonvascular central nervous system diseases. The foundation of this spectacular evolution was laid by the efforts of pioneering visionaries who often worked alone and under difficult, almost impossible, conditions. Ongoing device development and refinement have revolutionized the field at a dizzying, exhilarating pace. With a better understanding of the molecular basis of diseases and further advancements in gene therapy, neuroendovascular techniques have an enormous potential for application to the entire spectrum of central nervous system diseases as a minimally invasive vehicle for the delivery of biological factors. PMID- 11270536 TI - Surgical repair of clinoidal segment carotid artery aneurysms unsuitable for endovascular treatment. AB - OBJECTIVE: Clinoidal segment carotid artery aneurysms are surgically challenging lesions. The aneurysm neck originates proximal to the distal dural ring, and the aneurysms typically are larger. Therefore, endovascular techniques are often considered to be the primary treatment option. Treatment techniques and results for 40 clinoidal segment carotid artery aneurysms that were considered unsuitable for contemporary endovascular intervention are analyzed in this report. METHODS: Forty aneurysms in 33 female and 3 male patients were treated surgically. Fifteen patients had bilateral aneurysms; of these patients, four underwent bilateral craniotomies. Twenty-seven aneurysms were 10 to 14 mm in size, eight were 15 to 24 mm, and five were more than 25 mm. The most common presentation was visual loss, which occurred in 13 patients. Seven patients presented with subarachnoid hemorrhage. RESULTS: Thirty-seven aneurysms were directly repaired with clipping, two were trapped with bypass, and one was trapped without bypass. The complication rate was 10%, with one major stroke, two minor strokes, and one successfully treated brain abscess. CONCLUSION: Surgical treatment of clinoidal segment carotid artery aneurysms can produce acceptable outcomes. Specific preoperative and intraoperative techniques facilitate improved surgical results for aneurysms that are not treatable with contemporary endovascular techniques. PMID- 11270537 TI - Rheolytic catheter and thrombolysis of dural venous sinus thrombosis: a case series. AB - OBJECTIVE: The high morbidity and mortality rates associated with dural sinus thrombosis may be heightened by a delay in diagnosis, which necessitates prompt and effective treatment. Traditional treatment consists of the initiation of systemic anticoagulation with heparin and, more recently, regional thrombolysis with direct endovascular infusion of thrombolytic agents. We report our experience in a series of five patients in whom we accomplished mechanical clot lysis with the combination of a rheolytic device and balloon catheters. METHODS: Five patients with dural sinus thrombosis were treated with a combination of pharmacological and mechanical thrombolysis with the 5-French Angiolet rheolytic catheter (Possis Medical, Minneapolis, MN) and balloon catheters. The success of the procedure was determined by resolution of or improvement in the patient's neurological examination results and imaging features. RESULTS: All five patients demonstrated immediate improvement as observed on imaging studies or in terms of neurological status. Three patients required more than one intervention, and all but one patient continued to improve after the final intervention. Two of the five patients continued to experience mild residual neurological deficits, and two patients experienced complete recovery. The fifth patient had a delayed recurrence of thrombosis that required multiple interventions, and the patient has significant neurological deficits. Navigation of the dural sinuses was possible in all patients with the use of a microcatheter and was possible to a variable degree with the rheolytic catheter. Known complications of the procedures included two pseudoaneurysms at the femoral puncture site. CONCLUSION: Mechanical clot lysis is a powerful technique for immediate restoration of antegrade venous flow in dural sinus thrombosis. In most patients, the superior sagittal sinuses and contralateral transverse sinuses could be accessed with the 5-French rheolytic catheter. PMID- 11270538 TI - Is the aspect ratio a reliable index for predicting the rupture of a saccular aneurysm? AB - OBJECTIVE: The present retrospective study was undertaken to prove the reliability of the aspect ratio (aneurysm depth to aneurysm neck width) for predicting an aneurysmal rupture. The aspect ratio is considered a better geometric index than aneurysm size for determining the intra-aneurysmal blood flow. METHODS: We measured the aspect ratios and the sizes of aneurysms, as determined by examining angiographic films magnified 1.4x, in 129 patients with ruptured aneurysms and in 72 patients with 78 unruptured aneurysms. After categorizing the aneurysms into four groups on the basis of their locations (aneurysms of the anterior communicating artery, middle cerebral artery, internal carotid artery-posterior communicating artery [ICA-PComA], and other aneurysms), a statistical analysis of ruptured and unruptured aneurysms was performed. RESULTS: The mean aneurysm size was found to be statistically significant in the aneurysms at the ICA-PComA and in locations excluding the anterior communicating artery, the middle cerebral artery, and the ICA-PComA. However, the mean aspect ratio was statistically significant at all four locations. In patients with ruptured aneurysms, no ruptured aneurysms with an aspect ratio of less than 1.0 were found. The distribution of the ruptured group versus the unruptured group with an aspect ratio of less than 1.6 at each location was 13 versus 79%, respectively, at the anterior communicating artery, 11 versus 58% at the middle cerebral artery, 11% versus 85% at the ICA-PComA, and 7 versus 81% at other locations. CONCLUSION: The aspect ratio between ruptured aneurysms and unruptured aneurysms was found to be statistically significant, and almost 80% of the ruptured aneurysms showed an aspect ratio of more than 1.6, whereas almost 90% of the unruptured aneurysms showed an aspect ratio of less than 1.6. This study therefore suggests that the aspect ratio may be useful in predicting imminent aneurysmal ruptures. PMID- 11270539 TI - Cerebral hemodynamic parameters for patients with neurological improvements after extracranial-intracranial arterial bypass surgery: evaluation using positron emission tomography. AB - OBJECTIVE: The purpose of this study was to clarify the hemodynamic features of patients who experienced improved neurological function after extracranial intracranial arterial bypass surgery. With this aim, we retrospectively analyzed the results of their pre- and postoperative positron emission tomographic studies. METHODS: This study included 16 patients who exhibited stable neurological dysfunction just before extracranial-intracranial bypass surgery. All underwent pre- and postoperative positron emission tomographic studies. They were divided into groups, i.e., patients who did (Group 1, n = 6) or did not (Group 2, n = 10) manifest postoperative improvements in neurological functions. Positron emission tomographic parameters obtained in the middle cerebral artery territories were compared between the two groups. RESULTS: Comparison of the preoperative hemodynamic values on the affected side and the contralateral side demonstrated that the mean regional cerebral blood flow values were significantly lower on the affected side in both groups (Group 1, P < 0.005; Group 2, P < 0.05). For Group 1 patients, the mean regional oxygen extraction fraction (rOEF) and regional cerebral blood volume values were significantly higher on the affected side than on the contralateral side (P < 0.01 and P < 0.05, respectively). For Group 2 patients, the mean regional cerebral metabolic rate of oxygen (rCMRO2) value was significantly lower on the affected side than on the contralateral side (P < 0.05). The mean rOEF and rCMRO2 values on the affected side were significantly higher for Group 1 patients, compared with Group 2 patients, before surgery (P < 0.05 and P < 0.05, respectively). The preoperative regional cerebral blood flow and regional cerebral blood volume values on the affected side were similar for the two groups. Postoperative changes in mean regional cerebral blood flow and mean rOEF on the affected side were statistically significant for both groups. The mean rCMRO2 on the affected side for Group 2 was significantly lower than that for Group 1, even after bypass surgery (P < 0.05). CONCLUSION: Bypass surgery may improve neurological function for patients with significantly elevated rOEF values and rCMRO2 values near the normal level. These hemodynamic parameters may be useful for the identification of candidates for extracranial-intracranial bypass surgery. PMID- 11270540 TI - Spontaneous intracranial hypotension mimicking aneurysmal subarachnoid hemorrhage. AB - OBJECTIVE: An excruciating headache of instantaneous onset is known as a thunderclap headache. A subarachnoid hemorrhage is the prototypical cause, but other serious disorders may also present with a thunderclap headache, including cerebral venous sinus thrombosis, carotid artery dissection, and pituitary apoplexy. We report a group of patients with thunderclap headaches as the initial manifestation of spontaneous intracranial hypotension caused by a spinal cerebrospinal fluid leak. METHODS: Among 28 patients with spontaneous intracranial hypotension due to a documented spinal cerebrospinal fluid leak, four (14%) initially experienced an excruciating headaches of instantaneous onset. RESULTS: The mean age of the four patients (two men and two women) was 35 years (range, 24-45 yr). Nuchal rigidity was present in the three patients who sought early medical attention, and they underwent emergency computed tomographic scanning, lumbar puncture, and cerebral angiography to rule out an aneurysmal subarachnoid hemorrhage. The delay between the onset of headache and diagnosis of intracranial hypotension ranged from 4 days to 5 weeks. A fourth patient did not seek medical attention until 1 month after the ictus. CONCLUSION: Spontaneous intracranial hypotension should be included in the differential diagnosis of thunderclap headache, even when meningismus is present. PMID- 11270541 TI - Management of malignant pineal germ cell tumors with residual mature teratoma. AB - OBJECTIVE: The treatment of intracranial mixed germ cell tumors presents a unique challenge, since eradication of malignant tumor by radiation and/or chemotherapy may spare the benign tumor component. We reviewed our surgical experience with residual malignant pineal germ cell tumors after neoadjuvant therapy. METHODS: Between 1987 and 1997, 16 patients with malignant intracranial germ cell tumors were treated at the Mayo Clinic with a protocol of neoadjuvant chemotherapy and radiation therapy. After the diagnosis was confirmed by histopathological examination, all patients were treated with four cycles of etoposide and cisplatin as well as external beam radiation therapy (range, 3030-5940 cGy). Six patients had an incomplete response to therapy, as demonstrated by observation of residual tumor on magnetic resonance imaging scans. Initial pathology in these six patients was germinoma in four and combinations of yolk sac tumor, embryonal carcinoma, malignant teratoma, and germinoma in two. Two patients had synchronous pineal and suprasellar tumors, with leptomeningeal dissemination. Tumor markers were elevated in four of the six patients at presentation. RESULTS: All patients with residual pineal tumors underwent surgical resection via an infratentorial, supracerebellar approach. Pathological examination revealed mature teratoma in five patients and amorphous debris in one patient. No patient had recurrent malignancy. Significant neurological morbidity occurred in one patient, with no mortality. At a mean follow-up of 23 months, no recurrence on magnetic resonance imaging has been documented. CONCLUSION: Residual pineal tumor occurring after treatment of malignant intracranial germ cell tumor with neoadjuvant therapy is likely to be mature teratoma. Operative resection of these benign recurrences is safe and effective. PMID- 11270542 TI - Percutaneous controlled radiofrequency trigeminal rhizotomy for the treatment of idiopathic trigeminal neuralgia: 25-year experience with 1,600 patients. AB - OBJECTIVE: The objective of this study was to evaluate the effectiveness of percutaneous, controlled radiofrequency trigeminal rhizotomy (RF-TR). The outcome of 1,600 patients with idiopathic trigeminal neuralgia after RF-TR was analyzed after a follow-up period of 1 to 25 years. METHODS: A total of 1,600 patients with idiopathic trigeminal neuralgia underwent 2,138 percutaneous radiofrequency rhizotomy procedures between 1974 and 1999. Sixty-seven patients had bilateral idiopathic trigeminal neuralgia, and 36 of them were treated with bilateral RF TR; 1,216 patients (76%) were successfully managed with a single procedure, and the remainder were treated with multiple procedures. Benzodiazepines and narcotic analgesics were used for anesthesia because patient cooperation during the procedures was essential so that the physician could create selective, controlled lesions. RESULTS: The average follow-up time was 68.1 +/- 66.4 months (range, 12 300 mo). Acute pain relief was accomplished in 97.6% of patients. Complete pain relief was achieved at 5 years in 57.7% of the patients who underwent a single procedure. Pain relief was reported in 92% of patients with a single procedure or with multiple procedures 5 years after the first rhizotomy was performed. At 10 year follow-up, 52.3% of the patients who underwent a single procedure and 94.2% of the patients who underwent multiple procedures had experienced pain relief; at 20-year follow-up, 41 and 100% of these patients, respectively, had experienced pain relief. No mortalities occurred. After the first procedure was performed, early pain recurrence (<6 mo) was observed in 123 patients (7.7%) and late pain recurrence was observed in 278 patients (17.4%). Complications included diminished corneal reflex in 91 patients (5.7%), masseter weakness and paralysis in 66 (4.1%), dysesthesia in 16 (1 %), anesthesia dolorosa in 12 (0.8%), keratitis in 10 (0.6%), and transient paralysis of Cranial Nerves III and VI in 12 (0.8%). Permanent Cranial Nerve VI palsy was observed in two patients, cerebrospinal fluid leakage in two, carotid-cavernous fistula in one, and aseptic meningitis in one. CONCLUSION: Percutaneous, controlled RF-TR represents a minimally invasive, low-risk technique with a high rate of efficacy. The procedure may safely be repeated if pain recurs. PMID- 11270543 TI - Concepts and methods in chronic thalamic stimulation for treatment of tremor: technique and application. AB - OBJECTIVE: To rationalize the technique and reduce the costs associated with chronic deep brain stimulation of the thalamus for treatment of refractory tremor. METHODS: The efficacy and safety of a modification in surgical techniques was prospectively assessed in 94 patients with tremor. Bilateral electrodes were implanted in 29 patients, and 65 patients received unilateral implants. Forty five patients had Parkinson's disease tremor, 42 patients had essential tremor, and 7 patients had kinetic tremors of different causes. In all instances, intraoperative stimulations to analyze the thresholds of intrinsic and extrinsic responses were performed directly with the implanted leads. The electrodes were repositioned until satisfactory results were achieved. The pulse generators were implanted directly after the first step in the same operative session. Patients were not subjected to interoperative test stimulation trials. RESULTS: Postoperative improvement of tremor at a mean follow-up of 11.9 months was rated as excellent in 47 patients (50%), marked in 37 patients (39%), moderate in 8 patients (9%), and minor in 2 patients (2%). There was no persistent morbidity related to surgery. In patients with Parkinson's disease, the symptomatic improvement of tremor was rated as excellent in 51% of patients, marked in 36%, moderate in 11%, and minor in 2%. In patients with essential tremor, symptomatic outcome was classified as excellent in 57% of patients, marked in 36%, moderate in 5%, and minor in 2%. Six of the seven patients with kinetic tremor achieved marked symptomatic improvement, and one patient experienced moderate improvement. Forty patients experienced stimulation-related side effects. Side effects were mild in general, and they were reversible with a change in electrical parameters. They occurred more frequently in patients who had bilateral stimulation. CONCLUSION: Excellent to marked improvement of tremor is achieved in the majority of patients with physiological target determination via implanted leads in thalamic deep brain stimulation. Interoperative test stimulation trials are unnecessary. Modifications in technique may help to reduce the costs of the related hospital stay. PMID- 11270544 TI - Assessments of axial motor control during deep brain stimulation in parkinsonian patients. AB - OBJECTIVE: We tested the hypothesis that bilateral deep brain stimulation (DBS) in the globus pallidus internus or the subthalamic nucleus improves various components of postural and oromotor function and that some of the components correlate with changes in the Unified Parkinson's Disease Rating Scale (UPDRS) in patients with Parkinson's disease. METHODS: Six patients with Parkinson's disease were evaluated for four postural and two orofacial UPDRS items, and quantitative tests of posture adjustments and oromotor control were performed while the patients were on and off DBS. Measurements of postural adjustments included reactive force and latency before a voluntary step. The oromotor assessments involved velocity and amplitude changes during voluntary jaw movement. RESULTS: DBS significantly improved the total UPDRS motor score by an average of 44%, which included improvement of 18 to 54% in the postural and orofacial items. DBS also decreased foot lift-off latency significantly, but it produced a variable response to the preparatory postural force in the swing limb. DBS significantly improved jaw-opening velocity by 14 to 50% and jaw opening amplitude by 5 to 41%. Significant correlations for the percentage change from off and on DBS occurred among a few UPDRS items and foot lift-off latency and jaw-opening velocities. CONCLUSION: DBS in either the globus pallidus internus or the subthalamic nucleus induces improvements in bradykinesia of specific components of postural and oromotor control, which also can be measured by the postural and orofacial UPDRS items. In some Parkinson's disease patients, DBS results in improvements in force or amplitude control, although these changes are not reflected in changes in UPDRS postural and orofacial items. A battery of quantitative and clinical tests must be used to evaluate the effects of DBS on axial motor control adequately. PMID- 11270545 TI - Retrosigmoid approach to acoustic neuroma (vestibular schwannoma). AB - The retrosigmoid approach for the microsurgical removal of an acoustic neuroma (vestibular schwannoma) is described, and perioperative medical management of the patient is discussed. The techniques for monitoring facial and cochlear nerve function are presented. The supine-oblique position, skin incision, bone removal, dural opening, and initial exposure are outlined. Important points in the technique for removing acoustic neuromas and preserving hearing, when possible, are described and illustrated. PMID- 11270546 TI - Image-guided transorbital roof craniotomy via a suprabrow approach: a surgical series of 72 patients. AB - OBJECTIVE: Many subfrontal and orbitofrontal craniotomy techniques have been developed. We present our results with the transorbital roof craniotomy, a frontal craniotomy that incorporates the orbital roof and is performed via a suprabrow incision. This technique was used in 72 patients, primarily for tumor resection. METHODS: Charts were retrospectively reviewed for all patients undergoing transorbital procedures. A total of 72 patients underwent 82 transorbital craniotomies from September 1995 to July 1999. The primary indication for the transorbital approach was mass lesion of the orbit, anterior fossa, or parasellar region. RESULTS: A total of 47 women and 25 men with a mean age of 53 years underwent 82 procedures. The primary pathological finding was meningioma, which occurred in 40 patients (55.6%), followed by craniopharyngioma (6.9%), pituitary macroadenoma (6.9%), schwannoma (5.5%), and hemangioma (5.5%). Simpson Grade I or II resection was achieved in 54% of patients, with Simpson Grade III to V resection achieved in the remaining 46%. Forty-one patients presented with visual loss in 43 cases, with 44.2% experiencing postoperative visual improvement, 46.5% remaining unchanged, and 9.3% worsening. Overall morbidity was 18.4%, with cerebrospinal fluid leak being the most common complication (6.6%). No patients died. CONCLUSION: The transorbital roof craniotomy is an evolutionary approach that provides excellent exposure to the orbit, anterior fossa, and parasellar region with little significant morbidity and, in our series, no mortality. Although we have used this approach primarily for resection of mass lesions, future directions for this procedure will likely lie in treating vascular lesions and lesions of the interpeduncular fossa. PMID- 11270547 TI - Transforaminal lumbar interbody fusion: technique, complications, and early results. AB - OBJECTIVE: To demonstrate the safety, surgical efficacy, and advantages of the transforaminal approach for lumbar interbody fusion when combined with pedicle screw fixation. METHODS: We retrospectively reviewed the records of 22 patients (age range, 34-63 yr; mean, 49 yr) with Grade I or II spondylolisthesis who underwent transforaminal lumbar interbody fusion. Nineteen patients presented with low back pain and associated radiculopathy, and three presented with low back pain only. Transforaminal lumbar interbody fusion was performed at L4-L5 in 8 patients, L5-S1 in 11 patients, L3-L4 and L4-L5 in 2 patients, and L4-L5 and L5 S1 in 1 patient. Periodic follow-up took place 1 to 12 months after surgery (mean, 5.3 mo). Decompression is performed according to clinical circumstances. Pedicle screws are placed, and a discectomy is carried out. The cartilaginous endplates are removed. The interspace is gradually distracted, resulting in lost disc height being regained, and interbody fusion cages are positioned. The pedicle screw-and-rod construct is then compressed, restoring lumbar lordosis. RESULTS: Low back pain completely resolved in 16 patients, moderate relief from pain was achieved in 5 patients, and the pain was unchanged in one patient. Nonneurological complications included intraoperative durotomy in one patient and postoperative wound infection in two. In one patient, postoperative mild L5 motor paresis resolved. One patient had a temporary brachial plexopathy due to intraoperative positioning, and one patient had peripheral polyneuropathy secondary to prolonged intraoperative blood pressure cuff inflation. CONCLUSION: Transforaminal lumbar interbody fusion is a safe and effective method for achieving circumferential spinal fusion via a single-stage procedure. This procedure is particularly useful in restoring disc space height and lumbar lordosis. PMID- 11270548 TI - Computer-assisted visualization of arteriovenous malformations on the home personal computer. AB - OBJECTIVE: Arteriovenous malformations (AVMs) are difficult lesions to treat, partly because it is difficult to formulate a three-dimensional mental image of the nidus and its supplying arteries, draining veins, and arteries of passage. Our purpose is to develop personal computer software that allows better visualization of complex, three-dimensional, connected vascular anatomy for surgical planning. METHODS: Vessels are defined from magnetic resonance angiograms and are symbolically linked to form vascular trees. The nidus of the AVM is also defined by magnetic resonance angiography. These representations of the nidus and vasculature can be viewed together in a software program that allows the user to color-code groups of vessels or to selectively turn connected groups of vessels "off" to avoid obscuring the part of the image that the user wants to observe. Structures can be viewed from any angle. The vessels can also be shown intersecting any magnetic resonance angiogram slice or superimposed upon digital subtraction angiograms obtained from the same patient. RESULTS: We report results from two patients with AVMs in which our representations were compared with the findings during surgery. Our three-dimensional vascular trees correctly depicted the relationship of the nidus to feeding vessels in three dimensions. We show findings in an additional, unoperated patient for whom vessel trees were created from three-dimensional digital subtraction angiography data and compared with a volume rendering of the original data set. CONCLUSION: Computer-assisted, three-dimensional visualizations of complex vascular anatomy can be helpful in planning the surgical excision of AVMs. Software programs that produce these images can provide important information that is difficult to obtain by traditional techniques. This imaging method is also applicable to guidance of endovascular procedures and removal of complex tumors. PMID- 11270549 TI - Robot-assisted microsurgery: a feasibility study in the rat. AB - OBJECTIVE: Telerobotic surgery is a novel technology that can improve a surgeon's manual dexterity as well as the results achieved with microsurgical procedures. METHODS: A prototype Robot-Assisted MicroSurgery (RAMS) microdexterity enhancement system developed by the Jet Propulsion Laboratory and MicroDexterity Systems, Inc., was tested in 10 rats. Carotid arteriotomies were created and closed using either the RAMS system or conventional microsurgical techniques. The time required, the technical quality (vessel patency and suture line integrity), the error rate, and subjective difficulty were compared. RESULTS: All procedures were successfully completed using the RAMS system to manipulate the vessel but not to hold the needle or place the sutures. The precision, technical quality, and error rate of telerobotic surgery were similar to those of conventional techniques. However, the use of the RAMS system was associated with a twofold increase in the length of the procedure. CONCLUSION: Surgery using a microdexterity enhancement system, or RAMS prototype, is feasible. With further development, such as a stereotelevisualization and haptic feedback system, this system could be used for telerobotic surgery in neurosurgical practice. PMID- 11270550 TI - MIB-1 staining index of pediatric meningiomas. AB - OBJECTIVE: For adult meningiomas, the staining index (SI) for the anti-Ki-67 monoclonal antibody MIB-1 is well correlated with histological atypia and tumor recurrence. MIB-1 SIs for meningiomas in the pediatric population have not been previously reported. Meningiomas tend to be more histologically aggressive and to recur more frequently in children, compared with adults. The objectives of this study were to determine whether MIB-1 SIs are correlated with pathological atypia and recurrence among pediatric meningiomas and to compare the MIB-1 SIs of pediatric meningiomas with those of adult meningiomas. METHODS: MIB-1 SIs were assessed on paraffin-embedded sections of 14 pediatric meningiomas (patient age, 2-17 yr), 5 of which contained atypical or malignant features. For comparison with benign pediatric meningiomas, MIB-1 SIs were also assessed on paraffin embedded sections of 14 adult meningiomas (patient age, 38-90 yr), none of which displayed atypical or malignant features or recurred within a 5-month median follow-up period. RESULTS: MIB-1 SIs of pediatric meningiomas ranged from 1.2 to 31.6% (median, 9.1%). Significant differences were observed between the MIB-1 SIs for tumors with atypical or malignant features (median, 12.3%; range, 7.0-31.6%) and those for tumors without atypia (median, 7.0%; range, 1.2-12.6%; P = 0.045). There were six recurrences after gross total resection, during a 36.5-month median follow-up period. All five of the tumors with pathological atypia recurred; one tumor without atypia recurred. Significant differences were observed between MIB-1 SIs for nonrecurrent tumors (median, 6.6%; range, 1.2 12.2%) and those for recurrent tumors (median, 12.5%; range, 7.0-31.6%; P = 0.012). The median MIB-1 SI for adult control specimens was 8.8% (range, 1.2 19.3%), which did not differ significantly from that for pediatric meningiomas without atypia (P = 0.68). CONCLUSION: For this cohort of pediatric meningiomas, pathological atypia and the tendency to recur were correlated with elevated MIB-1 SIs. The median MIB-1 SI for pediatric meningiomas without histological atypia did not differ significantly from that for adult meningiomas without atypia, suggesting that the more aggressive clinical features of meningiomas in children may be attributable to factors other than the rate of cellular proliferation. PMID- 11270551 TI - Cathepsins B and L are markers for clinically invasive types of meningiomas. AB - OBJECTIVE: Meningiomas are benign neoplasms that derive from coverings of the brain. Approximately 10% of benign tumors progress into atypical, malignant tumors, thus constituting a subset of histopathologically benign tumors that are clinically invasive. The aim of this study was to evaluate cathepsins B and L and their inhibitors as new prognostic factors that could distinguish malignant from benign forms of meningiomas. METHODS: Using immunohistochemical analysis and specific monoclonal antibodies, we evaluated the levels of cathepsins B and L and the levels of the endogenous cysteine proteinase inhibitors stefin A and cystatin C in 88 meningiomas. Immunohistochemical scores were determined as the sum of the frequency (0-3) and intensity (0-3) of immunolabeling of the tumor cells. RESULTS: Of the 88 tumors studied, 67 were benign meningiomas and 21 were atypical meningiomas. Among the benign group, nine tumors had certain features of malignancy. These tumors were classified as border benign meningiomas, and the rest were classified as clear benign meningiomas. A high immunohistochemical score (4-6) for cathepsin B was more frequent in atypical tumors than in clear benign tumors (P < 0.001). Compared with clear benign tumors, higher cathepsin B immunohistochemical scores were found in atypical tumors (P < 0.001) and border benign tumors (P < 0.03). No statistical difference in immunohistochemical staining of cathepsin B was found between atypical meningiomas and border benign meningiomas. Higher expression of cathepsin L was found in atypical tumors as compared with clear benign tumors (P < 0.03), but it was not observed in border benign as compared with clear benign meningiomas. No immunostaining for stefin A and cystatin C was detected in any of the tumors. CONCLUSION: We show that the levels of cathepsin B and cathepsin L antigens are significantly higher in invasive types of benign meningioma. Specifically, cathepsin B may be used as a diagnostic marker to distinguish histomorphologically benign but invasive meningiomas from histomorphologically clear benign tumors. PMID- 11270552 TI - Intratumoral injection of lipopolysaccharide causes regression of subcutaneously implanted mouse glioblastoma multiforme. AB - OBJECTIVE: Anecdotal reports documented extended survival times for patients who developed infections at the site of resection of malignant gliomas. Hypothesized mechanisms for this phenomenon include immune responses triggered by lipopolysaccharide (LPS). This investigation assessed whether LPS could produce tumor regression in an in vivo model of malignant glioma. METHODS: Delayed brain tumor cells (2 x 10(6)) were injected subcutaneously into female BALB/c mice. LPS (300-500 microg) was injected intratumorally or subcutaneously at a contralateral site on Days 10, 17, and 24. Control animals received phosphate-buffered saline intratumorally or subcutaneously. Mice were killed on Day 28, and tumors were removed. Mean tumor masses for control animals and the two LPS-treated groups (intratumoral or contralateral subcutaneous treatment) were compared. Histological assessments of treated and control tumors were performed. RESULTS: Complete or nearly total tumor regression was achieved in all 20 mice with subcutaneous delayed brain tumor cell tumors treated intratumorally with 400 microg of LPS (mean tumor mass of 0.09 +/- 0.38 g versus 2.42 +/- 2.46 g for control animals, P < 0.0001). Intratumoral administration of 300 microg of LPS or subcutaneous injection of 300 or 400 microg of LPS at a contralateral site resulted in less consistent regression of subcutaneous tumors. Administration of 500 microg of LPS resulted in tumor regression similar to that observed with lower doses but was limited by treatment-related deaths in 40% of animals. Histological assessment revealed lymphocytic infiltration of LPS-treated tumors. CONCLUSION: Intratumoral injections of LPS caused dramatic regression of subcutaneously implanted delayed brain tumor cell mouse gliomas. Investigation of this antitumoral effect may improve treatment responses for patients with malignant gliomas. PMID- 11270553 TI - Therapeutic effects of sodium butyrate on glioma cells in vitro and in the rat C6 glioma model. AB - OBJECTIVE: Preliminary in vitro studies have indicated that sodium butyrate inhibits the proliferation of cultured glioma cells and induces cellular differentiation, making it potentially useful as a therapeutic agent for patients with glioblastoma multiforme. The purpose of this study was to expand on the preliminary research by investigating the effects of sodium butyrate on multiple cell lines, explanted cells from glioblastoma tumor specimens, and in vivo in the rat C6 glioma brain tumor model. METHODS: Four malignant glioma cell lines (A 172, T98G, U118MG, and C6) and two primary cell cultures derived from human glioblastoma tumor specimens were treated with 2 mmol/L sodium butyrate for up to 72 hours. Sodium butyrate-induced effects on cell morphology, proliferation, cell cycle distribution, migration, glial fibrillary acidic protein staining, and S100beta protein content were determined. For in vivo studies, a total of 64 male Wistar-Furth rats underwent operations to implant C6 glioma cells stereotactically or were used as controls. The rats were treated with escalating doses of sodium butyrate by microinfusion with Alzet minipumps (Durect Corp., Cupertino, CA). RESULTS: Sodium butyrate treatment in vitro produced changes in morphology and glial fibrillary acidic protein expression indicative of cellular differentiation. In cell lines and explanted cells, sodium butyrate consistently inhibited glioblastoma cell proliferation (to 51 +/- 6% that of controls) and migration (to 46 +/- 17%). Intratumoral infusion of 40 mmol/L sodium butyrate prolonged the survival of Wistar-Furth rats with intracerebral C6 tumors (P = 0.013) without detectable toxicity. CONCLUSION: These data support further consideration of direct interstitial infusion of sodium butyrate in a Phase I clinical study for patients with recurrent glioblastoma multiforme. PMID- 11270554 TI - Overexpression of cyclooxygenase-2 in rabbit basilar artery endothelial cells after subarachnoid hemorrhage. AB - OBJECTIVE: We investigated the expression in rabbit basilar arteries of cyclooxygenase (COX)-2, which is the inducible isoform of the enzyme of prostaglandin (PG) production, and the concentrations of the proinflammatory cytokine tumor necrosis factor alpha (TNFalpha) and representative PGs in the cerebrospinal fluid (CSF) after experimental subarachnoid hemorrhage (SAH). METHODS: Seven sets of basilar arteries were removed from control rabbits and from rabbits killed 1 and 3 days after induced SAH. The arteries were subjected to identical simultaneous immunolabeling for examination with a confocal microscope. One-half of each artery was stained for the constitutive form COX-1 and the other half for COX-2. CSF was sampled in control animals and at 6 hours, 1 day, and 3 days for assays of TNFalpha, PGE2, and 6-keto-PGF1 (metabolite of PGI2). RESULTS: COX-1 immunoreactivity in the endothelial layer was similar in the three groups. Weak endothelial COX-2 immunoreactivity was found in arteries of control animals. COX-2 staining was higher in the group killed at 3 days compared with the control group (P < 0.05). The levels of PGE2 and 6-keto PGF1alpha in the CSF peaked significantly at 6 hours, then decreased at 3 days to the basal level (PGE2) or significantly lower (6-keto-PGF1). TNFalpha was undetectable in control CSF, significantly higher (P < 0.001) at 6 hours, and undetectable at 3 days. CONCLUSION: After SAH, endothelial COX-1 immunoreactivity does not change, whereas overexpression of COX-2 occurs at 3 days. This induction does not seem linked to TNFalpha production, nor is it responsible for early raised levels of PGE2 and PGI2 in the CSF. We suggest that the role of induced COX-2 may be to modify gene expression and hence alter the properties of the vessel wall after SAH. PMID- 11270555 TI - A spinal thecal sac constriction model supports the theory that induced pressure gradients in the cord cause edema and cyst formation. AB - OBJECTIVE: Spinal cord cysts are a devastating condition that occur secondary to obstructions of the spinal canal, which may be caused by congenital malformations, trauma, spinal canal stenosis, tumors, meningitis, or arachnoiditis. A hypothesis that could explain how spinal cord cysts form in these situations has been presented recently. Therefore, a novel spinal thecal sac constriction model was implemented to test various aspects of this hypothesis. METHODS: Thecal sac constriction was achieved by subjecting rats to an extradural silk ligature at the T8 spinal cord level. Rats with complete spinal cord transection served as a second model for comparison. The animals underwent high-resolution magnetic resonance imaging and histological analysis. RESULTS: Thecal sac constriction caused edema cranial and caudal to the ligation within 3 weeks, and cysts developed after 8 to 13 weeks. In contrast, cysts in rats with spinal cord transection were located predominantly in the cranial spinal cord. Histological sections of spinal cords confirmed the magnetic resonance imaging results. CONCLUSION: Magnetic resonance imaging provided the specific advantage of enabling characterization of events as they occurred repeatedly over time in the spinal cords of individual living animals. The spinal thecal sac constriction model proved useful for investigation of features of the cerebrospinal fluid pulse pressure theory. Edema and cyst distributions were in accordance with this theory. We conclude that induced intramedullary pressure gradients originating from the cerebrospinal fluid pulse pressure may underlie cyst formation in the vicinity of spinal canal obstructions and that cysts are preceded by edema. PMID- 11270556 TI - Psychosurgery: a historical overview. AB - Neurosurgical treatment for psychiatric disorders has a long and controversial history. From the Stone Age use of trephining to release the demons of the spirit to the millimeter accuracy of stereotactic instruments currently used in the operating room, psychosurgery has enjoyed enthusiastic support as well as experiencing scorn. Today, psychosurgery is a minimally invasive and highly selective treatment that is performed for only a few patients with severe, treatment-refractory, affective, anxiety, or obsessive-compulsive disorders. Recent advances in technology and functional neuroanatomic techniques, as well as economic pressures to decrease the costs of caring for chronically ill patients, may provide an opportunity for psychosurgery to become a more attractive option for the treatment of psychiatric diseases. In this historical overview, the rise and fall of psychosurgery are described. A better understanding of the colorful history of this interesting topic should enable modern neurosurgeons and other health care professionals to meet the social, ethical, and technical challenges that are sure to lie ahead. PMID- 11270557 TI - Leeches for the unfortunate locksmith: self-inflicted posttraumatic transient cerebral blindness--mode of treatment and underlying mechanism (1826). AB - In 1826, Jean-Pierre Gama, a French military surgeon, treated a young locksmith who had self-inflicted posttraumatic transient cortical blindness. This may be the earliest detailed, firsthand description of this condition by a medically and scientifically trained observer. Gama's report sheds light on the concept of the mechanism of coup-contrecoup of cerebral concussion and its treatment in the early 19th century and on the germinating discipline of cerebral localization. PMID- 11270558 TI - Spontaneous resolution of Chiari I malformation and syringomyelia: case report and review of the literature. AB - OBJECTIVE AND IMPORTANCE: Indications for surgery and the surgical technique of foramen magnum decompression for patients with Chiari I malformation and syringomyelia are controversial issues. This case report supports the view that observation may be adequate for patients without progressive symptoms or with mild clinical symptoms. CLINICAL PRESENTATION: A 37-year-old woman presented with a 3-month history of burning dysesthesias and hypesthesia in her right arm. A neurological examination revealed hypesthesia in the right trigeminal distribution. A magnetic resonance imaging scan revealed a Chiari I malformation with syringomyelia between C2 and T2. No hydrocephalus was observed. CLINICAL COURSE: Because the patient's symptoms regressed spontaneously, surgery was not performed. Thirty-two months after her initial examination, the patient was asymptomatic. A second magnetic resonance imaging scan was obtained, which demonstrated complete spontaneous resolution of the Chiari I malformation and syringomyelia. CONCLUSION: We attribute the regression of the patient's symptoms to spontaneous recanalization of cerebrospinal fluid pathways at the foramen magnum, which most likely was due to rupture of the arachnoid membranes that had obstructed cerebrospinal fluid flow. PMID- 11270559 TI - A posterior tibial nerve neurilemoma unrecognized for 10 years: case report. AB - OBJECTIVE AND IMPORTANCE: Neoplasms of peripheral nerves can be obscured, especially during the early phase. The author reports a patient with a posterior tibial nerve neurilemoma (schwannoma). For a decade, the tumor was misdiagnosed as nonspecific S1 radiculopathy and psychogenic chronic pain syndrome. The patient's presentation and initial management are unique. CLINICAL PRESENTATION: A 40-year-old woman reported severe left foot and calf pain, numbness, and weakness. The symptoms were evident during three pregnancies, and they gradually progressed. The neuropathic pain was protracted, despite implantation of a dorsal column stimulator and administration of a wide variety of medications and therapies. The symptoms were unresponsive to both inpatient and outpatient treatments, which resulted in a misdiagnosis of psychogenic pain for more than a decade. Diagnostic scans obtained by computed tomography, ultrasonography, and nuclear scintigraphy confirmed a popliteal fossa mass. INTERVENTION: A high, large posterior tibial nerve neurilemoma was found intraoperatively, positioned just below the sciatic nerve bifurcation with extensive degenerative features and hemorrhages. Surgical resection provided immediate recovery. CONCLUSION: Peripheral nerve tumors are rarely acknowledged clinical entities. Chronic unexplained foot and calf pain and a positive Tinel's sign should raise suspicion of posterior tibial nerve neurilemoma. Even in patients who have had such tumors for a decade, surgical resection remains the treatment of choice. PMID- 11270560 TI - Cord compression secondary to cervical disc herniation associated with calcification of the ligamentum flavum: case report. AB - OBJECTIVE AND IMPORTANCE: Calcification of the ligamentum flavum is a rare disease that occurs almost exclusively in elderly Japanese people. We report the case of a young Caucasian woman who presented with a C5-C6 disc herniation associated with a cervical calcified ligamentum flavum. CLINICAL PRESENTATION: The patient presented with a cord compression syndrome of 76 hours' evolution. At exploration, a Brown-Sequard syndrome at the C6 level was found. Magnetic resonance imaging and computed tomography led to a correct diagnosis and planning for decompression. INTERVENTION: We operated on the patient through a combined anterior and posterior approach. After the patient underwent anterior discectomy with intersomatic arthrodesis, we performed posterior decompression. During the operation, we observed that the dura mater could not be separated from the ligamentum, so an en bloc excision of both structures was performed. Microscopic examination indicated that the excised ligamentum had calcification, and total integration of the dura mater into the structure of the ligamentum was demonstrated. To our knowledge, this circumstance has never been described before. A posterior C3-C7 arthrodesis was performed to prevent postoperative kyphosis. Recovery was successful, with total recovery from neurological deficits 4 months later. CONCLUSION: Calcification of the ligamentum flavum is a progressive disease that starts early in life and becomes symptomatic later in life when spinal stenosis occurs. Magnetic resonance imaging and computed tomography provide adequate diagnosis and allow proper surgical planning for decompression. The presence of hyperintense areas within the spinal cord parenchyma, in the absence of a traumatic antecedent, does not preclude a complete recovery. PMID- 11270561 TI - Intrasellar rhabdomyosarcoma: case report. AB - OBJECTIVE AND IMPORTANCE: There has been only one reported case of an intrasellar rhabdomyosarcoma, the origin of which was in the para-nasal sinus. The authors encountered a patient with an intrasellar rhabdomyosarcoma with no evidence of tumor at any additional sites. CLINICAL PRESENTATION: A 28-year-old otherwise healthy man complaining of headache exhibited left abducent nerve palsy and left visual disturbance. The patient was diagnosed as having a sellar tumor invading the left cavernous sinus. INTERVENTION: Near total removal of the tumor was achieved via a trans-sphenoidal approach. Histologically, the tumor was composed of small, round-to-elongated undifferentiated cells and large spindle cells with myoblastic features. Immunohistochemically, tumor cells were positive for antibodies against desmin, myoglobin, and alpha-smooth muscle actin. The tumor was identified as an embryonal rhabdomyosarcoma on the basis of the above pathological findings. Systemic investigation, including the nasal and para-nasal regions, failed to detect any additional tumors. Postoperative local radiation therapy and chemotherapy with the use of ifosfamide, etoposide, and vincristine brought about complete initial remission. CONCLUSION: Rhabdomyosarcoma should be considered in the differential diagnosis of a primary intrasellar neoplasm. PMID- 11270562 TI - Chronic motor cortex stimulation for phantom limb pain: a functional magnetic resonance imaging study: technical case report. AB - OBJECTIVE AND IMPORTANCE: Chronic motor cortex stimulation has provided satisfactory control of pain in patients with central or neuropathic trigeminal pain. We used this technique in a patient who experienced phantom limb pain. Functional magnetic resonance imaging (fMRI) was used to guide electrode placement and to assist in understanding the control mechanisms involved in phantom limb pain. CLINICAL PRESENTATION: A 45-year-old man whose right arm had been amputated 2 years previously experienced phantom limb pain and phantom limb phenomena, described as the apparent possibility of moving the amputated hand voluntarily. He was treated with chronic motor cortex stimulation. INTERVENTION: Data from fMRI were used pre- and postoperatively to detect shoulder and stump cortical activated areas and the "virtual" amputated hand cortical area. These sites of preoperative fMRI activation were integrated in an infrared-based frameless stereotactic device for surgical planning. Phantom limb virtual finger movement caused contralateral primary motor cortex activation. Satisfactory pain control was obtained; a 70% reduction in the phantom limb pain was achieved on a visual analog scale. Postoperatively and under chronic stimulation, inhibiting effects on the primary sensorimotor cortex as well as on the contralateral primary motor and sensitive cortices were detected by fMRI studies. CONCLUSION: Chronic motor cortex stimulation can be used to relieve phantom limb pain and phantom limb phenomena. Integrated by an infrared-based frameless stereotactic device, fMRI data are useful in assisting the neurosurgeon in electrode placement for this indication. Pain control mechanisms and cortical reorganization phenomena can be studied by the use of fMRI. PMID- 11270563 TI - Laboratory evaluation of the phoenix CRx diamond valve. AB - OBJECTIVE: To assess the long-term hydrodynamic properties of a new cerebrospinal fluid flow-regulating hydrocephalus shunt called the CRx Diamond valve (Phoenix Biomedical Corp., Valley Forge, PA). METHODS: Three samples of a Diamond valve were tested in the United Kingdom Shunt Evaluation Laboratory during a 40-day period. Tests were performed for long-term pressure-flow performance, overdrainage, susceptibility to ambient temperature changes, external pressure, reflux, presence of small particles in the reagent, mechanical durability, and magnetic resonance imaging compatibility. RESULTS: Tests demonstrated that the Diamond valve stabilized flow within the range of 0.36 to 0.62 ml/min when pressure varied from 14 to 23 mm Hg. Hydrodynamic resistance demonstrated pressure-dependent variability from 20 to 78 mm Hg/(ml/min). The time drift of hydrodynamic parameters was significant (P < 0.001). The valve was insensitive to changes in temperature, external pressure, rapid fluctuations of differential pressure, small particles in fluid, and reflux. CONCLUSION: The Diamond valve demonstrated the intended variable resistance, which increased as the pressure increased. This property may help it limit overdrainage related to body posture as well as nocturnal vasogenic waves. Flow through the valve stabilizes within a wide range, which may contribute to the prevention of excessive pressure buildup after implantation. However, shunt placement should be avoided in patients who present with normal baseline intracranial pressure but an increased incidence of high vasogenic intracranial pressure waves. PMID- 11270564 TI - CUSA EXcel ultrasonic aspiration system. PMID- 11270565 TI - The "unreachable" peaks of the neurointerventional mountaineers. PMID- 11270566 TI - Effects of radiation on cerebral vasculature: a review. PMID- 11270567 TI - Familial colloid cysts of the third ventricle. PMID- 11270568 TI - Microinvasive transaxillary thoracoscopic sympathectomy: technical note. PMID- 11270569 TI - Resolution of tonsillar herniation after supratentorial tumor resection: case report and review of the literature. PMID- 11270570 TI - Management of severe traumatic brain injury by decompressive craniectomy. PMID- 11270571 TI - A novel genetic screen identifies checkpoint-defective alleles of Schizosaccharomyces pombe chk1. AB - The protein kinase Chk1 is required in the fission yeast Schizosaccharomyces pombe for delaying cell cycle progression in response to DNA damage. Chk1 becomes phosphorylated when DNA is damaged by a variety of agents, including the anti tumor drug camptothecin. To further characterize the behavior of Chk1 in response to DNA damage, we used PCR-based mutagenesis of the chk1 gene coupled with in vivo gap repair to generate mutant alleles. Of 44 chk1 mutants recovered, six encode full-length proteins that confer a DNA damage-sensitive phenotype. All of the alleles render cells checkpoint-defective, but confer subtle differences in sensitivity to camptothecin or UV light. Mutant alleles were sequenced and served to identify regions of the protein that are critical for checkpoint function. PMID- 11270572 TI - Schizosaccharomyces pombe taf1+ is required for nitrogen starvation-induced sexual development and for entering the dormant GO state. AB - Environmental change, such as nutritional starvation, induces physiological and morphological alterations that enable fission yeast cells to survive. We isolated a novel gene, taf1+, required for the response to nitrogen starvation in the fission yeast Schizosaccharomyces pombe. taf1 disruptants could not mate upon nitrogen starvation, but could upon carbon starvation. taf1 disruptants had a defect in inducing stell+ expression under nitrogen starvation conditions. Furthermore, they lost viability quickly in nitrogen-depleted medium. Unlike wild type cells, starved taf1-cells had nuclear chromatin that were flat and adhered to the cell periphery. These results indicate that tqf1+ is required for nitrogen starvation-induced sexual development and entering the dormant G0 state. PMID- 11270573 TI - Regulation of the Aspergillus nidulans hisB gene by histidine starvation. AB - The hisB gene of the filamentous fungus Aspergillus nidulans encodes imidazole glycerol-phosphate dehydratase (E.C. 4.2.1.19), which catalyses the seventh enzymatic step in histidine biosynthesis. The gene was isolated and its deduced peptide sequence of 247 amino acids showed up to 54% identity with the IGPD enzymes of organisms comprising all three kingdoms. Expression of hisB cDNA in a Saccharomyces cerevisiae his3delta mutant strain functionally complemented the growth phenotype under histidine limitation. Addition of histidine did not affect hisB mRNA levels in A. nidulans wild-type cells. Histidine starvation conditions increased the hisB transcript level four-fold, suggesting regulation by a cross pathway regulatory network. Deletion of the complete hisB open reading frame in A. nidulans strain A234 resulted in histidine auxotrophy. Additionally, hisB deletion strains were blocked from sexual fruiting body formation on medium containing low concentrations of histidine. This developmental phenotype of the hisB deletion mutant strain correlated with the induction of the cross-pathway control system. PMID- 11270574 TI - Ambient alkaline pH prevents maturation but not synthesis of ASPA, the aspartyl protease from Penicillium roqueforti. AB - Penicillium roqueforti secretes an aspartyl protease, ASPA, which represents the main extracellular proteolytic activity. Alkaline pH of the medium plays a major role by inhibiting the enzymatic activity and stopping aspA expression in the presence of casein, an inducing protein. However, casein degradation by the mature enzyme produces peptides which can induce aspA expression at acidic and alkaline pH. ASPA synthesized as a proenzyme is processed at an acidic pH but not at an alkaline pH. The data indicate that, in P. roqueforti, alkaline pH has an indirect repressive effect by inhibiting ASPA maturation and the release of inducers. At an acidic pH, the mature enzyme degrades extracellular proteins and peptides are released to induce aspA. In contrast, at an alkaline pH, the proenzyme remains inactive, the inducing substances are consequently not produced and aspA is no longer expressed. PMID- 11270575 TI - L-Sorbose induces cellulase gene transcription in the cellulolytic fungus Trichoderma reesei. AB - L-Sorbose has previously been assumed to stimulate cellulase formation in an indirect manner, different from that of sophorose in Trichoderma reesei. Through Northern blot analysis however, L-sorbose was found to regulate coordinately six cellulase genes (including eg13, whose behavior has not been studied so far) at transcriptional level, as is the case with sophorose in T. reesei strains PC-3-7 and QM9414. Dot blot analysis showed that the proportions of each cellulase mRNA to cbh1 mRNA, the largest amount of mRNA transcribed in T. reesei, did not change when L-sorbose or sophorose was used as an inducer in the PC-3-7 and QM9414 strains. cbh2 and egl1 mRNAs were about 45-60% and 20-30% of the cbh1 transcript, whereas small amounts of mRNA, 1-2% of cbh1, were observed on other endoglucanase genes. Furthermore, the PC-3-7 strain showed an enhanced level of cellulase gene transcription, about two- and four- to six-fold higher than that of the QM9414 strain with sophorose and L-sorbose, respectively. PMID- 11270576 TI - Regulation of chitinase 33 (chit33) gene expression in Trichoderma harzianum. AB - We investigated the regulation of chit33 expression in Trichoderma harzianum CECT 2413. This gene encodes the Chit33 endochitinase, which is a major component of the fungus' chitinolytic enzyme system and is important for biocontrol. To this end, both Northern analysis and reporter gene fusions of a 1.4-kb fragment of the 5'-upstream sequences of chit33 to the Aspergillus niger goxA gene (encoding glucose oxidase) and the Aquorea victoria green fluorescent protein were used. Northern analysis and data obtained with the reporter systems were compatible, thus showing that the 1.4-kb fragment bears all necessary information for the regulation of chit33 gene expression. chit33 is weakly expressed during growth on chitin and Rhizoctonia solani cell walls. The addition of N-acetylglucosamine transiently induced chit33 expression in resting cells of the fungus. The addition of either glucose or glycerol prevented induction of chit33 gene expression by chitin or cell walls. Incubation of T. harzianum in the presence of low concentrations (0.1%, w/v) of glucose and high concentrations (38 mM) of ammonium sulfate, or in the presence of high concentrations (1%, w/v) of glucose and low concentrations (0.38 mM) of ammonium sulfate also stimulated chit33-mRNA accumulation, although to a lower degree than induction by N-acetylglucosamine. Transfer of T. harzianum cultures to either 40 degrees C or 4 degrees C initiated a very rapid expression of chit33 in the absence of an inducer, yet only at very low levels (5%) of the induced control. Confrontation experiments, using the gfp gene as a reporter and R. solani as a host, showed that chit33 is expressed only during but not before the stage of overgrowth on R. solani. These data show that Chit33 is an enzyme involved in mycoparasitism; and its formation is controlled by induction, by either carbon or nitrogen starvation and, to a low degree, also under conditions of temperature stress. PMID- 11270577 TI - Development of a strain-specific SCAR marker for the detection of Trichoderma atroviride 11, a biological control agent against soilborne fungal plant pathogens. AB - The genus Trichoderma includes biocontrol agents (BCAs) effective against soilborne plant pathogenic fungi. Several potentially useful strains for biological control are difficult to distinguish from other strains of Trichoderma found in the field. So, there is a need to find ways to monitor these strains when applied to natural pathosystems. We have used random amplified polymorphic DNA (RAPD) markers to estimate genetic variation among sixteen strains of the species T. asperellum, T. atroviride, T. harzianum, T. inhamatum and T. longibrachiatum previously selected as BCAs, and to obtain fingerprinting patterns. Analysis of these polymorphisms revealed four distinct groups, in agreement with previous studies. Some of the RAPD products generated were used to design specific primers. Diagnostic PCR performed using these primers specifically identify the strain T. atroviride 11, showing that DNA markers may be successfully used for identification purposes. This SCAR (sequence characterised amplified region) marker can clearly distinguish strain 11 from other closely related Trichoderma strains. PMID- 11270579 TI - Occupancy by oral administration of nicardipine of L-type calcium channels in rat brain. AB - The occupancy of L-type Ca2+ channels by treatment with an oral dose of the dihydropyridine-type Ca2+ antagonist nicardipine (sustained-release formulation) was evaluated in membrane preparations of rat frontal cortex and hippocampus using a radioligand binding assay technique, with [3H]-nicardipine as a ligand. Three hours after nicardipine administration, specific binding was decreased by about 15-20%, both in the frontal cortex and hippocampus. This indicates that oral nicardipine occupied approximately 15-20% of L-type Ca2+ channels. A progressive occupancy of Ca2+ channels was observed between six and 12 h after nicardipine administration. Twelve hours after drug administration, approximately 65-70% of Ca2+ channels were occupied. These findings indicate that oral treatment with 3 mg/kg of nicardipine (sustained-release formulation) occupies L type Ca2+ channels in rat brain by more than 40% from the 6th to the 24th h after drug administration. This suggests that an oral dose of nicardipine (sustained release formulation) in duces a significant occupancy of L-type Ca2+ channels in rat frontal cortex and hippocampus for about one day. The possible clinico therapeutic relevance of this observation is discussed. PMID- 11270578 TI - Antihypertensive drugs and sympathetic nervous system. AB - Several studies have demonstrated that essential hypertension is accompanied by sympathetic activation, which contributes to blood pressure elevation. Sympathetic activation also has adverse consequences in hypertensive patients beyond initiating blood pressure elevation. There is evidence that neural vasoconstriction has metabolic effects in skeletal muscle, impairing glucose delivery to muscles. In the liver, retarding of post prandial clearance of lipids contributes to hyperlipidemia. Cardiac sympathetic activation is a probable cause of sudden death in hearth failure. A trophic effect of sympathetic activation on cardiovascular growth is also likely, contributing to the development of left ventricular hypertrophy. Consequently, one of the major aims of antihypertensive therapy should be to attenuate sympathetic tone. It is possible that, among the antihypertensive drugs available, those inhibiting the sympathetic nervous system might best reduce cardiovascular risk. PMID- 11270580 TI - Effect of nicardipine treatment on the expression of neurofilament 200 KDa immunoreactivity in the brain of spontaneously hypertensive rats. AB - Neurofilaments (NFP) are components of neuronal cytoskeleton involved primarily in axonal transport and in the regulation of dynamic activities of nerve cells. NFP consist of three subunits denominated high- (200 kDa, NFP-H), intermediate- (160 kDa, NFP-I), and low-molecular weight (68 kDa, NFP-L) neurofilament proteins. Their function and polymerization depends on phosphorylation status, and is regulated by Ca2+ influx. Ca2+ overload enhances degradation of NFP and may compromise axonal transport. An increased susceptibility to ischemia occurs in hypertension, which is also a cause of brain damage. In this study, the expression of phosphorylated NFP (P-NFP) was investigated in the brain of spontaneously hypertensive rats (SHR) using immunohistochemical techniques with antibodies against the phosphorylated epitope of NFP RT-97. Microanatomical analysis included frontal cortex, occipital cortex, hippocampus and cerebellar cortex. The effect of long-term treatment with the dihydropyridine-type Ca2+ antagonist nicardipine on the expression of P-NFP was investigated as well. In hypertension a decreased P-NFP immunoreactivity was observed in frontal and occipital cortex, in the CA1 subfield of hippocampus and in the dentate gyrus, but not in the CA3 subfield of hippocampus or in the cerebellar cortex. Treatment with a daily dose of 3 mg/kg of nicardipine and 10 mg/kg of hydralazine significantly reduced systolic pressure in SHR. The above dose of nicardipine and to a lesser extent a non-hypotensive dose of the compound (0.1 mg/kg/day), but not hydralazine, increased P-NFP immunoreactivity in the cerebral cortex and hippocampus, except the CA3 subfield. The possibility that rescued P-NFP immunoreactivity by treatment with nicardipine depends on improved brain perfusion caused by the compound and/or by countering neuronal Ca2+ overload is discussed. PMID- 11270581 TI - Protective effect of treatment with nicardipine on cerebrovascular tree of spontaneously hypertensive rats. AB - The effect of an eight-week treatment with the Ca2+ channel blocker nicardipine on different-sized pial arteries and intracerebral arteries was assessed in spontaneously hypertensive rats (SHR) by microanatomical techniques. Normotensive Wistar-Kyoto (WKY) rats were used as normotensive reference animals. In SHR a significant increase of systolic blood pressure (SBP) in comparison with WKY rats was noticeable. An increased thickness of tunica media and luminal narrowing were also seen in medium- and small-sized pial arteries, and in intracerebral arteries of SHR in comparison with WKY rats. The media-to-lumen ratio was also increased in medium (diameter between 150 and 50 microm) and small-sized (diameter < than 50 microm) pial and intracerebral arteries. Treatment with nicardipine significantly reduced SBP, the thickness of tunica media, media-to-lumen ratio and increased luminal area of medium- and small-sized pial arteries and of intracerebral arteries. These findings demonstrate that treatment of SHR with nicardipine induces a moderate vasodilatation of both pial and intracerebral arteries regulating cerebrovascular resistance. This property may be useful in avoiding generalized or exaggerated cerebrovascular dilatation in hypertension which could be accompanied by impaired brain perfusion. PMID- 11270582 TI - The dopaminergic system in hypertension. AB - Dopamine exerts cardiovascular and renal actions mediated through interaction with specific dopamine receptors. Dopamine receptors are cell surface receptors coupled to G-proteins and classified into two main super families based on biochemical, pharmacological and molecular characteristics. The dopamine D1-like receptor super family includes D1 and D5 receptors, known also in rodents as D1A and D1B sites. These receptors are linked to stimulation of adenylate cyclase. The dopamine D2-like receptor super family includes D2, D3 and D4 receptors. These receptors are linked to inhibition of adenylate cylase or not related with this enzyme activity. They also interfere with opening of Ca+2 channels and are linked to stimulation of K+ receptors. Dopamine receptor subtypes are expressed in brain as well as in extracerebral structures such as the heart, blood vessels, carotid body, kidney, adrenal gland, parathyroid gland and gastrointestinal tract. In the kidney, which represents the peripheral organ where dopamine receptors were more extensively investigated, dopamine receptors are involved in regulation of hemodynamic, electrolyte and water transport, as well as renin secretion. Hypertension-related dopamine receptor changes were also investigated primarily in the kidney. Defective renal dopamine production and/or dopamine receptor function have been reported in human primary hypertension as well as in genetic models of animal hypertension. There may be a primary defect in D1-like receptors and an altered signalling system in the proximal tubules that lead to reduced dopamine-mediated effects on renal sodium excretion in hypertension. Studies on the influence of hypertension on dopamine D2-like receptors are sparse Disruption of either D1A or D3 receptors at the gene level causes hypertension in mice. Using peripheral blood lymphocytes as possible markers of the status of dopamine receptors in essential hypertension, no changes of dopamine D1-like receptors were noticeable, whereas an increase of dopamine D2-like receptors likely representing an up-regulation mechanism was reported. Available information collectively indicates an involvement of peripheral dopaminergic system in hypertension consisting either in impaired receptor transduction mechanisms and/or in receptor loss. A better knowledge of molecular bases of these changes may contribute to the development of specific therapeutic approaches in the future. PMID- 11270583 TI - Effect of antihypertensive treatment on peripheral nerve vasculature in spontaneously hypertensive rats. AB - The influence of hypertension and of treatment with the dihydropyridine-type Ca+2 antagonist nicardipine on peripheral nerve vasculature were investigated in spontaneously hypertensive rats (SHR). Male SHR were treated from the 16th to the 26th week of age with vehicle (control group), with nicardipine, at the hypotensive dose of 3 mg/kg/day, or at the nonhypotensive dose of 0.1 mg/kg/day or with an equihypotensive dose (10 mg/kg/day) of the nondihydropyridine-type vasodilator hydralazine. Age-matched normotensive Wistar-Kyoto (WKY) rats were left untreated and used as normotensive reference animals. In SHR a significant increase of systolic pressure values accompanied by sciatic nerve microvascular changes, involving primarily interfascicular arteries and to a lesser extent intrafascicular arteries, was observed. Treatment with the hypotensive dose of nicardipine countered hypertension-dependent microvascular changes occurring in both interfascicular and intrafascicular arteries. The nonhypotensive dose of nicardipine and hydralazine displayed a modest activity on interfascicular arteries, but significantly countered hypertension-related changes involving intrafascicular arteries. The above findings indicate the occurrence of hypertension-related changes of peripheral nerve microvasculature and of positive effects induced by appropriate pharmacological treatment. Further work is in progress to identify the functional relevance of microanatomical observations of the present study. PMID- 11270584 TI - Stroke unit: a cardio-cerebral approach. AB - Stroke units are special units where stroke patients receive, simultaneously, medical and physical treatment. Compared to general neurological and medical wards, stroke units show a significant reduction of short- and long-time mortality, and an improvement of long-term quality of life. Nevertheless, mortality in these units is still high (1-year mortality approximately 32%; 5 year mortality degrees 60%), and consequently, new approaches are needed to control stroke parameters during the acute phase, with the goal to reduce mortality rates. The philosophy of our stroke unit in Fermo (Italy), is to establish a strong association between heart and brain care by approaching each stroke patient as a cardiocerebral patient. In particular, we perform 12-lead Holter ECG monitoring, to prevent the vicious cycle affecting correct cerebral and cardiac functions, and to react to cardiac complications, mostly arrhythmias, that can worsen cerebral damage. Holter ECG monitoring allows a fast physiotherapeutic approach, a better evaluation of metabolic parameters, and collectively, a better global evaluation of the patient during the acute phase of disease. In two years of activity, 80 patients that were admitted to our stroke unit during 1998, and treated as cardio-cerebral patients, were followed-up. This combined treatment decreased the 1-year mortality rate by about 30%, in comparison with the 22% mortality rate reported in the literature. These results confirm the validity of stroke units, as well as of our approach based on cardio cerebral control of each stroke patient. PMID- 11270585 TI - Relationships between salt sensitivity of blood pressure and sympathetic nervous system activity: a short review of evidence. AB - Experimental and clinical studies provided evidence in favor of complex relationships between sympathetic nervous system activity and salt-sensitivity of blood pressure. Genetic and acquired metabolic alterations associated with a tendency to retain salt and water may generate salt-sensitivity of blood pressure and shift the pressure-natriuresis curve to the right, promoting an increase in blood pressure. Sympathetic activation is a factor contributing to this result. Chronic high dietary salt intake is followed by a derangement in mechanisms of central sympathetic inhibition and then by an enhanced peripheral sympathetic tone. This, in turn, may generate salt-sensitivity of blood pressure by affecting renal hemodynamics, tubular sodium and water handling. Insulin resistance and sodium and water retention are prompted by high-fat (as well as high carbohydrate) diets, and by an increase in body fat mass. Also, aging is a condition of impaired interactions of the above factors. A gain in weight due to reduced physical activity, not followed by a parallel decrease in calorie intake, brings to a fall in insulin sensitivity. In many cases, the natural age-related decline of renal function is associated with a reduced physical exercise, hyperinsulinemia and sodium retention; sympathetic nervous system activity is enhanced and causes an increase in blood pressure. PMID- 11270586 TI - Peripheral adrenergic system and hypertension. AB - Hypertension is a condition where adrenergic responsiveness, sympathetic activity and adrenoceptors are somewhat altered. Many techniques are available to assess human sympathetic nervous system activity. They each present limitations and disadvantages. Characterization and subdivision of the alpha and beta adrenoceptors, according to their localization and answer to different agonists, was facilitated in recent years by the extensive use of pharmacological and molecular biology techniques. Some adrenoceptor studies were conducted on animal models, human tissues and peripheral blood cells to assess their changes in various forms and stages of hypertension. Our group has pointed out that alpha1 adrenergic receptors expressed by human peripheral blood lymphocytes underwent changes of density in essential hypertensives, compared to normotensive control subjects. The importance of these findings could provide an assessment of alpha1 peripheral receptors with possible future clinical implications in the pathophysiology and treatment of hypertension. PMID- 11270587 TI - Role of the sympathetic nervous system in cardiac remodeling in hypertension. AB - The role of the sympathetic nervous system in the settings of cardiac hypertrophy is postulated on the basis of experimental and clinical evidence. Only recently, the intimate mechanisms underlying hypertrophic responses of cardiac cell have been explored. Recent evidence spots the role of adrenergic receptors in the activation of several intracellular pathways of signaling that lead to nuclear responses of myocardiocyte. The molecular structures involved in these pathways may represent novel targets of future therapeutic interventions for cardiac hypertrophy and vascular remodeling in hypertension. PMID- 11270588 TI - Sympathetic nervous system, diabetes, and hypertension. AB - Hypertension is twice as frequent in diabetic patients than in the general population. Its prevalence is higher in Type 2 than in Type 1 diabetes: in the former, the onset of hypertension often precedes the diagnosis of diabetes, whereas, in the latter it is strictly related to the presence of nephropathy. Sympathetic nerve overactivity is crucial in the pathogenesis of hypertension in diabetes. It can be related to the activation of the renin-angiotensin aldosterone (RAA) system in Type 1 diabetic patients with chronic renal failure, or to a condition of insulin resistance/hyperinsulinemia in Type 2 patients with the metabolic syndrome. In patients with early autonomic neuropathy, vagal impairment can lead to a relative predominance of sympathetic activity in the sympatho-vagal balance. In these patients, the onset of hypertension is frequently preceded by reduced nocturnal dipping. Sympathetic overactivity stimulates RAA activity, promotes sodium reabsorption, and increases heart rate, stroke volume and peripheral vascular resistance, thus inducing hypertension and increasing cardiovascular risk. A number of drugs acting either directly or indirectly on sympathetic activity are available for the treatment of hypertension in diabetic subjects. Opinions on the potential advantages of the metabolic profile of some of these drugs are as yet conflicting. PMID- 11270589 TI - Obesity and autonomic function in adolescence. AB - Hypertension and obesity are risk factors for coronary heart diseases in adults. In turn, childhood overweight and high blood pressure increase the risk of subsequent obesity and hypertension in adulthood. Human obesity is characterized by profound alterations of hemodynamic and metabolic states. Whether these alterations involve sympathetic nervous system control on cardiac function is controversial. We report the results of our study, conducted in a sample of obese adolescents by using power spectral analysis of heart rate variability. An increase in sympathetic tone coupled with a reduction in vagal tone was found. This allowed us to hypothesize that autonomic nervous system changes depend on the time course of obesity development. It is still unclear if treatment of obesity in adolescence prevents subsequent autonomic imbalance and hypertension. PMID- 11270590 TI - Sympathetic nervous system and chronic renal failure. AB - The aim of this work was to review evidence on the role of the sympathetic nervous system (SNS) in chronic renal failure (CRF). Three main points are discussed: 1) SNS and pathogenesis of arterial hypertension; 2) SNS and cardiovascular risk; 3) implication of SNS in arterial hypotension during hemodialysis. Several lines of evidence indicate the presence of a sympathetic hyperactivity in CRF, and its relationship with arterial hypertension. It is suggested that diseased kidneys send afferent nervous signals to central integrative sympathetic nuclei, thus contributing to the development and maintenance of arterial hypertension. The elimination of these impulses with nephrectomy could explain the concomitant reduction of blood pressure. Several experiments confirmed this hypothesis. Regarding SNS and cardiovascular risk, some data suggest that reduced heart rate variability identifies an increased risk for both all causes and sudden death, independently from other recognized risk factors. Symptomatic hypotension is a common problem during hemodialysis treatment, occurring in approximately 20-30% of all hemodialysis sessions and is accompanied by acute withdrawal of sympathetic activity, vasodilation and relative bradicardia. This reflex is thought to be evoked by vigorous contraction of a progressively empty left ventricle, activating cardiac mechanoceptors. This inhibits cardiovascular centers through vagal afferents, and overrides the stimulation by baroreceptor deactivation. Alternative explanations include cerebral ischemia and increased production of nitric oxide, which inhibit central sympathetic activity. It is hoped that therapies aimed at modulating sympathetic nerve activity in patients with CRF will ameliorate their prognosis and quality of life. PMID- 11270591 TI - Heart rate variability and left ventricular diastolic function in patients with borderline hypertension with and without left ventricular hypertrophy. AB - The relationships between heart rate variability (HRV), left ventricular mass and diastolic function in borderline hypertensive patients (BHT) were evaluated. 24 h Holter electrocardiogram (ECG) and blood pressure (BP) monitoring, M and 2 D echocardiogram and Doppler analysis in 42 BHT with and without left ventricular hypertrophy (LVH) and in 20 normotensive controls were assessed. From 24-h ECG, time domain indexes of HRV were calculated. Standard Deviation of all Cycles (SDNN) and Standard Deviation of the means of heart periods over five-minute intervals (SDANN) were significantly reduced in BHT with LVH but not in BHT without LVH. No significant differences of short-term variability measures were detectable, although a progressive decrease among control subjects and BHT with and without LVH was observed. Diastolic left ventricular compliance evaluated by early to late transmitral flow velocity ratio (E/A ratio) significantly declined from normotensive subjects to BHT with LVH. There was a significant positive correlation between E/A and SDNN and SDANN throughout all studied groups. This indicates that BHT with LVH has a reduced HRV compared to other groups. This impairment is probably related to left ventricular mass and left ventricular filling abnormalities. PMID- 11270592 TI - Baroreflex sensitivity in secondary hypertension. AB - A deranged baroreceptor control of the cardiovascular functions has been reported in essential hypertension. Studies performed in experimental animals and in humans using different approaches have documented an impairment of both baroreflex heart rate modulation (resetting and loss of sensitivity) and baroreceptor control of peripheral vasomotor tone (only resetting). Baroreflex alterations have been reported also in secondary forms of hypertension, but data are controversial. This paper reviews recent works concerning baroreflex function in secondary hypertension. Either structural changes of arterial wall (decrease of vascular distensibility) or functional processes (involving angiotensin II, aldosterone, catecholamines, nitric oxide) have been proposed as potential mechanisms responsible for baroreflex readjustments in secondary hypertension. It remains unclear, and it is difficult to define exactly, if baroreflex changes associated with secondary form of hypertension are primarily due to factors specific for different hypertensive conditions, or merely follow blood pressure elevation. PMID- 11270593 TI - MHV infection of the CNS: mechanisms of immune-mediated control. AB - Mice infected with neurotropic strains of mouse hepatitis virus (MHV) clear infectious virus; nevertheless, viral persistence in the central nervous system (CNS) is associated with ongoing primary demyelination. Acute infection induces a potent regional CD8+ T-cell response. The high prevalence of virus specific T cells correlates with ex vivo cytolytic activity, interferon-gamma (IFN-gamma) secretion and efficient reduction in virus. Viral clearance from most cell types is controlled by a perforin dependent mechanism. However, IFN-gamma is essential for controlling virus replication in oligodendrocytes. Furthermore, CD4+ T cells enhance CD8+ T-cell survival and effectiveness. Clearance of infectious virus is associated with a gradual decline of CNS T cells; nevertheless, activated T cells are retained within the CNS. The loss of cytolytic activity, but retention of IFN gamma secretion during viral clearance suggests stringent regulation of CD8+ T cell effector function, possibly as a means to minimize CNS damage. However, similar CD8+ T-cell responses to demyelinating and non demyelinating JHMV variants support the notion that CD8+ T cells do not contribute to the demyelinating process. Although T-cell retention is tightly linked to the presence of persisting virus, contributions to regulating the latent state are unknown. Studies in B-cell-deficient mice suggest that antibodies are required to prevent virus recrudescence. Although acute JHMV infection is thus primarily controlled by CD8+ T cells, both CD4+ T cells and B cells make significant contributions in maintaining the balance between viral replication and immune control, thus allowing host and pathogen survival. PMID- 11270594 TI - Persistence of respiratory syncytial virus in macrophages alters phagocytosis and pro-inflammatory cytokine production. AB - Functions of macrophage are known to be altered by acute infection with respiratory syncytial virus (RSV). However, it is unknown whether the persistent presence and expression of the RSV genome have any effect on the functions of these cells. We used a murine macrophage-like cell line (P388D1) persistently infected with RSV to determine: (i) phagocytic activity mediated by Fcgamma receptors, (ii) expression of Fcgamma receptors, and (iii) production of IL 1beta, IL-6 and TNF-alpha. Viral persistence was found to increase phagocytosis, expression of Fcgamma receptors and the production of IL-1beta and IL-6. In contrast the biological activity of secreted TNF-alpha decreased. In this study we give novel evidence that RSV persistence alters the biological activities of macrophages. PMID- 11270595 TI - Protective efficacy of rotavirus 2/6-virus-like particles combined with CT-E29H, a detoxified cholera toxin adjuvant. AB - Identifying a safe and efficacious mucosal adjuvant is crucial for the development of subunit vaccines against rotavirus and other mucosal pathogens. Moreover, recognition of determinants of protective immunity to rotavirus infection is essential to the design of the means to prevent or control this viral gastrointestinal disease. We have studied the kinetics of systemic and mucosal antibody responses elicited upon mucosal immunization of mice with rotavirus recombinant virus-like particles (rVLPs) alone or combined with a detoxified version of cholera toxin, CT-E29H. CT-E29H has been shown to maintain the adjuvant effect of parental cholera holotoxin. Both inbred BALB/c and outbred CD-1 mice were immunized with rotavirus VP2/6-rVLPs (2/6-VLPs) combined with CT E29H, orally or intranasally (i.n.), and the comparative efficacy of different formulations was then determined. Rotavirus-specific serum and fecal IgA, IgM, and IgG antibodies were determined by enzyme-linked immunoadsorbent assay (ELISA) weekly (or every other week) following vaccination. Animals then were challenged with a murine rotavirus strain, EDIM. The degree to which vaccinated animals were protected from the wild-type rotavirus challenge was reflected in the levels of viral antigen shed in stools (percent reduction in antigen shedding, PRAS). BALB/c mice immunized by either route produced rotavirus-specific serum IgA, IgM and IgG, as well as fecal IgA and IgG, but not IgM; however, the intranasal immunization induced stronger systemic IgG and IgM responses than did oral immunization. Similar levels of prechallenge rotavirus-specific fecal and serum IgA were detected in both the orally and the i.n. immunized groups. Two immunizations with 2-6VLPs and CT-E29H were sufficient to protect BALB/c mice, regardless of the route of administration. PRAS was 99.6, 98.8, and 98.8% for oral, i.n. and the oral + i.n. groups, respectively; in contrast vaccination with 2/6-VLPs alone was not protective (PRAS = 39%), indicating the critical role of CT-E29H in inducing protective levels of immune responses. Two of four outbred CD 1 mice that were immunized orally with 2/6-VLPs-CT-E29H showed no humoral responses (PRAS, 65%), but four of four i.n. immunized CD-1 mice displayed humoral responses (PRAS, 97.9%). Serum anti-VP6 and VP2 antibodies were detected in all immunoresponsive mice. The combined results in two strains of mice indicate that CTE29H is an effective mucosal adjuvant capable of inducing protective immune responses and suggest that intranasal administration is the preferred route of immunization. PMID- 11270596 TI - Induction of a virus-specific antibody response to foot and mouth disease virus using the structural protein VP1 expressed in transgenic potato plants. AB - We have recently communicated the oral and parental immunogenicity of the structural protein VP1 of foot and mouth disease virus (FMDV) expressed in different transgenic plants. Those results clearly indicated the necessity of increasing the expression of the foreign genes in the transgenic plant to avoid additional steps toward the purification and/or concentration of the antigen of interest. Here, we report the production of transgenic potatoes plants containing the VP1 gene cloned under the regulatory activity of either a single (pRok2) or a double (pRok3) copy of the S35 cauliflower mosaic virus (CaMV 35S) promoter, as a strategy for increasing the level of VP1 gene expression. The presence of the VP1 gene in the plants was confirmed by polymerase chain reaction (PCR) and its specific transcription activity was demonstrated by reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that, although the immunized animals presented a FMDV VP1 specific antibody response and protection against the experimental challenge, no significant differences were demonstrated in the immunizing activity of plant extracts obtained from the pRok2 or pRok3 transformed plants. These results confirm those previously obtained using other plant species allowing the possibility of using plants as antigen expression vectors, and demonstrated that at least in the potato system, the use of double CaMV 35S promoter does not cause a significant increase in the level of the VP1 expressed. PMID- 11270597 TI - Direct functional analysis of epitope-specific CD8+ T cells in peripheral blood. AB - The functional status of virus-specific CD8+ T cells is important for the outcome and the immunopathogenesis of viral infections. We have developed an assay for the direct functional analysis of antigen-specific CD8+ T cells, which does not require prolonged in vitro cultivation and amplification of T cells. Whole blood samples were incubated with peptide antigens for <5 h, followed by staining with peptide-MHC tetramers to identify epitope-specific T cells. The cells were also stained for the activation marker CD69 or for the production of cytokines such as interferon-gamma (IFNgamma) or tumor necrosis factor-alpha (TNFalpha). With the combined staining with tetramer and antibodies to CD69 or cytokines the number of antigen-specific CD8+ T cells as well as the functional response of each individual cell to the cognate antigen can be determined in a single experiment. Virus-specific CD8+ T cells that are nonfunctional, as well as those that are functional under the same stimulating conditions can be simultaneously detected with this assay, which is not possible by using other T-cell functional assays including cytotoxicity assay, intracellular cytokine staining, and enzyme-linked immunospot (ELISPOT) assay. PMID- 11270598 TI - Virulence of mousepox virus is independent of serpin-mediated control of cellular cytotoxicity. AB - We have investigated whether the differential virulence seen of two Ectromelia (Ect) strains, EctMoscow and ECtHampstead egg, is due to mutation or differential regulation of their serpins (SPI). Poxvirus encoded serine proteinase inhibitors (serpins) have been shown to interfere with cytolytic activity of leukocytes and can also determine virulence. We show that the deduced amino acid sequences of SPI-1, 2, and 3 are identical for the highly virulent EctMoscow and the low virulent EctHampstead strains and that the two viruses express similar potential to inhibit T-cell cytotoxicity, in particular, Fas-mediated target cell lysis, by allorective effectors. Virus titres in wild type B6 mice were effectively controlled very early after inoculation with EctHampstead as compared with EctMoscow, but lack of perforin renders B6 mice similarly susceptible to both virus strains. The data demonstrate that in Ect infection the perforin-mediated cytolytic pathway is not the primary target of serpins and suggest that the apparent attenuation of EctHampstead seen in B6 mice is due to control elements distinct from SPI-1, 2, and 3. PMID- 11270599 TI - Mapping of T-helper epitopes of Rinderpest virus hemagglutinin protein. AB - Rinderpest virus (RPV) is a highly contagious and often fatal disease of domestic and wild ruminants, caused by rinderpest virus of the genus Morbillivirus under the family Paramyxoviridae. Hemagglutinin (H) and fusion (F) proteins of this enveloped virus confer protective immunity against experimental challenge with virulent rinderpest virus. We have earlier demonstrated that immunization with a single dose of recombinant extracellular baculovirus expressing H protein elicits H-specific humoral and lymphoproliferative responses in cattle. The lymphoproliferative responses are predominantly BoLA class II restricted. In this work, we have analyzed lymphoproliferative responses of peripheral lymphocytes from immunized cattle to truncated H protein fragments expressed in E. coli for locating domains harboring Th epitopes. One region (aa 113-182) recognized by immune T cells is conserved in the H protein of measles virus, which was earlier shown to contain a dominant Th epitope in mouse. Synthetic peptides within this region of measles virus H protein were used to identify a Th epitope conserved in the H protein of RPV virus (aa 123-137) in cattle. A second Th epitope located at the C-terminus of RPV-H was mapped to the region corresponding to aa 512-609 using truncated protein fragments expressed in E. coli. The C-terminal epitope (aa 575-583) was mapped using synthetic peptides corresponding to measles virus H as well as RPV-H protein. PMID- 11270600 TI - A role for lipid rafts in immune cell signaling. AB - Cross-linking of surface receptors in hematopoietic cells results in the enrichment of these receptors in the rafts along with other downstream signaling molecules. A possible explanation how signal is transduced through the plasma membrane has arisen from the concept of raft. From the study of cellular responses in the plasma membrane which enrich members of the Src-family tyrosine kinase, rafts can function as centers of signal transduction by forming patches. Under physiological conditions, these elements synergize to transduce successfully a signal at the plasma membrane. Rafts are suggested to be important in controlling appropriate protein interactions in hematopoietic cells, and aggregation of rafts following receptor ligation may be a general mechanism for promoting immune cell signaling. PMID- 11270601 TI - Elimination of latently HIV-1-infected cells by lymphoblasts armed with bifunctional antibody. AB - The Fab' fragment of a monoclonal antibody (mAb) to CD3 and the F(ab')2 fragment of a mAb to human immunodeficiency virus 1 (HIV-1) gp41 were combined to generate a bifunctional antibody (BFA). The mAb to gp41 (IV1-4G6) has previously been shown to react with a number of HIV-1 strains and T-lymphoblastoid cells (TLBC) armed with the BFA (BFA-TLBC) effectively inhibited HIV-1 in primarily cultured lymphoblasts infected with the clinically isolated virus which was reactive to the mAb. Although BFA-TLBC could not cause cytolysis of 51Cr-labeled latently infected cells (OM-10.1) in 6 hr incubation, cocultivation of OM-10.1 cells with BFA-TLBC for 3 days or more eliminated the latently infected cells making the cells susceptible to BFA-TLBC. Therefore, BFA-TLBC may be beneficial for HIV infected patients in eradicating latently infected cells which can not be eliminated even with highly active antiretroviral therapy (HAART). PMID- 11270602 TI - PCR detection of Capnocytophaga species in dental plaque samples from children aged 2 to 12 years. AB - The purpose of this study was to detect the presence of Capnocytophaga sputigena, C. ochracea, and C. gingivalis in plaque samples from the toothbrushes of 122 children, using a polymerase chain reaction (PCR) method. The subjects were 25, 85, and 12 children with healthy gingiva, gingivitis, and periodontitis, respectively, ranging in age from 2-12 years old. Plaque samples were collected from all erupted tooth sites using a sterile toothbrush. The mean amount of DNA recovered from the samples was approximately 19.3 microg, which was deemed sufficient for performing a PCR-based survey. C. sputigena prevalence in healthy, gingivitis, and periodontitis subjects was 48.0%, 36.5% and 25.0%, respectively, that for C. ochracea was 100%, 89.4%, and 50.0%, respectively, and that for C. gingivalis was 96.0%, 84.7%, and 75.0%, respectively. The lowest age of positive subjects was approximately 2 years. Our results showed that C. sputigena was moderately prevalent, whereas C. ochracea and C. gingivalis were commonly detected in the oral cavities of the tested children, suggesting that all of these species become established in the early years. PMID- 11270603 TI - Relatedness between the coagulase gene 3'-end region and coagulase serotypes among Staphylococcus aureus strains. AB - The 3'-end region of the coagulase gene from 22 strains of Staphylococcus aureus including 10 standard serotype strains was sequenced, and five subgroups with 4-8 tandem repeating units were distinguished among the tested strains. Phylogenetic analysis of the 3'-end region of the coagulase gene indicated that strains belonging to the same serotype were clustered in the same branch. A phylogenetic tree of the deduced amino acid sequences revealed that the C-terminal region might not be responsible for the epitope of the coagulase protein. PMID- 11270604 TI - Intraperitoneal immune cell responses to Eubacterium saphenum in mice. AB - Oral asaccharolytic Eubacterium saphenum, which are newly isolated gram-positive rods and one of the predominant microorganisms in human periodontal pockets, were injected intraperitoneally in mice to elucidate their pathogenicity in periodontal diseases. Infiltrating immune cells in the peritoneal exudate were quantitated and intracellular T cell (CD4+/CD8+/gammadelta+) production of cytokines IL-4 and IFN-gamma which are related to cellular and humoral immunity, respectively, was determined. Neutrophils appeared first in peritoneal exudates, followed by macrophages and lymphocytes, after the injection of either E. saphenum or Porphyromonas gingivalis. Intracellular IL-4+ and IFN-gamma+ gammadelta T cells were detected in the exudates after the injection of E. saphenum (4.6 +/- 0.8% and 10.1 +/- 1.4%, respectively) and P. gingivalis (5.3 +/ 1.6% and 10.1 +/- 2.1%, respectively). The intracellular production of IL-4/IFN gamma in CD4+/CD8+ T cells was rather low indicating that the main response was from gammadelta T cells which initiated the immune reactions in mouse peritoneal cavities after injection of E. saphenum or P. gingivalis. Serum IgG and IgM levels were elevated in animals injected with E. saphenum and similarly with P. gingivalis. The present study showed that with slight differences, similar modes of cell response and cytokine and Ig production were observed after intraperitoneal injection of both E. saphenum and P. gingivalis, indicating that E. saphenum may play just as important a role in periodontal diseases as P. gingivalis. PMID- 11270605 TI - Rapid detection and quantification of five periodontopathic bacteria by real-time PCR. AB - The quantity of periodontopathic bacteria in plaque samples is an important determinant for understanding the etiologic role of bacteria. The real-time PCR method was used to detect and quantify periodontopathic bacteria, such as Actinobacillus actinomycetemcomitans, Bacteroides forsythus, Porphyromonas gingivalis, Treponema denticola, and Treponema socranskii, in saliva and subgingival plaque samples. There was good agreement between the results of conventional PCR and real-time PCR methods. Using the LightCycler system we were able to determine the amount of periodontopathic bacteria within an hour. The real-time PCR method was linear for samples containing from 10(3) to more than 10(8) cells (r2 = 0.999). The application of the real-time PCR method should be useful in the rapid detection and quantification of periodontopathic bacteria in clinical samples. PMID- 11270606 TI - Cross reaction of recombinant equine infectious anemia virus antigen to heterologous strains and application for serological survey among horses in the field. AB - Cross reactivity of equine infectious anemia virus (EIAV) antigen prepared using a recombinant baculovirus containing the p26 gene of strain P337-V70 was examined by the agar gel immunodiffusion (AGID) test and enzyme-linked immunosorbent assay (ELISA). Serum samples serially collected from 13 horses experimentally infected with six different EIAV strains (two or three horses per strain) were subjected to the test. Positive reactions were observed in the AGID test and ELISA before or soon after the first feverish period and continued persistently in most of the horses. The results with recombinant antigens were essentially the same as those with the virion antigen prepared from horse cell cultures both in the AGID test and ELISA. The reactivities of the antigens were further compared using serum samples collected from horses in 1999 in certain districts of Mongolia where equine infectious anemia has been prevalent, and from horses in Japan in 1973 when EIA had not been eliminated completely from Japanese horses. These results were completely concurrent. Generally, recombinant antigens have high specificity but low cross reactivity to heterologous strains. However, the present study showed that the recombinant EIAV p26 antigen has cross reactivity to the heterologous strain and is useful for diagnosis of EIA in the field. PMID- 11270607 TI - A comparison of complete genome sequences of the attenuated RC-HL strain of rabies virus used for production of animal vaccine in Japan, and the parental Nishigahara strain. AB - In order to establish the molecular basis of the pathogenicity of the attenuated RC-HL strain of rabies virus used for the production of animal vaccine in Japan, the complete genome sequence of this strain was determined and compared with that of the parental Nishigahara strain which is virulent for adult mice. The viral genome of both strains was composed of 11,926 nucleotides. The nucleotide sequences of the two genomes showed a high homology of 98.9%. The homology of the G gene was lower than those of N, P, M and L genes at both nucleotide and deduced amino acid levels, and the percentage of radical amino acid substitutions on the G protein was the highest among the five proteins. These findings raise the possibility that the structure of the G protein is the most variable among the five proteins of the two strains. Furthermore, we found two clusters of amino acid substitutions on the G and L proteins. The relevance of these clusters to the difference in the pathogenicity between the two strains is discussed. PMID- 11270608 TI - Hemadsorption expressed by cloned H genes from subacute sclerosing panencephalitis (SSPE) viruses and their possible progenitor measles viruses isolated in Osaka, Japan. AB - Most subacute sclerosing panencephalitis (SSPE) viruses, including our Osaka-1, 2, and -3 strains isolated in Osaka, have shown negative hemadsorption (HAD) by African green monkey red blood cells. This property has been thought to be characteristic of SSPE virus as compared to the positive reaction of the standard Edmonston strain of measles virus (MV). However, this assumption has become quite obscure because MV mutates frequently at the genetic level during its multiplication and also because recent field strains isolated by lymphoblastoid cell lines have shown negative HAD. To investigate the above issue, the nucleotide sequences of the hemagglutinin (H) genes from SSPE virus Osaka-1, -2, or -3 strains were compared to those of various MV field strains isolated in Osaka by Vero cells. The H gene sequences of three SSPE strains were relatively conserved without such biased hypermutation as had been observed in the matrix (M) gene of three SSPE strains. However, this analysis of the H gene sequence of the SSPE viruses enabled us to deduce possible progenitor MVs, which are in agreement with the deduction from the M gene analysis we reported previously. The HAD of Vero cells transfected with the cloned H cDNAs from the SSPE strains and their progenitors suggested that negative HAD of the SSPE viruses has been maintained as one of original properties of the progenitor MVs rather than having been acquired as an altered one during long-term persistent infection in the brains of patients with SSPE. PMID- 11270609 TI - Nitric oxide production and iNOS mRNA expression in mice induced by repeated stimulation with live Fusobacterium nucleatum. AB - There have been few studies on the detection of direct nitric oxide (NO) production and interferon-gamma (IFN-gamma) in vivo without using animal cell culture. We questioned whether NO and IFN-gamma could be produced at the site of infection. The peritoneal cavity of mice was used as the local infection model. NO and IFN-gamma in abdominal washings from these mice were measured directly at various times after injection of Fusobacterium nucleatum, a gram-negative rod periodontal pathogen. The mice were divided into three groups: those treated with live bacteria (LB), those treated with heat-killed bacteria (HKB) and those untreated: normal (N). These mice were compared on the basis of cell filtration, NO and IFN-gamma production by injection of live bacteria (LFn) or heat-killed bacteria (HKFn). In the LB group, the total cell number increased corresponding to an increase in neutrophils after injection of both LFn and HKFn. A low level of NO was constantly produced in abdominal washings, but a significant amount of NO was synthesized in the LB group only 12 hr to 24 hr after injection of LFn. At the same time iNOS enzyme activity and iNOS mRNA expression were detected. IFN gamma, which may contribute to enhance NO production, was also secreted at a high level from peritoneal exudate cells (PEC) at 12 hr and 24 hr in the LB group by stimulation of LFn. At 12 hr and 24 hr, iNOS positive cells in the LB group by infection of LFn were identified and shown to contain mostly macrophages. These findings indicate that live bacteria play important roles in NO production by macrophages. It is suggested that NO may contribute to the inflammatory response during F. nucleatum infection in periodontitis. PMID- 11270610 TI - Effect of constitutively expressed phoP gene on the localization of Salmonella typhimurium within Mac-1 positive phagocytes. AB - Intracellular localization of the wild-type (Spv+), the phoP-constitutively expressed strain (PhoPc), and the spv-deleted strain (Spv-) of Salmonella typhimurium was examined by the use of confocal laser scanning microscopy analysis of immunostained sections of mouse spleens after oral or subcutaneous inoculation. Only 40% of salmonellae of both the PhoPc and the Spv- strains were detected intracellularly within Mac-1 positive cells at day five after oral or day four after subcutaneous inoculation. In contrast, over 85% of salmonellae of the Spv+ strain were detected inside Mac-1 positive cells. In both inoculation trials, the splenic colony-forming unit values for the PhoPc and Spv- strains were significantly lower than the corresponding value for the Spv+ strain. These findings suggest that the constitutively expressed phoP gene of S. typhimurium attenuated virulence by limiting intracellular proliferation within mouse spleen phagocytes, and that the lack of spv genes had the same effect. PMID- 11270611 TI - Seroprevalence of antibodies against spotted fever group rickettsia among dogs and humans in Okinawa, Japan. AB - We evaluated serum antibodies against Rickettsia japonica in 517 dogs (430 stray dogs and 87 pet dogs) and 164 humans in Okinawa, Japan, by indirect immunofluorescence assay. The seropositive rate in stray dogs was significantly higher than that in pet dogs (30.7 versus 4.6%, P<0.01). This high prevalence rate is attributed to the understandably frequent environmental exposure of stray dogs to tick infestation. Human samples obtained from Okinawa and Sapporo also showed a significant difference in seropositive antibody percentages (45.1 and 12.0%, respectively, P<0.01). This result suggests that there has been pre exposure to spotted fever group rickettsia in humans in Okinawa. PMID- 11270612 TI - First detection of Ehrlichia platys in dogs and ticks in Okinawa, Japan. AB - We investigated Ehrlichia platys infection of dogs and ticks in Okinawa, Japan. Using E. platys specific primers, E. platys and HE3-R, PCR-positive results were obtained with 32.0% (64/200) of blood samples of dogs and 3.8% (3/77) of ticks. The nucleotide sequences of the amplified DNA fragment from the dogs and the ticks infesting them were identical, and the sequence corresponded to that of the E. platys Gzh981 strain. We concluded that there is a cyclic maintenance of E. platys between dogs and ticks in Okinawa. PMID- 11270613 TI - A non-surgical rat model of foreign body-associated urinary tract infection with Pseudomonas aeruginosa. AB - This study established a rat model of foreign body-associated urinary tract infection. A spiral polyethylene tube (PT) was placed transurethrally into the bladder without surgical manipulation, followed by transurethral inoculation with Pseudomonas aeruginosa. The persistence of P. aeruginosa in the kidneys and bladder was significantly enhanced by placement of the PT, whereas the bacteria were eliminated rapidly from the urinary tract in the animals without the PT. Scanning electron microscopy revealed a thick biofilm on the surface of the PT from the early stage of infection. Histopathologically, acute pyelonephritis was followed by chronic renal inflammation as well as continuous and sporadic polymorphonuclear leukocyte accumulation and hemorrhage in the pelvis and adjacent tissues, suggesting continuous ascending introduction of the bacteria from the biofilm adhering to the PT. We believe our model simulates the pathophysiology of foreign body-associated urinary tract infection characterized by biofilm formation on the surface of a foreign body. PMID- 11270614 TI - Serological analysis of human leptospirosis in the Philippines. AB - Leptospirosis in the Philippines is an underrepresented disease. To achieve an accurate means of serodiagnosis, we demonstrated antibodies to the prevalent Leptospira serovars in sera of 71 patients from three major hospitals in Manila by the microscopic agglutination test and Western blot analysis. Sera of 53 patients contained antibody against 8 serovars poi, tarassovi, manilae, pyrogenes, australis, grippotyphosa, javanica, and autumnalis. PMID- 11270615 TI - The significance of cytomegalovirus infection over the clinical course of adult T cell leukemia/lymphoma. AB - The significance of cytomegalovirus (CMV) infections developed over the clinical course of adult T-cell leukemia/lymphoma (ATLL) were evaluated in relation to the patient survival rate, ATL activity and immunocompetent cells. ATLL patients with CMV infections on admission exhibited a poor survival rate, while patients with CMV infections at any time after admission survived longer than those not infected with this virus. ATLL patients who exhibited a numbers of CMV infection on admission showed higher ATL activity and had lower numbers of CD8-positive and CD56-positive cells than those who developed CMV infections at any time after admission. Therefore, it appears likely that patients with CMV infections on admission were in an immunosuppressive state due to aggressive ATL activity. PMID- 11270616 TI - NF-kappaB activation and IkappaB alpha dynamism involved in iNOS and chemokine induction in astroglial cells. AB - This review will discuss the recent literature on the molecular mechanism of NF kappaB activation, with special focus on IkappaB alpha dynamism involved in iNOS- and chemokine-induction in glial cells. NF-kappaB, a heterotrimer composed of p50, p65 (Rel A) and IkappaB alpha, has been shown to be activated by elimination of the regulatory subunit IkappaB alpha from the heterotrimer. The elimination of IkappaB alpha (formation of active NF-kappaB, p50-p65) is due to phosplorylation of serines 32 and 36 of IkappaB alpha, followed by polyubiquitination and 26S proteasomal degradation of IkappaB alpha. Experiments using stable clones of rat C6 glioma cells transfected with dominant negative IkappaB alpha (serines 32 and 36 replaced by alanine) suggest that NF-kappaB activation (phosphorylation of IkappaB alpha) is involved in LPS/IFNgamma- or IL-1beta/IFNgamma-induced iNOS expression. Furthermore, the time courses of phosphorylation, ubiquitination of IkappaB alpha and proteasome activity after IL-1beta treatment also suggest that 26S proteasomal degradation of IkappaB alpha is more crucial for chemokine expression in glial cells. PMID- 11270617 TI - Regional expression of the splice variants of Kv4.3 in rat brain and effects of C terminus deletion on expressed K+ currents. AB - In situ hybridization and RT-PCR analyses have revealed that, among three Kv4.3 splice variants (a, b, and c) with distinct C-terminal cytoplasmic domains, the mRNA for Kv4.3a is abundant in cerebral cortex, cerebellum, olfactory bulb, and medulla oblongata, whereas the mRNA for Kv4.3c is localized mainly to hippocampus. Three new distinct splice variants of Kv4.3 (Kv4.3d, e and f), which consist of 601, 635, and 628 amino acids, respectively, and have distinct C terminal cytoplasmic domains, were isolated from rat brain by RT-PCR. Kv4.3b, d, e and f were expressed at much lower levels in brain. Mutagenesis which removed 149 amino acids in C-terminal domain of Kv4.3a significantly slowed its rate of recovery from inactivation as measured in heterologous expression in HEK293 cells. Surprisingly, however, neither the rate of inactivation nor voltage dependence of the activation and inactivation were changed. PMID- 11270618 TI - The novel kappa-opioid receptor agonist TRK-820 suppresses the rewarding and locomotor-enhancing effects of morphine in mice. AB - The effects of the novel kappa-opioid receptor agonist TRK-820 on the rewarding and locomotor-enhancing effects of morphine were investigated in mice. Morphine (1-5 mg/kg, s.c.) caused a dose-related preference for the drug-associated place. In contrast, TRK-820 (0.003-0.03 mg/kg, s.c.) did not produce a significant preference for either compartment of the test box. In combination studies, co injection of TRK-820 (0.01 and 0.03 mg/kg, s.c.) with morphine significantly suppressed the morphine (5 mg/kg, s.c.)-induced place preference, and this effect of TRK-820 was antagonized by pretreatment with nor-BN1 (3 mg/kg, s.c.), a selective kappa-opioid receptor antagonist. TRK-820 also suppressed morphine induced hyperlocomotion, and this suppression was also blocked by nor-BNI. These results suggest that TRK-820 suppresses the rewarding and locomotor-enhancing effects of morphine through the activation of kappa-opioid receptors. Thus, we propose that TRK-820 may be useful for controlling pain while reducing undesirable side-effects. PMID- 11270619 TI - Immunohistochemical and in situ hybridization studies of gonadotropin releasing hormone (GnRH) and its receptor in rat digestive tract. AB - GnRH(LH-RH) is first discovered in the hypothalamus and found to have a role in regulation of reproduction. With the study on it deepening, GnRH was demonnstrated that it also exists in a number of organs beyond the hypothalamus and acts on extrapituitary organs. To study whether digestive tract synthesizes GnRH and its receptor and, if it does, by what cells. In the experiment, the locallizations of GnRH and its receptors in rat digestive tract were studied using immunohistochemistry and in situ hybridization. The parietal cells of gastric gland, the villous and glandular epithelium in small and large intestine and parasympathetic ganglion cells of myenteric plexus showed GnRH immunoreactivity; GnRH mRNA hybridization signal was detected. The epithelium of gastric pit and the cells above in digestive tract showed GnRH receptor immunoreactivity; GnRH receptor mRNA hybridization signal was detected. The immunoreactive and signal materials distributed in cytoplasm of all positive cells, with nuclei being immunonegative and with no hybridization signal. These results suggested that the digestive tract can produce GnRH and express GnRH receptor; GnRH may also be a gastrointestinal hormone. PMID- 11270621 TI - Dihydropyridine calcium channel blockers inhibit ascorbic acid accumulation in human intestinal Caco-2 cells. AB - We investigated the effect of commonly used medications on the accumulation of ascorbic acid in human intestinal Caco-2 cells. Although ascorbic acid is negatively charged at physiological pH, anionic compounds including drugs and metabolites had little effect on its accumulation. On the other hand, hydrophobic 1,4-dihydropyridine compounds (nifedipine and nicardipine), but not other structurally unrelated calcium channel blockers, were found to be potent inhibitors. They inhibited both Na+-dependent and Na+-independent (K+ substituting Na+) accumulation of ascorbic acid. The inhibition was non competitive with a Ki of 108 microM and 9 microM for nifedipine and nicardipine, respectively. The efflux of ascorbic acid from cells was not affected. Previously, we reported a similar inhibition of ascorbic acid accumulation by estrogens. When nifedipine and estrogens were included in the buffer together, the combined inhibitory effect was less than additive implying that they may act through the same mechanism. The potential clinical significance of dihydropyridine usage on ascorbic acid status in human needs to be considered. PMID- 11270620 TI - The role of oxidative stress in developmental and reproductive toxicity of tamoxifen. AB - The antiestrogen tamoxifen (TAM) is widely used as a drug against breast cancer and is currently being tested as a chemopreventive agent. However, a number of studies showed genotoxic and carcinogenic effects of TAM. These effects are thought to be related to oxygen radical overproduction which occurs during TAM metabolic activation. There is no evidence, thus far, on TAM toxicity to embryos and gametes. The present study was designed to elucidate the mechanisms of TAM induced developmental, reproductive and cytogenetic toxicity towards sea urchin (SU) embryos with regard to the possibility of TAM-initiated oxidative stress. Embryo cultures from SU were subjected to long-term (throughout embryogenesis) or short-term (two hours) incubation with TAM at concentrations from 10(-8) to 10( 5) M. The experiments on TAM-induced toxicity to gametes were carried out with SU sperm, or unfertilized eggs, suspended in TAM (10(-8) to 10(-6) M). To assess the effects of TAM to embryos or to gametes, developmental defects, embryonic mortality, fertilization success, and cytogenetic abnormalities were scored. Oxidative damage to DNA and lipids was detected by measurements of 8OHdG levels and lipid peroxidation, respectively. Reactive oxygen species (ROS) production by eggs and embryos was recorded by luminol-dependent chemiluminescence (LDCL) and cytochrome c reduction methods. The changes in activities of SU superoxide dismutase (SOD) and catalase were also evaluated. TAM exerted: a) early embryonic mortality to exposed embryos and to the offspring of exposed eggs; b) developmental defects to the offspring of exposed sperm; c) decrease in sperm fertilization success, and d) cytogenetic effects in the offspring of exposed sperm or eggs. These morphological effects corresponded to the state of oxidative stress in SU embryos (increased oxidative damage to DNA and lipids and induction of antioxidant enzymes). Since TAM did increase significantly ROS production by embryos, it is suggested that TAM may be metabolically activated by SU embryonic oxidases and peroxidases, which in turn could be induced by TAM. The present study provides further support to the utilization of the SU system as a useful model to help elucidate mechanisms of chemical teratogenesis and carcinogenesis. PMID- 11270622 TI - NMDA-responsible, APV-insensitive receptor in cultured human astrocytes. AB - The present study was conducted to assess N-methyl-D-aspartate (NMDA)-responsible receptors in cultured human astrocytes by monitoring whole-cell membrane currents. NMDA generated currents, that are potentiated by glycine and blocked by Mg2+, with the current/voltage relation showing a reversal potential of +/- 0 mV. The currents were not inhibited by either the selective NMDA receptor antagonist, D-2-amino-5-phosphonovaleric acid (APV), or the non-selective ionotropic glutamate receptor antagonist, kynurenic acid. The currents were inhibited only by 19% in Ca2+-free extracellular solution. Furthermore, GDPbetaS, a broad G protein inhibitor, inhibited NMDA-induced currents to 82% of original levels. The results of the present study thus suggest that an NMDA-responsible, APV insensitive receptor with low Ca2+ permeability, distinct from the neuronal NMDA receptors, is expressed in human astrocytes and that the receptor is regulated in part by an unknown G-protein-linked receptor. PMID- 11270623 TI - Effect of androgenic anabolic steroids on sperm quality and serum hormone levels in adult male bodybuilders. AB - The purpose of this study was to assess the influence of the administration of high doses of androgenic anabolic steroids (AAS) on endocrine and semen parameters. Thirty volunteering bodybuilders were studied (ages ranging between 26.6 +/- 4.1 years). A history of anabolic steroid administration was recorded for fifteen subjects, and results of semen analysis and endocrine parameters were compared with data from fifteen bodybuilders not using steroids. In those subjects using AAS, eight had sperm counts under the lower normal limit (20 x 10(6) sperm/ml), three had azoospermia, two polyzoospermia, and two had normal sperm counts. The percentage of morphologically normal sperm was significantly reduced, only 17.7% had normal spermatozoa. In the control group, only one subject had oligozoospermia. The hormonal parameters revealed reduced FSH (1.5 +/ 3.2 vs 5.0 +/- 1.6, p < 0.001) and PRL (5.1 +/- 4.9 vs 9.2 +/- 4.4, p < 0.01) levels. LH, T, E2 and DHEA levels did not vary. PMID- 11270624 TI - Evidence for specific analgesic activity of a muscarinic agonist selected among a new series of acetylenic derivatives. AB - The central and peripheral effects of a series of Oxotremorine/Oxotremorine-M derivatives, previously characterized as muscarinic agonists in isolated preparations, were investigated in in vivo experiments. The molecules were tested for their antinociceptive activity (formalin licking and acetic acid writhing tests) and for their ability to induce tremor in mice. Peripheral cholinergic effects such as salivation, bradycardia, hypotension and intestinal hypermotility were studied in anaesthetized rats. All of the acetylenic compounds acted as muscarinic analgesics displaying the same order of potency shown in in vitro studies. The Oxotremorine-like subset showed a clearer distinction between doses producing antinociception and doses exerting undesirable central/peripheral side effects compared to the Oxotremorine-M derivatives. The most promising profile was displayed by the isoxazolin-3-one Oxotremorine-like derivative (compound 1a), which was characterized by a wider therapeutic window than that of the parent molecule Oxotremorine. Indeed, it produced atropine-sensitive analgesia (ID50 about 0.1 mg/kg i.p.) in the absence of tremorogenic (EC50 2.73 mg/kg i.p.) and cardiovascular effects while lethality occurred only at higher doses (LD50 19 mg/kg i.p.). These results suggest that such a derivative could be a candidate for further development of selective muscarinic analgesics. PMID- 11270625 TI - Metabolic fate of 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine by human spermatozoa: biosynthesis and biodegradation. AB - Human spermatozoa can synthesise 1-alkyl-2-acetyl-glycerophosphocholine (AAGPC) in small amount by acetyltransferase (AT) in absence of any stimulus, but can actively catabolise it by acetylhydrolase (HY). Seminal plasma, on the other hand, was devoid of anabolic enzyme albeit enrich in catabolic enzyme, suggesting as an active site for biodegradation of AAGPC secreted by spermatozoa. Both, AT and HY exhibited pH-optima in range of 7.0-7.6 at which spermatozoa are maximum viable and motile. Ionophore A23187 and EGTA inhibited AT, reversibly, whereas HY was inhibited by BSA, calcium-channel blockers, and phospholipase A2-inhibitors. Effect of aging-time on ejaculates exhibited decreased AT activity with increased HY activity along with unchanged calcium content of spermatozoa. Serotonin in vitro studies showed a pro-aggregator role on agglutination of spermatozoa. Viscid/long liquefaction time ejaculates exhibited raised AT activity and calcium contents with decreased HY activity in spermatozoa and high degree of agglutination. Studies with dithiothreitol-treatment indeed helped in liquefaction but levels of both enzymes remained status quo, suggesting existence of both pathways: remodelling of membrane phospholipids and de novo synthesis of AAGPC in spermatozoa, earlier being pre-dominant. We have proposed a role of AAGPC-Serotonin-Calcium in agglutination and liquefaction of spermatozoa, a vital aspect in normal fertility. PMID- 11270626 TI - Singlet oxygen generation from phosphatidylcholine hydroperoxide in the presence of copper. AB - This study pursued whether singlet oxygen ((1)O2) is generated from phosphatidylcholine hydroperoxide (PCOOH), the oxidized modification product of a major constituent of biomembranes and serum lipoproteins. The (1)O2 formation was detected, by utilizing the oxidation of 2,2,6,6-tetramethyl-4-piperidone (TMPD) by (1)O2 to yield 2,2,6,6-tetramethyl-4-piperidone-1-oxyl (TEMPONE), which generates electron spin resonance (ESR) signals. The TEMPONE signal was detected in human plasma with addition of PCOOH by ESR determination after introducing copper(II). The TEMPONE formation was proportional to the amounts of PCOOH added according to moles of active oxygen. The TEMPONE signal intensity was weakened significantly in the presence of beta-carotene and histidine in a concentration dependent manner, but was not at all decreased by mannitol, Mn-superoxide dismutase and catalase. In addition, HPLC-chemiluminescence analysis demonstrated that incubation with the PCOOH/Cu(II) combination oxidized cholesterol, a relatively oxidation-resistant component, to the cholesterol hydroperoxide. These results reveal that (1)O2 is generated from PCOOH in contact with copper(II). In conclusion, this in-vitro study provides directly the (1)O2 formation in living organisms following the advancement of peroxidation of constitutive lipids. PMID- 11270627 TI - Application of time delay spectrometry for rough surface characterization. AB - This paper describes the design and performance of an ultrasound measurement system, utilizing a swept frequency excitation signal, for analyzing the backscattered signal from planar rough surfaces. The implementation is in the form of a time delay spectrometry (TDS) system operating in reflection mode whose advantages are improved signal-to-noise ratio even with low peak power relative to conventional pulse-echo methods. Because of simultaneous transmission and reception, the TDS system requires two transducers. The TDS measurement system uses a swept frequency spectrum analyzer as the central analog processing unit. Both planar piston and focused transducers in the low MHz range were used for the measurements. Due to the statistical nature of rough surface backscatter, the mean of several statistically uncorrelated measurements is required to characterize the scattering behavior of a given rough surface. Backscatter data are obtained for a series of planar rough surfaces, in the form of scattering magnitude vs. frequency and vs. incident (=backscattered) angle. Analysis of the results reveals a good correlation between the root-mean-square (RMS) height and mean backscatter magnitude at 0 degrees incident angle, and between the ratio of RMS height to correlation length and the difference in mean backscatter magnitude between 0 degrees and 5 degrees. PMID- 11270628 TI - The role of internal reflection in transskull phase distortion. AB - Phase distortion due to reflection in transcranial ultrasound propagation is investigated. Understanding of these phase-dependent properties is motivated by efforts to construct a reliable prediction model for noninvasive ultrasound therapy in the brain. The present study measures the phase of an ultrasound wave after propagation through an ex vivo human skull and considers the dependence of this phase on reflections between the transducer and the skull surface in addition to reflections within the skull. Experiments are performed using a human calvarium fragment placed between an underwater ultrasonic transducer and a polyvinylidene difluoride hydrophone. Data are presented indicating the ultrasound phase dependence as a function of burst length and the distance of the transducer element from the skull at a driving frequency of 0.5 MHz. Experimental results are compared with predictions obtained from a propagation model which considers transmission at the skull interfaces as well as multiple reflections within the skull. It is concluded that by using short ultrasound bursts a distance may be indicated that beyond which the contributions of transducer reflections on the phase of the propagating wave may be neglected. Additionally, a comparison of the measurements with simulated data supports the contention that for reasonably small incident angles, reflection within the skull causes minimal phase shift. PMID- 11270629 TI - Performances estimation of a rotary traveling wave ultrasonic motor based on two dimension analytical model. AB - The understanding of ultrasonic motor performances as a function of input parameters, such as the voltage amplitude, driving frequency, the preload on the rotor, is a key to many applications and control of ultrasonic motor. This paper presents performances estimation of the piezoelectric rotary traveling wave ultrasonic motor as a function of input voltage amplitude and driving frequency and preload. The Love equation is used to derive the traveling wave amplitude on the stator surface. With the contact model of the distributed spring-rigid body between the stator and rotor, a two-dimension analytical model of the rotary traveling wave ultrasonic motor is constructed. Then the performances of stead rotation speed and stall torque are deduced. With MATLAB computational language and iteration algorithm, we estimate the performances of rotation speed and stall torque versus input parameters respectively. The same experiments are completed with the optoelectronic tachometer and stand weight. Both estimation and experiment results reveal the pattern of performance variation as a function of its input parameters. PMID- 11270630 TI - Inversion of synthetic and experimental acoustical scattering data for the comparison of two reconstruction methods employing the Born approximation. AB - This work is concerned with the reconstruction, from measured (synthetic or real) data, of a 2D penetrable fluid-like object of arbitrary cross-section embedded in a fluid of infinite extent and insonified by a plane acoustic wave. Green's theorem is used to provide a domain integral representation of the scattered field. The introduction therein of the Born approximation gives rise to a linearized form of the inverse problem. The actual inversion is carried out by two methods. The first diffraction tomography (DT), exhibits the contrast function very conveniently and explicitly in the form of a wave number/incident angle Fourier transform of the far backscattered field and thus requires measurements of this field for incident waves all around the object and at all frequencies. The second discretized domain integral equation with Born approximation method, is numerically more intensive, but enables a wider choice of configurations and requires less measurements (one or several frequencies, one or several incident waves, choice of measurement points) than the DT method. A comparison of the two methods is carried out by inversion of both simulated and experimental scattered field data. PMID- 11270631 TI - Characterization of wheat-flour-water doughs: a new method using ultrasound. AB - In this paper, an original method of evaluating the physical properties of wheat flour-water systems using high-frequency low-power ultrasound is presented. Most of the experiments were performed with a reflectance technique measuring the acoustic impedance of doughs. The velocity of propagation, attenuation and viscoelastic moduli have been evaluated for both compressional and shear ultrasonic waves in the interval 2-10 MHz for doughs of different hydrations. The 53% water content was found to be critical with respect to the presence of free water. The influence of the mixing and rest times on the longitudinal ultrasonic parameters is also studied. PMID- 11270632 TI - Flow Rate Profiler: an instrument to measure blood velocity profiles. AB - In this paper we present Flow Rate Profiler (FRP), an instrument for measuring the blood velocity by means of ultrasound-based techniques. The velocity is directly related to the shear rate, which is in turn proportional to the shear stress, a parameter expressing the pressure exerted by the blood on the vessel walls. The knowledge of this value is important in medicine to establish the state of the vessels, directly related to vascular diseases. FRP provides a non invasive measure of the blood velocity by exploiting the red corpuscles property of diffusing ultrasound waves: in practice blood velocity is determined by a cross-correlation technique, which analyses the time shift between correlated subsequent echo waves, instead of frequency shift characteristic of the Doppler technique. The acquired data are then processed on a personal computer by means of mathematical techniques based on the evaluation of the correlation function, giving a reconstructed velocity profile and showing a good adherence with experimental data, since the average error is nearly the 10%. The reconstructed profile is displayed to the operator, who can follow the vessel status in real time. A few comparisons between the reconstructed and the experimental profiles are also presented, together with a study on a small set of patients suffering from artery hypertension. PMID- 11270633 TI - Clad polymer buffer rods for polymer process monitoring. AB - Clad polymer buffer rods consisting of a polyetheretherketone (PEEK) core and a cladding made of a heat resistance epoxy aluminum composite are presented. The core has a uniform diameter or a taper shape. Ultrasonic measurement results indicate that the ultrasonic signal strength and signal to noise ratio of these clad rods are better than those of the non-clad PEEK rods for both longitudinal and shear waves because of the improved ultrasonic wave guidance in the core. Comparisons of these rods with those made of polymide and high-density polyethylene are given. Applications of these buffer rods for ultrasonic monitoring of polymer extrusion at temperatures up to 200 degrees C and pressures up to 180 psi are demonstrated. The monitoring results also reveal that within certain operating temperature and pressure range, clad polymer buffer rods show advantages over clad steel buffer rods. PMID- 11270634 TI - A miniature class V flextensional cymbal transducer with directional beam patterns: the double-driver. AB - The "double-driver" cymbal, a directional class V flextensional transducer, is described in this paper. Its basic structure is a bilaminar piezoelectric disk with metal caps as mechanical transformers and amplifiers. The directivity was accomplished by exciting the double-driver in a combined flexural and bending mode causing the sound pressure to add in one direction and cancel in the opposite direction. The cardioid beam pattern predicted by finite element modeling agreed well with the experimental measurements. A 3 x 3 double-driver array was constructed to demonstrate that under optimal conditions the array can provide a directional beam pattern with a front-to-back ratio of more than 20 dB. PMID- 11270635 TI - Cooperative effects in multi-bubble sonoluminescence. AB - Theoretical and experimental evidence is presented in support of quantum collective effects in sonoluminescence. PMID- 11270636 TI - Cytokine production by natural killer lymphocytes in follicular and luteal phase of the ovarian cycle in humans. AB - PROBLEM: The aim of this study was to test the hypothesis that, during luteal phase of the ovarian cycle, as compared with follicular phase, the cytokine productive capacity of peripheral natural killer (NK)-lymphocytes in humans is shifted towards a "Th2-type"-like response. METHOD OF STUDY: Intracellular Th1 and Th2 cytokine production by in vitro activated peripheral NK-lymphocytes in a whole blood preparation of the follicular and the luteal phase of the ovarian cycle was measured by flow cytometry. RESULTS: There was no difference in interferon (IFN)-gamma, interleukin (IL)-2, IL-4, and IL-10 cytokine production in activated NK-lymphocytes when comparing luteal phase with follicular phase of the ovarian cycle. However, there was a significant increase in peripheral NK lymphocyte number in luteal phase compared with follicular phase. CONCLUSION: The cytokine productive capacity of peripheral NK-lymphocytes is not shifted towards a "Th2-type"-like response in the luteal phase as compared with the follicular phase of the ovarian cycle in humans. PMID- 11270637 TI - Immunohistochemical staining of IL-1 alpha and IL-1 receptor antagonist but not IL-1 beta in cultures of sertoli cells. AB - The interleukin-1 (IL-1) system has been suggested to be involved in the cell cell cross talk within the testis. To identify a testicular cell source of IL-1 alpha, IL-1 beta and IL-1 receptor antagonist (IL-1ra), immature mouse Sertoli cells were isolated, purified, cultured and examined for the cellular compartment localization of these cytokines by immunohistochemical staining. Our results show that both Germ cells and Sertoli cells in unpurified Sertoli cell cultures (before hypotonic shock) and purified culture of Sertoli cells (after hypotonic shock) were stained for IL-1 alpha. The levels of this cytokine were increased in Sertoli cells when the purified cultures were stimulated with lipopolysaccharide (LPS) (5 microg/mL). However, we could not identify a positive staining for IL-1 beta when Sertoli cell cultures were stained for this cytokine, even after stimulation with various concentrations of LPS (0.1-10 microg/mL). On the other hand, immunohistochemical staining of isolated Sertoli cells without treatment with hypotonic shock (cultures containing Sertoli cells and Germ cells) for IL 1ra showed constitutive positive staining of both cell types (Sertoli cells and Germ cells). Our results, using immunohistochemical staining, may indicate the different expression of IL-1 alpha, IL-1 beta and IL-1ra in Sertoli cells. These results may suggest the involvement of IL-1 system in the autocrine and paracrine regulation of testicular cell functions. PMID- 11270638 TI - Interleukin-1beta-induced prostaglandin E2 production in human myometrial cells: role of a pertussis toxin-sensitive component. AB - PROBLEM: The objective of this study was to evaluate the possible role of pertussis toxin (PTX)-sensitive G-protein(s) in interleukin-1beta (IL-1) signaling in human myometrial cells (HMC). METHOD: Primary cultures of HMC were stimulated with human recombinant IL-1 alone or in combination with PTX. Prostaglandin (PG) E2 in the medium was measured by radioimmunoassay, cyclooxygenase type 2 (Cox-2) and IkappaB by western analysis, and the activities of two members of the mitogen-activated protein kinase (MAPK) family of enzymes, ERK-2 and JNK, by the phosphorylation of appropriate substrates. RESULTS: IL-1 increased PGE2 output during an 18-hr long incubation by 21.7-fold (n = 5 experiments). This increase was inhibited by 57% after pretreatment overnight with PTX. IL-1-induced expression of Cox-2 protein was also suppressed to a similar degree in PTX-treated HMC cultures. Degradation of the nuclear factor kappa B (NF-kappaB)-inhibiting protein (IkappaB), a critical step in IL-1 signaling to the nucleus, was significantly inhibited by PTX, as was IL-1-induced activation of ERK-2 and JNK. CONCLUSIONS: It is suggested that the occupied IL-1 receptor-generated signal in HMC is transmitted by multiple pathways. One is coupled to a PTX-sensitive G-protein upstream from the MAPK phosphorylation cascade. This, in turn, may interact with another signaling pathway, the activation of NF-kappaB, via the phosphorylation of the IkappaB kinase complex. PMID- 11270639 TI - Interleukin-6 up-regulates the oxytocin receptor in cultured uterine smooth muscle cells. AB - PROBLEM: Intra-uterine infection results in the production of inflammatory cytokines, including interleukin-6 (IL-6). Increased oxytocin-receptor (OTR) concentrations are associated with the onset of preterm labor. We hypothesize that infection up-regulates OTR expression through IL-6-induced transcription factors. METHOD OF STUDY: Primary cultures of human myometrium were treated for various time periods or with different concentrations of IL-6 and OTR mRNA as well as OTR binding were measured by means of reverse transcription polymerase chain reaction and 125I-ornithine-vasotocin-binding assay. To study underlying mechanisms of OTR changes with IL-6 treated, cells were also incubated with genistein or H7 (tyrosine and serine phosphorylation inhibitors), respectively. RESULTS: OTR mRNA increased 2.5-fold after 4 hr of IL-6 treatment and OTR binding 1.4-fold after 8 hr of cytokine stimulation. The IL-6-induced increase in binding was blocked by genistein and H7. CONCLUSIONS: IL-6 up-regulates uterine OTR mRNA expression and binding capacity in cultured human myocytes most likely through tyrosine and serine phosphorylation pathways involving the nuclear factor STAT-3. PMID- 11270640 TI - Soluble endothelial and platelet selectins in serum and ascitic fluid of women with ovarian hyperstimulation syndrome. AB - PROBLEM: The selectins, a group of cell adhesion molecules, are major mediators of inflammatory, immunologic, and angiogenic reactions. Their possible involvement and levels of the soluble isoform in serum and ascitic fluid of women with ovarian hyperstimulation syndrome (OHSS) were not previously evaluated. METHOD OF STUDY: This prospective, case-control study involved 16 women with OHSS. Ten matched women treated by controlled ovarian stimulation and eight healthy women with normal diagnostic laparoscopy served as controls. Serum and peritoneal fluid samples obtained from all subjects were assayed for soluble endothelial selectin (sES) and soluble platelet selectin (sPS) by specific enzyme linked immunosorbent assay. RESULTS: Significantly higher levels of sES (median, 17.1 [9-41] vs 9.3 [2.4 21.3] ng/mL [P = 0.03]) and sPS (median, 23 [13-277] vs 6.5 [1.6-28.7] ng/mL [P = 0.001]) were observed in the peritoneal fluid of women with OHSS than the basal levels of healthy, non-stimulated women. Women with OHSS had significantly lower sES serum levels than those treated by controlled ovarian hyperstimulation without OHSS, while their sPS serum levels were comparable. CONCLUSIONS: Ascitic fluid of women with OHSS contains appreciable amounts of soluble selectins, suggesting their ovarian origin and possible involvement in the syndrome. PMID- 11270641 TI - Association between interleukin concentration in follicular fluid and intracytoplasmic sperm injection (ICSI) outcome. AB - PROBLEM: The aim of this study was to determine the presence and concentration of interleukin IL-6, IL-8, and granulocyte-macrophage-colony-stimulating factor (GM CSF) in pre-ovulatory ovarian follicular fluid (FF) of patients undergoing controlled ovarian hyperstimulation for intracytoplasmic sperm injection (ICSI) therapy on one hand, and to find out whether these cytotokine concentrations could be used as a predictive parameter for ICSI outcome. DESIGN: The levels of IL-6, IL-8, and GM-CSF were measured from women that underwent ICSI therapy and the results were compared between the patients who became pregnant after IC PMID- 11270642 TI - Polymorphism of human leukocyte antigen-E gene in the Japanese population with or without recurrent abortion. AB - PROBLEM: The aim of this study was to investigate the gene frequencies and shared alleles of human leukocyte antigen (HLA)-E gene in Japanese couples with or without recurrent abortion. METHOD OF STUDY: Polymerase chain reaction (PCR) single strand conformation polymorphism (SSCP) analysis was carried out to detect polymorphism in exon 3 of the HLA-E gene in 30 Japanese couples with recurrent abortion and 38 normal Japanese couples with proven fertility. RESULTS: No point mutation was detected in exon 3 of HLA-E in both recurrent aborters and normal controls. HLA-EG and HLA-ER alleles were detected with frequencies of 66.7% and 33.3% in couples with recurrent abortion and 69.2% and 30.8 in normal couples, respectively. The gene frequency of HLA-EG was higher than that of HLA-ER, which is contrary to that found in Caucasian, African-American and Hispanic people but similar to Chinese people. The frequency of each allele was not significantly different between recurrent aborters and normal controls. The number of shared alleles between each couple with recurrent abortion is not significantly different from that with normal controls. CONCLUSION: Allele frequencies of HLA-E were suggested to be different in Asian people from those in other ethnic people. In light of no specific distribution pattern in recurrent aborters, HLA-E polymorphism does not seem to play a role in the pathogenesis of recurrent abortion. PMID- 11270643 TI - Pregnancy outcome in recurrent spontaneous abortion associated with antiphospholipid antibodies: a comparative study of intravenous immunoglobulin versus prednisone plus low-dose aspirin. AB - PROBLEM: To compare the use of intravenous immunoglobulins (IVIG) with prednisone plus low-dose aspirin (LDA) in treating pregnant women with a history of recurrent fetal loss having the antiphospholipid antibody (aPL), in terms of live birth rate and maternal and perinatal morbidity. METHOD: A prospective, two centers trial study included 82 recurrent aborters with aPL syndrome. Twenty-nine were treated with prednisone and LDA in one center, 53 received IVIG in the other center. Maternal and fetal outcomes and pregnancy complications were compared between groups. RESULTS: Live-birth rates were equivalent between groups (78 vs 76%). Mean birth weight was higher in the IVIG group than in the prednisone plus LDA group. In the prednisone- plus LDA-treated patients, gestational hypertension and gestational diabetes were found significantly more often than in the IVIG treated group (14 vs 5% and 14 vs 5%, respectively). CONCLUSION: In patients with aPL syndrome, IVIG treatment improved pregnancy outcome, with significantly lower pregnancy complication rates, when compared with prednisone plus LDA therapy. PMID- 11270645 TI - Does seminal plasma nitrite reflect nitrite + nitrate concentrations in leukocytospermic infertile men? PMID- 11270644 TI - Serum levels of inhibin B, unlike inhibin A and activin A, are not altered in women with preeclampsia. AB - PROBLEM: To investigate whether inhibin A, inhibin B, and activin A serum levels are altered in women with preeclampsia. METHOD OF STUDY: Serum samples of 20 women with preeclampsia (study group) and 20 normotensive pregnant women, matched for maternal and gestational age and parity, were assayed for inhibin A, inhibin B and activin A by specific enzyme-linked immunosorbent assay. RESULTS: Median serum concentrations of inhibin A and activin A were significantly higher among women with preeclampsia than in women with normotensive pregnancies, while inhibin B levels were comparable in both groups. Activin A levels were positively correlated with those of inhibins A and B, and inhibin A levels were positively correlated with diastolic blood pressure and inhibin B concentration in the study group. CONCLUSIONS: Inhibin A and activin A, but not inhibin B, serum levels are markedly increased in women with preeclampsia. These hormones might serve as an endocrine marker for preeclampsia. PMID- 11270646 TI - Adhesion of the dissimilatory Fe(III)-reducing bacterium Shewanella alga BrY to crystalline Fe(III) oxides. AB - Shewanella alga BrY adhesion to hydrous ferric oxide, goethite, and hematite was examined. Adhesion to each oxide followed the Langmuir adsorption model. No correlation between adhesion and Fe(III) oxide surface area or crystallinity was observed. Zeta potential measurements suggested that electrostatic interactions do not influence S. alga BrY adhesion to these minerals. Cell adhesion does not appear to explain the recalcitrance of crystalline Fe(III) oxides to bacterial reduction. PMID- 11270647 TI - Xanthomonas albilineans diversity and identification based on rep-PCR fingerprints. AB - PCR with BOX and ERIC primers was used to analyze DNA of Xanthomonas albilineans and other bacteria associated with sugarcane. Generated fingerprints permitted clear separation of X. albilineans from other bacteria and revealed variation within the species. Good agreement between fingerprint groups and geographic origin and serovars was observed. PMID- 11270648 TI - Effect of genomic location on horizontal transfer of a recombinant gene cassette between Pseudomonas strains in the rhizosphere and spermosphere of barley seedlings. AB - The use of genetically engineered bacteria in natural environments constitutes a risk of transfer of recombinant DNA to the indigenous bacteria. However, chromosomal genes are believed to be less likely to transfer than genes on mobilizable and conjugative plasmids. To study this assumption, horizontal transfer of a recombinant gene cassette inserted into the chromosome of a Pseudomonas stutzeri strain, into a mobilizable plasmid (pAGM42), and into a conjugative plasmid (pKJK5) isolated from barley rhizosphere was investigated. Horizontal transfer efficiencies of the gene cassette inserted into a conjugative plasmid was 8.20 x 10(-3) transconjugants/(donors x recipients)1/2 in the rhizosphere and 4.57 x 10(-2) transconjugants/(donors x recipients)1/2 in the spermosphere. Mobilization of the plasmid pAGM42 by the plasmids RP4 and pKJK5 was also detected at high levels in the microcosms, transfer efficiencies were up to 4.36 x 10(-3) transconjugants/(donors x recipients)1/2. Transfer of chromosomal encoded genes could not be detected in the microcosms by conjugation or transformation. However, transformation did occur by using the same bacterial strains under laboratory conditions. The rhizosphere and especially the spermosphere thus proved to be hot spot environments providing favorable conditions for gene transfer by mobilization and conjugation, but these environments did not support transformation at a detectable level. PMID- 11270649 TI - DNase-resistant DNA in the extracellular and cell wall-associated fractions of Frankia strains R43 and CcI3. AB - DNases were shown to be present in the extracellular fraction of Frankia strains R43 and CcI3. In spite of this, DNA was found in both the extracellular and cell wall fractions of these strains, and it was shown that extracellular DNA was resistant to the DNases secreted into the culture medium of both Frankia strains. Furthermore, Southern blot analysis under high stringency conditions revealed the chromosomal origin of the cell wall-adsorbed DNA (CW-DNA). Mobility gel band shift assays suggested that the extracellular DNA and the CW-DNA are engaged in complexes with other molecules, most likely proteins, which are probably responsible for the enzymatic resistance observed against extracellular DNase activities. In addition, it was shown that lysis of a small proportion of the cells in the exponential growth phase may account for the DNA being released into the supernatant and adsorbed to the cell wall. PMID- 11270650 TI - Enhanced tolerance against salt-stress and freezing-stress of Escherichia coli cells expressing algal bbc1 gene. AB - The expression of the eukaryotic bbc1 (breast basic conserved) gene (the bbc1 gene of the marine green alga Chlamydomonas sp. W-80 strain) enhanced the tolerance against salt-stress and freezing-stress in E. coli cells. The expression of the BBC1 protein in the E. coli cells carrying the algal bbc1 gene and that in the Chlamydomonas W-80 cells were examined by Western blotting analysis. The result suggests that the eukaryotic BBC1 protein expressed in the E. coli cells has a protective function against the cellular dehydration. PMID- 11270651 TI - Differences in the regulation of the intracellular Ca2+-dependent serine proteinase activity between Bacillus subtilis and B. megaterium. AB - A rise of the intracellular serine proteinase activity (ISP) during postexponential growth of Bacillus subtilis was decreased by a temperature upshift from 35 degrees to 42 degrees C. However, the amount of both molecular forms of the major intracellular serine proteinase ISP1 determined by immunoblotting was similar at both temperatures or even slightly increased at 42 degrees C. The evolution of the ISP activity in B. megaterium showed an opposite temperature dependence, being faster during growth at 42 degrees C. The amount of immunologically detected ISP1 again did not correlate well with the enzyme activity. Moreover, most of the ISP1 molecules in cell-free extracts from B. megaterium were inactive and were activated by increasing the CaCl2 concentration up to 30 mM--unlike B. subtilis, where the enzymic activity was unaffected by Ca2+ concentration. These data suggest that the ISP1 activity in the two bacillar species during postexponential growth is regulated posttranscriptionally, but that the regulatory mechanisms differ. PMID- 11270652 TI - Mechanism of the photocatalytic inactivation of Lactobacillus casei phage PL-1 by titania thin film. AB - The mechanism of the inactivation of Lactobacillus casei phage PL-1 suspended in a phosphate buffer by black-light (BL) -catalytic titanium dioxide (TiO2) thin film was studied. Generation of both superoxide anions (O2-) and hydroxyl radicals (*OH) was confirmed in the aqueous medium in which TiO2 film was settled with BL irradiation under gentle shaking. With BL-irradiation alone without TiO2 film, only O2- was generated to some extent. The genome DNA inside the phage particles was found to be fragmented by the treatment of PL-1 phages with BL catalytic TiO2 film. The phage inactivation by BL-catalytic TiO2 film was inhibited by the addition of albumin in a concentration-dependent manner. BL catalytic TiO2 film was considered to cause primarily the damage to the capsid protein through the generation of active oxygen species such as *OH, followed by damage to the genome DNA inside the phage particles. PMID- 11270653 TI - Comparative analysis between Pseudomonas aeruginosa genotypes and severity of symptoms in patients with unilateral or bilateral otitis externa. AB - Random amplified polymorphic DNA (RAPD) analysis was done on 32 isolates of Pseudomonas aeruginosa. These isolates were obtained from 22 patients who presented to the emergency room in a major medical center in Beirut, Lebanon, during a 5-month period with the diagnosis of either unilateral or bilateral otitis externa. Patients had yellowish to greenish discharge, moderate to severe external auditory canal swelling, moderate to severe pain, and periauricular cellulitis. None of these patients had intrinsic predisposing factors. An ear swab was obtained from both ears of patients, cultured on trypticase soy agar. P. aeruginosa was identified on the basis of pyocyanine production and API identification kits. RAPD analysis was done by using two primers (10 mer and 21 mer primers) and appropriate PCR conditions on extracted DNA. Our data have shown 23 RAPD patterns (A-W) distributed among the 32 P. aeruginosa isolates. RAPD patterns were reproducible. Twenty of 32 isolates were recovered from 10 patients with bilateral otitis externa. The remaining 12 of 32 isolates were recovered from 12 different patients with unilateral otitis externa. Eleven RAPD patterns (A,B,C,D,E,F,H,I,R,U,V) were associated with severe clinical symptoms, including severe pain, severe external auditory canal swelling, periauricular cellulitis, and a yellowish discharge. The remaining RAPD patterns were not associated with severe infections. This denotes a possible association between certain genotypes and severity of symptoms. PMID- 11270654 TI - Molecular markers for the characterization of Brazilian Cercospora caricis isolates. AB - Cercospora caricis is of interest as a potential mycoherbicide for control of purple nutsedge, Cyperus rotundus, which is considered to be the world's worst weed. The genetic variation of a collection of Brazilian Ce. caricis isolated from Cy. rotundus was analyzed by using RAPD, RFLP with a telomeric probe, [TTAGGG]18 and sequencing of the ITS1-5.8S-ITS2 regions of the ribosomal RNA gene. The Brazilian isolates were also compared with a Ce. caricis isolate from Florida, USA and with some other Cercospora species. A cluster of isolates from the Brazilian cerrado region was identified showing high genetic similarity. In contrast, isolates originating in other geographic regions of Brazil were less than 50% and 25% related to the former group according to similarity estimates produced from RAPD and telomeric RFLP analyses respectively. ITS sequence analysis did not support taxonomic division of the Brazilian strains, but did confirm the distant relatedness of these strains to the Ce. caricis isolate from Florida. The data indicate a need for an extensive molecular survey of Cercospora species associated with the Cyperaceae. PMID- 11270655 TI - Behavior of variable V3 region from 16S rDNA of lactic acid bacteria in denaturing gradient gel electrophoresis. AB - Separation of amplified V3 region from 16S rDNA by denaturing gradient gel electrophoresis (DGGE) was tested as a tool for differentiation of lactic acid bacteria commonly isolated from food. Variable V3 regions of 21 reference strains and 34 wild strains referred to species belonging to the genera Pediococcus, Enterococcus, Lactococcus, Lactobacillus, Leuconostoc, Weissella, and Streptococcus were analyzed. DGGE profiles obtained were species-specific for most of the cultures tested. Moreover, it was possible to group the remaining LAB reference strains according to the migration of their 16S V3 region in the denaturing gel. The results are discussed with reference to their potential in the analysis of LAB communities in food, besides shedding light on taxonomic aspects. PMID- 11270656 TI - Structure-activity relationships for six ketolide antibiotics. AB - Six structurally related 3-keto-substituted macrolide antibiotics (ketolides) were compared for concentration-dependent inhibitory effects on growth rate, viable cell number, and protein synthesis rates in Staphylococcus aureus cells. Inhibitory effects on 50S ribosomal subunit formation were also examined, as this is a second target for these antibiotics. A concentration range of 0.01 to 0.1 microg/ml was tested. An IC50 for inhibition of translation and 50S synthesis was measured for each compound, to relate structural features to inhibitory activity. ABT-773 was the most effective of the six compounds tested with an IC50 = 0.035 microg/ml. HMR 3004 was almost as effective with an IC50 = 0.05 microg/ml. Two 2 fluoroketolides (HMR 3562 and HMR 3787) were equivalent in their inhibitory activity with an IC50 = 0.06 microg/ml. Telithromycin (HMR 3647) had an IC50 = 0.08 microg/ml, and HMR 3832 was least effective with an IC50 = 0.11 microg/ml. Each antibiotic had an equivalent inhibitory effect on translation and 50S subunit formation. These results indicate specific structural features of these antimicrobial agents, which contribute to defined inhibitory activities against susceptible organisms. PMID- 11270657 TI - Purification, refolding of hybrid hIFNgamma-kringle 5 expressed in Escherichia coli. AB - The DNA sequence coding for plasminogen kringle 5 (pK5), an inhibitor of angiogenesis, was fused with that coding for interferon gamma and over-produced in the form of inactive inclusion bodies in E. coli. The amount of fusion protein was about 40% of total protein produced. The fusion protein contained in the inclusion bodies was solubilized in 8 M urea and purified by anion-exchange chromatography. We employed the orthogonal experimental design L16(4(5)) (5 factors, 4 levels, 16 experiments) procedure for researching the influence of denaturant, aggregation suppressor L-arginine, NaCl, pH, and glycine on the refolding procedure. Our results suggest that the presence of appropriate L arginine, NaCl, and denaturant in the refolding buffer inhibits the aggregation of the fusion protein and increases the yield of renatured protein with biological activity. The refolded fusion protein, gammaIFN/pk5, has in vitro anti endothelial cell proliferation activity. PMID- 11270658 TI - A unique pattern of mycelial elongation of Blakeslea trispora and its effect on morphological characteristics and beta-carotene synthesis. AB - The mycelial morphology of Blakeslea trispora was of crucial importance in the production of beta-carotene in submerged cultures of B. trispora. After the spores were inoculated, the time-course variation of mycelial morphology was closely examined under the microscope. With the addition of the non-ionic surfactant (Span 20: Sorbitan monolaurate, E493) to the culture medium, a unique pattern of mycelial elongation was observed: 1) slow formation of germ tubes from spores and 2) appearance of mycelia with very short length, which allowed a well dispersed growth of B. trispora without significant pellet aggregation. Span 20 appears to act like a paramorphogen. Without Span 20, however, the fungal culture finally formed a big clump of mycelium owing to heavy cross-linking of long mycelia. But the short mycelium maintained in the course of cultivation seemed to be irrelevant to growth inhibition, because the final concentration of dry mycelium was much higher with Span 20 after 3-day cultivation. The 20-fold increase in specific yield of beta-carotene (mg beta-carotene produced per g mycelium) was achieved with this drastic change in the pattern of mycelial elongation. The reason for this result might be more effective mass transfer and/or enhanced sensitivity to environmental oxidative stress in the well dispersed mycelial cultures of B. trispora. PMID- 11270659 TI - Impact of cellular metabolism on the biological effects of benzo[a]pyrene and related hydrocarbons. AB - Polycyclic aromatic hydrocarbons are ubiquitous contaminants in the environment. Benzo[a]pyrene (BaP), a prototypical member of this class of chemicals, has been extensively studied for its toxic effects in laboratory animals and human populations. BaP toxicity is often mediated by oxidative metabolism to reactive intermediates that interact with macromolecules leading to alterations in target cell structure and function. More recent evidence suggests that disruption of cellular signaling pathways involved in the regulation of growth and differentiation contribute significantly to the toxicity of BaP and its metabolites. This review summarizes recent advances in our understanding of biological mechanisms of BaP toxicity at the molecular level, and the role of metabolic intermediates in carcinogenesis, atherogenesis, and teratogenesis. PMID- 11270660 TI - Gap-junction communication in chemical carcinogenesis. PMID- 11270661 TI - Competitive inhibition of the transcription of rabbit CYP1A1 gene by upstream stimulatory factor 1 (USF1). AB - The induction of CYP1A1 by 3-methylcholanthrene occurs in neonatal but not in adult rabbits. The expression of aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (Arnt) mRNAs is seen even in adult rabbits. The CYP1A1 inducibility does not seem to be regulated by DNA methylation, known to inhibit the transcription of a gene(s). Preliminary experiments suggest that a constitutive factor(s) in adult liver nuclear extracts is bound to the core sequence of rabbit xenobiotic-responsive element (XRE). The sequence of rabbit XRE overlaps with that of the upstream stimulatory factor 1 (USF1)-binding site. The AhR/Arnt mediated activation of XRE-TK/Luc reporter gene in RK13 cells is blocked by transfection with a USF1 expression vector. These results indicate that the XRE of the rabbit CYP1A1 gene is recognized by the basic helix-loop-helix proteins to regulate the expression of CYP1A1 in both an agonistic (AhR/Arnt) and an antagonistic (USF1) manner. PMID- 11270662 TI - Mechanisms of 1,1-dichloroethylene-induced cytotoxicity in lung and liver. AB - Exposure to 1,1-dichloroethylene (DCE) causes lung and liver toxicities in mice. The lesions are characterized by damage preferentially to bronchiolar Clara cells in the lung and necrosis of centrilobular hepatocytes in the liver. The primary metabolites formed from DCE in lung and liver microsomal incubations are the epoxide, 2,2-dichloroacetaldehyde and 2-chloroacetyl chloride, which undergo hydrolysis and/or conjugation with glutathione (GSH). The major products formed are the epoxide-derived GSH conjugates 2-(S-glutathionyl) acetyl glutathione [B] and 2-S-glutathionyl acetate [C]. The hydrate of 2,2-dichloroacetaldehyde (acetal) is also detected. These metabolites are detected in vivo in murine lung and liver cytosol and in bile, and importantly, also in human lung and liver microsomal incubations. Formation of the epoxide is mediated mainly by CYP2E1. Immunohistochemical studies localized the epoxide-derived GSH conjugate [C] and cysteine-containing proteins in Clara cells and centrilobular hepatocytes. These findings are consistent with the premise that the lung and liver cytotoxicities induced by DCE are associated with in situ formation of the epoxide within the target cells. PMID- 11270663 TI - A life in science: biochemist-nutritionist-forensic toxicologist-pharmacologist. PMID- 11270664 TI - The sexual games of the body politic: fantasy and state violence in Northern Ireland. AB - This article analyzes the practice of strip searching women political prisoners in Northern Ireland as a violent technology of control aimed at breaking the political identity of prisoners. Focusing on a controversial case of a mass strip search carried out in 1992, the article examines the phantasmatic investements pervading this seemingly rational technology of control. Using a psychoanalytic notion of fantasy against the backdrop of a Foucaultian theory of power, this article argues that strip searches constitute a gendered form of political domination driven by, and performed within, a phantasmatic scenario of sexual violence. In this scenario both the political and gender identities of prisoners are re-inscribed with the power of a state acting as a male body politic. The article argues that the phantasmatic support of rational technologies of control betrays the contingent and shifting character of domination as well as its ambiguous effects. PMID- 11270665 TI - 'Eloquent chaos' in the oral discourse of killing fields survivors: an exploration of atrocity and narrativization. AB - If "narrative" implies a form of discourse in which sequenced events are meaningfully connected, an "anti-narrative" is a chaotic discourse form "of time without sequence, telling without mediation, and speaking about oneself without being fully able to reflect on oneself" (Frank 1995: 98). This paper examines narratives and anti-narratives in the oral discourses of survivors of the Cambodian killing fields. Through an extended analysis of two cases, we demonstrate the internal logic and "eloquence" of anti-narratives--i.e., the ways in which anti-narrative patterns vividly express and reveal a survivor's complex and continuing experience of atrocity. PMID- 11270666 TI - Applying Aristotle's doctrine of causation to Aboriginal and biomedical understanding of diabetes. AB - Aristotle's doctrine on causation identifies four distinct types of cause: formal, efficient, material, and final. Science is said to have differentiated itself from philosophy by concentrating solely on efficient causes. Nonetheless, when applied to narratives of causation, Aristotle's doctrine provides a useful heuristic to explore the issues such as Aboriginal and biomedical perceptions of causal factors for non-insulin dependent diabetes mellitus (NIDDM) on Manitoulin Island, Ontario. This paper also outlines two divergent causal stories for NIDDM and the associated moral positions regarding the 'righteous' pursuit of health. Biomedical narratives emphasize the role of lifestyle factors, particularly the impact of obesity, in causation. In the case of diabetes, the moral course of action is pursued through lifestyle choices. In contrast, Aboriginal narratives emphasize the role of genetics in causation. These narratives describe diabetes as collectively affecting Aboriginal people - thus identifying Aboriginal people as different. Aboriginal frameworks for health venture beyond the 'efficient' cause of biomedicine and thus the moral pursuit of health within this framework involves returning to an initial state of health and purity through traditional knowledge. PMID- 11270667 TI - Technology and effect: HIV/AIDS testing in Brazil. AB - Contemporary techno-scientific and medical developments are restructuring social interactions and the very processes by which individual subjectivity is formed. This essay elaborates on the experiential and ethical impact of such transformations from the perspective of people who, in ordinary and unexpected ways, act science and technology out. We carried out ethnographic research in an HIV/AIDS Testing and Counseling Center (CTA) in northeastern Brazil, combining participant observation with epidemiological analyses and clinical survey. We found a high demand for free testing by low-risk clients, largely working and middle class, experiencing anxiety and complaining of AIDS-like symptoms. Most of the clients were sero-negative and many returned for a second and third testing. We understand this to be a new techno-cultural phenomenon and call it imaginary AIDS. Throughout this essay, we describe CTA's routine practices, place these practices in historical, political, economic and cross-cultural perspective, and analyze the subjective data we collected from the clients of our pilot study. We explore how clinical epidemiological expertise and HIV testing technology are integrated into new forms of bio-politics aimed at specific marketable and disease-free populations, and on the affective absorption of bio-technical truth and the engendering of a technoneurosis in this testing center. PMID- 11270668 TI - Advanced glycation end-products: a review. AB - Advanced glycation end-products are a complex and heterogeneous group of compounds that have been implicated in diabetes related complications. At present it is not known if they are the cause or the consequence of the complications observed. We discuss the chemistry of advanced glycated end-product formation and their patho-biochemistry particularly in relation to the diabetic microvascular complications of retinopathy, neuropathy and nephropathy as well as their role in the accelerated vasculopathy observed in diabetes. The concept of carbonyl stress as a cause for advanced glycated end-product toxicity is mentioned. We discuss alterations in the concentrations of advanced glycated end-products in the body, particularly in relation to changes occurring with age, diabetes and its complications such as nephropathy. Problems relating to current methods of advanced glycated end-product detection and measurement are highlighted including the lack of a universally established method of detection or unit of measurement. Agents used for the treatment of advanced glycated end-product accumulation are reviewed, with an emphasis on the results of the recent phase III trials using aminoguanidine and diabetes related complications. PMID- 11270669 TI - Incidence of blindness in southern Germany between 1990 and 1998. AB - AIMS/HYPOTHESIS: A reduction of diabetes-related blindness by at least one third was declared a primary objective for Europe in 1989 (St. Vincent Declaration). To ascertain a potential change of incidence rates, we collected data on blindness in a German district (population: about 5 million) over 9 years. METHODS: We obtained complete lists of newly registered blindness-allowance recipients between 1990 and 1998 and population data on Wurttemberg-Hohenzollern, Germany. We estimated incidence rates of blindness in the general population and the diabetic population. To ascertain any time trend, we applied Poisson regression models. RESULTS: There were 6371 newly registered blindness allowance recipients (1990-1998). Of these 67% were women and 27 % had diabetes. Mean age was 71.7 years. Standardised results in the diabetic population (incidence rates per 100,000 person-years; standard: diabetic population; 95 % CI): 1990: 72 (61;82); 1991: 88 (76;100); 1992: 77 (67;88); 1993: 82 (71;93); 1994: 62 (53;72); 1995: 82 (71;93); 1996: 70 (60;80); 1997: 69 (59;79); 1998: 59 (49;68). The Poisson model estimated a 3 % decrease of incident blindness in the diabetic population for each year (Relative risk per year 0.97; CI: 0.95; 0.99). No significant change could be observed in the non-diabetic population (Relative risk: 0.99; CI: 0.98; 1.00). Relative risks for each year varied between sub-groups according to sex, diabetic status and cause of blindness between 0.94 and 1.01. CONCLUSION/INTERPRETATION: A slight reduction of incident blindness could be shown but a reduction by one third has not been reached. Several possible sources of bias in the data have to be considered. PMID- 11270670 TI - Undiagnosed coeliac disease and risk of autoimmune disorders in subjects with Type I diabetes mellitus. AB - AIMS/HYPOTHESIS: We tested the hypothesis that silent coeliac disease is more frequent than expected in both patients with Type I (insulin-dependent) diabetes mellitus and their first-degree relatives. We evaluated how the presence of other autoimmune disorders in diabetic patients and their first-degree relatives is related to silent, unrecognized coeliac disease. METHODS: Sera from 491 subjects with Type I diabetes, 824 relatives and 4,000 healthy control subjects were screened for anti-endomysial antibodies and all those subjects who tested positive for anti-endomysial antibodies underwent intestinal biopsy. RESULTS: We found that the prevalence of coeliac disease was 5.7 % among the diabetic patients and 1.9 % among the relatives, values significantly higher than those found among the control subjects (p < 0.0001; p < 0.001). The prevalence of autoimmune disorders in diabetic patients with coeliac disease was significantly higher than in subjects with Type I diabetes alone (p < 0.0001). The prevalence of autoimmune disorders in the relatives with coeliac disease was significantly higher than in those who tested negative for anti-endomysial antibodies (p = 0.01). CONCLUSION/INTERPRETATION: This report provides further confirmation of the high prevalence of undiagnosed coeliac disease among diabetic patients and their relatives. This interesting new finding is the increased presence of other autoimmune diseases in these patients, as well as in their relatives with a delayed diagnosis for coeliac disease. Patients newly diagnosed with coeliac disease showed excellent compliance with the gluten-free diet. This should encourage policymakers to consider introducing an easy-to-use screening programme for diabetic patients and their relatives into everyday clinical practice, in order to prevent coeliac-associated symptoms and the onset of additional, more serious auto-immune disorders. PMID- 11270671 TI - UKPDS 50: risk factors for incidence and progression of retinopathy in Type II diabetes over 6 years from diagnosis. AB - AIMS/HYPOTHESIS: To determine risk factors related to the incidence and progression of diabetic retinopathy over 6 years from diagnosis of Type II (non insulin-dependent) diabetes mellitus. METHODS: This report describes 1919 patients from within the United Kingdom Prospective Diabetes Study (UKPDS), with retinal photographs taken at diagnosis and 6 years later and with complete data available. Photographs were centrally graded for lesions of diabetic retinopathy using the modified Early Treatment of Diabetic Retinopathy Study Final scale. Risk factors were assessed after 3 months diet from the time of diagnosis of diabetes. Patients were seen every 3 months in a hospital setting. Biochemical measurements were done by a central laboratory. End points of vitreous haemorrhage and photocagulation were confirmed by independent adjudication of systematically collected clinical data. The main outcome measures were incidence and progression of retinopathy defined as a two-step Early Treatment of Diabetic Retinopathy Study (ETDRS) final scale change. RESULTS: Of the 1919 patients, 1216 (63 %) had no retinopathy at diagnosis. By 6 years, 22 % of these had developed retinopathy, that is microaneurysms in both eyes or worse. In the 703 (37 %) patients with retinopathy at diagnosis, 29 % progressed by two scale steps or more. Development of retinopathy (incidence) was strongly associated with baseline glycaemia, glycaemic exposure over 6 years, higher blood pressure and with not smoking. In those who already had retinopathy, progression was associated with older age, male sex, hyperglycaemia (as evidenced by a higher HbA1c) and with not smoking. CONCLUSION/INTERPRETATION: The findings re-emphasise the need for good glycaemic control and assiduous treatment of hypertension if diabetic retinopathy is to be minimised. PMID- 11270672 TI - Increased plasma leptin in gestational diabetes. AB - AIMS/HYPOTHESIS: Insulin resistance as well as marked changes in body weight and energy metabolism are associated with pregnancy. Its impact on plasma leptin is not known and was determined in this longitudinal study in both diabetic and normal pregnancy. METHODS: At 28 gestational weeks plasma concentrations of leptin and B-cell hormones were measured at fasting and after an oral glucose load (OGTT:75 g) in women with gestational diabetes and pregnant women with normal glucose tolerance and compared with women who were not pregnant (C). RESULTS: Plasma leptin (ng/ml) was higher (p < 0.001) in women with gestational diabetes (24.9 +/- 1.6) than in women with normal glucose tolerance (18.2 +/- 1.5) and increased in both groups when compared with the non-pregnant women (8.2 +/- 1.3; p < 0.0005). No change in plasma leptin concentrations was induced by OGTT in any group. Basal insulin release was higher (p < 0.05) in women with gestational diabetes compared with the pregnant women with normal glucose tolerance. Marked insulin resistance was confirmed by a 20 % lower (p < 0.05) insulin sensitivity in subgroup analysis and a decrease of almost 40% in fasting glucose/insulin ratio (p < 0.005) in women with gestational diabetes. Leptin correlated in women with gestational diabetes with basal plasma concentrations of glucose (p < 0.02), insulin (p < 0.004) and proinsulin (p < 0.01) as well as with BMI (p < 0.001) and overall pregnancy induced maternal weight gain (p < 0.009). With normalisation of blood glucose 8 weeks after delivery in women with gestational diabetes their plasma leptin decreased (p < 0.0005) to 17.3 +/- 1.9 ng/ml but did not completely normalize (p < 0.05 vs non-pregnant women). CONCLUSION/INTERPRETATION: Our data show that women with gestational diabetes without any change in plasma leptin upon oral glucose loading have increased plasma leptin concentrations during and after pregnancy, a clear association of plasma leptin with the respective concentration of glucose and insulin resistance as well as with changes in body weight, and a failure to normalize spontaneously BMI to the same extent as pregnant women with normal glucose tolerance when compared with matched control subjects. PMID- 11270673 TI - Ceramide impairs the insulin-dependent membrane recruitment of protein kinase B leading to a loss in downstream signalling in L6 skeletal muscle cells. AB - AIMS/HYPOTHESIS: Increased cellular production of ceramide has been implicated in the pathogenesis of insulin resistance and in the impaired utilisation of glucose. In this study we have used L6 muscle cells to investigate the mechanism by which the short-chain ceramide analogue, C2-ceramide, promotes a loss in insulin sensitivity leading to a reduction in insulin stimulated glucose transport and glycogen synthesis. METHOD: L6 muscle cells were pre-incubated with C2-ceramide and the effects of insulin on glucose transport, glycogen synthesis and the activities of key molecules involved in proximal insulin signalling determined. RESULTS: Incubation of L6 muscle cells with ceramide (100 micromol/l) for 2 h led to a complete loss of insulin-stimulated glucose transport and glycogen synthesis. This inhibition was not due to impaired insulin receptor substrate 1 phosphorylation or a loss in phosphoinositide 3-kinase activation but was caused by a failure to activate protein kinase B. This defect could not be attributed to inhibition of 3-phosphoinositide-dependent kinase-1, or to impaired binding of phosphatidylinositol 3,4,5 triphosphate (PtdIns(3,4,5)P3) to the PH domain of protein kinase B, but results from the inability to recruit protein kinase B to the plasma membrane. Expression of a membrane-targetted protein kinase B led to its constitutive activation and an increase in glucose transport that was not inhibited by ceramide. CONCLUSIONS/INTERPRETATION: These findings suggest that a defect in protein kinase B recruitment underpins the ceramide induced loss in insulin sensitivity of key cell responses such as glucose transport and glycogen synthesis in L6 cells. They also suggest that a stimulated rise in PtdIns(3,4,5)P3 is necessary but not sufficient for protein kinase B activation in this system. PMID- 11270674 TI - Myocardial fatty acid oxidation in patients with impaired glucose tolerance. AB - AIMS/HYPOTHESIS: Fatty acids are an important source of energy in the myocardium. Abnormal myocardial fatty acid metabolism could contribute to the deterioration of cardiac function frequently observed in patients with Type II (non-insulin dependent) diabetes mellitus. In our previous study, myocardial total uptake of non-esterified fatty acid (NEFA) was measured in patients with impaired glucose tolerance and found to be normal. This study aimed to investigate the subsequent metabolic steps and beta-oxidation of NEFA. METHODS: A total of 6 men with impaired fasting glucose (age 50 +/- 2 years, BMI 29 +/- 1 kg/m2, means +/- SEM) and 6 healthy men (50 +/- 1 years, 25 +/- 1 kg/ m2) were studied in the fasting state. Myocardial blood flow was measured with [15O]H2O and positron emission tomography and myocardial NEFA metabolism with [11C]palmitic acid. RESULTS: Myocardial blood flow was normal and not different between the impaired glucose tolerance and the control group (78 +/- 6 vs 73 +/- 13 ml/100 g/ min, NS). The [11C]palmitic acid uptake indices were similar between the groups (10.4 +/- 0.5 vs 11.2 +/- 0.8 ml/100 g/min, respectively, NS). The clearance of [11C]-palmitate from the myocardium, an index of NEFA beta-oxidation, was similar between the groups (half-times of activity 17.6 +/- 1.6 vs 19.5 +/- 2.3 min, respectively, NS) CONCLUSION/INTERPRETATION: The results indicate that myocardial NEFA uptake and beta-oxidation are not altered in patients with IGT. Thus, it is not likely that altered NEFA metabolism contributes to the deterioration of the cardiac function in patients with IGT or Type II diabetes. PMID- 11270675 TI - Regulation of protein kinase C by short term hyperglycaemia in human platelets in vivo and in vitro. AB - AIMS/HYPOTHESIS: Postprandial hyperglycaemia carries an increased risk of macrovascular disease even without Type II (non-insulin-dependent) diabetes mellitus. Chronic hyperglycaemia activates protein kinase C (PKC) in vitro and in vivo but it is not known whether PKC is regulated by short-term post-prandial hyperglycaemia in vivo in humans. We investigated whether PKC is regulated in vivo in hyperglycaemic and hyperinsulinaemic infusion tests and correlated the results to stimulations in vitro. METHODS: Protein kinase C regulation was measured in platelets obtained from 8 healthy subjects who were infused with glucose and insulin for 2 h attaining peak concentrations of 16 mmol/l glucose and in platelets from 8 healthy young subjects, 8 older subjects without diabetes, and 10 older subjects with Type II diabetes after incubation in vitro with 16 mmol/l glucose or glucose and insulin. For precise quantification, a shortened PKC beta1 standard protein was generated by bacterial expression and PKC alpha, beta1, beta2 and delta isoenzyme values were measured by immunoblot analyses. RESULTS: Hyperglycaemic and hyperinsulinaemic in vivo tests increased the amounts of PKC alpha, beta1 and beta2 in the membrane fraction of platelets to 225 +/- 87 %, 164 +/- 22 % and 302 +/- 135 %, respectively, when compared with the baseline values in young healthy volunteers (n = 8, p < 0.05). The expression of PKC delta did not change. In comparison to the recombinant PKC beta1 standard protein, 5 ng PKC beta1/ microg protein was measured before the test and 2 ng/microg were translocated to the membrane fraction after the infusion. No change in the absolute amount of PKC beta1 was detected. In contrast, after incubation in vitro PKC was not regulated by glucose or glucose and insulin in 8 young healthy subjects (age 26 +/- 0.7 years) and in 8 older, healthy subjects (age 64,8 +/- 4 years) although 100 nmol/l 12-O-tetradecanoylphorbol 13-acetate caused maximal activation. In marked contrast, PKC beta1 and PKC beta2, but not PKC alpha or PKC delta, were increased in vitro in the membrane fraction by 292 +/- 61% and 432 +/- 88% (p < 0.05) in 10 subjects with Type II diabetes mellitus matched for age, sex and BMI. CONCLUSION/INTERPRETATION: We found that short-term hyperglycaemia activates PKC alpha, beta1 and beta2 in platelets of healthy persons making them potential candidates for mediating the increased cardiovascular risk of postprandial hyperglycaemia. Hyperglycaemia and hyperinsulinaemia did not cause short-term activation of PKC in platelets in vitro suggesting the existence of additional stimuli. Subjects with Type II diabetes showed a markedly altered reactivity of platelet PKC beta in vitro indicating some diabetes-related regulation. PMID- 11270676 TI - Presence of glutamine:fructose-6-phosphate amidotransferase for glucosamine-6 phosphate synthesis in endothelial cells: effects of hyperglycaemia and glutamine. AB - AIMS/HYPOTHESIS: Recent studies show that glucosamine infusion impairs endothelium-dependent blood flow in normoglycaemic rats. The pathophysiological relevance of this finding, however, depends on whether de novo glucosamine synthesis occurs in endothelial cells. The aim of this study was to test the hypothesis of whether glutamine:fructose-6-phosphate amidotransferase (the first and key regulatory enzyme in hexosamine synthesis) is present for endothelial glucosamine synthesis. METHODS: Bovine venular, bovine aortic, human microvascular, human umbilical vein, and rat coronary microvascular endothelial cells were used to measure glutamine:fructose-6-phosphate amidotransferase activity. To determine glucosamine-6-phosphate synthesis in intact cells, they were incubated for 1 h in Krebs bicarbonate buffer containing 5, 15 or 30 mmol/l [U-14C]glucose and 0.5, 2 or 4 mmol/l glutamine. The [14C]Glucosamine-6-phosphate and its end products ([14C]UDP-N-acetylglucosamine and [14C]UDP Nacetylgalactosamine) were separated by HPLC. RESULTS: There were high glutamine:fructose-6-phosphate amidotransferase activities in all endothelial cells studied. Exposure of cells to 15 to 30 mmol/l glucose or 2 to 4 mmol/l glutamine increased enzyme activity. Glucosamine-6-phosphate, UDP-N acetylglucosamine and UDP-N-acetylgalactosamine syntheses increased with increasing extracellular concentrations of glucose from 5 to 30 mmol/l or of glutamine from 0.5 to 4 mmol/l. CONCLUSION/INTERPRETATION: Our results show the presence of glutamine:fructose-6-phosphate amidotransferase for de novo glucosamine synthesis in endothelial cells and the modulation of this pathway by hyperglycaemia and glutamine. As glucosamine inhibits endothelial nitric oxide synthesis, these findings could have important implications for impaired endothelium-dependent relaxation and vascular dysfunction in diabetes mellitus. PMID- 11270677 TI - Progression of large artery structural and functional alterations in Type I diabetes. AB - AIMS/HYPOTHESIS: Type I (insulin-dependent) diabetes mellitus is accompanied by reduced arterial distensibility and increased arterial wall thickness even in normotensive subjects with no micro-macrovascular complications. It is not known whether, and how fast, these subclinical markers of vascular damage develop over time. METHODS: We measured arterial wall distensibility in radial, common carotid artery and abdominal aorta in 60 normotensive patients (aged 35.0 +/- 1.2 years, means +/- SE) with Type I diabetes with no microvascular or macrovascular complications and in 20 healthy control subjects matched for age. Arterial distensibility was determined by continuous measurements of arterial diameter through echotracking techniques and by using either the Langewouters (radial artery) or the Reneman formula (carotid artery and aorta). The same echotracking techniques allowed us to ascertain the radial and carotid artery wall thickness. Data were collected before and after 23 +/- 1 months. RESULTS: In the first study, carotid artery distensibility was similar but radial artey and aortic distensibility was less (p < 0.01) in patients with diabetes than in control subjects (-39 % and 25 % respectively). This was accompanied by an increase (p < 0.01) in both radial (42 %) and carotid artery wall thickness (46 %). After 23 +/ 1 months diabetic subjects showed a further reduction in arterial distensibility (radial-12 %, p < 0.05; carotid-8 %, NS; aorta-20% p < 0.05) and an increase in arterial wall thickness (radial + 15 %; carotid 14%, p < 0,05). No change in distensibility and wall thickness values occurred in control subjects. CONCLUSION/INTERPRETATION: The early reduction in arterial distensibility and increase in arterial wall thickness characterizing uncomplicated normotensive Type I diabetes patients shows a measurable worsening over the short term. PMID- 11270678 TI - Hyperglycaemia in vitro alters the proliferation and mitochondrial activity of the choriocarcinoma cell lines BeWo, JAR and JEG-3 as models for human first trimester trophoblast. AB - AIMS/HYPOTHESIS: Early intrauterine growth delay in diabetes could be caused by a reduced growth of the placenta. Our study investigates whether hyperglycaemia in vitro reduces trophoblast proliferation. METHODS: First-trimester trophoblast cell models (BeWo, JAR and JEG-3 choriocarcinoma cells) were cultured for 24 and 48 h with 5.5 mmol/l D-glucose, 25 mmol/1 D-glucose (hyperglycaemia) and with an osmotic control. Cell number, total protein and nucleic acid content and mitochondrial activity (tetrazolium salt assay) were measured, the cell cycle analysed (FACS, cyclin B1 levels) and apoptosis (Annexin-V) measured. RESULTS: In BeWo cells hyperglycaemia reduced cell number, protein, nucleic acid and cyclin B1 levels. The reduced G2/M and increased G0/G1 population after 24 h reflects growth arrest at G0/G1. In JAR cells after 24 h the population was arrested in G0/G1, whereas after 48 h the G0/G1 block was abrogated and the cells were arrested at G2/M. The net effect was an unchanged cell number. In JEG-3 cells hyperglycaemia resulted in fewer cells after 24 h but not after 48 h indicating some adaptation. Mitochondrial activity was either stimulated (BeWo) or reduced (JAR, JEG-3) under hyperglycaemia. Some of these effects were also induced by hyperosmolarity alone. CONCLUSION/INTERPRETATION: Hyperglycaemia has the potential to inhibit the proliferation of first-trimester trophoblast cell models. The mechanisms leading to growth arrest and to changes in mitochondrial activity are complex and depend on differentiation. We hypothesise a hyperglycaemia-induced impairment of placental growth in the first trimester of a poorly controlled diabetic pregnancy. PMID- 11270679 TI - Streptozotocin-induced diabetes alters the oligomerization pattern of acetylcholinesterase in rat skeletal muscle. AB - AIMS/HYPOTHESIS: Diabetes mellitus has a serious effect on most of the properties of skeletal muscles. Changes in neuromuscular transmission are also involved in propagating the disease. METHODS: In our experiments, acetylcholinesterase was extracted from the fast extensor digitorum longus and slow soleus muscles of control, non-treated 6-week-diabetic and insulin-treated diabetic rats. The extracts were applied to velocity sedimentation and acetylcholinesterase activity was determined. RESULTS: We observed considerable differences in the distribution of individual acetylcholinesterase molecular forms in diabetic fast muscles. This included a 59% decline in G4 content together with a fivefold increase in A8 and a 53 % increase in A12 activity resulting in a shift of acetylcholinesterase profile characteristically towards slow muscles. These alterations were partly reversed by insulin treatment. CONCLUSION/INTERPRETATION: In slow muscles diabetes caused an increase in G4 activity without affecting the sedimentation profile. Decline in G4 content in fast muscles could contribute to enhanced desensitization of acetylcholine receptors in diabetes. PMID- 11270680 TI - Markers of renal tubular dysfunction measured annually do not predict risk of microalbuminuria in the first few years after diagnosis of Type I diabetes. AB - AIMS/HYPOTHESIS: Early detection of risk of microalbuminuria could prevent early renal damage. We investigated whether urine retinol binding protein and N-acetyl glucosaminidase could predict the risk of microalbuminuria in a large cohort of children followed from diagnosis of Type I (insulin-dependent) diabetes mellitus. METHODS: Subjects under 16 years of age within a georaphically defined region were recruited at diagnosis of Type I (insulin-dependent) diabetes mellitus. Annually, albumin-, retinol binding protein- and N-acetyl-glucosaminidase- to creatinine ratios were each measured in 3 urine samples. RESULTS: A total of 511 subjects were followed for a median of 6 years (range: 1-14). Microalbuminuria (males: > or = 3.5 mg/mmol; females: > or = 4.0 mg/mmol, in 2 out of 3 urines) developed in 78 subjects (36 male). The cumulative probability of microalbuminuria was 40% after 12 years duration of diabetes. Retinol-binding proteinuria (men: > or = 21 microg/mmol; women > or = 33 microg/mmol) developed in 217 subjects (152 men). The cumulative probability of retinol-binding proteinuria was 67 % after 12 years duration of diabetes. The cumulative probability of retinol-binding-proteinuria was 40 % before the onset of microalbuminuria and 59% in subjects who did not subsequently develop microalbuminuria. Retinol-binding-proteinuria developed at a higher rate with increasing HbA1c than microalbuminuria. N-acetyl-glucosaminidase-uria (males: > or = 56 micromol-pnp x h(-1) x mmol(-1); females: > or = 46 micromol-pnp h(-1) x mmol(-1)) developed in 477 subjects. The cumulative probability of N acetylglucosaminidase-uria was 98 % after 10 years of diabetes duration. The cumulative probability of N-acetyl-glucosaminidase-uria was 73 % in the years before the onset of microalbuminuria and 97 % in subjects without microalbuminuria. The probability of Nacetyl-glucosaminidase-uria was 99 % with an HbA1c greater than or equal to 14.5 %. CONCLUSIONS/INTERPRETATION: Raised amounts of urine retinol binding protein and N-acetyl-glycosaminidase are related to HbA1c and the duration of diabetes. They occur in the majority of subjects and are not early markers for the risk of microalbuminuria. PMID- 11270681 TI - Ramipril and aminoguanidine restore renal lysosomal processing in streptozotocin diabetic rats. AB - AIMS/HYPOTHESIS: We aimed to examine the time course for the diabetes-related changes in renal lysosomal processing and to determine whether these changes can be prevented by aminoguanidine or ramipril treatment. METHODS: The percentage desulphation of intravenously injected tritium labelled dextran sulphate ([3H]DSO4) in the urine, as determined by ion-exchange chromatography, was used as a marker of lysosomal sulphatase activity. Sulphatase activity was determined 1, 2, 3 and 4 weeks after the onset of diabetes in rats as well as in rats treated with either aminoguanidine or ramipril for twelve weeks. RESULTS: The amount of totally desulphated [3H]DSO4 in urine collected from control rats was 65.6 +/- 0.8%. This was significantly reduced in diabetic rats two (57.4 +/- 1.4% desulphated), three (56.8 +/- 1.3 % desulphated) and four (52.9 +/- 2.2% desulphated) weeks after the onset of diabetes. The significant decrease in the amount of totally desulphated [3H]DSO4 in the urine also found at 12 weeks after the onset of diabetes was not affected by drug treatment. There was no significant difference in the amount of partially desulphated [3H]DSO4 in the urine between all the study groups. However, the increase in totally sulphated [3H]DSO4 in the urine collected from diabetic rats (8.7 +/- 1.7 % sulphated) compared with that of control rats (2.2 +/- 0.5% sulphated) was normalised by treatment with both aminoguanidine (4.8 +/- 1.6% sulphated) or ramipril (4.5 +/- 0.8% sulphated). CONCLUSIONS/INTERPRETATION: These results raise the possibility that the diabetes-induced changes in renal lysosomal processing may be one of the initial events in the development of diabetic nephropathy. Aminoguanidine and ramipril, known for their different mechanism of action, seem to prevent diabetes induced changes in lysosomal processing either through their effects on enzyme activity within the lysosome or through their effects on the trafficking of molecules to and from the lysosome. PMID- 11270682 TI - Studies of variability in the PTEN gene among Danish caucasian patients with Type II diabetes mellitus. AB - AIM/HYPOTHESIS: Phosphatase and tensin homologue deleted from chromosome ten (PTEN) has recently been characterized as a novel member in the expanding network of proteins regulating the intracellular effects of insulin. By dephosphorylation of phosphatidyl-inositol-(3, 4, 5)-trisphosphate (PIP3) the PTEN protein regulates the insulin-dependent phosphoinositide 3-kinase (PI3K) signalling cassette and accordingly might function as a regulator of insulin sensitivity in skeletal muscle and adipose tissue. In this study we tested PTEN as a candidate gene for insulin resistance and late-onset Type II (non-insulin-dependent) diabetes mellitus in a Danish Caucasian population. METHODS: The nine exons of the PTEN, including intronic flanking regions were analysed by PCR-SSCP and heteroduplex analysis in 62 patients with insulin-resistant Type II diabetes. RESULTS: No mutations predicted to influence the expression or biological function of the PTEN protein but four intronic polymorphisms were identified: IVS1-96 A-->G (allelic frequency 0.22, 95 % CI: 0.12-0.32), IVS3 + 99 C-->T (0.01, CI: 0-0.03), IVS7-3 TT-->T (0.10, CI: 0.03-0.18) and IVS8 + 32 G-->T (0.35, CI: 0.23-0.47). The IVS8 + 32 G-->T polymorphism was used as a bi-allelic marker for the PTEN locus and examined in 379 patients with Type II diabetes and in 224 control subjects with normal glucose tolerance. The IVS8 + 32 G-->T polymorphism in the PTEN was not associated with Type II diabetes and it did not have any effect on body-mass index, blood pressure, HOMA insulin resistance index, or concentrations of plasma glucose, serum insulin or serum C peptide obtained during an oral glucose tolerance test (OGTT). CONCLUSION/INTERPRETATION: Variability in the PTEN is not a common cause of Type II diabetes in the Danish Caucasian population. PMID- 11270683 TI - Variants of neurogenin 3 gene are not associated with Type II diabetes in Japanese subjects. AB - AIMS/HYPOTHESIS: Neurogenin 3 (ngn3) is a transcription factor expressed in the endocrine precursor cells of the pancreas. It has recently been reported that ngn3-deficient mice show absence of pancreatic endocrine cells and die of postnatal diabetes. The purpose of this investigation was to screen for polymorphisms of the ngn3 gene and to test whether these polymorphisms are associated with Type II (non-insulin-dependent) diabetes mellitus in the Japanese subjects. METHODS: We screened ngn3 gene and upstream region by direct sequencing and estimated the prevalence of polymorphisms in 197 patients with Type II (non insulin-dependent) diabetes mellitus and 216 control subjects. RESULTS: We identified four novel polymorphisms, Ser199Phe (596C/T), -43insCA, -983C/T and 1822G/A. In an association study the allelic frequencies of the major allele of these four polymorphisms were 0.721, 0.914, 0.912 and 0.530 in diabetic patients, respectively, and 0.694, 0.905, 0.917 and 0.537 in control subjects, respectively. CONCLUSION/INTERPRETATION: Mutations and polymorphisms of ngn3 gene are not significantly associated with Type II (non-insulin-dependent) diabetes mellitus in the Japanese subjects. PMID- 11270684 TI - Variable effects of the APOC3-482C > T variant on insulin, glucose and triglyceride concentrations in different ethnic groups. AB - AIMS/HYPOTHESIS: The apolipoprotein C3-482C> T variant modulates insulin and glucose concentrations after an oral glucose tolerance test (OGTT) in young healthy white men. We evaluated the effect of this variant in different ethnic groups with different rates of Type II (non-insulin-dependent) diabetes mellitus and coronary heart disease. METHODS: We investigated the -482C > T in a population-based cross-sectional study of white subjects (n = 462), South Asians (n = 442) and subjects of West African and Afro-Caribbean origin (n = 462), whose OGTT and fasting plasma triglyceride concentrations had been measured. RESULTS: The -482T allele frequency differed between the three groups: 0.25 (95 % CI 0.22 0.28) in white subjects, 0.44 (0.41-0.47) in South Asians and 0.71 (0.68-0.74) in black subjects (p < 0.0001). A positive association was found between body mass index and genotype in black women (p = 0.009) and in black men (p = 0.056) but not in white subjects or South Asians. Associations between -482C > T and fasting insulin were found in white subjects, where -482T allele carriers had higher concentrations (adjusted for age and sex, p = 0.007, also including smoking and body mass index, p = 0.038). Higher triglyceride concentrations (p = 0.004 and p = 0.007 in the two models) but not glucose concentrations were also associated with -482C > T. In black subjects, decreased fasting insulin (p = 0.04) and fasting glucose (p = 0.004) were associated with -482C > T. No relation was observed between genotype and any post-load measured. CONCLUSIONS/INTERPRETATION. Allele frequencies of the -482C > T and associations with insulin, glucose and triglyceride concentrations vary considerably among ethnic groups. Although the results are consistent among white subjects across different studies, the associations among black subjects and South Asians differ. PMID- 11270686 TI - Silent coronary artery disease in diabetes--a feature of autonomic neuropathy or accelerated atherosclerosis? AB - Asymptomatic coronary artery disease and myocardial infarctions are common in diabetic subjects. The available clinical and epidemiological data suggest that the increased incidence of asymptomatic myocardial infarctions and coronary artery disease in diabetic patients mainly reflects accelerated coronary atherosclerosis and the proportion of silent disease relative to symptomatic disease or episodes is not increased in diabetes. In spite of the theoretical background, there is no convincing clinical or epidemiological evidence that diabetic autonomic neuropathy plays a major part in the lack of ischaemic pain. This is not surprising because the mechanisms of silent myocardial ischaemia are complex and controversial even without diabetes. PMID- 11270685 TI - Functional consequences of mutations in the MODY4 gene (IPF1) and coexistence with MODY3 mutations. AB - AIMS/HYPOTHESIS: The aim of this study was to examine the putative role of mutations in the insulin promoter 1 (IPF1) gene in early-onset diabetes. METHODS: We carried out mutation screening of the IPF1 gene in 115 Scandinavian families with at least two members with onset of diabetes younger than 40 years. The allele frequencies were also tested in 183 unrelated patients with late-onset Type II (non-insulin-dependent) diabetes mellitus and in 92 non-diabetic control subjects. RESULTS: Two novel IPF1 variants (G212R and P239Q) and one previously reported (D76N) IPF1 variant were identified in the 115 families (3.5%). The D76N variant was found in one MODY3 family (S315fsinsA of HNF1alpha) and also in two families with late-onset Type II diabetes. The P239Q variant was identified in two families with early-onset diabetes including one with MODY3 (R272C of HNF1alpha) and in three families with late-onset Type II diabetes. Despite the fact that the variants did not segregate completely with diabetes, the non diabetic carriers of the IPF1 variants had increased blood glucose concentrations (p < 0.05) and reduced insulin:glucose ratios (p < 0.05) during an oral glucose tolerance test compared with non-diabetic family members without these variants. In addition, when the G212R and P239Q variants were expressed in cells without IPF1 i.e.. Nes2y cells, both variants showed about a 50% reduction in their ability to activate insulin gene transcription compared to wild-type IPF1, as measured by reporter gene assay. CONCLUSION/INTERPRETATION: Although mutations in the IPF-1 gene are rare in early- (3.5 %) and late-onset (2.7 % ) Type II diabetes, they are functionally important and occur also in families with other MODY mutations. PMID- 11270687 TI - ICA12(SOX13) autoantibodies are unlikely to be a useful marker for pre-clinical Type I diabetes. PMID- 11270689 TI - Insulin fails to modulate the cardiac L-type Ca2+ current in Type II diabetes patients--a possible link to cardiac dysfunction in diabetes mellitus. PMID- 11270688 TI - Continuous glucose monitoring during the European Soccer cup semifinal, Italy against Holland. PMID- 11270690 TI - The Russ Prize. PMID- 11270691 TI - The Chicago School of Arrhythmology: an analysis of a revisionist view. PMID- 11270692 TI - The Chicago School of Electrocardiography and second-degree atrioventricular block: an historical perspective. PMID- 11270693 TI - Mapping of atrial activation patterns after inducing contiguous radiofrequency lesions: an experimental study. AB - High resolution mapping techniques are used to analyze the changes in atrial activation patterns produced by contiguous RF induced lesions. In 12 Langendorff perfused rabbit hearts, left atrial activation maps were obtained before and after RF induction of epicardial lesions following a triple-phase sequential protocol: (phase 1) three separate lesions positioned vertically in the central zone of the left atrial wall; (phase 2) the addition of two lesions located between the central lesion and the upper and lower lesions; and (phase 3) the placement of four additional lesions between those induced in the previous phases. In six additional experiments a pathological analysis of the individual RF lesions was performed. In phase 1 (lesion diameter = 2.8+/-0.2 mm, gap between lesions = 3+/-0.8 mm), the activation process bordered the lesions line in two (250-ms cycles) and four experiments (100-ms cycles). In phase 2, activation bordered the lesions line in eight (250-ms cycles, P < 0.01 vs control) and nine experiments (100-ms cycles, P < 0.001), and in phase 3 this occurred in all experiments except one (both cycles, P < 0.001 vs control). In the experiments with conduction block, the increment of the interval between activation times proximal and distal to the lesions showed a significant correlation to the length of the lesions (r = 0.68, P < 0.05, 100-ms cycle). In two (17%) experiments, sustained regular tachycardias were induced with reentrant activation patterns around the lesions line. In conclusion, in this acute model, atrial RF lesions with intact tissue gaps of 3 mm between them interrupt conduction occasionally, and conduction block may be frequency dependent. Lesion overlap is required to achieve complete conduction block lines. Tachycardias with reentrant activation patterns around a lesions line may be induced. PMID- 11270695 TI - Reversal of tachycardia induced cardiomyopathy following ablation of repetitive monomorphic right ventricular outflow tract tachycardia. AB - Radiofrequency catheter ablation was performed in four adults with myocardial dysfunction related to repetitive monomorphic ventricular tachycardia (RMVT) originating in the right ventricular outflow tract. Serial echocardiographic assessment of left ventricular function before and after radiofrequency catheter ablation of RMVT showed complete reversal of left ventricular dysfunction without arrhythmia recurrence during 31+/-28 months follow-up. PMID- 11270694 TI - Spatial aspects of ventricular repolarization in postinfarction patients. AB - QT dispersion has been proposed to reflect the heterogeneity of ventricular repolarization, but a poor reproducibility limits its clinical usefulness. Spatial vectorcardiographic descriptors constitute a novel approach to quantify ventricular repolarization. To test the ability of vectorcardiographic descriptors to discriminate among different subsets of postinfarction patients, 50 consecutively recruited patients with acute MI, 50 patients with history of an old (> 6 months) MI, and 50 healthy controls were evaluated. The maximum and minimum QT and JT intervals and QT and JT dispersion were manually measured from a digitally recorded 12-lead ECG. X, Y, and Z leads were reconstructed from the 12-lead ECG. The amplitude of the maximum spatial T vector (spatial T amplitude), the angle between the maximum spatial QRS and T vectors (spatial QRS-T angle), and the frontal plane QRS-T angle were automatically calculated. The spatial T amplitude and the spatial QRS-T angle did not differ between patients with a recent and those with an old MI (P = 1). QT dispersion was significantly lower in patients with an old MI than in patients with a recent one (P = 0.002). The spatial repolarization descriptors showed better short-term reproducibility than the dispersion indices. In conclusion, the spatial T amplitude and the spatial QRS-T angle are accurate measures of ventricular repolarization that do not differ between patients with recent and those with old MI. The different QT dispersion values observed in this study between the two post-MI groups should be considered cautiously because of the low accuracy of the manual measurements. PMID- 11270696 TI - RV function in stable and unstable VT: is there a need for hemodynamic monitoring in future defibrillators? AB - During electrophysiological investigation of 22 patients with VT or aborted sudden cardiac death, arterial and RV pressures were measured. The time courses of mean arterial pressure (MAP), RV pulse pressure (RVPP), RV pulse pressure integral (RVPPI), and maximum right ventricular dP/dt (RV dP/dtmax) were followed during the first 15 seconds after VT induction. Compared to basal (preinduction) conditions, the RVPPI decreased by 41+/-10% (mean +/- SD) after 10-15 seconds of VT in 11 patients with stable VT and by 75+/-8% in 11 patients with unstable VT (MAP < 60 mmHg 15 s after VT onset). RVPP decreased by 13+/-11% after 10-15 seconds of VT in the stable VT group and by 50+/-16% in the unstable VT group. For RV dP/dtmax, these decreases were 4+/-22% in the stable VT group and 37+/-24% in the unstable VT group. There was a good correlation between percent decrease in MAP and percent decrease in RVPPI, RVPP, and RV dP/dtmax at 5-10 seconds (r = 0.86, 0.81, and 0.73, respectively) and 10-15 seconds (r = 0.84, 0.82, and 0.69, respectively) after VT onset. There was hardly any overlap of distributions of the individual values with the RVPPI parameter between the two VT groups. Comparing and correlating the percent decrease in mean arterial pressure with the RVPPI, RVPP, and RV dP/dtmax during induced VT, RVPPI demonstrated the most significant and specific changes in discriminating stable from unstable rhythms. However, by comparing RVPPI and RVPP using the area under the receiver operating characteristic curves, there was no significant statistical difference between the two parameters. By integrating rate criteria, electrogram signal analysis, and RVPPI or RVPP as a hemodynamic criterion, detection and treatment algorithms could improve the performance of future implantable defibrillators and avoiding shocks in VTs that can be terminated by antitachycardia pacing. PMID- 11270698 TI - Clinical outcome of patients who develop PAF after CABG surgery. AB - This was a retrospective analysis of patients who had CABG surgery at our hospital over a 12-month period to determine the intermediate-term prognosis of those who had developed PAF after their operation before hospital discharge. Of 317 patients who were operated by a single surgical group, 116 (37%) had AF postoperatively of whom 112 had the paroxysmal form. Of these, 36 were treated with class I or III antiarrhythmic drugs and rate control drugs (group 1) and 76 were treated with rate control alone (group 2). Group 3 consisted of 151 randomly selected patients who did not have AF. All patients were reevaluated at 6 weeks to determine their rhythm and clinical status. Only one patient each in groups 1 and 2 was in AF 6 weeks after discharge. There was a trend toward a higher mortality and morbidity in group 2 patients. PAF after coronary surgery appears to be a self-limited disease process. In this cohort of patients, the rate of recurrence of AF after discharge was similar in patients receiving class I or class III antiarrhythmic drugs together with rate control agents compared to those receiving rate control drugs alone. PMID- 11270697 TI - Discrimination between ventricular and supraventricular tachycardia by dual chamber cardioverter defibrillators: importance of the atrial sensing function. AB - Although the addition of atrial sensing in dual chamber ICDs may improve the ability of the device to discriminate between supraventricular (SVT) and ventricular tachycardia (VT), atrial sensing errors may also negatively affect tachycardia classification. This prospective study evaluated the incidence of atrial sensing errors in a dual chamber ICD and their impact on VT/SVT discrimination. In 145 patients, a dual chamber ICD (Defender) was implanted. Analysis of 1,241 tachycardia episodes stored during a mean follow-up of 14+/-8 months revealed atrial sensing errors in 817 (66%) episodes. Upon expert review, device-based classification was confirmed in 509 (98%) of 522 SVT episodes. No false device-based SVT classification was related to atrial sensing errors. Of 719 episodes classified as VT by the device, 645 (90%) were confirmed. There were 74 episodes of false-positive VT detection. Of these, 63 were related to atrial sensing errors: atrial undersensing in 58 (92%) and atrial oversensing in 5 (8%) episodes. Atrial sensing errors led to incorrect VT/SVT discrimination in 51 (4%) of 1,241 episodes. Only the occurrence of paroxysmal atrial fibrillation and abdominal site of device implantation showed a significant influence on false VT/SVT discrimination. Atrial sensing errors are frequently encountered in dual chamber ICDs. Due to the VT/SVT discrimination algorithm, atrial sensing errors only led to misclassification in 4 % of all episodes, mainly due to atrial undersensing. No VT underdetection due to atrial oversensing occurred. PMID- 11270699 TI - The effects of aging on AV nodal recovery properties. AB - The purpose of this study was to assess the changes of AV nodal recovery properties with aging. Although in children and young adults it was found that there were age dependent changes in their AV nodal recovery properties, in the older population this information was not available. In 92 subjects (aged 16-92 years) without AV nodal disease or dual AV nodal pathway physiology, their AV nodal recovery curves were studied by delivering premature atrial extrastimuli coupled to basic atrial beats during cardiac electrophysiological study. Data were analyzed using linear regression and curve-fitting techniques. Patients were grouped by age, group I < 40 years (n = 33), group II 40-59 years (n = 26), and group III > 60 years (n = 33). The results showed that the AV nodal recovery curve did not change significantly in the aging process except that the AV nodal effective refractory period had a positive correlation with increasing age. The latter was significantly increased in group III when compared to group I or group II. For this parameter, when patients whose AV nodal refractory period was limited by the atrial refractory period were excluded, there was still a statistically significant increase in group III compared to group II (P < 0.05): group I (n = 27): 202+/-42 ms; group II (n = 17): 197+/-26 ms; and group III (n = 17): 224+/-46 ms. The results suggest that the AV nodal recovery curve remains unchanged once it reaches adulthood, with the exception that the nodal effective refractory period becomes slightly longer after age 60. PMID- 11270700 TI - Force and torque effects of a 1.5-Tesla MRI scanner on cardiac pacemakers and ICDs. AB - Magnetic resonance imaging (MRI) is a widely accepted tool for the diagnosis of a variety of disease states. However, the presence of an implanted pacemaker is considered to be a strict contraindication to MRI in a vast majority of centers due to safety concerns. In phantom studies, the authors investigated the force and torque effects of the static magnetic field of MRI on pacemakers and ICDs. Thirty-one pacemakers (15 dual chamber and 16 single chamber units) from eight manufacturers and 13 ICDs from four manufacturers were exposed to the static magnetic field of a 1.5-Tesla MRI scanner. Magnetic force and acceleration measurements were obtained quantitatively, and torque measurements were made qualitatively. For pacemakers, the measured magnetic force was in the range of 0.05-3.60 N. Pacemakers released after 1995 had low magnetic force values as compared to the older devices. For these devices, the measured acceleration was even lower than the gravity of the earth (< 9.81 N/kg). Likewise, the torque levels were significantly reduced in newer generation pacemakers (< or = 2 from a scale of 6). ICD devices, except for one recent model, showed higher force (1.03 5.85 N), acceleration 9.5-34.2 N/kg), and torque (5-6 out of 6) levels. In conclusion, modern pacemakers present no safety risk with respect to magnetic force and torque induced by the static magnetic field of a 1.5-Tesla MRI scanner. However, ICD devices, despite considerable reduction in size and weight, may still pose problems due to strong magnetic force and torque. PMID- 11270702 TI - Adjustment of the evoked response sensitivity after hospital discharge in pacemaker patients with automatic ventricular threshold tracking activated. AB - Automatic threshold tracking in cardiac pacemakers allows ventricular capture verification and self-adaptive pacing output regulation. The Autocapture algorithm detects the evoked response (ER) signal immediately after the pacing pulse to verify the efficacy of ventricular pacing. Before hospital delivery, the ER sensitivity must be programmed individually so that the pacemaker detects the ER signal adequately without sensing lead polarization. The aims of the study were to assess the frequency of patients in whom Autocapture could be activated and whether the ER sensitivity had to be adjusted after hospital discharge. The study included 44 patients who received the VVIR pacemaker Regency SR+ (St. Jude Medical) connected to the model 1450 T pacing lead. ER signal, lead polarization, and ER sensitivity were evaluated before hospital discharge and 1, 3, and 6 months after implantation. The system recommended activating Autocapture in 42 of 44 patients. The mean ER signal was 8.4+/-1.2 mV at discharge, 9.0+/-3.9 mV at month 1, 8.9+/-4.9 mV at month 3, and 9.3+/-4.5 mV at month 6. Polarization was 1.0+/-0.1 mV at discharge, 1.1+/-0.5 mV at month 1, 1.1+/-0.2 mV at month 3, and 1.1+/-0.5 mV at month 6. Mean ER sensitivity was 3.7+/-1.8 mV at discharge, 4.0+/ 1.8 mV after 1, 4.1+/-2.2 mV after 3, and 4.1+/-1.8 mV after 6 months. ER sensitivity could remain unadjusted in 14 patients. Programming to a less sensitive ER setting from 2.9+/-1.2 mV to 4.3+/-1.5 mV was possible in 21 patients. Programming to a more sensitive ER setting from 4.1+/-1.1 mV to 2.5+/ 0.9 mV was required in nine patients because of the decrease of the ER signal. The automatic threshold tracking algorithm Autocapture could be activated in 95% of patients. Programming to more sensitive ER settings was recommended in 21% of the patients after hospital discharge. Therefore, ER signal and polarization must be checked at each follow-up, as a decrease in ER signal amplitude can make reprogramming of the ER sensitivity necessary. There is no risk for the patient if the ER is not sensed, as high voltage backup stimulation is present. PMID- 11270701 TI - Extension of generator longevity by use of high impedance ventricular leads. AB - The resistance of a pacing lead negatively correlates to current consumption. A prospective, randomized trial was conducted to evaluate the effect of a high impedance ventricular lead (CapSure Z) on generator longevity compared to a conventional lead (CapSure SP) eighty-nine patients were included in the study (51 male, 37 female, age 70.0+/-10.3 years). Forty-six patients received a CapSure SP lead (5024 bipolar), and 43 patients received a CapSure Z lead (5034 bipolar) in a randomized fashion. Follow-up data collected at 5 days, 3, 6, and 12 months postimplant included: lead impedance, pacing and sensing thresholds, impulse energy, and estimated time to replacement. All parameters were collected via pacemaker telemetry; the time to replacement was calculated automatically by a programmed algorithm of the pacemaker. There was no difference in the performance of the atrial lead when a dual chamber device was indicated. The CapSure Z leads displayed statistically significant higher impedance values than the CapSure SP lead in all follow-up periods. There was no significant difference in lead related complications. No significant differences were observed between pacing and sensing thresholds in both groups. The CapSure Z leads provided a significant reduction in current drain, resulting in a reduction of mean energy consumption at the 12-month follow-up from 10.4+/-5.0 microJ in the CapSure SP group to 6.6+/-1.4 microJ in the CapSure Z group (median from 9.9 microJ to 6.9 microJ, respectively), providing an estimated increase in mean longevity of more than 1 year from 81.1+/-23.5 months in the CapSure SP group to 94.5+/-13.4 months in the CapSure Z group (median: 76.5 months to 95.0 months, respectively). The use of a high resistance lead for ventricular pacing appears to result in a clinically relevant extension of generator longevity. PMID- 11270703 TI - Is sinus node modification appropriate for inappropriate sinus tachycardia with features of postural orthostatic tachycardia syndrome? AB - Inappropriate sinus tachycardia and postural orthostatic tachycardia are ill defined syndromes with overlapping features. Although sinus node modification has been reported to effectively slow the sinus rate, long-term clinical response has not been adequately assessed. Furthermore, whether patients with postural orthostatic tachycardia would benefit from sinus node modification is unknown. The study prospectively assessed the short- and long-term clinical outcomes of seven consecutive female patients with postural orthostatic tachycardia syndrome and inappropriate sinus tachycardia who were treated with sinus node modification. The study was conducted in a tertiary care center. The electrophysiological and clinical responses were prospectively assessed as defined by autonomic function testing, including Valsalva maneuver, deep breathing, tilt table testing, and quantitative sudomotor axonal reflex testing. Among the study population (mean age was 41+/-6 years), 5 (71%) patients had successful sinus node modification. At baseline, heart rates were 101+/-12 beats/min before modification and 77+/-9 beats/min after modification (P = 0.001). With isoproterenol, heart rates were 136+/-9 and 105+/-12 beats/min (P = 0.002) before and after modification, respectively. The mean heart rate during 24 hour Holter monitoring was also significantly reduced: 96+/-9 and 72+/-6 beats/min (P = 0.005) before and after modification, respectively. Despite the significant reduction in heart rate, autonomic symptom score index (based on ten categories of clinical symptoms) was unchanged before (15.6+/-4.1) and after (14.6+/-3.6) sinus node modification (P = 0.38). Sinus rate can be effectively slowed by sinus node modification. Clinical symptoms are not significantly improved after sinus node modification in patients with inappropriate sinus tachycardia and postural orthostatic tachycardia. A primary subtle autonomic disregulation is frequently present in this population. Sinus node modification is not recommended in this patient population. PMID- 11270704 TI - Typical atrial flutterlike tachycardia developing after inferior vena cava tricuspid annulus isthmus ablation. PMID- 11270705 TI - Biventricular pacing as alternative therapy for dilated cardiomyopathy associated with congenital heart disease. AB - Biventricular, alternative, and multisite pacing are currently being explored to improve cardiac function among patients with medically refractory, end-stage dilated cardiomyopathies. Although, due to inherent myocardial abnormalities, patients with repaired congenital heart defects may be at a greater risk than others to develop heart failure, often requiring cardiac transplantation. The efficacy of biventricular pacing among these patients is unknown. This report presents a patient with successfully repaired congenital heart disease in infancy who developed a symptomatic dilated cardiomyopathy at 22 years of age. Following biventricular pacing, systemic ventricular function showed a 14% improvement in ventricular dP/dt. One month later, subjective symptoms improved and cardiac ultrasound illustrated a 125% increase in fractional area of change. Exercise stress testing showed a 17% improvement in aerobic work capacity. PMID- 11270706 TI - Inappropriate shock therapy in a heart failure defibrillator. AB - A 63-year-old male with dilated cardiomyopathy underwent implantation of a "heart failure" defibrillator capable of biventricular pacing. He received an inappropriate shock 5 hours after the procedure. Stored electrograms revealed that during each sinus beat the ventricular channel recorded up to three separate events. These resulted from far-field atrial sensing by the coronary venous lead, appropriate right ventricular sensing, then delayed left ventricular sensing (the result of left bundle branch block). As a consequence of far-field left atrial sensing the two subsequent ventricular electrograms fell within the VF zone. Following an atrial premature beat, VF detection criteria were satisfied and shock therapy delivered. Although coronary venous lead repositioning eliminated far-field atrial sensing, double counting of the widely split right and left ventricular electrograms still occurred during sinus rhythm. Shortening the programmed AV delay resulted in constant biventricular pacing with a single electrogram. PMID- 11270707 TI - Dysfunction of phrenic pacemakers induced by metallic rescue blankets. AB - Phrenic pacing can restore diaphragmatic contractions in patients with central respiratory paralysis. It relies on radiofrequency transmission of energy from an external unit to implanted receivers through circular coil antennas. The case of a patient is reported in whom severe hypoventilation occurred following the use of a metallic rescue blanket. The phenomenon was confirmed in two subsequent patients and during benchmark tests. Possible mechanisms include reflection and diffusion of high frequency waves by a Faraday-like effect. Patients with implanted devices relying on telemetric control or powering, and their care givers, should be warned against the use of metallic rescue sheets. PMID- 11270708 TI - Unipolar defibrillator? AB - An unusual case of "unipolar" pacing and myopotential over-sensing leading to an inappropriate ICD shock in a patient with an implanted defibrillator is reported. The reasons for unipolar behavior in a system using a committed bipolar device are discussed. PMID- 11270710 TI - Termination of pacemaker endless loop tachycardia by an atrial extrasystole. AB - This article documents the termination of a pacemaker endless loop tachycardia by a critically timed atrial extrasystole. Although predictable electrophysiologically, this mode of termination has not been previously reported. The observation is conceptually important because it provides the final link in establishing the similarity of endless loop tachycardia and spontaneous reentrant AV nodal or junctional tachycardias in terms of initiation and termination by single atrial or ventricular extrasystoles. PMID- 11270709 TI - Possible role of epicardial left ventricular programmed stimulation in Brugada syndrome. AB - A patient with recurrent syncope due to polymorphic ventricular tachycardia was diagnosed with Brugada syndrome. Programmed right ventricular stimulation could not induce arrhythmia. Epicardial stimulation from a left ventricular site through the coronary sinus led to polymorphic VT. The stimulation protocol for risk stratification in Brugada syndrome is discussed. PMID- 11270711 TI - Inactivation of a ventricular tachycardia preventive algorithm during automatic mode switching for atrial tachyarrhythmia. AB - A patient with a dual chamber implantable defibrillator and pause dependent VT in whom a rate smoothing algorithm failed to operate during automatic mode switching due to device idiosyncrasy is reported. Preventive measures are discussed. PMID- 11270712 TI - Essentials of randomized clinical trials. PMID- 11270713 TI - Consensus statement on indications, guidelines for use, and recommendations for follow-up of implantable cardioverter defibrillators. North American Society of Electrophysiology and Pacing. PMID- 11270714 TI - Allicin release under simulated gastrointestinal conditions from garlic powder tablets employed in clinical trials on serum cholesterol. AB - The failure of five recent clinical trials to show significant reduction in elevated serum cholesterol by a single brand of allicin-standardized garlic powder tablets is in contrast to many prior positive studies with the same brand. The hypocholesterolemic activity of garlic is mainly due to allicin, a compound that is produced by the acid-sensitive garlic enzyme, alliinase, only after tablet consumption. Therefore, the allicin-releasing ability of ten lots of these tablets--manufactured over the same years that the positive and negative clinical trials were conducted (1989-1997)--was determined under simulated gastrointestinal dissolution conditions, as defined by U.S. Pharmacopeia Method 724A. It was found that the older lots were more resistant to acid-disintegration (2.5 h vs. 1.3 h, P < 0.001) and that they released three times as much allicin (44% vs. 15 % of their potential, P < 0.001) as the newer lots. A second brand of tablets employed in a recent negative trial released no detectable amount of allicin, while a third set of tablets with high allicin release was used in a trial that gave positive effects. Hence, the persons involved in the recent negative clinical trials probably received considerably less allicin than did those in the older positive studies, possibly accounting for much of the discrepancy in the outcomes. In conclusion, clinical trials using garlic powder tablets to assess any effect of garlic that might be related to allicin, as most are, cannot be considered valid for garlic when the trial shows no effect, unless the expected allicin release from the tablets has at least been determined under standardized drug release conditions (USP 724A). PMID- 11270715 TI - The inhibitory effects of ginsenosides on protein tyrosine kinase activated by hypoxia/reoxygenation in cultured human umbilical vein endothelial cells. AB - 27 individual ginsenosides and aglycones, together with five extracts from ginseng roots, ginseng leaves, American ginseng roots, American ginseng leaves and non-saponin fraction from roots of Panax ginseng, were tested for their effects on protein tyrosine kinase (PTK) activation induced by an in vitro hypoxia/reoxygenation (H/R) model in cultured human umbilical vein endothelial cells (HUVEC). The results indicated that ginsenoside-Rb1 (3), -Rd (7), -Ra1 (1) and -Ro (27) showed significant inhibitory effects on PTK activation induced by H/R. Dose-response experiments revealed that ginsenoside-Rb1 was the most active compound and it completely blocked PTK activation at a wide range of concentrations. Most protopanaxadiol-type ginsenosides and some protopanaxatriol type saponins also showed significant effects on PTK activation. However, the crude extracts did not protect against H/R-induced PTK activation. PMID- 11270716 TI - Inhibitory effect of decoction of Perilla frutescens on cultured murine mesangial cell proliferation and quantitative analysis of its active constituents. AB - The leaves of Perilla frutescens (perilla) are a common herb used in Japan for garnishing raw seafood. Previously, we reported that a decoction of perilla leaves had suppressive effects on the progression of glomerulonephritis in an animal model of spontaneous IgA nephropathy. The objective of the present study was to isolate anti-nephritic constituents in the perilla decoction under the guidance of its in vitro anti-proliferative activity on cultured murine mesangial cells, and to measure the contents of the active constituents in decoctions prepared from various perilla chemotypes, which differ in their composition of essential oils and/or pigments. DNA synthesis of cultured mesangial cells induced by 1% fetal calf serum was significantly inhibited by the perilla decoction (IC50 values, 8.8 microg/ml). Caffeic acid, luteolin 7-O-[beta-glucuronosyl(1-->2)beta glucuronide], apigenin 7-O-[beta-glucuronosyl(1-->2)beta-glucuronide], scutellarin, and rosmarinic acid were isolated as active constituents. The contents of these phenolic compounds were not significantly different among chemotypes of P. frutescens. Considering the relation between the contents in the perilla decoction and the activities of these compounds, rosmarinic acid represents the in vitro anti-proliferative effect of perilla decoction. PMID- 11270717 TI - Extraction, isolation and characterisation of antitumor principle, alpha-hederin, from the seeds of Nigella sativa. AB - We have previously reported the in vitro cytotoxic activity of column fraction 5 (CC-5) of an ethanolic extract of Nigella sativa seeds. In this study, the effect of CC-5 was evaluated for its in vivo antitumor activity against i.p. (intraperitoneally) implanted murine P388 leukemia and s.c. (subcutaneously) implanted LL/2 (Lewis lung carcinoma) cells in BDF1 mice (C57BL/6 x DBA/2 mice). CC-5 at doses of 200 and 400 mg/kg b.w. prolonged the life span of these mice by 153% compared to DMSO-treated control mice. The antitumor activity of a 21-day treatment of CC-5 against s.c. implanted LL/2 was tested in mice using four experimental protocols as described in the methods. In protocols C and D, CC-5 at a dose of 400 mg/kg b.w. produced significant tumor inhibition rate (TIR) values of 60% (P < 0.001) and 70% (P < 0.001) respectively. Alpha-hederin, a triterpene saponin isolated from CC-5, when given i.p. for 7 days at doses of 5 and 10 mg/kg b.w. to mice with formed tumors, produced significant dose-dependent TIR values of 48% (P < 0.05) and 65% (p < 0.01) respectively on day 8 and 50% (P < 0.01) and 71% (P < 0.001), respectively, on day 15, compared to 81% (P < 0.01) on day 8 and 42% (P < 0.01) on day 15 in the cyclophosphamide (CP)-treated group. The underlying mechanism(s) of antitumor activity of alpha-hederin remain to be established. PMID- 11270718 TI - Inhibitors of bacterial topoisomerases: mechanisms of action and resistance and clinical aspects. AB - The quinolone class of inhibitors of bacterial type II topoisomerases has gained major clinical importance during the last years due to improvements in both pharmacokinetic and pharmacodynamic properties. These include favorable bioavailability allowing oral administration, good tolerability, high tissue concentrations as well as superior bactericidal activity against a broad spectrum of clinically relevant pathogens, like enterobacteria, Pseudomonas aeruginoso, Staphylococcus aureus, and Streptococcus pneumoniae. In addition, no enzymatic mechanism of drug inactivation exists in bacteria and no indications for transfer of clinically relevant resistance exist. Nevertheless, resistance is being increasingly reported, even for naturally highly susceptible species like Escherichia coli. The underlying mechanisms of resistance include alterations in both bacterial targets, DNA gyrase and topoisomerase IV, often combined with mutations affecting drug accumulation, e.g., by increased drug efflux, reduced drug influx, or both. Investigations aiming at understanding the molecular mechanisms of quinolone action and resistance in more detail should provide a basis for a rational design of more potent derivatives. In addition, a prudent use of these highly valuable "magic bullets" is necessary to preserve their potential for the future. PMID- 11270719 TI - Magnolol from Magnolia officinalis inhibits 11beta-hydroxysteroid dehydrogenase without increases of corticosterone and thymocyte apoptosis in mice. AB - Magnolol is an 11beta-hydroxysteroid dehydrogenase (11beta-HSD) inhibitor contained in Magnolia officinalis which is used in Chinese remedies. We have reported that glycyrrhetinic acid, a strong 11beta-HSD inhibitor isolated from licorice, induces apoptosis of murine thymocytes via accumulation of corticosterone. In this paper, we report that magnolol inhibited 11beta-HSD without increases in the blood concentration of corticosterone and in thymocyte apoptosis in mice. Oxidative activities of the enzyme (from corticosterone to 11 dehydrocorticosterone) in liver, kidney and thymus in vitro were examined 24 h after a single administration of magnolol. Magnolol inhibited the enzyme activity in kidney (P < 0.0001) and thymus (P < 0.002), while the activity in liver was not affected. Blood concentrations of corticosterone in the magnolol-treated mice were unexpectedly lower than those in the control animals (P < 0.002). This means that the inhibition of 11beta-HSD by magnolol did not increase the systemic level of corticosterone which is relevant to thymocyte apoptosis. Accordingly, our flow cytometric analysis of thymocytes after magnolol treatment showed no change in the number of apoptotic cells. We concluded that unlike glycyrrhetinic acid, magnolol selectively inhibited 11beta-HSD in kidney and thymus but not in liver, so that the blood concentrations of corticosterone could not exceed the control level. PMID- 11270720 TI - Reversal caused by n-butylidenephthalide from the deficits of inhibitory avoidance performance in rats. AB - The present study was designed to investigate the mechanism of action of n butylidenephthalide on the deficits of inhibitory avoidance performance induced by drugs in rats with piracetam as a positive control. n-Butylidenephthalide attenuated the scopolamine-induced and mecamylamine-induced acquisition impairment, and also attenuated the acquisition impairment induced by scopolamine plus mecamylamine. Furthermore, scopolamine methylbromide, a peripheral cholinergic muscarinic receptor antagonist, did not block the counteracting effect of n-butylidenephthalide on the scopolamine-induced acquisition impairment. n-Butylidenephthalide attenuated the impairment of inhibitory avoidance performance induced by the central acetylcholinergic neurotoxin AF64A administered intracisternally. From the above results, we suggest that n butylidenephthalide attenuated the deficits of inhibitory avoidance performance induced by drugs, which are the effects related to activating the central but not the peripheral cholinergic neuronal system via muscarinic and nicotinic receptors. PMID- 11270721 TI - Investigations of the mechanism of membrane activity of selected triterpenoid saponins. AB - The experiments with different triterpenoid saponins showed a significant difference in the microphysiometric activity between acylated and non-acylated compounds. Only acylated saponins in low concentrations were able to activate the metabolism of endothelial cells after short-time incubation in a transient manner lasting approximately 2 hours. The strongest effect was produced by virgaureasaponin b, an incubation with 4 microg/ml over 6 min induced a stimulation of the metabolic activity of 190% maximum after 30 min. This treatment did not induce non-reversible toxic effects, and could be repeated in the same way after a recovery period. The saponins without acylation in the glycosidic part did not induce metabolic stimulation, even in concentrations higher than observed for the acylated compounds. The current interpretation of the results is connected with the suggestion that especially acylated saponins are able to integrate into the cellular membrane transiently. This integration might induce pore-like structures which change the membrane permeability associated with alterations in the ionic homeostasis between intracellular and extracellular compartments. PMID- 11270722 TI - Effects of saponins on the water solubility of different model compounds. AB - The influence of saponins on the water solubility of compounds having poor aqueous solubility was investigated with the model compounds digitoxin, rutin and aesculin. The saponins that were tested represent the most abundant structural types. Only slight effects on aqueous solubility were obtained for all the model compounds. These effects include both enhancements and reductions in aqueous solubility depending on the saponin, the concentration of the saponin solution and the model compound. Structure-activity relationships of general significance were not observed. Hence, saponins generally should not be regarded as solubilizers. PMID- 11270723 TI - Abietane diterpenoids from the cones of Larix kaempferi and their inhibitory effects on Epstein-Barr virus activation. AB - Four known (1-3, 8) and four new abietane diterpenes, 15-hydroxy 8alpha,14alpha,12alpha,13alpha-diepoxyabietan-18-oic acid (4), 7alpha,8alpha,13beta,14beta-diepoxyabietan-18-oic acid (5), 18-nor-abieta-8,11,13 triene-4alpha,7alpha,15-triol (6), and abieta-8,11,13-triene-7alpha,15,18-triol (7) were isolated from the CHCl3 extract of the cones of Larix koempferi. A known compound, 13,14-seco-13,14-dioxoabiet-13-en-18-oic acid (8) was isolated from natural sources for the first time. Their structures were determined by chemical and spectroscopic methods including 1D and 2D NMR techniques. The absolute stereostructure of 5 was determined by X-ray crystallographic analysis. The inhibitory effects of these compounds on Epstein-Barr virus early antigen (EBV EA) activation induced by the tumor promoter, 12-O-tetradecanoylphorbol-13 acetate (TPA), were examined as a primary screening for antitumor promotors. PMID- 11270724 TI - Three new stilbene trimers from the lianas of Gnetum hainanense. AB - Three new stilbene trimers, gnetuhainins M-O (1-3), were isolated from the lianas of Gnetum hainanense. Their structures and relative configurations were determined by spectroscopic evidence, especially on 2D NMR analysis. The anti inflammatory activity has been tested for the isolated compounds. PMID- 11270725 TI - Isolation and frontier molecular orbital investigation of bioactive quinone methide triterpenoids from the bark of Salacia petenensis. AB - The crude dichloromethane bark extract of Salacia petenensis (Hippocrateaceae) from Monteverde, Costa Rica, shows antibacterial and cytotoxic activity. Bioactivity-directed separation led to the isolation of tingenone and netzahualcoyonol as the biologically active materials. Also isolated from the extract were 3-methoxyfriedel-2-en-1-one (a new natural product) and 29 hydroxyfriedelan-3-one. The structures of these compounds were elucidated on the basis of NMR spectral analysis. Molecular orbital calculations have been carried out using the semi-empirical PM3 and Hartee-Fock 3-21G ab initio techniques on the quinone-methide nortriterpenoids tingenone and netzahualcoyonol, as well as on the nucleotide bases adenine, guanine, cytosine, and thymine. The molecular orbital calculations suggest that a possible mode of cytotoxic action of quinone methide triterpenoids involves quasi-intercalative interaction of the compounds with DNA followed by nucleophilic addition of the DNA base to carbon-6 of the triterpenoid. PMID- 11270726 TI - Quantitative determination of secoiridoid glucosides in in vitro propagated plants of Gentiana davidii var. formosana by high performance liquid chromatography. AB - A simple protocol for in vitro mass propagation of Gentiana davidii var. formosana (Gentianaceae) has been developed. Multiple shoot development was achieved by culturing the stem node explants on Murashige and Skoog (MS) medium supplemented with 4.44 microM N6-benzyladenine (BA). The shoots were multiplied by subculturing on MS medium supplemented with 1.07-10.74 microM alpha naphthaleneacetic acid (NAA) and 8.88 microM BA. Shoots were rooted on MS basal medium supplemented with various auxins. Shoots rooted on growth regulator-free medium were transferred to peat moss:vermiculite mixture and acclimatized in the growth chamber. The contents of gentiopicroside and swertiamarin, the two important secoiridoid glucosides, in different plant material were determined by high performance liquid chromatography (HPLC). The analysis revealed that the content of gentiopicroside and swertiamarin in the aerial and underground parts of G. davidii var. formosana was higher than the marketed crude drug (underground parts of G. scabra) and varied with the age of the plant. PMID- 11270727 TI - Monoamine oxidase inhibitors from rhizoma of Coptis chinensis. AB - Three protoberberine alkaloids jatrorrhizine, berberine and palmatine were isolated from the monoamine oxidase (MAO) inhibitory fraction of the methanol extract of Coptis chinensis rhizoma. Jatrorrhizine was shown to inhibit non competitively both MAO-A and -B from rat brain mitochondria with the IC50 values of 4 and 62 microM, respectively. Berberine only competitively inhibited MAO-A with an IC50 values of 126 microM whereas palmatine exhibited, up to 200 microM, no inhibition on any type of the enzyme. The structure-activity relationship was briefly discussed. PMID- 11270728 TI - Inhibitory effect of tetrandrine on dopamine biosynthesis and tyrosine hydroxylase in PC12 cells. AB - The effect of tetrandrine, a bis-benzylisoquinoline alkaloid, on dopamine biosynthesis in PC12 cells was investigated. Tetrandrine at a concentration of 3.0 microM decreased dopamine content by 59.4% (IC50 = 2.4 microM) and intracellular tyrosine hydroxylase (TH) activity was inhibited by the treatment of tetrandrine (49.8% inhibition at 3.0 microM) compared with control. We next examined the effects of tetrandrine on the kinetics of PC12 TH. The PC12 TH was obtained from PC12 cells with minor purification. Tetrandrine inhibited the PC12 TH activity by 40.6% at a concentration of 45 microM and exhibited noncompetitive inhibition on the enzyme using L-tyrosine as a substrate (Ki = 60.8 microM). These results suggest that the inhibition of TH activity by tetrandrine may partially contribute to the decrease in dopamine biosynthesis in PC12 cells. PMID- 11270729 TI - Cytotoxicity of iridals, triterpenoids from Iris, on human tumor cell lines A2780 and K562. AB - Six different triterpenoids (known iridals) extracted from Iris germanica L. were purified and bioassayed on two cultured human tumor cell lines: A2780 and K562 (and for each one a drug-sensitive and a drug-resistant cell line). All of the tested iridals had an IC50 in the 0.1 to 5.3 microg/ml range. Some of them were shown to be more effective than doxorubicine. Toxicity of iridals on multi-drug resistance (MDR) expressing cell lines seemed to indicate that the effects of these triterpenoids are less affected by MDR than those of doxorubicine or taxol. PMID- 11270730 TI - Composition and antimicrobial activity of the essential oil of Salvia ringens. AB - The essential oils of Salvia ringens (samples A and B), were analyzed by means of GC and GC/MS. From the seventy-five identified constituents representing 99.82 and 99.86% of the oils, 1,8-cineole and alpha-pinene were the major components. Furthermore, sample B exhibited a very interesting antimicrobial profile after it was tested against six gram (+/-) bacteria and three pathogenic fungi. PMID- 11270731 TI - Composition and antimicrobial activity of the essential oil of Baccharis notosergila. AB - The essential oil of the aerial parts of Baccharis notosergila was examined by GC and GC-MS. Thirty-one constituents were identified representing 96.4% and alpha pinene, limonene, beta-caryophyllene, and spathulenol were found to be the major components. Furthermore, the oil was tested against eight gram-positive and negative bacteria and it was found that they exhibited moderate antibacterial activity. PMID- 11270732 TI - Antifungal chalcones from Maclura tinctoria. AB - Five prenylated flavonoids, including one new natural product, were isolated from an ethanol extract of the leaves of Maclura tinctoria (L.) Gaud. The new compound has been characterized as 2',4',4,2''-tetrahydroxy-3'-[3''-methylbut-3'' enyl]chalcone (1). The known compounds were identified as 2',4',4-trihydroxy-3' [3''-methylbut-3''-enyl]chalcone (isobavachalcone) (2), 4,2'-dihydroxy-2''-[1 hydroxy-1-methylethyl]-2'',3''-dihydrofurano[4'',5'':3',4']chalcone (bakuchalcone) (3), 4,4',5''-trihydroxy-6'',6'' dimethyldihydropyrano[2'',3'':5',6'']chalcone (bavachromanol) (4), and 5,7,3',4' tetrahydroxy-6,8-diprenylisoflavone (6,8-diprenylorobol) (5). All the isolated compounds were evaluated against the AIDS-related opportunistic fungal pathogens, Candida albicans and Cryptococcus neoformans. Compound 2 was active against both yeasts. PMID- 11270734 TI - Molluscicidal and trypanocidal activities of lapachol derivatives. AB - The activity of the potassium salt of lapachol against the snail Biomphalaria glabrata and its egg masses was tested. The obtained IC50 values (2.70 ppm and 1.43 ppm, respectively) are indicative of a strong activity. Lapachol derivatives were also assayed against infective trypomastigote blood forms of T. cruzi and the triacetoxy derivative of reduced lapachol showed relevant trypanocidal activity, killing 95.7% of the parasites at the concentration of 42 microg/mL. PMID- 11270733 TI - In vitro antiplasmodial activity of 4-phenylcoumarins from Exostema mexicanum. AB - The stem bark of Exostema mexicanum (Rubiaceae) is used in Latin American folk medicine as a quinine substitute for malaria treatment. Bioassay-guided fractionation of lipophilic and hydrophilic extracts from the stem bark and branches yielded two previously undescribed 4-phenylcoumarins: 4',8-dihydroxy-5,7 dimethoxy-4-phenylcoumarin (exomexin A) and 3',4'-dihydroxy-5,7,8-trimethoxy-4 phenylcoumarin (exomexin B). Together with five known derivatives the in vitro activities against a chloroquine-sensitive strain (poW) and a chloroquine resistant strain (Dd2) of Plasmodium falciparum have been evaluated. The most lipophilic compound, 4',5,7,8-tetramethoxy-4-phenylcoumarin (O-methylexostemin) revealed the strongest antiplasmodial activity (IC50 values: 3.6 microg/ml [poW], 1.6 microg/ml [Dd2]). PMID- 11270735 TI - Effects of silymarin on the acute stage of the trinitrobenzenesulphonic acid model of rat colitis. AB - The intestinal anti-inflammatory activity of several doses of silymarin was tested in the acute stage of the trinitrobenzenesulfonic acid (TNBS) model of rat colitis. The results obtained show that oral pre-treatment with 50 mg/kg of silymarin significantly attenuated macroscopic colonic damage as well as reduced colonic myeloperoxidase activity compared to non-treated colitic animals. The beneficial effect was accompanied by an improvement in the colonic oxidative status, which was altered in colonic inflammation, by preventing glutathione depletion and reducing malonyldialdehyde production. This suggests that the well known antioxidant properties of silymarin can participate in its intestinal anti inflammatory activity. In addition, a preservation in the colonic absorptive function was also observed, and this effect can also account for the colonic protective effect observed in this model of acute colitis. PMID- 11270736 TI - High yield of podophyllotoxin from leaves of Podophyllum peltatum by in situ conversion of podophyllotoxin 4- O-beta-D-glucopyranoside. AB - Rehydration of powdered tissues of Podophyllum peltatum L. prior to extraction with an organic solvent allows endogenous beta-glucosidases to hydrolyze lignan 4 O-beta-D-glucosides in situ and increase the yield of podophyllotoxin. Aqueous extraction of rhizomes and leaves of P. peltatum yielded 4- to 10-fold greater quantities of podophyllotoxin than the traditional ethanolic extraction. Most significantly, leaves were shown to contain over 52 mg of podophyllotoxin per g of dry weight (5.2%), exceeding levels previously reported from any source. These results point to the use of leaves harvested from cultivated P. peltatum as an attractive alternative to the destructive collection of natural populations. PMID- 11270737 TI - 7-feruloylloganin: an iridoid glucoside from stems of Lonicera insularis. AB - A new iridoid glucoside, 7-feruloylloganin (1), was isolated from stems of Lonicera insularis, along with six known lignans including (-)-pinoresinol, 9alpha-hydroxypinoresinol, balanophonin, erythro-1-(4-hydroxy-3-methoxyphenyl)-2 [4-[2-formyl-(E)-vinyl]-2-methoxyphenoxy]-propane-1,3-diol, threo-1-(4-hydroxy-3 methoxyphenyl)-2-[4-[2-formyl-(E)-vinyl]-2-methoxyphenoxy]-propane-1,3-diol and buddlenol A. The structure of 1 was determined by analyses of 2D NMR (1H-1H COSY, HMQC and HMBC) and HRFABMS. PMID- 11270738 TI - The influence of the radiofrequency excitation and conversion pulses on the lineshapes and intensities of the triple-quantum MAS NMR spectra of I = 3/2 nuclei. AB - A rigorous examination of the various multiple-quantum magic angle spinning sequences is carried out with reference to sensitivity enhancement in the isotropic dimension and the lineshapes of the corresponding MAS peaks in the anisotropic dimension. An echo efficiency parameter is defined here, which is shown to be an indicator of the performance aspects of the various sequences. This can be used in the design of further new experiments in this field. A consequence of such a systematic analysis has been the combination of a spin-lock pulse for excitation of multiple-quantum coherences and an amplitude-modulated pulse for their conversion to observable single-quantum coherences. This approach has resulted in an improved performance over other sequences with respect to both the anisotropic lineshapes and the isotropic intensities. PMID- 11270740 TI - Carbon-13 and fluorine-19 NMR spectroscopy of the supramolecular solid p-tert butylcalix(4)arene.alpha,alpha,alpha-trifluorotoluene. AB - The supramolecular 1:1 host-guest inclusion compound, p-tert-butylcalix[4]arene x alpha,alpha,alpha-trifluorotoluene, 1, is characterized by 19F and 13C solid state NMR spectroscopy. Whereas the 13C NMR spectra are easily interpreted in the context of earlier work on similar host-guest compounds, the 15F NMR spectra of solid 1 are, initially, more difficult to understand. The 19F[1H] NMR spectrum obtained under cross-polarization and magic-angle spinning conditions shows a single isotropic resonance with a significant spinning sideband manifold. The static 19F[1H] CP NMR spectrum consists of a powder pattern dominated by the contributions of the anisotropic chemical shift and the homonuclear dipolar interactions. The 19F MREV-8 experiment, which minimizes the 19F-19F dipolar contribution, helps to identify the chemical shift contribution as an axial lineshape. The full static 19F[1H] CP NMR spectrum is analysed using subspectral analysis and subsequently simulated as a function of the 19F-19F internuclear distance (D(FF) = 2.25 +/- 0.01 A) of the rapidly rotating CF3 group without including contributions from additional libration motions and the anisotropy in the scalar tensor. The shielding span is found to be 56 ppm. The width of the centerband in the 19F[1H] sample-spinning CP NMR spectrum is very sensitive to the angle between the rotor and the magnetic field. Compound 1 is thus an attractive standard for setting the magic angle for NMR probes containing a fluorine channel with a proton-decoupling facility. PMID- 11270739 TI - Dynamic carbon-13 MAS NMR: application to benzene ring flips in polyaryl ethers. AB - Carbon-13 dynamic MAS NMR is used to determine the pi-flip rates of the phenyl rings in the low-molecular-weight members of the polyaryl ethers series (phenyl O(-phenylene-O)n-phenyl). The first member in the series (diphenyl ether, n = 0) does not undergo measurable dynamic processes up to its melting point (28 degrees C). The second and third members (n = 1 and 2) exhibit, above room temperature, line broadening effects due to fast pi-flips of the terminal rings, while the spectra of the n = 1 homologue also exhibit line broadening for the inner phenylene ring. Kinetic parameters for the various pi-flip processes were derived by a detailed lineshape analysis of the MAS spectra. The measurements were extended to lower temperatures by time-reverse ODESSA experiments. The kinetic parameters derived from these experiments are, k(t)(300 K) = 31 s(-1), E(t) = 84 kJ/mol, and k(i)(300 K) = 1.3 s(-1), Ei = 77 kJ/mol for the n = 1 homologue and k(t)(300 K) = 3.2 s(-1), E(i) = 78 kJ/mol, for the n = 2 homologue, where the subscripts t and i refer to the terminal and inner benzene rings, respectively. For the simulation of the dynamic MAS spectra the Floquet expansion method was used. In an introductory chapter the Floquet method is reviewed with emphasis on the practical aspects of the computation procedure, on the sensitivity of the results to the isotropic and anisotropic chemical shift parameters, and on the form of the results in the limiting fast and slow exchange regimes. PMID- 11270741 TI - 27Al NMR study in UNiAl. AB - The ternary compound UNiAl exhibiting an antiferromagnetic order below T(N) = 19.3 K has been studied in the paramagnetic state using the 27Al NMR technique and different magnetically oriented samples. The quadrupole coupling constant e2qQ/h = 1.56 MHz is temperature independent. The dominant, longitudinal component of the Knight shift with respect to the hexagonal c axis, Kparallel, is positive and increases upon lowering the temperature down to 50 K. Much smaller in magnitude, the transverse component, Kperpendicular, is also positive and only slightly temperature dependent. The plots of the Knight shift vs magnetic susceptibility Kparallel(chi parallel) and Kperpendicular (chi perpendicular) form the same line, which implies that the transferred hyperfine field of 9.2 kOe/microB for 27Al nuclei should be considered isotropic. PMID- 11270742 TI - 27Al NMR study in ZrNiAl. AB - We have studied the microscopic properties of the hexagonal ZrNiAl, a model compound for a wide family of intermetallic compounds crystallizing in this type of structure, by using 27Al NMR spectroscopy. We have investigated the lineshape of static and MAS NMR spectra as a function of magnetic field strength (4.7-9.4 T) and temperature (5-300 K). Our data indicate that the 27Al NMR spectra result from a combined effect of quadrupole and anisotropic shift interactions. The 27Al nuclei are in an environment characterized by the quadrupole coupling constant e2qQ/h of 3.3 MHz, asymmetry parameter etaQ of 0.42, isotropic shift delta(iso) of 393 ppm, shift anisotropy delta(anis) = delta(zz) - (delta(xx) + delta(yy))/2 of 150 ppm, and asymmetry factor etaS of 0.5. They are found to be temperature independent. The spin-lattice relaxation rate measured at 7.05 T is proportional to the temperature with T1T = 135 s K. The mechanisms responsible for observed values of delta(iso), delta(anis), T1T, and the enhanced Korringa constant are discussed. PMID- 11270743 TI - Influence of sodium ion dynamics on the 23Na quadrupolar interaction in sodalite: a high-temperature 23Na MAS NMR study. AB - High-temperature 33Na MAS NMR experiments up to 873 K for a number of different sodalites (Na8[AlSiO4]6(NO3)2, Na8[AlSiO4]6(NO2)2, Na8[AlSiO4]6I2, Na7.9[AlSiO4]6(SCN)7.9 x 0.5H2O, Na8[AlGeO4]6(NO3)2, and Na7[AlSiO4]6(H3O2) x 4H2O) were carried out. The spectra of the first five sodalites consist of a quadrupolar MAS pattern with different quadrupolar coupling constants. The quadrupolar interaction for the thiocyanate sodalite, the nitrate aluminosilicate, and germanate sodalite decreases strongly passing a coalescence state on heating, while the quadrupolar interaction of the iodide and nitrite sample shows nearly no change. The basic hydrosodalite shows an asymmetric lineshape at room temperature and, between 350 and 370 K, a second line due to the evaporation of cage-water emerges. The linewidth increases with rising temperature. The temperature dependence of the quadrupolar interaction seems to be a function of the sodalite beta-cage expansion. Two conceivable jump mechanisms are proposed for a tetrahedral two-site jump between occupied and unoccupied tetrahedral sites. PMID- 11270744 TI - Molecular dynamics in solid riboflavin as studied by 1H NMR. AB - Spin-lattice relaxation times Tl and Tld as well as NMR second moment were employed to study the molecular dynamics of riboflavin (vitamin B2) in the temperature range 55-350 K. The broad and flat Tl minimum observed at low temperatures is attributed to the motion of two nonequivalent methyl groups. The motion of the methyl groups is interpreted in terms of Haupt's theory, which takes into account the tunneling assisted relaxation. An additional mechanism of relaxation in the high temperature region is provided by the motion of a proton in one of the hydroxyl groups. The Davidson-Cole distribution of correlation times for this motion is assumed. PMID- 11270745 TI - A multinuclear magnetic resonance study of intramolecular hydrogen bonding in some Schiff and Schiff-Mannich bases in solution and in the solid state. AB - Two Schiff bases, N,N'-bis(5-bromosalicylidene)-1,2-diaminoethane, BS, and 7-[(1 [5-bromo-2-hydroxyphenyl] methylidene)amino]-4-methylcoumarin, Sc, and two appropriate Schiff-Mannich bases, N,N'-bis[5-bromo-3 [(diethylamino)methyl]salicylidene]-1,2-diaminoethane, BSM, and 7-[(1-[5-bromo-3 [(diethylamino)methyl]-2-hydroxyphenyl] methylidene)amino]-4-methylcoumarin, SMc, capable of intramolecular hydrogen bonding have been investigated by multinuclear magnetic resonance methods in both solid and liquid phases. In all of the compounds under investigation tautomeric equilibrium involving an intramolecular hydrogen bond has been found. The Schiff-Mannich bases, which can form two different kinds of H bonds at room temperature, form relatively weak H bonds with the imino nitrogen atoms. At low temperatures the tautomeric proton exchange becomes slow on the NMR time scale and both hydrogen-bonded forms can be observed by 1H, 13C, and 15N NMR methods. In the solid state the tautomeric process is frozen and only one H-bonded form is present. On the basis of 13C and 15N CPMAS NMR spectra this is identified as the form with hydrogen bonds involving the imino groups. This conclusion is in good agreement with previous results obtained by X-ray diffraction methods. The investigated Schiff bases (BS and Sc) form relatively weak H bonds. The proton position in the hydrogen bridge, estimated from 15N and 13C chemical shifts, is very similar in both the solution and solid phases. In chloroform solution the observed tautomeric equilibria are almost insensitive to a temperature change within the range 223 to 303 K. PMID- 11270746 TI - Analytic solutions to compartmental models of the HIV/AIDS epidemic. AB - For many years compartmental models have provided useful insights into the spread of epidemics. Such models are usually fairly easy to set up, but even the simpler models have the disadvantage that they are intractable to analytic solution. In this paper we examine models of the HIV/AIDS epidemic, and show that the equations may be linearized in a piecewise manner over time, thus allowing analytic solutions to be obtained. Indications of the usefulness of this approach are provided. In particular, an analytic solution gives insight into the mechanism of the epidemic, together with a clearer picture of the sensitivity of results to changes in parameter values. Further, the processes of parameter estimation and the methodology of back-calculation also benefit from the provision of functional forms for the state variables. PMID- 11270747 TI - Nature of equilibria and effects of drug treatments in some simple viral population dynamical models. AB - We examine some simple mathematical models which have been recently employed to predict the evolution of population dynamical systems involving virus particles. They include: (1) A general two-component antibody-viral system; (2) A simplified two-component model for HIV-1 dynamics (3) An HIV-1 three-component model including virions and (4) A four-component HIV-1 dynamical model which includes both latently and actively infected cells. For each system we find equilibrium points and analyse their local stability properties in order to obtain a global phase portrait. Analytical methods are complemented with numerical solutions. In all four models there are at most two equilibrium points for physically meaningful values of the variables. As the viral growth rate parameter increases through a critical value, a transcritical bifurcation occurs. One critical point (P1) is always found at zero viral or infected cell levels and non-zero antibody or uninfected cell levels. For parameter values in their usual ranges, P1 is either an asymptotically stable node or a saddle point. When the critical point P2 occurs at biologically meaningful values, it is either an asymptotically stable node or an asymptotically stable spiral point. For all three HIV-1 models, the values of the parameters at which P2 makes a transition to physically meaningful values are precisely those at which P1 changes from an asymptotically stable node to an unstable saddle point. The global pictures for all four models are similar and examples are represented graphically. No limitcycle solutions were found in any of the models for parameter values in their usual ranges. In the four-component HIV-1 model, the effects of varying each parameter are found and conditions under which P2 changes from spiral point to node are investigated numerically. The effects of reverse transcriptase inhibitors and protease inhibitors, two classes of drugs used to treat HIV-1 infection, are examined in the three-component model for early HIV-1 dynamics. PMID- 11270748 TI - Modelling the growth of soil-borne fungi in response to carbon and nitrogen. AB - Growth of soil-borne fungi is poorly described and understood, largely because non-destructive observations on hyphae in soil are difficult to make. Mathematical modelling can help in the understanding of fungal growth. Except for a model by Paustian & Sch urer (1987a), fungal growth models do not consider carbon and nitrogen contents of the supplied substrate, although these nutrients have considerable effects on hyphal extension in soil. We introduce a fungal growth model in relation to soil organic matter decomposition dealing with the detailed dynamics of carbon and nitrogen. Substrate with a certain carbon : nitrogen ratio is supplied at a constant rate, broken down and then taken up by fungal mycelium. The nutrients are first stored internally in metabolic pools and then incorporated into structural fungal biomass. Standard mathematical procedures were used to obtain overall-steady states of the variables (implicitly from a cubic equation) and the conditions for existence. Numerical computations for a wide range of parameter combinations show that at most one solution for the steady state is biologically meaningful, specified by the conditions for existence. These conditions specify a constraint, namely that the 'energy' (in terms of carbon) invested in breakdown of substrate should be less than the 'energy' resulting from breakdown of substrate, leading to a positive carbon balance. The biological interpretation of the conditions for existence is that for growth the 'energy' necessary for production of structural fungal biomass and for maintenance should be less than the mentioned positive carbon balance in the situation where all substrate is colonized. In summary, the analysis of this complicated fungal growth model gave results with a clear biological interpretation. PMID- 11270749 TI - Analysis of a multiple equivalent cylinder model with generalized taper. AB - A multiple equivalent cylinder somatic shunt cable model for passive neurones is considered in which one or more tapering equivalent cylinders emanate from a uniformly polarized soma. Each tapering equivalent cylinder approximates the loss of dendritic trunk parameter in the one or more dendritic trees that it represents, relaxing certain symmetry conditions necessary for the Rall equivalent cylinder reduction and in particular allowing terminal branches to end at different electrotonic distances from the soma. The types of taper belong to a class of six specific forms which can account for strict taper, strict flare as well as tapering and flaring combinations. As such they can account for a wide variety of taper forms that arise in practice upon collapse of a dendritic tree to an appropriate equivalent cylinder involving taper. The uniform (i.e. non taper) and exponential taper forms are two of the six taper types. PMID- 11270750 TI - Biological implications of a discrete mathematical model for collagen deposition and alignment in dermal wound repair. AB - We develop a novel mathematical model for collagen deposition and alignment during dermal wound healing. We focus on the interactions between fibroblasts, modelled as discrete entities, and a continuous extracellular matrix composed of collagen and a fibrin based blood clot. There are four basic interactions assumed in the model: fibroblasts orient the collagen matrix, fibroblasts produce and degrade collagen and fibrin and the matrix directs the fibroblasts and determines the speed of the cells. Several factors which influence the alignment of collagen are examined and related to current anti-scarring therapies using transforming growth factor beta. The most influential of these factors are cell speed and, more importantly for wound healing, the influx of fibroblasts from surrounding tissue. PMID- 11270751 TI - Lotka-Volterra equations with chemotaxis: walls, barriers and travelling waves. AB - In this paper we consider a simple two species model for the growth of new blood vessels. The model is based upon the Lotka-Volterra system of predator and prey interaction, where we identify newly developed capillary tips as the predator species and a chemoattractant which directs their motion as the prey. We extend the Lotka-Volterra system to include a one-dimensional spatial dependence, by allowing the predators to migrate in a manner modelled on the phenomenon of chemotaxis. A feature of this model is its potential to support travelling wave solutions. We emphasize that in order to determine the existence of such travelling waves it is essential that the global relationships of a number of phase plane features other than the equilibria be investigated. PMID- 11270752 TI - The emerging European epidemic of variant Creutzfeldt-Jakob disease and bovine spongiform encephalopathy: lessons for Australia. PMID- 11270753 TI - Iron deficiency in children: food for thought. PMID- 11270754 TI - Measuring myocardial damage. PMID- 11270756 TI - Measurement of cardiac troponin I levels in the emergency department: predictive value for cardiac and all-cause mortality. AB - OBJECTIVE: To assess the predictive value of cardiac troponin I levels in cardiac and all-cause mortality in patients presenting to an emergency department. DESIGN: A prospective cohort study. SETTING: The emergency department of a major tertiary teaching hospital in metropolitan Melbourne over a six-week period in 1998. PATIENTS: All patients with requests for cardiac enzyme level measurement. MAIN OUTCOME MEASURES: Cardiac and all-cause mortality within 30 days of presentation. RESULTS: 424 patients (232 men, 192 women; age range, 16-93 years) were reviewed. The 30-day mortality rate was 7.3% (31/424); in patients with raised levels of both creatine kinase (CK)-MB isoenzyme and troponin I this rate was 27% (7/26; 95% CI, 13%-44%); and in those with troponin I levels above 2 microg/L, but normal CK-MB values, it was 24% (5/21; 95% CI, 5%-43%). The mortality rate in the group with normal results of cardiac markers was 4.3% (14/328; 95% CI, 2.1%-6.5%). Patients with minor increases in troponin I levels (minimal myocardial damage) showed an intermediate 30-day mortality rate (13%, 5/39; 95% CI, 2%-24%). Other predictors of 30-day mortality included age, presentation with shortness of breath, and electrocardiography (ECG) changes diagnostic of acute myocardial infarction or consistent with ischaemia. Cardiovascular causes were responsible for most of the deaths in patients with raised troponin I levels. Multivariate logistic regression analysis showed that raised levels of troponin (> 2.0 microg/L), but not of CK-MB, predict 30-day mortality rate. CONCLUSIONS: Compared with CK-MB, cardiac troponin I more accurately predicts 30-day mortality rates in patients presenting to the emergency department. Moreover, troponin I levels identify additional groups of patients at increased risk of death not so identified by measuring CK-MB values. PMID- 11270755 TI - Iron deficiency in Australian-born children of Arabic background in central Sydney. AB - OBJECTIVES: To determine the prevalence of iron depletion and deficiency, and iron-deficiency anaemia, along with risk factors for iron depletion, in Australian-born children aged 12-36 months of Arabic-speaking background. DESIGN: Community-based survey. SETTING: Central Sydney Area Health Service (CSAHS), NSW, April to August, 1997. PARTICIPANTS: All children born at five Sydney hospitals between 1 May 1994 and 30 April 1996, whose mothers gave an Arabic-speaking country of birth and resided in the area served by the CSAHS. MAIN OUTCOME MEASURES: Full blood count (haemoglobin, mean corpuscular haemoglobin, mean corpuscular volume), plasma ferritin concentration, haemoglobin electrophoresis, potential risk factors for iron depletion. RESULTS: Families of 641 of the 1,161 eligible children were able to be contacted, and 403 agreed to testing (response rate, 62.9% among those contacted). Overall, 6% of children had iron-deficiency anaemia, another 9% were iron deficient without anaemia, and 23% were iron depleted. Multiple logistic regression analysis showed three significant independent risk factors for iron depletion: <37 weeks' gestation (odds ratio [OR], 5.88, P=0.001); mother resident in Australia for less than the median time of 8.5 years (OR, 1.96, P=0.016); and daily intake of >600 mL cows' milk (OR, 3.89, P=<0.001). CONCLUSION: Impaired iron status is common among children of Arabic background, and targeted screening is recommended for this group. PMID- 11270757 TI - Diagnostic cervical zygapophyseal joint blocks for chronic cervical pain. AB - OBJECTIVES: (1) To determine the prevalence of cervical zygapophyseal joint pain in a specialist clinical setting; (2) to review the number of diagnostic blocks needed to identify the segmental level of the symptomatic joints; and (3) to determine the distribution of segmental levels of cervical zygapophyseal joint pain in a clinical setting. DESIGN AND SETTING: Retrospective audit of patients of three independent rehabilitation medicine specialists who had undergone cervical zygapophyseal joint blocks in hospital outpatient clinics and private rooms. PATIENTS: 97 patients aged 18-82 years with chronic neck pain (with or without headache) of more than six months' duration refractory to conservative therapies. INTERVENTION: Diagnostic fluoroscopic cervical third occipital and medial branch blocks of zygapophyseal joints. Diagnosis required confirmation by a repeat procedure. RESULTS: 35 of 97 patients (36%) had a confirmed symptomatic cervical zygapophyseal joint (95% CI, 27%-45%). The symptomatic segmental level was found at the first attempt by reference to a standard pain diagram in 83% of cases (29 of 35). The most common symptomatic levels were C3-4 (11/35; 31%) and C5-6 (10/35; 29%). CONCLUSION: The prevalence of cervical zygapophyseal joint pain estimated in this clinical study is lower than that found in previous research setting studies, but our requirement for confirmation by a repeat block (which many patients declined) makes our estimate conservative; it is likely that the true prevalence is higher. Zygapophyseal joints are clearly a common source of pain in patients presenting with chronic neck pain, with or without headache. Cervical zygapophyseal joint pain is readily diagnosable, enabling patients to seek further, targeted treatment. PMID- 11270758 TI - An epidemic of dengue 3 in far north Queensland, 1997-1999. AB - OBJECTIVES: To describe an epidemic of dengue type 3 that occurred in far north Queensland in 1997-1999 and its influence on the further development of dengue prevention and control strategies. DESIGN: Epidemiological and laboratory investigation of cases, entomological surveys and phylogenetic analysis of dengue virus isolates. MAIN OUTCOME MEASURES: Numbers and characteristics of confirmed cases; Breteau Index (BI; number of containers breeding Aedes aegypti per 100 premises); effect of control measures on mosquito populations; genetic homology of epidemic virus with other dengue virus isolates. RESULTS: The epidemic lasted 70 weeks and comprised 498 confirmed cases in three towns (Cairns, Port Douglas and Mossman); 101 patients (20%) were admitted to hospital. Median interval between symptom onset and notification was seven days (range, 0-53 days), and cumulative duration of viraemia of public health significance was 2,072 days. BIs in affected areas were high, particularly in Mossman (45) and Port Douglas (31). Control measures significantly reduced mosquito populations (assessed as number of ovitraps containing Ae. aegypti eggs and mean number of eggs per trap [P< 0.05 for both]). However, transmission persisted in several foci, in part due to undetected waterfilled containers breeding Ae. aegypti. The epidemic virus belonged to serotype 3; phylogenetic analysis suggested it was imported from Thailand. CONCLUSIONS: The epidemic had greater morbidity than other recent Queensland epidemics of dengue and was harder to control, necessitating substantial revision of the Dengue Fever Management Plan for North Queensland. The epidemic's severity supports the hypothesis that dengue viruses from South East Asia are more virulent than others. PMID- 11270759 TI - Acute hepatitis C virus infection in an Australian prison inmate: tattooing as a possible transmission route. AB - Clinically apparent hepatitis C virus (HCV) infection developed in a prison inmate after two tattooing episodes within the recognised incubation period for HCV infection. Seroconversion and HCV viraemia with subsequent resolution of hepatitis and loss of plasma viraemia were documented. Introducing licensed tattooists, and thereby improving infection control practices, may reduce the risk of hepatitis C virus infection in prisons. PMID- 11270761 TI - Understanding wellness in old age. PMID- 11270760 TI - Fibroadenoma of the breast. AB - Fibroadenoma of the breast is a common cause of a benign breast lump in premenopausal women. The consensus view is that women with fibroadenomas are not at significant increased risk of developing breast cancer. Diagnosis is based on the combination of clinical examination, imaging and non-surgical tissue biopsy (the triple test). A clinical diagnosis of fibroadenoma alone is unreliable and does not exclude malignancy even in younger women. The choice of imaging is mammography, combined with ultrasound in older women, and ultrasound alone in younger women. Tissue biopsy, by either fine-needle aspiration or core biopsy, is the most accurate means of establishing the diagnosis. Traditionally, symptomatic fibroadenomas were treated by surgical excision, and this option should always be offered. There is increasing evidence that a conservative approach is safe and acceptable, provided the result of an adequate triple test is both negative for cancer and consistent with a fibroadenoma. Patients who choose conservative management need to be informed of the limitation of the tests, and must be assessed promptly if there is symptomatic or clinical change. PMID- 11270762 TI - Diagnosing inhaled foreign bodies in children. PMID- 11270763 TI - Does drinking carrot juice affect cancer of the prostate? PMID- 11270764 TI - Changes in HIV-1 viral load trends with highly active antiretroviral therapy. PMID- 11270765 TI - Physiotherapists' use of medications and the influence of drug companies. PMID- 11270766 TI - Biopsychosocial profile of adults with intellectual disability. PMID- 11270767 TI - Reducing premature death and renal failure in Australian aboriginals: a community based cardiovascular and renal protective program. PMID- 11270768 TI - Selling clinical trials to the public. PMID- 11270769 TI - Sex bias in management of LUTS in men. PMID- 11270770 TI - Drug testing at the Sydney Olympics. PMID- 11270771 TI - Drug testing at the Sydney Olympic Games. PMID- 11270773 TI - Carcinosarcoma of the colon and spleen: a fleshy purple mass on colonoscopy. PMID- 11270774 TI - Differences in carcinogenesis by the length of carcinogen exposure period in rat colon. AB - To clarify the carcinogenic factors--whether it is the kind of carcinogen or their length of exposure--that determine whether colorectal cancer develops from an adenoma or develops de novo in the absence of an adenoma, we histopathologically analyzed a total of 229 rat colon tumors induced by administration of 1,2-dimethyl-hydrazine (DMH) or N-methyl-N'-nitro-N nitrosoguanidine (MNNG) for three or 15 weeks. In the three-week-exposure groups, 71% of DMH-induced carcinomas and 82% of MNNG-induced carcinomas coexisted with low-grade dysplasia (adenomatous remnant). However, in the 15-week-exposure groups, lowgrade dysplasia was observed in only 10% of DMH-induced and 27% of MNNG-induced carcinomas. Even in the tumors smaller than 20 mm3, it was observed in only 10% of DMH-induced and 32% of MNNG-induced carcinomas. Furthermore, carcinomas without low-grade dysplasia predominated from the initial period of tumor occurrence. Next, we investigated association of K-ras and APC gene mutations with these carcinogenesis patterns in 80 tumors. K-ras mutations were not detected in any tumors induced by three weeks of exposure. However, in the 15 week-exposure groups, this mutation was observed in 57% of DMH-induced tumors and 13% of MNNG-induced tumors. APC mutations in the region homologous to the human mutation cluster region were observed in only 6% of tumors. Thus, our results suggest that the carcinogenesis patterns in rat colon are dependent on the length of exposure to carcinogen and that K-ras mutations were partly involved in a subset of them. PMID- 11270775 TI - Percutaneous liver biopsy: what is the current approach? Results of a questionnaire survey. AB - The technique of performing liver biopsy varies among physicians, even within the same practice. In an attempt to determine whether gastroenteroogists/hepatologists differ in their approach compared to radiologists, we surveyed a nearly equal number of physicians in the Washington, DC, USA, metropolitan area with respect to 26 variables. While the technique can vary considerably, relatively few differences were seen between the groups. Only about half of gastroenterologists and hepatologists use ultrasound guidance, a biopsy, gun, and conscious sedation; radiologists routinely used a biopsy gun and conscious sedation and rarely require an overnight stay. PMID- 11270776 TI - Percutaneous liver biopsy using an ultrasound-guided subcostal route. AB - Percutaneous biopsy is considered one of the most important diagnostic tools to evaluate diffuse liver diseases. The introduction and widespread diffusion of ultrasounds in medical practice has improved percutaneous bioptic technique, while reducing postoperative complications. Although ultrasonography has become almost ubiquitous in prebiopsy investigation, only one third of biopsies are performed under ultrasound control. Moreover, the one-day procedure, reported in several studies to be safe and cost effective, accounted for only 4% of biopsies done. We report our experience of 142 percutaneous US-guided biopsies performed on 140 patients affected by chronic diffuse liver disease over a four-year period. Liver biopsies were performed under US guidance at the patient's bed using an anterior subcostal route. We evaluated postoperative pain, modifications of blood pressure and red cell count, hospital stay, morbidity and mortality rates, and adequacy of specimens for histologic examination. There was no operative mortality. As for major complications, one case of hemobilia occurred. As for minor complications, two cases of persistent postoperative pain required analgesic therapy. Patients were discharged the day following the procedure in all cases but two, who were discharged on the third and fifth postoperative days. Liver specimens were suitable for histologic diagnosis in all but one case, in which there were no portal spaces. According to our experience, we believe that hepatic biopsy guided by ultrasonography could replace blinded biopsy in the diagnosis of diffuse liver disease. The procedure is suitable to be performed safely on an outpatient basis. PMID- 11270772 TI - Complications of long-term home total parenteral nutrition: their identification, prevention and treatment. AB - The purpose of this review is to describe the most common complications of home total parenteral nutrition, their identification, treatment and prevention. Data sources were manuscripts and abstracts published in the English literature since 1968. Studies were selected for summarization in this review on the basis of clinical relevance to the practicing clinician. Home total parenteral nutrition is a relatively safe, life-saving method for nutrient delivery in patients with compromised gastrointestinal function. However, numerous complications, with associated morbidity and mortality, involving the delivery system and the gastrointestinal, renal, and skeletal systems may develop. Catheter-related complications are often preventable and treatable when they occur, although renal and bone abnormalities have elusive etiologies. PMID- 11270778 TI - Shared genetic risk factors in autoimmune liver disease. AB - To determine if shared genetic risk factors for autoimmune liver disease affect clinical manifestations, we evaluated 271 patients and 92 normal subjects by DNA based techniques. Genetic risk factors were intermixed in all conditions, and frequency varied according to disease type. DR4 distinguished autoimmune hepatitis (P = 0.0002) and primary biliary cirrhosis (P = 0.004) from primary sclerosing cholangitis. DR52 distinguished primary sclerosing cholangitis from autoimmune hepatitis (P = 0.0007) and primary biliary cirrhosis (P = 0.00007) and DR3 distinguished autoimmune hepatitis (P = 0.002) and primary sclerosing cholangitis (P = 0.0005) from primary biliary cirrhosis. Only the occurrence of DR4 in primary sclerosing cholangitis was lower than in normal subjects (P = 0.02). Patients with mixed genetic risk factors did not have distinctive features or manifestations of hybrid conditions. We conclude that patients with shared genetic risk factors do not have characteristic features nor do they have overlap syndromes. DR4 may be protective against primary sclerosing cholangitis. PMID- 11270777 TI - Influence of low-dose oral contraceptives, alcohol, and grapefruit on. AB - The aminopryine breath test (ABT) measures hepatic reserve in patients with acute and chronic liver disease and gives an assesment of the hepatic function in patients undergoing major liver surgery. Aminopyrine is metabolized by the mixed cytochrome P-450 system, which can be influenced by many foreign compounds and drugs. Whether these foreign compounds and drugs can influence the results of the ABT has seldomly been tested. We studied three groups: Healthy female volunteers, either normally menstruating or taking oral contraceptives, were asked to perform a [13C]ABT during the time of the menses and at midcylce. Healthy volunteers were asked to perform a ABT after consuming 30 g of alcohol. Healthy volunteers were asked to perform a ABT after consuming 250 ml of grapefruit juice. The 13C/12C ratio in expired air was measured by gas isotope ratio mass spectrometry. PMID- 11270779 TI - Evaluation of regional liver damage by magnetic resonance imaging with superparamagnetic iron oxide in rat liver. AB - The purpose of this study was to investigate whether regional liver damage could be detected by means of enhanced MR imaging with a superparamagnetic iron oxide (SH U 555A) in an ischemia-reperfusion model of rat liver. Ischemic liver damage was induced in the right lobe by vascular clamping for 0 (sham), 30 (I-30), 60 (I 60), and 90 minutes (I-90). There was no significant difference in relative enhancement (RE) between the ischemic and nonischemic lobes in the sham, I-30 and I-60 groups, while RE of the ischemic lobe was significantly lower than that of its nonischemic counterpart in the I-90 group as seen on SH U 555A enhanced proton density spin echo images (P < 0.05). Histological examination revealed that iron deposits were significantly smaller in the ischemic than the nonischemic lobe in the I-90 group (P < 0.05), although there was no significant difference in the number of Kupffer cells. Our results indicate that severe regional liver damage can be evaluated by MR imaging with SH U 555A. PMID- 11270780 TI - PYY potently inhibits pancreatic exocrine secretion mediated through CCK-secretin stimulated pathways but not 2-DG-stimulated pathways in awake rats. AB - Peptide YY (PYY) is an important modulator of stimulated pancreatic exocrine secretion. PYY acts proximal to the acinar cell but the exact site and mechanism of action are unknown. The aim of the present study is to determine the pathway through which PYY exerts its effect on the exocrine pancreas in awake rats under physiological condition. When pancreatic secretion was stimulated by graded doses of cholecystokinin (CCK) (14, 28, 58 pmol/kg/hr) with secretin (1.25, 2.5, 5.0 pmol/kg/hr) or CCK alone at 28 pmol/kg/hr, PYY1-36 dose-dependently inhibited pancreatic secretory responses. Moreover, PYY1-36 at 50 pmol/kg/hr almost completely blocked the stimulation by CCK (P < 0.01). Although background infusion of PYY1-36 or PYY3-36 at 12.5 pmol/kg/hr inhibited basal pancreatic fluid and protein secretion, but both of them only partly inhibited the subsequent 2-DG stimulated pancreatic fluid and protein secretion. Furthermore, PYY1-36 at 50 pmol/kg/hr failed to inhibit 2-DG-stimulated pancreatic secretion. These results confirm that PYY1-36 inhibits CCK-stimulated pancreatic secretion under all experimental conditions. However, in the awake, surgically recovered rat, PYY1-36 at both low and high doses failed to fully inhibit 2-DG-stimulated pancreatic secretion. Therefore, the site of PYY's inhibitory action on pancreatic secretion appears to be primarily on the CCK-stimulated pathway at a site proximal to the convergence of the CCK and 2-DG pathways. PMID- 11270781 TI - Apoptosis in rat spontaneous chronic pancreatis: role of the Fas and Fas ligand system. AB - The Fas/Fas ligand (FasL) system is suggested to be correlated to the onset of inflammation and apoptosis in various diseases. However, whether Fas and FasL are expressed in chronic pancreatitis is unknown. The aim of this study was to examine the expression of the Fas/FasL system and to analyze its correlation with apoptosis in a spontaneous chronic pancreatitis model (the WBN/Kob rat). Four week-old male WBN/Kob rats were fed a special pellet diet (MB-3). Different groups of rats were killed every four weeks, and pancreata were histopathologically examined. Fas and FasL mRNAs in the pancreas were detected with a reverse transcription-polymerase chain reaction method. The cellular localization of Fas and FasL mRNA and protein was determined with in situ hybridization (ISH) and immunohistochemistry (IHC). Apoptosis was detected with a terminal deoxynucleotidyltransferase-mediated method. Fas and FasL mRNA were expressed when the pancreas was still pathologically normal, and showed a biphasic peak at 12 and 20 weeks. ISH and IHC confirmed that Fas and FasL are expressed in the cytoplasm of acinar cells, ductal cells, and lymphocytes. An apoptotic index in acinar cells correlated to the expression of Fas and FasL mRNAs. These results suggest that the expression of the Fas/FasL system is involved in acinar cell apoptosis and the onset and progression of chronic pancreatitis in the WBN/Kob rat. PMID- 11270782 TI - Myosin light chain phosphorylation correlates with contractile force in guinea pig gallbladder muscle. AB - Acetylcholine (ACh)-induced gallbladder smooth muscle contraction involves myosin light chain phosphorylation (MLCP). ACh-induced contraction is dose dependent. Whether MLCP by ACh is also dose dependent and how it correlates with contractile force have not been carefully evaluated. This study investigated the correlation between gallbladder muscle contraction and MLCP. Guinea pig gallbladder muscle strips were studied isometrically and frozen after different doses of ACh (0, 0.1, 5, 100 microM) for different periods of incubation (0.5, 1, 2, 3, 4 min). MLCP was determined using gel electrophoresis. Both contraction and MLCP in response to ACh were concentration dependent. Peak MLCP to ACh 100 microM occurred at 30 sec. There was a high correlation between active force and MLCP (r = 0.991; P = 0.009 at 30 sec stimulation). Nifedipine 1 microM reduced ACh induced contraction and MLCP by a similar degree (31% and 33%, respectively). In conclusion, gallbladder contractile force significantly correlates with MLCP. This is consistent with the hypothesis that initiation of gallbladder cholinergic contraction is dependent on phosphorylation of myosin light chains. PMID- 11270783 TI - Direct effect of thyroxine on pig sphincter of Oddi contractility. AB - Sphincter of Oddi (SO) motility has an important role in the regulation of bile flow. SO function disturbances (stenosis or dyskinesia) may prevent normal bile flow and thus enhance the probability of common bile duct (CBD) stone formation. Previously we have shown that there is an increased prevalence of diagnosed hypothyroidism in CBD stone patients, compared with gallbladder stone patients or age-, sex-, and hospital-admission-adjusted controls. The present study was done to test the hypothesis that thyroxine directly effects the SO. The specificity of the effects of thyroxine were studied by comparing with triiodothyronine (T3), progesterone, cortisone, estrogen, and testosterone. For ex vivo studies three or four successive 1 to 1.5-mm SO rings were prepared from each pig and placed between two hooks in oxygenated physiologic salt solution at 37 degrees C. SO contraction was measured with isometric force displacement transducers and registered on a polygraph. Each SO ring was stimulated with KCl (125 mM), acetylcholine (ACh; 10 or 100 microM) and histamine (Hist; 10 or 100 microM) with and without thyroxine (10(-10) or 10(-8) M), T3 (10(-9) or 10(-7) M), progesterone (1 microM), cortisone (1 microM), estrogen (1 microM), or testosterone (1 nM) in the medium. KCI, ACh, and Hist induced strong contractions in the SO rings. The addition of thyroxine did not influence significantly the KCl-induced contractions, but the ACh- and Hist-induced contractions decreased by a mean of 37-44% (P < 0.001) and 54-56% (P < 0.001), respectively, as compared to the contractions without thyroxine. Triiodothyronine had a similar inhibitory effect to thyroxine, whereas cortisone, estrogen, and testosterone had no effect. Progesterone decreased the KCl-, ACh-, and Hist-induced SO contractions. In conclusion, physiological concentrations of thyroxine have an inhibitory effect on receptor-mediated ACh and Hist, but not on the nonspecific KCl-induced SO contraction ex vivo. The inhibitory effect is similar in thyroxine and triiodothyronine. Of the steroid hormones, only progesterone nonspecifically ameliorates SO contractions ex vivo. Because the effect of thyroxine on the SO is prorelaxing, the lack of thyroxine may result in an increased tension of the SO. PMID- 11270784 TI - Autoimmune pancreatitis detected as a mass in the head of the pancreas with contiguous fibrosis around the superior mesenteric artery. PMID- 11270785 TI - Intestinal transit in dogs is accelerated by volume distension during fat-induced jejunal brake. AB - Intestinal transit is accelerated by volume distension and slowed by nutrient load. We hypothesized that the accelerating effect of volume distension might overcome the slowing effect of nutrient. To test this hypothesis, we compared intestinal transit in five dogs equipped with duodenal and mid-intestinal fistulas. The proximal half of the small intestine was perfused with 60 mM oleate, while the distal half of the small intestine was either perfused with buffer (with distension) or left unperfused (without distension). We found that intestinal transit was slowed by oleate (with marker recovery reduced from 85.8 +/- 5.3 to 39.4 +/- 7.5%) (P < 0.01) and that volume distension accelerated intestinal transit so that marker recovery increased from 39.4 +/- 7.5 without distension to 60.8 +/- 6.3% with distension (P < 0.05). We concluded that intestinal transit is accelerated by volume distension during fat-induced jejunal brake. PMID- 11270786 TI - Mechanisms involved in enteropathy induced by administration of nonsteroidal antiinflammatory drugs (NSAIDS). AB - Mice received oral indomethacin (1 mg/mouse) daily for five days. It was found that severe gastroenteropathy (ie, paralytic stomach and necrotic intestine) was induced on the sixth day. Ulcer formation was also seen at many sites in the digestive tract, especially in the colon. In parallel with the increase in the number of leukocytes in the digestive tract, the proportion of granulocytes increased at various sites, for example, in the intraepithelium and lamina propria of the colon and the lamina propria of the appendix. The number of extrathymic T cells at these sites in the digestive tract, especially gammadelta T cells in the colon, increased. A functional assay revealed that granulocytes isolated from mice injected with indomethacin were activated in terms of their superoxide production upon stimulation. In conjunction with the data on the simultaneous activation of granulocytes in the liver and blood, the present results suggest that nonsteroidal antiinflammatory drugs (NSAIDs) have the potential to induce severe granulocytosis in specific sites of the body, possibly via their stimulatory effect on the sympathetic nervous system (ie, granulocytes bear adrenergic receptors on their surface). PMID- 11270787 TI - Effect of diet on changes in small intestinal blood flow following intracolonic administration of indomethacin to rats. AB - Liquid diet (LD) is known to be protective against indomethacin-induced enteropathy, which is thought to be associated with ischemic change. We tested the hypothesis that the solid component of diet modulates small intestinal blood flow (SIBF) following indomethacin administration. In the first experiment, male Wistar rats were divided into 18-hr-fasted and normal diet groups. Indomethacin (20 mg/kg) or vehicle was administered intracolonically. SIBF was measured on both the mesenteric and antimesenteric sides of the intestine, using the hydrogen gas clearance method. In the second experiment, rats were given LD alone or LD with increasing concentration of soluble/insoluble fiber for seven days. The baseline SIBF was significantly higher in the groups with normal diet and LD with fiber than in the fasting and LD groups. Following indomethacin administration, SIBF gradually decreased in the groups with normal diet and LD with insoluble fiber, while neither liquid diet nor fasting reduced SIBF. There was no difference in SIBF between the mesenteric and antimesenteric sides of the intestine in any group. Our findings suggest that solid components of diet increase basal SIBF and decrease SIBF following indomethacin administration. PMID- 11270788 TI - Intestinal permeability of x-ray constrast media iodixanol and iohexol during bacterial overgrowth of small intestines in rats. AB - To investigate the recovery of iodinated water-soluble contrast medium from small bowel with small morphological alterations, iohexol or iodixanol was instilled through an orogastric tube in rats 14 days after surgery that established a self filling blind loop in the jejunum. This rat model induced small bowel bacterial overgrowth with only minor abnormalities observed on histology and scanning electron microscopy. Animals with end-to-end anastomosis of the jejunum or unoperated rats served as controls. Compared with unoperated animals, urinary recovery of iohexol and iodixanol was significantly higher in both groups that underwent surgery. Moreover, the contrast medium recovery was numerically higher in the self-filling blind loop group given iodixanol than in the end-to-end anastomosis group, although not statistically significant, P = 0.09. Our results indicate that iohexol and iodixanol may detect small barrier impairments in the intestines. Iodixanol, the largest of the two, may seem to differentiate better between normal and minimally impaired intestinal barrier. PMID- 11270789 TI - Anti-endomysial antibody negative celiac disease: does additional serological testing help? AB - Anti-endomysium antibodies (AEM) fail to identify all untreated celiac disease (CD) patients. This study aims to determine if additional serology, in particular, IgA anti-tissue transglutaminase (tTG) antibodies, increases detection. Fifty-three biopsy-proven untreated CD patients (39 women, 14 men; median age 51 years) and 65 control patients with normal duodenal histology (46 women, 19 men; age range 17-90 years, median 45 years) were prospectively studied. Serum total IgA, IgA anti-tTG, IgA AEM, IgA anti-gliadin (AGA) and IgG AGA antibodies were measured. Thirteen (25%) CD patients were AEM negative. None were IgA deficient. Three AEM-negative CD patients had a raised IgA anti-tTG and IgA AGA. IgG AGA was raised in 10 AEM-negative CD patients, but also in 14/65 (22%) of controls. In conclusion, AEM-negative CD is common and detection is only modestly enhanced by testing for IgA anti-tTG antibodies. Duodenal biopsy is still recommended for the accurate diagnosis of CD. PMID- 11270790 TI - Extending gastric emptying scintigraphy from two to four hours detects more patients with gastroparesis. AB - Gastric emptying scintigraphy (GES) is usually performed for up to 2 hr to measure the gastric emptying (GE) of solids. Symptomatic patients, however, may have borderline results at 2 hr, making it difficult to determine whether a gastric motor disorder is present. The aim of this study was to assess whether extending GES to 4 hr is useful in evaluating patients for gastroparesis and to correlate the results of GES with patient symptoms. We studied 129 patients undergoing GES at Temple University Hospital between July 1998 and March 1999. Solid-phase GE was measured at 0, 0.5, 1, 2, 3, and 4 hr after ingestion of a 99mTc sulfur colloid-labeled egg meal. Dyspeptic symptoms of upper abdominal discomfort, early satiety, postprandial abdominal bloating, nausea, vomiting, and anorexia were graded as none, mild, moderate and severe (0, 1, 2 and 3, respectively) with the sum representing a total symptom score. Of 129 patients, 86 had normal GE at 2 hr; 26 of the 86 normal scans at 2 hr were delayed at 3 hr. Six of the 60 scans normal at 2 and 3 hr were delayed at 4 hr. Of 43 patients with delayed GE at 2 hr, 39 were delayed at 3 hr and 35 were delayed at 4 hr. Overall, the percentage of patients with delayed GE increased from 33% at 2 hr only to 58% using the results of the 2-, 3-, and 4-hr scans (P < 0.05). There was a significantly greater symptom score in patients with delayed GE compared to patients with normal GE (8.4 +/- 0.5 vs 7.1 +/- 0.5; P < 0.05). Conclusion, prolonging GES after ingestion of a 99mTc-labeled egg meal from 2 to 4 hr increased the number of symptomatic patients found to have delayed GE. These results suggest that GES should be performed for up to 4 hrs when the 2-hr result is normal. PMID- 11270791 TI - Effect of peroxynitrite on motor function of the opossum esophagus. AB - Nitric oxide (NO*) is a mediator of esophageal motility. Esophageal dysmotility accompanies esophagitis. During inflammation, superoxide and NO* form peroxynitrite (ONOO-), a reactive molecule that alters cellular function. We tested the hypotheses that ONOO- affects esophageal motility and is produced in association with esophagitis. Transverse muscle strips from the opossum esophagus were stimulated by an electrical field, and nitrotyrosine immunoblots were performed. Peroxynitrite, its decomposed form, or NaNO2 relaxed the lower esophageal sphincter (LES) and attenuated the off response. These effects were inhibited by oxyhemoglobin (Hgb). An antagonist of guanylate cyclase, 1H[1,2,4]oxadiazole[4,3]quinoxalin-1-one (ODQ), inhibited the LES relaxation produced by ONOO-. Nitrotyrosine, a footprint for ONOO- production, was detected in inflamed esophagus. These studies support the hypotheses that ONOO alters esophageal motor function and is formed in association with esophagitis. It is possible that some of the esophageal motor dysfunction seen with esophagitis may be related to the formation of ONOO-. PMID- 11270792 TI - Effect of intraduodenal and intravenous amino acids on proximal gastric motor function in man. AB - The present study was performed to investigate the effect of amino acids during the intestinal and postabsorptive phase of digestion on proximal gastric motor function measured with an electronic barostat. Eight healthy volunteers participated in three experiments performed during continuous infusion of: (1) intravenous and intraduodenal saline, (2) intraduodenal amino acids, and (3) intravenous amino acids. Both intraduodenal and intravenous amino acids induced gastric relaxation and increased gastric compliance. Only during intraduodenal amino acids did plasma CCK levels increase significantly. Correlation between intragastric volume measurements (with pressure set at MDP + 2 mm Hg) and plasma CCK levels was 0.90 (P < 0.001) during the early intestinal phase. Relaxation of the proximal stomach is related to plasma CCK in the early intestinal phase, whereas in the postabsorptive phase of amino acids other mechanisms play a role in proximal gastric relaxation. PMID- 11270793 TI - Effect of Helicobacter pylori infection on gastric emptying and gastrointestinal hormones in dyspeptic and healthy subjects. AB - There is no general agreement as regards the effect of Helicobacter pylori infection on gastric emptying in patients with functional dyspepsia. Food releases several gastrointestinal hormones, and some of these are known to contribute to the regulation of gastric emptying. The aim of this study was to investigate the influence of H. pylori on gastric emptying in dyspeptic and healthy subjects and to verify whether different hormone secretion patterns are affected by the presence of the bacterium. Twenty-seven patients affected by functional dyspepsia and 30 asymptomatic healthy subjects entered the study. H. pylori presence was assessed in controls by IgG antibodies to H. pylori and [13C] urea breath test, and that in patients by Warthin-Starry stain on gastric biopsies. After ingesting a standard solid-liquid meal, an ultrasound examination of gastric emptying was performed. Plasma concentrations of gastrin, cholecystokinin, and pancreatic polypeptide were measured in the fasting and postprandial period for 4 hours. The incidence of H. pylori infection was not higher in functional dyspepsia patients than in controls. As regards gastric emptying, no difference was detected between patients and controls with and without H. pylori infection. On the contrary, the presence of H. pylori infection determined alterations in gastrin levels, which were higher in controls than in patients. Basal CCK levels were higher in the H. pylori-negative patients than H. pylori-positive patients and controls. In conclusion, H. pylori infection seems not to cause alterations in gastric emptying, but rather alterations in gastrin levels. In contrast, the altered levels of CCK account for its involvement in the pathophysiology of H. pylori-negative dyspepsia. PMID- 11270794 TI - Effects of genotypically different strains of Helicobacter pylori on human microvascular endothelial cells in vitro. AB - Helicobacter pylori induces a number of disturbances in rodent gastric microcirculation in vivo. These events may result from direct necrotic or apoptotic damage to endothelial cells. This study therefore aimed to investigate the effects of genotypically different H. pylori strains on microvascular endothelial cell (MVEC) viability in vitro. Four H. pylori extracts were prepared from strains with different cagA or vacA status. MVECs were plated into 96-well plates and coincubated with 50 microl of extract or vehicle for 24, 48, 72, or 96 hr. An MPP assay quantified overall MVEC viability. The dual labeling of MVECs with propidium iodide and Hoechst 33342 distinguished between necrotic and apoptotic cell death, respectively, and allowed total number of viable cells to be determined. All strains of H. pylori decreased cell viability after 72 and 96 hr. Neither necrosis or apoptosis was observed. Counting total number of viable cells revealed decreased cell proliferation with all strains when compared to controls, again reaching significance at 72 and 96 hr. In conclusion, both the MTT assay and the diret cell counting technique demonstrated that all H. pylori strains induced cytostatic but not cytotoxic effects on MVECs. This suggests that microcirculatory disturbances observed in vivo may not be the result of direct endothelial cell damage. However, inhibition of angiogenesis may explain why ulcer healing is delayed in H. pylori-infected patients. PMID- 11270795 TI - Transmission of Helicobacter pyori in an animal model. AB - An experimental murine model was studied to evaluate the orogastrointestinal colonization of Helicobacter pylori and the animal-to-animal transmission. Balb/C mice were infected with H. pylori and housed with uninoculated mice in cages with and without a grate on the floor. Mice were killed after 7, 14, 30, and 45 days, and samples from the esophagus, stomach, small intestine, colon, and rectum were analyzed for H. pylori by PCR and immunohistochemistry and for histological changes. Bacterial colonization was assessed also by culture from stomach samples. H. pylori was cultured by stomach samples of infected mice at 7, 14, and 30 days. Using PCR and immunohistochemistry, H. pylori was detected in inoculated and uninoculated mice in all areas examined, with an high percentage of positive samples in the esophagus and stomach. Moreover transmission was detected, without differences, regardless of whether mice were housed with or without a grate on the floor, supporting an orooral animal transmission. PMID- 11270796 TI - Comparative analysis of colonization of Helicobacter pylori and glycolipids receptor density in Mongolian gerbils and mice. AB - The Mongolian gerbil has been used as an excellent experimental animal model for studying Helicobacter pylori infection because it can stably colonize and induce severe chronic gastritis, ulceration, and cancer-simulating human diseases in this animal. In contrast, H. pylori can only induce mild inflammation in many mouse models. The aim in this study is to clarify the difference of induction of pathological lesions in the two animal models. SPF ICR mice and Mongolian gerbils were inoculated with a clinically isolated strain of H. pylori. Six weeks after inoculation, bacteria colonizing the stomach were counted. Immunohistochemical staining and biochemical analyses of three putative receptor glycolipids were performed with monoclonal antibodies to the respective glycolipids. Significantly higher numbers of H. pylori were recovered from the stomachs of Mongolian gerbils than mice (5.77 +/- 0.46 log CFU vs 4.17 +/- 0.55 log CFU, P < 0.01). Immunohistochemical studies showed that sulfatide expression in the gastric mucosa of Mongolian gerbils was much stronger than that in mice, whereas the expression of Lewis(b) glycolipid and GM3 were almost equal. Quantitative analysis of each glycolipid by thin-layer chromatography confirmed the results of immunohistochemical study, showing 4.1 times higher sulfatide content in the Mongolian gerbil stomach. The content of both Lewis(b) and GM3 was almost equivalent in these two animals. In conclusions, higher levels of sulfatide expression, a putative adhesion receptor, in the gastric mucosa of Mongolian gerbils may allow abundant colonization by H. pylori, resulting in the development of gastric lesions in this animal model. PMID- 11270797 TI - Host-bacterial interaction: what role does Helicobacter pylori urease play? PMID- 11270798 TI - Reproducibility and intragastric variation of duodenogastric reflux using ambulatory gastric bilirubin monitoring. AB - Duodenogastric reflux has long been considered to be important in the pathogenesis of many gastric disorders that exhibit regional variation within the stomach. Ambulatory gastric bilirubin monitoring is a new technique and, although extensively validated, reproducibility and gastric regional variation have not been specifically addressed. Fourteen patients with symptoms of gastroesophageal reflux and 12 healthy subjects underwent 24-h ambulatory gastric bilirubin monitoring with the bilirubin sensor in the upper stomach. Gastric bilirubin monitoring with two simultaneous bilirubin probes, one in the upper stomach and the other in the antrum, was performed on a separate occasion. Gastric bilirubin exposure in the initial and repeat studies showed a good correlation (R = 0.60, P < 0.01). Gastric bilirubin exposure in the upper stomach and the antrum showed a high degree of correlation (R = 0.90, P < 0.01). In conclusion, reproducible results are obtained with ambulatory gastric bilirubin monitoring and duodenogastric reflux does not exhibit significant regional variation within the stomach. PMID- 11270799 TI - Colorectal adenomas and diet: a case-control study. Colorectal Adenoma Study Group. AB - It has been postulated that high intakes of animal fat and protein and low intakes of fiber, calcium, and antioxidants increase the risk of colorectal cancer. Whether specific types of protein such as that from red meat are important, and whether vegetables might be key protective factors will also be considered in this study. Dietary intake over the past year was studied according to the diet history method by means of a case-control study in 184 cases and matched controls. After adjustment for energy, relative weight, and social class, no associations were found for fat or protein in comparison with either control group. Unexpectedly, carbohydrate intake was inversely related with adenoma risk, the RR being 0.29 (0.10-0.81) for quintile 5 versus 1 in comparison with hospital controls. None of the antioxidants showed a significant protective effect except beta-carotene intake in comparison with hospital controls, the RR being 0.24 (0.11-0.50) for the highest versus the lowest quintile. There was, however, a statistically significant positive association between adenomas and meat consumption with the RR for the highest versus the lowest quintile. There was, however, a statistically significant positive association between adenomas and meat consumption with the RR for the highest versus the lowest quintile of intake being 3.6 (1.7-7.5) in comparison with hospital controls and 4.4 (1.6-12.1) in comparison with population controls. Our data support the protective role for carbohydrate intake and of beta-carotene intake in the etiology of colorectal adenomas and show a strong increased risk for developing adenomas in those with high meat intake. PMID- 11270800 TI - Short chain fatty acids differentially modulate cellular phenotype and c-myc protein levels in primary human nonmalignant and malignant colonocytes. AB - Short chain fatty acids may protect colonic mucosa against neoplastic transformation by modulating colonocyte phenotype, DNA synthesis, and c-myc levels. To test this hypothesis, nonmalignant and malignant human colonocytes were isolated from surgical specimens and treated with 10 mM acetate, propionate, or butyrate. Markers of cellular phenotype, DNA synthesis, and c-myc protein levels were assayed by alkaline phosphatase and dipeptidyl dipeptidase IV activities, [3H]thymidine labeling, and western blotting, respectively. Butyrate, in particular, exerted discordant effects on alkaline phosphatase (P < 0.05), and c-myc levels (P < 0.05, N > or = 6) in nonmalignant and malignant human colonocytes. DPDD was unaffected by any of the short chain fatty acids tested. [3H]Thymidine labeling was differentially stimulated by short chain fatty acids in both cell types and greater DNA synthesis rates were observed in malignant colonocytes (P < 0.005, N = 16). These data suggest that in vitro, butyrate, in particular, may differentially modulate phenotype, DNA synthesis, and c-myc in nonmalignant and malignant human colonocytes. PMID- 11270801 TI - Levels of sICAM-1, sVCAM-1 and MCP-1 in patients with hyperlipoproteinemia IIa and -IIb. AB - OBJECTIVE: Hyperlipoproteinemia is one of the factors that are involved in the development of atherosclerosis. One of the mechanisms through which these high plasma lipid levels trigger the formation of atherosclerotic lesions is a change in the expression of adhesion molecules on endothelial and smooth muscle cells. The aim of this study was to evaluate the plasma levels of soluble intracellular adhesion molecule-1 (sICAM-1), soluble vascular cell adhesion molecule-1 (sVCAM 1) and monocyte chemoattractant protein-1 (MCP-1) in patients with Type IIa (HLP IIa) and IIb (HLP-IIb) hyperlipoproteinemias. SUBJECTS: Twenty patients with HLP IIa, 20 patients with HLP-IIb and 23 control subjects were studied. To accurately evaluate adhesion molecule levels, we excluded those hyperlipemic patients and control subjects who had an inflammatory disease. METHODS: Plasma sICAM-1, sVCAM 1 and MCP-1 levels were measured by the ELISA method. RESULTS: sVCAM-1 levels in HLP-IIa and HLP-IIb patients (535 +/- 27 ng/ml and 545 +/- 22 ng/ml, respectively) did not differ significantly from those in the control group (558 +/- 20 ng/ml). sICAM-1 levels were significantly higher in patients with HLP-IIa and HLP-IIb (279 +/- 10 ng/ml and 322 +/- 12 ng/ml, respectively) compared to the control group (226 +/- 10 ng/ml). MCP-1 levels were significantly higher in HLP IIa and HLP-IIb patients (151 +/- 12 pg/ml vs 154 +/- 12 pg/ml, respectively) compared to healthy controls (98 +/- 4 pg/ml). sICAM-1 levels in the HLP-IIb group were significantly higher than in the HLP-IIa group. CONCLUSION: The results of the study suggest that lipid abnormalities affect the levels of adhesion molecules and chemokines in plasma. PMID- 11270802 TI - Caffeine metabolism before and after liver transplantation. AB - AIM: To study drug metabolism in patients before and after liver transplantation using caffeine as a probe drug. Forty-five patients undergoing liver transplantation for various liver diseases and who had well documented dossiers were selected for the study. Before the liver transplantation and 1 month, 1 year, and 6 years after liver transplantation, they were given 200 mg of caffeine by the oral route in the morning after voiding their bladder. Twenty-four-hour urine samples were collected and caffeine and metabolites were determined by HPLC: 1-methylurate (1U), 1-methylxanthine (1X), 1.7-dimethylurate (17U), 1.7 dimethylxanthine (17X), 7-methylxanthine (7X), 3-methylxanthine (3X), 1.3 dimethylurate (13U), 3.7-dimethylxanthine (37X), 1.3-dimethylxanthine (13X), 1.3.7-trimethylxanthine = caffeine (137X). Indices of enzyme activities were calculated from the following urinary elimination ratios: (AFMU+1U+1X)/17U for CYP1A2, 17U/17X for CYP2A6, 1U/1X for xanthine oxidase (XO), AFMU/(AFMU+1U+1X) for N-acetyltransferase (NAT-2). RESULTS: Compared with results obtained in a group of 70 healthy subjects, caffeine metabolism before liver transplantation was deeply depressed with a decreased elimination rate in the case of all metabolites and a decreased CYP1A2 activity. Caffeine metabolism began to return to the control values one month after transplantation. One year and 6 years after liver transplantation, quantitatively, the metabolism of caffeine was stable and not different from control, but with qualitative modifications. CYP1A2 activity was decreased with reduced urinary elimination rates of 1X and 17X. XO and CYP2A6 activities and 1U and 17U urinary elimination rates were increased. Immunosuppressive treatment was possibly responsible for the metabolic pathway changes. Almost the same modifications were observed in 9 patients after bone marrow transplantation who had been treated with the same immunosuppressive drugs, cyclosporine and azathioprine. During severe rejection phases in 6 of the liver transplant patients, caffeine metabolism was progressively decreased when the usual liver function tests showed moderate but uniform changes. CONCLUSION: Despite an apparent normal drug-metabolic function, immunosuppressive treatment induces stable variations in drugmetabolic pathways after liver transplantation which can be detected from the changes in caffeine metabolism. PMID- 11270803 TI - Pharmacokinetics of oral talinolol following a single dose and during steady state in patients with chronic renal failure and healthy volunteers. AB - OBJECTIVE: The objective of this study was to investigate the effect of renal impairment on the pharmacokinetics of the selective beta1-receptor antagonist talinolol. METHODS: Pharmacokinetic data were obtained in 12 healthy volunteers, 12 patients with renal impairment and 8 patients with terminal renal insufficiency after the oral administration of 100 mg talinolol and under steady state conditions (100 mg talinolol daily). Concentrations of talinolol in plasma, urine and dialysate during hemodialysis were measured with a validated HPLC method. RESULTS: Talinolol is absorbed quite rapidly from the gastrointestinal tract (tmax 2.5-4 h). Steady state conditions were reached within 3-4 days depending on renal function. The calculated mean elimination half-life (t(1/2z)) in healthy volunteers (11 male, 1 female) was about 12 h. After an oral dose of 100 mg, about 55% of the bioavailable talinolol is eliminated unchanged in the urine. This fraction is reduced to 25% in patients with moderate to severe renal failure. A strong correlation was found between the renal elimination of talinolol and creatinine clearance. In patients with renal failure, the delayed elimination leads to an increase in t(1/2z) and to a decrease in the apparent total body clearance. Steady state trough levels (c(min)ss) in these patients are about 2.2-fold higher than in volunteers. The hemodialysability of talinolol was low. CONCLUSION: The disposition of talinolol shows a strong dependence on the renal function. On the basis of the kinetic data for talinolol, dose reductions of 30-50% are recommended in subjects with moderate to severe renal impairment. PMID- 11270804 TI - Plasma and urine pharmacokinetics of the dopamine agonist alpha dihydroergocryptine in patients with hepatic dysfunction. AB - OBJECTIVE: The aim of this study was to evaluate the pharmacokinetic behavior of unchanged alpha-dihydroergocryptine (DHEC, Almirid, Desitin Arzneimittel GmbH, Hamburg, Germany, under licence of Polichem S.A., Luxembourg) and total DHEC (unchanged DHEC and pooled metabolites) in plasma and urine in patients with impaired hepatic function, following administration of single oral doses. METHODS: The study was carried out according to an open, uncontrolled, parallel group design, investigating two study groups: patients with hepatic dysfunction, i.e. with evidence of stable cirrhosis (n = 10) and age- and sex-matched healthy subjects (n = 8). Each subject received a single dose of 20 mg DHEC. Blood samples were taken at specified intervals up to 72 h after dosing and urine was collected fractionally for 24 h. Concentrations of unchanged DHEC were determined by RIA and concentrations of total DHEC (unchanged and pooled metabolites) by EIA. RESULTS: The plasma and urinary pharmacokinetics of DHEC and its metabolites were characterized by large variability. In patients with impaired hepatic function, the geometric mean Cmax and AUC(0-infinity) values for unchanged DHEC were 571.3 pg/ml (CV: 0.87) and 4038 pg x h/ml (CV: 1.04) and were approximately 2 times (2.04, 95% CI: 0.93 to 4.46 and 2.11, 95% CI: 0.58 to 7.73 for Cmax and AUC(0-infinity), respectively) larger than those measured in age-matched healthy controls. The 24-hour urinary excretion was approximately 3 times (3.41, 95% CI: 0.95 to 12.21) higher in patients with hepatic dysfunction. Similar results were obtained for total DHEC. CONCLUSIONS: The results reflect an increased systemic exposure in patients with impaired hepatic function which is not due to a reduced urinary excretion/elimination or reduced renal clearance. The most likely mechanism involved is a reduction in pre-systemic biotransformation. The observed range of effects on the pharmacokinetics of DHEC in patients with compromized hepatic function does not suggest the need to revise the dosage recommendations, since treatment with DHEC is generally started with low doses and is slowly up titrated according to the individual response and the occurrence of adverse effects. Nevertheless, lower maintenance doses are likely to be achieved. PMID- 11270806 TI - Bioequivalence of two aceclofenac tablet formulations after a single oral dose to healthy male Korean volunteers. AB - A bioequivalence study of aceclofenac tablets (test formulation: Dong-A, reference formulation: Airtal) was conducted in 16 healthy male Korean volunteers who received each medicine at a dose of 100 mg in a 2 x 2 crossover study. There was a one-week washout period between the doses. Plasma concentrations of aceclofenac were monitored by high-performance liquid chromatography over a period of 24 hours after the administration. AUCinf (the area under the plasma concentration-time curve from time zero to time infinity) was calculated by the linear-log trapezoidal method. Cmax (maximum plasma drug concentration) and tmax (time to reach Cmax) were compiled from the plasma concentration-time data. Analysis of variance was carried out using logarithmically transformed AUCinf and Cmax, and non-transformed tmax. There were no significant differences between the medications in AUCinf and Cmax. The point estimates and 90% confidence intervals for AUCinf (parametric) and Cmax (parametric) were 1.04 (0.93 to approximately 1.17) and 0.99 (0.91 to approximately 1.08), respectively, satisfying the bioequivalence criteria of the European Committee for Proprietary Medicinal Products and the US Food and Drug Administration Guidelines. The corresponding value for tmax was 0.75 (0.00 to approximately 1.00). Moreover, the modified Pitman-Morgan's adjusted F-test indicated that the bioavailabilities of aceclofenac in the 2 medications were comparable regarding intra- and interindividual variability. Therefore, these results indicate that the 2 medications of aceclofenac are bioequivalent and, thus, may be prescribed interchangeably. PMID- 11270805 TI - Bioequivalence study of a novel Solutab tablet formulation of amoxicillin/clavulanic acid versus the originator film-coated tablet. AB - With amoxicillin/clavulanic acid Solutab tablet, a new tablet formulation of amoxicillin/clavulanic acid (500/125), was developed. The aim of the present study was to demonstrate bioequivalence between the new tablet formulation, taken as an intact tablet and after prior dispersal, versus the originator product viz. Augmentan film-coated tablet. The study was performed in 48 healthy volunteers, according to an open, single-dose three-period, crossover design. Blood samples were taken prior to each administration and at 10 time points after dosing. Plasma concentrations of amoxicillin and clavulanic acid were determined by validated high performance liquid chromatography with UV detection. With regard to amoxicillin, the results were within the preset bioequivalence range of 0.8 to 1.25 for the ratios of the primary parameters AUC(0-t) and Cmax. In terms of clavulanic acid the 90% confidence intervals of the ratios for AUC(0-t) and Cmax versus the reference lay outside the predefined bioequivalence range of 0.75 to 1.33. This result, however, was mainly due to the large variability of the reference formulation compared to the amoxicillin/clavulanic acid Solutab tablet. Based on statistical indications that 3/48 subjects with extremely low levels on the reference formulation could be regarded as "outliers" and after excluding these subjects' data from the statistical analysis, results for clavulanic acid were within the predefined bioequivalence range of 0.75 to 1.33. Overall, the amoxicillin/clavulanic acid Solutab tablet provided, in comparison to the reference tablet, less variable levels of clavulanic acid, thus giving more appropriate protection to the available amoxicillin. Thirteen adverse events were reported post dosing by 7 subjects. There were no differences in incidence of adverse events between amoxicillin/clavulanic acid Solutab tablet taken intact or dispersed and Augmentan. PMID- 11270807 TI - Distribution of pyranose and furanose forms of 6-deoxyheptoses in water solution. AB - On the basis of 1H and 13C NMR spectroscopy studies, the proportion of pyranose and furanose forms of 6-deoxyheptoses in water solution was determined. Water solution of 6-deoxyheptoses contains all possible furanose and pyranose forms (except 6-deoxy-gluco-heptose for which only pyranose was found), although pyranose is dominant. PMID- 11270808 TI - Synthesis, analysis and rearrangement of novel unnatural glucosinolates. AB - As part of a structure activity study to examine the interaction of glucosinolates with leaf surfaces, a number of glucosinolates were synthesised bearing novel side chain functionalities. These included 7-carboxyheptyl, heptyl, and naphthyl side chains. For the carboxyheptyl glucosinolate, a novel intramolecular rearrangement reaction was observed during the final deprotection step, which generated an ester attached to the C-3 of glucose. Studies by 1H NMR spectroscopy showed that the hydrophobic side chain associated with one face of the glucose ring and it was proposed that this was the driving force for the rearrangement. Similar hydrophobic interactions were also observed between the heptyl and naphthyl side chains and the glucose. PMID- 11270809 TI - Synthesis of 4'-O-acetyl-maltose and alpha-D-galactopyranosyl-(1-->4)-D glucopyranose for biochemical studies of amylose biosynthesis. AB - The chemical synthesis of the title compounds as maltose analogs, in which the non-reducing end is modified by acetylation of the 4'-OH group or by reversing its configuration, is reported. For synthesis of the 4'-O-acetylated analog, beta maltose was converted into its per-O-benzylated-4',6'-O-benzylidene derivative followed by removal of the benzylidene acetal function and selective silylation at C-6'. Acetylation at C-4' of the obtained silylated compound followed by removal of the benzyl ether protecting groups and subsequent desilylation afforded the desired analog. The other maltose analog was synthesized via the glycosidation reaction between the glycosyl donor, O-(2,3,4,6-tetra-O-benzyl alpha/beta-D-galactopyranosyl)trichloroacetimidate and the glycosyl acceptor, phenyl 2,3,6-tri-O-benzyl-1-thio-beta-D-glucopyranoside followed by removal of the phenylthio group and debenzylation to provide the desired analog. PMID- 11270810 TI - Practical synthesis of a tetrasaccharide derivative corresponding to ristomycin A and ristocetin A. AB - A practical synthesis of fully benzoylated tetrasaccharide, whose free form is indispensable to the antibiotic ristomycin A for the process of dimerization and binding to the cell wall, was achieved via sequential assembly of the building blocks, allyl 3,4-di-O-benzoyl-alpha-D-glucopyranoside, 2,3,4-tri-O-benzoyl-alpha L-rhamnopyranosyl trichloroacetimidate, 2-O-acetyl-3,4,6-tri-O-benzoyl-alpha-D mannopyranosyl trichloroacetimidate, and 2,3,5-tri-O-benzoyl-alpha-D arabinofuranosyl trichloroacetimidate. A one-pot preparation of allyl 3,4-di-O benzoyl-2-O-tert-butyldimethylsilyl-6-O-triphenylmethyl-alpha-D-glucopyranoside is described, and regioselective glycosylation is carried out using perbenzoylated sugar trichloroacetimidates as glycosyl donors in the presence of a catalytic amount of trimethylsilyl trifluoromethanesulfonate (TMSOTf). PMID- 11270811 TI - Structure and conformation of a novel genetically engineered polysaccharide P2. AB - A new exocellular polysaccharide (P2) has been produced by the manipulation of a glycosyl transferase gene (aceP) involved in the biosynthesis of the polysaccharide acetan by the bacterium Acetobacter xylinum strain CKE5. The P2 polysaccharide has been studied by methylation analysis, reductive cleavage, and 1H and 13C NMR spectroscopy. The data are consistent with the structure predicted when the aceP gene is deactivated: [Molecular structure: see text]. The effect of cooling on proton NMR line width indicates a coil-helix transition in P2 at about 70 degrees C. PMID- 11270812 TI - Characterization of cytosolic sialidase from Chinese hamster ovary cells: part I: cloning and expression of soluble sialidase in Escherichia coli. AB - The cDNA of Chinese hamster ovary (CHO) cell cytosolic sialidase was amplified by RT-PCR and cloned into the pGEX-2T plasmid vector encoding for glutathione S transferase (GST). Screening revealed transformed Escherichia coli clones with the constructed plasmid encoding the CHO cell sialidase sequence. After isopropyl beta-D-thiogalactopyranoside (IPTG) induction, SDS-PAGE of the total protein extracts revealed a new protein of about 70 kDa, correlating with the molecular weight of a fusion protein composed of the GST (26 kDa) and the cloned cytosolic CHO cell sialidase (43 kDa). A soluble fusion protein was purified from sonified E. coli homogenates by one-step affinity chromatography on Glutathione Sepharose 4B, which showed sialidase activity towards 4-methyl-umbelliferyl-alpha-D-N acetylneuraminic acid (MUF-Neu5Ac) substrate. Induction of cells with 0.1, 0.5, and 1.0 mM IPTG revealed highest total protein amounts after induction with 1.0 mM IPTG, but highest specific activity for affinity chromatography purified eluates from cultures induced with 0.1 mM IPTG. Therefore, large scale production was performed by inducing cells during exponential growth in a 25 L bioreactor for 3 h with 0.1 mM IPTG after chilling the cell suspension to 25 degrees C. The amount of 26.46 mg of 40-fold purified GST-sialidase with a specific activity of 0.999 U/mg protein was obtained from crude protein extracts by one-step affinity chromatography. 2-Deoxy-2,3-dehydro-N-acetylneuraminic acid (Neu5Ac2en) and Neu5Ac were competitive inhibitors for the sialidase, the former being the more effective one using MUF-Neu5Ac as the substrate. The cytosolic sialidase is capable of desialylating a wide spectrum of different types of gangliosides using a thin-layer chromatography overlay kinetic assay without detergents. This is the subject of the accompanying paper (Muthing, J.; Burg, M. Carbohydr. Res. 2001, 330, 347-356). PMID- 11270813 TI - Characterization of cytosolic sialidase from Chinese hamster ovary cells: part II. Substrate specificity for gangliosides. AB - Cytosolic Chinese hamster ovary (CHO) cell sialidase has been cloned as a soluble glutathione S-transferase (GST)-sialidase fusion protein with an apparent molecular weight of 69 kD in Escherichia coli. The enzyme has then been produced in mg quantities at 25-L bioreactor scale and purified by one-step affinity chromatography on glutathione sepharose (Burg, M.; Muthing, J. Carbohydr. Res. 2001, 330, 335-346). The cloned sialidase was probed for desialylation of a wide spectrum of different types of gangliosides using a thin-layer chromatography (TLC) overlay kinetic assay. Different gangliosides were separated on silica gel precoated TLC plates, incubated with increasing concentrations of sialidase (50 degreesU/mL up to 1.6 mU/mL) without detergents, and desialylated gangliosides were detected with specific anti-asialoganglioside antibodies. The enzyme exhibited almost identical hydrolysis activity in degradation of GM3(Neu5Ac) and GM3(Neu5Gc). A slightly enhanced activity, compared with reference Vibrio cholerae sialidase, was detected towards terminally alpha(2-3)-sialylated neolacto-series gangliosides IV3-alpha-Neu5Ac-nLc4Cer and VI3-alpha-Neu5Ac nLc6Cer. The ganglio-series gangliosides G(D1a), G(D1b), and G(T1b), the preferential substrates of V. cholerae sialidase for generating cleavage resistant G(M1), were less suitable targets for the CHO cell sialidase. The increasing evidence on colocalization of gangliosides and sialidase in the cytosol strongly suggests the involvement of the cytosolic sialidase in ganglioside metabolism on intracellular level by yet unknown mechanisms. PMID- 11270814 TI - L-Altruronic acid formed by epimerization of D-galacturonic acid methyl esters during saponification of citrus pectin. AB - While searching for oligosaccharides containing rhamnose residues in the endopolygalacturonase (EPG) digest of saponified citrus pectin, we found several oligomers containing, in addition to galacturonic acid, a sugar previously unreported in pectin. The 1- and 2-D 1H NMR spectra of the oligosaccharides were consistent with the sugar being a uronic acid with its 2- and 3-hydroxyls being axial and 4-hydroxyl being equatorial. MALDI-TOF mass spectrometry indicated that the oligomers consisted solely of uronic acids. Reduction of the uronic acids in the oligosaccharides converted them to galactose and altrose. The altrose was found to be the L enantiomer by comparison of its trimethylsilyl (-)-2-butyl glycosides to those of authentic D-altrose and a racemic mixture. The sugar was not found in oligosaccharides prepared from EPG digestion of citrus pectin deesterified with pectin methylesterase rather than saponification. Thus, it appears that during saponification, a small proportion of the methylesterified galacturonic acid residues in pectins is epimerized at C-5 leading to formation of L-altruronic acid residues. PMID- 11270815 TI - Experimental evidence for a semi-flexible conformation for arabinoxylans. AB - Purified water-soluble arabinoxylans from wheat flour were deferuloylated and fractionated into six fractions by graded ethanol precipitation. Further fractionation by HPSEC on Sephacryl S500 resulted in 48 subfractions with low polydispersity index. Conformational characteristics (persistence length q, hydrodynamic parameter v and Mark-Houwink exponent a) were similar among all subfractions and fitted with a semi-flexible conformation, whatever their structural characteristics. Substitution degree of the xylan backbone by arabinose residues has no influence on the conformation of arabinoxylans. PMID- 11270816 TI - The hydrolysis of barley beta-glucan by the cellulase EC 3.2.1.4 under dilute conditions is identical to that of barley solubilase. AB - Barley beta-glucan solubilase is an enzyme that degrades barley beta-glucan in extracts obtained from barley flour. The solubilase preferentially attacks the longer blocks of beta-(1-->4) linkages, i.e., those containing at least nine glucosyl residues. There is strong evidence to suggest that the solubilase derives from fungi associated with the husk of the grain. It was found that cellulase (EC 3.2.1.4) from Trichoderma sp. shows similar activity under dilute conditions. Since fungi associated with the husk of the grain are known to produce these types of cellulases, there is no need, based on current evidence, to propose the existence of a unique enzyme, i.e., solubilase, for the solubilising behaviour of enzymes in the barley grain. PMID- 11270817 TI - The combined hydrolysis and hydrogenation of inulin catalyzed by bifunctional Ru/C. AB - A one-pot process for hydrolysis and hydrogenation of inulin to D-mannitol and D glucitol over a bifunctional Ru/C catalyst was developed. The hydrolysis is catalyzed by the carbon support, onto which acidity was introduced by pre oxidation. The effect of different carbon treatments on the hydrolysis of inulin was studied. Oxidation with ammonium peroxydisulfate resulted in a carbon with the highest hydrolysis activity. On this carbon, long chain inulin is hydrolyzed faster than inulin rich in short chains. The application of high pressure (up to 100 bar) increased the hydrolysis rate substantially. The combined process was successfully conducted with a Ru-catalyst supported on this oxidized carbon. PMID- 11270819 TI - A practical route to partially protected pyrrolidines as precursors for the stereoselective synthesis of alexines. AB - Either 3-O-benzoyl- (2a) or 3-O-benzyl-1,2-O-isopropylidene-beta-D-fructopyranose (2b) were regioselectively O-benzylated at C-4 to give 4a and 4b, respectively, which were transformed into 5-azido-3-O-benzoyl-4-O-benzyl- (6a) and 5-azido-3,4 di-O-benzyl-5-deoxy-1,2-O-isopropylidene-alpha-L-sorbopyranose (6b) by nucleophilic displacement of the corresponding 5-O-mesyl derivatives 5a and 5b by sodium azide in DMF, respectively. Compound 6b was also prepared from 4b in one step by the Mitsunobu methodology. Deacetonation of 6a and 6b gave the partially protected free azidouloses 8a and 8b, respectively, that were protected as their 1-O-TBDPS derivatives 9a and 9b. Hydrogenation of 9b over Raney nickel gave stereoselectively (2R,3R,4R,5S)-3,4-dibenzyloxy-2'-O-tert-butyldiphenylsilyl-2,5 bis(hydroxymethyl)pyrrolidine (12) which was identified by transformation into the well known (2R,3R,4R,5S)-3,4-dihydroxy-2,5-bis(hydroxymethyl)pyrrolidine (1, DGDP). PMID- 11270818 TI - Solid state NMR and X-ray diffraction studies of alpha-D-galacturonic acid monohydrate. AB - Crystalline alpha-D-galacturonic acid monohydrate has been studied by 13C CPMAS NMR and X-ray crystallography. The molecular dynamics were investigated by evaluating 13C spin-lattice relaxation in the rotating frame (T1rho) and chemical shift-anisotropy properties of each carbon. Only limited molecular motions can be detected in the low frequency (< 10(4) Hz) range by 13C relaxation time measurements (T1rho) and changes of chemical shift anisotropy properties as a function of temperature. X-ray analysis (at both ambient temperature and 150 K) shows that the acid has the usual chair-shaped, pyranose ring conformation, and that the acid and water molecules are linked, through all their O-H groups, in an extensively hydrogen-bonded lattice. PMID- 11270820 TI - Convenient synthesis of 4,6-di-O-benzyl-myo-inositol and myo-inositol 1,3,5 orthoesters. AB - Convenient high yielding methods for the preparation of 4,6-di-O-benzyl-myo inositol, myo-inositol 1,3,5-orthoformate and myo-inositol 1,3,5-orthoacetate, without involving chromatography are described. Myo-inositol was converted to racemic 2,4-di-O-benzoyl-myo-inositol 1,3,5-orthoformate by successive treatment with triethyl orthoformate and benzoyl chloride. The dibenzoate obtained on benzylation with benzyl bromide and silver(I) oxide gave 2-O-benzoyl-4,6-di-O benzyl-myo-inositol 1,3,5-orthoformate. Deprotection of the benzoate and the orthoformate with isobutylamine and aqueous trifluoroacetic acid, respectively gave 4,6-di-O-benzyl-myo-inositol in an overall yield of 67%. Myo-inositol orthoformate and orthoacetate were prepared and isolated as their tribenzoates. The free orthoesters were regenerated by deprotection of the benzoates by aminolysis with isobutylamine. PMID- 11270821 TI - Comparison of chemical and enzymatic synthesis of 2-acetamido-2-deoxy-D-mannose 6 phosphate: a new approach. AB - Chemical and enzymatic methods to synthesis of 2-acetamido-2-deoxy-D-mannose-6 phosphate (ManNAc-6-P) have been investigated. A new preparative method has been developed although some established procedures were tried. In this new method, a 6-O-acetyl or 4,6-di-O-acetyl group of the per-O-acetylated 2-acetamido-2-deoxy-D mannose (ManNAc) were regioselectively removed with an esterase from the yellow yeast, Rhodosporidium toruloides, followed by phosphorylation and O-deacetylation under mild conditions. 1H and 13C NMR data spectra of ManNAc-6-P were recorded. PMID- 11270822 TI - Conformational preferences in glycosylamines. implications for the exo-anomeric effect. AB - The conformational preferences about the C-N bond in N-(4-methoxyphenyl)-2,3,4,6 tetra-O-acetyl-alpha (1) and beta-D-glucopyranosylamine (2), in the solid state and in solution, have been investigated. The crystal structure of the axially substituted alpha anomer (1) indicates a conformational preference about the C-1 N bond in which nN-->sigma*C-O exo-anomeric interactions may be expressed, although this conformational preference is not displayed in solution. The solution conformation relieves steric interactions that result from expression of the exo-anomeric effect in the solid-state conformation. The conformational preference in the equatorially substituted beta anomer (2) both in solution and in the solid state is similar and permits expression of nN-->sigma*C-O exo anomeric interactions. The structural data for 1 and 2 indicate significant differences in O-5-C-1-N-1 bond angles but insignificant differences in each of the O-5-C-1 or C-1-N-1 bond lengths. The J(C-1-H-1 coupling constants in 1 and 2 indicate a greater coupling constant for the alpha anomer that is consistent with a dominant nO-->sigma*C-H orbital interaction in the beta anomer that weakens the C-1-H-1 bond. PMID- 11270823 TI - Sulfated beta-(1-->4)-galacto-oligosaccharides and their effect on angiogenesis. AB - Sulfated beta-(1-->4)-galacto-oligosaccharides were prepared from an arabino galacto-rhamno-galacturonan from Lupinus polyphyllus Lindl. by successive partial hydrolysis and SO3-pyridine sulfation in DMF. The resulting oligosaccharide polysulfates were analyzed by analytical GPC and the sulfate content was determined by ion chromatography. DP 5 and higher showed a pronounced antiangiogenic effect with scores of 0.9-1.2 for DP 7-9 using the CAM-assay. An interaction with the fibroblast growth factor FGF-2 was noticed for DP 4-12 depending on the degree of sulfation using the FGF-2-trypsin assay. PMID- 11270824 TI - Molecular and crystal structures of N-(beta-D-galactopyranosyl)pyridinium bromide and its per-O-acetylated derivative. AB - 1H NMR spectroscopy and X-ray diffraction data are described for N-(2,3,4,6-tetra O-acetyl-beta-D-galactopyranosyl)pyridinium bromide and N-(beta-D galactopyranosyl)pyridinium bromide. X-ray crystallography revealed that the O acetylated salt crystallizes with two molecules of water and one molecule of ethanol. PMID- 11270825 TI - Virtual placement of frontal ventricular catheters using frameless neuronavigation: an "unbloody training" for young neurosurgeons. AB - OBJECTIVE: To evaluate virtually the reliability of freehand puncture of the anterior horn of the lateral ventricle and to provide realistic, but unbloody training for young neurosurgeons. METHODS: Virtual placement of ventricular catheters was performed repeatedly by neurosurgical doctors and thereafter controlled by neuronavigation. With the help of a frameless stereotactic navigation device they virtually had to hit the anterior horn of the lateral ventricle on the MRI of 29 brains with normal ventricular sizes and 60 pathological ventricles, respectively. The catheter placement was simulated using the pointer of the navigation system (EasyGuide Neuro). The monitor screen was blinded, so that on-line control was impossible. Virtual elongation of the pointer tip was performed on the workstation and the position of the virtual catheter was evaluated on a printout. RESULTS: Virtual freehand catheter placement was performed 145 times into the MRIs of the normal brains. In 66 cases (45%) the site of the catheter tip was judged as accurate as shown by the navigation system. No difference concerning the number of correctly placed catheters was observed when comparing more and less experienced doctors. The results in the 60 pathological MRIs of patients differed with respect to the size of the ventricles: in narrow ventricles an accurate placement succeeded in 7 of 22 cases (32%), moderately enlarged ventricles were accurately hit in 15 out of 32 cases (46%) and wide ventricles in 5 of 6 attempts (83%), respectively. CONCLUSION: This setup is a simple, practicable tool for neurosurgical education. The virtual freehand placement of ventricular drains controlled by neuronavigation provides an unbloody training of a routine neurosurgical procedure in a realistic setting without the risk of injuring a patient. Neuronavigation systems can serve therefore as a link between learning from observation and handling the real situation. PMID- 11270826 TI - Relationships of virtual reality neuroendoscopic simulations to actual imaging. AB - Advances in computer technology have permitted virtual reality images of the ventricular system. To determine the relevance of these images we have compared virtual reality simulations of the ventricular system with endoscopic findings in three patients. The virtual fly-through can be simulated after definition of waypoints. Flight objects of interest can be viewed from all sides. Important drawbacks are that filigree structures may be missed and blood vessels cannot be distinguished clearly. However, virtual endoscopy can presently be used as a planning tool or for training and has future potential for neurosurgery. PMID- 11270827 TI - Clinical features in patients requiring reoperation after failed endoscopic procedures for hydrocephalus. AB - The aim of this study was to clarify the clinical features of patients at risk of secondary obstruction following endoscopic fenestration. Clinical notes and endoscopic findings for 15 patients treated with endoscopic procedures were retrospectively reviewed. Endoscopic third ventriculostomy (ETV) was performed as initial treatment in 4 patients with non-communicating hydrocephalus, including a neonate with myelomeningocele, and as an alternative to shunt revision in 4 patients. Two patients with non-communicating hydrocephalus caused by tumor or arachnoid cyst were also managed with third ventriculostomy. Four patients with loculated hydrocephalus underwent endoscopic septostomy. A child with an isolated fourth ventricle was treated with endoscopic aqueductoplasty. Of the 15 patients undergoing endoscopic procedure, 4 required reoperation. Of the 10 patients treated with ETV, only the neonate with myelomeningocele required a ventriculoperitoneal shunt because of failure of the initial procedure. Of the 4 patients treated with endoscopic septostomy, 2 children with loculated hydrocephalus following intraventricular hemorrhage (IVH) underwent a second septostomy. In a patient with an isolated fourth ventricle following posthemorrhagic hydrocephalus, recurrence was noted 8 months after the initial procedure. He underwent a second procedure using a stent implanted into the aqueduct to maintain CSF circulation. Sufficient stomal size or implantation of a stent may be required in the under-2-year age group with hydrocephalus accompanied by IVH and associated with myelomeningocele, in whom the risk of secondary obstruction may be high. PMID- 11270828 TI - The "optimal" burr hole position for endoscopic third ventriculostomy: results from 31 stereotactically guided procedures. AB - ETV is a well established and successful method in contemporary neurosurgery. With growing experience there is a more efficient patient selection and further advances in technical know how. We evaluated retrospectively a consecutive group of 27 patients who were treated in our institution by stereotactic guided ETV between 1992 and 1996. When reviewing their postoperative imaging studies (MRI/CT) we could measure the position of the burr hole as port of entry for the rigid endoscope in 17 out of 23 finally selected patients. The median lateral position was 28 mm (mean 26.5 mm) from the midline and 8 mm (mean 6.5 mm) anterior of the coronal suture. We conclude that the optimal burr hole position should be 3 cm lateral to the midline and 1 cm anterior of the coronal suture, in the patients with normal anatomical findings. PMID- 11270829 TI - Transient hyponatriemia complicated by seizures after endoscopic third ventriculostomy. AB - We present an infant who underwent endoscopic third ventriculostomy due to symptomatic hydrocephalus secondary to aqueductal stenosis. This is the first reported case of inappropriate secretion of antidiuretic hormone complicated by hyponatriemia and seizures following endoscopic surgery. The possibility of such a neuroendoscopic complication should alert neurosurgeons and close observation of serum electrolytes is highly recommended in the acute postoperative period, particularly in infants. PMID- 11270830 TI - Surgery of intrinsic cerebral neoplasms in eloquent areas under local anesthesia. AB - 28 patients with a mean age of 43.6 years were operated on for a cerebral neoplasm situated in close proximity to an eloquent area (24 speech area, 4 motor cortex) from 1996 to 1999. Preoperatively, all patients had undergone a detailed neuropsychological examination. In 10 patients aphasic disturbances could be detected. All patients underwent preoperative PET studies (methionine and (15)O labeled water with activation during speech or finger tapping). These were performed and co-registered with MRI data to demonstrate the topographical relationship between motor or language function and the tumor borders. Anesthesia was induced with i.v. administration of propofol (150-250 mg/h). Craniotomy was performed under local infiltration anesthesia. After opening of the dura, sedation was stopped and operation was continued with the patient being alert and co-operative. With close clinical observation during electrical cortex stimulation, a speech arrest could be triggered or avoided. The motor cortex was identified by recording the phase reversal of the contralateral SEP of the median nerve and by direct cortical stimulation. As soon as aphasic or motor disturbances appeared, the tumor removal was continued with the goal of avoiding these specific regions. In 27 patients, preexisting neuropsychological and neurological deficits did not worsen. Only one patient was left postoperatively with a major permanent aphasic deficit that was present preoperatively to a minor degree. The use of local anesthesia in craniotomy for surgery of intrinsic cerebral neoplasms in eloquent areas allows for a continuous and repetitive monitoring of speech and motor function during the removal of even those tumors that were previously considered inoperable. PMID- 11270831 TI - Early effects of PRS-irradiation for 9L gliosarcoma: characterization of interphase cell death. AB - We characterized the interphase cell death of 9L gliosarcoma after high-dose rate, low-energy photon irradiation using the Photon Radiosurgery System (PRS), a novel device for interstitial radiotherapy. Within 24 hours after irradiation with a dose of 18 Gray, 22.0% of cells underwent metabolic cell death, whereas dead cells in controls stayed less than 5.0% (p<0.005). In the majority of sensitive cells, loss of membrane integrity preceded the lethal morphological changes. The response was dose-dependent over the range of 9-18 Gray, but saturation was obtained over 18 Gray. On the other hand, a significant (p < 0.01) increase in the number of TUNEL-positive cells with apoptotic morphology was detected 6-24 hours after irradiation, but the fraction remained 1.9-2.1% of the population and was independent of the doses between 9 and 25 Gray. Apoptotic cells were rarely observed in the control cells (0.3-0.6%). Our data indicate that single high-dose irradiation induces both necrotic and apoptotic interphase cell death in 9L gliosarcoma, but rapid cell death mostly occurs through the non apoptotic pathway. PMID- 11270832 TI - Gamma knife radiosurgery of the brain stem cavernomas. AB - Over 6 years (1992-1998) 26 patients with brain stem cavernomas were treated using the Leksell gamma knife in Prague. 25 patients had a follow up of 6-66, median 24 months. Annual risk of bleeding before radiosurgery was 4%. After gamma knife treatment sudden impairment of neurodeficit reported as rebleeding was observed in 4 patients at 6-51 months, median 16.5 months, after radiosurgery. This represented a 6.8% risk of rebleeding after radiosurgery, which is not significantly different from the risk before radiosurgery. MRI or CT was performed in 24 patients 6-48, median 24, months after radiosurgery. There were no signs of rebleeding in any of the patients, nor any increase of the cavernoma. A decrease of cavernoma size was observed in 8 (33%) of patients. Temporary collateral edema after radiosurgery was detected in 5 (21%) of patients 3-12, median 11, months after radiosurgery. Neurodeficit was observed in 21 of 26 patients before radiosurgery. Improvement of the neurodeficit was detected in 9 (43%) of them 6-36, median 8, months after radiosurgery. Temporary morbidity caused by collateral edema or rebleeding occurred in 7 patients (28%) and permanent morbidity remained in 2 patients (8%). 2 patients died because of rebleeding 6 and 51 months after radiosurgery and the third patient for unrelated reason. Radiosurgery of the brain stem cavernomas was indicated when there was bleeding in the history or progressive neurodeficit and microsurgery was considered too risky. Leksell gamma knife radiosurgery of cavernomas has proved its low morbidity and zero mortality. In case of an insufficient effect of radiosurgery, or if the protective effect from rebleeding comes too late, morbidity and mortality can correspond to the natural course of the disease, as it was left without any treatment. PMID- 11270833 TI - Endoscopic surgery of the third ventricle: the subfrontal trans-lamina terminalis approach. AB - The authors describe an endoscopic approach to the anterior aspect of the third ventricle and demonstrate its use in the cadaver. This technique consists of a small supraorbital craniotomy and a subfrontal trans-lamina terminalis approach to the third ventricle. It may be helpful in the management of refractory third ventricular lesions that cannot be easily accessed endoscopically through the foramina of Monro. PMID- 11270834 TI - Titanium clamps for refixation of bone fragments in the repair of depressed skull fractures: technical note. AB - The rigid fixation of bone fragments in the repair of depressed skull fractures can be a problem, especially if not all fragments are replaceable. Usually, in these cases mini- or microplates are used. The Craniofix titanium clamp (Aesculap, Germany) was developed for the fixation of bone flaps after osteoplastic craniotomy. It consists of a screw holding together two 10-mm diameter metallic disks with concaved teeth pressing bone flap and cranium between the two disks. We used this system for the operative treatment of two patients with depressed skull fracture. A rigid fixation of the bone fragments was achieved in both cases. The postoperative three-dimensional CT scan showed a good fragment alignment. From the report of two cases we show that this system is a useful tool in the fixation of bone fragments in the repair of depressed skull fractures. PMID- 11270835 TI - The human tail associated with intraspinal lipoma: case report. AB - A case of a tail in a 9-month-old baby is reported. Computed tomography and magnetic resonance imaging clearly demonstrated the presence of spina bifida and lipoma continuous from the tail to the spinal canal. A few medical-historical aspects are discussed. The human tail may be related to spinal dysraphism and requires detailed neuroimaging investigation and microsurgery. PMID- 11270836 TI - Biochemical tests of growth hormone status in short children. PMID- 11270837 TI - Urea kinetic modelling: a measure of dialysis adequacy. PMID- 11270838 TI - Evaluation of effect of analytical imprecision in maternal serum screening for Down's syndrome. AB - Formulae to evaluate the effect of inter-assay analytical imprecision (expressed as the coefficient of variation) in maternal serum screening for Down's syndrome have been developed. Experimentally determined imprecision in Down's syndrome risk (based on maternal serum alpha-fetoprotein, unconjugated oestriol and human chorionic gonadotrophin) was found to be consistent with predicted values. Imprecision in the measurement of analytes becomes amplified when risk is calculated using the values of these analytes. A large separation between the means and small standard deviations for normal and affected pregnancies are the characteristics of the tests most useful in screening, but these attributes also result in the most imprecision in risk. In addition, the relative imprecision associated with Down's syndrome risk is not the same for all women screened. Combining tests for multivariate analyses results in a complex compounding of the errors. The need for strict quality control and test reproducibility is emphasized. The effect of analytical imprecision should be of particular concern to laboratories that provide screening for women of advanced maternal age. PMID- 11270839 TI - What is the role of genetic testing in diagnosis of haemochromatosis? PMID- 11270841 TI - Identification of an apolipoprotein(e) variant associated with type III hyperlipoproteinaemia in an indigenous Australian. AB - As a result of testing for lipid and apolipoprotein(e) (apo E) phenotype status of an indigenous Australian community, an apo E variant associated with type III hyperlipoproteinaemia has been identified. Apo E phenotype was determined by analysis of VLDL by isoelectric focusing, and genotype on DNA amplified by polymerase chain reaction, using two different restriction enzyme isotyping assays. Phenotypes and genotypes were discordant in samples from two subjects and an abnormal-sized restriction fragment was also observed in their genotyping gel patterns. DNA sequencing studies revealed this was due to a single nucleotide deletion, 3817delC, at amino acid 136 on apo E. This resulted in a new reading frame and the premature termination of the apo E protein due to a stop codon (TGA) at nucleotide 4105. The variant apo E null allele showed a recessive mode of inheritance and, in combination with the E2 allele, resulted in the type III hyperlipoproteinaemic phenotype but when inherited with the E4 allele had no marked effect on plasma lipids. PMID- 11270840 TI - Prevalence of macroamylasaemia using polyethylene glycol precipitation as a screening method. AB - Four hundred and fifty-four subjects with hyperamylasaemia were screened for the presence of macroamylase using polyethylene glycol precipitation (PEG) and the Beckman automated amylase assay based on the hydrolysis of maltotetraose. Twenty five subjects (5.5%) exhibited PEG precipitation values suggestive of macroamylasaemia (MA) (>52% loss of original amylase activity). Macroamylasaemia was confirmed by gel filtration chromatography (GFC) in 21/25 subjects, using albumin as a molecular weight marker. Biochemical and clinical details of the 21 subjects identified are presented and discussed. It is recommended that serum exhibiting more than 57% precipitation of the original amylase activity by PEG should be examined by GFC to confirm or exclude MA. PMID- 11270842 TI - Improvement of a standard MIRA method for urinary oxalate by elimination of reagent carry-over. PMID- 11270843 TI - Serum creatine kinase: an adjunct biochemical index of subclinical thyrotoxicosis? PMID- 11270844 TI - Seasonal variation in glycated haemoglobin in diabetics. PMID- 11270845 TI - Practical implications of in vitro stability of cardiac markers. PMID- 11270846 TI - Evaluation of thin-layer chromatography methods for laxative detection. PMID- 11270847 TI - Solid-phase extraction as an alternative to dichloromethane in analysis of urine free cortisol. PMID- 11270848 TI - Automated method for urine 5-hydroxyindole-acetic acid by high-performance liquid chromatography using Gilson ASTED sample preparation unit. PMID- 11270849 TI - Cider potomania. PMID- 11270850 TI - Cider potomania. PMID- 11270851 TI - Development of a 1 MV field-emission transmission electron microscope. AB - We developed a 1 MV field-emission transmission electron microscope. This paper reports details and specifications of the instrument. The microscope was designed to obtain a bright and coherent electron beam by using the field emission gun equipped with a pre-accelerating magnetic lens and the high-voltage power supply with high stability (0.5 ppm min(-1)). Using this microscope, the brightness of 1.8 x 10(10) A cm(-2) sr(-1) and the lattice resolution of 49.8 pm were attained. PMID- 11270852 TI - High-resolution electron microscopy of crystal defects in C70-toluene solvate. AB - Defects in C70 x C6H5CH3 were examined using transmission electron microscopy. Two types of defects with respect to (001) and [011] were observed. The former is an inter-grain of BCT phase in Amm2 orthorhombic matrix, whereas the latter is a twin. Origins of the observed defects were determined to be of a shearing nature in the directions of [010] and <011>. In addition, an attempt is made to explain the decagonal symmetry of the compound, as reported by several authors, with respect to strain relaxation by observing defects as well as a multiple-twinning model described by Fleming et al. (Phys. Rev. B44: 888 (1991)). The conversion of C70-toluene solvate into C70 pure phases is discussed in terms of the defects. PMID- 11270853 TI - The structure of an Al3Pd-based modulated structure studied by atomic-scale electron microscopy observations. AB - A modulated structure based on an Al3Pd structure in an Al70Co25Pd5 alloy has been studied by atomic-scale observations with high-resolution transmission electron microscopy (HREM) and high-angle annular detector dark-field (HAADF) technique using a scanning transmission electron microscope. From the electron microscopy observations, we can reveal an arrangement of atom columnar clusters, which produces peculiar shifts of diffraction spots in an electron diffraction pattern. Arrangements of transition-metal atoms in defect regions in the modulated structure are proposed from the observed HAADF image. PMID- 11270854 TI - Osmium-metal coating device using hollow-cathode plasma CVD method. AB - A novel osmium-metal coating device for SEM observation has been developed to prevent negative charge build-up on specimens by applying the hollow-cathode low voltage discharge plasma chemical vapour deposition (CVD) method. The CVD method using the hollow-cathode offers the following advantages. (i) The method can deposit osmium-metal at very low discharge voltage that is as low as half of that of the planar parallel electrode method. Therefore, the method avoids damage due to ion bombardment during the coating process. (ii) The method can minimize the quantity of the OsO4 gas by introducing directly into the hollow-cathode. This feature is important to prevent the air pollution caused by the purged gas. (iii) A large coating area is guaranteed because the Os ion is filled in the hollow cathode where the specimen is holed. (iv) The lower discharge voltage can be used by mixing Ar, N2 or air with the OsO4 gas as the environmental gas in the chamber. (v) The hybrid coating is also available by lining the appropriate metal material such as platinum (Pt) on the surface of the inside of the hollow cathode. The method uses the plasma CVD of Os metal as well as the ion-sputter deposition of the lined metal. PMID- 11270855 TI - Development of battery charging system for the FE gun system of HVEM. AB - We have developed a reliable and efficient battery-charging system for the field emission (FE) gun system of high voltage electron microscopes, where the operating condition of the whole FE gun system can be controlled and monitored through a bi-directional optical fiber system, and the control and monitoring circuits located on the high potential of 1 MV are driven by rechargeable Ni-Cd batteries. The power transmission from ground to the high potential is performed through filter condenser circuit. Under the condition that the filter condenser current is limited to 0.2 A(rms) (root-mean-square value), it is possible to transmit the maximum power as high as 65 W, which is enough for the daily operation of the FE gun system. The charging circuit has a function of protecting the batteries from over-charging. PMID- 11270856 TI - Simulation of atomic-scale high-angle annular dark field scanning transmission electron microscopy images. AB - Image simulations for high-angle annular dark field (HAADF) scanning transmission electron microscopy (STEM) based on the Bethe's eigen-value method are presented. The effects of aperture size and defocus of a probe-forming lens, both of which determine the shape of the probe, and the effect of the distortion, influencing the Bloch wave field on atomic columns channelled by electrons, on the HAADF image intensity are discussed in terms of dynamical effect. These effects are illustrated by our experimental atomic-scale HAADF-STEM images, detected in a detector range of 50-110 mrad. It is emphasized that the image simulations are indispensable for quantification of experimental HAADF-STEM images and as such provide a valuable compositional analysis for every atomic column along the incident beam. PMID- 11270857 TI - Image quality improvement using helium gas in low voltage variable pressure scanning electron microscopy. AB - An effective combination of the low voltage and variable pressure (VP) scanning electron microscopy (SEM) are discussed. In low voltage VP-SEM, helium gas is utilized for reducing the amount of scatter of the primary electron beam. Most samples can receive various benefits obtained from the combination of low voltage and low vacuum observation. Compared to a back-scattered electron (BSE) image in air, signal-to-noise ratio (SNR) of a BSE image taken with helium gas is 5.4 times under a pressure of 50 Pa and an accelerating voltage of 1.5 kV. PMID- 11270858 TI - Binding of influenza type A viruses to group B Streptococcus and haemagglutination by virus-bound bacteria. AB - We studied the bindings of human influenza A type viruses to group B Streptococcus (GBS), types Ia, II, III and IV, of sialic acid (SA)alpha2-3 linkage, using A/PR/8/34(H1N1) and A/Memphis/1/71(H3N2). The viruses were found to bind to all types of GBS, with the exception of PR/8/34 for GBSII, and to elute from GBSIa, III and IV at 37 degrees C, except GBSII. Electron microscopy confirmed these behaviours of the influenza viruses. The virus-binding capability of GBS types seemed to depend on the side chain length of the terminal SA. Treatment of GBSIa, III and IV, except enzyme-resistant type II, with bacterial neuraminidase resulted in the loss of virus-binding capability of GBS. These findings confirmed that SAalpha2-3 linkage of GBS capsules functions as receptor for human influenza viruses. When singular bacteria were prepared from mainly chain-like GBS with sonication, viruses were found to bind to them more efficiently. Untreated and sonicated GBS were both aggregated with high doses of virus. Furthermore, using A/Memphis/1/71(H3N2) and GBSII, we found that virus bound GBS, untreated or sonicated, caused haemagglutination (HA). The morphological evidence that chicken erythrocytes were bridged with virus-bound native GBSII, supporting occurrences of HA, was obtained. Statistical analysis suggested that HA by virus-bound sonicated (singular) GBS was mediated by bacteria bound by at least two or three virus particles. PMID- 11270859 TI - Developmental changes in the ultrastructure of the Sauromatum guttatum (Araceae) mitochondria. AB - Electron microscopic studies show that the electron density of the mitochondria of the thermogenic appendix of Sauromatum guttatum inflorescences changes during development. Two to 3 days before heat-production, the mitochondria accumulate osmiophilic, electron-dense material between the inner and outer membranes. During heat-production and the release of volatiles compounds the osmiophilic material disappears from the inter membrane space. Deposits with the same electron density are also found in the endoplasmic reticulum. It is concluded that the mitochondria may also have the capacity to accumulate that material in the inter membrane space. PMID- 11270860 TI - The capillary of the organum vasculosum laminae terminalis (OVLT) in rabbits is more permeable to horseradish peroxidase (HRP) than that in rats. AB - The distribution of the tracer, horseradish peroxidase (HRP), in the organum vasculosum laminae terminalis (OVLT) of rabbits and rats was examined by electron microscopy. In rabbits, the HRP-diaminobenzidine reaction products were heavily distributed in the OVLT and surrounding brain tissues 10 and 60 min after the injection of HRP (50 mg kg(-1), i.v.), and were retained in the parenchymal tissues at 24 h post-injection. The majority was found in numerous large phagosomes of macrophages located in the perivascular spaces of the vascular beds and in ependymal cells (tanycytes) in the parenchyma. A large amount of reaction product was also localized in the intercellular clefts of the parenchyma. HRP incorporation was seen in both nerve cells and ependymal cells in the OVLT at 10 min post-injection, but only in nerve cells in the preoptic area at 60 min post injection. In rats, however, a small amount of the reaction products was observed in the OVLT 10 min after the injection of HRP (50 and 70 mg kg(-1), i.v.), and the levels were markedly reduced at 60 min post-injection. No HRP-incorporation by nerve cells was seen. From these findings, we concluded that the capillary of the OVLT of the rabbit is more permeable to HRP than that of the rat. PMID- 11270861 TI - Monoliths as stationary phases for separation of proteins and polynucleotides and enzymatic conversion. AB - Monoliths are considered as a novel generation of stationary phases. They were applied for capillary electrochromatography and liquid chromatography exploiting every action principle such as ion-exchange, affinity recognition, reversed phase, and hydrophobic interaction. The fast separation was explained by convective transport of the solutes through the bed. The contribution of this mode of transport is similarly explained as done for the beds packed with particles with gigapores. For monolithic beds, the concept of an ultrashort bed was frequently used. This mode of operation allows very short separation time. In many cases a gradient elution is necessary to achieve separation. Examples of applications for protein and polynucleotide separation performed on monoliths are given. Enzymatic conversion was described showing the examples of several immobilzed enzymes. PMID- 11270862 TI - Sugar-lectin interactions investigated through affinity capillary electrophoresis. AB - The affinity interactions of Concanavalin A (Con A) with various saccharide oligomers (dextrins, dextrans, and selected N-linked glycans from various glycoproteins) have been investigated through a capillary electrophoresis approach. Con A has shown a notable binding discrimination between the alpha-1,6 linked dextran and alpha-1,4-linked dextrin oligomers. Both the binding capacity and binding discrimination appear to decrease with an increase in sugar chainlength. While the core structure of N-linked glycans is deemed to be responsible for the overall binding of various glycans to Con A, the presence of mannose units at the non-reducing ends was found to be very beneficial to the affinity interaction with Con A. Finally, a connection between the glycan-lectin interaction and glycoprotein-lectin interaction has also been suggested. PMID- 11270863 TI - Silicon microstructures for high-speed and high-sensitivity protein identifications. AB - Silicon microtechnology has been used to develop a microstructure toolbox in order to enable high accuracy protein identification. During the last 2 years we developed and applied monocrystalline silicon structures and established new automated protein analysis platforms. The development of a high throughput protein platform is presented where fully automated protein identifications are performed. It includes the reduction and alkylation of the protein sample in a standard 96- or 384-well plate format prior to injection of 1 microl samples into the continuous flow based microtechnology platform. The processed sample is transferred to a microchip nanovial array target using piezoelectric microdispensing. Identification is made by MALDI-TOF MS and a database search. After the initial sample reduction and alkylation period of 50 min the platform can digest and process protein samples at a speed of 100 samples in 210 min. An optional configuration of the platform, operating the dispenser in the 'static mode', enables on-target enrichment of low abundant proteins and peptides e.g. from 2DE samples. This makes detection at the low attomole level possible. PMID- 11270864 TI - Identification of multiple sources of charge heterogeneity in a recombinant antibody. AB - Seven forms of a therapeutic recombinant antibody that binds to the her2/neu gene product were resolved by cation-exchange chromatography. Structural differences were assigned by peptide mapping and HIC after papain digestion. Deamidation of light chain asparagine 30 to aspartate in one or both light chains is responsible for two acidic forms. A low potency form is due to isomerization of heavy chain aspartate 102; the Asp102 succinimide is also present in a basic peak fraction. Forms with both Asn30 deamidation and Asp102 isomerization modifications were isolated. Deamidation of heavy chain Asn55 to isoaspartate was also detected. Isoelectric focusing in a polyacrylamide gel was used to verify the assignments. All modifications were found in complementarity determining regions. PMID- 11270865 TI - Characterization of a recombinant monoclonal antibody by mass spectrometry combined with liquid chromatography. AB - In this report, we present the characterization of a humanized monoclonal antibody specific for the human epidermal growth factor receptor (hEGFR). Direct analysis by matrix assisted laser desorption ionization mass spectrometry (MALDI MS) of peptide mixtures and chromatographically isolated fractions allowed identification of 94.0% and 85.4% of the amino acid sequence of light and heavy chains, respectively. Microheterogeneity sources were identified in light and heavy chains and a previously unreported posttranslational modification for immunoglobulins was found. One N-glycosylation site was identified in the heavy chain with non-sialylated bianntenary fucosylated structures. This study is one of the first to assess the potential of MALDI-MS in combination with more conventional protein chemistry techniques for the characterization of monoclonal antibodies. PMID- 11270866 TI - Characterization and quality control of recombinant adenovirus vectors for gene therapy. AB - Highly purified recombinant adenovirus undergoes routine quality controls for identity, potency and purity prior to its use as a gene therapy vector. Quantitative characterization of infectivity is measurable by the expression of the DNA binding protein, an early adenoviral protein, in an immunofluorescence bioassay on permissive cells as a potency determinant. The specific particle count, a key quality indicator, is the total number of intact particles present compared to the number of infectious units. Electron microscopic analysis using negative staining gives a qualitative biophysical analysis of the particles eluted from anion-exchange HPLC. One purity assessment is accomplished via the documented presence and relative ratios of component adenoviral proteins as well as potential contaminants by reversed-phase HPLC of the intact virus followed by protein peak identification using MALDI-TOF mass spectrometry and subsequent data mining. Verification of the viral genome is performed and expression of the transgene is evaluated in in vitro systems for identity. Production lots are also evaluated for replication-competent adenovirus prior to human use. For adenovirus carrying the human IL-2 transgene, quantitative IL-2 expression is demonstrated by ELISA and cytokine potency by cytotoxic T lymphocyte assay following infection of permissive cells. Both quantitative and qualitative analyses show good batch to batch reproducibility under routine test conditions using validated methods. PMID- 11270867 TI - Automated LC-LC-MS-MS platform using binary ion-exchange and gradient reversed phase chromatography for improved proteomic analyses. AB - A simple multidimensional liquid chromatography system utilizing an isocratic pump and a HPLC system is described for the comprehensive proteomic analysis of complex peptide digest mixtures by coupled LC-LC-MS-MS techniques. A binary ion exchange separation was achieved through the use of a strong cation-exchange column followed by a reversed-phase column for data-dependent LC-MS-MS analysis of the unbound analytes, and following salt elution (and concomitant column reequilibration), the bound analytes. Off-line validation of the platform showed near quantitative recovery of fractionated peptides and essentially complete ion exchange partitioning. In comparative analyses of a highly complex peptide digest mixture a >40% increase in the number of peptide and protein identifications was achieved using this multidimensional platform compared to an unfractionated control. PMID- 11270868 TI - Proteomics of glycoproteins based on affinity selection of glycopeptides from tryptic digests. AB - Identification of glycoproteins in complex mixtures derived from either human blood serum or a cancer cell line was achieved in a process involving the steps of (1) reduction and alkylation, (2) proteolysis of all proteins in the mixture with trypsin, (3) affinity chromatographic selection of the glycopeptides with an immobilized lectin, (4) direct transfer of the glycopeptide fraction to a reversed-phase liquid chromatography (RPLC) column and further fractionation by gradient elution, (5) matrix-assisted laser desorption ionization mass spectrometry of individual fractions collected from the RPLC column, and (6) peptide identification based on a database search. The types of glycoproteins analyzed were; (1) N-type glycoproteins of known primary structure, (2) N-type glycoproteins of unknown structure, and (3) O-type glycoproteins glycosylated with a single N-acetylglucosamine. Identification of peptides from complex mixtures was greatly facilitated by either C-terminal sequencing with a carboxypeptidase mixture or by comparing chromatographic behavior and mass to standards, as in the case of a known protein. In addition, deglycosylation of peptides with N glycosidase F was necessary to identify N-type glycoproteins of unknown structure. The strength of this approach is that it is fast and targets specific molecular species or classes of glycoproteins for identification. The weakness is that it does not discriminate between glycoforms. PMID- 11270869 TI - Protein analysis of an individual Caenorhabditis elegans single-cell embryo by capillary electrophoresis. AB - We present a simple one-dimensional electrophoretic map of the expressed proteins in a Caenorhabditis elegans embryo. The embryo was taken from an adult nematode, injected into a 50-microm I.D. capillary, and lysed. The proteins were fluorescently labeled and then separated by capillary electrophoresis and detected by laser-induced fluorescence. Over 20 components were resolved in the 22-min separation. The dynamic range was outstanding for this separation, noise in the baseline was less than 0.01% the amplitude of the largest component. PMID- 11270870 TI - Matrix-assisted laser desorption-ionization mass spectrometry peptide mass fingerprinting for proteome analysis: identification efficiency after on-blot or in-gel digestion with and without desalting procedures. AB - In theory, peptide mass fingerprinting by matrix assisted laser desorption ionization mass spectrometry (MALDI-MS) has the potential to identify all of the proteins detected by silver staining on gels. In practice, if the genome of the organism investigated is completely sequenced, using current techniques, all proteins stained by Coomassie Brilliant Blue can be identified. This loss of identification sensitivity of ten to hundred-fold is caused by loss of peptides by surface contacts. Therefore, we performed digestion and transfer of peptides in the lower microl range and reduced the number of steps. The peptide mix obtained from in-gel or on-blot digestion was analyzed directly after digestion or after concentration on POROS R2 beads. Eight protein spots of a 2-DE gel from Mycobacterium bovis BCG were identified using these four preparation procedures for MALDI-MS. Overall, on-blot digestion was as effective as in-gel digestion. Whereas higher signal intensities resulted after concentration, hydrophilic peptides are better detected by direct measurement of the peptide mix without POROS R2 concentration. PMID- 11270871 TI - Analysis of regulatory phosphorylation sites in ZAP-70 by capillary high performance liquid chromatography coupled to electrospray ionization or matrix assisted laser desorption ionization time-of-flight mass spectrometry. AB - A methodology for the rapid and quantitative analysis of phosphorylation sites in proteins is presented. The coupling of capillary high-performance liquid chromatography (HPLC) to electrospray ionization mass spectrometry (ESI-MS) allowed one to distinguish phosphorylation sites based on retention time and mass difference from complex peptide mixtures. The methodology was first evaluated and validated for a mixture of non-, mono-, and dityrosine-phosphorylated synthetic peptides, corresponding to the tryptic fragment 485-496 (ALGADDSYYTAR) of the human protein tyrosine kinase ZAP-70. The limits of detection for the non-, mono- and diphosphorylated peptides were about 15, 40 and 100 fmol, respectively, when using a 300 microm I.D. column. Application of the method was extended to identify phosphopeptides generated from a trypsin digest of recombinant autophosphorylated ZAP-70, in particular with respect to quantifying the status at the regulatory phosphorylation sites Tyr-492 and Tyr-493. Combination of chromatographic and on-line tandem mass spectrometry data allowed one to ascertain the identity of the detected peptides, a prerequisite to analyses in more complex biological samples. As an extension to the methodology described above, we evaluated the feasibility of interfacing capillary HPLC to matrix assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS), using a micromachined piezoelectric flow-through dispenser as the interface. This enabled direct arraying of chromatographically separated components onto a target plate that was precoated with matrix for subsequent analysis by MALDI-TOF MS without further sample handling. PMID- 11270872 TI - Quantification of plasma-derived blood coagulation factor VIII by real-time biosensor measurements. AB - Plasma-derived blood coagulation factor VIII was analyzed in real time using biosensor technology. Monoclonal antibodies directed against the heavy and against the light chain of factor VIII were immobilized on different carboxymethyl dextran surfaces. Different factor VIII concentrations were injected over the antibody surfaces in parallel and response levels were determined from the dissociation phase at a fixed time after sample injection. Serial dilutions of plasma-derived factor VIII with known concentrations determined by a commercial FVIIIC:Ag ELISA were used as standards. A quantification limit of 0.9 I.U./ml with antibody 530p and 1.5 I.U./ml with antibody 531p was calculated. Intra-assay precision expressed as percent coefficient of variation was below 10% for concentrations above 0.6 I.U./ml. Inter-assay precision for antibody 530p was below 20% for concentrations higher than 0.6 I.U./ml. For 531p, inter-assay precision was below 10% for concentrations higher than 2 I.U./ml. A sensor chip lifetime in respect to regeneration of at least 100 cycles for both antibodies was found. The small sample requirement of 35 microl allows fast analysis of different FVIII products and the use of two monoclonal antibodies directed against two different FVIII domains provides additional information about the integrity of the FVIII molecule. PMID- 11270873 TI - Characterization of cellobiohydrolase I (Cel7A) glycoforms from extracts of Trichoderma reesei using capillary isoelectric focusing and electrospray mass spectrometry. AB - Capillary isoelectric focusing (CIEF) was used to profile the cellulase composition in complex fermentation samples of secreted proteins from Trichoderma reesei. The enzyme cellobiohydrolase I (CBH I, also referred to as Cel7A), a major component in these extracts, was purified from different strains and characterized using analytical methods such as CIEF, high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD), and capillary liquid chromatography-electrospray mass spectrometry (cLC-ESMS). ESMS was also used to monitor the extent of glycosylation in CBH I isolated from T. reesei strain RUT-C30 and two derivative mutant strains. Selective identification of tryptic N-linked glycopeptides was achieved using LC-ESMS on a quadrupole/time of-flight instrument with a mixed scan function. The suspected glycopeptides were further analyzed by on-line tandem mass spectrometry to determine the nature of N linked glycans and their attachment sites. This strategy enabled the identification of a high mannose glycan attached to Asn270 (predominantly Man8GlcNAc2) and single GlcNAc occupancy at Asn45 and Asn384 with some site heterogeneity depending on strains and fermentation conditions. The linker region of CBH I was shown to be extensively glycosylated with di-, and tri-saccharides at Thr and Ser residues as indicated by MALDI-TOF and HPAEC-PAD experiments. Additional heterogeneity was noted in the CBH I linker peptide of RUT-C30 strain with the presence of a phosphorylated di-saccharide. PMID- 11270874 TI - Purification of novel peptide antibiotics from human milk. AB - A strategy was established for the identification of novel antimicrobial peptides from human milk. For the generation of bioactive peptides human milk was acidified and proteolyzed with pepsin simulating the digest in infants stomachs. Separation of proteins and resulting fragments was performed by means of reversed phase chromatography detecting the antimicrobial activity of each fraction using a sensitive radial diffusion assay. In order to avoid the purification of the known abundant antimicrobial milk protein lysozyme, it was identified in HPLC fractions by its enzymatic activity and by matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS). On condition that lysozyme was not detectable and antibacterial activity of HPLC fractions was caused by a peptide, which was confirmed by proteolytic cleavage leading to a loss of activity, further purification was performed by consecutive chromatographic steps guided by the antibacterial assay. Using this strategy, an as yet unknown casein fragment exhibiting antimicrobial activity was purified in addition to antimicrobial lactoferrin fragments. The new antimicrobial peptide resembles a proteolytic fragment of human casein-K (residues 63-117) and inhibits the growth of gram positive, gram-negative bacteria, and yeasts. Our results confirm that antimicrobially-active peptides are liberated from human milk proteins during proteolytic hydrolysis and may play an important role in the host defense system of the newborn. PMID- 11270875 TI - On leadership, P values, cancer, and family. PMID- 11270876 TI - Primary venorrhaphy for traumatic inferior vena cava injuries. AB - Primary venorrhaphy for traumatic inferior vena cava (IVC) injury has been criticized because of the potential for stenosis, thrombosis, and embolism. A retrospective study was performed to evaluate the morbidity and outcome of this method. Thirty-eight patients at our institution had traumatic injuries to the IVC between 1994 and 1999. Thirty (79%) were from firearms, five (13%) from stab wounds, and three (8%) from blunt trauma. Six patients died in the emergency department. The remaining 32 patients underwent exploratory celiotomy with 23 survivors and nine intraoperative deaths for a mortality rate of 28 per cent (nine of 32). Vascular control was achieved by manual compression in 44 per cent and by local clamping directly above and below the injury in 38 per cent. All repairs were by primary venorrhaphy, and no patient was treated with patch angioplasty or venous reconstruction. Three patients had caval ligation. Follow up IVC imaging in 11 patients revealed that the IVC was patent in eight, narrowed in two, and thrombosed below the renal veins in one. One patient developed a pulmonary embolus. The vast majority of traumatic injuries to the IVC can be managed by direct compression or local clamping and primary venorrhaphy. Direct repairs are associated with a low thrombosis and embolic complication rate. PMID- 11270878 TI - Concomitant blunt enteric injuries with injuries of the liver and spleen: a dilemma for trauma surgeons. AB - Prompt identification of enteric injuries after blunt trauma remains problematic. With the increased utilization of nonoperative management of blunt abdominal trauma gastrointestinal disruptions may escape timely detection and repair. The purpose of this study was to evaluate blunt enteric injuries requiring operative repair in adult patients and the association of concomitant hepatic and/or splenic injuries. Over a 10-year period (January 1990 through December 1999) 1648 patients suffered blunt liver, spleen, and/or enteric injuries, with 87 (5.3%) of these requiring operative repairs of the enteric injury. These patients had enteric injury only (EI) (60.9%; 53 of 87), concomitant enteric/splenic injury (ESI) (10.3%; 9 of 87), concomitant enteric/hepatic injury (EHI) (13.8%; 12 of 87), and enteric/hepatic/splenic injury (EHSI) 14.9% (13 of 87). A delay in treatment of >8 hours from presentation of EI compared with either EHI or ESI was not significantly different between the two groups. EHSI had exploratory laparotomy more expeditiously related to hemodynamic instability. Mortality rates were higher with EHI related to hemorrhagic shock and/or severe traumatic brain injury. Morbidity was not related to a delay in diagnosis until the period of delay was greater than 24 hours. The nonoperative management of blunt solid organ injury does not delay the detection and treatment of concomitant bowel injuries compared with isolated blunt enteric injuries. Occult enteric injury with solid organ injury has a low incidence and represents a continuing challenge to the clinical acumen of the trauma surgeon. PMID- 11270877 TI - Serum amylase and lipase elevation is associated with intracranial events. AB - Serum amylase and lipase elevation has been observed in trauma patients and patients with traumatic intracranial bleeding. However, the causes of this elevation have not been clearly elucidated. A further question remains as to whether other intracranial events are associated with such enzyme elevation as well. We retrospectively reviewed 75 patients consecutively admitted to Cook County Hospital Neurosurgical Intensive Care Unit over a 3-month period for trauma, infection, tumor, or other space-occupying lesions with an unstable condition or neurological deficit. Eleven patients (15%) had elevated amylase and lipase levels. The patients were divided into two groups: Group I (n = 64) had normal and Group II (n = 11) had raised amylase and lipase levels [amylase 402 +/ 444 U/L with normal < or = 125 U/L and lipase 474 +/- 313 U/L with normal < or = 55 U/L]. All Group II patients suffered an intracranial event. Twenty-four Group I (38%) and 10 Group II (91%) patients required craniotomy (P < 0.01). No patients had clinical or radiographic evidence of pancreatitis. In summary, intracranial events are associated with serum amylase and lipase elevation probably through centrally activated pathways. Because of the lack of diagnostic value, routine pancreatic enzyme monitoring should not be performed in this patient population. PMID- 11270879 TI - Infectious complications following duodenal and/or pancreatic trauma. AB - Patients with pancreatic and/or duodenal trauma often have a high incidence of infectious complications. In this study we attempted to find the most important risk factors for these infections. A retrospective review of the records of 167 patients seen over 7 years (1989 through 1996) at an urban Level I trauma center for injury to the duodenum and/or pancreas was performed. Fifty-nine patients (35%) had isolated injury to the duodenum (13 blunt, 46 penetrating), 81 (49%) had isolated pancreatic trauma (18 blunt, 63 penetrating), and 27 (16%) had combined injuries (two blunt, 25 penetrating). The overall mortality rate was 21 per cent and the infectious morbidity rate was 40 per cent. The majority of patients had primary repair and/or drainage as treatment of their injuries. Patients with pancreatic injuries (alone or combined with a duodenal injury) had a much higher infection rate than duodenal injuries. The patients with duodenal injuries had significantly lower penetrating abdominal trauma indices, number of intra-abdominal organ injuries, and incidence of hypothermia. On multivariate analysis independent factors associated with infections included hypothermia and the presence of a pancreatic injury. Although injuries to the pancreas and duodenum often coexist it is the pancreatic injury that contributes most to the infectious morbidity. PMID- 11270880 TI - Ultrasound evaluation of the magnitude of pneumothorax: a new concept. AB - Pneumothorax is commonly seen in trauma patients; the diagnosis is confirmed by radiography. The use of ultrasound where radiographic capabilities are absent, is being investigated by the National Aeronautics and Space Administration. We investigated the ability of ultrasound to assess the magnitude of pneumothorax in a porcine model. Sonography was performed on anesthetized pigs in both ground based laboratory (n = 5) and microgravity conditions (0 x g) aboard the KC-135 aircraft during parabolic flight (n = 4). Aliquots of air (50-100 cm3) were introduced into the chest to simulate pneumothorax. Results were videorecorded and digitized for later interpretation. Several distinct sonographic patterns of partial lung sliding were noted including the combination of a sliding zone with a still zone and a "segmented" sliding zone. These "partial lung sliding" patterns exclude massive pneumothorax manifested by a complete separation of the lung from the parietal pleura. In 0 x g, the sonographic picture is more diverse; one x g differences between posterior and anterior aspects are diminished. Modest pneumothorax can be inferred by the ultrasound sign of "partial lung sliding." This finding, which increases the negative predictive value of thoracic ultrasound, may be attributed to intermittent pleural contact, small air spaces, or alterations in pleural lubricant. Further studies of these phenomena are warranted. PMID- 11270881 TI - Interleukin-10 attenuates proinflammatory cytokine production and improves survival in lethal pancreatitis. AB - Given that interleukin (IL)-10 (IL-10) serves as a potent down-regulator of specific proinflammatory cytokines we reasoned that its administration should improve outcome in situations in which the biological response to a severe inflammatory challenge is the critical determinant of survival. To test our hypothesis we administered IL-10 in the setting of lethal pancreatitis to determine its effect on proinflammatory cytokine production and survival. We divided Sprague-Dawley rats into three groups. Controls (Group 1, n = 5) received a sham laparotomy. We induced pancreatitis in Group 2 (n = 9) and Group 3 (n = 9) via laparotomy and intrapancreatic infiltration of one mL of 5 per cent sodium taurocholate. Group 2 was treated only with saline, whereas Group 3 was treated with 10,000 units of IL-10 (in saline) at 30 minutes, 3.5 hours, and 6.5 hours after induction of pancreatitis. Serial blood samples were obtained at 6.5 hours for measurement of amylase, IL-1, and IL-6. The Kaplan-Meier method, Wilcoxon test, and Student's t test were used for analysis. Seven-day survival was 100, 0, and 45 per cent in Groups 1, 2, and 3, respectively. Production of amylase, IL-1, and IL-6 was lower in the IL-10-treated group (Group 3) compared with the group treated with saline alone (Group 2, P < 0.05). We conclude that administration of IL-10 in the setting of otherwise 100 per cent lethal experimental pancreatitis significantly reduces production of amylase, IL-1, and IL-6 and improves survival. PMID- 11270882 TI - What is normal intra-abdominal pressure? AB - The causes and effects of increased intra-abdominal pressure and abdominal compartment syndrome have been well documented. However, there have been no large series to determine normal intra-abdominal pressure in hospitalized patients. The purpose of this study was to determine normal intra-abdominal pressure in randomly selected hospitalized patients and to identify factors that predict variation in normal intra-abdominal pressure. A total of 77 patients were prospectively enrolled between September 1998 and July 1999. Data obtained included patient demographics (i.e., age, gender, height, weight, and body mass index), reason for hospitalization and bladder catheterization, previous and current surgical status, comorbidities, and intra-abdominal pressures. Intra abdominal pressure readings were obtained through an indwelling transurethral bladder (Foley) catheter. Data were analyzed by analysis of variance and multiple regression analysis. There were 36 females and 41 males with a mean age of 67.7 years. Average weight, height, and body mass index were 79.6 kg, 1.70 m, and 27.6 kg/m2, respectively. Mean intraabdominal pressure was 6.5 mm Hg (range 0.2-16.2 mm Hg). Body mass index was positively related to intra-abdominal pressure (P < 0.0004). Gender, age, and medical and surgical histories did not significantly affect intra-abdominal pressure. However, using multiple regression analysis, a relationship between intra-abdominal pressure, body mass index, and abdominal surgery was discovered. Intra-abdominal pressure is related to a patient's body mass index and influenced by recent abdominal surgery. Thus, the normal intra abdominal pressure can be estimated in hospitalized patients by using the derived equation. Knowledge of the expected intra-abdominal pressure can then by used in recognizing when an abnormally high intra-abdominal pressure or abdominal compartment syndrome exists. PMID- 11270884 TI - The effects of triiodothyronine augmentation on antithrombin III levels in sepsis. AB - Sepsis and multisystem organ failure are often associated with disseminated intravascular coagulation and consumption of coagulation inhibitors such as antithrombin III (ATIII). The "sick euthyroid syndrome" is also seen in association with significant illnesses and consists of decreased levels of circulating triiodothyronine (T3). We evaluated whether T3 supplementation would affect ATIII levels in septic rats. Thirty Sprague-Dawley rats were divided into three groups: sham laparotomy (S) plus saline, cecal ligation and puncture (CLP) plus saline, and CLP plus T3 (3 ng/hour) via an osmotic minipump. Twenty-four hours after laparotomy blood was drawn, and T3 and ATIII levels were then compared with baseline values. T3 supplementation partially negated the sepsis induced decrease in circulating T3 levels. The levels are expressed as percentage change from the levels before surgery (S, -12.9 +/- 3.1; CLP, -60.0 +/- 5.3; CLP + T3, -34.9 +/- 4.3; mean +/- standard error; P < 0.05). T3 supplementation also statistically changed the percentage difference in ATIII levels toward the control (S, 9.6 +/- 2.8; CLP, -37.9 +/- 5.4; CLP + T3, -16.0 +/- 4.5; mean +/- standard error; P < 0.01). T3 supplementation reduced the sepsis-induced decrease in ATIII levels. Whether this was accomplished by decreased consumption or increased production of ATIII via the direct anabolic effect of T3 on acute-phase protein synthesis in the liver is unknown and warrants further investigation. PMID- 11270883 TI - Evaluation of serum calcium levels in predicting hypoparathyroidism after total/near-total thyroidectomy or parathyroidectomy. AB - Hospital stays for thyroid and parathyroid surgery have decreased significantly with selected patients staying under 8 hours. Strategies to recognize hypocalcemia postoperatively vary. We examined timed postoperative calcium levels to determine how long one needs to monitor patients for hypoparathyroidism. We analyzed 120 consecutive patients having total/near-total thyroidectomy and/or parathyroidectomy between April 1998 and October 1999. Total and ionized serum calcium levels were obtained at 8, 16, and 22 hours postoperatively. Strict criteria for significant hypoparathyroidism were defined as a symptomatic patient, a total calcium value of less than 7.2 mg/dL, or an ionized calcium value of less than 1.0 mmol/L. Eighteen patients (15%) met criteria for hypocalcemia. The 8-hour ionized calcium level identified 40 per cent of those that needed supplementation. With the inclusion of the 16-hour ionized calcium value 94.5 per cent of patients who met criteria were identified. Of the 74 patients who had not previously received calcium at 22 hours after surgery only one patient with hypocalcemia was identified. Serial calcium values postoperatively add to the costs associated with an overnight hospital stay. In addition to clinical examination an ionized calcium level 16 hours postoperatively is sufficient to identify significant hypoparathyroidism in the majority of patients. PMID- 11270886 TI - Acute colonic perforation associated with colorectal cancer. AB - Our purpose was to evaluate long-term outcome in patients presenting with acute colonic perforation in the setting of colorectal cancer. We conducted a retrospective review of 48 consecutive patients presenting with acute colonic perforation associated with colorectal cancer at a single institution. Patients presented either with free air or acute peritonitis. No patients with colonic obstruction were included. Forty-eight patients presented with colon perforation. Thirty-six had perforation at the tumor, 11 proximal to the tumor, and one distal to the primary tumor. Patients who perforated proximal to the tumor were older (74.5 +/- 2 vs 64.7 +/- 3; P < 0.04) and had a longer length of stay (46.8 +/- 17 vs 11.6 +/- 1 P < 0.001). Fourteen patients had stage II disease, 19 stage III, and 15 stage IV. Thirty-day mortality was 14 per cent (n = 7) with nine in hospital deaths. Of 30-day survivors 29 (60%) had curative resection (21 with local perforation and nine with proximal perforation). Of these 14 received adjuvant chemotherapy. Eleven patients (33%) had either unresectable or metastatic disease on exploration. Mean follow-up was 21.5 months. Ten patients developed metastatic disease after potentially curative resections. Of these nine patients had perforations of the primary tumor. Three patients developed local recurrence and all had local tumor perforations. One-year survival was 55 per cent (n = 16). Five-year disease-free survival was 14 per cent (n = 4). There were no long-term survivors after perforation proximal to the tumor, although disease stage was comparable in both groups. We conclude that perforation proximal to a cancer is associated with a higher perioperative mortality and worse long-term outcome when compared with acute perforations at the site of the tumor. Long-term survival requires both aggressive management of the concomitant sepsis and definitive oncologic surgery. PMID- 11270885 TI - Interferon alpha2b inhibits the murine melanoma cell line Cloudman S91 in vivo but not in vitro: a model for studying tumor cell-cytokine interactions. AB - Interferon alpha2b has recently been shown to improve outcome in patients with metastatic malignant melanoma. The high-dose interferon therapy used is however associated with significant systemic adverse effects. These adverse effects are likely related to the multitude of actions of interferon which in addition to its antineoplastic effects also possesses antiviral and immunomodulating properties. Elucidation of the mechanism of the antiproliferative effects of interferon may allow for the development of agents that possess the antineoplastic properties while being devoid of the other effects that make interferon toxic. In the animal model developed for this study tumors in mice receiving interferon alpha2b grew at a slower rate and achieved a small final tumor volume (3040 +/- 690 vs 1400 +/ 314 mm3 for the control and treated groups respectively, P < 0.05). Furthermore the final tumor weight in the treated group was significantly smaller (1.50 +/- 0.21 g vs 2.76 +/- 0.46 g for the treated and control groups respectively; P = 0.036). The (3-[4,5-Dimethylthiazol-2-y]-2,5-diphenyltetrazolium bromide) (MTT) colorimetric assay failed to reveal any direct effects of interferon alpha2b on this murine melanoma cell line. This antiproliferative effect of interferon alpha2b was in addition found to be independent of alterations in the expression of the angiogenic cytokines vascular endothelial growth factor, basic fibroblast growth factor, and transforming growth factor beta. PMID- 11270887 TI - Retrospective study of neostigmine for the treatment of acute colonic pseudo obstruction. AB - Acute colonic pseudo-obstruction (ACPO) typically develops postoperatively or after severe illness. Studies suggest that pharmacologic manipulation with intravenous (i.v.) neostigmine (NSM) may be an effective and less invasive treatment modality for ACPO with minimal side effects. The purpose of this study was to retrospectively assess the efficacy and incidence of complications of an i.v. NSM bolus in patients with ACPO. Eight patients with ten episodes of ACPO were treated with a bolus dose of NSM. Rapid and effective decompression of the colon was achieved in six episodes after a single dose of NSM at a mean of 22.8 +/- 13.5 minutes. In three episodes decompression occurred after a second dose of NSM at a mean of 44.7 +/- 37.7 minutes. One patient failed NSM treatment but responded to a Cystografin enema. One patient experienced significant bradycardia. NSM is a simple, safe, and effective treatment for ACPO and based on result comparison of this study and previous studies both bolus and slow infusion dosing practices of NSM are effective. The NSM bolus dosing side effect profile has been shown to include significant bradycardia, whereas when NSM was infused over one hour significant bradycardic episodes requiring treatment have not been encountered. We propose that a prospective study evaluating NSM dosing as an i.v. bolus versus an i.v. infusion would be useful in determining whether NSM infusion can be proven safer than bolus dosing for the treatment of ACPO. PMID- 11270888 TI - Age is not a contraindication to pancreaticoduodenectomy. AB - The incidence of pancreatic cancer has increased threefold over the last 40 years with the greatest rate of growth occurring in the elderly. In the past it was suggested that elderly patients tolerated pancreaticoduodenectomy less well than younger patients with higher mortality rates. This single-institution experience examines the question of whether age is a significant factor in relation to morbidity and mortality in patients undergoing pancreaticoduodenectomy. Between 1994 and 1999 outcomes of 122 patients who underwent pancreaticoduodenectomy were reviewed. There were 48 patients 70 years of age and older and 74 patients less than 70 years of age. Both groups were compared with respect to preoperative clinical prognostic determinates and perioperative factors affecting morbidity and mortality. There was no significant difference between the two groups comparing their comorbidities, use of preoperative antibiotics, intraoperative blood loss, or length of hospital stay (11.9 and 10.8 days respectively). The two groups were also similar with regard to pathologic diagnosis with pancreatic adenocarcinoma being the most frequently encountered neoplasm. There was one death in the less-than-70-year-old group and none in the older group. No significant difference in the rate of complications was appreciated. These data demonstrate that pancreaticoduodenectomy can be performed safely in patients 70 years of age and older with morbidity and mortality rates similar to those of younger individuals. PMID- 11270890 TI - A prospective study of the mesh-plug hernioplasty. AB - A prospective study of patients with symptomatic inguinal hernias was undertaken to determine the safety and efficacy of the mesh-plug hernioplasty. Between May 1, 1997 and March 1, 1999 a total of 309 mesh-plug hernioplasties were performed on 283 patients. There were 43 recurrent and 26 bilateral hernioplasties. There were 273 men and 10 women ranging in age from 15 to 94 years (mean 47 years). There were 199 indirect, 104 direct, and six femoral hernias. Mean operative time for primary hernioplasty was 26 minutes (range 20-34) and 35 minutes (range 31 40) for recurrent hernioplasty. All procedures were performed as outpatient surgery with mean recovery room time being 45 minutes (range 25-27) for primary hernioplasty. Two hundred sixty-six patients (94%) returned to normal activities within 3 days. All manual laborers (124 patients) returned to work without restriction on postoperative day 14. Only 43 patients (15%) required prescription pain medication. At one year postoperatively 283 patients (100% follow-up) have been examined and no recurrence has been detected. At 2 years postoperatively 135 patients (100% follow-up) have been examined and no recurrence has been detected. The mesh-plug hernioplasty uses a minimum of medical resources, is associated with a small amount of postoperative pain, and has an early return to normal activities and manual labor without a documented early recurrence in this study. PMID- 11270889 TI - Trimodality therapy for advanced gallbladder cancer. AB - We conducted a retrospective review of all patients who underwent surgical extirpation for stage III, stage IV, or recurrent carcinoma of the gallbladder. Between 1991 and 1999 ten patients underwent surgical resection for advanced gallbladder cancer. All patients received adjuvant therapy either pre- or postoperatively. Radiotherapy was used in all patients and chemotherapy in 90 per cent of patients. Two patients subsequently underwent resection for locally recurrent disease. An additional patient with stage II disease initially was also treated surgically for a local recurrence. Surgical management involved cholecystectomy and resection of various amounts of liver surrounding the gallbladder bed and regional lymphadenectomy. Contiguously involved structures were resected en bloc. Resection of recurrent disease included excision of all gross tumor. The median overall survival excluding the one 30-day mortality was 53.6 months (range 8-73 months). Four patients have survived 4 or more years, and currently four patients are alive and disease free at 73, 49, 33, and 8 months. Median disease-free interval after each resection of recurrent disease was 13.8 months (range 4-28 months). We conclude that trimodality therapy in selected patients with stage III, IV, or recurrent carcinoma of the gallbladder is possible and may result in prolonged survival. PMID- 11270891 TI - Risk of parathyromatosis after fine-needle aspiration. AB - Reoperative surgery for hyperparathyroidism (HPT) is fraught with hazard. When preoperative imaging studies are inconclusive or patient comorbidities are extensive fine needle-aspiration (FNA) is helpful to confirm the presence of suspected parathyroid tissue in the neck. Some surgeons refrain from using FNA because of the concern of tissue implantation (parathyromatosis). A retrospective review (1984-1996) of all patients diagnosed with HPT undergoing FNA of suspected parathyroid tissue was performed to document whether a correlation exists between FNA of suspected parathyroid tissue and subsequent development of parathyromatosis. Parathyromatosis was considered to have occurred when proven by histology or suspected on the basis of clinical studies. Of 81 patients with HPT undergoing ultrasound-guided FNA to assess abnormalities in the neck 41 patients with confirmed parathyroid tissue were identified. The indications for FNA in these 41 patients were: prior failed cervical exploration (n = 33), prior neck surgery and/or radiation (n = 2), inconclusive noninvasive imaging studies (n = 15), and severe comorbidities (n = 8). Mean follow-up was 5.8 years. No case of FNA-induced parathyromatosis was identified. FNA is useful to confirm the presence of parathyroid tissue in very select patients with hyperparathyroidism. FNA often eliminates the need for other imaging studies, may prevent a needless or likely fruitless re-exploration, and does not cause parathyromatosis. PMID- 11270892 TI - Moral issues in day-to-day palliative medicine and their relevance for the education of European general practitioners. AB - BACKGROUND: Considerations of moral problems in palliative medicine often deal with extreme situations. This study identified moral issues arising in routine palliative medicine. Their relevance for the education of European general practitioners is assessed. METHODS: Consecutive consultations of cancer patients with incurable disease were recorded in three outpatient clinics and one general practice in Belgium. Moral issues were identified by qualitative analysis of verbal transcripts of 30 of these consultations using the grounded-theory approach. The relevance of these issues for medical education was assessed by interviewing one educator of general practitioners from each of the 15 European Union states. RESULTS: Three core categories of moral issues were identified: telling the truth, patient control versus medical dominance, and handling the patient's life-world. The practical relevance of these issues was recognized by the educators. The suggested educational methods to deal with these topics were all active learning processes in small-group settings but varied otherwise. CONCLUSIONS: The moral issues identified in day-to-day palliative medicine may complement the problems evoked in the literature dealing with more extreme situations. An effort to study the appropriate way for medical education to deal with these topics may be indicated. PMID- 11270893 TI - A unique approach to multidisciplinary cancer education. AB - BACKGROUND: Almost every health care worker, regardless of specialty, interacts with individuals who have histories of cancer. Some health care workers are relatively unfamiliar with the cancer experience and those who are familiar with it are interested in additional information. METHOD: The Mayo Clinic Cancer Center Education Subcommittee designed a monthly educational program titled Cancer Connections: A Multidisciplinary Update. The goals of this program were to provide: 1) up-to-date information about cancer issues, 2) a better understanding of various team members' roles in caring for people with cancer, 3) insight into patients' responses to the diagnosis and treatment of their cancers, and 4) greater awareness of the resources available to cancer patients and their families. Each session consists of a "panel" of presenters: three healthcare professionals and a patient. RESULTS AND DISCUSSION: The success of this program is reflected in the regular high attendance and consistently positive evaluations of the participants. PMID- 11270894 TI - Effectiveness of an oral and pharyngeal cancer awareness program for health professionals. AB - BACKGROUND: Of approximately 31,000 patients diagnosed as having oral/pharyngeal cancers (OPCs) each year in the United States, about half will die of the disease within five years, for reasons associated with patient behaviors as well as delays in diagnosis by health care professionals. METHODS: To address an apparent lack of OPC knowledge of health care professionals, a brief, non-discipline specific, multi-component educational intervention was designed and presented to 352 health care professionals in community health centers and hospital training programs. OPC knowledge was assessed before and three months after the intervention. RESULTS: A 44% post-intervention response rate was obtained. There were significant (p < or = 0.05) increases in knowledge regarding oral sites at risk for OPC, etiologic factors, and early signs and symptoms. There were significant interactions (p < or = 0.01) between increases in knowledge and various health provider groups. A significant increase in perceived competency in OPC knowledge (p < or = 0.001) was reported, and significantly fewer participants felt the need for additional training. CONCLUSION: A brief, multi-component educational intervention can increase health care professionals' knowledge regarding OPC. PMID- 11270895 TI - Can assessment of psychosocial orientation assist continuing education program development in psychosocial oncology? AB - BACKGROUND: A pilot study was designed to aid in the development of a formal, interdisciplinary curriculum in psychosocial oncology for front-line health care professionals. METHOD: A 190-item questionnaire was distributed to psychosocial (PP) and non-psychosocial (NPP) oncology professionals attending a psychosocial skills workshop. A 38-item attitudinal survey of psychosocial orientation was used in an attempt to identify unperceived needs of the learners. RESULTS: Of the 150 questionnaires distributed, 104 (69%) were completed and returned. Overall scores for satisfaction with the workshop were high, and significantly higher in the PP group. No interdisciplinary difference existed in the preferred learning formats for future events, and both groups preferred interactive, experiential forums for developing skills relevant to patient management. The two groups' perceived learning needs differed. NPPs wanted to focus on skills such as communication, counseling, crisis intervention, palliative care, and coping with life-threatening illness. The attitudinal survey results demonstrated a significant difference between the psychosocial orientations of PPs and NPPs and suggested that NPPs would benefit from: 1) information to correct misconceptions about patients' psychosocial needs and experiences, 2) demonstrations of how to overcome contextual barriers to the delivery of psychosocial care. CONCLUSIONS: Front-line oncology professionals in many disciplines are interested in continuing education in psychosocial oncology. The attitudinal survey provided insight into unperceived learning needs that can help in designing future curricula. Its value as a tool to measure impact of these programs is worthy of future study. PMID- 11270896 TI - Age bias: a cause of underutilization of breast conservation treatment. AB - BACKGROUND: Breast conservation therapy (BCT) has been shown to result i about the same disease control and survival as modified radical mastectomy (MRM) for stage I and II breast cancers. Barriers to using BCT in patients with invasive breast cancer include "physician preference." This study was undertaken to investigate the bias of residents with respect to breast-conserving procedures. METHODS: Internal medicine and surgery residents were instructed about the efficacy of BCT. Subsequently, their opinions were assessed using a questionnaire concerning recommendations for BCT versus MRM as well as breast reconstruction after MRM in similar patients. Chi square tests were used for statistical analysis. RESULTS: Seventy-nine residents (54 medical, 25 surgical) participated. MRM was recommended for 38% of older (> 59 years old) versus 11% of younger patients (< 31 years old), p < 0.01. Furthermore, breast reconstruction was recommended for 96% of younger versus only 70% of older patients (p < 0.01). CONCLUSIONS: Residents are biased against older women in their recommendations for breast conservation and breast reconstruction. Educational efforts to decrease this age bias should be instituted during residency. PMID- 11270897 TI - Colorado family physicians' knowledge of hereditary breast cancer and related practice. AB - BACKGROUND: The objective of this study was to describe Colorado family physicians' (FPs') knowledge of hereditary breast cancer and related practice behaviors. METHOD: A survey was mailed to 400 practicing FPs randomly sampled from the board-certified members of the Colorado Academy of Family physicians. RESULTS: Overall, knowledge of hereditary breast cancer was inadequate. Less than half of the respondents knew that BRCA mutations account for 0-10% of all breast cancers; 38% knew the lifetime risk for non-carriers, and 17% responded that a known carrier would have a lifetime breast cancer risk of 50%. Just 45% knew that a BRCA1 mutation could pass from father to daughter. Similarly, only half reported an increased ovarian cancer risk for BRCA1 carriers, and an increased risk for male breast cancer for BRCA2 carriers. All respondents reported that taking a family cancer history was part of their regular clinical practices. The majority reported having referred no patient for cancer genetic counseling or testing within the prior year, with only two reporting having ordered BRCA1 or BRCA2 testing within the year. There was no significant sociodemographic or knowledge difference between the physicians who reported referring patients and the others (p > 0.05). There was interest in learning more about hereditary breast cancer, with rural physicians requesting Internet and teleconference courses. CONCLUSIONS: As cancer genetics emerges into the primary care arena. FPs recognize their knowledge deficit in this area. Future cancer genetic outreach for primary care providers statewide is necessary and would be welcomed, and may require a variety of educational and consultative approaches, depending on geographic location of practice. PMID- 11270898 TI - Are we ordering too many PSA tests? Prostate cancer diagnosis and PSA screening patterns for a single Veterans Affairs Medical Center. AB - BACKGROUND: Limits on the frequency of PSA testing and an endpoint for the age of the screened population have not been established. The numbers of performed serum PSA tests, cost evolution, and utilization patterns by various subspecialties in one medical center were analyzed to gain insight into trends in screening for early detection of prostate cancer and gather information about the appropriate use of PSA testing. METHOD: Computerized records were reviewed for numbers of PSA tests obtained, prostate biopsies performed, and prostate cancer cases diagnosed in the VA NJ-Health Care System from 1996 to 1998. In addition, PSA tests performed during two representative weeks in 1996 and 1997 were analyzed to evaluate a smaller cohort of patients with regard to age, consequences of the test results in their management, and subspecialties ordering the tests. RESULTS: PSA testing increased steadily between 1992 and 1998, with the most significant change (152% increase) between 1997 (9,410 tests) and 1998 (23,684). Prostate cancer diagnoses by biopsy were 164/434 (37.8%) in 1997 and 195/507 (38.5%) in 1998. For the 14,274 additional PSA tests obtained in 1998, 31 more prostate cancers were diagnosed. Prostate cancer diagnoses per PSA tests were 164/9,410 (1.8%) in 1997 and 195/23,684 (0.8%) in 1998. Primary care providers ordered 61% of the PSA tests. CONCLUSIONS: Most PSA tests at this institution were ordered by general practitioners, and the number of PSA tests ordered for men over 75 was high. The dramatic increase between 1997 and 1998 was not accompanied by a similar rise in the diagnosis of prostate cancer, raising the possibility of indiscriminate PSA testing or unnecessary repetition of testing. Guidelines for prostate cancer screening and continued PSA testing in the geriatric population may need further clarification. PMID- 11270899 TI - Men's attitudes toward prostate cancer and seeking prostate-specific antigen testing. AB - BACKGROUND: Although the Australian Cancer Society recommends against performing PSA tests to screen for prostate cancer, many Australian men currently undergo such screening. This study investigated attitudinal variables that may predict prostate cancer screening behaviors in this context. METHODS: A questionnaire was administered by mail in a two-phase procedure, first to a sample of 1,461 men (46% response), then to 919 men from the initial sample. Prostate cancer screening behaviors of men > 40 years old were examined. The questionnaire assessed worry about prostate cancer, perceived vulnerability to prostate cancer, belief in the efficacy of PSA testing for detection, having received a PSA test for detection, and the presence of urologic symptoms at the time of testing. RESULTS: Men who had had PSA testing with urologic symptoms at the time of the test were more worried about prostate cancer and perceived themselves as more vulnerable to prostate cancer compared with both asymptomatic tested and untested men. Men who had undergone PSA testing believed the test to be more effective in the detection of prostate cancer than did men who had not. CONCLUSIONS: Urologic symptoms act as a risk cue for men to prostate cancer. Asymptomatic men should be considered separately from symptomatic men in the investigation of psychological variables predictive of seeking screening for prostate cancer. These findings are discussed in terms of both the focus and design of interventions to alter prostate cancer screening behavior and their implications for the clinical management of men with urologic symptoms. PMID- 11270900 TI - Identifying women at risk for inherited breast cancer using a mammography registry. AB - BACKGROUND: Women at risk for inherited breast cancer have been identified for intervention studies through newly diagnosed relatives or from volunteers with a family history. This pilot study tested the use of a mammography registry to identify women at risk. METHOD: Fifty women with first-degree relatives diagnosed as having breast cancer before age 45 were randomly selected from the Vermont Breast Cancer Surveillance System. Thirty-three women (66%) completed a phone interview that included a three-generation family pedigree of breast and ovarian cancers. RESULTS: Fifty-one percent of the women were at higher risk for inherited breast cancer based on the family history. Eighteen percent of the first-degree relatives' breast cancers were pathologically confirmed. CONCLUSION: Mammography registries that collect similar family history data may be used to identify women at risk for inherited breast cancer. Many intervention studies would require improved methods to obtain pathologic confirmation. PMID- 11270901 TI - Breast cancer screening among Hmong women in California. AB - BACKGROUND: Breast cancer is the leading cause of cancer incidence and mortality among Asian American and Pacific Islander (AAPI) women in the United States. Hmong women are among those at the highest risk for health problems, due to high rates of poverty, language isolation, and cultural barriers. METHOD: One-on-one survey interviews were completed with 201 Hmong women aged 20 years and older in Fresno, Long Beach, Orange County, and San Diego to determine their breast cancer screening behaviors--breast self-examination (BSE), clinical breast examination (CBE), and mammography. RESULTS: Overall, 51% of all respondents had ever performed BSE. Among respondents aged 40 or older, 52% had ever had a CBE and only 30% had ever had mammography. DISCUSSION: Significant correlates of CBE and mammography screening are presented, and implications of findings for research and education are discussed. PMID- 11270902 TI - Two community outreach strategies to increase breast cancer screening among low income women. AB - BACKGROUND: Two approaches were designed for low-income women to promote their use of mammography screening. METHODS: During 1995-96, as part of a community outreach project in a Florida city, 1,157 women aged 45 years or older attended group education sessions on breast cancer screening, while another 1,450 participated in one-on-one talks about screening at display tables in various public places. County mammography registry data were used to assess changes in the use of mammography screening. RESULTS: Among women 55 years old or older, especially whites, the one-on-one approach was more often associated with subsequent mammography screening than was the group approach. African American women and Latina women appeared to benefit more from the group approach than from the one-on-one approach. CONCLUSIONS: Group or one-on-one breast cancer screening education can improve screening behaviors among low-income women, depending on the age and ethnicity of the women targeted. PMID- 11270903 TI - Are you having fun? PMID- 11270904 TI - Decision making in cancer education. PMID- 11270905 TI - The impact of a cancer education program on the knowledge base of participating students. AB - BACKGROUND: Partners in Research is a ten-week summer elective designed to provide cancer-related educational activities. This study was undertaken to evaluate the impact of the program on the general cancer knowledge of medical students, pharmacy students, and undergraduate biology majors. METHODS: The 24 students enrolled in 1999 were evaluated using a pretest and post-test with 75 multiple-choice questions. RESULTS: The mean test score increased significantly from 46.6% to 53.0% (p = 0.001). Improvements were significant for general cancer knowledge and three specific disease categories (breast, gastrointestinal, and skin cancers). CONCLUSIONS: The results indicate that the program does increase the cancer-related knowledge of students. PMID- 11270906 TI - The placebo enigma in antidepressant clinical trials. PMID- 11270907 TI - Are lorazepam-induced deficits in attention similar to those resulting from aging? AB - The purpose of this experiment was to compare, in three tasks of attention, the impairment caused by lorazepam (1 and 2.5 mg) administered to young volunteers with the impairment that results from aging. Performance on digit cancellation (DC), digit-symbol substitution (DSS), and Paced Auditory Serial Addition Task (PASAT) was significantly impaired by lorazepam (2.5 mg) and was significantly worse in the middle-aged group (mean +/- SEM, aged 58.9+/-0.8 years) compared with the younger, IQ-matched group (20.7+/-0.2 years). However, there were interesting differences in the extent of impairments among the three tests. In the DC test, lorazepam (2.5 mg) produced a significantly greater impairment than was seen in either the middle-aged men or middle-aged women. However, in the DSS test, the middle-aged women were significantly more impaired than either the middle-aged men or the young volunteers tested after lorazepam (2.5 mg). In the PASAT, both the lorazepam (2.5 mg) group and the middle-aged women were more impaired than the middle-aged men. These results raise the important possibility of gender differences in age-related decline of attentional processes. PMID- 11270908 TI - Absence of neuropsychologic deficits in patients receiving long-term treatment with alprazolam-XR for panic disorder. AB - Studies to date on the effects of benzodiazepines on neuropsychologic function have yielded conflicting data with respect to the type, severity, and duration of deficits that may be induced by these agents. As part of a placebo-controlled trial of alprazolam-XR (extended release) administered in combination with cognitive-behavioral therapy in patients with panic disorder, a battery of tests was used to measure neuropsychologic function. Thirty-eight outpatients were randomly assigned to receive either alprazolam-XR or placebo. Dosages were titrated up so that the alprazolam group (N = 18) received a mean dose of 4 mg/day (reduced in two patients because of sedative side effects). Neuropsychologic function after 6 weeks of therapy at the target dosage was compared with baseline assessments in each group. Both groups showed a statistically significant improvement from baseline to repeated assessments on measures of attention, executive functioning, psychomotor speed, and visual memory (p < 0.001); these gains were attributed to a practice effect. No significant changes were noted in measures of learning, verbal memory, or reaction time, and neither group showed any deterioration from baseline to retesting in any aspect of neuropsychologic function. These findings call into question the assumption that long-term benzodiazepine therapy produces significant neuropsychologic deficit in patients with diagnosed anxiety disorders. PMID- 11270909 TI - Fluvoxamine treatment of mixed anxiety and depression: evidence for serotonergically mediated anxiolysis. AB - Although increasing evidence suggests that selective serotonin reuptake inhibitor (SSRI) treatment may be effective for anxiety in addition to depression, SSRI anxiolysis has not been definitively related to the inhibition of serotonin (5 HT) transport. The gene that encodes for the human serotonin transporter (5-HTT) expresses its protein in neurons and in blood platelets, and both tissues respond to transport inhibition similarly in response to SSRI treatment. This study examined the relationship between the change in the 5-HTT's apparent affinity for 5-HT and the anxiolytic response in a group of 18 fluvoxamine-treated patients meeting Structured Clinical Interview for DSM-IV criteria for both generalized anxiety disorder and major depression. Significant decreases were found in both Hamilton anxiety and Hamilton depression scores over a 2-month treatment period. Robust increases were found in the apparent affinity constant (Km) for platelet 5 HT transport with treatment, and the increases covaried significantly with the decrease in anxiety (F = 4.97, p < 0.03). The pretreatment 5-HTT Km significantly correlated with the improvement in depression scores (r = 0.53, p < 0.03), consistent with the Hypothesis of Initial Conditions. These results suggest that the therapeutic effect of SSRI treatment can be linked to the magnitude and time course of 5-HT transport inhibition effected with fluvoxamine, a drug that seems to have an antianxiety effect of the same magnitude as its effect on depression. PMID- 11270910 TI - Double-blind clinical trial of sertraline treatment for alcohol dependence. AB - Clinical studies that have evaluated serotonergic medications to reduce alcohol consumption have yielded conflicting results. These studies primarily treated patients with alcohol dependence, excluding those with a current depressive disorder, in an effort to differentiate any medication effects directly on drinking from those on mood. Yet despite the exclusion of current depression, a group of alcohol-dependent patients who are not depressed can be highly heterogeneous. For example, this subgroup can include those with a lifetime depressive disorder. If these patients were more sensitive to serotonergic medications than patients without a lifetime depressive disorder, medication effects in a subgroup of patients who were not depressed could be obscured. Thus, the purpose of this study was to examine the efficacy of sertraline for treating alcohol dependence in patient groups that were differentiated by the presence or absence of lifetime depression. This study examined the effectiveness of sertraline (200 mg/day) or placebo for 14 weeks in 100 alcohol-dependent subjects with (N = 53) or without (N = 47) a lifetime diagnosis of comorbid depression. Sertraline treatment seemed to provide an advantage in reducing drinking in alcohol-dependent patients without lifetime depression, illustrated best with a measure of drinking frequency during treatment. However, sertraline was no better than placebo in patients with a diagnosis of lifetime comorbid depression, and current depression did not change the results. Treatment with selective serotonin reuptake inhibitors may be useful in alcohol-dependent patients who are not depressed. Subtyping those with alcohol dependence on the basis of the absence versus the presence of a lifetime depressive disorder may help to resolve conflicting findings in the literature on the treatment of alcohol dependence with serotonergic medications. PMID- 11270911 TI - Effect on sexual function of long-term treatment with selective serotonin reuptake inhibitors in depressed patients treated in primary care. AB - The purpose of this study was to prospectively examine the occurrence and severity of sexual dysfunction symptoms in depressed patients before and after 6 months of treatment with selective serotonin reuptake inhibitors. The study was part of a randomized, double-blind, controlled trial of sertraline or citalopram in patients with a DSM-III-R major depressive disorder treated by general practitioners. Three hundred eight patients (221 women and 87 men) were assessed at baseline and after 6 months of treatment by means of the Montgomery-Asberg Depression Rating Scale and five items from the Utvalg for Kliniske Undersogelser (UKU) Side Effect Scale covering different aspects of sexual functioning. As measured by the UKU Side Effect Scale, sexual desire and mean total score significantly improved in women, and sexual desire improved in men. Men reported no change in orgasmic dysfunction, erectile dysfunction, or mean total score, but there was a trend toward worsening of ejaculatory dysfunction. However, in the subgroup of women who reported no sexual problems at baseline, 11.8% reported decreased sexual desire, and 14.3% reported orgasmic dysfunction at week 24. The corresponding figures in the same subgroup of men were 16.7% and 18.9%, respectively, and as many as 25% experienced ejaculatory dysfunction after 24 weeks. There were no statistically significant differences between sertraline and citalopram in the magnitude or frequency of adverse sexual side effects. PMID- 11270912 TI - Fluoxetine pharmacokinetics and effect on CYP2C19 in young and elderly volunteers. AB - The objective of this study was to assess in both young and elderly volunteers the pharmacokinetics of fluoxetine and norfluoxetine and effects on cytochrome P450 (CYP) 2C19. Male volunteers aged 18 to 40 years (N = 14) or older than 65 years (N = 16) received fluoxetine 20 mg/day for 6 weeks and fluoxetine 40 mg/day for an additional 6 weeks. Blood was drawn over a 24-hour period after the initial dose and after 6 weeks and 12 weeks to determine AUC0-24, Cmax, and tmax; weekly to evaluate predose levels (C0); and over a 3-week period after discontinuation to evaluate washout (t1/2). Mephenytoin was used to assess CYP2C19 activity before and after 6 weeks and 12 weeks of fluoxetine. Fluoxetine AUC0-24, C0, and Cmax did not differ in young and elderly subjects. The norfluoxetine C0 was 22% lower in elderly subjects (p < .05), with comparable decreases in AUC0-24 and Cmax. In the elderly volunteers, the t1/2 for fluoxetine was 25% longer (5.0 vs. 4.0 days) and for norfluoxetine was 33% longer (20 vs. 15 days), although variability and sample size precluded statistical significance. Fluoxetine dosing inhibited CYP2C19 activity in both age groups, increasing the (S)- to (R)-mephenytoin ratio 3- to 4-fold (p < .01). The half-lives of fluoxetine and norfluoxetine at 40 mg/day were longer than commonly reported in the literature and may be longer in elderly subjects. Fluoxetine substantially inhibited the metabolism of the CYP2C19 substrate (S)-mephenytoin. PMID- 11270913 TI - Differential effects of fluvoxamine and other antidepressants on the biotransformation of melatonin. AB - Melatonin, the predominant product of the pineal gland, is involved in the maintenance of diurnal rhythms. Nocturnal blood concentrations of melatonin have been shown to be enhanced by fluvoxamine, but not by other serotonin reuptake inhibitors. Because fluvoxamine is an inhibitor of several cytochrome P450 (CYP) enzymes, the authors studied the biotransformation of melatonin and the effects of fluvoxamine on the metabolism of melatonin in vitro using human liver microsomes and recombinant human CYP isoenzymes. Melatonin was found to be almost exclusively metabolized by CYP1A2 to 6-hydroxymelatonin and N-acetylserotonin with a minimal contribution of CYP2C19. Both reactions were potently inhibited by fluvoxamine, with a Ki of 0.02 microM for the formation of 6-hydroxymelatonin and 0.05 microM for the formation of N-acetylserotonin. Other than fluvoxamine, fluoxetine, paroxetine, citalopram, imipramine, and desipramine were also tested at 2 and 20 microM. Among the other antidepressants, only paroxetine was able to affect the metabolism of melatonin at supratherapeutic concentrations of 20 microM, which did not reach by far the magnitude of the inhibitory potency of fluvoxamine. The authors concluded that fluvoxamine is a potent inhibitor of melatonin degradation. Because this inhibitory action is also found in vivo, fluvoxamine might be used as an enhancer of melatonin, which might offer new therapeutic possibilities of fluvoxamine. PMID- 11270914 TI - Diphenhydramine alters the disposition of venlafaxine through inhibition of CYP2D6 activity in humans. AB - CYP2D6 is the major enzyme involved in the metabolism of venlafaxine. Subjects with a low CYP2D6 activity have increased plasma concentrations of venlafaxine that may predispose them to cardiovascular side effects. In vitro and in vivo studies showed that diphenhydramine, a nonprescription antihistamine, can inhibit CYP2D6 activity. Therefore, the authors investigated in this study a potential drug interaction between diphenhydramine and venlafaxine. Fifteen male volunteers, nine with the extensive metabolizer (EM) and six with the poor metabolizer (PM) phenotype of CYP2D6, received venlafaxine hydrochloride 18.75 mg orally every 12 hours for 48 hours on two occasions (1 week apart): once alone and once during the concomitant administration of diphenhydramine hydrochloride (50 mg every 12 hours). Blood and urine samples were collected for 12 hours under steady-state conditions. In EMs, diphenhydramine decreased venlafaxine oral clearance from 104+/-60 L/hr to 43+/-23 L/hr (mean +/- SD; p < 0.05) without any effect on renal clearance (4+/-1 L/hr during venlafaxine alone and 4+/-2 L/hr during venlafaxine plus diphenhydramine). In PMs, coadministration of diphenhydramine did not cause significant changes in oral clearance and partial metabolic clearances of venlafaxine to its various metabolites. Diphenhydramine disposition was only slightly affected by genetically determined low CYP2D6 activity or concomitant administration of venlafaxine. In conclusion, diphenhydramine, at therapeutic doses, inhibits CYP2D6-mediated metabolism of venlafaxine in humans. Clinically significant interactions could be encountered during the concomitant administration of diphenhydramine and other antidepressant or antipsychotic drugs that are substrates of CYP2D6. PMID- 11270916 TI - Changes in quantitatively assessed tremor during treatment of major depression with lithium augmented by paroxetine or amitriptyline. AB - Tremor is a relatively frequent side effect of lithium and of antidepressants with serotonergic properties. It can be expected that combinations of lithium (which is itself serotonergic) with such antidepressants will enhance not only efficacy, but also the incidence of side effects, including tremor. To quantitatively monitor the effect of antidepressant augmentation of ongoing lithium therapy on tremor, lithium-maintained patients with a breakthrough episode of major depression were randomly assigned under double-blind conditions to receive paroxetine 20 mg/day (N = 14) or amitriptyline 75 mg/day (N = 17). The initial dosages could be increased after 2 weeks to 40 mg/day and 150 mg/day, respectively, and the patients were treated for 6 weeks. Tremor activity was assessed weekly, quantitatively by accelerometry and qualitatively with the Dosage Record and Treatment Emergent Symptom Scale. Statistical analysis detected no significant difference between the treatment groups with respect to changes in mean tremor activity relative to baseline. However, analysis of the pooled data showed that tremor increased significantly during the course of combined lithium and antidepressant therapy, with the greatest increments occurring independent of dosage approximately 3 weeks after initiation of combination treatment. Although the mean tremor activity subsided toward the end of treatment, tremor activity on the whole was still significantly greater after 6 weeks of combined lithium and antidepressant treatment than at the start of combination therapy. Increased tremor was not associated with decreased medication compliance, and no patient discontinued treatment because of increased tremor. Tremor frequency was not affected by the study treatments. PMID- 11270915 TI - Sustained response to open-label venlafaxine in drug-resistant major depression. AB - The aim of this study was to evaluate the response to venlafaxine in patients with treatment-resistant depression during an extension phase of an open-label study of venlafaxine. After completing the initial 8 weeks of the study, patients could continue venlafaxine treatment for an additional period of up to 10 months. Efficacy results are given for 149 patients with treatment-resistant depression. Response was defined as a 50% reduction in scores on the Montgomery-Asberg Depression Rating Scale (MADRS); 69% were responders after 8 weeks of treatment in the initial study phase, and 73% were responders at their final extension phase visit. The mean MADRS score was 32.8 before treatment, 12.9 by 8 weeks, and 10.8 at the final extension visit. There was a statistically significant reduction of 2.1 MADRS units from entry into the extension phase to the final extension visit. At extension entry, 36.7% patients were in remission, as defined by a MADRS score of less than 12, whereas at the final extension visit, this had increased to 49%. Improvement in Clinical Global Impressions Scale scores (both patient and physician ratings) was maintained throughout the extension period, with 88% of patients reporting some improvement (75% with "very much" or "much") and 92% of doctors noting some improvement in patients (79% with "very much" or "much") at the last extension visit. The safety profile during the extension phase of the study was similar to that found in the initial phase and in other studies. The most common study events were somnolence (21%), headache (18%), insomnia (16%), sweating (16%), constipation (14%), dry mouth (11%), nausea (10%), and dizziness (10%). Patients with resistant depression that was treated with venlafaxine maintained their response for up to 10 months after an 8-week phase of treatment and showed some evidence of further improvement. PMID- 11270917 TI - Anti-Inflammatory effects of antidepressants through suppression of the interferon-gamma/interleukin-10 production ratio. AB - There is some evidence that major depression--in particular, treatment-resistant depression (TRD)--is accompanied by activation of the inflammatory response system and that proinflammatory cytokines may play a role in the etiology of depression. This study was carried out to examine the effects of antidepressive agents, i.e., imipramine, venlafaxine, L-5-hydroxytryptophan, and fluoxetine on the production of interferon-gamma (IFN-gamma), a proinflammatory cytokine, and interleukin-10 (IL-10), a negative immunoregulatory cytokine. Diluted whole blood of fluoxetine-treated patients with TRD (mean age, 50.6+/-3.9 years) and age matched healthy controls (mean age, 51.6+/-1.7 years) and younger healthy volunteers (mean age, 35.4+/-9.6 years) was stimulated with phytohemagglutinin (1 microg/mL) and lipopolysaccharide (5 microg/mL) for 48 hours with and without incubation with the antidepressants at 10-6 M and 10(-5) M. IFN-gamma and IL-10 were quantified by means of enzyme-linked immunoassays. The ratio of IFN-gamma to IL-10 production by immunocytes was computed because this ratio is of critical importance in determining the capacity of immunocytes to activate or inhibit monocytic and T-lymphocytic functions. All four antidepressive drugs significantly increased the production of IL-10. Fluoxetine significantly decreased the production of IFN-gamma. All four antidepressants significantly reduced the IFN-gamma/IL-10 ratio. There were no significant differences in the antidepressant-induced changes in IFN-gamma or IL-10 between younger and older healthy volunteers and TRD patients. Tricyclic antidepressants, selective serotonin reuptake inhibitors, and serotonin-noradrenaline reuptake inhibitors, as well as the immediate precursor of serotonin, have a common, negative immunoregulatory effect by suppressing the IFN-gamma/IL-10 production ratio. It is suggested that the therapeutic efficacy of antidepressants may be related to their negative immunoregulatory effects. PMID- 11270919 TI - Relevance of liver enzyme elevations with four different neuroleptics: a retrospective review of 7,263 treatment courses. AB - Data on liver enzyme elevations were collected in a retrospective study of 7,263 treatment courses with haloperidol, clozapine, perphenazine, and perazine. Charts of 233 patients hospitalized between 1980 and 1992 at Tubingen University Psychiatric Clinic were selected because clinically relevant increases of liver enzymes had been detected during monotherapy with one of the four examined neuroleptics. At least one hepatic enzyme (mostly alanine aminotransferase [ALAT]) exceeded the established reference range of 3-fold elevations of ALAT, aspartate aminotransferase, gamma-glutamyl transpeptidase, and glutamate dehydrogenase and 2-fold elevations of alkaline phosphatase (AP) during monotherapy with clozapine in 15%, perazine in 7.6%, perphenazine in 4%, and haloperidol in 2.4% of the cases. If all liver enzyme abnormalities with any elevation greater than the conventional upper limits are considered, incidences were as follows: clozapine, 78%; perphenazine, 62%; perazine, 59%; and haloperidol, 50%. Testing for overall differences within the four neuroleptics resulted in significantly different incidences of liver enzyme elevations (chi2 test,p < 0.0001). Threefold increases of AP (>540 U/L) were seen in three patients receiving haloperidol (0.3%) only. Twofold increases of AP (>360 U/L) were distributed as follows: clozapine, 1%; haloperidol, 0.8%; perazine, 0.3%; and perphenazine, 0.1%. Only in the group with 1-fold elevations of AP (>180 U/L) were the differences within the drug regimens significant (clozapine, 40.3%; haloperidol, 33.2%; perphenazine, 23.4%; and perazine, 23.1%; chi2 test, p < 0.0001). In the period under study, no instance of icterus occurred. PMID- 11270918 TI - In vivo extrastriatal and striatal D2 dopamine receptor blockade by amisulpride in schizophrenia. AB - Amisulpride, a substituted benzamide with high affinity for dopamine D2 and D3 receptors only, has been reported to have therapeutic effects on both negative and positive schizophrenic symptoms, although at distinct dose ranges (50-300 mg/day vs. 400-1,200 mg/day). The purpose of this study was to investigate the binding of amisulpride to extrastriatal (i.e., thalamus and temporal cortex) and striatal D2 dopamine receptors with respect to plasma amisulpride determinations. Ten patients with schizophrenia treated with amisulpride over a wide range of doses (25-1,200 mg/day) were studied. Positron emission tomography images were acquired by using 76Br-FLB-457, a highly specific antagonist of the D2 and D3 dopamine receptors. Binding indexes (BI) in the regions studied were estimated with reference to values from six healthy subjects. A curvilinear relationship was demonstrated between plasma concentration of amisulpride and the BI in extrastriatal regions. The BI also varied as a function of plasma concentration in striatum. Furthermore, the data provide evidence for different binding profiles: low plasma concentrations (28-92 ng/mL) induced marked extrastriatal binding and low striatal binding, whereas higher plasma concentrations (>153 ng/mL) induced marked binding both in extrastriatal and striatal regions. Dose dependent differential binding profiles of amisulpride to D2 receptors in extrastriatal and striatal regions were demonstrated, and two therapeutic ranges of plasma concentrations for negative and positive schizophrenic symptoms, respectively, are suggested. PMID- 11270920 TI - Comparing guanfacine and dextroamphetamine for the treatment of adult attention deficit/hyperactivity disorder. AB - The objective of this study was to compare the efficacy of the alpha-2a agonist guanfacine with that of dextroamphetamine for the treatment of adult attention deficit/hyperactivity disorder (ADHD). Seventeen adult outpatients who met DSM-IV criteria for ADHD participated in a double-blind, placebo-controlled, crossover study comparing drug effects on ADHD symptoms. Measures of change included the DSM-IV ADHD Behavior Checklist for Adults and the Copeland Symptom Checklist for Adult Attention Deficit Disorders. Cognitive measures of attention included the Stroop and Controlled Oral Word Association Test using the letters "C," "F," and "L" (COWAT, CFL version). For each trial, the drug was administered daily and titered up to optimal doses of maximum efficacy but with a minimum of side effects, and then data were collected. Both drugs significantly reduced ADHD symptoms on the DSM-IV Adult Behavior Checklist for Adults over placebo (p < 0.05). The Stroop Color subscale showed significant improvement for both drugs (p < 0.05), but the Color-Word measures showed significant improvement for guanfacine only (p < 0.01). The average dose of guanfacine was 1.10 (SD = 0.60), and the most common side effect of guanfacine was fatigue. No subjects discontinued drug trials. This preliminary study indicates that guanfacine may be a well-tolerated treatment option for adult ADHD. PMID- 11270922 TI - Awareness of tardive dyskinesia in Asian patients with schizophrenia. AB - While ethnocultural differences in risk of tardive dyskinesia (TD) have been suggested, no previous studies have examined whether this factor also plays a role in lack of awareness of TD. This study examined this question in an Asian population with schizophrenia. Six hundred seven patients in a state mental hospital in Singapore were assessed using the Abnormal Involuntary Movement Scale (AIMS) and the Simpson-Angus Rating Scale. Of the 607 patients, 242 (39.9%) met criteria for TD, and 163 (67.4%) patients were not aware of the presence of TD. No significant differences in terms of age, gender, and duration of illness were found between those aware of their TD and those not aware. Daily neuroleptic doses and scores for the AIMS and Simpson-Angus Rating Scale were significantly different, although after logistic regression, only the Simpson-Angus Rating Scale scores remained significant. The finding that a large proportion of our patients lacked awareness of their TD is consistent with other reports in the West and provides evidence that this feature is characteristic of the illness rather than of a specific ethnocultural group. We found an association between lack of awareness and greater severity of extrapyramidal symptoms (EPS), suggesting that there may be a subtype of TD in which lack of awareness and greater vulnerability of developing EPS are features. PMID- 11270921 TI - Cytochrome P450 2D6 genotype and methadone steady-state concentrations. AB - A genetic polymorphism of cytochrome P450 2D6 has been described with the existence of poor (zero functional genes), extensive (one or two functional genes), and ultrarapid metabolizers (three or more functional genes). The authors measured the steady-state trough (R)- (i.e., the active enantiomer), (S)-, and (R,S)-methadone plasma levels in opiate-dependent patients receiving methadone maintenance treatment (MMT) and genotyped them for cytochrome P4502D6. The patients' medical records were reviewed to assess the outcome of the MMT with regard to the absence of illicit opiate consumption and to the absence of withdrawal complaints in ultrarapid and poor metabolizers. Of 256 patients included, 18 were found to be poor metabolizers, 228 to be extensive metabolizers, and 10 to be ultrarapid metabolizers. Significant differences were found between genotypes for (R)- (p = 0.024), (S)- (p = 0.033), and (R,S) methadone (p = 0.026) concentrations to dose-to-weight ratios. For (R)-methadone, a significant difference was found between ultrarapid metabolizers and poor metabolizers (p = 0.009), with the median value in the former group being only 54% of the median value in the latter group. These results confirm the involvement of cytochrome P450 2D6 in methadone metabolism. Although the difference was nonsignificant (p = 0.103), 13 (72%) of the 18 poor metabolizers and only 4 (40%) of the 10 ultrarapid metabolizers were considered successful in their treatment. More studies are needed to examine the influence of the ultrarapid metabolizer status on the outcome of the MMT. PMID- 11270923 TI - Antidepressive treatment with amitriptyline and paroxetine: comparable effects on heart rate variability. PMID- 11270924 TI - Venlafaxine-associated mania. PMID- 11270925 TI - Effects of selective serotonin reuptake inhibitors on sexual function. PMID- 11270926 TI - Lithium augmentation in venlafaxine-refractory depression. PMID- 11270928 TI - Olanzapine and Huntington's disease. PMID- 11270927 TI - Topiramate in the treatment of acute mania. PMID- 11270929 TI - Reply to comments on "Olanzapine treatment of children, adolescents, and adults with pervasive developmental disorders: an open-label pilot study". PMID- 11270931 TI - Piracetam in the treatment of tardive dyskinesia and akathisia: a case report. PMID- 11270930 TI - Olanzapine-associated priapism. PMID- 11270932 TI - Treatment of borderline personality disorder with mood instability with divalproex sodium: series of ten cases. PMID- 11270933 TI - Patient and rater education of expectations in clinical trials (PREECT). PMID- 11270934 TI - Interpretation of antibacterial susceptibility reports: in vitro versus clinical break-points. AB - Currently, antibacterial activity is measured primarily via in vitro laboratory tests. Clinicians rely heavily upon the reported susceptibility gained via in vitro laboratory tests when choosing an antibacterial agent. An evolving concept is to utilise pharmacodynamic and pharmacokinetic drug properties in addition to in vitro susceptibility reports to assess the potential effectiveness of an antibacterial agent against a specific pathogen. This article presents examples of the utility of these concepts in terms of optimal clinical use of common antibacterials as well as more informed interpretation of the in vitro literature. PMID- 11270935 TI - Role of global surveillance in combating bacterial resistance. AB - In recent years, one of the more alarming aspects of clinical microbiology has been the dramatic increase in the incidence of antibacterial resistance among pathogens causing nosocomial as well as community-acquired infections. Numerous antibacterial agents have lost their in vitro activity as a result of selective pressure exerted by antibacterial usage. There is a general consensus on the fact that emergence and spread of resistance may be delayed by improving hygiene measures, reducing inappropriate use of antibacterials, and adopting successful empirical therapy based on sound epidemiological data. As a consequence, worldwide international studies of antibacterial resistance surveillance have been established. Surveys such as the Alexander Project and the SENTRY Programme supply high quality data to participating countries, stimulate collaboration and provide the educational information required for clinical decision-making that may result in improved cure rates. PMID- 11270936 TI - Comparative review of topical ophthalmic antibacterial preparations. AB - The choice of an antibacterial is based on considerations of pharmacodynamic, pharmacokinetic and bacteriological characteristics, risk of selecting resistant mutants, and cost. In this article we review 16 commercially available ophthalmic antibacterial preparations. Fusidic acid and bacitracin are selective for gram positive bacteria whereas polymyxin B targets gram-negative species. Aminoglycosides and quinolones are broad spectrum antibacterials. The widespread use of an antibacterial increases risks of selecting resistance to it. Acquired resistance is well documented for fusidic acid and rifamycin, and newly described for quinolones. The bioavailability of an antibacterial agent depends on the target bacterial species, the site of infection and the integrity of the haemato aqueous barrier. Some agents (fusidic acid, quinolones) penetrate the cornea, passing into the anterior chamber of normal eyes at therapeutic concentrations, whereas others (polymixin B, bacitracin) have no penetrating powers and remain at the surface of the eye. Toxicity is mostly manifested by allergic reactions to excipients or active ingredients in topical antibacterial preparations. A few cases of haematological toxicity have brought suspicion on topical chloramphenicol, but the link has yet to be proven. Erythromycin and polymyxin B are probably okay to use as topical applications in pregnant women and nursing mothers. Costs of treatment must be evaluated as a whole (regimen, drug associations). Prices for a bottle of eyedrops may vary 3-fold. The cheapest drugs include chloramphenicol, polymyxin B and gentamicin, the most expensive being fusidic acid and the quinolones. PMID- 11270937 TI - Prophylaxis against herpesvirus infections in transplant recipients. AB - Herpesvirus infections are important after stem cell and organ transplant. During the last decades several antiviral agents have been introduced with efficacy against herpesviruses. These agents are the nucleoside analogues aciclovir, valaciclovir, famciclovir, and ganciclovir; the nucleotide analogue cidofovir; and the pyrophosphate analogue foscarnet. Several studies have been performed with antiviral agents with the aim to reduce morbidity and mortality associated with herpesvirus infections in transplant recipients. Aciclovir and valaciclovir have been examined in randomised, controlled trials in both solid organ and stem cell transplant patients, and were shown to be very effective for the prevention of herpes simplex virus (HSV) and varicella-zoster virus infections. In addition, these drugs were shown to reduce cytomegalovirus (CMV) infection and improve survival in allogenic stem cell transplant patients and to reduce CMV infection, CMV disease (aciclovir and valaciclovir), and acute rejection (valaciclovir) in renal transplant patients. Ganciclovir is very effective for the prevention of CMV infection and disease in both stem cell and solid organ transplant recipients. It can also be used in preemptive strategies in which the aim is to prevent CMV disease in patients who have ongoing CMV infection documented by antigenaemia or detection of CMV DNA. The latter strategy has the advantage of reducing the exposure to the drug and thereby the risk for toxicity. Foscarnet has also been shown to be effective as preemptive therapy for CMV in allogenic stem cell transplant patients and as therapy for aciclovir-resistant HSV infections. Finally cidofovir is an interesting agent with broad spectrum antiherpesvirus efficacy. However, because of the drug's toxicity profile, further studies are needed. PMID- 11270938 TI - Safety profiles for the HMG-CoA reductase inhibitors: treatment and trust. AB - Hypercholesterolaemia is a chronic condition that often requires life-long treatment, making the safety of lipid-lowering drugs a critical issue. 3-hydroxy 3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors ('statins') are commonly used as the pharmacotherapeutic treatment of choice for patients with hypercholesterolaemia. These agents have consistently demonstrated a positive safety and tolerability profile, and are recommended by the US National Cholesterol Education Program guidelines and by the European Joint Task Force for Prevention of Coronary Heart Disease to be used after, or in addition to, a first line approach with diet. Several large-scale clinical trials have shown HMG-CoA reductase inhibitors to be efficacious and well tolerated, and to be associated with a low rate of treatment withdrawal due to adverse events. These studies included mortality and morbidity end-points, and comprised both primary- and secondary-prevention trials. Hepatic, renal and muscular systems are rarely affected during HMG-CoA reductase inhibitor therapy and the few drug interactions that can occur with concomitantly administered drugs are well documented. There is no conclusive evidence linking HMG-CoA reductase inhibitors to the development of cancer in humans. In long term studies with various HMG-CoA reductase inhibitors, there was no increase in cancer rates compared with placebo. Thus, it can be concluded that HMG-CoA reductase inhibitors are well tolerated, effective treatments for hypercholesterolaemia. PMID- 11270941 TI - Omalizumab. AB - * Omalizumab is a recombinant humanised monoclonal antibody which specifically binds to the C epsilon3 domain of immunoglobulin (Ig) E, the site of high affinity IgE receptor binding. * Improvements in asthma symptoms and health related quality-of-life, and a significant reduction in the frequency of asthma exacerbations were seen in allergic asthmatic patients treated with omalizumab. * Omalizumab was also effective in the treatment of children with allergic asthma demonstrating improvements in health-related quality-of-life and significant dosage reductions of inhaled corticosteroids. * Administration of omalizumab to patients with allergic rhinitis resulted in a rapid dose-dependent suppression of serum free IgE levels. * Omalizumab significantly improved health-related quality of-life and nasal symptoms in patients with seasonal allergic rhinitis. Antihistamine requirements were also significantly reduced following treatment. * Adverse events were infrequent in clinical trials of omalizumab, and not significantly different from placebo. The most frequent drug-related event was mild to moderate urticaria. PMID- 11270940 TI - Treatment options for sleep apnoea. AB - Sleep apnoea (SA) is a common sleep disorder affecting 4 to 25% of the adult population. The most common form, obstructive SA, is characterised by recurrent upper airway obstruction during sleep associated with sleep disruption and hypoxaemia. There is increasing evidence that SA leads to impaired vigilance, quality of life, driving accidents and probably represents a vascular disease risk factor. Currently, the most effective treatments are aimed at increasing upper airway space by either air pressure [(continuous positive airway pressure (CPAP)], upper airway surgery or oral appliances. CPAP is the main treatment modality for moderate to severe SA but noncompliance approaches 50% in clinic populations. A number of pharmacological agents have been used in SA but at this stage, none are indicated in moderate to severe SA. PMID- 11270942 TI - Oseltamivir: a review of its use in influenza. AB - Oseltamivir is a prodrug of oseltamivir carboxylate (Ro 64-0802, GS4071), a potent and selective inhibitor of the neuraminidase glycoprotein essential for replication of influenza A and B viruses. Studies in volunteers with experimental human influenza A or B showed that administration of oral oseltamivir 20 to 200 mg twice daily for 5 days reduced both the quantity and duration of viral shedding compared with placebo. Subsequent assessment of the drug at a dosage of 75 mg twice daily for 5 days in otherwise healthy adults with naturally acquired febrile influenza showed that oseltamivir reduced the duration of the disease by up to 1.5 days and the severity of illness by up to 38% compared with placebo when initiated within 36 hours of symptom onset (earlier initiation of therapy was associated with faster resolution). The incidence of secondary complications and the use of antibacterials were also reduced significantly in oseltamivir recipients. A liquid formulation of oseltamivir (2 mg/kg twice daily for 5 days) has been shown to be effective in the treatment of children with influenza, and data presented in abstracts suggest that the drug may also be of use in high-risk populations such as the elderly or those with chronic cardiac or respiratory disease. In addition to treatment efficacy, the drug has demonstrated efficacy when used for seasonal or household prophylaxis. Oral oseltamivir (75 mg once or twice daily for 6 weeks) during a period of local influenza activity significantly prevented the development of naturally acquired influenza by >70% compared with placebo in unvaccinated otherwise healthy adults. The drug also demonstrated efficacy when used adjunctively in previously vaccinated high-risk elderly patients (92% protective efficacy). Short term administration of oseltamivir (75 mg once daily for 7 days) may significantly reduce the risk of illness in household contacts of infected persons when administered within 48 hours of symptom onset in the infected person. Oseltamivir 75 mg twice daily for 5 days was well tolerated in clinical trials in healthy adults and high-risk patients, with nausea and vomiting being the most commonly reported events. Gastrointestinal events were mild and transient and both nausea and vomiting were less likely when oseltamivir was taken with food. CONCLUSIONS: Oseltamivir is a well tolerated orally active neuraminidase inhibitor which significantly reduces the duration of symptomatic illness and hastens the return to normal levels of activity when initiated promptly in patients with naturally acquired influenza. It therefore represents a useful therapeutic alternative to zanamivir (especially in patients who prefer oral administration or who have an underlying respiratory disorder) and the M2 inhibitors amantadine and rimantadine (because of its broader spectrum of anti-influenza activity and lower likelihood of resistance) in patients with influenza. In addition, although annual vaccination remains the best means of influenza prevention, there may be a place for oseltamivir in providing household prophylaxis or adjunctive prophylaxis in high-risk vaccinated patients during an outbreak of the disease or for use in patients in whom vaccination is unsuitable or ineffective. PMID- 11270944 TI - More on bioplastique. PMID- 11270945 TI - Proceedings of the 2nd Annual Conference on Vaccine Research. 27-30 March 1999. Bethesda, Maryland, USA. PMID- 11270939 TI - New insights into the second generation antihistamines. AB - Second generation antihistamines are recognised as being highly effective treatments for allergy-based disease and are among the most frequently prescribed and safest drugs in the world. However, consideration of the therapeutic index or the benefit/risk ratio of the H1 receptor antagonists is of paramount importance when prescribing this class of compounds as they are used to treat non-life threatening conditions. There are many second generation antihistamines available and at first examination these appear to be comparable in terms of safety and efficacy. However, the newer antihistamines in fact represent a heterogeneous group of compounds, having markedly differing chemical structures, adverse effects, half-life, tissue distribution and metabolism, spectrum of antihistaminic properties, and varying degrees of anti-inflammatory effects. With regard to the latter, there is growing awareness that some of these compounds might represent useful adjunct medications in asthma therapy. In terms of safety issues, the current second generation grouping includes compounds with proven cardiotoxic effects and others with the potential for adverse drug interactions. Moreover, some of the second generation H1 antagonists have given cause for concern regarding their potential to cause a degree of somnolence in some individuals. It can be argued, therefore, that the present second generation grouping is too large and indistinct since this was based primarily on the concept of separating the first generation sedating compounds from nonsedating H1 antagonists. Although it is too early to talk about a third generation grouping of antihistamines, future membership of such a classification could be based on a low volume of distribution coupled with a lack of sedating effects, drug interactions and cardiotoxicity. PMID- 11270943 TI - Zafirlukast: an update of its pharmacology and therapeutic efficacy in asthma. AB - Zafirlukast is a selective and competitive orally administered inhibitor of the cysteinyl leukotrienes LTC4, LTD4 and LTE4. The drug is indicated for the prophylaxis and treatment of chronic asthma, and has been developed in response to mounting evidence indicating the importance of the cysteinyl leukotrienes in the pathogenesis of this disorder. The efficacy of zafirlukast 20 mg twice daily has been shown in double-blind placebo-controlled studies of up to 13 weeks' duration in patients aged > or = 12 years. Zafirlukast was consistently superior to placebo in improving objective measures of lung function and subjective measures such as symptom scores and use of as-required bronchodilator therapy. This dosage is also as effective when added to low-dosage inhaled corticosteroid therapy as doubling of corticosteroid dosages. Recent studies indicate superior efficacy over zafirlukast of twice-daily inhaled fluticasone propionate 88 microg or salmeterol 42 microg, although zafirlukast was nevertheless associated with clinical improvement. Data also show zafirlukast 40 mg to be of similar efficacy to pranlukast 225 mg (both twice daily). Overall, preliminary pharmacoeconomic data suggest that healthcare costs are reduced by zafirlukast therapy, although superior cost effectiveness has been reported with inhaled fluticasone propionate. and further studies are needed. Data are available to show improvements in patient-rated quality of life, and preference for and high rates of compliance with zafirlukast. In clinical trials, zafirlukast has shown an adverse event profile similar to that of placebo. Isolated reports of hepatic dysfunction in a small number of individuals receiving the drug have been received, and recommendations for monitoring of patients are in place. Although no causal relationship has been established between zafirlukast and Churg-Strauss Syndrome, patients undergoing corticosteroid dosage reductions require careful surveillance. CONCLUSIONS: zafirlukast is an effective and well tolerated agent for preventive monotherapy in mild to moderate persistent asthma. Emerging data indicate benefit of the drug when added to low-dosage inhaled corticosteroids and show that it may be a viable alternative to inhaled adjunctive treatments and increased corticosteroid dosages in some patients. Although inhaled fluticasone propionate and salmeterol have been associated with greater clinical improvement than zafirlukast in clinical studies, compliance considerations and the confirmed clinical efficacy relative to placebo of the drug denote zafirlukast as an effective alternative in treatment programmes based on individualised therapy. As experience with zafirlukast accumulates, it is expected that the drug will be positioned more definitively in national and international treatment guidelines. PMID- 11270946 TI - An ethical paradox: the effect of unethical conduct on medical students' values. AB - OBJECTIVE: To report the ethical development of medical students across four years of education at one medical school. DESIGN AND SETTING: A questionnaire was distributed to all four classes at the Wake Forest University School of Medicine during the Spring of 1996. PARTICIPANTS: Three hundred and three students provided demographic information as well as information concerning their ethical development both as current medical students and future interns. MAIN MEASUREMENTS: Results were analyzed using cross-tabulations, correlations, and analysis of variance. RESULTS: Results suggested that the observation of and participation in unethical conduct may have disparaging effects on medical students' codes of ethics with 35% of the total sample (24% of first years rising to 55% of fourth years) stating that derogatory comments made by residents/attendings, either in the patient's presence or absence, were "sometimes" or "often" appropriate. However, approximately 70% of the sample contended that their personal code of ethics had not changed since beginning medical school and would not change as a resident. CONCLUSIONS: Results may represent an internal struggle that detracts from the medical school experience, both as a person and as a doctor. Our goal as educators is to alter the educational environment so that acceptance of such behaviour is not considered part of becoming a physician. PMID- 11270947 TI - [Central hemodynamics and portal-hepatic blood flow in patients with acute and chronic virus hepatitis]. AB - AIM: To characterize central hemodynamics and portal-hepatic blood flow in patients with acute and chronic viral hepatitis. MATERIAL AND METHODS: Ultrasound investigation, tetrapolar rheography, hepatic scintigraphy were made in 149 patients with acute and chronic viral hepatitis B, C and B + C. RESULTS: The above patients had disturbances in central hemodynamics manifesting with developing myocardiodystrophy and hyperkinetic hemodynamics syndrome; decreased intensity and increased amplitude of respiratory fluctuations of volumic hepatic blood flow; impairment of peripheral blood flow. CONCLUSION: The above changes were most prominent in patients with chronic hepatitis B + C. PMID- 11270948 TI - [Changes in the levels of acute phase proteins in viral hepatitis]. AB - AIM: To investigate changes in the levels of acute phase proteins (APP) in the serum of patients with acute and chronic viral hepatitis of various etiology. MATERIAL AND METHODS: APP (prealbumin, albumin, transferrin, haptoglobin) were measured by automatic kinetic analyzer JCS II TM (Beckman) in 92 patients with acute and 61 patients with chronic viral hepatitis. Synthesis of albumin, prealbumin was depressed, of haptoglobin increased in patients with acute viral hepatitis irrespective of etiology at the height of intoxication. In chronic viral diseases APP content depended on the disease etiology. In chronic hepatitis C, B + C and D albumin, haptoglobin and transferrin were low in clinical exacerbation while in patients with chronic hepatitis B a fall in albumin, prealbumin and transferrin is accompanied with high haptoglobin. CONCLUSION: A complicated multidirectional changes in APP in patients with acute or chronic hepatitis serve, on the one hand, an integral diagnostic indication of hepatic function in conditions of systemic endotoxemia. On the other hand, endotoxemia is caused by lipopolysaccharides of gram-negative bacteria entering blood flow in great amounts in morphofunctional hepatic disorders caused by viral hepatitis of different etiology. PMID- 11270949 TI - [Clinico-diagnostic evaluation of changes in ATPase activity, lipid peroxidation and stability of red cell membranes in patients suffering from hemorrhagic fever with renal syndrome]. AB - AIM: To examine morphofunctional and metabolic features of erythrocytes affecting blood rheology in patients with hemorrhagic fever with renal syndrome (HFRS) for assessment of the disease severity. MATERIAL AND METHODS: 130 HFRS patients were examined using clinical, laboratory, serological tests and fluorescent antibody test. Activity of transport ATPase and content of lipid peroxidation (LPO) products in erythrocyte membranes were measured. These membranes stability was assessed by osmotic and acid resistance in different disease periods. RESULTS: Inhibition of Na+, K+, Ca+ active ATPase of erythrocyte membrane occurred in all the examinees, LPO products rose. The membrane stability was more disturbed in moderate and severe HFRS, especially in polyuretic period. CONCLUSION: Depression of ATPase activity, growth of LPO content in erythrocytes, their relationships can be used as indicators of red cell metabolic disorders, abnormal blood rheology, and eventually, in the disease prognosis. Early membrane defects detected by osmotic and acid resistance can improve the disease diagnosis and provide data on the condition's severity. PMID- 11270950 TI - [Results of spontaneous NBT-test in influenza patients]. AB - AIM: Assessment of leukocyte activity with spontaneous NBT-test in influenza patients regarding the disease stage, severity, complication and concomitant diseases. MATERIAL AND METHODS: 107 influenza patients aged 16-84 years were studied. 70 patients had no complications, 11 patients had early influenzal pneumonia, 26 patients had late viral-bacterial infection. Chronic concomitant diseases were diagnosed in 23 cases. Cytochemical examination of leukocyte activity was made in all the patients using spontaneous NBT-test. RESULTS: In mild influenza NBT-test results were within upper limits of normal value. In alleviation of the symptoms NBT-test parameters were low. In early influenzal and viral-bacterial pneumonia leukocyte activity was high and lowered to normal in late convalescence. CONCLUSION: Parameters of spontaneous NBT-test in influenzal patients were elevated depending on influenza stage, severity and complications. This fact is of differential-diagnostic importance. PMID- 11270951 TI - [Yersiniosis as a therapeutic problem]. PMID- 11270952 TI - [Age and symptoms of Lyme disease]. AB - AIM: Comparison of clinical symptoms in Lyme disease (LD) in various age groups. MATERIAL AND METHODS: 150 patients with verified LD were divided into 4 age groups: under 15 years (group 1), 16-40 years (group 2), 41-60 years (group 3), over 60 years (group 4). Antibodies to Borrelia burgdorteri were detected with indirect immunofluorescence and Western blot. RESULTS: LD clinical symptoms differed in the age groups. Patients of group 1 had more prevalent infectious syndrome with fever but they had no radiculoneuritis and polyneuritis. Patients of group 2 more frequently suffered of carditis and secondary erythema. Groups 3 and 4 were characterized by infectious syndrome, secondary erythema and aseptic meningitis, joint lesions being more frequent in group 3, nervous system lesions- in group 4. CONCLUSION: Age peculiarities of LD symptoms are very important. In particular, joint syndrome is responsible for lingering course of LD. PMID- 11270953 TI - [Pneumocystic pneumonia with lethal outcome in male with Wegener's granulomatosis]. PMID- 11270954 TI - [Activity of chymotrypsin-like proteinases in patients with ischemic heart disease, arterial hypertension and nonspecific aortic arteritis]. AB - AIM: To analyze activity of chymotrypsin-like plasma proteinases (CTP) in patients with various cardiovascular diseases. MATERIAL AND METHODS: CTP activity was studied in 82 patients with various cardiovascular diseases: 13 coronary heart disease patients with normal arterial pressure, 49 patients with essential hypertension (EH) and secondary arterial hypertension, 20 patients with nonspecific aortic arteritis. 28 donors served control. RESULTS: CTP activity in plasma of patients with EH rose 4 times compared to donors. If EH patients had concurrent diseases (CHD, chronic pyelonephritis, atherosclerosis of extracranial arteries), CTP activity may increase by 30-300%. In patients with nonspecific aortic arteritis CTP activity in blood plasma is 17.5 times higher than in donors. CONCLUSION: High CTP activity in cardiovascular patients may be explained by chymase and cathepsin G release into blood flow indicating activation of alternative to ACE pathway of angiotensin II production or the presence of the inflammatory process. PMID- 11270955 TI - [Various regimes of cardiostimulation in determination of anti-recurrence therapy of atrial fibrillation and flutter paroxysms in patients with ischemic heart disease]. AB - AIM: Choice of optimal cardiostimulation regimens using transesophageal pacing for design of antirecurrence antiarrhythmic therapy (AAT) in IHD patients. MATERIAL AND METHODS: 198 patients with IHD complicated by paroxysms of atrial fibrillation (AF) and atrial flutter (AFl) received AAT chosen at concurrent, frequent, volley UHF and slowly increasing UHF stimulation using transesophageal pacing. RESULTS: Slowly increasing UHF stimulation proved most effective both in detection and reproduction of induced paroxysms of AF and AFl. The duration of positive clinical effect of antirecurrence AAT of AF and AFl paroxysms in IHD patients determined at using this regimen of cardiostimulation averaged 3.1 +/- 0.3 years. Left atrial dilatation is an unfavorable prognostic criterion in respect to efficacy of the recurrence AAT. CONCLUSION: Slowly increasing UHF stimulation is most effective in determination of antirecurrence AAT of AF and AFl paroxysms in IHD patients. PMID- 11270956 TI - [Psychological aspects of systemic approach to intra-nosological diagnosis in arterial hypertension]. AB - AIM: To evaluate significance of the program of psychological examination of patients with essential hypertension (EH) for clinicians and characterize the patients psychologically. MATERIAL AND METHODS: 46 EH patients were examined for time of trouble, stress liability, psychological reaction to the disease, factors of vitality. 24-h blood pressure (BP) monitoring was made in 40 patients. RESULTS: It is shown that in EH more frequent are hypochondriac and anxiodepressive psychological types. "White coat" hypertensive reactions (primarily, a rise of systolic blood pressure by 12.5%, on the average) were observed in any psychological types but more in the hypochondriac type. Time of trouble and stress liability were high and correlated. Cluster analysis has identified patients with the highest mean BP. Their psychological analysis is presented. CONCLUSION: The proposed program of evaluation of psychological features of somatic patients allow to define main psychological factors influencing BP. This program is available for clinicians who are not professional psychologists. PMID- 11270957 TI - [Redergin treatment of hypertensive menopausal women]. AB - AIM: To assess a hypotensive effect of redergin (dihydroergotoxin)--agonist of dopaminergic receptors--in monotherapy (4.5-6 mg/day) and in combination with enalapril and amlodipin (10 mg/day). MATERIAL AND METHODS: Redergin in monotherapy or combined therapy was given to 106 hypertensive women in pre- or postmenopause and 24 hypertensive women of reproductive age. Antihypertensive effect was assessed by changes in arterial pressure, frequency and severity of hypertensive crises, diuresis, clinical symptoms of menopausal syndrome. RESULTS: A significant fall in arterial pressure, intensive diuresis, less frequent or absent hypertensive crises, relief of menopausal symptoms were observed on day 10 14 of redergin monotherapy of menopausal patients with mild hypertension and in combined treatment of menopausal women with moderate and severe hypertension. CONCLUSION: Antihypertensive and diuretic effect of redergin confirm a pathogenetic role of deficient dopaminergic activity in development of menopausal hypertension. PMID- 11270958 TI - [Platelet aggregation in patients with various forms of left ventricular hypertrophy and its changes at long-term follow-up and treatment]. AB - AIM: To study platelet aggregation (PA) in patients with left ventricular hypertrophy (LVH) in essential hypertension (EH) and hypertrophic cardiomyopathy (HTCM), and to assess aspirin treatment effects. MATERIAL AND METHODS: A general clinical examination, echocardiography, 24-h monitoring of ECG and arterial pressure, bicycle exercise, test for platelet aggregation, routine blood biochemical tests were performed in 30 males with EH and LVH, 30 males with HTCM and 10 healthy controls. RESULTS: It was found that the patients had high platelet sensitivity to aggregation inductors as well as high spontaneous aggregation. LVH severity correlated positively with activation of spontaneous and induced platelet aggregation. This dependence is unrelated to arterial pressure or the disease duration. Patients with episodes of silent myocardial ischemia had a more pronounced rise in spontaneous and induced platelet aggregation than those without silent ischemia. A long-term administration of aspirin significantly lowered spontaneous and induced platelet aggregation, depression of ST segment. The above changes occurred both in HTCM and in hypertensive patients with LVH. CONCLUSION: Platelet aggregation correlates with severity of LVH and episodes of silent myocardial ischemia both in patients with hypertension, LVH and HTCM. PMID- 11270959 TI - [Follow-up patients with sick sinus syndrome after pacemaker implantation]. AB - AIM: To study a clinical course and dynamics of arrhythmogenesis in patients with sick sinus syndrome (SSS) in varying regimes of permanent pacing. MATERIAL AND METHODS: Of 223 SSS patients, the SSS course and outcomes under pacing were studied in 148 bradyarrhythmia and 75 bradytachyarrhythmia patients. ECG variants and pacing regimes were regarded. RESULTS: Within 4-13-year follow-up, constant form of supraventricular tachyarrhythmia was diagnosed in atrial stimulation in 3.4% of cases, in ventricular stimulation in 32%. Thromboembolic complications and atrioventricular blocks of the II-III degree in atrial stimulation were registered, respectively, in 5.6 and 4.2% cases. Thromboembolism in ventricular stimulation occurred in 11.1%. Reoperation (conversion from atrial to ventricular stimulation, ablation of atrioventricular conjunction) was made for change of pacing method and regime in 11.2% because of changed arrhythmia type. Total mortality for the follow-up period was 14.3%, in bradycardia--9.5%, in bradytachycardia--24%. CONCLUSION: Continuous pacing improves quality of life in SSS patients. PMID- 11270960 TI - [HGV and TTV - new hepatitis viruses]. AB - AIM: To examine clinical, immunological and morphological features of HGV- and TTV-infections in patients with chronic hepatic diseases (CHD) and assess efficiency of treatment of HGV-seropositive patients. MATERIAL AND METHODS: 202 patients with CHD were examined for markers of HBV-, HCV-, HGV- and TTV infections. Some patients were subjected to puncture biopsy of the liver. Efficiency of interferon-alpha treatment of HGV and HBV/HCV coinfection was studied. RESULTS: HGV RNA and TTV DNA were detected in 19.8 and 11.8% of cases, respectively. Biochemical indices in patients with HGV and TTV monoinfections significantly differed from those in the control group while morphological changes in most of them corresponded to those with hepatitis. INF-alpha was given to 7 patients with HGV + HBV/HCV infections. A response was achieved in 3 months in 2 of them. CONCLUSION: The role of HGV and TTV in hepatic diseases pathology is still unclear. Further studies on detection and examination of patients infected with G and TT viruses are necessary. When choosing therapy, the presence of HGV RNA and TTV DNA in blood serum, virus genome in hepatocytes and histological changes in hepatic tissue should be considered. PMID- 11270961 TI - Update on the prevention, diagnosis, and treatment of Lyme disease. AB - With our better understanding of Lyme disease, we now know it is not the "great imitator" of disease it once was thought to be. Limited, identifiable syndromes can be related to Lyme disease. Most of the disease's manifestations resolve without treatment. Treatment with standard antibiotics is very effective at preventing the development of long-term sequelae. The Lyme disease vaccine is safe and effective at preventing transmission of Lyme disease. Future improvements in the care of patients with Lyme disease should focus on identifying the etiology and most effective therapies for patients with posttreatment chronic Lyme disease syndrome, determining the safety and efficacy of vaccination in children, and developing second generation vaccines with improved efficacy and dosing schedules, possibly through the addition of antigens expressed in the human host. PMID- 11270962 TI - Index of suspicion. Case #3. Diagnosis: X-linked agammaglobulinemia (XLA). PMID- 11270963 TI - [Re:"Heterotopic pregnancy: a report of 5 cases and review of the literature"]. PMID- 11270964 TI - Effect of tetrandrine on morphine dependence in isolated guinea pig ileum. AB - AIM: To evaluate the effects of tetrandrine (Tet) and nimodipine (Nim) on the morphine (Mor) withdrawal response in the isolated guinea pig ileum. METHODS: The withdrawal contracture was elicited by addition of naloxone (Nal) (1 mumol.L-1) to the isolated naive ileum incubated with Mor (3 mumol.L-1) at 37.5 degrees C for 4 h or to the ileum obtained from Mor-dependent guinea pig. RESULTS: When Nim (0.01, 0.05, and 0.1 mumol.L-1) or Tet (1, 10, and 50 mumol.L-1) was added 1 min before Nal in the naive ilea bathed in Krebs solution containing Mor, or when the ilea from Mor-dependent guinea pigs were incubated with Nim (0.01, 0.05, and 0.1 mumol.L-1) or Tet (1, 10, and 50 mumol.L-1) for 15 min, or when Nim (5 and 10 mg.kg-1, i.p.) or Tet (15 and 30 mg.kg-1, i.p.) was administered in vivo to Mor dependent guinea pigs, the Nal-precipitated withdrawal contracture was significantly decreased in a dose-dependent manner. CONCLUSION: Tet and Nim, Ca2+ channel blockers, could inhibit the Nal-precipitated Mor withdrawal response in the isolated guinea pig ileum. PMID- 11270965 TI - Properties of transient outward potassium current and inward rectifier potassium current in immature human atrial myocytes. AB - AIM: To study the properties of transient outward K+ current (Ito) and inward rectifier K+ current (IKl) in immature human heart. METHODS: Ito and IKl were recorded using whole-cell patch-clamp technique in atrial myocytes isolated from 12 immature (aged from 6 months to 5 a) human hearts. RESULTS: Ito was voltage dependent, activated and inactivated rapidly. The IC50 (95% confidence limits) of 4-AP on Ito was 0.64 (0.48-0.87) mmol.L-1. 4-AP 1 mmol.L-1 shifted V1/2 of activation from (6.6 +/- 2.0) mV to (19.8 +/- 3.0) mV (n = 4-10, P < 0.01). 4-AP 0.3 mmol.L-1 changed V1/2 of inactivation from (-49 +/- 4) mV to (-61.4 +/- 2.1) mV (n = 3, P < 0.01), but there were no obvious influence on voltage-dependent activation of Ito (P > 0.05). At the same concentration, the recovery time constant (tau value) was prolonged from (108 +/- 16) ms to (220 +/- 67) ms (n = 3 12, P < 0.01). IKl was also voltage-dependent. Its reverse potential was -40 mV. CONCLUSION: Both Ito and IKl are important K+ channel currents in immature human atrial myocytes. 4-AP can affect the inactivation and recovery of Ito at low concentration (0.3 mmol.L-1) and affect its activation at high concentration (1 mmol.L-1). PMID- 11270966 TI - Effects of berbamine on intracellular calcium concentration in cultured HeLa cells. AB - AIM: To study the involvement of Ca2+ signaling and the effects of berbamine (Ber) on intracellular calcium concentration ([Ca2+]i) elevated in cultured HeLa cells. METHODS: [Ca2+]i was measured by confocal microscopy in single HeLa cell loaded with Fluo 3-AM. The change of [Ca2+]i was represented by fluorescent intensity (FI). RESULTS: (1) In the presence of extracellular Ca2+ 1.3 mmol.L-1, the resting level of FI was 186 +/- 44, n = 49 cells from all control experiments, and KCl, NE, caffeine, and calcimycin (Cal) all induced [Ca2+]i elevations in cultured HeLa cells. (2) The resting level of FI was not affected by pretreatment with Ber. The FI increased by KCl 60 mmol.L-1, NE 100 micromol.L 1, and Cal 30 micromol.L-1 were attenuated (P < 0.05 or P < 0.01), the slope and the time to peak of FI increase were decreased and prolonged. (3) In the absence of extracellular Ca2+, caffeine 80 mmol.L-1-induced [Ca2+]i mobilization was not inhibited by Ber 100 micromol.L-1 pretreatment. (4) These effects of Ber were similar to those of verapamil (Ver) 10 mumol.L-1. CONCLUSION: Although it was derived from cervical cancer, the HeLa cells which were belong to the nonexcitable cell possessed the similar biological properties with excitable cells, and Ca2+ also played a crucial role in signal transduction processes. PMID- 11270967 TI - Effects of changrolin on potassium currents in guinea pig and rabbit single heart cells. AB - AIM: To elucidate whether or not changrolin (CRL) modifies the potassium currents (ITO, IK, and IKl) in myocardial cells. METHODS: A tight seal whole-cell patch clamp technique was used to record ITO, IK, and IKl in single cells isolated from guinea pig and rabbit hearts. RESULTS: At a clinically relevant concentration, CRL 50 mumol.L-1 inhibited the transient outward current (ITO) by 17.7% +/- 2.4% (n = 8) in rabbit atrial cells. The voltage-dependence of steady-state inactivation of ITO was not affected by CRL. This concentration of CRL did not influence the time-independent inward rectifier or the delayed rectifier K+ currents (IKl and IK, respectively) in rabbit and guinea pig ventricular cells. CONCLUSION: CRL inhibited ITO, but not IK nor IKl. PMID- 11270968 TI - Effects of ascorbic acid on human hepatoma cell proliferation and redifferentiation. AB - AIM: To examine the effects of ascorbic acid (AA) on hepatoma. METHODS: Choosing an all-trans tretinoin (Tre) as a positive control, cell growth, and cell redifferentiation tests by cell surface charges, biochemical changes, and cell growth in soft agar were measured. RESULTS: After being treated with AA 6 mmol.L 1, the growth curve and mitotic index of human hepatoma cells decreased remarkably, the cellular growth inhibitory rate amounted to 58.9%. The indices related with cell malignancy alleviated, such as cell surface charge obviously decreased, the electrophoresis rate dropped from 1.64 microns.s-1.V-1.cm-1 to 0.93, the average value of alpha-fetoprotein (alpha-FP) content decreased from 302 micrograms.g-1(protein) to 90, and gamma-glytamyl-transpeptidase (gamma-GT) activity from 0.81 U.g-1(protein) to 0.16. The index related with cell differentiation increased, such as the average level of tyrosine-alpha ketoglutarate transminase activity increased from 10.3 micromol.g-1(protein) to 41.2, and the colonogenic potential decreased 94.4%. CONCLUSION: AA can inhibit human hepatoma cells proliferation, induce redifferentiation, and reverse its malignant phenotypic characteristics. PMID- 11270969 TI - Tyrosine kinase participates in alpha 1A-adrenoceptor-mediated increase of intracellular calcium in human embryo kidney 293 cells. AB - AIM: To determine the role of protein-tyrosine kinase (PTK) in alpha 1A adrenoceptor-mediated increase of [Ca2+]i (intracellular calcium) in human embryo kidney (HEK) 293 cells expressed alpha 1A-adrenoceptor. METHODS: Effects of two PTK inhibitors: genistein and tyrphostin, were investigated on the increase of [Ca2+]i by using Fura-2, The activity of PTK was measured and the accumulation of [3H] InsPs were observed. RESULTS: Norepinephrine stimulated a rapid increase in [Ca2+]i to (371 +/- 31) nmol.L-1 in HEK 293 cells. Norepinephrine-induced increase of [Ca2+]i was inhibited by the tyrosine kinase inhibitors quercetin and tyrphostin by 23.8% and 21.4%, respectively, but the accumulation of [3H]InsPs induced by norepinephrine was not. The activity of the plasma-associated tyrosine kinase was increased to (1.73 +/- 0.72)-fold over the control by norepinephrine 10 mumol.L-1. The norepinephrine-activated PTK was inhibited by calphostin C and depletion of intra- and extra-cellular Ca2+. CONCLUSION: The PTK participates in mobilization of Ca2+ mediated by alpha 1A-adrenoceptors in HEK 293 cell lines. PMID- 11270971 TI - Pharmacokinetics of intragastric ipriflavone solid dispersion in rats. AB - AIM: To evaluate pharmacokinetic behavior of ipriflavone solid dispersion in rats. METHODS: The plasma concentrations of ipriflavone in rats were determined by HPLC with UV detector. RESULTS: Plasma concentration-time curves after ig ipriflavone solid dispersion 250 mg.kg-1 in rats were fitted with one-compartment model. Pharmacokinetic parameters were as follows: Ke = 0.21 h-1, T1/2Ke = 5.19 h, Ka = 1.71 h-1, T1/2Ka = 0.41 h, Tmax = 0.67 h, Cmax = 429 micrograms.L-1, AUC = 3916 micrograms.h.L-1; The relative bioavailability of ipriflavone solid dispersion was 323%. CONCLUSION: Ipriflavone in solid dispersion was absorbed more effectively than that in physical mixture in rats. PMID- 11270970 TI - Antimycoplasmal activities of (S)-(-)-9-fluoro-2,3-dihydro-3-methyl-10 -[4-(2 pyridyl)-1-piperazinyl]-7-oxo-7H-pyrido[1,2,3-de][1,4]benzoxazine -6-carboxylic acid (YH-6) in comparison with other antibiotics in vitro. AB - AIM: To determine the susceptibilities of Mycoplasma and Ureaplasma to (S)-(-)-9 fluoro-2,3-dihydro-3-methyl-10 -[4-(2-pyridyl)-1-piperazinyl]-7-oxo-7H pyrido[1,2,3-de][1,4]benzoxazine -6-carboxylic acid (YH-6) and to compare it with those referential quinolones, macrolides, and tetracyclines. METHODS: The minimum inhibitory concentration (MIC) were determined by microdilution method in vitro. RESULTS: The MIC of YH-6 for Ureaplasma urealyticum (Uu: 250 micrograms.L-1), Mycoplasma hominis (Mh: 500 micrograms.L-1), M orale (Mo: 125 micrograms.L-1) and M salivarium (Ms: 125 micrograms.L-1) were closely similar to those of macrolides (erythromycin and leucomycin) and were 2-8 folds greater than those of ofloxacin (Ofl). Uu and Mh easily induced resistance to erythromycin and tetracycline. They did not easily form resistance to quinolone (YH-6, Ofl), josamycin and tylosin. Tetracycline-resistance (Tcr) or erythromycin-resistance (EMr) strains of Uu (or Mh) had cross-resistance to erythromycin or tetracycline. However, they had no cross-resistance to quinolone, josamycin and tylosin. CONCLUSION: YH-6 was a highly active quinolone against Mycoplasma, but could hardly induce resistance to Uu. EMr- or Tcr- strains of Uu (or Mh) had no cross-resistance to YH-6. PMID- 11270972 TI - Effects of agmatine on afterdepolarizations induced by isoproterenol in guinea pig papillary muscles. AB - AIM: To study the effects of agmatine (Agm) on early afterdepolarizations (EAD) and delayed afterdepolarizations (DAD) induced by isoproterenol (Iso) in guinea pig papillary muscles. METHODS: EAD and DAD were recorded using intracellular glass microelectrode technique. RESULTS: (1) EAD and DAD induced by Iso 20 nmol.L 1 were markedly inhibited by pretreatment with Agm 1.0-2.0 mmol.L-1 in a concentration-dependent manner. (2) NG-nitro-L-arginine methyl ester (L-NAME, 0.5 mmol.L-1), a NOS inhibitor, did not affect the inhibitory effects of Agm (1.0 mmol.L-1) on EAD and DAD induced by Iso. (3) The inhibitory effects of Agm (1.0 mmol.L-1) on EAD and DAD induced by Iso (20 nmol.L-1) were eliminated by pretreatment with idazoxan (Ida, 0.1 mmol.L-1), an alpha-2 adrenergic receptor (alpha 2-AR) and imidazoline receptor (IR) antagonist. CONCLUSION: The inhibitory effects of Agm on EAD and DAD induced by Iso in papillary muscles is related to the reduction in calcium influx and mediated by alpha 2-AR and/or IR. PMID- 11270973 TI - Quantitative design of drug compatibility by weighted modification method. AB - AIM: To set up a new method for designing and quantitatively analyzing drug compatibility. METHODS: Drugs for compatibility were divided into 6 dose levels which were evenly distributed to 6 compound groups according to a fixed design. A new mathematical model was set up to fit the dose-effect data of 6 groups. The coefficients, obtained from the model, reflected the dose-effect relationship and the important degree of every drug in combination. According to the coefficients, the drugs in compatibility could be distinguished into principal drug, synergist, inferior, antagonist, and assistant. Because compatibility in the maximal effect group was nearly (or was) an optimal one in 6 groups, the doses in the group were taken as a base for further modification which considered interaction among drugs. The results of the modification were demonstrated by further experiment. This method was applied to design and to quantitatively analyze the compatibility of allantoin, metronidazole, and dexamethasone sodium phosphate by 2 effect indices in mice. RESULTS: This new method was able to effectively determine important degree of drugs in combination, and to optimize their doses for designing compatibility. CONCLUSION: This weighted modification method is a highly efficient, accurate, and practical means for designing and quantitatively analyzing drug compatibility. PMID- 11270974 TI - Specificity of inducible nitric-oxide synthase inhibitors: prospects for their clinical therapy. AB - Nitric oxide (NO) is a ubiquitous, naturally occurring molecule found in a variety of cell types and organ systems. It is a double-edged sword, beneficial as a messenger or modulator and for immunology self-defense, but potentially toxic. The formation and signal function of nitric oxide are mainly modulated by nitric-oxide synthase (NOS). Up to the present, a number of diseases, including circulatory shock, atherosclerosis, cardiac allograft rejection, chronic inflammation, cardiac infarction, cancer and so on, have been demonstrated that their pathogenesis may be involved in the sustained production of large quantities of nitric oxide. Animal studies and human studies have shown that specific inhibitors of inducible nitric-oxide synthase may be useful in the therapy of a variety of diseases associated with induction of nitric-oxide synthase. In this review, we compare and contrast these inhibitors along with examples of their use in the studies of medicine. PMID- 11270975 TI - Effects of RP58866 on transmembrane K+ currents in mammalian ventricular myocytes. AB - AIM: To determine effects of RP58866 on inward rectifier K+ current (IKl), transient outward K+ current (Ito) and delayed outward rectifier K+ current (IK) in isolated cardiac myocytes. METHODS: In isolated ventricular myocytes of guinea pig and dog, the effect of RP58866 on IKl, Ito, and IK were observed by the whole cell voltage-clamp technique. RESULTS: RP58866 decreased IKl in a concentration dependent manner, with an IC50 of (3.4 +/- 0.8) micromol.L-1 (n = 6) at -100 mV in guinea pig ventricular cells. In dog ventricular myocytes, RP58866 inhibited Ito with IC50 of (2.3 +/- 0.5) micromol.L-1 at +40 mV. In guinea pig ventricular cells, RP58866 at 100 micromol.L-1 decreased IK: IKstep by (58 +/- 13)% at +40 mV, and IKtail by (86 +/- 17)%, respectively. RP58866 inhibited IKstep with an IC50 of (7.5 +/- 0.8) micromol.L-1, and IKtail with an IC50 of (3.5 +/- 0.9) micromol.L-1. The envelope of tail analysis suggested that both IKr and IKs were inhibited. CONCLUSION: RP58866 inhibits IKl, Ito, and IK in cardiac myocytes with a similar potency, and is not a specific IKl inhibitor. PMID- 11270976 TI - Chlorpromazine inhibits hepatocyte apoptosis caused by withdrawal of phenobarbital in mice. AB - AIM: To study the inhibitory effect of chlorpromazine (Chl), verapamil, and aspirin on hepatocyte apoptosis induced by the cessation of phenobarbital (Phe) treatment in mice. METHODS: Liver DNA content, ratio of liver weight/body weight, DNA fragmentation, DNA electrophoresis, the end-labeling test (TUNEL), and the morphologic changes of liver cells as indices of liver mass and hepatocyte apoptosis were applied to investigate (1) the kinetic process of hepatocyte proliferation induced by Phe 75 mg.kg-1 i.p. and the regression of hyperplastic liver caused by withdrawal of Phe in mice, (2) the effect of Chl 25 mg.kg-1, verapamil 50 mg.kg-1 or aspirin 60 mg.kg-1 i.p. on mouse hepatocyte apoptosis, and (3) the time course of effects of Chl on the regression of liver size and DNA fragmentation content after withdrawal of Phe. RESULTS: The process of hepatocyte proliferation and regression induced by administration and withdrawal of Phe in mice consisted of 4 phases: proliferation, plateau, rapid regression, and slow regression phases. In the rapid regression phase, the typic changes of hepatocyte apoptosis were found, which was prevented in early period by the Ca(2+) calmodulin antagonist Chl, but not by verapamil or aspirin. CONCLUSION: The Ca(2+)-calmodulin played an important role in the hepatocyte apoptosis caused by withdrawal of Phe. PMID- 11270977 TI - Comet electrophoresis of blood nucleated cells in genotoxicity assessment. AB - AIM: Genotoxicity evaluations of several different chemicals including L-4 oxalysine, 10-Hydroxycamptothecin (HCT), 19-norprogesteron (ST1435), dimethyl sulfoxide (Me2SO), bleomycin (BLM), and mitomycin C (MMC). METHODS: Alkaline comet assay in vitro (single cell gel) (SCG). RESULTS: L-4-oxalysine and HCT did not cause directly DNA damage. ST1435, the subdermal implant progestin, had no effect on DNA damage until the dose level up to 4 mmol.L-1. Me2SO did not increase DNA damage at concentration below 2%, but showed a concentration dependent DNA damage at > or = 4%. Bleomycin and mitomycin C demonstrated a strong dose-dependent DNA damage. CONCLUSION: Comet assay as a tool to test the genotoxicity of suspected chemicals, is rapid, simple, sensitive, good reproducible, and inexpensive. PMID- 11270979 TI - Effects of ligustrazine, tanshinone II A, ubiquinone, and idebenone on mouse water maze performance. AB - AIM: To observe the effects of four drugs, ligustrazine (Lig), tanshinone II A (Tan), ubiquinone (Ubi) and idebenone (Ide), on learning and memory of mouse. METHODS: Mouse water maze was used to evaluate nootropic effect. RESULTS: In comparison with the defective model (only scopolamine 3 mg.kg-1, Tan 20 mg.kg-1, ig) shortened the escape latency dramatically from (36 +/- 19) s to (11 +/- 5) s (P < 0.01) and reduced errors from 7 +/- 5 to 1.5 +/- 1.3 (P < 0.05). Ubi 20 mg.kg-1 ig decreased the escape latency from (37 +/- 18) s to (17 +/- 12) s and errors from 8 +/- 5 to 2.1 +/- 2.7 (P < 0.01). Ide 120 mg.kg-1 (ig) reduced the errors from 8 +/- 6 to 3.4 +/- 2.9 (P < 0.05), but had no remarkable effect on the escape latency. Lig did not exhibit marked effect on the deficit. CONCLUSION: Tan, Ubi, and Ide improved scopolamine-caused spatial performance defects in mouse. PMID- 11270978 TI - Action sites of rotation and unit firing induced by l-stepholidine and DA agonists in basal ganglia of 6-OHDA-lesioned rats. AB - AIM: To elucidate the action sites of l-stepholidine (SPD) in the basal ganglia. METHODS: Counting the rotations after intra-nucleus microinjection and recording the neuron firing by microiontophoresis of SPD and DA agonists in the basal ganglia of 6-hydroxydopamine (6-OHDA)-lesioned rats. RESULTS: The DA immunoreactive substance was markedly reduced in the 6-OHDA-lesioned rats. The intra-neostriatum microinjection of apomorphine (Apo, D1/D2), SK&F 38393 (D1), and SPD elicited remarkable rotation, and the characteristics of SK&F 38393 produced rotation were of long latency and long duration. The intra-substantia nigra pars reticulata (SNR) injection of Apo, SK&F 38393, and SPD induced the rotation response, while the selective D2 agonist quinpirole hydrochloride (Ly171555) did not because of scarce D2 receptors in the SNR. The intraglobus pallidus (GP) injection of DA agonists and SPD failed to evoke rotation, but the GP nucleus still had the contribution to rotation elicited by i.p. injection of DA agonists and SPD in the 6-OHDA-lesioned rats with successive kainic acid (KA) lesion. Besides, the successive lesion of entopeduncular nucleus (EP) on rotation was less important than that of GP nucleus. The microiontophoresis of Apo and SPD into the SNR could evoke the neuron firing, but failed to activate the GP neurons, which were activated by sodium glutamate (Glu) and inhibited by gamma aminobutyric acid (GABA). CONCLUSION: The action sites of SPD-induced rotation and neuron firing via the D1 receptors are in the neostriatum and SNR instead of GP. The direct neurocircuit through SNR is the most important for rotation of 6 OHDA-lesioned rats. PMID- 11270980 TI - KN-62 provides neuroprotection against glutamate-induced excitotoxicity in neurons. AB - AIM: To study the effects of KN-62, an inhibitor of Ca(2+)-calmodulin dependent protein kinase II (CCDPK II), on the damage of cortical neurons and mechanisms of the loss of CCDPK II activity induced by sodium glutamate (Glu). METHODS: CCDPK II activity was measured by 32P incorporation and backphosphorylations of endogenous proteins were studied by autoradiography. RESULTS: 1) KN-62 provided partial protection against excitotoxical damage only before Glu (100 mumol.L-1, 10 min) treatment. 2) KN-62 markedly suppressed the loss of CCDPK II activity induced by Glu from 48.0% to 90.6%. 3) Backphosphorylation of endogenous proteins (especially the 50 kDa protein) reduced to 78.2% of control after treatment with Glu, and the reduction was protected with KN-62 added before Glu. CONCLUSION: KN 62 provided the protection against excitotoxicity and the loss of CCDPK II activity as well as backphosphorylation of endogenous proteins induced by Glu. The neuroprotection provided by KN-62 was due to the inhibition of autophosphorylation of CCDPK II. PMID- 11270981 TI - Protection of dopaminergic antagonists against anoxia-induced inhibition of Ca(2+)-calmodulin dependent protein kinase II activity in rat brain. AB - AIM: To study the effect of dopamine receptor antagonists on anoxia-induced inhibition of Ca(2+)-calmodulin dependent protein kinase II (CCDPK II) activity in rat hippocampus and striatum. METHODS: Using the rat hippocampal and striatal slices under 95% N2 + 5% CO2, the activity of CCDPK II was examined by 32P incorporation. RESULTS: Under anoxia for 30 min, the CCDPK II activity decreased to 29.2% and 27.0% of the control in rat hippocampal and striatal slices, respectively. Preincubation with Sch-23390 (a specific D1-like dopamine receptor antagonist), or domperidone (a specific D2-like dopamine receptor antagonist), resulted in a concentration-dependent attenuation of the anoxia-induced inhibition of CCDPK II activity which was preserved up to about 60%. CONCLUSION: Dopamine receptor stimulation is involved in anoxia-induced inhibition of CCDPK II activity in rat hippocampus and striatum. PMID- 11270982 TI - Nitric oxide-dependent mechanism of anti-ischemic myocardial protection induced by monophosphoryl lipid A. AB - Monophosphoryl Lipid A (MLA) is a detoxified derivative of endotoxin and was first derived and purified from bacterial lipopolysaccharide in 1980s. This pharmacological agent has been studied as a vaccine adjunct, anti-septic, or anti tumor agent by means of its immunomodulatory properties. In addition, MLA is one of the most well documented protective drugs against cardiac ischemia/reperfusion injury in various animal species. Mechanisms involved with the MLA-induced cardioprotection are still not fully understood. A key role for ATP-sensitive potassium channels and inducible nitric oxide synthase (iNOS) has been proposed. This article provides a brief overview on the updated understanding of MLA induced cardioprotection and focuses on the new evidence and insights that were brought into the field by a number of new publications during 1998-1999. Our recent study in a globally ischemic mouse heart model is particularly highlighted. An obligatory role for nitric oxide (NO) in mediating the delayed cardioprotective effect induced by MLA via induction of iNOS was double-confirmed by using S-methylisothiourea (SMT)--a specific inhibitors of iNOS as well as the iNOS gene knockout mice. A direct association of the MLA-induced infarct size reduction with increased NO production was also demonstrated in this study. Future studies should target on identifying the key type(s) of cytokine and the receptors as well as free radical-activated transcription factors that may be responsible for induction of iNOS and the subsequent anti-ischemic cardioprotection with MLA. Information gathered in the studies on MLA may eventually enhance our understanding in the mechanisms of delayed phase of myocardial preconditioning and its clinical applications. PMID- 11270983 TI - Endothelin-1 releases endothelium-derived endoperoxides and thromboxane A2 in porcine coronary arteries with regenerated endothelium. AB - AIM: To determine the role of endothelium-derived contracting factor (EDCF) in the response to endothelin-1 in arteries with regenerated endothelium. METHODS: Rings of porcine coronary arteries, with and without endothelium of previously deendothelialized left anterior descending coronary arteries and native left circumflex coronary arteries, were suspended in conventional organ chambers for the measurement of isometric force. RESULTS: In quiescent rings of the previously deendothelialized left anterior descending coronary artery treated with the NO synthase inhibitor nitro-L-arginine, endothelin-1 caused contractions which were larger in rings with than that in those without endothelium. Under the same experimental conditions, in the left circumflex coronary artery, the contractions to endothelin-1 were augmented markedly by the removal of the endothelium. In rings with endothelium of the previously deendothelialized left anterior descending coronary artery, indometacin (inhibitor of cyclooxygenase) and ridogrel (thromboxane A2 receptor antagonist and inhibitor of thromboxane synthase) inhibited contractions to endothelin-1. Dazoxiben (inhibitor of thromboxane synthase) inhibited, to the same extent as indometacin and ridogel, the response to higher concentrations of endothelin-1. The endothelium-dependent component of the response to lower concentrations of endothelin-1 was inhibited by indometacin and ridogrel, but not by dazoxiben. In rings without endothelium of both previously deendothelialized left anterior descending and native left circumflex coronary arteries, indometacin and ridogrel did not affect the contractions to endothelin-1. CONCLUSION: These findings suggest that in regenerated endothelium, high concentrations of endothelin-1 stimulate the release of thromboxane A2. Endoperoxides generated by activation of endothelial cyclooxygenase may be the endothelium-derived contracting factor(s) released in regenerated endothelium by lower concentrations of the peptide. PMID- 11270984 TI - Effect of oral administration of vitamin C on human aqueous humor ascorbate concentration. AB - AIM: To study oral administration of vitamin C on human aqueous humour ascorbate concentration. METHODS: High performance liquid chromatography (HPLC) coupled with electrochemical detector (ECD) was used. The effect of oral administration of various doses of ascorbic acid, 0 (control), 1.0, 1.5, 2.0, 3.0, and 5.0 g, on its concentration in aqueous humour, obtained from volunteer cataract patients was studied. RESULTS: The concentration of ascorbic acid in aqueous humour of control group (without administration of vitamin-C tablet or drug containing ascorbic acid was (254 +/- 119) mg.L-1. This study revealed that the administration of 2.0 g of ascorbic acid saturate the aqueous humour and further increase in the dose (3.0 g and 5.0 g) did not increase its concentration in aqueous humour, although its concentration was increased in plasma. CONCLUSION: Oral administration of 2.0 g of Vc is sufficient to saturate the aqueous humour where it may be helpful in controlling the intra-ocular pressure. PMID- 11270985 TI - Comparison of 12-chloroscoulerine enantiomers on animal behavior to dopamine receptors. AB - AIM: To compare the pharmacological characteristics of 12-chloroscoulerine (CSL) enantiomers to dopamine (DA) receptors. METHODS: Radioligand receptor binding assay with calf striatum and behavioral tests of mice or rats were used. RESULTS: In the competitive binding assay, the affinities (Ki) of l-CSL to D1 and D2 receptors were 5.7 nmol.L-1, while those of d-CSL for D1 and D2 receptors were 135 and 9150 nmol.L-1, respectively. The Ki of dl-CSL to D1 and D2 receptors were 8.9 and 9.6 nmol.L-1, respectively, which were slightly weaker than that of l CSL. In the behavioral experiments, CSL enantiomers 5-60 mg.kg-1 antagonized the stereotypy induced by apomorphine in rats, and 5-150 mg.kg-1 produced catalepsy. The enantiomers 10-60 mg.kg-1 reduced the mice jumping behavior induced by amphetamine + levodopa. l-CSL 10-80 mg.kg-1 antagonized the spontaneous locomotor activity of normal or amphetamine-treated mice. CONCLUSION: CSL enantiomers are antagonists to DA receptors: l-CSL > dl-CSL >> d-CSL. PMID- 11270986 TI - Desipramine and fluoxetine antagonized 5,7-dihydroxytryptamine-induced lesion on rat hippocampal and cortical neurons. AB - AIM: To assess the protective effect of desipramine (Des) and fluoxetine (Flu) on the neurons against the lesion induced by a selective serotonergic neurotoxin in vitro. METHODS: The 10-day cultured primary neurons of hippocampus and cortex of rat was exposed to 5,7-dihydroxytryptamine (5,7-DHT) to determine the optimal lesion concentration and duration. Before exposing to 5,7-DHT, Des and Flu was added to the medium for 30 min to observe the protective effects. RESULTS: The optimal concentration and duration for 5,7-DHT was 600 micromol.L-1 and 4 h, respectively. Both Des and Flu showed a protective effect in the dose range of 0.8 micromol.L-1 to 10 micromol.L-1 and 0.04 micromol.L-1 to 0.6 micromol.L-1, respectively, when the neurons were injured by 5,7-DHT 600 micromol.L-1 for 4 h. Flu showed a higher efficacy than Des. Both exhibited a more powerful protective effect on the hippocampal neuron than on the cortical neuron. CONCLUSION: The antidepressant effect of Des and Flu was attributed to their protective effect on the injured serotonergic neuron of the hippocampus and the cortex. PMID- 11270987 TI - Inhibitory effects of 8-(N,N-diethylamino)-n-octyl-3,4,5-trimethoxybenzoate (TMB 8) on intracellular Ca2+ elevated by neurotransmitters in brain cells. AB - AIM: To study the effects of TMB-8 on [Ca2+]i elevation induced by neurotransmitters in dissociated brain cells. METHODS: The brain cell suspension was made using a gentle trituration for 1 min with a polished pipette. The changes of [Ca2+]i were detected by the fluorescent indicator, Fura 2-AM. RESULTS: In the presence of extracellular Ca2+ 1.3 mmol.L-1, sodium glutamate (Glu), histamine (His), and serotonin (5-HT) markedly increased the [Ca2+]i which were reduced by TMB-8 30 mumol.L-1. TMB-8 3 mumol.L-1 produced inhibitory effects on the increase of [Ca2+]i by His and 5-HT in a Ca(2+)-free Hanks' solution. The increase of [Ca2+]i by His and 5-HT was reduced to control level by TMB-8 10 mumol.L-1. CONCLUSION: TMB-8 inhibited the [Ca2+]i elevation induced by Glu, 5 HT, and His in brain cells. PMID- 11270988 TI - Electrophysiologic effects of agmatine on pacemaker cells in sinoatrial node of rabbits. AB - AIM: To study the electrophysiologic effects of agmatine (Agm) on pacemaker cells in sinoatrial (SA) node. METHODS: Parameters of action potential (AP) in SA node were recorded using intracellular microelectrode technique. RESULTS: Agm not only slowed down the amplitude of action potential (APA), maximal rate of depolarization (Vmax), velocity of diastolic (phase 4) depolarization (VDD), and rate of pacemaker firing (RPF), but also prolonged 90% duration of action potential (APD90) in a concentration-dependent manner. The effects of Agm (10 mmol.L-1) could be blocked completely by pretreatment with idazoxan (0.15 mmol.L 1), an alpha 2-adrenergic receptor (alpha 2-AR) and imidazoline receptor (IR) antagonist. Pretreatment with NG-nitro-L-arginine methyl ester (L-NAME, 1 mmol.L 1), an NOS inhibitor, did not affect the electrophysiologic effects of Agm on pacemaker cells in SA node. Elevation of Ca2+ concentration (5 mmol.L-1) in perfusate antagonized the effects of Agm (10 mmol.L-1). Lemakalim (Lem, 30 mumol.L-1), an opener of ATP-sensitive potassium channels, partially inhibited the prolonging effect of Agm on repolarization. CONCLUSION: The electrophysiologic effects of Agm on pacemaker cells in SA node were likely attributed to the reduction in calcium influx and potassium efflux and mediated by alpha 2-AR and IR. PMID- 11270989 TI - Inhibitory effect of dopamine on Ca(2+)-calmodulin-dependent protein kinase II activity in rat hippocampal slices. AB - AIM: To study the effect of dopamine (DA) on Ca(2+)-calmodulin dependent protein kinase II (CCDPK II) activity in rat hippocampus. METHODS: Using rat hippocampal slices as an in vitro model, the activity of CCDPK II was examined by the method of 32P-incorporation. RESULTS: Exogenous DA reduced CCDPK II activity in hippocampal slices in a concentration- and time-dependent manner. Removal of extracellular calcium antagonized the DA-induced inhibition of CCDPK II activity, partially or completely. The activity of CCDPK II was markedly decreased by apomorphine (a nonselective DA receptor agonist), SKF38393 (a selective D1-like DA receptor agonist), or quinpirole (a selective D2-like DA receptor agonist). The inhibition of CCDPK II activity induced by exogenous DA was abolished by preincubation with Sch-23390, a selective D1-like DA receptor antagonist, or domperidone, a selective D2-like DA receptor antagonist. CONCLUSION: DA has an inhibitory effect on CCDPK II activity in rat hippocampus, related to stimulation of D1-like and D2-like receptors and calcium influx. PMID- 11270991 TI - Early and delayed protection by capsaicin against reperfusion injury in rat hearts. AB - AIM: To study early or delayed cardioprotection afforded by pretreatment with capsaicin. METHODS: The isolated rat heart was perfused in a Langendorff model. Heart rate, coronary flow, left ventricular pressure, and its first derivative (+/- dp/dtmax) were recorded, and the calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) and the release of creatine kinase (CK) were measured. RESULTS: Capsaicin (50 mg.kg-1, s.c.) improved the recovery of cardiac function and decreased the release of CK. CK was (2.12 +/- 0.40) and (0.26 +/- 0.04) u.min 1.g-1(wet wt) for ischemia-reperfusion (I/R) and capsaicin + I/R, respectively (P < 0.05). Capsaicin treatment caused an increase in the concentration of CGRP-LI in plasma. CGRP-LI was (135 +/- 12) and (304 +/- 45) ng.L-1 for vehicle + I/R and capsaicin + I/R, respectively (P < 0.05). After pretreatment with capsaicin to deplete the sensory nerve transmitter content, the cardioprotection and the increased level of CGRP by capsaicin were abolished. A delayed protection was shown in the hearts obtained from the rats pretreated with capsaicin 24 h or 48 h before the experiments. CONCLUSION: Pretreatment with capsaicin induces the early and delayed cardioprotection, which may be related to stimulation of CGRP release in the rat. PMID- 11270990 TI - Blocking L-calcium current by l-tetrahydropalmatine in single ventricular myocyte of guinea pigs. AB - AIM: To study the effect of l-tetrahydropalmatine (l-THP) on L-type calcium channel. METHODS: Patch clamp technique (whole cell recording) was used to record L-Ca2+ current in single cardiac myocyte. RESULTS: 1) l-THP 1, 10, and 100 micromol.L-1 reduced ICa-max from (999 +/- 93) pA to (700 +/- 111) pA, (582 +/- 66) pA, and (420 +/- 112) pA (n = 6, P < 0.01), respectively. 2) l-THP reduced the voltage at half-maximal inactivation (V1/2) of L-Ca2+ channel to more negative potentials by 9 mV (n = 5, P < 0.05). 3) l-THP caused both tonic and use dependent reduction of Ca2+ current. Tonic block of l-THP on Ca2+ current was 46% +/- 8% (n = 6, P < 0.01). The degree of use dependent blocking was 13.5% +/- 2.4% (n = 6, P < 0.05) at 1 Hz, the degree increased to 44% +/- 5% (n = 6, P < 0.01) at 3 Hz. 4) l-THP delayed half-recovery time of Ca2+ channel recovery from inactivity from (94 +/- 39) ms to (170 +/- 42) ms(n = 6, P < 0.01). CONCLUSION: l THP has a moderate inhibitory effect on L-Ca2+ current. PMID- 11270992 TI - Inhibitory effect of ligustrazine on proliferation of rabbit vascular smooth muscle cells after arterial injury. AB - AIM: To study the inhibitory effect of ligustrazine (Lig) on growth of cultured rabbit aortic vascular smooth muscle cells (VSMC) after balloon injury. METHODS: Twenty New Zealand white rabbits were subjected to arterial injury with a balloon catheter (810 kPa for three consecutive inflations, 1 min each time). The uptake of [3H]thymidine in primary cultural VSMC incubated with rabbit serum, which obtained from the animals treated without or with Lig (40 mg.kg-1.d-1, i.v.) for 21 d (7 d before and 14 d after the injury procedure) was determined. The determination was performed in direct addition of TMP to culture as well. And histological cross-sections of the blood wall were also analyzed. RESULTS: After balloon injury the intimal thickening (77 +/- 23) microns and lumen diameter narrowing (877 +/- 118) microns in dilated sites were increased significantly than the normal adjacent wall [(41 +/- 13) microns, P < 0.01; (1033 +/- 175) microns, P < 0.05, respectively]. Treatment with Lig decreased both intimal thickening (56 +/- 16) microns (P < 0.05) and lumen diameter narrowing (1023 +/- 157) microns (P < 0.05). Lig inhibited [3H]thymidine uptake in VSMC incubated with the serum obtained from these rabbits. Direct addition of Lig inhibited [3H]thymidine uptake in cultured VSMC in a dose-dependent manner (40-4000) micrograms/well. CONCLUSION: Lig shows a pronounced inhibitory effect on VSMC proliferation after balloon injury. PMID- 11270994 TI - Effects of 3-n-butylphthalide on production of vasoactive substances by cerebral and aortic endothelial cells. AB - AIM: The effects of dl-3-n-butylphthalide (dl-NBP), l-3-n-butylphthalide (l-NBP), and d-3-n-butylphthalide (d-NBP) on the production of nitric oxide (NO), epoprostenol (Epo) and endothelin-1 (ET-1) were investigated in cerebrovascular and aortic endothelium in culture. METHODS: Bovine cerebral endothelial cells (BCEC) and bovine aortic endothelial cells (BAEC) were cultured in Medium 199 in vitro. After incubation with dl-, l-, and d-NBP for 24 h, the release of NO, Epo, and ET-1 were analyzed by using spectrometry assay and radioimmunoassay (RIA) respectively. RESULTS: Low concentrations of dl- and l-NBP (0.1-10 mumol.L-1) enhanced nitrite and 6-ketoprostaglandin F1 alpha (6-ketoPGF1 alpha) production in both BAEC and BCEC after a 24-h incubation, and l-NBP has a potent effect on promoting Epo production in BCEC. The production of ET-1 secreted by BCEC and BAEC was increased after TNF alpha stimulation, this enhancement was not blunted by the simultaneous addition of dl-, l-, and d-NBP. CONCLUSION: 1) dl-NBP and l NBP increase NO production in both BCEC and BAEC. 2) l-NBP increases more Epo production in BCEC than that in BAEC, and dl-NBP has selective effect on increasing Epo production in BCEC. PMID- 11270993 TI - Effects of perindopril, propranolol, and dihydrochlorothiazide on cardiovascular remodelling in spontaneously hypertensive rats. AB - AIM: To investigate the effects of perindopril, propranolol, and dihydrochlorothiazide on artery wall thickening, left ventricular hypertrophy, and cardiac fibrosis in spontaneously hypertensive rats (SHR). METHODS: After measurement of systolic blood pressure (SBP), 16-wk-old Male SHR were randomly divided into 3 groups (each n = 10), given perindopril (Per, 5 mg.kg-1.d-1), propranolol (Pro, 40 mg.kg-1.d-1), dihydrochlorothiazide (DCT, 100 mg.kg-1.d-1) respectively by gavage for 12 wk. Sex-, age-, and number-matched untreated SHR and normotensive Wistar Kyoto rats (WKY) served as controls. When the treatment finished, body weights (BW) and SBP were measured before decapitation of the rats. The heart was excised rapidly, the left ventricle was weighed and then subjected to collagen content analysis. Vascular wall and lumen ratio from aorta, renal arteries and branch III vessels of mesenteric arteries were determined morphometrically. RESULTS: Treated rats in 3 groups showed a lower SBP and the ratio of left ventricle weight to body weight (LVW/BW) compared with WKY. Artery wall thickening was similarly inhibited in the treated groups. Per and Pro inhibited cardiac fibrosis, but collagen concentration increased in DCT treated SHR [collagen volume fraction (CVF): 19 +/- 4 vs SHR 14 +/- 4, P < 0.05; perivascular collagen fraction(PVCF): 84 +/- 7 vs SHR 79 +/- 5, P < 0.05]. CONCLUSION: Per and Pro inhibited, but DCT promoted, cardiac fibrosis. PMID- 11270995 TI - Inhibitory effect of MAP kinase antisense oligonucleotide on angiotensin II induced c-myc gene expression and proliferation of rat cardiac fibroblast. AB - AIM: To investigate the inhibitory effect of down-regulating mitogen activated protein kinase (MAPK) on c-myc gene expression and further on cardiac fibroblast proliferation. METHODS: Cultured neonatal rat cardiac fibroblasts was pretreated with a phosphorothioate-protected 17-mer antisense MAPK oligodeoxynucleotide (ODN) directed against the initiation of translation sites of the p42 and p44 MAPK isoforms by liposomal transfection. A 17-mer sense and mismatch sequence MAPK ODN were used as controls. After liposomal transfecting, cells were exposed to angiotensin II (Ang II) 10 nmol.L-1 for 5 min and then harvested in lysis buffer. MAPK activity was measured by Western blot and P-81 phosphocellulose filter paper method by using [gamma-32P]ATP and myelin basic protein as substrates. c-myc mRNA expression stimulated by Ang II for 30 min was measured by Northern blot. DNA synthesis and collagen protein synthesis induced by Ang II for 24 h were measured by [3H]thymidine incorporation and [3H]Proline incorporation, respectively. RESULTS: Antisense ODN 0.2 mumol.L-1 reduced Ang II-induced MAPK activities by 72%, MAPK protein expression by 80%, and suppressed c-myc mRNA expression by 97%, respectively. [3H]thymidine incorporation and [3H]proline incorporation in Ang II-induced cardiac fibroblast were inhibited by 59% and 58%, respectively. CONCLUSION: A 17-mer MAPK antisense oligonucleotide directed againsts the initiation of translation sites of MAPK could specifically inhibit Ang II-stimulated cultured neonatal rat cardiac fibroblast proliferation through down-regulating MAPK activity and further depleting c-myc mRNA expression. PMID- 11270996 TI - Ginkgolide A, B, and huperzine A inhibit nitric oxide production from rat C6 and human BT325 glioma cells. AB - AIM: To study the effects of ginkgolide A, B (Gin A, Gin B) and huperzine A (Hup A) on nitric oxide (NO) production from cultured astrocytes. METHODS: Nitrites in supernatants were measured with Griess assay. RESULTS: Hup A 0.001-100 mumol.L-1 time- and concentration-dependently inhibited the NO production from rat C6 astrocytoma cells. The NO production from C6 cells was concentration-dependently inhibited by the treatment with Gin A or Gin B 0.001-10 micromol.L-1 for 24 h. The NO production from human BT325 astrocytoma cells was concentration dependently inhibited by Hup A, Gin A, or Gin B 0.01-10 micromol.L-1 for 24 h. CONCLUSION: Gin A, Gin B, and Hup A inhibited astrocytes producing NO. PMID- 11270997 TI - Fibrinogenolytic properties of natrahagin (a proteinase from cobra venom) and its effect on human platelet aggregation. AB - AIM: To study the fibrinogenolytic properties of natrahagin and its effect on platelet aggregation. METHOD: SDS-PAGE, fibrinogenolytic activity assay, platelet aggregation. RESULTS: Upon incubation of fibrinogen with natrahagin at the ratio of 50:1 (w/w), A alpha-chains of fibrinogen were almost completely hydrolyzed in 5 min; however, at least 6 h was needed for the complete degradation of gamma chains. Fibrinogenolytic activity of natrahagin was 0.349 +/- 0.044 g.min-1.g-1 as determined by its ability to reduce the clottable fibrinogen. On the other hand, natrahagin concentration-dependently inhibited platelet aggregation induced by ristocetin in platelet-rich plasma and thrombin (80 U.L-1) in washed platelets with IC50 (95% confidence limit) of 56 (40-79) and 3.3 (1.4-8.0) mg.L-1. No inhibitory effect was found on collagen- and ADP-induced platelet aggregation even when the dose of natrahagin reached 200 mg.L-1. CONCLUSION: Natrahagin is an alpha, gamma-fibrinogenase with an inhibitory effect on platelet membrane glycoprotein Ib (GPIb)-dependent platelet aggregation. PMID- 11270998 TI - Platelet activating factor-induced P-selectin expression in platelets and its related signal transduction. AB - AIM: To study the intracellular signal transduction mechanisms of platelet activating factor (PAF)-induced platelet P-selectin expression. METHODS: Human blood platelets were used to test the effect of PAF-induced P-selectin expression using flow cytometry. RESULTS: PAF 20 nmol.L-1 elicited a moderate upregulation of P-selectin expression [(47.5 +/- 1.3)% vs control (3.8 +/- 0.9)%, P < 0.01]. Pretreatment with egtazic acid (EGTA) 2 mmol.L-1 and 5,5'- dimethyl-bis-(o aminophenoxy)-ethane-N,N,N',N'-tetracetic acid (BAPTA) 200 mumol.L-1 to block Ca2+ influx or chelate the intracellular calcium, respectively, reduced P selectin expression in response to PAF [(13.3 +/- 0.9)% and (16.8 +/- 1.9)% vs (47.5 +/- 1.3)% of PAF group, P < 0.01]. Inhibition of Na+/H+ exchange with amiloride (Ami) 400 mumol.L-1 resulted in an inhibition of P-selectin expression [(37.5 +/- 2.1)% vs (47.5 +/- 1.3)% of PAF group, P < 0.01]. Genistein (Gen) 300 mumol.L-1 to inhibit protein tyrosine phosphorylation showed similar effect [(29 +/- 4)% vs (47.5 +/- 1.3)% of PAF group, P < 0.01]. CONCLUSION: Multiple signal transduction pathways, including protein tyrosine phosphorylation, Na+/H+ exchange, and Ca2+ mobilization, mediated PAF-induced P-selectin expression. PMID- 11270999 TI - Pharmacokinetics of sustained-release capsule of 5-isosorbide mononitrate in 20 healthy Chinese young men. AB - AIM: To compare the pharmacokinetics of domestic and imported sustained-release capsule of 5-isosorbide mononitrate (5-IM). METHODS: A single and 5-d-repeated oral doses of 5-IM 50 mg were performed on 2 groups of 20 Chinese healthy subjects (10 subjects for each group) in a randomized crossover protocol. The 5 IM in plasma were measured by gas chromatography with electron-captured detector method. Data were analyzed automatically by using a CAPP program on a PC computer. RESULTS: Fitting the 5-IM concentration-time curves to one-compartment model or following trapezoidal rule, the parameters such as Tmax, Cmax, Ke, MRT, and AUC were calculated and there were no significant differences between the two kinds of capsule. The major pharmacokinetic parameters of domestic and imported 5 IM sustained-release capsule with a 5-d multiple dose were respectively: Cmax (677 +/- 103) and (702 +/- 76) micrograms.L-1; Tmax (5.1 +/- 2.0) and (5.6 +/- 1.3) h; MRT (11.5 +/- 0.5) and (11.4 +/- 0.7) h; AUC0-infinity (12,121 +/- 1346) and (12,352 +/- 988) micrograms.h.L-1. The fraction of drug absorbed in vivo was correlated well with the percentage amount of drug released in vitro at corresponding time (P < 0.05), and the fluctuation indices on d 5 in multiple dose study were not significantly different between the two formulations (P > 0.05). The relative bioavailability of the domestic capsule for single and multiple dose were 96% +/- 11% and 98% +/- 10%, respectively. CONCLUSION: Domestic 5-IM sustained-release capsule showed bioequivalence compared with the imported capsule and provided the same nitrate-low interval in the latter part of the 24-h dosing interval. PMID- 11271000 TI - Preparation and dissolution property of ipriflavone solid dispersion. AB - AIM: To prepare and identify ipriflavone (IP) solid dispersion, and determine its dissolution property. METHODS: The solvent method was used for preparation and differential scanning calorimetry (DSC), X-ray diffraction and infrared spectrophotometry for identification of IP solid dispersion. The dissolution of the dispersion was determined with paddle method. RESULTS: The dissolution of IP solid dispersion consisting of IP and povidone-k30 (PVP) (1:8) in artificial gastric juice is 6.15 times as high as that of IP alone. The DSC curves, X-ray diffraction patterns and infrared spectrophotometries of IP have been changed obviously by the dispersion. CONCLUSION: The dissolution of IP is increased by solid dispersion method. PMID- 11271001 TI - [The TT virus: review of the literature]. AB - OBJECTIVE: To review the literature on the TT virus. METHODS: The literature review was based on articles identified through MEDLINE between Jan. 1, 1997, and August 15, 1999. RESULTS: In 1997, a new DNA virus, designated TTV, was isolated and seemed to be associated with non A-G post-transfusion hepatitis. The virus was identified using a polymerase chain reaction (PCR) because serology was not routinely available. At least 16 genotypes were identified. Depending on the PCR technique used, the prevalence of infection ranged from 17% to 71% in a group of sera tested. The prevalence rate ranged from 1.2% to 62% among blood donors, from 0.5% to 83% among hemophiliacs and from 1% to 71% in cases of chronic hepatitis. The current hypothesis is that routes of infection were parenteral and orofecal. The pathogenesis of this virus, if it really exists, is not yet clearly established. It has been postulated that some interaction may exist between the TT virus and the hepatitis C virus. The use of interferon seems to decrease the TT viremia, according to results obtained outside the context of clinical trials. CONCLUSION: The pathogenesis of the TT virus needs to be rapidly established for transmission prevention and therapeutic intervention. PMID- 11271002 TI - Identification and genetic mapping of four novel genes that regulate leaf development in Arabidopsis. AB - Molecular and genetic characterizations of mutants have led to a better understanding of many developmental processes in the model system Arabidopsis thaliana. However, the leaf development that is specific to plants has been little studied. With the aim of contributing to the genetic dissection of leaf development, we have performed a large-scare screening for mutants with abnormal leaves. Among a great number of leaf mutants we have generated by T-DNA and transposon tagging and ethyl-methae sulfonate (EMS) mutagenesis, four independent mutant lines have been identified and studied genetically. Phenotypes of these mutant lines represent the defects of four novel nuclear genes designated LL1 (LOTUS LEAF 1), LL2 (LOTUS LEAF 2), URO (UPRIGHT ROSETTE). and EIL (ENVIRONMENT CONDITION INDUCED LESION). The phenotypic analysis indicates that these genes play important roles during leaf development. For the further genetic analysis of these genes and the map-based cloning of LL1 and LL2, we have mapped these genes to chromosome regions with an efficient and rapid mapping method. PMID- 11271003 TI - [How do we get rid of the tube?]. PMID- 11271004 TI - [Health care reform in Germany 2000]. PMID- 11271005 TI - [The demographic factor in Germany in the 21st century]. PMID- 11271006 TI - [Pediatric diabetes nursing consultant at the German Society for Diabetes]. PMID- 11271007 TI - [Developmental problems in children with chronic diseases on the example of type 1 diabetes mellitus]. PMID- 11271008 TI - Development and validation of clinical indicators for mental health nursing practice. AB - A national study was undertaken in Australia to develop and validate a set of clinical indicators for mental health nursing. Using survey and action research procedures, the indicators were developed in two stages. During stage one, focus group interviews involving 39 nurses were conducted at national conferences in Australia and New Zealand in order to provide a pool of indicator statements. A Delphi survey of an Australian sample of mental health nurses (n = 33) was then conducted to refine the indicators. In stage two, the refined indicators were tested and validated in selected clinical settings. A total of 1751 mental health nurses employed at 14 sites were involved in the second stage of the study. The resulting data were used to establish the set of national indicators that the Australian and New Zealand College of Mental Health Nurses will use in practice accreditation and benchmarking. PMID- 11271009 TI - The psychiatric consultation-liaison nurse: towards articulating a model for practice. AB - Psychiatric consultation-liaison nursing (PCLN) has experienced considerable growth in the past decade in Australia. In spite of this growth, a model for PCLN practice has not been adequately developed and articulated. The aim of this paper is to begin the process of articulating a model for PCLN practice in order to redress the paucity of literature in this area. The paper includes a brief history of PCLN; articulation of the components of the PCLN role, namely consultation, liaison and culture brokering; the impact of PCLN on quality improvement; and the importance of research articulation for the development of the PCLN role. PMID- 11271010 TI - Scoping mental health nursing education. AB - In late 1999 the National Mental Health Working Group of the Australian Health Ministers Advisory Council commissioned the Australian and New Zealand College of Mental Health Nurses to undertake a scoping study of mental health nursing. A final report will be submitted to the National Mental Health Working Group in February 2000. The purpose of this article is to draw attention to some of the systemic problems that confront the education of mental health nurses in Australia. Shortcomings in the preparation of undergraduate students of nursing for commencing practice in mental health nursing are described and comments are given on issues affecting the quality of postgraduate mental health nursing education. PMID- 11271011 TI - Lisa's lessons: a case study of mental health teaching and learning. AB - Practical approaches to the educational preparation of mental health nurses need to be shared in order to contribute to discipline development. This paper presents the results of an ethnographic study using case study and educational criticism to explore mental health classrooms and share practical approaches to teaching. PMID- 11271012 TI - Charting the future today: psychiatric and mental health nurses in cyberspace. AB - The development of the Internet is happening at a staggering pace and promises to have a dramatic impact on human relations. If nursing is to adapt to and benefit from these changes, consideration ought to be given to the experiences and opinions of nurses who have adapted to and use the technology. This paper provides an outline of the findings of an Email survey of psychiatric and mental health nurses who are experienced in using the Internet. Questions focused on what psychiatric and mental health nurses use the Internet for, how their use has changed, work-related benefits, and what impact they see the Internet having in the future. PMID- 11271013 TI - Scoping the Australian mental health nursing workforce. AB - This is the second of four articles on the Scoping Study of the Australian mental health nursing workforce conducted on behalf of the Australian and New Zealand College of Mental Health Nurses (ANZCMHN) for the Australian Health Ministers Advisory Council (AHMAC) National Working Group on Mental Health (NWGMH). Its purpose is to focus on overlooked issues in planning the mental health nursing workforce. Whereas it is acknowledged that there are problems in the supply of mental health nurses, it is argued that equal attention needs to be given to addressing the working conditions and rewards of mental health nurses. PMID- 11271014 TI - The physical and psychosocial consequences of opioid addiction: an overview of changes in opioid treatment. AB - This paper examines a selection of the current literature to gain information concerning opioids, addiction profiles, public opinion, legal issues and withdrawal protocols. Hundreds of Australians have died at a young age due to the complications of their own opioid misuse. This paper outlines what has been achieved in recent times in the management of people withdrawing from opioid misuse, as well as reviewing the new evidence that offers hope for faster opioid withdrawal and rehabilitation. PMID- 11271015 TI - Behavioural psychotherapy training for nurses in Australia: a pilot program. AB - This paper describes a pilot program that examined the feasibility of training for qualified mental health nurses in behavioural psychotherapy in response to the perceived need for improved client access to services. A 6-month course was conducted with four nurses from the in-patient mental health unit at Flinders Medical Centre, South Australia. They received a combination of workshop training and supervised practice by qualified and experienced nurse behavioural psychotherapists and were assessed throughout the period for clinical competency and level of knowledge in the subject. All four nurses completed the training satisfactorily. Each trainee treated four clients who presented with a range of anxiety disorders. The implications for further training of suitably qualified mental health professionals in the area are discussed. PMID- 11271016 TI - Permanent and stable housing for individuals living with a mental illness in the community: a paradigm shift in attitude for mental health nurses. AB - The provision of appropriate housing for individuals with a mental illness has been recognized by a number of researchers as a means to enhance effectiveness of treatment and rehabilitation services, to maintain treatment gains, and to decrease community opposition to deinstitutionalization. Whether community-based services, which are now meant to be the focus of treatment, are successful or not is crucially related to the nature and availability of accommodation. This paper argues a case for change in the current philosophical basis of, and services provided by, mental health professionals and agencies that are charged with the responsibility of meeting the housing needs of consumers of mental health services. This change, it is contended, needs to be to an approach that is more flexible, more supportive of the consumer, and in which the consumers are empowered to make decisions and choices about their housing needs. PMID- 11271017 TI - Architecture signifying social control: the restoration of asylumdom in mental health care? AB - Prior to the era of community care, the asylum has been an architectural manifestation of the power of psychiatry and the State. Asylumdom was a period in which custodial warehouses were used by the State to store sections of the population considered to be 'unreasonable'. There are signs, however, of a re birth of asylumdom in both the UK and Australia. For example, there is a projected growth in the number of 'secure units' in the community, and a growing concern with issues of security and risk management in mental health care. Furthermore, new technologies for containment and surveillance are being installed in acute inpatient psychiatric units. Once again the asylum will symbolize emphatically the authority of the State (and its agencies of social control), and re-emphasize the exclusion of 'unreason' from the 'reasonable' society. PMID- 11271018 TI - Brushing has made a sweeping change: use of the endoluminal FAS brush in haemodialysis central venous catheter management. AB - Common usage of central venous catheter (CVC) access for haemodialysis has presented the haemodialysis nurse with the challenge of maintaining CVCs as a viable form of access. The major complications seen with CVC use are obstruction and infection. A project was undertaken to identify the usefulness of the endoluminal fibrin analysis system (FAS) brush as an intervention in haemodialysis CVC management. The aims of the study were to identify: the reasons for brushing CVCs and the number of occasions brushing is indicated; how successful brushing is in unblocking and improving flow from CVCs, and the length of time the catheter remains patent following successful declotting. Seventeen patients were found suitable for CVC brushing and divided into two groups depending on the indication for brushing. In the group in which the catheter was brushed to restore flow, 73 per cent of brushings were successful, and in 50 per cent of those cases the CVC remained patent for 6 weeks. Sixty per cent of catheter brushings to improve flow were successful, and in 50 per cent of the CVCs flow was sustained over a 6-week period. Overall, the findings support the use of the endoluminal FAS brush for the applications trialled. PMID- 11271019 TI - Plethysmography: the new wave in haemodynamic monitoring--a review of clinical applications. AB - The plethysmograph, a useful, non-invasive circulatory assessment capability featured on most modern pulse oximeters, provides a waveform representation of pulsatile peripheral blood flow, from which can be drawn assessments of both the peripheral and central circulation. Implementation and maintenance of plethysmography monitoring is straightforward and uncomplicated by virtue of its non-invasiveness. Yet despite its capabilities, ease of use and widespread availability it remains an underutilised data source. Diagnostic and monitoring capabilities of the device include heart rate and rhythm monitoring, detection of myocardial and valvular dysfunction, assessment of intra-aortic balloon pump performance when pressure waveforms are unobtainable, detection and measurement of pulsus paradoxus, improved performance of the Allen's test and detection of peripheral vascular diseases, peripheral vasoconstriction and developing shock. This paper describes the range of established applications of plethysmography, reviews pertinent literature and describes the directions in which, in the absence of supportive literature, clinical practice is finding applications. PMID- 11271020 TI - Making research connections to improve clinical practice. PMID- 11271021 TI - Providing the best possible care: an overview of the current understanding of diabetic ketoacidosis. AB - Diabetic ketoacidosis remains a potentially fatal sequela of diabetes mellitus, despite advances in health care. Nurses, especially those in critical care areas, need to have a good understanding of the condition, including the physiological changes involved and treatment considerations. This paper reports on some of the literature relating to diagnosis, physiological changes and treatment that pertains to critical care nurses. Hyperglycaemia, ketosis and metabolic acidosis are indicative of the condition, with treatment directed towards reversal of dehydration, restoration of normoglycaemia and reversal of ketoacidosis, correction of electrolyte imbalances (especially hypokalaemia) and prevention of complications. Most important are constant monitoring and evaluation of the patient's condition and titration of treatment to individual needs. Nurses face the challenge of caring competently for the patient with DKA, which includes understanding the physiological changes as these influence management and treatment. PMID- 11271022 TI - ACCCN Ltd position statement on adult and paediatric resuscitation by nurses. PMID- 11271023 TI - Blood gas analysis may lead to iatrogenic anaemia in intensive care. PMID- 11271024 TI - Chest X-ray quiz. Left-sided chest pain. PMID- 11271025 TI - Endotracheal tube stability in the resuscitation environment. AB - Evidence presented in the anaesthetic and emergency medical services literature warns of the possibility of accidental or insidious displacement of endotracheal tubes in intubated patients. In particular, there is evidence that head movement in intubated patients can lead to displacement of the distal tip of an endotracheal tube in the trachea while its depth markings remain fixed in relation to the patient's lips or teeth. Immobilisation of the heads of all intubated patients should be considered, to prevent the possibility of accidental endotracheal tube displacement. In addition, all intubated patients must be clinically assessed for proper endotracheal tube placement and maintenance of adequate ventilation after every movement during resuscitation and diagnostic measures. PMID- 11271026 TI - Recognising our limits: the murky shores where science meets values and beliefs. PMID- 11271028 TI - Nurse-patient communication in the intensive care unit: a review of the literature. AB - Patient care within an intensive care unit (ICU) can be a difficult and stressful task for even the most experienced and skilled critical care nurse. Good communication between the patient, relatives and nurse is integral to quality care of the patient and should extend to the entire health-care team. This article reviews the literature on nurse-patient communication in the ICU. While numerous research studies have been completed, they are predominantly qualitative and descriptive. Recent studies have investigated the patients' perceptions and recollections of the communication that transpired between them and nurses while they were cared for within an ICU. The literature indicates that nurses communicate extremely poorly with patients, despite a high level of knowledge and skill with respect to communication. Tentative explanations of high stress levels, a preoccupation with physical care and technology, and the attraction to critical care areas of nurses with specific personality types are discussed as possible reasons for this. The need for further research into, and attempts to alleviate, this problem is clearly demonstrated. PMID- 11271027 TI - Measuring the health outcomes of general ICU patients: a systematic review of methods and findings. AB - Studies that have measured patient outcomes following a critical illness and admission to a general intensive care unit (ICU) have used a variety of methods and variables. Traditionally, the measures have been mortality and morbidity, but they now also include assessment of indices such as functional status, health status, quality of life and patient satisfaction. This review paper examines 31 previously published primary studies that measured patient outcomes from adult general ICUs. Inclusion criteria for the review process included papers with explicit measurement of patient outcomes beyond mortality rates: functional status, health status and quality of life. The paper includes summative descriptions of the reviewed papers, discussion of the various methods, results and limitations, and some synthesis of the pooled findings. The review indicated that the health status and activities of the majority of survivors who responded to the follow-up measures were similar to what they were before their critical illness. However, whether these subjects are representative of ICU survivors in general is not conclusive, as lack of methodological control over exclusions and losses to follow-up were evident. In addition, the heterogeneity of the ICU population in terms of age, acuity and diagnoses complicates any potential comparisons. There is an opportunity for further studies in this area by nurse researchers, in either intra- or multidisciplinary teams. Future studies should incorporate rigorous methodologies and a triangulated approach, in order to adequately examine patient outcomes following a critical illness and admission to a general ICU. PMID- 11271029 TI - Chest X-ray quiz. A ventricular aneurysm. PMID- 11271030 TI - Relatives' lived experiences of complementary therapies in a critical care department--a phenomenological study. AB - This phenomenological study examined relatives' lived experiences of complementary therapies in the Department of Critical Care Medicine at Royal Hobart Hospital. Participants in the study, 20 relatives of critically ill patients, were involved in a non-structured, audiotaped interview. Subsequently, transcripts were analysed using a phenomenological transformative process to identify common themes in the text. Study findings suggested four emerging conceptual categories, which with further analysis uncovered the essence of the phenomenon as extending and enriching a caring atmosphere. PMID- 11271032 TI - The practice characteristics of advanced nurse practitioners in acute and critical care settings. PMID- 11271031 TI - Use of continuous positive airway pressure (CPAP) in the critically ill- physiological principles. AB - CPAP therapy helps improve oxygenation in patients who are awake and able to maintain a good respiratory drive. In many cases this means that intubation and ventilation can be avoided. The main goals of CPAP are to minimise alveolar collapse, improve compliance, decrease work of breathing and improve ventilation/perfusion matching. These effects work together to improve arterial oxygenation. A clear, concise and straightforward discussion of CPAP is difficult to find in the existing literature. As CPAP has developed into a common therapy for patients with respiratory failure, it is essential that nurses using this therapy are familiar with the equipment and the physiological effects it produces. Assessment and management of the patient receiving CPAP therapy are also important. This paper will address the physiological principles of CPAP therapy so that nurses working with critically ill patients receiving CPAP therapy understand the system and are accurate and astute in their respiratory assessment, in order to provide optimum care. PMID- 11271033 TI - Tuberculosis: the latest. AB - Tuberculosis (TB) is a complex disease with a long history of human infection. This article provides an overview of TB and discusses ramifications for anesthesia practitioners. Specific individuals and groups are at an increased risk of developing TB. The acid-fast bacilli that causes TB is transmitted via the airborne route. The anesthesia provider must adapt his/her approach to the patient with TB so that protection is afforded that practitioner, colleagues, and the patient. Future patients are also shielded from potential exposure to acid fast bacilli. To minimize the risk of transmission of acid-fast organisms, the anesthesia practitioner must consider TB as a possible diagnosis when performing a preanesthetic evaluation. Standard precautions for bloodborne pathogens must be followed. In addition, appropriate respiratory precautions must be taken. Proper cleansing, decontaminating, sterilizing, or disposal of equipment must occur. Screening of anesthesia providers for the possibility of infection with TB is to be conducted at intervals recommended by the Occupational Safety and Health Administration. Each anesthetist has a responsibility to practice safely. In today's anesthesia practice, safety includes vigilance against TB. PMID- 11271034 TI - Prevention of occupational transmission of bloodborne diseases in clinical nurse anesthesia practice. AB - This article discusses the most commonly recognized bloodborne infectious diseases with which the nurse anesthetist comes into contact during the provision of clinical care: hepatitis B virus, hepatitis C virus, and the human immunodeficiency virus, along with the potential percutaneous and mucocutaneous routes of exposure. The prevention of occupational exposure to these pathogens is discussed by applying standard or universal precaution measures to anesthesia clinical practice. Treatment measures for accidental exposure to bloodborne pathogens is also discussed. PMID- 11271035 TI - Reprocessing anesthesia instruments and devices. AB - Reprocessing anesthesia instruments and devices can often present a challenge for anesthesia providers because anesthesia devices have become more complex, cross contamination with disease-forming pathogens can occur, and the importance of appropriate reprocessing may not be fully understood. Based on accepted practice recommendations, regulations, and research, reprocessing must be performed by skilled individuals who understand asepsis, cleaning, disinfection, and sterilization principles. This article describes the art of reprocessing and includes highlighted information on recommended practices, Spaulding's classifications, personal protective attire, precleaning, leak testing of flexible endoscopes, device disassembly, cleaning supplies and solutions, cleaning methods, rinsing, reassembly of the device, inspection, disinfection, and sterilization. PMID- 11271036 TI - Anesthetic drug interactions. Quarterly update. PMID- 11271037 TI - Patient safety and human error: the big picture. AB - For most of the past century, health care literature including many books written about health care and its quality have documented the problems of errors in health care delivery. That outcomes of care have differed significantly among hospitals has also inferred that perhaps the "best practices" or the appropriate resources may not have been used, although most of these study results have be adjusted for case mix. The Institute of Medicine's recent publication, "To Err is Human," represents their review of studies quantifying medical errors in health care and their recommendations for eliminating such errors to the extent possible. One should note that, while using the term "medical," it does not infer that all errors are made by physicians. It recommends shifting the focus of study from blaming the health providers to studying the "system" in which health care is provided, believing that most of the errors committed are not reckless but rather result from system variables. The Institute of Medicine's recommendations are broad and cover a variety of quality assurance mechanisms. It recommends mandatory reporting of these errors to a central agency via a state mechanism, with better and broader legislation to make peer review, for purposes of studying errors with a view toward making change in the system, privileged information, and not subject to subpoena. The American Medical Association and American Nurses Association, in their testimony before the US Senate Committee on Appropriations, Subcommittee on Labor, Health and Human Services, Education and Related Agencies, on December 13, 1999, support the recommendations in general with a few reservations. PMID- 11271038 TI - Infectious diseases in the operating room. AB - Patients with infectious diseases have special implications for infection control in the operating room. The increased use and abuse of antibiotics has ushered in a category of resistant organisms. These multiresistant organisms are spread by direct or indirect contact, primarily from the hands of caregivers or contact with contaminated environmental surfaces. Another category of infectious diseases is prions (pronounced pree-ons). Unlike other infectious diseases, human prions diseases are not spread through routine exposures such as direct contact, droplet, and airborne routes. The causative agent is highly resistant to traditional disinfecting and sterilization processes. This article provides an overview of the multiresistant infections of methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci, and Staphylococcus aureus with reduced susceptibility to vancomycin along with the human prions diseases Creutzfeldt Jakob disease, German-Straussler-Scheinker syndrome, kuru, and fatal familial insomnia. We provide a template of precautions that can be used in developing operating room and anesthesia infection control protocols for this patient population. PMID- 11271039 TI - Comparison of the length of stage II labor and incidence of forceps and cesarean delivery after epidural infusion of 0.125% bupivacaine with 2 mu/mL fentanyl as compared with 0.0625% bupivacaine with 2 mu/mL fentanyl. AB - A large, retrospective chart review was conducted to analyze the length of stage II labor and instrumental and cesarean-section delivery rates in nulliparous women who received either 0.0625% bupivacaine with 2 mu/mL fentanyl or 0.125% bupivacaine with 2 mu/mL fentanyl. Data collected included length of stage II labor, incidence of operative or instrumental delivery rates, concentration of bupivacaine used, and demographic data. Demographics obtained included maternal age, weight, and height, as well as neonatal gestational age, weight, and Apgar scores. Further investigated were additional analgesic requirements of supplemental boluses of local anesthetic between the groups. No differences in demographics were noted between the groups. Instrumental delivery rates were similar between the groups with an incidence of 17.5% in the 0.125% bupivacaine group versus a 15% incidence in the 0.0625% bupivacaine group. Cesarean delivery rate was 17% in the 0.125% bupivacaine group versus a 21% ratio in the 0.0625% bupivacaine group. Duration of stage II labor was noted to be prolonged in the 0.125% bupivacaine group but was not statistically significant. Based on this data, it can be concluded that the use of 0.125% bupivacaine with 2 mu/mL fentanyl does not cause a statistically significant increase in instrumental or cesarean delivery rates, nor does it have a detrimental effect on length of stage II labor. PMID- 11271040 TI - Intravenous regional anesthesia: new approaches to an old technique. AB - The pace of modern surgical procedures demands a fast and effective regional anesthesia technique. Intravenous regional anesthesia (IVRA) is such a technique. Traditionally, IVRA has been limited by tourniquet pain, inability to provide postoperative analgesia, and lack of a bloodless field for microsurgical repairs. Current research indicates that these limitations can be significantly improved with alteration of the block solution or alteration of the exsanguination technique. Additions to the local anesthetic such as meperidine, ketorolac, and clonidine have been shown to increase tourniquet tolerance and significantly improve postoperative analgesia. Additionally, when a bloodless field is required for microvascular surgery or nerve repairs, a re-exsanguination technique can be used. Advances in IVRA have made this technique an excellent choice for cases involving the hand, forearm, foot, and lower leg cases that least 60 minutes or less. PMID- 11271041 TI - Psoas compartment block. AB - The psoas compartment acts as a conduit for the nerve roots of the lumbar plexus. Originating at approximately the 12th thoracic vertebrae, this potential compartment continues on caudally, bordered posterolaterally by fascia of the quadratus lumborum and iliacus muscles, medially by the fascia of the psoas major muscle, and anteriorly by the transversalis fascia. This natural "gutter" acts as a repository for local anesthetic agents and provides an excellent method of unilateral anterior lower extremity anesthesia. After elicitation of a motor evoked response in the muscles of the anterior thigh, 30 to 40 milliliters of local anesthetic is incrementally injected into the compartment. Spread of the anesthetic to all roots of the plexus occurs in 15 to 20 minutes. Profound sensory and motor blockade can be achieved providing surgical anesthesia as well as long duration postoperative pain relief. PMID- 11271042 TI - Regional anesthesia and issues of coagulation status. AB - Regional anesthesia has generally been accepted as safe and is associated with a relatively low incidence of neurological injury. In 1993, the Food and Drug Administration approved the use of low molecular weight heparin for perioperative thromboprophylaxis. In the past 5 years, there has been an alarming increase in the number of intraspinal hematomas and neurological injury associated with its use in patients undergoing neuraxial anesthesia. This article reviews the coagulation cascade, the various laboratory studies used to assess the coagulation system, and issues related to neuraxial block in the patient with perioperative anticoagulation therapy. The various anticoagulant drugs and their potential to cause intraspinal hemorrhage are addressed with special attention paid to low molecular weight heparin. Finally, recommendations for managing the patient who receives these drugs is presented. PMID- 11271043 TI - Anesthetic drug interactions. Quarterly update. PMID- 11271044 TI - Rural health care and the nurse anesthetist. AB - As they work in all types of practice arrangements and settings with and without anesthesiologists, certified registered nurse anesthetists have been and continue to be the principle anesthesia providers in rural hospitals in the United States. As such, they are responsible for providing anesthesia services to about 1 quarter of the US population that resides in rural and frontier areas of this country. Rural health care is characterized by its necessity to provide a broad array of services with lesser resources than are typically available in metropolitan or urban areas. The rural population is characterized as having a higher proportion of elderly people and children under the age of 18, a higher incidence of chronic diseases, a lower mortality rate (albeit a slightly higher infant mortality rate), and a 40% higher mortality rate resulting from accidents. Rural residents are poorer and less likely to have job-related health insurance benefits or Medicare supplemental insurance. Despite the significant rise in the number of anesthesiologists in the past 10 to 15 years, there is no evidence that they are attracted to practice in these areas. As sole anesthesia providers in many of these rural hospitals, rural CRNAs have both common and unique problems and issues that confront them. However, from available reports, their communities are satisfied with their services, providing evidence of the capability of CRNAs to function satisfactorily without the anesthesiologist. PMID- 11271046 TI - Analogy. Clarification or obfuscation? PMID- 11271045 TI - Risk factors associated with postoperative pulmonary complications following total abdominal hysterectomy. AB - The purpose of this descriptive correlational study was to describe the relationships between risk factors and the development of postoperative pulmonary complications (PPCs) following total abdominal hysterectomy (TAH). As part of a large, prospective study, data were analyzed on a subset of women who had undergone TAH. Data collection included a preoperative interview and chest exam followed by a daily postoperative interview, chest exam, and review of the medical chart. A multicriteria definition of PPC was used for atelectasis/pneumonia. This study describes the incidence of PPCs in a TAH surgical population and provides foundational work to begin identifying important risk factors to guide pulmonary care. PMID- 11271047 TI - Outcome indicators for direct and indirect caregiving. AB - Informal caregiving and outcomes for caregiving are an important part of health care and of particular importance in nursing. The purpose of this research is to report the results of a survey mailed to nursing experts for validation of the outcome labels Caregiver Role Performance: Direct Care and Caregiver Role Performance: Indirect Care and their accompanying indicators. Experts were asked to rate how important the identified indicators were for assessing those two outcomes. In addition, the respondents were asked to what extent nursing interventions influence the achievement of each identified indicator for Caregiver Role Performance: Direct Care and Caregiver Role Performance: Indirect Care. In general, the validity of the concept analysis work by the caregiver focus group was supported. Ten indicators for Caregiver Performance: Direct Care were retained, 1 was dropped that was considered most appropriate for indirect care, and 3 new indicators were added to reflect the nurse experts surveyed. For Caregiver Performance: Indirect Care, all of the indicators were retained. PMID- 11271048 TI - Perimenopause and the quality of life. AB - The purposes of this study are to describe the frequency and distress of symptoms associated with perimenopause, to examine the changes in the quality of life (QOL) related to perimenopause, and to examine the relationships between symptoms associated with perimenopause and the QOL. A cross-sectional, correlational design was employed. Two hundred fourteen perimenopausal women completed the Women's Health Assessment Scale (WHAS) and the Quality of Life Scale. It was found that vasomotor symptoms were not central to the list of symptoms associated with perimenopause. More women reported psychosomatic complaints as opposed to vasomotor complaints. Compared to the premenopausal period, women during perimenopause experienced slightly, yet significantly decreased, levels of QOL. Multiple regression analysis demonstrated that the psychosomatic symptom category was the sole predictor of the QOL during perimenopause. In summary, psychosomatic symptoms occur most frequently and are most distressful for perimenopausal women in this study. It may be important to manage psychosomatic symptoms to improve the QOL for perimenopausal women. PMID- 11271049 TI - Nursing support during labor. AB - This descriptive study replicates and extends the previous work on nursing support during labor. Within 72 hours of giving birth, 88 participants rated 25 selected nursing behaviors on a Likert scale as to their perceived helpfulness, with two open-ended questions for additional comments on helpful nursing behaviors. Nursing behaviors were categorized as emotional, informational, or tangible support. The majority of behaviors considered most helpful were in the emotional support category. Sixteen of the 25 behaviors were considered helpful, with the most helpful behaviors being making the woman feel cared about as an individual, appearing calm and confident, and treating the woman with respect. Findings were consistent with those of a previous study and suggest that, regardless of the pain management used, nurses supporting childbearing women must not only be competent but also use a high degree of interpersonal skills in providing nursing care. PMID- 11271050 TI - Is arm span an accurate measure of height in young and middle-age adults? AB - Height is used to determine many important clinical measurements, but height may be difficult or impossible to measure accurately in some patients. The purpose of this study is to determine the accuracy of arm span as a measure of height in young and middle-age adults. A sample of 83 people between the ages of 20 and 61 years participated in this anthropometric study. Height and arm span were measured with a metal rule. A prediction equation was derived from regression analysis. Arm span is a valid measure of height in young and middle-age adults, and the accuracy is improved when using the prediction equation. PMID- 11271051 TI - Evaluating and managing burn wounds. AB - Central to successful management of those who have suffered a burn of any size or depth is proper and effective evaluation and management of the wound. If a careful initial evaluation is made, basic principles of management are applied, and the response to therapy is regularly monitored, reliably excellent results can be expected for the large majority of patients. PMID- 11271052 TI - The Stigma Scale: measuring body image and the skin. AB - People with chronic wounds or skin conditions often face not only physiological challenges, but also emotional ones as well. Measuring these emotional challenges and more broadly, the emotional challenges of having a skin defect, was the focus of this instrument development study. PMID- 11271053 TI - What's your assessment? Foreign body. PMID- 11271054 TI - Complementary, alternative, integrative: have nurses kept pace with their clients? AB - While medical literature reflects an interest in the use of complementary therapies, there is a paucity of studies in the nursing literature addressing the use of therapies by nurses, either on themselves or on their clients. While the utilization rate of complementary therapies by the general population has been estimated to be as high as 45%, and nurses are interacting with clients who use these therapies on a daily basis, little is known about nurses' attitudes, knowledge, or perceived efficacy of the therapies. PMID- 11271055 TI - Fungal contamination of outpatient examination rooms: is your office safe? AB - Dermatophyte and nondermatophyte fungi are ubiquitious in the envionment. This study demonstates the presence of dermatophyte fungi and saprophytic fungi contaminating step stools and flooring samples in office examination rooms. The use of protective coverings as well as other barriers will help to protect both patients and health care workers from acquiring nosocomial infection. PMID- 11271056 TI - Wound assessment and evaluation. Gangrene. PMID- 11271057 TI - Erythrodermic CTCL: a case study of treatment with extracorporeal photopheresis. AB - Erythrodermic cutaneous t-cell lymphoma (CTCL) is an aggressive variant of CTCL with multiple skin manifestations and symptoms. A brief overview of CTCL and a case study of one patient with erythrodermic CTCL who underwent successful treatment in a research protocol with extracorporeal photopheresis and UVADEX are presented. Nursing care, critical for patients with erythrodermic CTCL, is also described. PMID- 11271058 TI - What's your assessment? Wart. PMID- 11271059 TI - Geriatric dermatology in chronic care and rehabilitation. AB - The skin care of disabled elders living in nursing homes and adult congregate, subacute, and home health settings requires special effort and consideration. Practitioners who regularly assist elders in these situations may be unfamiliar with dermatology problems related to chronic disability. As the U.S. population ages, skin care in the elderly and disabled will continue to be a challenge. However, if the diseases and problems can be addressed in a knowledgeable and problem-oriented manner, treatment can be maximized. PMID- 11271060 TI - Wound assessment and evaluation. PMID- 11271061 TI - Acupuncture: National Institutes of Health Consensus Development Conference statement. PMID- 11271062 TI - Empowering leadership. PMID- 11271063 TI - Who's the customer? PMID- 11271064 TI - Dermatoses in African-Americans. AB - Many cutaneous diseases which afflict Caucasians also occur in Blacks. However, some of these diseases may occur more commonly in African-Americans or present in an "atypical" or different manner. It is important to be aware of these variations in presentation to properly diagnose and manage these diseases. There is also a subset of skin diseases which are unique to Black patients about which the clinician should be knowledgeable. Finally, because so many skin diseases can affect the pigmentary system, causing hypopigmentation or hyperpigmentation, the social and psychological impact of cutaneous disease at times may be more devastating in African-Americans. PMID- 11271073 TI - Index of suspicion. Case 2. Diagnosis: Leprosy. PMID- 11271074 TI - Is there more than one radiation-induced fecal incontinence syndrome? Re: Yeoh Ek et al.: chronic effects of therapeutic irradiation for localized prostatic carcinoma on anorectal function. IJROBP 2000; 47:915-924. PMID- 11271075 TI - Serial changes in tumor oxygenation during the early phase of radiation therapy in cervical cancer-are we quantitating hypoxia change? Re: Lying et al., IJROBP 2000; 46:935-946. PMID- 11271076 TI - VO2/DO2 relationship: how to get rid of methodological pitfalls? PMID- 11271077 TI - Hydrocortisone and the reduction of vasopressors in septic shock: therapy or only chart cosmetics? PMID- 11271078 TI - The state of research on multipurpose severity of illness scoring systems: are we on target? PMID- 11271079 TI - Intravenous amiodarone in intensive care. Time for a reappraisal? AB - Amiodarone is widely used in intensive care units for the treatment of a variety of arrhythmias. It is currently the drug of choice for supraventricular tachyarrhythmias in many units because of its combination of efficacy and safety. This review summarises the current state of knowledge regarding the short-term administration of intravenous amiodarone to control arrhythmias in perioperative, coronary care and intensive care patients. It outlines the electrophysiology, haemodynamics, pharmacokinetics and toxicity of the drug. In particular, it examines the recent concerns regarding acute pulmonary toxicity. PMID- 11271080 TI - Different dosages of dobutamine in septic shock patients: determining oxygen consumption with a metabolic monitor integrated in a ventilator. AB - OBJECTIVE: Oxygen consumption (VO2) obtained from respiratory gases by indirect calorimetry (VO2,IC) with a metabolic monitor integrated in a ventilator were to be compared to VO2 obtained by the Fick principle (VO2,Fick) in septic patients following an increase in oxygen delivery (DO2) induced by positive inotropic support. DESIGN: Prospective clinical study. SETTING: University Hospital, Surgical Intensive Care Unit (ICU). PATIENTS: Thirty patients suffering from sepsis. INTERVENTIONS: DO2 was increased by dobutamine infusion, starting with an initial dosage of 5 microg x kg x min, increased to a maximum of 10 microg x kg x min. MEASUREMENTS AND MAIN RESULTS: Dobutamine infusion induced a dosage-related increase in DO2 (from 577 +/- 192 to 752 +/- 202 ml x min x m2, p < 0.01), which was associated with a statistically significant increase in VO2,IC (from 173 +/- 30 to 188 +/- 28 ml x min x m2, p < 0.01) and in VO2,Fick (from 140 +/- 25 to 156 +/- 24 ml x min x m2, p < 0.01). The comparison between VO2,IC and VO2,Fick revealed differences (bias and precision--33 +/- 32 ml x min x m2). CONCLUSIONS: With a metabolic monitor integrated in a ventilator it was possible to carry out continuous monitoring of calorimetric data under clinical conditions. In contrast to previous studies using indirect calorimetry, this study showed a moderate correlation between VO2 and DO2 in septic patients using either method. The clinical relevance of this finding requires further investigation. Different factors (e. g. injectant temperature, pulmonary VO2) produced substantial differences between VO2,IC and VO2,Fick as previously shown. PMID- 11271081 TI - Plasma cortisol levels before and during "low-dose" hydrocortisone therapy and their relationship to hemodynamic improvement in patients with septic shock. AB - OBJECTIVES: To compare cortisol levels during "low-dose" hydrocortisone therapy to basal and ACTH-stimulated endogenous levels and to assess whether clinical course and the need for catecholamines depend on cortisol levels and/or pretreatment adrenocortical responsiveness. DESIGN AND SETTING: Prospective observational study in a medical ICU of a university hospital. PATIENTS: Twenty consecutive patients with septic shock and a cardiac index of 3.5 l/min or higher, started on "low-dose" hydrocortisone therapy (100 mg bolus, 10 mg/h for 7 days and subsequent tapering) within 72 h of the onset of shock. MEASUREMENTS AND RESULTS: Basal total and free plasma cortisol levels ranged from 203 to 2169 and from 17 to 372 nmol/l. In 11 patients cortisol production was considered "inadequate" because there was neither a response to ACTH of at least 200 nmol/l nor a baseline level of at least 1000 nmol/l. Following the initiation of hydrocortisone therapy total and free cortisol levels increased 4.2- and 8.5-fold to median levels of 3,587 (interquartile range 2,679-5,220) and 1,210 (interquartile range 750-1,846) nmol/l on day 1, and thereafter declined to median levels of 1,310 nmol/l and 345 nmol/l on day 7. Patients with "inadequate" steroid production could be weaned from vasopressor therapy significantly faster, although their plasma free cortisol concentrations during the hydrocortisone treatment period did not differ. CONCLUSIONS: (a) During proposed regimens of "low-dose" hydrocortisone therapy, initially achieved plasma cortisol concentrations considerably exceed basal and ACTH stimulated levels. (b) Cortisol concentrations decline subsequently, despite continuous application of a constant dose. (c) "Inadequate" endogenous steroid production appears to sensitize patients to the hemodynamic effects of a "therapeutic rise" in plasma cortisol levels. PMID- 11271082 TI - Low flow inflation pressure-time curve in patients with acute respiratory distress syndrome. AB - OBJECTIVE: In mechanically ventilated patients with ARDS, determination of the lower (LIP) and upper (UIP) inflection points of the static pressure-volume curve (P-V) is crucial for planning ventilatory strategies. Recently, a simple new method was proposed for measuring the P-V curve by inflating the lung with constant low flow [14]. We hypothesized that during low flow inflation LIP and UIP might be determined using the pressure-time curve (P-T) instead of P-V. METHODS: Eleven paralyzed patients with ARDS were studied. During volume control ventilation the patients were allowed to reach passive functional residual capacity (FRC) and then ventilator frequency, inspiratory to total breath duration ratio and tidal volume (VT) were set to 5 breaths/ min, 80% and 500 or 1,500 ml, respectively. With these settings, constant inspiratory flow (V'I) was administered for 9.6 s and ranged, depending on VT, between 0.05 and 0.15 l/s. P V and P-T were obtained at two levels of positive end-expiratory pressure (PEEP; 0 and 10 cm H2O), with V'I being achieved either fast (< 0.1 s, minimum delay) or slowly (0.4 s, maximum delay). RESULTS: With minimum flow delay for a given experimental condition, the shape of the P-T did not differ from that of P-V. In all cases P-T correctly identified the presence of LIP and UIP, which did not differ significantly between P-T and P-V. With maximum flow delay, compared to P V, the initial part of P-T was significantly shifted to the left. P-T did not identify the presence of UIP and LIP in one and two cases, respectively. CONCLUSIONS: Provided that constant flow is given relatively fast, P-T accurately determines the shape of P-V, as well as the LIP and UIP. Flow delay causes a leftward shift of the initial part of P-T, masking the presence of LIP and UIP in some cases. PMID- 11271083 TI - Continuous positive airway pressure facilitates spontaneous breathing in weaning chronic obstructive pulmonary disease patients by improving breathing pattern and gas exchange. AB - OBJECTIVE: To elucidate the effects of continuous positive airway pressure (CPAP) on breathing pattern, gas exchange and the ability to sustain spontaneous breathing (SB) in chronic obstructive pulmonary disease (COPD) patients with dynamic hyperinflation. DESIGN: Prospective study with two randomised trials of SB without and with CPAP in each patient. SETTING: Medical intensive care units (ICUs) in two university hospitals. PATIENTS: Nine dynamically hyperinflated, intubated COPD patients recuperating from acute exacerbation. INTERVENTIONS: One SB trial with CPAP (5-7.5 cmH2O), one without (control) in each patient. MEASUREMENTS: airway opening pressure, gas flow and thus breathing pattern, oxygen uptake, carbon dioxide excretion, arterial blood gases, dyspnoea and respiratory drive (P100). RESULTS: With CPAP, intrinsic positive end-expiratory pressure (PEEPi) fell from 11.4 to 6.3 cm H2O (p < 0.05). Eight patients sustained SB with CPAP for the maximum time planned (30 min), one failed after 18 min. In contrast, only four patients successfully completed the control trial, the others failing after 5-18 min (p < 0.05). Dyspnoea-gauged on a visual analogue scale by five patients--was less severe or occurred later with CPAP. Breathing with CPAP tended to be slower (18.9 vs 22.2 min(-1), p < 0.05) and deeper (tidal volume 370 vs 323 ml). At the end of the control run, PaCO2 was higher (60 vs 55 mmHg, p < 0.05) and still rising while being stable at the end of the CPAP trial. CONCLUSION: CPAP helps severely ill COPD patients sustain SB. Apparently it does so by promoting slower, deeper breathing and thus facilitating carbon dioxide elimination. PMID- 11271084 TI - Sampling rate causes bias in APACHE II and SAPS II scores. AB - OBJECTIVE: To study the effect of sampling rate of laboratory and haemodynamic data on severity scorings and predicted risk of hospital death. DESIGN: Prospective study. SETTING: Medical-surgical intensive care unit (ICU) with 23 beds in a university hospital. PATIENTS: Sixty-nine consecutive emergency admission patients. INTERVENTIONS: Blood samples were drawn from indwelling arterial lines for the laboratory tests of all variables contained in the APACHE II and SAPS II scores at 2-hourly intervals from the time of admission up to 24 h or earlier discharge or death of the patient. Haemodynamic data and temperature were collected either manually by the attending nurse once an hour or as 2-min median values automatically using a Clinical Information Management System (CIMS, Clinisoft, Datex-Ohmeda, Helsinki, Finland). Three sets of severity scores were obtained. (1) "Traditional" scores (haemodynamic data from manual records and laboratory values from tests taken at admission and subsequently on clinical basis only). (2) "CIMS" scores (haemodynamic data from 2-min median values and laboratory values prescribed on clinical indication) and (3) "High rate" scores (haemodynamic data from 2-min median values and laboratory values at 2-hourly intervals). Probability of hospital death was calculated using the SAPS II and APACHE II scores, respectively. RESULTS: Increasing the sampling rate of haemodynamic monitoring interval to 2-min from once per hour resulted in 7.8 % and 11.5 % increases (p < 0.001) in the APACHE II and SAPS II scores, respectively. The combined effect of increased sampling rate of haemodynamic and laboratory tests on the APACHE II and SAPS II scores was 14.4 % and 14.5 % compared to traditional scores (p < 0.001), respectively. The probability of hospital death increased from 0.23 and 0.21 ("traditional" SAPS II and APACHE II) to 0.31 and 0.25 ("high rate" SAPS II and APACHE II), respectively, and, because eight patients died, standardised mortality ratio (SMR) decreased from 0.53 to 0.41 (SAPS II) and from 0.60 to 0.50 (APACHE II). CONCLUSIONS: Increased sampling rate results in higher scores and lower SMR. Comparisons between hospitals using severity scores are biased due to differences in the sampling rates. PMID- 11271086 TI - Comparison of Sepsis-related Organ Failure Assessment (SOFA) score and CIS (cellular injury score) for scoring of severity for patients with multiple organ dysfunction syndrome (MODS). AB - OBJECTIVE: To evaluate the usefulness of cellular injury score (CIS) and Sepsis related Organ Failure Assessment (SOFA) score for determination of the severity of multiple organ dysfunction syndrome (MODS). DESIGN: A prospective observational study. SETTING: A medical and surgical intensive care unit (ICU) of a teaching hospital. PATIENTS: Forty-seven consecutive MODS patients. MEASUREMENTS AND RESULTS: SOFA score and CIS were measured every day for 12 months for 47 MODS patients. Comparison was made of the SOFA score and CIS for usefulness in the scoring of severity of MODS in 26 survivors and 21 non survivors. In addition, receiver operating characteristics (ROC) analysis was used to determine the usefulness of these two indexes as predictors of prognosis. No significant differences were found on admission between the survivors and non survivors, but significant differences between the two subgroups (p < 0.001) were found in maximum value within 1 week after admission and maximum value during the course of treatment for both indexes. Analysis of changes after admission indicated that significant differences between survivors and non-survivors began to appear on day 3 of admission for both indexes; at that time SOFA score began to deteriorate in the non-survivors while CIS began to improve in the survivors. ROC analysis demonstrated that the area under the ROC curve was 0.769 for SOFA scores and 0.760 for CIS. CONCLUSIONS: Both SOFA score and CIS sequentially reflected the severity of MODS. Furthermore, they were comparable in diagnostic value as predictors of prognosis. These findings may indicate the possibility that MODS is a summation of effects of cellular injury. In addition, sequential evaluation of both SOFA score and CIS would provide a more accurate prediction of prognosis than conventional methods. PMID- 11271085 TI - Validation of severity scoring systems SAPS II and APACHE II in a single-center population. AB - OBJECTIVE: To validate two severity scoring systems, the Simplified Acute Physiology Score (SAPS II) and Acute Physiology and Chronic Health Evaluation (APACHE II), in a single-center ICU population. DESIGN AND SETTING: Prospective data collection in a two four-bed multidisciplinary ICUs of a teaching hospital. PATIENTS AND METHODS: Data were collected in ICU over 4 years on 1,721 consecutively admitted patients (aged 18 years or older, no transferrals, ICU stay at least 24 h) regarding SAPS II, APACHE II, predicted hospital mortality, and survival upon hospital discharge. RESULTS: At the predicted risk of 0.5, sensitivity was 39.4 % for SAPS II and 31.6 % for APACHE II, specificity 95.6 % and 97.2 %, and correct classification rate 85.6 % and 85.5 %, respectively. The area under the ROC curve was higher than 0.8 for both models. The goodness-of-fit statistic showed no significant difference between observed and predicted hospital mortality (H = 7.62 for SAPS II, H = 3.87 for APACHE II; and C = 9.32 and C = 5.05, respectively). Observed hospital mortality of patients with risk of death higher than 60 % was overpredicted by SAPS II and underpredicted by APACHE II. The observed hospital mortality was significantly higher than that predicted by the models in medical patients and in those admitted from the ward. CONCLUSIONS: This study validates both SAPS II and APACHE II scores in an ICU population comprised mainly of surgical patients. The type of ICU admission and the location in the hospital before ICU admission influence the predictive ability of the models. PMID- 11271087 TI - Retrospective evaluation of the simplified Therapeutic Intervention Scoring System (TISS-28) in a surgical intensive care unit. AB - OBJECTIVE: To compare the simplified Therapeutic Intervention Scoring System (TISS-28) with its original version, to provide reference values of daily TISS-28 assessment and to describe its association with severity of illness in surgical patients. DESIGN: Retrospective evaluation of prospectively collected audit data; four documentation periods. SETTING: Ten-bed intensive care unit (ICU) in a surgical university hospital. PATIENTS: One thousand nine hundred eighty-six consecutive admissions (1,808 patients; 10,448 observation days) who stayed on ICU for at least 6 h. Patients were in hospital for abdominal, vascular or trauma surgery. The average age was 61.5 years, the mean APACHE II score on admission 10.3 points. INTERVENTIONS: None. MEASUREMENTS: Raw data for APACHE II score and TISS were recorded daily. TISS-28 was calculated retrospectively from the original TISS data. RESULTS: Average TISS-28 values (28.7 points; SD = 9.7) do not differ substantially from the original TISS values (28.2 points, SD = 10.9) and overall correlation is high (r = 0.935). Of the patients, 57.3 % left the ICU after 1-2 days as survivors with a mean daily TISS-28 of 20.0 points. Variability between documentation periods was higher with the original TISS. On average, patients with increasing severity of disease require an increasing amount of care. Survivors have lower TISS-28 values than non-survivors (27.6 vs 34.9). CONCLUSIONS: In a surgical ICU the simplified version of TISS with 28 items (TISS 28) sufficiently reflects the amount of intensive care provided and may provide useful additional information on severity of disease and prognosis. It should replace the original index, at least in these cases. PMID- 11271088 TI - A comparison of severity of illness scoring systems for elderly patients with severe pneumonia. AB - OBJECTIVE: To evaluate the predictive ability of three severity of illness scoring systems in elderly patients with severe pneumonia requiring mechanical ventilation compared to a younger age group. DESIGN: Prospective cohort study. SETTING: Two university-affiliated tertiary care hospitals. PATIENTS AND PARTICIPANTS: One hundred four patients 75 years of age and older and 253 patients younger than 75 years of age enrolled from medical intensive care units. MEASUREMENTS AND RESULTS: Probabilities of hospital death for patients were estimated by the Acute Physiology and Chronic Health Evaluation (APACHE) II, the Mortality Probability Model (MPM) II and the Simplified Acute Physiology Score (SAPS) II. Predicted risks of hospital death were compared with observed outcomes using three methods of assessing the overall goodness of fit. The actual mortality of the elderly group was 54.87 % (95 % confidence interval [CI]: 45.2 64.4 %) compared to 28.9 % (95 % CI, 23.3-34.4 %) in the younger age group. There was a significant difference in the predictive accuracy of the scoring systems as assessed by the c-index, which is equivalent to the area under the receiver operator characteristics (ROC) curve, between the two groups, but not within individual groups. Calibration was insufficient for APACHE II and SAPS II in the elderly cohort as in-hospital mortality was lower than the predicted mortality for both models. CONCLUSIONS: Although the three severity of illness scoring systems (APACHE II, MPM II and SAPS II) demonstrated average discrimination when applied to estimate hospital mortality in the elderly patients with severe pneumonia, MPM II had the closest fit to our database. Alternative modeling approaches might be needed to customize the model coefficients to the elderly population for more accurate probabilities or to develop specialized models targeted to the designed population. PMID- 11271089 TI - Predictive factors of death in primary lung cancer patients on admission to the intensive care unit. AB - OBJECTIVE: To assess the lung cancer patient's prognosis in the intensive care unit with early predictive factors of death. DESIGN: Retrospective study from July 1986 to February 1996. SETTING: Medical intensive care unit at a university hospital. PATIENTS: Fifty-seven patients with primary lung cancer admitted to our medical intensive care unit (MICU). MEASUREMENTS AND RESULTS: Data collection included demographic data (age, sex, underlying diseases, MICU admitting diagnosis) and evaluation of tumor (pathologic subtypes, metastases, lung cancer staging, treatment options). Three indexes were calculated for each patient: Karnofsky performance status, Simplified Acute Physiology Score (SAPS) II, and multisystem organ failure score (ODIN score). Mortality was high in the MICU: 66% of patients died during their MICU stay, and hospital mortality reached 75%. In multivariate analysis, acute pulmonary disease and Karnofsky performance status < 70 were associated with a poor MICU and post-MICU prognosis. For the survivors, long-term survival after MICU discharge depended exclusively on the severity of the lung cancer. CONCLUSIONS: We confirmed the high mortality rate of lung cancer patients admitted to the MICU. Two predictive factors of death in MICU were identified: performance status < 70 and acute pulmonary disease. PMID- 11271090 TI - Predictors of short-term mortality in critically ill patients with solid malignancies. AB - Admission of cancer patients with serious medical complications to the ICU remains controversial primarily because of the high short-term mortality rates in these patients. However, the cancer patient population is heterogeneous regarding age, underlying conditions, and curability of their disease, suggesting that large variations may occur in the effectiveness of intensive care within this subgroup of critically ill patients. OBJECTIVES: To identify factors predicting 30-day mortality in patients with solid tumors admitted to a medical ICU. PATIENTS AND METHODS: We conducted a retrospective study in 120 consecutive cancer patients (excluding patients with hematological malignancies) admitted to the medical ICU of a 650-bed university hospital between January 1990 and July 1997. Medical history, physical and laboratory test findings at admission, and therapeutic interventions within the first 24 h in the ICU were recorded. The study endpoint was vital status 30 days after ICU admission. Stepwise logistic regression was used to identify independent prognostic factors. RESULTS: The observed 30-day mortality rate was 58.7 % (n = 68), with most deaths (92 %) occurring in the ICU. Univariate predictors of 30-day mortality were either protective [prior surgery for the cancer (p = 0.01) and complete remission (p = 0.01)] or associated with higher mortality [Knaus scale C or D (p = 0.02), shock (p = 0.04), need for vasopressors (p = 0.0006) or for mechanical ventilation (p = 0.0001), SAPS II score greater than 36 (p = 0.0001), LOD score greater than 6 (p = 0.0001), and ODIN score > 2 (p = 0.0001)]. Three variables were independent predictors: previous surgery for the cancer (OR 0.20, 95 % CI 0.07-0.58), LOD score > 6 (OR 1.26, 95 % CI 1.09-1.44), and need for mechanical ventilation (OR 3.55, 95 % CI; 1.26-6.7). Variables previously thought to be indicative of a poor prognosis (i. e., advanced age, metastatic or progressive disease, neutropenia or bone marrow transplantation) were not predictive of outcome. CONCLUSION: When transfer to an ICU is considered an option by patients and physicians, 30-day mortality is better estimated by an evaluation of acute organ dysfunction than by the characteristics of the underlying malignancy. PMID- 11271091 TI - Costs of care, long-term prognosis and quality of life in patients requiring renal replacement therapy during intensive care. AB - OBJECTIVE: To assess (1) the long-term outcome of patients requiring renal replacement therapy (RRT) in terms of 6-month and 5-year mortality, (2) quality of life and (3) costs of the intensive care. DESIGN: A retrospective observational cohort study. SETTING: Twenty-three-bed multidisciplinary intensive care unit (ICU) in a tertiary care center. PATIENTS AND PARTICIPANTS: Out of 3,447 intensive care patients admitted, 62 patients with no end-stage renal failure required RRT. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: The incidence rate of acute renal failure (ARF) was 8/100,000 inhabitants/ year. The majority of patients (71%) had ARF in conjunction with multiple organ failure. The mortality in the ICU and in the hospital was 34 % and 45%, respectively. Mortality was 55% at 6 months and 65 % at 5 years. Renal function recovered in 82 % of the survivors during hospitalization. Loss of energy and limitations of physical mobility assessed by Nottingham Health Profile were the most frequently reported complaints at 6 months. Functional ability, as assessed by the Activities of Daily Living score was fairly good at 6 months. The cost per ARF 6 month survivor was $80,000. CONCLUSIONS: There was only a minor increase in mortality after discharge from hospital among patients treated for ARF in intensive care. The costs related to ARF in intensive care are high, but the almost complete physical and functional recovery seen in ARF survivors should be noted in cost-effective analyses. PMID- 11271092 TI - Low seroprevalence of Helicobacter pylori infection in patients with stress ulcer bleeding--a prospective evaluation of patients on a cardiosurgical intensive care unit. AB - OBJECTIVE: The pathogenesis of stress ulceration in seriously ill patients is uncertain and the pathogenic role of Helicobacter pylori infection is unknown. We therefore assessed the seroprevalence of patients of a cardiosurgical intensive care unit (ICU) with clinically important stress ulcer bleeding. We compared this prevalence with a control group matched for this kind of surgical intervention, missing history of peptic ulcer disease, age and gender. DESIGN: Prospective survey. SETTING: Cardiosurgical ICU in a university teaching hospital. PATIENTS AND PARTICIPANTS: Two thousand five hundred seventy cardiosurgical patients with intravenous ranitidine stress ulcer prophylaxis were screened for clinically important stress ulcer bleeding. Helicobacter pylori seropositivity was measured in all patients with a clinically important bleeding and in a control group of 245 consecutive cardiosurgical patients, matched for the kind of cardiosurgical intervention, age and gender. RESULTS: In 56 of 2,570 (2.1%) patients signs of clinically important bleeding were seen. Endoscopical examination revealed stress ulcer bleeding in 42 cases. The incidence of stress ulcer bleeding was 1.6%. The seropositivity of the group with ulcer bleeding was 45.2 % whereas 62.4 % of the patients in the control group were Helicobacter pylori positive (p = 0.08). CONCLUSIONS: Our results suggest that the Helicobacter pylori infection does not play a pathogenic role in stress ulcer bleeding. Prophylactic cure of Helicobacter pylori can not be recommended in this setting. PMID- 11271093 TI - Accuracy of totally implanted ports, tunnelled, single- and multiple-lumen central venous catheters for measurement of central venous pressure. AB - OBJECTIVE: To verify the accuracy of totally implanted ports, tunnelled central venous catheters (CVC), widely used in cancer patients, and multi-lumen catheters, used in intensive care units (ICUs), in measuring central venous pressure (CVP), using right atrial pressure (RAP) measured in a Swan-Ganz catheter as the reference standard. DESIGN: A prospective study, over a 10-month period. SETTING: A medical-surgical ICU in a comprehensive cancer centre. PATIENTS AND PARTICIPANTS: Patients who had both (1) a Swan-Ganz catheter and (2) either a tunnelled catheter, a single or a multi-lumen catheter, or a totally implanted port. INTERVENTIONS: RAP and CVP were measured simultaneously in each patient. MEASUREMENTS AND RESULTS: Fifty-six pairs of RAP-CVP measurements were performed in 35 patients: 6 tunnelled catheters, 6 non-tunnelled single-lumen catheters, 26 multiple-lumen catheters and 18 totally implanted ports were studied. RAP measured in the Swan-Ganz catheter and CVP measured in the CVC were strongly correlated (r = 0.94, p < 0.01), whatever the type of catheter studied. The mean difference between RAP and CVP was -0.39 +/- 1.73 (SD) mmHg. In 51 cases (91%), the difference was within the limits of agreement (-3.78 to 3.00 mmHg, Bland and Altman method). For the five cases with a difference of 4 mmHg (three totally implanted ports, one double- and one triple-lumen catheter), CVP was greater than RAP. CONCLUSIONS: CVP can be accurately measured in totally implanted ports, tunnelled or non-tunnelled single-lumen and multiple-lumen catheters. When the difference exceeds the limit of agreement, the discrepancy between the two measurements has limited significance in most cases. PMID- 11271094 TI - Hypothermia with indoor occurrence is associated with a worse outcome. AB - OBJECTIVE: To describe patients admitted to intensive care unit (ICU) for hypothermia, evaluate prognostic factors, and test the hypothesis that patients found indoors have a worse outcome. DESIGN AND SETTING: Retrospective clinical investigation in a medical ICU. PATIENTS: Eighty-one consecutive patients admitted to ICU, with a body temperature of 35 degrees C or lower and rewarmed passively or with minimally invasive techniques, over a 17-year period. MEASUREMENTS AND RESULTS: Patients were analyzed by age, gender, and causes of hypothermia and split into two groups (indoors and outdoors), according to the location where hypothermia occurred. Prognostic factors were determined by univariate method and stepwise logistic regression. The major complications were acute renal failure (43 %), aspiration pneumonia (22 %), rhabdomyolysis (22 %), and acute respiratory distress syndrome (12%). Principal comorbidities in the outdoor patients (21%) were alcohol and drug intoxication, and those in the indoor patients (79 %) were sepsis and neuropsychiatric disorders. Stepwise logistic regression identified two variables predictive of death: illness severity at admission (SAPS II > or = 40) and the location where hypothermia occurred (indoors versus outdoors). CONCLUSIONS: With equivalent body temperature, patients found indoors were more severely affected and died more frequently than those found outdoors. PMID- 11271095 TI - Outcome of bedside percutaneous tracheostomy with bronchoscopic guidance. AB - OBJECTIVE: To determine the morbidity and mortality of percutaneous dilational tracheostomy with bronchoscopic guidance when performed by medical intensivists. DESIGN: A retrospective analysis. SETTING: A tertiary care university hospital. PATIENTS: Fifty consecutive patients who underwent percutaneous dilational tracheostomy for prolonged mechanical ventilation. INTERVENTION: Bedside percutaneous dilational tracheostomy with bronchoscopic guidance. RESULTS: Seventeen women and 33 men with a mean age of 62 +/- 17 years. Operative mortality was 0 with four (8%) operative complications. Complications included one posterior tracheal abrasion, one anterior tracheal laceration, one episode of endobronchial hemorrhage requiring bronchoscopy, and one pneumothorax. Thirty-day mortality was 28% and overall mortality was 40%. All deaths were related to the patients' underlying disease. CONCLUSIONS: Percutaneous dilational tracheostomy with bronchoscopic guidance is a safe procedure when performed by experienced medical intensive care personnel in tertiary care institutions. Bronchoscopy helps to reduce the risk of major complications and aids in the management of minor complications. PMID- 11271096 TI - ICU nurse-to-patient ratio is associated with complications and resource use after esophagectomy. AB - OBJECTIVE: To determine if having a night-time nurse-to-patient ratio (NNPR) of one nurse caring for one or two patients (> 1:2) versus one nurse caring for three or more patients (< 1:2) in the intensive care unit (ICU) is associated with clinical and economic outcomes following esophageal resection. DESIGN: State wide observational cohort study. Hospital discharge data was linked to a prospective survey of ICU organizational characteristics. Multivariate analysis adjusting for case-mix, hospital and surgeon volume was used to determine the association of NNPR with in-hospital mortality, length of stay (LOS), hospital cost and specific postoperative complications. SETTING: Non-federal acute care hospitals (n = 35) in Maryland that performed esophageal resection. PATIENTS AND PARTICIPANTS: Adult patients who had esophageal resection in Maryland, 1994 to 1998 (n = 366 patients). MEASUREMENTS AND RESULTS: Two hundred twenty-five patients at nine hospitals had a NNPR > 1:2;128 patients in 23 hospitals had a NNPR < 1:2. No significant association between NNPR and in-hospital mortality was seen. A 39 % increase in median in-hospital LOS (4.3 days; 95% CI, (2, 5 days); p < 0.001), and a 32% increase in costs ($4,810; 95 % CI, ($2,094, $7,952) was associated with a NNPR < 1:2. Pneumonia (OR 2.4; 95 % CI (1.2, 4.7); p = 0.012), reintubation (OR 2.6; 95% CI(1.4, 4.5);p = 0.001), and septicemia (OR 3.6; 95 % CI(1.1, 12.5); p = 0.04), were specific complications associated with a NNPR < 1:2. CONCLUSIONS: A nurse caring for more than two ICU patients at night increases the risk of several postoperative pulmonary and infectious complications and was associated with increased resource use in patients undergoing esophageal resection. PMID- 11271097 TI - Management of coagulative disorders in the critically ill. PMID- 11271098 TI - Pneumopericardium, pneumomediastinum, pericarditis and mediastinal abscess secondary to diverticulitis of the sigmoid. PMID- 11271099 TI - Ammonium chloride poisoning: a misunderstood cause of metabolic acidosis with normal anion gap. PMID- 11271100 TI - Definition for septic syndrome should be re-evaluated. PMID- 11271113 TI - Caring for all: an old idea for a new millennium. PMID- 11271114 TI - Guide to safe prescribing. AB - In this article, we examine the common errors in prescribing medications and analyze the steps clinicians can take to develop and maintain good prescribing habits. Advanced practice nurses are encouraged to develop a personal formulary by using reliable resources and to use this formulary to write error-free prescriptions. PMID- 11271115 TI - Meeting the health needs of homeless or low-income persons: role of the nurse practitioner. AB - Providing health care for homeless or low-income families presents many challenges and rewards. Nurse practitioners are in a unique position to make a positive impact on the health needs of this population. However, they need to think about the ramifications of their approach, assessment techniques, and intervention strategies to optimize the health care offered. We share our experience in working with this population and lessons we have learned from review of the literature. PMID- 11271116 TI - Emotional regulation and alexithymia: treatment implications for nurse practitioners. AB - It has been estimated that between 50% and 80% of all visits to nurse practitioners are for symptoms that are vague, without a specific diagnosis or cause. Such symptoms as headaches and pain, as well as other illnesses, may stem from difficulties in emotional regulation. Understanding that these individuals suffer from emotional dysregulation and alexithymia provides guidelines for assessment and treatment protocols. Integrating the holistic therapeutic strategies delineated in this article is essential for expert advanced practice. PMID- 11271117 TI - Rational use of antibiotics for upper respiratory infections: an evidence-based approach. AB - Despite the lack of evidence of efficacy of antibiotic agents for treating upper respiratory tract infection (URI) symptoms (i.e., acute cough, sore throat, purulent nasal discharge, bronchitis, and the common cold), primary care providers frequently prescribe antibiotic agents for patients presenting with such symptoms. Far from being a harmless practice, prescribing antibiotics for conditions for which there is no proven benefit of such therapy contributes to a number of adverse consequences, including the development of antimicrobial resistance and an unnecessary expense to patients and the healthcare system as a whole. An evidence-based approach to practice can guide nurse practitioners in making the best clinical management decisions for patients presenting with URI symptoms. PMID- 11271118 TI - Risk factors for perinatal asphyxia at Queen Elizabeth Central Hospital, Malawi. AB - The aim of the study was to identify maternal risk factors for perinatal asphyxia in Malawi. Records of 100 mothers who delivered neonates with Apgar scores less than 6 at 5 minutes of birth during March to September 1998 were analyzed. The majority of the mothers were primigravidas (79%) and were within the normal childbearing ages of 20 to 34 years (61.2%). Sixty-one percent of the mothers started antenatal care at 20 to 28 weeks' gestation. Sixty-five percent of the mothers developed obstetric and medical problems that contributed to perinatal asphyxia, and of these, 12 mothers (18.5%) had more than one problem. The problems were premature labor and delivery (21%), preeclampsia (10%), cephalopelvic disproportion (8%), breech presentation (12%), prolonged second stage (11%), fetal distress (7%), cord prolapse (4%), antepartum hemorrhage (2%), prolonged rupture of membranes (1%), and malaria (1%). Forty-six percent had assisted deliveries, and these were cesarean section (18%), vacuum extraction (14%), breech delivery (12%), and forceps delivery (2%). Eighty-one percent of the neonates were admitted to the neonatal nursery, and of these, 56 neonates (67.1%) developed complications; the most common was hypoxic ischemic encephalopathy (38 neonates; 67.9%). Thirty-three percent of the neonates died within 6 days postdelivery. Morbidity and mortality related to perinatal asphyxia can be reduced if staff are knowledgeable and skilled in basic neonatal resuscitation and necessary equipment is available. Mothers should be encouraged to report early for antepartum and intrapartum care for adequate surveillance. The quality of neonatal care, with a focus on thermoregulation and infection prevention, needs to be improved. PMID- 11271119 TI - A Delphi survey to identify activities of nurse practitioners in primary care. AB - The practice activities of nurse practitioners (NPs) in primary care have not been well delineated. The purpose of this study is to identify the activities of NPs in the provision of primary care. Patient, practitioner, and healthcare system variables were collected to describe the sample and determine if these factors influenced the activities of NPs. Primary care NPs (n = 139) representing the practice areas of pediatrics (n = 12), family (n = 75), adult (n = 15), and women's health (n = 37) responded to serial questionnaires using Delphi technique asking them to identify activities they perform routinely in their practices. Using frequency analysis, activities common to the practice of all NPs were identified, and using Chi-square, activities of specialty NP practice were identified. Regression analysis was used to correlate NP, patient, and healthcare system variables with the activities performed as part of NP practice. Three lists of activities of NP practice were developed: core activities of all NP practice, specialty activities, and other activities. Initial analyses of variables that might influence NP practice were examined. These results can be useful to healthcare consumers choosing providers, as well as legislators, educators, employers of NPs, and other interested healthcare professionals. PMID- 11271120 TI - Nurse practitioners can cut costs and provide increased care. PMID- 11271121 TI - Early views of nurse practitioners: a Medline search. AB - Nurses have been functioning in expanded roles as nurse practitioners (NPs) since 1965. The role has been characterized by controversy regarding economics, scope of practice, level of independence, accountability, and training since its inception. This article examines the response of the medical community to NPs during the first 15 years of their development. The medical literature from 1967 through 1982 was examined through a Medline search to ascertain the issues, attitudes, and barriers to practice that evolved as a foundation for understanding the present response of medicine to NPs. PMID- 11271122 TI - Profiles of editorial board members: the DC connection. PMID- 11271123 TI - The disease of addiction. AB - Drug use is a preventable behavior, and addiction is a treatable disease. More than two decades of scientific research have yielded tremendous understanding concerning the nature of addiction and what to do about it. Many tools that can be used by health care providers to aid in the recognition, referral, and treatment for drug abuse and addiction also have been developed. To assist in these efforts, the National Institute on Drug Abuse recently has published the first ever science-based guide to drug treatment: Principles of Drug Addiction Treatment. This guide articulates some essential characteristics of addiction and its treatment and lays out the principles derived from two decades of scientific research that characterize effective treatment programs. This article elaborates many of those points. PMID- 11271124 TI - Addiction in patients with chronic pain. PMID- 11271125 TI - Assessment and treatment of addictions in primary care. AB - Most clinicians are faced with the challenge of providing care and treatment for patients who experience the chronic relapsing brain disease known as addiction. The purpose of this article is to increase awareness of techniques and tools available to primary care clinicians (PCCs) for assessing and treating addictions in the office or clinic setting. A review of the history, physical examination, laboratory tests, and diagnostics relevant to addictive illness will help PCCs to hone their skills in addiction management. Addiction screening instruments and brief interventions used in primary care are presented. Adjunct therapies designed to promote the biopsychosocial and spiritual well-being of patients who are addicted have shown promise. PMID- 11271126 TI - Women and substance use disorders. AB - In the United States, 45% of all women use alcohol, and nearly 26% smoke cigarettes. Approximately 13% use illicit substances, prescribed medications, or both in recreational ways. Although women use these substances less often than men, the health consequences of their use for women, particularly the use of alcohol, are either equivalent or greater. Women's social relationships play key roles in the onset of substance use disorders, treatment, and continued recovery. Major psychoactive substances include nicotine, alcohol, illicit drugs, and recreational use of nonprescribed medications. Biopsychosocial aspects unique to women are reviewed, followed by treatment approaches, concluding with implications for primary care providers. PMID- 11271127 TI - Recognition and management of addictive sexual disorders: guide for the primary care clinician. AB - With greater awareness of sexual exploitation, professional sexual misconduct, and the sexual issues of public figures, a growing awareness of the problem of sexual addiction is emerging. As a result of public awareness, more cases will be brought to the attention of primary care providers. When primary care providers are confronted by problematic sexual behavior that fits the parameters of addictive illness, they should know what the implications are in order to make appropriate clinical decisions and to evaluate treatment approaches. The purpose of this article is to summarize the nature of the problem, to review critical issues in assessment, to provide treatment options, and to suggest critical factors for monitoring progress. PMID- 11271128 TI - Pharmacology of substance abuse. PMID- 11271130 TI - For the recovering individual. PMID- 11271129 TI - Inhalant abuse. PMID- 11271131 TI - Gambling addiction. PMID- 11271132 TI - Marijuana. PMID- 11271133 TI - Perinatal drug and alcohol addiction: the role of the primary care provider. PMID- 11271134 TI - To whom am I listening? PMID- 11271135 TI - Becoming and remaining homeless: a qualitative investigation. AB - This article reports the qualitative findings of a multimethod study of the homeless population in Toronto, Canada. The qualitative component sought to identify how people become homeless and why some individuals remain homeless for an extended period of time or cycle in and out of homelessness (the chronically homeless). In-depth, semistructured interviews were conducted with 29 homeless adults. The findings suggest that people both become and remain homeless due to a combination of macro level factors (poverty, lack of employment, low welfare wages, lack of affordable housing) and personal vulnerability (childhood abuse or neglect, mental health symptoms, impoverished support networks, substance abuse). Chronically homeless individuals often reported experiences of severe childhood trauma and tended to attribute their continued homelessness to a substance abuse problem. It is concluded that both macro and individual level factors must be considered in planning programs and services to address the issue of homelessness in Canada. PMID- 11271136 TI - Depression in schizophrenia: nursing care as a generalized resistance resource. AB - Although depression can be a feature of acute psychosis and an aftermath of a psychotic episode (postpsychotic depression), data indicate that depressive syndromes in schizophrenia can be found years after the immediate postacute phase. Researchers acknowledge the clinical importance of recognizing that depressive symptoms are frequently described in patients diagnosed with schizophrenia. Using the framework of Antonovsky's (1972, 1987, 1992) Sense of Coherence as a guide, this article explores what nurses can do to facilitate development of generalized resistance resources (GRRs) in the physical, psychological, and emotional armamentarium of those diagnosed with schizophrenia, mood disorder not otherwise specified (NOS). PMID- 11271137 TI - Infertility as a transformational process: a framework for psychotherapeutic support of infertile women. AB - The purpose of this qualitative descriptive study was to investigate the phenomenon of infertility as experienced by infertile women. A purposive sample of 25 infertile women participated in the study. Data were extracted from taped interviews and the researcher's observational field notes. Data analysis was conducted according to the techniques described by Miles and Huberman (1994). Participant responses to interviews were categorized by examining the interview transcripts and identifying significant statements and meanings. Themes which emerged from the statements were then ascertained and cross-case comparisons were made in order to confirm or to reconsider these themes. Five key themes emerged from the data: failure to fulfill a prescribed societal norm, assault on personal identity, mourning, transformation, and restitution. The women experienced infertility as a transformational process in which they mourned their loss of reproductive function and parenting roles and struggled to make restitution for the perceived stigma and powerlessness associated with nonfulfillment of a prescribed societal norm, the exclusion from cherished societal rituals, and the deprivation of ties of descent. Findings from this study have provided a framework for increased awareness of the phenomenon of infertility and for the essential components of supportive counseling or psychotherapy, regardless of the outcome of the infertility experience. PMID- 11271138 TI - Hispanic women and AIDS: gendered risk factors and clinical implications. AB - Current theory and research indicate that Hispanic women's increased risk for AIDS is due to sociocultural and psychological factors that are influenced by gender. Hispanic women's low social status limits their ability to negotiate safer sex behaviors for AIDS prevention. Female gender identity, based upon the desire for interpersonal attachment, places Hispanic women at risk for AIDS through a fear of abandonment by male sexual partners who perceive safer sex behaviors as unmasculine. Male sexual partners' beliefs, practices, and behaviors, such as bisexuality and the practice of anal intercourse, increase Hispanic women's AIDS risk. The experience of involuntary sexual encounters and the lack of power to determine sexual behaviors and practices within these encounters also place Hispanic women at risk for AIDS. PMID- 11271139 TI - Marital conflict management skills, parenting style, and early-onset conduct problems: processes and pathways. PMID- 11271140 TI - African-American fathers in low-income, urban families: development, behavior, and home environment of their three-year-old children. PMID- 11271141 TI - Ten years after: examination of adolescent screening questions that predict future violence-related injury. PMID- 11271142 TI - Symptoms of depression in adolescents with epilepsy. PMID- 11271143 TI - Hopefulness and its characteristics in adolescents with cancer. PMID- 11271144 TI - Health status of children with special health care needs: measurement issues and instruments. PMID- 11271145 TI - Psychiatric comorbidity and 16-month trajectory of substance-abusing and substance-dependent juvenile offenders. PMID- 11271146 TI - Factors associated with the development of substance use disorder in depressed adolescents. PMID- 11271147 TI - Approaches to the measurement of depressive symptomatology in children with cancer: attempting to circumvent the effects of defensiveness. PMID- 11271148 TI - Asthma knowledge, roles, functions, and educational needs of school nurses. PMID- 11271149 TI - Pediatric registered nurse usage and perception of EMLA. PMID- 11271150 TI - High tech - high touch nursing care. PMID- 11271151 TI - Qualitative research interviews with children and their families. AB - Regulations about conducting research with children, development of the field of qualitative research, and sophistication of our knowledge about children has prompted increased recognition of children as research subjects are described. Guidelines for how to prepare and conduct qualitative interviews with children and their families are summarized in Table 1. While gathering research data from children and their families presents many challenges, qualitative interview methods are a potentially rich source of data. PMID- 11271152 TI - Violence in our schools. Sources of prevention for pediatric nurses. PMID- 11271153 TI - Pandora's box. Educational materials resource assessment. PMID- 11271154 TI - Agency for Health Care Policy and Research. Rehabilitation for traumatic brain injury in children and adolescents: summary. PMID- 11271155 TI - Asthma self-management programs for children. Part I: Description of the programs. PMID- 11271156 TI - Intermittent subcutaneous injections for symptom control in hospice care: a retrospective investigation. AB - An alternative route to oral medications used by some hospice programs is intermittent injections of medications using an indwelling subcutaneous butterfly needle. The nurse places the infusion sets and instructs caregivers on medication administration. Although this method has become more common in hospice care, it has not received much attention in part because of a lack of data to support its efficacy. This study describes the use of intermittent subcutaneous medications for symptom relief in a home hospice program. A chart review was conducted of the 191 patients who received medications by this route during three calendar years; 77% had cancer. The average duration of hospice care was 25 days; on average, intermittent subcutaneous medications were instituted 4 days prior to the patient's death. The main indications for this route were inability to swallow/somnolence (65%), and pain unresponsive to oral medication (19%). Symptoms to be controlled by this method were pain (88%), anxiety (72%), and dyspnea (4%). Morphine was used most frequently for pain, and Ativan was used most frequently for anxiety. Side effects from the medications and problems with this route of administration were rarely reported, thereby supporting the practicality of this method in hospice care. These results form the foundation for a prospective study that is documenting staff, patient, and caregiver variables that impact on the effectiveness and manageability of this method of symptom management in hospice care. PMID- 11271157 TI - The place of spiritual well-being in hospice patients' overall quality of life. AB - There is an increasing awareness of, and interest in the relationship between spirituality and health. This research examines spiritual well-being as one of six components of hospice patients' overall quality of life. Patients admitted over a four-month period were surveyed, using the Functional Assessment of Cancer Therapy scale (FACT-G), at admission, one month later, three months later, and six months later. Data showed spiritual well-being to be an important contributor to overall quality of life. The article concludes by advocating that providing spiritual care to hospice patients makes good business sense. PMID- 11271158 TI - Predictors of family members' satisfaction with hospice. AB - This large, long-term study of families served by hospice found that nearly 95 percent said hospice had been helpful. Still, about 30 percent of family members said there was something they wish hospice had done differently. Those who had some complaint were more likely than those who had no complaints to be women, to report the patient had needed a great deal of care, to have a history of depression and greater levels of distress before and after the patient's death, and to be dissatisfied with the support they received from family members and friends. PMID- 11271159 TI - Assessing readiness for death in hospice elders and older adults. AB - BACKGROUND: Readiness for death may affect the quality of the death experience and influence response to treatments. The psychologic vulnerability of the dying person is a major focus of palliative care. Accurate assessment of readiness for death may lead to earlier and more appropriate interventions. OBJECTIVE: The purpose of this study was to assess the psychometric properties of the revised readiness for death instrument. METHODS: Using a known groups technique and a cross-sectional study design, the revised instrument was administered to 52 elders in hospice care with a terminal diagnosis and 91 community dwelling adults without a terminal diagnosis. RESULTS: Instrument content validity (Kappa = 0.96) was supported by three expert panelists who were hospice researchers. Principal components factor analysis explained 43% of the variance and partially supported the proposed four-factor structure of the revised 26-item instrument. Internal consistency was acceptable (.76). Discriminant validity was significant as assessed by an independent t-test between two contrast groups (t = 5.98, p = 0.000). The factor analysis, reliability testing, and qualitative analysis of items supported deletion of 2 items. CONCLUSIONS: Results indicated that the revised instrument has sound psychometric properties but further testing with a larger sample of hospice subjects is needed to confirm the factor structure of the instrument. PMID- 11271160 TI - Physician attitudes toward palliative care at a community teaching hospital. AB - The goals of the study were to explore physicians' attitudes and opinions about palliative care and its implementation. Four focus groups composed of attending physicians were conducted by a professional facilitator at a community teaching hospital. The audio-tapes of the groups were carefully transcribed and analyzed according to rigorous qualitative methodology. Physicians perceived palliative care and pain control as important. Problems they perceived were a lack of education for physicians, residents, other health care professionals, and the general public; a lack of hospital support systems to implement palliative care appropriately, and a lack of knowledge and support regarding legal considerations. They believed that a palliative care unit was a reasonable tool to overcome many obstacles to good end-of-life care. PMID- 11271161 TI - Papers and abstracts from the 9th International Symposium on Antiphospholipid Antibodies. Tours, France, September 12-16, 2000. PMID- 11271162 TI - Hospitals to spend 200m pounds sterling to prevent spread of vCJD. PMID- 11271163 TI - Pen "amnesty" for doctors who shun drug companies. PMID- 11271164 TI - Plasma homocysteine and MTHFR C677T genotype in levodopa-treated patients with PD. PMID- 11271165 TI - The cingulate hidden hand. PMID- 11271166 TI - Localised myelitis caused by visceral larva migrans due to Ascaris suum masquerading as an isolated spinal cord tumour. PMID- 11271167 TI - Effect of lower limb position on ankle jerk assessment. PMID- 11271168 TI - Randomised controlled trial of occupational therapy at home: results at 1 year. PMID- 11271169 TI - Relation between Glasgow outcome score extended (GOSE) and the EQ-5D health status questionnaire after head injury. PMID- 11271171 TI - Richard H. Osborne, retiring editor: an appreciation, 1977. PMID- 11271170 TI - Frederick Osborn, demography's statesman, on his eightieth spring: March, 1969. PMID- 11271172 TI - Behavior Genetics Association April 6, 1973. Is genetic diversity compatible with human equality? 1973. PMID- 11271173 TI - Pathogen-induced expression of plant ATP: citrate lyase. PMID- 11271174 TI - Ernennung neuer korrespondierendeer Mitglieder (Appointments of new corresponding members). PMID- 11271175 TI - The use of ABO-compatible mismatched livers in the UK. AB - Elective blood group O liver recipients appear to wait longer than most other groups for matched donors. The aim of this study was to confirm the suspected differences in elective waiting times in the UK using data from the United Kingdom Transplant Support Service, and to determine some of the factors responsible for them. The findings were that potential group O recipients waited significantly longer than other groups for transplantation, and that 22% of group O livers were going to non-O recipients. AB, the group with the shortest waiting time, was receiving 74.5% mismatched (but compatible) grafts, from all other groups. PMID- 11271176 TI - Orthotopic liver transplantation for acute hepatic failure in children. AB - Thirty children received 35 liver transplants for fulminant or late-onset liver failure between March 1988 and May 1993. Aetiology included non-A non-B hepatitis in 12, Wilson's disease in 8, drug-induced hepatic failure in 6, hepatitis B in 1, hepatitis A in 1, tyrosinaemia in 1 and congenital haemochromatosis in 1. Three patients were retransplanted, one each for hepatic artery thrombosis, non-A non-B graft reinfection, and chronic rejection. Two of these three patients received a third transplant for chronic rejection and hepatic artery thrombosis. One patient in the retransplant group survived. Overall, graft and patient survival at a mean follow-up of 17 months were 49% and 57%, respectively. Mortality was related to vascular complications in three patients (hepatic venous obstruction, portal vein thrombosis and hepatic artery thrombosis). Two patients died of primary sepsis (cerebral aspergillosis and cytomegalovirus (CMV) pneumonitis in association with graft-versus-host disease). Systemic sepsis and multiorgan failure was documented as a cause of death in four children and sepsis in association with chronic rejection in a further three patients. One child died of respiratory failure 4 weeks after transplantation. Mortality in eight children less than 2 years was 75% and this was significantly greater than for older children (P < 0.003, Mantel Cox). Earlier referral, even in the absence of a definitive diagnosis and particularly in children under 2 years is advisable and may improve survival. PMID- 11271177 TI - Living-related liver transplantation for fulminant hepatic failure in children. AB - Liver transplantation is increasingly accepted as a choice of treatment for fulminant hepatic failure (FHF) since it has been proved to significantly improve the survival rate in these patients compared with other therapeutic modalities. We have successfully performed a total of 76 living related liver transplantations (LRLT) three of which were for FHF. The first case was an 11 year-old boy with FHF due to an unidentified cause. He had required plasmapheresis a total of 24 times and haemofiltration to save his life before LRLT. He was transplanted with a left lobe (420 g) graft, calculated as 1.05% of his weight (40 kg). He recovered hepatic function uneventfully and was discharged from hospital after 7 weeks. The second case was a 13-year-old girl who developed FHF with grade III encephalopathy due to acute Wilson's disease, and was referred to us. She underwent LRLT with a left lobe graft (440 g), estimated as 0.95% of her weight (47 kg), which functioned well after surgery. The third case was a 13 year-old girl with grade II encephalopathy due to acute Wilson's disease. She was 27% obese with a body weight of 58 kg. She underwent LRLT with ABO blood group incompatibility with a left lobe (352 g), estimated as 0.80% of her weight (modified 44 kg). She was discharged with sensorimotor neuropathy due to vitamin B deficiency. The present results suggest that LRLT is feasible for FHF both clinically and ethically, and that a partial liver graft weighing around 1% of the recipient's weight can maintain the recipient's life. We limit the diagnostic indication for LRLT to chronic liver disease, since an urgent situation may affect a voluntary decision for the patient's parents to donate the partial liver. However, LRLT is thought to be an acceptable choice of treatment provided it is requested by the patient and family. Furthermore, it is a potential option for resolving the graft shortage in paediatric liver transplantation, being independent of cadaver donor availability. PMID- 11271178 TI - Glucose metabolism in liver transplant recipients treated with FK 506 or cyclosporin in the European multicentre study. AB - From September 1990 to January 1992, 545 liver transplant patients were randomised to treatment with either FK506 and prednisolone or a conventional cyclosporin-based immunosuppressive regimen (CBIR). Eight European centres participated in the study. Adverse events were reported as defined by each centre. Hyperglycaemia was reported as an adverse event in 30.7% of patients receiving FK 506 compared with 20.5% in the CBIR group (P < 0.01). Diabetes mellitus was reported in 17.2% of patients treated with FK 506 and 9.5% of CBIR treated patients (P < 0.05). Treatment with insulin was required in 12.0% of patients in the DK 506 treatment group and in 5% in the CBIR group at 6 months. Initially, higher doses of FK 506 were used. During the study, the protocol was changed to allow a lower dose of FK 506. When the early and late cohorts of patients were compared, the incidence of diabetes mellitus fell from 23.9% to 10.5% in FK 506-treated patients but remained relatively constant in the CBIR group (10.4% to 8.7%). The median cumulative doses of i.v. and p.o. corticosteroids were significantly greater in the CBIR group. Thus, in the overall series, the incidence of diabetes mellitus was significantly greater in the FK 506 group as compared with the CBIR group. However, when a lower FK 506 dose was used during the second half of the study, the difference in the incidence of diabetes mellitus disappeared. PMID- 11271179 TI - Living related liver transplantation in children: a report of the first 58 recipients at the University of Chicago. AB - From November 1989 58 living donor liver transplants were performed in 56 children ranging in age from 1 month to 13 years. Donors were adults (> 18 years of age) with a close relationship to the recipient. ABO compatibility and normal donor health were required. Liver segments two and three were transplanted in 53 cases, and segments two, three and four in 5 cases. Actuarial patient survival at 2 years was 89%; graft survival was 76%. Six recipients died: four secondary to sepsis and two because of post-transplant lymphoproliferative disease. The main cause of graft loss was arterial thrombosis, occurring in six patients (10%). Since refinement of the technique, there have been few donor complications, but these have included a biliary tract injury and a hepatic artery thrombosis. Both donors are well, without long-term adverse sequelae. Overall, the outcome of living donor transplantation is excellent; morbidity has been encountered in a small number of donors. PMID- 11271181 TI - Portosystemic shunt for the treatment of portal vein thrombosis following orthotopic liver transplantation. AB - The efficacy of the portosystemic shunt operation for the treatment of portal vein thrombosis following orthotopic liver transplantation was demonstrated. From 1 July 1988 to 31 December 1991 42 portosystemic shunt operations were performed at our centre. In six of these cases portal vein thrombosis after orthotopic liver transplantation (OLT) was the indication for the procedure. All the patients retained adequate liver function but they demonstrated manifestations of significant portal hypertension, mainly variceal rebleeding. Two of the patients were children. Three patients underwent distal splenorenal shunt (DSRS), one mesocaval and one side-to-side splenorenal shunt and the last one side-to-side splenorenal shunt which was converted to DSRS 2 weeks later. All these patients were doing well after 30 months mean follow-up time without rebleeding or other signs of portal hypertension and none had so far required retransplantation. PMID- 11271180 TI - The effect of venovenous bypass on lactic acid levels during human liver transplantation (OLT). AB - Lactate determinations did not contribute to the quantification of the systemic and regional tissue oxygenation during OLT. Venous stasis was not an important factor in the tissue imbalance between oxygen supply and oxygen demand. PMID- 11271182 TI - Biliary complications in 100 adult liver transplantations: a retrospective clinical study. AB - Biliary complications were reviewed in 100 consecutive adult liver transplantations. Included in the study were 92 patients surviving for more than 1 month. In 86 transplantations biliobiliary anastomosis was performed with (n = 25) or without (n = 61) a T-tube. In six cases biliodigestive anastomosis (Roux en-Y) was performed. Biliary stricture caused by hepatic arterial thrombosis was not included. Biliary complications were seen in 17 cases: seven anastomotic strictures, four T-tube-related leakages, four anastomotic leakages, one leakage of unknown origin and one late cholangitis. Nine were surgically treated (six strictures and three leakages). Patients with primary sclerosing cholangitis had the highest biliary complication rate (36%). Early anastomotic strictures were associated with a higher rate of major bacterial infections (P = 0.03) and CMV disease (P = 0.08) than those without biliary complications. Biliobiliary anastomosis with a T-tube was associated with more complications (28%) than anastomoses without a T-tube (13%). To date, total patient survival including all 100 transplantations was 71% (median follow-up 3.3 years). We conclude that biliary complications are rather common but they do not affect survival and can be treated. Biliary T-tubes can be omitted. PMID- 11271183 TI - A simple method for predicting bone fractures in PBC patients after liver transplantation. AB - We introduce a simple serum test to predict which patients will have bone problems after liver transplantation. The crosslinked part of collagen I (s-ICTP) was measured in 21 patients with primary biliary cirrhosis before transplantation. Those with postoperative fractures had increased pretransplant values of s-ICTP compared with those without fractures. PMID- 11271184 TI - Transplantation for alcoholic cirrhosis: how does recurrence of disease harm the graft? AB - Many transplant centres are reluctant to accept alcoholic patients because of their supposed potential for alcoholic recidivism, resulting in graft failure and recurrence of alcoholic liver cirrhosis. From May 1982 to January 1993 80 patients received orthotopic liver transplantation (OLT) at our institution either for alcoholic cirrhosis exclusively (n = 58) or for a hepatoma in an alcoholic cirrhosis (n = 22). The outcome of these patients was analysed with particular attention to recurrence of liver disease. Overall survival in this group was 67% and 49% at 1 and 5 years, respectively, with a median follow-up of 45 months. Actuarial survival of patients transplanted since January 1989 (n = 46) and 84% and 82% at 1 and 2 years (median follow-up 31 months). Non-fatal clinical endpoints were analysed in those patients surviving for at least 3 months (n = 61). Return to alcohol abuse was documented in 16 patients at routine short-term out patient check-ups. All patients except one admitted to taking alcohol and showed changes in their laboratory test results. A specific pattern of liver function test values related to alcohol abuse was not detected and at the end of a relapse the liver function values usually returned to pre-event values. Only in one case was toxic injury of the liver related to alcoholic recidivism diagnosed on percutaneous liver needle biopsy or post-mortem examination. Compliance with the required immunosuppressive regimen and social rehabilitation after OLT were excellent. Unwillingness to offer OLT to individuals with alcoholic liver disease because of failure to demonstrate 100% long-term abstinence appears difficult to defend in the face of results showing good survival, compliance and social rehabilitation. The hypothesis of a higher sensitivity of the transplanted liver to a drinking episode and the redevelopment of alcoholic diesease in the new liver was not confirmed in our study population. PMID- 11271185 TI - The correlation of serum hyaluronan of liver donors with posttransplant liver function. AB - Primary organ non-function occurs in 2-17% of patients after liver transplantation (LTX). Liver function tests focus on hepatocytes, underestimating the importance of non-parenchymal cells such as endothelium. Liver endothelium metabolises hyaluronan (HA). The function of endothelium can be estimated by measuring the concentration of HA in serum. We evaluated two organ donor groups, the first with HA levels below 200 microg/l and the second with HA levels above 200 microg/l. Both groups degraded more than 60% of HA during the first 6 h after reperfusion. Determination of hyaluronic acid in patients after LTX gives valuable information about the functional state of liver endothelium. It does not necessarily parallel the release of transaminases. In this study, there was no correlation between the levels of HA in the donor and posttransplant liver function. High HA in organ donors are most likely a consequence of different degrees of trauma and a higher release of HA into the circulation rather than impaired endothelial function. PMID- 11271187 TI - Role of non-parenchymal liver cells in ischaemia-reperfusion liver injury: protective effects of muramyl dipeptide. AB - It has been suggested that non-parenchymal liver cells play a central role after ischaemia and reperfusion of the liver. Male Lewis rats were subjected to 90 min of warm liver ischaemia. Four groups were constituted: group 1, no treatment; group 2, muramyl dipeptide treatment, activation of Kupffer cells; group 3, dextran sulphate injection, Kupffer cell blockade; and group 4, gadolinium chloride administration, Kupffer cell blockade. Dextran sulphate (4 mg/100 g) and gadolinium chloride (GdCl2, 0.7 mg/100 g) were given intravenously on day 2. MDP was injected intravenously (500 mg/250 g) 24 h before and 10 min after the intervention. Mortality rates were assessed and serum transaminases, histology of the liver and Kupffer cell phagocytic activity were evaluated 6 h after the end of ischaemia. MDP treatment significantly (P < 0.001) reduced mortality (30%) in comparison with the non-treated group (60%). The mortality rate was significantly higher in the dextran sulphate-treated (80%) and gadolinium chloride-treated (90%) groups in comparison with group 1. A significant reduction in transaminase levels was observed after MDP treatment, while blockade of Kupffer cells resulted in higher serum transaminase levels. The extent of necrosis and congestion was improved by MDP administration, while disruption of the vascular and sinusoidal integrity of the liver and extensive areas of necrosis were observed in dextran sulphate and gadolinium chloride-treated rats. Sheep red blood cell 51Cr liver uptake was deeply depressed 6 h after the end of ischaemia in group 1 (10 +/- 1.2%/g tissue). MDP injection restored the Kupffer cell activity (30.6 +/- 3.22%/g tissue) while dextran sulphate and gadolinium chloride administration markedly decreased SRBC 51Cr liver uptake. Our findings demonstrate that MDP in able to protect the liver from ischaemic insult while blockade of Kupffer cells was deleterious in rats subjected to liver ischaemia. PMID- 11271186 TI - Changes in circulating levels of atrial natriuretic factor (ANF) during orthotopic liver transplantation in humans. AB - Atrial natriuretic factor (ANF) is a 28 amino acid peptide secreted by the atrial cardiocytes. Clearance is via the lung (50%) and the liver (25%). The main stimulus to ANF secretion is atrial distension but vasoconstrictors, sympathetic stimulation, catecolamines and tachycardia are able to enhance its circulating blood levels. ANF blood concentrations were measured during orthotopic liver transplantation in six postnecrotic cirrhotic patients. Significant increases in ANF blood levels occurred at the end of the anhepatic phase (P < or = 0.02 vs baseline) associated with low cardiac filling pressures (P < or = 0.02 vs baseline) and increased systemic vascular resistances (P < or = 0.02 vs preanhepatic phase). Aldosterone blood levels showed a similar behaviour, increasing significantly (P > or = 0.001 vs baseline) at the end of the anhepatic phase. ANF fell after reperfusion of the graft and returned towards baseline values at the end of the procedure. Since most of the total body clearance of ANF is performed by the lungs, its sharp increase at the end of the anhepatic phase could be considered a counterregulatory response to vasoconstricting stimulation and to fluid-sparing mechanisms in the presence of relative hypovolaemia. Its decrease after reperfusion could be related to volume normalization and partly to the enhanced clearance performed by the newly grafted liver. PMID- 11271188 TI - Intraoperative evaluation of big-endothelin plasma levels during liver transplantation in different vascular compartments. AB - Endothelin-1 (ET) is derived from its precursor big-ET, secreted by endothelial cells of multiple origin. The role of ET peptides in the physiological responses after orthotopic liver transplantation (OLT) was investigated. Venous big-ET plasma levels were analysed by RIA in 28 patients before and after OLT. Samples for analysis were taken intraoperatively from 12 patients from the caval, portal and hepatic veins and the radial artery at multiple time points. Highest caval levels were found during the anhepatic period and 60 min after reperfusion, followed by a drop and subsequent increase postoperatively. Highest levels in the hepatic and portal veins were detected during explanation and reperfusion. A different pattern was found in the radial artery. Values during rejection and infection were elevated compared with preoperative and postoperative levels. The heterogeneity of the kinetics points to different sites of ET generation, including liver and splanchnic circulation. It suggests a predominant paracrine secretion mode of ET peptides with various stimuli involved. Big-ET levels could reflect endothelial cell damage, as big-ET is generated intracellularly and biological activity is rather weak. PMID- 11271190 TI - Preservation of the recipient inferior vena cava in liver transplantation. AB - Twenty piggy-back (PB) liver transplantations (LT) were compared with 20 LT performed by the standard technique in order to evaluate whether or not the theoretical haemodynamic advantages of the preservation of the inferior vena cava (IVC) have any impact on the final results of the LT. Statistically significant differences were observed in the duration of the hepatectomy, which was longer for PB LT (192 min vs. 146 min), and in the duration of the anhepatic phase, which was shorter in that group (52 min vs. 76 min). There were no differences in the duration of the complete surgical procedure, consumption of blood products, incidence of postoperative acute renal failure, number of reoperations or survival. PMID- 11271189 TI - Pharmacokinetic interpretation of FK 506 levels in blood and in plasma during a European randomised study in primary liver transplant patients. The FK 506 European Study Group. AB - The efficacy and safety of FK 506 compared with cyclosporin were evaluated in a European multicentre study with primary liver transplant patients. The daily intravenous doses ranged from 0.15 to 15.9 mg and the daily oral doses from 0.5 to 30 mg. Trough concentrations of FK 506 in blood and plasma were determined by an enzyme immunoassay. Blood concentrations ranged from 0.5 to 391 ng/ml and from 0.5 to 616 ng/ml after intravenous and oral doses, respectively. The corresponding plasma levels ranged from 0.05 to 56 ng/ml and from 0.05 to 104 ng/ml, respectively. In comparison to the parallel US trial, the mean oral doses in this European study were about 20% lower and the mean blood concentrations were 40% lower. However, the efficacy in these two trials was similar. No significant relationship between blood levels and selected adverse events or serum creatinine concentrations were observed in the European study (6-month data). An analysis of plasma protein and albumin concentrations showed an increase to normal ranges 4-8 weeks post-transplantation. The level of both markers remained lower in patients who withdrew due to adverse events. PMID- 11271191 TI - Recipient hepatectomy with preservation of inferior vena cava reduces the need for veno-venous bypass in liver transplantation. AB - Recipient hepatectomy with inferior vena cava (IVC) preservation, the piggy back (PGB) technique, was adopted as our routine option in the management of the anhepatic phase of orthotopic liver transplantation (OLT) to avoid the use of veno-venous bypass (VV-BP). In the last 5 years, 119 OLT in adult patients have been performed in our unit. In the first period (47 OLT), VV-BP was used in 59% of the cases and cross-clamping in the rest. In the second period, following the introduction of the PGB technique, 72 OLT were performed. VV-BP was used in 5.5% of the cases, PGB technique in 87.5% and cross-clamping in 6.9%. There was a significant reduction in the need for VV-BP in the second period. Operating time and blood transfusion were significantly greater in the VV-BP group. No PGB technique related complications were observed. In conclusion, the PGB technique reduced the need for VVBP with consequent savings in time, blood transfusion and the cost of OLT. PMID- 11271192 TI - Effects of warm Carolina rinse on microvascular reperfusion injury in rat liver transplantation. AB - Recently, it has been demonstrated that the use of both cold Carolina rinse (CR, 4 degrees C) as well as warm Ringer's lactate (RL, 37 degrees C) attenuates microvascular perfusion failure and leukocyte (WBC) accumulation in liver grafts. The aim of this study was to analyse in vivo whether warming of CR can also lead to a reduction in microvascular reperfusion injury in rat liver transplantation. Syngeneic orthotopic liver transplantation, including arterial reconstruction, was performed in male Lewis rats (180-300 g). Livers were stored in University of Wisconsin (UW) solution for 24 h and rinsed with 15 ml CR which was either cold 4 degrees C (n = 7) or warm 37 degrees C (n = 8) prior to reperfusion. Hepatic microcirculation and WBC accumulation were assessed by intravital fluorescence microscopy, and graft function was determined by analysis of bile flow during the 90-min reperfusion period. Warm CR yielded significantly (P < 0.01) improved sinusoidal perfusion when compared with cold CR; however, the extent of WBC adherence in both sinusoids and postsinusoidal venules did not vary between the groups. In addition, bile flow was slightly increased after warm CR. We conclude that after 24 h of cold storage in UW solution, warming of CR may offer additional benefit in the prevention of microcirculatory reperfusion injury without affecting WBC accumulation. PMID- 11271193 TI - Hepatocyte isolation from pig livers after warm ischaemic injury. AB - Hepatocyte cultures have been used extensively for a wide variety of physiological, pharmacological and experimental studies. The warm ischaemic period before isolation is kept to a minimum to achieve a high yield of cells isolated and a good viability for culture. We have recently introduced a new concept of liver resuscitation after warm ischaemia that is based on a 3-h reperfusion period with an improved perfusate and simultaneous dialysis. In this study, we applied the new technique for hepatocyte isolation from livers subjected to 80 min of complete ischaemia at 37 degrees C. Cell yield was improved by a resuscitating perfusion from 58% to 73% and viability from 39% to 76%. PMID- 11271194 TI - MHC antigen presentation on the surface of hepatocytes: modulation during and after hypoxic stress. AB - Presentation and recognition of MHC antigen on the surface of cells is the basic process of initiating rejection and distinguishing "self and not self". This is considered to begin when a transplanted organ is reperfused with the blood of the recipient. In this study, the modulation of MHC I antigen presentation during preservation was investigated in a cell culture model using primary hepatocyte cultures of male Wistar rats. By incubation with different preservation solutions used in clinical transplantation, expression of the MHC antigen was observed during cold hypoxia. Primary hepatocyte cultures were isolated from male Wistar rats by a modified Seglen technique and seeded to glass slides, thus obtaining monolayer cultures. After 1 day of resting the cultures were incubated under different conditions using Krebs Henseleit solution (KH), Euro-Collins solution (EC), HTK solution of Bretschneider (HTK) and University of Wisconsin solution (UW) as incubation media. The conditions of incubation were warm normoxia (37 degrees C, pO2 100 mm Hg) and cold hypoxia (4 degrees C, pO2 < 0.1 mm Hg), which is the main condition of organ preservation. Incubation time was 6 h. Before starting the incubation reference cultures were fixed and every 60 min several cultures were withdrawn from the experiment and fixed also. MHC expression was studied by staining the cultures with monoclonal antibodies against rat MHC class I and class II. As expected, MHC class II was not present on the surface of hepatocytes, while MHC class I was demonstrated on the controls as well as on the cultures that were incubated using KH and EC independent of temperature and hypoxia or normoxia. At the end of the incubation they were still positive but not as strong as in the beginning. During incubation using HTK and UW, MHC I was not detectable at all phases of the experiment. In conclusion, hypoxic stress did not completely eradicate MHC I expression in rat hepatocytes, while the composition of the preservation medium may result in a hepatocyte surface negative for MHC I. Hydroxyethylstarch may be the substance in UW that covers the MHC antigen, while in HTK, mannitol or the histidine complex may play the same role. MHC negativity of the cells of a preserved transplant is another reason for the benefit of UW or HTK. PMID- 11271195 TI - SPC-100270, a protein kinase C inhibitor, reduced hypoxic injury due to reperfusion following orthotopic liver transplantation in the rat. AB - Recently, we reported that SPC-100270, a sphingosine derivative and inhibitor of protein kinase C (50-90 microM) in mixed micelle assays, reduced reperfusion injury resulting from hypoxia in a low-flow, reflow model of liver perfusion. Here we report that SPC-100270 has similar beneficial effects following liver transplantation in vivo. Rat liver transplantation was performed using nonarterial and rearterial techniques. Livers from syngenic rats were harvested surgically, prepared with vascular cuffs and a splint, and stored for 24 or 48 h in University of Wisconsin (UW) cold storage solution. Just prior to completion of vascular reconstruction, the organ was rinsed with 3 or 10 ml of Ringer's solution, vehicle, or a solution containing SPC-100270 (up to 500 microM). Following implantation surgery, low doses of SPC-100270 were ineffective at reducing both parenchymal and nonparenchymal cell death, yet significant (P < 0.05) reductions were observed with 500 microM. Further, nonparechnymal cell viability was improved nearly four fold by the drug. SPC-100270 (500 microM) tended to increase survival following 48 h cold storage in UW solution, but the improvement was not statistically significant. SPC-100270 also did not diminish carbon-centered free radical formation in transplanted livers from alcohol treated rats. Collectively, these data support the hypothesis that pretreatment of donor livers with an inhibitor of protein kinase C is effective in vivo at reducing reperfusion injury, particularly to nonparenchymal cells, following orthotopic liver transplantation in the rat. PMID- 11271196 TI - Experimental transplantation of hepatocytes in cases of toxic acute liver failure. An allograft model. AB - The aim of this experimental study was to modify a rat liver-cell harvesting technique and to evaluate the efficacy of allogeneic liver cell transplantation (Tx) using cyclosporin A immunosuppression in rats with N-dimethylonitrosamine (N DMNA)-induced acute liver failure (ALF). Twenty male Wistar rats, weighing 190 320 g, were used as donors. Hepatocytes were harvested by the use of a modification of the Seglen portal vein collagenase perfusion technique (type V/1.3 mg/ml), which resulted in the isolation of a mean of 8000 viable clusters of hepatocytes per donor (viability was measured using the trypan blue exclusion test). The male Lewis recipients and controls received 20 mg/kg N-DMNA i.v., and were then divided in three groups. Group 1 (n = 5) received no treatment, group 2 (n = 10) received 5000 clusters of freshly isolated hepatocytes (FIH) in the spleen 24 h after the administration of N-DMNA- and group 3 (n = 10) received 5000 clusters of FIH beneath the renal capsule, 24 h after the administration of N-DMNA. All groups were treated with cyclosporin A 20 mg/kg per day i.p.. SGOT and bilirubin values were measured and all surviving rats were sacrificed on day 14. All rats in group 1 died of histologically confirmed liver necrosis within 72 h. The 14-day survival was 60% in group 2 and 50% in group 3. The post-Tx SGOT values reached their maximum on days 3-4 (group 2, mean 754; group 3, mean 529) and were only slightly elevated on day 14 (group 2 = 75, group 3 = 48). The post Tx bilirubin values reached their maximum on days 3-5 (group 2 = 1.1, group 3 = 1) but failed to return to normal until day 14. Autopsy and histological examination of the surviving animals showed well-preserved hepatocellular spherical aggregates in the spleen and hepatocellular "cords" in the kidney accompanied by signs of regeneration of the native liver. We concluded that the hepatocyte Tx in a rat experimental allo-Tx model improved the survival rate and the SGOT values in cases of toxic ALF. Survival rates between the two different sites of Tx were similar. PMID- 11271197 TI - Effect of cold aerobic perfusion on nonparenchymal cell viability of rat livers. AB - We investigated the effect of oxygen supply on hepatic cellular viability during cold perfusion storage of rat livers. A perfluoro-N-methyldecahydroisoquinoline (FMIQ) emulsion is used as an oxygen carrier. The composition of the perfusate containing 20 w/v% FMIQ is essentially the same as the University of Wisconsin (UW) solution except for the exclusion of hydroxyethyl starch. Rat livers were perfused at 4 degrees C for up to 24 h with either UW solution (group I, oxygenated; group II, unoxygenated) or FMIQ solution (group III, oxygenated; group IV, unoxygenated). After perfusion storage, the livers were reperfused with warm (37 degrees C) oxygenated or cold (4 degrees C) unoxygenated Krebs-Henseleit bicarbonate buffer, and nuclear trypan blue uptake was measured as the index of cell death. With warm oxygenated reperfusion, there remained less than 2% noviable parenchymal cells up to 24 h, regardless of perfusate or oxygenation. In UW-perfused livers, the proportion of nonviable nonprenchymal cells (NPC) increased progressively regardless of oxygenation, the values in groups I and II in the periportal field at 24 h being 39.9 +/- 4.7% (mean +/- SD) and 36.5 +/- 4.2%, respectively. By contrast, in FMIQ-perfused livers, dye uptake by NPC was significantly reduced with oxygenation (16.9 +/- 5.7% and 39.4% +/- 9.1% at 24 h in groups III and IV; P < 0.001). With cold unoxygenated reperfusion, livers in groups I, II, and IV showed a significant decrease of nonviable NPC, while those in group III showed no significant changes. These data indicate that oxygen supply during perfusion storage of the liver may ameliorate lethal injury to NPC precipitated during reperfusion. PMID- 11271198 TI - The effect of a tumour necrosis factor antibody on the regenerative response after partial hepatectomy in rats. AB - In this study we investigated the effect of tumour necrosis factor (TNF) on the regenerative response after partial hepatectomy. Adult male rats were injected intravenously with an antibody to TNF immediately after partial hepatectomy. Animals were sacrificed at 0, 24, 48, 72 and 96 h postoperatively. Hepatic thymidine kinase (TK) activity, liver weight to body weight (LW/BW) ratio, and mitotic index (MI) were used as indices of hepatic regeneration. The rats treated with TNF-Ab had significantly lower levels of TK activity in the liver at 24 h postoperatively compared to the saline treated animals. Furthermore the peak hepatic TK activity was delayed to 48 h in the rats treated with TNF-Ab. The mitotic indices and LW/BW ratios in the TNF-Ab- and saline-treated animals were similar. These data suggest that TNF potentiates the regenerative response after partial hepatectomy. PMID- 11271199 TI - Kupffer cells participate in rejection following liver transplantation in the rat. AB - Recognition of foreign antigens involves macrophages which release mediators such as immunoactive interleukins, and in the liver, the resident macrophages (Kupffer cells) are activated following transplantation. Therefore, we evaluated the hypothesis that Kupffer cells participate in the rejection reaction following transplantation. Orthotopic liver transplantation was performed between different syngenic rat strains. Livers from Lewis rats were stored in lactated Ringer's solution for 1 h to minimize cold ischemic injury and transplanted into PVG recipients. At 24 h postoperatively, transaminases (AST) were elevated to values around 2000 U/l, total bilirubin was increased to values around 20 micromol/l, and five of six rats died within 3 days. Macroscopic and histological examination showed large areas of necrosis without cellular infiltration, characteristic of rejection. When donor rats were treated with gadolinium chloride (GdCl3, 10 mg/kg i.v. 24 h before storage of the liver) to inactivate the Kupffer cells, AST levels only rose to around 700 U/l, and the total bilirubin level was in the normal range (< 4 micromol/l). Survival was improved significantly by GdCl3, with five of seven rats surviving more than 1 month (P < 0.05) and four of seven rats surviving for at least 100 days without immunosuppressive drug therapy. Rejection was not totally prevented, however, since the surviving rats had elevated AST and bilirubin levels, and cellular infiltration in portal areas along with proliferation of bile canaliculi was observed. These data are consistent with the hypothesis that Kupffer cells participate in mechanisms of early rejection following liver transplantation. PMID- 11271200 TI - Effects of intrahepatic arterial and intraportal administration of FK 506 on liver allograft survival in rats. AB - Rejection is still the limiting factor for successful organ transplantation, and overdosage of immunosuppressive drugs often results in severe viral infection, side-effects and toxicity. Thus, more specific immunosuppression to lessen these side-effects is highly desirable. In this study, we compared the effects of FK 506 administered by different routes (hepatic artery, portal vein and systemic circulation) on the inhibition of rejection. FK 506 was given to recipient LEW rats with PVG liver grafts via the penile vein (systemic administration), portal vein or hepatic artery (local administration) for 3 or 7 successive days after liver transplantation. In control LEW rats without immunosuppression, the PVG liver allografts were rejected between 9 and 21 days after transplantation. Intravenous administration of FK 506 for 3 days (0.32 and 1.28 mg/kg daily) only had a marginal effect on prolonging liver allograft survival (21.1 +/- 12.5 and 32.0 +/- 24.0 days, respectively; control 14.1 +/- 4.1 days). However, systemic administration of FK 506 (0.08-1.28 mg/kg daily) for 7 days suppressed liver allograft rejection markedly (42.3 +/- 5.9 to 80.5 +/- 53.4 days; control 14.1 +/ 4.1 days), and 50% of the recipient rats survived for at least 60 days after liver transplantation. Moreover, when a low dose of FK 506 (0.32 mg/kg) was infused into the hepatic artery or portal vein of the transplanted liver for 3 days only, liver allograft survival times were prolonged markedly, and 54% of rats with grafts survived for at least 60 days. This effect was almost equal to that after 7 days systemic treatment with FK 506. In conclusion, 7 days' treatment with FK 506 administered systemically was an effective regimen for the suppression of liver allograft rejection in rats. Furthermore, local immunosuppression with low-dose, short-term (3 days) FK 506 treatment administered via the hepatic artery or portal vein of the transplanted liver dramatically improved allograft salvage. PMID- 11271201 TI - Therapeutic effect of 15-deoxyspergualin on acute graft rejection in canine liver transplantation. AB - Therapeutic effect of 15-deoxyspergualin (DSG) on acute rejection was investigated by examining hepatic functions and histological findings in a model of canine liver transplantation. When acute rejection (defined as an acute rise of hepatic functions) occurred, the recipients were treated with DSG alone or combined with a small amount of methylprednisolone (MP). The recipients in group 1 were administered no basic immunosuppressant, and those in groups 2 and 3 received cyclosporine as the basic drug. The rejections in groups 1 and 2 were treated with DSG combined with MP and those in group 3, with DSG alone. Amelioration or healing of hepatic dysfunction and histological abnormalities as seen in all groups except for one case in group 3. The observations in this study were quite similar to those in renal transplantation. Therefore, DSG therapy is expected to be useful for treating graft rejection even in clinical liver transplantation. PMID- 11271202 TI - Efficacy of prostacyclin analogue (OP-2507) in viable hepatic grafts from pigs with non-beating hearts. AB - We investigated whether the stable prostacyclin analogue (OP-2507; OP) would ameliorate warm ischemia-related injury of the liver graft under conditions of a nonbeating heart. Thirty-six mongrel pigs were arranged into 3 groups of 6 pairs. Group 1 pigs underwent orthotopic liver transplantation from heart-beating donors (HBD). In group 2, animals received liver grafts from nonheart-beating donors (NHBD), defined as 30 min of cardiac arrest. Group 3 pigs received grafts from NHBD, but the donor had been pretreated with OP by intraportal infusion (2 microg/kg x min for 30 min immediately before the induction of cardiac arrest). The grafts were preserved at 4 degrees C in Euro-Collins solution in which OP was dissolved at 200 microg/l. Five-day survival rates after transplantation improved significantly in OP-treated animals (3/6, for group 3), compared with 0/6 for group 2 (P < 0.05, generalized Wilcoxon test). Five of 6 animals survived more than 5 days in the HBD group (group 1). Although the serum transaminase activities and bile production did not differ in the early phase of recirculation among the groups, there was a significant improvement in the hepatic microcirculatory environment in the surviving groups (groups 1 and 3). Analysis of arterial prostanoid levels showed a substantial suppression of PGE2 release by OP treatment following reperfusion. Our data indicate that a stable prostacyclin analogue can be clinically useful for expanding the donor pool by improving the quality of the liver graft. PMID- 11271203 TI - Role of immunosuppression in recurrence after liver transplantation for diethylnitrosamine-induced tumors in rats. AB - Hepatocellular carcinoma is one of the world's most common malignant diseases, with an increasing incidence related to liver cirrhosis. The purpose of the study was to evaluate the role of immunosuppression in recurrence in rats transplanted after liver tumor induction by diethylnitrosamine (DENA), which has proved to be a reliable carcinogen. In 14-week-old Lewis rats weighing 200 g, tumors were induced by the oral administration (5 mg/100 ml in drinking water ad libitum) of DENA for 13 weeks. Orthotopic liver transplantation (OLT) was performed after 4 weeks' latency. In the Lewis/Lewis rats weighing 200 g, tumors sporin A (CsA) treatment, median survival was 199-days with no recurrence or metastasis. In the BN/Lewis group with no CsA (5 ats) median survival was 144 days. All rats died due to rejection. In the other BN/Lewis group (10 rats), OLT was followed by CsA administration (7.5 mg/kg). Median survival was 161 days. In three rats (218 days), there was liver tumor recurrence; in two rats (137.5 days), kidney and lung metastases were found. The remaining rats died of septic complications. In the Lewis/Lewis + CsA group (10 rats), median survival was 131 days with 5 recurrencies and/or metastases. Two rats are still surviving at 84 and 88 days. Our results suggest that the DENA model is reliable; it proved to have a similar carcinologic pattern to HCC in man. Moreover, immunosuppression seems to play an important role in determining recurrence. Further studies are needed to investigate the efficacy of chemotherapy agents pre- and post-transplantation. PMID- 11271204 TI - The validity of the MEGX test in correlation with histology after orthotopic rat liver transplantation. AB - Lidocaine metabolism (MEGX test) as an indicator for liver function in the assessment of different degrees of liver disease and as a predictor for liver outcome after transplantation is well established. Since reduced liver function is associated with an alteration in parenchymal and non-parenchymal cells, we evaluated whether MEGX values correlate with histology in an in vivo model of orthotopic rat liver transplantation (ORLT) to assess histological damage without taking biopsy specimens. Livers from syngeneic Lewis rats were transplanted with rearterialization after 15-30 h of cold storage in UW solution and rinsing with Carolina Rinse Solution prior to implantation. Forty-eight hours after transplantation, the MEGX test was performed and metabolites were measured with a commercial kit as described elsewhere. Biopsy specimens were taken and graded three degrees of damage (mild, moderate, and severe) in a double blind fashion by a pathologist. MEGX values were assigned to the histological results. Statistical analyses were done with a Mann-Whitney test (n = 58) for mean values. The mean MEGX values attributed to histologies with a mild, moderate, severe degree of damage were 159.96, 78.46 and 44.42 ng/ml, respectively. When the histological groups were compared with the mean MEGX values, mild vs moderate, mild vs severe and moderate vs severe were significant (P - 0.0001). In conclusion, MEGX values correlate significantly with histological grading in a linear fashion after ORLT. The MEGX test may be of clinical value because it reflects the histological pattern of livers and may reduce the necessity to take biopsy specimens before and after transplantation. PMID- 11271205 TI - Hepatitis C in liver transplant patients. AB - The aim of this study was to determine whether infection by the hepatitis C virus (HCV) recurs after orthotopic liver transplantation (OLT) and to define the natural history of post-transplantation chronic hepatitis due to HCV. Of 70 patients, 10 (14.3%) were found to have antibodies to HCV before transplantation. After OLT 14 of the 70 patients (20%) had positive anti-HCV antibodies: 8 of 10 positive pre-OLT (80%) and 6 of 60 negative pre-OLT (10%). Of 14 patients anti HCV+ post-OLT (57%), developed 8 chronic hepatitis: chronic persistent hepatitis in three patients, chronic lobular hepatitis in three patients and chronic, active hepatitis in two patients. We treated four patients with interferon obtaining normalization of transaminases in three of them after 6 months, but with a severe relapse in two. These results suggest that hepatitis C recurs in a majority, of liver transplant recipients and that morbidity is an important consideration. Interferon treatment of these patients requires further study to obtain conclusive results. PMID- 11271206 TI - Incidence and outcome of hepatitis C virus infection after liver transplantation. AB - The objective of this study was to determine the incidence and outcome of hepatitis C virus (HCV) infection after liver transplantation (OLT). Fifty-two transplanted patients were studied. Serum samples were examined for antibodies to HCV (anti-HCV) and HCV-RNA by PCR, before and after OLT. Patients were distributed into two groups: group 1 consisted of 24 patients (pretransplant anti HCV positive) and group 2 consisted of 28 patients (pretransplant anti-HCV negative). One year after OLT, HCV-infected patients were evaluated by liver biopsy. HCV-RNA was detected in 28 of the 52 (53.9%) patients after OLT. Twenty two patients in group 1 (96%) were reinfected. In group 2, acquired HCV infection was detected in six (21.4%) patients. At 6 and 12 months, one and five of six patients had seroconverted, respectively. Liver biopsy in 23 HCV-infected patients showed chronic hepatitis in 18 (78%) cases (2, chronic persistent hepatitis; 3, chronic lobular hepatitis and 13, chronic active hepatitis). Fourteen of the 23 (60.8%) patients were asymptomatic. Most symptomatic patients had chronic hepatitis with cholestasis. Overall, 18 of 20 cases of chronic hepatitis diagnosed in OLT recipients were HCV related. Mortality beyond 6 months after OLT was slightly higher in the HCV-infected group (P = 0.055). In conclusion, HCV reinfection is almost universal. Acquired HCV infection post-OLT is frequent. HCV-infected patients frequently develop chronic hepatitis. Most chronic hepatitis after transplantation are HCV related. PMID- 11271207 TI - Renal complications and development of hypertension in the European study of FK 506 and cyclosporin in primary liver transplant recipients. AB - We examined the occurrence of renal complications and hypertension in 540 primary liver recipients entered into the European liver trial comparing primary FK 506 to a cyclosporin A based immunosuppression regimen (CBIR). No difference in serious renal impairment or mean creatinine levels was observed with similar rates of "kidney failure" (FK 506 9.4% vs. CBIR 7.3%) and dialysis requirements (FK 506 12% vs. CBIR 11%). "Abnormal kidney function", a less serious parameter of renal impairment, was reported in 89 recipients (33%) in the FK 506 group versus 58 (21%) in the CBIR group (P < 0.01). Development of this complication was associated with elevated intravenous FK 506 dosing schedules, with the mean cumulative dose 43% higher than treated patients with unaffected kidney function. In a later cohort of patients where intravenous dosing was lower, no significant difference in renal complications was detectable. The 6-month prevalence rate of systemic arterial hypertension was noted to be lower in the FK 506-treated patients compared to the CBIR group [33 (17.2%) vs. 47 (25.7%)]. PMID- 11271208 TI - Incidence and clinical relevance of recurrent hepatitis C infection after orthotopic liver transplantation. AB - From September 1988 to November 1992 318 liver transplants were performed at our hospital. Of these patients 68 had end-stage cirrhosis due to non-A, non-B, hepatitis, 44 of whom (64.7%) had hepatitis C virus RNA in the serum. Of this subgroup 35 patients (79.5%) were also anti-HCV positive. Postoperatively most recipients remained anti-HCV positive and after 1 year more than 90% had HCV RNA in the serum. About 40% developed a mild, chronic hepatitis and 50% were carriers of HCV without histopathological signs. Two patients suffered from a temporary severe acute hepatitis and one patient had a fulminant liver failure due to reinfection. In general, in liver recipients transplanted for end-stage HCV hepatitis there was a high incidence of reinfection with HCV. The clinical course, however, was less severe than in hepatitis B recurrence. PMID- 11271209 TI - Recurrence of hepatitis C virus after liver transplantation. AB - The hepatitis C virus is a common cause of chronic hepatitis after orthotopic liver transplantation (OLT). We evaluated 95 consecutive patients who underwent OLT at our institute between March 1988 and November 1992 and who had a follow-up period longer than 3 months. All patients had a second-generation test (ELISA + RIBA) for HCV antibodies (HCV Ab) before and monthly after OLT; all had a polymerase chain reaction (PCR) test for detection of viral RNA after the operation. Whenever biochemical abnormalities (hypertransaminasemia 2 times the normal range) were seen, a percutaneous liver biopsy was performed. Forty-two HCV Ab+ patients before OLT remained positive after OLT. In this group the PCR test was positive in 32 cases (78.5%). In 13/42 (30.9%) cases (all PCR+) with hypertransaminasemia histological examination showed signs of viral C hepatitis (score of Knodell minimum 3, maximum 12, median 5.5). Of 53 HCV Ab patients before OLT, only 1 became HCV Ab+ and PCR+ 15 months after OLT. In the remaining 52 patients 15 were PCR+. Twenty of 53 patients (37.7%) had a liver biopsy because of hypertransaminasemia: in no case did histology show any signs of hepatitis C. In conclusion, viral C recurs often after OLT for post-hepatitic C cirrhosis. The histological graft lesions are in most cases moderate. We did not observe any deaths related to viral C infection in grafted patients. According to our results post-hepatic C cirrhosis remains a good indication for OLT. PMID- 11271210 TI - Results of liver transplantation in acute liver failure caused by viral hepatitis. AB - Fulminant liver failure due to acute viral hepatitis is the most common emergency indication for liver transplantation. The postoperative course is highly correlated with the type and duration of infection. The complication rate is lowest in fulminant hepatitis B patients and highest in subacute hepatitis C/NANB patients. PMID- 11271211 TI - Hepatitis C in liver transplantation: preliminary study of prognostic factors. AB - At the University of Miami liver transplantation for chronic liver disease in HCV positive patients has shown good results, with a 92% patients survival rate (follow up 8 to 57 months, median 21). None the less, we found that a large number of patients are expected to develop serious histological graft damage and may need retransplantation, which may place a further strain on the already scarce donor resources. We have conducted a preliminary investigation on the importance of parameters which may correlate with the prognosis of HCV grafts. We found no impact of HLA match or typing. An interesting hypothesis, which deserves further investigation, is that some HCV strains could be more virulent than others and play a role as an independent risk factor. We have identified six strains among our patients and the BK serotype shows a trend to be associated with a worse outcome. We have found that patients developing and maintaining higher liver enzyme levels (ALT and GGT) after transplant and those with higher levels of viremia may be at risk to develop serious damage to their grafts. PMID- 11271212 TI - Good and impaired response to ganciclovir treatment of severe CMV infections in liver transplant recipients. AB - In this study we investigated good and impaired clinical responses to ganciclovir treatment of severe CMV disease in 23 adult liver transplant patients. CMV episodes were diagnosed by direct immunodetection of CMV-specific antigens in blood leukocytes and by viral cultures. The patients were monitored weekly for CMV antigenemia during the antiviral treatment. Sixteen out of 23 patients recovered from CMV episodes with the standard ganciclovir therapy of 2 weeks. Seven patients demonstrated an impaired response to ganciclovir and had to be treated for longer than 2 weeks (29 +/- 9 days). The patients with an impaired response to ganciclovir also demonstrated higher CMV antigenemia levels compared to those with good a response, and all still had antigenemia after 2 weeks' therapy. Thus, most severe CMV infections in liver transplant patients subsided with ganciclovir treatment of 2 weeks, but impaired responses also occurred and patients had to be treated for several weeks with ganciclovir before they recovered from CMV. PMID- 11271213 TI - Factors associated with the development of post-transplant lymphoproliferative disease (PTLD) in Epstein-Barr virus (EBV)-seronegative adult liver transplant recipients. AB - Epstein-Barr virus (EBV) infection is recognized as the principal aetiological factor in the pathogenesis of post-transplant lymphoproliferative disease (PTLD), particularly when primary EBV infection occurs after transplantation. We analysed, using a time-dependent proportional hazards model, the factors associated with development of PTLD in 40 adult liver transplant recipients who were seronegative for EBV prior to transplantation. Of 40 patients, 13 (33%) had a tissue diagnosis of PTLD at a median time of 126 days after transplantation. The multivariate analysis showed that prior CMV disease, the number of steroid boluses given and the number of units of RBC and FFP transfused were significant risk factors for development of PTLD. PMID- 11271214 TI - Selection of small hepatocellular carcinoma improves long-term results of hepatic transplantation for malignancy. AB - Liver transplantation for advanced hepatocellular carcinoma is often followed by early tumour recurrence and death. At the beginning of the liver transplantation programme at Berlin Virchow we decided to offer liver transplantation only to patients with solitary tumours not exceeding a maximum diameter of 5 cm or to patients with two or three tumour nodes with a maximum diameter of 4 cm. From September 1988 to October 1993 435 liver transplants were performed in 403 patients. Of these, 32 patients (8%) had a histologically confirmed hepatocellular carcinoma (29 males, 3 females, median age 56 years). The overall actuarial survival according to Kaplan-Meier for the whole series of 32 patients with hepatocellular carcinoma was 82%, 78%, and 78% at 1, 2 and 3 years, respectively. Tumour size alone did not seem to be a relevant factor when comparing patients with tumours up to or larger than 3 cm in diameter. Patients with solitary tumours had a better prognosis than patients with multiple tumours. The largest difference was found between patients with stage I-III (UICC) tumours and those with stage IVA tumours: 1-, 2- and 3-year survival rates were 89% throughout in the former group, while the corresponding figures for patients with stage IVA tumours were 63%, 47% and 47%. Efforts should be made to identify stage IVA tumours preoperatively in order to use the precious resource of scarce donor livers in an optimal way. PMID- 11271215 TI - A prospective, randomized trial of pretransplant blood transfusions in cadaver kidney transplant candidates. Leuven Collaborative Group for Transplantation. AB - To assess the effect of pretransplant blood transfusions on the outcome of cadaveric kidney transplantation, a single-centre analysis was performed of 171 patients randomly assigned to receive no pretransplant transfusion (n = 85) or to receive at least three random blood transfusions (n = 86). After transfusion 18 of the latter patients developed circulating lymphocytotoxic T-cell antibodies, but the sensitization was only transient. At the time of transplantation, none was still sensitized. In both groups 60 patients have been transplanted. Patient and graft survival rates were significantly higher in the transfused group than in the non-transfused group. In the non-transfused patients the higher mortality was due to complications related to repeated anti-rejection therapy. Non transfused patients had more repeated acute rejection episodes than the transfused patients. The present study indicates that pretransplant blood transfusions still facilitate graft acceptance even in the setting of good HLA matching and with cyclosporine as the basic immunosuppressant. The risk of sensitization is very low. PMID- 11271216 TI - Two-year follow-up study of the efficacy and safety of FK 506 in kidney transplant patients. Japanese FK 506 Study Group. AB - We conducted a 2-year follow-up study of the efficacy and safety of FK 506 in 104 kidney transplant patients at 32 sites in Japan. The initial daily oral dose of FK 506 was 0.3 mg/kg, which was gradually reduced to 0.15 mg/kg by month 10 and remained stable thereafter. The mean trough level of FK 506 in whole blood and the mean serum creatinine level in year 2 were 7.9 ng/ml and 1.9 mg/dl, respectively. Patient and graft survival rates for all patients were 97% and 92%, respectively. Forty-six patients (44%) experienced rejection episodes, and 84% of these episodes occurred within 3 months after transplantation. The principal adverse reactions to FK 506 therapy were hyperglycaemia, renal dysfunction and hyperkalaemia. Most of these events were dose-dependent, and disappeared or ameliorated following reduction of the FK 506 dose. PMID- 11271217 TI - Pretreatment of kidney allografts with monoclonal antibodies to CD45: results of a multicentre study. CD45 Study Group. AB - The perfusion of human renal allografts with CD45-specific monoclonal antibodies (mAbs) may reduce their immunogenicity and the incidence of rejection. We performed a safety study in 40 patients receiving their first cadaveric renal transplant. Two milligrams each of the rat CD45 specific mAbs YTH 24.5 and YTH 54.12 were perfused into the grafts prior to transplantation. The patients were followed for 3 months. No patient died, and four grafts were lost, three to vascular causes not considered to be related to antibody perfusion and one to severe rejection. Two patients developed an anti-rat antibody response. Immunohistological double-labelling performed on cortical biopsies taken post perfusion and prior to wound closure showed that at least 60% of the CD45+ cells were coated by perfused anti-CD45 ('antibody uptake'). Among the patients studied for uptake and episodes of rejection, the incidence of rejection was 75% in 12 patients whose antibody uptake was < 95% compared with 22% in 18 patients with antibody uptake > or = 95% (P = 0.01). We conclude that this treatment was free of adverse effects and that there is a correlation between the uptake of antibody by passenger leucocytes and reduction in acute rejection episodes. PMID- 11271218 TI - Preventive OKT3 treatment with cyclosporine (Sandimmun) for second kidney transplantation. AB - A total of 793 kidney transplantations (KTx) were performed from November 1073 to March 1993. Two hundred and forty-two patients were treated with conventional immunosuppression (azathioprine + prednisolone) and all the others with cyclosporine (Sandimmun) and prednisolone (SIM + PRED). The survival of the second graft was less good in both therapeutic groups than that of the first ones, so we have started to use preventive immunotherapy with OKT3 (CILAG) in combination with SIM (both before operation) and PRED. We compared 32 SIM-PRED patients with 20 OKT3 + SIM + PRED patients. All underwent a second KTx. The two groups were found to be comparable and homogeneous with regard to 14 of 18 parameters analysed statistically. Statistically significant differences were found between the two groups as regards the frequency of acute rejection within 30 days (46.69% vs 20%), the delta creatinine value on the 1st and 2nd postoperative days (-4.3: -8 vs -8.6: -19.7%), patient survival after 4 years (78.2 vs 100%), and graft survival after 1 and 4 years (-58.9: -42.8 vs -83.5: 83.5%), with better results in the OKT3 group. We conclude that the preventive use of OKT3 simultaneously with SIM + PRED for the second KTx is the method of choice to prevent rejection and improve survival. This treatment results in patient and graft survival following the second KTx being as good as after the first KTx with SIM + PRED. PMID- 11271219 TI - Antilymphocyte globulins versus OKT3 as prophylactic treatment in highly sensitized renal transplant recipients. AB - Monoclonal antibodies were proposed as an effective prophylactic immunosuppressive treatment in highly sensitized patients (HSP). In this study we compared the results obtained in HSP treated with OKT3 or antilymphocyte globulins (ALG). From January 1989 to January 1993, 38 transplantations were performed in patients with high panel reactive antibodies (PRA > 50%). The group comprised 22 women and 16 men, mean age 45 +/- 2 (23-67) years; ten were second grafts and two were third grafts. Peak PRA was > or = 80% in 24 sensitized patients and 50-80% in 14 sensitized patients. Patients were randomly assigned to either prophylactic OKT3 (n = 15) or ALG (n = 23). Oral cyclosporin A (10 mg/kg) was started at day 8 in the OKT3 group and when the serum creatinine level decreased to 200 micromol/l in the ALG group. OKT3 was systematically withdrawn on day 10 but ALG was stopped only when total blood cyclosporin A concentration reached 150-200 ng/ml. In both groups, azathioprine (150 mg/day) and prednisolone were given. During the first months, 6/15 grafts were lost in the OKT3 group (three hyperacute rejections, one renal vein thrombosis, one steroid-resistant rejection, one death); in the ALG group 4/23 grafts were lost (one hyperacute rejection, two steroid-resistant rejections, one death). Side effects were significantly more frequent in the OKT3 group than in the ALG group. After 12 months of follow up, the graft survival was 71% (27/38) and did not significantly differ (log-rank test, NS) between the OKT3 (60%, 9/15) and the ALG group (78%, 18/23). We conclude that the use of the monoclonal antibody OKT3 as a prophylactic agent in HSP does not improve the early graft survival when compared with prophylactic ALG. Polyclonal antibodies, which react with many epitopes and are much better tolerated seem to offer a good strategy for induction therapy in this population. PMID- 11271220 TI - Safety and tolerability of a new oral formulation of cyclosporin A, Sandimmun Neoral, in renal transplant patients. AB - The current therapy with Sandimmun has been improved by the development of a new oral formulation of its active ingredient, Cyclosporine A and which is called Sandimmun Neoral. This new galenical formulation is based on the microemulsion technology and offers consistent oral absorption and pharmacokinetic predictability. In two studies of a 12 weeks duration each and including 466 stable renal transplant patients and 86 new renal transplant patients it was shown that Sandimmun Neoral is as well tolerated and as safe as Sandimmun. Stable renal transplant patients currently receiving Sandimmun can safely be switched to Sandimmun Neoral on a 1:1 dose level basis. However, as a result of the more consistent absorption of Sandimmun Neoral, poor absorbers with Sandimmun will become normal absorbers and than will need a considerable dose reduction to reach the same Cyclosporine A exposure. In new renal transplant patients kidney function seems to improve better and faster when Sandimmun Neoral is given as shown by creatinine and creatinine clearance values. In the Sandimmun Neoral group less patients experienced a rejection episode and time free of rejection was longer, this may reflect a better maintenance of immunosuppression. In addition, considerably lower doses (16% on average) are required for Sandimmun Neoral to achieve comparable cyclosporine A blood trough levels and these patients are also sooner stabilized in terms of Cyclosporine a therapy. PMID- 11271221 TI - Safety and steady-state pharmacokinetics of a new oral formulation of cyclosporin A in renal transplant patients. AB - The steady-state pharmacokinetics of a new oral formulation of cyclosporin A (Sandimmun Neoral, NOF, a microemulsion) was compared with those of the market formulation (Sandimmun, SIM) in stable renal transplant patients. Both formulations were administered as soft gelatin capsules every 12 h with doses adjusted to provide comparable trough concentrations (CminSS). Whole blood samples were obtained over a steady-state dosing interval (tau), and the cyclosporin A level was determined by a specific monoclonal RIA. Both formulations were well tolerated. The mean doses were 139 +/- 27 mg (SIM) vs. 120 +/- 19 mg (NOF), indicating a milligram dose-conversion factor of approximately 1:1 to yield comparable troughs. NOF exhibited a stronger correlation between AUCtauSS and CminSS (r2 = 0.821) compared with SIM (r2 = 0.288), due in part to less variability in the NOF profiles. Average increases of 39% in CmaxSS and 15% in AUCtauSS during treatment with NOF were not associated with any safety concerns over the 4-week exposure to Sandimmun Neoral, as evidenced by the absence of changes in blood pressure, hematologic and biochemical parameters (including serum creatinine and blood urea nitrogen, BUN) and ultrasound of the transplanted kidney. PMID- 11271222 TI - Neurological complications in the European multicentre study of FK 506 and cyclosporin in primary liver transplantation. AB - Neurological complications were examined in a multicentre, randomized, parallel group study of 545 patients undergoing primary liver transplantation to compare the efficacy and safety of FK 506- and cyclosporin A-based immunosuppressive regimens (CBIR). In an additional analysis, patients were divided into early and late randomized cohorts to detect the influence of protocol amendements that allowed for FK 506 dose reductions. Initial follow-up was for 6 months. Tremor, headache and insomnia were the most frequently reported adverse events involving the neurological system. Whereas these neurological symptoms were observed significantly more often in FK 506-treated patients (P < 0.05 vs. CsA for the overall population), this was no longer the case for the late FK 506 and CBIR cohorts. The risk of FK 506-treated patients developing tremor was related to the initial i.v. dose, the rate of administration of the i.v. dose and the daily dose (P < 0.01). Headache was significantly correlated with the FK 506 dose (P < 0.05), and insomnia was not related to any dosing variable. Major neurological symptoms, including psychosis, convulsion, coma, aphasia and intracranial haemorrhage, were reported with a low frequency (0.4-5.2%), and differences between both treatment groups were neither significant for the overall population nor for the early and late cohorts of FK 506 and CBIR. Data from the late cohorts showed no differences in the overall incidence of neurological adverse events between FK 506- and CBIR-treated patients. PMID- 11271223 TI - Prophylactic isradipine treatment after kidney transplantation: a prospective double-blind placebo-controlled randomized trial. AB - There is evidence that calcium antagonists may have a beneficial effect on cyclosporine-induced nephropathy after transplantation. We treated 50 consecutive non-diabetic patients receiving their first cadaveric transplant with isradipine, a dihydropyridine calcium antagonist, or placebo in a double-blind, randomized, placebo-controlled trial. There were no significant differences between the two groups as regards age, weight, sex, HLA matching and ischaemic periods. To achieve optimal vasodilation, treatment was started intravenously 2 h before the transplantation procedure, and continued orally afterwards for 3 months. The immunosuppressive treatment included rabbit antithymocyte globulin on day 0, and oral cyclosporine from day 5. In both groups 7 patients had primary non functioning grafts, but the incidence of never functioning kidneys due to vascular and thrombotic complications was significantly higher in the placebo group (0 vs 4 patients, P < 0.05). Hypertension was treated with oral labetolol in combination with guanfacine if necessary. In the placebo group antihypertensive medication had to be prescribed significantly more often (67% vs 33% of patients, P < 0.05), but resulted in similar blood pressure recordings in the two study groups. Cyclosporin A (CsA) plasma concentrations were also comparable but in the isradipine group a significantly higher dose of CsA was needed to achieve adequate levels (8.0 +/- 0.5 vs 6.2 +/- 0.5 mg/kg per day, P < 0.01). However, in the isradipine-treated patients creatinine clearance was significantly higher (66.1 +/- 4.5 vs 55.6 +/- 6.2 ml/min, P < 0.05) after 3 months. We conclude that isradipine is an effective antihypertensive agent after kidney transplantation. Isradipine ameliorates CsA-induced nephropathy and seems to protect against early postoperative vascular complications. PMID- 11271224 TI - Effects of PUVA therapy on kidney allografts: results of a randomized prospective double-blind study. AB - After successful experimental organ transplant studies on the efficacy of PUVA therapy combining donor pretreatment with the photosensitizer 8-methoxypsoralen (P) and the ex vivo irradiation of organs with long-wave ultraviolet light (UVA) prior to transplantation, we started in 1989 the first randomized, prospective, double-blind study to clarify the efficacy of PUVA therapy in human kidney transplantation. This study included 50 kidney donors, 25 of whom were PUVA treated. A total of 75 kidneys were transplanted in Berlin, Halle and Rostock. The complete data of these 75 recipients were available for the final evaluation. The PUVA group (n = 36) and the non-PUVA group (n = 39) were not statistically significantly different as to donor and recipient data. Regarding the results, no differences were seen in initial hospitalization time, early graft function, rejection rate, number and time of rejection episodes. After a follow-up of 24 months, both graft survival (PUVA vs. non-PUVA: 75% vs. 71.8%) and patient survival (97.2% vs. 97.4%, respectively) were comparably high. PUVA therapy did not influence the development of vascular rejection. Interestingly, the rate of late graft loss after the 6th posttransplant month was lower, but not statistically significantly so, in the PUVA than in the non-PUVA-group (2 vs. 6 graft losses). Thus, PUVA-pretreated kidneys may be associated with a reduced development of chronic rejection. PMID- 11271225 TI - Long-term graft survival rate of zero-mismatch kidney transplants for HLA-DRB1. AB - The study of a two-locus association between HLA-B and -DRB 1 revealed a significant 43 linkage disequilibrium. Donor-recipient HLA-DRB1 was determined by these 43 linkages. Zero-mismatch for HLA-DRB1 had a significant effect on the graft survival rate in living related and cadaver transplants. The 5-year graft survival rate was 94% in the zero-mismatch group for HLA-DRB1, 96% for related transplants, 92% for cadaver cases, and 94% in HLA identical siblings. A statistically significant difference was found between the zero-mismatch group for HLA-DRB1 and mismatch groups for HLA-DRB1 or HLA-DR (P < 0.01). The zero mismatch group for HLA-DRB1 had mismatches for HLA-A and/or HLA-B in 46 of 70 cases (66%). No significant differences in the rejection rate was observed between zero-mismatch and mismatch cases for HLA-A and/or -B in the zero-mismatch group for HLA-DRB1. In the second step, genotyping was conducted in 118 cases. The 5-year graft survival rate was 93% in the zero-mismatch group for HLA-DRB1 and 86% in mismatch group (not a significant difference). We concluded that zero mismatch transplant for HLA-DRB1 had a better long-term graft survival rate regardless of HLA class I. PMID- 11271226 TI - Factors responsible for delayed graft function and the impact of HLA-DR incompatibilities on rejection episodes in the early posttransplant period of renal allografts. HTK study group. AB - The analyses in this study demonstrated a significant effect of HLA-DR matching on the number of rejection treatments in the first 3 months after renal transplantation. PMID- 11271227 TI - Induction of donor-specific tolerance or sensitization as measured by sequential MLC reactivity up to 24 months after renal transplantation. PMID- 11271228 TI - Antibodies in alloimmunized uraemic patients treated with recombinant erythropoietin. AB - We have retrospectively analysed sera from 52 already sensitized uraemic patients collected over 1 year and compared erythropoietin (EPO)-treated with non-EPO treated patients. Significantly fewer (P<0.01) patients (33%) on dialysis because of the rejection of their kidney grafts received EPO than patients on dialysis because of underlying kidney disease (71%). EPO treatment reduced the number of additional blood transfusions, since 3/28 EPO-treated but 12/24 non-EPO-treated patients were given blood (P<0.05). Among the EPO-treated patients, 64% showed a loss of panel-reactive antibodies (PRA), as measured by the micro-lymphocytotoxic technique, while only 12.5% of the non-treated patients showed a loss of PRA (P<0.01). In the subgroup of transplanted patients, PRA loss was only found among the EPO-treated patients, but their number was small (P<0.05). The class, subclass and specificities of the antibodies, as determined by FACS (flow cytometry) analyses, showed no distinct differences between EPO- and non-EPO treated patients. The differences were significant between transfused and previously transplanted patients. PMID- 11271229 TI - Immunological risk factors are solely responsible for primary non-function of renal allografts. AB - Primary non-function (PNF) of renal allografts has been attributed to various risk factors, among them immunological ones, as well as unfavourable preservation conditions. To investigate the impact of these risk factory on the occurrence of PNF, 1335 consecutive kidney transplants performed at a single centre over a 10 year period were analysed. All patients received immunosuppression based on cyclosporine. As the method of analysis a conditional stepwise logistic regression model was chosen, comparing each graft suffering PNF with its partner kidney retrieved from the same donor. Thus, all donor-related variables could be omitted from the analysis, as they are the same in every pair of grafts. Risk factors analysed included panel-reactive antibodies, number of pretransplant transfusions, pregnancies, number of prior transplants, cold and second warm ischaemia time, mismatches on HLA loci A, B and DR and recipient age. The overall incidence of PNF was 87 grafts (6.5%). One patient suffered immediate rejection due to transplantation of an ABO incompatible graft. This case was excluded from further analysis. PNF occurred three times in recipients of living related grafts, twice in recipients of en-bloc grafts and four times in grafts, in which the paired kidney was either not transplanted or shipped outside the Eurotransplant region, so that no paired graft was available for matched case control analysis. Of the remaining 77 pairs, twice both organs of one donor failed immediately. The remaining 73 complete pairs were analysed. Two of the investigated risk factors have independently a significant impact on the occurrence of PNF. Increasing the number of pretransplant transfusions raises the relative risk of graft failure up to six fold (P=0.02), while a history of prior transplants bears a relative risk of 0.21E05 (P=0.005). Ischaemia has no significant impact on the occurrence of PNF. Our data strongly suggest that immunological rather than donor risk factors are responsible for the non-function of kidney grafts. PMID- 11271230 TI - Improved outcome of renal transplantation in amyloidosis. AB - We report our results in 96 patients with amyloidosis who received 105 cadaveric renal allografts. The graft survival of amyloidosis patients has improved with time and with improved immunosuppression. The graft survival of amyloidosis patients is comparable to the results in another systemic disease, i. e., diabetes, and only slightly inferior to those in primary renal disease, even though amyloidosis patients tolerate complications poorly and the patients are at high risk of dying during the first 3 months. PMID- 11271232 TI - Embolization of non-tolerated non-functioning kidney graft: alternative to surgical removal. AB - The results of treatment by percutaneous transcatheter embolization in eight cases of non-tolerated non-functioning kidney graft are presented. The symptoms resulting from non-tolerance of the renal graft were fever, pain and haematuria. Embolization was well tolerated in all eight cases and the only adverse effect was post-embolization self-limited fever in five cases. The symptoms of non tolerance of the graft disappeared immediately in all cases, with minimal morbidity and no mortality. In only one patient was it necessary to perform second embolization procedure to achieve permanent control of symptoms. We conclude that percutaneous embolization of non-tolerated non-functioning kidney graft is an effective procedure with significantly less morbidity than with surgical graft nephrectomy. PMID- 11271231 TI - Focus on intractable rejection: 6-month results of the European multicentre liver study of FK 506 and cyclosporin A. AB - The incidence of intractable rejection was evaluated during the course of a multicentre, randomised, parallel-group study comparing the efficacy and safety of FK 506 and conventional cyclosporin A-based immunosuppressive regimens in patients undergoing primary liver transplantation. A diagnosis of intractable rejection was made if there was histological evidence of unchanged or worsening acute rejection, or chronic rejection after two discrete courses of antirejection therapy. Antirejection regimens were specific to each centre. Patients who experienced intractable rejection could be withdrawn from the study. Patients who were withdrawn from the cyclosporin A treatment group could subsequently receive FK 506 therapy and vice-versa. Intractable rejection was diagnosed in 32/540 patients (5.9%): 7/267 patients (2.6%) in the FK 506 treatment group and 25/273 patients (9.2%) receiving cyclosporin A therapy (P < 0.01). Of these 32 patients, 25 were withdrawn from the study: 3 and 22, from the FK 506 and cyclosporin A treatment groups, respectively. All three patients withdrawn from the FK 506 treatment group are alive: two having undergone retransplantation. Of the 22 patients withdrawn from the cyclosporin A group and converted to FK 506 therapy, 6 were retransplanted, 4 of whom subsequently died. A further two patients died without retransplantation. Thus, in 14 of the 16 patients who were still alive at 6 months, the liver graft was saved after conversion to FK 506 treatment. The reduced incidence of intractable rejection in patients receiving treatment with FK 506, together with the successful rescue of patients developing intractable rejection while receiving cyclosporin A, suggests that FK 506 is an effective immunosuppressive agent following orthotopic liver transplantation. PMID- 11271233 TI - The influence of donor age on initial and long-term renal allograft outcome. Leuven Collaborative Group for Transplantation. AB - To investigate the impact of donor age on the immediate and long-term graft outcome, 808 primary cadaveric renal allograft recipients, transplanted between January 1983 and December 1992, were divided into six groups according to donor age: 10-19 years (n = 142), 20-29 years (n = 214), 30-39 years (n = 136), 40-49 years (n = 146), 50-59 years (n = 142), 60-69 years (n = 28). The six groups were comparable with regard to donor origin (local/distant), serum creatinine, cold ischemia and reanastomosis time, recipient sex, degree of presensitization, number of pretransplant blood transfusions, number of HLA-B and B/DR mismatches. The incidence of delayed graft function was linearly correlated with increasing donor age, from 11.9% (donors 10-19 years) to 39.3% (donors 60-69 years) (P<0.0001). Graft survival at 3 years was not influenced by donor age (from 89.3% for the youngest donors to 84.4% for donors 60-69 years). After the 3rd decade, the creatinine clearance linearly decreased with donor age (6.2 ml/min, P < 0.01). This progressive decline could not be attributed to the recipient age (-7 ml/decade for 485 recipients < 50 years, and -6.1 ml/decade for 323 patients > or = 50 years). Despite the decreased function in older kidneys, recipient renal function remained remarkably stable between 1 and 3 years after transplantation within each donor age group. PMID- 11271234 TI - Alloantigen-independent factors lead to signs of chronic rejection in long-term kidney isografts. AB - Chronic rejection is the most important cause of graft failure after the first year of transplantation. In addition to the effects of host immunological injury, antigen-independent factors may gain in importance over the long term. We therefore assessed the influence of such factors on rat kidney isografts functioning for prolonged periods and compared our findings with those observed in naive control rats and in allograft recipients. Functional, morphological, and immunohistological changes in the isografts became obvious 32 weeks after engraftment, and by 52 weeks the findings mimicked those of chronic allograft rejection. As allografted kidneys sustained these changes earlier on and more intensely, both alloantigen-independent and -dependent factors are thought to be implicated in this process. PMID- 11271235 TI - Expression of ICAM-1 and HLA class II in acute cellular and vascular rejection of human kidney allografts. AB - We studied the relationship between ICAM-1 and class II expression on graft tubular cells and the relationship with graft inflammation in 50 kidney transplants monitored with serial aspiration biopsies after transplantation. Of the 50 grafts, 26 had an acute rejection 17 +/- 10 days after transplantation, 5 also had acute vascular rejection (AVR) and 24 had no rejection. The initial post transplant ICAM-1 and class II expression was low in all grafts. All 21 grafts with acute cellular rejection (ACR) displayed ICAM-1 induction, with a peak at the beginning of acute blastogenic rejection and declining over 20 days to prerejection levels. Class II expression reached a peak later and also declined later to prerejection levels. In the grafts with irreversible AVR both ICAM-1 and class II expression remained elevated. The 24 grafts with no rejection displayed no ICAM-1 or class II induction on tubular cells during the follow-up. The differences between ICAM-1 and classs II expression in biopsies with rejection and with no rejection were statistically significant. The results demonstrate that ICAM-1 was induced early during ACR on the graft tubular cells and that it disappeared rapidly in reversible rejections. The induction of class II antigens was slightly slower but quantitatively greater. In the irreversible rejections with a combination of ACR and AVR both ICAM-1 and class II expression remained elevated. PMID- 11271236 TI - Human chronic kidney allograft rejection is accompanied by increased intraglomerular cathepsin B and L activity. AB - The major reason for late graft losses is chronic rejection. Recently, a large number of studies have indicated that proteolytic enzymes play an important role as mediators of glomerular injury. The cysteine proteinases cathepsins B and L degrade structural matrix proteins such as type I collagen and laminin. We investigated intraglomerular protease activities in 12 patients after kidney graftectomy because of end-stage renal disease following chronic rejection. A group of 12 patients undergoing nephrectomy because of cancer served as controls using only non-involved parts of the kidney. The activities of cathepsins B and L in homogenates of isolated glomeruli were measured fluorometrically methylcoumarylamide substrates and related to DNA content. In rejected kidney allografts we observed significantly enhanced intraglomerular cathepsin B activity and cathepsin B + L activity. PMID- 11271237 TI - Living kidney transplantation between spouses: results in 100 cases. AB - The use of unrelated living donors in kidney transplantation is still controversial but many transplant centres have accepted this procedure. The main argument against this approach is usually an ethical one. Because of this, at our institution we accept biologically unrelated donors only if they have an emotional closeness to the recipient. From January 1983 to October 1993, out of 654 kidney transplantations we performed at our institution, 364 kidney allografts were from living donors. Of these living donors, 245 were first-degree relatives of the recipient (LRD) while 119 were unrelated (LURD); 100 cases were spouses--wife to husband in 76 cases and husband to wife in 24 cases. Statistical analysis of the results (chi-square) revealed actuarial patient and graft survival rates of 89.8% and 86.8% at 1 year, 82.9% and 72.3% at 5 years and 72.3% and 60.3% at 9 years, respectively. In our series, the result of living donor kidney transplantation in this group were similar to those obtained in the LRD group, while they were significantly better than those from cadaver donors (P = 0.003). In conclusion, cadaver organs given the shortage of kidney transplantation between spouses may be a good alternative and can be performed successfully, providing a "gift of life" for both the patient and the family. PMID- 11271238 TI - Relationship between renal histology and later graft outcome. AB - We have created the chronic allograft damage index (CADI), which quantifies the early histopathological changes in renal allografts. In this study we showed that the CADI at 2 years after renal transplantation predicted the graft outcome 4 years later and that the CADI identified the risk group that proceeded to chronic rejection during subsequent years. PMID- 11271239 TI - Patient and graft survival in the European Multicentre Liver Study--FK 506 vs cyclosporin A. AB - A prospective randomised study was conducted to evaluate the efficacy and safety of FK 506 administered with corticosteroids compared with a cyclosporin A-based immunosuppressive regimen in patients undergoing primary liver transplantation. 545 patients were recruited in eight European centres, of whom 267 were randomised to FK 506 therapy and 273 to cyclosporin A-based therapy. The estimated Kaplan-Meier patient and graft survival figures of 82.9% and 77.5% respectively in the FK 506 group were higher than the comparable figures in the cyclosporin A group (77.5% and 72.6%, respectively). These differences did not reach statistical significance. Retransplantation rates, time to first rejection episode and number of rejection episodes were all lower (P < 0.001) in the FK 506 group. The infection rates were comparable between the two groups. During the study, the dose of FK 506 was reduced; this did not compromise efficacy and reduced the associated toxicity. FK 506 provides effective immunosuppression in patients undergoing primary liver transplantation and is associated with a lower incidence of rejection. PMID- 11271240 TI - Cyclosporin, nifedipine and gingival hyperplasia: a randomized controlled study. AB - Nifedipine increases the frequency and severity of gingival hyperplasia associated with CyA therapy in renal transplant recipients and this effect appears to be independent of whole-blood CyA levels. De novo malignancies have been reported arising in areas of gingival hyperplasia, in a group already at high risk of malignancy. Patients receiving CyA and nifedipine should receive advice regarding the need for strict oral hygiene to control the initial development of gingival hyperplasia, with severe cases being promptly referred for gingivectomy and histological examination. PMID- 11271241 TI - CD4 and CD8 monoclonal antibody therapy in canine renal allografts. AB - Therapy with CD4 and CD8 monoclonal antibodies was evaluated in dogs which received double-haplotype MHC-mismatched renal allografts. Neither CD4 nor CD8 monoclonal antibodies given alone prolonged allografts survival (creatinine > or = 300 micromol/l) beyond 7 days. However, combined therapy with CD4 and CD8 antibodies given up to day 10 did prolong allograft survival to a median of 14 days. A longer (21 day) course of CD4 and CD8 antibodies did not extend allograft survival further. The effect of prolonged antibody therapy was restricted by the occurrence of both an antiglobulin response and an anaphylactoid reaction to the monoclonal antibody preparation. When the CD4 and CD8 antibodies were combined with a pan-T-cell-depleting Thy-1 antibody, the survival of double-haplotype mismatched allografts was further prolonged (median 16 days). The median survival of single-haplotype mismatched renal allografts on this triple therapy was 21 days, with one surviving to day 36. PMID- 11271242 TI - Host leukocytes and their products in chronic kidney allograft rejection in rats. AB - Bilaterally nephrectomized Lewis recipients of Fisher 344 (F344) kidney allografts, treated with CyA (1.5 mg/kg/day x 10), develop progressive changes of chronic rejection. Treated F344-to-F344 acted as isograft controls. Proteinuria was determined sequentially. Grafts were harvested 8, 12 and 16 weeks after transplantation (n = 9/group/time period). Infiltrating host cells and their products were assessed in chronically rejecting grafts by histology and immunostaining using mAbs for monocyte/macrophages, T-cells, ICAM-1, LFA-1 and cytokines. For in vitro binding studies, snap-frozen sections of transplanted kidneys were incubated with monocytes/macrophages and lymphocytes isolated from peripheral blood (PBL) of naive animals. For in vivo migration studies, naive cell populations were labeled with Bis-Benzamide and transferred i.v. to grafted animals at weeks 8, 12 and 16 (n = 3/group); grafts were harvested 24 h later and cell localization assessed under immunofluorescence. Increasing numbers of ED1 + monocytes/macrophages in allografted kidneys peaked at 16 weeks, localizing preferentially in glomeruli, where IL-1, IL-6 and TNF-alpha expression had also become intense and correlated with progressive glomerulosclerosis. Binding studies corroborated these results. In vitro, a few monocytes/macrophages bound to glomeruli and vessels at 8 weeks; by 12 weeks, binding to glomeruli was high (72% of cells). In vivo, large numbers of transferred labelled monocytes/macrophages were found in kidney allografts at 12 weeks (23%, isografts; < 7%, P < 0.01). In contrast T cells (primarily CD4+) were a consistent feature in allografts elevated as compared to isografts and correlating with in vitro and in vivo binding patterns; associated cytokines included IL-2, IFN- and TNF-alpha. Functional data followed these results: urine protein excretion by allograft recipients increased from baseline at 8 weeks (12 mg/day) to > 50 mg/day at 16 weeks at which point animals were beginning to die of renal failure; proteinuria in isografted rats did not increase during this time period. These results suggest that monocyte/macrophage and CD4+ T cells and their products are important in chronic kidney allograft rejection, contributing to the progressive sclerosis and fibrosis. PMID- 11271243 TI - The relationship between reduced functioning kidney mass and chronic rejection in rats. AB - Interest has recently increased in the role of alloantigen-independent factors in chronic rejection. In this context, we examined the long-term effects of reduced functioning kidney mass in a F344-->LEW allograft (A) model. Animals were divided into three groups depending upon the amount of retained kidney. Renal arterial branches in the hilus were ligated so that one-third or two-thirds of the graft remained viable (1/3 and 2/3 groups, respectively); organs were left intact in the third (3/3) group. Urine protein concentrations were determined 4, 6, 8 and 10 weeks after engraftment and organs (five/group/time) were harvested and examined morphologically and immunohistologically. Proteinuria increased progressively in all 1/3, 2/3 and 3/3A animals, but faster in those with reduced kidney mass. This functional decline correlated well with increasing numbers of macrophages followed by interstitial fibrosis and glomerular sclerosis, which had become prominent by week 6 in group 1/3A and by 8 weeks in groups 2/3A and 1/3I (I, isografted), with animals beginning to die. IL-1, IL-6 and TNF production correlated well with the location and number of macrophages in all groups. These results suggest that kidney mass exerts a significant alloantigen-independent influence on chronic rejection. Allogenicity of the graft accelerates and amplifies the process. PMID- 11271244 TI - Cytomegalovirus (CMV) prophylaxis by acyclovir in pre-transplant CMV-positive renal transplant recipients. AB - Cytomegalovirus (CMV) infections, either primoinfection or reactivation, remain an important problem in organ transplantation. We therefore designed a prospective study in which pre-transplant CMV-positive renal transplant (RT) patients were randomized to receive for 3 months starting immediately after transplantation either acyclovir or nothing. Between April 1992 and January 1993, 53 cadaveric renal transplantations were performed in our institution. The immunosuppressive regimen included anti-thymoglobulins (ATG), azathioprine, steroids and cyclosporine A. Patients randomized in the acyclovir arm received the drug from day 1 to day 90 (D90) intravenously as long as the creatinine clearance was not above 10 ml/min and per os afterwards (3200 mg/day if the creatinine clearance was above 50 ml/min). CMV viraemia tests were systematically performed every 2 weeks until day 90 or when febrile episodes occurred. The patients were 53 adults who received a RT during the study period; 37 were included in the study of which 19 received acyclovir prophylaxis (group A) and 18, no prophylaxis (group B). The two groups did not significantly differ according to sex ratio, recipient's age, number of CMV-negative donors and number of days on ATG (10.76+/-6.16 vs. 8.28+/-4.21 days). There were significantly fewer viraemia episodes in group A (n = 6) than in group B (n = 13, P < 0.05); nevertheless, the percentage of symptomatic CMV viraemia was the same in both groups (35% vs. 38.5%). The onset of CMV viraemia occurred in the same period in both groups (39+/-13.8 days vs. 34.3+/-15 days; P = NS). The number of rejection episodes in the study period was the same in both groups (8 in each). We conclude from this prospective study that post-RT acyclovir prophylaxis reduces significantly the number of CMV viraemia episodes but does not delay their onset. Furthermore, it has no effect upon the percentage of symptomatic viraemias. PMID- 11271245 TI - Positive donor and negative recipient cytomegalovirus status is a detrimental factor for long-term renal allograft survival. AB - In 524 allogeneic cadaveric kidney transplants, the impact of cytomegalovirus (CMV) donor/recipient status on the incidence of CMV infection, CMV disease, early and long-term graft, and patient survival have been analyzed with respect to rejection episodes. Most CMV infections (59%) and diseases (17%) were found in CMV-negative recipients of CMV-positive kidneys. The 1-year function rate of CMV positive kidneys (75%) dropped about 10% below that of CMV-negative organs (85%), and in the case of CMV-negative recipients an additional graft loss of more than 10% happened within the 4th and 5th years (5-year graft survival pos./neg.: 56%). This detrimental effect was exaggerated if it coincided with antibody-treated rejection episodes. PMID- 11271246 TI - Prospective study of pulmonary function for cyclomegalovirus infection after renal transplantation. AB - Pulmonary involvement remains the main complication of cytomegalovirus (CMV) infection in renal transplantation (RT). Early diagnosis is required for the best management of patients, and preemptive therapy could be a successful approach. In order to define the predictive value of lung alveolocapillary abnormalities. We prospectively studied 26 renal transplant recipients for their diffusing capacity of carbon monoxide (DLCO) and their pulmonary clearance of a 99mTc-DTPA aerosol. Patients were studied before transplantation and then every 2 weeks up to the end of the 3rd month following RT. Viral blood cultures and serological determinations were performed every week during these first 3 months. Bronchoalveolar lavage (BAL) was done in case of CMV disease. Statistic analysis was done using Student's t-test and Pearson's chi-square test. During the 3-month follow-up, 13 patients remained free of CMV infection. Three non-infected and 8 infected patients showed DTPA clearance abnormalities (P < 0.05). Six non infected and 7 infected patients showed DLCO disturbance (P > 0.50). DTPA clearance were significantly modified on days 45 and 60 in the infected group. Serial DTPA scanning, in association with viral blood cultures, could be a useful test to avoid unnecessary BAL in a preemptive therapy strategy. PMID- 11271247 TI - Treatment of chronic hepatitis B and C with interferon-alpha in renal allograft recipients: preliminary results. AB - We evaluated the effects of treatment with interferon (IFN) on liver disease and renal allograft function in ten immunosuppressed cadaver kidney recipients. Two females and eight males (mean age 39 years) with biopsy-proven chronic active hepatitis (n = 8) or persistent hepatitis (n = 2) and serum positive for hepatitis B surface antigen (HBsAg) and HBe antigen (n = 5) or serum positive for anti-HCV antibodies (n = 3) or serum positive for HBsAg, anti-HCV and anti-HDV antibodies (n = 2) received 3 million units IFN thrice weekly of 6 months. All patients responded with a reduction in serum aminotransferase activity and in five of them liver function completely normalized. Three patients among five infected with HBV cleared HBeAg. During the follow-up period liver function remained stable in 9 patients after discontinuation of IFN therapy. Three patients lost their grafts due to rejection 1, 2, and 4 months after IFN therapy, respectively. In six patients renal function remained stable during and after IFN therapy. We conclude that in selected groups of renal allograft recipients IFN can be used safely and effectively for the treatment of chronic viral hepatitis. PMID- 11271248 TI - The impact of hepatitis C virus infection on liver disease in renal transplant recipients. AB - To assess the prevalence of hepatitis C virus (HCV) infection in renal transplant recipients and its impact on posttransplant liver disease, the sera from 176 recipients who had been followed for 1-20 years (mean 8.3 years) were tested for HCV-specific antibody using enzyme immunoassay. HCV-specific antibody was detected in 53 patients (30.1%) including 2 patients also positive for hepatitis B surface antigen (HBsAg). Among 167 HBsAg-negative patients, the presence of HCV specific antibody was associated with an increased incidence of chemically significant hepatitis (70.6% vs. 9.5% in anti-HCV-negative patients, P < 0.01). Hepatitis was more likely to be chronic in anti-HCV-positive patients than in anti-HCV-negative patients (P<0.05). Serious liver disease developed in 4 of 51 anti-HCV-positive, HBsAg-negative patients: liver failure causing death in 3 and hepatoma in 1. Liver biopsy specimens from anti-HCV-positive patients showed more aggressive histological lesions compared with those from anti-HCV-negative patients. We conclude that HCV infection is quite prevalent in our renal transplant recipients and plays a major role in posttransplant chronic liver disease. PMID- 11271249 TI - Cancer incidence in a kidney-transplanted population. AB - The cancer incidence in all Finnish kidney-transplant recipients up to 1991 was studied. In 2090 patients 94 cancers were diagnosed, with a calculated incidence of 14.2% at 15 years' follow-up. The standardised incidence rate (SIR) compared with the entire Finnish population was 2.7, and it remained stable throughout the follow-up period. The SIR for skin cancer was 20, for thyroid cancer 11, and for kidney cancer, non Hodgkin lymphomas, cancer of the colon, bladder and female genital organs, 7, 6, 5, 4 and 3 respectively. PMID- 11271250 TI - Non-Hodgkin's lymphoma after kidney or heart transplantation: frequency of occurrence during the first posttransplant year. AB - The incidence of non-Hodgkin's lymphoma was analysed in over 70,000 kidney transplant recipients and over 10,000 heart, heart-lung or lung transplant recipients. An increased incidence of lymphomas during the first posttransplant year was observed in cadaver kidney recipients as compared to related kidney recipients, in thoracic organ recipients as compared to kidney recipients, in heart-lung recipients as compared to heart or lung recipients, in patients transplanted in North America as compared to patients transplanted in Europe, in patients receiving cyclosporine in combination with azathioprine as compared to patients with other immunosuppressive regimens, and in patients receiving ATG/ALG or monoclonal OKT3 for rejection prophylaxis. PMID- 11271251 TI - The value of flow cytometric crossmatch in cardiac transplantation. AB - One of the major clinical problems in cardiac transplantation is that of moderate rejection of the graft, and over the past few years there is increasing evidence that humoral antibody may be important in graft prognosis. The sensitivity of the conventional cytotoxic crossmatch has been questioned, and an increased significance of there of the flow cytometric crossmatch (FCXM) to detect the presence of antibodies before transplantation has been reported. In this study we have examined the sera of 138 cardiac transplants (1988-1992) for the presence of donor-directed IgG and IgM antibodies using FCXM. Sera were collected immediately before transplantation and before the institution of immunosuppressive therapy. All pretransplant cytotoxic crossmatches were negative. After a minimum follow-up period of 3 months, the performance of the transplants was graded by endomyocardial biopsy: 1, no or mild evidence of rejection; 2, patients showing moderate rejection requiring increased immunosuppression. Of the 138 patients studied, 10 patients were excluded as they died within the first week of transplantation. Eight children were excluded since they were given prophylactic ATG (Merieux). A positive FCXM result was defined as showing values in excess of that found for the AB control sera. A significant association was found between the presence of both IgG to T and B cells and IgM to T cells and graft performance (P = 0.02 and 0.93, respectively). Indeed, IgM-directed T-cell antibodies were only found in patients with moderate rejecton. These two groups were mutually exclusive, so that the FCXM was able to identify the presence of moderate rejection in 55% of the patients. In conclusion, results show that pretransplant FCXM in cardiac transplantation provides a more sensitive assay of antibody status in recipients and has proved to be of prognostic value. PMID- 11271252 TI - Vascular cell adhesion molecule-1 (VCAM-1) is induced during cytomegalovirus infection in vascular structures of heart allografts. AB - This study was designed to investigate the expression of vascular cell adhesion molecule-1 (VCAM-1) and the counter-ligand VLA-4 (CD 49d) in frozen sections of endomyocardial biopsies (EMBs) of heart allografts in relation to the onset of cytomegalovirus (CMV) infection diagnosed by CMV antigenemia. Altogether, 105 EMBs were obtained from 21 heart transplant recipients. Serial EMBs from nine patients with CMV infection, from five patients with rejection, and from seven patients with a noncomplicated postoperative course were analyzed. Associated with CMV infection, capillary expression of VCAM-1 was significantly induced when compared to control biopsies (P < 0.0001). A striking difference in the expression of VCAM-1 during rejection and CMV infection was observed: in most rejecting biopsies only a few capillaries stained faintly for VCAM-1, whereas during CMV infection, multifocal intense staining was found (P < 0.0001). PMID- 11271253 TI - Enhancement of transplantation-associated atherosclerosis by CMV, which can be prevented by antiviral therapy in the form of HPMPC. AB - Effects of acute cytomegalovirus (CMV) infections on transplantation-associated atherosclerosis (TxAA) were studied in a rat model for chronic vascular rejection. Rats underwent orthotopic abdominal allogeneic aorta transplantation. Neo-intima formation and media thinning was quantitated by measurement of cross sectional surface areas (CSA) at day 50 post transplantation (Tx). Acute rat cytomegalovirus (RCMV) infection, established at the moment of maximum intimal proliferation and influx of inflammatory cells in the adventitia, resulted in enhanced neo-intima formation, accompanied by increased influx and proliferation of smooth muscle cells (smc) in the intima. This effect was completely inhibited by HPMPC, a very potent and selective inhibitor of CMV replication, indicating the virus specificity of the measured effects. Despite increased neointima formation, media thinning ("necrosis") was not affected by RCMV infection. PMID- 11271254 TI - Neurotoxicity after orthotopic liver transplantation in cyclosporin A- and FK 506 treated patients. AB - Neurotoxicity is a serious complication following orthotopic liver transplantation leading to increased morbidity and mortality. Neurotoxicity may be evoked by various perioperative factors, or may be due to drug-specific toxicity of immunosuppression. In the present study we evaluated the incidence of central nervous system (CNS) toxicity occurring within the early postoperative period of 121 patients, 61 randomly assigned to FK 506- and 60 to CsA-based immunosuppression as part of a multicentre study. The incidence of moderate or severe CNS toxicity was higher in patients treated with FK 506 (21.3%) than in patients receiving CsA (11.7%). The duration of symptoms was also greater in patients treated with FK 506 than in patients receiving CsA. The incidence of moderate or severe neurotoxicity after retransplantation was markedly greater in patients treated with FK 506 (100% of the patients). PMID- 11271255 TI - Presence of polymorphonuclear granulocytes during the early stage of transplant arteriosclerosis after prolonged ischemia in the rat. AB - The presence and function of polymorphonuclear granulocytes has been investigated, in particular, in the microcirculation in many short-term models of ischaemia/reperfusion injury. The aim of this study was to examine the presence of granulocytes in the aorta in a recently established long-term model of transplant arteriosclerosis, based on prolonged cold graft ischaemia time in the rat. Aortic grafts of PVG donors were subjected to two different cold ischaemia times of 1 and 4 h (n = 5 in each group) before an orthotopic transplantation to syngeneic recipients. The grafts were explanted shortly after various times post reperfusion (7.5 min 24 h) and examined with conventional staining, immunohistochemistry and transmission electron microscopy for the presence of granulocytes. The results showed the presence of these cells adherent to the endothelial layer or in the subendothelial layer in grafts with both ischaemia times and with a maximum seen 2 h after transplantation. The internal elastic lamina was interrupted at sites of granulocyte adherence. We concluded that the polymorphonuclear granulocyte may be involved in the ischaemia/reperfusion injury in this model, thus, contributing to the development of accelerated transplant arteriosclerosis. PMID- 11271256 TI - Effect of 15-deoxyspergualin on allograft arterio-sclerosis and growth factor synthesis in the rat. AB - Chronic rejection is a major cause for late graft loss. Typical vascular changes in the grafts are adventitial inflammation, disappearance of myocytes in the media and thickening of the intimal layer. We investigated the effect of a new immunosuppressive drug, 15-deoxyspergualin (DSG), on chronic rejection using our rat aortic allograft model. At the dose of 1.0 mg/kg per day, DSG significantly reduced all histopathological parameters of chronic rejection, thus, inhibiting the generation of allograft arteriosclerosis. Growth factor synthesis in the grafts was determined by reverse transcription reaction with oligo dT primers and semiquantitated by polymerase chain reaction. The expression of several growth factors, PDGF-BB, IGF-1, EGF an TGF-beta, was suppressed to 16-60% of the non treated allograft level. This indicated that DSG may work via suppression of growth factor synthesis and, thus, inhibits the generation of chronic rejection. PMID- 11271257 TI - The immunomodulatory effects of a novel agent, leflunomide, in rat cardiac allotransplantation. AB - We assessed the immunomodulatory effect of leflunomide (LEF) in a heterotopic abdominal model of rat heart transplantation using a major histocompatibility mismatch (DA X LEW). The endpoint of this study was cardiac rejection assessed by abdominal palpation of the ventricular impulse and confirmed by laparotomy and histology. In this study, LF was investigated using four dosages (5, 10, 20 and 30 mg/kg per day orally) against cyclosporine (CsA) (15 mg/kg per day orally) and FK 506 (1 mg/kg per day orally). The ability of LEF to prevent rejection and reverse ongoing acute rejection was assessed. The results showed that untreated hearts were fully rejected by day 5 and that LEF at 5 mg/kg was significantly better than any other dose tested, was superior to FK 1 mg/kg, and was as effective as CsA 15 mg/kg in preventing rejection after a short course of treatment. After a longer course, 10 and 20 mg/kg LEF proved more effective than 5 mg/kg in controlling rejection and as efficacious as 1 mg/kg FK and 15 mg/kg CsA. In the control of ongoing established early rejection. LEF proved to be equally effective, even at lower doses (5 mg/kg), to CsA 15 mg/kg and FK 1 mg/kg. In the control of ongoing established late rejection, 20 mg/kg LEF proved to be superior to 10 mg/kg LEF and 15 mg/kg CsA, and was as effective as FK 1 mg/kg. However, when higher doses of CsA (25 mg/kg) and FK (2 mg/kg) were tested against 20 mg/kg LEF in this mode of rescue, LEF proved as effective as CsA and superior to FK. In the assessment of drug toxicity using body weight as a parameter, 20 mg/kg LEF proved safer than any other LEF dose tested, and safer than 15 mg/kg CsA and 1 mg/kg FK in both short- and long term administration. We conclude that LEF is a relatively safe and potent immunosuppressant with promising clinical potential. PMID- 11271258 TI - Induction of class II molecules by cytomegalovirus in rat heart endothelial cells is inhibited by ganciclovir. AB - Cytomegalovirus (CMV) has been demonstrated to induce class II antigen expression in endothelial cells. To study whether ganciclovir (DHPG) has an effect on CMV induced class II expression, cultured rat heart endothelial cells were infected with rat CMV (RCMV) and treated with different DHPG concentrations. Class II antigens in endothelial cells were detected by a monoclonal antibody and immunoperoxidase technique. Control cells did not express class II antigen, but during RCMV infection 92% of cells were class II-positive. DHPG treatment (1, 10, 100 and 1000 microg/ml) decreased RCMV-induced class II expression from 73% to 59%, 6% and 0%, respectively. As DHPG inhibits CMV DNA polymerase, our present results suggest that DHPG affects RCMV-induced class II expression via the inhibition of RCMV DNA replication. PMID- 11271259 TI - Effect of ganciclovir prophylaxis on cytomegalovirus-enhanced allograft arteriosclerosis. AB - Clinical and experimental studies have established the accelerating role of cytomegalovirus (CMV) infection on cardiac allograft arteriosclerosis, i.e. chronic rejection. In this study, we investigated the effect of ganciclovir prophylaxis on the development of CMV-enhanced allograft arteriosclerosis. Rat aortic allografts were done from DA donors to WF recipients and either infected with 10(5) plaque-forming units of rat CMV (RCMV) 1 day after transplantation or left uninfected. RCMV-infected allografts received ganciclovir at an initial dose of 20 mg/kg and a maintenance dose of 10 mg/kg twice a day for 14 days. Grafts were removed at 7, 14 days, and 1 and 3 months after transplantation and processed for morphometry and autoradiography. The results of this study demonstrated that ganciclovir prophylaxis significantly delays and reduces RCMV enhanced allograft arteriosclerosis in the rat. PMID- 11271261 TI - Measuring left ventricular function after heart transplantation via digitization of M-mode echocardiograms. AB - The aim of the study was to assess the usefulness of M-mode echocardiography for noninvasive diagnosis of cardiac rejection. For this purpose, 292 M-mode images of 26 heart transplant recipients were analyzed. The echocardiographic images were digitized into an image analysis system. The curves of left ventricular diameter changes were obtained and its first differential calculated. A total of 23 parameters were measured. The most important parameters were: peak velocity of systolic diameter change, peak velocity of diastolic diameter change, time to peak velocity of systolic diameter change, time to peak velocity of diastolic diameter change (TPEAK-D), isovolumetric relaxation time (IVRT), rapid filling time (RFT), shortening fraction (SF), and mean velocity of circumferential fiber shortening (MVCF). The echocardiographic parameters were compared to biopsy results. In 18 patients, 23 biopsy-proven moderate rejections occurred. When rejection occurred, IVRT decreased 23% +/- 6% (P < 0.05), SF decreased 13% +/- 14% (P<0.05), MVCF decreased 18% +/- 18% (P<0.05), and TPEAK-D increased 27% +/- 27% (P<0.05). We concluded that the analysis of digitized M-mode images can identify heart transplant rejection. PMID- 11271260 TI - Is the incidence of cytomegalovirus disease following heart transplantation decreased by prophylactic ganciclovir and CMV-hyperimmunglobulin? AB - Ganciclovir (DHPG) was used for the prophylaxis of CMV disease after heart transplantation (HTx) in 20 patients (aged 52 +/- 8 years old). DHPG was used during the first 2 weeks post HTx, and during antirejection therapy with OKT3 or thymoglobulin (ATG), at a dosage of 3 mg/kg q 12 h in the case of a CMV+ donor (D) and/or CMV+ recipient (R). CMV-hyperimmunglobulin (-Ig, 1 ml/kg per week for 6 weeks) was added in the case of a CMV+ donor. A historical control group included 18 HTx patients (aged 53 +/- 10 years old). We excluded the combination of CMV- donor and CMV- recipient. Both groups received the same immunosuppression with methylprednisolone (MP), azathioprine, ATG, and cyclosporine A. The global incidence of CMV disease was 15% (3/20 patients) in the study group and 11% (2/18 patients) in the control group. Similar results were observed in the D+/R- combination (study group 40%, 2/5 patients; control group, 25%, 2/8 patients) and in cases of R+ irrespective of D status (study group, 7%, 1/15 patients; control group 0%, 0/10 patients). No difference was observed in both groups with respect to the incidence of CMV disease after antirejection therapy either with MP or with OKT3/ATG. At 1 year post HTx, no difference was found in the incidence of acute rejection, coronary artery disease or other etiology of infection or mortality. All patients CMV disease responded to a 14-day course of DHPG (5 mg/kg q 12 h). No relapsing disease was observed, and no patient died from CMV. Our results suggested that at the doses and time-scale used, DHPG, with or without CMV-Ig did not reduce the incidence of CMV disease after HTx. PMID- 11271262 TI - The effects of single lung transplantation in rats with monocrotaline-induced pulmonary hypertension. AB - Acute haemodynamic change after single lung transplantation for primary pulmonary hypertension was evaluated using a rat transplantation model. Inbred Fisher 344 rats were administered with 40 mg/kg monocrotaline in order to induce pulmonary hypertension. The rats whose mean pulmonary arterial pressure (PAP) was over 30.0 mm Hg received a left lung isograft from a normal donor after right heart catheterization. In the control group, PAP increased after single lung transplantation. On the other hand, in the pulmonary hypertensive group, PAP was significantly decreased 60 min after the transplantation, but 3 and 6 h after the transplantation, the PAP significantly increased again. On the day after the operation, it again decreased significantly. Left-to-right lung blood flow ratio was significantly increased in rats with pulmonary hypertension compared to rats with normal pulmonary pressure on both the 1st and 3rd postoperative days. The oedema of the grafted lung was more severe in the pulmonary hypertensive group than in the control group in the acute phase. In conclusion, single lung transplantation for pulmonary hypertension shifted pulmonary blood perfusion to the grafted lung and this shift made pulmonary oedema of the grafts more severe in the acute phase. These oedematous changes, which were more pronounced in the grafts in the pulmonary hypertensive rats, might have contributed to the transient rise in PAP in those rats after single lung transplantation. PMID- 11271263 TI - Pleural-changes in the lung allograft during acute rejection. AB - To ascertain the cause of pleural fibrosis in lung allografts, pleural changes were investigated in rat syngeneic and allogeneic lung grafts. The pleura of lung syngeneic grafts showed no pathological changes except for mild edema on the first day after transplantation. In lung allografts, recipient cells migrated into the subpleural tissue early after transplantation (latent phase). In the vascuar phase, recipient lymphocytes in the subpleural tissue increased in number, while almost all alveolar structures were free from infiltration. Both CD4-positive and CD8-positive cells infiltrated in almost equal numbers with macrophages. The subsets of infiltrating cells were similar to those of the perivascular and peribronchial areas. In the late vascular or alveolar phase, fibroblasts were observed among the infiltrating cells, and fibrotic changes started. In the destructive phase, collagen formation with marked pleural thickening was dominant. Pulmonary acute rejection should be treated at least up to the late vascular phase to prevent pleural fibrosis. PMID- 11271264 TI - Neutrophil elastase and obliterative bronchiolitis. AB - Bronchoalveolar lavage levels of elastase were assayed to determine the timing and magnitude of elevations in elastase relative to both fibrosis, as indicated by hyaluronate (HA) levels, and decline in FEV1 characteristic of the clinical syndrome of obliterative bronchiolitis (OB). Samples were collected from 48 heart lung or single lung transplant recipients. Regression analysis was performed and demonstrated that high levels of elastase occurred with active decline in lung function and in association with high levels of HA. This study suggested that intense neutrophil elastase release occurs concurrent with the development of OB and may contribute to the destruction of bronchiolar architecture. PMID- 11271265 TI - Paediatric incidence of acute rejection and obliterative bronchiolitis: a comparison with adults. AB - Obliterative bronchiolitis (OB) continues to be a major cause of morbidity and mortality following heart-lung transplantation. We compared the incidence of death from obliterative bronchiolitis in 19 children and 72 adults following heart-lung transplantation at our institutes. The incidence of death from OB at 2 years was 38% for children compared with 17% for adults, this difference was significant (Cox-Mantel Z value = 2.243, P < 0.05). The frequency of acute lung rejection and persistent lung rejection, previously described as risk factors for OB in adults, were significantly more common in children, P = 0.004 and P = 0.001, respectively. Average forced expiratory volume in 1 s was lower in children than in adults for each 3-month period after transplantation (P < 0.001). In conclusion, identified risk factors for the development of OB were more common, and the risk of death from OB was greater in children than in adults following heart-lung transplantation. PMID- 11271266 TI - Combined liver and pancreatic islets transplantation in man using cyclosporin immunosuppression. AB - We report a 1-year patient and graft survival after combined liver-pancreatic islet transplantation. The patient was affected by a pancreatic neuroendocrine carcinoma with extensive liver metastasis. Native pancreas and total liver removal was undertaken after a good response to chemotherapy, and the patient was still cancer-free 1 year later. Normal liver function and insulin independence was achieved, although islet response to glucose challenge remained delayed. Immunosuppression was maintained with cyclosporin monotherapy. PMID- 11271267 TI - Our experience with Roux-Y intestinal drainage in simultaneous kidney and pancreas transplantation. AB - Enteric drainage is a sound surgical technique in SKP, and it avoids the majority of urological as well as metabolic complications. We did not see an increase in intraabdominal complications or of graft loss due to rejection. Intestinal leak is rare and easily managed provided a Roux-Y loop of jejunum is used. Even though the number of patients was small and the follow-up short, the results of the RY group were at least comparable to the BD group. In view of our results, we plan to use this technique in all our future SKP patients. PMID- 11271268 TI - Simultaneous pancreas/kidney transplantation--the optimal therapy for type I diabetics with end-stage renal disease in Europe, too? AB - Contrary to the situation in the USA, the number of pancreatic transplantations declined during the last year in the Eurotransplant region. Whether the high postoperative morbidity and unsatisfactory graft function rates reported by many European centres can be overcome was investigated in a single centre study. In a consecutive series of 80 patients with simultaneous pancreas/kidney transplantations, postoperative morbidity due to graft pancreatitis and recurrent rejections was significant. Both of these complications, however, were treated successfully in the vast majority of patients. Graft thrombosis was almost completely prevented. Excellent function rates of the pancreatic grafts of 88% after 1 year and 83% after 5 years were achieved. Thus, simultaneous pancreas/kidney transplantation can be recommended as the optimal therapy for type I diabetics with end-stage renal disease in Europe, too. PMID- 11271270 TI - Combined pancreatic and kidney transplantation: en bloc retrieval and transplantation--a new surgical technique. AB - We designed and performed on two patients a new surgical procedure of en bloc kidney and pancreatic transplantation. The liver, pancreas and kidneys were removed en bloc in the donor. On the bench, the liver and the left kidney were separated from the bloc, leaving the pancreas and the right kidney for combined kidney and pancreatic transplantation. The portal vein was divided near to the emergence of the splenic vein. The coeliac axis was taken with an aortic patch. The left renal vein was cut at its entrance to the inferior vena cava (IVC) and the left renal artery was taken with an aortic patch. Reconstruction of the pancreatic vessels was performed with a double anastomosis: the portal vein was anastomosed to the hole in the IVC resulting from the section of the left renal vein and the splenic artery was anastomosed to the hole in the aorta resulting from the section of the left renal artery. The proximal ends of the aorta and IVC were closed with running sutures. In the recipient, the iliac vessels on the right side were dissected. Anastomosis of the distal part of the aorta and the IVC was performed with the right iliac vessels. Duodenocystostomy and reimplantation of the ureter were done according to the usual techniques. This new surgical technique allowed an easy vascular reconstruction of the pancreatic vessels. In the recipient, only one side was used for renal and pancreatic transplantation. Moreover, the length of the transplant procedure was significantly reduced. PMID- 11271269 TI - Long-term follow-up of lipid metabolism and rheologic properties after successful pancreas and kidney transplantation. AB - The long-term effect of pancreatic and kidney transplantation (spkt) on blood viscosity, lipid metabolism and skin microcirculation in insulin-dependent diabetes mellitus (IDDM) was studied because impaired rheological properties of blood may play a role in the development of diabetic micro- and macroangiopathy. 46 IDDM-patients (16 f/30 m; 23 +/- 34 y mean duration of diabetes; 60 +/- 14 mos mean follow up period) underwent spkt (Gr.I: n = 28) or solitary kidney (Gr.II: n = 18) transplantation, and were compared with healthy controls (C). Rheological measurements were performed with Mooney-Ewart rotation-viscosimeter determining whole blood viscosity (WBV), at shear rates 1, 5, 10, 20, 50, 100, 200 sec(-1). Triglycerides, total and HDL-, LDL- and VLDL cholesterol and fibrinogen were measured. Microcirculation was estimated by transcutaneous oxygen tension measurement (tcpO2) and laser speckle method, in the forefoot area. Hemoglobin A1 was normalized only in group I (I: 7.2 +/- 0.2%; II: 8.3 +/- 0.3%; C: < 8%). WBV at low shear (1, 5, 10) was increased in both groups, when compared to healthy controls (I: 12.4 +/- 2; 12.5 +/- 1; 6.8 +/- 0.5 mpas; II: 18.7 +/- 2; 13.4 +/- 15; 9.4 +/- 1 mpas; C: 7.5 +/- 0.5; 6.7 +/- 0.3; 5.4 +/- 0.2 mpas; P < 0.05). Plasma fibrinogen was elevated in both groups compared to normals: (I: 384 +/- 19; II: 448 +/- 20; C: 250 +/- 50 mg/dl; P < 0.05). There was a positive influence of spkt on skin microcirculation: tcpO2/prior tx: I: 44 +/- 3; II: 49 +/- 6 mmHg; post tx: I: 59 +/- 4; II: 42 +/- 3 mmHg. Laser speckle prior tx I: 3.3 +/- 0.3; II: 4.7 +/- 0.2 rel. U.; post tx: 3.8 +/- 0.2; II: 4.3 +/- 0.2 rel. U. Patients with progression of angiopathy showed still higher fibrinogen and shear rates (P < 0.05). There was no significant difference for total HDL-, LDL- and VLDL cholesterol. Despite normalization of glucose metabolism and significant improvement of microcirculation in spkt patients, fibrinogen and the shear rates are increased indicating a persisting "individual" vascular risk. It is suggested that an additional hemorheological approach in the treatment posttrransplant might prevent the progression of vascular complications. PMID- 11271271 TI - Does intrathymic injection of alloantigen-presenting cells before islet allo transplantation prolong graft survival? AB - Current immunosuppressive agents have potentially dangerous side-effects, are non specific and most are also diabetogenic. We investigated tolerance induction with intrathymic injection of purified antigen-presenting cells (APC) plus a single dose of antilymphocyte serum (ALS) intraperitoneally before allogeneic islet transplantation in the rat model WAG to Lewis (RT1u to RT1l). Purified donor APC [non-parenchymal cells (NPC) or dendritic cells (DC)] were prepared from liver and spleen, respectively. Isograft function for more than 120 days proved that islet isolation, purification and transplantation procedures were adequate. A total of WAG DC (4 x 10(5)) or NPC (2 x 10(6)) in 20 microl were injected into both lobes of the thymus of 140-210 g Lewis recipients followed by a single injection of ALS. Three days later, diabetes was induced with streptozotocin (60 mg/kg). Four days later allogeneic islets were grafted into the liver by intraportal injection of 3000 WAG islets. Control animals (n = 8) received 20 microl saline intrathymically instead of APC. Graft function was assessed by blood glucose measurements with glucose levels above 15 mmol/l on 3 consecutive days defined as graft rejection. Animals given DC (n = 9) or NPC (n = 8) intrathymically plus 1 ml of ALS, rejected their grafts in an accelerated fashion with a median survival time (MST) of 3 days. However, control animals rejected their grafts with a MST of 7 days, but with two animals surviving for more than 2 months. In conclusion, intrathymic inoculation with purified APC plus a single dose of ALS did not prolong allogeneic islet graft function but induced accelerated rejection of the islet allografts. PMID- 11271272 TI - Clinical experience with human anodal trypsinogen (HAT) for detection of pancreatic allograft rejection. AB - To date one of the major dilemmas in clinical pancreas transplantation is the lack of a reliable indicator for pancreas rejection. In a consecutive series of 52 patients undergoing simultaneous pancreas and kidney (SPK) transplantation with bladder drainage technique between October 1991 and December 1992 a new test using serial levels of serum human anodal trypsinogen (HAT) was evaluated for its efficacy to detect pancreas rejection. Postoperative baseline levels of HAT were compared to peak HAT values at time of rejection. HAT profiles at time of rejection were calculated and compared to profiles of urinary amylase, serum amylase, fasting blood sugar and serum creatinine. In this series one year patient survival was 97%, graft survival of the pancreas 86% and of the kidney 90%. In 71% of the patients at least one rejection episode occurred. At time of kidney-biopsy proven rejection with a concurrent serum creatinine rise a significant HAT level rise to more than 1000 ng/ml was observed from baseline levels of 200 ng/ml (P < 0.001) indicating kidney and pancreas rejection (73%). Urinary amylase levels decreased in the majority of rejection episodes at the same time from baseline levels to less than 20,000 U/l. In 25% of the rejection episodes a significant serum creatinine rise was observed without a HAT rise or urinary amylase decrease indicating kidney-only rejection, while in 2% a urinary amylase decrease and simultaneous HAT also was observed with a negative kidney biopsy indicating pancreas-only rejection. We feel that increase in HAT levels significantly correlates with pancreas rejection. After SPK, determination of HAT is an additional helpful non-invasive test. In pancreas transplantation alone HAT can be a useful indicator to detect rejection and facilitate timing of a pancreas biopsy and initiation of antirejection treatment. PMID- 11271273 TI - Neurological examinations after liver transplantation concerning patients under corticosteroid immunosuppression and either FK 506 or cyclosporin. AB - To study the neurological sequelae in liver transplanted recipients, 25 patients were followed up between 5 and 30 months after transplantation and another 14 patients were seen before and after transplantation. Physical examination took special note of tremor and polyneuropathy; additionally, patients estimated concentration and memory, tremor, paraesthesias and sleep disturbances on a self rating scale. Tremor was reported to be preexistent in 50% of the later FK 506 and cyclosporin group and only temporarily rose afterwards. Twenty-eight percent complained of tremor and 20% said that it interfered mildly with daily activity. Only 2 of 39 patients showed new signs of polyneuropathy. Concentration and memory improved significantly after transplantation. In the second group of patients, MRI, EEG, lumbar puncture and neuropsychological tests were done just before and routinely after transplantation, revealing numerous preexisting neurological deficits with only singular changes afterwards. PMID- 11271274 TI - Combined hepatocyte-islet transplantation: an allograft model. AB - Experimental hepatocyte transplantation (Tx) has been shown to improve the survival rate of acute hepatic failure (AHF) in different models. Histological and biochemical data from some studies suggest more satisfactory function of hepatocytes after combined hepatocyte-islet Tx. The aim of the present study was to compare the survival rate between two different sites of Tx (kidney subcapsular and spleen) of hepatocytes alone or combined with islets of Langerhans in rats with surgically induced AHF (90% hepatectomy) accross a major histocompatibility barrier (WAG to Lewis). Rats were divided into five groups (n = 6 in each group). Group 1 consisted of AHF without treatment, group 2, AHF followed by hepatocyte Tx into the spleen, group 3, AHF followed by hepatocyte Tx subcapsular into the kidney, group 4, AHF followed by combined hepatocyte islet Tx into the spleen, and group 5, AHF followed by combined hepatocyte islet Tx subcapsular into the kidney. The number of hepatocytes was 10(7) and the number of islets was 400. All rats received cyclosporin A (CsA) i.v. (20 mg/kg on days 0 4 and 10 mg/kg on days 5-30). Hepatocytes were harvested using a modification of the portal vein collagenase perfusion (type V 1.3 mg/ml) and islets, with the collagenase digestive technique (type XI 1 mg/ml). All Tx took place 24 h after AHF. All rats in group 1 died within 48 h. In groups 2 and 3, the combined survival rate was 33% 1 month after Tx, while in groups 4 and 5, the combined survival rate was 50% at 1 month. All surviving animals were sacrificed and histological examination showed well-preserved hepatocellular aggregates in the spleen and beneath the renal capsule, as well as islets around the clusters of hepatocytes. SGOT and SGPT values were also measured. We concluded that the combined Tx in a rat experimental allo-Tx model in cases of AHF improves the survival rate in comparison with hepatocyte Tx alone. The survival rate at the two different sites for combined Tx was similar. PMID- 11271275 TI - Metabolic intervention to affect canine pancreas recovery following ischemia during preservation by the two-layer method. AB - We have demonstrated that a high adenosine triphosphate (ATP) level in a canine pancreas during preservation by the two-layer method is an important determinant for the ultimate success of pancreatic transplantation. In this study, we investigated (a) the effect of factors that seemed to have an influence on energy metabolism in the canine pancreas at the tissue ATP level and (b) graft viability during preservation by the two-layer method. ATP tissue concentration was determined by high-performance liquid chromatography and graft viability was assessed on the basis of survival rate following autotransplantation. First, the pancreas was harvested from either 72-h-fasted (n = 5) or fed dogs (n = 5) and preserved by the two-layer Euro-Collins solution (EC)/perfluorochemical (PFC) method for 24 h. All the pancreatic grafts were viable in both fed and fasted groups. There was also no significant difference in ATP tissue concentration between the two groups (7.48 +/- 0.55 vs. 7.03 +/- 0.74 micromol/g dry weight, NS). Second, the pancreatic grafts subjected to 60 min of warm ischemia were preserved by either the two-layer (EC/PFC) or (EC + adenosine/PFC) method for 24 h. Without adenosine, ATP tissue concentration did not recover (1.62 +/- 0.26 after warm ischemia vs. 1.56 +/- 0.40 micromol/g dry weight after preservation, NS) and all the pancreatic grafts failed. However, provision of adenosine led to restoration of ATP tissue levels (1.90 +/- 0.53 vs. 7.23 +/- 2.17 micromol/g dry weight, P < 0.01) and four of five grafts functioned immediately and maintained normoglycemia after transplantation. These results clearly demonstrated that the nutritional state of the pancreatic graft before procurement had no influence on ATP tissue level as well as graft viability during 24-h preservation by the two layer method. On the other hand, provision of adenosine during 24-h preservation enhanced ATP synthesis of the pancreatic tissue, thereby improving viability of the ischemically damaged pancreas. PMID- 11271276 TI - Difference in energy metabolism between fresh and warm ischemic canine pancreases during preservation by the two-layer method. AB - We have demonstrated that adenosine triphosphate (ATP) is synthesized within a canine pancreas during preservation by the two-layer method and there is a direct correlation between a high ATP tissue level and good posttransplant outcome. The purpose of this study was to examine the difference in energy metabolism between fresh and warm ischemic pancreases during preservation by this method. First, fresh pancreases were preserved with simple cold storage in Euro-Collins solution (EC; group 1A), or by the two-layer method using EC (group 1B), EC with 2,4 dinitrophenol (DNP; group 1C), an uncoupler of oxidative phosphorylation, or modified EC (ECM; group 1d), which contained mannitol in place of glucose for 48 h. ATP tissue concentrations in group 1B were significantly higher than in group 1A (7.91 +/- 1.21 vs. 1.21 +/- 0.31 micromol/g dry weight, P < 0.01) but almost equal to group 1d (7.91 +/- 1.21 vs. 7.59 +/- 0.97 micromol/g dry weight, NS). DNP (group 1C) caused a significant decrease in tissue ATP levels in group 1A (0.61 +/- 0.07 vs. 7.91 +/- 1.21 micromol/g dry weight, P < 0.01). Second, pancreases subjected to 60 min of warm ischemia were preserved by simple cold storage with EC (group 2A) or the two-layer method using EC (group 2B) or EC with adenosine (group 2C) for 24 h. ATP tissue levels in groups 2A and 2B after preservation were 1.40 +/- 0.46 and 1.56 +/- 0.40 micromol/g dry weight and graft survival rates were 0/5 (0%) and 0/3 (0%), respectively. However, tissue ATP levels in group 2C after preservation were significantly higher compared with the value before preservation (7.23 +/- 2.17 vs. 1.90 +/- 0.53/g dry weight, P < 0.01) and graft survival rate was 4/5, 80%. Other nucleosides, hypoxanthine, inosine, and adenine did not substitute for adenosine. In addition, studies with [2-3 H] adenosine demonstrated that almost all of the adenosine was converted to adenine nucleotides. This study clearly demonstrated that fresh grafts synthesize ATP mainly via mitochondrial oxidative phosphorylation using endogenous substrates. However, after significant warm ischemia, pancreases produce ATP mainly via direct phosphorylation of exogenous adenosine during preservation by the two-layer method. PMID- 11271277 TI - Preservation studies using acinar cell cultures of the pancreas: stimulation of amylase/lipase release before and after hypoxic stress. AB - In clinical pancreas transplantation, postischemic (i. e. postpreservation) transplant pancreatitis is a major problem in some cases but also an interesting model of pancreatitis. In this study, the effect of simulated organ preservation of isolated acinar cells was evaluated as regards enzyme release (basic and cerulein-stimulated), using common preservation solutions as the incubation media. Primary pancreas acinar cell cultures were isolated from the pancreas of male Wistar rats using the modified method of Amsterdam and Jamieson. After resting the cells in culture flasks for 1 week, monolayer cultures were obtained. The basic enzyme release of amylase and lipase was measured as well as the effect of stimulation with cerulein (10(-8) M) and the effect of replacing the medium (without changing the consistence or temperature of the medium). In a second step, the cultures were incubated under conditions of cold hypoxia for 6 h (4 degrees C, P O2 < 0.1 mm Hg) using Krebs-Henseleit solution (KH), Euro-Collins solution (EC), HTK solution of Bretschneider (HTK) or University of Wisconsin solution (UW) as the incubation solution. After 6 h, the media were changed to warm normoxic KH, and a second stimulation test with cerulein was performed. The native microstructure of the cultures was observed as well. Enzyme release was elevated by a factor of 5 by stimulating the acinar cells with cerulein as well as by changing the medium in the experiments prior to the hypoxic incubation. After hypoxic incubation and change to KH, the morphology of the cultures was excellent, while the basic enzyme release was on a very low level, no matter which preservation solution was used during cold hypoxia. Stimulation with cerulein caused only minimal elevation of enzyme release during an observation period of 60 min. These observations show that cold storage in preservation solution provides maintenance of the cell morphology and sufficient down regulation of enzyme release of pancreatic acinar cells. Thus, acinar cells alone do not seem to be the pacemaker of pancreatitis after organ preservation. The presented experimental model will be the subject of extended evaluation in the future. PMID- 11271278 TI - The effect of deoxyspergualin (DSG) on rejection and graft-versus-host disease (GVHD) after small bowel transplantation. AB - Both rejection and graft-versus-host-disease may occur after fully allogeneic small bowel transplantation. In this study, we established unidirectional models of rejection and GVHD in rats and evaluated the efficacy of 15-deoxyspergualin (DSG). When F1 small bowel was transplanted into LEW rats (rejection model) the graft was acutely rejected. The administration of DSG (5 mg/kg per day for 10 days) significantly prolonged the survival, but was efficacious only when used prophylactically. When a unidirectional GVHD model (F1 --> LEW SBTx) was examined, the administration of DSG from day 0 after grafting greatly suppressed GVHD, resulting in more than 300 days survival. However, only cutaneous GVHD, but not fatal GVHD, was suppressed when the start of administration was postponed until day 4 after grafting. From in vitro studies, DSG inhibited natural killer cell activities to K-562 and skin epidermal cells. The response was well correlated with in vivo GVHD course. These results suggest that DSG is an effective immunosuppressant for both rejection and GVHD when used prophylactically. DSG exerted the effect more stongly against cutaneous GVHD than fatal GVHD by inhibiting natural killer systems. PMID- 11271280 TI - Clonal deletion and clonal anergy in allogeneic bone marrow chimeras prepared with TBI or TLI. AB - The evolution of Vbeta6-expressing C3H (H2k, Thy 1.2, Mls a-) lymphocytes was investigated in C3H recipients mice pretreated with total body irradiation (TBI) or total lymphoid irradiation (TLI) and infusion of AKR (H2k, Thy 1.1, Mls a+) cells. After TBI (9.5 Gy) all Vbeta6+ Thy 1.2 (C3H) cells, which are capable of reacting against the Mls a antigen that like expressed by AKR mice, were deleted in the thymus and the periphery in stable bone marrow (BM) chimeras obtained by infusion of 5 x 10(6) T-cell-depleted (TCD) AKR BM cells. When, in the opposite combination, 30 x 10(6) C3H spleen cells were infused into TBI-treated AKR cells, all animals developed graft-versus-host disease (GVHD) with no clonal deletion and in contrast, showed an increase in Vbeta6+ C3H cells. After injection of 30 x 10(6) AKR BM cells into TLI-treated C3H mice no C3H cells were detected in the thymus and only a small percentage in the periphery. Within these C3H cells Vbeta6+ cells were only partially deleted and anergized as they did not respond in vitro after stimulation with Mls a+ AKR cells or anti-Vbeta6 mAb. Cells suppressing anti-Mls a-reacting C3H cells were not found. After injection of 15 x 10(6) AKR cells more C3H cells were found in the thymus, but only a minority of Vbeta6+ cells persisted in the periphery of these animals. In conclusion in TBI prepared chimeras only clonal deletion occurred, whereas in TLI-prepared chimeras both clonal deletion and anergy occurred in maintaining tolerance. PMID- 11271281 TI - Characterization of improved renal transplant preservation mechanisms using PB-2 flush solution by HPLC assay. AB - Renal flush solutions have been found to be beneficial for extending organ viability, but their mechanisms of action are poorly understood. In order to delineate these mechanisms we studied the addition of mannitol and adenosine to a modified simple hypothermic intracellular flush solution (PB-2), and the relationships of renal adenine nucleotide (AN) concentrations with ischemia, reperfusion and viability. A significant (P < 0.05) and progressive decay in AN and increase in total degradation products (DP; hypoxanthine and xanthine) was noted during warm ischemia. Total AN and AMP were significantly higher after 50 h of cold storage in the PB-2 compared to the C-2 control cold flush group. Viability was associated with a significant (P < 0.01) regeneration of AN within 45 min of reperfusion. HPLC assay indicated that PB-2 cold flush solution enhances renal viability by both diminution of reperfusion injury enabling better reflow and primary preservation of AN. The latter process may appreciably contribute to the former process. PMID- 11271279 TI - Cellular mechanisms of alloimmune non-responsiveness in tolerant mixed lymphocyte chimeras induced by vascularized bone marrow transplants. AB - It has been demonstrated that the development of stable mixed lymphocyte chimerism is associated with alloimmune tolerance induction in vascularized bone marrow transplant (VBMT) recipients. The underlying mechanisms of immune non responsiveness in tolerant VBMT chimeras remains unclear. Our VBMT model involves the transplantation of a parental donor limb (Lewis rats) onto a hybrid (Lewis x Brown Norway) F1 recipient. Tolerogenic mechanisms and cellular immune regulation to self and host allodeterminants were investigated during the early post transplant phase of tolerance induction. Flow cytometric analysis of sIg+ depleted experimental peripheral blood lymphocytes from tolerant VBMT recipients demonstrated low level stable mixed immune chimerism. Chimeric cells tested for responsiveness against self-LEW determinants showed activated proliferation and immune dysregulation 30 days post-transplantation. However, direct immunocytolytic activity against LEW determinants was not found. Tolerant chimeras also demonstrated elevated cellular proliferation and cytolytic responses against host-specific BN allodeterminants at 30 days. Consistent with these in vitro findings, limited clinical signs compatible with GVH reactivity were evident in vivo at this time. Following this initial period, the tolerant VBMT animals returned to normal clinical condition and remained otherwise healthy throughout the study. Consistent with these results, VBMT chimeras then showed declining proliferative responses from the elevated values seen at 30 days against self-LEW determinants. Proliferative and immunocytolytic responses also decreased against host-specific BN allodeterminants from peak levels at 30 days. In conclusion, these results provide evidence that the initial phases of tolerance induction in VBMT chimeras consist of self- and alloimmune regulation that follow an early period of immune dysregulation. Sequential phases of immune dysregulation and re-regulation elucidated in VBMT stable mixed chimeras within the first 100-day period may represent important mechanisms of tolerance induction. PMID- 11271282 TI - Improved renal preservation with PB-3 flush solution during 72 h of cold storage. AB - Previously, PB-2 flush solution has been found to be superior to Collins 2 solution (C-2) in extending renal viability in the dog. To further characterize preservation mechanisms, we studied mitochondrial oxidative function during 72 h of cold storage comparing PB-3, UW-1, and C-2 flush storage solutions. Complex 1 dependent mitochondrial oxidative phosphorylation (MOP) was found to be significantly less (P < 0.001) at 5 h and 72 h of cold storage (P < 0.001) for the C-2 compared to the other flush groups. Complex 2 dependent MOP had parallel results, having significantly less function (P < 0.002) at 5 h and 72 h (P < 0.02) of cold storage in the C-2 group compared to the other groups. PB-3 and UW 1 solutions were noted to be comparable, suggesting possible equivalent preservation efficacy. Nevertheless the components of PB-3 at the level of MOP contributed significantly to better preservation compared to C-2 solution. PMID- 11271283 TI - Renal protective effect of liposomed superoxide dismutase in an experimental warm ischemia model. AB - Superoxide dismutase (SOD) is a potent scavenger of superoxide radicals produced during normothermic ischemia-reperfusion. Since it has a short half-life, its optimal effect is achieved when it is given prior to reperfusion. The inclusion of SOD in liposomes (lipo-SOD) prolongs its half-life (free SOD: 6 min; lipo-SOD: 4 h). The protective effect of lipo-SOD in a 60-min bilateral renal warm ischemia model was studied. We divided 60 male Wistar rats between two control groups and five study groups according to the drug used (SOD or lipo-SOD) and to the time of SOD administration (prior to ischemia or prior to reperfusion). SOD and lipo-SOD were both given at 20 mg/kg endovenously. Weight, diuresis, creatinine per 100 g (Cr/100 g), and creatinine clearance per 100 g (CrCl/100 g) were studied. Conventional renal histology was performed after reperfusion and on day 7. Renal malondialdehyde, 6 keto PGF 1 alpha, and TxB2 tissue levels were studied after reperfusion. Results showed that the renal protective effect of free SOD on warm ischemic-reperfusion injury depended on the time of administration, being more effective when given before reperfusion. On the other hand, the renal protective effect of liposomed SOD did not depend on the time of administration since efficacy was similar when given before reperfusion or before ischemia. The functional protective effect of liposomed SOD was similar to that of free SOD when they were given prior to reperfusion. Nevertheless, since histological damage observed with liposomed SOD was less than with free SOD, it is suggested that the liposomed galenic form may offer better protection against renal warm ischemia. In addition, liposomed SOD was better at preventing tissue prostanoid generation after renal warm ischemic-reperfusion injury than free SOD. We concluded that liposomed SOD shows a higher renal protective effect against warm ischemia than free SOD. PMID- 11271284 TI - In situ perfusion and UW solution used for storage did not decrease the incidence of ATN in kidneys harvested from hemodynamically unstable donors. AB - The incidence of acute tubular necrosis ATN after cadaveric kidney transplantation in our centre has been in the range of 50%. A prospective study was carried out in 1991 and 1992 to assess the effect of in situ perfusion and hypothermic storage of kidneys harvested from brain-dead haemodynamically stable and unstable donors. Three litres of Ringer's solution were used for in situ perfusion. In 40 cases, the kidneys were stored in Euro-Collins (EC) solution and in the other 78 cases, in University of Wisconsin (UW) solution. Among the factors that could contribute to ATN, we analysed warm ischaemia time, anastomosis time and cold storage time. Function was considered to be delayed if the patient required posttransplantation dialysis. The donors were considered haemodynamically unstable when hypotension before harvesting was present (BP < 70 mm Hg over 2 h) despite high doses (> 15 microg/kg per minute) of dopamine or when cardiac arrest occurred at the time of harvesting and oliguria had been present for at least 2 h. Haemodynamically stable donors with a BP greater than 80 mm Hg had a normal diuresis. In all donors in this group the dose of dopamine was lower than 10 microg/kg per minute. The study showed that storage in UW solution did not influence the incidence of ATN in kidneys harvested from haemodynamically unstable donors. Differences observed in our study were due to haemodynamic status preceding donor nephrectomy and length of cold storage time. PMID- 11271285 TI - A report of the eurotransplant randomized multicenter study comparing kidney graft preservation with HTK, UW and EC solutions. HTK study group. AB - Special attention has been focused in this randomized study on the primary function of renal allografts preserved with different solutions. Histidine Tryptophane-Ketoglutarate (HTK) and University of Wisconsin (UW) solutions provided a significantly lower incidence of delayed graft function compared to Euro-Collins solution. Improved renal function after transplantation was observed in the HTK and UW groups compared to the EC group. PMID- 11271286 TI - Comparison of UW versus HTK solution for myocardial protection in heart transplantation. AB - In order to evaluate the protective effect of University of Wisconsin (UW) solution in heart transplantation, a retrospective comparative study with histidine-tryptophane-ketoglutarate (HTK) solution was initiated. In group I, we included 160 patients with HTK preservation, while group II consisted of 50 patients who had their transplant protected with UW solution. All patients received standard quadruple drug therapy for immunosuppression. The average ischaemic time of the donor hearts in group I was 142+/-44 min, ranging from 83 to 235 min. Acute immediate perioperative graft failure occurred in six cases (3.8%). Statistical analysis including the chi-square test, revealed a significant increase in the incidence of acute perioperative graft failure when compared with duration of ischaemic time (P < 0.01). Within the first 30 postoperative days, 24 patients died (15% early mortality). The same statistical correlation was evident between the incidence of early mortality and duration of graft ischaemic time. The 30-day and 6-month survival rates were 81% and 78%, respectively. The average ischemic time of the donor hearts in group II was 193+/ 50 min ranging from 100 to 360 min, which was significantly longer in comparison with the group I (P < 0.05). Acute perioperative graft failure occurred once (2%); the patient was retransplanted successfully. Five patients died within the first 30 postoperative days (10% early mortality). There was no correlation between length of ischaemic time and incidence of acute graft failure or early mortality. The 30-day and 6-month survival rates were 90% and 88%, respectively and, thus, better when compared with group I. In both groups similar results were achieved with regard to postoperative NYHA status of the patients and incidence of cardiac arrhythmias. Myocardial preservation with HTK solution showed satisfying results as long as the ischaemic time did not exceed 4 h. The early functional results achieved with UW graft protection were excellent, even with ischaemic times longer than 4 h and not depending on lenght of ischaemic period. PMID- 11271287 TI - Successful cardiac presrevation for 12 hours using nondepolarizing cold cardioplegia. A canine model. AB - Isolated canine hearts were preserved for 12 h at 5 degrees C followed by normothermic reperfusion for 2 h. Dogs were divided into two groups: group 1 (n = 7) received a nondepolarizing preservation solution in multidose, and group 2 (n = 6) received single-flushed University of Wisconsin (UW) solution, both administered in multidose fashion. At the end of reperfusion, the myocardial adenosine triphosphate concentration and left ventricular systolic and diastolic function were preserved better in groupl than in group 2. Myocardial mitochondrial ultrastructural integrity was identical in the two groups. These results suggested that in a 12-h heart preservation, nondepolarizing solution administered in multidose fashion protects the myocardium from the deleterious effects of hypothermia and cardioplegia better than UW solution. PMID- 11271288 TI - A prospective randomized clinical trial of liver preservation using high-sodium versus high-potassium lactobionate/raffinose solution. AB - High-sodium as opposed to high-potassium lactobionate/raffinose preservation solution offers potential advantages in improving the quality of liver storage by reducing potassium-induced vasoconstriction and preventing hyperkalaemia on reperfusion. In our study we evaluated in a prospective trial (encompassing 40 consecutive cadaver donor hepatic retrievals and subsequent transplants) the efficacy of a high-sodium formulation versus the standard high-potassium solution. Quality of preservation was assessed by clinical indices of liver function in the intraoperative and early postoperative phases, including measurements of requirements for blood and blood products and potassium, circulating liver enzymes and bilirubin. Frequencies of acute rejection episodes and primary non-function were also recorded. No significant differences were evident in any of the measured parameters. Thus a sodium-based solution can be used for hepatic preservation, advancing the possibility that it may be possible to develop a single storage solution for clinical multi-organ donor operations. PMID- 11271290 TI - Delayed oxidation of intramitochondrial pyridine nucleotide oxido reduction state as compared with tissue oxygenation in human liver transplantation. AB - Intra- and post-operative oxygenation of graft liver and subsequent oxidation of the intramitochondrial redox state of pyridine nucleotide were studied in liver transplantation from living related donors with the arterial ketone body ratio (AKBR), the ratio of oxidized flavoprotein to reduced pyridine nucleotide (FP/PN ratio), and oxygen saturation of hemoglobin in liver tissue (hepatic SO2). The subjects involved in this study consisted of 20 pediatric patients. Hepatic SO2 was measured by near-infrared tissue spectroscopy. FP/PN ratio was measured by two-dimensional fluorometric scanning. Tissue oxygenation was normalized at the end of the operation. By contrast, AKBR remained at a low value at the end of the operation. At 24 and 48 h after the operation, the AKBR values increased to near the control value, indicating that it took 24 h for the intramitochondrial redox state to be normalized. The FP/PN ratio also remained at a low value at the end of the operation as compared with the control value. In conclusion, the observed delay in oxidation of the intramitochondrial redox state as compared with tissue oxygenation indicated the transition of the redox state associated with the changes in the metabolic state, and suggested the important role of microcirculation in the normalization of the redox state. PMID- 11271289 TI - High concentrations of adenosine are needed in Carolina rinse to prevent activation of Kupffer cells. AB - Adenosine is a key component of Carolina rinse solution, which was developed to prevent an O2-dependent reperfusion injury following liver transplantation. Kupffer cells are activated following liver transplantation, an effect minimized by adenosine. In this study, the concentration of adenosine was varied in Carolina rinse, Ringer's, and Ringer's containing hydroxyethyl starch. Following 24 h of cold storage in University of Wisconsin cold storage solution, colloidal carbon uptake was measured to assess phagocytosis by Kupffer cells. Carbon uptake was elevated significantly in this model from values of 136 +/- 15 in unstored control livers to 187 +/- 25 mg/g per hour following a rinse with 10 ml of Ringer's solution. Phagocytosis was reduced about 50% when 0.1 mM adenosine was added to the Ringer's solution; however, addition of up to 1 mM adenosine in Carolina Rinse did not prevent elevated phagocytosis. In contrast, values were reduced significantly by addition of 5-10 mM adenosine to Carolina Rinse or Ringer's containing hydroxyethyl starch. Thus, the higher concentrations of adenosine needed to reduce Kupffer cell activation can best be explained by binding of adenosine to hydroxyethyl starch. PMID- 11271291 TI - Successful 48-h liver preservation by controlling nutritional status of donor and recipient. AB - The nutritional status of the donor has been shown to affect the outcome of liver transplantation in the rat. It has been proposed that this may be due to inhibition of Kupffer cell induced injury to the reperfused organ, which leads to an inflammatory type response. In this study we investigated how altering the nutritional status of the recipient affects the outcome of liver transplantation after preservation of the liver for 44 or 48 h in the University of Wisconsin (UW) solution. The nutritional status of the rats was altered by either fasting or by feeding an essential fatty acid free diet (EFAD) for 2 months. This type of diet has been shown to reduce significantly the inflammatory response in rats. Survival after 44-h preservation of livers from fed donors (fed a standard laboratory diet) transplanted to fed recipients was 29% (2/7) but increased to 80% (4/5) when the recipient was fed the EFAD diet. After 48-h preservation, there were no survivors under either of these two dietary combinations. However, survival was 100% after 48-h preservation if the donor had been fasted for 4 days and the recipient was fed the EFAD. These results showed that the nutritional status of the donor and recipient are important factors in the outcome of liver transplantation. How nutritional factors affect liver preservation and transplantation are not clear but may be related to the inflammatory response regulated by Kupffer cells and circulating neutrophils in the liver, both of which are influenced by the diet of the animal. PMID- 11271292 TI - Neurological recovery after liver transplantation: a prospective study of 22 patients. AB - A group of 22 liver transplantation patients were examined pre- and postoperatively using clinical neurological, neurophysiological and neuroradiological methods. After the operation improvement was observed in neurological symptoms, and in neuropsychological and neurophysiological test results. Our study shows that liver recipients have a high prevalence of nervous system dysfunction and that successful transplantation is followed by significant improvement. PMID- 11271293 TI - Endothelin-1 is involved in hepatic sinusoidal vasoconstriction after ischemia and reperfusion. AB - Endothelin-1 (ET-1), a vasoactive peptide, causes a significant rise in portal vein pressure, which is most likely a result of severe vasoconstriction in the liver. In this study, the effect of ET-1 on sinusoidal vasoconstriction in the liver after ischemia and reperfusion was directly investigated using intravital microscopy. In anesthetized female Sprague Dawley rats (200-250 g) ischemia of the median and left liver lobes was induced for 90 min by temporary ligation of the left pedicle. After declamping and a 90-min reperfusion period, the livers were exposed for intravital microscopy. Using a Nikon MM-11 fluorescence microscope (545 nm, 330x), a CCD camera (Cohu FK 6990), and a SVHS video recording unit, the hepatic microcirculation was directly investigated. Besides sham groups, two ischemia groups were studied, receiving ET-1 antiserum (anti-ET 1; 0.5 ml; Peptide Inst., Osaka, Japan) or NaCl 0.9% (0.5 ml) 5 min prior to reperfusion of the liver (n = 6/group). Following a transient drop in the mean arterial blood pressure in the anti-ET-1-treated groups, comparable systemic hemodynamic conditions among the four groups were noted during intravital microscopic assessment at the end of the 90-min reperfusion period. Reduction in the sinusoidal diameters during postischemic reperfusion (7.7 +/- 0.5 microm) was prevented by anti-ET-1 treatment (9.6 +/- 0.25 microm; P < 0.01; mean + SEM) back to control values (9.6 +/- 0.32 microm), while most of other microcirculatory parameters did not show significant differences. The results supported further the role of ET-1 in dysregulation of the sinusoidal vascular tone in the liver, e.g., after ischemia and reperfusion. PMID- 11271294 TI - Immunodepressants ameliorate normothermic ischemia injury to the rat liver by down-regulating tumor necrosis factor, not by alleviation of lipid peroxidative injury. AB - Mechanisms by which immunodepressants (Cyclosporine, CsA; FK 506, FK; Azanthioprine, AZA) ameliorate warm ischemic injury of the liver were examined. Female Sprague-Dawley rats were subjected to 60-min of normothermic liver ischemia. Animals were assigned to one of four groups: group I, control with vehicle treatment; groups II, III, and IV, treatment with CsA (10 mg/kg), FK (1 mg/kg), and AZA (1 mg/kg), respectively. The immunosuppressive agents were given per os for 4 consecutive days prior to the induction of hepatic ischemia. In addition to a survival study, plasma levels of endotoxin, serum activities of tumor necrosis factor-alpha (TNF), plasma levels of phosphatidylcholine hydroperoxide (PCOOH) as a lipid peroxide, and serum alanine aminotransferase (ALT) were investigated in blood samples collected from the suprahepatic vena cava. A 7-day survival period was significantly higher in the immunosuppressed animals. Serum TNF levels were elevated and peaked at 3 h following reperfusion. When, the peak values were compared, the animals given immunodepressants had significantly lower levels of TNF (217.0 +/- 40.6 pg/ml for group I, 67.6 +/- 13.7 for group II, 87.9 +/- 28.3 for group III and 89.1 +/- 19.9 for group IV; Mean +/- SEM). Plasma PCOOH levels were also elevated following reperfusion, but with no statistical difference among the groups. Our data suggest that immunodepressants ameliorate warm ischemia/reperfusion injury through modulation of TNF production and not through a diminution of lipid peroxidative injury. PMID- 11271295 TI - A method for HLA-DQA typing by the PCR-SSP technique. AB - There is preliminary evidence that matching for HLA-DQ is important for kidney graft survival. We developed a method for HLA-DQA typing based on the PCR-SSP principle. The procedure consisted of three steps: DNA isolation, PCR amplification and visualization of the PCR product under UV light. For the identification of all currently known DQA1 alleles, we designed 18 different primers that allowed typing for the specificities DQA1*0101, *1012, *0103, *0104, *0201, *03, *0401, *0501 and DQA1*0601. For the typing of a single individual, 12 PCR mixes were needed, each containing a primer pair specific for a certain allele group, and a pair of control primers that amplified a non-polymorphic region. The time required for this procedure was approximately 3 h from the time of blood collection. Comparison of this method with DQA typing by the RFLP method in 151 individuals revealed only a single discrepancy. The method can be easily applied for prospective cadaver donor typing. PMID- 11271296 TI - Comparison of serological and DNA HLA-DR typing results for transplantation in Western Europe, Eastern Europe, North America and South America. AB - In a previous study, DNA typing revealed that 25% of serological HLA-DR typings of kidney transplants were incorrect. In the current study, we analyzed whether this error rate had improved in recent years, and whether there were differences according to geographical region. From 1988 to 1991 the error rate of serological typing improved slightly in Western Europe from 19% to 16%, and in North America, from 21% to 16%. In Eastern Europe, the error rate decreased from 49% to 33% in 1991, whereas the rate remained high in South America at 60% in 1988 and 72% in 1991. The high error rates in South America and Eastern Europe reflected a lack of good quality serological typing reagents. The 16% typing errors in Western Europe and North America demonstrated the current limit of serological techniques for cadaver donor typing and underlined the need for prospective DNA typing. PMID- 11271297 TI - Nephrotoxicity after orthotopic liver transplantation in cyclosporin A and FK 506 treated patients. AB - Nephrotoxicity represents a serious side-effect of immunosuppression following orthotopic liver transplantation. In order to preserve the therapeutic potential of cyclosporin (CsA) and FK 506 in human liver transplantation and to differentiate the nephrotoxic action of either drug in a clinical setting, we evaluated the incidence of early and late nephrotoxicity in 121 patients, 60 randomly assigned to CsA- and 61 to FK 506-based immunosuppression. Early postoperative renal insufficiency (between POD0 and 30; SCr 1.5-3 mg/dl) was observed to a similar extent in patients treated with CsA (36.7%) and FK 506 (42.6%). Early postoperative acute renal failure (ARF; SCr > 3 mg/dl) occurred in 18.3%, regardless of the immunosuppressive management. Approximately 50% of patients with ARF required hemodialysis (CsA: 11.7%; and FK 506: 8.3%). Mean onset of hemodialysis in CsA-treated patients was POD1 and in FK 506-treated patients, POD6, which demonstrated a different time course of drug-specific nephrotoxicity of CsA and FK 506 in early ARF. All patients with early postoperative ARF requiring hemodialysis survived more than 1 year (100% survival). New onset of late ARF (between POD30 and 365), however, occurred in 6.5% under FK 506 and in 1.7% under CsA immunosuppression due to severe infections with the multiple organ failure syndrome. This observation was consistent with the assumption of over-immunosuppression rather than a primary nephrotoxic effect. Mortality of patients with late ARF requiring hemodialysis was 100%. Late renal insufficiency appeared in 23.3% of CsA- and in 29.4% of FK 506-treated patients, and represented a slowly progressing form of drug-specific nephrotoxicity. These preliminary results demonstrated a similar outcome in terms of early and late nephrotoxicity, but longer follow-up will delineate its overall efficacy and toxicity in humans. PMID- 11271298 TI - HLA class II DNA-RFLP typing in 102 individuals from Northern Greece. AB - The serological identification of HLA class II alleles is often doubtful. Since accurate HLA typing is essential for the matching of donor-recipient pairs in allogeneic transplantation, an effort was made to establish DNA restriction fragment length polymorphism (RFLP) typing and to assess the correlation between the serological and RFLP techniques in the population of Northern Greece. One hundred and two healthy individuals (204 HLA-DR alleles) from Northern Greece were HLA-DR, DQ typed with both the microcytotoxicity and the Taq I RFLP method, using three exon-specific probes. DNA-RFLP typing revealed (1) concordant results with serology in 69.9% (142/204) of the alleles and (2) at least one HLA-DR allele discrepant to serology in 30.4% (62/204) of the alleles. Incorrect serological DR types (weak reactions or inability to distinguish between two alleles with a common epitope) were identified in 54 alleles (26.5%), while 3.9% (8/204) of serological "blank" alleles turned out to be definable alleles by RFPL. Of the individuals tested, 10.8% (11/102) were DR-homozygous by RFLP. This comparison of results obtained by serology and RFLP demonstrated the necessity of the clinical application of DNA typing, especially for organ transplantation where accurate HLA typing has an important influence on graft survival. PMID- 11271299 TI - A retrospective flow cytometric crossmatch study in transplant recipients with autoreactive lymphocytotoxic antibodies. AB - Our previous data shows renal transplant recipients with autoreactive lymphocytotoxic antibodies to have a reduced transplant survival when compared to patients without autoantibodies. This could have been due to the presence of weak IgG antibodies inhibited by the dithiothreitol used to remove IgM antibodies in the pretransplant cytotoxicity crossmatch. That possibility was investigated in a retrospective study of 52 recipients of 57 renal transplants who were recrossmatched using a more sensitive flow cytometry crossmatch (FCXM) to detect recipient IgG antibodies to donor T and/or B cell splenic lymphocytes. Fourteen of the 57 (24%) transplants failed. Six losses were within the 1st month posttransplant and four of these were immunological failures. None of the transplant failures had a positive pretransplant FCXM. These results showed that the recipients with autoantibodies did not have pretransplant IgG anti-donor antibodies. The transplant failures did not, therefore, relate to the presence of antibodies undetected by the dithiothreitol-treated cytotoxicity crossmatch. PMID- 11271300 TI - Selection of the most compatible graft recipient by computer program, used in regional tissue typing laboratory of Hellas (R.T.T.L.). AB - The purpose of this study was to present the new computer program that we developed in the regional tissue typing laboratory (R.T.T.L.) and use for the selection of the most suitable recipient for cadaveric allografts in a more efficient way. This new program was written and compiled in TURBO PASCAL 6.0, sorts all possible recipients to the HLA type of a donor, checks for the existence of splits and utilises them, and when requested, gives out results to more specific inquiries, i.e. all compatible recipients from 2DR2B2A to 1DR0B0A matching. It can also forecast success percentages according to different factors, i.e. combination of donor's and recipient's ages, HLA matching etc. The material used for this study were the patients who are registered in the formal cadaveric transplantation list of R.T.T.L. We have used this program since 2 January 1992 together with the old one. Since then we found that the new program is faster in sorting all the possible recipients of cadaveric renal allografts according to the criteria already mentioned. The total selection time, with all the criteria activated, averages a few seconds, whereas with the old program it took approximately 2 min just for the sorting of HLA matching, without any other criteria activated. In the printout of the final result of each inquiry are all the possible recipients in the sorted order together with relevant data (telephone number, address etc.). As a result, the laboratory personnel has been free from the tedious task of this sorting which was initially done by hand and the possibility of error has been eliminated. The program was developed exclusively by doctors and all the updates needed are done by the users. More important, however, is the fact that in many cases the time of cold ischaemia was reduced by more than 30 min with all the obvious advantages for the longevity of the graft's life. PMID- 11271301 TI - Profile of cytokine production in mixed kidney lymphocyte culture. AB - Kidney cells are an important source of immunoregulatory molecules that regulate cell-to-cell interactions, which is the key step in the generation of the immunoresponse to alloantigens. In this study we identified the cytokines that are produced by both lymphoid cells and kidney cells when coincubated in mixed kidney lymphocyte cultures (MKLC). The capacity of kidney cells to stimulate the proliferation of effector allogeneic lymphocytes was assayed by incubating irradiated kidney cells and lymphocyte. The cytokine secretion profile in MKLC was investigated by incubating monolayers of kidney cells with effector peripheral blood mononuclear cells (PBMC). The culture supernatants were harvested on days 1, 2, 3, 4, 5, 6, 7, and 8 and assayed for IL-1beta, IL-2, IL 6, and TNF alpha using an ELISA. Kidney cells, in comparison to PBMC stimulator cells were poor stimulators of the alloproliferation even when HLA expression was increased by IFN gamma treatment. Compared to lymphocyte or kidney cells incubated alone, MKLC induced a considerable stimulation of cytokine production. This increase in cytokine production was observed essentially for IL-2 and IL-6 (at day 3, a 10-fold increase in IL-2 and a 5-fold increase in IL-6). This study provided evidence that target kidney cells and effector lymphocyte interactions generate a number of cytokines such as IL-1beta, IL-2, IL-6, or TNF alpha. These cytokines are known to modulate allo-proliferation and generation of cytotoxic T lymphocytes (CTL). PMID- 11271302 TI - Interleukin-8 serum and urine concentrations after kidney transplantation. AB - We conducted a prospective study of 12 patients undergoing kidney transplantation. In these patients, we monitored interleukin-8 (IL-8) in both serum and urine before and after kidney transplantation. Levels of IL-8 were analyzed by a solid-phase double ligand ELISA method. Three patients with an uneventful recovery after transplantation showed IL-8 serum levels below the detection limit, whereas some small amounts were detected in the urine of these patients. IL-8 serum levels markedly increased with acute graft rejection and infection. Increments in serum and urine preceded clinical complications in all patients. Highest levels were observed in bacterial infection and lowest in acute rejection. Although nonspecific, IL-8 can be considered as an indicator molecule of inflammatory processes occurring during kidney transplantation. PMID- 11271303 TI - In vivo near-infrared monitoring of nitric oxide production and tissue oxygen sufficiency in rat liver allografts during the acute rejection reaction. AB - We established a new technique of in vivo near-infrared (NIR) spectroscopy that can estimate both nitric oxide (NO) production and tissue oxygen sufficiency in living organs during the alloimmune response. The present study was aimed at evaluating the potential of this technique for monitoring the rejection response utilizing the rat model of orthotopic liver transplantation without arterialization. The relative changes of nitrosyl-hemoglobin, oxyhemoglobin and oxidized-cytochrome oxidase in the graft livers were quantified by use of this method. Nitrosyl-hemoglobin in the allogenic grafts was elevated at the onset of the rejection response and was suppressed when the rejection reaction was treated by the administration of 15-deoxyspergualin. Oxy-hemoglobin and oxidized cytochrome oxidase were decreased in accordance with parenchymal disorder determined histologically. These results demonstrated that the new technique of in vivo NIR spectroscopy can assess simultaneously both the immune response and graft function after liver transplantation. PMID- 11271304 TI - FK 506 and cyclosporin each block antigen-induced T cell receptor signalling that is dependent on CD4 co-receptor and operates in the absence of detectable cytoplasmic calcium fluxes. AB - The T cell hybridoma "171", which lacks CD4 but expresses T cell receptor (TCR) for hen egg white lysozyme, requires introduction of wild-type CD4 for antigen mediated induction and secretion of interleukin-2 (IL-2). Mutant CD4, which fails to associate with the tyrosine kinase p56lck does not support IL-2 secretion, suggesting that a role of CD4 is to bring cytoplasmic p56lck into alignment for signal transduction to the IL-2 promotor. Using 171, 171-CD4 (wild-type) and 171 CD4 (mutant), we found that IL-2 secretion was inhibited by FK 506 and cyclosporin but not by rapamycin. However, this inhibition was not associated with calcium fluxes since no change in cytoplasmic free calcium levels ([Ca]i; resting level 80 nM) was detectable during antigen stimulation of the 171 or 171 CD4 cells. Thus, although FK 506 and cyclosporin inhibited calcium-dependent signalling to the IL-2 promoter via inhibition of the protein phosphatase calcineurin, it is possible that IL-2 induction via TCR/CD4 requires an FK 506 (and cyclosporin) sensitive step which is independent of cytoplasmic calcium changes. PMID- 11271305 TI - Effect of immunosuppressive agents FK 506 and cyclosporin and steroids on the expression of IL-6 and its receptor by stimulated lymphocytes and monocytes. AB - The interaction of interleukin-6 (IL-6) with its receptor (IL-6R) is not well understood. In the present study, we investigated the effect of different immunosuppressive agents on the expression of the couple IL-6/IL-6R on cultured lymphocytes and monocytes. IL-6 in culture supernatants from cultured monocytes were analyzed by ELISA. The expression of IL-6R was studied by flow cytometry. Dexamethazone, cyclosporin (CyA), and FK 506 at immunosuppressive concentrations induced a dose-dependent inhibition of IL-6 secretion from adherent monocytes (MO) stimulated with phytohemagglutinin (PHA). Dexamethazone was the most effective agent in inhibiting IL-6 secretion, while the inhibitory effect observed with 1 ng/ml FK 506 was comparable with that obtained with 100 ng/ml CyA. Unstimulated MO strongly expressed IL-6R (80% positive cells). Stimulation of MO with PHA resulted in a significant downregulation of IL-6R expression. Treatment of PHA-stimulated adherent MO with different concentrations of CyA and FK 506 induced a restoration of IL-6R expression. FK 506 was 100 time more effective in restoring IL-6R than CyA. This restoration of IL-6R was incomplete. FK 506, CyA, and steroids may exert their immunosuppressive effect by inhibiting IL-6 secretion and partially restoring MO IL-6R, which may be important in protecting the cell target against IL-6 autocrine stimulation. PMID- 11271306 TI - In vitro and in vivo effects of BT 563, an anti-interleukin-2 receptor monoclonal antibody. AB - BT 563, a murine anti-IL-2R MoAb, was found to be more potent than anti-Tac in inhibiting proliferation in the mixed lymphocyte reaction. Results obtained with 33B3.1 in these experiments were similar to those with BT 563. The anti-IL-2R MoAb 2A3 was shown to be a suitable agent for monitoring the effect of BT 563 on peripheral blood. IL-2R-positive cells were not detected in peripheral blood samples from 1 h after the first dose until 8 days after the last dose. Plasma trough levels were measured in patients receiving 5 or 10 mg daily. The administration of BT 563 to allograft recipients did not lead to clinically significant side effects. PMID- 11271307 TI - Mechanisms of unresponsiveness associated with pretransplant blood transfusion cyclosporine-induced mixed lymphocyte chimerism. AB - Multiple pretransplant blood transfusions while under limited daily cyclosporine cover (PTBT-CsA) induce extensive rat renal allograft survival and antigen specific non-responsiveness. The underlying mechanisms of this extensive allograft survival are not yet fully understood. We hypothesized that one of the potential contributing mechanisms to tolerance induction in PTBT-CsA-treated kidney recipients is the development of stable mixed chimerism, putatively due to the proliferation of stem cells capable of haematopoiesis in the transfused blood. BN rats served as whole blood and kidney donors. LEW rats served as recipients of the PTBT-CsA protocol and BN kidney transplants. Three weekly transfusions were given under concomitant limited CsA cover. Following these multiple primary sensitizations, antigen-specific splenic cellular responsiveness in vivo was normal in comparison with naive animals. However, these experimental splenocytes were non-specifically suppressed against third-party allodeterminants. At 100 days post-transplantation (T100) following tolerance induction to kidney allografts (secondary challenge), in vivo adoptive transfer experiments demonstrated the existence of potent splenic suppressor cells. In vitro suppressor cell assays confirmed that these cells were non-specific suppressor cells. However, following chimerism stabilization at T130, splenic antigen-specific suppressor cells became exclusively expressed in the tolerant animals, replacing the non-specific suppressor cells. At this time, splenic microchimerism was at peak levels and remained stable from T100 to T130. In conclusion, these findings demonstrate that sequential mechanisms of suppressor cell network expression are induced within a chimeric environment by blood-CsA immune modulation. Stable mixed lymphocyte chimerism and related immunomodulatory mechanisms may, therefore, play an important tolerogenic role in blood-CsA induced non-responsiveness and in the beneficial effect of blood transfusion. PMID- 11271308 TI - T-cell anergy: a consequence of interaction between T cells and allogeneic rat renal epithelial cells. AB - In order to study the immunogenicity of parenchymal cells within an allograft, renal tubular cells were propagated from both PVG and DA strain rats. These cells were induced to express class II major histocompatibility (MHC) antigens by stimulation for 4 days with interferon-gamma (IFN-gamma). It was found that resting lymphoid cells derived from Lewis rats responded vigorously after stimulation with irradiated splenic cells from PVG rats. However, stimulation with renal cells from PVG rats did not result in interleukin (IL-2) production or lymphoproliferation. Furthermore, lymphocytes from this mixture failed to respond to secondary stimulation by PVG splenic cells; lymphocytes primed by mixture with DA renal cells responded normally to secondary stimulation by PVG splenic cells. These results indicate that renal epithelial cells can specifically anergise allogeneic lymphocytes. PMID- 11271310 TI - Distribution and persistence of antigen-presenting cells after intrathymic injection. AB - Intrathymic injection of donor immune cells has been shown by previous studies to prolong survival of rat allogeneic tissues. The aim of this pilot study was to assess the distribution and the persistence of intrathymically (i.t.) injected purified antigen presenting cells (APC) over a period of time in the rat model DA to-WAG (RT1av to RT1u) using a specific monoclonal antibody (Mab) with RT1Aa class I polymorphic specificity (R3/13 clone). Purified non-parenchymal cells (NPC) or dendritic cells (DC) were prepared from liver and spleen of DA rats with a purity of greater than 60% and 90%, respectively, shown by selected Mab staining methods. DA NPC (1 x 10(6)) or DC (5 x 10(5)) in 20 microl were injected into both lobes of the thymus of WAG rats with or without 1 ml antilymphocyte serum (ALS) intraperitoneally. Thymus tissue was removed on days 3, 5, 10, 20 and 30, and processed for frozen sections and immunohistochemical staining with R3/13. Numerous DA-positive cells were detected in the first 3-10 days post-i.t. inoculation in both NPC- and DC-treated rats, with or without ALS. After day 10, the proportion of positive cells decreased in all cases except in rats given NPC and ALS, where similar numbers of R3/13-positive cells were seen throughout. These DA-positive cells were mostly found in the medullary portion of the thymus at the corticomedullary junction in close proximity to thymic dendritic interdigitating cells. We concluded from this pilot study that the injected cells remained in the thymus for a limited period. However, the immunosuppressive effect of ALS promoted some degree of persistence of donor APC in the thymus beyond 30 days. Further studies are in progress to reveal the specificity of these cells. PMID- 11271309 TI - Low-dose heparin: a novel approach in immunosuppression. AB - Very low subcutaneous doses of standard and low molecular weight heparin inhibited the traffic of sensitized lymphocytes to a graft site, reduced in situ mononuclear cell infiltration and prolonged skin allograft survival in mice. Similar effects were caused by low doses of oral heparin, while higher oral doses prolonged graft survival. Our results suggest that oral heparin may have immunosuppressive properties applicable in clinical transplantation. PMID- 11271311 TI - Early infiltration of renal allografts with 27E10-positive macrophages and graft outcome. AB - Recently, we have demonstrated that acute cellular rejection is correlated with a massive infiltration of 27E10-positive macrophages. To examine the distribution of macrophage differentiation markers in the infiltrate in the very early post transplantation period, two biopsies were taken intraoperatively, approximately 1 h following reperfusion, in each of 16 renal transplant recipients. One biopsy was taken for conventional histology and the other biopsy was snap-frozen. The sections were stained using an ABC indirect immunoperoxidase technique. A panel of monoclonal antibodies against three macrophage differentiation markers (27E10, 25F9 and RM3/1) was used to stain the sections. Using the early inflammation macrophage marker 27E10, there was an unexpected strong staining in 3 out of 16 biopsies. This severe infiltration of 27E10-positive macrophages with 10-20 macrophages per high power field (compared to 0-2 in others) was correlated in all cases with a poor outcome of the graft. All seven kidneys with no 27E10 positive infiltration showed a good function 6 weeks post-transplantation. The other macrophage markers, 25F9 and RM3/1, showed a less marked correlation with graft outcome. In conclusion, a massive infiltration of renal allografts with 27E10-positive macrophages 1 h post-transplantation may be a very early predictor of poor graft outcome. PMID- 11271312 TI - Optimal FK 506 dosage in patients under primary immunosuppression following liver transplantation. AB - In a retrospective study, we analysed the FK 506 dosage used in primary liver graft recipients enrolled in the European FK 506 multicenter trial conducted from September 1990 to January 1992. In addition, a second cohort of patients treated more recently in a single centre was investigated. The impact of different dosing strategies on the clinical course of the patients was analysed with special emphasis on the incidence of rejection episodes and FK 506 side-effects. Among the patients enrolled in the European FK 506 multicenter trial, those patients enrolled during the "early" phase of the study received a higher oral FK 506 dose [mean oral dosage on day 7 = 0.19 mg/kg body weight (bw) per day, n = 134] compared to patients enrolled during the "late" period of the study (mean oral dosage on day 7 = 0.14 mg/kg bw per day, n = 133). This lower dosage was the result of several protocol amendments performed to reduce the incidence of FK 506 side-effects. Lowering of the FK 506 dosage was accompanied by a reduction in the long-term prevalence of side-effects such as diabetes (n. s.) or hypertension (P < 0.05), while patient survival and rejection frequency remained constant. Patients treated in centres with online FK 506 blood level monitoring experienced significantly less hypertension, less episodes of diabetes and less rejection episodes compared to patients treated in centres without. The clinical course of those patients enrolled in the multicentre trial was compared with the course of a cohort of liver-grafted patients treated with FK 506 more recently in a single centre. These patients had a further reduction in the FK 506 dosage (0.10 mg/kg bw per day p.o. or less according to whole blood levels, with no intravenous FK 506 administration). When compared to patients enrolled in the multicentre trial, these patients experienced less side-effects (nephrotoxicity, hypertension, serious early neurotoxicity) while adaequate immunosuppression was maintained. PMID- 11271313 TI - Prolonged rat allograft survival induced by temporary elimination of alpha/beta T cells with monoclonal antibody. AB - We tested the ability of lewis (LEW; RT-1(1)) recipients to reject DA (RT-1av1) cardiac allografts following the selective elimination of alpha/beta T cells with the mouse monoclonal antibody R73. One group of adult rats (6 weeks old) received 1000 microg R73 i.p. on days 2 and 1 before transplantation, and 100 microg R73 every third day after transplantation up to day 18. Prolonged cardiac graft survival was noted (30, 30, 32, 51, 62, 108, > 500, > 500, > 500 days). Untreated controls (n = 10) rejected their grafts within 7 +/- 1 days. R73 therapy induced a dramatic decrease in alpha/beta T cells from 69% before treatment to 5% within the first 5 days, followed by an increase to 64% by day 8. The T cell increase was paralleled by the appearance of anti-mouse antibody. A second group of adult rats (10 weeks old) received the same treatment. These "older" recipients rejected their grafts within 20 +/- 5 days. Chronic R73 therapy from birth until the day of transplantation (100 microg R73 i.p. twice a week) resulted in graft survival of 37 +/- 9 days in eight animals. Two rats had a graft survival of more than 200 days. When chronic R73 therapy was continued to day 70 after transplantation, DA hearts were accepted well in all animals for more than 100 days. Alpha/beta T cells were virtually absent throughout the whole time of treatment. Antibodies against R73 were not detected. We concluded that selective elimination of alpha/beta T cells has a strong effect on allograft survival. PMID- 11271314 TI - Heart allograft survival in rats following immunization with soluble peptide MHC class I donor antigens: evidence for the role of indirect recognition in rejection. AB - The current series of experiments addressed the question of whether indirect priming with donor MHC antigens affects heart allograft survival. LEW (RT-1(1)) rats were immunized with a mixture of two peptides corresponding to the variable region of MHC class I locus Aa antigen (alpha1 and alpha2 domain). The recipients were transplanted with a DA (RT1-1a) heart 1 month after immunization, and graft survival was closely monitored by ECG. All peptide-treated recipients presented with anti-peptide antibodies at the time of transplantation and developed a strongly accelerated graft rejection. These findings indicated that indirect recognition of MHC I donor antigens promotes heart allograft rejection. PMID- 11271315 TI - Pretransplant serum IgA concentration and IgA-anti-Fab autoantibody activity as prognostic indicators of kidney graft survival. AB - IgA concentration and IgA-anti-Fab autoantibody activity were tested in pretransplant sera of 308 kidney graft recipients. Recipients with a serum IgA concentration of 2 g/l or greater had a 1-year graft survival rate of 83%, compared with a 68% rate in recipients with serum IgA of less than 2 g/l (P < 0.005). Serum IgA concentration and IgA-anti-Fab autoantibody activity were significantly associated (r = 0.38, P < 0.0001). Recipients with a high pretransplant IgA-anti-Fab activity had a significantly better graft survival rate (81%) than patients with low pretransplant IgA-anti-Fab (67%, P < 0.025). When IgA-anti-Fab and serum IgA were considered together, 137 recipients with high IgA-anti-Fab and high serum IgA had a 86% 1-year graft survival rate, which was significantly better than the 63% survival rate in patients with low IgA-anti Fab and low serum IgA (P < 0.0005). The pretransplant serum IgA level and IgA anti-Fab autoantibody activity were excellent predictors of kidney graft outcome. PMID- 11271316 TI - Mechanism of suppression of cloned human suppressor T cells. AB - We report the mechanism of suppression of suppressor T cell clone III-1-C5 using helper T cell clone III-1-B6, mitogen responses and rIL-2. Clone III-1-C5 suppressed the mixed lymphocyte reaction (MLR) by secreting alloantigen non specific, MHC non-restricted suppressor factor(s). Clone III-1-C5 did not suppress mitogen (PHA, Con A, PWM) response nor proliferation by exogeneous rIL 2. Clone III-1-C5 suppressed proliferation by clone III-1-B6, which augments proliferation by direct cell to cell contact with responder cells and not by soluble factors. These results indicated that suppressor T cells exhibit suppressive effects not only by inhibiting IL-2 synthesis but by inhibiting the direct effects of helper T-cells. PMID- 11271317 TI - Long-term survival of heart grafts in the presence of donor-specific cytotoxic T cell precursors (CTLp) in the peripheral blood. AB - To monitor their immunological status we determined donor and third-party specific cytotoxic T-cell precursor frequencies (CTLpf) in the peripheral blood of 15 heart transplant recipients. PBL samples were obtained at different time points before and after transplantation. Donor-specific CTLpf and third-party specific CTLpf were within the same range for all samples (1-1489/10(6) cells). The donor-specific CTLpf were not different between patients who had never had an acute rejection (AR) and patients who had an acute rejection as diagnosed by endomyocardial biopsy (EMB). No difference was observed between donor-specific CTLpf of samples taken on the day of transplantation and those obtained between 3 months and 3 years after transplantation. There was also no relationship between the donor-specific CTLpf in the PBL and the culturing success of lymphocytes from EMB taken at the same time. CTLpf were in the same range both when cultures could be propagated from the graft and when no cells grew out. We conclude that long term graft survival is possible in the presence of CTLpf in peripheral blood. PMID- 11271318 TI - Role of CTLA 4-Ig on induction of unresponsiveness to multiple minor alloantigens. AB - LPS blasts inhibited graft rejection in multiple minor incompatible strain combinations but not in presensitized animals or in major histoincompatible combinations. PMID- 11271319 TI - Injection of v-mos-transformed and irradiated macrophages leads to longlasting specific acceptance of MHC-allogeneic heart grafts and specific prolongation of skin graft survival in mice. AB - In vitro studies have revealed that the v-mos-transformed clone mos2 (mos2) of the murine macrophage cell line P388D1 (D1) (H-2d) is capable of inducing a state of specific unresponsiveness in MHC-allogeneic unprimed T cells. Here, we present data on the in vivo relevance of these findings. Male C57bl/6 mice (H-2b) were injected i.p. 6 times with 10(7) of the following irradiated cell types: D1, mos3, mos2, DBA/2-(H-2d) or C3H-(H-2k) spleen macrophages. DBA/2 and C3H skin or heart grafts were performed 10 days after the last injection. The normal rejection time for allogeneic skin was 7.5 days and for allogeneic hearts, was 12.8 days. After injection of D1 or mos3, DBA/2 skin grafts were rejected after 4.5 and 6.5 days, respectively, and the hearts, after 15.4 and 18.6 days, respectively. Third-partly C3H grafts were rejected normally (7.0 days). In contrast, injection of mos2 led to prolongation of DBA skin graft survival to 12.3 days. DBA/2 hearts were accepted for more than 160 days as revealed by heart beating. Again, C3H grafts were rejected normally (11.0 days). DBA/2 skin grafts on day 102 after heart grafting survived for 30 days, indicating hyporesponsiveness against these grafts. These results confirmed the in vitro findings. The mos2 cells obviously induced a state of specific unresponsiveness in otherwise unmanipulated recipients. However, the duration of this unresponsiveness induced by the injection of irradiated cells was dependent on the organ type. PMID- 11271320 TI - Split tolerance in chimeric GVH mice. AB - The i.v. inoculation of parental spleen cells into unirradiated adult F1 hybrid mice results in a graft-versus-host reaction (GVHR). In the strain combination B10D2 --> (B10.BRx B10.D2) F1, this reaction is associated with thymic injury and transient but profound cellular immune deficiency. We further analysed the immune status of these mice 60 days after GVHR induction. Phenotypic studies of spleen cells showed that these mice were repopulated with parental lymphocytes resulting in a high degree of chimerism (85%). At this time, the mice looked healthy and recovered a normal cytotoxic T cell response (CTL) against allogeneic cells. GVH chimeric splenocytes were unresponsive against F1 hybrid cells in mixed lymphocyte culture (MLC), but exhibited anti-F1 CTL reactivity. We also analysed the anti-F1 reactivity of these mice in vivo. GVH chimeric splenocytes were unable to induce GVHR after injection into a new F1 hybrid and F1 GVH mice specifically rejected F1 bone marrow (BM) cells after lethal irradiation. Grafting a neonatal parental thymus prevented the rejection of F1 BM cells and restored CTL alloreactivity. It is concluded that the chimeric state induced by GVHR is associated with a split tolerance and that a radiosensitive thymic dependent mechanism is involved in maintaining self-tolerance in these mice. PMID- 11271321 TI - Successive emergence of two CD8 subsets in primary CMV infection of allograft recipients. AB - Allograft recipients with cytomegalovirus (CMV) infection develop increased proportions of circulating CD8+ lymphocytes. A longitudinal study of 11 kidney and 5 liver allograft recipients with primary CMV infection but no other aetiological factor to explain graft dysfunction revealed selective imbalances in peripheral blood CD8+ T cell subsets. Initially, CMV viraemia was associated with elevated CD8+bright T cell numbers and T cell activation. Activation markers fell to normal when viral cultures became negative (before the end of the 1st month). During the 2nd-6th months, most (12/16) patients continued to have high CD8+ T cell counts (1050-2900 CD8+ cells/mm3), comprising an uncommon CD8+ T cell subset, as 45-73% of CD8+bright lymphocytes were CD3+ and TCRalphabeta+ but were not stained by anti-CD28, CD11b, CD16, CD56 and CD57 antibody. Unexpectedly, CD8+ CD57+ T cells, a hallmark of CMV infection, did not appear until the 2nd-6th months of primary CMV infection, and their numbers increased progressively thereafter. They became the predominant CD8+ T cell subset after about 6 months of infection and their persistence for several (up to 4) years was strongly correlated (r = 0.87) with expansion of CD8+ cells. Persistence of CD8 lymphocytosis was, thus, directly related to the rate of expansion of an uncommon CD8+ CD57- subset and its progressive replacement by CD8+ CD57+ T cells that were chronically elicited by CMV. PMID- 11271322 TI - Interactions between malignant tumor growth and allogeneic graft rejection in an experimental rat model. AB - We describe a combined tumor and simultaneous transplant model in rats in tended to investigate interactions between tumor growth and graft rejection. To study the influence of tumor growth on graft rejection. Novikoff hepatoma cells were injected subcutaneously into the back of Lewis rats. Eight days later, the grown solid tumor was resected, and allogeneic heart transplantation was performed. Four groups were formed, receiving 5-fluorouracil (5-FU), cyclosporin A (CsA), 5 FU + CsA, and placebo, respectively. In the corresponding groups, tumor injection was omitted. Graft survival was significantly prolonged when CsA was given 5-FU did not abrogate or augment CsA efficiency nor influence graft survival when given alone. In the corresponding control groups, graft survival was similar, thus excluding an immunomodulating effect of the prior tumor growth on graft survival. To study the reverse interaction of allogeneic graft on tumor growth, heart grafting and tumor cell injection were performed on the same day. In different groups, 5-FU, CsA, 5-FU + CsA, and placebo was given. For the control, no transplantation was carried out. The tumor was resected on the 8th postoperative day and examined by immunohistology. A slight decrease of tumor growth by 5-FU, but a marked increase by CsA were found, whereas the graft alone showed no immunomodulation. PMID- 11271323 TI - Cytokine gene expression profiles in human endomyocardial biopsy (EMB) derived lymphocyte cultures and in EMB tissue. AB - The reverse transcriptase polymerase chain reaction (RT-PCR) technique was used to analyse cytokine gene expression in relation to acute cardiac rejection. Expression of interleukins, IL-2, IL-4, IL-6 and IL-10 mRNA was studied in sequential endomyocardial biopsies (EMB) and in graft-infiltrating lymphocyte (GIL) cultures propagated from EMB taken after heart transplantation. The cytokine gene expression of GIL propagated from EMB taken during an episode of rejection and of immunological quiescence was comparable. In contrast, posttransplant EMB showed selective IL-2 gene expression when rejection was diagnosed. IL-4 mRNA was absent in pretransplant EMB but present in posttransplant EMB taken during periods of rejection and of immunological quiescence. Both IL-6 and IL-10 transcripts were found in pre- and posttransplant EMB. These findings confirmed that IL-2 is specifically involved in cardiac rejection, while IL-4 may play a role in immune responses leading to graft rejection or graft tolerance. PMID- 11271324 TI - Discrepancy between mRNA expression and production of IL-2 and IL-4 by cultured graft infiltrating cells propagated from endomyocardial biopsies. AB - We studied whether acute rejection correlated with the cytokine production pattern and mRNA expression of interleukin-2 (IL-2) and interleukin-4 (IL-4) in lymphocyte cultures derived from endomyocardial biopsies (EMB) that were stimulated with B cell lines of donor origin. Unstimulated biopsy cultures neither expressed mRNA nor produced IL-2 or IL-4. All stimulated biopsy cultures contained mRNA transcripts for IL-2 and IL-4. In contrast, we found different IL 2 and IL-4 production patterns. Within the first 90 days after heart transplantation (HTx), higher levels of IL-4 were measured in cultures derived from EMB with myocytolysis than in cultures from EMB without signs of myocytolysis. More than 90 days after HTx, this phenomenon was reversed and more IL-4 was produced in cultures derived from EMB without myocytolysis. These differences were not detected for IL-2 production. PMID- 11271325 TI - Inhibition of IL-2 synthesis by donor-specific suppressor T cells in a renal transplant recipient. AB - A study was conducted to elucidate the mechanism of donor-specific Mixed Lymphocyte Reaction (MLR and Cell Mediated Lymphotoxicity (CML) unresponsiveness in a renal transplant recipient with a long-term well-functioning kidney. The peripheral blood lymphocytes (PBL) of the recipient, who had not shown rejection since his transplantation 5 years previously, and those of his mother (donor), his father and two healthy third parties were examined. MLR, CML, semimicro MLR in a double chamber, interleukin-2 (IL-2) synthesis assay and limiting dilution assay were performed. This recipient showed donor-specific MLR and CML unresponsiveness. IL-2 assay showed that the PBL of the recipient produced less IL-2 against the donor than against the father and the third parties. The addition of exogenous recombinant IL-2 (rIL-2; Takeda Co.) to the priming MLR caused a recovery of CML against the donor. A limiting dilution assay indicated that cytotoxic T cell precursor (CTLp) frequencies against the donor and father did not differ. The suppressor assay in a double chamber indicated that the PBL of the recipient stimulated by the donor PBL had a non-specific suppressive effect on MLR, CML and IL-2 synthesis of the PBL across the Major Histocompatibility Complex (MHC) barrier. This suppressive effect was abolished by OKT3 or OKT8 monoclonal antibody and complement. Thus, the recipient had donor specific suppressor T cells that produced a humoral non-specific suppressive factor only when stimulated by the donor PBL, and this factor suppressed MLR and CML by inhibiting IL-2 synthesis of the PBL. PMID- 11271326 TI - Does concommitant splenectomy raise the mortality of liver transplant recipients? AB - Within a 17-month period, 130 orthotopic liver transplantations were performed in our hospital. Nine of these were retransplantations and were not included in our analysis. In the remaining 121 patients, splenectomy was performed in 34 patients, either synchronously with the transplant procedure (27 patients) or in the postoperative period (7 patients). Indications for splenectomy were lienalis steal syndrome in 15 patients and hypersplenism in 15 cases. The number of rejection episodes was fairly equal in both groups (splenectomized vs. non splenectomized, 61.7% vs. 63.9%, respectively). There was a marked difference in the frequency of infectious episodes (61.7% vs. 25.3%) that resulted in a decreased survival rate (77.5% vs. 95.4%) for splenectomized patients. Therefore, we recommend splenectomy only for very selected patients and investigate the banding of the splenic artery as an alternative. PMID- 11271327 TI - Expression of cell-matrix molecules and integrin receptors in human liver grafts during chronic rejection. AB - The inflammatory response of immune cells to target cells and cell-matrix molecules is regulated by several receptor-ligand molecules. As fibrosis develops in ongoing chronic rejection after liver transplantation, it is of interest to analyze patterns of integrin receptors and cell-matrix molecules in order to study the relation between immune cells and the stromal and parenchymal cells. In the present study, we demonstrated the expression of these molecules in chronic rejected human liver grafts using immunohistochemical techniques. The results showed a differential expression and induction of integrin receptors and cell matrix molecules on resident liver cells, especially on sinusoids, reflecting a state of chronic inflammation and a specific interaction between integrin receptors and cell-matrix molecules. The patterns of induced integrin receptors on graft-infiltrating cells was closely related to the local production of cell matrix molecules and reflected the final sequence of a stepwise progress of the inflammatory reaction. PMID- 11271329 TI - Xenogeneic ex vivo hemoperfusion of rhesus monkey livers with human blood. AB - In order to copy the clinical situation of concordant xenotransplantation, Rhesus Monkey livers were hemoperfused with human blood. Changes of immunological (TNFalpha, IL-1beta, IL-2, IL-2R, IL-6, IFNgamma, TXB2, 6kPGF1alpha, sICAM-1, sELAM-1, sHLA-I-Ag) and pathophysiological (GOT, GPT, LDH, CK) parameters were followed. Our experiment proves that all phenomena start in the first hour of xenogeneic blood circulation. Xenogeneic rejection in our concordant system is surprisingly severe. Preformed natural antibodies only cannot be the reason of such a damage. We think that beside other important immunological mechanisms, humoral mediators play a considerable role at the beginning of a xenogeneic rejection. PMID- 11271328 TI - FK 506 primary immunosuppression following emergency liver transplantation for fulminant hepatic failure. European FK 506 Study Liver Group. AB - The efficacy and safety of an FK 506-compared to a cyclosporin A based immunosuppression regimen was examined in liver recipients who underwent transplantation for fulminant hepatic failure in the European FK 506 liver study. A consistent trend towards improved patient and graft survival noted in the FK 506-treated patients was apparent from the first postoperative week (e. g. patient survival: day 7, 95.5% vs. 82.1% and month 6, 72.7% vs. 60.7%). Acute (in particular intractable) rejection was less frequent in the FK 506 group (e. g. cumulative intractable rejection rate at 6 months, 6.2% vs. 22.6%). In a single centre (Kings College Hospital), 17 patients were studied in more detail. The FK 506 treatment group had improved graft function, lower steroid requirements and episodes of infection. Accompanying these benefits, apache 111 and TISS scores were lower in this group in the early posttransplant period. Intensive care discharge was earlier and both treatment groups experienced similar toxicity. PMID- 11271330 TI - Production of proinflammatory cytokines and adhesion molecules in ex-vivo xenogeneic kidney perfusion. AB - Xenogeneic transplantation of solid organs is limited due to hyperacute rejection. In concordant systems, the mechanisms of rejection can be studied due to cross-reactivity of mediators with anti-human monoclonal antibodies. The aim of this study was to obtain information about the kinetics of proinflammatory cytokines and production of soluble adhesion molecules in the acute phase of reperfusion, eight kidneys from rhesus monkeys were perfused ex-vivo with human blood (group B/0) for 1 hour in a closed system. Blood levels of IL-1b, IL-6, TNFalpha, soluble ICAM, and E-selectin were measured using an ELISA technique under steady-state conditions. Cytokine levels rose significantly within the 60 min interval (IL-1b, 6.1 +/- 2.6-161.1 +/- 98.5 pg/ml; IL-6, 30.2 +/- 7.7-274.2 +/- 75.8 pg/ml; TNFalpha, 544.2 +/- 363.6-1651.0 +/- 25.7 pg/ml; P < 0.05). Immediately after the beginning of reperfusion, soluble ICAM-1 and selectin levels were abnormally high and rose constantly throughout the observation period, reaching significance at 60 min. High levels of proinflammatory cytokines may lead to an induction of adhesion molecules, thus, upregulating the leukocyte endothelial interaction in a complement-independent mechanism. Specific pretreatment with monoclonal antibodies against ICAM-1, LFA-1, or other soluble mediators may be useful in down-regulating hyperacute rejection in trans-species transplantation. PMID- 11271331 TI - Prolongation of discordant renal xenograft survival by depletion of complement. Comparative effects of systemically administered cobra venom factor and soluble complement receptor type 1 in a guinea-pig to rat model. AB - There is an increasing body of evidence to suggest that inhibition of complement activation may be a valuable approach to avert hyperacute rejection. In our study, the guinea-pig to rat discordant kidney xenograft model was adapted for the investigation of renal transplant function and an attempt was made to delay the hyperacute rejection using systemically administered cobra venom factor (CVF) and soluble complement receptor type 1 (sCR1). The saline-treated control recipients experienced a rapid transplant rejection with a xenograft survival averaging 10.5 +/- 2.1 min. Administration of a single 60 U/kg i.v. bolus of CVF significantly prolonged renal graft survival to 20.4 +/- 2.5 h, and by a single bolus of sCR1 (50 mg/kg) a prolongation of graft survival to 18.8 +/- 2.3 h was achieved. The grafts functioned only over periods of 2.5 +/- 0.3 and 2.3 +/- 0.2 h, respectively. No complications of sCR1 were noted. We concluded that complement inhibition by sCR1 may be an important component in the therapeutic approach aiming at the prevention of hyperacute rejection in human organ transplantation. PMID- 11271332 TI - Role of CD4+ T cells in the rat to mouse cardiac xenotransplantation. AB - T cell subsets involved in rejection of xenografts were analyzed using a rat to mouse cardiac xenotransplant model. Proliferating response and interleulin-2 (IL 2) production in recipients' spleen cells were almost completely abrogated by elimination of L3T4+ T cells, but not by elimination of Lyt2.1+ T cells. Cytotoxic T lymphocyte (CTL) activities were mediated by both L3T4+ and Lyt2.1+ T cells with the help of IL-2-producing L3T4+ T cells. Administration of anti-L3T4 monoclonal antibody (mAb) into recipient mice resulted in a significant prolongation of graft survival (mean graft survival was 29.2 days). Moreover, anti-L3T4 mAb treatment plus thymectomy led to indefinite graft survival. Anti rat endothelial cell (EC) antibody production in the grafted mice was remarkably suppressed by anti-L3T4 mAb treatment. In contrast, Lyt2.1 mAb treatment did not prolong the graft survival and did not suppress anti-EC antibody production. These results indicated the absolute requirement of L3T4+ T cells in the rejection of rat to mouse cardiac xenografts. PMID- 11271334 TI - Living kidney donation in Europe: legal and ethical perspectives--the EUROTOLD Project. AB - The demand for organ replacement by transplantation continues to outstrip supply, leading to unnecessary morbidity and health care costs. "Space capacity" (i. e. cadaver organs not currently harvested for transplant) has been tackled within Western Europe in particular by many strategies--medical, social, educational and legal--but with varying degrees of success. Despite this, the use of living donors has not been fully exploided in many European countries to fill this gap. The total picture is, however, one of marked differences between countries and between centres within countries. In Turkey and Greece, living donors generally account for 60-90% of all renal donors. Countries within Scandinavia also have a high rate of living donor use, especially Norway. By contrast the percentage is far more modest, for example in Spain, Ireland, France and Germany. In the United Kingdom the rate is only 6% with a range of between 0 and 20%. Sources of living donors also show substantial variations between countries, notably the extent to which non-genetically related donors are used. A European Commission sponsored study has been established to acquire a broad understanding of the interaction of ethical values, cultural traditions and social customs on willingness to donate. It will also aim to assess the effect of national laws on professional attitudes to living donor transplantation. This is a collaborative project between the University Department of Surgery, Leicester General Hospital, the Department of Law, De Montfort University and the Department of Mathematics, Newcastle University. Transplant units throughout Europe, including France, Germany, Switzerland, Norway, The Netherlands, Eire and Turkey are collaborating to exchange information and views on living donor transplantation. PMID- 11271333 TI - Primary nonfunction of islet xenografts: the role of macrophages. AB - Primary nonfunction (PNF) of hamster islets occurs when they are transplanted under the kidney capsule of diabetic Lewis rat recipients. PNF is absent if the islets are grafted into the liver. Previous results have indicated that macrophages might be involved in the phenomenon of PNF. To test this hypothesis, two procedures affecting macrophages were employed. First, recipients were pretreated with liposomes containing dichloromethylene disphosphonate (L-MDP), killing macrophages upon phagocytosis. Second, recipients were pretreated with the arginine analogue, L-NA, in order to inhibit the synthesis of nitric oxide (NO), a molecule believed to mediate beta cell dysfunction. Injection of L-MDP into the peritoneal cavity had no effect on PNF. However, co-transplantation of L MDP with hamster islets under the kidney capsule reduced PNF significantly. Eradication of Kupffer cells with L-MDP did not prolong the survival of islets transplanted into the liver, indicating that acute xenogeneic rejection in this model is not mediated by macrophages. A single injection of L-NA, 3 h before transplantation resulted in complete annihilation of PNF, all recipients became normoglycemic within 1 day and remained so for 1.4+/-0.5 days. These results confirmed the finding that macrophages and NO play a crucial role in PNF of islet grafts. PMID- 11271335 TI - Reasons for 50% reduction in the number of organ donors within 2 years--opinion poll amongst all ICUs of a transplant centre. AB - To detect the reasons for a massive decrease in the annual number of organ donors and as a means of evaluating the effectiveness of our information programme, a questionnaire was designed and sent to all intensive care units (ICUs) in our catchment area. We wished to obtain information about medical, organizational and capacity problems and negative occurrences that had happend during past retrievals. Although 60% of the answers we reiceved (87% feedback rate) mentioned the additional workload involved in treating an organ donor (and 88% had serious problems because of the shortage of nurses), less than 16% remembered a "lost" donor because of capacity problems. Eighty-six percent recognized our efforts to support them in any respect and were satisfied with the amount of "service" provided by the transplantation (TX) centre. About 45% remembered negative occurrences. More than 85% of all replies asked for more and continuing information related to organ donation and transplantation. We think that the key to a successful TX programme is a system of active care for the ICU staff in all peripheral hospitals; repeated mailing of updated information brochures, annual lectures about new developments, letters of thanks after each reported donor (including information on the fate of the organs), visiting donor ICU's accompanied by successfully transplanted recipients, etc.... The downwards trend of donor rates in our area clearly shows that it takes more than a stable legal situation to ensure the necessary amount of donor organs, even a very successful TX centre has to work hard to maintain a certain standard of knowledge, information and motivation amongst the staff of the peripheral hospitals. Moreover, the high turnover rate of ICU personnel requires a steady "flow of information" and cooperation between the "transplant people" and their coworkers outside to guarantee a permanent state of awareness concerning organ donation and transplantation. In fact, awareness seems to be the key issue: the activity of sending out the questionnaires was enough to raise the number of reported donors from 72 (estimated in July) to 96 (31 December 1992). PMID- 11271336 TI - Influence of donor criteria on postoperative graft function after orthotopic liver transplantation. AB - Evaluation of graft quality remains a major problem in liver transplantation. The aim of this retrospective analysis was to examine the impact of donor criteria on postoperative graft function. Between June 1986 and September 1993 324 liver transplantations were performed at our institution. Criteria for exclusion from analysis were postoperative thrombosis of graft vessels, retransplantation, death prior to the 5th postoperative day or missing donor criteria. For the eligible 255 transplantations the impact of the following donor criteria were examined: age (range 1-62 years, median 28 years), size/body weigt index, duration of intensive care, cause of death, circulatory condition, need for vasopressive support and liver function tests (bilirubin, GOT, GPT, GGT, LDH, ALP, prothrombin time (PT), creatinine, sodium). The following intraoperative factors were also assessed: type of protective solution, cold ischaemic time (CIT), anhepatic period and blood transfusions. Graft function during the first 5 postoperative days was categorized into four groups: (1) good function (GOT max < 1000 U/l, spontaneous PT > 50%, bile production > 100 ml/day); (2) fair function (GOT 1000 2500 U/l, clotting factor support < 2 days, bile < 100 ml/day); (3) poor function (GOT > 2500 U/l, clotting factor support > 2 days, bile < 20 ml/day); (4) primary non-function (retransplantation required within 7 days). A univariate analysis revealed duration of intensive care (P = 0.001), circulatory condition (P = 0.005), anhepatic period (P = 0.0004), blood transfusions (P = 0.03) and CIT (P = 0.039) as significant risk factors for postoperative graft function. Entering these factors in a multivariate regression model we identified creatinine (P = 0.007), duration of intensive care (P = 0.009) and the size/body weight index (P = 0.03) as donor-related factors of high significance. Analysis of the intraoperative data revealed the anhepatic period as the factor of highest significance (P = 0.0004) together with CIT (P = 0.02) and intraoperative blood transfusions (P = 0.008). A doubling of the number of days of intensive care resulted in a threefold increased risk of postoperative graft failure. Prolonged intensive care is a variable representing multiple risk factors. Accepting donors with a longer history of hypotension or who show signs such as elevated creatinine should be carefully considered. In patients with expected surgical difficulties resulting in an extended anhepatic period and a higher blood loss, transplantation of organs retrieved from donors with a long duration of intensive care and a long CIT should be avoided. PMID- 11271337 TI - Life quality of patients undergoing liver transplantation. AB - The aim of this study was to assess the life quality of a group of patients who had undergone liver transplantation using (1) a psychological test to evaluate family relations, work activity, emotional state and social relationships; (2) the quantification of hospital dependence and degree of fitness for work. Included in the study were 32 patients using the following criteria: diagnosis of hepatic cirrhosis and minimum posttransplant follow-up of 6 months. The average age of the study population was 44.8 +/- 10.5 years; there were 23 males and 9 females, with an average follow-up of 15 months. The psychological test used was the Quality of Life Scale (QLS) which consists of 21 items each scoring from 1 to 6 points. The questionnaire was completed before transplantation by all the patients, and after transplantation by 32 patients at 6 months, 20 at 12 months and 12 at 24 months. Hospital dependence was evaluated by the number of admissions and number of days per admission. Lastly, we compared fitness for work before transplantation and at 1 and 2 years after transplantation. The QLS test showed a post-transplant improvement in the four aspects assessed, particularly in the personal aspects (emotions and family) (P < 0.001). Hospital dependence following liver transplantation decreased significantly compared with the pretransplant situation (P < 0.01). Finally, the post-transplant percentage of unfitness for work decreased with time, reaching a significant differences 2 years after transplantation (P < 0.05). PMID- 11271338 TI - Corticosteroids and concomitant medication in the European multicentre study of FK 506 and cyclosporin in primary liver transplantation. AB - The steroid-sparing effect and the use of concomitant medication during the treatment of liver transplant patients with the novel immunosuppressant FK 506 were evaluated within the European multicentre, randomized, parallel-group study in liver transplantation. Patients undergoing primary liver transplantation were randomized to treatment with FK 506 (n = 267) or with a cyclosporin-based immunosuppressive regimen (n = 273). The total cumulative steroid usage was significantly reduced in the FK 506 treatment group, which is likely to have resulted from the lower incidence of acute rejection in these patients. The number of patients receiving antidiabetic, diuretic and antihypertensive therapy did not differ between the two treatment groups, even though the incidence of diabetes mellitus and oliguria was significantly higher in the FK 506 group. It can, therefore, be assumed that in a number of such cases the severity of these events was very mild necessitating no specific therapy. PMID- 11271339 TI - FK 506 rescue therapy for intractable liver allograft rejection. AB - Intractable liver allograft rejection remains an important cause of graft loss. In this present study, we evaluated the role of oral FK 506 in 30 rejection episodes resistant to conventional cyclosporin-based triple immunosuppression in a series of 28 patients. Rejection was reversed in 11 (91.7%) of 12 patients for intractable acute rejection and in 10 (58.8%) of 17 patients for chronic rejection. A progressive decline in serum bilirubin was observed within 14 days in those successfully salvaged and a serum bilirubin of less than 200 micromol/l at the time of FK 506 conversion in the chronic rejection subgroup was found to be good predictor of response (specificity 100%, sensitivity 60%). New onset diabetes mellitus (29%) and reversible renal impairment (32%) were the commonest adverse events observed. Eleven (52%) of the responding patients successfully discontinued corticosteroid medication and are currently on FK 506 monotherapy. FK 506 therapy has a significant impact in the control of both intractable acute and chronic allograft rejection with an acceptable toxicity profile. PMID- 11271340 TI - Comparison of short and long-term renal function in liver transplant patients receiving cyclosporin or FK 506. AB - Long-term renal function was compared in 49 liver recipients [25 patients received cyclosporin (CyA) and 24 patients received FK 506] followed for a period of 1 year. Creatinine (CR) and glomerular filtration rate (GFR) pretransplantation (pre-Tx) and at 1, 3, 5, and 12 months post-Tx were recorded, as well as incidences of hyperkalemia, post-Tx hypertension, and insulin dependent diabetes mellitus (IDDM) in the two groups. At 1 year post-Tx, the mean Cr had risen from baseline by 56% and 60% in the FK and CyA groups, respectively; the mean GFR had dropped by 32% in FK patients and by 27% in CyA patients. Acute nephrotoxicity occurred in 7/25 CyA patients (2/7 required dialysis) and 9/26 FK patients (7/9 required dialysis; 2/7 were switched to CyA). None remained on dialysis at 3 months. Renal insufficiency persisted at 1 year in 7/16 patients with early toxicity (CyA, 4; FK, 3) and in 3 of the remaining 36 pts (P < 0.001). Hyperkalemia occurred in 4/25 CyA, and in 12/24 FK patients (P < 0.025), post-Tx hypertension occurred in 15 CyA, and 7 FK patients (P < 0.05), and IDDM occurred in 4 CyA and 7 FK patients (P = ns). FK 506 and CyA, thus, exerted similar chronic renal effects. Although acute renal insufficiency improved upon dose reduction, renal impairment was permanent in some cases. PMID- 11271341 TI - Long-term use of FK 506 in living related liver transplantation. AB - FK 506 (Tacrolimus) was used with steroids to treat 61 pediatric patients who received living related partial liver transplantation. Fifty-two recipients survived and 9 died between 6 months and 3 years after transplantation. In the surviving patients, oral doses of Tacrolimus were tapered from 0.298 +/- 0.277 mg/kg daily at 1 month after transplantation to 0.078 +/- 0.054 at 24 months after transplantation. The 12 h trough levels of Tacrolimus were 12.6 +/- 7.1 ng/ml and 4.1 +/- 2.4 at 1 and 24 months after transplantation, respectively. The percentage of recipients free from steroids was 77%, 97%, and 94% at 6, 12, and 24 months after transplantation, respectively. Liver allograft rejection was encountered in seven recipients, five of whom were treated by steroid pulse therapy and a dose increase of Tacrolimus; the remaining two required OKT3. However, there was no episode of rejection that required retransplantation. Infectious complications encountered in 34 patients included 12 bacterial, 3 fungal, and 19 viral infections. Two recipients died one of fungal pneumonia and one of Epstein-Barr virus-associated lymphoproliferative disorder. Regarding adverse reactions of Tacrolimus, hypertension was observed in 28 patients, diabetes mellitus in 3, pancreatitis in 3, convulsion in 1, tremor in 12, itching in 5, and pigmentation in the oral mucosa in 2. Slightly increased values of creatinine were observed in most of the patients; however, an abnormal increase of serum of serum creatinine (> 1.0 mg/dl) was confined to the complicated cases. Improvement of somatic growth was observed in 21 patients (62%) and 13 (75%) at 12 and 24 months after transplantation, respectively. The long-term use of Tacrolimus is highly effective in terms of its immunosuppressive potential and reduced adverse reaction. Steady growth development can be expected in pediatric recipients free from steroids. PMID- 11271342 TI - Steroid withdrawal 3 months after liver transplantation--does FK 506 confer any advantage over cyclosporin? AB - Eighty-one liver recipients were randomised to FK 506 or cyclosporin (CyA) and azathioprine (AzA), both in combination with steroids. Twenty-seven FK 506 and 29 CyA/AzA patients continued in the trial 3 months after transplantation. Steroids were ceased in 23 (85%) FK 506 patients and in 27 (93%) CyA patients. After steroid withdrawal, 2 FK 506 and 4 CyA patients were excluded from the study, all for reasons other than rejection. The median follow-up was 16 months for the FK 506, and 19 months for CyA group. There were no acute rejection episodes or graft losses in the FK 506 group. None of the CyA patients lost their graft but three (13%) had episodes of acute rejection requiring steroids to be recommenced in two cases. There was no evidence of chronic rejection in any of the annual review biopsies in either group. Our results suggested no advantage of FK 506 over CyA in its steroid-sparing effect. PMID- 11271343 TI - Prospective randomized trial of steroid withdrawal in liver transplant patients: preliminary report. AB - Although steroid withdrawal has been successfully performed in heart and kidney transplant recipients, no controlled studies of SW have been carried out in liver transplant patients. To evaluate this possibility a prospective controlled study was carried out in 46 liver transplant recipients operated on after may 1991. They all received a sequential quadruple immunosuppression consisting of 3 mg/kg antithymocyte globulins (RATG) for the first 5 postoperative days, cyclosporin A (starting from day 3-5 and maintaining parenteral whole-blood trough levels at 200-300 ng/ml during the first month and at 150-250 thereafter), azathioprine (1 mg/kg per day for the first month) and steroids. Prednisone was started at a dose of 200 mg per day 1 and then tapered to 20 mg/day over the first posteroperative week; this dose was maintained until day 90 when the patients were randomly allocated either to long-term steroid therapy (0.1 mg/kg per day) or to steroid withdrawal. Minimum follow-up after randomization was 6 months (6-27 months). Liver biochemistry was checked at regular intervals throughout the follow-up period. Liver biopsies were performed whenever clinically indicated and also in the first 19 patients during readmission for annual review. The incidence ot acute and chronic rejection 90 days from liver transplantation was 2.5% in patients maintained on long-term therapy. No patient in the steroid-withdrawal group had experienced either an acute or a chronic rejection episode so far. Steroid-related complications did not differ significantly between the two groups. The most recent interim analysis showed that steroid withdrawal is a safe undertaking in liver transplant recipients and may be successfully accomplished in almost all patients. PMID- 11271344 TI - Simultaneous liver-renal grafting for polycystic disease. AB - Our purpose was to describe the first in our country and fourth worldwide simultaneous liver and renal grafting for polycystic disease. PMID- 11271345 TI - Adverse impact of high panel-reactive antibody (PRA) and positive cytotoxic crossmatch in liver transplantation. AB - Of 91 liver transplants (LTX) performed from October 1988 to December 1992, 13 (14.2%) of the patients received a liver from a lymphocytotoxic-positive crossmatch (CM) donor. Severe early rejection resulting in graft floss occurred in seven positive CM patients. Three of the remaining positive CM patients suffered several rejection episodes leading to chronic rejection and FK 506 was required as rescue treatment. A significant difference in mean panel-reactive antibody (PRA) of 8.6% and 56.9% was found in negative and positive CM patients, respectively (P = 0.012). A higher mean PRA (67.7%) was found in positive CM patients with rejection compared with positive CM patients without rejection (PRA 38%). Overall graft and patient survival were 31.9% and 35% in positive CM patients compared with 57.0% and 61.9% in negative CM patients. These differences were statistically significant (P = 0.023). In our experience the risk of developing severe acute rejection with graft failure and chronic rejection is related to PRA > 60% and positive CM. We recommend that in patients with PRA > 60%, the result of CM should be awaited before proceeding to LTX. PMID- 11271346 TI - The diagnostic value of GM-CSF and IL-6 determinations in patients after renal transplantation. AB - Cytokines are released by graft-infiltrating cells during cellular rejection. We studied the release of GMCSF and IL-6 and their prognostic significance in predicting rejection. Sequential measurements were made in serum and urine samples with an IL-6 specific cell line and a GMCSF ELISA. Biopsy tissue was snap frozen and examined with immunohistochemical methods. The IL-6 values for normal controls (CTR) and stable transplant patients (PTS) were 5-10 pg/ml in serum and 0-2.5 pg/ml in urine. In 51 biopsy-proven rejections (AR), serum IL-6 values at least doubled in 15 (sensitivity 29%, specificity 87%; 19 +/- 7 vs. 7 +/- 2 pg/ml; P = ns). In urine an increase was observed in 29 of 36 AR (sensitivity 80%, specificity 75%; 92 +/- 34 vs. 5 +/- 1 pg/ml; P < 0.05). After treatment, IL 6 decreased in urine in 26/29 PTS to 7 +/- 2 pg/ml (P < 0.05). In three PTS, rejection persisted, as did their elevated IL-6 urine values. In PTS with urinary tract infections, IL-6 increased in the serum of 13/19 and in the urine of 10/12. GMCSF in serum was not influenced by rejection; however, urine values increased in 22/33 AR (sensitivity 67%, specificity 96%; 22 +/- 5 vs. 4.8 +/- 0.3 pg/ml; P < 0.05). These values decreased (5 +/- 0.3; P < 0.05) after treatment. During infection, increased urinary GMCSF levels were observed in 2/9 PTS. Further analysis revealed a better correlation between elevated cytokine levels and rejection episodes in the early posttransplant period. In kidneys with acute rejection, IL-6 was found in the interstitium of all PTS tested. CTR tissue was negative. In PTS GMCSF was found in arterioles and in infiltrate; however, control tissue also showed some staining. Cytokine labeling in tissue could not be correlated with serum or urine values. We concluded: (1) serum IL-6 and GMCSF are of no value in rejection; (2) in urine, they reflect rejection, especially in the early posttransplant period; however, infection confounds the results; (3) IL 6 staining in tissue may be helpful, but requires more study. PMID- 11271347 TI - Comparison of modified one- and two-layer microsurgical vasovasostomy. PMID- 11271348 TI - Bilateral germ cell cancer of the testis: a report of 11 patients with a long term follow-up. PMID- 11271349 TI - Comparison of single and multiple sessions of percutaneous sclerotherapy of simple renal cysts. PMID- 11271350 TI - One year's clinical experience with unenhanced spiral computed tomography for the assessment of acute loin pain suggestive of renal colic. PMID- 11271351 TI - Anomalies of external urethral meatus in girls with non-neurogenic bladder sphincter dysfunction. PMID- 11271352 TI - [Foreign blood transfusion--ways out of a dilemma]. PMID- 11271353 TI - [Immunosuppresson from transfusion of blood and blood products intumor surgery]. PMID- 11271354 TI - [Is there a "critical hematocrit"?]. PMID- 11271355 TI - [Bacterial infection risks of allogenic and autologous bloodcomponents]. PMID- 11271356 TI - [Erythrocyte quality following mechanical autotransfusion with collected wound drainage blood]. PMID- 11271357 TI - [Wound blood irradiation in oncologic procedures--safe andeffective?]. PMID- 11271358 TI - [Autologous hemotherapy--procedures--a cost-benefit analysis]. PMID- 11271359 TI - [Does controlled hypotension have a place in foreign blood transfusion sparing procedures]. PMID- 11271360 TI - [Synthetic oxygen carriers as an alternative to foreign blood transfusion]. PMID- 11271361 TI - [The effectiveness of augmented acute normovolemic hemodilution (A-ANH)]. PMID- 11271362 TI - [Erythropoietin--an alternative to foreign blood transfusion]. PMID- 11271363 TI - [Storage dependent effects on the oxygen tranport capacity of erythrocyte concentrates]. PMID- 11271364 TI - Electron microscopic investigation of rat brain after brief cardiac arrest. AB - Rats were submitted to 10-min cardiac arrest, followed by resuscitation and survival for 1 day, 3 days or 1 week. Five regions of interest (CA1 and CA3 sector of hippocampus, dentate gyrus, reticular nucleus of thalamus and parietal cortex) where studied by light and electron microscopy at each of the survival times, and compared with non-ischemic control rats. Cell counts revealed delayed neuronal loss of about 30% after 3 days in both CA1 and CA3 sectors. Ischemic cell changes consisting of cytoplasmic condensation and nuclear pyknosis appeared in these regions on day 7 and --to a lesser degree-- also affected dentate gyrus, the reticular nucleus of thalamus and cerebral cortex. Ultrastructural alterations were evaluated using an ultrastructural injury catalogue. In all brain regions similar, although quantitatively differently expressed, changes occurred except ribosomal disaggregation, which was restricted to neurons of hippocampal CA1 sector on the first day after cardiac arrest. Progressive alterations included swelling of mitochondria and endoplasmic reticulum, which was most pronounced in CA1 and CA3 sectors of hippocampus, as well as chromatin aggregation and alterations of neuronal volume, which affected mainly the granule cells of dentate gyrus. Other alterations, such as osmiophilic inclusions or the formation of nuclear pore complexes, were transient with a maximum on the first day after cardiac arrest. Treatment with the free-radical scavenger alpha-phenyl N-tert-butyl nitrone (PBN) suppressed the formation of nuclear pores but otherwise did not markedly change the morphological outcome. In comparison to previous studies of global brain ischemia induced by arterial inflow occlusion of the same duration, the present data demonstrate remarkable preservation of tissue integrity in CA1 sector but also distinct changes in brain regions considered to be resistant to ischemic injury. Morphological alterations of brain after cardiac arrest do not follow the established pattern of selective vulnerability. PMID- 11271365 TI - Dynamic changes of magnetic resonance imaging abnormalities in relation to inflammation and glial responses after photothrombotic cerebral infarction in the rat brain. AB - This investigation analyzed the potential of high-resolution magnetic resonance imaging (MRI) at a field strength of 7T to depict leukocyte infiltration and glial responses after focal cerebral ischemia induced by photothrombotic occlusion of cerebral microvessels. For this purpose we superimposed multiparametric MRI (apparent diffusion coefficient, T2, perfusion-weighted, and gadolinium-DTPA-enhanced T1-weighted imaging) on tissue sections stained for phagocytes and astrocytes and, moreover, assessed the regional distribution of tissue pH and ATP content by invasive biochemical methods. Comparing the histological data with the various MRI parameters, high-resolution MRI did not allow a spatial discrimination between distinct areas of phagocyte accumulation or astroglial scar formation, based on image contrast or even quantitative parameter value differences. However, MRI parameters underwent characteristic changes and differentiated distinct stages of tissue remodeling between days 3 and 14 after photothrombosis. Low apparent diffusion coefficient (ADC) and high T2 values indicated an early stage (3 days) with necrosis and beginning glial activation. Normal ADC and reduced T2 elevation characterized an infarct with advanced glial activation and infiltration of hematogenous cells at 7 days after photothrombosis. Heterogeneous ADC together with T2 elevation reflected a late infarct stage (14 days) when pseudocystic degeneration and scar formation had occurred. PMID- 11271366 TI - Retrograde axonal transport of bismuth: an autometallographic study. AB - Bismuth subnitrate was injected into the triceps surae muscle of 3-month-old male Wistar rats. Sections of lumbar spinal cord (L4-L6) and corresponding dorsal root ganglia were developed by autometallography (AMG) to trace possible bismuth in neuronal somata resulting from retrograde axonal transport. At 3 days after treatment bismuth clusters could be traced by AMG in spinal cord motor neurons and in dorsal root ganglion cells ipsilateral to the injection site. Retrograde transport of bismuth could be avoided by ligation or intraneuronal injection of cholchicine into the sciatic nerve. At the ultrastructural level bismuth was found to be located exclusively in lysosome-like organelles in motor and sensory neuronal somata projecting to the injection site. The present study shows that bismuth is transported retrogradely in both sensory and motor axons if their terminals are exposed to bismuth ions. PMID- 11271367 TI - Pathological findings in the x-linked form of Charcot-Marie-Tooth disease: a morphometric and ultrastructural analysis. AB - Mutations in the connexin 32 gene (Cx 32) are associated with the x-linked form of Charcot-Marie-Tooth disease (CMTX) and segregate with a CMT 1 phenotype. The gap junction protein Cx 32 is expressed in myelinating Schwann cells and has been localized to regions of non-compacted cytoplasm in paranodes and in Schmidt Lanterman incisures. Mutant Cx 32 myelin proteins are predicted to impair Schwann cell functions. We have studied the resulting pathology in motor and sensory nerves from the probands of 13 CMTX kindreds with precisely defined genotype. This report provides a detailed descriptive and morphometric analysis of 14 CMTX nerve biopsy samples, taken at various stages in the development of the neuropathy and studied by light and electron microscopic examination. Findings indicated unusually prominent changes in paranodal myelin with resulting widened nodes of Ranvier, but with segmental demyelination being less common. In parallel early axonal cytoskeletal abnormalities were noted, which were followed later by axonal atrophy, degeneration and loss of myelinated nerve fibers, occurring in a length-dependent fashion. Regenerative sprouting was also unusually prominent. Ultrastructural abnormalities included a frequent dilatation of the adaxonal spaces, prominence of the adaxonal Schwann cell cytoplasm and widening of the Schmidt-Lanterman incisures. We conclude that mutations in Cx 32 gap junction protein lead to a compromise of Schwann cell functions and to impaired Schwann cell-axon interactions with subsequent pathology in both myelin and axons. PMID- 11271368 TI - Involvement of the cerebral cortex and autonomic ganglia in Machado-Joseph disease. AB - Machado-Joseph disease (MJD) is an autosomal dominant neurodegenerative disorder caused by expansion of a polyglutamine tract in the disease protein. In this study of brains from four autopsied MJD patients, we have demonstrated immunohistochemically that expanded polyglutamine stretches largely accumulate as inclusions in neuronal nuclei, and less frequently are distributed in the nucleoplasm in a diffuse pattern. These nuclear abnormalities involved many neurons covering a wide range of central and peripheral nervous system regions, including the cerebral cortex, thalamus and autonomic ganglia that have been categorized previously as spared regions by conventional pathological studies. These lesions, newly recognized by polyglutamine immunohistochemistry, may be responsible for the cerebral cortical dysfunctions or autonomic abnormalities pointed out in MJD patients by the recent clinical and neuroradiological studies. PMID- 11271369 TI - Neuropathology of progressive cognitive decline in chronically hypertensive rhesus monkeys. AB - Hypertension is an identified major risk factor for cerebrovascular disease, which is second only to Alzheimer's disease as a cause of dementia in the elderly. In addition, hypertension has been associated with a more subtle, progressive decline in cognitive function for which the neuropathology is not well understood. The present study was undertaken to explore this relationship in an experimental, nonhuman primate model, with hypertension produced by a coarctation of the thoracic aorta. Since prior studies with this model have shown a progressive decline in memory function, similar to that seen in human hypertension, as well as scattered microinfarcts in the cerebral white and gray matter, this study was designed to explore the relationship between these two. In addition to microinfarcts, the hypertensive monkeys with the highest arterial blood pressure also showed minute areas of focal gliosis without infarction. The number of these focal lesions showed a significant correlation with the severity of the hypertension, but not with the behavioral deficit. For four of these hypertensive monkeys, immunostaining demonstrated a pervasive, widespread activation of microglial cells and astroglial cells in the white matter as well as evidence of leaks in the blood-brain barrier, providing a more logical substrate for the cognitive decline. PMID- 11271370 TI - Up-regulation of cyclooxygenase-2 in inflammatory demyelinating neuropathy. AB - To clarify the role of prostaglandins in peripheral nerve demyelination, we examined the expression of cyclooxygenase-2 (COX-2) using selected nerve specimens from patients with chronic inflammatory demyelinating polyneuropathy. COX-2 protein was up-regulated in macrophages causing active demyelination. In situ hybridization revealed that COX-2 mRNA signals were strongly expressed on macrophages adhering to the demyelinating nerve fibers at the endoneurium. This observation may provide a rationale for application of neuroprotective strategies employing COX-2 inhibitors in inflammatory demyelinating neuropathies. PMID- 11271371 TI - Differential regulation of chromogranin A, chromogranin B and secretoneurin protein expression after transient forebrain ischemia in the gerbil. AB - The chromogranin/secretogranin family of proteins is widely distributed in the central nervous system, where they are stored in large dense-core vesicles. These proproteins are actively processed into small neuroactive peptides, which influence neurotransmitter release, microglial activation and monocyte migration. These properties suggest a possible role of chromogranins/secretogranins in the response that follows central nervous system injury. In the present study, the temporal pattern of expression and the distribution of chromogranin A, chromogranin B and secretoneurin, the major proteolytic product of secretogranin II, have been studied by immunohistochemistry after 5 min of transient forebrain ischemia in the Mongolian gerbil. A strong increase in the immunoreactivity for chromogranin A and secretoneurin was found in the CA3 pyramidal cell layer of the hippocampus, starting at 12 h, with a peak at 24 h and decrease at 48 h after transient forebrain ischemia. In the hippocampal formation, a rise in chromogranin A immunoreactivity was detected in neurons of the subiculum and the granule cell layer of the dentate gyrus. In addition, increase in the immunoreactivity for chromogranin A and secretoneurin was found in selected neurons of the neocortex. Chromogranin A and secretoneurin immunostaining patterns were similar in ischemic and control gerbils at 4 and 7 days following the ischemic insult. Chromogranin A and secretoneurin immunoreactivity in consecutive sections showed co-localization of both antigens but also selective overexpression of chromogranin A or secretoneurin in various neurons. No changes in chromogranin B immunoreactivity were detected across the time course following transient forebrain ischemia. These data indicate that changes in the expression of the chromogranin family of proteins after ischemia are selective for chromogranin A and secretoneurin. To our knowledge, this is the first study showing that the expression of the chromogranin family of proteins is differentially regulated after an ischemic insult in selected neuronal populations of the hippocampal formation and the cerebral cortex. Furthermore, the present data suggest a possible implication of chromogranin A and secretoneurin in the pathophysiology of transient forebrain ischemia. PMID- 11271372 TI - Distinct isoforms of tau aggregated in neurons and glial cells in brains of patients with Pick's disease, corticobasal degeneration and progressive supranuclear palsy. AB - We investigated isoform composition of aggregated tau protein in brains with Pick's disease (PiD), corticobasal degeneration (CBD) and progressive supranuclear palsy (PSP) by immunoblot analysis of sarkosyl-insoluble fractions of brain homogenates. We also examined the adjacent brain tissues immunohistochemically with a rabbit antibody, Ex10, which specifically recognizes exon 10 of tau. The Ex10 recognizes tau isoforms with four microtubule-binding repeats (4Rtau) but not those with three microtubule-binding repeats (3Rtau). Sarkosyl-insoluble tau from the brains of patients with CBD and PSP consisted of 4Rtau. Insoluble tau from the PiD brains contained both 3Rtau and 4Rtau, where 3Rtau predominated over 4Rtau. In brain tissues of CBD and PSP, Ex10 immunostained all neuronal and glial tau-positive structures. They included pre tangles, astrocytic plaques, tuft-shaped astrocytes, and oligodendroglial coiled bodies. In PiD brains, astrocytic inclusions were also positive for 4Rtau. However, the majority of, if not all, Pick bodies and oligodendroglial tau inclusions were negative for 4Rtau. Such results suggest that, in neurons and oligodendroglia, tau isoforms involved in the pathological processes differ between CBD/PSP and PiD, and are thus disease specific. This contrasts with the astrocytic tau isoforms that accumulate similarly in all three disorders. PMID- 11271373 TI - Expression of dystroglycan complex in satellite cells of dorsal root ganglia. AB - In Schwann cells, the transmembrane glycoprotein beta-dystroglycan composes the dystroglycan complex together with the extracellular glycoprotein alpha dystroglycan, which binds laminin-2 (alpha2/beta1/gamma1), a major component of the Schwann cell basal lamina. In the Schwann cell cytoplasm, beta-dystroglycan is anchored to a dystrophin isoform, Dp116. In this study, we investigated the expression of beta-dystroglycan, Dp116 and the laminin-alpha2 chain in satellite cells of rat dorsal root ganglia (DRGs). Immunohistochemical study showed that immunoreactivities for beta-dystroglycan and Dp116 were both localized to the outer rim of neuron-satellite cell and axon-Schwann cell units, indicating that both satellite and Schwann cells expressed these proteins in DRGs. Immunoreactivity for the laminin-alpha2 chain was detected in a similar location, indicating that the basal lamina surrounding satellite and Schwann cells in DRGs contained laminin-2. Ultrastructurally, immunoreactivity for the cytoplasmic domain of beta-dystroglycan as well as that for Dp116 was most intense in the cytoplasm just underlying the outer membrane of satellite cells. The immunoreactivity for laminin was associated with the outer surface of those cells, suggesting that it was localized in the surrounding basal lamina. These results indicate that the dystroglycan complex is expressed in the satellite cell outer membrane and involved in the adhesion with the basal lamina through the interaction with laminin-2. PMID- 11271374 TI - Inter- and/or intra-organ distribution of mitochondrial C3303T or A3243G mutation in mitochondrial cytopathy. AB - Fatal infantile mitochondrial cytopathy associated with a C3303T mutation in the mitochondrial tRNA(Leu(UuR)) gene has been reported clinically, biochemically and genetically. Here we have analyzed the percentage of this mutation in various autopsied tissues, and also in single muscle fibers using a micromanupulator, to evaluate the correlation between the pathology and heteroplasmic condition using polymerase chain reaction/restriction fragment length polymorphism. A 5-month-old Japanese girl was admitted to our hospital showing generalized muscle weakness, hepatomegaly, and cardiomegaly with lactic acidosis, and died at 6 months of age. Skeletal muscle showed severe degenerating myopathy found to be full of ragged red fibers (RRFs), an increased number of lipid droplets, and severe cytochrome c oxidase (COX) deficiency. Microscopically hepatocytes showed massive accumulation in lipid droplets, and the heart muscle showed a network pattern suggesting metabolic cardiomyopathy. The activities of respiratory chain enzyme complex I and complex IV in the skeletal muscle were significantly decreased to 23.4% and 5.0%, respectively, of the control value. The percentage of C3303T mutation in the patient tissues were variable, and ranged from 25% in the pancreas to 99% in the spinal cord. By single fiber analysis, the percentages of C3303T mutation in RRFs with COX negative (group 1; 42.4+/-7.0) and with COX positive (group 2; 58.2+/-5.8) were significantly higher than in non RRFs with normal COX staining (group 3; 10.7+/-6.3) (both P>0.001). Our patient showed a fatal infantile form of encephalopathy, myopathy and cardiomyopathy associated with widely distributed C3303T mutation in all of somatic cells. PMID- 11271375 TI - Autoradiography with [3H]PK11195 of spinal tract degeneration in amyotrophic lateral sclerosis. AB - The diagnostic hallmarks of amyotrophic lateral sclerosis (ALS) are degeneration of upper and lower motor neurons and of corticospinal tracts. Here, we demonstrate the suitability of the gliosis marker [3H]PK11195 for quantitative evaluation of tract degeneration in ALS in vitro. Binding of [3H]PK11195 was increased in lateral and ventral white matter of ALS spinal cords but not in the anterior horn, in spite of a dramatic loss in muscarinic binding sites and a high level of oxidatively modified protein. Labeling of activated microglia with [11C]PK11195 may also allow tract degeneration in ALS to be visualized in vivo. PMID- 11271376 TI - Postlesional transcriptional regulation of metabotropic glutamate receptors: implications for plasticity and excitotoxicity. AB - Metabotropic glutamate receptors (mGluRs) seem to be involved both in neuronal excitotoxicity evoked by pathological conditions such as ischemia, and in processes associated with neuronal plasticity and regulation of synaptic strength. Using semiquantitative reverse transcription-polymerase chain reaction, we measured the messenger RNA levels of mGluR2-5, 24 h and 7 days after experimentally induced focal cortical infarcts in rat motor cortex. The mRNA level of mGluR3 was strikingly decreased ipsilaterally after 24 h. On day 7, the expression of mGluR2 was down-regulated both ipsi- and contralaterally. Other receptors of this family showed either a slight or no regulation at both time points. The early changes in the expression pattern of mGluRs might be primarily linked to excitotoxicity processes which occur immediately after an ischemic lesion in cortex, whereas the delayed changes might represent one chain link in the cascade of compensatory mechanisms contributing to functional amelioration. PMID- 11271377 TI - Enhancement of central nervous system pathology in early simian immunodeficiency virus infection by dopaminergic drugs. AB - Human immunodeficiency virus infection (HIV) at late stages of the disease is accompanied by neurological complications, including motor, behavioral and cognitive impairment. Using simian immunodeficiency virus (SIV)-infected rhesus monkeys, an animal model of HIV infection, we found that during the asymptomatic SIV infection dopamine (DA) deficits are early components of central nervous system (CNS) dysfunction. To investigate the role of the DA system in SIV infection and to restore the DA deficiency, we administered selegiline, an agent with DAergic and neuroprotective properties, to SIV-infected monkeys. Selegiline increased DA availability but induced CNS vacuolization, SIV encephalitic lesions, and enhanced CNS viral replication during early SIV infection. The pathological changes seem to be mediated by DA, as treatment with L-DOPA, the precursor of DA, had similar effects. We propose that any natural or induced DAergic dysregulation which results in increased DA availability may potentiate HIV-associated neurological disease (ND). Our findings raise new questions regarding the pathogenesis of HIV-ND and generate concerns about the safety of dopaminergic drugs in the clinical management of HIV-infected patients. PMID- 11271378 TI - Apoptosis in the basal ganglia of the developing human nervous system. AB - Programmed cell death (PCD) plays a crucial role in the development of the central nervous system through controlling neuronal numbers and adequate synaptic connections. PCD has been considered to occur in the form of apoptosis. To examine how apoptosis occurs in the developing human brain, we performed a morphometric TUNEL study, using a commercially available kit (ApopTag Kit, Oncor Inc.). We examined apoptotic cells in the basal ganglia of 47 fetuses and newborns without macroscopical and microscopical evident congenital anomalies. Gestational age ranged from 12 to 40 weeks. The numerical density as well as the labeling index of TUNEL-positively labeled nuclei were evaluated. In the caudate nucleus and putamen, TUNEL-labeled cells were observed around the 12th week of gestation. The numerical density of total cells was significantly decreased, whereas the labeling index of apoptotic cells was significantly increased with advanced gestational age. In the globus pallidus, the numerical density of total cells decreased with advancing gestational age, while the labeling index of apoptotic cells increased between the 20th and 28th week, followed by a decrease until the 40th week. The analysis of TUNEL-positive cells revealed a different reaction pattern for the various basal ganglia with regard to the timing and degree of the apoptotic process in regulating cell numbers. PMID- 11271380 TI - Two novel glucose-6-phosphate dehydrogenase variants found in newborn mass screening for galactosaemia. AB - Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-linked recessive disorder in which haemolytic anaemia is the major symptom. The Beutler spot test employed in mass-screening for galactosaemia in newborns requires several intrinsic erythrocyte enzymes such as G6PD for its reaction and can theoretically detect G6PD deficiency apart from galactose-1-phosphate uridyltransferase deficiency. In this study, we detected two patients with G6PD deficiency using the quantitative Beutler test which was recently developed in our laboratory. Both patients lacked erythrocyte G6PD activity but exhibited no clinical symptoms. Molecular analysis in patients 1 and 2 revealed two novel missense mutations of C853T causing R285C and A1220C causing K407T, respectively. Molecular rather than enzymatic analysis was required in familial studies to detect and diagnose the carrier state. To date these patients have avoided oxidant stress and haemolytic diatheses have not been induced. CONCLUSION: Our results indicate that the quantitative Beutler test can detect glucose-6 phosphate dehydrogenase deficiency of class 1 and 2 and is therefore useful for early intervention and prevention of haemolytic diathesis in patients with this disorder. PMID- 11271379 TI - Carnitine-acylcarnitine translocase deficiency: phenotype, residual enzyme activity and outcome. AB - Carnitine-acylcarnitine translocase deficiency is a rare and life-threatening mitochondrial fatty acid beta-oxidation disorder. We describe a patient who, despite a severe clinical course and an extremely low carnitine-acylcarnitine translocase activity, is currently alive and in good health. We performed an extensive analysis of all previously published cases in order to evaluate the clinical features and prognostic factors. Reports on 21 patients with carnitine acylcarnitine translocase deficiency were obtained. Only 5 out of the 21 patients survived early childhood. At least 20 siblings are reported to have died of sudden unexplained death in the neonatal period. Although phenotype and residual enzyme activity have been suggested to be related to outcome, we were not able to establish such a relationship. CONCLUSION: Phenotype and residual enzyme activity do not appear to be major prognostic factors. Vigorous work-up in order to reach an expedite diagnosis and prompt medical intervention during acute episodes, especially in the neonatal period, may prevent fatal complications. PMID- 11271381 TI - Molybdenum balance studies in premature male infants. AB - Despite the fact that the trace element molybdenum (Mo) is essential, there is insufficient knowledge about the demands in infancy. Mo balances were therefore assessed under consideration of formula Mo concentrations ranging from 0.125 to 2.704 micromol/l. Sixteen premature male infants participated in the investigation. Their birth weights were between 1,500 and 1,990 g, the median (range) gestational age was 34 (32-36) weeks and the post-conceptual age at the time of study 37.4 (34.1-40.6) weeks. Twenty-four balance studies were performed and the materials analysed by atomic absorption spectroscopy. Infants with a "low" Mo intake received 0.024 (0.020-0.035) micromol/ kg per day, had a urinary excretion of 0.02 (0.008-0.045) and a retention of 0.0006 (-0.03 to 0.008) micromol/kg per day. Infants with a "high" intake received 0.284 (0.227-0.487) micromol/kg per day, had a urinary excretion of 0.243 (0.118-0.378) and a retention of 0.022 (-71.1 to 141.44) micromol/kg per day. Since the median urinary excretion exceeded 60% of the Mo intake at low and high intakes, sufficient resorption but minimal retention was assessed at low intakes of Mo. CONCLUSION: In view of the limited knowledge of long-term exposure to an elevated molybdenum intake and the substantial retention observed at higher intakes, upper limits should be set for molybdenum concentrations in preterm infant formulas. PMID- 11271382 TI - Intermittent hyperaldosteronism in a child due to an adrenal adenoma. AB - Aldosterone producing adenoma (APA) is a rare but potentially curable form of paediatric hypertension. We report a case of APA in a 9-year-old boy, suspected due to persistent hypokalaemia. Neither BP nor initial laboratory investigations disclosed the diagnosis and the presence of an APA was suggested by functional tests and radiological findings. Histologically, a cortical tumour was found associated with a marked medullary hyperplasia of both chromaffin and ganglion cells. CONCLUSION: This case reinforces the need for further investigations in patients with misleading clinical and laboratory data. PMID- 11271383 TI - A comparison of cytokine responses in respiratory syncytial virus and influenza A infections in infants. AB - Respiratory syncytial virus (RSV) infection is a major cause of bronchiolitis in infants while influenza A infection usually manifests as upper respiratory tract infection. We hypothesised that the immunological responses of infants to RSV infection and influenza A infection are different. This prospective study was undertaken to compare the cytokine responses during RSV and influenza A infection. Sera and nasopharyngeal aspirates (NPA) were collected from infants with a coryzal illness with or without wheeze who were admitted to the paediatric wards during 1998. Cytokines, adhesion molecules, RANTES, IgE and eosinophil cationic protein (ECP) were measured by enzyme linked immunosorbent assay or fluorescence enzyme immunoassay. The diagnosis of RSV and influenza infections was based on direct immunofluorescence and viral culture. Of the 39 infants studied, RSV infection was confirmed in 11 patients and Influenza A in 10 patients. All RSV patients and one influenza A patient had wheeze during the infection. The serum concentrations of interleukin (IL)-4 and IL-5, regulated upon activation normal T cell expressed and secreted (RANTES) and soluble intercellular adhesion molecule 1 (sICAM-1) in infants with RSV infection were significantly higher than those with influenza A infection (all P < 0.02). The concentration of tumour necrosis factor-alpha (TNF-alpha) in NPA was significantly lower in infants with RSV infection (P < 0.01). CONCLUSION: A predominant T helper cell type 2 cytokine and related immunological response was observed in infants with respiratory syncytial virus infection whereas a predominant pro-inflammatory cytokine response was observed in infants with influenza A infection. This may explain the different clinical manifestations of the two viral infections in infants. PMID- 11271384 TI - A characteristic EEG pattern in 4p-syndrome: case report and review of the literature. AB - Deletions on the short arm of chromosome 4 cause Wolf-Hirschhorn syndrome (WHS) and Pitt-Rogers-Danks syndrome (PRDS). WHS is associated with severe growth and mental retardation, microcephaly, a characteristic facies and congenital malformations. The PRDS phenotype is similar to WHS but generally less severe. Seizures occur in the majority of WHS and PRDS patients. Sgro et al. [17] described a stereotypic electroclinical pattern in four unrelated WHS patients, consisting of intermittent bursts of 2-3 Hz high voltage slow waves with spike wave activity in the parietal areas during drowsiness and sleep associated with myoclonic jerks. We report a patient with PRDS and the typical EEG pattern and review 14 WHS patients with similar EEG findings reported in the literature. CONCLUSION: Awareness and recognition of the characteristic electroclinical findings in Wolf-Hirschhorn syndrome and Pitt-Rogers-Danks syndrome might help in the early diagnosis of such patients. PMID- 11271386 TI - Malignant osteopetrosis with rickets. PMID- 11271385 TI - Neonatal and neurodevelopmental outcome in infants born before 30 weeks of gestation with absent or reversed end-diastolic flow velocities in the umbilical artery. AB - The objective of our study was to examine the outcome of infants born at a gestational age < 30 weeks with absent or reversed end-diastolic flow velocity (AREDFV) in the umbilical artery in comparison with gestational age-matched eutrophic controls. A group of 40 infants who had AREDFV were matched for gestational age and date of birth with 40 appropriate for gestational age infants. Perinatal outcome variables were retrospectively reviewed. In 16 out of the 40 matched pairs, a standardized neurological examination was done and, depending on age, the Kaufman Assessment Battery for Children or the Bayley Scales of Infant Development were applied at a corrected age of 13 to 100 months to assess neurodevelopmental outcome. The results were compared using Fisher's Exact Test or Mann Whitney U Tests as appropriate. In the AREDFV group, 26/40 (65%) survived until discharge compared to 39/40 (97.5%) in the control group (P < 0.001). AREDFV was associated with a higher rate of chronic lung disease, retinopathy of prematurity > or = grade III and impaired intestinal motility. More AREDFV infants suffered from permanent neurological sequelae compared with control infants: 44% versus 25% were mentally retarded (P = 0.033), and 38% versus 19% showed severe motor impairment (P = 0.073). CONCLUSION: Absent or reversed end-diastolic flow velocity is not only associated with a higher mortality and morbidity during the neonatal period, but the surviving infants of this high risk group have an increased risk for mental retardation and severe motor impairment as compared with appropriate for gestational age preterm infants of the same gestational age. PMID- 11271387 TI - Does the tuberous sclerosis complex include clivus chordoma? A case report. PMID- 11271389 TI - Effect of a high volume strategy high frequency oscillation on cerebral haemodynamics. PMID- 11271388 TI - Diagnosis and clinical course of congenital dyserythropoietic anaemia type 1 in a 12-year-old girl. PMID- 11271390 TI - Bilateral asynchronous adrenal adenoma in a girl with an incomplete form of Beckwith-Wiedemann syndrome. PMID- 11271391 TI - Joint laxity, scoliosis, and thoracic cage abnormalities in children with Beckwith-Wiedemann syndrome. PMID- 11271392 TI - Splanchnic oxygen delivery in exomphalos detected with near infrared spectroscopy. PMID- 11271393 TI - Cell volume regulation and transport mechanisms across the blood-brain barrier: implications for the management of hypernatraemic states. AB - Onset and correction of hypernatraemia constitute major hypertonic stresses for mammalian cells. Cells respond by activating specific osmoregulatory mechanisms allowing to preserve their volume and to adapt to their new environmental conditions. These processes have major implications in the management of hypernatraemia. In particular, cells chronically exposed to hypertonic conditions progressively accumulate organic osmolytes to maintain optimal intracellular electrolyte concentrations. During treatment of hypernatraemia, elevated intracellular organic osmotic content exposes cells to cellular oedema if sodium concentrations are rapidly corrected. In addition, circulating ions equilibrate slowly across the blood-brain barrier during acute changes in plasma osmolality. This can generate major brain water shifts and severe cerebral lesions related to brain shrinking or cerebral oedema. CONCLUSION: The basic mechanisms involved in brain ion and water transport are reviewed. A proper understanding of these processes is essential to develop appropriate treatment strategies in managing children with hypernatraemia. PMID- 11271394 TI - Ambulatory blood pressure monitoring in children with unilateral multicystic dysplastic kidney. AB - Multicystic dysplastic kidney (MCDK) is one of the most common congenital renal anomalies. Arterial hypertension is a potential complication of MCDK. Blood pressure (BP) has so far been measured only casually and the frequency of hypertension has been estimated to be between 0%-8%. Ambulatory blood pressure monitoring (ABPM) provides more precise information on BP than the casual BP measurement. The aim of this study was to investigate the BP profile in children with MCDK using ABPM. A group of 25 children (16 girls), with a mean age of 7.8 years (range 3.8-17.7 years) were investigated. ABPM was performed using the oscillometric SpaceLabs 90207 device. Hypertension was defined as mean systolic and/or diastolic BP during the day and/or in the night exceeding 95th percentile for ABPM. Five (20%) children showed hypertension, two of them had combined daytime and night-time hypertension and three had isolated nocturnal hypertension, although daytime BP was between the 90th-95th percentile in two of them. Children with ultrasonographical and/or laboratory signs of contralateral kidney abnormalities showed a higher incidence of hypertension than those without abnormalities (two of four versus 3 of 21). The mean night-time systolic and diastolic BP of children with MCDK was significantly higher than in healthy children (+ 0.50 and + 0.54 SDS, respectively, P = 0.012 and 0.03, respectively). Three of the hypertensive children were already nephrectomised. All five hypertensive children showed ultrasonographical and/or laboratory signs of contralateral kidney abnormalities. Hypertensive children had significantly higher microalbuminuria than normotensive children (6.9 +/- 3.2 mg/mmol creatinine versus 1.8 +/- 0.7, P = 0.03). The nocturnal BP fall (dip) was attenuated in five children, only one of whom was hypertensive. CONCLUSION: Arterial hypertension in children with multicystic dysplastic kidney is seen more often if based on ambulatory blood pressure monitoring than on casual blood pressure recordings. The main risk factor for developing hypertension is contralateral kidney damage. Ambulatory blood pressure monitoring should be performed in children with multicystic dysplastic kidney, especially in those with contralateral kidney abnormalities. PMID- 11271395 TI - Nasal glioma presenting as capillary haemangioma. AB - We report the case of a 5-month-old female infant with a congenital nasal tumour originally attributed to a capillary haemangioma. Doppler-flow ultrasound imaging revealed a solid mass surrounded by mildly enlarged vessels which had a flow pattern atypical of haemangioma. Histology showed non-malignant gliomatous cells with low proliferative activity. A diagnosis of nasal glioma was thus established and the patient underwent cranial MRT which excluded intracranial communication of the nasal glioma. Nasal gliomas arise from a skull defect, originating from the defective closure of the anterior neuroporus. They represent encephaloceles which have lost their intracranial connection. Nasal gliomas usually present shortly after birth as an intranasal obstruction or, as in our case, as a mostly extranasal tumour. CONCLUSION: Nasal glioma is often misdiagnosed as a capillary haemangioma. It can be distinguished from the latter by Doppler-flow ultrasonography. Magnetic resonance imaging is required to exclude intracranial communication. PMID- 11271396 TI - Connatal tuberculosis in an extremely low birth weight infant: case report and management of exposure to tuberculosis in a neonatal intensive care unit. AB - A case of connatal tuberculosis in an extremely low birth weight infant is reported. The patient was a female with a birth weight of 973 g born in the 27th week of pregnancy. She developed respiratory distress and signs of infection immediately after birth, which did not respond to mechanical ventilation, antibiotics, and corticosteroid therapy. Connatal tuberculosis was confirmed at 48 days of age by isolation of Mycobacterium tuberculosis from the infant's tracheal aspirate and the mother's menstrual discharge. The infant died of respiratory failure at 90 days of age. Mantoux tuberculin skin tests (TST) were performed on 99 infants, 144 medical staff members, and two family members. TST conversion occurred in three medical staff members, and preventive therapy with isoniazid was initiated. Eight exposed infants had normal chest X-rays and negative gastric aspirates for acid-fast bacilli and all received preventive isoniazid therapy. No case of tuberculosis developed during the 2-year follow-up period. CONCLUSION: Connatal tuberculosis should be considered in neonatal respiratory infection resistant to antibiotics. Prevention of transmission of tuberculosis on the neonatal intensive care unit by chemoprophylaxis is important. PMID- 11271397 TI - Prophylaxis of intravenous immunoglobulin and acyclovir in perinatal varicella. AB - Maternal chickenpox around the time of delivery can cause severe and even fatal illness in the newborn but an effectively preventive method has not yet been established. We proposed that a combination of intravenous immunoglobulin (IVIG) and acyclovir (ACV) intravenously could effectively prevent perinatal varicella. A group of 24 newborn infants whose mother had developed a varicella rash within 14 days before and after delivery were studied. Some 15 infants whose mothers' rash appeared within 7 days before and 5 days after delivery were categorised as an at-risk group and received IVIG prophylaxis (500 mg/kg) administered soon after birth or post-natal contact either alone or with intravenous acyclovir (5 mg/kg every 8 h) for a total of 5 days starting from 7 days after the onset of maternal rash. Of four infants receiving IVIG alone, two developed clinical varicella. None of ten infants receiving both IVIG and ACV contracted varicella. One infant receiving ACV alone had no varicella vesicles either. Of nine infants in the not at-risk group four had undetectable varicella-zoster virus antibody on admission and developed clinical varicella subsequently. CONCLUSION: The combination of intravenous immunoglobulin given soon after birth and prophylactic acyclovir intravenously administered 7 days after the onset of maternal rash can effectively prevent perinatal varicella. PMID- 11271399 TI - Modification of d-amphetamine-induced responses by baclofen in rats. AB - RATIONALE: d-Amphetamine is known to have effects on heart rate, body temperature and locomotor activity. However, it is not known if GABAB receptor stimulation modifies these actions of d-amphetamine. OBJECTIVES: Using telemetry recordings, this study examined the interactions between the GABAB receptor agonist baclofen and d-amphetamine, on heart rate responses, body temperature and locomotor activity, and whether these effects could be blocked by the GABAB receptor antagonist SCH 50911. METHODS: Rats were prepared with telemetry implants, which allowed continuous monitoring of heart rate, core temperature and spontaneous locomotor activity. Drugs were subsequently administered subcutaneously to the animals for simultaneous recording over a period of 180 min. RESULTS: d Amphetamine alone (0.3, 1.0 and 3.0 mg/kg) produced a dose-related increase in heart rate, but was without any effect on body temperature, whilst at 1.0 mg/kg, it significantly increased locomotor activity. Baclofen increased heart rate without affecting locomotor activity, and, at the highest dose of 10.0 mg/kg, it induced a significant reduction in body temperature. Graded doses of baclofen (3.0-10.0 mg/kg), when co-administered with d-amphetamine (1.0 mg/kg), did not modify the d-amphetamine-induced increase in heart rate, but the combination produced significant reductions of body temperature. The latter was only partially reversed by the GABAB receptor antagonist SCH 50911 (10.0 mg/kg). By contrast, the increase in locomotor activity by d-amphetamine was markedly blocked by baclofen in a dose-related manner, and this effect was abolished by SCH 50911 (10.0 mg/kg). SCH 50911 alone at doses of 3.0, 6.0 and 10.0 mg/kg had no effect on temperature or locomotor activity. Heart rate was increased significantly, but the magnitude of change was small. CONCLUSIONS: The results of the present study suggest that stimulation of GABAB receptors by baclofen completely blocks the locomotor stimulatory effects of d-amphetamine. Baclofen did not significantly modify the increase in heart rate produced by d-amphetamine and decreased body temperature, both alone and in combination with d-amphetamine. GABAB receptor ligands may well have potential as pharmacotherapies in the treatment of amphetamine abuse and dependence. PMID- 11271398 TI - Procalcitonin, IL-6, IL-8, IL-1 receptor antagonist and C-reactive protein as identificators of serious bacterial infections in children with fever without localising signs. AB - Fever without localising signs in very young children remains a diagnostic problem. Until present, a clinical scoring system combined with leucocyte count, urine analysis and determination of CRP are recognised as being helpful to identify patients at risk of serious bacterial illness. In this study we asked the question whether the determination of procalcitonin (PCT), interleukin (IL) 6, IL-8 and interleukin-1 receptor antagonist (IL- Ra) was superior to these commonly used markers for the prediction of a serious bacterial infection (SBI). Children, 7 days to 36 months of age, with a rectal temperature above 38 degrees C and without localising signs of infection were prospectively enrolled. For each infant, we performed a physical examination, a clinical score according to McCarthy, a complete white cell count, an urine analysis and a determination of CRP. We further determined PCT, IL-6, IL-8, and IL-1Ra concentrations and compared their predictive value with those of the usual management of fever without localising signs. Each infant at risk of SBI had blood culture, urine and cerebrospinal fluid cultures when indicated, and received antibiotics until culture results were available. A total of 124 children were included of whom 28 (23%) had SBI. Concentrations of PCT, CRP and IL-6 were significantly higher in the group of children with SBI but IL-8 and IL-1Ra were comparable between both groups. PCT showed a sensitivity of 93% and a specificity of 78% for detection of SBI and CRP had a sensitivity of 89% and a specificity of 75%. CONCLUSION: Compared to commonly used screening methods such as the McCarthy score, leucocyte count and other inflammatory markers such as interleukin-6, interleukin-8 and interleukin- receptor antagonist, procalcitonin and C-reactive protein offer a better sensitivity and specificity in predicting serious bacterial infection in children with fever without localising signs. PMID- 11271400 TI - Increased incidence of dyskinesias and other behavioral effects of re-exposure to neuroleptic treatment in social colonies of Cebus apella monkeys. AB - RATIONALE: Typical neuroleptic medications are still administered to as many as 40% of patients receiving antipsychotic treatment in the US. Intermittent administration or interruption of long-term neuroleptic medication for schizophrenia may increase the incidence of human tardive dyskinesias, and similarly may produce increasingly marked motor side-effects, parkinsonism, and other behavioral pathologies in non-human primates. OBJECTIVES AND METHODS: Given these similarities, we addressed the issue of prolonged and intermittent typical neuroleptic treatment and dopaminergic function during a 5-year, multi-phase study with social colonies of Cebus apella monkeys. In the previously reported phase 1, we examined the effects of 48 weeks of exposure to, followed by withdrawal from, fluphenazine decanoate (FPZ). Phase 3 reported here examined the effects of 18 weeks of re-exposure to FPZ in these same monkeys, 91 weeks after discontinuation of their phase 1 FPZ treatment. RESULTS: Analysis of blood plasma FPZ indicated levels of 0.22+/-0.08 ng/ml for the six injections during the re exposure period (n=54), comparable to the 0.24+/-0.07 ng/ml levels measured during our original treatment with this dose. Acute dyskinesias and dystonias increased by 300% upon re-exposure to FPZ; 15 of 18 FPZ-treated animals exhibited oral-buccal dyskinesias and all exhibited torticollis or retrocollis. Retreatment with FPZ was also associated with highly significant reductions in Self- and Environment-Directed Behavior and Directed Affiliation, effects similar to those seen during the original phase 1 FPZ treatment. Although FPZ re-treatment was associated with a significant reduction in Directed Aggression (an effect that was more robust than that observed during phase 1), in phase 3, we again observed an increase in Directed Aggression during early drug discontinuation when animals were in a stress-inducing situation. CONCLUSIONS: These results both support our phase 1 conclusion that typical neuroleptic medications may contribute to negative symptoms of schizophrenia and provide additional evidence for the possibility of increased aggression in stressful situations when medication is discontinued. Additionally, the results indicate that intermittent treatment with typical neuroleptics may dramatically increase the incidence of dystonias and dyskinesias. PMID- 11271401 TI - State-dependent effects of alcohol on recollective experience, familiarity and awareness of memories. AB - RATIONALE: Explicit memory (EM) is the memory for events which occurs with full awareness of where and how the recalled events took place, whereas implicit memory (IM) is the memory which is unfolded without any awareness of these events and usually becomes apparent when performance is facilitated by its presence. These two types of memory can be understood as different systems. Findings attempting to differentiate between the two systems in normal subjects have been controversial, with some researchers arguing that there is a single memory system and only the match in processes used during learning and later at retrieval can be important. OBJECTIVES: The present study compared the effects of alcohol (0.8 g/kg) or placebo administered prior to encoding and/or retrieval on measures of explicit and implicit memory in terms of recollective experience and familiarity. METHODS: At encoding subjects studied a list of 80 words presented in pairs. At retrieval, participants first carried out an implicit stem completion task, followed by an explicitly cued recall task (stem completion) which measured IM and EM respectively. After stem completion participants were required to indicate whether the items from the studied list were consciously recollected ("remember" response) or was known for a fact that were presented in the studied list ("know" response). In the IM task completed items from the studied list but not recognised by the subjects as such indicated memory without awareness. Studied items were of high and low associations. Forty-eight participants were tested in one of four drug conditions: alcohol-alcohol, placebo-placebo, placebo-alcohol, alcohol-placebo. RESULTS: In the implicit stem completion task, alcohol did not affect overall correct completion rates. However, participants who received alcohol prior to encoding reported lower awareness of correctly completed study items. In the cued recall task, alcohol also did not affect overall performance. However, participants in the same drug-state conditions (SS) reported greater recollection than familiarity with study material, whereas participants who encoded and retrieved material in different drug-state conditions (DS) reported recollection and familiarity to the same extent. In addition, DS participants showed more familiarity with study material compared to SS participants. Direct comparisons between IM and EM tasks demonstrated that alcohol at retrieval decreased the cued recall of items from high associations compared to placebo, but did not have any effect on implicit stem completion. CONCLUSIONS: In summary, these results demonstrate a dissociation of alcohol effects on measures of EM and IM. Alcohol administered prior to encoding reduced awareness of implicitly retrieved material, but did not impair IM per se, confirming previous findings with alcohol. In addition, the data provided new evidence for state-dependent retrieval effects on EM but not IM. It was also shown that for explicitly retrieved items, recollective experience benefits from same drug state, whereas familiarity benefits from different drug state between encoding and retrieval. PMID- 11271402 TI - Receptor-mediated regulation of serotonin output in the rat dorsal raphe nucleus: effects of risperidone. AB - OBJECTIVES: The present study was undertaken to characterize the regulation of serotonin (5-HT) efflux and neuronal activity in the dorsal raphe nucleus (DRN) as well as to examine the potential ability of the antipsychotic drug risperidone to interfere with these mechanisms. METHODS AND RESULTS: By using microdialysis in freely moving rats, it was found that administration of the alpha2 adrenoceptor antagonist idazoxan (0.25 mg/kg, SC), the 5-HT1B/D receptor antagonist GR 127,935 (1.0 mg/kg, SC) and risperidone (0.6 or 2.0 mg/kg, SC) increased 5-HT output in the DRN. Local DRN perfusion with GR 127,935 or risperidone via reversed dialysis (100 or 10-100 microM, respectively) enhanced 5 HT efflux in this area, whereas idazoxan (10-100 microM) failed to affect this parameter. Both systemic administration and reversed DRN dialysis of the D2/3 and 5-HT2A receptor antagonists raclopride (2.0 mg/kg, SC or 10-100 microM) and MDL 100,907 (1.0 mg/kg, SC or 10-100 microM), respectively, were without effect. Intraraphe dialysis of the 5-HT1B/D receptor agonist CP 135,807 (0.2 microM) decreased the efflux of 5-HT in the DRN, an effect which was antagonized by co administration of either GR 127,935 or risperidone (10 and 3.3 microM, respectively). By using single-cell recording, it was found that administration of GR 127,935 (50-400 microg/kg, IV) decreased, whereas CP 135,807 (2.5-20 microg/kg, IV) increased firing of 5-HT cells in the DRN. CONCLUSIONS: Our findings suggest a regulatory role of local 5-HT1B/D receptors on 5-HT efflux as well as cell firing in the DRN and indicate that risperidone may interfere with the regulation of 5-HT availability in this area primarily via blockade of 5-HT1D receptors. PMID- 11271403 TI - Nicotine self-administration and withdrawal: modulation of anxiety in the social interaction test in rats. AB - RATIONALE: Most smokers report smoking has an anxiolytic effect, which may contribute to nicotine dependence. OBJECTIVE: To examine effects in the social interaction test (SI) of anxiety after 4 weeks' self-administered nicotine (15 infusions of 0.03 mg/kg, totalling 0.45 mg/kg per day), and after 24 and 72 h of withdrawal. The effect of exposure to the operant chamber on withdrawal responses was also examined. METHODS: Animals were trained to self-administer saline or nicotine and after 4 weeks they were tested in SI after their daily self administration session. Animals were retested after 24 and 72 h withdrawal, when they were either taken directly from the home cage or were tested 5 min after a 30-min exposure to the operant chamber. RESULTS: Compared with the saline control group, the animals that had been self-administering nicotine for 4 weeks showed decreased social interaction with no decrease in locomotor activity, indicating a significant anxiogenic effect of the nicotine infusions. There was no change in social interaction after 24 and 72 h withdrawal from chronic nicotine, regardless of whether or not the rats were exposed to the operant chamber just prior to being tested. CONCLUSIONS: Nicotine self-administration is not maintained because of its anxiolytic effect, but despite, or because of, its anxiogenic effect. There was no evidence of an anxiogenic response after either 24 or 72 h of withdrawal and thus increased anxiety on withdrawal from nicotine does not seem to contribute to nicotine self-administration. PMID- 11271404 TI - Alterations in diurnal and nocturnal locomotor activity in rats treated with a monoamine-depleting regimen of methamphetamine or 3,4 methylenedioxymethamphetamine. AB - RATIONALE: The long-term neurochemical effects produced by the repeated administration of methamphetamine (MA) and 3,4-methylenedioxymethamphetamine (MDMA) are well documented; however, the functional consequences have not been clearly defined. OBJECTIVE: The present study was designed to investigate whether rats treated with a monoamine-depleting regimen of MA or MDMA exhibit disturbances in locomotor activity during the diurnal and nocturnal cycles. METHODS: Rats were treated with the vehicle or a monoamine-depleting regimen of MA or MDMA (10 mg/kg, IP, every 2 h for four injections on a single day). One week after drug treatment, the rats were placed in residential activity chambers and their locomotor activity was monitored for the next 7-day/night cycles. RESULTS: MA-treated rats exhibited depletions of striatal dopamine and serotonin content of approximately 70%, whereas MDMA-treated rats showed depletions of striatal serotonin content of approximately 50%. Rats treated with MA demonstrated a significant reduction in diurnal, but not nocturnal, locomotor activity, whereas MDMA-treated rats exhibited significant reductions in both diurnal and nocturnal locomotor activity. Analysis of the difference in activity between the nocturnal and diurnal cycles revealed that MA-treated animals exhibited a significantly greater change in activity as compared to that observed in vehicle- and MDMA-treated rats. CONCLUSIONS: Although it is unknown whether the adaptations in locomotor activity observed in MA- and MDMA-treated rats are due to the loss of dopamine and/or serotonin, these data suggest that the administration of a monoamine-depleting regimen of MA or MDMA results in alterations in light-cycle-dependent locomotor activity. PMID- 11271405 TI - Effects of nicotine on head-shakes and tryptophan metabolites. AB - RATIONALE: Nicotine appears to ameliorate the tics of Tourette syndrome. There is evidence that plasma concentrations of the tryptophan metabolite kynurenine may be elevated in this condition. Rodent head-shakes have been proposed as a putative model of Tourette syndrome and are potentiated by kynurenine. OBJECTIVES: To determine the effects of acute and chronic nicotine on mouse head shakes, and to study whether nicotine influences brain and plasma levels of kynurenine and certain of its further metabolites in this species. METHODS: Behavioural and biochemical studies. RESULTS: Acute (-10 min) administration of ( )-nicotine, or the nicotinic agonist (+)-epibatidine, dose dependently attenuated head-shakes induced by the 5-HT2A/2C receptor agonist +/-1-(2,5-dimethoxy-4 iodophenyl)-2-aminopropane (DOI). This attenuation was inhibited by the nicotinic receptor antagonist mecamylamine. Acute nicotine did not affect either spontaneous head-shakes or plasma and brain kynurenine. Fifteen hours after the last of twice daily injections of nicotine (1.6 mg/kg for 7 days), the frequency of spontaneous and DOI-induced head-shakes was significantly potentiated and there was a significant elevation of both plasma and brain kynurenine, although no differences were detected in plasma concentrations of tryptophan, kynurenic acid, 3-hydroxykynurenine or 3-hydroxyanthranilic acid. Brain levels of 5 hydroxytryptamine and 5-hydroxyindole acetic acid were also unaffected. In contrast, all these measures were unchanged 15 h after a single nicotine dose (1.6 mg/kg). CONCLUSIONS: The acute studies indicate that head-shakes induced by DOI are indeed inhibited by nicotinic receptor agonists and suggest that this is not a consequence of an increase in kynurenine. While a role for kynurenine or its metabolites in increasing the head-shake rate after chronic nicotine cannot be excluded, alternative explanations included alterations in the expression or functional status of nicotinic receptor components and further work will be required to characterise this effect. PMID- 11271406 TI - Time course of changes in cocaine self-administration behavior in rats during immunization with the cocaine vaccine IPC-1010. AB - RATIONALE: Following a 6-week immunization period consisting of three biweekly injections of the cocaine vaccine IPC-1010, the reacquisition of cocaine self administration behavior in rats was previously shown to be reduced in a manner that was dependant on serum antibody level. The present studies were conducted to examine additional issues relevant to the clinical use of the vaccine. OBJECTIVES: One experiment was conducted to address the issue of whether exposure to cocaine during the immunization period would influence the ability of the vaccine to block cocaine self-administration. A second experiment was conducted to determine if the reductions in drug-seeking behavior and drug intake by the vaccine were behaviorally specific, or if behavior maintained by a non-drug reinforcer would be similarly affected. METHODS: Identical second-order schedules of cocaine (1 mg/kg) or food pellet (45 mg) delivery were used in rats. In both studies, the time course of changes in behavior during the 6-week immunization period was examined in vaccine and alum-treated control rats following baseline and extinction conditions. RESULTS: The cocaine vaccine IPC-1010 induced average serum antibody levels of 0.07 mg/ml and significantly reduced self-administration behavior during the 2-week period following the third vaccine boost in a subgroup of rats with serum antibody levels greater than the average value. Cocaine self administration behavior at this time point significantly correlated with serum antibody level. IPC-1010 did not alter responding maintained by food throughout the immunization period although serum antibody levels reached a similar average of 0.06 mg/ml in this group of rats. CONCLUSIONS: These findings suggest that the reductions in drug-seeking behavior and drug intake after immunization with IPC 1010 did not result from a reduced ability of the rats to respond on the lever. Furthermore, daily exposure to cocaine during the immunization period did not influence the ability of the vaccine to reduce cocaine self-administration behavior that emerged gradually over time. These findings also confirm the need for a sufficiently high antibody level to blunt the reinforcing effects of cocaine. PMID- 11271407 TI - Contribution of the active metabolite, norcocaine, to cocaine's effects after intravenous and oral administration in rats: pharmacodynamics. AB - RATIONALE: Oral cocaine is more effective than IV cocaine by pharmacokinetic and pharmacodynamic analysis. One explanation is involvement of the active metabolite, norcocaine, in cocaine's effects. OBJECTIVES: To evaluate norcocaine's contribution to oral cocaine's effects, norcocaine's effects as a parent compound were determined and compared to those of cocaine using a differential reinforcement of low rate (DRL 45-s) schedule and spontaneous activity (large and small movements) after IV and PO routes of administration. METHODS: The effects of cocaine and norcocaine on DRL performance (shorter response and reinforcement rates) and spontaneous activity were investigated in 3 h sessions. The changes in effects across time (effect-time profiles) and dose response curves (DRCs) were constructed to evaluate the duration of action and potency (ED50) of both drugs. RESULTS: Under the DRL 45-s schedule, effect-time profiles showed both drugs via the two routes of administration significantly increasing and decreasing shorter-response rates and reinforcement rates, respectively. However, cocaine produced greater effects on shorterresponse rates than norcocaine, while both drugs produced comparable effects on reinforcement rates. For spontaneous activity, although IV cocaine, PO cocaine, and PO norcocaine dose- and time-dependently increased spontaneous activity, cocaine's effects were more profound than those of norcocaine. Effect-time profiles revealed that the duration of drug action was a function of dose, route, and behavioral paradigm used. According to ED50 values, IV cocaine was more effective than PO cocaine; however, PO cocaine was more effective than IV cocaine as judged by ED50 values corrected for absolute oral bioavailability. CONCLUSIONS: Norcocaine's contribution to oral cocaine's effects on DRL performance is evident. Other mechanism(s), such as a greater acute tolerance to IV cocaine's effects than to PO cocaine's effects, can be excluded. PMID- 11271408 TI - Distinct contributions of glutamate and dopamine receptors to temporal aspects of rodent working memory using a clinically relevant task. AB - RATIONALE: Understanding the mechanistic basis of working memory, the capacity to hold representation "on line," is important for delineating the processes involved in higher cognitive functions and the pathophysiology of thought disorders. OBJECTIVES: We compared the contribution of glutamate and dopamine receptor subtypes to temporal aspects of working memory using a modified rodent spatial working memory task that incorporates important elements of clinical working memory tasks. METHODS: A discrete paired-trial variable-delay T-maze task was used. Initial characterization studies indicated that performance on this task is stable at seconds-long retention intervals, is sensitive to retention interval and proactive interference, and is dependent on the integrity of the medial prefrontal cortex. RESULTS: Consistent with clinical findings, low dose amphetamine (0.25 mg/kg) produced a delay-dependent improvement in performance, while higher doses impaired performance at all retention intervals. D1 receptor blockade produced the predicted dose- and delay-dependent impairment. D2 receptor blockade had no effect. Activation of metabotropic glutamate 2/3 (mGluR2/3) receptors, which in the prefrontal cortex inhibits the slow asynchronous phase of glutamate release, also produced a delay-dependent impairment. Low doses of an AMPA/kainate antagonist had effects similar to the mGluR2/3 agonist. In contrast, NMDA receptor antagonist-induced impairment was memory load-insensitive, resulting in chance-level performance at all retention intervals. CONCLUSIONS: These findings suggest that activation of NMDA receptors is necessary for the formation of mnemonic encoding while modulatory components involving slow asynchronous release of glutamate and phasic release of dopamine contribute to the active maintenance of information during the delay period. PMID- 11271409 TI - 8-OH-DPAT, but not deramciclane, antagonizes the anxiogenic-like action of paroxetine in an elevated plus-maze. AB - OBJECTIVE: To investigate whether a 5-hydroxytryptamine (5-HT) reuptake inhibitor (paroxetine) has an anxiogenic-like effect and what possible pharmacological mechanism underlies that action. METHODS: We used the rat elevated plus-maze paradigm followed by measurement of locomotor activity. Some of the rats were subjected to handling and adaptation to the experimental situation, while the rest were naive to the test situation. Paroxetine was administered as a single treatment and in combination with the 5-HT1A receptor agonist (8-OH-DPAT) or 5 HT2A/2C receptor antagonist (deramciclane). RESULTS: The administration of paroxetine induced an anxiogenic-like action in rats adapted to handling, but not in handling naive animals. Treatment with paroxetine (0.1-2 mg/kg) reduced the number of open arm visits and time spent in open arms, and the ratio between open and total arm entries in the elevated plus-maze. Paroxetine also decreased the number of line crossings and head-dips. Paroxetine caused the strongest anti exploratory action at a dose of 0.5 mg/kg. Paroxetine did not suppress the locomotor activity of rats, showing that the described anti-exploratory effect was behaviourally specific to the plus-maze. Pretreatment with 8-OH-DPAT (0.05 mg/kg) completely reversed the anxiogenic-like action of paroxetine, whereas treatment with deramciclane (2 mg/kg) affected only the number of closed arm visits. Deramciclane (0.5-2 mg/kg) and 8-OH-DPAT (0.01-0.1 mg/kg) changed neither exploratory behaviour nor locomotor activity if given as single treatments to the habituated rats. CONCLUSION: The 5-HT reuptake inhibitor, paroxetine, at a low dose (0.5 mg/kg) induces an anxiogenic-like action in handling adapted rats. The effectiveness of 8-OH-DPAT against paroxetine probably supports a role of both pre- and postsynaptic 5HT-ergic mechanisms in the anxiogenic-like action of paroxetine. PMID- 11271410 TI - The relative contributions of ecstasy and cannabis to cognitive impairment. AB - RATIONALE: (+/-)-3,4-methylenedioxymethamphet-amine (MDMA; 'ecstasy'), a commonly used recreational drug, has typically been found to be related to poor cognitive function in humans. However, cannabis consumption may not have been adequately controlled for in these studies. OBJECTIVE: The present study was designed to further elucidate the relation between MDMA and cannabis in cognitive impairment. METHODS: Subjects who had used neither MDMA nor cannabis (controls; n=31), cannabis but not MDMA (cannabis users; n=18) and both MDMA and cannabis (MDMA/cannabis users; n=11) were compared on a battery of neuropsychological tests. RESULTS: The cannabis and MDMA/cannabis groups did not differ on any of the tests, whereas the combined cannabis and MDMA/cannabis groups performed more poorly than controls on tests of memory, learning, word fluency, speed of processing and manual dexterity. Further, apart from speed of processing where higher MDMA consumption predicted slower processing, covariate analysis revealed that the deficits were more closely related to cannabis than MDMA usage. CONCLUSION: The results suggest that cannabis is an important confound in studies of MDMA-related cognitive impairment, and that previously reported cognitive impairment in MDMA users may have been caused by coincident cannabis use. PMID- 11271411 TI - Acute hydrocortisone administration does not affect subjective responses to d amphetamine in humans. AB - RATIONALE: Stress and glucocorticoids facilitate and reinstate psychostimulant self-administration in rodents. However, the effects of stress and glucocorticoids on the subjective and behavioral effects of psychostimulants have not been well studied in humans. OBJECTIVES: To examine the effects of acute hydrocortisone pretreatment on the subjective and behavioral effects of d amphetamine. METHODS: Hydrocortisone (100 mg) and d-amphetamine (20 mg) were administered orally to 16 healthy male and female volunteers in a four-session, placebo-controlled, within-subject, crossover design. To prevent stomach irritation, subjects received rantidine hydrochloride before each experimental session. Dependent measures included self-reported mood and subjective effects (Addiction Research Center inventory, the profile of mood states, and a series of visual analogue scales), vital signs, salivary cortisol, and psychomotor performance. RESULTS: Hydrocortisone elevated salivary cortisol levels, produced modest dysphoria, and reduced subjects' reports of wanting more drug. However, hydrocortisone pretreatment did not affect any of the physiological, behavioral, or subjective effects of d-amphetamine. CONCLUSIONS: In contrast to the effects of glucocorticoids in rodent studies, these results indicate that an acute increase in cortisol does not enhance the psychostimulant effects of d amphetamine in humans. PMID- 11271412 TI - Following long-term training with citalopram, both mirtazapine and mianserin block its discriminative stimulus properties in rats. AB - RATIONALE: The discriminative stimulus (DS) properties of the selective serotonin (5-HT) uptake inhibitor (SSRI), citalopram, are mediated by 5-HT2C receptors. Interestingly, the "atypical" antidepressants, mianserin and mirtazapine, behave as antagonists at 5-HT2C receptors. OBJECTIVE: Herein, we evaluated the influence of mianserin and mirtazapine upon the DS effects of citalopram. METHODS: In a two lever drug discrimination procedure, rats initially trained to discriminate citalopram (2.5 mg/kg, i.p.) from saline were retrained with a lower dose of citalopram (0.63 mg/kg, i.p.). Subsequently, generalization and antagonist studies were conducted with mianserin and mirtazapine. RESULTS: Both dose dependently blocked, but did not generalize to, the DS properties of citalopram without markedly disrupting response rates. Their effective dose50s were 0.1 and 1.4 mg/kg, s.c., respectively. CONCLUSION: These observations are consistent with a role of 5-HT2C receptors in mediation of the interoceptive properties of SSRIs and suggest that the DS effects of citalopram are not related to its "antidepressant" properties per se. Finally, they underline the distinctive nature of mirtazapine and mianserin as compared to antidepressant agents which interact with 5-HT uptake sites. PMID- 11271413 TI - Appropriate use of "knockout" mice as models of depression or models of testing the efficacy of antidepressants. PMID- 11271415 TI - Green fluorescent protein as a reporter for gene expression in the mucoralean fungus Absidia glauca. AB - Mucoralean fungi (Zygomycota) are used for many industrial processes and also as important model organisms for investigating basic biological problems. Their genetic analysis is severely hampered by low transformation frequencies, by their strong tendency towards autonomous replication of plasmids instead of stable integration, and by the lack of reliable genetic reporter systems. We constructed plasmids for transforming the model zygomycete Absidia glauca that carry the versatile reporter gene coding for green fluorescent protein (GFP). gfp expression is controlled either by the homologous actin promoter or the promoter for the elongation factor of translation, EF1alpha. These plasmids also confer neomycin resistance and carry one of two genetic elements (rag1, seg1) that improve mitotic stability of the plasmid. The gfp constructs were replicated extrachromosomally and could be recovered from retransformed Escherichia coli cells. gfp expression was monitored by epifluorescence microscopy. The gfp reporter gene plasmids presented here for the model zygomycete A. glauca constitute the first reliable system that allows the monitoring of gene expression in this important group of fungi. PMID- 11271414 TI - A prescription to resist proscriptions for murine models of depression. PMID- 11271416 TI - Oxygen requirement for denitrification by the fungus Fusarium oxysporum. AB - The effects of dioxygen (O2) on the denitrification activity of the fungus Fusarium oxysporum MT-811 in fed-batch culture in a stirred jar fermentor were examined. The results revealed that fungal denitrifying activity requires a minimal amount of O2 for induction, which is repressed by excess O2. The optimal O2 supply differed between the denitrification substrates : 690 micromol O2 x h( 1) (g dry cell wt.)(-1) for nitrate (NO3-) and about 250 micromol O2 x h(-1) (g dry cell wt.)(-1) for nitrite (NO2-). The reduction of NO3- required more O2 than that of NO2- . With an optimal O2 supply, 80% and 52% of nitrogen atoms in NO3- and NO2-, respectively, were recovered as the denitrification product N2O. These features of F. oxysporum differ from those of bacterial denitrifiers that work exclusively under anoxic conditions. The denitrification activity of F. oxysporum MT-811 mutants with impaired NO3- assimilation was about double that of the wild type strain, suggesting competition for the substrate between assimilatory and dissimilatory types of NO3- reduction. These results showed that denitrification by F. oxysporum has unique features, namely, a minimal O2 requirement and competition with assimilatory NO3-. PMID- 11271417 TI - Significance of pantothenate for glucose fermentation by Oenococcus oeni and for suppression of the erythritol and acetate production. AB - The heterofermentative lactic acid bacterium Oenococcus oeni requires pantothenic acid for growth. In the presence of sufficient pantothenic acid, glucose was converted by heterolactic fermentation stoichiometrically to lactate, ethanol and CO2. Under pantothenic acid limitation, substantial amounts of erythritol, acetate and glycerol were produced by growing and resting bacteria. Production of erythritol and glycerol was required to compensate for the decreasing ethanol production and to enable the synthesis of acetate. In ribose fermentation, there were no shifts in the fermentation pattern in response to pantothenate supply. In the presence of pantothenate, growing O. oeni contained at least 10.2 microM HSCoA, whereas the HSCoA content was tenfold lower after growth in pantothenate depleted media. HSCoA and acetyl-CoA are cosubstrates of phosphotransacetylase and acetaldehyde dehydrogenase from the ethanol pathway. Both enzymes were found with activities commensurate with their function in ethanol production during heterolactic fermentation. From the kinetic data of the enzymes and the HSCoA and acetyl-CoA contents, it can be calculated that, under pantothenate limitation, phosphotransacetylase, and in particular acetaldehyde dehydrogenase activities become limiting due to low levels of the cosubstrates. Thus HSCoA deficiency represents the major limiting factor in heterolactic fermentation of glucose under pantothenate deficiency and the reason for the shift to erythritol, acetate, and glycerol fermentation. PMID- 11271418 TI - Glutamate decarboxylase activity in Trichoderma viride conidia and developing mycelia. AB - Glutamic acid decarboxylase (GAD) activity was measured in homogenates of conidia and both submerged and aerial mycelia of Trichoderma viride. The GAD activity in conidia had a temperature optimum at 30 degrees C and a pH optimum at pH 4. GAD was stimulated by EDTA (2 mM) and was insensitive to treatment with calmodulin antagonists calmidazolium (10 microM) or phenothiazine neuroleptics (60 microM). Cyclosporin A (up to 300 microM) partially inhibited GAD in the homogenate, but not in the supernatant obtained after centrifuging the homogenate. Attempts to release GAD activity from the homogenate using high ionic strength, detergents, or urea failed. Freezing-thawing led to the partial increase of activity in the conidial homogenate. These results indicate that GAD is a membrane-bound enzyme. The highest specific activity of GAD was present in the mitochondrial/vacuolar organellar fraction. Germination of conidia in the submerged culture led to a temporary decrease in GAD activity. After prolonged cultivation, the activity displayed quasi-oscillatory changes. The stationary state was characterized by a high GAD activity. The presence of gamma-aminobutyric acid in the submerged mycelia was demonstrated. In surface culture in the dark, GAD activity increased in a monophasic manner until conidia formation. The illumination of dark cultivated mycelia by a white-light pulse caused a dramatic increase in GAD activity. Light-induced changes were not observed in mutants with delayed onset of conidiation. In the dark or upon illumination by light pulse, the increase of GAD activity preceded the appearance of conidia. Thus, GAD activity in T. viride is closely associated with its developmental status and may represent a link between differentiation events and energy metabolism. PMID- 11271419 TI - Potassium uptake with low affinity and high rate in Enterococcus hirae at alkaline pH. AB - Two high-affinity K+ uptake systems, KtrI and KtrII, have been reported in Enterococcus hirae. A mutant, JEMK1, defective in these two systems did not grow at pH 10 in low-K+ medium (less than 1 mM K+), but grew well when supplemented with 10 mM KCl. In this mutant, we found an energy-dependent K+ uptake at pH 10 with a low affinity for K+ (Km of approximately 20 mM) and an extremely high rate [Vmax of 1.6 micromol x min(-1) (mg protein)(-1)]. Rb+ uptake [Km of approximately 40 mM and Vmax of 0.5 micromol x min(-1) (mg protein)(-1)], which was inhibited competitively by K+ and less prominently by Cs+, was also observed. The specificity of this transport is likely to be K+>Rb+>Cs+. This peculiar K+ transport plays a role as a salvage mechanism against defects in high-affinity systems in the K+ homeostasis of this bacterium. PMID- 11271420 TI - Virulent strains of Salmonella enteritidis disrupt the epithelial barrier of Caco 2 and HEp-2 cells. AB - To confirm the existence in nature of Salmonella enteritidis strains of different degrees of virulence and to elucidate the mechanisms underlying the effects of such strains on the epithelial barrier function, the consequences of infection of Caco-2 cells and HEp-2 cells with 15 S. enteritidis strains in a chicken infection model were examined. The more virulent strains of S. enteritidis, which are biofilm producers in adherence test medium, were able to disrupt HEp-2 and Caco-2 monolayers, as shown by transmonolayer electrical resistance and lactate dehydrogenase activity. In contrast, the low-virulence strains of S. enteritidis, which do not produce biofilms in adherence test medium, had no effect on the same cells. An avirulent rough mutant of Salmonella minnesota exhibited a pattern of behaviour similar to that of the low virulence strains of S. enteritidis, whilst a clinical Salmonella typhi strain caused rapid injury to the monolayers. The effect of supernatants of Salmonella cultures in adherence test medium on the integrity of Caco-2 cell monolayers indicated that the high-virulence S. enteritidis strains, but not the low-virulence strains, release a soluble factor when incubated under optimum biofilm-forming conditions, which enables the disruption of the integrity of Caco-2 monolayers. PMID- 11271421 TI - Different glycolytic pathways for glucose and fructose in the halophilic archaeon Halococcus saccharolyticus. AB - The glucose and fructose degradation pathways were analyzed in the halophilic archaeon Halococcus saccharolyticus by 13C-NMR labeling studies in growing cultures, comparative enzyme measurements and cell suspension experiments. H. saccharolyticus grown on complex media containing glucose or fructose specifically 13C-labeled at C1 and C3, formed acetate and small amounts of lactate. The 13C-labeling patterns, analyzed by 1H- and 13C-NMR, indicated that glucose was degraded via an Entner-Doudoroff (ED) type pathway (100%), whereas fructose was degraded almost completely via an Embden-Meyerhof (EM) type pathway (96%) and only to a small extent (4%) via an ED pathway. Glucose-grown and fructose-grown cells contained all the enzyme activities of the modified versions of the ED and EM pathways recently proposed for halophilic archaea. Glucose-grown cells showed increased activities of the ED enzymes gluconate dehydratase and 2 keto-3-deoxy-gluconate kinase, whereas fructose-grown cells contained higher activities of the key enzymes of a modified EM pathway, ketohexokinase and fructose-1-phosphate kinase. During growth of H. saccharolyticus on media containing both glucose and fructose, diauxic growth kinetics were observed. After complete consumption of glucose, fructose was degraded after a lag phase, in which fructose-1-phosphate kinase activity increased. Suspensions of glucose grown cells consumed initially only glucose rather than fructose, those of fructose-grown cells degraded fructose rather than glucose. Upon longer incubation times, glucose- and fructose-grown cells also metabolized the alternate hexoses. The data indicate that, in the archaeon H. saccharolyticus, the isomeric hexoses glucose and fructose are degraded via inducible, functionally separated glycolytic pathways: glucose via a modified ED pathway, and fructose via a modified EM pathway. PMID- 11271422 TI - Bypass of the requirement for cdc16p GAP function in Schizosaccharomyces pombe by mutation of the septation initiation network genes. AB - The onset of septum formation in the fission yeast Schizosaccharomyces pombe is signaled via the spglp GTPase-switch, which is part of the septation initiation network. This is negatively regulated by the two-component GTPase-activating protein (GAP) comprised of the products of the cdc16 and byr4 genes. Loss-of function mutations in either of these genes result in multiple rounds of septum formation without cell cleavage. In this work, we demonstrate that attenuation of the protein kinase cdc7p can rescue the lethality of a null allele of cdc16. This observation provides the basis for selection of chromosomal mutations and multicopy suppressors that attenuate the signaling of septation. Using this screen, mutations in all the previously described septation initiation network genes were obtained, with the exception of byr4, sid4 and plo1. We also demonstrate that increased expression of the dma1 gene can rescue the lethality of a null allele of cdc16. The implications for the regulation of septum formation in fission yeast are discussed. PMID- 11271423 TI - Mycoplasma hominis and Trichomonas vaginalis symbiosis: multiplicity of infection and transmissibility of M. hominis to human cells. AB - We recently reported that most Trichomonas vaginalis isolates cultured in vitro are infected by Mycoplasma hominis. In this work, we have characterized some aspects of the relationships between the two microorganisms. PCR, cultivation, and immunological methods revealed that the number of M. hominis organisms carried by T. vaginalis in culture varied from isolate to isolate, suggesting a specific multiplicity of infection. Moreover, infected T. vaginalis isolates were able to pass bacteria not only to M. hominis-free protozoa, but also to human derived epithelial cells. The in vitro transmission of the bacterium from T. vaginalis to both uninfected parasite isolates and human epithelial cells suggests a role for T. vaginalis as a carrier of the M. hominis infection in vivo. PMID- 11271424 TI - Nitrate respiratory metabolism in an obligately autotrophic hydrogen-oxidizing bacterium, Hydrogenobacter thermophilus TK-6. AB - Hydrogenobacter thermophilus strain TK-6 was observed to grow anaerobically on nitrate as an electron acceptor when molecular hydrogen was used as an energy source. Nitrite was detected as the product of a respiratory reaction. 15NO, 15N2O, and 15N2 were detected with Na15NO3 as an electron acceptor. Western immunoblot analysis showed that cell-free extracts from cells grown on nitrate reacted with antibodies against heme cd1-type nitrite reductase from Pseudomonas aeruginosa. The positive bands, which had molecular masses similar to that of the heme cd1-type nitrite reductase, were also stained by heme staining. These results indicate that nitrite reductase of strain TK-6 is a heme cd1-type enzyme. Activity of ATP:citrate lyase, one of the key enzymes of the reductive TCA cycle, was detected in cell-free extract of cells cultivated on nitrate, which indicates that the cycle operates during anaerobic growth. PMID- 11271425 TI - Molecular analysis of the grd operon coding for genes of the glycine reductase and of the thioredoxin system from Clostridium sticklandii. AB - A probe based on the sequence of the gene encoding selenoprotein A of glycine reductase of Clostridium sticklandii was used to obtain clones of adjacent DNA that encoded the other components of glycine reductase, proteins B and C, in addition to thioredoxin and thioredoxin reductase. The genes of the thioredoxin system and the glycine reductase were shown to be transcribed together, confirming an operon structure. In addition, a gene (grdX) encoding a 13.7-kDa protein of unknown function seemed to be associated with the reductase genes. Four potential promoters were identified by mapping the 5'-end of the mRNAs. The sequence of promoter P1 was shown to be similar to the sigma70 promoter consensus sequence. The other three promoters were similar to each other, but not to known promoter consensus sequences. The transcripts starting at each of the four promoters were terminated to about 80% at a predicted loop structure downstream of grdB; the remaining transcripts continued through this structure and covered the genes encoding both subunits of protein C and bmpA, a gene that was also expressed monocistronically. PMID- 11271426 TI - Perilymphatic pressure measurement in patients with Meniere's disease. AB - The MMS-10 Tympanic Displacement Analyser is a new device for measuring perilymphatic pressure in humans. This instrument was used in 70 patients with Meniere's disease (44 affected ears) and a group of 50 young normal hearing subjects. No significant differences in perilymphatic pressure measurements were found between the groups. Although measurement parameters showed large inter individual variation in a subgroup of 25 patients, the intra-individual correlation in the subgroup was good. In patients with Meniere's disease no relationship was found between perilymphatic pressure, hearing thresholds, blood pressure, gender or age. There was no difference between unilaterally and bilaterally affected patients. PMID- 11271427 TI - Tympanoplasty with and without mastoidectomy for non-cholesteatomatous chronic otitis media. AB - OBJECTIVES: Cases of non-cholesteatomatous chronic otits media (COM) were reviewed to determine whether mastoidectomy is helpful when combined with tympanoplasty for these conditions. STUDY DESIGN: A retrospective analysis of 251 ears with non-cholesteatomatous COM operated on by one surgeon (Y.M.) in an 11 year period was conducted. METHODS: Patients in group A (n = 147) were treated by tympanoplasty with mastoidectomy. Patients in group B (n = 104) were operated on without mastoidectomy. RESULTS: Graft success rates were 90.5% in group A and 93.3% in group B. There was no statistically significant difference. Graft success rates of discharging ears were 90.0% in group A and 85.7% in group B. Graft success rates of dry ears were 90.7% in group A and 94.4% in group B. There was no statistically significant difference between discharging ears and dry ears. The rates of the postoperative air-bone gap within 20dB were 81.6% in group A and 90.4% in group B, without a statistically significant difference. CONCLUSIONS: Mastoidectomy is not helpful in tympanoplasty for non cholesteatomatous COM, even if the ear is discharging. PMID- 11271428 TI - Sex distribution in children with tympanosclerosis after insertion of a tympanostomy tube. AB - Tympanostomy tube insertion is an accepted treatment for otitis media with effusion in children. Several clinical studies have shown that tube insertion may cause myringosclerosis. During the period 1988 to 1997 we treated 533 ears from 311 children who had otitis media with effusion by inserting tympanostomy tubes. Most of these (431 ears from 251 children) were re-examined in 1998 and sex and occurrence of myringosclerosis at the tube insertion site were noted. Myringosclerosis was observed in 31% of ears of girls treated with tubes, whereas in boys 71% of ears showed myringosclerosis. This difference between sexes may indicate a genetic predisposition such as that seen in atherosclerosis. PMID- 11271429 TI - Closed tympanoplasty in cholesteatoma surgery: long-term (10 years) hearing results using cartilage ossiculoplasty. AB - The aim of this retrospective study was to evaluate the long-term hearing results of using costal cartilage prostheses in ossicular chain reconstruction procedures in subjects operated on for a middle ear cholesteatoma with an intact canal wall tympanoplasty. Thirty-six patients (four with bilateral disease) followed up for 10 years who underwent an ossiculoplasty with a cartilage prostheses between January 1987 and December 1989 constituted the population studied. All the subjects underwent a staged intact canal wall tympanoplasty with mastoidectomy. Ossiculoplasty with total or partial chondroprosthesis was performed during the second stage. The long-term outcome was evaluated in terms of hearing according to the guidelines of the Committee on Hearing and Equilibrium (1995), and in terms of complications (anatomical and functional). In 18 patients a partial cartilage ossicular replacement prosthesis (PORP) was used, while in 22 a total cartilage ossicular replacement prosthesis (TORP) was used. In the PORP group the mean preoperative air-bone gap (ABG) was 22.4 dB hearing level (HL); before the second stage the ABG was 37.9 dB HL, at 2 years it was 12.1 dB HL, at 5 years 15.3 dB HL and at 10 years 15.8 dB HL. In the TORP group the mean preoperative ABG was 31.6 dB HL; before the second stage the ABG was 41.1 dB HL, at 2 years it was 14.4 dB HL, at 5 years 17 dB HL and at 10 years 18.5 dB HL. In both groups the number of cases with a postoperative ABG of < 20 dB HL remained stable (P > 0.05) over time. The failure rate was 17.5%, but only in 5% of cases was a functional revision needed. No cases of extrusion of the prostheses were encountered. The use of a chondroprosthesis is associated with functional results similar to those obtained by other authors. The efficacy of the prostheses remains stable over time and is associated with a very low rate of complications and failures. In this series no extrusion occurred and in no case did an infectious disease develop after cartilage transplantation. PMID- 11271430 TI - Pulmonary metastasectomy in the treatment of recurrent ameloblastoma of the maxilla and mandible: a case report. AB - Ameloblastoma is an aggressive benign tumor with frequent local recurrences. Although histologically benign, it occasionally metastasizes to many organs, most commonly to the lungs. The metastasis develops after multiple recurrences and many unsuccessful attempts at removal of the tumor. When distant metastasis occurs, the prognosis is poor and there is no effective treatment. A case of metastatic ameloblastoma of the mandible and maxilla is reported which was treated with pulmonary metastasectomy. The treatment options are discussed in relation to the literature. PMID- 11271431 TI - Intramuscular hemangioma of posterior neck muscles. AB - Intramuscular hemangiomas of the head and neck are rare tumors, sparsely reported. They usually present themselves as a mass which enlarges suddenly. A case of intramuscular hemangioma involving the posterior neck muscles is presented. Computed tomography scanning, magnetic resonance imaging and angiography revealed the vascular nature of this lesion. Surgery consisted of a wide excision. The patient is free of disease after a 4-year follow-up. PMID- 11271432 TI - New developments in the therapy of obstructive sleep apnea. AB - This review of the literature summarizes new trends in the diagnosis and treatment of obstructive sleep apnea (OSA) over the last 3 years. A literature search in Medline on 5 March 2000 using the keywords "OSA" and "OSAS" identified 123 papers. Another 86 articles were added from the references of the first 123 papers. New trends were observed concerning measurements of quality of life. There are new developments regarding conservative treatment, for example, nasal continuous positive airway pressure (nCPAP) therapy and oral devices. With regard to surgical treatment of OSA new surgical procedures, the radiofrequency technique, and the concept of multilevel surgery are discussed. After more than 25 years of interdisciplinary sleep medicine there still are some new developments of interest for ears, nose, and throat surgeons, which indicate the need for the involvement of otorhinolaryngologists in modern sleep medicine. PMID- 11271433 TI - Comparison of different 3D navigation systems by a clinical "user". AB - Three-dimensional navigation systems are routinely used in endoscopic skull base surgery, neurosurgery, maxillo-facial and endoscopic sinus surgery. Their precision can, however, change in the course of one experiment. We have compared five different 3D navigation systems and discuss here possible reasons for the limits of system precision. A plexiglass cube on which test points were marked served as a test-model. Two well-trained system users measured the distances between the test points in each of the five systems. The results were compared with reference data provided by the NUMEREX device at the Technical University of Vienna. The accuracy data shown by all these 3D navigation systems ranged from 0.0 mm to 6.67 mm. The accuracy data of a system calculated in advance did not always correspond with the system precision on the screen. The system precision in the center of the cube was higher than on its surface, which made us conclude that the angle between the tracker system and the pointing device touching the test point may be critical for system precision. Applying an automatic registration step did not result in greater system precision. Slice thickness and the angle of the pointing device seem to be responsible for system precision. PMID- 11271434 TI - Castleman's disease (giant lymph node hyperplasia) of the neck: a case report. AB - Castleman's disease (giant lymph node hyperplasia) is an uncommon cause of neck mass. Its cause and pathogenesis are still unknown. Giant lymph node hyperplasia (GLH) usually presents as an asymptomatic solitary mass and can occur anywhere in the head and neck. Diagnostic test results are always inconclusive. Excision and histopathological evaluation are the only ways to make a definitive diagnosis. The disease is curable by surgical excision. A case of GLH presenting as a solitary neck mass in a 68-year-old man is reported. There has been no recurrence during about 7 months. PMID- 11271435 TI - Results and complications of facial reanimation following cerebellopontine angle surgery. AB - The present study was undertaken to evaluate the results of a group of patients following treatment for cerebellopontine angle lesions who developed postoperative facial palsy and underwent facial nerve repair in order to reanimate the muscles of facial expression. A retrospective study was performed on 23 patients treated between 1988 and 1997 at the 2nd and 4th ENT chairs of University "La Sapienza" of Rome for facial palsy following cerebellopontine angle surgery. Tumors included acoustic neuromas (n = 3). Seventeen patients underwent hypoglossal-facial anastomoses [10 with end-to-end anastomoses, 4 with May's interposition "jump-nerve" grafts and 3 with partial (30%) use of the hypoglossal nerve plus a facial cross-over]. The remaining patients were operated on using a cable graft with the sural nerve (n = 2) and the great auricular nerve (n = 4). Postoperative facial function was determined by the House-Brackmann 6 scale classification The hypoglossal-facial anastomoses resulted in long-term grade III or IV findings. Cable grafts improved facial function from grade VI to grade III. None of the patients operated on with the modfied VII-XII anastomosis developed swallowing disturbances. The ten patients having traditional hypoglossal-facial anastomoses showed different degrees of tongue disability and retention of residue in the oral cavity. Surgical recovery of postoperative facial palsy can be obtained with various techniques according to the availability of the proximal facial nerve stump at the brain stem. Since a traditional hypoglossal-facial anastomosis procedure can be a source of a separate disability for the patient, techniques are preferred that leave the hypoglossal nerve mostly intact and uncompromised. PMID- 11271436 TI - Effect of bolus consistency on swallowing--does altering consistency help? AB - The influence of food bolus consistency on the pharyngeal wave during swallowing was investigated using a four-sensor manometry probe in 22 healthy volunteers. Pharyngeal pressures were recorded for 5 ml boluses of water, pudding and buttered bread via a manometry probe placed transnasally. The distal sensor was sited within the upper oesophageal sphincter (UOS); the three proximal sensors were then located 2, 4 and 6 cm above the UOS. The amplitude and timing of the swallow waveforms for pudding and buttered bread were recorded and compared with those for water. Increased bolus viscosity led to increased amplitude of the bolus wave and clearing contraction within the pharynx. In the UOS, increased bolus viscosity was associated with a larger pressure nadir (sub-atmospheric pressure) on opening and intra bolus pressure during transit. Bolus consistency also influenced the coordination of the swallow response with delayed pharyngeal clearance. The putative relevance of these findings to dietary modification for patients with neurological and neuromuscular dysphagia is discussed. PMID- 11271437 TI - Auditory brainstem and cochlear implants: functional results obtained after one year of rehabilitation. AB - Very little information has been published on the clinical outcome of auditory brainstem implants (ABI). The present paper evaluates results obtained in a patient affected by a bilateral acoustic neuroma in type II neurofibromatosis who received an implant during removal of the residual tumor. One year later surgical revision of the ABI was necessary because no auditory sensation was obtained after ABI activation. Twelve months after the surgical revision, 12 electrodes out of 15 evoked auditory sensation. The results of rehabilitation were compared with those obtained in a group of eight postlingually deaf patients with cochlear implants (CI). Twelve months postoperatively the CI patients identified 97.7 +/- 5.1% of bisyllabic words in a closed set while the ABI patient identified 86%. CI patients recognized 87.1 +/- 11.3% of sentences and 81.3 +/- 14.8% of words with contextual cues while the ABI patient recognized 75% and 65% respectively. Speech recognition improved more slowly in the ABI patient than in the CI patients and his scores for open-set words and sentences without lip reading and contextual cues were poorer. Although the results obtained in the ABI patient were not as good as those obtained in the CI patients, the ABI patient said his quality of life was improved. PMID- 11271438 TI - Different forms of dizziness occurring after cochlear implant. AB - Dizziness after cochlear implant (CI) was studied in a series of 94 consecutive adult patients receiving a cochlear implant, 46 (49.0%) of whom experienced dizziness post-operatively. In 29 patients, post-operative dizziness occurred soon after surgery and subsided within one month. Dizziness of the continuous type, lasting more than 6 months, was a complaint in only two patients. In addition to these already known forms of dizziness, spells of vertigo occurring later than one month after cochlear implant were experienced by 15 patients (delayed-V). The spells of delayed-V occurred suddenly and persisted for several hours. Moreover, 85.7% of delayed-V patients complained of hearing and tinnitus abnormalities during these spells. The clinical features of delayed-V were similar to those in patients with Meniere's disease. The preoperative bithermal caloric test showed a significantly higher response for the delayed-V group than the other groups (ANOVA: P < 0.05) in terms of slow phase eye velocity of caloric nystagmus. These findings suggest that inner ear lesions due to cochlear implant surgery develop gradually. Similarities in clinical features between delayed-V and Meniere's disease indicate the presence of labyrinthine hydrops. PMID- 11271439 TI - The privacy wars begin. PMID- 11271440 TI - See the big picture, understand the details. AB - Ray B. Lefton, DDS, FHFMA, MBA, CPA, is the CFO of Temple East Inc., a subsidiary of Temple University Health System, in Philadelphia, Pennsylvania. Lefton's primary responsibilities include all traditional financial duties. He also serves as Temple East's CIO and compliance officer, and has the departments of finance, medical records, registration, patient accounting, materials management, information systems, rehabilitation services, podiatry, and hospitalists reporting to him. He is a member of HFMA's Metropolitan Philadelphia Chapter. His organization has made great strides in reducing both denied and avoidable days and in lowering days in accounts receivable. PMID- 11271441 TI - Assessing an IDS's readiness to operate profitably under a risk contract. AB - An IDS that wishes to assume risk should understand first the factors that have caused IDSs to experience poor financial performance under risk contracts. Then, the IDS should clarify its goals for managed care contracting, assess its operational strengths and weaknesses, evaluate potential managed care payer partners, and understand the amount of risk associated with various contract options. Implementation issues that need to be addressed include aligning the financial incentives of physicians and the IDS, establishing systemwide policies and procedures for monitoring performance, and developing information systems capabilities. PMID- 11271442 TI - Internet-based eligibility verification lowers costs, improves payment timeliness. AB - Verification of claims eligibility traditionally is performed either through telephone communication between the payer's and provider's staff or through an EDI clearinghouse. These forms of verification offer inconsistent quality, often use out-of-date information, cost time and money, and may financially harm payers and providers by leading to delays in payments or claims denied after care has been rendered. Internet-based eligibility verification software reduces the number of rejected claims because it can access current information without using a clearinghouse. The software also allows earlier collection of copayments because these obligations can be identified accurately when care is rendered. By offering Internet-based eligibility verification, payers can help providers achieve financial benefits while gaining such benefits themselves. PMID- 11271443 TI - Realizing positive returns from your e-health investments. AB - Although the healthcare industry still is in the beginning stages of e-health adoption and has yet to achieve a significant return on e-health investments, healthcare organizations can anticipate increasingly positive results over time. To capitalize on the potential of e-health, financial managers need to adopt a clear, concise approach to valuing e-health investment options, making investment decisions, and managing an organization's return on invested capital. Key actions to take are developing a strategy, looking beyond cost reduction to other advantages of e-health strategies, understanding the needs of the stakeholders, becoming comfortable with experimentation, having realistic expectations, and instituting a process for valuing e-health investments that does not duplicate valuation processes for traditional information technology projects. PMID- 11271444 TI - Charge-process strategies for outpatient prospective payment. AB - One of the most important strategies for operating successfully under the outpatient prospective payment system is to establish an effective charging process. Achieving control of the process begins with a thorough assessment of chargemaster coding, how charges are structured in the chargemaster, and how hospital departments use the charges. Once problems are identified and corrected, the next step is to develop a system to maintain and monitor both the chargemaster and the charging process. Perhaps the most important component of the monitoring system is to include a mechanism to provide the necessary feedback to departments regarding errors related to the charge process. PMID- 11271445 TI - Accounting for a not-for-profit organization's fund-raising costs. AB - The AICPA's Statement of Position (SOP) 98-2 provides guidance on how a not-for profit entity should account for costs associated with activities that involve a fund-raising component in combination with one or more mission-related components. Costs may be allocated among the various components of such an activity as long as the activity meets certain criteria specified by SOP 98-2. These criteria are related to the activity's purpose, audience, and content. If the activity does not meet the criteria, then all costs of the activity must be shown as fund-raising costs. PMID- 11271446 TI - Improving group practice performance with benchmarking. AB - Group practices can use benchmarking to improve physician productivity to best practice levels. The benchmarking process can be broken down into two phases. In the first phase, the problem is identified. This phase involves identifying critical drivers, choosing an external benchmark, gathering internal data, identifying variances, and establishing targets. In the second phase, action is taken. This phase involves identifying actions to take, defining responsibilities, implementing the changes, and monitoring performance. Group practices that use benchmarking need to understand the tool's limitations. Benchmarks serve as roadmaps, but any action plan should be tailored to the practice and take a variety of factors into consideration. PMID- 11271447 TI - HCFA proposes PPS for inpatient rehabilitation services. PMID- 11271448 TI - Making a healthcare Web site a sound investment. PMID- 11271449 TI - Outsourcing investment management: creating and evaluating requests for proposals. PMID- 11271450 TI - A corporate culture match is vital to job success. PMID- 11271451 TI - Mesalamine maintenance therapy for Crohn's disease. PMID- 11271452 TI - Limitations of anti-guinea pig liver transglutaminase IgA in screening of celiac disease. PMID- 11271453 TI - Motilin-related peptide and ghrelin: lessons from molecular techniques, peptide chemistry, and receptor biology. PMID- 11271454 TI - Identification of motilin-related peptide. PMID- 11271455 TI - A comparative study of corneal endothelial changes induced by different durations of soft contact lens wear. AB - PURPOSE: To analyze the effect on the morphologic characteristics of the corneal endothelium of the duration of soft contact lens wearing periods. METHODS: Ninety soft contact lens wearers were divided into three groups: short-term users, for less than 5 years (n=60 eyes); intermediate-term users, from 6 years to 10 years (n=60); longterm users, for more than 10 years (n=60). Thirty non-contact lens wearers (60 eyes) were included as controls. All eyes were examined with a specular microscope. Analysis of covariance was used to detect any differences among the controls and the various soft contact lens subgroups. RESULTS: There was a significant correlation between duration of soft contact lens use and morphologic changes of corneal endothelium. All soft contact lens subgroups had a significantly greater coefficient of variation in cell size than non-contact lens users . The proportion of hexagonal cells and the mean corneal endothelial cell density in those using soft contact lenses for more than 6 years were significantly lower than in the control group . Soft contact lens wear was significantly correlated with decreasing corneal endothelial cell densities with time. CONCLUSION: The coefficient of variation in cell size may be a sensitive indicator of early morphologic changes of the corneal endothelium. As the decrease in cell density among the contact lens subgroups was significantly associated with the duration of soft contact lens wearing periods, it will be useful to investigate endothelial cell density for evaluation of corneal endothelial function concerned with contact lens wearing. PMID- 11271456 TI - Graft endothelium and thickness after penetrating keratoplasty, comparing mechanical and excimer laser trephination: a prospective randomised study. AB - PURPOSE: To assess the impact of nonmechanical trephination on the graft endothelium and thickness after penetrating keratoplasty (PK). METHODS: Inclusion criteria for this prospective, randomised, cross-sectional, clinical study were: (1) Treatment between October 1992 and December 1997; (2) one surgeon (G.O.H.N.); (3) primary central PK; (4) Fuchs' dystrophy (diameter 7.5/7.6 mm) or keratoconus (diameter 8.0/8.1 mm); (5) graft oversize 0.1 mm; (6) no previous intraocular surgery; (7) 16-bite double-running diagonal suture. In 179 patients (mean age 51+/-18 years), PK was performed using either the 193-nm Meditec MEL60 excimer laser ("Excimer") along metal masks with eight "orientation teeth/notches" (53 keratoconus, 35 Fuchs' dystrophy) or motor trephination with the Mikrokeratron (Geuder) ("Control": 53 keratoconus, 38 Fuchs' dystrophy). For donor trephination from the epithelial side an artificial anterior chamber was used in both groups. In 27% of the excimer and 29% of the control group a triple procedure was performed. Specular microscopy (EM-1000, Tomey) and pachymetry (SP-2000, Tomey) were performed before removal of the first suture (0.4+/-0.2 years postoperatively), before (1.1+/-0.4 years) and after (1.7+/-0.6 years) removal of the second suture but before any additional surgical intervention. RESULTS: Endothelial cell count: Neither "two-sutures-in" (1953+/-426/1804+/-385 cells/mm2, p=0.13), "one-suture-in" (1629+/-439/1765+/-440 cells/mm2, p=0.27), nor "all-sutures-out" (1259+/-493/1294+/-532 cells/mm2, p=0.83) differed significantly between Excimer and Control. Graft thickness: Neither "two-sutures in" (527+/-58/524+/-16 mucrom, p=0.89), "one-suture-in" (537+/-72/551+/-40 microm, p=0.86), nor "all-sutures-out" (576+/-53/565+/-62 microm, p=0.38) differed significantly between Excimer and Control. Cell count and corneal thickness were not significantly different comparing Fuchs' dystrophy and keratoconus or comparing PK only and triple procedures. Graft thickness and endothelial cell count correlated highly significantly inversely with "all sutures out" (P<0.0001). CONCLUSIONS: Excimer laser trephination from the epithelial side using an artificial anterior chamber in donors seems to have no disadvantages concerning the graft endothelium after PK. Endothelial cell loss was not increased in eyes with Fuchs' dystrophy compared with keratoconus or after triple procedures compared with PK only. PMID- 11271457 TI - Indocyanine green and fluorescein hyperfluorescence at the site of occlusion in branch retinal vein occlusion. AB - BACKGROUND: In patients with branch retinal vein occlusion (BRVO), we investigated the presence of indocyanine green (ICG) and fluorescein hyperfluorescence at the site of occlusion. We also assessed the association of this feature with the clinical outcome of these patients. METHODS: Both indocyanine green (ICG) videoangiography and fluorescein angiography (FAG) were performed in 21 eyes with BRVO of less than 1 month duration. Deterioration of the disease was defined clinically as an increase in retinal hemorrhages or retinal edema. Capillary nonperfusion was quantified with computer image analysis from the FAG pictures. RESULTS: ICG videoangiography showed focal hyperfluorescence along the venous wall at the site of the affected A-V crossing in 9 of the 21 eyes, and FAG showed this feature in 10 eyes. The ICG hyperfluorescence was more prominently and focally detected than the hyperfluorescence on FAG, which was sometimes diffusely seen throughout the whole occluded area. Eight of the nine eyes showing ICG hyperfluorescence had clinical deterioration with an increase in retinal hemorrhage or edema. This deterioration occurred more frequently in eyes with hyperfluorescence and/or late leakage than in ones without these features. The mean nonperfused area was significantly larger in eyes with hyperfluorescence than in eyes without these features. CONCLUSION: The ICG hyperfluorescence at the site of A-V crossing is associated with disease deterioration in patients with fresh BRVO. The ICG hyperfluorescence was more easily detectable than the hyperfluorescence on FAG, although the difference in sensitivity between the two methods is not great. PMID- 11271458 TI - Enzymatic heterogeneity of bovine retinal pigment epithelial cells in vivo and in vitro. AB - BACKGROUND: The retinal pigment epithelium (RPE) contains a variety of enzymes that are involved in the metabolism of the neural retina. It has been shown that the photoreceptor densities display regional variations. We aimed to find out whether the enzymes in the RPE also display regional differences. METHODS: Various enzymes were localized in RPE cells in situ and in cultured RPE cells. RPE cells of the entire tapetal region of adult and fetal bovine eyes were examined. The following enzymes were studied: Activities of aminopeptidases A and M, dipeptidylpeptidases I and II, and gamma-glutamyltranspeptidase were localized by histochemistry. Alkaline phosphatase, dipeptidylpeptidase IV, and cathepsin B were studied by histochemistry and immunocytochemistry. In addition, activities of two enzymes were localized in one cell. RESULTS: The distribution pattern of the enzymatic activities showed no marked regional differences. In contrast, pronounced cell-to-cell variability in the activities was detected for some of the enzymes tested. These intercellular differences were detected already in fetal retinae of early stages and persisted in RPE cultures. CONCLUSION: The heterogeneous distribution of enzymatic activities in RPE cells appears not to be caused exclusively by stimuli from the neural retina and from the choroid, because heterogeneity starts in the early fetal period and persists in culture. PMID- 11271459 TI - Immunology and growth characteristics of ocular basal cell carcinoma. AB - BACKGROUND: Knowledge about immunological features and growth characteristics of palpebral (ocular) basal cell carcinomas (BCCs) is limited. In particular, it is unclear whether ocular BCC represents in this regard a special BCC entity or not. METHODS: Twenty BCCs of the lid area (ocular BCCs) were investigated immunohistologically using monoclonal antibodies against CD4, CD8, CD45Ro, CD50, CD68, HECA-452, Ki67 (MIB1), and the p53 epitope. For comparison, nine BCCs excised distant from the eye (non-ocular BCCs) were evaluated. RESULTS: In BCCs the distribution of the immunocompetent cells investigated is markedly irregular. These cells are localized mainly around BCC islands. Only a few of them invade tumour cell aggregates. The CD4:CD8 ratio as detected by immunohistochemistry is >1 in 82% of ocular BCCs and in 88% of nonocular BCCs. Often there are dense infiltrations of CD68+ cells (macrophages) and HECA-452+ cells adjacent to tumour cell aggregates. The growth fraction [percentage of proliferating (Ki67+/MIB 1+) cells] varies from 0% to more than 30%. Proliferative activity is enhanced at the invasion front. Additionally, the amount of p53+ cells differs considerably among the BCCs. CONCLUSIONS: CD4+ T cells seem to be the most important cell population for BCC immunosurveillance, offering the chance for conservative interferon therapy. The role of CD68+ and HECA-452+ cells has to be further elucidated. In many tumours the large amount of proliferating cells contrasts to the usually slow growth of BCCs, indicating strong apoptotic processes. The results can be regarded only as semiquantitative. So far, ocular and nonocular BCCs exhibit no essential differences regarding immunocompetent cell infiltration and growth characteristics. According to this, palpebral BCCs are "normal" BCCs and not a special BCC variant. Therefore, results from dermatological research concerning BCC can be extended without limitations to their counterparts in the lid area. PMID- 11271460 TI - Optical coherence tomography (OCT) findings in normal retina and laser-induced choroidal neovascularization in rats. AB - BACKGROUND: To evaluate the optical coherence tomographic (OCT) features of the retina of rats, we compared the OCT images with the histological appearance of normal retinas and retinas with laser-induced choroidal neovascularization. METHODS: Twelve eyes of 12 adult pigmented rats (Brown-Norway) were used. Color fundus photography, fluorescein angiography (FAG), and OCT images of normal retinas and retinas with laser photocoagulation-induced choroidal neovascularization were studied. RESULTS: OCT showed a double-layered structure in the normal sensory retina with a highly reflective layer located in the inner retina and a low reflective layer located in the outer retina. The retinal pigment epithelium (RPE) and choriocapillaris were imaged as a layer with the highest reflection. On the first day after photocoagulation, OCT showed a disruption of the highly reflective layer corresponding to the RPE, and an enhanced reflectivity in the choroid under the lesion. Choroidal neovascularizations (CNVs) which appeared 2 weeks after photocoagulation was seen as a multi-layered, highly reflective area protruding from the RPE into the subretinal space A CNV beneath a subretinal hematoma was difficult to detect because of the low transmission of the scanning light through the hematoma. The histopathological appearance was well correlated with the OCT images. CONCLUSION: The two reflective bands in the OCT images were identified as coming from the inner layers of the retina and from the photoreceptors. The highest reflective band arose from the RPE and choriocapillaris. In the future, OCT combined with FAG or indocyanine-green angiography will be a useful tool for the evaluation of animal studies of choroidal neovascularization and other retinal diseases. PMID- 11271461 TI - Blue light-induced apoptosis in cultured retinal pigment epithelium cells of the rat. AB - BACKGROUND: We previously demonstrated that phagosome-free retinal pigment epithelium (RPE) cells in culture can be damaged directly by blue light (wavelength 440+/-10 nm) as observed by electron microscope. A low intensity (1.0 mW/cm2) of light induced only swelling of mitochondria, while a high intensity (4.0 mW/cm2) induced necrosis in the RPE. The aim of the present study was to investigate what intensity of blue light could induce apoptosis in cultured phagosome-free RPE. METHODS: Primary cultured RPE cells, harvested from Long Evans rats, that contained no phagosomes were exposed to a cool blue light (wavelength 440+/-10 nm). After exposure, transmission electron microscopy (TEM) and TdT-mediated dUTP nick-end labeling (TUNEL) staining were used to detect apoptosis in the RPE cells. To assess the relationship of oxidation to apoptosis by blue light, we added N-acetylcysteine (NAC) as a free radical scavenger and investigated its inhibitory effect on apoptosis. RESULTS: In RPE cells exposed to blue light of 2.7 mW/cm2 for 24 h, apoptotic bodies were found by TEM. In RPE cells exposed to blue light of 2.0 mW/cm2 for 60 h, apoptotic bodies, nuclear condensation and nuclear segmentation were observed by TEM and some RPE cells showed positive TUNEL staining. When 30 mM of NAC was added, TUNEL staining was negative. CONCLUSION: Our findings demonstrate that apoptotic cell death is induced by blue light exposure in cultured RPE cells in vitro. The findings of our previous experiments and those of the present study suggest that a higher intensity of blue light could induce necrosis, and moderately intense blue light could induce non-necrotic cell death or apoptosis, in RPE cells. Furthermore, it is suggested that blue light caused cell death by a free-radical-associated mechanism. PMID- 11271462 TI - Clinicopathological correlation in exudative age-related macular degeneration: recurrent choroidal neovascularization. AB - PURPOSE: To report the pathology of surgically removed submacular tissue in recurrent choroidal neovascularization after laser photocoagulation of classic choroidal neovascularization in age-related macular degeneration. METHODS: A recurrent subfoveal choroidal neovascular membrane was surgically removed in two patients. The recurrence was identified as a classic membrane on fluorescein angiography at the foveal border of the laser scar. A net was visualized in the early venous phase of the indocyanine green angiogram, with associated late hyperfluorescence. Both patients had undergone laser photocoagulation for a classic interpapillomacular choroidal neovascular membrane about 1 1/2 years earlier. The specimens were serially sectioned and stained with hematoxylin eosin, periodic acid-Schiff, Masson trichrome and phosphotungstic acid hematoxylin. RESULTS: The two specimens consisted of subretinal fibrovascular tissue with fibrin exudation. Fibrovascular tissue bordered subretinal fibrous tissue adherent to Bruch's membrane and remnants of the choroid in one patient. The fibrovascular portion most likely corresponded to the recurrence, whereas the fibrous portion represented the original membrane, being obliterated after photocoagulation. Some peripapillary tissue was additionally removed in the other patient. The latter lesion was invisible on fluorescein angiography but stained in the late phase of indocyanine green angiography and corresponded histopathologically to poorly vascularized intra-Bruch's fibrovascular tissue. Granular deposits, periodic acid-Schiff positive and metachromatically purple on Masson trichrome stain, representing diffuse drusen (basal laminar/linear deposits), were identified in the three specimens. CONCLUSION: A subretinal fibrovascular membrane corresponded with the classic recurrent choroidal neovascularization. PMID- 11271463 TI - Alteration of the vascular supply in the rabbit ciliary body by transscleral diode laser cyclophotocoagulation. AB - BACKGROUND: Besides the direct destruction of the non-pigmented ciliary epithelium by cyclodestructive procedures, further mechanisms are responsible for the decrease of intraocular pressure. This study evaluates the alteration of the ciliary body vascularization by contact transscleral diode laser cyclophotocoagulation in rabbit eyes. METHODS: Pigmented chinchilla bastard rabbits were used. Preliminary experiments were conducted to determine the parameters for diode laser cyclophotocoagulation of the pars plana or pars plicata. Then, treatment of the pars plicata (three rabbits) or pars plana (three rabbits) was performed in the right eye of six rabbits. After 2, 6 and 12 weeks histologic and transmission electron microscopic studies were performed. Furthermore, three rabbits received pars plicata cyclophotocoagulation of the right and pars plana cyclophotocoagulation of the left eye. After 2, 6 and 12 weeks, vascular casts of the ciliary body were investigated by scanning electron microscopy. RESULTS: Histologic and transmission electron microscopic studies showed a marked coagulation necrosis with subsequent ciliary body atrophy, destruction of the ciliary epithelium, pigment dispersion in the ciliary body stroma and peripheral anterior synechiae. Examination of vascular casts of the ciliary body revealed a marked rarefication of the capillary network within the treated areas of the ciliary body in all eyes and at every time of investigation. Anterior to the laser burns the capillary network was not markedly affected in the eyes with cyclophotocoagulation of the pars plana. After 3 months short vessel sprouts were seen, but regeneration was mostly incomplete. CONCLUSIONS: The vascular casting technique is an excellent method for the investigation of changes in ciliary body vascularization after cyclodestruction. This study is the first to demonstrate a marked rarefication of the ciliary body vascularization after diode laser cyclophotocoagulation using vascular casts. The results suggest that alteration of vascularization probably acts as a strong synergistic mechanism in the decrease of intraocular pressure after cyclophotocoagulation of the pars plicata. PMID- 11271464 TI - Ischemia increases prostaglandin H synthase-2 levels in retina and visual cortex in piglets. AB - BACKGROUND: Ischemia increases levels of prostaglandin H synthase-2 (PGHS-2) in neonatal brain and cerebral vasculature, but effects on the developing visual system are unknown. We examined the effects of ischemia on PGHS-2 mRNA and protein levels in the retina and visual cortex in anesthetized piglets. METHODS: Ten minutes of complete retinal and brain ischemia was induced by increasing intracranial pressure. After 2-12 h of reperfusion, samples of retina and visual cortex were collected for determinations of levels of PGHS-2 mRNA (RNase protection assay) or protein (immunohistochemistry and western blotting). Tissues also were obtained from control animals. RESULTS: Levels of PGHS-2 mRNA were undetectable in control animals but showed a dramatic increase at 2-4 h in the cortex and retina in animals exposed to ischemia. Detectable but limited PGHS-2 immunoreactivity (IR) was present in the retina and visual cortex from control animals. In piglets not subjected to ischemia, PGHS-2 IR was localized mainly to the outer limiting membrane and to the Muller cells. Ischemia induced a marked increase in PGHS-2 IR in the neural retina, with the greatest increase in the photoreceptor layer. PGHS-2 levels in whole retina also increased at 8 h after ischemia. In the intact visual cortex PGHS-2 IR was evident in layers II and V. Ischemia increased the intensity of IR in layers II/III as well as layer V. CONCLUSIONS: Detectable amounts of PGHS-2 protein are present in the piglet retina and visual cortex under normal conditions, but levels are markedly increased 8-12 h after ischemic stress. Enhanced PGHS-2 levels after ischemic stress may contribute to delayed pathological changes of the visual system in the neonate. PMID- 11271465 TI - Lack of association of mutations of the bestrophin gene with age-related macular degeneration in non-familial Japanese patients. AB - BACKGROUND: Heterozygous mutations of the bestrophin gene are associated with Best macular dystrophy (BMD). The bestrophin gene is specifically expressed in the retinal pigment epithelium. BMD is a hereditary form of macular degeneration that may develop subretinal neovascularisation similar to the wet type of age related macular degeneration (AMD). PURPOSE: To study whether mutations of the bestrophin gene occur in non-familial Japanese AMD patients. METHODS: A total of 85 non-familial AMD patients (average age 67.5 years; 71 male, 14 female) diagnosed by indocyanine green angiography were screened. Among them, 69 patients (81 %) were classified as having wet type AMD. Genomic DNA was purified from the total blood and used as the template for polymerase chain reaction (PCR). All the exons of bestrophin gene were amplified by PCR. Mutation analysis was performed by SSCP using the ABI Prism 310 Genetic Analyzer (Perkin Elmer). Nucleotide sequence was determined by direct sequencing of the PCR amplicons. As the control, 105 non-AMD patients (average age 62.0 years; 52 male, 53 female) were screened by the same method. RESULTS: Only one AMD patient had a specific polymorphism in exon 2, but no mutations leading to amino acid substitutions were found. In exon 2 and 3, two further polymorphisms were detected in all AMD patients as well as normal controls. CONCLUSION: No mutations were found in the bestrophin gene in nonfamilial Japanese patients with AMD or in normal controls. PMID- 11271466 TI - The global tuberculosis situation and the new control strategy of the World Health Organization. 1991. PMID- 11271467 TI - Quality oversight: why are rural hospitals less likely to be JCAHO accredited? AB - There is a large rural-urban disparity in the proportion of hospitals that are accredited by the Joint Commission on the Accreditation of Health Care Organizations (JCAHO). Several factors can influence whether a hospital participates in the accreditation process. A few of those factors include the hospital's size, case mix and ownership. However, even after controlling for many of these factors, hospitals in the most rural locations are less likely to be accredited by the JCAHO than urban hospitals. A survey was conducted to explore why rural hospitals are not participating in the accreditation process. Survey results show that the largest factor contributing to rural hospital deterrence to seeking accreditation is cost. Without accreditation by the JCAHO and compliance with their movement into performance measurement, quality monitoring of rural hospitals will fall further behind that of urban hospitals. Policy initiatives that make accreditation more financially feasible should be considered. PMID- 11271468 TI - A semi-empirical approach to the modeling of the electrophoretic mobility in free solution: application to polystyrenesulfonates of various sulfonation rates. AB - This work focuses on the understanding of the electrophoretic behavior of flexible chains of polystyrenesulfonates (PSSs) in free solution. It deals mainly with the variation of the electrophoretic mobility with (i) the polymerization degree (N) of fully sulfonated PSSs and (ii) the sulfonation rate of randomly sulfonated PSSs. In both cases, the electrophoretic mobility was modeled following a semi-empirical approach which involves parameters retaining a physical meaning. Fully sulfonated PSS oligomers, having a length smaller than or similar to the Debye length, exhibit a particular electrophoretic behavior, in between that observed for multicharged small molecules and that for polyelectrolytes. The electrophoretic mobility of these oligomers increases strongly with N, which is attributed to a hydrodynamic coupling between monomers. Then the mobility is maximum for an N of about 10, for which the PSS oligomers are still in a rod-like conformation. Afterwards, as N increases and the PSSs are larger than the Debye length, the electrophoretic mobility decreases slowly until it reaches a constant value corresponding to the free-draining behavior. Next, the electrophoretic behavior of long PSS (N about 1,200) differing in their sulfonation rates was investigated. The effective charge rates were determined independently by conductimetric measurements and the mobilities were modeled as a function of the sulfonation rate. The PSS behavior observed was compared to the one previously reported for classical polyelectrolytes having hydrophilic backbones, such as copolymers of poly(acryamide-coacrylic acid). A specific behavior has been pointed out for these partially sulfonated PSSs, which is attributed to the hydrophobicity of their backbone. Finally, it is shown that separations of PSSs of different sulfonation rates can be obtained with electrolytes containing an anionic surfactant or methanol. PMID- 11271469 TI - Electrokinetic flow in a planar slit covered by an ion-penetrable charged membrane. AB - The electrokinetic flow of an electrolyte solution in a planar slit covered by an ion-penetrable charged membrane layer is analyzed theoretically. An approximate analytical expression for the spatial variation in the electrical potential is derived, and the electroosmotic velocity, the total electric current, and the streaming potential of the system under consideration are evaluated. The effects of epsilon' (relative permittivity of liquid phase/relative permittivity of membrane layer), eta' (viscosity of liquid phase/viscosity of membrane layer) and the valence of anions (coions) on the volumetric flow rate and total current are examined. We show that the effect of the valence of cations (counterions) on the volumetric flow rate is less significant than that of epsilon' and that of eta'. However, the effect of epsilon' on the total current is less significant than that of the valence of cations and that of eta'. The variation of total current as a function of ionic strength is found to have a local minimum, regardless of whether a pressure gradient is applied or not. The absolute streaming potential has a local maximum as the concentration of fixed charge varies, which was not found in previous studies. PMID- 11271471 TI - High-resolution electrophoretic procedures for the identification of five Eimeria species from chickens, and detection of population variation. AB - To overcome limitations of conventional approaches for the identification of Eimeria species of chickens, we have established high resolution electrophoretic procedures using genetic markers in ribosomal DNA. The first and second internal transcribed spacer (ITS-1 and ITS-2) regions of ribosomal DNA were amplified by polymerase chain reaction (PCR) from genomic DNA samples representing five species of Eimeria (E. acervulina, E. brunetti, E. maxima, E. necatrix and E. tenella), denatured and then subjected to denaturing polyacrylamide gel electrophoresis (D-PAGE) or single-strand conformation polymorphism (SSCP) analysis. Differences in D-PAGE profiles for both the ITS-1 and ITS-2 fragments (combined with an apparent lack of variation within individual species) enabled the unequivocal identification of the five species, and SSCP allowed the detection of population variation between some isolates representing E. acervulina, which remained undetected by D-PAGE. The establishment of these approaches has important implications for controlling the purity of laboratory lines of Eimeria, for diagnosis and for studying the epidemiology of coccidiosis. PMID- 11271470 TI - An electrophoretic study of vitamin E status and expression of heat shock proteins in alveolar type II and liver cells. AB - Vitamin E is the most important lipophilic antioxidant. Oxidative injuries are prevented or minimized by vitamin E supplementation. Various physiological and pathological situations are accompanied by vitamin E deficiency. However, it is not clear whether alimentary vitamin E deficiency in itself constitutes oxidant stress that induces appropriate responses, which, in turn, can be avoided by adequate vitamin E supplies, or whether the remaining cellular antioxidants compensate a temporary vitamin E deficiency. We studied effects of the dietary vitamin E status on cellular vitamin E levels and on the expression of heat shock proteins (HSPs) in alveolar type II cells and liver. The expression of HSPs, representing an early and very sensitive marker of cellular stress, was compared with the activity of antioxidative enzymes. Vitamin E depletion caused a substantial increase in HSP32 in alveolar type II cells, whereas in liver there was a marked increase in HSP70. The activity of the antioxidant enzymes, however, did not change significantly. A reversal of HSP expression to almost normal levels was seen after vitamin E resupplementation. These results indicate that, under normal conditions, a suboptimal supply of vitamin E to rats exposes the alveolar type II cells and the liver to reversible cellular stress. PMID- 11271472 TI - Infrared fluorescent automated detection of thirteen short tandem repeat polymorphisms and one gender-determining system of the CODIS core system. AB - We used an infrared (IR) automated fluorescence monolaser sequencer for the analysis of 13 autosomal short tandem repeat (STR) systems (TPOX, D3S1358, FGA, CSF1PO, D5S818, D7S820, D8S1179, TH01, vWA, D13S317, D16S359, D18S51, D21S11) and the X-Y homologous gene amelogenin system. These two systems represent the core of the combined DNA index systems (CODIS). Four independent multiplex reactions, based on the polymerase chain reaction (PCR) technique and on the direct labeling of the forward primer of every primer pair, with a new molecule (IRDye800), were set up, permitting the exact characterization of the alleles by comparison with ladders of specific sequenced alleles. This is the first report of the whole analysis of the STRs of the CODIS core using an IR automated DNA sequencer. The protocol was used to solve paternity/maternity tests and for population studies. The electrophoretic system also proved useful for the correct typing of those loci differing in size by only 2 bp. A sensibility study demonstrated that the test can detect an average of 10 pg of undegraded human DNA. We also performed a preliminary study analyzing some forensic samples and mixed stains, which suggested the usefulness of using this analytical system for human identification as well as for forensic purposes. PMID- 11271473 TI - Detection of sequence variability of the collagen type IIalpha 1 3' variable number of tandem repeat. AB - The variable number of tandem repeat (VNTR) 3' of the collagen type II (COL2A1) gene has been shown to be highly variable with a complex molecular structure. In a previous pilot experiment we observed discordance between methods to genotype this informative marker. To further investigate the extent and molecular nature of this discordance, we genotyped a random sample of 207 Caucasian individuals with two genotyping methods and sequenced new alleles. We compared single-strand (SS) analysis, which is based on detection of size differences between the different alleles, and heteroduplex analysis (HA), which is sensitive to both size and sequence differences. Overall, 26% of discordance between the two methods was detected. Approximately two thirds of this discordance was caused by subdivision of SS-alleles 13R1 and 14R2 into HA-alleles 4A + 4B and 3B + 3C, respectively. Sequence analysis of the COL2A1 VNTR alleles 4B and 3C showed that these alleles differed in sequence, but not in size, from already described SS alleles, which explains why they escape detection by SS. The 4B allele is a frequent allele in the population (14%) and is, therefore, important to distinguish in association studies. We conclude that HA is a reliable method when the described optimized electrophoretic conditions are used. HA is a sensitive genotyping method to document allelic diversity at this locus, which can distinguish more alleles compared to the SS method. PMID- 11271474 TI - The IL-10 gene is not involved in the predisposition to inflammatory bowel disease. AB - Although genetic predisposition for inflammatory bowel disease (IBD) is well established, little is known about the accountable genes. The pathogenesis of IBD is characterized by an imbalanced activation of Th1- and Th2-lymphocytes. IL-10 represents an anti-inflammatory cytokine which downregulates the production of Th1-derived cytokines. To evaluate the role of the IL-10 gene in IBD, two polymorphisms in the promoter region (G/A at position -1082 and C/A at position 592) were genotyped in 142 patients with Crohn's disease (CD), 104 patients with ulcerative colitis (UC), and 400 healthy controls. Significant differences were not apparent, neither in the allele frequencies of either polymorphism, nor in the haplotype frequencies. Screening of the coding region of the IL-10 gene by polymerase chain reaction--single strand conformation polymorphism (PCR-SSCP) analysis revealed a rare sequence variation in exon 1 leading to an amino acid exchange (G-->A; G15R) in two patients with CD and five healthy controls. Therefore, polymorphisms of the IL-10 gene are not demonstrably involved in the predisposition of IBD in our cohorts of patients. PMID- 11271475 TI - Capillary zone electrophoresis of sub-microm-sized particles in electrolyte solutions of various ionic strengths: size-dependent electrophoretic migration and separation efficiency. AB - To gain insight into the mechanisms of size-dependent separation of microparticles in capillary zone electrophoresis (CZE), sulfated polystyrene latex microspheres of 139, 189, 268, and 381 nm radius were subjected to CZE in Tris-borate buffers of various ionic strengths ranging from 0.0003 to 0.005, at electric field strengths of 100-500 V cm(-1). Size-dependent electrophoretic migration of polystyrene particles in CZE was shown to be an explicit function of kappaR, where kappa(-1) and rare the thickness of electric double layer (which can be derived from the ionic strength of the buffer) and particle radius, respectively. Particle mobility depends on kappaR in a manner consistent with that expected from the Overbeek-Booth electrokinetic theory, though a charged hairy layer on the surface of polystyrene latex particles complicates the quantitative prediction and optimization of size-dependent separation of such particles in CZE. However, the Overbeek-Booth theory remains a useful general guide for size-dependent separation of microparticles in CZE. In accordance with it, it could be shown that, for a given pair of polystyrene particles of different sizes, there exists an ionic strength which provides the optimal separation selectivity. Peak spreading was promoted by both an increasing electric field strength and a decreasing ionic strength. When the capillary is efficiently thermostated, the electrophoretic heterogeneity of polystyrene microspheres appears to be the major contributor to peak spreading. Yet, at both elevated electric field strengths (500 V/cm) and the highest ionic strength used (0.005), thermal effects in a capillary appear to contribute significantly to peak spreading or can even dominate it. PMID- 11271476 TI - Derivatization, extraction and concentration of amino acids and peptides by using organic/aqueous phases in capillary electrophoresis with fluorescence detection. AB - We report a novel method that facilitates sample pretreatment and detection in amino acid analysis by coupling solvent extraction with capillary electrophoresis. Amino acids and peptides were fluorescently labeled, concentrated into an organic solvent, and then separated by capillary zone electrophoresis with fluorescence detection. To achieve this, acetophenone was first employed to dissolve the derivatizing reagent, fluorescamine. The products, which possessed both hydrophilic and hydrophobic moieties, could be extracted and concentrated into the organic phase by suppressing the deprotonation of carboxyl groups, thus enhancing the hydrophobicity of the resulting molecules through pH modification in the aqueous solution. Furthermore, by fine-tuning the pH value, individual amino acids and short peptide molecules could be separated selectively from the sample bulk. This convenient, chemically controllable concentration technique may be useful in sample concentration and purification of biologically related samples such as amino acids and short peptides. PMID- 11271477 TI - Formation of concentration gradient and its application to DNA capillary electrophoresis. AB - A new method to introduce the concentration gradient into the capillary has been developed and its application to DNA capillary electrophoresis is presented. The concentration gradient produced by mixing 5% w/v polyacrylamide-co-poly(N dimethylacrylamide) (PAM-co-PDMA) solution and 1 x Tris/N tris(hydroxymethyl)methyl-3-amino-propanesulfonic acid/EDTA (TT) + 5 M urea buffer was successfully achieved by using two programmable syringe pumps with strict control of dead volume, flow rate, and pressure balance. This method has the advantages of high stability, reproducibility, and versatility. The column with concentration gradient greatly improved the resolution, especially for the large DNA fragments, due to a decrease in band width broadening with time. A column containing 2-4% w/v gradient in four steps had a longer read length, shorter separation time and better resolution (after 380 base) than that of 4% w/v single concentration polymer solution. The number of steps in the gradient had almost no effect on the performance. The change in the average concentration by relocating the position of the same step gradient, i.e., a combination of different low concentration to high concentration polymer solution ratios, resulted in a different migration time, read length and resolution. PMID- 11271478 TI - Enantioseparation of dihydropyridine derivatives by means of neutral and negatively charged beta-cyclodextrin derivatives using capillary electrophoresis. AB - Employing capillary electrophoresis, the racemates of 29 acidic, neutral and basic dihydropyridines (DHPs) were separated by means of neutral and negatively charged cyclodextrins (CDs). Whereas the enantiomers of the acidic DHPs could be resolved with neutral CDs, mostly alpha- and beta-CD, the enantiomers of the neutral DHPs were only baseline-separated using the sulfobutyl ether-substituted beta-CD. Working in reversed polarity mode (detector at the anode) improved the peak shape and the resolution of the enantiomers. The racemates of the DHP bearing a secondary or tertiary amine function in the side chain at position 3 could be separated by using either the neutral gamma-CD or negatively charged CDs. The poor peak shape found with anionic CDs could be improved by the addition of methanol. The combination of gamma-CD and sulfated beta-CD allowed the detection of the minor enantiomer of lercanidipine (24) at less than 1% w/w. PMID- 11271479 TI - Competitive binding of a Tris run buffer with chiral crown ether in chiral capillary electrophoresis. AB - In capillary electrophoresis of primary amine racemates using (+)-(18-crown-6) tetracarboxylic acid (18C6H4) as a chiral selector, chiral recognition emanates from the differences in the complex formation between 18C6H4 and the two protonated amine enantiomers. The presence of buffer constituents such as tris(hydroxymethyl)aminomethane (Tris) or Na+, capable of forming complexes with 18C6H4, is thus detrimental to the chiral separation of primary amines. Such a competitive binding of buffer constituents was studied by comparing the electrophoretic mobilities of racemic analytes obtained in Tris/citric acid and triethylamine/citric acid buffers. We developed a simple fitting method to determine the competitive binding constant and applied it to the Tris buffer system. The competitive binding constant of Tris with 18C6H4 obtained at pH 3.0 was 27 +/- 4. PMID- 11271481 TI - Analysis of catechins and caffeine in tea extracts by micellar electrokinetic chromatography. AB - Cancer chemotherapy is a new and important medical science and much interest has been focused on catechins, not only for their antioxidant activity, but also because of their known antimutagenic and antitumorigenic properties. Green tea and black tea, which are among the most popular beverages consumed worldwide, contain many different catechins. We developed an analytical method capable of separating six different catechins and caffeine in tea by micellar electrokinetic chromatography in only 20 min without extensive sample preparation. Furthermore, we compared the amount of catechins and caffeine in several teas and different preparation modes. PMID- 11271480 TI - Rapid determination of fungicides in fruit juices by micellar electrokinetic chromatography: use of organic modifiers to enhance selectivity and on-column high-salt stacking to improve sensitivity. AB - A rapid, reliable method for the multiresidue analysis of eight commonly used fungicides by micellar electrokinetic chromatography (MEKC) was developed. Excellent separation of the eight fungicides (carbendazim, metalaxyl, captan, procymidone, folpet, captafol, vinclozolin and iprodione) is achieved within about 10 min by using optimized electrophoretic conditions that include the addition of a mixture of organic modifiers to the running buffer for improved resolution. The sensitivity of the method is enhanced by using an enrichment step that involves on-column high-salt stacking. Limits of detection in the microgram per-liter region and relative standard deviations from 2.1 to 5.9% are thus obtained for the fungicides without detracting from peak resolution. These results reveal that the high-salt stacking method provides highly improved sensitivity and enables highly flexible adjustment of the selectivity of the separation method. Also, the method surpasses other stacking alternatives used in MEKC and affords routine analyses of fruit juice containing fungicides at trace levels following a straightforward sample treatment. The robustness of the high salt stacking method as demonstrated in this work makes MEKC methods involving stacking procedures an attractive choice for routine analyses. PMID- 11271482 TI - A turning point in proteome analysis: sample prefractionation via multicompartment electrolyzers with isoelectric membranes. AB - Sample prefractionation, as obtained via multicompartment electrolyzers with isoelectric membranes, greatly enhanced the load ability, resolution and detection sensitivity of two-dimensional (2-D) maps in proteome analysis. This was demonstrated with different samples. In an Escherichia coli total cell extract, analysis by a 2-D map run in a pH 4-5 gradient showed many more spots when prefractionated, as compared with standard maps available in databases such as SWISS-2DPAGE. Analysis of human plasma in the pH 3-6 range showed an increase in the number of highly acidic proteins in the fractionated sample compared to whole plasma. With both samples no protein precipitation or smears occurred and much larger sample amounts could be loaded upon prefractionation, so that a large number of spots could be visualized by Coomassie staining, which is fully compatible with subsequent matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) analysis. PMID- 11271483 TI - An immobiline DryStrip application method enabling high-capacity two-dimensional gel electrophoresis. AB - In the field of proteomics the need to detect low-abundance cellular components, such as regulatory proteins, is of critical importance. Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) is one of the most commonly used separation tools for these biological investigations. In this paper we report an alternative micropreparative 2-D PAGE sample application method, called the "paper bridge loading" method. This method makes it possible to apply a larger sample volume to commercially available immobilized pH gradient (IPG) strips. The Vh products required for focusing are only marginally longer than those used in analytical experiments. The method was compared to traditional cup loading and in gel rehydration. With 18 cm long narrow-range Immobiline DryStrip pH 4.5-5.5, the "paper bridge" method allowed the application of 10 mg human plasma proteins compared to 3 mg with traditional loading methods. The corresponding figures using Escherichia coli sample was found to be 6 mg and less than 2 mg, respectively. The paper bridge method also showed the best results in terms of spot resolution and separation of high molecular weight proteins. PMID- 11271484 TI - Postelectrophoretic staining of proteins separated by two-dimensional gel electrophoresis using SYPRO dyes. AB - While the classical silver stain has been the method of choice for high sensitivity protein visualization on two-dimensional gel electrophoresis (2-D PAGE), post-electrophoretic fluorescent staining with the SYPRO group of dyes has emerged to challenge silver staining for proteome analysis. The latter offers improved sensitivity, higher dynamic range and easy handling. However, most of the published data were derived from analysis of 1-D gel separations. In this work, we have focused on three commercially available fluorescent dyes, SYPRO Ruby, SYPRO Orange and SYPRO Red (Molecular Probes, Eugene, OR, USA) and studied their sensitivity and dynamic range on 2-D PAGE. The use of a multiwavelength fluorescent scanner to image 2-D protein profiles visualized with fluorescent staining is discussed, and a detailed comparison with analysis by silver staining is also provided. These results demonstrate the advantages of using SYPRO dyes, which are in agreement with the literature based on 1-D gel electrophoresis, and give a more realistic understanding of the performance of these fluorescent dyes with 2-D PAGE. PMID- 11271485 TI - A modified silver staining protocol for visualization of proteins compatible with matrix-assisted laser desorption/ionization and electrospray ionization-mass spectrometry. AB - The growing availability of genomic sequence information, together with improvements in analytical methodology, have enabled high throughput, high sensitivity protein identification. Silver staining remains the most sensitive method for visualization of proteins separated by two-dimensional gel electrophoresis (2-D PAGE). Several silver staining protocols have been developed which offer improved compatibility with subsequent mass spectrometric analysis. We describe a modified silver staining method that is available as a commercial kit (Silver Stain PlusOne; Amersham Pharmacia Biotech, Amersham, UK). The 2-D patterns abtained with this modified protocol are comparable to those from other silver staining methods. Omitting the sensitizing reagent allows higher loading without saturation, which facilitates protein identification and quantitation. We show that tryptic digests of proteins visualized by the modified stain afford excellent mass spectra by both matrix-assisted laser desorption/ionization and tandem electrospray ionization. We conclude that the modified silver staining protocol is highly compatible with subsequent mass spectrometric analysis. PMID- 11271486 TI - A comparison of silver stain and SYPRO Ruby Protein Gel Stain with respect to protein detection in two-dimensional gels and identification by peptide mass profiling. AB - Proteomic projects are often focused on the discovery of differentially expressed proteins between control and experimental samples. Most laboratories choose the approach of running two-dimensional (2-D) gels, analyzing them and identifying the differentially expressed proteins by in-gel digestion and mass spectrometry. To date, the available stains for visualizing proteins on 2-D gels have been less than ideal for these projects because of poor detection sensitivity (Coomassie blue stain) or poor peptide recovery from in-gel digests and mass spectrometry (silver stain), unless extra destaining and washing steps are included in the protocol. In addition, the limited dynamic range of these stains has made it difficult to rigorously and reliably determine subtle differences in protein quantities. SYPRO Ruby Protein Gel Stain is a novel, ruthenium-based fluorescent dye for the detection of proteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels that has properties making it well suited to high throughput proteomics projects. The advantages of SYPRO Ruby Protein Gel Stain relative to silver stain demonstrated in this study include a broad linear dynamic range and enhanced recovery of peptides from in-gel digests for matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry. PMID- 11271487 TI - Investigating the reaction of a number of gel electrophoresis cross-linkers with beta-lactoglobulin by matrix assisted laser desorption/ionization-mass spectrometry. AB - A number of cross-linkers that are commonly used in polyacrylamide gels have been incubated with bovine beta-lactoglobulin B and the resulting reaction mixtures were examined by matrix assisted laser desorption/ionization-mass spectrometry. At concentrations of 0.1, 1, and 20 mM of each cross-linker incubated for 1 h with 50 pmol/microL of the protein, a reactivity scale can be expressed as polyethylene glycol diacrylate > N,N'-bisacrylylcystamine > bisacrylyl piperazine > N,N'-methylenebisacrylamide >> N,N'-diallyltartardiamide (PEGDA>BAC>BAP>Bis>>DATD). Relatively short incubation times indicated one of the five Cys residues as the target of reaction, which was confirmed by post-source decay measurements. Longer incubation times (24 h) with bisacrylamide extended the reaction to all five Cys residues and a number of Lys residues. A second consequence of longer reaction time is the involvement of both terminals of the cross-linker in the observed reaction. This experimental evidence is the first to demonstrate a different reactivity of both ends of one of the most commonly used cross-linkers. Investigation of solutions containing a cross-linker and acrylamide monomers provided useful information on the competition between the two identities for reaction with the protein. Possible implications of these experimental observations for isoelectric focusing separations in polyacrylamide gels are discussed. PMID- 11271488 TI - Separation and characterization of basic barley seed proteins. AB - Basic proteins in barley starchy endosperm from developing seeds were separated by two-dimensional (2-D) nonequilibrium pH gel electrophoresis. Total as well as partial extracts were analyzed. Edman degradation sequencing and immunological detection were performed after transfer of separated proteins onto membranes. Only one protein could be analyzed by N-terminal sequencing of blotted and separated proteins from the total extract. Fractionation of extracts was done using cation exchange chromatography, concanavalin A and heparin affinity chromatography. Internal sequences were determined after in-gel cleavage of proteins using trypsin or cyanogen bromide and separation of the fragments by reversed-phase chromatography or in a gel electrophoresis system for peptide separation. This resulted in a new protocol for obtaining internal sequences from proteins separated by 2-D electrophoresis. A total of 16 sequences, including nine internal sequences, were analyzed, permitting the identification of ten proteins, including five that appeared to have a blocked N-terminus. An additional protein was identified using immunological detection. Three protein sequences remained unidentified. Separated proteins were also analyzed with a glycan detection method. PMID- 11271489 TI - Alkaline proteins of Bacillus subtilis: first steps towards a two-dimensional alkaline master gel. AB - The genomic sequence of Bacillus subtilis, which is the best studied Gram positive bacterium, enabled us to obtain a theoretical two-dimensional (2-D) map, demonstrating that about one-third of this proteome has a theoretical alkaline isoelectric point (pI). This represents an important part of the entire proteome, which is not detectable in conventional 2-D gels (pH range 4-7). Sequence analysis revealed that 91% of the ribosomal proteins and a high amount of theoretical membrane proteins should be localized in the alkaline pH range requiring different protein extraction procedures. In order to find the pH range which gives the best resolution results for the alkaline proteins of B. subtilis, immobilized pH gradients (IPGs) with different pH ranges (pH 6-10, 6-11, 4-12, 9 12, and 3-10) were tested and optimized for IPG 4-12. Here we present a version of a first alkaline master 2-D gel for B. subtilis, which is a further complement of the already existing master gel (pH 4-7) in the Sub2D database. Almost 150 spots could be detected and 41 proteins have already been identified. PMID- 11271490 TI - Identification of Tuber borchii Vittad. mycelium proteins separated by two dimensional polyacrylamide gel electrophoresis using amino acid analysis and sequence tagging. AB - This paper reports the first results in the proteome analysis of Tuber borchii Vittad. mycelium, an ectomycorrhizal fungus poorly defined genetically, but known for its generation of edible fruit bodies known as white truffles. Employing isoelectric focusing on immobilized pH gradients, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we obtained an electropherogram presenting over 800 spots within the window of isoelectric points (pI) 3.5-9 and a molecular mass of 10-200 kDa. Different reducing agents were tested in the sample preparation buffers, and the standard lysis buffer plus 2% w/v polyvinylpolypyrrolidone allowed the best solubilization and resolution of the proteins. The T. borchii proteins separated in micropreparative gels were electroblotted onto polyvinylidene difluoride membranes and visualized by Coomassie staining. Twenty-three proteins were excised and analyzed by the combination of amino acid and N-terminal analysis. One protein was identified by matching its amino acid composition, estimated isoelectric point and molecular mass against the SWISS-PROT and EMBL databases. Four spots were successfully tagged by Edman microsequencing but no homologous sequences were found in databases. PMID- 11271491 TI - Differences between predicted and observed sequences in Saccharomyces cerevisiae. AB - We recently studied the protein composition of a Saccharomyces cerevisiae wine yeast strain (K310) of enological interest. About 2,500 spots of 8-250 kDa observed molecular mass were resolved by two-dimensional gel electrophoresis. Experimental molecular masses and isoelectric points were calculated for most of them. Twenty-seven proteins were subjected to Edman microsequencing. N-terminal sequences of 12/27 proteins were determined, whereas internal sequences of 6/27 proteins were obtained following in situ proteolysis. Comparison between the experimental data and those reported in the SWISS-PROT database revealed some differences between genotypic and phenotypic sequences. These are indicative of the changes a protein can undergo with respect to the primary structure coded by the genomic DNA. Our results highlight the need to complement genomic analysis with detailed proteomics in order to refine the vast amount of information provided by DNA sequencing and to find an exact correlation between genome and proteome. PMID- 11271492 TI - Investigation of Fasciola hepatica sample preparation for two-dimensional electrophoresis. AB - This paper investigates the preparation of Fasciola hepatica samples for two dimensional electrophoresis (2-DE). Whole samples were prepared by both hot sodium dodecyl sulfate (SDS) solubilisation and precipitation using trichloroacetic acid (TCA) to remove nonprotein contaminants and to inactivate endogenous proteases. Sample preparation had a marked influence on the 2-DE gel profile. TCA precipitation resulted in no measurable improvement in the profile observed, compared to the untreated control. Solubilisation of sample with hot SDS increased the number of protein spots, as did TCA precipitation with the addition of phosphotungstic acid. The preparation of excretory-secretory (ES) products poses problems due to both high salt concentrations and low protein concentration. All precipitation methods used to overcome this gave similar profiles, except acetone alone, which caused depletion of the larger proteins. TCA in acetone gave the best result, similar to that obtained by centrifugal filtration of the sample. Overcrowding of spots in some regions of the 2-DE gel occurred in the whole Fasciola hepatica sample. This problem was alleviated by differential solubilisation, which also resulted in the enrichment of some proteins. PMID- 11271493 TI - Proteome analysis of activated murine B-lymphocytes. AB - Proteins extracted from murine B-lymphocytes after in vitro stimulation by lipopolysaccharide were separated by two-dimensional (2-D) polyacrylamide gel electrophoresis and analyzed by matrix assisted laser desorption/ionization mass spectrometry. Structural information on the protein entities from 153 spots was obtained. Since many of these spots occur as members of spot families, a smaller number --98 genes-- was found to be coding for the identified spots. The elucidated proteins belong to groups of functional categories; we found 26 enzymes, 36 regulatory proteins, 15 chaperones, 15 structural proteins, 4 immunoglobulins, 1 ribosomal and 1 histone protein. A comparison between expected and observed molecular masses yields a good correlation for the majority of the compared spot entities. This set of proteins now identified in the context of a lymphocyte 2-D gel pattern should advance further studies on lymphocyte functions. PMID- 11271494 TI - Towards the proteome of Mycobacterium tuberculosis. AB - Human tuberculosis is caused by the intracellular pathogen Mycobacterium tuberculosis. Sequencing of the genome of M. tuberculosis strain H37Rv has predicted 3924 open reading frames, and enabled identification of proteins from this bacterium by peptide mass fingerprinting. Extracellular proteins from the culture medium and proteins in cellular extracts were examined by two-dimensional gel electrophoresis using immobilized pH gradient technology. By mass spectrometry and immunodetection, 49 culture filtrate proteins and 118 lysate proteins were identified, 83 of which were novel. To date, 288 proteins have been identified in M. tuberculosis proteome studies, and a list is presented which includes all identified proteins (available at http://www.ssi.dk/publichealth/tbimmun). The information obtained from the M. tuberculosis proteome so far is discussed in relation to the information obtained from the complete genome sequence. PMID- 11271495 TI - Proteomics approach in classifying the biochemical basis of the anticancer activity of the new olomoucine-derived synthetic cyclin-dependent kinase inhibitor, bohemine. AB - The aim of this study was to use two-dimensional electrophoresis (2-DE) coupled with multivariate principal component analysis (PCA) to characterize the quantitative changes in the protein composition of the CEM T-lymphoblastic leukemia cell line after treatment with bohemine (BOH), a synthetic olomoucin derived cyclin-dependent kinase inhibitor (CDKI). Cell classification, reflecting protein patterns, clearly distinguished two main groups: one group consists of 9, 12 and 24 h treated BOH cells while the second is represented by the 0 and 24 h control untreated cells and the 6 h BOH-exposed CEM lymphoblasts. Discriminant protein spots differentially expressed in the BOH-treated CEM cells were selected for identification by matrix assisted laser desorption/ionization-mass spectrometry (MALDI-MS) or electrospray ionization-tandem MS (ESI-MS/MS). Five of the selected protein spots were unequivocally identified as alpha-enolase, triosephosphate isomerase, eukaryotic initiation factor 5A, and alpha- and beta subunits of Rho GDP-dissociation inhibitor 1. These proteins, all significantly downregulated in CEM T-lymphoblast leukemia in the course of BOH treatment, are known to play an important role in cellular functions such as glycolysis, protein biosynthesis, and cytoskeleton rearrangement. These results indicate that the cellular effects of olomoucine-derived CDKIs are not dependent on their ability to inhibit CDKs and could be mediated by several factors such as a decrease in protein synthesis and/or glycolysis which in turn diminishes the ability of cancer cells to function. PMID- 11271496 TI - Towards a two-dimensional proteome map of Mycoplasma pneumoniae. AB - A Proteome map of the bacterium Mycoplasma pneumoniae was constructed using two dimensional (2-D) gel electrophoresis in combination with mass spectrometry (MS). M. pneumoniae is a human pathogen with a known genome sequence of 816 kbp coding for only 688 open reading frames, and is therefore an ideal model system to explore the scope and limits of the current technology. The soluble protein content of this bacterium grown under standard laboratory conditions was separated by 1-D or 2-D gel electrophoresis applying various pH gradients, different acrylamide concentrations and buffer systems. Proteins were identified using liquid chromatography-electrospray ionization ion trap and matrix-assisted laser desorption/ionization-MS. Mass spectrometric protein identification was supported and controlled using N-terminal sequencing and immunological methods. So far, proteins from about 350 spots were characterized with MS by determining the molecular weights and partial sequences of their tryptic peptides. Comparing these experimental data with the DNA sequence-derived predictions it was possible to assign these 350 proteins to 224 genes. The importance of proteomics for genome analysis was shown by the identification of four proteins, not annotated in the original publication. Although the proteome map is still incomplete, it is already a useful reference for comparative analyses of M. pneumoniae cells grown under modified conditions. PMID- 11271497 TI - Human ERp29: isolation, primary structural characterisation and two-dimensional gel mapping. AB - Recently we characterised a novel 29 kDa endoplasmic reticulum protein that is widely expressed in rat tissues, and named it ERp29. Several ERp29-like gene products have been reported in human tissues but uncertainty surrounds their relationships with each other and rat ERp29. To clarify these issues, ERp29 was isolated from human liver and characterised by primary structural analysis and two-dimensional gel mapping. Comparisons with rat ERp29 revealed striking homologies both in sequence and physical properties. Characterisation of the isoelectric heterogeneity and anomalous mass on two-dimensional gels enabled two reported homologues (UL35 and ERp31) to be identified as ERp29. Resolution of a sequence discrepancy led to unequivocal correlation of human ERp29 with the cognate cDNA previously named ERp31 and ERp28. Consequent links established to human genome and proteome projects showed that ERp29 is encoded by a gene on chromosome 12 that is expressed universally in human tissues. Together, these findings unified various ERp29 homologues as products of a single gene orthologous to rat ERp29 and established ERp29 as the only known member of a new protein class. Investigations of ERp29 function in human health and disease should benefit from the integrated links between genome, proteome and murine model organisms established here. PMID- 11271498 TI - Complementing genomics with proteomics: the membrane subproteome of Pseudomonas aeruginosa PAO1. AB - With the completion of many genome projects, a shift is now occurring from the acquisition of gene sequence to understanding the role and context of gene products within the genome. The opportunistic pathogen Pseudomonas aeruginosa is one organism for which a genome sequence is now available, including the annotation of open reading frames (ORFs). However, approximately one third of the ORFs are as yet undefined in function. Proteomics can complement genomics, by characterising gene products and their response to a variety of biological and environmental influences. In this study we have established the first two dimensional gel electrophoresis reference map of proteins from the membrane fraction of P. aeruginosa strain PA01. A total of 189 proteins have been identified and correlated with 104 genes from the P. aeruginosa genome. Annotated membrane proteins could be grouped into three distinct categories: (i) those with functions previously characterised in P. aeruginosa (38%); (ii) those with significant sequence similarity to proteins with assigned function or hypothetical proteins in other organisms (46%); and (iii) those with unknown function (16%). Transmembrane prediction algorithms showed that each identified protein sequence contained at least one membrane-spanning region. Furthermore, the current methodology used to isolate the membrane fraction was shown to be highly specific since no contaminating cytosolic proteins were characterised. Preliminary analysis showed that at least 15 gel spots may be glycosylated in vivo, including three proteins that have not previously been functionally characterised. The reference map of membrane proteins from this organism is now the basis for determining surface molecules associated with antibiotic resistance and efflux, cell-cell signalling and pathogen-host interactions in a variety of P. aeruginosa strains. PMID- 11271499 TI - Cross-matching marsupial proteins with eutherian mammal databases: proteome analysis of cells from UV-induced skin tumours of an opossum (Monodelphis domestica). AB - The identification and characterisation of Monodelphis proteins has required cross-species analysis. Protein expression was investigated in normal, nonirradiated adult fibroblasts and also in fibroblastic cells from a benign cutaneous tumour after chronic ultraviolet (UVB) exposure and a metastatic cutaneous tumour after intermittent exposure. Proteins were separated and visualised by two-dimensional gel electrophoresis (2-D PAGE) and a peptide mass fingerprint (PMF) was obtained for protein spots using matrix assisted laser desorption/ionisation-time of flight-mass spectrometry (MALDITOF-MS). Cross species PMF database analysis facilitated the identification of 120 proteins, constituting 46.5% of the proteins analysed. The identification of two proteins was confirmed by internal amino acid sequencing using tandem MS. Differential protein expression was observed between normal fibroblasts and those in tumours chronically or intermittently exposed. A number of tropomyosin and vimentin isoforms were expressed only in cells from the metastatic tumour induced by intermittent exposure to UV radiation. These results highlight the value of cross species PMF analysis for the rapid characterisation of proteins from a poorly defined species and also show how proteomics can be used to detect changes in protein expression in differentially treated cells. PMID- 11271500 TI - Identification of nolR-regulated proteins in Sinorhizobium meliloti using proteome analysis. AB - Extractable proteins from Sinorhizobium meliloti strains AK631 and EK698 (a Tn5 induced noIR-deficient mutant of AK631), grown in tryptone agar (TA) medium with or without the addition of the plant signal luteolin, were separated by two dimensional gel electrophoresis and compared. Analysis of silver-stained gels showed that the noIR mutant had 189 proteins that were significantly altered in their levels (101 protein spots up- and 88 downregulated). Coomassie-stained preparative two-dimensional (2-D) gels or polyvinylidene difluoride (PVDF) membranes blotted from preparative gels showed that at least 52 of the altered proteins could be reproducibly detected and isolated from the noIR mutant. These 52 altered protein spots were classified into five groups based on an assessment of protein abundance by computer analysis and the effect of the presence or absence of luteolin addition to the growth medium. N-terminal microsequencing of 38 proteins revealed that the most striking feature of the consequence of the noIR mutation is the number and broad spectrum of cellular functions that are affected by the loss of the NoIR function. These include proteins involved in the tricarboxylic acid (TCA) cycle, heat shock and cold shock proteins, protein synthesis, a translation elongation factor, oxidative stress and cell growth and maintenance functions. We propose that the NoIR repressor is a global regulatory protein which responds to environmental factors to fine-tune intracellular metabolism. PMID- 11271502 TI - Characterization of the pore structure of aqueous three-dimensional polyacrylamide gels with a novel cross-linker. AB - The properties of polyacrylamide hydrogels synthesized with a novel hexafunctional (three double bonds) cross-linker, hexahydro-1,3,5-triacryloyl-s triazine (1a), was evaluated and compared to the currently used tetrafunctional (two double bonds) cross-linker N,N-methylenebisacrylamide (Bis). A variety of characterization techniques that require very little sample preparation and data handling were chosen and include polymerization temperature profiles and conversions, water swelling, differential scanning calorimetry (DSC), polyacrylamide gel electrophoresis (PAGE), Gradiflow electrophoretic separation process and scanning electron microscopy (SEM). The alternative use of 1a compared to Bis results in polyacrylamide gels with larger pore sizes and a broad pore size distribution. PMID- 11271501 TI - Proteome analysis demonstrates complex replicon and luteolin interactions in pSyma-cured derivatives of Sinorhizobium meliloti strain 2011. AB - Sinorhizobium meliloti was studied by proteomic analysis to investigate the contribution made by plasmid-encoded functions on the intracellular regulation of this bacterium. Protein profiles of strain 2011 were compared with those from its mutant strains which were either cured of their pRme2011a (also called pSyma) plasmid (strain 818), or contained an extensive deletion of this plasmid (strain SmA146). Plasmid pSyma contains the nodulation and nitrogen fixation genes and is 1.4 Mbp with an estimated coding potential of 1,400 proteins. However, under the growth conditions used we could detect 60 differences between the parent strain and its pSyma-cured derivative, strain 818. While the majority of these differences were due to regulatory changes, such as up- and downregulation, some proteins were totally missing in some strains. These 60 proteins were classified into 21 subgroups, A to U, based on their measured protein levels when the cells were grown in the presence or absence of luteolin. Comparisons were made between the different strains to assess the possible interactions of the different proteins of the subgroups and plasmid pSyma. These results suggest that pSyma has a role in the regulation of the expression of genes from the other replicons (3.5 Mbp chromosome and the 1.7 Mbp pSymB plasmid) present in the S. meliloti cells. Proteome analysis provides a sensitive tool to examine the functional organisation of the S. meliloti genome and the intracellular gene interactions between replicons and will provide a powerful analytical tool to complement the genome sequencing of strain 1021. PMID- 11271503 TI - Growth arrest-specific gene 6 expression in proliferating rabbit vascular smooth muscle cells in vitro and in vivo. AB - Proliferation and migration of vascular smooth muscle cells (VSMCs) are involved in the processes of atherosclerosis and restenosis. The protein product of the growth arrest-specific gene 6 (Gas-6) has recently been identified as a ligand for the Axl/Rse/Mer tyrosine kinase receptor family, which may be involved in proliferation and migration of VSMCs. Here we show that Gas-6 gene expression is increased in proliferating VSMCs in tissue culture (2.5-fold increase by Northern blot) and following neointimal proliferation in a rabbit balloon-injury model (3 fold increase by Western blot). Neither platelet-derived growth factor (PDGF) nor thrombin stimulate the expression of Gas-6 in cultured VSMCs despite the ability of the PDGF, but not thrombin, to stimulate proliferation in growth-arrested cells. These data suggest a role for the Gas-6 regulatory system in VSMC proliferation, which may be a target for therapeutic interventions in the atherosclerotic process and restenosis after angioplasty. PMID- 11271504 TI - Determination of P2X1alpha-sarcoglycan (adhalin) expression levels in failing human dilated cardiomyopathic left ventricles. AB - This study is concerned with the molecular basis of human idiopathic dilated cardiomyopathy (DCM). This disorder affects the entire heart including both atria and ventricles. It is characterized by a progressive dilatation of the ventricles and loss of contractile power that results in an impaired cardiac output. Changes in cellular levels of dystrophin have been reported in patients with muscular dystrophies (Beckers and Duchenne) which manifest as DCM. However, previous studies using Western blots dos Remedios et al., Electrophoresis 1996, 17, 235 238) of samples of left ventricles from DCM patients showed no abnormalities in dystrophin content. P2X receptors are ATP-gated cation channels located in the sarcolemma. They are upregulated by a factor of about two in the atria of DCM patients compared with nondiseased control samples. A dystrophin-associated protein, alpha-sarcoglycan, has recently been shown to be an ecto-ATPase (an extracellular ATPase) capable of regulating ATP concentrations in the space between the cardiomyocytes. In this report we examine the relationship between changes in P2X1 receptors in left ventricle samples from DCM patients and the concentration of alpha-sarcoglycan. We found no evidence for upregulation of P2X1 receptors nor was the expression of alpha-sarcoglycan significantly altered. PMID- 11271505 TI - The role of ATP, ADP and divalent cations in the formation of binary and ternary complexes of actin, cofilin and DNase I. AB - Actin is the major cytoskeletal protein of virtually all eukaryotic cells. Actin assembly/disassembly is involved in a variety of cellular processes and actin binding proteins are essential in regulation of the pool of actin monomers. Cofilin and DNase I are actin-binding proteins, which form both binary (actin DNase 1, cofilin-actin) and ternary (cofilin-actin-DNase I) complexes with actin. Here we use native gel electrophoresis to examine the roles of ATP, ADP, Ca2+ and Mg2+ in the formation of these complexes as well as on the ability of actin to self-assemble. Conditions which favour actin polymerisation are: ATP (no Me2+) > or = ADP (no Me2+) > ADP-Ca2+ = ADP-Mg2+ > ATP-Mg2+ > ATP-Ca2+. Preferential conditions for the formation of the binary actin-cofilin complex are: ADP-Mg2+ > or = ADP-Ca2+ >> ATP-Ca2+ approximately equals ATP-Mg2+ approximately equals ADP No Me2+ approximately equals ATP-No Me2+. Actin forms a very tight complex with DNase I in the order: ATP-Ca2+ > or = ATP-Mg2+ approximately equals ADP-Mg2+ approximately equals ADP-Ca2+ > or = ADP-(no Me2+) > ATP-(no Me2+). Effectively, the complex does not form in the presence of ATP and the absence of free Me2+. Finally, the conditions which favour the formation of a ternary complex of cofilin-actin-DNase I resemble the actin-DNase I, namely: ATP-Ca2+ approximately equals ADP-Ca2+ approximately equals ADP-Mg2+ approximately equals ATPMg2+ ADP (no Me2+) > ATP-(no Me2+). PMID- 11271507 TI - A probabilistic model for the establishment of neuron polarity. AB - The main aim of this paper is to present a simple probabilistic model for the early stage of neuron growth: the specification on an axon out of several initially similar neurites. The model is a Markov process with competition between the growing neurites, wherein longer objects have more chances to grow, and parameter alpha determines the intensity of the competition. For alpha > 1 the model provides results which are qualitatively similar to the experimental ones, i.e. selection of one rapidly elongating axon out of several neurites while other less successful neurites stop growing at some random time. Rigorous mathematical proofs are given. PMID- 11271506 TI - Stochastic population models with interacting species. AB - A stochastic model concerning the evolution of a multi-species population is presented assuming species competition for a habitat. The model takes into account colonization, death and replacement for all individuals. Two cases are treated: (i) colonizations follow the hierarchic rule by which species of lower rank are always outcompeted by those of higher rank and (ii) there are no privileged species. In both cases, under suitable assumptions, a thorough description of the evolution of the population is obtained. The two models are finally compared and the corresponding evolutionary behaviors of the populations are discussed. PMID- 11271508 TI - Simulation of 'hitch-hiking' genealogies. AB - An ancestral influence graph is derived, an analogue of the coalescent and a composite of Griffiths' (1991) two-locus ancestral graph and Krone and Neuhauser's (1997) ancestral selection graph. This generalizes their use of branching-coalescing random graphs so as to incorporate both selection and recombination into gene genealogies. Qualitative understanding of a 'hitch hiking' effect on genealogies is pursued via diagrammatic representation of the genealogical process in a two-locus, two-allele haploid model. Extending the simulation technique of Griffiths and Tavare (1996), computational estimation of expected times to the most recent common ancestor of samples of n genes under recombination and selection in two-locus, two-allele haploid and diploid models are presented. Such times are conditional on sample configuration. Monte Carlo simulations show that 'hitch-hiking' is a subtle effect that alters the conditional expected depth of the genealogy at the linked neutral locus depending on a mutation-selection-recombination balance. PMID- 11271509 TI - Periodic solutions in a model of competition between plasmid-bearing and plasmid free organisms in a chemostat with an inhibitor. AB - We obtain necessary and sufficient conditions on the existence of a unique positive equilibrium point and a set of sufficient conditions on the existence of periodic solutions for a 3-dimensional system which arises from a model of competition between plasmid-bearing and plasmid-free organisms in a chemostat with an inhibitor. Our results improve the corresponding results obtained by Hsu, Luo, and Waltman [1]. PMID- 11271510 TI - Rethinking maternal-fetal conflict: gender and equality in perinatal ethics. PMID- 11271511 TI - Outpatient cervical ripening with intravaginal misoprostol. PMID- 11271513 TI - A comparative experimental study of the effects of diclofenac and ketoprofen on the small-bowel mucosa of canines. AB - The aim of this experimental study was to investigate the effect of diclofenac sodium and ketoprofen, two non-steroidal anti-inflammatory drugs (NSAIDs) with different excretion pathways, and the role of other enteric factors during simultaneous administration of these drugs on the development of mucosal lesions of the small intestine in canines. Twenty-five animals were divided into three groups. Group I included 10 canines, 5 with diclofenac sodium (group Ia) and 5 with ketoprofen administration (group Ib). Group II included 5 animals in which a segment of ileum was surgically isolated from the rest of the small intestine. Group III included 10 animals in two subgroups of 5; a segment of ileum was surgically isolated in both subgroups; groups IIIa received diclofenac and group IIIb ketoprofen. Histological examination of the specimens taken revealed macroscopic and microscopic mucosal lesions in 5/5 animals in group Ia, whereas none of the 5 animals in group Ib had any lesions. Group II did not reveal any mucosal lesions. Three out of 5 animals (60%) administered diclofenac in group IIIa had intestinal mucosal lesions, but none of the 5 revealed lesions in the isolated loop of ileum. No lesions were observed in the isolated loop or in the rest of the intestinal mucosa in the animals in group IIIb. Our results suggest that NSAIDs produce intestinal mucosal lesions not only when administered per mouth but also after intramuscular administration. Diclofenac, unlike ketoprofen, was responsible for the development of lesions in the intestinal mucosa. The role of drugs and/or their metabolites in the intestine and certain other factors must still be determined. PMID- 11271512 TI - Isolated working rat heart adaptation after abrupt changes in extracellular Ca2+ concentration. AB - In isolated working rat hearts, a rapid up- or downward change in perfusate Ca2+ concentration by the factors 2, 4, or 8 in the range between 1.25 and 10 mM resulted within 1 min in the well-known change of left ventricular contractility as evaluated by maximum left ventricular pressure change velocity (LVdP/dt max) and left ventricular end-diastolic pressure (LVEDP). However, within about 10 min thereafter, contractility showed an adaptive behaviour opposite to the initial change, with t 1/2 values for LVdP/dt max between 1.45 and 2.8 min. The adaptive LVdP/dt max reactions amounted to 10-35% of the initial change. With an abrupt fall from 10 mM to 1.25 mM Ca2+ (not tolerated by all hearts), LVdP/dt max decreased initially by 4.260 mmHg/s (LVEDP +9.7 mmHg) and increased thereafter by 524 mmHg/s (LVEDP -6.8 mmHg). The adaptive inotropic behaviour cannot be related to changes in heart rate or coronary flow and is not affected by thapsigargin (1 microM) or the amiloride derivative benzamil (10 microM). This suggests that sarcoplasmatic Ca2+-ATPase and sarcolemmal Na+-Ca2+ exchange do not play a decisive role in this adaptive behaviour. In conclusion, an intrinsic regulatory mechanism of the myocardium attenuates the inotropic effect of acute changes in Ca2+ concentration. This phenomenon might protect the heart against Ca2+ overload after an acute rise in catecholamine concentration. PMID- 11271514 TI - May pentoxifylline improve lung function after one-lung flooding? AB - One-lung flooding makes the intraoperative sonography of round pulmonary lesions possible. During the flooded phase, the flooded lung suffers a significant reduction in perfusion. After ischemia and reperfusion, neutrophil granulocytes lead to further tissue injuries. A study was made on four animals to determine whether administration of pentoxifylline--a potent inhibitor of granulocyte adhesion to the endothelium--improves lung function after one-lung flooding. Two animals were subjected to thoracotomy with extended hemodynamic monitoring. Thoracoscopy was performed on two other animals, which were extubated after the flooding liquid was drained and survived for 24 degrees h. A bolus of 1 mg/kg of pentoxifylline was administered at the time of thoracotomy/thoracoscopy. followed by continuous infusion of pentoxifylline at a rate of 1.5 mg/kg per hour until 30 degrees min after reventilation (thoracotomy), or until extubation, respectively. The control group consisted of animals employed in previous experiments. Except for pentoxifylline administration, they were subjected to identical experimental conditions. The control group for the thoracotomy experiment comprised 14 animals, that for the thoracoscopy experiment three animals. The experiments proved that 30 degrees min after the draining of the flooding liquid and reventilation, all four pentoxifylline-treated animals had a higher partial arterial oxygen pressure and a lower pulmonary shunt volume compared with the control animals. In the two animals that survived, a positive effect on lung function was no longer detectable 24 degrees h after extubation. The administration did not lead to a drop in the pulmonary arterial pressure and did not cause any hemodynamic changes other than a moderate tachycardia. PMID- 11271515 TI - Localization of annexin V in rat normal kidney and experimental glomerulonephritis. AB - The localization of annexin V, a calcium binding protein, was immunochemically and immunohistologically studied in experimental rat glomerulonephritis using annexin V polyclonal antibody. Plasma and urinary annexin V levels were measured by a sandwich enzyme-linked immunosorbent assay (ELISA). Urinary annexin V level, which was correlated with urinary L-lactate dehydrogenase activity, N-acetyl-beta D-glucosaminidase activity and protein level, increased time-dependently after the injection of nephritogenic antigen (bovine glomerular basement membrane), progressively increasing to attain a peak level at 4 weeks of 51.5 +/- 11.3 ng/h. However, plasma annexin V level showed no increase during the study period. Normal kidneys showed strong staining for annexin V in distal tubules, being particularly strong in tubules of the inner stripe of the outer medulla, but could not be detected in proximal tubules. Annexin V was seen in visceral epithelial cells. Bowman's capsule of the glomerulus, the vascular endothelium of arterioles and interlobular arteries, and vascular smooth muscle. In nephritis, the lumen of distal tubules and the luminal cell membrane were deeply stained, with leakage of annexin V being observed from tubular cells. In the present study, renal annexin V was markedly excreted into urine, and its urinary level reflected the severity of damage of renal tissue and the progression of nephritis. These changes of annexin V in the distal tubule and visceral epithelial cells may be of significance in cell injury of the kidney. PMID- 11271516 TI - Ischemia at 4 degrees C: a novel mouse model to investigate the effect of hypothermia on postischemic hepatic microcirculatory injury. AB - Hypothermia of the ischemic organ at 4 degrees C protects hepatic microcirculation from ischemia-reperfusion (IR) injury. The effect of hypothermia during ischemia was investigated in animal models using liver transplantation and storage of the harvested organ in cold preservation solutions. No investigation of the isolated influence of hypothermia at 4 degrees C of the ischemic organ on hepatic IR injury exists, due to the lack of an appropriate animal model. Therefore, the aim of our present study was to develop such a model using intravital video fluorescence microscopy (IVM). In C57BL/6 mice, a reversible isolated ischemia of the left liver lobe was induced for 90 min, followed by 240 min of reperfusion. The temperature of the ischemic organ was adjusted to either 4 degrees C or 37 degrees C by superfusion with 0.9% NaCl. Sham-operated animals without IR served as controls. The hepatic microcirculation was analyzed using IVM at 30 min and 240 min after reperfusion by quantifying sinusoidal perfusion and leukocyte-endothelial cell interaction in postsinusoidal venules. At the end of the experiment, blood and tissue samples were taken for measurement of liver enzyme activities and light and electron microscopy. Mean arterial pressure and body temperature were kept constant throughout the experiment, while the temperature of the ischemic liver lobe was adjusted to predefined levels. After normothermic ischemia, hepatic microvascular perfusion was significantly impaired compared with sham-operated animals. Perfusion failure was significantly reduced in hypothermic livers and did not differ from livers of the sham-group. Liver enzyme activities in the normotherimic group were significantly higher than in the sham and hypothermic groups. Light and electron microscopy revealed severe histological alterations at 37 degrees C ischemia, whereas at 4 degrees C ischemia only minimal lesions were encountered. Our novel model allows for isolated adjustment of ischemic liver lobe temperature without changing body temperature and systemic macrohemodynamic parameters. Hypothermia at 4 degrees C largely attenuates postischemic microvascular perfusion injury of the liver. PMID- 11271517 TI - Dendrimers and hyperbranched polymers as high-loading supports for organic synthesis. AB - Polymeric supports have become a big necessity for automated synthesis and combinatorial chemistry, yet, the loading capacities of most polystyrene resins are very limited (typically < 1.5 mmol x g(-1)). Dendrimers and hyperbranched polymers have been discussed for this application and now became readily available. These soluble polymers can either be used directly as high-loading supports for substrates, reagents, and catalysts or alternatively in hybrid polymers linked to conventional polystyrene resins. PMID- 11271518 TI - Mixed dimer and mixed trimer complexes of nBuLi and a chiral lithium amide. AB - Multinuclear and multidimensional NMR spectroscopy have shown that lithium (S)-N isopropyl-O-methyl-valinol (1-[6Li]) exists in a mixed 2:1 complex with nBu[6Li], (1-[6Li])2/nBu[6Li], in non-coordinating solvents such as hexane or toluene. A 6Li,1H-HOESY NMR spectrum indicates that the complex is a cyclic trimer with a large distance between the di-coordinated lithium and the carbanion of nBu[6Li]. Such arrangements are present in the solid state as previously reported by Williard and Sun. The exchange of lithium atoms within the trimer is slow at -33 degrees C. The exchange barrier (deltaG++) was determined to be 14.7 kcal x mol( 1) from quantitative 6Li,6Li-EXSY spectra. Addition of diethyl ether results in the formation of mixed dimers of (1-[6Li])/nBu[6Li], tetramers of nBu[6Li], and homodimers (1-[6Li])2. The apparent equilibrium constant of the mixed dimer was determined from the 6Li NMR integrals as K = 7. PMID- 11271519 TI - An experimental and theoretical study of the basicity of tetra-tert butyltetrahedrane. AB - The gas-phase basicity (GB) of tetra-tert-butyltetrahedrane (tBu4THD) was determined by FT-ICR mass spectrometry and comparison with reference compounds of known basicity. Its GB, 1035+/-10 kJ x mol(-1), makes tetra-tert butyltetrahedrane one of the strongest bases reported so far. Ab initio calculations [B3LYP/6-31G(d) and B3LYP/6-311 + G(d,p)//6-31G(d)] have been carried out in order to compare the high experimental basicity of tBu4THD with that estimated theoretically. Both B3LYP/6-31G(d) and QCISD(T) calculations were used to determine the reaction path which connects the initial tetrahedrane ammonium complex with the final products, protonated cyclobutadiene (CBDH+) and ammonia. PMID- 11271520 TI - [cyclo-CH2[Sn(CI2)CH2Si(Me2)]2O]: synthesis and complexation behaviour of a novel, cyclic, bidentate Lewis acid and its conversion into a tin-containing fluorosilane with intermolecular Si-F...Sn bridges. AB - Acid-catalysed hydrolysis of [CH2[(Sn(Ph2)CH2Si(OiPr)Me2]2] followed by subsequent reaction with mercuric chloride in acetone afforded the novel silicon- and tin-containing eight-membered ring [cyclo-CH2[Sn(Cl2)CH2Si(Me2)]2O] in good yield, the crystal structure of which is reported. 119Sn NMR and X-ray studies indicate that [cyclo-CH2[Sn(Cl2)CH2Si(Me2)]2O] acts as a bidentate Lewis acid towards chloride ions exclusively forming the 1:1 complex [(Ph3P)2N]+[cyclo CH2[Sn(Cl2)CH2Si(Me2)]2OCl]- upon addition of [(Ph3P)2N]+Cl- . Also reported are the synthesis and structure of [K(dibenzo[18]crown-6)]+[cyclo CH2(Sn(Cl2)CH2Si(Me2)]2OF]-, the first completely characterised organostannate with a C2SnCl2F- substituent pattern. No ring-opening polymerisation could be achieved for [cyclo-CH2[Sn(Cl2)CH2Si(Me2)]2O] or for its perphenylated derivative [cyclo-CH2[Sn(Ph2)CH2Si(Me2)]2O]. The reaction of [cyclo CH2[Sn(Cl2)CH2Si(Me2)]2O] with Me3O+BF4- gave the tin-containing fluorosilane [CH2[Sn(Cl2)CH2Si(F)Me2]2], in which the Si-F bond is activated by intermolecular Si-F...Sn interactions in the solid state. PMID- 11271521 TI - The 2-naphthylmethyl (NAP) group in carbohydrate synthesis: first total synthesis of the GlyCAM-1 oligosaccharide structures. AB - Total syntheses of the GlyCAM-1 (glycosylation-dependent cell adhesion molecule 1) oligosaccharide structures: [alpha-NeuAc-(2 --> 3)-beta-Gal-(1 --> 4)-[alpha Fuc-(1 --> 3)]-beta-(6-O-SO3Na)-GlcNAc-(1 --> 6)]-[alpha-NeuAc-(2 --> 3)-beta-Gal (1 --> 3)]-alpha-GalNAc-OMe (1) and [alpha-NeuAc-(2 --> 3)-beta-Gal-(1 --> 4) [alpha-Fuc-(1 --> 3)]-beta-GlcNAc-(1 --> 6)]-[alpha-NeuAc-(2 3)-beta-Gal-(1 --> 3)]-alpha-GalNAc-OMe (2) through a novel sialyl LewisX tetrasaccharide donor are described. Employing sequential glycosylation strategy, the starting trisaccharide was regio- and stereoselectively constructed through coupling of a disaccharide imidate with the monosaccharide acceptor phenyl-6-O-naphthylmethyl-2 deoxy-2-phthalimido-1-thio-beta-D-glucopyranoside with TMSOTf as a catalyst without affecting the SPh group. The novel sialyl Lewisx tetrasaccharide donor 3 was then obtained by alpha-L-fucosylation of trisaccharide acceptor with the 2,3,4-tri-O-benzyl-1-thio-beta-L-fucoside donor. The structure of the novel sialyl Lewisx tetrasaccharide was established by a combination of 2D DQF-COSY and 2D ROESY experiments. Target oligosaccharides 1 and 2 were eventually constructed through heptasaccharide which was obtained by regioselective assembly of advanced sialyl Lewisx tetrasaccharide donor 3 and a sialylated trisaccharide acceptor in a predictable and controlled manner. Finally, target heptasaccharides 1 and 2 were fully characterized by 2D DQF-COSY, 2D ROESY, HSQC, HMBC experiments and FAB mass spectroscopy. PMID- 11271523 TI - Reactivity of calix-tetrapyrrole SmII and SmIII complexes with acetylene: isolation of an "N-confused" calix-tetrapyrrole ring. AB - The nature of the substituents present on the calix-tetrapyrrole tetra-anion ligand [[R2C(C4H2N)]4]4- (R = [-(CH2)5-]0.5, Et) determines the type of reactivity of the corresponding SmII compounds with acetylene. With R = [-(CH2)5 ]0.5, dehydrogenation occurred to yield the nearly colorless dinuclear diacetylide complex [[[[-(CH2)5-]4-calix-tetrapyrrole]SmIII]2(mu-C2Li4)].THF as the only detectable reaction product. Conversely, with R = Et, acetylene coupling in addition to dehydrogenation resulted in the formation of a dimeric butatrienediyl enolate derivative [[(Et8-calix-tetrapyrrole)SmIII[Li[Li(thf)]2(mu OCH=CH2)]]2(mu,eta2,eta'2-HC=C=C=CH)]. Reaction of the trivalent hydride [(Et8 calix-tetrapyrrole)(thf)SmIII[(mu-H)[Li(thf)]]2 or of the terminally bonded methyl derivative [(Et8-calix-tetrapyrrole)(CH3)SmIII[[Li(thf)]2[Li(thf)2](mu3 Cl)]] with acetylene resulted in a mixture of the carbide [[(Et8-calix tetrapyrrole)SmIII]2(mu-C2Li4)].Et2O with the dimerization product [[(Et8-calix tetrapyrrole)SmIII[Li[Li(thf)]2(mu3-OCH=CH2)]]2-mu,eta2,eta'2-HC=C=C=CH)]. The same reaction also yielded a third product, a trivalent complex [[(Et8-calix tetrapyrrole)SmIII[Li(thf)2]]2], in which the macrocycle was isomerized by shifting the ring attachment of one of the four pyrrole rings. PMID- 11271522 TI - Highly efficient synthesis of linear pyrrole oligomers by twofold Heck reactions. AB - The twofold Heck reaction of the vinylpyrroles 3a and 3b with the iodobenzenes 4a c led to the linear pyrrole oligomers 5, 6, and 7. The synthesis of both symmetrical and unsymmetrical oligomers, such as 10a and 10b, was also accomplished by a Heck reaction of 8 and 9 and by a Heck reaction of 3a and 11 followed by a Wittig reaction and a second Heck reaction with 8. The pentacyclic oligomers 14 and 19 were prepared by a twofold Heck reaction of 13 with 4 and by a twofold Heck reaction of 15 with 16 followed by a Wittig reaction and a twofold Heck reaction with 8. PMID- 11271524 TI - Two-dimensional [TIn2] polyanions in BaTIn2 (T=Rh, Pd, Ir, Pt)--the collapse of the three-dimensional indium polyanion of BaIn2. AB - The title compounds were prepared from the elements by reactions in sealed tantalum tubes in a high-frequency furnace. The four compounds were investigated by X-ray diffraction both on powders and single crystals, and the structures of the rhodium and platinum compounds were refined from single-crystal data: Cmcm, a = 447.68(8), b = 1131.1(2), c = 805.6(2) pm, wR2 = 0.0561, 354 F2 values for BaRhIn2; a = 452.06(8), b = 1162.4(2). c = 801.5(1) pm, wR2 = 0.1427, 362 F2 values, for BaPtIn2: with 16 variables for each refinement. The structures are isopointal to MgCuAl2 and can be considered to be a transition metal (T) filled CaIn2 type, in which the indium atoms form a distorted network like hexagonal diamond (lonsdaleite). The indium substructure is cut apart in BaTIn2 and resembles together with the transition metal atoms a two-dimensional polyanion rather than a three-dimensional polyanion as found in the compounds CaTIn2, CaTSn2, and SrTIn2. Semiempirical band structure calculations support the assumption of a two-dimensional polyanion in which the strongest interactions are found for the T-In contacts. PMID- 11271525 TI - The self-assembly of a lipophilic guanosine nucleoside into polymeric columnar aggregates: the nucleoside strucutre contains sufficient information to drive the process towards a strikingly regular polymer. AB - Lipophilic guanosine derivatives act as self-assembled ionophores. In the presence of alkali metal ions in organic solvents, these G derivatives can form tubular polymeric structures. The molecular aggregates formed by 3',5'-didecanoyl 2'-deoxyguanosine (1) have been characterised by SANS and NMR spectroscopy. The polymer is structured as a pile of stacked G quartets held together by the alkali metal ions that occupy the column's central channel. The deoxyribose moieties, with their alkyl substituents, surround the stacked G quartets, and the nucleoside's long-chain alkyl tails are in intimate contact with the organic solvent. In this polymeric structure, there is an amazing regularity in the rotamers around the glycosidic bond within each G quartet and in the repeat sequence of the G quartets along the columns. In hydrocarbon solvents, these columnar aggregates form lyomesophases of the cholesteric and hexagonal types. PMID- 11271526 TI - Determination of absolute configuration of (pi-allyl) palladium complexes by NMR spectroscopy and stereoselective complexation. AB - The chiral chelating ligand N,N'-bis(phenylethyl)bispidine (1) forms a rigid cavity which accommodates (pi-allyl)palladium species with high selectivity. In the resulting complex, the absolute configuration of the pi-allyl ligand can be determined by the detection in NMR spectra of a few unambiguous interligand NOEs. Dynamic processes involving the pi-allyl ligand can be investigated. Depending on the analytical target, ligand (S,S)-1 or (R,R)-1 may be used. PMID- 11271527 TI - The symmetry-broken formalism applied to the electronic structure of an iminosemiquinone copper(II) catalyst: a key to the qualitative understanding of its reactivity. AB - A concise outline of the known derivation of the singlet-triplet energy-gap equations within the symmetry-broken wavefunction framework is given. They allow a computation of the singlet-triplet energy gap for molecules that exhibit a weak antiferromagnetic coupling of electrons. The accuracy of the equations is assessed by computation of the singlet-triplet gaps in model Na2 molecules. Various antiferromagnetic coupling strengths are simulated by the use of different Na-Na bond lengths in the computations. The singlet-triplet energy gaps obtained with the different equations are compared with the gaps computed with the more accurate coupled-cluster methods. Subsequently, the equations are applied to an iminosemiquinone copper(II) complex found previously to have remarkable catalytic properties. The application is performed by employing wave function equations but with quantities computed within the density functional framework. The electronic ground state of this complex is computed to be a singlet state, which is also the experimental finding. Moreover, the experimental geometry and the singlet-triplet gap are reasonably reproduced by the computation. A straightforward method to determine the magnetic orbitals is suggested and applied. We illustrate that the form of the magnetic orbitals indicates in a qualitative manner that hydrogen-atom abstraction should be a major reaction pathway of the iminosemiquinone copper(II) complex. Hydrogen-atom abstraction has been suggested previously to be the rate-determining step in a catalytic process initiated by the iminosemiquinone copper(II) complex. The results support the notion that the form of the magnetic orbitals might be a qualitative indicator for the reactivity of molecules that exhibit weak antiferromagnetic coupling. PMID- 11271528 TI - Enhanced reactivity of 2-rhodaoxetanes through a labile acetonitrile ligand. AB - New cationic, square-planar, ethene complexes [(Rbpa)RhI(C2H4)]+ [2a]--[2c]+ (Rbpa = N-alkyl-N,N-di(2-pyridylmethyl)amine; [2a]+: alkyl =R=Me; [2b]+: R = Bu; [2c]+: R = Bz) have been selectively oxygenated in acetonitrile by aqueous hydrogen peroxide to 2-rhoda(III)oxetanes with a labile acetonitrile ligand, [(Rbpa)RhIII(kappa2-C,O-CH2CH2O-)(MeCN)]+, [3a]+-[3c]+. The rate of elimination of acetaldehyde from [(Rbpa)RhIII(kappa2-C,O-CH2CH2O-)(MeCN)]+ increases in the order R = Me< R = Bu< R = Bz. Elimination of acetaldehyde from [(Bzbpa)RhIII(kappa2-C,O-CH2CH2O)(MeCN)]+ [3c]+, in the presence of ethene results in regeneration of ethene complex [(Bzbpa)RhI(C2H4)]+ [2c]+, and closes a catalytic cycle. In the presence of Z,Z-1,5-cyclooctadiene (cod) the corresponding cod complex [(Bzbpa)RhI(cod)]+ [6c]+ is formed. Further oxidation of [3c]+ by H2O2 results in the transient formylmethyl-hydroxy complex [(Bzbpa)RhIII(OH)[kappa1-C-CH2C(O)H]]+ [5c]+. PMID- 11271529 TI - The configurational stability of an enantioenriched alpha-thiobenzyllithium derivative and the stereochemical course of its electrophilic substitution reactions; synthesis of enantiomerically pure, tertiary benzylic thiols. AB - The lithium compound (S)-7, formed by deprotonation of the (S)-S-1-phenylethyl thiocarbamate (S)-10, is configurationally stable at -70 degrees C. Even at elevated temperatures it racemizes only very slowly. It represents the first essentially enantiopure alpha-thiocarbanion derivative and can be utilized in asymmetric synthesis. Most electrophiles (except proton acids) add to (S)-7 with complete stereoinversion. Cleavage of the substitution products leads to practically enantiopure, tertiary 1-phenylalkanethiols. PMID- 11271530 TI - Stirring effects on the spontaneous formation of chirality in the homoassociation of diprotonated meso-tetraphenylsulfonato porphyrins. AB - Homoassociates of the achiral title porphyrins in acid solutions show spontaneous symmetry breaking, which can be detected by circular dichroism (CD). The CD spectra are due to differential scattering and differential absorption contributions, the relative significance of which is related to the shape and size of the homoassociate. When an earlier model, designed for the association of these diprotonated porphyrins (J aggregates with the geometry of stepped sheets of intramolecular-stabilised zwitterions), was applied to an exciton-coupling point-dipole approximation, the folding of the one-dimensional homoassociates explained the CD signals detected. An effect of the vortex direction, caused by stirring or rotary evaporation, upon the exciton chirality sign was detected. In the case of H2TPPS3, the number of experiments performed gave a statistical significance to this effect. This vortex effect can be attributed to enhancement of the chirality fluctuations that originate in the diffusion-limited aggregation to high-molecular-weight homoassociates. In this sense, the phenomenon could be general for supramolecular systems that are obtained under kinetic control, and its detection would be possible when inherent chiral chromophores were being generated in the association process. PMID- 11271532 TI - Preparation and reactions of stannylated amino acids. AB - Hydrostannations of propargylic glycine esters with the new hydrostannation catalyst [Mo(CO)3(CNtBu)3] (MoBI3) gave rise to alpha-stannylated allylic esters in good yield and with high regioselectivity. The chelate Claisen rearrangements of these esters allow the syntheses of gamma,delta-unsaturated amino acids with a vinylstannane moiety in the side chain. The amino acids obtained can be further modified by cross-coupling with various types of electrophiles. PMID- 11271531 TI - Tetrathiafulvalene crowns: redox-switchable ligands. AB - A series of redox-responsive ligands that associate the electroactive tetrathiafulvalene core with polyether subunits of various lengths has been synthesized. X-ray structures are provided for each of the free ligands. The requisite structural criteria for reaching switchable ligands are satisfied for the largest macrocycles, that is, planarity of the 1,1',3,3'-tetrathiafulvalene (TTF) pi system and correctly oriented coordinating atoms. The ability of these ligands to recognize various metal cations as a function of the cavity size has been investigated by various techniques (LSIMS, 1H NMR, and UV/Vis spectroscopy, cyclic voltammetry). These systems exhibit an unprecedented high coordination ability among TTF crown ethers. Their switchable ligating properties have been confirmed by cyclic voltammetry, and metal-cation complexation has been illustrated by X-ray structures of three of the corresponding metal complexes (Pb2+, Sr2+, and Ba2+). Solid-state structures of these complexes display original packing modes with channel-like arrangements. PMID- 11271533 TI - A new type of calixarene: octahydroxypyridin. AB - A new type of calixarene has been synthesised. Like resorcinol and a number of resorcinol derivatives, 2,6-dihydroxypyridine has been proven to form cyclic tetramers in moderate yields with a number of aliphatic and two aromatic aldehydes in acidic media. The problem of the formation of configurational isomers can be reduced by increasing the reaction temperature and time. With electron-rich aromatic aldehydes, 2,6-dihydroxypyridine unexpectedly yields deep coloured acyclic quinoid systems. The octahydroxypyridine[4]arenes may have a high potential as receptors for molecular recognition and self organisation. PMID- 11271534 TI - Palladium-catalyzed amination of aryl bromides and aryl triflates using diphosphane ligands: a kinetic study. AB - [Pd(P-O-P)(Ar)]+ complexes with ligands that have wide bite angles are active catalysts for the coupling of aniline derivatives with aryl triflates. Kinetic studies show that for these systems a fast equilibrium that involves coordination of the amine precedes the deprotonation, which is the rate-limiting step of the reaction. This reaction is faster for compounds with a smaller P-Pd-P angle. When halide salts are present, the base sodium tert-butoxide is activated and adds to the palladium complex. This rate-limiting step is preceded by a fast equilibrium that involves decoordination of the halide. The initial reaction rate is faster for compounds with a larger P-Pd-P angle. This is due to the closer proximity of the oxygen to the Pd center, and this assists in the dissociation of the halide. PMID- 11271535 TI - Energetics of C-Cl, C-Br, and C-I bonds in haloacetic acids: enthalpies of formation of XCH2COOH (X=CI, Br, I) compounds and the carboxymethyl radical. AB - The standard molar enthalpies of formation of chloro-, bromo-, and iodoacetic acids in the crystalline state, at 298.15 K, were determined as deltafH(o)m(C2H3O2Cl, cr alpha)=-(509.74+/- 0.49) kJ x mol(-1), deltafH(o)m(C2H3O2Br, cr I)-(466.98 +/- 1.08) kJ x mol(-1), and deltafH(o)m (C2H3O2I, cr)=-(415.44 +/- 1.53) kJ x mol(-1), respectively, by rotating-bomb combustion calorimetry. Vapor pressure versus temperature measurements by the Knudsen effusion method led to deltasubH(o)m(C2H3O2Cl)=(82.19 +/- 0.92) kJ x mol( 1), deltasubH(o)m(C2H3O2Br)=(83.50 +/- 2.95) kJ x mol(-1), and deltasubH(o)m (C2H3O2I) = (86.47 +/- 1.02) kJ x mol(-1), at 298.15 K. From the obtained deltafH(o)m(cr) and deltasubH(o)m values it was possible to derive deltafH(o)m(C2H3O2Cl, g)=-(427.55 +/- 1.04) kJ x mol(-1), deltafH(o)m (C2H3O2Br, g)=-(383.48 +/- 3.14) kJ x mol(-1), and deltafH(o)m(C2H3O2I, g)=-(328.97 +/- 1.84) kJ x mol(-1). These data, taken with a published value of the enthalpy of formation of acetic acid, and the enthalpy of formation of the carboxymethyl radical, deltafH(o)m(CH2COOH, g)=-(238 +/- 2) kJ x mol(-1), obtained from density functional theory calculations, led to DHo(H-CH2COOH)=(412.8 +/- 3.2) kJ x mol( 1), DHo(Cl-CH2COOH)=(310.9 +/- 2.2) kJ x mol(-1), DHo(Br-CH2COOH)=(257.4 +/- 3.7) kJ x mol(-1), and DHo(I-CH2COOH)=(197.8 +/- 2.7) kJ x mol(-1). A discussion of the C-X bonding energetics in XCH2COOH, CH3X, C2H5X, C2H3X, and C6H5X (X=H, Cl, Br, I) compounds is presented. PMID- 11271536 TI - The facile preparation of weakly coordinating anions: structure and characterisation of silverpolyfluoroalkoxyaluminates AgAl(ORF)4, calculation of the alkoxide ion affinity. AB - Purified LiAlH4 reacts with fluorinated alcohols HORF to give LiAl(ORF)4 (RF= CH(CF3)2, 2a; -C(CH3)(CF3)2, 2b; -C(CF3)3, 2c) in 77 to 90% yield. The crude lithium aluminates LiAl(ORF)4 react metathetically with AgF to give the silver aluminates AgAl(ORF)4 (RF=-CH(CF3)2, 3a; -C(CH3)(CF3)2, 3b; -C(CF3)3, 3c) in almost quantitative yield. The solid-state structures of solvated 3a-c showed that the silver cation is only weakly coordinated (CN(Ag)=6-10; CN = coordination number) by the solvent and/or weak cation - anion contacts Ag-X (X=O, F, Cl, C). The strength of the Ag-X contacts of 3a-c was analysed by Brown's bond-valence method and then compared with other silver salts of weakly coordinating anions (WCAs), for example [CB11H6Cl6]- and [M(OTeF5)n]- (M=B, Sb, n=4, 6). Based on this quantitative picture we showed that the Al[OC(CF3)3]4 anion is one of the most weakly coordinating anions known. Moreover, the AgAl(ORF)4 species are certainly the easiest WCAs to access preparatively (20 g in two days), additionally at low cost. The Al-O bond length of Al(ORF)4- is shortest in the sterically congested Al[OC(CF3)3]4- anion-which is stable in H2O and aqueous HNO3 (35 weight%)--and indicates a strong and highly polar Al-O bond that is resistant towards heterolytic alkoxide ion abstraction. This observation was supported by a series of HF-DFT calculations of OR-, Al(OR)3 and Al(OR)4- at the MPW1PW91 and B3LYP levels (R= CH3, CF3, C(CF3)3). The alkoxide ion affinity (AIA) is highest for R=CF3 (AlA=384 +/- 9 kJ x mol(-1)) and R= C(CF3)3 (AIA=390 +/- 3 kJ x mol( 1)), but lowest for R=CH3 (AIA=363 +/- 7 kJ X mol(-1)). The gaseous AL(ORF)4 anions are stable against the action of the strong Lewis acid ALF3(g) by 88.5 +/- 2.5 (RF=CF3) and 63 +/- 12 kJ X mol(-1) (RF=C(CF3)3), while AL(OCH3)4- decomposes with -91 +/- 2 kJ X mol(-1). Therefore the presented fluorinated aluminates AL(ORF)4- appear to be ideal candidates when large and resistant WCAs are needed, for example, in cationic homogenous catalysis, for highly electrophilic cations or for weak cationic Lewis acid/base complexes. PMID- 11271537 TI - Gold complexes with dithiothiophene ligands: a metal based on a neutral molecule. AB - The gold complexes n-Bu4N[Au(alpha-tpdt)2] (5), n-Bu4N[Au(dtpdt)2] (4) and n Bu4N[Au(tpdt)2] (6) based on new dithiothiophene ligands (alpha-tpdt= 2,3 thiophenedithiolate, dtpdt=2,3-dihydro-5,6-thiophenedithiolate and tpdt = 3,4 thiophenedithiolate) have been prepared and characterised. These gold(III) complexes are diamagnetic, but they can be oxidised with iodine to the paramagnetic compounds [Au(alpha-tpdt)2] (8), [Au(dtpdt)2] (7) and n Bu4N[[Au(tpdt)2]n-2] (9), which were isolated as fine powders and which exhibit paramagnetic susceptibilities that are almost temperature independent with room temperature values of 2.5 x 10(-4), 2.0 x 10(-4) and 5 x 10(-4) emu x mol(-1), respectively. Interestingly, the neutral complex [Au(alpha-tpdt)2] (8) as a polycrystalline sample displays the properties of a metallic system with a room temperature electrical conductivity of 6 S x cm(-1) and a thermoelectric power of 5.5 microVK(-1); this is the first time that this metallic property has been observed in a molecular system based on a neutral species. PMID- 11271538 TI - Development of analogues of 1alpha,25-dihydroxyvitamin D3 with biased side chain orientation: methylated des-C,D-homo analogues. AB - The discovery that 1alpha,25-dihydroxyvitamin D3 is effective in the inhibition of cellular proliferation and in the induction of cellular differentiation has led to a search for analogues in which these activities and the classical calcemic activity of this hormone are separated. In this context, the synthesis and biological evaluation are reported of the three stereoisomeric CD-ring modified structural analogues in order to enforce a particular and different orientation of the 25-hydroxylated side chain. Comparison of the results of the biological evaluation and conformational analysis of the side chain suggests one defined and "active" geometry. PMID- 11271539 TI - Self-assembly of multinuclear coordination species with chiral bipyridine ligands: silver complexes of 5,6-CHIRAGEN(o,m,p-xylidene) ligands and equilibrium behaviour in solution. AB - The complexation reactions between Ag- and a series of enantiopure ligands belonging to the CHIRAGEN (from CHIRAlity GENerator) family (L1, L2, L3, based on (-)-5,6-pinene bipyridine) have been studied in solution. It has been shown that the length of the bridge plays a fundamental role in the self-assembly processes leading to different compounds: mononuclear complexes (with L3), mixtures of polynuclear complexes (with L2) and circular helicates (with L 1). Although the absolute configuration of the chiral centres in all three ligands is the same, the metal-centred chirality of L3 (delta) is inverted with respect to that in the other two complexes with L1 and L2 (delta). The metal configuration is thus opposite in the mononuclear complex with respect to the polynuclear species. Detailed thermodynamic studies were carried out for the Ag+ and L1 ligand system by 1H and 109Ag NMR spectroscopy (as a function of concentration, temperature and pressure). At low temperature and high pressure, the [Ag6L1(6)]6+ hexanuclear circular helicate forms a tetranuclear circular helicate [Ag4L1(4)]4+: 2[Ag6L1(6)]6+ <=> 3 [Ag4L1(4)]4+. The thermodynamics parameters, obtained by temperature and pressure variation, have the following values: K298 = (8.7 +/- 0.7) x 10(-5) mol x kg(-1), deltaHo = -15.65 +/- 0.8 kJ x mol(-1), deltaSo = 130.2 +/- 3 J x mol(-1) x K(-1) and deltaVo(256 K)= -160 +/- 12 cm3 x mol(-1). The reaction volume calculated according to Connolly's method indicates that the calculated structure of [Ag4L1(4)]4+ is plausible. Both the signs and large magnitudes of deltaSo and deltaVo are counterintuitive, yet can be understood by modelling methods. PMID- 11271540 TI - Provision of hormonal contraceptives without a mandatory pelvic examination: the first stop demonstration project. AB - CONTEXT: First Stop, an 18-month demonstration project that operated in 1996 1997, was designed to offer low-income adult women in California hormonal contraceptives without requiring a pelvic examination. METHODS: An evaluation was undertaken to assess the contraceptives adopted by First Stop clients, compare health risks of these women with risks among women using traditional family planning clinics and assess clients'satisfaction. Data on 2,065 First Stop clients and 1,507 women attending traditional clinics were collected through several self- and clinician-administered instruments, including questionnaires, a telephone survey and medical chart abstractions. RESULTS: After the initial First Stop visit, 38% of women adopted a more effective method than they had used at last sex, 4 7% remained with the same method, 12% switched to a less-effective method and 3% accepted no method. Of clients who were referred for additional medical care, 73% followed through on their referrals. Compared with clients at traditional clinics, First Stop clients were less likely to have a regular source of health care, but more likely to have made a health care visit in the past year. Most First Stop clients valued the project's services; 76% said it was important to be able to receive pills or injections without a pelvic examination. CONCLUSIONS: Programs that provide hormonal contraceptives without requiring a pelvic examination can expand low-income women's access to these methods and improve the chances that they will obtain other reproductive health services. PMID- 11271541 TI - Contraceptive failure in the first two years of use: differences across socioeconomic subgroups. AB - CONTEXT: While differences in levels of contraceptive use across socioeconomic subgroups of women have narrowed greatly over time, large disparities remain in rates of unintended pregnancy. One reason is variations in the effectiveness with which women and their partners use contraceptive methods. METHODS: Data on contraceptive use and accidental pregnancy from the 1988 and 1995 National Surveys of Family Growth were corrected for abortion underreporting and pooled for analysis. Use-failure rates were estimated for reversible methods during the first year, second year and first two years of use, for subgroups of women of various characteristics. RESULTS: The average failure rate for all reversible methods, adjusted for abortion underreporting, declines from 13% to 8% from the first year of method use to the second year. First-year failure rates are highest among women using spermicides, withdrawal and periodic abstinence (on average, 23 28% in the first year), and lowest for women relying on long-acting methods and oral contraceptives (4-8%). On average, they exceed 10% for all users except women aged 30-44, married women and women in the highest poverty-status category. The chance of accidental pregnancy does not differ significantly between method users younger than 18 and those aged 18-19. CONCLUSION: Both user and method characteristics determine whether contraceptive users will be able to avoid unintended pregnancy. Family planning providers should help clients to identify methods that they are most likely to use successfully, and counsel them on how to be consistent users and to avoid behaviors that contribute to method failure. PMID- 11271542 TI - A new HIV prevention framework. PMID- 11271543 TI - Choice of female-controlled barrier methods among young women and their male sexual partners. AB - CONTEXT: Little is known about the factors associated with the choice of female controlled, over-the-counter barrier contraceptive methods among women and their male sexual partners. METHODS: Predictors of method choice were assessed following an educational presentation on contraceptive use and risk reduction among 510 sexually active females aged 15-30 who were recruited in the San Francisco Bay Area. In addition, the primary partners of 160 of these women participated in the survey RESULTS: Twenty-two percent of women who enrolled in the study alone, 25% of those who enrolled with their main partner and 18% of these male partners chose female-controlled, over-the-counter barrier methods alone. The strongest predictor of this choice was current use of a hormonal contraceptive both for women who participated in the study on their own (odds ratio, 2.1) and for those who enrolled their partner in the study (odds ratio, 6.3). Female-controlled methods were also chosen significantly more often by teenagers than by older women; for example, among those who enrolled with a male partner, the odds ratio for selection of a female-controlled barrier method by women younger than 18 was 6.0. Among women who enrolled without a partner, those who had had multiple partners in the previous six months and those who were current users of male condoms were less likely to choose female-controlled methods (odds ratios, 0.7 and 0.5, respectively). CONCLUSIONS: Although the majority of participants did not choose female-controlled, over-the-counter barrier methods without also choosing male condoms, such female-controlled methods appear to offer an acceptable alternative for prevention of sexually transmitted infections. They may be a particularly attractive option for individuals using hormonal contraceptives and for teenage women. PMID- 11271544 TI - As if having HIV weren't enough. PMID- 11271545 TI - Union status, marital history and female contraceptive sterilization in the United States. AB - CONTEXT: Much of what is known about the choice of sterilization as a contraceptive method is based on data from married women or couples. Because of increasing rates of cohabitation, divorce and repartnering, however, the relationship context in which sterilization decisions are made has changed. METHODS: The 1995 National Survey of Family Growth includes the complete birth and union histories of 10,277 white, black and Hispanic women. The distribution of union status and marital history at the time of tubal sterilization was estimated for these three racial and ethnic groups among the 799 women who had had a tubal ligation in 1990-1995 before age 40. Cox proportional hazard regression models were used to estimate the effects of union status and marital history on the risk of tubal sterilization. The analysis controlled for the woman's age, parity, race and ethnicity education, region, experience of an unwanted birth and calendar period. RESULTS: Among women who obtained a tubal sterilization, most whites (79%) and Hispanics (66%) were married when they had the operation, compared with only 36% of black women. At the time of their sterilization, 46% of black women had never been married. Among all women, regardless of race and ethnicity and net of all controls, the probability of tubal sterilization is about 25% lower for single, never-married women than for cohabiting or married women. Cohabitation does not reduce the likelihood in comparison to marriage, however. Higher rates of tubal sterilization among Hispanic women are accounted for by their higher parity at each age; differences in parity or marriage by race only partially account for the relatively higher rates of tubal sterilization among black women. CONCLUSIONS: Because women currently spend greater proportions of their lives outside of marriage or in less stable cohabiting partnerships than they did in the past, they are increasingly likely to make the decision to seek sterilization on their own. As a result, the gender gap in contraceptive sterilization will likely increase. The possibility of partnership change is an important consideration in choosing sterilization as a contraceptive method. PMID- 11271547 TI - An assessment of the effectiveness of magnetic resonance imaging of the shoulder: literature review. AB - OBJECTIVE: To analyse and compare all papers published to date (August 2000) that quantify the effectiveness, defined as the impact of clinician's diagnosis or management plans, or patient outcome, of MRI of the shoulder. DESIGN: A computerised search of Index Medicus with a broad search strategy relating to shoulder MRI was performed. Manual assessment of all papers listed was undertaken with classification of each paper depending on whether it addressed questions of (1) technical performance, (2) diagnostic performance or (3) outcome. RESULTS: Four of 265 qualifying papers addressed aspects of effectiveness and these were reviewed. The impact on the clinician's diagnosis varied widely between papers: the primary diagnosis was altered in 23% to 68% of cases, and the management plans were subsequently changed in 15% to 61% of cases. Only one paper addressed the impact on patient health. CONCLUSIONS: The effectiveness of MRI of the shoulder depends on the clinical skills of the referring clinician and prevalence of disease in the study population. This will have implications when the effectiveness of an imaging technique between different institutions is compared, and this in turn will influence any comparisons of cost-effectiveness. PMID- 11271546 TI - Public or private providers? U.S. women's use of reproductive health services. AB - CONTEXT: U.S. women receive contraceptive and reproductive health services from a wide range of publicly funded and private providers. Information on trends in and on patterns of service use can help policymakers and program planners assess the adequacy of current services and plan for future improvements. METHODS: Women who reported in the 1995 National Survey of Family Growth that they had obtained any contraceptive or other reproductive health service in the past year were classified by their primary source of care, and the services they received, their characteristics and their primary source of care were analyzed. Logistic regression was used to test which factors predict women's use of publicly subsidized family planning clinics and of specific types of services. RESULTS: The percentage of women of reproductive age who obtained family planning services increased slightly between 1988 and 1995, primarily among women aged 30 and older. Nearly one in four women who received any contraceptive care visited a publicly funded family planning clinic, as did one in three who received contraceptive counseling or sexually transmitted disease (STD) testing and treatment. Women whose primary source of reproductive care was a publicly funded family planning clinic received a wider range of services than women who visited private providers; moreover, the former were significantly more likely to report obtaining contraceptive care or STD-related care, even after the effects of their background characteristics were controlled. Young, unmarried, minority, less educated and poor women were more likely than others to depend on publicly subsidized family planning clinics. Source of health insurance was one of the most important predictors of the use of public family planning clinics: Medicaid recipients and uninsured women were 3-4 times as likely as women with private insurance to obtain clinic care. CONCLUSIONS: Publicly funded family planning clinics are an important source of contraceptive and other reproductive health care, providing millions of U.S. women with a wide range of services. Since women's need for reproductive care and for publicly subsidized care is not likely to diminish, clinics may be financially challenged in their efforts to continue delivering this broad package of services to growing numbers of uninsured or disenfranchised women. PMID- 11271548 TI - Alveolar soft part sarcoma: MR and angiographic findings. AB - OBJECTIVE: To present the MR and angiographic findings of alveolar soft part sarcoma (ASPS). DESIGN AND PATIENTS: MR examinations (12 tumors of 10 patients) of ASPS performed at multiple hospitals were retrospectively reviewed. The tumors were found in the thigh (n=4), lower leg (n=4), femur (n=2, local metastasis), scalp (n=1) and arm (n=1). The MR signal characteristics including signal intensity, homogeneity and signal void of lesions and bony invasion including direct invasion or local metastasis were evaluated. Angiographic findings (n=4) and post-embolotherapy follow-up MR imaging (n=2) findings were also assessed. RESULTS: Local bony metastasis was found in two cases. Seven tumors showed heterogeneous high signal intensity on T - and T2-weighted images with good enhancement. One tumor had a very high signal on T1-weighted images. Eight tumors (67%) showed numerous signal voids in or near the tumors. All four angiographic studies showed numerous enlarged vessels, arteriovenous shunts and delayed washout. Two cases mimicked arteriovenous malformations on angiographic studies but MR images demonstrated solid soft tissue components as well as tortuous vessels. CONCLUSIONS: High signal on T1 -weighted image and numerous signal voids are highly suggestive of ASPS, although they are not universal as has been suggested and arteriovenous malformation should be included in the differential diagnosis. Local bony metastases in ASPS were seen in two cases and should be carefully investigated. PMID- 11271549 TI - Atypical calcific tendinitis with cortical erosions. AB - OBJECTIVE: To present and discuss six cases of calcific tendinitis in atypical locations (one at the insertion of the pectoralis major and five at the insertion of the gluteus maximus). PATIENTS AND RESULTS: All cases were associated with cortical erosions, and five had soft tissue calcifications. The initial presentation was confusing and the patients were suspected of having infection or neoplastic disease. CONCLUSION: Calcific tendinitis is a self-limiting condition. It is important to recognize the imaging features of this condition to avoid unnecessary investigation and surgery. PMID- 11271550 TI - Increased T2 signal intensity in the distal clavicle: incidence and clinical implications. AB - OBJECTIVE: The objectives of the current study were (1) to quantify the incidence of increased T2 signal in the distal clavicle and (2) to assess the clinical significance of this finding in patients with chronic acromioclavicular (AC) joint pain. DESIGN AND PATIENTS: Eight patients (five male and three female, 15 41 years of age) with disabling shoulder pain localized to the AC joint and marked increased T2 signal in the distal clavicle are presented. These eight patients underwent MR examination over a 25 month period (August 1996 to September 1998). The dictated reports of all shoulder MR examinations conducted over this same time period were reviewed retrospectively for the presence of signal abnormality in the distal clavicle. Clinical data and, in five patients, findings at shoulder arthroscopy or open surgery, were correlated with the results of MR imaging. One patient underwent arthroscopy on both shoulders. RESULTS: The selected eight patients each presented clinically with disabling shoulder pain localized to the AC joint. One patient is presented twice, as both shoulders were symptomatic (n=9). Plain film examination (9/9) failed to indicate a structural cause of shoulder pain in any of the patients. MR examination demonstrated abnormally increased T2 signal in the distal clavicle in all nine cases and no other cause for AC joint pain. Three patients responded to a course of conservative therapy. Six experienced refractory pain despite conservative therapy. Resection of the distal clavicle was performed in five of the six cases. All patients who underwent resection of the distal clavicle experienced complete resolution of AC joint pain. A retrospective review of the dictated reports for all shoulder MR imaging examinations performed at out institution over a 25 month period (August 1996 to September 1998; n=761) demonstrated a 12.5% incidence of abnormally increased T2 signal in the distal clavicle. CONCLUSIONS: Increased T2 signal in the distal clavicle is a relatively common finding (12.5%) on MR imaging examinations of the shoulder and in most cases is of no clinical significance. However, in patients with chronic AC joint pain and no other abnormality on plain film or MR imaging, increased T2 signal may represent an early manifestation of, or a process similar to, osteolysis of the distal clavicle. Patients with this presentation who continue to suffer from disabling pain following conservative therapy may benefit from surgical resection of the distal clavicle. PMID- 11271551 TI - Pedicle marrow signal intensity changes in the lumbar spine: a manifestation of facet degenerative joint disease. AB - OBJECTIVE: Signal intensity changes in lumbar pedicles, similar to those described in vertebral body endplates adjacent to degenerated discs, have been described as an ancillary sign of spondylolysis on MRI. The purpose of this study was to determine whether pedicle marrow signal intensity changes also occur in association with facet degenerative joint disease. DESIGN: Eighty-nine lumbar spine MRI examinations without spondylolysis were reviewed for marrow signal intensity changes in pedicles and vertebral bodies as well as for facet degenerative joint disease. RESULTS: Five percent (46/890) of lumbar pedicles in 23 patients had marrow signal intensity changes. Ninety-one percent (42/46) of the abnormal pedicles had adjacent degenerative joint disease of the facets, while only 21% (189/890) of normal pedicles had adjacent facet degenerative joint disease (p<0.001). Eighty-nine percent (41/46) of the pedicles with marrow signal intensity changes had adjacent degenerative disc disease. CONCLUSIONS: Pedicle marrow signal intensity changes are not a specific sign of spondylolysis; they are commonly seen with adjacent facet degenerative joint disease in the absence of spondylolysis. Pedicle marrow signal intensity changes are probably a response to abnormal stresses related to abnormal motion or loading caused by the degenerative changes in the spinal segment. PMID- 11271553 TI - Synovial sarcoma arising in association with a popliteal cyst. AB - Synovial sarcoma is a relatively common soft tissue sarcoma particularly in the adolescent and young adult. We report an unusual case of a synovial sarcoma arising within a popliteal cyst in a 13-year-old female presenting with bilateral popliteal cysts. MR imaging demonstrated the cyst with evidence of subacute haemorrhage and a discrete nodule of tumour. PMID- 11271552 TI - Intraosseous glomus tumor of the fibula. AB - Glomus tumor is a rare, benign vascular tumor and intraosseous glomus tumor, which arises primarily within bone, is even rarer. Fewer than 20 cases have been reported in the literature. We present the case of a 34-year-old woman with glomus tumor primarily in the midshaft of the fibula that radiologically mimicked chondromyxoid fibroma, aneurysmal bone cyst or adamantinoma, together with a review of other reported cases. PMID- 11271554 TI - Popliteal amyloidoma presenting with leg ischemia in a chronic dialysis patient. AB - The authors report a case of bilateral popliteal amyloidoma causing stenosis of the popliteal artery and vein. This patient had been treated with hemodialysis for 26 years. The diagnosis was made with MR angiography. A popliteal tumor of the right knee was resected surgically and the histologic examination showed deposition of amyloid. After resecting the popliteal tumor, the severe leg pain and intermittent claudication improved. This report suggests that popliteal amyloid tumors should be considered in a patient undergoing long-term hemodialysis who complains of leg pain and intermittent claudication. PMID- 11271555 TI - Sarcomatoid chordoma: chordoma with a massive malignant spindle-cell component. AB - We report a case of chordoma containing a spindle cell sarcomatoid component with a gradual transition from conventional chordoma. Immunohistochemically, many tumor cells in both conventional chordoma and sarcomatoid components were positive for cytokeratins (AE1/AE3, CAM5.2) and epithelial membrane antigen as well as vimentin. This report provides a rare example of sarcomatoid chordoma. Familiarity with this type of bone tumor should help to avoid confusion with dedifferentiated chordoma and other spindle cell sarcomas or carcinomas. PMID- 11271556 TI - Intra-articular membranous interposition detected by MRI in developmental dysplasia of the hip. AB - Intra-articular membranous interposition was detected by MRI in the hip joint with residual subluxation of a girl aged 5 years 10 months. This structure, which had low signal intensity on both T1- and T2-weighted images, separated the femoral head from the acetabulum. Histological examination revealed chondrometaplasia, which suggested that this interposition might be transformed to a surface cartilaginous tissue of the secondary acetabulum often observed in residual subluxation of the hip. PMID- 11271557 TI - [SBU report on asthma and chronic obstructive lung disease. A strong support prior to difficult decisions in clinical situations]. PMID- 11271558 TI - [SBU report on asthma and COPD. Useful and easy-to-read also for stressed general practitioners]. PMID- 11271559 TI - [SBU report on asthma and chronic obstructive lung disease. A systematic review of knowledge which makes the choice of therapy easier]. PMID- 11271560 TI - [Oxygen therapy undisputed in severe, but doubtful in moderate, hypoxia. Comment to meta-analysis of home oxygen therapy in chronic obstructive lung disease]. AB - The review included randomized controlled trials on patients with chronic obstructive pulmonary disease (COPD) receiving domiciliary long-term oxygen therapy (LTOT). The authors identified six articles concerning four randomized controlled trials but could not perform any meta-analysis due to the heterogenous patient populations and treatments. From these trials they conclude that LTOT improves survival in patients with severe hypoxemia (arterial PaO2 less than 8 kPa) but has no effect in patients desaturating only at night or in patients with moderate hypoxemia. They also sensibly remark that it is possible that statistically significant improvements in some physiological variables have little measurable impact on subjects perceived quality of life or survival. The conclusion that survival benefit is demonstrated also in patients with hypoxemia in the range 7,4-8 kPa is debatable, with the strongest evidence pointing against benefit. These patients are better classified as moderately hypoxemic. In Sweden, they comprise 20% of new patients starting on LTOT. For them, the effect of LTOT should be evaluated individually in terms other than survival or quality of life. A more recently published trial supports the conclusion that domiciliary nocturnal oxygen therapy has no impact on survival in nocturnal desaturation without severe daytime hypoxemia. There is also new evidence that the type of oxygen equipment might have a decisive impact on the quality of life in mobile patients receiving LTOT--improved quality of life with liquid oxygen and poorer quality of life with concentrator and conventional (heavy) gas cylinder. The importance of optimum equipment selection for each patient has been overlooked but merits further investigation. PMID- 11271561 TI - [Policy document. Catheter-based diagnosis and treatment of coronary diseases]. AB - Diagnostic coronary angiography and percutaneous coronary interventions (PCI) are rapidly developing fields. In-house thoracic surgery backup is no longer a prerequisite for PCI. The demand for physicians trained in interventional cardiology has created a need to formalise such education. The Swedish societies of cardiology and thoracic radiology have agreed on a policy document establishing the details of this education. It is the responsibility of the tutor to decide when the pupil has achieved adequate skills. PMID- 11271562 TI - [The diabetic hand--complications of diabetes]. AB - Diabetes mellitus is common in Western countries. Secondary complications in various organs occur after type 1 as well as type 2 diabetes. Complications, such as nephropathy, retinopathy and neuropathy, are well known. Specific efforts to treat complications in the lower leg--"the diabetic foot"--have been emphasized. Various complications to diabetes occur also in the upper extremity and hand- "the diabetic hand"--and include not only more specific diabetic-related conditions like limited joint mobility but conditions related to the non-diabetic hand, such as trigger finger, Dupuytren's contracture and peripheral nerve compression lesions are also seen. Following surgical treatment of the latter conditions extra care of patients with diabetes mellitus--for example physiotherapy and help by an occupational therapist--may have to be provided. This review describes complications in the hand in diabetes mellitus. PMID- 11271563 TI - [National Diabetes Registry. A teaching pedagogical instrument for quality assurance]. AB - The National Diabetes Registry of Sweden presents results from its third year of registration. Quality of care data from more than 41,000 patients with diabetes treated at medical and children's departments as well as primary health care centers are compared with national goals concerning metabolic and blood pressure control, and information is provided concerning the prevalence of lipid treatment, nephropathy, smoking etc. The national HbA1c goal of 6.5% is reached by 30% of patients treated at medical departments and by 50% in primary health care. Concerning patients with onset before 30 years of age, this goal is reached by 25%, which indicates that the national goal is difficult to reach and that individual goals must be set for individual patients. PMID- 11271564 TI - [Anti-reflux surgery is possible also in children with multiple disabilities. Good results after Nissen fundoplication in 10-years experience]. AB - 82 children with gastroesophageal reflux, 43 of whom were seriously neurologically impaired, were operated according to Nissen. A retrospective study compares the results, complications and recurrences between children with and children without neurological disability. There were no major differences between the groups; four of the neurologically impaired children and one without impairment had died from unrelated causes. In 62 children, 31 of whom were neurologically impaired, the operation was completely successful, 13 children (six of whom were neurologically impaired) had minor problems, two (both neurologically impaired) had major problems. The conclusion is that serious neurological impairment can not be considered a contraindication to antireflux surgery. PMID- 11271565 TI - [Intracranial endoscopy]. PMID- 11271566 TI - [When cholera held the inhabitants of Uppsala in an iron grip]. PMID- 11271567 TI - [Why is not blood pressure determination routinely performed in sitting position in Sweden? Overtreatment of false hypertension could be avoided]. PMID- 11271568 TI - ["Health allowance" should not function outside big cities]. PMID- 11271569 TI - [Competition is beneficial also in sparsely populated areas]. PMID- 11271570 TI - [Report from a bucket]. PMID- 11271571 TI - [Child abuse--from the perspective of the child]. PMID- 11271572 TI - [Postoperative radiotherapy increases mortality in women with breast cancer diagnosed by mass screening]. PMID- 11271573 TI - [It's unfortunate to put dialogue against evidence]. PMID- 11271574 TI - [Sweden behind when it comes to influenza vaccination. Better vaccination programs in many western countries]. PMID- 11271576 TI - [Feedback on prescribing profiles at a primary health center. Important element in quality assurance of drug prescription]. AB - DU90%--the number of drugs that account for 90% of DDDs--and adherence to guidelines in this segment were proposed by the Swedish Medical Quality Council (MKR) as indicators for assessing the quality of drug prescribing. We tested these indicators at a primary health care (PHC) center in Stockholm. Bar-coded prescriptions purchased at pharmacies were compared with the guidelines issued by the regional drug committee. The data were presented and discussed at the PHC center. Although the DU90% method neither examines the appropriateness of use nor provides outcome data, it was shown to be an inexpensive, flexible and simple method for assessing the general quality of drug prescribing. PMID- 11271575 TI - [More elderly persons should be informed about the importance of vaccination against influenza. Extensive needs of information also among the health personnel]. AB - In Sweden yearly vaccination against influenza for persons over 65 years of age has been recommended since 1997. In the Goteborg area approximately 33.6 percent of this age group were vaccinated during the autumn 1999. The most common reasons given for not being vaccinated were lack of confidence in the vaccine and a belief that influenza is not a serious disease. The study shows that both doctors and the target group need more information about the benefits of influenza vaccination. PMID- 11271577 TI - [Catheter remnants can cause intestinal obstruction and perforation. PEG catheters should be removed with the guidance of a gastroscope]. PMID- 11271578 TI - [Orthopedic inpatients--an afflicted and complex patient group]. AB - We studied all patients (n = 147) discharged from the orthopedic wards at Karnsjukhuset, Skovde, during a four week period. Two thirds had been admitted via emergency rooms, were older, used more medicines and had more diagnoses than the elective patients, and they required a great deal of medical resources. 38% of the emergency patients did not require specific orthopedic care. 47% of these were more than 80 years of age, had several diagnoses, and had been in-patients in at least two departments during the past year. There is a need for enhanced cooperation and information between departments in order to better provide care for elderly patients with more than one diagnosis. PMID- 11271579 TI - [The case method--a new student-activating method in medical education]. AB - The case method is a student-activating method, used together with problem based learning (PBL) and traditional teaching methods in the curriculum for undergraduate medical education at the Faculty of Medicine in Lund/Malmo, Sweden. The case method provides training in the solving of clinical problems and is thus especially useful at the clinical level of medical education, and in an integrated and problem based curriculum. The case method consumes less teaching resources than PBL, and might thus be useful in a situation with increasing numbers of medical students. PMID- 11271581 TI - [The evil of Nazi ideology. Destructive effects of the insanity on health services and other functions in the society]. PMID- 11271580 TI - [Large social inequalities behind women's risk of coronary disease. Unskilled work and family strains are crucial factors]. AB - There is a clear and consistent association between lower social economic position and increased risk for coronary heart disease (CHD). This association is even stronger in women than men. In the Stockholm Female Coronary Risk study, compared with executives/professionals, women with un/semiskilled occupations had a four-fold increased risk for developing CHD. Using similar comparisons, a three fold increased risk for a poor CHD prognosis was observed after a 5-year follow up. Family stress was an important factor contribution to the socioeconomic differences in women's cardiovascular health. Both family- and work-related factors should be considered in strategies geared to reducing social inequalities in women's cardiovascular health. PMID- 11271582 TI - [Sufferings of Jewish physicians during the Nazi terror]. PMID- 11271583 TI - [Patients' rights and physicians' professional independence. some thoughts on prioritization and choice of treatment]. PMID- 11271584 TI - [Nijinsky, divine dancer and choreographer. He outlined the geometry of dance and life]. PMID- 11271586 TI - [A reply: Criticism is based on an objective basis in spite of all. Attention deficit disorder with hyperactivity]. PMID- 11271585 TI - [Irrelevant criticism under scientific cover. Attention deficit disorder with hyperactivity]. PMID- 11271587 TI - [An attitude which is being exploited. Attention deficit disorder with hyperactivity]. PMID- 11271588 TI - [A reply: General survey is necessary. Attention deficit disorder with hyperactivity]. PMID- 11271589 TI - [Demosthenes did not suffer of stammering but of rhinolalia aperta]. PMID- 11271590 TI - [The number of reported cases to the HSAN is not unreasonably great]. PMID- 11271591 TI - [Misleading marketing concerning thrombosis survey]. PMID- 11271598 TI - Dietary vitamin E and rainbow trout (Oncorhynchus mykiss) phagocyte functions: effect on gut and on head kidney leucocytes. AB - The effects of vitamin E (deficiency or supplementation) on the non-specific immune system in rainbow trout, Oncorhynchus mykiss, were evaluated. Rainbow trout were fed daily a semi-purified diet supplemented with vitamin E at 0, 28 and 295 mg x kg(-1) of diet. After 80 days of experimental feeding, the phagocytic function (respiratory burst evaluated by the CL response, phagocytosis) from gut leucocytes and head kidney enriched macrophages was measured; head kidney cell pinocytosis and serum lysozyme activity were also analysed. The results showed that some phagocyte functions were influenced by dietary vitamin E. When fish were fed the high dietary dose of vitamin E an enhancement of phagocytosis was found, but only significantly for the leucocytes isolated from the gut of rainbow trout; moreover, an impaired response was also observed in the fish fed no vitamin E for 80 days. However, no significant differences were noticed on the oxidative burst (CL) response of both gut and head kidney cells according to the dietary dose of vitamin E. Pinocytosis evaluated on head kidney cells was not influenced by dietary vitamin E. Fish fed vitamin E at 295 mg x kg(-1) had a lower serum lysozyme activity than those fed with vitamin E at 28 mg x kg(-1) and the fish fed no vitamin E for 80 days had an impaired activity. Thus, the present results demonstrate that altered dietary levels of vitamin E modulates the phagocytic functions of gut leucocytes in rainbow trout; moreover, the vitamin E diet effect seems to be greater on the local intestinal response as compared to systemic (head kidney). Taken together, this study confirms the crucial role of gut phagocytes in mucosal non-lymphoid defences in fish. PMID- 11271599 TI - Uptake and processing of a Vibrio anguillarum bacterin in Artemia franciscana measured by ELISA and immunohistochemistry. AB - Nauplii of Artemia franciscana were incubated in two different concentrations (undiluted and 1:9 in autoclaved sea water) of a divalent bacterin composed of two different serovars of Vibrio anguillarum. In order to investigate uptake and further processing of a bacterin in the live feed organism A. franciscana, immunohistochemistry was applied, visualising the presence of whole bacterial cells and antigens from the bacterin in individual nauplii. By using ELISA, it was shown that approximately 1.5-2-5 x 10(5) cells were incorporated into each Artemia under the conditions used. Maximum incorporation of cells was measured after 30 min, whereas after 60 min there was a decline to levels of 0.9-1.6 x 10(5) cells per Artemia. Immediately after incubation in the bacterin solution, the nauplii were transferred to a culture of the alga Isochrysis galbana, in order to simulate transfer of the nauplii to rearing tanks for fish larvae. From the ELISA, it could be concluded that the incorporated bacterial cells were excreted from the Artemia nauplii rapidly, however a large variation among different nauplii could be visualised by immunohistochemistry. PMID- 11271600 TI - Enhanced lysozyme production in Atlantic salmon (Salmo salar L.) macrophages treated with yeast beta-glucan and bacterial lipopolysaccharide. AB - Atlantic salmon head kidney macrophages grown in the presence of particulate yeast beta-glucan and bacterial lipopolysaccharide (LPS) showed increased production of lysozyme in the culture supernatants compared to non-treated controls. The increased lysozyme production started at day 3 and was five- to six fold higher compared to controls at day 6 in culture. Beta-glucan showed an approximate linear dose-response curve between 1 and 250 microg x ml(-1) whereas LPS showed a dose-response curve with a well-defined optimum concentration (10 microg x ml(-1)). The increase in lysozyme activity was accompanied by an accumulation of lysozyme gene transcript in the stimulated cells. Recombinant human tumor necrosis factor alpha, known for its ability to stimulate lysozyme in human macrophages and to elevate respiratory burst activity of rainbow trout macrophages, failed to stimulate lysozyme production of Atlantic salmon macrophages. Macrophages isolated from fish suffering from a non-lethal Ichthyobodo necator infection displayed a highly increased ability to produce lysozyme in response to both beta-glucan and LPS. As in higher vertebrates, lysozyme production may reflect the differentiation stage of the Atlantic salmon macrophages as well as a direct activation of lysozyme gene transcription by biological response modifiers. The rather late increase in lysozyme production induced by beta-glucan and LPS may thus be explained by stimulation of differentiation of the macrophages in culture eventually combined with direct activation of transcription of the lysozyme gene. PMID- 11271601 TI - Antibodies against Vibrio salmonicida lipopolysaccharide (LPS) and whole bacteria in sera from Atlantic salmon (Salmo salar L.) vaccinated during the smolting and early post-smolt period. AB - The antibody response to Vibrio salmonicida LPS was studied by ELISA and immunoblot after vaccination of salmon with an aqueous vaccine containing the bacterium. The vaccination of the groups took place from February to July. Antibodies to LPS were present in all sera. Sera from unvaccinated fish and samples collected a short time after vaccination contained low antibody levels. The antibody levels in vaccinated groups increased with time after vaccination except for fish vaccinated during the smolting period. For these fish groups decreasing levels were found in the autumn. Vaccination provided high levels of antibodies to LPS at least 6 months later, even with low water temperatures at primary and secondary vaccination. Fish vaccinated during smolting at higher water temperatures reached high antibody levels a short time after vaccination, but for these groups a decrease took place resulting in the lowest antibody levels of the vaccinated groups in September. Immunoblots showed that antibody reacted with low molecular weight components corresponding to the O-side chain. Immunoblots using whole bacteria as antigen showed a few immunoreactive bands and individual reaction patterns with respect to location of the bands and immunodominance. PMID- 11271602 TI - Effects of intrinsic and extrinsic factors on the haemocyte profile of the prawn, Macrobrachium rosenbergii. AB - The giant freshwater prawn Macrobrachium rosenbergii was investigated for its total haemocyte count (THC) based on season, sex, size and feeding rate. The THC, when the prawns were subjected to injections of foreign materials was also investigated. The prawns displayed the highest and lowest THC in autumn and winter respectively, with no significant difference between male and female, or among animals with a body weight range of 7-115 g. The prawns displayed the lowest THC at D3 stage, and the highest in C stage. The prawns displayed the lowest THC when they were fed at 0.1% feeding rate among feeding rates of 0.1%, 0.5%, 1.0% and 1.5% body weight x day(-1). Prawns injected with carbon powder and Enterococcus showed increased THC during the first 6 h. Prawns injected with saline and carbon powder had the lowest THC after 30 h, and recovered to the normal value after 54 h. Prawns injected with Enterococcus showed the lowest THC after 42 h, and showed delayed recovery. PMID- 11271604 TI - Immune parameters of immunised cod (Gadus morhua L.). AB - The immune response of cod (Gadus morhua L.) is unusual in that specific antibody response is limited or absent. In the present study cod was immunised with haptenated and non-haptenated protein antigen at two different temperatures and the antibody response monitored over a period of 18 months. Other humoral parameters of immunological importance were also analysed, namely total immunoglobulin concentration, anti-protease and spontaneous haemolytic activity. No specific antibody response was detected but increased activity of non-specific anti-TNP antibodies was observed 10-12 weeks after immunisation, irrespective of the antigen used. This antibody activity was attributed to the adjuvant used (FCA) and did not cross react with other antigens tested. Other parameters were probably not influenced by the immunisation but seasonal fluctuations were indicated. The immunoglobulin level appeared to peak in August-September and the anti-protease activity and the haemolytic activity in October-January. PMID- 11271603 TI - The gill is a major organ for antibody secreting cell production following direct immersion of sea bass (Dicentrarchus labrax, L.) in a Photobacterium damselae ssp. piscicida bacterin: an ontogenetic study. AB - Extremely high numbers of antibody secreting cells (ASC) were observed in the gills of sea bass fry immunised at three different age/sizes (initial weight of 0.1, 2 and 5 g) by direct immersion in a Photobacterium damselae spp. piscicida bacterin. The relatively low ASC production in the head kidney and spleen suggests that the systemic compartment was only slightly stimulated upon immersion vaccination. There was no response of corresponding magnitude in the gut as the one observed in the gills. A clear age effect was observed in the ASC response of the different groups, especially visible in the gills. Significantly higher numbers of specific ASC were observed in the gills of the two oldest groups (initial weight of 2 and 5 g) compared with the youngest fish (initial weight of 0.1 g), but the oldest groups were not significantly different from each other. Additionally, a more rapid response was observed with the ageing of the fish, with peak responses in all the organs at day 18, 16 and 8 post immunisation in the smallest to largest fish, respectively. There was no evidence that direct immersion exposure to P. damselae ssp. piscicida at the earliest stages used in the present study (0.1 g) was tolerogenic. In the context of present knowledge, this study strongly supports the importance of the route of immunisation to locally stimulate ASC and the importance that the gills might have in specific responses. PMID- 11271605 TI - Phagocytosis of Loma salmonae (Microsporidia) spores in Atlantic salmon (Salmo salar), a resistant host, and chinook salmon (Oncorhynchus tshawytscha), a susceptible host. AB - The in vitro phagocytosis of Loma salmonae spores by macrophages of Atlantic salmon and two strains of chinook salmon were investigated. Opsonisation of L. salmonae with plasma factors increased uptake by head kidney macrophages. Macrophages of Atlantic salmon, which are resistant to the parasite, had a significantly higher phagocytic index (PI) than those of chinook salmon, a susceptible species. This may indicate a possible mechanism contributing to resistance in Atlantic salmon or that L. salmonae is able to evade or suppress initial binding by macrophages of chinook. Non-specific binding or lectinophagocytosis was also suggested by significantly higher PI of spores from EDTA treated plasma when compared with no plasma or heat treated plasma. In comparison, uptake of Baker's yeast Saccharomyces cerevisiae by phagocytes was not significantly different between fish species and strains for all treatments. PMID- 11271606 TI - Costs and effectiveness of community postnatal support workers. Researchers must now focus on effectiveness with specific groups of women. PMID- 11271607 TI - Mental health services for people with learning disabilities. Medical needs are important too. PMID- 11271608 TI - Differential diagnoses for asthma should include mediastinal masses. PMID- 11271609 TI - Improvement in prescribing can be measured only over time. PMID- 11271610 TI - Emphasis has shifted from medical ethics to bioethics. PMID- 11271611 TI - Stapled haemorrhoidectomy offers substantial benefits. PMID- 11271612 TI - [Recurrent hemoptysis. 65-year-old post-kidney transplant patient with bilateral thoracic wall tumors]. PMID- 11271613 TI - [66-year-old patient with anemia, thrombocytopenia and changes in plasma coagulation after surgery of femoral neck fracture]. PMID- 11271614 TI - [Cytokines: classification, receptors, mechanisms of action]. PMID- 11271615 TI - [Sudden deafness and tinnitus]. PMID- 11271617 TI - [Pediatric emergency in the house call. Comment on the contribution by K. Chelius]. PMID- 11271618 TI - [Suppression of synthesis of tumor necrosis factor]. PMID- 11271619 TI - [Cytokine analysis. What is feasible--what is useful?]. PMID- 11271620 TI - [Modulation of cytokines in chronic inflammatory bowel diseases]. PMID- 11271616 TI - [Good sense and nonsense of antitussive agents]. PMID- 11271621 TI - [Cytokines and anti-cytokine therapy in rheumatoid arthritis]. PMID- 11271622 TI - [Cytokines and obstructive respiratory tract diseases]. PMID- 11271623 TI - [Cytokines in heart diseases]. PMID- 11271624 TI - [Cytokines and chronic renal replacement measures]. PMID- 11271625 TI - [Lipid therapy. Prevention of arteriosclerotic cardiovascular diseases]. PMID- 11271626 TI - Type II nitric oxide synthase (NOS2) expression correlates with lymph node status in oral squamous cell carcinoma. AB - In tumour biology, nitric oxide (NO) has a complex array of concentration dependent actions, including both inhibitory and promoting effects. It is thought that the levels of NO found in many human cancers lead to enhanced angiogenesis and tumour dissemination. In the current study, we assessed the immunohistochemical expression of the enzyme type II nitric oxide synthase (NOS2) in 41 cases of oral squamous cell carcinoma and correlated the findings with lymph node status. A significant relationship was found between NOS2 expression and lymph node metastasis (P<0.0002). Furthermore, lymph node metastasis correlated with the degree and intensity of staining seen (P<0.001). No correlation was found between the size of the primary tumour, degree of tumour differentiation or smoking status and NOS2 staining. Western blotting confirmed NOS2 protein expression in select cases. As with many other human tumours, NOS2 is not a ubiquitous finding in oral cancer. Its expression may be of value in assessing lymph node status prior to surgery, and it represents a target for possible therapeutic manipulation. PMID- 11271627 TI - Sequential immunohistochemical p53 expression in biopsies of oral lichen planus undergoing malignant evolution. AB - Transformation in squamous cell carcinoma (SCC) may occur in a small percentage of patients affected by oral lichen planus (OLP), but the pathogenesis remains to be elucidated. Overexpression of p53 protein was investigated immunohistochemically in 28 cases of OLP, followed up by sequential biopsies for up to 96 months. In 15 cases (Group 1), no dysplastic changes or neoplastic transformation occurred during the follow-up period; in 7 cases, OLP and SCC were synchronously observed (Group 2), whereas in another 6 cases (Group 3) SCC developed several months or years after diagnosis of OLP. The percentage of p53 positive epithelial cells at first diagnosis was significantly higher in the cases of Groups 2 and 3 than in those of Group 1. In contrast, evaluation of growth fraction by MIB-1 monoclonal antibody did not show any statistical differences among the three groups. Although no conclusions can be drawn about the molecular pathway leading to neoplastic transformation of OLP, or about the role of p53, the results indicate that immunohistochemical evaluation of p53 expression may be a practical tool to select cases of OLP with a high risk of neoplastic transformation. PMID- 11271628 TI - Comparison of apoptosis and apoptosis-related gene products between extranodal oral B-cell lymphoma and maxillofacial nodal B-cell lymphoma. AB - Twenty-seven cases of primary extranodal oral B-cell lymphoma and 22 cases of primary maxillofacial nodal B-cell lymphoma were investigated for the presence of apoptotic cells and the expression of apoptosis-related gene products by terminal deoxynucleotidyl-mediated dUTP nick end labeling (TUNEL) and immunohistochemistry. The majority of extranodal oral diffuse large B-cell lymphomas (17/25, 68%) and maxillofacial nodal diffuse large B-cell lymphomas (14/16, 88%) contained no or less than 10% apoptotic cells. Whereas the majority of extranodal oral diffuse large B-cell lymphomas (18/25, 72%) and maxillofacial nodal diffuse large B-cell lymphomas (13/16, 81%) contained more than 10% of Ki 67-positive cells. Bcl-2-, Bax-, p53- and Ki-67-positive rates were higher in maxillofacial nodal diffuse large B-cell lymphomas than in extranodal oral diffuse large B-cell lymphomas, but only Bax (chi2 test, 0.01 .05) were fat grams, r = 0.031; fat as percentage of kilocalories, r = 0.023; fiber grams per 1,000 kcal; r = 0.230; and fruit/vegetable servings, r = 0.186. Correlations with total functional network scores were: fat grams, r = 0.022 (P > .05); fat as percentage of kilocalories, r = -0.108 (P > .05); fiber grams per 1,000 kcal, r = 0.276, P < .05; and daily fruit/vegetable servings, r = 0.326, P < .05. APPLICATIONS/CONCLUSIONS: Intensive and skillful dietary intervention can succeed whether or not clients bring strong internal locus of control or social support to the diet change program. PMID- 11271694 TI - Nutrient intake of obese female binge eaters. AB - OBJECTIVE: To compare the 6-month change in selected nutrients and number of binge days (from 7-day food records) between obese binge eaters randomly assigned to either a behavioral self-management (BSM) or waiting list control (WLC) group. Within each of the 2 groups, the average intake of selected nutrients on binge and nonbinge days at baseline and at 6 months were compared. DESIGN: A randomized, controlled, intervention study with assessments at entry and 6 months later. SUBJECTS: Forty-six women in the BSM group and 36 in the WLC group completed the 6-month measurement. Participants were 25 to 50 years of age, 30 to 90 pounds overweight, did not have a history of physical or psychological illnesses, and scored 20 or greater on the binge eating scale. INTERVENTION: Participants in the BSM intervention received 6 months of weekly, 1-hour classes taught by registered dietitians. Participants in the WLC group were not contacted during the 6 months. OUTCOME MEASURES: The main outcome measures were change in energy consumed (kilocalories); percentage of energy from fat, protein, and carbohydrate; grams of fiber/1,000 kcal; and change in the number of self reported binge days. STATISTICAL ANALYSES: Weight at 6 months was compared using a 2-sample t test. The change in the number of binge days at 6 months and the amount of change in selected nutrients by group was compared using the 2-sample t test. The paired t test was used to compare the average nutrient intakes on binge and nonbinge days within groups. RESULTS: No significant difference was found in the 6-month change between groups in any of the selected nutrients. The BSM group reported a greater reduction in binge days between baseline and 6 months compared with the WLC group (mean 1.0 vs 1.7, P < 0.03). Within the BSM group at 6 months, energy intake and percentage of energy from fat on nonbinge days were significantly reduced compared with binge days. At baseline within the WLC group, energy intake increased and percentage of energy from protein decreased significantly on nonbinge days compared with binge days. Within the WLC group at 6 months, energy intake and percentage of energy from fat significantly decreased and percentage of energy from protein significantly increased on nonbinge days. CONCLUSIONS: Our results suggest that collecting dietary information from participants identified with binge eating disorder is challenging. Dietitians who conduct behavioral weight management programs may require additional training in identifying and understanding the psychological characteristics of participants with binge-eating disorder. PMID- 11271695 TI - Validity of a telephone-administered 24-hour dietary recall in telephone and non telephone households in the rural Lower Mississippi Delta region. AB - OBJECTIVE: To determine if 24-hour dietary recall data are influenced by whether data are collected by telephone or face-to-face interviews in telephone and non telephone households. DESIGN: Dual sampling frame of telephone and non-telephone households. In telephone households, participants completed a 24-hour dietary recall either by face-to-face interview or telephone interview. In non-telephone households, participants completed a 24-hour dietary recall either by face-to face interview or by using a cellular telephone provided by a field interviewer. SUBJECTS/SETTING: Four hundred nine participants from the rural Delta region of Arkansas, Louisiana, and Mississippi. MAIN OUTCOME MEASURES: Mean energy and protein intakes. STATISTICAL ANALYSES PERFORMED: Comparison of telephone and non telephone households, controlling for type of interview, and comparison of telephone and face-to-face interviews in each household type using unpaired t tests and linear regression, adjusting for gender, age, and body mass index. RESULTS: Mean differences between telephone and face-to-face interviews for telephone households were -171 kcal (P = 0.1) and -6.9 g protein (P = 0.2), and for non-telephone households -143 kcal (P = 0.6) and 0.4 g protein (P = 1.0). Mean differences between telephone and non-telephone households for telephone interviews were 0 kcal (P = 1.0) and -0.9 g protein (P = 0.9), and for face-to face interviews 28 kcal (P = 0.9) and 6.4 g protein (P = 0.5). Findings persisted when adjusted for gender, age, and body mass index. No statistically significant differences were detected for mean energy or protein intake between telephone and face-to-face interviews or between telephone and non-telephone households. APPLICATIONS/CONCLUSIONS: These data provide support that telephone surveys adequately describe energy and protein intakes for a rural, low-income population. PMID- 11271696 TI - A practical, theory-based approach to establishing school nutrition advisory councils. AB - This article describes a process for establishing school nutrition advisory councils (SNACs) as an integral part of a school environment approach to promoting the nutritional health of students. The application of social cognitive theory as the conceptual framework for SNACs is discussed and the steps for establishing councils, including describing the school food environment, recruiting council members and convening the council, are reviewed. Actions taken by SNACs to positively affect the school nutrition environment are also described. SNACs are 1 component of the Teens Eating for Energy and Nutrition at School (TEENS) study, a group randomized, school-based intervention trial conducted in 16 middle schools in the Minneapolis-St Paul, Minn, metropolitan area. TEENS seeks to promote healthful dietary behaviors among young adolescents to reduce future cancer risk. Primary outcome measures include increasing fruit and vegetable intake and decreasing fat consumption. SNACs were established in the intervention schools to assess the overall school food environment and to advance school-level policy that promotes a healthful food environment. PMID- 11271697 TI - Prophylactic and therapeutic uses of probiotics: a review. AB - Probiotics, live microbial food supplements that beneficially affect the host by improving its intestinal microbial balance, are quickly gaining interest as functional foods in the current era of self-care and complementary medicine. Microbes have been used for years in food and alcoholic fermentations and relatively recently have undergone scientific scrutiny to examine their purported health benefits. Some of the claims for which research supports a beneficial effect of probiotic consumption include: improving intestinal tract health, enhancing the immune system, synthesizing and enhancing the bioavailability of nutrients, reducing symptoms of lactose intolerance, decreasing the prevalence of allergy in susceptible individuals, and reducing risk of certain cancers. The mechanisms by which probiotics exert their effects are largely unknown, but may involve modifying gut pH, antagonizing pathogens through production of antimicrobial and antibacterial compounds, competing for pathogen binding and receptor sites as well as for available nutrients and growth factors, stimulating immunomodulatory cells, and producing lactase. Selection criteria, efficacy, food and supplement sources and safety issues around probiotics are reviewed. Nutrition professionals can provide a tremendous service by helping clients overcome negative perceptions of all bacteria and, when appropriate, by developing individualized dietary plans to take advantage of the benefits probiotics may confer. PMID- 11271698 TI - Folate, zinc, and vitamin B-12 intake during pregnancy and postpartum. PMID- 11271699 TI - Breast-feeding practices of Native American mothers participating in WIC. PMID- 11271700 TI - Differences in dietary patterns of Vietnamese, white, African-American, and Hispanic adolescents in Worcester, Mass. PMID- 11271701 TI - Older women living in subsidized housing report low levels of nutrition support. PMID- 11271702 TI - Mutagenicity and genotoxicity of sorbic acid-amine reaction products. AB - Sorbic acid (E200) and its salts (potassium and calcium sorbate: E202 and E203) are allowed for use as preservatives in numerous processed foods. Sorbic acid had a conjugated system of double bonds which makes it susceptible to nucleophilic attack, sometimes giving mutagenic products. Under conditions typical of food processing (50-80 degrees C), we analysed the cyclic derivatives resulting from a double addition reaction between sorbic acid and various amines. Mutagenesis studies, involving Ames' test and genotoxicity studies with HeLa cells and plasmid DNA, showed that none of the products studied presented either mutagenic or genotoxic activities. PMID- 11271703 TI - 3-monochloropropane-1,2-diol (3-MCPD) in soy sauces and similar products available from retail outlets in the UK. AB - A survey of the level of 3-monochloropropane-1,2-diol (3-MCPD) in soy sauces and similar products available in the UK is reported. The survey was carried out by the Joint Ministry of Agriculture, Fisheries and Food/Department of Health Food Safety and Standards Group (JFSSG) to check for compliance with the Food Advisory Committee's (FAC) recommended limit for 3-MCPD of 0.01 mg/kg following reports that soy sauces in several European countries had been found to contain high levels (up to 124 mg/kg) of 3-MCPD. Forty samples of soy sauce and similar products purchased from retail outlets were analysed using a GC-MS procedure which had been formally validated by an earlier collaborative trial. 3-MCPD was undetectable in 21 (52%) of samples analysed in the survey, with a further five samples containing very low levels of between 0.01 and 0.02 mg/kg. Five samples (13%) contained 3-MCPD at levels in the range 0.020-1 mg/kg while nine samples (23%) were found to contain 3-MCPD at levels greater than 1 mg/kg, with the highest level being 30.5 mg/kg. PMID- 11271704 TI - Comparison of extraction techniques for the recovery of veterinary drug residues from animal tissues. AB - Four different techniques for the extraction of veterinary drugs were compared. The use of a high speed mixer/emulsifier, an ultrasonic bath, a Stomacher and an end-over-end mixer were used to extract both incurred and fortified residues of chlortetracycline, sulphadiazine and flumequine from chicken muscle. For each drug, similar analytical recoveries from fortified muscle samples were obtained using each of the extraction techniques. However, for each analyte the highest drug concentration detected in incurred samples was obtained following preparation using a mixer/emulsifier extraction. Residue concentrations obtained using sonication, Stomacher or end-over-end mixer procedures were as low as 20% of those obtained using the mixer/emulsifier. This highlights the need to measure both incurred and spiked samples during method validation. A survey of published methods indicated that 75% of laboratories use some form of high-speed homogenization for the extraction of drug residues from tissue. However only 52% reported detection of incurred residues. More than two-thirds of methods that used other forms of extraction did not measure incurred residues. The use of such methods has implications for the statutory detection of veterinary drug residues. PMID- 11271705 TI - Solid-phase microextraction coupled with high performance liquid chromatography: a complementary technique to solid-phase microextraction-gas chromatography for the analysis of pesticide residues in strawberries. AB - Solid-phase microextraction coupled with high performance liquid chromatography has been studied for the analysis of methiocarb, napropamide, fenoxycarb and bupirimate in strawberries. The strawberries were blended and centrifuged. Then, an aliquot of the resulting extracting solution was subjected to solid-phase microextraction (SPME) on a 60 microns polydimethylsiloxane/divinylbenzene (PDMS/DVB) fibre for 45 min at room temperature. The extracted pesticides on the SPME fibre were desorbed into SPME/high performance liquid chromatography (HPLC) interface for HPLC analysis with diode-array detection (DAD). The method is organic solvent-free for the whole extraction process and is simple and easy to manipulate. The detection limits were shown to be at low microgram kg-1 level and the linear response covered the range from 0.05 to 2 mg kg-1 of pesticides in strawberries with a regression coefficient larger than 0.99. A good repeatability with RSDs between 2.92 and 9.25% was obtained, depending on compounds. PMID- 11271706 TI - Estimation of the dietary intake of pesticide residues, lead, cadmium, arsenic and radionuclides in France. AB - Estimation of the dietary exposure of French consumers to 10 pesticides (omethoate, oxydemeton, phosalon, phosphamidon, triazophos, dicofol (op' + pp'), parathion ethyl, dichlorvos, procymidon and vinchlozolin), three heavy metals (lead, cadmium and arsenic) and three radionuclides (134caesium, 137caesium and 131iodine) from collected duplicate portion in mass catering establishments in 1998/1999 are reported, and compared with those from previous French studies as well as those from other countries. Dietary exposure estimates appear to be reassuring, in that Estimated Daily Intake (EDI) estimates are generally low, representing at maximum only 4% of the Acceptable Daily Intake (ADI) for pesticide residues and 28% of the Provisional Tolerable Weekly Intake (PTWI) for heavy metals. Moreover, none of the three radionuclides has been found in duplicate meals. When comparisons are possible, estimated dietary exposures for heavy metals are lower than those from previous French studies and similar or above those from other countries. PMID- 11271707 TI - Long-term survey of patulin in apple juice concentrates produced in Turkey. AB - A liquid chromatographic method described by us elsewhere was evaluated for a long-term survey of patulin in apple juice concentrates. Patulin was separated on a reversed phase C18 LC column with water-acetonitrile (99:1) as the mobile phase and quantitated with a photodiode array (PDA) detector. Relatively low amounts of patulin (< 5 micrograms/l for single strength juice at 11.2 degrees Bx) were detected in apple juice concentrates and confirmed by PDA detector, comparing the corresponding UV spectra with that of patulin standard. Four hundred and eighty two apple juice concentrates produced through 1996-99 were analysed for their patulin contents. Year-to-year variations in patulin levels of apple juice concentrates were found out to be statistically significant. Patulin contamination levels of apple juice concentrates tended to decrease through the years and averaged 63, 43, 19 and 31 micrograms/l in 1996, 1997, 1998 and 1999, respectively. Percentages of concentrates exceeding the maximum permitted concentration of 50 micrograms/l were 52%, 34%, 8% and 8% for 1996, 1997, 1998 and 1999, respectively. PMID- 11271708 TI - Migration of bisphenol-A diglycidyl ether (BADGE) and its reaction products in canned foods. AB - Bisphenol-A diglycidyl ether (BADGE) is used as an additive or starting agent in coatings for cans. The presence of hydrochloric acid in the organosol (PVC-based) lacquers results in formation of chlorohydroxy compounds of BADGE. These compounds, as well as BADGE itself, are potential migrants into the preserved food and are of toxicological concern. In the present investigation the presence of BADGE and the chlorohydroxy compounds (BADGE.HCl and BADGE.2HCl) in various kinds of canned foods from 30 brands have been determined by HPLC with fluorescence detection. BADGE was found in levels up to 5.1 mg/kg in the food and only in food from cans containing BADGE.HCl and BADGE.2HCl in the lacquers. BADGE was found both in fish in oil and in fish in tomato sauce, however, the highest amounts were found in the fatty foodstuffs. BADGE.HCl and BADGE.2HCl were found in concentrations up to 2.4 mg/kg and 8.3 mg/kg, respectively. Unlike BADGE, BADGE.2HCl was found in similar concentrations in fish in oil and in fish in tomato sauce. In aqueous and acidic foodstuffs BADGE readily hydrolyses into mono and dihydrolysed products (BADGE.H2O and BADGE.2H2O). In this study BADGE.H2O was not found in any food sample, whereas BADGE.2H2O was found in levels up to 2.6 mg/kg. The Scientific Committee for Food (SCF) of the European Commission has proposed that a limit of restriction of 1 mg/kg food shall include BADGE itself and BADGE.H2O, BADGE.HCl, BADGE.2HCl and BADGE.HCL.H2O. The present results indicate that the migration of BADGE.HCl and BADGE.2HCl, compounds with almost no data on toxicity, implies a greater problem than BADGE.H2O and BADGE.2H2O. PMID- 11271709 TI - A mistake in the JECFA recommendations on polydimethylsiloxane (PDMS). PMID- 11271710 TI - Impact of childhood rape and aggravated assault on adult mental health. AB - Associations among childhood assault (rape, aggravated assault, or both) and indices of adult mental health (posttraumatic stress disorder, major depressive episode) were examined in a national probability sample of 4,008 (weighted) women. Relationships among assault characteristics and these adult mental health indices were also investigated. Findings suggested particularly deleterious effects for childhood aggravated assault and rapes that caused additional physical injury. PMID- 11271711 TI - Adolescent sexual offenders: incidence of childhood maltreatment, serious emotional disturbance, and prior offenses. AB - Adolescents incarcerated for sexual offenses were compared to those incarcerated for other crimes on measures of prior child abuse and neglect, serious emotional disturbance, and involvement in child welfare and the juvenile justice system. Sexual or physical abuse was more common than neglect among sexual offenders with fewer than three prior maltreatment reports. Sexual offenders were twice as likely to be receiving special education services for severe emotional disturbance, and were usually incarcerated later than other offenders. Implications for research and practice are discussed. PMID- 11271712 TI - The economics of child sex-offender rehabilitation programs: beyond Prentky & Burgess. AB - In a 1990 article in this journal, Prentky and Burgess examined cost effectiveness of the rehabilitation of child molesters. Their estimates were based on the tangible costs of incarceration and particular recidivism rates. This paper extends those findings by estimating the intangible costs of child sexual abuse and a range of recidivism rates. The result is to focus greater attention on the efficacy of treatment programs and the potential economic damage done to children by child molesters. PMID- 11271713 TI - Exposure to violence, coping resources, and psychological adjustment of South African children. AB - The effects of exposure to direct and vicarious political, family, and community violence on the adjustment of 625 six-year-old black South African children was examined. Ambient community violence was most consistently related to children's psychosocial outcomes. Resources in the form of individual child resilience, maternal coping, and positive family relationships were found to mitigate the adverse impact in all the assessed domains of children's functioning. PMID- 11271714 TI - A prolegomenon on restraint of children: implicating constitutional rights. AB - News media and advocacy groups have brought to public attention a disturbing number of recent deaths proximal to the use of physical restraints. This paper examines the evidence indicating that use of these procedures can be dangerous to patients; explores the theoretical basis and practical application of restraints; and argues not only that their use may be unethical as a therapeutic intervention, but that it may have constitutional implications. PMID- 11271715 TI - Expectations and experiences of participants in ongoing adoption reunion relationships: a qualitative study. AB - Expectations of the adoption reunion process, responses to disappointments, and factors that influence reunion outcomes are investigated in a qualitative study of ten adult adoptees and ten birthmothers. Themes derived from interviews with participants are analyzed and explored, and implications for clinical practice and research are presented. PMID- 11271716 TI - Effects of war and organized violence on children: a study of Bosnian refugees in Sweden. AB - Data from 99 school-aged Bosnian refugee children living in Sweden were analyzed to reveal the patterns of war stress experienced and the relation between these stressors and current psychological problems. A significant pattern of associations emerged. When children had experienced much stress, talking about their experiences seemed to exacerbate their negative effects. PMID- 11271718 TI - A model of predictors and outcomes of outness among lesbian and bisexual women. AB - In a structural equation model of 2,401 lesbian and bisexual women, three variables of lesbian sexual identity were found to predict outness, which predicted lower psychological distress, which, in turn, predicted lower suicidality. The model held true for the subsamples of European-American and African-American women, and to a lesser extent for those of Latina, Asian American, Native-American, and Jewish women. PMID- 11271717 TI - A job and a home: social networks and the integration of the mentally disabled in the community. AB - Formerly hospitalized psychiatric patients participated in a program including skills in apartment living and employment. Contrasted to a comparison group, program graduates showed more multifunctional and more reciprocal relationships with other people. Results highlight the importance of work, housing, and the utilization of community services, and suggest the quality of supportive interpersonal relations as a criterion of program success. PMID- 11271720 TI - Ego development, self-perception, and self-complexity in adolescence: a study of female psychiatric inpatients. AB - A study of two groups of female psychiatric inpatients, differing in level of ego development, explored domains of self-perception that best predicted global self worth and symptom clusters that best predicted second-order factors of self perception. Findings revealed quantitative and qualitative differences in self complexities, and more positive self-perceptions among the higher ego-level group in scholastic competence, job competence, and behavioral conduct. Results are discussed from a developmental perspective. PMID- 11271719 TI - Quality of care for psychiatric emergency service patients presenting with substance use problems. AB - Psychiatric emergency service assessments of 683 patients were observed to better understand the quality of care substance users receive and the effects of clinicians' attitudes toward their patients on such care. Findings run counter to those of previous reports in that substance users, once recognized, are likely to receive better care than other patients. PMID- 11271721 TI - Subsyndromal depressive symptoms and major depression in postpartum women. AB - This study documents differences in the psychosocial functioning of women three months postpartum with subclinical depression, major depression prior to the birth of the baby, major depression both pre- and post-birth, and no depression. An understanding of these differences may have implications for intervention insofar as maternal depression places at risk not only the mother's functioning but her infant's development, as well. PMID- 11271722 TI - Depressive symptoms and suicidality in physically abused children. AB - Depressive symptoms and suicidality were assessed in 114 children 6-12 years old, of whom 41 had been physically abused, 38 neglected, and 35 neither abused nor neglected. The physically abused children manifested significantly higher levels of depressive symptomatology and suicidality than did the other two groups. Implications for research and clinical practice are discussed. PMID- 11271723 TI - Human immunodeficiency viruses types 1 and 2 infection among replacement blood donors in Mumbai (Bombay). AB - Between 1993-96, blood donated by 12,235 replacement blood donors was screened by various Enzyme Linked Immunosorbent Assays for detecting antibodies to Human Immunodeficiency Viruses types 1 and 2 according to the guidelines specified by Indian--Food and Drug Administration. 222 replacement blood donors (1.81%) were found to be seropositive for antibodies to Human Immunodeficiency Virus types 1 and 2. Furthermore, the ImmunoComb II HIV 1 & 2 BiSpot rapid sandwich ELISA in a comb format was used for differentially identifying HIV-1 and/or HIV-2 infection among these blood donors in Mumbai. Our data indicates that there is a low seroprevalence of HIV-1-2 infection among replacement blood donors in Mumbai (Bombay). Among them, while HIV-1 is still the predominant virus, dual HIV-1-2 and HIV-2 only infections are steadily increasing. PMID- 11271724 TI - Glutathione levels in health and sickness. AB - We speculate that the glutathione (GSH) status of human subjects could be an indicator of health and functional age. In this regard, in a study in which, 80 young and 40 elderly healthy individuals were selected as control. We also studied 145 patients with chronic illnesses namely, ischaemic heart disease, diabetes, preeclampsia, cataract, chronic renal failure and leukaemia (age 52 +/- 8.6 years). We observed that all the subjects had high malonadildehyde and low glutathione levels as compared to control. These early observations support the hypothesis that oxidative stress may have an important aetiological rule and antioxidants a potential therapeutic role. PMID- 11271726 TI - Abuse of neuroleptic drug by psychiatric patients. AB - Due to the depaminergic neurotrasmission in the mesocortical and mesolimbic reward systems, the neuroleptic drugs have been considered free from the risk of abuse or dependence because of their antidopaminergic properties. Here, two cases of neuroleptic abuse have been described and a possibility of reward circuits other than mesocortical and mesolimbic systems operational behind this phenomenon has been postulated. There is a need to clinically recognise the abuse potential of neuroleptic drugs. PMID- 11271725 TI - Drug resistance in tuberculosis patients in Jodhpur district. AB - Drug resistant tuberculosis is a serious problem in control of tuberculosis. To assess this problem in Jodhpur district, Sputum samples of symptomatic quarry workers and cases of pulmonary tuberculosis attending District Tuberculosis Clinic (DTC) Jodhpur were tested for culture of Mycobacterium tuberculosis complex (MTB) and their sensitivity to antituberculous drugs, using proportion method. Primary drug resistance to isoniazid was observed in 16.67%, to streptomycin in 16.67%, to ethambutol in 6.67% and to rifampicin in 6.67%. Acquired resistance to isoniazid was 61.76%, to streptomycin was 51.52%, to rifampicin was also 70.59%, and to ethambutol was 39.39%. Proportion of Multi drug resistant (MDR) TB defined as resistant to at least isoniazid and and rifampicin, was 3.3% in new cases (primary drug resistance) and 38.2% in old cases (acquired drug resistance), the later may be due to inadequate treatment, the the history of which was present in most cases. Adequate treatment of such cases with effective regimens is of vital importance to prevent the spread of MDR TB. PMID- 11271727 TI - Reappearance of Vibrio cholerae serogroup 0139 in Yavatmal during June-August 1998. PMID- 11271728 TI - Salmonella senftenberg: ear infection. A case report. AB - A case of Otitis media, a rare complication of Salmonella senftenberg infection is reported. PMID- 11271729 TI - Effects of crude drugs on glucose uptake in 3T3-L1 adipocytes. AB - In this study, various water-extracted crude drugs from Radix Asparagi, Radix Ginseng, Radix Scutellariae, Cortex Lycii Radicis, Cortex Phellodendri and Radix Ophiopogonis were investigated in their effects on [3H]-2-deoxyglucose uptake in 3T3-L1 adipocytes. Following treatment of cells with various crude drugs for 60 mim, the basal [3H]-2-deoxyglucose uptake in cultured 3T3-L1 cells was changed by Radix Asparagi from 140 pmole/min/mg protein of control to 513 (0.1 mg/ml), 201 (1 mg/ml) and 97 (10 mg/ml). Glucose uptake was changed to 324 (0.1 mg/ml), 146 (1 mg/ml) and 46 (10 mg/ml) with Radix Ginseng. In the presence of Radix Scutellariae, glucose uptake was changed to 215 (0.1 mg/ml), 213 (1 mg/ml) and 34 (10 mg/ml). In the presence of Cortex Lycii Radicis, glucose uptake was 230 (0.1 mg/ml), 188 (1 mg/ml) and 38 (10 mg/ml). In the case of Cortex Phellodendri and Radix Ophiopogonis, uptake was changed to 142 (0.1 mg/ml), 132 (1 mg/ml), 24 (10 mg/ml) and 489 (0.1 mg/ml), 374 (1 mg/ml), 344 (10 mg/ml), respectively. In insulin-stimulated cells, the [3H]-2-deoxyglucose uptake was changed by Radix Asparagi from 570 pmole/min/mg protein of the control to 816 (0.1 mg/ml), 674 (1 mg/ml) and 532 (10 mg/ml). After incubation with Radix Ginseng, the glucose uptake was changed to 254 (0.1 mg/mi), 123 (1 mg/mi) to 76 (10 mg/mi). In the presence of Radix Scutellariae, the glucose uptake was changed to 315 (0.1 mg/ml), 265 (1 mg/ml) and 33 (10 mg/ml). After incubation of Cortex Lycii Radicis, the uptake activity was changed to 281 (0.1 mg/ml), 248 (1 mg/ml) and 37 (10 mg/ml). In the case of Cortex Phellodendri and Radix Ophiopogonis, the activity of glucose uptake was measured as 747 (0.1 mg/ml), 523 (1 mg/ml), 33 (10 mg/ml) and 753 (0.1 mg/ml), 740 (1 mg/ml), and 421 (10 mg/ml), respectively. These results indicate that the water-extracted materials of Radix Asparagi and Radix Ophiopogonis increase the glucose uptake in basal and insulin-stimulated 3T3-L1 adipocytes. PMID- 11271730 TI - A survey on the intestinal parasites of the school children in Kaohsiung county. AB - The present study on the prevalence of intestinal parasites infection, from September to December 1999, was conducted among school children in Taoyuan Hsiang, Kaohsiung county. The investigated areas included three (Jiannshan, Shingjong and Taoyuan) primary schools. The overall infection rate in 305 children determined by Merthiolate-Iodine-Formaldehyde Concentration method of stools was 17%. Four confirmed species of helminthes (Ascaris lumbricoides, Hookworm, Trichuris trichiura and Hymenolepis nana) and three species of protozoa (Giardia lamblia, Entamoeba coli and Blastocystis hominis) were detected. Males and females had the infection rates of 24% and 11%. The infection rates of aboriginal and non-aboriginal children were 17% and 14%, respectively. Grade 1 and Grade 6 had the highest infection rate (21%). Following tape perianal examination of 302 children, the overall infection rate of Enterobius vermicularis was 25%. Males and females had the infection rates of 24% and 26%. The infection rates of aboriginal and non-aboriginal children were 27% and 11%, respectively. Grade 1 had the highest infection rate (37%). Based on these data, the infection rate of intestinal parasites among rural primary school children in Kaohsiung county remains high. PMID- 11271731 TI - Preliminary development and testing of instruments to measure self-care agency and social support of women in Taiwan. AB - The purpose of this study was to adapt and test the reliability and validity of the Personal Resource Questionnaire 85-Part 2 (PRQ85-Part 2) and Exercise of Self Care Agency Scale (ESCA) for women in Taiwan. Using a 3-phase study, the instruments were first translated into Chinese, back translated into English, and examined for content validity. In the second phase, the Chinese versions of the instruments were administered to 30 women, aged 60 to 75 years, and 34 women, aged 22 to 38 years. Modifications were made in the wording of items to make them more meaningful for the older women. In phase three, the instruments were administered to 284 women, 60 to 88 years of age. Indices of content validity were 1.0 for both the PRQ85-Part 2 and ESCA. Alpha reliability coefficients ranged from .84 to .90 for the PRQ85-Part 2 and from .86 to .92 for the ESCA. A test-retest reliability coefficient after one week was .80 for the PRQ85-Part 2 and .91 for the ESCA. The findings of this study indicate that both instruments are culturally appropriate for use with women in Taiwan. PMID- 11271732 TI - The study on core conflictual relationship of short-term counseling. AB - This study utilizes the Core Conflictual Relationship Theme Method (CCRT; Luborsky and Crits-Christoph, 1997) on process study of short-term counseling. The purpose was to evaluate the formation and variation of interpersonal conflicts presented by nine clients, so as to enhance the understanding of the core conflicts of each client and how these clients were influenced by short-term counseling. CCRT was conducted by two raters to analyze the content of core conflictual relationship themes. The raters started with taking a twenty-minute vignette from four tapes of the counseling sessions which were the 1st, 2nd, 11th and the 12th. A transcript was taken for each session and the raters searched for relationship episodes (REs) in it. Further, other main persons were identified in REs. Three components, including wish(W), response from others(RO), and response of self(RS) were investigated and analyzed. The raters calculated the appearance frequency of each component and transferred them into formulation of relationship theme in written language. The findings are as follows. (1) The interrater reliabilities between the two raters over W, RO, RS are significant (Kappa = .21 .82, P < .01) (2) The most frequently described other main persons in 201 REs were 'clleague' (N = 87, 43. 3%), the second 'self' (N = 42, 32.8%) and the third 'family members' (N = 41, 20. 4%). The first three described wishes were 'to assert self and be independent' (N = 66, 32.8%), 'to be close and accepted' (N = 48, 23.9%), and 'to be loved and understood' (N = 35, 17.4%) (3) The change of pervasiveness scores of each component in terms of positive and negative responses evaluated by t-test were found to decrease significantly in NRO (r = 2.73, p < .05). The score was most significantly decreased in 'rejecting and opposing' item (t = -3.83, p < .01). Conclusions and suggestions regarding application of the Core Conflictual Relationship Theme analytical and categorization methods in the Chinese context are presented. PMID- 11271733 TI - An application of the situational interview to selection of nursing personnel. AB - The study was designed to develop a situational interview technique to replace the unstructured interview commonly used in the selection process of nursing personnel and explore the applicability of incorporating the situational interview and psychological tests into the development of a regression model of nursing personnel selection. The authors collected from several teaching hospitals in Kaohsiung job-related critical incidents which were used to construct the questionnaire for the situational interview and the performance appraisal checklist. A sample of subjects, 198 in the first year but only 116 the second year, were interviewed and evaluated with the situational interview, the Nursing Image Scale, the Work Values Inventory, and part of the Gordon Personal Profile-Inventory as predictor measures and the performance appraisal checklist as the criterion measure. The results showed that the reliability and validity coefficients of the situational interview were close to or even better than those obtained by the previous researchers in the same field. The reliability and validity coefficients of the performance appraisal checklist indicated its applicability and suitability. Multiple regression analysis indicated that the situational interview as well as some traits measured by the personality and values tests could predict the performance appraisal scores of nursing personnel. PMID- 11271734 TI - Prosthesis folding as a cause of the saline breast implant partial deflation 12 years after augmentation mammaplasty: a case report. AB - In this paper, I report a case of delayed slow leakage (partial deflation) from an inflatable saline breast implant, 12 years after augmentation mammaplasty. The patient had a history of breast foreign body injection and developed a complication with multiple painful indurated mass. She received a subcutaneous mastectomy to remove all the injected breast foreign body in November 1986 and an inflatable breast implant was given for augmentation simultaneously. Owing to the accidental deflation of the left implant, the breast implant was removed and replaced in January 1999. Examination of the removed breast implant revealed a linear fold (13 cm in length) on the lower pole of the prosthesis and a small leaking hole (microscopic tear) in the middle of that fold. The possible mechanisms of implant deflation may include faulty manufacturing process, fold flaw leak (crease fold failure) and capsular contracture. In this case report the capsular contracture had caused prosthesis folding and the prosthesis had become partial deflation finally. Besides the better surgical intervention to prevent capsular contracture, I believe that choosing a correct implant size preoperatively and filling that implant to its optimal level will best minimize the possibility of breast silicone shell folding. PMID- 11271735 TI - Partial anomalous pulmonary venous connection draining into superior vena cava- two cases reports. AB - Anomalous pulmonary venous connection is a relatively common associated anomaly in patients with atrial septal defect (ASD), particularly among those with the sinus venosus type. The incidence of partial anomalous pulmonary venous connection (PAPVC) is higher than 0.7% in the general population and 10% in patients with ASD. In this study, we present two cases with initial impression of ASD, the sinus venosus type in one and the secundum type in the other. The one with the sinus venosus type was found to have a PAPVC that drained into SVC, and the other was suspected of having the same problem because an abnormal shunt was found during cardiac catheterization. This speculation could not be proved, however, due to transesophageal echocardiogram failure. Because we feared the possibility of cardiac defects other than ASD, we performed a minimally invasive operation using a small midline incision instead of the submammary incision and did a full median sternotomy on the patient to look for other complicating coexistent cardiac defects. This patient and the former one were both proven intraoperatively to have a PAPVC that drained into SVC with sinus venosus ASD. The operation to correct an ASD is a rudimentary procedure, and it often becomes a common type of minimally invasive operation among young cardiac surgeons with limited experience. A submammary incision under the impression of simple ASD may meet with certain complications. Therefore, after our experience with the latter case, we do the minimally invasive operation using a small midline incision, which can be easily extended if need be. PMID- 11271736 TI - The effect of the teaching physician rule on residency education. AB - BACKGROUND AND OBJECTIVES: In 1996, the Health Care Finance Administration implemented the Teaching Physician Rule (TPR) to clarify the responsibilities of attending physicians when they are supervising residents and billing Medicare for that service. We measured some of the effects of the TPR on family practice residency training. METHODS: After pilot testing, a questionnaire was mailed to the directors of all family practice residency programs in the United States. The directors were asked to provide a similar questionnaire to a senior resident. RESULTS: Of 449 residency directors, 310 (69%) responded. Eighty percent of residencies apply the TPR to at least some patient encounters. Residency directors reported that the TPR had an overall negative effect on their residency. Residents reported a more negative impression of the rule than did the directors. On average, residency directors reported that the mandated level of supervision in the outpatient setting increased faculty attending time by .24 FTE. CONCLUSIONS: The TPR was perceived by residency directors and senior residents to have some negative effect on family practice residency programs, at least in part by increasing the need for more faculty time for supervision. PMID- 11271737 TI - Fetal biometry: a comparison of family physicians and radiologists. AB - BACKGROUND AND OBJECTIVES: While performance and reading of obstetric sonograms is a skill widely taught in family practice residency programs, no prior studies have compared ultrasound performance and interpretation in residency programs with that of hospital radiologists. This study compares results of fetal biometry for gestational age determination performed sequentially by faculty-supervised residents and radiologists. METHODS: This retrospective chart review selected cases from among all gravidas who had ultrasound performed in a family practice residency clinic between January 1992 and April 1999. Biometry was performed by residents under the supervision of faculty preceptors who had ultrasound training and experience. A patient was included if (1) results of both a family practice ultrasound and a radiologist-read hospital ultrasound were present in the chart, (2) both studies were done before 36 weeks gestation, and (3) the family practice examination preceded the hospital study. The difference in expected date of confinement between resident and radiologist ultrasound was calculated, and this difference was evaluated for statistical significance by a paired sample t test. RESULTS: Ninety-two ultrasound pairs were assessed, a sample size that provided .90 power to detect a gestational age estimate difference of 3 days between family practice and radiologist interpretations. The normally distributed observed mean difference in gestational age estimates was only 1.5 days. CONCLUSIONS: This study found no difference in gestational age assessment performed by closely supervised family practice residents in comparison to radiologists. PMID- 11271738 TI - The lack of screening for diabetic nephropathy: evidence from a privately insured population. AB - BACKGROUND: We examined the performance of screening tests for diabetic nephropathy in a population of privately insured individuals. METHODS: Administrative data from a large private health plan were analyzed. Continuously insured persons with diabetes (ages 30-62) with > or = one office visit during the study year (July 1995 to June 1996) were included (n = 4,758). Outcome variables included a urinalysis for protein and a test for microalbuminuria. The likelihood of test performance according to age, gender, insurance plan type, total office visits, diabetes office visits, and specialty of predominant physician was examined both in bivariate analyses and a logistic regression. RESULTS: Among the 4,623 patients without evidence of nephropathy, only 16.5% had a urinalysis test conducted sometime in the year. All individuals (2.1% of sample) who received a microalbuminuria test also received a urinalysis. Individuals with indemnity or PPO plans were more likely to be screened than individuals in point-of-service plans. Patients with more visits and more diabetes visits were more likely to be screened. In the regression with family practice as the reference category, general internists were the only physician specialty more likely to have screened patients. CONCLUSIONS: The majority of patients with diabetes mellitus do not receive annual screening for microalbuminuria or urinary protein. PMID- 11271739 TI - Shortchanging adolescents: room for improvement in preventive care by physicians. AB - BACKGROUND AND OBJECTIVES: Behaviors developed in adolescence influence health later in life. Adolescents seldom visit physicians to discuss health-related behaviors. Instead, physicians must incorporate health counseling into the exams for which the adolescents do come. We studied the frequency and duration of adolescents' consultations with family physicians and pediatricians involving counseling about diet and nutrition, exercise, weight reduction, cholesterol reduction, HIV transmission, injury prevention, and tobacco use. METHODS: Data were analyzed from the National Ambulatory Medical Care Survey for the 3-year period from 1995 through 1997. This survey uses a multistage national probability sample of patient visits to nonfederal, office-based physicians. We described patterns of counseling provided to adolescents and compared patterns for family physicians/general practitioners and pediatricians. RESULTS: Of 91,395 physician reported visits analyzed, 4,242 (4.6%) were by adolescents ages 12-19. Visits to family physicians and pediatricians accounted for 1,846 (43.5%) of these visits. Counseling about any of the seven areas studied was included in 15.8% of family physician visits and 21.6% of pediatrician visits. The length of consultation increased from 13.8 to 17.6 minutes if counseling was included. CONCLUSIONS: Adolescents visits physicians infrequently. When they do, few receive counseling on critical adolescent health issues. Both family physicians and pediatricians have room for improvement. PMID- 11271740 TI - Family medicine faculty development fellowships and the medically underserved. AB - OBJECTIVE: This study measured the prevalence of service in federally designated medically underserved communities (FD-MUC) by Title VII-funded, full-time faculty development fellowship alumni. METHODS: A two-stage survey of alumni of full time, family medicine faculty development fellowships was completed. Alumni were dichotomized as serving in an FD-MUC or not. RESULTS: Of the 105 fellowship alumni identified, 81% (n = 85) responded; 42% (n = 36) were serving in an FD MUC. Of alumni serving in an FD-MUC, the mean full-time equivalent service time was 73%. Of the demographic variables measured, only race was significantly associated with FD-MUC service, and minorities were more likely to practice in an FD-MUC. Respondents serving in FD-MUCs were more satisfied with their relationships with nonphysician health professionals, salary and income, and their role in making organizational and administrative decisions than those not serving in FD-MUCs. CONCLUSIONS: Title VII has the broad policy objective of increasing access to medical care by improving the supply and distribution of physicians and recruitment of minority health professionals. Alumni of faculty development programs have a high service rate in FD-MUCs, and minority alumni are significantly more likely to practice in these sites. PMID- 11271741 TI - Electronic medical records: the family practice resident perspective. AB - BACKGROUND AND OBJECTIVES: Few studies have included family practice residents' perceptions regarding the use of electronic medical records (EMR) in a residency program. This study determined residents' perceptions of EMR systems and what variables influenced those perceptions. Specifically, we studied how EMR training and previous computer background influenced resident perception of difficulty in EMR implementation, time efficiency, preventive care opportunities, accuracy of medical records, and desired future use of EMR systems. Questions targeted the use of the EMR in the resident continuity clinic. METHODS: Survey questionnaires about the use of EMR systems in the residents' continuity clinic were mailed to residency directors and residents of 219 family practice residency programs. Respondents were given the opportunity to comment on benefits of and concerns about EMR. RESULTS: Resident response rate was 46% in programs using EMR systems. Findings revealed that the length of EMR training a resident received was unrelated to the perceived adequacy of training but was related to the residents' assessment of the difficulty of implementing the EMR in their continuity clinic. Residents who perceived training to be adequate and perceived a relative ease of implementing the EMR were more likely to perceive the EMR to be beneficial and were more likely to choose the EMR over traditional paper records for future use. Computer background/experience was not related to perceived satisfaction with the EMR, nor was it related to perceived difficulty of implementation, adequacy of training, or anticipated future use of an EMR system. CONCLUSIONS: Although residents recognize the benefits of the EMR, our study demonstrates an overall ambivalence and frustration toward EMR systems currently in use in family practice residency continuity clinics. However, the training they receive regarding EMR use in their residency may influence not only the perceived ease of EMR implementation but attitudes regarding the ability of the EMR to assist them with preventive opportunities, time management, and medical record accuracy. This may in turn have influence on the use of EMR systems in their practices after residency. PMID- 11271742 TI - Leading change versus managing care: the role of change agent in family medicine. PMID- 11271743 TI - Bias shown in study of better care for patients with skin disease? PMID- 11271744 TI - Support urged for accreditation of faculty development fellowships. PMID- 11271745 TI - The call for moral leadership. PMID- 11271746 TI - On the rise and fall of videotaping programs. PMID- 11271748 TI - What is a physician? PMID- 11271747 TI - Doctor lounge talk. AB - The discussions that take place in the doctor's lounge are essential to the well being of the preceptor. Doctor lounge talk should be filtered for students with their level of development in mind. PMID- 11271749 TI - Interpretation of the American Board of Family Practice In-training Examination. AB - The American Board of Family Practice (ABFP) In-training Examination (ITE) is the most frequently used method of evaluation among accredited family practice programs. Despite its common use, there are still misunderstandings regarding the proper interpretation of the ABFP ITE results. The ABFP ITE is a norm-referenced test that allows for comparison of a resident's performance relative to other residents in the same year of training; it does not measure absolute performance. There is a paucity of evidence supporting the validity of the ABFP ITE. The eight specialty areas and clinical set problems lack sufficient reliability to be interpreted per individual resident. The composite score of the ABFP ITE has sufficient reliability to be used as part of a formal and comprehensive evaluation system. Group comparisons using the 10 ABFP ITE measures are appropriate. PMID- 11271751 TI - A social relations variance partitioning of dyadic behavior. AB - The authors investigate the relative importance of actor and interaction partner as determinants of dyadic behavior. Using the social relations model (D. A. Kenny, 1994a; D. A. Kenny & L. La Voie, 1984), the authors estimate the variance attributable to each determinant plus the reciprocity of behavioral responses from 7 studies. The authors find evidence for moderate behavioral consistency in a person's behavior across interaction partners, little or no evidence that people consistently engender the same behavioral response from others, and preliminary evidence of unique responding to particular partners. They also consider several methodological issues concerning behavioral measurement as well as the implications of the results for the study of accuracy. PMID- 11271750 TI - The role of transesophageal echocardiography in the diagnosis and management of patients with aortic perivalvular abscesses. AB - Aortic valve abscesses (AVAs) are a devastating complication of aortic valve endocarditis. Over 8 years, 25 patients were diagnosed with AVA by transesophageal echo (TEE). Management and outcomes were then analyzed. Eleven (44%) AVAs involved prosthetic valves, and 6 (24%) occurred in congenitally malformed valves. Twenty patients (80%) underwent surgical intervention; the rest were treated medically. Eleven (44%) of the patients died [6 (30%) surgery patients and all the medical patients]. Eight of 11 (73%) patients who died were culture positive for Staphylococcus aureus. All patients with congenitally malformed aortic valves underwent surgical intervention and survived. We conclude that: (1) despite advances in therapy and diagnosis, patients with AVAs have a high mortality rate; (2) prognosis with AVA is especially poor when S aureus is the infectious organism; (3) patients with AVAs in congenitally malformed valves have a great outcome with surgery; (4) patients treated medically have a very poor prognosis; earlier identification by TEE may be critical to improving survival. PMID- 11271753 TI - A comprehensive meta-analysis of the predictive validity of the graduate record examinations: implications for graduate student selection and performance. AB - This meta-analysis examined the validity of the Graduate Record Examinations (GRE) and undergraduate grade point average (UGPA) as predictors of graduate school performance. The study included samples from multiple disciplines, considered different criterion measures, and corrected for statistical artifacts. Data from 1,753 independent samples were included in the meta-analysis, yielding 6,589 correlations for 8 different criteria and 82,659 graduate students. The results indicated that the GRE and UGPA are generalizably valid predictors of graduate grade point average, 1st-year graduate grade point average, comprehensive examination scores, publication citation counts, and faculty ratings. GRE correlations with degree attainment and research productivity were consistently positive; however, some lower 90% credibility intervals included 0. Subject Tests tended to be better predictors than the Verbal, Quantitative, and Analytical tests. PMID- 11271752 TI - Theories of reasoned action and planned behavior as models of condom use: a meta analysis. AB - To examine how well the theories of reasoned action and planned behavior predict condom use, the authors synthesized 96 data sets (N = 22,594) containing associations between the models' key variables. Consistent with the theory of reasoned action's predictions, (a) condom use was related to intentions (weighted mean r. = .45), (b) intentions were based on attitudes (r. = .58) and subjective norms (r. = .39), and (c) attitudes were associated with behavioral beliefs (r. = .56) and norms were associated with normative beliefs (r. = .46). Consistent with the theory of planned behavior's predictions, perceived behavioral control was related to condom use intentions (r. = .45) and condom use (r. = .25), but in contrast to the theory, it did not contribute significantly to condom use. The strength of these associations, however, was influenced by the consideration of past behavior. Implications of these results for HIV prevention efforts are discussed. PMID- 11271754 TI - From physical time to the first and second moments of psychological time. AB - After examination of the status of time in experimental psychology and a review of related major texts, 2 opposite approaches are presented in which time is either unified or fragmented. Unified time perception views, usually guided by Weber's law, are embodied in various models. After a brief review of old models and a description of the major contemporary models of time perception, views on fragmented time perception are presented as challenges for any unified time view. Fragmentation of psychological time emerges from (a) disruptions of the Weber function, which are caused by the types of interval presentation, by extensive practice, and by counting explicitly or not; and (b) modulations of time sensitivity and perceived duration by attention and interval structures. Weber's law is a useful guide for studying psychological time, but it is also reasonable to assume that more than one so-called central timekeeper could contribute to perceiving time. PMID- 11271755 TI - Event structure in perception and conception. AB - Events can be understood in terms of their temporal structure. The authors first draw on several bodies of research to construct an analysis of how people use event structure in perception, understanding, planning, and action. Philosophy provides a grounding for the basic units of events and actions. Perceptual psychology provides an analogy to object perception: Like objects, events belong to categories, and, like objects, events have parts. These relationships generate 2 hierarchical organizations for events: taxonomies and partonomies. Event partonomies have been studied by looking at how people segment activity as it happens. Structured representations of events can relate partonomy to goal relationships and causal structure; such representations have been shown to drive narrative comprehension, memory, and planning. Computational models provide insight into how mental representations might be organized and transformed. These different approaches to event structure converge on an explanation of how multiple sources of information interact in event perception and conception. PMID- 11271756 TI - From mice to men: what can we learn about personality from animal research? AB - The author explores the viability of a comparative approach to personality research. A review of the diverse animal-personality literature suggests that (a) most research uses trait constructs, focuses on variation within (vs. across) species, and uses either behavioral codings or trait ratings; (b) ratings are generally reliable and show some validity (7 parameters that could influence reliability and 4 challenges to validation are discussed); and (c) some dimensions emerge across species, but summaries are hindered by a lack of standard descriptors. Arguments for and against cross-species comparisons are discussed, and research guidelines are suggested. Finally, a research agenda guided by evolutionary and ecological principles is proposed. It is concluded that animal studies provide unique opportunities to examine biological, genetic, and environmental bases of personality and to study personality change, personality-health links, and personality perception. PMID- 11271757 TI - Coping with stress during childhood and adolescence: problems, progress, and potential in theory and research. AB - Progress and issues in the study of coping with stress during childhood and adolescence are reviewed. Definitions of coping are considered, and the relationship between coping and other aspects of responses to stress (e.g., temperament and stress reactivity) is described. Questionnaire, interview, and observation measures of child and adolescent coping are evaluated with regard to reliability and validity. Studies of the association of coping with symptoms of psychopathology and social and academic competence are reviewed. Initial progress has been made in the conceptualization and measurement of coping, and substantial evidence has accumulated on the association between coping and adjustment. Problems still remain in the conceptualization and measurement of coping in young people, however, and aspects of the development and correlates of coping remain to be identified. An agenda for future research on child-adolescent coping is outlined. PMID- 11271759 TI - Comparison of chlorine and a prototype produce wash product for effectiveness in killing Salmonella and Escherichia coli O157:H7 on alfalfa seeds. AB - Outbreaks of Salmonella and Escherichia coli O157:H7 infections associated with alfalfa and other seed sprouts have occurred with increased frequency in recent years. This study was undertaken to determine the efficacy of a liquid prototype produce wash product (Fit), compared with water and chlorinated water, in killing Salmonella and E. coli O157:H7 inoculated onto alfalfa seeds. We investigated the efficacy of treatments as influenced by seeds from two different lots obtained from two seeds suppliers and by two methods of inoculation. The efficacy of treatments was influenced by differences in seed lots and amount of organic material in the inoculum. Significant (alpha = 0.05) reductions in Salmonella populations on seeds treated with 20,000 ppm of chlorine or Fit for 30 min ranged from 2.3 to 2.5 log10 CFU/g and 1.7 to 2.3 log10 CFU/g, respectively. Reductions (alpha = 0.05) in E. coli O157:H7 ranged from 2.0 to 2.1 log10 CFU/g and 1.7 to more than 5.4 log10 CFU/g of seeds treated, respectively, with 20,000 ppm of chlorine or Fit. Compared with treatment with 200 ppm of chlorine, treatment with either 20,000 ppm of chlorine or Fit resulted in significantly higher reductions in populations of Salmonella and E. coli O157:H7. None of the treatments eliminated these pathogens as evidenced by their detection on enrichment of treated seeds. Considering the human health and environmental hazards associated with the use of 20,000 ppm of chlorine, Fit provides an effective alternative to chlorine as a treatment to significantly reduce bacterial pathogens that have been associated with alfalfa seeds. PMID- 11271758 TI - Quantitative determination of the role of lettuce leaf structures in protecting Escherichia coli O157:H7 from chlorine disinfection. AB - Viability of Escherichia coli O157:H7 cells on lettuce leaves after 200 mg/liter (200 ppm) chlorine treatment and the role of lettuce leaf structures in protecting cells from chlorine inactivation were evaluated by confocal scanning microscopy (CSLM). Lettuce samples (2 by 2 cm) were inoculated by immersing in a suspension containing 10(9) CFU/ml of E. coli O157: H7 for 24+/-1 h at 4 degrees C. Rinsed samples were treated with 200 mg/liter (200 ppm) chlorine for 5 min at 22 degrees C. Viability of E. coli O157:H7 cells was evaluated by CSLM observation of samples stained with Sytox green (dead cell stain) and Alexa 594 conjugated antibody against E. coli O157:H7. Quantitative microscopic observations of viability were made at intact leaf surface, stomata, and damaged tissue. Most E. coli O157:H7 cells (68.3+/-16.2%) that had penetrated 30 to 40 microm from the damaged tissue surface remained viable after chlorine treatment. Cells on the surface survived least (25.2+/-15.8% survival), while cells that penetrated 0 to 10 microm from the damaged tissue surface or entered stomata showed intermediate survival (50.8 +/-13.5 and 45.6+/-9.7% survival, respectively). Viability was associated with the depth at which E. coli O157:H7 cells were in the stomata. Although cells on the leaf surface were mostly inactivated, some viable cells were observed in cracks of cuticle and on the trichome. These results demonstrate the importance of lettuce leaf structures in the protection of E. coli O157:H7 cells from chlorine inactivation. PMID- 11271760 TI - Efficacy of washing with a commercial flatbed brush washer, using conventional and experimental washing agents, in reducing populations of Escherichia coli on artificially inoculated apples. AB - Conventional and experimental washing formulations were applied with a commercial flatbed brush washer under conditions representative of commercial practice to determine their efficacy in decontaminating apples inoculated with a nonpathogenic Escherichia coli strain. Golden Delicious apples (18 kg) inoculated with E. coli were mixed with approximately 109 kg of uninoculated Fuji apples (distinctly different in appearance) in a wet dump tank containing 1,325 liters of water at 20 degrees C for 15 min. The combined apples were washed in a flatbed brush washer with the following washing solutions: water at 20 degrees C, water at 50 degrees C, 200 ppm of chlorine (pH 6.4) at 20 degrees C, 8% trisodium phosphate at 20 degrees C, 8% trisodium phosphate at 50 degrees C, 5% hydrogen peroxide at 20 degrees C, 5% hydrogen peroxide at 50 degrees C, 1% APL Kleen 245 at 50 degrees C, and two-stage washing treatments using the combination of 1% APL Kleen 245 at 20 or 50 degrees C followed by 5% hydrogen peroxide at 35 or 50 degrees C. None of the washing treatments tested under the conditions of this experiment significantly reduced the E. coli populations on the inoculated apples or in cider made from these apples, probably as a consequence of the inability of this washing system to inactivate or remove the bacterial cells in inaccessible calyx and stem areas of apples. These results are important because they demonstrate the need for new fruit washing technology that can overcome this limitation. Also, there was no significant cross-contamination of the Fuji apples in the dump tank. Significant cross-contamination of cider, made with uninoculated apples, occurred in the hammer mill and/or the press cloth when these units were not sanitized following a trial with inoculated apples. PMID- 11271761 TI - Polymerase chain reaction for the direct detection of Brucella spp. in milk and cheese. AB - A polymerase chain reaction test was developed to detect Brucella spp. directly in milk and cheese and optimized using primers for the BSCP-31 gene. A total of 46 cheese samples produced with sheep and goats milk were assayed, and Brucella spp. was detected in 46% of them, especially in cheese made from sheep milk. This method is of remarkable epidemiologic interest because it is an indirect test indicating the sanitary quality of milk used in dairy industries. The method showed good sensitivity and specificity. It is faster and less expensive than the conventional bacteriological assays. PMID- 11271762 TI - Application of multiple antimicrobial interventions for microbial decontamination of commercial beef trim. AB - Commercially produced, irregularly sized (range, 100 to 400 cm2), uninoculated beef trim was treated by a previously optimized multihurdle antimicrobial process under spray system or hot air gun with set-up speed (1 cm/s): W (water wash at 65 psi for five passes) + HW (82 degrees C water at 30 psi for three passes) + HA (510 degrees C air for five passes) + L (2% [vol/vol] room temperature lactic acid wash at 30 psi for three passes). After treatment, the trim was finely ground, vacuum packaged, and stored at 4 degrees C for up to 20 days. At regular intervals (0, 5, 10, 15, and 20 days of storage at 4 degrees C), the ground beef was analyzed to measure mesophilic aerobic bacteria (APC), coliforms, psychrotrophic bacteria (PCT), and presumptive lactic acid bacteria (PLAB) and compared with the untreated control. The numbers of APC, coliforms, PCT, and PLAB were reduced to nearly nondetectable levels immediately after treatment, with significant differences compared with the control (P < 0.05), then started to increase after 5 to 10 days of storage at 4 degrees C. After 20 days, microbial populations of treated ground beef were significantly lower than those of nontreated ground beef for the numbers of APC, coliforms, PCT, and PLAB (P < 0.05), with differences of 1.2, 2.4, 1.6, and 1.6 log CFU/g, respectively. Based on microbial reduction and quality aspects, the multihurdle antimicrobial process was identified as an effective intervention to reduce coliforms on beef trim. PMID- 11271763 TI - Extent of microbial contamination in United States pork retail products. AB - To determine the extent of microbiological contamination of U.S. pork, 384 samples of retail pork were collected from 24 stores in six cities, including (i) whole-muscle, store-packaged pork; (ii) fresh, store-packaged ground pork and/or pork sausage; (iii) prepackaged ground pork and/or pork sausage; and (iv) whole muscle, enhanced (injected or marinated; 60% store-packaged, 40% prepackaged) pork. Additional samples (n = 120) of freshly ground pork and/or pork sausage were collected from two hot-boning sow/boar sausage plants, two slaughter and fabrication plants, and two further-processing plants. Samples were analyzed for aerobic plate counts (APC), total coliform counts (TCC), Escherichia coli counts (ECC), and incidences of Salmonella spp., Listeria monocytogenes, Campylobacter jejuni, Campylobacter coli, and Yersinia enterocolitica. Mean log APC and TCC were highest (P < 0.05) for store-ground pork, while whole-muscle, enhanced products and prepackaged ground products had the lowest (P < 0.05) APC. Mean log APC and TCC were higher (P < 0.05) in samples from the slaughter and fabrication plants than in samples from hot-boning and further processing plants. Mean log ECC were lower (P < 0.05) in samples from further-processing plants compared to slaughter and fabrication plants and hot-boning, sow and boar sausage plants. L. monocytogenes was detected in 26.7% of plant samples and 19.8% of retail samples and was present more frequently in ground products. Y. enterocolitica was detected most often in whole-muscle, store-packaged cuts (19.8%) and in store ground product (11.5%). Salmonella spp. were found in 9.6% of retail samples and 5.8% of plant samples, while C. jejuni and C. coli were found in 1.3% of retail samples and 6.7% of plant samples. Pork products exposed to the most handling and processing appeared to be of the poorest microbiological quality. These results should be useful in risk assessments that are directed at the identification of actions that could enhance food safety. PMID- 11271764 TI - Quantitative investigation of the effects of chemical decontamination procedures on the microbiological status of broiler carcasses during processing. AB - The effects of elevated chlorine concentrations (25 ppm) added to water in the final carcass washing equipment on total viable counts (TVCs 22 degrees C) and Escherichia coli and Enterobacteriaceae levels on poultry carcasses were investigated. Mean TVC counts on neck skin samples were significantly reduced when pre-evisceration and postwash samples were compared with log10 4.98 to 4.52 CFU/g recovered, respectively (P < or = 0.05). No significant reductions in TVC counts were observed in control samples at corresponding sampling points subjected to wash water containing 1 to 2 ppm chlorine. E. coli and Enterobacteriaceae counts were not significantly altered following final carcass washing in the processing plant. A second trial assessed the microbial decontamination capabilities of sodium triphosphate (TSP) on broiler carcasses. Neck skin samples from carcasses were obtained before final washing (control), following a 15-s dip in potable water and after dipping in a 10% TSP solution (pH 12) for 15 s. Reductions in E. coli and Enterobacteriaceae counts were all statistically significant for both water and TSP-treated samples when compared with corresponding controls (P < or = 0.01). The TSP treatment resulted in higher reductions of log10 1.95 and 1.86/g for E. coli and Enterobacteriaceae, respectively. In contrast, reductions of log10 0.37 and 0.3 l/g were observed for E. coli and Enterobacteriaceae counts when water-dipped carcasses were compared with corresponding controls. Significantly, Salmonella was not detected in any of the TSP-treated carcasses, while log10 1.92 and 1.04/g were found in control and water-dipped samples, respectively. Thermophilic Campylobacter counts were significantly lower in both treatment groups when compared with corresponding controlsresulting in log10 0.55 and 1.71/g reductions for water- and TSP-dipped carcasses, respectively (P < or = 0.01). PMID- 11271765 TI - Presence and level of Campylobacter, coliforms, Escherichia coli, and total aerobic bacteria recovered from broiler parts with and without skin. AB - This study was undertaken to determine if broiler chicken parts without skin are less contaminated with Campylobacter than those with skin. Samples were taken in a commercial plant from defeathered carcasses before evisceration. Bacterial counts from rinse of aseptically removed meat samples were lower than those from stomached skin samples. No Campylobacter were recovered from meat collected from the breasts or thighs, and only 2 of 10 drumstick meat samples had detectable levels of Campylobacter. However, 9 of 10 breast skin, 10 of 10 thigh skin, and 8 of 10 drumstick skin samples were positive for Campylobacter, with between 2 and 3 log10 CFU/g of Campylobacter. Breasts, thighs, and drumsticks were removed from broiler carcasses following evisceration before entering the chill tank. There was a significant difference (50 to 90%) in the levels of Campylobacter on breasts, thighs, and drumsticks with and without skin. Similar trends were noted for coliform, Escherichia coli, and total aerobic bacterial counts from samples collected in the plant. Broiler part samples were also collected at retail outlets. These samples were either skin on and skinned in the laboratory or skin off at purchase. Aseptic removal of skin from broiler breasts, thighs, and drumsticks did not cause change in Campylobacter, coliform, E. coli, or total aerobic counts recovered from the skinned part. Likewise, parts purchased without skin did not have different bacterial counts than paired parts purchased with the skin on. Consumers should not expect to significantly lower the number of bacteria present on a chicken breast, thigh, or drumstick by removing the skin. PMID- 11271766 TI - Novel quantitative assays for estimating the antimicrobial activity of fresh garlic juice. AB - Novel agar diffusion and broth dilution assays were developed for quantitatively estimating the antimicrobial activity of fresh garlic juice. Bacteria found to be inhibited by garlic juice in agar diffusion assay included two gram-positive and five gram-negative species. Leuconostoc mesenteroides was not inhibited. Escherichia coli B-103 (HB101, with pJH101, ampicillin resistant, 100 microg ml( 1)) was inhibited and chosen as the standard culture for quantitative assays. The agar diffusion assay was based on the slope ratio method, where the slope of dose response for garlic juice was divided by the slope of dose response for methylmethane thiosulfonate (MMTSO2). Juice from fresh garlic varied in activity between 1.76 and 2.31 microg of MMTSO2 per mg of garlic juice. The activity of juice decreased during 11 months of storage of garlic cloves at 5 degrees C from 2.31 to less than 0.1 microg of MMTSO2 per mg of juice. The broth dilution assay also used the E. coli B-103 culture, which permitted selective enumeration of this bacterium when 100 microg ml(-1) of ampicillin was incorporated into the enumerating agar. Selective enumeration was essential since the garlic juice was not sterile and, thus, contained natural flora. Growth of E. coli was unaffected by 0.1%, delayed by 0.25%, and completely inhibited at 0.5 and 2% garlic juice in broth during 24 h of incubation at 37 micro C. The minimum inhibition concentration of garlic juice by broth dilution assay was, thus, estimated to be 0.5%, which is equivalent to 3.46 microg of MMTSO2 per mg of garlic juice by the agar diffusion assay. PMID- 11271767 TI - Antibacterial effects of different food-related phosphates using Aeromonas hydrophila. AB - Aeromonas hydrophila is considered to be an emergent food-related bacterium. Phosphates are used as additives, mainly in meat products, to improve the quality of these foods. The antibacterial properties of phosphates are also well known. In this work, two A. hydrophila strains in early exponential phase were used: (A) A. hydrophila ATCC 7965 and (B) A. hydrophila derived from food, isolated in our laboratory. MIC and MBC studies were performed to assess the antibacterial effects of four phosphates assayed in brain heart infusion broth (BHI) and modified complete defined synthetic medium (mCDS) as compared to cooked ground meat medium (CM). The MBC values of the phosphates in CM were significantly higher than MIC values in BHI broth and mCDS medium (P < 0.05). In the two latter media, the growth of both A. hydrophila strains was totally inhibited by concentrations between 0.5 and 3.0%. Although all the assayed phosphates proved to have bactericidal effects on A. hydrophila, 0.5% sodium acid pyrophosphate (SAPP) exhibited greater effects in both strains and was selected for subsequent experiments. The bacteriolytic effect of SAPP was spectrophotometrically determined (260 nm of absorbance) by means of the leakage of intracellular nucleotides and microscopically confirmed by the presence of massive gelatinous aggregates. These were identified by enzymes (RNase, DNase, and proteinase) that hydrolyzed the nucleotides and proteins released during cellular lysis in the presence of SAPP. It was concluded that 0.5% SAPP can have bactericidal and bacteriolytic effects in early exponential phase A. hydrophila cells. PMID- 11271768 TI - Screening for clostridium botulinum type A, B, and E in cooked chilled foods containing vegetables and raw material using polymerase chain reaction and molecular probes. AB - A molecular method was used for the detection of Clostridium botulinum spores of type A, B, and E in commercial cooked and pasteurized vegetable purees and in the raw materials (vegetables and other ingredients). The method allowed the detection of less than 8 spores/g of product for C. botulinum type A, less than 1 spore/g for proteolytic type B, less than 21 spores/g for nonproteolytic type B, and less than 0.1 spore/g for type E. Thirty-seven samples of raw vegetables and ingredients were tested for the presence of C. botulinum type A, B, and E; 88 and 90 samples of vegetable purees were tested, respectively, for the presence of C. botulinum type A and B and for the presence of C. botulinum type E. All samples were negative, suggesting that the prevalence of C. botulinum in these vegetable purees and the raw ingredients is probably low. PMID- 11271769 TI - Rapid assessment of the bacteriological quality of raw milk using ATP bioluminescence. AB - Research was conducted to assess the practical use of an ATP bioluminescence assay to evaluate the bacteriological quality of raw milk. Filtration was used to precondition samples before ATP determination, which was measured in relative light units (RLUs). The Lumac ATP bioluminescence assay results were compared with standard plate counts (SPCs) of samples to estimate the microbial load for 246 raw milk samples that were split and either tested immediately or subjected to two preliminary incubation temperatures, 12.8 and 15.6 degrees C, for 18 h. Linear regression analysis procedures were used to analyze the data. Preincubation treatments were analyzed separately. For all treatments, linear regression coefficients were significantly different from zero (P < 0.01). The R2 values calculated using log10-transformed SPC and log10-transformed RLUs for samples tested immediately and samples preliminarily incubated at 12.8 and 15.6 degrees C were 0.58, 0.78, and 0.80, respectively. The R2 for all samples combined was 0.78. Differences in regressions among treatments were tested using a multiple slope and intercept model. Treatment intercepts and slopes were significantly different (P < 0.01). A linear regression equation was used to predict SPC from ATP values. Comparison of predicted values with actual SPCs indicated that ATP could be useful in predicting SPC in raw milk. PMID- 11271770 TI - The effect of nisin on the keeping quality of reduced heat-treated milks. AB - Milk was subjected to a combination process involving reduced heat treatment (RHT) of 117 degrees C for 2 s and nisin (75 and 150 IU ml(-1)). The microbial activity and other quality aspects were compared with a RHT control (without nisin) and with a ultrahigh temperature (UHT) milk processed at 142 degrees C for 2 s. Nisin was found to inhibit microbial growth for products stored without refrigeration, and RHT-nisin samples stored at 30 degrees C showed very low spoilage rates during 150 days, although not low enough to satisfy requirements for commercial sterility. RHT-nisin samples could be distinguished from and were preferred to the UHT control. Significant browning occurred during storage at 30 degrees C and above but was less in the RHT-nisin milk samples compared with the UHT milk. In RHT-nisin milk samples stored at 20 and 10 degrees C, no microbial activity could be detected in most samples after storage for 1 year. The effectiveness of this combination of RHT, nisin, and low storage temperatures against gram-positive spore-forming bacteria suggests potential for use of nisin in extended shelf life products. PMID- 11271771 TI - Biogenic amines in vacuum-packaged and carbon dioxide-controlled atmosphere packaged fresh pork stored at -1.50 degrees C. AB - Biogenic amines are formed in foods as a result of amino acid decarboxylation catalyzed by bacterial enzymes. When consumed in sufficient quantities, these compounds will cause headache, hypertension, fever, and heart failure. Technologies such as vacuum packaging and carbon dioxide-modified atmosphere packaging (CO2-MAP), when combined with low-temperature storage (-1.5 degrees C), allow fresh pork to have a storage life long enough for export to overseas markets. During low-temperature storage of pork in these packaging systems, the lactic acid bacteria (LAB), which possess the enzymes for biogenic amine formation, dominate the microflora. The objectives of this study were to determine the quantities of biogenic amines in packaged fresh pork, to monitor LAB growth, and to determine the storage life by sensory evaluation. Vacuum packaged and CO2-MAP pork were stored at -1.5+/-0.5 degrees C for 9 and 13 weeks, respectively. Phenylethylamine, putrescine, cadaverine, histamine, tyramine, spermidine, and spermine concentrations were determined weekly by high performance liquid chromatography and capillary gel electrophoresis. LAB and carnobacteria were enumerated weekly. Samples were evaluated for odor and appearance. The CO2-MAP was successful in delaying bacterial growth and the development of unacceptable off-odors compared with the vacuum packaging. The storage lives of the vacuum-packaged and CO2-MAP pork were 5 and 13 weeks, respectively. High-performance liquid chromatography was the superior method for biogenic amine quantification. Tyramine and phenylethylamine in pork of both packaging treatments approached levels considered to be potentially toxic. Given Canada's increasing role in the export of fresh meat to foreign markets, it is recommended that the formation of biogenic amines in vacuum-packaged and CO2-MAP pork be further investigated. PMID- 11271772 TI - Microbiological quality and shelf life modeling of ready-to-eat cicorino. AB - Growth of the microbial population resident in ready-to-use fresh cut cicorino, a variety of Chichorium intybus, was determined in the various preparation steps with the aim of defining the hazard critical control points. The investigation concerned cut cicorino from two producers. During the process a 1- to 1.5-fold increase of microbial counts was observed, and the retail product showed values of 10(5) to 10(6) CFU/g. The shelf life of the product was kinetically modeled in order to check the effects of storage temperature and assess the microbial indexes most relevant for hygiene and quality of the distributed product. A modified Gompertz function described the kinetics of microbial growth and allowed definition of a stability time and its dependence on temperature. Stability times were of 0.3, 3.7, and 4.7 days at storage temperatures of 20, 10, and 5 degrees C, respectively. Q10 (the fold decrease of stability time for an increase of 10 degrees C) was 3.85. The results from this study may be used to predict the effects of temperatures experienced in the distribution chain on bacterial levels in cicorino. PMID- 11271773 TI - Influence of turkey meat on residual nitrite in cured meat products. AB - A response surface experimental design was employed to estimate residual nitrite level at various initial nitrite concentrations, percent turkey meat in the formula, and heat quantity (F) values using a typical wiener as the test system. Pork and mechanically separated turkey were used as the meat ingredients. Residual nitrite and pH were measured at day 1, 7 days, 14 days, and 49 days after processing. Protein, fat, salt, moisture, and CIE (L*a*b*) color values were also determined. Results showed that the effect of turkey meat on residual nitrite level was significant (P < 0.01). An increased amount of turkey meat in the formula resulted in lower residual nitrite levels at a fixed pH. The residual nitrite level was initially proportional to initial nitrite concentration, but it became a nonsignificant factor during longer storage time. Differences in heat quantity had a significant effect (P < 0.05) on residual nitrite level initially. Greater heat quantity decreased residual nitrite level in finished cured meat products at a fixed pH. However, this effect became nonsignificant during longer storage. Reduction of residual nitrite in wieners because of turkey meat addition at a fixed pH was due to characteristics of the turkey tissue, but the mechanism of action remains unknown. It was also established that commercial wieners had a higher pH if poultry meat was included in the formulation. PMID- 11271774 TI - Mutagenicity and identification of mutagenic compounds of fumes obtained from heating peanut oil. AB - Since the fume of cooking oil has been reported to increase the risk of lung cancer, the objectives of this study were to evaluate the mutagenicity and to find the mutagens in the fumes of peanut oil heated to the smoke point. Peanut oil prepared from roasted peanut kernel showed a lower smoke point, less unsaturated fatty acids, more fume formation, and stronger mutagenicity than that from unroasted kernel. Further investigation of mutagenic compounds was performed by the Ames test and gas chromatography/mass spectrometry analysis. Among the 12 compounds identified from the neutral fraction of methanol extract, four compounds at a dose of 10 microg per plate were mutagenic to Salmonella Typhimurium TA98 and TA100 in the order of trans-trans-2,4-decadienal > trans trans-2,4-nonadienal > trans-2-decenal > trans-2-undecenal. Results report the enal compounds formed as the mutagens in the fumes of peanut oil and indicate that inhaling cooking fumes might cause carcinogenic risk. PMID- 11271775 TI - Roquefortine C occurrence in blue cheese. AB - Several strains of Penicillium are used for the production of mold-ripened cheeses, and some of them are able to produce mycotoxins. The aims of the research were the determination of roquefortine C and PR toxin in domestic and imported blue cheeses, the identification of the penicillia used as starter, and the investigation of their capacity for producing toxins in culture media. Roquefortine C was always found in the cheeses at levels ranging from 0.05 to 1.47 mg/kg, whereas the PR toxin was never found. The identification of the fungal strains present in the domestic cheeses included Penicillium glabrum, Penicillium roqueforti, and Penicillium cyclopium in the Gorgonzola "dolce" and Penicillium roqueforti in the Gorgonzola "naturale"; in one case, the presence of Penicillium crustosum was observed. The strains isolated from the foreign cheeses belonged to P. roqueforti. The strains were able to produce between 0.18 and 8.44 mg/liter of roquefortine in yeast extract sucrose medium and between 0.06 and 3.08 mg/liter and less than 0.05 mg/liter when inoculated in milk at 20 degrees C for 14 days and 4 degrees C for 24 days, respectively. Linear relations between production of roquefortine in culture media and cheeses did not emerge. PR toxin ranged from less than 0.05 to 60.30 mg/liter in yeast extract sucrose medium and was produced in milk at 20 degrees C from only one strain. The low levels and the relatively low toxicity of roquefortine make the consumption of blue cheese safe for the consumer. PMID- 11271776 TI - Difficulty in recovering inoculated Campylobacter jejuni from dry poultry associated samples. AB - We inoculated 5 cm2 of clean chick pads, 5 g of clean pine shavings, and fresh unsanitized broiler breeder eggshell halves with a cell suspension of Campylobacter jejuni in physiological saline. Inoculation levels were 10(2), 10(3), or 10(4) cells per sample. The samples were allowed to remain at room temperature for 15, 30, or 60 min before addition of enrichment broth. When chick pad samples were inoculated with 102 cells, by 15 min 40% of the samples had detectable levels of Campylobacter, and by 30 to 60 min Campylobacter could be detected in only 20% of the samples. With samples of pine shavings, only 25% of those inoculated with 103 cells were positive for Campylobacter after 15 min and only 5% were positive for Campylobacter after 30 min. When 104 cells were inoculated onto litter, Campylobacter was recovered from 20% of the samples at 15 min and 15% of the samples after 30 min. Eggshells were also found to be a harsh environment. When the inoculum was 102 at 15 min, 8 of 10 samples were positive for Campylobacter but at 60 min only 10% of the samples remained positive for Campylobacter. The current cultural methods may not be adequate for recovering low numbers of Campylobacter from dry samples. Campylobacter may be present but culturally undetectable in the commercial hatchery and hatchery environment. PMID- 11271777 TI - Effect of sodium chlorate on Salmonella typhimurium concentrations in the weaned pig gut. AB - Salmonella cause economic losses to the swine industry due to disease and compromised food safety. Since the gut is a major reservoir for Salmonella, strategies are sought to reduce their concentration in pigs immediately before processing. Respiratory nitrate reductase activity possessed by Salmonella also catalyzes the intracellular reduction of chlorate (an analog of nitrate) to chlorite, which is lethal to the microbe. Since most gastrointestinal anaerobes lack respiratory nitrate reductase, we conducted a study to determine if chlorate may selectively kill Salmonella within the pig gut. Weaned pigs orally infected with 8 x 10(7) CFU of a novobiocin- and nalidixic acid-resistant strain of Salmonella Typhimurium were treated 8 and 16 h later via oral gavage (10 ml) with 0 or 100 mM sodium chlorate. Pigs were euthanized at 8-h intervals after receiving the last treatment. Samples collected by necropsy were cultured qualitatively and quantitatively for Salmonella and for most probable numbers of total culturable anaerobes. A significant (P < 0.05) chlorate treatment effect was observed on cecal concentrations of Salmonella, with the largest reductions occurring 16 h after receiving the last chlorate treatment. An observed treatment by time after treatment interaction suggests the chlorate effect was concentration dependent. Chlorate treatment may provide a means to reduce foodborne pathogens immediately before harvest. PMID- 11271778 TI - Study of inactivation of Lactobacillus plantarum in orange-carrot juice by means of pulsed electric fields: comparison of inactivation kinetics models. AB - The inactivation kinetics of Lactobacillus plantarum was studied in orange-carrot juice using high intensity pulsed electric fields. The results indicated that under the treatment conditions applied, 28.6, 32.0, and 35.8 kV/cm and treatment times ranging from 10.2 to 46.3 micros, the inactivation of L. plantarum obtained was up to 2.5 decimal reductions. Experimental and literature data were fitted to Bigelow, Hulsheger et al. and Peleg models and to Weibull frequency distribution function. Weibull was the one that best interpreted the data with accuracy factor values closer to 1. PMID- 11271779 TI - Description of a simple detection assay for in situ production of bacteriocin on meat. AB - Using a modification of the agar diffusion assay, in situ bacteriocin production on meat was analyzed using cooked meat medium (CMM) and sterile pork tissue (lean and fat) with Carnobacterium piscicola LV17 as the producer and Carnobacterium divergens LV13 as the indicator strains. Contrary to what is observed in APT broth, bacteriocin production by C. piscicola LV17 occurred with growth at low inoculum levels (< or =10(4) CFU/cm2 or g of meat) on disks (10 cm2) of pork fat tissue (pH 6.58) and on CMM particles (pH 7.0) but not on disks of lean tissue (pH 5.61). The assays described in this study do not required sophisticated equipment and would be useful to study bacteriocin production on meat products stored under various conditions. PMID- 11271780 TI - Aflatoxin B1 degradation by flavobacterium aurantiacum in the presence of reducing conditions and seryl and sulfhydryl group inhibitors. AB - This study was undertaken to determine the effects of reducing conditions (L cysteine) and seryl (phenylmethylsulfonyl fluoride) and sulfhydryl (divalent cadmium) group inhibitors on aflatoxin B1 (AFB1) degradation by Flavobacterium aurantiacum. High-performance liquid chromatography was used to determine AFB1 concentrations in 72-h cultures of F. aurantiacum. The addition of 0.1, 1, or 10 mM L-cysteine did not have any significant effect on AFB1 degradation by these cultures after incubation for 4, 24, or 48 h (P > 0.05). The addition of 0.1 mM phenylmethylsulfonyl fluoride did not significantly decrease AFB1 degradation (P > 0.05), but 1 mM phenylmethylsulfonyl fluoride significantly decreased AFB1 degradation after 4, 24, and 48 h of incubation (P < or = 0.05). No significant difference in AFB1 degradation was obtained with 0.1 mM Cd2+ after 4, 24, or 48 h of incubation (P > 0.05). The addition of 1 and 10 mM Cd2+ significantly decreased AFB1 degradation compared with the cells containing AFB1 alone after 4 and 24 h (P < or = 0.05). The addition of chelators, 1 mM EDTA and 1 mM o phenanthroline, did not result in removal of inhibition of AFB1 degradation observed with 1 and 10 mM Cd2+. Higher concentration of chelators (>1 mM) are necessary to overcome the inhibitory effect. Further work on the cellular fractions and/or crude enzyme preparations is necessary to determine if indeed sulfhydryl and seryl groups of the enzymes are involved in AFB1 degradation (by maintaining either the structure or function of the enzyme). PMID- 11271781 TI - Effects of gamma radiation on the allergenic and antigenic properties of milk proteins. AB - This study was carried out to evaluate the application of food irradiation technology as a method for reducing milk allergies. Bovine alpha-casein (ACA) and beta-lactoglobulin (BLG) were used as milk proteins. Using milk-hypersensitive patients' immunoglobulin E (IgE) and rabbit IgGs individually produced to ACA and BLG, the changes of allergenicity and antigenicity of irradiated proteins were observed by competitive indirect enzyme-linked immunosorbent assay. Allergenicity and antigenicity of the irradiated proteins were changed with different slopes of the inhibition curves. The disappearance of the band on sodium dodecyl sulfate polyacrylamide gel electrophoresis and increase of the turbidity showed that solubility of the proteins decreased by radiation, and this decrease might be caused by agglomeration of the proteins. These results indicated that epitopes on milk allergens were structurally altered by gamma irradiation. PMID- 11271782 TI - Leptin: of mice and men? AB - A major advance in the understanding of the control of appetite, food intake, and energy expenditure came with the discovery of leptin. Leptin concentrations correlate with adipose tissue mass, and leptin acts via the central nervous system (CNS) to reduce food intake and increase energy expenditure. A variety of different neurotransmitters have been implicated in mediating the CNS effects of leptin. In humans, leptin deficiency is unlikely to be a major cause of obesity. Most humans are not leptin deficient, but have a leptin concentration raised in proportion to their fat mass. A recent clinical trial looking at the use of recombinant leptin in treating human obesity has resulted in only variable amounts of weight loss. The role of leptin extends beyond the control of food intake and energy expenditure. Leptin reverses many of the physiological responses to starvation. It is suggested that the main role of leptin might be in response to food deprivation and not in obesity. PMID- 11271783 TI - Preoperative assessment of prognostic factors in breast cancer. AB - The adoption of preoperative diagnostic strategies involving fine needle aspiration cytology (FNAC) or core biopsy is well established, allowing the planning of operating lists and bed occupancy and patient involvement in therapeutic management. In addition to diagnosis, however, pathologists are increasingly being asked to provide pathological prognostic information from preoperative samples. This leader describes techniques for predicting prognosis and response to treatment on these specimens and some of the problems inherent in the determination of prognosis on small samples. For example, although histological grade can be assessed relatively reliably on either core or FNAC samples, the evaluation of tumour type (which includes an overall assessment of the architecture of a given tumour) may be less reliable on small preoperative samples. Other well recognised histological prognostic factors, such as vascular channel invasion or tumour size, cannot be determined accurately on small preoperative samples. For those patients who might benefit from neoadjuvant treatment, predicting the response to such treatments--for example, by the assessment of oestrogen receptor status--can readily be performed on either core biopsy or FNAC. In the future, other molecular markers such as C-erbB-2 might also prove beneficial in predicting response to newly developed treatments. PMID- 11271784 TI - An evaluation of three commercial kits for use as screening methods for the detection of leptospiral antibodies in the UK. AB - AIMS: To compare three commercial screening tests--the PanBio leptospiral IgM enzyme linked immunosorbent assay (ELISA), the Biolisa leptospiral IgM ELISA, and the indirect haemagglutination assay (IHA)--with the microscopic agglutination test (MAT) and two "in house" ELISAs--urease and horseradish peroxidase (HRP)- for the detection of leptospiral antibodies in a local UK and Eire population. METHOD: Two hundred sera submitted for a differential diagnosis of leptospirosis were tested by all methods. A further 142 sera from patients with antibodies to toxoplasma, Epstein-Barr virus (EBV), hepatitis A virus, rheumatoid factor, Borrelia burgdorferi, Mycoplasma pneumoniae, syphilis, cytomegalovirus, and Q fever were tested for crossreactivity. RESULTS: Compared with the MAT, sensitivity and specificity were found to be: PanBio, 90%/94%; Biolisa with sorbent, 100%/85%; and IHA, 54%/95%. Seven of 200 trial sera gave false negative results with PanBio; 14 of 200 trial sera gave false positive results with Biolisa with sorbent, as did a further 25 of the 142 sera tested for potential crossreactivity. Two of 142 sera gave crossreactions with PanBio and IHA (one each). CONCLUSIONS: The degree of false positivity seen with the Biolisa suggests that the recommended positive value of > or = 26 Eu/ml should be reassessed using pools of sera from local populations. When the cut off value was reassessed, using a value of > or = 40 Eu/ml, a sensitivity and specificity of 96% and 94%, respectively, was achieved. Even the modified Biolisa appears to be over sensitive and to show a high degree of non-specificity. The IHA, although specific (95%), lacked sensitivity in this study. The PanBio appeared to be the most suitable as a screening test for leptospiral IgM in the UK, although it would be advisable for all positive test results to be confirmed by a different enzyme immunoassay and the MAT. PMID- 11271785 TI - Topoisomerase II alpha and II beta expression in childhood acute lymphoblastic leukaemia: relation to prognostic factors and clinical outcome. AB - BACKGROUND/AIMS: Many regimens used in the treatment of childhood acute lymphoblastic leukaemia (ALL) include Daunorubicin or Etoposide, which act as topoisomerase poisons. It has been suggested that there may be a relation between topoisomerase expression and response to topoisomerase poisons, based mainly on results from in vitro studies. Therefore, the aim of this study was to investigate this relation in a clinical setting and determine whether topoisomerase II alpha and II beta might be of predictive value in ALL. METHODS: Cellular expression of topoisomerases II alpha and II beta was assessed in 177 cases of ALL by immunohistochemistry using monoclonal antibodies to the two enzymes. The percentages of cell nuclei showing positive staining for topoisomerase II alpha and II beta expression were assessed. RESULTS: Taking the series as a whole, a clear separation of survival curves was seen with the established prognostic markers white blood cell (WBC) count, CD10 status, and sex. However, topoisomerase II alpha and II beta expression showed no relation to survival. No association was found between the topoisomerases and the prognostic markers CD10 and WBC count; however, topoisomerase II alpha expression was found to be related to sex, with expression being lower in girls (p = 0.002). CONCLUSIONS: These results suggest that the response to topoisomerase poisons cannot be predicted by the assessment of topoisomerase II alpha and II beta expression as defined by immunohistochemistry. PMID- 11271787 TI - Scientific dishonestry: European reflections. AB - Scientific dishonesty has attracted increased attention around the world during the past three to four decades. Europe became aware of the problem later than the USA, but has within the past 10 years created national control systems for all biomedical projects, not only those supported by public money. The prevalence of the problem can only be calculated indirectly by referring to population figures as denominators. Measured this way, figures from Denmark as a whole show: 1-2 cases referred/million inhabitants/year, 1 case treated/million inhabitants/year, 1 case of scientific dishonesty/million inhabitants/5 years. For Finland, 1-2 cases were referred/million inhabitants/1-2 years; for Norway, similar figures of 1/4 million inhabitants/year were calculated. Figures from the Danish national independent control body 1993-7 show the distribution of the types of cases that were charged, with numbers of confirmed cases in parentheses: fabrication, 2 (1); plagiarism, 3 (0); theft, 2 (0); ghost authorship, 2 (1); false methodological description, 3 (1); twisted statistics, 2 (0); suppression of existing data, 4 (0); unwarranted use of data, 4 (0); and authorship problems, 8 (1). This survey emphasises the need for national guidelines, an independent national control body, and initiatives for strong preventive actions. PMID- 11271786 TI - Expression of acidic fibroblast growth factor (aFGF) and fibroblast growth factor receptor 4 (FGFR4) in breast fibroadenomas. AB - BACKGROUND/AIM: Fibroadenomas are benign tumours composed of both glandular and fibrous tissue. The mechanisms regulating the growth of these tumours and the relation between the stromal and epithelial cells are poorly understood. Acidic fibroblast growth factor (aFGF) is a well known fibroblast activator, which acts through four specific cell surface receptors, among which, fibroblast growth factor receptor 4 (FGFR4) is highly specific. The aim of this study was to evaluate the distribution of aFGF and FGFR4 in specific cell types of fibroadenomas to understand their possible role in the growth of these breast lesions. METHODS: Formalin fixed and paraffin wax embedded tissues from 15 fibroadenomas and peritumoral normal breasts were investigated for the expression of aFGF and FGFR4 using immunohistochemistry. The presence of aFGF mRNA was also investigated using in situ hybridisation. RESULTS: Immunoreactivity for aFGF and FGFR4 was seen in epithelial cells, but it was lacking in myoepithelial cells of both normal tissues and fibroadenomas. Strong FGFR4 immunoreactivity was found in stromal fibroblasts, which were also weakly positive for aFGF. aFGF mRNA was detected in epithelial cells and in some stromal fibroblasts. CONCLUSIONS: These results suggest a paracrine/autocrine modulation of epithelial and stromal cells of fibroadenomas through an aFGF-FGFR4 interaction. This interaction might regulate various cell functions and the growth of fibroadenomas. PMID- 11271789 TI - Association of human beta-herpesviruses with the development of cervical cancer: bystanders or cofactors. AB - BACKGROUND/AIM: Human papillomaviruses (HPVs) are important, but not sufficient, for the development of cervical cancer. All three human beta-herpesviruses- cytomegalovirus (CMV) and human herpesviruses (HHV) types 6 and 7--have been detected in the cervix. In addition, CMV and HHV-6 can interact with HPVs in vivo. This study examined the possible role of beta-herpesviruses in cervical cancer development. METHODS: HPV, CMV, HHV-6, and HHV-7 were detected by the polymerase chain reaction using cervical scrapes taken at colposcopy from 388 women. HPV types were identified using restriction fragment length polymorphisms. Colposcopy guided biopsies were taken from abnormal areas, and the histological findings were regarded as the final diagnoses. The associations between herpesvirus infection and the degree of cervical lesion were analysed with respect to HPV status. RESULTS: Of the 388 women, 51.8% had a normal cervix, 14.4% had cervical intraepithelial neoplasia grade 1 (CIN1), 8.2% had CIN2, 19.3% had CIN3, and 6.2% had invasive carcinoma. Overall, the positive rates for high, intermediate, and low risk HPVs were 18.8%, 21.4%, and 5.2%, respectively. Fifteen patients harboured HPVs for which the genotype could not be identified. Positive rates for CMV, HHV-6, and HHV-7 were 9.5%, 3.6%, and 3.4%, respectively. HPV positive patients carried a higher risk for high grade lesions (CIN2/3 or carcinoma) (odds ratio (OR), 5.24; 95% confidence interval (CI), 3.19 to 8.62; chi 2 = 51.79; p < 0.001), whereas those positive for CMV, HHV-6, or HHV-7 did not. Thirteen of 131 patients with high grade lesions had HPV/herpesvirus coinfections, but no association with the cervical lesion was noted. Furthermore, positive rates for herpesviruses among HPV negative, high/intermediate risk HPV negative, and high risk HPV negative subgroups were similarly low and without a significant association. CONCLUSIONS: The ubiquitous nature of herpesviruses may pose difficulty in elucidating their pathogenic role. These results indicate that CMV, HHV-6, and HHV-7 are bystanders rather than cofactors in the oncogenesis of cervical cancer. PMID- 11271788 TI - Nuclear beta catenin expression is related to unfavourable outcome in oropharyngeal and hypopharyngeal squamous cell carcinoma. AB - AIMS: To investigate the expression of alpha, beta, and gamma catenins in oropharyngeal and hypopharyngeal squamous cell carcinoma and their relations to each other, as well as to clinical data, tumour differentiation, and prognosis. METHODS: Primary tumours for analysis were obtained from 138 patients diagnosed with squamous cell carcinoma of the oropharynx or hypopharynx between 1975 and 1998 in eastern Finland. Immunohistochemistry was used to evaluate the expression of alpha, beta, and gamma catenins. The expression patterns of all catenins were related to clinical data and survival. RESULTS: The expression patterns of all three catenins were significantly interrelated. Reduced gamma catenin expression was significantly associated with poor histological differentiation. No association was found between alpha or beta catenin expression and clinicopathological characteristics. In univariate analysis, patients whose tumours had nuclear beta catenin expression had shorter overall survival than patients with no nuclear expression. In Cox multivariate analysis, nuclear beta catenin expression, tumour status (T class), and Karnofsky performance index were independent prognostic factors of overall survival. CONCLUSIONS: Reduced expression of gamma catenin is associated with dedifferentiation in primary squamous cell carcinoma of the oropharynx and hypopharynx. The fact that nuclear beta catenin expression independently predicts short overall survival suggests that it might be a valuable prognostic marker in pharyngeal squamous cell carcinoma. PMID- 11271790 TI - Why oral calcium supplements may reduce renal stone disease: report of a clinical pilot study. AB - AIMS: To investigate whether increasing the daily baseline of gut calcium can cause a gradual downregulation of the active intestinal transport of calcium via reduced parathyroid hormone (PTH) mediated activation of vitamin D, and to discuss why such a mechanism might prevent calcium oxalate rich stones. To demonstrate the importance of seasonal effects upon the evaluation of such data. METHODS: Within an intensive 24 hour urine collection regimen, daily calcium supplementation (500 mg) was given to five stone formers for a 10 week period during a six month crossover study. In a further population of patients on follow up for previous renal stone disease, observations were made on 1066 24 hour urine samples collected over five years in respect of seasonal effects relevant to the interpretation of the study. RESULTS: In the group of patients on calcium supplements the following results were found. During calcium supplementation, the proportion of urine calcium to oxalate was higher (increased calcium to oxalate molar ratio), the 24 hour urine product of calcium and oxalate did not rise, and urine oxalate was lower during the first six weeks of supplementation. Twenty four hour urine calcium was 10.2% higher than baseline in the final four weeks of the 10 weeks of supplementation. Twenty four hour urine phosphate was 11.4% lower during the first six weeks of supplementation, but then rose while the patients were still on supplementation; renal tubular reabsorption of phosphate (TmP/GFR) mirrored the urine phosphate changes inversely. PTH was higher after stopping supplementation, but 1,25-(OH)2-cholecalciferol changes were not detected. In the 1066 urine samples collected over five years the following results were found. Calcium and oxalate excretion correlated positively and not inversely. Urine calcium and phosphate excretion were 5.5% and 2.5% higher, respectively, in "light" months of the year compared with "dark" months. A post summer decline in both urine calcium and urine phosphate was relevant to the interpretation of the study. CONCLUSIONS: Regular calcium supplementation does not raise the product of calcium and oxalate in urine and the proportion of oxalate to calcium is reduced. The underlying mechanisms of the changes seen in phosphate, calcium, and PTH and the observations on 1,25-(OH)2-cholecalciferol are not clear. Observed changes in phosphate could possibly be part of a calcium regulating feedback loop operating over a period of weeks. In evaluating these mechanisms background seasonal effects are important. It is possible that "programming" of the gut mucosa in terms of calcium transport is a major determinant of the relation between calcium and oxalate concentrations in urine and their relative abundance. Increased oral calcium, in association with a reduction of the relative proportion absorbed, may be pertinent to the prevention of calcium oxalate rich stones. PMID- 11271791 TI - Rapid quantitative assessment of gastric corpus atrophy in tissue sections. AB - BACKGROUND/AIMS: Grading of Helicobacter pylori induced atrophic gastritis using the updated Sydney system is severely limited by high interobserver variability. The aim of this study was to set up a quantitative test of gastric corpus mucosal atrophy in tissue sections and test its reproducibility and correlation with the Sydney scores of atrophy. METHOD: Mucosal atrophy was assessed in 124 haematoxylin and eosin stained corpus biopsy specimens by two experienced gastrointestinal pathologists (EB, JL) according to the updated Sydney system as none (n = 33), mild (n = 33), moderate (n = 33), or pronounced (n = 25). In each specimen, the proportions of glands, stroma, infiltrate, and intestinal metaplasia in the glandular zone were measured as volume percentages using a point counting method. The optimal point sample size, intra-observer and interobserver reproducibility, discriminative power for degrees of atrophy, and correlations with H pylori status were evaluated. RESULTS: Counting 400 points in 200 fields of vision provided the smallest sample size that still had excellent intra-observer and interobserver reproducibility (r > or = 0.96). Overall, the volume percentage of glands (VPGL), infiltrate (VPI), and stroma (VPS) correlated well with the Sydney scores for atrophy (p < or = 0.003). However, no differences were found between non-atrophic mucosa and mild atrophy. No correlation was found between age and either the Sydney grade of atrophy or the VPGL or VPS. In non atrophic mucosa and mild atrophy, H pylori positive cases showed a significantly higher VPI than did H pylori negative cases. A lower VPGL was seen in H pylori positive cases than in H pylori negative cases in the mild atrophy group. VPS did not correlate with H pylori status within each grade of atrophy. CONCLUSION: Point counting is a powerful and reproducible tool for the quantitative analysis of mucosal corpus atrophy in tissue sections. These data favour the combination of "none" and "mild" atrophy into one category, resulting in a three class grading system for corpus atrophy, when using the updated Sydney system. PMID- 11271793 TI - Mixed apocrine/endocrine ductal carcinoma in situ of the breast coexistent with lobular carcinoma in situ. AB - An unusual mixed form of ductal carcinoma in situ (DCIS) of the breast is described, which exhibits a biphenotypic morphology encompassing a range of differential diagnostic DCIS subtypes. In addition, immunophenotypic and ultrastructural studies demonstrate neuroendocrine and apocrine differentiation, raising questions regarding appropriate classification and biological behaviour. In two cases, coexistence of this mixed form of DCIS with lobular carcinoma in situ (LCIS) in the same duct lobular units is an additional unusual feature that might, at least in some cases, indicate a closer relation between them. PMID- 11271795 TI - Clear cell adenocarcinoma of the colon arising in endometriosis: a rare variant of primary colonic adenocarcinoma. PMID- 11271794 TI - Primary peripheral T cell lymphoma of the endometrium. AB - A case of a primary peripheral T cell lymphoma arising in the endometrium is presented. Primary lymphomas of the female genital tract are rare, with endometrial lymphomas and those of T cell type being rarer still. Extensive investigations revealed no other sites of disease and the patient was treated by hysterectomy and chemotherapy. She remains well 33 months later. We believe that this case is exceptionally unusual. PMID- 11271792 TI - Neutrophil disorders and their management. AB - Neutrophil disorders are an uncommon yet important cause of morbidity and mortality in infants and children. This article is an overview of these conditions, with emphasis on clinical recognition, rational investigation, and treatment. A comprehensive list of references is provided for further reading. PMID- 11271796 TI - Retroperitoneal extraskeletal osteosarcoma. PMID- 11271798 TI - Dietary dangers: ingestion of a bread bag clip. PMID- 11271797 TI - Immunohistochemical demonstration of oestrogen and progesterone receptors. PMID- 11271801 TI - Purification and characterization of a thermostable alpha-amylase from Bacillus stearothermophilus. AB - A soil isolate of Bacillus stearothermophilus was found to synthesize thermostable alpha-amylase. The enzyme was purified to homogeneity by ammonium sulfate fractionation and IECC on DEAE-cellulose column. The purified enzyme was considered to be a monomeric protein with a molar mass of 64 kDa, as determined by SDS-PAGE. The enzyme showed a wide range of pH tolerance and maximum activity at pH 7.0. The temperature tolerance was up to 100 degrees C with more than 90% catalytic activity; the maximum activity was observed at 50 degrees C. Divalent metal ions exhibited inhibitory effect on the enzyme activity. However, proteinase inhibitor did not react positively. PMID- 11271800 TI - Physiological activity of some organophosphorus compounds and their effects on the physico-chemical properties of model membranes. AB - The effect of the newly synthesized phosphonic compound dibutyl 2-octylamino-2 propanephosphonate (DBOP) on the growth of the aquatic plant Spirodela oligorrhiza and stability of red blood cells (RBC) and planar lipid membranes (BLM) was studied to determine its physiological activity and, if possible, correlate this activity to compound-induced changes in the mechanical properties of the model membranes. The measure of the phytotoxicity was the DBOP concentration causing 50% plant growth retardation, while measures of stability of model membranes were 100% hemolysis of RBC and a critical concentration of DBOP causing BLM destruction in no more that 3 min. These data were compared with those for dibutyl 1-butylamino-1-cyclohexanephosphonate (DBBC) and diethyl 9 butylamino-9-fluorenephosphonate (DEBF) known for their physiological activities. Both DBBC and DEBF influenced Spirodela growth significantly less than DBOP Destabilization of the model membrane caused by DBBC and DBOP was similar whereas DEBF exerted a weak influence on RBC and BLM stability. The results indicate that the physiological activities of DBOP and DEBF are not limited to the lipid phase of biological membranes and may involve also disturbance of metabolic processes. PMID- 11271799 TI - From no-confidence to nitric oxide acknowledgement: a story of bacterial nitric oxide reductase. AB - The review briefly summarizes current knowledge of the bacterial nitric-oxide reductase (NOR). This membrane enzyme consists of two subunits, the smaller one contains haem C and the larger one two haems B and nonhaem iron. The protein sequence and structure of metal centres demonstrate the relationship of NOR to the family of terminal oxidases. The binuclear Fe-Fe reaction centre, consisting of antiferromagnetically coupled haem B and nonhaem iron, is analogous to Fe-Cu centre of terminal oxidases. The data on the structure and function of NOR and terminal oxidases suggest that all these enzymes are closely evolutionally related. The catalytic properties are determined most of all by the relatively high toxicity of nitric oxide as a substrate and the resulting strong need to maintain its concentration at nanomolar levels. A kinetic model of the action of the enzyme comprises substrate inhibition. NOR does not conserve the free energy of nitric oxide reduction because it does not work as a proton pump and, moreover, the protons coming into the reaction are taken from periplasm, i.e. they do not cross the membrane. PMID- 11271803 TI - Copper accumulation by Aspergillus awamori. AB - Aspergillus awamori accumulated Cu2+ from aqueous solutions. The level of copper uptake was dependent on the ambient metal concentration. The process consisted of two phases: a fast initial phase and a slower secondary phase. Chelation of these ions occurs by chemical, equilibrated and saturable mechanism, following the mathematical models of Langmuir and Freundlich, with better performance on the Langmuir model. Data transformation allowed us to calculate the kinetic constants of the sorption reaction. PMID- 11271802 TI - Detoxication of the herbicide diuron by Pseudomonas sp. AB - A strain of bacteria able to detoxicate the herbicide diuron in pure culture was isolated from sites contaminated with different urea herbicides. Diuron was used as a sole source of carbon and energy by this isolate which is a Gram-negative, aerobic, rod-shaped bacterium with a single polar flagellum, and grows at 40 degrees C. The strain has been identified as Pseudomonas sp. PMID- 11271804 TI - Two forms of yeast plasma membrane H(+)-ATPase: comparison of yield and effects of inhibitors. AB - Classical isolation procedure for plasma membrane H(+)-ATPase of Saccharomyces cerevisiae based on fractional centrifugation yielded always a roughly two-fold greater amount of membranes when starting from glucitol-preincubated than from glucose-preincubated yeast. This difference persisted all the way to the purified plasma membranes and to the purified H(+)-ATPase. The ATP-hydrolyzing activity by plasma membranes was roughly twice greater in glucose-preincubated cells than in the D-glucitol-preincubated ones while the purified enzyme was 7 times more active after glucose than after glucitol. Effects of diethylstilbestrol, suloctidil, erythrosin B, vanadate and dicarbanonaboranuide were very similar on plasma membrane-localized and purified ATPases of both forms, suggesting that both preparations contain the two ATPase forms, the glucose-preincubated one being richer in the activated form while the glucitol-preincubated one contains less of it. PMID- 11271805 TI - Use of synchronously excited fluorescence to assess the accumulation of membrane potential probes in yeast cells. AB - Evaluation of emission spectra of fluorescent probes used for the monitoring of membrane potential in microbial cells can be greatly facilitated by using synchronously excited spectroscopy (SES). This method permits the suppression of undesirable spectrum components (contributions due to scattered light or cell autofluorescence) and leads to considerable increase in monitored emission intensity and to narrowing of spectral peaks. It allows an efficient fractional decomposition of the probe fluorescence spectra into their free and bound dye fluorescence components. The usefulness of the method was tested by monitoring the accumulation of the fluorescent membrane potential probe diS-C3(3) in yeast cells, which serves as a qualitative measure of the membrane potential. PMID- 11271806 TI - Dimorphism in Benjaminiella poitrasii: involvement of intracellular endochitinase and N-acetylglucosaminidase activities in the yeast-mycelium transition. AB - The chitinase and N-acetylglucosaminidase activities in cell-wall-bound and free fractions in the dimorphic fungus Benjaminiella poitrasii were studied as a function of morphological (unicellular yeast-mycelium) transition. The specific activities of chitinases of cell-wall-free, particularly in the membrane fraction, were significantly different in the yeast and mycelial forms. During the yeast-mycelium transition, the N-acetylglucosaminidase activity isolated in a membrane preparation increased steadily. The activity of the yeast cells (0.83 +/ 0.17 nkat/mg protein) increased 17-fold to 14.2 +/- 1.7 nkat/mg protein in 1-d old mycelial cells. The endochitinase activity increased 12-fold between 6 and 12 h and thereafter practically remained unchanged up to 24 h. A reverse trend in the chitinolytic activities was observed during the mycelium-yeast transition. Isoelectrofocussing (pH range 3.5-10) of mixed membrane fraction free of particulate fraction of parent and morphological (Y-5, yeast-form) mutant cells separated endochitinase and N-acetylglucosaminidase activity into two pH ranges, viz. 4.3-5.7 and 6.1-7.7, respectively. The predominant N-acetylglucosaminidase activity observed at pH 6.9 and 7.1 for the parent strain membrane fraction was undetected in the mutant preparation. The results suggested that the membrane bound (either tightly or loosely) chitinolytic enzymes, particularly, N acetylglucosaminidase, significantly contributed to the morphological changes in B. poitrasii. PMID- 11271808 TI - Progesterone side-chain degradation by some species of Aspergillus flavus group. AB - Seventy isolates belonging to 6 species and one variety of A. flavus group were shown to degrade the progesterone side-chain to yield delta 4-androstene-3,17 dione and testosterone. The isolates of five species (A. flavo-furcatis, A. flavus, A. oryzae, A. parasiticus and A. tamarii) possessed enzyme systems catalyzing the opening of ring D and formed testololactone as final steroid metabolite in addition to their ability to produce the above mentioned two products. 11 beta-Hydroxy-delta 4-androstene-3,17-dione was formed by only A. flavus and A. tamarii while 11 beta-hydroxytestosterone was produced by A. flavo furcatis, A. parasiticus and A. subolivaceus. The chromatographic resolution of the mixture products obtained (when the selective isolate of each species reacted with 1 g of progesterone) revealed that 60-75% of progesterone was converted into delta 4-androstene-3,17-dione (8-30%), testosterone (7-33%), testololactone (14 37%) and other products (3-40%). The most bioconversion activity was exhibited by A. oryzae, followed by A. parasiticus. The highest values of delta 4-androstene 3,17-dione (30% of added progesterone) and testosterone (33%) were formed by A. flavus var. columnaris while those of testololactone (37%) were produced by A. oryzae. A systematic variation could be observed between the different tested species of A. flavus group with respect to the transformation reactions of progesterone. Comparative biotransformation results showed that essential differences exist between the tested species in this group; this biochemical differentiation may supplement the morphological and other physiological criteria used in the identification of the different species in the A. flavus group. PMID- 11271807 TI - Immuno-electron localization of DNA in chondriolites of Saccharomyces cerevisiae mitochondria. AB - Under electron microscope, the matrix of sectioned mitochondria exhibits ribosomes and an oval, electron-transparent zone which is devoid of ribosomes and is named chondriolite. Fine fibers or clumps of an electron-dense material appeared in this zone after several fixation and contrasting steps and were identified with mitochondrial DNA by cytologists. To verify this assumption, we labeled DNA by a monoclonal antibody and a secondary antibody coupled to immunogold. The label was observed in the nucleus and in the chondriolite zone of sectioned mitochondria. Because the ultrastructure of chondriolites resembles that of nucleoids of prokaryotes, we suggest the term mitochondrial nucleoid for the zone of mitochondrial matrix devoid of ribosomes and containing DNA. PMID- 11271809 TI - Ligninolytic enzyme complex of Armillaria spp. AB - Ten strains belonging to five species of European Armillaria (Fr.:Fr.) Staude were examined for their ability to produce laccase, lignin peroxidase, manganese dependent peroxidase and manganese-independent peroxidase. No lignin peroxidase activity was observed in any of the strains. Manganese-dependent peroxidase production by all tested strains was low. Difference in the ratio of laccase to manganese-independent peroxidase in strains of A. gallica in comparison to all other species was detected. PMID- 11271810 TI - Protein overexport in a Saccharomyces cerevisiae mutant is not due to facilitated release of cell-surface proteins. AB - Saccharomyces cerevisiae strain MW11 is a temperature-sensitive mutant which exports twenty times more proteins at 37 degrees C than parental or wild-type strains do. To understand the mechanism underlying the protein overexport in the mutant the possibility of an altered cell-wall structure leading to facilitated release of cell-surface proteins was studied. Data on calcofluor white and zymolyase sensitivities, resistance to killer 1 toxin and determination of exported acid phosphatase and invertase did not provide evidence for alterations in the cell-wall structure that could explain the protein overexport phenotype. The results were obtained in experiments when transcription of mutated gene was discontinued which permits the full expression of the protein overexport phenotype. PMID- 11271811 TI - Isolation and identification of xylitol-producing yeasts from agricultural residues. AB - Selected yeast strains isolated from corn silage and viticulture residues were screened for their capacities to convert D-xylose into xylitol A conventional TLC was adapted for easy determination of xylose and xylitol in the culture supernatant solutions. This technique is suitable for the first steps of a screening program to select xylitol-producing yeasts from natural environments. Candida tropicalis ASM III (NRRL Y-27290), isolated from corn silage, appears to be a promising strain for xylitol production with a high yield (0.88 g xylitol per g of xylose consumed). PMID- 11271813 TI - Verification of hypocholesterolemic effect of fermented milk on human subjects with different cholesterol levels. AB - The possible hypocholesterolemic effect of acidophilus milk was evaluated on 27 human subjects having different levels of serum cholesterol, i.e. < 2.0 (group C1), 2.0-2.2 (C2), 2.2-2.5 (C3) and > 2.5 g/L (C4). The acidophilus milk was prepared by fermentation of low-fat milk with Lactobacillus acidophilus and was fed to each volunteer at the rate of 200 mL/d for 20 d. Blood samples from the volunteers were collected and analyzed for lipid profile twice prior to, during and after feeding, keeping a gap of 10 d between two collections. A significant decrease (p < 0.05) in average total cholesterol was found in the C2 and C3 groups, amounting to 21 and 12%, respectively. The average LDL cholesterol decreased in C2, C3 and C4 groups by 0.54, 0.26 and 0.46 g/L, respectively. In the C2 group, the LDL/HDL and total/HDL ratio was also reduced by 1.4 and 1.3, respectively. However, in the C1 group, the average total and LDL cholesterol level did not show any significant change but serum triacylglycerols and VLDL cholesterol showed a significant (p < 0.05) increase of 0.53 and 0.11 g/L, respectively. Regression analysis of the data revealed a square trend in most of the parameters over time period. Overall, the feeding had the best effect in the subjects with lipidemic status of borderline cholesterol level (2.0-2.2 g/L) group. PMID- 11271812 TI - Ultrastructure of two oil-degrading bacteria isolated from the tropical soil environment. AB - Two oil-degrading bacteria identified as Pseudomonas aeruginosa and Micrococcus luteus were isolated from crude-oil-polluted soils in Nigeria. The organisms were grown on n-hexadecane and sodium succinate and then examined for the presence of hydrocarbon inclusions. Inclusion bodies were found in n-hexadecane-grown cells and were absent in succinate-grown cells. Formation of hydrocarbon inclusion bodies appears to be a general phenomenon among hydrocarbon utilizers. PMID- 11271814 TI - Methanogenesis in rumen ciliate cultures of Entodinium caudatum and Epidinium ecaudatum after long-term cultivation in a chemically defined medium. AB - The methanogenic activity in the presence of Entodinium caudatum and Epidinium ecaudatum was well preserved after long-term cultivation. Microscopic observation revealed that methane production in the presence of E. caudatum was probably caused by their intracellular methanogenic activity, while methane production in the presence of E. ecaudatum f caudatum et ecaudatum could be attributed to both the methanogenic bacterial fraction of their external surface and their intracellular activity. Methane production per protozoan cell of E. caudatum and E. ecaudatum was 2.1 nmol per cell per d and 6.0 nmol per cell per d, respectively. E. caudatum was responsible for almost the entire methane production in the culture. The activity of free methanogens constituted approximately 50% of the total methane production in the E. ecaudatum culture. Decrease of digestibility of substrates and differences in the fermentation end products accompanied the inhibition of methanogenesis in both cultures by penicillin G, streptomycin, chloramphenicol, 2-bromoethanesulfonate, and pyromellitic diimide. E. caudatum appeared to be more sensitive than E. ecaudatum to the compounds tested. Hydrogen recoveries based on both volatile fatty acids and methane production suggested that the methanogenic population appeared not to be fully able to consume hydrogen produced in the protozoan cultures. The culture conditions tested were found to be suitable for experiments on the relationship between rumen ciliates and rumen bacteria. PMID- 11271815 TI - Effect of dichromate on population and growth of various protozoa isolated from industrial effluents. AB - Three protozoa belonging to genera Euglena, Vorticella and Stylonychia collected from industrial wastes were cultured in a medium containing inorganic salts, basically meant for the growth of algae. Protozoa showed rapid growth in the medium. Hexavalent chromium (K2Cr2O7) at a concentration of 5 micrograms/L in the medium adversely affected the growth of protozoa. At the end of eight days of Cr administration, the population of Euglena, Vorticella and Stylonychia increased 8 , 4.5- and 10-fold, respectively, as against 30-, 6.75- and 50-fold increase in the control cultures. No apparent death phase and no change in activity or morphology of protozoa was observed at this Cr concentration. The protozoa were also exposed to different metal ions, viz. Pb (2.42 mmol/L), Cr (0.48 mmol/L), Cd (0.36 mmol/L), administered in the culture medium for a period of 2 years. The metal tolerance for S. mytilus and V. microstoma was Pb > Cr > Cd. E. proxima could not tolerate any of the long-term metal treatments. Because of the ability of these protozoa to tolerate high concentrations of heavy metals, their potential role in remediation of heavy metals from industrial wastewater is considered. PMID- 11271816 TI - The membrane potential of Methanobacterium thermoautotrophicum under different external conditions. AB - The membrane potential (delta psi) of whole cells of Methanobacterium thermoautotrophicum strain delta H was estimated under different external conditions using a TPP(+)-sensitive electrode. The results show that the delta psi values of M. thermoautotrophicum at alkaline pHout (8.5) are comparable with delta psi values under slightly acidic conditions (pH 6.8; 230 and 205 mV, respectively). On the other hand, the size of colonies on Petri dishes was remarkably smaller at pH 8.5 than at 6.8. The delta psi was insensitive to relevant ATPase inhibitors. At pH 6.8, the protonophore 3,3',4',5 tetrachlorosalicylanilide (TCS) strongly inhibited delta psi formation and ATP synthesis driven by methanogenic electron transport. On the other hand, at pH 8.5 the CH4 formation and ATP synthesis were insensitive to TCS and a protonophore resistant delta psi of approximately 150 mV was determined. The finding of a protonophore-resistant delta psi at pH 8.5 indicates that at alkaline pHout these cells can switch from H(+)-energetics to Na(+)-energetics, when the delta [symbol: see text] H+ becomes limited. The results strongly support the hypothesis that at alkaline pHout Na+ ions might fully substitute for H+ in these cells as the coupling ions. PMID- 11271817 TI - Construction of promoter-probe shuttle vectors for Escherichia coli and corynebacteria on the basis of promoterless alpha-amylase gene. AB - We constructed new promoter-probe vectors for E. coli and corynebacteria based on the promoterless alpha-amylase gene originating from Bacillus subtilis. Vectors pJUPAE1 and pJUPAE2 are suitable for isolation of transcriptionally active fragments from plasmids, phages or genomic DNA. alpha-Amylase activity can be easily visually detected on agar plates containing a chromogenic substrate, or by direct measurement of alpha-amylase activity. PMID- 11271818 TI - Isolation and characterization of a novel phytase from Penicillium simplicissimum. AB - Eighty-three isolates from different soil samples exhibited the potential for producing active extracellular phytase. The most active fungal isolate with phytase activity was identified as Penicillium simplicissimum. In shaking culture with enrichment medium, the highest extracellular phytase activity of the producing strain was 3.8 U/mL. The crude enzyme filtrate was purified to homogeneity using ultrafiltration. IEC and gel filtration chromatography. The molar mass of the purified enzyme was estimated to be 65 kDa on SDS-PAGE. The saccharide identification with periodic acid-Schiff reagent (PAS) and activity recognition by 1-naphthyl phosphate was all positive. The isoelectric point of the enzyme, as deduced by isoelectric focusing, was pH 5.8, the optimum pH and temperature being pH 4.0 and 55 degrees C, respectively. The purified enzyme revealed broad substrate specificity and was strongly inhibited by Fe2+, Fe3+ and Zn2+; however, no inhibition was found by EDTA and PMSF. Phytase activity was inhibited when 2 mmol/L of dodecasodium phytate was added and the Km for it was determined to be 813 mmol/L. PMID- 11271819 TI - Formation of myo-inositol phosphates by Aspergillus niger 3-phytase. AB - Kinetics of phytate hydrolysis by Aspergillus niger phytase and correlation between the amount of released phosphate and creation of lower myo-inositol phosphates were investigated. Phytase was able to hydrolyze myo-inositol hexakis , pentakis-, tetrakis-, and trisphosphates. Finally, about 56% of total phosphate were released and myo-inositol bisphosphate was detected as the end-product. PMID- 11271820 TI - Antibacterial effects of trisubstituted quinazoline derivatives. AB - Five trisubstituted quinazolones and eight trisubstituted quinazoline-4-thiones have been tested for antibacterial effects by a microdilution method. Four derivatives exerted a significant effect on E. coli, P. aeruginosa, S. aureus and B. subtilis (IC50 < 100 mg/L). In the bacterium P. aeruginosa six quinazolines showed a higher antibacterial effect than ampicillin. The most sensitive to the effects of the quinazolines was S. aureus; a concentration of 100 mg/L of six derivatives induced a bacteriostatic effect on S. aureus. The quinazoline-4 thiones were generally more active than the quinazolones. All the tested concentrations of the four most effective quinazolines influenced the specific growth rate. PMID- 11271821 TI - Antifungal properties of substituted 1-phenyl-5-mercaptotetrazoles and their oxidation product, 5-bis-(1-phenyltetrazolyl)disulfide. AB - The antifungal effect of substituted 1-phenyl-5-mercaptotetrazoles was tested with Candida tropicalis, C. pseudotropicalis, C. mogii, Trichosporon cutaneum, Cryptococcus albidus and S. cerevisiae. Candida strains exhibited the lowest sensitivity to the compounds; the most sensitive was S. cerevisiae. The MIC values ranged from 40 to > 1000 mg/mL. The antifungal effect of halogenated compounds decreased in the series of bromo > chloro > fluoro derivatives. The electrochemical oxidation of substituted 1-phenyl-5-mercaptotetrazole derivatives in an acetonitrile medium was studied as a model for the enzymic oxidation of the substance, including study of the effect of water, perchloric and trifluoromethanesulfuric acids on E1/2 and I1. 5-Bis-(1 phenyltetrazolyl)disulfide, the compound with no antifungal effect, has been identified as the main oxidation product of 1-phenyl-5-mercaptotetrazole. PMID- 11271822 TI - Polysaccharide hydrolases of Aureobasidium pullulans. AB - The polysaccharide hydrolase activity of a group of selected strains of the genus Aureobasidium pullulans was investigated using a new gel testing assay. A total of 31 strains were tested for alpha-amylase, alpha-glucosidase and glucoamylase, beta-glucosidase, lichenase, cellulase, xylanase and xylosidase, mannanase and mannosidase production during growth of microorganisms on respective meshed polysaccharide gels. Attempts were made to increase the polysaccharide hydrolase activity through selection of some A. pullulans strains by passaging them on the respective modified xylanase- and cellulase-containing gels. The individual saccharide degradation cleavage products were investigated by chromatography. PMID- 11271823 TI - Findings of mycobacteria in insectivores and small rodents. AB - The organs of 30 insectivorous mammals and 62 rodents from areas inhabited by people or livestock where cattle paratuberculosis or mycobacterial infections of swine had been found to occur were examined by cultivation during the monitoring of occurrence and spread of mycobacterioses in cattle and swine. Mycobacteria were found in the organs of 3 insectivores (10%) and 6 rodents (9.7%). Mycobacterium chelonae was isolated from the organs of the lesser white-toothed shrew (Crocidura suaveolens) and the common vole (Microtus arvalis), and M. vaccae and M. avium subsp. avium (IS901+, serotype 1) from the organs of the common shrew (Sorex araneus). M. avium subsp. avium (IS901+, serotype 1) was also isolated from the organs of the yellow-necked mouse (Apodemus flavicollis). Slow growing mycobacteria of group III (according to Runyon) were isolated from the organs of the mouse (Mus musculus sensu lato) and the yellow-necked mouse (A. flavicollis). These findings had no connection with the epizootological situation in the nearby livestock. M. fortuitum was isolated from the organs of the common vole (M. arvalis) caught in a field within easy reach of a swine breeding herd. M. fortuitum was also identified in the lymph nodes and droppings of this swine herd, as well as in the straw, scrapings from the floor of stalls, troughs and banisters, as well as from larvae and imagoes of dipterous insects. These results demonstrate the possibility that insectivores and small rodents can spread the causative agents of mycobacteria in wild and domestic animals. PMID- 11271824 TI - Effect of Al, Co, and Pb ions on growth of Frankia spp. in a mineral medium. AB - Growth of three Frankia strains associated with roots of Casuarina tree, treated with different concentrations of metal ions, was measured as total protein content. One strain was highly resistant to different aluminum ion concentrations up to 10 mmol/L. The other two strains were more sensitive to the higher aluminum concentrations (1.25-10 mmol/L). Growth inhibition by cobalt and lead concentrations varied, depending on the tested strain. Stimulation occurred only at cobalt concentrations of 0.33 and 0.65 mmol/L for one strain. PMID- 11271825 TI - Use of the molecular typing methods to evaluate the control of Listeria monocytogenes contamination in a raw milk and dairy products. AB - Nineteen serogroup 1/2a Listeria monocytogenes strains isolated from raw milk, dairy products and salt water in one dairy were analyzed. Pulsed field gel electrophoresis (PFGE) and ribotyping were used to determine whether these strains isolated over a 8-month period are epidemiologically related. The samples of raw milk were contaminated by different L. monocytogenes clones. The clones isolated from dairy products (with the exception of one sample) and salt water were identical. Comparative genetic analysis of the clones isolated from raw milk, salt water and dairy products revealed the source of contamination and identified the L. monocytogenes strain involved in this process. PMID- 11271826 TI - Detection of feline immunodeficiency provirus by seminested polymerase chain reaction. AB - Specific primers for the detection of the FIV provirus in peripheral blood mononuclear cells (PBMC) by seminested PCR (snPCR) were developed. Forty cats (patients from veterinary hospitals) were investigated for the presence of FIV serum antibodies by immunoblot and for the presence of provirus in PBMC by snPCR. Seventeen of the 40 examined samples (42.5%) showed the presence of antibodies against FIV. The total number of animals that were found positive in snPCR was 19 (47.5%). Fourteen of the seropositive animals (35%) were positive by snPCR whereas three seropositive animals (7.5%) turned out to be snPCR negative. Of twenty-three animals that were negative by immunoblot, five (12.5%) were found to be positive by snPCR. PMID- 11271827 TI - Effect of spermine on proliferation of hyphae of Glomus fistulosum, an arbuscular mycorrhizal fungus, in maize roots. AB - Effects of two oligoamines, putrescine and spermine, on proliferation of intraradical hyphae in surface disinfected root segments were studied under axenic conditions in vitro. No significant effects of putrescine were observed. Spermine significantly stimulated hyphal growth at a concentration of about 1.5 mumol/L. High concentration (> 150 mumol/L) caused a strong inhibition of hyphal growth and of the percentage of root segments bearing proliferating hyphae. DL alpha-difluoromethylornithine, a metabolic inhibitor of polyamine synthesis, caused a significant inhibition of proliferation of the hyphae only in the presence of 2 mumol/L spermine. PMID- 11271828 TI - Antifungal activity of venenatine, an indole alkaloid isolated from Alstonia venenata. AB - The indole alkaloid venenatine exhibited antifungal activity against some plant pathogenic and saprophytic fungi. Venenatine in an aqueous acetic acid solution inhibited spore germination of all the 10 tested fungi, Fusarium udum, Alternaria brassicicola, Ustilago cynodontis and Aspergillus flavus showed an especially high sensitivity towards this compound, exhibiting germination levels below 10%. The spore germination and colony development of the parasitic fungus Erysiphe pisi, which causes powdery mildew in pea (Pisum sativum), on excised leaves of pea was also significantly affected. Pre-inoculation rather than post inoculation treatment of the leaves was more inhibitory against spore germination and colony development. PMID- 11271829 TI - Cold stress induces switchover of respiratory pathway to lactate glycolysis in psychrotrophic Rhizobium strains. AB - Two psychrotrophic strains of Rhizobium, DDSS69, a non-cold acclimated strain, and ATR1, a cold acclimated strain, were subjected to cold stress. A 4-fold increase in the specific activity of lactate dehydrogenase (LDH) was characteristic for cold stressed cells of DDSS69, whereas ATR1 showed a higher LDH activity in general, which increased 1.5-fold under cold stress. Cold sensitive mutants of DDSS69 which could not grow below 15 degrees C, in contrast to the wild type which could grow at 5 degrees C, were isolated using Tn5-tagged mutagenesis. These mutants showed a 40% lower LDH activity than the wild type grown at 5 degrees C that was comparable to the wild type grown at 15 degrees C. High specific activity of succinic dehydrogenase (SDH) at 28 degrees C in both strains and mutants indicated that aerobic respiration via the citrate cycle is the normal mode of saccharide utilization. Shifts to lower temperatures decreased the specific activity of SDH. However, alcohol dehydrogenase (ADH) activity remained very low in both the strains and the mutants at low temperatures indicating that a shift from aerobic saccharide metabolism to anaerobic one under cold stress involves lactate glycolysis rather than alcohol fermentation. There was an increase in membrane-bound ATPase activity under cold stress which is correlated to higher LDH activity. These data show that, in psychrotrophic Rhizobium strains, cold stress induces a switchover of respiratory metabolism from aerobic to anaerobic pathway, especially lactate glycolysis. PMID- 11271830 TI - Serotypic characterization of group A rotaviruses associated with children's diarrhea in Slovakia. AB - A total of 368 rotavirus RNA-positive (PAGE) stool samples collected continually during 1992-95 from infants and young children under five years of age hospitalized with acute gastroenteritis were serotyped using an enzyme immunoassay with VP7-specific monoclonal antibodies (ELISA with MAbs) for serotypes G1-G4. The serotype was identified in 106 stool samples (29%). Comparison of electropherotype and serotype profile in individual samples did not show any remarkable correlation. The members of three electropherotypes (A, C, K) belonged to all 4 serotypes. The representatives of two electropherotypes (E, G) and of mixed electropherotype did not react with any of the specific monoclonal antibodies used. The distribution of the serotypes was scored as 52, 13, 14 and 7.5% for G1 G4, respectively, whereas 13% of samples reacted with two or more type-specific monoclonal antibodies. The G1 serotype dominated during the period followed. PMID- 11271831 TI - Changes in potential denitrification and respiration during the cold storage of soils. AB - The denitrification potential in moderately fertilized soil sampled four times during 1995 decreased significantly after cold storage, at 4 +/- 2 degrees C for 1 week. Prolonged storage (up to 24 weeks) resulted in a further decrease of denitrification potential which dropped to 38-54% of the original values. Similarly, denitrification potential decreased substantially during the first week of storage in differently fertilized soils. After 24 weeks of storage, denitrification potential dropped to 29-55% of that in fresh soils. The effects of storage at 4 +/- 2 degrees C on denitrification potential and respiration (determined as carbon dioxide evolution) were in general the same in moderately fertilized soils from four different sites: in all soils, depression of both the denitrification potential and potential respiration was found after 8 weeks. However, the extent to which the parameters were decreased differed from case to case. Not only the duration and storage conditions but also unidentified soil parameters are important for the persistence of biological activity in stored soils. PMID- 11271832 TI - Mitochondria--tool for taxonomic identification of yeasts from Saccharomyces sensu stricto complex. AB - Mitochondrial genomes of Saccharomyces and close relatives previously used for transplacement of mitochondria to S. cerevisiae were examined. The origins of replication in mitochondrial DNA, the presence of nuclear and mitochondrial polymorphic loci and the ability to produce mitochondrial respiration-deficient mutants were used to reclassify some collection yeasts and to assign others into four separate subgroups. The first included isolates identical to Saccharomyces cerevisiae (S. italicus, S. oviformis, S. chevalieri and S. capensis) which possess 5 or more replication origins. The second group consists of S paradoxus (var douglasii) mitochondrial genome with the equal number of ori sequences but incompatible mitochondria. The third group represents Saccharomyces sensu stricto petite-positive species (S. carlsbergensis, S. heterogenicus, S. uvarum, S. willianus) with 1-2 origins of replication significantly different from S. cerevisiae. In addition, the locus between tRNA(fMet) and tRNA(Pro) is about one half of the 1400 bp members of S. cerevisiae complex. The last group includes isolates that do not belong to Saccharomyces sensu stricto group as they are petite-negative and devoid of any S. cerevisiae-like replication origins. PMID- 11271833 TI - Possible causes of nicarbazin residues in chicken tissues. AB - Two experiments were carried out to investigate possible causes of nicarbazin residues in broiler chicken tissues. The first experiment was designed to establish whether feeding nicarbazin as stipulated in the product license can result in 4,4'-dinitrocarbanilide (DNC) tissue residues exceeding the JECFA MRL (200 micrograms/kg). It was shown that the MRL was exceeded in the livers of broilers housed on deep litter, but not in those of broilers housed on wire flooring. Muscle DNC concentrations were well below the MRL. The higher residual tissue concentrations in birds housed on deep litter were attributed to faecal recycling. The second experiment was to establish the relationship between nicarbazin-contaminated withdrawal ration up to the point of slaughter and DNC residues in the tissues of broilers that had not been previously exposed to nicarbazin. Tissue DNC concentrations were found to be proportional to feed concentrations. The housing method caused no significant difference in tissue residues. Meal containing nicarbazin at a concentration of 2.4 mg/kg or greater caused liver DNC residues above the JECFA MRL. Violative residues may, therefore, occur in chickens not exposed to nicarbazin during rearing, but fed withdrawal ration contaminated at 2.4 mg/kg or greater, or in chickens housed on deep litter and fed nicarbazin-medicated meal according to the product license even when the withdrawal ration is nicarbazin-free. PMID- 11271834 TI - Fish for human consumption: risk of contamination by mercury. AB - Total mercury concentrations were measured in the muscle of different kinds of fish: megrim (Lepidorhombus boscii), common sole (Solea vulgaris), striped mullet (Mullus barbatus), anglerfish (Lophius piscatorius), and black-bellied angler (Lophius budegassa), caught in the South Adriatic Sea (South Italy). The highest total mercury levels were found in anglerfish (0.61-2.22 mg/kg wet wt, mean 1.26 +/- 0.58), followed by black-bellied angler (0.22-1.62 mg/kg wet wt, 0.68 +/- 0.36), megrim (0.05-0.92 mg/kg wet wt, 0.39 +/- 0.30), striped mullet (0.10-0.63 mg/kg wet wt, 0.31 +/- 0.13) and common sole (0.05-0.44 mg/kg wet wt, 0.19 +/- 0.15). According to current regulations, 62.5% of anglerfish (Lophius piscatorius) and 23% of black-bellied angler (Lophius budegassa) samples showed concentrations exceeding the peak value of 1 mg/kg, while only 25% of samples of megrim (Lepidorhombus boscii), and 8.3% of striped mullet (Mullus barbatus), exceeded the peak value fixed at 0.5 mg/kg. Correlations between total mercury concentration and specimen weight were evident in all the species examined. PMID- 11271836 TI - The effects of T-2 toxin exposure on liver drug metabolizing enzymes in rabbit. AB - High doses of T-2 toxin are known to decrease protein synthesis and mono oxygenase activities in rat liver. The purpose of this study was to investigate whether exposure at a low dose could alter the normal metabolism of the xenobiotic by the liver. Three doses of T-2 toxin, dissolved in olive oil, were orally and daily administered to New Zealand white rabbits for five days. At 0.5 mg/kg, three of the five animals died, whereas only a weak decrease in body weight gain and moderate signs of toxicity occurred in rabbits receiving 0.25 mg/kg/day, and the body weight increased without signs of toxicity at 0.1 mg/kg/day. At 0.25 mg/kg/day, total liver microsomal P450 content, and the activities of aminopyrine and benzphetamine N-demethylases, pentoxyresorufin O depentylase, glutathione S-transferases accepting 1-chloro-2,4-dinitrobenzene and 1,2-dichloro-4-nitrobenzene as substrates, were decreased. By contrast, ethylmorphine and erythromycin N-demethylases, ethoxyresorufin and methoxyresorufin O-dealkylases, aniline hydroxylase, and UDP glucuronyltransferase accepting p-nitrophenol as substrate, were unaffected. The expression of P450 1A1, 1A2, 2A1, and 2B4, but not P450 2C3 and 3A6, were also decreased, whereas microsomal conjugated dienes, fluorescent substances, and malondialdehyde contents were increased. At 0.1 mg/kg/day, neither significant effects on drug metabolizing enzymes nor microsomal oxidative damages were obtained. Taken together, these results suggest that a short exposure time to the mycotoxin would not be associated with significant changes in the normal metabolism of xenobiotics by the liver. PMID- 11271835 TI - Assessment of the levels of chlorinated pesticides in breast milk in Kuwait. AB - Breast milk samples, randomly collected from 32 Kuwaiti donors were analysed for chlorinated pesticides. DDE residues ranged from 127 to 3333 micrograms/kg, averaging, 833 micrograms/kg, expressed on a fat weight basis. DDT levels ranged from 0.6 to 67 micrograms/kg fat and averaged 12.4 micrograms/kg, fat. High DDE/DDT ratios were found, which indicated that the exposure to DDT, in most cases, happened quite some time ago. In addition many of the samples also contained isomers of HCH, heptachlor-epoxide, aldrin, dieldrin and endrin. Residue levels of non-DDT pesticides were generally low. Overall levels of DDT pesticides in Kuwaiti milk samples were lower than levels reported from other Middle Eastern countries, although methodologies may not be directly comparable. PMID- 11271838 TI - Toxicological evaluation of commercial mineral water bottled in polyethylene terephthalate: a cytogenetic approach with Allium cepa. AB - The aim of this study was to ascertain the possible toxicological effects of chemicals released into mineral water packaged in polyethylene terephthalate (PET) bottles. Two commercial mineral waters, bottled both in PET and glass and stored under different conditions, were examined using the Allium cepa test. The influence of the water samples on macroscopic (root length, colour and form) and microscopic (root tip mitotic index, chromosome aberrations) parameters was examined. The water samples were analysed after: (A) controlled-condition storage (no direct light exposure and 18 +/- 2 degrees C), (B) storage at 40 degrees C for 10 days, in the dark (migration test in accordance with 82/711/EEC), and (C) exposure to sunlight and varying temperatures (18-38 degrees C, mean temperature 25 +/- 3 degrees C). The two water samples bottled in PET induced cytogenetic aberrations regardless of the storage conditions. These signs of toxicity were evident even only 8 weeks after bottling, which is well within the recommended expiry date. Storage conditions were very important, as is suggested by the finding that chromosomal aberrations were particularly apparent after exposure to direct sunlight. However, as plant systems are not considered as primary screening tools by current international guidelines for mammalian systems, extrapolation of the results from this test system to other systems and, eventually, to human beings should be based on results from a battery of assays covering various metabolic pathways. PMID- 11271837 TI - Deoxynivalenol and ochratoxin A in German wheat and changes of level in relation to storage parameters. AB - The occurrence of the mycotoxins deoxynivalenol (DON) and ochratoxin A (OTA) in the winter wheat of 1997 and 1998 grown under organic farming conditions was investigated using ELISAs (R-Biopharm) for quantification. The influence of delayed drying of the grain after harvest on the development of DON and OTA was determined in storage trials (moisture: 17% and 20%; temperature: 20 degrees C; duration: four and six weeks). The Tox5 PCR assay was used both to detect Fusarium species with the potential to produce trichothecenes and as a measure of their relative DNA content during the storage trials. The intensity of the PCR signals was correlated with the DON concentration. Fusarium species were identified microscopically by standard methods. All the freshly harvested grain samples were contaminated with DON and showed further increases in the DON concentration during storage. OTA contamination was found in 14.3% of the 1997 samples and in 24.1% of the 1998 samples. OTA increased during storage trials of the 1997 samples but not in the 1998 samples. PMID- 11271839 TI - Properties of polyethylene films with incorporated benzoic anhydride and/or ethyl and propyl esters of 4-hydroxybenzoic acid and their suitability for food packaging. AB - Benzoic anhydride and ethyl and propyl esters of 4-hydroxybenzoic acid (ETP and PRP, respectively, also termed parabens) incorporated into low density polyethylene (LDPE) film were studied with regard to migration into food and food simulants at 6 degrees C and 25 degrees C, and changes in selected properties of the film were investigated. Antimicrobials were incorporated into polymer film in concentrations of 5 g/kg and 10 g/kg. The addition of parabens into the polymer was more difficult than benzoic anhydride due to their volatility. For benzoic anhydride, 30-40% and 10-20% of the added amount was found to leach from the film into aqueous and olive oil food simulants, respectively. The migration into both water and olive oil followed a very similar course in the case of parabens. Migration levels over 90% and in the range of 70% to 80%, relative to the amount of agent in the film, were determined for ETP and PRP respectively. The incorporation of antimicrobials into the film significantly changed the functional characteristics of the packaging material, i.e. permeability of oxygen, carbon dioxide and water vapour, tensile strength, coefficient of friction, sealing strength and transparency. Shelf life tests with packaged cheese and toasted bread demonstrated the efficiency of the film containing 10 g/kg of BA against mould growth on the food surface during storage at 6 degrees C. PMID- 11271840 TI - Influence of various parameters on the browning of potassium sorbate in the presence of amines. AB - Potassium sorbate can undergo oxidation to form carbonyl moieties and cause browning. This investigation studied the fate of the compounds produced by auto oxidation of potassium sorbate and measured the browning in the presence of amines. Experimental plans in which four factors were combined (temperature, oxygen, amine and light) led to the observation that the interaction between oxygen and high temperature (75 degrees C) favoured browning, independently of the presence of amine. At 4 degrees C, the amine seemed to cause a decrease in the proportion of carbonyl-containing compounds which would appear to participate in carbonyl-amine reactions. At 75 degrees C, the amine forms adducts with the potassium sorbate. In parallel, high temperature favours auto-oxidation reactions that produce carbonyls. GC/MS and NMR analysis of the reaction products of potassium sorbate/amine mixtures led to the identification of cyclic products. PMID- 11271841 TI - Analysis of benzo[a]pyrene in spiked fatty foods by second derivative synchronous spectrofluorimetry after microwave-assisted treatment of samples. AB - A simple, rapid and inexpensive method has been developed for the determination of benzo[a]pyrene (B(a)P, a known carcinogen) in foods with a high fat content. In-house validation was carried out by checking B(a)P recovery +/- precision from those samples by a simple spiking procedure. The method involves extracting the fat from a freeze-dried product and the saponification of the B(a)P-containing lipid fraction assisted by microwave energy. After partitioning in hexane, and purification by passage through a silica cartridge, the B(a)P-containing hexane eluate is analysed by second derivative synchronous spectrofluorimetry. The method was found to have a recovery of 90 +/- 5%. The detection and quantification limits in food (0.05 and 0.12 microgram/kg, respectively) indicate that the B(a)P maximum that the European Union (EU) intends to set for foods (1 microgram/kg), can be monitored by this method. PMID- 11271842 TI - Effect of novel water-soluble polymeric forms of sorbic acid against Fusarium oxysporum f.sp. radicis-cucumerinum. AB - New controlled release water-soluble formulations of sorbic (2,4-hexadienoic) acid were prepared and their inhibitory activity on mycelium growth of Fusarium oxysporum f.sp. radicis-cucumerinum was evaluated. The new products are epoxidized polymers of polyvinylpyrrolidone (PVP) containing covalently bonded sorbic acid (polymeric esters of sorbic acid) and complexes of PVP with hydrogen bonded sorbic acid, characterized by controlled release of sorbic acid. It was shown that the polymeric complexes of sorbic acid with PVP were more effective fungicidal agents than sorbic acid polymeric esters. In all cases the activity of polymeric derivatives (esters and complexes) was increased by lowering the molecular weight of the polymeric carriers. Controlled release formulations of these polymeric derivatives are new promising products due to their low toxicity, wide range of efficient concentrations for application and ability to regulate lyophilicity. Our data contribute to the understanding of the action mechanism of various polymeric sorbic acid formulations and can result in products which are particularly suitable for food and feed protection applications. PMID- 11271843 TI - The determination of total SO2 in grape juice. A comparison among five methods. AB - The EC official method of total SO2 analysis in grape juice was modified in 1990. The main improvements concerned the amount and concentration of H3PO4 used during the distillation to recover the combined SO2 and the standardization of the distillation time at 15 min. This comparative study evaluated the total SO2 level of 12 grape juices determined by five methods, including distillation, iodimetric and enzymatic-based methods. Attention was focused on the total SO2 legal limit of 10 mg/l fixed in Europe for grape juice. Analysis of variance disclosed a significant difference among the total SO2 content in grape juices determined by five methods. Each analytical method showed limits in relation of their ability to release the combined SO2. In particular, the SO2 bonded to phenolic compounds is partially released at low pH in the acidified juice leading to higher results. PMID- 11271844 TI - Dietary intake exposure to sulphites in Italy--analytical determination of sulphite-containing foods and their combination into standard meals for adults and children. AB - The theoretical risk of exceeding the Acceptable Daily Intake (ADI) for sulphites has mostly been examined on the basis of a worst-case scenario. In order to examine the real situation the determination of residue sulphite levels in ready to-consume foods is required. The aim of this paper is to assess the actual sulphite content of diets obtained from a combination of realistic meals high in sulphite-containing foods. Food products available in Italy containing added sulphites were identified. Overall, 211 samples of foods and beverages (including 85 samples of wine) were collected. The determination of sulphite residues was carried out on the foods which were prepared according to normal domestic practice. It was shown that the diets obtained from these foods would lead to an intake of 23 mg/day in children and 50 mg/day in adults (both slightly above the ADI for respectively a 30 kg child and a 60 kg adult). Among all sulphite containing foods, the highest contributors to the intake were dried fruit and wine, both ingested without further treatment. The analysis of specific consumption data confirmed the existence of a risk of exceeding the ADI related to sulphite residue levels in wine. PMID- 11271846 TI - Dietary change, energy balance and body weight regulation among migrating students. AB - This study was conducted to examine how subjects modulate their food intake and energy balance when they migrate from a low energy density food intake pattern to one of high energy density. It was hypothesised that an increase in the energy density of food consumed would result in increased body weight of the migrating subjects unless food intake and energy balance could be modulated. Food selection, food intake, basal metabolic rate (BMR) and anthropometric measurements were made on 53 female and 56 male newly arrived overseas students. All subjects were from Malaysia, but the data was collected at Oxford Brookes University where the subjects were studying. Food intake using 3-day food diaries and food frequency questionnaires (FFQs). BMR and anthropometric measurements including body weight were measured on arrival in the UK and after 3 and 6 months' stay. Student's t-tests and analysis of variance (ANOVA) were used to compare the data. A significant difference (P < 0.05) was found between the energy density of the foods consumed in Malaysia and after 3 and 6 months in the UK. There was also a significant decrease (P < 0.05) in protein consumed. However, there were no differences in total energy intake. From results of the FFQs, differences were found in food selection due mainly to the lack of availability of certain foods in UK supermarkets. No significant differences were found in the BMR and anthropometric measurements made at the start of the study and later assessments. It appears that Malaysian students are able to remain in energy balance and are weight stable at least during the first 6 months of residence in the UK, despite the wider choice of energy dense food available. This suggests that at least in the short term, subjects are able to modulate their food intake in response to changes in the energy densities and free choice of food. PMID- 11271845 TI - Microbiological and chemical detection of incurred penicillin G, oxytetracycline, enrofloxacin and ciprofloxacin residues in bovine and porcine tissues. AB - Incurred penicillin G, oxytetracycline, enrofloxacin and ciprofloxacin residues in bovine and porcine muscle and kidney samples were analysed by microbiological and chemical methods, the former using Bacillus subtilis BGA as a test organism on agar media of pH 6, pH 7.2 and pH 8 and the latter using liquid chromatography. Least squares fits between the logarithms of the chemically obtained concentrations of the antimicrobials and the widths of the inhibition zones were used to estimate the inhibition zone widths corresponding to the maximum residue limit concentrations. In vitro sensitivities were determined with standard antimicrobial solutions. The results indicate that if B. subtilis BGA is used as a test organism, muscle tissue cannot be used as test material for screening oxytetracycline, enrofloxacin and ciprofloxacin residues on the plates used in this study, while penicillin G can be screened from muscle tissue. Because of the numerous factors causing or increasing variation in the analysis, the inhibition zone caused by a given antibiotic concentration cannot be predicted precisely. Therefore, a positive agar diffusion test needs to be confirmed chemically. If a kidney sample gives a positive agar diffusion test result, the antimicrobial concentration in a muscle sample from the same carcass should be checked chemically. PMID- 11271848 TI - Implementation of ISO 9000 to the food industry: an overview. AB - Since the early 1980s, manufacturing industries worldwide have seen a revolution in the way they operate. Consumers have become more and more demanding and the key to company/firm survival is the recognition of customers' satisfaction. In a way, companies have been forced to enhance the quality of their processes and their products. Some of them chose to establish internal quality systems whereas others have opted for employing a general quality system standard, such as the BS 5750 or the ISO 9000 series. In Greece, a great part of the companies (about 80%) employ quality system standards, among which the highest percentage (80%) belong to food manufacturing industries. PMID- 11271847 TI - Time spent outdoors and seasonal variation in serum concentrations of 25 hydroxyvitamin D in Korean women. AB - The vitamin D status of 179 Korean women between the ages of 20 and 75 were measured by the use of high performance liquid chromatography (HPLC). Related biochemical indices such as iPTH, alkaline, phosphatase, creatinine, albumin, Ca, Mg, and P were also measured. Factors such as demographic characteristics, intake of foods containing vitamin D, and proxy measure of sunlight exposure (time spent outdoors) were assessed to determine their effect on vitamin D status and used in the analysis. The purpose of this study was twofold. The first was to define reference data for the distribution of vitamin D status and to explore the relationship between vitamin D and the variables that affect the vitamin D status in Korean women. The second was to analyze the risk factors of the vitamin D status and the relation between the factors. The results of this study will provide valuable information regarding the role of vitamin D in Korean women. The mean serum 25-hydroxyvitamin D (25-OHD) level was 25.8 ng/ml. Of the total subjects, 16.5% showed vitamin D deficiency (i.e. s-25-OHD < 10 ng/ml). Serum 25 OHD was inversely related to iPTH and alkaline phosphatase. There were significant changes in serum 25-OHD level from the pre- to the post-menopausal women with a positive correlation between vitamin D intake and serum at the 25 OHD level. Significant seasonal variation of serum 25-OHD and PTH were noted in 26 of the subjects and the serum 25-OHD level also correlated with sunlight exposure especially at 12.00 p.m. to 2.00 p.m. as assessed by the time spent outdoors. The relative importance of the two sources of vitamin D such as dietary intake (33.6% explained) and endogenous production of the time spent outdoors (19.7% explained) in serum 25-OHD were also evaluated. Multiple regression analysis revealed that the effects of aging on serum 25-OHD could largely be accounted for. Both the decline in dietary vitamin D intake and the time spent outdoors were closely related to the decreasing serum 25-OHD level. Among the determinants of low serum 25-OHD were age, dietary vitamin D intake, serum calcium level and dietary calcium intake, and serum alkaline phosphatase. PMID- 11271849 TI - Study of the effect of lactobacillus GG supplementation in combination with and without arginine aspartate on lipoproteins and liver peroxidation in cholesterol fed rats. AB - The effect of diet integration with lactobacillus GG and arginine aspartate administered singly or together to rats submitted to a cholesterol-enriched diet have been evaluated by measuring both the changes in the levels of cholesterol and triglycerides and the variations of the most indicative parameters of peroxidation in plasma lipoproteins and livers. The administration of lactobacillus GG alone is able to induce a significant hypocholesterolaemic effect while the arginine aspartate singly or together with the lactobacillus does not seem to promote any significant hypocholesterolaemic effect. The cholesterol levels (expressed as mg x dL-1) are in fact: 45.5 for the control diet; 185.4 for the cholesterol-enriched diet; 131.1 for the cholesterol-enriched diet + lactobacillus; 178.2 for the cholesterol enriched diet + arginine aspartate and 122.4 for the cholesterol-enriched diet + lactobacillus + arginine aspartate. On the contrary, the co-administration of lactobacillus and arginine aspartate gives rise to a very high preventive activity against the cholesterol induced peroxidation damages both in the plasma lipoproteins and in the liver. Such preventive activity is higher by far than that obtainable when lactobacillus or arginine aspartate are administered singly to the rats. PMID- 11271850 TI - The effect of browning intensity on the protein quality of qurshallah. AB - The effect of browning intensity of qurshallah, a popular bakery product in Jordan leavened by ammonium bicarbonate and baked twice, on the protein quality was investigated. Light, medium and dark toasted qurshallah samples baked at the same temperature but for different times were used. Data on the proximate analysis of the three different levels of toasted qurshallah indicated significant differences (P < 0.05) regarding the moisture content of the three toasting levels, and in the fat content between the light and the dark toasted samples. Net protein utilization (NPU) using Sprague-Dawley rats, was determined for diets containing the toasted qurshallah samples. Food consumption and gain in body weight of rats decreased with increasing toasting level. Dark toasting of qurshallah caused loss in body weight of animals. The values for NPU operative (NPUop), NPU Standardized (NPUst) and net dietary protein as a percentage of total energy (NDpE%) for light, medium and dark toasted qurshallah diets were significantly different (P < 0.05). The highest values obtained were for the light toasted (75.5, 82.0 and 6.6 respectively), and the lowest values for the medium toasted qurshallah (56.4, 56.3 and 3.7 respectively). The results indicate that intense browning of chemically leavened bakery products might be deleterious to health and should be avoided. PMID- 11271851 TI - Use of palm mid-fraction in dark chocolate as base filling centre at different storage temperatures. AB - Dark chocolates filled with palm mid-fraction (PMF) were stored at different temperatures to evaluate the physical and chemical changes. Storage at low temperature (18 degrees C) reduces the PMF migration to negligible extent. Higher storage temperatures (30 and 35 degrees C) increased the PMF migration from the filling centre into the chocolate coating. As a consequence of fat migration, fatty acid composition, triglyceride composition, hardness, solid fat content, melting point and polymorphic structure changed, leading to bloom formation, which started by fat migration and was influenced by recrystallization tendency within the chocolate coating. PMID- 11271852 TI - Vitamin E and beta-carotene affect natural killer cell function. AB - Vitamin E supplementation has been shown to contribute in immunoregulation, antibody production, and resistance to implanted tumors. Similarly beta-carotene has been shown to down-regulate growth factors which contribute towards proliferation of pre-malignant cells. We embarked upon a study to evaluate the effect of vitamin E and beta-carotene on natural killer (NK) cells, which perform tumor surveillance role in the mammalian body. Mouse splenocytes or human peripheral blood lymphocytes were used as NK cells with murine YAC-1 lymphoma or human K-562 lymphoma cells, respectively, as target cells. The NK cells were treated with vitamin E or beta-carotene while target cells were labeled with sodium 51chromate. Both cell types were then reacted for 4 hours. The NK cell tumorolytic activity was measured by the chromium release assay. Oral administration of alpha-tocopherol at a dose of 100 mg/d in mice showed a significant increase in NK cell activity. Similarly, treatment of NK cells with alpha-tocopherol in vitro at doses 0.5 mg/ml, 1-0 mg/ml, and 2.0 mg/ml increased the tumorolytic activity of NK cells. Tocotrienol showed a similar response at ten times lower dose. When NK cells were treated with varying concentrations of palm vitee (mixture of alpha-tocopherol and tocotrienol), maximum effect was observed at the dose mixture of 12 micrograms and 24 micrograms alpha-tocopherol and tocotrienol, respectively. When murine NK cells were treated in vitro with beta-carotene at doses ranging from 2 ng/mg to 200 ng/ml, a decrease in tumorolytic effect was observed. However, human NK cells after treatment with beta-carotene at doses ranging from 0.1 microgram/ml to 10 micrograms/ml showed a significant increase in tumorolytic function. NK cells were also obtained from mice that had been parenterally administered beta-carotene and alpha-tocopherol. These experiments showed no significant increase in the NK cell function. PMID- 11271853 TI - Effect of palm oil on plasma lipoprotein concentrations and plasma low-density lipoprotein composition in non-human primates. AB - Palm oil (PO) contains approximately 43% of palmitic acid. It is the most abundant saturated fatty acid in the diet and it is generally considered the primary cholesterol (C)-raising fatty acid. However, the effect of palmitic acid on plasma cholesterol appears to depend on the cholesterol content of the diet. The aim of this study was to determine the effect of PO with either a high-fat, high-C or moderate-fat, moderate-C diet on lipoprotein C and low-density lipoprotein (LDL) composition. Fifty adult, male vervet monkeys were randomly assigned to the high-fat diet group (HFD: 35%E fat, approximately 0.106 mg C/kJ; n = 30) and the moderate-fat diet group (MFD: 30%E fat, approximately 0.027 mg C/kJ; n = 30). Baseline LDL-C, high-density lipoprotein (HDL)-C and body weight were used to stratify the vervets into comparable experimental groups within each dietary group. The HFD group was divided into two groups of 10 each: one group continued with the HFD in which 8.1%E was derived from lard (AF); in the other group, AF was substituted isocalorically with PO. The MFD group was divided into three groups of 10 each: one group continued with the MFD in which 11.8%E was derived from AF; in the other two groups, the AF was substituted isocalorically with either sunflower oil (SO) or PO. This article presents preliminary results on plasma lipoproteins and LDL composition after 6 months of dietary intervention. Plasma total and LDL-C was higher in all the groups, but the mean changes elicited by PO with either the HFD or MFD were no different from that observed with AF and SO. There was no difference in the mean change of LDL molecular weight within the HFD and MFD. It is concluded that PO is no different from AF (HFD and MFD) or SO (MFD) in its cholesterolaemic effect. PMID- 11271854 TI - Effects of administration of alpha-tocopherol and tocotrienols on serum lipids and liver HMG CoA reductase activity. AB - Male hamsters were fed on semi-synthetic diets containing commercial corn oil (CO), isolated corn oil triglycerides (COTG), COTG supplemented with 30 ppm of alpha-tocopherol (COTGTL) and COTG supplemented with 81 ppm of alpha-tocopherol (COTGTH) as the dietary lipid for 45 days. Male albino guinea pigs were fed on commercial chow pellets and treated with different dosages of tocopherol and tocotrienols intra-peritoneally for 6 consecutive days. Serum and liver were taken for analysis. Our results show that stripping corn oil of its unsaponifiable components resulted in COTG which yielded lower serum total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) and raised high density lipoprotein cholesterol (HDL-C) and serum triglycerides (TG) levels. These results indicate that the COTG with its fatty acids are responsible for the hypocholesterolemic effect exhibited by corn oil. However, supplementing the COTG diet with alpha-tocopherol (alpha-T) at 30 ppm significantly raised the serum TC, LDL-C and TG levels, but did not alter the HDL-C level, indicating that alpha-T is hypercholesterolemic. Supplementing the COTG diet with alpha-T at 81 ppm raised the serum TC level but to a lesser extent as compared to that obtained with 30-ppm alpha-T supplementation. The increased TC, in this case, was reflected mainly by an increased in HDL-C level as the LDL-C level was unchanged. The TG level was also raised but to a lesser extent than that obtained with a lower alpha-T supplementation. The liver HMG CoA reductase (HMGCR) activity was exhibited (56%) by the COTG as compared to CO. Supplementation of alpha-T at 30 ppm to the COTG diet resulted in further inhibition (76%) of the liver HMGCR activity. On the contrary, supplementation of alpha-T at 81 ppm to COTG diet resulted in a highly stimulatory effect (131%) on the liver HMGCR activity. Short term studies with guinea pigs treated intra-peritoneally with alpha-T showed that at low dosage (5 mg) the HMGCR activity was inhibited by 46% whereas increasing the dosage of alpha-T to 20 mg yielded lesser inhibition (18%) as compared to that of the control. Further increase in the dosage of alpha-T to 50 mg actually resulted in 90% stimulation of the liver HMGCR activity as compared to the control. These results clearly indicate that the effect of alpha-T on HMGCR activity was dose-dependent. Treatment of the guinea pigs with 10 mg of tocotrienols (T3) resulted in 48% inhibition of the liver HMGCR activity. However, treatment with a mixture of 5 mg of alpha-T with 10 mg of T3 resulted in lesser inhibition (13%) of the liver HMGCR activity as compared to that obtained with 10 mg of T3. The above results indicate that the alpha-T is hypercholesterolemic in the hamster and its effect on liver HMGCR is dose dependent. T3 exhibited inhibitory effect on liver HMGCR and alpha-T attenuated the inhibitory effect of T3 on liver HMGCR. PMID- 11271855 TI - Effect of palm vitamin E on the healing of ethanol-induced gastric injury in rats. AB - The effect of palm vitamin E on the healing of ethanol-induced gastric lesions and various biochemical parameters were investigated. The study was divided into two phases. In the first phase of the study, 42 rats of Sprague Dawley species (200-250 gm weight) were randomly divided into two groups fed with a normal diet (control) or palm vitamin E enriched diet (150 mg/kg) for 3 weeks. The rats were killed after 3 weeks of feeding. Gastric tissue contents of malondialdehyde (MDA), prostaglandin E2 and acid were measured. In the second phase of the study 42 rats were divided into two groups. Group 1 was fed normal rat pellets (control) and group 2 was fed palm vitamin E enriched pellets (150 mg/kg food) for 3 weeks. After 3 weeks of feeding gastric mucosal injury was induced by an orogastric tube administration of 0.5 ml 100% ethanol. The rats were killed at 1 hour, 4 hours and 1 week after ethanol exposure for semiquantitative determination of ulcer index and gastric acid concentration. Gastric tissue MDA and mucus were measured only at 1 week after ethanol exposure. In the first phase of the study we found that palm vitamin E only caused a significant reduction in gastric MDA. However, it showed no significant effects on prostaglandin E2 and gastric acid concentration. In the second phase of the study, the mean ulcer index of palm vitamin E supplemented group killed after 1 week of ethanol exposure was significantly lower compared to the respective control. However, there was no significant difference in ulcer index in rats killed at 1 hour and 24 hours after ethanol exposure. The gastric acid concentration was significantly higher in the group treated with palm vitamin E killed 1 week after ethanol exposure compared to control. The gastric tissue MDA was significantly lower in the palm vitamin E supplemented group compared to control. There was no significant difference in gastric mucus content of the both groups. The ulcer healing which occurred in the presence of a high gastric acid suggests that the effect of palm vitamin E on the healing of gastric lesions was not mediated via a reduction in gastric acid nor was it mediated through increasing prostaglandin E2 or mucus production. The most probable mechanism is via reducing lipid peroxidation as reflected by a significant decreased in gastric tissue MDA content. PMID- 11271857 TI - Effects of exchanging 4%en between dietary stearic and palmitic acid on hamster plasma lipoprotein metabolism. AB - This study was designed to determine whether the exchange of specific fatty acids (palmitic (16:0) for stearic (18:0)), would exert differential effects on plasma and lipoprotein lipids, when diets contained approximately 30%en from fat with adequate levels of linoleic acid (18:2). Thirty-two male Golden Syrian hamsters were fed isocaloric purified diets with comparable amounts of 18:2 (approximately 10.5%en). The 18:0-rich diet (50% cocoa butter, 41% safflower oil, 9% sunflower oil) provided 4.8%en 16:0 and 5.3%en from 18:0, while the 16:0-rich diet (59% palm oil, 36% safflower oil, 5% olive oil) provided 8.7%en from 16:0 and 1.2%en from 18:0, resulting in a 16:0/18:0 exchange of approximately 4%en. Both diets contained negligible amounts of lauric and myristic acid (< 0.2%en), approximately 9.5%en from oleic acid and 77 mg cholesterol/1000 kcal. Animals were fed their respective diets for 4 weeks at which point various lipid and lipoprotein parameters were measured. There were no significant difference between dietary groups for any of the measured parameters, which included body weights, food consumption, plasma lipids, lipoprotein lipid and apoprotein concentrations, as well as lipoprotein compositions. Additionally, estimated diameters of various lipoprotein particles were not affected by the fatty acid exchanges employed. Thus these data suggest that when total fat is restricted to 30%en and 18:2 levels are approximately 10%en, a 4%en exchange between 16:0 and 18:0 (representing intakes of approximately 9 g/d/2000 kcal diet) produces comparable plasma lipids. PMID- 11271856 TI - Red palm oil as a source of beta-carotene in a school biscuit used to address vitamin A deficiency in primary school children. AB - The effect of a biscuit with red palm oil as a source of beta-carotene was compared with the effect of a biscuit with beta-carotene from a synthetic source on the vitamin A status of primary school children in a randomised controlled trial. Children aged 5-11 years (n = 265) were randomly assigned to one of three groups: (1) placebo biscuit; (2) biscuit with synthetic beta-carotene as a vitamin A fortificant; and (3) biscuit with red palm oil as a source of beta carotene. The two non-placebo biscuits were designed to provide 34% of the RDA for vitamin A per serving (4 x 15 g biscuits). The biscuits were distributed daily during the school week and compliance was closely monitored and recorded. Children were assessed at baseline and after 6 months of intervention. Mean serum retinol in all three groups increased significantly compared to baseline (P < 0.0001). The prevalence of low serum retinol levels (< 20 micrograms/dL) dropped from 50 to 24.4% in the control group, from 48.2 to 16.9% in the synthetic beta carotene group, and from 50.6 to 22.8% in the red palm oil group. There was no significant treatment effect compared to the control group in either the synthetic beta-carotene or red palm oil group. The increase in the control group was probably due to a school feeding scheme (providing 33% of the RDA for vitamin A) introduced during the latter part of the study. Our results were thus confounded and the 'true' effect of the red palm oil biscuit on vitamin A status could not be established. The study has, however, shown that red palm oil can be incorporated in a biscuit and that the end product with regard to taste and appearance was well accepted by the school children. A follow-up study in a school where there is no school feeding is indicated. PMID- 11271858 TI - The effect of palmitic acid on lipoprotein cholesterol levels. AB - The present study assessed the effect of high versus low palmitic acid intakes of plasma lipoprotein cholesterol levels and on rates for endogenous synthesis of cholesterol in normal and hypercholesterolemic subjects. On day 21 of each diet treatment, a fasting blood sample was drawn for lipoprotein determination and to provide a measure of the background level of deuterium. A priming dose of deuterium was consumed and a second blood sample obtained 24 hours after the first sample. Isotope ratio mass spectrometry was used to determine the incorporation of deuterium into the newly synthesized cholesterol molecule and fractional synthetic rates were calculated. Four diets were formulated to provide combinations of two levels of 16:0 at two levels of 18:2n-6. Subjects received each of the four diet treatments for 21 days, followed by washout periods of 21 days. Serum total cholesterol and LDL-cholesterol was not significantly affected by the high level of 16:0 when diets also contained a high level of 18:2n-6. Fractional synthesis rates of cholesterol observed for each diet treatment did not differ significantly, suggesting no relationship between the endogenous synthesis of cholesterol and dietary 16:0 content. The results indicate that 16:0 has no effect on serum lipoprotein profiles in the presence of recommended intakes for 18:2n-6. PMID- 11271859 TI - Comparing palmolein with different predominantly monounsaturated oils: effect on plasma lipids. AB - Our group has compared plasma lipids in randomised crossover trials in which approximately half the fat intake of subjects was changed from palmolein to each of three predominantly monounsaturated oils in a series of experiments in free living volunteers. With canola oil total cholesterols were lower than on palmolein; part of this reduction was due to lower HDL-cholesterol (HDL-c) so that total cholesterol/HDL-c was only 2% lower. With olive oil mean total cholesterols were the same as on palmolein but HDL-cholesterols were a little lower and LDL-cholesterols a little higher. Plasma lipid fatty acid patterns confirmed the diet change, showing 5% higher 16:0 on palmolein and 11% higher 18:1 on olive oil. To test the possibility that lack of effect of the extra palmitic acid in the palmolein-olive oil comparison was because subjects were young, thin and active, comparison of a third oil, high oleic sunflower oil (HOSO) with palmolein was made in both young and middle-aged subjects. Plasma total and LDL-cholesterols were 7% lower in the whole group on HOSO but HDL-c was also 5% lower so total cholesterol/HDL-c was only 3% lower than on palmolein. There was no difference in lowering of LDL-c on HOSO between young and older subjects. In comparisons of all three predominantly monounsaturated oils with palmolein a higher HDL-c on palmolein reduced the presumed health benefit of lower total cholesterols on canola and high oleic sunflower oil. The reason for no reduction of total cholesterol on olive oil compared with palmolein was presumably due to the higher linoleic and higher phytosterols in palmolein and higher squalene in the olive oil. PMID- 11271860 TI - Vitamin E and factors affecting atherosclerosis in rabbits fed a cholesterol-rich diet. AB - The present study aims to examine the effects of a palm-oil-derived vitamin E mixture containing tocotrienol (approximately 70%) and tocopherol (approximately 30%) on plasma lipids and on the formation of atherosclerotic plaques in rabbits given a 2% cholesterol diet. Eighteen New Zealand White rabbits (2.2-2.8 kg) were divided into three groups; group 1 (control) was fed a normal diet, group 2 (AT) was fed a 2% cholesterol diet and group 3 (PV) was fed a 2% cholesterol diet with oral palm vitamin E (60 mg/kg body weight) given daily for 10 weeks. There were no differences in the total cholesterol and triacylglycerol levels between the AT and PV groups. The PV group had a significantly higher concentrations of HDL-c and a lower TC/HDL-c ratio compared to the AT group (P < 0.003). The aortic tissue content of cholesterol and atherosclerotic lesions were comparable in both the AT and PV groups. However, the PV group had a lower content of plasma and aortic tissue malondialdehyde (P < 0.005). Our findings suggest that despite a highly atherogenic diet, palm vitamin E improved some important plasma lipid parameters, reduced lipid peroxidation but did not have an effect on the atherosclerotic plaque formation. PMID- 11271861 TI - Tocotrienols inhibit growth of ZR-75-1 breast cancer cells. AB - The vitamin E component of palm oil provides a rich source of tocotrienols which have been shown previously to be growth inhibitory to two human breast cancer cell lines: responsive MCF7 cells and unresponsive MDA-MB-231 cells. Data presented here shows that the tocotrienol-rich fraction (TRF) of palm oil and individual fractions (alpha, gamma and delta) can also inhibit the growth of another responsive human breast cancer cell line, ZR-75-1. At low concentrations in the absence of oestrogen tocotrienols stimulated growth of the ZR-75-1 cells, but at higher concentrations in the presence as well as in the absence of oestradiol, tocotrienols inhibited cell growth strongly. As for MCF7 cells, alpha tocopherol had no effect on growth of the ZR-75-1 cells in either the absence or presence of oestradiol. In studying the effects of tocotrienols in combination with antioestrogens, it was found that TRF could further inhibit growth of ZR-75 1 cells in the presence of tamoxifen (10(-7) M and 10(-8) M). Individual tocotrienol fractions (alpha, gamma, delta) could inhibit growth of ZR-75-1 cells in the presence of 10(-8) M oestradiol and 10(-8) M pure antioestrogen ICI 164,384. The immature mouse uterine weight bioassay confirmed that TRF could not exert oestrogen antagonist action in vivo. These results provide evidence of wider growth-inhibitory effects of tocotrienols beyond MCF7 and MDA-MB-231 cells, and with an oestrogen-independent mechanism of action, suggest a possible clinical advantage in combining administration of tocotrienols with antioestrogen therapy. PMID- 11271863 TI - Introducing computer-aided instruction into a traditional histology course: student evaluation of the educational value. AB - Both the World Health Organisation and the World Federation for Medical Education have endorsed integration of computer technology into medical education. In line with this and for more practical reasons, second-year medical students were introduced to commercial and in-house computer-aided instruction (CAI) packages in the traditional histology course. Although CAI was well received, light microscopy was still regarded as a valuable skill in the undergraduate curriculum. Its application was viewed to be important in future medical studies, as well as in general practice. It was also perceived to be a more self-directed form of learning than CAI. Students' use of CAI packages was related to CAI meeting course objectives and the level of interactivity. PMID- 11271862 TI - Computer-assisted planimetry associated with Sturge-Weber syndrome. AB - Photographs of the optic nerve head (optic disc) in the eye are used for the clinical assessment of the disease glaucoma. These images are usually subjectively assessed by a clinician. A case of Sturge-Weber Syndrome which includes glaucoma as a symptom, is presented here. Narrowing of the rim of the optic disc was measured using custom-made measurement software confirming glaucomatous progression. To maximize the information obtained from optic disc images, low cost software can assist with quantifying disc parameters aiding clinical interpretation. PMID- 11271864 TI - The photography of neonatal bereavement at Wythenshawe Hospital. AB - A large number of babies are stillborn or die in the neonatal period in Britain every year. This is only a small percentage of births overall, but these babies represent a considerable loss to the parents, and their hopes and ideals both for themselves and for their child. One of the key aids to grieving is memory, and in those cases where babies are born dead or die in utero, a history of the baby needs to be created. Photography can help create this history, and bereaved parents and families have expressed the importance of photographic images in assisting with their grieving process. PMID- 11271865 TI - The Wellcome Trust Medical Photographic Library Digitization Project: a case study. AB - This study shows the planning process and the first stages of the implementation of a project to digitize over 120,000 photographic negatives and transparencies from The Wellcome Trust's Medical Photographic Library collection. It shows how and why decisions were made and the resulting resourcing implications. Following the planning and proposal stages it details the results of tests carried out after equipment installation, and the subsequent revisions of original estimates and amendments to decisions on resolution levels. The implementation of the scanning project is described along with some modifications made to the work processes as a result of early experience of the project. In conclusion, it suggests that a constant process of review of processes, equipment and planning is necessary to maintain a successful digitization project. PMID- 11271866 TI - Medical illustration and the electronic patient record. PMID- 11271867 TI - Chronic fatigue in general practice: economic evaluation of counselling versus cognitive behaviour therapy. AB - BACKGROUND: There is a paucity of evidence relating to the cost-effectiveness of alternative treatment responses to chronic fatigue. AIM: To compare the relative costs and outcomes of counselling versus cognitive behaviour therapy (CBT) provided in primary care settings for the treatment of fatigue. DESIGN OF STUDY: A randomised controlled trial incorporating a cost-consequences analysis. SETTING: One hundred and twenty-nine patients from 10 general practices across London and the South Thames region who had experienced symptoms of fatigue for at least three months. METHOD: An economic analysis was performed to measure costs of therapy, other use of health services, informal care-giving, and lost employment. The principal outcome measure was the Fatigue Questionnaire; secondary measures were the Hospital Anxiety and Depression Scale and a social adjustment scale. RESULTS: Although the mean cost of treatment was higher for the CBT group (164 Pounds, standard deviation = 67) than the counselling group (109 Pounds, SD = 49; 95% confidence interval = 35 to 76, P < 0.001), a comparison of change scores between baseline and six-month assessment revealed no statistically significant differences between the two groups in terms of aggregate health care costs, patient and family costs or incremental cost-effectiveness (cost per unit of improvement on the fatigue score). CONCLUSIONS: Counselling and CBT both led to improvements in fatigue and related symptoms, while slightly reducing informal care and lost productivity costs. Counselling represents a less costly (and more widely available) intervention but no overall cost-effectiveness advantage was found for either form of therapy. PMID- 11271868 TI - Chronic fatigue in general practice: is counselling as good as cognitive behaviour therapy? A UK randomised trial. AB - BACKGROUND: Fatigue is a common symptom for which patients consult their doctors in primary care. With usual medical management the majority of patients report that their symptoms persist and become chronic. There is little evidence for the effectiveness of any fatigue management in primary care. AIM: To compare the effectiveness of cognitive behaviour therapy (CBT) with counselling for patients with chronic fatigue and to describe satisfaction with care. DESIGN OF STUDY: Randomised trial with parallel group design. SETTING: Ten general practices located in London and the South Thames region of the United Kingdom recruited patients to the trial between 1996 and 1998. Patients came from a wide range of socioeconomic backgrounds and lived in urban, suburban, and rural areas. METHOD: Data were collected before randomisation, after treatment, and six months later. Patients were offered six sessions of up to one hour each of either CBT or counselling. Outcomes include: self-report of fatigue symptoms six months later, anxiety and depression, symptom attributions, social adjustment and patients' satisfaction with care. RESULTS: One hundred and sixty patients with chronic fatigue entered the trial, 45 (28%) met research criteria for chronic fatigue syndrome; 129 completed follow-up. All patients met Chalder et al's standard criteria for fatigue. Mean fatigue scores were 23 on entry (at baseline) and 15 at six months' follow-up. Sixty-one (47%) patients no longer met standard criteria for fatigue after six months. There was no significant difference in effect between the two therapies on fatigue (1.04 [95% CI = -1.7 to 3.7]), anxiety and depression or social adjustment outcomes for all patients and for the subgroup with chronic fatigue syndrome. Use of antidepressants and consultations with the doctor decreased after therapy but there were no differences between groups. CONCLUSION: Counselling and CBT were equivalent in effect for patients with chronic fatigue in primary care. The choice between therapies can therefore depend on other considerations, such as cost and accessibility. PMID- 11271869 TI - Access to complementary medicine via general practice. AB - BACKGROUND: The popularity of complementary medicine continues to be asserted by the professional associations and umbrella organisations of these therapies. Within conventional medicine there are also signs that attitudes towards some of the complementary therapies are changing. AIM: To describe the scale and scope of access to complementary therapies (acupuncture, chiropractic, homoeopathy, hypnotherapy, medical herbalism, and osteopathy) via general practice in England. DESIGN OF STUDY: A postal questionnaire sent to 1226 individual general practitioners (GPs) in a random cluster sample of GP partnerships in England. GPs received up to three reminders. SETTING: One in eight (1226) GP partnerships in England in 1995. METHOD: Postal questionnaire to assess estimates of the number of practices offering 'in-house' access to a range of complementary therapies or making National Health Service (NHS) referrals outside the practice; sources of funding for provision and variations by practice characteristics. RESULTS: A total of 964 GPs replied (78.6%). Of these, 760 provided detailed information. An estimated 39.5% (95% CI = 35%-43%) of GP partnerships in England provided access to some form of complementary therapy for their NHS patients. If all non responding partnerships are assumed to be non-providers, the lowest possible estimate is 30.3%. An estimated 21.4% (95% CI = 19%-24%) were offering access via the provision of treatment by a member of the primary health care team, 6.1% (95% CI = 2%-10%) employed an 'independent' complementary therapist, and an estimated 24.6% of partnerships (95% CI = 21%-28%) had made NHS referrals for complementary therapies. The reported volume of provision within any individual service tended to be low. Acupuncture and homoeopathy were the most commonly available therapies. Patients made some payment for 25% of practice-based provision. Former fundholding practices were significantly more likely to offer complementary therapies than non-fundholding practices, (45% versus 36%, P = 0.02). Fundholding did not affect the range of therapies offered, and patients from former fundholding practices were no more likely to pay for treatment. CONCLUSION: Access to complementary health care for NHS patients was widespread in English general practices in 1995. This data suggests that a limited range of complementary therapies were acceptable to a large proportion of GPs. Fundholding clearly provided a mechanism for the provision of complementary therapies in primary care. Patterns of provision are likely to alter with the demise of fundholding and existing provision may significantly reduce unless the Primary Care Groups or Primary Care Trusts are prepared to support the 'levelling up' of some services. PMID- 11271871 TI - General practitioners' views on the early diagnosis of dementia. AB - This study investigated the views on the early diagnosis of dementia from over 1000 general practitioners (GPs) from 12 Health Authority areas in England and Wales. Almost half of the GPs did not believe it was beneficial to make an early diagnosis, yet most admitted they needed more training in the area. In areas where there had been specific efforts to contact and educate local GPs, the GPs were far more likely to believe in the value of early diagnosis. PMID- 11271870 TI - Ethical and research dilemmas arising from a questionnaire study of psychological morbidity among general practice managers. AB - A questionnaire-based research project enquiring into the psychological health of general practice managers found that 5% of managers admitted to suicidal ideas. This paper explores the moral issues raised when research conducted at a distance uncovers information about participants which indicates that they may be at increased risk of harm. It examines whether the authors of such studies have responsibilities towards their research participants beyond those of analysing and properly interpreting the data supplied to them. The paper is an exercise in self-reflection and self-criticism; not all the questions posed and explored by it can be answered definitively. Implications for planning studies of this kind are discussed. PMID- 11271872 TI - How do general practices manage requests from patients for 'same-day' appointments? A questionnaire survey. AB - Same-day appointment requests are common and can be problematic for general practices that run appointment systems. In a questionnaire survey sent to 90 general practices in one health authority area (response rate 88%), a variety of management systems existed for dealing with same-day appointment requests. Managing the requests was found to be a significant cause of stress for many general practitioners. Registrars, locums, and practice nurses play only a small part in meeting patient demands and few practices operate telephone triage to help manage these requests. PMID- 11271873 TI - The William Pickles Lecture. Issues of risk: 'this unique opportunity'. PMID- 11271874 TI - Ageing Britain--challenges and opportunities for general practice. PMID- 11271875 TI - Sexually transmitted infections in primary care: a need for education. AB - General practitioners and practice nurses require the clinical skills that will enable them to detect sexually transmitted infections in the context of a shift to having no, or insidious symptoms. They need to be able to confirm the diagnosis and have clear models for management and referral. Primary care and genitourinary medicine need to work more closely together to increase mutual understanding and clarify the issues which surround referral and attendance. Sexual health risk assessment through the investigation of sexual history is a helpful way forward in both differential diagnosis and in targeting sexual health promotion and care. Many aspects of these clinical skills are specific to the primary care context. There is a need for improved undergraduate, postgraduate, and in-service training. Multidisciplinary educational approaches are ideal for the subject of sexual health. Primary care groups offer a potential way forward to help develop quality in primary care and some are developing health improvement programmes that aim to address sexual health issues. PMID- 11271877 TI - Roles of President and Chairman of the RCGP. PMID- 11271876 TI - Setting up a database of medical error in general practice: conceptual and methodological considerations. AB - Though common and the cause of much morbidity and health cost, medical error has until recently attracted little attention from primary care workers. A database that logs medical error, operating within the context of clinical governance initiatives at the level of Primary Care Groups, could provide an appropriate framework within which to scrutinise and identify systematic organisational features associated with risk of serious adverse events. This paper discusses some of the key conceptual and methodological issues that need to be resolved before such a database can be implemented in general practice and considers these deliberations in the light of the Chief Medical Officer for England's recent report, An organisation with a memory. PMID- 11271878 TI - Reciprocity. PMID- 11271879 TI - Shared decision-making. PMID- 11271880 TI - Can anyone pass the summative assessment MCQ? PMID- 11271881 TI - NHS National Plan. PMID- 11271882 TI - Evaluating primary care research networks--exposing a wider agenda. PMID- 11271884 TI - Concordance in medicine. PMID- 11271883 TI - Concordance in medicine. PMID- 11271885 TI - To syringe or not to syringe, safety is the question! PMID- 11271886 TI - Counsellors in general practice. PMID- 11271887 TI - Treating depression in primary care. PMID- 11271888 TI - Learning from complementary medicine. PMID- 11271889 TI - Learning from complementary medicine. PMID- 11271890 TI - The future general practitioner. PMID- 11271891 TI - Is it time to review the idea of compliance with guidelines? PMID- 11271893 TI - Commentary. Are current tumour response criteria relevant for the 21st century? PMID- 11271892 TI - Blood pressure control in treated hypertensive patients: clinical performance of general practitioners. AB - BACKGROUND: The blood pressure of many treated hypertensive patients remains above recommended target levels. This discrepancy may be related to general practitioners' (GPs') actions. AIM: To assess clinical performance of GPs in blood pressure control in treated hypertensive patients and to explore the influence of patient and GP characteristics on clinical performance. DESIGN OF STUDY: Cross-sectional study conducted on 195 GPs with invitations to participate made via bulletins and by letter. SETTING: One hundred and thirty-two practices in the southern half of The Netherlands from November 1996 to April 1997. METHOD: Performance criteria were selected from Dutch national hypertension guidelines for general practice. GPs completed self-report forms immediately after follow-up visits of hypertensive patients treated with antihypertensive medication. RESULTS: The GPs recorded 3526 follow-up visits. In 63% of these consultations the diastolic blood pressure (DBP) was 90 mmHg or above. The median performance rates of the GPs were less than 51% for most of the recommended actions, even at a DBP of > or = 100 mmHg. Performance of non-pharmacological actions increased gradually with increasing DBP; prescribing an increase in antihypertensive medication and making a follow-up appointment scheduled within six weeks rose steeply at a DBP of > or = 100 mmHg. Patient and GP characteristics contributed little to clinical performance. Action performance rates varied considerably between GPs. CONCLUSION: GPs seem to target their actions at a DBP of below 100 mmHg, whereas guidelines recommend targeting at a DBP of below 90 mmHg. PMID- 11271894 TI - Effects of radiographic contrast media on proliferation and apoptosis of human vascular endothelial cells. AB - The aim of the study was to determine the effects of radiographic contrast media (RCM) on proliferation and apoptosis of human vascular endothelial cells. Human umbilical vein endothelial cells (HUVECs) were exposed for either 1 min or 15 min to RCM (diatrizoate, ioxaglate, iopromide, iotrolan) at an iodine concentration of 250 mgl ml-1. Controls were complete growth medium (CGM) and saturated mannitol (osmotic control). [3H]thymidine incorporation was used to determine cell proliferation 24 h after exposure. Apoptosis was determined at 1 h and 6 h by terminal uridine nick end labelling (TUNEL), time lapse video microscopy (TLVM) and DNA electrophoresis. Mean proliferation rates (%) (+/- SEM) (p-values compared with the CGM control) at 1 min and 15 min, respectively, were: diatrizoate: 31.9 (10.6), 5.8 (1.5) (p < 0.001); ioxaglate: 48.4 (10.9), 20.4 (4.5) (p < 0.001); iopromide: 63.4 (8.7), 58.2 (10.2) (p < 0.05); iotrolan: 84.7 (7.3), 72.8 (12.4) (p = ns); saturated mannitol 50.5 (9.6), 45.9 (10.0) (p < 0.001). Mean apoptotic indices (%) (+/- SEM) at 1 h and 6 h following 1 min exposure, respectively, were: CGM: 0.25 (0.13), 0.23 (0.08); diatrizoate: 2.18 (0.19), 2.69 (0.34) (p < 0.001); ioxaglate: 1.90 (0.23), 1.69 (0.02) (p < 0.05); iopromide: 0.59 (0.04), 0.33 (0.02) (p = ns); iotrolan: 0.30 (0.07), 0.27 (0.1) (p = ns); saturated mannitol 2.11 (0.24), 1.4 (0.1) (p < 0.05). After 15 min exposure, apoptosis rates at both 1 h and 6 h, respectively, were: iotrolan: 0.29 (0.17), 0.51 (0.16) (p = ns); diatrizoate: 3.19 (0.81), 11.66 (1.75) (p < 0.001); ioxaglate: 1.88 (0.14), 2.87 (0.20) (p < 0.05); iopromide: 1.06 (0.11), 1.52 (0.15) (p < 0.05); saturated mannitol 1.62 (0.09), 4.63 (0.74) (p < 0.05). TLVM and DNA electrophoresis confirmed the occurence of apoptosis after exposure to RCM. In conclusion, saturated mannitol and all tested RCM, with the exception of iotrolan, (diatrizoate > ioxaglate > iopromide) reduced proliferation and increased apoptosis of HUVECs. The effects were more pronounced with ionic RCM and seem to depend on osmolality as well as the chemical structure of these agents. Endothelial injury and apoptosis may be responsible for some of the side effects associated with intravascular use of RCM. PMID- 11271895 TI - Increased hepatic arterial blood flow after decreased portal supply to the liver parenchyma owing to intrahepatic portosystemic venous shunt: angiographic demonstration using helical CT. AB - This study was conducted to investigate the haemodynamics of the liver parenchyma in the presence of intrahepatic portosystemic venous shunt. 3 patients with intrahepatic portosystemic venous shunts and 24 patients with normal intrahepatic haemodynamics underwent both CT arterial portography and CT during hepatic arteriography. Angiographic findings with helical CT were compared, and CT attenuated values were measured in both groups. The liver parenchyma on CT arterial portography had lower attenuation than on CT during hepatic arteriography in all patients with intrahepatic portosystemic venous shunts. Overall average CT attenuation was 92.2 +/- 7.7 Hounsfield units (HU) on CT arterial portography and 149.9 +/- 8.5 HU after CT during hepatic arteriography, with the opposite findings in all patients without intrahepatic portosystemic venous shunt: CT attenuation 142.0 +/- 25.7 HU on CT arterial portography and 100.7 +/- 16.4 HU after CT during hepatic arteriography. In conclusion, the portal venous supply to the liver parenchyma decreased due to intrahepatic portosystemic venous shunts, with a compensatory increase in hepatic arterial blood supply. PMID- 11271896 TI - Crohn's disease of aphthous type: serial changes in intestinal lesions. AB - In Crohn's disease (CD), aphthous lesions are regarded as possible precursors of typical intestinal involvement. To determine the natural course of intestinal lesions in CD of aphthous type, the clinical course of 10 patients was retrospectively investigated during a period ranging from 6 to 16 years after diagnosis. The criterion for inclusion was confirmed aphthous lesions within the gastrointestinal tract with histologically verified epithelioid granuloma. The degrees of aphthous lesions in the small intestine and the colon were graded by small bowel radiography, barium enema examination and colonoscopy. Five patients developed typical CD during a period ranging from 0.8-3.3 years. The site of involvement was the ileum in three patients, the colon in one patient and both the ileum and the colon in one patient. Typical small intestinal CD occurred in four of seven patients with marked aphthous lesions of the small intestine, whereas colonic CD occurred in two of eight patients with such aphthous lesions of the colon. These findings suggest that CD of aphthous type is not necessarily a precursor of clinically overt disease. This may especially be the case for colonic aphthous lesions. PMID- 11271897 TI - The influence of clinical information on the reporting of CT by radiologists. AB - The aim of the study was to determine whether clinical information alters the CT report. This prospective blinded study consisted of 50 consecutive patients who attended a Department of Radiology for CT. Each study was interpreted by two of three consultant radiologists, before and after knowledge of the clinical information. 19 reports were changed after clinical information was known. Clinical follow-up was available in 15 cases. In ten cases the reports were more accurate after clinical information and in five cases the reports were less accurate. In three of the five cases where accuracy was reduced, the clinical information was incorrect. It was concluded that clinical information affects the CT report. If the information is accurate it has a beneficial effect; if it is inaccurate it has a detrimental effect. The more complex the investigation, the more important the clinical information. There was a correlation between readers regarding the influence of clinical information. Correct clinical information therefore improves the radiology report. It is the responsibility of the clinician to provide this information in an accurate and legible form. PMID- 11271898 TI - Influence of anode/filter material and tube potential on contrast, signal-to noise ratio and average absorbed dose in mammography: a Monte Carlo study. AB - The comparative performance of mammographic X-ray systems that use different anode/filter combinations has been assessed for screen-film and digital imaging. Monte Carlo techniques have been used to calculate average glandular dose as well as contrast and signal-to-noise ratio for imaging two test details. Five anode/filter combinations have been studied to establish the potential for dose saving or image quality improvement. For screen-film mammography, it was found that little benefit is gained by changing from a standard 28 kV molybdenum/molybdenum spectrum for breasts up to 6 cm thick. For thicker breasts, where the tube potential for the standard technique might be increased, 20% improvement in contrast can be achieved without dose penalty using molybdenum/rhodium or rhodium/rhodium spectra, whereas dose savings of more than 50% can be attained whilst maintaining contrast using tungsten/rhodium or rhodium/aluminium spectra. In digital mammography, a molybdenum/molybdenum spectrum delivers the lowest dose for a 2 cm breast, but gives the highest dose for thicker breasts. Tungsten/rhodium or rhodium/aluminium spectra provide the lowest doses at greater thicknesses. It is concluded that for screen-film mammography, molybdenum/molybdenum is the spectrum of choice for all but the thickest or most glandular breasts. In digital mammography, an alternative spectrum is preferable for breasts thicker than 2 cm. PMID- 11271899 TI - Image quality and dose in film-screen magnification mammography. AB - Extended exposure times in magnification mammography are a result of the reduced X-ray tube currents required for a small focal spot. The consequences of this are the potential for reduced image quality through motion blur during exposure as well as the onset of film reciprocity law failure. Previous investigators have suggested increasing the X-ray tube potential as a practical mechanism for reducing exposure times in magnification mammography and have demonstrated negligible image quality degradation at least up to 32 kVp. This paper describes a film-screen magnification mammography study that expands upon this previous work to investigate the magnitude of the reduction of breast mean glandular dose and exposure time and the changes in subjective image quality (visibility of low contrast details in an RMI 152 phantom) with increases in tube potential between 28 kVp and 35 kVp. Measures of changes in the radiographic contrast and in the scatter-to-primary ratio (SPR) in magnification geometry as a function of tube potential were also obtained. Evidence for reciprocity law failure was also assessed. For a constant film optical density, increasing the X-ray tube potential from 28 kVp to 35 kVp reduced the mean glandular dose from 3.9 mGy to 2.7 mGy and reduced the exposure time from 3.2 s to 1.0 s. Over this range, the detection rate of fibrils and microcalcification-mimicking specks did not vary with tube potential at the 0.05 level of significance. It was found that only the low contrast mass detail detection rate at 35 kVp was significantly less than that at 28 kVp. The measured radiographic contrast decreased with tube potential and the SPR increased with tube potential. However, both changes were weak, and linear regressions determined that the 95% confidence intervals of the slopes relating both contrast and SPR with tube potential encompassed zero. It is concluded that magnification mammography performed at 34 kVp yields significant reductions in exposure time and mean glandular dose, with a detail detection capability similar to that at 28 kVp. PMID- 11271900 TI - Morphological study of the femur in osteopetrotic (op/op) mice using microcomputed tomography. AB - Osteopetrosis is an inherited metabolic disorder in which normal bone remodelling is inhibited. Its primary cause is considered to be disruption of the functional balance between osteoblasts and osteoclasts as well as a reduction in the quantity of osteoclasts. The purpose of this study was to observe morphological characteristics of the femur in osteopetrotic (op/op) mice, in which the pathogenic mechanism of osteopetrosis has been shown to operate, using microcomputed tomography (micro-CT), in addition to macro-anatomical observations. Previously, micro-CT has been used mainly for morphometry of a small portion of cancellous bone. However, with recent improvements it has become possible for three-dimensional (3D) observations of bone using slightly larger samples to be made. In this study, the accuracy of reconstructed 3D images of the femur produced by micro-CT was confirmed by comparison with the original femur. In addition, an arbitrary cross-section could be displayed on the screen for detailed examination of its internal structure, and the volume percentage of trabecular bone in a particular region of interest could be measured in three dimensions. The results of this study revealed that the femur was smaller and malformed in the op/op mouse compared with the controls, and the differences were greater at the age of 18 weeks than at 5 weeks. In the control mice, bone marrow occupied a large space in the centre of the body of the femur, whereas this area was occupied by calcified bone tissue in the mutant mice. Moreover, when 3D bone density was measured in the region of interest, the value was greater in the mutants than in the controls at both 5 weeks and 18 weeks of age. This study also showed that micro-CT can be applied to 3D morphometric analyses. PMID- 11271901 TI - A study of the application of paediatric reference levels. AB - Radiation exposure of paediatrics is of particular concern because of the greater health detriment. In this study, the application of patient dose reference levels to paediatric radiographic examinations has been investigated. The relationships between entrance surface dose and patient age and size parameters have been studied in three hospitals. The data have been used to derive conversion factors to describe relationships between doses for children of different ages. The usefulness of equivalent patient diameter, weight and age as variables relating to doses has been examined. Simple conversion factors in look-up tables have been derived that link doses for patients of a variety of ages and sizes for particular examinations. It is proposed that factors of this type could be applied to data for individual patients to allow a wider range of ages to be included in any group. This would enable sufficient dose data to be collected from non-specialist hospitals for comparison with reference levels. It would facilitate identification of hospitals where doses are higher so that changes could be made to radiographic practice. PMID- 11271902 TI - Evaluation of combined therapy with chemoembolization and irradiation for large hepatocellular carcinoma. AB - The effects of combined transcatheter arterial chemoembolization (TACE) and radiotherapy in patients with large hepatocellular carcinoma (HCC) were analysed retrospectively. A total of 107 patients with large unresectable HCC was treated with TACE followed by external beam irradiation. The largest dimension of the tumours ranged from 5 cm to 18 cm. Acute effects, survival rates, toxicity and prognostic factors were analysed. Follow-up ranged from 4 months to 98 months (median 24 months). An objective response, i.e. reduction of tumour area greater than 50%, was achieved in 48.6% of cases. In 64.9% of the cases with increased alpha-feto protein (AFP) values, AFP level underwent a reduction of more than 25%. The cumulative survival rates at 1, 3 and 5 years were 59.4%, 28.4% and 15.8%, respectively (median survival 18 months). The combination therapy was generally well tolerated. Only two patients died from liver failure or variceal bleeding associated with therapy. The Cox proportional hazards model showed that the number of tumours and the irradiation dose were independent prognostic factors. The results indicate that combined TACE with radiotherapy is a promising therapeutic approach for large unresectable HCC. Prospective controlled trials to ascertain the real potential benefit of this approach are required. PMID- 11271903 TI - Fluid collections detected by ultrasound following uncomplicated colorectal surgery. AB - The aim of the study was to assess the incidence and site of intraperitoneal fluid collections following uncomplicated colorectal surgery and to identify factors relating to the presence of such collections. 38 patients (22 males) with a mean age of 67 years (range 38-85 years) undergoing uncomplicated colorectal procedures were studied prospectively. Patients underwent abdominal and pelvic ultrasound on Day 3 and Day 7 following surgery. The number, site and volume of collections were recorded. Ultrasound-detected fluid collections were present in 26% on Day 3 and 25% on Day 7 following laparotomy. The presence of a collection was not related to the amount of residual volume after peritoneal lavage with normal saline prior to operative closure, to intraoperative blood loss or to the presence of drains. The right upper quadrant was the commonest site of intraperitoneal collections. In the absence of additional clinical signs, the presence of such collections is not an indication for intervention. PMID- 11271904 TI - Gas exchange parameters in radiotherapy patients during breathing of 2%, 3.5% and 5% carbogen gas mixtures. AB - The gas mixture carbogen may be breathed by patients to enhance the oxygenation level and therefore the radiosensitivity of tumours. However, owing to the high CO2 content, its inhalation is associated with patient intolerance. Our aim was to determine a suitable carbon dioxide and oxygen gas mixture with similar enhancement of arterial oxygenation to 5% carbogen and with improved patient tolerance. 14 patients entered the study; of those 14, 8 were able to tolerate 2%, 3.5% and 5% carbogen mixtures as well as a control gas for sufficient time to allow successful arterial blood gas sampling. Gas exchange parameters were measured using a carbon dioxide monitor and a blood gas analyser. Arterial carbon dioxide tension ranged from 2.9 kPa to 6.82 kPa whilst breathing the carbogen mixtures, and arterial oxygen tension increased at least three-fold from basal values. There were no significant changes in the respiratory rate, heart rate and blood pH. The results suggest that 2% CO2 in O2 enhances arterial oxygen levels to a similar extent as 3.5% and 5% CO2 and that it is well tolerated. PMID- 11271905 TI - Massive intrathoracic haemorrhage after CT-guided lung biopsy. AB - CT-guided lung biopsy is now widely performed for tumorous lesions in the lung, and both its usefulness in this context and the associated complications have been well described in the literature. Although severe complications are rare, we describe a case in which massive intrathoracic haemorrhage developed after lung biopsy and necessitated emergency operation for control. Intraoperative findings suggested that the source of the haemorrhage was a fibrous, cord-like substance present at the site of adhesion associated with old tuberculosis. We attributed this haemorrhage to a pneumothorax, which developed after lung biopsy and caused the new vessels penetrating the centre of the fibrous, cord-like substance to stretch and rupture. Numerous cases have been reported of spontaneous haemopneumothorax precipitated by spontaneous pneumothorax and resulting from the rupture of such vessels. PMID- 11271906 TI - Fatal lipid embolism following intraarterial angiography at an early stage of arteriosclerosis. AB - Intraarterial angiography was performed on a patient with peripheral arterial occlusive disease (Fontaine IIb). No relevant risk factors were known, and a previous angiography had been undertaken without incident. After administration of contrast medium, the patient complained of acute pain in the lower abdomen and both legs, and a sudden rise in blood pressure was observed. The patient subsequently lost consciousness and died within 1.5 h. Postmortem examination showed that death was due to peripheral atheromatous microembolism of lipids, and not cholesterol as is usual in these cases. The differential diagnosis is discussed and a review of the literature is presented. PMID- 11271907 TI - Fatal haemorrhagic myocarditis secondary to cyclophosphamide therapy. AB - Haemorrhagic myocarditis is a rare but important complication of cyclophosphamide therapy. Echocardiographic identification of the disorder can be made. We believe that the ultrasound features of this disorder have not been previously reported. PMID- 11271908 TI - Liver involvement in hereditary haemorrhagic telangiectasia: assessment with 99Tcm-phytate radionuclide angiography and 123I-IMP transrectal portal scintigraphy. AB - Abnormal hepatic haemodynamics and function in a 43-year-old woman with hereditary haemorrhagic telangiectasia (HHT) were evaluated using 99Tcm-phytate angiography and iodine-123-iodoamphetamine transrectal portal scintigraphy. Radionuclide angiography demonstrated hyperdynamic perfusion of the liver owing to intrahepatic arteriovenous fistulae (AVF), entry of tracer into the systemic circulation through intrahepatic portosystemic shunts and an increase in recirculating blood flow caused by these vascular disorders. Heterogeneous distribution of tracer also suggested the presence of chronic hepatic injury. Transrectal portal scintigraphy showed large portosystemic shunts. Other imaging techniques confirmed the presence of the AVF but failed to identify the portosystemic shunts. Non-invasive radionuclide studies are helpful in the evaluation of hepatic involvement of HHT. PMID- 11271909 TI - MR appearances of the locked knee. AB - This review illustrates the MR appearances of commonly encountered problems that can present as a "locked knee", as well as several unusual causes. Internal derangement of menisci, particularly bucket handle tears, predominate. Loose bodies as a result of trauma/degeneration and lesions such as cysts of the cruciate ligaments and focal pigmented villonodular synovitis are also illustrated. While meniscal tears are the major cause of "locked knee" in clinical practice, interference with normal knee kinematics is non-specific with regard to the diagnosis. Emphasis is therefore given to less frequently seen abnormalities that lead to a mechanical block of knee extension. PMID- 11271910 TI - Case of the month. TB or not TB? PMID- 11271911 TI - MRI screening for acoustic neuroma. PMID- 11271912 TI - Judgement postponed in misconduct case. PMID- 11271913 TI - Communication skills: the case for early training. PMID- 11271914 TI - Correlations of measurements of subclinical claw horn lesions in dairy cattle. AB - Measurements were made of the extent of sole and white line lesions on the claws of 115 Holstein-Friesian cows on at least three and at most 16 occasions, and some cows were followed up to their third lactation. All the measurements were made between 12 weeks before calving and 45 weeks after calving. In total, 1016 repeated observations were made. Correlations were calculated between pairs of claws, between types of lesion (sole and white line), and between pairs of the different measurements (number of lesions, proportion of the claw affected, maximum severity score and proportion of the claw affected weighted for severity). The outer hind claws had the greatest extent of lesions of both types. Spearman correlation coefficients and confidence intervals measured the strength of the association. All the associations between claws were positive, suggesting that the lesions did not occur in isolation. Sole and white line lesions were not associated at individual observation points. Lesions on the left and right claws were markedly similar, except for sole lesions on the two inner hind claws, and for white line lesions on the two outer hind claws. PMID- 11271915 TI - Efficacy of moxidectin 2 per cent gel against naturally acquired strongyle infections in horses, with particular reference to larval cyathostomes. AB - The efficacy of moxidectin 2 per cent equine gel against naturally acquired strongyle infections was assessed in 18 ponies which had grazed on contaminated pasture before being housed for eight weeks. Twenty-four hours before the treatment, two randomly selected ponies were euthanased and their worm burdens were determined. Eight of the remaining 16 ponies were treated with moxidectin 2 per cent gel while the other eight were given a placebo gel. Eight weeks later the 16 animals were necropsied and their worm burdens established. A 100 per cent efficacy was recorded against adult and lumenal L4 cyathostomes and adult Strongylus and Triodontophorus species. Digest recoveries of larval cyathostomes indicated a 90.8 per cent (P<0.002) reduction in early L3 and a 99.9 per cent (P<0.001) reduction in developing stages. There was a reduction in faecal egg output of between 96 and 100 per cent in the treated animals compared with the controls. PMID- 11271916 TI - Sick sinus syndrome in nine West Highland white terriers. AB - Sick sinus syndrome is a clinical term used to describe the clinical signs of sinus node dysfunction. This paper describes the clinical data from nine West Highland white terriers, eight females and one male, in which a diagnosis of sick sinus syndrome was made. The most common clinical signs were episodic weakness and presyncope. Electrocardiographic findings included sinus bradycardia, sinus arrest with or without escape complexes, disturbances of atrioventricular conduction, paroxysmal supraventricular tachycardia, or some combination of these dysrhythmias. The main radiographic changes were mild right-sided cardiomegaly in five cases, and a slight increase in bronchial and interstitial markings in four, but there was no evidence of congestive heart failure in any of the dogs. Echocardiography revealed mild to moderate mitral endocardiosis in three cases with no other significant abnormalities. The dogs' responses to parenteral atropine were variable and were not necessarily related to their response to oral anticholinergic agents. Five of the dogs were initially treated with propantheline bromide, but in only two of them were the clinical signs controlled in the long term. Six of the dogs were successfully treated by the implantation of a transvenous pacemaker. PMID- 11271917 TI - Osteoblastic osteosarcoma in a fattening pig. PMID- 11271918 TI - Properties of a new CDV isolate from a raccoon dog (Nyctereutes procyonoides viverrinus) in Japan. PMID- 11271919 TI - Outbreak of contagious ecthyma in camels in Israel. PMID- 11271920 TI - Closure of the Thurso veterinary investigation centre. PMID- 11271921 TI - Bulk tank milk failures. PMID- 11271922 TI - The farming crisis and animal welfare. PMID- 11271923 TI - Gastrointestinal parasites in ostriches (Struthio camelus). PMID- 11271924 TI - Fleas, hosts and locations. PMID- 11271926 TI - Electronic training devices for dogs. PMID- 11271925 TI - Angiostrongylus vasorum in a dog in Surrey. PMID- 11271927 TI - The year 2000. Looking back and looking forward. PMID- 11271928 TI - Eye tracking disturbances in schizophrenia. AB - PURPOSE: To study the frequency of different types of eye tracking disturbances in schizophrenia. MATERIALS AND METHODS: Smooth pursuit eye movements were studied by electro-oculography (EOG) in 22 schizophrenic patients (ICD-10 criteria) and 15 age and sex-matched controls. The studied parameters included average pursuit gain, number of saccades, the frequency of different types of saccades (catch-up, back-up, anticipatory saccades), and disturbances during fixation. The results were analysed statistically. RESULTS: The average pursuit gain was significantly affected in patients for target velocity of 30 degrees/sec (p = 0.007). The catch-up and back-up saccades were more common in cases than controls but the difference was not significant (p = 0.39 and 0.36 respectively). The anticipatory saccades were significantly more frequent in cases than controls (p < 0.0001) for both 15 degrees/sec and 30 degrees/sec target velocities. This was also correlated with the duration of illness. CONCLUSION: Anticipatory saccades are significantly more frequent during eye tracking in schizophrenia and appear to be an objective marker for the disease. PMID- 11271929 TI - Inferior limbal-conjunctival autograft transplantation for recurrent pterygium. AB - PURPOSE: To study the safety and efficacy of inferior limbal-conjunctival autograft (LCAT) transplantation in the surgical management of recurrent pterygium. METHODS: Prospective non-comparative case series. Inferior limbal conjunctival autografting was performed on 11 patients (11 eyes) with recurrent pterygium. Pterygium recurrence was considered a surgical failure. RESULTS: Recurrence of pterygium was noted in two (18.2%) eyes, after a mean follow up of 16.2 +/- 0.9 months (range: 10-19 months). Neither recurrence required further surgical treatment. Nonprogressive pseudopterygium formation was noted at the donor site in five (45.5%) eyes. CONCLUSION: Inferior LCAT appears to be a safe and effective option in the management of recurrent pterygium. In patients with suspected or proven glaucoma, this may be the procedure of choice, if mitomycin C is contraindicated. PMID- 11271930 TI - Tonometry in normal and scarred corneas, and in postkeratoplasty eyes: a comparative study of the Goldmann, the ProTon and the Schiotz tonometers. AB - PURPOSE: Clinical comparison of intraocular pressure (IOP) measured with the Goldmann applanation tonometer (GAT), the ProTon tonometer (PT), and the Schiotz tonometer (ST), in normal eyes, eyes with scarred corneas and postkeratoplasty eyes. MATERIAL AND METHODS: The IOP readings with GAT, PT, and ST were compared in 125 eyes with normal corneas (Group A), 17 eyes with scarred corneas (Group B), and in 21 postkeratoplasty eyes (Group C). The data were statistically analysed at 95% confidence interval; linear regression analysis and paired t-test were done. RESULTS: The mean differences and their standard deviation [SD] between GAT and PT readings, and GAT and ST readings respectively were: [1] in Group A: -0.23 [SD 2.75] mmHg and +0.24 [SD 3.18] mmHg respectively; [2] in Group B: -1.8 [SD 12.67] mmHg and -4.5 [SD 9.95 mmHg; and [3] in Group C: +0.24 [SD 8.72] mmHg and -0.12 [SD 8.7] mmHg. They were not statistically significant. In Group A the 95% confidence interval between GAT and PT readings was -5.27 mmHg to 5.73 mmHg, and between GAT and ST readings, -6.12 mmHg to 6.59 mmHg. Ninety six [77%] eyes with the PT and 84 [69%] eyes with ST measurements were within 3 mmHg of GAT pressure. The correlation coefficients [r] for PT and ST were 0.93 [P = 0.0000] and 0.88 [P = 0.0000] respectively. In Group B 95% confidence interval between GAT and PT readings was -27.17 mmHg to 23.51 mmHg, and between GAT and ST measurement, -24.37 mmHg to 15.44 mmHg. The correlation coefficients [r] for the PT and ST were 0.112 [P = 0.660] and 0.630 [P = 0.006] respectively. In group C, the 95% confidence interval between GAT and PT measurements was -17.20 mmHg to 17.67 mmHg, and between GAT and ST measurements, -17.51 mmHg to 17.27 mmHg. The correlation coefficients [r] for the PT and the ST were 0.780 [P = 0.0000] and 0.740 [P = 0.0001] respectively. CONCLUSIONS: In clinical practice PT appears to have a higher level of accuracy than ST in normal corneas. In scarred corneas and post-penetrating keratoplasty eyes, because of high SD for mean differences and wide confidence interval of 95%, both PT and ST are inaccurate in measuring IOP as compared to GAT in such eyes. PMID- 11271931 TI - VISION 2020. The right to sight. PMID- 11271932 TI - Intraocular cilia associated with perforating injury. AB - PURPOSE: To report a case series of penetrating injury complicated by occurrence of intraocular cilia. METHODS: Retrospective analysis of charts of 11 eyes of 11 patients with penetrating injury and intraocular cilia, presenting between September 1978 and November 1998. Ten eyes underwent surgery for trauma-related problems such as cataract, vitritis, retinal detachment etc., at which time intraocular cilia were removed. One eye did not have surgery and continues to harbour cilia at the posterior perforation site. RESULTS: Metallic wire was responsible for injury in 6 of 11 eyes with intraocular cilia. Five eyes had significant intraocular inflammation. The cilia were located in the anterior segment in 4 eyes; in the posterior segment in 6 eyes and in both in one eye. At the last follow up, 72.7% had 6/18 or better vision. Poor vision in the rest was due to recurrent retinal detachment (2 eyes) and macular scarring (1 eye). CONCLUSION: Intraocular cilia are more commonly associated with injury by a metallic wire. The presentation and management of an injured eye does not seem to be influenced by the presence of cilia in the eye. PMID- 11271933 TI - Possible role of polyamines in gyrate atrophy. AB - PURPOSE: Gyrate atrophy (GA) is marked by hyperornithinemia and lowered ornithine amino transferase (OAT). However there are patients of GA without hyperornithinemia and those with hyperornithinemia without GA. Some cases of GA have been reported to have low lysine. The purpose of the study was to determine if polyamines, the metabolites of ornithine, and lysine have any diagnostic role in GA. METHODS: Ornithine in plasma was estimated by two-dimensional paper chromatography, with elution of the coloured spot, and the absorbance measured using a spectrophotometer at 560 nm. OAT assay in lymphocytes was done spectrophotometrically using ornithine as substrate. Blood and urinary polyamines were extracted with n-butanol, benzoylated and analysed with HPLC; putrescine, spermine, spermidine, and cadaverine were assayed individually at 254 nm with the UV detector using ODS, G18 column with 63% methanol as solvent. RESULTS: Of the 7 patients investigated, 6 had features typical of GA. One was diagnosed to have atypical retinitis pigmentosa (case 3). The first five cases had elevated ornithine and diminished OAT, but cases 6 and 7 had near-normal ornithine and case 7 had near-normal OAT. However, all 7 patients had increased levels of total polyamines in urine compared to normals. Five had increased putrescine and three had increased spermine. All the 7 had decreased cadaverine in urine. Thus, though there were inconsistencies with ornithine and OAT, all the 7 patients had elevated polyamines from ornithine and decreased cadaverine. CONCLUSION: In addition to estimating ornithine and OAT in GA, it is suggested that urinary polyamines may be analysed as the latter appears to correlate better with the clinical condition and help in the diagnosis to a greater extent. Moreover, while ornithine is an innocuous amino acid, polyamines are known to damage DNA and proteins. PMID- 11271935 TI - Open angle glaucoma as a manifestation of Waardenburg's syndrome. AB - Waardenburg's syndrome is a rare, autosomal dominant disorder, with several clinical signs, each with variable penetrance. We report this case of Waardenburg's syndrome with bilateral open-angle glaucoma with unique gonioscopic findings. PMID- 11271934 TI - Papilloedema with peripapillary retinal haemorrhages in an acquired immunodeficiency syndrome (AIDS) patient with cryptococcal meningitis. AB - A case of cryptococcal meningitis in an AIDS patient who presented with optic disc edema, bilateral retinal and peripapillary haemorrhages is reported. PMID- 11271936 TI - Molecular genetics of cataract. AB - Studies on hereditary congenital cataracts have led to the identification of genes involved in formation of these cataracts. Knowledge of the structure and function of a particular gene and the effect of disease-associated mutations on its function are providing insights into the mechanisms of cataract. Identification of the disease gene requires both the relevant clinical data as well as genetic data on the entire pedigree in which the disease is found to occur. Genes for hereditary cataract have been mapped by genetic linkage analysis, in which one examines the inheritance pattern of DNA markers throughout the genome in all individuals of the pedigree, and compares those with the inheritance of the disease. Cosegregation of a set of markers with disease implies that the disease gene is present at the same chromosomal location as those markers. The genes so far identified for hereditary cataracts in both humans and animal models encode structural lens proteins, gap junction proteins, membrane proteins and regulatory proteins involved in lens development. Understanding of the mechanisms of hereditary cataract may also help us understand the manner in which environmental and nutritional factors act on the lens to promote opacification. PMID- 11271937 TI - Micrococcal endophthalmitis following extracapsular cataract extraction with foldable silicone intraocular lens implantation. AB - A case of postoperative endophthalmitis caused by micrococci, after phacoemulsification and foldable silicone intraocular lens (IOL) implantation is reported. PMID- 11271938 TI - Ascaris lumbricoides in the lacrimal passage. AB - Ascariasis is caused by the roundworm, Ascaris lumbricoides. We report an additional case of live Ascaris lumbricoides removed from the lacrimal puncta of a 10-year-old boy. PMID- 11271939 TI - Kearns Sayre syndrome: an atypical presentation. AB - Kearns Sayre syndrome is a rare presentation which usually involves a triad of factors: external ophthalmoplegia, retinal pigmentary degeneration, and heart block. We present a clinically and histopathologically confirmed case of Kearns Sayre syndrome that involved no retinal pathology. PMID- 11271940 TI - A rare case of both eyelids swelling: isolated conjunctival amyloidosis. AB - Amyloid is an eosinophilic, amorphous protein that has been reported to deposit in virtually any tissue or organ and when extensive, may attain tumourous proportions. We present a rare case where both the upper and lower palpebral conjunctiva were affected by amyloid deposition. PMID- 11271941 TI - Unilateral visual impairment in an urban population in southern India. AB - This study assessed the prevalence and causes of unilateral visual impairment in the urban population of Hyderabad city as part of the Andhra Pradesh Eye Disease Study. Stratified, random, cluster, systematic sampling was used to select 2,954 subjects from 24 clusters representative of the population of Hyderabad. Eligible subjects underwent detailed eye examination including logMAR visual acuity, refraction, slitlamp biomicroscopy, applanation tonometry, gonioscopy, dilatation, cataract grading, and stereoscopic evaluation of fundus. Automated threshold visual fields and slitlamp and fundus photography were done when indicated by standardised criteria. Unilateral visual impairment was defined as presenting distance visual acuity < 6/18 in the worse eye and > or = 6/12 in the better eye, which was further divided into unilateral blindness (visual acuity < 6/60 in the worse eye) and unilateral moderate visual impairment (visual acuity < 6/18-6/60 in the worse eye). A total of 2,522 subjects (85.4% of eligible), including 1,399 > or = 30 years old, participated in the study. In addition to the 1% blindness and 7.2% moderate visual impairment (based on bilateral visual impairment criteria) reported earlier in this sample, 139 subjects had unilateral visual impairment, an age-gender-adjusted prevalence of 3.8% (95% confidence interval 2.7-4.9%). The major causes of this visual impairment 39.9% were refractive error (42.9%), cataract (14.4%), corneal disease (11.5%), and retinal disease (11.2%). Of this unilateral visual impairment was blindness. The major causes of unilateral blindness were corneal disease (23.2%), cataract (22.5%), retinal disease (18%), and optic atrophy (12.9%). On the other hand, the predominant cause of unilateral moderate visual impairment was refractive error (67%) followed by cataract (9%). Of the total unilateral visual impairment, 34.3% was present in those < 30 years old and 36.2% in those 30-49 years old. Unilateral visual impairment afflicts approximately 1 in 25 persons in this urban population. A large proportion of this unilateral visual impairment is present in younger age groups. The causes of unilateral visual impairment, like those of bilateral visual impairment in this population, are varied, suggesting therefore, that in addition to the current focus of eye care in India predominantly on cataract, other causes of visual impairment need to be addressed as well. PMID- 11271942 TI - Adaptation of WHOQOL as health-related quality of life instrument to develop a vision-specific instrument. AB - The WHOQOL instrument was adapted as a health-related QOL instrument for a population-based epidemiologic study of eye diseases in southern India, the Andhra Pradesh Eye Disease Study (APEDS). A follow-up question was added to each item in WHOQOL to determine whether the decrease in QOL was due to any health reasons including eye-related reasons. Modifications in WHOQOL and translation in local language were done through the use of the focus groups including health professionals and people not related to health care. The modified instrument has 28 items across 6 domains of the WHOQOL and was translated into the local language, Telugu, using the pragmatic approach. It takes 10-20 minutes to be administered by a trained interviewer. Reliability was within acceptable range. This health-related QOL instrument is being used in the population-based study APEDS to develop a vision-specific QOL instrument which could potentially be used to assess the impact of visual impairment on QOL across different cultures and for use in evaluating eye-care interventions. This health-related QOL instrument could also be used to develop other disease-specific instruments as it allows assessment of the extent to which various aspects of QOL are affected by a variety of health problems. PMID- 11271943 TI - Is there a minimum endothelial cell count for a clear cornea after penetrating keratoplasty? PMID- 11271944 TI - Predictors of failure of transoesophageal cardioversion of common atrial flutter. AB - BACKGROUND: Common atrial flutter is due to a re-entry circuit in the right atrium. It is possible to entrain and interrupt this arrhythmia with transoesophageal pacing (TEAP) in a substantial percentage of patients. The aim of this study is to evaluate factors associated with failure of transoesophageal cardioversion of common atrial flutter. METHODS: One hundred consecutive patients underwent an attempted transoesophageal cardioversion of their common atrial flutter. In order to detect factors associated with failure of this procedure, the following were considered: (a) age and gender; (b) underlying heart disease; (c) time of onset of the arrhythmia; (d) antiarrhythmic treatment at the time of cardioversion; (e) flutter cycle length, (f) A/V deflection ratio at the site of transoesophageal pacing; and (g) longitudinal and transverse diameters of right and left atrium on the echocardiogram. RESULTS: In 84 of 100 patients, TEAP modified the atrial flutter circuit: in 23 of these, sinus rhythm was restored; in 31 patients, flutter was converted into atrial fibrillation which spontaneously reverted to sinus rhythm; and in remaining 30 patients, persistent atrial fibrillation was obtained. In 16 cases, no modification in atrial flutter circuit was obtained by TEAP (Group 2). Using univariate analysis, this group of patients showed no significant difference in flutter cycle length, a smaller A/V ratio at the site of TEAP, a longer transverse diameter of left atrium and a shorter transverse diameter of right atrium. Analysis of the therapy at cardioversion shows that no Group 2 patients was on intravenous amiodarone, while a greater percentage of patients of the former group was on chronic amiodarone treatment. A logistic regression model applied to the data showed that flutter cycle length, transverse diameter of left atrium and A/V deflection ratio at the site of TEAP were independent variables with influence on the failure rate. CONCLUSION: Transoesophageal pacing is able to modify the circuit of common atrial flutter in a large percentage of patients, and can convert this arrhythmia to sinus rhythm in more than 50% of cases. Failure of this procedure is associated with electrophysiological parameters (flutter cycle length, A/V ratio at the site of TEAP), anatomical factors (left and right atrial diameters) and treatment in use at the time of TEAP. PMID- 11271946 TI - Internal cardioversion as a first-line method of cardioversion? PMID- 11271945 TI - The long QT syndrome. PMID- 11271947 TI - How to approach left-sided accessory pathway ablation using intracardiac echocardiography. PMID- 11271948 TI - Limitations of the use of spectral analysis of heart rate variability for the estimation of cardiac sympathetic activity in heart failure. AB - Spectral analysis of heart rate variability has gained popularity as a simple, non-invasive tool for assessing autonomic function in both normal subjects and in patients in a variety of clinical settings. However, the use of this method as a means of estimating the magnitude of cardiac sympathetic activation in individual patients with heart failure has proved disappointing, with a lack of concordance with more direct measures of sympathetic outflow. This review will describe the rationale involved in using sympathetic indices obtained from spectral analysis of heart rate variability to assess cardiac sympathetic outflow in normal subjects and patients with heart failure. The specific limitations and technological concerns that dictate how it may most effectively be used in this patient population will be discussed. PMID- 11271949 TI - Scatterplots of RR and RT interval variability bring evidence for diverse non linear dynamics of heart rate and ventricular repolarization duration in coronary heart disease. AB - OBJECTIVE: QT interval prolongation and increased spatial QT dispersion are important factors increasing the risk in coronary heart disease. The authors studied the spontaneous beat-to-beat variability of ventricular repolarization (RT intervals) in normal subjects and in patients after myocardial infarction (MI) in order to define the determinants of abnormal temporal dispersion. METHODS: Seventy-six patients with a history of MI (17 female, 59 male, aged 52 +/- 10 years) comprised the study group. Forty-seven patients had preserved left ventricular ejection fraction (EF > or = 40%, MI-A) and 29 patients had left ventricular dysfunction (EF < 40%, MI-B). Twenty healthy volunteers (6 female, 14 male, aged 25 +/- 5 years) were included as the control group. An ECG signal of 512 heartbeats was recorded in the supine position. After analogue-to-digital conversion (16 bit, 2 kHz), the fiducial points of the R wave and T wave were determined. The RR and RT variability (V) assessed in the time domain as the standard deviations of RR and RT (ms), as well as the coefficients of scatterplots of RR and RT intervals. RESULTS: As expected, the standard deviation of RR was significantly reduced in MI patients. The magnitude of RTV in the time domain was similar in the controls and in both subgroups of MI patients. The complexity of heart rate variability (HRV) was slightly, but significantly, reduced in the MI-B group, but not significantly in the MI-A heart group. The complexity of RTV behaved in the opposite manner, being increased in both MI subgroups with the lower mean in the MI-B patients. The different behaviour of HRV and RTV was indicated by the increased ratio of RR/RT coefficients, which reached a significantly greater value in the MI-B group. CONCLUSION: The authors have described different patterns of scatterplot of short-term HRV and RTV in normal subjects, which confirmed that RTV is a less complex phenomenon than HRV. In patients after MI, the complexity of HRV diminishes, while the complexity of RTV increases. These opposing changes are more pronounced in patients with left ventricular dysfunction. A possible prognostic value of this feature is unknown and remains to be elucidated in future prospective studies. PMID- 11271950 TI - Safety and efficacy of low-energy cardioversion of 500 patients using two different techniques. AB - AIM: To present some safety and efficacy issues of low-energy internal cardioversion of chronic atrial fibrillation from 500 consecutive procedures performed with two different techniques, using either two single-coil catheters, or a single twin-coil catheter. METHODS AND RESULTS: Low-energy internal cardioversion was carried out in 368 patients by means of two defibrillation catheters: the former was positioned in the right atrium and the latter either in the left pulmonary artery (212 patients), or in the distal coronary sinus (156 patients). In the remaining 132 patients, a single twin-coil catheter was positioned with the distal coil either in the pulmonary artery (75 patients) or in the coronary sinus (57 patients), while the proximal coil was in the right atrium. The external defibrillator delivered truncated biphasic shocks (6/6 ms, tilt 50%), with a voltage of 10-400 V. In 283 patients (57%) external cardioversion had been unsuccessfully tried before low-energy internal cardioversion. After a total of 1118 shocks, the overall success rate was 92.2% (91.3% with two catheters and 94.7% with the single catheter); the success rate was 93.4 and 91.3% with the coronary sinus and the pulmonary artery approach, respectively. The mean energy used was 6.5 +/- 3.4 J (voltage: 320 +/- 45 V); no difference was found between the twin catheter (6.3 +/- 3.1 J) and the single catheter approach (6.9 +/- 3.7 J), while the coronary sinus configuration required a significantly lower energy than the pulmonary artery configuration (5.6 +/- 2.9 vs 7.2 +/- 3.8 J, P < 0.05). The duration of the current atrial fibrillation episode was the only clinical characteristic statistically different between the 461 successfully cardioverted patients and the 39 failures (295 vs 727 days, P < 0.01). No complication was recorded during or after the delivery of the therapy; no procedure had to be terminated because of patient's intolerance. CONCLUSIONS: Low-energy internal cardioversion is a safe and effective procedure for converting chronic atrial fibrillation, confirmed by this large multicentre experience. The newly available twin-coil catheter seems to achieve a slightly better success rate compared with the traditional two-catheter technique, and is associated with the same safety profile. PMID- 11271951 TI - Optimal atrioventricular delay setting determined by evoked QT interval in patients with implanted stimulus-T-driven DDDR pacemakers. AB - Cardiac function is improved by optimizing the atrioventricular (AV) delay. An automatic optimizing function of AV delay may be necessary to achieve the most favourable haemodynamic state in paced patients. The QT interval may change when cardiac function is improved by optimizing the AV delay. The QT or stimulus-T interval is used as a sensor for rate-responsive pacemakers. Evoked (e) QT interval is measured as the time duration from the ventricular pace pulse (stimulus) and the T-sense point that is the steepest point of the intracardiac T wave (stimulus-T interval). The relationship between AV delay, eQT interval and cardiac function was studied in 10 patients (73 +/- 10 (SD) years old) with an implanted stimulus-T-driven DDDR pacemaker. Cardiac output (CO) and pulmonary capillary wedge pressure (PCWP) were measured by Swan-Ganz catheter. The AV delay was prolonged stepwise by 30 ms. Electrocardiogram event markers which indicated ventricular spike and sensed T wave were recorded, and the interval between two event markers was measured as eQT interval. When AV delay was changed from 240 ms to the AV delay at which CO was maximal (172 +/- 33 ms), eQT interval prolonged from 346 +/- 60 to 353 +/- 62 ms (P < 0.01). There was a significant positive correlation between the optimal AV delay at which CO was maximal (172 +/- 33 ms) and the optimal AV delay which was predicted from the maximum eQT interval (179 +/- 37 ms, r = 0.92, P < 0.001). When AV delay was changed from 240 ms to the predicted optimal AV delay, CO increased from 4.2 +/- 0.7 to 4.5 +/- 0.81.min-1 (P < 0.001) and PCWP was decreased from 7.1 +/- 4.0 to 5.7 +/- 3.1 mmHg (P < 0.05). In conclusion, the optimal AV delay can be predicted from the eQT interval which is sensed by an implanted pacemaker. Automatic setting of the optimal AV delay may be achieved by the QT sensor of an implanted pacemaker. PMID- 11271952 TI - Differences in pacing from the atrial appendage and the lateral atrial free wall on left ventricular filling and haemodynamics during DDD pacing. AB - INTRODUCTION: Atrioventricular sequential pacing involves stimulation from electrodes in the right atrium, generally the atrial appendage (RAA) and the right ventricular apex. The appendage, however, may be unsuitable if a stable position cannot be achieved. The aim of this study was to assess the haemodynamic consequences of different atrial stimulation sites during DDD pacing. METHODS: In 12 consecutive patients (mean age 67 +/- 7 years) who underwent DDD pacemaker implantation, an additional temporary bipolar pacing electrode was positioned on the right atrial free wall. Pacing was performed alternating from the two locations at 85, 100 and 120 beats per minute (bpm). Paced atrioventricular delay was set at 180 ms. Cardiac output and mitral inflow measurements were performed using Doppler echocardiography. RESULTS: Pacing at 85 and 100 bpm resulted in a significantly higher A-peak velocity from the RAA compared with the right atrial free wall. Cardiac index was consistently higher from the RAA location (2.4 +/- 1.2 vs 2.1 +/- 0.91. min-1 m-2 at 85 bpm, 2.71 +/- 1.4 vs 2.35 +/- 1.11. min-1 m 2 at 100 bpm and 2.94 +/- 1.5 vs 2.61 +/- 1.41. min-1 m-2 at 120 bpm, P < 0.05). CONCLUSION: Stimulation from the RAA was superior to stimulation from the right atrial free wall with respect to left ventricular filling and cardiac output. Compared with stimulation from the right atrial free wall, RAA pacing resulted in an increase of 10-15% in cardiac output. PMID- 11271953 TI - Permanent pacemaker implantation via the femoral vein: an alternative in cases with contraindications to the pectoral approach. AB - BACKGROUND: This paper presents a consecutive series of permanent pacemakers (PPM) implanted via the femoral vein in patients with contraindications to pacing systems via the superior vena cava (SCV). The femoral vein approach is a less invasive and feasible alternative to epicardial lead placement. METHODS: Twenty seven patients had femoral pacemakers inserted. Indications for femoral vein pacemaker insertion were: SVC/subclavian obstruction (12 patients, 44.4%), previous infection in SVC leads (four patients, 14.8%), mastectomy and/or radiotherapy to chest (four patients, 14.8%), multiple leads in SVC (two patients, 7.4%), recurrent erosion (two patients, 7.4%), abnormal anatomy (one patient, 3.7%), painful pacemaker pocket (one patient, 3.7%) and burns (one patient, 3.7%). Fifty-one leads, 25 atrial and 26 ventricular, were inserted. The majority of leads were active fixations (96% of atrial leads and 85% ventricular leads). RESULTS: During a mean follow-up of 36.5 months (range 0.9-116.5), six additional unplanned procedures were performed in four patients. Atrial lead displacement occurred in five leads (20%). There were no ventricular lead displacements. In two patients, box revision for pre-erosion was required. One patient had persistent pain at the site of abdominal pacemaker generator. Infection, thromboembolic events, thromophlebitis, evidence of lower limb venous occlusion and lead fracture did not occur. CONCLUSION: Femoral vein PPM are a simple and feasible alternative in patients in whom the SVC approach is contraindicated. PMID- 11271954 TI - Adverse events with transvenous left ventricular pacing in patients with severe heart failure: early experience from a single centre. AB - AIMS: Assessment of complications following implantation of transvenous ventricular electrodes to pace the left ventricle. METHODS AND RESULTS: Twenty eight patients with severe cardiac failure and left bundle branch block were prospectively followed for adverse effects of implantation of a left ventricular transvenous pacing system. Immediate follow-up was associated with loss of left ventricular pacing in nine patients (32%). This was due to lead dislodgement in four cases (corrected by re-operation in three of these cases), and due to increased threshold in five cases (corrected by programming a higher pacing amplitude in all five cases, but with intermittent diaphragmatic contraction in one case). After 1 month, one patient died, one patient with severe coronary heart disease suffered a myocardial infarction, and left ventricular pacing was lost in two patients. Pericardial effusion, new significant ventricular arrhythmias or other adverse effects were not observed. After a mean follow-up of 16 +/- 9.2 months, pacing leads remained stable and no late complications related to the transvenous left ventricular epicardial pacing were observed. CONCLUSION: Placement of a permanent lead in a tributary of the coronary sinus is feasible without serious adverse effects during the first month. The only frequent adverse event was lead dislodgement; a finding which emphasizes the need for development of specially designed leads for this application. PMID- 11271955 TI - Influence of atrial flutter ablation on right to left inter-atrial conduction. AB - AIMS: Ablation of the atrial isthmus between the tricuspid annulus and the inferior vena cava changes P-wave morphology during low lateral right atrial pacing. For better understanding of the mechanism of this alteration, the sequence of activation of the inter-atrial septum and the left atrium were compared before and after ablation of the isthmus between the inferior vena cava and the tricuspid annulus. METHODS AND RESULTS: In 13 patients, left atrial mapping was performed using a duodecapolar electrode catheter advanced to the far distal coronary sinus. The inter-atrial septum was mapped using a right atrial duodecapolar electrode catheter. Conduction times were measured during low lateral right atrial pacing from the pacing artefact and during sinus rhythm from the earliest right atrial electrogram to every intra-cardiac electrogram before and after the ablation. During low lateral right atrial pacing, isthmus ablation resulted in a significant delay in every left atrial lead. Changes were maximal at the posterior aspect of the left atrium and minimal at its anterior aspect. No significant change was discernible on the inter-atrial septum. During sinus rhythm, atrial activations remained unchanged. CONCLUSION: Electrocardiographic changes of P-wave morphology result from alteration in the sequence of left atrial activation rather than that of the inter-atrial septum. PMID- 11271956 TI - Sotalol vs metoprolol for ventricular rate control in patients with chronic atrial fibrillation who have undergone digitalization: a single-blinded crossover study. AB - AIMS: To compare the effects of sotalol and metoprolol on heart rate, during isotonic (ITE) and isometric (IME) exercise and daily activities, in digitalized patients with chronic atrial fibrillation. METHODS AND RESULTS: The study had a randomized, single-blinded, crossover design. Twenty-three patients with chronic atrial fibrillation received placebo for 4 weeks, followed by a 4-week period of treatment with sotalol and metoprolol in random order. At the end of each period, the patients were assessed with 24-h ECG monitoring, a cardiopulmonary exercise test and a handgrip manoeuvre. Both agents produced a lower heart rate than placebo at rest and at all levels of isotonic exercise (P < 0.001) without affecting oxygen uptake. Sotalol produced a lower heart rate than metoprolol only at submaximal exercise (116 +/- 9 bpm for sotalol vs 125 +/- 11 bpm for metoprolol, P < 0.001). During isometric exercise, sotalol produced a lower maximum heart rate than did metoprolol (113 +/- 22 vs 129 +/- 18 bpm, respectively). Both agents produced a lower mean heart rate than placebo over 24 h (P < 0.001 for all), while sotalol produced a lower mean heart rate than metoprolol during the daytime (P < 0.01). CONCLUSION: Sotalol is a safe and effective agent for control of heart rate in digitalized patients with atrial fibrillation. Sotalol is superior to metoprolol at submaximal exercise, resulting in better rate control during daily activities. PMID- 11271957 TI - Avoiding inappropriate ventricular tachycardia detection due to T-wave oversensing in an implantable cardioverter defibrillator: a novel application of the electrogram width criterion. AB - Appropriate sensing is an essential function of an implantable cardioverter defibrillator (ICD). T-wave oversensing by an ICD can be a serious problem in some patients, causing overestimation of the heart rate, inappropriate tachyarrhythmia detection and therapy delivery. Decreasing the sensitivity or programming longer refractory periods can sometimes overcome T-wave oversensing, but these measures may interfere with the ability of the ICD to correctly detect tachyarrhythmias. This report proposes a novel application of the electrogram (EGM) width criterion using a recently introduced detection enhancement algorithm intended to improve the specificity of ventricular tachycardia detection, to avoid T-wave oversensing. Based on the course of a case with persistent T-wave oversensing and review of previously published reports, oversensing problems in ICDs and management strategies are discussed. PMID- 11271958 TI - Psychosocial determinants of perceived vulnerability to harm among adult drinkers. AB - OBJECTIVE: Perceived vulnerability to harm is widely acknowledged as a determinant of behavior change, but little is known about why some drinkers believe that they are personally "at risk" for problems while others do not. This study examined perceived vulnerability to alcohol-related harm in relation to epidemiological risk status on a standardized problem-drinking measure and two psychosocial measures of drinking context: (1) typical reasons for drinking and cutting down and (2) social network influences related to alcohol use. We evaluated the general hypothesis that these psychosocial variables would independently affect perceived vulnerability to alcohol-related harm, over and above epidemiological risk status. METHOD: Adults between the ages of 18 and 79 (N = 430; 249 women, 173 men, 8 gender unknown) completed a questionnaire about drinking behavior and drinking-related social and motivational context. RESULTS: There was a positive relationship between problem-drinking status and perceived risk of experiencing harm, and no support for the idea that objectively "at-risk" drinkers believe that they are less likely to personally experience harm than comparable peers. Drinking motives and social network variables each significantly improved the prediction of perceived vulnerability when epidemiological risk status was controlled. CONCLUSIONS: Interventions designed to alter drinkers' risk perceptions should take into account the reasons that people have for drinking and the social network context of alcohol use, in addition to whether or not individuals are "problem drinkers." PMID- 11271959 TI - Self-reports of physical, sexual and emotional abuse in an alcoholism treatment sample. AB - OBJECTIVE: There is a growing appreciation that emotional, physical and sexual abuse events are frequently part of the life histories of individuals in treatment for alcohol disorders. The present study examines reports of lifetime abuse in a clinical trial for treatment of alcohol dependency. METHOD: Data were obtained from baseline assessments conducted with participants (N = 1,726; 1,307 men) entering Project MATCH, a multisite clinical trial conducted at nine geographically dispersed research sites. Differences on a broad range of participant characteristics were examined by gender and by reported abuse type. RESULTS: More than half (59%) of the participants reported lifetime abuse. Women were more likely (77%) to report abuse than were men (54%). A lower proportion of men than women (6% vs 31%) reported experiencing both physical and sexual abuse. Gender differences were found on the majority of psychosocial measures. Comparisons of the psychosocial measures by abuse type generally indicated that participants without abuse histories had better functioning than did participants reporting abuse. CONCLUSIONS: The high frequency of lifetime abuse in this geographically dispersed sample underscores the necessity for including assessment of emotional, physical and sexual abuse with alcoholism treatment seeking populations. Participants reporting such events may require other treatment in addition to that for alcohol dependency. PMID- 11271960 TI - Alcohol dependence among cocaine-dependent outpatients: demographics, drug use, treatment outcome and other characteristics. AB - OBJECTIVE: Concurrent dependence on alcohol is common among those seeking treatment for cocaine dependence. More information is needed about differences between those with and without concurrent alcohol dependence, including possible special treatment needs or outcome differences. METHOD: Data were obtained from 302 adults (70% men) enrolled in outpatient treatment for cocaine dependence. Individuals who did and those who did not meet criteria for alcohol dependence were compared on demographics, drug use, treatment outcome and other variables. RESULTS: With regard to cocaine use, alcoholics were more likely than nonalcoholics to report an intranasal route of administration, use of cocaine in social settings, more simultaneous use of cocaine and alcohol, and more adverse consequences of their cocaine use. With regard to alcohol use, alcoholics reported consuming alcohol more frequently and in larger amounts, had longer drinking histories and were more likely than nonalcoholics to report increases in alcohol consumption when using cocaine. Alcoholics were heavier cigarette smokers than nonalcoholics and reported more severe employment, legal, family and psychiatric problems. There were overall improvements in both groups from intake through 12 months after treatment. With regard to treatment retention and cocaine abstinence, alcoholics had better outcomes than nonalcoholics when treated with intensive behavioral counseling plus incentives, but the reverse was true when treated with control treatments. CONCLUSIONS: Compared with nonalcoholic cocaine dependent subjects, codependent patients exhibit a wider array of problems, many of which merit professional attention. Both alcoholics and nonalcoholics exhibit substantial improvements during treatment, with alcoholics perhaps requiring extra treatment efforts for successful outcomes. PMID- 11271961 TI - Drinking levels, alcohol problems and secondhand effects in substance-free college residences: results of a national study. AB - OBJECTIVE: This study examines alcohol use, associated problems and secondhand effects among residents of substance-free and alcohol-free housing on U.S. college campuses. METHOD: In the spring of 1999, a nationally representative sample of students completed survey questionnaires regarding alcohol use and related behaviors. The responses of 2,555 (61.25% female) students living in different types of residences (substance-free, alcohol-free and unrestricted) at the 52 campuses at which these housing options existed were compared. RESULTS: Substance-free residences were not substance-free; however, residents drank less heavily and experienced fewer alcohol-related problems and secondhand effects than students living in unrestricted housing. They were less likely (three fifths) to engage in heavy episodic drinking. The difference between students in substance-free and unrestricted housing was greatest for students who had not been heavy episodic drinkers in high school and for those on campuses with lower overall levels of heavy episodic drinking. In contrast, students who lived in alcohol-free halls were no less likely to be heavily involved in alcohol use than were students in unrestricted housing. CONCLUSIONS: Residence in substance-free housing was associated with lower likelihood of heavy episodic drinking in college for students who were not heavy episodic drinkers in high school. Whether or not this is a causal relationship or a result of self-selection needs to be examined in a prospective study. These living arrangements are also associated with lower levels of secondhand effects. College administrators may want to consider offering or increasing their substance-free housing options as one possible method of decreasing heavy student drinking. PMID- 11271962 TI - Social influence processes and college student drinking: the mediational role of alcohol outcome expectancies. AB - OBJECTIVE: Social influences are among the most robust predictors of adolescent substance use and misuse. Studies with early adolescent samples have supported the need to distinguish among various types of social influences to better delineate relations between social factors and alcohol use and problems. METHOD: The first major goal of the present study (N = 399, 263 women) was to examine unique relations between particular facets of social influence and alcohol use and problems in a relatively heavy-drinking population (i.e., college students). We hypothesized that active social influences (offers to drink alcohol) and passive social influences (social modeling and perceived norms) would demonstrate positive associations with measures of alcohol use and problems. We also tested the hypothesis that alcohol outcome expectancies would mediate associations between social influences and drinking behaviors. RESULTS: Structural equation modeling analyses provided strong support for the first hypothesis. Social modeling demonstrated the strongest association with alcohol use and problems, and active social influences demonstrated significant associations with both use and problems. Perceived norms were related to alcohol use, but not directly with alcohol problems. Support for the second hypothesis was positive but limited to one type of social influence. Strong evidence for a mediational role of outcome expectancies was found for relations between social modeling and alcohol use and problems. CONCLUSIONS: Together, these findings demonstrate the unique and relative contribution of active and passive social influences and provide limited support for a hypothesized process by which social factors influence cognitions and alcohol-related behaviors. PMID- 11271963 TI - Family, religious, school and peer influences on adolescent alcohol use: a longitudinal study. AB - OBJECTIVE: In this study, the cross-temporal relationship between family social support and adolescent alcohol use was examined. A primary aim was to investigate the mechanisms through which family social support affects drinking among youth. Another aim was to examine reciprocal relationships among the study variables. METHOD: Four-wave (with 6-month intervals) panel survey data collected from 840 middle adolescent boys (n = 443) and girls (n = 397) attending a suburban school district in western New York were analyzed using structural equation modeling with maximum likelihood estimation. RESULTS: Analyses revealed that family social support was indirectly associated with decreased alcohol consumption among the respondents, primarily through variables measuring religiosity, school grades and peer alcohol use. In addition, adolescent alcohol use was directly associated with subsequent increases in peer alcohol use and later decreases in school performance. Results also showed that receiving good grades in school predicted moderate increases in family social support. CONCLUSIONS: The findings of this study are discussed in terms of the interrelationships that exist among multiple socializing influences and alcohol use among adolescents. PMID- 11271964 TI - Use of anabolic-androgenic steroids in adolescence: winning, looking good or being bad? AB - OBJECTIVE: To investigate the prevalence of anabolic-androgenic steroid (AAS) use among Norwegian adolescents and to contrast three perspectives on AAS use: performance enhancement in sports competition, body image and eating concerns, and AAS-use as belonging to a cluster of problem behaviors. METHOD: A nationally representative sample of 8,877 (53.8% female) Norwegian youths (15-22 years of age) were surveyed (response rate 78%). Sports participation included measures of participation in strength sports, participation in competitive sports, strength training and perceived athletic competence. Body image and eating concerns included measures of disordered eating, perceived physical appearance and satisfaction with body parts. Problem behavior was measured by three dimensions of conduct problems (overt destruction, overt nondestruction and covert destruction), illicit drug use and sexual involvement. RESULTS: Information about AAS was obtained from 8,508 subjects. Lifetime AAS use was 0.8% (1.2% male and 0.6% female), 12-month prevalence was 0.3% and 5.1% had been offered AAS. AAS use did not vary according to sports involvement and demographics. Logistic regression analyses showed that AAS use was associated with such problem behavior as marijuana (cannabis) involvement and overt nondestruction (e.g., aggressive type conduct problems) and, to some extent, with involvement in power sports and disordered eating. AAS users differed little from those who had been offered but refrained from using AAS, except that they were more likely to be current marijuana users. CONCLUSIONS: Adolescent AAS use seems primarily to be another type of problem behavior and only secondarily is it associated with strength sport participation and disordered eating. PMID- 11271965 TI - A preliminary evaluation of the potential usefulness of the diagnoses of polysubstance dependence. AB - OBJECTIVE: The concept of polysubstance dependence (PD) has been defined several ways over the years. However, few clinicians and researchers appear to use this label in a manner consistent with any of the major diagnostic manuals. This article evaluates the prevalence and characteristics associated with PD in participants in a large collaborative study. METHOD: In DSM-IV, PD characterizes people who do not meet criteria for dependence on any one substance but, when all drugs of abuse are considered have experienced three or more of the seven dependence items across the substances. In this study, structured face-to-face interviews were administered to 8,834 men and women as part of the Collaborative Study on the Genetics of Alcoholism. The 198 subjects (2.2%) with a slightly expanded concept of the DSM-IV disorder were compared with men and women with dependence on alcohol, marijuana or stimulants, subjects with substance abuse and those with no substance use disorder. RESULTS: In this dataset, compared with subjects with a specific substance dependence, those with PD were slightly more educated and less likely to be divorced or separated, and they had fewer substance-related problems. At the same time, those with PD had more substance problems than did subjects who only met criteria for abuse. These basic conclusions were unchanged among the subset of 59 subjects who met the more restricted, classical DSM-IV PD criteria. CONCLUSIONS: The data indicate that, while relatively rare, subjects with PD might differ in potentially important ways from those with dependence or abuse on specific drugs. A large prospective study of a group with carefully defined PD is needed. PMID- 11271966 TI - Can methodological features account for patient-treatment matching findings in the alcohol field? AB - OBJECTIVE: Despite enthusiasm for the potential of matching patients to alcohol treatments to improve outcomes, consistent findings have not emerged. This review considers the extent to which methodological factors may account for the pattern of findings from research on Patient x Alcohol Treatment interactions. METHOD: We focused on 55 studies that compared more than one type of alcohol treatment and included formal statistical tests for interactions. We examined four predictors of the number of significant interactions found in the 55 studies: (1) the number of statistical tests for interactions conducted, (2) the average number of participants, (3) whether or not participants were randomized to treatment and (4) the proportion of tested interactions that were hypothesis- or rationale driven, as opposed to exploratory. RESULTS: Only the number of statistical tests for interactions predicted the number of patient-treatment interactions identified per study (zero-order r = 0.47; r2 = 0.22). A substantial number of tests for interactions (43) was conducted, on average, per study. Only a minority of the studies (33%) included enough participants to have a reasonable probability (0.80) of identifying a medium-sized matching effect. CONCLUSIONS: Drawing general conclusions regarding matching patients to alcohol treatments is hampered because Type I error has contributed to the matches identified, studies in this area are often underpowered, and the combinations of patient and treatment variables that have been tested are few relative to the numerous possible combinations. To be productive, future research will need to focus on patients at the extremes of matching dimensions and on distinct treatments. (J Stud. Alcohol 62: 62-73, 2001) PMID- 11271967 TI - Reports of alcohol-related harm: telephone versus face-to-face interviews. AB - OBJECTIVE: To assess the effect of mode of administration in alcohol surveys (telephone vs face-to-face interviews), prevalence rates of self-reported harms due to alcohol were compared for two datasets with equivalent measures. METHOD: Two national alcohol surveys were used: the 1990 Warning Labels Survey, in which random digit dialing was used to generate a sample of 2,000 adults interviewed by telephone, and the 1990 National Alcohol Survey (face-to-face interviews), a probability sample of U.S. adults living in households (N = 2,058). Both surveys included identical items on five areas of alcohol-related harm, yielding one composite index of any harm reported in the last 12 months that was compared between the two surveys for current drinkers. RESULTS: After controlling for demographic characteristics and alcohol use, the telephone survey yielded significantly higher rates of alcohol-related health harm, work harm and "any harm" as compared to the in-person survey. The interaction between heavier drinking (five or more drinks during 1 day, weekly or more often) and method of data collection was significant for health harm and any harm. Respondents in the telephone survey who drank 5+ less than weekly were more likely than those interviewed in person to report health harm due to alcohol use; those in the telephone survey who drank 5+ weekly or more often were more likely to report any harm. CONCLUSIONS: Possible explanations for differences between the surveys include anonymity and fewer social desirability issues associated with telephone surveys, as well as potentially differing cognitive requirements in telephone versus face-to-face interviews. PMID- 11271968 TI - Measuring readiness-to-change substance misuse among psychiatric outpatients: I. Reliability and validity of self-report measures. AB - OBJECTIVE: The high rates of comorbid substance use disorders among persons living with severe and persistent mental illness (SPMI) have increased interest in assessing and enhancing motivation to change substance misuse in this population. This study provides evidence for the psychometric adequacy of three self-report measures of readiness-to-change. METHOD: The sample consisted of 84 persons (65% men) with co-occurring substance abuse or dependence and an SPMI. After a psychiatric assessment, participants completed three measures of readiness-to-change, which yielded seven subscales: (1) the Stages of Change Readiness and Treatment Eagerness Scale (ambivalence about change, recognition of substance-related problems, taking steps), (2) Decisional Balance Scale (pros of using, cons of using) and (3) the Alcohol and Drug Consequences Questionnaire (costs of quitting, benefits of quitting). RESULTS: All of the subscales were stable over time, and 6 of the 7 subscales demonstrated excellent internal consistency. Reliability indices were comparable when analyses were repeated on subsets of participants defined by diagnosis, cognitive function, positive symptoms and negative symptoms. A pattern of theoretically meaningful intercorrelations provided convergent evidence of validity, and a general lack of relationships with demographic variables and indices of psychiatric status provided discriminant evidence of validity. CONCLUSIONS: These findings support efforts to quantify readiness-to-change substance misuse among persons with an SPMI. PMID- 11271970 TI - Cloninger's temperament and character dimensions in young adulthood and their relation to characteristics of parental alcohol use and smoking. AB - OBJECTIVE: We examined the relationship of parental alcohol use (i.e., the frequency of alcohol intake and getting drunk) and smoking to Cloninger's temperament dimensions (Novelty Seeking, Harm Avoidance, Reward Dependence and Persistence) and character dimensions (Self-Directedness, Cooperativeness and Self-Transcendence) in young adulthood. METHOD: We used a 14-year longitudinal study of 1,849 (1,101 female) randomly selected healthy adolescents and young adults, and their parents. Alcohol consumption and smoking were self-reported by the parents. Offspring temperament and character were measured by the Temperament and Character Inventory (TCI) 14 years later. RESULTS: Maternal and paternal frequency of alcohol intake, getting drunk and smoking were associated with offspring temperament and character dimensions, particularly Novelty Seeking, in young adulthood for both men and women. CONCLUSIONS: The results support the relevance of Cloninger's concepts and the TCI in identifying subjects with unique characteristics related to their family histories. Possible mediating mechanisms are discussed. PMID- 11271969 TI - Association of outpatient alcohol and drug treatment with health care utilization and cost: revisiting the offset hypothesis. AB - OBJECTIVE: This study examines the hypothesis that treatment reduces medical utilization and costs of patients with substance use problems. METHOD: Adult patients (N = 1.011; 67% men) entering the outpatient chemical dependency recovery program at Sacramento Kaiser Permanente over a 2-year period were recruited into the study. Medical utilization and costs were examined for 18 months prior and 18 months after intake. To account for overall changes in utilization and cost, an age, gender and length-of-enrollment matched nonpatient control group (N = 4,925) was selected from health-plan members living in the same service area. Multivariate analyses controlling for age and gender were conducted using generalized estimating equation methods, allowing for correlation between repeated measures and nonnormal distributions of the outcome variable. RESULTS: The treatment cohort was less likely to be hospitalized (odds ratio [OR] = 0.59; p < .01) and there was a trend for having spent fewer days (rate ratio [RR] = 0.77; p < .10) in the hospital in the posttreatment period compared to pretreatment period. These patients were also less likely to visit the emergency room (ER) (OR = 0.64; p < .01) and had fewer ER visits (RR = 0.81; p < .01) following treatment. Inpatient, ER and total medical costs declined by 35%, 39% and 26%, respectively (p < .01). Reductions in cost were greater for the treatment cohort when compared with the matched sample (p < .05). Among women, there were significant reductions (p < .05) in inpatient, ER and total costs for the study cohort when compared with the matched sample; among men, the reductions in inpatient and ER cost (but not total cost) were significantly larger (p < .05) for the study cohort when compared with the matched sample. For the treatment cohort, the change in medical cost was not significantly different by gender. Changes in cost were significantly different across the various age groups (p < .05) for the study cohort and the matched sample. Among those in the group aged 40-49 years, the decline in cost for study cohort was significantly larger (p < .05) than for the matched sample. CONCLUSIONS: For patients with substance use disorders entering treatment, there was a substantial decline in inappropriate utilization and cost (hospital and ER) in the posttreatment period. The disaggregated pattern of posttreatment decline in utilization and cost is suggestive of long-term reductions that warrant a longer follow-up. PMID- 11271971 TI - Examination of the neighborhood activation theory in normal and hearing-impaired listeners. AB - OBJECTIVE: Experiments were conducted to examine the effects of lexical information on word recognition among normal hearing listeners and individuals with sensorineural hearing loss. The lexical factors of interest were incorporated in the Neighborhood Activation Model (NAM). Central to this model is the concept that words are recognized relationally in the context of other phonemically similar words. NAM suggests that words in the mental lexicon are organized into similarity neighborhoods and the listener is required to select the target word from competing lexical items. Two structural characteristics of similarity neighborhoods that influence word recognition have been identified; "neighborhood density" or the number of phonemically similar words (neighbors) for a particular target item and "neighborhood frequency" or the average frequency of occurrence of all the items within a neighborhood. A third lexical factor, "word frequency" or the frequency of occurrence of a target word in the language, is assumed to optimize the word recognition process by biasing the system toward choosing a high frequency over a low frequency word. DESIGN: Three experiments were performed. In the initial experiments, word recognition for consonant-vowel-consonant (CVC) monosyllables was assessed in young normal hearing listeners by systematically partitioning the items into the eight possible lexical conditions that could be created by two levels of the three lexical factors, word frequency (high and low), neighborhood density (high and low), and average neighborhood frequency (high and low). Neighborhood structure and word frequency were estimated computationally using a large, on-line lexicon based Webster's Pocket Dictionary. From this program 400 highly familiar, monosyllables were selected and partitioned into eight orthogonal lexical groups (50 words/group). The 400 words were presented randomly to normal hearing listeners in speech-shaped noise (Experiment 1) and "in quiet" (Experiment 2) as well as to an elderly group of listeners with sensorineural hearing loss in the speech-shaped noise (Experiment 3). RESULTS: The results of three experiments verified predictions of NAM in both normal hearing and hearing-impaired listeners. In each experiment, words from low density neighborhoods were recognized more accurately than those from high density neighborhoods. The presence of high frequency neighbors (average neighborhood frequency) produced poorer recognition performance than comparable conditions with low frequency neighbors. Word frequency was found to have a highly significant effect on word recognition. Lexical conditions with high word frequencies produced higher performance scores than conditions with low frequency words. CONCLUSION: The results supported the basic tenets of NAM theory and identified both neighborhood structural properties and word frequency as significant lexical factors affecting word recognition when listening in noise and "in quiet." The results of the third experiment permit extension of NAM theory to individuals with sensorineural hearing loss. Future development of speech recognition tests should allow for the effects of higher level cognitive (lexical) factors on lower level phonemic processing. PMID- 11271973 TI - Variation in speech perception scores among children with cochlear implants. AB - OBJECTIVE: The objective of this study was to identify common factors affecting speech perception scores in children with cochlear implants. DESIGN: Speech perception data for 167 implanted children were collected at two cochlear implant centres in Melbourne and Sydney. The data comprised audition-alone scores on open set word and sentence tests. Children were selected on the basis that they had a Nucleus 22-electrode cochlear implant. The average age of the children was 5 yr. Information was also collected about 12 factors that may have influenced speech perception scores for each child. Analysis of covariance was used to identify factors that significantly affected speech perception scores. Pearson pairwise correlation coefficients were also calculated for all factors analyzed. RESULTS: The analyses in this study identified factors that accounted for 51%, 34%, and 45% of the variance in phoneme, word and sentence perception scores. Scores decreased by 1.4 to 2.4% per year of profound deafness prior to implantation. Children who normally use oral communication scored significantly higher than children normally using sign or simultaneous oral and sign communication. Children implanted in Sydney scored higher on average than children implanted in Melbourne. CONCLUSIONS: The results show that a significant part of the variation in speech perception scores is systematically related to audiological and environmental factors for each child. The reasons for significant differences between children using different communication modes or from different clinics were not identified. PMID- 11271972 TI - Sudden cortical hearing loss for speech: a case report. AB - A 35-yr-old male patient experienced a sudden loss of speech understanding due to a bilateral cerebrovascular disease. A detailed summary of audiological and neurological findings was presented. Findings indicated that the presence of Pa waves of the Middle Latency Response (MLR) may be a positive prognosis for improvement in hearing thresholds and speech understanding. PMID- 11271974 TI - Articulatory changes with short-term deactivation of the cochlear implants of two prelingually deafened children. AB - OBJECTIVE: The purpose of this investigation was to determine how suddenly diminished auditory feedback affects articulatory behaviors for prelingually deafened children with cochlear implants (CIs). DESIGN: Two 6-yr-old children served as participants. Considering their level of hearing impairment, one child had above average speech perception and production skills while the other child had exceptionally good speech perception and production abilities. Baseline data were collected four times over the course of 2 days with the children wearing their CIs. For three additional days, data were collected while the children wore their CIs (ON condition) and then again after their CIs had been deactivated for 1 hr (OFF condition). Variables assessed included amount of jaw opening, F1, F2, nasal air flow, voice onset time (VOT), voicing duration, and the magnitude and duration of intraoral air pressure (Po). Findings were related to each other and to previously reported phonatory findings from the same two children (Higgins, McCleary, & Schulte, 1999) to determine whether changes in articulatory variables in the OFF condition were consistent with a direct effect of diminished auditory feedback or an indirect influence of suprasegmental parameters. RESULTS: Both children exhibited consistent and significant changes in articulatory parameters in the absence of auditory feedback. Such changes occurred more often for the child who had especially proficient speech perception and production skills. Some changes (i.e., reduced Po for [p] and reduced nasal air flow for [m]) appeared related to the influence of suprasegmental parameters, in particular, reduced subglottal air pressure. Other effects (i.e., increased F2 for [a] and reduced VOT for [p]) were suggestive of changes in the children's abilities to maintain appropriate articulatory placements and timing. Finally, a few changes (reduced jaw opening for [i] and increased Po duration for [p]) may have reflected compensatory strategies to maintain correct tongue placement and enhance temporal distinctions in the absence of auditory feedback. CONCLUSIONS: Based on the data of our two participants, it appears that some prelingually deafened children with CIs and good speech perception/production skills rely on auditory feedback to maintain articulatory precision. In the absence of auditory feedback, such children may demonstrate changes in articulatory placement and timing. In addition, data from one of our participants are consistent with the idea that some children may be aware that their articulatory control is compromised in the absence of auditory feedback and attempt to compensate by altering articulatory durations or the range of associated articulatory movements. PMID- 11271975 TI - Predicted and measured speech recognition performance in noise with linear amplification. AB - OBJECTIVE: The purpose of this study was to investigate the applicability of the Speech Intelligibility Index (SII) in hearing aid fitting. It was hypothesized that estimated speech intelligibility, based on the SII, could be a more reliable measure than real speech recognition results for comparing hearing aid characteristics. DESIGN: The test subjects were 29 elderly persons (66 to 80 yr) with mild-to-moderate hearing loss, who were using monaurally fitted linear hearing aids. They were selected from the files at Sahlgrenska hearing clinic. Speech recognition scores were obtained at fixed speech-to-noise ratios with Phonemically Balanced (PB) words in speech-weighted noise and in low-frequency noise. A Just-Follow-Conversation (JFC) test was performed with connected speech presented in the same background noises. The subjects were tested without hearing aid and with their hearing aids set at three different frequency responses. Predicted speech recognition scores were calculated for each condition based on the SII, complemented with a correction for sensorineural hearing impairment. The calculations involved speech and noise spectra, pure tone thresholds and insertion gain responses. RESULTS: For each condition, the measured speech recognition scores were, on average, well predicted by the calculated scores. The intra-individual standard deviation of the predicted scores was estimated to be about one percent unit. The group results of the JFC test were in agreement with the word recognition results for the aided conditions, but a floor effect was observed for the unaided conditions. CONCLUSIONS: Speech intelligibility prediction based on the modified SII is a valid estimate of speech recognition performance of hearing-impaired persons with mild-to-moderate hearing loss. Estimated intelligibility based on the SII is more reliable than actually measured speech recognition performance, for comparing amplification conditions within subjects. PMID- 11271976 TI - An examination of several characteristics that affect the prediction of OSPL90 in hearing aids. AB - OBJECTIVE: Investigators at the National Acoustic Laboratories have provided a theoretical derivation and experimental validation of a formula for setting the maximum output of hearing aids (Dillon & Storey, 1998; Storey, Dillon, Yeend, & Wigney, 1998). Given that measurement of discomfort levels for setting maximum output can be both time-consuming and of questionable reliability, the use of a prescriptive formula warrants consideration. In this article, an extensive data base was considered and issues of normal hearing, clinical protocol, age and gender were investigated in an effort to further determine optimal maximum output settings. DESIGN: Data were gathered from five previous investigations, for a total of 433 subjects (total ears = 710). Threshold of discomfort (TD) measures were obtained using one of two adaptations of the Ascending Method of Limits, one with category anchors and one without. RESULTS: Subjects with normal hearing had significantly lower TDs than subjects with hearing loss. A different regression line for measured TDs as a function of hearing level was noted for subjects whose hearing threshold levels fell between 20 and 60 dB HL and those with thresholds above 60 dB HL. When all effects (hearing level, method, age and gender) were considered in a single predictive model for the two threshold groups, only method and threshold were significant predictors of TD. However, for the subjects with thresholds between 20 and 60 dB HL, less than 4% of the variance in TD measures could be accounted for by those factors. For subjects with threshold above 60 dB HL, 22% of the variance in TD measures could be accounted for by those variables. CONCLUSIONS: For both groups of subjects (20 to 60 dB HL and above 60 dB HL) methodology and hearing thresholds are significant predictors of discomfort levels. Age and gender are not. Given the small variance accounted for by any of these factors, measures of discomfort using standardized methodologies seem warranted. PMID- 11271977 TI - Delay and temporal integration in medial olivocochlear bundle activation in humans. AB - OBJECTIVE: Contralateral suppression of the transient-evoked otoacoustic emissions (TEOAEs) provides a means of studying auditory efferent function, but the temporal dynamics of the reflex are not fully understood. The most fundamental parameter is the time-course of activation of contralateral suppression. The stimulus parameters are likely to be important; this may include temporal dynamics of the suppressor itself. This investigation thus was devoted to the further study of 1) delay of contralateral suppression of TEOAEs-effect of delay of the ipsilateral probe-and 2) temporal variation of the suppressor-effect of amplitude modulation of the contralateral noise stimulus. DESIGN: Measurements were made in three samples of normal-hearing subjects (N(total) = 71), employing well-established methods of TEOAE assessment. RESULTS: Statistically significant contralateral suppression occurred some 60 msec after onset of the contralateral noise; thereafter, the effect was essentially constant (i.e., to >180 msec). The results for click delays less than 60 msec, nevertheless, were systematic and readily fitted by a sloping straight line (dB suppression versus time) reminiscent of the concept of threshold power integration. The onset of suppression may thus be characterized by a time constant. The delay of suppression also was found to be reduced by contralateral amplitude-modulated noise. CONCLUSIONS: These findings reinforce a growing consensus in the literature that, despite initiation perhaps some milliseconds after onset of the contralateral stimulus, there is a substantial delay, i.e., in the tens of milliseconds, before maximal suppression is achieved. The exact time constant of this effect appears to depend upon the combination of probe and suppressor levels, including the temporality of the suppressor. These factors are likely to delimit the role/influence of this reflex in real-world function, favoring perhaps more-or-less sustained suppression that is activated in a time-varying sound environment. PMID- 11271978 TI - Brief report: the cochlear microphonic as an indication of outer hair cell function. AB - The extra-cellular cochlear microphonic is believed to be generated predominantly by outer hair cells and therefore it would seem reasonable to assume that the presence of a cochlear microphonic excludes outer hair cell dysfunction. Indeed, a diagnosis of auditory neuropathy might be, and has been, made on the basis of a cochlear microphonic present with an abnormal auditory brainstem response. Animal studies, however, have shown that the cochlear microphonic recorded from the round window is dominated by cellular generators located in the base of the cochlea. Primarily on this basis, it is argued that the presence of a cochlear microphonic does not exclude outer hair cell pathology and so outer hair cell integrity should not necessarily be inferred from the presence of the cochlear microphonic alone. In contrast, the absence of an otoacoustic emission in such cases is consistent with outer hair cell dysfunction. PMID- 11271979 TI - The biology and treatment of chronic myelogenous leukemia. AB - Over the past 2 decades, our understanding of the pathobiological events underlying chronic myelogenous leukemia (CML) has grown. At the same time, effective transplant and nontransplant treatment approaches to CML have been developed that increase the options available to newly diagnosed patients, and that can cure or prolong survival in this formerly incurable disease. Newly diagnosed patients presenting with extreme leukocytosis or thrombocytosis may benefit from immediate therapy with hydroxyurea (Hydrea) and pheresis. After stabilization, eligible patients may elect to undergo immediate transplant. The majority, however, should begin therapy with either interferon-alpha and cytarabine, or they should be entered into the STI-571 trials. PMID- 11271980 TI - Lung cancer vaccine demonstrates enhanced antitumor immunity. PMID- 11271981 TI - Breast cancer during pregnancy. AB - The care of a pregnant breast cancer patient is a challenging clinical situation that historically has placed the welfare of the mother in conflict with that of the fetus. For the woman in this situation, the emotions usually associated with pregnancy can be overshadowed by the emotions aroused by a diagnosis of breast cancer and its subsequent treatment. The majority of published information on the management of breast cancer during pregnancy has consisted of retrospective chart reviews, case reports, and anecdotes. There is a paucity of published data from the prospective study of women who are pregnant at the time of their breast cancer diagnosis. This review will endeavor to address the diagnosis, staging, and subsequent treatment of breast cancer during pregnancy. The limited information available for this group of women on the outcomes of labor, delivery, and neonatal health will also be reviewed. This review will not specifically address pregnancy that occurs after diagnosis and subsequent treatment for breast cancer. However, some data, particularly those of an epidemiologic nature, address breast cancer diagnosed during pregnancy or within the year following delivery. PMID- 11271982 TI - Clinical trials referral resource. Current clinical trials in myelodysplatsic syndromes. PMID- 11271983 TI - Current management of opioid-related side effects. AB - The optimal management of opioid-related side effects is hampered by a lack of comparative studies of management strategies. The prevalence of such side effects is influenced by the extent of disease, the patient's age, the presence of coexistent renal and hepatic disease, pulmonary disease, and cognitive dysfunction, a prior opioid history, use of polypharmacy, dose of opioid drug being administered, and the route of administration. The most common opioid related side effects are constipation, sedation, nausea, vomiting, and cognitive disturbance. Less frequent side effects include urinary retention, perceptual distortion, respiratory depression, and myoclonus. In an era emphasizing quality of life in cancer care, clinicians need to be aware of (1) factors that influence the prevalence of opioid-related side effects, (2) effective management strategies, and (3) how to recognize when symptoms are opioid related as opposed to caused by other etiologies, such as the patient's disease process or treatment approaches. The use of validated instruments and repeated assessment enhances such an evaluation and subsequent treatment. This article delineates the current optimal management of opioid-related nausea and vomiting, constipation, cognitive side effects, myoclonus, and respiratory depression. PMID- 11271984 TI - Maintenance therapy for superficial bladder cancer. AB - Transurethral resection remains the standard for first-line treatment of transitional cell carcinoma of the bladder. This technique clearly defines the pathologic grade and is essential in determining the clinical stage of the bladder tumor. Intravesical therapy is an important adjunct to transurethral resection in the management of patients with superficial bladder cancer, many of whom are at risk for disease recurrence and progression. Pharmacotherapy consisting of cytotoxic and immunomodulating agents has demonstrated utility against superficial transitional cell carcinoma. Bacillus Calmette-Guerin and mitomycin (Mutamycin) remain the more commonly used and most effective agents in the prophylaxis against recurrence and progression of superficial bladder transitional cell carcinoma. Many studies have examined their efficacy at different schedules. This article reviews the traditional intravesical agents that are useful in the therapy and prophylaxis of superficial transitional cell carcinoma of the bladder. It also addresses their long-term efficacy when used as maintenance therapy in higher-risk patients. PMID- 11271985 TI - Is there a role for intraperitoneal chemotherapy in the management of ovarian cancer? AB - Phase I and II clinical trial data have demonstrated the safety, pharmacokinetic advantage, and potential for enhanced cytotoxicity associated with the intraperitoneal administration of antineoplastic agents in the management of ovarian cancer. In two randomized phase III studies comparing the intraperitoneal and intravenous administration of cisplatin (Platinol) as initial therapy for small-volume residual advanced ovarian cancer, intraperitoneal delivery of the agent produced superior progression-free and overall survival. Reluctance to employ intraperitoneal cisplatin in the standard management of ovarian cancer appears to be related to the added time, effort, and potential morbidity associated with the approach, as well as a general preference for the less toxic, less complicated carboplatin (Paraplatin)-based regimen. However, existing data support the use of this unique method of drug delivery in carefully selected patients outside of the clinical trial setting. PMID- 11271986 TI - Excited states at surfaces: Fano profiles in STM spectroscopy of adsorbates. AB - The Fano-Anderson model for a discrete state embedded within a continuum is revisited within the context of excitation and decay processes which lead to some manifestations of Fano lineshape profiles. The phenomenon of resonance tunneling between an STM tip and a metal surface upon which there are isolated adsorbed atoms is discussed and the relationship between the spectroscopic signature of such systems and that of the Fano profile is taken up. Recent experimental studies of Kondo systems of magnetic adsorbates such as Co and Ce adsorbed on noble metal (111) surfaces have motivated this work. PMID- 11271987 TI - Quantum dynamics of the dissociation of H2 on Cu(100): dependence of the site reactivity on initial rovibrational state. AB - We perform six-dimensional (6D) quantum wavepacket calculations for H2 dissociatively adsorbing on Cu(100) from a variety of rovibrational initial states. The calculations are performed on a new potential energy surface (PES), the construction of which is also detailed. Reaction probabilities are in good agreement with experimental findings. Using a new flux analysis method, we calculate the reaction probability density as a function of surface site and collision energy, for a variety of initial states. This approach is used to study the effects of rotation and vibration on reaction at specific surface sites. The results are explained in terms of characteristics of the PES and intrinsically dynamic effects. An important observation is that, even at low collision energies, reaction does not necessarily proceed predominantly in the region of the minimum potential barrier, but can occur almost exclusively at a site with a higher barrier. This suggests that experimental control of initial conditions could be used to selectively induce reaction at particular surface sites. Our predictions for site-reactivity could be tested using contemporary experimental methods: The calculations predict that, for reacting molecules, there will be a dependence of the quadrupole alignment of j on the incident vibrational state, v. This is a direct result of PES topography in the vicinity of the preferred reaction sites of v = 0 and v = 1 molecules. Invoking detailed balance, evidence for this difference in preferred reaction site of v = 0 and 1 molecules could be obtained through associative desorption experiments. PMID- 11271989 TI - The role of rotational excitation in the activated dissociative chemisorption of vibrationally excited methane on Ni(100). AB - We have measured the sticking probability of methane excited to v = 1 of the v3 antisymmetric C-H stretching vibration on a clean Ni(100) surface as a function of rotational state (J = 0, 1, 2 and 3) and have investigated the effect of Coriolis-mixing on reactivity. The data span a wide range of kinetic energies (9 49 kJ mol-1) and indicate that rotational excitation does not alter reactivity by more than a factor of two, even at low molecular speeds that allow for considerable rotation of the molecule during the interaction with the surface. In addition, rotation-induced Coriolis-splitting of the v3 mode into F+, F0 and F- states does not significantly affect the reactivity for J = 1 at 49 kJ mol-1 translational energy, even though the nuclear motions of these states differ. The lack of a pronounced rotational energy effect in methane dissociation on Ni(100) suggests that our previous results for (v = 1, v3, J = 2) are representative of all rovibrational sublevels of this vibrational mode. These experiments shed light on the relative importance of rotational hindering and dynamical steering mechanisms in the dissociative chemisorption on Ni(100) and guide future attempts to accurately model methane dissociation on nickel surfaces. PMID- 11271988 TI - Energy disposal during desorption of D2 from the surface and subsurface region of Ni(111). AB - The recombination of surface and subsurface D atoms on Ni(111) has been studied using resonance-enhanced multiphoton ionisation (REMPI) to measure the internal state and translational energy distributions of the desorbing product. By detecting D2 formed during temperature-programmed desorption we were able to examine the reaction between subsurface and surface D atoms, and the recombination of two D atoms chemisorbed on the surface. Translational energy distributions for D2 formed by recombination of surface D are very sensitive to coverage. Desorption from a low coverage surface produced a translational energy release of 2.6 kT, but a thermal rotational distribution, reflecting an entrance channel barrier to dissociative chemisorption on the clean Ni(111) surface. Sticking probabilities predicted from detailed balance are consistent with molecular beam adsorption measurements. Desorption from D coverages above 0.5 ML resulted in a sub-thermal energy release, desorption being mediated by a molecular precursor state with D2 dissociation occurring via a non-activated, trapping-dissociation channel. In contrast, the reaction of subsurface D produces translationally hot D2, with a mean energy approaching 8 kTs at 180 K. This is consistent with the energetics for direct recombination of a chemisorbed D atom with a metastable subsurface D atom, which overcomes an activation barrier to resurface of between 0.35 and 0.47 eV depending on D concentration. The energy release decreases at higher temperature, probably as a result of a reduction in the energy of resurfacing D as the subsurface D concentration drops. This low energy component is attributed to accommodation of resurfacing D which is unable to react directly, followed by slow thermal desorption via the high coverage, surface D recombination channel. No internal rotational or vibration excitation was found in D2 formed by reaction of subsurface D. PMID- 11271990 TI - Long-lived adsorbate states on metal surfaces. AB - It has been shown recently that the peculiarities of the band structure of a metal can qualitatively influence the electron tunnelling between an adsorbate and a metal surface, the so-called resonant charge transfer (RCT). The presence of a projected band gap along the normal to the surface in the case of Cu(111) has been shown to lead to a blocking of the RCT in the case of Cs/Cu(111), resulting in the existence of a very long-lived excited state. Such long-lived states are potentially very important for surface reaction mechanisms invoking a transient state as an intermediate. Various systems: Cs, model M- negative ion of p pi symmetry, CO adsorbed on Cu(111), are investigated in order to determine the conditions for the blocking of the RCT and the existence of long-lived states. PMID- 11271991 TI - Structure and dynamics of excited electronic states at the adsorbate/metal interface: C6F6/Cu(111). AB - Excited state electron transfer at the adsorbate/metal interface represents a key step in molecular electronic devices. The dynamics of such processes are governed by ultrafast energy relaxation which can be probed directly by time-resolved two photon photoemission (2PPE). Using 2PPE spectroscopy we investigate the energetics and lifetimes of the unoccupied electronic states of C6F6 adsorbed on Cu(111) as a model system for electron transfer at organic/metal interfaces. With increasing C6F6 layer thickness we find a pronounced decrease in the energetic position of the lowest unoccupied state, which is accompanied by a strong increase in its lifetime as well as a decrease in the effective electron mass. The frequently employed dielectric continuum model which describes delocalized (quantum well) states within adsorbate layers does not give a consistent explanation of these findings. By adsorption of Xe overlayers onto C6F6/Cu(111) we can show that, even for one monolayer of C6F6, the excited state must be localized predominantly inside the C6F6 layer and thus originates from a molecular state (presumably an antibonding sigma* orbital). With increasing coverage this state becomes more delocalized within the adsorbate layer, which reduces the coupling to the metal substrate and thus enhances the excited state lifetime. PMID- 11271992 TI - Excitation mechanisms and photochemistry of adsorbates with spherical symmetry. AB - By comparing the photo-stimulated desorption of Xe from an oxidized Si(100) surface with the photochemistry of methane on metal surfaces, we try to deduce a common concept in the excited state and the excitation mechanism responsible for the photo-induced processes. Xe atoms are desorbed from the oxidized Si(100) surface by the irradiation of photons in the range 1.16-6.43 eV. Two velocity components with average kinetic energy 0.85 and 0.25 eV are observed in the time of-flight distributions. The fast component appears only if the photon energy exceeds approximately 3 eV, but the slow component is present over the entire photon energy range. By analogy with the photochemistry of methane on metal surfaces, the excitation mechanism responsible for the fast component is postulated to be a transition from the 5p state of Xe to the excited state originating from strong hybridization between the 6s state of Xe and the dangling bond at a surface silicon atom bonded to oxygen inserted in the dimer bond. In this scheme an excited electron is transferred from the adsorbate to the substrate, which is the reverse direction to the substrate-mediated excitation frequently assumed in surface photochemistry. PMID- 11271994 TI - Electronic mechanism of STM-induced diffusion of hydrogen on Si(100). AB - We have observed a scanning tunneling microscopy (STM) induced lateral transfer of a single hydrogen atom on the Si(100) surface. The transfer rate of the hydrogen atom is proportional to the electron dose, indicating an electron assisted transfer mechanism. Measurements of the relations between the transfer rate and the sample bias and temperature give further support for an electronic mechanism. The bias dependence of the transfer rate shows a peak, and from a first principles electronic structure calculation we show that the position of the peak is related to the energy of a localized surface resonance. We propose that the hydrogen transfer is related to inelastic hole scattering with this surface resonance. We develop a microscopic model for the hydrogen transfer, and using the experimental data we extract information on the resonance lifetime and the transfer yield per resonant electron. The transfer takes place by tunneling through a small excited state transfer barrier. The transfer rate is increased if the hydrogen atom before the resonant excitation is vibrationally excited, and this gives rise to an increasing transfer rate with increasing sample temperature. PMID- 11271993 TI - Controlling organic reactions on silicon surfaces with a scanning tunneling microscope: theoretical and experimental studies of resonance-mediated desorption. AB - The dynamics of tip-induced, resonance-mediated bond-breaking in complex organic adsorbates is studied theoretically and experimentally. Desorption of benzene from a Si(100) surface is found to be efficient and sensitive to voltage, the measured yield rising from below 10(-10) to ca. 10(-6) per electron within a ca. 0.8 V range at low (< 100 pA) current. A theoretical model, based upon first principles electronic structure calculations and quantum mechanical wavepacket simulations, traces these observations to multi-mode dynamics triggered by a transition into a cationic resonance. The model is generalized to provide understanding of, and suggest a means of control over, the behaviour of different classes of organic adsorbates under tunneling current. PMID- 11271995 TI - Inelastic interactions of tunnel electrons with surfaces. AB - Inelastic interactions of electrons emitted from the tip of a scanning tunnelling microscope (STM) are used to desorb individual hydrogen atoms from a Ge(111) surface. It is observed that the inelastic interactions depend not only on the electron energy and the current intensity but also on the electron emission regime of the STM tip. Quite surprisingly, it is found that tunnel electrons interact inelastically much less efficiently than field emitted electrons even though the electrons are in resonance with the Ge-H unoccuppied orbital. PMID- 11271996 TI - The initiation and characterization of single bimolecular reactions with a scanning tunneling microscope. AB - A scanning tunneling microscope (STM) operating at 9 K in ultrahigh vacuum was used to initiate a bimolecular reaction between isolated hydrogen sulfide and dicarbon molecules on the Cu(001) surface. The reaction products ethynyl (CCH) and sulfhydryl (SH) were identified by inelastic electron tunneling spectroscopy (STM-IETS) and by sequentially removing hydrogen atoms from an H2S molecule using energetic tunneling electrons. For comparison, the thermal diffusion and reaction of H2S and CC at 45 K and H2O and CC at 9 K were also observed. PMID- 11271998 TI - Theoretical aspects of tunneling-current-induced bond excitation and breaking at surfaces. AB - We have performed a density functional study of the electronic structure, images and vibrationally inelastic tunneling in the scanning tunneling microscope and vibrational damping by excitation of electron-hole pairs of CO chemisorbed on the (111) and (100) faces of Cu. We find that the 2 pi* molecular orbital of CO turns into a broad resonance with parameters that differ significantly from those suggested by inverse and two-photon photoemission measurements. The calculated vibrational damping rate for the internal stretch mode and relative changes in tunneling conductance across vibrational thresholds are in agreement with experiment. The non-adiabatic electron-vibration coupling is well described by the Newn-Anderson model for the 2 pi*-derived resonance whereas this model is not able to describe the non-adiabatic coupling between the tunneling electrons and the vibration. We believe that this model misses an important mechanism for vibrational excitation in tunneling that involves the change of tunneling amplitude by deformation of the tails of the one-electron wavefunctions with vibrational coordinate. PMID- 11271997 TI - Effect of the projected band gap on the formation of negative ions in grazing collisions from Cu surfaces. AB - The formation of negative ions (H-, O-, S-, F-, Cl-) is studied for grazing scattering of fast ions from Cu(110) and Cu(111) surfaces. In a detailed experimental and theoretical investigation we reveal that the projected L-band gap of the Cu metal affects charge transfer in a specific manner. From the analysis of the negative ion fractions as functions of projectile velocity we conclude that, for the Cu(111) surface the electronic 2D surface state continuum plays an essential role in the projectile-surface electron transfer. PMID- 11271999 TI - Dynamics of charge transfer states on metal surfaces: the competition between reactivity and quenching. AB - The dynamics of excited states of adsorbates on surfaces caused by charge transfer is studied. Both negative and positive charge transfer processes are possible. In particular we are interested in positive charge transfer from a metal surface to molecular or atomic oxygen adsorbed on the surface. Once the negatively charged oxygen on the surface loses an electron it becomes chemically activated. The ability of this species to react depends on the quenching time or back transfer. The analysis of these processes is based on a set of diabatic potential energy surfaces each representing a different charged oxygen species. The dynamics is followed by solving the multichannel time-dependent Schrodinger equation or Liouville von Neumann equation. Due to the nonadiabatic character of these reactions large isotope effects are predicted. PMID- 11272000 TI - Self-trapped excitons at the quartz(0001) surface. AB - We have studied self-trapped excitons in alpha-quartz using density functional theory (DFT), both in the crystal and at the (0001) surface. The excitons are triplet excited states that distort the crystal locally. They have a long lifetime, of the order of a millisecond, and become thermally equilibrated. We have calculated the drop in the exciton energy as it approaches the surface from the interior of the crystal. In the subsurface layer of the -OH terminated (0001) surface, the energy has dropped by 0.7 eV. Another 0.4 eV drop occurs as the exciton enters the surface layer, where it breaks off an OH radical. The drop in energy can be understood from the greater ease of structural distortion at the surface. These calculations illustrate that excitons formed in the bulk could migrate out to the surface and form chemically active surface species. Molecules adsorbed at the surface could also serve as traps for the excitons and could, in principle, be induced to undergo structural or chemical transitions. PMID- 11272001 TI - Abstractive chemisorption of O2 on Al(111). AB - We present experimental evidence that abstraction is a common mechanism (approximately 50%) in the dissociative chemisorption of oxygen on Al(111) at a translational energy of 0.5 eV. As a result of this mechanism, individual isolated O-atoms are observed in scanning tunneling microscopy (STM). At this translational energy ordinary dissociative chemisorption processes also occur, resulting in pairs of adatoms. The ejected O-atoms originating from the abstraction reaction are detected in the gas phase using laser spectrometry. Together, these observations provide compelling evidence for the abstraction mechanism. PMID- 11272002 TI - Chemical selectivity in the remote abstractive chemisorption of ICl on Al(111). AB - The interaction of ICl and Al(111) involves remote dissociation in its chemisorption process. In remote dissociation, an electron harpoons from an Al(111) surface to an ICl gas molecule to initiate the chemisorption process. We have determined that ICl can chemisorb onto Al(111) by non-activated direct chemisorption, and the sticking probability of this direct channel is 0.65 +/- 0.03. Furthermore, low energy ICl molecules that do not undergo remote dissociation can chemisorb onto Al(111) by precursor-mediated chemisorption. Not only is the interaction of ICl and Al(111) reactive, it is chemically selective. Studies with Auger spectroscopy reveal that the ratio of chlorine atoms to iodine atoms on the Al(111) is 0.32 +/- 0.10 at low (0.042 +/- 0.002) surface coverage. Time-of-flight mass spectrometry studies also show that chlorine atoms are the only species scattered from the surface after ICl interacts with Al(111). These results indicate that iodine-selective abstraction, in which the iodine atom of ICl chemisorbs to the aluminium surface while the chlorine atom is ejected into the gas phase, is the dominant mechanism in this reaction. Iodine-end first collisions are more reactive than chlorine-end first collisions because the lowest unoccupied molecular orbital (LUMO) of ICl is primarily composed of iodine atomic orbitals, and it is the LUMO that interacts with the harpooning electron from the aluminium. PMID- 11272003 TI - Charge-transfer reactions in atom scattering from ionic surfaces: a time dependent wavepacket approach. AB - A diabetic description of charge transfer between atoms and ionic surfaces is presented, specifically examining the F/LiF(100) and F/KI(100) systems for which experiment shows ion formation to be very efficient. Potential energy surfaces describing the energetics for these systems have been generated with a semi empirical scheme. At the site of charge exchange, there is a curve-crossing between the ground state and the state representing charge capture by the projectile. Quantum dynamics calculations with time-dependent wavepacket methods give an initial ion-formation probability of unity for all cases considered. At lowest energies, the ions cannot escape the surface, giving an effective threshold for negative-ion production very close to that observed in experiment. Re-neutralization by charge transfer back to the conduction band of the solid is also examined. PMID- 11272004 TI - Direct and indirect DIET and DIMET from semiconductor and metal surfaces: what can we learn from 'toy models'? AB - Desorption induced by electronic transitions (DIET) and its variant DIMET (M = 'Multiple'), are among the simplest possible "reactions" of ad-species involving ultra-short lived electronically excited states at surfaces. The non-adiabatic bond-cleavage can be enforced, for example, with laser irradiation or with electrons or holes emitted from the tip of a scanning tunnelling microscope (STM). The transient creation of excited intermediates can proceed directly (localised to the adsorbate-substrate complex), or indirectly (i.e., through the substrate). To understand the basic processes, simple one-mode two-state "toy models" such as the Menzel-Gomer-Redhead (MGR) or the Antoniewicz scenarios have proven very useful in the past. We adopt and extend MGR- and Antoniewicz-type models together with numerically exact open-system density matrix theory to address a few actual problems/experiments in DI(M)ET: (1) Direct, laser-induced desorption of H(D) from Si(100) surfaces which has been realised in the continuous-wave DIET regime only recently [T. Vondrak and X.-Y. Zhu, Phys. Rev. Lett., 1999, 82, 1967], is studied and compared to so-far hypothetical femtosecond laser desorption. The possibility of controlling the reaction by shaping the laser pulses is addressed. (2) For the same system, temperature effects are studied for electron- or hole-stimulated desorption with an STM [T. C. Shen, C. Wang, G. C. Abeln, T. R. Tucker, J. W. Lyding, Ph. Avouris and R. E. Walkup, Science, 1995, 268, 1590; C. Thirstrup, M. Sakurai, T. Nakayama and K. Stokbro, Surf. Sci., 1999, 424, L329]. A modified version of Gadzuk's "sudden transition and averaging" approach is adopted which accounts for temperature dependent excited state lifetimes. (3) For photodesorption of NO from Pt(111), based on quantum dynamical simulations possible experimental tests involving static electric fields are suggested to address the relevance of the recently challenged [F. M. Zimmermann, Surf. Sci., 1997. 390, 174], "negative ion resonance" model of the Antoniewicz type. PMID- 11272005 TI - Photoinduced charge-transfer reaction at surfaces. Part I. (HCl)m..Nan/LiF(001) + hv (640 nm)-->(HCl)m - 1 Cl-Nan+/LiF(001) + H(g). AB - A sub-monolayer of atomic sodium, Nan, was deposited on LiF(001) at 50 K and characterized by temperature-programmed desorption, X-ray photoelectron spectroscopy, and titration with HCl. The Nan was dosed with HCl to form (HCl)m..Nan/LiF(001), which was then irradiated by 640 nm laser-radiation to induce a charge-transfer (CT) reaction. Reaction-product atomic H(g) was observed leaving the surface, by two-color Rydberg-atom time-of-flight (TOF) spectroscopy. These H-atoms gave evidence of arising from the photoinduced harpooning reaction between the sodium clusters, Nan, on the substrate, and (HCl)m adsorbed on the Nan. The translational energy distribution, its vibrational structure, and the angular distribution of H(g) gave information regarding the harpooning event. Translationally and vibrationally excited HCl(g) was shown, by resonance-enhanced multiphoton ionization (REMPI), to be formed as an alternate product; by way of (HCl)m..Nan/LiF(001) + 602 nm-->(HCl)m - 1 Nan/LiF(001) + HCl(g)(v > or = 0). PMID- 11272006 TI - Semiclassical treatment of reactions at surfaces with electronic transitions. AB - The semiclassical treatment of reactions at surfaces with electronic transitions based on the fewest-switches algorithm is compared with full quantum mechanical results. As a model system the ionization probability in I2 scattering from a diamond surface is chosen. In the calculations we treat the molecular distance from the surface and one surface oscillator coordinate explicitly. Furthermore, we also consider molecular rotation in the semiclassical calculations. The semiclassical results agree with the quantum results although some discrepancies remain, as far as the phase coherence is concerned. We identify energy transfer to molecular and surface degrees of freedom as a possible mechanism that could explain the experimental dependence of the ionization probability on the incident kinetic energy of the molecule. PMID- 11272007 TI - Production, purification and properties of microbial phytases. AB - Phytases (myo-inositol hexakisphosphate phosphohydrolase, EC 3.1.3.8) catalyse the release of phosphate from phytate (mycoinositol hexakiphosphate). Several cereal grains, legumes and oilseeds, etc., store phosphorus as phytate. Environmental pollution due to the high-phosphate manure, resulting in the accumulation of P at various locations has raised serious concerns. Phytases appear of significant value in effectively controlling P pollution. They can be produced from a host of sources including plants, animals and micro-organisms. Microbial sources, however, are promising for their commercial exploitations. Strains of Aspergillus sp., chiefly A. ficuum and A. niger have most commonly been employed for industrial purposes. Phytases are considered as a monomeric protein, generally possessing a molecular weight between 40 and 100 kDa. They show broad substrate specificity and have generally pH and temperature optima around 4.5-6.0 and 45-60 degrees C. The crystal structure of phytase has been determined at 2.5 A resolution. Immobilization of phytase has been found to enhance its thermostability. This article reviews recent trends on the production, purification and properties of microbial phytases. PMID- 11272008 TI - Applications of pectinases in the commercial sector: a review. AB - Pectinases are one of the upcoming enzymes of fruit and textile industries. These enzymes break down complex polysaccharides of plant tissues into simpler molecules like galacturonic acids. The role of acidic pectinases in bringing down the cloudiness and bitterness of fruit juices is well established. Recently, there has been a good number of reports on the application of alkaline pectinases in the textile industry for the retting and degumming of fiber crops, production of good quality paper, fermentation of coffee and tea, oil extractions and treatment of pectic waste water. This review discusses various types of pectinases and their applications in the commercial sector. PMID- 11272009 TI - Phytoextraction: a cost-effective plant-based technology for the removal of metals from the environment. AB - Phytoremediation is an emerging technology that uses plants to clean up pollutants (metals and organics) from the environment. Within this field of phytoremediation, the utilization of plants to transport and concentrate metals from the soil into the harvestable parts of roots and above-ground shoots is usually called phytoextraction. Most traditional remediation methods do not provide acceptable solutions for the removal of metals from soils. By contrast, phytoextraction of metals is a cost-effective approach that uses metal accumulating plants to clean up these soils. Subsequently, the harvestable parts, rich in accumulated metals, can be easily and safely processed by drying, ashing or composting. Some extracted metals can also be reclaimed from the ash, generating recycling revenues. Phytoextraction appears a very promising technology for the removal of metal pollutants from the environment and may be, at present, approaching commercialization. PMID- 11272010 TI - The potential for short rotation energy forestry on restored landfill caps. AB - This review examines the potential for producing biomass on restored landfills using willow and poplar species in short rotation energy forestry. In southern England, the potential production may be about 20 t ha(-1) of dry stem wood annually. However, actual yields are likely to be constrained by detrimental soil conditions, including shallow depth, compaction, low water holding capacity and poor nutritional status. These factors will affect plant growth by causing drought, waterlogging, poor soil aeration and nutritional deficiencies. Practical solutions to these problems include the correct placement and handling of the agricultural cap material, soil amelioration using tillage and the addition of organic matter (such as sewage sludge), irrigation (possibly using landfill leachate), the installation of drainage and the application of inorganic fertilizers. The correct choice of species and clone, along with good site management are also essential if economically viable yields are to be obtained. Further investigations are required to determine the actual yields that can be obtained on landfill sites using a range of management inputs. PMID- 11272011 TI - Remediation of dyes in textile effluent: a critical review on current treatment technologies with a proposed alternative. AB - The control of water pollution has become of increasing importance in recent years. The release of dyes into the environment constitutes only a small proportion of water pollution, but dyes are visible in small quantities due to their brilliance. Tightening government legislation is forcing textile industries to treat their waste effluent to an increasingly high standard. Currently, removal of dyes from effluents is by physio-chemical means. Such methods are often very costly and although the dyes are removed, accumulation of concentrated sludge creates a disposal problem. There is a need to find alternative treatments that are effective in removing dyes from large volumes of effluents and are low in cost, such as biological or combination systems. This article reviews the current available technologies and suggests an effective, cheaper alternative for dye removal and decolourisation applicable on large scale. PMID- 11272012 TI - Biofiltration--the treatment of fluids by microorganisms immobilized into the filter bedding material: a review. AB - Biofiltration is distinguished from other biological waste treatments by the fact that there is a separation between the microorganisms and the treated waste. In biofiltration systems the microorganisms are immobilized to the bedding material, while the treated fluid flows through it. In recent decades, a vast amount of literature has been written on single experiments involving the treatment of fluids by immobilized microorganisms. Several artificial immobilization methods have been examined and impressive results have been achieved in the treatment of fluids with one of the artificial immobilization methods the entrapment of microorganisms within polymer beads. This method, even though it needs to be improved, seems to have a future potential in commercial biofiltration systems. The methods of artificial immobilization of microorganisms within biofiltration systems have several advantages, but also suffer from several disadvantages in comparison to the treatment of fluids by naturally attached microorganisms. Understanding the mechanisms and forces responsible for the attachment of microbes to the bedding material, in attempt to improve this attachment, is of the utmost importance. Further improvement of the artificial entrapment of microorganisms within polymers will allow the exploitation of the advantages of this method in the treatment of fluids. The aim of this review essay is to introduce the main principles of two immobilization processes - the self attachment of microorganisms to the bedding material and the artificial entrapment of microorganisms within polymer beads. Both treatments of liquids and gases with each immobilization process are discussed. The advantages and disadvantages of each immobilization process are pointed out and different aspects of the fluid treatment with the two immobilization processes are compared. PMID- 11272013 TI - The treatment of pulp and paper mill effluent: a review. AB - The manufacture of paper generates significant quantities of wastewater; as high as 60 m3/tonne of paper produced. The raw wastewaters from paper and board mills can be potentially very polluting. Indeed, a recent survey within the UK industry has found that their chemical oxygen demands can be as high as 11000 mg/l. This paper reviews the processes involved in paper making and examines the effects which they could have on the environment. It also evaluates the treatment processes which are used to minimise these effects. In line with the majority of UK practice, it focuses mainly on aerobic biological treatment and, in particular, on the activated sludge process. This means that there is an in-depth discussion about the problems associated with filamentous bacteria and sludge "bulking". The paper also discusses the way in which anaerobic digestion can be applied to the treatment of liquid wastes from the manufacture of paper. PMID- 11272014 TI - Characteristics of wood ash and influence on soil properties and nutrient uptake: an overview. AB - Wood industries and power plants generate enormous quantities of wood ash. Disposal in landfills has been for long a common method for removal. New regulations for conserving the environment have raised the costs of landfill disposal and added to the difficulties for acquiring new sites for disposal. Over a few decades a number of studies have been carried out on the utilization of wood ashes in agriculture and forestry as an alternative method for disposal. Because of their properties and their influence on soil chemistry the utilization of wood ashes is particularly suited for the fertility management of tropical acid soils and forest soils. This review principally focuses on ash from the wood industry and power plants and considers its physical, chemical and mineralogical characteristics, its effect on soil properties, on the availability of nutrient elements and on the growth and chemical composition of crops and trees, as well as its impact on the environment. PMID- 11272015 TI - Utilization of by-products from the tequila industry. Part 2: Potential value of Agave tequilana Weber azul leaves. AB - The leaves of the agave plant are left in the field after harvesting the heads for tequila production. Different types of agave leaves were isolated, classified, and their content in the total plant determined. The usable fractions were collected and their properties determined. Of the total wet weight of the agave plant, 54% corresponds to the agave head, 32% corresponds to materials which could be usable for sugar and fiber production which leaves 14% of the wet plant without apparent utility. The fractions with higher total reducing sugars (TRS) content were the fresh fraction of partially dry leaves stuck to the head and the leaf bases with a TRS content of 16.1% and 13.1%, respectively. The highest TRS concentration (16-28%) is in the agave head which is used for tequila production. The leaves are 90-120 cm long and 8-12 cm wide and contain fiber bundles that are 23-52 cm long and 0.6-13 mm wide. The ultimate fiber length is approximately 1.6 mm with an average width of 25 microns. There are several types of leaf fibers that can be utilized depending on what part of the plant they come from and what product is desired. Agave leaf fibers were pulped using a soda pulping process and the pulp was hand formed into test sheets. Test sheets made from pulped agave leaf fibers had a breaking length comparable to paper made from both pine and eucalyptus fibers, but the tear index and burst index were lower than the other two papers. PMID- 11272016 TI - Organic matter components, aggregate stability and biological activity in a horticultural soil fertilized with different rates of two sewage sludges during ten years. AB - The effects of the application as fertilizer during ten years of two sewage sludges (aerobically and anaerobically digested, at rates of 400, 800, and 1200 kg of N/ha yr), on the aggregate stability and contents of related organic matter components, microbial biomass and levels of five enzymatic activities (alkaline phosphomonoesterase, phosphodiesterase, urease, arylsulphatase and dehydrogenase) were investigated. The application of both sludges at mid and high rates gave rise to significant increases of organic matter, humified substances and humic acids, but no effects on carbohydrates, microbial gums and aggregate stability were observed. As for biological activity in soils, the high variability of data led to a general absence of statistical significance despite the large differences between treatments observed. Significant increases of phosphodiesterase activity were nevertheless produced by the high rate of aerobic sludge and the mid and high rates of the anaerobic sludge. PMID- 11272017 TI - Seasonal variation in the biomass and agar yield from Gracilaria cervicornis and Hydropuntia cornea from Brazil. AB - Seasonality of biomass and agar yield from two agarophytes (G. cervicornis and H. cornea) was determined. The biomass from G. cervicornis was higher (390 g m-2) during the dry season and lower during the rainy season (129 g m-2). The data analysis for G. cervicornis revealed a significant seasonal variation (P < 0.05). H. cornea did not show a clear seasonal variation and was present only from March to August. The peak in biomass for this species was recorded in April (383 g m-2) and was significantly different from the other months (P < 0.05). The agar yield for G. cervicornis varied from 11% to 20%, with generally higher values recorded during the dry season. The agar yield showed a highly significant variation (P < 0.001). Agar yield from H. cornea ranged from 29% to 41%, with a peak recorded in June. The results above indicate that H. cornea can be considered a good candidate for commercial use. PMID- 11272018 TI - Nitrogen, carbon and phosphorus mineralization in soils from semi-arid highlands of central Mexico amended with tannery sludge. AB - Tannery sludge contains valuable nutrients and could be used as a fertilizer to pioneering vegetation in heavily eroded soils of the semi-arid highlands of central Mexico. Soil collected under and outside the canopy of mesquite (Prosopis laeviginata), huizache (Acacia tortuoso) and catclaw (Mimosa biuncifera), and cultivated with maize (Zea mays) and beans (Phaesolus vulgaris) was amended with 1.5 g tannery sludge kg-1 soil or 210 kg dry sludge ha-1 or left unamended. Amended and unamended soils were incubated aerobically for 70 days at 22 +/- 2 degrees C and CO2 production, available P, and inorganic N concentrations were monitored. The CO2 production rate, total C and P, available P, biomass C and P were larger under the canopy of the vegetation than outside of the canopy. The soils were depleted of N as more than 50 mg N kg-1 soil could not be accounted for in the first days of the incubation. Nitrification showed a lag, which lasted 28 days, and concentration of available P remained constant or increased slightly. Application of tannery sludge to soil increased CO2 production with 6.5 mg CO2 kg-1 soil d-1 and inorganic N with 30 mg N kg-1 soil after 70 days, but available P did not increase. Application of tannery sludge increased C and N mineralization and could thus provide valuable nutrients to a pioneer vegetation. Although no inhibitory effects on the biological functioning of the soil were found, further investigation into possible long-term environmental effects are necessary. PMID- 11272019 TI - Carbon and char residue yields from rapid pyrolysis of kraft black liquor. AB - The yields of char residue, fixed carbon, and inorganic carbonate were measured for oxidized black liquor char residues produced in a laboratory laminar entrained-flow reactor (LEFR) at heating rates of 4000-13,000 degrees C/s. The char residue yields at the end of devolatilization thus obtained decreased nearly linearly with temperature, from 75% at 700 degrees C to 58% at 1100 degrees C. There were explainable differences in the char residue yields from the liquor used in this study and those used in other studies. Char residue yields seemed to depend mainly on the temperature to which the particles or droplets were exposed and were not very sensitive to heating rate. Fixed carbon yields behaved similarly to those of the char residue. The fixed carbon remaining at the end of devolatilization decreased from 67% at 700 degrees C to about 45% at 1100 degrees C. The carbonate content in black liquor changed very little before and after devolatilization. PMID- 11272020 TI - Supercritical CO2 pretreatment of lignocellulose enhances enzymatic cellulose hydrolysis. AB - The supercritical carbon dioxide (SC-CO2) pretreatment of lignocellulose for enzymatic hydrolysis of cellulose was investigated. Aspen (hardwood) and southern yellow pine (softwood) with moisture contents in the range of 0-73% (w/w) were pretreated with SC-CO2 at 3100 and 4000 psi and at 112-165 degrees C for 10-60 min. Each pretreated lignocellulose was hydrolyzed with commercial cellulase to assess its enzymatic digestibility. Untreated aspen and southern yellow pine (SYP) gave final reducing sugar yields of 14.5 +/- 2.3 and 12.8 +/- 2.7% of theoretical maximum, respectively. When no moisture was present in lignocellulose to be pretreated, the final reducing sugar yield from hydrolysis of SC-CO2 pretreated lignocellulose was similar to that of untreated aspen. When the moisture content of lignocellulose was increased, particularly in aspen, significantly increased final sugar yields were obtained from enzymatic hydrolysis of SC-CO2-pretreated lignocellulose. When the moisture content of lignocellulose was 73% (w/w) before pretreatment, the sugar yields from the enzymatic hydrolysis of aspen and southern yellow pine pretreated with SC-CO2 at 3100 psi and 165 degrees C for 30 min were 84.7 +/- 2.6 and 27.3 +/- 3.8% of theoretical maximum, respectively. The SC-CO2 pretreatments of both aspen and SYP with moisture contents of 40, 57, and 73% (w/w) showed significantly higher final sugar yields compared to the thermal pretreatments without SC-CO2. PMID- 11272021 TI - Testing of alkaline and enzymatic hydrolysis pretreatments for fat particles in slaughterhouse wastewater. AB - Four pretreatments to hydrolyse and/or reduce the size of fat particles in slaughterhouse wastewater (SHW) were tested: sodium hydroxide and three lipases of plant, bacterial and animal (pancreatic) origin. Hydrolysing agents and SHW containing between 2.5 and 3 g/l of fat particles were mixed at room temperature for 4 h. Additions of 5-400 meq NaOH/l did not increase soluble COD (SCOD) in SHW, but the average particle size was reduced to 73% +/- 7% of the initial average particle size (D(in)) at NaOH concentrations ranging from 150 to 300 meq/l. Pretreatment with pancreatic lipase PL-250 reduced the average particle size to a maximum of 60% +/- 3% of D(in). As D(in) was decreased from 359 to 68 microns, the enzyme concentration required to obtain the maximum particle size reduction increased from 200 to 1000 mg/l. A 4-h pretreatment with PL-250 also increased the free long-chain fatty acid (LCFA) concentration to a maximum of 15.5 mg/l, indicating some solubilization of the pork fat particles in SHW. SCOD was not significantly increased by the pretreatment, but SCOD was not found to be a good indicator of enzymatic lipolysis because of enzyme adsorption on the fat particle surface. Pancreatic lipase appeared more efficient with beef fat than pork fat, possibly because beef fat contains less polyunsaturated fatty acids than pork fat. The bacterial lipase LG-1000 was also efficient in reducing average fat particle size, but high doses (> 1000 mg/l) were required to obtain a significant reduction after 4 h of pretreatment. SCOD was not increased by pretreatment with LG-1000. No particle size reduction or changes in SCOD were noted after 4 h of pretreatment with the plant lipase EcoSystem Plus. It was concluded that PL-250 was the best pretreatment to hydrolyse fat particles in SHW. However, its impact on the efficiency of a downstream anaerobic digestion process remains to be tested. PMID- 11272022 TI - Enzyme-catalyzed synthesis of alkyl-beta-glucosides in a water-alcohol two-phase system. AB - The enzymic synthesis of alkyl-beta-glucosides by water-immiscible alcohols was studied in stirred flasks as well as in a tubular enzymatic reactor. In the first case, direct alkylation of beta-D-glucose from hexanol using immobilized beta glycosidase gave a higher conversion yield and final product concentration than that using the enzyme in its free state (yield 10 against 8% mol/mol, concentration 2 against 1.6 g/l). Direct glycosylation of beta-D-glucose from hexanol resulted in a higher reaction performance (yield 10%) than that from octanol (yield 5%). However, the two different incubation temperatures tested (37 degrees C and 50 degrees C), showed no significant differences concerning final product concentrations. The more interesting results were obtained by transglycosylation of methyl-1-beta-glucose from hexanol, with a conversion yield of 21% mol/mol (product amount 4 g/l). However, the transgalactosylation of lactose from hexanol, catalyzed by a fungal beta-galactosidase, showed only a feeble reactivity. The feasibility of enzymic alkylation was also tested in a tubular enzymatic reactor; hexyl-1-beta-glucoside was produced via direct glycosylation from hexanol catalyzed by free beta-glycosidase with a final concentration 1.3-2.3 g/l and a yield varying between 11% and 20% mol/mol. PMID- 11272023 TI - Constitutive dechlorination of chlorinated ethenes by a methanol degrading methanogenic consortium. AB - The ability of granular methanogenic sludge to dechlorinate chloroethenes was investigated with unadapted sludge from an upflow anaerobic sludge blanket (UASB) reactor fed with methanol. The sludge degraded chlorinated ethenes, but the degradation rates were low. The addition of primary substrate was necessary to sustain dechlorination. The dechlorinating activity seemed to be constitutively present in the anaerobic bacteria. Usually, one chlorine atom was removed via reductive hydrogenolysis. Only trichloroethene (TCE) was converted to substantial amounts of vinylchloride (VC). 1,1-Dichloroethene (1,1DCE) was observed to be an important intermediate in the dechlorination by unadapted granular sludge, although previously this compound had not been commonly observed. Furthermore, the dechlorination of 1,1DCE was faster than the dechlorination of the other chloroethenes. PMID- 11272024 TI - Studies on carboxymethyl cellulase produced by an alkalothermophilic actinomycete. AB - A novel alkalothermophilic actinomycete having optimum growth at pH 9 and 50 degrees C was isolated from self-heating compost from the Barabanki district of Uttar Pradesh, India. Based on its morphology, susceptibility of spores to heat and novobiocin, guaninecytosine content of chromosomal DNA and cell wall composition, the organism was classified under Thermomonospora. The alkalothermophilic actinomycete produced 23 IU/ml carboxymethyl cellulase (CMCase). The CMCase was purified by fractional ammonium sulphate precipitation followed by cellulose affinity chromatography and Sephacryl S-200 gel filtration. The CMCase had a molecular weight of 38 KD and pI of 4.1. The enzyme exhibited optimum activity at pH 5 and temperature 50 degrees C. The CMCase showed pH stability in the range 7-10. The enzyme retained 100% activity at 50 degrees C for 72 h and had half-lives of 7 and 3 h at 60 degrees C and 70 degrees C, respectively. The CMCase was stable in the presence of commercial detergents such as Ariel, Henko and Surf Excel, indicating its potential as an additive to laundry detergents. PMID- 11272025 TI - Algal growth control by a barley straw extract. AB - In recent years, there has been an apparent increase in the occurrence of harmful algal blooms occurring in potable waters. The potential of a simple barley straw extract to inhibit algal growth was assessed. Algal growth in lakewater was inhibited by the addition of barley straw (1% w/v), with the chlorophyll a concentration remaining below the original level (40 micrograms l-1) throughout the experiment. In contrast, in the presence of wheat straw, algal biomass increased, reaching a final chlorophyll a concentration of 1160 micrograms l-1 after 28 days. Analysis of the remaining particulate straw at the end of the experiment showed that the lignin content of barley straw had increased significantly from 10-33% (w/w). Further, a preparation of a simple aqueous extract from the decomposed-barley straw was found to inhibit the cyanobacteria Microcystis sp. and the algal species Scenedesmus, with chlorophyll a levels some 10-fold lower than in untreated flasks. This study shows that a decomposed-barley extract, even in a very dilute concentration (0.005%) was capable of inhibiting the growth of Microcystis sp., a commonly occurring cyanobacterium which produces the toxin microcystin and has been responsible for some of the most serious pernicious algal blooms in the UK. PMID- 11272026 TI - Molecular weight distribution of Pinus radiata kraft mill wastewater treated by anaerobic digestion. AB - Kraft mill is responsible for massive discharge of highly polluted effluents. The main characteristics of this effluent are high toxicity and low biodegradability due to tannin, lignin and chlorophenol compounds. The composition may vary dramatically depending, for instance, on the utilised feedstock and process. The purpose of this work was to investigate the molecular weight distribution of Pinus radiata kraft pulping wastewater treated by anaerobic digestion by using two types of anaerobic reactors: fixed bed and sludge blanket. Anaerobic sludge blanket (UASB) and anaerobic filter (AF) were operated. In both reactors, the total alkalinity ranged between 1.0 and 1.5 g CaCO3/l, while the organic load rate (OLR) was increasing during operation from 1.2 to 3.3 gCOD/l d. COD and total phenolic compounds (UV215) removal ranged between 30-50% and 13-20%, respectively, while the BOD5 removal ranged 60-90%. However only a partial biodegradation (10-43%) of tannin and lignin was observed. Results from ultrafiltration analyses indicated that the fraction with a molecular weight (MW) < 1000, COD and colour decreased after anaerobic treatment, but the total phenolic compounds increased. In the 1000 < MW < 10,000 fraction, there was no change in COD, UV215 and colour. In the > 10,000 MW fraction, colour and COD fraction increased by 14% and 5%, respectively, after anaerobic treatment. It can be concluded from this study, that treatment with UASB or AF reactors is not enough, under the conditions tested, for a large COD removal from Pinus radiata wastewater. PMID- 11272027 TI - Simultaneous saccharification and fermentation of lignocellulosic wastes to ethanol using a thermotolerant yeast. AB - Simultaneous saccharification and fermentation (SSF) studies were carried out to produce ethanol from lignocellulosic wastes (sugar cane leaves and Antigonum leptopus leaves) using Trichoderma reesei cellulase and yeast cells. The ability of a thermotolerant yeast, Kluyveromyces fragilis NCIM 3358, was compared with Saccharomyces cerevisiae NRRL-Y-132. K. fragilis was found to perform better in the SSF process and result in high yields of ethanol (2.5-3.5% w/v) compared to S. cerevisiae (2.0-2.5% w/v). Increased ethanol yields were obtained when the cellulase was supplemented with beta-glucosidase. The conversions with K. fragilis were completed in a short time. The substrates were in the following order in terms of fast conversions: Solka floc > A. leptopus > sugar cane. PMID- 11272028 TI - National inventory of organic wastes for use as growing media for ornamental potted plant production: case study in Spain. AB - An inventory of materials suitable for use as growing media for ornamental potted plant production in Spain has been prepared. Special attention has been paid to solid organic wastes generated by production, industrial and consumer activities. Information obtained from this study has been organised into two data bases. Data base 1 contains the "General Characteristics" file of more than 105 materials. In this file, data are available regarding generation points, material availability, uses, cost, disposal expenses, etc. Data base 2 is comprised of the "Specific Properties" file of 63 materials selected from data base 1. The main physical, chemical and biological properties of these materials as container media have been characterised, and the results obtained have been compiled. Finally, a computerised data bank has been created which can be found in the home page of the Spanish Ministry of Agriculture, Fisheries and Food (http://agritel2.mapya.es/sustratos/). PMID- 11272029 TI - The rational use of antibiotics in the treatment of brain abscess. AB - The Working Party was instituted to investigate the rationale of therapeutic antibiotic usage in patients with brain abscess and to make recommendations for current practice. A systematic review of English language publications on brain abscess over the last 25 years was carried out using electronic databases and secondary sources, and data were evaluated. Few publications were identified where the microbiological procedures were adequately described and many authors continue to report sterile pus in a proportion of cases. The vast majority of reports were retrospective neurosurgical assessments in which details of laboratory procedures and antibiotic regimens were missing. There are no published reports of controlled clinical trials or comparative therapeutic studies. The recommendations made by the Working Party are based on relevant published information and the expertise of Working Party members. Recommendations vary according to the location of the abscess which reflects the likely source of the infection and therefore the bacterial types most likely to be present in aspirated pus. Bacteria with multiple resistance to antimicrobial agents do not feature significantly in cases of brain abscess. PMID- 11272030 TI - Combined transhorizontal-supracerebellar approach for microvascular decompression of trigeminal neuralgia. AB - A combined approach for microvascular decompression of trigeminal neuralgia is reported. Arachnoid dissection of the cerebellar horizontal fissure (transhorizontal approach) allowed easy identification of the root exit zone of the trigeminal nerve. The superior semilunar lobule was pulled back rostrally so that retraction of the acoustic nerve was minimal. After identification of the offending vessels, the supracerebellar artery was mobilized and dissected out through the supracerebellar route, which was also less invasive to the acoustic nerve than the standard approach. By combining these two approaches, the whole surface of the trigeminal nerve can be observed easily. Thus, the offending vessels can be readily identified, mobilized and moved away from the trigeminal nerve with minimal retraction of the acoustic nerve. PMID- 11272031 TI - A clinical audit of the Hakim programmable valve in patients with complex hydrocephalus. AB - The objective of the investigation was to determine the pattern of use of the Hakim (Medos) programmable valve implanted in patients with complex hydrocephalus and their clinical outcome. A prospective audit of patients with complex hydrocephalus undergoing Hakim programmable valve implantation between 1989 and 1994 in the United Kingdom and Ireland, was followed-up for a minimum of 5 years. Surgical practice and complications were audited together with clinical outcome. One-hundred-and-thirty-nine patients (80 male, 59 female; mean age 43.4 years; median 47 years; range 1 month-84 years) with complex hydrocephalus due to a wide range of aetiologies were implanted with the Hakim programmable valve. Eighty eight (63%) had large or massive ventricles prior to implantation; seven (5%) were slit. Fifty-five (40%) had previously been shunted with a fixed pressure system. One-hundred-and-thirty-one (94%) of the Hakim programmable shunts were ventriculoperitoneal; four (3%) ventriculoatrial; two (1.4%) cystoperitoneal; and two (1.5%) lumboperitoneal. The initial opening pressure selected ranged from 50 to 200 mmH2O (median 120). Valves were reprogrammed on average 1.7 times with 143 reprogrammings in the first year after implantation; 67 in the second; 19 in the third; three in the fourth; two in the fifth. Forty-nine (36%) valves were never reprogrammed after implantation. During the 5 years audit period, there were 70 (50%) shunt revisions, 40 of which were performed within 1 year of implantation. Thirty-six (27%) shunts were removed. There were 24 (18%) shunt infections. Subdural collections were identified in 37(27%) patients after Hakim programmable valve implantation; 10 (27%) required surgical drainage. Five (3.7%) patients developed symptomatic slit ventricles after Hakim programmable valve implantation. Headache was improved following reprogramming in 27(71%) of the 38 patients with refractory headache. After Hakim programmable valve implantation, patients underwent an average of 4.6 CT scans (range 1-25); 0.3 MRI (range 1-5) and 1.8 skull radiographs (range 1-20). The mean hospital stay per patient over 5 years was 26 days (range 1-110 days). Five years after implantation, the Glasgow Outcome scale was favourable in 64% of patients. The Hakim programmable valve is useful in the management of patients with complex hydrocephalus and may reduce the need for shunt revision for headache. Non-haemorrhagic, post-shunting, subdural collections identified on routine postoperative CT may be treated by reprogramming. PMID- 11272032 TI - The effects of chronic nitric oxide synthase suppression on glioma pathophysiology. AB - Nitric oxide synthase (NOS) is strongly expressed in glioma and has an important role in tumour blood flow (TBF) regulation. Whether manipulation of NOS function within a tumour can have any therapeutic effect is unknown. This study therefore evaluated the pathophysiological effects of chronic systemic NOS inhibition on experimental rodent glioma blood flow, growth and necrosis. To determine the duration and pathophysiological effects of systemic NOS inhibition, Ng-nitro-L arginine methyl ester (L-NAME) was given to rats bearing C6 glioma acutely (single dose i.v., 30 mg kg) or for either 4 or 7 days (i.p. 75 mg kg day) prior to study. TBF and local cerebral blood flow (LCBF) were measured using C14 iodoantipyrine quantitative autoradiography. Tumour volume, tumoural necrosis and tumoural NOS were measured using conventional neuropathology and immunocytochemistry. Acute and 4-day L-NAME administration produced significant TBF reductions (-48 and -39%, respectively) with less marked changes in LCBF (-35 and -15%, respectively). Seven-day L-NAME administration reduced tumour volume (p = 0.12), increased tumoural necrosis (p < 0.05), but immunohistochemistry showed no difference in tumoural NOS expression. These results confirm that NOS has a significant role in the pathophysiology of experimental glioma, and that in this glioma model the effects of chronic systemic NOS inhibition are, for the period under study, predominately anti-tumoural. Whether chronic NOS inhibition is useful as an adjunct in glioma therapy or provides the opportunity for novel therapeutic approaches requires further study. PMID- 11272033 TI - Fatal posterior fossa pneumocephalus due to hydrogen peroxide irrigation of lumbar wound. AB - Fatal brain stem failure developed suddenly in a 40-year-old male undergoing irrigation of an infected wound consequent to lumbar disc space infection. CT of head revealed posterior fossa pneumacephalus compressing the brain stem, which most likely developed from ingress of nascent oxygen under pressure through a defect in the thecal sac. PMID- 11272034 TI - Effective diagnosis of spinal cord compression using telemedicine. AB - We report the case of a man with an acute cervical cord lesion which was diagnosed after a teleconsultation with a neurologist at a distant neurological centre. We show that the ability to assess patients clinically using real-time videolinks is also of use in detecting those patients who would benefit from specialist neurosurgical intervention, but who might otherwise be denied it because of their location. PMID- 11272035 TI - A method for the resection of cranial tumours and skull reconstruction. AB - A new technique for the resection of cranial tumours and subsequent reconstruction using stereolithographic (SL) biomodelling and customized cranioplastic implants has been developed. The technique is based on a custom model of the tumour and surrounding skull from which the resection of the tumour and shape of the cranioplasty can be determined. A patient with a hyperostotic fronto-orbital meningioma was selected. CT was performed and SL biomodels manufactured. The surgeon marked the resection margin on the biomodel and a customized resection template was fashioned. The tumour was then resected from the biomodel and a customized acrylic implant was manufactured to reconstruct the defect. At surgery the tumour was exposed in a routine fashion and the template used to mark the resection margin. Once resected, the defect was reconstructed with the custom cranioplastic implant. The technique facilitated accurate surgical resection of the tumour and subsequent reconstruction. The surgeon reported several advantages of the technique including increased confidence, reduced operating time (at least 1 h), excellent cosmetic results, accuracy, and simplicity. The patient reported that the opportunity to see the biomodel, template and implant improved her understanding of the procedure. PMID- 11272036 TI - Endoscopic fenestration of a symptomatic cyst of the septum pellucidum. AB - We present the case of a 25-year-old woman suffering from chronic severe headaches and syncope associated with an expanding cyst of the septum pellucidum. After a simple endoscopic fenestration of the cyst the symptoms resolved, resulting in an excellent clinical outcome. This case shows that simple endoscopic fenestration may produce a rapid and maintained symptomatic improvement in patients with an expanding cyst of the septum pellucidum. PMID- 11272037 TI - The treatment of 'acquired tonsillar herniation' in pseudotumour cerebri. AB - Acquired tonsillar herniation and syrinx formation are recognized complications of a lumboperitoneal shunt used to treat conditions of increased CSF volume. Treatment of this complication is by ventriculoperitoneal shunt or foramen magnum decompression. We describe the first case of acquired tonsillar herniation in a pseudotumour cerebri (following lumboperitoneal shunt insertion) that responded partly to ventriculoperitoneal shunt insertion and more completely to foramen magnum decompression. Our case demonstrates that a ventriculoperitoneal shunt is not always sufficient in treating this complication and reversing visual deterioration. Foramen magnum decompression may therefore be a sight saving procedure in pseudotumour cerebri with acquired tonsillar herniation and may be important in understanding the pathogenesis of conditions of increased CSF volume. PMID- 11272038 TI - Spontaneous cerebrospinal fluid rhinorrhoea and pneumocephalus following temporal lobectomy for epilepsy. AB - A 30-year-old female presented with headache, CSF rhinorrhoea, mild right facial weakness, 2 months following temporal lobectomy for epilepsy. CT revealed marked intraventricular pneumocephalus with breached air cells in the pneumatized lower part of temporal bone. The dural and bony defects repaired successfully with complete resolution of the pneumocephalus. PMID- 11272039 TI - Arachnoid cyst with spontaneous rupture into the subdural space. AB - Spontaneous rupture of an arachnoid cyst into the subdural space is an unusual complication. Only six cases have been reported in the literature. We report here an additional case and review the literature concerning arachnoid cysts. The possible pathogenesis of this condition is discussed. PMID- 11272040 TI - Epidermoid cyst of the cavernous sinus: clinical features, pathogenesis and treatment. AB - Epidermoid cyst may rarely arise within the lateral wall of the cavernous sinus (CS) and produce clinical symptoms mimicking the Tolosa-Hunt syndrome. Pathogenically, it is suggested that some of the subpopulation of cells arising from the neural crest which give rise to skin, may remain entrapped in the meninges around the nerves in the lateral wall in an uncompleted stage of maturation and may develop epidermoid cysts. A case is presented of a 68-year-old man with a 4-week history previously diagnosed as Tolosa-Hunt syndrome. MRI is the investigation of choice. Surgical resection can result in excellent recovery of preoperative deficits of cranial nerves (CN) III through VI. PMID- 11272041 TI - Radiation-induced cavernous angioma mimicking metastatic disease. AB - Patients with carcinoma of the lung typically have a limited life expectancy especially after developing metastatic disease in the brain. New enhancing lesions in the brain are usually felt to represent new areas of metastasis. Recently, there have been several case reports of cavernous angiomas appearing years after radiation to the brain, typically in children. We present a case of a 41-year-old gentleman with carcinoma of the lung with metastasis to the brain who received postoperative radiation. Five-and-a-half years later he presented with a new enhancing lesion of the brain with surrounding vasogenic oedema, thought to represent a metastatic tumour. It proved is the a radiation-induced cavernous angioma. PMID- 11272042 TI - Pituitary tumour presenting with trigeminal neuralgia as an isolated symptom. AB - Invasion of the cavernous sinus by pituitary adenoma may cause involvement of cranial nerves III, IV, V and VI. However, trigeminal neuralgia as an isolated, initial symptom is very unusual. A patient with a pituitary adenoma, who had intractable trigeminal neuralgia as the only complaint which resolved following surgical removal is presented. PMID- 11272043 TI - Bacillus cereus meningitis complicating cerebrospinal fluid fistula repair and spinal drainage. AB - Non-anthrax Bacillus species are rare, but serious causes of bacterial meningitis in those either immunocompromised or treated with CSF diversion. Although resistant to first-line antibiotics, they usually respond to chloramphenicol. We report a case of fulminant Bacillus cereus meningitis that complicated lumbar spinal drainage which proved resistant to all first-line antibiotics including chloramphenicol. PMID- 11272044 TI - Torticollis due to atlanto-axial rotatory fixation following general anaesthesia. AB - Atlanto-axial rotatory fixation is uncommon, but should be suspected in any patient developing a torticollis during the recovery period of an operation performed on the head and neck under general anaesthesia. The present case report shows that the condition can occur in adults as well as children. A high degree of suspicion is required to instigate appropriate imaging. CT provides unequivocal proof and will lead to early successful treatment. PMID- 11272046 TI - Migration of a ventricular access device. PMID- 11272045 TI - Rapid growth in a cavernoma. AB - The availability of magnetic resonance imaging has greatly increased the detection of cavernous malformations of the central nervous system in both symptomatic and asymptomatic patients. These lesions may be responsible for previously unexplained neurological events or may even have been incorrectly diagnosed. We describe a patient presenting with focal neurological signs in whom an initial diagnosis of malignant glioma had been made. Following excision, the pathological diagnosis was cavernoma. This vascular lesion has continued to show rapid growth and aggressive behaviour despite multiple surgical resections. The indications for operative and non-operative intervention will be discussed. PMID- 11272047 TI - Foreign body granuloma after craniotomy for tumor: a diagnostic dilemma. PMID- 11272048 TI - Image-guided transoral clipping of basilar aneurysm. PMID- 11272049 TI - Cystic lesions associated with intracranial meningiomas. Report of three cases. PMID- 11272050 TI - Sacral and skull base chordoma: metastases or second primary? PMID- 11272051 TI - Comparative efficacy of intravenous cefotaxime and trimethoprim/sulfamethoxazole in preventing infection. PMID- 11272052 TI - Management of tuberculomas of the craniovertebral junction. PMID- 11272053 TI - Neuroendoscopy combined with frameless neuronavigation. PMID- 11272054 TI - [Color and duplex Dopplerometry in the prenatal diagnosis of congenital developmental defects]. AB - OBJECTIVE: Impact of Doppler system in detection of congenital malformations during prenatal diagnosis. DESIGN: Prospective clinical study. SETTING: Department of Medical Genetics and Foetal Medicine, Department of Obstetrics and Gynaecology University Hospital Olomouc. METHODS: In 34 pregnancies with prenataly diagnosed congenital malformation the umbilical artery pulsatility index (PI) and middle cerebral artery PI were assessed. RESULTS: Abnormal Doppler patterns were obtained in 18 patients (chromosomal abnormalities--3x, gastroschisis--2x, diaphragmatic hernia--2x, urinary tract abnormalities--2x, internal hydrocephalus--4x, microcephaly, anencephaly, encephalocoele, non-immune hydrops of the fetus, thoracopagus--1x). CONCLUSIONS: In accordance to the literature abnormal Doppler patterns in umbilical artery were obtained in trisomy 21 foetuses. The use of umbilical artery PI or ductus venosus PI in the first trimester of pregnancy as an additional parameter for screening purposes needs to be confirmed by further investigation (needs to be further explored). Pulsed Doppler waveform analysis gives a possibility to appreciate the functional state of the foetus. In our patient group abnormal Doppler patterns were found in such substantial congenital malformations that they indicated pregnancy termination themselves. On the other hand in several similar morphological defects normal Doppler patterns were found. In these cases, in regard to the type of malformation, the Doppler appreciation of foetal functional state was not prognostically significant. PMID- 11272056 TI - [Effect of maternal O2 inhalation on oxygen saturation in the parturient (SpO2) and the fetus (FSpO2)]. AB - OBJECTIVE: The aim of the study was to evaluate the impact of different concentrations of inspirated O2 on the SpO2 values of the foetus and the mother during the 1st and 2nd stages of labor. DESIGN: Prospective study. SETTING: 1st Department of Gynecology and Obstetrics, Medical Faculty of Masaryk University, Brno. METHODS: 17 non-risk patients were enrolled in the study. The SpO2 levels of the mother and the foetus were monitored simultaneously in 10 min intervals with the room air inspiration entrainment O2 mask and the inflatable face mask during 1st and 2nd stages of labor. RESULTS: No changes in maternal SpO2 values were revealed in the three different O2 inspiration regimens neither in the 1st, nor in the 2nd stages of labor. The mean value was 98% +/- 1.6 SD. As for the foetus the SpO2 values were increased by 6% (+/- 7.9 SD) after 40% concentration of O2 and 7.7% (+/- 8.8 SD) after 98% O2 inspiration during the 1st stage of labour. These values have decreased promptly after the cessation of the O2 inspiration. The mean SpO2 values at room O2 concentration were 46.3% +/- 7.9 SD for the 1st stage of labor and 43.7% +/- 4.8 SD for the 2nd stage of labor. CONCLUSION: The O2 inspiration during the labor has no impact on the maternal SpO2 values but increases the SpO2 values of the foetus during the 1st stage of labor. We were not able to evaluate the impact of the O2 inspiration on foetal SpO2 values during the 2nd stage of labor. PMID- 11272055 TI - [Prenatal diagnosis of congenital defects in the Czech Republic in 1998]. AB - OBJECTIVE: Presentation of prenatal diagnostics results in the year 1998 and their comparison with the period 1990-1997. DESIGN: A retrospective study. SETTING: Subchair of Medical Genetics, Postgraduate Medical Institute, Videnska 800, Praha 4, Czech Republic. METHODS: Processing of prenatal diagnostics data obtained from departments of Medical Genetics from all the Czech Republic. The comparison with livebirths with congenital malformation was made possible using the data from Statistics Reports "Congenital malformation in a foetus/newborn" which are filed in the Institute of Health Information and Statistics. For a comparison on the international level the data published by the International Clearinghouse for Birth Defects Monitoring Systems were used. RESULTS: 553 foetuses with congenital malformation were diagnosed in 1998. In the same year, 406 pregnancies were terminated from this reason. Totally, 445 particular malformations were found in the above mentioned foetuses. Most often, inborn chromosomal aberrations (and Down Syndrome especially) were detected. Down Syndrome itself was diagnosed in 101 cases, out of which in 98 cases the pregnancy was terminated. Taking into account 61 cases of Down Syndrome in livebirths in 1998, the sensitivity of prenatal diagnostics of this defect is 61.64%. Finally, a comparison of our results with data from different congenital malformation registers all over the world was made. CONCLUSION: An increase of the prenatal diagnostics of congenital malformation as well as its improvement has taken place in the Czech Republic in last years. This improvement is accompanied by an increase of the number of prenatally diagnosed cases and also by an increase of the number of terminated pregnancies as its consequence. An incidence of severe congenital malformations therefore decreases in livebirths. An efficiency of prenatal diagnostics is fully comparable to the results published by major congenital malformation registers worldwide. PMID- 11272057 TI - [Trends in the reproductive health of women in the Czech Republic 1993-1997. V. International comparisons]. AB - OBJECTIVE: 1) Overall objective is to detect changes in reproductive health of women due to changes in the system of social and health care. 2) The particular objective of this part of the study (part V) is to compare the incidence of induced abortions and contraceptive use in the Czech Republic and selected developed countries during last 20 years. DESIGN: Retrospective comparative epidemiological study. SETTING: Institute for the Care of Mother and Child, Praha 4-Podoli. METHODS: Input data for the Czech Republic use came from the results of the four previous parts of the study. The data from similar studies in selected countries were divided into two groups. Seven countries of Central Europe formed the first group, the second group consisted of 14 countries of Western and Northern Europe and 4 developed countries over-the-ocean. The both groups were ordered according to the reliability of reported data. RESULTS: The cause of higher number of induced abortions and lower number of women using effective contraception in the Czech Republic compared to other developed countries falls into sixties when different social approach towards induced abortion legislation and concurrent development of planned parenthood began. This is being documented by comparisons of data on use of induced abortions and contraceptive use among women of different age groups, while using internationally accepted indicators. Although the number of induced abortions did not change substantially in western countries during last 20 years (there has been even slight increase in some of them since 1993), the number of induced abortions in the Czech Republic decreased markedly, and this fact, together with increased use of contraception since 1993 led to decrease of difference in between these two variables, compared to western countries. CONCLUSIONS: The 2-3 fold higher number of induced abortions in the Czech Republic than in countries of the second group before 1993 decreased substantially; there were only 19.3 induced abortions per 1000 women in the Czech Republic in 1997 compared to average of 14.1 induced abortions in developed countries. Use of effective contraceptive methods is lower in the Czech Republic due to low use of sterilization. PMID- 11272058 TI - [Homocysteinemia--its significance in gynecology and obstetrics]. AB - OBJECTIVE: Metabolic study on plasmatic levels of homocysteine (Hcy) in healthy women during normal or pathological pregnancy accompanied with corresponding levels of Hcy in amniotic fluid and foetal sera. Increased levels of Hcy- hyperhomocysteinaemia is respected as an independent risk factor accelerating the early development of vessel damage and causing the neural tube defects (NTD). DESIGN: Basic study to get our own data about Hcy in Czech healthy and population at risk of pregnant and non-pregnant women. SETTING: Department of Obstetrics and Gynaecology, 1st Faculty of Medicine, Charles University, Prague. METHODS: Total homocysteine in plasma, amniotic fluid and foetal sera was estimated by chromatographic method with use of fluorescence detection. RESULTS: Normal homocysteine in preclimacteric healthy nonpregnant women is: 9.7 +/- 1.6 mumol/l with evident age-dependence. In healthy climacteric women are higher levels of Hcy (corresponding to the men values): 11.8 +/- 2.6 mumol/l. After use of hormonal contraceptives the plasmatic levels of Hcy decrease: 7.2 +/- 2.0 mumol/l. In physiological pregnancies Hcy reachs the lowest values: 4.4 +/- 1.7 mumol/l with any evident oscillations during pregnancy. In women in childbed period was Hcy 8.4 +/- 2.1 mumol/l observed. In pathological pregnanciesare its levels slightly elevated: 6.3 +/- 2.1 mumol/l, most evident in placental abruptions: 7.5 +/- 1.7 mumol/l. In pregnant women with susp. results of screening on M. Down only unsignificant increase of Hcy was observed: 6.12 +/- 2.4 mumol/l. In amnial fluids of healthy pregnant women are levels of Hcy are quite low: 4.1 +/- 1.2 mumol/l with any oscillations during pregnancy. In foetal sera of pregnancies at risk (NTD, susp., trisomy, inborn errors of metabolism): 3.6 +/- 1.4 mumol/l of Hcy was detected. The foetoplacental quotient for Hcy is 0.62. CONCLUSION: Average values for Hcy were established in physiological as well as in pathological pregnancies and till now only limited diagnostic significance has been observed. The hyperhomocysteinaemia mentioned in previous papers was not in NTD observed because our pregnant patients were regularly supplemented with all critical vitamins (folate, B6, B12). PMID- 11272059 TI - [Noninvasive screening for genital chlamydiosis in adolescents in Brno]. AB - OBJECTIVE: To run a screening for genital chlamydiosis in adolescents living in the town Brno as the first action of this type in the Czech Republic and to use the results of the screening for the elaboration of recommendations for running similar actions on the national scale. DESIGN: Prospective epidemiological study. SETTING: Veterinary Research Institute, Brno; Institute of Clinical Biochemistry, 1st Faculty of Medicine, Charles University and General Faculty Hospital, Prague; Section of Public Health, Municipal Authorities of Brno; Bioplus Ltd., Brno; Regional Hygienic Services, Brno; Faculty Hospital Brno; with technical assistance of teachers and students of two Medical Assistant Schools in Brno. METHODS: Students (337 females and 15 males) of two Medical Assistant Schools, older than 18 years, were used as probands within the study. Sediments of the first portions of urine collected from the individual probands were tested for the presence of Chlamydia trachomatis using the direct fluorescent test, ELISA, and the ligase chain reaction. RESULTS: Positive reactions in any of the three tests were found in 31 of the 352 probands (8.8%). Positive and doubtful reactions in the direct fluorescent antibody test were obtained in 11 (35.5%) and 3 (9.6%) of the 31 reactors, respectively. The corresponding values for ELISA were 9 (23.0%) and 8 (25.8%), respectively, and those for the ligase chain reaction 3 (16.6%) and 3 (16.6%), respectively. The overall prevalence of 8.8% is higher than the European mean. CONCLUSIONS: The first limited screening for genital Chlamydia infections in the Czech Republic was run in Brno. Urinary samples were collected from 337 females and 15 males aged approximately 18 years. The presence of Chlamydia trachomatis in the urinary sediment was demonstrated by the direct fluorescent test, ELISA, and the ligase chain reaction. The established prevalence of 8.8% exceeded the European mean (3.9%). PMID- 11272060 TI - [[Endometrial ablation--prospective 5-year follow-up study]. AB - OBJECTIVE: Objective of the study is to evaluate the effectiveness of endometrial ablation in patients with persistent uterine bleeding who are unresponsive to conservative therapy. DESIGN: Prospective clinical study. SETTING: Department of Obstetrics and Gynaecology, 1st Medical Faculty, Charles University and General Faculty Hospital in Prague, Czech Republic. METHODS: 100 women with intractable uterine bleeding were subjected to undergo endometrial ablation. 44 patients were treated preoperatively not only with danazol or progestins but also with Norethisteron acetas to stop the acute bleeding preoperatively and 65 gave no preoperative drug administration. Under appropriate anesthesia the cervix was dilated to 100 mm and the uterine cavity was distended with Purisol (Sorbitol and Manitol). Roller-ball coagulation technique combined with loop highfrequency endoresection was used in most of the patients (85 patients). The findings of small uterine myoma(s) were not consider as a contraindication of the endometrial ablation. RESULTS: At 51-2 months 42 (42%) of patients reported amenorrhea, 51 (51%) hypomenorrhea, 5 (5%) eumenorrhea and 2 (2%) nochange. The mean time to complete operation was 30 minutes (range 15-45 minutes). The procedure was completed in all 100 women and we had no serious complications. CONCLUSION: It is concluded, that endometrial ablation is safe and effective hysteroscopic procedure in the cases of abnormal uterine bleeding for women with normal uterine morfological findings or small uterine myoma(s) considering the follow up 51-2 month of the study. PMID- 11272061 TI - [Liver function tests during administration of triphasic contraceptives containing Norgestimate]. AB - OBJECTIVE: To evaluate serum levels of liver enzymes and bilirubin before and after six cycles of use of a triphasic oral contraceptive containing 35 micrograms of ethinylestradiol and 180/215/250 micrograms of norgestimate (Pramino, Janssen-Cilag). DESIGN: A prospective, open-label, non-comparative, multicentric phase IV study. SETTING: Department of Obstetrics and Gynaecology, 1st Medical Faculty, Charles University, Prague. METHOD: Before the start and after six cycles of Pramino use, the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyltransferase (GMT), and bilirubin were determined in women. As the analyses were performed in different laboratories, the evaluation involved: 1. Subgroups of women examined in laboratories with the same reference range; 2. A group of women examined in laboratories with the upper reference range within the mean +/- SD interval; 3. Percentage variations of the measured values from a concrete upper reference limit. RESULTS: When evaluating subgroups of women from laboratories with the same standards, significant decreases in AST and bilirubin were seen in some subgroups, a rise in GMT in one subgroup; the other changes were non-significant. When assessing the entire group and percentage variations from standard, a mild rise in GMT was again seen; however, the values remain deep within the normal range. Values above the upper reference limit at the start of the study either do not change significantly throughout the study, or they normalize spontaneously. CONCLUSION: No clinically significant changes in liver function tests occurred in users of a triphasic oral contraceptive containing norgestimate along with 35 micrograms EE over a period of six cycles of use. The results, as judged by their dynamics, suggest liver function tests are not a useful tool for routine monitoring a healthy combined oral contraceptive user. PMID- 11272062 TI - [Hyperandrogenic syndrome (polycystic ovary syndrome)--diagnostic criteria, differential diagnosis, clinical signs and laboratory findings (symptomatology), late risks]. PMID- 11272063 TI - [Early fetal karyotyping and its role in prenatal diagnosis]. AB - OBJECTIVE: Shift of indicated invasive examination in prenatal diagnostics towards the earlier phases of pregnancy with preservation of quality of cytogenetic detection. DESIGN: Cytogenetic and molecular-cytogenetic analysis of the chorionic villi after long term culture. SETTING: Department of Medical Genetics and Foetal Medicine, Faculty of Medicine, Palacky University Olomouc, Faculty Hospital in Olomouc. METHODS: Cultivation of fibroblasts developing from chorionic villi after enzymatic or mechanical disintegration and their karyotyping. Using fluorescent in situ hybridisation to identify the most common chromosomal aneuploidies and to determine gonosomes in indicated cases. RESULTS: Testing and optimisation of long term culture method and its routine use. Method was utilised so far in 12 patients and successfulness was 83%. Additional fluorescent in situ hybridisation was performed in 6 cases. CONCLUSION: Using long term culture method of chorionic villi as reliable and routine tool in prenatal diagnostics. PMID- 11272064 TI - [An improvement in the amniotic fluid collection technique]. AB - OBJECTIVE: Elaboration of a maximally safe and comfortable technique of collection of amniotic fluid (AMC). DESIGN: For practice. SETTING: Gyncentrum Ltd., Hloubetinska 3, Praha 9, Dept. of Medical Genetics GENNET, Klimentska 1, Praha 1. METHOD: In 81 patients AMC was performed using a needle with an external diameter of 0.53 mm, by the sampling system Vacutainer of Becton-Dickinson Co. CONCLUSION: The method elaborated by the authors appears to be perspective for use in invasive procedures in the sphere of prenatal diagnosis. PMID- 11272066 TI - [Ambulatory hysteroscopy--a new trend in the diagnosis and treatment of intrauterine pathology]. AB - OBJECTIVE: An analysis of ambulatory hysteroscopic procedures. DESIGN: Retrospective study. SETTING: Dept. of Obstetrics and Gynecology of the 1st Faculty of Medicine, Charles University and the General Faculty Hospital, Apolinarska 18, Prague 2, Czech Republic. METHODS: The sample consists of 225 patients who underwent ambulatory hysteroscopy, during the period between September 1999 to February 2000. The sample makes up 46.3% of the total number of hysteroscopies performed during that period. The procedures were performed under a paracervical block, only rarely in combination with analgesic sedation. RESULTS: Indications for the procedure were most often abnormal uterine bleeding peri- or postmenopausally, 51.2%, abnormal ultrasound findings in 9.8%, and endometrial polyps in 1.8%. In only 23.11% of cases, hysteroscopy was performed in the frame of TCS (transcervical surgery), the remaining procedures were diagnostic. The paracervical block was a great benefit to the comfort of the ambulatory procedures. In 93%, the procedure was evaluated as comfortable, in 4.8% there was discomfort, and in only 2.2% general anesthesia was used due to significant pain. We did not observe any early or late complications. CONCLUSION: Hysteroscopy can be performed as an ambulatory procedure, assuming quality equipment, an experienced surgeon, and properly administered local anesthesia. Our results demonstrate that hysteroscopy is not only a diagnostic procedure, but can be used for certain procedures in the area of TCS (with low or intermediate level of difficulty). PMID- 11272065 TI - [Radical trachelectomy in the treatment of early cervical carcinoma]. AB - In recent years a new less radical methods in the treatment of early cervical carcinoma has been introduced. The radical trachelectomy with laparoscopic pelvic lymphadenectomy represents one of these options. This procedure is special in that it not only treats the cervical cancer in acceptable oncological fashion but at the same time preserves the fertility potential of the patients. Thus, this surgery represents a midway point in between conisation and radical hysterectomy. Based on our initial experience in this study the technique of radical trachelectomy is analysed in detail and at the same time the current literature on the subject is reviewed. We conclude that after careful selection of the patients this more conservative approach leads to the same results comparable to classical radical surgery. PMID- 11272067 TI - [Can hysteroscopy reliably detect malignancy? Analysis of 1200 hysteroscopy findings]. AB - OBJECTIVE: The aim of this study was retrospective analysis of how accurate was per-operational visual evaluation of malign process in an uterus cavity during hysteroscopy. And to evaluate whether increasing experience of hysteroscopiers leads to significant accuracy considering the neoplasm of an uterus cavity. SETTING: Department of Gynaecology and Obstetrics, Havirov. METHOD: In Havirov Hospital, 1,200 hysteroscopies altogether were performed in the period from December 1995 to March 1999. In this group, there were 26 cases of histologically verified endometrial cancer. The authors retrospectively attempted to evaluate how accurately the suspected disorder was already stated during the per operational hysteroscopy. The advantage of comparing the sub-group was taken in the first 690 hysteroscopies, of which the complex analysis was published in Cs. Gynekologie 5/98, and in the sub-group of 510 hysteroscopies performed in the following period, to state whether experience can more precisely define the per operational malignity recognition. The statistical analysis was performed by means of the Fischer exact test of numerical charts. Among other things, the MEDLINE database was used during discussion. RESULTS: The endometrial cancer was encountered 26 times altogether, it means in 2.2% cases of hysteroscopies. Carcinoma in situ occurred three times, the stage IA three times, IB 17 times, IC three times. A hysteroscopier described the negative finding incorrectly 13 times altogether, it means 50% of all cases. The sensitivity and the specificity of hysteroscopy for endometrial cancer prediction was 50% and 99.5% (P < 0.01). The comparison of the first sub-group results (16 cases of endometrial cancer, sensitivity 75%, specificity 99.7%, (P < 0.01) and the second sub-group (10 cases of endometrial cancer, sensitivity 10%, specificity 99.2%, P = 0.09%) indicates that even increasing experience of a hysteroscopier does not more precisely define per-operation malign consideration. CONCLUSION: The authors have come to the conclusion that the pre-operation consideration of intrauteral pathology during hysteroscopy does not allow to assess precisely whether there is a neoprocess of an uterus cavity, or not. Even growing experience does not define with more precision verification of malign disorders especially at early stages of this illness. Hysteroscopy always has to be supplemented with endometrium biopsy. PMID- 11272068 TI - [Termination of 128 pregnancies in the 2nd trimester using prostaglandin 15 methyl F2 alpha]. AB - OBJECTIVE: The critical analysis of the authors' standard protocol of termination of pregnancy during the second trimester. DESIGN: The prospective nonrandomised study. SETTING: Department of Gynecology and Obstetrics, 1th Faculty of Medicine, Prague. METHODS: We used a synthetic prostaglandin analog carboprost, 15-methyl prostaglandin F2 alpha for induction of abortion during second trimester. It was administered in a single dose 500 micrograms. It was given into the amniotic cavity through the transabdominal puncture. At the same time dinoprostonum gel (0.5 mg) was given into vagine. Peridural analgesia has used since beginning of contractions. We investigated indication, mean period of induction, correlation between the period and indication and week of pregnancy so as a type and number of complication. RESULTS: From October 1998 till January 2000 128 pregnancy were terminated by intraamniotic prostaglandins administration. After a single administration 67.2% of women aborted within 24 hours. In 32.8% the intraamniotic administration was repeated twice. The mean induction period, i.e. the interval between the administration and abortion of the foetus was 28 hours. We didn't detect a correlation between the period of induction and the week of gestation or indication. COMPLICATIONS: A major blood loss replaced by transfusion of erythrocyte mass at 4 cases, a rest in uterus at 3 cases, once perforation of uterus during revision of the uterine cavity, once sectio minor for bleeding, once major bleeding conjoined with septic shock, once phlebotrombosis and at three headache (in connection with peridural analgesia). CONCLUSION: The therapeutic effect was achieved in all instances. One case of sectio minor was connected with strong vaginal bleeding. The method fulfils condition for second trimester termination of pregnancy--must be safe, rapid, psychologically feasible and associated with a minimal risk of long-term consequences. The disadvantage of the method is the price of prostaglandins and necessity to repeat administration in 32.8% of patients. PMID- 11272069 TI - [Treatment of infertility using oocytes without the zona pellucida]. AB - OBJECTIVE: The study of zona free oocytes fertilization using intracytoplasmic sperm injection (ICSI) with the aim to increase the pregnancy rate in the assisted reproduction. DESIGN: Pilot study. SETTING: Assisted Reproduction Centre, Gynecology Obstetrics Department of the 1st Medical Faculty and General Faculty Hospital, Charles University, Prague. METHODS: Zona free oocytes were fertilized using ICSI and cultured in standard conditions in vitro for 5 days until the blastocyst stage. Blastocysts were cryopreserved using 8% glycerol. RESULTS: The fertilization process of zona free oocyte was physiological, oocytes expulsed the polar body and in standard time, i.e. 18 hour post insemination, two distinct, morphologically normal pronuclei were apparent in cytoplasm. During early cleavage, from the first to the third day of in vitro culture, the dynamics of cell division was normal, the three-dimensional arrangement of cells was more flattened than in normal, zona intact embryo. The fourth and fifth day of culture, the morphological appearance of embryo corresponded to normal development. CONCLUSION: The fertilization of zona free oocytes using ICSI can give morphologically normal zygote and after 5 days culture in vitro these zygotes can develop into blastocyst stage. This method can increase the pregnancy rate in patients with repeatedly low number of oocytes or with defects of zona pellucida. PMID- 11272071 TI - [Clinical problems and complications in laparoscopic access]. AB - OBJECTIVE: To analyse entry-related laparoscopic technique and complications. DESIGN: Literary review. SETTING: Department Gynaecology and Obstetrics, Endoscopic Training Centre, Hospital Kladno. METHOD: Peri- and postoperative complications of laparoscopic entry were assessed and analysed from database Medline and Current Contents. RESULTS: There is helpful to classify intra abdominal entry complications occurring after laparoscopy into the two types (type I and II). The risk of bowel damage, calculated from analysed studies was 0.4/1,000 and for major vessel injuries the risk was 0.2/1,000. CONCLUSION: The need to perforate the abdominal wall to perform laparoscopic intraabdominal surgery will probably always be associated with risk of damaging structures beneath. With good techniques and appropriate case selection, this risk of such complications should be occure less than 1 per 1,000 laparoscopies. None of the existing techniques or technologies completely eliminate the risk of type II damage. PMID- 11272070 TI - [The suspensory system of the vagina--present views]. AB - OBJECTIVE: To describe vaginal support levels according to the DeLancey's classification, their appearance in transversal magnetic resonance images and clinical categories caused by their defects. DESIGN: Review. SETTING: Department of Obstetrics and Gynecology, 1st Medical Faculty, Charles University, Prague; Department of Radiodiagnostics, Charles University, Medical Faculty in Hradec Kralove. METHODS: A review of literature, examinations of female pelvis with magnetic resonance imaging (MRI) and 3 dimensional computer reconstructions were made. RESULTS: We performed 20 MRI studies and 5 computer spatial reconstructions of female pelvis. Selected results of a review of literature and our MRI examinations were correlated with various types of vaginal descensus. CONCLUSIONS: The questions about the supporting structures of female vagina are still not fully answered. Great advance was achieved by some recent pathological studies and by the introduction of modern magnetic resonance units in this field. The DeLancey's classification goes well in accord with clinical examination, imaging methods and cadaver studies. PMID- 11272072 TI - [Anemia in pregnancy--review. Part 2]. AB - OBJECTIVE: To review the clinical risks of iron deficiency anemia (IDA) in pregnant women: a list of the possible disorders of the mother, fetus and the newborn. A discussion about the clinical value of iron administration in gestation. DESIGN: Review article. SETTING: Department of Obstetrics and Gynecology, 1st Faculty of Medicine and the General Faculty Hospital, Charles University, Prague. Apolinarska 18, Prague 2, 128,00. METHODS: Analysis of the results in literature (texts in medical journals, monographies, textbooks, internet--"Medline") and authors' clinical experience. CONCLUSIONS: Routine iron supplementation in pregnancy is still a controversial issue. The key question is, whether improving the mother's laboratory parameters helps to improve her clinical status and the clinical outcome of pregnancy. There is no doubt that iron supplementation in pregnancy decreases the incidence of anemia and increases the level of iron stores in the 2nd and 3rd trimester of gestation as well as in the puerperium. Even with the presence of many recent studies there still exists a lack of proper evidence, that routine iron administration in pregnancy leads to improvement of the clinical status of the mother and fetus. Up to this time there is not sufficient proof either in favour of or against iron supplementation in pregnancy. In conclusion, there is a need for further research (randomized, controlled, clinical trials focused on the clinical outcomes of pregnancy, with a sufficient amount of pregnant women and with representative statistical evaluation; or careful metaanalysis of the existing studies) to reach definite results about the importance of iron administration and about the treatment of asymptomatic anemia in gestation. PMID- 11272073 TI - [Meconium and its significance]. AB - OBJECTIVE: A review of meconium patophysiology and its contribution to the incidence of perinatal infection. DESIGN: Review article. SETTING: Department of Gynaecology and Obstetrics, Charles University and Faculty Hospital Plzen, Czech Republic. METHOD: The reported incidence of meconium-stained amniotic fluid varies between 7 and 22%. The patophysiology of the presence of meconium in the amniotic fluid is not sufficiently explained. Meconium in fetal bowels is under hormonal and neurol control. The presence of the meconium-stained amniotic fluid was always considered to be a potential risk for the fetal and neonatal well being. The review is further divided in to three chapters. (II. Meconium and meconium aspiration syndrome, III. Meconium and postnatal neurological handicap). RESULTS AND CONCLUSION: The first chapter on deals with meconium risk in the development of perinatal infection: intraamniotic infection/chorioamnionitis, postnatal endometritis, infection of the abdominal wound after Caesarean and neonatal infection. The incidence of clinical chorioamnionitis is 15% with the presence of meconium compared to 3% in controls. The incidence of puerperal endometritis is 10% in comparison to 3% under normal conditions. Two main mechanisms of development (or coincidence) of intraamniotic infection in the presence of meconium exist. 1) Infection may be a cause of meconium passage. 2A) Alteration of Zn/P ratio in the amniotic fluid can promote bacterial growth. 2B) Meconium attached to macrophages or absorbed by phagocytosis can impair cellular immune response. The antibiotic prophylaxis is discussed. PMID- 11272074 TI - [Overview of basic approaches in assessing the moral status of the human embryo]. AB - The introduction of in vitro fertilization in 1978 provoked a broad debate about the ethical problems of technological intervention on the human reproduction. It is obvious, that the issue of whether the human embryo is or is not a person seems to be most important in current debates on the morality of selected abortion, cryopreservation and embryo experimentation. The seeking of the moral status of embryo addresses this fundamental question in a multidisciplinary manner. It is clear, that the philosophical or ontological approach to the embryo has a direct bearing upon a practical issues. This work deals with the moral status of embryo. The author claims, that we can establish his or her moral status by two different way. First it is a substantial approach. Then the embryo is the person since the conception. Second is the historical approach. Then the person is a concept of human being, that is unintelligible apart from social and historical context. But when we are not sure whether embryo is a person or not, it is wise to treat the embryo like a person. PMID- 11272075 TI - [Dr. Josef Hynie, (8 May 1900--23 March 1989)]. PMID- 11272076 TI - [Organization continues at the hospital: perspectives of change in the service of hospital personnel]. PMID- 11272077 TI - [Normative evolution of nursing]. PMID- 11272078 TI - [Autonomy and quality in the history of the Italian National Association of Nurses and the International Council of Nursing]. PMID- 11272079 TI - [Methods and instruments of quality: accreditation]. PMID- 11272080 TI - [Continuous improvement of the quality of nursing care]. PMID- 11272081 TI - [Certification of nursing care in a hospital environment according to the norms of the Italian Hospital Association]. PMID- 11272082 TI - [Accreditation of nursing services in France]. PMID- 11272083 TI - [Health professions and accreditation: current status]. PMID- 11272084 TI - [Professional accreditation: from the viewpoint of the citizens]. PMID- 11272085 TI - [Research and evaluation of Internet resources for the practice and teaching of nursing]. AB - Recent changes in the Italian health care system are causing a complex redefinition of the traditional principles of nursing. Among the new principles that are being proposed, the implementation of a clinical practice based on the evidence generated by the medical research community appears to be prominent. However, objective time constraints in finding and evaluating the available information have often hampered the achievement of this highly desirable goal. In this perspective, exploitation of the intrinsic quickness of the internet-based information retrieval systems has the potential to effectively circumvent the problem. To provide nurses with a proper training in a timely search and evaluation of on-line data, we have designed and developed a guide to those websites providing clinical information. This guide consists of (1) reviews of existing websites, and (2) proposal of a standardized model for selection, evaluation, and description of existing and newly appearing websites. We believe that this guide might increase the capability of nurses to effectively exploit the medical and scientific information resources available on the net. PMID- 11272086 TI - Intensive care and limits of technology. AB - The development of technology in the area of intensive care has allowed a fundamental improvement in patient care and supervision and has increased nursing staff's opportunity for autonomy. However the nurses state that they are poorly equipped to utilize or implement resources which they say would benefit the consumer. PMID- 11272087 TI - [Huber needle in situ inpatients under continuous infusion chemotherapy: results of a study, Phase II]. AB - PURPOSE: Chemotherapy administered as a continuous infusion is a widely used treatment in oncology. Huber needle deserves close attention during chemotherapy, but no data are reported on how long it can be left in situ without change. We therefore evaluated the feasibility of leaving in situ the needle for a prolonged time. METHODS: Patients candidated to continuous infusion chemotherapy were considered eligible for the study. The needle was changed at the end of the 21 day period when the patient started a new cycle of chemotherapy. On that occasion the site of injection was evaluated while replacing the needle. RESULTS: On 129 evaluable patients submitted to continuous infusion chemotherapy, 124 patients did not demonstrate any adverse cutaneous reaction. Five patients (3.8%) presented sores but we were able to continue the treatment leaving in situ the needle. CONCLUSION: Our results demonstrated that the needle can be left in situ for the entire time the patient is at home between cycles of chemotherapy. This procedure avoids patient stress and anxiety due to unjustified substitutions of the needle. PMID- 11272088 TI - [Considerations on patients' rights]. AB - In the present paper the patient's bill of rights are considered according to the new legislation of the Country. The involvement of nursing and private Associations are taken into consideration. PMID- 11272089 TI - [The effectiveness of foot reflexotherapy on chronic pain associated with a herniated disk]. AB - Foot reflexology is both a diagnostic technique and therapy. It is an alternative therapy which is considered useful in pain management. Its effectiveness as a therapy has been studied at the Mestre hospital where a clinical study has been undertaken to determine the effectiveness of reflexology in the reduction of pain. A group of 40 persons suffering almost exclusively from a lumbar-sacral disc hernia received three treatments of reflexology massage for a week. The results found that 25 persons (62.5%) reported a reduction in pain, (rating at 0.75 on a scale of 0-4). These results however did not take into consideration the relationship between the effectiveness of foot reflexology and variables such as the persons physicality (Body Mass Index), or their psychological or social status. PMID- 11272090 TI - [Control of infections transmitted by blood]. PMID- 11272091 TI - Overexpression of epidermal growth factor and epidermal growth factor-receptor mRNAs in dyshormonogenetic goiters. AB - Thyroid malignancy has been induced by long-term endogenous thyrotropin (TSH) stimulation in experimental animals, leading to local and distant metastasis. It has been postulated that constant and prolonged endogenous TSH stimulation in dyshormonogenetic thyroid tissues could result in thyroid neoplasia. The possible role of growth factors and oncogenes in goitrogenesis and favoring neoplasia has also been mentioned. Overexpression of certain growth factors and/or their receptors, and of oncogenes implicated in growth promotion may play a significant role in the relatively frequent finding of thyroid malignancy in congenital goiters. In this study the expression of epidermal growth factor (EGF), epidermal growth factor receptor (EGF-R), transforming growth factor-beta (TGF-beta), c myc, and p53 mRNAs was determined in 14 thyroid tissue samples: 6 from patients with thyroid peroxidase (TPO) gene mutations, 4 with thyroglobulin (Tg) gene defects and 4 normal thyroid tissues. EGF mRNA overexpression was seen in 7 of 10 dyshormonogenetic tissues (3.5 to 12.0 arbitrary optical densitometry units [AODU]) and considered significantly higher (p < 0.01) when compared to normal thyroid tissues (0.25 to 0.32 AODU). Moreover, overexpression of EGF-R mRNA was present in 6 of 10 dyshormonogenetic tissues (2.23 to 13.03 AODU) and considered significantly higher (p < 0.01) when compared to normal thyroid tissues (0.42 to 0.65 AODU). There was no difference in c-myc, p53, and TGF-beta mRNAs expression between dyshormonogenetic and normal tissues. The overexpression of EGF and EGF-R mRNAs found in dyshormonogenetic tissues may suggest that this growth factor may play a role in cellular proliferation and contribute to goiter formation. PMID- 11272092 TI - Butyrate alters the expression and activity of cell cycle components in anaplastic thyroid carcinoma cells. AB - Anaplastic thyroid carcinoma (ATC) is the most malignant and aggressive form of thyroid cancer. Most patients die within months of diagnosis, primarily due to the absence of effective chemotherapeutic strategies. Identifying alternative therapies is necessary to increase long-term survival. Butyrate elicits a number of responses from cancer cells both in vitro and in vivo including growth repression, cell cycle arrest, differentiation, and apoptosis. Even though many types of cancer cells have been studied, little is known of the response of ATC cells to this drug. In this study, we report that butyrate induces differential cell cycle arrest (arrest in G1 and G2/M phases) in an ATC cell line that correlates with changes in the expression, phosphorylation, and activity of key components of the cell cycle machinery. Exposure to butyrate increases the expression of the cyclin-dependent kinase inhibitors, p21/Cip1 and p27/Kip1, decreases the expression of cyclin A and cyclin B, inhibits the phosphorylation of the retinoblastoma protein (pRb), and decreases the activity of cdk1 and cdk2 associated kinases. These results suggest that butyrate may be useful in the clinical treatment of ATC. PMID- 11272093 TI - Activin A and activin receptors in thyroid cancer. AB - Proliferation is controlled by a network of mitogenic and growth inhibitory factors. Transforming growth factor-beta1 (TGF-beta1) and activin A are the most important growth inhibitors of benign follicular epithelial cells of the human thyroid. The effects of these substances on malignant primary thyrocytes are not known. We have examined the growth regulatory effects of activin A and TGF-beta1 in primary cultures derived from four papillary cancers, two follicular thyroid cancers, and three benign thyroid tissues. Malignant cells demonstrated resistance to activin and TGF-beta1 or reversal to a weak but significant mitogenic effect (p < 0.001). We also evaluated the activin receptor transcription pattern. Isoforms alk4-1, 4-2, and 4-3 were found in benign (n = 12) and malignant (n = 22) tissues. Two subtypes of type I and type II activin receptors were demonstrated. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) demonstrated a significant threefold downregulation of alk4-1 receptors in papillary (n = 25) and follicular (n = 18) thyroid cancers as compared to normal thyroids (n = 12) (p < 0.001). To our knowledge these are the first data to demonstrate reversal of activin and TGF-beta1 effects in thyroid malignancy and to demonstrate changes of the type Ib activin receptor expression in thyroid malignancy. PMID- 11272094 TI - Association of the HLA-DRB1*0301 and HLA-DQA1*0501 alleles with Graves' disease in a population representing the gene contribution from several ethnic backgrounds. AB - Graves' disease (GD) is the most frequent cause of hyperthyroidism. Although the etiology is not completely elucidated, there are several lines of evidence suggesting multifactorial mechanisms. Genetic, constitutional, and environmental factors are involved in its pathogenesis. Major histocompatibility complex (MHC) class II alleles have been associated with GD in several populations of distinct ethnic backgrounds and there is increasing evidence supporting an association between GD and HLA-DR3 in Caucasian populations. The MHC class II alleles were evaluated in 75 Brazilian patients presenting with GD and in 166 control individuals from the same geographic area. HLA-DRB, DQB, and DQA alleles were identified using polymerase chain reaction (PCR)-amplified DNA hybridized with sequence-specific probes. The HLA-DRB1*0301 allele was significantly increased in patients (34/75, 45.3%) as compared with controls (37/166, 22.3%, p = 0.009), conferring a relative risk (RR) of 2.8 and an etiologic fraction (EF) of 0.287. The HLA-DQA1*0501 allele was also overrepresented in patients (48/71, 67.6%) in relation to controls (24/71, 33.8%; p = 0.004), conferring an RR of 3.74 and an EF of 0.351. The susceptibility conferred by HLA-DQA1*0501 was independent of the HLA-DRB1*0301 allele. On the other hand, the HLA-DQB1*0602 allele was significantly decreased in patients (6/75, 8.0%) in relation to controls (53/166, 31.9%, p = 0.0008), conferring an RR of 0.18 and a preventive fraction of 0.267. Although the Brazilian population comprises individuals of several ethnic backgrounds, these results corroborate the participation of the HLA-DRB1*0301 and HLA-DQA1*0501 alleles as susceptibility markers for GD, and emphasize the participation of the HLA-DQB1*0602 allele as conferring protection against the development of the disease. PMID- 11272095 TI - Absence of sodium/iodide symporter gene mutations in differentiated human thyroid carcinomas. AB - Decrease or loss of the sodium iodide (Na+/I-) symporter (NIS) activity influences the suitability of using radioiodine to detect and treat metastatic thyroid tissues. In previous studies, the presence of the NIS transcript, albeit at lower expression levels, has been shown in most thyroid differentiated carcinomas. In this study we searched for point mutations or other genetic alterations that may be responsible for an altered function of the NIS protein in tumors that still express NIS transcripts. Tumoral cDNAs derived from seven differentiated thyroid carcinomas (DTC), five papillary and two follicular, were analyzed by direct sequencing after polymerase chain reaction (PCR) amplification of the structural gene of the Na+/I- symporter. Neither mutations nor other genetic abnormalities were detected in any tumor sample examined. The data indicate that mutations or other genetic alterations of the NIS structural gene are not a major cause of the reduced iodide uptake in DTC. PMID- 11272096 TI - Identifying differentially expressed genes associated with metastasis of follicular thyroid cancer by cDNA expression array. AB - Patients with follicular thyroid carcinoma have a higher incidence of metastasis than papillary thyroid carcinoma when thyroid cancer is diagnosed. The cDNA expression array technology is utilized herein to profile differentially expressed genes from metastatic human follicular thyroid carcinoma and reveal new tumor markers as well as target genes for therapeutic intervention. Tissue samples were obtained during surgical resection of the thyroid follicular carcinoma and metastatic tissue in the brain of the same patient. Two identical Atlas human cDNA expression arrays were hybridized with 32P-labeled cDNA probes derived from RNA of either primary thyroid cancer or metastatic tissue. Parallel analysis of the hybridized signals allowed us to identify the alteration of gene expression in the metastasis process. Eighteen genes significantly overexpressed and 40 genes significantly underexpressed were identified in the metastatic thyroid cancer. Genes that displayed an altered expression were associated with the processes of cell cycle regulation, apoptosis, DNA damage response, angiogenesis, cell adhesion and mobility, invasion, and immune response. An expression profile of genes that are associated with metastasis process of follicular thyroid cancer was also discussed. Further investigation is required to understand the precise relationship between the altered expression of these genes and the metastasis process of follicular thyroid cancer. PMID- 11272097 TI - Megalin in thyroid physiology and pathology. AB - Megalin, a member of the low density lipoprotein endocytic receptor family, is expressed on the apical surface of thyroid epithelial cells, directly facing the follicle lumen, where colloid is stored in high concentrations. Studies in vivo and with cultured thyroid cells have provided evidence that megalin expression on thyroid cells is TSH-dependent. Thyroglobulin (Tg), the major protein component of the colloid and the precursor of thyroid hormones, binds to megalin with high affinity and megalin mediates in part its uptake by thyrocytes. Tg internalized by megalin avoids the lysosomal pathway and is delivered by transepithelial transport (transcytosis) to the basolateral membrane of thyrocytes, from which it is released into the bloodstream. This process competes with pathways leading to thyroid hormone release from Tg molecules, which occurs following internalization of Tg molecules from the colloid by other means of uptake (fluid phase endocytosis or endocytosis mediated by low affinity receptors) that result in proteolytic cleavage in the lyosomes. During transcytosis of Tg, a portion of megalin (secretory component) remains complexed with Tg and enters the circulation, where its detection may serve as a tool to identify the origin of serum Tg in patients with thyroid diseases. Tg endocytosis via megalin is facilitated by the interaction of Tg with cell surface heparan sulfate proteoglycans, which occurs via a carboxyl terminal heparin binding site of Tg functionally related with a major megalin binding site. Although autoantibodies against megalin can be found in the serum of approximately 50% of patients with autoimmune thyroiditis, a role of megalin in this and other thyroid diseases remains to be established. PMID- 11272098 TI - Thyroid antibodies and fetal loss: an evolving story. AB - This article reviews the literature on thyroid antibodies and miscarriage. In 1990, in a study designed to determine the incidence and etiology of postpartum thyroiditis, a serendipitous finding emerged revealing an association between thyroid antibodies and spontaneous miscarriage. Subsequently, four other studies, performed on three different continents, have confirmed the correlation. Six studies have evaluated the relationship between thyroid antibodies and recurrent abortion, defined as three or more spontaneous miscarriages. The majority of the studies (67%) reported a statistically significant increase in the incidence of thyroid antibodies in the recurrent abortion group as compared to controls. Four intervention trials have evaluated the impact of immunosuppressive therapy in women with thyroid antibodies. Although all of the trials revealed a decrease in the incidence of recurrent abortion, each study was limited by methodological concerns. A recently developed murine model of pregnancy has also demonstrated increased fetal loss in female mice immunized with thyroglobulin when mated with allogeneic males. The implications of these data generated over the last decade are discussed. PMID- 11272099 TI - High frequency of incidental diagnosis of extrathyroidal neoplastic diseases at the fine-needle aspiration biopsy of laterocervical lymph nodes in patients with thyroid nodules. AB - This study was undertaken to evaluate the frequency of the incidental diagnosis of extrathyroidal lymph node diseases at ultrasound-guided fine-needle aspiration biopsy/cytology (FNAB/C) being done to check the presence of metastatic thyroid cancer in 30 subjects with thyroid nodule (TN) and enlarged cervical lymph nodes (CLN). The patients in whom cytology suggested the presence of malignancy in the TN or in the CLN underwent surgical removal for histologic diagnosis. The spectrum of diseases revealed by this survey included: (1) 10 benign diseases including 1 case of Piringer-Kuchinka lymphadenitis with benign TN; (2) 10 metastatic thyroid cancers (2 anaplastic and 8 papillary cancers); (3) 3 benign TN associated with metastatic invasion of cervical lymph nodes from lung (2 cases) and breast (1 case) cancer; (4) 1 Hodgkin's lymphoma of the cervical lymph nodes with hyperplastic TN; (5) 3 nodal lymphomas with benign thyroid nodule and 2 cases of thyroid lymphoma with nodal invasion; and (6) 1 nodal sarcoidosis with benign TN. The results of this study demonstrate that important neoplastic and hematologic diseases affecting the cervical lymph nodes may frequently be incidentally detected using ultrasonography (US) and FNAB/C in the diagnostic procedure for thyroid nodule. PMID- 11272100 TI - Routine measurement of serum calcitonin is useful for early detection of medullary thyroid carcinoma in patients with nodular thyroid diseases. AB - BACKGROUND: Medullary thyroid carcinoma (MTC) is characterized by a high concentration of serum calcitonin. Routine measurement of serum calcitonin concentration has been advocated for detection of MTC among patients with nodular thyroid diseases. However, a minimal to moderate increase of serum calcitonin concentration has been frequently observed in diseases other than MTC. Fine needle aspiration cytology (FNAC) is not a reliable method for detection of MTC. Therefore, we evaluated the usefulness of routine measurement of serum calcitonin concentration in patients with nodular thyroid diseases, and studied the validity of pentagastrin stimulation test and FNAC in these patients. SUBJECTS AND METHODS: We performed routine measurement of serum calcitonin concentrations in 1,448 patients (male, 285, female, 1,163) with nodular thyroid diseases. The average age was 46 years (range, 14-86 years). Initial examination included thyroid examination, thyroid scan or ultrasonography, measurements of serum free triiodothyronine) (T3), free thyroxine (T4), thyrotropin (TSH) levels, and antithyroid autoantibodies. FNAC was performed in all patients who had palpable or visible thyroid nodule by ultrasonography, and pentagastrin stimulation test was performed in 39 patients who consented. Serum calcitonin concentration was measured with a two-site immunoradiometric assay using commercial kits. We also measured the serum calcitonin concentration in 407 healthy subjects without thyroid or nonthyroid diseases. RESULTS: Serum calcitonin concentration was 10 pg/mL or less in 403 normal subjects (99.0 percentile), and 11-13 pg/mL in the remaining 4 subjects. We found that 56 (3.87%) of 1,448 patients with nodular thyroid diseases had serum calcitonin level above 10 pg/mL. Ten patients (0.69%) with histologically confirmed MTC were detected by the routine measurement of serum calcitonin. The prevalence of MTC was 5.2% in 194 patients with thyroid carcinoma. Five of 10 patients with MTC had basal serum calcitonin level above 100 pg/mL. The remaining 5 patients had minimal or moderate elevation of basal serum calcitonin (range, 12-86 pg/mL). Serum calcitonin concentration increased to more than 100 pg/mL by pentagastrin in all patients with MTC (2.4- to 37.7 fold increase). FNAC suggested MTC in only 2 patients (22.2%), and failed to diagnose MTC in 7 patients. FNAC was not performed in 1 patient with MTC, because he had no visible mass by ultrasonography. CONCLUSION: These results suggested that routine measurement of serum calcitonin is useful in the early detection of MTC among patients with nodular thyroid diseases. Pentagastrin stimulation test may also be a reliable way for evaluating thyroid nodular patients with mild or moderate elevation of serum calcitonin concentrations. However, FNAC was not sensitive in detecting MTC. We recommend routine measurement of serum calcitonin concentration in patients with nodular thyroid diseases. PMID- 11272101 TI - Plasma free fatty acids in neonates with congenital hypothyroidism. AB - Since the introduction of neonatal mass screening for congenital hypothyroidism (CH), numerous cases have been detected. It is of interest that even severely hypothyroid neonates rarely exhibit bradycardia, hypothermia, or inactivity, which have been recognized as typical signs of CH. Regarding neonates and young infants, few reported data are available on the effects of thyroid hormones on energy expenditure. Plasma free fatty acids (FFAs), markers for lipolysis, play essential roles in maintaining physiologic homeostasis. To study fuel utilization in CH neonates, we measured heart rates, plasma FFA, and thyroid hormones before and after levothryoxine (LT4) replacement therapy. Fifty-five screen-detected CH neonates and 29 age-matched normal neonates for controls were enrolled. The CH neonates were divided into two groups according to serum thyroid hormone levels: a mildly hypothyroid group (n = 37), serum thyrotropin (TSH) less than 100 microIU/mL and free thyroxine (FT4) 0.6 ng/dL or more; and a severely hypothyroid group (n = 18), TSH 100 microIU/mL or more and FT4 less than 0.6 ng/dL. Twenty four of the 55 patients had their heart rates measured by electrocardiography. Fasting blood samples were taken from the subjects during physical movements. Serum levels of TSH, FT4, FFA, and other blood chemicals, measured on an autoanalyzer system in our hospital, were compared before and after LT4 substitution therapy. The following results were obtained. The mean plasma FFA values before LT4 replacement were 208.5 +/- 89.4 microEq/L in the mildly hypothyroid group, 228.5 +/- 114.7 microEq/L in the severely hypothyroid group, and 213.9 +/- 97.7 microEq/L in controls. No statistical differences were noted among the three values. Two months after LT4 replacement therapy, at the age of 3 months, plasma FFA concentrations significantly increased in both groups compared with those before the therapy. Control infants also showed a significant increase in plasma FFA concentrations from 1 to 3 months of age. There were no significant differences in plasma FFA concentrations among the three groups at the age of 3 months. No significant correlations were found between plasma FFA and serum thyroid hormones. From these results it is suggested that in neonates and young infants, thyroid hormones do not play major roles in mobilization of fats through the adrenergic regulation of lipolysis for energy supply. This may be one of the reasons for the unexpectedly mild signs and symptoms in the screen-detected hypothyroid neonates. PMID- 11272102 TI - An evaluation of thallium imaging for detection of carcinoma in clinically palpable solitary, nonfunctioning thyroid nodules. AB - OBJECTIVE: An evaluation of thallium imaging for differentiating benign from malignant lesions in clinically palpable solitary, nonfunctioning, thyroid nodules. METHODS: Seventy-eight patients presenting with a clinically palpable solitary nonfunctioning thyroid nodule were imaged with 3 mCi thallium-201 with a pinhole acquisition at 20 minutes and 3 hours after injection. Thallium uptake was assessed as grade 1, less than the rest of the gland; grade 2, same as the rest of the gland; and grade 3, more than the rest of the gland. All patients underwent surgery and the histology was compared with the thallium scan results. RESULTS: Of the 78 patients presenting with solitary thyroid nodule, 13 were malignant and 65 were benign. Twenty-four patients with benign disease showed no uptake of thallium at 3 hours (grade 1). Thirty-two patients with benign disease and 2 patients with malignant lesion had grade 2 uptake at 3 hours. Eleven patients with malignant disease and 9 with benign disease had grade 3 uptake at 3 hours. CONCLUSIONS: All malignant lesions had at least grade 2 and most had grade 3 uptake at 3 hours. All lesions with grade 1 uptake at 3 hours were benign, enabling malignancy to be excluded in one-third of cases. Thallium imaging is a useful adjunct to fine-needle cytology in evaluation of solitary thyroid nodules especially when the latter is inconclusive. PMID- 11272103 TI - Graves' disease after 131I therapy for toxic nodule. PMID- 11272104 TI - Medullary thyroid carcinoma and lack of renal agenesis. PMID- 11272105 TI - Quantitative analysis of disturbed cell maturation in dysplastic lesions of the respiratory tract epithelium. AB - Autoradiographic patterns of [3H]thymidine incorporation, nuclear/cytoplasmic ratios (N/C), and the percentage of dark epithelial cells were analyzed in a group of epithelial lesions induced by 7,12-dimethylbenz[a]anthracene (DMBA) in rat tracheal transplants. It was found that similar lesions of different age exhibit the same labeling indices (LIs), therefore the lesions of different age were subsequently pooled in the following groups and studied by high resolution light microscopic autoradiography: squamous metaplasia without or with only mild atypia, squamous metaplasia with moderate atypia, squamous metaplasia with severe atypia, carcinoma in situ, and microinvasive carcinoma. Normal tracheal and esophageal epithelia were also analyzed. Whereas the normal tracheal basal layer exhibited an LI smaller than 1%, a clear difference between the carcinomas (in situ and invasive) on one hand (LI approximately 32%) and all the remaining epithelia on the other hand (LI approximately 18%) was detected. The LIs of the suprabasal cells exhibited a statistically significant difference between the squamous epithelia without atypia (LI approximately 2%) and the group comprising all the atypical lesions (LI approximately 9%). Gradients of increasing N/C (nucleus-cytoplasm ratios) values could be observed as the lesions increased in severity, especially in the middle and surface layers (e.g., in the surface layer regular metaplasia N/C = 0.08, squamous metaplasia with moderate atypia N/C = 0.26, and carcinoma in situ N/C = 0.50). Dark cells were absent in the normal esophageal epithelium, were present in moderate numbers in the basal layer of regular squamous metaplasia (18%), and increased markedly in the atypical epithelial lesions (approximately 50% in the atypical squamous metaplasias and 70% in carcinoma in situ). In the suprabasal layer dark cells increased from 3% in squamous metaplasia with moderate atypia to 28% in metaplasia with severe atypia and 56% in carcinoma in situ. The results confirm in a quantitative fashion that disturbances of cell maturation and cell proliferation are key features of dysplastic lesions induced by chemical carcinogens, and suggest the use of objective parameters for evaluation and classification of preneoplastic alterations. PMID- 11272106 TI - In vitro formation and properties of beta- and gamma-hydroxy-N nitrosohexamethyleneimine. AB - Stable hydroxylated metabolites of N-nitrosohexamethyleneimine (NO-HEX) account for about one-third of the metabolites produced in in vitro reactions using uninduced rat liver microsomes post-microsomal supernatant. The ratio of gamma- to beta-hydroxyNO-HEX thus produced is 3:1. Each of these isomers exists in two conformeric forms that can be separated easily at room temperature by h.p.l.c. on C18 columns. The ratio of isomers does not vary with substrate concentration or time of reaction, which suggests that one liver enzyme is responsible for the formation of both isomers. Different ratios of conformeric forms of gamma hydroxyNO-HEX produced by liver and lung microsomes indicate that these organs have different enzymes that perform the same functions. Anti-syn (E-Z) configurational assignments for the conformers were based on data obtained from 13C-n.m.r. studies. Although isolated conformers were stable at room temperature, when heated above 70 degrees C or dissolved in aprotic solvents, after several hours an equilibrium mixture was formed. The conformers of beta-hydroxyNO-HEX reached equilibrium in less than half the time required by gamma-hydroxy conformers. The results of i.r. spectroscopic studies suggest that hydrogen bonding may be involved in the stabilization of the conformers. Upon modification of the hydroxy group separable conformers are not detected; this is consistent with the suggestion that the hydroxy group may be involved in the stabilization of separable conformeric forms. We speculate that the occurrence of relatively stable and separable conformers may be due to intramolecular hydrogen bonding between the hydroxy and nitroso groups. PMID- 11272107 TI - Histogenesis of pseudo-ductular changes induced in the pancreas of guinea pigs treated with N-methyl-N-nitrosourea. AB - Tumors of the exocrine pancreas of the inbred strain 13 guinea pigs, induced by N methyl-N-nitrosourea, reveal duct-like glandular differentiation and marked desmoplastic reaction of the stroma, characteristic of adenocarcinoma of human pancreas. During the course of induction of these tumors in the guinea pigs by N methyl-N-nitrosourea, atypical pseudo-ductular proliferations were encountered in the pancreas which appeared to be precursor lesions for pancreatic carcinoma. The histogenesis of these pseudo-ductular lesions was studied by light and electron microscopy. The earliest changes consisted of dilatation of acinar lumina with decrease of apical cytoplasm and increased mitotic activity of the acinar cells. The actively proliferating, well-formed pseudo-ductules were lined by cuboidal or flattened epithelium containing a prominent nucleus and scant cytoplasm with few or no discernible zymogen granules by light microscopy. By electron microscopy, the cells lining the pseudo-ductules displayed features of immature or embryonic pancreatic acinar cells characterized by prominent nucleoli, marked decrease in rough endoplasmic reticulum with increase of free ribosomes, atypical zymogen granules and abundant microfilaments and microtubules. In two guinea pigs, transition from pseudo-ductular changes to adenocarcinoma was clearly evident. On the basis of these findings, it is proposed that the pseudo-ductular lesions of the guinea pig pancreas, and possibly those occuring in other species, are derived from acinar cells as a consequence of carcinogen induced cell proliferation leading to immature or dedifferentiated phenotypes. This hypothesis can, in part, be confirmed by immunocytochemical localization of pancreatic acinar cell specific secretory proteins and lectins in these pseudo-ductules. PMID- 11272108 TI - Investigation of the mechanism for enhancement of radiation transformation in vitro by 12-O-tetradecanoylphorbol-13-acetate. AB - We have performed experiments designed to investigate the mechanism for the enhancement of radiation transformation in vitro by 12-O-tetradecanoylphorbol-13 acetate (TPA). Two types of experiments, involving C3H 10T1/2 cells and 100 rad X ray exposures with subsequent TPA treatment, are reported here. In one set of experiments, cultures were initially seeded at differing initial cell densities prior to irradiation. In the other series of experiments, cultures exposed to 100 rads and TPA contained the same initial cell densities (about 300 viable cells per dish); these cultures were allowed to reach confluence and were then reseeded at various cell densities to allow a second cycle of growth to confluence (which involved different numbers of cell divisions). We have observed that the number of transformed foci which ultimately developed per dish treated with 100 rads and TPA remains approximately constant even though the cell density (initial or reseeded) is varied over several orders of magnitude. The yield of transformants per dish which occurred following this treatment was similar to those previously observed in cultures irradiated with higher doses (400-600 rads) of X-rays alone. Our results suggest that a similar mechanism for the induction of malignant transformation may be involved for high doses of radiation alone or for a low dose of radiation followed by TPA treatment. PMID- 11272109 TI - Ethylene dibromide and disulfiram: studies in vivo and in vitro on the mechanism of the observed synergistic carcinogenic response. AB - Two possible mechanisms for the reported carcinogenic synergism between ethylene dibromide (EDB) and disulfiram have been investigated in vivo and in vitro, the first involving increased production of an EDB-derived glutathione mustard and the second increased production of bromoacetaldehyde. Consistent with both of these suggested mechanisms, repeated administrations of disulfiram to rats inreased liver glutathione-S-transferase activity and decreased liver low Km aldehyde dehydrogenase activity. However, when added to a rat liver S-9 fraction in vitro, disulfiram decreased transferase activity and only depressed the dehydrogenase activity after a period of preincubation. Although the mutagenic potency of EDB to Salmonella typhimurium was slightly enhanced in vitro by the addition of a rat liver S-9 fraction, the further addition of disulfiram to the assay medium produced no additional change. Similarly, the addition of a range of S-9 and S-0.5 liver fractions derived from disulfiram-treated rats also failed to enhance significantly its mutagenic potency over the normal S-9 fraction. The general implications of these findings are discussed. PMID- 11272110 TI - Isolation of mutants temperature-sensitive for expression of the transformed state from chemically transformed C3H/10T1/2 cells. AB - 58 MCA Cl 16 is an oncogenic methylcholanthrene-transformed variant of the non transformed mouse embryo fibroblast cell line, C3H/10T1/2 Cl 8. Using two different protocols, we have isolated six temperature-sensitive mutants from N methyl-N'-nitro-N-nitrosoguanidine treated cultures of 58 MCA Cl 16. C3H/10T1/2 Cl 8, 58 MCA Cl 16 and the six mutant lines were characterized with respect to several properties associated with the transformed state: morphology, saturation density, anchorage independence, cell surface morphology and growth in medium containing 1% fetal calf serum. In general, C3H/10T1/2 cells behaved as non transformed, whether grown at 33 degrees C or 39.5 degrees C. The transformed parental line and all six mutants behaved as transformed cells at 33 degrees C. At 39.5 degrees C, only the parental transformed line retained the transformed phenotype. Three of the mutants revert towards non-transformed behavior at 39.5 degrees C for all of the properties tested. The remaining mutants are temperature sensitive for some, but not all, transformed characteristics. Thus, while the expression of these transformed properties is sometimes coupled, we have been able to dissociate the expression of traits such as saturation density, anchorage independence and transformed morphology from each other. These mutants should prove to be valuable tools in the study of the mechanisms which underly the expression of the chemically-induced transformed state. PMID- 11272111 TI - The enzyme-catalysed conversion of anti-benzo[a]pyrene-7,8-diol 9,10-oxide into a glutathione conjugate. AB - Anti-BP-7,8-diol 9,10-oxide (r-7,t-8-dihydroxy-t-9, 10-oxy-7,8,9,10 tetrahydrobenzo[a]pyrene) was converted in the presence of a rat-liver supernatant fraction and glutathione into a water-soluble metabolite that was identified as a glutathione conjugate. The formation of the glutathione conjugate appears to be catalysed by glutathione S-transferases, present in the rat-liver supernatant, because the amount of conjugate formed was reduced considerably when anti-BP-7,8-diol 9,10-oxide was incubated with glutathione either in the absence of the supernatant fraction or in the presence of heat-denatured supernatant fraction. PMID- 11272112 TI - Fine structure of rat liver during chronic intoxication with two heterocyclic N nitrosamines: N-nitrosopiperidine and the non-carcinogen, 2,2',6,6'-tetramethyl-N nitrosopiperidine. AB - A comparative fine structural investigation was carried out on the long-term effects on rat liver of chronic exposure to the carcinogen, N-nitrosopiperidine (NPIP) and its non-carcinogenic derivative, 2,2',6,6'-tetramethyl-N nitrosopiperidine. Some morphological alterations apparently related to drug metabolism and toxicity were common to both compounds. Others, notably that involving the rough ER, were specific to NPIP and resembled those that are induced by other hepatocarcinogens and are characteristic of hepatic cell tumour. It is suggested that the rough ER lesion is associated with initiation and provides a morphological "marker" for the induction of liver neoplasia. PMID- 11272114 TI - Metabolism of carcinogenic 2-hydroxybenz. AB - The rates of metabolism of the carcinogenic 2-hydroxybenzo[a]pyrene (2-OH-B[a]P) and the non-carcinogenic 3- and 9-hydroxybenzo[a]pyrenes in cultured cell systems have been studied and compared. While 70-80% of the non-carcinogens are converted to water-soluble derivatives by hamster embryo fibroblasts in 24 h, carcinogenic 2-OH-B[a]P is metabolized at a slower rate (45% in 24 h), comparable to that for the parent hydrocarbon, benzo[a]pyrene (B[a]P). Analysis of extracellular organic solvent-soluble metabolites of 2-OH-B[a]P in cultured hamster embryo fibroblasts, using h.p.l.c., indicates the presence of a single major metabolite, which has been identified by mass spectroscopy as a dihydroxy derivative of B[a]P. At least one additional major organic solvent-soluble metabolite is formed in cultures of either mouse epidermal epithelial cells or human foreskin fibroblasts, indicating a different balance of metabolic pathways in these cell systems. The greater persistence of carcinogenic 2-OH-B[a]P in cells and its higher concentration in the cell cytoplasm compared with the non-carcinogenic phenols may be related to its relatively high biological activity. Differences in metabolism of 2-OH-B[a]P in several cultured cell systems indicate the importance of an appropriate choice of activating system in understanding the relationship between metabolism and carcinogenesis. PMID- 11272113 TI - The inhibition of the transplacental blastomogenic effect of nitrosomethylurea by postnatal administration of buformin to rats. AB - N-Nitrosomethylurea (NMU) (20 mg/kg) was i.p. administered to rats on the 21st day of pregnancy. A decrease of glucose utilisation in the oral glucose tolerance test was found in 3 month old female progeny of NMU-treated rats. The serum insulin level did not differ from control, but serum cholesterol level was higher in offspring of NMU-treated rats. The ability of diethylstilboestrol to inhibit compensatory ovarian hypertrophy was decreased in female hemicastrated 3 month old rats whose mothers were treated with NMU. Postnatal administration of the antidiabetic drug buformin decreased the malignant neurogenic tumor incidence 3.5 times (to rats transplacentally treated with NMU). PMID- 11272115 TI - Malignant transformation by the DNA-protein crosslinking agent trans-Pt(II) diamminedichloride. AB - Trans-Pt(II) diamminedichloride (trans-Pt) produced high levels of DNA-protein crosslinks in mouse C3H-10T1/2 cells and mouse 3T3 cells. Trans-Pt induced malignant transformation in these cell types at all doses tested. In 10T1/2 cells trans-Pt induced moderate levels of transformation similar to that induced by equitoxic doses of X-rays. Trans-Pt did not act as a tumor promoter and enhance the transformation frequency seen with X-rays when cells were irradiated and then treated with this agent. PMID- 11272116 TI - Quantitative assay for carcinogen altered differentiation in mouse epidermal cells. AB - Basal epidermal cells can be selectively maintained as a monolayer in culture medium containing a low ionic calcium concentration of 0.01-0.10 mM. Cessation of proliferation, maturation and shedding of squamous sheets can be induced in this population by increasing the calcium concentration above 0.1 mM. Since alterations in the regulation of proliferation and differentiation are associated with epidermal carcinogenesis in vivo, it appeared reasonable that changes in the phenotypic response to calcium might follow exposure to carcinogens in vitro. Support for this hypothesis was provided by the observation that malignant epidermal cells continued to proliferate when switched from low to high calcium medium, and could thus be selected from a mixture of such cells and a large excess of normal cells which did not survive after induced differentiation. Normal primary epidermal cells were plated in low calcium medium, treated on day 3 with a chemical carcinogen, maintained for 3-9 weeks in low calcium (0.02 mM) and then switched to high calcium medium (1.4 mM). After an additional 4 weeks, surviving epithelial colonies were fixed, stained with rhodamine and counted. Treatment of cultures with 7,12-dimethylbenz[a]anthracene or N-methyl-N'-nitro-N nitrosoguanidine yielded 4-10 fold more colonies than solvent controls. Colony number was proportional to carcinogen dose for both agents, and increased with time in low calcium prior to selection by calcium increase. Cells obtained from colonies in treated cultures demonstrated characteristic epidermal morphology and keratinization, and could be subcultured, but did not grow in agar or produce tumors in syngeneic hosts. This model system represents a quantitative assay for carcinogen altered epithelial cell differentiation and may select for an early property of preneoplastic epidermal cells. PMID- 11272117 TI - Differential properties among clones of simian virus 40-transformed human epithelial cells. AB - Monolayer cultures of human prostatic epithelial cells were exposed to SV40 virus at 35th population doubling. Clones were isolated from infected plates after growth had ceased on the control plates. The nuclei of these clones were virtually all positive for viral T-antigen by immunofluorescence. When the properties of three of these lines were compared to those of normal cells, they were found to have altered morphology, ultrastructure, chromosomes, and growth behavior. All transformed lines had reduced serum dependence and were capable of growing in soft agar. However, their reduced serum dependence was not due to reduced growth factor requirements because each subline's response to growth factors was different. PMID- 11272118 TI - Similar defects in DNA repair and replication in the pigmented xerodermoid and the xeroderma pigmentosum variants. AB - The "pigmented xerodermoid" was previously defined on the basis of mild clinical symptoms that suggested it might be similar to but distinct from xeroderma pigmentosum (XP). XP and pigmented xerodermoid cell cultures were irradiated with ultraviolet light and unscheduled DNA synthesis, strand breakage during repair, chain growth during semiconservative DNA replication with or without caffeine, and the recovery of DNA replication were determined. It is concluded that a pigmented xerodermoid cell culture is indistinguishable from the XP variant and the former term is therefore redundant. The defect common to these cell types appears to be the loss of a gene product that permits normal cells to replicate DNA without interruption at damaged sites (u.v.-induced pyrimidine dimers). The consequence of this loss is that replication forks are blocked more frequently and at lower doses in XP variant cells. The correlation between this defect and high levels of actinic carcinogenesis in these patients points to an important role for perturbations in DNA replication in human carcinogenesis. PMID- 11272119 TI - Factors affecting the performance of the styles cell transformation test. AB - The effects of certain factors on the performance of the cell transformation test of Styles were examined by testing the demonstrability of transformation of cells of a subclone of BHK21 C13 in response to treatment with 4-nitroquinoline-1 oxide, N-methyl-N'-nitro-N-nitrosoguanidine, benzo[a]pyrene and 2 acetamidofluorene. An important requirement for success was supplementation of the soft agar medium with a serum which supported microcolony formation by a high proportion of the cells. Increasing the concentration of an unsuitable serum improved the results obtained; this suggests that the serum was inadequate rather than inhibitory. Alteration of the concentration of S-9 fraction used to activate the precarcinogens benzo[a]pyrene and 2-acetamidofluorene had little effect on the induction of transformation by either. A clearer distinction between the transformation frequencies for control and treated cells was usually obtained when 500 microm rather than 200 microm was the minimum diameter set for definition of transformation. It is suggested that, to assess the validity of results obtained with compounds which appear to induce transformation only at highly toxic levels, transformation frequencies for treated cells should be compared with those for control cells seeded at comparable densities. PMID- 11272120 TI - Studies on the mechanism by which a tumor promoter inhibits binding of epidermal growth factor to cellular receptors. AB - This study analyzes the mechanism by which the tumor promoter 12-0-tetradecanoyl phorbol 13-acetate (TPA) inhibits binding of epidermal growth factor (EGF) to its membrane receptors in HeLa cells. Kinetic studies indicate that inhibition of EGF binding occurs within a few minutes after exposure of cells to TPA; delayed addition of TPA causes a reduction in previously bound EGF. With prolonged exposure to TPA, the cells become refractory to TPA inhibition of EGF binding. Evidence was obtained that TPA acts by causing the dissociation of EGF-receptor complexes present on the cell surface and not by increasing the proteolytic degradation or internalization of EGF. Evidence that the TPA inhibition of EGF receptor binding is not a direct consequence of TPA binding to the "active site" of EGF receptors was obtained by the differential effects of pH, temperature or exposure time and that TPA does not inhibit EGF binding when added to isolated plasma membrane fragments. Studies with a variety of inhibitors suggest that the TPA inhibition of EGF-receptor binding does not require macromolecular synthesis, energy metabolism, or cytoskeletal changes. Taken together, our results suggest that TPA inhibition of EGF-receptor binding results from TPA-induced changes in the membrane microenvironment of EGF receptors. PMID- 11272121 TI - Inhibition of epidermal growth factor binding to cultured mouse epidermal cells by tumor promoters. AB - The tumor promotor 12-O-tetradecanoylphorbol-13-acetate (TPA) caused partial inhibition of the binding of [125I]epidermal growth factor ([125I]EGF) to intact mouse epidermal cells (HEL/37). The proportion of [125I]EGF binding which was insensitive to TPA inhibition decreased with increasing EGF concentration. The partial inhibition of [125I]EGF binding was not due to destruction of TPA or to covalent linkage of EGF to receptor sites. The binding of [125I]EGF to HEL/37 cells showed a curvilinear Scatchard plot; this was converted into a linear plot in the presence of TPA. We conclude that TPA acts to inhibit interactions between EGF receptors or between EGF receptors and some other membrane component(s). PMID- 11272122 TI - Mutagenic, transforming and promoting effect of pickled vegetables from Linxian county, China. AB - A recent epidemiological survey in China showed that there is a regional distribution of esophageal cancer and a correlation between mortality for this cancer and environmental factors, especially the consumption of pickled vegetables. A series of experimental studies with pickled vegetable extract were done using different in vitro biological systems. The results showed that pickled vegetable extract induced 6-thioguanine-resistant mutants in V79 cells and increased sister chromatid exchanges in the same cells and in Syrian hamster embryo cells. Pickled vegetables extract induced transformed foci in Syrian hamster embryo cells and in 3-methylcholanthrene initiated C3H/10T1/2 cells. The mutagenic, transforming and promoting activities of pickled vegetable extract seen in vitro conform with in vivo results and provide evidence for the presence of a mutagen and/or a carcinogen in pickled vegetable extract. A possible role of pickled vegetables consumption in the etiology of esophageal cancer is discussed. PMID- 11272123 TI - Ultrastructural analysis of pancreatic carcinogenesis. III. Multifocal cystic lesions induced by N-nitroso-bis(2-hydroxypropyl)amine in the hamster exocrine pancreas. AB - Hamsters were given weekly injections of the pancreatic carcinogen N-nitroso bis(2-hydroxypyropyl)-amine for up to 44 weeks. The cystic foci induced by this treatment, which are putative precursor lesions, were studied by high resolution light microscopy and electron microscopy. They were found to consist mainly of cells that closely resembled those of hamster pancreatic adenomas and adenocarcinomas, confirming their status as precursors of these tumors. However, they additionally contained cells that possessed acinar cell characteristics, raising the possibility that dedifferentiated acinar cells might be involved in the histogenesis of the foci and thus of the pancreatic tumors. PMID- 11272124 TI - Two-stage carcinogenesis in NMRI mice: intravaginal application of 7,12 dimethylbenz[a]anthracene as initiator followed by the phorbol ester 12-O tetradecanoylphorbol-13-acetate as promoter. AB - In this study a new modification of the systemic 2-stage carcinogenesis experiment is described. Tumors were induced in NMRI-mice using a single intravaginal application of the carcinogen 7,12-dimethylbenz[a]anthracene (DMBA) (initiator) followed by intravaginal treatment with 12-O-tetradecanoylphorbol-13 acetate (TPA) (promotor) as a cocarcinogen over a period of 52 weeks. The substances were applied using small tampons. Development of preneoplastic lesions or neoplasias was not found in the group of untreated control animals (n = 50) and in TPA treated animals (n = 50). However, mice which had been treated with TPA alone showed pronounced hyperplasia. When DMBA was applied without subsequent treatment with TPA (n = 100), formation of preneoplastic lesions was frequently observed. Fibroepitheliomas (papillomas), carcinomas and sarcomas, typically located intravaginally and at the external region of the vagina, were observed after the combined treatment with DMBA and TPA (n = 100). In addition, benign as well as malignant tumors were found, mainly in the forestomach, in DMBA treated animals and more frequently after application of DMBA and TPA. Tumors of the lung and leukemias were identified more frequently in comparison to the normal spontaneous tumor rate in NMRI mice, and were probably in part caused by the promoting substance TPA. On the other hand, no significant tumor formation in the mammary gland and in the ovaries was observed after DMBA/TPA application. The tumors observed in these organs were, however, caused by treatment with the carcinogen DMBA only. This new modification of the 2-stage experiment, i.e. the direct intravaginal application of initiating and promoting substances, has special relevance for the elucidation of the mechanisms involved in the development of carcinomas located at the portio vaginalis uteri in humans. PMID- 11272125 TI - The influence of molecular size and partition coefficients on the predictability of tumor initiation in mouse skin from mutagenicity in Salmonella typhimurium. AB - Initiation of tumors in mouse skin is well correlated with mutagenesis in hamster V79 cells (correlation coefficient r = 0.88), but is poorly correlated with bacterial mutagenicity in the Ames assay (r = 0.58). Efforts to understand the difference in the degrees of correlation led to the observation that if the bacterial mutagenicity data are combined with a molecular size or partition factor, the correlation of initiating activity with this combined parameter approaches that found with the hamster cell mutagenesis. The improvement in correlation (r = 0.84) leads to the suggestion that when a broad size range of compounds is considered a most important factor in accounting for the poor correlation initially observed may be a difference between mammalian and bacterial cells in permeability or target access. PMID- 11272126 TI - Methylnitrosourea induction of thymomas in AKR mice requires one or two "hits" only. AB - Induction of thymomas by methylnitrosourea in many strains of mice requires 3 "hits". AKR mice develop thymomas spontaneously late in life, probably because of their large load of viral leukemia oncogenes. It was expected therefore, and so found, that methylnitrosourea induces thymomas in AKR mice with only 1 or 2 "hits". The viral oncogene therefore appears to function as a dominant "hit" gene cooperating with the chemical carcinogen. PMID- 11272127 TI - Cyclopenta-polycyclic aromatic hydrocarbons: potential carcinogens and mutagens. AB - Cyclopenta-polycyclic aromatic hydrocarbons (cyclopenta-PAHs) are a group of compounds that have been detected as environmental pollutants. Perturbation molecular orbital (PMO) calculations on their presumed ultimate carcinogenic metabolites, the cyclopenta-PAH expoxides, predict that they may have a greater biological hazard than the classic PAHs. PMID- 11272128 TI - Impaired restriction endonuclease cleavage of DNA modified with N-methyl-N nitrosourea. AB - This paper examines the ability of several restriction enzymes to cleave lambda DNA treated in vitro with N-methyl-N-nitrosourea. An impaired digestion was observed with all tested enzymes suggesting that the sequence specific restriction endonucleases are either unable to recognize carcinogen-modified sequences or to insert endonucleolytic cuts at such sites. PMID- 11272129 TI - The discovery of type 1 diabetes. AB - The etiological heterogeneity of idiopathic diabetes has been recognized for 25 years, and subdivision into type 1 and type 2 diabetes is fundamental to the way we think about the disease. Review of the literature suggests that the concept of type 1 diabetes as an immunemediated disease emerged rapidly over the period from 1974 to 1976 and showed many of the features of a classic paradigm shift. A few key observations triggered recognition and acceptance of the new paradigm, but the necessary context was provided by scientific developments in areas mainly unrelated to diabetes. The disease paradigm established by 1976 is still widely accepted, and its essential features have been modified only in detail by the revolution in molecular biology that has occurred over the intervening period. Notwithstanding, some of the underlying assumptions remain imprecise, unchallenged, or unconfirmed. Appreciation of the historical origin and subsequent evolution of these fundamental concepts could stimulate critical analysis and help prepare the way for a new paradigm. PMID- 11272130 TI - Ghrelin, an endogenous growth hormone secretagogue, is a novel orexigenic peptide that antagonizes leptin action through the activation of hypothalamic neuropeptide Y/Y1 receptor pathway. AB - Ghrelin, an endogenous ligand for growth hormone secretagogue (GHS) receptor originally isolated from the stomach, occurs in the hypothalamic arcuate nucleus and may play a role in energy homeostasis. Synthetic GHSs have activated the hypothalamic arcuate neurons containing neuropeptide Y (NPY), suggesting the involvement of NPY in some of ghrelin actions. This study was designed to elucidate the role of ghrelin in the regulation of food intake. A single intracerebroventricular (ICV) injection of ghrelin (5-5,000 ng/rat) caused a significant and dose-related increase in cumulative food intake in rats. Ghrelin (500 ng/rat) was also effective in growth hormone-deficient spontaneous dwarf rats. Hypothalamic NPY mRNA expression was increased in rats that received a single ICV injection of ghrelin (500 ng/rat) (approximately 160% of that in vehicle-treated groups, P < 0.05). The ghrelin's orexigenic effect was abolished dose-dependently by ICV co-injection of NPY Y1 receptor antagonist (10-30 microg/rat). The leptin-induced inhibition of food intake was reversed by ICV co injection of ghrelin in a dose-dependent manner (5-500 ng/rat). Leptin reduced hypothalamic NPY mRNA expression by 35% (P < 0.05), which was abolished by ICV co injection of ghrelin (500 ng/rat). This study provides evidence that ghrelin is an orexigenic peptide that antagonizes leptin action through the activation of hypothalamic NPY/Y1 receptor pathway. PMID- 11272131 TI - Overexpression of 1-acyl-glycerol-3-phosphate acyltransferase-alpha enhances lipid storage in cellular models of adipose tissue and skeletal muscle. AB - Plasma nonesterified fatty acids (NEFA) at elevated concentrations antagonize insulin action and thus may play a critical role in the development of insulin resistance in type 2 diabetes. Plasma NEFA and glucose concentrations are regulated, in part, by their uptake into peripheral tissues. Cellular energy uptake can be increased by enhancing either energy transport or metabolism. The effects of overexpression of 1-acylglycerol-3-phosphate acyltransferase (AGAT) alpha, which catalyzes the second step in triglyceride formation from glycerol-3 phosphate, was studied in 3T3-L1 adipocytes and C2C12 myotubes. In myotubes, overexpression of AGAT-alpha did not affect total [14C]glucose uptake in the presence or absence of insulin, whereas insulin-stimulated [14C]glucose conversion to cellular lipids increased significantly (33%, P = 0.004) with a concomitant decrease (-30%, P = 0.005) in glycogen formation. [3H]oleic acid (OA) uptake in AGAT-overexpressing myotubes increased 34% (P = 0.027) upon insulin stimulation. AGAT-alpha overexpression in adipocytes increased basal (130%, P = 0.04) and insulin-stimulated (27%, P = 0.01) [3H]OA uptake, increased insulin stimulated glucose uptake (56%, P = 0.04) and conversion to cellular lipids (85%, P = 0.007), and suppressed basal (-44%, P = 0.01) and isoproterenol-stimulated OA release (-45%, P = 0.03) but not glycerol release. Our data indicate that an increase in metabolic flow to triglyceride synthesis can inhibit NEFA release, increase NEFA uptake, and promote insulin-mediated glucose utilization in 3T3-L1 adipocytes. In myotubes, however, AGAT-alpha overexpression does not increase basal cellular energy uptake, but can enhance NEFA uptake and divert glucose from glycogen synthesis to lipogenesis upon insulin stimulation. PMID- 11272132 TI - Nitric oxide increases glucose uptake through a mechanism that is distinct from the insulin and contraction pathways in rat skeletal muscle. AB - Insulin, contraction, and the nitric oxide (NO) donor, sodium nitroprusside (SNP), all increase glucose transport in skeletal muscle. Some reports suggest that NO is a critical mediator of insulin- and/or contraction-stimulated transport. To determine if the mechanism leading to NO-stimulated glucose uptake is similar to the insulin- or contraction-dependent signaling pathways, isolated soleus and extensor digitorum longus (EDL) muscles from rats were treated with various combinations of SNP (maximum 10 mmol/l), insulin (maximum 50 mU/ml), electrical stimulation to produce contractions (maximum 10 min), wortmannin (100 nmol/l), and/or the NO synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA) (0.1 mmol/l). The combinations of SNP plus insulin and SNP plus contraction both had fully additive effects on 2-deoxyglucose uptake. Wortmannin completely inhibited insulin-stimulated glucose transport and only slightly inhibited SNP stimulated 2-deoxyglucose uptake, whereas L-NMMA did not inhibit contraction stimulated 2-deoxyglucose uptake. SNP significantly increased the activity of the alpha1 catalytic subunit of 5'AMP-activated protein kinase (AMPK), a signaling molecule that has been implicated in mediating glucose transport in fuel-depleted cells. Addition of the NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME) (1 mg/ml) to the drinking water of rats for 2 days failed to affect the increase in muscle 2-deoxyglucose uptake in response to treadmill exercise. These data suggest that NO stimulates glucose uptake through a mechanism that is distinct from both the insulin and contraction signaling pathways. PMID- 11272133 TI - Effects of chronic central nervous system administration of agouti-related protein in pair-fed animals. AB - The melanocortin receptor (MC3-R and MC4-R) antagonist, agouti-related protein (AGRP), is a potent stimulant of food intake. We examined the effect of chronic intracerebroventricular (ICV) AGRP treatment on energy metabolism and pituitary function in ad libitum fed rats and rats administered AGRP and then pair-fed to a saline control group. Chronic ICV AGRP (83-132) administration (1 nmol/day for 7 days) significantly increased food intake and body weight in ad libitum fed animals compared with saline-treated controls (body weight on day 7: 272 +/- 6 [saline] vs. 319 +/- 8 g [AGRP ad libitum fed]; P < 0.001). A significant increase in the epididymal fat pad weight, interscapular brown adipose tissue (BAT) weight, and plasma leptin was also observed in the ad libitum fed group. In the AGRP pair-fed group, a significant increase in the epididymal fat pad weight, BAT weight, and plasma leptin was again observed, suggesting that AGRP caused metabolic changes independent of increased food intake. BAT uncoupling protein 1 (UCP-1) content was significantly decreased compared with saline controls in both the AGRP ad libitum fed (21 +/- 8% of saline control; P < 0.01) and AGRP pair-fed groups (24 +/- 7% of saline control; P < 0.01). Plasma thyroid-stimulating hormone (TSH) was significantly suppressed compared with saline controls in both the AGRP ad libitum fed and AGRP pair-fed groups (3.5 +/- 0.3 [saline] vs. 2.7 +/ 0.4 [AGRP ad libitum fed] vs. 2.1 +/- 0.2 ng/ml [AGRP pair-fed]; P < 0.01). This study demonstrates that independent of its orexigenic effects, chronic AGRP treatment decreased BAT UCP-1, suppressed plasma TSH, and increased fat mass and plasma leptin, suggesting that it may play a role in energy expenditure. PMID- 11272134 TI - PKCdelta activation: a divergence point in the signaling of insulin and IGF-1 induced proliferation of skin keratinocytes. AB - Insulin and insulin-like growth factor-1 (IGF-1) are members of the family of the insulin family of growth factors, which activate similar cellular downstream pathways. In this study, we analyzed the effects of insulin and IGF-1 on the proliferation of murine skin keratinocytes in an attempt to determine whether these hormones trigger the same signaling pathways. Increasing doses of insulin and IGF-1 promote keratinocyte proliferation in an additive manner. We identified downstream pathways specifically involved in insulin signaling that are known to play a role in skin physiology; these include activation of the Na+/K+ pump and protein kinase C (PKC). Insulin, but not IGF-1, stimulated Na+/K+ pump activity. Furthermore, ouabain, a specific Na+/K+ pump inhibitor, abolished the proliferative effect of insulin but not that of IGF-1. Insulin and IGF-1 also differentially regulated PKC activation. Insulin, but not IGF-1, specifically activated and translocated the PKCB isoform to the membrane fraction. There was no effect on PKC isoforms alpha, eta, epsilon, and zeta, which are expressed in skin. PKC8 overexpression increased keratinocyte proliferation and Na+/K+ pump activity to a degree similar to that induced by insulin but had no affect on IGF 1-induced proliferation. Furthermore, a dominant negative form of PKCdelta abolished the effects of insulin on both proliferation and Na+/K+ pump activity but did not abrogate induction of keratinocyte proliferation induced by other growth factors. These data indicate that though insulin or IGF-1 stimulation induce keratinocyte proliferation, only insulin action is specifically mediated via PKC8 and involves activation of the Na+/K+ pump. PMID- 11272135 TI - Regulation of glycogen synthase kinase-3 in human skeletal muscle: effects of food intake and bicycle exercise. AB - Studies of skeletal muscle from rodents performed both in vivo and in vitro suggest a regulatory role of glycogen synthase kinase (GSK) 3 in glycogen synthase (GS) activation in response to insulin. Recently, hyperinsulinemic clamp studies in humans support such a role under nearly physiological conditions. In addition, in rats the activation of GS in skeletal muscle during treadmill running is time-related to the deactivation of GSK3. We investigated whether GSK3 was deactivated in human muscle during low- (approximately 50% VO2max for 1.5 h) and high-intensity (approximately 75% VO2max for 1 h) bicycle exercise as well as food intake. We observed a small but significant increase in GSK3alpha (10-20%) activity in biopsies obtained from vastus lateralis after both low- and high intensity exercise, whereas GSK3beta activity was unaffected. Subsequent food intake increased Aktphosphorylation (approximately 2-fold) and deactivated GSK3alpha (approximately 40%), whereas GSK3beta activity was unchanged. GS activity increased in response to both exercise and food intake. We conclude that GSK3alpha but not GSK3beta may have a role in the regulation of GS activity in response to meal-associated hyperinsulinemia in humans. However, in contrast to findings in muscle from rats, exercise does not deactivate GSK3 in humans, suggesting a GSK3-independent mechanism in the regulation of GS activity in muscle during physical activity. PMID- 11272136 TI - Prolonged islet graft survival in NOD mice by blockade of the CD40-CD154 pathway of T-cell costimulation. AB - Allorejection and recurrence of autoimmunity are the major barriers to transplantation of islets of Langerhans for the cure of type 1 diabetes in humans. CD40-CD154 (CD40 ligand) interaction blockade by the use of anti-CD154 monoclonal antibody (mAb) has shown efficacy in preventing allorejection in several models of organ and cell transplantation. Here we report the beneficial effect of the chronic administration of a hamster anti-murine CD154 mAb, MR1, in prolonging islet graft survival in NOD mice. We explored the transplantation of C57BL/6 islets into spontaneously diabetic NOD mice, a combination in which both allogeneic and autoimmune components are implicated in graft loss. Recipients were treated either with an irrelevant control antibody or with MR1. MR1 administration was effective in prolonging allograft survival, but did not provide permanent protection from diabetes recurrence. The autoimmune component of graft loss was studied in spontaneously diabetic NOD mice that received syngeneic islets from young male NOD mice. In this combination, a less dramatic yet substantial delay in diabetes recurrence was observed in the MR1-treated recipients when compared with the control group. Finally, the allogeneic component was explored by transplanting C57BL/6 islets into chemically induced diabetic male NOD mice. In this setting, long-term graft survival (>100 days) was achieved in MR1-treated mice, whereas control recipients rejected their grafts within 25 days. In conclusion, chronic blockade of CD154 results in permanent protection from allorejection and significantly delays recurrence of diabetes in NOD mice. PMID- 11272137 TI - Metabolic effects of restoring partial beta-cell function after islet allotransplantation in type 1 diabetic patients. AB - Successful intraportal islet transplantation normalizes glucose metabolism in diabetic humans. To date, full function is not routinely achieved after islet transplantation in humans, with most grafts being characterized by only partial function. Moreover, the duration of full function is variable and cannot be sufficiently predicted with available methods. In contrast, most grafts retain partial function for a long time. We hypothesized that partial function can restore normal protein and lipid metabolism in diabetic individuals. We studied 45 diabetic patients after islet transplantation. Labeled glucose and leucine were infused to assess whole-body glucose and protein turnover in 1) 6 type 1 diabetic patients with full function after intraportal islet transplantation (FF group; C-peptide > 0.6 nmol/l; daily insulin dosage 0.03 +/- 0.02 U x kg(-1) body wt x day(-1); fasting plasma glucose < 7.7 mmol/l; HbA1c < or = 6.5%), 2) 17 patients with partial function (PF group; C-peptide > 0.16 nmol/l; insulin dosage < 0.4 U x kg(-1) body wt x day(-1)), 3) 9 patients with no function (NF group; C peptide < 0.16 nmol/l; insulin dosage > 0.4 U x kg(-1) body wt x day(-1)), and 4) 6 patients with chronic uveitis as control subjects (CU group). Hepatic albumin synthesis was assessed in an additional five PF and five healthy volunteers by means of a primed-continuous infusion of [3,3,3-2H3]leucine. The insulin requirement was 97% lower than pretransplant levels for the FF group and 57% lower than pretransplant levels for the PF group. In the basal state, the PF group had a plasma glucose concentration slightly higher than that of the FF (P = 0.249) and CU groups (P = 0.08), but was improved with respect to the NF group (P < 0.01). Plasma leucine (101.1 +/- 5.9 micromol/l) and branched-chain amino acids (337.6 +/- 16.6 micromol/l) were similar in the PF, FF, and CU groups, and significantly lower than in the NF group (P < 0.01). During insulin infusion, the metabolic clearance rate of glucose was defective in the NF group versus in the other groups (P < 0.01). Both the basal and insulin-stimulated proteolytic and proteosynthetic rates were comparable in the PF, FF, and CU groups, but significantly higher in the NF group (P = 0.05). In addition, the PF group had a normal hepatic albumin synthesis. Plasma free fatty acid concentrations in the PF and FF groups were similar to those of the CU group, but the NF group showed a reduced insulin-dependent suppression during the clamp. We concluded that the restoration of approximately 60% of endogenous insulin secretion is capable of normalizing the alterations of protein and lipid metabolism in type 1 diabetic kidney recipients, notwithstanding chronic immunosuppressive therapy. The results of the present study indicate that "success" of islet transplantation may be best defined by a number of metabolic criteria, not just glucose concentration/metabolism alone. PMID- 11272138 TI - Double-stranded RNA-dependent protein kinase is not required for double-stranded RNA-induced nitric oxide synthase expression or nuclear factor-kappaB activation by islets. AB - Environmental factors, such as viral infection, have been implicated in the destruction of beta-cells during the development of autoimmune diabetes. Double stranded RNA (dsRNA), produced during viral replication, is an active component of a viral infection that stimulates antiviral responses in infected cells. Previous studies have shown that treatment of rat islets with dsRNA in combination with gamma-interferon (IFN-gamma) results in a nitric oxide-dependent inhibition of glucose-stimulated insulin secretion. This study examines the role of nuclear factor-kappaB (NF-kappaB) and the dsRNA-dependent protein kinase (PKR) in dsRNA + IFN-gamma-induced nitric oxide synthase (iNOS) expression and nitric oxide production by rat, mouse, and human islets. Treatment of rat and human islets with dsRNA in the form of polyinosinic-polycytidylic acid (poly IC) and IFN-gamma resulted in iNOS expression and nitric oxide production. Inhibitors of NF-kappaB activation-the proteasome inhibitor MG-132 and the antioxidant pyrrolidine-dithiocarbamate (PDTC)-prevented poly IC + IFN-gamma-induced iNOS expression and nitric oxide production. Incubation of rat islets for 3 h or human islets for 2 h with poly IC alone or poly IC + IFN-gamma resulted in NF-kappaB nuclear translocation and degradation of the NF-kappaB inhibitor protein, IkappaB, events that are prevented by MG-132. PKR has been shown to participate in dsRNA-induced NF-kappaB activation in a number of cell types, including mouse embryonic fibroblasts. However, poly IC stimulated NF-kappaB nuclear translocation and IkappaB degradation to similar levels in islets isolated from mice devoid of PKR (PKR-/-) and wild-type mice (PKR+/+). Furthermore, the genetic absence of PKR did not affect dsRNA + IFN-gamma-induced iNOS expression, nitric oxide production, or the inhibitory actions of these agents on glucose-stimulated insulin secretion. These results suggest that 1) NF-KB activation is required for dsRNA + IFN-gamma-induced iNOS expression, 2) PKR is not required for either dsRNA-induced NF-kappaB activation or dsRNA + IFN-y-induced iNOS expression by islets, and 3) PKR is not required for dsRNA + IFN-gamma-induced inhibition of glucose-stimulated insulin secretion by islets. PMID- 11272140 TI - Stimulated endocrine cell proliferation and differentiation in transplanted human pancreatic islets: effects of the ob gene and compensatory growth of the implantation organ. AB - Neogenesis is crucial for the maintenance of beta-cell mass in the human pancreas and possibly for the outcome of clinical islet transplantation. To date, no studies have reported a stimulation of human beta-cell neogenesis in vivo. Therefore, we investigated whether human alpha-, beta-, and duct cell growth can be stimulated when human islets are xenotransplanted to obese hyperglycemic hyperinsulinemic ob/ob mice immunosuppressed with anti-lymphocyte serum. Moreover, we wanted to study whether beta-cell growth and duct-to-beta-cell differentiation were induced in the hepatocyte growth factor (HGF)-dependent compensatory kidney growth model. For that purpose, we evaluated human islets grafted to nude (nu/nu) mice before uninephrectomy of the contralateral kidney for DNA-synthesis and duct cell expression of the beta-cell-specific transcription factor Nkx 6.1 as an estimate of differentiation. Human islet grafts were well preserved after 2 weeks when transplanted to ob/ob mice during anti-lymphocyte immunosuppression. Both human beta-cells (P < 0.01) and duct cells (P < 0.001) were growth stimulated when islets were transplanted to ob/ob mice. We also observed a correlation between increased duct cell proliferation and increased organ donor age (P = 0.02). Moreover, duct (P < 0.05) and beta-cell (P < 0.05) proliferation, as well as duct cell Nkx 6.1 expression (P < 0.05), were enhanced by the compensatory kidney growth after uninephrectomy. We conclude that it is possible to stimulate human beta-cell neogenesis in vivo, provided that the recipient carries certain growth-stimulatory traits. Furthermore, it seems that duct cell proliferation increases with increasing organ donor age. Altogether, these data and previous results from our laboratory suggest that human beta-cell neogenesis becomes more dependent on differentiation and less dependent on proliferation with increasing age. PMID- 11272139 TI - Effects of glucose and amino acids on free ADP in betaHC9 insulin-secreting cells. AB - Stimulation of insulin release by glucose is widely thought to be coupled to a decrease in the activity of ATP-sensitive K+ channels (KATP channels) that is caused by a decreased concentration of free ADP. To date, most other investigators have reported only on total cellular ADP concentrations, even though only a small fraction of all ADP is free and only the free ADP affects KATP channels. We tested the hypothesis that amino acids elicit insulin release via a decrease in the activity of KATP channels owing to a decrease in the level of free ADP. We estimated the concentration of free ADP in betaHC9 hyperplastic insulin-secreting cells based on the cell diameter and on luminometric measurements of ATP, phosphocreatine, and total creatine. The concentration of free ADP fell exponentially as the concentration of glucose increased. A physiological mixture of amino acids greatly stimulated insulin release at 0-30 mmol/l glucose but affected the concentration of free ADP only to a minor degree and significantly so only at < or = 2 mmol/l glucose. In the presence of 2 deoxyglucose and NaN3, amino acids were unable to stimulate insulin release. When KATP channels were held open with diazoxide (and the plasma membrane partially depolarized with high extracellular KCl), amino acids still stimulated insulin release. We conclude that amino acid-induced insulin release depends on two components: a yet-unknown amino acid sensor and KATP channels, which serve to attenuate hormone release when cellular energy stores are low. We propose that glucose-induced insulin release may be regulated similarly by two components: glucokinase and KATP channels. PMID- 11272141 TI - Defective stimulus-secretion coupling in islets of Psammomys obesus, an animal model for type 2 diabetes. AB - Psammomys obesus is a model of type 2 diabetes that displays resistance to insulin and deranged beta-cell response to glucose. We examined the major signaling pathways for insulin release in P. obesus islets. Islets from hyperglycemic animals utilized twice as much glucose as islets from normoglycemic diabetes-prone or diabetes-resistant controls but exhibited similar rates of glucose oxidation. Fractional oxidation of glucose was constant in control islets over a range of concentrations, whereas islets from hyperglycemic P. obesus showed a decline at high glucose. The mitochondrial substrates alpha ketoisocaproate and monomethyl succinate had no effect on insulin secretion in P. obesus islets. Basal insulin release in islets from diabetes-resistant P. obesus was unaffected by glucagon-like peptide 1 (GLP-1) or forskolin, whereas that of islets of the diabetic line was augmented by the drugs. GLP-1 and forskolin potentiated the insulin response to maximal (11.1 mmol/l) glucose in islets from all groups. The phorbol ester phorbol myristic acid (PMA) potentiated basal insulin release in islets from prediabetic animals, but not those from hyperglycemic or diabetes-resistant P. obesus. At the maximal stimulatory glucose concentration, PMA potentiated insulin response in islets from normoglycemic prediabetic and diabetes-resistant P. obesus but had no effect on islets from hyperglycemic P. obesus. Maintenance of islets from hyperglycemic P. obesus for 18 h in low (3.3 mmol/l) glucose in the presence of diazoxide (375 pmol/l) dramatically improved the insulin response to glucose and restored the responsiveness to PMA. Immunohistochemical analysis indicated that hyperglycemia was associated with reduced expression of alpha-protein kinase C (PKC) and diminished translocation of lambda-PKC. In summary, we found that 1) P. obesus islets have low oxidative capacity, probably resulting in limited ability to generate ATP to initiate and drive the insulin secretion; 2) insulin response potentiated by cyclic AMP-dependent protein kinase is intact in P. obesus islets, and increased sensitivity to GLP-1 or forskolin in the diabetic line may be secondary to increased sensitivity to glucose; and 3) islets of hyperglycemic P. obesus display reduced expression of alpha-PKC and diminished translocation of lambda-PKC associated with impaired response to PMA. We conclude that low beta cell oxidative capacity coupled with impaired PKC-dependent signaling may contribute to the animals' poor adaptation to a high-energy diet. PMID- 11272142 TI - Lipotoxicity of the pancreatic beta-cell is associated with glucose-dependent esterification of fatty acids into neutral lipids. AB - Prolonged exposure of isolated islets to supraphysiologic concentrations of palmitate decreases insulin gene expression in the presence of elevated glucose levels. This study was designed to determine whether or not this phenomenon is associated with a glucose-dependent increase in esterification of fatty acids into neutral lipids. Gene expression of sn-glycerol-3-phosphate acyltransferase (GPAT), diacylglycerol acyltransferase (DGAT), and hormone-sensitive lipase (HSL), three key enzymes of lipid metabolism, was detected in isolated rat islets. Their levels of expression were not affected after a 72-h exposure to elevated glucose and palmitate. To determine the effects of glucose on palmitate induced neutral lipid synthesis, isolated rat islets were cultured for 72 h with trace amounts of [14C]palmitate with or without 0.5 mmol/l unlabeled palmitate, at 2.8 or 16.7 mmol/l glucose. Glucose increased incorporation of [14C]palmitate into complex lipids. Addition of exogenous palmitate directed lipid metabolism toward neutral lipid synthesis. As a result, neutral lipid mass was increased upon prolonged incubation with elevated palmitate only in the presence of high glucose. The ability of palmitate to increase neutral lipid synthesis in the presence of high glucose was concentration-dependent in HIT cells and was inversely correlated to insulin mRNA levels. 2-Bromopalmitate, an inhibitor of fatty acid mitochondrial beta-oxidation, reproduced the inhibitory effect of palmitate on insulin mRNA levels. In contrast, palmitate methyl ester, which is not metabolized, and the medium-chain fatty acid octanoate, which is readily oxidized, did not affect insulin gene expression, suggesting that fatty-acid inhibition of insulin gene expression requires activation of the esterification pathway. These results demonstrate that inhibition of insulin gene expression upon prolonged exposure of islets to palmitate is associated with a glucose dependent increase in esterification of fatty acids into neutral lipids. PMID- 11272144 TI - Hyperinsulinism of infancy: the regulated release of insulin by KATP channel independent pathways. AB - Hyperinsulinism of infancy (HI) is a congenital defect in the regulated release of insulin from pancreatic beta-cells. Here we describe stimulus-secretion coupling mechanisms in beta-cells and intact islets of Langerhans isolated from three patients with a novel SUR1 gene defect. 2154+3 A to G SUR1 (GenBank accession number L78207) is the first report of familial HI among nonconsanguineous Caucasians identified in the U.K. Using patch-clamp methodologies, we have shown that this mutation is associated with both a decrease in the number of operational ATP-sensitive K+ channels (KATP channels) in beta-cells and impaired ADP-dependent regulation. There were no apparent defects in the regulation of Ca2+- and voltage-gated K+ channels or delayed rectifier K+ channels. Intact HI beta-cells were spontaneously electrically active and generating Ca2+ action currents that were largely insensitive to diazoxide and somatostatin. As a consequence, when intact HI islets were challenged with glucose and tolbutamide, there was no rise in intracellular free calcium ion concentration ([Ca2+]i) over basal values. Capacitance measurements used to monitor exocytosis in control and HI beta-cells revealed that there were no defects in Ca2+-dependent exocytotic events. Finally, insulin release studies documented that whereas tolbutamide failed to cause insulin secretion as a consequence of impaired [Ca2+]i signaling, glucose readily promoted insulin release. Glucose was also found to augment the actions of protein kinase C- and protein kinase A-dependent agonists in the absence of extracellular Ca2+. These findings document the relationship between SUR1 gene defects and insulin secretion in vivo and in vitro and describe for the first time KATP channel independent pathways of regulated insulin secretion in diseased human beta-cells. PMID- 11272143 TI - Dysregulation of insulin secretion in children with congenital hyperinsulinism due to sulfonylurea receptor mutations. AB - Mutations in the high-affinity sulfonylurea receptor (SUR)-1 cause one of the severe recessively inherited diffuse forms of congenital hyperinsulinism or, when associated with loss of heterozygosity, focal adenomatosis. We hypothesized that SUR1 mutations would render the beta-cell insensitive to sulfonylureas and to glucose. Stimulated insulin responses were compared among eight patients with diffuse hyperinsulinism (two mutations), six carrier parents, and ten normal adults. In the patients with diffuse hyperinsulinism, the acute insulin response to intravenous tolbutamide was absent and did not overlap with the responses seen in either adult group. There was positive, albeit significantly blunted, acute insulin response to intravenous dextrose in the patients with diffuse hyperinsulinism. Graded infusions of glucose, to raise and then lower plasma glucose concentrations over 4 h, caused similar rises in blood glucose but lower peak insulin levels in the hyperinsulinemic patients. Loss of acute insulin response to tolbutamide can identify children with diffuse SUR1 defects. The greater response to glucose than to tolbutamide indicates that ATP-sensitive potassium (KATP) channel-independent pathways are involved in glucose-mediated insulin release in patients with diffuse SUR1 defects. The diminished glucose responsiveness suggests that SUR1 mutations and lack of KATP channel activity may contribute to the late development of diabetes in patients with hyperinsulinism independently of subtotal pancreatectomy. PMID- 11272145 TI - Characterization of a KATP channel-independent pathway involved in potentiation of insulin secretion by efaroxan. AB - Efaroxan, like several other imidazoline reagents, elicits a glucose-dependent increase in insulin secretion from pancreatic beta-cells. This response has been attributed to efaroxan-mediated blockade of KATP channels, with the subsequent gating of voltage-sensitive calcium channels. However, increasing evidence suggests that, at best, this mechanism can account for only part of the secretory response to the imidazoline. In support of this, we now show that efaroxan can induce functional changes in the secretory pathway of pancreatic beta-cells that are independent of KATP channel blockade. In particular, efaroxan was found to promote a sustained sensitization of glucose-induced insulin release that persisted after removal of the drug and to potentiate Ca2+-induced insulin secretion from electropermeabilized islets. To investigate the mechanisms involved, we studied the effects of the efaroxan antagonist KU14R. This agent is known to selectively inhibit insulin secretion induced by efaroxan, without altering the secretory response to glucose or KCl. Surprisingly, however, KU14R markedly impaired the potentiation of insulin secretion mediated by agents that raise cAMP, including the adenylate cyclase activator, forskolin, and the phosphodiesterase inhibitor isobutylmethyl xanthine (IBMX). These effects were not accompanied by any reduction in cAMP levels, suggesting an antagonistic action of KU14R at a more distal point in the pathway of potentiation. In accord with our previous work, islets that were exposed to efaroxan for 24 h became selectively desensitized to this agent, but they still responded normally to glucose. Unexpectedly, however, the ability of either forskolin or IBMX to potentiate glucose-induced insulin secretion was severely impaired in these islets. By contrast, the elevation of cAMP was unaffected by culture of islets with efaroxan. Taken together, the data suggest that, in addition to effects on the KATP channel, imidazolines also interact with a more distal component that is crucial to the potentiation of insulin secretion. This component is not required for Ca2+-dependent secretion per se but is essential to the mechanism by which cAMP potentiates insulin release. Overall, the results indicate that the actions of efaroxan at this distal site may be more important for control of insulin secretion than its effects on the KATP channel. PMID- 11272146 TI - Physiological increase in plasma leptin markedly inhibits insulin secretion in vivo. AB - The demonstration of leptin receptors on the pancreatic beta-cells suggests the possibility of direct actions of leptin on insulin secretion. In vitro studies on islets or perfused pancreas and beta-cell lines produced inconsistent results. We performed an in vivo study to distinctly examine whether leptin has an effect on glucose-stimulated insulin secretion. Young chronically catheterized Sprague Dawley rats (n = 28) were subjected to a 4-h hyperglycemic clamp study (approximately 11 mmol/l). At minute 120 to 240, rats were assigned to receive either saline or leptin (0.1, 0.5, and 5 microg x kg(-1) x min) infusion. Leptin decreased plasma insulin levels abruptly, and an approximately twofold decrease in plasma insulin levels compared with saline control was sustained over the 2 h of the study (14.8 +/- 5.8 vs. 34.8 +/- 2.6 ng/ml with leptin and saline infusion, respectively, P < 0.001). Moreover, a dose-dependent decrease in plasma insulin levels was noted (r = -0.731, P < 0.01). Since milrinone, an inhibitor of cAMP phosphodiesterase (PDE) 3, did not reverse the effect of leptin on glucose induced insulin secretion, its action may be independent of PDE3. These findings suggest that acute physiological increase in plasma leptin levels acutely and significantly inhibits glucose-stimulated insulin secretion in vivo. The site of leptin effects on insulin secretion remains to be determined. PMID- 11272147 TI - Metabolic regulation by leucine of translation initiation through the mTOR signaling pathway by pancreatic beta-cells. AB - Recent findings have demonstrated that the branched-chain amino acid leucine can activate the translational regulators, phosphorylated heat- and acid-stable protein regulated by insulin (PHAS-I) and p70 S6 kinase (p70S6k), in an insulin independent and rapamycin-sensitive manner through mammalian target of rapamycin (mTOR), although the mechanism for this activation is undefined. It has been previously established that leucine-induced insulin secretion by beta-cells involves increased mitochondrial metabolism by oxidative decarboxylation and allosteric activation of glutamate dehydrogenase (GDH). We now show that these same intramitochondrial events that generate signals for leucine-induced insulin exocytosis are required to activate the mTOR mitogenic signaling pathway by beta cells. Thus, a minimal model consisting of leucine and glutamine as substrates for oxidative decarboxylation and an activator of GDH, respectively, confirmed the requirement for these two metabolic components and mimicked closely the synergistic interactions achieved by a complete complement of amino acids to activate p70s6k in a rapamycin-sensitive manner. Studies using various leucine analogs also confirmed the close association of mitochondrial metabolism and the ability of leucine analogs to activate p70s6k. Furthermore, selective inhibitors of mitochondrial function blocked this activation in a reversible manner, which was not associated with a global reduction in ATP levels. These findings indicate that leucine at physiological concentrations stimulates p70s6k phosphorylation via the mTOR pathway, in part, by serving both as a mitochondrial fuel and an allosteric activator of GDH. Leucine-mediated activation of protein translation through mTOR may contribute to enhanced beta-cell function by stimulating growth related protein synthesis and proliferation associated with the maintenance of beta-cell mass. PMID- 11272148 TI - Expression and distribution of lactate/monocarboxylate transporter isoforms in pancreatic islets and the exocrine pancreas. AB - Transport of lactate across the plasma membrane of pancreatic islet beta-cells is slow, as described by Sekine et al. (J Biol Chem 269:4895-4902, 1994), which is a feature that may be important for normal nutrient-induced insulin secretion. Although eight members of the monocarboxylate transporter (MCT) family have now been identified, the expression of these isoforms within the exocrine and endocrine pancreas has not been explored in detail. Using immunocytochemical analysis of pancreatic sections fixed in situ, we demonstrated three phenomena. First, immunoreactivity of the commonly expressed lactate transporter isoform MCT1 is near zero in both alpha- and beta-cells but is abundant in the pancreatic acinar cell plasma membrane. No MCT2 or MCT4 was detected in any pancreatic cell type. Second, Western analysis of purified beta- and non-beta-cell membranes revealed undetectable levels of MCT1 and MCT4. In derived beta-cell lines, MCT1 was absent from MIN6 cells and present in low amounts in INS-1 cell membranes and at high levels in RINm5F cells. MCT4 was weakly expressed in MIN6 beta-cells. Third, CD147, an MCT-associated chaperone protein, which is closely colocalized with MCT1 on acinar cell membranes, was absent from islet cell membranes. CD147 was also largely absent from MIN6 and INS-1 cells but abundant in RINm5F cells. Low expression of MCT1, MCT2, and MCT4 contributes to the enzymatic configuration of beta-cells, which is poised to ensure glucose oxidation and the generation of metabolic signals and may also be important for glucose sensing in islet non-beta cells. MCT overexpression throughout the islet could contribute to deranged hormone secretion in some forms of type 2 diabetes. PMID- 11272149 TI - Alpha- and beta-cell responses to small changes in plasma glucose in the conscious dog. AB - The responses of the pancreatic alpha- and beta-cells to small changes in glucose were examined in overnight-fasted conscious dogs. Each study consisted of an equilibration (-140 to -40 min), a control (-40 to 0 min), and a test period (0 to 180 min), during which BAY R3401 (10 mg/kg), a glycogen phosphorylase inhibitor, was administered orally, either alone to create mild hypoglycemia or with peripheral glucose infusion to maintain euglycemia or create mild hyperglycemia. Drug administration in the hypoglycemic group decreased net hepatic glucose output (NHGO) from 8.9 +/- 1.7 (basal) to 6.0 +/- 1.7 and 5.8 +/- 1.0 pmol x kg(-1) x min(-1) by 30 and 90 min. As a result, the arterial plasma glucose level decreased from 5.8 +/- 0.2 (basal) to 5.2 +/- 0.3 and 4.4 +/- 0.3 mmol/l by 30 and 90 min, respectively (P < 0.01). Arterial plasma insulin levels and the hepatic portal-arterial difference in plasma insulin decreased (P < 0.01) from 78 +/- 18 and 90 +/- 24 to 24 +/- 6 and 12 +/- 12 pmol/l over the first 30 min of the test period and decreased to 18 +/- 6 and 0 pmol/l by 90 min, respectively. The arterial glucagon levels and the hepatic portal-arterial difference in plasma glucagon increased from 43 +/- 5 and 4 +/- 2 to 51 +/- 5 and 10 +/- 5 ng/l by 30 min (P < 0.05) and to 79 +/- 16 and 31 +/- 15 ng/l by 90 min (P < 0.05), respectively. In euglycemic dogs, the arterial plasma glucose level remained at 5.9 +/- 0.1 mmol/l, and the NHGO decreased from 10 +/- 0.6 to -3.3 +/ 0.6 pmol x kg(-1) x min(-1) (180 min). The insulin and glucagon levels and the hepatic portal-arterial differences remained constant. In hyperglycemic dogs, the arterial plasma glucose level increased from 5.9 +/- 0.2 to 6.2 +/- 0.2 mmol/l by 30 min, and the NHGO decreased from 10 +/- 1.7 to 0 pmol x kg(-1) x min(-1) by 30 min. The arterial plasma insulin levels and the hepatic portal-arterial difference in plasma insulin increased from 60 +/- 18 and 78 +/- 24 to 126 +/- 30 and 192 +/- 42 pmol/l by 30 min, after which they averaged 138 +/- 24 and 282 +/- 30 pmol/l, respectively. The arterial plasma glucagon levels and the hepatic portal-arterial difference in plasma glucagon decreased slightly from 41 +/- 7 and 4 +/- 3 to 34 +/- 7 and 3 +/- 2 ng/l during the test period. These data show that the alpha- and beta-cells of the pancreas respond as a coupled unit to very small decreases in the plasma glucose level. PMID- 11272150 TI - Central infusion of histamine reduces fat accumulation and upregulates UCP family in leptin-resistant obese mice. AB - Leptin resistance has recently been confirmed not only in animal obese models but in human obesity. Evidence is rapidly emerging that suggests that activation of histamine signaling in the hypothalamus may have substantial anti-obesity and antidiabetic actions, particularly in leptin-resistant states. To address this issue, effects of central, chronic treatment with histamine on food intake, adiposity, and energy expenditure were examined using leptin-resistant obese and diabetic mice. Infusion of histamine (0.05 pmol x g body wt(-1) x day(-1)) into the lateral cerebroventricle (i.c.v.) for 7 successive days reduced food intake and body weight significantly in both diet-induced obesity (DIO) and db/db mice. Histamine treatment reduced body fat weight, ob gene expression, and serum leptin concentration more in the model mice than in pair-fed controls. The suppressive effect on fat deposition was significant in visceral fat but not in subcutaneous fat. Serum concentrations of glucose and/or insulin were reduced, and tests for glucose and insulin tolerance showed improvement of insulin sensitivity in those mice treated with histamine compared with pair-fed controls. On the other hand, gene expression of uncoupling protein (UCP)-1 in brown adipose tissue and UCP-3 expression in white adipose tissue were upregulated more in mice with i.c.v. histamine infusion than in the pair-fed controls. These upregulating effects of histamine were attenuated by targeted disruption of the H1-receptor in DIO and db/db mice. Sustained i.c.v. treatment with histamine thus makes it possible to partially restore the distorted energy intake and expenditure in leptin-resistant mice. Together, i.c.v. treatment with histamine contributes to improvement of energy balance even in leptin-resistant DIO and db/db mice. PMID- 11272151 TI - Targeted disruption of histamine H1-receptor attenuates regulatory effects of leptin on feeding, adiposity, and UCP family in mice. AB - Histamine neurons are widely distributed in the brain and suppress food intake through the histamine H1 receptor (H1-R) in the hypothalamus. To examine the role of neuronal histamine in leptin signaling pathways, we investigated the effects of H1-R knockout (H1KO) mice on both food intake and mRNA expressions of uncoupling proteins (UCPs) as regulated by leptin, and concomitantly on basal changes in both expression of hypothalamic neuropeptides and diet-induced fat deposition in adipose tissues. H1KO mice showed no change in daily food intake, growth curve, body weight, or adiposity. Reflecting no specificity in these parameters, H1KO mice induced no basal changes in mRNA expression of hypothalamic neuropeptides, ob gene, or peripheral UCPs. Loading H1KO mice with a high-fat diet accelerated fat deposition and ob gene expression compared with the controls. Leptin-induced feeding suppression was partially attenuated in H1KO mice, indicating involvement of histamine neurons in feeding regulation as a downstream signal of leptin. Upregulation of fat UCP mRNA and reduction of body fat induced by central infusion of leptin were attenuated in the H1KO mice. These results show that H1KO mice are a novel leptin-resistant model and that H1-R is a key receptor for downstream signaling of leptin in the brain that contributes to regulation of feeding, fat deposition, and UCP mRNA expression. PMID- 11272152 TI - Effects of short-term improvement of insulin treatment and glycemia on hepatic glycogen metabolism in type 1 diabetes. AB - Insufficiently treated type 1 diabetic patients exhibit inappropriate postprandial hyperglycemia and reduction in liver glycogen stores. To examine the effect of acute improvement of metabolic control on hepatic glycogen metabolism, lean young type 1 diabetic (HbA1c 8.8 +/- 0.3%) and matched nondiabetic subjects (HbA1c 5.4 +/- 0.1%) were studied during the course of a day with three isocaloric mixed meals. Hepatic glycogen concentrations were determined noninvasively using in vivo 13C nuclear magnetic resonance spectroscopy. Rates of net glycogen synthesis and breakdown were calculated from linear regression of the glycogen concentration time curves from 7:30-10:30 P.M. and from 10:30 P.M. to 8:00 A.M., respectively. The mean plasma glucose concentration was approximately 2.4-fold higher in diabetic than in nondiabetic subjects (13.6 +/- 0.4 vs. 5.8 +/- 0.1 mmol/l, P < 0.001). Rates of net glycogen synthesis and net glycogen breakdown were reduced by approximately 74% (0.11 +/- 0.02 vs. 0.43 +/- 0.04 mmol/l liver/min, P < 0.001) and by approximately 47% (0.10 +/- 0.01 vs. 0.19 +/- 0.01 mmol/l liver/min, P < 0.001) in diabetic patients, respectively. During short-term (24-h) intensified insulin treatment, the mean plasma glucose level was not different between diabetic and nondiabetic subjects (6.4 +/- 0.1 mmol/l). Net glycogen synthesis and breakdown increased by approximately 92% (0.23 +/- 0.04 mmol/l liver/min, P = 0.017) and by approximately 40% (0.14 approximately 0.01 mmol/l liver/min, P = 0.011), respectively. In conclusion, poorly controlled type 1 diabetic patients present with marked reduction in both hepatic glycogen synthesis and breakdown. Both defects in glycogen metabolism are improved but not normalized by short-term restoration of insulinemia and glycemia. PMID- 11272153 TI - Hypoglycemic detection does not occur in the hepatic artery or liver: findings consistent with a portal vein glucosensor locus. AB - Our laboratory has previously demonstrated that hypoglycemic detection occurs in the portal vein, not the liver. To ascertain whether hypoglycemic detection may also occur in the hepatic artery, normoglycemia was established across the liver via a localized hepatic artery glucose infusion. Male mongrel dogs (n = 7) were infused with insulin (5.0 mU x kg(-1) x min(-1)) via the jugular vein to induce systemic hypoglycemia. Animals participated in two hyperinsulinemic-hypoglycemic clamp experiments distinguished by the site of glucose infusion. During the liver irrigation protocol, glucose was infused via the hepatic artery (HA protocol) to maintain liver normoglycemia as systemic glucose concentrations were systematically lowered over 260 min (nadir = 2.2 +/- 0.01 mmol/l). During control experiments, glucose was infused peripherally (PER protocol) to control reductions in blood glucose. Arterial glucose concentrations were not significantly different at any time between the two protocols (P = 0.73). Hepatic artery and liver glucose concentrations were significantly elevated in the HA versus PER protocol throughout the duration of the progressive hyperinsulinemic hypoglycemic clamp. During the PER protocol, epinephrine and norepinephrine concentrations increased significantly above basal values (0.53 +/- 0.06 and 0.85 +/- 0.2 nmol/l, respectively) to plateaus of 4.4 +/- 0.86 (P = 0.0001) and 3.6 +/ 0.69 nmol/l (P = 0.001), respectively. There were no significant differences between the two protocols in the epinephrine (P = 0.81) and the norepinephrine (P = 0.68) response to hypoglycemia. The current findings indicate that glucosensors important to hypoglycemic detection do not reside in the hepatic artery. Furthermore, these data confirm our previous findings that glucosensors important to hypoglycemic detection are not present in the liver, but are in fact localized to the portal vein. PMID- 11272154 TI - Protection against oxidative stress-induced insulin resistance in rat L6 muscle cells by mircomolar concentrations of alpha-lipoic acid. AB - In diabetic patients, alpha-lipoic acid (LA) improves skeletal muscle glucose transport, resulting in increased glucose disposal; however, the molecular mechanism of action of LA is presently unknown. We studied the effects of LA on basal and insulin-stimulated glucose transport in cultured rat L6 muscle cells that overexpress GLUT4. When 2-deoxy-D-glucose uptake was measured in these cells, they were more sensitive and responsive to insulin than wild-type L6 cells. LA, at concentrations < or = 1 mmol/l, had only small effects on glucose transport in cells not exposed to oxidative stress. When cells were exposed to glucose oxidase and glucose to generate H2O2 and cause oxidative stress, there was a marked decrease in insulin-stimulated glucose transport. Pretreatment with LA over the concentration range of 10-1,000 pmol/l protected the insulin effect from inhibition by H2O2. Both the R and S isomers of LA were equally effective. In addition, oxidative stress caused a significant decrease (approximately 50%) in reduced glutathione concentration, along with the rapid activation of the stress-sensitive p38 mitogen-activated protein kinase. Pretreatment with LA prevented both of these events, coincident with protecting insulin action. These studies indicate that in muscle, the major site of insulin-stimulated glucose disposal, one important effect of LA on the insulin-signaling cascade is to protect cells from oxidative stress-induced insulin resistance. PMID- 11272155 TI - Peroxisome proliferator-activated receptor (PPAR)-alpha activation lowers muscle lipids and improves insulin sensitivity in high fat-fed rats: comparison with PPAR-gamma activation. AB - Peroxisome proliferator-activated receptor (PPAR)-alpha agonists lower circulating lipids, but the consequences for muscle lipid metabolism and insulin sensitivity are not clear. We investigated whether PPAR-alpha activation improves insulin sensitivity in insulin-resistant rats and compared the effects with PPAR gamma activation. Three-week high fat-fed male Wistar rats were untreated or treated with the specific PPAR-alpha agonist WY14643 or the PPAR-gamma agonist pioglitazone (both 3 mg x kg(-1) x day(-1)) for the last 2 weeks of high-fat feeding. Like pioglitazone, WY14643 lowered basal plasma levels of glucose, triglycerides (-16% vs. untreated), and leptin (-52%), and also muscle triglyceride (-34%) and total long-chain acyl-CoAs (LCACoAs) (-41%) (P < 0.05). In contrast to pioglitazone, WY14643 substantially reduced visceral fat weight and total liver triglyceride content (P < 0.01) without increasing body weight gain. WY14643 and pioglitazone similarly enhanced whole-body insulin sensitivity (clamp glucose infusion rate increased 35 and 37% and glucose disposal 22 and 15%, respectively, vs. untreated). Both agents enhanced insulin-mediated muscle glucose metabolic index (Rg') and reduced muscle triglyceride and LCACoA accumulation (P < 0.05). Although pioglitazone had more potent effects than WY14643 on muscle insulin sensitization, this was associated with its greater effect to reduce muscle LCACoA accumulation. Overall insulin-mediated muscle Rg' was inversely correlated with the content of LCACoAs (r = -0.74, P = 0.001) and with plasma triglyceride levels (r = -0.77, P < 0.001). We conclude that even though WY14643 and pioglitazone, representing PPAR-alpha and PPAR-gamma activation, respectively, may alter muscle lipid supply by different mechanisms, both significantly improve muscle insulin action in the high fat-fed rat model of insulin resistance, and this effect is proportional to the degree to which they reduce muscle lipid accumulation. PMID- 11272156 TI - Free fatty acids induce peripheral insulin resistance without increasing muscle hexosamine pathway product levels in rats. AB - To evaluate the role of the hexosamine biosynthesis pathway (HBP) in fat-induced insulin resistance, we examined whether fat-induced insulin resistance is additive to that induced by increased HBP flux via glucosamine infusion and, if so, whether such additive effects correlate with muscle HBP product levels. Prolonged hyperinsulinemic (approximately 550 pmol/l) euglycemic clamps were conducted in conscious overnight-fasted rats. After the initial 150 min to attain steady-state insulin action, rats received an additional infusion of saline, Intralipid, glucosamine, or Intralipid and glucosamine (n = 8 or 9 for each) for 330 min. At the conclusion of clamps, skeletal muscles (soleus, extensor digitorum longus, and tibialis anterior) were taken for the measurement of HBP product levels. Intralipid and glucosamine infusions decreased insulin-stimulated glucose uptake (Rd) by 38 and 28%, respectively. When the infusions were combined, insulin-stimulated Rd decreased 47%, significantly more than with Intralipid or glucosamine alone (P < 0.05). The glucosamine-induced insulin resistance was associated with four- to fivefold increases in muscle HBP product levels. In contrast, the Intralipid-induced insulin resistance was accompanied by absolutely no increase in HBP product levels in all of the muscles examined. Also, when infused with glucosamine, Intralipid decreased insulin action below that with glucosamine alone without changing HBP product levels. In a separate study, short-term (50 and 180 min) Intralipid infusion also failed to increase muscle HBP product levels. In conclusion, increased availability of plasma free fatty acids induces peripheral insulin resistance without increasing HBP product levels in skeletal muscle. PMID- 11272157 TI - Transgenic complementation of leptin-receptor deficiency. I. Rescue of the obesity/diabetes phenotype of LEPR-null mice expressing a LEPR-B transgene. AB - Mice homozygous for the Leprdb3J (db3J) mutation are null for all known isoforms of the leptin receptor (LEPR). These animals are obese, hyperphagic, cold intolerant, insulin resistant, and infertile. Mice homozygous for the Leprdb (db) mutation (lacking the B isoform only) have the same phenotype as db3J animals. To better understand the function(s) of the LEPR isoforms in vivo, we generated db3J/db3J and db/db mice bearing a transgene (neuron-specific enolase [NSE]-Rb) expressing the B isoform of LEPR, the isoform capable of activating the signal transducer and activator of transcription (STAT) pathway, under the control of the neuron-specific enolase enhancer/promoter. The NSE-Rb transgene was expressed in the brain, with low levels of expression in adrenals, testis, and white adipose tissue. LEPR-B transgene expression in NSE-Rb db3J/db3J mice partially corrected the increased fat mass, hyperphagia, and glucose intolerance while restoring fertility in males and rescuing the cold intolerance in both sexes. The body weights of NSE-Rb transgenic mice that possessed the full complement of short LEPR isoforms (NSE-Rb db/db mice) were similar to those of NSE-Rb db3J/db3J mice, suggesting that the short LEPR isoforms play little role in body weight regulation. Based on quantitative analysis of hypothalamic neuropeptide gene expression in the transgenic animals, we infer full restoration of leptin sensitivity to proopiomelanocortin (POMC) neurons, partial correction of leptin sensitivity in agouti gene-related protein (AGRP)/neuropeptide Y (NPY) neurons, and a lack of effect on leptin sensitivity of melanin concentrating hormone neurons. Thus, hypothalamic POMC and AGRP/NPY neurons are primary candidates as the mediators of the effects of the NSE-Rb transgene on energy homeostasis, ingestive behavior, the neuroendocrine system, and glucose metabolism. PMID- 11272158 TI - Redistribution of sudomotor responses is an early sign of sympathetic dysfunction in type 1 diabetes. AB - Patients with diabetic neuropathy typically have decreased sweating in the feet but excessive sweating in the upper body. Previous studies of sudomotor function in diabetes have included patients with longstanding disease. The present study was designed to test for the early presence of sudomotor dysfunction and to characterize its relation to glycemic control and other aspects of peripheral nerve function. A total of 37 patients (10 males, 27 females) enrolled in a longitudinal study, in which autonomic function was evaluated annually for 3 years. Patients enrolled 2-22 months after the diagnosis of type 1 diabetes. Forty-one age- and sex-matched healthy control subjects were also studied. Sweat production in response to acetylcholine stimulation was dramatically increased in the forearm at the time of the first evaluation (1.67 +/- 0.24 micro/cm2 in the diabetic patients vs. 1.04 +/- 0.14 microl/cm2 in the control subjects, P < 0.05). Likewise, the ratio of sweating in the forearm to sweating below the waist was higher in the diabetic patients (0.553 +/- 0.07 microl/cm2) than in the control subjects (0.385 +/- 0.04 microl/cm2, P < 0.05). Forearm sweat was negatively associated with the renin-toprorenin ratio and vanillylmandelic acid (VMA) excretion (P < 0.025), tests of sympathetic nerve function. The ratio of sweating in the forearm to sweating in the foot was likewise increased in diabetic patients with poor glycemic control. We interpret this redistribution of sudomotor responses to be indicative of sympathetic nerve injury and conclude 1) that the sympathetic nervous system is especially vulnerable to the adverse effects of chronic hyperglycemia and 2) that sympathetic dysfunction can be detected very early in type 1 diabetes. PMID- 11272159 TI - Cyclic stretch and hypertension induce retinal expression of vascular endothelial growth factor and vascular endothelial growth factor receptor-2: potential mechanisms for exacerbation of diabetic retinopathy by hypertension. AB - Systemic hypertension exacerbates diabetic retinopathy and other coexisting ocular disorders through mechanisms that remain largely unknown. Increased vascular permeability and intraocular neovascularization characterize these conditions and are complications primarily mediated by vascular endothelial growth factor (VEGF). Because systemic hypertension increases vascular stretch, we evaluated the expression of VEGF, VEGF-R2 (kinase insert domain-containing receptor [KDR]), and VEGF-R1 (fms-like tyrosine kinase [Flt]) in bovine retinal endothelial cells (BRECs) undergoing clinically relevant cyclic stretch and in spontaneously hypertensive rat (SHR) retina. A single exposure to 20% symmetric static stretch increased KDR mRNA expression 3.9 +/- 1.1-fold after 3 h (P = 0.002), with a gradual return to baseline within 9 h. In contrast, BRECs exposed to cardiac-profile cyclic stretch at 60 cpm continuously accumulated KDR mRNA in a transcriptionally mediated, time-dependent and stretch-magnitude-dependent manner. Exposure to 9% cyclic stretch increased KDR mRNA expression 8.7 +/- 2.9 fold (P = 0.011) after 9 h and KDR protein concentration 1.8 +/- 0.3-fold (P = 0.005) after 12 h. Stretched-induced VEGF responses were similar. Scatchard binding analysis demonstrated a 180 +/- 40% (P = 0.032) increase in high-affinity VEGF receptor number with no change in affinity. Cyclic stretch increased basal thymidine uptake 60 +/- 10% (P < 0.001) and VEGF-stimulated thymidine uptake by 2.6 +/- 0.2-fold (P = 0.005). VEGF-NAb reduced cyclic stretch-induced thymidine uptake by 65%. Stretched-induced KDR expression was not inhibited by AT1 receptor blockade using candesartan. Hypertension increased retinal KDR expression 67 +/- 42% (P < 0.05) in SHR rats compared with normotensive WKY control animals. When hypertension was reduced using captopril or candesartan, retinal KDR expression returned to baseline levels. VEGF reacted similarly, but Flt expression did not change. These data suggest a novel molecular mechanism that would account for the exacerbation of diabetic retinopathy by concomitant hypertension, and may partially explain the principal clinical manifestations of hypertensive retinopathy itself. Furthermore, these data imply that anti-VEGF therapies may prove therapeutically effective for hypertensive retinopathy and/or ameliorating the deleterious effects of coexistent hypertension on VEGF-associated disorders such as diabetic retinopathy. PMID- 11272160 TI - The effect of diabetes on expression of beta1-, beta2-, and beta3-adrenoreceptors in rat hearts. AB - Diabetic hearts exhibit decreased responsiveness to stimulation by beta adrenoreceptor (beta-AR) agonists. This decrease in activity may be due to changes in expression and/or signaling of beta-AR. Recently we showed that right atrial strips from 14-week streptozotocin (STZ)-induced diabetic rat hearts exhibit decreased responsiveness to beta1-AR agonist stimulation, but not to beta2-AR agonist. In the present study, we investigated the effects of long-term diabetes on the expression of cardiac beta1-, beta2-, and beta3-ARs and looked at whether these changes could be restored with insulin treatment. Using reverse transcription-polymerase chain reaction (RT-PCR), PAGE, and Western blot analysis, we found that beta1-AR mRNA and protein levels decreased by 34.9 +/- 5.8 and 44.4 +/- 5.8%, respectively, in 14 week-STZ-treated diabetic rat hearts when compared with age-matched controls. On the other hand, mRNA levels encoding beta2- and beta3-ARs increased by 72.5 +/- 16.6 and 97.3 +/- 26.1%, respectively. Although the latter translated into a proportional increase in beta3-AR protein levels (100.0 +/- 17.0%), beta2-AR protein levels decreased to 82.6 +/- 1.1% of control. Insulin treatment for 2 weeks, after 12 weeks of untreated diabetes, partially restored beta1-AR mRNA and protein levels to 60.1 +/- 8.4 and 83.2 +/- 5.0%, respectively, of control. Although insulin treatment minimally attenuated the rise in mRNA levels encoding beta2- and beta3-ARs, the steady-state levels of these proteins returned to near control values. These data suggest that the decreased responsiveness of diabetic hearts to stimulation of beta-AR agonists may be due to a decrease in beta1-AR and an increase beta3-AR expression. PMID- 11272161 TI - Acute hyperinsulinism modulates plasma apolipoprotein B-48 triglyceride-rich lipoproteins in healthy subjects during the postprandial period. AB - The role of postprandial insulin in the regulation of postprandial lipid metabolism is still poorly understood. The roles of hyperinsulinemia and insulin resistance in the alteration of postprandial lipid metabolism are not clear either. To improve knowledge in this area, we submitted healthy men to acute hyperinsulinemia in two different ways. In the first study, we compared in 10 men the effects of four isolipidic test meals that induce different degrees of hyperinsulinemia on postprandial lipid metabolism. Three different carbohydrate sources were compared according to their glycemic indexes (GIs; 35, 75, and 100 for white kidney bean, spaghetti, and white bread test meals, respectively); the fourth test meal did not contain any carbohydrates. Postprandial plasma insulin levels were proportional to the GIs (maximal plasma insulin concentrations: 113 +/- 16 to 266 +/- 36 pmol/l). We found a strong positive correlation during the 6 h postprandial period between apolipoprotein (apo) B-48 plasma concentration and insulin plasma concentration (r2 = 0.70; P = 0.0001). In a second study, 5 of the 10 subjects again ingested the carbohydrate-free meal, but during a 3-h hyperinsulinemic- (550 +/- 145 pmol/l plasma insulin) euglycemic (5.5 +/- 0.8 mmol/l plasma glucose) clamp. A biphasic response was observed with markedly reduced levels of plasma apoB-48 during insulin infusion, followed by a late accumulation of plasma apoB-48 and triglycerides. Overall, the data obtained showed that portal and peripheral hyperinsulinism delays and exacerbates postprandial accumulation of intestinally derived chylomicrons in plasma and thus is involved in the regulation of apoB-48-triglyceride-rich lipoprotein metabolism, in the absence of insulin-resistance syndrome. PMID- 11272162 TI - Type I diabetes manifested solely by 2-h oral glucose tolerance test criteria. AB - The clinical presentation of type 1 diabetes usually involves symptoms such as polyuria and polydipsia. However, investigators in the Diabetes Prevention Trial of Type 1 Diabetes (DPT-1) have detected a group of subjects with type 1 diabetes who have a different phenotype. These subjects are asymptomatic, have normal (<6.1 mmol/l) (group A) or impaired (6.1- <7.0 mmol/l) (group B) fasting glucose, but have 2-h glucose values >11.1 mmol/l on their oral glucose tolerance tests (OGTT). Of the 585 OGTTs performed on islet cell antibody (ICA)-positive relatives with insulin autoantibodies (IAA) or low first-phase insulin response (FPIR), normal glucose tolerance (NGT) was found in 427 subjects; impaired glucose tolerance (IGT) was found in 87 subjects, and diabetes was found by 2-h OGTT criteria alone in 61 subjects. Despite marked differences in 2-h glucose values (NGT 5.8 +/- 1.1 mmol/l, IGT 8.9 +/- 0.9 mmol/l, and group A 13.5 +/- 2.5 mmol/l), there were no significant differences in fasting glucose values among NGT (4.8 +/- 0.5 mmol/l), IGT (5.03 +/- 0.5 mmol/l), and group A (4.99 +/- 0.7 mmol/l) categories. Mean FPIR was higher in subjects with NGT compared with subjects with IGT and subjects diagnosed by 2-h OGTT criteria alone. However, the correlation between FPIR and 2-h glucose value was low (r2 = 0.114). Multivariate analysis demonstrated that additional independent variables provide smaller contributions to the 2-h glucose value. In conclusion, there are asymptomatic type 1 diabetic subjects whose diabetes was diagnosed by the 2-h criteria on OGTT alone. Despite the importance of beta-cell dysfunction in the pathogenesis of type I diabetes, factors other than impaired FPIR must also contribute to postprandial glucose tolerance in these subjects. PMID- 11272163 TI - The islet in type 2 diabetes: back to center stage. PMID- 11272164 TI - Changes in the expression of transcription factors in pancreatic AR42J cells during differentiation into insulin-producing cells. AB - Pancreatic AR42J cells possess both exocrine and neuroendocrine properties and convert to insulin-producing cells upon treatment with activin A and hepatocyte growth factor (HGF). We studied changes in the mRNA expression of various transcription factors during the course of differentiation. Among the transcription factors studied, expression levels of Pax4 and neurogenin3 changed significantly. These two factors were not detected in naive cells, whereas their mRNA levels were markedly increased after treatment with activin A and HGF. Thus, these two factors were induced by activin A. Transfection of Pax4 did not induce any changes in morphology or expression of pancreatic polypeptide (PP). Furthermore, introduction of antisense Pax4 did not affect the conversion into insulin-producing cells induced by activin A and HGF. In contrast, transfection of neurogenin3 induced morphological changes similar to those induced by activin A. In addition, transfection of neurogenin3 induced the expression of PP. Conversely, introduction of antisense neurogenin3 blocked the differentiation of AR42J cells induced by activin A and HGF. These results indicate that activin A regulates the expression of neurogenin3, which is critical for the differentiation of AR42J into endocrine cells. PMID- 11272165 TI - beta-cell genes and diabetes: quantitative and qualitative differences in the pathophysiology of hepatic nuclear factor-1alpha and glucokinase mutations. AB - Mutations in the beta-cell genes encoding the glycolytic enzyme glucokinase (GCK) and the transcription factor hepatocyte nuclear factor (HNF)-1alpha are the most common causes of maturity-onset diabetes of the young (MODY). Studying patients with mutations in these genes gives insights into the functions of these two critical beta-cell genes in humans. We studied 178 U.K. and French MODY family members, including 45 GCK mutation carriers and 40 HNF-1alpha mutation carriers. Homeostasis model assessment of fasting insulin and glucose showed reduced beta cell function in both GCK (48% controls, P<0.0001) and HNF-1alpha (42% controls, P<0.0001). Insulin sensitivity was similar to that of control subjects in the GCK subjects (93% controls, P = 0.78) but increased in the HNF-1alpha subjects (134.5% controls, P = 0.005). The GCK patients showed a similar phenotype between and within families with mild lifelong fasting hyperglycemia (fasting plasma glucose [FPG] 5.5-9.2 mmol/l, interquartile [IQ] range 6.6-7.4), which declined slightly with age (0.017 mmol/l per year) and rarely required pharmacological treatment (17% oral hypoglycemic agents, 4% insulin). HNF-1alpha patients showed far greater variation in fasting glucose both between and within families (FPG 4.1-18.5 mmol/l, IQ range 5.45-10.4), with a marked deterioration with age (0.06 mmol/l per year), and 59% of patients required treatment with tablets or insulin. Proinsulin-to-insulin ratios are increased in HNF-1alpha subjects (29.5%) but not in GCK (18.5%) subjects. In an oral glucose tolerance test, the 0- to 120-min glucose increment was small in GCK patients (2.4+/-1.8 mmol/l) but large in HNF 1alpha patients (8.5+/-3.0 mmol/l, P< 0.0001). This comparison shows that the clear clinical differences in these two genetic subgroups of diabetes reflect the quantitative and qualitative differences in beta-cell dysfunction. The defect in GCK is a stable defect of glucose sensing, whereas the HNF-1alpha mutation causes a progressive defect that alters beta-cell insulin secretion directly rather than the sensing of glucose. PMID- 11272166 TI - Hyperglycemia contributes to impaired insulin response in GK rat islets. AB - Insulin secretion and glucose metabolism were compared in pancreatic islets from type 2 diabetic GK rats treated with phlorizin or vehicle. Treatment of control and GK rats with phlorizin for 30 days did not affect body weight, islet glucose utilization, or islet glucose oxidation. In phlorizin-treated GK rats, glucose induced insulin release was about twofold higher at 11.0 and 16.7 mmol/l glucose compared with vehicle, treated GK rats, whereas phlorizin had no effect on control Wistar rats. However, also in phlorizin-treated GK rats, the amount of insulin released by the islets was significantly less than that from control rats (5.29+/-0.33 vs. 7.50+/-1.31 pmol x min(-1) islet(-1) at 16.7 mmol/l glucose; P<0.001). Islet glucose-6-phosphatase activity was significantly higher in GK rats than in control rats; phlorizin treatment significantly decreased this activity. These findings demonstrate that hyperglycemia per se constitutes an important factor for impaired insulin release in GK rats. Correction of hyperglycemia normalizes islet glucose-6-phosphatase activity, which may be an underlying factor for the partial improvement of glucose-induced insulin release. PMID- 11272167 TI - beta-cell glucotoxicity in the Psammomys obesus model of type 2 diabetes. AB - Deficient insulin secretion and relative hyperproinsulinemia are characteristic features of type 2 diabetes. The gerbil Psammomys obesus appears to be an ideal natural model of the human disease because it shows increased tendency to develop diet-induced diabetes, which is associated with moderate obesity. The disease is characterized by initial hyperinsulinemia, progressing to hypoinsulinemia associated with depleted pancreatic insulin stores and an increased proportion of insulin precursor molecules in the blood and islets. Although the proinsulin translational efficacy was found to be increased in hyperglycemic animals, insulin mRNA levels were not augmented and exhibited a gradual decrease with disease progression. The development of hyperglycemia was associated with a transient increase in beta-cell proliferative activity, as opposed to a prolonged increase in the rate of beta-cell death, culminating in disruption of islet architecture. The hypothesis that glucotoxicity is responsible in part for these in vivo changes was investigated in vitro in primary islet cultures. Islets from diabetes-prone P. obesus cultured at high glucose concentrations displayed changes in beta-cell function that mimic those observed in diabetic animals. These changes include deficient insulin secretion, depleted insulin content, an increased proportion of insulin precursor molecules, a progressive increase of DNA fragmentation, and a transient proliferative response. Furthermore, insulin mRNA was not increased by short-term exposure of P. obesus islets to elevated glucose in vitro. It is proposed that beta-cell glucotoxicity in P. obesus results from the inability of proinsulin biosynthesis to keep pace with chronic insulin hypersecretion. The resulting depletion of the insulin stores may be related to deficient glucose-regulated insulin gene transcription, possibly due to defective PDX-1 (pancreatic duodenal homeobox factor-1) expression in the adult P. obesus. An additional glucotoxic effect involves the loss of beta-cell mass in hyperglycemic P. obesus as a result of progressive beta-cell death without an adequate increase in the rate of beta-cell proliferation. PMID- 11272168 TI - Lipotoxicity of beta-cells in obesity and in other causes of fatty acid spillover. AB - A recently identified function of leptin is to protect nonadipose tissues from the nonoxidative metabolic products of long-chain fatty acids (FAs) during periods of overnutrition by increasing the beta-oxidative metabolism of surplus FAs and reducing lipogenesis. When this protective system fails, harmful products of nonoxidative metabolism such as ceramide increase in nonadipose tissues, including the pancreatic islets and heart, and cause nitric oxide-mediated lipotoxicity and lipoapoptosis. The triacylglycerol content in nonadipocytes provides a useful index of overall nonoxidative metabolism. In normal animal tissue, triacylglycerol is maintained within a narrow range; even when the caloric intake is excessive, compensatory FA-induced upregulation of oxidation prevents overaccumulation. However, if leptin is deficient or if leptin receptors (Ob-R) are nonfunctional, this autoregulatory system does not operate, and triacylglycerol content rises in nonadipose tissues. This provides a source of excess FAs that enter potentially toxic pathways of nonoxidative metabolism leading to apoptosis of certain tissues. FA overload in skeletal muscle causes insulin resistance; in myocardium, it impairs cardiac function; and in pancreatic islets, it causes beta-cell dysfunction, apoptosis, and diabetes. All abnormalities in these tissues can be blocked by troglitazone, an inhibitor of FA accumulation. PMID- 11272169 TI - Overstimulation and beta-cell function. AB - Previous and present evidence ascribes an important role to overstimulation of beta-cells for the secretory abnormalities associated with type 2 diabetes. The abnormality most clearly linked to overstimulation is the elevated ratio of circulating proinsulin to insulin. Evidence obtained in human pancreatic islets suggests that aberrations in insulin oscillations that occur in type 2 diabetes could at least in part be linked to abnormalities in cytoplasmic Ca2+ oscillations induced by overstimulation. Furthermore, in a transplantation model, we have obtained evidence for long-lasting, perhaps irreversible, effects of overstimulation, implying that this is a causative factor for the well-recognized deterioration of insulin secretion with increasing duration of type 2 diabetes. The mechanisms behind the effects of overstimulation are only partly clarified, but it is clear that reduced insulin secretion after overstimulation is only partly explained by decreased insulin stores. In cultured human pancreatic islets, overstimulation by high glucose leads to a rise in cytoplasmic Ca2+ levels, which persists after normalization of the glucose levels. Persistent elevation of cytoplasmic Ca2+ may trigger apoptosis, thus participating in long term irreversible deterioration of beta-cell function. These data provide sufficient rationale for clinical studies to test the beneficial effects of relative beta-cell rest in type 2 diabetic patients. PMID- 11272170 TI - Sialylated form of the neural cell adhesion molecule (NCAM): a new tool for the identification and sorting of beta-cell subpopulations with different functional activity. AB - To clarify the relationship between variations in beta-cell mass and pancreatic function, we investigated the possibility to analyze, quantify, and sort beta cell subpopulations with different functional maturity. To this aim, we tested the reliability of the sialylated form of neural cell adhesion molecule (NCAM) (PSA-NCAM) as a marker of beta-cell functional activity. Islet cells isolated from adult rats were analyzed for their PSA-NCAM abundance using an anti-PSA-NCAM antibody. We found that PSA-NCAM is expressed only in beta-cells. The PSA-NCAM labeling was also studied with a fluorescence-activated cell sorter. We showed that the beta-cell population is heterogeneous for PSA-NCAM labeling. To directly determine the relationship between PSA-NCAM labeling and beta-cell activity, in vitro insulin secretion studies were performed on sorted beta-cell subpopulations using a perifusion technique. Two beta-cell subpopulations were analyzed: one that was highly labeled for PSA-NCAM and another that was poorly labeled. Insulin secretion from high PSA-NCAM-labeled beta-cells was significantly higher than that in low PSA-NCAM-labeled beta-cells. This differential expression in the beta cell population was well correlated with differences in glucose responsiveness. PSA-NCAM seems thus suitable for use as a tool to identify beta-cell subpopulations according to their glucose responsiveness. PMID- 11272171 TI - beta-cell-specific expression of insulin and PDX-1 genes. PMID- 11272173 TI - Expression profiling of pancreatic beta-cells: glucose regulation of secretory and metabolic pathway genes. PMID- 11272172 TI - Establishment of a tet-on gene expression system in glucose-responsive and unresponsive MIN6 cells. PMID- 11272174 TI - Potential role of the early response gene c-myc in beta-cell adaptation to changes in glucose concentration. PMID- 11272175 TI - Defective glucose-regulated insulin gene expression associated with PDX-1 deficiency in the Psammomys obesus model of type 2 diabetes. PMID- 11272176 TI - IRS proteins and beta-cell function. AB - Insulin receptor substrate (IRS) proteins mediate a variety of the metabolic and growth-promoting actions of insulin and IGF-1. After phosphorylation by activated receptors, these intracellular signaling molecules recruit various downstream effector pathways including phosphatidylinositol 3-kinase and Grb2. Ablation of the IRS-2 gene produces a diabetic phenotype; mice lacking IRS-2 display peripheral insulin resistance and beta-cell dysfunction characterized by a 50% reduction in beta-cell mass. In contrast, deletion of IRS-1 retards somatic growth and enhances beta-cell mass. IRS1-/- mice are 50% smaller than controls but have a twofold increase in pancreatic beta-cell mass. Thus, observations from these recently developed animal models implicate the IRS signaling systems in the response of classical insulin target tissues, and they suggest a critical role for these proteins in the regulation of beta-cell function. In humans, type 2 diabetes generally occurs when insulin-secretory reserves fail to compensate for peripheral insulin resistance. Study and identification of the signals downstream of IRS proteins in beta-cells may provide unique insights into the compensatory mechanisms by which these cells respond to insulin resistance. Therefore, the intent of this review is to summarize recent observations regarding the regulation of beta-cell function by members of the IRS protein family. PMID- 11272177 TI - Endocrine pancreas in insulin receptor-deficient mouse pups. AB - Insulin receptor (IR)-deficient pups rapidly become hyperglycemic and hyperinsulinemic and die of diabetic ketoacidosis within a few days. Immunocytochemical analysis of the endocrine pancreas revealed that IR deficiency did not alter islet morphology or the number of beta-, alpha-, delta-, and pancreatic polypeptide (PP) cells. The lack of IR did not result in major changes in the expression of islet hormone genes or of beta-cell-specific marker genes encoding pancreas duodenum homeobox-containing transcription factor-1 (PDX-1), glucokinase (GCK), and GLUT2, as shown by reverse transcriptase-polymerase chain reaction analysis. The serum glucagon levels in IR-deficient and nondiabetic littermates were comparable. Finally, total insulin content in the pancreas of IR deficient pups was gradually depleted, indicating sustained insulin secretion, not compensated for by increased insulin biosynthesis. These findings are discussed in light of recent results suggesting a role of IR in beta-cell function. PMID- 11272178 TI - Online monitoring of stimulus-induced gene expression in pancreatic beta-cells. AB - Fluorescent proteins have been extensively used as protein "tags" to study the subcellular localization of proteins and/or their translocation upon stimulation or as markers for transfection in transient and stable expression systems. However, they have not been frequently used as reporter genes to monitor stimulus induced gene expression in mammalian cells. Here we demonstrate the use of fluorescent proteins to study stimulus-induced gene transcription. The general applicability of the approach is exemplified by doxycyclin-(Tet-On) and phorbol 12-myristate 13-acetate-induced (c-fos) promoter activation, with green fluorescent protein (GFP) and red fluorescent protein (DsRed) as semiquantitative and immediate reporters, of transcription activation. Under the control of beta cell-specific promoters, such as the rat insulin 1 promoter or the rat upstream glucokinase promoter, this approach allowed us to monitor online glucose-induced gene transcription in primary beta-cells at the single-cell level as well as in the context of the islet of Langerhans. Applying discretely detectable fluorescent proteins, for example GFP and DsRed, enabled us to simultaneously monitor stimulus-induced transcription by two different promoters in the same cell. PMID- 11272179 TI - Compensatory responses in mice carrying a null mutation for Ins1 or Ins2. AB - Intrauterine growth retardation and postnatal acute diabetes result from insulin deficiency in double homozygous null mutants for Ins1 and Ins2 (Duvillie B, et al., Proc. Natl. Acad. Sci. USA 94:5137-5140, 1997). The characterization of single homozygous null mutants for Ins1 or Ins2 is described here. Neither kind of mutant mice was diabetic. Immunocytochemical analysis of the islets showed normal distribution of the endocrine cells producing insulin, glucagon, somatostatin, or pancreatic polypeptide. Analysis of the expression of the functional insulin gene in Ins1-/- or Ins2-/- mice revealed a dramatic increase of Ins1 transcripts in Ins2-/- mutants. This compensatory response was quantitatively reflected by total pancreatic insulin content similar for both types of mutants and wild-type mice. Moreover, both mutants had normal plasma insulin levels and normal glucose tolerance tests. The determination of beta-cell mass by morphometry indicated beta-cell hyperplasia in the mutant mice. The beta cell mass in Ins2-/- mice was increased almost threefold, which accounts for the increase of Ins1 transcripts in Ins2-/-mutants. This study thus contributes to evaluate the potential of increasing the beta-cell mass to compensate for low insulin production. PMID- 11272180 TI - Beta-cell adaptation and decompensation during the progression of diabetes. AB - Inadequate beta-cell function is an essential component of all forms of diabetes. The most obvious problem is a failure to maintain sufficient beta-cell mass and function to cope with whatever insulin resistance is present. The most striking functional defect is a loss of acute glucose-induced insulin secretion (GIIS). This review discusses the ways in which beta-cells successfully adapt to increased demand and then decompensate as diabetes develops. Successful adaptation is achieved through increased beta-cell mass and increased insulin secretion. The hypothesis is explored that beta-cells exposed to the diabetic milieu lose their differentiation, which leads to loss of specialized functions such as GIIS. This concept has been strengthened by the finding of dedifferentiation of beta-cells in a rat model of partial pancreatectomy that includes a reduction of insulin gene expression, which may further contribute to decreased insulin production. Another finding was increased expression of c-Myc, which probably contributes to an increase in the expression of lactate dehydrogenase and the development of beta-cell hypertrophy. Arguments are developed that the beta-cell changes found in diabetes are better correlated with increased glucose levels than with non-esterified fatty acid levels, thus supporting the importance of glucose toxicity. PMID- 11272181 TI - beta-cell dysfunction and failure in type 2 diabetes: potential mechanisms. AB - Type 2 diabetes is characterized by a progressive loss of beta-cell function throughout the course of the disease. The pattern of loss is an initial defect in early or first-phase insulin secretion, followed by a decreasing maximal capacity of glucose to potentiate all nonglucose signals. Last, a defective steady-state and basal insulin secretion develops, leading to complete beta-cell failure requiring insulin treatment. This functional loss exceeds the expected impact of a 20-50% loss of beta-cells reported at autopsy, which has been associated with amyloid deposits. This review summarizes the nature of the amyloid deposition process and its association with disproportionate hyperproinsulinemia. It reviews recent studies in IAPP (islet-amyloid polypeptide, or amylin) transgenic mice developing islet amyloid deposits and hyperglycemia to suggest that the process of amyloid fibril formation impairs function early and leads to beta-cell failure and eventual death. Based on the known association of amyloid deposits and relative hyperproinsulinemia, it is hypothesized that fibril formation begins during impaired glucose tolerance after other factors cause the initial defects in early insulin secretion and insulin action. Thus, the process that leads to beta-cell loss is implicated in the deposition of amyloid and the late unrelenting progressive hyperglycemia now found in all patients despite current therapies. PMID- 11272183 TI - Decreased insulin secretion in type 2 diabetes: a problem of cellular mass or function? AB - Type 2 diabetes is characterized by diminished or inappropriate secretion of insulin, which could be a defect of either islet cell function or beta-cell mass. Quantitation of islet cell populations in postmortem pancreas demonstrates little change of beta-cell mass in type 2 diabetes. Reduction of islet cell mass (up to 30%) is associated largely with islet amyloid deposition, and the degree of amyloidosis is independent of the duration of the disease. Insulin secretory capacity is dependent on both function and mass of cells. beta-Cell secretion is heterogeneous; increasing glucose concentrations result in recruitment of beta cells into the secretory pool, indicating a large reserve of secretory capacity that can be recruited in insulin resistant conditions. The Starling curve of islet function describes the relationship of insulin secretion to increasing levels of insulin resistance and hyperglycemia in type 2 diabetes. Longitudinal studies in Macaca mulatta monkeys show that insulin resistance is accompanied by increased islet mass and onset of diabetes is associated with deposition of amyloid and reduction of beta-cells. Increasing the function of unresponsive beta cells rather than the mass of cells may be a more effective therapeutic target for type 2 diabetes. PMID- 11272182 TI - A model for glucose control of insulin secretion during 24 h of free living. AB - The aim of this work was to develop a mathematical model describing the functional dependence of insulin secretion on plasma glucose concentrations during 24 h of free living. We obtained hourly central venous blood samples from a group of healthy volunteers who spent 24 h in a calorimetric chamber, where they consumed standardized meals. Insulin secretory rates were reconstructed from plasma C-peptide concentrations by deconvolution. The relationship between insulin release and plasma glucose concentrations was modeled as the sum of three components: a static component (describing the dependence on plasma glucose concentration itself, with an embedded circadian oscillation), a dynamic component (modeling the dependence on glucose rate of change), and a residual component (including the fraction of insulin secretion not explained by glucose levels). The model fit of the individual 24-h secretion profiles was satisfactory (within the assigned experimental error of glucose and C-peptide concentrations). The static component yielded a dose-response function in which insulin release increased quasi-linearly (from 40 to 400 pmol/min on average) over the range of 4 9 mmol/l glucose. The dynamic component was significantly different from zero in coincidence with meal-related glucose excursions. The circadian oscillation and the residual component accounted for the day/night difference in the ability of glucose to stimulate insulin release. Over 24 h, total insulin release averaged 257+/-58 nmol (or 43+/-10 U). The static and dynamic component together accounted for approximately 80% of total insulin release. The model proposed here provides a detailed robust description of glucose-related insulin release during free living conditions. In nondiabetic subjects, non-glucose-dependent insulin release is a small fraction of total insulin secretion. PMID- 11272184 TI - Human type 2 diabetes: morphological evidence for abnormal beta-cell function. AB - The exact nature of the beta-cell defect in type 2 diabetic patients is still unclear. beta-Cell mass reduction has been reported but remains controversial. A preliminary study of a large series of patients has demonstrated that in most, the beta-cell defect is not related to a decreased beta-cell mass. Amyloid deposits are observed in the islets of some type 2 diabetic patients but also in normoglycemic subjects. Because it has been claimed that these deposits interfere with beta-cell function, we evaluated in situ the effect of insular amyloid deposits on beta-cell transcription and translation. Pancreases were obtained at autopsy from 28 normoglycemic patients and 41 type 2 diabetic patients. Staining with hemaluneosin and Congo red was used to analyze the general features of the islets and the presence of amyloid deposits, respectively. Immunohistochemistry for proinsulin was performed with an antibody recognizing the junction between B chain and C-peptide, thus specifically labeling the Golgi area where proinsulin is produced. In seven patients, we evaluated insulin gene transcription by in situ hybridization of proinsulin mRNA combined with Congo red staining, and we evaluated insulin storage by double immunostaining for insulin and amylin. In many type 2 diabetic patients, the islets appeared entirely normal. Amyloid deposits were found in 57% of diabetic subjects and 33% of normoglycemic age matched control subjects. The percentage of amyloid-infiltrated islets varied from 0.4 to 74%. beta-Cells from amyloid-containing islets still had specific Golgi proinsulin labeling. In obese type 2 diabetic patients, the number of beta cells with abnormal expression of proinsulin in the whole cytoplasm was significantly higher than in normoglycemic control subjects. Proinsulin mRNA was significantly reduced in islets with amyloid deposits when compared with amyloid free islets, but the mean reduction did not exceed 16%. Insulin was still present in the beta-cells of amyloid-containing islets, and its amount, estimated by measurement of the insulin-labeling optical density, was not statistically different from that in amyloid-free islets. In conclusion, even in amyloid containing islets, beta-cells maintain active insulin transcription and translation and normal insulin storage. Taking into account that in most cases only a small proportion of islets are infiltrated by amyloid, the limited reduction in proinsulin mRNA is unlikely to play a major role in the pathogenesis of diabetes. PMID- 11272185 TI - A model for assessing insulin secretion and its control under free-living conditions. PMID- 11272186 TI - beta-cell adaptation to hyperglycemia. PMID- 11272187 TI - The focal form of persistent hyperinsulinemic hypoglycemia of infancy. PMID- 11272188 TI - Oophorectomy promotes islet amyloid formation in human islet amyloid polypeptide transgenic mice. PMID- 11272189 TI - beta-cell neogenesis in type 2 diabetes. PMID- 11272190 TI - No decrease of the beta-cell mass in type 2 diabetic patients. PMID- 11272191 TI - Prospects for treatment of type 2 diabetes by expansion of the beta-cell mass. PMID- 11272192 TI - beta-cell turnover: its assessment and implications. AB - The pancreatic beta-cells are responsible for the maintenance of the body's glucose levels within a very narrow range; their population is dynamic and undergoes compensatory changes to maintain euglycemia. The structural parameters that allow mass changes (replication, neogenesis, cell volume changes, and cell death) can now be assessed and have proved to be powerful tools. Changes in one parameter can dramatically affect the beta-cell mass. Unfortunately, conclusions are often drawn on measurements that do not assess beta-cell mass but only relative volumes. Throughout the lifetime of a mammal, low levels of beta-cell replication and apoptosis are balanced and result in a slowly increasing mass. The balance allows gradual replacement of the beta-cell population; thus, beta cells should be considered a slowly renewed tissue. Two major implications of beta-cell turnover are that 1) at any time, the beta-cells would be at different ages and 2) any limitation on replacement could have dire consequences for glucose homeostasis. PMID- 11272193 TI - Regulation of beta-cell mass by hormones and growth factors. AB - Substantial new information has accumulated on molecular mechanisms of pancreas development, regulation of beta-cell gene expression, and the role of growth factors in the differentiation, growth, and regeneration of beta-cells. The present review focuses on some recent studies on the mechanism of action of cytokines such as growth hormone (GH) and prolactin (PRL) in beta-cell proliferation and gene expression-in particular, the role of signal transducers and activators of transcription (STAT) proteins. The implication of the discovery of suppressors of cytokine signaling (SOCS) proteins for the interaction between stimulatory and inhibitory cytokines, including GH, PRL, leptin, and the proinflammatory cytokines interleukin-1 and interferon-gamma, in beta-cell survival is not yet clear. Recent studies indicate a role of cell adhesion molecules and the delta-like protein preadipocyte factor 1/fetal antigen 1 (Pref 1/FA-1) in cytokine-induced beta-cell growth and development. Surprisingly, glucagon-like peptide-1 (GLP-1) was recently found to stimulate not only insulin secretion but also beta-cell replication and differentiation, which may present a new perspective in treatment of type 2 diabetes. Together with the intriguing reports on positive effects of insulin on both beta-cell growth and function, a picture is emerging of an integrated network of signaling events acting in concert to control beta-cell mass adaptation to insulin demand. PMID- 11272194 TI - Endocrine pancreas plasticity under physiological and pathological conditions. AB - Endocrine pancreas plasticity may be defined as the ability of the organ to adapt the beta-cell mass to the variations in insulin demand. For example, during late pregnancy and obesity, the increase of the beta-cell mass, in association with beta-cell hyperactivity, contributes to insulin oversecretion in response to insulin resistance. There is increasing evidence that the ability of the beta cell mass to expand in adult mammals is much higher than previously thought. During pregnancy, placental hormones, especially placental lactogens, are mainly responsible for the changes in beta-cell mass. The factors involved in beta-cell growth in obesity are far from clear, although increased free fatty acids seem to be the main candidate. Many data suggest that the impairment of insulin secretion in type 2 diabetes is partly related to reduction of beta-cell mass, at least relative to prevailing insulin demand. This defect may originate from genetic predisposition, but the situation is likely worsened by environmental factors such as hyperglycemia (glucotoxicity) and hyperlipidemia (lipotoxicity). Better understanding of beta-cell growth and regeneration mechanisms may allow new strategies in the treatment of type 2 diabetes based on early limitation of beta cell damage and/or restoration of a functional beta-cell mass. PMID- 11272195 TI - Insulin promoter factor-1 controls several aspects of beta-cell identity. PMID- 11272196 TI - Regulatory elements involved in human pdx-1 gene expression. AB - PDX-1 was shown to be expressed early during development in cells of both exocrine and endocrine origin; later it becomes restricted primarily to beta cells where it regulates the expression of beta-cell-specific genes and mediates the glucose effect on insulin gene transcription. Therefore, it was important to identify the molecular mechanisms that specifically govern the expression of pdx 1 in the mature beta-cell. To address this question, we analyzed 7 kb of the 5' flanking region of the human pdx-1 gene. By transient transfections of beta- and non-beta-cell lines with different 5' and 3' deletions of that region, a strong beta-cell-specific enhancer element located between -3.71 and 3.46 kb was revealed. We also sequenced about 4.5 kb of the human 5' flanking region and compared it with that of the mouse pdx-1 gene. This comparison revealed three short conserved regions, designated PH1, PH2, and PH3. We showed that HNF-3beta can bind and stimulate the activity of the human PH1 and PH2 elements in non-beta cells. Results reported by Wu et al. (7) and Sharma et al. (6) also indicate that expression of the mouse pdx-1 is controlled by an HNF-3-like element. Thus, it can be stated that at least some aspects of pdx-1 expression rely on the transcription factor HNF-3beta. Because HNF-3beta is not restricted to beta cells, the selective transcription of pdx-1 is likely to rely on additional factors. Our findings that the PH1 enhancer element binds both HNF-3beta and PDX 1 and that mutations in each individual site dramatically impair its transcriptional activity suggest that these factors cooperate with one another. We therefore propose that a possible feedback mechanism might control the expression of pdx-1 at different stages during development. PMID- 11272197 TI - Na-Pi cotransporter expressed during the differentiation of pancreatic AR42J cells. PMID- 11272198 TI - Expression of dominant-negative STAT5 inhibits growth hormone- and prolactin induced proliferation of insulin-producing cells. PMID- 11272199 TI - Novel pancreatic precursor cell lines for studying beta-cell differentiation. PMID- 11272200 TI - Role of apoptosis in pancreatic beta-cell death in diabetes. AB - Apoptosis is a physiological form of cell death that occurs during normal development, and critical mediators of this process include caspases, reactive oxygen species, and Ca2+. Excessive apoptosis of the pancreatic beta-cell has been associated with diabetes. Consequently, apoptosis research has focused on how infiltrating macrophages or cytotoxic T-cells might kill pancreatic beta cells using cytokines or death receptor triggering. Meanwhile, the intracellular events in the target beta-cell have been largely ignored. Elucidation of such targets might help develop improved treatment strategies for diabetes. This article will outline recent developments in apoptosis research, with emphasis on mechanisms that may be relevant to beta-cell death in type 1 and type 2 diabetes. Several of the models proposed in beta-cell killing converge on Ca2+ signaling, indicating that the pancreatic beta-cell may be an ideal system in which to carefully dissect the role of Ca2+ during apoptosis. PMID- 11272202 TI - Developmental biology of the pancreas. AB - All pancreatic cell types (endocrine, exocrine, and ductal) are derived from the same endodermal dorsal and ventral anlage, which grow together to form the definitive pancreas. Golosow and Grobstein were pioneers in the field of pancreatic developmental research, as were Wessells and Cohen, who already in the 1960s performed classic embryological experiments describing the morphogenesis of the pancreas and the epithelio-mesenchymal interactions that are instrumental for proper pancreas development. Recent findings suggest that follistatin and fibroblast growth factors represent some of these key mesenchymal factors that actively promote at least pancreatic exocrine development. The true endodermal origin of the pancreatic endocrine cells became evident by experiments performed by the groups of LeDouarin and Rutter in the 1970s. The newly acquired insights regarding the specification of pancreatic endocrine cells as controlled by the notch signaling pathway (i.e., similar to the mechanisms by which neurons are specified during neurogenesis) have provided a novel understanding of the long acknowledged similarities between neurons and the pancreatic endocrine cells. Last, the identification of a number of distinct transcription factors operating at various levels of pancreatic development and in different cell types has provided useful information both on pancreas development and on various pancreatic disorders such as diabetes. Interestingly, four of the hitherto defined five different maturity-onset diabetes of the young (MODY) genes correspond to transcription factors, and, in addition, several transcription factors have also been linked to type 2 diabetes. PMID- 11272201 TI - Roles of ATP-sensitive K+ channels in cell survival and differentiation in the endocrine pancreas. AB - To determine the roles of the ATP-sensitive K+ (K(ATP)) channels in endocrine pancreas more directly, two types of genetically engineered Kir6.2 mice were developed: mice expressing a dominant-negative form of Kir6.2 specifically in beta-cells (Kir6.2G132S Tg mice) and mice lacking Kir6.2 (Kir6.2-/- or Kir6.2 null mice). The Kir6.2G132S Tg mice show severe impairment of K(ATP) channel function only in the beta-cells, whereas Kir6.2 null mice are completely defective in K(ATP) channel function in all of the cells in which Kir6.2 is a constituent of the K(ATP) channels, because of the disruption of Kir6.2. Both types of mice show abnormal architecture of the pancreatic islets. The number of beta-cells in Kir6.2G132S Tg mice decreases markedly with age, whereas that in Kir6.2-/- mice decreases slightly. alpha-Cells, which are normally present only in the periphery of pancreatic islets, also appear in the center of the islets in both Kir6.2G132S Tg and Kir6.2-/- mice. Interestingly, the number of peptide YY (PYY) and glucagon-positive cells is markedly increased in Kir6.2 null mice, whereas the number of PP cells and delta-cells is not altered. Apoptotic cells are detected by the TdT-mediated dUTP nick-end labeling (TUNEL) method at a high frequency in both Kir6.2G372S Tg and Kir6.2-/- mice compared with the respective controls. Thus, studies of Kir6.2G372S Tg and Kir6.2-/- mice indicate that K(ATP) channels play an important role in cell survival and differentiation in the endocrine pancreas. PMID- 11272203 TI - Role of pancreatic beta-cells in the process of beta-cell death. AB - Studies on the pathogenesis of type 1 diabetes have mainly focused on the role of the immune system in the destruction of pancreatic beta-cells. Lack of data on the cellular and molecular events at the beta-cell level is caused by the inaccessibility of these cells during development of the disease. Indirect information has been collected from isolated rodent and human islet cell preparations that were exposed to cytotoxic conditions. This article reviews in vitro experiments that investigated the role of beta-cells in the process of beta cell death. beta-Cells rapidly die in necrosis because of toxic levels of oxidizing radicals or of nitric oxide; they progressively become apoptotic after prolonged culture at low glucose or with proinflammatory cytokines. Their susceptibility to necrosis or apoptosis varies with their functional state and thus with the environmental conditions. A change in cellular phenotype can alter its recognition of potentially cytotoxic agents and its defense mechanisms against cell death. These observations support the view that beta-cells are not necessarily passive victims of a cytotoxic process but can actively participate in a process of beta-cell death. Their role will be influenced by neighboring non beta-cells, which can make the islet internal milieu more protective or toxic for the beta-cells. We consider duct cells as potentially important contributors to this local process. PMID- 11272204 TI - beta-cell apoptosis: stimuli and signaling. AB - Pancreatic beta-cells are sensitive to a number of proapoptotic stimuli. Thus, apoptosis is an important part of the physiological neonatal remodeling of the endocrine pancreas, and a number of pathological stimuli involved in type 1 and type 2 diabetes have been shown to elicit beta-cell apoptosis. Factors of relevance to type 1 diabetes include proinflammatory cytokines, nitric oxide, and reactive oxygen species as well as Fas ligand. Recent findings that free fatty acids, glucose, sulfonylurea, and amylin cause beta-cell apoptosis in vitro suggest that programmed cell death may also be involved in the pathogenesis of type 2 diabetes. Furthermore, there is evidence favoring a convergence in signaling pathways toward common effectors of beta-cell apoptosis elicited by stimuli implicated in the pathogenesis of type 1 and type 2 diabetes. Therefore, recent studies involving the stimuli and signaling pathways of beta-cell apoptosis-in particular, mitogen- and stress-activated protein kinases-will be reviewed. It is concluded that immunological, inflammatory, and metabolic signals cause beta-cell apoptosis, and the possibility that these signals converge toward a common beta-cell death signaling pathway should be investigated further. PMID- 11272205 TI - beta-cell apoptosis and defense mechanisms: lessons from type 1 diabetes. AB - Increased evidence suggests that apoptosis is the main mode of beta-cell death in early type 1 diabetes. Cytokines mediate beta-cell apoptosis, and in this article, we discuss some of the cytokine-modified genes that may contribute to beta-cell survival or death. The gene encoding for the inducible form of nitric oxide synthase is induced by interleukin (IL)-1beta or IL-1beta plus gamma interferon in rodent and human islets, respectively. This leads to nitric oxide (NO) formation, which contributes to a major extent to beta-cell necrosis and to a minor extent to the process of beta-cell apoptosis. The main mode of cell death induced by cytokines in human beta-cells is apoptosis, whereas cytokines lead to both necrosis and apoptosis in rat and mouse beta-cells. It is suggested that the necrotic component in rodent islets is due to NO-induced mitochondrial impairment and consequent decreased ATP production. Human islets, possessing better antioxidant defenses, are able to preserve glucose oxidation and ATP production, and can thus complete the apoptotic program after the death signal delivered by cytokines. We propose that this death signal results from cytokine-induced parallel and/or sequential changes in the expression of multiple proapoptotic and prosurvival genes. The identity of these "gene modules" and of the transcription factors regulating them remains to be established. PMID- 11272206 TI - Imidazoline compounds protect against interleukin 1beta-induced beta-cell apoptosis. AB - Imidazoline compounds have been considered for the treatment of type 2 diabetes. We have now investigated the effects of imidazolines on interleukin (IL)-1beta induced beta-cell apoptosis and the signal transduction pathways involved. Inhibition of Ca2+ influx into beta-cells by D-600, a blocker of voltage-gated L type Ca2+ channels, suppressed IL-1beta-induced apoptosis. Our data show that calcineurin, Ca2+/calmodulin-dependent serine/threonine protein phosphatase 2B, is responsible for the effect of Ca2+ on beta-cell apoptosis. We also demonstrate that IL-1beta-mediated apoptosis correlates with expression of inducible nitric oxide synthase (iNOS) and the increase in intracellular production of nitric oxide. An inhibitor of cGMP-dependent protein kinase (PKG), KT5823, suppressed IL 1beta-induced apoptosis, suggesting the involvement of a PKG-dependent pathway in the apoptotic process. One of the major findings in this study is that imidazoline compounds RX871024 and efaroxan, suggested as prototypes of a new generation of drugs against type 2 diabetes, can protect against IL-1beta-induced apoptosis in pancreatic beta-cells, possibly by their inhibition of the expression of iNOS, a key element in the IL-1beta-induced apoptotic pathway in pancreatic beta-cells. These data suggest that imidazoline compounds should be explored as a potential therapeutic agent for the treatment of both type 1 and type 2 diabetes. PMID- 11272207 TI - Effects of serum from patients with type 1 diabetes on primary cerebellar granule cells. AB - Type 1 diabetes is an autoimmune disease of unknown etiology. Our previous work has shown that a factor present in serum from type 1 diabetic patients causes increased Ca2+ channel activity and apoptotic DNA fragmentation in pancreatic beta-cells. Here we examined the effects of type 1 diabetic serum on primary cerebellar granule cells (CGCs). In CGCs, exposure to type 1 diabetic serum did not cause increased apoptosis or changes in Ca2+ channel activity. However, patient serum did cause modulation of Ca2+ signals in a cell type with triangular soma that exhibited low voltage-gated Ca2+ currents. This cell was present primarily in cultures exposed to type 1 diabetic serum. The presence of low voltage-gated Ca2+ currents and long neuronal dendrites indicated that this unique cell was of neuronal origin and not of glial origin. PMID- 11272208 TI - Molecular regulation of Fas expression in beta-cells. PMID- 11272209 TI - Early development of beta-cells is impaired in the GK rat model of type 2 diabetes. AB - The Goto-Kakisaki (GK) rat is a genetic model of type 2 diabetes obtained by selective inbreeding of mildly glucose-intolerant Wistar rats. Previous studies have shown that at birth, the beta-cell mass of the GK rat is severely reduced compared with that of the Wistar rat. Therefore, beta-cell deficit could be the primary defect leading to type 2 diabetes in this model. To identify the abnormality at the origin of the beta-cell mass deficit, we compared the fetal development of GK and Wistar rats. Our study reveals that during early development (embryonic day 12-14 [E12-14]), GK fetuses present a delayed global growth that progressively recovers: at birth, no size or weight difference persists. However, from E18 onward, the weight and DNA content of the pancreas and liver are reduced by 30% in the GK fetuses. Cell proliferation is reduced in the GK pancreas from E16 to E20. Whereas apoptotic cells are scarce in the Wistar fetal pancreas, a wave of apoptosis from E16 to E18 was detected in the GK pancreas. Analysis of pancreas differentiation revealed that from E12 to E14, there are no significant differences in the number of alpha- and beta-cells between the GK and Wistar pancreas. However, by E16, the average number of beta cells in the GK pancreas represents only 50% that of the Wistar pancreas, and this difference persists until birth. The number of alpha-cells was reduced by 25% from E18 to E21. To determine whether the defect in GK pancreas development depends on intrinsic pancreatic factors or on endocrine extrapancreatic factors, we performed in vitro cultures of E12 pancreatic rudiments. The cultures show that in vitro, the growth and endocrine differentiation of the GK and Wistar pancreatic rudiments are identical. Thus, impaired development of the GK pancreas probably results from insufficiency of extrapancreatic factor(s) necessary for the growth and survival of fetal pancreatic cells. PMID- 11272210 TI - beta-cell function and viability in the spontaneously diabetic GK rat: information from the GK/Par colony. AB - The GK rat model of type 2 diabetes is especially convenient to dissect the pathogenic mechanism necessary for the emergence of overt diabetes because all adult rats obtained in our department (GK/Par colony) to date have stable basal mild hyperglycemia and because overt diabetes is preceded by a period of normoglycemia, ranging from birth to weaning. The purpose of this article is to sum up the information so far available related to the biology of the beta-cell in the GK/Par rat. In terms of beta-cell function, there is no major intrinsic secretory defect in the prediabetic GK/Par beta-cell, and the lack of beta-cell reactivity to glucose (which reflects multiple intracellular abnormalities), as seen during the adult period when the GK/Par rats are overtly diabetic, represents an acquired defect (perhaps glucotoxicity). In terms of beta-cell population, the earliest alteration so far detected in the GK/Par rat targets the size of the beta-cell population. Several convergent data suggest that the permanently reduced beta-cell mass in the GK/Par rat reflects a limitation of beta-cell neogenesis during early fetal life, and it is conceivable that some genes among the set involved in GK diabetes belong to the subset of genes controlling early beta-cell development. PMID- 11272211 TI - beta-cell genes and diabetes: molecular and clinical characterization of mutations in transcription factors. AB - beta-Cell transcription factor genes are important in the pathophysiology of the beta-cell, with mutations in hepatocyte nuclear factor (HNF)-1alpha, HNF-4alpha, insulin promoter factor (IPF)-1, HNF-1beta, and NeuroD1/BETA2, all resulting in early-onset type 2 diabetes. We assessed the relative contribution of these genes to early-onset type 2 diabetes using linkage and sequencing analysis in a cohort of 101 families (95% U.K. Caucasian). The relative distribution of the 90 families fitting maturity-onset diabetes of the young (MODY) criteria was 63% HNF 1alpha, 2% HNF-4alpha, 0% IPF-1, 1% HNF-1beta, 0% NeuroD1/ BETA2, and 20% glucokinase. We report the molecular genetic and clinical characteristics of these patients including 29 new families and 8 novel HNF-1alpha gene mutations. Mutations in the transactivation domain are more likely to be protein truncating rather than result in amino acid substitutions, suggesting that a relatively severe disruption of this domain is necessary to result in diabetes. Mutations in the different transcription factors result in clinical heterogeneity. IPF-1 mutations are associated with a higher age at diagnosis (42.7 years) than HNF 1alpha (20.4 years), HNF-1beta (24.2 years), or HNF-4alpha (26.3 years) gene mutations. Subjects with HNF-1beta mutations, in contrast to the other transcription factors, frequently present with renal disease. A comparison of age at diagnosis between subjects with different types and locations of HNF-1alpha mutations did not reveal genotype-phenotype correlations. In conclusion, mutations in transcription factors expressed in the beta-cell are the major cause of MODY, and the phenotype clearly varies with the gene that is mutated. There is little evidence to indicate that different mutations within the same gene have different phenotypes. PMID- 11272212 TI - Preparation, premedication, and surveillance. AB - The endoscopic literature published during the past year has once again confirmed that there is significant variation from country to country regarding whether or not patients wish to receive conscious sedation during endoscopy (and particularly colonoscopy) - and there may even be variation from one endoscopic unit to another within the same country. Particular attention has been given to attempts to identify "ideal" candidates for conscious sedation, and to the importance of providing patients with information before the procedure. It has been shown that patients who receive detailed information about a medical procedure beforehand are able to benefit from this. The role of benzodiazepines, particularly midazolam, was investigated in studies emphasizing that the dosage should be kept to the minimum that is compatible with patient comfort and successful performance of the procedure. There have been few publications comparing propofol with midazolam. As expected, in view of the known pharmacological properties of the two drugs, the quality of sedation was better and the recovery time was shorter in patients who were treated with propofol. However, important questions are still open regarding the narrow therapeutic range of propofol and the methods by which it is administered (by endoscopists or by anesthesiologists). An important aspect of sedation procedures is the prevention of hypoxia and cardiopulmonary complications. Recent endoscopic reports have added little further information concerning the well-known risk of oxygen desaturation during conscious sedation. Performing endoscopy in unsedated patients reduces, but does not eliminate, the risk of hypoxia. Among the various risk factors, it has been found that chronic respiratory failure and coronary heart disease are factors predictive of severe desaturation and relevant electrocardiographic changes. The use of electronic monitoring techniques with pulse oximetry is recommended as a standard procedure during digestive endoscopy; however, it has been observed that when supplemental oxygen is administered, pulse oximetry no longer reflects normal ventilatory function and does not detect episodes of severe CO2 retention. Transcutaneous measurement of PCO2 therefore seems more reliable as a means of assessing hypoventilation. Several papers have proposed "ideal formulas" for bowel preparation for endoscopic procedures. Various regimens have been proposed as alternatives to polyethylene glycol electrolyte lavage solution (PEG-ELS) and sodium phosphate compounds, with different results. On the whole, there is still little information regarding the best and most cost-effective method of bowel cleansing for colonoscopy and flexible sigmoidoscopy. PMID- 11272213 TI - Gastroesophageal reflux disease and Barrett's esophagus. AB - Gastroesophageal reflux disease (GERD) is a common clinical problem. Circumstantial evidence continues to suggest that infection with Helicobacter pylori may protect some patients from developing GERD and its complications. An empirical trial of a proton-pump inhibitor may now be a reasonable alternative to endoscopy or 24-hour pH testing for the diagnosis of GERD. Long-term follow-up data covering more than over a decade indicate that proton-pump inhibitors are effective and safe agents for the treatment of GERD. Furthermore, a strategy of proton-pump inhibitors first may be the most cost-effective approach to GERD. It remains unclear why some patients with GERD develop Barrett's esophagus, whereas others do not. Recent studies demonstrate the importance of pulses of acid or bile in increasing cell proliferation and cyclooxygenase-2 expression in Barrett's epithelium cell cultures. Short-segment Barrett's esophagus is now clearly associated with an increased risk of dysplasia or cancer compared to intestinal metaplasia of the cardia, and the cancer risk in this condition is similar to that with long-segment Barrett's esophagus. However, the overall cancer risk in patients with Barrett's esophagus is lower than previously estimated, at approximately 0.5% annually. Ablation techniques continue to show promise, but are not yet ready for routine clinical use. Endoscopic mucosal resection is a new treatment option for selected patients with high-grade dysplasia or superficial esophageal adenocarcinoma. PMID- 11272214 TI - Treatment of esophageal and gastric tumors. AB - In the diagnosis of early cancer, there are differences in the pathological criteria used by Western and Asian (Japanese) pathologists. The Vienna classification advocated by pathologists offers standard pathological criteria common to all endoscopists, and it has clarified the indications for the treatment of superficial lesions, including high-grade dysplasia and mucosal cancer. Endoscopic mucosal resection (EMR) is increasingly being used in the treatment of early cancer. Experience with EMR in the treatment of Barrett's esophagus with cancer has been reported, and the preliminary results are encouraging. Some technical variations and improvements in EMR procedures have been described. As an injection agent, the use of mucinous substances such as sodium hyaluronate has been reported. A cutting knife with an insulated tip has been designed, making the use of the precutting technique much safer. Studies have been conducted comparing the freehand technique with the cap technique for EMR, and it was found that the cap technique is generally better. Ablative treatment has also been used in many cases, with satisfactory results. In advanced cancer, self-expanding metallic stents have been used for palliative treatment, with generally satisfactory results. The range of applications for therapeutic endoscopy has continued to expand during the last two years in the treatment of esophageal and gastric tumors. PMID- 11272215 TI - Variceal bleeding and portal hypertension: still a therapeutic challenge? AB - In the primary prevention of variceal hemorrhage, beta-blockers continue to be the first-line treatment. Newer nonselective beta-blockers with anti-alpha1 adrenergic activity, such as carvedilol, appear to have a better impact on reducing the hepatic venous pressure gradient than propranolol. The addition of isosorbide mononitrate appears to improve the effectiveness of beta-blockers in primary prophylaxis, but not that of somatostatin in the treatment of acute variceal hemorrhage. The use of vasoactive drugs alone in acute variceal bleeding has not proved to be more effective than endoscopic treatment. The advent of endoscopic variceal ligation (EVL) has strengthened the role of endoscopy in the management of bleeding esophageal varices. EVL has improved the results, particularly in terms of lowering the treatment-related morbidity, compared with endoscopic variceal sclerotherapy (EVS). However, the variceal recurrence rate after initial eradication with EVL is relatively high. In contrast to synchronous combined therapy with EVL plus EVS, metachronous combination of EVL and low-dose EVS may improve the results of EVL alone. For bleeding fundic varices, obliteration using cyanoacrylate is currently the treatment of choice. Endosonography (EUS) is coming into more widespread use in the assessment of variceal eradication and in further attempts to improve the results of endoscopic injection therapy. According to two meta-analysis studies, transjugular intrahepatic portosystemic shunt (TIPS) is not yet capable of replacing endoscopic treatment in the secondary prevention of variceal bleeding. PMID- 11272216 TI - Inflammatory bowel disease. AB - The pathogenesis of inflammatory bowel disease (IBD) continues to be explored, with the emphasis on genetically determined host immune dysregulation as it applies to interactions with luminal bacteria. The role of endoscopic ultrasound in the evaluation of IBD continues to be studied. Recent advances in the evaluation of enterocutaneous fistulas have been made through the use of hydrogen peroxide to enhance definition of the fistula course and characteristics of the neighboring bowel, as evaluated by transabdominal ultrasound. Refinements have been made in the use and interpretation of the results of surveillance colonoscopy for detection of colorectal cancer and dysplasia in IBD, but a consistent approach still needs to be applied by individual practitioners. New data exist on the usefulness of methotrexate for maintenance of remission, while a lack of efficacy has been demonstrated for mesalamine in the prevention of postoperative recurrence. The role of anti-tumor necrosis factor (TNF) antibody therapy in the treatment of IBD continues to be investigated. PMID- 11272217 TI - Diagnostic endoscopic retrograde cholangiopancreatography. AB - Since the introduction of magnetic resonance cholangiopancreatography (MRCP), the focus in endoscopic retrograde cholangiopancreatography (ERCP) has shifted from diagnosis to treatment - a change that has organizational implications in relation to teaching and providing access to ERCP. Most of last year's papers on ERCP described refinements of the technique and indication, tissue sampling, and efforts to reduce complications. Many studies compared MRCP with various other imaging methods and with histopathological findings. Bile duct stones and strictures are still the main target, but new entities for evaluation with MRCP have been found: primary sclerosing cholangitis, choledochal cysts, chronic pancreatitis, pancreatic injury, and postoperative abnormalities. Helical computed-tomographic cholangiography is still attracting some interest. This review closes with a comparison of ERCP with MRCP with regard to availability, legal aspects, operator-dependency, and cost-effectiveness. PMID- 11272218 TI - Endoscopic ultrasonography. AB - Twenty years after the introduction of endoscopic ultrasonography, many papers on the topic are still being published every year in the medical literature. Along with established clinical indications, such as gastrointestinal and pancreatic cancer staging and differential diagnosis of submucosal tumors, new applications have been suggested, such as mediastinal and liver tumor sampling with fine needle aspiration. Improved accuracy and cost-effectiveness have been demonstrated in comparison with other imaging techniques. Reports of large series of fine-needle aspiration procedures have described a high level of accuracy for EUS in the diagnosis of lymph nodes and perivisceral masses. Pancreatic and ampullary tumors still represent a major challenge, as shown by numerous articles describing differential-diagnostic criteria and cytological sampling techniques. A few papers have also been published on the topic of portal hypertension, but it seems questionable whether there is any real advantage for endoscopic ultrasonography over traditional endoscopy here. New techniques such as radiofrequency tumor ablation are promising, while others such as three dimensional imaging and the use of contrast enhancement have not yet met with routine clinical application. Finally, some of the papers published during the last year have studied the technique of endoscopic ultrasonography itself, dealing with issues of outcome, current clinical availability and use, and the learning curve. Evidently, endoscopic ultrasonography is still widely underused - not only among general practitioners and physicians in other specialties, but even by gastroenterologists. Although endoscopic ultrasonography is already 20 years old, considerable efforts are still needed, therefore, to ensure that it becomes more widely accepted in clinical practice. PMID- 11272219 TI - Recurrent cholangitis following "successful" treatment for gallbladder cancer. PMID- 11272220 TI - Hemostatic clip in gastrointestinal bleeding. PMID- 11272221 TI - Concomitant manifestation of achalasia and sphincter of Oddi dysfunction. AB - We report the case of a 44-year-old woman who suffered from biliary-type pain after cholecystectomy, dysphagia, and weight loss. Examinations revealed sphincter of Oddi dysfunction (SOD) and achalasia. Complete relief of symptoms was achieved by endoscopic sphincterotomy and pneumatic dilation of the distal esophagus. This case report demonstrates for the first time the concomitant manifestation of two motility disorders of the upper gastrointestinal tract, i.e. achalasia and SOD. At present, any causal relationship seems speculative; however, both diseases were successfully treated using endoscopic procedures. PMID- 11272222 TI - Three cases of fistulae arising from gastrointestinal tract treated with endoscopic injection of Histoacryl. AB - Treatment of gastrointestinal fistula usually consists of conservative management and surgery as definite therapeutic measure. Histoacryl can be used to treat gastrointestinal fistula with no response to conservative management in surgically high-risk patients. We report here three cases of gastrointestinal fistula successfully treated with endoscopic injection of Histoacryl through a catheter into the internal opening and fistulous tract. PMID- 11272223 TI - Guidelines of the French Society of Digestive Endoscopy: endoscopic mucosectomy. PMID- 11272224 TI - Intraductal papillary mucinous tumor of the bile duct: why not? PMID- 11272225 TI - Accidental arytenoid banding: an unusual complication of single-shot ligation therapy. PMID- 11272226 TI - Multiple strictures in jejunal Crohn's disease: push enteroscopy dilation. PMID- 11272227 TI - Proximal displacement of biliary stent with distal perforation and impaction in the pancreas. PMID- 11272229 TI - Endosonographic Features of advanced gastric carcinoma invading the colon: is it easy to differentiate from pancreatic invasion? PMID- 11272228 TI - Usefulness of Maalox for detection of the precise bleeding points and confirmation of hemostasis on gastrointestinal hemorrhage. PMID- 11272231 TI - [First aid service in Germany]. PMID- 11272230 TI - Rapid worsening of esophageal varices in a patient with small hepatocellular carcinomas after percutaneous ethanol injection therapy. PMID- 11272232 TI - [When generic non-prescription drugs are administered and yet everything goes well.... The problem of medical studies on infants and children]. PMID- 11272233 TI - [Ethics questions concerning infant death]. PMID- 11272234 TI - [Alternative medicine and parent involvement ... possibilities of a student project]. PMID- 11272235 TI - [Lusting for sweets--psychological aspects of sweet foods]. PMID- 11272236 TI - [Current requirements of pediatric care--pedagogic answers]. PMID- 11272237 TI - [Intervention based on nursing theory in families with a chronically ill child I]. PMID- 11272238 TI - [Lack of treatment in growing patient groups--congenital heart defects in adults]. PMID- 11272239 TI - [22nd annual meeting of the German Pediatric Nursing Association, Stuttgart, Germany 2000]. PMID- 11272240 TI - [Status of the controversial discussion of the pathogenesis and treatment of chronic otitis media with effusion in childhood]. AB - BACKGROUND: Chronic otitis media with effusion is one of the most common diseases in childhood. The causes of otitis media are unclear as far as their importance is concerned. METHOD: Extensive search in the German and Anglo-saxon literature with following presentation of the results. RESULTS: The original idea of a mechanical obstruction of the eustachian tube by the adenoids seems to be insufficient. Inflammatory mechanisms as a result of pathogenic germs in the middle ear fluid stand opposite to immunomodulating mechanisms as relevant causes of chronic otitis media with effusion. There are references which lead to the importance of allergic co-mechanisms individually. The controversial discussion of the pathophysiology of chronic otitis media with effusion is illustrated by the different opinions of optimal treatment modalities. Varying medical treatment opportunities exist while centrally the importance of the use of antibiotics is controversial. Based on the absence of long lasting effects of medical treatments, there are groups supporting a wait-and-see policy. Concerning the long-term effects of operative treatment, the importance of isolate tubes insertion in opposition to a combined treatment modality (adenotomy and tubes) is controversial. The actual debate focusses on adenotomy or adenotonsillectomy without operating the middle ear as treatment modality in patients with otitis media with effusion. CONCLUSIONS: Economic and social importance of this disease forces further investigations in pathogenesis and optimizing of medical and operative treatment modalities. PMID- 11272241 TI - [Epithelium-stroma interaction in cholesteatoma of the middle ear]. AB - BACKGROUND: Local Infiltration and bony destruction are characteristic features of cholesteatomas. The aim of the study was assessment of cell ploidy, proliferation rates and expression of cell adhesion molecules to analyze the pathogenetic role of matrix (epithelium) in cholesteatoma. The cellbiologic parameters were compared to clinical findings. PATIENTS AND METHOD: Tissue samples from 48 patients with cholesteatomas were analyzed by: routine histology, quantitative DNA-cytometry with the DNA indices: 2cDeviation Index (2cDI) and 5c Exceeding Rate (5c ER), immunhistochemical analysis of proliferation rate (ki67 MIB1 and PCNA), cell adhesion molecules, cell-cell interaction: E-Cadherin, alpha 1 beta 6-Integrin, Inter-Cellular-Adhesion-Molecule (I-CAM), cell-matrix interaction: CD44v4/5, CD 44v6, alpha v-, beta 3-Integrinchains and vascular-Cell Adhesion-Molecule (V-CAM). Clinical data included patient age, history of ear disease, pre-operative audiometry, intra-operative size and extension of the cholesteatoma, destruction of ossicles and petrous bone. For comparison healthy squamous epithelium was obtained from the external ear canal of 10 patients during stapes surgery. RESULTS: Ossicular destructions were found in 34 cases. Three patients had mesotympanic cholesteatomas, four patients had mesotympanic and epitympanic involvement. In 37 patients cholesteatomas extended into the mastoid and in four patients the perilabyrinth and the petrous apex were reached. DNA-cytometric examination of matrix showed normal diploid values and no aneuploid cells (DNA-content > 5c) in all patients. The proliferation rates of the matrix were increased in comparison to normal epithelium. Cell adhesion molecules for intercellular bindings were expressed in similar pattern in cholesteatomas and in normal epithelium. Cell adhesion molecules for cell matrix bindings showed new or increased expression in cholesteatomas. No significant correlation between proliferation and clinical findings could be established. CONCLUSION: The study confirms previous suggestions that the growth of cholesteatomas is not stimulated by the matrix. The increased proliferation of the matrix is a result of the inflammatory process in the cholesteatoma and is correlated to the size of the cholesteatoma. On a cellular or molecular level no correlation between bone destruction through the cholesteatoma and proliferation rate of the cholesteatomas could be established. These findings support suggestions that the perimatrix of the cholesteatomas is the main pathogenetic factor. PMID- 11272242 TI - [Unilateral hearing loss in childhood. An empirical analysis comparing bilateral hearing loss]. AB - BACKGROUND: Unilateral cochlear hearing loss is considered as a risk factor for auditory, verbal-communicative, behavioral and academic development. An early diagnosis is therefore necessary. METHOD/PATIENTS: 182 consecutive patients with an age up to 10 years were diagnosed with permanent hearing loss in the mild to profound range in a defined 5-years-period (1.10.1994-30.9.1999) in the Pedaudiologic Outpatient Clinic of the University Gottingen. Fifty children (27.5%) had a unilateral hearing loss (30 sensorineural, 20 conductive caused by aural atresia with or without microtia), 132 a bilateral one. This paper presents the data of the consecutive series of the 30 sensorineural unilaterally hearing impaired children (> 25 dB). RESULTS: A slight male predominance was present (53.3%). The left ear was affected in 43.3%, the right ear in 56.7%. In the majority of the cases the hearing loss was severe and profound. The hearing impairments were diagnosed by the median age of 69.5 months and all aided by the median age of 70 months. The etiology remained unknown in 60 per cent of the cases. Hearing aid acceptance at the first follow-up (on average after 6 months) was found to be 79 per cent. CONCLUSIONS: The data suggest the relevance and necessity of a pedaudiometric prevention. They demonstrate the urgent necessity of a molecular genetic cause investigation. Recessive sensorineural hearing loss with onset in infancy may exist with no antecedent family history. PMID- 11272243 TI - [DNA repair if mucous membrane cells and lymphocytes with the comet assay]. AB - BACKGROUND: Exogenous and endogenous risk factors are involved in human carcinogenesis of the head and neck. Noteworthy hereditary factors include mutagen sensitivity and the individual's capacity for DNA repair. Repair mechanisms influence different phases of mutation and malignant transformation. The present study introduces a highly sensitive method for evaluating the repair capacity of human mucosal cells and lymphocytes. METHOD: Human epithelia of the nose and peripheral lymphocytes were incubated with the tobacco-related carcinogen N'nitrosodiethylamine (NDEA). The solvent dimethylsulfoxide (DMSO) served as negative control. Following repair times of 0 min, 15 min and 30 min, the cells were subjected to a modified version of the alkaline microgel electrophoresis technique (Comet assay). The data were digitally analyzed after fluorescent staining. RESULTS: Using the Comet assay, DNA repair could be quantified in human mucosal cells and in lymphocytes. The majority of DNA strand breaks induced by NDEA were repaired within 15 min in both cell types. CONCLUSIONS: Up to now, the Comet assay has been the preferred method for demonstrating substance-induced DNA damage. It has been used in repair studies involving lymphocytes, bacterial systems and animal-derived cells. A modified version of this method, however, can be used to quantify DNA repair in human mucosal cells and peripheral lymphocytes targeted by carcinogens. It is thus possible to evaluate an endogenous factor involved in the development of malignant transformations in mucosal cells of the upper aerodigestive tract. PMID- 11272244 TI - [Lymph node metastasis in head-neck tumors]. AB - BACKGROUND: One of the most important criteria of malignancy of head and neck cancer are the cervical lymph metastases. Being significant for the therapeutical plan is how tumor depending parameters like T-stage, degree of differentiation and tumor localisation will influence the N-stage and therefore the extension of neck dissection. METHOD: To evaluate the pattern of formations of metastases and the success of therapy a retrospective study was performed on 405 patients with carcinoma of the oral cavity (n = 47), the oropharynx (n = 117), the hypopharynx (n = 47) and the larynx (n = 193). RESULTS: By the time of surgery carcinoma of the hypopharynx were most frequently accompanied by cervical metastases (80%), followed by carcinoma of the oropharynx (70%), the oral cavity (52%) and the larynx (26%). Occurrence and extension of regional lymph node metastases correlated well with T-stages and degree of differentiation. After surgical therapy locoregional recurrence could be observed in 5.2% of the patients. Five year-survival rate was reduced to 50% on patients with positive lymph nodes. The different tumour sites showed preferred patterns of metastatic spread, without complete avoidance of certain levels. CONCLUSION: For the decision on indication and extent of neck dissection the preoperative diagnostic (ultrasound, CT-scan, MRI), localisation of tumour, T-stage, degree of differentiation and the knowledge of typical metastatic spread must be considered. PMID- 11272245 TI - [Schwannoma of the tongue]. AB - BACKGROUND: Schwannoma, in the English literature mostly called neurilemmoma, are well known in the ear-nose-and-throat-medicine as benign tumors of the nerve sheath, for example as neuroma of the acoustic nerve. They are seldom found at other peripheral nerves of the head. The localisation in the tongue has so far only been described in some individual cases. CASE REPORT: We present a case of a 28-year-old woman with a schwannoma of the tongue, measuring 4.0 x 2.2 x 3.2 cm. The case history and the radiological investigations suggested a malignant tumor. During the operation the lump revealed itself as being encapsulated. Under the histological examination it was discovered to be a Schwannoma. CONCLUSIONS: We would like to add a further case of a schwannoma of the tongue to the so far published cases, thus allowing this possibility to be considered in general diagnosis. PMID- 11272246 TI - [Low-grade malignant peripheral nerve sheath tumor of the neck soft tissues]. AB - BACKGROUND: Malignant Peripheral Nerve Sheath Tumours (MPNST) either grow sporadically, after radiation or chemotherapy respectively. In many cases they are associated with Neurofibromatosis I. Because of the multiform histologic picture they are often difficult to differentiate from other soft tissue tumours. PATIENT: We present the case of a sporadic MPNST which developed from the vagus nerve of a 39-year-old patient following radiation of the neck 7 years before. After complete excision there has been no recurrence up to now. RESULTS AND CONCLUSIONS: Sporadic MPNST of the head and neck are comparatively rare. With regard to the strong association with Neurofibromatosis I and the difficult differential diagnosis to other soft tissue tumours the emphasis should be put on excluding further manifestations of Neurofibromatosis I and of secondary tumours. PMID- 11272247 TI - [Juvenile hyaline fibromatosis (JHF)]. AB - The juvenile hyaline fibromatosis (JHF) is a rare tumorous autosomal recessive disease of the connective tissue. The etiopathogenesis of the disease is unknown. Typical diagnostic criteria are multiple hyaline subcutaneous fibroma, filamentous tumors of the skin, gingival hypertrophy, muscle contractures of the extremities and multiple osteolytic bone destructions. Involvement of visceral organs is not described. Mental development and life expectancy are normal. Until today, no causal treatment of the JHF exists. Surgical excision of the dermal tumors is indicated for functional and aesthetic improvement. Complete excision should be executed in the early phase of their development. This has shown the best functional and aesthetic results, especially in facial areas. The progress of the disease is individual. Due to frequent recurrencies, repeated resections and revisions may be necessary. Interdisciplinal counseling and therapy, particularly physiotherapy, may partially improve the state of function. PMID- 11272248 TI - [Sinus floor augmentation with simultaneous implant insertion using recombinant human osteogenic protein-1]. AB - INTRODUCTION: The purpose of this study was to examine whether the combination of an osteoinductive protein (recombinant human osteogenic protein-(rhOP-1 = bone morphogenetic protein-7) with natural bovine bone mineral (BioOss) would improve ossification and the bone-implant contact (BIC) in a sinus floor augmentation with simultaneous placement of implants. MATERIAL AND METHODS: In this study, the maxillary sinus floors in five miniature pigs were augmented with 3 ml BioOss containing 420 micrograms rhOP-1 on the test side and 3 ml BioOss alone on the control side. At the time of augmentation a titanium implant (ITI) was inserted from a laterocaudal direction. RESULTS: After 6 months of healing the sites of augmentation were removed and examined in non-decalcified sections by microradiography, fluorescence microscopy of sequentially labelled specimens and by histometry. On both sides, significant amounts of newly formed bone were observed. However, on the test sites, the BIC in the augmented area was 80.0% versus 38.6% on control sites. It can be concluded that the application of bone morphogenetic proteins caused a more rapid and enhanced osseointegration of simultaneously placed implants when compared to the bone substitute alone. DISCUSSION: Therefore recombinant human osteogenic protein-1 delivered by natural bone mineral has the potential to become a clinical alternative for autogenous bone grafts in sinus floor augmentation. PMID- 11272249 TI - [Diving medicine aspects in otorhinolaryngology. II. Diving-specific illnesses and dysfunctions as well as assessment for diving fitness]. AB - Scuba diving with compressed air has become a recreational sport that can be performed at all stages of adulthood. The human body including all gas-filled cavities are exposed to an increased ambient pressure during a dive. In the present review article, specific aspects of diving related disorders are that are of importance in the otolaryngology field are presented and discussed: the multitude of causes for divers' vertigo and the so called divers ear. Furthermore, useful recommendations in the assessment of physical fitness for diving are presented. This review will provide a background and foundation for both, an adequate treatment of these diseases and a critical and responsible health education of the diver. PMID- 11272250 TI - [Interesting case no. 41. Lymph node metastasis of a germ cell tumor]. PMID- 11272251 TI - [Classification of complex signal patterns with artificial neural networks in hearing and voice diagnosis]. PMID- 11272252 TI - [Laser surgery of laryngeal and hypopharyngeal carcinoma. II]. PMID- 11272253 TI - Fluoroquinolones inhibit preferentially Streptococcus pneumoniae DNA topoisomerase IV than DNA gyrase native proteins. AB - The genes encoding the subunits of DNA topoisomerase IV (parC and parE) and DNA gyrase (gyrA and gyrB) of Streptococcus pneumoniae were cloned and overproduced in Escherichia coli by using the T7promoter-T7 RNA polymerase system. The four subunits were separately purified to near homogeneity by column chromatography. Protein purification was achieved by DEAE-sepharose, heparin-agarose, and hydroxylapatite chromatography. DNA topoisomerase IV was reconstituted when ParC and ParE were combined at a 3.8-fold excess of ParE. The reconstituted topoisomerase IV showed to generate efficient ATP-dependent DNA decatenation activity. The DNA gyrase ATP-dependent supercoiling activity was reconstituted by mixing equimolar amounts of the two gyrase subunits. The inhibitory effects of four representative fluoroquinolones on the DNA decatenation activity of topoisomerase IV and DNA supercoiling of gyrase have been examined and compared. All four compounds were more active in inhibiting topoisomerase IV than gyrase. Moreover, there was a positive correlation between the inhibitory activity against topoisomerase IV decatenation and DNA gyrase supercoiling. The classification of the four fluoroquinolones, considering their inhibitory activities in decatenation, supercoiling and growth was the following: clinafloxacin > trovafloxacin > sparfloxacin > ciprofloxacin. These results suggest these drugs primarily target topoisomerase IV of S. pneumoniae, and gyrase secondarily, in agreement with genetic data. PMID- 11272254 TI - Clonal origin of the type 37 Streptococcus pneumoniae. AB - It has been recently reported that different type 37 clinical isolates of Streptococcus pneumoniae have an identical tts gene directing the formation of type 37 capsular polysaccharide. Here we show that type 37 S. pneumoniae strains isolated in two different continents (Europe and America) some 60 years apart frequently gave rise to nontypable variants upon in vitro cultivation. The tts gene from three independent nontypable mutants was PCR amplified and sequenced showing different classes of inactivating mutations. Furthermore, pulsed-field gel electrophoresis and multilocus sequence typing demonstrated that the type 37 pneumococcal isolates studied so far constitute a highly related strain cluster (a clonal complex), and strongly suggested that every type 37 pneumococcus has spread globally from a single, old clone. PMID- 11272255 TI - Mutations in the interdomain loop region of the tetA(A) tetracycline resistance gene increase efflux of minocycline and glycylcyclines. AB - A novel class of tetracyclines, the glycylcyclines, have been shown to be active against bacterial strains harboring genes encoding tetracycline efflux pumps. However, two veterinary Salmonella isolates that carried tetracycline resistance determinants of the tetA(A) class were found to have reduced susceptibility to glycylcyclines, especially two early investigational glycylcyclines, DMG-MINO and DMG-DMDOT. These isolates were also quite resistant to tetracycline and minocycline. The isolates, one a strain of S. cholerasuis and the other, S. typhimurium, both carried the same novel tetA(A) variant, based on DNA sequencing, with one determinant plasmid encoded and the other located on the chromosome. This tetA(A) variant was cloned and shown to provide reduced susceptibility to the glycylcycline class although GAR-936, a glycylcycline currently in clinical development, was the least affected. The novel tetA(A) gene carries two mutations in the largest cytoplasmic loop of the efflux pump, which causes a double frameshift in codons 201, 202, and 203. This "interdomain region" of the efflux pump has generally been regarded as having no functional role in the efflux of tetracycline but the double frameshift is most likely responsible for the enhanced resistance observed and points to an interaction that was previously unrecognized. Mutants of the tetA(B) class with decreased susceptibility to the glycylcyclines were also generated in vitro. These all carried mutations in the portion of the tetA(B) gene encoding a transmembrane spanning region of the efflux pump. The laboratory-generated mutants point to the tight constraints in substrate recognition of the transmembrane-spanning region and may suggest that it will be the interdomain region of the pump that is likely to be the locus of future glycylcycline resistance mutations as these compounds enter clinical use. PMID- 11272257 TI - Streptococcus pyogenes collected in Torino (northwest Italy) between 1983 and 1998: survey of macrolide resistance and trend of genotype by RAPD. AB - We surveyed macrolide resistance in 1,086 isolates of Streptococcus pyogenes, collected between 1983 and 1998, from throat swabs of children with untreated pharyngotonsillitis living in Torino (northwest Italy). In 1983 and 1985, the frequency of erythromycin resistance was 10%, and from 1990 to 1992 it was 4%. However, it rose to 16.6% in 1994 and reached 51% in 1996 before decreasing to 38.5% in 1998. Characterization of the phenotype of resistant isolates revealed the prevalence of constitutive resistance (CR) in 1996, whereas the M phenotype, characterized by resistance to 14- and 15-membered macrolides with susceptibility to clindamycin and streptogramin B, prevailed in 1998. Moreover, in 1997 we observed an increase in the frequency of autoagglutinating bacteria and, in 1998, of OF-negative S. pyogenes. Meanwhile, penicillin tolerance, assessed in the isolates collected from 1990 to 1996, decreased and disappeared. Random amplification of polymorphic DNA (RAPD) was used to obtain the genomic profile of 32 S. pyogenes strains. Four main DNA profiles were demonstrated, generally related to the macrolide-resistance phenotype and for the major part to the T serotype. These results indicate that RAPD is reliable as a first screening method in the epidemiological characterization of resistant S. pyogenes. PMID- 11272256 TI - Correlation of pncA sequence with pyrazinamide resistance level in BACTEC for 21 mycobacterium tuberculosis clinical isolates. AB - Mutations in the pncA gene, encoding pyrazinamidase, are considered the major mechanism of pyrazinamide (PZA) resistance in Mycobacterium tuberculosis, but resistant strains containing the wild-type gene have been described. The correlation of pncA sequence with PZA resistance level was examined for 21 M. tuberculosis clinical isolates. Susceptibility patterns were determined for 100, 300, and 900 microg/ml concentrations of the drug in BACTEC. Insertions and deletions and a substitution in the putative promoter region led to high-level resistance, whereas substitutions within the open reading frame seemed to confer variable levels of resistance. Variable resistance levels and PZase activities were also observed among isolates lacking pncA mutations. The high-level resistance (900 microg/ml) in pncA wild-type isolates highlights the clinical significance of these isolates. These data also suggest that there may still be more than one alternative mechanism leading to PZA resistance in M. tuberculosis isolates. PMID- 11272258 TI - Detection of multiresistant ceftazidime-susceptible Klebsiella pneumoniae isolates lacking TEM-26 after class restriction of cephalosporins. AB - A multitude of extended spectrum beta-lactamases (ESBLs) have evolved in response to the use of late generation cephalosporins. In those hospitals where Klebsiella pneumoniae and other bacteria possessing these enzymes flourish, many interventions have been applied to reduce this trend. We instituted a policy of class restriction of cephalosporins in our hospital in 1996 that led to a 44% reduction in ceftazidime-resistant K. pneumoniae hospital-wide and an 87% decrease in the surgical intensive care unit. Another interesting outcome of this strategy was the identification of multiresistant K. pneumoniae, which was now susceptible to ceftazidime. Characterization of these novel isolates demonstrated that the TEM-26 enzyme, which was responsible for ceftazidime resistance in our earlier described outbreak, was lacking in most of the isolates examined. Among the remaining ceftazidime-resistant K. pneumoniae, TEM-26 was also absent, and new enzymes that hydrolyze ceftazidime were detected. Loss of ceftazidime hydrolyzing beta-lactamases was observed after in vitro passage of ceftazidime resistant K. pneumoniae on antibiotic-free media. These findings suggest that class restriction of cephalosporins may increase susceptibility among extended spectrum beta-lactamase-producing pathogens. PMID- 11272259 TI - Streptogramin resistance and shared pulsed-field gel electrophoresis patterns in vanA-containing Enterococcus faecium and Enterococcus hirae isolated from humans and animals in Spain. AB - The present study was performed to determine if any of the 45 vanA-containing Enterococcus faecium or 18 vanA-containing E. hirae strains were shared by chickens (32 E. faecium/l7 E. hirae) and humans (13 E. faecium/1 E. hirae) using pulsed field gel electrophoresis (PFGE) and to study quinupristin-dalfopristin (Q D) resistance. Seven of the 45 E. faecium isolates (from 2 outpatients and from 5 poultry products) were resistant to Q-D (MIC > or = 16 microg/ml); one strain was shown to have satA by PCR and sequencing and, in the other six isolates, the recently described satG gene was demonstrated. Six different PFGE patterns were detected among the 7 Q-D E. faecium-resistant isolates. None of the E. hirae isolates showed Q-D resistance. Among the 45 vanA -containing E. faecium strains, 25 unrelated clones were found by PFGE with highly diverse patterns and an indistinguishable PFGE pattern was observed in vanA-containing E. faecium strains from two humans and two poultry products. A single PFGE pattern was detected in 17 of 18 vanA-containing E. hirae isolates, obtained from one human and 16 chicken samples. Based on the presence of indistinguishable PFGE patterns among VR E. faecium and E. hirae from humans and chickens, we conclude that horizontal transfer of these strains could occur between both groups. PMID- 11272260 TI - Detection and characterization of vancomycin-resistant enterococci in farm animals and raw meat products in Italy. AB - The emergence of vancomycin-resistant enterococci (VRE) in Europe has been ascribed to the long-time use of the glycopeptide antibiotic avoparcin as feed additive in food animals, until its ban in 1997 in EU. The pres- ence of VRE in food of animal origin is believed to represent a potential risk for the consumer. We studied the fecal carriage of VRE in broiler chickens and slaughter pigs in Italy before the avoparcin ban and eval- uated the impact of avoparcin withdrawal on the presence of VRE in raw meat products. Broilers and pigs were both found to be frequently colonized by VRE, as 36% and 24.6% of the flocks or the herds, respec- tively, were positive. Molecular typing of VRE strains by PFGE showed that animals housed in different pens within the same farm were colonized by clonally related strains. After the avoparcin ban, a decrease in the rate of VRE contamination in meat products was observed. Such a decrease was statistically significant in poultry (from 18.8% to 9.6%) but not in pork products (from 9.7% to 6.9%). The majority of VRE from all sources carried the vanA resistance gene and included Enterococcus faecium, E. faecalis, E. hirae, E. durans, and E. gallinarum. None of the strains carried the vanB gene, whereas constitutively resistant vanC-positive strains were frequently found. Our results show that avoparcin withdrawal has been successful in reducing VRE contamination in meat products. However, this measure needs to be complemented by a prudent use of glycopeptide antibiotics in human medicine. PMID- 11272261 TI - Antimicrobial drug resistance in non-typhoidal salmonellas from humans in England and Wales in 1999: decrease in multiple resistance in Salmonella enterica serotypes Typhimurium, Virchow, and Hadar. AB - In 1999 the incidence of multiple drug resistance (to four or more antimicrobials) in non-typhoidal salmonellas from humans in England and Wales fell in isolations of Salmonella enterica serotypes Typhimurium, Virchow, and Hadar. This fall has been most noticeable in S. Typhimurium, where 59% of isolates were multiresistant compared to 81% in 1996. The main reason for this has been a 75% decline in isolations of multiply-resistant S. Typhimurium definitive phage type (DT) 104 (MR DT104) since 1996. Nevertheless MR DT104 remains second to S. Enteritidis phage type 4 as the most common strain in cases of human salmonellosis in England and Wales. Multiple resistance has also remained high in S. Hadar, with 49% of isolates resistant to four drugs or more compared to 56% in 1996. Isolates with decreased sensitivity to ciprofloxacin (minimal inhibitory concentration: 0.25-1.0 mg/L) have increased in incidence in S. Enteritidis, S. Virchow, and S. Hadar; in S. Hadar 70% of isolates were resistant to ciprofloxacin at this level. It is hoped that Codes of Practice introduced by some pharmaceutical companies, governments, professional organisations, and others to combat the unnecessary prophylactic use of fluoroquinolones in animal husbandry will not result in a reduction in the incidence of resistance to ciprofloxacin in salmonella organisms causing infections in humans. PMID- 11272262 TI - Effectiveness of a vancomycin restriction policy in changing the prescribing patterns of house staff. AB - After noting a rise in vancomycin-resistant enterococci (VRE) infections, we initiated a program to decrease inappropriate vancomycin use that focused on improvement of house staff prescribing practices. The initial intervention in June, 1995, encouraging house staff to follow hospital guidelines for vancomycin use and eliciting support from service chiefs in this effort, had little impact. A more intensive educational intervention, beginning in January, 1996, involved concurrent review of all vancomycin orders and one-on-one discussion with the house staff regarding the rationale for the order by an infectious diseases clinical pharmacist. When usage was deemed inappropriate, the pharmacist asked that vancomycin be discontinued, but no automatic stop orders were issued. During the next two and one-half years, this second intervention proved effective at decreasing inappropriate use from 39% to 16.8% +/- 2.4% (p = 0.005). This change was primarily due to a decrease in appropriate vancomycin prophylaxis by cardiothoracic surgery. VRE infections decreased from 0.29/100 patients discharged prior to initiating the program to 0.13/100 patients discharged after the second intervention (p = 0.01). This educational program, although labor intensive, preserved house staff decision-making skills related to antibiotic prescribing at the same time that it decreased inappropriate vancomycin use. PMID- 11272263 TI - Reaping a biotech blunder. PMID- 11272264 TI - So you want a girl? PMID- 11272265 TI - Post-genome, Celera now shoots for profits. PMID- 11272266 TI - New mad cow hideout: the medicine chest. PMID- 11272267 TI - Proteinase 3, Wegener's autoantigen: from gene to antigen. AB - Proteinase 3 (PR3) is one of four serine protease homologues in the azurophilic granules of neutrophils and granules of monocytes. It is of importance that anti neutrophil cytoplasmic antibodies (ANCA) in patients with Wegener's granulomatosis (WG) are mainly directed against PR3 only. Furthermore, PR3 is overexpressed in a variety of acute and chronic myeloid leukemia cells. Cytotoxic T lymphocytes specific for a PR3-derived peptide have been shown to specifically lyse leukemia cells that overexpress PR3. This review will focus on PR3 and the characteristics of PR3 that might implicate this particular antigen in the pathogenesis of WG and as target for immunotherapy in myeloid leukemias. We will discuss the genetic localization and gene regulation of PR3, the processing, storage, and expression of the PR3 protein, and the physiological functions of PR3, and compare this with the three other neutrophil-derived serine proteases: human leukocyte elastase, cathepsin G, and azurocidin. Three main differences are described between PR3 and the other serine proteases. This makes PR3 a very intriguing protein with a large array of physiological functions, some of which may play a role in ANCA-associated vasculitidis and myeloid leukemia. PMID- 11272268 TI - Chronic granulomatous disease: more than the lack of superoxide? AB - Chronic granulomatous disease (CGD) is an inherited disease characterized by severe and recurrent bacterial and fungal infections manifested in most cases in early childhood. Phagocytic cells of CGD patients are unable to produce superoxide anions, and their efficiency in bacterial killing is significantly impaired. Recent work has shown alterations in the electrophysiological properties of CGD granulocytes, which might contribute to the pathogenesis of the disease. The new aspects that we discuss in this review concern the proton channel function of gp91phox (the electron-transporting subunit of the NADPH oxidase) and the electrogenic activity of the active enzyme complex, which can affect the transmembrane trafficking of several ions. Based on the reviewed data, we also propose a hypothesis that the absence of a functional NADPH oxidase in CGD neutrophils could result in altered ion compositions within intracellular and intraphagosomal spaces during the process of phagocytosis. PMID- 11272269 TI - More than destructive: neutrophil-derived serine proteases in cytokine bioactivity control. AB - In addition to the mechanisms inducing the expression and secretion of cytokines under distinct pathophysiological conditions, the fate of cytokines after secretion at sites of inflammation is a field of growing interest. Proteolysis has been suggested to be a fundamental mechanism of regulating the activities of various components of the cytokine network. Evidence grows that besides highly specific cytokine converting proteases such as interleukin-1beta-converting enzyme or tumor necrosis factor-converting enzyme, neutrophil-derived serine proteases are intimately involved in the modulation of the activities of cytokines and their receptors. Particularly at sites of inflammation, high amounts of the active serine proteases elastase, cathepsin G, and proteinase 3 are released from infiltrating polymorphonuclear cells in close temporal correlation to elevated levels of inflammatory cytokines, strongly indicating that these proteases are involved in the control of cytokine bioactivity and availability. PMID- 11272270 TI - Activation-induced expression of CD1d antigen on mature T cells. AB - In the present study, we investigated the expression of human CD1d antigen on activated mature T cells. Expression of this glycoprotein was found to be highly regulated and dependent on PHA stimulation. Flow cytometry studies using the NOR3.2 antibody, which recognized CD1d under denaturing conditions, showed a clear increase in its expression after PHA stimulation. Expression of this molecule after PHA activation was confirmed by analysis of its corresponding transcript by RT-PCR. A single band representing mRNA for CD1d membrane isoform was observed in activated PBMC as well as in ER3 CD1D-transfected and MOLT-4, pre T cell lines, which were used as controls. Western blot analysis revealed an activation-dependent increase in CD1d protein expression when PBMC and enriched T cells were activated for different time periods. Activation-dependent expression of CD1d antigen was also confirmed in allogenic-activated T cells, suggesting that this event could have biological significance. Finally, immunocytochemical studies showed the presence of this protein at the plasma membrane accompanied by a cytoplasmic and perinuclear distribution. Results presented herein provide the first experimental evidence showing that CD1d antigen is present on circulating, activated T lymphocytes, suggesting that its expression is dependent on the activation state of the cells. Elucidation of the molecular mechanisms implicated in the activation-dependent expression of this nonclassical antigen will provide new insights into the understanding of antigen presentation and immune regulation. PMID- 11272271 TI - Antibodies recognizing CD24 LAP epitope on human T cells enhance CD28 and IL-2 T cell proliferation. AB - Membrane expression of the CD24 molecule on activated T lymphocytes is not elucidated fully. We previously described the intracellular and cell-surface expression of the CD24 sialic acid-dependent epitope(s) on phytohemagglutinin activated peripheral blood mononuclear cells. However, the CD24 core protein was not detected previously on human T cells. This study reinvestigated the expression and role of CD24 in T cell subsets. We analyzed binding of anti-CD24 monoclonal antibodies (mAbs) to sialic and leucine-alanine-proline (LAP) epitopes in resting and activated, normal T lymphocytes. CD24 LAP and CD24 sialic epitopes were detected on activated CD4- and CD8-positive cells. Although expression of CD24 sialic epitopes remained stably expressed in interleukin (IL)-2-dependent cultures, T cell expression of the LAP epitope was transient. Anti-LAP antibodies strongly enhanced the response of T cells to a combination of anti-CD3/CD28 mAbs and enhanced proliferative response induced by recombinant IL-2. We found similarities in the tissue distribution and function of the human CD24 LAP molecule and the murine, heat-stable antigen, which suggests that CD24 might function as a signaling molecule on human T cells. PMID- 11272272 TI - Characteristics of the free cytosolic calcium timelag following IgE-mediated stimulation of human basophils: significance for the nonreleasing basophil phenotype. AB - These studies examine characteristics of the quiescent period (timelag) of the free cytosolic calcium ([Ca++]i) elevation that follows stimulation of human basophils through the IgE receptor. Previous studies established that the [Ca++]i timelag was sensitive to the rate of ligand binding, but little else is known about this response characteristic. The [Ca++]i timelag could be lengthened using antigenic stimulation that is rapid but only weakly induces secretion: tenfold differences in the "strength" of the stimulus, as assessed by histamine release, are associated with threefold differences in the timelag. Inhibiting p53/56lyn kinase with low concentrations of the specific inhibitor, PP1, lengthened the [Ca++]i timelag dramatically. PP1 was also found to delay the onset of syk phosphorylation and histamine release. Staurosporine and genistein, which are known to inhibit early tyrosine kinases, had, at best, only modest effects on the [Ca++]i timelag. Specific inhibitors of protein kinase C (PKC) had no effect on the [Ca++]i timelag, and direct activation of PKC with PMA had only very modest effects on the timelag. Contrary to expectations, basophils with the so-called nonreleasing phenotype demonstrated an IgE-mediated [Ca++]i response at the single-cell level. However, the length of [Ca++]i timelag in nonreleasing basophils was threefold longer than normally found in releasing basophils. Furthermore, the [Ca++]i response was significantly more asynchronous than in releasing basophils and lacking in a sustained [Ca++]i elevation. These studies indicate that the [Ca++]i timelag following stimulation through the IgE receptor is sensitive to inhibition of lyn kinase but not other agents that have been demonstrated to inhibit early tyrosine kinases previously. However, only one characteristic of the [Ca++]i response phenotype of nonreleasing basophils--the [Ca++]i timelag but not the absence of a sustained [Ca++]i elevation--could be mimicked by inhibition of lyn kinase with PP1. PMID- 11272273 TI - Differential effects of anti-Fc gamma RIIIb autoantibodies on polymorphonuclear neutrophil apoptosis and function. AB - Anti-Fc gamma receptor IIIb (Fc gammaRIIIb) human autoantibodies (Ab) have been classified previously into three groups, based on the results of an indirect immunofluorescence (IIF) test and an enzyme-linked immunosorbent assay (ELISA): IIF+/ELISA+ (group A), IIF+/ELISA- (group B), and IIF-/ELISA+ (group C) sera. In this study, differential effects between IIF+ autoAb, recognizing cell-bound Fc gammaR, and those ELISA+, recognizing only cell-free Fc gammaR, were studied on polymorphonuclear neutrophils (PMN). Neither group A nor B autoAb was cytotoxic, although both prolonged the survival of PMN by delaying spontaneous apoptosis. By the same extent, the PMN-binding antisera stimulated the appearance of a CD11b(dim) population, following a 12-h incubation. This event was associated with a lowered expression of beta2 integrin molecules, resulting in altered PMN function. Treatment with groups A and B autoAb reduced adhesiveness and respiratory burst. This impairment of the responses was more pronounced when the cells originated from donors NA1+ NA1+ rather than donors NA2+ NA2+. From our observations, the influences of anti-Fc gammaRIIIb autoAb on PMN survival, as well as function and subsequent dysregulation of the inflammatory response, have proven somewhat dependent on their target antigens, as determined by IIF coupled with ELISA and Fc gammaRIIIb polymorphism. PMID- 11272275 TI - Increase of CCR1 and CCR5 expression and enhanced functional response to MIP-1 alpha during differentiation of human monocytes to macrophages. AB - Chemokines and their receptors regulate migration of leukocytes under normal and inflammatory conditions. In this study, we analyzed the CC chemokine receptor (CCR) expression of monocytes differentiating in vitro to macrophages. We observed a time-dependent change of expression and functional responsiveness of CCR1, CCR2, and CCR5 within 48 h. Whereas freshly harvested monocytes were strongly attracted by monocyte chemotactic protein 1 (MCP-1), a specific ligand for CCR2, only a weak response was observed to macrophage inflammatory protein 1alpha (MIP-1alpha), which binds to CCR1 and CCR5. In striking contrast, differentiated macrophages displayed a strong chemotactic response to MIP-1alpha and only a weak response to MCP-1. These findings were paralleled by intracellular calcium shifts. During the time course of monocyte to macrophage differentiation, mRNA levels and surface expression of CCR2 decreased, whereas that of CCR1 and CCR5 increased. The time-dependent switch from CCR2 on monocytes to CCR1 and CCR5 on mature macrophages reflects a functional change belonging to the differentiation process of monocytes to macrophages and may form the basis for a differential responsiveness of monocytes and macrophages to distinct sets of chemokines. PMID- 11272274 TI - Inhibitory effect of serine protease inhibitors on neutrophil-mediated endothelial cell injury. AB - To investigate the inhibitory effect of serine protease inhibitors (SPI) on neutrophil-mediated endothelial cell (EC) injury, we analyzed the in vitro cytotoxicity of radiolabeled human umbilical vein EC (HUVEC) mediated by neutrophils in the presence of SPI. The EC injury was inhibited dose-dependently by urinary trypsin inhibitor (ulinastatin, UTI) and ONO-5046, which have the ability to inactivate neutrophil elastase, but not by gabexate mesilate, nafamostat mesilate, aprotinin, and argatroban, which have no ability to inactivate neutrophil elastase. In addition, when UTI and ONO-5046 were added to the tumor necrosis factor alpha-primed neutrophils alone, they showed a dose dependent inhibition of the intracellular elastase activity, but the other SPI did not, for either flow cytometry or confocal microscopy. Therefore, UTI and ONO 5046 may protect EC against the neutrophil-mediated injury not only by inactivating the extracellular elastase secreted by neutrophils, but also by acting directly on neutrophils and suppressing the production and secretion of activated elastase from them. PMID- 11272276 TI - CpG-containing oligodeoxynucleotides induce TNF-alpha and IL-6 production but not degranulation from murine bone marrow-derived mast cells. AB - Mast cells are sentinel cells critical to the initiation of innate immune and inflammatory responses, particularly at mucosal surfaces. To fulfill this function they can be activated by several pathogen-associated stimuli to produce cytokines with or without concurrent degranulation. We examined the ability of immunostimulatory DNA sequences including CpG motifs, which are found in increased quantities in bacterial DNA, to activate mouse bone marrow-derived mast cells (mBMMC). Mast cells were treated with a range of doses of CpG-containing oligodeoxynucleotides or control oligodeoxynucleotides without CpG within their sequence. There was a dose-dependent increase in the production of both interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-alpha) by mast cells treated with the CpG-containing oligodeoxynucleotides. The cytokine levels induced were directly related to the number of CpG within a given length of sequence. Treatment with oligonucleotides containing 3CpG induced an eightfold increase in TNF production over control incubated mast cells. Other cytokines, including granulocyte-macrophage colony-stimulating factor, IL-4, interferon gamma, and IL-12 were not induced by oligonucleotide treatment. Neither CpG containing oligodeoxynucleotides nor control oligodeoxynucleotides induced degranulation of mast cells. Bacterial DNA from Escherichia coli also induced IL 6 from mBMMC but neither calf thymus DNA nor methylase-treated E. coli DNA had such an effect. Examination of the uptake of Texas red-labeled CpG and non-CpG containing oligodeoxynucleotides revealed that they were both similarly taken up by the mBMMC. These results have important implications for the mechanism by which mast cells respond to bacteria and for the potential role of mast cells in DNA vaccination. PMID- 11272277 TI - Stromal derived factor-1 alpha (SDF-1 alpha) induces CD4+ T cell apoptosis via the functional up-regulation of the Fas (CD95)/Fas ligand (CD95L) pathway. AB - Stromal-derived factor-1alpha (SDF-1alpha), the high-affinity ligand of CXC chemokine receptor 4 (CXCR4), induced a progressive increase of apoptosis when added to the Jurkat CD4+/CXCR4+ T cell line. The SDF-1alpha-mediated Jurkat cell apoptosis was observed in serum-free or serum-containing cultures, peaked at SDF 1alpha concentrations of 10-100 ng/ml, required 3 days to take place, and was completely blocked by the z-VAD-fmk tripeptide caspase inhibitor. Although SDF 1alpha did not modify the expression of TNF-alpha or that of TNF-RI and TNF-RII, it increased the expression of surface Fas/APO-1 (CD95) and intracellular Fas ligand (CD95L) significantly. Moreover, the ability of SDF-1alpha to induce apoptosis was inhibited by an anti-CD95 Fab' neutralizing antibody. These findings suggest a role for SDF-1alpha in the homeostatic control of CD4+ T-cell survival/apoptosis mediated by the CD95-CD95L pathway. PMID- 11272278 TI - Differences in the induction of CD8+ T cell responses by subpopulations of dendritic cells from afferent lymph are related to IL-1 alpha secretion. AB - The major subset of dendritic cells (DC) from bovine afferent lymph expresses the SIRP alpha MyD-1 antigen, but not CD11a or the antigen recognized by mAb CC81, and potently stimulates CD4+ and CD8+ T lymphocyte proliferation. The minor subpopulation, that is CD11a+ CC81+ MyD-1-, effectively stimulates CD4+ but not CD8+ T lymphocyte proliferation. CD11a+ CC81+ MyD-1- DC did not induce anergy or death or secrete an inhibitory factor. However, supernatant from cultures of CD8+ T cells with CD11a- CC81- MyD-1+ DC significantly enhanced proliferation of CD8+ T cells in response to CD11a+ CC81+ MyD-1- DC, an effect that was blocked by interleukin (IL)-1alpha, but not IL-1beta, specific mAb. The proliferation of CD8+ T cells with CD11a+ CC81+ MyD-1- DC was also enhanced by adding IL-1alpha. IL-1beta slightly enhanced proliferation, whereas IL-2, IL-6, IL-12, and IL-15 had no effect. We conclude that the failure to stimulate CD8+ T cell proliferation results from the lack of IL-1alpha synthesis by this population, which may have important consequences in vivo. PMID- 11272279 TI - Involvement of protein kinases in the potentiation of lipopolysaccharide-induced inflammatory mediator formation by thapsigargin in peritoneal macrophages. AB - We have explored the regulatory roles played by Ca2+-dependent signaling on lipopolysaccharide (LPS)-induced nitric oxide (NO), prostaglandin E2 (PGE2), tumor necrosis factor alpha (TNF-alpha), and interleukin-6 (IL-6) release in mouse peritoneal macrophages. To elevate intracellular Ca2+, we used thapsigargin (TG) and UTP. Although LPS alone cannot stimulate NO synthesis, co-addition with TG, which sustainably increased [Ca2+]i, resulted in NO release. UTP, via acting on P2Y6 receptors, can stimulate phosphoinositide (PI) turnover and transient [Ca2+]i increase, however, it did not possess the NO priming effect. LPS alone triggered the release of PGE2, TNF-alpha, and IL-6; all of which were potentiated by the presence of TG, but not of UTP. The stimulatory effect of LPS plus TG on NO release was inhibited by the presence of Ro 31-8220, Go6976, KN-93, PD 098059, or SB 203580, and abolished by BAPTA/AM and nuclear factor kappaB (NF-kappaB) inhibitor, PDTC. PGE2, TNF-alpha, and IL-6 release by LPS alone were attenuated by Ro 31-8220, Go6976, PD 098059, SB 203580, and PDTC. Using L-NAME, soluble TNF alpha receptor, IL-6 antibody, NS-398, and indomethacin, we performed experiments to understand the cross-regulation by the four mediators. The results revealed that TNF-alpha up-regulated NO, PGE2, and IL-6 synthesis; PGE2 up-regulated NO, but down-regulated TNF-alpha synthesis; and PGE2 and IL-6 mutually up-regulated reciprocally. Taken together, murine peritoneal macrophages required a sustained [Ca2+]i increase, which proceeds after TG, but not UTP, stimulation, to enhance LPS-mediated release of inflammatory mediators, particularly for NO induction. Activation of PKC-, ERK-, and p38 MAPK-dependent signaling also are essential for LPS action. The positive regulatory interactions among these mediators might amplify the inflammatory response caused by endotoxin. PMID- 11272280 TI - Colostral neutrophils express Fc alpha receptors (CD89) lacking gamma chain association and mediate noninflammatory properties of secretory IgA. AB - Colostrum plays an important role in protecting newborn infants against acute gastrointestinal and respiratory infections. IgA antibodies have been considered the major effector component; however, the role of their receptors on colostral phagocytes, especially neutrophils, has not been studied. Here, we demonstrate that CD15+ colostrum neutrophils express IgA Fc receptors (Fc alphaR, CD89) at levels similar to those of blood neutrophils. Most colostral cells (70%) bear secretory IgA (SIgA) on their surface (and intracellularly), whereas blood cells do not. The Fc alphaR on colostral neutrophils was identified as the a.1 isoform with a similar molecular mass (55-75 kDa) as that identified for blood neutrophils. Removal of N-linked carbohydrates revealed a major protein core of 32 kDa for both cell types. In contrast, co-immunoprecipitation and immunoblot experiments using a mild detergent, digitonin, revealed a lack of gamma chain association with Fc alphaR (gamma-less) exclusively on colostral neutrophils. The functional role of these gamma-less Fc alphaR cells was evaluated by measuring superoxide release and killing of SIgA-coated enteropathogenic E. coli. No increase in superoxide release was observed in colostral cells compared with blood neutrophils, whereas optimal release was obtained with PMA stimulation. Furthermore, despite similar bacterial phagocytosis index between both cell types, IgA-mediated bacterial-killing was not detectable with colostral neutrophils, whereas killing was detectable on blood cells. These results reveal exclusive expression of gamma-less Fc alphaR on colostral neutrophils associated with receptor hyperoccupation by IgA and with low, bacterial-killing activity, which suggest that this receptor may mediate noninflammatory effects of SIgA. PMID- 11272281 TI - Cloning, mRNA distribution, and functional expression of an avian counterpart of the chemokine receptor/HIV coreceptor CXCR4. AB - The chemokine signaling system, which coordinates the basal and emergency trafficking of leukocytes, presumably coevolved with the hematopoietic system. To study its phylogenetic origins, we used the open reading frame (ORF) of the human chemokine receptor CXCR4 as a genomic probe, since in mammals it is the most highly conserved chemokine receptor known. CXCR4 cross-hybridized to genomic DNA from mouse and chicken, but not zebrafish, Drosophila, or Caenorhabditis elegans. Accordingly, we cloned the corresponding chicken cDNA. The ORF is 359 codons long versus 352 for human CXCR4, and encodes a protein 82% identical to human CXCR4. In a calcium flux assay of receptor function, CHO-K1 cells stably transfected with the chicken cDNA responded specifically to human SDF-1, the specific ligand for CXCR4, but not to a panel of other chemokines tested at 100 nM. SDF-1 activated the cells in a dose-dependent manner (EC50 approximately 5 nM), whereas parental CHO-K1 cells did not respond. The CHO-K1 cell transfectants also bound 125I-SDF-1 specifically. Leukocytes from chicken peripheral blood expressed chCXCR4 mRNA and responded to human SDF-1 in a calcium flux assay with an EC50 similar to that for chCXCR4-transfected CHO cells, suggesting that this response is mediated by native chCXCR4. Analysis of chicken genomic DNA with the chicken cDNA as probe revealed a pattern consistent with a single copy gene, and the absence of any closely related genes. mRNA was detected in brain, bursa, liver, small and large intestine, embryonal fibroblasts, and blood leukocytes, but not in stomach or pancreas. These results, which identify the first functional non viral, non-mammalian chemokine receptor, suggest that the origins of a functional chemokine system extend at least to birds and suggest that, as in mammals, CXCR4 functions in many avian tissues. PMID- 11272282 TI - Phase Ia study of a hypoxic cell sensitizer doranidazole (PR-350) in combination with conventional radiotherapy. AB - A phase Ia study of a 2-nitroimidazole nucleoside analog radiosensitizer doranidazole was conducted to evaluate its toxicity and pharmacokinetics in patients undergoing conventional external beam radiotherapy. Twenty-nine patients, aged 40-74 years, with a WHO performance status of 0-2 and with adequate organ functions, were entered in the study. Single administration of doranidazole was investigated first with 13 patients and then a course of five consecutive daily administrations was tested in 16 patients. Doranidazole was given i.v. 25 min before irradiation. Doranidazole doses of 400, 800, 1300 and 2000 mg/m2 were evaluated in the former study, and daily doses of 800, 1300 and 2000 mg/m2 were investigated in the latter study. All patients tolerated doranidazole administration. Although a transient decrease in the 24-h creatinine clearance rate was observed in five patients (one in the single administration study and four in the repeat administration study), this was not considered to be the dose-limiting toxicity. Other toxicities (hematological and gastrointestinal), which may not be related to doranidazole administration, were also mild and were not dose limiting. No neurotoxicity was observed. The average maximum concentration, area under the time-concentration curve and half-life of doranidazole in serum were 172-194 microg/ml, 502-582 microg x h/l and 4.2-4.6 h, respectively, at 2000 mg/m2. At the tested doses, administration of doranidazole was tolerable and achieved serum concentrations at which reasonable radiosensitization could be expected. A phase Ib/II study to evaluate the feasibility and efficacy of up to 30 repeat administrations seems to be warranted. PMID- 11272283 TI - Approach to a multiparametric sensor-chip-based tumor chemosensitivity assay. AB - Although not widely practiced by oncologists, in vitro tumor chemosensitivity assays (TCA) have proved to increase the lifetime of tumor patients in prospective clinical trials. By individualizing cancer therapy, they can support the clinician's decision which is usually based on empirically retrieved data and thereby prevent inadequate chemotherapy. We present the first results of a new sensor-chip-based technology which might be useful for a multiparametric TCA. In particular, the aspect of dynamic on-line data generation on intact cellular specimens is a major difference to alternative assays. A series of experiments has been performed on cell lines and human tumor explants. Cell cultures and tumor tissue explants were placed on miniaturized silicon and glass sensor chips. The sensor data currently analyze metabolic profiles (rates of extracellular acidification and cellular oxygen consumption) and changes in cell morphology (monitoring of electric impedance). With the cell lines, drug-associated cellular signals have been detected with all three parameters, while primary explants so far caused metabolic responses only. In particular, cellular respiration or mitochondrial activity seems to be a most sensitive indicator of acute cytotoxic effects. The experimental results were achieved using different test versions. Besides giving a status report, the theoretical potential and current problems of sensor chip technology in TCA is discussed. PMID- 11272284 TI - A hollow fiber model for in vitro studies of cytotoxic compounds: activity of the cyanoguanidine CHS 828. AB - The hollow fiber assay is currently used as an in vivo model for anticancer drug screening in nude mice, but it can also be used as an in vitro model. In the current study, an in vitro hollow fiber model was used to study the effect and mode of induced cell death of a new cyanoguanidine, CHS 828. Human leukemia, adenocarcinoma and lymphoma cell lines as well as primary cultures of human tumor cells from patients with chronic lymphocytic leukemia (CLL) and ovarian cancer (OC) and normal human lymphocytes were cultured in semipermeable hollow fibers. The fibers were incubated for 3 or 14 days prior to CHS 828 exposure for 72 h, followed by determination of living cell density by MTT staining. For cell morphology, using harvested cultures on cytospin slides had technical advantages compared to using paraffin sections of the formalin-fixed fibers. CHS 828 showed higher antitumor activity on CLL and normal human lymphocyte cultures compared to OC cultures, and cell lines cultured 3 days were more sensitive than those cultured 14 days. Morphological examination of CHS 828-treated cultures revealed a mixture of apoptosis and necrosis. PMID- 11272285 TI - Inhibition of mitotic cyclin B and cdc2 kinase activity by selenomethionine in synchronized colon cancer cells. AB - Selenomethionine (SeMet), an organic selenium compound, has been demonstrated to have significant chemopreventive activity. However, the mechanism of action of SeMet has yet to be identified. Previously, our laboratory found that treatment of cells with SeMet induced apoptosis and altered the cell cycle. These observations have led to further analysis of the cell cycle effects of SeMet in colon cancer cells. Synchronized HCT 116 colon cancer cells treated with 100 microM SeMet for 66 h were found to have a transient delay in G2/M phase of the cell cycle at 18 and 24 h after treatment. With this was observed an inhibition of cell growth. Coincidentally with this delay was a decrease in mitotic cyclin B RNA expression at 18 h after treatment. In addition, the cdc2 kinase activity of HCT 116 cells was decreased at 18 h. Morphological studies indicate an increase in the number of treated cells (45%) undergoing apoptosis at 66 h compared to control cells (27%). These studies demonstrate that modulation of mitotic cyclin expression and cdc2 kinase activity play a role in the ability of SeMet to inhibit tumor cell growth. A consequence of this prolonged arrest is apoptosis. PMID- 11272286 TI - Induction of apoptosis in human leukemia K-562 and gastric carcinoma SGC-7901 cells by salvicine, a novel anticancer compound. AB - Salvicine (a novel diterpenoid quinone compound) exhibited a marked antitumor activity on human solid tumor cell lines and BALB/c-nu human carcinoma xenografts in our earlier studies, and it has been chosen as a candidate anticarcinogenic compound in the preclinical research stage. The present study was undertaken in order to observe whether or not the antitumor effect of salvicine is associated with its ability to induce apoptosis. Our results show that salvicine is capable of inhibiting cell proliferation and inducing characteristic changes of apoptosis in both human leukemia K-562 and gastric carcinoma SGC-7901 cells. These effects are dose and time dependent. The results of this study strongly suggest that the antitumor effect of salvicine is associated with its ability to induce apoptosis. Meanwhile, this study also shows that the activity of salvicine against K-562 and SGC-7901 cells is similar with regards to both growth inhibition and apoptosis induction, further indicating that salvicine causes these particular effects on solid tumor cells. PMID- 11272287 TI - Evaluation of combretastatin A-4 prodrug in a non-Hodgkin's lymphoma xenograft model: preclinical efficacy. AB - Combretastatin A-4 prodrug (CA4P) is a new antitubulin agent currently in phase I/II clinical trials against solid tumors. We have previously reported on the in vitro activity of CA4P against a panel of malignant human B-lymphoid cell lines. In this study, we investigated the antitumor and the antiangiogenic activity of CA4P in our diffuse large cell lymphoma WSU-DLCL2-SCID mouse model. WSU-DLCL2 cells (10(7)) were injected s.c. into 5-week-old female ICR-SCID mice. Tumor bearing mice were treated at the CA4P maximum tolerated dose (MTD) of 800 mg/kg in different dose/schedules. CA4P showed significant antitumor activity against this lymphoma model. Best results were seen when MTD was given in two and four divided doses (400 and 200 mg/kg, respectively). CA4P given in four divided doses (4 x 200 mg/kg) showed a log10 kill of 1.01, T/C of 11.7% and T-C of 12 days. Immunohistochemical staining using anti-CD31 antibody after 6, 24, 48 and 120 h treatment revealed a significant decrease in the number of tumor blood vessels after 24 h (about 80%). Only the periphery of treated tumors revealed the presence of blood vessels. Morphological examination of the tumors after tetrachrome staining showed a necrotic center in tumors of CA4P-treated animals. New blood vessel formation was noted to emerge in tumor tissues as early as 48 h following a single dose of CA4P. The G2/M arrest observed in vitro was not detected in vivo indicating predominance of the antiangiogenic effects with regard to antitumor efficacy in vivo. We conclude that CA4P has antiangiogenic activity in this lymphoma model and the use of this agent should be explored clinically in the treatment of non-Hodgkin's lymphoma. PMID- 11272288 TI - Platinum(II) and palladium(II) complexes with 2-acetylpyridine thiosemicarbazone: cytogenetic and antineoplastic effects. AB - The effect of three novel complexes of Pt(II) and three complexes of Pd(II) with 2-acetylpyridine thiosemicarbazone (HAcTsc) on sister chromatid exchange (SCE) rates and human lymphocyte proliferation kinetics on a molar basis was studied. Also, the effect of Pt(II) and Pd(II) complexes against leukemia P388 was investigated. Among these compounds, the most effective in inducing antitumor and cytogenetic effects were the complexes [Pt(AcTsc)2] x H2O and [Pd(AcTsc)2] while the rest, i.e. (HAcTsc), [Pt(AcTsc)Cl], [Pt(HAcTsc)2]Cl2 x 2H2O, [Pd(AcTsc)Cl] and [Pd(HAcTsc)2]Cl2, displayed marginal cytogenetic and antitumor effects. PMID- 11272289 TI - Interferon-alpha-induced pure red cell aplasia following chronic myelogenous leukemia. AB - We report a case of pure red cell aplasia (PRCA) that resulted from interferon (IFN)-alpha therapy for chronic myelogenous leukemia. PRCA improved within 1 month after IFN-alpha was discontinued. This case indicates the involvement of IFN-alpha in the pathogenesis of PRCA. PMID- 11272290 TI - In vitro targeting of a cytotoxic analog of luteinizing hormone-releasing hormone AN-207 to ES-2 human ovarian cancer cells as demonstrated by microsatellite analyses. AB - Targeting of cytotoxic agents represents a modern approach to the treatment of various cancers, that improves the efficacy and reduces peripheral toxicity. Recently we developed a powerful cytotoxic analog of luteinizing hormone releasing hormone (LHRH), AN-207, designed to be targeted to tumors that express LHRH receptors. This analog consists of the superactive derivative of doxorubicin (DOX), 2-pyrrolino-DOX (AN-201), linked to [D-Lys6]LHRH carrier. In the present study we investigated the cytocidal effects of AN-207 and AN-201 on the LHRH receptor-positive ES-2 ovarian cancer cells. The targeting of AN-207 to ES-2 cells in the presence of LHRH receptor-negative UCI-107 ovarian cancer cells was also evaluated by semi-quantitative polymerase chain reaction (PCR) amplification of microsatellite markers. Ligand competition assays showed a single class of high-affinity and low-capacity binding sites in ES-2 cells with a mean dissociation constant (KD) of 3.93 +/- 0.1 nM and a mean maximal binding capacity (Bmax) of 271 +/- 26.1 fmol/mg membrane protein. Kinetic assays indicated that AN 207 caused cell death in a concentration- and time-dependent manner in ES-2 cells, but not in UCI-107 cells, while the kinetics of cytotoxic effects of AN 201 were similar in both cell lines. To investigate targeting, ES-2 cells were co cultured with UCI-107 cells, treated with 10 nM AN-207 or AN-201 for different times and then cultured for 48 h in the absence of cytotoxic agents. Genomic DNA was extracted for microsatellite analyses using different markers. Semi quantitative analyses of the intensity of the alleles that correspond to each cell line indicated that AN-207 was selectively targeted to ES-2 cells, while AN 201 showed no selectivity for either cell line. These results extend our previous findings that AN-207 can be targeted to ovarian cancers and other tumors that express receptors for LHRH. Cytotoxic analogs of LHRH, such as AN-207, should be considered for treatment of LHRH receptor-positive tumors. PMID- 11272291 TI - Phosphodiesterase 3 as a potential target for therapy of malignant tumors in the submandibular gland. AB - Phosphodiesterase (PDE) 3s have been characterized in human neoplastic submandibular gland intercalated duct HSG cells. There have been no reports on PDE3 in malignant salivary gland cells. PDE3 activity was detected in homogenates of HSG cells. About 75% of PDE3 activity in HSG cells was recovered in supernatant fractions and 25% in particulate fractions. PDE3A and 3B mRNAs were detected by reverse transcription-polymerase chain reaction in RNA from HSG cells. The nucleotide sequences of the fragments were identical to those of human PDE3A and 3B. The PDE3-specific inhibitor, cilostamide, inhibited the growth of HSG cells. Our results indicate that PDE3s may be important in the growth of HSG cells. PDE3 thus appears to be a potential new target for antiproliferative therapies. PMID- 11272292 TI - The homocamptothecin BN 80915 is a highly potent orally active topoisomerase I poison. AB - BN 80915, a lead compound of the homocamptothecin (hCPT) family, has entered clinical trials. BN 80915 is a difluoro-hCPT where the six-membered alpha hydroxylactone ring of camptothecin (CPT) is replaced by a seven-membered beta hydroxylactone ring. Preclinical data reported here show that in spite of the modification to the crucial E-ring of CPTs, BN 80915 retains topoisomerase I poisoning activity as shown in living HT29 cells as well as in cell-free assays, where BN 80915 always performs better than SN-38 or TPT. In antiproliferative assays BN 80915 is also very potent as evidenced by IC50s values consistently lower than those of SN38 in sensitive cell lines as well as in their related multidrug-resistant lines overexpressing P-glycoprotein or multidrug resistance associated protein. Furthermore, in human plasma, in contrast to CPT analogs, the hydrolysis of BN 80915 is slow, leading to improved plasma stability, and irreversible, thus avoiding toxicity related to the accumulation of active principle during excretion in the urinary tract. These findings may account for the good in vivo efficacy observed in PC3 xenograft experiments where BN 80915 administered orally at very low doses doubled the tumor growth delay in comparison to CPT-11 administered i.p. Altogether, these results strongly support further development of BN 80915. PMID- 11272293 TI - Hope for patients with asplenia or hyposplenism. PMID- 11272294 TI - The science and politics of cancer screening. PMID- 11272295 TI - Fluoxetine and side effects in the geriatric population. PMID- 11272297 TI - Treatment of plantar fasciitis. AB - Plantar fasciitis is a common cause of heel pain in adults. The disorder classically presents with pain that is particularly severe with the first few steps taken in the morning. In general, plantar fasciitis is a self-limited condition. However, symptoms usually resolve more quickly when the interval between the onset of symptoms and the onset of treatment is shorter. Many treatment options exist, including rest, stretching, strengthening, change of shoes, arch supports, orthotics, night splints, anti-inflammatory agents and surgery. Usually, plantar fasciitis can be treated successfully by tailoring treatment to an individual's risk factors and preferences. PMID- 11272296 TI - Treatment of depression in the elderly. PMID- 11272298 TI - Plasma viral load testing in the management of HIV infection. AB - The polymerase chain reaction assay, branched DNA assay and nucleic acid sequence based amplification assay quantitate human immunodeficiency virus (HIV) RNA levels. Plasma viral load (PVL) testing has become a cornerstone of HIV disease management. Initiation of antiretroviral drug therapy is usually recommended when the PVL is 10,000 to 30,000 copies per mL or when CD4+ T-lymphocyte counts are less than 350 to 500 per mm3 (0.35 to 0.50 x 10(9) per L). PVL levels usually show a 1- to 2-log reduction within four to six weeks after therapy is started. The goal is no detectable virus in 16 to 24 weeks. Periodic monitoring of PVL is important to promptly identify treatment failure. When feasible, the same assay should be used for serial PVL testing in the individual patient. At least two PVL measurements usually should be performed before antiretroviral drug therapy is initiated or changed. PVL testing may be helpful in the rare instance of indeterminate HIV antibody testing, especially in a patient with recent infection. PMID- 11272300 TI - Screening for cancer: evaluating the evidence. AB - Many patients expect to undergo screening tests for cancer. In evaluating screening procedures, physicians must take into account the known effects of lead time, length and screening biases, all of which can result in an overestimation of the benefits of screening. The gold standard by which a screening test is evaluated remains the prospective, randomized controlled trial, demonstrating reduced morbidity and mortality. The magnitude of benefit from screening is best expressed in terms of the number of patients needed to screen. This value ranges from approximately 500 to 1,100 for proven screening interventions. These concepts are illustrated by controversies in current screening recommendations for cancers of the cervix, lung, colon, breast and prostate, which together account for more than 50 percent of cancer deaths in the United States. PMID- 11272299 TI - Detection, education and management of the asplenic or hyposplenic patient. AB - Fulminant, potentially life-threatening infection is a major long-term risk after splenectomy or in persons who are functionally hyposplenic as a result of various systemic conditions. Most of these infections are caused by encapsulated organisms such as pneumococci, Haemophilus influenzae and meningococci. A splenectomized patient is also more susceptible to infections with intraerythrocytic organisms such as Babesia microti and those that seldom affect healthy people, such as Capnocytophaga canimorsus. Most patients who have lost their spleens because of trauma are aware of their asplenic condition, but some older patients do not know that they are asplenic. Other patients may have functional hyposplenism secondary to a variety of systemic diseases ranging from celiac disease to hemoglobinopathies. The identification of Howell-Jolly bodies on peripheral blood film is an important clue to the diagnosis of asplenia or hyposplenia. Management of patients with these conditions includes a combination of immunization, antibiotic prophylaxis and patient education. With the increasing prevalence of antibiotic-resistant pneumococci, appropriate use of the pneumococcal vaccine has become especially important. PMID- 11272301 TI - Photo quiz. Pigmented preauricular papules. PMID- 11272303 TI - A "hopeless" patient. PMID- 11272302 TI - AAP develops guidelines for early detection of dislocated hips. PMID- 11272304 TI - Trauma program bolsters case for better equipment with benchmarking study. AB - Gregory Jurkovich, MD, FACS, head of trauma at Harborview Medical Center in Seattle, contended that patients with severe head injuries did better when they had intercranial pressure monitors. But it is an expensive proposition that some might question. So it was nice to get support from national benchmarking data that the best trauma programs did just what his physicians did at Harborview. PMID- 11272305 TI - Are you doing your best for women patients? AB - Report card data show a nation failing to make the grade. When the National Women's Law Center in Washington, DC, released its premier issue of Making the Grade on Women's Health: A National and State-by-State Report Card in August, a lot of policy-makers and state health leaders took notice. But now even health plans are taking a look at the data. PMID- 11272306 TI - Outsourcing satisfaction gives system a boost. AB - One might expect Kristin Baird, RN, MHA, the author of a book about customer service in health care and vice president of business development at the small central Wisconsin health system Watertown Area Health Services, to know the value of measuring patient satisfaction. That assumption is correct. And since the system and its hospital, five clinics, and two senior housing complexes already engage in external benchmarking of financial and quality indicators, you might expect that they did the same with patient satisfaction. But here, you'd be wrong. PMID- 11272307 TI - New technology helps data management. AB - Are the new Web-based and wireless systems for you? The age of technology has many benefits for the health care industry. Information is easier to come by, easier to access. But how do you bring all the pieces together? Two new systems may help health care organizations manage their data and information better in the future. PMID- 11272308 TI - First derivative spectrophotometric, TLC-densitometric, and HPLC determination of acebutolol HCL in presence of its acid-induced degradation product. AB - Three methods are presented for the determination of acebutolol HCl in presence of its acid-induced degradation product. The first method was based on measurement of the first derivative amplitude of acebutolol HCl at 266.6 nm. The second method was based on separation of acebutolol HCl from its acid-induced degradation product followed by densitometric measurement of the spots at 230 nm. The separation was carried out on silica gel 60 F254, using ethanol-glacial acetic acid (4:1, v/v) as mobile phase. Second order polynomial equation was used for the regression line. The third method was based on high performance liquid chromatographic (HPLC) separation of acebutolol HCl from its acid-induced degradation product on a reversed phase, ODS column using a mobile phase of methanol-water (55:45, v/v) with UV detection at 240 nm. The first derivative spectrophotometric method was utilized to investigate the kinetics of the acid degradation process at different temperatures. PMID- 11272309 TI - The quantification of endogenous steroids in bovine aqueous humour and vitreous humour using isotope dilution GC-NCI-MS. AB - Pentafluorobenzyloxime-trimethylsilyl derivatives of androgens, progestogens and corticosteroids were prepared and used for the analysis of these steroids in bovine aqueous humour and vitreous humour by GC-MS method. Appropriate deuteriated isotopomers of the parent steroids were labelled with deuterium via simple synthetic procedure and used as internal standards. The concentration (ng ml(-1), +/- S.E.M.) of these steroids in bovine aqueous humour and vitreous humour were found to be as follow: (1) aqueous humour (n = 17); hydrocortisone (n = 17; 2.40 +/- 0.54), progesterone (n = 15; 0.06 +/- 0.01), 4-androstene-3,17 dione (n = 8; 0.15 +/- 0.07) and testosterone (n = 4; 0.14 +/- 0.04); and (2) bovine vitreous humour (n = 19); hydrocortisone (n = 19; 1.78 +/- 0.25), progesterone (n = 18; 0.09 +/- 0.01), 4-androstene-3,17-dione (n = 19; 0.11 +/- 0.02), 11-deoxycorticosterone (n = 12; 29.27 +/- 6.42), 17alpha hydroxyprogesterone (n = 6; 5.55 +/- 3.12). The concentration of corticosterone, 11-deoxycorticosterone and 17alpha-hydroxyprogesterone and testosterone and corticosterone were below the limit of detection in aqueous humour and vitreous humour, respectively. PMID- 11272310 TI - Development of a capillary electrophoresis assay for the determination of carvedilol enantiomers in serum using cyclodextrins. AB - A capillary electrophoresis method using cyclodextrins as the chiral selectors was developed for the determination of carvedilol enantiomers in serum. Several types of cyclodextrins were evaluated. The effect of cyclodextrin concentration on enantiomer resolution was investigated. Best results were obtained using 10 mM hydroxypropyl-beta-cyclodextrin in the run buffer. The effect of voltage on efficiency was assessed. Other electrophoretic conditions were optimized. The method was validated for carvedilol enantiomers in serum. Linearity of detection was assessed over the concentration range of 50-4000 ng/ml of each enantiomer in serum. Intra- and inter-assay variability obtained were under 8% for both enantiomers. PMID- 11272311 TI - Rapid and simple determination of mycophenolic acid in human plasma by ion-pair RP-LC with fluorescence detection. AB - Mycophenolic acid (MPA) is an immunosuppressive drug given as the prodrug of mycophenolate mofetil (MMF). In order to investigate the pharmacokinetics of MPA, a simple, specific, sensitive and reliable method has been established for the quantitative determination of MPA in plasma from renal transplant recipients. The method involves a single-step protein precipitation procedure and a specific determination by ion-pair reversed-phase high-performance liquid chromatography (HPLC) with fluorescence detection. Separation was achieved on a C18 column (150 x 4.6 mm, 5 microm) with a mobile phase composed of borate buffer (pH 10.0; 50 mM)--acetonitrile--tetrabutylammonium bromide (200 mM) (75:25:1, v/v/v). The fluorescence detector was set at 310 (excitation) and 430 nm (emission). Following protein precipitation with ice-cold acetonitrile, clear supernatants (50 microl) were injected into the HPLC system. The retention time of MPA was approximately 4.5 min. The HPLC run time was 8 min. The assay was linear in concentration range 0.2-20.0 microg/ml for MPA in human plasma. Precision of the assay in the concentration range examined was from 0.89 to 3.21% for the intra assay run and from 3.01 to 4.35% for the inter-assay run. A limit of detection was 0.05 microg/ml at a signal-to-noise ratio of 3. This validated method was then applied to the determination of MPA concentrations in renal transplant recipients after oral administration of 0.75 g of MMF. PMID- 11272312 TI - Spectrophotometric determination of ampicillin, dicluxacillin, flucloxacillin and amoxicillin antibiotic drugs: ion-pair formation with molybdenum and thiocyanate. AB - A sensitive spectrophotometric method is developed for the determination of some antibiotic drugs such as ampicillin (amp), dicluxacillin (dicl), flucloxacillin (fluc) and amoxicillin (amox). The method involves the formation of ion-pairs between these drugs under investigation and inorganic complex of Mo (V) thiocyanate followed by its extraction with methylene chloride. The optimum conditions for the ion-pairs formation are established. The method permits the determination of amp, dicl, fluc and amox over a concentration range of 1.5-77.5, 3-75, 1.5-79 and 7.5-75 microg ml(-1) respectively. The sensitivity (S) is found to be 0.017, 0.061, 0.014 and 0.073 microg cm(-2) for amp, dicl, fluc and amox, respectively. The method is simple, rapid, reproducible and accurate within +/- 1%. The method is applicable for the assay of the four drugs under investigation in different dosage forms and the results are in good agreement with those obtained by the official method. PMID- 11272313 TI - Directly coupled HPLC-NMR and HPLC-MS approaches for the rapid characterisation of drug metabolites in urine: application to the human metabolism of naproxen. AB - High resolution nuclear magnetic resonance (NMR) spectroscopy is a very powerful tool for the structural identification of xenobiotic metabolites in complex biological matrices such as plasma, urine and bile. However, these fluids are dominated by thousands of signals resulting from endogenous metabolites and it is advantageous when investigating drug metabolites in such matrices to simplify the spectra by including a separation step in the experiment by directly-coupling HPLC and NMR. Naproxen (6-methoxy-alpha-methyl-2-naphthyl acetic acid) is administered as the S-enantiomer and is metabolised in vivo to form its demethylated metabolite which is subsequently conjugated with beta-D-glucuronic acid as well as with sulfate. Naproxen is also metabolised by phase II metabolism directly to form a glycine conjugate as well as a glucuronic acid conjugate at the carboxyl group. In the present investigation, the metabolism of naproxen was investigated in urine samples with a very simple sample preparation using a combination of directly-coupled HPLC-1H NMR spectroscopy and HPLC-mass spectrometry (MS). A buffer system was developed which allows the same chromatographic method to be used for the HPLC-NMR as well as the HPLC-MS analysis. The combination of these methods is complementary in information content since the NMR spectra provide evidence to distinguish isomers such as the type of glucuronides formed, and the HPLC-MS data allow identification of molecules containing NMR-silent fragments such as occur in the sulfate ester. PMID- 11272314 TI - New reagents for detection of faecal occult blood. AB - Imipramine hydrochloride (IPH) and desipramine hydrochloride (DPH), two widely used antidepressant drugs, are proposed as new reagents for detection of faecal occult blood. The usefulness of IPH and DPH in occult blood detection has been examined and compared with benzidine and stanoccult methods. The results show that the proposed reagents are selective and sensitive and gives reproducible results. The proposed methodology is much less subject to vegetable peroxidase, iron and vitamin C interference and can be performed on patients who are on a normal diet. PMID- 11272315 TI - Determination of diclofenac sodium, flufenamic acid, indomethacin and ketoprofen by LC-APCI-MS. AB - A sensitive, selective and accurate high-performance liquid chromatography-mass spectrometry (LC-MS) assay for the determination of selected non-steroidal anti inflammatory drugs (NSAIDs), namely diclofenac sodium (DIC), flufenamic acid (FLU), indomethacin (IND) and ketoprofen (KET), either individually or in mixtures, was developed. The examined drugs were injected onto Shim-pack GLC-CN column and were eluted with a mobile phase consisting of acetonitrile and 20 mM ammonium acetate solution (5:1 v/v)/pH 7.4 at a flow rate l ml min(-1). The mass spectrometer, operated in the single ion monitoring mode, was programmed to admit the negative ions [M-H] at m/z 295.9 (DIC), 280.1 (FLU), 355.8 (IND) and 252.9 (KET), respectively. The calibration curves were linear (r > or = 0.9993) over the concentration range 50-300 ng ml(-1) (FLU, DIC) and 100-500 ng ml(-1) (KET, IND) with detection limits of 0.5-4.0 ng. The mean predicted concentrations for the analytes were in the range -5.9 and 5.2% of the nominal concentrations. Within-day and between-day precision were in the range of 0.8-9.1% of the R.S.D. Mean recovery percentages of the individual compounds from laboratory-made mixtures and pharmaceutical formulations were (99.5-101.5%) and (100.6-102.2%), respectively. PMID- 11272317 TI - Analysis of binary mixtures of losartan potassium and hydrochlorothiazide by using high performance liquid chromatography, ratio derivative spectrophotometric and compensation technique. AB - A new simple, precise, rapid and selective reversed-phase high performance liquid chromatographic (HPLC) and two spectrophotometric methods have been described for resolving binary mixture of losartan potassium and hydrochlorothiazide in the pharmaceutical formulations. The first method, is based on HPLC on a reversed phase column using a mobile phase 0.01 N sodium dihydrogen phosphate:methanol:acetonitrile (8:2:1 v/v/v) (pH 5.5) with detection at 265.0 nm. The second method, is depend on ratio derivative spectrophotometry, the amplitudes in the first derivative of the ratio spectra at 238.360 nm and at 230.423 nm were selected to simultaneously determine losartan potassium and hydrochlorothiazide in the mixture. The third method, based on compensation technique is presented for the derivative spectrophotometric determination of binary mixtures with overlapping spectra. By using ratios of the derivative maxima or the derivative minimum, the exact compensation of either component in the mixture can be achieved, followed by its determination. The accuracy and precision of the methods have been determined and they have been validated by analysing synthetic mixtures containing losartan potassium and hydrochlorothiazide. The methods do not require any separation step. The methods were also applied to the determination of losartan potassium and hydrochlorothiazide in pharmaceutical preparations. The analytical results were quite good in all cases. PMID- 11272316 TI - Study of partition of nitrazepam in bile salt micelles and the role of lecithin. AB - The effect of trihydroxy (sodium cholate and sodium glycocholate) and dihydroxy (sodium deoxycholate and sodium glycodeoxycholate) bile salt micelles on the spectrophotometric properties and on the solubility of nitrazepam in aqueous solution, at 25.0 degrees C and at ionic strength 0.1 M in sodium chloride, has been assessed. From the results obtained it was possible to calculate the partition coefficients (Kp) of nitrazepam between aqueous and micellar phases. The partition coefficients of nitrazepam have also been determined in mixed micelles of cholate or deoxycholate with lecithin (egg yolk phosphatidylcholine), which were used as a model of the gastrointestinal tract. Drug partition was found to depend on the bile acid (number of hydroxyl groups and conjugation with glycine), and our data indicate further that addition of lecithin to bile salt micelles decreases the values of the partition coefficients in the mixed micelles at physiological pH. PMID- 11272318 TI - Assessing molecular similarity/diversity of chemical structures by FT-IR spectroscopy. AB - FT-IR spectra have been investigated for their ability to distinguish compounds which are chemically diverse and to produce clusters of compounds which makes sense chemically. Principal component analysis (PCA) was applied to the analysis of a small database of FT-IR spectra. The effect of the data pretreatment step of log transformation on spectral data pattern was also visualized by using PCA plots. The method of sequential projection pursuit (SPP) was applied to detect inhomogeneities in the data. Finally, cluster analysis of these spectra, depending on unweighted pair-group average linkage, was carried out. PMID- 11272319 TI - Simultaneous identification and quantitative determination of neomycin sulfate, polymixin B sulfate, zinc bacytracin and methyl and propyl hydroxybenzoates in ophthalmic ointment by TLC. AB - A thin layer chromatographic-densitometric method for identification and quantitation of neomycin sulfate, polymixin B sulfate, zinc bacytracin and auxiliary substances (methyl and propyl hydroxybenzoates) in ophthalmic ointment was developed. To separate these constituents the silica gel coated TLC plates and two mobile phases were used. The suitable mobile phases were: methanol-n butanol-ammonia 25%-chloroform (14:4:9:12, v/v/v/v) for determination of antibiotics and n-pentane-glacial acetic acid (66:9, v/v) for methyl and propyl hydroxybenzoates. The antibiotic chromatograms were detected by using ninhydrin ethanol solution, while densitometric measurements were made at lambda = 550 nm. Hydroxybenzoates were identified by UV measurements at lambda = 260 nm. The constituents under consideration were well separated at sufficient detection level. The recovery for all constituents ranged from 98.08% to 104.95%. PMID- 11272320 TI - Non-ionic micellar affinity capillary electrophoresis for analysis of interactions between micelles and drugs. AB - Micellar affinity capillary electrophoresis (MACE) was introduced to evaluate the affinity of various kinds of drugs as benzoic acid, salicylic acid, trinitrophenol, p-hydroxybenzoic acid and o-acetylsalicylic acid. Non-ionic micelles as Brij 35 (polyethylenglycol dodecylether), Tagat (polyoxyethylene (20) glycerol monooleate) and Tween 20 (polyoxyethylen sorbitan monolaurate) were used as a pseudostationary phase in capillary electrophoresis. For polyvinyl alcohol (PVA) coated capillary was used in this examinations. The drugs had negative electrophoretic mobilities at a pH value of pH 7.2. The negatively charged drugs migrated toward the anode and were related by their interaction with the micelles. The difference in the mobility of the drugs owing to the presence of the micelles reflected the interaction between these drugs and the micelles. Equations were derived to calculate the capacity factor k' from the migration times in the presence of micelles t' and in the absence of micelles t, the partition coefficients Pwm and the Gibbs free energy. The drugs show different interaction and affinity with the micelles in the systems. Strong interaction was observed between benzoic acid and the micelles. Furthermore, a linear relationship (R = 0.999) was obtained between deltaG(o) and ln Pwm in the micellar solubilization of drugs. These results show that deltaG(o) can give us information on the affinity and on the partition behaviour of the drugs in these systems. PMID- 11272321 TI - 13C/12C isotope ratio MS analysis of testosterone, in chemicals and pharmaceutical preparations. AB - The 13C/12C ratio can be used to detect testosterone misuse in sport because (semi)-synthetic testosterone is supposed to have a 13C abundance different from that of endogenous natural human testosterone. In this study, gas chromatography/combustion isotope ratio mass spectrometry (GC/C/IRMS) analysis for the measurement of the delta 13C/1000 value of testosterone from esterified forms of 13 pharmaceutical preparations, six reagent grade chemicals and three bulk materials (raw materials used in pharmaceutical proarations) obtained world wide was investigated after applying a strong acidic solvolytic procedure. Mean delta 13C/1000 values of non esterified (free) testosterone from chemicals and bulk materials of several testosterone esters were in the range: -25.91/ 32.82/1000 while the value obtained for a (semi)-synthetic, reagent grade, free testosterone was -27.36/1000. The delta 13C/1000 results obtained for testosterone from the pharmaceuticals investigated containing testosterone esters were quite homogeneous (mean and S.D. of delta 13C/1000 values of free testosterone: 27.43 +/- 0.76/1000), being the range between -26.18 and 30.04/1000. Values described above were clearly different from those reported by several authors for endogenous natural human testosterone and its main metabolites excreted into the urine in non-consumers of testosterone (delta 13C/1000 range: from -21.3 to -24.4/1000), while they were similar to those of urinary testosterone and metabolites from individuals treated with testosterone esters and testosterone precursors. This finding justifies the fact that administration of these pharmaceutical formulations led to a statistical decrease of carbon isotope ratio of urinary testosterone and its main metabolites in treated subjects. PMID- 11272322 TI - Isolation and identification of a degradation product in a capsule formulation containing the elastase inhibitor, DMP 777. AB - An unexpected degradation product, greater than 0.10%, was observed in a DMP 777 capsule formulation stored at 40 degrees C/75% r.h. for 3 months and 25 degrees C/60% r.h. for 2 years. The degradant of interest was prepared in quantity by refluxing the drug substance in dilute acid. A preparative HPLC method was developed to separate the various degradants and to collect each as a separate fraction. Each fraction was analyzed by the analytical HPLC gradient test method to assure positive identification of each peak and to correlate each peak to the original capsule sample. Key isolated degradation products were used for structure elucidation with mass spectrometry and NMR. The major degradant of interest in the capsule formulation was found to be a carboxylic acid resulting from the acid hydrolysis of an amide bond. PMID- 11272323 TI - Determination of cisapride in pharmaceutical preparations using derivative spectrophotometry. AB - Derivative spectrophotometric and high performance liquid chromatographic methods (HPLC) were described for the determination of cisapride in pharmaceutical preparations. Spectrophotometrically, cisapride was determined by measuring the 1D-values at 264, 300 nm and 2D-values at 276, 290 and 276-290 nm. Beer's Law was obeyed in the range 2-12 microg ml(-1). The HPLC method depends upon using micropack-Si-10 column at ambient temperature with a mobile phase consisting of methanol-concentrated ammonia (99.25:0.75) at a flow rate of 1 ml min(-1). Quantitation was achieved by UV detection at 272 nm using quinine as internal standard. Calibration curve was linear over the concentration range 2-10 microg ml(-1). Both derivative spectrophotometry and HPLC methods showed good linearity, precision and reproducibility. No interference was found from tablet or suspension matrices at the selected derivative wavelengths and chromatographic conditions. The proposed methods were successfully applied to the assay of commercial tablets and suspension. The procedures were rapid, simple and suitable for quality control applications. PMID- 11272325 TI - A simple HPLC method for quantitation of enalaprilat. AB - A reversed-phase high performance liquid chromatography (HPLC) method with UV detection has been developed for the determination of enalaprilat. The method produced linear response over the wide concentration range of 1-200 microg/ml, with an average accuracy of 97.35 +/- 4.93%, as well as average intra- and iter day variations of 3.72 and 5.18%, respectively. The limits of detection and quantitation of the method were 0.125 and 0.5 microg/ml, respectively. The method was selective with respect to resolution of the peaks of enalaprilat and enalapril maleate. PMID- 11272324 TI - Determination of assay and impurities of gamma irradiated chloramphenicol in eye ointment. AB - A sample preparation method was developed to isolate chloramphenicol and its radiolytic products from an oily ointment base. The isolation method suspended the eye ointment in n-hexane at 45 degrees C, and isolated the target compounds as residue by centrifugation. It was found that the main element to ensure a satisfactory isolation was keeping the sample solution at 45 degrees C during sample preparation. Linearity, precision, accuracy and suitability of the method were confirmed valid for both assay and impurity tests. This isolation method was ideal for assay, unique for extraction of unexpected and complex radiolysis products, and had a number of advantages compared to the pretreatment methods described in The United Stares Pharmacopoeia and British Pharmacopoeia, in terms of accuracy, precision, and easy handling. The effect of gamma-irradiation on chloramphenicol eye ointment was studied by HPLC, after applying the developed sample preparation method. The present assay and impurity test methods with HPLC were confirmed to be suitable for irradiated chloramphenicol in eye ointment. Formation of radiolytic products induced by gamma-irradiation was evidenced by the impurity test. The assay test showed that active ingredient of chloramphenicol eye ointment decreased by 3.3% at an irradiation dose of 25 kGy and by 11.1% at 50 kGy. PMID- 11272326 TI - NMR characterization of a novel bile acid sequestrant, DMP 504. AB - DMP 504, a potential bile acid sequestrant for the treatment of hypercholesterolemia, is a highly insoluble, cross-linked polymer which does not lend itself to ordinary means of characterization used for drug substances in the pharmaceutical industry. Therefore, alternative characterization techniques have been sought. As part of an effort into extensive characterization of DMP 504 drug substance, nuclear magnetic resonance (NMR) was employed to provide insight into details of the DMP 504 polymer structure. The primary motivation for determining the structure of the polymer chain is to relate the DMP 504 structure to its performance properties as a bile acid sequestrant. Characterization of the polymer chain and understanding of the structural basis of its properties is essential in optimizing and controlling the manufacture of reproducible drug substance. NMR has proven a versatile tool for the description of polymer structure and dynamics because of the wide range of nuclear interactions affecting the NMR signal. This allows the design of experiments to elicit information about specific polymer interactions or properties. The methods of sample preparation utilized to obtain NMR spectra of the insoluble polymer, as well as a discussion and comparison of results for the characterization of DMP 504 obtained using several different NMR techniques will be presented. PMID- 11272327 TI - A simple and efficient high-performance liquid chromatographic assay for etomidate in plasma. AB - The development and validation of an effective and simplified LC assay for the quantitation of etomidate in beagle plasma is described. The methodology employs a rapid and simple protein precipitation procedure in combination with previously reported chromatographic conditions. Using a 0.3 ml aliquot of plasma, the assay is linear in the concentration range of 50 to 5000 ng/ml, with an extraction efficiency between 97 to 104% and accuracy between 98 and 105%. PMID- 11272328 TI - Validation of a CE assay for the analysis of isomeric aminopyridines and diaminopyridines. AB - A capillary electrophoresis assay for the analysis of aminopyridines and diaminopyridines has been developed and validated. The compounds were separated using a 100 mM sodium acetate buffer at pH 5.15, an applied voltage of 20 kV and a capillary of 60 cm effective length, N-(1-naphthyl)ethylenediamine was used as internal standard to compensate for injection errors and minor fluctuations of the migration times. The detection wavelength was set at 240 nm and not optimized for a specific derivative. The assay was validated with respect to specificity, linearity, range, limit of quantitation and detection, precision, and robustness. Within certain limits, the assay also allowed the detection and determination of the other aminopyridine derivatives at the 0.1% level as demonstrated for 3,4 diaminopyridine. PMID- 11272329 TI - Oxidative degradation study of nitrendipine using stability indicating, HPLC, HPTLC and spectrophotometric method. AB - The objective of this study is to develop validated stability indicating HPLC (A), HPTLC (B) and spectrophotometric (C) method for the estimation of nitrendipine. The stability indicating capability of the assays is proved using forced degradation, by exposing drug solution to sunlight, acidic and alkaline medium. The chromatogram and UV spectrum showed nitrendipine well resolved from the degradation product. Degradation of drug is found faster in acidic (0.1 N hydrochloric acid) medium as compared to alkaline (0.1 N sodium hydroxide) medium at 100 degrees C. Also, photodegradation is studied, with special emphasis on the effect of solvents like methanol (1), chloroform (2), dichloromethane (3), acetone (4) and ethyl acetate (5), on the rate of photodegradation. The degradation of title compound followed first order kinetics in all cases. Estimation of the drug is carried out by the stability indicating methods mentioned, using one point standardization within the linearity range of interest. The methods are compared in respect of performance precision and accuracy. Major route of degradation in all cases is found to be oxidation and degradation product is confirmed as dehydronitrendipine by the use of relevant UV, IR and 1H NMR spectrometry. PMID- 11272330 TI - Improved high performance liquid chromatographic analysis of omeprazole in human plasma. AB - A simple high-performance liquid chromatographic method was developed for the determination of omeprazole in human plasma. Omeprazole and the internal standard, chloramphenicol, were extracted from alkalinized plasma samples using dichloromethane. The mobile phase was 0.05 M Na2HPO4-ACN (65:35, v/v) adjusted to pH 6.5. Analysis was run at a flow rate of 1.0 ml/min at a detection wavelength of 302 nm. The method was specific and sensitive with a detection limit of 2.5 ng/ml at a signal-to-noise ratio of 4:1. The limit of quantification was set at 5 ng/ml. The calibration curve was linear over a concentration range of 5-1280 ng/ml. Mean recovery value of the extraction procedure was about 96%, while the within and between day coefficient of variation and percent error values of the assay method were all less than 14%. PMID- 11272331 TI - Antithrombotic treatment in protection against thrombogenic effects of 5 fluorouracil on vascular endothelium: a scanning microscopy evaluation. AB - Cardiotoxicity is a serious side effect of treatment of malignant diseases with 5 fluorouracil (5-FU). The underlying pathophysiologic mechanism remains unclear but clinical data suggest that the endothelium of coronary arteries may be involved. Experimental studies indicate that the endothelium is especially susceptible to 5-FU and support the hypothesis that a thrombogenic effect of 5 FU, secondary to its direct toxic effect on the endothelium, is one of the pathophysiologic mechanisms behind 5-FU-induced cardiotoxicity. In the present study we evaluate the role of antithrombotic treatment with dalteparin as protection against the thrombogenic effect of 5-FU on the vascular endothelium in a rabbit model. The effects on the vascular endothelium of 5-FU, dalteparin, and the combination of these two substances were evaluated with scanning electron microscopy 1, 3, 7, 14, and 30 days after treatment and compared with a control group. Very severe damage to the endothelium was seen in 5-FU-treated animals, often leading to intima disruption and denudation of underlying structures, with accompanying platelet accumulation and fibrin formation. The most extensive damage was observed on Day 3 after treatment. The cytotoxic effect of 5-FU was partly reversible. The combination of 5-FU and dalteparin gave lower scores on Day 3 because of less evidence of thrombotic events. However, the reversibility of the endothelial damage was poorer in this group, as well as in the group that received dalteparin alone. The findings support the hypothesis that antithrombotic treatment with dalteparin can protect against the thrombogenic effect of 5-FU, secondary to its direct toxic effect on the vascular endothelium. However, the study indicates that dalteparin per se has a toxic effect on the endothelium that is different from that of 5-FU. PMID- 11272332 TI - Characterization and identification of local defects in glass. AB - The most relevant defects in glasses are categorized and investigated by appropriate microanalytical techniques. Since these defects very often present a real challenge because of complex chemical and mineralogical properties, a multimethod approach is necessary to supplement or confirm the findings from scanning techniques. The combination of electron probe microanalysis/energy dispersive x-ray (EPMA/EDX) and laser ablation inductively coupled plasma mass spectroscopy (LA-ICP-MS) allows the determination of element trace concentrations in a knot, a glassy defect, thus finally enabling the identification of a special source of the defect from otherwise nondistinguishable refractories. The type of crystals can be determined exactly by the use of EPMA and x-ray dispersion (XRD), stones (a crystalline agglomerate) are analyzed by EPMA/EDX pointing to a possible source for this defect; results on metallic inclusions, "filled bubbles," and surface defects are reported and defect sources are discussed. Since close cooperation with the production departments and knowledge of production techniques and conditions are necessary for the diagnosis to take appropriate countermeasures, an approach is presented which systematically accumulates all information into a data base. The structure of such "expert systems" is described, which leads to correlation of appearance, analytical data, and source of the defects for even more accurate diagnosis and faster reaction. PMID- 11272333 TI - Natural color scanning electron microscopy based on the frequency characteristics of the human visual system. AB - The principles of image formation in natural color scanning electron microscopy (NC-SEM) are discussed in detail. This method is based on the frequency characteristic of the human visual system. It is shown that the Mach effect and masking effect are important in the characteristics. The former, which can enhance structural details, is visually similar to the edge effect in secondary electron (SE) images, and the latter is required for proper representation of very degraded color information obtained from a light microscope. When using these effects suitably, an NC-SEM image with the resolution equivalent to that of an SEM image can be acquired, though it is composed of an SEM image and a special video microscopy (VM) image with a resolution much lower than the SEM image of the identical view. The NC-SEM is more effective than the SEM in observation. interpretation, and analysis of various samples with important color information. PMID- 11272334 TI - C-banding visualized by atomic force microscopy. AB - C-banding is a method used for studying chromosome rearrangements near centromeres and for investigating polymorphisms. In human chromosomes, the C bands are located at the centromere of all the chromosomes and the distal long arm of the Y chromosome. In this study, we aimed to detect the structural changes in chromosomes during the stages of C-banding by atomic force microscopy. We observed crater-like structures in the chromosomes after 2xSSC (saline sodium citrate) treatment and measured the relative difference between the heights of chromatid and centromere of the chromosomes. Results showed that the relative difference was 3 nm in chromosomes 1, 9, 16, and Y, whereas in the other chromosomes this value was 11.6 nm. After Giemsa staining, the relative difference increased by a factor of 16 in chromosomes 1, 9, 16, and Y. The other chromosomes showed no such increase, which is in accordance with our suggestion that nonhiston proteins associated with DNA in constitutive heterochromatin can make the constitutive heterochromatin resistant to C-banding. PMID- 11272335 TI - Scanning electron microscopy of nonconductive specimens at critical energies in a cathode lens system. AB - A method for scanning electron microscopy imaging of nonconductive specimens, based on measurement and utilisation of a critical energy, is described in detail together with examples of its application. The critical energy, at which the total electron yield curve crosses the unit level, is estimated on the basis of measurement of the image signal development from the beginning of irradiation. This approach, concentrated onto the detected signal as the only quantity crucial for the given purpose of acquiring a noncharged micrograph, evades consequences of any changes in an irradiated specimen that influence the total electron yield curve and possibly also the critical energy value. Implementation of the automated method, realised using a cathode lens-equipped scanning electron microsope (SEM), enables one to establish a mean rate of charging over the field of view and its dependence on the electron landing energy. This dependence enables one to determine the energy of a minimum damage of the image of the given field of view. Factors influencing reliability and applicability of the method are discussed and examples of noncharged micrographs of specimens from both life and material science fields are presented. PMID- 11272336 TI - Investigation of foreign substances in food. AB - Scanning electron microscopy (SEM) together with energy-dispersive spectroscopy (EDS) and Fourier transform infrared spectroscopy (FTIR) were used to investigate foreign substances from seven categories of foreign substances in food. (1) Naturally occurring foreign substances--Using FTIR, a foreign substance was identified as a natural resin probably from the product. (2) Foreign substances introduced during food processing. Scanning electron microscopy-EDS was used to identify a foreign material found on surf clams as calcium phosphate from a product/ingredient interaction. Using SEM-EDS, a crystalline material in a meat product was identified as calcium salts of chloride and phosphate. Fourier transform infrared spectroscopy was used to identify foreign material that clogged an aerosol valve as chipboard. Using SEM-EDS, the metal in the heel of a glass bottle was identified as copper sulfide-containing metal inclusion. (3) Insects, reptiles, and rodents--Scanning electron microscopy was used to determine that a mouse found in food was not processed with the food, but entered the container after it left the factory. (4) Glass fragments--Glass from various sources can be distinguished from one another using SEM-EDS either by the level of the major elements in glass or by the presence of elements in one glass, but not in another. (5) Glass-like particles--Using SEM-EDS, glass-like particles found on beets were determined to be a fatty acid. (6) Metal foreign objects- Using SEM-EDS, metals from a variety of sources can be easily distinguished. For example, a tin-soldered container can be distinguished from a lead-soldered can. Using SEM-EDS, the metal fiber found on the bottom of a two-piece can likely enter the can during the final stage of the manufacture of the drawn and ironed food can. (7) Drug capsule identification--Fourier transform infrared spectroscopy was used to determine that a pill found in food was ibuprofen. PMID- 11272337 TI - The swelling behavior and wet morphology of water-absorbable polymer materials. AB - In this research work, a low-temperature examination method for scanning electron microscopy (SEM) is introduced. A water-absorbable polymer material, the sulfonated polyethylene (SPE) ion-exchange hollow fiber membrane, was used for the experiments. With this low-temperature technique, the wet morphology of the water-absorbable sulfonated polyethylene hollow fiber membrane was revealed. The results obtained from this investigation offer some important information to explain the behavior of the SPE hollow fiber membranes when they are applied in pervaporation separation of water/organic solvent mixtures, such as water/ethylene glycol, water/ethanol, and so forth. PMID- 11272338 TI - Three-dimensional localisation of fluorescence resonance energy transfer in living cells under two-photon excitation. AB - Three-dimensional (3-D) imaging of fluorescence resonance energy transfer (FRET) in human cells under two-photon excitation was demonstrated in this study. A sample was prepared by expressing a donor and an acceptor in living cells and using an antibody to secure the proximity of contact between the donor and the acceptor. The quenching of fluorescence emission of a donor in the double labelled cells indicates the presence of FRET that occurred in these living cells. Because of the quadratic relation of the excitation power, 3-D localisation of FRET becomes possible. PMID- 11272340 TI - Perspectives. HHS's new big cheese: compassionate conservative who wielded the imperial veto. PMID- 11272339 TI - A calix[4]arene based calcium-selective optode membrane: measuring the absorbance maximum wavelength shift. AB - An absorbance based calcium ion sensor is presented which applies a new immobilized chromogenic calix[4]arene. The membrane consists of a hydrogel/1,3 pentanedial mixture and the 1,3-bis(indoaniline)-derived 2,4-bis [(ethylcarbonyl)methoxyl-calix[4]arene as calcium receptor. The indicator entrapped within the gel was not rapidly washed out. Upon calcium complexation a bathochromic shift up to 70 nm was observed along with an increase of the absorption coefficient. The dynamic range of calcium measurements is from 3 to 10 mM with a point of inflection of the calibration curve at a calcium concentration of about 7 mM. The concentration of calcium ions can be calculated from either absorbance at a distinct wavelength or--preferably when indicator leaching occurs -from the wavelength shift of the absorbance occurring when calcium levels are increased. PMID- 11272341 TI - Maternal depressive symptoms and child behavior problems in a nationally representative normal birthweight sample. AB - OBJECTIVE: To evaluate the association between maternal depressive symptoms and child behavior problems in a nationally representative sample of U.S. mothers of normal birthweight babies. METHODS: We analyzed data from the 1988 National Maternal and Infant Health Survey (NMIHS) and a 1991 follow-up survey. Depressive symptoms were measured at both surveys using the CES-D, and child behavior problems were assessed by maternal self-report at follow-up. RESULTS: Approximately 28% of the 5303 mothers reported depressive symptoms at a mean of 17 months after delivery, as did 20% at 36 months. In multivariate analyses, women with depressive symptoms at either or both surveys were significantly more likely than women without depressive symptoms to report that their children had frequent temper tantrums or difficulty getting along with other children, and were difficult to manage, unhappy, or fearful. Compared to women without depressive symptoms, the risks of reporting three out of the five child behavior problems for women with depressive symptoms were OR = 1.6 (CI = 1.1-2.1), 1988 only; OR = 2.3 (CI = 1.6-3.3), 1991 only; and OR = 3.6 (2.6-5.0), both 1988 and 1991. CONCLUSIONS: Study findings indicate that a substantial proportion of mothers of young children in the United States experience depressive symptoms and that their children are at significantly increased risk of maternally reported behavior problems. Our results suggest that efforts to identify and treat depression in new mothers should be increased and that mothers whose children are found to have behavior problems should be assessed for depression. PMID- 11272342 TI - Use of a comprehensive state birth data system to assess mother's satisfaction with length of stay. AB - OBJECTIVE: To assess length of stay, home visits, and mothers' feelings after full implementation of a law requiring a 48-hour minimum stay for women with normal vaginal deliveries. METHODS: The New Jersey Electronic Birth Certificate System (EBC) was used to capture demographic characteristics, and length of stay (LOS), and to select a sample of women with low risk uncomplicated vaginal deliveries. A follow-up mail survey (with a nonresponder phone component) enhanced the information available on the EBC. RESULTS: The response rate for women included in the sample was 82.1% (1276/1555). The mean length of stay was 1.9 days. Sixty-two percent (787) of women thought their LOS after delivery was just right. Women who thought their stays were too short tended to be older, married, working outside the home, or have an LOS of less than 48 hours. The most common reasons for thinking the LOS was too short was a need for rest and concern about the baby. DISCUSSION: The combination of an augmented electronic birth certificate system and a follow-up survey proved to be a rapid, reliable, and inexpensive method of assessment. The mothers' desires for rest, education on the care of her newborn, and reassurance that any medical complications could be handled, are paramount concerns that need to be taken into account by payers and practitioners wanting to decrease LOS. PMID- 11272343 TI - Perceptions of hormonal contraceptive safety and side effects among low-income Latina and non-Latina women. AB - OBJECTIVES: We explored perceptions of the safety and side effects of oral and injectable hormonal contraceptives among low-income women at high risk of unintended pregnancy. METHODS: Overall safety perceptions, specific health concerns, and the relationship between these safety perceptions and contraceptive choices were determined by focus groups and questionnaires obtained from white non-Latina (n = 19), English-speaking (n = 21), and Spanish-speaking Latina women (n = 19). RESULTS: Uncertainty or ambivalence about the safety of oral and injectable contraceptives was reported by 41% and 70% of respondents respectively, while 20% considered these methods to be mostly harmful. Personal experiences and stories from social networks proved to be more salient than medical opinions in shaping safety perceptions. Side effects and concerns about long-term health effects were common themes. While white non-Latina women focused predominantly on physical side effects, emotional side effects also contributed to Latinas' decisions about contraceptive switching. Spanish-speaking Latinas differed from English-speaking Latinas in other attitudinal dimensions, contraceptive use prevalence, and access to contraceptive services. CONCLUSION: Low-income mothers lacked confidence in method safety and had many concerns about the side effects of oral and injectable contraceptives. Because such concerns can be a barrier to contraceptive use, these perceptions need to be corrected to encourage more effective use of hormonal methods and to prevent unintended pregnancies. Culturally appropriate interventions should focus on client-provider interactions, social networks, and access to care. PMID- 11272344 TI - Changes in births to foreign-born women after welfare and immigration policy reforms in California. AB - OBJECTIVES: To determine whether passage of welfare and immigration policies was followed in California by changes in births to foreign-born women in California with respect to total numbers, payer sources, prenatal care use, or health outcomes. METHODS: Comparison of births to foreign-born and US-born women from 1990 to 1997 using adjusted odds ratios generated with multivariate logistic regression. RESULTS: Policies passed in 1994 and 1996 were followed by decreases in adjusted odds of births to foreign-born women with prenatal Medicaid coverage, without a corresponding increase in uninsured foreign-born women. There was no decline in the use of prenatal care by foreign-born women, and no worsening of birth outcomes after passage of the reforms. Foreign-born women, however, remained more likely to have inadequate prenatal care than US-born women, and the improvement in outcomes that occurred for US-born women from 1994 to 1997 did not occur for foreign-born women. CONCLUSIONS: In spite of the fact that pregnant immigrant women remained eligible for Medicaid after passage of welfare and immigration policies in California, the volume of births to foreign-born women using Medicaid declined. The lack of a corresponding increase in births to uninsured foreign-born women appears to have prevented deterioration in the use of prenatal care or birth outcomes. PMID- 11272345 TI - Risk factors for late or no prenatal care following Medicaid expansions in California. AB - OBJECTIVES: To describe the characteristics and risk factors of women with only third-trimester (late) or no prenatal care. METHODS: A statewide postpartum survey was conducted that included 6364 low-income women delivering in California hospitals in 1994 and 1995. RESULTS: The following factors appeared most important, considering both prevalence and association with late or no care: poverty, being uninsured, multiparity, being unmarried, and unplanned pregnancy. Forty-two percent of women with no care were uninsured, and uninsured women were at dramatically increased risk of no care. Over 40% of uninsured women with no care had applied for Medi-Cal prenatally but did not receive it. Risks did not vary by ethnicity except that African American women were at lower risk of late care than women of European background. Child care problems were not significantly associated with either late or no care, and transportation problems (not asked of women with no care) were not significantly related to late care. CONCLUSIONS: Lack of insurance appeared to be a significant barrier for the 40% of women with no care who unsuccessfully applied for Medi-Cal prenatally, indicating a need to address barriers to Medi-Cal enrollment. However, lack of financial access is unlikely to completely explain the dramatic risks associated with being uninsured. In addition to eliminating barriers to prenatal coverage, policies to reduce late/no care should focus on pre-pregnancy factors (e.g., planned pregnancy and poverty reduction) rather than on logistical barriers during pregnancy. PMID- 11272346 TI - Baltimore's consumer ombudsman and assistance program: an emerging public health service in Medicaid Managed Care. AB - OBJECTIVES: To present Baltimore City Health Department's (BCHD) experience in developing and operating an ombudsprogram for Maryland's Medicaid Managed Care HealthChoice Program as an innovative public health response to its MCH Title V assurance functions. METHODS: This paper presents a case study that 1) describes Baltimore City's Consumer Ombudsman and Assistance Program (COAP) in terms of development, function, structure, and resources; 2) provides a summary of its first 30 months' experiences, both quantitatively and qualitatively; and 3) describes COAP's successes, value, and limitations with respect to its three essential roles--a) conflict resolution for individuals, b) education for consumers, providers and advocates, and c) catalyst for quality improvement. OUTCOMES: Over 1300 cases (involving enrollment, access, billing, and treatment issues) were referred to COAP by the State's Complaint Resolution Section during the first 2 1/2 years of HealthChoice. Ombudsman interventions resulted in conflict resolution for enrollees using a continuum of education, mediation and advocacy, and in generating systematic data for systems change through collaboration with state and community public health, managed care organization, provider, consumer, and advocacy officials and groups. CONCLUSIONS: Public health ombudsprograms can effectively assist and educate enrollees; and provide concurrent, or real-time, information for consumer, provider, health plan, and advocacy groups, as well as public policymakers and legislators to better inform systems improvement and innovation. The community-based ombudsman role is an effective mechanism to ensure appropriate care for MCH populations and others with special needs. Such efforts can be funded by federal/state Medicaid administrative funds and are a sound investment in assuring access to comprehensive care for vulnerable populations. PMID- 11272347 TI - Enhancing technology skills of maternal and child health professionals. AB - OBJECTIVE: Information technology skills are essential for effective and efficient practice in maternal and child health (MCH). METHODS: Prior to the beginning of a web-based analytic skills training course, enrolled MCH professionals confidence in using e-mail and other Internet skills and beliefs about their usefulness were assessed. RESULTS: The assessment showed that participants believed strongly in the value of the specific technology skills but confidence in the ability to use these skills was low. CONCLUSION: An online technology skills training module was developed based on skills needed for the course to ensure that course participants learned and practiced the technology skills necessary to successfully complete the online analytic skills course. We describe the development, implementation, and results of the online technology skills module. PMID- 11272348 TI - Gamma vs beta irradiation: which is superior? AB - BACKGROUND: The debate over which radioisotope, gamma-emitting or beta-emitting, is ideal for vascular brachytherapy, is as old as the field itself. Gamma isotopes such as Ir-192 have been successfully used in large-scale clinical trials. However, radiation protection and safety remains a practical issue when high energy gamma rays are used in interventional suites. METHODS: In this report, we have evaluated dosimetric properties of currently available radiation sources, gamma and beta, in terms of their utility in vascular brachytherapy. Published data have been analyzed in comparing the depth doses of these two radiation modalities. RESULTS AND CONCLUSIONS: Our evaluation shows that significant dosimetric differences do exist between gamma and beta sources. The superiority of one over the other in clinical utility will more likely be determined by the outcome of human trials. PMID- 11272349 TI - First vascular brachytherapy, catheter used in human coronaries. PMID- 11272350 TI - Is the pathology of radiation injury different in small vs large blood vessels? PMID- 11272351 TI - Intracoronary radiation therapy for restenosis prevention: status of the clinical trials. AB - BACKGROUND: Vascular brachytherapy is an evolving field for the prevention of restenosis. The rationale for such therapy is that radiation prevents neointimal proliferation and vessel constriction following vascular injury. In recent years, numerous animal experiments have demonstrated that low doses of radiation when applied following intracoronary intervention reduces proliferation and alters the restenosis rate. This novel approach introduced specific vascular radiation biology and radiation physics aspects, new devices, and platforms to deliver the radiation intraluminally. METHODS: A variety of beta or gamma isotopes for use in vascular brachytherapy has been suggested. These isotopes need to be delivered intracoronarily to provide the therapeutic dose to the target cell via a safe and user-friendly platform. Two platforms of delivery are available and in practice: catheter-based systems and radioactive stenting. However, these novel applications have to be proven in the clinical arena. RESULTS: We have been conducting clinical trials with intracoronary brachytherapy since 1994 and are gathering information regarding the feasibility, safety, and effectiveness of this technology. Currently we are in the midst of an intensive clinical investigation using various devices, isotopes doses, and techniques. This report summarizes all the clinical work that has been conducted so far and is currently under investigation in this field. CONCLUSIONS: Without having the results of the majority of the trials described in this paper, it is difficult at this stage to draw firm conclusions regarding the future of this technology. However, the information presented will serve as a tool for the reader to judge the direction of vascular brachytherapy for prevention of restenosis. PMID- 11272352 TI - Initial studies with gamma radiotherpay to inhibit coronary restenosis. AB - BACKGROUND: Early studies indicate brachytherapy is a potential clinical treamtent for restenosis after coronary angioplasty procedures. The objective of this study was to evaluate the safety and efficacy of gamma radiation plus stenting in patients' previous restenosis. METHODS: In patients with previous coronary restenosis, balloon dilatation and/or coronary stenting was undertaken and then patients were randomly assigned to receive either Iridium-192 or placebo. Quantitative coronary angiographic, intravascular ultrasonographic, and clinical follow-up was obtained. RESULTS: Of the 55 patients enrolled, 26 were treated with Iridium-192 and 29 were in the placebo group. Late luminal loss was significantly lower in the treated group compared to the placebo group (0.38 +/- 1.06 mm vs. 1.03 +/- 0.97 mm, p = 0.03). Restenosis (stenosis of > or = 50% at follow-up) was found in 17% of treated patients compared to 54% of placebo patients (p = 0.01). The need for target lesion revascularization was observed in 12% of patients in the treated group compared to 45% in the placebo group (p = 0.01). CONCLUSIONS: In this initial trial, at 12 months follow-up, patients with previous restenosis were benefited by catheter-based gamma radiation therapy. PMID- 11272353 TI - Two-year follow-up after intracoronary gamma radiation therapy. AB - BACKGROUND: Neointimal hyperplasia and unfavorable remodeling have been demonstrated to be the major limitation to endovascular revascularization procedures. Intracoronary gamma radiation therapy has been shown to reduce the restenosis index. However, the late effects of these novel procedures are unknown. MATERIALS AND METHODS: To evaluate the long-term effects on clinical and angiographic outcome of endovascular gamma radiation therapy following percutaneous transluminal coronary angioplasty (PTCA), serial angiography over a 2-year period was performed in 21 patients (22 lesions) who were treated with 192Ir in doses of 20-25 Gy after PTCA. Angiograms were analyzed using quantitative methods (QCA). RESULTS: The mean late loss between PTCA and 6 months was 0.20 +/- 0.59 and 0.13 +/- 0.84 between 6 months and 2 years. At 6 months, angiographic binary restenosis was present in six arteries (27.2%). At 2 years, binary restenosis was observed in six arteries (27.2%), including one patient who had developed restenosis and excluding one patient with spontaneous regression. Two early pseudoaneurysms and two late aneurysms were observed at 6 months, with little increase at 2 years. No other angiographic complication was observed. None of the patients or medical staff developed complications or illnesses that could be related to the effects of the radiation procedure. CONCLUSIONS: Gamma radiation therapy decreases late luminal loss, is safe and free of unexpected complications at 6 months follow-up, with no significant changes or late complications at 2-years' follow-up. PMID- 11272354 TI - External beam radiation therapy to prevent postangioplasty dialysis access restenosis: a feasibility study. AB - PURPOSE: To evaluate the feasibility and short-term side-effects of postangioplasty external beam radiation for patients with compromised arteriovenous dialysis accesses. MATERIALS AND METHODS: Ten patients with compromised arteriovenous dialysis accesses were studied. Following confirmation of access compromise by an angiogram, patients were treated by a standard angioplasty +/- stent deployment. The target volume incorporated a margin of 1 cm beyond the angioplastied segment. The first 5 patients were treated to a dose of 12 Gy in two 6-Gy fractions spaced 48 h apart, whereas the next 5 patients were treated to 8 Gy in two similarly separated 4-Gy fractions. Five of the patients had at least one prior access that had failed. The current access had been in use for 6-52 months, and 5 of the 10 patients had at least one episode of compromise involving the current access. The length of stenosis ranged from 2 to 9 cm (mean 4.4 cm). All patients were followed clinically for adequacy of dialysis; a radiological follow-up with a fistulogram was performed for all in 3-monthly intervals. Follow-up ranged from 4 to 10 months, with a median follow-up of 6 months. RESULTS: There were no procedure-related complications. Three patients developed a restenosis at the site of the original stenosis, and one patient developed a restenosis at the edge of the stent. As part of the natural history of this process, five patients have also developed new lesions elsewhere in the proximal veins requiring radiological intervention. None of the patients showed any radiation-related side effects, either in the skin/ soft tissues, or in the vasculature on follow-up angiograms. CONCLUSIONS: Several different radiotherapeutic approaches are being currently investigated to prevent postangioplasty restenosis. There are several issues involved with the use of endovascular brachytherapy in these patients. On the other hand, given the superficial location of the dialysis access, we believe that external beam radiation offers a safe and simple method of radiation therapy in this setting. The clinical efficacy of this modality needs to be established through a randomized phase III trial. PMID- 11272356 TI - The experimental animal models for assessing treatment of restenosis. AB - Coronary restenosis after percutaneous interventions remains a major clinical problem. The assessment of therapies for the prevention of restenosis relies on the use of experimental models. This review describes the most frequently used animal models of coronary artery retenosis and the intraspecies differences among them, particularly in the extent and composition of the neointimal thickening. These differences in neointima formation should be considered in the interpretation of effective antiproliferative therapies before they are transferred into clinical trials. PMID- 11272357 TI - Comparability of the external vs internal location of radiation in inhibiting neointimal hyperplasia. AB - PURPOSE: One of the most controversial issues in utilizing radiation to inhibit restenosis is the importance of the location of the radiation source. The experimental results from using external forms of radiation have been contradictory and conflicting. In this study, we undertook to externally place a brachytherapy catheter source and to determine if a dose-response effect could be observed, as has been demonstrated with endovascular locations. MATERIALS AND METHODS: Neointimal hyperplasia was created in a rat carotid artery model by a balloon catheter technique. Immediately following injury, treatment groups received irradiation via an externally located high-dose rate (HDR) 192Ir brachytherapy catheter. This system allows for a more uniform dose delivery compared with endovascular radiation sources. Radiation was delivered to a 2-cm length of the injured vessel at doses of 5, 10, or 15 Gy and the animals were sacrificed at various time points following treatment (24 h to 6 months). Serial sections of tissue were stained immunohistochemically with primary antibodies for CD11b, platelet-derived growth factor (PDGF), and alpha-smooth muscle actin. RESULTS: Radiation doses of 5, 10, and 15 Gy inhibited the appearance of neointimal hyperplasia in a dose- and time-dependent manner. That is, doses of 5 15 Gy allowed for varying degrees of neointimal hyperplasia at 3 weeks posttreatment, with a greater resurgence of monocyte/macrophage activity at 5 Gy than at 10 or 15 Gy, where an absence of macrophage activity and PDGF expression was noted. From 2 to 6 months, the 10 and 15 Gy doses were again more suppressive of neointimal hyperplasia than 5 Gy, and at 6 months posttreatment the doses were approximately 25% and 50% effective, respectively. CONCLUSIONS: The demonstrated effectiveness of external brachytherapy provides "proof of principle," that it is the radiation dose delivered to the arterial wall, and not the location of the source, which is critical to a successful outcome. Ablation of the resident monocyte/macrophage population (or prevention of their activation) occurs with low to moderate doses of irradiation, leading to the absence of a cytokine cascade as evi denced by a lack of PDGF expression. A favorable therapeutic ratio exists, therefore, for radiation treatment of the arterial vasculature to prevent neointimal hyperplasia postangioplasty. PMID- 11272358 TI - Review of endovascular brachytherapy physics for prevention of restenosis. AB - Intraluminal irradiation of coronary and peripheral arteries has been shown to reduce neointimal hyperplasia following balloon angioplasty, thereby inhibiting restenosis. Several irradiation techniques are being investigated, including temporary intravascular insertion of high activity gamma- or beta-emitting seeds and wires; inflation of dilatation balloon catheter with radioactive liquid or gas; insertion of miniature x-ray tubes via coronary catheters; permanent implantation of radioactive stents; and postangioplasty fractionated external beam irradiation. Unlike conventional brachytherapy, intravascular treatment of restenosis requires accurate knowledge of dose at distances of 0.5-5 mm from the radioactive source. This requirement presents special problems with regard to source calibration and dose specification, because dose gradients at such close distances from a radioactive source are extremely large. This makes it virtually impossible to define the characteristics of an ideal radiation source without some knowledge of the location and radiosensitivity of the target tissues, plus the radiotolerance of normal tissues. Hence, the current debate over whether beta or gamma sources are to be preferred. Imprecise knowledge of dose-volume effects for coronary arteries, plus uncertainties in the biological time sequencing of restenosis fuel a second debate on whether external beam treatments may be efficacious, and whether or not permanent radioactive stents may prove superior to high dose, single fraction brachytherapy. We review here the dosimetric properties of the various irradiation techniques and isotopes that have been proposed, including aspects of radiation safety, dose homogeneity, and practical aspects of source delivery. PMID- 11272359 TI - On the depth of penetration of photons and electrons for intravascular brachytherapy. AB - PURPOSE: To investigate the depth dose characteristics of various radionuclides under consideration for intravascular brachytherapy (IVB). MATERIALS AND METHODS: In the past few years, various preclinical studies have shown that 10-30 Gy of ionizing radiation delivered by a brachytherapy treatment may inhibit restenosis following angioplasty. A number of new delivery systems using various radionuclides have been developed and are being investigated for IVB. Typical target size for IVB is in the range of millimeters, in contrast to conventional brachytherapy for cancer in which the target may be 1-5 cm in size. The question addressed in this paper is: whether lower energy photon emitters and even beta emitters, which are not commonly used for intracavitary brachytherapy of cancer, may provide a depth of penetration adequate for IVB. To explore this issue, radial dose functions for photons and electrons in the range of 1-10 mm in water were calculated using Monte Carlo simulation. Reference depth for normalization of the radial dose funtion was chosen to be 2 mm. RESULTS: Radial dose functions have been calculated for monoenergetic photons with energies of 0.01, 0.02, 0.04, 0.06, 0.08, 0.10, 0.20, 0.40, and 1.00 MeV and monoenergetic electrons with energies of 0.5, 1.0, 1.5 and 2.0 MeV. Also, the same calculations have been performed for 192Ir, 125I, and 103Pd gamma or x-ray sources as well as 90Sr-90Y, 32P, and 188Re beta-emitting sources. Results are also provided for selected cases in a simulated calcified lesion in water. CONCLUSIONS: It is concluded that photons above an energy of 20 keV and electrons above an energy of 1.0 MeV are acceptable from the point of view of adequate depth of penetration for IVB in tissue. PMID- 11272355 TI - Radiobiological principles in intravascular irradiation. AB - BACKGROUND: There is wide interest in the use of intravascular irradiation to control restenosis following balloon angioplasty. Part of the mechanism of restenosis appears to be the proliferation of smooth muscle cells (SMCs), triggered to divide by the damage caused by the angioplasty, and this proliferation can be inhibited by irradiation with y-rays or beta-rays. METHODS: In vitro data for the survival of smooth muscle cells exposed to radiation was used to model the likely control of restenosis by radiation. Physical and biological data were used to estimate differences in biological effectiveness of gamma-rays and beta3-rays, as well as the effect on cell killing of extending the exposure time. RESULTS: Based on the radiosensitivity of SMCs, measured in vitro, and the limited proliferative potential of these normal somatic cells, it is possible to understand and to model quantitatively how a single acute gamma-ray dose in the range of 15-20 Gy can inhibit restenosis. The few successful trials carried out to date where radiation has been shown to inhibit restenosis have all involved the gamma-emitter Iridium-192. The use of this radionuclide involves radiation safety problems and an inconveniently long treatment time. Consequently, there is much interest in developing a beta-emitting source that would solve both problems, and a number of different possibilities are being pursued. This development introduces two new problems discussed in this paper. First, beta-rays in the megavoltage range are less effective biologically than gamma-rays in the kilovoltage range, but the magnitude of the difference is not well known. Second, in the case of a single large dose, such as that proposed to inhibit restenosis, the biological effect will vary substantially as the exposure time varies from 1 to 20 min. If the clinical data are to be compared between centers using a variety of beta and y-emitting radionuclides, these factors will need to be taken into account. CONCLUSIONS: Doses of > 15 Gy are unlikely to result in elimination of the restenosis problem but should delay onset of restenosis for a significant period; the larger the dose, the longer the delay. Successful trials of endovascular radiation completed to date involve y-rays, while many systems being developed are based on beta-emitting radionuclides. Experimental data are urgently needed so that allowances can be made for the difference in dose-rate and radiation quality between gamma and beta-emitting radionuclides. PMID- 11272360 TI - The American Brachytherapy Society perspective on intravascular brachytherapy. AB - BACKGROUND: Recent clinical studies indicate that intravascular brachytherapy (IVB) can reduce the rate of restenosis substantially after angioplasty procedures. However, no clinical guidelines exist for optimal therapy. METHODS: The members of the IVB Subcommittee of the American Brachytherapy Society (ABS) identified the areas of consensus and controversies in IVB to issue the ABS perspective on IVB, based on analysis of published reports and the clinical experience of the members in brachytherapy. RESULTS: IVB is still experimental. The long-term efficacy, toxicity, the target tissue, and dose required for IVB are not established. The ABS recommends that IVB procedures must be performed, with careful attention to radiation-related issues, in the context of controlled multidisciplinary clinical trials with the approval of the institutional review board, the Nuclear Regulatory Commission, the Food and Drug Administration, and under an Investigational Device Exemption. The therapeutic radiologist, with a qualified radiation physicist, is responsible for dose prescription and delivery and needs to be present during the IVB procedure as part of this multidisciplinary team. The long-term outcome from these studies should be reviewed critically and published in peer-reviewed journals. The ABS endorsed the dosimetric guidelines of the American Association of Physicists in Medicine Task Group 60 (AAPM TG-60) report. The ABS recommends that dose specification be defined clearly; to allow comparisons between studies, the dose should be prescribed at 2 mm from the source for intracoronary brachytherapy and at an average luminal radius of +2 mm for peripheral vascular brachytherapy. The prescription doses at the above point is generally in the 12-18 Gy range. Comprehensive procedures for quality assurance, radiation protection, and emergencies should be in place before initiating an IVB program. Higher energy beta sources, lower energy gamma sources, dose-volume histograms, and correlation of three-dimensional reconstructions of delivered dose with patterns of failure are areas for further research. CONCLUSION: The ABS perspective on IVB is presented to assist the interventional team in developing protocols for the use of IVB in the prevention of restenosis. Long-term outcome data with a standardized reporting system are needed to establish the role of brachytherapy in preventing vascular restenosis. Endovascular brachytherapy is a new and evolving modality, and these recommendations are subject to modifications as new data become available. PMID- 11272361 TI - A dosimetric comparison of conventional vs conformal external beam irradiation of a stented coronary artery utilizing a new fluoroscopic imaging detector system. AB - Purpose. To determine whether conformal external photon beam irradiation may prevent or reduce the rate of restenosis of a stented coronary artery following percutaneous transluminal coronary angioplasty (PTCA). Optimal conformal external beam irradiation with limited cardiac dose requires adequate visualization of the stented vascular segment. With existing image intensifiers, identification of a coronary stent is poorly localized. We propose using an amorphous silicon panel detector to observe the movement of the stent during the cardiac cycle. BACKGROUND: Long-term radiation-induced coronary complications can be minimized by: (a) reducing the radiation field sizes, (b) fractionating the total dose over several days, and (c) applying multiple treatment beams. Localization of the movement of the stent during the cardiac cycle may allow for the design of radiation fields that conform to the stented vessel segment. This scheme may permit gating the radiation beam on or off relative to movement of the stent within or outside the radiation fields, respectively. METHODS: Using a new solid state amorphous silicon planar detector, with a dynamic range of 12 bits, fluoroscopic images of a Palmaz-Schatz coronary stent were obtained. The stent was centered in a polystyrene phantom 20 cm thick and imaged using a 90-kVp, 3.5 ma, source-detector and source stent distances of 114 and 100 cm, respectively. With the solid-state silicon detector, the stent was identified in a single video frame (1/30 s). This fast image acquisition should allow for mapping the motion of the stent during the cardiac cycle. The stent movement during the cardiac cycle may then be correlated with the QRS complex in the electrocardiogram. CONCLUSIONS: The localization of a coronary stent during the cardiac cycle under fluoroscopy permits delivery of small conformal external radiation fields to treat stented coronary arteries, while minimizing radiation dose to surrounding normal cardiac tissue and vasculature. The best selection of treatment beam angles will be provided by high resolution fluoroscopic images of the stented region obtained from different beam directions. The three-dimensional movement of the stent, indexed in time with the QRS complex, will provide an important measure for gating radiation beams for conformal treatment delivery. PMID- 11272362 TI - Endovascular beta irradiation for prevention of restenosis using solution radioisotopes: pharmacologic and dosimetric properties of rhenium-188 compounds. AB - PURPOSE: Irradiation of the arterial wall with beta particles has been shown to be effective in inhibiting neointimal hyperplasia following percutaneous transluminal coronary angioplasty (PTCA). In this study, we describe the use of 188W/188Re generators to obtain 188Re (half-life 16.9 h, maximal beta energy of 2.12 MeV) as a new candidate radioisotope for endovascular irradiation. We have evaluated two [188Re]-compounds as candidates for use as solution-based radiation sources that would allow conventional liquid-filled balloon inflation for delivery of radiation to the vessel wall. While balloon rupture at nominal inflation pressures is a very rare event, (<1 per 10,000 at high pressure), radioisotope release could potentially result in significant dose to radiation sensitive organs. We have thus evaluated the biodistribution, dosimetry, and kinetics of excretion in rats of two 188Re-labeled compounds that are proposed for intravascular therapy. MATERIALS AND METHODS: Rhenium-188 was obtained as [188Re]-sodium perrhenate by saline elution of an alumina-based 188W/188Re generator system (>500 mCi). High specific volume solutions of the [188Re]-sodium perrhenate (>50 mCi/ml) were obtained by post-elution concentration of the generator bolus by passage through a tandem silver cation/anion column system. Rhenium-188-labeled benzoylthioacetyltriglycine (MAG3) was prepared by stannous ion reduction of [188Re]-perrhenate in the presence of the benzyl-MAG3 substrate, and was characterized as a single radioactive component. Rhenium-188-perrhenate and [188Re]-MAG3 were administered to separate groups of Fischer rats, which were sacrificed at various times and the tissue distribution of 88Re determined in the major organs. Excretory products were also collected daily from separate groups of rats for each agent over 7 days. The effects of perchlorate and iodide preblocking and postdisplacement of thyroid uptake of [188Re]-perrhenate were also evaluated. RESULTS: Organ uptake values were modest for both agents [<0.25 % injected dose(ID)/gram of tissue at 6 h] for all organs evaluated except for the thyroid, with the intestines and intestinal contents showing the highest uptake values (0.72-1.97 %ID/gram). Whereas thyroid uptake of 188Re after injection of [188Re]-MAG3 was low (0.16 %ID/gram), uptake after injection of [188Re] perrhenate was higher and could be blocked by pretreatment with perchlorate (intravenous [IV]) or displaced by perchlorate posttreatment. Also, oral or IV iodide pre- or postadministration could also significantly block or displace thyroid uptake of [188Re]-perrhenate. Both [188Re] agents were excreted primarily via the urinary bladder. The excretion half-life of [188Re]-perrhenate was about 7 h; in contrast, the [188Re]-MAG3 complex showed 50% excretion in less than 2 h. The large intestines received the most significant adsorbed dose, with values of 2.0 cGy/ mCi for [188Re]-perrhenate and 4.6 x 10(-3) cGy/mCi for [188Re]-MAG3. CONCLUSIONS: Rhenium-188-MAG3 shows more rapid urinary bladder excretion in rats than perrhenate and both agents show low organ uptake. Thyroid uptake of free [188Re]-perrhenate can be blocked or displaced with oral perchlorate administration. For the projected use of [188Re]-MAG3 for balloon inflation required for irradiation of the arterial wall, calculated organ dose values are within acceptable limits in the unlikely event of low pressure balloon rupture. Rhenium-188-MAG3 in solution is thus a new candidate for balloon dilation providing uniform endovascular irradiation following PTCA for restenosis therapy. PMID- 11272363 TI - Pathology of radiation-induced coronary artery disease in human and pig. AB - PURPOSE: External beam mediastinal radiation-induced accelerated coronary atherosclerotic heart disease in humans has been recognized, especially when the condition occurs in young persons. The purpose of the present study was to compare external beam radiation-induced accelerated coronary atherosclerosis in humans with that seen in the pig coronary arteries following radioactive stent placement. METHODS: A literature review of radiation-induced coronary artery disease was performed. In addition, clinical records and coronary histology from the Armed Forces Institute of Pathology Registry were reviewed from patients who had received external beam radiation for mediastinal malignancies. Coronary arteries from pigs that had radioactive coronary stent placement were evaluated from our stent pathology laboratory and analyzed for comparison with the human disease. RESULTS: In humans, the characteristics of the intimal plaque in accelerated atherosclerosis postradiation therapy were similar to that seen in typical atherosclerotic coronary disease in the absence of radiation therapy. However, medial thinning and adventitial fibrosis were the distinguishing pathologic arterial changes secondary to radiation seen in humans. Radioactive stent placement in pig coronary arteries produced similar changes to that observed in humans post-external beam radiation, consisting of medial injury and adventitial thickening accompanied by intimal foam cell collections, calcification, and necrotic core formation containing cholesterol clefts resulting in severe luminal narrowing. CONCLUSIONS: Radiation, delivered via external beam or radioactive stent, induces changes of intimal atherosclerosis with medial thinning and adventitial scarring in human and pig. Therefore, pending completion of long-term clinical studies, caution should be exercised before the widespread use of brachytherapy is advocated for the treatment and prevention of coronary restenosis. PMID- 11272364 TI - Clinical experience with a spiral balloon centering catheter for the delivery of intracoronary radiation therapy. AB - PURPOSE: To develop and evaluate an intravascular radiation delivery catheter that incorporates a centering mechanism and that allows side branch and distal artery perfusion. METHODS AND MATERIALS: The Galileo Centering Catheter (Guidant Vascular Interventions, Houston, TX) incorporates a rapid exchange tip design. A unique spiral balloon allows centering and facilitates perfusion to the distal artery and side branches. The catheter contains a dedicated dead-end lumen for source wire delivery to the lesion site. The treatment area is precisely defined by radiopaque markers. RESULTS: In three clinical trials to date, radiation (or placebo) was delivered successfully to 300 of 312 patients (96%). With balloon inflation, TIMI grade 2 or 3 flow was achieved in side branches in 82% and in the distal artery in 77% of patients. Despite treatment (dwell) times ranging from 87 to 948 s (mean = 250 s), only 8% of patients required fractionation of treatment. CONCLUSION: The Galileo Centering Catheter is a safe and highly effective method for delivering intracoronary radiation therapy. Its unique design provides centering of the source while allowing side branch and distal coronary perfusion during treatment. This catheter would facilitate intracoronary radiation therapy and allow uniform and reproducible dose delivery to the target in the artery wall. PMID- 11272365 TI - Progress in clinical trials for coronary arterial restenosis using beta radiation sources. AB - Vascular brachytherapy has yet taken the leap into progression from emerging technology to standard of care dependent on the outcome of clinical trials. Data collected from early pilot trials and on-going clinical trials indicate that such a progression is achievable. Three-year follow-up data from patients treated with intracoronary radiation for the prevention of restenosis are now available. Data from larger trials are being assessed using angiographic and intravascular ultrasound analysis. The beta radiation studies are demonstrating different levels of efficacy, raising new issues regarding dosimetry and potential complications. Past trials have examined the use of vascular brachytherapy for recurrence prevention of restenosis in patients with in-stent restenosis. New data related to the use of liquid-filled balloon systems and radioactive stent are also being collected. This article updates the current status of clinical trials in vascular brachytherapy utilizing beta emitters, highlighting preliminary results and assessing their implications for the development of this field. PMID- 11272366 TI - Theoretical assessment of dose-rate effect in endovascular brachytherapy. AB - PURPOSE: Several prospective randomized clinical trials utilizing endovascular brachytherapy after coronary angioplasty have shown promising preliminary results. Numerous clinical trials have been initiated to evaluate different delivery systems and source types. In this study, the dose-rate effect is investigated using a biophysical model derived from linear-quadratic formalism. MATERIALS AND METHODS: The dose-rate effect is quantified using the Dale's formulation, which is based on a linear-quadratic model. This model converts the total absorbed dose into the biological equivalent dose (BED) based on the dose rate, total dose, treatment duration, biological endpoint (alpha/beta ratio), and sublethal damage repair constant. The calculations are performed for two common source configurations used in current clinical trials (192Ir and 90Sr/Y). RESULTS: At smaller radial distance, the dose rate is higher, hence BED increases due to the increase in the relative effectiveness per unit dose (RE) and absorbed dose for a given treatment duration. For 90Sr/Y source, a similar trend is observed; however, it is at a much greater magnitude. The RE for 192Ir is close to unity, which is equivalent to that of external beam irradiation. CONCLUSION: Although current clinical trials in endovascular brachytherapy report similar absorbed dose, the biological effects may be different due to the extremely high gradient of dose rate near the sources, a variety of isotopes and delivery systems, and different dose prescriptions. If the theoretical predictions in this study are validated in clinical trials, the proposed model can be useful to compare different protocols, design new delivery systems and isotopes, and optimize how radiation is delivered. PMID- 11272367 TI - Theoretical assessment of late cardiac complication from endovascular brachytherapy for restenosis prevention. AB - PURPOSE: In this study, a theoretical assessment of late cardiac complication from endovascular brachytherapy is performed using the integrated logistic model. MATERIALS AND METHODS: Calculation were performed for various lengths of Ir-192 sources using alpha/beta = 3.2 for the endpoint of chronic ischemia, TD50/5 = 7,000 cGy, and TD5/5 = 5,000 cGy. The dose distribution over a standard heart was divided into volume elements with uniform dose (dose-volume histogram). Using linear-quadratic equation, the dose in each of the volume elements was converted into dose equivalent to standard fractionation external beam irradiation. The normal tissue complication probability (NTCP) for each volume element was calculated and combined together to arrive at the cumulative risk of late cardiac complication. The NTCP was plotted against the dose prescribed at 2-mm radial distance for four treatment lengths. RESULTS: (1) The overall risk of late cardiac toxicity (chronic ischemia within 5 years) was estimated to be less than 1% for current clinical trials using Ir-192. (2) There is a volume effect with higher risk for larger irradiated volume, which can come from longer treatment time, the same dose prescribed at a greater radial distance, and a longer source train. (3) The NTCP vs. dose demonstrates a sigmoidal relationship. There is a threshold dose (about 500 cGy), below which the risk is minimal; the gradient of the curve is greater for longer treatment length. CONCLUSION: If the prediction from this model is validated with clinical data, it will contribute to guidelines for dose prescription, dose escalation, evaluation of new source design, and multivessel treatment. PMID- 11272368 TI - Effects of a positron-emitting vanadium-48 nitinol stent on experimental restenosis in porcine coronary arteries: an injury-response study. AB - BACKGROUND: The major limitation of coronary stenting is restenosis due to exaggerated neointimal thickening. We evaluated a positron-emitting V48 nitinol stent in a porcine coronary model of restenosis. METHODS AND RESULTS: Pigs (n = 16) received a control nonradioactive and a V48 stent (1.5 or 10.6 muCi) randomized to the left anterior descending artery (LAD) and right coronary artery (RCA). Histology, morphometric variables, and strut injury scores were evaluated after 32 days. Peristrut fibrinoid deposits were greater in the high-dose group (p < 0.0001). Control stent area stenosis (AS) and mean neointimal thickness (NIT) correlated with injury (r = 0.81 and 0.79, respectively). Higher-dose stents reduced AS by 20% (0.57 +/- 0.13 vs. 0.71 +/- 0.16; p = 0.029) and mean NIT by 35% (0.44 +/- 0.16 vs. 0.71 +/- 0.24mm; p = 0.001) compared with controls. Lower-dose 1.5-muCi stents did not differ from controls. NIT over individual struts was reduced in the high-dose group compared with controls by 0.18 mm for grade 1 injury, 0.31 mm for grade 2, and 0.38 mm for grade 3 (p < 0.02 for all comparisons). CONCLUSIONS: 1.5-muCi V48 nitinol stents did not influence vessel histology or restenotic parameters in pig coronary arteries. In contrast, 10.6 muCi stents created a distinctive histological picture consisting of increased fibrinoid deposits on the neointimal-facing side of the struts without cellular organization. Higher dose radioactive stents significantly reduced AS and mean NIT. The reduction in neointimal thickening was greatest when the depth of strut penetration into the vascular wall was most severe. PMID- 11272370 TI - Effectiveness of fractionated external beam radiation in the inhibition of vascular restenosis. AB - BACKGROUND: From the clinical oncologic experience, fractionation of the radiation dose offers a better therapeutic window, both with respect to effectiveness and unwanted side effects. Thus, we undertook a pilot study in a rodent model, using a single dose of 15 Gy compared with fractionation schedules of 5 or 10 daily applications of 3 Gy. MATERIALS AND METHODS: Using a previously described rat angioplasty model, animals were randomly assigned to one of four groups: unilateral balloon injury, sham irradiation; unilateral balloon injury, bilateral 15 Gy single dose irradiation; unilateral balloon injury, bilateral 3 Gy x 5 daily fractions; or unilateral balloon injury, bilateral 3 Gy x 10 daily fractions. RESULTS AND CONCLUSIONS: All three radiation groups demonstrated a clear inhibition of neointimal hyperplasia. We therefore offer evidence for the effectiveness of fractionated radiation as a means to inhibit vascular restenosis in a rat carotid model. However, the 3 Gy x 5 schedule was less effective than either the 3 Gy x 10 schedule or the 15 Gy single dose. This study must be repeated using longer time points to provide proof of principle. PMID- 11272369 TI - Radioactive beta-emitting solution-filled balloon treatment prevents porcine coronary restenosis. AB - PURPOSE: Intracoronary gamma or beta radiation from centrally located sources at the time of overstretch balloon injury inhibits neointimal proliferation. In an effort to deliver homogeneous, centered radiation fields in a technically straightforward fashion, we studied the effects of a beta-emitting solution used as a balloon inflation fluid to deliver radiation at the time of coronary injury. METHODS: Twenty-one coronary arteries in 13 juvenile swine underwent irradiation (control and 11 or 25 Gy media dose). Radiation was delivered using a perfusion balloon inflated with an Re-188 solution. Subsequently, overdilatation percutaneous transluminal coronary angioplasty was performed at the pretreated segment. Histopathologic and histomorphometric analysis was performed at 30 days after injury on the entire irradiated artery. RESULTS: Balloon overdilation was associated with significant vascular injury and marked neointimal proliferation in control and low-dose (11 Gy)-treated arteries. High-dose radiation (25 Gy) significantly inhibited neointima formation compared with controls (neointimal area: 0.49 +/- 0.29 mm2 vs. 1.51 +/- 0.22 mm2, respectively; p = 0.02) and low dose radiation (neointimal area 1.75 +/- 0.54 mm2, p > 0.1 compared with controls). CONCLUSIONS: Liquid Re-188 is an effective beta-emitting vehicle to deliver intracoronary radiation and prevent restenosis in this model. Intracoronary radiation treatment using aqueous radioisotope sources is technically straightforward and provides the optimally achievable radiation dose distribution. PMID- 11272371 TI - Augmentation of the expression of proangiogenic genes in cardiomyocytes with low dose laser irradiation in vitro. AB - BACKGROUND AND OBJECTIVE: Several reports suggest that low power red laser light (LPRLL) is capable of affecting cellular processes in the absence of significant thermal effect. The objective of the present study was to determine the effect of LPRLL on proliferation of fetal cardiomyocytes in vitro and on the expression of proangiogenic genes, transforming growth factor-beta (TGF-beta), and vascular endothelial growth factor (VEGF). STUDY DESIGN/MATERIALS AND METHODS: All cell cultures were irradiated with single-dose LPRLL using a He-Ne continuous wave laser (632 nm) with different doses. The effect of LPRLL on new DNA synthesis was studied by 3H thymidine-incorporation assay. VEGF and TGF-beta expression by cardiomyocytes was studied by reverse transcription-polymerase chain reaction (RT PCR). RESULTS: We observed that a dose-dependent increase in cardiomyocytes proliferation can be obtained with LPRLL and that there is a significant increase in VEGF and TGF-beta mRNA expression by cardiomyocytes. CONCLUSIONS: These data may have significant importance leading to the establishment of new methods for myocardial photoangiogenesis and photoregeneration as well as in vitro proliferation of cardiac myocytes. PMID- 11272372 TI - Methods to improve dose uniformity for radioactive stents in endovascular brachytherapy. AB - Intravascular brachytherapy (IVBT) has rapidly gained acceptance as a new treatment modality for reducing restenosis and improving the success rate of percutaneous transluminal coronary angioplasty (PTCA). Recent clinical results on patients treated with beta-emitting 32P stents suggest that radiation reduces in stent restenosis but may exacerbate neointimal growth at the edges of the stents. This has been referred to as the "candy wrapper effect." It is well known that radioactive stents yield extremely inhomogeneous dose distributions, with low doses delivered to tissues in between stent struts, at the ends of the stent, and also at depth. Some animal model studies suggest that low doses of radiation may stimulate rather than inhibit neointimal growth in an injured vessel, and it is hypothesized that dose inhomogeneity at the ends of a stent may contribute to the candy wrapper effect. We present here a theoretical study comparing dose distributions for beta stents vs. gamma stents; "dumbbell" radioactive loaded stents vs. uniformly loaded stents; and stents with alternate strut design. Calculations demonstrate that dose inhomogenieties between stent struts, at the ends of stents, and at depth can be reduced by better stent design and isotope selection. Prior to the introduction of radioactive stents, criteria for stent design included factors such as trackability, flexibility, strength, etc. We show here that if stent design also includes criteria for strut shape and spacing that improved dose distributions are possible, which in turn could reduce the candy wrapper effect. PMID- 11272373 TI - Low-energy 103Pd gamma (X-ray) source for vascular brachytherapy. AB - PURPOSE: This article describes the merits of 103Pd, a low energy x-ray emitter, as a new and potentially superior candidate for intraluminal brachytherapy. 103Pd can be ion implanted into different materials, designs, and devices for vascular brachytherapy. METHODS: The mass-analyzed ion implantation process has been used to embed the desired activity of 103Pd into the surface of 316L stainless steel stents. The low-energy, 21-keV x-ray from 103Pd delivers reasonably homogeneous radiation to the vessel wall and loses intensity rapidly beyond the immediate vicinity of the source. Shielding for 103Pd x-rays is trivial and the issues regarding safety, health hazards, storage, and handling are easily manageable. RESULTS: Experimental data on 103Pd ion-implanted stents show a clean gamma-ray spectrum devoid of any radioimpurity. Activity measurements within a batch demonstrate little stent-to-stent variation (approximately 2%), excellent axial and radial uniformity (<10%), and minimal dissolution in a saline environment (approximately 0.02%). The dosimetry shows the focused radiation field for 103Pd stents and rapid fall-off beyond a few millimeters of the stent. Furthermore, the 103Pd dosimetry indicates that a 350 muCi stent will deliver approximately 14 Gy at 1-mm distance, over its lifetime. CONCLUSION: 103Pd is an appropriate source for vascular brachytherapy. It has an appropriate combination of half-life (16.93 days) and energy (21 keV). The half-life of 103Pd delivers the dose with an acceptable dose rate for stent applications while, at the same time, allowing for manageable shipping, storage, and/or disposal. PMID- 11272374 TI - Clinical review: irradiation for lower extremity arterial occlusive disease. AB - Lower extremity atherosclerosis, a disease of aging, is both widespread and increasing in prevalence-it is estimated that almost 100,000 patients per year in the United States require operative bypass for lower extremity ischemia. It is an axiom of vascular surgery that essentially every bypass graft will eventually fail. Many if not most such failures are due to the process of intimal hyperplasia at one or both anastomoses. The search for a "cure" for intimal hyperplasia has been long, but thus far unrewarding. Recent advances in therapeutic irradiation, however, offer a potential solution to this problem. This review is designed to acquaint the radiation oncologist with the basic concepts behind lower extremity atherosclerosis and its treatment, and to introduce briefly the special problems inherent in considering irradiation of an end-to-side anastomosis. PMID- 11272375 TI - Radiation safety requirements for cardiovascular brachytherapy. AB - Cardiovascular brachytherapy, the use of high intensity radiation to inhibit the growth of neointimal tissue after coronary revascularization by either balloon angioplasty or other methods is being tested in a number of clinical trials to assess the efficacy of the treatment. This new use of radiation to aleviate the suffering of individuals with coronary artery disease has excited many interventionalists and has caused others to view the new technique with skepticism. There are a number of operational and safety concerns to face in incorporating this treatment modality into the cardiac catheterization laboratory. Delivering the radiation dose to the patient with a minimum of radiation exposure to both patient and operating personnel requires close attention to the physical characteristics of the radiation source as well as the administrative and regulatory requirements imposed on the facility by federal and state regulators. The insertion of the source into the proper artery and location is the task of the cardiologist in collaboration with the radiation oncologist. The determination of the appropriate radiation dose is the responsibility of the medical physicist. The safe handling of the radioisotope source is the responsibility of the radiation safety specialist. State and federal regulations dictate minimum requirements of safety in the handling of radioactive sources used in cardiovascular brachytherapy. These requirements involve close monitoring of the patient and operating personnel to insure that radiation exposures are minimized. They involve the restricted access of nonessential personnel to the cath lab during the treatment. The entrances to the cath lab must be monitored to prevent unauthorized entry. Operating personnel must be closely monitored to maintain radiation exposures as low as reasonably achievable (ALARA). The patient must be monitored to insure that the source is implanted for the prescribed time and the patient's exposure is also ALARA consistent with the medical benefit expected. Public corridors must be monitored to prevent public exposures to the radiation emanating from the patient. The radiation exposure field around the patient during a typical gamma treatment presents what the regulators define as a high radiation area. This means that the exposure levels are in excess of 100 milli-rem (mrem) per hour at 30 cm from the patient. In fact, the exposure levels around the patient for a typical treatment are in the roentgens per hour range. The use of beta particle emitting radionuclides (Sr90/Y90 and P32) presents a much lower safety problem. But the use of radioactive materials in the cath lab still presents new safety concerns such as training, monitoring, record keeping, and public relations among the cath lab technologists. PMID- 11272376 TI - Delayed healing and increased thrombosis following intracoronary radiation in balloon injured porcine coronary arteries. PMID- 11272378 TI - Innovations and challenges to intravascular brachytherapy dosimetry. PMID- 11272377 TI - Assessing the advantages and disadvantages of novel radiation therapy for vascular restenosis: injury score, artery size, and short lengths. PMID- 11272379 TI - The future of complementary and alternative medicine. PMID- 11272380 TI - Incorporating new (mind/body, alternative, complementary, or integrative) medicine into everyday care. AB - Use of a variety of therapies and techniques--ranging from acupuncture to yoga to herbal therapies--that are designed to relieve medical conditions and illnesses and/or emphasize the mind, body, and spirit connections are gaining popularity among patients in the United States. For years, many hospitals, plans, clinicians, and insurers ignored these therapies when using "conventional" therapies. But, times are changing: A movement is now afoot to determine whether these therapies and techniques can be successfully integrated with current health care treatments to provide quality care to patients. PMID- 11272382 TI - [New aspects in managing mandibular fractures]. AB - BACKGROUND: The aim of osteosynthetic mandibular fracture treatment is a bony bridging of the fracture gap. The gap distance and the fragment movement determine the micromovement in the gap tissue. They are known to be the main factors associated with the clinical outcome of fracture treatment. LITERATURE: The term fracture stability and the underlying mechanically modulated tissue reactions are not well described in the literature. MODEL: In this article we describe experimental results of strain related bone regenerate reactions in vivo. Additionally, in an in vitro model of osteosynthetic fracture treatment micromovements in the sense of bone strains were determined in the gap area. Implications for the osteosynthetic treatment of mandibular fractures are discussed on the basis of our biological and biomechanical results. PMID- 11272381 TI - [In vivo studies of screw-bone contact of drill-free screws and conventional self tapping screws]. AB - Screw-bone contact (SBC) and bone remodeling of titanium drill-free screws or self-tapping screws should be compared. Each 10 titanium self-tapping miniscrews or microscrews, and each 10 titanium drill-free miniscrews or microscrews were inserted into the anterior wall of the frontal sinus of 5 Gottingen minipigs. Intraperitoneal injections of fluochromes (Xylenol, Calcein, Alizarincomplexon and Tetrazyklin) were performed between the 2nd and 9th postoperative week. The pigs were sacrificed after 6 months, the screw-bone blocks were resected and microradiographic, histologic and fluorescence microscopic examinations were carried out. In drill-free screws mean SBC was 88.4 (miniscrews) or 93.8% (microscrews). In self-tapping miniscrews mean SBC was 54.9, in microscrews 81%; the differences were significant in statistical analysis (t-test: p < 0.05). In fluorescence microscopy the amount of bone remodeling (ratio of residual versus newly formed bone) was measured. Significantly more of the residual bone was found in the region of the screw threads of drill-free screws (miniscrews: mean 71.8, microscrews: mean 67.9%) than in the region of screw threads of self tapping screws (miniscrews: mean 33.1, microscrews: mean 42.4%; t-test: p < 0.05). The superior SBC of drill free screws in comparison to SBC of self-tapping screws and the higher amount of original bone in the threads of drill free screws explain the sufficient holding strength of drill free screws in clinical tests. Both results are important for osteosynthesis in regions where thin cortical bone is present, as in the central midface. PMID- 11272383 TI - [CT-computer-template-assisted planning of implant and magnet position in epi prosthetic management of facial defects]. AB - BACKGROUND: The aesthetic result of a prosthetic reconstruction in the facial area depends on precise, long-term stable positioning of the facial prosthesis. For fixation of the facial prosthesis and contouring of the soft tissue, good long-term success can be achieved by implant-magnet systems. The precise positioning of the implants and magnets when only little bone is available is difficult and without any planning is often not possible. COMPUTER EVALUATION: Computer-supported evaluation of CT data with the aid of CT and drill templates can optimise the planning and carrying out of surgery. This procedure is illustrated with reference to the treatment of a patient following obital exenteration due to an embryonic rhabdomyosarcoma. The programs SIM/Plant and coDiagnostiX were compared with one another when carrying out the CT evaluation and planning the orbital prosthesis. In so doing, the great advantage of the definition of the CT window (grey levels), the freely definable panorama sections and cross-sections, as well as the three-dimensional, spatial representation using the programme coDiagnostiX on a conventional PC are made clear. The results of planning can be applied and implemented during surgery, with the aid of the CT and drill templates. DISCUSSION: The CT computer template-supported procedure is suitable for routine clinical use and can be recommended for planning of implant fixed facial prosthesis in the orbital area when little bone is available or in the case of difficult anatomical relationships. PMID- 11272384 TI - [Risks and complications of membrane-guided bone regeneration. Retrospective analysis]. AB - AIM OF THE STUDY: The aim of this retrospective study was to determine the number of premature membrane exposures and to evaluate possible predisposing factors. PATIENTS AND METHODS: A total of 72 patients were treated with ePTFE membranes (GoreTex Augmentation Material) for membrane-guided bone regeneration (GBR) of periimplant bone defects (median age 38.5 years, minimum 18 years, maximum 68 years; 39% male and 61% female patients, data collection between 03/1993 and 02/2000). Sixty-one percent of the membranes were applied in the upper jaw and 39% in the lower jaw; 61% of all GBR procedures were performed in conjunction with single tooth replacements. RESULTS: Premature membrane exposure (PME) was found at 44% of the sites. Of these PMEs 72% occurred after a primary, uneventful healing period subsequent to membrane implantation. Statistical analysis revealed a significant correlation between the parameters "premature membrane exposure", "membrane covering the alveolar ridge" (P = 0.0004) and "membrane located beneath the incision line" (P = 0.0004). Additionally a tendency for correlation was calculated for the parameters "defect configuration" (P = 0.08) and "smoking" (P = 0.04). CONCLUSIONS: The defect morphology, the flap and incision design and the smoking habits of the patients have to be carefully considered if guided bone regeneration is planned. Non-space-maintaining defect configurations, smoking habits and crestal incisions crossing the planned membrane location seem to be contraindications for the use of ePTFE membranes. In such cases, alternative treatment methods should be considered. PMID- 11272385 TI - [Frey syndrome]. AB - AIMS: The incidence of Frey's syndrome after parotidectomy as cited in the literature varies distinctively. Strategies for successful treatment are also assessed differently. PATIENTS: Between 1980 and 1994 a total of 372 parotidectomies were performed in 364 patients at the Bochum University Hospital. RESULTS: After an average of 18 months following parotidectomy, 86 patients (23.5%) developed Frey's syndrome. Thirty-five patients were treated with scopolamine ointment. The symptoms improved in nine cases after an average of 25 months of therapy. Of the patients receiving no treatment (n = 20), seven improved after an average follow-up of 20 months. Therapy with scopolamine ointment did not elicit significantly better results compared to no treatment at all. CONCLUSION: Gustatory sweating after parotidectomy still has to be regarded as an unpleasant complication which is difficult to cure. PMID- 11272386 TI - [Detection of p53 mutation in mouth mucosa smears of patients with oral squamous epithelial carcinoma]. AB - Tumour-suppressor gene p53 encodes for an important cell-cycle regulatory protein and is therefore probably important for the development of many malignant diseases, e.g. squamous cell carcinoma of the mouth. This gene has mutated most frequently in connection with the development of cancer, so it has been well explored. Hence we chose it to find out whether swabs of the oral mucosa are suitable for supplying material for the detection of mutations in a gene that is connected with the development of oral squamous cell carcinomas, because swabs are easier to obtain than biopsies. We examined biopsies, swabs from the tumour, and swabs from mucosa that appeared healthy from 32 patients with oral squamous cell carcinoma and mucosal swabs from 35 healthy persons with polymerase chain reaction (PCR) and temperature gradient gel electrophoresis (TGGE). Fourteen of the 32 patients with a tumour showed mutations of p53, and in all cases the mutation could be demonstrated both in the biopsy and in the tumour swab. In four cases the mutation was also found in the swab of normal mucosa. Our investigations revealed that swabs are a suitable method for obtaining material for the detection of gene mutations in oral squamous cell carcinomas. PMID- 11272387 TI - [Forensic evaluation of injuries to nerves and jaw bone after wisdom tooth extraction from the viewpoint of current jurisprudence]. AB - The removal of lower wisdom teeth is one of the most frequent operations in oral surgery. Iatrogenic lesions of the lingual and inferior alveolar nerve and fractures of the lower jaw are rare, but can lead to actions for damage and compensation for personal suffering. The aim of our investigation was to elaborate recommendations for the oral surgeon from the legal point of view. Therefore we investigated in two data banks all court decisions concerning wisdom teeth and analysed them in regard to the obligation to provide information, documentation and the minimum requirements of the surgical treatment. In 57% of all court decisions the obligation to provide information was omitted by the surgeon. Surprisingly, regarding injuries of the lingual nerve we found different opinions from various law courts. In some former decisions only the fact of the damage was equivalent to a lack of care, while recently other courts had the opinion that an injury of the lingual nerve could also be caused in spite of careful treatment. With regard to lower jaw fracture all courts emphasized that the patient must be informed about the risk by the surgeon. Moreover, minimum requirements for the removal of wisdom teeth were given. In conclusion, the recent court decisions restrict the liability for complications but demand the observance of minimum requirements for the surgical treatment. Additionally, all courts stress that informed consent about the risk of nerve damage of jaw fracture must be obtained before the removal of wisdom teeth. PMID- 11272389 TI - [Peri-osseous intracranial translocation of titanium osteosynthesis plates and screws after fronto-orbital advancement]. AB - BACKGROUND: Perossoeus intracranial translocation or passive intracranial transmission of titanium osteosynthesis plates and screws in the growing skull following surgical craniosynostosis corrections, also referred to as the PIT effect, has been described in the literature since 1995. It is a phenomenon which has not received due attention until recently and is explained by appositional and resorptional remodeling processes in the growing skull. CASE REPORT AND DISCUSSION: An impressive case of the PIT effect with a total intracranial dislocation of titanium plates and screws is used to demonstrate the problems associated with this phenomenon and to discuss the few clinical case reports in the English-language literature. The obvious advantages of a resorbable material are pointed out; however, it is still uncertain as to whether the resorption process is fast enough to avoid the PIT effect if used clinically. PMID- 11272388 TI - [Reconstruction of the orbits with polylactate implants: animal experimental results after 12 months and clinical prospects]. AB - In earlier experiments healing of large orbital wall defects in sheep occurred undisturbed by osteoconductive bone growth along biodegradable membranes when there was no interference with additional bone grafts or titanium miniplate osteosynthesis. In this experiment similar bilateral defects were reconstructed with poly(L/DL 80/20) lactide implants using a microporous membrane 0.5 mm thick without further support on one side, an 0.25 mm microporous membrane supported by solid polylactide buttresses and stabilized by polylactide dowels on the opposite side. After 12 months we found a symmetrical reconstruction of the normal anatomy of the orbits in CT and X-ray examinations. In contrast, histologic investigations revealed massive foreign-body reactions around degrading buttress implants and dowels especially. Milder reactions occurred in some orbits along the membranes as well, in contrast to our earlier experiments with 4-month follow up. None of the implants had degraded completely 12 months after surgery. In our 12-month long-term survey, polylactide microporous membranes confirmed their osteoconductive potential in orbital wall reconstruction. Nevertheless, massive polylactic implants should not be considered for clinical application in the orbit because of significant late foreign-body reactions. PMID- 11272390 TI - [Calcium pyrophosphate arthropathy (pseudogout) of the temporomandibular joint]. AB - The temporomandibular joint is rarely affected by crystal deposition disease (gout and pseudogout). Only the temporomandibular joint was affected in a 69-year old patient with calcium pyrophosphate dihydrate deposition disease (pseudogout). This case is described here. Clinically presenting as painless swelling, computed tomography and magnetic resonance imaging showed signs of a chronic destructive arthritis. The evidence for calcium pyrophosphate dihydrate crystals was provided by histological examination. Diagnostic criteria, differential diagnosis and possibilities for treatment are discussed on the basis of our own experience and the literature. PMID- 11272391 TI - Stacking the shelves with data. PMID- 11272392 TI - Technology opens 'golden window'. PMID- 11272393 TI - HIPAA may boost technologies. PMID- 11272394 TI - A glimpse of a rosier future? PMID- 11272396 TI - Crunching data on the Web. PMID- 11272395 TI - On the hunt for new recruits. PMID- 11272397 TI - An ASP enables a slow approach. PMID- 11272398 TI - The devil's in the details. PMID- 11272399 TI - Medical errors reporting & prevention. Weathering the storm ahead. PMID- 11272400 TI - Getting around with hand-helds. PMID- 11272401 TI - Health care welcomes new 'CeOs'. PMID- 11272402 TI - Molecular approach to ayurveda. AB - In ayurvedic system of medicine, it is considered that a living system is made of panch-mahabuta, in the form of Vata, pitta and kapha at the physical level and satwa, raja and tama at the mental level. This covers the psychosomatic constitution and commonly known as the Tridosh theory. The imbalance in these body humours is the basic cause of any type of disease manifestation. Till date, several objective parameters have been proposed to monitor the level of these basic humours but none of them is complete. In this exercise, now it is proposed to consider free radical theory of diseases as one of the objective parameters. To be more specific, vata can be monitored in terms of membrane bound signal transduction, pitta as the process of phosphorylation and de-phosphorylation of different proteins (signalling moieties and enzymes) and kapha can be viewed as the degree of gene expression as protein synthesis. This can be correlated with the ojas of the body or total body defence mechanism. PMID- 11272403 TI - Integrins and disintegrins: the candidate molecular players in sperm-egg interaction. AB - Fertilization includes sperm-egg recognition, adhesion, binding, fusion and egg activation. Integrin (ITG) receptors which are adhesion molecules are expressed on mouse, hamster and human gametes and their potential ligands also have been identified. Role of ITGs during fertilization is supported by inhibition of sperm egg adhesion and/or fusion by means of anti-ITG mAbs, Arg-Gly-Asp (RGD) or disintegrin-like peptides. This review includes the current understanding of the molecular mechanism that regulates sperm-egg interaction and implications of this knowledge for assessing the fertility potential of men and immunocontraceptive method. PMID- 11272404 TI - Localization of estrogen and progesterone receptors in the endometrium of common marmosets Callithrix jacchus. AB - In the present study, changes in the immunohistochemical localization of endometrial estrogen receptor (ER) and progesterone receptor (PR) during various stages of the ovarian cyclicity in common marmoset, have been reported. Ovarian cyclicity was monitored by estimating plasma estradiol and progesterone. During the early follicular phase, weak ER immunolocalization was observed in the endometrial stroma. During the late follicular phase under the influence of rising estradiol levels, stromal ER localization was intense. During the luteal phase, ER localization was absent in the stroma indicating that high concentrations of progesterone suppressed ER. PR localization was not observed in the stroma during the early follicular phase, while weak staining was seen in the stroma during the late follicular phase. PR localization was maximum during the mid luteal phase. However in marmoset, endometrial ER and PR localization was restricted only to the stroma. This unique feature may be due to the characteristic reproductive profile of this nonmenstruating species and needs to be studied further. Thus it can be hypothesized that in the marmoset endometrium, steroid hormone mediated effects possibly occur directly in the stroma and are then transmitted to the epithelium by autocrine/paracrine action of growth factors and cytokines. PMID- 11272405 TI - Radiosensitization of a mouse melanoma by withaferin A: in vivo studies. AB - The radiosensitizing effect of a plant withanolide, withaferin A, on the B16F1 mouse melanoma was studied in vivo. Treatment of 100 mm3 tumours with 10 to 60 mg/kg withaferin A intraperitoneally produced a dose dependent increase in growth delay and volume doubling time. Injection of 30-50 mg/kg withaferin A, followed by 30 Gy local gamma irradiation, significantly enhanced the tumour response. No systemic or local adverse reactions were noted in these groups. The drug was most effective when injected intraperitoneally 1 h before irradiation. However, neither the individual agents nor their combination could produce any complete response (tumour cure). Melanoma is a relatively radioresistant tumour. The present results indicate that the radiation response of this tumour can be significantly enhanced by pretreatment with withaferin A. PMID- 11272406 TI - Isoprenoid pathway and free radical generation and damage in neuropsychiatric disorders. AB - Two substances which are products of the isoprenoid pathway, can participate in lipid peroxidation. One is digoxin, which by inhibiting membrane Na(+)-K+ ATPase, causes increase in intracellular Ca2+ and depletion of intracellular Mg2+, both effects contributing to increase in lipid peroxidation. Ubiquinone, another products of the pathway is a powerful membrane antioxidant and its deficiency can also result in defective electron transport and generation of reactive oxygen species. In view of this and also in the light of some preliminary reports on alteration in lipid peroxidation in neuropsychiatric disorders, a study was undertaken on the following aspects in some of these disorders (primary generalised epilepsy, schizophrenia, multiple sclerosis, Parkinson's disease and CNS glioma)--1) concentration of digoxin, ubiquinone, activity of HMG CoA reductase and RBC membrane Na(+)-K+ ATPase 2) activity of enzymes involved in free radical scavenging 3) parameters of lipid peroxidation and 4) antioxidant status. The result obtained indicates an increase in the concentration of digoxin and activity of HMG CoA reductase, decrease in ubiquinone levels and in the activity of membrane Na(+)-K+ ATPase. There is increased lipid peroxidation as evidenced from the increase in the concentration of MDA, conjugated dienes, hydroperoxides and NO with decreased antioxidant protection as indicated by decrease in ubiquinone, vit E and reduced glutathione in schizophrenia, Parkinson's disease and CNS glioma. The activity of enzymes involved in free radical scavenging like SOD, catalase, glutathione peroxidase and glutathione reductase is decreased in the above diseases. However, there is no evidence of any increase in lipid peroxidation in epilepsy or MS. The role of increased operation of the isoprenoid pathway as evidenced by alteration in the concentration of digoxin and ubiquinone in the generation of free radicals and protection against them in these disorders is discussed. PMID- 11272407 TI - Role of KATP channels in reduced antinociceptive effect of morphine in streptozotocin-induced diabetic mice. AB - The nociceptive effect was measured using withdrawal latency in tail flick test in mice rendered diabetic by administering streptozotocin (200 mg/kg, i.p.). The antinociceptive effect of morphine (4 and 8 mg/kg, s.c.) and cromakalim, a KATP channel opener, (0.3, 1 and 2 micrograms, i.c.v.) was significantly reduced in diabetic mice. Moreover, co-administration of cromakalim(0.3 microgram) did not alter the reduced antinociceptive effect of morphine(4 mg/kg) in diabetic mice. Spleenectomy in diabetic mice restored the decrease in antinociceptive effect of morphine and cromakalim. Multiple dose treatment with insulin to maintain euglycaemia for 3 days in diabetic mice prevented the decrease in antinociceptive effect of morphine and cromakalim. However, hyperglycaemic tyrode's buffer did not alter the pD2 value of morphine in isolated guinea pig ileum suggesting that hyperglycaemia does not interfere with mu receptor mediated responses in vitro. The results suggest that hyperglycaemia induced decrease in antinociceptive effect of morphine and cromakalim may be due to alteration in KATP channels. Some unknown factor from spleen in diabetic mice may be responsible for this alteration in KATP channels in diabetic mice. PMID- 11272408 TI - Acute hepatotoxicity of DDT: effect on glucocorticoid receptors and serum transcortin. AB - The hepatotoxic effect of 1,1 bis (p-chlorophenyl) 2,2,2 trichloroethane (DDT) treatment for 10 consecutive days has been examined in Wistar rats. DDT exposure increased relative liver weight, dose dependently, with a marked decrease of glycogen content and profound histological changes including cytoplasmic vacuolization, signs of necrosis and nuclear enlargement. The hepatomegaly induced by DDT (50 and 100 mg/kg body weight day-1) appeared not to be accompanied by a significant alteration of the hepatic glucocorticoid receptor concentration and affinity while, serum corticosteroid binding globulin level increased slightly with the lower dose of the pesticide. It is concluded that a short-term exposure to DDT did not lead to a status stress and, therefore, the hepatotoxic effect of organochlorine seemed not to be mediated by endogenous glucocorticoids. PMID- 11272410 TI - Stereological study of rat spleen following acute ethanol treatment. AB - To investigate the acute effect of ethanol (4 g/kg, i.p.) on spleen adult female Wistar rats were treated intraperitoneally with: a) ethanol (4 g/kg body wt), b) naltrexone (5 mg/kg body wt) followed 45 minutes later by ethanol (4 g/kg body wt) and c) naltrexone (5 mg/kg body wt) alone. Untreated and saline-treated rats were used as controls. Twenty hours after the ethanol treatment the animals were sacrificed and the spleens were removed. A piece of tissue from the central part of each organ was fixed in Bouin's solution. Paraffin sections were stained with hematoxylin-eosin and analysed using stereological measurements. The volume densities of the following tissue compartments: red pulp, white pulp (divided in follicles, periarterioral lymphatic sheath and marginal zone) and the connective tissue were determined. Stereological analysis also included parameters of follicles: the areal numerical density (the number of follicles per 1 mm2 of tissue section), the numerical density (the number of follicles per mm3 of tissue) and the mean follicle diameter. The immunoarchitecture of the spleen was preserved following acute ethanol treatment. Unlike other parameters that were unaffected, ethanol evoked a decrease in both volume density of follicle and the mean follicle diameter. Naltrexone pretreatment had no influence on ethanol induced changes. The data obtained indicate that a single dose of ethanol has a profound effect on rat spleen affecting the follicles, but the mechanism of its action remains to be elucidated. PMID- 11272409 TI - Effect of intraamniotic vitamin A on palatal closure of fetal rats. AB - On day 15 of gestation, intraamniotic vitamin A in a dose of 150 IU was administered to the fetal rats to examine its effect on palatal closure. Fetuses subjected to only amniocentesis acted as control for the study. The fetuses were recovered on day 19, 20 and 21, respectively. Vitamin A resulted in poor development of palatine shelves. There was no clear demarcation of the base and the free margins of the shelves were either rounded or blunted with poor attempt towards closure. In the vitamin A group, the incidence of cleft palate were similar in all three days while there was a gradual decline with increasing gestational age in the amniocentesis group. The results suggest that unlike amniocentesis, in vitamin A treated fetuses, there was no attempt towards a delayed closure of the palate. PMID- 11272411 TI - Chromosome banding studies in an Indian mullet: evidence of structural rearrangements from NOR locations. AB - C-, G- and NOR bands have been studied in the female sex of Rhinomugil corsula. (Mugilidae, Pisces) by deploying the conventional methodologies with suitable modifications of minor nature. The diploid metaphase complements contained 48 acrocentric chromosomes. The localization of C-band heterochromatin was found to be mostly at or near the centromeric regions of the acrocentric chromosomes. The G-type bands were not so well defined, but some of the G-banded chromosomes also contained C-bands. Interestingly, silver-positive NORs were found at the telomeric ends of five acrocentric chromosomes, including one homologous pair having NORs in both chromatids, while one chromosome showed NORs in both of its chromatids and the other two had only one NOR localized at one of its chromatids. This would suggest that one homologue of the second pair of NOR-bearing chromosomes possibly underwent a chromatid exchange with a non-NOR bearing chromosome. This is quite a unique situation not reported earlier in any species of fish., though some other form of NOR-polymorphism/heteromorphism has rarely been reported. Therefore, further exploration in natural populations of this species to examine the other sex and to verify if there also exists other chromosomally polymorphic races (in respect of NOR-polymorphism) of this species, would be rewarding. PMID- 11272412 TI - Resistance of legume seeds to the bruchid, Callosobruchus maculatus: metabolites relationship. AB - A study was initiated to categorize the seeds of various wild and cultivar legume varieties on the basis of their relative resistance to the bruchid, C. maculatus, and to correlate the important primary and secondary metabolites (non-protein anti-metabolites) in these seeds to the developmental parameters of the bruchid. In general, the wild seed varities showed greater amount of resistance to the bruchid attack when compared to that of the cultivar varieties. All the cultivar varieties studied showed higher amounts of primary metabolites, namely, proteins, carbohydrates, lipids and free amino acids thus showing a positive correlation between the primary metabolites content and the infestation rate. The wild varieties, however, showed significantly lower amounts of these primary metabolites and consequently a lower level of infestation. The non-protein anti metabolites such as total phenols, ortho- dihydroxy phenols and tannis were significantly lower in the cultivars. The wild varieties, in contrast, revealed higher amounts of these secondary metabolites showing a negative correlation between these secondary metabolites content and the infestation rate. The study revealed that these non-protein anti-metabolites are important in conferring resistance to the seeds. PMID- 11272413 TI - In vivo qualitative changes of 31P NMR in stressed maize roots vis-a-vis carbon substrate determining the degree of stress. AB - High resolution 31P nuclear magnetic resonance spectroscopy was used to investigate the changes in phosphate metabolism and intracellular pH in intact maize (Zea mays L) root segments to hyper osmotic shock. The results were compared with the happenings under field conditions, when the stress was given gradually. Effect of sugar substrate on adaptation of tissue to both kinds of situations was also studied. The hyper osmotic shock resulted in large vacuolar alkalinization and a decrease in pH across tonoplast membrane. There was gradual build up of phosphocholine and decrease in glucose 6P and UPDG levels. In gradual stress, the root segments were able to adapt to the stress and maintained pH gradient across tonoplast, with marginal alkalinization of vacuoles. The presence of sugar substrate reduced the impact of stress significantly, commensurate with the increased activity of plasmalemma H(+)-ATPase. The latter providing the driving force for uptake of organic molecules and ions required for osmoregulation. PMID- 11272414 TI - Comparison of protein profiles and enzymes in non-mycorrhizal and mycorrhizal roots of Pennisetum pedicellatum. AB - Pennisetum pedicellatum plants were inoculated with Glomus mosseae, G. aggregatum and Gigaspora margarita. There were both quantitative and qualitative changes in the protein pattern of inoculated plants. Gi. margarita induced increase in protein in the plants. Acid phosphatase, alkaline phosphatase, superoxide dismutase and chitinase activities were high at the beginning of infection, but declined as the infection advanced. Gi. margarita was an efficient fungus in enhancing enzyme activity and proteins in roots compared with G. mosseae and G. aggregatum. Protein profile revealed the presence of 12 peaks in mycorrhizal plants compared with 8 in nonmycorrhizal plants. PMID- 11272415 TI - Modulation of antigenicity of mycelial antigens during developmental cycle of Karnal bunt (Tilletia indica) of wheat. AB - Indirect enzyme linked immunosorbent assays (ELISA) were developed using polyclonal antibodies against soluble cytoplasmic (SCA) and insoluble cell wall antigens (ICWA) for monitoring modulation of mycelial antigens during growth cycle of T. indica. With SCA, continuous decrease in ELISA reactivity was observed in maturing fungus cultures, suggesting that SCA were expressed predominantly during early vegetative phase and their decreasing role was apparent as the fungus matures possibly towards sporogenous mycelium. In case of ICWA, the reaction profile showed an increase up to exponential phase of growth probably due to increase in the cell division and branching of mycelium. But later, ICWA antibody reactivity was decreased which may be due to conversion of mycelial phase to sporogenous phase, a quiescent stage of growth. Characterization of changes in antigenic configuration during developmental cycle of Tilletia indica by these antibodies could prove to be useful in identification of developmentally related and virulence marker(s). PMID- 11272416 TI - Agrobacterium mediated transformation of Vigna sesquipedalis Koern (asparagus bean). AB - Agrobacterium mediated transformation of Vigna sesquipedalis was achieved using cotyledonary node explants prepared from 5 days old seedlings germinated on B5 basal medium, and transformed using Agrobacterium tumefaciens strain EHA101, carrying the phosphinothricin-N-acetyltransferase gene and neomycin-3 phosphotransferase-II gene as selectable markers and GUS gene as a screenable marker. Gene transfer was achieved by inoculation of cotyledonary node explants with a bacterial suspension and a further cocultivation with Agrobacterium suspension for 3 days on B5 basal medium. Only 10% of the explants were transformed with EHA101 and exhibited transient expression of GUS genes, while 2% of shoots exhibited stable integration of genes and developed into plants. Transgenic character of tissues was confirmed by GUS assay and Southern analysis. Histological analysis of GUS gene expression directly after cocultivation revealed a high competence of subepidermal cell layers of cotyledonary node and associated cotyledons for transformation with Agrobacterium. PMID- 11272417 TI - In vitro multiplication of Peganum harmala--an important medicinal plant. AB - The frequency of shoot regeneration and multiplication of P. harmala was influenced by the type of explant and kind and concentration of hormones. Of the various seedling explants, cotyledonary node exhibited maximum shoot regeneration frequency from axillary region on MS medium supplemented with 5 microM BAP. Addition of 0.1 microM NAA enhanced the efficacy of BAP for multiple shoot regeneration as well as improved the growth of shoots. BAP (5 microM) in combination with NAA (0.1 microM) was found to be the optimal for inducing an average of 4-5 shoots per explant in 75% of the cultures within 5 weeks. Replacement of BAP with other cytokinins at equimolar concentration of BAP i.e. 5 microM was not effective in inducing multiple shoots. Regenerated shoots were rooted on MS medium containing IBA (8 microM) with 80% efficiency. The plantlets were successfully established in soil where 80% of them developed into morphological normal plants. PMID- 11272418 TI - Plant regeneration from leaf explants of mulberry: influence of sugar, genotype and 6-benzyladenine. AB - A protocol for plant regeneration from leaf explants was developed for tropical mulberry varieties. Effect of sugars, 6-benzyladenine and genotype on shoot regeneration was studied. Highest percentage of shoot regeneration (80 +/- 6) was obtained with genotype S799 on medium containing glucose and 8.9 microM 6 benzyladenine. Genotypes Mandalaya and MIHP, having thicker leaves with waxy cuticle, showed poorer regeneration ability than S799 and Sujanpur-5, which have thinner leaves and cuticle. Histological studies revealed that shoots regenerated from sub-epidermal cells. PMID- 11272419 TI - Effect of eugenol and tincture of crataegus (TCR) on in vitro oxidation of LDL + VLDL isolated from plasma of non-insulin dependent diabetic patients. AB - The present study was carried out to study the effect of antioxidants on oxidised LDL + VLDL and found that vitamin E, eugenol and tincture of crataegus (antioxidants) inhibited oxidation of (LDL + VLDL) similar to standard antioxidant (butylated hydroxy toluene). Vitamin C acted as an antioxidant at lower concentration, and prooxidant at higher concentration. PMID- 11272420 TI - Detection of tomato leaf curl geminivirus in its vector Bemisia tabaci. AB - Geminiviruses are single-stranded DNA plant infecting viruses that cause major losses in important crops in tropical and subtropical countries. Tomato leaf curl virus (TLCV) belonging to the genera Begomovirus, is a whitefly-transmitted geminivirus that causes a severe leaf curl disease in tomato (Lycopersicon esculentum). The importance of this disease has prompted a great need for a rapid identification of TLCV in its hosts and vector. Polymerase chain reaction (PCR) is the most sensitive approach to detect a minute amount of viral nucleic acid. It is the most ideal method to amplify geminiviruses as they replicate via a double-stranded, circular DNA form. In this study, geminivirus specific degenerated primers were employed to detect TLCV occurring in its vector whitefly Bemisia tabaci by PCR based approach. One primer pair, amplified TLCV DNA fragment of about 1.1 Kb representing partly replicase gene, intergenic region and partly coat protein gene was used. When a set of primer targeted to the core region of the coat protein gene of geminivirus was used, a PCR amplified fragment of about 0.5 Kb was obtained. This approach is highly useful for an early detection of TLCV occurring in very small amount in the vector B. tabaci. Its implications in geminivirus management strategies and their differentiation and being discussed. PMID- 11272421 TI - Antifungal activity of bicyclic heterocyclic-1,2-diazole. AB - A novel series of heterocyclic-1,2 diazole namely N1-iso nicotinoyl-5,5'-dimethyl cyclohexane-4-(sulpha/substituted phenylazo)-1,2-diazoles have been synthesized. The compounds were screened for the anti-fungal properties against building fungi. The fungal species used for this purpose were Aspergillus niger and Pencillium frequentans. It was found that out of a series of 25 compounds, fourteen have shown significant fungicidal properties against both the above species. Minimum inhibition concentration was observed between 100 and 200 ppm for most of the compounds. PMID- 11272422 TI - Study on cephalopod's ink for anti-retroviral activity. AB - Aqueous extracts of ink from four cephalopods, adult and young Sepiella inermis and Loligo duvaucelli were tested against Moloney murine leukaemia virus reverse transcriptase (MMLV RT). Ink from young cephalopods, S. intermis and L. duvaucelli showed strong inhibition of MMLV RT. PMID- 11272423 TI - Dashboard keeps system focused on strategic goals. PMID- 11272424 TI - Data and added reimbursement in Anthem OB/GYN program lead to improved patient care. PMID- 11272425 TI - Wireless technology the link between MDs, transcriptionists. AB - A technological and operational makeover of a Miami physician practice has cut the red tape involved in transcribing and maintaining chart entries. Routing information also has become easier and less susceptible to errors. PMID- 11272426 TI - DM approach to renal disease slashes utilization, boosts outcomes. PMID- 11272427 TI - Use data benchmarks to boost labor productivity. AB - The key to effective and long-lasting improvement in labor productivity is to equip middle managers with the right tools, says a former hospital administrator now serving as a consultant. One strategy is to look hard at typically ignored labor benchmarks. PMID- 11272428 TI - Transition to an electronic health system with long-term strategy. PMID- 11272429 TI - [Acute cholecystitis in patients over 80 years of age: indication for immediate surgical treatment]. AB - The Authors analyse the results of very early surgical treatment in 43 patients over 80 years of age. The severity of morphological changes in the gallbladder and complicated course of the disease are the main factors causing an unfavorable effect on the results of a later treatment. Echography allows, in a high percentage of cases, to confirm the clinical doubt of acute cholecystitis. Cholecystectomy was carried out in all the patients within the first 6 hours (very early surgery-VES). The overall mortality rate was 7%; the morbility rate was 18%. PMID- 11272430 TI - [Acquired abnormalities of the biliary tract. Preoperative diagnosis and surgical risk in the laparoscopic era]. AB - Acquired anomalies of the biliary tract are rare. The aim of this work was to examine their frequency and to assess potential associated danger when performing a laparoscopic cholecystectomy. A retrospective analysis of clinical charts of 3.870 patients undergoing elective cholecystectomy between 1959 and 1997 was performed. Eighteen cases of choledoco-duodenal fistula, 9 of cholecysto-duodenal and 12 of cholecysto-choledochal fistulas were observed. Two cases of acquired absence of the cystic duct and one cholecysto-colic fistula were also encountered. The traditional contrastographic radiology showed to be more accurate in defining presence and nature of the acquired anomalies. Etiopathogenesis of the main anomalies and consequent risks in performing laparoscopic cholecystectomy were discussed. PMID- 11272431 TI - [The Allgrove syndrome]. AB - Allgrove syndrome, rare autosomal recessive pathology, characterized by adrenocortical insufficiency, achalasia and alacrimia, rises usually in pediatric age, while its observation in adults is very rare. A clinical case appeared in an adult, observed by the Authors, is the reason to evaluate the etiopathogenetic mechanisms of this syndrome, with particular attention about potential role of genetic abnormalities and about others factors, again not well known. PMID- 11272432 TI - Adenomas with severe dysplasia and early carcinoma of the colon-rectum: our experience of 27 patients. AB - Authors report their experience in the treatment of adenomas with severe dysplasia and early cancer of the colon-rectum confirming that the endoscopic resection of these lesions is safe and curative when completely removed and no submucosal invasion is found. Between 1995 and 1999, 219 patients underwent colonoscopy and 287 polyps were removed. Histologic examination showed 217 adenomatous polyps (75.6%), 58 non adenomatous (20.2%) and 12 early carcinomas (4.2%). Severe dysplasia was found in 15 adenomas. There were no complications. The Authors focused on 12 patient with early carcinoma and 15 with severe dysplasia. The mean follow-up was 34 months. No recurrences were observed, supporting that this procedure is the treatment of choice in selected cases. PMID- 11272433 TI - [Pseudo-thrombophlebitic syndrome of the leg caused by cyst in the inner popliteal hamstrings. A case report]. AB - The Authors present a clinical case of pseudo-thrombophlebitic syndrome caused by a cyst of the inner popliteal hamstrings. Pointing out anatomic characteristics of the area involved together with the physiopathology and etiology of the formation that fill the space of the inner popliteal hamstrings, in agreement with the observations reported in literature, and intervene in determining a great number of venous blockage observed in the leg. In conclusion, they observe how a correct clinical-anamnestic approach is important in the diagnosis and consequent prompt and aimed therapy, and how this approach should use the modern instruments available today for a definite diagnosis in the presence of a syndrome of venous stasis of the lower limbs. PMID- 11272435 TI - [Role of ultrasound-guided percutaneous alcohol administration in the treatment of solitary cysts of the liver]. AB - Non-parasitic liver cysts represent a frequent pathology, but rarely they cause compressive symptoms so to require a suitable treatment. The Authors report a case of solitary large cyst localized in the left liver lobe, causing compressive symptoms in a cardiopathic old woman in whom surgical treatment was contraindicated. The treatment consisted in US-guided percutaneous puncture, decompression and sclerotherapy by 300 cc of ethylic alcohol (90 degrees). The patient showed a moderate alcoholic intoxication, associated with fever. This treatment been repeated fifteen days after for the large dimension of the cyst. Twelve months later the results were excellent with a complete relief of compressive symptoms and a remarkable reduction of the cyst at the CT scan. The Authors conclude that, in case of non operable large liver cysts, percutaneous ethanol injection is the treatment of choice, with low cost and morbidity, and mild recurrence rate. PMID- 11272434 TI - ["Strawberry" gallbladder: review of the literature and our experience]. AB - The term "strawberry gallbladder" refers to an anatomo-pathological aspect which is included in the wider chapter of gallbladder cholesterolosis. Introducing their experience, the Authors summarize the hypotheses through the years proposed about the etiopathogenesis of this condition the clinical symptoms that it can produce and the diagnostic strategies used to identify it. Moreover the Authors underline that the "strawberry gallbladder" continues to be reported with a significant frequency. They also confirm the opportunity of surgical treatment of symptomatic patients as a valid alternative to medical therapy which is not always effective, it is long lasting, often complex and not completely side effects lacking. PMID- 11272436 TI - [Idiopathic varicocele. Diagnosis and follow-up with CW-Doppler and Echo-Color Doppler: our experience]. AB - The Authors outline their experience on routine use of Doppler velocimetry in pre and post-operation studies on patients affected with idiopathic varicocele. This instrumental study, characterized by non-invasive techniques and repeatability, if correctly carried out, allows both accurate identification of anatomical alterations which are at the bottom of varicocele and correct indications for surgical treatment. Moreover the Authors confirm the importance of spermatic veins ligature following the Ivanissevich method in the treatment of male infertility. PMID- 11272437 TI - Laparoscopic cholecystectomy in adult patients with sickle cell disease. AB - Cholecystectomy is a common surgical procedure performed in patients with sickle cell disease (SCD). Postoperative complications, including acute painful vaso occlusive crisis and acute chest syndrome, have been described frequently after either traditional or laparoscopic cholecystectomy (LC). It's still not clear if preoperative blood transfusion, hyperhydration, intraoperative body temperature conservation may reduce complications rate. The Authors reviewed the charts of seven patients with SCD operated on LC for symptomatic gallbladder lithiasis and describe their perioperative management. In 3 patients preoperative endoscopic removal of stones was achieved. Five patients with HB lower than 9 g/dl and/or HbS higher than 40% were transfused preoperatively and all the patients were hyperhydrated. Intraoperative monitoring was achieved for early recognition of ventilation to perfusion mismatch and acid-base balance or temperature modifications. The Authors reported only one case of postoperative lower extremities pain. This study suggests that LC is a safe procedure in SCD if appropriate monitoring and perioperative management are achieved. PMID- 11272438 TI - [Colonic pouch versus direct colo-anal anastomosis in the reconstruction after total resection of the mesorectum: review of the literature]. AB - Good results in terms of control of the disease and 5 years survival have encouraged the use of the sphincter-saving technique for the treatment of cancers of the lower rectum. However, after this operation a percentage of patients complains of functional abnormalities such as increased bowel frequency and modification of continence especially within 12-18 months after surgery. This review analyzes the results of coloanal anastomosis following rectal excision trying to evaluate if the construction of a colonic pouch allows to limit or to prevent the functional anorectal abnormalities that usually follow a straight colo-anal anastomosis. PMID- 11272439 TI - [Robots in surgery]. PMID- 11272440 TI - [Advanced gastric adenocarcinoma with RER+ phenotype presents a good prognosis after 5 years of curative resection]. AB - Microsatellite instability (MIN) has been found both in advanced and early gastric cancer. To find out the step played by MSI in gastric carcinogenesis, links between RER+ phenotype and clinical and pathological aspects have been studied. In this work our purpose is to analyze the relationship between MIN+ advanced gastric cancer and prognosis at 5 years after radical surgery. We investigated 34 patients affected by gastric cancer who underwent R0 surgical resection from February 1991 to October 1994. After that, they underwent a four monthly follow-up for a minimum of 5 years. Genetic abnormalities have been searched including (a) those occurring in common-type CIN carcinomas and (b) those characteristic of MIN cancers. DNA extraction showed the presence of microsatellite instability (MIN) in 9 (26%) of the samples (vs. 74% of chromosomal instability CIN); none of them was M+ (vs. 12% of CIN cancers). Recurrence occurred in 2 out of 9 of the MIN cancers (22%) and in 21 out of 25 CIN cancers (84%). In conclusion, our data suggest that advanced gastric cancers with mutator phenotype show a better outcome at 5 years than the CIN phenotype. PMID- 11272441 TI - Fat embolism, ARDS, coma, death: the four horsemen of the fractured hip. AB - The pathophysiology of fat embolism syndrome (FES) is presented in the context of total joint arthroplasty. The current literature is reviewed with recommendations for surgical technique, anesthetic and pulmonary management. Diagnosis is quite difficult but can be established by imaging techniques such as MRI, SPECT, and transcranial Doppler sonography. Early steroid treatment may limit morbidity. PMID- 11272442 TI - Childhood disabilities in medical education at the John A. Burns School of Medicine (JABSOM). PMID- 11272443 TI - Clinical and epidemiological observations regarding the 1998 Kauai murine typhus outbreak. AB - Five cases of murine typhus occurring on southwestern Kauai in 1998 are described, following an investigation by the Department of Health. Two cases also had concurrent leptospirosis. Recent habitat changes of peridomestic animals and their fleas may have increased the risk for developing murine typhus. Increased suspicion of typhus by island physicians and more aggressive rodent control activities are recommended. PMID- 11272444 TI - Pleomorphic adenoma. AB - Pleomorphic adenoma is the most common neoplasm of the salivary glands. Though a benign lesion, proper recognition and management of this process is needed to avoid increasing enlargement of the mass, facial nerve impairment, risk of malignant degeneration, and recurrence after surgical resection. The epidemiology, diagnosis, and treatment options for this neoplasm are discussed. PMID- 11272445 TI - Radiology case of the month. Painless mass in my right thigh. Rectus femoris tear of the musculotendinous junction presenting as a pseudotumor. PMID- 11272446 TI - Dr Andrew W. Smyth. PMID- 11272447 TI - Management of good-grade intracranial aneurysms. AB - In this retrospective study over a 6-year period, 205 patients in good neurological condition with aneurysms were treated at Louisiana State University Health Sciences Center--Shreveport. Of the 205 patients, there were 120 patients in Hunt and Hess Grade I, 52 patients in grade II, and 32 patients in grade III; 84 aneurysms were unruptured. Ninety-six patients reached the hospital within 3 days of subarachnoid hemorrhage, 77 on the first day. The majority underwent surgery within 3 days of presentation (N = 141). A good outcome was observed in 190 patients; 3 patients died (1 in grade II and 2 in grade III). The outcome was significantly influenced by Hunt & Hess Grade at admission, location of the aneurysm, and size of the aneurysm. Aneurysms of the posterior circulation and aneurysms with larger sacs had significantly worse outcomes than aneurysms in the anterior circulation. No patients in grade I died, and there was no mortality in the patients with unruptured aneurysms. The overall mortality was 1.5% and the morbidity was 4.5%. PMID- 11272448 TI - Management dilemmas in patients with hereditary renal adysplasia. AB - We report a neonatal case of right renal aplasia with left dysplastic kidney and mild pulmonary hypoplasia. The respiratory insufficiency gradually improved on high frequency oscillation and conventional ventilation. Severe hypotension necessitated the use of inotrops. Anuria and electrolyte imbalances were managed by peritoneal dialysis. At age 13 days the baby had a small bowel perforation, developed septic shock and, after discussion with the multi-disciplinary team and the family, inotropic support was withdrawn and the baby died. The family history revealed the father had a newborn from a previous marriage who died secondary to bilateral renal agenesis. Renal studies in the father showed agenesis of the kidney with normal renal functions suggesting the diagnosis of hereditary renal adysplasia, an autosomal dominant condition with variable expression. This case illustrates the importance of renal ultrasound of the parents and siblings of affected newborns with structural kidney anomalies. A general consensus is lacking as to which infants with bilateral renal adysplasia should be aggressively treated. PMID- 11272449 TI - Knowledge and attitudes of family physicians about clinical practice guidelines and the care of patients with type 2 diabetes mellitus. AB - This study examined the attitudes of Louisiana family physicians toward clinical practice guidelines in general and specifically how attitudes and familiarity with the American Diabetes Association Clinical Recommendations (ADACR) correlated with knowledge and evidence-based "best practice" in the care of type 2 diabetes. Surveys were mailed to a random sample of 278 eligible physicians from which a 32% response rate was obtained (n = 90). Family physicians' general attitudes towards guidelines were neutral. Attitude correlated significantly with knowledge of the ADACR (P = .03) but not with "best practice". Despite low scores for knowledge, all but one of the ADACR were adhered to by more than 85% of respondents. Physician attitudes do not appear to be barriers to guideline implementation. Results may be used to focus studies of processes and outcomes in guideline implementation. PMID- 11272450 TI - A smoking cessation helpline for Louisiana smokers--and a new resource for medical professionals. AB - The Louisiana Office of Public Health Tobacco Control and Prevention Program received funding from the Louisiana State Legislature to expand smoking cessation activities in the state. The Tobacco Control Program conducted a review of other states' current smoking cessation programs, cessation programs in Louisiana, and epidemiological evidence in planning the expansion of the Louisiana tobacco smoking cessation program. As a result, a smoking cessation helpline has been developed. Medical clinicians in Louisiana have a powerful impact on their patients' smoking cessation success. Simple discussion by physicians with their patients about how to quit smoking has improved successful quitting rates by 40%. Approximately 70% of smokers visit a physician each year and 60% visit a dentist, so there is clearly an opportunity to reach many persons at risk. The telephone smoking cessation helpline at 1-800-LUNG-USA is a tool for clinicians to use when advising their patients on ways to improve their health and prevent disease. PMID- 11272451 TI - Homogenized is best. PMID- 11272452 TI - ECG of the month. Contenders. Ventricular parasystole. PMID- 11272453 TI - [Determination of the potassium balances in diary cows and the examination of daily and lactation period-associated variations. AB - The objectives of this study were to evaluate a method of determining the internal and external potassium balances in dairy cows and to determine daily and lactation period-associated variations of these balances. Ingested potassium, milk potassium concentration (LK) and potassium urinary fractional clearance (FeK) defined the potassium external balance. Serum (SK) and red blood cell (CIEK) potassium concentrations defined the potassium internal balance. Whole blood, urine, and milk were collected at different times of the day from 19 cows, grouped in 5 lactation periods. Significant differences were observed for LK between lactation periods. Significant variations occurred during the day for CIEK, LK, and FeK. Thus, in order to monitor potassium balances during a time period, samples should be collected at the same time each day. This method of determining both potassium balances simultaneously could be useful in the study of the bovine hypokalemia syndrome. PMID- 11272454 TI - Effects of epidural lidocaine anesthesia on bulls during electroejaculation. AB - Two experiments were conducted to determine whether caudal epidural lidocaine anesthesia reduces a stress response to electroejaculation. In the 1st experiment, changes in cortisol and progesterone concentrations in serial blood samples were used to assess the stress response to restraint (control), transrectal massage, caudal epidural injection of saline, electroejaculation after caudal epidural injection of lidocaine, and electroejaculation without epidural lidocaine. In the 2nd experiment, behavioral responses were subjectively scored in bulls that were electroejaculated with or without caudal epidural lidocaine anesthesia. Cortisol and progesterone concentrations were significantly elevated after electroejaculation, whether or not bulls received caudal epidural anesthesia. Elevations in cortisol and progesterone were lower and fewer bulls vocalized during electroejaculation when given caudal epidural anesthesia; however, the differences were not significant. PMID- 11272455 TI - Canadian beef quality audit 1998-99. AB - The second beef quality audit was conducted in Canada in 1998-99 to determine the prevalence of quality defects in slaughtered cattle and to monitor changes since the first audit in 1995. Approximately 0.6% of the number of cattle processed annually in Canada were evaluated. Brands were observed on 49% and tag was observed on 43% of the hides. Both brands and tag had increased from 1995. Seventy percent of the cattle were polled and 5% had full horns; thus, the number of horned cattle had decreased from 1995. Bruises were found on 54% of the carcasses, which was a decrease from 78% in 1995. Sixty-eight percent of the bruises were minor, 28% major, and 4% critical in severity. The distribution of bruises on the carcass was 17% on the chuck, 36% on the rib, 30% on the loin, and 16% on the round. Grubs were observed on 0.008% of the carcasses, and surface injection site lesions were observed on 0.2% of the whole carcasses, a decrease from the 1.3% seen in 1995. Seventy-two percent of the livers were passed for human food and 14% for pet food; 14% were condemned. Approximately 64% of the liver losses were due to abscesses. Five percent of the heads and tongues and 0.3% of the whole carcasses were condemned. The hot carcass weight was highly variable in all cattle, averaging 353 kg (s = 43). The average ribeye area was 90 cm2 (s = 13). Both hot carcass weight and ribeye area had increased from 1995. The average grade fat was 9 mm (s = 5), ranging from 0 mm to 48 mm. Lean meat yield averaged 58.8% (s = 4.6). One percent of the carcasses were devoid of marbling, 17% were Canada A, 49% were Canada AA, 32% were Canada AAA, and 1% were Canada Prime, which was an increase in marbling from 1995. Dark cutters were found in 1% of all carcasses; 1% of steers, 0.5% of heifers, 3% of cows, and 14% of bulls. Three percent of the carcasses were underfinished and 13% were overfinished. The number of overfinished carcasses had increased from 1995. Stages, steers with bullish traits, were infrequently observed in 0.5% of the steers, and 0.2% of the steers and 0.3% of the heifers had poor conformation. Yellow fat was not observed in any steers or heifers, but it was found on 65% of the cow carcasses. Only 0.6% of the heifers had an aged carcass, based on skeletal maturity. Based on August 1998 to July 1999 prices, it was estimated that the Canadian beef industry lost $82.62 per head processed, or $274 million annually, from quality nonconformities, which was an increase from 1995. Additional improvements in management, feeding, handling, genetics, marketing, and grading are needed in the beef industry to reduce quality defects. PMID- 11272456 TI - Successful euthanasia of a juvenile fin whale. AB - A stranded juvenile fin whale was successfully euthanized with an intravenous injection of sedative and cardioplegic drugs. Veterinarians may face a number of serious difficulties if called to perform this task, and advance preparation is required for successful euthanasia of these animals. PMID- 11272457 TI - Seroprevalence of Neospora caninum in beef cattle in northern Alberta. AB - Blood samples were collected from 1806 pregnancy-tested cows from 174 herds at a northern Alberta auction mart in the fall of 1998. One hundred sixty-two (9.0%) of these samples were positive for antibodies to N. caninum. Thirty-five of 260 samples (13.5%) collected from the same region in the 1980s were also serologically positive for N. caninum. PMID- 11272458 TI - Laminitis in a mature elk hind (Cervus elaphus). AB - Laminitis should be considered as a differential diagnosis in elk presenting with shifting leg lameness, reluctance to move, recumbency and hoof wall ridging. Eliminating the underlying cause and corrective trimming lead to a good prognosis for recovery. PMID- 11272459 TI - Monensin toxicosis in 2 sheep flocks. AB - Several lambs in 2 sheep flocks died suddenly and others were examined for generalized weakness and dyspnea. Postmortem findings were suggestive of degenerative myocardial and skeletal muscle myopathy, which was confirmed histologically. Feed analysis revealed toxic levels of monensin and ionophore toxicosis was diagnosed. PMID- 11272460 TI - Immune-mediated vasculitis in a shar-pei with swollen hock syndrome. AB - A castrated male shar-pei was presented for episodes of lethargy, swelling of the tarsal joints, and polydipsia with polyuria. Histological examination of biopsies from skin overlying the tarsi and direct immunoperoxidase immunohistochemical staining confirmed immune complex vasculitis, suggesting a role for immune complex deposition in the pathogenesis of shar-pei fever. PMID- 11272461 TI - Partial mandibulectomy as the treatment of a comminuted mandibular fracture in a dog. PMID- 11272463 TI - Diverse career opportunities abound in veterinary medicine. PMID- 11272462 TI - Congenital hepatic fibrosis in calves. PMID- 11272464 TI - An ethicist's commentary on the swine infected with antimicrobial-resistant Salmonella. PMID- 11272465 TI - Regulation of differentiated osteoclasts. AB - Osteoclasts respond to many factors, including endocrines, cytokines, cell-cell interactions, and cell-matrix contacts. For mature osteoclasts, the first level of control occurs through signaling that follows binding to an appropriate substrate. Mononuclear and multinucleate osteoclasts are activated when cell surface integrins, notably but not exclusively alphavbeta3 integrins, bind to calcified matrices. The binding process results in actin ring formation and deployment of adhesive proteins into a ring shape such that a seal is formed. As this ring forms, components of the ruffled border assemble from diffuse distribution to form the resorption apparatus, which includes the vacuolar ATPase, carbonic anhydrase, and other key molecules. This review focuses on the control of osteoclast activity, beginning with attachment and ruffled border assembly. Direct and indirect regulation by PTH/PTHrP, genomic and nongenomic effects of estrogen, and gene expression of ruffled border components, carbonic anhydrase, and vacuolar ATPase are reviewed. Finally, the need to understand complex signaling pathway interaction is discussed. PMID- 11272466 TI - Role of mitochondria in apoptosis. AB - Apoptosis is characterized by biochemical processes that are largely conserved throughout evolution. The basic elements of the system comprise caspases, their activators and inhibitors, and regulators of mitochondrial integrity. New evidence reveals the role of mitochondria as the central coordinators of apoptosis. Accordingly, some caspases are sequestered within the mitochondria, and mitochondria contain additional proapoptotic factors. Bcl-2 and Bax homologs regulate the integrity of the mitochondrial outer membrane, which may also serve as a scaffold for the apoptotic machinery. PMID- 11272467 TI - Methylated DNA sequences in genomic imprinting. AB - Genomic imprinting is a special form of epigenetic system that determines the parent-of-origin-specific, or monoallelic, expression of a small number of genes, termed "imprinted" genes. Considerable sequence and methylation analysis of imprinted genes has revealed a common theme: Regions of allele-specific methylation inherited from the gametes, or primary differentially methylated regions (DMRs), are associated with CpG islands and repeat elements, and this overall structure suggests functional significance. For at least three imprinted genes the sequence of the primary DMR constitutes an element able to regulate gene activity in cis--a chromatin insulator and a promoter of an antisense transcript. In these cases the unique feature of imprinting appears to be in the ability to switch the regulatory capacity of these elements on or off by the absence or presence of inherited methylation. Increasing evidence therefore suggests that genomic imprinting for at least some genes constitutes the regulation of gene regulatory elements by methylation. An important challenge now is to determine how the differential methylation of primary DMR sequences is established in the germ line. If methylation is the primary imprint, then the processes establishing it are the primary imprinting mechanisms. Trans-acting factors that are expressed in one sex of germ line and not the other are likely to be involved, and their ability to methylate may be mediated through repeat elements associated with the sequence of primary DMRs. PMID- 11272468 TI - Osteopoiesis: the early development of bone cells. AB - An understanding of the disorders of bone formation clearly requires insights into the complex regulatory events occurring during the evolution of bone precursor cells into osteoblasts. Moreover, a rational approach to therapeutic interventions that might alter the clinical course of bone disorders must take into consideration the exact nature of the developmental control mechanism(s) being affected during the disease process. The process of osteopoiesis involves the proliferation and maturation of primitive precursor cells into functional osteoblasts. The bone cell lineage originates from mesenchymal stem cells that commit to the osteogenic cell lineage becoming osteoprogenitor cells, preosteoblasts, osteoblasts, and osteocytes. In order to understand how different regulatory signals coordinate bone cell development, it is important to study the responses of bone progenitor cells to different microenviromental signals. This requires that lineage markers be identified for the various populations of bone cells and their precursors, that cell separation techniques be established so that cells of the osteogenic lineage can be purified at different stages of differentiation, and that these isolated cells are studied under serum-free, chemically defined conditions. This review focuses on the current understanding of bone progenitor cell development, examining the various types of precursor cells, their responses to cytokines and other extracellular influences, and recent observations on the biochemical and molecular control of lineage-specific gene expression. Although the emphasis is on human cells, the importance of work using rodent cells goes without saying, and is addressed where relevant. PMID- 11272469 TI - Regulation of apoptosis by E1A and Myc oncoproteins. AB - E1A and c-myc are oncogenes that can deregulate the cell cycle and promote transformation under conditions where normal cell-cycle checkpoints are inactivated. In situations where cell-cycle checkpoints are intact, the E1A and c Myc proteins potently induce apoptosis, a property that is believed to be the end result of a cellular response to uncontrolled growth-promoting signals. p53 is a key regulator of E1A and c-myc-induced apoptosis and, together with the oncoproteins, may transcriptionally activate numerous genes whose products influence, or are themselves, members of the core apoptotic machinery. The upstream signaling events and the ultimate apoptotic pathways activated by E1A and c-Myc are discussed in this review. PMID- 11272470 TI - Mechanisms of tumor metastasis to bone. AB - Bone metastases occur in approximately 80% of patients with advanced cancer. They are characterized by cancer cell growth and bone destruction that cause pain, fractures, anemia, and hypercalcemia. At diagnosis, bone metastases are usually incurable owing to their advanced development. However, the early stages in their formation are asymptomatic and begin as single micrometastatic cells from the blood stream. These cells can be detected by molecular analysis of bone marrow in approximately 30% of patients at the time of cancer diagnosis, but not all single micrometastatic cells develop into clinically significant bone metastases. A synergistic relationship exists between the micometastasis and the bone environment creating favorable conditions for the development and growth of disseminated tumor cells. Such bone metastases induce osteolysis or new bone formation, releasing growth factors and cytokines, which in turn amplify this pathological mechanism. The underling hypothesis, first proposed by Paget in 1889, is that the growth of disseminated tumor cells in bone is dependent on the fertility of the soil or bone itself. This article explores the most current opinions in this area of study and presents a comprehensive summary of the major factors involved. PMID- 11272471 TI - Control of chromatin remodeling. AB - Chromatin structure has a pivotal role in the regulation of gene expression. Transcriptional activation or the repression of a gene require the recruitment of multiple chromatin remodeling complexes. Chromatin remodeling complexes modulate the higher order structure of chromatin, facilitate or hinder the binding of transcription factors, and aid in or prevent the establishment of a transcriptional preinitiation complex. Two types of chromatin remodeling complexes have been extensively studied--ATP-dependent chromatin remodeling complexes and histone-modifying enzymes--which include histone acetyltransferases, histone deacetylases, and histone kinases. Transcriptional activators and repressors are responsible for recruitment of one or more of these large, multisubunit chromatin remodeling complexes. In this review, the features of the chromatin remodeling complexes and the modes of their recruitment are presented. PMID- 11272472 TI - Adaptive learning: interventions for verbal and motor deficits. AB - Advances in basic and clinical neuroscience are uniting to form new optimism for treatment and rehabilitation of persons with a variety of neurologic disorders. Both cognitive and motor systems have shown remarkable degrees of plasticity in response to incoming stimuli. Understanding the brain (and spinal cord) capacity for change will lead to new topics for research as well as new approaches to rehabilitation. Adaptive learning has been shown to be a fundamental part of the developmental process and has been used in remediation of a variety of language difficulties. Using such principles to approach motor functions also is showing promise. Expanding these observations to encompass other areas of disease and rehabilitation is an area for further research. Interdisciplinary approaches including the fields of computer technology, imaging, and genetic analysis will provide new tools. Contribution of new concepts within adaptive learning must address such topics as the relation between motor and sensory responses, measures that accurately indicate cognitive health, the brain and spinal cord areas involved in particular learning tasks, the optimal time windows for intervention, and the importance of behavior and motivation in treatment and rehabilitation. PMID- 11272473 TI - Neuroimaging of recovery of function after stroke: implications for rehabilitation. AB - Stroke is a leading cause of morbidity and mortality in individuals. Many patients have good functional recovery after stroke. The mechanisms of recovery remain largely unknown. Neuroimaging of patients recovering from stroke may provide important insight into the mechanisms of recovery as well as assist in the development of new rehabilitation techniques. The first part of this article reviews previous neuroimaging studies that have monitored the reorganization within the motor and language areas after stroke. In the second section, a unifying theory based on John Hughlings Jackson's "Principles of Compensation" is presented as a possible theory for recovery of function. In the final portion of the article, possible implications and future applications of neuroimaging studies for rehabilitation are presented. PMID- 11272474 TI - Predicting discharge destination for patients with severe motor stroke: important functional tasks. AB - Many patients with severe stroke are capable of returning to the community after receiving rehabilitation services. The purpose of this study was to describe outcomes of patients with stroke in FIM-FRG STR1, a classification based on the Functional Independence Measure, and identify important functional tasks associated with discharge to home. FIM-FRG STR1 is one of nine subpopulations of stroke that have been identified based on motor/cognitive FIM subscale score and age. We reviewed the program evaluation data of 259 cases of stroke from 1993 to 1996. We performed a descriptive analysis of the data and a logistic regression analysis to determine which tasks measured by the FIM were associated with discharge destination, a key indicator of rehabilitation success. We found that three admission FIM variables (bladder management, toilet transfers, memory) and three discharge FIM variables (upper body dressing, bed/chair transfers, comprehension) were associated with discharge destination with up to 75% accuracy. The implications of these findings are discussed. PMID- 11272475 TI - Effects of postlesion experience on behavioral recovery and neurophysiologic reorganization after cortical injury in primates. AB - Previous studies have shown that after injury to the hand representation in primary motor cortex (M1), size of the spared hand representation decreased dramatically unless the unimpaired hand was restrained and monkeys received daily rehabilitative training using the impaired fingers. The goal of this study was to determine if restriction of the unimpaired hand was sufficient to retain spared hand area after injury or if retention of the spared area required repetitive use of the impaired limb. After infarct to the hand area of M1 in adult squirrel monkeys, the unimpaired hand was restrained by a mesh sleeve over the unimpaired arm. Monkeys did not receive rehabilitative training. Electrophysiologic maps of M1 were derived in anesthetized monkeys before infarct and 1 month after infarct by using intracortical microstimulation. One month after the lesion, the size of the hand representation had decreased. Areal changes were significantly smaller than those in animals in a previous study that had received daily repetitive training after infarct (p < 0.05). Areal changes were not different from those in a group of animals that received neither rehabilitative intervention nor hand restraint after injury. These results suggest that retention of hand area in M1 after a lesion requires repetitive use of the impaired hand. PMID- 11272476 TI - The effects of ipsilateral forearm movement and contralateral hand grasp on the spastic hand opened by electrical stimulation. AB - The purpose of this study was to investigate the effects of ipsilateral arm movement and contralateral hand grasp on the spastic hand opened by open-loop electrical stimulation. The major problem of applying proper electrical stimulation is variable spasticity, the intensity of which changes with posture and movements of other parts of the body. Electrical stimulation was applied to extensor digitorum communis and ulnar nerve to open the affected hand. Different procedures were then used to assess the effects of moving the ipsilateral forearm and contracting the contralateral normal hand. Electrical stimulation opened the spastic hand in more than 95% of trials in all subjects, whether stimulation was applied before or after the movement of the forearm. Moving the ipsilateral forearm did have an effect on opening the hand, and making adjustment of stimulation intensities was necessary in all subjects. The stimulation opened the spastic hand during the contraction of the contralateral normal hand. Electrical stimulation could open the spastic hands most of the time, in the presence of ipsilateral forearm movement and contralateral normal hand contraction. If electrical stimulation was applied before the ipsilateral forearm was moved toward the target, stimulation intensities needed to be adjusted. PMID- 11272477 TI - Controlling an artificial arm with foot movements. AB - A history of prosthesis control techniques is outlined. An alternative approach to hand and arm prosthesis control is proposed, and a working-prototype model is described. Commercial artificial limbs have function limited by a small number of user-interface control channels, and they are awkward to operate because joints are turned on and off in a serial fashion and not in parallel. Because of natural anatomic and physiologic similarities between the upper and lower extremities, the foot and leg is an ideal control interface. A stocking laced with multiple sensors can provide input on multiple joint positions, and this information used to control homologous movements in a prosthetic upper extremity. One significant advantage of this form of control is simultaneous activation of multiple joints. PMID- 11272478 TI - Mirror movements complicate interpretation of cerebral activation changes during recovery from subcortical infarction. AB - In recovered stroke patients, performance of motor tasks with the affected limb has been reported to activate cortical areas ipsilateral to the affected side. The better to determine the causal role these areas play in recovery of motor function, we assessed cerebral activation during motor activity longitudinally after hemiparesis due to cerebral infarction. A secondary goal was to ascertain the relation between mirror movements and activation ipsilateral to motor activity. Positron emission tomography with oxygen-15 water measured regional cerebral blood flow during wrist movement early and late in the course of recovery from hemiparesis. Surface electromyography recorded muscular activity, and computer-assisted video analysis quantified movement during the scans. Mirror movements, movements contralateral to the instructed movement of the hemiparetic arm, were often seen. Activation of motor areas in the hemisphere ipsilateral to the affected limb roughly correlated with presence of mirror movements. Other changes in cerebral activation were small, when the task was controlled for rate, but high-rate-specific recruitment of ipsilateral cortical areas occurred in one case. However, the common occurrence of mirror movements, particularly with effortful tasks, complicates interpretation of data regarding the role of the ipsilateral hemisphere in recovery. PMID- 11272479 TI - Minimizing discomfort with surface neuromuscular stimulation. AB - The purpose of this study was to evaluate the effects of stimulus parameters, electrode types, and electrode positions on the perception of discomfort during lower extremity surface neuromuscular stimulation. Ten normal and eight neurologically impaired (four incomplete spinal cord and four stroke) subjects were enrolled. Neurologically impaired subjects had some sensation, although it was often reduced. Parameters of the stimulation were varied in a way that produced the same level of ankle dorsiflexion, as measured with a goniometer. Discomfort was assessed after each stimulation with a 0-10 verbal scale (0, no discomfort; 10, worst pain). Increasing the pulse frequency was associated with increased discomfort for subjects in both groups (p > 0.05). Increasing the pulse duration was associated with increased discomfort in the neurologically impaired subjects (p > 0.05), but not in the normal subjects (p > 0.05). The electrode size and type had no effects on discomfort (p > 0.05). Stimulation of the peroneal nerve over the fibular head was better tolerated than the direct motor point stimulation of the tibialis anterior motor point (p < 0.05). The data suggest that to minimize discomfort, surface stimulation should be applied over nerves rather than motor points, and frequency and pulse duration should be set as low as possible for a given degree of contraction. PMID- 11272480 TI - Recovery rates after stroke and their impact on outcome prediction. AB - Current assessments do not provide reliable factors predictive of outcome from stroke for stroke survivors of intermediate age and severity of deficit. We sought to investigate whether early rate of functional improvement can facilitate prediction of functional outcome, length of stay, and disposition beyond that afforded by age and initial severity of deficit. Prospective study of consecutive admissions to acute rehabilitation (N = 244) with diagnosis of ischemic or hemorrhagic stroke. Independent measures were age, marital status, living situation, social situation, lag from symptom onset to rehabilitation, stroke type, admission score on the Functional Independence Measure (FIM), rate of FIM change (ROFC) as assessed by the best weekly FIM change in the first 3 weeks of rehabilitation. Dependent measures were functional status on discharge as assessed by a modification of Steinman's method, length of stay, and discharge disposition. Logistic regression analyses on each of the dependent measures identified significant factors, and interactions of significant factors were assessed by analysis of variance on continuous dependent variables. Cross tabulations using significant factors from the logistic regression analyses were performed to identify groups with homogeneous outcomes. Groups with >80% homogeneity were considered likely to have predictive value. Discharge functional status: Admission FIM (AFIM) again fractionated the population into groups with poor outcome (AFIM <50 remained dependent), good outcome (AFIM >70 achieved nondependence), and an intermediate group with unpredictable outcome. In this intermediate group, ROFC had significant effect only for a small number of patients (n = 9) with rapid improvement (ROFC >25) who achieved nondependence. LOS: AFIM >70 had less than average LOS, ROFC = 10-15 FIM/week had longer than average LOS. LOS was significantly prolonged in patients with poor outcomes. Disposition: AFIM >70 and age <60 were strongly associated with home discharge. Patients not living at home before admission were not discharged home. Married patients had a greater tendency to home discharge than did those not married. ROFC had no bearing on disposition. ROFC has an independent influence on outcome but was sufficiently powerful in our sample to identify reliably only a very small subset of patients with otherwise indeterminate prognosis. LOS seems inordinately prolonged in patients with poor outcomes. Both of these results can guide efficient rehabilitation management. PMID- 11272481 TI - Nifedipine and nimodipine competitively inhibit uridine kinase and orotidine phosphate decarboxylase: theoretical relevance to poor outcome in stroke. AB - Nifedipine and nimodipine, dihydropyridine calcium channel blockers, are commonly used as antihypertensive and antianginal agents in patients at risk for stroke. At least one stroke trial suggests that patients receiving calcium channel blockers at the time of an acute stroke have worse outcomes than those receiving other or no antihypertensive medications. We hypothesize that the poor outcome may not be related to blood pressure changes but instead may be mediated by competitive inhibition of important enzymes of pyrimidine synthesis whose products are needed to repair nerve cell membranes after an acute stroke. Both drugs acted as competitive inhibitors of the only enzymes that are known to synthesize the nucleotide uridine-5'-phosphate: uridine kinase and orotidine-5' phosphate decarboxylase. Nifedipine produced Ki values of 28 microM for uridine kinase and 105 microM for orotidine-5'-phosphate decarboxylase. Nimodipine produced Ki values of 20 microM for uridine kinase and 18 microM for orotidine-5' phosphate decarboxylase. For uridine kinase, these inhibitors bound more tightly than the physiologic substrates uridine or cytidine. For the decarboxylase, the inhibitors bound less tightly than the normal physiologic substrate orotidine-5' phosphate. Additional experiments are needed to determine whether the concentrations of nifedipine or nimodipine, and of cytidine, uridine, and orotidine-5'-phosphate in human brain, are such that this inhibition would affect stroke outcome. PMID- 11272482 TI - Experimental transmission of cutaneous chytridiomycosis in dendrobatid frogs. AB - In a series of three experiments during March-October, 1998, two species of captive-bred poison dart frogs (Dendrobates tinctorius and D. auratus) were exposed to Batrachochytrium dendrobatidis, a recently-described chytridiomycete fungus (chytrid) that was originally isolated from a blue poison dart frog (D. azureus). All frogs exposed to the chytrids developed a fatal skin disease, whereas none of the control frogs developed skin lesions. The most consistent clinical sign in chytrid-exposed frogs was excessive shedding of skin. Gross lesions were subtle, usually affected the legs and ventrum, and consisted of mild skin thickening and discoloration. Microscopic examination of shed skin pieces and/or skin imprints demonstrated the presence of chytrids and was used for ante mortem and post mortem confirmation of chytrid infection. Histologically, there was epidermal hyperkeratosis, hyperplasia, and hypertrophy associated with low to moderate numbers of chytrids in the keratinized layers. These experiments demonstrated that Batrachochytrium dendrobatidis can be a fatal pathogen in poison dart frogs. The experimentally-induced disease in these frogs resembled cases of cutaneous chytridiomycosis that have recently been described in several other species of captive and wild amphibians. PMID- 11272483 TI - Pathology of brucellosis in bison from Yellowstone National Park. AB - Between February 1995 and June 1999, specimens from seven aborted bison (Bison bison) fetuses or stillborn calves and their placentas, two additional placentas, three dead neonates, one 2-wk-old calf, and 35 juvenile and adult female bison from Yellowstone National Park (USA) were submitted for bacteriologic and histopathologic examination. One adult animal with a retained placenta had recently aborted. Serum samples from the 35 juvenile and adult bison were tested for Brucella spp. antibodies. Twenty-six bison, including the cow with the retained placenta, were seropositive, one was suspect, and eight were seronegative. Brucella abortus biovar 1 was isolated from three aborted fetuses and associated placentas, an additional placenta, the 2-wk-old calf, and 11 of the seropositive female bison including the animal that had recently aborted. Brucella abortus biovar 2 was isolated from one additional seropositive adult female bison. Brucella abortus was recovered from numerous tissue sites from the aborted fetuses, placentas and 2-wk-old calf. In the juvenile and adult bison, the organism was more frequently isolated from supramammary (83%), retropharyngeal (67%), and iliac (58%) lymph nodes than from other tissues cultured. Cultures from the seronegative and suspect bison were negative for B. abortus. Lesions in the B. abortus-infected, aborted placentas and fetuses consisted of necropurulent placentitis and mild bronchointerstitial pneumonia. The infected 2-wk-old calf had bronchointerstitial pneumonia, focal splenic infarction, and purulent nephritis. The recently-aborting bison cow had purulent endometritis and necropurulent placentitis. Immunohistochemical staining of tissues from the culture-positive aborted fetuses, placentas, 2-wk-old calf, and recently-aborting cow disclosed large numbers of B. abortus in placental trophoblasts and exudate, and fetal and calf lung. A similar study with the same tissue collection and culture protocol was done using six seropositive cattle from a B. abortus-infected herd in July and August, 1997. Results of the bison and cattle studies were similar. PMID- 11272484 TI - Evaluation of the fluorescence polarization assay and comparison to other serological assays for detection of brucellosis in cervids. AB - The complement fixation test (CFT), competitive enzyme immunoassay (CELISA), indirect enzyme immunoassay (IELISA) and fluorescence polarization assay (FPA) were evaluated for the detection of antibodies to Brucella abortus and Brucella suis biotype 4 in caribou (Rangifer tarandus caribou), elk (Cervus elapus), red deer (Cervus elapus), and reindeer (Rangifer tarandus tarandus). When combining the data the FPA and the CELISA were determined to be the most suitable tests for serodiagnosis of Cervidae. The overall actual sensitivity of the CFT and the IELISA was 100%. The overall actual sensitivity for the CELISA and FPA was 99%. The overall relative specificity of the CFT (including treatment of anti complementary data as positive or negative for analysis), the CELISA, the IELISA and the FPA were 65%, 93%, 99%, 99%, and 99%, respectively. The specificities of the buffered plate agglutination test (BPAT), the CFT, the CELISA, the FPA and the IELISA for 55 elk vaccinated with B. abortus strain 19 and tested 4 mo post vaccination were 14%, 31%, 51%, 84%, and 2%, respectively. The FPA is the diagnostic test of choice because it has sensitivity and specificity values comparable to the CELISA; it has the capability to distinguish vaccinal antibody and antibody resulting from exposure to cross-reacting organisms such as Yersinia enterocolitica 0:9 from antibody to Brucella spp. in most cases; it is technically simple to do; it is adaptable to field use and it is relatively inexpensive. PMID- 11272485 TI - Elimination of rabies from red foxes in eastern Ontario. AB - The province of Ontario (Canada) reported more laboratory confirmed rabid animals than any other state or province in Canada or the USA from 1958-91, with the exception of 1960-62. More than 95% of those cases occurred in the southern 10% of Ontario (approximately 100,000 km2), the region with the highest human population density and greatest agricultural activity. Rabies posed an expensive threat to human health and significant costs to the agricultural economy. The rabies variant originated in arctic foxes: the main vector in southern Ontario was the red fox (Vulpes vulpes), with lesser involvement of the striped skunk (Mephitis mephitis). The Ontario Ministry of Natural Resources began a 5 yr experiment in 1989 to eliminate terrestrial rabies from a approximately 30,000 km2 study area in the eastern end of southern Ontario. Baits containing oral rabies vaccine were dropped annually in the study area at a density of 20 baits/km2 from 1989-95. That continued 2 yr beyond the original 5 yr plan. The experiment was successful in eliminating the arctic fox variant of rabies from the whole area. In the 1980's, an average of 235 rabid foxes per year were reported in the study area. None have been reported since 1993. Cases of fox rabies in other species also disappeared. In 1995, the last bovine and companion animal cases were reported and in 1996 the last rabid skunk occurred. Only bat variants of rabies were present until 1999, when the raccoon variant entered from New York (USA). The success of this experiment led to an expansion of the program to all of southern Ontario in 1994. Persistence of terrestrial rabies, and ease of elimination, appeared to vary geographically, and probably over time. Ecological factors which enhance or reduce the long term survival of rabies in wild foxes are poorly understood. PMID- 11272486 TI - Hemoprotozoa of freshwater turtles in Queensland. AB - Blood smears from 27 turtles (15 Emydura signata, nine Elseya latisternum, and three Chelodina longicollis) from southeastern Queensland (Australia) were examined for infections by hemoprotozoan parasites between January and June 1999. Infections were found in 26 (96%) of the turtles. Twenty five (93%) were infected with the adeleorin coccidian Haemogregarina clelandi, eight (30%) with the hemosporidian Haemoproteus chelodinae, 11 (41%) with the kinetoplastid flagellate Trypanosoma chelodinae, and eight (30%) with a novel Trypanosoma sp. Despite the high prevalence and intensity of infections, there was no evidence of clinical disease in any of the turtles. PMID- 11272487 TI - Rabies and canine distemper in an arctic fox population in Alaska. AB - Two oil field workers were attacked by a rabid arctic fox (Alopex lagopus) in the Prudhoe Bay oil field (Alaska, USA) prompting officials to reduce the local fox population. Ninety-nine foxes were killed during winter 1994. We tested foxes for prevalence of rabies and canine distemper. Exposure to rabies was detected in five of 99 foxes. Of the five, only one fox had rabies virus in neural tissue as determined by the direct fluorescent antibody test. The other four foxes had been exposed to rabies, but had apparently produced antibodies and did not have an active infection. No evidence of canine distemper was detected as determined by the absence of distemper antibodies in serum and distemper virus in neural tissue. PMID- 11272488 TI - Aleutian mink disease parvovirus in wild riparian carnivores in Spain. AB - Serious declines in populations of native European mink (Mustela lutreola) have occurred in Europe. One responsible factor may be infectious diseases introduced by exotic American mink (Mustela vison). In order to investigate a possible role for Aleutian mink disease parvovirus (ADV), we surveyed native riparian carnivores and feral American mink. When serum samples from 12 free-ranging European and 16 feral American mink were tested, antibodies to ADV were detected from three of nine European mink. ADV DNA was detected by polymerase chain reaction in whole cell DNA from four of seven carcasses; two American mink, one European mink and a Eurasian otter (Lutra lutra). Lesions typical of Aleutian disease were present in one of the American mink. A portion of the ADV VP2 capsid gene was sequenced and the results suggested that two sequence types of ADV were circulating in Spain, and that the Spanish ADVs differed from other described isolates from North America and Europe. Future conservation and restoration efforts should include measures to avoid introduction or spread of ADV infection to native animals. PMID- 11272489 TI - Seroepizootiology of selected infectious disease agents in free-living birds of prey in Germany. AB - Four hundred forty-eight blood plasma samples from free-living birds of prey from Berlin and the Brandenburg area in eastern Germany were tested for antibodies against Newcastle disease virus (NDV), falcon herpesvirus (FHV), owl herpesvirus (OHV), and Chlamydia psittaci. Antibodies to NDV were detected in 6 (2%) of 346 tested diurnal birds of prey, whereas none of the owls (n = 55) was positive. The positive samples originated from two common buzzards (Buteo buteo), three ospreys (Pandion haliactus) and one marsh harrier (Circus aeruginosus). Titers varied between 1:8 and 1:32. Of 253 birds of prey one osprey (<1%) tested positive for antibodies to FHV with low titer of 1:6. This is the first detection of antibodies against FHV in an osprey. Furthermore, antibodies against OHV could be found in one tawny owl (Strix aluco) and one common buzzard (2 of 253, 1%) with low titers of 1:6. Of 422 birds of prey 267 (63%) tested positive for antibodies to Chlamydia psittaci with titers varying between 1:5 and 1:256 which reflects the ubiquitous occurrence of Chlamydia psittaci in these birds of prey. PMID- 11272491 TI - Hematological and biochemical reference intervals for wild caught Eurasian otter from Spain. AB - Hematologic and serum chemistry reference intervals were determined from 33 wild caught Eurasian otters (Lutra lutra lutra) between November 1995 and May 1998 during a reintroduction project. Blood was obtained by jugular venipuncture after administration of ketamine and medetomidine. The mean, standard deviation, and range for 19 hematology parameters and 28 serum chemistry values are presented. There were no significant differences between sexes in most analytes. The results are in agreement with those reported previously for Eurasian otters with the exception of higher leukocyte and neutrophil counts, lower eosinophil and lymphocyte counts and higher activities for aspartate aminotransferase and creatine kinase. The Eurasian otters have lower erythrocyte counts but higher mean corpuscular volume and mean corpuscular hemoglobin values than the river otter (Lutra canadensis) in North America. PMID- 11272490 TI - Experimental adenovirus hemorrhagic disease in white-tailed deer fawns. AB - Infection with a newly described endotheliotropic adenovirus was the cause of a 1993 epizootic reminiscent of hemorrhagic disease in California mule deer (Odocoileus hemionus columbianus and O. hemionus hemionus). Pulmonary edema and intestinal luminal hemorrhage, or necrotizing stomatitis associated with systemic or localized vasculitis, respectively, were common lesions seen in animals that died during the epizootic. In order to determine if white-tailed deer (Odocoileus virginianus) also are susceptible to infection and fatal disease with the deer adenovirus, eight white-tailed deer fawns (4- to 6-mo-old) were inoculated with purified deer adenovirus. Four were inoculated intravenously and four were inoculated through the mucous membranes. Seven days post-inoculation, one of the fawns inoculated intravenously died. Pulmonary edema and hemorrhagic enteropathy were associated with pulmonary and intestinal vasculitis with systemic multiorgan distribution of endotheliotropic adenovirus as demonstrated by transmission electron microscopy and immunohistochemistry. Adenovirus was reisolated from lung homogenates of the fawn that died of adenovirus hemorrhagic disease. PMID- 11272492 TI - Cloning, sequencing, and expression of interferon-gamma from elk in North America. AB - Eradication of Mycobacterium bovis relies on accurate detection of infected animals, including potential domestic and wildlife reservoirs. Available diagnostic tests lack the sensitivity and specificity necessary for accurate detection, particularly in infected wildlife populations. Recently, an in vitro diagnostic test for cattle which measures plasma interferon-gamma (IFN-gamma) levels in blood following in vitro incubation with M. bovis purified protein derivative has been enveloped. This test appears to have increased sensitivity over traditional testing. Unfortunately, it does not detect IFN-gamma from Cervidae. To begin to address this problem, the IFN-gamma gene from elk (Cervus elaphus) was cloned, sequenced, expressed, and characterized. cDNA was cloned from mitogen stimulated peripheral blood mononuclear cells. The predicted amino acid (aa) sequence was compared to known sequences from cattle, sheep, goats, red deer (Cervus elaphus), humans, and mice. Biological activity of the recombinant elk IFN-gamma (rElkIFN-gamma) was confirmed in a vesicular stomatitis virus cytopathic effect reduction assay. Production of monoclonal antibodies to IFN gamma epitopes conserved between ruminant species could provide an important tool for the development of reliable, practical diagnostic assays for detection of a delayed type hypersensitivity response to a variety of persistent infectious agents in ruminants, including M. bovis and Brucella abortus. Moreover, development of these reagents will aid investigators in studies to explore immunological responses of elk that are associated with resistance to infectious diseases. PMID- 11272494 TI - Necropsy findings and environmental contaminants in common loons from New York. AB - Diagnostic and analytical findings are presented for 105 common loons (Gavia immer) found dead or debilitated in New York (USA) from 1972-99. Aspergillosis (23% of cases) and ingestion of lead fishing weights (21%) were the most common pathologies encountered. Stranding on land, shooting, other trauma, gill nets, air sacculitis and peritonitis, and emaciation of uncertain etiology accounted for most of the remaining causes of disease or death. Analysis for total mercury in the liver of 83 loons yielded a geometric mean (gm) of 10.3 mg/kg (wet basis) and range of 0.07 to 371 mg/kg, with emaciated birds generally showing higher levels. Organochlorine contaminant levels in brain were generally low, principally consisting of PCB's (gm = 2.02 mg/kg) and DDE (0.47 mg/kg). PMID- 11272493 TI - Hematological, protein electrophoresis and cholinesterase values of free-living nestling peregrine falcons in Spain. AB - Protein electrophoresis, hematological and cholinesterase values were determined in 32 nestling free-living peregrine falcons (Falco peregrinus) (15- to 27-days old) in order to establish normal reference values for this population. The following values (mean +/- SD) were observed: prealbumin 0.31 +/- 0.04 g/dl, albumin 1.25 +/- 0.06 g/dl, alpha1 and alpha2-globulin 0.23 +/- 0.02 and 0.16 +/- 0.02 g/dl respectively, beta-globulin 1.02 +/- 0.05 g/dl, gamma-globulin 0.060 +/ 0.08 g/dl, total protein 3.79 +/- 0.18 g/dl, 21.26 +/- 1.30 white blood cells/microl (1 x 10(3)), 2.17 +/- 0.07 red blood cells/microl (1 x 10(6)), packed cell volume 37.58 +/- 0.82%, hemoglobin 20.96 +/- 0.29 g/dl, heterophils 61.14 +/- 2.50% and cholinesterase 1,184 +/- 75 IU/L. There were no difference in any of these parameters among males and females. The hematological values obtained could be considered as representative values in free-living nestling peregrine falcons. PMID- 11272495 TI - Epidermoptid mange in Laysan albatross fledglings in Hawaii. AB - Mange caused by the epidermoptid mite Myialges nudus (Acari: Epidermoptidae) is described in 31 dead fledgling Laysan albatrosses (Phoebastria immutabilis) from Midway Atoll (Hawaii, USA) sampled from 18 June to 10 July 1990 and from 21 June to 22 July 1991. This is the first record for this parasite from this host. Mites were collected from the skin; were located primarily in the stratum corneum; and were associated with mild to severe granulomatous inflammation, hyperkeratosis, dermal edema, ballooning degeneration of keratinocytes, neovascularization, and subdermal fibrosis. The severity of inflammation in some birds suggested that dermatitis due to M. nudus could be a significant cause of morbidity, or even mortality, in these birds. PMID- 11272496 TI - Ectoparasites of the island fox on Santa Cruz Island. AB - The ectoparasite fauna for island foxes (Urocyon littoralis) on Santa Cruz Island (California, USA) in April (wet season) and September (dry season) 1998 was evaluated. Three taxa of ectoparasites were identified. These were fleas (Pulex irritans), lice (Neotrichodectes mephitidis), and ticks (Ixodes pacificus). Ectoparasite abundances varied seasonally. Typical of insular endemic species, island foxes may be especially vulnerable to the introduction of novel disease organisms and their vectors. PMID- 11272497 TI - Feline leukemia virus in a captive bobcat. AB - An 11-mo-old captive-bred male neutered bobcat (Felis rufus) presented with lethargy, anorexia, leukopenia, neutropenia, lymphopenia, and nonregenerative anemia. The animal was diagnosed as feline leukemia virus (FeLV) positive by immunofluorescent antibody and enzyme-linked immunosorbant assay (ELISA) testing. It died despite supportive care. Pathologic examination revealed multifocal non suppurative encephalitis, diffuse interstitial pneumonia, multifocal hepatocellular necrosis, non-suppurative peritonitis, and lymphoid depletion. FeLV was isolated from peripheral blood mononuclear cells, bone marrow, spleen, and lymph node. FeLV-specific gag sequences were amplified by DNA polymerase chain reaction (PCR) and aligned with known domestic cat FeLV's. The source of the virus was speculated to be a domestic cat that was a surrogate nurse. Case reports of FeLV in nondomestic felids are few, and FeLV does not appear to be enzootic in wild felids, except European wildcats (Felis silvestris) in France and Scotland. Introduction of FeLV into free-living and captive nondomestic felid populations could have serious consequences for their health and survival. Measures to prevent the introduction of this virus to nondomestic felids are warranted. PMID- 11272498 TI - Serological responses and immunity to superinfection with avian malaria in experimentally-infected Hawaii amakihi. AB - Six of seven Hawaii Amakihi (Hemignathus virens) with chronic malarial infections had no increases in peripheral parasitemia, declines in food consumption, or loss of body weight when rechallenged with the homologous isolate of Plasmodium relictum 61 to 62 days after initial infection. Five uninfected control amakihi exposed at the same time to infective mosquito bites developed acute infections with high parasitemias. Reductions in food consumption and loss of body weight occurred in all control birds and three of these individuals eventually died. When surviving birds were rechallenged >2 yr later with either the same parasite isolate or an isolate of P. relictum collected on the island of Kauai, all individuals were immune to superinfection. Chronically infected birds developed antibodies to a common suite of malarial antigens ranging in size from 22 to 170 kDa that were detectable as early as 8 days post infection on immunoblots of SDS polyacrylamide gels. Antibodies to this suite of malarial antigens persisted as long as 1,248 days after initial infection and were consistently detectable at times when parasites were not easily found by microscopy on Giemsa-stained blood smears. The immunoblotting method that is described here appears to be an effective technique for identifying birds with chronic, low-intensity malarial infections when circulating parasites are not easily detectable by microscopy. Hawaiian honeycreepers that are capable of recovering from acute infections develop concomitant immunity to superinfection, making them functionally immune in areas where malaria transmission has become endemic. PMID- 11272499 TI - Ehrlichiosis in a moose calf in Norway. AB - A case of granulocytic ehrlichiosis in a moose calf (Alces alces) in Norway is described. The animal was heavily infested with ticks (Ixodes ricinus), and died from a Klebsiella pneumoniae septicemia. Examination of blood smears from the calf revealed cytoplasmic inclusions (morulae) typical of infection with Ehrlichia phagocytophila in the granulocytes. Ehrlichia sp. was detected by polymerase chain reaction (PCR) in blood from the calf, and in the ticks. Sequence determination identified it as E. phagocytophila. This is the first report of ehrlichiosis in moose. PMID- 11272500 TI - Saprolegniosis in salmonids and their eggs in Japan. AB - An epizootic of the fungal infection saprolegniosis that occurred in freshwater cultured salmons and their eggs at some hatcheries in Hokkaido (Japan) was investigated. In almost all cases, the initial clinical sign was characterized by the growth of cotton-like mycelia on the fishs' body surface, especially the head, adipose fin, and caudal fin, but the mycelia were not visible to the naked eye in the internal organs. Thirty-three strains isolated from lesions were classified in the genus Saprolegnia according to their morphological and biological characteristics on hemp seed cultures at various temperatures. Fifteen of the strains were identified as Saprolegnia parasitica, 16 were identified as S. salmonis, and two were identified as S. australis. PMID- 11272501 TI - Interspecific variability of prevalence in blood parasites of adult passerine birds during the breeding season in Alaska. AB - Blood parasite prevalence based on microscopic examination of stained blood smears was determined in adults of 11 passerine bird species sampled during their breeding season (May and June 1997-98) in interior Alaska (USA). These species included primarily Nearctic migratory species such as the dark-eyed junco (Junco hyemalis) and neotropical migratory species such as the blackpoll warbler (Dendroica striata), alder flycatcher (Empidonax alnorum), Swainson's thrush (Catharus ustulatus), northern waterthrush (Seiurus noveboracensis), and bank swallow (Riparia riparia) as well as one long-distance palearctic migrant, the arctic warbler (Phylloscopus borealis). The more prevalent parasites were Leucocytozoon dubreuili (73% of the sampled turdinids), L. fringillinarum (42% of the sampled fringillids and parulids), and Trypanosoma avium (39% of the sampled hosts). Other parasites (H. fallisi: 18% of the sampled turdinids; Haemoproteus paruli: 14% of the sampled parulids; H. fringillae: 5% of the sampled fringillids; microfilariae: 4% of the sampled hosts) were observed less frequently. Plasmodium vaughani was found only in two yellow warblers (Dendroica petechia). Overall parasite prevalence varied between 0% in the alder flycatcher to >80% in Swainson's thrush, arctic warbler, and Townsend's warbler (Dendroica townsendi). Prevalence of various hematozoa also was bird species-dependent. No relationship was observed between prevalence and either foraging (aerial versus trees/shrubs) or nesting habits (ground versus arboreal) or general location of the wintering area of the different species examined. Prevalence also was unrelated to average dates of arrival on breeding grounds and, therefore, to potential duration of exposure to local insect vectors before capture. Differences in blood parasite prevalence among species breeding in a same region and in the same type of habitat may result from differences in host specificity such as immunological resistance to infection or blood meal preference by potential vectors and/or in behavioral adjustments/physiological traits that alter exposure to vectors. PMID- 11272503 TI - Lumpy jaw in wild sheep and its evolutionary implications. AB - The distribution and prevalence of mandibular osteomyelitis, lumpy jaw, and other dental anomalies in wild sheep were investigated and their biological and evolutionary implications were assessed. Our survey was based on 3,363 mandibles of wild sheep and 1,028 from domesticated varieties. Lumpy jaw is widespread in wild sheep of North America, but it is rare or absent in wild sheep from Eurasia. Among the subspecies of Ovis spp. in North American, the thinhorn sheep (Ovis dalli) were the most seriously impacted, with a prevalence in Dall's sheep (O. dalli dalli) of 23.3% and 29.3% in Stone's sheep (O. dalli stonei). Among the bighorns (O. canadensis), the Rocky Mountain subspecies (O. canadensis canadensis) had a higher rate (12.1%) than other subspecies. Lumpy jaw was not documented in the desert sheep of Baja California (O. canadensis cremnobates, O. canadensis weemsii). Based on data from affected thinhorn sheep, it appears there is an inverse relationship between age of a subspecies in a long term evolutionary context and susceptibility to lumpy jaw. In Eurasian wild sheep lumpy jaw is rare or absent with prevalences ranging from 0 to 7.1% among suspecies, and in domesticated breeds the prevalence averaged 5.0%. The impact of lumpy jaw on different age classes or longevity is equivocal, although females are more susceptible than males. Lumpy jaw appears to effect horn development in males. PMID- 11272502 TI - Serologic survey for Toxoplasma gondii in lynx from interior Alaska. AB - Two hundred fifty-five lynx (Felis lynx) carcasses were collected from trappers in Interior Alaska (USA). Serosanguinous fluids were collected from the chest cavity of each carcass. These fluids were tested for evidence of exposure to Toxoplasma gondii by means of a modified agglutination test using formalin fixed tachyzoites and mercaptoethanol. Thirty-nine of the samples had titers greater than or equal to the threshold (> or = 25). Antibody prevalence differed between areas, and was directly related to age of the host. PMID- 11272504 TI - Evaluation of ewe vaccination as a tool for increasing bighorn lamb survival following pasteurellosis epizootics. AB - We conducted field and laboratory experiments to evaluate whether treating pregnant bighorn ewes with a combination of an experimental Pasteurella trehalosi and Mannheimia haemolytica (formerly P. haemolytica) vaccine and a commercially available bovine P. multocida and M. haemolytica vaccine would increase lamb survival following a pneumonia epidemic. Three free-ranging bighorn herds affected by pasteurellosis outbreaks between November 1995 and June 1996 were included in the field experiment. Post-epidemic lamb survival was low in all three herds in 1996, with November lamb:ewe ratios of < or = 8:100. In March 1997, thirty-six ewes (12/herd) were captured and radiocollared. Half of the ewes captured in each herd were randomly selected to receive both vaccines; the other half were injected with 0.9% saline solution as controls. Lambs born to radiocollared ewes were observed two or more times per week and were considered to have survived if they were alive in October 1997, about 6 mo after birth. Lamb survival differed among herds (range 22% to 100%), and survival of lambs born to vaccinated ewes was lower (P = 0.08) than survival of lambs born to unvaccinated ewes. Bronchopneumonia (pasteurellosis) was the dominant cause of mortality among lambs examined. We concurrently evaluated vaccine effects on survival of lambs born to seven captive ewes removed from the wild during the 1995-96 epidemic. Antibody titers were high in captive ewes prior to vaccination, and vaccines failed to enhance antibody titers in treated captive ewes. None of the captive born lambs survived. These data suggest that, using existing technology, vaccinating bighorn ewes following pneumonia epidemics has little chance of increasing neonatal survival and population recovery. PMID- 11272505 TI - Bovine tuberculosis in free-ranging carnivores from Michigan. AB - During a survey of carnivores and omnivores for bovine tuberculosis conducted in Michigan (USA) since 1996, Mycobacterium bovis was cultured from lymph nodes pooled from six coyotes (Canis latrans) (four adult female, two adult male), two adult male raccoons (Procyon lotor), one adult male red fox (Vulpes vulpes), and one 1.5-yr-old male black bear (Ursus americanus). One adult, male bobcat (Felis rufus) with histologic lesions suggestive of tuberculosis was negative on culture but positive for organisms belonging to the Mycobacterium tuberculosis complex when tested by polymerase chain reaction. All the tuberculous animals were taken from three adjoining counties where M. bovis is known to be endemic in the free ranging white-tailed deer (Odocoileus virginianus) population. There were two coyotes, one raccoon, one red fox, and one bobcat infected in Alpena county. Montmorency County had two coyotes and one raccoon with M. bovis. Two coyotes and a bear were infected from Alcona County. These free-ranging carnivores/omnivores probably became infected with M. bovis through consumption of tuberculous deer. Other species included in the survey were opossum (Didelphis virginiana), gray fox (Urocyon cinereoargenteus), and badger (Taxidea taxus); these were negative for M. bovis. PMID- 11272506 TI - Serum antigen 85 levels in adjunct testing for active mycobacterial infections in orangutans. AB - Diagnosis of active mycobacterial disease in orangutans (Pongo pygmaeus) has been impeded by high levels of non-specific intradermal skin test reactivity to mycobacterial antigens. This may be due in part to cross reactivity between antigens, tuberculin concentrations used or other species-specific factors. Antigen 85 (Ag85) complex proteins are major secretory products of actively growing mycobacteria, and measurement of serum Ag85 could provide a method for determining active mycobacterial infections that was not dependent on host immunity. Serum Ag85 was measured by dot-immunobinding assay using monoclonal anti-Ag85, purified Ag85 standard and enhanced chemiluminescence technology in coded serum samples from 14 captive orangutans from a zoo in Colorado, 15 semi captive orangutans in Malaysia, and 19 free-ranging wild orangutans in Malaysia. Orangutans from Colorado (USA) were culture negative for Mycobacterium tuberculosis and M. avium, although all had laboratory suspicion or evidence of mycobacterial infection; median serum Ag85 was 10 microU/ml (range, <0.25-630 microU/ml). Of the semi-captive orangutans, six were skin test reactive and two were culture positive for M. avium on necropsy. Median serum Ag85 for this group was 1,880 microU/ml (0.75-7,000 microU/ml), significantly higher than that of Colorado zoo or free-ranging Malaysian orangutans. Median serum Ag85 in the latter group was 125 microU/ml (range, 0.75-2,500 microU/ml). These data suggest that suggest that additional studies using more specific reagents and more samples from animals of known status are appropriate. PMID- 11272507 TI - Host range and dynamics of mycoplasmal conjunctivitis among birds in North America. AB - An epidemic of conjunctivitis among house finches (Carpodacus mexicanus) caused by Mycoplasma gallisepticum (MG) bacterial infections was first described in 1994. The disease exhibits high primary host specificity, but has been isolated from a limited number of secondary avian hosts at various times and locations. We used records from the House Finch Disease Survey, a continent-wide, volunteer monitoring project, to document the host range of conjunctivitis in birds at feeding stations and to investigate how disease in house finches might influence the spread of conjunctivitis to other hosts. Between 1994 and 1998, participants recorded 675 cases of conjunctivitis in 31 species other than house finches in eastern North America. Seventy five % of these cases were observed among three species: American goldfinches (Carduelis tristis), purple finches (Carpodacus purpureus) and house sparrows (Passer domesticus). The proportion of sites with diseased wintering populations of the three species increased over the 4 yr study and coincided with range expansion of conjunctivitis in house finches. Sites with diseased house finches present were significantly more likely to report conjunctivitis in each of the three species during the same month. These observations are most consistent with transmission of an infectious agent (presumably MG) from house finches to these secondary hosts via spillover of localized epidemics, rather than sustained interspecific transmission. PMID- 11272508 TI - Characterization of the mycoplasmal conjunctivitis epizootic in a house finch population in the southeastern USA. AB - An epidemiological study of the prevalence of mycoplasmal conjunctivitis in the house finch (Carpodacus mexicanus) was conducted in Auburn (Alabama, USA) between March 1998 and February 1999. Clinical disease was observed in 4% of the 1,214 finches trapped and examined. This rate is comparable to the average annual prevalence observed in this population since 1996, although the prevalence of clinical disease observed in the peak months of September through November was lower than in previous years. Clinically ill birds were observed in all months of the study. To estimate the prevalence of recovering and asymptomatic, infected birds, we tested a subset of 334 house finches serologically for exposure to Mycoplasma gallisepticum (MG) using the serum plate agglutination (SPA) assay. The prevalence of clinical disease in this subsample was slightly higher (7%) than in the entire sample, reflecting the fact that the serological survey was initiated in the late summer when the prevalence of MG infection peaks in our study population and a sampling bias for symptomatic birds. The serological survey indicated that 13% of this subpopulation had been exposed to MG. We also tested 46 of 334 finches by polymerase chain reaction (PCR) to detect MG in seropositive, asymptomatic birds. Use of the PCR in conjunction with the SPA detected six asymptomatic, infected birds that may represent potential carriers or birds in the early stages of infection. The decreasing prevalence of clinical disease observed during the peak months suggests a changing host-parasite relationship. Continued surveillance of this population, employing both clinical observation and serological analysis will be useful in characterizing these changes over time. PMID- 11272509 TI - Serologic survey of Brucella spp. antibodies in some marine mammals of North America. AB - A serologic survey of anti-Brucella spp. antibodies was undertaken on 2,470 samples of 14 North American marine mammal species collected between 1984-97. Serum or blood from eight species of cetaceans and six species of pinnipeds was sampled from Pacific, Atlantic, and Arctic oceans. Two competitive enzyme-linked immunosorbent assays (C-ELISA's), using specific monoclonal antibodies to Brucella abortus cell wall components, were used to detect anti-Brucella spp. antibodies in the samples. Sera from 33 cetaceans and 61 pinnipeds gave inhibition values, in one or both of the tests, which exceeded the threshold that indicates Brucella spp. exposure in cattle. Seropositive animals were identified from Pacific, Atlantic, and Arctic oceans. While Brucella spp. was not isolated, differences in the response of seropositive cetacean and pinniped sera in the two assays suggest that two antigenically distinct species or biovars of Brucella spp. are present. No pathology consistent with clinical brucellosis was noted in any of the animals tested although detailed examination was not conducted on all carcasses. PMID- 11272510 TI - Section 401(k), 403(b) and 457(b) plans: who can do what? AB - This article examines the three main types of supplementary pension arrangements. It focuses on their origins and development, employer eligibility, deferral and contribution limits, funding arrangements and other technical requirements as they have been affected by recent developments. PMID- 11272511 TI - What businesses are doing to attract and retain employees--becoming an employer of choice. AB - This article explains why the new economy has made many businesses anxious to become employers of choice. The author uses a case study to show how one firm achieved this goal and explains the role that innovative benefits practices and other HR policies can play in attracting and retaining qualified employees. PMID- 11272512 TI - The legal challenge to ERISA preemptions. AB - Managed care, once celebrated as a vehicle to halt the increasing cost of health care, has come under increasing fire from patients and health care providers, accused of cutting costs and managing care at the expense of patients. The Employee Retirement Income Security Act of 1974 (ERISA) has been a shield for managed care organizations (MCOs), buffeting them from liability for quality-of care issues. Lawsuits and legislation are chipping away at the protective shield of ERISA as MCOs find themselves more liable for their decisions and for the care provided by physicians with whom the MCO contracts and provides financial incentives for controlling cost of medical care. PMID- 11272513 TI - Communications planning. AB - Whether you are communicating a totally new benefit program or announcing a change in an existing one, effective planning will make the difference between success and failure of your communications campaign. Effective benefits communication is more than just distributing a summary plan description or conducting an employee meeting. It's more than spending a lot of money on elaborate brochures or expensive videotapes, or posting information on the intranet. More and more, benefits communicators are expected to communicate in a way that influences employees to make changes in their personal lives, a tall order that requires careful planning and coordination. PMID- 11272514 TI - Group disability income: new policies emphasize employee appeal. AB - Group disability insurance is a voluntary benefit that offers advantages to both employers and employees. Employees who want this kind of coverage realize that their employers have done much of the homework for them in terms of comparing benefits, rates and contract provisions. Employers recognize an opportunity to add a benefit that may help attract and retain valuable employees with little additional cost or administrative burden. PMID- 11272515 TI - Managing prescription drug costs. AB - This article reviews prescription drug cost trends, touching on issues such as cost versus value, demographic changes, direct-to-consumer advertising, effects of the Internet and disease management. The author also discusses pharmacy benefit managers, as well as various utilization management strategies like benefit plan design and the use of formularies. PMID- 11272517 TI - Vision care: a compensation cornerstone. AB - Vision care is a benefit that provides advantages for both employers and employees, and it has become an essential element of most competitive benefit packages. Well-designed vision care programs offer participants choice, options, self-help and value. An option that employers may want to consider is reduced prices on laser vision correction procedures. PMID- 11272516 TI - Human rights issues and employee benefit plans. AB - Canadians have human rights protections at both provincial and federal levels of government. At the federal level, the most important legislative enactments are the Canadian Charter of Rights and Freedoms (the "Charter") and the Canadian Human Rights Act. Provincially and territorially, human rights are legislatively safeguarded primarily by provincial human rights codes. Both federally and provincially, human rights may also be impacted by a variety of other statutes and regulations such as employment standards acts, workers' compensation acts, occupational health and safety acts, and pay equity legislation. PMID- 11272518 TI - Taking the pulse of personalized and online employee communication strategies: the second annual survey of major employer trends. AB - In their efforts to build workforce commitment, many employers are using personalized communication to reach out to employees. Benefacts, the personalized communication service of Aon Consulting, recently surveyed employers nationwide regarding their uses of personalized communication, especially in the online environment. The survey looked at organizations' current online and print communication strategies as well as their future plans. This article describes the needs of employees to understand organizational purpose and to find a balance between work and their personal lives. This examination of the uses of personalized communication to meet these needs and the analysis of current industry trends will help benefit professionals focus on their own communication plans as they strive to meet the challenges of today's workforce. PMID- 11272519 TI - Direct determination of the single-ion anisotropy in a one-dimensional magnetic system by high-field EPR spectroscopy; synthesis, EPR, and X-ray structure of NixZn1-x(C2O4)(dmiz)2. AB - The synthesis, X-ray structure, and EPR measurements of the integer-spin linear chain antiferromagnet [Ni(ox)(dmiz)2] (where ox = C2O4(2-) and dmiz = 1,2 dimethylimidazole) are presented. The sign and size of the single-ion zero field splitting (Zfs) of the divalent Ni have been determined by high field/high frequency EPR spectroscopy. The spectra of powder samples of the derivatives [NixZn1-x(C2O4)(dmiz)2] for x = 0.09 and 0.07, at frequencies ranging from 110 to 440 GHz allowed the accurate determination of the zfs parameters D and E, with D = 1.875(4) cm(-1) and E = 0.38 cm(-1). The X-ray structure has been determined from measurements on a single crystal with x = 0.07. Structural parameters are as follows: a = 14.5252(7) A, b = 12.1916(8) A, c = 8.6850(8) A,beta = 97.460(6)degrees in space group C2/c. The zigzag chain contains octahedrally coordinated metal ions with two cis-oriented N-coordinated dmiz ligands and two cis-oriented, tetradentate bridging oxalato(2-) ligands, together resulting in a MN2O4 donor set. The structure was refined to a conventional R value of 0.073 for 1,051 observed reflections. Zn-O distances are 2.167(5) A and Zn-N = 2.098 A. Coordination angles vary for cis angles from 78.4 to 100.7 degrees, with trans angles varying from 163.9 degrees to 165.5 degrees. PMID- 11272520 TI - Influence of anionic ligands (X) on the nature and magnetic properties of dinuclear LCuDgX3.nH2O complexes (LH2 standing for tetradentate Schiff base ligands deriving from 2-hydroxy-3-methoxybenzaldehyde and X being Cl, N3C2, and CF3COO). AB - The monometallic precursor L1Cu (L1H2 standing for 1,3-bis((3 methoxysalicylidene)amino)-2,2'-dimethylpropane) reacts with GdC13 x 6H2O to afford a dinuclear complex which crystallizes in the orthorhombic space group Pca2(1) (No. 29) in a cell having the dimensions a = 9.0246(11) A, b = 16.5198(14) A, c = 20.286(2) A, and Z = 4. Analysis of the structural data shows that it may be formulated as [L1CuCl2Gd(H2O)4]Cl x 2H2O. The cationic dinuclear unit possesses a CuO2Gd bridging core which is almost planar. The complex displays a ferromagnetic interaction (10.1 cm(-1) which is the largest yet reported for a structurally characterized dinuclear (Cu-Gd) complex. Lower magnetic interactions are observed for neutral L1CuGdX3 x H2O complexes (X = N3C2, CF3COO). Consideration of the magnetic and structural data obtained for various dinuclear (Cu-Gd) complexes leads to a correlation between the magnitude of the magnetic interaction and the exponential of the dihedral angle between the two halves of the CuO2Gd bridging core. PMID- 11272522 TI - In pursuit of the molybdenum(III) tris(thiolate) fragment: unusual structure of a dimolybdenum mu-nitrido complex. AB - The new molybdenum nitrido-thiolate complex N triple bond Mo(SAd)3 (Ad = 1 adamantyl) was prepared by a ligand exchange route involving reaction of Ti(SAd)(OiPr)3 with Chisholm's nitrido-butoxide complex N triple bond Mo(OtBu)3. In an effort to abstract the nitrido nitrogen from N triple bond Mo(SAd)3, the compound was treated with Mo(N[tBu]Ph)3, a three-coordinate molybdenum(III) complex. This resulted in formation of the unusual and thermally unstable (mu nitrido)dimolybdenum complex (AdS)3Mo(mu-N)Mo(N[tBu]Ph)3, which was isolated and characterized. An X-ray study revealed (AdS)3Mo(mu-N)Mo(N[tBu]Ph)3 to possess an unsymmetrical Mo-(mu-N)-Mo linkage, the Mo-thiolate fragment exhibiting a substantially longer bond to the bridging nitrogen atom. The structure of (AdS)3Mo-(mu-N)Mo(N[tBu]Ph)3 is noteworthy, displaying trigonal monopyramidal coordination at the (mu-N)-Mo-thiolate Mo center. Since N triple bond Mo(N[tBu]Ph)3 is a good leaving group, (AdS)3Mo(mu-N)Mo(N[tBu]Ph)3 should be a source of the reactive Mo(SAd)3 fragment. In all the studied reactions of the (mu nitrido)dimolybdenum complex one of the observed products was N triple bond Mo(N[tBu]Ph)3. Two products containing the Mo(SAd)3 fragment were observed: (AdS)3Mo triple bond Mo(SAd)3 and [(ON)Mo(mu-SAd)(SAd)2]2. Upon treatment with pyridine, the tris(thio-1-adamantyl)-(nitrosyl)molybdenum dimer forms the pyridine adduct (AdS)3Mo(NO)(py), which is a monomer. PMID- 11272521 TI - Is ferromagnetism an intrinsic property of the CuII/GdIII couple? 1. Structures and magnetic properties of two novel dinuclear complexes with a mu-phenolato-mu oximato (Cu,Gd) core. AB - Two original dinuclear (Cu(II),Gd(III)) complexes (1 and 2) deriving from polydentate nonsymmetrical Schiff base ligands LiH2 have been prepared. Formally they differ by the length of the diamino chain. They crystallize in the orthorhombic Pbca (No. 61) (1) and in the monoclinic P2(1/n) (No. 14) (2) space groups. The cell parameters are a = 12.6295(7) A, b = 20.7894(9) A, c = 18.3301(13) A, and Z = 8 for 1 and a = 12.7246(16) A, b = 13.5691(17) A, c = 14.5310(19) A, beta = 94.629(16) degrees , and Z = 4 for 2. These structural studies show that in both complexes the CuII and GdIII ions are doubly bridged by a phenolato oxygen atom and an oximato (N-O) pair. The bridging network is not planar. The more important distortions are observed for the complex having the larger diamino chain. Unexpectedly the latter complex presents an antiferromagnetic interaction, but the related J value is small (J approximately equal to -0.49 cm(-1)). In the former complex the interaction is ferromagnetic (J approximately equal to 3.5 cm(-1)) as it is for complexes containing (CuO2Gd) bridging cores which yield J values varying from 1.4 to 10.1 cm(-1). PMID- 11272523 TI - Substitution and hydrogenation reactions on rhodium(I)-ethylene complexes of the hydrotris(pyrazolyl)borate ligands T (T = Tp, TpMe2). AB - The bis(ethylene) Rh species TpMe2Rh(C2H4)2(1*) (TpMe2 = tris(3,5-dimethyl-1 pyrazol-1-yl)hydroborate) has been obtained from [RhCl(C2H4)2]2 and KTpMe2. Complex 1* easily decomposes in solution to give mainly the butadiene species TpMe2Rh(eta74-C4H6). In the solid state its thermal decomposition follows a different course and the allyl TpMe2RhH(syn-C3H4Me) is cleanly obtained as a mixture of exo and endo isomers. The complexes Tp'Rh(C2H4)2 (Tp' = Tp, TpMe2) afford the monosubstituted species Tp'Rh(C2H4)(PR3) upon reaction with PR3 but react differently with L = CO or CNR: the Tp compound gives dinuclear [TpRh]2(mu L)3 complexes, while, in the case of 1*, TpMe2Rh(C2H4)(L) species are obtained. The ethylene ligand of complexes TpMe2Rh(C2H4)(PR3) is labile, and several peroxo compounds of composition TpMe2Rh(O2)(PR3) have been isolated by their reaction with O2. All the mononuclear Rh(I) complexes are formulated as 18e- trigonal bipyramidal species on the basis of IR and NMR spectroscopic studies. A series of dihydride complexes of Rh(III) of formulation Tp'RhH2(PR3) have been prepared by the hydrogenation of the corresponding ethylene derivatives. Complexes [TpRh]2(mu CNCy)3, TpMe2Rh(C2H4)(PEt3), and TpMe2Rh(O2)(PEt3) have been further characterized by X-ray diffraction studies. PMID- 11272524 TI - Tuning metal-to-metal charge transfer of mixed-valence complexes containing ferrocenylpyridine and rutheniumammines via solvent donicity and substituent effects. AB - A homogeneous series of heterobimetallic complexes of [R-Fc(4-py)Ru(NH3)5](PF6)2 (R = H, Et, Br, acetyl; Fc(4-py) = 4-ferrocenylpyridine) have been prepared and characterized. The mixed-valence species generated in situ using ferrocenium hexafluorophosphate as the oxidant show class II behavior, and the oxidized sites are ruthenium centered. deltaE(1/2), E(1/2)(Fe(III)/Fe(II)) - E(1/2)(Ru(III)/Ru(II)), an upper limit for deltaGo that is an energetic difference between the donor and acceptor sites, changes sharply and linearly with Gutmann solvent donor number (DN) and Hammett substituent constants (sigma). The solvent-dependent and substituent-dependent intervalence transfer bands were found to vary almost exclusively with deltaE(1/2). The activation energy for the optical electron transfer versus deltaE(1/2) plot yields a common nuclear reorganization energy (lambda) of 0.74 +/- 0.04 eV for this series. The equation that allows one to incorporate the effect of both solvent donicity and substituents on optical electron transfer is Eop = lambda + deltaGo, where deltaGo = (deltaGo)intrinsic + (deltaGo)solvent donicity + (deltaGo)substituent effect (deltaGo )intinnsic with a numerical value of 0.083 +/- 0.045 eV was obtained from the intercept of the deltaE(1/2) of [H-Fc(4-py)Ru(NH3)5]2+,3+,4+ versus DN plot. (deltaGo)solvent donicity was obtained from the average slopes of the deltaE(1/2) of [R-Fc-(4-py)Ru(NH3)5]2+,3+,4+ versus DN plot, and (deltaGo)substituent effect was obtained from the average slopes of the corresponding deltaE(1/2) versus sigma plot. The empirical equation allows one to finely tune Eop of this series to Eop = 0.82 + 0.019(DN) + 0.44sigma eV at 298 K, and the discrepancy between the calculated and experimental data is less than 6%. PMID- 11272525 TI - Azo anion radical complexes of osmium and related nonradical species. AB - The reaction of [Os(H)(Br)(CO)(PPh3)3], 5, with 2-(phenylazo)pyridine (pap) in boiling dry heptane has afforded the azo anion radical complex [Os(pap. )(Br)(CO)(PPh3)2], 6a, as the major product and [Os(pap)(H)(CO)(PPh3)2]Br, 7, as a minor byproduct. Upon replacing pap by the better pi-acceptor azo-2,2' bipyridine (abp) in the above synthesis, the radical complex [Os(abp. )(Br)(CO)(PPh3)2], 6b, becomes the sole product. It is proposed that 6 is formed via homolytic cleavage of the Os-H bond in 5; in the formation of 7, the Os-Br bond of 5 is heterolytically cleaved. The X-ray structures of 6b and 7.CH2Cl2 have been determined. In 6b, the N-N length is 1.35(2) A, consistent with the anion radical description; in 7.CH2Cl2 the length is 1.27(1) A. The spin-bearing extended Huckel HOMO in a model of 6 is found to be approximately 70% azo-pi* in character associated with a small metal contribution. An electronic band observed in the range 600-700 nm in solutions of 6 is assigned to the HOMO --> LUMO transition, the LUMO being 95% pyridine-pi* in character. One-electron paramagnetic 6 displays well-defined anisotropic EPR features near g = 2.00. The anisotropy arises from the metal character of HOMO and is magnified by the large spin-orbit coupling in osmium. In a moisture-free environment 6 is indefinitely stable in the solid state, but in CH2Cl2-MeCN solution 6a is rapidly oxidized by air, affording [Os(pap)(Br)(CO)(PPh3)2]+, 6a+, which has been isolated as the diamagnetic PF6- salt; 6b+PF6- has been similarly prepared. The voltammetric reduction potentials of the 6+/6 couple follow the order 6a+/6a < 6b+/6b, and the carbon monoxide stretching frequencies follow the order 6a < 6b and 6a+ < 6b+. These trends are consistent with the pi-acidity order pap < abp. Crystal data are as follows: (6b, C47H38BrN4OOsP2) monoclinic, space group P21/c (no. 14), a = 10.215(4) A, b = 17.634(7) A, c = 22.473(8) A, beta = 97.67(3) degrees , Z = 4; (7.CH2Cl2, C49H42BrCl2N3OOsP2) monoclinic, space group P2(1/n) (no. 14), a = 15.323(7) A, b = 15.201(6) A, c = 19.542(7) A, beta = 92.51(3) degrees, Z = 4. PMID- 11272526 TI - Preparations and electrochemical properties of pyrazine-bridged ruthenium binuclear complexes exhibiting molecular hysteresis. AB - Three binuclear Ru complexes cis-,cis-[(NH3)4(L)Ru-pz-Ru(NH3)4(dmso)](PF6)4 (L = NH3 (4), pyridine (5), benzonitrile (6); dmso = dimethyl sulfoxide) have been prepared, and their electrochemical behavior, exhibiting molecular hysteresis, is reported. Simulations of cyclic voltammograms and thin-layer cyclic voltammograms have provided redox potentials, isomerization rates, and interconversion rates of the complexes. The rates of the conversions between two isomeric intermediate states have been determined to be5 x 10(-6) and 4 x 10(-4) s(-1) for the complex 4, 4 x 10(-5) and 4 x 10(-4) s(-1) for the complex 5, and 2 x 10(-4) and 5 x 10( 5) s(-1) for the complex 6. The equilibrium parameters between these states are discussed in relation to the redox potentials of the complexes. PMID- 11272528 TI - Iminoacylation. 3. Formation of platinum(IV)-based metallaligands due to facile one-end addition of vic-dioximes to coordinated organonitriles. AB - The reaction of vic-dioximes with the organonitrile platinum(IV) complexes trans [PtCl4(RCN)2] (R = Me, CH2Ph, Ph, vic-dioxime = dimethylglyoxime; R = Me, vic dioxime = cyclohexa-, cyclohepta-, and cyclooctanedione dioximes) proceeds rapidly under relatively mild conditions and affords products of one-end addition of the dioximes to the nitrile carbon, i.e. [PtC4(NH=C(R)ON=[spacer]=NOH)2] (1-6) (R = Me, CH2Ph, Ph, spacer = C(Me)C-(Me) for dimethylglyoxime; R = Me, spacer = C[C4H8]C, C[C5H10]C, C[C6H12]C for the other dioximes), giving a novel type of metallaligand. All addition compounds were characterized by elemental analyses (C, H, N, C1, Pt), FAB mass spectrometry, and IR and 1H, 13C[1H], and 195Pt NMR spectroscopy. X-ray structure determination of the dimethylformamide bis-solvate [PtCl4(NH=C(Me)ON=C(Me)C(Me)=NOH)2] x 2DMF (la) disclosed its overall trans geometry with the dimethylglyoxime part in anti configuration and the amidine one end (rather than N,N-bidentate) coordination mode of the N-donor ligands. When a mixture of cis- and trans-[PtC4(MeCN)2] in MeCN was treated with dimethylglyoxime, the formation of, correspondingly, cis- and trans [PtCl4(NH=C(Me)ON=C(Me)C(Me)=NOH)2] (1) was observed and cis-to-trans isomerization in DMSO-d6 solution was monitored by 1H, 2D [1H,15N] HMQC, and 195Pt NMR spectroscopies. Although performed ab initio calculations give evidence that the trans geometry is the favorable one for the iminoacylated species [PtCl4 (ligand)2], the platinum(IV) complex [PtCl4(NH=C(Me)ON=C[C4Hs]C=NOH)2] (4) was isolated exclusively in cis configuration with the two metallaligand "arms" held together by intramolecular hydrogen bonding between the two peripheral OH groups, as it was proved by single-crystal X-ray diffractometry. The classic substitution products, e.g. [PtC12(N,N-dioximato)2] (12-15), are formed in the addition reaction as only byproducts in minor yield; two of them, [PtCl2(C7H11N2O2)2] (14) and [PtCl2(C8H13N2O2)2] (15), were structurally characterized. Complexes (12-15) were also prepared by reaction of the vic-dioximes with [PtCl4L(Me2SO)] (L = Me2SO, MeCN), but monoximes (Me2C=NOH, [C4H8]C=NOH, [C5H10]C=NOH, PhC(H)=NOH, (OH)C6H4C(H)= NOH) react differently adding to [PtCl4(MeCN)(Me2SO)] to give the corresponding iminoacylated products [PtCl4(NH=C(Me)ON=CRR')(Me2SO)](7-11). PMID- 11272527 TI - Stereoselective formation of seven-coordinate titanium(IV) monomer and dimer complexes of ethylenebis(o-hydroxyphenyl)glycine. AB - Reactions between the antitumor agent titanocene dichloride (Cp2TiCl2) and the hexadentate ligand N,N'-ethylenebis-(o-hydroxyphenylglycine) (H4ehpg) have been investigated in aqueous solution and the solid state. The racemic ligands give crystals of the monomer [Ti(ehpg)(H2O)] x (11/3)H2O (1), while the meso ligand gives the oxo-bridged dimer [[Ti(Hehpg)(H2O)]2O] x 13H2O (2). Complex 1 crystallizes in the monoclinic space group C2/c with a = 24.149(4) A, b = 14.143(3) A, c = 19.487(3) A, beta = 105.371(13) degrees, V = 6417.7(19) A3, Z = 12, and R(F) = 0.0499 for 4,428 independent reflections having I > 2sigma(I), and contains seven-coordinate pentagonal-bipyramidal TiIV with two axial phenolate ligands (Ti-O, 1.869(2) A). The pentagonal plane contains the two N-atoms at 2.210(2) A, two carboxylate O-atoms at 2.061(2) A, and a water molecule (Ti-OH2, 2.091(3) A). Complex 2 crystallizes as an oxygen-bridged dimer in the triclinic space group P-1 with a = 12.521(6) A, b = 14.085(7) A, c = 16.635(8) A, alpha = 80.93(2) degrees beta = 69.23(2) degrees, gamma = 64.33(2) degrees , V = 2472(2) A3, Z = 4, and R(F) = 0.0580 for 5956 independent reflections having I > 2sigma(I). Each seven-coordinate, pentagonal-bipyramidal TiIV has a bridging oxide and a phenolate as axial ligands. The pentagonal plane donors are H2O, two carboxylate O-atoms, and two NH groups, which form H-bonds to O-atoms both in the same half-molecule (O...N, 2.93-3.13 A) and in the other half-molecule (O...N, 2.73-2.75 A); the second phenoxyl group of each Hehpg ligand is protonated and not coordinated to TiIV, but H-bonds to a nearby amine proton (O...N, 2.73-2.75 A) from the same ligand and to a nearby H2O (O...O, 2.68 A). In contrast to all previously reported crystalline metal-EHPG complexes containing racemic ligands, in which the N(S,S)C(R,R) or N(R,R)C(S,S) form is present, complex 1 unexpectedly contains the N(S,S)C(S,S) and N(R,R)C(R,R) forms. This is attributed to the presence of ring strain in seven-coordinate TiIV complexes. Moreover, the rac ligands selectively form crystals of monomeric 1, while the meso ligand selectively forms crystals of the dimer 2 (N(R,R)C(R,S) or N(S,S)C(S,R)). Complexes 1 and 2 exhibit phenolate-to-TiIV charge-transfer bands near 387 nm, and 2D NMR studies indicate that the structures of 1 and 2 in solution are similar to those in the solid state. Complex 1 is stable over the pH range 1.0 7.0, while 2 is stable only between pH 2.5 and pH 5.5. Cp2TiCl2 reacts with EHPG at pH* 7.0 to give complex 1 with a t 1/2 of ca. 50 min (298 K), but complex 2 was not formed at this pH* value. At pH* 3.7, the reaction is very slow: 1 forms with a half-life of ca. 2.5 d, and 2 after ca. 1 week at ambient temperature. The relevance of these data to the possible role of serum transferrin as a mediator for the delivery of TiIV to tumor cells is discussed. PMID- 11272529 TI - A Cu(II)-mediated C-H oxygenation of sterically hindered tripyridine ligands to form triangular Cu(II)3 complexes. AB - Two sterically hindered tris-pyridyl methane ligands, tris(6-methyl-2 pyridyl)methane (L1) and bis(6-methyl-2-pyridyl)pyridylmethane (L2), are newly synthesized. Under aerobic conditions, Ln (n = 1 or 2) reacts with CuX2 (X = Cl or Br), oxygenated at the methine position to LnOH or LnOMe. The former alcoholate ligand creates trinuclear Cu(II) complexes [Cu3(X)(LnO)3](PF6)2 [(X, n) = (Br, 1) 1, (C1, 1) 2, (Br, 2) 3, or (C1, 2) 4] in which the alkoxide oxygen atoms bridge copper centers. The crystal structures of 1-4 are presented along with their magnetic susceptibility data. The weak antiferromagnetic coupling between the Cu(II) centers in this trinuclear arrangement is due to weak interaction of the magnetic orbitals (dz2) which are oriented along three alternate sides in a hexagon of the Cu3O3 core in 1-4. Under anaerobic conditions, L1 reacts with CuBr2 to form a square pyramidal complex [CuL1Br2] (9) with the ligand facially capping. [Cu(Br)2(L1OMe)] (10) was obtained after the suspension of 9 in MeOH was stirred under air for 48 h. In the presence of cyclohexene, 9 is converted to [Cu(Br)(L1)]m (m = 1 or 2) 5 quantitatively to give trans- 1,2-dibromocyclohexane, indicating that Br2 is generated during the reaction. The FAB MS spectrum of [18O]-1 prepared by the reaction of L1 with CuBr2 under 18O2 shows that the ligand of [18O]-1 is L1(18O-.) L1(18OH), L1OCD3, and bis(6-methyl-2-pyridyl) ketone were obtained from reaction of L1 with CuBr2 in CD3OD under 18O2. These results indicate that the origins of the O atom in L1OH and L1OMe are O2 and MeOH, respectively. On the basis of these results, a mechanism of the oxygenation of L1 in the present system will be proposed. PMID- 11272530 TI - Polynuclear magnesium and magnesium-titanium species. Syntheses and crystal structures of [Mg4(mu3,eta2-ddbfo)2(mu,eta2-ddbfo)2(mu,eta1-ddbfo)29eta1 ddbfo2],. AB - Tetranuclear magnesium complexes with chelating alkoxo ligands have been synthesized with the aim of investigating coordinatively unsaturated magnesium sites able to bind TiX4 (X = Cl, OR), of the type necessary for the formation of the active centers in polymerization catalysts. The magnesium compound [Mg4(mu3,eta2-ddbfo)2(mu,eta2-ddbfo)2(mu,eta1-ddbfo)2(eta1-ddbfo)2] x 2CH2Cl2 (1) (ddbfo = 2,3-dihydro-2,2-dimethyl-7-benzofuranoxide) was prepared by the reaction of MgBu2 with ddbfoH in dichloromethane. Complex 1 exists as a centrosymmetric tetranuclear species with two different types of magnesium centers corresponding to octahedral MgO6 and trigonal bipyramidal MgO5 geometry. Compound 1 is monoclinic, space group P2(1/c), with a = 12.053(2) A, b = 13.323(3) A, c = 17.069(3) A, beta = 98.50(3) degrees , and Z = 4. The reaction of 1 with methanol in tetrahydrofuran (THF) gave compound [Mg4(mu3-OMe)2(mu,eta2-ddbfo)2(mu,eta1 ddbfo)2(eta1-ddbfo)2(CH3OH)5] x CH3OH x THF (2). During this reaction one of the two five-coordinate MgO5 centers in 1 is completed by a methanol molecule and becomes octahedral in 2. Species 2 belongs to the P2(1/n) monoclinic space group, with a = 13.323(3) A, b = 20.768(4) A, c = 27.584(6) A, beta = 104.26(3) degrees , and Z = 4. Compound [Mg4(mu3,eta2-thffo)2(mu,zeta2-thffo)2(mu,eta1-thffo)2[mu OTi(DIPP)3]2] x 2CH2Cl2 (3) is formed as a result of substitution of two thffo (thffo = 2-tetrahydrofurfuroxide) ligands bonded to the five-coordinate magnesium atom in [Mg4(thffo)8] by bulky OTi(DIPP)3 (DIPP = diisopropylphenolate) groups. Crystals of 3 are monoclinic, space group P2(1/n), with a = 17.069(3) A, b = 18.421(4) A, 17.815(4) A, beta = 90.77(3) degrees , and Z = 4. The X-ray crystal structures of complexes 1-3 are discussed in terms of explaining the role of the coordinatively unsaturated magnesium site in chiral catalyst active center formation. PMID- 11272531 TI - Pentasulfide S5(2-) as the first tridentate chelating ligand in Ru(P(OE)3)3S5(E = Me and Et). AB - Room temperature stirring of H2Ru(P(OE)3)4 (E = Me and Et) and elemental sulfur in benzene afforded the optically active compounds Ru(P(OMe)3)3S5 (1) and Ru(P(OEt)3)3S5 (2). Compounds 1 and 2 are crystallized in the trigonal space group P31 with a = 14.231(10) A, c = 10.24(1) A, V = 1794(2) A3, and Z = 3, and orthorhombic space group P212121 with a = 15.393(5) A, b = 18.126(6) A, c = 12.421(4) A, V = 3465(1) A3, and Z = 4, respectively. Solutions of 1 and 2 did not show any optical activity since the bulk materials are racemic mixtures. The X-ray analyses also reveal that in both compounds polysulfide S5(2-) ion acts as a novel tridentate ligand, resulting in an asymmetric bicyclic RuS5 unit having three- and five-membered rings around the ruthenium atom. Fragmentation of the S5(2-) ring to S2- ion was observed in the presence of sulfur-abstracting reagents such as PR3 (R = Ph, OMe, and OEt) and also to S2(2-) ion when the compounds were reacted with RuCl2(P(OE)3)4 (E = Me and Et). PMID- 11272532 TI - Optical spectroscopy and density functional calculations of chromium(V)-doped YVO4 and YPO4: influence of the second coordination sphere. AB - Low temperature polarized single-crystal absorption and luminescence spectra of Cr(V)-doped YVO4 and the powder luminescence spectrum of Cr(V)-doped YPO4 are reported and discussed. A rich fine structure and strong polarization effects are observed in the near-infrared. Due to a strong interaction of the Cr(V) ion with two Y3+ ions in the second coordination sphere of the Cr(V) ion, the electronic ground state is different from the one expected on the basis of an angular overlap calculation in which only the four oxygen ligands are taken into account. This effect of the Y3+ ions on the ground state of Cr(V) is confirmed by a density functional calculation and by literature EPR data. CrO4(3-) bending modes are responsible for the fine structure in the d-d transition and the resulting distortion in the emitting excited state. PMID- 11272533 TI - Luminescent mononuclear and binuclear cyclometalated palladium(II) complexes of 6 phenyl-2,2 bipyridines: spectroscopic and structural comparisons with platinum(II) analogues. AB - The mononuclear cyclometalated Pd(II) complexes [Pd(L1)X] (HL1 = 6-phenyl-2,2' bipyridine; X = Cl, la; Br, 1b; I, 1c), [Pd(L1)PPh3]+ (1d), [Pd(L2-5)Cl] [2a-5a, HL2-5 = 4-(aryl)-6-phenyl-2,2'-bipyridine; aryl = phenyl (2), 4-chlorophenyl (3), 4-tolyl (4), 4-methoxyphenyl (5)] and the binuclear derivatives [Pd2(L1-5)2(mu dppm)]2+ (1e-5e, dppm = bis(diphenylphosphino)methane) and [Pd2(L1)2(mu dppCs)]2+, (1f, dppC5 = 1,5-bis(diphenylphosphino)pentane) were prepared. The crystal structures of 1d(ClO4), 1e(ClO4)2 x DMF, and 2e(ClO4)2 have been determined by X-ray crystallography. The magnitude of the Pd-Pd distances in le and 2e (3.230(1) and 3.320(2) A, respectively) suggest minimal metal-metal interaction, although pi-stacking of the aromatic ligands (interplanar separations 3.34 and 3.35 A, respectively) is evident. All complexes display low energy UV absorptions at lambda approximately 390 nm, which are tentatively assigned to 1MLCT transitions; red shifts resulting from Pd-Pd interactions in the binuclear species are not apparent. The complexes in this work are non emissive at 298 K, but the cationic derivatives exhibit intense luminescence at 77 K. The structured emissions of 1d and 1f in MeOH/EtOH glass (lambdamax 467-586 nm) and all cationic species in the solid state (lambdamax 493-578 nm) are assigned to intraligand excited states. Complexes le-5e display dual emissions in MeOH/EtOH glass at 77 K, and the broad structureless bands at lambdamax 626-658 nm are attributed to pi-pi excimeric IL transitions. A comparison between the photophysical properties of Pd(II) and Pt(II) congeners is presented. PMID- 11272535 TI - Electrochemical properties of vanadium(III,IV,V)-salen complexes in acetonitrile. Four-electron reduction of O2 by V(III)-salen. AB - The coordination chemistry and electrochemistry of complexes of vanadium(III,IV,V) with salen (H2 salen = N,N'-ethylenebis(salicylideneamine) were reexamined in an attempt to uncover the origin of puzzling results reported in a previous study (Inorg. Chem. 1994, 33, 1056). Microelectrodes were utilized to allow measurements in the absence of supporting electrolyte. The source of the puzzling results was identified and the modifications required in the previous interpretations are specified. Corrected values of formal potentials and diffusion coefficients are also given. The acid-induced disproportionation of V(IV)O(salen), as originally proposed by Bonadies et al. (J. Chem. Soc., Chem. Commun. 1986, 1218; Inorg. Chem. 1987, 26, 1218), was largely supported by the present results. The equilibrium constant for this disproportionation reaction was measured. The stoichiometry and kinetics of the reaction between O2 and the V(III)-salen complex were examined, and a possible mechanism for this four electron reduction of O2 is suggested. PMID- 11272534 TI - Synthesis and structures of bis(dithiolene)molybdenum complexes related to the active sites of the DMSO reductase enzyme family. AB - Structural analogues of the reduced (Mo(IV)) sites of members of the DMSO reductase family of molybdoenzymes are sought. These sites usually contain two pterin-dithiolene cofactor ligands and one protein-based ligand. Reaction of [Mo(MeCN)3(CO)3] and [Ni(S2C2R2)2] affords the trigonal prismatic complexes [Mo(CO)2(S2C2R2)2] (R = Me (1), Ph (2)), which by carbonyl substitution serve as useful precursors to a variety of bis(dithiolene)molybdenum-(IV,V) complexes. Reaction of 1 with Et4NOH yields [MoO(S2C2Me2)2]2- (3), which is readily oxidized to [MoO(S2C2Me2)2]1- (4). The hindered arene oxide ligands ArO- afford the square pyramidal complexes [Mo(OAr)(S2C2R2)2]1- (5, 6). The ligands PhQ- affordthe trigonal prismatic monocarbonyls [Mo(CO)(QPh)(S2C2Me2)2]1- (Q = S (8), Se (12)) while the bulky ligand ArS- forms square pyramidal [Mo(SAr)(S2C2R2)2]- (9, 10). In contrast, reactions with ArSe- result in [Mo(CO)(SeAr)(S2C2R2)2]1-(14, 15), which have not been successfully decarbonylated. Other compounds prepared by substitution reactions of 1 and 2 include the bridged dimers [Mo2(mu Q)2(S2C2Me2)4]2- (Q = S (7), Se (11)) and [Mo2(mu-SePh)2(S2C2Ph2)4]2- (13). The complexes 1, 3-5, 7-10, 12-14, [Mo(S2C2Me2)3] (16), and [Mo(S2C2Me2)3]1- (17) were characterized by X-ray structure determinations. Certain complexes approach the binding arrangements in at least one DMSO reductase (5/6) and its Ser/Cys mutant, and in dissimilatory nitrate reductases (9/10). This investigation provides the initial demonstration of the new types of bis(dithiolene)molybdenum(IV) complexes available through [Mo(CO)2(S2C2R2)2] precursors, some of which will be utilized in reactivity studies. (Ar = 2,6 diisopropylphenyl or 2,4,6-triisopropylphenyl.) PMID- 11272536 TI - High-frequency EPR study of the ferrous ion in the reduced rubredoxin model. AB - High-frequency (94-371 GHz) EPR data are reported for powdered samples of [PPh4]2[Fe(SPh)4], an accurate model for the reduced site of rubredoxins. This is the first HFEPR investigation of an S = 2 ferrous complex, illustrating the utility of this technique for the investigation of integer-spin systems. A full matrix diagonalization approach is used to simulate spectra over the 94-371 GHz frequency range, providing the spin-Hamiltonian parameters g, D, and E. It is observed that g is anisotropic, characterized by gx = gy = 2.08 and gz = 2.00, and that D = +5.84 cm(-1) and E = +1.42 cm(-1), where the uncertainty in each parameter is estimated as +/- 2%. The spin-Hamiltonian for [PPh4]2[Fe(SPh)4] is related to fundamental properties, such as the crystal-field splitting and the spin-orbit coupling of Fe2+. It is shown that the conventional spin-Hamiltonian accurately represents the electronic structure of the Fe2+ ion in this molecule. Through a comparison with Fe(SPh)4(PPh4)2, the zero-field splitting of the Fe2+ site in reduced rubredoxin is estimated to be D = +5.3 cm(-1) and E = +1.5 cm( 1). This is one of the few HFEPR investigations of a rhombic, high-spin system; as such, it is a step toward the eventual investigation of similar Fe2+ sites in proteins. PMID- 11272537 TI - Catalytic two-electron reductions of N2O and N3- by myoglobin in surfactant films. AB - Myoglobin (Mb), in films of dimethyldidodecylammonium bromide (ddab) on graphite electrodes, is used as a catalyst to mediate the electrochemical reduction of nitrous oxide (N2O) as well as the isoelectronic ion azide (N3-) in aqueous solutions. The electrocatalytic reductions are characterized by a rate-dependent irreversibility in cyclic voltammograms of Mb/ddab in the presence of the substrates. Bulk electrolysis shows that the reduction of 15N15NO by Mb/ddab yields 15N15N as shown by GC/MS. The catalytic reduction of azide results in almost quantitative formation of ammonia. These electrocatalytic processes are rationalized as two-electron reductions, with the catalyst cycling between the Fe(I) and Fe(III) states of Mb. To our knowledge, this is the first characterization of N2O reduction by an Fe porphyrin or heme protein. PMID- 11272538 TI - Understanding the orientation and dynamic motion of planar heterocyclic N-donor ligands by exploiting the symmetry properties of mixed-ligand mu-oxorhenium(V) dinuclear complexes. AB - Factors influencing the orientation and dynamic motions of planar N-donor heterocyclic ligands (L) are of interest since such features have broad relevance in metallobiochemistry [Marzilli, L. G.; Marzilli, P. A.; Alessio, E. Pure Appl. Chem. 1998, 70, 961-968]. We found that mu-oxorhenium(V) dinuclear complexes [ReOCl2LsLt]-O-[ReOCl2LsLt] bearing either symmetrical (L = py = pyridine; 3,5 lut = 3,5-lutidine) or lopsided (L = Me3-Bzm = 1,5,6-trimethylbenzimidazole) cis L ligands are particularly useful for studying these factors. NMR data showed that terminal (Lt) and stacked (Ls) ligands were exchanged by approximately 180 degrees rotation about the Re-O-Re bond system. Such exchange occurred, however, between degenerate chiral conformers. Here we report a combined X-ray structural and solution NMR investigation of the AA + CC (racemic) and AC (meso) forms of two mixed-ligand mu-oxorhenium dimers that bear one lopsided and one symmetrical ligand on each Re atom, namely, Re2O3-Cl4(py)2(Me3Bzm)2 (1rac and 1meso) and Re2O3Cl4(3,5-lut)2(Me3Bzm)2 (2rac and 2meso). The presence of two different cis L ligands in 1 and 2 breaks the local symmetry at each Re atom, so that, in the racemic dimers, the exchange of terminal and stacked ligands leads to nondegenerate conformers. Overall, NMR data showed that the unsymmetrical dimers 1 and 2 undergo two dynamic processes contemporaneously, namely, 180 degrees rotation about the Re-N(py or 3,5-lut) bond and coupled rotation about the Re-O Re/Re-N bonds. Both processes reach the slow exchange limit below -80 degrees C. Rotation of py in 1 occurs faster than that of 3,5-lut in 2; this difference is attributed to the higher steric demands of 3,5-lut compared to py. For both dimers NMR data provided compelling evidence of the preferred conformers in solution, including ligand orientations. The low-T solution structure of 1meso and 2meso is chiral, the same as that found in the solid state for 2meso, where the Me3Bzm on one Re atom is stacked with the 3,5-lut on the other Re atom. The remaining Me3Bzm and 3,5-lut, one on each Re atom, are both terminal. In solution the coupled Re-O-Re/Re-N rotations interconvert the two halves of each meso dimer to yield the same overall stable chiral conformation. For the racemic dimers, however, this process does not interconvert one enantiomer into the other, but instead interconverts two rotamers, R1 and R2, each of which is chiral. We found that, in the case of both 1rac and 2rac, the conformer with stacking symmetrical ligands (R1) is roughly 1 order of magnitude more stable than that with stacking Me3Bzm ligands (R2). Moreover, the solution conformation of R1 is the same as that found in the solid state of 1rac. Solution- and solid-state data indicate that the key interaction favoring the observed conformations is very likely the electrostatic attraction between the delta+ H2 atoms on the Me3Bzm ligands and the negative O and Cl groups in the core of the dimers. Finally, for both meso and racemic dimers we were also able to elucidate the preferred pathways of the coupled dynamic motions and establish that, very likely, the two halves of the dimers swing back and forth by approximately 130 degrees through the anti eclipsed form. PMID- 11272539 TI - Mechanism of CO exchange on cis-[M(CO)2X2]- complexes (M=Rh,Ir;X=Cl, Br, or I). AB - The CO exchange on cis-[M(CO)2X2]- with M = Ir (X = Cl, la; X = Br, 1b; X = I, 1c) and M = Rh (X = Cl, 2a; X = Br, 2b; X = I, 2c) was studied in dichloromethane. The exchange reaction [cis-[M(CO)2X2]- + 2*CO is in equilibrium cis-[M(*CO)2X2]- + 2CO (exchange rate constant: kobs)] was followed as a function of temperature and carbon monoxide concentration (up to 6 MPa) using homemade high gas pressure NMR sapphire tubes. The reaction is first order for both CO and cis-[M(CO)2X2]- concentrations. The second-order rate constant, k2(298) (=kobs)[CO]), the enthalpy, deltaH*, and the entropy of activation, deltaS*, obtained for the six complexes are respectively as follows: la, (1.08 +/- 0.01) x 10(3) L mol(-1) s(-1), 15.37 +/- 0.3 kJ mol(-1), -135.3 +/- 1 J mol(-1) K(-1); 1b, (12.7 +/- 0.2) x 10(3) L mol(-1) s(-1), 13.26 +/- 0.5 kJ mol(-1), -121.9 +/- 2 J mol(-1) K(-1); 1c, (98.9 +/- 1.4) x 10(3) L mol(-1) s(-1), 12.50 +/- 0.6 kJ mol(-1), -107.4 +/- 2 J mol(-1) K(-1); 2a, (1.62 +/- 0.02) x 10(3) L mol(-1) s( 1), 17.47 +/- 0.4 kJ mol(-1), -124.9 +/- 1 J mol(-1) K(-1); 2b, (24.8 +/- 0.2) x 10(3) L mol(-1) s(-1), 11.35 +/- 0.4 kJ mol(-1), -122.7 +/- 1 J mol(-1) K(-1); 2c, (850 +/- 120) x 10(3) L mol(-1), s(-1), 9.87 +/- 0.8 kJ mol(-1), -98.3 +/- 4 J mol(-1) K(-1). For complexes la and 2a, the volumes of activation were measured and are -20.9 +/- 1.2 cm3 mol(-1) (332.0 K) and -17.2 +/- 1.0 cm3 mol(-1) (330.8 K), respectively. The second-order kinetics and the large negative values of the entropies and volumes of activation point to a limiting associative, A, exchange mechanism. The reactivity of CO exchange follows the increasing trans effect of the halogens (Cl < Br << I), and this is observed on both metal centers. For the same halogen, the rhodium complex is more reactive than the iridium complex. This reactivity difference between rhodium and iridium is less marked for chloride (1.5: 1) than for iodide (8.6:1) at 298 K. PMID- 11272540 TI - Novel synthetic routes to several new, differentially substituted ruthenium tris(4,4 disubstituted-2,2-bipyridine) complexes. AB - The stepwise synthesis of several novel Ru(tris(pp)) complexes (pp = 4,4' disubstituted-2,2'-bipyridine; substituent = H, Me, chiral ester, or chiral amide) is described, where the pp ligands may be the same, or different, in each complex. All of the complexes detailed have been resolved into their pure delta- and lambda-enantiomers or diastereomers. The complexes, which are prepared starting from RuCl3, contain novel ligand architectures, with a range of chiral esters and amides attached to the 4,4'-positions of the bpy ligands. It was postulated that these chiral groups would be capable of inducing chirality at the metal center, but our investigations have shown this not to be the case, and in all reactions completely racemic products were formed. Resolution by chiral HPLC, and the subsequent characterization of the products through NMR, UV-vis, and circular dichroism (CD) spectroscopy, has been carried out; the characteristics of the CD spectra have been discussed with respect to the electron-donating/ withdrawing ability of the groups at the 4,4'-positions. The X-ray crystal structure of the optically pure complex lambda-[Ru(dmbpy)2(4,4'-bis((R)-(+)-alpha phenylethylamido)-2,2'-bipyridine)] x 2PF6 x 2CHCl3 was obtained and solved using direct methods. This result, in conjunction with the CD spectra, enabled the complete and unambiguous assignment of the stereocenters of all of the novel Ru(tris(bpy)) complexes prepared in this investigation. PMID- 11272541 TI - Diastereoselective preparation and characterization of ruthenium bis(bipyridine) sulfoxide complexes. AB - A new concept in the synthesis of optically active octahedral ruthenium complexes was realized for the first time when cis- or trans-Ru(bpy)2Cl, (cis- or trans-1) was reacted with either (R)-(+)- or (S)-(-)-methyl p-tolyl sulfoxide (2 or 3); this novel asymmetric synthesis leads to the diastereoselective formation of the ruthenium bis(bipyridine) complex cis-delta-[Ru(bpy)2(2)Cl]Cl (4) (49.6% de) or cis-lambda-[Ru(bpy)2(3)Cl]Cl (5) (48.4% de), respectively. cis- or trans Ru(dmbpy)2Cl2 (cis- or trans-6) (dmbpy = 4,4'-dimethyl-2,2'-bipyridine) also reacts with 2 or 3, leading to the diastereoselective formation of cis-delta [Ru(dmbpy)2(2)Cl]Cl (7) (59.5% de) or cis-lambda-[Ru(dmbpy)2(3)Cl]Cl (8) (57.2% de), respectively. The diastereoselectivity of these reactions is governed solely by the chirality of the sulfoxide nucleophile. This represents the first process by which a sigma-bonded ligand occupying only a single coordination site has had such an important influence on the stereochemical outcome of a ruthenium bis(bipyridine) complex formation. These novel complexes were fully characterized by elemental analysis and IR, UV/vis, and 1H, 13C, and 2D NMR spectroscopy. An investigation into the chiroptical properties of these novel ruthenium bis(bipyridine) sulfoxide complexes has been carried out, and circular dichroism spectra are used to assign absolute stereochemistry. PMID- 11272542 TI - Charge effects on oxygen atom transfer. AB - The rhenium(V) complex [(HCpz3)ReOCl2]+ ([1]+), the tris(pyrazolyl)methane analogue of the known tris(pyrazolyl)borate complex (HBpz3)ReOCl2 (2), has been prepared. The two complexes are strikingly similar, as are the phosphine oxide adducts [(HCpz3)ReCl2(OPPh3)]Cl ([3]Cl) and (HBpz3)ReCl2(OPPh3) (4), which have been characterized by X-ray crystallography. Comparison of the bimolecular reduction of [1]BF4 and 2 by triarylphosphines reveals a pronounced charge effect, with the cationic species being reduced by PPh3 about 1,000 times faster than its neutral analogue in CH2Cl2 at room temperature. Ligand substitution of the adducts [3]+ and 4 is dissociative, with the cationic complex dissociating phosphine oxide about 56 times more slowly than the neutral compound. The relative impact of charge on ground and transition states in atom transfer reactions is discussed. PMID- 11272543 TI - Synthesis, characterization, and chiral properties of CoIII2AgI3 pentanuclear, CoIII4ZnII4 octanuclear, and CoIII mononuclear complexes with aza-capped hexadentate-N3S3 thiolate ligands: crystal structures of. AB - The reaction of an S-bridged Co2(III)Ag3(I) pentanuclear complex, [Ag3[Co(aet)3]2][BF4]3 (aet = NH2CH2CH2S-), with paraformaldehyde in basic acetonitrile, followed by adding aqueous ammonia, produced an aza-capped Co2(III) Ag3(I) complex, [Ag3[Co(L)]2]3+ ([1]3+) (L = N(CH2NHCH2CH2S-)3). The crystal structure of [1]3+ was determined by X-ray crystallography. [1][PF6]3 x H2O, empirical formula C18H44Ag3Co2F18N8OP3S6, crystallizes in the tetragonal space group 142m with a = 13.012(1) A, c = 24.707(2) A, and Z = 4. In [1]3+ the two aza capped [Co(L)] units are linked by three Ag(I) atoms, such that the two Co(III) atoms are encapsulated in a macrobicyclic metallocage, [Ag3(I)(L)2]3-. [1]3+ was converted to an aza-capped Co4(III)Zn4(II) octanuclear complex, [Zn4O[Co(L)]4]6+ ([2]6+), by reaction with I- in the presence of Zn2+ and ZnO in water. The crystal structure of [2]6+ was also determined by X-ray crystallography. [2][PF6]6 x 8H2O, empirical formula C36H100Co4F36N16O9P6S12Zn4, crystallizes in the monoclinic space group P2(1/n) with a = 14.33(7) A, b = 25.67(10) A, c = 24.83(6) A, beta = 101.3(3) degrees , and Z = 4. In [2]6+ each of four [Co(L)] units is bound to each trigonal Zn3(II) face of the tetrahedral [Zn4(II)O]6+ core, such that each Co(III) atom is encapsulated in a macrobicyclic [Zn4(II)O(L)] fragment. Treatment of [2]6+ with a basic aqueous solution resulted in a cleavage of the Zn-S bonds to produce an aza-capped Co(III) mononuclear complex, [Co(L)] ([3]), from which [1]3+ is readily reproduced by the reaction with Ag+ in water. All the reactions were found to proceed with retention of the absolute configuration (delta or lambda) of the Co(III) chiral centers; deltadelta-[1]3+, deltadeltadeltadelta-[2]6+, and A-[3] were derived from deltadelta-[Ag3[Co(aet)3]2]3+. The contributions to circular dichroism (CD) from the triple helicity in [1]3+, besides from the asymmetric N and S donor atoms and the Co(III) chiral centers in [1]3+ and [2]6+, were estimated by comparing the CD spectra of deltadelta-[1]3+, deltadeltadeltadelta-[2]6+, and delta-[3]. PMID- 11272544 TI - Neocuproine-extended porphyrin coordination complexes. 2. Spectroscopic properties of the metalloporphyrin derivatives and investigations into the HOMO ordering. AB - The synthesis of a porphyrin compound, 1, containing a 2,9-dimethyl-1,10 phenanthroline moiety that is fused at the beta-pyrrole positions is reported. The absorption spectra of the free-base, copper(II), and zinc(II) derivatives have been studied. On the basis of absorption band intensities, the HOMO of the free base (H21) and its copper and zinc complexes (Cu1 and Zn1) was determined to be of a1u symmetry. Relative to H21, compounds Cul and Znl show enhanced spectral changes upon external metal ion binding. Although the HOMO is the same in all three compounds, the energy gap between the two highest occupied orbitals is greater for Cu1 and Zn1 than it is for the free-base compound. Several metal ions (Ni2+, Cu+, Cu2+, Zn2+, Li+) were examined in their binding to the phenanthrolinic group by measuring the resulting changes in the absorption spectra. It is shown that the observed changes in the absorption spectra are insensitive to the nature of the metal ion coordinated by the phenanthroline moiety. Significant differences in the absorption and emission spectra between Zn1 and [Zn(Zn1)2]2+ clearly demonstrate that the porphyrin pi-system is strongly affected by the binding of metal ions at the fused phenanthrolinic moiety. PMID- 11272545 TI - Toward ligand identification within a CCHHC zinc-binding domain from the NZF/MyT1 family. AB - A family of proteins that contain presumed zinc-binding domains with the consensus sequence Cys-X4-CysX4-His-X7-His-X5-Cys has recently been identified, but the metal binding and structural properties of these domains have not been investigated. This consensus is striking because of the presence of five conserved potential zinc-binding residues. A peptide corresponding to the third putative zinc-binding domain from the transcription factor NZF-1 (hereafter NZF 13) has been synthesized and characterized. The UV-visible absorption spectroscopic properties of the cobalt(H) complex of this peptide demonstrate that metal binding is tetrahedral, and the position of the visible absorption bands suggests coordination by three cysteinates and one histidine. To identify which of the two conserved histidine residues acts a metal-binding residue, two histidine to alanine variant peptides were also synthesized. Both variant peptides bound cobalt(II) in a tetrahedral fashion; replacement of the first of the two histidines has a somewhat larger effect on the detailed shape of the absorption spectral features than does replacement of the second histidine. These results suggest that the metal-coordinating residues (italicized) are Cys-X4-Cys X4-His-X7-His-Xs-Cys. However, simultaneous substitution of both histidine residues with alanine generated a peptide with much more dramatically affected metal binding properties. These observations suggests that the relatively modest effects observed for the singly substituted peptides may be due to metal interactions involving the remaining histidine. Because of these phenomena, further studies will be required to establish more conclusively the roles of the two histidine residues in metal binding and the potential significance of the apparent alternative histidine coordination. PMID- 11272546 TI - Electron transfer. 140. Reactions of riboflavin with metal center reductants. AB - Riboflavin (I) is reduced in separable steps by indium(I), vanadium(II), europium(II), and titanium(III) in 0.02-1.0 M H+, yielding first the radical ion, II (lambdamax = 495 nm), and then the dihydro compound, III. The initial reduction with InI yields 2 equiv of the radical, but kinetic profiles exhibit no irregularity due to intervention of In(II), indicating that participation by the dipositive state is much more rapid than the In(I) reaction. Predominant paths involve the protonated form of the flavin, RbH+, and that of the radical, RbH2.+. Formation of the radical with excess V(II) and Ti(III) (but not with In(I)) is strongly autocatalytic, reflecting rapid comproportionation involving the flavin and the dihydro compound. The V(II) and Ti(III) rates for both steps greatly exceed the substitution-controlled limits for these states and therefore pertain to outer-sphere processes. The very high ratio kEu/kv for the first step, however, points to an inner-sphere reduction by the lanthanide cation. A kinetic inversion is observed for In(I) (kRbH.+ > kRbH2.+), implying a bridged reduction path for the initial step with this center as well. PMID- 11272548 TI - Addition of ammonia to AlH3 and BH3. Why does only aluminum form 2:1 adducts? AB - The electronic structures of the mono- and bisammonia adducts EH3NH3 and EH3(NH3)2, E = B and Al, have been investigated using ab initio electronic structure methods. Geometries were optimized at the MP2/cc-pVTZ level. Higher level correlated methods (MP4(SDTQ), QCISD(T), CCSD(T)), as well as the G2 and CBS-Q methods, were used to obtain accurate bond dissociation energies. The E-N bond dissociation energy (De) is computed near 33 kcal/mol (E = B) and 31 kca/mol (E = Al), respectively. Whereas the Al-N bond energy pertaining to the second ammonia molecule in AlH3(NH3)2 is 11-12 kcal/mol, only a transition-state structure may be located for the species BH3(NH3)2. We analyze factors which may distinguish Al from B with respect to the formation of stable bisamine adducts. The most significant difference relates to electronegativity and hence the propensity of boron to engage in predominantly covalent bonding, as compared with the bonding of aluminum with ammonia, which shows substantial electrostatic character. Neither steric factors nor the participation of d-orbitals is found to play an important role in differentiating aluminum from boron. The lesser electronegativity of third-row elements appears to be the critical common feature allowing the formation of hypercoordinate complexes of these elements in contrast to their second-row analogues. Consideration of some group 14 analogues and hard/soft acid/base effects supports this view. PMID- 11272547 TI - Co(III) complexes with coordinated carboxamido nitrogens and thiolato sulfurs as models for Co-containing nitrile hydratase and their conversion to the corresponding sulfinato species. AB - Recent spectroscopic data suggest that the Co(III) site in Co-containing nitrile hydratase is ligated to carboxamido nitrogens and thiolato sulfurs and most possibly one or more of the bound thiolates exist as sulfenato and/or sulfinato groups. The absence of any Co(III) complex with such coordination makes it quite difficult to predict the reactivity of this kind of Co(III) site. In this paper, the Co(III) complexes of two designed ligands PyPepSH2 (1) and PyPepRSH2 (2) have been reported. The two complexes, namely, (Et4N)[Co(PyPepS)2] (3) and Na[Co(PyPepRS)2] (4) are the first examples of Co(III) complexes with carboxamido nitrogens and thiolato sulfurs as donors. The average Co(III)-Namido and Co(III) S distances in these complexes lie in the range 1.90-1.92 and 2.22-2.24 A, respectively. Reaction of H2O2 with both complexes readily affords Na[Co(PyPepSO2)2] (5) and Na[Co(PyPepRSO2)2] (6), species in which the thiolato sulfurs are converted to sulfinato (SO2) groups. Such conversion also occurs when solutions of 3 and 4 are exposed to dioxygen in the presence of activated charcoal. These reactions are clean and the S --> SO2 transformation does not introduce significant changes in the metric parameters of these complexes. The reactivity of 3 and 4 indicates that the bound Cys-sulfurs around the biological Co(III) site could be oxidized to sulfinato groups. PMID- 11272549 TI - Electronic absorption and MCD spectra for Pt(AuPPh3)8(2+) and Au(AuPPh3)8(3+) cluster complexes in poly(methyl methacrylate) thin films at 295 and 10 K. AB - Electronic absorption and 8 T magnetic circular dichroism (MCD) spectra are reported for nitrate salts of Pt(AuPPh3)8(2+) and Au(AuPPh3)8(3+) in poly(methyl methacrylate) (PMM) thin films at 295 and 10 K in the vis-UV region from 1.6 to 3.6 microm(-1) (1 microm(-1) = 10(4) cm(-1). Enhanced resolution is observed at low temperature, especially for Pt(AuPPh3)8(2+), which emphasizes the differences in the nature of the low-energy excited configurations and states between Pt(AuPPh3)8(2+) and Au(AuPPh3)8(3+). The absorption and MCD spectra for Pt(AuPPh3)8(2+) are interpreted in terms of a combination of excitations from filled Pt 5d orbitals to empty Au framework 6s orbitals and intraframework Au8(2+) (IF) transitions, whereas the spectra for Au(AuPPh3)8(3+) are ascribed entirely to Au IF transitions. PMID- 11272550 TI - A novel synthesis of hexakis(trifluoromethyl)cyclotriphosphazene. Single-crystal X-ray structures of N3P3(CF3)6 and N3P3F6. AB - Reaction of hexafluorocyclotriphosphazene (N3P3F6) with trimethyl(trifluoromethyl)silane in the presence of a catalytic amount of cesium fluoride in THF produced hexakis(trifluoromethyl)cyclotriphosphazene [N3P3(CF3)6] in 90% isolated yield. N3P3(CF3)6 is fully characterized by melting point, IR, NMR (19F, 13C, 31p), MS, and elemental analysis data. Single-crystal X-ray structures of N3P3(CF3)6 and N3P3F6 are reported. PMID- 11272551 TI - Oxidative azavinylidene formation in the reaction of 1,3-diphenylisobenzofuran with osmium nitride complexes. PMID- 11272552 TI - Comparing the isoelectronic complexes [RuTp(CH3CN)3]PF6 (Tp = hydridotris(pyrazolyl)borate) and. PMID- 11272553 TI - After more than 60 years, a new NaTl type Zintl phase:KTl at high pressure. PMID- 11272554 TI - Synthesis and structural characterization of a 4-coordinate molybdenum(VI) dioxo diaryloxide, MoO2(O-2,6-t-Bu2C6H3)2.HO-2,6-t-Bu2C6H3. PMID- 11272555 TI - Reactivity of the iron porphyrin Fe(TPP)(NO) with excess NO. Formation of Fe(TPP)(NO)(NO2) occurs via reaction with trace NO2. PMID- 11272557 TI - Ammonolysis of mono(pentamethylcyclopentadienyl) titanium(IV) derivatives. AB - Ammonolyses of mono(pentamethylcyclopentadienyl) titanium(IV) derivatives [Ti(eta5-C5Me5)X3] (X = NMe2, Me, Cl) have been carried out in solution to give polynuclear nitrido complexes. Reaction of the tris(dimethylamido) derivative [Ti(eta5-C5Me5)(NMe2)3] with excess of ammonia at 80-100 degrees C gives the cubane complex [[Ti(eta5-C5Me5)]4(mu3-N)4] (1). Treatment of the trimethyl derivative [Ti(eta5-C5Me5)Me3] with NH3 at room temperature leads to the trinuclear imido-nitrido complex [[Ti(eta/5-CsMes)(mu-NH)]3(mu3-N)] (2) via the intermediate [[Ti(eta5-C5Me5)Me]2(mu-NH)2] (3). The analogous reaction of [Ti(eta5-C5Me5)Me3] with 2,4,6-trimethylaniline (ArNH2) gives the dinuclear imido complex [[Ti(eta5-C5Me5)Me])2(mu-NAr)2] (4) which reacts with ammonia to afford [[Ti(eta5-C5Me5)(NH2)]2(mu-NAr)2] (5). Complex 2 has been used, by treatments with the tris(dimethylamido) derivatives [Ti(eta5-C5H5-nRn)(NMe2)3], as precursor of the cubane nitrido systems [[Ti4(eta5-C5Me5)3(eta5-C5H5-nRn)](mu3-N)4] [R = Me n = 5 (1), R = H n = 0 (6), R = SiMe3 n = 1 (7), R = Me n = 1 (8)] via dimethylamine elimination. Reaction of [Ti(eta5-C5Me5)Cl3] or [Ti(eta5 C5Me5)(NMe2)Cl2] with excess of ammonia at room temperature gives the dinuclear complex [[Ti2(eta5-C5Me5)2Cl3(NH3)](mu-N)] (9) where an intramolecular hydrogen bonding and a nonlineal nitrido ligand bridge the "Ti(eta5-C5Me5)Cl(NH3)" and "Ti(eta5-C5Me5)Cl2" moieties. The molecular structures of [[Ti(eta5-C5Me5)Me]2 (mu-NAr)2] (4) and [[Ti2(eta5-C5Me5)2Cl3(NH3)](mu-N)] (9) have been determined by X-ray crystallographic studies. Density functional theory calculations also have been conducted on complex 9 to confirm the existence of an intramolecular N H...Cl hydrogen bond and to evaluate different aspects of its molecular disposition. PMID- 11272556 TI - Influence of carrier ligand NH hydrogen bonding to the O6 and phosphate group of guanine nucleotides in platinum complexes with a single guanine ligand. AB - Coordinated N,N',N"-trimethyldiethylenetriamine (Me3dien) has several possible configurations: two have mirror symmetry (R,S configurations at the terminal nitrogens) and the terminal N-Me's anti or syn with respect to the central N-Me (anti-(R,S) and syn-(R,S) isomers, respectively), and two are nonsymmetrical (R,R and S,S configurations at terminal nitrogens, rac denotes a 1:1 mixture of the two isomers). For each configuration, two Me3dienPtG atropisomers can be formed (anti or syn orientation of central N-Me and G 06, G = guanine derivative), and these can be observed since the terminal N-Me's decrease the rate of G rotation about the Pt-N7 bond. In symmetrical syn-(R,S)-Me3dienPtG derivatives with G = 9 EtG and 3'-GMP, the anti rotamer, which can form O6-NH H-bonds, was slightly favored over the syn rotamer but never more than 2:1. This anti rotamer is also favored by lower steric repulsion between the terminal N-Me's and G O6; thus, the contribution of O6-NH H-bonding to the stability of the anti rotamer could be rather small. With G = 5'-GMP, an O6-NH H-bond in the anti rotamer and a phosphate-NH H-bond in the syn rotamer can form. Only the syn rotamer was detected in solution, indicating that NH H-bonds to 5'-phosphate are far more important than to O6, particularly since steric factors favor the anti rotamer. Interconversion between rotamers was faster for syn-(R,S)- than for rac-Me3dien derivatives. This appears to be determined by a smaller steric impediment to G rotation of two "quasi equatorial" N-Me's, both on one side of the platinum coordination plane (syn-(R,S) isomer), than one "quasi equatorial" and one "quasi axial" N-Me on either side of the coordination plane (rac isomer). PMID- 11272558 TI - Electronic structure of [(CpCr)[(CO)3M]]mu-Cot (M = Cr, Fe): a theoretical study. AB - The electronic structure of two cyclooctatetraene-bridged dinuclear first-row transition metal complexes of the type [(CpM)[(CO)3M']]mu-Cot (M = Cr; M' = Fe (1), Cr (2)) was investigated by complete active space self-consistent field (CASSCF) calculations. In this context the differences in the binding capabilities of the complex fragments CpM and (CO)3M are discussed on the basis of extended Huckel molecular orbital (MO) calculations. The geometries used for the CASSCF calculations for complex 1 were obtained from the crystal structure. For 2 a model structure was established by geometry optimization using density functional methods. The CASSCF results agree well with the experimental findings and provide insight into the binding situation of the two compounds. Complex 1 can be regarded as being composed of a chromocene-like subunit CpCr(eta5-C5H5) and the fragment (CO)3Fe(eta3-C3H3). A direct metal-metal bond is found, involving one initially singly occupied orbital of each fragment, leading to a doublet ground state for 1 with the remaining unpaired electron localized at the chromium center. For 2 no such direct metal-metal bond can be recognized. A very weak direct metal-metal interaction is induced by electron donation from the Cot2 ligand into a formally unoccupied metal-metal binding orbital combination. In the quartet ground state all three unpaired electrons are localized at the chromium center of the formally doubly positive charged CpCr unit, on which complex fragment [(CO)3Cr(eta5-Cot)]2- acts like a cyclopentadienyl ligand. The coordination sphere of the chromium center of the CpCr unit resembles that of a metallocene metal center and its metal 3d occupation scheme corresponds to that of vanadocene. PMID- 11272559 TI - Trimerization of NaC2N3 to Na3C6N9 in the solid: ab initio crystal structure determination of two polymorphs of NaC2N3 and of Na3C6N9 from X-ray powder diffractometry. AB - Sodium dicyanamide NaC2N3 was found to undergo two phase transitions. According to thermal analysis and temperature-dependent X-ray powder diffractometry, the transition of alpha-NaC2N3 (1a) to beta-NaC2N3 (1b) occurs at 33 degrees C and is displacive. 1a crystallizes in the monoclinic system, space group P21/n (no. 14), with a = 647.7(1), b = 1494.8(3), c = 357.25(7) pm, beta = 93.496(1) degrees, and Z = 4. The structure was solved from powder diffraction data (Cu Kalpha1, T = 22 degrees C) using direct methods and it was refined by the Rietveld method. The final agreement factors were wRp = 0.072, Rp = 0.053, and RF = 0.074. 1b crystallizes in the orthorhombic system, space group Pbnm (no. 62), with a = 650.15(5), b = 1495.1(2), c = 360.50(3) pm, and Z = 4. The structure was refined by the Rietveld method using the atomic coordinates of 1a as starting values (Mo Kalpha1, T = 150 degrees C). The final agreement factors were wRp = 0.044, Rp = 0.034, RF = 0.140. The crystal structures of both polymorphs contain sheets of Na+ and N(CN)2- ions which are in la nearly and in 1b exactly coplanar. Above 340 degrees C, 1b trimerizes in the solid to Na3C6N9 (2). 2 crystallizes in the monoclinic system, space group P21/n (no. 14), with a = 1104.82(1), b = 2338.06(3), c = 351.616(3) pm, beta = 97.9132(9)degrees, and Z = 4. The structure was solved from synchrotron powder diffraction data (lambda = 59.733 pm) using direct methods and it was refined by the Rietveld method. The final agreement factors were wRp = 0.080, Rp = 0.059, and RF = 0.080. The compound contains Na+ and the planar tricyanomelaminate C6N9(3-). The phase transition from 1b to 2 is reconstructive. It occurs in the solid-state without involvement of other phases or intermediates. The crystal structures of 1b and 2 indicate that there is no preorientation of the N(CN)2- in the solid before their trimerization to C6N9(3 ). PMID- 11272560 TI - Crystal structure and replacement reaction of coordinated water molecules of the heteropoly compounds of sandwich-type tungstoarsenates. AB - Six new heteropoly compounds in the [M4(H2O)2(As2W15O56)2]16- series (M = CuII, MnII, CoII, NiII, ZnII, CdII), previously unknown, were synthesized and characterized by means of IR, UV-vis, CV, 183W NMR, TG-DSC, and elemental analyses. The synthetic method used in preparing this type of heteropoly compounds was different from that in preparing the corresponding tungstophosphates in that the starting materials were transition metal chlorides in 1.5 times the stoichiometric amount and the required pH value is lower than 2. The crystal structure of Na16[Cu4(H2O)2(As2W15O56)2].47H2O was solved in triclinic, P1 symmetry, with a = 12.721(3) A, b = 24.516(5) A, c = 26.450(5) A, alpha = 89.90(3) degrees, beta = 77.32(3) degrees, gamma = 89.96(3)degrees, V = 8048(3) A3, Z = 2, and R = 0.0966. This anion is isostructural with the previously reported [Cu4(H2O)2(P2W15O56)2]16-, having a rhombic tetrameric cluster Cu4O16 sandwiched by two trivacant Dawson-Wells anions [As2W15O56]12-. The range of the bond lengths of the equatorial Cu-O bonds is 1.83-2.05 A, while that of the axial Cu-O bonds is 2.30-2.39 A. The distortion of the Cu4O16 cluster is smaller in the As species than in the P species. Two copper atoms in the Cu4O16 cluster are coordinated by water molecules. The replacement reactions of the coordinated water molecules of this series of heteropoly compounds in aqueous solutions and in selected organic solvents are also reported here for the first time. The results show that [Fe(CN)6]4-, [Fe(CN)6]3-, H2NCH2CH2NH2, etc., can replace the coordinated water to form its characteristic color in aqueous solutions, while in organic solvents the coordinated water molecules are lost, leaving unshared coordination positions that can be occupied by some organic ligands such as pyridine, lactic acid, and acetone to restore the octahedral coordination of M2+. The crystallographic morphologies of this series of heteropolyanions after phase transfer are dependent on different transition metal ions present in the central M4O16 clusters although the anions are isostructural with each other. PMID- 11272562 TI - The related compounds MThTe3 (M = Mn, Mg) and ACuThSe3 (A = K, Cs): syntheses and characterization. AB - Single crystals of MnThTe3 (1) and MgThTe3 (2) grow as small black plates from the stoichiometric reaction of the elements, the former at 1,000 degrees C and the latter at 900 degrees C with the aid of a Sn flux. Both compounds crystallize in the space group Cmcm of the orthorhombic system with four formula units in cells of dimensions a = 4.2783(6) A, b = 13.8618(11) A, and c = 9.9568(15) A for 1 and a = 4.2854(6) A, b = 14.042(2) A, and c = 9.9450(14) A for 2 at T = 153(2) K. KCuThSe3 (3) forms as red blocks from a stoichiometric mixture of K2Se, Cu, Th, and Se at 800 degrees C, and CsCuThSe3 (4) forms as yellow blocks from a stoichiometric mixture of Cs2Se3, Cu, Th, and Se at 850 degrees C. Compounds 3 and 4 also crystallize in the space group Cmcm of the orthorhombic system with four formula units in cells of dimensions a = 4.1832(8) A, b = 14.335(3) A, and c = 10.859(2) A for 3 and a = 4.2105(7) A, b = 15.715(3) A, and c = 10.897(2) A for 4 at 153(2) K. Compounds 1 and 2 are isostructural with each other as well as with several uranium analogues and comprise pseudolayered structures with slabs of corner-shared MTe6 octahedra alternating with slabs of cap- and edge-shared ThTe8 bicapped trigonal prisms. The slabs are bonded together through the sharing of edges and vertices of the various polyhedra to form three-dimensional structures. Compounds 3 and 4 are two-dimensional layered structures that are closely related to 1 and 2. In 3 and 4, ThSe6 octahedra form the same slabs as MTe6 in 1 and 2 and Cu atoms occupy the tetrahedral holes in the layers. Alkali metal cations occupy bicapped trigonal prismatic sites between the layers. Neither structure type has short Q-Q interactions, and therefore the oxidation states of all atoms are straightforwardly assigned on the assumption of Th4+. Magnetic susceptibility measurements on compound 1 show a ferromagnetic transition at 70 K and a magnetic moment of 5.9(2) muB per Mn ion, indicating low spin Mn2+. PMID- 11272561 TI - Gold and silver complexes with the ferrocenyl phosphine FcCH2PPh2. AB - Linear gold(I) and silver(I) complexes with the ferrocenyl phosphine FcCH2PPh2 [Fc = (eta5-C5H5)Fe(eta5-C5H4)] of the types [AuR(PPh2CH2Fc)], [M(PPh3)(PPh2CH2Fc)]OTf, and [M(PPh2CH2Fc)2]OTf (M = Au, Ag) have been obtained. Three-coordinate gold(I) and silver(I) derivatives of the types [AuCl(PPh2CH2Fc)2] and [M(PPh2CH2Fc)3]X (M = Au, X = ClO4; M = Ag, X = OTf) have been obtained from the corresponding gold and silver precursors in the appropriate molar ratio, although some of them are involved in equilibria in solution. The crystal structures of [AuR(PPh2CH2Fc)] (R = Cl, C6F5), [AuL(PPh2CH2Fc)]OTf (L = PPh3, FcCH2PPh2), [Au(C6F5)3(PPh2CH2Fc)], and [Ag(PPh2CH2Fc)3]OTf have been determined by X-ray diffraction studies. PMID- 11272563 TI - Novel Mn(II)Mn(III)Mn(II) trinuclear complexes with carbohydrate bridges derived from seven-coordinate manganese(II) complexes with N-glycoside. AB - Reactions of MnX2.nH2O with tris(N-(D-mannosyl)-2-aminoethyl)amine ((D-Man)3 tren), which was formed from D-mannose and tris(2-aminoethyl)amine (tren) in situ, afforded colorless crystals of [Mn((D-Man)3-tren)]X2 (3a, X = Cl; 3b, X = Br; 3c, X = NO3; 3d, X = 1/2SO4). The similar reaction of MnSO4.5H2O with tris(N (L-rhamnosyl)-2-aminoethyl)amine ((L-Rha)3-tren) gave [Mn((L-Rha)3-tren)]SO4 (4d), where L-rhamnose is 6-deoxy-L-mannose. The structures of 3b and 4d were determined by X-ray crystallography to have a seven-coordinate Mn(II) center ligated by the N-glycoside ligand, (aldose)3-tren, with a C3 helical structure. Three D-mannosyl residues of 3b are arranged in a delta(ob3) configuration around the metal, leading to formation of a cage-type sugar domain in which a water molecule is trapped. In 4d, three L-rhamnosyl moieties are in a delta(lel3) configuration to form a facially opened sugar domain on which a sulfate anion is capping through hydrogen bonding. These structures demonstrated that a configurational switch around the seven-coordinate manganese(II) center occurs depending on its counteranion. Reactions of 3a, 3b, and 4d with 0.5 equiv of Mn(II) salt in the presence of triethylamine yielded reddish orange crystals formulated as [[Mn((aldose)3-tren)]2Mn(H2O)X3.nH2O (5a, aldose = D-Man, X = Cl; 5b, aldose = D-Man, X = Br; 6d, aldose = L-Rha, X = 1/2SO4). The analogous trinuclear complexes 6a (aldose = L-Rha, X = Cl), 6b (aldose = L-Rha, X = Br), and 6c (aldose = L-Rha, X = NO3) were prepared by the one-pot reaction of Mn(II) salts with (L-Rha)3-tren without isolation of the intermediate Mn(II) complexes. X-ray crystallographic studies revealed that 5a, 5b, 6c, and 6d have a linearly ordered trimanganese core, Mn(II)Mn(III)Mn(II), bridged by two carbohydrate residues with Mn-Mn separations of 3.845(2)-3.919(4) A and Mn-Mn-Mn angles of 170.7(1)-173.81(7) degrees. The terminal Mn(II) atoms are seven-coordinate with a distorted mono-face-capped octahedral geometry ligated by the (aldose)3-tren ligand through three oxygen atoms of C-2 hydroxyl groups, three N-glycosidic nitrogen atoms, and a tertiary amino group. The central Mn(III) atoms are five coordinate ligated by four oxygen atoms of carbohydrate residues in the (aldose)3 tren ligands and one water molecule, resulting in a square-pyramidal geometry. In the bridging part, a beta-aldopyranosyl unit with a chair conformation bridges the two Mn(II)Mn(III) ions with the C-2 mu-alkoxo group and with the C-1 N glycosidic amino and the C-3 alkoxo groups coordinating to each metal center. These structures could be very useful information in relation to xylose isomerases which promote aldose-ketose isomerization by using divalent dimetal centers such as Mn2+, Mg2+, and Co2+. PMID- 11272564 TI - Rhodium complexes with the chelating and binucleating ligands P(CH2CH2Py)nPh3-n (Py = 2-pyridyl; n = 1,2): structures and fluxional behavior. AB - Several rhodium(I) complexes of the type [RhX(CO)(PePy2)], [Rh(diene)(PePy)]+, and [Rh(diene)(PePy2)]+ (PePyn = P(CH2CH2Py)nPh3-n; Py = 2-pyridyl; n = 1, 2) have been prepared. The two former are square planar; the latter are pentacoordinated for diene = tetrafluorobenzobarrelene or norbornadiene (confirmed by X-ray diffraction), but an equilibrium of 4- and 5-coordinate isomers exists in solution for diene = 1,5-cyclooctadiene. The fluxional behavior of all these complexes is studied by NMR spectroscopy. The complex [Rh(NBD)(PePy2)]PF6.Cl2CH2 crystallizes in the monoclinic space group P21/n with a = 8.455(1) A, b = 18.068(3) A, c = 19.729(3) A, beta = 99.658(3)degrees, and Z = 4. The complexes [Rh(diene)(PePy2)]+ react with CO to give the dimeric complex [Rh2(CO)2[P(CH2CH2Py)2Ph]2](BF4)2 with the pyridylphosphine acting as P,N chelating and P,N-bridging. PMID- 11272565 TI - Synthesis and characterization of nickel(II) bis(alkylthio)salen complexes. AB - NiX2(2-RSC6H4CH=NCH2CH2N=CHC6H4SR-2) (NiX2L; L = 5) (1a, X = Br, R = C6H13; 1b, X = Cl, R = C12H25) and NiX2(2-C6H13SC6H4CH2NHCH2CH2NHCH2C6H4SC6H13-2) (NiX2L; L = 6) (2a, X = Br; 2b, X = Cl; 2c, X = OClO3) were prepared from ligands 5 and 6, respectively. The 1:2 metal-ligand complex Ni(OClO3)2(2 RSC6H4CH2NHCH2CH2NHCH2C6H4SR-2)2 3, was obtained from an EtOH solution of 2c. The characterization of paramagnetic 1-3 included single-crystal X-ray diffraction studies of 1a and 3. Complex 2c converted into 3 in the presence of excess ligand 6 in CHCl3. PMID- 11272566 TI - Monochlorogallane: physical properties and structure of the gaseous molecule H2Ga(mu-Cl)2GaH2 as determined by vibrational, electron diffraction, and ab initio studies. AB - Monochlorogallane, synthesized by the metathesis of gallium(III) chloride with an excess of trimethylsilane at ca. 250 K, has been characterized by chemical analysis, by its IR, Raman, and 1H NMR spectra, and by the products of its reaction with trimethylamine. The vibrational spectra of the vapor species isolated in solid Ar, N2, or CH4 matrixes at ca. 12 K imply the presence of only one species, viz. the dimer with an equilibrium structure conforming to D2h symmetry. The structure of this molecule has been determined by gas-phase electron diffraction (GED) measurements augmented by the results of ab initio molecular orbital calculations. An equilibrium structure with D2h symmetry has been assumed in the analysis of the electron diffraction pattern. However, as the molecule has a very low frequency Ga(mu-Cl)2Ga ring-puckering mode, a dynamic model was used to describe it with the aid of a set of pseudoconformers spaced at even intervals (deltadelta = 5 degrees, deltamax, = 20 delta) around the ring puckering angle delta and Boltzmann-weighted according to a quartic potential V(delta) = V4delta4 + V2delta2. The differences in bond distances and angles between the different pseudoconformers were constrained to the values derived from the ab initio calculations employing second-order Moller-Plesset (MP2) methods (with all the electrons included in the correlation calculations) and a 6 311G(d) basis set. The results for the weighted average of the principal distances (ralpha) and angles ( UO2(H2O)4(2+) + H2O and UO2(H2O)5(2+) + H2O --> UO2(H2O)6(2+) indicates that the thermodynamics favors the second (associative) reaction in gas phase at 0 K, while the thermodynamics of water transfer between the first and second coordination spheres, UO2(H2O)5(2+) --> UO2(H2O)4(H2O)2+ and UO2(H2O)5(H2O)2+ --> UO2(H2O)6(2+), favors the first (dissociative) reaction. The energy difference between the associative and dissociative reactions is small, and solvation has to be included in ab initio models in order to allow quantitative comparisons between experimental data and theory. Theoretical calculations of the activation energy were not possible because of the excessive computing time required. On the basis of theoretical and experimental studies, we suggest that the water exchange in UO2(H2O)5(2+) follows a dissociative interchange mechanism. The rates of exchange of water in UO2(oxalate)F(H2O)2- (and UO2(oxalate)F2(H2O)2- studied previously) are much slower than in the aqua ion, kex = 1.6 x 10(4) s(-1), an effect which we assign to hydrogen bonding involving coordinated water and fluoride. The kinetic parameters for the exchange of water in UO2(H2O)52+ and quenching of photo excited *UO2(H2O)5(2+) are very near the same, indicating similar mechanisms. PMID- 11272580 TI - High-pressure synthesis of the polymorph of layer structured compounds MNX (M = Zr, Hf; X = Cl, Br, I). AB - Transition metal nitride halides MNX (M = Zr, Hf; X = Cl, Br, I) have two types of layer structured polymorphs, the alpha-form with the FeOCl type and the beta form with the SmSI type. Both polymorphs consist of corrugated double M-N layers sandwiched between halogen layers, but with different atomic arrangements within the layers. The beta-form had been considered to be a high-temperature polymorph, because some beta-forms were obtained by thermal treatment of the corresponding alpha-forms. Here, the alpha-form was successfully transformed into the beta-form under high-pressure and high-temperature conditions; the new members of the beta form were prepared for the first time from alpha-HfNBr, alpha-ZrNI, and alpha HfNI using a high pressure of 3-5 GPa at 900 degrees C. The beta-form should be characterized as the high-pressure form rather than the corresponding high temperature polymorph. This is the first high-pressure study on the polymorphs of metal nitride systems. PMID- 11272581 TI - Synthesis and spectroscopy of N3P3X5OCH=CH2 (X = Cl, F, OCH3, OCH2CF3, N(CH3)2) and N3P3X4(OCH=CH2)2 (X = Cl, N(CH3)2). Correlations of ultraviolet photoelectron spectroscopy and nuclear magnetic resonance data to electronic and geometrical structure. AB - The syntheses of the vinyloxycyclotriphosphazene derivatives N3P3X5OCH=CH2 (X = OMe, OCH2CF3) and the N3P3(NMe2)4(OCH=CH2)2 isomeric mixture along with improved preparations of N3P3X5OCH=CH2 (X = F, NMe2) are reported. The interactions between the vinyloxy function and the cyclophosphazene in these and the previously reported N3P3Cl5 (OCH=CH2) and N3P3F6-n(OCH=CH2)n (n = 1-4) have been examined by ultraviolet photoelectron spectroscopy (UPS) and NMR spectroscopy. The UPS data for the chloro and fluoro derivatives show a strong electron withdrawing effect of the phosphazene on the olefin that is mediated with decreasing halogen substitution. The 1H and 13C NMR data for N3P3X5OCH=CH2 (X = F, Cl, OMe, OCH2CF3, NMe2) show significant changes as a function of the phosphazene substituent. There is a linear correlation between the beta-carbon chemical shift on the vinyloxy unit and the phosphorus chemical shift at the vinyloxyphosphorus centers. The chemical shifts of the different phosphorus centers on each ring are also related in a linear fashion. These relationships may be understood in terms of the relative electron donor-acceptor abilities of the substituents on the phosphazene ring. The 1H NMR spectra of the N3P3(NMe2)4(OCH-CH2)2 isomeric mixture allow for assignment of the relative amounts of cis and trans isomers. A model for the observed cis preference in the formation of N3P3Cl4(OCH=CH)2 is presented. PMID- 11272582 TI - Low-valent gallium, indium, and tin compounds that contain a highly fluorinated tris(pyrazolyl)borate ligand: syntheses and characterization of. AB - Syntheses and characterization of gallium(I), indium(I), and tin(II) complexes of the [HB(3,5-(CF3)2Pz)3]- ligand (where [HB(3,5-(CF3)2Pz)3]- = hydrotris(3,5 bis(trifluoromethyl)pyrazolyl)borate)) are reported. X-ray crystal structures of [HB(3,5-(CF3)2Pz)3]In and [HB(3,5-(CF3)2Pz)3]Sn(CF3SO3) show monomeric structures in the solid state. The In-N and Sn-N bond distances are longer than the corresponding bond distances of nonfluorinated analogues. NMR data of the gallium(I) adduct [HB(3,5-(CF3)2Pz)3]Ga are very similar to those of the indium(I) analogue suggesting similar solution structures. PMID- 11272583 TI - Ruthenium complexes with a terminal hydrazido ligand. Synthesis, spectroscopy, and X-ray crystal structure. AB - Bis(1,1-diphenylhydrazido(1-))ruthenium(IV) porphyrins, [Ru(IV)(Por)(NHNPh2)2] (Por = TPP, TTP, 4-Cl-TPP, 4-MeO-TPP), were prepared in approximately 60% yields through the reaction of dioxoruthenium(VI) porphyrins, [Ru(VI)(Por)O2], with 1,1 diphenylhydrazine in ethanol. This new type of ruthenium complex has been characterized by 1H NMR, IR, UV-vis, and FABMS with elemental analysis. The crystal structure of [Ru(IV)(TTP)(NHNPh2)2], which reveals an eta1-coordination mode for both hydrazido axial ligands, has been determined. The average Ru-NHNPh2 distance and Ru-N-N angle were found to be 1.911(3) A and 141.1(3) degrees, respectively. The porphyrin ring exhibits a ruffling distortion that is unprecedentedly large for ruthenium complexes with simple porphyrinato ligands (such as TTP). This is probably due to the steric effect of the axial hydrazido(1 ) ligands. PMID- 11272584 TI - Structural characterization of anti- and syn-allyltricarbonyliron bromide: rotational spectra, quadrupole coupling, and density functional calculations. AB - Rotational transitions for two distinct structural isomers of allyltricarbonyliron bromide have been clearly observed in the cold molecular beam of a pulsed-beam Fourier transform microwave spectrometer. Rotational transitions exhibiting quadrupole splitting patterns for each isomer were measured for the 79Br and 81Br isotopomers. Both isomers are accidental near prolate symmetric tops. The measured rotational constants for the 79Br isotopomer are A(anti) = 920.6148(2) MHz, B(anti) = 582.8866(12) MHz, C(anti) = 581.3027(12) MHz, A(syn) = 919.5055(1) MHz, B(syn) = 584.1865(1) MHz, and C(syn) = 581.6392(1) MHz. Analysis of the isotopic substitution data and possible transition assignments indicates that these molecules have Cs symmetry. Both isomers are found to have a dipole component along the a axis. However, the anti isomer has a "c" type dipole component, whereas a "b" dipole component is found for the syn isomer. It was found necessary to carefully analyze both rotational constants and the quadrupole coupling data in order to determine the correct assignment of dipole moment components for each isomer. This change in dipole assignments implies that there is a switch of inertial axes upon isomerization resulting from a subtle shift of the allyl center of mass coordinates, upon reorientation of the allyl ligand. The X-ray and DFT calculated structures for the anti isomer are in excellent agreement with the present data. No previous structural data for the syn isomer were available, and the present analysis strongly supports the expected conformation. PMID- 11272586 TI - Pentafluorosulfanylnitramide salts. AB - The synthesis and properties of a new class of inorganic salts, named pentafluorosulfanylnitramide salts (or pentafluorosulfanylnitraminic acid salts) [Z+SF5NNO2-], are described. A number of SF5-nitramide salts (Z+SF5NNO2-) were successfully prepared via nucleophilic displacements from carbamates and/or ion exchange techniques, but some salts [M(SF5NNO2)x; M = Li, Mg, Al] decomposed during isolation procedures and appear to be unstable in the solid state. Single crystal X-ray diffraction was used to fully characterize the Z+SF5NNO2-, and their properties/structures are compared with those of the corresponding dinitramide salts (or dinitraminic acid salts), Z+N(NO2)2-. X-ray crystallography revealed major structural differences between N(NO2)2- and SF5N(NO2)- salts concerning the N-N distances and the angles subtended at the central nitrogen atom. In the N(NO2)2- salts, there are two nonequivalent N-N (average lengths 1.372(2) and 1.354(2) A) distances and an average N-N-N angle of 115.8(3) degrees (falls between sp3 and sp2 hybridization). In the SFsNNO2- salts, the average N-N distance is much shorter, 1.308(9) A, and the average N-N-S angle is 120.0(5) degrees (closely fits sp2 hybridization). The SF5NNO2- salts show a remarkable metrical similarity for the SF5 moiety in all structures, indicating a lack of sensitivity to its steric and electronic environment. This is in marked contrast to N(NO2)2-, where there is a wide variation in conformations adopted by these anions which can be related to their environment. PMID- 11272585 TI - Cisplatin--DNA cross-link models with an unusual type of chirality-neutral chelate amine carrier ligand, N,N dimethylpiperazine (Me2ppz): Me2ppzPt(guanosine monophosphate)2 adducts that exhibit novel properties. AB - Most simple cis-PtA2G2 complexes that model the G-G cross-link DNA lesions caused by the clinically used anticancer drug cis-PtCl2(NH3)2 undergo large fluxional motions at a rapid rate (A2 = two amines or a diamine; G = guanine derivative). The carrier amine ligands in active compounds have NH groups, but the fundamental role of the NH groups has been obscured by the dynamic motion. To assess carrier ligand effects, we examine retro models, cis-PtA2G2 complexes, in which dynamic motion has been reduced by the incorporation of steric bulk into the carrier ligands. In this study we introduce a new approach employing the chirality neutral chelate (CNC) ligand, Me2ppz (N,N'-dimethylpiperazine). Because they lie in the Pt coordination plane, the methyl groups of Me2ppz do not clash with the 06 of the base of G ligands in the ground state, but such clashes sterically hinder dynamic motion. NMR spectroscopy provided conclusive evidence that Me2ppzPt(GMP)2 complexes (GMP = 5'- and 3'-GMP) exist as a slowly interconverting mixture of two dominant head-to-tail (HT) conformers and a head-to-head (HH) conformer. Since the absence of carrier ligand chirality precluded using NMR methods to determine the absolute conformation of the two HT conformers, we used our recently developed CD pH jump method to establish chirality. The most abundant HT Me2ppzPt(5'-GMP)2 form had A chirality. Previously this chirality was shown to be favored by phosphate-cis G NIH hydrogen-bonding interligand interactions; such interactions also favor the HT conformers over the HH conformer. For typical carrier ligands, G O6 and phosphate interactions with the carrier ligand NH groups also favor the HT forms. These latter interactions are absent in Me2ppzPt(GMP)2 complexes, but the HT forms are still dominant. Nevertheless, we do find the first evidence for an HH form of a simple cis-PtA2G2 model with A2 lacking any NH groups. In previous studies, the absence of the HH conformer in cis-PtA2G2 complexes lacking carrier NH groups may be due to the presence of out-of-plane carrier ligand bulk. Such bulk forces both G O6-G O6 and G O6-carrier ligand clashes, thereby disfavoring the HH form. The major DNA cross link adduct has the HH conformation. Thus, for anticancer activity, the small bulk of the NH group may be more important than the H-bonding interaction. PMID- 11272587 TI - Metal-ligand interactions in bis(isodicyclopentadienyl)iron complexes. PMID- 11272588 TI - Reactivity of the [Mo3(mu3-S)(mu2-S2)3Br6]2- anion toward the imidodiphosphinochalcogenido ligands. PMID- 11272589 TI - Reaction between the (3,1) isomer of Ru2(F5ap)4Cl and CN-. Synthesis, structural determination, and electrochemistry of Ru2(F5ap). PMID- 11272590 TI - EXAFS study of uranyl nitrate dimer at high and low temperature. PMID- 11272591 TI - Synthesis, structure, and magnetic properties of the charge-transfer salt ferromagnet decamethylchromocenium dimethyl dicyanofumarate, T(Curie) = 5.7 K. PMID- 11272592 TI - Transition or change? PMID- 11272593 TI - Assessment of unidimensionality of physical functioning in patients receiving therapy in acute, orthopedic outpatient centers. AB - Physical functioning is a common construct of interest for patients receiving rehabilitation. This report describes the assessment of hierarchial structure, unidimensionality and reproducibility of item calibrations along the continuum of physical functioning defined by the PF-10 of the MOS SF-36. Three new questions specific to patients with upper extremity impairments were added, and item calibrations were compared across several groups of patients with different musculoskeletal impairments. Reproducibility of item calibrations over testing times was supported. Item order was dependent on impairment in a clinically logical pattern. Construct validity of the physical functioning scale was supported and improved with the new questions for patients with upper extremity impairments as well as for patients with some lower level extremity impairments. PMID- 11272595 TI - Measuring the capacity of families to provide unpaid support for a disabled family member: using BIGSTEPS to identify primary and secondary dimensions. AB - The Principal Components subroutine of the BIGSTEPS computer program (Rasch analysis) was used to identify primary and secondary dimensions within an item set composed of variables pertaining to family capacity to provide assistance to a family member with disability. Data were obtained through interviews with family caregivers of patients in a major rehabilitation hospital, both during patients' inpatient rehabilitation stay and 3 and 6 months after inpatient discharge. Rasch analysis revealed the primary dimension within these data to be family capacity to provide unpaid instrumental help (18 items) and the strongest secondary dimension to be stressors on the caregivers as assessed during the inpatient stay (5-11 items). A scale consisting of the caregiver stressor items significantly predicted patients' functional gain at 3 months after discharge from inpatient rehabilitation (r = -.21) and at 6 months (r = -.20), and also the number of days that the patient had spent in a nursing facility as of 6 months after discharge (r = +.17). Caregiver stress, burnout, and quality of life at 3 months and 6 months were also significantly predicted. These findings strongly suggest the importance of more definitive research into family stressors that affect long-term patient outcomes. PMID- 11272594 TI - Identifying shortcomings in the measurement of service quality. AB - SERVPEFR, the performance component of the Service Quality Scale (SERVQUAL), has been shown to measure five underlying dimensions corresponding to Tangibles, Reliability, Responsiveness, Assurance, and Empathy (Parasuraman, Zeithaml, & Berry, 1988). This paper describes three separate studies employing SERVPERF in an Australian context. In the first of these studies (N = 113), a shortened 15 item version of the SERVPERF scale (SERVPERF-R) was found to be suitable for use in an Australian small business setting. A five-factor structure was identifiable but the factors were highly correlated, suggesting that they were not clearly distinct. The tendency for marked negative skewness observed by other researchers was also noted here. A follow-up study involving three other small businesses (N = 212) used Rasch analysis to test assumptions about the spread of items on the underlying continuum. These analyses indicated that there is an even, though narrow, spread of items across the continuum. The Rasch analysis suggested that the items in both SERVPERF and SERVPERF-R are too easy to rate highly and that more "difficult" items need to be added to the scale. The third study (N = 122) was conducted using a version of SERVPERF-R that included seven new items intended to extend the range of the scale. The new items, however, did not achieve this desirable outcome. The implications for service quality assessment are discussed. PMID- 11272596 TI - The stability of health status measurement (SF-36) in a working population. AB - This study tests the stability of health status measurement (SF-36) in a working population. A total of 4,225 employees from two sectors (one state agency, one private company) enrolled in three health plans at Trigon BlueCross/BlueShield of Virginia. An eight-dimension short-form health survey (SF-36) was first tested on a cross-sectional basis for its validity. Then, a panel study was established to test for the stability of health status instrument over time. Structural equation modeling built on equality constraint conditions was the statistical technique for this study. Data were collected through two-wave mail surveys. Both comprehensive (original eight scales) and parsimonious (revised five scales) models of health status were found fit into the data quite well. Furthermore, the revised parsimonious model was shown highly stable over time. Within a working population aged 18 to 64, people are relatively healthy. Their perception of health issues is reflected mainly on "physical health status," as indicated by physical functionings or role limitations. The high stability of revised health status model warrants the possibility of using a more concise health status instrument for the majority of people in working force. PMID- 11272597 TI - New directions in pediatric rehabilitation measurement: the growing challenge. AB - The Center on Rehabilitation Effectiveness (CRE) was created in 1998 at Boston University's Sargent College of Health and Rehabilitation Sciences. An important reason for the creation of the Center was demand by purchasers of health services and patients for high quality yet cost-effective rehabilitation programs, particularly with respect to pediatric services. This demand for accountability has created many pressures and challenges for the psychometric community. These challenges include: pediatric rehabilitation assessments that are conceptually grounded in rehabilitation theory; new instruments that are short yet sensitive enough to detect meaningful disability restrictions and that are sensitive enough to measure meaningful change; and new scales that offer real meaningful comparisons across patients. The purpose of this paper is to explain how the Center for Rehabilitation Effectiveness will meet these growing challenges. PMID- 11272599 TI - Pseudolikelihood estimation of the Rasch model. AB - An estimation method is proposed for the Rasch model on the basis of the pseudolikelihood theory of Arnold and Strauss (1988). A simulation study was conducted to compare the proposed maximum pseudolikelihood estimates with the well known conditional maximum likelihood and unconditional maximum likelihood estimates for the item parameters of the Rasch model. The results show great similarity between the methods. PMID- 11272598 TI - Naturalistic assessment of functional performance in school settings: reliability and validity of the School AMPS scales. AB - The School Assessment of Motor and Process Skills (School AMPS) is an assessment tool designed to be used by occupational therapists to measure the effectiveness of a student's ability to perform school tasks in naturalistic classroom settings. Rater reliability, internal scale validity, and person response validity of the School AMPS was investigated by examining the goodness-of-fit of raters, motor and process skill items, and students to the many-faceted Rasch model used in the development of the School AMPS. Five of six raters demonstrated acceptable goodness-of-fit (MnSq < or = 1.4 and z < 2). All 36 motor and process skill items demonstrated acceptable goodness-of-fit. Of the 208 students in the study, 93.7% demonstrated acceptable goodness-of-fit on the School AMPS motor scale and 88.9% demonstrated acceptable goodness-of-fit on the School AMPS process scale. The results of this study support the rater reliability, scale validity, and person response validity for the School AMPS as a tool to be used to evaluate the effectiveness of student performance of school tasks in the classroom setting. PMID- 11272600 TI - Perspectives. Disease management for Medicare: how many paradigms can we shift today? PMID- 11272601 TI - Quantitation of peripheral blood cytomegalovirus DNA for monitoring recurrent cytomegalovirus retinitis in pediatric solid organ transplant recipients. AB - Cytomegalovirus (CMV) infection is a major concern following solid organ transplantation, especially in the pediatric population who remain at high risk of primary infection. CMV disease leads not only to increased patient and graft morbidity, but also to increased health care costs. This study describes the usefulness of a quantitative CMV polymerase chain reaction (PCR) technique for monitoring peripheral blood CMV DNA in pediatric recipients of kidney and liver allografts who had recurrent CMV retinitis. The incidence of CMV disease in 28 pediatric transplant recipients was 28.6%, one-half of whom developed retinitis. Two of these patients had recurrent retinitis on cessation of anti-viral treatment. A peripheral blood CMV DNA copy number of > or =500/microg of DNA was associated with recrudescence of the retinitis in these patients. We conclude that the measurement of peripheral blood CMV DNA by PCR is a useful tool for the surveillance of disease resolution and recurrence. This is particularly important in patients with CMV retinitis, who may remain asymptomatic for a period of time, despite recurrences. PMID- 11272602 TI - The impact of acute rejection on chronic rejection: a report of the North American Pediatric Renal Transplant Cooperative Study. AB - The North American Pediatric Renal Transplant Cooperative Study (NAPRTCS) group has analyzed its database from January 1987 to October 1998. During this time we enrolled 6,395 transplants: of these, 5,323 were primary and 1,072 were repeat transplants. Overall, 30.8% (483/1,566) of the grafts failed as a result of chronic rejection. For living donor (LD) grafts, the failure rate as a result of chronic rejection was 32% (175/553), and it was 30% (308/1,013) for cadaveric donor (CD) transplants. A proportional hazards model identified first acute rejection, multiple rejections, and a late acute rejection as risk factors for the development of chronic rejection. Additional risk factors for the development of chronic rejection were African-American race, a repeat transplant, and a cyclosporin A (CsA) dose of < 5 mg/kg/day. Our analysis found that one acute rejection episode increases the risk of chronic rejection graft failure three fold. Patients with two or more acute rejections have a 12-fold increased relative risk (RR) of chronic rejection graft loss (CRGL). A late acute rejection (> 365 days post-transplant) increases the RR by six-fold. Two or more acute rejections, when the first is a late initial rejection, increases the RR 26-fold. Based on this information we have initiated a multicenter trial of intervention in patients with one or more acute rejections. PMID- 11272604 TI - Does treatment of bladder dysfunction prior to renal transplant improve outcome in patients with posterior urethral valves? AB - Fourteen patients with posterior urethral valves and end-stage renal failure were urodynamically evaluated in order to identify and correct any bladder dysfunction before renal transplant. Of the 14 patients, during filling, six had normal bladder function, two had an over-distended bladder (one with instability), one had instability, four had poor compliance, and one had a very reduced bladder capacity. During the voiding phase, one had a myogenic failure and another had detrusor-sphincteric dyssynergia. Four patients underwent bladder augmentation. Three were managed with anticholinergic therapy and two required clean intermittent catheterization owing to post-voiding residual urine. A renal transplant was performed in all of these patients at a mean age of 8.2 years. We compared outcome in these 14 patients with bladder dysfunction treated before transplantation with outcome in a matched control group of 14 transplant patients. Graft function and survival were similar in both groups. We believe that urodynamic studies must be included in the pretransplant evaluation of patients with posterior urethral valves in order to diagnose any bladder dysfunction and commence appropriate treatment to avoid any negative influence on graft function. PMID- 11272603 TI - Survival after acute graft failure in pediatric thoracic organ transplant recipients. AB - Survival among recipients of repeat thoracic organ transplantation, particularly in the setting of acute graft failure (AGF), is lower than survival after a primary transplant. This has created controversy over the fair allocation of scarce organs. We reviewed our experience to assess the effectiveness of aggressive therapy and retransplantation in pediatric patients with AGF. Between November 1994 and March 1998, 52 patients aged 49 days to 16.9 years (median age 4.7 years) underwent thoracic organ transplantation (32 primary and 4 repeat heart, 16 primary and 4 repeat lung, and 3 primary heart-lung transplants). Acute graft failure occurred in nine (4 heart, 3 lung, 2 heart-lung transplants), six of whom were supported with extracorporeal membrane oxygenation (ECMO), and four of whom underwent repeat transplant. Six of the nine survived, including all of those who were retransplanted, and five of the nine were alive 1 year later. The average postoperative hospital stay after receiving a second organ was 46.5 days vs. a postoperative 22-day stay in recipients without AGF (p = 0.07). We conclude that the decision to allocate institutional and professional resources to the aggressive support of patients with AGF must be made at the level of the individual transplant center. However, we feel that the outcome of aggressive support and retransplantation justifies the allocation of organs to these patients and suggests that the current policies governing organ allocation for patients with early graft failure should be re-examined. PMID- 11272605 TI - Administration of recombinant human granulocyte-macrophage colony-stimulating factor to children undergoing allogeneic marrow transplantation: a prospective, randomized, double-masked, placebo-controlled trial. AB - Granulocyte-macrophage colony-stimulating factor (GM-CSF) was administered to 40 pediatric patients undergoing partially matched related, or closely matched unrelated, allogeneic marrow transplants. This trial was set up in a prospective, randomized, double-masked, placebo-controlled manner to establish if the administration of GM-CSF to such patients enhanced neutrophil recovery in this allogeneic transplant setting. The GM-CSF group had a significantly shorter time to neutrophil recovery to > 500 x 10(9) cells/L (15 days) than the placebo group (p = 0.0036). In addition, the GM-CSF group had a significantly shorter neutrophil recovery time to > 1,000 x 10(9) cells/L (18 days) than the placebo group (p = 0.0053). The primary objective of this study was met by showing that GM-CSF enhanced neutrophil recovery in this allogeneic setting. However, within the study group of patients, there was no effect of GM-CSF on the incidence or severity of graft-vs.-host disease (GvHD), one of the secondary end-points of the study. With regard to the other secondary end-points, there was no effect of GM CSF on marrow cellularity, duration of systemic antibiotics given for real infections or as prophylaxis to prevent infections, risk of significant infections (as defined by systemic culture of virus, fungus, or bacteria), and duration or cost of hospitalization, platelet recovery, and nutritional support. With the secondary end-points, it will be necessary to study larger numbers of pediatric patients to identify differences that are small in this study group. In conclusion, GM-CSF can be safely administered to children with few, if any, significant side-effects. Additional work remains to facilitate earlier discharge of patients and decreased antibiotic usage, to offset the cost of using a neutrophil growth factor. PMID- 11272607 TI - Do six-antigen-matched cadaver donor kidneys provide better graft survival to children compared with one-haploidentical living-related donor transplants? A report of the North American Pediatric Renal Transplant Cooperative Study. AB - Since 1991, more than 50% of pediatric transplant recipients have received a living donor (LD) kidney, and approximately equals 85% of these allografts were one-haploidentical parental kidneys. Short-term (1 yr) and long-term (5 yr) graft survival of LD kidneys are 10% and 15% better, respectively, than that of cadaver donor (CD) kidneys. Because of these results, children are frequently not placed on a cadaver waiting list until the possibility of a LD is excluded--a process that may take up to 1 yr. The hypothesis for this study was that the graft outcome of a six-antigen-matched CD kidney is superior to that of a one haploidentical LD kidney, and that children are at a disadvantage by not being placed on a CD list whilst waiting for a LD. The database of the North American Pediatric Renal Transplant Cooperative Study (NAPRTCS) for 11 yrs (1987-98), was reviewed to identify children who were recipients of a six-antigen-matched CD kidney (primary and repeat transplants), and those who were recipients of a one haploidentical LD kidney (primary and repeat transplants). Using standard statistical methods, the morbidity, rejection episodes, post-transplant hospitalizations, renal function, long- and short-term graft survival, and half life of primary recipients were compared in the two groups. Unlike adult patients, only 2.7% (87/3313) of CD recipients in the pediatric age range received a six-antigen-matched kidney, and the annual accrual rate over 11 yrs was never higher than 4%. Comparison of 57 primary six-antigen-CD kidneys (PCD) with 2,472 primary LD (PLD) kidneys revealed that morbidity, rejection rates, and ratios were identical in the two groups. Renal function and subsequent hospitalizations were also identical in the two groups. Five-year graft survival of the PCD group was 90% compared with 80% for the PLD group, and the half-life of the PCD group was 25 +/- 12.9 yrs compared with 19.6 +/- 1.3 yrs. Our data suggest that the six-antigen-matched CD kidney may have less graft loss as a result of chronic rejection and would therefore confer a better long-term outcome. Based on these findings we recommend that all children, whilst waiting for a LD work-up, be listed with the United Network for Organ Sharing (UNOS) registry for a CD kidney. PMID- 11272606 TI - Follow-up of chimerism, including T- and B-lymphocytes and granulocytes in children more than one year after allogeneic bone marrow transplantation. AB - In bone-marrow-transplanted children, early detection of graft failure, relapse, and other potentially treatable problems is facilitated by the use of polymerase chain reaction (PCR) assays that monitor whether blood and marrow cells are of recipient or donor origin. Presence of mixed donor-recipient chimerism (MC) within the first year after BMT frequently correlates with clinical problems. To study if MC detected one year or more post-BMT was also often associated with clinical problems, the chimeric status in 33 children surviving 1-11 yr (median: 2 yr) after BMT was investigated. A PCR with a sensitivity of 1-2%, using fluorescent primers analyzing DNA fragment length polymorphisms, was applied. T- and B-cells and granulocytes were immunomagnetically isolated and tested separately for all patients. Of the 33 patients, of whom 21 had received pretreatment including total body irradiation (TBI), 27 (82%) exhibited full donor chimerism. Six children (18%), four of whom had received pretreatment without TBI, had MC. In three of these children, all with aplastic anemia, isolated T-cell MC had not posed apparent clinical problems. In two patients, both with MC including B-cells, immune hemolytic anemia was observed. A sixth patient with AML presented with MC and relapse. In two of the six children MC was detected only by cell subset analysis. In conclusion, analysis of MC in leukocyte subsets is more informative than analysis of whole blood only and may reveal clinically important variations in the origin of different cell populations. The prevalence of MC is lower after the first year post-BMT, and when present is less often associated with clinical problems. PMID- 11272608 TI - Risk factors for bone mineral density loss in pediatric renal transplant patients. AB - Bone mineral density (BMD) is decreased in both adult and pediatric renal transplant recipients. To investigate the risk factors associated with this decrease in BMD post-renal transplant, we studied 33 children, aged 7-22 yr, who had received a renal transplant from 0.3 to 10 yr prior to this study. BMD analysis of the total body, spine, and femur was carried out by using dual-energy X-ray absorptiometry (DEXA). Age, weight, Tanner stage, time on dialysis prior to transplantation, cumulative corticosteroid dosage, and cyclosporin A (CsA) dosage since transplantation, and use of corticosteroid therapy prior to transplantation, were recorded. Spine, femur, and total body BMD Z-scores were greater than two standard deviations (2 SD) below the mean in 45%, 42%, and 17% of patients, respectively. Age correlated inversely with total body and spine BMD Z-scores (p = 0.001 and p = 0.008); no child under 14 yr of age had a total body or spine BMD Z-score greater than 2 SD below the mean for age. Patients at a Tanner stage of 4 or 5 had lower total body and spine BMD Z-scores than did patients at Tanner stages 1-3 (p = 0.043). Time post-transplant correlated inversely with both spine and total body BMD Z-score (p = 0.013 and p = 0.023). Only total body BMD Z-score correlated inversely with cumulative corticosteroid dose (in g, p = 0.045). BMD did not correlate with cumulative CsA dose. Black patients tended to have decreased total body BMD compared with Caucasian patients. In pediatric renal transplant patients, decreases in BMD start in adolescence. Risk factors for BMD loss in these patients include increasing age, time post-transplant, increasing Tanner stage, and ethnicity. Longitudinal studies in these patients and strategies to improve BMD are needed. PMID- 11272609 TI - Epstein-Barr virus-associated post-transplant lymphoproliferative disease after bone marrow transplantation mimicking graft-versus-host disease. AB - In contrast to solid organ transplantation (Tx), the incidence of post-transplant lymphoproliferative disease (PTLD) after hematopoietic stem cell Tx (HSCT) is generally low. This risk, however, is significantly elevated in patients receiving human leukocyte antigen (HLA) mis-matched or T-cell-depleted grafts, or after treatment for severe graft-versus-host disease (GvHD). An 18-yr-old patient with positive Epstein-Barr virus (EBV) serology received a fully matched, unmanipulated bone marrow graft from an unrelated EBV-positive donor for treatment of acute myeloid leukemia (AML) in second complete remission. GvHD prophylaxis was performed with cyclosporin A (CsA) and a short course of methotrexate. Four months after Tx, the patient developed ulcerative tonsillitis that was unresponsive to antibiotic treatment. Diarrhea appearing simultaneously was interpreted as gastrointestinal GvHD and steroids were added to CsA. A few days later the patient was admitted to hospital because of generalized seizure and pneumonia. Despite reduction of immunosuppression, intensification of anti viral treatment, and subsequent mechanical ventilation, the patient died of acute respiratory distress 6 days later. Autopsy demonstrated disseminated EBV-induced, multi-nodular lymphoma infiltration of the entire colon but no signs of GvHD. Moreover, both lungs, paratracheal lymph nodes, kidneys, thyroid gland, and liver were infiltrated with large B-cell non-Hodgkin's lymphomas. This case underlines the rapid and aggressive course of EBV-induced disseminated PTLD after HSCT, initially mimicking intestinal GvHD because of massive colonic lymphoma infiltration. Tissue biopsies should be performed early for establishing correct diagnosis, thus enabling specific therapy, e.g. infusion of donor leukocytes with cytotoxic T-lymphocytes. PMID- 11272610 TI - Post-transplant epididymitis and orchitis following Listeria monocytogenes septicaemia. AB - We report the occurrence of epididymitis and orchitis 1 week after the onset of Listeriosis in an 11-month-old boy receiving an orthotopic liver transplantation for biliary atresia. Immunologic implications of Listeria monocytogenes-induced testicular inflammation are discussed, and the potential role of immunosuppression with tacrolimus is also discussed. PMID- 11272611 TI - Asymptomatic inferior vena cava abnormalities in three children with end-stage renal disease. PMID- 11272613 TI - The role of granulocyte-macrophage colony-stimulating factor in paediatric bone marrow transplantation. PMID- 11272612 TI - Chronic rejection in pediatric renal transplantation: where are we? PMID- 11272614 TI - What happens if the heart won't start? PMID- 11272615 TI - Impact of acute rejection on development of chronic rejection in pediatric renal transplant recipients. AB - For pediatric kidney transplant recipients, chronic rejection has become the predominant cause of graft loss. This article reviews risk factors for chronic rejection and what can be done to lower the risk of chronic rejection for future transplant recipients. PMID- 11272616 TI - Objectivity in psychosocial measurement: what, why, how. AB - This article raises and tries to answer questions concerning what objectivity in psychosocial measurement is, why it is important, and how it can be achieved. Following in the tradition of the Socratic art of maiuetics, objectivity is characterized by the separation of meaning from the geometric, metaphoric, or numeric figure carrying it, allowing an ideal and abstract entity to take on a life of its own. Examples of objective entities start from anything teachable and learnable, but for the purposes of measurement, the meter, gram, volt, and liter are paradigmatic because of their generalizability across observers, instruments, laboratories, samples, applications, etc. Objectivity is important because it is only through it that distinct conceptual entities are meaningfully distinguished. Seen from another angle, objectivity is important because it defines the conditions of the possibility of shared meaning and community. Full objectivity in psychosocial measurement can be achieved only by attending to both its methodological and its social aspects. The methodological aspect has recently achieved some notice in psychosocial measurement, especially in the form of Rasch's probabilistic conjoint models. Objectivity's social aspect has only recently been noticed by historians of science, and has not yet been systematically incorporated in any psychosocial science. An approach to achieving full objectivity in psychosocial measurement is adapted from the ASTM Standard Practice for Conducting an Interlaboratory Study to Determine the Precision of a Test Method (ASTM Committee E-11 on Statistical Methods, 1992). PMID- 11272617 TI - Rasch analysis in measurement of physician work. AB - This report describes the use of Rasch analysis of paired comparisons in measuring physician work. The method examines the ranking of a series of related medical or surgical procedures by survey of physicians experienced in the provision of such services. Each service in the group is paired with every other service. The physicians select which of a pair of services from the group under study represents the greater amount of physician work. When the results are analyzed by Rasch method, the resulting measures (in logits) provide a scale (yardstick) upon which each service can be placed relative to all the other services in the group. In this way, a rank ordering of the services is accomplished that is both linear and objective. The Rasch measures can than be charted against existing work values to refine the assigned relative values for such services. The result of applying this method to a series of spinal operations indicates that this technique could be expanded to many other groups of related medical services, with improved relative valuation of physician work. PMID- 11272618 TI - Uses of Rasch scaling in the measurement of cognitive development and growth. AB - This report describes two types of findings: (a) the consistency between two major cognitive tests in terms of their developmental scales based on item response theory, and (b) the initial development of ideas and methods for the revival of the classic concept of ratio IQ. The ratio IQ (e.g., Stanford-Binet, 1937) was formed by the division of mental age (derived from test performance) by chronological age multiplied by 100. Following a multitude of criticisms about the scaling qualities of the ratio IQ, it was mostly abandoned by the major intelligence batteries, beginning with the Wechsler scales in the 1940's, in favor of standard scores. This study presents a new approach to age equivalence scores as a basis for mental age, and the calculation of ratio IQ, based on Rasch model item response theory. The new ratio IQ was compared statistically with standard-score IQ (mean 100, SD 15) from the Leiter International Performance Scale--Revised (Leiter-R) and from the Woodcock-Johnson Psychoeducational Battery -Revised (WJ-R). The essential element of the new ratio IQ is the W-scale, a Rasch-based score employed in the WJ-R and in the Leiter-R. Mental age was estimated from the W-scale estimate of ability and chronological age from a W scale age equivalence for each month of age. Statistical results showed a highly similar growth curve for the W-scale scores on the Leiter-R and the WJ-R, even though the two scales have different content and standardization samples. Also, high correlations were found between the new ratio IQ and standard-score IQ (e.g., correlations ranging from .87 to .95 depending on age range). Criterion related evidence of validity was found in the correlation of .82 between the new ratio IQ and the Wechsler Intelligence Scale for Children--Third Edition (WISC III) standard-score IQ and in correlations with achievement-test scores. Finally, the ratio IQ showed predictable mean differences between groups of children with typical cognitive ability, cognitive delay levels of performance and giftedness. The standard deviation of the new ratio IQ was somewhat variable across age groups, however, so new interpretive guidelines would be needed if the new index is to employed in published tests. Implications of the scaling methods are discussed. PMID- 11272619 TI - Biologic patterns of disability. AB - We describe the use of a mathematical/statistical method (i.e., Rasch analysis) to elucidate biological patterns of disability present in the functional ability of persons undergoing medical rehabilitation. Two measures chosen for illustration are the FIM Instrument for inpatients and the Body Movement and Control (BMC) measure for outpatients. In order to meet the assumptions necessary for application of linear statistics to clinical measurement studies, Rasch analysis was used to transform ordinal scales into linear measures. Another unique feature of Rasch analysis is that it allows evaluation of the difficulty of items and the abilities of persons being tested, separately, on the same metric. Also, the difficulty represented by each item may be arranged along a hierarchy from easy to hard. The hierarchies of functional ability items are dependent upon the specific patterns of disability related to underlying pathophysiology. For inpatients, initial analyses of the 18 items of the FIM Instrument demonstrated separate hierarchies for the 13 motor items and for the 5 cognition items. Subsequent analyses demonstrated five distinct patterns for the 13 motor items of: brain dysfunction, orthopedic conditions, pain conditions, ambulatory spinal cord dysfunction, and wheelchair users with spinal cord dysfunction. Two patterns were identified for cognition: stroke with right body hemiparesis and all others. For outpatients, the BMC measure of physical functioning is used to demonstrate that pathophysiologic conditions are expected to affect the hierarchial pattern of items differently. This was noted to be the case for persons with lower body dysfunction, low back pain, and neck pain/upper limb dysfunction. Based upon the item responses, sitting, reaching and standing appear to represent items most useful for discriminating between the three conditions in terms of the functional consequences. Rasch analysis, among other advantages, enables investigation of the subtle relationships among items and is a useful method to evaluate underlying biological patterns of disability. A clinician, using a map that shows the expected relationships between item scores, may observe that a particular patient matches or does not match the expected pattern. Such insights may help the clinician in monitoring the responses of the patient to treatment efforts. PMID- 11272620 TI - FIM levels as ordinal categories. AB - Data collected on rating scales have generally been analyzed without verifying that the scales have functioned as intended. The FIM levels are precisely conceptualized and meticulously defined. Their effective empirical functioning as ordinal categories merits continual monitoring. Ordinality implies that each succeeding level represents a higher level of functioning. Further, as a patient improves in functioning each ordinal level in turn is expected to be observed. Taking advantage of the clarity of Rasch theory, guidelines are suggested that prompt the analyst to investigate whether the rating categories are cooperating to produce observations on which useful measurement and prudent inference about patient status can be based. PMID- 11272621 TI - Food: sustenance and symbol. PMID- 11272623 TI - Dietary intake, body mass index, exercise, and alcohol: are college women following the dietary guidelines for Americans? AB - Study findings suggest that college women practice diet and health behaviors that contradict the 1995 Dietary Guidelines for Americans. To confirm this hypothesis, the authors surveyed the diet, exercise, and health habits of 60 female students enrolled in three university aerobics courses. They measured height and weight to calculate body mass index (BMI) and assessed physical activity, using the Self Reported Physical Activity scale. To estimate food and nutrient intake, they used 3-day food records. Participants reported diets that were nutritionally adequate but exceeded national recommendations for fat, sugar, and sodium, and their reports of exercise habits suggested that the lifestyles of 66% of the respondents were sedentary. Although the students' mean BMIs suggested healthy weights, 25% of the women were classified as overweight. A majority of the participants were following at least 1 of the 7 dietary guidelines; however, no participant was adhering to all proposed behaviors. PMID- 11272622 TI - The eating disorders NOS diagnostic profile among college women. AB - The authors examined a proposed profile of eating-related behaviors, associated features, developmental issues, and help-seeking behavior among college women, using an eating disorder response program. The most common symptom scenario was a pattern of regular binge eating, together with daily exercise and occasional purging. The most common associated features were distressing or dysfunctional overconcern about body image and self-esteem, usually with day-to-day stress and intermittent depression. The women who fit this pattern also presented developmental issues of perfectionism, conflictual relationships with parents, and struggles for independence; and they tended to be ambivalent about seeking services. Implications for practice, including the need to develop a framework for eating disorder responses on campus that includes preventive programs and developmental interventions to target emerging and moderate concerns are discussed; limitations and the preliminary nature of the findings are explicated. PMID- 11272624 TI - Dietetics majors' weight-reduction beliefs, behaviors, and information sources. AB - One hundred twenty-eight female dietetics majors aspiring to be registered dietitians were surveyed to identify and assess their reasons for wanting to lose weight and the weight-loss techniques and information sources they used and would recommend to clients. Fisher's exact tests were used to analyze behavioral data, and binomial tests to determine whether proportions of students achieving their desired weight-loss outcomes were significantly greater than 50%. Most dieters wanted to lose weight to improve their appearance and increase their self-esteem. Sound weight-loss techniques that were used and recommended include increased exercise, low-fat foods and snacks, and portion control. Accurate information sources used and recommended included food labels and college nutrition courses. Unsound weight-loss techniques and potentially inaccurate information sources were also used and would be recommended by a few students. Findings suggest a need for more learning opportunities focusing on enhancement of self-esteem and weight management. PMID- 11272625 TI - Primary care for young African American men. AB - Young African American men in the inner city have higher rates of mortality and morbidity from potentially preventable causes than other American men of the same age. They suffer disproportionately high rates of preventable illness from violence, sexually transmitted diseases, and HIV infection. These young men present with problems related to sexual concerns, mental health issues, substance abuse, and violence. They also report substantial risk-taking behaviors, including unprotected sex, substance use, and weapon carrying, as well as exposure to violence. Access to and use of preventive primary care services has been limited for these patients in the past because of financial barriers and competing social issues. Racism and historical oppression have created barriers of mistrust for young men of color. Factors that contribute to their adverse health status, as well as ways to address these problems, are discussed. PMID- 11272626 TI - Planning and execution of a successful hepatitis B immunization program. AB - Today's college students occupy a "window" between older adults, whose need for hepatitis B vaccination is minimal, and younger people who receive the vaccine as part of their childhood immunization series. However, because of the high-risk activities that are often part of student behavior, college students are among the individuals who are at the highest risk for this disease; 75% of all reported hepatitis B cases occur in persons between the ages of 15 and 39 years. It is therefore imperative that college health professionals take seriously their responsibility to educate students about issues related to hepatitis B and make available programs that enable the students to receive immunizations. This article briefly describes a successful hepatitis B education and immunization program at an institution that has no mandatory prematriculation immunization requirements. PMID- 11272627 TI - Health professional students' occupational exposures to blood-borne pathogens: primary and secondary prevention strategies. AB - Health science students, along with the health professionals they hope to become, are at increased risk for certain occupational injuries and illnesses. One of these risks is occupational exposure to blood-borne pathogens, such as human immunodeficiency virus (HIV) and hepatitis, which may result in severe illnesses or even death. Two case studies demonstrate postexposure care of exposed individuals at the University of Texas Medical Branch Student Health Services before and after policy changes and prevention strategies were strengthened in response to exposure incidents. PMID- 11272628 TI - Pulmonary hypertension in COPD: old and new concepts. AB - Pulmonary hypertension is a common complication in chronic obstructive pulmonary disease (COPD). Its presence and severity is closely related to disease prognosis. Remodelling of pulmonary vessels is the principal causative factor of pulmonary hypertension in COPD. In advanced COPD, pulmonary vascular remodelling is related to the severity of arterial hypoxaemia. However, recent studies have shown that structural abnormalities and alterations of vascular function are also apparent in patients with mild COPD who do not have hypoxaemia and even in smokers with normal lung function. Pulmonary endothelium plays a crucial role in the regulation of vascular tone and cell growth of the vessel wall. Alterations of endothelial function in pulmonary arteries are apparent at the early stages of chronic obstructive pulmonary disease evolution. Potential mechanisms of endothelial damage at these initial stages include the effects of cigarette smoke components and inflammatory changes. The resultant alteration of pulmonary endothelium by these factors might predispose patients with mild chronic obstructive pulmonary disease to further vascular damage by additional factors, such as hypoxaemia, ultimately leading to pulmonary hypertension. PMID- 11272629 TI - Current classification of idiopathic interstitial pneumonias. PMID- 11272630 TI - The clinical role of bronchoalveolar lavage in the setting of idiopathic interstitial pneumonias. PMID- 11272631 TI - Difficult, therapy-resistant asthma: definition and clinical features. AB - Difficult therapy-resistant asthma may be defined as poorly controlled asthma in terms of chronic symptoms, episodic exacerbations, persistent and variable airways obstruction despite a continued requirement for short-acting beta 2 agonists despite the use of high doses of inhaled steroids. It is necessary to confirm the diagnosis of asthma, to exclude alternative diagnosis, and to make sure that adequate asthma treatment is given and being adhered to. Difficult therapy-resistant asthma is likely to consist of several clinical subgroups characterized by the temporal sequence of exacerbations and symptoms, the chronicity and rapidity of symptoms and response to treatment, such as brittle asthma, chronic difficult asthma, and fatal asthma. Exacerbating factors such as gastrooesophageal reflux, sinorhinitis, allergen exposure, drugs, respiratory tract infections and psychosocial factors must be addressed. PMID- 11272632 TI - Ear and nose involvement in systemic diseases. AB - A whole range of otolaryngeal manifestations may occur as complications or represent the first symptom and sign of a variety of systemic diseases. Otolaryngologists are often the first physicians to recognize that otolaryngeal abnormalities are symptomatic of a broader disease and mandate a systemic approach to the problem. In the present study, the authors focus primarily on ear, nose and throat manifestations that may occur in the context of systemic diseases, discussing clinical manifestations and reviewing the salient histologic, laboratory, and serologic features. PMID- 11272633 TI - The link between allergic asthma and rhinitis. PMID- 11272634 TI - Rhinological aspects of cystic fibrosis. AB - Cystic fibrosis is a genetic disorder of the exocrine glandular function leading to the formation of thick mucus in the nasal and paranasal cavities. Nearly all the patients develop sinus disease but not all have complaints. Two different sinus diseases can be found: chronic sinusitis and nasal polyposis. Only the symptomatic cases have to be treated. Medical treatment consists of antibiotherapy for chronic sinusitis and nasal steroid sprays for nasal polyps. Sinus surgery is frequently needed in case there is a persistence of symptoms (and before lung transplantation). PMID- 11272635 TI - The common cold as a trigger of asthma. AB - The common cold is a viral disease with predominant symptoms from the upper airways. Rhinovirus is the most important common cold virus, and rhinovirus infection is predominantly transmitted by direct contact (nasal secretion-hand (object)-hand-mucous membrane in eye and nose). The viral disease results in the release of IL-8 from nasal epithelial cells, causing a neutrophil-dominated inflammation in the nose. The biochemical mediators, causing nasal symptoms, have not yet been identified. A common cold is the most important cause of exacerbations of asthma in children and also in adults. The rhinovirus infection induces airway inflammation, bronchial hyperresponsiveness and asthma symptoms. However, the mode of action of the virus-induced inflammation on the asthma disease is poorly understood. As a routine, physicians give oral corticosteroid and increase the dosage of inhaled corticosteroid during a common cold-induced exacerbation of asthma, but there do not seem to be any placebo-controlled trials in support of this practice. PMID- 11272636 TI - Lung sounds in asthma and chronic obstructive pulmonary disease. PMID- 11272638 TI - Clinical assessment in the diagnosis of pulmonary embolism. PMID- 11272637 TI - Diagnosis of pneumonia in primary care. PMID- 11272639 TI - The smoking cessation clinic. AB - Tobacco smoking is a disease and the leading cause of preventable death in industrialized countries. The control of smoking is a priority for all health professionals. The best measure for counteracting this disease is to promote smoking cessation among current smokers. In fact smoking cessation results not only in an immediate reduction in mortality and morbidity, but also a decrease in the disease prevalence and consequently in the probability that youngsters are "infected". A smoking cessation clinic can be an easy and effective way to treat tobacco use and dependence. It gives intensive treatments to smokers motivated to quit, ensuring a higher success rate, but also treats "difficult" patients. Any district authority can start a clinic because it is a "low resource-low budget" structure: its staff is composed of three part-time professionals (a physician, a nurse, a psychologist); it operates with a few, cheap technical instruments and uses only evidence based treatments. The difficulties concern some existing historical and cultural gaps: so far, health staff have not included this activity in their routine, are poorly trained about smoking cessation and have smoking habits of their own. Moreover, in many countries smoking cessation interventions are not recognized as services covered by the National Health Service or private insurance. Thus it is necessary to educate and update staff on smoking cessation. To obtain smoke free hospitals and premises, special smoking cessation programmes are offered to health staff and the smoking cessation clinic is the setting where this activity preferentially takes place. To satisfy the requirements for a smoking cessation clinic, scientific societies can play an important role as providers of continuing professional education and as solicitors for the government to encourage allocation of resources to these activities. PMID- 11272640 TI - Smoking cessation clinic: an Italian experience. PMID- 11272641 TI - What's new in the COPD management? PMID- 11272642 TI - Noninvasive positive pressure ventilation in COPD patients with chronic respiratory insufficiency. PMID- 11272643 TI - A novel EF-hand calcium-binding protein in the flagellum of the protozoan Tritrichomonas suis. AB - The cloning and characterization of Ts-p41, an EF-hand calcium-binding protein of the protozoan parasite Tritrichomonas suis is described. A T. suis cDNA library was screened with monospecific antibodies affinity purified on an immunoreactive 41 kDa antigen in a Triton X-114 membrane-protein fraction. The resulting cDNA fragments turned out to be derived from 2 different genes encoding closely related Ts-p41 variants. The deduced amino acid sequences contained 6 EF-hand domains perfectly matching the canonical consensus motif and a putative C terminal prenylation site. Northern and Southern hybridizations revealed that Ts p41 was highly expressed and encoded by a gene-family. A cDNA encoding Ts-p41 was expressed as recombinant protein in Escherichia coli. By overlay with 45Ca it was demonstrated that the native and recombinant Ts-p41 proteins bind Ca2+. In immunofluorescence, epitopes recognized by anti-Ts-p41 antibodies were distributed as well on the anterior flagella as on the recurrent flagellum of the parasite. Our findings with the parabasalid T. suis suggest that multiple EF-hand bearing calcium-binding proteins might be a common phenomenon associated with flagellar motility. PMID- 11272644 TI - Eimeria telekii n.sp. (Apicomplexa: Coccidia) from Lemniscomys striatus (Rodentia: Muridae): morphology, pathology and phylogeny. AB - Using a combination of morphological, life-cycle and molecular data, we describe a new apicomplexan parasite Eimeria telekii n.sp. from a striped grass mouse Lemniscomys striatus captured in Kenya. Oocysts are oval to spherical or ellipsoidal, 20.4 x 15.7 (15.5-25.0 x 12.0-20.0) microm with a colourless, smooth and bilayered wall. Sporocysts are ellipsoidal, 11.2 x 7.8 (10.0-12.0 x 7.0-9.0) microm with a small Stieda body and granular sporocyst residuum and contain 2 elongated, banana-shaped sporozoites with a single refractile body. Life-cycle, pathogenicity and host specificity of this parasite were studied in laboratory bred Lemniscomys barbarus and BALB/c mice. Two asexual stages and the sexual phase took place within the enterocytes of the caecum and colon of L. barbarus but not in inoculated BALB/c mice. An infectious dose of 5000 oocysts caused severe clinical illness and mortality in 2/2 (100%) L. barbarus. Phylogenetic analysis of the small subunit rRNA gene of E. telekii and members of the genera Eimeria, Cyclospora and Isospora placed E. telekii within the eimerian rodent clade. PMID- 11272645 TI - Modifications of the cuticular hydrocarbon profile of Apis mellifera worker bees in the presence of the ectoparasitic mite Varroa jacobsoni in brood cells. AB - Varroa jacobsoni is an ectoparasite of Apis mellifera which invades brood cells, on 8-day-old larvae several hours before cell capping. Reproduction of the parasite takes place in the capped brood cells during the nymphose of the bee. Cuticular hydrocarbons of unparasitized bees and of bees parasitized by Varroa jacobsoni were extracted and analysed by gas chromatography (GC) coupled with mass spectrometry (GC-MS). Three developmental stages of worker honey bees were studied: larvae, pupae and emergent adults. The comparison between unparasitized and parasitized hosts was performed with Principal Components Analysis coupled with a multivariate variance analysis. The cuticular hydrocarbon profiles of honey bees were qualitatively similar, for the 3 developmental stages and regardless of the presence of Varroa in the cells. Nevertheless, comparison of the relative proportions of hydrocarbons showed that the cuticular profiles of pupae and emergent adults parasitized by 1 mite and of larvae parasitized by 2 mites were significantly different from the corresponding unparasitized individuals. Such modifications could be regarded (i) as a cause of the multi infestation in larvae during invasion of brood and (ii) as a consequence of stress and/or removal of proteins contained in the haemolymph of the host during its development. PMID- 11272646 TI - Evaluation of density-dependent fecundity in human Schistosoma mansoni infections by relating egg counts to circulating antigens through Deming regression. AB - Regression analysis of the relationship of serum circulating anodic and cathodic antigens (CAA and CCA), as a possible direct measure of worm burden, and fecal egg counts allows the study of phenomena like density-dependent fecundity in human Schistosoma mansoni infections. For a reliable analysis, variations in egg count measurements as well as in circulating antigen levels have to be taken into account, and an accurate estimation of these variations (represented by parameter lambda in the so-called Deming regression) is of great importance. From a new, extensive data set of repeated measurements of fecal egg counts and CAA and CCA concentrations we determined the respective values for parameter lambda, and (re)analysed the relationship between circulating antigens and egg counts in 3 data sets from Burundi, Senegal and Zaire by Deming regression. For comparison, ordinary linear regression was performed as well, which considerably biased the regression lines for CCA, but not for CAA. The analyses resulted in a clearly non proportional relationship between egg counts and CAA, and, to a lesser extent, CCA. Assuming that egg counts and antigen measurements directly reflect egg production and worm burdens, respectively, our findings reinforce the indication of density-dependent fecundity in schistosomiasis mansoni, as suggested by others. PMID- 11272647 TI - Fasciola hepatica miracidia are dependent on respiration and endogenous glycogen degradation for their energy generation. AB - It is generally accepted that free-living stages of parasitic helminths are dependent on aerobic degradation of endogenous energy sources for their energy generation. This concept, however, is not the result of extensive experimental evidence, but originated mainly intuitively as oxygen is widely available in their habitat and these stages generally have a small size. Schistosoma mansoni, the sole parasitic helminth whose energy metabolism has been studied throughout its life-cycle indeed has aerobically functioning free-living stages. However, large differences exist in energy metabolism between adult stages of distinct parasitic helminths, and caution should be taken in predicting that all free living stages of all parasitic helminths have the same, aerobic energy metabolism. Hence, this report studied the energy metabolism of Fasciola hepatica miracidia and demonstrated that F. hepatica miracidia are also dependent on aerobic degradation of their endogenous glycogen stores by glycolysis and on Krebs cycle activity for energy generation. However, in contrast to S. mansoni, F. hepatica miracidia cannot function anaerobically, as inhibition of the respiratory chain blocked motility and carbohydrate degradation, and finally resulted in death of the miracidia. Therefore, this report demonstrated that differences exist between miracidia of distinct species, in pre-adaptation of their energy metabolism to the occasional hypoxic conditions within their next host. PMID- 11272648 TI - Development of a PCR-based method for the detection of Opisthorchis viverrini in experimentally infected hamsters. AB - Opisthorchis viverrini infection is an endemic disease that causes a serious public health problem in southeast Asia, especially in northeast Thailand. We have developed a PCR method using a pair of primers named OV-6F/OV-6R for detecting O. viverrini eggs in stool samples and compared it with Stoll's egg count method. The primers were designed based on the pOV-A6 specific DNA probe sequence which gave a 330 base pair product. The PCR method can detect a single egg in artificially inoculated faeces or as little as 2 x 10(-17) ng of O. viverrini genomic DNA. The method gave 100% sensitivity in all hamster groups except in animals exposed to the lowest intensity of infection (1 metacercaria/hamster). In the first month of infection, the PCR method was more sensitive than using the egg-count method in all infected groups especially in the light infections. The PCR method was also successfully used in monitoring a therapeutic study. Since the PCR method showed no cross-reaction with Heterophyid flukes, it can be useful for specific identification of O. viverrini eggs in stool samples without the risk of false positives. It also has great potential for application in clinical epidemiological studies. PMID- 11272649 TI - A self-fertile species of Steinernema from Indonesia: further evidence of convergent evolution amongst entomopathogenic nematodes? AB - More than 20 species of the entomopathogenic nematode Steinernema have been described; to date, all reproduce exclusively by cross-fertilization of male and female individuals. Steinernema sp. strain T87 from Indonesia was found to consist largely of self-fertile hermaphrodites. Progeny were produced by morphological females both in insects (Galleria mellonella) and in hanging drops of insect haemolymph inoculated with a single infective juvenile. Sperm were present in the oviduct of unmated morphological females. Approximately 1% of infective juveniles developed into males, and males were also present in the second generation where they constituted 1-6% of the population. Under the same conditions the related species Steinernema longicaudum strain CB2B displayed typical steinernematid reproduction: cross-fertilization and a 1:1 sex ratio. It is argued that the development of hermaphroditism in Steinernema sp. T87 represents convergent evolution with Heterorhabditis, the other major genus of entomopathogenic nematode. PMID- 11272650 TI - Differential impact of a shared nematode parasite on two gamebird hosts: implications for apparent competition. AB - If the deleterious effects of non-specific parasites are greater on vulnerable host species than on reservoir host species then exclusion of the vulnerable host through apparent competition is more likely. Evidence suggests that such a mechanism occurs in interactions between the ring-necked pheasant (Phasianus colchicus), the grey partridge (Perdix perdix), and their shared caecal nematode Heterakis gallinarum. Modelling of the system predicts that the reduced parasite impact on the pheasant compared to the partridge results in the force of infection transmitted from pheasants to partridges being sufficient to cause partridge exclusion. Since the parasite impacts are currently estimated from correlational work, controlled infections were conducted to experimentally compare the impact of H. gallinarum on the two hosts and verify cause and effect. While challenged partridges showed reduced mass gain, decreased food consumption, and impaired caecal activity, in comparison to controls, the only detectable effect of parasite challenge on the pheasant was impaired caecal activity. The impact of H. gallinarum on challenged partridges conforms with previous correlational data, supporting the prediction that parasite-mediated apparent competition with the ring-necked pheasant may result in grey partridge exclusion. However, the observed decrease in the caecal activity of challenged pheasants could imply that H. gallinarum may also have an impact on the fecundity and survival of pheasants in the wild, particularly if food is limiting. If this is the case, the associated decrease in the force of infection to which the partridge is exposed may be sufficient to change the model prediction from partridge exclusion to pheasant and partridge coexistence. PMID- 11272651 TI - Mutation scanning analysis of microsatellite variability in the second internal transcribed spacer (precursor ribosomal RNA) for three species of Metastrongylus (Strongylida: Metastrongyloidea). AB - This study investigated sequence variability in the second internal transcribed spacer of ribosomal DNA for 3 species of Metastrongylus (porcine lungworms). The ITS-2 region was amplified by PCR from individuals of M. elongatus, M. pudendotectus and M. salmi, and then subjected directly to single-strand conformation polymorphism analysis (SSCP), which allowed the direct display of sequence variation within and among individuals representing each species. There were marked differences in SSCP profiles among species, making this approach useful for species identification. For individual species, representative bands were excised from electrophoretic gels, reamplified by PCR and subjected to direct sequencing. For all 3 taxa, variability in the ITS-2 was related chiefly to the presence of microsatellites. Eight different microsatellites were identified, namely (A)n, (TG)n, (TCG)n, (TA)n, (TATG)n, (G)n, (TACA)n and (T)n. Considerable variability in microsatellite repeat number (ranging from 1 to 23) was found among individual nematodes of a species and between species. The microsatellites were located to specific stem or loop regions in the predicted ITS-2 precursor rRNA secondary structure. The results may suggest that slipped strand mispairing in microsatellite regions contributes to sequence variability in the ITS-2 of Metastrongylus species under structural constraint as a consequence of microsatellite location in the precursor rRNA. Similar studies of the ITS-2 for a wide range of parasitic nematodes may lead to a better understanding of concerted evolution in these organisms. PMID- 11272652 TI - Characterization of 5-HT receptors in the parasitic nematode, Ascaris suum. AB - The pharmacological profiles of the 5-hydroxytryptamine (5-HT) receptors on Ascaris suum pharyngeal and somatic body wall muscles were investigated. The mechanisms involved following activation of these receptors were also studied. 5 HT activated and maintained pumping in isolated pharynxes with an EC-50 value of 44+/-1.7 microM. The 5-HT agonists, tryptamine, sumatriptan 8-OH-DPAT and 5 carboxyamidotryptamine all failed to stimulate pumping. The 5-HT2 antagonist, ketanserin, initially excited and then inhibited pumping while the 5-HT3 antagonist, ondansetron, had no effect. 5-HT and 5-HT agonists, 8-OH-DPAT, 5 carboxyamidotryptamine, alpha-methyl-5-HT and tryptamine all inhibited ACh induced contractions of a somatic body wall muscle strip. Ketanserin partially blocked the inhibitory effect of alpha-methyl-5-HT and ACh-induced contractions while the 5-HT uptake blocker, fluoxetine, potentiated the effect of 5-HT on ACh induced contractions. Basal levels of cAMP, 1540+/-232 pmol/mg, in pharyngeal muscle and 1721+/-134 pmol/mg, somatic body wall muscle, were both increased by forskolin. 5-HT had no effect on pharyngeal muscle cAMP levels but raised cAMP levels in somatic body wall muscle, e.g. 100 micron 5-HT, raised the level to 2851+/-212 pmol/mg and 1000 microM raised levels to 4578+/-1234 pmol/mg. 5-HT, 1000 microM, increased inositol phosphate levels in pharyngeal muscle. These results provide some evidence for a 5-HT2-like receptor on pharyngeal muscle. In contrast, the situation on somatic body wall muscle is more confusing since the pharmacological profile partly indicates a 5-HT2-like receptor but this receptor is linked to a rise in cAMP levels. Further studies are required to resolve the position but they must be based on the rational design of ligands specifically for nematode 5-HT receptors and not simply using ligands developed for the classification of mammalian 5-HT receptors. Such a design must take into account data from molecular biology studies of nematode 5-HT receptors. PMID- 11272653 TI - Characterization of a putative nitric oxide synthase in the neuromuscular system of the parasitic nematode, Ascaris suum. AB - In this paper we report on the biochemical presence of nitric oxide synthase (NOS)-like activity in Ascaris suum tissue and examine the pharmacological effect of NO donors on A. suum muscle strip preparation. NOS activity was determined by monitoring the formation of [3H]citrulline from [3H]L-arginine and NO formation via the oxyhaemoglobin assay. Neuromuscular tissue from A. suum which stained positively for NADPH diaphorase, contained NOS activity. Neither NOS activity nor NADPH diaphorase staining was detected in intestinal tissue. The absence of Ca2+, NADPH and other co-factors normally required for mammalian neuronal NOS activity only partially reduced the formation of both citrulline and NO by A. suum neuromuscular homogenate. The results of the biochemical assays indicate the presence of an enzyme capable of producing NO and citrulline, but with a different profile from that of rat neuronal NOS. We also present preliminary evidence for the action of NO (NO donors) in the neuromuscular system of A. suum. PMID- 11272654 TI - Methods for estimation of associations between multiple species parasite infections. AB - Human populations are often infected with more than one species of parasite, especially in developing countries where overall rates of parasitism are high. Infections with multiple parasite species may not necessarily be independent within an individual as physiological, immunological or ecological factors may result in positive or negative associations between infections with different parasite species. A general framework for estimation of these associations is presented. Data from over 215000 individuals are analysed and the associations between geohelminth (Ascaris lumbricoides, Trichuris trichiura and hookworm) and malaria species are investigated. A method is presented for analysing data from multiple communities and testing whether the associations in different communities are equal. Overall estimates of the associations between species are obtained for each country and continent where data were available. Associations between geohelminth species were, in general, found to be positive whilst both positive and negative associations were found between the different Plasmodium species. There was evidence for significant geographical heterogeneity between the associations. A method for using these parameter estimates to predict the distribution of multiple infections when only marginal prevalence data are available is described and demonstrated. PMID- 11272655 TI - What is reality? PMID- 11272656 TI - A shining city set upon a hill. PMID- 11272657 TI - Drugs and devices "10 years after". PMID- 11272658 TI - Use of low-molecular-weight heparin and glycoprotein IIb/IIIa inhibitors in acute coronary syndromes. PMID- 11272659 TI - Primary angioplasty for acute myocardial infarction. PMID- 11272660 TI - Intracoronary brachytherapy: an evolving modality for treatment of restenosis following percutaneous coronary intervention. PMID- 11272661 TI - Pharmacotherapy of systolic heart failure: emphasis on mortality outcomes. AB - Mortality outcomes in clinical trials have become the foundation for the evidence based management of patients with systolic heart failure. This review was intended to give the clinician a better understanding of newer pharmacological strategies in this patient population. PMID- 11272662 TI - Current approaches to evaluation and management of patients with ventricular arrhythmias. AB - The clinical manifestations of ventricular arrhythmias encompass a broad spectrum, from complete absence of symptoms to sudden death. Although our understanding of the pathophysiology and natural history of these arrhythmias has advanced significantly over the past decade, large gaps in our knowledge remain, especially in patients with heart failure not due to coronary artery disease. We have learned much about the appropriate roles of antiarrhythmic drugs and implantable defibrillators in the prevention of sudden death. Studies performed over the past decade have made clear that the primary treatment for patients at high risk for life-threatening ventricular arrhythmias should be the implantable defibrillator. However, specific syndromes causing ventricular tachyarrhythmias are being recognized, and care must be individualized. Although hospital mortality from acute myocardial infarction has decreased as a result of newer therapies, sudden death after hospital discharge remains an important problem, causing at least 30% of post-infarction deaths, even in patients who have received thrombolytic therapy. Two independent studies have confirmed that patients with asymptomatic non-sustained ventricular tachycardia in the presence of left ventricular ejection fraction < .40 after myocardial infarction who have sustained ventricular tachycardia inducible by electrophysiologic study are at significant risk for sudden death. This risk is significantly reduced by ICD, but not pharmacologic, antiarrhythmic therapy. Our major challenge at this time is not how best to treat high risk patients, but how best to identify them prior to events. Finally, physicians should be aware that many symptomatic ventricular tachycardias are now curable at low risk, using catheters to deliver radiofrequency energy. PMID- 11272664 TI - Shock, sepsis, UTI, and DRGs: a mistranslation. PMID- 11272663 TI - Delaying the diagnosis of abdominal pain with analgesics. PMID- 11272665 TI - Trends and patterns in health insurance coverage, 1991-2000. PMID- 11272666 TI - Point of view--protecting patients. PMID- 11272667 TI - Is brachytherapy comparable with radical prostatectomy and external-beam radiation for clinically localized prostate cancer? AB - PURPOSE: The aim of this study was to summarize the prostate brachytherapy literature and provide a comparative analysis of brachytherapy, radical prostatectomy, and external-beam radiation therapy outcomes for early-stage carcinoma of the prostate gland. MATERIALS AND METHODS: Published literature on brachytherapy, radical prostatectomy, and external-beam radiation therapy for clinically localized carcinoma ofthe prostate gland was reviewed. In addition, MEDLINE searches were performed to ensure completeness of the knowledge base. RESULTS: For patients with low-risk features, the biochemical results of prostate brachytherapy are as favorable as the most positive radical prostatectomy and external-beam radiation therapy series. In most studies, patients with intermediate- and high-risk disease have more durable biochemical outcomes when treated with brachytherapy (with or without external-beam radiation therapy). Long-term urinary morbidity is primarily restricted to patients with a history of transurethral resection. Significant bowel dysfunction is uncommon. Although erectile dysfunction occurs in approximately 50% of patients at 5 years, 80% respond favorably to sildenafil. Multiple postoperative dosimetric studies supported the ability of brachytherapists to adequately encompass the target volume. Compared with radical prostatectomy and external-beam radiation therapy, the total cost of prostate brachytherapy is 20% less. CONCLUSIONS: With prostate specific antigen-based follow-up as long as 10 years, the results of prostate brachytherapy for low-risk patients are as favorable as the most positive radical prostatectomy and external-beam radiation therapy series. In most reports, intermediate- and high-risk patients have more durable biochemical outcomes when managed by brachytherapy approach (with or without external-beam radiation therapy). Serious complications following brachytherapy are relatively rare. PMID- 11272668 TI - Complete laparoscopic approach for radical cystectomy and continent urinary diversion (sigma rectum pouch). AB - Technical and manual progress made in recent years now enables large uro oncological procedures to be performed by means of laparoscopy. We report the first successful radical laparoscopic cystectomy and laparoscopic construction of a continent urinary diversion. Laparotomy can be avoided completely. The advantages are clear reduction of blood loss and postoperative morbidity with faster convalescence. PMID- 11272669 TI - Orthotopic bladder replacement to the urethra following salvage radical cystoprostatectomy in men with failed radiation therapy. AB - PURPOSE: Salvage cystoprostatectomy has evolved as a safe and potentially curative treatment option for patients with radiation recurrent bladder cancer. Although orthotopic bladder replacement remains the preferred form of urinary diversion, there is minimal information about its role in salvage cystectomy series. We describe our limited experience in this regard. MATERIAL AND METHODS: We evaluated the operative characteristics and outcome of two patients with muscle invasive transitional cell carcinoma (TCC) of the urinary bladder after failed high-dose radiation therapy (mean 6,490 cGy). Both patients underwent salvage cystoprostatectomy with Studer-type ileal neobladder reconstruction. Existing literature on the topic is reviewed. RESULTS: Final histopathology showed pT3 N0 M0 TCC urinary bladder with no recent evidence of tumor recurrence. There was no mortality or major perioperative complication. Mean surgery time was 590 minutes, and mean blood loss was 1,600 mL, with 3.5 U of packed cell transfusion per patient. Mean length of stay was 15 days. Postoperative complications included urinary tract infection in both cases. Prolonged urinary leakage, metabolic derangements, and loose stools were seen in one case. At a mean follow-up of 17 months, both patients have well-preserved upper tracts, normal renal function, good capacity neobladders, and satisfactory postvoid emptying. Both patients are fully continent. CONCLUSIONS: Salvage cystoprostatectomy with orthotopic bladder replacement is a safe and effective management option in a select group of radiation recurrent bladder cancer patients. PMID- 11272671 TI - Feasibility of early catheter removal after radical retropubic prostatectomy. AB - PURPOSE: This study was conducted to determine if early (1 week) removal of the urethral catheter after radical prostatectomy is feasible. MATERIAL AND METHODS: Eighty patients underwent surgery from 1992 to 1999. Of the 78 patients with analyzable results, 22 (28%) had the catheter removed after 3 weeks (group 1) and 56 (72%) after 1 week (group 2). RESULTS: Median follow-up of 49 months revealed no mortality or major morbidity in the two groups of patients. Urinary functions were satisfactory and similar in both groups of patients. Group 2 patients had an improved continence rate of 92%, achieving full continence at 3 months, compared to 59% for group 1. The mean duration of hospitalization of 8.2 days for group 2 was better than the 12.1 days for group 1. CONCLUSIONS: Early removal of the catheter after radical prostatectomy was feasible, did not impose any short- or long-term morbidity, and may offer some benefits. PMID- 11272670 TI - Utility of capromab pendetide (ProstaScint) imaging in the management of prostate cancer. AB - PURPOSE: Capromab pendetide (ProstaScint) is an indium In 111 ((111)In)-labeled monoclonal antibody to prostate-specific membrane antigen (PSMA) used to image prostate cancer. The appropriate techniques for obtaining images with this modality and the appropriate clinical indications for this study are in the process of being optimized. MATERIALS AND METHODS: From 1994 to 2000, 631 monoclonal antibody imaging studies with (111)In capromab pendetide were performed. The accuracy and utility of this modality in the primary staging of patients with disease at high risk of metastasis and patients with recurrent or residual disease after primary therapy were evaluated. RESULTS: In high-risk patients evaluated for risk of lymph node metastases prior to pelvic lymph node dissection, capromab pendetide imaging was found to have a positive predictive value (PPV) of 62%, negative predictive value (NPV) of 72%, sensitivity of 62%, and specificity of 72%. In patients evaluated with capromab pendetide imaging for prostatic fossa recurrence using prostatic fossa needle biopsy as the gold standard, capromab pendetide imaging was found to have a PPV of 50%, NPV of 70%, sensitivity of 49%, and specificity of 71%. CONCLUSIONS: The sensitivity and NPV of (111)In capromab pendetide imaging are better than those of computed tomography and magnetic resonance imaging for detection of soft-tissue and nodal metastases from prostate cancer. The utility of this modality has been demonstrated in the primary staging of patients with disease at high risk of metastasis. Patients with recurrent or residual disease after primary therapy also may benefit from capromab pendetide imaging prior to selection of salvage therapy. Innovative methods for the use of capromab pendetide imaging in radiation therapy treatment planning are under development. PMID- 11272673 TI - Case 2. Umbilical discharge. A 3-month-old female has persistent drainage from her umbilicus. PMID- 11272672 TI - Case 1. Abdominal mass. A 1-year-old with a left abdominal mass and gross hematuria. PMID- 11272674 TI - Case 3. Adolescent scrotal fullness. An 11-year-old male with left scrotal fullness. PMID- 11272675 TI - Case 4. Abdominal wall abnormality. Newborn with congenital anomaly. PMID- 11272676 TI - Case 5. 31-month-old female with recurrent urinary tract infection. PMID- 11272677 TI - Treatment of interstitial cystitis with a quercetin supplement. AB - PURPOSE: Interstitial cystitis (IC) is a disorder of unknown etiology with few effective therapies. Oral bioflavonoid therapy utilizing quercetin recently proved to be clinically effective in men with chronic pelvic pain syndrome, a disorder with similarities to IC. We therefore tested in an open-label trial a quercetin-based supplement in patients with clinically proven IC. MATERIALS AND METHODS: Twenty-two patients (5 men and 17 women; average age 53.1 years) with classically documented IC received one capsule of Cysta-Q complex (equivalent to 500 mg of quercetin) twice a day for 4 weeks. Symptoms were assessed before and after therapy by the IC problem and symptom indices as well as by global assessment of pain (range 0-10). RESULTS: Two patients did not complete the study. In the remaining 20 patients, improvement was seen in all three parameters tested. After 4 weeks of treatment, the mean (+/- SEM) problem index improved from 11.3 +/- 0.6 to 5.1 +/- 0.7 (p = .000001), the mean symptom index improved from 11.9 +/- 0.9 to 4.5 +/- 0.5 (p = .000001), and the mean global assessment score improved from 8.2 +/- 0.4 to 3.5 +/- 0.4 (p = .000001). None of the patients experienced any negative side effects, and all but one patient had at least some improvement in every outcome measure. CONCLUSION: Oral therapy with the quercetin supplement Cysta-Q was well tolerated and provided significant symptomatic improvement in patients with IC. Larger, randomized, placebo controlled trials appear warranted based on these preliminary open-label results. PMID- 11272678 TI - Oral gabapentin (neurontin) treatment of refractory genitourinary tract pain. AB - PURPOSE: Refractory genitourinary pain is a common but difficult condition to treat. Examples of chronic genitourinary pain include orchalgia, interstitial cystitis, pain after bladder suspension surgery, nonbacterial prostatitis, and genital pain related to lumbosacral neuropathy. We report our experience with oral gabapentin treatment for this condition. Gabapentin is an anticonvulsant with unclear but therapeutic effects on neurologic pain. MATERIALS AND METHODS: Twenty-one patients referred with refractory genitourinary pain were treated with oral gabapentin. There were 9 men and 12 women. In the male patients, the location of pain was testicle (4), bladder (2), penis (1), or prostate (2). In female patients, the pain was located in the urethra (4), bladder (6), vulva (1), or vagina (1). The dose of gabapentin was titrated from 300 up to 2,100 mg/day. Subjective pain severity and 10-cm visual pain scale was used before and 6 months after therapy. RESULTS: The mean dose of gabapentin was 1,200 mg/day (range 300 2,100 mg). Ten of 21 patients reported subjective improvement of their pain. The remaining patients did not perceive any improvement. Gabapentin was well tolerated; only 4 patients dropped out due to side effects. The most common adverse effects were dizziness and drowsiness. Five of 8 patients with interstitial cystitis reported improvement. CONCLUSIONS: Although only 10 of 21 patients improved with gabapentin, this cohort included only patients with refractory genitourinary pain that failed a wide range of prior treatments. Gabapentin belongs in the armaterium of the urologist who treats genitourinary pain. PMID- 11272679 TI - Vesicoureteral reflux after ureteroneocystostomy: indications for postoperative voiding cystography. AB - PURPOSE: The aim of this study was to determine the risk factors for vesicoureteral reflux following ureteral reimplantation to identify a population that can be safely excluded from postoperative voiding cystography. MATERIALS AND METHODS: We retrospectively reviewed the medical records of 273 patients who underwent ureteroneocystostomy for vesicoureteral reflux between 1990 and 1998 and recorded the postoperative renal ultrasonography and voiding cystography results. RESULTS: There were 273 patients (534 ureters) who underwent ureteral reimplantation. We recorded the grade of preoperative hydronephrosis and vesicoureteral reflux and noted several preoperative and intraoperative variables, such as dysfunctional voiding, breakthrough infections, renal scarring, bladder trabeculations, type of reimplant, and postoperative urinary tract infections. With a mean follow-up of 20.6 months, persistent postoperative vesicoureteral reflux was noted in 11 patients (4%). Persistent postoperative reflux was noted in 11 patients (4%) or 12 renal units (2.2%). Reflux resolution rates for 534 renal units and 273 patients after routine follow-up voiding cystourethrogram (VCUG) was 97.8% (renal units) and 96% (patients), respectively. Contralateral vesicoureteral reflux developed in 4 (5.1%) of the 78 patients who underwent unilateral reimplantation. Two patients (0.7%) had postoperative ureteral obstruction. The risk factors for persistent postoperative reflux were identified as preoperative and postoperative hydronephrosis, renal scarring, and tapered reimplantations. The type of reimplant did not correlate with outcome. CONCLUSIONS: Vesicoureteral reflux after ureteral reimplantation is uncommon (4%). Because of the high success rate of ureteral reimplants and the benign course of those patients with persistent low-grade postoperative reflux, it is safe and efficient to eliminate postoperative VCUG in most patients who had a simple ureteral reimplantation for reflux. However, in some higher-risk patients, such as those with preoperative hydronephrosis, renal scarring, and ureteral tapering, postoperative voiding cystography may be indicated to assure resolution of vesicoureteral reflux. PMID- 11272680 TI - Postoperative cystography is unnecessary following renal transplantation with parallel incision extravesical ureteroneocystostomy. AB - PURPOSE: To determine the need for postoperative cystography following extravesical ureteroneocystostomy for renal transplantation. MATERIALS AND METHODS: The clinical courses of 200 consecutive kidney transplant recipients who underwent urinary tract reconstruction by parallel incision extravesical ureteroneocystostomy were reviewed. RESULTS: Five of the 200 recipients did not have the study because of early mortality (1) or medical problems (4). Grade I vesicoureteral reflux was present in 5 (3%) of 182 unstented allograft ureters and 5 of 13 stented allograft ureters. Two patients (1%) underwent repeat ureteroneocystostomy, one for obstruction and one for extravasation. The cystograms were normal in both patients. CONCLUSIONS: Routine retrograde cystography is unnecessary following urinary tract reconstruction by parallel incision extravesical ureteroneocystostomy. PMID- 11272681 TI - Comparison and clinical evaluation of hand-assist devices for hand-assisted laparoscopy. AB - Hand-assisted laparoscopic surgery (HALS) is being used increasingly in urologic laparoscopy, particularly for laparoscopic nephrectomy. Hand-assist devices (HADs) facilitate the intra-abdominal placement of the hand during laparoscopy. There are currently three HADs available in the United States: the Pneumo Sleeve, the Handport, and the Intromit. The performance of each HAD is assessed regarding usage options, maintenance of pneumoperitoneum, device failure, exchange of intra abdominal hands, adaptation to obese patients, and specimen removal. The use of these devices is reviewed based on our experience in more than 100 cases of HALS. PMID- 11272682 TI - Southern Illinois University (SIU) sling-bone anchored semitendinosus. AB - We describe a new type of pubovaginal sling that uses autologous fascia and minimizes the discomfort of harvest. The harvest is technically easy. The results are comparable to those of other slings previously described. PMID- 11272683 TI - Use of in situ spermatic cord patch for inguinal lymph node dissection. AB - Inguinal lymph node dissection for diagnosis of metastatic squamous cell carcinoma of the penis can cause significant morbidity and mortality for patients due to local wound breakdown, lymphedema, and vascular erosion. Various methods have been described to cover exposed femoral vessels to preserve their integrity, the most common being transposition of the sartorius muscle. We describe the successful use of in situ spermatic cord for coverage of the femoral artery and vein after inguinal lymph node dissection for squamous cell carcinoma of the penis in two patients. To our knowledge, this has not been previously described and is a simple and successful alternative way to cover the femoral vessels after inguinal lymphadenectomy. PMID- 11272684 TI - Practical approach to terminate urinary extravasation: percutaneous fistula tract embolization with N-butyl cyanoacrylate in a case with partial nephrectomy. AB - A 35-year-old woman who underwent partial nephrectomy had prolonged postsurgical urinary extravasation that led to a percutaneous fistula. A double-J catheter used as a ureteral stent during surgery was in place. A percutaneous pigtail nephrostomy was inserted on the 15th postoperative day but drainage continued. Antegrade pyelography demonstrated extravasation at the lower pole calyx. The double-J stent was removed on the 21st postoperative day, and a retrograde pyelogram showed no obstruction. Because drainage still was excessive on the 25th postoperative day, the fistula tract was embolized percutaneously with N-butyl cyanoacrylate, a tissue adhesive material. Drainage ceased immediately after the procedure, and control pyelography confirmed no extravasation. The patient was discharged on the 28th postoperative day. The patient had no additional complications at 36-month follow-up. PMID- 11272685 TI - Hand-assisted laparoscopic nephroureterectomy: description of technique. AB - PURPOSE: Traditional treatment of transitional cell carcinoma of the upper urinary tract (UTTCC) has been nephroureterectomy by open surgical techniques, often requiring two incisions. Our experience and technique for hand-assisted laparoscopic nephroureterectomy (HALNU) is reviewed. MATERIALS AND METHODS: Thirty-two patients had HALNU performed by one of three surgeons from August 1998 to October 2000. The distal ureter and bladder cuff was resected laparoscopically and sutured closed in 15 patients and resected by combined cystoscopic and laparoscopic approach in 17 patients. RESULTS: The indication for surgery was UTTCC for 29 patients and benign conditions in 2 patients. The mean operating time (including initial cystoscopy) was 372 minutes (281-530), and the mean blood loss was 541 cc (50-3500). The mean hospital stay was 5.5 days (3-12). There were no positive surgical margins, local recurrences, trocar site seeding, or wound seeding. CONCLUSIONS: HALNU is an effective minimally invasive approach for the treatment of UTTCC. PMID- 11272686 TI - Effectiveness of denuding the intestinal mucosa by submucosal injection in the porcine model. AB - PURPOSE: Lack of mucosal regrowth on denuded bowel segments is considered a prerequisite for successful grafting of cultured urothelial cells for bladder augmentation. This study was designed to establish a technique for clean and complete de-epithelialization of the intestinal mucosa. MATERIALS AND METHODS: A segment of the small bowel was isolated in six microminipigs. The isolated segment was detubularized and the submucosa injected with HEPES-buffered saline, elevating the mucosa from the underlying tissue and allowing removal with ease. One portion was resected and, along with a portion of the removed mucosa and a sample of the full-thickness bowel, submitted for histologic evaluation. Another portion of the bowel segment remained denuded while the final denuded segment was covered with polyglactin mesh. After 3 days, 1 week, and 3 weeks, specimens from each of the bowel segments were collected for histologic assessment. Immunoperoxidase labeling was performed to confirm the presence or absence of mucosal regrowth. RESULTS: All intestinal segments showed no mucosal regrowth at the 3-day, 1-week, and 3-week incubation periods. Minimal reaction between the denuded bowel segment and the mesh was evident in all of the pigs. CONCLUSIONS: In the porcine model, intestinal mucosa can be removed completely using submucosal injection. This technique may be useful in future clinical studies involving bladder augmentation using denuded bowel. PMID- 11272687 TI - Acute and chronic urticaria. Challenges and considerations for primary care physicians. AB - Urticaria and angioedema are common dermatologic problems seen by primary care physicians. A carefully taken history, physical examination, specific tests, and skin biopsy often provide useful diagnostic information. In patients with chronic urticaria, urticarial vasculitis and diseases that mimic urticaria need to be ruled out. A variety of treatment options are available for patients with urticaria and urticarial vasculitis. Pharmacologic therapy is useful when the specific cause is undetermined. When a trigger has been identified, the patient must avoid exposure to it. Patient education is an important component of management and should include instructions on crisis management, particularly for patients who have angioedema or a tendency for anaphylaxis. PMID- 11272688 TI - Risky behaviors with asymptomatic HIV infection? PMID- 11272689 TI - Painful skin erosions and fever in an infant. Eczema herpeticum. PMID- 11272690 TI - Angina. PMID- 11272691 TI - Brugada syndrome also linked to sudden cardiac death. PMID- 11272693 TI - The spectrum of acute bronchitis. Using baseline factors to guide empirical therapy. AB - The optimal therapy for acute bronchitis depends on the causative pathogen and the presence or absence of underlying lung disease. Because there is no fast, reliable way to identify the pathogen, physicians have to rely on clinical judgment and epidemiologic characteristics. In this article, Drs Flaherty, Saint, Fendrick, and Martinez discuss how an evidence-based approach to treatment may help ensure that efficacious therapy is available in the future. PMID- 11272692 TI - Update on screening for type 2 diabetes. The why, who, how, and what of testing and diagnosing. AB - Type 2 diabetes can cause serious complications even as it remains undiagnosed. Screening is recommended only in people with risk factors for the disease. The ADA recommends FPG as the test of choice, but RPG is also a practical alternative because it is easier and more convenient. Frequency of rescreening if the first screen result is normal depends on the number of risk factors present. Physicians should realize the importance of a confirmatory test and not base a diagnosis of diabetes on a single value unless the value is so high that the diagnosis is unequivocal. PMID- 11272694 TI - Hepatic encephalopathy. Metabolic consequence of cirrhosis often is reversible. AB - Hepatic encephalopathy is a well-recognized clinical complication of chronic liver disease. About 30% of patients with cirrhosis die in hepatic coma. Hepatic encephalopathy can occur in patients with fulminant liver disease without evidence of portal-systemic shunting. These patients have increased intracranial pressure and brain edema with a deleterious clinical course and poor prognosis unless liver transplantation is available. The pathogenesis of portal-systemic hepatic encephalopathy probably is multifactorial, although the predominant causative agent appears to be ammonia. The molecular basis of neurotoxicity of ammonia or other agents implicated in the condition is poorly understood. Therapy includes timely recognition and correction of precipitating factors. Once the condition is manifested, standard therapy is acute administration of lactulose, a disaccharide that is undigested in the small intestine. Its beneficial action is not fully understood. The use of oral antibiotics and BCAAs is of some benefit in patients who do not respond to lactulose. Limitation of protein in the diet may be useful for short periods but is not recommended for long-term use because of potential worsening of already poor nutrition. Several experimental therapies based on potential pathogenetic mechanisms have not resulted in improved outcomes over standard therapy with lactulose. However, future research will likely focus on the correction of alterations in neurotransmission. It is hoped that newer therapies will provide protection from the putative neurotoxins that cause secondary defects in neurotransmission. PMID- 11272695 TI - Bleeding esophageal varices. How to treat this dreaded complication of portal hypertension. AB - Bleeding esophageal varices, one of the most feared complications of portal hypertension, contribute to the estimated 32,000 deaths annually attributed to cirrhosis. Successful control requires knowledge of the pertinent anatomy, underlying pathophysiology of portal hypertension, and natural history of gastro esophageal varices. Drs Hegab and Luketic review these topics and discuss the various prophylactic and therapeutic approaches to management, including pharmacologic agents, endoscopic sclerotherapy, and trans-jugular intrahepatic portosystemic shunt (TIPS). PMID- 11272696 TI - The doctor glut revisited. How much has changed in the physician workforce? PMID- 11272697 TI - Minimizing ascites. Complication of cirrhosis signals clinical deterioration. AB - Ascites is the most common complication of cirrhosis. Its development is associated with a grave prognosis; 50% of patients die within 2 years of diagnosis. An understanding of the analysis of ascitic fluid is essential for the appropriate management of patients with liver disease and ascites. The management of patients with ascites involves a combination of dietary, medical, and surgical approaches. Furthermore, patients with ascites are at risk for ascitic fluid infections and neurohormonal dysregulation that can lead to hepatorenal syndrome. Early recognition of these complications allows therapeutic interventions that minimize further clinical deterioration in already chronically ill patients. PMID- 11272698 TI - NMR-restrained docking of a peptidic inhibitor to the N-terminal domain of the phosphoenolpyruvate:sugar phosphotransferase enzyme I. AB - Starting from the NMR structure of the binary complex between the N-terminal domain of the unphosphorylated enzyme I (EIN) of the phosphoenolpyruvate:sugar phosphotransferase (PTS) and the histidine-containing phosphocarrier protein (HPr), a molecular model of the phosphorylated transition state of the related complex was established using constrained simulated annealing. The coordinates of the phosphorylated EIN enzyme were then used in a second step for flexible docking of a decapeptide inhibitor of EIN whose enzyme-bound conformation itself was determined by NMR using transferred nuclear Overhauser effects. Two phosphorylation models of the peptide inhibitor were investigated and shown to be both functional. Interestingly, one model is very similar to that of the complex between EIN and its natural substrate HPr. The present study demonstrates that NMR-guided flexible docking constitutes an interesting tool for docking highly flexible peptide ligands and facilitates the upcoming protein-based design of nonpeptide EIN inhibitors for discovering new antibiotics. PMID- 11272699 TI - Simulation of carbohydrate-protein interactions: computer-aided design of a second generation GM1 mimic. AB - The oligosaccharide of ganglioside GM1 [Galbeta1-3GalNAcbeta1-4(NeuAcalpha2 3)Galbeta1-4Glcbeta1-1Cer] is the cellular target of two bacterial enterotoxins: the cholera toxin (CT) and the heat-labile toxin of E. coli (LT). We recently reported that the pseudosaccharide 2 [Galbeta1-3GalNAcbeta1-4(NeuAcalpha2 3)DCCHD] is a high-affinity ligand for CT. and thus a functional mimic of GM1 (Bernardi, A., Checchia, A., Brocca, P., Sonnino, S. and Zuccotto, F., J. Am. Chem. Soc., 121 (1999) 2032-2036). In this paper we describe the design of a second-generation mimic, formally obtained from 2 by inverting the configuration of a single stereocenter, thus transforming a N-acetyl galactosamine into a N acetyl glucosamine. The design process involved modeling of the free ligand and its LT complex, followed by qualitative and quantitative comparison with the corresponding structures of 2. The protocol employed relied on both conformational search and molecular dynamics methodologies to account for the flexibility of both the ligand and the protein receptor. The conformational search of the LT:inhibitor complex showed that, compared to 2, the new compound can insert one more hydroxy group within the protein binding site. Molecular dynamics simulations showed that, in turn, this may trigger a series of rearrangements and reorientations of side chains and crystallographic water molecules in the toxin, leading to new H-bond contacts which may result in enhanced affinity of the new inhibitor. FEP calculations were performed by mutating the structure of 2 in solution and in the protein complex, and the prediction was made that the second-generation mimic should be a stronger binder than its parent compound. PMID- 11272700 TI - The configurational dependence of binding free energies: a Poisson-Boltzmann study of Neuraminidase inhibitors. AB - The linear finite difference Poisson-Boltzmann (FDPB) equation is applied to the calculation of the electrostatic binding free energies of a group of inhibitors to the Neuraminidase enzyme. An ensemble of enzyme-inhibitor complex conformations was generated using Monte Carlo simulations and the electrostatic binding free energies of subtly different configurations of the enzyme-inhibitor complexes were calculated. It was seen that the binding free energies calculated using FDPB depend strongly on the configuration of the complex taken from the ensemble. This configurational dependence was investigated in detail in the electrostatic hydration free energies of the inhibitors. Differences in hydration energies of up to 7 kcal mol(-1) were obtained for root mean square (RMS) structural deviations of only 0.5 A. To verify the result, the grid size and parameter dependence of the calculated hydration free energies were systematically investigated. This showed that the absolute hydration free energies calculated using the FDPB equation were very sensitive to the values of key parameters, but that the configurational dependence of the free energies was independent of the parameters chosen. Thus just as molecular mechanics energies are very sensitive to configuration, and single-structure values are not typically used to score binding free energies, single FDPB energies should be treated with the same caution. PMID- 11272701 TI - Does a diol cyclic urea inhibitor of HIV-1 protease bind tighter than its corresponding alcohol form? A study by free energy perturbation and continuum electrostatics calculations. AB - The cyclic urea inhibitors of HIV-1 protease generally have two hydroxyl groups on the seven-membered ring. In this study, free energy perturbation and continuum electrostatic calculations were used to study the contributions of the two hydroxyl groups to the binding affinity and solubility of a cyclic urea inhibitor DMP323. The results indicated that the inhibitor with one hydroxyl group has better binding affinity and solubility than the inhibitor with two hydroxyl groups. Therefore, removal of one hydroxyl group from DMP323 may help to improve the properties of DMP323. This is also likely to be true for other cyclic urea inhibitors. The study also illustrated the difficulty in accurate modeling of the binding affinities of HIV-1 protease inhibitors, which involves many possible protonation states of the two catalytic aspartic acids in the active site of the enzyme. PMID- 11272702 TI - Ligand-receptor docking with the Mining Minima optimizer. AB - The optimizer developed for the Mining Minima algorithm, which uses ideas from Genetic Algorithms, the Global Underestimator Method, and Poling, has been adapted for use in ligand-receptor docking. The present study describes the resulting methodology and evaluates its accuracy and speed for 27 test systems. The performance of the new docking algorithm appears to be competitive with that of previously published methods. The energy model, an empirical force field with a distance-dependent dielectric treatment of solvation, is adequate for a number of test cases, although incorrect low-energy conformations begin to compete with the correct conformation for larger sampling volumes and for highly solvent exposed binding sites that impose little steric constraint on the ligand. PMID- 11272703 TI - Warmr: a data mining tool for chemical data. AB - Data mining techniques are becoming increasingly important in chemistry as databases become too large to examine manually. Data mining methods from the field of Inductive Logic Programming (ILP) have potential advantages for structural chemical data. In this paper we present Warmr, the first ILP data mining algorithm to be applied to chemoinformatic data. We illustrate the value of Warmr by applying it to a well studied database of chemical compounds tested for carcinogenicity in rodents. Data mining was used to find all frequent substructures in the database, and knowledge of these frequent substructures is shown to add value to the database. One use of the frequent substructures was to convert them into probabilistic prediction rules relating compound description to carcinogenesis. These rules were found to be accurate on test data, and to give some insight into the relationship between structure and activity in carcinogenesis. The substructures were also used to prove that there existed no accurate rule, based purely on atom-bond substructure with less than seven conditions, that could predict carcinogenicity. This results put a lower bound on the complexity of the relationship between chemical structure and carcinogenicity. Only by using a data mining algorithm, and by doing a complete search, is it possible to prove such a result. Finally the frequent substructures were shown to add value by increasing the accuracy of statistical and machine learning programs that were trained to predict chemical carcinogenicity. We conclude that Warmr, and ILP data mining methods generally, are an important new tool for analysing chemical databases. PMID- 11272705 TI - Incidence of zoonotic diseases in military working dogs serving in Operations Desert Shield and Desert Storm. AB - The United States deployed 118 military working dogs (MWDs) to the Persian Gulf theater during Operations Desert Shield and Desert Storm. This study is a retrospective descriptive study of medical records of these deployed dogs, with the objective to determine whether there were infectious or parasitic diseases with a zoonotic potential in a sentinel population of MWDs that may be of concern to Persian Gulf veterans. Fifty-one percent of visits to veterinary treatment facilities during deployment were for illness or injury. Potential zoonotic conditions accounted for 21% of the total visits, 41% of the "sick-call" visits, and 63% of presentations for illness to veterinary treatment facilities. This study did not determine whether the diseases treated were transmitted between MWDs and the troops. Although the etiologic agents were not determined in these cases, no evidence was found supporting new or reemerging illnesses in this population of dogs. PMID- 11272704 TI - A theoretical approach to the influence of the macrocycle conformation on the molecular electronic structure in Mg-porphyrins. AB - Nonplanar saddled (sad) ruffled (ruf) and domed (dom) conformations of the Mg porphyrin (MgP) macrocycle in several degrees of deformation have been computed. These symmetrical distortion modes were induced in unsubstituted macrocycle using molecular definitions for calculations which permits us to achieve a systematical variation of the nonplanarity varying only a convenient geometrical parameter of the molecule. Series of nonplanar macrocycles like those synthesized in previous works employing peripheral substitutions are obtained. The procedure here used to induce deformations gives the possibility of investigating the modulator role of the out-of-plane distortions on the geometry and electronic properties of the porphyrin avoiding additional influences due to the substituents or the surrounding protein scaffolding. PMID- 11272706 TI - Psychiatric profiles in the U.S. Air Force: a clinical interpretation of Air Force Instruction 48-123. AB - The foundations of our current system for profiling military psychiatric patients were laid during World War II, well before the development of the first version of the Diagnostic and Statistical Manual of Mental Disorders. The general principles and terminology remain in use today through Air Force Instruction 48 123, Medical Examination and Standards. The terminology used is clearly outdated, making it difficult to use and risking misuse, deploying the wrong person or denying deployment to an appropriate person. Our objective is to review the current standards for making psychiatric profiles in the U.S. Air Force and propose a practical interpretation of the current Air Force Instruction. Considerable research remains to be done to improve our profile system, especially in light of the development of effective treatments for many psychiatric illnesses. Although prognostic data are available for some illnesses, little research has been done on military populations and essentially none of it considers the rigors of military deployment. Diagnosis, prognosis, duty environments, and demands of duties all must be considered in making profile decisions. Reductionistic approaches more simple than this will serve neither the commander nor the airman. PMID- 11272707 TI - Tuberculosis infection after humanitarian assistance, Guantanamo Bay, 1995. AB - Upon redeployment to Fort Lewis, Washington, from Operation Sea Signal in Guantanamo Bay, Cuba, 5% of a military police unit was identified as positive for purified protein derivative (PPD). A case-control study was conducted to document the number of converters and to identify risk factors among the soldiers for PPD conversion while in Cuba. Forty-six of the soldiers (3.7% of the unit) met the criteria for PPD conversion as a result of deployment. Forty-four converters and 84 controls completed surveys. Logistic regression showed that statistically significant independent risk factors for PPD conversion included working around coughing migrants (odds ratio [OR] = 6.73, 95% confidence interval [CI] = 2.2 20.4) and birthplace outside the United States (OR = 4.89, CI = 1.3-18.5). Contact in the psychiatric hospital (OR = 0.22, CI = 0.05-0.90) and contact with migrants with known tuberculosis (OR = 0.16, CI = 0.05-0.54) appeared to be protective factors, possibly because known tuberculosis patients and hospitalized patients most likely would be on treatment and rendered noninfectious. With the U.S. military's involvement in humanitarian and refugee operations in countries highly endemic for tuberculosis, service members are at increased risk of acquiring tuberculosis infection. Detection of tuberculosis infection and appropriate treatment should become a higher priority within the U.S. military. PMID- 11272708 TI - Attitudes and practices of military family physicians regarding obesity. AB - This study's objective was to define the current attitudes and practices of military family physicians regarding obesity. The authors mailed a cross sectional survey to 267 military family physicians selected randomly from the 1997 Uniformed Services Academy of Family Physicians membership database. A total of 214 surveys (80%) were returned. Most respondents believed that they should be role models to patients (93%) and felt obligated to counsel patients regarding obesity (90%). Fifty-six percent did not consider counseling obese patients professionally satisfying. Most correctly identified obesity as a risk factor for several diseases, except colon cancer (35%). Fifty-four percent correctly identified the current World Health Organization definition of obesity. A notable minority ascribed negative attributes of sadness (18%) and lack of self-control (25%) to obese individuals. The results of this survey indicate knowledge gaps and professional ambivalence regarding obesity in the study group. Methods of increasing family physician effectiveness in modifying this important risk factor deserve further study. PMID- 11272709 TI - Magnetic resonance imaging in a military setting: a utilization analysis. AB - A prospective study was conducted to evaluate the use of magnetic resonance imaging (MRI) by orthopedic surgeons and residents versus the use of MRI by non orthopedically trained health care providers in diagnosing knee pathology. Fifty patients initially evaluated by members of one of these groups who underwent subsequent knee MRI evaluation were selected to participate. Two orthopedic examiners individually examined all patients, recording clinical diagnosis and the merit of MRI evaluation in each case. Clinical accuracy, sensitivity, and specificity were compared between groups based on MRI findings. Diagnostic accuracy was similar; however, the orthopedic group displayed greater sensitivity, suggesting better clinical assessment. The study examiners observed both groups using MRI equally inappropriately and found 62% of the imaging studies unjustified. We conclude that knee MRI is used inappropriately in the current military setting. An algorithm is proposed to guide the future use of MRI in the diagnosis and management of knee pathology. PMID- 11272710 TI - Incidence and outcomes of total hip arthroplasty among U.S. Army aviators. AB - Among Army aviators, the incidence of total hip arthroplasty (THA) is unknown. This study analyzes the incidence and aeromedical disposition of THA among Army aviators. The U.S. Army Epidemiology Data Register was queried for a 10-year period from calendar year 1987 to 1996. The aviators selected for this study cohort were all qualified and on active flight status before undergoing THA. Data collected included age, gender, diagnoses, and aeromedical dispositions. There were 214,003 aviator-years of observation. Eleven aviators underwent 14 THAs. The overall incidence of THA was 0.05 per 1,000 aviator-years of observation. Of the 11 aviators who underwent THA during the study period, 4 received aeromedical suspension from flying duties (36%). THA is a rare medical event among Army aircrew members. Most are able to return to full flying duties with a waiver. Aircrew members younger than 50 years with THA are more likely to be suspended from aviation duties. PMID- 11272711 TI - U.S. Army noncombat munitions injuries. AB - OBJECTIVE: The object of this study was to determine the types of noncombat injuries secondary to munitions sustained by U.S. Army soldiers. METHODS: A retrospective review of all noncombat munitions injuries reported to the U.S. Army Safety Center from August 1989 to September 1996 was conducted. RESULTS: There were 742 incidents reported, resulting in 894 injured soldiers. The most common types of injuries were thermal burns, puncture wounds, and lacerations. The extremities were the most common anatomical location injured. The most common activities associated with injuries were combat training exercises, munitions firing, and rendering munitions safe. CONCLUSION: This study demonstrates a distinctive injury pattern for each category of munitions. Military readiness will be improved if we train all personnel to be familiar with the injury patterns and the most common situations associated with injury. By informing unit commanders which activities are associated with increased risk of injury, they may better prepare preventive measures to decrease the number of noncombat injuries. PMID- 11272712 TI - Values identified in different groups of Air Force nurses. AB - Fundamental personal values are reflected in the choices and decisions made in every aspect of our lives. This descriptive study identified values held by a convenience sample of 224 Air Force nurses stationed at four U.S. Air Force medical facilities. Study participants identified seven of eight literature supported values in the categories "important" or "very important" across the demographic factors of age, gender, educational level, military rank, marital status, and years of Air Force or civilian nursing experience. These seven values were ability utilization, achievement, altruism, autonomy, economic reward, economic security, and personal development. Personnel using this information may ease the transition process to military nursing, facilitate job placement to positions reflecting personally held values, and provide valuable insight for Air Force nurse recruiters who have limited knowledge of the nursing profession. In all, this would promote job satisfaction and Air Force nurse retention. PMID- 11272713 TI - Implementing a pain management service at an Army Medical Center. AB - In 1999, the Joint Commission for Accreditation of Healthcare Organizations published comprehensive pain management standards. Previous research has shown that pain control in people with cancer remains a significant problem in health care, even though cancer pain can be managed effectively in up to 90% of patients. In addition, postoperatively, many patients fail to have adequate pain control because of staff failures to routinely assess pain and pain relief. Many patients, if not questioned, silently tolerate unrelieved pain. National guidelines were published that address both acute and cancer pain in 1992 and 1994, respectively. In 1995, Tripler Army Medical Center dedicated $300,000 to create a 24-hour Pain Management Service to improve pain management. This article describes the structure of, educational programs offered by, and system changes implemented by the Pain Management Service. PMID- 11272715 TI - Work and sleep hours of U.S. Army aviation personnel working reverse cycle. AB - A one-page questionnaire was administered to 157 aviation personnel from three Army posts to determine when Army aviation personnel work and sleep while on reverse cycle. This project was undertaken as a first step to developing tailor made fatigue reduction strategies for shift workers in Army aviation. The results indicated that 97.6% of the surveyed aviation personnel had experience working night shift/reverse cycle at some point in their careers, with 69.4% working the night shift within the past 6 months. Of those who had experience working the night shift, 36.2% reported usually working from early in the afternoon to early in the morning, with 52.2% of personnel returning home by 4:00 a.m.; however, 28.3% arrived home after 8:00 a.m. Almost 62% of the respondents indicated that they did not feel they received adequate daytime sleep some of the time or at all while on reverse cycle/night shift. Research is needed to address the issue of helping aviation personnel sleep during the daylight hours, both for training exercises and for deployment. Once the work/rest schedule for a unit is known, countermeasures such as light therapy or gradual changes in scheduling can be tailored to meet the specific needs of the individual or unit. PMID- 11272716 TI - Acute pancreatitis in patients with hemorrhagic fever with renal syndrome. AB - The objective of this study was to determine the prevalence and clinical significance of acute pancreatitis in patients in whom hemorrhagic fever with renal syndrome (HFRS) has been diagnosed. We retrospectively reviewed all patients with a diagnosis of HFRS at our institution from 1994 to 1998. The review included medical records, laboratory results, radiologic examinations, and one autopsy report. From 1994 to 1998, 13 patients received diagnoses of HFRS that were confirmed by serology. In 9 patients (69%), serum amylase, serum lipase, or both were assessed during hospitalization. Seven (78%) of the 9 patients had pancreatitis. Four (57%) of these 7 patients with HFRS and pancreatitis had associated pulmonary edema, and 1 patient had rhabdomyolysis. In our small retrospective case series, acute pancreatitis in patients with HFRS was much more common than previously recognized. Patients with HFRS and pancreatitis had increased morbidity. However, the treatment for the associated pancreatitis was conservative. PMID- 11272714 TI - Variations in feminine hygiene practices of military women in deployed and noncombat environments. AB - There is limited information on how military women manage feminine hygiene practices in combat and noncombat environments. The purpose of this study was to describe feminine hygiene practices of military women in deployed and noncombat (normal) environments. A nonexperimental descriptive research design was used. The study used a survey questionnaire, the Deployed Female Health Practice Questionnaire, which was developed specifically for military women to report their experiences with hygiene issues. Significant differences between deployed and normal environments were found in the areas of types of menses management products used and in douching and handwashing practices. Continuing education about safe feminine hygiene practices will help military women cope better in deployed (field) environments. Recommendations suggest further study on intervention strategies for hygiene management practices. PMID- 11272717 TI - Oral health of ambulatory care patients. AB - OBJECTIVES: This project assessed the clinical oral health status of Veterans Administration (VA) patients and examined the relationship between oral health and both sociodemographic factors and dental care utilization. METHODS: Data were collected on 538 users of VA ambulatory medical care. Oral health was assessed by clinical examinations, and dental use and sociodemographic information are based on self-report. RESULTS: Younger, more educated VA patients with higher incomes had more teeth, fewer untreated and treated root caries, and were less likely to be edentulous or to have dentures. Dental utilization emerged as the most important aspect of veterans' oral health status, even after sociodemographic factors were controlled. Compared with the general population, veterans have poorer oral health with the exception of coronal caries. CONCLUSION: Compared with national studies, VA patients appear to have worse oral health. The importance of sociodemographic factors and dental utilization that has been found in other studies applies to veterans' oral health as well. PMID- 11272718 TI - Active surveillance of birth defects among U.S. Department of Defense beneficiaries: a feasibility study. AB - Since the Vietnam War, concern regarding the association of military exposures and birth defects has grown. The possibility of such associations remains a source of unease. To determine if such an association exists, birth defects surveillance among military families must be conducted. This project compared health record abstraction (active surveillance) with screening of Department of Defense electronic medical data (passive surveillance) to detect birth defects among San Diego County military families during the period January 1, 1997, through June 30, 1998. A total of 171 of 5,351 infants (3.2%) were identified as having a major defect, consistent with national civilian rates. There was approximately 80% concurrence between passive and active surveillance birth defect data, suggesting that a hybrid system of electronic data, supplemented with active surveillance in a specific region, represents a feasible and cost effective surveillance program for the geographically dispersed military population. PMID- 11272719 TI - Incorporating new recipes into the Armed Forces Recipe File: determination of acceptability. AB - As part of a project of decrease fat, cholesterol, and sodium in soldiers' diets, new ethnic and breakfast items were developed and standardized for 100 portions. Acceptability data were collected after initial recipe development, during recipe validation at a collaborating university, and in an actual Army garrison. Acceptability was determined using a nine-point hedonic scale; products rating > or = 6.0 in initial tests were prepared in garrison. Acceptability data were compared among test settings, ethnic categories, and food type. When grouped by ethnic categories, acceptability ratings varied more than when grouped by food type. Ratings varied most between development and validation settings (7.2 vs. 6.6; p < 0.05) and least between validation and actual Army settings (6.6 vs. 6.6; not significant). Because acceptability ratings were similar between the validation site and the Army garrison, future recipe development may continue without additional testing at actual Army garrisons, leading to more timely armed forces recipe file additions. PMID- 11272720 TI - A Naval Academy midshipman with ehrlichiosis after summer field exercises in Quantico, Virginia. AB - A case of human ehrlichiosis (caused by infection with Ehrlichia chaffeensis) is presented. The patient was a female Naval Academy midshipman with a 26-day history of daily field training with the U.S. Marines near Quantico, Virginia. She presented with a several-day history of myalgias, fever, and frontal headache. During her clinical course, she developed fever to 104 degrees F, dry cough, dyspnea on exertion, arthralgias, and nephrotic syndrome. She did not develop a rash. Laboratory studies were significant for thrombocytopenia, equivocal Lyme enzyme immunosorbent assay with a negative confirmatory western immunoblot, equivocal Rocky Mountain spotted fever acute serology without a convalescent increase in immunoglobulin G, and immunoglobulin G/immunoglobulin M serology positive for human monocytic ehrlichiosis. She manifested known sequelae for this emerging disease, including dyspnea, pedal edema, increased transminases, and nephrotic syndrome. PMID- 11272722 TI - Is the excited-state H-atom transfer in hypericin concerted? AB - The excited-state intramolecular H-atom transfer of hypericin (Hyp) was investigated as a function of pH in monodispersed reverse micelles formed by sodium bis(2-ethylhexyl)sulfosuccinate/heptane/water and in complexes with Tb3+ under conditions in which one of the two carbonyl groups of Hyp is incapable of accepting a hydrogen atom. The results of pump-probe transient absorption experiments provide no evidence for a concerted H-atom transfer mechanism. PMID- 11272721 TI - Randomized controlled trial of concurrent hepatitis A and B vaccination. AB - Hepatitis A and B viruses are threats to deployed military forces. The objective of this study was to determine the feasibility of concurrent vaccination against hepatitis A and B viruses. One hundred five healthy persons, 20 to 49 years of age and without serologic markers to hepatitis A or B viruses, were randomized to receive an inactivated hepatitis A vaccine (HEP A; 25 units in 0.5 mL), recombinant hepatitis B vaccine (HEP B; 10 micrograms in 1.0 mL), or both (HEP A & B) concurrently in separate arms. Vaccines were administered intramuscularly at 0, 1, and 6 months. Sera obtained at 1, 2, 6, 7, and 12 months after the first dose were tested for quantitative antibody to hepatitis A virus (anti-HAV) and antibody to hepatitis B surface antigen. Local reactions (e.g., pain) were reported by less than half of the volunteers and were similar at the site of HEP A, whether given alone or concurrently. However, more persons complained of pain (usually mild) at the HEP B site when HEP B was given concurrently with HEP A compared with HEP B alone (43% vs. 15%, 34% vs. 9%, and 42% vs. 15% for doses 1, 2, and 3, respectively; p < 0.05 for each dose). Among persons immunized with HEP A alone or HEP A & B, the proportion with > or = 10 mIU/mL anti-HAV was 83% in both groups 1 month after dose 1 and 100% at months 2, 7, and 12. The geometric mean concentrations of anti-HAV increased from 21 mIU/mL at month 1 to 2,649 and 2,312 mIU/mL in the HEP A and HEP A & B groups, respectively, at month 7. The response to HEP B was similar whether administered alone or concurrently. Antibody responses were similar in those receiving HEP A or HEP B concurrently or alone, but more subjects reported pain (usually mild) at the HEP B site after concurrent vaccination than after HEP B alone. Further work should be conducted to approve HEP A for patients younger than 2 years of age and to develop combined HEP A and HEP B vaccines in the United States. PMID- 11272724 TI - UV-induces formation of hydrogen peroxide based on the photochemistry of ketoprofen. AB - Ketoprofen (KP) is a potent nonsteroidal anti-inflammatory drug. However, application to the skin is problematic because the photosensitizing properties of the benzophenone moiety may cause phototoxic effects when the treated skin region is exposed to UVA light. Using capillary electrophoresis with electrochemical detection we are able to differentiate the peroxides formed during illumination of KP-containing solutions of linoleic acid. Contrary to other profens a high amount of hydrogen peroxide was found among the reaction products. For investigation of the skin damaging effect human keratinocytes were used as models. Cell viability, DNA synthesis efficiency and intracellular concentration of peroxides were determined. Viability and proliferation behavior was not altered under the influence of KP. While lower concentrations of KP (10-100 nM) led to a protection against the UVA-induced (8 J/cm2) cell proliferation damage, higher concentrations (10-100 microM) led to an amplification of the proliferation decrease. With UVB irradiation at relevant doses the effects were lower than using UVA. Furthermore, intracellular peroxide content was increased after UV irradiation and KP addition. In conclusion some efforts have to be done to avoid these side effects in the use of KP for topical or transdermal application. PMID- 11272723 TI - Solvatochromic effects in the electronic absorption and nuclear magnetic resonance spectra of hypericin in organic solvents and in lipid bilayers. AB - The natural product hypericin was tested in recent years as a biological photosensitizer with a potential for viral and cellular photodamage. We thus studied extensively its spectroscopy and membrane partitioning. Absorption, fluorescence excitation and emission spectra of the sodium salt (HyNa) were measured in 36 protic and aprotic, polar and apolar, solvents. Electronic transition bands as well as vibrational progressions were identified. Aggregation in some nonpolar solvents and protonation in organic acids were demonstrated. Modeling solvatochromism was done by Lippert equation, by the ET(30) parameter and by the Taft multiparameter approach. In all cases, separation into protic and aprotic solvents gave much better fits to the models. 13C chemical shift data could also be correlated with solvent polarity. They correlated best with Lippert's delta f polarity measure, but tended to fall into two distinct solvent groups--each along different lines--corresponding to protic and aprotic media, respectively. This interesting phenomenon suggests that in the case of the charged and slightly water soluble HyNa, two mechanisms of solvation are involved, each resulting in its own line equation. In aprotic media, dipole dipole interaction is the predominant solvation mechanism. In protic solvents, the most effective means of solvation is likely to be hydrogen bonding. When intercalated into the liposomal phospholipid bilayer, HyNa is oriented at an angle to the interface, thus experiencing a gradient of solvent polarities: a highly polar environment (similar to methanol) for C-2/5, suggesting that they lie not far from the interface; a moderately polar environment (similar to that of n-propanol) for C-6a/14a, which are somewhat deeper within the bilayer; and a more lipophilic environment (akin to n-hexanol) for C-10/11. The fluorescence excitation peak in liposomes also correlates with an aprotic medium of relatively high polarity, as might be excepted from a molecule in a shallow position in the bilayer. PMID- 11272725 TI - Alternative forms of formamidopyrimidine-DNA glycosylase from Arabidopsis thaliana. AB - Formamidopyrimidine-DNA glycosylase (FPG) catalyzes the initial steps in the repair of DNA containing oxidized purines. Two complementary DNA clones encoding homologs of bacterial FPG, designated Atfpg-1 and Atfpg-2, have been isolated from Arabidopsis thaliana. They are products of alternative splicing of the transcript of a single gene. Proteins encoded by both clones, AtFPG-1 and AtFPG 2, engineered to contain oligohistidine sequences on their C-terminal ends, were expressed in Escherichia coli and purified, and their activities were assayed. Both proteins cleaved DNA that contained apurinic sites, indicating that they have abasic lyase activity. AtFPG-1, but not AtFPG-2, showed significant cleavage of a double-stranded oligonucleotide that contained 8-oxo-guanine, indicating that the structural differences between the two proteins influence their enzymatic activities. However, both proteins were able to cleave the same sites in DNA that was treated with visible light in the presence of methylene blue. PMID- 11272726 TI - Diffuse solar UV radiation and implications for preventing human eye damage. AB - Ocular UV exposure is a function of both the direct and diffuse components of solar radiation. Broadband global and diffuse UV measurements were made in the morning, noon and afternoon. Thirty sets of measurements were made in summer and 50 in each of the other seasons at each of the periods in full sun. Corresponding sets were made in the shade of Australian evergreen trees: 42 trees in summer and 50 in each of the other seasons. The percentage diffuse UV was higher for the shorter 320-400 nm range (UVB) than for 280-320 nm (UVA). The percentage diffuse UVB ranged from 23 to 59%, whereas the percentage diffuse UVA ranged from 17 to 31%. The percentage diffuse UV was lower at noon than in the morning and afternoon with the difference more pronounced for the UVB. The average percentage diffuse UVB over all the measurements in the tree shade for the morning, noon and afternoon was 62, 58 and 71%, respectively, and the average percentage diffuse UVA was 52, 51 and 59%, respectively. PMID- 11272727 TI - Effects of UVB irradiance on conidia and germinants of the entomopathogenic Hyphomycete Metarhizium anisopliae: a study of reciprocity and recovery. AB - We tested the effects of irradiances of 920 and 1200 mW m-2 (weighted irradiance) on the conidia and germinants of the entomopathogenic Hyphomycete Metarhizium anisopliae. The conidia were exposed to the two irradiances for 1, 2, 4, 6, 7 or 8 h. Increased exposure decreased relative percent culturability. The inactivation provoked by the irradiance of 1200 mW m-2 was higher than for the 920 mW m-2, with a reduction in the 50% lethal time (LT50) from 6 h 40 min to 4 h 26 min. Reciprocity was not observed when conidia in water suspension and germinants in different stages of the germinative process were exposed to a 17.3 kJ m-2 total dose at both irradiance levels. Although nonreciprocity was observed in all situations, its magnitude varied as a function of metabolic state and/or cell-cycle phase in which the conidia were at the exposure time. The least difference between the effects of the two irradiance levels was observed when nongerminating conidia in suspension were exposed, and the greatest was observed when conidia were exposed during an advanced germination phase. Doses of 6.6 and 17.3 kJ m-2 supplied through the two irradiance levels delayed the germination of the surviving conidia. At both doses, delay was greater during exposure to the higher irradiance. Nonreciprocity was higher for the 17.3 kJ m-2 dose. Nonreciprocity magnitude, in addition to depending on the conidial physiological state, also depended on dose. The results demonstrate the importance of evaluating the impact of the increase in irradiance during the different stages of the fungal life cycle, especially during the stages which are more sensitive to UV, and not simply in dormant conidia. PMID- 11272728 TI - Wavelength-specific activation of MAP kinase family proteins by monochromatic UV irradiation. AB - The depletion of stratospheric ozone causes related increase in UV light below about 310 nm, which significantly affects biological and ecological systems. To understand the wavelength-specific effects of UV light, Molt4 cells (human T lymphoma cells) were irradiated with a series of monochromatic UV lights and the activities of three members of the mitogen-activated protein (MAP) kinase group were examined. Extracellular signal-regulated kinase was specifically activated within 1 min after UV irradiation in the range 320-360 nm. In contrast, P38 kinase was activated by 270-280 nm light with a peak at 1 min after irradiation. c-Jun N-terminal kinase activation was observed in a narrow range of UV light with a sharp peak at 280 nm occurring in 10 min. JNK translocated from the cytosol to the nucleus upon irradiation, while P38 remained in the cytosol even after UV irradiation. The activation of three MAP kinases was prevented by antioxidant reagents, suggesting that an oxidative signal initiates these responses. These results confirm that UV light affects various cellular functions through the activation of intracellular signaling systems including MAP kinase family proteins. However, the UV-induced activities of the separate MAP kinases show distinctly different dose, time and wavelength dependencies. PMID- 11272729 TI - Supramolecular structure of self-assembled synthetic zinc-13(1)-oxo-chlorins possessing a primary, secondary or tertiary alcoholic 3(1)-hydroxyl group: visible spectroscopic and molecular modeling studies. AB - Zinc-chlorin 3 (see Fig. 2 in text) possessing a tertiary 3(1)-hydroxyl group and a 13-keto group was synthesized as a model for the antenna chlorophylls of green bacteria. Self-aggregation of 3 in nonpolar organic media was examined and compared to 1 and 2 possessing a primary and secondary 3(1)-hydroxyl group, respectively. Zinc-chlorin 3 self-aggregated in 1 vol% CH2Cl2-hexane to form oligomers and showed a red-shifted Qy maximum at 704 nm compared to the monomer (648 nm in CH2Cl2). This red-shift is larger than that of 2S (648-->697 nm) and comparable to that of 2R (648-->705 nm), but smaller than that of 1 (648-->740 nm), indicating that while a single 3(1)-methyl group (prim-OH-->sec-OH) suppressed close and/or higher aggregation, the additional 3(1)-methyl group (sec OH-->tert-OH) did not further suppress aggregation. The relative stability of the aggregates was in the order 1 > 2R-3 > 2S as determined by visible spectral analyses. Molecular modeling calculations on dodecamers of zinc-chlorins 1, 2R and 3 gave similar well-ordered energy-minimized structures, while 1 stacked more tightly than 2R and 3. In contrast, 2S gave a relatively disordered (twisted) structure. The calculated dodecameric structures could explain the visible spectral data of 1-3 in nonpolar organic media. PMID- 11272730 TI - 5-Aminolaevulinic acid methyl ester transport on amino acid carriers in a human colon adenocarcinoma cell line. AB - The transport mechanisms of 5-aminolevulinic acid methyl ester (5-ALA-ME) have been studied in a human adenocarcinoma cell line (WiDr) by means of 14[C]-labeled 5-ALA-ME. The transport was found to be partly Na+ dependent, while the extracellular Cl- concentration did not affect the uptake. The transport of 5-ALA ME into WiDr cells was dependent on the incubation temperature and was found to be completely blocked by the inhibitors of energy metabolism, 2-deoxyglucose and sodium azide. WiDr cells were treated with 10 mM of 14 different amino acids and the substrate specificity of the 5-ALA-ME transporter(s) was analyzed by treating the cells with 23 microM or 1 mM 14[C]-labeled 5-ALA-ME. The transport of 5-ALA ME was found to be inhibited to the highest extent, i.e. about 60%, by the nonpolar amino acids L-alanine, L-methionine, L-tryptophan and glycine. The uptake of 5-ALA-ME followed an exponential decay with increasing concentration of glycine, reaching a maximum inhibition of uptake of 5-ALA-ME of 55%. Sarcosine, a specific inhibitor of system Gly, did not significantly inhibit 5-ALA-ME transport. In contrast to transport of 5-ALA, 5-ALA-ME does not seem to be taken up by system BETA transporters. In conclusion, the cellular uptake of 5-ALA-ME into WiDr cells seems to be due to active transport mechanisms, involving transporters of nonpolar amino acids. PMID- 11272731 TI - Activation of the IL-10 gene promoter following photodynamic therapy of murine keratinocytes. AB - Photodynamic therapy (PDT), an anticancer treatment modality, has recently been shown to be an effective treatment for several autoimmune disease models including antigen-induced arthritis. PDT was found to induce the expression of IL 10 messenger RNA (mRNA) and protein in the skin, and this expression has similar kinetics to the appearance of PDT-induced suppression of skin-mediated immune responses such as the contract hypersensitivity (CHS) response. Some aspects of the UVB-induced suppression of the immune response have been linked to the induction of IL-10. IL-10 has been shown to inhibit the development and activation of Th1 cells, which are critical for many cell-mediated immune responses, including CHS. We have examined the effect of PDT and UVB irradiation on the activity of the IL-10 gene promoter and on IL-10 mRNA stability using the murine keratinocyte line, PAM 212. In vitro PDT induces IL-10 mRNA and protein expression from PAM 212 cells, which can be correlated with an increase in AP-1 DNA binding activity and activation of the IL-10 gene promoter by PDT. Deletion of an AP-1 response element from the IL-10 gene promoter was shown to abrogate the PDT-induced promoter activity indicating that the AP-1 response element is critical to IL-10 induction by PDT. In addition, PDT results in an increase in IL 10 mRNA stability, which may also contribute to the increased IL-10 expression in PAM 212 cells following PDT. In vitro UVB irradiation also results in activation of the IL-10 promoter. However, in contrast to PDT, UVB-induced activation of the IL-10 promoter is not AP-1 dependent and did not increase IL-10 mRNA stability. PMID- 11272732 TI - In vivo fluorescence spectroscopy of nonmelanoma skin cancer. AB - In vivo and ex vivo tissue autofluorescence (endogenous fluorescence) have been employed to investigate the presence of markers that could be used to detect tissue abnormalities and/or malignancies. We present a study of the autofluorescence of normal skin and tumor in vivo, conducted on 18 patients diagnosed with nonmelanoma skin cancers (NMSC). We observed that both in basal cell carcinomas (BCC) and squamous cell carcinomas (SCC) the endogenous fluorescence due to tryptophan residues was more intense in tumor than in normal tissue, probably due to epidermal thickening and/or hyperproliferation. Conversely, the fluorescence intensity associated with dermal collagen crosslinks was generally lower in tumors than in the surrounding normal tissue, probably because of degradation or erosion of the connective tissue due to enzymes released by the tumor. The decrease of collagen fluorescence in the connective tissue adjacent to the tumor loci was validated by fluorescence imaging on fresh frozen tissue sections obtained from 33 NMSC excised specimens. Our results suggest that endogenous fluorescence of NMSC, excited in the UV region of the spectrum, has characteristic features that are different from normal tissue and may be exploited for noninvasive diagnostics and for the detection of tumor margins. PMID- 11272733 TI - Inhibition of cutaneous UV light-induced tumor necrosis factor-alpha protein production by Allotrap 1258, a novel immunomodulatory peptide. AB - Peptides derived from the heavy chain of the HLA Class-I molecules have been shown to modulate immune responses both in vivo and in vitro. Using a computer aided rational drug design approach, novel immunomodulatory peptides were designed based on peptide 2702.75-85, derived from HLA-B2702. Several peptides were identified which had increased immunomodulatory activity, including peptides RDP1258 and its D-isomer the peptide Allotrap 1258. The present study using Skh/hr hairless mouse skin model evaluated the in vivo effects of Allotrap 1258 on acute UVB-induced skin inflammation. Here we demonstrate that intraperitoneal administration of Allotrap 1258 1 h prior to UV exposure resulted in significantly diminished levels of UV-induced tumor necrosis factor (TNF)-alpha protein production in the epidermis but had no effect on other parameters of the acute UV-induced inflammatory response. By virtue of its ability to suppress TNF alpha protein production, Allotrap 1258 could prove to be an effective modulator of inflammatory responses. PMID- 11272734 TI - Postirradiation hyperthermia selectively potentiates the merocyanine 540 sensitized photoinactivation of small cell lung cancer cells. AB - Lung cancer has long been considered a disease that might benefit from the dose escalation of radio/chemotherapy afforded by a stem cell transplant. However, the clinical experience with high-dose chemotherapy and autologous bone marrow transplantation in lung cancer has been disappointing, with most trials showing little or no improvement in long-term survival. Unfortunately, lung cancer has a tendency to metastasize to the bone marrow, and lung cancer cells are known to circulate in the peripheral blood. Therefore, there is concern that autologous stem cell grafts from lung cancer patients may reinoculate recipients with live tumor cells. Photochemical purging of stem cell grafts with Merocyanine 540 (MC540) is highly effective against a wide range of leukemia and lymphoma cells and is well tolerated by normal hematopoietic stem and progenitor cells. Most solid tumor cells (including lung cancer cells), however, are only moderately sensitive or refractory to MC540-mediated photodynamic therapy (PDT). We report here that postirradiation hyperthermia (< or = 42 degrees C, 3 h) potentiates the MC540-mediated photoinactivation of both wild-type (H69) and cisplatin-resistant mutant (H69/CDDP) small cell lung cancer cells by several orders of magnitude, while only minimally enhancing the depletion of normal human granulocyte/macrophage progenitor cells. Our data suggest that postirradiation hyperthermia provides a simple and effective means of extending the utility of MC540-PDT to the purging of stem cell grafts contaminated with lung cancer and possibly other solid tumor cells. PMID- 11272735 TI - Differences in the response of wheat, soybean and lettuce to reduced blue radiation. AB - Although many fundamental blue light responses have been identified, blue light dose-response curves are not well characterized. We studied the growth and development of soybean, wheat and lettuce plants under high-pressure sodium (HPS) and metal halide (MH) lamps with yellow filters creating five fractions of blue light. The blue light fractions obtained were < 0.1, 2 and 6% under HPS lamps, and 6, 12 and 26% under MH lamps. Studies utilizing both lamp types were done at two photosynthetic photon flux levels, 200 and 500 mumol m-2 s-1 under a 16 h photoperiod. Phytochrome photoequilibria was nearly identical among treatments. The blue light effect on dry mass, stem length, leaf area, specific leaf area and tillering/branching was species dependent. For these parameters, wheat did not respond to blue light, but lettuce was highly sensitive to blue light fraction between 0 and 6% blue. Soybean stem length decreased and leaf area increased up to 6% blue, but total dry mass was unchanged. The blue light fraction determined the stem elongation response in soybean, whereas the absolute amount of blue light determined the stem elongation response in lettuce. The data indicate that lettuce growth and development requires blue light, but soybean and wheat may not. PMID- 11272736 TI - Evidence for yellow light suppression of lettuce growth. AB - Researchers studying plant growth under different lamp types often attribute differences in growth to a blue light response. Lettuce plants were grown in six blue light treatments comprising five blue light fractions (0, 2, 6% from high pressure sodium [HPS] lamps and 6, 12, 26% from metal halide [MH] lamps). Lettuce chlorophyll concentration, dry mass, leaf area and specific leaf area under the HPS and MH 6% blue were significantly different, suggesting wavelengths other than blue and red affected plant growth. Results were reproducible in two replicate studies at each of two photosynthetic photon fluxes, 200 and 500 mumol m-2 s-1. We graphed the data against absolute blue light, phytochrome photoequilibrium, phototropic blue, UV, red:far red, blue:red, blue: far red and 'yellow' light fraction. Only the 'yellow' wavelength range (580-600 nm) explained the differences between the two lamp types. PMID- 11272737 TI - Exciplex-type behavior and partition of 3-substituted indole derivatives in reverse micelles made with benzylhexadecyldimethylammonium chloride, water and benzene. AB - The fluorescence properties of 3-methylindole (MI), 3-indoleacetic acid (IAA), 3 indoleethyltrimethylammonium bromide (IETA), L-tryptophan (Trp) and tryptamine hydrochloride (TA) were studied in reverse micelles solutions made with the cationic surfactant benzylhexadecyldimethylammonium chloride (BHDC) in benzene as a function of the molar ratio water/surfactant R (= [H2O]/[BHDC]). The fluorescence quenching of the model compound MI by benzene in cyclohexane solutions and by BHDC in benzene solutions were also studied in detail. The fluorescence of MI in benzene is characteristic of a charge-transfer exciplex. The exciplex is quenched by the presence of BHDC, due to the interactions of the surfactant ion pairs with the polar exciplex. In reverse micelle solutions at low R values, all the indoles show exciplex-type fluorescence. As R increases, the fluorescence behavior strongly depends on the nature of the indole derivative. The anionic IAA remains anchored to the cationic interface and its fluorescence is quenched upon water addition due to the increases of interface's micropolarity. For IETA, TA and Trp an initial fluorescence quenching is observed at increasing R, but a fluorescence recovery is observed at R > 5, indicating a probe partition between the micellar interface and the water pool. For the neutral MI, the fluorescence changes with R indicate the partition of the probe between the micellar interface and the bulk benzene pseudophase. A simple two site model is proposed for the calculation of the partition constants K as a function of R. In all cases, the calculation showed that even at the highest R value, about 90% of the indole molecules remain associated at the micellar interface. PMID- 11272738 TI - Effects of scopolamine on working memory in rats in a delayed matching-to position task, employing a subject-centered procedure. AB - The effect of scopolamine hydrobromide on a delayed matching-to-position task was examined while controlling for two confounding factors, i.e., mediating behavior and slow performance on a task. The task was given on the basis of a subject centered method in which delay intervals are dependent on subjects' performance. The performance of individual subjects, rather than averaged group performance, was taken as the focus of the analysis. The results indicated that scopolamine had an effect not only on speed of performing the task but also on the length of the retention interval. The effects differed considerably among individuals: the effects on both the length of the retention interval and the speed of performance were found for two of the five subjects. An effect on speed of performance alone was found for one subject. No effects on either measure were found in a further two subjects. PMID- 11272739 TI - The Guilt Inventory. AB - The Guilt Inventory contains subscales measuring trait guilt, state guilt, and moral standards. Previous research has suggested the reliability of these scales and the validity of their interpretations. The items, coding, and scoring procedure for the Guilt Inventory are presented here as well as additional evidence regarding the reliability and measurement characteristics of its subscales. PMID- 11272740 TI - Workplace romance in the public sector: sex differences in reactions to the Clinton-Lewinsky affair. AB - This study examined reactions of part-time MBA students (n = 199) and undergraduate business students (n = 220) to the affair involving U.S. President Bill Clinton and White House intern Monica Lewinsky prior to Clinton's impeachment by the House of Representatives. Consistent with research on attitudes toward workplace romance in the private sector, women believed that this affair occurring in the public sector represented a more serious problem for the nation and more than men were inclined to prefer that some type of action, e.g., resignation or impeachment, be taken. Implications of the results are discussed. PMID- 11272741 TI - Supervisors' tactics of influence and subordinates' tolerance for disagreement. AB - Self-reported scores of 156 subordinates on tolerance for disagreement were associated with their perceptions of supervisors' use of referent, expertise, and reward-based tactics and less use of legitimate and punishment-based tactics of influence. PMID- 11272742 TI - Comparison: attitudes toward homosexuality of international and American college students. AB - Using the Index of Attitudes Toward Homosexuals to study the attitudes of 34 Asian students and 32 American students toward lesbians and gay men showed these Asian students were more likely to harbor homophobic attitudes than these American students. There were no significant sex differences between groups. PMID- 11272743 TI - Validating cluster assignments. AB - In the absence of a statistically based assessment of cluster assignments of observations in terms of their similarity along selected variables the validity of a set of clusters could be developed if clusters withstand both empirical scrutiny and "make sense" to expert informants. PMID- 11272744 TI - Interracial dating: attitudes and experience among American college students in California. AB - Dating and marriage crossing ethnic, racial, and cultural lines have become increasingly common in the United States. This study examined two aspects, interracial dating behavior and attitudes toward romantic involvement, in four ethnic groups of college students: Euro-American, Latino, Asian-American, and African-American. Subjects (196 men, 367 women) were surveyed with regard to their willingness to be romantically involved interracially or interculturally along with their actual interracial dating experience. Analysis indicated a high willingness in all ethnic groups to be romantically involved as well as an absence of sex difference with regard to both attitude and experience. However, there were differences in both attitude and experience among ethnic groups. PMID- 11272745 TI - Knowledge and attitudes about HIV/AIDS of a sample of school teachers in South Africa. AB - A questionnaire was administered to 160 black school teachers randomly chosen from one rural region of the Northern Province of South Africa. Their ages ranged from 26 to 57 years. Scores indicated very poor general knowledge about transmission of HIV/AIDS and moderately high supportive attitudes about dealing with HIV inside and outside of the classroom. Pearson product-moment correlations of .3 and .6 suggested weak association of knowledge about transmission and general knowledge with a supportive attitude. PMID- 11272746 TI - An unusual birthmark case thought to be linked to a person who had previously died. AB - The following case report describes a Burmese subject with an unusual birthmark and birth defects thought by local people to be linked to events surrounding the death of his mother's first husband. The nature of the link is explored, including how the assumption of a linkage could have led to subsequent events. PMID- 11272747 TI - Self-disclosure and occupational stress in Chinese professionals. AB - The relationship of self-disclosure with occupational stress and symptoms of stress was examined among 243 Hong Kong Chinese adult professionals. Self disclosure was negatively correlated with both occupational stress and psychological symptoms of stress for disclosures of personal feelings and social relationships when disclosing to a Best Friend, indicating a stress-buffering effect. There was a positive correlation between occupational stress and psychological symptoms of stress for disclosure of personal feelings and information about social relationships when disclosing to Mother. While both sexes reported similar occupational stress, the total self-disclosure of women was higher than for men, and this was especially true for intimate topics. The implications of the results were discussed. PMID- 11272748 TI - Psychological empowerment as a criterion for adjustment to a new job. AB - Interview data from 120 professionals and managers, collected by telephone after they experienced a job change, were matched with personality test scores from an employment testing center. Four dimensions of psychological empowerment (self determination, meaning, competence, and impact) were tested as criteria, and four personality traits (achievement, endurance, locus of control, and self-esteem), measured prior to the job change, were used as predictors. In a multiple regression analysis the personality traits accounted for 26% of the variance in overall empowerment, and locus of control emerged as an important antecedent. Additional analyses pointed to perceived managerial support, sex, and rank as possible moderators. Ideas for further research and theoretical extension are discussed. PMID- 11272749 TI - Personality correlates of alienation in a university sample. AB - In a study of alienation among urban university students, 85 men and 136 women completed the Gould Manifest Alienation Measure and the 44-item Big Five Inventory. Multivariate analysis of variance disclosed that the 77 college students high on Alienation, irrespective of sex, scored higher on the Neuroticism scale and lower on the Conscientiousness and Openness scales of the Big Five Inventory. None of the interactions between sex and scores on the Big Five Inventory was significant. Results suggest that college students scoring high on alienation can survive in a supportive university environment, although they appear to experience increased anxiety and tension and have a relatively constricted awareness of their environments. Students scoring high on alienation may also be more tolerant of deviant behavior given their perceived irrelevance of social norms. PMID- 11272750 TI - A Monte Carlo investigation of the Fisher Z transformation for normal and nonnormal distributions. AB - The Fisher transformation of the sample correlation coefficient r (1915, 1921) and two related techniques by Gayen (1951) and Jeyaratnam (1992) are examined for robustness to nonnormality. Monte Carlo analyses compare combinations of sample sizes and population parameters for seven bivariate distributions. The Fisher, Gayen, and Jeyaratnam approaches are shown to provide useful results for a bivariate normal distribution with any population correlation coefficient rho and for nonnormal bivariate distributions when rho = 0. In contrast, the techniques are virtually useless for nonnormal bivariate distributions when rho not equal to 0.0. Surprisingly, small samples are found to provide better estimates than large samples for skewed and symmetric heavy-tailed bivariate distributions. PMID- 11272751 TI - Personality characteristics and coping styles of women working in and in training for nontraditional blue collar jobs. AB - 43 women employed in nontraditional blue collar jobs were compared with 27 women in training for such jobs on a number of variables linked to job success and satisfaction. The purpose of the study was to determine whether women enter blue collar trades with characteristics that predispose them to successful work-role transitions or whether they adopt different coping styles and behavioral characteristics over time in order to fit into their work environments. Questionnaire packets that contained the Self-efficacy Scale, the Personal Assertion Analysis, the Ways of Coping Scale-Revised, and the Bem Sex Role Inventory were completed by 70 women. Participants in training were more androgynous and higher than expected on both problem-focused and emotion-focused coping. Possible explanations and suggestions for research are discussed. PMID- 11272752 TI - Psychology of the scientist: LXXXIII. An assessment of Herek's critique of the Cameron group's survey studies. AB - Herek's criticisms of the research on sexuality by Paul Cameron's research group are evaluated. While the Cameron group's research has many limitations, these limitations are not uncommon in contemporary research, especially research that concerns specialized, hard-to-find populations. The best response from a scientific perspective, it is argued, is better research, not merely critical comments on existing research. PMID- 11272753 TI - Examination of factor structure for the consumers' responses to the Value Consciousness Scale. AB - The psychometric properties of the Value Consciousness Scale developed by Lichtenstein, Netemeyer, and Burton in 1990 were examined in a retail grocery study (N = 497). Original assessment of scale properties was undertaken using two convenience samples in a nonretail setting and additional scale performance has been documented by the scale authors. This study furthers previous research by (1) examining performance on the items in the retail grocery setting and (2) utilizing an appropriately rigorous sampling procedure. A confirmatory factor analysis indicated that the Value Consciousness Scale does not exhibit unidimensional properties, and one must be cautious if this scale is used in applications of market segmentation until further clarification can be provided. PMID- 11272754 TI - Revision of texts with word-processing. AB - The aim of the research was to study the use of a word processor to improve writing and revising skill by 28 pairs of primary school children. Analysis confirmed greater difficulty in the drafting of texts than with traditional handwriting, but the increase in the number of words and errors does not imply improvement in lexical and orthographical aspects. The guided revision of texts on a word processor may support beginning writers' expression and, with increasing ease, allow them to use greater precision in the process of revision. PMID- 11272755 TI - Association of need for cognition with judgments of height, weight, and body fat covariation. AB - 559 college students, assessed for Need for Cognition, judged whether height, weight, and body fat were correlated using judgment probes that controlled for framing and conditional format. A principal components analysis of Need for Cognition scores identified two factors underlying the scale, which may be important in judgment outcome. In addition, judgments about correlations among height, weight, and body fat were similar to those of previous studies. Furthermore, the hypothesis that Need for Cognition would be related to the tendency to judge a negative correlation between height and body fat, i.e., a possible illusory correlation, was confirmed. Results are discussed relative to the Elaboration Likelihood Model of Persuasion. PMID- 11272756 TI - Motivations for use of opiates among addicts seeking treatment in Shiraz. AB - This study assessed the characteristics and motivation for substance use among addicts referred to the Shiraz Self-identified Center, an out-patient treatment facility. Data were gathered by a semistructured interview from 306 consecutive addicts seeking treatment and referred from July to September, 1998. Their mean age was 37 yr., and the majority (73.9%) were married. Of these addicts, 28.4% were workers, 13.4% drivers, and 11.4% were unemployed. Modeling or social pressure (43.1%) was identified as the first and enjoyment (fun) was the second most common reason given for opiate use. The majority (97.1%) used opium and 71.9% used alcohol; however, only 2.6% reported current use of alcohol. Other subjects were current users of cigarettes (72.2%), opium (67%), heroin (35%), hashish (2%), hallucinogens (0.3%), and cocaine (0.3%). The most common reason given for currently using opiates was habit (56.5%). About 36% of the subjects reported that they had frequently used opiates for more than a decade. These findings are quite different from those carried out in the West, although there is some overlap. Cultural attitudes toward drug use likely affect the types and amount of use. PMID- 11272757 TI - Expression of unfavorable emotions in Japanese college students with alexithymic characteristics. AB - We examined the association of alexithymic characteristics and the expression of unfavorable feelings such as anger and hostility in a sample of 489 Japanese college students. Analysis suggested that alexithymic college students are prone to indicate not only emotional instability but also cynical hostility and anger. On the contrary, alexithymic college students indicated significantly higher scores on Anger-in and Anger Control, which may be related to Japanese sociocultural aspects. In particular, Anger Control was stronger in the men who were alexithymic, suggesting that they may unconsciously struggle for the suppression or control of unfavorable feelings such as anger and hostility to stabilize their inner emotions. PMID- 11272758 TI - Creating comparability among reliability coefficients: the case of Cronbach alpha and Cohen kappa. AB - Cronbach alpha and Cohen kappa were compared and found to differ along two major facets. A fourfold classification system based on these facets clarifies the double contrast and produces a common metric allowing direct comparability. A new estimator, coefficient beta, is introduced in the process and is presented as a complement to coefficient alpha in estimating the psychometric properties of test scores and ratings. PMID- 11272759 TI - Who buys their textbooks online? AB - A pilot study of 82 undergraduates indicated that skills with computers and Internet use predicted purchasing textbooks online. PMID- 11272761 TI - "Prefrontal deficits" discriminate young offenders from age-matched cohorts: juvenile delinquency as an expected feature of the normal distribution of prefrontal cerebral development. AB - A total of 36 male adolescent delinquents and 19 age-matched students from a male secondary school were administered several different performance-based tests. Although estimates of intelligence did not differ significantly between the groups, the incarcerated delinquents displayed more impulsivity, lower conceptual level, less conceptual flexibility, and poorer critical thinking than the reference group. The linear combination of only three variables, critical thinking, conceptual level, and numbers of errors during a conditioned spatial association task, correctly classified 89% of the 55 subjects. The results were considered consistent with the hypothesis that delayed or different development of complex functions associated with the left and (particularly) the right prefrontal cortices and their limbic inputs may be responsible for antisocial behavior. This variation will always be present due to the statistical variations in ontogeny and less than optimal parental structure within a consistent but small proportion of any generational cohort. PMID- 11272760 TI - Ego Function Assessment of substance abusers: standardization and reliability. AB - This study examined the reliability of Ego Function Assessment in a self-report inventory which yields 12 function measures. 89 substance abusers in an urban treatment facility completed the inventory; 81 completed the inventory again after 2 wk. Test protocols of the first administration were assessed for interitem consistency utilizing a hierarchical algorithm which revised scales to yield optimal alphas. Retest scores were used to evaluate the stability of the 12 scales over 2 wk. Standardization, reliability, and stability data are presented for the original and revised scales. The possibility of response sets in the items are explored, and data patterns are discussed in terms of state vs trait issues in ego functioning. PMID- 11272762 TI - Obligation conditionals in a nonstandard conditional selection task: general versus specific reasoning strategies as a false dichotomy. AB - The paper reports a study comparing performance on obligation and causal conditionals in the explicit standard order conditional selection task. Analysis indicated that both general and specific reasoning strategies are used when a request is given to disprove obligation conditionals, contradicting the prevalent view that only one of the two kinds of strategy is used and confirming the 1993 suggestions of Evans and later ones by Evans and Over. The incidence of general reasoning strategies in this situation is reduced, compared to that for other kinds of conditional in a disproof task probably because a specific reasoning strategy is available that is easier to use. PMID- 11272763 TI - Evaluation of a distance education course in students' decision-making and beliefs about careers. AB - To assess the influence of a distance course on careers in students' career decision-making and beliefs, 178 distance education college students in the Open University in Taiwan completed the Career Decision Scale, the Career Beliefs Checklist, and a demographic sheet. Unlike most prior research, the course was not associated with career decision-making and beliefs of the students, but students mentioned that career beliefs influenced their decision-making. Three way multivariate analysis of covariance showed a main effect for age groups on career indecision. On a follow-up questionnaire to which 143 of the same subjects responded, the students indicated their concern about specific factors in career decision-making. The research findings have important implications for the redesign of the career education course for adults and for research. PMID- 11272766 TI - Medicine in your palm. PMID- 11272765 TI - The Internet. Part II: A simplified guide on how to use. PMID- 11272764 TI - Limitations in ethnic research on the MMPI-A. AB - This article describes two common limitations in research on responses of ethnic groups on the MMPI-A, lack of external correlates and combining scale scores for males and females. PMID- 11272768 TI - What a mouse can do for a medical practice. PMID- 11272767 TI - Health care and new technology. PMID- 11272769 TI - How do you handle special therapeutic requests? PMID- 11272771 TI - "Me first!"--my biggest mistake. PMID- 11272770 TI - The Practice Administrators Committee of MCMS. AB - Beginning with this issue, Maryland Medicine will occasionally include a column provided by members of the Practice Administrator's Committee of the Montgomery County Medical Society. The editorial board believes their viewpoint may be helpful to physicians in the day-to-day business issues they confront. PMID- 11272772 TI - The bombing of Avranches. PMID- 11272773 TI - Cyberspeak. PMID- 11272774 TI - The "god" on my desk. PMID- 11272775 TI - The Internet. Part One: A guide to history. PMID- 11272776 TI - Effective local treatment for severe ocular inflammatory disease: toward the Holy Grail. PMID- 11272777 TI - Management of diabetic retinopathy by general practitioners in Victoria. AB - PURPOSE: To compare the self-reported management of diabetic retinopathy by general practitioners to the National Health and Medical Research Council of Australia (NHMRC) Guidelines for the Management of Diabetic Retinopathy. METHODS: In 1994, a stratified (by urban/rural practice location) sample of 500 general practitioners in Victoria was surveyed in regard to their management of diabetic retinopathy. Following the release of the NHMRC Guidelines for the Management of Diabetic Retinopathy in 1997, these same general practitioners were sent a two page questionnaire related to their management of diabetic retinopathy. RESULTS: Completed questionnaires were received from 228 general practitioners (59% of original participants). Only 37% (79/216) of the general practitioners reported that they had received a copy of the guidelines. Of the general practitioners who had received the guidelines, 18% (14/79) said that they had not read them at all, while 65% (51/79) had read them partially and 18% (14/79) had read them in their entirety. At follow up, less than half (98/214) of general practitioners reported examining 50% or more of their patients for diabetic retinopathy, compared with 104/214 at baseline. General practitioners who had read the guidelines were more likely to report that not being sure what to do when changes were detected was a minor barrier or was not a barrier to them performing dilated ophthalmoscopy (93% vs 83%, chi2(1) = 3.67, P = 0.055). Nearly all of the general practitioners reported that they refer their patients with diabetes to an ophthalmologist or optometrist at least every 2 years as recommended. Seventy-six per cent (170/224) of the general practitioners felt that 70% or more of their patients complied with their instructions to visit an ophthalmologist or optometrist. CONCLUSION: The NHMRC guidelines for diabetic retinopathy appear to have had a positive effect on some of the attitudes of general practitioners who have read them, but more effort is needed to disseminate the guidelines to all general practitioners and to increase their uptake. PMID- 11272778 TI - Contrast sensitivity following focal laser photocoagulation in clinically significant macular oedema due to diabetic retinopathy. AB - PURPOSE: To evaluate the influence of focal aser photocoagulation on contrast sensitivity in diabetic patients with clinically significant macular oedema (CSMO). METHODS: A prospective non-comparative interventional study was performed on a group of patients with CSMO at Dr Rajendra Prasad Centre for Ophthalmic Sciences, New Delhi, a tertiary eye care centre. Thirteen diabetc patients (14 eyes) with CSMO and no history of prior photocoagulation were recruited for this study. Direct focal photocoagulation of all leaking microaneurysms was performed using an argon green laser (514 nm). A contact lens was used as a slit lamp delivery system. Evaluation of the best corrected Snellen visual acuity, contrast senstivity, slit lamp biomicroscopy, macular status on direct ophthalmoscopy and fluorescein angiography was carried out 1 month and 3 months after laser photocoagulation. RESULTS: Following direct focal laser photocoagulation, focal CSMO resolved completely in all but one eye, 4-8 weeks later, as seen on slit lamp biomicroscopy and/or fluorescein angiography. Post-treatment, visual acuity remained stationary in eight eyes, improved by one line in three eyes, by two lines in two eyes and by three lines in one eye. The mean +/- SD pretreatment and post-treatment decimal visual acuities were 0.49+/-0.30 and 0.59+/-0.28, respectively. The mean +/- SD pre-laser contrast sensitivity score was 121.3+/ 83.6, which increased significantly to a mean +/- SD of 151.6+/-80.5 fo lowing direct focal photocoagulation. CONCLUSION: Focal argon laser photocoagulation in CSMO in diabetics helps in improving the contrast sensitivity and stabilizes the visual acuity. The changes in contrast sensitivity and visual acuity are independent of each other. PMID- 11272779 TI - Safety and efficacy of intravitreal triamcinolone for cystoid macular oedema in uveitis. AB - PURPOSE: To report the safety and efficacy of intravitreal triamcinolone in the treatment of inflammatory cystoid macular oedema (CMO) in six patients who were resistant to other forms of therapy. METHODS: An open-label unmasked prospective nonrandomized pilot study of six patients with idiopathic uveitis and visually significant macular oedema, resistant to periocular and/or systemic corticosteroid treatment, was carried out. Baseline examination and investigations were performed, including fundus fluorescein angiography, and the patients were given a single intravitreal injection of triamcinolone (4 mg/0.1 mL). The primary outcome measure was angiographic resolution of CMO. Patients were reviewed at intervals of 2-4 weeks for 12 months. RESULTS: A single intravitreal injection of triamcinolone induced clinical and angiographic resolution of inflammatory macular oedema in all patients for varying periods of time up to 6 months. Five patients experienced increased intraocular pressure to 30 mmHg or greater which required treatment. Two patients developed posterior subcapsular cataract. CONCLUSION: One injection of intravitreal triamcinolone was an effective short-term treatment for resistant CMO in uveitis. As with steroids given by other routes, raised intraocular pressure and cataract may occur. As it was so effective in these eyes with resistant CMO, a larger study is warranted to evaluate this form of therapy. PMID- 11272780 TI - Indocyanine green anterior segment angiography for studying conjunctival vascular changes after trabeculectomy. AB - BACKGROUND: The aim of the study was to evaluate the use of indocyanine green (ICG) for angiography of the anterior segment to characterize conjunctival and episcleral vasculature changes after trabeculectomy. METHODS: This was a prospective evaluation of anterior segment ICG angiography in 10 eyes of 10 patients undergoing trabeculectomy for the first time. Trabeculectomy was performed with intraoperative sponge application of 5-fluorouracil (5 cases) or mitomycin C (5 cases). Anterior segment ICG angiography was performed prior to surgery, then at 2 weeks and 2 months after surgery. RESULTS: With ICG, the anterior segment vessels were well delineated, including deep episcleral veins, which have not been clearly shown in previous angiographic techniques. Late phases of the angiogram could also be studied. The vascular alterations after trabeculectomy noted included oss of vascularity over the bleb area and vascular anastomoses along the perimeter of the avascular bleb. CONCLUSIONS: Angiography using ICG has potential as an investigative tool to study the conjunctival and episcleral vasculature changes after trabeculectomy. PMID- 11272781 TI - Dacryocystorhinostomy for epiphora in the presence of a patent lacrimal system. AB - PURPOSE: To evaluate the success rate of dacryocystorhinostomy (DCR) for epiphora in patients with a clinically patent lacrmal drainage system. METHODS: A series of 51 patients, 12 men and 39 women, who underwent DCR surgery for epiphora with a clinically patent lacrimal drainage system in the background of normal eyelid examination, were evaluated. All patients underwent fluorescein dye disappearance testing and Jones 1 and 2 testing with dacryocystograms in borderline cases. A standard DCR was performed with bicanalicular silicone tubing inserted in all cases. The average time for the removal of the silicone tubes was 9.6 weeks. RESULTS: In 48 cases (94%) there was improvement in symptoms with minimal or no significant epiphora postoperatively. CONCLUSIONS: Epiphora with a patent lacrimal drainage system obstruction can be successfully treated by DCR based on fluorescein dye disappearance tests and Jones 1 and 2 tests, with dacryocystography in borderline cases. PMID- 11272783 TI - Transforming growth factor-beta1 induces alpha-smooth muscle actin expression and fibronectin synthesis in cultured human retinal pigment epithelial cells. AB - BACKGROUND: Proliferative vitreoretinopathy is a serious complication of retinal detachment, yet its pathogenesis is not fully understood. Retinal pigment epithelial cells and glial cells are found in the fibrous membranes in proliferative vitreoretinopathy. Many cytokines are involved in the pathology. Transforming growth factor (TGF)-beta, a cytokine found in serum, has been shown to be an important factor regulating the synthesis of fibrous extracellular matrix in proliferative vitreoretinopathy. METHODS: Cultured human retinal pigment epithelial cells were used in the experiments. The effects of TGF-beta1 on phenotype and function in retinal pigment epithelial cells were recorded as changes in the expression of alpha-smooth muscle actin and fibronectin synthesis using immunohistochemistry and enzyme-linked immunosorbent assay, respectively. RESULTS: TGF-beta1 induced the expression of alpha-smooth muscle actin (P < 0.0001, n = 3), and significantly increased the synthesis of fibronectin by cultured human retinal pigment epithelial cells (P < 0.01, n = 4). CONCLUSIONS: Elevated levels of TGF-beta1 in proliferative vitreoretinopathy may contribute to phenotype changes in retinal pigment epithelial cells leading to matrix deposition and contraction. Since the elevated levels of TGF-beta1 may emanate from a number of diverse sources in proliferative vitreoretinopathy, developing an antagonist to TGF-beta1 may offer an approach to the treatment of proliferative vitreoretinopathy. PMID- 11272782 TI - Comparison of subretinal fluid levels of two 0.3% ciprofloxacin-containing eye drops. AB - PURPOSE: Two ophthalmic solutions of 0.3% ciprofloxacin eye drops are available in Turkey: Ciloxan and Siprogut. A previous study by the same authors was the first to report vitreous penetration of ciprofloxacin-containing eye drops. The aim of the present study was to compare the levels of drug found in the subretinal fluid by the two products following local administration. METHODS: Forty-three patients undergoing conventional retinal detachment surgery received either Ciloxan (22 patients) or Siprogut (21 patients). Beginning 6 h before surgery, two drops of solution were instilled onto the operative eye every 30 min for the first 3 h and then hourly for the next 3 h. Subretinal fluid samples were collected 30 min after administration of the last dose and were assayed for ciprofloxacin levels using a method involving high-performance liquid chromatography with fluorometric detection. RESULTS: The minimum and maximum subretinal fluid concentrations measured were 0.11 microg/mL and 0.65 microg/mL, respectively, with Ciloxan, and 0.08 microg/mL and 0.62 microg/mL, respectively wth Siprogut. There was no statistical difference between the subretinal fluid ciprofloxacin levels of the two products. The subretinal fluid drug evels attained by both products were below the minimum inhibitory concentrations of common ocular pathogens. CONCLUSIONS: Ciloxan and Siprogut can penetrate subretinal fluid. The ocular bioavailability of ciprofloxacin after local administration is equivalent for both pharmaceutical products. PMID- 11272784 TI - Orbital involvement in cherubism. AB - Cherubism is a rare, inherited condition characterized by fibro-osseous lesions of the maxilla and mandible. It has recently been localized to chromosome 4p16.3. The fullness of the lower half of the face and retraction of the lower lids gives the characteristic 'eyes raised to heaven' cherubic appearance. A case report of a 7-year-old girl with extensive orbital involvement of cherubism is presented. The patient underwent multidisciplinary surgery for the bony lesions, which extended from the maxillary antrum into adjacent structures, including extensive extraperiosteal orbital involvement. Cherubism may have orbital manifestations including lower lid retraction, proptosis, diplopia, globe displacement and visual loss due to optic atrophy. Ophthalmologists should be aware of the syndrome, its ophthalmic features and overall management. PMID- 11272785 TI - Ockham's glaucoma. AB - The combination of characteristic optic nerve head cupping, arcuate visual field loss and ocular hypertension would usually be thought sufficient to diagnose glaucoma. Only in the absence of elevated intraocular pressure, when normal tension glaucoma may be suspected, would intracranial imaging normally be performed to exclude occult pathology. A case is presented which illustrates the continuing need for vigilance, and an open mind, years after an apparently straight-forward diagnosis has been made. PMID- 11272786 TI - Carotid artery ectasia coexistent with primary open angle glaucoma. AB - A 60-year-old smoker presented with high intraocular pressure in the right eye with a right afferent pupil defect and visual field suggestive of primary open angle glaucoma in the right eye only, when an examination 2 years earlier had revealed no hint of ocular pathology. Radiological investigations demonstrated prominent ectasia of the internal carotid arteries extending into the proximal middle cerebral arteries. The changes in the carotids extended throughout the cavernous sinus regions, encroached on the under surface of the optic chiasm and were closely related to the internal aspects of both optic canals. In primary open angle glaucoma management, neural imaging is not normally recommended; however, neural imaging investigations should be considered if the presentation is not typical of a chronic bilateral optic neuropathy PMID- 11272787 TI - Trichiasis in Aboriginal people of the Kimberley region of Western Australia. AB - BACKGROUND: Trachoma is a significant public health problem in the Kimberley region of Western Australia. The prevalence of follicular trachoma in the region has been well documented and control of active trachoma is well established. In contrast, trichiasis prevalence in recent years is less well documented. This study aimed to determine trichiasis prevalence in the Kimberley population and to document an appropriate intervention. METHODS: A collaborative programme was established involving the Kimberley Public Health Unit, Kimberley Aged Care Services and the visiting ophthalmology service. After training, medical students screened the aged-care population for trichiasis and the aged-care services staff were educated about identification and referral procedures for patients with trichiasis. The Kimberley Public Health Unit was responsible for ensuring specialist assessment, and correction, of trichiasis and appropriate post treatment follow up. RESULTS: A total of 597 Kimberley Aboriginal people aged 50 years or more were screened for trichiasis, representing 42% of that age group. Trichiasis was suspected in 40 people. Of the 28 people with suspected trichiasis who underwent ophthalmological assessment, 17 were found to have trachomatous trichiasis. The observed prevalence of trachomatous trichiasis was 2.8%. The trichiasis screening programme has been adopted by most aged-care organizations in the Kimberley. CONCLUSIONS: The results show that trichiasis continues to affect elderly Aboriginal people, especially those from Halls Creek Shire. Health professionals and aged-care workers should be alert to the possibility of this preventable cause of blindness and work collaboratively to ensure that patients receive timely and appropriate treatment. PMID- 11272788 TI - Meiosis in primary trisomics of rye: considerations for models of chromosome pairing. AB - Meiotic chromosome pairing of primary trisomics of rye was analysed by electron microscopy in surface-spread prophase I nuclei and compared with light microscopic observations of metaphase I cells. Despite the large-sized chromosomes of rye, prophase I trivalent frequencies were close to the two thirds expected on a simple model with two terminal independent pairing initiation sites per trisome (set of three homologous chromosomes). Direct observations mostly reveal one pairing partner switch (PPS) in prophase I trivalents, which confirms this supposition. There were no significant differences between the number of trivalent and bivalent plus univalent configurations observed at prophase and metaphase I; therefore, synapsed segments form chiasmata. In all of the trisomics, the three homologues showed variations not only in the number of telomeric C-bands but also in the amount of heterochromatin of these bands, which allowed identification of chromosomes or chromosome arms associated in most metaphase I configurations. In trisomics for chromosomes 2, 3 and 5, some metaphase I chromosome configuration frequencies did not fit those expected under the assumption of random chromosome association among all partners, suggesting the existence of preferences for pairing between two given chromosome arms of the trisome. No preferential associations either at metaphase I or pachytene were observed in the trisomics for chromosome 6. The fit between theoretical pairing models and the experimental data is also discussed. PMID- 11272789 TI - Karyotype comparison and phylogenetic relationships of Pipistrellus-like bats (Vespertilionidae; Chiroptera; Mammalia). AB - Detailed karyotype descriptions of 20 Pipistrellus-like bat species belonging to the family Vespertilionidae are presented. For the first time, chromosomal complements of four species, i.e. Pipistrellus stenopterus (2n = 32), P. javanicus (2n = 34), Hypsugo eisentrauti (2n = 42) and H. crassulus (2n = 30) are reported. A Pipistrellus kuhlii-like species from Madagascar represents a separate species distinguished from the European Pipistrellus kuhlii (2n = 44) by a diploid chromosome number of 42. Banded karyotypes are presented for the first time for Scotozous dormeri, Hypsugo capensis, Hesperoptenus blanfordi, Tylonycteris pachypus and robustula. Chromosomal evolution in the family Vespertilionidae is characterized by the conservation of entire chromosomal arms and reductions in diploid chromosome number via Robertsonian fusions. Less frequently, centric fissions, para- and pericentric inversions and centromere shifts were found to have occurred. In several cases a certain type of chromosomal change predominates in a karyotype. Examples of this are the acquisition of interstitial heterochromatic bands in Tylonycteris robustula, and centric shifts in P. javanicus, H. eisentrauti and Hesp. blanfordi. The species examined here belong to three tribes, i.e. Pipistrellini, Vespertilionini and Eptesicini, which are distinguished by chromosomal characteristics. According to our results, the species Pipistrellus (Neoromicia) capensis belongs to the Vespertilionini and not to the Pipistrellini. We therefore propose to elevate the subgenus Neoromicia to generic rank. PMID- 11272790 TI - Cytogenetics of nine species of mediterranean blennies and additional evidence for an unusual multiple sex-chromosome system in Parablennius tentacularis (Perciformes, Blenniidae). AB - The chromosomal complements of nine species of Blenniidae (Aidablennius sphylnx, Blennius ocellaris, Lypophris adriaticus, L. pavo, L. trigloides, Parcablennius gattorugine, P. ponticus, P. sanguinolentus, P. tentacularis) from the Adriatic Sea were analysed with several banding methods and in-situ hybridization. In all species, the diploid set consists of 48 mostly acrocentric chromosomes and has a similar location (terminal centromeric) of NORs, except for L. pavo (interstitial pericentric) and P. ponticus (terminal on the long arm). There are major differences in karyotype with regard to the amount and distribution of heterochromatin. Parablennius tentacularis shows a distinctive sex-chromosome system involving 2n = 48 males with a large totally heterochromatic Y chromosome, and males with 2n = 47. This difference is likely to be the consequence of a translocation of an autosome on the original Y. This finding constitutes an additional instance of the great variability in origins of multiple sex chromosome systems in vertebrates. PMID- 11272791 TI - Chromosomal location and evolution of a satellite DNA family in seven sturgeon species. AB - The Hind III satellite DNA family, isolated from the Acipenser naccarii genome, was used as a probe for fluorescent in-situ hybridization (FISH) on the karyotype of seven sturgeon species, six belonging to the genus Acipenser and one to Huso. All species except one (A. sturio) exhibit from 8 to 80 chromosome hybridization signals, mainly localized at the pericentromeric regions. Eight chromosomes with weak hybridization signals are present in H. huso and A. ruthenus, which are characterized by a karyotype with about 120 chromosomes. The species with 240-260 chromosomes, A. transmontanus, A. naccarii, A. gueldenstaedtii, and A. baerii, show from 50 to 80 signals, prevalently localized around centromeres. Moreover, A. transmontanus and A. gueldenstaedtii show from 4 to 8 chromosomes with a double signal. The phylogenetic and evolutionary relationships among sturgeon species are discussed on the basis of number and morphology of signal-bearing chromosomes and on the localization of signals. PMID- 11272792 TI - Mapping of 13 horse genes by fluorescence in-situ hybridization (FISH) and somatic cell hybrid analysis. AB - We report fluorescence in-situ hybridization (FISH) and somatic cell hybrid mapping data for 13 different horse genes (ANP, CD2, CLU, CRISP3, CYP17, FGG, IL1RN, IL10, MMP13, PRM1, PTGS2, TNFA and TP53). Primers for PCR amplification of intronic or untranslated regions were designed from horse-specific DNA or mRNA sequences in GenBank. Two different horse bacterial artificial chromosome (BAC) libraries were screened with PCR for clones containing these 13 Type I loci, nine of which were found in the libraries. BAC clones were used as probes in dual colour FISH to confirm their precise chromosomal origin. The remaining four genes were mapped in a somatic cell hybrid panel. All chromosomal assignments except one were in agreement with human-horse ZOO-FISH data and revealed new and more detailed information on the equine comparative map. CLU was mapped by synteny to ECA2 while human-horse ZOO-FISH data predicted that CLU would be located on ECA9. The assignment of IL1RN permitted analysis of gene order conservation between HSA2 and ECA15, which identified that an event of inversion had occurred during the evolution of these two homologous chromosomes. PMID- 11272793 TI - Conservation of the rat X chromosome gene order in rodent species. AB - We constructed the comparative cytogenetic maps of X chromosomes in three rodent species, Indian spiny mouse (Mus platythrix), Syrian hamster and Chinese hamster, using 26 mouse cDNA clones. Twenty-six, 22 and 22 out of the 26 genes, which were mapped to human, mouse and rat X chromosomes in our previous study, were newly localized to X chromosomes of Indian spiny mouse, and Syrian and Chinese hamsters, respectively. The order of the genes aligned on the long arm of human X chromosome was highly conserved in rat and the three rodent species except mouse. The present results suggest a possibility that the rat X chromosome retains the ancestral form of the rodent X chromosomes. PMID- 11272795 TI - Two-color fluorescence labeling of early and mid-to-late replicating chromatin in living cells. PMID- 11272794 TI - Chromosome regions enriched in hyperacetylated histone H4 are preferred sites for endonuclease- and radiation-induced breakpoints. AB - Previously, we have shown that breakpoints induced by the endonucleases AluI, BamHI and DNase I in CHO chromosomes are localized mainly in G-light bands. Neutrons and gamma rays produced similar breakpoint clusters to endonucleases in most CHO chromosomes. Here we compare endonuclease- and radiation-induced breakpoint maps with hyperacetylation patterns of histone H4. The H4 acetylation pattern in chromosomes is similar to the pattern of G-light, or R-bands, and breakpoints are clustered in highly acetylated chromosome regions. These findings indicate that chromosomal aberrations occur more frequently in active than in inactive chromatin. PMID- 11272796 TI - The epidemiology of TT virus (TTV) infection in a hepatitis C and B virus hyperendemic area of southern Taiwan. AB - TT virus (TTV) is a newly isolated DNA virus from the serum of a patient with posttransfusion hepatitis of unknown etiology in 1997. To evaluate the clinical and molecular characteristics of TT virus (TTV) in a hepatitis C virus (HCV) and B (HBV) hyperendemic area (Masago), 200 residents were enrolled in the study. The sera were tested for alanine aminotransferase (ALT), HCV RNA and GB virus C/Hepatitis G virus (HGV) RNA, TTV DNA, HBsAg, anti-HCV and antibodies to HGV E2 protein (anti-E2). TTV DNA was positive in 99 of the 200 sera with a prevalence rate of 49.5%. The prevalence of HBsAg, anti-HCV, HCV RNA, HGV RNA, anti-E2 and HGV exposure (defined as positive for serum HGV RNA and/or anti-E2) was 38.9%, 69.5%, 64.5%, 17.0%, 25.5% and 39.5%, respectively. Neither clinical nor virological factors were associated with TTV viremia. The rate of ALT abnormality was significantly elevated in HCV RNA-positive (34.9%) than -negative (7.0%) residents (p < 0.001). HCV viremia was the only factor significantly associated with ALT elevation by multiple logistic regression (odds ratio: 6.96; 95% C.I.: 2.60-18.7). We concluded that in this HCV/HBV hyperendemic area, the prevalence of TTV DNA was high. No significant clinical factor was observed to be associated with TTV infection. TTV infection is not related to abnormal ALT levels and ALT abnormality was mainly attributable to HCV but not TTV, HBV or HGV infection. PMID- 11272797 TI - Endoscopic third ventriculostomy in the management of obstructive hydrocephalus caused by aqueductal stenosis. AB - Long-term extracranial shunting for hydrocephalus has numerous drawbacks related to shunt malfunction and infection. The outcomes have been very disappointing in some cases. We have treated twenty one patients with obstructive hydrocephalus, without mortality or morbidity, using a flexible endocope to perform third ventriculostomy. Favorable outcomes were achieved in 95.2% of the cases. Our results are superior to those previously reported. Most of the patients remained shunt independent after treatment and had obtained long-term stabilization. Flexible endoneurosurgical management is simple and safe, and it allows in situ observation and performance of biopsies. Therefore, in patients with obstructive hydrocephalus due to aqueductal stenosis, endoscopic third ventriculostomy should be seriously considered as the primary surgical management. PMID- 11272799 TI - The effect of electroacupuncture on shoulder subluxation for stroke patients. AB - In this study, we evaluated the effect of electroacupuncture on shoulder subluxation for stroke patients. Twenty hemiplegic patients with shoulder subluxation were randomly and equally divided into two groups. The subjects in the control group received conventional therapy, and the subjects in the study group were treated with electroacupuncture and conventional therapy for four weeks. The visual analog scale (VAS) for shoulder pain, motor function status, anthropometry, and X-ray assessment were used to evaluate the status of shoulder subluxation before and after treatment. The results indicated that the pain scores decreased in the study groups significantly more than those in the control group. The degrees of shoulder reduction, including the measurement of anthropometry and X-ray assessment in the study group, were more than those of the control group. However, the motor function status showed no significant difference between two groups. It is concluded that electroacupuncture can be an effective adjuvant management in the treatment of shoulder subluxation for stroke patients. PMID- 11272798 TI - Knee isokinetic strength and body fat analysis in university students. AB - Many factors such as anthropometric variables influence strength performance. This study is to determine the relationship between knee isokinetic strength and body composition, and to compare the gender differences. Test-retest reliability had been performed within one week for all measurement methods before the formal study. Fifty-eight 20-25 year-old university students, 32 females and 26 males, participated in this study. Isokinetic strength of the knee flexion and extension was measured at two angular velocities of 60 degrees/sec and 120 degrees/sec. Body composition was measured by bioelectrical impedance analysis (BIA) and skinfold caliper. The others variables including height, body weight, body mass index (BMI), and waist to hip ratio were measured or calculated. The results showed that the intra-class correlation coefficients for isokinetic knee strength were between 0.83 and 0.93, and body composition and anthropometric variables were between 0.83 and 0.98. Isokinetic knee strength was significantly correlated with body height, body weight, BMI, waist and hip ratio and percent of body fat estimated by skinfold caliper (r = -0.56 to 0.64). The correlation between isokinetic strength with percent of body fat estimated by BIA (r = -0.60 to 0.74; p < 0.001) and with fat free mass (r = 0.64 to 0.78; p < 0.001) was even higher. Although male subjects had significantly greater mean values in body height, body weight, waist to hip ratio and isokinetic strength than female subjects, the MANCOVA showed that the effect of gender on knee isokinetic strength would be eliminated when the covariant variable, the percent of body fat measured by BIA and BMI was controlled in the analysis model. In conclusion, knee isokinetic strength was significantly negatively correlated with proportion of fat and positively correlated with fat free mass. The magnitude of strength difference between males and females could be explained by differences in body fat proportion and BMI in this study. Therapist would take the body fat composition, fat free mass, and BMI into consideration in knee muscle strength measurement. Less body fat and higher BMI will contain more fat free mass that produces more muscle strength. PMID- 11272801 TI - Unusual bilateral peritoneopleural communication associated with cirrhotic ascites: detected by TC-99m sulphur colloid peritoneoscintigraphy. AB - Hydrothorax is an infrequent but well-recognized complication in patients on continuous ambulatory peritoneal dialysis (CAPD) and patients with cirrhotic ascites. It is usually found on the right side; an incidence on both sides is rarely reported. We describe the scintigraphic diagnosis of unusual bilateral peritoneopleural communication in a patient with cirrhotic ascites. PMID- 11272800 TI - Correlation between clinical activity score and thyroid autoantibodies in patients with thyroid ophthalmopathy. AB - To investigate the relation between clinical activity score (CAS) and thyroid autoantibodies of thyroid ophthalmopathy, we measured the level of TSH receptor antibody (TRAb), antithyroglobulin antibody (ATA), and antimicrosomal antibody (AMA) in 41 patients with thyroid ophthalmopathy. The results of thyroid autoantibodies level were compared with CAS. Under the multiple regression and correlation analysis among CAS and the levels of TRAb, ATA, and AMA, no correlation was shown in this study. In conclusion, there is no correlation of thyroid ophthalmopathy among CAS and the levels of TRAb, ATA, and AMA in our study. If we use these three kinds of thyroid autoantibodies to match the activity of thyroid ophthalmopathy, it seems to be inappropriate. Further search of other simplified index to reflect the activity of ophthalmopathy should be encouraged. PMID- 11272802 TI - Adrenal myelolipoma--report of two cases. AB - Adrenal myelolipoma, which is composed of hematopoietic and adipose elements, is a rare benign tumor. Most adrenal myelolipomas are asymptomatic and are found incidentally. We report two cases of adrenal myelolipoma. One was a middle-aged woman with right flank pain. Tumor size increased 8 years later. The other patient was a 63-year-old man presenting with right flank soreness. The right adrenal tumor was found by abdominal sonography. Both of them received adrenalectomy to relieve symptoms and the pathologic results showed adrenal myelolipoma. The clinical, radiologic and pathologic characteristics of these two cases are discussed together with a review of the literature. PMID- 11272804 TI - A novel monoclonal antibody specific for lymphatic endothelium. AB - The difficulty of identifying and differentiating lymphatic and blood microvessels in tissue sections can be overcome by a monoclonal antibody specific for lymphatic endothelium. Unfortunately, the only known antibody also reacts with the endothelium of some blood vessels. The technique of double immunization (passive, with an antiserum to blood endothelium, and active, with a suspension of lymphatic endothelial cells) was, therefore, used to increase the chances of recognizing specific lymphatic antigens by the mouse immune system. The monoclonal antibody obtained, LyMAb, a G1 immunoglobulin, reacted strongly with the endothelium of bovine thoracic duct, mesenteric collecting vessels and lymphatic vessels of gallbladder and lymph nodes and moderately with those of the intestinal wall. Blood vessels (intercostal arteries, azygos vein and blood microvessels of all organs tested) were consistently negative. The antibody was species-specific and did not react with formalin-fixed, paraffin-embedded sections. Cross-reactivity was limited to some connective tissue fibres and scattered cells in the lymph node parenchyma, intestinal villi and hepatic lobules. PMID- 11272803 TI - Histochemical differentiation of autometallographically traceable metals (Au, Ag, Hg, Bi, Zn): protocols for chemical removal of separate autometallographic metal clusters in Epon sections. AB - Nano-sized clusters of gold atoms, or alternatively silver, mercury, bismuth, or zinc sulphide/selenide molecules, can be autometallographically silver-enhanced by being placed in a developer containing reducing molecules and silver ions, i.e. an autometallographic developer. A specific recipe has been worked out for each autometallographically traceable metal, and in cases where two or more autometallographic catalysts are present in the same section it is feasible to distinguish one from the other by chemical removal of one or the other of the metals. In the present study we present protocols that allow differentiation and control of specificity of the established autometallographically detectable metals. It is recommended to implement a multi-element analysis, e.g. proton induced X-ray emission on a few samples to secure the histochemical data. PMID- 11272806 TI - Localization of gamma-tubulin to the basal foot associated with the basal body extending a cilium. AB - The human oviduct epithelium primarily consists of ciliated cells and secretory cells. Solitary cilia usually extend from the apical surface of the secretory cells. We investigated the localization of gamma-tubulin in the ciliary basal apparatus of both cell types by fluorescence immunohistochemistry and immunoelectron microscopy. In addition to basal bodies, gamma-tubulin was identified in the lateral basal foot, especially the basal foot cap. This observation is consistent with previous observations that microtubules radiate from the basal foot and the basal foot serves as the microtubule organizing centre. PMID- 11272805 TI - E-, N- and P-cadherin, and alpha-, beta- and gamma-catenin protein expression in normal, hyperplastic and carcinomatous human prostate. AB - The expression of E-, N- and P-cadherin, alpha-, beta- and gamma-catenin, and actin was studied by immunohistochemistry, ELISA, and Western blot analysis in normal prostates, and in the prostates of men with benign prostatic hyperplasia and men with prostatic carcinoma, in order to evaluate their possible role in the pathogenesis of these diseases. Present results reveal that the immunophenotype of hyperplastic prostates differs from those of both normal and carcinomatous prostates in the intracellular distribution (observed by immunohistochemistry) and the intensity (measured by ELISA) of immunoreactions to cadherins, catenins, and actin. Hyperplastic prostates differ form normal prostates in the weaker immunoreaction to the three cadherin types, the two catenins, and actin, as well as in the intracellular distribution of P-cadherin, beta- and gamma-catenin, and actin. Differences between benign prostatic hyperplasia and prostatic carcinoma are less marked because hyperplastic prostates differ from carcinomatous prostates only in the weaker immunoreactions to P-cadherin, and alpha-catenin. The most remarkable findings in this study were: (1) alpha-catenin production was elevated in prostatic carcinoma in comparison with benign prostatic hyperplasia and normal prostate; and (2) P-cadherin expression in benign prostatic hyperplasia is reduced with regard to those of normal and carcinomatous prostates. It may be concluded that a decreased immunoreaction to cadherins, catenins, and actin, as well as changes in the intracellular distribution of actin in prostatic cells are not necessarily suggestive of malignancy, because these alterations are also present in BPH, and thus, the loss of cadherin-catenin mediated adhesion alone is not sufficient to establish an invasive phenotype. PMID- 11272807 TI - Confocal microscopic evidence of decreased alpha-actin expression within rabbit cerebral artery smooth muscle cells after subarachnoid haemorrhage. AB - Our objective was to determine whether subarachnoid haemorrhage modifies cerebral artery smooth muscle cell phenotype and the contractile protein alpha-actin measured 7 days after haemorrhage. We used a rabbit subarachnoid haemorrhage model and immunofluorescence labelling of alpha-smooth muscle actin, vimentin and desmin. The paired comparison between the haemorrhage and sham rabbits was performed using confocal laser-scanning microscopy. We found in the haemorrhage group significantly less intense alpha-actin immunostaining (p = 0.036) and more intense vimentin immunostaining (p = 0.043) but no significant change in the intensity of desmin staining. Our results indicate an absolute decrease after subarachnoid haemorrhage in the amount of functional alpha-actin and in the light of the literature may suggest a certain degree of dedifferentiation of smooth muscle cells in the cerebral artery wall. PMID- 11272808 TI - Quantitative distribution of NADPH-diaphorase-positive myenteric neurons in different segments of the developing chicken small intestine and colon. AB - NADPH-diaphorase (NADPH-d) was used as a marker for neuronal nitric oxide synthase in order to investigate the nitrergic neurons of the developing myenteric ganglia on whole-mount preparations in the proximal and distal segments of the small intestine and in the colon of the chicken embryo, between incubation days 12 and 19. Neurons that were positive for NADPH-d were counted in randomly selected myenteric ganglia. The data obtained from each area and each age group were subjected to two-way analysis of variance (ANOVA) and the Student-Newman Keuls test. Between incubation days 12 and 19, the originally narrow-meshed myenteric plexus with its high ganglionic density progressively became wide meshed and the ganglionic density decreased significantly. Quantitative analysis further revealed a significant decrease in the NADPH-d-positive nerve cell density with age. At the same time, the constant or even increasing number of nitrergic cells per ganglion may indicate that the decreasing cell density may be a result of the growth of the bowel with decreasing ganglion density rather than a decrease in the total number of myenteric nitrergic cells. Regional differences in the dynamics of the quantitative changes were revealed. A significant decrease in the nitrergic cell number appeared earlier in the proximal than in the distal segments of the small intestine or in the colon. In contrast, the significant decline of the ganglionic density was first noticed in the colon at the same time. PMID- 11272810 TI - Effects of various decalcification protocols on detection of DNA strand breaks by terminal dUTP nick end labelling. AB - To analyse DNA strand breaks by terminal deoxy(d)-UTP nick-end labelling (TUNEL) in calcified tissues including bones and teeth, it is important to decalcify the tissues first. However, the effects of decalcifying reagents on the integrity of DNA are largely unknown. In the present study, we evaluated the usefulness of various decalcifying reagents including 10% EDTA (pH 7.4), 5% trichloroacetic acid (TCA), 5% formic acid, 5% HCl, 10% nitric acid, Plank-Rychlo's solution, Morse's solution and K-CX solution in TUNEL staining. Mouse maxilla was selected as the experimental system. Apoptotic cells naturally occurring in the epithelium were analysed. Tissues were assessed by soft X-ray imaging to confirm complete decalcification. The time required for decalcification of the tissue was 7 days with 10% EDTA and 2 days with other decalcifiers. Decalcified tissues were stained with Methyl/Green-Pyronine Y or 4',6-diamidino-2-phenylindole for assessment of DNA integrity. Nuclei of epithelial cells were strongly positive for both dyes after decalcification with 10% EDTA, 5% TCA, Morse's solution and 5% formic acid. The other reagents failed to retain DNA. Our results demonstrated good TUNEL staining of the maxilla treated with 10% EDTA or 5% TCA. Based on the required time for processing and the signal-noise ratio, we recommend 5% TCA as the decalcifying reagent to analyse for DNA strand breaks. PMID- 11272809 TI - Ultrastructural immunogold cytochemistry with autoimmune human sera and an antibody to uridine implicate human mast cell granules in RNA biology. AB - Human mast cells are professional secretory cells that store synthetic products in large granules filling their cytoplasm. Unlike many secretory cells, the principal synthetic organelle, ribosome-rich endoplasmic reticulum, is a minor component of their cytoplasm. Sightings of nonmembrane-bound ribosomes in and near their secretory granules stimulated detailed ultrastructural studies of various RNA species to implicate secretory-storage granules in RNA biology. In the work reported here, postembedding immunogold ultrastructural cytochemistry indicates that human mast cells contain uridine, an integral ingredient of RNA, and ribonucleoproteins, known to associate with small nuclear RNAs important for splicing RNA precursors, several ribonucleoproteins with possible functions in other aspects of RNA biology and ribonucleoproteins known to associate with ribosomes. These findings should catalyse future work toward establishing the full functional repertoire of secretory-storage granules. PMID- 11272811 TI - A mainstay of functional food science in Japan--history, present status, and future outlook. AB - The development of food science in the near future probably depends on the advance in functional food science, the concept of which was proposed first in Japan nearly 15 years ago. The new science has been internationally distributed and accepted as conceptually being beyond nutrition. In Japan, however, it traced a unique path of progress in the form of a product-driven rather than concept driven science. Actually, a number of substances and products with potential for disease risk reduction rather than simply for health maintenance have been investigated for their body-modulating functions. Some of them have been applied in practice to the industrialization of functional foods in terms of "foods for specified health uses" legally defined by new legislation. A variety of sophisticated methods have been introduced as well, including the so-called "XYZ" evaluation system, database construction for assessment of the function, and even the DNA microarray technique. The Ministry of Agriculture, Forestry, and Fisheries (MAFF) and the Ministry of Health and Welfare (MHW) also commenced their scientific as well as political activity, with its spread to industries which almost simultaneously began to vigorously investigate functional food products for enlargement of the food market. With all of this as a background, the Japan Liaison of the International Union of Food Science and Technology (IUFoST) hold a function food science symposium on behalf of related scientific bodies including the Japan Section of the International Life Science Institute (ILSI). This paper is an overview compiled from 12 presentations made in the symposium, with the aim of internationally publicizing the activity of functional food science in Japan. PMID- 11272812 TI - Purification and properties of thiosulfate dehydrogenase from Acidithiobacillus thiooxidans JCM7814. AB - A key enzyme of the thiosulfate oxidation pathway in Acidithiobacillus thiooxidans JCM7814 was investigated. As a result of assaying the enzymatic activities of thiosulfate dehydrogenase, rhodanese, and thiosulfate reductase at 5.5 of intracellular pH, the activity of thiosulfate dehydrogenase was measured as the key enzyme. The thiosulfate dehydrogenase of A. thiooxidans JCM7814 was purified using three chromatographies. The purified sample was electrophoretically homogeneous. The molecular mass of the enzyme was 27.9 kDa and it was a monomer. This enzyme had cytochrome c. The optimum pH and temperature of this enzyme were 3.5 and 35 degrees C. The enzyme was stable in the pH range from 5 to 7, and it was stable up to 45 degrees C. The isoelectric point of the enzyme was 8.9. This enzyme reacted with thiosulfate as a substrate. The Km was 0.81 mM. PMID- 11272813 TI - Isolation of antidiabetic components from white-skinned sweet potato (Ipomoea batatas L.). AB - We have already reported that white-skinned sweet potato (Ipomoea batatas L.) (WSSP) shows antidiabetic activity in streptozotocin (STZ) induced diabetic rats and genetically diabetic models (yellow KK, db/db mice and Zucker fatty rats). In this study, isolation and purification of the antidiabetic component of WSSP were attempted. Almost all antidiabetic activity was found in the cortex of WSSP. The fractionation of the antidiabetic component in the WSSP cortex was done by the following methods: dialysis of the water extract, 85% ethanol precipitation, 15% trichloroacetic acid (TCA) treatment, butyl-, phenyl-hydrophobic column chromatography, and ultrafiltration treatment. The antidiabetic component was not eliminated during dialysis and was soluble in 85% ethanol and 15% TCA, but it passed through a filter that allows the passage of substances of a molescular weight of 30,000. The uniformity of this isolated active component was analyzed using HPLC. A single peak was seen with three different columns (C8 reverse-phase column, anion exchange QA column, and gel filtration column (GFC)), indicating that the component is a uniform substance. The molecular weight of this antidiabetic component was estimated to be 22,000 by GFC analysis. This active component was presumed to be an acidic glycoprotein because it contained protein and sugar and was adsorbed onto the QA column at pH 7.0. PMID- 11272814 TI - Membrane-bound sugar alcohol dehydrogenase in acetic acid bacteria catalyzes L ribulose formation and NAD-dependent ribitol dehydrogenase is independent of the oxidative fermentation. AB - To identify the enzyme responsible for pentitol oxidation by acetic acid bacteria, two different ribitol oxidizing enzymes, one in the cytosolic fraction of NAD(P)-dependent and the other in the membrane fraction of NAD(P)-independent enzymes, were examined with respect to oxidative fermentation. The cytoplasmic NAD-dependent ribitol dehydrogenase (EC 1.1.1.56) was crystallized from Gluconobacter suboxydans IFO 12528 and found to be an enzyme having 100 kDa of molecular mass and 5 s as the sedimentation constant, composed of four identical subunits of 25 kDa. The enzyme catalyzed a shuttle reversible oxidoreduction between ribitol and D-ribulose in the presence of NAD and NADH, respectively. Xylitol and L-arabitol were well oxidized by the enzyme with reaction rates comparable to ribitol oxidation. D-Ribulose, L-ribulose, and L-xylulose were well reduced by the enzyme in the presence of NADH as cosubstrates. The optimum pH of pentitol oxidation was found at alkaline pH such as 9.5-10.5 and ketopentose reduction was found at pH 6.0. NAD-Dependent ribitol dehydrogenase seemed to be specific to oxidoreduction between pentitols and ketopentoses and D-sorbitol and D-mannitol were not oxidized by this enzyme. However, no D-ribulose accumulation was observed outside the cells during the growth of the organism on ribitol. L Ribulose was accumulated in the culture medium instead, as the direct oxidation product catalyzed by a membrane-bound NAD(P)-independent ribitol dehydrogenase. Thus, the physiological role of NAD-dependent ribitol dehydrogenase was accounted to catalyze ribitol oxidation to D-ribulose in cytoplasm, taking D-ribulose to the pentose phosphate pathway after being phosphorylated. L-Ribulose outside the cells would be incorporated into the cytoplasm in several ways when need for carbon and energy sources made it necessary to use L-ribulose for their survival. From a series of simple experiments, membrane-bound sugar alcohol dehydrogenase was concluded to be the enzyme responsible for L-ribulose production in oxidative fermentation by acetic acid bacteria. PMID- 11272815 TI - Effects of pH and metal ions on antioxidative activities of catechins. AB - The Effects of pH on antioxidative activities of catechol, pyrogallol, and four catechins, and effects of metal ions (Al3+, Ca2+, Cd2+, Co2+, Cr3+, Cu2+, Fe2+, Fe3+, K+, Mg2+, Mn2+, Na+, and Zn2+) on antioxidative activities of (-) epigallocatechin gallate (EGCG) were studied by an oxygen electrode method. The antioxidative activities of catechins were high and constant at pH 6-12, but decreased in acidic and strong alkaline solutions. Copper(II) ion the most strongly increased the antioxidative activity of EGCG among these metal ions examined, but iron(II) ion largely inhibited the antioxidative activity of EGCG. These effects are discussed considering the formation of metal complexes with catechins and the change in oxidation potentials. PMID- 11272816 TI - Purification and characterization of bifunctional alginate lyase from Alteromonas sp. strain no. 272 and its action on saturated oligomeric substrates. AB - A marine bacterium (strain No. 272) isolated from sea mud in Omura Bay produced an alginate lyase and was classified as an Alteromonas species. The enzyme was purified from the culture medium of the bacterium by DEAE-Cellulofine, Sephadex G 100 gel chromatography to an electrophoretically homogeneous state in the presence and absence of SDS. The molecular mass of the enzyme was 23 and 33.9 kDa on Sephadex G-100 column chromatography and SDS-polyacrylamide gel electrophoresis, respectively, with an isoelectric point of 3.8. The predominant secondary structure of the enzyme was found to be most likely beta-structure by circular dichroism. The enzyme was most active at pH 7.5-8.0 and stable around pH 5-11. The enzyme was more labile in Tris-HCI buffer (pH 7.0) to heat treatment, than in phosphate buffer (pH 7.0). No of metal ions significantly affected the enzyme activity. The enzyme acted on sodium alginate in an endo-type manner and on two components of alginate, poly-alpha1,4-L-guluronate and poly-beta1,4-D mannuronate, as judged by routine ultraviolet assay (235 nm) and circular dichroic spectral changes of the substrates. However, the coexisting poly alpha1,4-L-guluronate and poly-beta1,4-D-mannuronate apparently interacted with the enzyme in a competitive manner. Although the enzyme depolymerized alginate in an endo-type, it did not act on trimeric guluronate and mannuronate, but on the tetramers or more. The kinetic analyses showed that kcat/Km for each oligomer was larger for the guluronate oligomers than for the mannuronate ones, and that the subsite structure of the enzyme most likely consisted of six binding sites from the intrinsic reaction rate constant (kint) and intrinsic substrate binding constant (Kint). PMID- 11272817 TI - Detoxification effect of iron-encaging zeolite-processed water in tributyltin intoxicated Euglena gracilis Z. AB - In our previous paper, we reported the restoration promoting effects of mineral encaging zeolite-processed water, especially of a Fe-encaging one, on tributyltin chloride (TBTCl)-intoxicated Euglena gracilis. This present study extends the investigation on the behavior of TBTCl and a xenobiotic enzyme, cytochrome P-450, in Euglena cells incubated with or without Fe-encaging zeolite-processed water (FeZW). Subcellular fractionation of TBTCl-intoxicated Euglena cells, atomic absorption spectrophotometry, and GC analyses showed that TBTCl was rapidly incorporated into the cells to halt cell motility. GC-MS showed that FeZW promoted conversion of TBTCl to dibutyltin (DBT) as the major metabolite in the microsomal fraction of the cells. An in vitro incubation system with heat-treated microsomes did not convert TBTCl to DBT. The contribution of cytochrome P-450 in the microsomal fraction was suggested by an immunochemical method. The results suggest that the improvement of detoxification by FeZW in the TBT-intoxicated Euglena cells should be due to activation of biotransformation system of the Euglena cells by FeZW. PMID- 11272818 TI - Development of assay system for immunoglobulin production regulatory factors using whole cell cultures of mouse splenocytes. AB - We tried to establish an assay system for screening and assessment of immunoregulatory factors using whole cell cultures of mouse splenocytes and found that splenic adhesive cells markedly increased immunogobulin (Ig) production of splenocytes. In the absence of adhesive cells, lipopolysaccharides, pokeweed mitogen, and phytohemagglutinin stimulated the production of IgA, IgG, and IgM in a class-dependent manner. Adhesive cells increased more markedly Ig production of splenocytes stimulated with these mitogens. When mouse splenocytes were cultured with milk proteins in the absence of adhesive cells, lactoferrin, beta lactoglobulin, alpha-casein, and beta-casein stimulated IgA and IgG production. Adhesive cells increased IgA production of splenocytes stimulated with milk proteins, especially. These results suggest that the assay system is useful for assessment of Ig production-regulating factors. PMID- 11272819 TI - Cloning and analysis of valerophenone synthase gene expressed specifically in lupulin gland of hop (Humulus lupulus L.). AB - Resin and essential oil derived from hop (Humulus lupulus L.) cones are very important compounds for beer brewing, and they specifically accumulate in the lupulin gland of hop cones. In order to identify the genes responsible for the biosynthetic pathway of these compounds and use the identified genes for hop breeding using Marker Assisted Selection and transformation techniques, genes expressed specifically in the lupulin gland were cloned and sequenced. One of them was suggested to be similar to the chalcone synthase gene from the DNA sequence. The translation product of the gene had the activity of valerophenone synthase, which catalyzes a part of the synthesis reaction of alpha-acid and beta acid. Northern analysis showed that the valerophenone synthase gene seemed to be expressed specifically in the lupulin gland. PMID- 11272820 TI - A calcium-binding protein with four EF-hand motifs in Streptomyces ambofaciens. AB - A gene (cabA) encoding a calcium-binding protein was cloned from Streptomyces ambofaciens. CabA was 180 amino acid residues long and contained four typical EF hand motifs bearing high sequence similarity to the calcium-binding sites in calmodulin. Consistent with this, CabA showed distinct calcium-binding activity, comparable to bovine brain calmodulin. cabA was transcribed throughout growth, as found by S1 nuclease mapping. Southern hybridization experiments showed that a single copy of cabA was present in various Streptomyces species. A hypothetical relationship between CabA and aerial mycelium formation in this strain was examined, since S. ambofaciens showed calcium-dependent aerial mycelium formation. However, disruption of cabA or overexpression of cabA in S. ambofaciens caused no detectable phenotypic changes. PMID- 11272821 TI - Syntheses and biological activities of pyranyl-substituted cinnamates. AB - Twenty-two kinds of pyranyl-substituted cinnamates were synthesized by the reaction of 4-hydroxy-6-(2-phenylethyl)-2H-pyran-2-one or 4-hydroxy-6-methyl-2H pyran-2-one (HMP) with a variety of substituted cinnamic acids, and their antifungal and plant growth inhibitory activities were investigated. Among the compounds prepared, 6-methyl-2-oxo-2H-pyran-4-yl 3-(4-isopropylphenyl)propenoate (H5) showed the strongest antifungal activity against Rhizoctonia solani and Sclerotium dellfinii, and 6-methyl-2-oxo-2H-pyran-4-yl 3-(2 methylphenyl)propenoate (H2) had the highest plant growth inhibitory activity toward Brassica rapa. PMID- 11272822 TI - Intraspecific divergence of Saccharomyces kluyveri as revealed by the nucleotide sequences of 18S-28S rRNA spacer regions and alpha-galactosidase MEL genes. AB - In the five strains classified as the yeast Saccharomyces kluyveri, several substitutions were observed in the two internal transcribed spacer regions between 18S and 28S rRNA. A PCR reaction with primers targeted to the MEL1 gene of Saccharomyces cerevisiae amplified fragments of the expected size, and those sequences showed significant divergence in the strains of S. kluyveri. PMID- 11272823 TI - Synthesis of the racemate of the stereoisomer at C-6a of BE-40644, a bioactive metabolite of Actinoplanes sp. with a sesquiterpene-substituted p-benzoquinone structure. AB - BE-40644 is a tetracyclic metabolite of Actinoplanes sp. A 40644 possessing a sesquiterpene-substituted p-benzoquinone structure with cis-fused B/C ring stereochemistry that inhibits the human thioredoxin system as the well as the growth of several cancer cell lines. Its B/C trans-fused stereoisomer at C-6a was synthesized as a racemate starting from geranylacetone and 3,5-dihydroxybenzoic acid. PMID- 11272824 TI - Synthesis of corollosporine, an antibacterial metabolite of the marine fungus Corollospora maritima. AB - Corollosporine [(+/-)-3-hexyl-3,7-dihydroxy-1(3H)-isobenzofuran-1-one], an antibacterial metabolite of the marine fungus, Corollospora maritima, was synthesized by four different routes from 3-hydroxyphthalic anhydride or 2 methoxybenzoic acid as the starting material to verify its proposed structure. PMID- 11272825 TI - Monomolecular layer formation of ferritin molecules on an amphiphilic cyclodextrin derivative at the air/water interface. AB - A monomolecular layer of ferritin molecules was formed by adsorption from the subphase onto a Langmuir film of an amphiphilic beta-cyclodextrin (beta-CD) derivative at the air/water interface. The course of the adsorption of ferritin molecules was monitored by measuring the surface pressure and the resulting film was observed by transmission electron microscopy (TEM). These results show the potential of the amphiphilic CD derivative to work as a milder template for protein molecules at the air/water interface. PMID- 11272826 TI - An efficient synthesis of C2-symmetric chiral binaphthyl ketone catalysts. AB - An efficient synthesis of C2-symmetric chiral binaphthyl ketones 1a and b, effective catalysts for asymmetric epoxidation, is reported. The key features of this synthesis are Co(salen)-catalyzed macrolactonization of racemic 1,1' binaphthyl-2,2'-dicarboxylic acid monoglycidyl esters 3a and b and lipase catalyzed enantioselective acylation of resulting 11-membered cyclic binaphthyl alcohols 4a and b. PMID- 11272827 TI - A simple, rapid, and highly efficient gene expression system for multiheme cytochromes c. AB - The genes of tetraheme cytochrome c3 and hexadecaheme high-molecular-weight cytochrome c from Desulfovibrio vulgaris could be overexpressed as holoproteins in Shewanella oneidensis TSP-C using pUC-type vectors of E. coli. Surprisingly, S. oneidensis was transformed directly by pUC-type vectors through electroporation. The yields of the recombinant proteins in this expression system were much higher than the previously reported ones. PMID- 11272828 TI - Isolation of the rpoD gene encoding the principal sigma factor of the deep-sea piezophilic bacterium Shewanella violacea strain DSS12 and its overexpression in Escherichia coli. AB - The gene encoding the principal a factor (rpoD) of the piezophilic bacterium Shewanella violacea was cloned and sequenced. The rpoD gene was found to encode a polypeptide consisting of 614 amino acid residues, showing 75.6 and 64.3% identity to those of Escherichia coli and Pseudomonas putida, respectively. Comparison with E. coli sigma70 and P. putida sigma70 showed that significant similarity exists in four conserved regions known to be required for promoter recognition and core binding. Using an expression plasmid harboring the rpoD gene, the S. violacea sigma70 factor was overexpressed in E. coli and successfully purified to near homogeneity. PMID- 11272829 TI - In vitro absorption and metabolism of nobiletin, a chemopreventive polymethoxyflavonoid in citrus fruits. AB - The polymethoxyflavonoid (PMF), nobiletin (NOB), specifically occurs in citrus fruits, and is currently believed to be a promising anti-inflammatory and antitumor promoting agent. In the present study, we investigated the in vitro absorption and metabolism of NOB and compared them with those of the polyhydroxyflavonoid (PHF), luteolin (LT). NOB preferentially accumulated in a differentiated Caco-2 cell monolayer, which is a model for small intestinal epithelial cells, while LT did not. Treatment of NOB with a rat liver S-9 mixture led to the formation of 3'-demethyl-NOB, while that of LT did not. We thus suggest that PMFs including NOB have properties distinct from those of general flavonoids for absorption and metabolism in vitro. PMID- 11272830 TI - Apoptosis-inducing activity of a driselase digest fraction of green tea residue. AB - We enzymatically digested green tea residue with Driselase, a crude preparation containing cellulase, pectinase and proteases, in order to examine the potential usefulness of the residue. A fraction of the digest soluble in 70% ethanol was found to induce the death of U937 human histiocytic lymphoma cells by apoptosis. Other enzyme preparations gave similar products with cell death-inducing activity of varing potency. The green tea residue may therefore be a useful source of potential agents with anti-cancer activity. PMID- 11272831 TI - Uptake and accumulation of exogenous docosahexaenoic acid by Chlorella. AB - Tuna oil or its hydrolysate was added to a culture of Chlorella for its nutritional fortification as a feed for rotifer. Exogenous docosahexaenoic acid (DHA) in its free form was taken up by the cells of Chlorella vulgaris strain K 22 and by other strains, but tuna oil was not taken up by the cells. Accumulated DHA was found by electron microscopy in the cells in oil droplets. All strains of Chlorella used in these experiments took up exogenous DHA into the cells. It seems that the structure of the cell wall did not affect the uptake of DHA into the Chlorella cells. PMID- 11272832 TI - Characterization of PBZ1, a probenazole-inducible gene, in suspension-cultured rice cells. AB - Probenazole (PBZ) induces non-race specific resistance in rice plants against rice blast fungus and PBZ1 was identified as a PBZ-inducible gene from rice. The induction of PBZ1 expression in suspension-cultured rice cells was investigated. Northern blot analysis indicated that PBZ1 was induced by PBZ in a dose-dependent manner. Enzyme-linked immunosorbent assay (ELISA) showed a dose and time dependent accumulation of PBZ1 protein. Both mRNA and protein analysis showed that PBZ1 was not induced by salicylic acid or an active metabolite, 1,2 benzisothiazole-1,1-dioxide. PMID- 11272833 TI - Sequence analysis and overexpression of a pectin lyase gene (pel1) from Aspergillus oryzae KBN616. AB - A gene (pel1) encoding pectin lyase (Pel1) was isolated from a shoyu koji mold, Aspergillus oryzae KBN616, and characterized. The structural gene comprised 1,196 bp with a single intron. The ORF encoded 381 amino acids with a signal peptide of 20 amino acids. The deduced amino acid sequence showed high similarity to those of Aspergillus niger pectin lyases and Glomerella cingulata PnlA. The pel1 gene was successfully overexpressed under the promoter of the A. oryzae TEF1 gene. The molecular mass of the recombinant pectin lyase substantially coincided with that calculated based on nucleotide sequence. PMID- 11272835 TI - Primary culture of chicken hepatocytes as an in vitro model for determining the influence of dioxin. AB - An easy method for primary culture of chicken hepatocytes was developed to study the influence of dioxin on birds. Chicken hepatocytes could maintain gene expression and protein secretion of albumin for a long period in serum-free medium with free atmosphere exchange at 37 degrees C. Moreover, the cells showed a sensitive response to 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD) by monitoring the expression of P450 1A, theta GST (theta-GST) and albumin genes. PMID- 11272834 TI - Effects of the Escherichia coli sfsA gene on mal genes expression and a DNA binding activity of SfsA. AB - The sfsA gene was identified as one of the sfs genes the over-expression of which stimulates maltose fermentation of the Mal- Escherichia coli strain MK2001 (crp*1, cya:Km(r)). Expression from the malPQ promoter, which was measured using a chromosomally integrated malPp-lacZ fusion, was induced by over-expressing the sfsA gene in the crp*1, cya:Km(r) strain. The level of the MalE protein was increased in crp*1, cya:Km(r) cells over-producing SfsA. The SfsA protein was purified to homogeneity and tested for DNA binding activity. The purified SfsA protein binds to DNA non-specifically. All these results may suggest that SfsA functions as a DNA binding protein to induce the mal genes in coordination with CRP*1. PMID- 11272837 TI - Crystallization and structural analysis of intact maltotetraose-forming exo amylase from Pseudomonas stutzeri. AB - The intact maltotetraose-forming exo-amylase from Pseudomonas stutzeri (G4-1), which has a raw starch binding domain, has been crystallized. The structure was identified (PDB entry 1GCY) by the molecular replacement method using the structure of its catalytic domain (G4-2). The result showed that the raw starch binding domain is in a disordered state, the corresponding electron densities being almost invisible. Superposition of these two enzyme forms showed evidence for the possible location of the raw starch binding domain (SBD). This crystal is a novel case, in that it forms a regular lattice incorporating flexibly bound SBD in the channel of crystal packing of the catalytic domains. PMID- 11272836 TI - Molecular cloning, characterization, and expression of wheat cystatins. AB - We cloned four kinds of cDNAs of wheat cystatins (WCs), WC1, WC2, WC3, and WC4, from the seed. They had 47-68% amino acid sequence similarities to other plant cystatins. WC1, WC2, and WC4 had 63-67% similalities to one another while 93% of amino acids were identical between WC1 and WC3. This suggested that WCI, WC2, and WC4 should be regarded as the isoforms of wheat cystatins. The mRNAs for WC1, WC2, and WC4 were all expressed in seed at an early stage of maturation and, after that, their quantities decreased gradually. However, each of the mRNAs was again expressed one day after the start of germination and the expression continued for the following five days. WC1 seemed to be expressed at a higher level than WC2 and WC4. Immunostaining for looking at site-specific expression of each WC demonstrated that both WC1 and WC4 existed in the aleuron layer and embryo, but in the endosperm the only existing species was WC1. Differences in mRNA level and tissue localization found for the WCs may suggest their differential physiological roles. PMID- 11272838 TI - A cryptic plasmid, pAO1, from a compost bacterium, Bacillus sp. AB - The complete nucleotide sequence of a new cryptic plasmid, pAO1 isolated from a compost bacterium Bacillus sp., has been analyzed. Analysis of the PCR-based 16S rRNA sequence showed the bacterium harboring pAO1 was closely related to Bacillus pallidus. The plasmid pAO1 was 3,325 bp in size. Two open reading frames, ORF1 and ORF2, encoding putative polypeptides of 248 and 290 amino acids, respectively, were identified within the sequence. The ORF1 has a limited sequence similarity to an integrase/recombinase, while the ORF2 has high similarity with the replication protein of pBC1 from Bacillus coagulans. A putative origin sequence for a plus-strand was located between ORFs. Southern blot analysis indicates this plasmid replicates via a rolling circle-type mechanism. PMID- 11272839 TI - Translational regulation of the mRNA encoding the eukaryotic translation initiation factor 4E in Xenopus. AB - We have cloned the cDNA for Xenopus eukaryotic translation initiation factor 4E (eIF4E). Here we show that translation of a luciferase mRNA that contains the 5' untranslated region derived from Xenopus eIF4E is active in fertilized eggs, but is repressed in oocytes. The results suggest that the expression of Xenopus eIF4E is regulated at the translation level. PMID- 11272840 TI - Analysis of the isoform of Xenopus euakryotic translation initiation factor 4E. AB - We have found two isoforms of the eukaryotic translation initiation factor 4E (eIF4E) in Xenopus laevis. These proteins differ in length by 18 amino acids. Overexpression of either of the two eIF4E proteins modestly increase translation in Xenopus oocytes. The results suggest that both of these two isoforms function in translation. PMID- 11272841 TI - Enterostatin (VPDPR) has anti-analgesic and anti-amnesic activities. AB - Enterostatin (VPDPR), an anorexigenic peptide derived from the amino terminus of procolipase, significantly inhibited analgesia induced by the mu-opioid agonist morphine (5 mg/kg, s.c.) after i.c.v. administration to mice at a dose of 100 nmol. On the other hand, VPDPR (approximately 200 nmol, i.c.v.) did not attenuate analgesia induced by the kappa-opioid agonist D-Phe-D-Phe-D-Nle-D-Arg-NH2 (100 microg/mouse, i.c.v.) or delta-opioid agonist DTLET (4 nmol/mouse, i.c.v.). VPDPR (100 nmol, i.c.v.) significantly improved amnesia induced by scopolamine (0.2 mg/kg, i.p.) in mice. However, VPDPR did not enhance memory in normal mice at the same dose. PMID- 11272843 TI - New 11(15-->1)abeotaxane, 11(15-->1),11(10-->9)bisabeotaxane and 3,11 cyclotaxanes from Taxus yunnanensis. AB - Chemical examination of the seeds of Chinese yew, Taxus yunnanensis Cheng et L. K. Fu resulted in the isolation of an 11(15-->1)abeotaxane, an 11(15-->1), 11(10- >9)bisabeotaxane and two 3,11-cyclotaxanes. The structures of these new taxoids were established as 13alpha-acetoxy-5alpha-cinnamoyloxy-11(15-->1)abeotaxa 4(20),11-diene-9alpha,10beta,15-triol (1), 20-acetoxy-2alpha-benzoyloxy-4alpha, 5alpha, 7beta, 9alpha, 13alpha-pentahydroxy-11(15-->1), 11(10-->9) bisabeotax-11 eno-10,15-lactone (2), 2alpha,10beta-diacetoxy-5alpha-cinnamoyloxy-9alpha-hydroxy 3,11 -cyclotax-4(20)-en-13-one (3) and 10beta-acetoxy-2alpha,5alpha,9alpha trihydroxy-3,11-cyclotax-4(20)-en-13-one (4) on the basis of spectral analyses. PMID- 11272842 TI - Improved procedure for the enantiometric synthesis of 1-hydroxy/acetoxy-2,6 diaryl-3,7-dioxabicycl. AB - Short enantiomeric syntheses of the 1-hydroxy/acetoxy-3,7 dioxabicyclo[3.3.0]octane lignans, paulownin, and (+)-phrymarin I and II, were accomplished by starting from the chiral synthon, (R)-(+)-3-hydroxybutanolide, and employing photocyclization as the key step. PMID- 11272844 TI - Importance of the carbohydrate-binding module of Clostridium stercorarium Xyn10B to xylan hydrolysis. AB - The Clostridium stercorarium xylanase Xyn10B is a modular enzyme comprising two thermostabilizing domains, a family 10 catalytic domain of glycosyl hydrolases, a family 9 carbohydrate-binding module (CBM), and two S-layer homologous (SLH) domains [Biosci. Biotechnol. Biochem., 63, 1596-1604 (1999)]. To investigate the role of this CBM, we constructed two derivatives of Xyn10B and compared their hydrolytic activity toward xylan and some preparations of plant cell walls; Xyn10BdeltaCBM consists of a catalytic domain only, and Xyn10B-CBM comprises a catalytic domain and a CBM. Xyn10B-CBM bound to various insoluble polysaccharides including Avicel, acid-swollen cellulose, ball-milled chitin, Sephadex G-25, and amylose-resin. A cellulose binding assay in the presence of soluble saccharides suggested that the CBM of Xyn10B had an affinity for even monosaccharides such as glucose, galactose, xylose, mannose and ribose. Removal of the CBM from the enzyme negated its cellulose- and xylan-binding abilities and severely reduced its enzyme activity toward insoluble xylan and plant cell walls but not soluble xylan. These findings clearly indicated that the CBM of Xyn10B is important in the hydrolysis of insoluble xylan. This is the first report of a family 9 CBM with an affinity for insoluble xylan in addition to crystalline cellulose and the ability to increase hydrolytic activity toward insoluble xylan. PMID- 11272845 TI - Volatile flavor components of ripe and overripe ki-mikans (Citrus flaviculpus Hort. ex Tanaka) in comparison with Hyuganatsu (Citrus tamurana Hort. ex Tanaka). AB - The volatile flavor components of ripe and overripe ki-mikan (Citrus flaviculpus Hort. ex Tanaka) peel oil samples, which had been isolated by cold-pressing, were investigated by capillary GC and GC-MS, and compared with the Hyuganatsu (Citrus tamurana Hort. ex Tanaka) flavor. Limonene (ripe fruit, 82.44%; overripe fruit, 73.10%) was the most abundant compound in the ki-mikan oil, this being followed by gamma-terpinene (8.83% and 13.74%), trans-beta-farnesene (1.76% and 3.12%) and myrcene (1.54% and 1.13%). The composition of overripe ki-mikan oil was characterized by higher amounts of aliphatic and sesquiterpene hydrocarbons, monoterpene and sesquiterpene alcohols, ketones and esters than that of ripe ki mikan oil. Monoterpene hydrocarbons, especially limonene (84.78%), were predominant in Hyuganatsu oil. The CPO composition of ki-mikan was qualitatively similar to that of Hyuganatsu, but differed quantitatively. The content of sesquiterpene hydrocarbons was higher in the ki-mikan oil samples than in Hyuganatsu oil, while ketones showed the opposite predominance. These differences were more evident in the trans-beta-farnesene and l-carvone contents. The ratio of both these compounds could be used to distinguish ki-mikan oil from Hyuganatsu oil. PMID- 11272846 TI - Molecular cloning of rat USF2 cDNA and characterization of splicing variants. AB - The complete nucleotide sequence of rat USF2 cDNA was determined. In addition to the full length clone (USF2FL), four isoforms (delta1, delta2, delta3, and delta4) suggested to be generated by alternative splicing were isolated. USF2delta1 and delta2 lacked 27 and 67 internal amino acid residues, respectively. USF2delta3 and delta4 lacked most of the entire sequence but encoded short peptides of an N-terminal portion of USF2FL. Overexpression of USF2FL increased the transcription of the human high affinity IgE receptor (FcepsilonRI) alpha chain gene through specific binding to the CAGCTG motif in the first intron. On the other hand, overexpression of USF2delta1 or delta2 reduced the transcription of the human FcepsilonRI alpha chain gene. Both USF2FL and USF2delta1 bound to CACGTG as well as CAGCTG, while USF2delta2 bound to CACGTG but not to CAGCTG. These results suggested the presence of a different and definitive role of each variant in the expression of the alpha chain gene. PMID- 11272847 TI - Purification and some properties of ubiquinol oxidase from obligately chemolithotrophic iron-oxidizing bacterium, Thiobacillus ferrooxidans NASF-1. AB - Ubiquinol-oxidizing activity was detected in an acidophilic chemolithotrophic iron-oxidizing bacterium, T. ferrooxidans. The ubiquinol oxidase was purified 79 fold from plasma membranes of T. ferrooxidans NASF-1 cells. The purified oxidase is composed of two polypeptides with apparent molecular masses of 32,600 and 50,100 Da, as measured by gel electrophoresis in the presence of sodium dodecyl sulfate. The absorption spectrum of the reduced enzyme at room temperature showed big peaks at 530 and 563, and a small broad peak at 635 nm, indicating the involvement of cytochromes b and d. Characteristic peaks of cytochromes a and c were not observed in the spectrum at around 600 and 550 nm, respectively. This enzyme combined with CO, and its CO-reduced minus reduced difference spectrum showed peaks at 409 nm and 563 nm and a trough at 431 nm. These results indicated that the oxidase contained cytochrome b, but the involvement of cytochrome d was not clear. The enzyme catalyzed the oxidations of ubiquinol-2 and reduced N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride. The ubiquinol oxidase activity was activated by the addition of albumin and lecithin to the reaction mixture and inhibited by the respiratory inhibitors KCN, HQNO, NaN3, and antimycin A1, although the enzyme was relatively resistant to KCN, and the divalent cation, Zn2+, compared with ubiquinol oxidases of E. coli. PMID- 11272848 TI - Soy protein hydrolyzate with bound phospholipids reduces serum cholesterol levels in hypercholesterolemic adult male volunteers. AB - This study was done to evaluate the effects of soy protein hydrolyzate with bound phospholipids (c-SPHP), on the serum cholesterol levels in hypercholesterolemic subjects over a three-month period. Subjects were Taiwanese adult male volunteers whose serum total cholesterol levels were above 220 mg/dl. Twenty-one subjects were divided into three groups randomly, and each group was given c-SPHP zero, 3, or 6 g per day. Test diets were orally administered in a powdered drink form that contained c-SPHP or casein hydrolyzate (placebo). The subjects were given the test diet four times daily. The study consisted of a two-week pre-feeding period, a three-month feeding period, followed by a two-week post-feeding period. After 3 months of c-SPHP administration, 3 g per day, serum total cholesterol decreased significantly from the initial level (15.0%, p<0.01) and compared with the placebo group (p<0.05). Furthermore, LDL-cholesterol decreased significantly (27.7%, p<0.01) and the LDL/HDL ratio also decreased significantly (47.4%, p<0.01) from the initial levels. These effects of c-SPHP were dose-dependent. This study suggests that c-SPHP has remarkable improving effects on the serum cholesterol levels in hypercholesterolemic subjects. PMID- 11272849 TI - Direct expression of the extracellular portion of human FcepsilonRIalpha chain as inclusion bodies in Escherichia coli. AB - The extracellular portion of the alpha chain of the human high-affinity IgE receptor (FcepsilonRIalpha) was expressed as inclusion bodies in Escherichia coli. In immunoblot analysis, two bands were reactive to human IgE and mouse anti human FcepsilonRIalpha monoclonal antibodies. N-terminal sequencing showed that the two bands were equivalent to the soluble FcepsilonRIalpha with a methionine residue at the N-terminus (Met-1-172) and 23-172, in which the N-terminal 22 residues of the soluble FcepsilonRIalpha have been removed, possibly by degradation in E. coli cells. IgE-binding to CHO cells expressing FcepsilonRI was inhibited by the addition of the recombinant products prepared by the refolding procedure from inclusion bodies. The system for the expression of soluble human FcepsilonRIalpha in E. coli presented in this study and its further improvement would be useful for the production of the protein as a potent therapeutic and for analysis of the IgE-FcepsilonRIalpha interaction. PMID- 11272850 TI - Domain construction of cherry-tomato lectin: relation to newly found 42-kDa protein. AB - In the early stage of ripening of cherry-tomato fruits (Lycopersicon esculentum var. cherry), the lectin activity increased logarithmically and reached a plateau at day 10 after flowering. During purification of lectin from ripe and unripe fruits, a 42-kDa protein was found abundantly in unripe fruits. The protein cross reacted with anti-cherry-tomato-lectin serum, retained chitin-binding ability, but showed no lectin activity. Comparative studies between the structure of the lectin and the 42-kDa protein were done. N-Terminal amino acid sequences of the lectin, peptides derived from the S-pyridylethylated lectin, and fragments generated by limited proteolysis of the native lectin showed that the lectin was comprised of three domains, Hyp-rich, Cys-rich, and Gln-rich, and the alignment of them was as this order from the N-terminus. Studies on the 42-kDa protein showed that it contained two of the three domains, Cys-rich and Gln-rich, but the amino acid sequence analysis showed that the protein should be a product of another gene. PMID- 11272851 TI - High-level secretory production of phospholipase A1 by Saccharomyces cerevisiae and Aspergillus oryzae. AB - Phospholipase A1 (PLA1) is a hydrolytic enzyme that catalyzes the removal of the acyl group from position 1 of lecithin to form lysolecithin. The PLA1 gene, which had been cloned from Aspergillus oryzae, was expressed in Saccharomyces cerevisiae and A. oryzae. Through the modification of the medium composition and the feeding conditions of substrate, the production level of PLA1 by S. cerevisiae was increased to a level fivefold higher than that indicated in a previous report. In the case of A. oryzae, introduction of multicopies of PLA1 expression units, and the morphological change from the pellet form to the filamentous form were effective for the enhancement of PLA1 production. We succeeded in producing 3,500 U/ml of PLA1 using an industrial-scale fermentor. PMID- 11272852 TI - Light therapy. PMID- 11272853 TI - First Nations women's encounters with mainstream health care services. AB - Health care encounters are important areas for study because they reflect social, political, economic, and ideological relations between patients and the dominant health care system. This study examines mainstream health care encounters from the viewpoint of First Nations women from a reserve community in northwestern Canada. Perspectives from critical medical anthropology and the concept of cultural safety provided the theoretical orientation for the study. Critical and feminist ethnographic approaches were used to guide in-depth interviews conducted with 10 First Nations women. Findings were organized around two broad themes that characterized women's descriptions of "invalidating" and "affirming" encounters. These narratives revealed that women's encounters were shaped by racism, discrimination, and structural inequities that continue to marginalize and disadvantage First Nations women. The women's health care experiences have historical, political, and economic significance and are reflective of wider postcolonial relations that shape their everyday lives. PMID- 11272854 TI - Cognitive impairment, disruptive behaviors, and home care utilization. AB - Disruptive behaviors by an elder with cognitive impairment, such as violence, abrupt mood swings, and overt inappropriate sexual behaviors have been known to create caregiver distress, but little is known about how these behaviors may influence the use of home care services by that elder or their informal caregiver. The purpose of this study is to explore the associations between type of cognitive impairment (cognitive impairment with no dementia, and dementia), selected disruptive behaviors, and the use of four home care services (homemaking/cleaning, personal care, in-home nursing, home-delivered meals). Secondary data analysis from the 1991-1992 Manitoba Study on Health and Aging data set was conducted, using a sample of 124 community-dwelling elders and their unpaid caregivers. Multivariate analysis revealed that disruptive behaviors were significantly associated with the use of two services: personal care and home delivered meals. Being cognitively impaired with dementia was significant for only home-delivered meals. Overall functional status of the elder emerged as a consistent predictor. PMID- 11272855 TI - Critical incidents precipitating institutionalization of a relative with Alzheimer's. AB - Despite the stressors of caring for a relative with Alzheimer's disease (AD), families institutionalize their loved one only as a last resort. What constitutes this last resort? Predictors of and risks for institutionalization have been widely examined by researchers for two decades. To date, values underpinning the critical incident leading to institutionalization of a relative with AD have not been explored. The purpose of this secondary analysis of 20 interviews with family caregivers who had recently institutionalized their relative with AD was to examine underlying values that precipitated the move. Mitchell's definition, in 1983, of a critical incident was used as a guiding framework. Three major themes were identified: "I couldn't forgive myself if something happened," "It was ruining my life," and "I had no choice." The ability to identify underlying values precipitating critical events may help nurses plan interventions to assist increasing numbers of families faced with institutionalizing a relative with AD as our population ages. PMID- 11272856 TI - Modeling women's quality of life after cardiac events. AB - Quality of life (QOL) is presented as a global, unidimensional, and subjective assessment of one's life. This study examined the impact of perceived health status, hope, and optimism on QOL in 93 women after suffering a cardiac event. Construct validity was examined by estimating a model where QOL was measured with four indicators, and perceived health was measured with the SF-36 Health Survey. Hope was measured with the Herth Hope Index and dispositional optimism was measured with the Life Orientation Test. The unidimensionality of QOL and its response to health status, hope, and optimism were tested. Fit indices suggested that the theoretical relations posited were compatible with the data, (chi 2(42) = 44.125, p = .382, RMSEA = .0001, GFI = .942). The model explained 66% of the variance in QOL. Modeling suggested the presence of a complex latent concept composed of hope and optimism that influenced QOL. PMID- 11272857 TI - Pain management practices of nurses caring for older patients with osteoarthritis. AB - Osteoarthritis is a prevalent chronic illness in older people. Management of the ensuing pain is of critical importance in preventing disability and maintaining independence. This qualitative study explored the pain management techniques used by 10 RNs working in home health nursing. Four categories emerged from the interview data: knowing how to assess, knowing about pain treatments, trying but frustrated, and needing more knowledge. These categories were reduced into two constructs: Understanding Pain and Wanting to Provide Good Nursing Care. Clinical implications included supplementing pain management strategies by adding to the assessment base, expanding pain management techniques, and increasing knowledge about aging processes and pain control. PMID- 11272858 TI - Knowledge of and attitudes toward sex among Chinese adolescents. AB - This study was conducted to examine the knowledge of and attitudes toward sex of 178 Chinese secondary school students in Hong Kong. The data were collected using a questionnaire that comprised three parts: the Chinese version of the Mathtech Knowledge Test, the Chinese version of the Mathtech Attitude and Value Scale, and a demographic sheet seeking sociodemographic information. In general, students demonstrated a low level of sexual knowledge, especially in relation to adolescent marriage, the probability of pregnancy, and adolescent sexual activity. With regard to attitudes, students indicated positive attitudes toward importance of family and importance of birth control. Male students in comparison with their female counterparts had a higher level of agreement with premarital intercourse and the use of pressure and force in sexual activity. PMID- 11272859 TI - Meaningful nursing research. AB - Dr. Fawcett's editorial (Western Journal of Nursing Research, August 2000), staunchly criticizes research journals and researchers for their lack of attentiveness to nursing discipline-specific knowledge. She suggests that this lack of attentiveness posits nursing for extinction; in doing so, she raises fundamental questions about the relationship between nursing and nursing science. Despite a plethora of nursing literature, professional nursing remains an intangible concept for many nurses. The author addresses this issue from the perspective of her sense of self as a nurse. PMID- 11272861 TI - [Possibilities and limits of harvesting and using human embryonic stem cells]. AB - The availability of human embryonic stem cells as well as recent results about tissue specific stem cells residing in various organs of the human body have provided novel tools for basic sciences and medicine, which may result in a substantial increase of knowledge and the development of novel therapeutic concepts for the treatment of presently incurable diseases. Stem cell technologies, however, also raise fundamental ethical problems related to the sourcing and use of the cells. On the basis of a description of cultured stem cells derived from human embryos (ES cells), aborted human fetuses (EG cells), and tissues of the adult organism, respectively, the present article analyses ethical problems specifically related to each of these procedures in the context of the ethical principles underlying German law. PMID- 11272860 TI - [Psychotropic substances in the area of preclinical intensive care medicine]. AB - The aim of preclinical intensive care is to prevent severe consequences by providing immediate therapeutic aid to emergency patients in a life-threatening condition. We investigated how the abuse of drugs and alcohol influences the frequency and the category of emergency cases as well as the patient outcome. 250 emergency cases were analysed in a prospective study in the area of Heidelberg, Germany, from the summer of 1995 to the spring of 1999. The analysis is based on patient data obtained from the German standardised Emergency Record and on the clinical outcome and influence of drugs and alcohol. A serious level of drugs or alcohol was detected in 17% of the patients (the average level of alcohol was 1.97 promille). 81% of these intoxicated patients were given a NACA (National Advisory Committee for Aeronautics) score of less than 4 (not life-threatening) and therefore were not in need of the a presence of a physician. PMID- 11272862 TI - [Psychological and emotional factors of work experience as an issue for occupational medicine practice in the hospital]. AB - The consideration of psychomental and psychological load of employees is not part of the traditional working sphere of occupational physicians. Medical diagnostics and check-up are still regarded as the primary area of responsibility. This fact might be seen as a drawback with regard to the increasing importance of work related psychomental load on the one hand and the actual requirements for high quality occupational medicine on the other. The goal of a study in 64 occupational physicians in hospitals was to find out how occupational physicians estimate their own competence for dealing with psychomental problems of their clients, whether employees trust them, and which personal attitudes can be found towards the importance of work-related psychomental strain. The self-estimation of personal competence was--in contradiction to the results of former studies focusing on employees' views--surprisingly positive. A significant influence is the amount of work within the health-care sector compared to others. This might be interpreted as a selective effect and leads to the need for large-scale studies. Agreement could be found concerning the lack of chances for qualification in this sphere. Hence, further education and training in professional work with psychomental problems at the workplace has to be extended. PMID- 11272863 TI - [Organization of a fluoridation program with fluoride gel within the scope of a systematic quality assurance concept in dental health promotion]. AB - It is known from epidemiological studies that besides the instruction in oral hygiene and healthy nutrition the application of fluorides mainly contributes to improved oral health. In the district of Tuttlingen a fluoride varnish and a fluoride gel are used in oral prevention. The gel is applied every 14 days in collaboration with 207 parents and teachers. This cooperation should improve oral self-care and lead to an increased consciousness of preventive measures. The entire programmme is part of the concept of quality management in oral prevention in the district. Based on health reporting, sanitary priorities and targets are described with the period of time required for attaining the set targets. The achieved preventive measures are evaluated with regard to their effectiveness and efficiency for an optimised use of the available means and resources. PMID- 11272865 TI - [Kinetics of arsenic in human blood after a fish meal]. AB - It is well known that fish contains high amounts of arsenic (As) compounds (mean values per wet weight [mg x kg-1] Ballin [1]: 41; Falconer et al. [2]: 14; Staveland et al. [3]: 5.2), which are mainly represented by organic As compounds, especially by arsenobetaine. It is generally assumed that arsenobetaine is rapidly eliminated via the urine and therefore seems to be non-toxic for humans. However, the kinetics of arsenobetaine in human blood are unknown to date. Therefore, the following experiments were performed: 14 women of 24 to 32 years of age voluntarily ingested 179 to 292 g of cooked plaice fillet containing 44 (minimum) to 276 (maximum) mg As x kg-1 dry weight. Hence, the volunteers ingested 2.5 (minimum) to 20 (maximum) mg As per person, equivalent to 0.04 to 0.35 mg As x kg-1 body weight. The element As was measured by atomic absorption spectrometry using the graphite furnace technique in order to detect the total amount of As including that of the stable arsenobetaine. In the blood, the highest As values of 55 +/- 5.8 micrograms x L-1 (median) were found 2 hours after fish ingestion. Subsequently the As concentrations declined reaching 16 +/- 0.69 micrograms x L-1 (median) 48 hours after fish ingestion. In respect of the As values in blood recorded between 2 and 10 hours after fish ingestion, rapid elimination could be observed leading to a half-life of 7.1 hours (first value) recorded by linear regression analysis. With regard to the As values in blood between 10 and 48 hours after fish ingestion, a lower elimination rate was estimated with a longer half-life of 63 hours (second value). The reason for this delayed elimination of As is not known. The results indicate that As mainly absorbed as arsenonetaine due to ingestion of fish is not eliminated as fast as had been expected on the basis of published data. As long as it is not known what happens to arsenobetaine remaining for longer periods in the blood with a half life of 63 hours, caution is advised regarding the general opinion that arsenobetaine is rapidly eliminated and non-toxic for human consumption. PMID- 11272864 TI - [Evaluation of health policy intervention on the community level--the "Neighborhood Coordination of Health and Social Care" Model Project in North Rhine-Westphalia]. AB - Between October 1995 and December 1998 the pilot project 'Local Coordination of Health and Social Care' was conducted in 28 communities of the state of North Rhine-Westphalia. The project has been evaluated by two university research teams. The aim of the project was basically to establish new structures of health planning and coordination at the community level, in order to improve health reporting and health care as well as health promotion. To realize this aim round tables, working groups and project-offices were implemented in the communities. The evaluation was focused on the following question: What were the conditions (structures) and processes that influenced the project outcomes? Qualitative and quantitative methods were applied (interviews, standardised self-administered questionnaires, analyses of documents) to this end. Evaluation of structures showed that most communities succeeded in integrating relevant health policy actors into the newly created round tables and working groups. Working climate and achievements were evaluated favourably by most of the involved actors. All communities succeeded in developing and enacting recommendations for programmes, and about 40% of these programmes were implemented during the project. The probability of programme implementation was particularly high if the programme was based on reliable local data and if execution was effected only on the community level. The possibly beneficial effects on health care and welfare produced by the new programmes could not be assessed within the short project period. The paper concludes with a brief discussion of practical consequences for future health policy at community level. PMID- 11272866 TI - [Determining the need for medical rehabilitation services of employed members of the legal pension fund. A recommendation from social medicine and social legal viewpoints]. AB - Assessing health care needs in populations has become a major activity of public health medicine worldwide. Its methodology has been developing mainly in the English-speaking world. Concept, methods, and techniques have not yet reached Germany though recently the national expert advisory council for the concerted action in health care (Sachverstandigenrat fur die Konzertierte Aktion im Gesundheitswesen) provided first "official" definitions of demand, supply, and need to identify over- and undersupply in health care. This article aims at defining, from a combined sociolegal and sociomedical perspective, the need for medical rehabilitation measures among insurees of German pension funds. According to section 15 SGB VI rehabilitation is conceived as a medically coordinated multimodal-multidisciplinary intervention with a cognitive-behavioural orientation. To objectify the need for rehabilitation a series of 9 questions was developed enquiring inter alia about the presence of a disease or disability, the extent or "amplification" of the disorder, its course pattern, the implied risk of permanent work disability and likely success of rehabilitation. Nonspecific back pain served as a paradigmatic condition. One of the main problems encountered is the presently small evidence base to arrive at the necessary prognostic and therapeutic judgements. PMID- 11272867 TI - [Life expectancy and the need for nursing care in Germany]. AB - Effects of further gains in life expectancy on the health and autonomy of the elderly population are a matter of controversy. Health indicators, which integrate information on both mortality and morbidity could help to clarify whether the additional years are spent in health or disease. Since the introduction of a statutory long-term care insurance in Germany, national data on the prevalence of dependency are available. These data were used for the calculation of dependency-free life expectancy and life expectancy in a state of dependency according to Sullivans method. The calculations are based on 71.5 million insured at mid-year 1999 and on the period life-table for the years 1995/97. At the age of 65 the average duration of dependency is 15.4 months for men and 29.4 months for women. Men can expect to spend 91.4% of their remaining lifetime dependency-free, whereas the dependency-free proportion among women is only 86.9%. The distribution of severity grades of dependency is similar for both sexes. Women, however, spend 35.4% of the total duration of dependency in institutional care (10.4 months), men only 22.1% (3.5 months). Information from long-term care insurance appears suitable for the monitoring of time trends in the health of the elderly population and for projections of future needs for health and social services. PMID- 11272868 TI - Cardiac troponin I in patients with chest pain. PMID- 11272869 TI - Scoring systems in trauma. AB - BACKGROUND: Trauma is a major cause of mortality and morbidity worldwide. Methods of assessing outcome have evolved with management of trauma victims. RESULTS AND DISCUSSION: The wide variety of scoring instruments available to assess the injured patient may be divided into three groups: anatomical, physiological and combined systems. Anatomical systems depend on an accurate description of the injuries sustained. Physiological systems measure the effects of injury on the patient's physiological reserves. Combined systems contain elements of both anatomical and physiological scores. Prospectively, scoring systems help in description, triage, treatment decisions and estimating outcome. Retrospective scoring is helpful in audit, in quality control, in comparing treatment methods or centres, and in identifying unexpected outcomes. Limitations may be inherent in the system or may reflect inaccurate or incomplete data collection. PMID- 11272870 TI - The role of cardiac troponin I in determining the necessity for exercise electrocardiography in low risk patients with chest pain. AB - BACKGROUND: Assessment of non-cardiac chest pain places a considerable burden on healthcare resources. The current practice of serial electrocardiographs (ECGs), serum creatinine phosphokinase and by pre-discharge exercise electrocardiography gives an average in-hospital stay of 3.7 days. AIMS: This study assess the use of a sensitive assay for cardiac troponin I (cTnI) to identify a low risk group for whom exercise ECG may not be indicated. METHOD: Ninety-five patients with acute chest pain and with peak cTnI < 0.1 ng/ml and a non-diagnostic resting ECG were studied. Patients were divided into two groups. Group one had normal range cTnI (< 0.03 ng/ml). Group two had minimal elevation of cTnI (0.03-0.099 ng/ml). Average follow-up was 172 days. RESULTS: Nineteen patients had minimal elevation in cTnI of whom five developed significant ST shift on exercise and five had adverse events. No patient with a normal range cTnI had a positive stress test and none suffered an adverse event (p < 0.001). CONCLUSION: CTnI in the normal range can identify patients with acute chest pain who have a negligible event rate and for whom exercise electrocardiography is not required. PMID- 11272871 TI - The potential for drug interactions with statin therapy in Ireland. AB - BACKGROUND: Seven percent of acute hospital admissions result from adverse drug reactions, of which 25% are due to drug interactions. Adverse effects of statin drugs occur in 3% of patients, mainly due to co-prescribing with other lipid lowering agents or agents that alter their metabolism. AIM: The aim of this study was to investigate co-prescribing of the frequently-used statin medications with interacting drugs. METHODS: Data from the General Medical Services (GMS) scheme of the Eastern Health Board from January to December 1998 were used in this study. Using the coding index for statins, co-prescribing was identified when concomitant medications were administered under the same GMS claim number. RESULTS: Of 7,602 patients prescribed statins, co-prescribing of simvastatin, atorvastatin and fluvastatin with competing substrates or inhibitors of their metabolism occurred in 32, 26 and 13.4% of prescriptions issued. Thirty-four per cent of patients on simvastatin, 28% on atorvastatin and 16% on fluvastatin were prescribed medications with drug interaction potential. CONCLUSION: Co prescribing of statins with competing substrates or inhibitors of their metabolism occurred in up to one-third of prescriptions issued. When statins are co-prescribed with recognised inhibitors of drug metabolism, pravastatin, which does not undergo significant hepatic metabolism, is the statin of choice. PMID- 11272872 TI - Hepatitis C infection in an Irish antenatal population. AB - BACKGROUND: Hepatitis C infection (HCV) has an estimated seroprevalence of 1-2% in women of child-bearing age and vertical transmission rate of 5-15%. AIMS: To characterise the current trends of HCV in an Irish antenatal population. METHODS: Infants of HCV seropositive women, born 1994 to 1999, were referred to the Paediatric Infectious Diseases service. Maternal details were collected retrospectively. RESULTS: 296 HCV seropositive women were studied. 244 (82%) were infected through intravenous drug use (IVDU), 25 (8%) through heterosexual contact and 13 (7%) via blood products. Nine women had no identifiable risk factors. Coinfection with other blood borne viruses was uncommon (4.7% HIV, 3.4% hepatitis B). Of 84 women tested for HCV-RNA, 46 (55%) were positive. Eighty three (26%) delivered prematurely; the caesarean section rate was 11%. CONCLUSIONS: HCV is increasingly detected in antenatal clinics. Heterosexual contact is a mode of spread. Maternal HCV viraemia can be variable in pregnancy. Further study of HCV in pregnancy is needed to define the impact of pregnancy on HCV, accurately predict infant outcome and selectively target interventions to women at greatest risk of transmission. PMID- 11272873 TI - Has there been a turning point in the numbers of AIDS and HIV antibody positive cases in Ireland? AB - BACKGROUND: Major developments in the prevention and treatment of human immunodeficiency virus (HIV) infection and acquired immunodeficiency syndrome (AIDS) have taken place in recent years. Changes in the size of the HIV and AIDS epidemic need to be monitored to assess these developments and plan future services. AIMS: To describe temporal trends in the incidence of HIV and AIDS in Ireland, describing separately the major risk groups, and to explore possible associations between these trends with developments in care. METHODS: The annual numbers of HIV and AIDS in Ireland were analysed to determine whether there has been a turning point in incidence. RESULTS: For AIDS, there has been an overall decrease in numbers since 1993, with both homosexual and intravenous drug users (IDU) risk groups exhibiting a decrease. For HIV antibody positive individuals, overall numbers have remained constant. However, there has been an upward trend in the heterosexual risk group and a downward trend in the IDU risk group. Thus both AIDS and HIV numbers exhibit turning points. CONCLUSIONS: Declines in HIV and AIDS incidence in the homosexual and IDU risk groups are indirectly attributable to health safety and treatment programmes. The increase in HIV antibody positive cases among heterosexuals may reflect a relapse in safe sex behaviour. PMID- 11272874 TI - Comparison of basal broth media for the optimal laboratory recovery of Campylobacter jejuni and Campylobacter coli. AB - BACKGROUND: Campylobacter species are the most common known bacterial cause of acute diarrhoeal disease and incidences are increasing yearly in Ireland. Concern is expressed that this increase may be due to poor laboratory techniques in recognising these organisms as they are fastidious and technically demanding to culture. AIMS: To evaluate and compare the efficacy of seven commercially available basal broth media for the laboratory cultivation of thermophilic C. jejuni and C. coli isolated from foods and clinical specimens, as well as for optimal laboratory passage. METHODS: Four strains of Campylobacter spp., including two C. jejuni and two C. coli isolated from pigs were used to compare the quantitative growth of cells on seven commercially available basal culture broths. These included nutrient broth (NB), nutrient broth no. 2 (NB2), Mueller Hinton broth (MH), brucella broth (BB), brain heat infusion broth supplemented with 0.6% (w/v) yeast extract (BHIYE), tryptone soya broth supplemented with 0.6% (w/v) yeast extract (TSBYE) and Campylobacter broth (KAMPY) employing a spectrophotometrical growth assay model. RESULTS: Mean absorbance results of the broth media comparison for each strain and statistical analysis of variance allowed the basal media to be ranked into the order, based on proliferation of organisms: BHIYE > TSBYE > BB > NB2 > MHB > KAMPY > NB CONCLUSION: These studies support laboratory employment of either BHIYE or TSBYE for optimal recovery of C. jejuni and C. coli and avoidance of nutrient broth. PMID- 11272875 TI - Validation of serological tests for Helicobacter pylori infection in an Irish population. AB - BACKGROUND: Serological tests for Helicobacter pylori using laboratory and 'office' formats are commonly used, easy to perform, inexpensive and widely available. Local validation of test performance is required. AIMS: This study examined the performance of a laboratory and 'office' ELISA in a population of Irish dyspeptics presenting for endoscopy. METHODS: Consecutive patients presenting for endoscopy had blood drawn at sedation. Samples were analysed using two ELISA formats; a standard laboratory format and an 'office' ELISA test card. H. pylori infection was diagnosed by analysis of antral and corpus biopsies using the rapid urease test, culture and histology. A combination of two positive invasive tests was considered indicative of infection. RESULTS: The sensitivity and specificity of laboratory ELISA was 82.4% and 85% respectively while the values for the 'office' ELISA were 87.7% and 85.7% respectively. In patients under 45 years sensitivities and specificities of the 'office' test exceeded 90%. The two serological tests agreed in 87.5% of subjects. CONCLUSIONS: Both tests performed satisfactorily. However, indeterminate results impaired the usefulness of the laboratory ELISA particularly when using a new cut-off. The 'office' ELISA performed particularly well in young patients. A simpler test using antigens from locally prevalent strains to optimise accuracy is awaited. PMID- 11272876 TI - The Short Form 36 (SF-36) Health Survey: normative data for the Irish population. AB - BACKGROUND: Generic measures of quality of life have a wide application in health research. They measure disease impact by comparing scores in patient groups with a healthy population. They also facilitate comparative studies between different patient groups. The SF-36 Health Survey quantifies respondents' perceptions of their functioning in eight dimensions of daily life. AIM: The aim of this study was to set normative values for the SF-36 in the Irish population aged 18 years and over. METHOD: A random sample of 800 subjects was drawn from the electoral register using the RANSAM method of sampling. RESULTS: Two hundred and ninety five (37%) valid questionnaires were returned for analysis. The SF-36 was found to have acceptable internal consistency and validity. Normative values for the total population are presented, in addition to results for males and females across seven age groups. Ageing was associated with a decline in the physical dimensions of health. CONCLUSIONS: There was no evidence to suggest that there were significant differences in health status between males and females, or between this Irish sample and the published norms for the US population. PMID- 11272878 TI - Aggressive management leads to improved survival in patients with small cell lung carcinoma. AB - BACKGROUND: Small cell lung carcinoma (SCLC) accounts for 17-25% of all cases of lung cancer, and remains the most lethal form of this disease. AIMS: We sought to determine whether an aggressive treatment policy led to an increase in median survival in patients with SCLC in our institution. METHODS: From 1985 to 1993, patients with SCLC were often treated conservatively on the basis of advanced age or poor performance status. From 1993 to 1998, a more aggressive management policy was adopted. All patients were treated with chemotherapy. Radiotherapy was administered, where appropriate, following the completion of chemotherapy. The medical records of 66 patients were analysed and clinical outcomes were compared. RESULTS: Median survival in the 1993-98 group (332 days) was significantly better compared to the 1985-93 group (194 days) (p = 0.02). In patients with limited disease, median survival in the 1993-98 group (489 days) was also significantly better compared to the 1985-93 group (254 days) (p = 0.04). The difference in median survival in extensive disease was not significant (p = 0.09). CONCLUSIONS: The presented data suggest that appropriate aggressive management of patients with SCLC leads to a significant increase in median survival. This survival benefit is most apparent in patients with limited disease. PMID- 11272877 TI - Recurrent pleomorphic adenoma: uninodular versus multinodular disease. AB - BACKGROUND: While treatment of previously untreated pleomorphic adenoma is relatively straightforward, recurrent pleomorphic adenoma presents a management problem with increased risk of injury to the facial nerve and an increased risk of malignant transformation in recurrent disease. AIMS: The objectives of this study were to review the management of recurrent pleomorphic adenoma in our unit to identify factors that might help treatment of future cases. METHODS: We reviewed the management of pleomorphic adenoma at our department over an eight year period from 1990-1998 and present our experience of recurrent pleomorphic adenoma of the parotid gland and parapharyngeal space. RESULTS: Twelve patients were treated with recurrent pleomorphic adenoma. In 10 of these, the site of recurrence was in the parotid gland with the remainder occurring in the parapharyngeal space. Type of recurrence was uninodular or multinodular, the former being easier to treat. Three patients required adjuvant radiotherapy. None of our patients had permanent facial nerve damage. CONCLUSIONS: In order to prevent recurrence of pleomorphic adenoma of the parotid gland, we recommend formal superficial parotidectomy for first time surgery. PMID- 11272879 TI - Rupture of the male membranous urethra. AB - BACKGROUND: Management of traumatic rupture of the male membranous urethra remains controversial. Long-term morbidity can include urinary incontinence, urethral stricture and erectile dysfunction. AIMS: To review management and outcome of urethral rupture to improve treatment protocols. METHODS: A retrospective study of 47 patients presenting with traumatic urethral rupture over 25 years was performed. RESULTS: All patients underwent emergency suprapubic catheterisation, 32 patients had open surgical realignment at 1-2 weeks; 78% of whom developed strictures. Ten patients unsuitable for early repair underwent delayed transabdominal transpubic urethroplasty at three months: 40% of whom developed strictures. Five patients with partial rupture were managed by cystoscopy and urethral catheter. Erectile dysfunction correlated to initial injury rather than treatment. CONCLUSIONS: If the patient is stable and requires emergency laparotomy for other abdominal injuries, he should have immediate realignment of the urethra. Early realignment of the urethra at laparotomy at 1-2 weeks can be combined with orthopaedic fixation of pelvic fractures. Patients who remain unstable due to associated injuries should have delayed urethroplasty at three months. PMID- 11272880 TI - Universal precautions--do Irish anaesthetists comply? AB - BACKGROUND: Anaesthetists are at high risk from blood-borne pathogens. Universal Precautions (UP) include the routine use of appropriate barrier precautions and techniques to reduce the likelihood of exposure to blood, body fluids and tissues that may contain pathogens. The compliance of Irish anaesthetists with these precautions has not been studied. AIM: To study the attitudes of Irish anaesthetists to Universal Precautions. METHOD: A postal questionnaire was sent to 210 anaesthetists currently practising in Ireland. The questionnaire was based on a model used in Australia and New Zealand. RESULTS: There was a 50% response rate to the survey. Only 15% of respondents had taken a risk history from a patient in the preceding four weeks. Resheathing of needles was commonplace. The effectiveness of hepatitis B immunisation was rarely checked and only 66% of respondents believe implementation of Universal Precautions to be practical. CONCLUSION: Irish anaesthetists comply poorly with Universal Precautions. PMID- 11272881 TI - Diagnostic advances in acute mesenteric ischaemia. PMID- 11272882 TI - Medical writing in Irish. PMID- 11272883 TI - The medical sciences in twentieth-century Ireland. PMID- 11272884 TI - Richard Carmichael 1779-1849. PMID- 11272885 TI - Isolated resectable pancreatic metastasis 10 years post gastrectomy. PMID- 11272886 TI - Impacted cancellous allograft in the proximal femur: early histological observations. PMID- 11272887 TI - Management of head injury in a regional hospital. PMID- 11272888 TI - Finding a workable balance: regulation of genetic testing in the human genome era. PMID- 11272890 TI - A comparative study of fibrous dysplasia and osteofibrous dysplasia with regard to Gsalpha mutation at the Arg201 codon: polymerase chain reaction-restriction fragment length polymorphism analysis of paraffin-embedded tissues. AB - Fibrous dysplasia and osteofibrous dysplasia are both benign fibro-osseous lesions of the bone and are generally seen during childhood or adolescence. Histologically, the features of these bone lesions sometimes look quite similar, but their precise nature remains controversial. Mutation of the alpha subunit of signal-transducing G proteins (Gsalpha), with an increase in cyclic adenosine monophosphate (cAMP) formation, has been implicated in the development of multiple endocrinopathies of the Albright-McCune syndrome and in the development of fibrous dysplasia. We studied Gsalpha mutation at the Arg201. codon in seven cases of fibrous dysplasia (six monostotic lesions and one polyostotic lesion) and seven cases of osteofibrous dysplasia using formalin-fixed, paraffin-embedded tissue, by means of polymerase chain reaction-restriction fragment length polymorphism and direct sequencing analysis. All of the seven cases of fibrous dysplasia showed missense point mutations in Gsalpha at the Arg201 codon that resulted in Arg-to-His substitution in three cases and Arg-to-Cys substitution in four cases. On the other hand, the seven cases of osteofibrous dysplasia and the normal bone used as a control showed no such mutation. These data suggest that fibrous dysplasia and osteofibrous dysplasia have different pathogeneses and that the detection of Gsalpha mutation at the Arg201 codon is quite useful for distinguishing between these lesions. PMID- 11272889 TI - Molecular profiling of clinical tissues specimens: feasibility and applications. PMID- 11272891 TI - Evidence by spectral karyotyping that 8q11.2 is nonrandomly involved in lipoblastoma. AB - We report two cases of lipoblastoma with chromosome 8-related aberrations, ie, a 92,XXYY,t(7;8Xp22;q11.2)x2 [8]/46,XY[16] in Case 1 and a 46,XY,-8,-13,add(16) (q22),+mar, +r [cp13]/46,XY[7] in Case 2. Using spectral karyotyping and fluorescence in situ hybridization techniques, the karyotype of Case 2 was redesignated as 46,XY, r(8), del(13)(q12), der(16)ins(16;8)(q22; q24q11.2)[cp13]/46,XY[7]. This report delineates a new chromosome rearrangement, ie, der(16)ins(16;8)(q22; q24q11.2) in lipoblastoma, and also confirms the t(7; 8)(p22;q11.2), reported only once previously, as a recurrent translocation involved in such a tumor. These findings provide valuable information for clinical molecular cytogenetic diagnosis of lipoblastoma. Furthermore, this report highlights the value of cytogenetic and molecular cytogenetic analysis in differential diagnosis of childhood adipose tissue tumors and adds to the number of lipoblastomas reported with chromosomal abnormalities at 8q11.2. PMID- 11272892 TI - Concomitant oncoprotein detection with fluorescence in situ hybridization (CODFISH): a fluorescence-based assay enabling simultaneous visualization of gene amplification and encoded protein expression. AB - We sought the validation of a three-color fluorescence-based system that simultaneously profiles Her2/neu oncogene copy by fluorescence in situ hybridization (FISH) and Her-2/neu encoded protein by the use of a versatile alkaline phosphatase chromogen fast red K in either fluorescence or bright-field mode. Nuclei were counterstained with DAPI. Nineteen infiltrating ductal carcinomas of breast were comprehensively evaluated for Her-2/neu amplification/overexpression by direct and indirect FISH using digoxigenin (DigFISH) and direct fluorescently labeled probes, autoradiographic RNA:RNA in situ hybridization, and immunohistochemistry using monoclonal antibody CB11. CODFISH results correlated well with DigFISH, direct-label FISH, mRNA expression, and oncoprotein expression as assessed with CB11, and enabled simultaneous visualization of gene copy and protein. In addition, qualitative immunohistochemistry may be followed by CODFISH gene copy enumeration to clarify ambiguous cases. PMID- 11272894 TI - PCR analysis of the immunoglobulin heavy chain gene in polyclonal processes can yield pseudoclonal bands as an artifact of low B cell number. AB - Polymerase chain reaction (PCR)-based analysis for detecting immunoglobulin heavy chain gene (IgH) rearrangements in lymphoproliferative disorders is well established. The presence of one or two discrete bands is interpreted as a monoclonal proliferation, whereas a smear pattern represents a polyclonal population. Prompted by our observation of discrete bands in histologically reactive processes with a relative paucity of B cells, we sought to determine whether low numbers of B cells in biopsy specimens could artifactually produce pseudomonoclonal bands. We performed IgH PCR analysis on serially diluted DNA samples from 5 B cell non-Hodgkin's lymphomas (B-NHLs), 5 reactive lymph nodes, 5 reactive tonsils and 10 microdissected germinal centers from a lymph node with follicular hyperplasia. We also assessed multiple aliquots of DNA samples from small biopsy specimens of reactive lymphocytic processes from the stomach (5 cases). PCR products were evaluated using high resolution agarose or polyacrylamide gels, and DNA sequencing was performed on IgH PCR products from two reactive germinal centers, which yielded monoclonal bands of identical size. All 5 B-NHLs harboring monoclonal B cell populations yielded single discrete bands, which were maintained in all dilutions. By contrast, all of the reactive lesions with polyclonal patterns at 50 ng/microl starting template concentration showed strong pseudomonoclonal bands at dilutions of 1:1,000 to 1:1,500 in placental DNA. Two of the microdissected reactive germinal centers that showed bands of identical size on duplicate reactions were proven to have different IgH sequences by sequencing. We conclude that specimens containing low numbers of polyclonal B cells may produce pseudomonoclonal bands on IgH PCR analysis. IgH PCR analysis should be performed on multiple aliquots of each DNA sample, and only samples that yield reproducible bands of identical size can be reliably interpreted as monoclonal. PMID- 11272893 TI - Quantitative mRNA expression analysis from formalin-fixed, paraffin-embedded tissues using 5' nuclease quantitative reverse transcription-polymerase chain reaction. AB - Analysis of gene expression and correlation with clinical parameters has the potential to become an important factor in therapeutic decision making. The ability to analyze gene expression in archived tissues, for which clinical followup is already available, will greatly facilitate research in this area. A major obstacle to this approach, however, has been the uncertainty about whether gene expression analyses from routinely archived tissues accurately reflect expression before fixation. In the present study we have optimized the RNA isolation and reverse transcription steps for quantitative reverse transcription polymerase chain reaction (RT-PCR) on archival material. Using tissue taken directly from the operating room, mRNAs with half-lives from 10 minutes to >8 hours were isolated and reverse transcribed. Subsequent real-time quantitative PCR methodology (TaqMan) on these cDNAs gives a measurement of gene expression in the fixed tissues comparable to that in the fresh tissue. In addition, we simulated routine pathology handling and demonstrate that this method of mRNA quantitation is insensitive to pre-fixation times (time from excision to fixation) of up to 12 hours. Therefore, it should be feasible to analyze gene expression in archived tissues where tissue collection procedures are largely unknown. PMID- 11272895 TI - A multi-site study for detection of the factor V (Leiden) mutation from genomic DNA using a homogeneous invader microtiter plate fluorescence resonance energy transfer (FRET) assay. AB - The goal of this multicenter study was to evaluate the second-generation Invader technology for detecting the factor V (Leiden) mutation directly from genomic DNA of different sample types. Invader assay results were compared with polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) or allele specific PCR (AS-PCR) analysis. The Invader assay is a PCR-independent methodology that uses a microtiter plate format. In the assay, a specific upstream Invader oligonucleotide and a downstream probe hybridize in tandem to a complementary DNA template and form a partially overlapping structure. The Cleavase VIII enzyme recognizes and cuts this structure to release the 5' flap of the probe. This flap then serves as an Invader oligonucleotide to direct cleavage of a fluorescence resonance energy transfer (FRET) probe in a second invasive cleavage reaction. Cleavage of this FRET probe results in the generation of a fluorescent signal. The results of the Invader assay were 99.5% concordant with the PCR-based methods. Of the 372 samples tested once, only two gave discordant results (one from operator error and one from unknown causes), but were concordant on retesting. These results indicate that a simple microtiter plate based Invader assay can reliably genotype clinical patient samples for the factor V (Leiden) point mutation directly from genomic DNA without prior target amplification. PMID- 11272896 TI - New challenges for biomedical research utilizing human biological materials. PMID- 11272897 TI - Hepatosplenic and subcutaneous panniculitis-like gamma/delta T cell lymphomas are derived from different Vdelta subsets of gamma/delta T lymphocytes. AB - Gamma/delta T cell lymphomas (gamma/delta TCL) represent rare, often aggressive types of T cell malignancy that are clinically and pathologically diverse. Most gamma/delta TCL occur as a hepatosplenic or subcutaneous type. To date, analysis of the T cell receptor delta (TCRS) gene repertoire of hepatosplenic gamma/delta TCL (gamma/delta HSTCL) and subcutaneous panniculitis-like gamma/delta TCL (gamma/delta SPTCL) has been reported only in a limited number of cases. In this study we analyzed 11 gamma/delta HSTCL and 4 gamma/delta SPTCL by polymerase chain reaction and immunostaining to determine their usage of the Vdelta subtypes (Vdelta1-6). It is noteworthy that 10 of 11 gamma/delta HSTCL expressed the Vdelta1 gene. The remaining case also expressed T cell receptor delta (TCRS) as determined by flow cytometry and TCRdelta rearrangement in Southern blot. However, the Vdelta gene expressed by this lymphoma could not be determined, which suggests usage of an as yet unidentified Vdelta gene. In striking contrast to the gamma/delta HSTCL, all 4 gamma/delta SPTCL expressed the Vdelta2 gene. Our data demonstrate that gamma/delta HSTCL are preferentially derived from the Vdelta1 subset of gamma/delta T lymphocytes, whereas gamma/delta SPTCL are preferentially derived from the Vdelta2 subset. The pattern of Vdelta gene expression in HSTCL and SPTCL corresponds to the respective, predominant gamma/delta T cell subsets normally found in the spleen and skin. This finding suggests that gamma/delta TCL are derived from normal gamma/delta T lymphocytes which reside in the affected tissues. Furthermore, the selective, lymphoma type specific Vdelta gene segment usage may provide a molecular tool to distinguish better among various types of gamma/delta TCL lymphoma particularly in the clinically advanced, widely disseminated cases. PMID- 11272898 TI - Detection of microsatellite instability by fluorescence multiplex polymerase chain reaction. AB - We have created a clinical molecular diagnostic assay to test for microsatellite instability (MSI) at multiple loci simultaneously in paraffin-embedded surgical pathology colon resection specimens. This fluorescent multiplex polymerase chain reaction (PCR) assay analyzes the five primary microsatellite loci recommended at the 1997 National Cancer Institute-sponsored conference on MSI for the identification of MSI or replication errors in colorectal cancer: Bat-25, Bat-26, D2S123, D5S346, and D17S250. Amplicon detection is accomplished by capillary electrophoresis using the ABI 310 Genetic Analyzer. Assay validation compared 18 specimens previously assessed by radioactive PCR and polyacrylamide gel electrophoresis detection to results generated by the reported assay. Germline and tumor DNA samples were amplified in separate multiplex PCR reactions, sized in separate capillary electrophoresis runs, and compared directly to identify novel length alleles in tumor tissue. A concordance of 100% between the two modalities was achieved. The multiplex assay routinely detected a subpopulation of 10% tumor alleles in the presence of 90% normal alleles. A novel statistical model was generated that corroborates the validity of using results generated by analysis of five independent microsatellites to achieve a single overall MSI diagnosis. The assay presented is superior to standard radioactive monoplex PCR, polyacrylamide gel electrophoretic analysis, primarily due to the multiplex PCR format. PMID- 11272899 TI - Multiplex genotype analysis of invasive carcinoma and accompanying proliferative lesions microdissected from breast tissue. AB - To understand the genetic basis of breast cancer in a comprehensive way, purported precursor lesions need to be analyzed at a large number of genetic marker loci and compared with each other and with the invasive components. However, the microscopic size of most of these lesions and the very small amount of material that can be obtained through microdissection limit the number of loci that can be included in the analysis. To address this issue, a multiplex genotyping approach has been developed. With this approach, polymorphic sequences at 28 marker loci were amplified simultaneously from the micro-dissected components in 5-microm paraffin-embedded breast tissue sections. The genotypes of the lesions were determined after resolving the amplified allelic products by denaturing gradient gel electrophoresis. Because the material isolated from each lesion in a single 5-microm section was sufficient for several 28-locus assays and several successive tissue sections with the same set of lesions may be prepared, it is possible to determine the genotype of each lesion at hundreds of genetic marker loci that may well cover the human genome. Analyzing a sufficient number of cases may yield information that could be used to understand the genetic basis of breast cancer development in a comprehensive way. PMID- 11272900 TI - Clinical categories of neuroblastoma are associated with different patterns of loss of heterozygosity on chromosome arm 1p. AB - Deletion of the short arm of chromosome 1 is frequently observed in neuroblastoma (NB). We performed loss of heterozygosity (LOH) analysis of 120 well characterized NB to better define specific regions of 1p loss and any association with clinical and biological prognostic features (DNA index, MYCN, age, and stage). All categories of disease were represented including 7 ganglioneuromas, 8 stage 4S, 33 local-regional (stages 1, 2, and 3), and 72 stage 4 NB according to the International Neuroblastoma Staging System. Patients were consistently treated with stage-appropriate protocols at a single institution. Sixteen highly informative, polymorphic loci mapping to chromosome 1 were evaluated using a sensitive, semi-automated, fluorescent detection system. Chromosome arm 1p deletions were detected in all categories of tumor except ganglioneuroma. Frequent LOH was detected at two separate regions of 1p and distinct patterns of losses were associated with individual clinical/biological categories. Clinically aggressive stage 4 tumors were predominantly diploid with extensive LOH frequently detected in the region of 1ptel to 1p35 (55%) and at 1p22 (56%). The shortest region of overlap for LOH at 1p36 was between D1S548 and D1S1592 and for 1p22 was between D1S1618 and D1S2766. Local-regional tumors were mostly hyperdiploid with short regions of loss primarily involving terminal regions of 1p36 (42%). Most spontaneously regressing stage 4S tumors (7/8) were hyperdiploid without loss of 1p36 or 1p22. These findings suggest that genes located on at least two separate regions of chromosome arm 1p play a significant role in the biology of NB and that distinct patterns of 1p LOH occur in individual clinical/biological categories. PMID- 11272902 TI - Detection of clonally restricted immunoglobulin heavy chain gene rearrangements in normal and lesional skin: analysis of the B cell component of the skin associated lymphoid tissue and implications for the molecular diagnosis of cutaneous B cell lymphomas. AB - A monoclonal B cell population is the hallmark of B cell neoplasms including cutaneous B cell lymphomas (CBCLs). We modified and tested several polymerase chain reaction (PCR)-based assays involving amplification of immunoglobulin heavy chain (IgH) gene rearrangements to optimize assays specifically for cutaneous lymphoid infiltrates. We achieved greatest sensitivity with an assay employing IgH consensus primers complementary to the framework 3 portion of the upstream variable region and the downstream joining region. We studied 12 CBCLs, 6 nodal lymphomas and 7 cell lines. In 17/25 of these B cell neoplasms (84%), we detected one or two dominant bands, consistent with one or both IgH alleles being rearranged in the neoplastic B cell clone. As expected, IgH PCR assays produced diffuse smears in agarose gels or complex ladders in polyacrylamide gels when polyclonal B cell controls (blood and tonsil) were analyzed. However, in normal skin and non-CBCL skin lesions, one or a small number of discrete bands were sometimes detected. In certain cases, this made it difficult to distinguish true positives (monoclonal CBCL) from false positives (clonally restricted benign B cells). Correlation with immunophenotyping confirmed that false positive results were confined to samples with sparse or immunohistologically undetectable B cell infiltrates. Pseudoclonal bands showed variable sizes in repeat PCR reactions and could be distinguished from monoclonal bands by polyacrylamide gel electrophoresis of pooled triplicate PCR products. These findings suggest that molecular diagnosis using IgH PCR assays is best suited for B-cell-rich infiltrates, and can be problematic when applied to suspected T-cell-rich CBCLs, cutaneous T cell lymphomas, or other lesions containing only few B cells unless one is cognizant of the potential pitfalls. Furthermore, these results demonstrate the presence of rare B cells in normal skin and immunohistologically defined cutaneous T cell infiltrates. This correlates with recent reports of sparse B cells within the lymph draining from normal skin and may represent molecular evidence for a trafficking B cell component of the skin-associated lymphoid tissue (SALT). It also suggests a candidate B cell subset for the pathogenesis of cutaneous lymphoid hyperplasia and CBCLs. PMID- 11272901 TI - COL1A1-PDGFB fusion transcripts in fibrosarcomatous areas of six dermatofibrosarcomas protuberans. AB - The fibrosarcomatous transformation of dermatofibrosarcoma protuberans (DFSP) has been considered for some time to be associated with an adverse clinical outcome. However, the molecular and cellular mechanism underlying the tumor progression remains undetermined. As the chimeric gene, COL1A1-PDGFB, has been proposed to play an important role in the histogenesis of DFSP, we conducted a reverse transcription-polymerase chain reaction assay to ascertain whether the COL1A1 PDGFB fusion transcripts can be detected in both conventional DFSP and fibrosarcomatous components of DFSP with fibrosarcomatous areas (DFSP-FS), using a simple method of microdissection on sections of archival formalin-fixed, paraffin-embedded tumor specimens from six DFSP-FS cases. The COL1A1-PDGFB fusion transcripts could be detected in FS areas in five of the six cases, whereas conventional DFSP areas of all cases expressed the chimeric mRNA. A subsequent sequence analysis of the polymerase chain reaction products confirmed that the detected messages were derived from identical gene fusions in the two different components of each of the five cases. Our results verify that the COL1A1-PDGFB fusion transcripts are preserved in the FS areas of most DFSP-FSs. The expression of the fusion transcripts in both conventional DFSP and FS areas of DFSP-FS supports a common histogenesis of the two components. PMID- 11272903 TI - 1999 Association for Molecular Pathology annual meeting. PMID- 11272904 TI - Cold-temperature plastic resin embedding of liver for DNA- and RNA-based genotyping. AB - The standard practice of tissue fixation in 10% formalin followed by embedding in paraffin wax preserves cellular morphology at the expense of availability and quality of DNA and RNA. The negative effect on cellular constituents results from a combination of extensive cross-linking and strand scission of DNA, RNA, and proteins induced by formaldehyde as well as RNA loss secondary to ubiquitous RNase activity and negative effects of high temperature exposure during paraffin melting, microscopic section collection, and tissue adherence to glass slides. An effective strategy to correlate cellular phenotype with molecular genotype involves microdissection of tissue sections based on specific histopathological features followed by genotyping of minute representative samples for specific underlying molecular alterations. Currently, this approach is limited to short length polymerase chain reaction amplification (<250 bp) of DNA, due to the negative effects of standard tissue fixation and processing. To overcome this obstacle and permit both cellular morphology and nucleic acid content to be preserved to the fullest extent, we instituted a system of cold-temperature plastic resin embedding based on the use of the water-miscible methyl methacrylate polymer known as Immunobed (Polysciences, Warminster, PA). The system is simple, easy to adapt to clinical practice, and cost-effective. Immunobed tissue sections demonstrate a cellular appearance equivalent or even superior to that of standard tissue sections. Moreover, thin sectioning (0.5-1.0 microm thickness) renders ultrastructural evaluation feasible on plastic-embedded blocks. Tissue microdissection is readily performed, yielding high levels of long DNA and RNA for genomic and transcription-based correlative molecular analysis. We recommend the use of Immunobed or similar products for use in molecular anatomical pathology. PMID- 11272906 TI - Molecular genetic pathology: coming of age in the molecular world. PMID- 11272907 TI - Quantitative fluorescence in situ hybridization in lung cancer as a diagnostic marker. AB - The diagnosis of lung cancer is quite often hampered by the existence of various cell types within samples such as biopsies or pleural effusions. We have established a new marker for image cytometry of interphase tumor cells of the lung by using the most recurrent and early cytogenetic event in lung cancer, the loss of the short arm of chromosome 3. The method is based on the detection of the imbalance between the long and the short arms of chromosome 3 by performing two-color fluorescence in situ hybridization on both arms. Fourteen tumors were analyzed after short-term culture and compared with the corresponding cytogenetic data obtained from metaphase analysis. Results on interphase nuclei and control experiments on metaphases were the same, with imbalance ratios ranging from 1.0 to 2.0 (mean value 1.6, median 1.5). To assess the clinical significance of this approach, three pleural effusions were analyzed. Data showed that normal cells within the sample could have been distinguished from the tumor cells based on different imbalance values between the long and the short arms. Thus, our method allows refined detection of lung tumor cells within samples containing heterogeneous cell populations. PMID- 11272905 TI - A comparison of MyoD1 and fetal acetylcholine receptor expression in childhood tumors and normal tissues: implications for the molecular diagnosis of minimal disease in rhabdomyosarcomas. AB - Detection of minimal residual disease or micrometastases in rhabdomyosarcoma (RMS) has been an unresolved problem in 70 to 80% of RMS patients. In patients with alveolar type RMS, which harbors chromosomal translocations and produces tumor-specific fusion products, polymerase chain reaction (PCR)-based diagnosis is clear-cut. In the more frequent embryonal RMS, however, no such PCR-based marker has been described. Recently it has been suggested that the PCR-based detection of MyoD1 may be a valuable adjunct in the diagnosis of minimal disease in embryonal RMS. We report here that MyoD1 mRNA is not specific for RMS, but can be amplified from ex vivo samples of many other childhood tumors and some normal tissues. By contrast, simultaneous amplification of alpha and gamma subunit message of the fetal type acetylcholine receptor (AChR), by a novel duplex PCR, and the quantification of both transcripts resulting in a alpha/gammaAChR ratio <1 was 100% sensitive in alveolar (n = 8) and embryonal (n = 10) RMS. Moreover, gammaAChR was not detected in other childhood (n = 27) or adult tumors (n = 12), or normal tissues, except thymus. The high sensitivity and specificity of the method were confirmed by the successful detection of five cases of cytologically or molecularly verified RMS bone marrow micrometastases among 47 bone marrow samples from childhood tumor patients. By contrast, MyoD1 showed no amplification because of its low level of transcription. We conclude that mRNA of the fetal type AChR is a more specific and (about 100 times) more sensitive marker for the molecular detection of RMS than MyoD1, and thus appears to be a promising candidate for the detection of minimal disease in RMS lacking tumor-specific translocations. PMID- 11272908 TI - Goals and objectives for molecular pathology education in residency programs. The Association for Molecular Pathology Training and Education Committee. AB - Increasing knowledge of the molecular basis of disease and advances in technology for analyzing nucleic acids and gene products are changing pathology practice. The explosion of information regarding inherited susceptibility to disease is an important aspect of this transformation. Pathology residency programs are incorporating molecular pathology education into their curricula to prepare newly trained pathologists for the future, yet little guidance has been available regarding the important components of molecular pathology training. We present general goals for pathology training programs for molecular pathology education. These include recommendations to pathology residents for the acquisition of both basic knowledge in human genetics and molecular biology and specific skills relevant to microbiology, molecular oncology, genetics, histocompatibility, and identity determination. The importance of residents gaining facility in integrating data gained via nucleic acid based-technology with other laboratory and clinical information available in the care of patients is emphasized. PMID- 11272909 TI - QSAR approach for the selection of congeneric compounds with a similar toxicological mode of action. AB - The selection of compounds with a similar toxicological mode of action is a key problem in the study of chemical mixtures. In this paper, an approach for the selection of chemicals with similar mode of action, based on the analysis of structural similarities by means of QSAR and chemometric methods, is described. As a first step, a complete representation of chemical structures for examined chemicals (phenylureas and triazines) by different sets of molecular descriptors allows a preliminary exploration of similarity using multi-dimensional scaling (MDS). The use of genetic algorithm (GA) to select the most relevant molecular descriptors in modeling toxicity data makes it possible to develop predictive toxicity models. The final step is a similarity analysis, based again on MDS, using selected molecular descriptors, really relevant in describing the toxicological effect. PMID- 11272910 TI - Development of QSARs to investigate the bacterial toxicity and biotransformation potential of aromatic heterocylic compounds. AB - A series of aromatic heterocyclic and hydrocarbon compounds were tested for toxicity and biotransformation potential against two contrasting lux-marked whole cell microbial biosensors. Toxicity was determined by inhibition of light output of a Pseudomonas fluorescens construct that expresses lux constitutively. Biotransformation was tested by increase in light output of P. fluorescens HK44 (pUTK21), which expresses lux when in the presence of a metabolic intermediate (salicylate). The data were then modelled against physical/chemical properties of the compounds tested to see if quantitative structure-activity relationships (QSARs) could be derived. Toxicity was found to be accurately predicted by log Kow (R2 = 0.95, Q2 = 0.88), with the basic (pyridine-ring containing) heterocycles modelled separately. The biotransformation data were best modelled using lowest unoccupied molecular orbital (LUMO) energies (R2 = 0.90, Q2 = 0.87). PMID- 11272911 TI - Influence of soil moisture on the sequestration of organic compounds in soil. AB - A study was conducted as a part of continuing investigation of the effect of soil moisture on the sequestration of organic compounds aged in the soil. Here, experiments focused on the effects of moisture changes within the soil before, during, and after contaminant addition. The extractability of aged (68 d) phenanthrene was greater from soil that had been subjected to wetting and drying cycles prior to solute addition as compared to soil initially maintained at constant moisture. The recovery of phenanthrene added to moist soil was increased relative to extractability from soil that was air-dried at the time of the contaminant addition. Repeated wetting and drying of soil after the addition of atrazine or phenanthrene resulted in decreased extractability of the compounds as compared to samples maintained at constant moisture. A method for rapidly sequestering contaminants is proposed and may be useful in limiting the time required for laboratory studies involving "aged" contaminants. These data build upon the findings of earlier work from our laboratory and indicate that changes in the moisture conditions of soil can affect the availability of sequestered contaminants possibly through alterations in the structure of the natural solid. PMID- 11272912 TI - Prediction of partition coefficient and toxicity for benzaldehyde compounds by their capacity factors and various molecular descriptors. AB - The log Kow and log Sw values of 14 substituted benzaldehyde compounds were determined by the shake-flask method. Acute toxicities of 14 substituted benzaldehyde compounds to Daphnia magna were recorded. Their capacity factors (k') were determined by reversed phased high-performance liquid chromatography (RP-HPLC) on C18 column and methanol-water eluent. Molecular connectivity indices, the linear solvation energy relationships (LSER) parameters, and quantum chemical parameters were calculated for the tested chemicals and used to develop quantitative structure-retention relationship (QSRR) and quantitative structure property/activity relationship (QSPR/QSAR). Results demonstrated that the molecular connectivity indices, LSER parameters, and quantum chemical parameters could be used to predict the k' for compounds studied, LSER method was more accurate. The results also show that chromatographic retention data, log k', can be used to predict log Kow and log Sw for tested compounds. The log k'w can be directly utilized as hydrophobic descriptors to predict the toxicity to D. Magna for benzaldehyde compounds. PMID- 11272913 TI - The use of bioluminescent biotests for study of natural and laboratory aquatic ecosystems. AB - A set of bioluminescent tests was developed to monitor water quality in natural and laboratory ecosystems. It consisted of four bioluminescent systems: luminous bacteria, coupled enzyme system NADH:FMN-oxidoreductase-luciferase and triplet enzyme systems with alcohol dehydrogenase and trypsin. The set of biotests was applied for a small forest pond (Siberia, Russia), laboratory microecosystems polluted with benzoquinone and a batch culture of blue-green algae. Thereby effects of natural water compared to those of models of heavy pollution and "bloom" of blue-greens on the bioluminescent tests were revealed. The set of biotests was not affected by a natural seasonal variability of water quality in the unpolluted pond, but responded to the heavy pollution and the "bloom" of blue greens. The set of biotests could be recommended as the alarm test to control the acute toxicity of natural water bodies. PMID- 11272914 TI - Effects of bisphenol A on energy balance and accumulation in brown adipose tissue in rats. AB - Some environmental contaminants have the potential to affect humans or animals by mimicking the effects of hormones. Bisphenol A (BPA) is a weak estrogen agonist when tested using in vitro or in vivo bioassays. In addition to the well documented effects of estrogens on reproductive functions, ovarian hormones also have salient effects on mammalian energy balance and feeding behavior. In this study, we investigated the effects of BPA on body weight and food intake of ovariectomized adult female rats. Treatment with doses of 4 or 5 mg/day for 15 days resulted in a significant reduction of body weight gain with no reduction in food intake. A dose of 1 mg/day did not affect feeding or weight gain. BPA was detected in the blood, brain and adipose tissues of the BPA-treated animals but not in the vehicle control group. There was a preferential concentration of BPA in brown adipose tissue. These results indicate that BPA can affect energy balance and that brown adipose tissue may be a primary tissue into which BPA accumulates in mammals. PMID- 11272915 TI - Mercury in sediment and fish from North Mississippi Lakes. AB - Sediments and/or fish were collected from Sardis, Enid and Grenada Lakes, which are located in three different watersheds in North Mississippi, in order to assess mercury contamination. The mean total mercury concentration in sediments from Enid Lake in 1997 was 0.154 mg Hg/kg, while 1998 sediment concentrations in Sardis, Enid and Grenada Lakes were 0.112, 0.088 and 0.133 mg Hg/kg, respectively. Sediment mercury concentrations in 1999 were similar in all three lakes but, generally lower than 1998. Mean total mercury concentrations in edible fillets of fish collected from Enid Lake in 1998 were above the human health FDA action level (> 1.0 mg Hg/kg) for bass (1.40), crappie (1.69) and gar (1.89); however, tissue concentrations were less than 1.0 mg Hg/kg in carp (0.63) and catfish (0.82). Human hazard indexes for each species was > or = 1 for both adults and children, indicating that there is a potential for toxic effects to occur. In addition, calculated consumption limits indicate that adults may consume 4-12 oz. of fish per month, depending on the species consumed. For children, 2 oz. per month may be consumed. Further studies are needed to determine the exact environmental consequences and human health impacts associated with mercury contamination in North Mississippi and the Southeastern United States. PMID- 11272917 TI - Effects of ozone on chlorophyll and quantum yield of tobacco (Nicotiana tabacum L.) varieties. AB - Plants of Bel-W3 and of seven commercial tobacco varieties (Nicotiana tabacum L.) were exposed to two relatively low ozone concentrations (90 or 135 ppb) for 20 consecutive days, for 8 h per day. Ozone caused necrotic and chlorotic spots, acceleration of leaf senescence, depression of photosynthetic mechanism, chlorophyll diminution and greater destruction of chl a than of chl b. The higher sensitivity of chl a was also confirmed by exposure of segments of leaves in test tubes to high ozone concentration (>1000 ppb) as well as by bubbling of ozone in extracts of chlorophyll in vitro. The quantum yield (QY) of photosynthesis was positively correlated with the chlorophyll content and negatively correlated with the visible injury and the chl b/a ratio. PMID- 11272916 TI - Evaluation of an SOS-Chromotest-based approach for the isolation and detection of sediment-associated genotoxins. AB - A series of experiments was conducted to evaluate an approach advanced by the St. Lawrence Centre (SLC) of Environment Canada for assessing the genotoxic potential of sediments. The SLC method entails the extraction, isolation and solvent exchange of the organic constituents in sediment, and the testing of these solubilized extracts with the SOS Chromotest (Escherichia coli PQ37). A total of five sediments, three variously contaminated by organic compounds and two reference materials certified for persistent organic chemicals, were Soxhlet extracted. Each of the five extracts was then split, with a portion remaining in crude form and another portion fractionated into two molecular-weight classes of organic contaminants, thus yielding a total of 15 extract samples. The ability of the SOS Chromotest to detect genotoxins in the various organic extracts was evaluated and compared with that of the Ames Fluctuation Assay (Salmonella typhimurium, strain TA100). The intra-laboratory variance associated with the SOS Chromotest was also assessed. Procedural details are presented and results are discussed. The SOS Chromotest results were in good agreement with those of the Ames Fluctuation Assay, especially after metabolic activation. However, the E. coli PQ37 system was slightly more sensitive than the Salmonella assay for detecting genotoxins in the sediment extracts. The SOS Chromotest was also the most discriminating of the two assays, generating SOS-induction factors that were consistent with the organic contamination gradient reported in the sediment samples. The removal of macromolecules from the dichloromethane extracts by size exclusion chromatography prior to testing enhanced the sensitivity of both test systems. The intra-laboratory variance of the SOS Chromotest ranged from 0.24% to 23.82%, depending on the extract sample. As applied in this study, the SOS Chromotest can serve as a sensitive test for screening the genotoxic potential of uncharacterized sediment extracts. A more sensitive assay would be appropriate, however, as a confirmation for definitive investigations, especially for the detection of direct-acting genotoxins. PMID- 11272918 TI - Acute toxicity of cadmium to fish Labeo rohita and copepod Diaptomus forbesi pre exposed to CaO and KMnO4. AB - 96-h LC50 values of cadmium (Cd) to fish Labeo rohita and the copepod Diaptomus forbesi, determined by static bioassays, were, respectively, 89.5 and 10.2 mg/l. LC50 values increased significantly when fish pre-exposed to 100-350 mg/l CaO or 0.5-1.5 mg/l KMnO4 for 4 d and the copepod to 20-70 mg/l CaO or 0.25-1.0 mg/l KMnO4 for same period. The LC50 values also increased when the pre-exposure period of CaO was increased to 12 d at concentration 100 mg/l for fish and 20 mg/l for copepod. All fish died when pre-exposed to 1.5 mg/l KMnO4 for 8 d. But LC50 values of Cd to copepod increased when pre-exposure period of 0.5 mg/l KMnO4 was increased from 4 to 8 d. PMID- 11272919 TI - Surfactant-based oil dispersant toxicity to developing nauplii of Artemia: effects on ATPase enzymatic system. AB - The paper deals with the toxicity of a surfactant-based oil dispersant to the ATPase activities of two naupliar stages of Artemia (instar I & II). Both instars were exposed to sub-lethal and lethal concentrations derived from acute toxicity data. The chosen concentrations were near to LOECs and NOECs. An eightfold difference indicated between the instars was instar-exposure time dependent. The most prominent effects were the inhibition and the stimulation of Na+/K+-ATPase and Mg2+-ATPase activities, respectively. The cause of these effects was related to the dispersant components, the surfactants. The pattern stimulation/inhibition of Mg2+-ATPase and Na+/K+-ATPase activities could be used to indicate toxic stress by surfactant-based oil dispersants since previous studies with other contaminants have shown different ATPase activity patterns. PMID- 11272920 TI - Risk assessment of etofenprox (Vectron) on non-target aquatic fauna compared with other pesticides used as Simulium larvicide in a tropical environment. AB - Within the rotational scheme developed by the Programme to fight the resistance of Simulium damnosum to chemical larvicides, there was an operational gap at discharges between 5 and 70 m3 s(-1) for the treatment of rivers where resistance to organophosphates was present. The use of permethrin and carbosulfan was precluded because of risk of environmental impact and, Bacillus thuringiensis ser. H-14 treatments were not envisageable due to cost and logistics constraints. Among the possible complementary groups of larvicides tested, the pseudo pyrethroids, held promise, because of a mode of action similar to that of pyrethroids, but along with a usually lower toxicity for fish. Etofenprox, one of the pseudo-pyrethroids tested, shows a global detachment of non-target insects in 24 h close to that of pyraclofos, an organo-phosphorus compound (27 against 23%). In laboratory conditions, six times the operational dose which is 0.03 mg l(-1) 10 min, is needed to cause 50% mortality of Caridina sp. (a small shrimps species) and 30 times this same dose for 95% mortality. For fish species, a safety margin of 400-800 times the operational dose is observed for Oreochromis niloticus and 200-400 times for Tilapia zillii. PMID- 11272921 TI - Tissue distribution, excretion, and metabolism of 1,2,7,8-tetrachlorodibenzo-p dioxin in the rat. AB - A tissue distribution, excretion, and metabolism study was conducted using a relatively non-toxic dioxin congener, i.e., 1,2,7,8-tetrachlorodibenzo-p-dioxin (1278-TCDD), to gain a better understanding of mammalian metabolism of dioxins. Conventional, bile duct cannulated, and germ free male rats were administered mg/kg quantities as a single oral dose. Elimination of 1278-TCDD was largely complete by 72 h. Distribution of [14C]1278-TCDD was low in all tissues examined. Metabolites were identified in urine, bile, and feces by negative ion FAB-MS and 1H-NMR, or GC/MS. The major fecal metabolite was a NIH-shifted hydroxylated TCDD. The bile contained a glucuronide conjugate of this hydroxy TCDD, and a diglucuronide conjugate of a dihydroxy-triCDD. The major metabolites in urine were glucuronide and sulfate conjugates of 4,5-dichlorocatechol. PMID- 11272922 TI - Partial compensation of the sublethal effect of deltamethrin on the sex pheromonal communication of Trichogramma brassicae. AB - Pyrethroid insecticides are widely used and lead to a sizable environmental pollution that could interfere with the population biology of insects. Trichogramma is a beneficial insect used in biological control and which natural populations contribute to the control of Lepidopterus pests. In this work, we determined the effect of a sublethal dose of deltamethrin on the sex pheromonal communication of Trichogramma. The dose used (LD 0.1) induces no detectable mortality (the theoretical mortality is only one insect over 1000) and can be a good representation of contamination by this insecticide from environmental pollution. The insecticide was shown to have opposite effects on the sex pheromonal communication of Trichogramma, depending on which sex was exposed (Delpuech, J.M., Legallet, B., Terrier, O., Fouillet, P., 1999. Chemosphere 38, 729-739). We show that, when both sexes are simultaneously exposed to the insecticide, this effect is only partially neutralized. The mean response of treated males responding to the sex pheromone from treated females is not significantly different from that of controls, but the kinetics of their response is not the same. When both sexes are treated, the response of males to the sex pheromone is lower at the beginning but their response does not decrease during time contrary to controls and becomes finally higher than that of controls. Therefore, the sublethal effect of deltamethrin in the field can be either advantageous or disadvantageous depending on the difficulty in finding females and their scarcity. PMID- 11272924 TI - [Nosocomial infections: risks an level of exigiency]. PMID- 11272923 TI - [The Roissy-Charles de Gaulle airport: nurses and airplanes]. PMID- 11272925 TI - [Scars: prevention is better than treatment]. PMID- 11272926 TI - [Documents and quality of health care]. PMID- 11272928 TI - [What is PMSI? (Program of Medicalization of Information Systems)]. PMID- 11272927 TI - [Saturnism: an emergency plan]. PMID- 11272929 TI - [Evaluation of the contribution of alveolar technology applied to protection in stomies (surgical openings)]. PMID- 11272930 TI - [Preparation of a research report for publication: materials and method]. PMID- 11272931 TI - [Assessment of the critical patient at admission. An indicator of quality of care]. AB - The type of information recorded by nurses at admission of critical patients to the Intensive Care Unit was described and the relation between the information recorded and the presence of absence of endotracheal intubation in the patient admitted was analyzed. A sample of 214 admission records of patients admitted to our unit in 1998 was studied using a data sheet based on Virginia Henderson assessment questionnaires. The presence or absence of 71 variables classified into four sections was analyzed: personal data, general data, Virginia Henderson basic needs, and other assessment data. Most data collected at admission were objective data obtained by observation and/or physical examination of the patient. These data were contained in two sections: "Virginia Henderson basic needs" (normal breathing, food and water intake, excretion, mobility, maintaining posture, conserving body temperature, skin hygiene and integrity, and avoiding danger) and "other assessment data" (medical treatment, diagnostic and therapeutic tests, and hemodynamic monitoring). Information about the patient's background in the section "general data" was obtained less frequently. Subjective data obtained from interviews was clearly limited. These data are included in the "Virginia Henderson basic needs" (sleep, rest, dressing and undressing, communicating, values and beliefs, feeling of satisfaction, absence of boredom, and intellectual stimulation). PMID- 11272932 TI - [Measurement of care time in patients with acute myocardial infarction admitted to a general ICU: evaluation and improvement]. AB - Acute myocardial infarction (AMI) requires early and safe nursing care, particularly with respect to initiating and following up thrombolytic treatment, the most effective therapy according to the literature. Time is decisive. Recommended door-to-needle time should not exceed 35 minutes (from patient's arrival to injection of the thrombolytic agent in the ICU). This quality of care study centered on the measurement of four partial times and their sum. These times corresponded to different phases a patient with AMI undergoes from arrival at the hospital emergency room center to thrombolysis in the ICU. The intrahospital delay in patient care was examined. Times were recorded on a specific register of all patients with priority I AMI (clear criteria for fibrinolysis) who were seen at our center. Total time to fibrinolysis in the ICU was 60 minutes (excessive intrahospital delay). A corrective intervention plan was designed and implemented, which reduced the delay to an acceptable 30 minutes. This improved the quality of care of AMI patients at our center. PMID- 11272933 TI - [Analysis of costs and cost per diagnostic group of critically burned patients in the Spanish public health care system]. AB - The financing of the National Institute of Health (INSALUD) of Spain will soon be based on Diagnosis-Related Groups (DRGs). Knowledge of the real cost of different DRGs is fundamental to ensure adequate financing and to establish criteria for comparisons between centers. Our public health system has no data on the real cost of critically burned patients and their DRGs. This retrospective descriptive study was carried out in a Major Burns Unit (MBU) and included all patients admitted between January and December 1996. Real total cost of the care of critical burned patients, cost per patient, and cost per DRG related with critical burn patients were calculated for the study period. Financing by Weighed Care Units (WCU) was compared with real costs. The total cost of the care of critical burn patients was 346,298,872 Spanish pesetas and the cost per patient was 4,439,729 ptas. WCU financing was 322,021,616 ptas and 4,128,482 ptas, respectively. The DRG with the highest total cost was 458 (non-extensive burns with skin grafts, 106,372,016 ptas). The DRG with the highest average cost was 472 (extensive burns with surgical procedure, 5,401,119 ptas). The DRG with the highest cost per stay was 457 (extensive burns without surgical procedure, 404,683 ptas). For the first time in Spain, the cost of DRGs related with critical burn patients is described. This information is necessary for DRG-based allocation of funds and for establishing criteria to compare centers. The real cost of critical burn patients exceeded WCU financing. PMID- 11272934 TI - [Monitoring patients on mechanical ventilation]. AB - The clinical situation of a patient with acute respiratory failure, indicates the level of care and mechanical ventilatory support required. The aims of mechanical ventilation are to correct hypoventilation, improve oxygenation and oxygen transport and reduce the work of breathing. Nursing care of patients under mechanical ventilation varies according to their state and the ventilation mode. When establishing a care plan, nurses identify the objectives, which will be useful to measure the interventions. These interventions include aspects related to monitoring and indicate whether or not the objectives are met. In this article the items that allow monitoring of a patient under mechanical ventilation grouped into general monitoring and respiratory monitoring are reviewed. PMID- 11272935 TI - Comparing central venous catheters. PMID- 11272936 TI - Discovering yoga. PMID- 11272937 TI - Interpreting the highs and lows of platelet counts. PMID- 11272938 TI - Breaking down cultural barriers. PMID- 11272939 TI - Putting a stop to chicken pox. PMID- 11272940 TI - How to safeguard delivery of high-alert i.v. drugs. PMID- 11272941 TI - Unforgettable patient. Going home. PMID- 11272942 TI - What makes bradycardia tick tick tick tick tick tick tick? Tick. PMID- 11272943 TI - Nursing grand rounds. Portrait of a lady. PMID- 11272945 TI - Protecting a patient with ruptured cerebral aneurysm. PMID- 11272944 TI - Interstate licensure: nursing beyond your state's borders. PMID- 11272946 TI - Photo guide. Sound advice about hearing aids. PMID- 11272947 TI - HIV update. Faster treatments slows the spread. PMID- 11272948 TI - Action stat. Epiglottitis. PMID- 11272949 TI - Risky phlebotomy with a syringe. PMID- 11272950 TI - Just the croup. PMID- 11272951 TI - Myths & facts ... about latex allergy. PMID- 11272953 TI - Bilateral tension pneumothorax during jet ventilation. PMID- 11272952 TI - Reaching out to Mrs. Cooper. PMID- 11272954 TI - Pearl Harbor, the Korean Conflict, and COL Mildred Irene Clark. PMID- 11272955 TI - Understanding the new Medicare Outpatient Prospective Payment System. PMID- 11272956 TI - Preanesthesia detection of equipment faults by anesthesia providers at an academic hospital: comparison of standard practice and a new electronic checklist. AB - We hypothesized that our institutional standard practice for preanesthesia equipment checkout, based in part on US Food and Drug Administration recommendations, failed to detect a significant number of faults (absent or nonfunctional equipment). We designed a new, computer-based, highly interactive electronic checklist that emulated the checklist methods used in aviation and military settings and compared it to our standard practice in the detection of faulty equipment. Using a randomized, cross-over design, anesthesia providers searched for prearranged faults over a 2-day period using both the electronic and standard approaches. Faults (easy and difficult) found, faults missed, and time to complete the checkout were recorded. The electronic checklist was superior to standard practice in the detection of "easy" and "difficult" equipment faults. However, even when the electronic checklist was used, a high proportion of difficult faults were missed. Whether the failure represents a need for improved checkout procedures and provider training or better equipment design will require further study. PMID- 11272957 TI - Dislocation of the mandible: a case report. AB - Dislocation of the mandible is a possible complication of direct tracheal laryngoscopy. The temporomandibular joint (TMJ) is unique in that any movement of the bone always causes movement in both joints simultaneously. The entire TMJ is surrounded by a ligamentous capsule and is stabilized by 3 ligaments. Four muscles of mastication move the mandible with great power. The lateral pterygoid muscle has nearly horizontal muscular fibers and is chiefly responsible for dislocating the mandibular condyle and articular disc past the articular eminence into the infratemporal fossa, causing the patient great pain and distress. If mandibular dislocation should occur, prompt recognition and treatment of the dislocation is recommended. There are steps, used by dentists, which can be employed by the nurse anesthetist to relocate the mandible. The technique for intraoral bimanual relocation of the mandible is described. PMID- 11272958 TI - Faculty perceptions of characteristics needed for clinical success at military nurse anesthesia programs. AB - In this exploratory descriptive study, an investigator-developed survey tool was used to describe military clinical faculty's perception of the characteristics that nurse anesthesia students need for success in the clinical portion of graduate education. Study participants consisted of 29 clinical faculty from the Army, Air Force, and Navy (100% response). The survey tool consisted of a quantitative part with 35 characteristics in 4 categories: academic knowledge, nursing knowledge, clinical skills, and personal characteristics. Seven qualitative questions made up the second part of the survey. The faculty rated 28 of the 35 characteristics as essential or important for clinical success. All categories contained characteristics rated important or higher, with personal characteristics and clinical awareness receiving the highest ratings. Qualitative analysis of the 7 additional questions further supported the importance of personal characteristics and clinical awareness. The findings give a description of the successful student as perceived by faculty. If a student successfully completes the didactic portion of the education, personal characteristics and clinical awareness have a large role in success in the clinical portion. Knowing what characteristics are thought important allows the characteristics to be studied further. With the emphasis in this study on personal characteristics, ways to assess the difficult-to-measure personal traits need to be developed. PMID- 11272959 TI - Assessment of blood pressure discrepancies in third-trimester hypertensive gravidas. AB - Previous studies have associated hypertension with discrepancies between right arm and left arm blood pressure (BP) measurements. The purpose of this study was to determine if there were clinically (defined as > or = 10 mm Hg disparity) and statistically significant differences between right arm and left arm BP measurements (systolic, diastolic, or mean) in 34 third-trimester hypertensive gravidas. Thirty-four third-trimester normotensive gravidas were used as controls. No subjects were in active labor. This study used a cross-sectional, 2 group design with convenience sampling. The protocol for BP measurement followed guidelines of the American Heart Association and the instrument manufacturer. The results showed a greater range in BP differences between arms for the hypertensive group in the systolic (0.67-26.67 mm Hg) and mean (0.25-67 mm Hg) pressures compared with the normotensive group (systolic, 0-14.33 mm Hg; mean, 0 12 mm Hg). The mean difference in BP between arms was greater for the hypertensive group compared with the normotensive group. Using a 1-tailed t test, the mean difference was statistically significant (P < or = .05) for the systolic pressure (P = .027) and for the mean pressure (P = .022), but not the diastolic pressure (P = .168). The frequency of clinically significant differences (> or = 10 mm Hg) was greater for the hypertensive group than for the normotensive group (13 vs 4). These differences in frequencies were not statistically significant with chi-square analysis (systolic, P = .074; diastolic, P = .303; mean, P = .303). These findings indicated BP discrepancies between arms exist in both normotensive and hypertensive gravidas, with a greater range and frequency of differences in the hypertensive group. This study supports the American Heart Association's recommendation of bilateral BP assessment. PMID- 11272960 TI - AANA Journal course: update for nurse anesthetists--intraoperative fluid management for the pediatric surgical patient. AB - Intraoperative fluid management for the pediatric surgical patient is a critical element of the anesthetic care plan. In contrast with adult patients, the fluid management is systematized by the use of established protocols that calculate fluid on a per kilogram basis. Children are relatively volume sensitive, and mismanagement of fluid and electrolytes can contribute to morbidity and mortality in infants and young children undergoing even the simplest procedures. Failure to correct volume deficiencies can lead to multisystem failure and death. Inappropriate overhydration can result in pulmonary edema and respiratory problems that can prove fatal. Regardless of the fluid management plan, perioperative fluid management must be flexible and take into account the physiologic development and age of the pediatric patient. The goals of intraoperative fluid management are to restore intravascular volume, maintain cardiac output, and, ultimately, ensure provision of oxygen to the tissues. PMID- 11272961 TI - Council on Accreditation of nurse anesthesia educational programs. PMID- 11272962 TI - Mammography screening for older women with and without cognitive impairment. AB - No upper age limit exists for Medicare benefits for mammography screening, but benefits for women older than age 75 remain unclear. From a clinical perspective, it would be useful to know if there is an upper age limit for women beyond which screening for breast cancer will not extend life. Using a decision-analysis model, the author examined the utility of screening using cohorts of women age 75 to 79, 80 to 84, and 85 and older, with and without cognitive impairment. The analysis evaluated different scenarios of the benefit of biennial screening versus no screening for women who had no prior screening and women who had participated in a regular screening program. Screening increased Quality Adjusted Life Years (QALYs) at all ages. Marginal savings in life expectancy adjusted for quality of life for women with no prior screening ranged from 43.5 days for healthy 75 to 79-year-old women to 25.9 days for women older than age 85. Among cognitively impaired women who were never screened, savings ranged from 20 to 5.5 days for the three age cohorts. Biennial screening among women who had been screened continuously resulted in substantially smaller life expectancy savings, from 3.3 days for healthy individuals age 75 to 79 to less than 1 day for women older than age 85. Cost effectiveness analysis indicated the reduction in costs associated with managing recurrent disease gained by early diagnosis with mammography was greatest among the population with no prior screening. Although the increase in QALYs was consistently lower for cognitively impaired women than for their healthy counterparts, the presence of cognitive impairment did not alter the finding that screening increased QALYs. PMID- 11272964 TI - Documenting productive behaviors. Using the functional behavior profile to plan discharge following stroke. AB - The Functional Behavior Profile (FBP) is presented as a clinically useful measurement tool to guide placement decisions following stroke. The measure provides the practitioner with a tool to identify the nature and extent of the behavioral capabilities and problems caregivers will face if they choose to manage their loved ones at home. The FBP yields information for planning treatment. Individuals with scores of 84 or lower were five times more likely to need supervision after discharge than those with scores of 85 or higher. The FBP was able to correctly classify 69% of the 45 patients in this study. Additionally, the FBP identified key behaviors that discriminated between patients who go home compared to those who require supervised care. PMID- 11272963 TI - Diffusion of innovation. A model for implementation of prompted voiding in long term care settings. PMID- 11272965 TI - Reliability and validity of a community needs assessment instrument for nursing homes in Taiwan. AB - Several studies have been conducted which identify the lack of nursing homes in the Taiwan area as a major problem. However, no research instruments were found which measure the perception of need for nursing homes. The purpose of this study was to evaluate the psychometric properties of a community needs assessment instrument for nursing homes in Taiwan, called the Perception Evaluation Tool (PET). A randomly selected sample of 319 subjects participated in the study. Results suggest the PET has appropriate reliability and validity. PMID- 11272966 TI - Policy, polls, and pills. PMID- 11272967 TI - From one who's been there. PMID- 11272968 TI - Improving quality of care in nursing facilities. Gerontological clinical nurse specialist as research nurse consultant. AB - It is becoming increasingly common for nursing facilities to use Quality Indicators (QI) derived from Minimum Data Set (MDS) data for quality improvement initiatives within their facilities. It is not known how much support facilities need to effectively review QI reports, investigate problems areas, and implement practice changes to improve care. In Missouri, the University of Missouri Columbia MDS and Nursing Home Quality Research Team has undertaken a Quality Improvement Intervention Study using a gerontological clinical nurse specialist (GCNS) to support quality improvement activities in nursing homes. Nursing facilities have responded positively to the availability of a GCNS to assist them in improving nursing facility care quality. PMID- 11272969 TI - Patients undergoing some form of infusion therapy. PMID- 11272970 TI - A comparison of continuous infusion and intermittent flushing methods in peripheral intravenous catheters in neonates. AB - The purpose of this study was to compare two methods of maintaining peripheral intravenous devices in neonates: continuous infusion (CI) and intermittent flushing (IF). There was no significant difference in the mean duration of patency between the two groups, but there was a significant difference with respect to reasons for removal or loss of patency. The main reason for removal in the CI group was infiltration or phlebitis, and in the IF group the reason was occlusion. PMID- 11272971 TI - The effect of a 1-hour training program on the incidence of bacteremia in pediatric patients receiving parenteral nutrition. AB - The effect of a 1-hour nurse training program on the frequency of bacteremia in patients receiving parenteral nutrition was evaluated in a pediatric tertiary center. All of the nurses had previous instruction on aseptic techniques in nursing school. The current program focused on aseptic management of intravenous catheters and implanted subcutaneous ports in patients receiving parenteral nutrition (PN). One hundred eighty-four nurses had a 1-hour training session in groups of three to five. The frequency of bacteremia in children receiving PN was not reduced (9.2% versus 8.9%), and there was no significant difference in time from the start of PN to the diagnosis of bacteremia (P = 0.31). The authors conclude that a 1-hour training session for the nursing staff was not sufficient. It is suggested that staff training for prevention of bloodstream infections associated with intravascular devices should cover a wider range of topics and take place over a longer period of time. PMID- 11272973 TI - Local complications of nursing interventions on peripheral veins. AB - The authors analyze the causes of complications after nursing interventions on peripheral veins. Twenty-one percent of all venipunctures involve complications, mainly subcutaneous hematomas (62% of all complications), paravasal injection of drugs (28%), spontaneous rupture of the vein (6%), obliteration of the vessel (2%), superficial phlebitis (2%), external bleeding from the vein (0.5%), and local allergic reactions (0.5%). The complications are cause mainly by incorrect technique. Recommendations are offered for preventing complications associated with peripheral infusion treatments. PMID- 11272972 TI - Effect of two different short peripheral catheter materials on phlebitis development. AB - One of the most common causes of phlebitis in hospitalized patients is intravenous catheters. The material of the catheter is a determining factor in the development of phlebitis, as are factors such as age, gender, and medical diagnosis of the patient. The aim of this study, conducted in the coronary care unit of a 384-bed hospital in Ankara, Turkey, was to determine the effect of two different short peripheral catheters on phlebitis development caused by i.v. treatment. Overall, 255 patients constituted the study sample (130 with Teflon, 125 with Vialon catheters). Both groups were followed up for phlebitis development for 6 days. The total phlebitis rate was 36.8%, with almost half of the patients (49.2%) in the Teflon catheter group and 24.0% of patients in the Vialon catheter group. A significant statistical relationship was found between phlebitis rate and variables such as gender, catheter material, and indwelling time. The results of the study demonstrate that Vialon catheters are associated with less risk of catheter-induced phlebitis than are Teflon catheters. PMID- 11272974 TI - Intravenous line management and prevention of catheter-related infections in America. AB - Advanced medical treatment is practiced both in hospitals and in homecare. Because most patients receiving advanced treatments need intravenous therapy, i.v. management is becoming increasingly important. Medical staffs are required to evaluate various i.v. technologies from the perspective of infection control and to select the most appropriate i.v. access line for the patient. For this reason, a seminar was held to exchange the latest information on i.v. line management and infection control in Japan and America. Guest speakers from the Intravenous Nurses Society of America were invited. The article provides a summary of the seminar. PMID- 11272975 TI - 5-fluorouracil extravasation following port failure. AB - A case is presented of cytotoxic extravasation as a result of an implantable venous port being perforated by a standard Huber needle. A patient receiving 5 fluorouracil via a dual reservoir port, implanted within the left chest wall, presented with hemoserous discharge from the right needle entry site. The left chest wall was warm to touch, erythematous, and swollen. Subcutaneous infiltration was suspected, and the infusion was ceased. A venogram was performed demonstrating significant extravasation around the left reservoir only. On port removal, inspection showed the Huber needle had penetrated the base plate on the left side. It is recommended that this complication be added as a possible sequelae of central venous port use. PMID- 11272976 TI - Teenage pregnancies. A baseline audit. PMID- 11272977 TI - Amniocentesis and the specialist midwife. The developing role. PMID- 11272978 TI - The conundrum of perineal care. PMID- 11272979 TI - The influence of antenatal classes on pain relief in labour. A review of the literature. PMID- 11272980 TI - Guidelines for supervisors of midwives in Yorkshire. PMID- 11272982 TI - Antepartum haemorrhage. Teaching to enhance practice. PMID- 11272981 TI - Supervision of midwives. What makes it special? PMID- 11272983 TI - Does your midwifery unit have a blame culture? PMID- 11272984 TI - Pleased to meet you. Julia Magill-Cuerden. PMID- 11272985 TI - In praise of an unusual midwife. PMID- 11272986 TI - [Drugs--magic or science?]. PMID- 11272988 TI - [Nursing rights and nursing reality]. PMID- 11272987 TI - [To fight headache]. PMID- 11272989 TI - [Are balloon catheters not as good as believed?]. PMID- 11272990 TI - [3M Cavilon--a non-irritating dermatologic agent]. PMID- 11272991 TI - [The effect of BIOPTRON-light in rheumatology]. PMID- 11272992 TI - [Dimensions of diseases]. PMID- 11272993 TI - [Opinions and attitudes of intensive care nurses on the effect of open visits on patients, family members, and nurses]. AB - The policy of family visits to patients admitted to the intensive care unit has been liberalized in recent years. This change has been progressive in our unit and family members now spend long periods of time with patients. An analysis was made of the beliefs, opinions and attitudes of nurses toward family visits and the relation between the beliefs of nurses and their attitude toward the effect of an open visiting policy on patients, family members and nurses. A descriptive correlation study was carried out in the Polyvalent Intensive Care Unit. The sample included 46 nurses who completed a self-administered, anonymous questionnaire. This questionnaire contained a Likert type scale analyzing the opinions of nurses regarding the effect of visits and a differential semantic scale analyzing nurses' attitudes toward visits by family members. The opinion that visits had a positive effect achieved a mean value of 3.001 on a scale with a maximum value of 4. The score obtained on the scale of attitudes toward an open visiting policy was 6.005, with a maximum value of 7. The correlation between opinions and attitudes was significant and positive (r = 0.523, p > 0.0001). Comparison of sociodemographic and other variables disclosed no statistically significant differences, except for the variables attitude and having children (t = -2.254, p = 0.03), which obtained a higher score. It is concluded that the opinions of nurses regarding the positive effect of open visits depended on their attitudes. For the most part, they were satisfied with the current visiting policy. PMID- 11272994 TI - [Pressure sores: evaluation of the systematic use of special surfaces for managing pressure sores in the intensive care unit of the Tarrasa Hospital, Spain]. AB - Pressure sores are a major challenge for healthcare systems. Patients admitted to intensive care units are an important risk group for pressure sores. Systematic use of a protocol employing special surfaces to manage pressure is a basic measure for preventing pressure sores in institutionalized patients. In a study carried out in the Intensive Care Unit of the Hospital of Terrassa (Spain), the incidence of pressure sores before and after introducing a prevention protocol that included the systematic use of special surfaces to manage pressure was compared. The incidence of pressure sores in patients admitted in 1998 and 1999 was studied. Sore locations were recorded. The results of three periods were examined: before introducing the protocol (6.4% incidence), after introducing a protocol (1.1%), and after use of the protocol was consolidated (0%). The results of the study suggest that the systematic use of special surfaces to manage pressure, in accordance with a prevention protocol, is a basic measure for reducing the incidence of pressure sores in patients admitted to intensive care units. PMID- 11272995 TI - [Postural technique in prone position: hemodynamic and respiratory parameters and complications]. AB - Therapeutic strategies used in the treatment of adult respiratory distress syndrome (ARDS) recommend placing the patient in prone position as an effective method for optimizing ventilation-perfusion parameters. We evaluated the therapeutic effect of postural treatment in prone position with the following goals: Comparison of hemodynamic and respiratory parameters before and after placing the patient in prone position, while in prone position, and before and after postural treatment. Complications associated with turning and time in prone position. A prospective study was made of 30 turns in 15 patients admitted to the Polyvalent ICU between January 1999 and April 2000 for medical-surgical pathology, mean age 55.4 +/- 16.3 years, diagnosed as ARDS during their stay in the ICU, Lung Injury Score (Murray) > 2.5, and a medical prescription for prone position. Nurses were experienced in postural treatment in prone position in accordance with two protocols, the technique for placing the patient in prone position and nursing care for patients in prone position. Comparison of paired means of the hemodynamic variables MBP, HR and CVP did not disclose any statistically significant difference between the time before turning, while in prone position, and before and after postural treatment. Comparison of paired means of the respiratory variables PaO2/FiO2, Sat O2, tidal volume (TV), PCO2, pH, and PaO2/FiO2 ratio showed a significant increase after placing the patient in prone position, another increase after 2 hours in prone position, and before and after postural treatment. Sat O2 increased significantly 2 hours after turning, and remained raised while the patient was in prone position. TV increased significantly before and after postural treatment. The reduction in PCO2 occurred after 2 hours in prone position. Comparisons before and after postural treatment disclosed a clinically significant difference but no change in pH. Following the protocol for placing the patient in prone position, no complications were associated with the turning procedure (accidental loss of TOT, tracheostomy, SNG, urinary catheter, vascular catheters, chest tubes, and drainage tubes). While patients were in prone position, stage II and III UPP, palpebral and/or conjunctival edema, and intolerance of enteral feeding occurred, but our results do not indicate that these complications appeared solely as a result of prone position. PMID- 11272996 TI - [Ethical guidelines for caregiving]. PMID- 11272997 TI - [Preparation of an article on research for publication]. PMID- 11272999 TI - Subacute combined degeneration one century later. The neurotrophic action of cobalamin (vitamin B12) revisited. PMID- 11273000 TI - Alzheimer-type I astrogliopathy (AIA) and its implications for dynamic plasticity of astroglia: a historical review of the significance of AIA. AB - Alzheimer-type I astrogliopathy (AIA) is an uncommon neuropathological phenomenon encountered in Wilson's disease and less often in acquired hepatic encephalopathy. Since its first description in 1912 it has received little attention. However, after 1971, when the nature of its morphogenesis began to be recognized and it was shown that it could be reproduced experimentally, its significance has been increasingly appreciated. Two intriguing characteristics of the dynamic plasticity of astroglia were revealed from the studies of the inter relationships between AIA and Alzheimer-type II astrogliopathy (AIIA); normal astroglia and AIIA; and reactive astrogliosis and AIIA, namely, the compensatory "rebound" phenomenon of Alzheimer astrogliopathy, and a dual cellular origin for reactive astrogliosis taking place in both normal and dystrophic astrocytes. More recently the presence of AIA and AIIA has been reported in a case of anoxic encephalopathy, and also in a case of Marchiafava-Bignami's disease. In this review, dependable criteria for the identification of the pathological features of AIA are discussed and emphasized. Both types of Alzheimer astrogliopathy may be used as pathologic markers with specific morphological and immunocytochemical characteristics to study in detail the disturbances of metabolic interactions between the astrocyte-neuron coupling and the exact mechanisms of the dynamic plasticity of astroglia. PMID- 11272998 TI - [Effect of two different tracheal cannula change schedules on microbiological contamination and patient comfort]. AB - Tracheostomy is a commonly used technique in intensive care units, but there are no uniform criteria governing the periodicity with which tracheal cannulas should be changed. The objective of our study was to evaluate if different cannula change schedules modified microbiological contamination and reduced the pain and bleeding related with cannula changes. In a comparative study of two groups, a control group in which the cannula was change every 48 hours and an experimental group in which the cannula was changed every 5 days were studied. Demographic differences, tracheostomy technique, microbiological study of the cannula, bronchial aspirate and stoma, clinical signs of stomal infection and secretions, and chest radiography were compared in the two groups. With each cannula change, we evaluated bleeding, pain, type of ventilation, hemodynamic disturbances, airway obstruction, opening of a false airway, oxygen saturation before and after cannula change, and recovery time. The study included 29 patients and 97 cannulas. In a homogeneous sample, the patients in the experimental group had a normal chest radiograph for a significantly longer time (p = 0.005). The stomas of the experimental group produced significantly less seepage (p = 0.04) and pain (p = 0.004). When the tracheostomy technique was correlated with the stoma, surgical tracheostomy performed in the unit showed significantly more reddening (p < 0.004) and seeping (p < 0.001). We conclude that prolonging cannula changes to every 5 days did not increase the incidence of contamination and reduced the pain of tracheostomized patients. PMID- 11273001 TI - The modulations of NCAM polysialylation state that follow transient global ischemia are brief on neurons but enduring on glia. AB - To investigate the role of polysialylated neural cell adhesion molecule (NCAM PSA)-mediated plasticity after injury, we examined the temporal and spatial expression of NCAM PSA immunoreactivity in the medial temporal lobe following global ischemia. Male Mongolian gerbils were subjected to bilateral common carotid artery occlusion for 5 min and killed at increasing times post-occlusion. The well-characterized delayed CAl pyramidal cell death was observed 5-7 days post-occlusion. At post-occlusion days 1-2 there was a small but significant increase of NCAM PSA-positive hippocampal granule cells followed by an equally significant decrease at post-occlusion day 5. In contrast, a substantial increase in glial PSA expression was observed in all hippocampal regions at 1-7 days post occlusion that was associated generally with stellate astroglia and specifically with the radial processes of glia traversing the granule cell layer of the dentate gyrus. Administration of the glutamate antagonist 2,3-dihydroxy-6-nitro-7 sulfamoyl-ben-zo(F)quinoxaline significantly blocked the ischemia-induced modulation of neuronal and glial NCAM PSA expression. Astroglial NCAM polysialylation became attenuated by 35 days post-occlusion except in the CAI area of cell death. The temporal and regional pattern of polysialylated NCAM expression in the ischemic gerbil hippocampus implicates this neuroplastic marker in mechanisms of neurotrophic-dependent repair/remodeling that ensue following transient interruption of blood flow. PMID- 11273002 TI - Alpha2 receptor binding in the medulla oblongata in the sudden infant death syndrome. AB - The sudden infant death syndrome (SIDS) is the leading cause of postnatal infant mortality in the United States. Its etiology remains unknown. We propose that SIDS, or a subset of SIDS, is due to a failure of autoresuscitation, a protective brainstem response to asphyxia or hypoxia, in a vulnerable infant during a critical developmental period. Gasping is an important component of autoresuscitation that is thought to be mediated by the "gasping center" in the lateral tegmentum of the medulla, a region homologous in its cytoarchitecture and chemical anatomy to the intermediate reticular zone (IRZ) in the human. Since we found that [3H]para-aminoclonidine ([3H]PAC) binding to alpha2-adrenergic receptors localizes to this region in human infants and, thereby provides a neurochemical marker for it, we tested the hypothesis that [3H]PAC binding to alpha2-adrenergic receptors is decreased in the IRZ in SIDS victims. Using quantitative tissue autoradiography with [3H]PAC as the radioligand and phentolamine as the displacer, we analyzed alpha2-receptor binding density in the IRZ, as well as in 7 additional sites for comparison, in 10 SIDS and 10 control medullae. There were no significant differences in alpha2 receptor binding in the IRZ, vagal nuclei, or other medullary sites examined between SIDS and control cases. These results suggest that the putative gasping defect in the IRZ in SIDS victims is not related to [3H]PAC binding to alpha2-adrenergic receptors. PMID- 11273004 TI - Early and progressive accumulation of reactive microglia in the Huntington disease brain. AB - Microglia may contribute to cell death in neurodegenerative diseases. We studied the activation of microglia in affected regions of Huntington disease (HD) brain by localizing thymosin beta-4 (Tbeta4), which is increased in reactive microglia. Activated microglia appeared in the neostriatum, cortex, and globus pallidus and the adjoining white matter of the HD brain, but not in control brain. In the striatum and cortex, reactive microglia occurred in all grades of pathology, accumulated with increasing grade, and grew in density in relation to degree of neuronal loss. The predominant morphology of activated microglia differed in the striatum and cortex. Processes of reactive microglia were conspicuous in low grade HD, suggesting an early microglia response to changes in neuropil and axons and in the grade 2 and grade 3 cortex, were aligned with the apical dendrites of pyramidal neurons. Some reactive microglia contacted pyramidal neurons with huntingtin-positive nuclear inclusions. The early and proximate association of activated microglia with degenerating neurons in the HD brain implicates a role for activated microglia in HD pathogenesis. PMID- 11273003 TI - Insulin-like growth factor-I and over-expression of Bcl-xL prevent glucose mediated apoptosis in Schwann cells. AB - Schwann cells (SCs), the myelinating cells of the peripheral nervous system, are lost or damaged in patients suffering from diabetic neuropathy. In the current study, 2 model systems are used to study the mechanism of SC damage in diabetic neuropathy: the streptozotocin (STZ)-treated diabetic rat and cultures of purified SCs in vitro. Electron microscopy of dorsal root ganglia from STZ treated rats reveals classic ultrastructural features of apoptosis in SCs, including chromatin clumping and prominent vacuolation. Bisbenzamide staining of SCs cultured in hyperglycemic defined media shows nuclear blebbing of apoptotic cells. Insulin-like growth factor-I (IGF-I) is protective. LY294002, a phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor, blocks the effect of IGF I. High glucose induces caspase cleavage in apoptotic SCs--an effect that is blocked by bok-asp-fmk (BAF), a caspase inhibitor. Although Bcl-xL expression remains unchanged in experimental conditions, over-expression of Bcl-xL protects SCs from apoptosis. In summary, hyperglycemia induces caspase activation and morphologic changes in SCs consistent with apoptotic death, both in vivo and in vitro. Over-expression of Bcl-xL, or IGF-I, signaling via PI 3-kinase, protects SCs from glucose-mediated apoptosis in vitro. IGF-I may be useful in preventing hyperglycemia-induced damage to SCs in patients suffering from diabetic neuropathy. PMID- 11273005 TI - Absence of detectable IL-1beta production in murine prion disease: a model of chronic neurodegeneration. AB - Murine prion disease is accompanied by a modified inflammatory response characterized by early but prolonged microglial activation and T-lymphocyte recruitment. In this model, we look at the profile of cytokine production, particularly IL-1beta. Mice inoculated with prion-infected brain homogenate show typical signs of prion disease. We were unable to detect any IL-1beta using immunohistochemistry, with various fixation protocols, or ELISA between 8 and 24 wk post-inoculation. Also, there was no increase in mRNA for IL-1beta, IL-6, IFNgamma, and iNOS as measured by quantitative RT-PCR. Using the same procedures and examining tissues at the same time, IL-1beta immunostaining was detected in infiltrating inflammatory cells in mouse brains injected with LPS or in a delayed type hypersensitivity response in the brain. Soluble IL-1beta was also increased, as measured by ELISA, and there was an increase in mRNA species for IL-1beta, IL 6, TNFalpha but not IFNgamma or iNOS in these brains. These data reveal that chronic neurodegeneration seen in prion disease does not induce production of a range of proinflammatory mediators despite showing marked microglial activation and raise the question as to whether IL-1beta would exacerbate the neurodegeneration as it does in acute neurodegeneration following head injury and stroke. PMID- 11273007 TI - Alzheimer-associated neuronal thread protein-induced apoptosis and impaired mitochondrial function in human central nervous system-derived neuronal cells. AB - In Alzheimer Disease (AD), dementia is due to cell loss and impaired synaptic function. The cell loss is mediated by increased apoptosis, predisposition to apoptosis, and impaired mitochondrial function. Previous studies demonstrated that the AD7c-NTP neuronal thread protein gene is over-expressed in AD beginning early in the course of disease, and that in AD, AD7c-NTP protein accumulation in neurons co-localizes with phospho-tau-immunoreactivity. To determine the potential contribution of AD7c-NTP over-expression to cell loss in AD, we utilized an inducible mammalian expression system to regulate AD7c-NTP gene expression in human CNS-derived neuronal cells by stimulation with isopropyl-1 beta-D-thiogalactopyranoside (IPTG). IPTG induction of AD7c-NTP gene expression resulted in increased cell death mediated by apoptosis, impaired mitochondrial function, and increased cellular levels of the p53 and CD95 pro-apoptosis gene products as occur in AD. In addition, over-expression of AD7c-NTP was associated with increased levels of phospho-tau, but not amyloid-beta immunoreactivity. These results suggest that AD7c-NTP over-expression may have a direct role in mediating some of the important cell death cascades associated with AD neurodegeneration, and further establish a link between AD7c-NTP overexpression and the accumulation of phospho-tau in preapoptotic CNS neuronal cells. PMID- 11273008 TI - Second primary glioblastoma. AB - Although characterized by a highly variable phenotype and multiple genetic alterations, glioblastomas are considered monoclonal in origin. We here report on a 64-yr-old patient who developed a second glioblastoma in the left frontal lobe 10 yr after surgical resection of a glioblastoma of right frontal lobe. The first tumor contained 2 p53 mutations, in codon 213 (CGA-->TGA, Arg-->stop) and codon 306 (CGA-->TGA, Arg-->stop), further, 1 missense PTEN mutation (codon 257, TTC- >TTA, Phe-->Leu) and a silent PTEN mutation (codon 154, TTC-->TTT, Phe-->Phe). The second glioblastoma also contained multiple, but different mutations: p53 mutations in codons 158 (CGC-->CAC, Arg-->His) and 273 (CGT-->TGT, Arg-->Cys), and a PTEN mutation in codon 233 (CGA-->TGA, Arg-->Stop). Both neoplasms had a homozygous p16 deletion. The discordant pattern of mutations indicates that the second glioblastoma was not a recurrence but an independent second glioblastoma. The presence in these neoplasms of multiple mutations in tumor suppressor genes suggests the involvement of a novel disease mechanism but there was no indication of a DNA mismatch repair deficiency or of an inherited tumor syndrome. PMID- 11273009 TI - Pharmacognosy in the 21st century. AB - The term pharmacognosy as a constituent scientific discipline of pharmacy has been in use for nearly 200 years, and it refers to studies on natural product drugs. During the last half of the 20th century, pharmacognosy evolved from being a descriptive botanical subject to one having a more chemical and biological focus. At the beginning of the 21st century, pharmacognosy teaching in academic pharmacy institutions has been given new relevance, as a result of the explosive growth in the use of herbal remedies (phytomedicines) in modern pharmacy practice, particularly in western Europe and North America. In turn, pharmacognosy research areas are continuing to expand, and now include aspects of cell and molecular biology in relation to natural products, ethnobotany and phytotherapy, in addition to the more traditional analytical method development and phytochemistry. Examples are provided in this review of promising bioactive compounds obtained in two multidisciplinary natural product drug discovery projects, aimed at the elucidation of new plant-derived cancer chemotherapeutic agents and novel cancer chemopreventives, respectively. The systematic study of herbal remedies offers pharmacognosy groups an attractive new area of research, ranging from investigating the biologically active principles of phytomedicines and their mode of action and potential drug interactions, to quality control, and involvement in clinical trials. PMID- 11273006 TI - Acute cytoskeletal alterations and cell death induced by experimental brain injury are attenuated by magnesium treatment and exacerbated by magnesium deficiency. AB - Traumatic brain injury results in a profound decline in intracellular magnesium ion levels that may jeopardize critical cellular functions. We examined the consequences of preinjury magnesium deficiency and post-traumatic magnesium treatment on injury-induced cytoskeletal damage and cell death at 24 h after injury. Adult male rats were fed either a normal (n = 24) or magnesium-deficient diet (n = 16) for 2 wk prior to anesthesia and lateral fluid percussion brain injury (n = 31) or sham injury (n = 9). Normally fed animals were then randomized to receive magnesium chloride (125 micromol, i.v., n = 10) or vehicle solution (n = 11) at 10 min postinjury. Magnesium treatment reduced cortical cell loss (p < 0.05), cortical alterations in microtubule-associated protein-2 (MAP-2) (p < 0.05), and both cortical and hippocampal calpain-mediated spectrin breakdown (p < 0.05 for each region) when compared to vehicle treatment. Conversely, magnesium deficiency prior to brain injury led to a greater area of cortical cell loss (p < 0.05 compared to vehicle treatment). Moreover, brain injury to magnesium deficient rats resulted in cytoskeletal alterations within the cortex and hippocampus that were not observed in vehicle- or magnesium-treated animals. These data suggest that cortical cell death and cytoskeletal disruptions in cortical and hippocampal neurons may be sensitive to magnesium status after experimental brain injury, and may be mediated in part through modulation of calpains. PMID- 11273010 TI - Increased vigabatrin entry into the brain by polysorbate 80 and sodium caprate. AB - The effects of a non-ionic surfactant, polysorbate 80, and the sodium salt of the saturated fatty acid, sodium caprate (C10), as potential brain absorption enhancers for vigabatrin were studied. Vigabatrin is an enzyme-activated irreversible inhibitor of gamma-aminobutyric acid (GABA) transaminase that increases brain and cerebrospinal GABA concentrations in animals and man. Before intravenous administration, a range of concentrations of the surfactants were tested using erythrocyte lysis or the red blood cell lysis test to establish the non-toxic concentration range. Vigabatrin was dissolved in 0.1% polysorbate 80 and 0.1% sodium caprate and administered intravenously in doses of 4 mL kg(-1) to male Wistar rats (230-250 g; n = 3). Rats were killed 2 h after drug and surfactant administration and the brains were immediately removed and homogenized in 0.4 M perchloric acid. Selected ion monitoring electrospray mass spectrometry was used to determine the concentration of vigabatrin and GABA directly from the perchloric acid extract of the rat brain. This method was developed to increase the speed and efficiency of the analysis by removing the need for complex extraction and derivatization procedures while retaining the specificity of the mass spectrometer as a detector. The stability of both vigabatrin and GABA in perchloric acid was established by monitoring their pseudo molecular ions in standard solutions at timed intervals over 24 h. Although the detection level for vigabatrin and GABA was at least 50 pg, only GABA was detected in rat brain. Vigabatrin caused a small increase in whole brain GABA. However, GABA levels were higher in the samples with vigabatrin + enhancer than in the samples where vigabatrin alone was administered. One-way analysis of variance indicated a significant effect of the surfactants on GABA levels (F (5,17) = 11.86, P < 0.01) and vigabatrin absorption was presumed. The rectal temperature of the rats is lowered by the presence of vigabatrin in the brain. Vigabatrin alone decreased rectal temperature by 6%. When given with either polysorbate 80 or sodium caprate, the extent of temperature lowering was significantly greater (P < 0.001). There was no significant difference after 2 h between polysorbate 80 + vigabatrin, and sodium caprate + vigabatrin. PMID- 11273011 TI - In-vitro and in-vivo evaluation of pH-responsive polymeric micelles in a photodynamic cancer therapy model. AB - pH-sensitive polymeric micelles of randomly and terminally alkylated N isopropylacrylamide copolymers were prepared and characterized. Aluminium chloride phthalocyanine (AlClPc), a second generation sensitizer for the photodynamic therapy of cancer, was incorporated in the micelles by dialysis. Their photodynamic activities were evaluated in-vitro against EMT-6 mouse mammary tumour cells and in-vivo against EMT-6 tumours implanted intradermally on each hind thigh of Balb/c mice. pH-sensitive polymeric micelles were found to exhibit greater cytotoxicity in-vitro than control Cremophor EL formulations. In the presence of chloroquine, a weak base that raises the internal pH of acidic organelles, in-vitro experiments demonstrated the importance of endosomalllysosomal acidity for the pH-sensitive polymeric micelles to be fully effective. Biodistribution was assessed by fluorescence of tissue extracts after intravenous injection of 2 micromol kg(-1) AlClPc. The results revealed accumulation of AlClPc polymeric micelles in the liver, spleen and lungs, with a lower tumour uptake than AlClPc Cremophor EL formulations. However, polymeric micelles exhibited similar activity in-vivo to the control Cremophor EL formulations, demonstrating the higher potency of AlClPc polymeric micelles when localized in tumour tissue. It was concluded that polymeric micelles represent a good alternative to Cremophor EL preparations for the vectorization of hydrophobic drugs. PMID- 11273012 TI - Encapsulation of bovine serum albumin in poly(lactide-co-glycolide) microspheres by the solid-in-oil-in-water technique. AB - Non-aqueous protocols to encapsulate pharmaceutical proteins into biocompatible polymers have gained much attention because they allow for the minimization of procedure-induced protein structural perturbations. The aim of this study was to determine if these advantages could be extended to a semi-aqueous encapsulation procedure, namely the solid-in-oil-in-water (s/o/w) technique. The model protein bovine serum albumin (BSA) was encapsulated into poly(lactide-co-glycolide) (PLG) microspheres by first suspending lyophilized BSA in methylene chloride containing PLG, followed by emulsification in a 1% aqueous solution of poly(vinyl alcohol). By variation of critical encapsulation parameters (homogenization intensity, BSA:PLG ratio, emulsifier concentration, ratio of organic to aqueous phase) an encapsulation efficiency of > 90% was achieved. The microspheres obtained showed an initial burst release of < 20%, a sustained release over a period of about 19 days, and a cumulative release of at least 90% of the encapsulated BSA. Different release profiles were observed when using different encapsulation protocols. These differences were related to differences in the microsphere erosion observed using scanning electron microscopy. Release of BSA was mainly due to simple diffusion or to both diffusion and microsphere erosion. Fourier-transform infrared studies were conducted to investigate the secondary structure of BSA during the encapsulation. Quantification of the alpha-helix and beta-sheet content as well as of overall structural changes showed that the secondary structure of encapsulated BSA was not more perturbed than in the lyophilized powder used initially. Thus, the encapsulation procedure did not cause detrimental structural perturbations in BSA. In summary, the results demonstrate that the s/o/w technique is an excellent alternative to the water-in-oil-in-water technique, which is still mainly used in the encapsulation of proteins in PLG microspheres. PMID- 11273013 TI - Pharmacokinetics of diaspirin cross-linked haemoglobin in a rat model of hepatic cirrhosis. AB - The aim of the study was to evaluate the effect of cirrhosis on the disposition of the haemoglobin-based oxygen carrier, diaspirin cross-linked haemoglobin (DCLHb). Cirrhosis was induced in male Sprague-Dawley rats (200-250 g) by inhalational exposure to carbon tetrachloride (CCl4), over a period of 6 weeks. Pharmacokinetic evaluation was performed after a single intravenous bolus administration of DCLHb (400 mg kg(-1)). Serum biochemistry, including aspartate transaminase, alkaline phosphatase, bile acids, serum albumin, and serum creatinine, were measured in CCl4-treated (n = 6) and age-matched control (n = 6) rats. After 6 weeks, the jugular vein and carotid artery were cannulated for bolus DCLHb administration (400 mg kg(-1)) and blood sampling, respectively, in both groups of rats. Cirrhosis produced significant (P < 0.05) elevations in alkaline phosphatase (497.4 +/- 84.8 U L(-1) vs 241.2 +/- 5.1 U L(-1)), aspartate transaminase (920.5 +/- 190.9 U L(-1) vs 238.2 +/- 118.1 U L(-1)) and bile acids (333.8 +/- 77.3 mg dL(-1) vs 43.8 +/- 4.2 mg dL(-1)) compared with the control group. No significant renal dysfunction was observed as a result of CCl4 exposure. Plasma DCLHb concentrations declined approximately log-linearly. Systemic clearance of DCLHb was estimated to be 2.2 +/- 0.7 mL h(-1) in the treatment group and was slightly, but not significantly, less in the control group (3.6 +/- 1.7 mL h(-1)). There was also a trend toward a longer elimination half-life in the treatment group (4.7 +/- 2.2 h) compared with the control group (3.8 +/- 0.8 h), although this difference was not statistically significant. Cirrhosis does not significantly alter the disposition of DCLHb perhaps due to increased extra-hepatic metabolism by the reticulo-endothelial system. PMID- 11273014 TI - Antimicrobial activity of tannin components from Vaccinium vitis-idaea L. AB - Reactive oxygen species have been implicated as important pathological mediators in many clinical disorders, including periodontal disease. As a possible alternative for the treatment of periodontal disease, the antimicrobial activity of six tannins isolated from Vaccinium vitis-idaea L., with confirmed antioxidant activity, were assayed by the agar dilution method against selected periodontal pathogens, Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Prevotella intermedia. The results showed that epicatechin-(4beta-->8) epicatechin-(4beta-->8, 2beta-->O-->7)-catechin had strong antimicrobial activity against P. gingivalis and P. intermedia, but not A. actinomycetemcomitans. The other tannins tested did not show antimicrobial activity. We conclude that tannins isolated from V. vitis-idaea L. with antimicrobial activity could potentially be used for the treatment of periodontal disease. PMID- 11273015 TI - Evaluation of anti-inflammatory potential of Bergenia ciliata Sternb. rhizome extract in rats. AB - The methanol extract of the rhizome of Bergenia ciliata Sternb. (Saxifragaceae) has been evaluated for anti-inflammatory potential using two acute rat models (carrageenan- and serotonin (5-HT)-induced rat paw oedema) and a chronic rat model (cotton pouch-induced granuloma). Phenylbutazone (100 mg kg(-1)), a non steroidal anti-inflammatory agent, was used as a standard. The methanol extract (100, 200 or 300 mg kg(-1)) exhibited significant (P < 0.05) anti-inflammatory activity in all the animal models. At 300 mg kg(-1) the methanol extract exhibited maximum inhibition of 32.4+/-2.89% in carrageenan-induced rat paw oedema while the standard showed an inhibition of 44.1+/-2.7% after 3 h of drug treatment. In the serotonin-induced rat paw oedema model, 300 mg kg(-1) methanol extract suppressed oedema by 45.33+/-2.09%, whereas the standard produced an inhibition of 53.5+/-4.3%. In the cotton pouch granuloma model the methanol extract inhibited significantly (P < 0.001) the granuloma weight in a dose dependent manner. In this model, 300 mg kg(-1) extract produced a maximum inhibition of 31.4+/-1.09% in granuloma weight compared with 41.1+/-1.32% reduction in granuloma weight for the standard. The methanol extract of B. ciliata exhibited significant anti-inflammatory potential at the dose levels examined. PMID- 11273016 TI - Anti-fibrotic effects of a hot-water extract from Salvia miltiorrhiza roots on liver fibrosis induced by biliary obstruction in rats. AB - The anti-fibrotic effects of a hot-water extract form the traditional Chinese medicinal herb Salvia miltiorrhiza (Labiatae) on liver fibrosis induced by biliary obstruction was studied in rats. Liver fibrosis was induced in male Sprague-Dawley rats by bile duct ligation and scission (BDL). After surgery, the hot-water extract of S. miltiorrhiza roots (100 mg kg(-1), p.o.) was administered daily for 28 days. The concentrations of aspartate transaminase, alanine transaminase, alkaline phosphatase, total bilirubin and total cholesterol in serum and hydroxyproline and malondialdehyde contents in liver were significantly increased in BDL rats. Treatment with the extract of S. miltiorrhiza significantly reduced (P < 0.01) the serum aspartate transaminase, alanine transaminase, alkaline phosphatase, and total cholesterol concentrations in BDL rats. The liver hydroxyproline content in BDL rats treated with extract was also reduced to 68% of that in BDL control rats (P < 0.01). The liver malondialdehyde content in BDL rats treated with the extract was also reduced to 47% of that in BDL control rats (P < 0.01). The morphological characteristics of fibrotic livers were improved in BDL rats treated with extract. Immunohistochemical examination of fibrotic liver showed that the extract of S. miltiorrhiza markedly reduced protein expression of alpha-smooth muscle cell-like actin, which indicates that hepatic stellate cell activation was inhibited during liver fibrosis development. The results indicate that the hot-water extract of S. miltiorrhiza roots inhibits fibrosis and lipid peroxidation in rats with liver fibrosis induced by biliary obstruction. PMID- 11273017 TI - Potency, affinity constants and receptor reserves for noradrenaline and adrenaline on aortae from aged normo- and hypertensive rats. AB - Previously we have determined the potency, affinity constants (K(A) values), and alpha1-adrenoceptor reserves for noradrenaline and adrenaline on the thoracic aortae of 20-week-old Wistar Kyoto normotensive (WKY) and spontaneously hypertensive rats (SHRs). This study has investigated whether these parameters were altered on the thoracic aortae by ageing, and in hypertension/heart failure. The effects of phenoxybenzamine on the contractile responses of the aortae of 20 month-old WKYs and SHRs were determined. The pD2 values for noradrenaline and adrenaline were 7.1 and 7.0, respectively, on the aortae of 20-month-old WKYs, and similar values were obtained on age-matched SHRs. On the aortae of 20-month old WKYs, the K(A) values for noradrenaline and adrenaline were 1.85 and 1.95 x 10(-6) M, and the receptor occupancies required for 50% maximum responses were 16 and 24%, respectively. There were lower affinities, by approximately twofold, but similar receptor reserves for noradrenaline and adrenaline on the aortae of age matched SHRs. In comparison with the aortae of 20-week-old WKYs and SHRs, there was a 5-fold loss of sensitivity to noradrenaline and adrenaline between 20 weeks and 20 months. Between 20 weeks and 20 months there was a 50-fold loss of affinity with ageing and a further twofold loss with hypertension/heart failure, and an increase in alpha1-adrenoceptor reserves for noradrenaline and adrenaline between 20 weeks and 20 months. There were no differences in the sensitivity and affinity, and minor changes in the alpha1-adrenoceptor reserves for noradrenaline and adrenaline between the aortae of 20-month-old WKYs and SHRs. In contrast there were major changes in these parameters in the ageing of the WKY aorta from 20 weeks to 20 months. There were no additional changes in the sensitivity and alpha1-adrenoceptor reserves, but a small additional change in affinity for noradrenaline and adrenaline in hypertension/heart failure on the aortae of 20 month-old SHRs. PMID- 11273018 TI - Imidazolines inhibit secretory responses of rat colonic mucosa to calcium dependent but not cyclic AMP-dependent secretagogues. AB - The purpose of this study was to investigate whether imidazolines have an anti secretory action on intestinal epithelial cells. Muscle-stripped preparations of rat colon and monolayers of T84 human colonic epithelial cells were set up in Ussing chambers for measurement of short-circuit current. In rat colon acetylcholine, histamine, vasoactive intestinal polypeptide and forskolin elicited secretory responses which were recorded as increases in short-circuit current. Secretory responses to acetylcholine were inhibited in a concentration dependent manner by the imidazolines phentolamine, idazoxan and clonidine. The effect of clonidine was not reversed by pre-incubation of mucosal preparations with yohimbine. Secretory responses to vasoactive intestinal polypeptide were unaffected by the three imidazolines. Phentolamine reduced responses of colonic mucosa to histamine but had no effect on responses to forskolin. Responses to vasoactive intestinal polypeptide and forskolin were significantly reduced in the presence of barium. In T84 cell monolayers phentolamine significantly reduced responses to acetylcholine. Three imidazolines, two with alpha-adrenoceptor antagonist properties and one with alpha-agonist properties, have anti-secretory effects in rat colonic mucosal preparations. The anti-secretory action appears to discriminate between calcium-dependent and cyclic AMP-dependent secretagogues, inhibiting the former but not the latter. PMID- 11273019 TI - Effect of chronic and acute administration of fluoxetine and its additive effect with morphine on the behavioural response in the formalin test in rats. AB - Serotonergic systems are involved in the central regulation of nociceptive sensitivity. Fluoxetine, a selective inhibitor of the reuptake of serotonin (5 hydroxytryptamine, 5-HT), was administered orally (0.16, 0.32, 0.8 mg kg(-1) daily for 7 days), intraperitoneally (0.04, 0.08, 0.16 mg kg(-1) day(-1) for 7 days and a single dose of 0.32 mg kg(-1)) and intracerebroventricularly (10 microg/rat) to rats and nociceptive sensitivity was evaluated using the formalin test (50 microL of 2.5% formalin injected subcutaneously). The effect of fluoxetine was also studied in the presence of 5,7-dihydroxytryptamine creatinine sulfate (5,7-DHT) and after co-administration with morphine. Oral (0.8 mg kg( 1)), intraperitoneal (0.16 and 0.32 mg kg(-1)) and intracerebroventricular (10 microg/rat) fluoxetine induced antinociception in the late phase of the formalin test. Furthermore, intrathecal administration of 5-HT (100 microg/rat) induced an analgesic effect. The analgesic effect of fluoxetine (0.16 and 0.32 mg kg(-1), i.p.) and 5-HT (100 microg/rat, i.t.) was abolished by pre-treatment with 5,7-DHT (100 microg/rat, i.t.). In addition, the analgesic effect of 5-HT (100 microg/rat, i.t.) was decreased by pre-treatment with naloxone (2 mg kg(-1), i.p.). Morphine (5 mg kg(-1), i.p.) induced analgesia that was increased by fluoxetine (0.32 mg kg(-1), i.p.). These results suggest that fluoxetine has an antinociceptive effect in tonic inflammatory pain through functional alteration of the serotonergic system and also potentiates the analgesic effect of morphine. PMID- 11273020 TI - Antioxidative effect of fluvastatin, an inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, on peroxidation of phospholipid liposomes. AB - The antioxidative effect of fluvastatin sodium (fluvastatin), a 3-hydroxy-3 methylglutaryl coenzyme A reductase inhibitor, on lipid peroxidation of phosphatidylcholine (PC) liposomes was investigated in various peroxidizing systems. Fluvastatin markedly inhibited the formation of thiobarbituric acid reactive substances in iron (II)-supported peroxidation of liposomes (IC50 = 1.2 x 10(-5) M). The order of magnitude of inhibition of each drug on the peroxidation was: butylated hydroxytoluene > fluvastatin > or = probucol >> pravastatin. Moreover, concentrations of fluvastatin ranging from 1 x 10(-6) to 1 x 10(-4) M inhibited peroxyl radical-mediated peroxidation of liposomes induced by water-soluble and lipid-soluble radical generators, 2,2'-azobis (2 amidinopropane) dihydro-chloride and 2,2'-azobis (2,4-dimethylvaleronitrile), respectively. However, pravastatin showed no effect against peroxyl radical mediated peroxidation. These results indicate that fluvastatin acted non enzymatically as an effective inhibitor against lipid peroxidation of PC liposomes and that the antioxidative effects of fluvastatin may be due to the scavenging action of fluvastatin on liposomal lipid peroxidation induced by peroxyl radicals generated in the aqueous and lipid phases. PMID- 11273021 TI - Effects of long-term pretreatment with isoproterenol on inotropic responsiveness to alpha-adrenoceptor stimulation: study in isolated perfused rat hearts. AB - The effects of chronic pretreatment with isoproterenol (5 mg kg(-1)) daily for 10 days on cardiac alpha-adrenergic responsiveness in Langendorff heart preparations were investigated. Isoproterenol pretreatment caused cardiac hypertrophy (29%) as shown by a significant increase in the ratio of ventricular dry weight to body weight. In preparations from isoproterenol-pretreated rats, both maximum increases in left ventricular systolic pressure and heart rate elicited by isoproterenol (10(-12) to 10(-4) M) were significantly reduced (the isoproterenol concentration producing 50% of the maximum positive inotropic and chronotropic responses was enhanced almost 32- and 4-fold, respectively), while the positive inotropic response to phenylephrine (10(-12) to 10(-4) M) was significantly enhanced (the phenylephrine concentration producing 50% of the maximum positive inotropic effect was reduced almost 100-fold), compared with saline-pretreated rats. In preparations from both groups, phenylephrine infusion induced non significant changes in heart rate and its positive inotropic response was reduced in the presence of propranolol (10(-7) M) in the perfusion medium. Even under beta-adrenoceptor blockade, the curve for the phenylephrine-induced positive inotropic effect remained shifted upward after isoproterenol pretreatment. Chronic isoproterenol pretreatment induces the expected cardiac beta-adrenoceptor desensitization while simultaneously enhancing the positive inotropic responsiveness to phenylephrine in Langendorff heart preparations. These findings support the hypothesis that cardiac alpha1-adrenoceptor stimulation may contribute to the maintenance of myocardial function under conditions in which beta-adrenoceptor function is compromised. PMID- 11273022 TI - Antioxidants, vitamin C and dithiothreitol, activate membrane-bound guanylate cyclase in PC12 cells. AB - Antioxidants and antioxidant enzymes are known to protect against cell death induced by reactive oxygen species. However, apart from directly quenching free radicals, little is known about the effect of antioxidants on hormone-activated second messenger systems. We previously found that antioxidants such as 17-beta estradiol and resveratrol activate membrane-bound guanylate cyclase GC-A, the receptor for atrial natriuretic factor (ANF), in PC12 cells. It is possible that other antioxidants may also activate membrane-bound guanylate cyclase GC-A. The aim of this study was to determine if dithiothreitol (DTT), vitamin C, and vitamin E activate membrane-bound guanylate cyclase GC-A in PC12 cells. The results showed that both DTT and vitamin C increased cGMP levels in PC12 cells, whereas vitamin E had no effect. DTT and vitamin C inhibited membrane-bound guanylate cyclase activity stimulated by ANF in PC12 cells. In contrast, DTT and vitamin C had no effect on soluble guanylate cyclase activity stimulated by substance P. Furthermore, NO synthase inhibitors L-NAME and aminoguanidine did not affect DTT- and vitamin C-stimulated guanylate cyclase activity. The results indicate that DTT and vitamin C, but not vitamin E, activate membrane-bound guanylate cyclase GC-A in PC12 cells. PMID- 11273023 TI - Inhibition of oxotremorine-induced desensitization of guinea-pig ileal longitudinal muscle in Ca2+-free conditions. AB - The aim of this study was to investigate the differences between oxotremorine induced and acetylcholine (ACh)-induced desensitization, particularly under Ca2+ free conditions, in guinea-pig ileal longitudinal muscle, and to elucidate the different mechanisms of desensitization that might exist between these two muscarinic agonists. Pretreatment of the tissue with 10(-7)-10(-5) M oxotremorine (desensitizing treatment) in normal Tyrode solution caused desensitization of the responses to ACh, as did the desensitizing treatment with ACh. However, Ca2+-free conditions significantly reduced oxotremorine-induced desensitization, contrary to the previous findings that Ca2+-free conditions enhanced ACh-induced desensitization. The desensitizing treatment with oxotremorine caused suppression of the responses to high K+ (tonic phase), as did the ACh treatment. Ca2+-free conditions removed this suppression, whereasthis condition enhanced ACh-induced suppression of the K+ response. A protein kinase C inhibitor, 1-(5 isoquinolinesulfonyl)-2-methylpiperazine (10(-4) M) had no effect on oxotremorine induced desensitization of the ACh response. The results suggest that a voltage gated Ca2+ channel was involved in oxotremorine-induced desensitization, as in ACh-induced desensitization, but that the process of inactivation of Ca2+ channels was different between oxotremorine and ACh, and that oxotremorine induced desensitization was due not only to Ca2+ channel, but also to other unknown factors. Protein kinase C did not participate in oxotremorine-induced desensitization. PMID- 11273024 TI - Preparation and characterization of biodegradable or enteric-coated microspheres containing the protease inhibitor camostat. AB - We have produced biodegradable or enteric-coated microspheres containing camostat mesylate, a protease inhibitor, using a water-oil-water emulsion solvent evaporation method. The characteristics of the microspheres were determined. When polylactic acid, a biodegradable polymer, was used as a wall material, the optimized microsphere obtained showed a loading efficiency of almost 95% and had a mean diameter of 30 microm. This microsphere showed a sustained-release profile, with nearly 25% of drug being released at seven days in a dissolution test. When hypromellose acetate succinate (AS-HG type, with a high content of succinyl group) was used as an enteric wall material, optimized microspheres showed a loading efficiency of almost 80%. In this case, pH 3.0 citrate buffer was used as an internal aqueous phase, and citrate buffer containing 0.5% polyvinylalcohol was used as the external aqueous phase. These microspheres showed a rapid release profile in pH 6.8 buffer, whereas the release was extremely slow in pH 1.2 buffer. Hypromellose acetate succinate microspheres were also prepared containing 10% (w/w) N-benzoyl-dl-arg-4-nitroanilide as a model substrate for trypsin, with or without 5% (w/w) camostat. These microspheres were incubated in pH 6 or 7 buffer containing trypsin at 37 degrees C. When camostat was included in the microspheres, the substrate was protected from attack by trypsin, while in the absence of camostat, the released substrate was immediately attacked by trypsin to produce the degradation product N-benzoyl-dl-arginine. PMID- 11273025 TI - A sensitive and reliable method for the detection of lipid peroxidation in biological tissues. AB - A simple, accurate and cost effective method has been designed for the determination of lipid peroxidation in biological tissue samples. The method was a modification and improvement on existing methods available for lipid peroxidation determination. Solid-phase extraction was used to separate the thiobarbituric acid-malondialdehyde complex from thiobarbituric acid-reactive substances and HPLC was performed using a C18 (Waters Spherisorb, 5 microm, 250 x 4.6 mm i.d.) column to achieve isolation of the complex. The procedure was validated with respect to linearity of calibration (0.998), precision, sensitivity and limits of quantitation (1 nmol mL(-1)) and detection (0.5 nmol mL(-1)). Resorcinol was used as an external standard. The method was tested by inducing free radical generation with a known free radical generator, quinolinic acid, in rat brain homogenate. The results showed that the method presented allowed detection of lipid peroxidation products at concentrations in the nanomolar (nM) range compared with the micromolar (microM) range detected by other methods, thus rendering it suitable for use with biological samples. In addition, the modified method allowed for detection of the purified lipid peroxidation products, thus eliminating the possibility of simultaneous detection of impurities that absorb at the same wavelength. PMID- 11273026 TI - Synthesis and in-vitro antimicrobial activity of new 1,2,4-triazoles. AB - We have described the synthesis of new 1,2,4-triazoles and have evaluated their antimicrobial profile. Antitubercular activity was determined in triplicate using the Lowenstein-Jensen medium. A loopful of Mycobacterium tuberculosis suspension was inoculated on the surface of each Lowenstein-Jensen media containing the test compounds (100, 10 or 1 microg mL(-1)). To evaluate in-vitro antibacterial activity, compounds (50, 5 or 0.5 microg) were evaluated against B. subtilis, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Staphylococcus typhi by the disc diffusion method. To evaluate antifungal activity Sabourauds Dextrose agar medium was used. Some of the compounds (5, 0.5 or 0.05 microg mL(-1)) were screened for activity against Aspergillus niger 88 and Aspergillus niger 90 and others were screened for activity against T. rubrum TR1, T. rubrum R6, T. rubrum R7 and T. mentagrophyte M1, using the cup plate method. Our results show that the triazoles with a pyrazine moiety at position 3 were more active as antitubercular and antifungal agents compared with the triazoles which had a pyrazine moiety at position 4 of the molecule. PMID- 11273027 TI - Stimulation of insulin release in rats by Die-Huang-Wan, a herbal mixture used in Chinese traditional medicine. AB - Die-Huang-Wan is a herbal mixture widely used in Chinese traditional medicine to treat diabetic disorders. We have investigated the effect of Die-Huang-Wan on plasma glucose concentration in-vivo. Die-Huang-Wan was administered orally (5.0, 15.0 or 26.0 mg kg(-1)) to three rat models. Wistar rats were used as the normal animal model, rats with insulin-resistance (induced by the repeated thrice daily injection of human long-acting insulin) were used as the non-insulin-dependent diabetic model, and streptozotocin-induced diabetic rats were used as the insulin dependent diabetic model. In normal rats, approximately 1 h after oral administration of Die-Huang-Wan the plasma glucose concentration decreased significantly in a dose-dependent manner, from 5 to 26.0 mg kg(-1). A similar effect was observed in rats with insulin-resistance. However, this effect was not observed in streptozotocin-induced diabetic rats, even at an oral dose of 26.0 mg kg(-1). These results suggested an insulin-dependent action, a view supported by the increase of plasma insulin-like immunoreactivity in normal rats receiving Die Huang-Wan. The results indicated that Die-Huang-Wan had an ability to stimulate the secretion of insulin and this preparation seemed helpful in improving the diabetic condition, especially hyperglycaemia in type-II diabetes. PMID- 11273029 TI - Detection of deoxyribonuclease I and II activities in Japanese quail oocytes. AB - Birds exhibit physiological polyspermy, i.e. numerous spermatozoa enter the germinal disc of an oocyte and form pronuclei during fertilisation. However, only one of them unites with the female pronucleus to form a zygote nucleus; the supernumerary spermatozoal nuclei degenerate at the early cleavage stages. To establish a factor responsible for spermatozoal degeneration, the presence of DNase activity was studied in vitro in extracts of Japanese quail oocytes using lambda DNA/HindIII as a substrate. The experimental conditions were designed to reveal the presence of either DNase I or DNase II activities, separately. Degradation of the substrate DNA was evaluated by electrophoresis on agarose gels stained with ethidium bromide. High activities of DNase I and DNase II were found in the germinal discs of the largest vitellogenic oocytes. DNase I activity was estimated to be about 3 x 10(-3) Kunitz units and DNase II about 4 x 10(-2) Kunitz units per germinal disc. DNase I activity in an oocyte seems to increase during oogenesis since DNA degradation by the extracts from the germinal discs of the largest vitellogenic oocytes was much higher than by those from previtellogenic and small vitellogenic oocytes. The presence of high DNase I and II activities in the largest vitellogenic oocytes would point to their role in degradation of DNA from supernumerary spermatozoa entering the ovum during polyspermic fertilisation in birds. The enzymes could be a factor, or one of the factors, in the late block to polyspermy in the cytoplasm of avian eggs. It is suggested here that the DNase activities might also be responsible for poor efficiency in obtaining transgenic birds by microinjection of exogenous DNA into the fertilised chick ovum. PMID- 11273028 TI - Role of nitric oxide in hypodipsia of rats with obstructive cholestasis. AB - Cholestasis is associated with the overproduction of nitric oxide (NO), and NO acts as an inhibitory mechanism when thirst is stimulated by water deprivation or by angiotensin II. Due to the presence of hypodipsia in the cholestatic condition, we have compared the rate of water intake between bile duct-ligated (cholestatic) and sham-operated rats. We have evaluated the effect of NO synthesis inhibition by N(G)-nitro-L-arginine (L-NNA, 10 mg kg(-1)/day) on the rate of water intake in cholestatic rats. The results showed that plasma alkaline phosphatase activity (a marker of liver damage) increased after bile-duct ligation, and that its elevation was partially (but significantly) prevented by treatment with L-arginine. A two-week bile-duct obstruction induced a significant decrease in the rate of water intake compared with sham-operated animals (35.87 +/- 1.45 vs 42.37 +/- 1.99 mL/day, P < 0.05). This effect was corrected by the daily administration of L-NNA. Surprisingly, L-arginine (200 mg kg(-1)/day) showed similar activity as L-NNA in cholestatic rats and increased water intake, but not in control animals. Systemic NO synthesis inhibition corrected the decrease in water intake observed in cholestatic rats. This suggests an important role for NO in the pathophysiology of hypodipsia in cholestatic subjects. The effect of chronic L-arginine administration observed in cholestatic rats but not seen in the control rats could be explained theoretically by the amelioration of cholestasis-induced liver damage by chronic L-arginine administration in bile duct-ligated rats. PMID- 11273030 TI - Mitogen-activated protein kinase regulates normal transition from metaphase to interphase following parthenogenetic activation in porcine oocytes. AB - The decrease in maturation-promoting factor (MPF) activity precedes that in mitogen-activated protein kinase (MAPK) activity after egg activation, but the cellular functions of this delayed inactivation of MAPK are still unclear. The present study was conducted to examine the essential role of MAPK activity for supporting the transition from metaphase to interphase in porcine oocytes matured in vitro. The increases in the phosphorylated forms of MAPK and the activities of MAPK and histone H1 kinase (H1K) were shown in oocytes arrested at the metaphase II (MII) stage. After additional incubation of MII-arrested oocytes in medium with added U0126, a specific inhibitor of MAPK kinase, 24% of oocytes completed the second meiotic division and underwent entry into interphase with pronucleus (PN) formation, but not second polar body (PB-2) emission. The intensities of the phosphorylated forms of MAPK and the activities of MAPK and H1K in matured oocytes treated with U0126 were significantly decreased by the treatment with U0126. Electrostimulation to induce artificial activation caused both H1K and MAPK inactivation; the inactivation of H1K preceded the inactivation of MAPK and sustained high levels of MAPK activity were detected during the period of PB-2 emission. However, the time sequence required for MAPK inactivation was significantly reduced by the addition of U0126 to the culture medium following electrostimulation, resulting in the dramatic inactivation of MAPK distinct from that of H1K. In these oocytes, PB-2 emission was markedly inhibited but little difference was found in the time course of PN formation compared with oocytes not treated with U0126. These findings suggest that the decrease in MAPK activity is partly involved in driving matured oocytes out of metaphase to induce PN development, and that the delayed MAPK inactivation after the onset of MPF inactivation in activated oocytes has a crucial role for PB-2 emission to accomplish the transition from meiosis to mitosis. PMID- 11273031 TI - The effect of various capacitation active compounds and capacitation time on the in vitro fertility and protein tyrosine phosphorylation profiles of bovine sperm. AB - In this paper the effects of capacitation and fertilisation stimulating compounds (heparin, caffeine, glucose, D-penicillamine, bovine serum (BOS), bovine serum albumin (BSA), polyvinyl alcohol (PVA)) were analysed in several in vitro fertilisation protocols. Attention was paid to the rate of penetrated oocytes, kinetics of penetration and to polyspermic fertilisation. Cryopreserved bovine sperm and in vitro matured bovine oocytes were used throughout all the fertilisation experiments. As detected in the first 8 h fertilisation experiment with non-incubated sperm, the supplementation of medium with heparin, BOS and glucose supported the fertilisation rate most effectively (100%), including the kinetics of pronuclei formation (52.4%). The absence of BOS resulted in a decreased fertilisation rate (62.7%) as well as a delay in pronuclei formation (13.6%), similar to that after substitution of heparin with caffeine (73.0% and 25.4%, respectively). The penetration rate in the control medium with BOS (without heparin and caffeine) was surprisingly high, especially in medium without glucose (62.2%). The positive effect of glucose on sperm penetration was observed mainly in a chemically defined medium with PVA. High polyspermy rates were observed throughout all experiments in the media containing heparin or caffeine and BOS as the macromolecular component. D-Penicillamine was not shown to be a fertilisation-stimulating molecule. However, as detected in the second experiment in which oocytes were fertilised with 5 h incubated sperm, its positive effect on the prolongation of a fertile life span of cryopreserved spermatozoa was significant. The presence of either caffeine or heparin in the fertilisation medium (FM) with BOS during sperm incubation induced tyrosine phosphorylation of an approximately 90 kDa protein, detected after 5 h of sperm incubation. The absence of BOS reduced tyrosine phosphorylation of this protein in fertilisation medium with heparin. The percentage of motile spermatozoa and those with intact acrosomes were monitored throughout all experiments. PMID- 11273032 TI - Expression and localisation of heat shock protein 70 in cultured bovine oocytes and embryos. AB - Effects of elevated in vitro temperature on in vitro produced early bovine embryos were analysed in order to determine its impact on the expression of heat shock protein 70 (hsp70). In vitro matured bovine oocytes, 2-cell and 8-cell embryos, and day 9 hatched blastocysts subjected to control and elevated temperature conditions were analysed by semiquantitative reverse transcription polymerase chain reaction methods for hsp70 mRNA expression. Results revealed the expression of hsp70 mRNA under control conditions and that early embryos can respond to heat stress by transcribing hsp70 mRNA. Confocal laser scanning microscopy used to localise the hsp70 protein in oocytes and embryos revealed that the distribution of hsp70 in the ooplasm of immature and mature oocytes is unaffected by exposure to elevated temperatures and that this protein was closely associated with the meiotic spindle, indicating its possible role in stabilising this structure. In 8-cell embryos derived under control conditions, hsp70 was evenly distributed in the cytoplasm but appeared as aggregates in some embryos exposed to elevated temperature. In heat-stressed hatched blastocysts, a more even distribution was noted following heat stress relative to corresponding controls, indicating their competence to respond to elevated temperature. PMID- 11273033 TI - Mammalian sperm molecules that are potentially important in interaction with female genital tract and egg vestments. AB - Fertilisation is a highly programmed process by which two radically different cells, sperm and egg, unite to form a zygote, a cell with somatic chromosome numbers. Development of the zygote begins immediately after sperm and egg haploid pronuclei come together, pooling their chromosomes to form a single diploid nucleus with the parental genes. Mammalian fertilisation is the net result of a complex set of molecular events which allow the capacitated spermatozoa to recognise and bind to the egg's extracellular coat, the zona pellucida (ZP), undergo the acrosome reaction, and fuse with the egg plasma membrane. Sperm-zona (egg) interaction leading to fertilisation is a species-specific carbohydrate mediated event which depends on glycan-recognising proteins (glycosyltransferases/glycosidases/lectin-like molecules) on sperm plasma membrane (receptors) and their complementary glycan units (ligands) on ZP. The receptor-ligand interaction event initiates a signal transduction pathway resulting in the exocytosis of acrosomal contents. The hydrolytic action of the sperm glycohydrolases and proteases released at the site of sperm-egg interaction, along with the enhanced thrust generated by the hyperactivated beat pattern of the bound spermatozoon, are important factors regulating the penetration of egg investments. This review focuses on sperm molecules believed to be important for the interaction with the female genital tract, passage through cumulus oophorus and attachment to ZP, induction of the acrosome reaction, secondary binding events, and passage through the ZP. An understanding of the expression and modifications of molecules thought to be important in multiple events leading to fertilisation will allow new strategies to block these modifications and alter sperm function. PMID- 11273034 TI - A gap-junction-mediated signal, rather than an external paracrine factor, predominates during meiotic induction in isolated mouse oocytes. AB - This study was carried out to compare the possible role of a secreted paracrine factor versus that of a gap-junction-transmitted signal in mediating meiotic induction in isolated mouse oocytes from PMSG-primed, immature mice. In the first set of experiments, oocyte-cumulus cell complexes (OCC) were pretreated for 3 h with 2 mM dbcAMP or FSH, washed, and the oocytes then cultured for 17-18 h in 40 microl drops containing either 300 microM dbcAMP or 4 mM hypoxanthine (HX). Each set of pretreated oocytes was cultured under three different conditions: (1) intact cumulus-cell-enclosed oocytes (CEO); (2) denuded oocytes (DO), cultured alone after removal of cumulus cells; and (3) co-cultured cumulus cells and oocytes (CC/DO), where the cumulus cells were removed in the same drop with a mouth-operated pipette and cultured alongside the oocytes. When pretreated with high dbcAMP or FSH, maturation was stimulated in CEO when cultured in either inhibitor (by 41.4-53.7%). Pretreatment failed to affect the maturation rate in DO. DO maturation was not altered appreciably by co-cultured cumulus cells when arrest was maintained with dbcAMP. However, an increase in maturation of 21-23% was observed in CC/DO in the HX-containing cultures that was not dependent on prior treatment with a meiosis-inducing stimulus. When DO were co-cultured with intact, FSH-treated OCC, there was no evidence of a positive factor secreted by the stimulated complexes, despite the fact that oocytes within the OCC were induced to resume maturation. In a second series of experiments the gap junction inhibitor, 18alpha-glycyrrhetinic acid (GA), was utilised. An initial experiment determined that GA dose-dependently blocked OCC metabolic coupling (0.2% coupling at 10 microM compared with 13.6% in controls). When HX-arrested CEO and DO were cultured for 17-18 h in medium containing increasing concentrations of GA, meiotic maturation was induced in CEO but not DO, suggesting that the cumulus cells provided a positive stimulus in the absence of functional gap junctional communication. No effect of GA was seen in dbcAMP-arrested oocytes. A kinetics experiment showed that when CEO were cultured in dbcAMP +/- FSH, meiotic induction was initiated after 3 h and germinal vesicle breakdown reached 60% by 6 h. When GA was added to the cultures at different times after the initiation of culture (0, 2, 3, 4 and 5 h), meiotic induction was immediately blocked. In addition, measurement of OCC coupling revealed that no reduction in coupling occurred during this induction period in the absence of GA. It is concluded that cumulus cells can secrete a positive factor, but that this is normally overridden by inhibitory influences transmitted through the gap junction pathway in intact complexes. Furthermore, upon exposure of complexes to a meiosis-inducing stimulus, a positive gap-junction-mediated signal now predominates to trigger germinal vesicle breakdown, and this signal is utilised throughout the induction period. PMID- 11273035 TI - A combination of calcium ionophore and puromycin effectively produces human parthenogenones with one haploid pronucleus. AB - Parthenogenetic activation with various combinations of the calcium ionophore A23187 and protein synthesis or phosphorylation inhibitors was investigated as a means of producing human parthenogenones with one haploid pronucleus. Unfertilised human aged oocytes exposed to 5 microM A23187 for 5 min were treated with 10 microg/ml puromycin (puromycin group, 46 oocytes) or 2 mM 6 dimethylaminopurine (DMAP group, 42 oocytes) for 5 h. Oocytes treated only with A23187 served as a control (control group, 40 oocytes). After washing the oocytes, they were incubated for up to 37 h. Evidence of activation (pronuclear formation) and cleavage was observed 18 h and 42 h after A23187 treatment, respectively. Activation rates in the puromycin and DMAP groups were significantly higher than in the control group (91% (42/46) and 77% (34/44) vs 20% (8/40), p < 0.05, respectively). In the puromycin group, 81% (34/42) of the activated oocytes showed one pronucleus with the second polar body (2ndPB), whereas none (0/34) of the activated oocytes in the DMAP group extruded the 2ndPB. The cleavage rate in the puromycin group was significantly lower than in the DMAP group (38% vs 68%, p < 0.05). The activated oocytes which had one pronucleus with the 2ndPB in the puromycin group showed a haploid set of chromosomes (10/13). In conclusion, the combination of A23187 and puromycin is effective for producing human parthenogenones with one haploid pronucleus. PMID- 11273036 TI - Activation of bovine oocytes matured in vitro by injection of bovine and human spermatozoa or their cytosolic fractions. AB - The aim of this study was to investigate whether bovine spermatozoa possess so called sperm factor in the cytosolic fraction (CF) which activates bovine oocytes, and whether bovine oocytes matured in vitro are activated by microinjection of CF extracted from spermatozoa of other species. In the first experiment, bovine and human spermatozoa were microinjected into ooplasm of bovine oocytes matured in vitro. Secondly, CF from bovine and human spermatozoa were injected into bovine oocytes. In the third, CF from human spermatozoa was injected into human unfertilised oocytes obtained 18-20 h after clinical intracytoplasmic sperm injection (ICSI). We found that microinjection of bovine spermatozoa into bovine oocytes induced oocyte activation, as shown by resumption of meiosis and formation of a female pronucleus, at a significantly higher rate than the bovine sham injection (63.0% vs 43.0%; p < 0.05). On the other hand, there was no significant difference in activation rate between the human sperm injection (35.9%) and the human sham injection (22.9%). Furthermore, microinjection of bovine sperm CF into bovine oocytes induced oocyte activation at a significantly higher rate than the human CF injection or sham injection (75.9% vs 14.8%, 20.4%; p < 0.01). Formation of a single female pronucleus and second polar body extrusion was observed in 95.1% of activated oocytes after bovine sperm CF injection. When human sperm CF was injected into human unfertilised oocytes, the activation rate was significantly higher than following sham injection (76.9% vs 44.0%; p < 0.05). These results indicate the presence of sperm factor in bovine sperm CF which activate bovine oocytes, and suggest the possibility that sperm factor has species-specificity at least between bovine and human. PMID- 11273037 TI - In vitro fertilisation of mouse oocytes reconstructed by transfer of metaphase II chromosomes results in live births. AB - The interaction between nucleus and cytoplasm can be explored through nuclear transfer. We describe here another tool to investigate this interaction: MII meiotic apparatus transfer (MAT) between mouse oocytes. In this study, the MII oocyte meiotic apparatus or spindle from C57BL/6 mice, a black strain, was transferred into an enucleated metaphase oocyte from Kunming mouse, a white strain. The results showed that the enucleation rate by treating oocytes with 3% sucrose was 100%, but the electrofusion efficiency was very low, with only 17.6% of reconstructed karyoplast-recipient cytoplasm pairs fused. When the fused oocytes were exposed to spermatozoa from C57BL/6 mice, 9 of 11 (82%) were fertilised. Eight reconstructed embryos at 1- to 4-cell stages were transferred into the oviducts of two synchronously pregnant Kunming strain fosters and one delivered two normal C57BL/6 offspring. This study indicates that MII meiotic apparatus or spindle sustains normal structure and function after micromanipulation and electrofusion. MAT provides a model for further research on the application of this technique to assisted human reproduction. PMID- 11273038 TI - Histochemical localisation of versican, aggrecan and hyaluronan in the developing condylar cartilage of the fetal rat mandible. AB - We investigated the histochemical localisation of versican, aggrecan and hyaluronan in the developing condylar cartilage of the fetal rat mandible at d 15 17 of gestation. At d 15 of gestation, immunostaining for versican was detected in the anlage of the future condylar process (condylar anlage), although the staining intensity showed a considerable regional variation. At d 16 of gestation, a metachromatically stained matrix firstly appeared in the condylar anlage. Aggrecan, hyaluronan and versican were simultaneously detected in this newly formed condylar cartilage. At d 17 of gestation, immunostaining for versican became restricted to the perichondrium and was barely detected in the cartilage. Colocalisation of versican and aggrecan was also seen in the cranial base cartilage at d 14 of gestation. These results indicate that although versican is replaced by aggrecan during the transition from prechondrogenic tissue to cartilage, both molecules were temporally colocalised in the newly formed cartilage. A hyaluronan-rich, low-versican area was identified in the posterior end of the condylar anlage during d 15-17 of gestation. The existence of this area is a unique structural feature of the developing condylar cartilage. PMID- 11273039 TI - Os incae: variation in frequency in major human population groups. AB - The variation in frequency of the Inca bone was examined in major human populations around the world. The New World populations have generally high frequencies of the Inca bone, whereas lower frequencies occur in northeast Asians and Australians. Tibetan/Nepalese and Assam/Sikkim populations in northeast India have more Inca bones than do neighbouring populations. Among modern populations originally derived from eastern Asian population stock, the frequencies are highest in some of the marginal isolated groups. In Central and West Asia as well as in Europe, frequency of the Inca bone is relatively low. The incidence of the complete Inca bone is, moreover, very low in the western hemisphere of the Old World except for Subsaharan Africa. Subsaharan Africans show as a whole a second peak in the occurrence of the Inca bone. Geographical and ethnographical patterns of the frequency variation of the Inca bone found in this study indicate that the possible genetic background for the occurrence of this bone cannot be completely excluded. Relatively high frequencies of the Inca bone in Subsaharan Africans indicate that this trait is not a uniquely eastern Asian regional character. PMID- 11273040 TI - Evidence of a hypermineralised calcified fibrocartilage on the human femoral neck and lesser trochanter. AB - Femoral neck fractures are a major cause of morbidity and mortality in elderly humans. In addition to the age-related loss of cancellous bone, changes to the microstructure and morphology of the metaphyseal cortex may be a contributing factor in osteoporotic hip fractures. Recent investigations have identified a hypermineralised tissue on the neck of the femur and trochanteric region that increases in fractional area with advancing age in both males (Boyce & Bloebaum, 1993) and females (Vajda & Bloebaum, 1999). The aim of this study was to determine if the hypermineralised tissue previously observed on the proximal femur is calcified fibrocartilage. Regional variations in the fractional area of hypermineralised tissue, cortical bone, and porosity of the cortical bone along the neck of the femur and lesser trochanter were also quantified. Comparison of back scattered electron and light microscope images of the same area show that regions of hypermineralised tissue correlate with the regions of calcified fibrocartilage from tendon and capsular insertions. The hypermineralised tissue and calcified fibrocartilage had similar morphological features such as the interdigitations of the calcified fibrocartilage into the bone, lacunar spaces, and distinctly shaped pores adjacent to the 2 tissues. Regions of the neck that did not contain insertions were covered with periosteum. There were no regional differences (P > 0.05) on the superior and inferior femoral neck in terms of the percentage area of hypermineralised calcified fibrocartilage, cortical bone, or cortical bone porosity. The lesser trochanter exhibited regional differences in the fractional area of hypermineralised calcified fibrocartilage (P = 0.007) and cortical bone (P = 0.007) but not porosity of the cortical bone (P > 0.05). The effects of calcified fibrocartilage on femoral neck periosteal expansion, repair, and mechanics are unknown, but may play a role in osteoporotic fractures and intracapsular fracture healing. PMID- 11273041 TI - Dermal fibroblasts participate in the formation of new muscle fibres when implanted into regenerating normal mouse muscle. AB - Both in vitro and in vivo studies have described the conversion of fibroblasts to myogenesis when in the presence of dysfunctional myogenic cells. Myogenic conversion of fibroblasts subjected to a normal, as opposed to a diseased muscle environment has only been reported in vitro. The primary aim of this work was to determine if fibroblasts can convert to a myogenic lineage and contribute to new fibre formation when implanted into the regenerating muscle of a normal mouse. Dermal fibroblasts were prepared from neonatal mouse skin and labelled prior to implantation with the fluorescent nuclear marker 4',6-diamidino-2-phenylindole (DAPI). Cells were implanted into muscles of host mice that had been subjected to either cold/crush or minced muscle injury. Some host muscles were x-irradiated to deplete the muscle of endogenous muscle precursor cells. Muscles were removed at 3 wk postimplantation and analysed both histologically and for the presence of DAPI labelled nuclei. Fibres containing DAPI labelled central nuclei indicated that the implanted cells had participated in the regenerative process. Mouse dermal fibroblasts therefore do contribute to muscle fibre formation in regenerating normal mouse muscle but the extent of their contribution is dependent on the nature of the trauma induced in the host muscle. The study also showed that regeneration was more successful in muscles which had not been irradiated, which is contrary to the previous studies where dermal fibroblasts were introduced into myopathic mouse muscle. PMID- 11273042 TI - Immunolocalisation of sodium channel NaG in the intact and injured human peripheral nervous system. AB - The voltage-gated 'glial' sodium channel NaG belongs to a distinct molecular class within the multi-gene family of mammalian sodium channels. Originally found in central and peripheral glia, NaG has since been detected in neurons in rat dorsal root ganglia (DRG) and may play a role in Schwann cell-axon interactions. We have studied the presence of NaG-like immunoreactivity in the intact and injured human peripheral nervous system using a specific affinity-purified antibody. Nerve fibres in normal and injured peripheral nerves and normal skin exhibited intense NaG-immunoreactivity. Numerous NaG-immunoreactive nerve fibres surrounded neuronal cell bodies within postmortem control DRG, and in DRG avulsed from the spinal cord (i.e. after traumatic central axotomy). There were no significant differences in the pattern of NaG immunostaining between control and avulsed DRG, or with delay after injury. Generally, the neuronal cell bodies were only very weakly immunoreactive to NaG, indicating that the NaG immunoreactivity was predominantly in Schwann cells/myelin. In accord, we demonstrated NaG immunostaining in cultured human and rat Schwann cells, and in distal nerve after wallerian degeneration. NaG thus appears to be a useful new marker for Schwann cells in the human PNS, and a role in neuropathy deserves investigation. PMID- 11273044 TI - Patterns of age-dependent changes in the numbers of lymph follicles and germinal centres in somatic and mesenteric lymph nodes in growing C57Bl/6 mice. AB - The timing of the first appearance of lymph follicles and germinal centres in various lymph nodes, and the ways in which numbers of these and IgM-synthesising cells increase within the nodes, were investigated in male and female C57Bl/6N mice aged from 4 d to 16 wk. The lymphoid organs examined were the Peyer's patches, spleen, somatic (submandibular, deep cervical, brachial, axillary, inguinal and popliteal) and visceral (mesenteric and lumbar) lymph nodes. Primary follicles appeared in most somatic lymph nodes 6 d after birth. The number of follicles per node then increased rather sharply in larger lymph nodes and slowly in smaller nodes, up to 28 d of age, reaching a level which varied according to the location of the node. Thereafter, the number of follicles in the somatic lymph nodes increased only slightly to moderately, reaching a peak or plateau at 8-12 wk. In the mesenteric (ileocaecal) nodes, primary follicles first appeared at 12 d, then increased linearly during the suckling period and after weaning to reach a plateau at 8 wk of age. Germinal centres appeared in the submandibular and mesenteric nodes at 28 d and their numbers increased consistently in the latter, while remaining low in the former. The impact of possible 'natural' exogenous antigen stimulation of the various lymph nodes was estimated from the presence of IgM-synthesising cells and germinal centres. Differences between the patterns of age-dependent changes in the numbers of lymph follicles observed in the somatic and mesenteric lymph nodes during their ontogeny are discussed in relation to differences in the magnitude of the exogenous antigen stimulatory effect. We also found that the variations in the numbers of lymph follicles produced in somatic lymph nodes at different locations during the first 28 d after birth reflected differences in the dimensions of the body regions drained by a particular somatic lymph node at this stage of development. PMID- 11273043 TI - Innervation of NADPH diaphorase-containing neurons correlated with acetylcholinesterase, tyrosine hydroxylase, and neuropeptides in the pigeon cloaca. AB - The motility of the avian cloaca is under neural control, but little is known about the neural network that accomplishes this function. This present study was designed to determine the distribution of nitric oxide-synthesising neurons in the pigeon cloaca by enzyme histochemistry for reduced nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d). NADPH-d-positive staining was seen in the neurons and fibres in the cloaca. The highest density of nerve fibres was noted in the coprodeum and the lowest in the proctodeum. In the coprodeum, NADPH d neurons were found singly, formed small groups of 2-10 neurons, or were seen in plexuses in the muscle layer, lamina propria, or around the arterioles. Several NADPH-d-positive neurons were also observed in the ganglia of the cloaca. NADPH-d fibres ran in the muscle layer, lamina muscularis mucosae and lamina propria, or surrounded blood vessels. The distribution pattern of acetylcholinesterase (AChE) stained neurons and fibres in the cloaca was similar to that of NADPH-d. Double staining for NADPH-d and AChE showed colocalisation of the 2 enzymes in many neurons of the cloaca. Tyrosine hydroxylase (TH)-immunoreactive nerve fibres originating outside the cloaca were also noted. In the urodeum and proctodeum, neurons or fibres positive for NADPH-d, AChE or TH were scattered in the lamina propria. Nerve fibres immunoreactive for calcitonin-gene related peptide, galanin, methionine-enkephalin, substance P, and vasoactive intestinal peptide were found sparsely in the cloaca. Our results demonstrate that nitrergic neurons constitute a subpopulation which is closely associated with the cholinergic system in the pigeon cloaca. PMID- 11273045 TI - Distribution of tracheal and laryngeal mucous glands in some rodents and the rabbit. AB - We used scanning electron microscopy to count the number of mucous gland openings in the tracheae and lower portion of the larynges of the rat, guinea pig, hamster, mouse and rabbit. Cells of the airway surface epithelium were removed by protease digestion better to visualise the gland openings. The distribution of glands was further studied by conventional histology and by PAS/Alcian blue staining of whole mounts. In all rodent species, gland openings in the larynx occurred with a frequency of 1-2 per mm2. Mice had no gland openings in their tracheae, and hamsters, only a handful. Rat tracheae contained 126+/-42 gland openings (+/-S.D.; n = 6) at a frequency of approximately 0.6 per mm2 at the top of the trachea and approximately 0.15 per mm2 at the bottom. Guinea pig tracheae contained 153+/-90 gland openings (+/-S.D.; n = 5), with 54% being in the top 40% of the trachea. In both rat and guinea pig, tracheal glands were found in the ventral aspect between the cartilaginous rings, and were absent from the dorsal membranous portion. Gland openings in most species were simple circles of approximately 50 microm diameter. However, glands in the rat trachea generally opened obliquely into shallow (approximately 20 microm deep) oval troughs (approximately 150 x 75 microm), which had their long axes oriented from head to tail. In the rabbit, there was no evidence of tracheal or laryngeal glands histologically. However, the tracheal and laryngeal surfaces contained numerous pits (approximately 30 microm diameter) distributed evenly over and between cartilages at a frequency of approximately 4 per mm2. These may correspond to the 'nests' of goblet cells described by others. PMID- 11273046 TI - Fibrocartilage in the transverse ligament of the human acetabulum. AB - Biomechanical experiments on isolated hip joints have suggested that the transverse ligament acts as a bridle for the lunate articular surface of the acetabulum during load bearing, but there are inherent limitations in such studies because the specimens are fixed artificially to testing devices and there are no modifying influences of muscle pull. Further evidence is thus needed to substantiate the theory. Here we argue that if the horns of the lunate surface are forced apart under load, the ligament would straighten and become compressed against the femoral head. It would thus be expected to share some of the features of tendons and ligaments that wrap around bony pulleys and yet previous work has suggested that the transverse ligament is purely fibrous. Transverse ligaments were removed from 8 cadavers (aged 17-39 y) and fixed in 90% methanol. Cryosections were immunolabelled with antibodies against collagens (types I, II, III, VI), glycosaminoglycans (chondroitins 4 and 6 sulphate, dermatan sulphate, keratan sulphate) and proteoglycans (aggrecan, link protein, versican, tenascin). A small sesamoid fibrocartilage was consistently present in the centre of each transverse ligament, near its inner surface at the site where it faced the femoral head. Additionally, a more prominent enthesis fibrocartilage was found at both bony attachments. All fibrocartilage regions, in at least some specimens, labelled for type II collagen, chondroitin 6 sulphate, aggrecan and link protein, molecules more typically associated with articular cartilage. The results suggest that the ligament should be classed as containing a 'moderately cartilaginous' sesamoid fibrocartilage, adapted to withstanding compression. This supports the inferences that can be drawn from previous biomechanical studies. We cannot give any quantitative estimate of the levels of compression experienced. All that can be said is that the ligament occupies an intermediate position in the spectrum of fibrocartilaginous tissues. It is more cartilaginous than some wrap-around tendons at the wrist, but less cartilaginous than certain other wrap-around ligaments, e.g. the transverse ligament of the atlas. PMID- 11273048 TI - Gluteus minimus: observations on its insertion. AB - In 17 adult and 3 full term fetuses the hip joints were dissected to expose the insertion of gluteus minimus. An attachment of the deep surface of the tendon as it passes over the hip joint capsule was found in every case. Histological examination of this insertion confirmed the presence of short intramuscular tendons firmly anchoring the tendon to the capsule. It is suggested that this attachment retracts the capsule during hip joint motion, thereby preventing capsular entrapment. PMID- 11273047 TI - Specialised cell types in the chorioallantoic membrane express carbonic anhydrase during chick embryogenesis. AB - The expression of carbonic anhydrase in the chorioallantoic membrane (CAM) of the chick embryo was investigated by means of the histochemical localisation of the enzyme catalytic sites and the immunohistochemical identification of its isoenzymatic forms. The results show that carbonic anhydrase is developmentally expressed in a subset of cells both in the ectodermal and the endodermal epithelium. The distribution patterns from both methodological approaches indicated that carbonic anhydrase is a marker of the villus cavity cells and the mitochondria-rich cells in the ectodermal and the endodermal epithelium, respectively. Such a cell-specific pattern of the enzyme expression provides a further contribution to characterising the heterogeneous cell population of the chick CAM and supports specific functional involvement for the distinct cell types in CAM-mediated processes, such as calcium transport, maintenance of acid base balance and water and electrolyte reabsorption, during chick embryogenesis. PMID- 11273049 TI - Carpal bone movements in gripping action of the giant panda (Ailuropoda melanoleuca). AB - The movement of the carpal bones in gripping was clarified in the giant panda (Ailuropoda melanoleuca) by means of macroscopic anatomy, computed tomography (CT) and related 3-dimensional (3-D) volume rendering techniques. In the gripping action, 3-D CT images demonstrated that the radial and 4th carpal bones largely rotate or flex to the radial and ulnar sides respectively. This indicates that these carpal bones on both sides enable the panda to flex the palm from the forearm and to grasp objects by the manipulation mechanism that includes the radial sesamoid. In the macroscopic observations, we found that the smooth articulation surfaces are enlarged between the radial carpal and the radius on the radial side, and between the 4th and ulnar carpals on the ulnar side. The panda skilfully grasps using a double pincer-like apparatus with the huge radial sesamoid and accessory carpal. PMID- 11273050 TI - Fourier analysis methodology of trabecular orientation measurement in the human tibial epiphysis. PMID- 11273051 TI - Biphalangeal fifth toe: an increasingly common variant? PMID- 11273052 TI - An overstretched hypothesis? PMID- 11273053 TI - Prevention of hypertension in adulthood by breastfeeding? PMID- 11273054 TI - Risk management in hypertrophic cardiomyopathy. PMID- 11273055 TI - Targeting voracious appetite of malaria-infected red-blood cell. PMID- 11273056 TI - Rehabilitation for patients with Parkinson's disease. PMID- 11273057 TI - Environmental public-health surveillance systems for chemical incidents. PMID- 11273058 TI - Scotland the brave: collective responsibility for personal care for the elderly and for the young disabled. PMID- 11273059 TI - Early nutrition in preterm infants and later blood pressure: two cohorts after randomised trials. AB - BACKGROUND: Despite data relating body size in early life to later cardiovascular outcomes, the hypothesis that nutrition affects such outcomes has not been established. Breastfeeding has been associated with lower blood pressure in later life, but previous studies have not controlled for possible confounding factors by using a randomised design with prospective follow-up. We undertook such a study to test the hypothesis that early diet programmes blood pressure in later life in children randomly assigned different diets at birth. METHODS: Blood pressure was measured at age 13-16 years in 216 (23%) of a cohort of 926 children who were born prematurely and had participated at birth in two parallel randomised trials in five neonatal units in the UK. Dietary interventions were: donated banked breastmilk versus preterm formula and standard term formula versus preterm formula. FINDINGS: Children followed up at age 13-16 years were similar to those not followed up in terms of social class and anthropometry at birth. Mean arterial blood pressure at age 13-16 years was lower in the 66 children assigned banked breastmilk (alone or in addition to mother's milk) than in the 64 assigned preterm formula (mean 81.9 [SD 7.8] vs 86.1 [6.5] mm Hg; 95% CI for difference -6.6 to -1.6; p=0.001). In non-randomised analyses, the proportion of enteral intake as human milk in the neonatal period was inversely related to later mean arterial pressure (beta=-0.3 mm Hg per 10% increase [95% CI -0.5 to 0.1]; p=0.006). No differences were found in the term formula (n=44) versus preterm formula (n=42) comparison. INTERPRETATION: Breastmilk consumption was associated with lower later blood pressure in children born prematurely. Our data provide experimental evidence of programming of a cardiovascular risk factor by early diet and further support the long-term beneficial effects of breastmilk. PMID- 11273060 TI - Clinical picture: occult ectopic ACTH syndrome. PMID- 11273061 TI - Relation between severity of left-ventricular hypertrophy and prognosis in patients with hypertrophic cardiomyopathy. AB - BACKGROUND: A previous study suggested that severe left-ventricular hypertrophy (maximum wall thickness > or = 30 mm) in patients with hypertrophic cardiomyopathy is associated with a risk of sudden cardiac death sufficient to warrant consideration for implantation of a cardioverter defibrillator (ICD). However, the prognostic significance of left-ventricular hypertrophy in relation to other clinical risk factors is poorly characterised. METHODS: We studied 630 patients consecutively referred to one hospital in London, UK (mean age 37 years [SD 16]; 382 male; mean follow-up 59 months). Patients underwent two-dimensional and doppler echocardiography, upright exercise testing, and Holter monitoring. FINDINGS: 39 patients died suddenly or had an appropriate ICD discharge; nine died from progressive heart failure; 11 from other cardiovascular causes and 23 from non-cardiac causes. There was a trend towards higher probability of sudden death or ICD discharge with increasing wall thickness (p=0.029, relative risk per 5 mm increment 1.31 [95% CI 1.03-1.66]). Of the 39 patients who died suddenly or had an ICD discharge, ten had a wall thickness of 30 mm or more. Patients with wall thickness of 30 mm or more had higher probability of sudden death or ICD discharge than patients with wall thickness less than 30 mm (p=0.049, 2.07 [1.00 4.25]. When considered together, the number of additional risk factors (one to three) was a better predictor of risk of sudden death or ICD discharge than wall thickness (p=0.0001, relative risk per additional factor 2.00 [1.43-2.79] vs p=0.058, 1.26 per 5 mm increment [0.99-1.60]). There was no relation between the pattern of hypertrophy and survival. INTERPRETATION: The risk of sudden death associated with a wall thickness of 30 mm or more in patients without other risk factors is insufficient to justify aggressive prophylactic therapy. Most sudden deaths occurred in patients with wall thickness less than 30 mm, so the presence of mild hypertrophy cannot be used to reassure patients that they are at low risk. PMID- 11273062 TI - Association between premature mortality and hypopituitarism. West Midlands Prospective Hypopituitary Study Group. AB - BACKGROUND: Four retrospective studies have reported premature mortality in patients with hypopituitarism with standard mortality ratios (SMRs) varying between 1.20 and 2.17. Patients with hypopituitarism have complex endocrine deficiencies, and the mechanisms underpinning any excess mortality are unknown. Furthermore, the suggestion has emerged that endogenous growth-hormone deficiency might account for any excess mortality. We aimed to clarify these issues by doing a large prospective study of total and specific-cause mortality in patients with hypopituitarism. METHODS: We followed up 1014 UK patients (514 men, 500 women) with hypopituitarism from January, 1992, to January, 2000. 573 (57%) patients had non-functioning adenomas, 118 (12%) craniopharyngiomas, and 93 (9%) prolactinomas. SMRs were calculated as the ratio of observed deaths to the number of deaths in an age-matched and sex-matched UK population. FINDINGS: The number of observed deaths was 181 compared with the 96.7 expected (SMR 1.87 [99% CI 1.62 2.16], p<0.0001). Univariate analysis indicated that mortality was higher in women (2.29 [1.86-2.82]) than men (1.57 [1.28-1.93], p=0.002), in younger patients, in patients with an underlying diagnosis of craniopharyngioma (9.28 [5.84-14.75] vs 1.61 [1.30-1.99], p<0.0001), and in the 353 patients treated with radiotherapy (2.32 [1.71-3.14] vs 1.66 [1.30-2.13], p=0.004). Excess mortality was attributed to cardiovascular (1.82 [1.30-2.54], p<0.0001), respiratory (2.66 [1.72-4.11], p<0.0001), and cerebrovascular (2.44 [1.58-4.18], p<0.0001) causes. There was no effect of hormonal deficiency on mortality, except for gonadotropin deficiency, which, if untreated was associated with excess mortality (untreated 2.97 [2.13-4.13] vs treated 1.42 [0.97-2.07], p<0.0001). Multiple regression analyses identified age at diagnosis, sex, a diagnosis of craniopharyngioma, and untreated gonadotropin deficiency as independent significant factors affecting mortality. INTERPRETATION: Patients with hypopituitarism have excess mortality, predominantly from vascular and respiratory disease. Age at diagnosis, female sex, and above all, craniopharyngioma were significant independent risk factors. Specific endocrine-axis deficiency, with the exception of untreated gonadotropin deficiency, does not seem to have a role. PMID- 11273063 TI - Effect of highly active antiretroviral therapy on diagnoses of sexually transmitted diseases in people with AIDS. AB - BACKGROUND: There has been an increase in high-risk sexual behaviour and sexually transmitted diseases (STD) during the time period when highly active antiretroviral therapy (HAART) became widely available. We examined whether taking HAART increased the risk of acquiring an STD--an epidemiological marker of unsafe sex--in people with AIDS. METHODS: We did a computerised match of people in the San Francisco STD and AIDS registries. People with AIDS who were diagnosed before 1999 and alive in November, 1995, or later, were classified as having had an STD after AIDS diagnosis or not having had an STD after AIDS diagnosis. We used a Cox proportional hazards model to see whether use of antiretroviral therapy was associated with acquiring an STD after AIDS, after adjustment for sex, age, race, HIV-1 risk category, and CD4 count at AIDS diagnosis. FINDINGS: People with AIDS who had had HAART showed an independent increase in the risk of developing an STD (hazard ratio 4.10; 95% CI 2.84-5.94). Americans of African origin, younger age, and higher CD4 count at AIDS diagnosis were also associated with acquiring an STD after AIDS. The number of people living with AIDS who acquired an STD increased over time from 60 (0.66%) in 1995 to 113 (1.32%) in 1998 (p<0.001). INTERPRETATION: We have shown that people on HAART are more likely to develop an STD, an epidemiological marker of unsafe sex. More intensive risk-reduction counselling and STD screening for people with AIDS is needed. PMID- 11273064 TI - Association between polymorphism in regulatory region of gene encoding tumour necrosis factor alpha and risk of Alzheimer's disease and vascular dementia: a case-control study. AB - BACKGROUND: Deposition of beta-amyloid in the brains of patients with Alzheimer's disease is thought to precede a chain of events that leads to an inflammatory response by the brain. We postulated that genetic variation in the regulatory region of the gene for the proinflammatory cytokine tumour necrosis factor alpha (TNF-alpha) leads to increased risk of Alzheimer's disease and vascular dementia. METHODS: A polymorphism in the regulatory region of the TNF-alpha gene was analysed in a case-control study. The polymorphism (C-850T) was typed in 242 patients with sporadic Alzheimer's disease, 81 patients with vascular dementia, 61 stroke patients without dementia, and 235 normal controls. These groups of individuals were also genotyped for the apolipoprotein E polymorphism, and the vascular dementia and stroke groups were typed at the HLA-DR locus. FINDINGS: The distribution of TNF-alpha genotypes in the vascular dementia group differed significantly from that in the stroke and normal control groups, giving an odds ratio of 2.51 (95% CI 1.49-4.21) for the development of vascular dementia for individuals with a CT or TT genotype. Logistic regression analysis indicated that the possession of the T allele significantly increased the risk of Alzheimer's disease associated with carriage of the apolipoprotein E epsilon4 allele (odds ratio 2.73 [1.68-4.44] for those with apolipoprotein E epsilon4 but no TNF-alpha T, vs 4.62 [2.38-8.96] for those with apolipoprotein E epsilon4 and TNF-alpha T; p=0.03). INTERPRETATION: Possession of the TNF-alpha T allele significantly increases the risk of vascular dementia, and increases the risk of Alzheimer's disease associated with apolipoprotein E. Although further research is needed, these findings suggest a potential role for anti-inflammatory therapy in vascular dementia and Alzheimer's disease, and perhaps especially in patients who have had a stroke. PMID- 11273065 TI - Leg pain in a patient with chronic hepatitis C. PMID- 11273066 TI - Congenital absence of the inferior vena cava: a rare risk factor for idiopathic deep-vein thrombosis. AB - Congenital absence of the inferior vena cava (AIVC) is a rare vascular defect, commonly reported as a fortuitous finding. The presence of AIVC in patients with DVT is underestimated because AIVC cannot be detected by compression B-mode ultrasonography. By use of computed tomography, we diagnosed four cases of AIVC in young patients with idiopathic DVT over a 5 year period. Based on the occurrence of DVT in patients below 30 years in our area during the same period, we estimate that AIVC is present in about 5% of cases of DVT in young patients. Computed tomography or angiography should be used for the diagnosis of idiopathic DVT in young patients. PMID- 11273067 TI - Herpes simplex virus 1 transmission through corneal transplantation. AB - Genetic characterisation of herpes simplex virus type 1 (HSV-1) DNA isolated from a donor cornea before and after corneal transplantation demonstrated the transmission of HSV-1 through transplantation. This study is the first to provide conclusive evidence for the transmission of HSV-1 by penetrating keratoplasty with subsequent reactivation of donor-derived HSV-1 in the transplanted cornea. PMID- 11273068 TI - Risk of clinical pelvic inflammatory disease attributable to an intrauterine device. AB - Use of the intrauterine device (IUD) is avoided because of perceived risk of pelvic inflammatory disease (PID) associated with sexually transmitted Infections (STI). Calculation of the risk of clinical pelvic inflammatory disease showed that the estimated risk was low (0.15%), even with a high STI prevalence. This estimated risk argues for making IUDs more available. PMID- 11273069 TI - Genetic-susceptibility factor and malignant mesothelioma in the Cappadocian region of Turkey. AB - Erionite present in stones used to build the villages of Karain and Tuzkoy, Turkey, mined from nearby caves, is purported to cause mesothelioma in half of the villagers. We constructed genetic epidemiology maps to test whether some villagers were genetically predisposed to mesothelioma. Analysis of a six generation extended pedigree of 526 individuals showed that mesothelioma was genetically transmitted, probably in an autosomal dominant way. This finding should lead to preventive strategies to lower the incidence of mesothelioma in future generations, and close monitoring of high-risk individuals might allow early detection and cure. PMID- 11273071 TI - Fatal aplastic anaemia associated with clopidogrel. AB - Clopidogrel, an inhibitor of platelet aggregation, was initially thought to be free of the side-effects of ticlopidine. We describe a man who developed aplastic anaemia after 5 months of treatment with clopidogrel. There were no other plausible causes. We suggest that his fatal aplastic anaemia might have been induced by clopidogrel. PMID- 11273070 TI - Impact of 24 hour critical care physician staffing on case-mix adjusted mortality in paediatric intensive care. AB - The 24 h availability of intensive care consultants (intensivists) has been shown to improve outcomes in adult intensive care units (ICU) in the UK. We tested whether such availability would improve standardised mortality ratios when compared to out-of-hours cover by general paediatricians in the paediatric ICU setting of a medium-income developing country. The standardised mortality ratio (SMR) improved significantly from 1.57 (95%CI 1.25-1.95) with non-specialist care to 0.88 (95%CI 0.63-1.19) with intensivist care (rate ratio 0.56, 95% CI 0.47 0.67). Mortality odds ratio decreased by 0.234, 0.246 and 0.266 in the low, moderate and high-risk patients. 24 h availability of intensivists was associated with improved outcomes and use of resources in paediatric intensive care in a developing country. PMID- 11273072 TI - India tries to rebuild 600,000 lives after earthquake. PMID- 11273073 TI - Evidence-based prescribing made simple. PMID- 11273074 TI - New US HIV treatment guidelines urge a more conservative approach. PMID- 11273075 TI - What next after the first transgenic monkey? PMID- 11273076 TI - Bush's debut: playing both sides on the abortion issue. PMID- 11273077 TI - South Africa's AIDS activists say new neviripine programme is not enough. PMID- 11273078 TI - Australian government loosens its grip on the pharmaceutical industry. PMID- 11273079 TI - Brazil and USA at loggerheads over production of generic antiretrovirals. PMID- 11273080 TI - Child-injury death rates--do international comparisons help? PMID- 11273081 TI - Reliable assessment of the effects of treatment on mortality and major morbidity, II: observational studies. AB - Observational studies and randomised trials can contribute complementary evidence about the effects of treatment on mortality and on major non-fatal outcomes. In particular, observational studies have an important role in the identification of large adverse effects of treatment on infrequent outcomes (ie, rare, but serious, side-effects) that are not likely to be related to the indications for (or contraindications to) the treatment of interest. Such studies can also provide useful information about the risks of death and disability in particular circumstances that can help to generalise from clinical trials to clinical practice. But, due to their inherent potential for moderate or large biases, observational studies have little role in the direct assessment of any moderate effects of treatment on major outcomes that might exist. Instead, sufficiently large-scale evidence from randomised trials is needed to assess such treatment effects appropriately reliably. Wider appreciation of the different strengths and weaknesses of these two types of epidemiological study should increase the likelihood that the most reliable evidence available informs decisions about the treatments doctors use--and patients receive--for the management of a wide range of life-threatening conditions. PMID- 11273082 TI - The uses of error: the complexity of general practice. PMID- 11273083 TI - Bristol, Shipman, and clinical governance: Shewhart's forgotten lessons. AB - During the past century, manufacturing industry has achieved great success in improving the quality of its products. An essential factor in this success has been the use of Walter A Shewhart's pioneering work in the economic control of variation, which culminated in the development of a simple yet powerful graphical method known as the control chart. This chart classifies variation as having a common cause or special cause and thus guides the user to the most appropriate action to effect improvement. Using six case studies, including the excess deaths after paediatric cardiac surgery seen in Bristol, UK, and the activities of general practitioner turned murderer Harold Shipman, we show a central role for Shewhart's approach in turning the rhetoric of clinical governance into a reality. PMID- 11273084 TI - The heart remembers: clinical implications. PMID- 11273085 TI - Glucose tolerance in adults after prenatal exposure to famine. PMID- 11273086 TI - Laparoscopic cholecystectomy. PMID- 11273087 TI - Galactorrhoea, hyperprolactinaemia, and protease inhibitors. PMID- 11273088 TI - Galactorrhoea, hyperprolactinaemia, and protease inhibitors. PMID- 11273089 TI - Galactorrhoea, hyperprolactinaemia, and protease inhibitors. PMID- 11273090 TI - Galactorrhoea, hyperprolactinaemia, and protease inhibitors. PMID- 11273091 TI - Difficulties in polio eradication. PMID- 11273092 TI - Difficulties in polio eradication. PMID- 11273093 TI - Difficulties in polio eradication. PMID- 11273094 TI - Difficulties in polio eradication. PMID- 11273095 TI - Difficulties in polio eradication. PMID- 11273096 TI - Fetal stimulation and activity state. PMID- 11273097 TI - Guiding hands of our teachers. PMID- 11273099 TI - Guiding hands of our teachers. PMID- 11273098 TI - Guiding hands of our teachers. Hand-hygiene Liaison Group. PMID- 11273101 TI - Facing the biological weapons threat. PMID- 11273100 TI - Guidance on rosiglitazone for type 2 diabetes mellitus. PMID- 11273102 TI - From protocol into practice: who needs the research? PMID- 11273103 TI - Politics and public health: democracy--or what? PMID- 11273104 TI - Circadian rhythm in ischaemic heart disease. PMID- 11273106 TI - Silent gastroesophageal reflux in patients with bronchial asthma. AB - Gastroesophageal reflux (GER) is a common occurrence in patients with asthma. We performed a prospective study to detect GER in patients with asthma using pH monitoring. Twenty consecutive patients (mean age 48 +/- 14 yr, range 23-70; 13 men) with asthma of unexplained etiology were evaluated. Esophageal and gastric pH were studied both while introducing and pulling out pH probe. Upper GI endoscopy was done in all the patients before doing pH monitoring. Esophageal pH was recorded at 25 cm, 30 cm and 35 cm from the incisors. Mean esophageal pH values while introducing pH probe were 2.9 +/- 1.08 (0-5.1), 2.5 +/- 1.2 (0-5) and 1.6 +/- 1.5 (0-4.6) at 25 cm, 30 cm and 35 cm from incisors respectively. The gastric pH was 0.5 +/- 0.4 (0-2). While pulling out pH probe, pH values were 1.5 +/- 1.4 (0-4.7), 2.4 +/- 1.5 (0-5.1) and 2.9 +/- 1.4 (0-5.4) at 35 cm, 30 cm, and 25 cm from incisors respectively. Of 20 patients, 19 had pH of 4 or lower at 25 cm, 30 cm and 35 cm from incisors. The mean pH value was lower at 35 cm than at 30 cm and 25 cm (1.6 +/- 1.5 vs 2.5 +/- 1.2 and 2.9 +/- 1.08, p < 0.04). In conclusion majority of adults with asthma have silent GER as detected on pH monitoring. PMID- 11273105 TI - Prognostic significance of thyroid antibodies in hyperthyroid patients treated with antithyroid drugs. AB - Lymphocytic infiltration of the thyroid gland in patients with hyperthyroidism is associated with the presence of serum antithyroidal microsomal antibodies (TMA) and serum antithyroglobulin antibodies (TGA). The aim of this study was to evaluate the clinical significance of TMA and TGA during and after treatment of hyperthyroidism with antithyroidal drugs. One hundred and fifty-four hyperthyroid patients were treated for 18 months with methimazole and then followed up for 18 months or more (mean, 24.8 +/- 12.6 months). Patients were classified into three group. group I, patients negative for TGA and TMA before and during 18 months of treatment, group II patients positive for TMA but negative for TGA before and during 18 months treatment and group III patients who were positive for both TGA and TMA before and during treatment. The relapse rates after discontinuation of treatment in these group were 44.7% (17 of 38), 29% (18 of 62) and 11.1% (6 of 54), respectively. The value in group I was significantly higher than that in group III (P < 0.01). These results show that presence of TMA and TGA influence the prognosis of patients with hyperthyroidism treated with methimazole with regard to relapse. Those patients who had both antibodies were least likely to have a relapse and those who had neither antibody before and during treatment were most likely to have a relapse of hyperthyroidism. PMID- 11273107 TI - Respiratory symptoms in Indian children exposed to different cooking fuels. AB - Smoke emission from fuels is an important source of indoor air pollution. Children spend considerable time indoors. It is therefore important to determine whether air contaminants from indoor air sources affect incidence of respiratory illness, cause symptoms and changes in pulmonary function status in them. Two hundred children in the age group of 7-15 were selected randomly. They were stratified according to the fuel used in their homes and respiratory symptoms were inquired from them according to a questionnaire recommended by the American Thoracic Society. The most symptomatic children were those whose households used kerosene (52%) and mixed fuels (46%) although different symptoms were present in varying extent in all 4 groups of children. Cough, cold, congestion or phlegm for one week or more occurred more frequently with mixed fuel use followed by kerosene. The present study thus showed that mixed fuel and kerosene fuel had worst effects on respiratory system in children whose households used these fuels. PMID- 11273108 TI - A comparative evaluation of pharmacokinetics of conventional and slow-release carbamazepine formulation in newly treated patients of epilepsy: a random evaluation. AB - Study was conducted to compare the pharmacokinetic profile of conventional and slow-release carbamazepine formulations in Indian epileptic patients. Twenty consecutive untreated patients of partial seizures were randomly assigned to receive either conventional carbamazepine (200 mg thrice a day) or slow-release carbamazepine formulation (200 mg thrice a day), 10 patients in each group. The serum carbamazepine concentrations were measured on 10th day and 20th day of treatment. The blood samples were collected before the morning dose. In the conventional treatment group five patients experienced side effects as compared to two in the slow-release group. On 10th day mean serum carbamazepine levels were significantly higher in conventional group (8.27 +/- 1.39 micrograms/ml) in comparison to slow release group (4.28 +/- 3.89 micrograms/ml). The difference was statistically significant (p < 0.05). On 20th day carbamazepine levels fell significantly in conventional group only (8.27 +/- 1.39 micrograms/ml to 5.76 +/- 2.32 micrograms/ml, p < 0.05). At this stage the difference in mean carbamazepine levels of two groups became insignificant (p < 0.05). In conclusion, controlled release formulations provide more steady serum concentrations of carbamazepine along with better tolerability. PMID- 11273109 TI - Exercise-induced ventricular arrhythmias in congestive heart failure and role of ACE inhibitors. AB - Ventricular arrhythmias are considered to be related to left ventricular (LV) dysfunction. ACE inhibitors though improve LV function their beneficial role on exercise-induced ventricular arrhythmias is not established. To study the effects of ACE inhibitors on exercise capacity vis-a-vis their role on exercise-induced ventricular arrhythmias, 25 patients of congestive heart failure (CHF) of various etiologies in NYHA Class II and III were subjected to a prospective randomised controlled trial. The control group comprising of 12 patients received conventional treatment (digitalis and diuretics) and the test group was given enalapril/captopril in addition as tolerated. They were followed up for 3 months. Exercise testing on treadmill and monitoring of clinical and biochemical parameters were done at the beginning and end of study in all cases. Ventricular arrhythmias observed during exercise and post-exercise for 10 minutes was analysed using Lown's grading for frequency and severity of ventricular arrhythmia. The mean exercise duration showed significant improvement on ACE inhibitor as compared to the control group (p < 0.05) however there was no significant change in the grades of arrhythmia. Serum electrolytes and other bio chemical parameter were within normal range. It is concluded that effect of ACE inhibitor on improving functional capacity in CHF is independent of it's any effect on exercise-induced ventricular arrhythmias. PMID- 11273110 TI - Acquired drug resistance in tuberculosis in Harayana, India. AB - Sputa of 200 treatment failure cases of pulmonary tuberculosis over a period of 1991-1995 were subjected to culture and sensitivity testing against commonly used anti-tuberculosis drugs. Out of 200 cases, 75% revealed resistance to one or more anti-TB drugs, resistance to isoniazid was observed in 72% cases, it was 49% for rifampicin, and 37% for streptomycin, while the resistance rate for other drugs was low. Majority of patients had resistance to two or three drugs concomitantly while resistance to 4, 5 and 6 drugs was of very low order and resistance to reserved drugs like kanamycin, ethionamide and cycloserine was encountered infrequently (1%). Out of 200 treatment failure patients multidrug resistance (MDR) was seen in 59% cases as 16% revealed resistance to isoniazid alone and strains in 22% cases were sensitive to all drugs. The study concludes that acquired MDR against first line antituberculosis drugs had increased as significant resistance against 3 drug combinations was observed although resistance against 2 drugs concomitantly was insignificant. Most ominous acquired drug resistance was seen against rifampicin in our region. The trends of drug resistance in the country and Haryana State are compared and their implications on outcome of chemotherapy are discussed. PMID- 11273112 TI - Pictorial CME. PMID- 11273111 TI - Risk stratification by treadmill testing in acute myocardial infarction following thrombolytic therapy. AB - Survivors of acute myocardial infarction (AMI) should have risk stratification for assessment of their future risk of cardiovascular events. One of the important means of risk stratification is by treadmill test (TMT). Most of the algorithms for assessment were done in the prethrombolytic era. But in the post thrombolytic era, risk stratification by TMT should be properly evaluated. Fifty males with confirmed AMI with age ranging from 38-62 years (mean 48 years) were tested with a symptom limited (Modified Bruce Protocol) TMT. The patients were followed up for a minimum of 6 months (range 6-10 months). Out of 50 patients, 38 reported for follow up. Among them 22 (Group A) had cardiac events and 16 (Group B) had no events. Among the patients (Group A), 6 had unstable angina, 7 had reinfarction, 2 had sudden death, 4 had coronary artery bypass grafting (CABG) and 3 had angioplasty. Comparison between the two groups, A and B in TMT parameters like ST segment depression > 2.5 mm (12 vs 9), no. of leads where ST depression occurred (66 vs 48) during exercise, mean work capacity (8.1 vs 7.9 mets), mean systolic blood pressure response were all statistically insignificant. Though TMT was believed to be a good prognostic indicator to assess further cardiac events after AMI, its efficacy in risk stratification after thrombolysis is yet to be determined. This study does not show its worth in post MI risk assessment. PMID- 11273113 TI - Nonsystemic vasculitic neuropathy. AB - The clinical, electrophysiological and pathological features and prognosis of 9 patients with nonsystemic vasculitic neuropathy are described. Nonsystemic vasculitic neuropathy accounted for 3% of cases of biopsy proven cases of various neuropathies and formed 56% of vasculitic neuropathy. Both clinically and on electrophysiological testing, mononeuritis multiplex was the form of neuropathy in 5 patients and 3 had sensory neuropathy. All the patients had a necrotizing vasculitis on nerve biopsy. Axonal degeneration was seen in teased fibers in all the patients. Eight patients showed good functional recovery one was left with mild bilateral claw hands. PMID- 11273114 TI - Yoga therapy in chronic bronchitis. AB - Fifteen patients of chronic bronchitis received yoga therapy in the form of pranayam and 8 types of 'asans' for a period of 4 weeks. They had a perceptible improvement in dyspnoea as was measured by visual analog. Lung function parameters (VC, FEV1, and PEFR) also improved after the practice of yoga. This preliminary study indicates that, yoga may be an useful adjunct to other conventional form of therapy for COPD. PMID- 11273116 TI - Vertigo: approach to the diagnosis and management. PMID- 11273115 TI - Addition of iodine to water by a filter based on polyiodide resin technology used at household level. PMID- 11273117 TI - Antibiotic policy. PMID- 11273118 TI - Current developments in the management of subacute sclerosing panencephalitis. PMID- 11273119 TI - Non-ulcer dyspepsia. PMID- 11273120 TI - Intrapleural streptokinase in multiloculated empyema thoracis. PMID- 11273121 TI - Rare presentations of idiopathic myelofibrosis: spontaneous rupture of spleen; pyoderma gangrenosum; and urologic obstruction. PMID- 11273122 TI - Carcinoid tumour of the caecum. PMID- 11273123 TI - High protein diet induced periodic behavioural disturbances in a case of compensated liver disease. PMID- 11273124 TI - Oncocytic variant of adrenal carcinoma presenting as Cushing's syndrome. PMID- 11273125 TI - Unsuspected filariasis coexisting with leishmaniasis in a splenic aspirate. PMID- 11273126 TI - Carcinoma colon presenting as paraneoplastic sensorimotor neuropathy. PMID- 11273127 TI - Emphysematous pyelonephritis successfully managed by medical therapy. PMID- 11273128 TI - Extent and pattern of self medication among inpatients in a north-Indian referral hospital. PMID- 11273129 TI - Worsening of steroid depending bronchial asthma following rifampicin administration. PMID- 11273130 TI - Clinical profile of sarcoidosis in Himachal Pradesh. PMID- 11273131 TI - MRI in excluding idiopathic intracranial hypertension. PMID- 11273132 TI - PAM responsive Gullian Barre like syndrome with low serum cholinesterase. PMID- 11273133 TI - Actin assembly at membranes controlled by ARF6. AB - The small GTPase, ADP-ribosylation factor-6 (ARF6), has been implicated in regulating membrane traffic and remodeling cortical F-actin. Using real-time video analysis of actin assembly in living cells, we investigated the function and mechanism of ARF6 in control of actin assembly. Expression of an activated form of ARF6 that mimicks the GTP-bound form of the GTPase induced actin assembly resulting in the movement of vesicle-like particles, some of which contain markers for pinosomes. Activated ARF6 also stimulated actin assembly at foci on the ventral surface of the cell and stimulated fluid phase pinocytosis. Particle motility induced by ARF6 involved Arp2/3 complex, tyrosine kinase activity, phospholipase D (PLD) and D3-phosphoinositides, but not phosphatidylinositol 4,5 bisphosphate (PI(4,5)P2). We conclude that ARF6 regulates actin assembly for pinosome motility and at foci on the ventral cell surface. PMID- 11273134 TI - Acute renal failure with neurological involvement in adults with measles: a new syndrome. PMID- 11273135 TI - Prognostic significance of eye changes in cerebral malaria. AB - AIMS: Eye in Plasmodium falciparum malaria are described by various workers all over the world but its prognostic significance is not clear because of conflicting observation by different authors from different regions. No such study is available on Indian adult patients of cerebral malaria. So we want to describe our observations on various eye abnormalities in these patients and study its prognostic significance. METHODOLOGY: Two hundred and fourteen adult (> 14 years) patients of strictly defined cerebral malaria admitted in classified malaria ward in this tertiary level health care station were studies. Detailed ophthalmoscopic examination was done through dilated pupils at the time of admission, daily thereafter, at the time of discharge and at weekly intervals in those with persistent changes at the time of discharge. RESULT: Retinal haemorrhage was found in 25 (11.68%) patients, papilloedema in 17 (7.94%), blurring of disc margins in 25 (11.68%), retinal oedema in six (2.8%), disc pallor in five (2.33%), vitreous haemorrhage and hard exudate in one (0.46%) each and subconjunctival haemorrhage in six (2.8%) patients. The mortality associated with individual finding was not statistically significant except disc pallor. CONCLUSION: None of the above finding except disc pallor (p < 0.05) was associated with statistically significant mortality (p > 0.05); whereas any of the fundus findings as a whole was related to statistically significant mortality (p < 0.05). PMID- 11273136 TI - Role of oxygen free radicals in causing endothelial damage in acute myocardial infarction. AB - OBJECTIVES: This work was done in order to study the oxidant and anti-oxidant status in a disease resulting from endothelial injury. The disease selected for study was acute myocardial infarction. METHODS: Sixty patients of acute myocardial infarction were selected after being diagnosed in accordance to the guidelines laid down by the WHO. Thirty subjects were included as controls. Plasma levels of certain markers of oxidative stress and anti oxidant activity were measured in all the subjects. Malonaldehyde (MDA) and nitrite (NO2) were measured as markers of free radical mediated endothelial injury, and superoxide dismutase (SOD) enzyme as an indicator of antioxidant activity. RESULTS: It was found that the plasma levels of MDA and nitrite were significantly elevated in the patients of acute myocardial infarction compared to the control group (7.29 +/- 3.28 v/s 4.57 +/- 0.63 nmol/ml and 12.85 +/- 8.71 v/s 0.97 +/- 0.25 microM respectively), thereby indicating that oxygen free radicals cause endothelial damage in them. The superoxide dismutase levels were also found to be elevated in these patients (5.57 +/- 1.47 v/s 3.91 +/- 0.66 U/ml). CONCLUSION: These results indicate that acute myocardial infarction is a state of enhanced free radical activity, which causes endothelial damage. The elevated SOD levels may imply that the body attempts to combat this oxidative stress by raising it's level of anti oxidants. PMID- 11273137 TI - Comparison of four nonculture diagnostic tests for Chlamydia trachomatis infection. AB - OBJECTIVE: Chlamydia trachomatis (CT) is one of the commonest sexually transmitted diseases leading to urethritis, epididymitis, prostatitis in men and urethritis, cervicitis, endometritis and pelvic inflammatory disease, sometimes complicated by infertility and ectopic gestation in women. Since culture of fastidious bacteria in a monocellular medium is not available in most laboratories we compared direct immunofluorescence antigen detecting test (DFA) with three other nonculture tests-antigen detecting enzyme immunoassay (EIA), Papanicolaou staining (Pap) and Geimsa stain for endocervical swabs from women in reproductive age group. METHODS: Three hundred and fifty seven women between 16 and 41 years of age and attending family welfare clinics of IRR were evaluated for the presence pap smears. In 100 cases DFA staining was compared with Geimsa staining. RESULTS: DFA test was positive in 60/357 (16.8%), EIA in 29 (8.1%) of cases and Pap smear in 37 (10%) cases. In the second group DFA was positive in 17 (17%) and Geimsa in 10 (10%) cases. CONCLUSION: Amongst the four tests DFA showed maximum sensitivity. ELISA is less expensive but has lower sensitivity. Pap stain also has less sensitivity and good specificity, the quality of smear is likely to affect the diagnosis. Though Geimsa stain is cheapest, for chlamydial cervicitis in our experience it was not as sensitive as DFA. Thus each laboratory must decide the method depending on its resources. PMID- 11273139 TI - Effect of esophageal and gastric distention on bronchial hyper-responsiveness in patients with bronchial asthma. AB - OBJECTIVE: Over-eating is said to aggravate asthma though the mechanism is still unclear. We tried to study the mechanism by causing distention in oesophagus and stomach. METHODS: Fifteen patients with nocturnal asthma were studied in a random cross-over design. The esophagus and stomach of the subjects were distended with a balloon. The effect of the distention on the airways was measured by taking forced expiratory volume one second (FEV1), forced vital capacity (FVC) and bronchial hyper-responsiveness (BHR). RESULTS: Distention of stomach caused significant reduction in FEV1 on FEV1/FVC ratio but similar distention of esophagus did not. Histamine PD20 was decreased by 0.43 (SEM 0.28) doubling dose with gastric distention. However, with oesophageal distention no significant change was observed in PD20. CONCLUSION: It can be concluded that gastric distention leads to broncho-constriction as measured by FEV1, FEV1/FVC ratio along with increase in BHR probably by inducing airway inflammation. Therefore asthmatic patients should be advised to avoid large meals. PMID- 11273138 TI - Magnesium deficiency potentiates free radical production associated with myocardial infarction. AB - OBJECTIVE: Oxidative injury and magnesium deficiency may accompany cardiovascular disease states and the study was planned to find out whether magnesium deficiency promotes oxidative injury. METHODS: Serum malonaldehyde (MDA), magnesium, vitamin E and total glutathione levels (GSH) were estimated in 22 patients with acute myocardial infarction and 15 healthy controls. RESULTS: Low levels of Mg, GSH, vitamin E and elevated levels of MDA were observed in patients of acute myocardial infarction. Statistically significant correlations were observed between Mg and MDA, MDA and GSH, Mg and vitamin E. CONCLUSION: Our findings suggest that Mg deficiency can potentiate oxidative injury to post ischaemic myocardium and that antioxidants may have a role in protection against the prooxidant influence(s) of Mg deficiency. PMID- 11273140 TI - Dietary and serum iron, body iron stores and coronary heart disease. AB - OBJECTIVES: To determine the role of body-iron stores as measured by serum iron, total iron binding capacity (TIBC), transferrin, ferritin and ferritin:transferrin ratio (FTR) in patients with coronary heart disease (CHD). METHODS: A case-control study was performed in 58 newly diagnosed CHD patients and 24 controls who were evaluated using clinical history, dietary history and biochemical examination. Dietary iron was determined by history; serum iron and TIBC were measured biochemically and ferritin by enzyme-linked immunoassay. Case control comparisons were performed by non-parametric Mann-Whitney test. RESULTS: There was no significant difference in mean age, prevalence of diabetes, hypertension and smoking, and dietary intake of calories and fats in cases and controls. Dietary iron intake was 11.2 +/- 3.4 mg/day in cases and 11.3 +/- 3.8 mg/day in controls (p > 0.05). Serum fasting glucose, cholesterol, LDL cholesterol, HDL cholesterol and triglycerides were not significantly different in cases and controls (p > 0.05). LDL/HDL ratio (4.17 +/- 1.4 vs. 4.62 +/- 2.3) and total cholesterol/HDL ratio (6.47 +/- 1.6 vs. 6.91 +/- 2.4) were also similar. In the whole study group serum iron (54.8 +/- 35.7 mcg/dl), transferrin (11.6 +/- 7.4%) and ferritin (52.4 +/- 57.8 ng/ml) levels were low. In cases as compared to controls serum iron (56.9 +/- 31 vs. 49.6 +/- 45 mcg/dl; z = 1.707, p = 0.088) and transferrin saturation (12.5 +/- 7.8 vs. 9.5 +/- 6.2%; z = 1.83, p = 0.066) were slightly more. Ferritin levels (48.8 +/- 55 vs. 60.9 +/- 64 ng/ml; z = 2.048, p = 0.040) as well as FTR (5.51 +/- 8.6 vs 7.47 +/- 6.1, z = 2.054, p = 0.040) was significantly lower in cases. CONCLUSIONS: In Indian CHD patients the body iron stores are lower as compared to controls. PMID- 11273141 TI - Detection of anti-nuclear antibodies from filter paper blood clots using indirect immunoenzyme technique: preliminary experience and results. AB - BACKGROUND: The facilities to detect antinuclear antibodies (ANA) patterns in patients with systemic rheumatic diseases/connective tissue disorders (CTD) using indirect immunofluorescence (IIF) technique (the gold standard) are sparse; the technique is technically difficult and expensive. A simpler technique, such as the indirect immunoenzyme (IIE) which uses light microscopy, ought to be evaluated for widespread use in our setting. OBJECTIVE: To study the feasibility and relevance of IIE in demonstrating ANA patterns, both from serum and filter paper blood clots (FPBC), in patients with CTD. METHODS: In this pilot study, ANA were detected from sera and FPBC of 21 patients with proven CTD using IIE; paired FPBC and serum samples were simultaneously collected in 10 patients. All samples, coded randomly, were tested by IIE and IIF, along with positive and negative controls. RESULTS: Using IIE, the results of the ANA patterns obtained from FPBC eluates and sera were similar; homogenous (SLE-6, PSS-1, RA-4), speckled (SLE-8, PSS-2, Overlap CTD-1) and centromere (PSS-1). Four SLE patients showed mixed pattern; sensitivity of IIE for lupus was hundred percent. On comparing the results with the serum IIF, the Kappa statistic of agreement was 1 (perfect) and 0.4 (fair) for FPBC-IIF and FPBC-IIE respectively; the results matched between serum IIF and FPBC-IIE in 8 of the 10 paired samples tested. CONCLUSIONS: IIE can demonstrate ANA both from sera and FPBC. This pilot study besides demonstrating positive trends for further probe also creates an awareness for such a feasible technique. However a larger sample size would be required to carry out its evaluation as an alternative to IIF and as a screening technique. PMID- 11273143 TI - Does nasal breathing cause frictional trauma in allergic rhinitis? AB - OBJECTIVES: Frictional stress on the walls of a tube increases with increased air flow and as the diameter of the tube is reduced. High values of frictional stress may occur in the nose during nasal obstruction which could damage the nasal mucosa particularly when the mucosa is inflamed and fragile as in allergic rhinitis. The effect of nasal airflow induced frictional stress on the nasal mucosa was studied in patients with allergic rhinitis. METHODS: We studied nasal peak flow rate in eight patients with allergic rhinitis and nasal obstruction comparing the change in peak expiratory flow after they breathed for 30 minutes through an obstructed and a patent nostril. Patients were studied in the right and left lateral decubitus positions to increase and decrease the resistance in the lower and upper nostril respectively and thus minimize any effects of cyclical changes in nasal resistance. Subjects breathed for 30 minutes through the upper patent nostril (schedule 1) and for a further 30 minutes through the lower obstructed nostril (schedule 2). Nasal peak expiratory flow rate was measured in both nostrils separately in both positions after each schedule. RESULTS: There was a significant reduction in mean (SD) nasal peak flow rate ( 12.8 (4.06) L/min) after subjects had breathed for 30 minutes through the obstructed nostril. There was no significant change in nasal peak flow rate after subjects had breathed through the patent nostril, or in the nostril that had no flow for 30 minutes. CONCLUSIONS: These findings are compatible with the hypothesis that frictional stress due to airflow through an obstructed nostril induces trauma and swelling of the nasal mucosa of patients with allergic rhinitis. PMID- 11273142 TI - Evaluation of efficacy and safety of losartan potassium in the treatment of mild to moderate hypertension as compared to enalapril maleate. AB - AIM: To study the effect of losartan potassium in the treatment of mild to moderate hypertension and to compare its efficacy and adverse effect profile with enalaparil maleate. MATERIAL AND METHODS: One hundred and forty five patients with mild to moderate essential hypertension were enrolled in this randomized, double blind, controlled, parallel and multicentric study. Seventy two patients received losartan potassium 50 mg and seventy three received enalapril maleate 5 mg. RESULTS: Losartan potassium reduced the DBP to < 90 mm Hg in 59% of the patients at the end of 8 weeks compared to 45% in the enalapril maleate group. DBP was reduced by 10 or > than 10 mm Hg in 89% of the patients with losartan as compared to the baseline whereas it was 80% in the enalapril group. Percentage of side effects seen in losartan and enalapril groups were 12 and 22 respectively. CONCLUSION: Losartan potassium is an efficacious antihypertensive agent in mild to moderate hypertension. It also has fewer side effects when compared to enalapril maleate. PMID- 11273144 TI - Clinical profile of non-O1 strain-O139 of Vibrio cholerae in the region of Ambajogai, Maharashtra. AB - OBJECTIVES: To study clinical profile of the newly emerged novel strain non-O1, O139 of Vibrio cholerae, in the region of Ambajogai, District Beed of Maharashtra. METHODS: Out of 208 patients of acute gastroenteritis, 41 revealed to be positive for Vibrio cholerae by recommended method of stool examination. All the strains were sent to National Institute of Cholera and Infectious Diseases, Calcutta for confirmation. RESULTS: Out of 41 cases, 12 were of Vibrio cholerae O1, 29 Non-O1, of which nine found to be O139 strain. All patients were from 2-80 years of age with low-socioeconomic status and maximum incidence was in August (64.70%), presented with severe rice watery loose motions. Vomiting was observed in 26 (63.41%), more so in patients of O139 infection (88.88%) than four (33.33%) of O1 infection. Sweating was observed in three patients (33.33%) of O139 infection, cramps in gastrocnemis muscles in three patients (33.33%) of O139 infection and two (16.66%) of O1 infection. Signs of dehydration were mild to moderate in four patients (33.33%) of O1 infection; severe dehydration in six (66.66%), moderate in two (22.22%) and mild in one patient (11.11%) of O139 infection. While dehydration was severe in four (20%), moderate in one (5%) and mild in three patients (15%) of Non-O1 infection (excluding O139 cases). Clinical features were more severe in patients of serotype O139 than the patients of O1 and Non-O1 (excluding O139 cases). However all patients responded to intravenous fluids, oral rehydration and antibiotics (tetracycline) within 24-48 hours without any mortality. CONCLUSIONS: This study reflects the first emergence of Non-O1, strain O139 during the year 1997 with severe and critical clinical features in Ambajogai region causing high morbidity in the form of severe dehydration and peripheral circulatory collapse which requires early and correct diagnosis and prompt treatment. PMID- 11273145 TI - Pictorial CME. Myocardial bridging. PMID- 11273146 TI - Adult subacute mountain sickness--a syndrome at extremes of high altitude. AB - BACKGROUND: Subacute mountain sickness is distinct syndrome of congestive cardiac failure seen in lowlanders during prolonged stay at extreme high altitude (> 5800 mtrs). OBJECTIVES: To study the clinical and investigative profile of subacute mountain sickness amongst Indian soldiers stationed at extreme altitude. MATERIAL AND METHODS: Symptomatic individuals who were stationed above 5000 mtrs were evacuated to 3000 mtrs and clinically screened for signs of congestive cardiac failure. ECG and X-ray chest; hematological and biochemical parameters were evaluated. Response to rest, oxygen and diuretics were studied and they were evacuated to the plains. They were followed up at the plains for a period of two weeks at the end of which chest X-ray and ECG were repeated. RESULTS: Eight patients were diagnosed over a period of one month who had classical features of congestive cardiac failure. The mean age was 28.75 years, the mean altitude 5828.47 mtrs and the mean duration of stay 17.35 weeks. The most common symptom was exertional dyspnea (6 of 8 cases) and the most common sign bilateral pedal edema (7 of 8 cases). Two patients had deep venous thrombosis. Clinical, ECG and X-ray evidence of pulmonary hypertension was seen in seven cases. The mean hemoglobin was 18 gm%. Response to oxygen and diuretics was dramatic. Clinical findings and investigations reverted to normal after two weeks of stay on the plains. CONCLUSION: This brief study of subacute mountain sickness reemphasizes the role of pulmonary hypertension as the initiating event. Other factors are salt and water retention and polycythemia. Brisk response to diuretics and oxygen and restoration of normalcy on deinduction to the plains establishes the reversibility of the syndrome. PMID- 11273147 TI - Role of losartan therapy in the management of diabetic hypertension. AB - The management of diabetic hypertension requires meticulous selection of agents in the antihypertension armamentorium. There may be several associated factors to be considered while treating a hypertensive diabetic. These include hyperglycemia, dyslipidemia, proteinuria, left ventricular hypertrophy and heart failure to name a few. Losartan is the first of a new class of agents in the list of antihypertensive drugs. By its selective angiotension II receptor (subtype AT1) blocking action it is postulated to bring about a more complete inhibition of the renin-angiotensin system. Thus, it might produce all the benefits of angiotensin converting enzyme (ACE) inhibitor therapy with the freedom from cough so commonly seen with the use of ACE inhibitors. This review attempts to analyze the possible benefits of losartan therapy in diabetes. PMID- 11273148 TI - First unprovoked seizure--to treat or not to treat? AB - It has always been a controversial subject whether or not to treat a patient with first episode of seizure. The actual decision whether or not to treat patients who present with initial seizure must be individualized. It depends on probability of having a recurrence and on the perceived risk/benefit ratio of treatment. In a large majority of patients, it is prudent to defer treatment with antiepileptic drugs until a second episode has occurred, unless it is a remote symptomatic seizure, associated with definite epileptiform abnormalities in EEG or a partial seizure. However, in an occasional patient, treatment may be indicated even after a first seizure; for example, in patients involved in certain occupations like driving or operating dangerous machinery. PMID- 11273149 TI - Medical philosophy. PMID- 11273150 TI - Renal failure, encephalitis and measles in a young woman. PMID- 11273151 TI - Tuberculous abdominal lymphadenopathy causing reversible renovascular hypertension. AB - Renovascular hypertension is an important but not so common cause of hypertension. Rarely the extrinsic compression of renal arteries by retroperitoneal structures may be responsible for hypertension. Lymphadenopathy due to metastasis has been reported to cause renovascular hypertension. The present case reports hypertension which resulted from tubercular abdominal lymphadenopathy in a twenty one years male. The patient was treated with antitubercular medication with which his hypertension also got controlled. An unusual cause of hypertension and a rare complication of tuberculosis is reported. PMID- 11273152 TI - Neurocysticercosis presenting as midbrain syndrome. AB - Brainstem is infrequently involved in patients with neurocysticercosis, usually, it occurs in association with disseminated form of neurocysticercosis. We are reporting two cases who had large multiple cysticercus lesions and presented as acute midbrain syndrome. The diagnosis of neurocysticercosis was established by presence of characteristic granulomatous extraaxial lesions around the midbrain, and in cerebral parenchyma, along with strongly positive ELISA for cysticercal antigen in cerebrospinal fluid as well as in serum. Both patients responded well to corticosteroids. However, repeat follow-up CT scan income case did not show significant alteration in the size of the lesion. PMID- 11273153 TI - Hypocomplementemic urticarial vasculitis and lower cranial nerve palsies. AB - A 55 years post menopausal lady presented with puffiness of face, and a pruritic urticarial rash over face and upper trunk of one week duration with accompanying dysphagia. Clinical examination revealed an urticarial rash over face and upper trunk, two small ulcers over floor of mouth and evidence of bilateral VIII, IX and Xth cranial nerve palsies. Hypocomplementemia, negative immune profile and evidence of vasculitis on skin biopsy suggested a diagnosis of hypocomplementemic urticarial vasculitis. The patient responded to a course of steroids. PMID- 11273154 TI - Guillain-Barre syndrome following antirabies vaccine. PMID- 11273155 TI - Encephalopathy, intermediate syndrome and delayed polyneuropathy in acute black Danadar (Phorate 10 CG) poisoning. PMID- 11273156 TI - Splenic abscess. PMID- 11273157 TI - An unusual presentation of Sprengel's deformity. PMID- 11273158 TI - Cerebral venous thrombosis due to protein C deficiency. PMID- 11273159 TI - Acquired hyperhomocysteinemia, megaloblastosis with subacute combined degeneration and deep venous thrombosis. PMID- 11273160 TI - Adverse drug reactions postal survey-bronchial asthma and angioedema with nimesulide. PMID- 11273162 TI - Hypochronic anemia in chronic renal failure--role of aluminium. PMID- 11273161 TI - Hypoglycaemic complication of the quinine. PMID- 11273163 TI - Conventional medical journalism is to stay. PMID- 11273164 TI - Cost effectiveness in renal function tests. PMID- 11273165 TI - Stroke like presentation of Creutzfeldt Jakob disease: an unusual variant. PMID- 11273166 TI - Impaired fasting glucose (IFG) versus impaired glucose tolerance (IGT): the present scenario. PMID- 11273167 TI - Give a boost to infectious diseases. PMID- 11273168 TI - Clinical profile of neurobrucellosis--a report on 12 cases from Bikaner (north west India). AB - OBJECTIVE: To study the spectrum of neurobrucellosis in a prospective study at Bikaner which is supposed to be uncommon in India. METHOD: This study was done on admitted patients of brucellosis from June 1996 to June 1999 in whom the diagnosis was done by history of exposure to animals, fever and arthralgia and demonstration of brucella antibody titres in serum 1:160. CSF examination was done in all the patients. All cases were treated by combination of doxycycline 100 mg twice daily, rifampicin 900 mg daily for 6-8 weeks and injection streptomycin 0.75 to 1 gm i.m. per day for initial 14 days. Detailed neurological examination and antibody titres of serum and CSF again measured at the end of treatment. RESULTS: Twelve out of 92 patients revealed evidence of neurobrucellosis in which four cases were of meningoencephalitis, two cases of myelitis leading to spastic paraparesis, five cases of polyradiculoneuropathy and one case of polyneuroradiculomyeloencephalopathy. The treatment regimen used was associated with a high cure rate and significant reduction in antibody titres in serum and CSF. CONCLUSION: Neurobrucellosis is an uncommon but serious manifestation affecting central and peripheral nervous system. The clinical profile of the disease mimicks closely to commonly seen neurological infective diseases like tubercular meningitis, viral encephalitis, aseptic meningitis, cerebral malaria and viral encephalopathy. Serum and CSF testing for brucella antibody titre is an important test for the diagnosis. Blood culture is not an ideal test for neurobrucellosis because of low yield and longer time required for the diagnosis. High degree of suspicion is prudent for the diagnosis. High degree of cure rate can be achieved by treatment with present regimen in a disease which is otherwise having high mortality and morbidity. PMID- 11273169 TI - Interictal brain 99m Tc-HMPAO SPECT study in cases of epilepsy with single ring enhancing CT lesion. AB - BACKGROUND: Contrast enhancing single ring or disc lesion (CESRL), a frequent finding in the CT scan of Indian patients with seizures, has a diverse etiology. Underlying cause in many of these cases remains conjectural. Some of these ring lesions show spontaneous resolution without any specific treatment and in others their temporal profile remains unpredictable. MATERIAL AND METHODS: In a prospective study we studied 17 cases of epilepsy, with the CT scan finding of single contrast enhancing ring lesion. Detailed clinical evaluation and interictal EEG was performed. CT scan and SPECT study was done initially and were then repeated. RESULTS: On the second CT, one lesion disappeared and 2 reduced in size, three showed no change in size. Third CT showed complete disappearance in one case and no change in the second case. Initial SPECT study was abnormal in all 17 cases, showing areas of hypoperfusion corresponding to the anatomical location of ring lesion. On follow up, patients with decreased ring size on CT showed decrease in area of perfusion deficit on the SPECT, but cases in which the lesion disappeared on the CT, the SPECT perfusion abnormality continued to persist, though to a lesser extent. CONCLUSION: Persistence of perfusion defects suggest the presence of altered underlying physiology. Hence, early withdrawal of antiepileptic drugs after disappearance of ring lesion on CT may lead to seizure recurrence. SPECT studies repeated after 3 months may help to prognosticate cases with CESRL and also help in deciding the optimum duration of antiepileptic therapy in individual cases. PMID- 11273170 TI - Behavioural disturbances in dementia. AB - OBJECTS: To assess the frequency of behavioural disturbances in patients suffering from dementia and their relation to dementia severity. METHODS: The study evaluated 75 patients referred to the Memory Clinic at our hospital. Patients meeting DSM-IV criteria for dementia were included in the study. Activities of daily living (ADL) and cognitive functioning were also assessed. Patients were then rated for dementia severity using the Clinical Dementia Rating scale. Clinical data regarding behavioural disturbances was obtained from the patient (where possible) and an informant (usually the primary caregiver) who had contact with the patient at least thrice a week. RESULTS: Results showed that behavioural disturbances were present in more than 60% of the sample. Psychotic and activity associated disturbances were most common, and that these were seen more frequently in Alzheimer's disease than any other type of dementia. Further, these disturbances occurred most frequently in dementias of moderate severity but reduced as the dementia progressed further. CONCLUSION: Our results indicate that behavioural disturbances are a prominent part of dementia and that judicious use of psychiatric medication should form an important aspect of management. PMID- 11273171 TI - Antiretroviral drugs in the treatment of people living with human immunodeficiency virus: experience in a south Indian tertiary referral centre. AB - OBJECTIVE: A decrease in the number of new acquired immunodeficiency syndrome (AIDS) cases and AIDS--related deaths was seen in developed countries since 1996 due to the use of new combination of antiretroviral drugs. This retrospective study discusses the use of antiretroviral drugs in the treatment of people living with human immunodeficiency virus (HIV) in a developing country setting. METHODS: A retrospective case note analysis was done of patients receiving antiretroviral therapy at YRG Centre for AIDS Research and Education between Aug. 1996 and Feb. 1999. Out of 936 persons with HIV treated at this centre, 6.1% of the patients were prescribed three groups of drugs: Group A was the combination of the reverse transcriptase inhibitors (nRTI) zidovudine 600 mg daily and lamivudine 300 mg daily, Group B was the combination of zidovudine 600 mg daily, lamivudine 300 mg daily with protease inhibitor (PI) ritonavir 1200 mg daily and Group C was the combination of zidovudine 600 mg daily and lamivudine 300 mg daily with indinavir 2400 mg daily. Twenty HIV positive pregnant women were given zidovudine 500 mg daily during the third trimester (Group D) to reduce the vertical transmission of HIV. RESULTS: The mean CD4 gain was 188.0 cells/micro litre in Group A, 118.8 cell/microlitre in Group B and 223.3 cells/microlitre in Group C with a mean duration of 4.3, 3.1 and 3.5 months respectively. Many patients stopped antiretroviral drugs due to high cost of therapy. CONCLUSION: Hence, physicians should prescribe antiretroviral drugs only after ensuring that the patients can afford and will comply with a longterm treatment. Prescribing guidelines should be available to those working in this field and should be adhered to so that emergence of resistant strains could be prevented. PMID- 11273172 TI - Foreign bodies in gut. AB - BACKGROUND: Foreign body ingestion is common and a frightening experience to the patients and relatives. We report our experience with 102 patients, (78 children and 24 adults), with foreign body ingestion. METHODS: After locating foreign bodies radiologically, 34 (43.6%) foreign bodies in children and 13 (54.2%) foreign bodies in adults were removed endoscopically. General anesthesia was used in 32 children and overtube was used for all sharp foreign bodies. RESULTS: In our study, 78 (76%) patients were below 12 years of age. Coins (79.5%) were commonest foreign bodies in children while dentures (25%) were commonest in adults. Foreign bodies were most commonly sited in stomach (25.6%) in children and esophagus (58.3%) in adults. In 41 (52.6%) children and in three (12.5%) adults, i.e. total 44 out of 102 (43.1%) patients passed foreign bodies spontaneously. The largest foreign body that passed spontaneously was 4-cm long nail in a child. In 34 (43.6%) children and in 13 (54.2%) adults foreign bodies were removed endoscopically. Only 3.8% children and 33.3% adults required surgery. CONCLUSIONS: There was no mortality in our series. Majority of foreign bodies do not require any intervention. Sharp foreign bodies are commonest indication for surgery. However, endoscopic removal is safe, effective and is the method of choice for most patients. PMID- 11273173 TI - Influence of smoking and hypertension on left ventricular mass. AB - OBJECTIVE: The present study was designed to observe the influence of smoking and hypertension on left ventricular mass (LVM), both individual and the combined effect. METHODS: This study was conducted in the Department of Medicine, SMS Medical College Hospital, Jaipur. Hundred patients were included in the study. They were divided into four groups--non smoker normotensives, non-smoker hypertensives, smoker normotensives and smoker hypertensives. They were also divided into smokers and nonsmokers and hypertensives and normotensive. Patients of age group 31-65 years of both sexes were chosen. Patients with secondary hypertension, diabetes mellitus, dyslipidemia, cardiomyopathies, valvular heart disease women on oral contraceptives were excluded from the study on the basis of history and relevant laboratory tests. An M-mode echocardiography was done and the LV mass was calculated by the formula proposed by Devereux et al. LV mass = 0.8 [1.04 (IVSd + LVIDd + LVPWd)3 - (LVIDd)3] + 0.6 gms RESULTS: The mean LV mass (in gms) in the four groups of nonsmoker normotensives, nonsmoker hypertensives, smoker normotensives and smoker hypertensives are 106.77 +/- 25.78, 165.3 +/- 42.55, 154.53 +/- 24.6 and 228.78 +/- 56.88 respectively. The comparison of mean LV mass (in gms) of smokers and nonsmokers were 191.66 +/- 40.74 and 136.04 +/- 36.16 (P < 0.001) respectively. The comparison of the mean LV mass (in gms) of hypertensives and nonhypertensives were 197.25 +/- 49.72 and 126.29 +/- 25.19 (P < 0.001) respectively. Comparison of the mean LV mass (in gms) of patient with two risk factors i.e., smokers hypertensives with patients without any risk factor i.e., nonsmoker normotensive were 228.78 +/- 56.88 and 106.77 +/- 25.78 (P < 0.001) respectively. While comparison of the mean LV mass (in gms) of patients with one risk factor each i.e., smoker normotensives and nonsmoker hypertensives were 165.3 +/- 42.55 and 154.53 +/- 24.6 (P < 0.05) respectively. CONCLUSION: Both smoking and hypertension cause an increase in LV mass, hypertension causing a more increase, than smoking, individually. Both smoking and hypertension combine together to increase the LV mass, more than either of them individually. Smoking by itself can cause an increase in LV mass independent of hypertension. PMID- 11273174 TI - Haemodynamic effects of pan masala in healthy volunteers. AB - OBJECTIVES: We studied acute haemodynamic effects of pan masala (powdered mixture of areca nut, slaked lime, catechu, and condiments) in healthy volunteers. METHODS: Fifty one males (mean age 28.6 +/- 10 years) were evaluated. One pouch (4 g) of pan masala without tobacco was given to each subject under fasting state and effects on pulse and blood pressure (BP) recorded. RESULTS: At baseline the pulse rate was 75.1 +/- 9.0 per minute, systolic BP was 119.1 +/- 10.8 mm Hg, and diastolic BP was 78.0 +/- 7.5. The pulse rate increased to 87.5 +/- 11.4 at ten minutes (+16.9 +/- 12.6%, p < 0.001) and fell to 76.7 +/- 9.1 at 30 minutes (p = ns). Systolic BP increased to 122.3 +/- 11.7 mm Hg at 10 minutes (+2.73 +/- 5.1%, p < 0.001) and was 120.8 +/- 10.8 at 30 minutes; while diastolic BP was 80.8 +/- 7.3 at 10 minutes (+3.83 +/- 6.1%, p < 0.001) and 79.4 +/- 7.6 at 30 minutes. CONCLUSIONS: Pan masala intake causes acute increase in pulse and BP. PMID- 11273175 TI - Mepacrine therapy in niclosamide resistant taeniasis. AB - OBJECTIVE: To evaluate the efficacy of mepacrine (quinacrine) in patients with niclosamide resistant Taenia saginata infection. METHODS: Eighty six cases with niclosamide resistant Taenia saginata (unresponsive to 2-8 courses of niclosamide) were treated with quinacrine (1 g) administered orally or via a nasogastric tube, and followed at 2, 4, 8 and 12 weeks for recurrence of passage of proglottids and presence of Taenia eggs in the stool examinations. Pre and post-therapy egg counts were obtained and egg viability was tested by staining with methylene blue. RESULTS: Eighty-one (94.2%) patients responded promptly with passage of the worm within 4-72 hours. The egg counts showed a drastic fall in 79 cases and a fall in viability from a median of 100% to 0% was observed. Only one patient demonstrated a relapse at 4 weeks. Gastrointestinal side effects occurred in 9 cases but were controlled easily by symptomatic therapy. CONCLUSION: We conclude that quinacrine is a safe, inexpensive, effective and generally well tolerated drug for the treatment of niclosamide resistant Taenia saginata infestations. PMID- 11273176 TI - Cyclospora infection in acquired immunodeficiency syndrome. AB - BACKGROUND: Diarrhea is a common clinical manifestation of human immunodeficiency virus (HIV) infection. The important protozoan pathogens causing diarrhea include cryptosporidium, microsporidium, Isospora belli and cyclospora besides giardia lamblia and entamoeba histolytica. OBJECTIVE: The objective of the present study was to determine the incidence of cyclospora (a coccidian protozoan) infection in HIV infected patients. METHODOLOGY: Faecal smears were stained by modified acid fast staining method to demonstrate oocysts of cyclospora. RESULTS: Out of 334 faecal specimens which were studied, cyclospora were identified in 22 cases (6.6 percent); and in 50 percent of the patients, there was a mixed infection with another protozoan parasite namely cryptosporidium. CONCLUSION: Identification of this parasite is important because cyclosporiasis can be treated with trimethoprim-sulfamethoxazole. Outbreaks of cyclospora infection have been linked to waterborne transmission. Though adequately chlorinated water is free of coliforms, it can still contain cyclospora. PMID- 11273177 TI - Non invasive ventilation. AB - Non-invasive ventilation refers to the technique of providing ventilatory support to a patient without an endo/orotracheal airway. It is a promising and rapidly upcoming new technique and is being used as first line therapy in a wide variety of conditions causing respiratory failure. The major indications for its use include respiratory failure due to a variety of causes (chest wall abnormalities, neuromuscular disease, COPD), weaning and stabilization of cardio-respiratory status before and after surgery. Patients who are candidates for this modality usually have a hypercapnic respiratory failure but are able to protect the airway and cooperate with treatment. The biggest advantage of the technique is its simplicity and avoidance of complications of intubation like trauma, infection and delayed complications like tracheal stenosis. Patient comfort is significantly improved and important functions like speech, swallowing and cough are preserved. Several purpose built ventilators are available for use including pressure preset and volume present machines, each of which have their own advantages and disadvantages in clinical practice. A range of patient interfaces is available. The initiation of non-invasive ventilation is much easier as compared to invasive ventilation and can be done for most patients in an intermediary care unit thereby cutting down treatment costs and saving precious intensive care beds. Titration of ventilatory parameters can usually be done using simple tests like oxymetry and blood gases. Several technique related problems like skin pressure sores, nasal symptoms and abdominal distension can be managed with simple measures. Non invasive ventilation has got a special and evolving role in management of COPD, both in acute exacerbations and chronic respiratory failure. In short, the advantages of this form of ventilation are numerous and physicians must familiarize themselves with this new technique, facilities for which should be available in all hospitals admitting patients with respiratory failure. PMID- 11273178 TI - Pictorial CME:Temporal arteritis. PMID- 11273179 TI - Mitochondrial cytopathies. PMID- 11273180 TI - Obesity--current perspective. PMID- 11273181 TI - Stress and Ayurveda: Selye--Mehta dialogue in context of the current findings. PMID- 11273182 TI - Subdural haematoma with spontaneous resolution--rare manifestation of idiopathic thrombocytopenic purpura. AB - Spontaneous intracranial haemorrhage presenting as subdural haematoma is an extremely rare presentation in adults due to idiopathic thrombocytopenic purpura. There are only five such case reports available in the literature and four had surgical evacuation of haematoma, while only one had spontaneous resolution. We report the case of a middle aged female who presented with bilateral papilloedema and later diagnosed to have subdural haematoma as a complication of idiopathic thrombocytopenic purpura, which had a spontaneous resolution. PMID- 11273183 TI - Disseminated histoplasmosis as an acquired immunodeficiency syndrome defining disease. PMID- 11273184 TI - Neuroleptic malignant syndrome occurring in a suspected case of systemic lupus erythematosus. AB - The case of a 40 year old female patient of SLE who developed fatal neuroleptic malignant syndrome following administration of serenase (haloperidol) is reported. Relevant literature has been reviewed. PMID- 11273185 TI - Familial infantile myaesthenia. AB - We report a family of two brothers with familial infantile myaesthenia which is an autosomal recessive congenital myaesthenic syndrome. It is a presynaptic neuro muscular junction disorder, responsive to treatment and has got good prognosis. PMID- 11273186 TI - Thrombocytosis--report of a case. AB - Thrombocytosis is a common clinical condition. Drugs used for thrombocytosis are hydroxy urea, alkylating agents and interferon alpha. We are reporting a case of chronic myeloid leukemia with thrombocytosis who did not respond to hydroxy urea but was controlled with the addition of a new drug anagrelide. Anagrelide is a platelet reducing agent with no serious side effects. It may be the main weapon against thrombocytosis associated with chronic myeloid leukaemia (CML) and essential thrombocytosis in the coming years. PMID- 11273187 TI - Emerging resistance of Plasmodium falciparum to artemisinine and related compounds. PMID- 11273188 TI - Diagnostic dilemma: aspergillosis. AB - Two cases of varied forms of Aspergillosis are reported who were being diagnosed and treated on different lines. One case, who was treated on lines of allergic bronchitis, had very high total eosinophil count and, fleeting pulmonary infiltrates over a period of 5 years along with history of cough, fever and weight loss. Aspergillus fumigatus was grown on sputum culture. On the background of a long standing history of bronchial asthma with evidence of peripheral as well as central eosinophilia, fleeting pulmonary infiltrates and A. fumigatus grown on sputum culture, we kept the diagnosis of Allergic Bronchopulmonary aspergillosis (ABPA) and put the patient on steroids and Itraconazole. Patient showed good response to therapy. Another case, a 50 year old male, presented to us with clinical picture of subacute myelitis. Being a known case of ABPA and on steroid therapy for long duration, we kept the diagnosis of invasive aspergillosis. Growth of Aspergillus fumigatus on sputum culture on three occasions and MR imaging of spine further supported our view. Aspergillosis of the lung do not have characteristic clinico-radiological features of permit the diagnosis and should be considered in the differential diagnosis of tuberculosis, pneumonia, bronchiectasis, lung abscess and bronchial asthma. PMID- 11273189 TI - Refractory hypokalemia in metastatic adrenocorticotrophic hormone--secreting pituitary carcinoma. PMID- 11273190 TI - Pontine hematoma and one and a half syndrome. PMID- 11273191 TI - Poland's syndrome--complicated with renal hypertension. PMID- 11273192 TI - Low trypsin levels in type-1 diabetes: an index of low exocrine output. PMID- 11273193 TI - The study of prolonged fevers. PMID- 11273194 TI - Ascaris psychosis: an unusual presentation of round worm infestation. PMID- 11273195 TI - Telepathology: advantages and problems. PMID- 11273196 TI - High altitude koilonychia developing as a result of cold environment and hypoxia- a fact or a myth? PMID- 11273197 TI - Penetrating cardiac injury: a dreaded outcome of diwali (fire work) festival. PMID- 11273199 TI - Epidemiological studies in developing countries. PMID- 11273198 TI - Randomized controlled trial on the effective dose of anti-snake venom in case of snake bite with systemic envenomation. PMID- 11273200 TI - Scorpion sting. PMID- 11273201 TI - Making room for a digital image view. PMID- 11273202 TI - No sure bets on micrometastases. PMID- 11273203 TI - Worries surface in wake of waives. PMID- 11273204 TI - Back to the drawing board. PMID- 11273205 TI - IOM lab recommendations right on the money. PMID- 11273206 TI - Labs slow to adopt some system features. PMID- 11273207 TI - Capitol scan. Key wins in self-referral rule. PMID- 11273208 TI - Relationship between HIV-1 Env subtypes A and D and disease progression in a rural Ugandan cohort. AB - OBJECTIVE: To investigate the role of HIV-1 envelope subtypes on disease progression in a rural cohort of Ugandan adults where two major HIV-1 subtypes (A and D) exist. METHODS: Participants of a clinical cohort seen between December 1995 and December 1998 had blood collected for HIV-1 subtyping. These included prevalent cases (people already infected with HIV at the start of the study in 1990) and incident cases (those who seroconverted between 1990 and December 1998). HIV-1 subtyping was carried out by heteroduplex mobility assay and DNA sequencing in the V3 env region. Disease progression was measured by the rate of CD4 lymphocyte count decline, clinical progression for the incident cases as time from seroconversion to AIDS or death, to first CD4 lymphocyte count < 200 x 10(6)/l and to the World Health Organization clinical stage 3. All analyses were adjusted for age and sex. RESULTS: One hundred and sixty-four individuals, including 47 prevalent and 117 incident cases, had V3 env subtype data of which 65 (40%) were subtyped as A and 99 as D. In the incident cases, 44 (38%) were subtyped as A and 73 as D. There was a suggestion that for most end-points A had a slower progression than D. The cumulative probability of remaining free from AIDS or death at 6 years post-seroconversion was 0.72 [95% confidence interval (CI), 0.50 to 0.85] for A and 0.58 (95% CI, 0.42 to 0.71) for D, and the adjusted hazard ratio of subtype D compared to A was estimated to be 1.39 (95% CI, 0.66 to 2.94; P = 0.39). The estimated difference in rates of decline in square root CD4 lymphocyte counts was -0.41 per year (95% CI, -0.98 to 0.15; P = 0.15). CONCLUSION: This study suggests that although subtype A may have a slower progression than D, HIV-1 envelope subtype is not a major factor in determining the progression of HIV-1 disease in a rural population in Uganda. PMID- 11273209 TI - Inhibition of Tat transactivation by the RNA polymerase II CTD-phosphatase FCP1. AB - OBJECTIVES: To asses the role of the RNAPII carboxy-terminal domain (CTD) phosphatase FCP1 on HIV-1 Tat-mediated transactivation. DESIGN: Construction of expression vectors encoding FCP1 phosphatase and analysis of their functions on Tat activity. METHODS: Basal and Tat-mediated transactivation of HIV-1 long terminal repeat (LTR)-driven transcription was compared, by transient transfections, in the presence of FCP1 phosphatase. Protein interactions were analysed by in vitro binding assays. RESULTS: FCP1 specifically and effectively represses Tat transactivation but not HIV-1 LTR-basal transcription. Protein interaction assays demonstrated that FCP1 specifically and directly binds Tat in vitro. CONCLUSION: The specific and efficient inhibitory function of FCP1 highlights the important role of this CTD-phosphatase in Tat-mediated transactivation, and it suggests that FCP1 might represent a specific target for modulation of Tat activity in infected cells. PMID- 11273210 TI - Clinical and laboratory guidelines for the use of HIV-1 drug resistance testing as part of treatment management: recommendations for the European setting. The EuroGUidelines Group for HIV resistance. AB - Viral drug susceptibility is associated with virologic response to new treatments. Standardized drug resistance tests are now available, and data from some clinical trials suggest that the use of drug resistance testing may be associated with improved virologic outcome. However, drug resistance testing is complex in terms of performance, interpretation and clinical application. HIV-1 drug resistance testing is used across Europe in patient management, but not in a consistent manner. This is due to differences in the national approaches to treatment, treatment management and reimbursement, as well as availability of tests. National guidelines only exist in some countries. In addition, the laboratory quality assurance and quality control standards are not applied uniformly. The EuroGuidelines Group was established to formulate clinical as well as laboratory guidelines for the use of HIV-1 drug resistance testing that are specific for the European setting. The group is comprised of academic clinicians and virologists, scientist from the industry and representatives of the patient community. The panel of experts will review these guidelines and update them on a yearly basis as new scientific evidence becomes available. PMID- 11273211 TI - Placebo-controlled trial of prednisone in advanced HIV-1 infection. AB - OBJECTIVE: To examine the safety and the immunologic and virologic consequences of corticosteroid use in HIV-1 infection. METHODS: A randomized, double-blinded, placebo-controlled trial of corticosteroid administration in 41 patients with advanced HIV-1 infection. Patients had a baseline median CD4 cell count of 131 x 10(6) cells/l at enrollment and 85% had a history of opportunistic infection. All but one of the patients had been taking stable antiretroviral regimen, including a protease inhibitor in 36, for a median duration of 158 days. Patients were randomized to 8 weeks of prednisone 0.5 mg/kg daily or placebo. RESULTS: No AIDS defining events occurred; two patients in each group developed oral candidiasis, and two patients on prednisone developed mild herpes simplex flares. None who developed oral candidiasis or herpes simplex was receiving prophylaxis and each responded promptly to therapy. In the prednisone group, two patients developed hyperglycemia and one diabetic increased insulin requirements. CD4 cell counts and plasma HIV-1 RNA levels did not change, but plasma tumor necrosis factor alpha levels and CD38+ CD8+ cells decreased significantly in those taking prednisone. CONCLUSION: Short-term prednisone administration is well tolerated and reasonably safe in advanced HIV-1 disease and decreases immune activation without effects on HIV-1 RNA levels or CD4 cell counts. These results suggest that, in stable HIV-1 disease, these immune activation markers are more likely consequences of but not inducers of HIV-1 replication. PMID- 11273212 TI - Premature atherosclerosis in HIV-infected individuals--focus on protease inhibitor therapy. AB - OBJECTIVE: Lipid disorders associated with the use of protease inhibitors may contribute to the premature development of atherosclerosis. The purpose of the present study was to determine whether the administration of a protease inhibitor containing regimen to middle-aged (30-50 years) HIV-infected individuals for 6 months or longer is associated with an increased prevalence of atherosclerosis. METHODS: High-resolution B-mode ultrasound imaging was used to visualize the femoral and carotid arteries of 68 HIV-negative and 168 HIV-infected individuals, including 136 patients who had received protease inhibitors for 26.8 +/- 8.9 months (mean +/- SD). Atherogenic plaques were defined as a thickening of the intima-media > or = 1200 mm. RESULTS: The proportion of participants with one or more plaques was higher in the HIV-infected group in comparison with the HIV negative group (55 versus 38%; P = 0.02), and so was the prevalence of cigarette smoking (61 versus 46%; P = 0.03) and hyperlipidaemia (56 versus 24%; P < 0.001). The presence of plaque was independently associated with age, male gender, plasma low-density lipoprotein cholesterol levels and smoking. In univariate logistic regression analysis, an association was also found with HIV infection. Among HIV infected subjects protease inhibitor therapy was not associated with the presence of plaque. CONCLUSIONS: A large proportion of the middle-aged HIV-infected individuals examined during this study had one or more atherosclerotic plaques within the femoral or carotid arteries. The presence of peripheral atherosclerosis within this population is not associated with the use of protease inhibitors, but rather with 'classic' cardiovascular risk factors such as smoking and hyperlipidaemia, which are amenable to interventions. PMID- 11273213 TI - Pneumocystis carinii pneumonia and cytomegalovirus infection in children with vertically acquired HIV infection. AB - OBJECTIVES: The outcome of Pneumocystis carinii pneumonia (PCP) in HIV-infected infants is poor, and the role of cytomegalovirus (CMV) co-infection in the course and outcome of PCP is unclear. This study describes the prevalence, clinical characteristics, management and changes in survival over time of vertically HIV infected infants developing PCP and/or CMV infection. METHODS: Data on children with HIV, born in the UK and Ireland and reported to the National Study of HIV in Pregnancy and Childhood, with PCP and/or CMV were combined with clinical information collected from reporting paediatricians. RESULTS: By April 1998, 340 vertically HIV-infected children had been reported, of whom 93 had PCP and/or CMV, as their first AIDS indicator disease; 85 (91%) were infants. Among infants with PCP, 79% were born to mothers not diagnosed as HIV infected, and there was an independent and statistically significant association with breast-feeding, being black African, and developing CMV disease. Median survival after PCP and/or CMV was significantly better in those born between 1993 and 1998 compared with those born before 1993 (P = 0.009), and worse than after other AIDS diagnoses (P = 0.01). Infants with dual infection were more likely to be ventilated (P = 0.003) and receive corticosteroids (P = 0.002) than those with PCP alone. CONCLUSION: Although survival from PCP and CMV has improved over time, these remain serious and potentially fatal infections among infants in whom maternal HIV status is not recognized in pregnancy. Breast-feeding increases the risk of combined PCP and CMV infection, which is associated with severe disease. PMID- 11273214 TI - Neurocognitive performance enhanced by highly active antiretroviral therapy in HIV-infected women. AB - OBJECTIVE: To determine whether highly active retroviral therapy (HAART) is associated with better neurocognitive outcome over time among HIV-infected women with severely impaired immune function. METHODS: A semiannual neurocognitive examination on four tasks was administered: Color Trail Making, Controlled Oral Word Association, Grooved Pegboard and Four-Word Learning. This protocol was initiated in the HIV Epidemiological Research study (HERS) study when a woman's CD4 cell count fell to < 100 x 10(6) cells/l. Immune function (CD4), viral load status and depression severity (CESD) were also assessed semi-annually, along with an interview to determine medication intake and illicit drug use. RESULTS: HAART was not available to any participant at the time of enrollment (baseline), while 44% reported taking HAART at their most recent visit (mean duration of HAART 36.3 +/- 12.6 months). HAART-treated women had improved neurocognitive performance compared with those not treated with HAART. Women taking HAART for 18 months or more showed the strongest neurocognitive performance with improved verbal fluency, psychomotor and executive functions. These functions worsened among women not taking HAART. Substance abuse status, severity of depressive symptoms, age and educational level did not influence the HAART treatment effects on neurocognitive performance. Neurocognitive improvements were strongly associated with the magnitude of CD4 cell count increases. CONCLUSIONS: HAART appeared to produce beneficial effect on neurocognitive functioning in HIV infected women with severely impaired immune systems. Benefits were greatest for women who reported receiving HAART for more than 18 months. PMID- 11273215 TI - Effectiveness of potent antiretroviral therapies on the incidence of opportunistic infections before and after AIDS diagnosis. AB - OBJECTIVES: To determine the effectiveness of potent antiretroviral therapy in reducing opportunistic infections (OI) as both a presenting event and subsequent to an AIDS-defining event. DESIGN AND METHODS: A total of 543 seroconverters and 1470 men with AIDS were compared for the time to development of OI as the presenting AIDS event and as a subsequent event in the 1984-1989, 1990-1992, 1993 1995, and 1996-1998 periods, when the major treatments were no therapy, monotherapy, combination therapy, and potent antiretroviral therapy, respectively. RESULTS: The seroconverters suffered 132 OI and the participants with AIDS had 717 OI. The relative hazard (RH) of OI as the presenting AIDS event declined by 81% in the calendar period when potent antiretroviral therapy was available compared with the monotherapy period. Declines were observed for Mycobacterium avium complex, cytomegalovirus disease, and esophageal candidiasis, but were statistically significant only for Pneumocystis carinii pneumonia. The RH of OI as a secondary infection dropped by 77% in the last calendar period compared with the monotherapy period. A significant decline was observed for all four OI. Prophylactic drug use did not increase in the era of potent antiretroviral therapy. CONCLUSION: The hazard of OI in the era of potent antiretroviral therapy has declined dramatically compared with the era of monotherapy, despite the concurrent decrease in the use of prophylactic drugs. Physicians should consider whether it is necessary to include prophylactic drugs as part of the complex drug regimen for patients on potent antiretroviral therapy. PMID- 11273216 TI - Duration of ruptured membranes and vertical transmission of HIV-1: a meta analysis from 15 prospective cohort studies. AB - OBJECTIVE: To test the a priori hypothesis that longer duration of ruptured membranes is associated with increased risk of vertical transmission of HIV. DESIGN: The relationship between duration of ruptured membranes and vertical transmission of HIV was evaluated in an individual patient data meta-analysis. METHODS: Eligible studies were prospective cohort studies including at least 100 mother-child pairs, from regions where HIV-infected women are counselled not to breastfeed. Analyses were restricted to vaginal deliveries and non-elective Cesarean sections; elective Cesarean section deliveries (those performed before onset of labour and before rupture of membranes) were excluded. RESULTS: The primary analysis included 4721 deliveries with duration of ruptured membranes < or = 24 h. After adjusting for other factors known to be associated with vertical transmission using logistic regression analysis to assess the strength of the relationship, the risk of vertical HIV transmission increased approximately 2% with an increase of 1 h in the duration of ruptured membranes [adjusted odds ratio, 1.02; 95% confidence interval, 1.01-1.04; for each 1 h increment]. There were no significant interactions of duration of ruptured membranes with study cohort or with any of the covariates, except maternal AIDS. Among women diagnosed with AIDS, the estimated probability of transmission increased from 8% to 31% with duration of ruptured membranes of 2 h and 24 h respectively (P < 0.01). CONCLUSIONS: These results support the importance of duration of ruptured membranes as a risk factor for vertical transmission of HIV and suggest that a diagnosis of AIDS in the mother at the time of delivery may potentiate the effect of duration of ruptured membranes. PMID- 11273217 TI - Sexual risk behaviour relates to the virological and immunological improvements during highly active antiretroviral therapy in HIV-1 infection. AB - OBJECTIVES: To evaluate the effect of highly active antiretroviral therapy (HAART) on the sexual behaviour of homosexual men, we conducted (i) an ecological study of time trends in sexual behaviour and sexually transmitted diseases; (ii) a HAART-effect study focused on the practice of unprotected anogenital sex. DESIGN: Subjects were participants in the ongoing Amsterdam Cohort Studies (ACS) among homosexual men, initiated in 1984. Data for (i) represented all ACS visits by HIV-1-positive and -negative participants who entered ACS at or below 30 years of age and were followed until 35 years (n = 1062). Data for (ii) represented all ACS visits of HIV-1-positive men from 1992 to 2000 (n = 365), of whom 84 were HAART recipients with at least 2 months of behavioural follow-up. RESULTS: (i) After HAART became generally available in July 1996, unprotected sex was practised more frequently and the incidence of gonorrhoea was higher compared to March 1992-June 1996 among HIV-1-negative and -positive men, respectively. (ii) Among HIV-1-positive men, a higher level of unprotected sex with casual partners was observed after HIV-1 RNA became undetectable and CD4 cell counts increased with the use of HAART. Notably, in individuals who did not receive HAART, high HIV-1-RNA levels (above 10(5) copies/ml) were likewise related to unprotected sex with casual partners. CONCLUSION: Data support the need for the reinforcement of safe sex prevention messages among HIV-1-negative men, and our data also provide a lead for redirecting and tailoring current prevention strategies to the needs of HIV-1-positive men. PMID- 11273218 TI - Method of feeding and transmission of HIV-1 from mothers to children by 15 months of age: prospective cohort study from Durban, South Africa. AB - OBJECTIVE: To determine the risk of HIV transmission by infant feeding modality. DESIGN AND SETTING: A prospective study in two hospitals in Durban, South Africa. PARTICIPANTS: A total of 551 HIV-infected pregnant women enrolled in a randomized trial of vitamin A. INTERVENTIONS: Women self-selected to breastfeed or formula feed after being counselled. Breastfeeders were encouraged to practice exclusive breastfeeding for 3-6 months. MAIN OUTCOME MEASURES: Cumulative probabilities of detecting HIV over time were estimated using Kaplan-Meier methods and were compared in three groups: 157 formula-fed (never breastfed); 118 exclusively breastfed for 3 months or more; and 276 mixed breastfed. RESULTS: The three feeding groups did not differ in any risk factors for transmission, and the probability of detecting HIV at birth was similar. Cumulative probabilities of HIV detection remained similar among never and exclusive breastfeeders up to 6 months: 0.194 (95% CI 0.136-0.260) and 0.194 (95% CI 0.125-0.274), respectively, whereas the probabilities among mixed breastfeeders soon surpassed both groups reaching 0.261 (95% CI 0.205-0.319) by 6 months. By 15 months, the cumulative probability of HIV infection remained lower among those who exclusively breastfed for 3 months or more than among other breastfeeders (0.247 versus 0.359). CONCLUSION: Infants exclusively breastfed for 3 months or more had no excess risk of HIV infection over 6 months than those never breastfed. These findings, if confirmed elsewhere, can influence public health policies on feeding choices available to HIV-infected mothers in developing countries. PMID- 11273219 TI - Vaginal lavage with chlorhexidine during labour to reduce mother-to-child HIV transmission: clinical trial in Mombasa, Kenya. AB - OBJECTIVES: To evaluate the effect of vaginal lavage with diluted chlorhexidine on mother-to child transmission of HIV (MTCT) in a breastfeeding population. METHODS: This prospective clinical trial was conducted in a governmental hospital in Mombasa, Kenya. On alternating weeks, women were allocated to non-intervention or to intervention consisting of vaginal lavage with 120 ml 0.2% chlorhexidine, later increased to 0.4%, repeated every 3 h from admission to delivery. Infants were tested for HIV by DNA polymerase chain reaction within 48 h and at 6 and 14 weeks of life. RESULTS: Enrolment and follow-up data were available for 297 and 309 HIV-positive women, respectively, in the non-lavage and the lavage groups. There was no evidence of a difference in intrapartum MTCT (17.2 versus 15.9%, OR 0.9, 95% CI 0.6-1.4) between the groups. Lavage solely before rupture of the membranes tended towards lower MTCT with chlorhexidine 0.2% (OR 0.6, 95% CI 0.3 1.1), and even more with chlorhexidine 0.4% (OR 0.1, 95% CI 0.0-0.9). CONCLUSION: The need remains for interventions reducing MTCT without HIV testing, often unavailable in countries with a high prevalence of HIV. Vaginal lavage with diluted chlorhexidine during delivery did not show a global effect on MTCT in our study. However, the data suggest that lavage before the membranes are ruptured might be associated with a reduction of MTCT, especially with higher concentrations of chlorhexidine. PMID- 11273221 TI - Predictors of HIV risk behavior among Russian men who have sex with men: an emerging epidemic. AB - BACKGROUND: Russia is experiencing one of the sharpest increases in HIV incidence in the world. Almost no research has examined patterns of risk behavior among Russian men who have sex with men (MSM). DESIGN AND METHODS: A total of 434 MSM were surveyed in all of St. Petersburg's gay-identified clubs during June 2000. Men completed questionnaires about their sexual practices, AIDS risk knowledge, safer sex attitudes, behavior change intentions, perceived safer sex norms, and fatalism. RESULTS: Most MSM were bisexual; 79% had female partners in their lives and 37% had female partners in the previous 3 months. Sexually transmitted disease treatment was reported by 32% of the men, 23% had sold sex to gain money, and knowledge about critical HIV risk-reduction steps was low. Of all men surveyed, 38% had unprotected anal sex in the previous 3 months, consistent condom use was reported by only 30% of men, and most recent anal intercourse occasions 37% of particpants'. Regression analyses showed that high-risk behavior was predicted by poor safer sex attitudes, weak behavior change intentions, low knowledge about AIDS risk, perceived peer norms that did not support safer sex, and having a boyfriend. CONCLUSION: To avert a widespread epidemic, HIV prevention interventions for Russian MSM are critically needed. Factors predicting risk were consistent with those found among MSM in other countries early in the HIV epidemic. However, unique cultural factors, including frequent bisexual behavior, the 'newness' of openly gay communities in Russia and lack of community experience in dealing with AIDS, require HIV prevention program tailoring. PMID- 11273220 TI - Continued high HIV-1 incidence in a vaccine trial preparatory cohort of injection drug users in Bangkok, Thailand. AB - BACKGROUND: A large epidemic of HIV-1 subtype B began among injection drug users (IDUs) in Bangkok in 1988. Despite ongoing prevention efforts, HIV-1 prevalence among IDUs remained at 30-50% through the 1990s. OBJECTIVES: To measure the incidence of HIV-1 infection and related risk factors to guide prevention efforts and to evaluate the feasibility of conducting an HIV vaccine efficacy trial. DESIGN AND METHODS: A prospective cohort study in which IDUs attending methadone treatment programs in Bangkok were screened during 1995-1996 for enrollment into the study. IDUs found to be HIV-seronegative on two occasions were offered enrollment with follow-up visits every 4 months. On each visit participants were evaluated with a questionnaire and serologic testing. RESULTS: A total of 1209 HIV-negative IDUs were enrolled. Through the end of 1998, the overall HIV-1 incidence rate was 5.8 (95% confidence interval, 4.8-6.8) per 100 person-years of follow-up. HIV-1 subtypes E and B accounted for 79 and 21% of infections, respectively. On multivariate analysis, HIV-1 seroconversion was primarily associated with the frequency of heroin injection, the sharing of injection equipment, and incarceration, especially with drug injection. Sexual behavior was not associated with increased risk for HIV-1. Risk factors for infection with HIV 1 subtypes E and B were similar. CONCLUSION: HIV-1 transmission risk remains high among Bangkok IDUs despite methadone treatment and other current prevention strategies. There is an urgent need to address this ongoing epidemic, especially in jails and prisons. This study led to the initiation in 1999 of a phase III HIV 1 vaccine efficacy trial in this population. PMID- 11273222 TI - Mitochondrial toxicity hypothesis for lipoatrophy: a refutation. PMID- 11273223 TI - Low lipolytic enzyme activity in patients with severe hypertriglyceridemia on highly active antiretroviral therapy. PMID- 11273224 TI - An argument for routine therapeutic drug monitoring of HIV-1 protease inhibitors during pregnancy. PMID- 11273225 TI - Large hepatic mitochondrial DNA deletions associated with L-lactic acidosis and highly active antiretroviral therapy. PMID- 11273226 TI - Drug interaction between St John's wort and nevirapine. PMID- 11273227 TI - Topoisomerase II inhibitor induced leukemia in a patient with AIDS. PMID- 11273228 TI - Factors associated with the response to antiretroviral therapy among HIV-infected patients with and without a history of injection drug use. PMID- 11273229 TI - Dynamics of seminal plasma HIV-1 decline after antiretroviral treatment. PMID- 11273230 TI - On the recent sharp increase in HIV detections in Cuba. PMID- 11273231 TI - Reduction of HIV transmission in HIV-discordant couples wishing to conceive. PMID- 11273232 TI - Failure of postexposure prophylaxis after sexual exposure to HIV. PMID- 11273233 TI - Mucosal IgA in exposed, uninfected subjects: evidence for a role in protection against HIV infection. PMID- 11273234 TI - GMC's proposals for revalidation. Effective revalidation system looks at how doctors practise and quality of patients' experience. PMID- 11273235 TI - GMC's proposals for revalidation. Purpose of revalidation process must be agreed on. PMID- 11273236 TI - GMC's proposals for revalidation. Appraisal is helpful only if done well. PMID- 11273237 TI - GMC assessment of Fergusson case was not at fault. PMID- 11273238 TI - Routine vaccination and child survival in Guinea-Bissau. Author's reply to commentary. PMID- 11273239 TI - Routine vaccination and child survival in Guinea-Bissau. Lessons can be learnt from this study. PMID- 11273240 TI - Routine vaccination and child survival in Guinea-Bissau. WHO responds to Guinea Bissau report. PMID- 11273241 TI - Vaccinations as risk factors for ill health in veterans of the Gulf war. Conclusion may be flawed by inadequate data. PMID- 11273242 TI - Tracheal stenosis can occur 20 years after intubation. PMID- 11273243 TI - Contraception in general practice before teenage pregnancy. Inappropriate selection of cases and controls biased study. PMID- 11273244 TI - Contraception in general practice before teenage pregnancy. Not all teenagers are sexually active. PMID- 11273245 TI - Conjunctival impression cytology for detection of subclinical vitamin A deficiency--can it be used routinely? PMID- 11273246 TI - Conjunctival impression cytology for detection of early vitamin A deficiency in children. AB - Children presenting with recurrent infections have a high risk of developing vitamin A deficiency. Conjunctival impression cytology (CIC) was used in the present study in such children to detect subclinical deficiency and to monitor the outcome after therapy. Seventy children with history of recurrent infections, and 10 healthy children in the age group of six months to five years were included in the study. CIC was performed using millipore filter paper and stained with PAS stain. A three tier grading system was used consisting of normal, borderline abnormal and abnormal for interpretation. Vitamin A supplementation was given in children in the latter two categories. Repeat cytology showed reversal to normal in these children. Hence in children with high risk of developing vitamin A deficiency, it is suggested to do CIC for detection and monitoring it. PMID- 11273247 TI - Consumption of 'country liquor' and its relation to alcoholic liver disease in Mumbai. AB - The amount of alcohol intake required for the development of liver disease has been determined in Western populations; corresponding figures in Indians, many of whom consume locally brewed liquors, are not known. We studied 328 patients from a public hospital in Mumbai who admitted to regular alcohol consumption, to determine the pattern of alcohol consumption and its relation to liver disease. Liver disease was more common in those who consumed illicitly-brewed as compared to licit liquor. Daily drinking, volume of consumption > 200 ml per day, and duration of drinking > 14 years were each significantly more common in those with liver disease. A cumulative intake of > 2000 ml. years, calculated as the product of volume (ml per day) and duration (years), was a reliable cut-off level for association with liver disease (sensitivity 65%, specificity 77%) and cirrhosis (sensitivity 70%, specificity 59%). The content of alcohol in these liquors, estimated in 23 samples, ranged from 23-36.1 g/100 ml, being lower in the illicit liquors. Thus, in Mumbai, alcoholic liver disease occurs more commonly with consumption of illicit liquor (despite its lower alcohol content); liver involvement appears earlier and with lower consumption levels than in the West. PMID- 11273248 TI - Estimation of fasting and post oral glucose serum insulin levels in hypertensive and obese subjects. AB - Insulin resistance and consequent hyperinsulinemia has been documented as a frequent occurrence in hypertension and obesity. Fasting and post oral glucose serum insulin levels serve as reliable markers of the state of insulin resistance. In this study, fasting serum insulin levels, as measured by radio immunoassay method, were found to be more increased in obese individuals than in those with hypertension alone. While, post-oral glucose load serum insulin levels increased more in hypertensive individuals than those with obesity alone. In subjects who had obesity and hypertension together, the fasting serum insulin levels did not show much change while post oral glucose load serum insulin levels greatly increased suggesting a compounding effect. PMID- 11273249 TI - Exposure to Parthenium hysterophorous pollen extract leads to bronchospasm in stable patients of bronchial asthma. AB - Role of Parthenium hysterophorous as an allergen evoking bronchial hyper responsiveness was assessed in twenty five adult patients with stable asthma and ten healthy controls. Assessment was made with the help of skin prick and bronchial provocation tests (BPT) using a commercially available Pathenium extract. Eleven patients (44%) of the study group had a positive skin reaction and 4 (16%) showed a significant fall in FEV1 and PEFR (p < 0.05) on bronchial provocation. In the control group only one patient (10%) had a positive skin test while there was none with a positive bronchial challenge. There was a significant fall in the mean values of FEV1 and PEFR over base line after the BPT in patients of asthma than controls. It is concluded that a significant proportion of bronchial asthma patients are sensitized to Parthenium hysterophorous and it may act as a cofactor in seasonal exacerbation of their symptoms. PMID- 11273250 TI - The value of sonography for estimation of liver volume by using a simple geometric formula ascertained in cadaveric livers. AB - Accurate assessment of liver size and its volume are important. However, as the clinical methods do not produce reliable results especially when the liver is shrunken, and the previous attempts to accurately assess liver volume by radio isotopes, CT scans and computer assisted ultrasonography have not gained popularity due to high cost and complex techniques, the need to devise a simpler technique for estimation of liver volume continues. In doing so, we estimated volume of 10 cadaveric livers by water displacement technique to serve as the reference value. Thereafter, assuming the shape of liver like a right-angled pyramid, we calculated its volume by a simple geometric formula of '1/2 abc'. However, a reduction of 15% was made from this to compensate for depression on the inferior surface of liver. This method was subsequently implemented to assess liver volume of 14 healthy individuals and 20 patients of fulminant hepatic failure (FHF) by using ultrasonography. Our findings revealed smaller liver volume in Indians as compared to the reported Western figures, and a significantly smaller liver volume in females as compared to males. The liver volume of 6 FHF patients who died was significantly smaller (696.5 +/- 143.5 cm3) as compared to that of the 14 FHF patients who survived (1083 +/- 365.3 cm3). Moreover, mortality rate was 100 per cent in 3 patients of FHF who showed markedly shrunken globular liver with a liver volume of less than 500 cm3. Thus, a markedly reduced liver size in FHF patients suggests a poor prognosis. However, since the number of FHF patients in the present series is small, it is our contention that a larger series is mandatory to confirm the findings of the present study. PMID- 11273251 TI - Pictorial CME. Presentation of lethargy and pellagra lesions diagnosed as hypothyroidism. PMID- 11273253 TI - A pilot study of polyunsaturated phosphatidyl choline in fulminant and subacute hepatic failure. AB - Present pilot study was conducted to evaluate efficacy and safety of polyunsaturated phosphatidyl choline (PPC) in a phase III clinical trial in patients of fulminant and subacute hepatic failure over one year period in a prospective randomised blinded controlled design. We found that in patients of fulminant hepatic failure, recovery period from encephalopathy was faster and mortality rate lower in the test group of patients who received PPC in a dose of 350 mg thrice daily for 6 to 8 weeks as compared to the control groups who did not receive it. In the patients of subacute hepatic failure, recovery from encephalopathy was faster, mortality rate lower and regression of ascites was significantly higher (P = 0.0022) in test group of patients who received PPC as compared to the control group. However, as the number of patients in the present pilot study is small, we propose that larger clinical trials are warranted in this direction to prove the efficacy and safety of PPC in fulminant and subacute hepatic failure. PMID- 11273254 TI - Plaque rupture or erosion? The enigma of acute myocardial infarction. PMID- 11273255 TI - Leptin--the fat controller. AB - Obesity is a common health disorder in humans and is inherited genetically. Though several theories have been proposed in the past to understand the mechanisms underlying the control of obesity, the recent discovery of leptin (OB) has made the obesity research interesting. OB, a product of ob gene is a 16 KD protein, secreted by the adipocytes. It acts through its receptor (OB-R), which is a product of db gene. ob and OB-R in conjunction with neuropeptide Y, melanocyte stimulating hormone and melanocortin-4 receptor have been found to control adiposity. Though several issues pertaining to ob need to be addressed, it is anticipated that future treatment of obesity may depend on our understanding of the action(s) of leptin and its associated molecules and receptors. PMID- 11273252 TI - Evaluation and selection of living related kidney donors--our experience in a government hospital. AB - In a state-funded, live related-donor kidney transplantation programme, 616 eligible end stage renal failure (ESRF) patients were seen over a four-year period. 73% of them had potential related donors, 64% of whom were willing to donate. Fear of surgery, non-congenial pre-morbid relationships and discouragement by family members were the most common reasons for unwillingness to donate. After investigations, 76% of the willing donors were found to be fit. ABO incompatibility, lymphocyte cross-match positivity and anatomic abnormalities were the most common grounds for non-acceptance. Sixty eight percent of the willing, fit donors finally donated their kidneys, patient-death and donor recipient withdrawal before surgery accounting for the remaining. One hundred and forty eight patients underwent renal transplantation. Two-thirds of the donors were females, mothers (37%) forming the single largest group. Eight five percent of the recipients were males. Overall, only 35% of the eligible ESRF patients had related, willing and fit donors attesting to the need for an active, cadaver donor transplantation programme. PMID- 11273256 TI - Tachyarrhythmias in Wolff-Parkinson-White syndrome diagnosis and management. PMID- 11273257 TI - Stress echocardiography. PMID- 11273258 TI - Cytomegalovirus pneumonia in a rheumatoid arthritis patient on low dose methotrexate. PMID- 11273259 TI - Syndrome of inappropriate antidiuretic hormone secretion occurring in association with urinary tract infection. PMID- 11273260 TI - Unusual manifestations of thalamic strokes. AB - Four patients of thalamic strokes with different symptoms are reported. The first had thalamic haemorrhage and developed delayed blepharospasm. The second patient had occlusion of posterior cerebral artery causing infarction of lateral thalamus and occipital lobes. The remaining two patients exhibited ipsilateral hemisensory loss and hemiataxia in absence of hemiparesis (thalamic ataxia). Both had circumscribed lesions in lateral thalamus. 'Thalamic ataxia' has a distinct localizing value. Thalamic strokes produce heterogenous clinical manifestations attributed to the involvement of different nuclei. PMID- 11273262 TI - Percutaneous drainage of hydatid cyst an alternative to surgery. PMID- 11273261 TI - Emphysematous pyelonephritis. AB - Emphysematous pyelonephritis is a severe form of acute pyelonephritis, characterised by fever, abdominal pain, nausea and vomiting, associated with intraparenchymal and perirenal gas production. It is often diagnosed radiologically, by plain films of abdomen, ultrasonogram and/or CT scan and often needs surgical drainage. We report a case which could be diagnosed clinically because of extensive surgical emphysema in a diabetic patient which was successfully managed by a combined medical and surgical approach. PMID- 11273263 TI - Congenital valvular aortic and pulmonary stenosis. PMID- 11273264 TI - Penile carcinoma in a seropositive HIV patient. PMID- 11273265 TI - Cysticercal dementia. PMID- 11273266 TI - Amoebic liver abscess in pregnancy. PMID- 11273267 TI - Intermittent hydrarthrosis. PMID- 11273268 TI - Acute leukemia with uncommon extramedullary lesions. PMID- 11273269 TI - Motor ataxia--an unusual presentation of mushroom poisoning. PMID- 11273270 TI - An evaluation of cost variation in different brands of antihypertensive agents. PMID- 11273271 TI - Diagnostic value of ELISA and ADA in tuberculosis--need for more specific tests. PMID- 11273272 TI - Tuberculous meningitis. PMID- 11273273 TI - Scientific superstition. PMID- 11273274 TI - Hepatopulmonary syndrome. PMID- 11273275 TI - Kimura's disease associated with nephrotic syndrome. PMID- 11273276 TI - Appeal for uniformity in the dosage formulations. PMID- 11273277 TI - Is mitral valve area estimation a sine qua non in the evaluation of mitral stenosis? PMID- 11273278 TI - Accredited continuing medical education for the association members--how do we go ahead? PMID- 11273279 TI - Selective preservation of reproductive function in Sheehan's syndrome. PMID- 11273280 TI - Prevalence of islet cell antibodies and its correlation with glucose and insulin response to oral glucose tolerance test in 1st degree relatives of insulin dependent diabetes mellitus probands. AB - 93 first degree relatives (1st DR) of insulin dependent diabetes mellitus (IDDM) patients were investigated for detection of islet cell antibodies (ICA) and beta cell functional status. ICA were detected in 26.9% Ist DR subjects (25/93), equally in parents, siblings and offspring. Normal (n = 16), impaired (n = 5) and diabetic (n = 4), glucose curves were seen in 1st DR. Low insulin levels were observed in parents and siblings with normal glucose tolerance test (N-GTT) at 90 min (p < 0.05), and (p < 0.0005) relatives with impaired glucose tolerance and diabetes. Insulin release to glucose (IRG-insulinogenic index) in control group was 352 +/- 42 mu U/mg. From the group of 25 ICA positive cases, 4 had mean IRG of 394 +/- 70 mu U/mg (group A) comparable to control, and had N-GTT; 12 had mean IRG of 107 +/- 15.9 mu U/mg (group B) significantly low (P < 0.005) compared to controls and group A and 9 showed IRG of 75 +/- 29.3 mu U/mg, lower than group B (NS) with abnormal response to glucose load. Loss of insulin secretory ability thus can precede hyperglycemia by years. The ICA positive relatives were grouped based on the immunological status with their probands. ICA status in probands does not give an idea about ICA status in their relatives. This indepth study leads to understand the correlation of genetic, metabolic and immunological parameters for early detection of IDDM in first degree relatives. PMID- 11273281 TI - Profile of young acute myocardial infarction in Harayana. AB - Profile of acute myocardial infarction (AMI) in young patients (below 40 years) was studied in a rural/semi-urban population. Out of the total 338 patients who were admitted to ICCU over a period of one year, 65 (19.2%) were aged 40 years or below (Range 14-40 years). Male:female ratio was 20:1. Majority of these young patients were thinly built, engaged in heavy physical work and belonged to lower socio-economic group. Smoking was the most common risk factor (87%); other risk factors were few. Majority of these young patients ignored chest pain and reported late to the hospital. However, despite this, incidence of complications/mortality was less in comparison to their older counterparts. The overall mortality was only 6% as compared to 21% in older age group. The study focuses our attention to the rising incidence of AMI in young individuals even in populations least prone to ischaemic heart disease. Smoking was the only modifiable risk factor which needs to be curbed with full force. PMID- 11273282 TI - Diagnostic yield in computed tomography guided stereotactic biopsies. AB - Fifty three patients underwent computerised tomography (CT) guided stereotactic biopsies from different CT defined zones of attenuation with the Leksell stereotactic apparatus from October 1993 through January 1995. Multiple lesions were seen in 16 cases and 3 of them had multiple rim enhancing lesions. Astrocytoma was the most common histological diagnosis and thalamus was the commonest site of these tumours. The overall positivity rate was 98.2%. Positive yield from the centre of the lesion, peripheral and perilesional areas was 92.1%, 54.7% and 17.6%, respectively. The definite pathological diagnosis was made in 81.1% of cases. Post-operative neurological worsening was seen in 6 patients, of which 2 recovered without any surgical treatment, in 1 patient ventriculo peritoneal shunt was done post-biopsy whereas in another evacuation of hamatoma was done which relieved headache and vomiting while 2 patients (3.7%) died. PMID- 11273283 TI - The spectrum of chronic autoimmune hepatitis. AB - Autoimmune hepatitis (AIH) is characterised by the presence of periportal hepatitis coupled with the presence of autoantibodies in the serum. We report our experience with 10 cases (females--8, males--2) who presented to the rheumatology clinic with either articular or extra-articular manifestations. Three patients (1 SLE, 1 Sjogren's and 1 RA), satisfied the criteria for an underlying rheumatic disease (secondary AIH) while, others had primary AIH. Median duration of hepatic involvement was 6 months and the varied presentations were noted. Articular disease ranged from arthralgias, palindromic arthritis to persistent non erosive/non-deforming arthritis (Jaccoud's arthritis). Autoimmune thrombocytopenia was seen in 2 and autoimmune hypo and hyperthyroidism were seen in 3 patients each. Anti-nuclear antibody was positive in 9/10 (6 with speckled pattern and 3 with homogenous pattern) and anti-mitochondrial antibodies were positive in three. Hepatitis C virus (HCV) markers were positive in 1, who probably had viral hepatitis with dominant autoimmune features. All have been started on steroids (5 patients--1 mg/kg dose, 1 patient--0.5 mg/kg dose and 3 patients--0.25 mg/kg dose). The HCV positive patient was on a low dose steroid (0.25 mg/kg) and interferron treatment was contemplated before she was lost to follow up. Four patients are also on azathioprine in the dose of 2 mg/kg/day. Of the 6 patients who are under regular follow up, the liver parameters have normalised in 5 and one showed hypoalbuminaemia with normal enzyme levels at the last follow up. PMID- 11273285 TI - Interictal brain 99m Tc-HMPAO SPECT study in chronic epilepsy. AB - We performed interictal brain 99m Tc-HMPAO study in eight cases of chronic epilepsy. SPECT (single photon emission computed tomography) study showed abnormality in seven cases. As compared to computed tomography (CT) scan and electroencephalogram (EEG) which were positive in 25% and 62% cases respectively it showed abnormality in 87%. In two cases where CT scan and EEG both were normal, SPECT showed areas of hypoperfusion. In one case where EEG indicated a bilateral focus, SPECT study showed a clearly defined unilateral focal hypoperfusion defect. Areas of hyperperfusion were not seen in any of our cases. Our results indicate that HMPAO SPECT is more sensitive than CT scan and EEG, in localising an epileptogenic focus in cases of chronic epilepsy. PMID- 11273284 TI - Helicobacter pylori infection and erosive gastritis. AB - One hundred and eleven patients were included in the study. Thirty seven had erosive gastritis, thirty four chronic gastritis and forty were controls without any gastrointestinal diseases confirmed by symptoms and upper gastrointestinal endoscopy. Patients with erosive gastritis were divided into non-steroidal anti inflammatory drug (NSAID) users and non-users. H pylori status was determined by urease test, serology and/or histology. The prevalence of H pylori was compared between the various groups. The prevalence of H pylori infection in erosive gastritis, chronic gastritis and controls was 68%, 76% and 65%, respectively, the difference was not significant (P > 0.05), 8 out of 11 patients with erosive gastritis and NSAID use (73%) were positive for H pylori. Likewise 17/26 patients with erosive gastritis without NSAID use (65%) were positive for H pylori (P > 0.05). Body of the stomach (65%) was the commonest site for erosions compared to antrum (43%) or fundus (27%) (P < 0.02). H pylori infection does not predispose to erosive gastritis. NSAID use does not affect H pylori prevalence. Routine H pylori eradication is, therefore, not indicated in patients with erosive gastritis infection. Body of the stomach is the most predominant site for erosions. PMID- 11273286 TI - Pictorial CME. Electrocardiogram reveals complete left bundle branch block. PMID- 11273287 TI - Central venous catheter related infections in a tertiary care hospital. AB - Intravascular catheters are increasingly important causes of nosocomial infections. Catheter related complications range from local exit site or tunnel infections to frank bacteremias. A semiquantitative method of culture of central venous catheters (CVC) was done in our hospital from January to December 1996. A total of 119 catheter tips sent to the Microbiology Department were cultured and 11 (9.24%) showed significant growth with associated blood stream infection. 14 (11.76%) of the CVCs showed scanty or less than 15 colonies in roll or contents and there was no associated blood stream infection. 7 (5.88%) showed moderate to heavy growth in roll and contents and there was no blood stream infection. The age groups ranged from 2 months to 66 years. The results of the study indicate that Gram negative organisms formed the predominant isolates. Gram negative isolates included Klebsiella species, Enterobacter species, E. coli species, Serratia and non-fermenting Gram negative bacilli. Coagulase negative staphylococcus which is often believed to be an important pathogen was not associated with bacteremia or septicemia in our hospital, during this study period. Considering the fact that 1553 operations were performed during the study period, the infection rate through CVC's would work out to a negligible 0.71%. PMID- 11273289 TI - Accredited continuing medical education for all API members. PMID- 11273288 TI - Angiotensin converting enzyme inhibitors and cough--a north Indian study. AB - Cough is an important side effect of Angiotensin Converting Enzyme Inhibitor (ACEI) therapy. The incidence of cough was investigated in a prospective 8 week study in 250 hypertensive patients receiving ACEI alone or in combination with other agents. Enalapril (5-20 mg/day), Lisinopril (5-20 mg/day), Captopril (25-75 mg/day) or Ramipril (5-15 mg/day) was prescribed to patients, who were followed up at weekly visits. Cough developed in 73 of the 250 patients i.e. an incidence of 29.2%. Females had a higher incidence of cough as compared to males--37.9% versus 15.5% (p < 0.001) and there was no significant difference in the cough incidence in the various age groups. A dry, non-productive cough developed in all patients within 4 weeks of ACEI initiation. Increased nocturnal intensity of cough was reported by 79.4% patients. Cough incidence was 34.4%, 24.3% and 18.1% in patients on Enalapril, Ramipril and Lisinopril, respectively. Cough was not dose related and was not related to smoking. There was no statistically significant difference among patients on ACEI alone or in combination with beta blockers, calcium channel blockers or diuretics. Of the 18 patients with ACEI induced cough who received Indomethacin, 50 mg bid, 8 reported complete cure and cough was reduced in intensity in the remaining ten. PMID- 11273290 TI - Chronic hepatitis--changing trends. AB - The clinical expressions, courses and consequences of hepatitis caused by different viruses (A,B,C,D,E,G) are different. Diagnosis of hepatitis is incomplete unless its etiology is specified and for chronic hepatitis, the etiology is apparent in almost all cases when autoimmune and metabolic diseases are also included. Hence, the classification based only on histology is not adequate and emphasis should also be on the etiology. The prognostic indices governing a response to interferon therapy in patients with chronic viral hepatitis have advanced with the knowledge of role played by viral genotypes, serum ferritin, hepatic iron concentration, viral quantification, and severity of histology. There have been recent changes in the definition and classification of autoimmune hepatitis as well as there is availability of newer immunosuppressive agents with encouraging results. PMID- 11273291 TI - Drug safety--whose concern? PMID- 11273292 TI - Patient compliance to drug therapy. PMID- 11273293 TI - 74-year old woman with hypercalcemia. PMID- 11273294 TI - Vaccination in India. PMID- 11273295 TI - Regular ovulatory menstrual cycles in a case of Sheehan's syndrome. AB - In this report we describe an unusual case of postpartum pituitary necrosis who had clinical and biochemical suggestion of decreased thyrotroph, somatotroph, lactotroph, and corticotroph reserve but continued to have regular ovulatory menstrual cycles. PMID- 11273296 TI - Pregnancy in Sheehan's syndrome: a report of three cases. AB - Postpartum pituitary necrosis (Sheehan's syndrome) is a relatively common clinical disorder in developing and underdeveloped areas of the world. Sheehan's syndrome has a spectrum of presentations. Spontaneous pregnancy in such patients is a rare occurrence. Three patients with clinical and hormonal evidence of postpartum pituitary necrosis conceived spontaneously during the follow up period. Pregnancy though rare may occur either due to sparing or recovery of gonadotroph function in such patients. PMID- 11273298 TI - Calcinosis universalis in a case of progressive systemic sclerosis. PMID- 11273297 TI - Two cases of neurosarcoidosis presenting as peripheral neuropathy and stroke in young. PMID- 11273299 TI - Spontaneous remission in idiopathic hyperprolactinemia. AB - In this report we describe a 37 year old lady who was demonstrated to have hyperprolactinemia causing amenorrhea-galactorrhea syndrome. Computerized tomography scan done twice did not reveal any sellar or suprasellar abnormality and there was no clinical or biochemical evidence of primary hypothyroidism. She had regression of galactorrhea, resumed regular menstrual cycles, and conceived twice on bromocriptine therapy. Following her second delivery she noticed spontaneous remission of galactorrhea and, prolactin levels estimated multiple times were normal. PMID- 11273300 TI - Xanthoma in a young woman with systemic lupus erythematosus. PMID- 11273301 TI - Seasonal relapse of healed duodenal ulcer. PMID- 11273302 TI - Diabetic ketoacidosis in Friedreich's ataxia--not a mere coincidence. PMID- 11273303 TI - Crescentic glomerulonephritis in association with traumatic arteriovenous fistula following gun shot injury. PMID- 11273304 TI - Orally administered serratiopeptidase: can it work? PMID- 11273305 TI - Another way of looking at the efficacy of pralidoxime in organophosphate poisoning. PMID- 11273306 TI - Pralidoxime in treatment of organophorus poisoning. PMID- 11273307 TI - Angiotensin converting enzyme estimation in pyrexia of unknown origin with bilateral hilar lymphadenopathy--an essential investigation. PMID- 11273308 TI - Idiopathic pulmonary hemosiderosis. PMID- 11273309 TI - Need for injection benzathine penicillin-9 lacs. PMID- 11273310 TI - Rheumatic pulmonary valve disease. PMID- 11273311 TI - Zoster associated pain. PMID- 11273312 TI - Herpes zoster and post-herpetic neuralgia--a clinical trial of aspirin in chloroform for anodyne. AB - Pain associated with Herpes Zoster (HZ) and Post-herpetic Neuralgia (PHN) has been a challenging task to manage with ease. Topical aspirin dissolved in chloroform is an effective means of reducing pain due to HZ and PHN in most patients. The locus of pain origin and analgesia induced by topical aspirin is supposed to be at cutaneous free nerve ending pain receptors. The present study was conduced in fifty two patients of HZ and PHN. Pain intensity before and after the application of drug was measured with help of Sort Form McGill Pain Questionnaire (SE-MPQ). Most of the patients experienced relief of pain within 1 5 minutes after the aspirin-chloroform application. Maximum relief was achieved in about 30-40 minutes and persisted for 5-6 hrs. In the beginning 3-4 applications were required but frequency decreased gradually as the pain abated. PMID- 11273313 TI - Left ventricular mural thrombus following myocardial infarction--a follow up study. AB - Left Ventricular mural thrombus detected by echocardiography in 41 patients after myocardial infarction (MI) were followed up for 4 years. Thirty eight patients were males and mean age of study population was 52.4 years. Echocardiography revealed predominant mural type of thrombi (38 patients) and none showed mobility. All of them showed regional wall motion abnormality (RWMA) and Left Ventricular (LV) aneurysm was found in 28 patients. Embolic events were observed in 6 patients and 1 patient died following embolic stroke. Follow up study revealed persistent left ventricular thrombus in 19 patients and risk factors detected were severe LV dysfunction and LV aneurysm. Six patients had spontaneous resolution and 6 had resolution of the thrombus after anticoagulants. While anticoagulant therapy was very effective in preventing embolism after recent MI (within 3 weeks), it was found not useful in chronic LV thrombi. We observed ongoing embolic risk in chronic LV thrombi with LV aneurysms but a randomised trial is needed to decide the role of anticoagulants in such situation. PMID- 11273314 TI - Association of HLA B27 antigen in Indian patients of ankylosing spondylitis and other autoimmune diseases. AB - One thousand three hundred and forty clinically suspected patients of Ankylosing Spondylitis (AS) and other autoimmune diseases and 5000 controls were studied to detect the association of HLA B27 antigen amongst them. Other alleles studied include HLA B7, B40 (B60), B22(B55), B13, etc. Our findings show a considerable and consistent association of HLA B27 with AS irrespective of the community to which the patient, belonged his hygiene or socio-economic conditions. We also found that people in the age group of 21-39 were the most vulnerable, when number of affected individuals or severity of the disease were taken into consideration. Male members showed a preponderance over females in HLA B27 positivity. Detection of HLA B27 could help in the diagnosis of AS. Patients suffering from other autoimmune diseases such as rheumatoid arthritis, psoriasis, Reiter's syndrome and uveitis and patients with inflammatory bowel disease, colitis, eczema, bacillary or fungal infection were also found to be HLA B27 positive. A study of other alleles shows that even they sometimes associate AS and other autoimmune diseases. PMID- 11273315 TI - Prevalence of stroke in rural community--an overview of Indian experience. AB - Various prevalence studies of stroke conducted in different regions of rural India have been analyzed and compared with prevalence study of stroke in 51,165 rural population of Haryana. The prevalence of stoke varies in different regions of country and ranges from 40 to 270/100,000 rural population. The prevalence rates correspond to that in urban areas in same region but is much lower than stroke prevalence in metropolitan cities in India and from reported prevalence of 400-800/100,000 in Western countries. Ethnic, socio-economic and dietary factors may be responsible for this variance. PMID- 11273316 TI - Effect of enalapril therapy on ventilatory pulmonary function tests in hypertensive patients. AB - Fifty newly diagnosed nonsmoker patients suffering from mild to moderate hypertension (diastolic BP 90 to 114 mmHg), randomly selected and not having respiratory or other systemic diseases which may affect pulmonary functions were subjected to thorough interrogation and clinical examination. Twenty five normal age and sex matched healthy volunteers served as control. All patients and controls were subjected to ventilatory pulmonary function tests (VPFT), done by computerized spirometer. Hypertensive patients were put on oral enalapril, doses were titrated and maintained on 2.5 to 10 mg once daily. Twenty percent of the total hypertensive patients reported mild to moderate dry cough and was more frequently observed among females (27%). Significant decline was observed in MEF 50% and MEF 25% of vital capacity values (p 0.0204 and 0.0001) after 10 days of enalapril therapy. These two VPFT parameters showed significantly higher decline among patients who developed cough as compared to patients who did not develop cough. Decline in VPFT parameters were directly related to doses of enalapril. PMID- 11273317 TI - The changed clinical spectrum of malaria due to drug resistance. AB - The clinical spectrum of 14 cases of Plasmodium falciparum malaria (PF) who received empirical treatment and suffered from initial prolonged mild illness culminating into severe complicated malaria are presented. The empirical treatment (ET) consisted of adequate doses of chloroquine in 9, chloroquine with pyrimethamine-sulphadoxine combination in 3 and pyrimethamine-sulphadoxine alone in 2 cases. Moderate fever and weakness persisted for 7 to 28 days leading to anaemia and progressive hepatosplenomegaly in all patients. Other clinical features noticed included jaundice in 5, sudden shock with pulmonary oedema in 4, cerebral malaria and renal failure in 3 each and multiorgan in 4 cases. Subsequent investigations revealed PF rings in 9 cases, mixed PF and vivax infection in 3 and PF gametocytaemia only in 2 patients. Seven patients received quinine, 4 quinine with doxycycline and 3 were given quinine followed by injection artemether. Exchange transfusion was carried out in two cases. Four patients died. The empirical treatment with first line antimalarials alters the clinical profile of resistant PF, makes it milder temporarily, delays in confirming the diagnosis and leads to high mortality. There is urgent need for more diligent early workup for these patients who linger on with moderate pyrexia, progressive hepatosplenomegaly, anaemia and jaundice after ET till better diagnostic methods are available to avoid the prolonged illness and high mortality. PMID- 11273318 TI - Human immunodeficiency virus infection in a tertiary care hospital--clinical and microbiological profile. AB - Infection with the Human Immunodeficiency Virus (HIV) has an increasing, direct and significant impact on the hospital, especially the emergency services. Out of a total of 39,876 patients screened, 1061 patients were reactive for anti-HIV antibodies. The incidence of the infection showed a remarkable progression from 0.89% in 1992-1993 to 5.6% in 1997, among the seropositive patients. The number of patients with signs and symptoms related to HIV infection has also shown a 2 fold rise (from 42% in 1992-1993 to 87% in 1997). The prevalence of HIV 1 in the seropositive patients is 93%, HIV 2 alone is 2.3% and 3.1% had a mixed infection with both HIV 1 and 2. Secondary infection with Mycobacteria ranked high (25%) among the symptomatic patients, with pulmonary and the disseminated varieties being more common. PMID- 11273319 TI - Pictorial CME. AIDS-related cryptococcus infection presenting with molluscum contagiosum-like skin lesions. PMID- 11273320 TI - Role of pinaverium bromide in south Indian patients with irritable bowel syndrome. AB - The effect of pinaverium bromide in controlling gastrointestinal symptoms in 61 patients with irritable bowel syndrome was studied, as an open trial. Individually, there was significant relief in abdominal discomfort/pain as well as in bowel symptoms in most of the patients. Abdominal pain was reduced in 49%, stool consistency improved in 74%, straining and urgency decreased in 71% and mucus decreased in 64%. Tolerance to the drug administered was good and side effects reported were few. PMID- 11273321 TI - Telephone mouthpiece as a possible source of hospital infection. AB - A bacterial culture from telephone mouthpiece showed that 47 percent of the instruments carried pathogenic bacteria. Wiping with a disinfectant swab reduced the number of contaminated telephones. But for complete elimination of bacterial contamination changes in design of mouthpiece holes or type of instrument or using a polythene plastic cover over mouthpiece is recommended. PMID- 11273322 TI - Aspirin: aspiring for a century (1899 to 1999) and the willow bark is still batting! PMID- 11273323 TI - Life threatening status epilepticus: a review with case studies. PMID- 11273324 TI - Sleep apnea/hypopnea syndrome. PMID- 11273325 TI - Scorpion sting. AB - Acute life threatening systemic involvement (cardiovascular and central nervous system) occur due to scorpion poisoning, often reported from rural part of world comprises the majority of developing countries. The most serious symptoms are hypertension, impaired left ventricular systolic function, hypotension and pulmonary oedema. Scorpion antivenin neutralizes circulatory and tissue venin. It has no action on effective effectors (Receptors). Prazosin possesses pharmacological properties that render it most suitable in antagonizing the toxicological effects of scorpion venom. PMID- 11273326 TI - Mini-hemithorax without lower rib crowding--a clue to diagnosis of unilateral hypoplasia of the lung. PMID- 11273328 TI - Jelly fish sting poisoning. PMID- 11273327 TI - Myelofibrosis with myeloid metaplasia due to tuberculosis. PMID- 11273329 TI - Pulmonary mucormycosis. PMID- 11273330 TI - Primary tuberculosis of the esophagus. PMID- 11273331 TI - Periodic hypersomnia. PMID- 11273332 TI - Spastic paraparesis in alcoholic cirrhosis with portal systemic encephalopathy. PMID- 11273333 TI - Inadvertent use of ciprofloxacin as a single agent in undiagnosed tuberculosis. PMID- 11273334 TI - Nephrotic syndrome and hypokalemic paralysis. PMID- 11273335 TI - Nuclear scan and sonogram in renal tuberculosis. PMID- 11273336 TI - Risk factors in NIDDM. PMID- 11273337 TI - Hepatomegaly due to primary amyloidosis. PMID- 11273338 TI - Hypovitaminosis D in patients attending a rheumatology clinic at a tertiary referral centre. PMID- 11273339 TI - Ankle brachial index in peripheral vascular disease in diabetes mellitus. PMID- 11273340 TI - Choice of ACE inhibitor in the therapy of hypertension. PMID- 11273341 TI - Pulsed Doppler echocardiographic study of left ventricular diastolic function in patients with idiopathic dilated cardiomyopathy. AB - Dilated cardiomyopathy is basically regarded as a disease of left ventricular systolic dysfunction. There are only a few studies evaluating diastolic function in patients with dilated cardiomyopathy. To assess the LV diastolic function, 25 patients with idiopathic dilated cardiomyopathy and 20 age and sex matched normal subjects were studied with transmitral spectral tracings derived from pulsed Doppler echocardiography. All cardiomyopathy patients were in New York Heart Association class III to IV with dilated left ventricles and reduced systolic function (mean ejection fraction of 36.6 +/- 6.7 Vs 65 +/- 6 in normal subjects, p < 0.001). Patients with cardiomyopathy demonstrated an increased ratio of early to late diastolic velocity (E/A) (1.89 +/- 0.59 Vs 1.50 +/- 0.27 m/sec, p < 0.05), short deceleration time (E-E/2) (57.05 +/- 13.36 Vs 70.20 +/- 16.56 msec, p < 0.01) and short isovolumic relaxation time (IVRT) (53.5 +/- 22.7 Vs 72 +/- 12 msec, p < 0.05) as compared to normal subjects. The early filling fraction (EFF) was higher (0.71 +/- 0.11 Vs 0.66 +/- 0.06, p < 0.05) and atrial filling fraction (AFF) was lower (0.28 +/- 0.11 Vs 0.33 +/- 0.06, p < 0.05) in cardiomyopathy patients than in normal subjects. Our observations in a select group of dilated cardiomyopathy patients with advanced disease demonstrate a restrictive pattern on pulsed Doppler echocardiography. PMID- 11273342 TI - Simultaneous radiochemotherapy in the treatment of inoperable, locally advanced head and neck cancers. AB - The results of radiation therapy alone in locally advanced head and neck cancers are dismal with 5 year locoregional control rates not exceeding 15%. The addition of concomitant chemotherapy with cisplatin and more recently carboplatin has shown promising results. Twenty patients of inoperable stage III and IV oral or oropharyngeal cancers were treated with concomitant chemoradiation with carboplatin 300 mg/m2 i.v. on days 1, 21 and 42 of radiation therapy. Twelve (60%) patients had a complete remission. Thirteen patients were alive at a median follow up of 11 months. The treatment was well tolerated with only 2 patients requiring treatment interruptions for mucositis. Longer follow up would reveal any improvement in overall survival. The relative ease with which carboplatin/RT was administered suggests that other agents might be added as well. PMID- 11273343 TI - Elective coronary artery stenting--immediate and follow up results. AB - Elective coronary artery stenting was performed in 242 consecutive patients in our centre for complex lesions (Type B, C), proximal lesions, restenotic lesions, total occlusion and venous grafts. The procedural success rate was 94.21%. Three patients (1.23%) required emergency coronary artery bypass surgery. Acute and sub acute thrombosis rate was 1.26% and 4.13%, respectively. There was one in hospital death (0.41%). 164 patients were followed up clinically for a mean period of 11 +/- 6 months (range 1 month to 30 months). Angiographic follow up was done in 68 patients with a restenosis rate of 16.17%. PMID- 11273344 TI - Early development of transient hypothyroidism after I131 therapy for thyrotoxicosis. AB - 295 patients of Graves' disease were studied for early development of transient hypothyroidism (TH) and its prognostic value following I131 therapy. 278 patients received I131 < 10 mci (6.4 +/- 1.7 mci) and 17, a dose of > 10 mci (12.6 +/- 2.6). TH was diagnosed on the basis of low T4 regardless of TSH within the first year after I131 therapy followed by normal T4 and TSH. 32 patients developed TH following administration of < 10 mci I131 and it was symptomatic in 10 patients. No instance of TH after high dose of I131 was noted. I131 uptake > 60% at 2 hours before treatment was a risk factor for developing TH (odds ratio 2.6, 95% confidence interval 0.8-9.6). At diagnosis of TH basal TSH was high in 53%, normal in 32%, or low in 15%; Hypothyroidism recognized during the first six months with basal TSH of 50 microU/ml or higher ruled out TH. Development of TH and its hormonal profile did not influence long term thyroid functions. As no prognostic factors predicted TH before I131 therapy or at the time of diagnosis, re-evaluation of thyroid functions later is essential to avoid unnecessary chronic replacement therapy, if hypothyroidism has been diagnosed within a few months of I131 treatment. PMID- 11273345 TI - Experience in adult population in dengue outbreak in Delhi. AB - A dengue outbreak has recently hit the Indian capital. We studied the clinical profile of adult patients. Five hundred and sixty patients of dengue infection were admitted in a specially created ward according to the criteria laid down by WHO. Haematemesis (28.28%), epistaxis (26.78%) and malena (14.28%) were some of the common presentations. Similarly lymphadenopathy, especially cervical (30.89%), palatal rashes (26.96%) and hepatomegaly (23.75%) were the most commonly encountered findings on physical examination. Most of the cases were of dengue fever with haemorrhage and only 2.5% cases were classified under dengue haemorrhagic fever or dengue shock syndrome. The average hospital stay was 3.4 days but only 9.8 hours in the eleven patients who died, suggesting their late arrival in preterminal situation giving little time for resuscitation. Thrombocytopenia was not a feature and only 12.85% patients had platelet count less than 70,000/cmm. Most of the patients who were admitted with thrombocytopenia, showed normalization in their platelet counts in next few days. Serological examination demonstrated evidence of recent dengue infection in 41.17% patients. Few patients required blood or platelet concentrate transfusion. Eleven patients died, three due to DIC, one of intracranial haemorrhage and seven due to massive gastric haemorrhage. Rest of the patients recovered completely. Thus we can conclude that recent outbreak in Delhi was of dengue fever with haemorrhage and mortality was very low in patients who came early to the hospital. PMID- 11273346 TI - Psychosocial aspects of seropositive HIV patients. AB - Acquired immunodeficiency syndrome, has become a serious global health problem with an enormous biomedical impact and diverse psychological and clinical manifestation. A study was conducted wherein seropositive HIV patients admitted in a medical ward were assessed for demographic profile, presenting physical illness, mode of contacting the infection, psychiatric morbidity and associated psychosocial factors. The results revealed male preponderance and heterosexual unprotected exposure as the common mode of contacting the illness. It was observed that majority of the patients presented with tuberculosis. Psychiatric assessment revealed a high rate of depressive and anxiety syndromes. Thus it was concluded that the HIV-infected population has a higher prevalence of psychiatric disorders as compared to the general population. The reason for this includes psychological, biological and social factors. PMID- 11273347 TI - Pictorial CME. Histoplasmosis in HIV seropositivity. PMID- 11273348 TI - An open clinical trial of benazepril--a new ACE inhibitor in mild-moderate hypertension. AB - Benazepril hydrochloride, a new non-sulfhydryl ACE inhibitor (ACEI) was studied in a titrated dose of 10 mg-20 mg once a day for 6 weeks in 42 mild to moderate adult hypertensive patients with sitting diastolic blood pressure (SDBP) 95-114 mm Hg. The pre-drug SDBP(mean +/- SE) of 102.5 +/- 0.8 mm Hg showed a significant reduction to 87.5 +/- 0.93 mm Hg at the end of treatment. BP was controlled (SDBP < or = 90 mm Hg) in 34 (81%) patients and a drop of at least 10 mm Hg from the pre-treatment SDBP value was noted in 34 (81%) patients. Common adverse reaction was cough in 8(19%) patients. Clinically significant changes in laboratory evaluations were not seen in any patient. Study showed that benazepril in a dose range of 10 to 20 mg per day is an effective agent for treatment of mild to moderate hypertension. PMID- 11273349 TI - Myasthenia gravis. PMID- 11273350 TI - Hormone replacement therapy in postmenopausal women for cardioprotection. PMID- 11273351 TI - Coenzyme Q in cardiovascular disease. AB - Coenzyme Q10 or ubiquinone normally present in many plant and animal cells is an antioxidant. Coenzyme Q10 deficiency has been observed in patients with congestive heart failure, angina pectoris, coronary artery disease, cardiomyopathy, hypertension, mitral valve prolapse and after coronary revascularization. Coenzyme Q10 is involved in the synthesis of ATP and hence is useful in preventing cellular damage during ischaemia-reperfusion injury. The clinical benefits are mainly due to its ability to improve energy production, antioxidant activity, and membrane stabilizing properties. Several studies showed that coenzyme Q could be useful in patients with congestive heart failure, angina pectoris, cardiomyopathy, coronary artery disease and in the preservation of myocardium. Coenzyme Q10 is normally present in the low density lipoprotein cholesterol fraction and inhibits its oxidation. It can also regenerate vitamin E. Coenzyme Q10 is known for producing minor gastrointestinal discomfort and elevation in SGOT and LDH when used. PMID- 11273352 TI - Hypernephroma presenting as right atrial mass. PMID- 11273353 TI - Colonic tuberculosis mimicking ulcerative colitis. PMID- 11273354 TI - Pure red cell aplasia with thymoma. PMID- 11273355 TI - Pure red cell aplasia associated with thymoma. PMID- 11273357 TI - Atypical presentation of herpes simplex encephalitis in acquired immune deficiency syndrome. PMID- 11273356 TI - Pure red cell aplasia with thymoma in a middle aged man. PMID- 11273358 TI - Clinico-radiological profile of Kluver-Bucy syndrome. PMID- 11273359 TI - Leptomeningeal and brain metastasis in a young patient of signet ring cell carcinoma of stomach. PMID- 11273360 TI - Hypopituitarism following snake bite. PMID- 11273361 TI - Consumer product information: how rational the information is? PMID- 11273362 TI - Clinical resistance to ciprofloxacin in Salmonella typhi. PMID- 11273363 TI - Is it Yellowman syndrome? PMID- 11273364 TI - Efficacy of invasive vascular closure devices for femoral hemostasis. PMID- 11273365 TI - What happens to patients who teach? AB - BACKGROUND: Over the last 35 years, the use of lay people as teaching patients has evolved from novel to routine. PURPOSE: Although many studies have examined the effectiveness, reliability, and validity of using trained patients to evaluate medical students and residents, little is known about the effect on patients of being teachers. This study addresses that question. METHODS: Patients in-training with actual musculoskeletal conditions completed 3 questionnaires: the SF-36 to evaluate perceived health status, the Chubon Life Situation Survey to examine perceived quality of life, and 3 visual analogue scales to assess the relationship with their personal physicians. One year later, the same questionnaires were readministered to patients who were actively teaching and those who had dropped out of training. RESULTS: In comparison to those who dropped out of training, patients who were actively teaching demonstrated a trend toward lower health status but higher quality of life; they also reported more satisfying relationships with their personal physicians. CONCLUSIONS: This study provides evidence that patient-teachers appear to benefit from teaching despite lowered health status. Similar to other studies, instructors' personal relationships with their respective physicians were positively influenced. PMID- 11273366 TI - Impacting faculty teaching and student performance: nine years' experience with the Objective Structured Clinical Examination. AB - BACKGROUND: The impetus for administering the 2nd-year Objective Structured Clinical Examination (OSCE) came from the great variability in student performance observed by 3rd-year clerkship directors. PURPOSE: To document the effects of the OSCE on faculty teaching, student performance, and the curriculum over 9 years of administration of the examinations to more than 1,000 second-year medical students. METHOD: A 20-station OSCE was administered to all medical students at the end of their 2nd year. Using predetermined criteria, clinical faculty served as evaluators in each station. A mix of 1st-, 3rd-, and 4th-year medical students were recruited to serve as simulated patients. Faculty evaluators and examinees completed a questionnaire evaluating their experience with the OSCE. Students received a report card of their performance. Small-group leaders of the Introduction to Clinical Medicine course received feedback on their group's performance on each station compared to the class mean. Summative data on class performance was reported to the curriculum committee. The academic status committee received data on students who performed unsatisfactorily. RESULTS: Faculty and examinee ratings of the OSCE experience were very positive. Over the 9-year period, student performance improved showing less variability and significantly fewer failed stations. CONCLUSION: The OSCE has proven to be a technically feasible, authentic evaluation method yielding valuable information for decisions regarding student performance, faculty teaching, and curriculum planning. PMID- 11273367 TI - Standardized patient assessment of ambulatory clerks: effect of timing and order of the clerkship. AB - BACKGROUND: A standardized patient examination may assess unique learning in an ambulatory clerkship but, as with written tests, may be affected by student maturation. PURPOSE: To explore the effect of timing and order of a medicine ambulatory clerkship on student performance of a standardized patient examination (SPE). METHODS: All students rotating through an ambulatory medicine clerkship in 1 academic year completed an SPE designed to reflect specific learning objectives of the clerkship as well as nonclerkship case content. Students were grouped according to prior inpatient clerkship experience. RESULTS: When compared to students with only ambulatory experience, students with both inpatient and ambulatory experiences in internal medicine did not perform better on the ambulatory cases of the SPE but did perform better on nonclerkship cases. Performance on the SPE was not affected by month of training. At completion of the inpatient clerkship, students with prior ambulatory experience did not perform better than students with inpatient-only experience on the National Board of Medical Examiners Medicine Subject Examination. CONCLUSIONS: The SPE is an appropriate assessment tool for the ambulatory clerkship when case content is linked to learning objectives of the clerkship. Unlike other knowledge-based assessments, the SPE is not affected by student maturation. PMID- 11273368 TI - The Script Concordance test: a tool to assess the reflective clinician. AB - BACKGROUND: The Script Concordance (SC) test is a new assessment tool. It is designed to probe whether knowledge of examinees is efficiently organized for clinical actions. That kind of organization of knowledge is named a script. The SC test places examinees in written, but authentic, clinical situations in which they must interpret data to make decisions. PURPOSE: The SC test is designed to measure the degree of concordance that exists between examinees' scripts and scripts of a panel of experts. The objective of this article is to provide interested educators with the practical "how to" information needed to build and use an SC test. METHODS: The theoretical background of the SC test is described. The principles of construction of an SC test are presented, including the writing of clinical cases, the choice of item format, the validation of the test, and the elaboration of the scoring system. RESULTS: A series of studies have shown that the SC test has interesting psychometric properties, in terms of reliability, face validity, and construct validity. Results from these studies are succinctly presented and commented. CONCLUSION: The SC test is a simple and direct approach to testing organization and use of knowledge. It has the strong advantage for a testing method of being relatively easy to construct and use and to be machine scorable. It can be either paper- or computer-based and can be used in undergraduate, postgraduate, or continuing medical education. PMID- 11273369 TI - Assessing the measurement properties of a clinical reasoning exercise. AB - BACKGROUND: A challenge for Problem-Based Learning (PBL) schools is to introduce reliable, valid, and cost-effective testing methods into the curriculum in such a way as to maximize the potential benefits of PBL while avoiding problems associated with assessment techniques like multiple-choice question, or MCQ, tests. PURPOSE: We document the continued development of an exam that was designed to satisfy the demands of both PBL and the scientific principles of measurement. METHODS: A total of 102 medical students wrote a clinical reasoning exercise (CRE) as a requirement for two consecutive units of instruction. Each CRE consisted of a series of 18 short clinical problems designed to assess a student's knowledge of the mechanism of diseases that were covered in three subunits located within each unit. Responses were scored by a student's tutor and a 2nd crossover tutor. RESULTS: Generalizability coefficients for raters, subunits, and individual problems were low, but the reliability of the overall test scores and the reliability of the scores across 2 units of instruction were high. Subsequent analyses found that the crossover tutor's ratings were lower than the ratings provided by one's own tutor, and the CRE correlated with the biology component of a progress test. CONCLUSION: The magnitude of the generalizability coefficients demonstrates that the CRE is capable of detecting differences in reasoning across knowledge domains and is therefore a useful evaluation tool. PMID- 11273370 TI - Validation and use of an instrument to measure the learning environment as perceived by medical students. AB - BACKGROUND: Aiming to inform curriculum changes in medical school, we developed, administered, and validated a 31-question survey to measure the learning environment as perceived by medical students. DESCRIPTION: We administered the survey annually in 4 medical school classes in a Southeastern medical school from May 1994 through May 1997 (N = 619). EVALUATION: The survey responses reflected 3 dimensions of the medical school learning environment: the teacher-learner relationship (T-L R), the physician-patient relationship (Phys-Pt R), and self efficacy. We found that the 3 dimensions are equally valid and reliable for all students, but that the mean values on all 3 dimensions differed by year in school and number of survey responses. CONCLUSIONS: As students progress through school, they perceive deteriorating T-L Rs, feel diminishing self-efficacy, and accord less value to the Phys-Pt R. Based on these results, we developed training programs for faculty members to promote teaching attributes known to facilitate relationship formation between teacher and learner, and learner-centered and self directed learning. PMID- 11273371 TI - The program for professional values and ethics in medical education. AB - BACKGROUND: Medical educators are very interested in the teaching and evaluation of professional attitudes and behaviors among medical students, residents, and faculty. At Tulane University School of Medicine, we created the Program for Professional Values and Ethics in Medical Education (PPVEME) to return the focus of our curriculum to the physician-patient-community relationship and to the nurturing of professionalism. DESCRIPTION: PPVEME brings together students, residents, and faculty into learning teams that create longitudinal curricula about five themes: integrity, communication, teamwork, leadership, and service. The emphasis is on learner-driven self- and group-reflection about shared experiences, thus modeling essential professional attributes. EVALUATION: The program is evaluated using surveys, student and faculty focus groups, and portfolios developed by student volunteers on each team. The first program event, a retreat for entering medical students, was highly successful. CONCLUSIONS: PPVEME is an attempt to construct a medical school learning environment around professionalism. Evaluation over time will tell how successfully that has been accomplished. PMID- 11273372 TI - Proposed use of two-part interactive modeling as a means to increase functional skills in children with a variety of disabilities. AB - PURPOSE: Many behavior modification and intervention programs are based on operant procedures developed for animal subjects, but few use modeling procedures in which one student observes interactions between two proficient trainers. We show how such procedures, which successfully trained Grey parrots (Psittacus erithacus) to produce and comprehend elements of human language, can be adapted for use with children with three types of disabilities: (a) autism with limited social and language skills, (b) developmental delay with physical handicaps and lack of language skills, and (c) hyperactivity with impaired cognitive and social skills. SUMMARY: Children were evaluated before entering the program and outcomes were recorded to determine improvement levels. No child reached totally normative (physical age-appropriate) levels, but all significantly improved their social and communication skills and use of contextually appropriate behavior. CONCLUSIONS: A two-trainer modeling system can be a valuable intervention tool for children whose disabilities involve social and communicative skills. PMID- 11273373 TI - Double duty: students' perceptions of Tulane's MD-MPH dual degree program. AB - PURPOSE: Although MD-MPH programs exemplify the initiative for collaboration between schools of medicine and public health and address the expanding requirements for effective medical practice, information on such programs is scant. Perspectives and motivations of students enrolled in a 4-year MD-MPH program are explored to benefit existing and new programs as well as to inspire future research. SUMMARY: A questionnaire, based on previously identified themes, was mailed to all 110 students enrolled in the MD-MPH program at Tulane University. The typical respondent felt prepared for the program, expected to practice medicine full time, and expected to practice internationally up to 3 months annually. Perceived enhancements and barriers to dual degrees are addressed. CONCLUSIONS: Increased awareness of MD-MPH programs at the undergraduate level might be beneficial. Respondents valued the broader perspectives on the doctor-patient-society triad and additional career opportunities gained through their combined studies. Findings of this study can facilitate program planning and improvement elsewhere. PMID- 11273374 TI - Student perspectives on primary care preceptorships: enhancing the medical student preceptorship learning environment. AB - BACKGROUND: Medical students participate in a longitudinal (3-year) primary care preceptorship to assist them in developing skills in interviewing and examining patients in an ambulatory care setting. PURPOSE: To identify from a student's perspective important context and process issues in a longitudinal preceptorship. METHODS: The investigators used an "editing" style of analysis to identify significant themes across 24 medical student focus groups held between October 1995 and December 1997. RESULTS: Significant themes emerged from the data analysis that describe important features of what makes the preceptorship work for students. The main themes are active teaching, active learning, a trusting relationship, sufficient time, and a shared understanding of preceptorship objectives. The potential benefits to students in an enhanced learning environment are comfort, confidence, responsibility, skills, knowledge, reinforcement, learning opportunities, teaching opportunities, and models for practice. CONCLUSIONS: We offer recommendations for enhancing longitudinal preceptorships for preceptors, students, and leaders in medical education. PMID- 11273375 TI - Evaluating a clerkship curriculum: description and results. AB - BACKGROUND: A comprehensive and up-to-date curriculum requires periodic formal review to ensure it continues to meet learners' needs. PURPOSE: This study describes a model for evaluating a surgery clerkship curriculum designed to determine the appropriateness of its learning objectives to the general professional education of a physician. METHODS: A survey was mailed to graduates who pursued generalist residencies. Respondents estimated the number of patients encountered annually with specified presenting complaints or disease entities and the percentage of time these were referred to surgeons. For 23 technical procedures, respondents estimated the frequency done annually and whether remaining proficient in the skill was considered important. RESULTS: The majority of graduates reported the need to remain proficient in 19 technical procedures. Numerous patient problems were identified as requiring careful instruction so that learners know when and when not to refer for surgical intervention. CONCLUSIONS: The clerkship was modified to include skills and topics not previously included or appropriately emphasized. PMID- 11273376 TI - Measuring critical thinking in problem-based learning discourse. AB - BACKGROUND: Critical thinking (CT) is a composite of skills linked to problem based learning (PBL). PURPOSES: This study has 3 purposes: (a) to determine if PBL discourse could be coded for CT, (b) to demonstrate reliable coding, and (c) to determine whether a CT ratio would provide a valid measure to compare 2 PBL groups. METHODS: Using prior research, we refined the code for a content analysis of PBL transcripts. Raters coded 6 hr of transcripts and computed CT ratios for each of the 5 CT stages. Average interrater agreement was 85.5%. CT ratios appeared to differ between 2 PBL groups delivered in 2 modalities. RESULTS: PBL discourse could be coded following a CT framework. Independent raters reliably applied the code, and the resulting CT ratios detected tenable differences. CONCLUSIONS: This approach could provide useful information about the effect of case modality. PMID- 11273378 TI - A suggested outline for writing curriculum development journal articles: the IDCRD format. AB - BACKGROUND: During the past decade, medical school and residency faculty have been active in developing and revising curricula for medical education programs. Many of these curriculum development efforts ultimately are published in peer reviewed professional journals as articles or abstracts. Unlike research publications, no uniform format currently exists for reporting curriculum development efforts in the peer-reviewed literature. SUMMARY: A suggested format for organizing curriculum development manuscripts consists of the introduction, development, curriculum, results, and discussion (IDCRD). Detailed descriptions of each section are discussed herein. CONCLUSIONS: The IDCRD manuscript outline is intended to provide useful guidance to medical educators in publishing their curriculum development efforts. Journal editors are encouraged to recognize the importance of providing uniform descriptions of curricula so that readers can benefit from the experience of others and replicate successful curriculum efforts. PMID- 11273377 TI - Interim evaluation of the Rural Health Scholars Program. AB - BACKGROUND: The imperative to address physician maldistribution has been directed in part at medical schools. DESCRIPTION: The Rural Health Scholars Program (RHSP) is an enrichment initiative that has been implemented at 2 medical schools to increase the number of students likely to practice primary care in rural, underserved areas. It is a longitudinal program that includes a skill-building workshop; a 5-week summer preceptorship with community-based preceptors in rural, underserved areas; and opportunities to return to preceptorship sites during 3rd- and 4th-year rotations. Students also attend community-based and teleconference seminars and workshops, as well as informal social gatherings. EVALUATION: A static-group comparison design was used to compare program participants with nonparticipants regarding residency program types and locations. CONCLUSIONS: The RHSP is meeting some interim objectives conducive to its long-term goal of developing physicians who will practice primary care medicine in rural, underserved areas of North Carolina. PMID- 11273379 TI - The evaluation of a workshop to promote interactive lecturing. AB - BACKGROUND: The lecture is the most widely used method of teaching in medical education. Although effective lecturing has been described in the literature, many question whether problem-solving skills or attitudes can be transmitted using the traditional lecture. Introducing interactive techniques can promote learner participation and, as a result, can lead to a higher level of learning. This article assesses the effectiveness of interactive learning. DESCRIPTION: A 4 hr workshop has been offered for 4 consecutive years to faculty members in the Faculty of Medicine at McGill University to allow participants to explore interactive techniques and incorporate them into their lectures. For this study, an evaluation was conducted of the workshop given in Spring 1996. The experimental group consisted of the first 60 faculty members to register for the workshop, and the comparison group comprised the 40 individuals on the waiting list. Three instruments were used in the evaluation. An immediate postworkshop questionnaire was completed by the participants. Six months after the session, a 6-month postworkshop questionnaire was completed by the experimental and comparison groups that explored the use of interactive lecturing techniques since the workshop. In addition, 23 individuals from the experimental group and 14 from the comparison group were videotaped 6 months after the session and were scored on a videotape observational grid by an independent rater. EVALUATION: Overall, the workshop was deemed very useful by the majority of the participants. On the 6 month postworkshop questionnaire, the only difference found in the demographic data between the 2 groups was in the number of years of teaching experience. The experimental group had given more interactive lectures over the past 6 months and had used more audience responses, certain types of questions, audience surveys, live interviews, verbal and written cases, and study guides. From the videotape observational data, the experimental group scored higher in questioning and engaging the audience, and in using nonverbal gestures. As well, this group received higher ratings for their interactivity and for the students' responsiveness. CONCLUSIONS: Interactive lectures can increase student participation and involvement in the large class lecture. This 4-hr workshop, designed to promote the use of interactive lecturing techniques, can be considered successful based on self-reports from participants as well as from observational data. PMID- 11273380 TI - Ambulatory morning report: an underutilized educational modality. AB - BACKGROUND: Many medicine residency training programs include a lecture-based preclinic conference series as part of the ambulatory educational curriculum when more effective teaching formats might be available. Our institution has replaced this lecture-based teaching format with an ambulatory morning report modeled after the inpatient paradigm. This study compares the efficacy of these 2 teaching models and defines the desired characteristics of this new teaching strategy. DESCRIPTION: We first conducted a background study by obtaining permission to use test questions from the Medical Knowledge Self-Assessment Program to develop pre- and postambulatory rotation tests to validate our ambulatory curriculum. Forty-three of 44 interns completed both pre- and postambulatory block testing. The mean score on these tests improved from 67% to 81%. Although this overall improvement achieved statistical significance, test question subgroup analysis clearly indicated that the improved knowledge in test items relating to our preclinic conference topics contributed negligibly to the overall statistical improvement. As such, we ultimately replaced our preclinic conference with a morning report format. In this study, 82 residents were enrolled in a 2-year prospective observational study. This study group completed an ambulatory curriculum in which the 1st year was completely lecture based, and the 2nd year included the morning report format. We were thus able to survey residents' opinions regarding the effectiveness of the 2 very different teaching formats used in consecutive years. EVALUATION: The survey results from those residents exposed to both teaching formats over 2 years revealed a high degree of satisfaction with the ambulatory morning report format. When comparing the long term educational value, 94% of the residents found the morning report format more effective than the lecture-based preclinic conference. In addition, many desirable characteristics of the morning report also were defined. CONCLUSIONS: Lecture-based preclinic conferences might not be the most effective way of conveying information over the long term. Residents seem to prefer the more interactive morning report format. When organizing such a teaching format, attention should be directed toward the characteristics that were felt to be desirable by those we are trying to teach. PMID- 11273381 TI - Toward setting a research agenda for systematic reviews of evidence of the effects of medical education. AB - PURPOSE: To provide an update on, and a preliminary research agenda for, best evidence medical education (BEME). SUMMARY: Efforts related to evidence-based medical education are summarized briefly, including BEME, the newly formed Campbell Collaboration, and the Cochrane Collaboration's Effective Practice and Organization of Care review group. A list of topics and priorities for which evidence of effectiveness in medical education should be systematically reviewed is provided based on the results of a session at the July 2000 annual meeting of the Society of Directors of Research in Medical Education. The highest ranked topics clustered around four major conceptual areas: (a) curricular design, (b) learning and instructional methods, (c) testing and assessment, and (d) outcomes. CONCLUSIONS: BEME is gaining momentum with growing numbers of people becoming involved as well as an increased number of pertinent workshops, publications, and Web sites. The work of creating pertinent systematic reviews of the medical education literature is at hand. PMID- 11273382 TI - Residency program director evaluations do not correlate with performance on a required 4th-year objective structured clinical examination. AB - BACKGROUND: Assessment of resident performance is a complex task. PURPOSE: To correlate performance on a 4th-year objective structured clinical examination (OSCE) with residency program director assessment, class rank, and U.S. Medical Licensing Examination (USMLE) scores. METHODS: We surveyed program directors about the performance of 50 graduates from our medical school chosen to represent the highest (OSCEHI) and lowest (OSCELO) 25 performers on our required 4th-year OSCE. Program directors were unaware of the OSCE scores of the graduates. RESULTS: OSCE scores did not correlate with Likert scores for any survey parameter studied (r < .23, p > .13 for all comparisons). Similarly, program director evaluations did not correlate with class rank or USMLE scores (r < .26, p > .09 for all comparisons). CONCLUSIONS: We concluded that program director evaluations of resident performance do not appear to correlate with objective tests of either clinical skills or knowledge taken during medical school. These findings suggest that more structured and objective evaluative tools might improve postgraduate training program assessment of trainees. PMID- 11273383 TI - Recognition and source memory as multivariate decision processes. AB - Recognition memory, source memory, and exclusion performance are three important domains of study in memory, each with its own findings, it specific theoretical developments, and its separate research literature. It is proposed here that results from all three domains can be treated with a single analytic model. This article shows how to generate a comprehensive memory representation based on multidimensional signal detection theory and how to make predictions for each of these paradigms using decision axes drawn through the space. The detection model is simpler than the comparable multinomial model, it is more easily generalizable, and it does not make threshold assumptions. An experiment using the same memory set for all three tasks demonstrates the analysis and tests the model. The results show that some seemingly complex relations between the paradigms derive from an underlying simplicity of structure. PMID- 11273385 TI - Category effects on estimates of stimuli: perception or reconstruction? AB - The present study examined a common category effect that has been reported in the literature: the tendency for estimates of individual stimuli to be biased toward the central value of the presented set of stimuli. Both encoding and reconstruction accounts of this central-tendency effect are considered. Plain vertical lines and vertical lines embedded in the Muller-Lyer illusion were estimated while still in view or from memory. Although bias due to the Muller Lyer illusion remained constant across the two conditions, bias due to the context set (category) occurred only when stimuli were estimated from memory. The results suggest that the category bias occurs at a later stage of processing than the Muller-Lyer effect and offer support for a reconstruction account of category effects on stimulus estimation. PMID- 11273384 TI - What the eyes say about speaking. AB - To study the time course of sentence formulation, we monitored the eye movements of speakers as they described simple events. The similarity between speakers' initial eye movements and those of observers performing a nonverbal event comprehension task suggested that response-relevant information was rapidly extracted from scenes, allowing speakers to select grammatical subjects based on comprehended events rather than salience. When speaking extemporaneously, speakers began fixating pictured elements less than a second before naming them within their descriptions, a finding consistent with incremental lexical encoding. Eye movements anticipated the order of mention despite changes in picture orientation, in who-did-what-to-whom, and in sentence structure. The results support Wundt's theory of sentence production. PMID- 11273386 TI - The attractiveness of nonface averages: implications for an evolutionary explanation of the attractiveness of average faces. AB - Researchers have argued that humans' attraction to average faces reflects an evolved psychological mechanism to identify high-quality mates. If this direct selection account is correct, there is no reason to expect a similar averageness bias for stimuli that are irrelevant to reproductive fitness. The current study, however, found a strong relationship between averageness and attractiveness for dogs, wristwatches, and birds. The most parsimonious explanation is that humans have a general attraction to prototypical exemplars, and that their attraction to average faces is a reflection of this more general attraction. We tested whether a general preference for familiar stimuli can account for the attractiveness of averageness. This account was not supported for dogs or birds, but could not be ruled out for watches. PMID- 11273387 TI - Specificity of learning: why infants fall over a veritable cliff. AB - Nine-month-old infants were tested at the precipice of safe and risky gaps in the surface of support. Their reaching and avoidance responses were compared in two postures, an experienced sitting posture and a less familiar crawling posture. The babies avoided reaching over risky gaps in the sitting posture but fell into risky gaps while attempting to reach in the crawling posture. This dissociation between developmental changes in posture suggests that (a) each postural milestone represents a different, modularly organized control system and (b) infants' adaptive avoidance responses are based on information about their postural stability relative to the gap size. Moreover, the results belie previous accounts suggesting that avoidance of a disparity in depth of the ground surface depends on general knowledge such as fear of heights, associations between depth information and falling, or knowledge that the body cannot be supported in empty space. PMID- 11273388 TI - Taxonic structure of infant reactivity: evidence from a taxometric perspective. AB - Previously, we proposed a theoretical framework that classified infants into qualitative categories of reactivity, rather than on a continuous dimension. The present research used an objective statistical procedure (maximum covariance analysis, or MAXCOV) to determine if a qualitative latent structure, consistent with our theoretical conjectures, would be found to underlie quantitative indices of reactivity to stimuli in a sample of 599 four-month-old infants. Results of the MAXCOV analysis showed clear evidence of a latent discontinuity underlying the behavioral measures of infant reactivity. The base rate of the latent class (or taxon) was estimated at 10%. Infants within the putative high-reactivity taxon, compared with infants not in the taxon, were elevated on measures of behavioral inhibition at 4.5 years. These results provide objective empirical support for a central tenet in our theoretical model by supporting the taxonicity of infant reactivity. PMID- 11273389 TI - Prosocial foundations of children's academic achievement. AB - The present longitudinal research demonstrates robust contributions of early prosocial behavior to children's developmental trajectories in academic and social domains. Both prosocial and aggressive behaviors in early childhood were tested as predictors of academic achievement and peer relations in adolescence 5 years later. Prosocialness included cooperating, helping, sharing, and consoling, and the measure of antisocial aspects included proneness to verbal and physical aggression. Prosocialness had a strong positive impact on later academic achievement and social preferences, but early aggression had no significant effect on either outcome. The conceptual model accounted for 35% of variance in later academic achievement, and 37% of variance in social preferences. Additional analysis revealed that early academic achievement did not contribute to later academic achievement after controlling for effects of early prosocialness. Possible mediating processes by which prosocialness may affect academic achievement and other socially desirable developmental outcomes are proposed. PMID- 11273391 TI - Aversive racism and selection decisions: 1989 and 1999. AB - The present study investigated differences over a 10-year period in whites' self reported racial prejudice and their bias in selection decisions involving black and white candidates for employment. We examined the hypothesis, derived from the aversive-racism framework, that although overt expressions of prejudice may decline significantly across time, subtle manifestations of bias may persist. Consistent with this hypothesis, self-reported prejudice was lower in 1998-1999 than it was in 1988-1989, and at both time periods, white participants did not discriminate against black relative to white candidates when the candidates' qualifications were clearly strong or weak, but they did discriminate when the appropriate decision was more ambiguous. Theoretical and practical implications are considered. PMID- 11273390 TI - The cultural bounds of maternal accommodation: how Chinese and American mothers communicate with deaf and hearing children. AB - Children with special needs typically require family accommodation to those needs. We explore here the extent to which cultural forces shape the accommodations mothers make when communicating with young deaf children. Sixteen mother-child dyads (8 Chinese, 8 American) were videotaped at home. In each culture, 4 mothers interacted with their deaf children, and 4 interacted with their hearing children. None of the deaf children knew sign language, nor spoke at age level. We found that mothers adjusted their communicative behaviors to their deaf children, but in every case, those adjustments were calibrated to cultural norms. American mothers, for example, increased their use of gesture with deaf children but stopped far short of the Chinese range--despite the obvious potential benefits of gesturing to children who cannot hear. These findings provide the first cross-cultural demonstration that children are, first and foremost, inculcated into their cultures and, only within that framework, then treated as special cases. PMID- 11273392 TI - Sadness and susceptibility to judgmental bias: the case of anchoring. AB - In a wide range of empirical paradigms, sadness has been associated with more extensive and detail-oriented thinking than happiness, resulting in reductions in judgmental bias that arise from reliance on stereotypes and other simple decision heuristics. It was hypothesized that anchoring would constitute a significant exception to this general pattern. Recent research on anchoring indicates that an active thought process underlies the emergence of this bias. If sad people are likely to think more actively about the judgmental anchor than their neutral-mood counterparts, their subsequent judgments should be more likely to be assimilated toward this reference point. This prediction was confirmed in two experiments demonstrating that sad people are indeed more susceptible to anchoring bias than are people in a neutral mood. Moreover, this effect generalized over judgments in positive, neutral, and negative content domains. PMID- 11273393 TI - Searching for one versus two identical targets: when visual search has a memory. AB - A recent study has suggested that observers' visual explorations of the external world can proceed unimpaired when the visual environment precludes the operation of memory processes (as, for instance, when the display elements change locations every 100 ms). One theoretical limitation of this study was that distractors were the only elements that had the potential to be tagged during visual search. The present study sought to clarify the amnesic-search hypothesis by investigating whether memory processes can guide search in other contexts in which targets also have the potential to be tagged. Accordingly, the experimental conditions of the previous study were repeated using a different search task in which observers had to decide whether one target or two were present among a variable number of similar distractors. Under these search conditions, the present findings provided strong evidence that memory processes can guide visual search. PMID- 11273394 TI - In search of remembrance: evidence for memory in visual search. AB - Observers searched for a target among distractors while the display items traded places every 110 ms. Search was slower when the target was always relocated to a position previously occupied by a distractor than when the items remained in place, showing the importance of memory for locations in a visual search task. Experiment 2 repeated a previous study in which items could move to any location within the display, but used a larger range of set sizes than tested in the earlier study. A cost in search times to relocating items was found at the larger set sizes, most likely reflecting that the probability that the target would replace a distractor increased with the set size. The findings provide strong evidence for the role of memory for locations within trials in a visual search task. PMID- 11273395 TI - False memories in women with self-reported childhood sexual abuse: an empirical study. AB - Although controversy exists about the validity of memories of childhood abuse, little is known about memory function in individuals reporting childhood abuse. This study assessed memories for previously presented words, including the capacity for false memory of critical lures not actually present in the word list, in 63 subjects, including abused women with posttraumatic stress disorder (PTSD), abused women without PTSD, and men and women without abuse or PTSD. Abused women with PTSD had a higher frequency of false recognition memory of critical lures (95%) than abused women without PTSD (78%), nonabused women without PTSD (79%), or nonabused men without PTSD (86%). PTSD women also showed poorer memory for studied words and increased insertions of non-studied words other than critical lures. These findings are consistent with a broad range of memory alterations in abused women with PTSD. PMID- 11273396 TI - Neighborhood deprivation affects children's mental health: environmental risks identified in a genetic design. AB - The possibility that neighborhood conditions affect children's development has captured much attention because of its implications for prevention. But does growing up in deprived neighborhoods matter above and beyond a genetic liability to behavior problems, if genetically vulnerable families tend to concentrate in poor neighborhoods? A nationwide study of 2-year-old twins shows that children in deprived neighborhoods were at increased risk for emotional and behavioral problems over and above any genetic liability. Environmental factors shared by members of a family accounted for 20% of the population variation in children's behavior problems, and neighborhood deprivation accounted for 5% of this family wide environmental effect. The results suggest that the link between poor neighborhoods and children's mental health may be a true environmental effect, and demonstrate that genetic designs are environmentally informative and can be used to identify modifiable risk factors for promoting child health. PMID- 11273397 TI - The role of inhibition in the regulation of sequential action. AB - We investigated the regulation of sequential action using a new paradigm. Participants learned a sequence of seven stimulus categories and then monitored for them during successive displays. All displays were instances of these categories, presented in pseudorandom order. On each trial, participants monitored for an instance of Category 1, pressed a key on a computer keyboard, then monitored for an instance of Category 2, pressed a key on the keyboard, and so on for all seven categories. Thus, a perfect trial contained exactly seven responses. Intrusion errors were classified as a function of ordinal distance from the current serial position (n). Fewer intrusion errors were made at near serial positions than at far ones, suggesting a gradient of lateral inhibition. In addition, more intrusions were made on n + 1 categories than n - 1 categories, suggesting greater availability of intended than completed goals. In accord with current models of sequential action, the results indicate lateral and self inhibition as important mechanisms in regulation of sequential action. PMID- 11273398 TI - Of wealth and death: materialism, mortality salience, and consumption behavior. AB - Theoretical work suggests that feelings of insecurity produce materialistic behavior, but most empirical evidence is correlational in nature. We therefore experimentally activated feelings of insecurity by having some subjects write short essays about death (mortality-salience condition). In Study 1, subjects in the mortality-salience condition, compared with subjects who wrote about a neutral topic, had higher financial expectations for themselves 15 years in the future, in terms of both their overall worth and the amount they would be spending on pleasurable items such as clothing and entertainment. Study 2 extended these findings by demonstrating that subjects exposed to death became more greedy and consumed more resources in a forest-management game. Results are discussed with regard to humanistic and terror-management theories of materialism. PMID- 11273399 TI - Psychological Science in the Public Interest: the case for juried analyses. AB - The inaugural issue of Psychological Science in the Public Interest (PSPI), a new publishing initiative by the American Psychological Society, accompanies this issue of Psychological Science. The report it contains, "Psychological Science Can Improve Diagnostic Decisions," by John Swets, Robyn Dawes, and John Monahan, represents a careful effort by those authors to summarize the potential of modern psychological science to enhance real-world diagnostic decisions. Such decisions (Is a cancer present? Will this individual commit violence? Will an impending storm strike? Will this applicant succeed?) are prevalent and crucial to the lives of individuals and to the well-being of our society. Subsequent issues of PSPI will address other important topics of public interest in areas where psychological science may have the potential to inform and improve public policy. Each of those reports will also represent the efforts of a distinguished team of scientists to report the available evidence, and the implications of that evidence, fairly and comprehensively. In this article, we describe the goals, procedures, and potential of PSPI. PMID- 11273400 TI - The causal influences of attributions on emotions: a procedural priming approach. AB - According to attributional theories of emotion, feelings of guilt presuppose that the causes of a negative event are located within the individual (internal attribution), whereas feelings of anger presuppose that the causes of the eliciting event are located outside the individual (external attribution). This study tested whether these attributions in fact exert the claimed causal influence on emotional experiences. The study employed a procedural priming technique in which neutral events were repeatedly attributed either to oneself (internal attribution) or to another person (external attribution). Subsequently, participants were exposed to a negative event that was ambiguous as to its causes. The results reveal that the prior repeated use of internal attributions enhanced the tendency to experience guilt, whereas the repeated use of external attributions enhanced the tendency to experience anger. These findings support the assumption that attributions exert a causal influence on emotions. PMID- 11273401 TI - Exploring Hindu Indian emotion expressions: evidence for accurate recognition by Americans and Indians. AB - Subjects were presented with videotaped expressions of 10 classic Hindu emotions. The 10 emotions were (in rough translation from Sanskrit) anger, disgust, fear, heroism, humor-amusement, love, peace, sadness, shame-embarrassment, and wonder. These emotions (except for shame) and their portrayal were described about 2,000 years ago in the Natyasastra, and are enacted in the contemporary Hindu classical dance. The expressions are dynamic and include both the face and the body, especially the hands. Three different expressive versions of each emotion were presented, along with 15 neutral expressions. American and Indian college students responded to each of these 45 expressions using either a fixed-response format (10 emotion names and "neutral/no emotion") or a totally free response format. Participants from both countries were quite accurate in identifying emotions correctly using both fixed-choice (65% correct, expected value of 9%) and free-response (61% correct, expected value close to zero) methods. PMID- 11273402 TI - Is infant-directed speech prosody a result of the vocal expression of emotion? AB - Many studies have found that infant-directed (ID) speech has higher pitch, has more exaggerated pitch contours, has a larger pitch range, has a slower tempo, and is more rhythmic than typical adult-directed (AD) speech. We show that the ID speech style reflects free vocal expression of emotion to infants, in comparison with more inhibited expression of emotion in typical AD speech. When AD speech does express emotion, the same acoustic features are used as in ID speech. We recorded ID and AD samples of speech expressing love-comfort, fear, and surprise. The emotions were equally discriminable in the ID and AD samples. Acoustic analyses showed few differences between the ID and AD samples, but robust differences across the emotions. We conclude that ID prosody itself is not special. What is special is the widespread expression of emotion to infants in comparison with the more inhibited expression of emotion in typical adult interactions. PMID- 11273403 TI - A search asymmetry reversed by figure-ground assignment. AB - We report evidence demonstrating that a search asymmetry favoring concave over convex targets can be reversed by altering the figure-ground assignment of edges in shapes. Visual search for a concave target among convex distractors is faster than search for a convex target among concave distractors (a search asymmetry). By using shapes with ambiguous local figure-ground relations, we demonstrated that search can be efficient (with search slopes around 10 ms/item) or inefficient (with search slopes around 30-40 ms/item) with the same stimuli, depending on whether edges are assigned to concave or convex "figures." This assignment process can operate in a top-down manner, according to the task set. The results suggest that attention is allocated to spatial regions following the computation of figure-ground relations in parallel across the elements present. This computation can also be modulated by top-down processes. PMID- 11273404 TI - Multiplicative effects of intention on the perception of bistable apparent motion. AB - When viewing ambiguous displays, observers can, via intentional efforts, affect which perceptual interpretation they perceive. Specifically, observers can increase the probability of seeing the desired percept. Little is known, however, about how intentional efforts interact with sensory inputs in exerting their effects on perception. In two experiments, the current study explored the possibility that intentional efforts might operate by multiplicatively enhancing the stimulus-based activation of the desired perceptual representation. Such a possibility is suggested by recent neurophysiological research on attention. In support of this idea, when we presented bistable apparent motion displays under stimulus conditions differentially favoring one motion percept over the other, observers' intentional efforts to see a particular motion were generally more effective under conditions in which stimulus factors favored the intended motion percept. PMID- 11273405 TI - Age differences in the control of looking behavior: do you know where your eyes have been? AB - Previous research has shown that during visual search young and old adults' eye movements are equivalently influenced by the appearance of task-irrelevant abrupt onsets. The finding of age-equivalent oculomotor capture is quite surprising in light of the abundant research suggesting that older adults exhibit poorer inhibitory control than young adults on a variety of different tasks. In the present study, we examined the hypothesis that oculomotor capture is age invariant when subjects' awareness of the appearance of task-irrelevant onsets is low, but that older adults will have more difficulty than young adults in inhibiting reflexive eye movements to task-irrelevant onsets when awareness of these objects is high. Our results were consistent with the level-of-awareness hypothesis. Young and old adults showed equivalent patterns of oculomotor capture with equiluminant onsets, but older adults misdirected their eyes to bright onsets more often than young adults did. PMID- 11273406 TI - Pigeons flexibly time or count on cue. AB - In Experiment 1, pigeons were presented with a sequence of light flashes and cued to peck a key for reward either after a fixed time or after a fixed number of flashes. Curves that showed the rate of key pecking over time within trials indicated that peak rates of response were reached near the fixed time on timing cued trials and near the fixed number of flashes on counting-cued trials. In Experiment 2, the key cue was shifted from timing to counting or from counting to timing midway through a trial. The peak times reached after the cue change indicated that pigeons kept track of time while cued to count but did not count while cued to time. These findings suggest a basic asymmetry in the dual-mode model of timing and counting. PMID- 11273407 TI - Exaggerating temporal differences enhances recognition of individuals from point light displays. AB - Humans are very good at perceiving each other's movements. In this article, we investigate the role of time-based information in the recognition of individuals from point light biological motion sequences. We report an experiment in which we used an exaggeration technique that changes temporal properties while keeping spatial information constant; differences in the durations of motion segments are exaggerated relative to average values. Participants first learned to recognize six individuals on the basis of a simple, unexaggerated arm movement. Subsequently, they recognized positively exaggerated versions of those movements better than the originals. Absolute duration did not appear to be the critical cue. The results show that time-based cues are used for the recognition of movements and that exaggerating temporal differences improves performance. The results suggest that exaggeration may reflect general principles of how diagnostic information is encoded for recognition in different domains. PMID- 11273408 TI - Differences, not ratios, control choice in an experimental analogue to foraging. AB - In choice between outcomes with different delays to reinforcement, most theories require that choice be governed by the ratio of the delays, not by the difference between them, a requirement also consistent with Weber's law. Instead, delay reduction theory and optimal-foraging theory stipulate, under conditions of the present experiments, that the difference between the delays, and not the ratio between them, controls choice. This prediction was assessed using a procedure, widely used in foraging experiments, in which pigeons chose between accepting and rejecting either of two delays when offered. Across conditions, the delays either differed by a constant amount, with the ratio between the delays varying, or differed by changing amounts, with the ratio between the delays constant. In each of six experiments, rate of acceptance of the longer delay depended only on the difference between the two delays and not on the ratio between them, supporting delay-reduction and foraging theory. PMID- 11273409 TI - Phonology matters: the phonological frequency effect in written Chinese. AB - Does phonology play a role in silent reading? This issue was addressed in Chinese. Phonology effects are less expected in Chinese than in alphabetical languages like English because the basic units of written Chinese (the characters) map directly into units of meaning (morphemes). This linguistic property gave rise to the view that phonology could be bypassed altogether in Chinese. The present study, however, shows that this is not the case. We report two experiments that demonstrate pure phonological frequency effects in processing written Chinese. Characters with a high phonological frequency were processed faster than characters with a low phonological frequency, despite the fact that the characters were matched on orthographic (printed) frequency. The present research points to a universal phonological principle according to which phonological information is routinely activated as a part of word identification. The research further suggests that part of the classic word-frequency effect may be phonological. PMID- 11273410 TI - Perception-action dissociations of a walkable Muller-Lyer configuration. AB - These studies examined the role of spatial encoding in inducing perception-action dissociations in visual illusions. Participants were shown a large-scale Muller Lyer configuration with hoops as its tails. In Experiment 1, participants either made verbal estimates of the extent of the Muller-Lyer shaft (verbal task) or walked the extent without vision, in an offset path (blind-walking task). For both tasks, participants stood a small distance away from the configuration, to elicit object-relative encoding of the shaft with respect to its hoops. A similar illusion bias was found in the verbal and motoric tasks. In Experiment 2, participants stood at one endpoint of the shaft in order to elicit egocentric encoding of extent. Verbal judgments continued to exhibit the illusion bias, whereas blind-walking judgments did not. These findings underscore the importance of egocentric encoding in motor tasks for producing perception-action dissociations. PMID- 11273411 TI - Are real moods required to reveal mood-congruent and mood-dependent memory? AB - While simulating, or acting as if, they were either happy or sad, university students recounted emotionally positive, neutral, or negative events from their personal past. Two days later, subjects were asked to freely recall the gist of all of these events, and they did so while simulating a mood that either did or did not match the one they had feigned before. By comparing the present results with those of a previous study, in which affectively realistic and subjectively convincing states of happiness and sadness had been engendered experimentally, we searched for--and found--striking differences between simulated and actual moods in their impact an autobiographical memory. In particular, it appears that the mood-congruent effects elicited by simulated moods are qualitatively different from those evoked by induced moods, and that only authentic affects have the power to produce mood-dependent effects. PMID- 11273412 TI - Self-regulatory failure: a resource-depletion approach. AB - Three studies were conducted to test the behavioral consequences of effortful self-regulation. Individuals with chronic inhibitions about eating were exposed to situations varying in level of self-regulatory demand. Subsequently, participants' ability to self-regulate was measured. Two studies manipulated self regulatory demand by exposing participants to good-tasting snack foods, whereas a third study required participants to control their emotional expressions. As hypothesized, exerting self-control during the first task led to decrements in self-control on a subsequent task. Moreover, these effects were not due to changes in affective state and occurred only when self-control was required in the first task. These findings are explained in terms of depletion of self regulatory resources, which impairs successful volitional control. PMID- 11273414 TI - Development of a single-code/default coding strategy in pigeons. AB - We tested the hypothesis that pigeons could use a cognitively efficient coding strategy by training them on a conditional discrimination (delayed symbolic matching) in which one alternative was correct following the presentation of one sample (one-to-one), whereas the other alternative was correct following the presentation of any one of four other samples (many-to-one). When retention intervals of different durations were inserted between the offset of the sample and the onset of the choice stimuli, divergent retention functions were found. With increasing retention interval, matching accuracy on trials involving any of the many-to-one samples was increasingly better than matching accuracy on trials involving the one-to-one sample. Furthermore, following this test, pigeons treated a novel sample as if it had been one of the many-to-one samples. The data suggest that rather than learning each of the five sample-comparison associations independently, the pigeons developed a cognitively efficient single-code/default coding strategy. PMID- 11273415 TI - The acquisition of an appetite. AB - Unlike older animals, weanling-age rats do not seek water to drink when they are dehydrated, despite the fact that a physiological sensitivity to dehydration is present very soon after birth. We demonstrate here that the appetitive behaviors needed to approach and obtain water become linked to dehydration only as a result of specific postnatal learning experience. Preventing early experience with dehydration retards the developmental emergence of dehydration-induced, water oriented behavior in young rats. But a single pairing of water with dehydration can establish an appetitive response. These findings reveal a critical role of early learning in the development of goal-oriented behavior. Such a learning process is potentially characteristic of other behavioral systems, from the most basic appetites to complex motives. PMID- 11273413 TI - Functional neuroanatomy of the cognitive process of mapping during discourse comprehension. AB - We used functional magnetic resonance imaging (fMRI) to identify brain regions involved in the process of mapping coherent discourse onto a developing mental representation. We manipulated discourse coherence by presenting sentences with definite articles (which lead to more coherent discourse) or indefinite articles (which lead to less coherent discourse). Comprehending connected discourse, compared with reading unrelated sentences, produced more neural activity in the right than left hemisphere of the frontal lobe. Thus, the right hemisphere of the frontal lobe is involved in some of the processes underlying mapping. In contrast, left-hemisphere structures were associated with lower-level processes in reading (such as word recognition and syntactic processing). Our results demonstrate the utility of using fMRI to investigate the neural substrates of higher-level cognitive processes such as discourse comprehension. PMID- 11273416 TI - Expression without recognition: contributions of the human amygdala to emotional communication. AB - A growing body of evidence from humans and other animals suggests the amygdala may be a critical neural substrate for emotional processing. In particular, recent studies have shown that damage to the human amygdala impairs the normal appraisal of social signals of emotion, primarily those of fear. However, effective social communication depends on both the ability to receive (emotional appraisal) and the ability to send (emotional expression) signals of emotional state. Although the role of the amygdala in the appraisal of emotion is well established, its importance for the production of emotional expressions is unknown. We report a case study of a patient with bilateral amygdaloid damage who, despite a severe deficit in interpreting facial expressions of emotion including fear, exhibits an intact ability to express this and other basic emotions. This dissociation suggests that a single neural module does not support all aspects of the social communication of emotional state. PMID- 11273417 TI - On the difficulty of noticing obvious features in patient appearance. AB - Medical students and experts were given head-and-shoulder photographs of patients, each showing a key feature of the patient's problem. Three quarters of these pictures were taken from textbooks. Noticing these supposedly obvious features was difficult and strongly influenced by contextual factors. Both experts and students gained about 20% in diagnostic accuracy by having the key features verbally described for them, although these were clearly visible on the photographs. Conversely, both experts and students reported seeing more of these features when the correct diagnosis was suggested to them. This facilitation resulted from an increase in sensitivity to depicted features, rather than a response bias. The properties of these features that allow such failures of noticing are discussed. PMID- 11273418 TI - Parts outweigh the whole (word) in unconscious analysis of meaning. AB - In unconscious semantic priming, an unidentifiable visually masked word (the prime) facilitates semantic classification of a following visible related word (the target). Three experiments reported here provide evidence that masked primes are analyzed mainly at the level of word parts, not whole-word meaning. In Experiment 1, masked nonword primes composed of subword fragments of earlier viewed targets functioned as effective evaluative primes. (For example, after repeated classification of the targets angel and warm, the nonword anrm acted as an evaluatively positive masked prime.) Experiment 2 showed that this part-word processing was potent enough to oppose analysis at the whole-word level. Thus, smile functioned as an evaluatively negative (!) masked prime after repeated classification of smut and bile. Experiment 3 found no priming when masked word primes contained no parts of earlier targets. These results suggest that robust unconscious priming (a) is driven by analysis of part-word information and (b) requires previous classification of visible targets that contain the fragments later serving as primes. Contrary to a widely held view, analysis of subliminal primes appears not to function at the level of analysis of complete words. PMID- 11273419 TI - Synchronizing visual and language processing: an effect of object name length on eye movements. AB - Are visual and verbal processing systems functionally independent? Two experiments (one using line drawings of common objects, the other using faces) explored the relationship between the number of syllables in an object's name (one or three) and the visual inspection of that object. The tasks were short term recognition and visual search. Results indicated more fixations and longer gaze durations on objects having three-syllable names when the task encouraged a verbal encoding of the objects (i.e., recognition). No effects of syllable length on eye movements were found when implicit naming demands were minimal (i.e., visual search). These findings suggest that implicitly naming a pictorial object constrains the oculomotor inspection of that object, and that the visual and verbal encoding of an object are synchronized so that the faster process must wait for the slower to be completed before gaze shifts to another object. Both findings imply a tight coupling between visual and linguistic processing, and highlight the utility of an oculomotor methodology to understand this coupling. PMID- 11273420 TI - Misremembrance of options past: source monitoring and choice. AB - This study reveals that when remembering past decisions, people engage in choice supportive memory distortion. When asked to make memory attributions of options' features, participants made source-monitoring errors that supported their decisions. They tended to attribute, both correctly and incorrectly, more positive features to the option they had selected than to its competitor. In addition, they sometimes attributed, both correctly and incorrectly, more negative features to the nonselected option. This pattern of distortion may be beneficial to people's general well-being, reducing regret for options not taken. At the same time, it is problematic for memory accuracy, for accountability, and for learning from past experience. PMID- 11273421 TI - Noise exclusion in spatial attention. AB - Precue validity affects the performance of perceptual tasks. These spatial attention effects have been variously attributed to facilitation of processing, capacity allocation, or noise reduction. We used a new attention-plus-external (stimulus)-noise paradigm and model to identify the mechanisms of attention in cue-validity paradigms. A new phenomenon is reported: a large effect of location cue validity in an orientation identification task that specifically occurs when the stimulus is embedded in external (environmental or stimulus) noise. This result identifies the mechanism of the effect as external-noise exclusion, distinguished from stimulus enhancement that manifests itself only in noiseless stimulus environments. PMID- 11273422 TI - Preschoolers' magnitude comparisons are mediated by a preverbal analog mechanism. AB - We report a study of 3- to 5-year-olds who performed a magnitude-comparison task. Stimuli were a series of pairs of arrays that sometimes differed in numerosity, and the children were asked to point to the more numerous array in each pair. The proportion of accurate responses was above chance for all age groups. However, error patterns were consistent with analog models of magnitude representation. Errors varied systematically with the ratio of stimulus pairs. Items with a 2:3 ratio were harder than items with a 1:2 ratio. Performance on posttests of verbal counting ability was variable, but did not predict performance on the numerical discrimination task. We argue that neither verbal counting nor nonnumerical perceptual strategies can explain these results. This study supports the hypothesis that adults and children share preverbal, analog representations of magnitude. PMID- 11273423 TI - Language development in profoundly deaf children with cochlear implants. AB - Although cochlear implants improve the ability of profoundly deaf children to understand speech, critics claim that the published literature does not document even a single case of a child who has developed a linguistic system based on input from an implant. Thus, it is of clinical and scientific importance to determine whether cochlear implants facilitate the development of English language skills. The English language skills of prelingually deaf children with cochlear implants were measured before and after implantation. We found that the rate of language development after implantation exceeded that expected from unimplanted deaf children (p < .001) and was similar to that of children with normal hearing. Despite a large amount of individual variability, the best performers in the implanted group seem to be developing an oral linguistic system based largely on auditory input obtained from a cochlear implant. PMID- 11273424 TI - Reflexive joint attention depends on lateralized cortical connections. AB - Joint attention, the tendency to spontaneously direct attention to where someone else is looking, has been thought to occur because eye direction provides a reliable cue to the presence of important events in the environment. We have discovered, however, that adults will shift their attention to where a schematic face is looking--even when gaze direction does not predict any events in the environment. Research with 2 split-brain patients revealed that this reflexive joint attention is lateralized to a single hemisphere. Moreover, although this phenomenon could be inhibited by inversion of a face, eyes alone produced reflexive shifts of attention. Consistent with recent functional neuroimaging studies, these results suggest that lateralized cortical connections between (a) temporal lobe subsystems specialized for processing upright faces and gaze and (b) the parietal area specialized for orienting spatial attention underlie human reflexive shifts of attention in response to gaze direction. PMID- 11273425 TI - Involuntary listening aids seeing: evidence from human electrophysiology. AB - It is well known that sensory events of one modality can influence judgments of sensory events in other modalities. For example, people respond more quickly to a target appearing at the location of a previous cue than to a target appearing at another location, even when the two stimuli are from different modalities. Such cross-modal interactions suggest that involuntary spatial attention mechanisms are not entirely modality-specific. In the present study, event-related brain potentials (ERPs) were recorded to elucidate the neural basis and timing of involuntary, cross-modal spatial attention effects. We found that orienting spatial attention to an irrelevant sound modulates the ERP to a subsequent visual target over modality-specific, extrastriate visual cortex, but only after the initial stages of sensory processing are completed. These findings are consistent with the proposal that involuntary spatial attention orienting to auditory and visual stimuli involves shared, or at least linked, brain mechanisms. PMID- 11273426 TI - The relative contributions of recognition and search-evaluation processes to high level chess performance: comment on Gobet and Simon. PMID- 11273427 TI - Using a cognitive architecture to examine what develops. AB - Different theories of development propose alternative mechanisms by which development occurs. Cognitive architectures can be used to examine the influence of each proposed mechanism of development while keeping all other mechanisms constant. An ACT-R computational model that matched adult behavior in solving a 21-block pyramid puzzle was created. The model was modified in three ways that corresponded to mechanisms of development proposed by developmental theories. The results showed that all the modifications (two of capacity and one of strategy choice) could approximate the behavior of 7-year-old children on the task. The strategy-choice modification provided the closest match on the two central measures of task behavior (time taken per layer, r = .99, and construction attempts per layer, r = .73). Modifying cognitive architectures is a fruitful way to compare and test potential developmental mechanisms, and can therefore help in specifying "what develops." PMID- 11273429 TI - The legal basis for the Danish Committee on Scientific Dishonesty. AB - The author, a High Court Judge, has chaired the Danish Committee on Scientific Dishonesty (DCSD) since its establishment in 1992. The Committee has worked in the health sector, but from 1999 the scope has been broadened to cover all fields of science. The article describes how the work is organised and the experiences gained. It is stressed, that the difficulty in connection with scientific dishonesty is, first and foremost, to organise a system suitable for investigating cases effectively, professionally, and with proper respect to the fundamental legal rights of the parties involved. The Committee has also spent much effort in determining what can be termed scientific dishonesty and what falls outside this category but which may, nevertheless, be characterised as breaching of good scientific practice. It is emphasised that these rules are not arbitrarily established by the Committee, but formulated in accordance with norms general accepted by opinion leaders in the scientific community. PMID- 11273428 TI - Evolving research misconduct policies and their significance for physical scientists. AB - Scientific misconduct includes the fabrication, falsification, and plagiarism (FFP) of concepts, data or ideas; some institutions in the United States have expanded this concept to include "other serious deviations (OSD) from accepted research practice." It is the absence of this OSD clause that distinguishes scientific misconduct policies of the past from the "research misconduct" policies that should be the basis of future federal policy in this area. This paper introduces a standard for judging whether an action should be considered research misconduct as distinguished from scientific misconduct: by this standard, research misconduct must involve activities unique to the practice of science and must have the potential to negatively affect the scientific record. Although the number of cases of scientific misconduct is uncertain (only the NIH and the NSF keep formal records), the costs are high in terms of the integrity of the scientific record, diversions from research to investigate allegations, ruined careers of those eventually exonerated, and erosion of public confidence in science. Existing scientific misconduct policies vary from institution to institution and from government agency to government agency; some have highly developed guidelines that include OSD, others have no guidelines at all. One result has been that the federal False Claims Act has been used to pursue allegations of scientific misconduct. As a consequence, such allegations have been adjudicated in federal courts, rather than judged by scientific peers. The federal government is now establishing a first-ever research misconduct policy that would apply to all research funded by the federal government regardless of which agency funded the research or whether the research was carried out in a government, industrial or university laboratory. Physical scientists, who up to now have only infrequently been the subject of scientific misconduct allegations, must nonetheless become active in the debate over research misconduct policies and how they are implemented since they will now be explicitly covered by this new federal wide policy. PMID- 11273430 TI - New common federal definition of research misconduct in the United States. PMID- 11273431 TI - Ethics and the responsibility of science. Background paper for the World Science Conference, Budapest June 26-July 1, 1999. PMID- 11273432 TI - From case management to prevention of scientific dishonesty in Denmark. AB - In 1992, The Danish Medical Research Council established a national committee on scientific dishonesty with the twofold task of handling cases of scientific misconduct and taking preventive initiatives. Scientific dishonesty was proven in only five cases, but in another nine cases lesser degrees of deviations from good scientific practice were found. The experiences from a total of 24 treated cases indicated that three key areas were at the basis of most of the accusations and the deviations from good practice: uncertainty about 1) authorship, about 2) rights and duties to use scientific data and about 3) agreements at the initiation of joint studies. As a consequence guidelines on good practice have been issued on these key subjects. PMID- 11273433 TI - Scientific misconduct: ongoing developments. PMID- 11273434 TI - Sociology and psychology within the scope of scientific dishonesty. AB - A survey is undertaken based on qualitative analyses of the cases of scientific misconduct from the Danish Committee on Scientific Dishonesty's first five years of collecting data, with additional information from selected international sources, in which underlying psychological motivations can be judged. PMID- 11273435 TI - Responding to allegations of scientific misconduct: the procedure at the French National Medical and Health Research Institute. AB - Institutions in France are not yet well prepared to respond to allegations of scientific misconduct. Following a serious allegation in late 1997, INSERM, the primary organization for medical and health-related research in France, began to reflect on this subject, aided by scientists and jurists. The conclusions have resulted in establishing a procedure to be followed in cases of alleged misconduct, and also in reinforcing the application of good laboratory practices within each laboratory. Guidelines for authorship practices and scientific assessment must also be considered. Even though each institution must remain responsible for responding to allegations of scientific misconduct within its doors, INSERM would like to see national, European, and international co ordination about the methods of such response. PMID- 11273436 TI - Safeguarding good scientific practice: new institutional approaches in Germany. AB - After summarising three recent case histories of alleged scientific misconduct in Germany, the efforts of the Deutsche Forschungsgemeinschaft (German Research Council) and the Hochschulrektorenkonferenz (German Rectors' Conference) to promote academic and procedural safeguards in favour of professional self regulation in science and scholarship are described in outline. PMID- 11273438 TI - Misconduct in science and the German law. AB - In the past, only norms and rules developed for other types of illegal activities could be applied to misconduct in science in Germany. But only particularly blatant cases of misconduct can be dealt with efficiently in this way. Nowadays, a couple of very important funding agencies and research institutions have enacted special procedures that apply in cases of suspected scientific misconduct. A strongly decentralised system of dealing with misconduct in science is being established in Germany. PMID- 11273437 TI - Scientific misconduct: an international perspective. PMID- 11273439 TI - Ethical implications in the allocation of scarce medical resources in Poland. AB - The health care system in Poland is undergoing major change and it is possible that these changes could affect clinical research. Therefore, the situation of funding of health care is important for the future of medical research in this country. Some questions relevant in this field will be addressed. Since funds for health care and scientific research remain inadequate, their allocation raises moral, economic, legal and organisational dilemmas. The clinical aspects of resource allocation also include physicians' responsibilities towards their patients. Scientific research, clinical medicine, and clinical research have a common denominator: they rely on trust. The physician should be a fiduciary of the patient as well as being a researcher for the benefit of the patient and for society. Some physicians and researchers, despite unethical conduct, escape disclosure and punishment, but decision-makers who wrongly allocate funds for health care and research are never held accountable for their actions. PMID- 11273441 TI - Protecting research integrity. AB - It is not controversial to state that acts of fraud do not belong in the academic world. What is debated is the best way to minimise the risk of fraudulent behaviour. Broadly speaking there are two different approaches to this problem. They differ with regard to whether the main focus is on internal or external control. In this article I argue that the main emphasis should be on internal structures in order to achieve the desired end. Only when the internal structures are in place is it meaningful to adopt external, supportive means to the same end. Invitation to the academic project as such, education and training in research ethics and good research practice, the implementation of good documentation procedures and the implementation of a procedure for investigation of suspicions of fraud which is characterised by efficiency, impartiality and competence are the four primary ingredients in the cure. The first three are suggested to build up the necessary foundation before a structure of investigation procedures are established. PMID- 11273440 TI - Principles of good clinical practice (GCP) in clinical research. AB - Good Clinical Practice is an international quality standard for conducting trials that involve participation of human subjects. Currently, the most widely accepted international document forming the base for GCP is the ICH Harmonised Tripartite Guideline for GCP, which defines in detail the responsibilities and obligations of parties engaged in clinical research. The purpose of this paper is to analyse how compliance with GCP provides protection of the trial subjects and assures quality and credibility of the data obtained. PMID- 11273442 TI - The Medical Research Council's approach to allegations of scientific misconduct. AB - The UK's Medical Research Council (MRC) introduced a specific policy and procedure for inquiring into allegations of scientific misconduct in December 1997; previously cases had been considered under normal disciplinary procedures. The policy formally covers staff employed in MRC units, but those in receipt of MRC grants in universities and elsewhere are expected to operate under similar policies. The MRC's approach is stepwise: preliminary action; assessment to establish prima facie evidence of misconduct; formal investigation; sanctions; and appeal. Strict time limits apply at all stages. The procedure will be evaluated after two years. The indications so far are that the procedure is robust, and its clarity and transparency have been an asset to all parties. The MRC is also convinced that it is equally important to achieve a working culture that fosters integrity. Thus education and training in good research practices are fundamental to the prevention of research misconduct. PMID- 11273443 TI - The American experience: lessons learned. AB - This paper discusses ten lessons learned since 1989 about handling allegations of scientific misconduct involving biomedical and behavioral research supported by the U.S. Public Health Service. PMID- 11273444 TI - Genetic research: can we control it? PMID- 11273445 TI - An analysis of moral issues affecting patenting inventions in the life sciences: a European perspective. AB - Following the 1980 US Supreme Court decision to allow a patent on a living organism, debate has continued on the moral issues involved in biotechnology patents of many kinds and remains a contentious issue for those opposed to the use of biotechnology in industry and agriculture. Attitudes to patenting in the life sciences, including those of the research scientists themselves, are analysed. The relevance of morality to patent law is discussed here in an international context with particular reference to the law of the European Patent Convention administered by the European Patent Office (EPO). The EPO has been the principal forum for opposition to such patents and the few cases under dispute in the EPO are reviewed, including patents for the onco-mouse, human relaxin gene, and the PGS herbicidally resistant plant (gmo). Morality provisions in the European Parliament and Council Directive 98/44/EC are also summarised. PMID- 11273447 TI - Bribery and extortion: can restaurants help? AB - Examples of tipping suggest that the distinction between tipping, bribery and extortion can be questioned. Some well known ideas about bribery will not work if extended to tipping and, indeed, these analyses may founder whether or not tipping, bribery and extortion merge. I suggest that more case study analysis as well as a discussion of the relationship between character and actions are needed. PMID- 11273446 TI - Against over-estimating the role of ethics in technology development. AB - The role of ethics in technology development has been often questioned, especially in the early days of societal reflection of technology. However, the situation has changed dramatically. Ethical consideration now is generally declared to be indispensable in shaping technology in a socially acceptable and sustainable way. The expectations of ethics are large; often even a kind of "New Ethics" is postulated. In the present paper an over-estimation of the role of ethics for technology development is rejected. It is argued that ethical reflection is, indeed, indispensable in certain problem areas and situation types; but there is, on the other hand, space for technology development free from the requirement for ethical reflection. The absence of a requirement for ethical reflection, however, always has to be considered relative to some "morale provisoire" (provisional morality) as an accepted normative framework within which technology development may occur without explicit ethical reflection. If this framework, however, is doubted or is shown to be insufficient the situation changes completely. Ethical reflection in this case becomes necessary, to consider this normative framework in order to offer modifications or supplements. PMID- 11273448 TI - Rethinking technology, revitalizing ethics: overcoming barriers to ethical design. AB - This paper explores the role of ethics in design. Traditionally, ethical questions have been seen as marginal issues in the design of technology. Part of the reason for this stems from the widely held notion of technology being "out of control." This notion is a barrier to what I call "ethical design" because it implies that ethics has no role to play in the development of technology. This view, however, is challenged by recent work in the field of Science and Technology Studies (STS). Looking into the dynamics of technological change, STS scholars argue that human choices are present at every stage of a technology's development and, furthermore, that human values are reflected in the very design of artifacts. This alternative view suggests that ethics can and should be included in the design process. Drawing on examples from the privacy arena, I point to some of the potential advantages of addressing ethical concerns early on in the design of a technology. I conclude with some general strategies for bringing ethics back into design. PMID- 11273449 TI - Equity of access: adaptive technology. AB - In this age of information technology, it is morally imperative that equal access to information via computer systems be afforded to people with disabilities. This paper addresses the problems that computer technology poses for students with disabilities and discusses what is needed to ensure equity of access, particularly in a university environment. PMID- 11273450 TI - Email, voicemail, and privacy: what policy is ethical? AB - Business people repeatedly asked Computer Professionals for Social Responsibility (CPSR) to recommend a policy to deal with email and voicemail. After many such requests to our organization, we attempted to construct guidelines that we could endorse. This paper outlines the guidelines that we proposed and the public reaction to them. The paper discusses the tensions inherent in a business environment, and the means of identifying ethical behavior for both companies and their employees. PMID- 11273451 TI - The development of computer ethics: contributions from business ethics and medical ethics. AB - In this essay, we demonstrate that the field of computer ethics shares many core similarities with two other areas of applied ethics. Academicians writing and teaching in the area of computer ethics, along with practitioners, must address ethical issues that are qualitatively similar in nature to those raised in medicine and business. In addition, as academic disciplines, these three fields also share some similar concerns. For example, all face the difficult challenge of maintaining a credible dialogue with diverse constituents such as academicians of various disciplines, professionals, policymakers, and the general public. Given these similarities, the fields of bioethics and business ethics can serve as useful models for the development of computer ethics. PMID- 11273452 TI - Ethics in the classroom: a reflection on integrating ethical discussions in an introductory course in computer programming. AB - In this paper, we describe our recent approaches to introducing students in a beginning computer science class to the study of ethical issues related to computer science and technology. This consists of three components: lectures on ethics and technology, in-class discussion of ethical scenarios, and a reflective paper on a topic related to ethics or the impact of technology on society. We give both student reactions to these aspects, and instructor perspective on the difficulties and benefits in exposing students to these ideas. PMID- 11273453 TI - Discourse and moral responsibility in biotechnical communication. PMID- 11273454 TI - Discourse ethics for agricultural biotechnology: its limits and its inevitability -a response to Jamieson. PMID- 11273455 TI - "E. O. Wilson as moralist". PMID- 11273456 TI - The use of genetic test information in insurance: the argument from indistinguishability reconsidered. AB - In the bioethical literature, discrimination in insurance on the basis of genetic risk factors detected by genetic testing has been defended and opposed on various ethical grounds. One important argument in favour of the practice is offered by those who believe that it is not possible to distinguish between genetic and non genetic information, at least not for practical policy purposes such as insurance decision-making. According to the argument from indistinguishability, the use of genetic test information for insurance purposes should be permitted, because genetic test information is no different from non-genetic medical information in any relevant respect, therefore it would be inconsistent to prohibit the former whilst permitting the latter. This paper discusses and defends this argument and suggests a new, more tenable foundation. PMID- 11273457 TI - On the philosophical analysis of genetic essentialism. Commentary on: "The use of genetic test information in insurance: the argument from indistinguishability reconsidered". PMID- 11273458 TI - Measuring consensus about scientific research norms. AB - In this paper, we empirically explore some manifestations of norms for the conduct of science. We focus on scientific research ethics and report survey results from 606 scientists who received funding in 1993 and 1994 from the Division of Molecular and Cellular Biology of the Biology Directorate of the National Science Foundation. We also report results for 91 administrators charged with overseeing research integrity at the scientists' research institutions. Both groups of respondents were presented with a set of scenarios, designed by fractional factorial methods, describing different kinds of scientific conduct that in the eyes of some would likely be unethical. Respondents then were asked to evaluate each of these scenarios for how unethical the behavior might be and what kinds of sanctions might be appropriate. We use the responses to consider the nature of consensus around norms related to the practice of science and in particular, similarities and differences between scientists and science administrators. Implications for policy are also discussed. PMID- 11273459 TI - Moral responsibility and the 'ignorant scientist'. AB - The question whether a scientist can be responsible for an outcome of her work which she does not foresee, and so is ignorant of, is addressed. It is argued that ignorance can be a ground for the attribution of responsibility, on condition that there are general principles, rules or norms, that the subject should be aware of. It is maintained that there are such rules which inform the practice of science as a social institution. PMID- 11273460 TI - Trustworthiness in explanation: the obligation to explain well. AB - 'Scientific integrity' certainly requires that data and references be beyond reproach. However, issues within the theory of scientific explanation suggest that there may be more to it than just this. While it is true that some contemporary, pragmatic analyses of explanation suffer from the 'problem of relevance' (an inability to ensure that explanations which are paradigmatic technically are relevant to the question being posed), it does not seem to be true that the addition of formal, metaphysical constraints is necessary to solve this problem. I argue that, when viewed as requests for help with an epistemic problem, explanation-seeking questions reveal the existence of a set of moral criteria centered in trust which, when satisfied, prevent trivial or irrelevant explanations from being offered, thereby broadening the concept of 'scientific integrity'. PMID- 11273461 TI - The wisdom of nature in integrating science, ethics and the arts. AB - This paper deals with an approach to the integration of science (with technology and economics), ethics (with religion and mysticism), the arts (aesthetics) and Nature, in order to establish a world-view based on holistic, evolutionary ethics that could help with problem solving. The author suggests that this integration is possible with the aid of "Nature's wisdom" which is mirrored in the macroscopic pattern of the ecosphere. The corresponding eco-principles represent the basis for unifying soft and hard sciences resulting in "deep sciences". Deduction and induction will remain the methodology for deep sciences and will include conventional experiments and aesthetic and sentient experiences. Perception becomes the decisive factor with the senses as operators for the building of consciousness through the subconscious. In this paper, an attempt at integrating the concepts of the "true", the "right" and the "beautiful" with the aid of Nature's wisdom is explained in more detail along with consequences. PMID- 11273462 TI - The Israel Press Council: review and suggestions for improvement. AB - The aim of this essay is to review the work of the Israel Press Council. The essay considers the history of the Press Council, analysing the way it has developed, its work, and how it reached its current status. It is argued that the existing situation is far from satisfactory, and that the media should advance more elaborate mechanisms of self-control, empowering the Press Council with greater authority and equipping it with substantive ability to sanction. PMID- 11273463 TI - An approach to integrating "professional responsibility" in engineering into the capstone design experience. AB - ABET 2000 Criteria encourages development of proficiency in professional responsibility in engineering as part of the undergraduate curriculum. This paper discusses the use of industrially sponsored capstone design projects to encourage active discussion of professional responsibility in engineering that naturally occurs during the engineering design process. The paper also discusses student participation in designing responses and approaches to issues such as engineering ethics. The paper includes specific examples of topics addressed by students and the approaches developed (by students) in addressing these issues. PMID- 11273464 TI - Service-learning and engineering ethics. AB - This paper explores ways in which service-learning programs can enhance ethics education in engineering. Service-learning programs combine volunteer work and academic study. The National Society for Professional Engineers (NSPE) and American Society for Civil Engineers (ASCE) codes of ethics explicitly encourage engineers to seek opportunities, beyond their work-related responsibilities, to serve their communities. Examples of how this can be encouraged as a part of the educational experiences of engineering students are explored. PMID- 11273465 TI - Examples of real world engineering ethics problems. AB - Nine examples are presented illustrating the kinds of problems encountered in actual practice by conscientious engineers. These cases are drawn fom the records of the IEEE Ethics Committee, and from the experience of the ethics helpline initiated recently by the Online Ethics Center for Engineering and Science. They range from situations in which companies try to cheat one another to those in which human health and safety are jeopardized. In one case, an engineer learned that even a quiet resignation can prove very costly in a personal sense. Some ways in which professional societies might make ethical practice of engineering somewhat easier are mentioned. PMID- 11273466 TI - Redefining and refining the process of clinical diagnosis. PMID- 11273467 TI - A study on relative contributions of the history, physical examination and investigations in making medical diagnosis. AB - Here we report an attempt to quantitate the relative contributions of the history, physical examination and investigations in making medical diagnosis. In this prospective study of 100 patients, with new or previously undiagnosed conditions, we listed their differential diagnosis with confidence score; after the history, after physical examination and after the investigations. In two patients no definite final diagnoses could be arrived even after extensive investigation--these two cases were excluded from the study. In seventy seven patients (78.58%) patients, the history led to diagnosis. The physical examination led to diagnosis in eight patients (8.17%); and investigations led to diagnosis in 13 patients (13.27%). The confidence in correct diagnosis increased from 6.36 on a scale of one to ten after the history to 7.57 after physical examination and 9.84 after investigations--implying that history, physical examination and investigation have their own limitation at each stage and an integrative approach is needed in making a medical diagnosis with more emphasis on history. PMID- 11273468 TI - Comparative study of visceral and parietal pleural biopsy in the etiological diagnosis of pleural diseases. AB - OBJECTIVE: The present study was planned to evaluate the efficacy and diagnostic reliability of conoctional parietal pleural biopsy to a technique of visceral pleural biopsy. METHOD: Study comprises of 54 diagnosed cases of pleural effusion and after establishing the clinical diagnosis for probable etiological causes. Then parietal pleural biopsy using absents punch biopsy needle and vesceral pleural biopsy using Prabhudesai et al technique was taken in all these patients. Size of the tissue yield; percentage of biopsies; diagnostic yield and sensitivity for these two techniques were compared. RESULTS: A definitive etiological diagnosis could be reached in 52 out of 54 patients on the basis of pleural biopsy (33 tuberculous, 16 malignant and 3 pyogenic), 23 (69.7%) tuberculous effusion patients were diagnosis by visceral pleural biopsy and 14 (42.4%) by parietal pleural biopsy out of 33 diagnosed tuberculous effusion cases. While for the 16 malignant effusions the visceral pleural biopsy showed suggestive histological change in 13 (81.25%) patients and the parietal pleural biopsy in seven (43.8%) with five (31.25%) of these patients being positive by both. All three pyogenic effusions showed only nonspecific inflammatory change in both pleurae. CONCLUSION: The mean size of biopsy sample obtained with modified Prabhudesai et al technique was significantly larger than that of the parietal pleural biopsy with Abrams punch (4.85 mm2 V/s 2.5 mm2 with P < 0.01). Adequate pleural tissue was identifiable in 94.4% and 90.7% of cases, respectively. The modified Prabhudesai et al technique proved to be effective safe and easily learnt. Visceral pleural sampling using this technique is a definite superior addition to the present diagnostic armamentarium of an idiopathic pleural effusion and its routine application together with parietal pleural biopsy will help to establish a definitive diagnosis in majority of patients with idiopathic pleural effusions. PMID- 11273469 TI - Nutritional risk factors in esophageal cancer. AB - OBJECTIVE: The present case-control study was undertaken with the objective to study the nutritional risk factors associated with esophageal cancer. METHODOLOGY: One hundred and fifty diagnosed esophageal cancer patients and an equal number of healthy individuals constituted the patient and control groups, respectively. Dietary consumption pattern during the preceding 20 years prior to the diagnosis of esophageal cancer was assessed utilising the standard food frequency questionnaire method. Information on alcohol consumption, smoking habits, chewing of betel leaf with tobacco was also collected. RESULTS: Multivariate analysis revealed that the risk of esophageal cancer was 7.81 times (p < 0.01) higher with daily consumption of alcohol. The risk increased to 3.16 times (p < 0.01) with the daily habit of chewing of betel leaf with tobacco. Nearly a two fold risk was observed when the consumption of "other vegetables" was less than four times per week. A 1.95 times (p < 0.01) increase in risk was observed with the daily habit of bidi smoking. CONCLUSION: Cancers in general are multifactorial in origin, and several environmental interactions are possible. It is not easy to quantify the contribution of diet to cancer risk. However, the results of the present study suggested that nutritional factors do play a role. PMID- 11273470 TI - Non-epileptic manifestations in patients with single enhancing computed tomography lesions. AB - OBJECTIVES AND METHODS: Single enhancing lesions are common computed tomographic (CT) abnormality in patients with epilepsy. In this series we are reporting 13 unusual cases with varied non-epileptic neurological manifestations in patients with ring or disk enhancing CT lesions. RESULTS: Acute, stroke like non-vascular focal neurological deficits (hemiparesis in four patients, and crural monoparesis, Broca's aphasia, homonymous hemianopia, hemichorea in one patient each) were frequent non-epileptic manifestations. Episodic vascular type of headache was seen in three patients, one patient had headache because of raised intracranial pressure. One patient presented with acute confusional state. All these patients were treated symptomatically, and with oral corticosteroids. CT lesions disappeared in 8-12 weeks time in all patients except in one patient with chorea where the lesion calcified. Significant clinical improvements were noted in all the patients. CONCLUSIONS: Several non-epileptic manifestations can also be associated with single enhancing CT lesions, and like epileptic disorders these disorders also have a benign course. Corticosteroids, probably, hasten the clinical improvement and produce early resolution of the CT lesions. PMID- 11273472 TI - Etiology based prevalence of Budd-Chiari syndrome in eastern India. AB - BACKGROUND: Diagnosis of Budd-Chiari syndrome (BCS) is often missed unless its possibility has been kept in mind. Obstruction of inferior vena cava (IVC) is reportedly the most frequent cause of BCS in Afro-Asian variety. AIM: An attempt was made to classify BCS (in an eastern Indian population) etiopathologically. PATIENTS AND METHODS: Thirty consecutive cases of BCS presenting over a period of five years were included. Following a thorough physical examination, necessary investigations (including coagulation profile, ultrasonography (with Doppler study) of hepatobiliary tract, hepatic vein and IVC angiography (n = 22) and liver biopsy (n = 26, including autopsy in two cases) were performed. RESULTS: Mean age at presentation was 32.7 +/- 10.36 years (range 12-60 years) with M:F = 21:9. Clinical presentations included, hepatomegaly in 28 (93.3%), ascites in 27 (90%), splenomegaly in 15 (50%), pain in abdomen in 26 (86.6%), jaundice in 10 (33.3%), back veins in 20 (66.6%) and gastrointestinal bleeding in three (10%) cases. Amongst the total of 30 patients, four, six and 20 cases presented as fulminant, acute and chronic BCS respectively. Twenty four cases of BCS could be diagnosed by ultrasonography alone, while the remainder required angiography for diagnosis. IVC and hepatic vein angiography revealed membranous obstruction in nine, partial stricture of IVC in six, and IVC and/or hepatic vein block in others. The etiopathological nature in 30 cases were as follows: idiopathic membranous obstruction in nine (30%), hepatocellular carcinoma in six (20%), idiopathic stricture in six (20%) cases and one case (3.3%) each of the following: cholangiocarcinoma, renal cell carcinoma, chronic pancreatitis, hydatid cyst in liver, protein S deficiency, oral contraceptive use, nephrotic syndrome (with antithrombin III deficiency), polycythemia rubra vera and chronic lymphatic leukemia. CONCLUSION: Idiopathic membranous obstruction and stricture of IVC are the commonest cause of BCS in the eastern part of India. Hepatocellular carcinoma is also a common cause, presenting in the fulminant form. Ultrasonography may be a helpful screening test for BCS, but IVC and hepatic vein catheterisation is essential for a complete work up of these patients. PMID- 11273471 TI - Serial neuro-electrophysiological studies in acute organophosphate poisoning- correlation with clinical findings, serum cholinesterase levels and atropine dosages. AB - OBJECTIVES AND METHODS: A prospective evaluation of the correlation between the serial clinical findings, serum cholinesterase levels, electrodiagnostic abnormalities and the daily atropine requirement was undertaken in 29 patients with confirmed acute organophosphate poisoning (OPP). RESULTS: Clinical weakness conforming to the pattern found in 'Intermediate Syndrome' was noted in 19 patients (65.55%). It was associated with all types of organophosphate compounds and occurred in all patients in whom the serum cholinesterase on admission was less than 200 units. Three types of electrodiagnostic abnormalities were noted: single supramaximal electrical stimulus induced repetitive response, a decrement- increment response to 30 Hz repetitive nerve stimulation (RNS) and a decremental responses to 30 Hz RNS. The 30 Hz decremental response correlated best with the presence of clinically detectable weakness (sensitivity = 61.72%; specificity = 81.54%; positive predictive value = 73.91%; negative predictive value = 71.62%). Time trends evaluation revealed that the peak daily atropine dosages were given at a mean of 1.76 +/- 0.83 days in comparison to a mean nadir of serum cholinesterase of 2.48 +/- 1.97 days and a mean nadir of 9:1 ratio of 2.65 +/- 1.76 days. The 2-tailed correlation coefficient analysis and simple regression analysis revealed a positive correlation between serum cholinesterase levels and the 9:1 ratios (correlation coefficient: 0.59). A negative correlation was observed between the 9:1 ratios and the daily atropine requirement (correlation coefficient: -0.57) and between serum cholinesterase levels and daily atropine requirement (correlation coefficient: -0.49). CONCLUSIONS: At admission, level of serum cholinesterase of less than 200 units is a predictor and the 30 Hz RNS decremental response could be a useful marker for the 'Intermediate Syndrome'. PMID- 11273474 TI - Bone scanning for bone metastasis in carcinoma cervix. AB - AIM: Bone metastasis in cervical cancer is rare. With the aim of defining the frequency of bony metastasis in patients of carcinoma cervix, with clinical suspicion of metastasis, we performed bone scan in 38 such patients. RESULTS: Twelve out of the 38 patients were confirmed as having metastasis. All the patients were also detected by bone scan (100% sensitivity). CONCLUSION: Bone scan should be investigation of choice for screening patients of carcinoma cervix with symptoms suggestive of metastasis in all stage of the disease. Bone scan is the most sensitive method for detection of bone metastasis. Bone scan offers the additional advantages of allowing a review of the kidney size to look for ureteric involvement and subsequent hydronephrosis. PMID- 11273473 TI - Doxycycline in the treatment of rheumatoid arthritis--a pilot study. AB - OBJECTIVE: To assess the efficacy and safety of doxycycline as a disease modifying anti-rheumatic drug (DMARD) in rheumatoid arthritis (RA) and compare it with methotrexate, a standard DMARD. MATERIAL AND METHODS: A single (assessor) blind prospective study with 15 patients of RA randomized to doxycycline and 14 to methotrexate. Baseline disease characteristics were similar in both groups. RESULTS: All disease activity measures studied viz. tender and swollen joint counts, physician and patient global assessment, visual analogue pain scale, health assessment questionnaire and ESR improved in both the treatment groups after six months of treatment. The difference between doxycycline and methotrexate was not statistically significant. No major side effects necessitating drug withdrawal were reported from either group. The side effects were few and mostly gastrointestinal. CONCLUSION: Doxycycline is a safe disease modifying drug in RA whose effect is sustained at six months. It compared favourably with methotrexate over a six month follow up. PMID- 11273475 TI - Pictorial CME. Choroid tubercles. PMID- 11273476 TI - Serum insulin assay: an important therapeutic tool in management of freshly diagnosed type 2 diabetes mellitus. AB - OBJECTIVES: The study was performed to see that, whether metabolic control and response to treatment in freshly diagnosed patients of type 2 diabetes mellitus is affected by primary pathology (hyperinsulinemia/inappropriate insulin secretion). METHODS: One hundred and eight freshly diagnosed patients of type 2 diabetes mellitus with age range from 30-65 years were followed for a period of three months. The blood glucose, serum triglyceride, and serum insulin levels were determined in each patient. Patients were found to have either higher or normal to low serum insulin values at fasting, and accordingly patients were distributed into two groups; group one (normal to low initial fasting serum insulin level i.e. < or = 30 microU/ml) and group two (high fasting serum insulin level i.e. > or = 30 microU/ml). Each group was further divided into two subgroups A and B. Subgroup A was treated with glipizide and B with metformin. RESULTS: Diabetic patients who had fasting hyperinsulinemia (n = 53, 100%) had blood pressure > or = 140/90 at the time of presentation. Patients who had fasting serum insulin within normal range only 30% (n = 17) had hypertension. Patients of group one had good recovery from hyperglycemia and reduction in triglyceride values when treated with sulphonylurea (subgroup A) as compared to patients treated with biguanide (subgroup B). On the contrary patients of group two showed poor glycemic control, increase in blood pressure and rise in serum triglyceride titre when treated with sulphonylurea (subgroup A) while in the same group biguanide effectively produced euglycemia with normalization of blood pressure and decrease in triglyceride levels (subgroup B). CONCLUSION: Assessment of initial serum insulin levels is helpful guide to decide about the type of oral hypoglycemic agent to be used in freshly diagnosed patients to type 2 diabetes mellitus. PMID- 11273477 TI - Prevalence of hepatitis GB virus C/hepatitis G virus infection in blood donors in India. AB - BACKGROUND: Hepatitis GBV-C/HGV is a newly described RNA virus with a parenteral route of transmission. It has been implicated in fulminant hepatitis and chronic viral hepatitis. We undertook to study the prevalence of GBV-C/HGV infection in blood donors of a tertiary care hospital in India. METHOD: Serum of 221 consecutive blood donors was tested for HBsAg, anti-HCV by EIA and HGV RNA by RT PCR. Two sets of primers; one from the 5'non-coding region and other from NS5a region of the HGV genome, were used for amplification. RESULTS: Prevalence of HGV RNA was found to be very low in healthy blood donors. Only two of the 221 (0.9%) donors were found to be HGV RNA positive. HBsAg and AntiHCV were found to be present in 5.43% (12/221) and 1.31% (3/221) respectively. Dual infection was seen in two of the 221 (0.9%) patients; one patient had HBsAg and HGV RNA positivity, while the other, had HBsAg and AntiHCV positivity. CONCLUSION: GBV-C/HGV is an uncommon infection in healthy blood donors in India, especially when compared to the prevalence of HBV and HCV infection. It is therefore unlikely to be an important cause of transfusion associated hepatitis in India. PMID- 11273478 TI - Emerging role of cardiac troponins in acute coronary syndromes. PMID- 11273480 TI - Nature vs. nurture. PMID- 11273479 TI - Diabetes and the liver. PMID- 11273481 TI - Pulmonary alveolar microlithiasis: role of transbronchial lung biopsy. PMID- 11273482 TI - Japanese encephalitis with movement disorder and atypical magnetic resonance imaging. AB - With the advent of magnetic resonance imaging, brain lesions associated with Japanese encephalitis are increasingly being recognized and correlated with movement disorder. Bilateral haemorrhagic thalamic infarcts on MRI, suggested as a characteristic finding in Japanese encephalitis were conspicuous by their absence in this case report of Japanese encephalitis. PMID- 11273484 TI - Neuro-Behcet's disease. AB - Behcet's disease (BD) is a multifocal disorder with an immunogenetic basis, which persists over many years. Initial descriptions mentioned oral and genital ulcers with uveitis. Later a number of other manifestations were added, like skin, joint and neurological. The involvement of nervous system (Neuro-Behcet's) is reportedly uncommon. We hereby report four cases of Neuro-Behcet's, i.e.; two cases of strokes involving multiple areas of the central nervous system and two cases had features of benign intracranial hypertension. All cases had mucocutaneous lesions or other system involvement. Cases satisfied the international study group criteria for diagnosis of BD. All cases were pathergy test positive. In comparison with the literature from Turkey and Greece, which reports a high pathergy positivity, reports from India have shown only few cases to be positive. The prognosis of Neuro-Behcet used to be poor but has recently been improved with reduced mortality, although whether this can be attributed to treatment with steroids and/or cytotoxic agents remains uncertain. PMID- 11273483 TI - Robinow syndrome. AB - Robinow syndrome is a rare congenital abnormality. It is characterized by mesomelic brachymelia, hemivertebrae, dysmorphic facies, genital hypoplasia, micropenis, clinodactyly, camptodactly, hypoplastic nails and moderate short stature. We are documenting the case on the account of its rarity and additional features. PMID- 11273485 TI - Apert syndrome variant with overlapping features of Crouzon syndrome. AB - A rare variant of Apert syndrome having overlapping features of Crouzon syndrome is described. The salient features of the two syndromes are briefly discussed and overlapping features are highlighted. A possible genetic explanation for the same is mentioned. PMID- 11273486 TI - Pancytopenia due to hemophagocytic syndrome as the presenting manifestation of tuberculosis. AB - We report here an unusual hematologic manifestation of tuberculosis. A 40 year old male presented with pancytopenia due to hemophagocytic syndrome. Further investigations revealed he had tuberculosis. PMID- 11273487 TI - Superior mesenteric artery syndrome causing acute intestinal obstruction. PMID- 11273488 TI - Lightning injury with survival. PMID- 11273489 TI - Pheochromocytoma with base of skull metastasis. PMID- 11273491 TI - Medicine in the next millennium. PMID- 11273490 TI - Adverse drug reaction reporting and monitoring: role of practising physicians. PMID- 11273492 TI - Comparison of two regimens on eradication of Helicobacter pylori. PMID- 11273493 TI - Step care treatment of falciparum malaria--a strategy to prevent drug resistance. PMID- 11273494 TI - Effect of weight loss on plasma high density lipoprotein cholesterol. PMID- 11273495 TI - Aluminium phosphide (ALP) is a widely used fumigant posticide. PMID- 11273496 TI - Self injection of insecticide. PMID- 11273497 TI - Is interferon-alpha useful in acute prolonged hepatitis B? PMID- 11273498 TI - Interferon-alpha in the treatment of acute prolonged hepatitis B virus infection. AB - OBJECTIVE: Acute hepatitis B virus (HBV) infection is a self-limiting disease which usually recovers within 4-12 weeks. At the present moment, there is no specific treatment of acute HBV infection. This study investigates the efficacy of interferon-alpha (IFN) therapy in acute prolonged HBV infection to prevent its progression into chronic stage. METHODS: We enrolled a total number of 54 patients for the study in the span of 8 years. Group A patients (n = 20) received IFN-alpha 5 million units (MU) subcutaneously (s.c.) thrice a week for 12 weeks and Group B patients (n = 34) were placed on placebo therapy as control for 12 weeks, with a follow-up for one year. RESULTS: Seroconversion (disappearance of HBsAg, HBeAg, serum HBV DNA and appearance of anti-HBe) in Group A occurred in 16 patients (80%) within 24 weeks of illness, whereas in Group B seroconversion was observed only in 18 patients (53%) within 24 weeks. During follow-up upto one year, two more patients showed seroconversion in Group A but none in Group B. While on treatment no casualty was reported in Group A but one patient died of hepatic coma in Group B. Our observation revealed that in acute prolonged (> 12 weeks but < 24 weeks) hepatitis B, spontaneous seroconversion rate was 53% but with moderate dose of IFN therapy (5 MU, s.c., thrice weekly) from 12 weeks onwards, the seroconversion rate came out to be 80% (upto 24 weeks) which increased upto 90% when followed-up for one year. CONCLUSIONS: IFN-alpha treatment in acute prolonged (> 12 weeks) HBV infection is safe and may prevent its progression to the chronic stage. PMID- 11273499 TI - Near drowning in cold water. AB - Drowning and near drowning is a common cause of accidental death all over the world; specially in road traffic accidents over bridges, swimming pool and boat tragedies. Cold water drowning resulting in hypothermia can lead to instant death before actual drowning. Five cases of near drowning (ND) in cold water, who presented with varied clinical picture like coma with decerebrate rigidity and fixed dilated pupils, hypertension with coma and delayed pulmonary oedema (Secondary drowning) are reported. Energetic management with continuous positive airway pressure was very rewarding in all patients with ND except in one who had transient organic psychosis persisting for two weeks followed by minimal cognitive defect in the form of slow mentation, lack of drive and mild irritability (Bender Gestald Test Score of 53). We have tried to analyse some of the clinical features of ND and the sequel associated with it. The management as well as recent developments in the field are also discussed. PMID- 11273500 TI - Aspergillosis of central nervous system: a study of 21 patients seen in a university hospital in south India. AB - AIM: To study the pattern of central nervous system aspergillosis in a tropical country. MATERIAL AND METHODS: Case records of patients with CNS aspergillosis seen by the authors in a university hospital in south India were reviewed. RESULTS: Of the 21 patients seen during the study period, 16 (76%) patients had intracranial invasion by contiguous spread from paranasal sinuses and one had from ear. Predisposing risk factors were present in two (12.5%) patients with sinocranial aspergillosis and in both patients with disseminated form. Skull bases syndromes were the presenting features in 13 patients, six patients presented with features of intracranial space occupying lesion and two patients had stroke like presentation. CT scans showed intracranial extradural contrast enhancing mass lesions in the anterior, middle or posterior cranial fossa in addition to mass lesions in the paranasal sinuses in 13 patients with sinocranial aspergillosis and in seven orbital lesions. Intracerebral contrast enhancing mass lesion was the CT finding in both the patients with solitary cerebral aspergillus granuloma and in the patient with otocranial aspergillus granuloma. Well-formed granuloma with dense fibrosis was the histological feature in patients with sinocranial and otocranial aspergillosis. Angioinvasion was the pathological feature in both the patients with disseminated form of aspergillosis. Surgical treatment was sub-radical in our series. Survival rates were not good even after surgical and antifungal chemotherapy. CONCLUSIONS: This study suggests that in this part of the world sinocranial aspergillosis is the most common form of histologically verified CNS aspergillosis. Associated predisposing factors and immune status of the host determine the clinical syndrome and type of pathology in patients with CNS aspergillosis. PMID- 11273501 TI - Hypothalamo-pituitary-adrenal axis function in asthmatics taking low dose inhaled beclomethasone dipropionate. AB - OBJECTIVES: Anti-inflammatory drugs, particularly inhaled corticosteroids remain the mainstay of treatment of bronchial asthma. However, these drugs have potential side effects. This study was undertaken to evaluate the effects of inhaled beclomethasone dipropionate (400 and 800 micrograms) over a period of six months on the hypothalamo-pituitary-adrenal axis (HPA) suppression. METHODS: Assessment of the hypothalamo-pituitary-adrenal axis function was carried out by tetracosactrin test at time zero, (before start of treatment), three months, and six months. The baseline values served as the controls for each patient. Serum cortisol was estimated by radioimmuno assay. The response to short tetracosactrin test was classified as normal if serum cortisol levels rose at least 200 nmol/L to a minimum of 500 nmol/L. RESULTS: There were seven patients who were inhaling beclomethasone dipropionate in a dose of 400 micrograms/day and another seven patients were taking the same drug in a dose of 800 micrograms/day. There was no side effect of the drug in any patient except in one patient who had dysphonia. The mean basal cortisol levels were normal in all the subjects at 0, 3 and 6 months of therapy. Tetracosactrin stimulation test was also normal in all patients at all the times who were receiving the dose of 400 micrograms/day. However, one patient (14%) receiving 800 micrograms/day had HPA axis suppression at six months. Two patients in this group also had low basal cortisol levels. There was no clinical evidence of such suppression/deficiency. CONCLUSION: Beclomethasone dipropionate in a dose of 800 micrograms/day may suppress the hypothalamo-pituitary-adrenal axis if used for long periods (six months). However, this may not have any clinical significance. PMID- 11273502 TI - Retardation of coronary atherosclerosis with yoga lifestyle intervention. AB - BACKGROUND: Yoga has potential for benefit for patients with coronary artery disease though objective, angiographic studies are lacking. MATERIAL AND METHODS: We evaluated possible role of lifestyle modification incorporating yoga, on retardation of coronary atherosclerotic disease. In this prospective, randomized, controlled trial, 42 men with angiographically proven coronary artery disease (CAD) were randomized to control (n = 21) and yoga intervention group (n = 21) and were followed for one year. The active group was treated with a user-friendly program consisting of yoga, control of risk factors, diet control and moderate aerobic exercise. The control group was managed by conventional methods i.e. risk factor control and American Heart Association step I diet. RESULTS: At one year, the yoga groups showed significant reduction in number of anginal episodes per week, improved exercise capacity and decrease in body weight. Serum total cholesterol, LDL cholesterol and triglyceride levels also showed greater reductions as compared with control group. Revascularisation procedures (coronary angioplasty or bypass surgery) were less frequently required in the yoga group (one versus eight patients; relative risk = 5.45; P = 0.01). Coronary angiography repeated at one year showed that significantly more lesions regressed (20% versus 2%) and less lesions progressed (5% versus 37%) in the yoga group (chi-square = 24.9; P < 0.0001). The compliance to the total program was excellent and no side effects were observed. CONCLUSION: Yoga lifestyle intervention retards progression and increases regression of coronary atherosclerosis in patients with severe coronary artery disease. It also improves symptomatic status, functional class and risk factor profile. PMID- 11273503 TI - Medical problems in surgical patients. AB - OBJECTIVES: There has been an increase in surgical cases due to physical violence, accidents and weapon related injuries. This study was undertaken to assess the medical problems in general surgical cases and due to various injuries. METHODS: All general surgical cases and casualties arising out of weapon related, accidents and blunt injuries admitted to a zonal hospital over a period of one and half years were studied. Only cases who developed a medical illness due to surgical cause, anaesthetic or surgery were included. Evaluation and treatment was done alongwith the surgeon till discharge/death. Details were analysed to ascertain the type of surgical illness, medical complication and the outcome of treatment. RESULTS: There were seven hundred sixty two (53.8%) general surgery cases and six hundred fifty four (46.2%) cases due to various injuries. After excluding cases with prior known medical illness, thirty seven patients were studied. There were eight (1.05%) patients out of seven hundred sixty two general surgery cases and twenty nine (4.43%) out of six hundred fifty four injury cases. Weapon related injury cases were the maximum. Their medical problems related to the organ injury, fat embolism and sepsis. Soft tissue injury was next common, they all developed renal failure. Vehicle accident victim(3) were few and developed fat embolism, aspiration. Two patients out of thirty seven succumbed to post anaesthetic complications. CONCLUSION: The incidence of medical problem in injury related cases are more than in general surgery cases. The type of injury contributes to the medical problem. Increase in mortality and morbidity is because of emergency nature of surgery. This problem needs special study. PMID- 11273504 TI - Adequacy assessment of oxygen therapy. AB - OBJECTIVES: Oxygen administration in the wards is usually not according to prescription and therefore the patients requiring oxygen therapy does not get optimal benefits. We planned to assess the need and adequacy of oxygen therapy as was given in wards of SMS Hospital, Jaipur. METHODOLOGY: We studied sixty-six patients in medical and surgical wards who were receiving oxygen therapy through various modes of delivery. Oxygen therapy system was checked in detail and oxygen saturation (SaO2) was measured by pulse oximetery. RESULTS: In our study, we found that no oxygen was flowing from cylinder head in 24 cases (35.5%) while in another 23 cases (35.2%) oxygen was flowing at lower than prescribed flow rates. Leakage in tubes and connections were found in nine cases (13.4%). None of our case was receiving oxygen as per prescription. After correction of faults, all patients showed improvement in SaO2. The criteria of starting oxygen therapy were met only in 47 patients (69%) as per American College of Chest Physicians (ACCP). CONCLUSIONS: Oxygen therapy should be administered according to guidelines. Proper monitoring of oxygen therapy is recommended to ensure adequate oxygenation and to save precious oxygen from wastage. Pulse oximeter is a simple, noninvasive and reliable method to assess it. PMID- 11273505 TI - Ocular cysticercosis--a review of 25 cases. AB - OBJECTIVE: To study the clinical manifestations, management and outcome of cases of ocular cysticercosis in India. METHODS: Retrospective analysis of records of patients presenting to the ophthalmology clinics, with cysticercosis, during years 1990-98. RESULTS: A total of 25 patients had ocular cysticercosis, during the period studied. Majority could be surgically removed without residual effects. However four patients (16%) had no useful vision left in the involved eye. CONCLUSION: Cysticercosis is still endemic in India and contributes to preventable blindness. Improving sanitation and health awareness are the only ways to prevent this infection. PMID- 11273506 TI - Colonic tuberculosis: colonoscopic appearance and clinico-pathologic analysis. AB - BACKGROUND: There has been a resurgence of interest in intestinal tuberculosis because of acquired immunodeficiency syndrome (AIDS) epidemic sweeping our country. Role of colonoscopy and colonoscopy directed histology for diagnosing the disease have been emphasised since last few years. AIMS AND OBJECTIVES: To know the colonoscopic features in patients with intestinal tuberculosis and to study the clinicopathological findings in the same. METHODS: We studied twenty one patients with intestinal tuberculosis referred to us between 1993-1997. Colonoscopy was done in all patients and biopsy specimens were collected from the site of lesion during the procedure. RESULTS: Ileocaecal disease was found in 9 patients, ileocaecal with contiguous ascending colon involvement in eight and segmental colonic tuberculosis in four cases. The colonoscopic findings included nodules in seven patients, nodules with ulcerations in three, ulcerations alone in seven, nodules with strictures in three and polypoidal mass in one patient. Eight cases revealed granuloma on histopathology. CONCLUSIONS: Though bacteriological and histological assessment of tissue is essential to differentiate tuberculosis from other disorders, we stress the importance of colonoscopic appearances in diagnosing tuberculosis. We also recommend antituberculous chemotherapy in patients with high clinical suspicion of tuberculosis on the basis of colonoscopic appearance alone after ruling other causes on histopathological examination. PMID- 11273507 TI - Pictorial CME. Haemophilic arthropathy. PMID- 11273508 TI - ACE inhibitors--revisited. PMID- 11273509 TI - Role of echocardiography in acute myocardial infarction. PMID- 11273510 TI - Nutrition in dialysis patients. AB - Malnutrition is a common clinical problem in dialysis patients, which is multifactorial in origin. It is most often found in a patient of chronic renal failure (CRF) during the period when the glomerular filtration rate (GFR) falls below 10 ml/min, but dialysis is yet to be started. The loss of proteins, aminoacids and other essential nutrients during the procedure of dialysis may further aggravate the malnutrition. Poor nutrition in dialysis patients is associated with increased morbidity and mortality in the form of delayed wound healing, malaise, fatigue, increased susceptibility to infection and poor rehabilitation. In view of the above consequences, all patients on dialysis must undergo nutritional assessment. It is very vital to maintain good nutritional status in-patients on dialysis by adequate protein and calories intake, appropriate supplementation of iron, calcium, minerals and water-soluble vitamins and, of course, the supplementation should be individualised. Nutritional needs are enhanced in presence of stresses like infection or surgery to limit excessive tissue catabolism and therefore, these are the situations, which demand intensive nutrition therapy. Total parenteral nutrition (TPN) may be required for patients on dialysis in intensive care unit, using a central venous catheter. However, enteral route is always preferred to parenteral ones, whenever possible. Even after adequate dialysis has been given, dietary counselling is often required for both hemodialysis and peritoneal dialysis patients to ensure that they ingest the recommended amount of protein, calories and essential micronutrients. PMID- 11273511 TI - Mitral valve prolapse syndrome. PMID- 11273512 TI - Atrioventricular nodal reentrant tachycardia with atrioventricular block. PMID- 11273513 TI - Infective endocarditis as a cause of fever in hemodialysis patients. AB - Vascular access infections are common in maintenance hemodialysis patients especially with dual lumen cuffed catheter. Persistent infections may lead to valvular seeding and the development of infective endocarditis. Though antibiotic therapy may often suffice, many patients may require surgical correction which carries a high risk of mortality. However appropriate preoperative therapy may considerably reduce the risk of surgery in maintenance hemodialysis patients. PMID- 11273514 TI - Citric acid treatment of diabetic foot: a simple and effective approach. PMID- 11273515 TI - Stevens-Johnson syndrome due to carbamazepine. AB - Two patients with psychotic disorders who developed Stevens-Johnson Syndrome while on treatment with carbamazepine is reported due to its rarity. Dermatological side-effects of carbamazepine may be more common in psychiatric as compared to neurological patients. PMID- 11273516 TI - Cerebral infarction in a young male following viper envenomation. AB - Neurological deficits can occur following viper bite. It is usually due to intracerebral or subarachnoid bleed as a result of depletion of clotting factors. A healthy 21-year old male developed motor aphasia and right hemiplegia within two hours of being bit by a viper. Brain CT scan revealed a left frontal lobe infarction. The possible mechanisms for cerebral infarction in this patient are hypotension, endothelial injury, hypercoagulability and vascular. PMID- 11273517 TI - Bleeding complication during coumarin therapy due to amiodarone and azithromycin. AB - A patient with mechanical heart valves developed bleeding, after the introduction of amiodarone and azithromycin. Though the anticoagulant effect could be neutralized, the patient succumbed to heart failure. Any new drug prescribed to patients on anticoagulant must be assessed for its potential for interaction and warrants frequent prothrombin time testing. PMID- 11273518 TI - Primary non-Hodgkin's lymphoma of the larynx presenting with protracted cough. PMID- 11273519 TI - Rhinocerebral mucormycosis presenting as polyneuritis cranialis. PMID- 11273520 TI - Familial parkinsonism with peripheral neuropathy. PMID- 11273521 TI - Cerebral infarction in the territory of anterior cerebral artery in a woman with antiphospholipid syndrome. PMID- 11273522 TI - Randomised control trial on the effective dose of anti-snake venom in cases of snake bite with systemic envenomation. PMID- 11273523 TI - Effect of antioxidant therapy on serum superoxide dismutase activity in patients with type-2 diabetes mellitus. PMID- 11273524 TI - Ciprofloxacin in the management of soft tissue infections in diabetes mellitus. PMID- 11273525 TI - Newer antimalarials. PMID- 11273526 TI - Hemiparesis secondary to malaria. PMID- 11273527 TI - Severe hyponatremia in a chronic schizophrenic patient. PMID- 11273528 TI - Pancreatic tuberculosis. PMID- 11273529 TI - Brainstem haemorrhage simulating transient ischemic attack. PMID- 11273530 TI - Rifampicin resistance in tuberculosis--are we back to the future? PMID- 11273531 TI - Characterization by single strand conformation polymorphism of mutations in the rpoB gene of rifampin-resistant Mycobacterium tuberculosis in strains from Vancouver, Mexico City and New Delhi. AB - OBJECTIVE: The study was conducted to evaluate usefulness of single strand conformation polymorphism (SSCP) over DNA sequencing in the diagnosis of rifampicin (Rif)-resistant tuberculosis. METHODS: Forty seven isolates of Mycobacterium tuberculosis (MTB) Rif-resistant and 25 Rif-sensitive were obtained from Vancouver, Mexico city and New Delhi and were analyzed by polymerase chain reaction (PCR) amplification of rpoB gene and the mutations were identified by DNA sequencing and SSCP. RESULTS: The mutations observed by DNA sequencing in 47 RIF-resistant isolates showed that the most common mutation among Vancouver isolates was in codon 526, Hist-->Arg and in Mexico isolates was in codon 531, Ser-Leu and New Delhi isolates was in codon 516, Asp-->Val. Using fluorescence based PCR-SSCP, it was possible to distinguish Rif-resistant isolates from Rif sensitive isolates. CONCLUSION: DNA sequencing is a highly accurate method for the detection of mutations associated with drug resistance in tuberculosis but is more expensive and requires special equipment and personnel. SSCP is a simple, accurate method and suitable for analysis of large number of samples and the results are available in less than 72 hours. PMID- 11273532 TI - Seizures in patients with human immunodeficiency virus infection. AB - OBJECTIVE: To study the significance of new onset seizure in patient with human immunodeficiency virus (HIV) infection. METHODS: Patients infected with HIV with the new onset seizure were enrolled in the study. Seizure type was classified. Adequate work up was done to search for a cause of their initial seizure. All patients were administered antiepileptic drugs in addition those detected to have treatable opportunistic infection were treated for the same. RESULTS: A total of 455 patients of HIV infection were admitted to this centre during study period, of these twenty three patients had new onset seizures. Seizures were generalized tonic-clonic in fifteen patients (65.21%), partial motor in six patients (26.08%) and partial motor with secondary generalization in two patients (8.69%). Recurrence of seizures was observed in 69.56% patients. Identified causes included cerebral toxoplasmosis in seven patients (30.43%), cryptococcal meningitis in four (17.39%), tuberculoma in three (13.04%), AIDS dementia complex in one (4.34%) and progressive multifocal leucoencephalopathy in one (4.34%). In seven patients (30.43%) seizures were not associated with any identifiable cause. Phenytoin was used for control of seizures and no side effects to the drug were noted during the brief period of follow up. CONCLUSION: Majority of patients with HIV infection and new onset seizures have secondary brain lesion as the cause of seizure. High rate of seizure recurrence mandates therapy of solitary seizure in patients with HIV infection. PMID- 11273533 TI - Oxidative stress after acute myocardial infarction: effect of thrombolytic treatment. AB - OBJECTIVE: Treatment with thrombolysis plays a crucial role in salvaging the myocardium in myocardial infarction (MI) patients, but reperfusion of ischaemic areas may itself be associated with reperfusion injury mediated by free radical induced oxidation. Hence the present study was planned to evaluate oxidative stress in patients receiving thrombolytic therapy during MI and to compare them with those not receiving thrombolysis. METHODS: Thiobarbituric acid reactive substance (TBARS) was used as a marker of lipid peroxidation in 30 patients after acute MI. Thirteen were treated by intravenous thrombolysis and 17 served as control. Also, vitamin E levels were estimated in these patients. RESULTS: Patients treated with thrombolysis showed a fall in vitamin E and increase in TBARS within first hours. The decrease in vitamin E was independent of a change in cholesterol. However, the levels were similar at 72 hours. CONCLUSION: The results indicate increased free radical production after MI and reperfusion also increases in free radical production and antioxidants may have a part in improving thrombolytic reperfusion of ischaemic myocardium. PMID- 11273535 TI - Poor diagnostic accuracy and applicability of Siriraj stroke score, Allen score and their combination in differentiating acute haemorrhagic and thrombotic stroke. AB - OBJECTIVE: To evaluate the relevance of bed side clinical diagnostic scoring systems--Siriraj stroke score (SSS), Allen score and their combined use for differentiating acute haemorrhagic and thrombotic stroke. MATERIAL AND METHODS: The study was conducted on 240 admitted patients of stroke over a period of two years. SSS was calculated immediately and Allen score, 24 hours after admission. CT scan was done immediately and 48 hours after admission if required. The sensitivity, specificity, positive predictive value, negative predictive value and diagnostic gain were calculated for both the scores. Comparability between the scores and CT scan finding was determined with the help of kappa statistic programme. Receiver operating characteristic curves (ROC) were plotted to assess the diagnostic accuracy of both scores over a range of cut-off points. RESULTS: One hundred and thirty four patients (55.83%) had infarction and 106 patients (44.17%) had haemorrhage. SSS was applicable in 66.25% patients (159 out of 240) while Allen score was applicable in only 61.25% patients (147 our of 240). The sensitivity, specificity, positive predictive value, negative predictive value and diagnostic gain for SSS were 73%, 85%, 85%, 71% and 30% for infarction and 85%, 73%, 71%, 85% and 27% for haemorrhage respectively, whereas the corresponding figures for Allen score were 91%, 60%, 77%, 82% and 18% for infarction and 60%, 91%, 82%, 77% and 41% for haemorrhage respectively. There was overall moderate comparability between SSS and Allen score for diagnosing supratentorial stroke (k = 0.396). The comparability of these scores in terms of certain results was worse (k = 0.143). However when the results that were within the diagnostic range with both the scores were considered, the agreement in diagnosing infarction and haemorrhage was almost perfect (k = 0.874). While considering CT scan finding as gold standard for differentiation of infarction and haemorrhage, the overall accuracy of SSS and Allen score was seventy eight percent. CONCLUSION: (a) Applicability of SSS only in 66.25% patients and wrong diagnosis in 22.01% patients does not reflect its usefulness because adequate management of stroke requires a gold standard diagnosis which is only possible by immediate CT scan. (b) Allen score is not useful because it can be assessed only after 24 hours of onset of stroke. This deprives the management to all thrombotic patients in speculated time window of modern management. PMID- 11273534 TI - Effects of cyclo-oxygenase and thromboxane synthetase inhibitors on right atrial prostaglandins. AB - OBJECTIVES: The aim of this study was to find out the effects of cyclo-oxygenase and thromboxane synthetase inhibitors on right atrial prostacyclin and thromboxane A2 levels. METHODS: The study consisted of a total of 50 patients subjected to coronary bypass surgery. These patients were divided into two groups, Group I and Group II each consisting of 25 patients. In Group I patients, the right atrial tissues were studied for effects of indomethacin and U63557A on the prostaglandin levels. In Group II patients, the right atrial tissues were studied for effects of Aspirin and U63557A on the prostaglandin levels. RESULTS: In Group I patients, the atrial tissues pretreated with indomethacin showed a fall in the levels of 6 keto PGF1 alpha from 153.5 +/- 28.4 pg/0.1 mg to 59.7 +/- 11.6 pg/0.1 mg and of TXB2 from 41.6 +/- 1.2 pg/0.1 mg to 17.2 +/- 3.2 pg/0.1 mg. In the atrial tissues of Group I treated with U63557A the levels of 6 keto PGF1 alpha fell to 145.4 +/- 26.8 pg/0.1 mg and the levels of TXB2 fell to 14.7 +/- 2.8 pg/0.1 mg. In Group II patients, the atrial tissues pretreated with aspirin, showed a fall in the levels of 6 keto PGF1 alpha from 142.1 +/- 2.8 pg/0.1 mg to 17.5 +/- 0.8 pg/0.1 mg. In the atrial tissues pretreated with U63557A, the levels of 6 keto PGF1 alpha fell to 131.2 +/- 2.9 pg/0.1 mg and the levels of TXB2 fell to 14.4 +/- 0.7 pg/0.1 mg. CONCLUSIONS: The study showed that human right atrial tissues are capable of producing TXA2 in addition to prostacyclin. Indomethacin and aspirin by inhibiting generation of cyclic endoperoxides inhibited synthesis of both prostacyclin and TXA2. In contrast a thromboxane synthethase inhibitor U63557A selectively inhibited TXA2 without significant effects on prostacyclin synthesis. PMID- 11273536 TI - Stressful life-events, anxiety, depression and coping in patients of irritable bowel syndrome. AB - A lot of research has pointed to a complex interaction between stressful life events, psychiatric morbidity and the irritable bowel syndrome (IBS). AIM: The present study aimed to determine the: stressful life-events in patients with irritable bowel syndrome patients in comparison to normal controls; effect of these events in causing clinically significant anxiety and depression; and the effect of psychopathology i.e. anxiety and depression on coping skills in these patients. METHODOLOGY: Thirty patients with irritable bowel syndrome were compared with thirty matched normal controls, on the presumptive stressful life events scale and the mechanisms of coping scale. Further, among the irritable bowel syndrome patient group, the anxious and depressed subgroups were separated from the non-anxious and non-depressed subgroups using the hospital anxiety and depression scale. Positive and negative coping mechanisms between these subgroups were compared. RESULTS: Significantly higher stress scores were found in the irritable bowel syndrome patient group than normal controls. Not all, but slightly more than fifty percent of irritable bowel syndrome had definite and clinically significant anxiety and/or depression. Those IBS patients with either definite depression tended to use predominantly negative coping styles as compared to those IBS patients without anxiety or depression. CONCLUSION: Stressful life-event scores are significantly higher in IBS patients than in normal controls. Although not all of these patients have anxiety and/or depression, a significant number of patients show evidence of the same. Presence or absence of anxiety and/or depression influences how the patient with IBS copes with illness. Therefore, though further studies on the issue are required, we suggested that, as a supplement to medical management, recognition and treatment of anxiety and depression in this subgroup of IBS patients with psychotropic drugs and cognitive therapy for gaining more positive coping skills, may require special attention in the management of irritable bowel syndrome. PMID- 11273537 TI - Spectrum of renal diseases in Indian adults. AB - INTRODUCTION: Inspite of nephrology as a specialty since seventies, there is still paucity of data regarding the spectrum of renal diseases in India. Available literature from few hospitals shows data on specific clinical syndrome of renal diseases or specific renal diseases rather than the overall spectrum as a whole. This information will be useful for better resource management. MATERIAL AND METHODS: We studied spectrum of renal diseases among 14,796 patients presenting for the first time to nephrology outpatients between January 1987 to Oct. 1998. Majority of patients in our clinic were adults. Patients 14 years or below who mostly attend pediatric renal unit of the hospital were excluded from the analysis. Till 1991, the study was retrospective but after 1991, patients were followed prospectively. Patients were grouped according to classical renal syndrome. After the initial presentation, patients were followed subsequently till their last follow-up in the clinic or till the time of reporting the present data. RESULTS: Mean age of patients was 38.69 +/- 15.5 years with male predominance in majority of presentations. Chronic renal failure (CRF), nephrotic syndrome (NS), nephritic syndrome and hypertension were the four common presentations seen in 47.8%, 15.03%, 4.6% and 4.9% cases respectively. Other presentations were acute renal failure (1.9%), urinary tract infection (2.9%), stone disease (4.6%), obstructive uropathy (2.1%), isolated haematuria (1.2%) and asymptomatic urinary abnormalities (0.3%). Chronic glomerulonephritis was seen in 49.4% cases of CRF followed by diabetic nephropathy in 28.4% cases. Of the nephrotic syndrome cases, primary glomerulonephritis was seen 58.5% cases, of which minimal change disease was the commonest cause in 38% cases. Of the secondary glomerular diseases, diabetic nephropathy was commonest cause of NS (53%) followed by amyloidosis (16.4%) and lupus (8.3%). Tuberculosis was the commonest cause of renal amyloidosis seen in 50% cases. Of the nephritic syndrome, post-infective glomerulonephritis was commonest cause followed by rapidly progressive glomerulonephritis being the second commonest cause. In the hypertensive group, essential hypertension was the commonest cause followed by renovascular hypertension. CONCLUSION: It is the first large study of its kind presenting the spectrum of renal diseases in the tertiary-care government hospital of the country and we expect the disease pattern to be reasonably similar in other similar government hospital of the country. Chronic renal failure, nephrotic syndrome and diabetes are three major diseases, with which we have to deal maximum. As CRF in young male patients is the largest load, with its wide social and economical implications in the Indian context, we must gear up to organise ourselves for providing the best possible care to these patients with the limited resources. PMID- 11273539 TI - Diabetic autonomic neuropathy causing gall bladder dysfunction. AB - OBJECTIVE: The objective of the study was to study gall bladder volume in fasting and 45 minutes post-prandial, by real time ultrasound in healthy controls and diabetic patients with and without autonomic neuropathy and to compare them. METHOD: Age, Sex and body mass index (BMI) matched 50 healthy subjects and 10 patients with insulin dependent diabetes mellitus and 40 patients with noninsulin dependent diabetes mellitus were evaluated according to National diabetes Data Group of National Institute of Health (1979) criteria: 1. Fasting (overnight) venous plasma glucose concentration of > 140 mg/dl on two separate occasions. 2. Following ingestion of 75 gms of glucose, venous plasma glucose concentration of > 200 mg/dl at second hour and at one other occasion during two hour test. Autonomic neuropathy was assessed by the presence of symptoms like dysphagia, abdominal fullness, nausea, vomiting, diarrhea +/- nocturnal, faecal incontinence or constipation, dysuria, urinary incontinence, the gustatory sweating, impotence etc. and were confirmed by standing test for orthostatic hypotension, hand grip test, Valsalva test and deep breaths test. RESULT: The study showed that: 1. Patients of diabetes mellitus had statistically significant larger fasting gall bladder volumes and these values were highly significant amongst patients with autonomic neuropathy. 2. Patients of diabetes mellitus and statistically significant larger post fatty meal gall bladder volume and these values were highly significant in patients with autonomic neuropathy. CONCLUSIONS: We therefore conclude that impaired gall bladder contraction was found amongst patients of diabetes mellitus with autonomic neuropathy. The mechanism responsible for cholecystoparesis is attributed to vagal neuropathy. Incomplete gall bladder emptying leads to sequestration of cholesterol and nidus formation. Therefore gall bladder functions should be evaluated routinely in such patients and early intervention is recommended. PMID- 11273538 TI - Effect of intraesophageal acid instillation on airway reactivity in patients with asthma. AB - OBJECTIVE: To study the change in airway reactivity due to presence of acid in lower esophagus and its reversibility by antacid. METHOD: In this double blind study 12 subjects with asthma and gastroesophageal reflux received acid (N/10 hydrochloric acid) and antacid (mixture of magnesium trisilicate and aluminum hydroxide) perfusion in lower esophagus via a nasogastric tube. The four combinations were antacid-antacid (control), antacid-acid, acid-antacid and acid acid. Airway reactivity (Histamine PD20) was recorded after each perfusion. RESULTS: Histamine PD20 significantly decreased (airway reactivity increased) (p < 0.05) with all three combinations containing acid as compared to control. No significant difference in airway reactivity was observed if the antacid was given before or after the acid. CONCLUSION: Presence of acid in lower esophagus can increase airway reactivity. This effect lasts longer than the presence of acid in esophagus itself. PMID- 11273540 TI - Comparative bioavailability of two formulations of azithromycin. AB - AIM: To compare the bioequivalence of two brands of azithromycin capsules in healthy male volunteers for regulatory purpose. METHOD: A single oral dose of 500 mg of either test (Panacea Biotec Ltd.) or reference (Pfizer India Ltd.) preparation of azithromycin was administered to 12 volunteers in double blind randomised cross over fashion. Serum levels of azithromycin were analysed using microbiological assay. The pharmacokinetic parameters studied were Cmax, Tmax, AUC, t1/2, Ke, CL and MRT. In vitro dissolution tests were conducted for both the preparations and compared with in vivo absorption. RESULTS: The mean peak serum azithromycin concentration of 0.516 +/- 0.008 microgram/ml was observed at 2.33 +/- 0.22 h with test brand and was similar to that of reference brand with Cmax of 0.494 +/- 0.011 microgram/ml at 2.71 +/- 0.26 h. The statistical difference between all the other paharmacokinetic parameters were insignificant. CONCLUSION: Both the brands of azithromycin can be considered to be bioequivalent on the basis of results obtained. PMID- 11273541 TI - pictorial CME. Lingual goitre. PMID- 11273542 TI - Tuberculosis and acquired immunodeficiency syndrome. AB - India has the largest HIV-infected population in the world with over 4 million infected. The current evidence indicates that the doubling time of the epidemic in India is less than 2 years. Those infected with Mycobacterium tuberculosis add to the gravity of the situation enhancing both morbidity and mortality in these dually infected patients. With the incomprehensive diagnostic facilities available, it is wise to exploit the situation with utilization of tuberculin test with a different cut off criterion for the early diagnosis of mycobacterial infection amongst HIV positive population, especially when there are financial constraints and lack of diagnostic facilities to get CD4 cell count. PMID- 11273543 TI - Chronobiology and chronotherapy: current perspectives. AB - To summarise, one can say that with the time to come, chronotherapy might become the order of the day in treatment of many systemic diseases whether they are cardiovascular, respiratory, infectious etc. This approach will certainly help in better control of signs and symptoms while simultaneously protecting the individual from untoward side effects and providing them with a better quality of life. PMID- 11273544 TI - A spectrum of dystonias-clinical features and update on management. AB - Dystonia is an interesting disorder characterized by involuntary movement of the body part or parts leading to abnormal deformed postures. The usual signs and symptoms are local pain, spasm and abnormal movements. Sensory trick is an important clinical phenomenon and is characteristic of dystonia. It is usually separated from other movement disorders such as chorea, athetosis, tics and myoclonus clinically. Various non-dystonic conditions simulate dystonia and need to be separated in view of different line of management. Improved understanding in molecular biology has helped in understanding of the disease. Confusing neuropathology and neurochemistry have deterred the finding of an effective drug, however empirical use of few drugs have improved the gloomy situation. Few conditions such as dopa-responsive dystonia have definite treatment. Recently use of botulinum toxin has provided beneficial response in hyper muscular contraction states such as dystonia and spasticity, Surgery and other non-medical therapies are effective in few situations. PMID- 11273545 TI - Medical text books in India. PMID- 11273546 TI - Need for bringing in a change in biochemistry curriculum to make it clinically oriented? AB - OBJECTIVES: This study was conducted to (a) assess the views of medical students and doctors regarding relevance of biochemistry training, (b) explore if they have any suggestion to bring in any improvement in contents of biochemistry curriculum and mode of teaching. METHODS: In 1997-98, a structured questionnaire was filled up by 114 medical students and 118 doctors. RESULTS: As many as 62/114 (55%) medical students and 40/118 (34%) doctors believed that it is not important to remember minute details of biochemical reactions (p value < 0.0001). Among medical students, 108/110 (98.2%) agree that a clinician should be invited to seminars for developing skills of interpretation of laboratory investigations; whilst 110/118 (93.2%) doctors expressed similar view, p value ns. Approximately 92% responders favored that departments biochemistry and physiology should co ordinate on the topics of common interest in order to save time and effort. What is the most informative and effective way of teaching biochemistry?' in response to this question only 0.9% responders opted lecture as the best option. Seminars with active participation of medical students was preferred by 93.2% responders. About 6.9% responders reckoned that symposium prepared by a more than one teacher. In response to the question whether it is possible to cover pre-clinical subjects in 12 months so as to allow spiral mode of curriculum, 73% of all the responders agreed that it would be good idea, there was no difference of opinion among the doctors and medical students. On the other hand, 27% were strongly opposed to this suggestion. CONCLUSIONS: We suggest that there is a need to modify the contents, methods of teaching, and curriculum organization of training in clinical biochemistry. How best the curriculum can be made problem oriented needs to be debated among medical educationists. PMID- 11273547 TI - Undifferentiated leukemia following treatment of gastric cancer with 5 fluorouracil, doxorubicin, mitomycin C chemotherapy. AB - Secondary leukemias are an increasingly well recognized complication of cancer chemotherapy. We report on the development of undifferentiated leukemia in a forty one year lady, eighteen months after treatment of gastric cancer with 5 fluorouracil (5 FU), doxorubicin and mitomycin C. She developed progressive weakness and pancytopenia over a period of few months. Leukemic blasts which were undifferentiated by immunophenotyping were seen in her peripheral blood. She died of intracranial haemorrhage during induction chemotherapy for the leukemia. Undifferentiated leukemia is a rare complication of FAM chemotherapy for gastric cancer. PMID- 11273548 TI - Chilaiditi syndrome with hypertension. AB - Chilaiditi syndrome is interposition of the intestine between liver and diaphragm. It is often asymptomatic but there were cases presented as acute pain in the abdomen, needing corrective surgical procedure; or as mistaken renal colic, or as suspected subphrenic abscess, or as pneumoperitonium. The interposition of proximal transverse colon was found to be more common than the small intestine. Chilaiditi syndrome was associated with colonic volvulus. The colonic interposition then progressed from mild abdominal discomfort to intermittent bowel obstruction. Some patients needed surgical operation like hepatic extraperitonealization, after replacement of the dislocated gastroenteric tract, bringing the superior surface of the liver again into direct contact with the related diaphragmatic dome. A rare case of Chilaiditi syndrome incidentally associated with hypertension and ischaemic heart disease, in a male aged 50 years is reported. PMID- 11273549 TI - Recurrent haematuria in coexisting IgA nephropathy and interstitial nephritis. AB - We report here a case of recurrent hematuria in a young man who presented with deteriorating renal function due to interstitial nephritis, secondary to, probably enteric fever. Immunofluorescence studies showed IgA nephropathy and ultrastructural studies revealed thin basement membrane nephropathy. PMID- 11273551 TI - Ciprofloxacin-induced severe cutaneous reaction and haemolysis in a young adult. PMID- 11273550 TI - Ofloxacin induced arthropathy in patients with multi-drug resistance tuberculosis. AB - Some of the 4-fluroquinolones have been reported to induce arthropathy, but so far there are no case reports of arthropathy due to ofloxacin. Two cases of ofloxacin induced arthropathy in multi-drug resistance (MDR) tuberculosis are reported and possible drug interaction with other anti-tubercular drugs is discussed. PMID- 11273552 TI - Primary hypothyroidism presenting as a pituitary gland enlargement with regression following thyroxine therapy. PMID- 11273553 TI - Multiple myeloma with cutaneous dissemination. PMID- 11273554 TI - Idiopathic polymyositis presenting as interstitial lung disease. PMID- 11273555 TI - Tuberculosis: a rare cause of splenic abscess. PMID- 11273556 TI - Blind conjunctival biopsy as an aid to the diagnosis of sarcoidosis. PMID- 11273557 TI - Acute intermittent porphyria: a cause of posterior leukoencephalopathy syndrome. PMID- 11273558 TI - Stroke like presentation of Creutzfeldt-Jakob disease. PMID- 11273559 TI - Essential secondary and tertiary care to non-communicable diseases. PMID- 11273560 TI - A survey of primaquine prescribing habits in the city of Mumbai. PMID- 11273561 TI - Bone remodeling on the Web. PMID- 11273562 TI - The Yin and Yang of nuclear receptors: symposium on nuclear receptors in brain, Oegstgeest, The Netherlands, 13-14 April 2000. AB - Novel aspects of nuclear receptors and their function in brain were discussed at a recent Symposium in Oegstgeest, The Netherlands. Presentations covered the diversity of these receptors, their target genes, proteins involved in transcriptional regulation, functional consequences of nuclear receptor activation and their relevance for human pathology. By elucidating the signalling pathway of nuclear receptors in brain, potential targets for therapeutic treatment of brain disorders can be identified. PMID- 11273563 TI - Insights into the role of estrogen in the male genital tract: a report on an estrogen and male reproduction workshop, Isola Capo Rizzuto, Italy, 23-24 September 1999. AB - The first International Workshop on estrogen and male reproduction was held in Isola Capo Rizzuto, Italy and was organized by the University of Calabria (COSENZA), the University of Naples and the University of Modena and Reggio Emilia. The workshop, which was attended by general scientists, endocrinologists and andrologists, addressed the impressive recent progress in this area of male reproduction. Owing to space limitations, this report focuses on only a few of the recent advances related to the role of estrogen in the male genital tract. PMID- 11273564 TI - Hunting out rare diseases on the Web. PMID- 11273566 TI - Gene imprinting on the Web. PMID- 11273567 TI - The Second International Symposium on the Developmental Aspects of Androgen Excess, Toronto, Canada, 20 June 2000. AB - Clinical hyperandrogenism, in particular polycystic ovary syndrome (PCOS), affects 4-7% of women of reproductive age, making it one of the most common human reproductive endocrinological abnormalities. However, our understanding of the developmental aspects of these disorders remains limited. The Second International Symposium on the Developmental Aspects of Androgen Excess (Toronto, Canada, 20 June 2000) was held with the purpose of fostering greater investigative communication, consensus and focus. It was felt that a better understanding of PCOS phenotypes was needed; that an aggressive attempt should be made to continue to expand the molecular genetic studies of the disorder; that research into the role and mechanism(s) underlying the associated defects in insulin action and signaling should be continued; that longitudinal studies, particularly those focusing on the role of intrauterine stress and malnutrition, and premature adrenarche, on the development of PCOS were warranted; and that an improved understanding of the molecular defects in steroidogenesis present in PCOS is needed. PMID- 11273569 TI - Diabetes on the Web. PMID- 11273565 TI - Should we watch what we eat and drink? Report on the International Workshop on Hormones and Endocrine Disrupters in Food and Water: possible impact on human health, Copenhagen, Denmark, 27-30 May 2000. AB - The aim of this Workshop was to bring together scientists with different backgrounds, including clinical endocrinologists, basic researchers and epidemiologists, to discuss the complex and controversial topic of endocrine disrupters, and their impact on human health. Nearly 250 scientists attended the Workshop, and 50 lectures and 90 posters were presented and discussed. The most important scientific findings and reviews are to be published in Human Reproduction and Human Reproduction Update. Some of the highlights are presented here. PMID- 11273568 TI - Endocrinology and pathological images on the Web. PMID- 11273570 TI - Protective role of cerebrospinal fluid in brain injuries. PMID- 11273571 TI - By the way, doctor... I work in a store and frequently lift boxes that are pretty heavy. One of my friends wears one of those thick leather back belts and swears that the extra support helps her back. My doctor doesn't have any opinion on whether these belts are a good idea. Do you? PMID- 11273573 TI - Basic Science. PMID- 11273574 TI - Measuring the negative mood component of stress experiences: description and psychometric properties of a short adjective check-list of stress responses. AB - Negative Mood (NM) is a 19-items adjective check-list developed to assess negative mood and stress responses. The items of the scale reflect dimensions such as depressed mood, anxiety, anger and time urgency. The data were collected from four different samples, two random population samples and two smaller selective samples. The psychometric properties of the NM showed high internal consistency (Cronbach's Alpha). When analysed with principal components analysis, three factors emerged, anxiety/depression, time pressure, and anger. The factors emerged in all four samples, within samples and between gender. The NM factors were tested for convergent and discriminative validity by correlating them with other more established measures of different aspects of negative mood. The results showed high convergent and discriminative validity for two of the NM factors, i.e., anxiety/depression, and anger, whereas the results for the third factor, time pressure, were more ambiguous. This scale has proven to be useful in capturing some vital dimensions of negative affect across different kinds of populations. PMID- 11273575 TI - Improving subjective health and reducing absenteeism in a natural work life intervention. AB - A natural one-year work-life intervention to improve occupational health and reduce absenteeism was designed as a field experiment. The intervention allowed the employees in the health care sector of a municipality to take up to five days of self-administered sick leave with full financial compensation up to four times a year. 165 employees in the intervention group and 100 employees in the control group filled out a questionnaire before and after the intervention. The result showed no evidence of misuse of this sick-leave option and some positive subjective health effects were found among those who used the option. Slight improvements were found in musculoskeletal problems and for cold/influenza. There were no effects on overall absenteeism. The question of the impact of local cultures on interventions to improve occupational health is also discussed. PMID- 11273576 TI - Dieting behavior in Norwegian adolescents. AB - In a sample of Norwegian adolescents (n = 1117) 27, 4% of the girls and 9% of the boys reported that they were dieting. Using multiple regression analyses, we were able to predict 50% of girls dieting behavior and 24% of boys dieting behavior. Psychological concerns relating to weight and eating (WEC) as well as perceptions of feeling fatter than others were the only significant predictors of dieting in girls, whilst dieting in boys could also be predicted by their Body Mass Index. Dieting girls who scored high on the Weight and Eating Concerns Inventory were found to suffer constipation and binge eating attacks more often than other dieting girls. This was taken as an indication that these girls were at risk of developing an eating disorder. PMID- 11273577 TI - Consumer satisfaction and attributions of improvement among fully recovered schizophrenics. AB - As people with serious mental illness are viewed as consumers rather than patients, their views and needs are increasingly seen as essential to the treatment process. The aim of the present study is to add valuable knowledge about how to help patients by reporting what fully recovered schizophrenics, as consumers, had found helpful in their treatment as well as which factors contributed to their recovery. In addition to a semistructured interview, two instruments were used: the "UCLA Social Attainment Survey Premorbid Adjustment Scale" (SASPAS) to measure patients' premorbid adjustment and the "Global Assessment Scale" (GAS) to obtain an assessment of the present general functioning of the subjects. The results showed that a majority of the recovered subjects emphasized their own will-power and the human qualities in their therapists as helpful to them in the process towards recovery. The therapeutic relationships were characterized by the kindling and sustaining of hope in the clients. PMID- 11273578 TI - Psychometric critique of acculturation psychology: the case of Iranian migrants in Norway. AB - The presumptions, terminology, psychometrics, statistical analyses, and ethics of the fourfold acculturation paradigm are criticized in detail. Illustrative data came from Iranian refugees in Norway (N = 80) answering: 1) the Satisfaction with Life Scale (SWLS), 2) Zung's Self-Rating Depression Scale (ZSRDS), 3) ipsative fourfold scales of Integration, Assimilation, Separation, and Marginalization, 4) orthogonal scales of attitudes towards Norwegian and Iranian cultures, measured independently and using balanced reverse-keying, and 5) ipsative forced-choice preferences for cultural practices of Norway, Iran, both, or from other societies as well. Iranians in Norway favored global multiculturalism and, as a group. did not show distress. The SWLS and ZSRDS were correlated, but the measures of acculturation failed to replicate one another. As unconstrained ipsative measures, the fourfold scales showed acquiescence response bias contamination and doubtful operationalization of scale constructs. Recommendations are discussed for improving acculturation research. PMID- 11273579 TI - Negative emotions and coronary heart disease: causally related or merely coexistent? A review. AB - Negative emotions have been claimed to be a cause of coronary heart disease (CHD) as well as a consequence of cardiovascular disorders. Early case studies of cardiac disorders of soldiers in battle drew attention to the possibility that strong negative emotional states could cause CHD. Subsequent reports of reactions to natural disasters supported the notion that intense negative emotions could precipitate somatic disorders such as CHD. Since then, numerous studies have investigated relations between negative emotions and CHD. Over the years, retrospective studies have found, for example, that negative emotions are often present before the occurrence of CHD. Cross-sectional studies have indicated that symptoms of depression and anxiety are often present in CHD patients. Prospective studies have shown that the likelihood of CHD tends to be higher for people with negative emotions than for those without them. The main symptoms of negative emotional states that seem to be most closely associated with CHD are nervousness, getting easily upset, feeling fatigue, being indecisive, having sleep disturbances, being usually worried about something, and feeling that others would be better off if oneself were dead. Although the findings appear to support the notion of causal connections between negative emotions and CHD, they fail to provide conclusive proof of such relations. An alternative explanation that could also account for the findings is simply that negative emotions and CHD often coexist. PMID- 11273580 TI - Colour-cueing in visual search. AB - Several studies have shown that people can selectively attend to stimulus colour, e.g., in visual search, and that preknowledge of a target colour can improve response speed/accuracy. The purpose was to use a form-identification task to determine whether valid colour precues can produce benefits and invalid cues costs. The subject had to identify the orientation of a "T"-shaped element in a ring of randomly-oriented "L"s when either two or four of the elements were differently coloured. Contrary to Moore and Egeth's (1998) recent findings, colour-based attention did affect performance under data-limited conditions: Colour cues produced benefits when processing load was high; when the load was reduced, they incurred only costs. Surprisingly, a valid colour cue succeeded in improving performance in the high-load condition even when its validity was reduced to the chance level. Overall, the results suggest that knowledge of a target colour does not facilitate the processing of the target, but makes it possible to prioritize it. PMID- 11273581 TI - Measuring adult attachment: a construct validation of two self-report instruments. AB - This study reports the Swedish construct validation of two translated attachment style scales. The factor structure of the attachment construct was investigated via exploratory and confirmatory factor analyses of attachment scores from 515 students of a Swedish university. Results supported the expected two-factor solution, but found a three-factor solution to be a viable alternative. In addition, the attachment scales were compared with the Big Five personality inventory (NEO-PI), using a sample of 87 Swedish students, and found to have expected correlation to this scale. PMID- 11273582 TI - Dimensionality of psychosocial responses to marital disruption. AB - The purpose of the study was to assess whether psychosocial responses to marital disruption were best arranged along one single dimension ranging from maximum positive to maximum negative responses, or whether positive and negative responses constituted two separate and distinct dimensions. Participants were 658 recently divorced individuals. A confirmatory factor analysis revealed that a two factor model provided a better fit to the data than a one-factor model. Moreover, a number of external variables differentiated between positive and negative responses, indicating that such responses reflect two separate domains. Thus, the findings support the notion that psychosocial responses after marital disruption comprise a bidimensional rather than a unidimensional pattern. The findings are discussed within the general framework of positive-negative asymmetry and the idea that positive and negative evaluations belong to different psychological system. PMID- 11273583 TI - Fourth International Conference on Colorectal Cancer: adjuvant treatment of colon cancer--introduction. PMID- 11273584 TI - Efficacy of adjuvant fluorouracil and leucovorin in stage B2 and C colon cancer. International Multicenter Pooled Analysis of Colon Cancer Trials Investigators. AB - The International Multicenter Pooled Analysis of Colon Cancer Trials (IMPACT) investigators have now completed two large systematic reviews of adjuvant therapy trials in colon cancer. The IMPACT 1 study pooled data from three separate trials each comparing the efficacy of 5-fluorouracil (5-FU)/leucovorin with observation alone as adjuvant treatment for 1,526 patients with Dukes' B or C colon cancer. The results showed that treatment with 5-FU/leucovorin significantly reduced mortality by 22% (P = .029) and events such as relapse, second tumor, or death by 35% (P < .0001) after 3 years of follow-up. The side effects associated with 5 FU/leucovorin were clinically acceptable. The IMPACT 1 study also showed a clear benefit of adjuvant treatment for patients with Dukes' C colon cancer, but not for stage-B patients. After up to 10 years of follow-up, 5-FU/leucovorin significantly reduced mortality by 30% for patients with Dukes' C disease (P = .003), but only reduced mortality by 8% in patients with Dukes' B colon cancer (P = .658). The aim of the IMPACT 2 study was to determine whether 5-FU/leucovorin is an effective adjuvant treatment for patients with Dukes' B2 colon cancer. Results were pooled from five separate trials that randomized 1,016 patients. After a median of 5.75 years of follow-up, B2 patients receiving 5-FU/leucovorin did not have a significant increase in overall survival or event-free survival. At 5 years, the hazard ratio for overall survival was 0.86 (90% confidence interval, 0.68 to 1.07) and for event-free survival was 0.83 (90% confidence interval, 0.72 to 1.07). 5-Fluorouracil/leucovorin was not recommended as a standard adjuvant treatment for all patients with Dukes' B2 colon cancer. PMID- 11273585 TI - Should Dukes' B patients receive adjuvant therapy? A statistical perspective. AB - The benefit of adjuvant therapy, such as 5-fluorouracil (5-FU) combined with leucovorin, is a matter of debate for patients with Dukes' B colon cancer. Several approaches have been taken to address this issue. Initially, studies were conducted to assess treatment benefits in both Dukes' B and Dukes' C patients. These studies identified an overall benefit of adjuvant treatment and enrolled enough Dukes' C patients to determine a treatment benefit for adjuvant 5 FU/leucovorin in this subpopulation. However, the individual studies were insufficiently powered to detect a treatment benefit in Dukes' B patients. An analysis of four separate studies (National Surgical Adjuvant Breast and Bowel project) compared the benefit of adjuvant treatment in Dukes' B patients with that in Dukes' C patients and showed similar relative reductions in mortality and disease-free survival in Dukes' B and in Dukes' C patients. The Liver Infusion Meta-Analysis Group also reported similar relative benefits from a portal vein infusion of 5-FU-based chemotherapy in Dukes' B and Dukes' C patients. The International Multicenter Pooled Analysis of Colon Cancer Trials B2 study, which combined data from patients with Dukes' B colon cancer in five separate trials, failed to show a statistically significant benefit of adjuvant 5-FU/leucovorin compared with surgery alone. We review the advantages and limitations of different approaches to detect treatment benefits in patients with Dukes' B colon cancer, and we argue that there is a need for a meta-analysis of all adjuvant trials to reliably address this question. PMID- 11273586 TI - Update of clinical trials with edrecolomab: a monoclonal antibody therapy for colorectal cancer. AB - Edrecolomab is a murine IgG2a monoclonal antibody that recognizes the tumor associated antigen Ep-CAM. Its antitumor effects are mediated through antibody dependent cellular cytotoxicity, complement-mediated cytolysis, and the induction of an anti-idiotypic network. An initial study of 189 patients with resected stage III colorectal cancer showed that edrecolomab reduced the relative risk of mortality by 32% compared with observation alone (P < .01). Edrecolomab has now been investigated in two large phase III studies of patients with stage III colon cancer, either as a single agent or in combination with 5-FU-based chemotherapy. Preliminary safety data have shown that edrecolomab is well tolerated when used as monotherapy and adds little to chemotherapy-related side effects when used in combination. Edrecolomab is also being studied as monotherapy following resection of stage II colon cancer, and in combination with chemotherapy in patients with resected stage II or III rectal cancer. In conclusion, edrecolomab is a novel biological therapy for the adjuvant treatment of colorectal cancer. Completed and ongoing trials may support its use as monotherapy in stage II colon cancer or in combination with chemotherapy in stage III colon cancer and stage II/III rectal cancer. PMID- 11273587 TI - A United Kingdom coordinating committee on cancer research study of adjuvant chemotherapy for colorectal cancer: preliminary results. AB - Standard adjuvant chemotherapy for colorectal cancer consists of 5-fluorouracil with leucovorin or levamisole. The large, multicenter, randomized, double-blind QUASAR (Quick and Simple and Reliable) trial investigated whether treatment with a higher dose of leucovorin or the addition of levamisole to 5-fluorouracil and leucovorin improved survival. In the QUASAR study, 4,927 patients with colorectal cancer with no evidence of residual disease following resection, were randomized to receive fluorouracil (370 mg/m2) with high-dose (175 mg) or low-dose (25 mg) leucovorin and either levamisole (50 mg) or placebo. The fluorouracil and leucovorin regimen was given either monthly (as six 5-day courses with 4 weeks between the start of each course) or weekly (as 30 once-weekly doses). Levamisole or placebo was given three times daily for 3 days, repeated every 2 weeks for 12 courses. The primary endpoint was death from any cause. Survival was similar with both high- and low-dose leucovorin (70.1% v 71.0% at 3 years; P = .43) as well as recurrence rates (36.0% v 35.8%; P = .94), and with levamisole compared with placebo (69.4% v 71.5%; P = .06) as well as recurrence rates (37.0% v 34.9%; P = .16). Monthly and weekly treatments were equally effective (although this was a nonrandomized comparison), while weekly treatment was associated with significantly fewer toxic effects (neutropenia, mucositis, and diarrhea). High dose leucovorin was not associated with a survival or recurrence benefit when compared with low-dose leucovorin. The ongoing QUASAR-1 trial aims to establish whether adjuvant chemotherapy has any worthwhile survival benefit in colorectal cancer patients with an uncertain indication following surgical resection. PMID- 11273588 TI - Randomized adjuvant study comparing two schemes of 5-fluorouracil and leucovorin in stage B2 and C colon adenocarcinoma: study design and preliminary safety results. Groupe d'Etude et de Recherche Clinique en Oncologie Radiotherapies. AB - The aim of this randomized open-label study was to compare a bimonthly with a monthly regimen of 5-fluorouracil (5-FU) and leucovorin for the adjuvant treatment of colon and high-rectum adenocarcinoma. The bimonthly regimen was administered for 2 consecutive days every 14 days as d,L-leucovorin 200 mg/m2 or L-leucovorin 100 mg/m2 as a 2-hour infusion followed by 5-FU bolus of 400 mg/m2 and a 600 mg/m2 5-FU 22-hour continuous infusion (LVSFU2). In the monthly regimen, d,L-leucovorin 200 mg/m2 or L-leucovorin 100 mg/m2 15-minute infusion followed by a 400 mg/m2 15 minute 5-FU bolus was administered for 5 consecutive days every 28 days (FUFOL). Nine hundred five patients with recently resected stage B2 or C colon or high-rectum adenocarcinoma (inferior pole of the tumor subperitoneal) were recruited into the study. Patients were randomized in a 2 x 2 factorial design to receive either LV5FU2 or FUFOL for 24 or 36 weeks. Characteristics of the patients in the two different treatment groups were similar at baseline. Compliance was good. Mean 5-FU dose intensities were 930 mg/ m2/wk and 463 mg/m2/wk for LVSFU2 and FUFOL, respectively. The incidence of maximal grade III-IV toxicities for LVSFU2 and FUFOL was neutropenia 6% and 16% (P < .001), diarrhea 4% and 10% (P < .001), and mucositis 2% and 7% (P < .001), respectively. Maximum grade III-IV toxicities in the LV5FU2 treatment group were significantly lower than in the FUFOL group (10% v 26%; P < .001). Although patients in the LV5FU2 group received twice the dose of 5-FU compared with those in the FUFOL group, LV5FU2 was shown to be less toxic. Efficacy data will be available in 2001. PMID- 11273589 TI - North Central Cancer Treatment Group--Mayo Clinic trials in colon cancer. AB - The adjuvant treatment of colon cancer is now accepted as an effective therapy following surgical resection of the primary tumor in patients at high risk for relapse. Results of studies conducted in the last 10 years have confirmed the benefits of using various 5-fluorouracil (5-FU)-based chemotherapy regimens to decrease recurrence rates and improve patient survival. The North Central Cancer Treatment Group-Mayo Clinic studies have made a significant contribution to establishing the role of adjuvant therapy in colon cancer. Our first study to suggest the effectiveness of adjuvant chemotherapy in patients with stage III disease was reported in 1989. The study used a combination of 5-FU and the anthelminthic agent levamisole, tested because of its ability to positively modulate the human cellular immune system. The results of this study were confirmed by a larger Intergroup study (0035) showing significant decreases in relapse and death rates in stage III colon cancer patients treated with 5-FU plus levamisole compared with surgery alone. Clinical trials conducted throughout the 1990s tested various 5-FU based regimens against the standard 5-FU and levamisole combination. From these trials, the combination of 5-FU plus leucovorin emerged as the standard surgical adjuvant treatment of colon cancer. The efficacy of treatment with this regimen for 6 months was similar to that of a 1-year 5-FU plus levamisole regimen. These findings led to the design of trials in which 5-FU was combined with both leucovorin and levamisole. No differences in efficacy were found in any of the various combinations tested. Current clinical trials are investigating the use of newer agents such as CPT-II in the adjuvant setting. The results of these trials may further improve the efficacy of adjuvant therapy in patients with high-risk colon cancer. PMID- 11273590 TI - 5-fluorouracil/leucovorin versus capecitabine in patients with stage III colon cancer. AB - Capecitabine is an orally administered fluoropyrimidine carbamate that is preferentially converted to 5-fluorouracil in tumors relative to normal tissue. Three different dosing regimens were investigated in phase I studies: continuous monotherapy, intermittent monotherapy, and intermittent therapy supplemented with leucovorin. These studies defined the maximum tolerated dose for each regimen. Dose-limiting toxicities were diarrhea, hand-foot syndrome, and leukopenia. Each phase I study recommended a dose; these were directly compared in a phase II study. This phase II study showed that the intermittent regimen of capecitabine monotherapy was the most favorable on the basis of tumor response rates, time to progression, dose intensity, and toxicities. The intermittent regimen of capecitabine has been compared with the Mayo Clinic regimen of 5 fluorouracil/leucovorin in two large, phase III studies of patients with advanced colorectal cancer. The combined data from the two studies showed that capecitabine was superior to 5-fluorouracil/leucovorin in terms of tumor response rate (22% v 13%; P < .0001) and similar in terms of time to disease progression and overall survival. Capecitabine is now being compared with the Mayo Clinic regimen as an adjuvant treatment for patients with Dukes' C colon cancer. PMID- 11273591 TI - Follow-up of stage B and C colorectal cancer in the United States and France. AB - The optimal postoperative follow-up strategy for patients with resected Dukes' Stage B and C colorectal cancer is controversial. Recently published guidelines support a minimal regimen of carcinoembryonic antigen measurements every 2 to 3 months for at least 2 years, history and physical examination every 3 to 6 months for 3 years, then annually, and colonoscopy every 3 to 5 years. Based on documented practice on the part of surgeons, this regimen would be regarded as intensive. Analyses of relapses following adjuvant therapy support an even more aggressive schedule, with the goal of maximizing the proportion of patients who may be operated on with curative intent (currently about 20% of those who relapse). Additional considerations that may influence the approach to such patients include the identification of second primary tumors (2% over 7 years observation), and the known improvement in quality of life and survival associated with early versus delayed initiation of chemotherapy. However, with the annual investment of resources estimated to be as high as 175 million dollars in the United States alone, a systematic study of such interventions is needed to provide support survival, quality of life, and economic evidence. PMID- 11273592 TI - National Surgical Adjuvant Breast and Bowel Project trials in colon cancer. AB - During the last decade, the National Surgical Adjuvant Breast and Bowel Project (NSABP) has completed six adjuvant chemotherapy trials comparing different adjuvant therapy regimens or adjuvant therapy versus surgery alone. A seventh trial is ongoing. These trials have contributed to defining the role of adjuvant therapy in colon cancer. Patients eligible for inclusion in NSABP trials had been diagnosed as having stage II or III colon cancer with no evidence of gross residual or metastatic disease. The follow-up strategies were similar in the reported trials with follow-up every 3 months for the first 2 years, then every 6 months for the next 3 to 5 years, and annually thereafter. The NSABP C-01 protocol was a three-arm trial comparing an adjuvant semustine/vincristine/5 fluorouracil (5-FU) regimen (MOF) to a Bacille Calmette-Guerin treatment, and to surgery alone. The C-02 protocol investigated whether portal vein infusion of 5 FU improved survival outcome compared with surgery alone. Protocol C-03 compared a semustine/vincristine/5-FU regimen to a 5-FU plus leucovorin (LV) (5-FU/LV) regimen. The NSABP C-04 protocol was a three-arm trial comparing 5-FU/LV, 5-FU plus levamisole, and 5-FU/LV plus levamisole. The NSABP C-05 trial compared 5 FU/LV to 5-FU/LV plus alpha-interferon. Results of NSABP C-01, C-02, C-03, C-04, and C-05 trials are summarized in this report. Patient accrual has completed in the NSABP C-06 trial comparing 5-FU/LV with oral tegafur and plus uracil leucovorin. The NSABP is currently conducting another trial (C-07) comparing 5 FU/LV with 5-FU/LV plus oxaliplatin. The role of adjuvant chemotherapy in stage II colon cancer is also discussed in this report. A recent pooled analysis of studies C-01, C-02, C-03, and C-04 has indicated that the relative treatment benefit in stage II disease is at least equal to the benefit in stage III colon cancers, and concluded that adjuvant chemotherapy also should be considered as the standard of care for stage II colon cancer patients. PMID- 11273593 TI - Cholinesterase inhibitors, beta-amyloid precursor protein and amyloid beta peptides in Alzheimer's disease. AB - The extracellular deposition of amyloid beta-peptide (Abeta) in the form of cerebrovascular amyloid and extracellular plaques is one of the major neuropathological manifestations of Alzheimer's disease (AD). Abeta is generated proteolytically from the large beta-amyloid precursor protein (APP). APP is cleaved by a group of proteases called "secretase" to generate soluble derivatives of APP (sAPP), which are secreted in human plasma, CSF and cultured cells. Neurochemically, there is a severe loss of cholinergic neurons and a decreased synthesis of acetylcholine in neocortex in AD. Current approved AD drugs, such as aricept and tacrine, are based on the use of cholinesterase inhibitors (ChEIs) and have been reported to improve memory deficits and cognitive decline in some patients with AD. To compare the effects of ChEIs on APP processing, we have tested a series of ChEIs such as tacrine, physostigmine, metrifonate, phenserine and cymserine in cultured human neuroblastoma cells. We analyzed levels of sAPP by immunochemical techniques with APP-specific antibodies and assayed levels of Abeta by a sensitive sandwich ELISA. Based on these results, ChEIs can be divided into three groups: the first group of ChEIs had no effect on sAPP secretion, the second decreased the sAPP secretion only, and third group affected the secretion of sAPP and Abeta. The difference in the action of metrifonate, physostigmine, phenserine and tacrine on APP processing is independent of their selectivity for the cholinesterase enzymes. This possibly is due to the different targets that are used by ChEIs. Studying the effects of ChEIs on different targets is useful to maximize the benefit of ChEIs for the treatment of AD subjects. PMID- 11273594 TI - Hydration changes for DNA intercalation reactions. AB - The hydration changes that accompany the DNA binding of five intercalators (ethidium, propidium, proflavine, daunomycin, and 7-aminoactinomycin D) were measured by the osmotic stress method with use of the osmolytes betaine, sucrose, and triethylene glycol. Water uptake was found to accompany complex formation for all intercalators except ethidium. The difference in the number of bound water molecules between the complex and the free reactants (Deltan(w)) was different for each intercalator. The values found for Deltan(w) were the following: propidium, +6; daunomycin, +18; proflavine, +30; and 7-aminoactinomycin D, +32. For ethidium binding to DNA a value of Deltan(w) = +0.25(+/-0.3) was found, indicating that within experimental error no water was released or taken up upon complex formation. Intercalation association constants measured in D2O were found to increase relative to values measured in H2O for all compounds except ethidium. A positive correlation between the ratio of binding constants (K(D2O)/K(H2O)) and Deltan(w) was found. These combined studies identify water as an important thermodynamic participant in the formation of certain intercalation complexes. PMID- 11273596 TI - Encapsulation of functional moieties within branched star polymers: effect of chain length and solvent on site isolation. AB - Porphyrin and pyrene photoactive cores have been encapsulated within an isolating polymeric shell using an efficient and general strategy based on the use of dendritic initiators for the ring-opening polymerization of epsilon-caprolactone to yield functional core star polymers. The isolation of the core functionalities has been studied using fluorescence quenching and fluorescence resonance energy transfer (FRET) techniques as well as solvatochromic probes. With increasing chain length as well as solvent polarity, enhanced site isolation of the core has been observed. These findings have been correlated to actual molecular dimensions independently measured by pulsed field gradient spin-echo (PGSE) NMR. The developed synthetic methodology offers a rapid route to efficient encapsulation of functional moieties and therefore has potential for the design of new materials. PMID- 11273595 TI - Phenylisoserine: a versatile amino acid for the construction of novel beta peptide structures. AB - The N-Boc O-tert-butyldimethysilyl-substituted hexa-beta-peptide methyl ester 18 was constructed from the O-TBS ether of (-)-(2R, 3S)-phenylisoserine. By NMR, it was determined that this homo beta-peptide adopts a highly stable beta-strand type secondary structure in chloroform solution, which is stabilized by both hydrophobic interactions involving the OTBS methyl groups of residues i and i + 2, and inter-(five-membered)/intra (six-membered)-residue H-bonding interactions. These interactions are systematically repeated along the peptide chain and, thereby, operate in concert to stabilize the observed conformation of 18. PMID- 11273597 TI - Surface reconstruction and the difference in surface acidity between gamma- and eta-alumina. AB - The surfaces of the gamma and eta forms of alumina are well known to differ significantly in Lewis acidity. Surface reconstructions observed on gamma- and eta-alumina are also qualitatively different, despite the close similarity of the bulk structures. Here we demonstrate through first-principles calculations that subtle differences in the bulk point defect distribution between these two forms of alumina give rise to the major differences in the mode of surface reconstruction and correlate with the different levels of Lewis acidity. PMID- 11273598 TI - The magnesium-ene cyclization stereochemically directed by an allylic oxyanionic group and its application to a highly stereoselective synthesis of (+/-) matatabiether. Allylmagnesium compounds by reductive magnesiation of allyl phenyl sulfides. AB - The first example of a magnesium-ene cyclization stereochemically directed by an allylic oxyanionic group is demonstrated by a highly stereoselective synthesis of the bicyclic terpene matatabiether 10. The synthetic method is particularly valuable, not only because of the stereochemical control and the utility of the versatile hydroxyl group introduced into the product, but also because the precursor of the allylmagnesium is an allyl phenyl sulfide, which is more stable and more easily prepared in a connective fashion than the usual allyl halide precursor. Since the presence of lithium ions encourages undesirable proton transfer to the cyclized organometallic and is detrimental to the stereochemical control, the conversion of the allylic thioether to the allylmagnesium utilizes a lithium-free method involving direct reductive magnesiation in the presence of the magnesium-anthracene complex. PMID- 11273600 TI - Chiral studies in amorphous solids: the effect of the polymeric glassy state on the racemization kinetics of bridged paddled binaphthyls. AB - Optical activity, used here for the first time to gain information about the amorphous solid state, allows previously unavailable insight into the dynamic properties of polymer glasses and their effect on a chemical process. This is accomplished by dispersing in polymer glasses atropisomeric bridged binaphthyls with appended oligophenyl paddles of varying sizes and studying the racemization kinetics as a function of temperature. The racemization occurs by a simple one dimensional twisting motion and, without effect on the intrinsic mechanism, sweeps out a variable volume of the matrix as the paddle length is increased. The racemization is limited by the polymer matrix only for probes with a minimum paddle size and only when the time scale for racemization is comparable to the time scale for segmental motion of the polymer matrix. The high barrier for this racemization is unique in probe studies of glasses and causes these overlapping time scales to occur significantly below the glass transition temperature. These measurements yield a clear quantitative view of the role of segmental dynamics on the racemization kinetics of the binaphthyls and allow the important demonstration, via the transition from first-order to stretched exponential kinetics, that heterogeneous dynamics persist deep within the glassy state. PMID- 11273599 TI - Total syntheses of tumor-related antigens N3: probing the feasibility limits of the glycal assembly method. AB - The total syntheses of two octasaccharide antigens isolated from human milk, 1 and 2, and their corresponding allyl glycosides, 3 and 4, have been achieved by utilizing the glycal method. Convergent assembly of the core hexasaccharides and concurrent introduction of two alpha-L-fucosyl moieties at the late stage of the syntheses provided these complex carbohydrates in a concise manner. With synthetic material obtained, biological evaluations of these antigens as potential gastrointestinal cancer immunotherapeutic agents have been initiated. PMID- 11273601 TI - Dendrimeric organochalcogen catalysts for the activation of hydrogen peroxide: improved catalytic activity through statistical effects and cooperativity in successive generations. AB - Dendrimeric polyphenylsulfides, -selenides, and -tellurides are prepared in high yield using propyloxy spacers to connect the phenylchalcogeno groups to the dendrimeric core. The selenides and tellurides catalyze the oxidation of bromide with hydrogen peroxide to give positive bromine species that can be captured by cyclohexene in two-phase systems. The corresponding sulfides show no catalytic activity. The increase in the rate of catalysis followed statistical effects for 1, 6, and 12 phenyltelluro groups. However, the increase in the rate of catalysis exceeds statistical contributions for the first few generations with 1, 3, 6, and 12 phenylseleno groups and suggested cooperativity among phenylseleno groups. The increase in catalytic rate was lost upon replacing all but one phenylseleno group with phenoxy groups. On the basis of H2O2 consumed, the dendrimer with 12 phenylseleno groups has a turnover number of >60 000 mol of H2O2 consumed per mole of catalyst. PMID- 11273603 TI - Photophysical properties of long rodlike meso-meso-linked zinc(II) porphyrins investigated by time-resolved laser spectroscopic methods. AB - The molecular design of directly meso-meso-linked porphyrin arrays as a new model of light-harvesting antenna as well as a molecular photonic wire was envisaged to bring the porphyrin units closer for rapid energy transfer. For this purpose, zinc(II) 5,15-bis(3,5-bis(octyloxy)phenyl)porphyrin (Z1) and its directly meso meso-linked porphyrin arrays up to Z128 (Zn, n represents the number of porphyrins) were synthesized. The absorption spectra of these porphyrin arrays change in a systematic manner with an increase in the number of porphyrins; the high-energy Soret bands remain at nearly the same wavelength (413-414 nm), while the low-energy exciton split Soret bands are gradually red-shifted, resulting in a progressive increase in the exciton splitting energy. The exciton splitting is nicely correlated with the values of cos[pi/(N + 1)] according to Kasha's exciton coupling theory, providing a value of 4250 cm(-1) for the exciton coupling energy in the S(2) state. The increasing red-shifts for the Q-bands are rather modest. The fluorescence excitation anisotropy spectra of the porphyrin arrays show that the photoexcitation of the high-energy Soret bands exhibits a large angle difference between absorption and emission dipoles in contrast with the photoexcitation of the low-energy exciton split Soret and Q-bands. This result indicates that the high-energy Soret bands are characteristic of the summation of the individual monomeric transitions with its overall dipole moment deviated from the array chain direction, while the low-energy Soret bands result from the exciton splitting between the monomeric transition dipoles in line with the array chain direction. From the fluorescence quantum yields and fluorescence lifetime measurements, the radiative coherent length was estimated to be 6-8 porphyrin units in the porphyrin arrays. Ultrafast fluorescence decay measurements show that the S(2) --> S(1) internal conversion process occurs in less than 1 ps in the porphyrin arrays due to the existence of exciton split band as a ladder-type deactivation channel, while this process is relatively slow in Z1 (approximately 1.6 ps). The rate of this process seems to follow the energy gap law, which is mainly determined by the energy gap between the two Soret bands of the porphyrin arrays. PMID- 11273602 TI - Chirality organization of ferrocenes bearing podand dipeptide chains: synthesis and structural characterization. AB - A variety of ferrocenes bearing podand dipeptide chains have been synthesized to form an ordered structure in both solid and solution states and have been investigated by 1H NMR, FT-IR, CD, and X-ray crystallographic analyses. Conformational enantiomerization through chirality organization was achieved by the intramolecular hydrogen bondings between the podand dipeptide chains. The single-crystal X-ray structure determination of the ferrocene 2 bearing the podand dipeptide chains (-D-Ala-D-Pro-OEt) revealed two C2-symmetric intramolecular hydrogen bondings between CO (Ala) and NH (another Ala) of each podand dipeptide chain to induce the chirality-organized structure. The molecular structures of the ferrocene 1 composed of the podand L-dipeptide chains (-L-Ala-L Pro-OEt) and 2 are in a good mirror image relationship, indicating that they are conformational enantiomers. An opposite helically ordered molecular arrangement was formed in the crystal packing of 2 as compared with 1. The ferrocene 2 exhibited induced circular dichroism (CD), which appeared at the absorbance of the ferrocene moiety. The mirror image of the CD signals between 1 and 2 was observed, suggesting that the chirality-organized structure via intramolecular hydrogen bondings is present even in solution. The ferrocene 4 bearing the podand dipeptide chains (-Gly-L-Leu-OEt) also showed an ordered structure in the crystal based on two intramolecular hydrogen bondings between CO (Gly) and NH (another Gly) of each podand dipeptide chain, together with intermolecular hydrogen bondings between CO adjacent to the ferrocene unit and NH (neighboring Leu) to create the highly organized self-assembly. A different self-assembly was observed in the crystal of the ferrocene 5 composed of the podand dipeptide chains (-Gly-L Phe-OEt), wherein each molecule is bonded to two neighboring molecules through two pairs of symmetrical intermolecular hydrogen bonds to form a 14-membered intermolecularly hydrogen-bonded ring. These ordered structures based on the intramolecular hydrogen bondings in the solution state are also confirmed by 1H NMR and FT-IR. PMID- 11273605 TI - Stark effects in gas-phase electronic spectra. Dipole moment of aniline in its excited S(1) state. AB - Measurements of the Stark effect on the rotationally resolved S(1)<--S(0) fluorescence excitation spectrum of aniline are reported, providing quantitative information about the degree of charge transfer in the electronic transition. We find that mu(a)(S(1)) = 2.801 +/- 0.007 D, a value that is approximately 150% larger than the ground state, mu(a)(S(0)) = 1.129 +/- 0.005 D. The enhanced value of the dipole moment in the S(1) state is attributed to more efficient electron donation by the quasi-planar amino group to the aromatic ring. PMID- 11273604 TI - Diels-Alder topochemistry via charge-transfer crystals: novel (thermal) single crystal-to-single-crystal transformations. AB - The solid-state [4+2] cycloaddition of anthracene to bis(N-ethylimino)-1,4 dithiin occurs via a unique single-phase topochemical reaction in the intermolecular (1:1) charge-transfer crystal. The thermal heteromolecular solid state condensation involves the entire crystal, and this rare crystalline event follows topochemical control during the entire cycloaddition. As a result, a new crystalline modification of the Diels-Alder product is formed with a crystal packing similar to that of the starting charge-transfer crystal but very different from that of the (thermodynamically favored) product modification obtained from solution-phase crystallization. Such a single-phase transformation is readily monitored by X-ray crystallography at various conversion stages, and the temporal changes in crystallographic parameters are correlated with temperature-dependent (solid-state) kinetic data that are obtained by 1H NMR spectroscopy at various reaction times. Thus, an acceleration of the solid-state reaction over time is found which results from a progressive lowering of the activation barrier for cycloaddition in a single crystal as it slowly and homogeneously converts from the reactant to the product lattice. PMID- 11273606 TI - Light-harvesting and photocurrent generation by gold electrodes modified with mixed self-assembled monolayers of boron-dipyrrin and ferrocene-porphyrin fullerene triad. AB - Three different kinds of mixed self-assembled monolayers have been prepared to mimic photosynthetic energy and electron transfer on a gold surface. Pyrene and boron-dipyrrin were chosen as a light-harvesting model. The mixed self-assembled monolayers of pyrene (or boron-dipyrrin) and porphyrin (energy acceptor model) reveal photoinduced singlet-singlet energy transfer from the pyrene (or boron dipyrrin) to the porphyrin on the gold surface. The boron-dipyrrin has also been combined with a reaction center model, ferrocene-porphyrin-fullerene triad, to construct integrated artificial photosynthetic assemblies on a gold electrode using mixed monolayers of the respective self-assembled unit. The mixed self assembled monolayers on the gold electrode have established a cascade of photoinduced energy transfer and multistep electron transfer, leading to the production of photocurrent output with the highest quantum yield (50 +/- 8%, based on the adsorbed photons) ever reported for photocurrent generation at monolayer-modified metal electrodes and across artificial membranes using donor acceptor linked molecules. The incident photon-to-current efficiency (IPCE) of the photoelectrochemical cell at 510 and 430 nm was determined as 0.6% and 1.6%, respectively. Thus, the present system provides the first example of an artificial photosynthetic system, which not only mimics light-harvesting and charge separation processes in photosynthesis but also acts as an efficient light to-current converter in molecular devices. PMID- 11273607 TI - Trapping of an activated HF molecule inside a double four-ring unit: a quantum chemical model of the microporous fluorinated gallium phosphate ULM-18. AB - The penetration of a proton into the prenucleation building unit of a microporous gallophosphate and its interaction with an encapsulated fluorine anion have been investigated by means of DFT calculations. The inorganic part of the fluorinated gallophosphate ULM-18 has been modeled by a neutral, double four-ring (D4R) unit of formula [(GaOH)4(HPO4)4-H2O] encapsulating the fluorine ion. Assuming the cage to be rigid and to retain throughout the calculations the geometry determined from X-ray diffraction (XRD), the position of F(-) has been optimized, either as an isolated guest species or in the presence of an incoming proton. In agreement with the XRD structure, the fluorine atom has been shown to occupy in both cases a nonsymmetric position in the cage, being attached to three gallium atoms out of four. The distribution of the molecular electrostatic potential inside and outside the (F(-))@[(GaOH)4(HPO4)4-H2O] system has provided indications concerning the pathways that could be used by an incoming proton to penetrate the D4R unit and to approach the fluorine anion. The migration of a proton from an external site of fixation to the interior of the D4R unit has been found possible through two faces out of six. In both cases, the process has been found exothermic by approximately 0.17 eV and the energy barrier was estimated to approximately 0.8 eV. Inside the gallophosphate cage, the proton first adopts a position typical of a strong F...H...O bond made possible through an important shift of the fluorine anion away from the tripod of bonded gallium atoms. Then, the F(-)...H(+) system can easily evolve back and forth on a flat potential curve between one of the F...H...O bonded conformations and a situation characterized by the cleavage of the H...O link and the formation of a moderately activated F-H molecule, with the fluorine still attached to three gallium atoms. PMID- 11273608 TI - Acid-base chemistry of a carbenium ion in a zeolite under equilibrium conditions: verification of a theoretical explanation of carbenium ion stability. AB - The 1,3-dimethylcyclopentenyl carbenium ion (C7H11(+)) was reproducibly prepared on zeolite HZSM-5 using a pulse-quench reactor, and then each of a number of bases was coadsorbed into the catalyst channels to either compete with the cation for protonation or to possibly react with it as a nucleophile. For seven bases with proton affinities (PA) between 142 and 212.1 kcal/mol, there was no reaction with C7H11(+). Coadsorption of smaller amounts of dimethylacetamide (PA = 217 kcal/mol) also produced no reaction, but with a higher loading, a proton was transferred from the carbenium ion to the base to leave 1,3-dimethylcyclopenta 1,3-diene in the zeolite as a neutral olefin. Deprotonation was the primary reaction with coadsorption of either pyridine (PA = 222 kcal/mol) or trimethylphosphine (PA = 229.2 kcal/mol). The estimated experimental deprotonation enthalpy for C7H11(+), approximately 217 kcal/mol in the zeolite, is in excellent agreement with MP4/6-311G gas-phase value of 215.6 kcal/mol. Coadsorption of either NH3 (PA = 204.0 kcal/mol) or PH3 (PA = 188 kcal/mol) does not deprotonate the carbenium ion, but these species do react as nucleophiles to form onium ion derivatives of C7H11(+). Analogous onium complexes with pyridine or trimethylphosphine formed in lower yields due to steric constraints in the zeolite channels. The essential experimental observations were all predicted and explained by density functional calculations (B3LYP/6-311G) and extensions of our recently developed theory of carbenium ion stability in zeolites. In addition, we report theoretical geometries for several complexes which contain unusual C-H- - X hydrogen bonds. PMID- 11273609 TI - A single transition state serves two mechanisms: an ab initio classical trajectory study of the electron transfer and substitution mechanisms in reactions of ketyl radical anions with alkyl halides. AB - Molecular dynamics has been used to investigate the reaction of a series of ketyl anion radicals and alkyl halides, CH2O(*)(-) + CH3X (X = F, Cl, Br) and NCCHO(*)( ) + CH3Cl. In addition to a floppy outer-sphere transition state which leads directly to ET products, there is a strongly bound transition state that yields both electron transfer (ET) and C-alkylated (SUB(C)) products. This common transition state has significant C-- C bonding and gives ET and SUB(C) products via a bifurcation on a single potential energy surface. Branching ratios have been estimated from ab initio classical trajectory calculations. The SUB(C) products are favored for transition states with short C--C bonds and ET for long C--C bonds. ET reactivity can be observed even at short distances of r(C)(-)(C) = ca. 2.4 A as in the transition state for the reaction NCCHO(*)(-) + CH3Cl. Therefore, the ET/SUB(C) reactivity is entangled over a significant range of the C--C distance. The mechanistic significance of the molecular dynamics study is discussed. PMID- 11273610 TI - Reactions of group V metal atoms with water molecules. Matrix isolation FTIR and quantum chemical studies. AB - Laser-ablated group V metal atoms (V, Nb, Ta) were co-deposited with water molecules in excess argon. The V atoms reacted with water to form the inserted HVOH molecule spontaneously. The Nb atoms reacted with water to form the NbOH2 complex and the inserted HNbOH molecule. Broad-band photolysis produced the H2VO and H2NbO molecules as well as the VO and NbO monoxides. For Ta + H2O reactions, neither TaOH2 nor HTaOH was observed, while the H2TaO molecule was produced on annealing, and the H2 elimination process was not observed on photolysis. The aforementioned species were identified via isotopic substitutions as well as density functional calculations. Qualitative analysis of the possible reaction paths leading to the observed products is proposed. The results have been compared with our earlier works concerning the Sc and group IV metal atoms with water reactions in order to observe existent trends for the early transition metal atoms. PMID- 11273611 TI - Platinum dioxide cation: easy to generate experimentally but difficult to describe theoretically. AB - A formal platinum(V) dioxide cation [Pt,O2](+) can be generated in the gas phase by successive oxidation of Pt(+) with N2O. The ion's reactivity is in keeping with the dioxide structure OPtO(+), rather than with [Pt,O2](+) isomers having intact O-O bonds, e.g., the dioxygen complex Pt(O2)(+) and peroxo species PtOO(+). Inter alia due to the high ionization energy of the neutral counterpart (11.2 eV), the [Pt,O2](+) cation is a rather aggressive reagent toward oxidizable neutrals. [Pt,O2](+) is even capable of activating inert substrates such as H2, CO, and CH4. Further, a sequence for the catalytic conversion CO + N(2)O --> CO2 + N2 is described with a turnover number of >100 for the catalytically active species PtOn(+) (n = 0-2). As a consequence of the high reactivity, however, the observed selectivities with most substrates are rather poor. For example, the reaction of PtO2(+) with ethane gives rise to 10 different product channels. In an attempt to analyze the structural features and different minima of the [Pt,O2](+) system, extensive ab initio studies are performed. While correlated ab initio methods describe the system reasonably well, density functional theory turns out to be much less accurate in terms of both structural and energetic descriptions. PMID- 11273612 TI - Photodissociation of ethylene sulfide at 193 nm: a photofragment translational spectroscopy study with VUV synchrotron radiation and ab initio calculations. AB - Photodissociation of ethylene sulfide at 193 nm has been studied using photofragment translational spectroscopy and ab initio theoretical calculations. Tunable synchrotron radiation was used as a universal but selective probe of the reaction products to reveal new aspects of the photodissociation dynamics. The channel giving S + C2H4 was found to be dominated by production of ground-state sulfur atoms (S(3P):S(1D) = 1.44:1), mostly through a spin-forbidden process. The results also suggest the presence of a channel giving S(3P) in conjunction with triplet ethylene C2H4 (3B(1u)) and allow insight into the energy of the latter species near its equilibrium geometry, in which the two methylene groups occupy perpendicular planes. In addition, a channel leading to the production of H2S with C2H2 also has been observed. Our experimental results are supported and elaborated by theoretical calculations. PMID- 11273613 TI - Long-range magnetic order in Mn[N(CN)2]2(pyz) (pyz = pyrazine). Susceptibility, magnetization, specific heat, and neutron diffraction measurements and electronic structure calculations. AB - Using dc magnetization, ac susceptibility, specific heat, and neutron diffraction, we have studied the magnetic properties of Mn[N(CN)2]2(pyz) (pyz = pyrazine) in detail. The material crystallizes in the monoclinic space group P2(1)/n with a = 7.3248(2), b = 16.7369(4), and c = 8.7905 (2) A, beta = 89.596 (2) degrees, V = 1077.65(7) A(3), and Z = 4, as determined by Rietveld refinement of neutron powder diffraction data at 1.35 K. The 5 K neutron powder diffraction data reflect very little variation in the crystal structure. Interpenetrating ReO3-like networks are formed from axially elongated Mn(2+) octahedra and edges made up of mu-bonded [N(CN)2](-) anions and neutral pyz ligands. A three dimensional antiferromagnetic ordering occurs below T(N) = 2.53(2) K. The magnetic unit cell is double the nuclear one along the a- and c-axes, giving the (1/2, 0, 1/2) superstructure. The crystallographic and antiferromagnetic structures are commensurate and consist of collinear Mn(2+) moments, each with a magnitude of 4.15(6) mu(B) aligned parallel to the a-direction (Mn-pyz-Mn chains). Electronic structure calculations indicate that the exchange interaction is much stronger along the Mn-pyz-Mn chain axis than along the Mn-NCNCN-Mn axes by a factor of approximately 40, giving rise to a predominantly one-dimensional magnetic system. Thus, the variable-temperature magnetic susceptibility data are well described by a Heisenberg antiferromagnetic chain model, giving g = 2.01(1) and J/k(B) = -0.27(1) K. Owing to single-ion anisotropy of the Mn(2+) ion, field induced phenomena ascribed to spin-flop and paramagnetic transitions are observed at 0.43 and 2.83 T, respectively. PMID- 11273615 TI - Fabrication of structured porous film by electrophoresis. PMID- 11273616 TI - Controlling photoinduced electron transfer within a hydrogen-bonded porphyrin phenoxynaphthacenequinone photochromic system. PMID- 11273617 TI - The first intermolecular transition metal-catalyzed [5+2] cycloadditions with simple, unactivated, vinylcyclopropanes. PMID- 11273614 TI - Unsupported Pt(0)-Tl(I) bonds in the simple [Pt(PPh2Py)3Tl](+) complexes. PMID- 11273618 TI - Isolation and characterization of light actinide metallofullerenes. PMID- 11273619 TI - Formation of high-quality CdTe, CdSe, and CdS nanocrystals using CdO as precursor. PMID- 11273621 TI - The formation of dimensionally ordered silicon nanowires within mesoporous silica. PMID- 11273620 TI - Comparison of protein backbone entropy and beta-sheet stability: NMR-derived dynamics of protein G B1 domain mutants. PMID- 11273622 TI - The first crystalline alkali metal salt of a benzenoid radical anion without a stabilizing substituent and of a related dimer: X-ray structures of the toluene radical anion and of the benzene radical anion dimer potassium-crown ether salts. PMID- 11273623 TI - Temporal cognition and the phenomenology of time: a multiplicative function for apparent duration. AB - The literature on time perception is discussed. This is done with reference both to the "cognitive-timer" model for time estimation and to the subjective experience of apparent duration. Three assumptions underlying the model are scrutinized. I stress the strong interplay among attention, arousal, and time perception, which is at the base of the cognitive-timer model. It is suggested that a multiplicative function of two key components (the number of subjective time units and their size) should predict apparent duration. Implications for other cognitive domains are drawn, and in particular an analogy is suggested between apparent duration and apparent movement. PMID- 11273624 TI - Consciousness during dreams. AB - Two aspects of consciousness are first considered: consciousness as awareness (phenomenological meaning) and consciousness as strategic control (functional meaning). As to awareness, three types can be distinguished: first, awareness as the phenomenal experiences of objects and events; second, awareness as meta awareness, i.e., the awareness of mental life itself; third, awareness as self awareness, i.e., the awareness of being oneself. While phenomenal experience and self-awareness are usually present during dreaming (even if many modifications are possible), meta-awareness is usually absent (apart from some particular experiences of self-reflectiveness) with the major exception of lucid dreaming. Consciousness as strategic control may also be present in dreams. The functioning of consciousness is then analyzed, following a cognitive model of dream production. In such a model, the dream is supposed to be the product of the interaction of three components: (a) the bottom-up activation of mnemonic elements coming from LTM systems, (b) interpretative and elaborative top-down processes, and (c) monitoring of phenomenal experience. A feedback circulation is activated among the components, where the top-down interpretative organization and the conscious monitoring of the oneiric scene elicitates other mnemonic contents, according to the requirements of the dream plot. This dream productive activity is submitted to unconscious and conscious processes. PMID- 11273625 TI - The location problem for color subjectivism. AB - According to color subjectivism, colors are mental properties, processes, or events of visual experiences of color. I first lay out an argument for subjectivism founded on claims from visual science and show that it also relies on a philosophical assumption. I then argue that subjectivism is untenable because this view cannot provide a plausible account of color perception. I describe three versions of subjectivism, each of which combines subjectivism with a theory of perception, namely sense datum theory, adverbialism, and the virtual color proposal, and argue that each version faces serious objections. Considering these three theories of perception to be exhaustive of those available to the subjectivist, I conclude that subjectivism is untenable and that the scientifically motivated argument for this view is unsound. I then offer the diagnosis that the philosophical assumption on which this argument relies is mistaken. PMID- 11273626 TI - Spatial location in color vision. AB - Ross argues that the location problem for color-the problem of how it is represented as occupying a particular location in space-constitutes an objection to color subjectivism. There are two ways in which the location problem can be interpreted. First, it can be read as a why-question about the relation of visual experience to the environment represented: Why does visual experience represent a patch of color as located in this part of space rather than that? On this interpretation, the subjectivist can answer Ross's objection by appealing to the physical location of reflectance rather than color. Second, it can be read as a how-question about visual representation itself: How does visual experience put together the experience of a color with the experience of its being located in space? This version makes the location problem a problem about visual experience itself and renders the ontology of color irrelevant to its solution. The location problem is thus no more a problem for the color subjectivist than for the color realist. PMID- 11273627 TI - The location problem reconsidered: a reply to Ross. PMID- 11273628 TI - The data problem for color objectivism. PMID- 11273629 TI - Putting color back where it belongs. AB - I disagree with Ross about the location of colors: They are in the brain, not in the external world. It is difficult to deny that there are colors in our conscious visual experience, and if we take the causal theory of perception seriously, we cannot identify these colors with the beginning of the causal chain in perception (external objects in the distal stimulus field), but we must search for them at the end of the causal chain (in the brain). Several lines of compelling evidence from cognitive neuroscience (e.g., synesthesia, dreaming, and achromatopsia) demonstrate unambiguously that color is in the brain. Furthermore, it seems that Ross has failed to consider one substantial version of subjectivism in his article. This monistic approach to color and consciousness appears to be the least implausible alternative when we try to understand what colors are and where they reside. PMID- 11273630 TI - Color, mental location, and the visual field. PMID- 11273631 TI - Subjectivism, physicalism, or none of the above? Comments on Ross's "The location problem for color subjectivism". PMID- 11273632 TI - Explaining metamers: right degrees of freedom, not subjectivism. PMID- 11273633 TI - What colors? whose colors? PMID- 11273634 TI - In defense of color psychophysicalism. PMID- 11273635 TI - Commentary on P.W. Ross: the location problem for color subjectivism. PMID- 11273636 TI - Color realism: toward a solution to the "hard problem". PMID- 11273640 TI - Selective introduction of antisense oligonucleotides into single adult CNS progenitor cells using electroporation demonstrates the requirement of STAT3 activation for CNTF-induced gliogenesis. AB - We have developed a novel method in which antisense DNA is selectively electroporated into individual adult neural progenitor cells. By electroporation of antisense oligonucleotides against signal transducer and activator of transcription 3 (STAT3) we demonstrate that ciliary neurotrophic factor (CNTF) is an instructive signal for astroglial type 2 cell fate specifically mediated via activation of STAT3. Activation of the mitogen-activated protein kinase (MAPK) signaling pathway induced only a transient increase in glial fibrillary acidic protein (GFAP) expression, and inhibition of this signaling pathway did not block the induction by CNTF of glial differentiation in progenitor cells. In addition we show that microelectroporation is a new powerful method for introducing antisense agents into single cells in complex cellular networks. PMID- 11273639 TI - pRb2/p130 gene overexpression induces astrocyte differentiation. AB - There are many data on the activity of the RB gene in neural differentiation and apoptosis, but the role of pRb2/p130 in neuronal and glial maturation has been far less investigated. To elucidate the role of pRb2/p130 in astrocyte development we overexpressed this protein in astrocytoma and normal astrocyte cultures by adenoviral-mediated gene transfer. In astrocytoma cells, p130/RB2 overexpression resulted in a significant reduction of cell growth and in an increased G(0)/G(1) cell population. We did not observe any induction of programmed cell death as determined by TUNEL reaction. Interestingly, pRb2/p130 overexpression induced astrocyte differentiation. Astrocyte cell cycle arrest and differentiation seemed to proceed through a way distinct from the p53 pathway. PMID- 11273641 TI - Correlation between putative inhibitory molecules at the dorsal root entry zone and failure of dorsal root axonal regeneration. AB - The molecular mechanisms involved in preventing regenerating dorsal root axons from entering the spinal cord at the dorsal root entry zone (DREZ) are obscure. We used immunohistochemistry, in situ hybridization, and electron microscopy to study axonal regeneration after dorsal rhizotomy in adult rats and its relationship to cellular changes and the distribution of putative growth inhibitory molecules in this region. Astrocyte processes, ending as bulb-shaped expansions, grew up to 700 microm into the basal lamina tubes of injured roots, where regenerating axons were also present. Some of these axons approached or reached the DREZ but grew no further; others turned back toward the ganglion, suggesting the presence of repulsive cues in or near the DREZ. Tenascin-C mRNA and protein and CSPG stub immunoreactivity were strongly upregulated in the roots after rhizotomy, but were only weakly expressed in the DREZ. Tenascin-R immunoreactivity was confined to CNS tissue, and unaffected by rhizotomy. Large, rounded GFAP-negative, NG2-immunoreactive cells, a few of which were OX42 positive, were found in the DREZ following rhizotomy. Astrocyte processes projecting into the roots were tenascin-R and NG2 negative. Hence, only NG2 expressing cells and tenascin-R were appropriately situated to inhibit regeneration through the DREZ. PMID- 11273642 TI - Brn-3a activates the expression of Bcl-x(L) and promotes neuronal survival in vivo as well as in vitro. AB - The determination of cell fate plays a critical role during the later stages of embryogenesis and the early postnatal period-a time during which approximately half of neurons born during neurogenesis undergo programmed cell death. It has previously been reported that the type IV POU domain transcription factor Brn-3a plays a role in the maturation and survival of sensory neuronal populations. Indeed we have shown that the long form of Brn-3a is capable of activating expression of the antiapoptotic Bcl-2 gene and enhancing neuronal survival in cultures of sensory neurons. In this study, we report the identification of another antiapoptotic family member, Bcl-x(L), as a target gene of Brn-3a in sensory neurons, providing a further mechanism by which Brn-3a determines sensory neuronal fate during development. Bcl-x(L) gene expression is activated by Brn-3a in sensory but not in sympathetic neurons and its expression is reduced by antisense inhibition of Brn-3a expression in sensory neurons. Most importantly, both Bcl-x(L) expression and neuronal survival are enhanced by the overexpression of Brn-3a in dorsal root ganglion in vivo in a model of sciatic nerve injury in the intact animal. PMID- 11273643 TI - Inhibitory proteoglycan immunoreactivity is higher at the caudal than the rostral Schwann cell graft-transected spinal cord interface. AB - To begin to evaluate the influence that proteoglycans may have on the success of Schwann cell (SC) transplants to induce axonal regrowth across a complete transection lesion and beyond, we determined the pattern of expression of inhibitory chondroitin sulfate proteoglycans (CSPGs) 3 weeks after transplantation into completely transected adult rat thoracic spinal cord. Using immunohistochemistry, we observed that: (1) CSPGs recognized by CS-56 antibody are present on astrocytes, fibroblasts, and SCs in the distal graft, and at lesion and cystic cavity borders; (2) CS-56 immunoreactivity (IR) is greater at the caudal SC graft-host cord interface than the rostral interface; (3) phosphacan-IR, also greater at the caudal interface, is associated with astrocytes, fibroblasts, as yet unidentified cells, and extracellular matrix; (4) neurocan-IR is present on astrocytes and as yet unidentified cells in grey and white matter; and (5) NG2-IR is associated with matrix near SC grafts, unidentified cells mainly in white matter, and lesion borders and cysts. Neither oligodendrocytes nor activated macrophages/microglia were immunostained. In sum, the CSPGs studied are increased at 3 weeks, especially at the caudal SC graft cord interface, possibly contributing to an inhibitory molecular barrier that precludes regrowing descending axons from entering the caudal host cord. PMID- 11273644 TI - Expression and function of ganglioside 9-O-acetyl GD3 in postmitotic granule cell development. AB - We have shown previously that the Jones monoclonal antibody (Jones mAb) recognizes 9-O-acetyl GD3 expressed during periods of neuronal migration and neurite outgrowth in the developing rat nervous system. In the present study we investigated the expression of this ganglioside in the developing cerebellum and correlated this expression with granule cell migration. Electron microscopic immunocytochemistry revealed that around the peak of cerebellar neuronal migration (7-day-old rat), 9-O-acetyl GD3 was localized at the contact sites between migrating granule cells and radial glia in the external granular layer and prospective molecular layer. In addition, using microexplant and slice cultures of the postnatal rat cerebellum, we tested whether the ganglioside detected by our antibody contribute to the regulation of neuronal migration in the cerebellar cortex. We have shown that the Jones mAb blocks the migration of neurons in a dose-dependent manner. These findings suggest strongly that 9-O acetyl GD3 is involved in granule cell migration in the developing cerebellum. PMID- 11273645 TI - Structural diversity despite strong evolutionary conservation in the 5' untranslated region of the P-type dystrophin transcript. AB - Analysis of the 5'-flanking regions of the Purkinje (P-) dystrophin genes and mRNAs in different species revealed strong sequence conservation but functional diversity. Multiple transcription initiation sites were identified in cerebella and muscles, tissues expressing P-dystrophin. The predominant initiation site was conserved, with another muscle-specific site located upstream. Despite sequence homology, significant tissue- and species-specific structural diversity in the P type 5'-ends exists, including alternative splicing within the 5'-untranslated region combined with alternative splicing of intron 1. One amino terminus is conserved in mammals and, to a lesser extent, in chicken. However, alternative usage of ATG codons may result in a choice of N-termini or translation of short upstream ORFs in different species. Promoter activity of a fragment upstream of the cap site was shown by transient expression in myoblasts and in vivo following intramuscular injection. It is tissue- and developmentally regulated. Analysis of promoter deletions suggests the existence of negative regulatory elements in the proximal region. PMID- 11273646 TI - Developmental expression of the small-conductance Ca(2+)-activated potassium channel SK2 in the rat retina. AB - Small-conductance Ca(2+)-activated potassium (SK) channels are present in most central neurons, where they mediate the afterhyperpolarizations (AHPs) following action potentials. SK channels integrate changes in intracellular Ca(2+) concentration with membrane potential and thus play an important role in controlling firing pattern and excitability. Here, we characterize the expression pattern of the apamin-sensitive SK subunits, SK2 and SK3, in the developing and adult rat retina using in situ hybridization and immunohistochemistry. The SK2 subunit showed a distinct and developmentally regulated pattern of expression. It appeared during the first postnatal week and located to retinal ganglion cells and to subpopulations of neurons in the inner nuclear layer. These neurons were identified as horizontal cells and dopaminergic amacrine cells by specific markers. In contrast to SK2, the SK3 subunit was detected neither in the developing nor in the adult retina. These results show cell-specific expression of the SK2 subunit in the retina and suggest that this channel underlies the apamin-sensitive AHP currents described in retinal ganglion cells. PMID- 11273647 TI - The metabolism and imaging in live cells of the bovine prion protein in its native form or carrying single amino acid substitutions. AB - Prion diseases are probably caused by an abnormal form of a cellular glycoprotein, the prion protein. Recent evidence suggests that the prion strain causing BSE has been transmitted to humans, thereby provoking a variant form of Creutzfeldt-Jacob disease. In this work, we analyzed the behavior of normal and malformed isoforms of the bovine PrP in transfected mammalian cell lines. Biochemical and immunocytochemical assays were complimented with imaging of live cells expressing fusion constructs between PrP and GFP. Bovine homologues of human E200K and D178N (129M) mutations were used as models of pathogenic isoforms. We show that the GFP does not impair the metabolism of native and mutant bPrPs and is thus a valid marker of PrP cellular distribution. We also show that each amino acid replacement provokes alterations in the cell sorting and processing of bPrP. These are different from those ascribed to both murine mutant homologues. However, human and bovine PrPs carrying the D178N genotype had similar cellular behavior. PMID- 11273648 TI - Autocrine regulation of norepinephrine transporter expression. AB - The norepinephrine transporter (NET) is a neurotransmitter scavenger and site of drug action in noradrenergic neurons. The aim of this study was to identify mechanisms that regulate NET expression during the development of quail (q) sympathetic neuroblasts, which develop from neural crest stem cells. Neurotrophin 3 (NT-3) and transforming growth factor beta1 (TGF-beta1) cause an increase of qNET mRNA levels in neural crest cells. When combined, the growth factors are additive in increasing qNET mRNA levels. Both NT-3 and TGF-beta1 are synthesized by neural crest cells. Onset of NET expression precedes the onset of neural crest stem cell emigration from the neural tube. In older embryos, qNET is expressed by several crest-derived and noncrest tissues. The data show that qNET expression in presumptive sympathetic neurons is initiated early in embryonic development by growth factors that are produced by neural crest cells themselves. Moreover, the results support our previous observations that norepinephrine transport contributes to the regulation of the differentiation of neural crest stem cells into sympathetic neurons. PMID- 11273649 TI - CD81 regulates neuron-induced astrocyte cell-cycle exit. AB - Astrocytes respond to contact with neurons by cell-cycle arrest and complex process formation. In our effort to discover the molecular mechanisms that underlie this phenomenon we have identified a known tetraspanin, CD81, as a critical component of astrocyte responses to neuronal differentiation signals. Here we show that CD81 is expressed on the surface of the astrocyte and that its expression level can be modulated by contact with neurons. Further, using three separate antibodies, 2F7, Eat1, and Eat2, which recognize unique epitopes in the extracellular domains of the CD81 protein, we show that there is a unique domain, recognized by Eat1, that is required for astrocyte cell-cycle withdrawal in response to neurons. This is likely due to conformational changes in the CD81 molecule, as inclusion of 2F7 actually augments neuron-induced astrocyte growth arrest. The critical nature of CD81 in normal astrocyte-neuron biology was confirmed by using mice in which CD81 had been deleted by homologous recombination. Astrocytes null at the CD81 locus were blind to the proliferative arrest encoded on the neuronal cell surface. Taken together, these data strongly suggest that CD81 is a critical regulator of neuron-induced astrocytic differentiation. PMID- 11273650 TI - Transplantation of mammalian olfactory progenitors into chick hosts reveals migration and differentiation potentials dependent on cell commitment. AB - In vertebrates, interneurons of the olfactory bulb are continuously generated postnatally and throughout life at the subventricular zone of the forebrain. From there, the neuronal progenitors migrate tangentially in a typical chain-like structure to the olfactory bulb in which they differentiate as interneurons. We have used a mouse/chick xenograft strategy to explore the migration and differentiation potential of the mouse olfactory progenitors in a heterochronic and heterotypic environment. We compared the migration of primary cells derived from the subventricular zone of adult or newborn lateral ventricule with the behavior of in vitro amplified cells derived from the same structures. We show that in the chick environment, olfactory bulb progenitors from newborn brain tissue perform chain migration along the neural crest cell routes, whereas grafted neurosphere-derived-cells migrate as isolated cells. These results, together with in vitro observations, allow us to propose that neuronal chain migration is a community effect independent of environmental cues but which is closely regulated by the differentiation program of the cells. We established that the progenitor cells performing chain migration are already committed, while neurosphere-derived-cells are able to integrate and differentiate as components of the peripheral nervous system. PMID- 11273651 TI - Molecular determinants of metabotropic glutamate receptor 1B trafficking. AB - The metabotropic glutamate receptor mGluR1 undergoes alternative splicing to generate isoforms differing in C-terminal sequence. The mechanism by which these isoforms give different functional responses to agonists in vitro is so far unclear. Using the native mGluR1 and CD2-mGluR1 chimeric molecules, as well as their C-terminal truncations and mutants, we identified an endoplasmic reticulum (ER) retention signal Arg-Arg-Lys-Lys within the C-terminal sequence of mGluR1b. Its presence results in a much reduced cell surface expression of the receptor and chimeric molecules in cell lines and their restricted trafficking in neurones. This motif is also present in the C-terminus of mGluR1a, but its effect is overcome by a region of the mGluR1a-specific C-terminal sequence (amino acids 975-1098). Our results indicate that these splice variants of mGluR1 utilize different targeting pathways and suggest that this may be a general phenomenon in the metabotropic glutamate receptor gene family. PMID- 11273652 TI - Characterization of the deg-3/des-2 receptor: a nicotinic acetylcholine receptor that mutates to cause neuronal degeneration. AB - The nicotinic acetylcholine receptor family (nAChR) is a large family of acetylcholine-gated cation channels. Here we characterize the Caenorhabditis elegans DEG-3/DES-2 nAChR, a receptor identified due to its involvement in neuronal degeneration. Pharmacological analysis of a DEG-3/DES-2 receptor expressed in Xenopus oocytes shows that this receptor is preferentially activated by choline. This choline sensitivity of the DEG-3/DES-2 channel can explain its role in neuronal degeneration, as shown by the toxic effects of choline on oocytes expressing the mutant DEG-3/DES-2 channel. We also show that in C. elegans the DEG-3/DES-2 receptor is localized to nonsynaptic regions, including the sensory endings of chemosensory neurons. This localization is in agreement with a role for this receptor in chemosensation of choline, as inferred from a defect in chemotaxis for choline seen in deg-3 mutants. Thus, this work also provides evidence for the diversity of nonsynaptic activities associated with nAChRs. PMID- 11273654 TI - The Role of Personality Traits and Goal Orientations in Strategy Use. AB - The aim of this study was to contribute to the development of an integrated theory on individual learning differences. To that end, theories on learning styles, personality, and achievement motivation were combined in an explanatory model (tested with structural equation modelling). Goal orientations play an important role in this model, situated between personality traits and theories of intelligence, on the one hand, and learning strategy constructs (surface learning and deep learning), on the other. Surface-level strategies were related to entity theory beliefs and ego orientation as well as to conscientiousness, agreeableness, and effort orientation. Deep-level strategies were only directly related to task orientation and intellect. The relations found shed more light on what individual differences in learning consist of and help explain regularities in learning behavior. Copyright 2001 Academic Press. PMID- 11273655 TI - Effects of Headings on Text Summarization. AB - A summarization task was used to study whether headings influence readers' representations of the topic structure of a text. College students (Experiments 1 3) and sixth- and eighth-graders (Experiment 3) summarized a multiple topic text that (a) included headings introducing every new subtopic, (b) included headings introducing half of the new subtopics, or (c) included no headings. In all experiments, topics were more likely to be included in a summary if they were signaled than if they were not signaled. This effect was magnified when the text was only half signaled: Signaled topics were more likely to appear in a summary if only half the text topics were signaled than if all of the topics were signaled; however, unsignaled topics were less likely to appear in a summary if half of the text topics were signaled than if none of the text topics were signaled. The findings demonstrate that readers rely heavily on headings in a task that emphasizes attention to a text's topic structure. It is suggested that previously observed signaling effects on text recall are mediated by effects on how readers represent a text's topic structure. Copyright 2001 Academic Press. PMID- 11273653 TI - Abnormalities of synapses and neurons in the hippocampus of neuropsin-deficient mice. AB - In the present study, we produced null-mutant mice of neuropsin, an extracellular matrix serine protease, to examine the neural functions of this protein particularly in the hippocampus. Golgi-Cox impregnation and Nissl-staining revealed morphological change of cell soma in the mutant mice compared to wild type mice. However, Golgi-Cox impregnation revealed no apparent change in the dendritic arborization and spine density. Quantitative electronmicroscopic analysis revealed that number of asymmetrical synapses were significantly decreased in the stratum radiatum, the major terminal field of Schaffer collaterals, whereas free boutons still holding synaptic vesicles but with no synaptic specialization were increased in number in the same microscopic fields. An increased number of parvalbumin-immunoreactive cells (known as fast spiking cells) in mutant was also observed. These results strongly suggest that neuropsin is involved in connectivity of a group of CA1 synapses and consequently in the hippocampal networking. PMID- 11273656 TI - Beliefs about Learning in Children's Understanding of Science Texts. AB - This study examined elementary school children's beliefs about learning and assessed the influences of such beliefs on their understanding of science texts. Eighty-three children, 46 from Grade 4 and 37 from Grade 6, were administered a questionnaire on children's implicit notions of learning. Children were also asked to read a science text and complete several tasks that assessed their understanding of text information. Results indicated that older children were more likely to hold constructivist views of learning, and they also performed better than younger children on the text-processing tasks. As well, children's views of learning were significantly related to depth of text understanding when age effects were controlled. This study extends current research on epistemological beliefs of university and high school students. Implications of children's beliefs about learning and their roles in knowledge construction are discussed. Copyright 2001 Academic Press. PMID- 11273657 TI - Map Edges: Focal Points for Facilitating Text Recall. AB - In two experiments undergraduate students were shown maps with features located either along an edge or within the interior of the map. Next, participants were asked to read a related text. Thereafter, they were asked to recall as much of the text information as possible and to reconstruct the map. In both experiments, the results consistently showed that students recall significantly more information when features are located along the edges of the maps. These findings will help give teachers and designers of classroom displays a better understanding of how to create maps that will facilitate the recall of related information. Copyright 2001 Academic Press. PMID- 11273659 TI - Chinese Children's Incidental Learning of Word Meanings. AB - We investigated the effects of metalinguistic awareness and the internal structure of Chinese characters on children's incidental learning of word meanings while reading. The participants were 241 Taiwanese children from six fourth-grade classes. They were randomly assigned to read one of two texts, and then their knowledge of unfamiliar characters from both texts was assessed. Two kinds of factors that might influence incidental learning of character meanings were examined: features of the unfamiliar characters (radical helpfulness, phonetic regularity, and contextual support) and child characteristics (radical awareness, phonetic awareness, general vocabulary, and prior knowledge of target characters). Results showed that children could incidentally learn characters during normal reading. Children who had more radical or phonetic awareness knew more characters, especially when the character contained a radical that gave a clue to meaning. However, contrary to expectation, radical helpfulness and phonetic regularity did not contribute to character learning. Characters were easier to learn when contextual support was strong. Copyright 2001 Academic Press. PMID- 11273658 TI - Systematic Mathematical Errors and Cognitive Load. AB - The hypothesis that the intrinsic nature of algebraic bracket tasks causes an uneven distribution of cognitive load during computation was tested in three experiments with grades 8 and 9 students. In Experiment 1, students were given problems which required two successive brackets to be expanded; each bracket required two operations (computations) to be completed. It was discovered that more errors were made during the calculation of the second bracket than the first, and more errors were made during the second operation than the first operation within each bracket. Verbal protocols collected in Experiment 2 indicated that most errors were caused by failures in working memory rather than poorly learned rules. In Experiment 3, a dual-task methodology showed that the cognitive nature of brackets affected working memory performance. It was concluded that the cognitive load experienced by problem solvers on these tasks varied across operations and caused the observed error pattern. Copyright 2001 Academic Press. PMID- 11273660 TI - Nonverbal Displays as Indicants of Task Difficulty. AB - Accurately determining when students are having difficulty with cognitive tasks is important in educational settings. This study investigated whether college students emitted observable displays of cognitive difficulty when engaged in solitary problem-solving tasks. Participants high and low in self-monitoring tendencies were videotaped while solving both hard and easy problems. Ten-second segments of the videotapes were rated for displayed difficulty levels. Results indicate that college students do emit nonverbal displays indicating task difficulty: Students' displayed significantly less difficulty while solving easy problems than while solving hard problems. Results also indicated that the difficulty displays of low self-monitors were more discernible than the difficulty displays of high self-monitors. Copyright 2001 Academic Press. PMID- 11273661 TI - Frame of Reference Effects Following the Announcement of Exam Results. AB - The internal/external frame of reference (I/E) model by Marsh (e.g., 1990a) assumes two central information sources for the constitution of domain-specific academic self-concepts: (1) social comparisons (external frame of reference), in which students compare their own achievements with those of their classmates; and (2) intraindividual comparisons (internal frame of reference), in which students compare their own achievements in one subject with their achievements in other subjects. In path analyses, it has been found that the latter type of comparison leads to negative paths from achievement indicators in subject A to self-concept measures in subject B. To investigate the actual impact of achievement feedback and of the frames of reference on changes in self-concept variables, we analyzed math and German self-concepts immediately following the announcement of exam results in each subject. Participants were 258 7th- to 9th-graders. Path analyses using structural equation modeling supported the validity of the I/E model with respect to the impact of simultaneously given current exam results. In particular, German achievement had a negative impact on subsequent math self concept. Copyright 2001 Academic Press. PMID- 11273664 TI - Optic nerve changes in zinc-deficient rats. AB - In this study the optic nerve changes in zinc (Zn)-deficient rats are examined. Zinc is one of the essential trace elements and is known to be related to optic nerve diseases such as ethambutol neuropathy. However, the effect of Zn on the optic nerve has not been studied experimentally in animals. We used 3 week old weanling male Wistar Kyoto rats weighing 40-50 g. Rats were fed a Zn-deficient diet containing 0.007 mg of Zn per 100 g, all other nutrients and distilled and deionized water. The same water supplemented with 3 mg Zn per 100 g was given to the control group. After 4 or 7 weeks on a Zn-deficient diet, the optic nerve was examined with an electron microscope. A recovery group was fed a Zn-containing diet for 5 weeks after 7 weeks on the Zn-deficient diet. The serum Zn levels of the deficient group were significantly decreased at both 4 and 7 weeks. Most of the Zn-deficient rats showed hair loss around the eyes and on the extremities. Ultrastructural findings were as follows. The number of myelinated axons of Zn deficient rats at 4 and 7 weeks were significantly decreased and the myelin sheaths were significantly thinner in the Zn-deficient groups and in the recovery group. Unmyelinated axons were more numerous than in the control rats. Destruction of myelin and proliferation of glial cells were found in the optic nerves of Zn-deficient rats. This study suggests that the optic nerve needs Zn for the maintenance of its cell structure and even if Zn is supplied to the Zn deficient rats, destruction of the myelin structure may continue. Zn-deficiency induce a decrease of myelinated nerve fibers, and it is thought that optic neuropathy in patients treated with some drugs such as ethambutol may be a secondary change due to Zn-deficiency following drug administration. PMID- 11273665 TI - Down-regulation of Na-K-Cl cotransport by protein kinase C is mediated by protein phosphatase 1 in pigmented ciliary epithelial cells. AB - The role of protein phosphatases in the regulation of Na-K-Cl cotransport was examined in human pigmented ciliary epithelial (PE) cells. Both a 37 kDa form and a 72 kDa form of protein phosphatase 1 (PP1) could be immunologically detected. The protein phosphatase inhibitor calyculin A stimulated Na-K-Cl cotransport by 89 +/- 12% at 10 n M, whereas okadaic acid had no effect at concentrations less than 100 n M. Calyculin A had no significant effect on either Na-K ATPase or ouabain-insensitive, bumetanide-insensitive 86Rb+uptake. These data suggest that PP1 plays a role in the inhibition of Na-K-Cl cotransport in PE cells. Treatment of cells with phorbol 12-myristate, 13-acetate (PMA), a protein kinase C (PKC) activator caused an 82% inhibition of Na-K-Cl cotransport. When cells were first treated for 5 min with PMA, 10 n M calyculin A stimulated Na-K-Cl cotransport by 53% compared to 101% by calyculin A alone. Treatment of cells with PMA after stimulation of Na-K-Cl cotransport by calyculin A resulted in a prompt 56% drop in cotransport activity. These data suggest that maximal inhibition of Na-K-Cl cotransport by PKC requires PP1 activity, but that a part of PKCs inhibitory effect is independent of PP1. The effect of PKC activation on PP1 was further examined by determining PP1 activity in cells pretreated with PMA. PP1 activity increased 38+/-8% in cells exposed to 1 microM PMA for 5 min. This stimulation was blocked by 100 n M staurosporine or 1 microM bisindolylmaleimide, two PKC inhibitors. An isomer which does not activate PKC (4 alpha phorbol didecanoate), did not stimulate PP1 activity. Thus PKC activation leads to an increase in PP1 activity in PE cells. Pretreatment of cells with the protein kinase A (PKA) inhibitor PHI 14-22 resulted in a partial reduction in calyculin A stimulation of cotransport, suggesting that PP1 and PKA function in a kinase-phosphatase regulatory loop. To determine whether other protein kinases might also be involved, several protein kinase inhibitors were tested, including KT5823 (protein kinase G, type II-specific), KN62 (calmodulin activated kinase-specific) and ML7 (myosin light chain kinase-specific). None prevented activation of Na-K Cl cotransport by calyculin A, suggesting that these kinases are not involved in the activation of Na-K-Cl cotransport. PMID- 11273666 TI - Ligand-binding characterization of xanthophyll carotenoids to solubilized membrane proteins derived from human retina. AB - The macula of the human retina contains extraordinarily high concentrations of lutein and zeaxanthin, xanthophyll carotenoids that appear to play an important role in protecting against age-related macular degeneration, the leading cause of blindness among the elderly. It is likely that the uptake and stabilization of these carotenoids is mediated by specific xanthophyll-binding proteins. In order to purify and characterize such a binding protein, a carotenoid-rich membrane fraction derived from human macula or peripheral retina was prepared by homogenization, differential centrifugation, and detergent solubilization. Further purification was carried out using ion-exchange chromatography and gel filtration chromatography coupled with continuous photodiode-array monitoring for endogenously associated xanthophyll carotenoids. The most highly purified preparations contained two major protein bands at 25 and 55 kDa that consistently co-eluted with endogenous lutein and zeaxanthin. The visible absorbance spectrum of the binding protein preparation closely matches the spectral absorbance of the human macular pigment, and it is bathochromically shifted about 10 nm from the spectrum of lutein and zeaxanthin dissolved in organic solvents. Binding of exogenously added lutein and zeaxanthin is saturable and specific with an apparent Kd of approximately 1 microM. Canthaxanthin and beta-carotene exhibit no significant binding activity to solubilized retinal membrane proteins when assayed under identical conditions. Other potential mammalian xanthophyll-binding proteins such as albumin, tubulin, lactoglobulin and serum lipoproteins possess only weak non-specific binding affinity for carotenoids when assayed under the same stringent binding conditions. This investigation provides the first direct evidence for the existence of specific xanthophyll-binding protein(s) in the vertebrate retina and macula. The possible roles of xanthophyll-binding proteins in normal macular function and in the pathogenesis of age-related macular degeneration remain to be elucidated. PMID- 11273668 TI - A histomorphometric study of corneal endothelial cells in normal human fetuses. AB - The purpose of this study was to investigate the histomorphometric change in the normal development of human fetal corneal endothelial cells. Eighty one human fetal corneas, ranging from 12 to 40 weeks of gestation, were examined. For determination of gross parameters, corneal diameter and height were measured. Then the corneal endothelium including Descemet's membrane was stained with hematoxylin-eosin using a flat preparation method. In addition to histologic examination under the light microscope, computer-assisted image analysis was performed to determine the cell area, coefficient of variation in cell area and cell density, in both central and peripheral cornea, from each specimen. Total cell count per cornea was obtained by multiplying endothelial cell density by corneal surface area. Linear and nonlinear regression analysis of gestational age and each parameter were used to model corneal endothelial development during the prenatal period. Fetal cornea grows rapidly throughout the prenatal period. During the same period, mean cell area and total cell count also increases gradually, but there is a steep increase in the total cell count in the early period and of the cell area in the late period. The mean cell density decreases rapidly from 16 015 to 6167 cell x mm(-2). There was no significant difference in all parameters except cell density, between the central and peripheral cornea and the difference in cell density was only 2%. In the early prenatal period, there is a rapid increase of total cell count by mitosis, whereas in the late period enlarged endothelial cells cover the rapidly widening inner corneal surface without a significant change in the total cell count. PMID- 11273667 TI - Voltage-dependent calcium channels in the rat retina: involvement in NMDA stimulated influx of calcium. AB - Rises in intracellular Ca2+ induced by activation of glutamate receptors are of ultimate importance for neuronal excitability and pathophysiological processes. In the present study, we aimed to elucidate the types of voltage-dependent Ca2+ channels involved in the NMDA-stimulated influx of Ca2+ into the isolated rat retina by using selective blockers. Additionally, the number of binding sites for radioligands labelling L- ([3H]nitrendipine), N- ([125I]omega-conotoxin MVIIA) and P/Q-type ([125I]omega-conotoxin MVIIC) Ca2+ channels was assessed in the rat retina and, for further comparison, in the rat cortex. Incubation of isolated rat retinas with 100 microM NMDA produced a three-fold increase in the influx of 45Ca2+ that was completely blunted by MK-801, a NMDA receptor antagonist, and partially attenuated (approximately 20%) by tetrodotoxin, a Na+ channel blocker. The L-type Ca2+ channel blocker nifedipine reduced NMDA-stimulated Ca2+ influx in a dose-related fashion, with a maximum reduction of approximately 50%. Similar effects were observed with verapamil and diltiazem. Blockers of N- and P/Q-type Ca2+ channels had no significant effect on the influx of Ca2+ evoked by NMDA. Co2+, a non-specific Ca2+ channel blocker, caused an inhibition of NMDA stimulated Ca2+ influx similar to that of nifedipine. Therefore, of all voltage dependent Ca2+ channels, L-type channels appear to make the greatest contribution (up to 50%) to the NMDA-stimulated influx of Ca2+ into the isolated rat retina. This finding contrasts with evidence obtained in brain neurones supporting a role for L-, N- and P/Q-type channels in NMDA-evoked Ca2+ signals. A comparison of the number of radioligand binding sites associated with L-, N- or P/Q-type Ca2+ channels in the rat cortex and retina revealed that such a difference cannot be ascribed to a distinct expression pattern of these channels in both tissues, although some variations were found. Interestingly, a different affinity of [3H]nitrendipine for L-type Ca2+ channels in the rat retina and cortex was observed which may reflect the expression of different classes of L-type channels in these tissues. The ability of L-type Ca2+ channel blockers to attenuate NMDA stimulated Ca2+ influx may underlie their neuroprotective effects in the retina. PMID- 11273669 TI - The presence of a human UV filter within the lens represents an oxidative stress. AB - It has recently been demonstrated that, with age, UV filters such as 3 hydroxykynurenine glucoside, bind to proteins in the human lens. This covalent interaction leads to colouration of the normal lens, and results from the instability of the kynurenine side chain. Other primate UV filters, in addition to containing the same side chain, can also be readily oxidized. One such compound is 3-hydroxykynurenine (3OHKyn). It has been proposed that oxidation of bound and/or free UV filters, such as 3OHKyn may give rise to the lens colouration associated with age-related nuclear cataract. Therefore it has become important to understand the oxidation of 3OHKyn within the lens. In this study, intact bovine lenses (which lack UV filters) were incubated with 3OHKyn and various lens parameters monitored. The effect of exposure to hyperbaric oxygen (HBO) was also assessed, both alone, and in combination with prior 3OHKyn incubation. Glutathione (GSH), protein sulfhydryl and protein-bound sulfhydryl levels, as well as soluble protein content and gel filtration profiles, were obtained for cortical and nuclear regions after defined periods of incubation. The presence of the primate UV filter, 3OHKyn, at concentrations similar to those present in the human lens, was shown to produce considerable oxidative stress within the lens, as judged by its effect on GSH. This effect was noted under normobaric conditions, but was exacerbated by increased oxygen. Exposure of lenses to HBO caused a marked fall in GSH in cortical and nuclear regions. This effect was exaggerated in the presence of 3OHKyn. HBO treatment also lead to a fall in protein sulfhydryl content, however, this was only partial (approximately 1 mol SH per mol protein) and changed only slowly, even with extended periods of exposure to HBO, suggesting that most crystallin sulfhydryl groups may be buried. 3OHKyn did not appreciably affect this oxidation although it did cause an increase in the level of protein-bound sulfhydryl. HBO treatment produced a more than two-fold increase in protein-bound sulfhydryl content in the cortex. There was little influence of 3OHKyn alone on protein solubility, even with extended periods of incubation, however, incubation for 72 hr in the presence of HBO caused a significant increase in insoluble protein particularly in the nucleus. This insolubilization was further increased in the presence of 3OHKyn. FPLC profiles showed that the proportion of gamma and beta crystallins in the soluble fraction decreased following HBO, suggesting that these may be involved in disulfide bond formation. This study demonstrates that a readily oxidized compound, such as the primate UV filter 3OHKyn, represents an oxidative stress within the lens and that such oxidative processes can be exacerbated if the concentration of oxygen within the lens is increased. We speculate that this factor may account for the evolution of unusually high levels of glutathione reductase in human lenses. PMID- 11273670 TI - Age-dependent changes in the expression of matrix components in the mouse eye. AB - Although the presence of 'cartilage-specific' collagens in the eye has been documented earlier, very little is known about their synthesis rates during ocular development, growth and aging. The purpose of the present study was to follow changes in the mRNA levels and distribution of key components of the extracellular matrix in the eyes of normal and transgenic Del1 mice, harboring a short deletion mutation in the type II collagen gene, during ocular growth and aging. Total RNAs extracted from mouse eyes were studied by Northern analysis for mRNA levels of type I, II, III, VI, IX and XI collagens, biglycan, fibromodulin and decorin. A predominant finding of the present study was the marked reduction in the mRNA levels of type I and II collagens in the eye upon aging. The changes in the mRNA levels of type III and VI collagen and proteoglycans were smaller. Localization of type II and IX collagen in the eye was performed by immunohistochemistry. Despite the reduction in the type II collagen mRNA levels, immunohistochemistry confirmed widespread distribution of the protein also in aging mouse eyes, suggesting its slow turnover. Although the Del1 mutation caused gradual degenerative lesions in the eyes, the distribution of the protein remained essentially unchanged. The widespread distribution and marked downregulation of type II collagen production in the mouse eye upon aging probably explain the gradual development of degenerative lesions, particularly in the eyes of transgenic Del1 mice, where production of mutant type II collagen chains also contributes to the process. PMID- 11273671 TI - Enzymatic, clinical and histologic evaluation of corneal tissues in experimental fungal keratitis in rabbits. AB - Mycotic keratitis, being frequently refractive to most of the currently available antifungal therapy, continues to pose a therapeutic challenge to the clinician. In keratitis of infectious etiology stromal dissolution may be brought about by a combination of agent and host factors. An understanding of the source and nature of corneal tissue damage is essential for evolving more effective therapeutic modalities in the treatment of fungal keratitis. In the present study, we have characterized the extracellular proteases produced in vitro by corneal fungal pathogens namely the Aspergillus flavus and Fusarium solani when collagen was provided as the sole nitrogen source. In addition, fungal infected rabbit corneas were investigated for proteolytic activities and nature of inflammatory reaction. Gelatin zymography detected protease bands with molecular mass ranging from 100 to 200 kDa in the culture extracts of A. flavus, and a single major band of molecular mass approximately 200 kDa in the culture extracts of F. solani. A basal proteolytic activity of mass 65 kDa was visualized in all uninfected and infected rabbit corneal extracts. Infected corneas in addition revealed the presence of additional proteolytic species of mass 92 and 200 kDa. The enzyme inhibitory profile suggested that fungal cultures in vitro contained predominantly serine protease activity and to a lesser extent metalloprotease activity. However, fungal infected corneal homogenates showed the presence of metalloproteinase activity alone, the enzymatic activities entirely being sensitive to ethylene diamine tetra acetate (EDTA), a metalloprotease inhibitor. Interestingly, the serine proteolytic activity detected in fungal cultures in vitro was not present in the fungal infected corneas in vivo. However, the possible role of fungal serine proteases in the activation of corneal matrix metalloproteinases (MMPs) cannot be ruled out. Based on the criteria of molecular mass, proteolytic activity in the presence of calcium at neutral pH, and sensitivity to inhibition by a metalloprotease inhibitor, the 65 and 92 kDa gelatinases were identified as MMP 2 and MMP 9, respectively. The expression of 92 and 200 kDa gelatinases correlated positively with the amount of polymorphonuclear cells present in the infected tissues. Activated resident corneal cells or inflammatory cells may largely contribute to the increased proteolytic activities in fungal infected corneas resulting in tissue matrix degradation in fungal keratitis. PMID- 11273672 TI - Involvement of L-DNase II in nuclear degeneration during chick retina development. AB - During the development of the neural retina, 50% of the neurons die physiologically by apoptosis. In the chick embryo, the apoptotic wave starts at E8 and ends at E18, with a peak at E11. The onset of apoptosis is accompanied by the activation of several degradative enzymes. Among these, the activation of the endonucleases leads to the degradation of the genomic DNA of the cell which is thought to be the final event in apoptosis. Here, we have investigated the endonucleases activated during apoptosis associated with retinal development. We have found that Ca2+-Mg2+-dependent endonucleases, as well as acid endonucleases are activated. The results obtained in vitro using purified nuclei from chicken retina indicate that the endonuclease activity resulting from the activation of L DNase II, an acid DNase is responsible for most of the DNA degradation observed in these cells. PMID- 11273673 TI - Stress fiber formation is required for matrix reorganization in a corneal myofibroblast cell line. AB - Corneal wound healing fibroblasts (myofibroblasts) develop a muscle-like contractile apparatus composed of prominent microfilament bundles (stress fibers) and express alpha-smooth muscle actin (alpha-SMA). In this study, gelsolin, an actin filament-severing protein, was overexpressed in a alpha-SMA-expressing corneal myofibroblast cell line (TRK43) to assess whether intact stress fibers are required for in vitro matrix organization and wound contraction. Stably integrated gelsolin was introduced by electroporation of an expression construct (pREPCG8) into cultured cells. Thirty-seven clones were isolated with half of the clones showing a fibroblastic phenotype while the remaining half appeared epithelioid. One fibroblastic clone, GS56, and one epithelioid clone, GS44, were selected for detailed characterization. The GS56 cells appeared highly elongated and spindle-shaped and had prominent stress fibers and focal adhesions. GS44 cells showed disruption of stress fibers and a cortical f-actin organization as well as the down regulation of alpha-SMA expression by immunocytochemistry and Western blotting. Both phenotypes showed enhanced gelsolin expression; however, fractionation of cell extracts demonstrated differences in the subcellular distribution of gelsolin with GS44 cells having markedly reduced and GS56 cells having markedly increased cytoskeletal gelsolin. In an in vitro wound contraction assay, epithelioid GS44 cells showed a significantly impaired ability to contract a collagen matrix compared to that of TRK43 cells, CT9 or GS56 transfectants. Loss of stress fibers in GS44 cells also correlated with enhanced cell motility. Together, these results demonstrate that the ability to form microfilament bundles or stress fibers is required for matrix organization and contraction by corneal myofibroblasts. Although no clear explanation is available, we suspect that differences in gene insertion of the gelsolin overexpression vector may have led to differential intercellular localization of gelsolin and its effect on stress fiber formation in the two cell lines. PMID- 11273674 TI - Retinal degeneration following failed photoreceptor maturation in 5A11/basigin null mice. AB - 5A11/Basigin is a member of the immunoglobulin gene superfamily which plays an important role in cell-cell interactions in the developing neural retina. These studies were initiated to investigate the distribution of 5A11/Basigin within the mouse retina, as well as the cytoarchitectural and biochemical effects on the retina after the inactivation of the 5A11/Basigin gene in a mouse strain. Immunocytochemical analyses indicated that mouse 5A11/Basigin is located on the surface of Muller cells, the apical and basal surfaces of the retinal pigmented epithelium, and blood vessels. Lower expression levels were found on photoreceptor cell bodies and a portion of the inner segments. Inactivation of the 5A11/Basigin gene in mice resulted in the failure of photoreceptor cells to fully mature. This failed development eventually lead to the degeneration, death and removal of most of the photoreceptors several months after birth. Biochemical analyses indicated that expression of Muller cell specific proteins, including glutamine synthetase and carbonic anhydrase-II, was not effected; however, opsin protein expression never achieved normal adult levels in the 5A11/Basigin null mice. Also, 5A11/Basigin null retinas were considered 'reactive' based on elevated glial fibrillary acidic protein expression. The results presented here suggest that 5A11/Basigin expression on Muller cells and/or the retinal pigmented epithelium is necessary for photoreceptor outer segment biochemical development and structural maintenance. However, the exact role that 5A11/Basigin plays during retinal development remains to be determined. PMID- 11273675 TI - Latanoprost exerts neuroprotective activity in vitro and in vivo. AB - Prostaglandins may influence cyclo-oxygenase (COX-2) and nitric oxide (NO) synthase activity, thus interfering with ischemia-induced neurotoxic processes. The prostaglandin synthetic derivative, latanoprost was tested in different in vivo and in vitro models of neuronal damage in order to study its influence on these processes. Ischemia was induced in rats by bilateral occlusion of the carotid arteries for 30 min. Latanoprost (0.01 mg x kg(-1)per die, i.p. for 3 days) or the ionotropic glutamate receptors antagonist, MK-801 (0.1 mg x kg( 1)per die, i.p. for 3 days) were equal in preventing lactate accumulation in retinal tissue of animals subjected to acute ischemia. Similar results were obtained in animals with retinal ischemia induced by increasing intraocular pressure to 120 mm Hg for 45 min. PGF2alpha, PGE2, latanoprost and acid of latanoprost (PhXA85) reduced the release of LDH from primary cultures of human retinal cells in vitro subjected to glutamate (10 microM) or hypoxia/re oxygenation exposure. This effect was observed only at concentrations of 1-0.01 microM for PGF2alpha and PGE2, and of 0.1-0.001 microM for latanoprost (0.01 microM-0.1 nM for PhXA85). The COX-2 activity in cultured retinal cells exposed to glutamate was measured as PGE2 production when latanoprost was applied compared to arachidonic acid (AA) at different molar concentrations. The COX-2 activity was reduced by arachidonic acid (0.1-0.01 microM) as well as by latanoprost (0.1-0.001 microM) and PhXA85 (0.01-0.001 microM) in retinal cells exposed to glutamate. Inhibition of inducible NO synthase was also found with the same drug concentrations. These results suggest that latanoprost exerts a neuroprotective activity in vitro and in vivo. This effect seems to be present only at low concentrations of the drug. A negative feedback on neuronal COX-2 activity may be possibly involved. PMID- 11273676 TI - Melatonin synthesis in the rat harderian gland: age- and time-related effects. AB - The Harderian gland is considered as an extrapineal source of melatonin. In the pineal gland, melatonin is known to present a circadian rhythm with high concentration during nighttime in all species studied. We determined in Wistar male rats the effects of age and time of day on melatonin synthesis in the Harderian gland. We compared Harderian gland melatonin content and the hormone synthesizing enzymes, serotonin N-acetyltransferase and hydroxyindole-O methyltransferase, in young (4 months) and old (22 months) animals at six circadian stages and found that melatonin synthesis in the Harderian gland was unaffected by age. We also studied the Wistar rat Harderian gland at ten different circadian stages and found that the Harderian gland did not exhibit a daily rhythm in its melatonin content. This study shows that, by contrast to the pineal gland, melatonin in Wistar rat Harderian gland does not exhibit daily variations and that aging does not affect the melatonin content of the gland. PMID- 11273678 TI - Neurospora at the millennium. PMID- 11273677 TI - Map kinases in fungal pathogens. AB - MAP kinases in eukaryotic cells are well known for transducing a variety of extracellular signals to regulate cell growth and differentiation. Recently, MAP kinases homologous to the yeast Fus3/Kss1 MAP kinases have been identified in several fungal pathogens and found to be important for appressorium formation, invasive hyphal growth, and fungal pathogenesis. This MAP kinase pathway also controls diverse growth or differentiation processes, including conidiation, conidial germination, and female fertility. MAP kinases homologous to yeast Slt2 and Hog1 have also been characterized in Candida albicans and Magnaporthe grisea. Mutants disrupted of the Slt2 homologues have weak cell walls, altered hyphal growth, and reduced virulence. The Hog1 homologues are dispensable for growth but are essential for regulating responses to hyperosmotic stress in C. albicans and M. grisea. Overall, recent studies have indicated that MAP kinase pathways may play important roles in regulating growth, differentiation, survival, and pathogenesis in fungal pathogens. PMID- 11273680 TI - Polarity-defective mutants of Aspergillus nidulans. AB - We have identified two polarity-defective (pod) mutants in Aspergillus nidulans from a collection of heat-sensitive lethal mutants. At restrictive temperature, these mutants are capable of nuclear division but are unable to establish polar hyphal growth. We cloned the two pod genes by complementation of their heat sensitive lethal phenotypes. The libraries used to clone the pod genes are under the control of the bidirectional niaD and niiA promoters. Complementation of the pod mutants is dependent on growth on inducing medium. We show that rescue of the heat-sensitive phenotype on inducing media is independent of the orientation of the gene relative to the niaD or niiA promoters, demonstrating that the intergenic region between the niaD and the niiA genes functions as an orientation independent enhancer and repressor that is capable of functioning over long distances. The products of the podG and the podH genes were identified as homologues of the alpha subunit of yeast mitochondrial phenylalanyl--tRNA synthetase and transcription factor IIF interacting component of the CTD phosphatase. Neither of these gene products would have been predicted to produce a pod mutant phenotype based on studies of cellular polarity mutants in other organisms. The implications of these results are discussed. PMID- 11273679 TI - The phylogenetics of mycotoxin and sclerotium production in Aspergillus flavus and Aspergillus oryzae. AB - Aspergillus flavus is a common filamentous fungus that produces aflatoxins and presents a major threat to agriculture and human health. Previous phylogenetic studies of A. flavus have shown that it consists of two subgroups, called groups I and II, and morphological studies indicated that it consists of two morphological groups based on sclerotium size, called "S" and "L." The industrially important non-aflatoxin-producing fungus A. oryzae is nested within group I. Three different gene regions, including part of a gene involved in aflatoxin biosynthesis (omt12), were sequenced in 33 S and L strains of A. flavus collected from various regions around the world, along with three isolates of A. oryzae and two isolates of A. parasiticus that were used as outgroups. The production of B and G aflatoxins and cyclopiazonic acid was analyzed in the A. flavus isolates, and each isolate was identified as "S" or "L" based on sclerotium size. Phylogenetic analysis of all three genes confirmed the inference that group I and group II represent a deep divergence within A. flavus. Most group I strains produced B aflatoxins to some degree, and none produced G aflatoxins. Four of six group II strains produced both B and G aflatoxins. All group II isolates were of the "S" sclerotium phenotype, whereas group I strains consisted of both "S" and "L" isolates. Based on the omt12 gene region, phylogenetic structure in sclerotium phenotype and aflatoxin production was evident within group I. Some non-aflatoxin-producing isolates of group I had an omt12 allele that was identical to that found in isolates of A. oryzae. PMID- 11273681 TI - Development and characterization of a genetic linkage map of Cryptococcus neoformans var. neoformans using amplified fragment length polymorphisms and other markers. AB - A segregating population of single basidiospore isolates from a sexual cross was used to generate the first moderately dense genetic linkage map of Cryptococcus neoformans var. neoformans (Serotype D). Polymorphic DNA markers were developed using amplified fragment length polymorphisms, random amplified polymorphic DNA, and gene-encoding sequences. These markers were used to analyze 100 meiotic progeny. All markers were tested for distorted segregation with a goodness of fit test. Of the total of 181 markers, 148 showed balanced (1:1) segregation ratios. Segregation distortion was observed for 33 markers. Based on all the markers, a linkage map was generated that consists of 14 major linkage groups with 127 markers, several small linkage groups, and 2 linkage groups that consist only of highly skewed markers. The genetic distance of the linkage map is 1356.3 cM. The estimated total haploid genome size for C. neoformans var. neoformans was calculated using Hulberts method and yielded a map size of 1917 cM. The number of major linkage groups correlates well with the proposed number of 13 chromosomes for C. neoformans var. neoformans. Several genes, including CAP64, CnLAC, and the mating-type locus, were mapped, and their associations were consistent with published data. To date, 6 linkage groups have been assigned to their corresponding chromosomes. This linkage map should provide a framework for the ongoing genome sequencing project and will be a useful tool for studying the genetics and pathogenicity of this important medical yeast. PMID- 11273682 TI - Hex-1, a gene unique to filamentous fungi, encodes the major protein of the Woronin body and functions as a plug for septal pores. AB - We have identified a gene, named hex-1, that encodes the major protein in the hexagonal crystals, or Woronin bodies, of Neurospora crassa. Analysis of a strain with a null mutation in the hex-1 gene showed that the septal pores in this organism were not plugged when hyphae were damaged, leading to extensive loss of cytoplasm. When grown on agar plates containing sorbose, the hex-1(-) strain showed extensive lysis of hyphal tips. The HEX-1 protein was predicted to be 19,125 Da. Analysis of the N-terminus of the purified protein indicated that 16 residues are cleaved, yielding a protein of 17,377 Da. A polyclonal antibody raised to the HEX-1 protein recognized multiple forms of the protein, apparently dimers and tetramers that were resistant to solubilization by sodium dodecyl sulfate and reducing reagents. Treatment of the protein with phosphatase caused dissociation of these oligomers. Preparations enriched in Woronin bodies contained catalase activity, which was not detected in comparable fractions from the hex-1(-) mutant strain. These results support the hypothesis that the Woronin body is a specialized peroxisome that functions as a plug for septal pores. PMID- 11273683 TI - Differential expression of chitin synthase III and IV mRNAs in ascomata of Tuber borchii Vittad. AB - A full-length genomic clone encoding a class III chitin synthase (CHS) and one DNA fragment corresponding to a class IV CHS were isolated from the mycorrhizal fungus Tuber borchii and used for an extensive expression analysis, together with a previously identified DNA fragment corresponding to a class II CHS. All three Chs mRNAs are constitutively expressed in vegetative mycelia, regardless of the age, mode of growth, and proliferation capacity of the hyphae. A strikingly different situation was observed in ascomata, where class III and IV, but not class II, mRNAs are differentially expressed in a maturation stage-dependent manner and accumulate, respectively, in sporogenic and vegetative hyphae. These data, the first on the expression of distinct Chs mRNAs during fruitbody development, point to the different cellular roles that can be played by distinct chitin synthases in the differentiation of spores of sexual origin (CHS III) or in ascoma enlargement promoted by the growth of vegetative hyphae (CHS IV). PMID- 11273684 TI - Microtubules Are required for motility and positioning of vesicles and mitochondria in hyphal tip cells of Allomyces macrogynus. AB - We have used video-enhanced light microscopy and digital image processing to characterize the intracellular motility and positioning of vesicles ( approximately 1-microm diameter) and mitochondria in growing hyphal tip cells of Allomyces macrogynus. These observations were coupled with cytoskeletal inhibitory experiments to define the roles of the microtubule and actin cytoskeletons in organelle translocation and positioning. Vesicles and mitochondria were abundant in apical and subapical hypha regions. Vesicles traveled along paths that were parallel to the longitudinal axis of the cell. Anterograde (i.e., toward the hyphal apex) and retrograde (i.e., away from the hyphal apex) movements of vesicles occurred at average rates of 4.0 and 2.2 microm/s, respectively. Bidirectional travel of vesicles along common paths was noted in the cortical cytoplasm. Mitochondria were aligned mostly parallel to the long axis of the hypha, except those extending into the hyphal apex, which were oriented toward the Spitzenkorper. In regions of the subapical hypha mitochondria were often restricted to the cortical cytoplasm and nuclei occupied the central cytoplasmic region. Mitochondria displayed rapid anterograde movements reaching speeds of 3.0 microm/s, but primarily maintained a constant position relative to either the advancing cytoplasm or the lateral cell wall. Cytoskeletal disruption experiments showed that the positioning of mitochondria and motility of vesicles and mitochondria were microtubule-based and suggested that the actin cytoskeleton played uncertain roles. PMID- 11273687 TI - Identification of symbiotic bacteria (Photorhabdus and Xenorhabdus) from the entomopathogenic nematodes Heterorhabditis marelatus and Steinernema oregonense based on 16S rDNA sequence. AB - Two species of entomopathogenic nematodes, Heterorhabditis marelatus and Steinernema oregonense, were described recently from the west coast of North America. It is not known whether the bacterial symbionts of these nematodes are also unique. Here we compared partial 16S rRNA sequences from the symbiotic bacteria of these two nematodes with sequence from previously described Photorhabdus and Xenorhabdus species. The 16S sequence from the new Xenorhabdus isolate appears very similar to, although not identical to, that of X. bovienii, the common symbiont of S. feltiae. The new Photorhabdus isolate appears to be very distinct from other known Photorhabdus species, although its closest affinities are with the P. temperata group. We also verified a monoxenic association between each isolate and its nematode by amplifying and sequencing bacterial 16S sequence from crushed adult and juvenile nematodes and from bacterial cultures isolated from infected hosts. PMID- 11273688 TI - The pathogenicity of Bacillus thuringiensis ssp. kurstaki to gamma-irradiated Cadra cautella (Walker) (Lepidoptera: Phycitidae). AB - The present investigation deals with the effects of Bacillus thuringiensis ssp. kurstaki on various biological parameters of gamma-irradiated Cadra cautella. The pathogen, irradiation, and their combinations significantly affected the insects by increasing their mortality and developmental periods, reducing the pupal and adult survival (%), and the adult longevity and reproductive potential were also significantly reduced. It was observed that irradiation-pathogen combinations produced additive to synergistic effects on C. cautella. PMID- 11273689 TI - Effects of exogenous nutrients on conidial germination and virulence against the silverleaf whitefly for two hyphomycetes. AB - Exogenous protein and sugar sources were tested for their impact on conidial germination of two silverleaf whitefly (Bemisia argentifolii) pathogens: Beauveria bassiana and Paecilomyces fumosoroseus. In liquid culture, sugars stimulated only 5-27% germination of B. bassiana and < or =11% germination of P. fumosoroseus, whereas, yeast extract or peptone stimulated 95-100% germination. In the absence of additional nutrients, agar alone stimulated approximately 50% germination. Storing spores for different periods of time did not alter their general response to exogenous nutrients. When spores were germinated before being applied to third instar B. argentifolii, mortality was as much as 2.45 times greater and occurred more rapidly than that for fresh spores. For ungerminated conidia, the mean time to death from infection was 5.45 (SE = 0.16) and 4.74 (SE = 0.08) days for application rates of 37 and 144 conidia x mm(-2), respectively. When conidia were germinated before application, infection times dropped to 4.58 (SE = 0.16) and 4.45 (SE = 0.10) days, respectively. A likely explanation for the greater pathogenicity and virulence of germinated over ungerminated B. bassiana conidia is that only a fraction of the spores applied to whitefly nymphs actually germinate on the cuticle. For some specialized applications, such as greenhouse production systems, it may be beneficial to germinate spores immediately prior to application. PMID- 11273690 TI - Histology of herniations through the body wall and cuticle of zooplankton from the Laurentian Great Lakes. AB - Zooplankton of the Laurentian Great Lakes developed hernial protrusions whose gross appearance matches those on zooplankton described elsewhere in the world. We have carried out a histologic and cytologic analysis of the protrusions and found that they are composed of apparently degenerating or necrotic tissue(s) that has been expressed from the organism through the process of herniation. At their base the protrusions are continuous with viable tissue(s) within the organism through a fissure in the exoskeleton. Our observations lead us to suspect that these hernial protrusions are lethal. The development of such protrusions in zooplankton may be a worldwide phenomenon, but the cause of the herniation remains a mystery. PMID- 11273691 TI - Disease prevalence and transmission of Microsporidium phytoseiuli infecting the predatory mite, Phytoseiulus persimilis (Acari: Phytoseiidae). AB - Isolated colonies of the predatory mite, Phytoseiulus persimilis, were used to gain information regarding prevalence and transmission of Microsporidium phytoseiuli. Two colonies of P. persimilis were reared on spider mite (Tetranychus urticae)-infested bean plants in isolated cages. Disease prevalence of predators from Colony 1 remained relatively low (between 0 and 15%) over 57 weeks of observation whereas disease prevalence of predators from Colony 2 increased over 3 months (from 12 to 100%). Disease prevalence among predators from Colony 1 had increased to 100% 2 months after weekly sampling had ceased for this colony and periodic sampling confirmed that disease prevalence among individuals of both colonies remained at 100%. Microsporidian spores were not detected in randomly chosen samples of T. urticae prey mites that were removed and examined biweekly during this period. Although numerous microsporidian spores were observed in smear preparations of fecal pellets examined by light microscopy, spores were not observed on leaf surfaces or predator feces when examined by SEM. The latter appeared as intact aggregates composed of numerous dumbbell-shaped crystals and it is unlikely that spores are liberated from intact fecal pellets onto leaf surfaces. Vertical transmission of M. phytoseiuli was 100%; horizontal transmission was low (14.3%) and occurred only when immature P. persimilis were permitted to develop in contact with infected immature and adult predators. The mean number of eggs produced per mated pair was highest when uninfected females were mated with uninfected males (63.2 eggs per mated pair). Although mean egg production decreased when one or both parents were infected, not all differences were significant. Male predatory mites did not contribute to infection of their progeny. Results suggest that routine examination of P. persimilis for microsporidian spores is essential for the management of M. phytoseiuli within P. persimilis colonies. Low disease prevalence and lack of obvious disease signs or symptoms, as in the case of M. phytoseiuli, increase the probability that these pathogens will escape notice unless individuals are routinely examined for pathogens. PMID- 11273692 TI - The association of Western flower thrips, Frankliniella occidentalis, with a near Erwinia species gut bacteria: transient or permanent? AB - Associations between insects and gut bacteria are ubiquitous. It is possible to make a distinction between permanent associations (called symbiosis), in which the same type of bacteria is present in more than one generation of the insect, and transient associations. Transient bacteria are ingested together with food but do not settle in the insect gut in such a way that they will be passed on to the next generation. In this study, we describe the permanent association between Western flower thrips (Frankliniella occidentalis), a polyphagous insect species that is a major pest worldwide, and one type of gut bacteria. On the basis of direct microscopic observations and counts of bacteria, it was found that thrips from the populations studied contained large numbers of bacteria in their hindgut. Bacteria were isolated from their host and grown on 10 different agar media. The number of bacteria isolated on agar media equaled the number of direct counts. All isolates had the same colony morphology. On the basis of their 16S rDNA sequence these bacteria were identified as Enterobacteriaceae, closely related to Escherichia coli. Isolates tested with API 20E biochemical tests were Erwinia species. This was the only type of bacteria found in all thrips individuals on any of the 10 different agar media. Universal primers, which would potentially pick up DNA from any bacterium present in the insect, were applied on crude DNA extracts from thrips with bacteria. We only found 16S rDNA sequences similar to those of the isolated thrips gut bacteria. The same type of bacteria was present in all life stages of the thrips and was found to persist in the thrips populations for at least 2 years (more than 50 generations). PMID- 11273693 TI - Growth and transmission of gut bacteria in the Western flower thrips, Frankliniella occidentalis. AB - The Western flower thrips (Frankliniella occidentalis), a polyphagous insect with global distribution, has a permanent association with a near Erwinia species TAC bacterium in its hindgut. Since this bacterium is able to grow outside the thrips, it is a facultative symbiont that is not completely dependent on the host. In this study we address the question of how the association is maintained and how bacteria are transmitted to newly hatched thrips larvae. Bacteria are passed on to new thrips via the food source. No evidence was found for vertical transmission from mother to offspring via the egg. Gut bacteria show unlimited growth during the larval (feeding) stages, and in the second instar stage 100% of the larvae become infected with high numbers of bacteria. In the prepupal and pupal stage, the number of bacteria declines, but increases again during the adult phase. A method to rear aposymbiotic (bacteria-free) thrips is described which enables studies on the impact of bacteria on the fitness of thrips. PMID- 11273694 TI - Ultrastructural findings on microsporidia spore wall as seen by ruthenium tetroxide fixation. PMID- 11273695 TI - A simple method for routine maintenance and preservation of entomophthoraceous cultures. PMID- 11273696 TI - Replication of insect iridescent virus 6 in a whitefly cell line. PMID- 11273697 TI - Troponin organization on relaxed and activated thin filaments revealed by electron microscopy and three-dimensional reconstruction. AB - The steric model of muscle regulation holds that at low Ca(2+) concentration, tropomyosin strands, running along thin filaments, are constrained by troponin in an inhibitory position that blocks myosin-binding sites on actin. Ca(2+) activation, releasing this constraint, allows tropomyosin movement, initiating actin-myosin interaction and contraction. Although the different positions of tropomyosin on the thin filament are well documented, corresponding information on troponin has been lacking and it has therefore not been possible to test the model structurally. Here, we show that troponin can be detected on thin filaments and demonstrate how its changing association with actin can control tropomyosin position in response to Ca(2+). To accomplish this, thin filaments were reconstituted with an engineered short tropomyosin, creating a favorable troponin stoichiometry and symmetry for three-dimensional analysis. We demonstrate that in the absence of Ca(2+), troponin bound to both tropomyosin and actin can act as a latch to constrain tropomyosin in a position on actin that inhibits actomyosin ATPase. In addition, we find that on Ca(2+) activation the actin-troponin connection is broken, allowing tropomyosin to assume a second position, initiating actomyosin ATPase and thus permitting contraction to proceed. PMID- 11273699 TI - Expanding the genetic code: selection of efficient suppressors of four-base codons and identification of "shifty" four-base codons with a library approach in Escherichia coli. AB - Naturally occurring tRNA mutants are known that suppress +1 frameshift mutations by means of an extended anticodon loop, and a few have been used in protein mutagenesis. In an effort to expand the number of possible ways to uniquely and efficiently encode unnatural amino acids, we have devised a general strategy to select tRNAs with the ability to suppress four-base codons from a library of tRNAs with randomized 8 or 9 nt anticodon loops. Our selectants included both known and novel suppressible four-base codons and resulted in a set of very efficient, non-cross-reactive tRNA/four-base codon pairs for AGGA, UAGA, CCCU and CUAG. The most efficient four-base codon suppressors had Watson-Crick complementary anticodons, and the sequences of the anticodon loops outside of the anticodons varied with the anticodon. Additionally, four-base codon reporter libraries were used to identify "shifty" sites at which +1 frameshifting is most favorable in the absence of suppressor tRNAs in Escherichia coli. We intend to use these tRNAs to explore the limits of unnatural polypeptide biosynthesis, both in vitro and eventually in vivo. In addition, this selection strategy is being extended to identify novel five- and six-base codon suppressors. PMID- 11273698 TI - Solution structure of a type I dockerin domain, a novel prokaryotic, extracellular calcium-binding domain. AB - The type I dockerin domain is responsible for incorporating its associated glycosyl hydrolase into the bacterial cellulosome, a multienzyme cellulolytic complex, via its interaction with a receptor domain (cohesin domain) of the cellulosomal scaffolding subunit. The highly conserved dockerin domain is characterized by two Ca(2+)-binding sites with sequence similarity to the EF-hand motif. Here, we present the three-dimensional solution structure of the 69 residue dockerin domain of Clostridium thermocellum cellobiohydrolase CelS. Torsion angle dynamics calculations utilizing a total of 728 NOE-derived distance constraints and 79 torsion angle restraints yielded an ensemble of 20 structures with an average backbone r.m.s.d. for residues 5 to 29 and 32 to 66 of 0.54 A from the mean structure. The structure consists of two Ca(2+)-binding loop-helix motifs connected by a linker; the E helices entering each loop of the classical EF-hand motif are absent from the dockerin domain. Each dockerin Ca(2+)-binding subdomain is stabilized by a cluster of buried hydrophobic side-chains. Structural comparisons reveal that, in its non-complexed state, the dockerin fold displays a dramatic departure from that of Ca(2+)-bound EF-hand domains. A putative cohesin-binding surface, comprised of conserved hydrophobic and basic residues, is proposed, providing new insight into cellulosome assembly. PMID- 11273700 TI - The complete nucleotide sequence of a plant root-inducing (Ri) plasmid indicates its chimeric structure and evolutionary relationship between tumor-inducing (Ti) and symbiotic (Sym) plasmids in Rhizobiaceae. AB - The Ri (root-inducing) plasmid in Agrobacterium rhizogenes and Ti (tumor inducing) plasmid in Agrobacterium tumefaciens have provided the fundamental basis for the construction of plant vectors and transgenic plants. Recently, the determination of the first complete nucleotide sequence of the Ti plasmid (pTi SAKURA) has been successful. To understand the general structure of these oncogenic T-DNA transfer plasmids, the whole nucleotide sequence of a mikimopine type Ri plasmid, pRi1724, was analyzed. The plasmid is 217,594 bp in size, and has 173 open reading frames (ORFs) in total, which are asymmetrically distributed. Except for 27 ORFs, which are unknown, 173 ORFs were classified into 12 groups as follows: three for DNA replication, nine for plasmid modification, 22 for conjugation, 26 for virulence, 11 for T-DNA gene, 19 for mikimopine/mikimopine-lactam transport, ten for an unknown opine metabolism, seven for transcriptional regulator, five for sugar transport, five for glycerol metabolism, four for chemoreceptor and 32 for others. The elucidated chimeric structure of pRi1724 interestingly indicates that the evolution of Rhizobiaceae plasmids seems to have kept interactions among the plasmids; especially, the genes and elements for a conjugal transfer of pRi1724 had clearly closer kinship to those of a Sym (symbiotic) plasmid, pNGR234a in Rhizobium sp. than those of Ti plasmids. By using sequencing and Northern analysis, we examined the metabolic pathway and gene expression of mikimopine, which is probably an Ri-specific opine. PMID- 11273701 TI - Charging levels of four tRNA species in Escherichia coli Rel(+) and Rel(-) strains during amino acid starvation: a simple model for the effect of ppGpp on translational accuracy. AB - Escherichia coli strains mutated in the relA gene lack the ability to produce ppGpp during amino acid starvation. One consequence of this deficiency is a tenfold increase in misincorporation at starved codons compared to the wild-type. Previous work had shown that the charging levels of tRNAs were the same in Rel(+) and Rel(-) strains and reduced, at most, two- to fivefold in both strains during starvation. The present reinvestigation of the charging levels of tRNA(2)(Arg), tRNA(1)(Thr), tRNA(1)(Leu) and tRNA(His) during starvation of isogenic Rel(+) and Rel(-) strains showed that starvation reduced charging levels tenfold to 40-fold. This reduction corresponds much better with the decreased rate of protein synthesis during starvation than that reported earlier. The determination of the charging levels of tRNA(2)(Arg) and tRNA(1)(Thr) during starvation were accurate enough to demonstrate that charging levels were at least fivefold lower in the Rel(-) strain compared to the Rel(+) strain. Together with other data from the literature, these new data suggest a simple model in which mis-incorporation increases as the substrate availability decreases and that ppGpp has no direct effect on enhancing translational accuracy at the ribosome. PMID- 11273702 TI - An expressed sequence tag (EST) data mining strategy succeeding in the discovery of new G-protein coupled receptors. AB - We have developed a comprehensive expressed sequence tag database search method and used it for the identification of new members of the G-protein coupled receptor superfamily. Our approach proved to be especially useful for the detection of expressed sequence tag sequences that do not encode conserved parts of a protein, making it an ideal tool for the identification of members of divergent protein families or of protein parts without conserved domain structures in the expressed sequence tag database. At least 14 of the expressed sequence tags found with this strategy are promising candidates for new putative G-protein coupled receptors. Here, we describe the sequence and expression analysis of five new members of this receptor superfamily, namely GPR84, GPR86, GPR87, GPR90 and GPR91. We also studied the genomic structure and chromosomal localization of the respective genes applying in silico methods. A cluster of six closely related G-protein coupled receptors was found on the human chromosome 3q24-3q25. It consists of four orphan receptors (GPR86, GPR87, GPR91, and H963), the purinergic receptor P2Y1, and the uridine 5'-diphosphoglucose receptor KIAA0001. It seems likely that these receptors evolved from a common ancestor and therefore might have related ligands. In conclusion, we describe a data mining procedure that proved to be useful for the identification and first characterization of new genes and is well applicable for other gene families. PMID- 11273703 TI - The mitochondrial proteins Ssq1 and Jac1 are required for the assembly of iron sulfur clusters in mitochondria. AB - Mitochondria of the yeast Saccharomyces cerevisiae contain three different Hsp70 chaperones, Ssc1, Ecm10 and Ssq1. Ssc1 is an essential protein that mediates the import of nuclear-encoded proteins into the organelle and their subsequent folding. The nucleotide state of Ssc1 is thereby regulated by the nucleotide exchange factor Mge1. Here, we show that Mge1 interacts with Ssq1 in an ATP dependent manner, suggesting that Mge1 also regulates Ssq1 function. In contrast to Ssc1, Ssq1 does not associate with the Tim44 subunit of the protein translocating complex, indicating a different function of both chaperones. Mutants in Ssq1 were reported to have low levels of iron sulfur (FeS) cluster containing enzymes. Employing an assay that allowed us to monitor the conversion of the apoform of mitochondrial ferredoxin into its FeS-containing holoform, Ssq1 was demonstrated to be required for the FeS cluster assembly in mitochondria. The mitochondrial DnaJ homolog Jac1 is crucial for this process, whereas Mdj1 function is dispensable. Furthermore, the presence of frataxin is necessary for FeS cluster assembly into ferredoxin suggesting a role for frataxin at the level of the formation of holo-ferredoxin. PMID- 11273704 TI - Structural and thermodynamic studies on mutant RNA motifs that impair the specificity between a viral replicase and its promoter. AB - The 3'-end region of the genomic RNA of brome mosaic virus forms a tRNA-like structure that is critical for its replication. Previous studies have shown that in this region, a stem-loop structure, called SLC, is necessary and sufficient for the binding of the RNA replicase, and for RNA replication. Recently, we determined the high-resolution NMR structure of SLC, which demonstrated that a 5' AUA-3' triloop region is an important structural element for the enzymatic recognition. We proposed that the 5'-adenine of the triloop, which is rigidly fixed ("clamped") to the stem, is a key recognition element for the replicase. To elucidate the role of this "clamped base motif" for the enzymatic recognition, we have now investigated the solution conformations of several stem-loop molecules with mutant triloops, 5'-UUA-3', 5'-GUA-3', 5'-CUA-3' and 5'-UUU-3', that destroy the enzymatic recognition. For the GUA and UUA mutants, we have obtained high resolution solution structures using 2D NMR. All four mutants have very similar thermodynamic stabilities, and all have the same secondary structures, a triloop with a five base-paired stem helix. In addition, they have quite similar sugar puckering patterns in the triloop region. The NMR structures of the GUA and UUA show that the 5' nucleotide of the triloop (G6 in GUA or U6 in UUA) lacks the strong interactions that hold its base in a fixed position. In particular, the U6 of UUA is found in two different conformations. Neither of these two mutants has the clamped base motif that was observed in the wild-type. While UUA also shows global change in the overall triloop conformation, GUA shows a very similar triloop conformation to the wild-type except for the lack of this motif. The absence of the clamped base motif is the only common structural difference between these two mutants and the wild-type. These results clearly indicate that the loss of function of the UUA and GUA mutants comes mainly from the destruction of a small key recognition motif rather than from global changes in their triloop conformations. Based on this study, we conclude that the key structural motif in the triloop recognized by the replicase is a solution-exposed, 5'-adenine base in the triloop that is clamped to the stem helix, which is called a clamped adenine motif. PMID- 11273705 TI - SuperStar: improved knowledge-based interaction fields for protein binding sites. AB - SuperStar is an empirical method for identifying interaction sites in proteins, based entirely on experimental information about non-bonded interactions occurring in small-molecule crystal structures, taken from the IsoStar database. We describe recent modifications and additions to SuperStar, validating the results on a test set of 122 X-ray structures of protein-ligand complexes. In this validation, propensity maps are generated for all the binding sites of these proteins, using four different probes: a charged NH(+)(3) nitrogen atom, a carbonyl oxygen atom, a hydroxyl oxygen atom and a methyl carbon atom. Next, the maps are compared with the experimentally observed positions of ligand atoms of these types. A peak-searching algorithm is introduced that highlights potential interaction hot spots. For the three hydrogen-bonding probes - NH(+)(3) nitrogen atom, carbonyl oxygen atom and hydroxyl oxygen atom - the average distance from the ligand atom to the nearest SuperStar peak is 1.0-1.2 A (0.8-1.0 A for solvent inaccessible ligand atoms). For the methyl carbon atom probe, this distance is about 1.5 A, probably because interactions to methyl groups are much less directional. The most important addition to SuperStar is the enabling of propensity maps around metal centres - Ca(2+), Mg(2+) and Zn(2+) - in protein binding sites. The results are validated on a test set of 24 protein-ligand complexes that have a metal ion in their binding site. Coordination geometries are derived automatically, using only the protein atoms that coordinate to the metal ion. The correct coordination geometry is derived in approximately 75 % of the cases. If the derived geometry is assumed during the SuperStar calculation, the average distance from a ligand atom coordinating to the metal ion to the nearest peak in the propensity map for an oxygen probe is 0.87(7) A. If the correct coordination geometry is imposed, this distance reduces to 0.59(7)A. This indicates that the SuperStar predictions around metal-binding sites are at least as good as those around other protein groups. Using clustering techniques, a non redundant set of probes is selected from the set of probes available in the IsoStar database. The performance in SuperStar of all these probes is tested on the test set of protein-ligand complexes. With the exception of the "ether oxygen" probe and the "any NH(+)" probe, all new probes perform as well as the four probes introduced first. PMID- 11273706 TI - Solution structure, domain features, and structural implications of mutants of the chromo domain from the fission yeast histone methyltransferase Clr4. AB - The encapsulation of otherwise transcribable loci within transcriptionally inactive heterochromatin is rapidly gaining recognition as an important mechanism of epigenetic gene regulation. In the fission yeast Schizosaccharomyces pombe, heterochromatinization of the mat2/mat3 loci silences the mating-type information encoded within these loci. Here, we present the solution structure of the chromo domain from the cryptic loci regulator protein Clr4. Clr4 is known to regulate silencing and switching at the mating-type loci and to affect chromatin structure at centromeres. Clr4 and its human and Drosophila homologs have been identified as histone H3-specific methyltransferases, further implicating this family of proteins in chromatin remodeling. Our structure highlights a conserved surface that may be involved in chromo domain-ligand interactions. We have also analyzed two chromo domain mutants (W31G and W41G) that previously were shown to affect silencing and switching in full-length Clr4. Both mutants are significantly destabilized relative to wild-type. PMID- 11273707 TI - Phosphorylation-induced structural changes in the amyloid precursor protein cytoplasmic tail detected by NMR. AB - The cytoplasmic tail of the amyloid precursor protein (APPc) interacts with several cellular factors implicated in intracellular signaling or proteolytic production of amyloid beta peptide found in senile plaques of Alzheimer's disease patients. APPc contains two threonine residues (654 and 668 relative to APP695, or 6 and 20 relative to APPc) and a serine residue (655 or 7, respectively) that are known to be phosphorylated in vivo and may play regulatory roles in these events. We show by solution NMR spectroscopy of a 49 residue cytoplasmic tail peptide (APP-C) that in all three cases, phosphorylation induces changes in backbone dihedral angles that can be attributed to formation of local hydrogen bonds between the phosphate group and nearby amide protons. Phosphorylation of S7 also induces chemical shift changes in the hydrophobic cluster (residues I8-V13), indicating additional medium-range effects. The most pronounced changes occur upon phosphorylation of T20, a neuron-specific phosphorylation site, where the N terminal helix capping box previously characterized for this region is altered. Characterization of torsion angles and transient hydrogen bonds indicates that prolyl isomerization of the pThr-Pro peptide bond results from both destabilization of the N-terminal helix capping box and stabilization of the cis isomer by transient hydrogen bonds. The significant population of the cis isomer (9 %) present after phosphorylation of T20 suggests a potential role of selective recognition of cis versus trans isomers in response to phosphorylation of APP. Together, these structural changes indicate that phosphorylation may act as a conformational switch in the cytoplasmic tail of APP to alter specificity and affinity of binding to cytosolic partners, particularly in response to the abnormal phosphorylation events associated with Alzheimer's disease. PMID- 11273708 TI - Comparison of the structural and dynamical properties of holo and apo bovine alpha-lactalbumin by NMR spectroscopy. AB - In the presence of 0.5 M NaCl at pH 7.1, the Ca(2+)-free apo form of recombinant bovine alpha-lactalbumin (BLA) is sufficiently stabilised in its native state to give well-resolved NMR spectra at 20 degrees C. The (1)H and (15)N NMR resonances of native apo-BLA have been assigned, and the chemical-shifts compared with those of the native holo protein. Large changes observed between the two forms of BLA are mainly limited to the Ca(2+)-binding region of the protein. These data suggest that Na(+) stabilises the native apo state through the screening of repulsive negative charges, at the Ca(2+)-binding site or elsewhere, rather than by a specific interaction at the vacant Ca(2+)-binding site. The hydrogen exchange protection of residues in the Ca(2+)-binding loop and the C-helix is reduced in the apo form compared to that in the holo form. This indicates that the dynamic behaviour of this region of the protein is substantially increased in the absence of the bound Ca(2+). Real-time NMR experiments show that the rearrangements of the structure associated with the conversion of the holo to apo form of the protein do not involve the detectable population of partially unfolded intermediates. Rather, the conversion appears to involve local reorganisations of the structure in the vicinity of the Ca(2+)-binding site that are coupled to the intrinsic fluctuations in the protein structure. PMID- 11273709 TI - Distinct cysteine sulfhydryl environments detected by analysis of Raman S-hh markers of Cys-->Ser mutant proteins. AB - Very little is known about the character or functional relevance of hydrogen bonded cysteine sulfhydryl (S-H) groups in proteins. The Raman S-H band is a unique and sensitive probe of the local S-H environment. Here, we report the use of Raman spectroscopy combined with site-specific mutagenesis to document the existence of five distinguishable hydrogen-bonded states of buried cysteine sulfhydryl groups in a native protein. The 666 residue subunit of the Salmonella typhimurium bacteriophage P22 tailspike contains eight cysteine residues distributed through the elongated structure. The tailspike cysteine residues display an unusual Raman S-H band complex (2500-2600 cm(-1) interval) indicative of diverse S-H hydrogen-bonding interactions in the native trimeric structure. To resolve specific Cys contributions to the complex Raman band we characterized a set of tailspike proteins with each cysteine replaced by a serine. The mutant proteins, once folded, were structurally and functionally indistinguishable from wild-type tailspikes, except for their Raman S-H signatures. Comparison of the Raman spectra of the mutant and wild-type proteins reveals the following hydrogen bond classes for cysteine sulfhydryl groups. (i) Cys613 forms the strongest S H...X bond of the tailspike, stronger than any heretofore observed for a protein. (ii) Cys267, Cys287 and Cys458 form robust S-H...X bonds. (iii) Moderate S-H...X bonding occurs for Cys169 and Cys635. (iv) Cys290 and Cys496 form weak hydrogen bonds. (v) It is remarkable that Cys287 contributes two Raman S-H markers, indicating the population of two distinct hydrogen-bonding states. The sum of the S-H Raman signatures of all eight mutants accurately reproduces the composite Raman band of the wild-type tailspike. The diverse cysteine states may be an outcome of the folding and assembly pathway of the tailspike, which though lacking disulfide bonds in the native state, utilizes transient disulfide bonds in the maturation pathway. This Raman study represents the first detailed assessment of local S-H hydrogen bonding in a native protein and provides information not obtainable directly by other structural probes. The method employed here should be applicable to a wide range of cysteine-containing proteins. PMID- 11273711 TI - Mapping protein family interactions: intramolecular and intermolecular protein family interaction repertoires in the PDB and yeast. AB - In the postgenomic era, one of the most interesting and important challenges is to understand protein interactions on a large scale. The physical interactions between protein domains are fundamental to the workings of a cell: in multi domain polypeptide chains, in multi-subunit proteins and in transient complexes between proteins that also exist independently. To study the large-scale patterns and evolution of interactions between protein domains, we view interactions between protein domains in terms of the interactions between structural families of evolutionarily related domains. This allows us to classify 8151 interactions between individual domains in the Protein Data Bank and the yeast Saccharomyces cerevisiae in terms of 664 types of interactions, between protein families. At least 51 interactions do not occur in the Protein Data Bank and can only be derived from the yeast data. The map of interactions between protein families has the form of a scale-free network, meaning that most protein families only interact with one or two other families, while a few families are extremely versatile in their interactions and are connected to many families. We observe that almost half of all known families engage in interactions with domains from their own family. We also see that the repertoires of interactions of domains within and between polypeptide chains overlap mostly for two specific types of protein families: enzymes and same-family interactions. This suggests that different types of protein interaction repertoires exist for structural, functional and regulatory reasons. PMID- 11273710 TI - Dramatic stabilization of an SH3 domain by a single substitution: roles of the folded and unfolded states. AB - The N-terminal SH3 domain of the Drosophila drk protein (drkN SH3) exists in equilibrium between folded and unfolded states under non-denaturing buffer conditions. In order to examine the origins of this instability, we have made mutations in the domain and characterized the thermodynamics and kinetics of folding. Results of substitutions of negatively charged residues to neutral amino acid residues suggest that the large electrostatic potential of the domain does not play a dominant role in the instability of the domain. Sequence alignment of a large number of SH3 domains reveals that the drkN SH3 domain has a threonine (T22) at a position corresponding to an otherwise highly conserved glycine residue in the diverging beta-turn connecting the beta3 and beta4 strands. Mutation of T22 to glycine results in significant stabilization of the drkN SH3 domain by 2.5 kcal/mole. To further characterize the basis for the stabilization of the T22 mutant relative to wild-type, we made additional mutant proteins with substitutions of residue T22. A strong correlation is seen between protein stability or folding rate and propensity for native beta-turn structure at this position. Correlation of folding rates with AGADIR predictions of non-native helical structure in the diverging turn region, along with our previous NMR evidence for non-native structure in this region of the unfolded state of the drkN SH3 domain, suggests that the free energy of the unfolded state also plays a role in stability. This result highlights the importance of both folded and unfolded states for understanding protein stability. PMID- 11273712 TI - Consistency analysis of similarity between multiple alignments: prediction of protein function and fold structure from analysis of local sequence motifs. AB - A new method to analyze the similarity between multiply aligned protein motifs (blocks) was developed. It identifies sets of consistently aligned blocks. These are found to be protein regions of similar function and structure that appear in different contexts. For example, the Rossmann fold ligand-binding region is found similar to TIM barrel and methylase regions, various protein families are predicted to have a TIM-barrel fold and the structural relation between the ClpP protease and crotonase folds is identified from their sequence. Besides identifying local structure features, sequence similarity across short sequence regions (less than 20 amino acid regions) also predicts structure similarity of whole domains (folds) a few hundred amino acid residues long. Most of these relations could not be identified by other advanced sequence-to-sequence or sequence-to-multiple alignments comparisons. We describe the method (termed CYRCA), present examples of our findings, and discuss their implications. PMID- 11273714 TI - Introduction: what do we know today about form and function of cardiac ion channels? PMID- 11273713 TI - Target recognition by EcoKI: the recognition domain is robust and restriction deficiency commonly results from the proteolytic control of enzyme activity. AB - We report a genetic and biochemical analysis of a target recognition domain (TRD) of EcoKI, a type I restriction and modification enzyme. The TRDs of type I R-M systems are within the specificity subunit (HsdS) and HsdS confers sequence specificity to a complex endowed with both restriction and modification activities. Random mutagenesis has revealed that most substitutions within the amino TRD of EcoKI, a region comprising 157 amino acid residues, have no detectable effect on the phenotype of the bacterium, even when the substitutions are non- conservative. The structure of the TRD appears to be robust. All but one of the six substitutions that confer a restriction-deficient, modification deficient (r(-)m(-)) phenotype were found to be in the interval between residues 80 and 110, a region predicted by sequence comparisons to form part of the protein-DNA interface. Additional site-directed mutations affecting this interval commonly impair both restriction and modification. However, we show that an r(-) phenotype cannot be taken as evidence that the EcoKI complex lacks endonuclease activity; in response to even a slightly impaired modification efficiency, the endonuclease activity of EcoKI is destroyed by a process dependent upon the ClpXP protease. Enzymes from mutants with an r(-)m(-) phenotype commonly retain some sequence-specific activity; methylase activity can be detected on hemimethylated DNA substrates and residual endonuclease activity is implied whenever the viability of the r(-)m(-) bacterium is dependent on ClpXP. Conversely, the viability of ClpX(-) r(-)m(-) bacteria can be used as evidence for little, or no, endonuclease activity. Of 14 mutants with an r(-)m(-) phenotype, only six are viable in the absence of ClpXP. The significance of four of the six residues (G91, G105, F107 and G141) is enhanced by the finding that even conservative substitutions for these residues impair modification, thereby conferring an r( )m(-) phenotype. PMID- 11273715 TI - The cardiac sodium channel: gating function and molecular pharmacology. AB - Cardiac sodium (Na) channels are dynamic molecules that undergo rapid structural changes in response to the changing electrical field in the myocardium. Inherited mutations in SCN5A, the gene encoding the cardiac Na channel, provoke life threatening cardiac arrhythmias, often by modifying these voltage-dependent conformational changes. These disorders (i.e. the long QT syndrome and Brugada syndrome) may serve as valuable models for understanding the mechanistic linkages between Na channel dysfunction and cardiac arrhythmias in more common, acquired conditions such as cardiac ischemia. In addition, the balance between therapeutic and adverse effects from Na channel blockade by antiarrhythmic compounds may be shifted by subtle alterations in Na channel function. This review examines recent studies that tie key loci in the Na channel primary sequence to its dynamic function, while examining the emerging themes linking Na channel structure, function, and pharmacology to inherited and acquired disorders of cardiac excitability. PMID- 11273716 TI - Ryanodine receptors/calcium release channels in heart failure and sudden cardiac death. AB - Calcium (Ca2+) ions are second messengers in signaling pathways in all types of cells. They regulate muscle contraction, electrical signals which determine the cardiac rhythm and cell growth pathways in the heart. In the past decade cDNA cloning has provided clues as to the molecular structure of the intracellular Ca2+ release channels (ryanodine receptors, RyR, and inositol 1,4,5-trisphosphate receptors, IP3R) on the sarcoplasmic and endoplasmic reticulum (SR/ER) and an understanding of how these molecules regulate Ca2+ homeostasis in the heart is beginning to emerge. The intracellular Ca2+ release channels form a distinct class of ion channels distinguished by their structure, size, and function. Both RyRs and IP3Rs have gigantic cytoplasmic domains that serve as scaffolds for modulatory proteins that regulate the channel pore located in the carboxy terminal 10% of the channel sequence. The channels are tetramers comprised of four RyR or IP3R subunits. RyR2 is required for excitation-contraction (EC) coupling in the heart. Using co-sedimentation and co-immunoprecipitation we have defined a macromolecular complex comprised of RyR2, FKBP12.6, PKA, the protein phosphatases PP1 and PP2A, and an anchoring protein mAKAP. We have shown that protein kinase A (PKA) phosphorylation of RyR2 dissociates FKBP12.6 and regulates the channel open probability (P(o)). In failing human hearts RyR2 is PKA hyperphosphorylated resulting in defective channel function due to increased sensitivity to Ca2+-induced activation. PMID- 11273717 TI - Inward rectifiers in the heart: an update on I(K1). AB - The cardiac inward rectifier potassium current (I(K1)), present in all ventricular and atrial myocytes, has been suggested to play a major role in repolarization of the action potential and stabilization of the resting potential. The molecular basis is now ascribed to members of the Kir2 sub-family of inward rectifier K channel genes, and the availability of recombinant expression systems has led to elucidation of the mechanism of inward rectification, as well as additional regulatory mechanisms involving intracellular pH and phosphorylation. In vivo manipulation of the genes encoding I(K1)and regulatory proteins now promise to provide new insights to the role of this conductance in the heart. This review details recent advances and considers the prospects for further elucidation of the role of this conductance in cardiac electrical activity. PMID- 11273718 TI - Ca2+-dependent regulation of cardiac L-type Ca2+ channels: is a unifying mechanism at hand? AB - Ca2+ entry (I(Ca)) through cardiac L-type Ca2+ channels (LTCC) drives critical cellular processes ranging from contraction to gene expression, and, when disordered, is implicated in arrhythmias and hypertrophy. LTCC activation occurs by cell membrane depolarization, but LTCCs are also regulated by auxiliary proteins, phosphorylation, and intracellular CA2+([Ca2+]i). LTCC regulation by [Ca2+]i is especially intriguing because increased [Ca2+]i signals dual and conflicting commands for I(Ca)inactivation and facilitation. A recent explosion of work has shed new light on the mechanisms and molecular identity of domains necessary for [Ca2+]i-dependent regulation of LTCC. PMID- 11273719 TI - Mitochondrial K(ATP)channel opening during index ischemia and following myocardial reperfusion in ischemic rat hearts. PMID- 11273721 TI - p38 MAPK activity is not increased early during sustained coronary artery occlusion in preconditioned versus control rabbit heart. AB - Our aim was to test the hypothesis that cardioprotection achieved with ischemic preconditioning (PC) involves increased activity of p38 mitogen-activated protein kinase (MAPK) early during sustained coronary artery occlusion. Using the isolated buffer-perfused rabbit heart model of regional ischemia, we quantified p38 MAPK activity (pmol/min/mg protein: by biochemical assay) at 5 and 10 min into coronary occlusion in hearts that first received PC ischemia or no intervention (controls), and in non-ischemic shams. Control hearts exhibited significant increases in p38 MAPK activity, averaging 883+/-142 and 1135+/-179 at 5 and 10 min of occlusion, v 144+/-49 in shams (P<0.05 and P<0.01). p38 MAPK activity was not, however, augmented with PC; rather, at 5 min into occlusion, activity was attenuated, averaging 432+/-72 (P=N.S. v sham). This early, modest reduction in p38 MAPK activity may be physiologically relevant: in additional hearts subjected to 30 min of sustained coronary occlusion and 2 h of reperfusion, infarct size (by tetrazolium staining: expressed as a % of the risk region) was 54+/-5% in hearts treated with SB 203580 (confirmed in our study to inhibit p38 MAPK activity at 5 min into occlusion) v 70+/-5% in vehicle controls (P<0.05). Thus, cardioprotection achieved with ischemic preconditioning in rabbit heart does not involve augmentation of p38 MAPK activity early during sustained coronary occlusion. PMID- 11273720 TI - The molecular genetic basis for hypertrophic cardiomyopathy. AB - Hypertrophic cardiomyopathy (HCM), a relatively common disease, is diagnosed clinically by unexplained cardiac hypertrophy and pathologically by myocyte hypertrophy, disarray, and interstitial fibrosis. HCM is the most common cause of sudden cardiac death (SCD) in the young and a major cause of morbidity and mortality in elderly. Hypertrophy and fibrosis are the major determinants of morbidity and SCD. More than 100 mutations in nine genes, all encoding sarcomeric proteins have been identified in patients with HCM, which had led to the notion that HCM is a disease of contractile sarcomeric proteins. The beta -myosin heavy chain (MyHC), cardiac troponin T (cTnT) and myosin binding protein-C (MyBP-C) are the most common genes accounting for approximately 2/3 of all HCM cases. Genotype phenotype correlation studies suggest that mutations in the beta -MyHC gene are associated with more extensive hypertrophy and a higher risk of SCD as compared to mutations in genes coding for other sarcomeric proteins, such as MyBP-C and cTnT. The prognostic significance of mutations is related to their hypertrophic expressivity and penetrance, with the exception of those in the cTnT, which are associated with mild hypertrophic response and a high incidence of SCD. However, there is a significant variability and factors, such as modifier genes and probably the environmental factors affect the phenotypic expression of HCM. The molecular pathogenesis of HCM is not completely understood. In vitro and in vivo studies suggest that mutations impart a diverse array of functional defects including reduced ATPase activity of myosin, acto-myosin interaction, cross bridging kinetics, myocyte contractility, and altered Ca2+ sensitivity. Hypertrophy and other clinical and pathological phenotypes are considered compensatory phenotypes secondary to functional defects. In summary, the molecular genetic basis of HCM has been identified, which affords the opportunity to delineate its pathogenesis. Understanding the pathogenesis of HCM could provide for genetic based diagnosis, risk stratification, treatment and prevention of cardiac phenotypes. PMID- 11273722 TI - Angiotensin II receptor antagonist blocks the expression of connexin43 induced by cyclical mechanical stretch in cultured neonatal rat cardiac myocytes. AB - Mechanical forces have profound effects on cardiomyocytes. To test whether angiotensin II is a potential mediator of stretch-induced effects on gap junctions, we used the angiotensin II (AT1) receptor antagonist, losartan, to investigate the cyclical stretch-induced expression of connexin43 (Cx43), the major cardiac muscle gap junction channel protein. Cultured neonatal rat cardiomyocytes grown on a flexible membrane base were stretched by vacuum to 20% of maximum elongation, at 60 cycles/min. The levels of Cx43 protein began to increase as early as 2 h after stretch was applied, reached a maximum of six-fold over the control by 24 h and remained at this level another 24 h (i.e. up to 48 h after stretch was applied). These increases of Cx43 protein at 24 h were largely (73%) and completely (100%) attenuated (P<0.001) by the addition (30 min before stretch) of 10 n M and 100 n M losartan, respectively. Similarly, the Cx43 mRNA levels in stretched cardiomyocytes rose 89% (P<0.01) above control (non-stretched cells) mRNA levels. This increase also was blocked by losartan. Cyclical stretch increased (and losartan decreased) the immunohistochemical labeling of Cx43 and significantly increased release of angiotensin II into the culture media from 7.5+/-0.6 ng/ml to 23.8+/-1.0 ng/ml (P<0.01) after a 1 h stretch. These findings indicate that cyclical mechanical stretch augments angiotensin II production and Cx43 gene expression in cultured cardiomyocytes, partially through mediation of the AT1 receptors, and suggests interaction between the cardiomyocyte local rennin-angiotensin system and Cx43 in response to stretch. PMID- 11273723 TI - Dual influence of disease and increased load on diaphragm muscle in heart failure. AB - We have recently shown that mitochondrial function and energy metabolism are altered in the myocardium as well as in slow and fast locomotor muscles of rats subjected to prolonged congestive heart failure (CHF) suggesting a generalized metabolic myopathy in heart failure. Here, we investigate whether the diaphragm of CHF animals, which experiences both increased work and the general systemic influence of heart failure, will also be susceptible to altered energy metabolism. Biopsies were obtained from the costal diaphragm of failing rats 8 months after aortic banding. A marked increase in type I and type IIa myosin heavy chains at the expense of types IIx and IIb, suggests an adaptation towards a slower phenotype. Glycolytic enzymes decreased in CHF diaphragm with an increase in the H:M lactate dehydrogenase isoenzyme ratio. These results suggest a reorientation of the diaphragm muscle towards a slow, fatigue-resistant phenotype. However, maximal oxidative capacity assessed in saponin-permeabilized fibers in the presence of ADP was considerably reduced in CHF diaphragm (7.7+/ 0.4 v 11.8+/-0.7 micromol O2/min/g dry weight in sham P<0.001), suggesting an alteration in oxidative phosphorylation. Furthermore, ADP sensitivity of CHF mitochondria was significantly increased (apparent Km for ADP 308+/-21 v 945+/ 106 microM in sham P<0.001), whereas sensitivity to ADP in the presence of creatine was comparable (Km 79+/-12 v 90+/-11 microM in sham). In heart failure, therefore, the diaphragm muscle seems to adapt towards a more slow and economical contraction as a result of increased workload, but this adaptation is limited by the disease-induced altered mitochondrial function. PMID- 11273724 TI - Rest-dependence of twitch amplitude and sarcoplasmic reticulum calcium content in the developing rat myocardium. AB - Post-rest contractile response was studied in isolated ventricular muscle from rats aged 1 to 90 days. Amplitude of rapid cooling contractures (RCC) was taken as an index of the sarcoplasmic reticulum (SR) Ca2+ content. We observed that: (a) developed tension (per cross-section area) increased with age; (b) time to peak twitch force and relaxation half-time decreased from 87+/-6 to 56+/-2 ms and from 68+/-6 to 36+/-1 ms, respectively, from the neonatal period to adulthood; (c) post-rest twitch potentiation was observed at all ages, with greater relative potentiation in younger preparations, although relative potentiation of [Ca2+]i transient amplitude was similar in young and adult isolated ventricular myocytes; (d) rest did not significantly affect the amplitude of RCC in muscle or caffeine evoked [Ca2+]i transients in myocytes at any studied age; (e) favoring Ca2+ efflux via Na+-Ca2+ exchange (NCX) during rest reversed twitch potentiation and caused a similar decrease in RCC amplitude ( approximately 40%) at all ages; (f) stimulation of Ca2+ influx via NCX during rest increased RCC amplitude ( approximately 40%) only in immature preparations. However, when this procedure was repeated after partial SR Ca2+ depletion, increase in RCC amplitude was not significantly age-dependent. We conclude that post-rest twitch potentiation is already present early after birth and does not require rest-dependent changes in SR Ca2+ content at any studied age. Our results suggest that NCX is close to equilibrium during rest in both adult and developing rat myocardium, and does not seem to mediate diastolic net Ca2+ fluxes which may affect the SR Ca2+ content. PMID- 11273725 TI - Mutations that alter the surface charge of alpha-tropomyosin are associated with dilated cardiomyopathy. AB - Proteins in cardiac myocytes assemble into contractile units known as sarcomeres. Contractile force is generated by interaction between sarcomeric thick and thin filaments. Thin filaments also transmit force within and between myocytes. Mutations in genes encoding the thin filament proteins actin and tropomyosin cause hypertrophic cardiomyopathy. Mutations affecting functionally distinct domains of actin also cause dilated cardiomyopathy (DCM). We used a non positional candidate gene approach to test further the hypothesis that dysfunction of sarcomeric thin filaments, due to different mutations in the same gene, can lead to either hypertrophic or dilated cardiomyopathy. Mutational analyses of alpha-tropomyosin 1 were performed in patients with idiopathic DCM. We identified two mutations that alter highly conserved residues and that, unlike hypertrophic cardiomyopathy-associated mutations, cause localized charge reversal on the surface of tropomyosin. Therefore, substitution of different amino acid residues in the same thin filament proteins is associated with the distinct phenotypes of cardiac hypertrophy or congestive heart failure. PMID- 11273727 TI - Regional localization and abundance of calcitonin gene-related peptide receptors in guinea pig heart. AB - Calcitonin gene-related peptide (CGRP) is a neurotransmitter that is released within the heart during myocardial ischemia. The present study was done to determine the regional localization and abundance of CGRP receptors in the guinea pig heart. CGRP binding sites in 20 microm frozen sections of heart were labeled using [125I]CGRP. Non-specific binding was determined in the presence of 1 microm unlabeled CGRP or CGRP(8-37). Significant amounts of specific CGRP binding were identified in atrial and ventricular myocardium, all portions of the conducting system, coronary arteries, the aorta and pulmonary trunk and intracardiac ganglia. Specific binding of CGRP to the left atrium was two-fold higher than binding to the right atrium (0.667+/-0.052 v 0.340+/-0.029 fmol/mg tissue, n=5, CGRP(8-37)group). In contrast to the atria, a lower and uniform density of CGRP receptors occurred in contractile tissue of the ventricular myocardium (e.g. 0.239+/-0.013 fmol/mg left ventricle, n=5). The highest concentration of CGRP receptors in guinea pig cardiac tissue occurred at the bundle of His and the bundle branches (0.752+/-0.087 and 0.713+/-0.138 fmol/mg tissue, respectively, n=5). CGRP receptors were localized to coronary vessels throughout the heart and to the ascending aorta and pulmonary trunk. Lastly, intracardiac ganglia exhibited moderate levels of specific [125I]CGRP binding (0.475+/-0.043 fmol/mg, n=5). These findings support the concept that CGRP can have direct effects on atrial and ventricular function as well as coronary flow. The high density of CGRP receptors in the distal conducting system and the presence of CGRP receptors in intracardiac ganglia further suggest that CGRP could have important effects on cardiac conduction velocity and parasympathetic regulation of the heart. PMID- 11273726 TI - Stage-specific differential activation of mitogen-activated protein kinases in hypertrophied and failing rat hearts. AB - Mitogen-activated protein kinases (MAPKs) are involved in the early development of cardiac hypertrophy, but their roles in chronic left ventricular hypertrophy (LVH) are unclear. We studied the angiotensin (Ang) II-induced cardiac MAPK activation of the hypertensive Dahl salt-sensitive (DS) rats in the subacute developing LVH stage, the chronic compensated LVH stage, and the congestive heart failure (CHF) stage. In the isolated, coronary-perfused heart preparation, Ang II infusion (1x10(-6)mol/l) activated extracellular signal-regulated kinase (ERK), c Jun N-terminal kinase (JNK) and p38-MAPK in the LV myocardium. No substantial differences were observed in the Ang II-induced ERK activation between the normotensive control DS rats and the hypertensive DS rats in either stage. In contrast, the Ang II-induced activation of JNK and p38-MAPK was augmented in the subacute LVH stage of the hypertensive DS rats, but then progressively attenuated in the chronic LVH and CHF stages. Chronic treatment with an angiotensin converting enzyme inhibitor, temocapril (20 mg/kg/day), ameliorated the responsiveness of the JNK/p38-MAPK activation, suggesting that the decreased JNK/p38-MAPK activation is a consequence of negative feedback regulation for the activated cardiac renin-angiotensin system in chronic LVH and CHF. Thus, the Ang II-induced activation of multiple cardiac MAPK pathways are differentially regulated, depending on the stages of chronic hypertrophic process. The JNK and p38-MAPK activation may be involved in the early development of adaptive LVH. However, the responsiveness of the cardiac JNK/p38-MAPK pathways progressively decreased in chronic LVH and CHF under the chronic activation of tissue renin angiotensin system. PMID- 11273728 TI - Cumulative inactivation of the outward potassium current: a likely mechanism underlying electrical memory in human atrial myocytes. AB - The influence of the mode of cell stimulation on the outward K+ current (I(o)) was studied in whole-cell patch-clamped human atrial myocytes. Acceleration of the rate of membrane depolarization at 1 Hz or during prolonged 5-s test pulses at 0.1 Hz increased the rate and extent of I(o) inactivation, resulting in enhanced inactivating (4.9+/-0.6 v 6.3+/-0.7 pA/pF) and suppressed maintained (5.9+/-1.2 v 3.2+/-0.3 pA/pF) current components. These alterations were associated with a leftward shift of the voltage-dependency of I(o), and persisted on returning to a control depolarization protocol (750-ms test pulses delivered at 0.1 Hz). The effects of increasing external K+ concentrations (40 m m) on the kinetics of I(o) were more pronounced following both rapid and prolonged depolarization (changes in I(t)/I(o)caused by 40 m m K+: 8.9+/-3.5% v 15.5+/-3.1% before and after prolonged depolarization; and 9.2+/-1.2% v 15.4+/-1.7% before and after rapid depolarization). The phosphatase inhibitor, okadaic acid, enhanced the effect of rapid and prolonged depolarization on I(o)whereas the inhibition of Ca2+/calmodulin-dependent protein kinase II (CaMK-II) with KN-62 or KN-93, or by intracellular application of the autocamtide-2-related inhibitory peptide, suppressed it. In conclusion, rapid and prolonged membrane depolarization both cause a cumulative increase in the rate and extent of I(o)inactivation. This process involves slow potassium channel inactivation mechanisms, is regulated by CaMK-II, and may contribute to the electrical memory of the atrial myocardium. PMID- 11273729 TI - Activation of p38 MAPK induced by a multi-cycle ischaemic preconditioning protocol is associated with attenuated p38 MAPK activity during sustained ischaemia and reperfusion. AB - The role of p38 mitogen-activated protein kinase (MAPK) in ischaemic preconditioning remains controversial. Since most previous studies focussed on events only during sustained ischaemia, the aim of this study was to establish the activation pattern of p38 MAPK during a multicycle preconditioning protocol, sustained ischaemia as well as reperfusion and to correlate these events with functional recovery of the isolated perfused rat heart. Isolated perfused rat hearts were preconditioned by 3x5 min global ischaemia followed by 25 min global ischaemia and 30 min reperfusion. Non-preconditioned hearts were subjected to 25 min global ischaemia and 30 min reperfusion. Hearts were freeze-clamped and p38 MAPK activation in tissue lysates was assessed by standard Western blotting techniques, using a dual phospho-p38 MAPK antibody as well as a non-radioactive IP-kinase assay. The results showed that transient dual phosphorylation and activation of p38 MAPK occurs during a 3x5 min preconditioning protocol: the activation was maximal during the first episode, becoming progressively lower during the second and third episodes. p38 MAPK activation was significantly less during both sustained ischaemia and reperfusion in preconditioned hearts, when compared with non-preconditioned hearts. Attenuation of p38 MAPK activity during sustained ischaemia and reperfusion was associated with improved functional recovery. The effect of inhibition of p38 MAPK activation on cardioprotection was further evaluated in adult, isolated cardiomyocytes. Administration of SB 203580 (1-10 microM) before and during the preconditioning protocol, had no effect on cell morphology and viability after 2 h hypoxia, compared to untreated preconditioned cardiomyocytes. When administered to non-preconditioned cells before the onset of 2 h hypoxia, it caused a significant improvement in both morphology and viability. In summary, the results suggest that attenuation of the kinase activity during sustained ischaemia and reperfusion may be an essential element of the preconditioning process. PMID- 11273731 TI - Protein kinase C-alpha and -epsilon modulate connexin-43 phosphorylation in human heart. AB - We have previously demonstrated that protein kinase C (PKC)- alpha expression is significantly elevated in failing human left ventricle, with immunostaining showing increased PKC- alpha localization at the intercalated disks of cardiomyocytes. In the present study we sought to determine, in the failing heart, if PKC- alpha interacted with connexin-43 (Cx-43) both spatially and functionally, and to compare the association of PKC- alpha/Cx-43 with that of PKC epsilon, a PKC isozyme that does not significantly increase in failing hearts. The possibility of a PKC- alpha or PKC- epsilon/Cx-43 association in non-failing hearts was also investigated. Co-immunoprecipitation of PKC- alpha or PKC- epsilon and Cx-43 in non-failing and failing left ventricle was achieved using antibodies to PKC- alpha or Cx-43. Confocal microscopy confirmed that PKC- alpha distribution within the cardiomyocyte included co-localization with connexin-43 in both failing and non-failing myocardium. In a similar manner, confocal imaging of PKC- epsilon showed cardiomyocyte distribution in both cytosol and membrane, and colocalization of PKC- epsilon with Cx-43. Recombinant PKC- alpha or - epsilon increased PKC activity significantly above endogenous levels in the co immunoprecipitated Cx-43 complexes (P<0.05). However, phosphorylation of purified human Cx-43 (isolated from failing human left ventricle) by recombinant PKC- alpha or PKC- epsilon resulted in only PKC- epsilon mediated Cx-43 phosphorylation. Thus, in the human heart PKC- alpha, PKC- epsilon, and Cx-43 appear to form a closely associated complex. Whereas only PKC- epsilon directly phosphorylates Cx-43, both PKC isoforms result in increased phosphorylation within the Cx-43 co-immunoprecipitated complex. PMID- 11273730 TI - MEK1/2-ERK1/2 mediates alpha1-adrenergic receptor-stimulated hypertrophy in adult rat ventricular myocytes. AB - We examined the relative roles of the mitogen-activated protein kinases (MAPK) in mediating the alpha1-adrenergic receptor (alpha1-AR) stimulated hypertrophic phenotype in adult rat ventricular myocytes (ARVM). Norepinephrine (NE; 1 microM) in the presence of the beta -AR antagonist propranolol (Pro; 2 microM) caused activation of Ras (>six-fold), MAPK/ERK kinase 1 and 2 (MEK1/2, >10-fold) and extracellular signal-regulated kinases 1 and 2 (ERK1/2, approximately 30-fold) within 5 min, as determined by kinase activity assays and Western blots using phospho-specific antibodies. Conversely, p38 and c-Jun amino-terminal kinases (JNK) were not activated by NE/Pro. Activated MEK1/2 signals remained detectable at 2 h, and activated ERK1/2 remained detectable at 48 h. The alpha1-AR selective inhibitor prazosin (100 nM) completely inhibited the NE/Pro-stimulated activation of Ras, MEK1/2 and ERK1/2. The MEK inhibitor PD98059 caused a concentration dependent inhibition of NE/Pro-stimulated protein synthesis (as assessed by [3H]leucine incorporation and cellular protein accumulation) and ERK1/2 activation, with approximately 50% inhibition at a concentration between 10 and 50 microM, which is consistent with the known IC50 values of PD98059 for MEK1 (4 microM) and MEK2 (50 microM). Thus, these data show that alpha1-AR stimulated hypertrophy in ARVM is dependent on the MEK1/2-ERK1/2 signaling pathway. PMID- 11273732 TI - Angiotensin blockade inhibits increased JNKs, AP-1 and NF- kappa B DNA-binding activities in myocardial infarcted rats. AB - Inhibition of the renin-angiotensin system has been shown to prevent left ventricular remodeling after myocardial infarction. However, the effect of angiotensin on the signal transduction pathway of left ventricular remodeling after myocardial infarction is as yet unknown. The purpose of this study was to measure myocardial MAPKs and AP-1, NF- kappa B, and Sp-1 DNA-binding activities after myocardial infarction. Moreover, we evaluated the effects of angiotensin converting enzyme (ACE) inhibitor and angiotensin receptor blocker (ARB) on signal transduction pathway. Myocardial infarction was produced by ligation of the coronary artery in Wistar rats. Temocapril (ACE inhibitor) (3 and 30 mg/kg/day) and candesartan cilexitil (ARB) (1 and 10 mg/kg/day) were orally administered once a day. After ligation of the left descending coronary artery, JNKs (p46JNK and p55JNK) increased to 2.0- (P<0.01) and 2.8-fold (P<0.01) at 7 days, respectively. ERKs (p44ERK and p42ERK) and p38 activities did not increase significantly. AP-1 and NF- kappa B binding activities increased at 5 days, reached their peak 2.2- and 2.0-fold at 7 days. Sp-1 did not change. ACE inhibitor and ARB inhibited JNKs, NF- kappa B and AP-1 activities. Increased JNKs, AP-1, NF- kappa B, and Sp-1 DNA-binding activities were suppressed by both drugs in the infarcted region. Doppler-echocardiography showed that ACE inhibitor and ARB prevented the dilatation of left ventricular cavity at 14 days and improved diastolic filling pattern. JNKs, AP-1 and NF- kappa B activation in myocardial infarcted rats could be responsible for left ventricular remodeling after myocardial infarction and angiotensin may be related to the activation of these signals. PMID- 11273733 TI - Thyroid hormone increases pacemaker activity in rat neonatal atrial myocytes. AB - The effects of thyroid hormone (3,3',5-triiodo- L -thyronine, T3) on pacemaker activity were studied with electrophysiological and pharmacological approaches using spontaneously beating neonatal atrial myocytes cultured from 2-day-old rats. Treatment with T3 (10(-8)m) for 24-48 h led to a positive chronotropic effect. The beating rate of T3-treated cells was 244+/-19 beats/min and for control cells it was 122+/-10 beats/min (P<0.05). Action potentials were recorded and showed that the predominant effect of T3 was to increase the diastolic depolarization rate (99.5+/-9.8 in T3-treated group v 44.0+/-7.8 mV/s in untreated group). Some cells that exhibited pacemaker activity lacked a pacemaker current (I(f)) under voltage clamp conditions I(f)was recorded in 5 of 12 spontaneously active control cells and in 6 of 10 T3-treated cells. In those cells exhibiting the pacemaker current, the I(f)density was significantly larger in T3-treated cells (-7.9+/-2.6 pA/pF v-1.8+/-0.5 pA/pF in control). The L-type Ca2+ current density was similar in the two groups (at -7 mV, -7.5+/-1.5 in treated group v-8.6+/-1.0 pA/pF in control). In the presence of T3, the Na+-Ca2+ exchanger current (I(Na/Ca)) density was larger (e.g. at +60 mV, it was 4.8+/-0.5 v 3.5+/-0.2 pA/pF in control cells, P<0.05). As intracellular Ca2+ is extruded from the cell, the electrogenic Na+-Ca2+ exchanger causes a declining inward current, which may contribute to the pacemaker potential-this declining inward current was demonstrated using the action potential voltage clamp technique and was shown to be larger in T3-treated myocytes. Our data demonstrate that thyroid hormone enhances pacemaker activity and that this may be due in part to an increased Na+-Ca2+ exchanger activity. PMID- 11273734 TI - Targeted deletion of the A3 adenosine receptor confers resistance to myocardial ischemic injury and does not prevent early preconditioning. AB - We used mice with genetic disruption of the A3 adenosine receptor (AR) gene (A3AR(-/-)mice) to assess the in vivo role of the A3AR in modulating myocardial ischemia/reperfusion injury and preconditioning (PC). Surprisingly, infarct size induced by 30 min of coronary artery occlusion and 24 h of reperfusion was 35% smaller in A3AR(-/-)compared to wild-type mice (A3AR(+/+)). The reduction in infarct size was not the result of differences in heart rate, body temperature or increased cardiac expression of A1ARs. However, neutrophil infiltration within infarcted regions was less in A3AR(-/-)mice. Furthermore, ischemic PC induced by either a single episode (one 5 min occlusion) or multiple episodes (six 4 min occlusions) of ischemia produced equivalent reductions in infarct size in A3AR(-/ )and A3AR(+/+)mice. These results indicate that, in the mouse, (i) A3ARs play an injurious role during acute myocardial ischemia/reperfusion injury, possibly by exacerbating the inflammatory response, and (ii) A3ARs are not necessary for the development of the early phase of ischemic PC. PMID- 11273735 TI - Mitochondrial K(ATP) channel opening is important during index ischemia and following myocardial reperfusion in ischemic preconditioned rat hearts. AB - We have previously demonstrated that K(ATP)channel openers administered just prior to and throughout reperfusion induce cardioprotection in the blood-perfused canine heart. However, a recent report suggests that the mitochondrial K(ATP)channel is only a trigger of ischemic preconditioning (IPC). These recent data are, however, in contrast to most previous investigations that suggested that activation of the mitochondrial K(ATP)channel is an important downstream mediator of cardioprotection. Therefore, we examined the role of the mitochondrial K(ATP)channel as a downstream mediator of IPC in a rat model by administering the selective mitochondrial K(ATP)channel antagonist, 5 hydroxydecanoate (5-HD), at several points during IPC. Infarct size (IS) was determined by tetrazolium chloride staining and expressed as a percent of the area at risk (AAR). Control animals had an IS/AAR of 58.4+/-0.6 and IS/AAR was reduced to 6.2+/-1.7 following IPC. 5-HD (10 mg/kg), attenuated cardioprotection when administered either 5 min prior to the IPC stimulus (40.4+/-1.4), during the reperfusion phase of the IPC stimulus (39.7+/-5.9), or 5 min prior to reperfusion during prolonged ischemia (34.3+/-6.9). Additionally, when 5-HD was administered at 5 mg/kg during the reperfusion phase of index ischemia plus 5 min prior to IPC or plus during the reperfusion phase of IPC, cardioprotection was also attenuated (36.3+/-5.5 and 43.8+/-6.9, respectively). These data suggest that activation of the mitochondrial K(ATP) channel is an important downstream regulator of myocardial protection with effects lasting into the reperfusion period following prolonged ischemia. PMID- 11273736 TI - Expression of molecular markers for bone formation increases during experimental acute otitis media. AB - Bony tissues are integral parts of the function of the middle ear and the protection of adjacent vital structures. To explore the reaction of middle ear bone to acute otitis media, rats were challenged with Streptococcus pneumoniae and Haemophilus influenzae. Local changes were monitored for up to 1 month. After reverse transcription, competitive polymerase chain reaction was used to determine the expression levels of two molecular markers of bone formation, osteocalcin and procollagen I, and the two cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-alpha, in the bone. Middle ear bone responded rapidly to bacterial challenge, and the reaction depended upon the causative agent. On day 1, IL-6 and TNF-alpha transcripts were detected in the bone from all middle ears. After a short period of decreased expression of osteocalcin, during which the otitis diagnosis could not be made clinically, the levels of bone formation markers increased dramatically. The maximum levels of these markers were reached on days 6 and 14 for animals challenged with H. influenzae and pneumococci, respectively. Infections induced by pneumococci had a longer duration, and after the initial phase the production of osteocalcin and procollagen transcript were significantly higher in the pneumococcus-infected animals. The results indicate that even in an uncomplicated infection, the bone of the bulla reacts to an acute otitis media with a short period of inhibited osteoblast activity followed by a longer period of new bone formation. PMID- 11273737 TI - Modulating effects of mucoregulating drugs on the attachment of Haemophilus influenzae. AB - Non-typable Haemophilus influenzae (NTHI) is one of the three major pathogens implicated in human respiratory infections. The ability to attach with pharyngeal epithelial cells is an important factor for infection and virulence. In the present study we describe the effects of two mucoregulating drugs, S carboxymethylcysteine (S-CMC) and ambroxol, on the attachment of NTHI to pharyngeal epithelial cells. There was a significant (P < 0.0001, < 0.001 and <0.01) decrease of attachment (8.8 +/ 2.4, 9.2+/-2.5 and 15.4 +/- 5.7 bactreria/cell) compared with the control (17.5 +/- 2.9, 15.5 +/- 3.1 and 18.8 +/ 6.8 bacteria/cell) after cells were treated wth S-CMC at a dose of 100, 10 and 1 microg/ml. After attachment assay, cells treated with S-CMC (100 microg/ml) showed a significant decrease (P < 0.01) of attached bacteria (3.1 +/- 0.8 bacteria/cell) compared with the control (5.9 +/- 1.8 bacteria/cell). Treatment of cells with ambroxol did not influence bacterial attachment. By scanning electron microscopic observation it was found that NTHI attaches to the surface elevations (microplicae) of human pharyngeal epithelial cells. Atomic force microscopic observation revealed that the surface potential of microplicae decreased significantly in cells treated with S-CMC compared with the untreated control cells. As bacteria with negative surface charge attach to the positively charged domain, i.e. microplicae of human pharyngeal epithelial cells, this study suggests that the decrease of attachment of NTHI with epithelial cells after treatment with S-CMC was possibly due to the decrease of surface charge. This study suggests that S-CMC decreases the episodes of respiratory infections in patients with respiratory diseases both by inhibiting the attachment of bacteria to the upper respiratory tract, and by detaching the adherent one. PMID- 11273738 TI - Identification of the mycobacterial DNA-binding protein 1 region which suppresses transcription in vitro. AB - We recently identified a novel DNA-binding protein from Mycobacterium bovis bacillus Calmette-Guerin (BCG), termed mycobacterial DNA-binding protein 1 (MDP1). MDP1 inhibited the in vitro syntheses of DNA, RNA and protein, and reduced growth rates of bacteria transformed with MDP1 genes. In this study, we examined the DNA binding regions of MDP1 by using a set of synthetic peptides. One dominant region was determined on peptide 4 composed of amino acids, at positions 31-50. The peptide 4 inhibited syntheses of both DNA and RNA in vitro. The critical amino acids residues for these functions were analysed utilizing synthetic peptides substituted with Ala. This domain was perfectly conserved in MDP1 homologues in mycobacteria, but not observed in other known DNA binding proteins. These results indicate mycobacteria possess a unique nuclear protein, which might be involved in growth regulation of these organisms. PMID- 11273739 TI - Murine monoclonal antibodies to PorA of Neisseria meningitidis show reduced protective activity in vivo against B:15:P1.7,16 subtype variants in an infant rat infection model. AB - The major outer membrane protein PorA of Neisseria meningitidis is the target for bactericidal serosubtyping antibodies and is currently considered as a potential vaccine candidate against group B meningococcal disease. Although the minor antigenic variability of the PorA has been increasingly recognized and described, its implication for vaccine design remains unclear. In this study, the protective activity of murine monoclonal PorA specific antibodies against four isogenic meningococcal P1.7,16 target strains, the prototype P1.7,16a and three loop 4 point mutation variants (designated P1.7,16b to d) constructed from reference strain H44/76 (B:15:P1.7,16a), was evaluated in the infant rat infection model. All monoclonal antibodies had been obtained by immunization of mice with outer membrane protein preparations from meningococcal serosubtype P1.7,16 reference strain H44/76. A challenge dose of 10(5)cfu/pup was given i.p. 1-2 h after the i.p. injection of 1:100 diluted antibodies, and the development of bacteremia was assessed by culturing blood samples taken 6 h after challenge. MN14C11.6, a reference monoclonal antibody for serosubtype P1.7 epitope located in predicted loop 1 (VR1) identical in all the variants, was equally protective against all loop 4 variants. The three P1.16 specific monoclonal antibodies tested (MN5C11G, MN12H2 and 62D12-8) all completely protected animals against the prototype P1.7,16a, variably against the P1.7,16b and P1.7,16c, but not against the P1.7,16d variant. Our findings therefore suggest that certain subtype variants may escape protection in vivo conferred by PorA specific antibodies. PMID- 11273740 TI - Chlamydia pneumoniae facilitates monocyte adhesion to endothelial and smooth muscle cells. AB - Chlamydia pneumoniae has been linked to atherosclerotic heart disease. However, there is a limited knowledge by which C. pneumoniae gain access to atheromatous lesions. The adhesion of C. pneumoniae -infected circulatory component(s) to endothelium and smooth muscle cells represents the first step in an inflammatory response. We examined the ability of viable as well as heat inactivated C. pneumoniae to infect human monocytes and subsequently the ability of infected monocytes to adhere to human coronary artery endothelial cells (HCAEC) and human coronary smooth muscle cells (HCSMC). Our results demonstrate susceptibility of monocytes to in vitro chlamydial infection. Inclusions of varying sizes and intensities were observed 3-5 days after inoculation with viable C. pneumoniae. Monocytes infected with heat inactivated organisms revealed no inclusions, in keeping with the observations of uninfected monocytes. Moreover, monocytes infected with viable C. pneumoniae adhered preferentially to HCAEC and HCSMC, as compared to uninfected monocytes or monocytes harbouring heat inactivated Chlamydia. PMID- 11273741 TI - The impact of Haemophilus ducreyi cytolethal distending toxin on cells involved in immune response. AB - The Haemophilus ducreyi cytolethal distending toxin (HdCDT) induces cell cycle arrest and thereby inhibits cell proliferation of many cultured mammalian cell lines. We investigated the effect of HdCDT on circulating human hematopoietic cells, including T- and B-cells, monocytes and polymorphonuclear cells (PMN). Lymphocytes were stimulated with T- and B-cell specific mitogens, whereas monocytes and PMN with endotoxin. HdCDT inhibited the mitogen-induced proliferation of T-cells in a dose-dependent manner as assayed by [(3)H] thymidine incorporation and MTT assays. Similarly to T-cells, HdCDT also inhibited the proliferation of B-cells and consequently the immunoglobulin production, measured by ELISPOT and ELISA assays. In contrast, the HdCDT did not affect monocytes or PMN, as measured by MTT assay. The TNF-alpha production by monocytes and the phagocytic ability of PMN were neither affected. The monocytic cell line THP-1 was, however, sensitive to the toxin, seen as a reduction of proliferation and viability after exposure to HdCDT. In conclusion, exposure to HdCDT significantly affects the proliferation and other biological activities of stimulated human T- and B-cells, while circulating monocytes and PMN are not sensitive to HdCDT. The sensitivity of cells of the acquired immune system to HdCDT may hamper specific host response to H. ducreyi and contribute to persistence of chancroid lesions. PMID- 11273742 TI - Invasion of human type II pneumocytes by Burkholderia cepacia. AB - Burkholderia cepacia is known to invade and survive within respiratory epithelial cells. Previous studies have employed transformed cell lines and it is not known whether the bacterium is capable of manifesting the same phenomena in primary cell culture. Two strains of B. cepacia of environmental (NCTC 10661) and clinical origin (C1359) were examined for their ability to invade and survive (over a 24 h period) within type II pneumocytes in primary culture using a gentamicin-ceftazidime antibiotic protection assay. Both strains of B. cepacia were capable of invasion of type II pneumocytes in primary culture. Strain C1359 was capable of multiplying intracellularly as indicated by a seven-fold increase in the numbers of bacteria from 4-24 h, whereas strain 10661, although unable to replicate intracellularly, was found to survive in the pneumocytes for at least 24 h. Future studies on the invasiveness of B. cepacia can employ A549 cells as a valid surrogate for primary cell culture assays which are time-consuming, labour intensive and expensive to perform. PMID- 11273743 TI - Role of endotoxin in the pathogenesis of haemorrhagic septicaemia in the buffalo. AB - The pathogenesis of haemorrhagic septicaemia in buffalo infected with Pasteurella multocida is poorly understood. However, the characteristic of sudden onset leading to the rapid death of infected animals is similar to that seen in other clinical conditions known to involve endotoxic shock. The objectives of the work were to assess the contribution of endotoxaemia to the disease's pathogenesis and to characterize the pathophysiological reaction, including the acute phase response, of buffalo to experimental infection with P. multocida serotype B:2, the bacterium responsible for the disease in Asia. After intranasal infection of eight buffaloes with a culture of a field isolate of P. multocida serotype B:2, three animals succumbed to the disease at 26-30 h post-infection (p.i.) and five survived. Rectal temperatures of infected animals rose to a peak at about 10 h p.i. and surviving animals showed a second peak in rectal temperature at 36 h p.i. Endotoxin was present only in serum of non-surviving animals 3-5 h before death or killing during which time concentrations increased rapidly, correlating with the development of overt clinical signs and reductions in rectal temperature, concentrations of white blood cells, serum thyroxine, iron, copper and zinc, an increase in serum haptoglobin and cortisol and the detection of a low-grade bacteraemia. A strong acute phase response was maintained in surviving animals with a progressive rise in serum haptoglobin over 96 h p.ia slow rise in the serum copper concentration from 24 h p.i. and an increase, from about 65 h p.iin serum alpha(1)-acid glycoprotein. The findings demonstrate that a progressive endotoxaemia and associated sequelae correlates with the development of overt haemorrhagic septicaemia disease and sudden death in buffalo. PMID- 11273744 TI - Time averaging and fitting of nonlinear metabolic changes: the issue of the time index choice applied to 31P MRS investigation of muscle energetics. AB - We present an exact analytical method dedicated to fitting time-dependent exponential-like changes in MR spectra. As an illustration, this method has been applied to fitting metabolic changes recorded by 31P MRS in human skeletal muscle occurring during a rest-exercise-recovery protocol. When recording metabolic changes with the accumulative method, the time averaging of the MR signals implies the choice of a time index for fitting any changes in the features of the associated MR spectra. A critical examination of the different ways (constant, linear, and exponential) of choosing the time index is reported. By numerical analysis, we have calculated the errors generated by the three methods and we have compared their sensitivity to noise. In the case of skeletal muscle, both constant and linear methods introduce large and uncontrolled errors for the whole set of metabolic parameters derived from [PCr] changes. In contrast, the exponential method affords a reliable estimation of critical parameters in muscle bioenergetics in both normal and pathological situations. This method is very easy to implement and provides an exact analytical solution to fitting changes in MR spectra recorded by the accumulative method. PMID- 11273745 TI - On the importance of exchangeable NH protons in creatine for the magnetic coupling of creatine methyl protons in skeletal muscle. AB - The methyl protons of creatine in skeletal muscle exhibit a strong off-resonance magnetization transfer effect. The mechanism of this process is unknown. We previously hypothesized that the exchangeable amide/amino protons of creatine might be involved. To test this the characteristics of the creatine magnetization transfer effect were investigated in excised rat hindleg skeletal muscle that was equilibrated in either H2O or D2O solutions containing creatine. The efficiency of off-resonance magnetization transfer to the protons of mobile creatine in excised muscle was similar to that previously reported in intact muscle in vivo. Equilibrating the isolated muscle in D2O solution had no effect on the magnetic coupling to the immobile protons. It is concluded that exchangeable protons play a negligible role in the magnetic coupling of creatine methyl protons in muscle. PMID- 11273747 TI - Magnet design with high B(0) homogeneity for fast-field-cycling NMR applications. AB - The design, construction, and performance of a low-inductance solenoidal coil with high B(0) homogeneity for fast-field-cycling NMR is presented. It consists of six concentric layers. The conductor width is varied to minimize the B(0) inhomogeneity in the volume of the sample. This is done using an algorithm which takes the real shape of the conductor directly into account. The calculated coil geometry can be manufactured easily using standard computerized numeric control equipment, which keeps the costs low. The coil is liquid cooled and produces a B(0) field of 0.95 T at 800 A. The field inhomogeneity in a cylindrical volume (diameter 5 mm, length 10 mm) is about 10 ppm, and the inductance is 190 microH. Switching times below 200 micros can be achieved. During 6 months of operation the coil has shown good stability and reliability. PMID- 11273753 TI - Fourier-transform EPR at high-field/high-frequency (3.4 T/95 GHz) using broadband stochastic microwave excitation. AB - Stochastic excitation with a full-width-half-maximum bandwidth of 250 MHz was used to perform Fourier-transform (FT) high-field/high-frequency electron paramagnetic resonance (EPR) at 3.4T/95 GHz (W-band). Thereby, the required microwave peak power is reduced by a factor of tau(p)/T1 as compared to equivalent pulsed FT EPR in which the spin system with spin-lattice relaxation time T1 is excited by a single microwave pulse of length tau(p). Stochastic EPR is particularly interesting under high-field/high-frequency conditions, because the limited output power of mm microwave sources, amplifiers, and mixers makes pulse FT EPR in that frequency domain impossible, at least for the near future. On the other hand, FT spectroscopy offers several advantages compared to field swept magnetic resonance methods, as is demonstrated by its success in NMR and X band EPR. In this paper we describe a novel stochastic W-band microwave bridge including a bimodal induction mode transmission resonator that serves for decoupling the microwave excitation and signal detection. We report first EPR measurements and discuss experimental difficulties as well as achieved sensitivity. Moreover, we discuss future improvements and the possibility for an application of stochastic W-band FT EPR to transient signals such as those of photoexcited radical pairs in photosynthetic reaction centers. PMID- 11273750 TI - Water modeled signal removal and data quantification in localized MR spectroscopy using a time-scale postacquistion method. AB - We have previously shown the continuous wavelet transform (CWT), a signal processing tool, which is based upon an iterative algorithm using a lorentzian signal model, to be useful as a postacquisition water suppression technique. To further exploit this tool we show its usefulness in accurately quantifying the signal metabolites after water removal. However, due to the static field inhomogeneities, eddy currents, and "radiation damping," the water signal and the metabolites may no longer have a lorentzian lineshape. Therefore, another signal model must be used. As the CWT is a flexible method, we have developed a new algorithm using a gaussian model and found that it fits the signal components, especially the water resonance, better than the lorentzian model in most cases. A new framework, which uses the two models, is proposed. The framework iteratively extracts each resonance, starting by the water peak, from the raw signal and adjusts its envelope to both the lorentzian and the gaussian models. The model giving the best fit is selected. As a consequence, the small signals originating from metabolites when selecting, removing, and quantifying the dominant water resonance from the raw time domain signal are preserved and an accurate estimation of their concentrations is obtained. This is demonstrated by analyzing (1H) magnetic resonance spectroscopy unsuppressed water data collected from a phantom with known concentrations at two different field strengths and data collected from normal volunteers using two different localization methods. PMID- 11273754 TI - Electron spin relaxation time measurements using radiofrequency longitudinally detected ESR and application in oximetry. AB - Longitudinally detected ESR (LODESR) involves transverse ESR irradiation with a modulated source and observing oscillations in the spin magnetization parallel to the main magnetic field. In this study, radiofrequency-LODESR was used for oximetry by measuring the relaxation times of the electron. T1e and T2e were measured by investigating LODESR signal magnitude as a function of detection frequency. We have also predicted theoretically and verified experimentally the LODESR signal phase dependence on detection frequency and relaxation times. These methods are valid even for inhomogeneous lines provided that T1e>>T2e. We have also developed a new method for measuring T1e, valid for inhomogeneous spectra, for all values of T1e and T2e, based on measuring the spectral area as a function of detection frequency. We have measured T1e and T2e for lithium phthalocyanine crystals, for the nitroxide TEMPOL, and for the single line agent Triarylmethyl (TAM). Furthermore, we have collected spectra from aqueous solutions of TEMPOL and TAM at different oxygen concentrations and confirmed that T1e values are reduced with increased oxygen concentration. We have also measured the spin lattice electronic relaxation time for degassed aqueous solutions of the same agents at different agent concentrations. T1e decreases as a function of concentration for TAM while it remains independent of free radical concentration for TEMPOL, a major advantage for oxygen mapping. This method, combined with the ability of LODESR to provide images of exogenous free radicals in vivo, presents an attractive alternative to the conventional transverse ESR linewidth based oximetry methods. PMID- 11273755 TI - Mechanism of oxygen response in carbon-based sensors. AB - The mechanism of oxygen response in several newly synthesized oxygen-sensitive chars was studied with the use of EPR spectroscopy. The results suggest that the compounds contain two basic types of paramagnetic centers (PC). The change in oxygen concentration leads to a mutual and reversible transformation of PCs in chars, which is reflected in EPR parameters. The adsorbed molecular oxygen progressively disturbs the wave functions of the PCs and so breaks the Heisenberg exchange between them. At high oxygen concentration, the 2D dipole-dipole interaction between PCs at the surface comes into play and determines the EPR lineshape. A suggested model quantitatively describes the evolution of the basic EPR parameters of each PC as a function of oxygen concentration. PMID- 11273757 TI - Symmetric echo acquisition for absolute-value display in solid-state NMR imaging. AB - A method of solid-state NMR imaging that permits echo Fourier transformation (FT) has been devised using a magic echo train. The echo FT imaging can be implemented simply by modifying the gradient pulse sequence in the previous magic echo imaging (TREV-16TS) so that the one-dimensional k-space trajectory follows the sampling points which are symmetric about the k origin. The implemented ability of echo FT improves the performance of the magic echo imaging: the sensitivity gained by radical2, the phase correction is made unnecessary, and the digital resolution is doubled. One- and two-dimensional imaging experiments have been conducted on some solid samples, confirming the improved performance and revealing a TREV-16TS adjustment parameter that is critical for the successful echo FT imaging. PMID- 11273758 TI - Simultaneous determination of orientational and order parameter distributions from NMR spectra of partially oriented model membranes. PMID- 11273760 TI - Two-voxel localization sequence for in vivo two-dimensional homonuclear correlation spectroscopy. AB - The combination of localized 2D 1H MR correlation spectroscopy and Hadamard encoding allows the simultaneous acquisition of multiple volumes of interest without an increase in the experimental duration, compared to single-voxel acquisition. In the present study, 2D correlation spectra were acquired simultaneously within 20 to 40 min in two voxels located in each hemisphere of the rat brain. An intervoxel distance of 20% of the voxel size was sufficient to limit spatial contamination. The following cerebral metabolites gave detectable crosspeaks: N-acetylaspartate, the glutamate/glutamine pool, aspartate, phosphoethanolamine, glucose, glutathione, taurine, myo-inositols, lactate, threonine, gamma-aminobutyric acid, and alanine. Most of the metabolites were measured without contamination of other resonances. PMID- 11273761 TI - In vivo 31P echo-planar spectroscopic imaging of human calf muscle. AB - Localized phosphorus-31 NMR spectra of human calf muscle in vivo were obtained by means of echo-planar spectroscopic imaging (EPSI) with a 1.5-T whole-body scanner. The technique permits the measurement of two-dimensional 31P SI data at a minimum acquisition time of 2.4 s (8x8 voxels, TR=300 ms). With 9.4 min measurement time (TR=1100 ms, 64 averages) and 25x25x40 mm spatial resolution in vivo the 31P NMR signal-to-noise ratio (S/N) of the phosphocreatine (PCr) resonance was about 45; the multiplets of nucleoside 5'-triphosphates were resolved. Spectral quality permits quantitative assessment of the PCr signal in a measurement time that is shorter by a factor of 2 or more than the minimum measurement time feasible with chemical-shift imaging. In a functional EPSI study with a time resolution of 20.5 s on the calf muscle of volunteers, spectra showed a 40% decrease of the PCr signal intensity (at rest: S/N congruent with12) upon exertion of the muscle. PMID- 11273765 TI - Characterization of hydrogen bond lengths in Watson-Crick base pairs by cross correlated relaxation. AB - Hydrogen bond lengths in Watson-Crick base pairs can be characterized by cross correlated relaxation between 1H chemical shift anisotropy and dipole-dipole coupling of 1H and its hydrogen bond acceptor 15N. As a reference, the cross correlated relaxation between 1H chemical shift anisotropy and dipole-dipole coupling of 1H and its hydrogen bond donor 15N is used. With the two measured cross-correlated relaxation rates, an apparent hydrogen bond length can be determined, which is composed by the hydrogen bond length multiplied by a term representing the amplitude of inter-base motions. Data are presented for the 15N3 1H3...15N1 hydrogen bonds in A=T base pairs of the Antennapedia homeodomain-DNA complex with a correlation time of global rotational diffusion of 20 ns. PMID- 11273766 TI - Characterization of the cholesteric phase of filamentous bacteriophage fd for molecular alignment. AB - Residual dipolar couplings arise from small degrees of alignment of molecules in a magnetic field and have proven to provide valuable structural information. Colloidal suspensions of rod-shaped viruses and bacteriophages constitute a frequently employed medium for imparting such alignment onto biomolecules. The stability and behavior of the liquid crystalline phases with respect to solution conditions such as pH, ionic strength, and temperature vary, and characterization should benefit practical applications as well as theoretical understanding. In this Communication we describe the pH dependence of the cholesteric liquid crystalline phase of the filamentous bacteriophage fd and demonstrate that the alignment tensor of the solute protein is modulated by pH. We also report the interesting observation that the relative sign of the residual dipolar coupling changes at low pH values. In addition, we demonstrate that the degree of alignment inversely scales with the lengths of the phage particles for phages with identical mass and charge per unit length. PMID- 11273768 TI - Augmenting transgene expression from carcinoembryonic antigen (CEA) promoter via a GAL4 gene regulatory system. AB - Though extensively studied, the use of tissue- or cell-type-specific promoters to target transgene expression is hampered by their weak activity. We hypothesized that this problem could be addressed by using a GAL4 gene regulatory system, wherein a weak, tissue-specific promoter would drive expression of the GAL4/VP16 fusion protein (GV16), which in turn would transactivate a minimal synthetic promoter, GAL4/TATA (GT), upstream of a transgene. To test this hypothesis, we constructed adenoviral vectors expressing a lacZ or GV16 gene driven by a carcinoembryonic antigen (CEA) promoter (Ad/CEA-LacZ or Ad/CEA-GV16) and evaluated levels of transgene expression they produced in cultured cells and in subcutaneous tumors after intratumoral administration. In CEA-positive cells, treatment with Ad/CEA-GV16 + Ad/GT-LacZ versus Ad/CEA-LacZ increased transgene expression 20- to 100-fold. In CEA-negative cells, treatment with Ad/CEA-GV16 + Ad/GT-LacZ increased transgene expression to a much lower degree (6- to 8-fold). In addition, analysis of Bax gene-mediated cell death revealed that this system can be used to avoid Bax's toxic effects on CEA-negative cells without compromising its ability to kill CEA-positive cells in vitro and in vivo. Thus, the combination of a tissue-specific promoter with the GAL4 gene regulatory system could be useful for targeting transgene expression. PMID- 11273769 TI - In utero delivery of adeno-associated viral vectors: intraperitoneal gene transfer produces long-term expression. AB - Recombinant adeno-associated viruses (rAAV) are promising gene transfer vectors that produce long-term expression without toxicity. To investigate future approaches for in utero gene delivery, the efficacy and safety of prenatal administration of rAAV were determined. Using luciferase as a reporter, expression was assessed by whole-body imaging and by analysis of luciferase activity in tissue extracts, at the time of birth and monthly thereafter. Transgene expression was detected in all injected animals. Highest levels of luciferase activity were detected at birth in the peritoneum and liver, while the heart, brain, and lung demonstrated low-level expression. In vivo luciferase imaging revealed persistent peritoneal expression for 18 months after in utero injection and provided a sensitive whole-body assay, useful in identifying tissues for subsequent analyses. There was no detectable hepatocellular injury. Antibodies that reacted with either luciferase or rAAV were not found. AAV sequences were not detected in germ-line tissues of injected animals or in tissues of their progeny. In utero AAV-mediated gene transfer in this animal model demonstrates that novel therapeutic vectors and strategies can be rapidly tested in vivo and that rAAV may be developed to ameliorate genetic diseases with perinatal morbidity and mortality. PMID- 11273770 TI - An immunomodulatory procedure that stabilizes transgene expression and permits readministration of E1-deleted adenovirus vectors. AB - Immune responses against E1-deleted adenovirus vectors and/or their transgene products result in the rapid elimination of vector-transduced cells and the generation of neutralizing antibodies. Different strategies of immunomodulation to stabilize transgene expression at therapeutic levels and to permit productive vector readministration have been examined. Our previous studies have shown that depletion of macrophages from spleen and liver decreases hepatic inflammation, significantly prolongs transgene expression, and delays the onset of humoral immune responses after systemic administration of an E1-deleted adenovirus vector. In the present study, we have examined the effects of macrophage depletion in combination with temporary blockade of CD40 ligation on E1-deleted adenovirus vector-mediated gene transfer. Alone, each of these treatments significantly inhibited the humoral immune response against the transgene product and prolonged its expression. Together, these treatments completely stabilized transgene expression and inhibited the production of neutralizing anti-adenovirus antibodies, permitting successful vector readministration. Animals rendered immunologically unresponsive to vector and transgene antigens regained their ability to mount productive immune responses against the vector after recovery of immune function, but remained unresponsive to the transgene product. These experiments demonstrate that this treatment is transient and antigen-specific. PMID- 11273772 TI - Intracellular delivery of a Tat-eGFP fusion protein into muscle cells. AB - The Tat protein from HIV-1, when fused with heterologous proteins or peptides, can traverse biological membranes in a process called "protein transduction," delivering its cargo into cells. A Tat-eGFP fusion protein was purified from bacteria to study the transduction kinetics of Tat fusion proteins into cultured myoblasts and in the muscle tissue. Correctly folded Tat-eGFP reaches a maximum intracellular level in nearly 30 min, while its endogenous fluorescence is first detected only after 14 h. The nuclear localization signal from the basic domain of Tat was not sufficient to confer nuclear localization to Tat-eGFP, suggesting that the nuclear import pathway used by the exogenously added Tat-eGFP might be sensitive to the folding state of eGFP. In mice, the direct delivery to the muscle tissue using subcutaneous injections or the intra-arterial pathway led to few positive fibers in the muscle periphery or surrounding the blood vessels. Muscles injected with Tat-eGFP showed intense labeling of the extracellular matrix (ECM), suggesting that, although Tat fusion proteins can transduce muscle fibers, their binding by components of the ECM surrounding myofibers could interfere with the intracellular transduction process. PMID- 11273771 TI - Correction of liver disease following transplantation of normal rat hepatocytes into Long-Evans Cinnamon rats modeling Wilson's disease. AB - To establish the efficacy of cell therapy in Wilson's disease, we used the Long Evans Cinnamon (LEC) rat model with atp7b gene mutation and copper toxicosis. Several groups of LEC rats were established, including animals pretreated with retrorsine to exacerbate copper toxicosis and inhibit proliferation in native hepatocytes followed by partial hepatectomy to promote liver repopulation. Hepatocytes from normal, syngeneic LEA rats were transplanted intrasplenically. Animal survival, biliary copper excretion, and hepatic copper were determined. The magnitude of liver repopulation was demonstrated by measuring serum ceruloplasmin and hepatic atp7b mRNA. Long-term survival in LEC rats treated with retrorsine, partial hepatectomy, and cell transplantation was up to 90%, whereas fewer than 10% of animals pretreated with retrorsine, without cell therapy, survived, P < 0.001. Liver repopulation occurred gradually after cell transplantation, ranging from <25% at 6 weeks, 26 to 40% at 4 months, and 74 to 100% at 6 months or beyond. Liver repopulation restored biliary copper excretion capacity and lowered liver copper levels. Remarkably, liver histology was completely normal in LEC rats with extensive liver repopulation, compared with widespread megalocytosis, apoptosis, oval cell proliferation, and cholangiofibrosis in untreated animals. These data indicate that liver repopulation with functionally intact cells can reverse pathophysiological perturbations and cure Wilson's disease. PMID- 11273773 TI - Transduction of liver cells by lentiviral vectors: analysis in living animals by fluorescence imaging. AB - Viral vectors based on lentiviruses, such as the human immunodeficiency virus, are able to transduce a broad spectrum of nondividing cells in vivo. This ability of lentiviral vectors makes them an attractive vehicle for gene transfer into the liver. In order to determine the requirements for efficient lentiviral gene transfer, we used a fluorescence imaging system, which allows the detection of cells and tissues that express fluorescent reporter genes (e.g., green fluorescence protein) in the living animal. We show that the latest generation of lentiviral vectors efficiently transduces the murine liver. Further analysis demonstrated that neither cell-cycle activation nor division of liver cells is a prerequisite for lentiviral gene transfer in vivo. PMID- 11273774 TI - Two point mutations increase targeted transduction and stabilize vector association of a modified retroviral envelope protein. AB - The current strategy of targeting retroviral vector transduction by inserting a peptide ligand into the envelope protein has met with several obstacles. These modified proteins redirected vector binding to a new cognate receptor on a specific cell type but gave little or no gene transfer because they did not fuse the vector and target cell membranes. They dissociated readily from vectors and often required coassembly of wild-type envelope protein. Here we report a novel strategy to overcome the fusion and stability defects of modified retroviral envelope proteins. We inserted a prototypic ligand, the receptor binding domain of amphotropic murine leukemia virus, into an ecotropic murine leukemia virus envelope protein mutant containing glutamine 227-to-arginine plus aspartate 243 to-tyrosine substitutions. This modified protein increased transduction redirected to human cells expressing the amphotropic receptor to a level within 10-fold that of wild-type amphotropic virus, an increase of as great as 2000-fold over transduction by modified protein lacking the mutations. In addition to suppressing the fusion defect, these mutations unexpectedly stabilized the association of the modified protein with vector particles. Insertion of clinically relevant ligands into this envelope mutant should improve the efficiency and reliability of retroviral transduction of specific cell types for gene therapy applications. PMID- 11273775 TI - Generation and characterization of E1/E2a/E3/E4-deficient adenoviral vectors encoding human factor VIII. AB - The use of adenoviral vectors for gene therapy has been limited due to host immune responses directed toward the vector and/or transgene and vector toxicity. To decrease adenoviral vector immunogenicity and toxicity, we attenuated viral gene expression by eliminating E1, E2a, E3, and E4 early genes from the adenoviral backbone. Two highly attenuated, fourth-generation (Av4) E1/E2a/E3/E4 deficient adenoviral vectors encoding human factor VIII (FVIII) under the control of a liver-specific albumin promoter were generated. One Av4 vector (Av4DeltaE4FVIII) was deficient in the entire E4 coding region and the second vector contained a deletion of the E4 region, except for open reading frame 3 (orf 3; Av4orf3FVIII). The Av4 vectors were compared to an E1/E2a/E3-deficient third-generation vector (Av3H8101) containing an analogous transgene expression cassette in vitro and in vivo following intravenous administration in hemophiliac mice. In vitro transduction of Hep3B cells revealed at all three vectors expressed functional FVIII. However, the Av4DeltaE4FVIII vector could not be scaled-up for in vivo studies. Both Av3H8101 and Av4orf3FVIII initially expressed similar levels of FVIII in hemophiliac mice. However, at 3 months, animals treated with the Av4orf3FVIII vector no longer expressed FVIII while Av3H8101 treated mice displayed persistent FVIII expression. Liver enzyme analyses of plasma samples revealed that the Av4orf3FVIII vector was significantly less hepatotoxic than the Av3H8101 vector. These data demonstrate that further attenuation of the adenoviral vector backbone by removal of the majority of the E4 coding region significantly diminished vector toxicity; however, the duration of transgene expression was reduced. PMID- 11273776 TI - Successful treatment of primary and disseminated human lung cancers by systemic delivery of tumor suppressor genes using an improved liposome vector. AB - Delivery of therapeutic genes to disseminated tumor sites has been a major challenge in the field of cancer gene therapy due to lack of an efficient vector delivery system. Among the various vectors currently available, liposomes have shown promise for the systemic delivery of genes to distant sites with minimal toxicity. In this report, we describe an improved extruded DOTAP:cholesterol (DOTAP:Chol) cationic liposome that efficiently delivers therapeutic tumor suppressor genes p53 and FHIT, which are frequently altered in lung cancer, to localized human primary lung cancers and to experimental disseminated metastases. Transgene expression was observed in 25% of tumor cells per tumor in primary tumors and 10% in disseminated tumors. When treated with DOTAP:Chol-p53 and -FHIT complex, significant suppression was observed in both primary (P < 0.02) and metastatic lung tumor growth (P < 0.007). Furthermore, repeated multiple treatments revealed a 2.5-fold increase in gene expression and increased therapeutic efficacy compared to single treatment. Finally, animal survival experiments revealed prolonged survival (median survival time: 76 days, P < 0.001 for H1299; and 96 days, P = 0.04 for A549) when treated with liposome-p53 DNA complex. Our findings may be of importance in the development of treatments for primary and disseminated human lung cancers. PMID- 11273777 TI - Intracranial injection of recombinant adeno-associated virus improves cognitive function in a murine model of mucopolysaccharidosis type VII. AB - Mucopolysaccharidosis type VII (MPS VII) is a lysosomal storage disease caused by the lack of beta-glucuronidase (GUSB) activity. GUSB deficiency leads to the progressive accumulation of undegraded glycosaminoglycans (GAGs) in cells of most tissues, including the brain, and is associated with mental retardation. Reduction of lysosomal storage in the central nervous system and prevention of cognitive dysfunction may require intracranial delivery of a therapeutic agent during the newborn period that provides a continuous source of GUSB. Therefore, we injected recombinant adeno-associated virus encoding human GUSB into both the anterior cortex and the hippocampus of newborn MPS VII mice. Total GUSB activity in the brain approached normal levels by 18 weeks. Although GUSB activity was concentrated near the injection sites, lysosomal distension was reduced in most areas of the brain. In addition to histopathologic evidence of GAG reduction, the previously undescribed accumulation of GM2 and GM3 gangliosides in the brain was also prevented. Furthermore, GUSB expression and reduced lysosomal distension correlated with improvements in cognitive function as measured in the Morris Water Maze test. These findings indicate that localized overexpression of GUSB has positive effects on the pathology and cognitive function and does not have overt toxicity. PMID- 11273778 TI - Fibronectin fragment CH-296 inhibits apoptosis and enhances ex vivo gene transfer by murine retrovirus and human lentivirus vectors independent of viral tropism in nonhuman primate CD34+ cells. AB - The fibronectin fragment CH-296 improved gene transfer to cytokine-mobilized nonhuman primate CD34+ cells irrespective of tropism to the MoMLV, GaLV, and VSV G envelope proteins using murine stem cell virus (MSCV) and human immunodeficiency virus-1 (HIV-1)-based retrovirus vectors. For the HIV-1 lentivirus vector, CH-296 enhanced gene transfer in the absence of added hematopoietic growth factors necessary for ex vivo stem cell expansion. In the presence of CH-296, apoptosis of CD34+ cells was inhibited, and in mobilized peripheral blood CD34+ cells, cell division was stimulated as measured by cell history/tracking experiments. PMID- 11273779 TI - Regression of human mammary adenocarcinoma by systemic administration of a recombinant gene encoding the hFlex-TRAIL fusion protein. AB - The tumor necrosis factor (TNF)-related apoptosis-inducing ligand, TRAIL, is a new member of the TNF family. It can specifically induce apoptosis in a variety of human tumors. To investigate the possibility of employing the TRAIL gene for systemic cancer therapy, we constructed a recombinant gene encoding the soluble form of the human Flt3L gene (hFlex) at the 5' end and the human TRAIL gene at the 3' end. Such design allows the TRAIL gene product to be secreted into the body circulation. We have also demonstrated that the addition of an isoleucine zipper to the N-terminal of TRAIL greatly enhanced the trimerization of the fusion protein and dramatically increased its anti-tumor activity. The fusion protein reached the level of 16-38 microg/ml in the serum after a single administration of the recombinant gene by hydrodynamic-based gene delivery in mice. A high level of the fusion protein correlated with the regression of a human breast tumor established in SCID mice. No apparent toxicity was observed in the SCID mouse model. In addition, the fusion protein caused an expansion of the dendritic cell population in the C57BL/6 recipient mice, indicating that the hFlex component of the fusion protein was functional. Thus, the hFlex-TRAIL fusion protein may provide a novel approach, with the possible involvement of dendritic cell-mediated anti-cancer immunity, for the treatment of TRAIL sensitive tumors. PMID- 11273780 TI - Nonviral gene delivery to the lateral ventricles in rat brain: initial evidence for widespread distribution and expression in the central nervous system. AB - The use of DNA for nonviral gene expression depends on several factors. These include (i) delivery and accessibility to the targeted tissue, (ii) protection from extracellular degradation, (iii) sufficient uptake by cells of interest, and (iv) protection from intracellular degradation to allow translation of adequate levels of intracellular or secreted proteins. As an initial step in demonstrating the feasibility of nonviral, cationic lipid-mediated gene therapy, we present evidence for the successful delivery and expression of heat shock protein Hsp70 and reporter gene enzymes in the central nervous system (CNS) of the rat after injection into the lateral ventricle. Gene delivery is accomplished using optimized formulations of plasmid DNA, which have been complexed with the cationic lipid MLRI. Results from DNA vectors encoding for green fluorescent protein (GFP), luciferase, and Hsp70 are reported. Standard immunofluorescent methods were used to demonstrate widespread expression of the reporter proteins and of Hsp70. Stereology analysis has been completed on three coronal sections, which illustrates the distribution of expression along the longitudinal axis. These initial findings support the further development of nonviral, lipid mediated gene delivery technology for transient expression of protective, intracellular proteins and represent an important step leading to in vivo studies to identify potential clinical benefits. PMID- 11273781 TI - Characterization of the cyclooxygenase-2 promoter in an adenoviral vector and its application for the mitigation of toxicity in suicide gene therapy of gastrointestinal cancers. AB - The application of adenoviral molecular chemotherapy for systemic malignant disease using herpes simplex virus thymidine kinase has been limited by ectopic transgene expression in the liver due to the vector hepatotropism. The aim of this study was to mitigate this hepatotoxicity using the promoter of cyclooxygenase-2, inactive in liver but active in many gastrointestinal cancers. To analyze the specificity of transgene expression driven by cyclooxygenase-2 (cox-2) promoters, promoters of two different lengths were incorporated into adenoviral vectors to drive luciferase expression. The specific cytocidal effect and in vivo toxicity were analyzed with thymidine kinase expression vectors. The specificity of the cox-2 promoter was well preserved in the adenoviral vector. In vivo, the cox-2 promoter (-1432/+59) showed very little activity in the liver but attained high activity, comparable to that of the cytomegalovirus promoter, in cyclooxygenase-2-positive subcutaneous tumors. The cox-2 promoter-driven thymidine kinase-expressing vectors showed a cytocidal effect specifically in cyclooxygenase-2-positive cells. When mice were treated with the thymidine kinase expressing vector and ganciclovir, the cox-2 promoter successfully mitigated the fatal hepatotoxicity, which was observed with the cytomegalovirus promoter-driven vector. The cox-2 promoter successfully mitigated the adverse effects of adenoviral suicide gene therapy by minimizing transgene expression in the liver. PMID- 11273782 TI - Infection of human airway epithelia with H1N1, H2N2, and H3N2 influenza A virus strains. AB - Three subtypes of influenza A virus cause human disease: H1N1, H2N2, and H3N2. Although all result in respiratory illness, little is known about how these subtypes infect differentiated airway epithelia. Therefore, we assayed A/PR/8/34 (H1N1), A/Japan/305/57 (H2N2), and X31 (H3N2) influenza virus strains for binding and infection on fully differentiated primary cultures of airway epithelia isolated from human bronchus, grown on semiporous filters at an air-liquid interface. In this model system, viral infectivity was highest when virus was applied to the apical versus the basolateral surface; Japan was most infectious, followed by PR8. The X31 strain showed very low levels of infectivity. Confocal microscopy and fluorescence-resonance energy transfer studies indicated that Japan virus could enter and fuse with cellular membranes, while infection with X31 virions was greatly inhibited. Japan virus could also productively infect human trachea explant tissues. These data show that influenza viruses with SAalpha2,3Gal binding specificity, like Japan, productively infect differentiated human airway epithelia from the apical surface. These data are important to consider in the development of pseudotyped recombinant viral vectors for gene transfer to human airway epithelia for gene therapy. PMID- 11273783 TI - Linear DNAs concatemerize in vivo and result in sustained transgene expression in mouse liver. AB - The short duration of transgene expression remains a major obstacle for the implementation of nonviral DNA vectors in clinical gene therapy trials. Here, we demonstrate stable, long-term transgene expression in vivo by transfecting a linear DNA expression cassette (LDNA) into mouse liver. Interestingly, despite similar quantities and cellular distribution of injected DNAs in their livers, mice receiving LDNA encoding human alpha1-antitrypsin (hAAT) expressed approximately 10- to 100-fold more serum hAAT than mice injected with closed circular (cc) DNA for a period of 9 months (length of study). Furthermore, when a linear human factor IX expression cassette was delivered to factor IX-deficient mice, sustained serum concentrations of more than 4 microg/ml (80% of normal) of the human clotting factor and correction of the bleeding diathesis were obtained. Southern blot analyses indicate that, unlike ccDNA, LDNA rapidly formed large, unintegrated concatemers in vivo, suggesting that transgene persistence from plasmid-based vectors was influenced by the structure of the vector in transfected cells. No differences in transgene expression or DNA molecular structures were observed when AAV ITRs were included to flank the hAAT expression cassette in both ccDNA- and LDNA-treated animals. Linear DNA transfection provides an approach for achieving long-term expression of a transgene in vivo. PMID- 11273784 TI - Correction of deficient CD34+ cells from peripheral blood after mobilization in a patient with congenital erythropoietic porphyria. AB - Congenital erythropoietic porphyria (CEP) is an inherited disease due to a deficiency in the uroporphyrinogen III synthase (UROS), the fourth enzyme of the heme pathway. It is characterized by accumulation of uroporphyrin I in the bone marrow, peripheral blood, and other organs. The onset of most cases occurs in infancy and the main symptoms are cutaneous photosensitivity and hemolysis. For severe transfusion-dependent cases, when allogeneic cell transplantation cannot be performed, autografting of genetically modified primitive/stem cells is the only alternative. In the present study, efficient mobilization of peripheral blood primitive CD34(+) cells was performed on a young adult CEP patient. Retroviral transduction of this cell population with the therapeutic human UROS (hUS) gene resulted in both enzymatic and metabolic correction of CD34(+)-derived cells, as demonstrated by the increase in UROS activity and by a 53% drop in porphyrin accumulation. A 10-24% gene transfer efficiency was achieved in the most primitive cells, as demonstrated by the expression of enhanced green fluorescent protein (EGFP) in long-term culture-initiating cells (LTC-IC). Furthermore, gene expression remained stable during in vitro erythroid differentiation. Therefore, these results are promising for the future treatment of CEP patients by gene therapy. PMID- 11273786 TI - Pharmacokinetic population study to describe cefepime lung concentrations. AB - Pharmacokinetic parameters of cefepime in 2 g plasma and lung tissue bid over 3 days to achieve the steady-state was studied in 16 patients (15 male, one female) subjected to lung surgery for bronchial epithelioma. The aims of this study were firstly to quantify cefepime lung diffusion with cefepime lung concentrations in comparison with cefepime serum concentrations, and secondly to estimate population pharmacokinetic parameters of cefepime in lung tissue using NONMEM. The mean characteristics of patients were: age, 60 years (range, 51-69 years), weight, 73 kg (range, 62-87 kg) and creatinine clearance, 77 ml/min (range, 62-92 ml/min). Both serum sample (two per patient) and lung sample (one per patient) cefepime concentrations were analysed by HPLC with UV detection. Five groups were made according to the time of sampling after the last cefepime intravenous infusion at the fifth infusion: 0.5 h (n=2), 2 h (n=5), 4 h (n=3), 8 h (n=3) and 12 h (n=3). The cefepime concentration ratio between lung and serum was calculated for each group and statistical analysis show no significant difference between groups. The mean concentration ratio between lung and serum was 101% (range, 70-130%). To explain this observation a two-compartment pharmacokinetic model with a population approach was used to describe pharmacokinetic parameters of cefepime both in lung and in serum. Serum was assimilated at the central compartment and lung was the peripheral compartment. NONMEM was used to estimate the mean and the variance of the pharmacokinetic parameters. Central volume of distribution (V(d)), steady-state volume of distribution (V(ss)), central clearance (CL) and transfer constants (K(cp)) from serum to lung and (K(pc)) from lung to serum were estimated. Central elimination half-life t(1/2Kbeta)was extrapolated from elimination constant beta. Results were: V(d)= 15.62 +/- 2.56 l, V(ss)= 17.58 +/- 2.58 l, CL = 3.65 +/- 1.25 l/h, beta = 0.234 h(-1), t(1/2beta)= 2.96 hours, K(cp)= 12.25 +/- 8.56 h(-1)and K(pc)= 0.242 +/- 0.085 h( 1). The results show that cefepime diffusion in lung occurs quickly without lagtime and in similar concentrations to that in serum. PMID- 11273787 TI - Relationship of airway responsiveness with airway morphometry in normal and immunized rabbits. AB - Airway responses to chemical stimuli occur over a wide range of concentrations, with overlap between severe, moderate and mild asthmatic groups and with normal healthy individuals. Mathematical modelling has suggested that relative thickness of the airway wall may account for this range of responsiveness. We have investigated whether in vivo airway responsiveness varies as a function of airway wall thickness in terms of airway smooth muscle area in normal and immunized New Zealand White (NZW) rabbits. Airway responsiveness to inhaled methacholine (MCh) was determined in vivo under neuroleptanalgesia. Subsequently, ex vivo responsiveness to MCh (pD(2)=-log EC(50)) of isolated bronchi from the same animal was established. Smooth muscle area per mm basement membrane (SM/mmBM) was also measured morphometrically in the tested bronchi and the findings related to in vivo and ex vivo responsiveness. We found no relationship between airway responsiveness in vivo and pD(2)values in either immunized or control rabbits. In both control and immunized rabbits, no correlation was found between SM/mmBM and in vivo airway responsiveness. Only in immunized animals with a PCA titre >0, was there a significant correlation (=-0.5986, P<0.05) between SM/mmBM and pD(2). We conclude that airway smooth muscle area per se is not the sole contributor of airway responsiveness in vivo in normal rabbits. PMID- 11273788 TI - Comparison of in vitro surface properties of clove oil-phospholipid suspensions with those of ALEC, Exosurf and Survanta. AB - Dipalmitoyl phosphatidylcholine, the main component of lung surfactant is ineffective as a replacement surfactant due to its poor adsorption. We studied clove oil as a possible additive for improving the surface activity of protein free phospholipid suspensions. We added low doses of clove oil, to phospholipid suspensions and studied the surface properties by in vitro analysis using a pulsating bubble surfactometer and a Wilhelmy balance. Survanta, ALEC and Exosurf were used as controls for comparison. The test surfactants, which were phospholipid-oil suspensions at 1% concentration, in buffer containing either 2 or 5 mM calcium, were pulsated at 40 cpm in a pulsating bubble surfactometer. The phospholipids studied were dipalmitoyl phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), binary mixtures of PC:PE (2:3) and PC:PG (2:3). The addition of clove oil (CO) to each of the above phospholipids was in the ratio of nine parts of phospholipid to one part of oil. The presence of CO caused a significant improvement in the adsorption and minimum surface tension of all the phospholipid suspensions studied. The mixtures PC with CO, both in the presence of 2 and 5 mM calcium, and PCPE with CO at 2 mM calcium concentration had surface properties significantly better than those of ALEC and Exosurf and equivalent to those of Survanta. The addition of clove oil helps improve the surface properties of phospholipids. PMID- 11273790 TI - The effect of acute alteration in oxygen tension on the bronchodilator response to salbutamol in vitro and in vivo in man. AB - These studies examine the effect of acute hypoxia on airway smooth muscle relaxation in response to salbutamol in vitro in human isolated bronchi from non asthmatics and in vivo in-patients with asthma. Isometric responses were measured from rings of human bronchi pre-constricted with methacholine under oxygen tensions of 95% (hyperoxia), 20% (normoxia) and 4% (hypoxia). Once contractions had plateaued, concentration - response curves were conducted to salbutamol (10( 9)-10(-4)m). Twelve stable asthmatic patients were studied in a randomised double blind fashion. On two study days following baseline measurements, patients were randomised to receive either oxygen (FiO(2)1.0) or a hypoxic gas mixture (FiO(2)0.15) followed by three incremental doses of nebulised salbutamol at 15 min intervals. On two further study days nebulised saline was administered instead of salbutamol. In isolated bronchi, salbutamol-induced relaxations were significantly (P< 0.001) greater in hyperoxia and normoxia (P< 0.01) when compared to hypoxia. Among patients with asthma no significant differences were found in the mean maximum % change in forced expiratory volume (FEV(1)) from baseline between the hypoxic and hyperoxic study days on which nebulised salbutamol was administered. We conclude that acute hypoxia attenuates airway smooth muscle relaxation in response to salbutamol in vitro but has no effect on salbutamol-induced bronchodilation in in-patients with asthma. PMID- 11273789 TI - Effect of the cyclooxygenase-2 inhibitor celecoxib on bronchial responsiveness and cough reflex sensitivity in asthmatics. AB - Cyclooxygenase (COX), an essential enzyme in the pathway of prostaglandin formation from arachidonic acid, exists in two isoforms. Cyclooxygenase-1 (COX-1) is expressed under normal physiologic conditions, whereas COX-2, the inducible isoform, is associated with inflammation. Recent studies have linked COX-2 induction to the asthmatic inflammatory response, but potentially beneficial results, such as enhanced production of antiinflammatory and bronchoprotective substances, may also occur. The aim of the present study was to investigate the effect of selective COX-2 inhibition on bronchial responsiveness and cough reflex sensitivity. Eight adult subjects with stable asthma underwent spirometry, bronchoprovocation challenge with methacholine, and cough challenge testing with capsaicin, before and after a 7 day course of the COX-2 inhibitor celecoxib (200 mg orally, twice daily) and placebo, in a randomized, double-blind, crossover fashion. No significant changes in pulmonary function, bronchial responsiveness, or cough reflex sensitivity were induced by celecoxib. It appears, therefore, that 1 week of therapy with celecoxib does not significantly affect basal airway tone, nor the afferent airway receptors controlling bronchoconstriction and cough. However, the results of this trial cannot be extrapolated to subjects with severe asthma, or those suffering an asthmatic exacerbation. In such conditions of enhanced inflammatory response, the role of selective COX-2 inhibition remains to be elucidated. PMID- 11273791 TI - Secretory leukocyte protease inhibitor, but not alpha-1 protease inhibitor, blocks tryptase-induced bronchoconstriction. AB - Alpha-1-protease inhibitor (alpha(1)-PI) and secretory leukocyte protease inhibitor (SLPI) are two natural airway serine protease inhibitors. While inhibition of neutrophil elastase is a function common to both alpha(1)-PI and SLPI, we showed previously that they exhibit different patterns of protection against antigen-induced changes in airway function in allergic sheep. Specifically, the protective effect seen with SLPI was similar to the profile of action of synthetic tryptase inhibitors in the model. Based on these data, and the fact that tryptase is a serine protease, we hypothesized that SLPI, but not alpha(1)-PI, would block tryptase-induced bronchoconstriction. To test this, we compared the responses to inhaled tryptase in five sheep without treatment or after treatment with either aerosol alpha(1)-PI (10 mg) or aerosol SLPI (50 mg). The doses of alpha(1)-PI and SLPI selected had been shown to be effective in previous antigen-provocation studies. Treatments were given 30 min before aerosol challenge with tryptase (500 ng). Tryptase alone increased (mean+/-SEM) pulmonary resistance (R(L)) 142 +/- 24% over baseline. Pretreatment with alpha(1)-PI had no effect on the tryptase response (R(L)increased 122 +/- 20%). Pretreatment with SLPI, however, blocked the tryptase-induced response (R(L) increased only 40 +/- 4% P<0.05 vs. tryptase). These are the first studies comparing the inhibitory activity of SLPI and alpha(1)-PI on inhaled tryptase-induced bronchoconstriction. We conclude that, in vivo, SLPI, but not alpha(1)-PI, can block tryptase-induced bronchoconstriction and that this activity may explain the differential effects of these two serine protease inhibitors on antigen-induced airway responses in allergic sheep. PMID- 11273792 TI - Heparin inhibits allergen-induced eosinophil infiltration into guinea-pig lung via a mechanism unrelated to its anticoagulant activity. AB - There is considerable interest in the discovery of novel molecules for the treatment of allergic diseases and several recent studies have demonstrated that heparin can inhibit airway responses in subjects with asthma. However, heparin is also an anticoagulant which is potentially an unwanted effect in a molecule for treating asthma and allergic diseases. Recently, though, there have been a number of molecules described that are heparin-like but devoid of anticoagulant activity. The aim of this study was to evaluate whether the ability of heparin to inhibit allergen-induced eosinophil infiltration could be mimicked by analogues of heparin, some of which lack anticoagulant activity. We evaluated the effects of heparin and a number of modified heparins for their ability to inhibit allergen induced eosinophil infiltration into airways of suitably sensitised guinea-pigs assessed by bronchoalveolar lavage. Heparin and various modified heparins inhibited allergen-induced eosinophil infiltration into guinea-pig lung, including modified heparin preparations lacking anticoagulant activity. Our results suggest that heparin can inhibit eosinophil infiltration into lung tissue via a mechanism unrelated to its ability to act as an anticoagulant. Our results suggest that it may be possible to develop novel antiinflammatory agents for the treatment of asthma and allergic diseases related to the structure of heparin. PMID- 11273793 TI - Effect of tachykinins on airway function in cynomolgus monkeys. AB - Tachykinins have been implicated as important mediators of asthma. This study used neurokinin A (NKA) and substance P (SP) to evaluate the effect of tachykinins on airway mechanics in cynomolgus monkeys. NK(1)-(CP 99,994) and NK(2)-(SR 48968) receptor antagonists were used to evaluate the role of NK(1)and NK(2)receptors on responses to NKA and SP. Lung resistance (R(L)) and dynamic lung compliance (C(Dyn)) were measured in anesthetized, mechanically ventilated cynomolgus monkeys following aerosol or intravenous challenge with NKA, SP or the standard bronchoconstrictor, histamine. Inhaled NKA or SP had variable effects on R(L)and C(Dyn)whereas aerosolized histamine (0.01-1 mg/ml) dose-dependently increased R(L)and decreased C(Dyn). Intravenous NKA (1-100 microg/kg), SP (1-30 microg/kg) or histamine (1-100 microg/kg) increased R(L)and decreased C(Dyn). Pretreatment with SR 48968 (0.1 and 1 mg/kg, i.v.) blocked bronchoconstrictor responses to i.v. NKA, whereas CP 99,994 (0.1 and 1 mg/kg, i.v.) was without effect. Bronchoconstrictor responses to i.v. SP were partially blocked by SR 48968 and CP 99,994. In conclusion, both NKA and SP produce bronchonconstriction in cynomolgus monkeys and this effect is more pronounced when they are given by the iv route. Furthermore, both NK(1)and NK(2)receptors are involved in the bronchonconstrictor response to exogenously administered tachykinins in cynomolgus monkeys. PMID- 11273794 TI - Relative lung bioavailability of generic sodium cromoglycate inhalers used with and without a spacer device. AB - The relative lung bioavailability of sodium cromoglycate following inhalation has been evaluated using urinary drug excretion in nine healthly volunteers. Each inhaled four 5 mg sodium cromoglycate doses from a generic metered dose inhaler (MDI) and when it was attached to large volume spacer (MDI + VOL). A breath actuated MDI was also evaluated either used on its own (EB) or attached to a small volume spacer tube (EBO). The mean (SD) urinary excretion of sodium cromoglycate in the first 30 min post-inhalation was 34.1 (20.2), 211.7 (123.5), 29.3 (19.5) and 52.8 (36.0) microg following MDI, MDI+VOL, EB and EBO, respectively. The cumulative mean (SD) urinary excretion of sodium cromoglycate over the 24 h post-inhalation was 364.7 (266.2), 1227.1 (459.0), 280.2 (155.4) and 429.5 (176.7) microg. A metered dose inhaler attached to a large volume spacer delivers more sodium cromoglycate to the lungs than any other inhalation method. PMID- 11273795 TI - Bronchoconstrictor reactivity to NKA in allergic dogs: a comparison to histamine and methacholine. AB - Airway hyperresponsiveness to neurokinin A (NKA) occurs in inflammatory airway diseases like asthma. In this study, bronchoconstrictor reactivity to NKA was measured in beagle dogs neonatally sensitized to and challenged with ragweed. Comparisons were made to histamine and methacholine. Lung resistance (R(L)) and dynamic lung compliance (C(Dyn)) were measured in anesthetized, spontaneously breathing dogs before and after aerosol challenge with NKA, histamine or methacholine. The concentration of these agents increasing R(L)by 25% above baseline (PC(25)) was calculated before and 24 h after aerosolized ragweed challenge. Before ragweed, the bronchoconstrictor reactivity to NKA was four-fold higher in ragweed-sensitized dogs (PC(25)=0.036+/-0.006%) compared to non sensitized controls (PC(25)=0.177+/-0.030%, P<0.05). On the other hand, there was no difference in the bronchoconstrictor reactivity to histamine or methacholine between these two groups. Twenty-four hours after ragweed challenge to sensitized dogs, NKA reactivity was unchanged from pre-ragweed values but histamine and methacholine reactivity was increased by 2-3-fold. These results demonstrate airway hyperresponsiveness to NKA, histamine and methacholine in allergic beagle dogs although hyperresponsiveness to NKA exists in these allergic dogs before an antigen challenge. This animal model may prove to be useful to evaluate the role of tachykinins in hyperractive airway diseases. PMID- 11273796 TI - Effect of inhaled ciclesonide on airway responsiveness to inhaled AMP, the composition of induced sputum and exhaled nitric oxide in patients with mild asthma. AB - To assess the efficacy of ciclesonide, a novel corticosteroid pro-drug, we compared its effect on lung function, airway responsiveness to inhaled AMP, the composition of induced sputum, and the level of exhaled nitric oxide (NO) with the effect of budesonide in patients with asthma. Fifteen non-smoking steroid naive patients (mean FEV(1), 94%pred) inhaled either 400 microg ciclesonide or 400 microg budesonide as a single morning dose for two weeks each separated by a > or =3 week wash-out period. The study was performed in a double-observer, randomized, cross-over design. FEV(1)increased significantly during treatment with budesonide (3.38 vs. 3.64 l P=0,003), but not after ciclesonide (3.60 vs. 3.69 l). PC(20)FEV(1)of AMP increased (P<0,001, each) after both budesonide (4.59 vs. 32.48 mg/ml, 2.8 doubling doses) and ciclesonide (3.92 vs. 20.00 mg/ml, 2.4 doubling doses). The percentage of sputum eosinophils was significantly reduced after ciclesonide (7.9 vs. 3.4% P=0.01), but not budesonide (6.0 vs. 4.3%). After both budesonide and ciclesonide, a significant (P<0.001) reduction in the level of exhaled NO occurred. In none of the parameters studied, the changes differed significantly between treatment with budesonide or ciclesonide. These data suggest that ciclesonide is equi-effective to budesonide with regard to its potency to reduce the airway responsiveness to inhaled AMP as well as airway inflammation in patients with mild asthma. PMID- 11273797 TI - Antibiotic treatment and baseline severity of disease in acute exacerbations of chronic bronchitis: a re-evaluation of previously published data of a placebo controlled randomized study. AB - The study was designed to extend retrospectively the analysis of a previously reported study on chronic bronchitis patients with acute exacerbations treated with amoxicillin-clavulanic acid or matched placebo. We retrospectively re clustered patients on the basis of severity of baseline lung function: Cluster 1 (104 patients) mean screening FEV(1)32.67+/-6.83 (SD); Cluster 2 (109 patients) mean screening FEV(1)54.12+/-5.56; Cluster 3 (122 patients) mean screening FEV(1)71.54+/-5.51. The success rate in the antibiotic group was significantly greater compared to the placebo group (P<0.001). When clinical improvement was analysed on the basis of patient re-clustering, 31.4% of Cluster 1 (severe COPD) patients treated with amoxicillin/clavulanate showed clinical improvement, whereas success was recorded in 58.8%. Conversely, 13.2% of Cluster 1 patients receiving placebo improved and 17% successfully recovered (P<0.001). Mild and moderate COPD patients (Clusters 2 and 3) were grouped together. In these two groups, 31.2% and 53.6% of patients receiving antibiotic treatment showed improvement or recovery, respectively, compared to 29.2% improvements and 30.2% successful recoveries among placebo-treated patients (P<0.001). In placebo treated patients the improvement/success vs. failure rate was significantly different in Cluster 1 patients compared to Cluster 2+3 subjects (P<0.01, (2)test). The differences in final FEV(1)values in the treatment group and placebo group were significantly different (P<0.01) in favour of the active treatment group. Among more severe patients (Cluster 1), the comparison between screening and follow up FEV(1)values showed an improvement following antibiotic treatment and worsening after placebo (P<0.01). In Clusters 2 and 3 the difference between screening and follow up FEV(1)values was not significant for both treatment groups. Our patients with severe functional impairment and higher number of exacerbations per year are those who derive the greatest benefit from antibiotic treatment. PMID- 11273798 TI - Comparison of PDE 4 inhibitors, rolipram and SB 207499 (ariflo), in a rat model of pulmonary neutrophilia. AB - Using a rat model of lipopolysaccharide (LPS)-induced pulmonary inflammation, the antiinflammatory activity of SB 207499 was evaluated and compared to that of the prototypic type-4 phosphodiesterase (PDE4) inhibitor, rolipram. In dose-response experiments, we found that rats exposed to 10 microg or 100 microg of intratracheal (it) LPS developed a prominent pulmonary inflammation, due to a significant increase in the number of recoverable bronchoalveolar lavage neutrophils. The pulmonary neutrophilia, provoked by the challenge of 10 microg LPS/rat, was significant at 2 h, peaked by 16 h, declined thereafter but remained elevated for up to 48 h. Additionally, the exposure of rats to 10 microg LPS caused the local pulmonary production of TNF- alpha. In contrast to the cellular influx, TNF- alpha production peaked at 2 h and rapidly declined to negligible levels by 8 h. While low levels were detected, the levels of IL-1 beta in bronchoalveolar lavage did not significantly differ from saline challenged animals. Rats pretreated with rolipram or SB 207499, displayed dose-dependent inhibition of the LPS-induced pulmonary inflammation. Nevertheless, the pulmonary production of TNF- alpha and IL-1 beta was unaffected by either SB 207499 or rolipram. When provoked with the 10 microg dose of LPS, adrenalectomized rats produced a similar 24 h induction of pulmonary neutrophilia. Pretreatment of adrenalectomized rats with the PDE4 inhibitors showed similar inhibitory results to those obtained in normal rats. In summary, we have shown, using a rat model of LPS-induced pulmonary neutrophilic inflammation, that the inhibitory activities of rolipram or SB207499 are not linked to the production of TNF- alpha or the inhibition of IL-1 beta, and occur independently of endogenous catecholamine or corticosteroid release. PMID- 11273800 TI - Prey detection in selective plankton feeding by the paddlefish: is the electric sense sufficient? AB - The long rostrum of the paddlefish Polyodon spathula supports an extensive array of ampullary electroreceptors and has been proposed to function as an antenna for detecting planktonic prey. Evidence in support of this hypothesis is presented in experiments that preclude the use of other sensory mechanisms for plankton detection. Paddlefish swimming in a recirculating observation chamber are shown to feed normally in the dark when prey-related chemical and hydrodynamic sensory cues are masked or attenuated. Specifically, we demonstrate that the spatial distribution of plankton captured by paddlefish is little changed when the plankton are individually encapsulated in agarose, when a high background concentration of plankton extract is added to the chamber, when the nares are plugged and under turbulent water flow conditions. Paddlefish also discriminate between encapsulated plankton and 'empty' agarose particles of the same size. Although capture distributions differed somewhat under certain conditions, the general pattern and effectiveness of prey capture were not disrupted by these procedures. These results support the conclusion that paddlefish, as zooplanktivores, rely on their passive electric sense for prey detection. PMID- 11273801 TI - Paddlefish strike at artificial dipoles simulating the weak electric fields of planktonic prey. AB - The freshwater paddlefish Polyodon spathula (Polyodontidae) feeds primarily on the water flea (Daphnia sp.), and previous studies suggest that these fish detect their planktonic prey using their rostral electrosensory system. Zooplankton produce direct-current and oscillating alternating-current electric fields containing multiple frequencies and amplitudes. We asked whether an inanimate electric field is sufficient to elicit paddlefish strikes equivalent to their feeding behavior. Juvenile paddlefish respond to artificial dipole stimuli by investigating the electric field and striking at the dipole electrode tips. These behavioral responses, scored as strikes, exhibit a bandpass characteristic with a maximum response between 5 and 15 Hz. Responses were less frequent at higher (20, 30, 40, 50 Hz) and lower (0.1, 0.5, 1 Hz) test frequencies, with a steep drop-off below 5 Hz. Strike rates also varied with stimulus intensity. Response frequency was greatest at 0.25 microA peak-to-peak amplitude, with reduced responses at lower and higher amplitudes (0.125 and 1.25 microA). Striking behavior was also influenced by water conductivity: strike rate was reduced at higher water conductivity. Dipole-elicited strikes exhibit behavioral plasticity. Fish habituate to repetitive dipole stimuli that are not reinforced by prey capture, and they dishabituate after food reinforcement. These experiments characterize paddlefish feeding strikes towards dipole electrodes at signal frequencies and intensities simulating the electric fields of zooplankton, their natural prey, and demonstrate that electric fields are sufficient to elicit feeding behavior. The results support the conclusion that paddlefish use their passive electrosensory system for planktivorous feeding. PMID- 11273802 TI - Count and spark? The echo response of the weakly electric fish Gnathonemus petersii to series of pulses. AB - Weakly electric fish of the pulse type electrolocate objects in the dark by emitting discrete electric organ discharges (EODs) separated by intervals of silence. Two neighbouring pulse-type fish often reduce the risk of discharging simultaneously by means of an 'echo response': one fish will respond to a neighbour's EOD with a discharge of its own following at a fixed short latency so that its EOD will occur long before the next EOD of its neighbour. Although working elegantly for two partners, this simple strategy should fail in larger groups because two fish could discharge in response to the same EOD of a third fish. Here, I show that the mormyrid fish Gnathonemus petersii could use a simple mechanism to reduce this problem. Individuals were stimulated with two closely spaced pulses, the second following so as to coincide with an echo given in response to the first. All the fish examined were able to respond more to the second pulse so that most of their echoes did not collide with the second pulse. An analysis was made of how echoing more to the second pulse depends on (i) the delay at which the stimulus followed the last spontaneous EOD, (ii) the spontaneous firing rate, (iii) the intensity of the stimulus, (iv) the number of stimulus pulses, (v) the interval between stimulus pulses, and (vi) the level of previous stimulation with double pulses. The results suggest that echoing more in response to the second pulse is probably because the first pulse causes an after effect whose inferred properties would be compatible with the properties of the mormyromast afferences thought to be involved in the echo response. PMID- 11273803 TI - Why do cubomedusae have only four swim pacemakers? AB - The classic view of swimming control in scyphozoan and cubozoan jellyfish involves a diffuse motor nerve net activated by multiple pacemaker sites that interact in a simple resetting hierarchy. Earlier modeling studies of jellyfish swimming, utilizing resetting linkages of multiple pacemakers, indicated that increases in pacemaker number were correlated with increases in the rate and regularity of network activity. We conducted a similar study using the cubozoan jellyfish Carybdea marsupialis, concentrating not only on the adaptive features of multiple pacemaker networks but also on the mechanism of pacemaker interaction. The best fit for our experimental data is a model in which pacemakers express a degree of independence. Thus, our results challenge the idea that pacemaker interactions in scyphozoan and cubozoan medusae are based on a strict resetting hierarchy. Furthermore, our data suggest that the combination of semi-independent linkage of pacemakers with the small pacemaker number characteristic of cubomedusae is important in (i) maintaining a biphasic modulatory capability in the swimming system, and (ii) allowing behaviorally appropriate directional responses to asymmetrical sensory inputs in the radially arranged jellyfish nervous system. PMID- 11273804 TI - Scale effects on the attachment pads and friction forces in syrphid flies (Diptera, Syrphidae). AB - To test the role of constructional and dimensional factors in the generation of friction force by systems of setose attachment pads, six species of syrphid fly (Platycheirus angustatus, Sphaerophoria scripta, Episyrphus balteatus, Eristalis tenax, Myathropa florea and Volucella pellucens) were studied using light and scanning electron microscopy. Flies were selected according to their various body mass and attachment pad dimensions. Such variables as pad area, setal density, the area of a single setal tip and body mass were individually measured. A centrifugal force tester, equipped with a fibre-optic sensor, was used to measure the friction forces of the pads on a smooth horizontal surface made of polyvinylchloride. Friction force, which is the resistance force of the insect mass against the sum of centrifugal and tangential forces, was greater in heavier insects such as Er. tenax, M. florea and V. pellucens. Although lighter species generated lower frictional forces, the acceleration required to detach an insect was greater in smaller species. The area of attachment pads, setal tip area and setal density differed significantly in the species studied, and the dependence of these variables on body mass was significant. The frictional properties of the material of the setal tips were not dependent on the dimensions of the fly species. Similar results were obtained for the frictional properties of the pulvillus as a whole. Thus, the properties of the secretion and the mechanical properties of the material of the setal tips are approximately constant among the species studied. It is concluded that differences in friction force must be related mainly to variations in the real contact area generated by the pad on the smooth surface. The real contact area can be estimated as the summed area of the broadened setal tips of the pad in contact with the surface. The real contact area depends on such morphological variables as setal density and the area of a single setal tip. Although individual variables vary among flies with different dimensions, they usually compensate such that smaller setal tip area is partially compensated for by higher setal density. PMID- 11273805 TI - Copper transport by lobster hepatopancreatic epithelial cells separated by centrifugal elutriation: measurements with the fluorescent dye Phen Green. AB - The hepatopancreas of the American lobster (Homarus americanus) possesses four types of epithelial cells arranged along blind-ended tubules. At the distal tips of these tubules, stem cells termed E-cells differentiate into three other cell types, R-cells, F-cells and B-cells, each of which have different absorptive and secretory roles in the biology of the overall organ. This investigation uses centrifugal elutriation to separate the individual hepatopancreatic epithelial cell types of Homarus americanus and to investigate their plasma membrane copper transport properties using the copper-sensitive fluorescent dye Phen Green. Results show highly dissimilar endogenous concentrations of copper in each cell type and within the vacuoles (vesicles) released from these cells during the centrifugation process ([copper] in vacuoles>E-cells>R-cells>F-cells approximately B-cells). All four cell types were able to absorb copper from external concentrations ranging from 0.01 to 8 micromol l(-1), but considerable differences in transport rates occurred between the cell types. External calcium (0--10 mmol l(-1)) stimulated the uptake of external copper in a saturable fashion, suggesting the occurrence of carrier-mediated metal uptake. Addition of the Ca(2+) channel blocker verapamil (30 micromol l(-1)) to the external medium reduced the uptake rate of copper by all four cell types, but to different extents in each type of cell. External zinc (0--1000 nmol l(-1)) was a competitive inhibitor of copper influx in E- and R-cells, suggesting that the two metals shared the same binding and transport mechanism. A model is proposed which suggests that copper may enter all hepatopancreatic epithelial cell types by a divalent cation antiport process that exchanges intracellular Ca(2+) (or other cations) with either external copper or zinc. Verapamil-sensitive Ca(2+) channels may allow access of external calcium to cytoplasmic exchange sites on the antiporter or to activator sites on the same transport protein. The results suggest that elutriation is an excellent technique for the separation of complex invertebrate organ systems into their separate cell types and for analyzing the physiological properties of each cell type in isolation. PMID- 11273807 TI - Boxfishes (Teleostei: Ostraciidae) as a model system for fishes swimming with many fins: kinematics. AB - Swimming movements in boxfishes were much more complex and varied than classical descriptions indicated. At low to moderate rectilinear swimming speeds (<5 TL s( 1), where TL is total body length), they were entirely median- and paired-fin swimmers, apparently using their caudal fins for steering. The pectoral and median paired fins generate both the thrust needed for forward motion and the continuously varied, interacting forces required for the maintenance of rectilinearity. It was only at higher swimming speeds (above 5 TL s(-1)), when burst-and-coast swimming was used, that they became primarily body and caudal-fin swimmers. Despite their unwieldy appearance and often asynchronous fin beats, boxfish swam in a stable manner. Swimming boxfish used three gaits. Fin-beat asymmetry and a relatively non-linear swimming trajectory characterized the first gait (0--1 TL s(-1)). The beginning of the second gait (1--3 TL s(-1)) was characterized by varying fin-beat frequencies and amplitudes as well as synchrony in pectoral fin motions. The remainder of the second gait (3--5 TL s(-1)) was characterized by constant fin-beat amplitudes, varying fin-beat frequencies and increasing pectoral fin-beat asynchrony. The third gait (>5 TL s(-1)) was characterized by the use of a caudal burst-and-coast variant. Adduction was always faster than abduction in the pectoral fins. There were no measurable refractory periods between successive phases of the fin movement cycles. Dorsal and anal fin movements were synchronized at speeds greater than 2.5 TL s(-1), but were often out of phase with pectoral fin movements. PMID- 11273806 TI - Ultrastructural design of anuran muscles used for call production in relation to the thermal environment of a species. AB - I examined the aerobic trunk muscles, which are used for call production, of male frogs from species that breed in different thermal environments to test the hypothesis that cold-adapted frogs should have fewer capillaries per unit mitochondrial volume in oxidative muscles than warm-adapted frogs because of reduced mitochondrial function at low temperatures. The species of interest were the cold-temperate Pseudacris crucifer and the warm-tropical Hyla microcephala in the family Hylidae, and the cold-temperate Rana sylvatica and the warm-temperate Rana clamitans in the family Ranidae. Trunk-muscle mitochondrial volume, V(V)(mt,f), was proportionally higher in species with higher mean calling rates (number of notes per hour), irrespective of the familial affinity of a species and the thermal environment in which it vocalized. Trunk-muscle capillary length density, J(V)(c,f), was significantly lower in P. crucifer than in H. microcephala because of significantly higher mean fiber area, a macro (f). Conversely, trunk-muscle J(V)(c,f) was similar in the two ranid species. Using total capillary length, J(c), and total mitochondrial volume, V(mt,m), as a measure of maximal oxygen supply and demand, respectively, in trunk muscles, J(c) to-V(mt,m) ratios were significantly lower in cold-adapted P. crucifer (4.3 km cm(-3)) and R. sylvatica (4.8 km cm(-3)) than in warm-adapted H. microcephala (7.1 km cm(-3)) and R. clamitans (6.4 km cm(-3)). In contrast, J(c)-to-V(mt,m) ratios in the more anaerobic gastrocnemius muscle of these species was not related to the thermal environment of a species, which may reflect capillaries conforming to microcirculatory functions, e.g. lactate removal, that take precedence over oxygen delivery. Mitochondrial cristae surface area, S(V)(im,mt), in P. crucifer trunk and gastrocnemius muscles (37.7+/-1.6 and 35.9+/-1.5 m(2 )cm(-3) respectively) was, on average, similar to mammalian values, suggesting equivalent structural capacities of muscle mitochondria in these two taxa. Taken together, the present data suggest that trunk-muscle respiratory design may reflect a capillary supply commensurate with maximal levels of oxygen delivery set by mitochondria operating at different environmental temperatures. P. crucifer and H. microcephala trunk muscles were also characterized by a high lipid content, which contrasted with a near absence of trunk-muscle lipids in R. sylvatica and R. clamitans. The extraordinarily high lipid content of P. crucifer trunk muscles (26 % of muscle volume) may serve as an auxiliary oxygen pathway to mitochondria and thus compensate in part for this tissue's reduced capillary/fiber interface. The effect of potentially high depletion rates of trunk-muscle lipid stores on metabolic rates of male frogs while calling is discussed. PMID- 11273808 TI - Flexibility in flight behaviour of barn swallows (Hirundo rustica) and house martins (Delichon urbica) tested in a wind tunnel. AB - The flight behaviour of barn swallows (Hirundo rustica) and house martins (Delichon urbica) was tested in a wind tunnel at 15 combinations of flight angles and speeds. In contrast to that of most other small passerines, the intermittent flight of hirundines rarely consists of regular patterns of flapping and rest phases. To vary mechanical power output, both species used intermittent flight, controlling the number of single, pulse-like wingbeats per unit time. House martins in descent tended to concentrate their wingbeats into bursts and performed true gliding flight during rest phases. Barn swallows mainly performed partial bounds during brief interruptions of upstrokes, which they progressively prolonged with decreasing flight angle. Thus, identification of distinct flapping phases to calculate wingbeat frequencies was not feasible. Instead, an effective wingbeat frequency for flight intervals of 20 s, including partial bounds, was introduced. The effective wingbeat frequencies of house martins (N=3) ranged from 2 to 10.5 s(-1), those of barn swallows (N=4) from 2.5 to 8.5 s(-1). In both hirundine species, effective wingbeat frequency was found to decrease almost linearly with decreasing flight angle. With changes in air speed, wingbeat frequency varied according to a U-shaped curve, suggesting a minimum power speed of roughly 9 m s(-1). The duration of the down- and upstrokes varied systematically depending on flight angle and air speed. PMID- 11273809 TI - Could osmotaxis explain the ability of blue petrels to return to their burrows at night? AB - Like many other species of petrel, blue petrel (Halobaena caerulea) are able to return to their nest burrows at night in complete darkness. Since petrels have a well-developed olfactory system, we carried out an experiment to test whether blue petrels use olfaction to localise their nest burrows. Incubating birds were injected intranasally with a zinc sulphate solution, which reversibly impairs the sensitivity of the olfactory mucosa; control birds were treated with physiological saline solution. None of the anosmic birds returned to their burrows, whereas all the birds treated with saline solution did. Our results suggest that olfactory cues are necessary for blue petrels to find their burrows. PMID- 11273810 TI - Physiological and biochemical correlates of brood size and energy expenditure in tree swallows. AB - Intra-population variation in many fitness-related traits (e.g. clutch size) is often attributed to variation in individual parental quality. One possible component of quality is the level at which each individual can expend energy while provisioning dependent young. We used breeding tree swallows (Tachycineta bicolor) to test whether adults with large, natural-sized broods and/or nestlings in good nutritional condition had relatively high daily energy expenditures (DEEs). Adults with high DEEs were predicted to have large internal organs and high metabolic capacities. We first measured the growth rate of nestlings in natural broods of five, six and seven over a 4-day period and then measured parental DEE using doubly labelled water. Adults were then dissected for analyses of body composition and to determine maximum enzyme activities in the pectoral muscle. Although the total mass gain of large broods was greater than that of small broods, parental DEE was independent of brood size. We hypothesize that adults matched their clutch size (and consequently, brood size) to their individual foraging efficiencies. When statistically controlling for the effects of brood size, in one of two years there was a positive correlation between DEE and brood mass. This suggests that among individuals rearing the same-sized broods there were reproductive benefits of a relatively high DEE. There was no correlation between either brood size or DEE and the mass of any internal organ or the metabolic capacity of the pectoral muscle. PMID- 11273811 TI - High-frequency initial pulses do not affect efficiency in rat fast skeletal muscle. AB - This study investigated the effects of high-frequency initial pulses on the efficiency (=total work output/high-energy phosphate consumption) of rat fast skeletal muscle. In situ rat medial gastrocnemius muscles performed 15 repeated shortening contractions (2 s(-1); velocity 50 mm s(-1)) with occluded blood flow while activated with triplets of 400 Hz followed by 60 Hz trains (T400;60) or with constant-frequency trains of either 60 or 91 Hz. All stimulation patterns consisted of six pulses. After the last contraction, the muscles were quickly freeze-clamped and analysed for metabolite levels. The calculated efficiencies were 20.4+/-3.0 mJ micromol(-1 )P (N=7), 19.4+/-1.8 mJ micromol(-1 )P (N=8) and 19.6+/-2.5 mJ micromol(-1 )P (N=7; means +/- s.d.) for T400;60, 60 and 91 Hz stimulation respectively (P>0.05). It is concluded that, although high-frequency initial pulses can enhance muscle performance, the efficiency of rat fast skeletal muscle did not differ from that for submaximal constant-frequency stimulation patterns. PMID- 11273812 TI - Regulation of Na(+) transport across leech skin by peptide hormones and neurotransmitters. AB - An increase in intracellular cyclic AMP concentration stimulates transepithelial Na(+) transport across the skin of the leech Hirudo medicinalis, but it is unclear how cytosolic cyclic AMP levels are elevated in vivo. In search of this external stimulus, we performed Ussing chamber experiments to test several peptide hormones and neurotransmitters for their effect on Na(+) transport across leech dorsal integument. Although all the peptide hormones under investigation significantly affected ion transport across leech integument, none of them mimicked the effect of an experimental rise in intracellular cyclic AMP level. The invertebrate peptides conopressin and angiotensin II amide inhibited short circuit-current- (I(sc)) and amiloride-sensitive Na(+) transport (I(amil)), although to slightly different degrees. The vertebrate peptide hormones 8 arginine-vasopressin and 8-lysine-vasopressin both produced an inhibition of I(amil) comparable with that caused by angiotensin II amide. However, 8-lysine vasopressin reduced I(sc), whereas 8-arginine-vasopressin induced a moderate increase in I(sc). The neurotransmitter dopamine, which occurs in the leech central nervous system in relatively large amounts, and its precursor l-dopamine both induced large decreases in I(sc) and I(amil). However, the reactions evoked by the catecholamines showed no pronounced similarity to the effects of intracellular cyclic AMP. Two other neurotransmitters known to occur in leeches, serotonin (5-hydroxytryptamine) and gamma-n-aminobutyric acid (GABA), had no influence on transepithelial ion transport in leech skin. PMID- 11273814 TI - [Pediatric lung transplantation: a reality]. PMID- 11273813 TI - Branchial CO(2) receptors and cardiorespiratory adjustments during hypercarbia in Pacific spiny dogfish (Squalus acanthias). AB - Adult Pacific spiny dogfish (Squalus acanthias) were exposed to acute (approximately 20 min) hypercarbia while we monitored arterial blood pressure, systemic vascular resistance (R(S)), cardiac output (V(b)) and frequency (fh) as well as ventilatory amplitude (V(AMP)) and frequency (f(V)). Separate series of experiments were conducted on control, atropinized (100 nmol kg(-1)) and branchially denervated fish to investigate putative CO(2)-chemoreceptive sites on the gills and their link to the autonomic nervous system and cardiorespiratory reflexes.In untreated fish, moderate hypercarbia (water CO(2 )partial pressure; Pw(CO2)=6.4+/-0.1 mmHg) (1 mmHg=0.133 kPa) elicited significant increases in V(AMP) (of approximately 92 %) and f(V) (of approximately 18 %) as well as decreases in fh (of approximately 64 %), V.(b) (approximately 29 %) and arterial blood pressure (of approximately 11 %); R(S) did not change significantly. Denervation of the branchial branches of cranial nerves IX and X to the pseudobranch and each gill arch eliminated all cardiorespiratory responses to hypercarbia. Prior administration of the muscarinic receptor antagonist atropine also abolished the hypercarbia-induced ventilatory responses and virtually eliminated all CO(2)-elicited cardiovascular adjustments. Although the atropinized dogfish displayed a hypercarbic bradycardia, the magnitude of the response was significantly attenuated (36+/-6 % decrease in fh in controls versus 9+/-2 % decrease in atropinized fish; means +/- s.e.m.).Thus, the results of the present study reveal the presence of gill CO(2) chemoreceptors in dogfish that are linked to numerous cardiorespiratory reflexes. In addition, because all cardiorespiratory responses to hypercarbia were abolished or attenuated by atropine, the CO(2) chemoreception process and/or one or more downstream elements probably involve cholinergic (muscarinic) neurotransmission. PMID- 11273815 TI - [Effectiveness of a clinical test in the preselection of children with suspected fragile X syndrome]. AB - BACKGROUND: Fragile X syndrome (FXS) is the most frequent hereditary cause of mental retardation. It can be diagnosed by molecular genetic techniques, but clinical suspicion is made less likely by it variable expression. OBJECTIVE: To assess the effectiveness of a six-item checklist in the preselection of children who are candidates for FXS genetic study. MATERIAL AND METHODS: We studied 70 male patients aged between 2 and 10 years with mental retardation of unknown cause. In all patients a checklist with six clinical criteria (mental retardation, history of familial mental retardation, long face, large ears, autistic-like behaviour, and attention deficit disorder with hyperactivity) measured from 0-2 points was applied and molecular genetic studies using polymerase chain reaction and Southern-blot were performed. RESULTS: In 14 of the 70 children (20%) molecular study confirmed full mutation (200 CGG repeats). A score of six points in the test had the greatest discriminatory power and was reached by 14 patients (100%) with mutation, but only by 2of 56patients (3.5%) without mutation. The most accurate diagnostic model was the association of mental retardation, attention deficit disorder with hyperactivity, large ears and a history of familial mental retardation followed by long face and autistic-like behaviour. CONCLUSIONS: The six-item checklist improved the preselection of children with suspicion of FXS, which was later confirmed by molecular genetic techniques. PMID- 11273816 TI - [Molecular diagnosis of fragile X syndrome with polymerase chain reaction: application of a diagnostic protocol in 50 families from northern Spain]. AB - OBJECTIVES: The aim of this study was to develop a rapid, non-radioactive and effective method for the molecular diagnosis of fragile X syndrome (FXS) by the polymerase chain reaction (PCR) of the CGG repeat and to establish a protocol to be used in: a)ruling out FXS in patients with non-specific mental retardation; b)determining the exact genotype of affected individuals; c)studying all at-risk individuals from families with FXS and identifying asymptomatic carriers, and d)offering accurate genetic and reproductive counselling to families with FXS. MATERIALS AND METHODS: Samples from 438 individuals from 50 families with FXS were studied using three different PCR tests: the first to detect ethidium bromide through ultraviolet light, the second to detect digoxigenin and CSPD after blotting and hybridisation with the (CGG)5 oligoprobe, and the third to amplify and detect the DXS548 microsatellite. RESULTS: Of the 438 individuals studied, 121 had full mutations (60 males and 61 females), 86 had pre-mutations (7 males and 79 females), 16 showed mosaic patterns and 215 had no mutations. PCR techniques amplified up to 120-150 repeats, and direct study with probes was required when no bands or only one band was detected in females. PCR was more accurate than genomic DNA Southern blot analysis in pre-mutated carriers. In one family, recombination between the FRAXA locus and the DXS548 microsatellite was found. CONCLUSIONS: These non-radioactive PCR protocols permit rapid and accurate diagnosis of FXS. They and are especially useful in prenatal diagnosis and in the identification of carriers. PMID- 11273817 TI - [Idiopathic childhood occipital epilepsy]. AB - OBJECTIVE: To describe the clinical and electroencephalographic (EEG) features, as well as the outcome of children diagnosed with idiopathic childhood occipital epilepsy (COE) in our hospital. METHODS: A retrospective review of the clinical records of children diagnosed with COE in the previous 10years was carried out with description of clinical and EEG features and neuroimaging studies. The outcome of patients followed-up for at least 5 years was also reviewed. RESULTS: Ten children were studied: two with type I (Gastaut) COE, six with type II (Panayiotopoulos) COE, and two with intermediate forms of the disorder. Patients with type I COE suffered daytime seizures with visual symptoms (hallucinations and amaurosis) followed by versive motor partial complex seizures with secondary generalized seizures. Age of onset was late childhood and the seizures reappeared in adolescence when therapy was discontinued. Patients with type II COE had nocturnal seizures consisting of tonic deviation of the head and eyes, some degree of disturbance of consciousness and hypotony followed by vomiting and hemiclonic movements or generalized tonic-clonic seizures. In five children, the first presenting symptom was status epilepticus. In all patients the age of onset was between 1 and 4 years. The results of neuroimaging studies were normal. EEG records showed normal baseline activity with slow wave spikes in the occipital region that disappeared or were reduced by eye opening. CONCLUSIONS: Our clinical and EEG findings are similar to those of other published studies. Type II COE frequently presents as status epilepticus and can be confused with other neurologic emergencies. Ictal EEG is useful to clarify the diagnosis. In type II COE, ictal symptomatology may overlap with migraine with aura. Although designated benign, patients with type I COE may develop learning problems and continue to have seizures throughout childhood. PMID- 11273818 TI - [Mid-term results of percutaneous closure of atrial septal defect in children]. AB - OBJECTIVE: To evaluate the medium-term results of percutaneous closure of atrial septal defect. METHODS: Twenty-two children (mean weight, 23 11kg; mean age, 5.7 2.4 years) underwent percutaneous atrial septal defect closure under general anesthesia. The procedure was monitored by transesophageal echocardiography. DAS Angel Wings (n4) and the Amplatzer device (n18) were used. RESULTS: Mean pulmonary artery pressure was 13 2.8mmHg, mean pulmonary vascular resistance was 1.50.5U/m2 and mean Qp/Qs flow ratio was 2.2 0.6. The mean diameter of the defects was 14.5 6.3mm by transesophageal echocardiography OmniPlane measurement and 15.95.3mm using balloon occlusion reference. A total of 31 devices were used: 4 Angel Wings and 27Amplatzer devices. Twelve Amplatzer devices were withdrawn through the introducer without complications, 5 due to a discrepancy in the size of the left auricle, 4 because they were too small to stabilize in the septum and 3 due to defective opening in the left auricle. In 19 patients implantation was successful. In 17 patients transthoracic color Doppler echocardiography carried out 24 hours after the procedure showed a minimal shunt which was no longer present 1 month later. The mean time of discharge was 38 12 hours after the procedure. After a mean follow up 15 6 months the patients remain asymptomatic with no clinical or technical problems. CONCLUSION: The success rate of percutaneous closure of atrial septal defects in well-selected patients was high and presented no complications. PMID- 11273819 TI - [Radiofrequency catheter ablation in children with Wolff-Parkinson-White syndrome and sudden cardiac death who had been resuscitated]. AB - INTRODUCTION: Sudden death may be the first manifestation of the Wolff-Parkinson White syndrome, especially in children and adolescents. OBJECTIVES: The aim of this study was to evaluate the usefulness of radiofrequency catheter ablation in children with Wolff-Parkinson-White syndrome with aborted sudden death. METHODS AND RESULTS: We report four patients with Wolff-Parkinson-White syndrome who survived cardiac arrest. The patients were aged from 2.5 months to 16 years. The two first patients were lactating infants; in the first sudden death occurred during digoxin treatment for supraventricular tachycardia secondary to Wolff Parkinson-White syndrome and in the second the syndrome was diagnosed after an episode of sudden death. In these patients a free wall accessory pathway (left posterior and left lateral, respectively) was successfully ablated using a transseptal approach. The third patient was diagnosed with asymptomatic Wolff Parkinson-White syndrome; sudden death occurred during exercise. In the fourth patient, sudden death occurred after intravenous therapy with adenosine triphosphate and amiodarone for rapid atrial fibrillation. In both patients, one accessory pathway, located in right posteroseptal and right anterior free wall, respectively, was ablated. After a mean follow-up of 43.5 26.4 months, no recurrence of sudden death had occurred and electrocardiogram showed sinus rhythm without delta wave. The third patient presented severe sequelae of hypoxemic encephalopathy, which persisted during the follow-up. CONCLUSIONS: Radiofrequency catheter ablation is the treatment of choice in Wolff-Parkinson-White syndrome with episodes of aborted sudden death. PMID- 11273820 TI - [Pediatric lung transplantation]. AB - Approximately 700 transplants world-wide have been performed in patients aged less than 18 years; in contrast, over 11,000 lung transplants have been performed in adults. The major diagnostic group is cystic fibrosis. An emerging group of patients are infants born with congenitally based pulmonary diseases such as surfactant protein B deficiency. Survival in children is very similar to that in adults, although it is generally perceived that children are at higher risk. For instance, no children have been transplanted for chronic obstructive lung disease (a low-risk diagnostic group) while this disease comprises approximately 40% of all adult lung transplantations. Immunosuppression generally consists of cyclosporine, azathioprine and prednisone. Post-transplant complications in children are similar to those observed in adults. Around 40% of patients will develop bronchiolitis obliterans approximately 3 years after transplantation and this remains the major impediment to long term survival. Donor shortage is also a significant obstacle, especially in adolescents. As a partial solution to this problem, living donor lung transplantation has taken on a greater role in our program. Although this is a complex, expensive treatment strategy, lung transplantation remains the most effective therapy for end-stage pulmonary parenchymal and vascular diseases even in children. PMID- 11273821 TI - [Recommendations on the use of formulas for the treatment and prevention of adverse reactions to cow milk proteins]. PMID- 11273822 TI - [Assessment of the quality of scientific evidence in Anales Espanoles de Pediatria]. AB - Evidence-based medicine is a new scientific paradigm that aims to use medical literature more effectively in guiding medical practice. The aim of this study was to assess the quality of scientific evidence in Spanish pediatric articles. Original articles published in Anales Espanoles de Pediatria during a 6.5year period (n733) were compared with those published in Pediatrics during a 1.5year period (n300). The quality of scientific evidence in Anales was high in only 3% of original articles (randomized clinical trials). It was average in 30.4% (non randomized clinical trials, cohort studies, case-control studies) and poor in 66.6% (descriptive studies, case reports, etc.). Only 10.2% of articles used appropriate methodological concepts according to evidence-based medicine. These concepts were mainly "soft" (odds ratio, relative risk, confidence interval) and no "hard" concepts (number needed to treat, likelihood ratio, odds pretest) were detected. The pediatric specialty showing the highest quality of scientific evidence, greatest use of appropriate methodological concepts and greatest statistical accessibility was pneumology. The first step in improving the quality of scientific evidence would be to establish collaboration between epidemiologists and/or biostatisticians. The evidence-based bibliometric indicators found in Pediatrics serve as a gold standard for Anales. PMID- 11273823 TI - [Pneumococci: a new microorganism in the newborn?]. AB - OBJECTIVE: To study the clinical characteristics and susceptibility to antimicrobial agents of Streptococcus pneumoniae invasive infection in our neonatal unit. METHODS: Data from newborns with Streptococcus pneumoniae invasive infection in the last 12 years were retrospectively collected. RESULTS: Eight cases of invasive infection were identified. Gestational age ranged from 30 to 38 weeks (median: 34 weeks) and birth weight ranged from 1,680g to 4,460g (median: 2,480g). Risk factors related to infection were identified in 7 patients. Although infection manifested as shock in 4 patients and meningitis in 1, evolution was favorable in all patients. Penicillin resistance was found in 3 patients. CONCLUSIONS: Streptococcus pneumoniae produces serious disease in neonates. Because of the increasing prevalence of penicillin-resistant pneumococci, the relationship between the percentage of mothers colonized with pneumococci and neonatal infection should be determined to develop new prevention and treatment strategies in newborn infants. PMID- 11273824 TI - [Transitory neonatal diabetes]. AB - Transitory neonatal diabetes mellitus is a rare carbohydrate metabolism disorder that usually occurs between the ages of 2 days and 6 months. We report the case of an asymptomatic newborn treated with NPH insulin, in whom genetic study revealed an alteration associated with neonatal diabetes. The patient was a low birth weight infant born after 37 weeks' gestation to a previously childless mother with gestational diabetes controlled by diet. There were familial antecedents of diabetes. Physical examination revealed only syndactylia of the second and third toes. Asymptomatic hyperglycemia higher than 200mg/dl was detected on the second day of life. Treatment with regular subcutaneous insulin was started on the fourth day of life with irregular response. On the forty-first day of life treatment with NPH insulin was started with better response, permitting the reduction of regular insulin until its suppression 15 days later. Treatment with NPH insulin was stopped when the patient was 9 months old. During this time concentrations of insulin, cortisone, peptide C, insulin antibodies, anti-TPO, anti-TG, anti-GAD, anti-tyrosine-phosphatase and glycosylate hemoglobin were normal. Abdominal echography showed no abnormalities. Karyotype: 46 XX, der(6)dup(q22-q23) (long arm duplication of chromosome number 6).In conclusion, NPH insulin could provide an alternative to regular insulin in the treatment of transitory neonatal diabetes mellitus. Its association with genetic alterations could alter prognosis. PMID- 11273825 TI - [Double total anomalous pulmonary venous connection]. AB - We present a newborn infant with right atrial isomerism, complex congenital heart malformation and anomalous pulmonary venous connection, reliably diagnosed by 2-D Doppler color echocardiography. The infant had no significant obstetric antecedents. The neonatal clinical picture included cyanosis, heart murmur and respiratory distress. The infant was treated with prostaglandin from the age of 24 hours until his death after surgery. The 2-D echo Doppler color flow mapping showed findings that suggested right atrial isomerism, severe left ventricular hypoplasia, pulmonary atresia and ductus arteriosus. The pulmonary veins flowed together to a posterior cardiac chamber from which an emissary vertical venous vessel connected with a left superior aneurysmal sack. Two venous channels emerged from this sack: one connecting to the innominate vein and the other to the atrium. The malformations were confirmed by cardiac catheterization. On the sixth day of life, the patient underwent anastomosis between the posterior venous chamber with the atrium, a modified Blalock-Taussig shunt implant, and ductus closure but died during surgery. The association between complex cardiac anomalies and uncommon obstructive total anomalous pulmonary venous connection in the context of right atrial isomerism is lethal and few neonates survive surgical repair. Two-dimensional echo color flow Doppler is a reliable diagnostic technique and an indispensable guide in angiography. PMID- 11273826 TI - [Blade atrial septostomy in an infant with hypoplastic left heart syndrome awaiting heart transplantation]. AB - Blade atrial septostomy was performed in a 36 day-old infant diagnosed with hypoplastic left heart syndrome and restrictive atrial septal defect. The patient was awaiting cardiac transplantation. The defect was enlarged using a 9.4-cm blade. The mean atrial gradient was reduced from 19 to 2mmHg. After septostomy the patient showed marked clinical improvement and aortic oxygen saturation increased from 68% to 88% with no complications. Twenty-five days later orthotopic heart transplantation was successfully performed. PMID- 11273827 TI - [Kwashiorkor as a symptom of abuse and neglect in Barcelona]. AB - Child abuse can take various forms and the presenting signs and symptoms can sometimes lead to diagnostic error. Faced with a malnourished child physicians should not forget that one of the possible causes of the malnutrition could be abuse. Because of rising immigration, forms of child abuse uncommon among the Spanish industrialized population may increase.An African girl, who had been resident in Spain for several years, presented with symptoms of kwashiorkor disease. This, combined with observation of the child's social environment and her own account, led to a diagnosis of neglect. This is a rare clinical presentation of abuse in our socio-economic milieu. PMID- 11273828 TI - [Multiple vertebral defect with associated anomalies]. PMID- 11273829 TI - [Ring chromosome 13 and congenital coagulation factor deficiency]. PMID- 11273830 TI - [Further thoughts on pure oxygen]. PMID- 11273832 TI - [Diagnostic codification of cardiopulmonary arrest in emergencies in pediatrics]. PMID- 11273833 TI - [Diagnostic codification of syncope in emergencies in pediatrics]. PMID- 11273834 TI - [Clarifications on diagnostic codification in emergencies in pediatrics]. PMID- 11273838 TI - Got soy? PMID- 11273839 TI - The APOE gene and diets--food (and drink) for thought. PMID- 11273840 TI - Fatty acids and early human growth. PMID- 11273841 TI - Dietary modulation of endothelial function: implications for cardiovascular disease. AB - The vascular endothelium is the primary site of dysfunction in many diseases, particularly cardiovascular disease. A variety of risk factors, including smoking, hypercholesterolemia, hyperhomocysteinemia, hypertension, and diabetes mellitus, adversely affect endothelial function. Emerging evidence suggests an important role of dietary factors in modulating endothelial function. In particular, n-3 fatty acids, antioxidant vitamins (especially vitamins E and C), folic acid, and L-arginine appear to have beneficial effects on vascular endothelial function, either by decreasing endothelial activation or by improving endothelium-dependent vasodilation in patients at high risk of cardiovascular disease as well as in healthy subjects. These effects may serve as one potential mechanism through which these nutrients reduce the risk of cardiovascular disease, as observed in epidemiologic studies and several clinical trials. This article reviews clinical and experimental evidence regarding the role of these nutrients in modulating endothelial function and their potential to prevent cardiovascular disease. PMID- 11273842 TI - Evaluation of body fat in fatter and leaner 10-y-old African American and white children: the Baton Rouge Children's Study. AB - BACKGROUND: Only a few published studies in children used several methods to compare body fat in large groups of fatter and leaner multiethnic children. We hypothesized that the preferred methods of determining body fat may differ in children with larger compared with smaller amounts of body fat, in boys compared with girls, and in African Americans compared with whites. OBJECTIVE: Our objective was to evaluate several methods of predicting body fat in 10-12-y-old white and African American boys and girls. DESIGN: The body fat of 129 African American and white boys and girls aged 10-12 y, distributed equally by sex and race, was measured with use of dual-energy X-ray absorptiometry (DXA), underwater weighing (densitometry), isotope dilution (H(2)18O), bioelectrical impedance, skinfold thicknesses, corporal diameters, and circumferences. RESULTS: With use of DXA as the criterion variable, body fat was bimodally distributed in the boys and skewed to higher values in the girls. Biceps skinfold thickness had the highest predictive value of any single skinfold thickness compared with DXA fat. All formulas for estimating body fat from skinfold thicknesses, body density, or impedance performed better in the children in the upper one-half of the fat distribution (the fatter children) than in those in the lower one-half (the leaner children). Body mass index was highly correlated with body fat (R2 = 0.77); there was a good correlation for the fatter children (R2 = 0.66) and no correlation for the leaner children (R2 = 0.09). The hydration of the fat-free mass was significantly higher in the fatter children than in the leaner ones (79.2% compared with 76.7%). CONCLUSIONS: These data are consistent with the hypothesis that all methods of estimating body fat work better in children with larger amounts of body fat. The best formulas use skinfold thicknesses, bioelectrical impedance, and a 4-compartment model. PMID- 11273843 TI - Occasional physical inactivity combined with a high-fat diet may be important in the development and maintenance of obesity in human subjects. AB - BACKGROUND: A better understanding of the environmental factors that contribute to obesity is imperative if any therapeutic effect on the increasing prevalence of overweight and obesity in the United States is to be achieved. OBJECTIVE: This study examined the effect of the interaction of diet composition and physical inactivity on energy and fat balances. DESIGN: Thirty-five normal-weight and obese subjects were randomly assigned to either a 15-d isoenergetic high carbohydrate (HC) or high-fat (HF) diet according to a crossover design. During the first 14 d, body weight and physical activity were maintained. On day 15, subjects spent 23 h in a whole-room indirect calorimeter and were fed a diet similar to that consumed during the previous 7 d while remaining physically inactive. RESULTS: Energy intakes required to maintain body weight stability during the first 14 d were similar between diets. Normal-weight and obese subjects consuming both diets had a positive energy balance on the sedentary day (day 15), suggesting that subjects were less active in the calorimeter. There was no significant effect of diet composition on total energy balance and total protein-energy balance on day 15; however, carbohydrate balance was more positive with the HC (2497.8 +/- 301.2 kJ) than with the HF (1159 +/- 301.2 kJ) diet (P = 0.0032). Most importantly, fat balance was more positive with the HF (1790.8 +/- 510.4 kJ) than with the HC (-62.8 +/- 510.4 kJ) diet (P = 0.0011). CONCLUSION: Chronic consumption of a high-carbohydrate diet could provide some protection against body fat accumulation in persons with a pattern of physical activity that includes frequent sedentary days. PMID- 11273844 TI - Beta-oxidation of linoleate in obese men undergoing weight loss. AB - BACKGROUND: In animals, the whole-body content and accumulation of linoleate can be measured and compared with its intake to determine linoleate beta-oxidation. This method can also provide quantitative information about the beta-oxidation of linoleate in humans. OBJECTIVES: The objectives of the study were to 1) use the wholebody fatty acid balance method to quantify whole-body concentrations of linoleate in humans, 2) estimate the distribution of linoleate between adipose and lean tissue, and 3) assess the effect of weight loss on linoleate stores and beta-oxidation in obese humans. DESIGN: Nine healthy obese men underwent supervised weight loss for 112 d (16 wk). Magnetic resonance imaging data and fatty acid profiles from fat biopsies were both used to determine linoleate stores in adipose and lean tissue and in the whole body. Linoleate beta-oxidation was calculated as intake - (accumulation + excretion). RESULTS: Mean weight loss was 13 kg and linoleate intake was 24 +/- 6 mmol/d over the study period. Whole body loss of linoleate was 37 +/- 18 mmol/d, or 28% of the level before weight loss. Combining the intake and whole-body loss of linoleate resulted in linoleate beta-oxidation exceeding intake by 2.5-fold during the weight-loss period. CONCLUSIONS: All dietary linoleate is beta-oxidized and at least an equivalent amount of linoleate is lost from the body during moderate weight loss in obese men. The method studied permits the assessment of long-term changes in linoleate homeostasis in obese humans and may be useful in determining the risk of linoleate deficiency in other conditions. PMID- 11273846 TI - Moderate dietary fat consumption as a risk factor for ischemic heart disease in a population with a low fat intake: a case-control study in Korean men. AB - BACKGROUND: Dietary fat intake is associated with the incidence of ischemic heart disease (IHD) in Western countries. In populations in which both the average dietary fat consumption and the incidence of IHD are lower than in Western countries, the association of dietary fat intake with IHD incidence remains unknown. OBJECTIVE: We conducted a case-control study to examine the association of dietary fat with IHD incidence in Korean men. DESIGN: The case group consisted of 108 patients with electrocardiogram-confirmed myocardial infarction or angiographically confirmed (> or =50% stenosis) IHD who were admitted to a university teaching hospital in Seoul, Republic of Korea. The controls were 142 age-matched patients admitted to the departments of ophthalmology and orthopedic surgery at the same hospital. Dietary fat intake was assessed by a nutritionist using a semiquantitative food-frequency questionnaire. Body mass index (BMI), cigarette use, alcohol intake, exercise, and history of disease were determined during an interview and examination. RESULTS: In a univariate analysis, the mean percentages of energy from total fat, saturated fatty acids, and monounsaturated fatty acids were significantly higher in the cases than in the controls. BMI, smoking, and a history of hypertension were associated with the occurrence of IHD. In multiple logistic analyses, total fat intake was a significant risk factor (odds ratio: 1.08 for 1% of energy intake; 95% CI: 1.02, 1.14) after adjustment for BMI and smoking. CONCLUSION: In a population with a relatively low fat intake (19% of energy intake), a moderate increase in total fat intake may be a risk factor for IHD. PMID- 11273845 TI - Influence of a stearic acid-rich structured triacylglycerol on postprandial lipemia, factor VII concentrations, and fibrinolytic activity in healthy subjects. AB - BACKGROUND: An elevated postprandial lipid concentration is believed to be atherogenic and to increase the risk of thrombosis. OBJECTIVE: The objective was to test whether the consumption of a stearic acid-rich structured triacylglycerol has adverse effects on postprandial fibrinolytic activity and lipemia, factor VII coagulant (FVII:c) activity, and activated FVII (FVIIa) concentrations. DESIGN: A randomized crossover design was used to compare the effects on middle-aged healthy men (n = 17) and women (n = 18) of meals enriched with cocoa butter, high oleate sunflower oil (oleate), or a structured triacylglycerol containing stearic acid. RESULTS: The mean increases from fasting in plasma triacylglycerol 3 h after the oleate, cocoa butter, and structured triacylglycerol meals were 1.36 (95% CI: 1.17, 1.56), 1.39 (1.17,1.63), and 0.65 (0.50, 0.82) mmol/L, respectively. Tissue plasminogen activator activity increased and plasminogen activator type 1 activity decreased after all 3 meals. Plasma FVII:c increased after the oleate and cocoa butter meals but not after the structured triacylglycerol meal. The values 6 h after the oleate and cocoa butter meals were 11.3% (7.0%, 15.6%) and 9.9% (4.7%, 15.2%), respectively, and were significantly different (P < 0.0001 and P = 0.001, respectively) from the value after the triacylglycerol meal [2.1% (-1.1%, 5.3%)]. Plasma FVIIa increased after all 3 meals, more so after the oleate and cocoa butter meals than after the structured triacylglycerol meal. CONCLUSION: The consumption of stearic acid in the form of a structured triacylglycerol leads to less of an increase in plasma triacylglycerol and in FVII:c than does a meal enriched in cocoa butter or oleate. PMID- 11273847 TI - The effect of soy protein with or without isoflavones relative to milk protein on plasma lipids in hypercholesterolemic postmenopausal women. AB - BACKGROUND: Clinical trial data and the results of a meta-analysis suggest a hypocholesterolemic effect of soy protein. The effect may be partially attributable to the isoflavones in soy. Few studies have examined the separate effects of soy protein and isoflavones. OBJECTIVE: The objective of this study was to determine the effect of soy protein and isoflavones on plasma lipid concentrations in postmenopausal, moderately hypercholesterolemic women. DESIGN: This was a randomized, double-blind, placebo-controlled clinical trial with 3 treatment groups. After a 4-wk run-in phase during which the women consumed a milk protein supplement, the subjects were randomly assigned to 12 wk of dietary protein supplementation (42 g/d) with either a milk protein (Milk group) or 1 of 2 soy proteins containing either trace amounts of isoflavones (Soy- group) or 80 mg aglycone isoflavones (Soy+ group). RESULTS: LDL-cholesterol concentrations decreased more in the Soy+ group (n = 31) than in the Soy- group (n = 33) (0.38 compared with 0.09 mmol/L; P = 0.005), but neither of these changes was significantly different from the 0.26-mmol/L decrease observed in the Milk group (n = 30). The results for total cholesterol were similar to those for LDL cholesterol. There were no significant differences in HDL-cholesterol or triacylglycerol concentrations between the 3 groups. CONCLUSIONS: The difference in total- and LDL-cholesterol lowering between the 2 soy-protein supplements suggests an effect attributable to the isoflavone-containing fraction. However, the unexpected LDL-cholesterol lowering observed in the Milk group, and the fact that there was no significant difference between either soy group and the Milk group, suggests that changes may have been due to other factors related to participation in the study. PMID- 11273849 TI - APOE polymorphism and the hypertriglyceridemic effect of dietary sucrose. AB - BACKGROUND: The E4 allele of the apolipoprotein gene (APOE) is associated with a greater serum cholesterol response to dietary changes in fat and cholesterol. However, less is known about the interaction between APOE polymorphism and other macronutrients in the diet. OBJECTIVE: We evaluated the interaction between APOE polymorphism and dietary fat and carbohydrate, particularly sucrose, in relation to serum lipid concentrations. DESIGN: A total of 284 men and 130 women with coronary artery disease (mean age: 61 y; range: 33-74 y) participated in the cross-sectional EUROASPIRE study. Serum lipids and fatty acids in cholesteryl esters (CEs) were measured and APOE genotypes were determined. Dietary intake was examined by using a 4-d food record. RESULTS: Patients were grouped by APOE genotype: E2 (E2/E2 and E2/E3; n = 21), E3 (E3/E3; n = 245), and E4 (E4/E2, E4/E3, and E4/E4; n = 148). Patients with the E2 allele had lower LDL-cholesterol concentrations and tended to have higher triacylglycerol concentrations than did patients with the E3 or E4 allele; concentrations were not significantly different between the last 2 groups. In regression analysis, significant predictors of serum triacylglycerol were the interaction between sucrose intake and the E2 allele, proportion of n-3 fatty acids in CEs, body mass index, and diabetes. A high sucrose intake was associated with high triacylglycerol concentrations only in patients with the E2 allele. Interaction between saturated fat intake and the E2 allele, proportion of linoleic acid in CEs, and fiber intake predicted serum cholesterol. CONCLUSION: Coronary artery disease patients with the E2 allele will likely have a greater triacylglycerol response to high dietary sucrose intakes than will patients with the E3 or E4 allele. PMID- 11273848 TI - Alcohol drinking determines the effect of the APOE locus on LDL-cholesterol concentrations in men: the Framingham Offspring Study. AB - BACKGROUND: The effect of alcohol drinking on LDL-cholesterol concentrations is unclear. The reported variability may be due to interactions between genetic factors and alcohol intake. OBJECTIVE: The purpose of the study was to examine whether variation at the apolipoprotein E gene (APOE) locus modulates the association between alcohol drinking and LDL cholesterol. DESIGN: We used a cross sectional design in a healthy population-based sample of 1014 men and 1133 women from the Framingham Offspring Study. RESULTS: In male nondrinkers (n = 197), LDL cholesterol was not significantly different across APOE allele groups [APOE*E2 (E2), APOE*E3 (E3), and APOE*E4 (E4)]. However, in male drinkers (n = 817), differences were observed (P: < 0.001); those with the E2 allele had the lowest concentrations. LDL cholesterol in men with the E2 allele was significantly lower in drinkers than in nondrinkers but was significantly higher in drinkers than in nondrinkers in men with the E4 allele. This APOE-alcohol interaction remained significant (P < 0.001) after age, body mass index, smoking status, and fat and energy intakes were controlled for. In women, the expected effect of APOE alleles on LDL cholesterol occurred in both drinkers (n = 791; P < 0.001) and nondrinkers (n = 342; P < 0.001). Multiple linear regression models showed a negative association (P < 0.05) between alcohol and LDL cholesterol in men with the E2 allele but a positive association in men with the E4 allele. No significant associations were observed in men or women with the E3 allele. CONCLUSION: In men, the effects of alcohol intake on LDL cholesterol are modulated in part by variability at the APOE locus. PMID- 11273850 TI - American ginseng (Panax quinquefolius L.) attenuates postprandial glycemia in a time-dependent but not dose-dependent manner in healthy individuals. AB - BACKGROUND: We previously showed that 3 g American ginseng administered 40 min before an oral glucose challenge significantly reduces postprandial glycemia in subjects without diabetes. Whether this effect can be replicated with doses <3 g and administration times closer to the oral glucose challenge is unclear. OBJECTIVE: Our objective was to study the dosing and timing effects of American ginseng on postprandial glycemia. DESIGN: In a random crossover design, 12 healthy individuals [X +/- SEM age: 42 +/- 7 y; body mass index (BMI; in kg/m2): 24.1 +/- 1.1] received 16 treatments: 0 (placebo), 1, 2, or 3 g American ginseng at 40, 20, 10, or 0 min before a 25-g oral glucose challenge. Capillary blood was collected before administration and at 0, 15, 30, 45, 60, and 90 min after the start of the glucose challenge. RESULTS: Two-way analysis of variance showed that the main effects of treatment and administration time were significant (P < 0.05). Glycemia was lower over the last 45 min of the test after doses of 1, 2, or 3 g ginseng than after placebo (P < 0.05); there were no significant differences between doses. The reductions in the areas under the curve for these 3 doses were 14.4 +/- 6.5%, 10.6 +/- 4.0%, and 9.1 +/- 6%, respectively. Glycemia in the last hour of the test and area under the curve were significantly lower when ginseng was administered 40 min before the challenge than when it was administered 20, 10, or 0 min before the challenge (P < 0.05). CONCLUSIONS: American ginseng reduced postprandial glycemia in subjects without diabetes. These reductions were time dependent but not dose dependent: an effect was seen only when the ginseng was administered 40 min before the challenge. Doses within the range of 1-3 g were equally effective. PMID- 11273851 TI - Low-dose vitamin B-6 effectively lowers fasting plasma homocysteine in healthy elderly persons who are folate and riboflavin replete. AB - BACKGROUND: Current data suggest that physiologic doses of vitamin B-6 have no significant homocysteine-lowering effect. It is possible that an effect of vitamin B-6 was missed in previous trials because of a much greater effect of folic acid, vitamin B-12, or both. OBJECTIVE: The aim of this study was to investigate the effect of low-dose vitamin B-6 supplementation on fasting total homocysteine (tHcy) concentrations in healthy elderly persons who were made replete with folate and riboflavin. DESIGN: Twenty-two healthy elderly persons aged 63-80 y were supplemented with a low dose of vitamin B-6 (1.6 mg/d) for 12 wk in a randomized, double-blind, placebo-controlled trial after repletion with folic acid (400 microg/d for 6 wk) and riboflavin (1.6 mg/d for 18 wk); none of the subjects had a vitamin B-12 deficiency. RESULTS: Folic acid supplementation lowered fasting tHcy by 19.6% (P < 0.001). After folic acid supplementation, baseline tHcy concentrations ranged from 6.22 to 23.52 micromol/L and 10 subjects had suboptimal vitamin B-6 status (plasma pyridoxal-P < 20 nmol/L). Two-way analysis of variance showed that the significant improvement in vitamin B-6 status in response to vitamin B-6 supplementation (on the basis of both pyridoxal P: and the erythrocyte aspartate aminotransferase activation coefficient) was reflected in a significant reduction in plasma tHcy of 7.5%. CONCLUSIONS: Low dose vitamin B-6 effectively lowers fasting plasma tHcy in healthy subjects who are both folate and riboflavin replete. This suggests that any program aimed at the treatment or prevention of hyperhomocysteinemia should include vitamin B-6 supplementation. PMID- 11273852 TI - Folate intake of the Dutch population according to newly established liquid chromatography data for foods. AB - BACKGROUND: Determining folate intake is difficult because existing folate data in food-composition tables are scarce and unreliable. OBJECTIVE: The purposes of this study were first to analyze 125 of the most important foods that contribute to folate intake in the Netherlands and second to estimate the folate intake of a representative sample of the population. DESIGN: We analyzed the folate content of foods by using a newly developed HPLC trienzyme method combined with an affinity chromatography cleanup step. These results were then used to estimate the folate intake of persons aged 1-92 y who participated in the second Dutch National Food Consumption Survey (DNFCS) in 1992 (n = 6218). RESULTS: For 35 important folate-containing foods, the mean relative folate contents measured by HPLC were 66%, 80%, and 77% of values for comparable foods included in the British food-composition table; the Ministry of Agriculture, Fisheries and Food table; and the US Department of Agriculture database, respectively. P values for comparison of relative values with 100% were 0.001, 0.171, and 0.144, respectively. The mean dietary folate intake of the DNFCS participants was 182 +/ 119 microg/d. Intake of supplement users (n = 86) was 344 microg/d, with 147 microg/d from supplements. On the basis of these findings, 42% of men and 54% of women do not meet current Dutch recommendations of 60 microg/d for children and 200 microg/d for adults. CONCLUSIONS: Total folate quantities in foods, analyzed by HPLC, are approximately 25% lower than amounts listed in recent food composition tables estimated by use of the microbiological method. On the basis of these new data, approximately 50% of a representative Dutch population sample does not meet the current recommendations for folate intake. PMID- 11273854 TI - Concurrent micronutrient deficiencies in lactating mothers and their infants in Indonesia. AB - BACKGROUND: Deficiencies of vitamin A, iron, and zinc are prevalent worldwide, affecting vulnerable groups such as lactating women and infants. However, the existence of concurrent deficiencies has received little attention. OBJECTIVE: The aim was to investigate the extent to which deficiencies of vitamin A, iron, and zinc coexist and the nutritional relation between lactating mothers and their infants. DESIGN: In a cross-sectional survey in rural West Java, Indonesia, 155 lactating mothers and their healthy infants were assessed anthropometrically and blood, urine, and breast-milk samples were obtained. RESULTS: Marginal vitamin A deficiency was found in 54% of the infants and 18% of the mothers. More than 50% of the mothers and infants were anemic and 17% of the infants and 25% of the mothers were zinc deficient. There was a strong interrelation between the micronutrient status of the mothers and infants and the concentrations of retinol and beta-carotene in breast milk. Vitamin A deficiency in infants led to an increased risk of anemia and zinc deficiency (odds ratios: 2.5 and 2.9, respectively), whereas in mothers the risk of anemia and iron deficiency (odds ratios: 3.8 and 4.8, respectively) increased. In infants, concentrations of insulin-like growth factor I were related to concentrations of plasma retinol and beta-carotene but not to zinc. CONCLUSIONS: Micronutrient deficiencies were prevalent in West Java. The micronutrient status of lactating mothers and that of their infants were closely related; breast milk was a key connecting factor for vitamin A status. Furthermore, concurrent micronutrient deficiencies appeared to be the norm. PMID- 11273853 TI - Serum concentrations of beta-carotene and alpha-tocopherol are associated with diet, smoking, and general and central adiposity. AB - BACKGROUND: Previous studies of associations between diet, obesity, and blood concentrations of alpha-tocopherol and beta-carotene have been equivocal. Furthermore, most studies used only body mass index (BMI) as an obesity measure. OBJECTIVES: Our objectives were to examine the associations between energy and nutrient intakes, alcohol consumption, tobacco use, and serum cholesterol and serum concentrations of alpha-tocopherol and beta-carotene, and to examine the associations between different measures of general and central adiposity and serum concentrations of alpha-tocopherol and beta-carotene. DESIGN: This was a cross-sectional, population-based study of 253 men and 276 women aged 46-67 y. Nutrient data were collected by a modified diet history method. Measures of obesity included BMI, percentage of body fat (impedance analysis), waist-to-hip ratio, and waist circumference. The associations between serum nutrient concentrations and the other factors were examined by multiple linear regression. RESULTS: Twenty-one percent of men and 34% of women used antioxidant supplements. The mean BMI was 26.1 in men and 25.4 in women. Serum beta-carotene concentration was positively associated with serum cholesterol concentration, fiber intake, and beta-carotene intake, and negatively associated with smoking and all measures of obesity. In men, serum beta-carotene concentration was not significantly associated with central adiposity after adjustment for body fat. Serum alpha tocopherol concentration was positively correlated with serum cholesterol, obesity, and vitamin E intake. In women, serum alpha-tocopherol concentration was also positively associated with intakes of ascorbic acid and selenium. Serum alpha-tocopherol concentration was associated with central adiposity after adjustment for body fat. CONCLUSION: Serum beta-carotene and alpha-tocopherol concentrations have different associations with diet, smoking, general adiposity, and central adiposity. PMID- 11273855 TI - Intake of major nutrients by women in the Maternal Phenylketonuria (MPKU) Study and effects on plasma phenylalanine concentrations. AB - BACKGROUND: Women with untreated phenylketonuria (PKU) often have poor reproductive outcomes. OBJECTIVE: We assessed the effects of intakes of major nutrients on plasma phenylalanine concentrations and we measured phenylalanine hydroxylase activity and phenylalanine intakes in pregnant women with PKU. DESIGN: Dietary intakes and plasma phenylalanine concentrations were compared in 4 subject groups defined on the basis of plasma phenylalanine concentrations: group 1 (n = 23), <360 micromol/L by 10 wk gestation and 120-360 micromol/L throughout the remainder of pregnancy; group 2 (n = 46), <600 micromol/L but not <360 micromol/L by 10 wk gestation and 120-600 micromol/L throughout the remainder of pregnancy; group 3 (n = 24), <600 micromol/L by 10 wk gestation but >600 micromol/L at least once thereafter; group 4 (n = 147), never <600 micromol/L. RESULTS: Except in the first trimester, mean intakes of phenylalanine, energy, and fat tended to be greater in group 1 than in the other groups. The mean protein intake of group 1 tended to be greater than that of the other groups. Intakes of protein (P < 0.0001), fat (P < 0.0001), and energy (P < 0.007) were negatively correlated with maternal plasma phenylalanine concentrations. It appeared that genotype did not affect phenylalanine tolerance. CONCLUSIONS: Maternal genotype appeared to have little influence on phenylalanine requirements during the first trimester. Early decline and maintenance of maternal plasma phenylalanine concentrations at <360 micromol/L and mean protein intake greater than the recommended dietary allowance (RDA) with mean energy intake near the RDA resulted in the best reproductive outcomes. Inadequate intakes of protein, fat, and energy may result in elevated plasma phenylalanine concentrations and may contribute to poor reproductive outcomes. PMID- 11273856 TI - Essential fatty acid composition of plasma phospholipids and birth weight: a study in term neonates. AB - BACKGROUND: Essential fatty acids (EFAs) in umbilical cord blood samples are associated with attained birth weight in premature infants and low-birth-weight neonates. OBJECTIVE: The objective was to investigate relations between the EFA composition of cord and maternal plasma phospholipids and birth weight in term neonates. DESIGN: This was a cross-sectional study in 627 singletons born at term. The plasma phospholipid EFA composition of the mothers was determined by gas-liquid chromatography at study entry (< or = 16 wk gestation), at delivery, and in cord plasma at birth. Birth weights were normalized to SD scores. RESULTS: In cord plasma, the dihomo-gamma-linolenic acid concentration was positively related to weight SD scores. Both arachidonic acid (AA) and docosahexaenoic acid (DHA) were negatively related to weight SD scores. EFA-status indicators showed similar negative associations, whereas eicosatrienoic acid concentrations were positively related to neonatal size. In maternal plasma, proportions of n-3 long chain polyenes (LCPs) and n-6 LCPs decreased during pregnancy. Larger decreases in AA, DHA, n-3 LCP, and n-6 LCP fractions were observed in mothers of heavier babies. Higher concentrations of LCPs in maternal plasma were, however, not related to a larger infant size at birth. CONCLUSIONS: A lower biochemical EFA status in umbilical cord plasma and a larger decrease in maternal plasma LCP concentrations are associated with a higher weight-for-gestational-age at birth in term neonates. Our findings do not support a growth-stimulating effect of AA or DHA; however, they do suggest that maternal-to-fetal transfer of EFAs might be a limiting factor in determining neonatal EFA status. PMID- 11273857 TI - Infant plasma trans, n-6, and n-3 fatty acids and conjugated linoleic acids are related to maternal plasma fatty acids, length of gestation, and birth weight and length. AB - BACKGROUND: Arachidonic acid (AA) and docosahexaenoic acid (DHA) are important for growth and neural development. trans Fatty acids (TFAs) may inhibit desaturation of linoleic acid (LA) and alpha-linolenic acid (ALA) to AA and DHA, respectively. Conjugated linoleic acids (CLAs) also alter lipid metabolism and body fat. OBJECTIVE: We determined the associations of birth outcome with maternal and infant plasma concentrations of TFAs, CLAs, AA, and DHA. DESIGN: In healthy women, we sampled maternal blood at 35 wk gestation (n = 58) and umbilical cord blood at birth (n = 70). RESULTS: Mean (+/- SEM) TFA concentrations (% by wt) in infant plasma were as follows: triacylglycerol, 2.83 +/- 0.19 (range: 0.63-12.79); phospholipid, 0.67 +/- 0.03 (0.11-1.33); and cholesteryl ester, 2.04 +/- 0.01 (0.86-4.24). LA, AA, DHA, TFA, and CLA concentrations in infant phospholipids correlated with the same fatty acid in maternal plasma phospholipids (n = 44; P < 0.05). Infant plasma cholesteryl ester and triacylglycerol TFAs and cholesteryl ester CLAs (r = -0.33, -0.42, and -0.49, respectively) were significantly inversely related to length of gestation. Triacylglycerol and cholesteryl ester AA were positively related to length of gestation (r = 0.41 and 0.37, respectively) and birth weight (r = 0.27 and 0.23, respectively). Inverse correlations occurred between infant plasma TFA and DHA concentrations in triacylglycerols (r = -0.33) and between TFA and AA concentrations in cholesteryl esters (r = -0.23). CONCLUSION: The results suggest possible important effects of TFAs and of AA on fetal growth and length of gestation. PMID- 11273858 TI - Assessing the effect of fatty acids on prostate carcinogenesis in humans: does self-reported dietary intake rank prostatic exposure correctly? AB - BACKGROUND: Dietary fatty acids may influence prostate carcinogenesis. Although the standard for assessing dietary effects in humans is the semiquantitative food frequency questionnaire, the extent to which self-reported intake correctly ranks prostatic exposure is unknown. OBJECTIVE: The objective was to examine the correlation between reported intakes of different fatty acids and their concentrations in prostate tissue. DESIGN: This was a cross-sectional study of 52 men undergoing surgical resection of the prostate gland. Usual dietary intake of saturated, total unsaturated, oleic, and linoleic fatty acids over the previous year was estimated with use of a 122-item version of the Health Habits and History Questionnaire. Concentrations in prostate tissue were measured directly by use of gas chromatography in healthy tissue collected at the time of surgery and were expressed as a percentage of total fatty acids. Correlations with 4 measures of dietary intake [g/d, g/d adjusted for total daily energy intake, % of total fat (as g/d), and % of total energy] were evaluated by Spearman's rank order correlation coefficients. RESULTS: Linoleic acid concentrations in prostate tissue were significantly correlated with dietary intake expressed as g/d adjusted for total energy [r = 0.29 (95% CI: 0.03, 0.49), P = 0.04], % of total fat [r = 0.36 (0.14, 0.550), P = 0.008], and % of total energy [r = 0.28 (0.04, 0.49), P = 0.042], but not as g/d. Although mean concentrations of saturated, total unsaturated, and oleic fatty acids in prostate tissue resembled mean intakes for the group, prostatic concentrations did not correlate with individual intakes. CONCLUSION: Self-reported intake of fatty acids is a satisfactory marker of prostatic exposure at the group level, but, with the exception of linoleic acid, does not correctly rank individuals with respect to intensity of exposure. PMID- 11273859 TI - Bone mineral content in girls perinatally infected with HIV. AB - BACKGROUND: Early diagnostic efforts and advances in multidrug therapy have considerably prolonged the survival time of children infected perinatally with HIV. Despite these advances, few studies have addressed calcium status and bone growth in HIV-infected children. OBJECTIVE: Our objective was to examine the effect of HIV infection on calcium status and bone growth in children. DESIGN: We measured calcitropic hormones, urinary calcium excretion, bone mineral content, and body composition in 19 young girls aged 9.2 +/- 2.6 y (range: 5.9-15.2 y) who were infected perinatally with HIV. RESULTS: Serum concentrations of 1,25 dihydroxyvitamin D [1,25(OH)(2)D] and parathyroid hormone concentrations were elevated above normal ranges in 25% and 12% of these girls, respectively. Urinary calcium excretion normalized for creatinine excretion was also elevated (Ca/Cr >0.18) in 17% of these children despite suboptimal calcium intakes (679 +/- 437 mg/d). Total-body bone mineral content, measured with the use of dual-energy X ray absorptiometry, averaged 845.1 +/- 279.0 g and was on average 2.7 z scores below age- and race-matched values reported in non-HIV-infected healthy girls. Significant positive correlations were found between an indirect marker of bone resorption in urine (N:-telopeptide) and 1,25(OH)2D (P < 0.02, r2 = 0.586, n = 9), and between serum N-telopeptide and total alkaline phosphatase (P < 0.001, r2 = 0.541, n = 17), suggesting that calcium insufficiency may be increasing bone resorption in this group. CONCLUSIONS: Young girls with HIV infection had low bone mass and evidence of calcium insufficiency. Nutritional counseling of children with HIV infection should emphasize adequate calcium intakes because of the importance of this age period in bone mineral acquisition. PMID- 11273860 TI - Relation between body composition, fat distribution, and lung function in elderly men. AB - BACKGROUND: Body composition changes with age, with increases in fat mass and visceral fat and declines in skeletal muscle mass; lung function also declines with age. Age-related changes in body composition and fat distribution may be associated with the pulmonary impairment observed in the elderly. OBJECTIVE: Our goal was to evaluate the relations between body composition, fat distribution, and lung function in elderly men. DESIGN: We studied 97 men aged 67-78 y with body mass indexes (BMIs; in kg/m2) ranging from 19.8 to 37.1. Body composition was evaluated by using dual-energy X-ray absorptiometry and fat distribution was evaluated by using waist and hip circumferences, waist-to-hip ratio, and sagittal abdominal diameter (SAD). Spirometry was done in all subjects and the distance walked by each subject during a 6-min walking test was evaluated as was leg strength. RESULTS: A significant negative correlation was found between adiposity, fat distribution indexes, forced vital capacity (FVC), and forced expiratory volume in 1 s (FEV1). A positive correlation was found between fat free mass and FVC. After adjustment for age, height, and weight, SAD still correlated negatively with FVC and FEV1 (r = -0.367 and -0.348, respectively; P < 0.01), whereas percentage body fat and fat mass correlated negatively and fat free mass correlated positively with FVC (r = -0.313, -0.323, and 0.299, respectively; all P < 0.01). After the sample was subdivided by tertile of fat free mass adjusted for age and BMI, FVC and FEV1 were significantly lower in the lowest fat-free mass tertile (P < 0.01). Stepwise multiple regression analysis performed with use of lung function variables as the dependent variables and age, height, fat mass, fat-free mass, waist circumference, and SAD as the independent variables showed that 3 variables entered the regression for predicting FVC: height, which entered the regression first; SAD, which entered second; and fat free mass, which entered third. Only 2 variables entered the regression for predicting FEV1: height, which entered the regression first, and SAD, which entered second. CONCLUSION: Our cross-sectional data show a significant association between body composition, fat distribution, and lung function in elderly men. PMID- 11273861 TI - Functional and metabolic early changes in calf muscle occurring during nutritional repletion in malnourished elderly patients. AB - BACKGROUND: Metabolic alterations in skeletal muscle associated with malnutrition and the potential reversibility of such alterations during refeeding are not fully understood. OBJECTIVE: We characterized early changes in muscle during refeeding in malnourished, hospitalized elderly subjects. DESIGN: Muscle function, metabolism, and mass were evaluated in 24 clinically stable patients (11 were malnourished) by using isokinetic plantar flexor torque measurements and nuclear magnetic resonance (NMR) imaging for medial gastrocnemius mass assessment and 31P and 13C NMR spectroscopy for inorganic phosphate (Pi), phosphocreatine, and glycogen quantitation. RESULTS: Malnourished subjects had lower muscle mass (P < 0.02) and tended to have lower strength than did control subjects. In malnourished subjects, muscle strength increased after refeeding (P < 0.01) whereas muscle mass was unchanged. The ratio of Pi to ATP was lower in malnourished than in control subjects (P < 0.001) and increased during refeeding (P < 0.01). The mean ratio of phosphocreatine to ATP was lower in malnourished than in control subjects (P < 0.01) and increased to control values after refeeding. Muscle glycogen showed a scattered distribution for malnourished subjects; the mean value did not differ significantly from that of control subjects, either at baseline or after refeeding. CONCLUSIONS: The lower ratio of phosphocreatine to ATP in malnourished subjects could have resulted from either lower total muscle creatine or reduced oxidative capacities. High or normal glycogen associated with a low Pi-to-ATP ratio in malnourished subjects suggested preferential use of lipid over carbohydrate for energy supply, which is known to reduce muscle performance. The data suggest that normalization of muscle metabolite content after refeeding improves muscle strength in malnourished subjects. PMID- 11273862 TI - Association between urinary potassium, urinary sodium, current diet, and bone density in prepubertal children. AB - BACKGROUND: Our understanding of the role of nutrients in bone development in children is limited. OBJECTIVE: We examined the associations between urinary potassium, urinary sodium, usual dietary intake, and bone mineral density (BMD) in prepubertal children. DESIGN: This was a cross-sectional study of 330 boys and girls aged 8 y. Urinary measures were assessed in a single, timed, overnight urine specimen. Usual diet was assessed with a food-frequency questionnaire completed by a parent or guardian. BMD at the femoral neck, lumbar spine, and total body was measured by dual-energy X-ray absorptiometry. RESULTS: Urinary potassium correlated significantly with BMD at all sites (femoral neck: r = 0.20, P < 0.001; lumbar spine: r = 0.19, P = 0.001; total body: r = 0.24, P < 0.001). After adjustment for confounders (primarily lean body mass), this association was lower in magnitude but remained significant at 2 sites with a consistent trend at the third (femoral neck: P = 0.15; lumbar spine: P = 0.046; total body: P = 0.028). Urinary sodium was not associated with BMD at any site. No nutrient or food intake estimate was associated with BMD, although urinary potassium correlated significantly with potassium intake (r = 0.14, P = 0.016) and fruit and vegetable intake (r = 0.12, P = 0.033). CONCLUSIONS: Urinary potassium was associated with both dietary intake and BMD independent of lean body mass in these well-nourished, calcium-replete young children. These findings should be confirmed in further longitudinal studies. Nevertheless, this association is likely to represent dietary intake of potassium and suggests that measurement of urinary potassium is superior to food-frequency questionnaires for assessing potassium intake in this age group. PMID- 11273864 TI - Estimates of body water compartments with use of bioelectrical impedance analysis. PMID- 11273865 TI - Viral infections after renal transplantation. AB - Viral infections are a leading cause of posttransplantation morbidity and mortality. A number of recent developments have altered our understanding and management of these disorders. The pathogenetic roles of several viruses, including human herpesviruses 6 and 8, have been newly established. Molecular based diagnostic tests now make more rapid diagnosis possible. The licensing of new potent antiviral agents offers a wider choice of drugs for viral prophylaxis and treatment. The use of more potent immunosuppressive agents is responsible in part for the increasing incidence of some viral infections, but this varies among drugs, and individual viruses differ in their sensitivity to immunosuppressive agents. This review summarizes the natural history, diagnosis, prevention, and treatment of many common viral infections after renal transplantation. PMID- 11273866 TI - Aldosterone as a mediator of progressive renal disease: pathogenetic and clinical implications. AB - End-stage renal disease is an enormous public health burden with an increasing incidence and prevalence. This escalating prevalence suggests that newer therapeutic interventions and strategies are needed to complement current antihypertensive approaches. Although much evidence shows that angiotensin II mediates progressive renal disease, recent evidence also implicates aldosterone as an important pathogenetic factor in progressive renal disease. Several lines of experimental evidence show that selective blockade of aldosterone, independent of renin-angiotensin blockade, reduces proteinuria and nephrosclerosis in the spontaneously hypertensive stroke-prone rat model and reduces proteinuria and glomerulosclerosis in the subtotally nephrectomized rat model (ie, remnant kidney). Although pharmacological blockade with angiotensin II-receptor blockers and angiotensin-converting enzyme inhibitors reduces proteinuria and nephrosclerosis and/or glomerulosclerosis, selective reinfusion of aldosterone restores these abnormalities despite continued renin-angiotensin blockade. Based on this theoretic construct, randomized clinical studies will be initiated to delineate the potential renal-protective effects of antihypertensive therapy using aldosterone-receptor blockade. This is a US government work. There are no restrictions on its use. PMID- 11273867 TI - Hypocalcemia: a pervasive metabolic abnormality in the critically ill. AB - Hypocalcemia has been reported in critically ill patients, most commonly in association with sepsis syndrome. However, the severity and incidence of hypocalcemia in nonseptic but critically ill patients has not been well defined. Therefore, the goal of this study was to identify and compare the frequency and degree of hypocalcemia in critically ill patients with differing underlying illnesses (those admitted to medical, surgical, trauma, neurosurgical, burn, respiratory, and coronary intensive care units [ICUs]; group A; n = 99). Results were compared with the frequency and degree of hypocalcemia in non-critically ill ICU patients (initially admitted to an ICU but discharged within 48 hours; group B; n = 50) or hospitalized non-ICU patients (group C; n = 50). Incidences of hypocalcemia (ionized calcium [Ca] < 1.16 mmol/L [less than normal]) were 88%, 66%, and 26% for groups A, B, and C, respectively (P: < 0.001). In group A, the frequency of hypocalcemia did not depend on the ICU setting or presence of sepsis. However, the occurrence of hypocalcemia correlated with both Acute Physiology and Chronic Health Evaluation II score (r = -0.39; P: < 0.001) and patient mortality (eg, hazard ratio for death, 1.65 for Ca decrements of 0.1 mmol/L; P: < 0.002). Hypomagnesemia, number of blood transfusions, and presence of acute renal failure were each associated with depressed Ca levels. A weak association (r = -0.12; P: = 0.09) was noted between serum Ca level and QT interval. Clinical concern stemming from hypocalcemia was underscored by the substantial use of intravenous (IV) Ca therapy ( approximately 2 to 3 g IV). We conclude that hypocalcemia is extremely common in hospitalized patients (up to 88%) and correlates with severity of illness, but not with a specific illness per se. Whether it directly impacts patient survival remains unknown. Resolution of this issue appears to be critical because of the frequency with which it leads to high-dose IV Ca therapy. PMID- 11273869 TI - How echogenic is echogenic? Quantitative acoustics of the renal cortex. AB - The echogenicity of the cortex is an important parameter in interpreting renal sonograms that suggest changes in cortical structure. Echogenicity is currently measured qualitatively, and no attempts have been made at quantification. We developed a method to quantify renal cortical echogenicity in reference to the liver and evaluated its reproducibility, dependence on scanning variables, and potential utility. Sonograms of the right kidney were digitized, and the mean pixel density of regions of the renal cortex and liver was measured and normalized to the gray scale. Echogenicity was expressed as the ratio of the brightness (inverse of mean pixel density) of the cortex to that of the liver. The mean coefficient of variation among measurements performed on multiple sonograms from the same study was 2.8%, and the coefficient of variation among multiple measurements performed on the same kidney over 1 year was 1.8%. The correlation between measurements obtained by two different individuals on identical images was 0.92, with a mean variation of 3.0%. Echogenicity was not significantly affected by type of scanner or probe frequency, but varied inversely with gain. However, the effect of gain was very small within the useful range. Water loading after an overnight fast increased echogenicity in all cases, with a mean increase of 6.4%. Echogenicity of normal kidneys was significantly less than that of the liver (range, 0.810 to 0.987), and in clinical sonograms analyzed retrospectively but blindly, echogenicity correlated with the qualitative gradations of echogenicity originally assigned. The most echogenic kidneys were 62% brighter than normal kidneys, many times greater than the variability of the measurement. We conclude that quantification of renal cortical echogenicity is feasible and reproducible and may be useful in detecting and following renal disease. Echogenicity of the renal cortex is less than that of the liver in healthy subjects and is influenced by the state of diuresis. PMID- 11273868 TI - Screening for primary aldosteronism without discontinuing hypertensive medications: plasma aldosterone-renin ratio. AB - The traditional workup for primary aldosteronism is cumbersome and requires discontinuing antihypertensive medications, which is inconvenient and potentially dangerous. A simple and accurate screening test that can be used without modifying medications is needed. The plasma aldosterone-renin ratio (ARR) is a valid screening assay for primary aldosteronism, but antihypertensives are usually discontinued before obtaining this ratio, limiting its utility. The present prospective study is designed to examine the validity of the ARR as a screening test for primary aldosteronism if the ratio is measured randomly while patients continue antihypertensive therapy. During the 18-month study period, 90 patients were referred to the hypertension clinic with poorly controlled hypertension. ARR was measured in random blood samples in all 90 patients while maintaining their prescribed antihypertensive medications. Those with elevated ARRs (>100 ng/dL / ng/mL/h) underwent further diagnostic workup, including adrenal computed tomography and/or magnetic resonance imaging and adrenal iodine 131 norcholesterol uptake scan. Fifteen patients (17%) had elevated ARRs greater than 100:1. Ten of 15 patients were found to have adrenal adenoma on diagnostic workup, and adenoma was later confirmed by histological examination after surgical removal in these 10 patients. Five patients were found to have adrenal hyperplasia; all 5 patients responded to antialdosterone treatment. Thus, all 15 patients had good control of blood pressure after surgery and/or antialdosterone medications. No patient showed a falsely elevated ARR. Data suggest that the ARR is a valid screening assay for primary aldosteronism in patients with poorly controlled blood pressure, and discontinuation of antihypertensive medications is not needed for this test. PMID- 11273870 TI - Macrophage subclasses and proliferation in childhood IgA glomerulonephritis. AB - We immunohistologically compared the number of intraglomerular infiltrating cells in 14 children with poststreptococcal acute glomerulonephritis (PSAGN) and 20 children with immunoglobulin A glomerulonephritis (IgAGN) with histological characteristics similar to those of PSAGN to explain the difference in clinicopathological characteristics between these two diseases. Immunohistological study was performed in kidney tissues from these patients by using monoclonal antibodies of T-cell marker (CD3 and CD45RO), B-cell marker (CD20), neutrophil marker (CD15), macrophage marker (CD68), four subclasses of macrophages (early-stage, acute-stage, chronic-stage, and mature inflammatory macrophage marker), and proliferating cell nuclear antigen (PCNA). The 34 patients were classified into three stages according to the time from the detection of urinary abnormalities to biopsy. Intraglomerular immunopositive cells were expressed as the number of cells per glomerulus. There were more intraglomerular positive cells of CD15, CD68, and the four macrophage subclasses in PSAGN than IgAGN. The number of intraglomerular infiltrating macrophages decreased with time in PSAGN, whereas the number of macrophages in IgAGN remained constant at all stages. Intraglomerular infiltration of acute-stage inflammatory macrophages alone was evident in IgAGN. Both the number of intraglomerular proliferating macrophages (PCNA-positive plus CD68-positive cells) and proportion of proliferating macrophages/total macrophages were greater in IgAGN than PSAGN. Normal urinalysis results were evident in all patients with PSAGN during follow up, and urinary abnormalities persisted in 18 patients with IgAGN. In conclusion, differences in the maturity of infiltrating macrophages and number of proliferating macrophages are associated with the different clinicopathological characteristics in children with PSAGN and IgAGN. PMID- 11273871 TI - Excessive body weight as a new independent risk factor for clinical and pathological progression in primary IgA nephritis. AB - Experimental evidence suggests a role for obesity in the formation and progression of some glomerular lesions, but data for human glomerulonephritis are lacking. In a cohort of 162 incident patients with biopsy-proven immunoglobulin A (IgA) nephropathy, we assessed whether the presence of an elevated body mass index (BMI >/= 25 kg/m(2)) at the time of the first renal biopsy (RB1) correlated with clinical data at RB1 (24-hour proteinuria, arterial hypertension, and renal function), pathological data (global optical score [GOS] with detailed pathological indices), and clinical progression to both arterial hypertension and chronic renal failure (CRF). In both univariate and multivariate analyses, the presence of an elevated BMI at RB1 was significantly associated with the severity of pathological renal lesions (GOS and vascular, tubular, and interstitial indices). Hypertension-free survival was significantly less in overweight patients (P: < 0.0001) compared with those with normal weight. In a Cox regression analysis for hypertension-free survival including 24-hour proteinuria greater than 1 g, GOS, and metabolic parameters, only elevated BMI and GOS were independent factors for the development of arterial hypertension. CRF-free survival was also significantly less in patients with an excessive BMI. In a multivariate Cox regression analysis for CRF-free survival, hypertension, GOS, and BMI at RB1 were independent risk factors for CRF. In IgA nephropathy, excessive body weight and/or BMI are underestimated predictive factors for the development of arterial hypertension and, ultimately, CRF. PMID- 11273872 TI - Renal vascular walls in patients with preeclampsia superimposed on essential hypertension. AB - This study was performed to clarify the relationship between changes in contractile proteins in renal vascular walls and the prognosis of hypertension during pregnancy. Twenty preeclamptic patients underwent renal biopsies after delivery and were divided into the following three groups: group I, patients with persistent hypertension after delivery (n = 7; mean age, 34.8 +/- 1.4 years [SE]); group II, patients who became normotensive after delivery and hypertensive again during follow-up (n = 5; mean age, 34.8 +/- 1.6 years), and group III, patients who became normotensive after delivery (n = 8; mean age, 28.0 +/- 1.0 years). We also examined age-matched healthy controls (group IV; n = 7; mean age, 34.9 +/- 1.5 years). Renal biopsy specimens were immunohistochemically stained by the avidin-biotinylated peroxidase complex method using antimonoclonal smooth muscle cell myosin heavy chain isoform antibodies (SM-1, SM-2) and antimonoclonal alpha-smooth muscle cell actin antibody (actin). We estimated and semiquantitatively scored the degree of staining in each section. In interlobular arteries, SM-1, SM-2, and actin staining in group I were significantly reduced compared with group IV (SM-1, SM-2, P: < 0.05; actin, P: < 0.01). In afferent arterioles (Afs), SM-1, SM-2, and actin staining were reduced in group I. SM-2 staining in group I was significantly reduced compared with the other three groups (versus group II, P: < 0.05; versus groups III and IV, P: < 0.01). These findings suggest that phenotypic changes in vascular smooth muscle cells (especially the disappearance of SM-2 in Afs) reflect the stage of underlying essential hypertension and can predict from the change in hypertension during pregnancy whether it will persist after delivery. PMID- 11273874 TI - Suspected iron dextran-related adverse drug events in hemodialysis patients. AB - Despite the use of recombinant erythropoietin, anemia remains a significant problem for patients with end-stage renal disease, in part related to chronic dialysis-related blood loss and resultant iron deficiency. Because oral iron preparations have been relatively ineffective and poorly tolerated in this population, intravenous (IV) iron dextran has been widely prescribed, despite a finite risk for adverse effects associated with its use. We analyzed data from Fresenius Medical Care North America (FMCNA) clinical variance reports to determine the incidence of suspected iron dextran-related adverse drug events (ADEs) and associated patient characteristics, dialysis practice patterns, and outcomes. We used a case-cohort study design, comparing individuals who experienced suspected ADEs with the overall FMCNA population. Among 841,252 IV iron dextran administrations from October 1998 through March 1999, there were 165 reported suspected ADEs, corresponding to an overall rate of 0.000196%, or approximately 20 per 100,000 doses. Forty-three patients (26%) required an independent emergency department evaluation, 18 patients (11%) required hospitalization, and 1 patient (0.6%) died. Dyspnea (43%), hypotension (23%), and neurological symptoms (23%) were the most common major ADEs; nausea (34%), vomiting (23%), flushing (27%), and pruritus (25%) were the most common other ADEs. ADEs were 8.1-fold more common among patients administered Dexferrum (American Regent Laboratories, Inc, Shirley, NY) compared with those administered InFed (Watson Pharmaceuticals, Phoenix, AZ). In summary, serious adverse reactions to IV iron dextran are rare in clinical practice. The risk appears to depend on the specific formulation of IV iron dextran. Otherwise, iron dextran related ADEs are difficult to predict. PMID- 11273873 TI - Erythropoietin deficiency causes anemia in nephrotic children with normal kidney function. AB - Anemia in persistent nephrotic syndrome (NS) has been described in a few case reports but has not been studied systematically. We present a group of 19 children with NS who developed anemia before the deterioration of kidney function. The aim of our study is to determine whether erythropoietin (EPO) and/or iron deficiency are causative factors and to evaluate the effect of EPO replacement therapy. Serum EPO levels, iron status, and vitamin B(12) concentrations were measured in nephrotic patients with anemia (NS-A) and compared with those of nephrotic children with normal hemoglobin (Hb) levels (NS NHb; n = 13). Two control groups consisted of age-matched patients without kidney disease or hypoxemia with either iron deficiency anemia (IDA; n = 19) or normal Hb concentrations (NHb; n = 16). Most NS-A patients experienced persistent steroid-resistant NS, whereas most NS-NHb children had steroid-responsive NS. Although serum iron, ferritin, and B(12) levels were significantly lower in NS-A children, appropriate replacement therapy that resulted in normalization of ferritin and/or cobalamin levels did not lead to correction of the anemia. NS-A patients had greater EPO levels than those without anemia (21.6 +/- 3.3 versus 5.5 +/- 0.8 IU/L; P: < 0.001), but their response to anemia was inappropriately low compared with IDA children (EPO, 94.6 +/- 15.1 IU/L) despite similar Hb concentrations. EPO therapy for 4 to 9 months in 6 NS-A children with Hb levels less than 9 g/dL led to resolution of the anemia. In conclusion, anemia is a common feature of persistent NS that develops before the deterioration of kidney function. Depletion of iron stores may contribute to the development of anemia, but iron replacement therapy is ineffective. Nephrotic patients have EPO deficiency with a blunted response to anemia. The EPO deficiency is amenable to EPO therapy, which is recommended for this group of patients. PMID- 11273875 TI - Melatonin prevents oxidative stress resulting from iron and erythropoietin administration. AB - Intravenous iron (Fe) and recombinant human erythropoietin (rHuEPO) are routine treatments in the management of anemia in patients with chronic renal failure. We investigated the oxidative stress acutely induced by these therapies and whether pretreatment with oral melatonin (MEL) would have a beneficial effect. Nine patients (four women) were studied within 1 month of entering a chronic hemodialysis program in the interdialytic period. Plasma malondialdehyde (MDA), red blood cell glutathione (GSH), and catalase (CAT) activity were measured in blood samples obtained before (baseline) and 1, 3, and 24 hours after the administration of Fe (100 mg of Fe saccharate intravenously over 1 hour) or rHuEPO (4,000 U intravenously). One hour before these treatments, patients were administered a single oral dose of MEL (0.3 mg/kg) or placebo. Each patient was studied on four occasions, corresponding to studies performed using either placebo or MEL in association with intravenous Fe and rHuEPO administration. Baseline data showed increased oxidative stress in patients with end-stage renal failure. Increments in oxidative stress induced by Fe were more pronounced at the end of the administration: MDA, baseline, 0.74 +/- 0.09 nmol/mL; 1 hour, 1.50 +/- 0.28 nmol/mL (P: < 0.001); GSH, baseline, 2.51 +/- 0.34 nmol/mg of hemoglobin (Hb); 1 hour, 1.66 +/- 0.01 nmol/mg Hb (P: < 0.001); and CAT activity, baseline, 27.0 +/- 5.7 kappa/mg Hb; 1 hour, 23.3 +/- 4.2 kappa/mg Hb (P: < 0.001). rHuEPO induced increments in oxidative stress were more pronounced (P: < 0.001) at 3 hours (MDA, 1.24 +/- 0.34 nmol/mL; GSH, 1.52 +/- 0.23 nmol/mg Hb; CAT activity, 18.0 +/- 3.1 kappa/mg Hb). MEL administration prevented the changes induced by Fe and rHuEPO and had no adverse side effects. These studies show that intravenous Fe and rHuEPO in doses commonly used to treat anemia in chronic hemodialysis patients acutely generate significant oxidative stress. Oral MEL prevents such oxidative stress and may be of clinical use. PMID- 11273876 TI - Efficacy of folinic versus folic acid for the correction of hyperhomocysteinemia in hemodialysis patients. AB - The effectiveness of intravenous folinic acid or intravenous folic acid for the treatment of hyperhomocysteinemia of hemodialysis patients is unknown. In a randomized, controlled, double-blind trial, 66 hemodialysis patients were administered either 15 mg of folic acid or an equimolar amount (16.1 mg) of folinic acid intravenously three times weekly. Normalization of total homocysteine (tHcy) plasma levels after 4 weeks of treatment was achieved in 10 patients (30.3%) in the folic-acid group and 6 patients (18.2%; P: = 0.389) in the folinic-acid group (normalization at any time during the study period in 39.4% and 33.3% of the patients; P: = 0.798). The relative reduction in tHcy plasma levels at week 4 was 32.2% in the folic-acid group and 34.1% in the folinic-acid group. A high baseline tHcy plasma concentration (P: = 0.00001), methylenetetrahydrofolate reductase (MTHFR) 677TT/1298AA genotype (P: = 0.03540), and low red blood cell folate concentrations (P: = 0.02285) were associated with a better relative response to treatment. Normalization of tHcy plasma levels was dependent on a lower baseline tHcy level (P: = 0.01976), younger age (P: = 0.00896), and MTHFR 677TT/1298AA or 677CT/1298AC genotypes (P: = 0.00208 and P: = 0.02320, respectively). A 4-week course of intravenous folinic acid is not superior to intravenous folic acid in reducing elevated tHcy plasma levels in hemodialysis patients. The response to treatment is predicted by tHcy plasma level, red blood cell folate content, and MTHFR genotype. PMID- 11273877 TI - Cefazolin dialytic clearance by high-efficiency and high-flux hemodialyzers. AB - Cefazolin dialytic clearance has not been determined in patients undergoing hemodialysis with high-efficiency or high-flux dialyzers. The objective of this study is to determine the pharmacokinetics and dialytic clearance of cefazolin and develop dosing strategies in these patients. Twenty-five uninfected subjects undergoing chronic thrice-weekly hemodialysis were administered a single dose of intravenous cefazolin (15 mg/kg) after their standard hemodialysis session. Fifteen subjects underwent hemodialysis with high-efficiency hemodialyzers, and 10 subjects underwent hemodialysis with high-flux hemodialyzers. Blood and urine samples were collected serially over the interdialytic period, during the next intradialytic period, and immediately after the next hemodialysis session. Serum and urine concentrations of cefazolin were determined by high-performance liquid chromatography. Differential equations describing a two-compartment model were fit to the cefazolin serum concentration-time data over the study period, and pharmacokinetic parameters were determined. Mean dialytic clearance values for cefazolin were significantly greater in the high-flux group compared with the high-efficiency group (30.9 +/- 6.52 versus 18.0 +/- 6.26 mL/min, respectively; P: < 0.05). Cefazolin reduction ratios were significantly greater (0.62 +/- 0.08 versus 0.50 +/- 0.07; P: < 0.005) in the high-flux group compared with the high efficiency group and correlated well with equilibrated urea reduction. The pharmacokinetic model developed from patient data was used to simulate cefazolin serum concentration data for high-efficiency and high-flux dialyzers. Cefazolin doses of 15 or 20 mg/kg after each hemodialysis session maintained adequate serum concentrations throughout a 2- or 3-day interdialytic period regardless of hemodialyzer type. PMID- 11273878 TI - The case for daily dialysis: its impact on costs and quality of life. AB - Research suggests daily hemodialysis may improve clinical outcomes. To date, a comprehensive review of its implications on quality of life has not been performed, and little is known about its economic impact. We conducted an economic evaluation comparing short daily or nocturnal hemodialysis with thrice weekly conventional in-center dialysis. Data on the quality of life and clinical effects of daily dialysis were obtained from more than 60 reports from 13 daily dialysis programs around the world (n = 197). Cost data were derived principally from the US Renal Data System, Centers for Disease Control, and Medicare Payment Advisory Commission. Resource use during daily hemodialysis was modeled after two ongoing programs in the United States. Results suggest that patients feel better and direct treatment costs could be reduced with daily dialysis. Costs are sensitive to assumptions about the effect of daily dialysis on hospital days. Reductions of at least 8% in hospital days are required for these modalities to be cost saving compared with documented reductions of 30% to 100%. Larger well controlled studies of daily versus conventional dialysis would be helpful to determine whether daily dialysis fulfills these promises. Medicare policy, which limits payment for most patients to three dialysis treatments weekly, poses a disincentive to more widespread adoption among dialysis centers. Given this constraint to broader acceptance, we address several policy options to gain a better understanding of the potential risks and benefits of daily dialysis. PMID- 11273879 TI - Hemodynamic reproducibility during blood flow measurements of hemodialysis synthetic grafts. AB - We have previously shown that graft blood flow (Qa) has a poor accuracy in predicting graft thrombosis. In this study, we determined whether hemodynamic variation helps explain this poor predictive accuracy. We also determined whether standardized timing of Qa measurements, which is widely recommended, will promote measurement reproducibility. We analyzed variations in mean arterial pressure (MAP) in seven consecutive dialysis sessions for 51 patients and determined the influence of MAP on Qa (by ultrasound dilution). We used a pooled coefficient of variation (CV) to summarize MAP variation within individual patients (computed as +/-2 CVs). MAPs from the seven sessions varied widely, and most variation was present with the first MAPs at the beginning of the sessions. These first MAPs varied by +/-23%, whereas variation for the entire session was +/-28%. The influence of MAP on Qa was determined by measuring the two together during consecutive thirds of a single session. The percentage of change in MAP (DeltaMAP) and Qa (DeltaQa) from the first to middle or last thirds of the session varied over wide ranges: -37% to 86% and -43% to 78%, respectively. The DeltaQa versus DeltaMAP correlation was relatively strong for changes between the first and middle thirds (r = 0.666) and first and last thirds (r = 0.646) of the session (both P: < 0.01). We conclude that MAP varies far more widely during dialysis than previously recognized. This variation is associated with large changes in Qa that may impair accuracy in predicting thrombosis. This wide MAP variation also indicates hemodynamic reproducibility is not feasible when measuring Qa. Thus, we do not recommend standardized timing of Qa measurements during dialysis. A practical method of addressing poor Qa reproducibility may be to take frequent measurements so that trends can be recognized before thrombosis occurs. PMID- 11273880 TI - Transcutaneous oxygen tension in patients with calciphylaxis. AB - Calciphylaxis is a severe complication of chronic renal failure, confined almost exclusively to patients on dialysis therapy. Histological characteristics of calciphylaxis include small-vessel calcifications of skin, subcutaneous tissue, and visceral organs. These vascular changes promote tissue ischemia that often results in tissue necrosis. In this study, we investigated the extent of skin ischemia in patients with calciphylaxis by means of transcutaneous oxygen tension (TCPO(2)) measurement, a noninvasive test that accurately assesses skin oxygenation. TCPO(2) levels were measured in 21 patients with calciphylaxis and 21 age- and sex-matched patients without evidence of calciphylaxis (controls). TCPO(2) levels were measured bilaterally at the chest, anterior abdomen, and upper thigh while patients breathed room air and after a 30-minute exposure to 100% fraction of inspired oxygen (FIO(2)). Compared with controls, patients with calciphylaxis showed significantly lower TCPO(2) levels at each body region. In both controls and patients with calciphylaxis, lower TCPO(2) levels correlated with increased weight and use of hemodialysis. No correlation with serum parathyroid hormone (PTH), serum calcium, or serum phosphorus values was present, although 39% of the patients with calciphylaxis had markedly elevated PTH values (sixfold greater than normal; >300 pg/dL). Low TCPO(2) levels in patients with calciphylaxis were documented in body regions with and without skin lesions. In patients with calciphylaxis, extremely low TCPO(2) values ( 200 mCi/m(2)) are at high risk to develop severe renal failure caused by TMA lesions. The histopathologic lesions are identical to those found after external radiotherapy, which suggests a causal relationship between (90)Y-DOTATOC and renal TMA. PMID- 11273888 TI - Should we still use iron dextran in hemodialysis patients? PMID- 11273889 TI - If daily dialysis is the answer, what is the question? PMID- 11273887 TI - Von Hippel-Lindau disease masquerading as autosomal dominant polycystic kidney disease. AB - The diagnostic confusion in differentiating the various causes of renal cystic diseases in adults is well documented. This confusion can include misclassifications between autosomal dominant polycystic kidney disease (ADPKD) and von Hippel-Lindau disease (VHL). We describe such a case of VHL. A review of the literature and of the patients in our database regarding typical features of each disease, mean age of onset, and frequency of these features was undertaken to provide helpful differentiating features. Pancreatic cysts are one differentiating feature. In VHL, pancreatic cysts can occur in 70% of patients, often are multiple, and rarely may cause exocrine or endocrine insufficiency. Pancreatic islet cell tumors occur. In ADPKD, pancreatic cysts are found in only 9% of patients, usually are single and asymptomatic, generally occur in conjunction with cystic liver disease, and are not found in children or unaffected family members. Pancreatic malignancies do not occur with increased frequency in ADPKD. A different pattern, especially in patients without a strong family history of ADPKD, may be a clue to VHL masquerading as ADPKD. Genetic mutation screening of the VHL gene should be used in these patients. PMID- 11273890 TI - Can ACE inhibitors prevent chronic allograft failure? PMID- 11273891 TI - A 26-year-old man with kidney allograft failure and foot pain. PMID- 11273892 TI - Emergence of interventional nephrology as a new subspecialty. PMID- 11273893 TI - Creatinine clearance overestimated glomerular filtration rate in a heavy tea drinker. PMID- 11273894 TI - 7-day ambulatory monitoring for adults with hypertension and diabetes. PMID- 11273897 TI - Lupus nephritis in a child with AIDS. AB - Concomitant acquired immunodeficiency syndrome (AIDS) and lupus nephritis is an exceptional feature in white patients. A white boy with maternofetal human immunodeficiency virus (HIV) infection had no medical follow-up until he presented at 12 years of age with a nephrotic syndrome, macrohematuria, renal failure, pancytopenia, and low CD4(+) cell count. A renal biopsy revealed severe lupus nephritis (World Health Organization class IV) with specific immune deposits in the absence of any clinical sign of systemic lupus erythematosus or specific autoantibodies at the time of diagnosis. The treatment consisted of methylprednisolone pulses followed by oral prednisone; antiretroviral triple therapy was started a few weeks later, which contributed to clinical and biologic improvement. To our knowledge, this is the first case report of lupus-like nephritis in a white child with AIDS, whose outcome might be improved significantly by a combination of steroids and antiretroviral therapy. PMID- 11273898 TI - Oncogenic Cushing's syndrome and nephrotic syndrome in the same patient. AB - The association of the nephrotic syndrome with malignancy is well established. This case report describes a patient who presented with renal cell carcinoma and went on to develop ectopic adrenocorticotropic hormone (ACTH) syndrome as well as the nephrotic syndrome. The patient's presentation was with hypokalemia refractory to conventional medical therapy. Oncogenic Cushing's syndrome and minimal change disease were diagnosed. We discuss the differentiation of Cushing's disease and Cushing's syndrome as well as oncogenic nephrotic syndrome and therapeutic options. PMID- 11273899 TI - Reversible posterior leukoencephalopathy syndrome in hepatitis C virus-positive long-term hemodialysis patients. AB - Chronic hepatitis C virus (HCV) infection is quite prevalent in long-term hemodialysis (HD) patients. Patients who are candidates for renal transplantation might be treated, before grafting, with interferon-alpha (IFN-alpha). Among 39 HCV-positive long-term HD patients treated with IFN-alpha, we observed three cases of reversible posterior leukoencephalopathy syndrome (PLES). PLES included headaches in three patients, confusion in three patients, cortical blindness in two patients, visual hallucinations in one patient, seizures in three patients, and respiratory distress in one patient in a context of fluid overload and severe hypertension in all cases. The three patients were receiving IFN-alpha and recombinant erythropoietin therapies simultaneously for de novo anemia. Contrast enhanced computed tomography scan or magnetic resonance imaging showed low density areas in the occipital lobes (in three patients), frontal lobes (in one patient), and temporal lobes (in one patient). After withdrawal of IFN-alpha and recombinant erythropoietin therapies, hemodiafiltration, and symptomatic treatment of seizures and hypertension, PLES was reversible within 1 week in one patient, 10 days in one patient, and 2 months in the third patient. Our case reports show the occurrence of reversible PLES in HCV-positive long-term HD patients treated with IFN-alpha. Physicians caring for HCV-positive long-term HD patients treated with IFN-alpha need to be particularly cautious when these patients receive simultaneously recombinant erythropoietin and when IFN-alpha therapy induces a weight loss, which indicates a reduction in dry weight. PMID- 11273900 TI - Reversible posterior leukoencephalopathy in a patient with minimal-change nephrotic syndrome. AB - A 9-year-old boy with nephrotic syndrome was transferred to our hospital because of acute renal failure and disturbance of consciousness after high-dose methylprednisolone therapy. He developed severe headache, visual disturbance, and generalized seizures. Brain computed tomography (CT) scan revealed multiple, bilateral, low-density areas in the parieto-occipital lobes. Magnetic resonance imaging (MRI) disclosed a high signal intensity area on T2-weighted images and a low signal intensity area on T1-weighted images in the same lesion. Follow-up brain CT scan and MRI, 2 weeks after the first studies, showed complete resolution of the abnormal lesions, which suggested the diagnosis of reversible posterior leukoencephalopathy syndrome (RPLS). Hypertension and high-dose methylprednisolone administration to the patient in the nephrotic state may be causes of this uncommon syndrome in this case. This is the first report of RPLS in nephrotic syndrome with hypertension not associated with cyclosporine administration. PMID- 11273902 TI - Pseudohypertension in a patient with diffuse scleroderma. AB - Pseudohypertension is the artifactual elevation of blood pressure that occurs secondary to noncompressible blood vessels. It has been described in patients with uremia, diabetes mellitus, and severe atherosclerosis. If unrecognized, the condition may lead to inappropriate and potentially harmful therapy. We report a case of pseudohypertension in a 65-year-old man with diffuse scleroderma. His blood pressure as assessed by conventional sphygmomanometry was at least 240/135 to 145 mm Hg. Intra-arterial blood pressure was found to be 107/52 mm Hg. The severe rise in blood pressure as measured by sphygmomanometry led to the concern of scleroderma renal crisis and potentially harmful therapy. Intra-arterial pressure monitoring confirmed the presence of pseudohypertension, however. This is the first reported case of pseudohypertension in a patient with diffuse scleroderma. PMID- 11273901 TI - Extreme metabolic alkalosis treated with normal bicarbonate hemodialysis. AB - Metabolic alkalosis (MA), defined as a primary increment in plasma bicarbonate concentration, is a common complication in hospitalized patients and is associated with high morbidity and mortality in severe cases. One of the major routes of compensation for MA (ie, the secretion of an alkaline urine) is lost in renal failure patients. We report three cases involving four episodes of extreme MA with an arterial pH value greater than 7.60, serum bicarbonate concentration greater than 55 mmol/L, and stupor or seizure. Profound vomiting or massive gastric drainage combined with concurrent oliguric renal failure was the underlying mechanism for severe MA. Hydration and normal central venous pressure failed to improve the MA. The extreme MA was reversed quickly and safely by conventional hemodialysis with normal bicarbonate dialysate of 25 to 28 mmol/L. To our knowledge, this is the first reported successful use of normal bicarbonate dialysate in the treatment of severe MA. We also found that either H(2) blockers or proton-pump inhibitors have a prophylactic effect on the formation of MA. PMID- 11273903 TI - Broken kidney: traumatic fracture of a renal allograft. AB - Although many renal transplant recipients are protective of their grafts and often express concern about the possibility of traumatic injury, such injury has been reported only rarely. We describe a case report of a boy who sustained an injury to his renal allograft after a bicycle accident. Despite a dramatic anatomic abnormality seen on magnetic resonance imaging, he suffered only minor and self-limited symptomatic and clinical consequences. We reviewed the literature addressing traumatic injuries affecting renal allografts. The long term consequences of such injuries on allograft function and survival are unknown. PMID- 11273904 TI - In search of a reliable awareness monitor. PMID- 11273905 TI - Investigations of the bispectral index monitor in pediatric anesthesia: first things first. PMID- 11273906 TI - Explicit intraoperative recall at a Bispectral Index of 47. PMID- 11273907 TI - The effect of insulin cardioplegia on atrial fibrillation after high-risk coronary bypass surgery: a double-blinded, randomized, controlled trial. AB - Atrial fibrillation after coronary bypass (CABG) surgery is an important cause of morbidity and increased resource utilization. Insulin-enhanced cardioplegia may reduce postoperative arrhythmias by improving aerobic myocardial metabolism and mitigating the deleterious effects of ischemia. We performed a double-blinded, randomized, controlled clinical trial to determine if insulin-enhanced cardioplegia decreases the risk of post-CABG atrial fibrillation in a high-risk patient population. We randomized 501 patients undergoing urgent CABG to receive insulin-enhanced (Humulin R 10 IU/L, Insulin group, n = 243) or standard (Control group, n = 258) blood cardioplegia during cardiopulmonary bypass. Patients were monitored by using continuous electrocardiography for a minimum of 3 days postoperatively. All standard cardiac medications, including beta-adrenergic blockers, were continued postoperatively. Insulin-enhanced cardioplegia did not result in a significant reduction in postoperative atrial fibrillation. Furthermore, we failed to detect a difference in the incidence of conduction defects, ventricular tachycardia, or pacemaker requirements between insulin and placebo patients. Atrial fibrillation was the most common arrhythmia, occurring in 31% of all patients. Independent predictors of atrial fibrillation were elderly age, preoperative atrial fibrillation, and renal insufficiency. Right bundle branch block was the most common conduction abnormality. Predictors of right bundle branch block were elderly age, female sex, and circumflex coronary artery disease. The incidence of postoperative ventricular tachycardia, left bundle branch block, and permanent pacemaker requirement was small. We conclude that insulin-enhanced cardioplegia does not reduce the incidence of postoperative atrial fibrillation in high-risk CABG patients. IMPLICATIONS: We conducted a double-blinded, randomized, placebo-controlled trial of insulin-enhanced cardioplegia in 501 patients undergoing urgent coronary bypass surgery. Insulin did not decrease the incidence of postoperative atrial fibrillation when compared with placebo. We also failed to demonstrate a difference in the incidence of other postoperative arrhythmias between the two groups of patients. PMID- 11273908 TI - Extracellular calcium modulates the effects of protamine on rat myocardium. AB - We studied the effects of protamine (10-300 microg. mL(-1)) as well as its interaction with heparin in rat left ventricular papillary muscles in vitro at calcium concentrations of 0.5 and 1 mM under low (isotony) and high (isometry) loads. Protamine induced a negative inotropic effect that was less pronounced at calcium 0.5 mM (active force at protamine 300 microg/mL, 84 +/- 20 vs 57 +/- 15% of baseline, P: < 0.05); whereas at calcium 1 mM there was a marked contracture of the muscle. For the smallest concentrations of protamine and at calcium 0.5 mM, we observed a moderate positive inotropic effect that was suppressed by nifedipine. Protamine induced a negative lusitropic effect under low load and decreased postrest potentiation, suggesting an impairment in the functions of the sarcoplasmic reticulum. Heparin was able to inhibit and reverse the negative inotropic effect of protamine. The negative inotropic effect of protamine is enhanced by an increase in extracellular calcium concentration. This negative inotropic effect is probably related to calcium overload and impairment in sarcoplasmic reticulum functions, and heparin can block these effects. IMPLICATIONS: The negative inotropic effect of protamine is enhanced by an increase in extracellular calcium concentration. This negative inotropic effect is probably related to calcium overload and impairment in sarcoplasmic reticulum functions, and heparin can block these effects. PMID- 11273909 TI - Apolipoprotein E polymorphisms and age at first coronary artery bypass graft. AB - Apolipoprotein E (apoE) polymorphisms are heritable determinants of total and low density lipoprotein cholesterol. The impact of apoE4 genotypes on the severity of atherosclerosis has been debated; however, recent studies have identified a correlation between apoE4 genotype and atherosclerosis. We assessed the impact of apoE4 genotype on age at first coronary artery bypass graft (CABG), hypothesizing that patients with the apoE4 allele are predisposed to coronary artery disease and present earlier for coronary revascularization. We assessed individual apoE genotypes and age in 560 patients undergoing primary CABG, by using analysis of variance (ANOVA) and controlling for gender. Because of the small number of patients in individual genotype groups, we compared patients with one or more copies of the apoE4 allele with those having no copies of the allele, again controlling for gender. A comparison of patients with one or more copies of the apoE4 allele with patients without the allele showed an earlier age at first CABG for those with the allele (P: = 0.032). Gene-dose analysis was also significant (P: = 0.012); patients with two copies of the allele presented at 54.2 +/- 6.9 yr. We report that the apoE4 allele is linked to age at first CABG. Identifying at-risk individuals may help prevent atherosclerosis. Further study is needed to define the mechanism of this association, and to define which coronary intervention is appropriate, based on long-term outcome. IMPLICATIONS: A correlation exists between apolipoprotein E (apoE) genotypes and the severity of atherosclerosis. We hypothesized that patients with the apoE4 allele are predisposed to coronary artery disease and present earlier for coronary artery bypass graft (CABG). Individuals with the apoE4 allele presented earlier for CABG, and the apoE4 allele is linked to age at first CABG. PMID- 11273911 TI - The relationship between plasma concentration of mature adrenomedullin and jugular venous oxygen saturation during and after cardiopulmonary bypass. AB - Adrenomedullin (AM), a vasodilatory peptide, increases during cardiac surgery. However, the physiological role of AM during cardiac surgery remains unclear. AM dilates cerebral arterioles and increases cerebral blood flow in rats. Therefore, we investigated whether AM is related to cerebral oxygen balance during cardiac surgery. In nine patients undergoing coronary artery bypass grafts, plasma concentrations of mature AM from the radial artery (mAMa) and jugular bulb (mAMj) were measured, and jugular venous oxygen saturation was obtained before surgery (baseline), before aortic cross-clamp (preclamp), after aortic declamp (postclamp), and 20 min after weaning from the cardiopulmonary bypass (post-CPB). Plasma concentrations of mAMa and mAMj were significantly increased at postclamp (P < 0.01 for both) and post-CPB (P < 0.01 for both) compared with baseline values. SjO(2) correlated with plasma mAMj concentrations at preclamp (r = 0.79, P < 0.01), postclamp (r = 0.71, P < 0.05), and post-CPB (r = 0.72, P < 0.05), as well as with mAMa concentrations at preclamp (r = 0.79, P < 0.01) and postclamp (r = 0.72, P < 0.05). This suggests a relationship between AM and cerebral oxygen balance during cardiac surgery. IMPLICATIONS: Plasma concentrations of mature form adrenomedullin, a vasodilatory peptide, was correlated with jugular venous oxygen saturation during cardiac surgery. This suggests a relationship between adrenomedullin and cerebral oxygen balance during cardiac surgery. PMID- 11273910 TI - Is perioperative plasma aprotinin concentration more predictable and constant after a weight-related dose regimen? AB - To determine whether a weight-related dose had advantages over a fixed, large dose regimen, we measured plasma concentrations of aprotinin by using an enzyme linked immunosorbent assay method at set time points in 30 patients having heart surgery with cardiopulmonary bypass. A weight-related dose comprising a preincision bolus injection of 40,000 kallikrein-inhibiting units (KIU)/kg (5.6 mg/kg) with the same amount given in the oxygenator prime was compared with a large-dose regimen of 2 x 10(6) KIU (280 mg) preincision bolus and addition to prime, together with an infusion of 500,000 KIU/h (70 mg/h). Peak plasma concentration in the Weight-Related group was less variable than with the fixed dose regimen. Forty percent of patients allocated to the fixed-dose regimen had an aprotinin concentration of more than 400 KIU/mL, compared with none in the Weight-Related group; this suggests a relative overdosing in the early surgical period in the Fixed-Dose group. There was great individual variability between patients in the time-concentration curves for aprotinin, with no difference between the two regimens. The weight-related dose regimen benefited by not requiring an intraoperative infusion while achieving the same plasma concentrations of aprotinin. IMPLICATIONS: Peak plasma concentrations of aprotinin were less variable with a weight-related dose schedule. This has implications for safety with regard to control of anticoagulation and cost in patients with small body mass. Plasma concentrations varied greatly with time between patients. This observation has implications for determining an optimal dose on the basis of aprotinin's currently known mechanisms of action. PMID- 11273912 TI - Oxygenation during one-lung ventilation: the effects of inhaled nitric oxide and increasing levels of inspired fraction of oxygen. AB - We studied whether inhaled nitric oxide (NO) would improve arterial oxygen tension (PaO(2)) and reduce the occurrence of oxygen saturation of hemoglobin (O(2)Hb) < 90% during one-lung ventilation (OLV). One-hundred-fifty-two patients were ventilated either with or without NO (20 ppm) with an inspired fraction of oxygen (FIO(2)) of either 0.3, 0.5, or 1.0 during OLV. Anesthesia was induced and maintained with propofol, remifentanil, and rocuronium IV, and lung separation was achieved with a double-lumen tube. During OLV, we set positive end-expiratory pressure at 5 cm H(2)O, peak pressure at 30 cm H(2)O, and end-tidal CO(2) at 30 mm Hg. The nonventilated lung was opened to room air and collapsed. During OLV, three consecutive measurements were performed every 10 min. The operated lung was temporarily ventilated if pulse oximetric saturation (SpO(2)) decreased to < 91%. SpO(2) <9 1% occurred in 2 of the 152 patients. SpO(2) overestimated O(2)Hb by 2.9% +/- 0.1%. NO failed to improve oxygenation or alter occurrence of O(2)Hb < 90% during OLV across all time points and all levels of FIO(2). Increasing FIO(2) increased oxygenation and decreased occurrence of O(2)Hb < 90% (P: < 0.001). At FIO(2) = 1, PaO(2) was higher (P < 0.01) and O(2)Hb < 90% rate tended to be lower (P = 0.1) during right versus left lung ventilation. PaO(2) was higher in patients undergoing pneumonectomy and lobectomy than in those undergoing metastasectomy or video-assisted operations (P < 0.05). IMPLICATIONS: Inhaled nitric oxide failed to improve oxygenation during one-lung ventilation. Oxygenation during one-lung ventilation was improved with increasing levels of FIO(2) during ventilation of the right versus the left lung and with increasing pathology of the nonventilated lung. PMID- 11273913 TI - Thoracic epidural anesthesia combined with general anesthesia: the preferred anesthetic technique for thoracic surgery. AB - Thoracic epidural anesthesia (TEA) combined with general anesthesia (GA) as well as total-IV anesthesia (TIVA) are both established anesthetic managements for thoracic surgery. We compared them with respect to hypoxic pulmonary vasoconstriction, shunt fraction and oxygenation during one-lung ventilation. Fifty patients, ASA physical status II-III undergoing pulmonary resection were randomly allocated to two groups. In the TIVA group, anesthesia was maintained with propofol and fentanyl. In the TEA group, anesthesia was maintained with TEA (bupivacaine 0.5%) combined with low-dose concentration 0.3-0.5 vol% of isoflurane (end-tidal). Changing from two-lung ventilation to one-lung ventilation caused a significant increase in cardiac output (CO) in the TIVA group, whereas no change was observed in the TEA group. One-lung ventilation caused significant increases in shunt fraction in both groups which was associated per definition with a significant decrease in PaO(2) in both groups but PaO(2) remained significantly increased in the TEA group (P < 0.05). We conclude that both anesthetic regimens are safe intraoperatively. However, TEA in combination with GA did not impair arterial oxygenation to the same extent as TIVA, which might be a result of the changes in CO. Therefore, patients with preexisting cardiopulmonary disease and impaired oxygenation before one-lung ventilation might benefit from TEA combined with GA. IMPLICATIONS: Fifty patients underwent lung surgery through the opened chest wall requiring ventilation of only one lung. Patients were randomly assigned to receive either general anesthesia alone or in combination with regional anesthesia via a catheter in the back. Oxygen content in the blood and blood pressure was better maintained in the group receiving the combination of general with regional anesthesia. PMID- 11273915 TI - Co-oximetry interference by hemoglobin-based blood substitutes. AB - The blood substitutes now being developed from molecularly modified hemoglobin interfere with a wide variety of clinical analyzers, but their effects on cooximeters are unknown. Therefore, we investigated the effects of five hemoglobin-based blood substitutes on the measurements of eight different oximeters and cooximeters: the AVL Omni 6, the AVOXimeters 1000 and 4000, the Ciba Corning (now Bayer) CC270 CO-Oximeter, the Instrumentation Laboratory Synthesis 35, the IL482 and IL682 CO-Oximeters, and the Radiometer OSM3 Hemoximeter. The five blood substitutes in this study were obtained from Apex Bioscience (Research Triangle Park, NC), Baxter Healthcare Corp. (Deerfield, IL), Biopure Corp. (Cambridge, MA), Hemoglobin Therapeutics, and Hemosol, Inc. (Etobicoke, Ontario, Canada). A cooximeter control was used to compare the eight different instruments' measurements on unaltered human hemoglobin. The instruments yielded measurements of total hemoglobin concentration in undiluted blood substitutes that were generally not more variable than those on the control material. By contrast, when compared with readings on controls, the test instruments yielded measurements of the fractional concentrations of oxy-, deoxy , carboxy-, and methemoglobin that showed greater instrument-to-instrument disparities and larger standard deviations about the all-instrument means. In some cases, the interference was even more obvious: five of six cooximeters gave negative carboxyhemoglobin readings on one particular product. Our findings indicate that the instruments will give less accurate but clinically useful measurements in the presence of these hemoglobin-based blood substitutes. IMPLICATIONS: We investigated the effects of five hemoglobin-based blood substitutes on the measurements of eight different cooximeters. Some blood substitutes caused obvious interference, such as negative carboxyhemoglobin readings; however, the findings indicate that cooximeters will generally give less accurate but clinically useful measurements in the presence of the hemoglobin-based blood substitutes that were tested. PMID- 11273916 TI - Unsuccessful placement of transesophageal echocardiography probe because of esophageal pathology. PMID- 11273914 TI - Voluven, a lower substituted novel hydroxyethyl starch (HES 130/0.4), causes fewer effects on coagulation in major orthopedic surgery than HES 200/0.5. AB - Hydroxyethyl starch (HES) solutions are effective plasma volume expanders. Impairment of coagulation occurs with large HES volumes infused perioperatively. Therefore, a lower substituted novel HES (Voluven; Fresenius Kabi, Bad Homburg, Germany) was developed to minimize hemostatic interactions, and was compared with HAES-steril (Fresenius Kabi) (pentastarch) regarding safety and efficacy. We performed a prospective, randomized, double-blinded study in 100 major orthopedic surgery patients. Because the 95% confidence interval (-330 mL; +284 mL) for the treatment contrast Voluven-HAES-steril was entirely included in the predefined equivalence range (+/- 500 mL), comparable efficacy was established. Voluven interfered significantly less than HAES-steril with coagulation factor VIII levels and partial thromboplastin time postoperatively. Total amounts of red blood cells transfused were comparable between the Voluven and HAES-steril groups, but a significantly reduced need for homologous red blood cells was observed in the Voluven group. We conclude that in large-blood-loss surgery, Voluven has a comparable efficacy with HAES-steril and may reduce coagulation impairment, possibly leading to a smaller number of allogeneic blood transfusions. IMPLICATIONS: Hydroxyethyl starches are common plasma volume expanders, but may interfere with coagulation at large doses. We tested a novel hydroxyethyl starch specification (Voluven; Fresenius Kabi, Bad Homburg, Germany) which was developed to reduce hemostatic interactions while preserving its efficacy in restoring plasma volume in comparison to HAES-steril (pentastarch; Fresenius Kabi) in major orthopedic surgery. PMID- 11273917 TI - Life-threatening hypoxemic respiratory failure after repair of acute type a aortic dissection: successful treatment with venoarterial extracorporeal life support using a prosthetic graft attached to the right axillary artery. PMID- 11273918 TI - The effect of bispectral index monitoring on anesthetic use and recovery in children anesthetized with sevoflurane in nitrous oxide. AB - The utility of bispectral index (BIS) monitoring to guide anesthetic administration has been demonstrated in adults. This prospective, randomized observer-blinded study was designed to evaluate the effect of BIS monitoring on anesthetic use and recovery characteristics in pediatric patients. After data collection in 38 historical controls, 202 patients age 0-18 yr were randomized into one of two groups: standard practice (SP) and BIS guided (BIS). Patients age 0-3 yr undergoing inguinal hernia repair (IH) and patients age 3-18 yr undergoing tonsillectomy and/or adenoidectomy (TA) were selected. All patients were anesthetized with sevoflurane in 60% N(2)O/O(2). Hernia patients also received a caudal epidural anesthetic before surgery. In the BIS group, anesthetic delivery was adjusted in an effort to achieve a target BIS of 45-60 during maintenance and 60-70 during the last 15 min of the procedure. BIS was recorded throughout surgery in all patients, but data were unavailable to the anesthesiologist in the SP group. In the TA patients, BIS monitoring was associated with a significant reduction in end-tidal sevoflurane concentration during maintenance (2.4 +/- 0.6%, SP and 1.8 +/- 0.4% BIS, mean +/- SD) and during the last 15 min of the procedure (2.1 +/- 0.7, SP and 1.6 +/- 0.6, BIS). There was a 25%-40% decrease in measured recovery times. In the patients 0-6 mo of age undergoing IH, sevoflurane concentrations during maintenance (2.0 +/- 0.4% SP, 0.9 +/- 0.8 BIS), during the last 15 min (1.6 +/- 0.4% SP, 0.6 +/- 0.6% BIS), and at the end of the procedure (1.1 +/- 0.6% SP, 0.3 +/- 0.3% BIS) were smaller in the BIS group. Emergence and recovery measures were unaffected by BIS titration. In the children 6 mo-3 yr of age, there were no significant differences between the SP and BIS groups in anesthetic use or recovery measures. IMPLICATIONS: Bispectral index monitoring in children results in less anesthetic use and faster recovery than standard practice. PMID- 11273919 TI - Stress response in infants undergoing cardiac surgery: a randomized study of fentanyl bolus, fentanyl infusion, and fentanyl-midazolam infusion. AB - There have been significant changes in the management of neonates and infants undergoing cardiac surgery in the past decade. We have evaluated in this prospective, randomized, double-blinded study the effect of large-dose fentanyl anesthesia, with or without midazolam, on stress responses and outcome. Forty five patients < 6 mo of age received bolus fentanyl (Group 1), fentanyl by continuous infusion (Group 2), or fentanyl-midazolam infusion (Group 3). Epinephrine, norepinephrine, cortisol, adrenocortical hormone, glucose, and lactate were measured after the induction (T1), after sternotomy (T2), 15 min after initiating cardiopulmonary bypass (T3), at the end of surgery (T4), and after 24 h in the intensive care unit (T5). Plasma fentanyl concentrations were obtained at all time points except at T5. Within each group epinephrine, norepinephrine, cortisol, glucose and lactate levels were significantly larger at T4 (P values < 0.01), but there were no differences among groups. Within groups, fentanyl levels were significantly larger in Groups 2 and 3 (P < 0.001) at T4, and among groups, the fentanyl level was larger only at T2 in Group 1 compared with Groups 2 and 3 (P < 0.006). There were no deaths or postoperative complications, and no significant differences in duration of mechanical ventilation or intensive care unit or hospital stay. Fentanyl dosing strategies, with or without midazolam, do not prevent a hormonal or metabolic stress response in infants undergoing cardiac surgery. IMPLICATIONS: We demonstrated a significant endocrine stress response in infants with well compensated congenital cardiac disease undergoing cardiac surgery, but without adverse postoperative outcome. The use of large-dose fentanyl, with or without midazolam, with the intention of providing "stress free" anesthesia, does not appear to be an important determinant of early postoperative outcome. PMID- 11273920 TI - The effects of sevoflurane and halothane anesthesia on cerebral blood flow velocity in children. AB - We compared cerebral blood flow velocity during anesthesia with sevoflurane and halothane in 23 children admitted for elective surgery (age, 0.4-9.7 yr; median age, 1.9 yr; ASA physical status I-II). Inhaled induction was performed in a randomized sequence with sevoflurane or halothane. Under steady-state conditions, cerebral blood flow velocity (systolic [V(s)], mean [V(mn)], and diastolic [VD]) were measured by a blinded investigator using transcranial pulsed Doppler ultrasonography. The anesthetic was then changed. CBFV measurements were repeated after washout of the first anesthetic and after steady-state of the second (equivalent minimal alveolar concentration to first anesthetic). The resistance index was calculated. VD and V(mn) were significantly lower during sevoflurane (V(mn) 1.35 m/s) than during halothane (V(mn) 1.50 m/s; P = 0.001), whereas V(s) was unchanged. The resistance index was lower during halothane (P < 0.001). Our results indicate lower vessel resistance and higher mean velocity during halothane than during sevoflurane. IMPLICATIONS: The mean cerebral blood flow velocity is significantly decreased in children during inhaled anesthesia with sevoflurane than during halothane. This might be relevant for the choice of anesthetic in children with risk of increased intracranial pressure, neurosurgery, or craniofacial osteotomies. PMID- 11273921 TI - Sensory stimuli and anxiety in children undergoing surgery: a randomized, controlled trial. AB - We assessed the effectiveness of a behavioral intervention aimed at reducing the anxiety of children undergoing anesthesia and surgery. The intervention consisted of dimmed operating room (OR) lights (200 Lx) and soft background music (Bach's "Air on a G String," 50-60 dB). Only one person, the attending anesthesiologist, interacted with the child during the induction of anesthesia. Children undergoing anesthesia and surgery were randomly assigned either to a low sensory stimulation group (LSSG, n = 33) or to control group (n = 37). By using validated behavioral measures of anxiety (mYPAS) and compliance (ICC), children were evaluated at the preoperative holding area and during the induction of anesthesia. On postoperative Days 1, 2, 3, 7, and 14, the behavioral recovery of the children was assessed by using the Post Hospitalization Behavior Questionnaire. We found that the LSSG was significantly less anxious compared with the control group on entrance to the OR (P = 0.03) and on the introduction of the anesthesia mask (P = 0.003). Also, the compliance during the induction of anesthesia was significantly better in children assigned to the LSSG (P = 0.02). The incidence of postoperative behavioral changes, however, did not differ significantly between the two groups (P = ns). We conclude that children who are exposed to low-level sensory stimuli during the induction of anesthesia and who are exposed to background music exhibit lower levels of anxiety and increased compliance. IMPLICATIONS: Children are less anxious and show increased compliance during induction when exposed to a single care-provider in a dimmed, quiet operating room with background music. PMID- 11273922 TI - Sonoclot analysis in healthy children. AB - Although use of the Sonoclot device (Sienco, Inc., Morrison, CO) has been reported in isolated pediatric cases and in small reports in neonates, there are no published data for normal pediatric patients. As the device is used in situations of abnormal coagulation, such as cardiac and liver transplantation surgery, our aim was to determine normal data ranges in healthy pediatric surgical patients. Blood was withdrawn after anesthetic induction, and the Sonoclot activated clotting time, rate of clot formation, time to peak amplitude, and peak amplitude was compared among four pediatric groups (< 12 mo, 13-24 mo, 25-48 mo, 49 mo-9 yr) and an adult group. The Sonoclot activated clotting time in the < 12-mo and the Adult groups were shorter than the oldest group of children (P < 0.05), although all were within the anticipated normal range, and there were no significant differences in clot rate, peak amplitude, and time to peak amplitude among groups without apparent trends with increasing age. These Sonoclot variables quantify adequate global clot formation in pediatric patients and will facilitate clinical coagulation management with appropriate pediatric normal ranges, avoiding the application of extrapolated adult data to children. IMPLICATIONS: Sonoclot variables are presented for 95 healthy pediatric surgical patients in four age groups, with small differences found in the Sonoclot (Sienco, Inc., Morrison, CO) activated clotting time between two groups and no significant differences in three other variables among groups. PMID- 11273923 TI - Prophylactically-administered rectal acetaminophen does not reduce postoperative opioid requirements in infants and small children undergoing elective cleft palate repair. AB - Rectal acetaminophen (Ac) is often administered prophylactically at anesthesia induction for postoperative pain management in small children and is thought to have an opioid-sparing effect. We assessed in this double-blinded, prospective, randomized study early opioid requirements after three doses of Ac (10, 20, and 40 mg/kg versus placebo) in 80 children (ASA physical status I, age 11.4 +/- 9.9 mo) undergoing cleft palate repair. Single Ac plasma concentrations were measured. Pain scores assessed in the postanesthesia care unit of > or = 4 of 10 resulted in the IV administration of 25 microg/kg piritramide, a popular European mu receptor agonist (lockout time, 10 min; maximum 0.125 mg/kg). There were no significant differences between groups with regard to the early postoperative pain scores and the overall cumulative IV opioid requirements. Maximal plasma concentrations achieved were only subtherapeutic (Ac 10 mg/kg: 8 microg/mL; Ac 20 mg/kg: 13 microg/mL; Ac 40 mg/kg: 21 microg/mL after 122, 122, and 121 min, respectively). We conclude that rectal Ac up to 40 mg/kg has no opioid-sparing effect, does not result in analgesic Ac plasma concentrations, and lacks proof of its efficacy in infants and small children undergoing cleft palate repair, whereas titrated IV opioid boluses produced rapid and reliable pain relief. IMPLICATIONS: Acetaminophen is widely used prophylactically for postoperative analgesia in children and is thought to have an opioid-sparing effect. We showed that rectal acetaminophen up to 40 mg/kg administered at anesthesia induction lacked proof of efficacy, whereas IV opioid boluses resulted in reliable pain relief in children undergoing cleft palate repair. PMID- 11273924 TI - A failure of the chain-link mechanism on the Ohmeda Excel 210 anesthetic machine. PMID- 11273925 TI - Increased T-wave amplitude after accidental intravascular injection of lidocaine plus bupivacaine without epinephrine in sevoflurane-anesthetized child. PMID- 11273927 TI - The preemptive analgesic effect of intraarticular bupivacaine and morphine after ambulatory arthroscopic knee surgery. AB - Intraarticular (IA) morphine provides effective postoperative analgesia after arthroscopic knee surgery. Some investigators have suggested that the preemptive administration of opioids may reduce postoperative analgesic requirements and hypersensitivity. We evaluated the analgesic effect of administering IA morphine either before or after surgical incision in patients undergoing arthroscopic knee surgery under local anesthesia. Forty patients undergoing arthroscopic meniscectomy were randomized into two groups. All patients received IA bupivacaine 0.25% before and after surgery together with IV sedation using midazolam and propofol. The Preemptive IA Morphine group received a single 3-mg dose of morphine with their preoperative bupivacaine. The Post-IA Morphine group received 3 mg of morphine at the completion of surgery with the postoperative bupivacaine. After surgery, pain scores, the time to first opioid use, and 24-h analgesic use were recorded. Analgesic duration, defined as the time from completion of surgery until first opioid use, was significantly longer in those patients receiving preoperative (953 +/- 209 min) versus postoperative (556 +/- 121 min) IA morphine. The 24-h acetaminophen and oxycodone use was less in the Preemptive group (2.2 +/- 1.2 pills) versus the Postoperative group (3.0 +/- 1.2 pills). We conclude that IA morphine provides a longer duration of postoperative analgesia with less 24-h opioid use when administered before surgery. IMPLICATIONS: The administration of intraarticular morphine 3 mg before arthroscopic knee surgery provides a longer duration of analgesia with less 24-h opioid use compared with the administration of the drug at the completion of surgery. PMID- 11273926 TI - Fast-tracking children after ambulatory surgery. AB - This study was designed to determine the feasibility and benefits of fast tracking children after ambulatory surgery. One-hundred-fifty-five healthy children undergoing surgical procedures lasting <90 min were studied in a randomized manner. After surgery, children who met predefined recovery criteria in the operating room were entered into one of the study groups. Seventy-one patients (control) were first admitted to the postanesthesia care unit (PACU) and then to the second-stage recovery unit (SSRU). Eighty-four children bypassed the PACU and were directly admitted to the SSRU (Fast-Track group). The demographic data, airway management, and surgical procedures were similar in both groups of patients. During the recovery phase, 62.0% of the PACU group patients and 40.5% of the Fast-Track patients received analgesics (P = 0.01). The total recovery time was 79.1 +/- 48.3 min in the Fast-Track group and 99.4 +/- 48.6 min in the Control group (P = 0.008). A larger percentage of parents in the Fast-Track group (31% vs 16%) reported that their child was restless on arrival at the SSRU (P = 0.037). There were no clinically significant adverse events. However, adequate pain control must be provided before transfer to SSRU. In conclusion, fast tracking children after ambulatory surgery is feasible and beneficial when specific selection criteria are used. IMPLICATIONS: The results of this study show that the total recovery time is shorter in children who are fast-tracked (bypass the postanesthesia care unit) after ambulatory surgery. A higher percentage of parents of the Fast-Track group felt that their child was restless on arrival at the second-stage recovery unit. Fast-tracking children after ambulatory surgery is feasible and beneficial when specific selection criteria are used. PMID- 11273928 TI - The in vitro effects of ketamine at large concentrations can be attributed to a nonspecific cytostatic effect. PMID- 11273929 TI - Thiopental is a competitive inhibitor at the human alpha7 nicotinic acetylcholine receptor. AB - The nicotinic acetylcholine receptors (nAChRs) in the central nervous system may be a potential target for the anesthetic effects of thiopental. We evaluated the mechanism of action of thiopental on the human alpha7 nAChR by using 2-electrode voltage clamp methodology. Concentration response curves for agonist were prepared in the presence of 25-250 microM of thiopental. Inhibition by the S- and R-thiopental enantiomers was compared with inhibition by racemic thiopental. We found that thiopental acts as a competitive inhibitor at the human alpha7 nAChR. Inhibition is independent of membrane potential and the K(i(apparent)) is 13 microM of thiopental. The clinical 50% effective concentration for thiopental in humans is 25 microM. Thus, with a K(i(apparent)) of 13 microM, inhibition of the human alpha7 nAChR is within a clinically relevant range. The S- and R enantiomers of thiopental cause inhibition indistinguishable from the inhibition caused by racemic thiopental. This discordance makes it unlikely that the alpha7 nAChR plays a role in loss of righting reflex induced by thiopental in mice, although nicotinic inhibition by thiopental may mediate other anesthetic effects and side effects. IMPLICATIONS: The receptors for nicotine in the brain may be involved in the mechanism of general anesthetics. We have shown that a human receptor for nicotine is inhibited by the anesthetic barbiturate thiopental, at concentrations used clinically. The nicotinic receptor thus may mediate some of the actions of this drug. PMID- 11273930 TI - The effects of propofol in the area postrema of rats. AB - Propofol has an antiemetic effect that may be mediated by gamma-aminobutyric acid (GABA) influences on the serotonin system, the mechanism of which is not known. We used three techniques, immunohistochemistry, High Performance Liquid Chromatography, and electrophysiology, to define propofol's effects on the rat's brainstem. Paired male Wistar rats received propofol, 20 mg/kg/hr, or Intralipid for 6 h. The brains were then subjected to immunohistochemical analysis of serotonin. In a separate experiment after a propofol or Intralipid infusion, cerebrospinal fluid (CSF) was extracted from the fourth ventricle and analyzed for the amount of serotonin and 5-hydroxyindoleacetic acid. Electrophysiological neuronal recordings were made in the area postrema (AP) in response to propofol with and without a GABA or serotonin antagonist. Results showed that immunohistochemical staining for serotonin in the propofol rats was significantly increased (28 +/- 12%) in the dorsal raphe and decreased in the AP (17 +/- 6%) compared with control. There were no significant changes in the isoflurane anesthetized animals. Both serotonin and 5-hydroxyindoleacetic acid in the CSF of the fourth ventricle at the level of the AP were significantly reduced by 63% and 36%, respectively. Both propofol and pentobarbital injections reduce AP neuronal activity, but only the propofol response was blocked by bicuculline, a GABA antagonist. We conclude that the reduced levels of serotonin in the AP and the CSF may explain the antiemetic property of propofol. Propofol may also directly act on AP neurons via a GABA(A) receptor to reduce their activity. IMPLICATIONS: Propofol may produce its antiemetic effect by depleting the area postrema of serotonin as well as by a direct gamma-aminobutyric acid-mediated inhibition. PMID- 11273931 TI - Scheduling a delay between different surgeons' cases in the same operating room on the same day using upper prediction bounds for case durations. AB - At some surgical suites, elective cases are only scheduled if they can be completed during regularly scheduled hours. At such a surgical suite, a surgeon may be scheduled to perform one or more cases in an operating room (OR), to be followed by another surgeon who will perform one or more cases. Scheduling a delay between the two surgeons' cases will improve the likelihood that the second surgeon's case(s) will start on time. We show that the mathematics of calculating a scheduled delay between the different surgeons' cases in the same OR on the same day is that of calculating an upper prediction bound for the duration of the second surgeon's case(s). We test an analytical expression for the upper prediction bound for the last one case of the day in an OR, and a Monte Carlo simulation method for the last two cases. We show that these 90% upper prediction bounds are at least as long as the actual durations for 90% +/- 0.2% of single cases and 92% +/- 0.6% of pairs of cases. We conclude that our methodology can be used to calculate an appropriate, and reasonably accurate, scheduled delay between two surgeons' cases in the same OR on the same day. IMPLICATIONS: We show how to use a statistical analysis of historical case duration data to calculate an appropriate and accurate scheduled delay between two surgeons' cases in the same operating room on the same day. PMID- 11273932 TI - Statistical analysis of postanesthesia care unit staffing at a surgical suite with frequent delays in admission from the operating room--a case study. PMID- 11273933 TI - Ventilatory support by continuous positive airway pressure breathing improves gas exchange as compared with partial ventilatory support with airway pressure release ventilation. AB - In acute lung injury, airway pressure release ventilation (APRV) with superimposed spontaneous breathing improves gas exchange compared with controlled mechanical ventilation. However, the release of airway pressure below the continuous positive airway pressure (CPAP) level may provoke lung collapse. Therefore, we compared gas exchange and hemodynamics using a crossover design in nine pigs with oleic acid-induced lung injury during CPAP breathing and APRV with a release pressure level of 0 and 5 cm H(2)O. At an identical minute ventilation (V(E) 8 L/min) spontaneous breathing averaged 55%, 67%, and 100% of V(E) during the two APRV modes and CPAP, respectively. Because of the concept of APRV, mean airway pressure was highest during CPAP and lowest during APRV with a release pressure of 0 cm H(2)O. Shunt was reduced to almost half during CPAP (6.6% of Q(t)) compared with both APRV-modes (13.0% of Q(t)). Cardiac output and oxygen consumption, in contrast, were similar during all three ventilatory settings. Thus, in our lung injury model, CPAP was superior to partial ventilatory support using APRV with and without positive end-expiratory pressure. This may be attributable to beneficial effects of spontaneous breathing on gas exchange as well as to rapid lung collapse during the phases of airway pressure release below the CPAP level. These findings may suggest that the amount of mechanical ventilatory support using the APRV mode should be kept at the necessary minimum. IMPLICATIONS: Oxygenation is better with continuous positive airway pressure breathing than with partial mechanical ventilatory support using airway pressure release ventilation. Therefore, mechanical ventilatory support achieved by a cyclic release of airway pressure during APRV should be kept at the minimum level that enables enough ventilatory support for patients to avoid respiratory muscle fatigue. PMID- 11273934 TI - ONO1714, a new inducible nitric oxide synthase inhibitor, attenuates sepsis induced diaphragmatic dysfunction in hamsters. AB - Sepsis causes impairment of diaphragmatic contractility and endurance capacity. Nitric oxide (NO) produced via inducible NO synthase (iNOS) has been implicated in the pathogenesis. Peroxynitrite, a NO-derived powerful oxidant, may be responsible for infection-induced diaphragmatic muscle failure. Therefore, we examined whether ONO1714, a new selective iNOS inhibitor, prevents sepsis-induced diaphragmatic dysfunction. Fifty male Golden-Syrian hamsters were randomly divided into five groups: hamsters that underwent sham laparotomy alone and received saline injection (Group Sham), those that underwent cecal ligation with puncture (CLP) and received saline injection (Group Sepsis), those that underwent sham laparotomy and received injection of ONO1714 0.3 mg/kg (Group Sham ONO1714high), those that underwent CLP and received ONO1714 0.1 mg/kg (Group Sepsis-ONO1714low), and those that underwent CLP and received ONO1714 0.3 mg/kg (Group Sepsis-ONO1714high). ONO1714 or saline was intraperitoneally injected 10 min before surgery. Diaphragmatic contractility was assessed in vitro using diaphragm muscle strips excised 24 h after operation. Diaphragm fatigability was assessed by time until tension decreased to 50% of the initial value (T50%) during fatigue trials. Twitch, tetanic tensions, and T50% during fatigue trials were reduced in Group Sepsis. Pretreatment with ONO1714 dose-dependently attenuated sepsis-induced diaphragmatic contractile profiles and endurance capacity. CLP increased plasma nitrite/nitrate (NOx; stable NO metabolites), and diaphragm malondialdehyde (MDA; a product of lipid peroxidation), positive immunostaining for nitrotyrosine (peroxynitrite footprint), and iNOS activity. ONO1714 attenuated the increase. This beneficial effect of ONO1714 may be attributable, in part, to inhibition of peroxynitrite-induced lipid peroxidation in the diaphragm. IMPLICATIONS: Sepsis impairs diaphragmatic contractility and endurance capacity, which may be involved in acute respiratory failure. Pretreatment with ONO1714, a new selective inducible nitric oxide synthase inhibitor, attenuated sepsis-induced diaphragmatic dysfunction in hamsters. PMID- 11273935 TI - The effects of positive end-expiratory pressure during active compression decompression cardiopulmonary resuscitation with the inspiratory threshold valve. AB - The use of an inspiratory impedance threshold valve (ITV) during active compression-decompression (ACD) cardiopulmonary resuscitation (CPR) improves perfusion pressures, and vital organ blood flow. We evaluated the effects of positive end-expiratory pressure (PEEP) on gas exchange, and coronary perfusion pressure gradients during ACD + ITV CPR in a porcine cardiac arrest model. All animals received pure oxygen intermittent positive pressure ventilation (IPPV) at a 5:1 compression-ventilation ratio during ACD + ITV CPR. After 8 min, pigs were randomized to further IPPV alone (n = 8), or IPPV with increasing levels of PEEP (n = 8) of 2.5, 5.0, 7.5, and 10 cm H(2)O for 4 consecutive min each, respectively. Mean +/- SEM arterial oxygen partial pressure decreased in the IPPV group from 150 +/- 30 at baseline after 8 min of CPR to 110 +/- 25 torr at 24 min, but increased in the PEEP group from 115 +/- 15 to 170 +/- 25 torr with increasing levels of PEEP (P <0.02 for comparisons within groups). Mean +/- SEM diastolic aortic minus diastolic left ventricular pressure gradient was significantly (P < 0.001) higher after the administration of PEEP (24 +/- 0 vs 17 +/- 1 mm Hg with 5 cm H(2)O of PEEP, and 26 +/- 0 vs 17 +/- 1 mm Hg with 10 cm H(2)O of PEEP), whereas the diastolic aortic minus right atrial pressure gradient (coronary perfusion pressure) was comparable between groups. Furthermore, systolic aortic pressures were significantly (P < 0.05) higher with 10 cm H(2)O of PEEP when compared with IPPV alone (68 +/- 0 vs 59 +/- 2 mm Hg). In conclusion, when CPR was performed with devices designed to improve venous return to the chest, increasing PEEP levels improved oxygenation. Moreover, PEEP significantly increased the diastolic aortic minus left ventricular gradient and did not affect the decompression phase aortic minus right atrial pressure gradient. These data suggest that PEEP reduces alveolar collapse during ACD + ITV CPR, thus leading to an increase in indirect myocardial compression. IMPLICATIONS: Inspiratory impedance during active compression-decompression cardiopulmonary resuscitation improves perfusion pressures, and vital organ blood flow during cardiac arrest. Increasing levels of positive end-expiratory pressure during performance of active compression-decompression cardiopulmonary resuscitation with an inspiratory impedance valve improves oxygenation, and increases the diastolic aortic-left ventricular pressure gradient and systolic arterial blood pressure. PMID- 11273936 TI - A lack of evidence of superiority of propofol versus midazolam for sedation in mechanically ventilated critically ill patients: a qualitative and quantitative systematic review. AB - Propofol and midazolam are often used for sedation in the intensive care unit. The aim of this systematic review was to estimate the efficacy and harm of propofol versus midazolam in mechanically ventilated patients. A systematic search (Medline, Cochrane Library, Embase, bibliographies), any language, up to June 1999 was performed for reports of randomized comparisons of propofol with midazolam. Data from 27 trials (1624 adults) were analyzed. The average duration of sedation varied between 4 and 339 h. In 10 trials, the duration of adequate sedation was longer with propofol (weighted mean difference 2.9 h; 95% confidence interval [CI], 0.2-5.6 h). In 13 trials (mostly postoperative), sedation lasted 4 to 35 h; in 9 of those, average weaning time from mechanical ventilation with propofol was 0.8-4.3 h; with midazolam it was 1.5-7.2 h (weighted mean difference 2.2 h [95% CI, 0.8 to 3.7 h]). In 8 trials, sedation lasted 54 to 339 h; there was a lack of evidence for difference in weaning times. Arterial hypotension (relative risk 2.5 [95% CI, 1.3 to 4.5]; number-needed-to-treat, 12), and hypertriglyceridemia (relative risk 12.1 [95%CI, 2.9 to 49.7]; number-needed-to treat, 6) occurred more often with propofol. The duration of adequate sedation time is longer with propofol compared with midazolam. In postoperative patients with sedation <36 h, weaning is faster with propofol. IMPLICATIONS: The duration of adequate sedation time is longer with propofol compared with midazolam. In postoperative patients with sedation < 36 h, weaning is faster with propofol. PMID- 11273937 TI - Stroke volume variation as a predictor of fluid responsiveness in patients undergoing brain surgery. AB - Changes in arterial blood pressure induced by mechanical ventilation allow assessment of cardiac preload. In this study, stroke volume variation (SVV), which is the percentage change between the maximal and minimal stroke volumes (SV) divided by the average of the minimum and maximum over a floating period of 30 s, continuously displayed by the PiCCO continuous cardiac output monitor, was evaluated as a predictor of fluid responsiveness. Fifteen patients undergoing brain surgery were included. During surgery, graded volume loading was performed with each volume loading step (VLS) consisting of 100 mL of 6% hydroxyethylstarch given for 2 min. Successive responsive VLSs were performed (increase in SV > 5% after a VLS) until a change in SV of < 5 % was reached (nonresponsive). A total of 140 VLSs were performed. Responsive and nonresponsive VLSs differed in their pre-VLS values of systolic blood pressure, SV, and SVV, but not in the values of heart rate and central venous pressure. By using receiver operating characteristic analysis, the area under the curve for SVV (0.870, 95% confidence interval [CI]: 0.809 to 0.903) was statistically more than those for central venous pressure (0.493, 95% CI: 0.397 to 0.590, P = 7 x 10(-10)), heart rate (0.593, 95% CI: 0.443 to 0.635, P = 5.7 x 10(-10)), and systolic blood pressure (0.729, 95% CI: 0.645 to 0.813, P: = 4.3 x 10(-3)). An SVV value of 9.5% or more, will predict an increase in the SV of at least 5% in response to a 100-mL volume load, with a sensitivity of 79% and a specificity of 93%. IMPLICATIONS: Stroke volume variation may be used as a continuous preload variable and in combination with the continuously measured cardiac output, defining on-line the most important characteristics of cardiac function, allowing for optimal fluid management. PMID- 11273938 TI - Microvascular endothelial dysfunction and its mechanism in a rat model of subarachnoid hemorrhage. AB - After subarachnoid hemorrhage (SAH), large cerebral arteries are prone to vasospasm. Using a rat model of SAH, we examined whether cortical microvessels demonstrate vasomotor changes that may make them prone to spasm and whether endothelial dysfunction may account for any observed changes. Two days after percutaneous catheterization into the cisterna magna, 0.3 mL of autologous blood was injected into the subarachnoid space. The brain tissue was harvested 20 min later, and microvessels were dissected from the parietal cortex. Vasomotor responses to the thromboxane analog U46619, the protein kinase C agonist phorbol acetate, endothelin-1, adenosine diphosphate, nitroprusside, and isoproterenol were examined in vitroin cerebral arterioles from the control, sham-operated, and SAH animals. Endothelial nitric oxide synthase (NOS3) messenger RNA and protein concentration was measured by northern and western blotting, respectively. Arterioles from the SAH animals demonstrated attenuated dilation to the endothelium-dependent dilator adenosine diphosphate and accentuated constriction to endothelin-1, while responses to the other agents tested were unchanged. NOS3 protein concentration was decreased, but NOS3 messenger RNA was increased after SAH. After SAH, cortical arterioles demonstrate endothelial dysfunction, which may be the basis for microvascular spasm. This is in part related to decreased NOS3, which occurs despite an increase in its transcription. IMPLICATIONS: Acute microvascular endothelial dysfunction may occur after subarachnoid hemorrhage and contribute to microvascular spasm. PMID- 11273939 TI - The effects of an increase of central blood volume before spinal anesthesia for cesarean delivery: a qualitative systematic review. AB - We evaluated in this qualitative systematic review the efficacy of increasing central blood volume on the incidence of hypotension after spinal anesthesia for elective cesarean delivery. Randomized controlled trials investigating any method of increasing central blood volume before the initiation of obstetric spinal anesthesia were sought by using MEDLINE (1966-2000), Embase (January 1988-April 2000), and the Cochrane Library (Issue 1, 2000). Additional reports from retrieved and review articles, hand searching of non-MEDLINE journals, and abstracts of major anesthesia meetings (1994-1999) were located. The primary outcome was the incidence of hypotension. Secondary outcomes included: ephedrine use, Apgar scores, umbilical cord pH values, and maternal nausea and vomiting. Twenty-three articles met our inclusion criteria with the use of crystalloid preload, colloid preload, and mechanical methods of increasing central volume. Crystalloid preload was inconsistent in preventing hypotension, whereas colloid appeared to be effective in all but one study. Leg wrapping and thromboembolic stockings decreased the incidence of hypotension compared with leg elevation or control. Few differences in fetal outcomes or maternal nausea and vomiting were reported. Increasing central blood volume by using colloid and leg wrapping decreases but does not abolish the incidence of hypotension before spinal anesthesia for elective cesarean delivery. IMPLICATIONS: We performed a systematic review to determine whether fluid loading reduced the incidence of low blood pressure after spinal anesthesia for cesarean delivery. Although no technique totally eliminates the occurrence of hypotension, colloid administration (starch or gelatin containing fluids) and leg wrapping were the most effective. PMID- 11273940 TI - The effects of opioids on isolated human pregnant uterine muscles. AB - We determined the effects of fentanyl, sufentanil, morphine, and meperidine on the spontaneous contractility of isolated human pregnant uterine muscle strips. Uterine specimens were obtained from normal full-term parturients undergoing elective lower-segment cesarean delivery. Longitudinal muscle strips were prepared and mounted vertically in tissue chambers to record their isometric tension. Opioid concentration-response curves were constructed after rhythmic contractions were established. The responses were also examined in the presence of opioid receptor blocker, nitric oxide synthase inhibitor, beta-adrenoceptor blocker, or cyclooxygenase inhibitor. Fentanyl and meperidine inhibited uterine contractility in a concentration-dependent manner, their concentration that inhibited 50% being 2.3 x 10(-6) and 1.0 x 10(-3)M, respectively. Sufentanil and morphine had no significant effects on uterine contractility. Pretreatment with either naloxone, N(G)-nitro-L-arginine methyl ester, atenolol, or indomethacin did not affect the uterine responses to opioids. These results demonstrate that fentanyl and meperidine may have direct inhibitory effects on the contractility of the human uterus, though at supraclinical concentrations. IMPLICATIONS: Opioids do not have a significant effect on spontaneous contractions of gravid human uterine muscle at their clinically relevant concentrations. PMID- 11273941 TI - The effect of intravenous ketorolac on opioid requirement and pain after cesarean delivery. AB - Nonsteroidal antiinflammatory drugs, including ketorolac, are widely used for postoperative analgesia. This randomized, double-blinded trial compared IV ketorolac or saline combined with meperidine patient-controlled epidural analgesia (PCEA) after cesarean delivery. Fifty healthy parturients scheduled for elective cesarean delivery under combined spinal-epidural anesthesia received PCEA plus either IV ketorolac (Group K) or saline (Group C) for 24 h. The ketorolac dose was modified, after six patients had been studied, based on new product information recommending a maximum of 120 mg ketorolac over 24 h. Group K (n = 24) and Group C (n = 20) were demographically similar. During the first 24 h, Group K used significantly less meperidine (P < 0.05). Postoperative pain at rest and with movement, and patient satisfaction, did not differ significantly between groups, except that worst pain at 12 h was less in Group K (P < 0.005). The two groups were similar with respect to patient recovery and side effects. IV ketorolac, as an adjunct to PCEA after cesarean delivery, produced a meperidine dose-sparing effect of approximately 30%, but did not significantly improve pain relief, reduce opioid-related side effects, or change patient outcome. PMID- 11273942 TI - The effects of thoracic epidural analgesia with bupivacaine 0.25% on ventilatory mechanics in patients with severe chronic obstructive pulmonary disease. AB - Optimal analgesia is important after thoracotomy in pulmonary-limited patients to avoid pain-related pulmonary complications. Thoracic epidural anesthesia (TEA) can provide excellent pain relief. However, potential paralysis of respiratory muscles and changes in bronchial tone might be unfavorable in patients with end stage chronic obstructive pulmonary disease (COPD). Therefore, we evaluated the effect of TEA on maximal inspiratory pressure, pattern of breathing, ventilatory mechanics, and gas exchange in 12 end-stage COPD patients. Pulmonary resistance, work of breathing, dynamic intrinsic positive end-expiratory pressure, and peak inspiratory and expiratory flow rates were evaluated by assessing esophageal pressure and airflow. An increase in minute ventilation (7.50 +/- 2.60 vs 8.70 +/ 2.10 L/min; P = 0.04) by means of increased tidal volume (0.46 +/- 0.16 vs 0.53 +/- 0.14 L/breath; P = 0.003) was detected after TEA. These changes were accompanied by an increase in peak inspiratory flow rate (0.48 +/- 0.17 vs 0.55 +/- 0.14 L/s; P = 0.02) and a decrease in pulmonary resistance (20.7 +/- 9.9 vs 16.6 +/- 8.1 cm H(2)O. L(-1). s(-1); P = 0.02). Peak expiratory flow rate, dynamic intrinsic positive end-expiratory pressure, work of breathing, PaO(2), and maximal inspiratory pressure were unchanged (all P > 0.50). We conclude that TEA with bupivacaine 0.25% can be used safely in end-stage COPD patients. IMPLICATIONS: Thoracic epidural anesthesia with bupivacaine 0.25% does not impair ventilatory mechanics and inspiratory respiratory muscle strength in severely limited chronic obstructive pulmonary disease patients. Thus, thoracic epidural anesthesia can be used safely in patients with end-stage chronic obstructive pulmonary disease. PMID- 11273943 TI - The clinical use of small-dose tetracaine spinal anesthesia for transurethral prostatectomy. AB - In a double-blinded study, we compared conventional dose tetracaine (8 mg), small dose tetracaine (4 mg) with added fentanyl and epinephrine, and small-dose tetracaine (4 mg) with added fentanyl subarachnoid anesthesia. Forty-five patients scheduled for transurethral resection of prostate (TURP) under subarachnoid anesthesia were randomly assigned to Group 1 (8 mg hyperbaric tetracaine), Group 2 (4 mg hyperbaric tetracaine, 10 microg fen-tanyl, and 0.2 mg epinephrine), and Group 3 (4 mg hyperbaric tetracaine, 10 microg fentanyl, and 0.2 mL saline). Evaluations were performed after spinal anesthesia. Subarachnoid block was successful in all patients except one in Group 1, who required general anesthesia by mask. The median peak sensory levels 10 min after the induction of spinal anesthesia in Group 1 was T8, which was significantly higher than Group 2 and Group 3 (P < 0.05). The time of sensory and motor recovery in Group 3 was less than in Groups 1 and 2 (P < 0.05). Hypotension was observed in four patients in Group 1 and none in Groups 2 and 3. We conclude that small-dose 4-mg hyperbaric tetracaine plus 10 microg fentanyl might provide adequate anesthesia and fewer side effects for TURP when compared with the conventional (8 mg) dose. IMPLICATIONS: Small-dose hyperbaric tetracaine (4 mg with 10 microg fentanyl added) may provide adequate anesthesia and fewer side effects for transurethral resection of the prostate. PMID- 11273944 TI - A comparison of ropivacaine with fentanyl to bupivacaine with fentanyl for postoperative patient-controlled epidural analgesia. AB - Ropivacaine for patient-controlled epidural analgesia (PCEA) may facilitate postoperative patient mobilization because it causes less motor block than bupivacaine. Forty patients undergoing abdominal surgery were randomized in a double-blinded manner to the following: 0.05% bupivacaine/4 microg fentanyl, 0.1% bupivacaine/fentanyl, 0.05% ropivacaine/fentanyl, or 0.1% ropivacaine/fentanyl for standardized PCEA. We measured pain scores, side effects, and PCEA consumption for 42 h. Lower-extremity motor function was assessed with electromyography and isometric force dynamometry. Analgesia was equivalent among groups. Local anesthetic use was more in the 0.1% Ropivacaine and 0.1% Bupivacaine groups (77% increase, P = 0.001). Motor function decreased during PCEA (10%-35% decrease from preoperative, P < 0.001) and was equivalent among groups. Eight patients were transiently unable to ambulate. These patients used more local anesthetic (45 vs 33 mg mean, P < 0.05) with additional decrease in motor function (32%, P < 0.004) compared with ambulating patients. Other side effects were mild and equivalent among solutions. PCEA with bupivacaine/fentanyl and ropivacaine/fentanyl as 0.05% or 0.1% solutions appears clinically equipotent. Lower-extremity motor function decreases, but is unlikely to result in prolonged inability to ambulate. Use of a 0.05% solution may be advantageous to decrease local anesthetic use and prevent transient motor block. IMPLICATIONS: Patient-controlled epidural analgesia with bupivacaine/fentanyl and ropivacaine/fentanyl as either 0.05% or 0.1% solutions are clinically similar. Lower-extremity motor function will decrease with the use of any of these combinations, but is unlikely to result in the inability to walk. PMID- 11273945 TI - Soy-containing diet suppresses chronic neuropathic sensory disorders in rats. AB - Partial sciatic nerve ligation (PSL) in rodents produces chronic neuropathic sensory disorders resembling neuropathic pain in humans. We previously reported that levels of allodynia and hyperalgesia after PSL injury were markedly attenuated by consumption of soy-containing diets. Here we aimed to show that dietary effect on pain behavior is not specific to a certain laboratory. For this purpose, experiments were conducted in a different laboratory (Baltimore rather than Jerusalem) and a different rat strain (Wistar rather than Sabra), with additional and different testing methods (radiant heat from a lamp rather than a CO(2) laser). Rats were fed two soy-free diets and a soy-containing one for 28 days. The sensitivity of rats to nonnoxious and noxious stimuli was determined before PSL injury, and levels of neuropathic sensory disorders were determined after it. We found that consuming the soy-containing diet prevented development of tactile and heat allodynia, but not mechanical hyperalgesia. This dietary effect was not correlated with calorie intake and weight gain or dietary concentration of fat and carbohydrates. We conclude that, regardless of experimental site, diet markedly affects chronic neuropathic sensory disorders in rats and should be standardized in animal models of pain. IMPLICATIONS: Levels of chronic sensory disorders in a rat model of allodynia and hyperalgesia after partial sciatic nerve ligation depend on the consumption of a soy-containing diet. Further studies are needed to determine the role of diet in humans with chronic pain. PMID- 11273946 TI - Local anesthesia does not block mustard-oil-induced temporomandibular inflammation. AB - Temporomandibular joint (TMJ) disorders and rheumatoid arthritis are two conditions in which neurogenic mechanisms may play a critical role. We investigated the neurogenic contribution underlying acute TMJ inflammation by evaluating effects of local anesthetic blockade of afferent innervation on the development of mustard oil (MO)-induced edema in the rat TMJ area. Groups of eight adult male Sprague-Dawley rats were anesthetized by intraperitoneal alpha chloralose and urethane. A saline injection into the right TMJ followed by MO (1% to 60%) 6 min later elicited dose-dependent edema development (P < 0.05, repeated measures analysis of variance). Lidocaine (5%) or bupivacaine (0.5%) followed by MO (1% or 40%) did not produce edema development different from saline controls (P > 0.05, repeated measures analysis of variance). The failure of local anesthetic blockade to prevent MO-induced edema is not consistent with MO acting through a neurogenic mechanism, as traditionally perceived. IMPLICATIONS: Inflammation found in temporomandibular disorders and rheumatoid arthritis may result from mediators released by pain-sensing neurons. Local anesthesia failed to block simulated neurogenic temporomandibular inflammation in a rat model, suggesting that functional neuronal input may not be necessary for the promotion of inflammation. PMID- 11273947 TI - Local anesthetics attenuate lysophosphatidic acid-induced priming in human neutrophils. AB - Lysophosphatidic acid (LPA) is an intercellular phospholipid mediator with a variety of actions that suggest a role in stimulating inflammatory responses. We therefore studied its actions on neutrophil (PMN) motility and respiratory burst. Because local anesthetics (LA) inhibit LPA signaling and attenuate PMN responses, we also investigated the effects of LA on these actions. Chemotaxis of human PMNs under agarose toward LPA (10(-10)-10(-3) M) was studied, with and without 1 h prior incubation in lidocaine (10(-9)-10(-4) M). Priming as well as activating effects of LPA on PMNs were measured using a cytochrome-c assay of superoxide anion (O2-) production. PMNs were incubated with lidocaine, tetracaine, or S-(-) ropivacaine (all at 10(-6)-10(-4) M) for 10 min or 1 h to assess interference with LPA signaling. LPA demonstrated chemoattractive effects towards human PMNs; this effect was concentration-dependently attenuated by lidocaine. LPA alone did not activate PMNs. However, it acted as a priming agent. LA in clinically relevant concentrations decreased (O2-) production induced by LPA/N formylmethionine-leucyl-phenylanaline. LPA acts as a chemoattractant and priming agent; however, it does not activate PMNs. LA, in clinically relevant concentrations, attenuate chemotactic and metabolic responses as a result of LPA. These results may explain the antiinflammatory effect of local anesthestics. IMPLICATIONS: Lysophosphatidic acid (LPA) influences two functions of human neutrophils, migration and metabolic activity. It acted as a chemoattractant and a priming-but not activating-agent. Responses to LPA were attenuated by local anesthetics in clinically relevant concentrations. PMID- 11273948 TI - Pelvic hematoma after an ilioinguinal nerve block for orchialgia. PMID- 11273949 TI - The successful treatment of a spino-subcutaneous fistula after bone marrow harvest by using an epidural blood patch. PMID- 11273950 TI - Intravenous ketoprofen in thyroid and parathyroid surgery. AB - We compared the ketoprofen-propacetamol combination relative to propacetamol alone in thyroid and parathyroid surgery in terms of postoperative analgesic efficacy, bleeding, and incidence of nausea and vomiting to determine whether ketoprofen results in any benefit in this type of surgery. Patients were distributed in two parallel groups to be managed by anesthesiologists habitually prescribing (Ketoprofen group) or not prescribing (Control group) ketoprofen in this situation. The same anesthetic technique was used for all patients. Postoperative analgesia consisted of 2 g of propacetamol every 6 h and morphine boluses if the pain score measured by the numerical rating scale pain exceeded 40 (3 mg IV every 10 min in the recovery room, then 5 mg SC every 4 h in the ward). The Ketoprofen group received 100 mg of ketoprofen IV during surgery (starting on resection of specimen) and 8 h later. In the recovery room, patients received oxygen if the SpO(2) while they were breathing room air was < 95% on admission and at 1 and 2 h. Pain scores, opioid consumption, the volume of the cervical draining fluid, and the concentration and mass of hemoglobin in this fluid collected over 24 h were recorded. The 214 patients were distributed into two groups (n = 107 in each group) that were comparable in terms of age, weight, sex, duration of surgery, type of endocrinopathy, surgeon involvement, and the intraoperative dose of sufentanil (P > 0.2). The Ketoprofen group had lower numerical rating scale (P < 0.05), received less morphine during the first 24 h after surgery (7.4 +/- 5 vs 11.7 +/- 6 mg, P < 0.05), had fewer nausea and vomiting episodes (21 vs 38, P < 0.05), and were less likely to require oxygen breathing after 1 h in the recovery room (33 vs 59 patients, P < 0.05). The two groups had the same 24-h volume of cervical draining fluid (72.5 +/- 43 vs 70 +/- 42 mL, P > 0.2) and the same concentration (5.9 +/- 3.4 vs 6.4 +/- 2.8 g per 100 mL, P > 0.1) and mass of hemoglobin (3.9 +/- 2.8 vs 4.2 +/- 2.5 g, P > 0.2) in this collected fluid. Two cervical hematomas necessitating reintervention occurred in the Control group, compared with none in the Ketoprofen group. Ketoprofen reduces the pain score after thyroid and parathyroid surgery, as well as morphine requirements and related adverse effects, without increasing the risk of cervical bleeding. IMPLICATIONS: In a prospective open study, ketoprofen reduced the pain score after thyroid and parathyroid surgery, as well as morphine requirements and related adverse effects, without increasing the risk of cervical bleeding. PMID- 11273951 TI - Ketorolac is not nephrotoxic in connection with sevoflurane anesthesia in patients undergoing breast surgery. AB - Ketorolac, which may cause renal vasoconstriction by cyclooxygenase inhibition, is often administered to patients anesthetized with sevoflurane that is metabolized to inorganic fluoride (F(-)), another potential nephrotoxin. We assessed this possible interaction using urine N-acetyl-beta-D-glucosaminidase indexed to urinary creatinine (U-NAG/crea) as a marker of proximal tubular, beta2 microglobulin as a tubular, urine oxygen tension (P(u)O(2)) as a medullary, and erythropoietin as a marker of tubulointerstitial damage. Thirty women (ASA physical status I-II) undergoing breast surgery were included in our double blinded study. They were allocated into two groups receiving either ketorolac 30 mg IM (Group K) or saline (Group C) at the time of premedication, at the end of, and 6 h after anesthesia maintained with sevoflurane. Urine output, U-NAG/crea, P(u)O(2,) serum creatinine, urea, and F(-) were assessed. Blood loss was larger in Group K (465 +/- 286 mL vs 240 +/- 149 mL, mean +/- SD, P < 0.05). The MAC doses of sevoflurane were similar. U-NAG/crea increased during the first 2 h of anesthesia and serum F(-) peaked 2 h after the anesthesia without differences between the groups. There were no statistically significant changes in P(u)O(2), erythropoietin, beta2-microglobulin, serum creatinine, urea, or urine output during anesthesia or the recovery period in either group. Our results indicate that the kidneys are not affected by ketorolac administered in connection with sevoflurane anesthesia. IMPLICATIONS: The different kinetics of N-acetyl-beta-D glucosaminidase indexed to urinary creatinine and serum inorganic fluoride during and after sevoflurane anesthesia suggest that the observed mild renal tubular function deterioration is not caused by inorganic fluoride. Administration of ketorolac IM is therefore considered safe in adequately hydrated healthy adult patients given sevoflurane anesthesia. PMID- 11273952 TI - Lactate is correlated with the indocyanine green elimination rate in liver resection for cirrhotic patients. AB - The role of lactate in liver ischemia-reperfusion injury in cirrhosis has not been clarified. Fifty patients with hepatocellular carcinoma who underwent partial liver resection under Pringle's maneuver were included in this study. We performed the indocyanine green clearance test before the operation and three times during the surgery to calculate its elimination rate. Blood lactate and base excess were measured at the corresponding times. Systolic and diastolic systemic arterial pressure, heart rate, cardiac index, and esophageal temperature were monitored. Aminotransferase levels were recorded the day before the operation, 1 h after the operation, and on the first and third postoperative days. We calculated the increase or decrease in lactate levels during the preischemic, ischemic, and postischemic phases, and examined the correlation between these results and the changes in indocyanine green elimination rate and some clinical factors. The lactate levels increased before reperfusion and began to decrease after reperfusion. The lactate increase and decrease during the ischemic and postischemic phases correlated with the change in indocyanine green elimination rate (P < 0.0001 and P = 0.02 for the respective phases). The lactate increase during the preischemic phase correlated with the duration of the preischemic phase (P < 0.0001). In cirrhotic patients who undergo liver resection with Pringle's maneuver and who do not show postoperative liver failure, the blood lactate profile might be a reliable indicator of liver metabolic capacity during surgery. IMPLICATIONS: In cirrhotic patients who underwent liver resection with Pringle's maneuver, the lactate increase and decrease during the ischemic and postischemic phases correlated with the change in the indocyanine green elimination rate. The blood lactate profile might be a reliable indicator of liver metabolic capacity during surgery. PMID- 11273953 TI - Postoperative left vocal cord dysfunction caused by Ortner's cardiovocal syndrome. PMID- 11273954 TI - Creation of individual soaking compartments within a glutaraldehyde fume hood. PMID- 11273955 TI - Immediate or early extubation: where do we start? PMID- 11273956 TI - Statistical clarification. PMID- 11273957 TI - Difficult insertion of interscalene brachial plexus catheter. PMID- 11273958 TI - New intravenous catheter not suitable for trans-tracheal jet ventilation. PMID- 11273959 TI - Endotoxin augments cerebral hyperemic response to halothane by inducing nitric oxide synthase. PMID- 11273960 TI - Improved energetics may explain the favorable effect of insulin infusion on bupivacaine cardiotoxicity. PMID- 11273961 TI - Use of intracuff lidocaine during general anesthesia. PMID- 11273962 TI - Proper probe positioning for infants with compromised ventilation from transesophageal echocardiography. PMID- 11273963 TI - Warm up. PMID- 11273964 TI - Evidence-based sports medicine. PMID- 11273965 TI - Do mouthguards prevent concussion? PMID- 11273967 TI - Dental evaluation of scuba diving mouthpieces using a subject assessment index and radiological analysis of jaw position. AB - OBJECTIVE: To compare two experimental scuba mouthpieces with a commercially available design. METHODS: A laboratory study using six men to assess effort, muscle pain, muscle fatigue, facial discomfort, tooth discomfort, and loss of lip sensation using a visual analogue scale. Cephalometric radiographs and analysis of jaw position with each mouth piece were also used. RESULTS: Fully customised mouthpieces caused the least discomfort, muscle pain, fatigue, and effort. They also resulted in the least mandibular displacement from the resting position. Radiographic analysis of jaw position showed that the fully customised design resulted in the least displacement from normal jaw position. CONCLUSIONS: A fully customised design gives the greatest comfort, least effort, and least mandibular displacement. This design is recommended, particularly for divers who experience temporomandibular dysfunction associated with diving. PMID- 11273966 TI - Diabetes and extreme altitude mountaineering. PMID- 11273968 TI - What do under 15 year old schoolboy rugby union players think about protective headgear? AB - OBJECTIVES: When protective headgear is designed, the attitudes of the intended users needs to be taken into account, as well as safety performance criteria. The aim of this study was therefore to determine the attitudes of schoolboy rugby union players towards protective headgear. METHODS: A survey of 140 rugby union players (82.4% response rate) from 10 randomly selected school teams in metropolitan Sydney was conducted at the end of the 1999 playing season. All players were aged 14-16 years. All teams had participated in a trial of headgear during the 1999 season in which six of the teams had been assigned to a headgear trial arm and four teams to a control arm. Players completed a self report questionnaire during a supervised session at school. The questionnaire collected information on recent head injuries, use of protective equipment, and attitudes towards headgear. RESULTS: Some form of protective equipment was always worn by 76.1% of players: 93.6% reported using a mouthguard and 79.3% a helmet/headgear during the 1999 season. The two most important reasons for wearing headgear were related to safety concerns. Players with no recent head/neck injury were more likely to report that they felt safer when wearing headgear (p<0.001) and less likely to cite a previous injury as a motivating factor for wearing headgear (p<0.001) than those who had sustained a recent head/neck injury. Of the players who wore headgear during the 1999 season, 67% said that they played more confidently when they wore headgear, but 63% said that their head was hotter. Few players reported that their head was uncomfortable (15%) or that it was hard to communicate (3%) when they wore headgear. The main reasons for not wearing headgear were related to its design features: uncomfortable (61%) and it was hot (57%). CONCLUSIONS: The primary reason cited by players for wearing headgear is safety. Receiving an injury would also motivate non-wearers to wear headgear. Players report that they are more confident and able to tackle harder if they wear headgear, suggesting that a belief in its protective capabilities may influence behaviour. These attitudes need to be addressed in the design of effective headgear as well as in strategies to promote its use. PMID- 11273969 TI - Echocardiographic characteristics of male athletes of different age. AB - Two dimensionally guided M mode and Doppler echocardiographic data for 578 male subjects (106 non-athletic and 472 athletes) were analysed from two aspects: (a) in the young adult category (19--30 years of age), competitors in different groups of sports were studied; (b) in the different age groups (children, 10--14 years; adolescent juniors, 15--18 years; young adults, 19--30 years; adults, 31- 44 years; older adults 45--60 years), data for athletes and non-athletes were compared. Morphological variables were related to body size by indices in which the exponents of the numerator and denominator were matched. Morphological signs of athletic heart were most consistently evident in the left ventricular muscle mass: in the young adult group, the highest values were seen in the endurance athletes, followed by the ball game players, sprinters/jumpers, and power athletes. A thicker muscular wall was the main reason for this hypertrophy. Internal diameter was only increased in the endurance athletes, and this increase was more evident in the younger groups. The E/A quotient (ratio of peak velocity during early and late diastole) indicated more effective diastolic function in the endurance athletes. The values for E/A quotient also suggested that regular physical activity at an older age may protect against age dependent impairment of diastolic function. PMID- 11273970 TI - Contact dermatitis in students practicing sports: incidence of rubber sensitisation. AB - BACKGROUND: Over the last few years, changes in cutaneous homoeostasis resulting from sports activities have been reported. In particular, alterations in sweating mechanisms, the hydrolipid barrier, and surface bacterial flora, together with exposure to atmospheric conditions and the need to use medicaments, detergents, and other topical substances, predispose subjects to allergic contact dermatitis. OBJECTIVE: To evaluate the incidence of allergic contact dermatitis in a group of young people practising sports activities. METHODS: Patch tests were performed to confirm the diagnosis of irritant or allergic dermatitis; in addition, the radioallergoabsorbent test (RAST) to latex was evaluated in the group studied. RESULTS: Allergic contact dermatitis caused by thiourams (23.3%) and mercaptobenzothiazole (20.9%) was prevalent. Other haptens, such as benzocaine and nickel, which are contained in clothing, equipment, topical medicaments, and creams used for massage, were also allergenic. In two cases, RAST positivity to latex was registered. CONCLUSIONS: -The results suggest that close contact with sports equipment may increase the incidence of allergic contact dermatitis. Students practising certain sports may have "professional" allergic contact dermatitis to additives used in the production of rubber. PMID- 11273971 TI - Ankle injuries in basketball: injury rate and risk factors. AB - OBJECTIVES: To determine the rate of ankle injury and examine risk factors of ankle injuries in mainly recreational basketball players. METHODS: Injury observers sat courtside to determine the occurrence of ankle injuries in basketball. Ankle injured players and a group of non-injured basketball players completed a questionnaire. RESULTS: A total of 10 393 basketball participations were observed and 40 ankle injuries documented. A group of non-injured players formed the control group (n = 360). The rate of ankle injury was 3.85 per 1000 participations, with almost half (45.9%) missing one week or more of competition and the most common mechanism being landing (45%). Over half (56.8%) of the ankle injured basketball players did not seek professional treatment. Three risk factors for ankle injury were identified: (1) players with a history of ankle injury were almost five times more likely to sustain an ankle injury (odds ratio (OR) 4.94, 95% confidence interval (CI) 1.95 to 12.48); (2) players wearing shoes with air cells in the heel were 4.3 times more likely to injure an ankle than those wearing shoes without air cells (OR 4.34, 95% CI 1.51 to 12.40); (3) players who did not stretch before the game were 2.6 times more likely to injure an ankle than players who did (OR 2.62, 95% CI 1.01 to 6.34). There was also a trend toward ankle tape decreasing the risk of ankle injury in players with a history of ankle injury (p = 0.06). CONCLUSIONS: Ankle injuries occurred at a rate of 3.85 per 1000 participations. The three identified risk factors, and landing, should all be considered when preventive strategies for ankle injuries in basketball are being formulated. PMID- 11273972 TI - Is glucose/amino acid supplementation after exercise an aid to strength training? AB - BACKGROUND: The precise timing of carbohydrate and amino acid ingestion relative to a bout of resistance exercise may modulate the training effect of the resistance exercise. OBJECTIVE: To assess whether regular glucose/amino acid supplementation immediately after resistance exercise could enhance the gain in muscle strength brought about by resistance training. METHODS: Seven untrained participants with a median age of 23 years and mean (SD) body mass 68.9 (13.5) kg resistance trained on a leg extension machine for five days a week for 10 weeks, using four sets of 10 repetitions. Alternate legs were trained on successive days, one leg each day. Subjects ingested either a supplement including 0.8 g glucose/kg and 0.2 g amino acids/kg, or placebo, on alternate training days immediately after training. Therefore the supplement was always ingested after training the same leg (supplement leg). Isometric, isokinetic, and 1 repetition maximum (RM) strength were measured before, during, and after training. Blood samples were analysed to determine the acute responses of insulin and glucose to resistance exercise and supplementation or placebo. RESULTS: Serum insulin concentration peaked 20 minutes after supplement ingestion at ninefold the placebo level, and remained significantly elevated for at least 80 minutes (p<0.01). Isometric, isokinetic, and 1 RM strength improved on both supplement and placebo legs (p<0.05). There were no significant differences in the gain in strength between the supplement leg and the placebo leg (p>0.05). CONCLUSION: Regular glucose/amino acid supplementation immediately after resistance exercise is unlikely to enhance the gain in muscle strength brought about by resistance training. PMID- 11273973 TI - Benefits from aerobic exercise in patients with major depression: a pilot study. AB - BACKGROUND: Several reports indicate that physical activity can reduce the severity of symptoms in depressed patients. Some data suggest that even a single exercise bout may result in a substantial mood improvement. OBJECTIVE: To evaluate the short term effects of a training programme on patients with moderate to severe major depression. METHODS: Twelve patients (mean (SD) age 49 (10) years; five men, seven women) with a major depressive episode according to the Diagnostic and Statistical Manual of the American Society of Psychiatry (DSM IV) criteria participated. The mean (SD) duration of the depressive episode was 35 (21) weeks (range 12--96). Training consisted of walking on a treadmill following an interval training pattern and was carried out for 30 minutes a day for 10 days. RESULTS: At the end of the training programme, there was a clinically relevant and statistically significant reduction in depression scores (Hamilton Rating Scale for Depression: before, 19.5 (3.3); after, 13 (5.5); p = 0.002. Self assessed intensity of symptoms: before, 23.2 (7); after, 17.7 (8.1); p = 0.006. Values are mean (SD)). Subjective and objective changes in depression scores correlated strongly (r = 0.66, p = 0.01). CONCLUSIONS: Aerobic exercise can produce substantial improvement in mood in patients with major depressive disorders in a short time. PMID- 11273974 TI - Short term power output in relation to growth and maturation. AB - OBJECTIVE: To examine short term power output during growth and maturation using a multilevel modelling approach. METHODS: Body mass, stature, and triceps and subscapular skinfold thicknesses of boys and girls, aged 12.2 (0.4) years (mean (SD)) at the onset of the study, were measured at age 12, 13, and 17 years. Sexual maturation, classified according to Tanner's stage of pubic hair development, was assessed on the first two occasions and assumed to be stage 5 at 17 years. Peak power (PP) and mean power (MP) were assessed on each occasion using the Wingate anaerobic test. RESULTS: Initial models, founded on 417 determinations of short term power output, identified body mass, stature, and age as significant explanatory variables of both PP and MP. The values for girls were significantly lower than those for boys, and a significant age by sex interaction described a progressive divergence in the MP of boys and girls. The introduction of sum of two skinfold thicknesses produced a model with an improvement in fit as indicated by a significant change in log likelihood. The stature term was negated and the body mass term increased. The age and sex terms were reduced but remained significant. The age by sex interaction term remained a significant explanatory variable for MP. Maturity effects were non-significant additional explanatory variables in all models of power output. CONCLUSION: The values of PP and MP for boys are higher than those for girls, and, for MP, sex differences increase with age. Body mass and skinfold thicknesses are significant influences on both PP and MP, but age exerts a positive but non-linear effect on power output independent of body size and fatness. PMID- 11273975 TI - Warm up practices of golfers: are they adequate? AB - BACKGROUND: Although it is widely recommended that golfers warm up before play or practice to enhance their physical performance and reduce their injury risk, it is not known to what extent they actually undertake such warm up procedures. OBJECTIVE: To collect information about the proportion of golfers who actively warm up and to determine the types of warm up behaviours. METHODS: This study was conducted over three weeks at three different golfing venues: a private golf course, a public golf course, and a golf driving range. Golfers' warm up behaviours, defined as any form of preparative exercise, were recorded by direct observation by two independent observers. RESULTS: The sample consisted of 1040 amateur golfers (852 men and 188 women) aged at least 18 years. Only 54.3% (95% confidence interval 49.8 to 58.8) performed some form of warm up activity. Air swings on the tee were the most commonly observed warm up activity, with 88.7% (95% confidence interval 85.9 to 91.5) of golfers who warmed up performing these. CONCLUSIONS: Only a small proportion of amateur golfers perform appropriate warm up exercises. To improve on this, golfers should be educated about the possible benefits of warming up and be shown how to perform an appropriate warm up routine. PMID- 11273976 TI - Sports doctors' resuscitation skills under examination: do they take it seriously? AB - As 64% of sports medicine doctors were unable to show proficiency at basic life support and assessment and management of a seriously injured patient with a potential spinal injury in the last two examinations for a University of Bath diploma in sports and exercise medicine, it was decided that a reminder is required of the importance of acquiring, at the very least, some basic resuscitation skills. An analysis and comment on the results from the first aid component of the examination is also presented. PMID- 11273977 TI - Sural nerve injury associated with neglected tendo Achilles ruptures. AB - Two patients are described with delayed presentation of a ruptured tendo Achilles, each exhibiting signs of sural nerve dysfunction. Recovery occurred in each case after operative repair. PMID- 11273978 TI - Avulsion fracture of the extensor carpi radialis longus in a rugby player: a case report. AB - Avulsion fracture of the base of the second metacarpal is an unusual injury, and the cause in the few cases reported in the literature was a fall on a volarly flexed wrist. A case of this rare injury suffered in a sport related accident by a semiprofessional rugby player is reported. It was treated with open reduction and internal fixation after failure of conservative management. PMID- 11273980 TI - Effectiveness of stretching to reduce injury. PMID- 11273979 TI - Embolisation of a traumatic aneurysm of the posterior circumflex humeral artery in a volleyball player. AB - Repetitive minor vascular injuries caused by physical activity in athletes may lead to ischaemia of the upper extremities. In volleyball players in particular, traumatic aneurysm of the posterior circumflex humeral artery has been reported to be a cause of ischaemia of the arm and hand. Such an aneurysm is described here; it was treated successfully with endovascular embolisation. PMID- 11273981 TI - The ups and downs of high altitude mountaineering. PMID- 11273982 TI - Professionalism and injuries in rugby union. PMID- 11273984 TI - Preprescription genotyping: not yet ready for prime time, but getting there. PMID- 11273985 TI - How many medicines do patients with heart failure need? PMID- 11273986 TI - Distribution of Chlamydia pneumoniae in the human arterial system and its relation to the local amount of atherosclerosis within the individual. AB - BACKGROUND: Chlamydia pneumoniae has been suggested to play a role in the origin of atherosclerosis. We studied the prevalence of C pneumoniae at multiple locations in the arterial system within the same individual. Studying the association between atherosclerosis and C pneumoniae within the individual excludes confounding by interindividual variability. METHODS AND RESULTS: Postmortem, the presence in the intima/plaque and media of C pneumoniae membrane protein was determined by use of a C pneumoniae-specific monoclonal antibody. In 24 individuals, 33 arterial locations were studied (n=738 segments). Area stenosis was determined in adjacent cross sections. In all individuals, immunostaining of C pneumoniae was observed in >/=1 artery. The highest prevalences were observed in the abdominal aorta (67%), internal and common iliac arteries (41%), and coronary arteries (33%). The lowest prevalences were observed in the radial (0%) and cerebral (2%) arteries. Within the individual, area stenosis was larger in cross sections with immunoreactivity compared with cross sections without immunoreactivity (31.0+/-11.9% versus 14.3+/-6.1%, respectively; P:<0.001). In the individual, immunoreactivity was observed in 15+/-10% of the arteries (range, 3% to 45%). Between individuals, the percentage of arteries with immunoreactivity to C pneumoniae was associated with the average area stenosis throughout the arterial system (r(2)=0.56, P:<0.001). CONCLUSIONS: C pneumoniae was mostly observed at locations that are related to clinically relevant features. Within the individual, the distribution of C pneumoniae is associated with the distribution of atherosclerosis. The role of the microorganism in atherosclerotic disease remains to be elucidated. PMID- 11273987 TI - Ascorbate restores endothelium-dependent vasodilation impaired by acute hyperglycemia in humans. AB - BACKGROUND: Endothelium-dependent vasodilation is impaired in patients with insulin-dependent and non-insulin-dependent diabetes mellitus and restored by vitamin C administration, implicating a causative role for oxidant stress. Hyperglycemia per se attenuates endothelium-dependent vasodilation in healthy subjects. Accordingly, this study investigated whether impaired endothelium dependent vasodilation caused by hyperglycemia in nondiabetic humans is restored by administration of the antioxidant vitamin C. METHODS AND RESULTS: Endothelium dependent vasodilation was measured by incremental brachial artery administration of methacholine chloride (0.3 to 10 microg/min) during euglycemia, after 6 hours of hyperglycemia (300 mg/dL) created by dextrose (50%) intra-arterial infusion, and with coadministration of vitamin C (24 mg/min) during hyperglycemia. Endothelium-dependent vasodilation was significantly diminished by hyperglycemia (P:=0.02 by ANOVA) and restored by vitamin C (P:=0.04). In contrast, endothelium dependent vasodilation was not affected by equimolar infusions of mannitol, with and without vitamin C coinfusion (P:=NS). Endothelium-independent vasodilation was measured by incremental infusion of verapamil chloride (10 to 300 microg/min) without and with coadministration of N:(G)-monomethyl-L-arginine (L-NMMA). In the absence of L-NMMA, endothelium-independent vasodilation was not significantly altered during hyperglycemia (P:=NS) but was augmented by vitamin C (P:=0.04). The coadministration of L-NMMA eliminated the vitamin C-related augmentation in verapamil-mediated vasodilation. CONCLUSIONS: Vitamin C administration restores endothelium-dependent vasodilation impaired by acute hyperglycemia in healthy humans in vivo. These findings suggest that hyperglycemia may contribute in part to impaired vascular function through production of superoxide anion. PMID- 11273988 TI - Ischemic preconditioning prevents endothelial injury and systemic neutrophil activation during ischemia-reperfusion in humans in vivo. AB - BACKGROUND: Endothelial dysfunction leading to neutrophil infiltration of tissues has been implicated in tissue injury caused by ischemia-reperfusion (IR). Tissue injury during IR can be reduced by prior ischemic preconditioning (IPC). In humans, it is unclear whether endothelial dysfunction occurs during IR or whether IPC offers protection against endothelial dysfunction and inflammatory cell activation. We studied the effects of experimental IR on endothelial and neutrophil function in the human forearm in vivo and examined the protection afforded by IPC. METHOD AND RESULTS: The forearm was made ischemic for 20 minutes by inflating a blood pressure cuff to 200 mm Hg. We assessed endothelial function of conduit (radial artery flow-mediated dilation) and resistance vessels (blood flow responses to intra-arterial infusion of the endothelium-dependent dilator acetylcholine) in healthy volunteers before and after IR. IR reduced flow mediated dilation of the radial artery at 15 minutes of reperfusion (7.7+/-1.5% to 3.5+/-0.9%) and the dilator response of resistance vessels to acetylcholine at 15, 30, and 60 minutes of reperfusion. IR did not reduce the dilator response of the radial artery to glyceryltrinitrate and only caused a small reduction of glyceryltrinitrate-induced dilation of resistance vessels at 60 minutes of reperfusion. IR caused an increase in neutrophil CD11b expression and platelet neutrophil complexes in the circulating blood. IPC (three 5-minute episodes of ischemia) before IR prevented endothelial dysfunction and neutrophil activation. CONCLUSIONS: A clinically relevant period of ischemia-reperfusion causes profound and sustained endothelial dysfunction and systemic neutrophil activation. IPC attenuates both of these effects in humans. PMID- 11273989 TI - Electromechanical mapping for detection of myocardial viability in patients with ischemic cardiomyopathy. AB - BACKGROUND: We evaluated the ability of electromechanical mapping of the left ventricle to distinguish between nonviable and viable myocardium in patients with ischemic cardiomyopathy. METHODS AND RESULTS: Unipolar voltage amplitudes and local endocardial shortening were measured in 31 patients (mean+/-SD age, 62+/-8 years) with ischemic cardiomyopathy (ejection fraction, 30+/-9%). Dysfunctional regions, identified by 3D echocardiography, were characterized as nonviable when PET revealed matched reduction of perfusion and metabolism and as viable when perfusion was reduced or normal and metabolism was preserved. Mean unipolar voltage amplitudes and local shortening differed among normal, nonviable, and viable dysfunctional segments. Coefficient of variation for local shortening exceeded differences between groups and did not allow distinction between normal and dysfunctional myocardium. Optimum nominal discriminatory unipolar voltage amplitude between nonviable and viable dysfunctional myocardium was 6.5 mV, but we observed a great overlap between groups. Individual cutoff levels calculated as a percentage of electrical activity in normal segments were more accurate in the detection of viable dysfunctional myocardium than a general nominal cutoff level. The optimum normalized discriminatory value was 68%. Sensitivity and specificity were 78% for the normalized discriminatory value compared with 69% for the nominal value (P:<0.02). CONCLUSIONS: Endocardial ECG amplitudes in patients with ischemic cardiomyopathy display a wide scatter, complicating the establishment of exact nominal values that allow distinction between viable and nonviable areas. Individual normalization of unipolar voltage amplitudes improves diagnostic accuracy. Electroanatomic mapping may enable identification of myocardial viability. PMID- 11273990 TI - Acetylcholine release in human heart atrium: influence of muscarinic autoreceptors, diabetes, and age. AB - BACKGROUND: An imbalance of sympathetic and parasympathetic drive to the heart is an important risk factor for cardiac death in patients with coronary heart disease, diabetes, and renal insufficiency. The amount of neurotransmitter released from peripheral autonomic nerves is modulated by presynaptic receptor systems. In analogy to alpha-autoreceptors on sympathetic nerves, muscarinic autoreceptors activated by endogenous acetylcholine may exist on parasympathetic nerves in the human heart. METHODS AND RESULTS: We developed a technique to study acetylcholine release from human atria and investigated muscarinic autoreceptor function. A pharmacological and molecular approach was used to characterize the subtype involved. Of the 5 muscarinic receptor subtypes cloned, only mRNA encoding for M(2)- and M(3)-receptors were detected. Potencies of several muscarinic antagonists against the release-inhibiting effect of the nonselective muscarinic agonist carbachol at the cardiac autoreceptor were correlated with published data for human cloned M(1)- through M(5)-receptors. CONCLUSIONS: This analysis clearly indicates that acetylcholine release in human atria is controlled by muscarinic M(2)-receptors. Blockade of these receptors by atropine doubles the amount of acetylcholine released at a stimulation frequency of 5 Hz. In atria of patients >70 years of age and patients with late diabetic complications, acetylcholine release is reduced. Locally impaired cardiac acetylcholine release may therefore represent a pathophysiological link to sudden cardiac death in elderly and diabetic patients. PMID- 11273991 TI - Pharmacogenetic interactions between beta-blocker therapy and the angiotensin converting enzyme deletion polymorphism in patients with congestive heart failure. AB - BACKGROUND: Activation of the renin-angiotensin and sympathetic nervous systems adversely affect heart failure progression. The ACE deletion allele (ACE D) is associated with increased renin-angiotensin activation; however, its influence on patient outcomes remains uncertain, and the pharmacogenetic interactions with beta-blocker therapy have not been previously evaluated. METHODS AND RESULTS: We prospectively followed 328 patients (age, 56.1+/-11.9 years) with systolic dysfunction (left ventricular ejection fraction, 0.24+/-0.08) to assess the impact of the ACE D allele on transplant-free survival (median follow-up, 21 months). Transplant-free survival was compared by genotype for the whole cohort and separately in patients with (n=120) and those without beta-blocker therapy (n=208) at the time of entry. Transplant-free survival was significantly poorer for patients with the D: allele (1-year percent survival II/ID/DD=94/77/75; 2 year=78/65/60; ordered log-rank test, P:=0.044). In patients not treated with beta-blockers, the adverse impact of ACE D allele was dramatically increased (1 year percent survival II/ID/DD=95/75/67; 2-year=81/61/48; P:=0.005). In contrast, in patients receiving beta-blocker therapy, no influence of ACE genotype on transplant-free survival was evident (1-year percent survival II/ID/DD=91/80/86; 2-year=70/71/77; P:=0.73). CONCLUSIONS: In a cohort of patients with systolic dysfunction, the ACE D allele was associated with a significantly poorer transplant-free survival. This effect was primarily evident in patients not treated with beta-blockers and was not seen in patients receiving therapy. These findings suggest a potential pharmacogenetic interaction between the ACE D/I polymorphism and therapy with beta-blockers in the determination of heart failure survival. PMID- 11273993 TI - Scaling exponent predicts defibrillation success for out-of-hospital ventricular fibrillation cardiac arrest. AB - BACKGROUND: -Defibrillator shocks often fail to terminate ventricular fibrillation (VF) in out-of-hospital cardiac arrest (OOHCA), and repeated failed shocks can worsen the subsequent response to therapy. Because the VF waveform changes with increasing duration of VF, it is possible that ECG analyses could estimate the preshock likelihood of defibrillation success. This study examined whether an amplitude-independent measure of preshock VF waveform morphology predicts outcome after defibrillation. Methods and Results-Clinical data and ECG recordings from an automated external defibrillator were obtained for 75 subjects with OOHCA in a suburban community with police first responders and a paramedic based emergency medical system. An estimate of the fractal self-similarity dimension, the scaling exponent, was calculated off-line for the VF waveform preceding shocks. Success of the first shock was determined from the recordings. Return of pulses and survival were determined by chart review. The first shock resulted in an organized rhythm in 43% of cases, and 17% of cases survived to hospital discharge. A lower mean value of the scaling exponent was observed for cases in which the first defibrillation resulted in an organized rhythm (P:=0.004), for cases with return of pulses (P:=0.049), and for cases surviving to hospital discharge (P:<0.001). Receiver operator curves revealed the utility of the scaling exponent for predicting the probability of restoring an organized rhythm (area under the curve=0.70) and of survival (area under the curve=0.84). CONCLUSIONS: -The VF waveform in OOHCA can be quantified with the scaling exponent, which predicts the probability of first-shock defibrillation and survival to hospital discharge. PMID- 11273992 TI - Upregulation of beta(3)-adrenoceptors and altered contractile response to inotropic amines in human failing myocardium. AB - BACKGROUND: Contrary to beta(1)- and beta(2)-adrenoceptors, beta(3)-adrenoceptors mediate a negative inotropic effect in human ventricular muscle. To assess their functional role in heart failure, our purpose was to compare the expression and contractile effect of beta(3)-adrenoceptors in nonfailing and failing human hearts. METHODS AND RESULTS: We analyzed left ventricular samples from 29 failing (16 ischemic and 13 dilated cardiomyopathic) hearts (ejection fraction 18.6+/-2%) and 25 nonfailing (including 12 innervated) explanted hearts (ejection fraction 64.2+/-3%). beta(3)-Adrenoceptor proteins were identified by immunohistochemistry in ventricular cardiomyocytes from nonfailing and failing hearts. Contrary to beta(1)-adrenoceptor mRNA, Western blot analysis of beta(3)-adrenoceptor proteins showed a 2- to 3-fold increase in failing compared with nonfailing hearts. A similar increase was observed for Galpha(i-2) proteins that couple beta(3) adrenoceptors to their negative inotropic effect. Contractile tension was measured in electrically stimulated myocardial samples ex vivo. In failing hearts, the positive inotropic effect of the nonspecific amine isoprenaline was reduced by 75% compared with that observed in nonfailing hearts. By contrast, the negative inotropic effect of beta(3)-preferential agonists was only mildly reduced. CONCLUSIONS: Opposite changes occur in beta(1)- and beta(3)-adrenoceptor abundance in the failing left ventricle, with an imbalance between their inotropic influences that may underlie the functional degradation of the human failing heart. PMID- 11273994 TI - Cardiac output and central distribution of blood flow in the human fetus. AB - BACKGROUND: The objectives of this study were to establish reference ranges for left and right cardiac output and to investigate blood flow distribution through the foramen ovale, ductus arteriosus, and pulmonary bed in human fetuses. METHODS AND RESULTS: A prospective study was performed in 222 normal fetuses from 13 to 41 weeks of gestation with high-resolution color Doppler ultrasound. Cardiac output and ductal flow were calculated by use of vessel diameter and the time velocity integral. Pulmonary blood flow was expressed as the difference between right cardiac output and ductal flow. Foramen ovale flow was estimated as the difference between pulmonary flow and left cardiac output. Gestational age specific reference ranges are given for left, right, and biventricular output and volume of ductal blood flow, showing an exponential increase with gestational age. Median ratio of right to left cardiac output was 1.42 and was not associated with gestational age. Right cardiac output was 59% and left cardiac output was 41% of biventricular cardiac output. Median biventricular cardiac output was estimated to be 425 mL. min(-1). kg(-1) fetal weight. Ductal blood flow was 46%, estimated pulmonary flow was 11%, and estimated foramen ovale flow was 33% of biventricular output. CONCLUSIONS: The study establishes reference ranges for fetal cardiac output and offers insights into the central blood flow distribution in human fetuses from 13 weeks to term. There is a clear right heart dominance. The estimated ratio of pulmonary blood flow to cardiac output is higher than in fetal lamb studies. PMID- 11273995 TI - Regional wall motion and abnormalities of electrical depolarization and repolarization in patients after surgical repair of tetralogy of Fallot. AB - BACKGROUND: Abnormal depolarization-repolarization in patients with repaired tetralogy of Fallot (TOF) is a risk factor for malignant ventricular tachycardia and sudden death. It is unclear whether ECG abnormalities are associated with abnormal regional right ventricular (RV) function. METHODS AND RESULTS: Seventy four patients (37 patients <18 and 37 >18 years old) who had had TOF repair at 4.0 years old (0.1 to 47 years old) were examined when they were 18.7 years old (1.7 to 61.1 years old), as were 112 control subjects with normal hearts. Regional function was evaluated with tissue Doppler imaging of the RV and left ventricular (LV) free wall and the septum. Myocardial velocities were sampled continuously from base to apex. Synchronous ECG was analyzed for QRS, QT, and JT duration and QRS, QT, and JT dispersion. All 74 TOF patients had normal LV myocardial velocities. Forty-eight patients (24 patients <18 and 24 >18 years old) had reversed myocardial velocities in diastole in the RV free wall, which were associated with reversed systolic myocardial velocities in 22 and additional reverse diastolic myocardial velocities in the septum in 19. Those 48 patients had a longer QRS duration (151+/-31 versus 124+/-27 ms) and greater QRS (47+/-18 versus 29+/-12 ms), QT (73+/-27 versus 52+/-22 ms), and JT (96+/-31 versus 67+/ 35 ms) dispersion. Compared with normal control subjects, all 74 TOF patients had decreased systolic and diastolic myocardial velocities and a longer isovolumic relaxation time. CONCLUSIONS: RV wall-motion abnormalities are a common finding late after TOF repair and are associated with repolarization-depolarization abnormalities. These data further underscore a likely mechanoelectrical interaction as an important part of the pathogenesis of RV disease in these patients. PMID- 11273996 TI - Role of cardiac nerves in the cardiovascular response to cocaine in conscious dogs. AB - BACKGROUND: Although the cardiovascular toxicity of cocaine is well recognized, considerable controversy remains as to the relative contribution of local norepinephrine reuptake inhibition versus central stimulatory effects of cocaine in eliciting its cardiovascular actions. The purpose of the present study was to determine the role of cardiac nerves in mediating the left ventricular (LV) and coronary hemodynamic responses to cocaine. METHODS AND RESULTS: We studied the cardiovascular response to acute cocaine administration (1 mg/kg) in 10 intact, conscious dogs and 6 dogs with ventricular denervation (VD). There were no significant differences in baseline hemodynamic parameters or plasma catecholamines between the 2 groups. In response to acute cocaine, LV and coronary hemodynamic responses were enhanced in the VD dogs. The enhanced systemic pressor and heart rate responses in VD dogs suggest that cardiac nerves mitigate the response to cocaine through ventricular mechanoreceptors rather than mediating the responses. CONCLUSIONS: These data suggest that peripheral blockade of norepinephrine reuptake is not the principal mechanism of the acute cardiac effects of cocaine. Rather, cardiac nerves modulate the effects of cocaine through baroreflex mechanisms. Thus, individual differences in baroreflex sensitivity may explain the hemodynamic variability observed in response to cocaine. PMID- 11273997 TI - Red wine does not reduce mature atherosclerosis in apolipoprotein E-deficient mice. AB - BACKGROUND: Red wine polyphenols and ethanol reduce fatty streak formation (early atherosclerosis) in various animal models. These experimental results support the observation that alcoholic beverages protect against myocardial infarction in humans. However, fatty streaks may not reflect the pathology of mature and clinically relevant atherosclerosis. The present study examined the effects of red wine polyphenols and ethanol on mature atherosclerosis in apolipoprotein E deficient mice. METHODS AND RESULTS: Eighty-four 7-week-old mice were randomized to receive water, red wine (diluted to 6% ethanol v/v), 6% ethanol v/v, or red wine powder in water. All mice were fed a normal chow diet. At 26 weeks of age, the mice were killed. HDL cholesterol was raised 12.0% (95% CI, 4.0% to 20.0%) and 9.2% (95% CI, 1.5% to 16.9%) by red wine and ethanol, respectively. At the end of study, all mice exhibited advanced atherosclerosis in the aortic bulb, whereas less mature atherosclerosis predominated in the brachiocephalic trunk. The amount of atherosclerosis in the aortic bulb and the brachiocephalic trunk were similar in all groups (P:=0.92 and P:=0.14, respectively). To evaluate whether ethanol or red wine polyphenols were protective by stabilizing atherosclerotic plaques rather than reducing their size, we measured the percentage of collagen-poor areas in left coronary sinus plaques as a morphological criterion of plaque stability. The percentage of collagen-poor areas did not differ between groups (P:=0.71). CONCLUSIONS: Neither ethanol nor red wine polyphenols reduced mature atherosclerosis or changed the content of collagen in plaques in apolipoprotein E-deficient mice. PMID- 11273998 TI - Aspirin (5 mmol/L) inhibits leukocyte attack and triggered reactive cell proliferation in a 3D human coronary in vitro model. AB - BACKGROUND: Leukocyte attack (LA) and the triggered reactive proliferation of smooth muscle cells (SMCs) are key events for the development of early atherosclerosis and restenosis. In the present study, we used a 3D human coronary in vitro model of LA (3DLA model) to examine the effect of high-dose aspirin on the adhesion and chemotaxis of leukocytes and the reactive proliferative response of SMCs. METHODS AND RESULTS: For dose-finding, the effect of aspirin (1, 2, 5, and 10 mmol/L) on the tumor necrosis factor-alpha-induced upregulation of intercellular adhesion molecule-1 was analyzed in monocultures of human coronary endothelial cells (HCAEC) and the SMCs of the human coronary media (HCMSMC). In cytoflow and Northern blot experiments, the expression of intercellular adhesion molecule-1 was slightly reduced after incubation with 5 mmol/L aspirin, and strong inhibition was found after incubation with 10 mmol/L. In 3DLA models, HCAECs and HCMSMCs were cultured on both sides of a porous filter. For LA, human monocytes or CD4(+) lymphocytes were seeded on the HCAEC side of the 3DLA unit. A dose of 5 mmol/L aspirin inhibited the adherence of monocytes or CD4(+) lymphocytes by 50% (P:<0.01) and the chemotaxis of monocytes by 90% (P:<0.01). The reactive proliferative response of cocultured HCMSMCs after LA, as measured by the uptake of bromodeoxyuridine, was significantly reduced by 83% after selective monocyte attack (P:<0.001) and by 42% after selective CD4(+) lymphocyte attack (P:<0.05). CONCLUSIONS: A local concentration of 5 mmol/L aspirin should be accepted as the lowest rational concentration for the beneficial in vitro effects of high-dose aspirin to be reproduced in clinical studies. PMID- 11273999 TI - H(mox-1) constitutes an adaptive response to effect antioxidant cardioprotection: A study with transgenic mice heterozygous for targeted disruption of the Heme oxygenase-1 gene. AB - BACKGROUND: Heme oxygenase-1 (H(mox-1)) has been implicated in protection of cells against ischemia/reperfusion injury. METHODS AND RESULTS: To examine the physiological role of H(mox-1), a line of heterozygous H(mox-1)-knockout mice was developed by targeted disruption of the mouse H(mox-1) gene. Transgene integration was confirmed and characterized at the protein level. A 40% reduction of H(mox-1) protein occurred in the hearts of H(mox-1)(+/)(-) mice compared with those of wild-type mice. Isolated mouse hearts from H(mox-1)(+/)(-) mice and wild type controls perfused via the Langendorff mode were subjected to 30 minutes of ischemia followed by 120 minutes of reperfusion. The H(mox-1)(+/)(-) hearts displayed reduced ventricular recovery, increased creatine kinase release, and increased infarct size compared with those of wild-type controls, indicating that these H(mox-1)(+/)(-) hearts were more susceptible to ischemia/reperfusion injury than wild-type controls. These results also suggest that H(mox-1)(+/)(-) hearts are subjected to increased amounts of oxidative stress. Treatment with 2 different antioxidants, Trolox or N:-acetylcysteine, only partially rescued the H(mox-1)(+/)(-) hearts from ischemia/reperfusion injury. Preconditioning, which renders the heart tolerant to subsequent lethal ischemia/reperfusion, failed to adapt the hearts of the H(mox-1)(+/)(-) mice compared with wild-type hearts. CONCLUSIONS: These results demonstrate that H(mox-1) plays a crucial role in ischemia/reperfusion injury not only by functioning as an intracellular antioxidant but also by inducing its own expression under stressful conditions such as preconditioning. PMID- 11274000 TI - Endothelium-derived hyperpolarizing factor : identification and mechanisms of action in human subcutaneous resistance arteries. AB - BACKGROUND: Both a vascular endothelial cytochrome P450 (CYP450) product of arachidonic acid metabolism and the potassium ion (K(+)) have been identified as endothelium-derived hyperpolarizing factors (EDHFs) in animal vascular tissues. We studied the relative importance of EDHF, nitric oxide (NO), and prostacyclin (PGI(2)) as vasodilators in human subcutaneous arteries. We also examined the mechanisms underlying the vasodilator action of EDHF to elucidate its identity. METHODS AND RESULTS: Subcutaneous resistance arteries were obtained from 41 healthy volunteers. The contribution of EDHF to the vasodilation induced by acetylcholine was assessed by inhibiting production of NO, PGI(2), and membrane hyperpolarization. The mechanisms underlying the relaxation evoked by K(+) and EDHF were elucidated. EDHF was found to account for approximately 80% of acetylcholine-mediated vasorelaxation. Its action was insensitive to the combination of barium and ouabain, whereas barium and ouabain reversed K(+) mediated vasorelaxation. EDHF-mediated vasorelaxation, however, was sensitive to the phospholipase A(2) inhibitor oleyloxyethyl phosphorylcholine and the CYP450 inhibitor ketoconazole. CONCLUSIONS: EDHF is the major contributor to endothelium dependent vasorelaxation in human subcutaneous resistance arteries. A product of phospholipase A(2)/CYP450-dependent metabolism of arachidonic acid and not K(+) is the likely identity of EDHF in human subcutaneous resistance arteries. PMID- 11274001 TI - Prevention of autoimmune myocarditis through the induction of antigen-specific peripheral immune tolerance. AB - BACKGROUND: Autoimmunity to cardiac antigens, in particular cardiac myosin, has been observed in humans with myocarditis and in animals with experimental inflammatory heart disease. Current treatments for myocarditis are in many cases immunosuppressive and might lead to increased cardiac damage by reducing host defenses against infectious agents. Therefore, we sought to develop an antigen specific approach to inhibit autoimmunity in mice with myosin-induced experimental autoimmune myocarditis. METHODS AND RESULTS: Syngeneic splenocytes, coupled with cardiac myosin by use of ethylene carbodiimide, were administered intravenously before disease induction, and the effects of this peripheral tolerization on myosin-induced myocarditis were assessed. This antigen-specific immunotherapy significantly reduced both the incidence and severity of myocarditis, with the prevention of myocyte necrosis, mononuclear cell infiltration, and fibrosis. Myosin-specific delayed-type hypersensitivity and antibody production were significantly reduced, demonstrating that peripheral tolerance affected both T- and B-cell responsiveness to the autoantigen. CONCLUSIONS: These results suggest that the induction of antigen-specific peripheral immune tolerance may be an effective approach for the treatment of myocarditides with autoimmune involvement. PMID- 11274002 TI - Change in character of mitral prosthesis regurgitant murmur with bundle-branch block: correlation of auscultatory and echocardiographic findings. PMID- 11274003 TI - Four-dimensional cardiac imaging with multislice computed tomography. PMID- 11274004 TI - Anatomy of the atrioventricular conduction system. PMID- 11274005 TI - Geographic miss: a cause of treatment failure in radio-oncology applied to intracoronary radiation therapy. PMID- 11274006 TI - New insights into the progression of aortic stenosis: implications for secondary prevention. PMID- 11274007 TI - Pseudo-myocardial infarction versus pseudo-pseudo-myocardial infarction. PMID- 11274009 TI - Pitfalls in the measurement of circulating vascular endothelial growth factor. AB - BACKGROUND: Vascular endothelial growth factor (VEGF) is a protein with antiapoptotic, mitogenic, and permeability-increasing activities specific for vascular endothelium. VEGF mRNA, which has five isoforms, is produced by nonmalignant cells in response to hypoxia and inflammation and by tumor cells in constitutively high concentrations. Because VEGF plays a crucial role in physiological and pathophysiological angiogenesis, measurements of circulating VEGF are of diagnostic and prognostic value, e.g., in cardiovascular failures, inflammatory diseases, and malignancies. However, there are major quantitative differences in the published results. This review attempts to identify reasons for these disparities. APPROACH: The literature was reviewed through a Medline search covering 1995 to 2000. A selection of exemplary references had to be made for this perspective overview. CONTENT: Data are included from studies on healthy humans, gynecological patients, and persons suffering from inflammatory or malignant diseases. The results indicate that competitive immunoassays detect the total amount of circulating VEGF, which enables observations regarding the increase in VEGF in pregnancy and preeclampsia to be made. In these cases, capture immunoassays utilizing neutralizing antibodies are insufficient because of an accompanying increase in VEGF-binding soluble receptors (sFlt-1). Measurements of circulating free VEGF are useful for study of malignant diseases, which are associated with both genetically and hypoxia-induced overproduction of VEGF. The VEGF isoform specificity of the antibodies is also critical because both VEGF(121) and VEGF(165) are secreted. It is important to consider that platelets and leukocytes release VEGF during blood clotting. CONCLUSIONS: Future efforts should concentrate on the balance between free VEGF, total VEGF, and sFlt 1. Plasma, rather than serum, should be used for analysis. PMID- 11274010 TI - Carcinoembryonic antigen as a marker for colorectal cancer: is it clinically useful? AB - BACKGROUND: Carcinoembryonic antigen (CEA) is one of the most widely used tumor markers worldwide. Its main application is mostly in gastrointestinal cancers, especially in colorectal malignancy. Although in use for almost 30 years, the clinical value of CEA in colorectal cancer is still not clear. METHODS: The literature relevant to the clinical value of CEA in colorectal cancer was reviewed. Particular attention was paid to studies involving metaanalyses and guidelines issued by Expert Panels. RESULTS: Although of little use in detecting early colorectal cancer, high preoperative concentrations of CEA correlate with adverse prognosis. Serial CEA measurements can detect recurrent colorectal cancer with a sensitivity of approximately 80%, a specificity of approximately 70%, and can provide a lead time of approximately 5 months. CEA is the most frequent indicator of recurrence in asymptomatic patients and currently is the most cost effective test for the preclinical detection of resectable disease. CEA is most useful for the early detection of liver metastasis in patients with diagnosed colorectal cancer. Overall, however, little evidence is available that monitoring of all patients with diagnosed colorectal cancer leads to enhanced patient outcome or quality of life. CONCLUSIONS: Currently, the most useful application of CEA is in the detection of liver metastasis from colorectal cancers. Because of the relative success of surgery in resecting hepatic metastases, serial determinations of the marker are recommended for detecting cancer spread to the liver. In the future, preoperative concentrations of CEA may be included with the standard staging procedures for assessing prognosis. PMID- 11274011 TI - Preoperative serum prostate-specific antigen (PSA) below 10 microg/l predicts neither the presence of prostate cancer nor the rate of postoperative PSA failure. AB - Recent information on the relationship of serum prostate-specific antigen (PSA) to prostate cancer and new reports on death rates in men warrant a reassessment of how we diagnose and treat prostate cancer. We now know for the first time that the annual death rate from prostate cancer in men > or =65 years of age is only 226 per 100 000 men. At least 40 000 of 100 000 men over age 65 (40%) have invasive prostate cancer as judged by examination of prostates in 3- to 4-mm step sections. Thus, only 1 of every 177 men 65 years of age or older (226 in 40 000) with invasive prostate cancer dies annually from his cancer. Serum PSA between 2 and 10 microg/L is used almost universally as an indication to biopsy the prostate. When 10-20 biopsies are commonly taken, it is not surprising that approximately 40% of men are biopsy-positive for prostate cancer. Despite this reliance on serum PSA as an indication for biopsy, data at Stanford show no clinically useful relationship between preoperative serum PSA (in the range 2-10 mg/L) and the volume of Gleason grade 4/5 cancer or the volume of Gleason grades 3, 2, and 1 cancer, nor can we show any useful relationship of such preoperative PSA concentrations (2-10 microg/L) to biochemical PSA failure rates after radical prostatectomy. We urgently need a better serum marker for prostate cancer. Because PSA biochemical failure rates after radical prostatectomy are directly proportional to the amount of Gleason grade 4/5 cancer in the prostate, a serum marker of Gleason grade 4/5 carcinoma could be ideal. PMID- 11274012 TI - DNA melting analysis for detection of single nucleotide polymorphisms. AB - BACKGROUND: Several methods for detection of single nucleotide polymorphisms (SNPs; e.g., denaturing gradient gel electrophoresis and denaturing HPLC) are indirectly based on the principle of differential melting of heteroduplex DNA. We present a method for detecting SNPs that is directly based on this principle. METHODS: We used a double-stranded DNA-specific fluorescent dye, SYBR Green I (SYBR) in an efficient system (PE 7700 Sequence Detector) in which DNA melting was controlled and monitored in a 96-well plate format. We measured the decrease in fluorescence intensity that accompanied DNA duplex denaturation, evaluating the effects of fragment length, dye concentration, DNA concentration, and sequence context using four naturally occurring polymorphisms (three SNPs and a single-base deletion/insertion). RESULTS: DNA melting analysis (DM) was used successfully for variant detection, and we also discovered two previously unknown SNPs by this approach. Concentrations of DNA amplicons were readily monitored by SYBR fluorescence, and DNA amplicon concentrations were highly reproducible, with a CV of 2.6%. We readily detected differences in the melting temperature between homoduplex and heteroduplex fragments 15-167 bp in length and differing by only a single nucleotide substitution. CONCLUSIONS: The efficiency and sensitivity of DMA make it highly suitable for the large-scale detection of sequence variants. PMID- 11274013 TI - zeta-, epsilon-, and gamma-Globin mRNA in blood samples and CD71(+) cell fractions from fetuses and from pregnant and nonpregnant women, with special attention to identification of fetal erythroblasts. AB - BACKGROUND: Information about the appearance of gamma-, epsilon-, and zeta-globin mRNAs in fetal erythroblasts during gestation and about the presence and amounts of these mRNAs in pregnant and nonpregnant women is important from the perspective of using these molecules as a marker of fetal erythroblasts. A specific marker is necessary for isolation and identification of fetal nucleated red blood cells from maternal blood samples for use in antenatal diagnosis of fetal genetic or chromosomal abnormalities. METHODS: We used a very sensitive reverse transcription-PCR (RT-PCR) method, coamplification analysis of gamma- and epsilon-globin cDNA, and quantitative analysis of gamma-globin mRNA based on competitive RT-PCR to investigate these aspects. RESULTS: All adult whole-blood samples were negative for epsilon- and zeta-globin mRNA. Analyses of CD71(+) cell fractions showed that specimens from 19 of 20 nonpregnant and 10 of 14 pregnant women (at 9-13 weeks of gestation) were positive for gamma-globin mRNA (Fisher's exact test, P = 0.13), and those from 3 of 20 nonpregnant and 5 of 14 pregnant women were positive for zeta-globin mRNA (Fisher's exact test, P = 0.23). No epsilon-globin mRNA was detected in CD71(+) cell fractions from 1-mL blood samples from adults. CD71(+) cell fractions from eight fetal blood samples (at 17 20 weeks of gestation) were positive for all three globin mRNAs. We found no statistically significant difference between the amounts of gamma-globin mRNA in pregnant and nonpregnant women. CONCLUSIONS: This study indicates that epsilon globin mRNA might function as a marker for fetal CD71(+) cells early in pregnancy. Although gamma-globin mRNA can be detected in CD71(+) cell fractions from most adults, these transcripts also may be of use because of a marked difference between adult and fetal values. PMID- 11274014 TI - Specific reverse transcription-PCR quantification of vascular endothelial growth factor (VEGF) splice variants by LightCycler technology. AB - BACKGROUND: Overexpression of vascular endothelial growth factor (VEGF) is associated with increased angiogenesis, growth and invasion in solid tumors, and hematologic malignancies. The expression of isoforms of VEGF, which mediate different effects, can be discriminated by splice-variant-specific quantitative reverse transcription-PCR (RT-PCR), but current methods have only modest sensitivity and precision and suffer from heteroduplex formation. METHODS: We used a real-time RT-PCR assay on the LightCycler system. Applicability for detection of different VEGF mRNAs and total VEGF message was tested on seven healthy tissues (each pooled from healthy donors) and seven correlated malignant tissues. Results were normalized to beta(2)-microglobulin mRNA. Amplification of VEGF splice variants was performed exclusively with variant-specific reverse primers, whereas forward primer and fluorescent probe were common to obtain similar RT-PCR kinetics. RESULTS: Highly specific detection of VEGF splice variants was achieved with minor intra- and interassay variation (<0.22 threshold cycle). Total VEGF expression was higher in malignant tissues. In healthy tissues, the mRNA encoding diffusible variants VEGF(121) and VEGF(165) constituted on average 78% (SD = 9.3%) of the total VEGF message, and the cell adherent variant VEGF(189) constituted on average 22% (SD = 5.4%). In contrast, in malignant tissues VEGF(121) and VEGF(165) accounted for 94% (SD = 7.6%) and VEGF(189) only 6% (SD = 3.7%). CONCLUSIONS: Because of the ability for quantification of VEGF splice variants with high specificity, sensitivity, and reproducibility, this new LightCycler assay is superior to conventional semiquantitative competitive RT-PCR and immunological assays and may contribute to better understanding of VEGF-mediated angiogenesis. PMID- 11274015 TI - C677T and A1298C polymorphisms of the methylenetetrahydrofolate reductase gene: incidence and effect of combined genotypes on plasma fasting and post-methionine load homocysteine in vascular disease. AB - BACKGROUND: Moderately increased plasma concentrations of total homocysteine (tHcy) have been shown to be an important risk factor for vascular diseases. Two common polymorphisms of the methylenetetrahydrofolate reductase (MTHFR) gene, the thermolabile C677T and a more recently reported A1298C polymorphism, may contribute to hyperhomocysteinemia. METHODS: Using PCR and restriction fragment length polymorphism analysis, we studied the prevalence of the C677T and A1298C MTHFR genotypes and the combined effect of these polymorphisms on plasma tHcy concentrations, as measured by HPLC with fluorometric detection, both fasting and post-methionine load (PML), in 1238 individuals. RESULTS: The prevalences of the C677T and A1298C genotypes did not differ significantly in 772 individuals with documented coronary artery disease (CAD), 137 individuals with deep-vein thrombosis (DVT), and 329 individuals without documented vascular disease. Individuals homozygous for the 677T allele had significantly increased fasting tHcy, particularly in the presence of low folate, compared with individuals homozygous for the wild-type allele. Neither the 1298AC nor the 1298CC genotype was associated with significantly increased fasting or PML tHcy concentrations irrespective of serum folate. Of the nine combined MTHFR genotypes, six were present in >10% of the population. Of these, the difference in mean fasting tHcy reached statistical significance (P<0.005) only in individuals with the 677TT/1298AA genotype compared with individuals with the wild-type 677CC/1298AA genotype. Differences in mean fasting tHcy did not reach statistical significance in individuals heterozygous for both MTHFR variants. We detected two 677CT/1298CC and three 677TT/1298AC individuals; only one, an 677TT/1298AC individual, had increased tHcy (both fasting and PML). No individuals had the 677TT/1298CC genotype. CONCLUSIONS: The prevalences of the C677T and A1298C polymorphisms did not differ among individuals with CAD, DVT, or those without documented vascular disease. In contrast to the C677T polymorphism, the A1298C polymorphism is not associated with increased fasting tHcy. Although the two polymorphisms usually exist in trans configuration, crossover may occur rarely to form recombinant chromosomes. PMID- 11274016 TI - Determination of RhD zygosity: comparison of a double amplification refractory mutation system approach and a multiplex real-time quantitative PCR approach. AB - BACKGROUND: Rh isoimmunization and hemolytic disease of the newborn still occur despite the availability of Rh immunoglobulin. For the prenatal investigation of sensitized RhD-negative pregnant women, determination of the zygosity of the RhD positive father has important implications. The currently available molecular methods for RhD zygosity assessment, in general, are technically demanding and labor-intensive. Therefore, at present, rhesus genotype assessment is most commonly inferred from results of serological tests. The recent elucidation of the genetic structure of the prevalent RHD deletion in Caucasians, as well as the development of real-time PCR, allowed us to explore two new approaches for the molecular determination of RhD zygosity. METHODS: Two methods for RhD zygosity determination were developed. The first was based on the double Amplification Refractory Mutation System (double ARMS). The second was based on multiplex real time quantitative PCR. For the double ARMS assay, allele-specific primers were designed to directly amplify the most prevalent RHD deletion found in RhD negative individuals in the Caucasian population. The multiplex real-time quantitative PCR assay, on the other hand, involved coamplification and quantification of RHD-specific sequences in relation to a reference gene, albumin, in a single PCR reaction. A ratio, DeltaCt, based on the threshold cycle, was then determined and reflects the RHD gene dosage. RESULTS: The allele specific primers of the double ARMS assay reliably amplified the RHD-deleted allele and therefore accurately distinguished homozygous from heterozygous RhD positive samples. The results were in complete concordance with serological testing. For the multiplex real-time quantitative PCR assay, the DeltaCt values clearly segregated into two distinct populations according to the RHD gene dosage, with mean values of 1.70 (SD, 0.17) and 2.62 (SD, 0.29) for the homozygous and heterozygous samples, respectively (P: <0.001, t-test). The results were in complete concordance with the results of serological testing as well as with the double ARMS assay. CONCLUSION: Double ARMS and real-time quantitative PCR are alternative robust assays for the determination of RhD zygosity. PMID- 11274018 TI - Improved diagnostic classification of alcohol abusers by combining carbohydrate deficient transferrin and gamma-glutamyltransferase. AB - BACKGROUND: Biochemical markers can provide objective evidence of high alcohol consumption. However, currently available markers have limitations in their diagnostic performance. METHODS: The diagnostic values of the most frequently used markers [carbohydrate-deficient transferrin (CDT), gamma-glutamyltransferase (GGT), aspartate aminotransferase, alanine aminotransferase, and mean corpuscular volume] were studied in an analysis of six different clinical studies (n = 1412) on alcohol abusers and social drinkers. The purpose of the analyses was to determine whether a combination of markers would improve the diagnosis of subjects. RESULTS: Discrimination between alcohol abusers and social drinkers, as measured by the areas under nonparametric ROC plots, was significantly better (P<0.001) for the new combined marker [gamma-CDT = 0.8. ln(GGT) + 1.3. ln(CDT)] than for any of the separate markers or combination of CDT or GGT with other markers. The cutoff values for gamma-CDT (6.5) can be taken to be the same among males and females. CONCLUSIONS: The combined variable gamma-CDT is a powerful tool to discriminate alcohol abusers from social drinkers and is recommended for clinical use. PMID- 11274017 TI - Highly sensitive, automated immunoassay for immunoglobulin free light chains in serum and urine. AB - BACKGROUND: Bence Jones proteins or monoclonal immunoglobulin kappa and lambda free light chains (FLCs) are important markers for identifying and monitoring many patients with B-cell tumors. Automated immunoassays that measure FLCs in urine and serum have considerable clinical potential. METHODS: Sheep antibodies, specific for FLCs, were prepared by immunization with pure kappa and lambda molecules and then adsorbed extensively against whole immunoglobulins. The antibodies were conjugated onto latex particles and used to assay kappa and lambda FLCs on the Beckman IMMAGE protein analyzer. RESULTS: The unconjugated antibodies showed minimal cross-reactivity with intact immunoglobulins or other proteins. With latex-conjugated antibodies, kappa and lambda FLCs could be measured in normal sera and most normal urine samples. Patients with multiple myeloma had increased concentrations of the relevant serum FLC, whereas both FLCs were increased in the sera of patients with systemic lupus erythematosus. CONCLUSIONS: We developed sensitive, automated immunoassays for kappa and lambda FLC measurements in serum and urine that should facilitate the assessment of patients with light chain abnormalities. PMID- 11274019 TI - Biochemical markers of bone formation in patients with plasma cell dyscrasias and benign osteoporosis. AB - BACKGROUND: Myeloma-induced bone loss is related to an uncoupling of bone formation and bone resorption. The aim of the present study was to assess the potential clinical value of biochemical markers of bone formation in the work up of patients with plasma cell dyscrasias. METHODS: Serum total alkaline phosphatase, bone-specific alkaline phosphatase (BAP), and osteocalcin (OC) were measured in 43 patients with newly diagnosed multiple myeloma (MM), in 40 patients with monoclonal gammopathy of undetermined significance (MGUS), in 40 patients with untreated benign vertebral osteoporosis (OPO), and in 48 healthy adults. RESULTS: In MM and MGUS patients, serum BAP, but not serum OC, was lower than in healthy controls (P<0.05). Serum OC was higher in patients with OPO than in healthy controls (P<0.05). The strongest associations between markers were found in OPO patients and in healthy adults. MM patients with early-stage disease or without detectable osteolysis had decreased serum BAP values (P<0.05). Serum OC was higher in MM patients with stage III disease (P<0.05) than in healthy controls. MM patients with OPO-like bone involvement had lower BAP values than sex- and age-matched MGUS patients with OPO-like bone involvement and patients with benign OPO (P<0.05). CONCLUSIONS: In patients with plasma cell dyscrasias, serum BAP, rather than serum OC, appears to reflect a suppressed bone formation rate and may be helpful in the differentiation between benign and myeloma-induced OPO. However, the overall clinical use of biochemical markers of bone formation in patients with plasma cell dyscrasia appears limited. PMID- 11274021 TI - Measurement of circulating forms of prostate-specific antigen in whole blood immediately after venipuncture: implications for point-of-care testing. AB - BACKGROUND: The purpose of this study was to validate the use of whole-blood samples in the determination of circulating forms of prostate-specific antigen (PSA). METHODS: Blood samples of hospitalized prostate cancer and benign prostatic hyperplasia patients were collected and processed to generate whole blood and serum samples. Three different rapid two-site immunoassays were developed to measure the concentrations of total PSA (PSA-T), free PSA (PSA-F), and PSA-alpha(1)-antichymotrypsin complex (PSA-ACT) to detect in vitro changes in whole-blood samples immediately after venipuncture. The possible influence of muscle movement on the release of PSA from prostate gland was studied in healthy men by measuring the rapid in vitro whole-blood kinetics of PSA forms before and after 15 min of physical exercise on a stationary bicycle. RESULTS: Rapid PSA-T, PSA-F, and PSA-ACT assays were designed using a 10-min sample incubation. No significant changes were detected in the concentrations of PSA-T, PSA-F, and PSA ACT from the earliest time point of 12-16 min compared with measurements performed up to 4 h after venipuncture. Physical exercise did not influence the concentrations of the circulating forms of PSA. Hematocrit-corrected whole-blood values of PSA-T and PSA-F forms were comparable to the respective serum values. Calculation of the percentage of PSA-F (PSA F/T ratio x 100) was similar irrespective of the sample format used, i.e., whole blood or serum. CONCLUSIONS: We found that immunodetectable PSA forms are likely at steady state immediately after venipuncture, thus enabling the use of anticoagulated whole-blood samples in near-patient settings for point-of-care testing, whereas determinations of PSA (e.g., PSA-T, PSA-F, or PSA-ACT) performed within the time frame of the office visit would provide results equivalent to conventional analyses performed in serum. PMID- 11274020 TI - Evaluation of a fully automated serum assay for C-terminal cross-linking telopeptide of type I collagen in osteoporosis. AB - BACKGROUND: Biochemical markers of bone turnover can provide prognostic information about the risk of osteoporotic fracture and are useful tools for monitoring efficacy of antiresorptive therapy. A serum-based automated assay may be of better clinical value than urinary markers because of lower imprecision and day-to-day within-person variability. Our aim was to evaluate the technical and clinical performances of a new, fully automated assay for serum C-terminal cross linking telopeptide of type I collagen (CTX), a marker of bone resorption. METHODS: Serum CTX was measured on the Elecsys 2010 automated analyzer (Roche). Results were compared with those of the manual ELISA. We measured serum CTX concentrations in 728 healthy women, ages 31-89 years. We investigated the ability of this assay to predict the rate of postmenopausal forearm bone loss evaluated by four repeated bone mineral density measurements using dual-x-ray absorptiometry in 305 women followed prospectively for 4 years. Finally, in a cohort of healthy, untreated, postmenopausal women, we compared baseline serum CTX in 55 women who subsequently had a fracture (20 vertebral and 35 peripheral fractures) with values in the 380 women who did not fracture during a mean 5 years of follow-up. RESULTS: The within- (n = 21) and between-run (n = 21) CVs were <4.1% and 5.7%, respectively. In 728 healthy women, serum CTX concentrations (automated) correlated with those of the manual ELISA (r = 0.82; P<0.0001). The median long-term within-person variability assessed by four repeated measurements over 3 months in 18 postmenopausal women was 9.4%. Compared with 254 premenopausal women, serum CTX was 39% (P<0.0001) higher in 45 perimenopausal women and 86% (P<0.0001) higher in 429 postmenopausal women (mean age, 64 years). Baseline serum CTX correlated negatively with changes of bone mass measured at the mid (r = -0.23; P<0.0001) and distal (r = -0.27; P<0001) radius. Postmenopausal women with serum CTX greater than the mean + 2 SD values in premenopausal women accounted for 42% of the population, lost bone at the mid radius on average eightfold more rapidly than the other women (-0.27% +/- 2.92% vs. -2.25% +/- 3.95%; P<0.0001), and had increased risk of fracture with a relative risk (95% confidence interval) of 1.8 (1.01-3.1) after adjustment for physical activity. CONCLUSIONS: The automated assay for serum CTX is precise and predicts rate of bone loss and fracture risk in postmenopausal women. Because it is convenient to use and has high throughput, this serum bone resorption marker may be useful for the investigation of patients with osteoporosis. PMID- 11274022 TI - Optimization of beta-quantification methods for high-throughput applications. AB - BACKGROUND: Risk of cardiovascular disease is assessed, in part, by laboratory measurement of the concentrations of several lipoproteins. beta-Quantification is a method of lipoprotein measurement that uses ultracentrifugation to partially separate lipoprotein classes. Although beta-quantification is used largely in clinical and basic research, methods have not been described to allow the analysis of a large number of small-volume specimens with a short turnaround time. We report two variations of the traditional 5-mL method used by the Lipid Research Clinics Program that overcome these shortcomings. METHODS: Two lower volume modifications of the traditional 5-mL beta-quantification method were developed. The methods used either 1 or 0.23 mL of specimen and required substantially less time for analysis (20 and 6 h, respectively) than the 5-mL method (2.5 days). The goal was to develop ultracentrifugation methods such that the concentration of cholesterol in the bottom fraction, from which LDL cholesterol concentration is calculated, agreed with the 5-mL method. Fresh serum specimens (n = 45) were analyzed by the three methods to determine comparability of the methods based on the recovery of cholesterol in the bottom fraction after ultracentrifugation. To evaluate intrarun precision, replicate specimens (n = 17) were analyzed in a single run for each method. This experiment also evaluated how quickly the fractions would remix after separation by ultracentrifugation. For the 1-mL method, accuracy of the measurement of LDL- and HDL-cholesterol concentrations and the interrun precision were established by analysis of frozen serum specimens provided by the CDC, which established target values for the pools using reference methods. RESULTS: No clinically significant differences in cholesterol concentrations in the bottom fraction were observed for the 1- and 0.23-mL methods, which had mean biases of 0.8% and 1.5% relative to the 5-mL method, respectively. Intra- and interrun variability was acceptable for each method, e.g., <1.8% for cholesterol in the bottom fraction. Ultracentrifuged specimens were stable for at least 4 h with no evidence of contamination of cholesterol in the bottom fraction. For comparison specimens provided by the CDC, the 1-mL method met the accuracy and precision goals of the National Cholesterol Education Program for the measurement of HDL- and LDL-cholesterol concentrations (goals: total error <13% and <12%, respectively), with total errors of 6.45% and 5.43%, respectively. CONCLUSIONS: Both the 1- and 0.23-mL beta-quantification methods are suitable substitutes for the traditional 5-mL method for use in clinical and basic research for the determination of LDL-cholesterol concentration. Both methods provide much higher throughput and require substantially less specimen volume. The 0.23-mL method can be performed in 1 day, but it is slightly less precise than the 1-mL method. In our laboratory setting, as many as 80 specimens are routinely processed per day using the 1-mL method. PMID- 11274023 TI - Fatty acid ethyl esters in liver and adipose tissues as postmortem markers for ethanol intake. AB - BACKGROUND: Fatty acid ethyl esters (FAEEs) are nonoxidative metabolites of ethanol. FAEEs are found in liver, pancreas, and adipose tissues up to 24 h after consumption of ethanol, and on that basis, they are potentially useful markers for ethanol intake. In this study with rats, we investigated the efficacy of using FAEEs in liver and in adipose tissue as postmortem markers for premortem ethanol ingestion. METHODS: An animal study was conducted in which test rats received injections of ethanol and control rats received injections of normal saline. The rats were killed 2 h after the injections. The bodies of the animals were stored at 4 degrees C up to 12 h, and samples of liver and adipose tissues were collected at different time intervals and processed for FAEE quantification. In another set of experiments, the rats received injections and were killed as described above, but bodies of animals from both groups were stored at 4, 25, or 37 degrees C for up to 72 h, and liver samples were collected and processed for FAEE quantification. RESULTS: FAEEs were detected up to 12 h after death in liver and adipose tissue samples from the bodies of ethanol-treated animals stored at 4 degrees C; negligible amounts were detected in the bodies of animals that received normal saline. Adipose tissues contained higher amounts of FAEEs than liver, as well as more species: eight FAEE species in adipose tissue and five in liver tissue. Higher concentrations of FAEEs were detected in livers of treated animals stored at 25 degrees C for up to 48 h than in livers of controls stored under the same conditions. CONCLUSIONS: For at least 12 h after death, FAEEs in liver and adipose tissues are useful postmortem markers of premortem ethanol ingestion. PMID- 11274024 TI - Measurement of immunoreactive angiotensin-(1-7) heptapeptide in human blood. AB - BACKGROUND: The renal enzyme renin cleaves from the hepatic alpha(2)-globulin angiotensinogen angiotensin-(1-10) decapeptide [Ang-(1-10)], which is further metabolized to smaller peptides that help maintain cardiovascular homeostasis. The Ang-(1-7) heptapeptide has been reported to have several physiological effects, including natriuresis, diuresis, vasodilation, and release of vasopressin and prostaglandins. METHODS: To investigate Ang-(1-7) in clinical settings, we developed a method to measure immunoreactive (ir-) Ang-(1-7) in 2 mL of human blood and to estimate plasma concentrations by correcting for the hematocrit. A sensitive and specific antiserum against Ang-(1-7) was raised in a rabbit. Human blood was collected in the presence of an inhibitor mixture including a renin inhibitor to prevent peptide generation in vitro. Ang-(1-7) was extracted into ethanol and purified on phenylsilylsilica. The peptide was quantified by radioimmunoassay. Increasing doses of Ang-(1-7) were infused into volunteers, and plasma concentrations of the peptide were measured. RESULTS: The detection limit for plasma ir-Ang-(1-7) was 1 pmol/L. CVs for high and low blood concentrations were 4% and 20%, respectively, and between-assay CVs were 8% and 13%, respectively. Reference values for human plasma concentrations of ir-Ang-(1 7) were 1.0-9.5 pmol/L (median, 4.7 pmol/L) and increased linearly during infusion of increasing doses of Ang-(1-7). CONCLUSIONS: Reliable measurement of plasma ir-Ang-(1-7) is achieved with efficient inhibition of enzymes that generate or metabolize Ang-(1-7) after blood sampling, extraction in ethanol, and purification on phenylsilylsilica, and by use of a specific antiserum. PMID- 11274025 TI - Plasma protein contents determined by Fourier-transform infrared spectrometry. AB - BACKGROUND: Fourier-transform infrared (FT-IR) spectrometry has been used to measure small molecules in plasma. We wished to extend this use to measurement of plasma proteins. METHODS: We analyzed plasma proteins, glucose, lactate, and urea in 49 blood samples from 35 healthy subjects and 14 patients. For determining the concentration of each biomolecule, the method used the following steps: (a) The biomolecule was sought for which the correlation between spectral range areas of plasma FT-IR spectra and concentrations determined by comparison method was greatest. (b) The IR absorption of the biomolecule at the most characteristic spectral range was calculated by analyzing pure samples of known concentrations. (c) The plasma concentration of the biomolecule was determined using the FT-IR absorption of the pure compound and the integration value obtained for the plasma FT-IR spectra. (d) The spectral contribution of the biomolecule was subtracted from the plasma FT-IR spectra, and the resulting spectra were saved for further analyses. (e) The same method was then applied to determining the concentrations of other biomolecules by sequentially comparing the resulting FT-IR spectra. RESULTS: Results agreed with those obtained by clinical methods for the following biomolecules when analyzed in the following order: albumin, glucose, fibrinogen, IgG(2), lactate, IgG(1), alpha(1)-antitrypsin, alpha(2)-macroglobulin, transferrin, apolipoprotein (Apo)-A(1), urea, Apo-B, IgM, Apo-C(3), IgA, IgG(4), IgG(3), IgD, haptoglobin, and alpha(1)-acid glycoprotein. CONCLUSION: FT-IR spectrometry is a useful tool for determining concentrations of several plasma biomolecules. PMID- 11274027 TI - Decreased serum zinc in fructose malabsorbers. PMID- 11274026 TI - Semiautomated DNA mutation analysis using a robotic workstation and molecular beacons. AB - BACKGROUND: Our increasing knowledge of the genetic basis of inheritable diseases requires the development of automated reliable methods for high-throughput analyses. METHODS: We investigated the combination of semiautomated DNA extraction from blood using a robotic workstation, followed by automated mutation detection using highly specific fluorescent DNA probes, so-called molecular beacons, which can discriminate between alleles with as little as one single-base mutation. We designed two molecular beacons, one recognizing the wild-type allele and the other the mutant allele, to determine genotypes in a single reaction. To evaluate this procedure, we examined the C677T mutation in the methylenetetrahydrofolate reductase (MTHFR) gene, which is associated with an increased risk for cardiovascular disease and neural tube defects. DNA was isolated from 10 microL of fresh EDTA-blood samples by use of a robotic workstation. The DNA samples were analyzed using molecular beacons as well as conventional methods. RESULTS: Both methods were compared, and no differences were found between outcomes of genotyping. CONCLUSIONS: The described assay enables robust and automated extraction of DNA and analysis of up to 96 samples (10 microL of blood per sample) within 5 h. This is superior to conventional methods and makes it suitable for high-throughput analyses. PMID- 11274028 TI - ELISA methodology for detection of modified osteoprotegerin in clinical studies. PMID- 11274029 TI - Umbilical cord and maternal plasma thiol concentrations in normal pregnancy. PMID- 11274030 TI - Effect of storage on phenylalanine and tyrosine measurements in whole-blood samples. PMID- 11274031 TI - Mean serum concentration of vitamin D-binding protein (Gc globulin) is related to the Gc phenotype in women. PMID- 11274032 TI - Effect of hemoglobin variants (Hb J, Hb G, and Hb E) on HbA1c values as measured by cation-exchange HPLC (Diamat). PMID- 11274033 TI - Errors caused by the use of D,L-octanoylcarnitine for blood-spot calibrators. PMID- 11274034 TI - Semiautomation of nucleic acid-based assays for Chlamydia trachomatis and Neisseria gonorrhoeae. PMID- 11274035 TI - Sensitive method for detection and semiquantification of Bence Jones protein by cellulose acetate membrane electrophoresis using colloidal silver staining. PMID- 11274036 TI - Stability of busulfan in frozen plasma and whole blood samples. PMID- 11274037 TI - Gas chromatographic-mass spectrometric measurement of 15-deoxy-delta(12,14) prostaglandin J(2), the peroxisome proliferator-activated receptor gamma ligand, in urine. PMID- 11274039 TI - Two single-tube tetra-primer assays to detect the CYP2C19*2 and *3 alleles of S mephenytoin hydroxylase. PMID- 11274038 TI - Plasma cholesterol concentrations in twin children: estimates of genetic and environmental influences. PMID- 11274040 TI - ARMS allele-specific amplification-based detection of mutant p53 DNA and mRNA in tumors of the breast. PMID- 11274041 TI - Chemiluminescent assay for serum thyroglobulin in the management of patients with thyroid carcinoma. PMID- 11274042 TI - Diagnosis of alpha-L-iduronidase deficiency in dried blood spots on filter paper: the possibility of newborn diagnosis. PMID- 11274043 TI - Cardiac troponin T is not detected in Western blots of diseased renal tissue. PMID- 11274044 TI - The chemical work of Alexander and Jane Marcet. AB - Alexander Marcet was an authority on urinary calculi and their analysis when few medical practitioners appreciated the usefulness of chemistry in the explanation and treatment of disease. In An Essay on The Chemical History and Medical Treatment of Calculous Disorders, he described the discovery of an xanthine stone. He drew line illustrations of simple chemical apparatus useful for bedside analysis. His microtechnique used drops of solution and pinhead pieces of calculi; reagents were acids and alkalies and the blowpipe in conjunction with a small alcohol lamp. He reported the earliest description of a disorder later named "alcaptonuria". Marcet's work and that of a few others, on the chemical composition of urine and calculi, laid the foundations of our present knowledge. Between 1807 and 1820, his lectures to the medical students at Guy's Hospital were illustrated by experiments. Jane Haldimand Marcet wrote the very popular CONVERSATIONS: on Chemistry (16 editions in Great Britain). Her book dominated elementary chemical instruction during the first half of the 19th century. She followed Lavoisier's scheme of classification and explained chemical reactions in terms of affinity, aggregation, gravitation, and repulsion. Her advocacy that experimentation accompany lecture was new. The availability of serious scientific education in the new women's academies set the stage for increasing women's involvement in science. She also published a series of CONVERSATIONS: The topics were Political Economy, Natural Philosophy, and Vegetable Physiology. PMID- 11274045 TI - Measurement of total protein is a useful inclusion in liver function test profiles. PMID- 11274046 TI - Effects of blood-collection systems and tubes on hematologic, chemical, and coagulation tests and on plasma hemoglobin. PMID- 11274047 TI - Autoimmune hypoglycemia presenting as seizure one week after surgery. PMID- 11274050 TI - LIS1 and dynein motor function in neuronal migration and development. PMID- 11274051 TI - A mutation in a mitochondrial transmembrane protein is responsible for the pleiotropic hematological and skeletal phenotype of flexed-tail (f/f) mice. AB - We have studied the flexed-tail (f) mouse to gain insight into mammalian mitochondrial iron metabolism. Flexed-tail animals have axial skeletal abnormalities and a transient embryonic and neonatal anemia characterized by pathologic intramitochondrial iron deposits in erythrocytes. Mitochondrial iron accumulation is the hallmark of sideroblastic anemias, which typically result from defects in heme biosynthesis or other pathways that lead to abnormal erythroid mitochondrial iron utilization. To clone the f gene, we used positional cloning techniques, and identified a frameshift mutation in a mitochondrial transmembrane protein. The mutated gene, Sfxn1, is the prototype of a novel family of evolutionarily conserved proteins present in eukaryotes. PMID- 11274052 TI - Naked cuticle targets dishevelled to antagonize Wnt signal transduction. AB - In Drosophila embryos the protein Naked cuticle (Nkd) limits the effects of the Wnt signal Wingless (Wg) during early segmentation. nkd loss of function results in segment polarity defects and embryonic death, but how nkd affects Wnt signaling is unknown. Using ectopic expression, we find that Nkd affects, in a cell-autonomous manner, a transduction step between the Wnt signaling components Dishevelled (Dsh) and Zeste-white 3 kinase (Zw3). Zw3 is essential for repressing Wg target-gene transcription in the absence of a Wg signal, and the role of Wg is to relieve this inhibition. Our double-mutant analysis shows that, in contrast to Zw3, Nkd acts when the Wg pathway is active to restrain signal transduction. Yeast two hybrid and in vitro experiments indicate that Nkd directly binds to the basic-PDZ region of Dsh. Specially timed Nkd overexpression is capable of abolishing Dsh function in a distinct signaling pathway that controls planar-cell polarity. Our results suggest that Nkd acts directly through Dsh to limit Wg activity and thus determines how efficiently Wnt signals stabilize Armadillo (Arm)/beta-catenin and activate downstream genes. PMID- 11274053 TI - Regulation of DAF-2 receptor signaling by human insulin and ins-1, a member of the unusually large and diverse C. elegans insulin gene family. AB - The activity of the DAF-2 insulin-like receptor is required for Caenorhabditis elegans reproductive growth and normal adult life span. Informatic analysis identified 37 C. elegans genes predicted to encode insulin-like peptides. Many of these genes are divergent insulin superfamily members, and many are clustered, indicating recent diversification of the family. The ins genes are primarily expressed in neurons, including sensory neurons, a subset of which are required for reproductive development. Structural predictions and likely C-peptide cleavage sites typical of mammalian insulins suggest that ins-1 is most closely related to insulin. Overexpression of ins-1, or expression of human insulin under the control of ins-1 regulatory sequences, causes partially penetrant arrest at the dauer stage and enhances dauer arrest in weak daf-2 mutants, suggesting that INS-1 and human insulin antagonize DAF-2 insulin-like signaling. A deletion of the ins-1 coding region does not enhance or suppress dauer arrest, indicating a functional redundancy among the 37 ins genes. Of five other ins genes tested, the only other one bearing a predicted C peptide also antagonizes daf-2 signaling, whereas four ins genes without a C peptide do not, indicating functional diversity within the ins family. PMID- 11274054 TI - An SH2-domain-containing kinase negatively regulates the phosphatidylinositol-3 kinase pathway. AB - SHK1 is a novel dual-specificity kinase that contains an SH2 domain in its C terminal region. We demonstrate that SHK1 is required for proper chemotaxis and phagocytosis. Mutant shk1 null cells lack polarity, move very slowly, and exhibit an elevated and temporally extended chemoattractant-mediated activation of the kinase Akt/PKB. GFP fusions of the PH domain of Akt/PKB or the PH-domain containing protein CRAC, which become transiently associated with the plasma membrane after a global stimulation with a chemoattractant, remain associated with the plasma membrane for an extended period of time in shk1 null cells. These results suggest that SHK1 is a negative regulator of the PI3K (phosphatidylinositol-3 kinase) pathway. Furthermore, when a chemoattractant gradient is applied to a wild-type cell, these PH-domain-containing proteins and the F-actin-binding protein coronin localize to its leading edge, but in an shk1 null cell they become randomly associated with the plasma membrane and cortex, irrespective of the direction of the chemoattractant gradient, suggesting that SHK1 is required for the proper spatiotemporal control of F-actin levels in chemotaxing cells. Consistent with such functions, SHK1 is localized at the plasma membrane/cortex, and we show that its SH2 domain is required for this localization and the proper function of SHK1. PMID- 11274055 TI - Mitogen-activated protein kinase phosphatase is required for genotoxic stress relief in Arabidopsis. AB - Genotoxic stress activates complex cellular responses allowing for the repair of DNA damage and proper cell recovery. Although plants are exposed constantly to increasing solar UV irradiation, the signaling cascades activated by genotoxic environments are largely unknown. We have identified an Arabidopsis mutant (mkp1) hypersensitive to genotoxic stress treatments (UV-C and methyl methanesulphonate) due to disruption of a gene that encodes an Arabidopsis homolog of mitogen activated protein kinase phosphatase (AtMKP1). Growth of the mkp1 mutant under standard conditions is indistinguishable from wild type, indicating a stress specific function of AtMKP1. MAP kinase phosphatases (MKPs), the potent inactivators of MAP kinases, are considered important regulators of MAP kinase signaling. Although biochemical data from mammalian cell cultures suggests an involvement of MKPs in cellular stress responses, there is no in vivo genetic support for this view in any multicellular organism. The genetic and biochemical data presented here imply a central role for a MAP kinase cascade in genotoxic stress signaling in plants and indicate AtMKP1 to be a crucial regulator of the MAP kinase activity in vivo, determining the outcome of the cellular reaction and the level of genotoxic resistance. PMID- 11274056 TI - Closely related proteins MBD2 and MBD3 play distinctive but interacting roles in mouse development. AB - MBD2 and MBD3 are closely related proteins with consensus methyl-CpG binding domains. MBD2 is a transcriptional repressor that specifically binds to methylated DNA and is a component of the MeCP1 protein complex. In contrast, MBD3 fails to bind methylated DNA in murine cells, and is a component of the Mi-2/NuRD corepressor complex. We show by gene targeting that the two proteins are not functionally redundant in mice, as Mbd3-/- mice die during early embryogenesis, whereas Mbd2-/- mice are viable and fertile. Maternal behavior of Mbd2-/- mice is however defective and, at the molecular level, Mbd2-/- mice lack a component of MeCP1. Mbd2-mutant cells fail to fully silence transcription from exogenous methylated templates, but inappropriate activation of endogenous imprinted genes or retroviral sequences was not detected. Despite their differences, Mbd3 and Mbd2 interact genetically suggesting a functional relationship. Genetic and biochemical data together favor the view that MBD3 is a key component of the Mi 2/NuRD corepressor complex, whereas MBD2 may be one of several factors that can recruit this complex to DNA. PMID- 11274057 TI - hMSH3 and hMSH6 interact with PCNA and colocalize with it to replication foci. AB - Proliferating cell nuclear antigen (PCNA) has been implicated in eukaryotic postreplicative mismatch correction, but the nature of its interaction with the repair machinery remained enigmatic. We now show that PCNA binds to the human mismatch binding factors hMutSalpha and hMutSbeta via their hMSH6 and hMSH3 subunits, respectively. The N-terminal domains of both proteins contain the highly conserved PCNA-binding motif Qxx[LI]xx[FF]. A variant of hMutSalpha, lacking this motif because of deletion of 77 N-terminal residues of the hMSH6 subunit, no longer was able to interact with PCNA in vitro and failed to restore mismatch repair in hMSH6-deficient cells. Colocalization of PCNA and hMSH6 or hMSH3 to replication foci implies an intimate link between replication and mismatch correction. We postulate that PCNA plays a role in repair initiation by guiding the mismatch repair proteins to free termini in the newly replicated DNA strands. PMID- 11274058 TI - Tn7 recognizes transposition target structures associated with DNA replication using the DNA-binding protein TnsE. AB - We report that the bacterial transposon Tn7 selects targets by recognizing features associated with DNA replication using the transposon-encoded DNA-binding protein TnsE. We show that Tn7 transposition directed by TnsE occurs in one orientation with respect to chromosomal DNA replication, indicating that a structure or complex involved in DNA replication is likely to be a critical determinant of TnsE insertion. We find that mutant TnsE proteins that allow higher levels of transposition also bind DNA better than the wild-type protein. The increased binding affinity displayed by the TnsE high-activity mutants indicates that DNA binding is relevant to transposition activity and suggests that TnsE interacts directly with target DNAs. In vitro, TnsE interacts preferentially with certain DNA structures, indicating a mechanism for the TnsE mediated orientation and insertion preference. The pattern of TnsE-mediated insertion events around the Escherichia coli chromosome provides insight into how DNA replication forks proceed in vivo. PMID- 11274059 TI - Topoisomerase IV, alone, unknots DNA in E. coli. AB - Knotted DNA has potentially devastating effects on cells. By using two site specific recombination systems, we tied all biologically significant simple DNA knots in Escherichia coli. When topoisomerase IV activity was blocked, either with a drug or in a temperature-sensitive mutant, the knotted recombination intermediates accumulated whether or not gyrase was active. In contrast to its decatenation activity, which is strongly affected by DNA supercoiling, topoisomerase IV unknotted DNA independently of supercoiling. This differential supercoiling effect held true regardless of the relative sizes of the catenanes and knots. Finally, topoisomerase IV unknotted DNA equally well when DNA replication was blocked with hydroxyurea. We conclude that topoisomerase IV, not gyrase, unknots DNA and that it is able to access DNA in the cell freely. With these results, it is now possible to assign completely the topological roles of the topoisomerases in E. coli. It is clear that the topoisomerases in the cell have distinct and nonoverlapping roles. Consequently, our results suggest limitations in assigning a physiological function to a protein based upon sequence similarity or even upon in vitro biochemical activity. PMID- 11274060 TI - Drosophila Brain Tumor is a translational repressor. AB - The Drosophila brain tumor (brat) gene encodes a member of the conserved NHL family of proteins, which appear to regulate differentiation and growth in a variety of organisms. One of the founding family members, Caenorhabditis elegans LIN-41, is thought to control posttranscriptional gene expression. However, the mechanism by which LIN-41, or any other NHL protein, acts has not been clear. Using a yeast "four-hybrid" interaction assay, we show that Brain Tumor is recruited to hunchback (hb) mRNA through interactions with Nanos and Pumilio, which bind to the RNA to repress its translation. Interaction with the Nanos/Pumilio/RNA complex is mediated by the Brat NHL domain; single amino acid substitutions in this domain compromise quaternary complex assembly in vitro and hb regulation in vivo. Thus, recruitment of Brat is necessary for translational repression and the normal development of posterior embryonic pattern. In addition to regulating abdominal segmentation, previous genetic analysis has shown that Brat, Nanos, and Pumilio govern a variety of developmental processes. We examined the role of Brat in two of these processes-regulation of maternal Cyclin B mRNA in the embryo and regulation of imaginal disc development. The results of these experiments suggest that NHL domain proteins are recruited to various mRNAs by combinatorial protein-protein interactions. PMID- 11274063 TI - High-resolution ultrasonic imaging and characterization of the ciliary body. AB - PURPOSE: To develop a means for noninvasive in vivo visualization of the ciliary processes using very-high-frequency (50 MHz) ultrasound and to develop quantitative morphologic descriptors that may relate to physiologic function. METHODS: The region of the ciliary body was scanned with very-high-frequency ultrasound, both in rabbits and in normal human subjects. Data were acquired in a series of planes so that the spacing between them was less than the beam width of the transducer in its focal plane. Three-dimensional perspective images were constructed, representing the anatomy of the angle region, including the ciliary processes. The automatically detected boundaries of the ciliary processes were analyzed to compute their periphery, area, shape factor, and fractal dimension. These measures were compared between the human and the rabbit eye and analyzed for periodicities related to the spacing of successive processes. RESULTS: Three dimensional images allowed visualization of the radial arrangement of the processes. All biometric descriptors were significantly different between the rabbit and human eye and showed periodicities consistent with spacing between processes. CONCLUSIONS: The methods described in this report are sensitive descriptors of the state of the ciliary processes. These techniques may be of value in measurement of changes in the ciliary body associated with disease, medical therapy, and aging. PMID- 11274061 TI - A novel embryonic poly(A) binding protein, ePAB, regulates mRNA deadenylation in Xenopus egg extracts. AB - An in vitro system that recapitulates the in vivo effect of AU-rich elements (AREs) on mRNA deadenylation has been developed from Xenopus activated egg extracts. ARE-mediated deadenylation is uncoupled from mRNA body decay, and the rate of deadenylation increases with the number of tandem AUUUAs. A novel ARE binding protein called ePAB (for embryonic poly(A)-binding protein) has been purified from this extract by ARE affinity selection. ePAB exhibits 72% identity to mammalian and Xenopus PABP1 and is the predominant poly(A)-binding protein expressed in the stage VI oocyte and during Xenopus early development. Immunodepletion of ePAB increases the rate of both ARE-mediated and default deadenylation in vitro. In contrast, addition of even a small excess of ePAB inhibits deadenylation, demonstrating that the ePAB concentration is critical for determining the rate of ARE-mediated deadenylation. These data argue that ePAB is the poly(A)-binding protein responsible for stabilization of poly(A) tails and is thus a potential regulator of mRNA deadenylation and translation during early development. PMID- 11274064 TI - Lack of blood-brain barrier properties in microvessels of the prelaminar optic nerve head. AB - PURPOSE: To define the blood-brain barrier (BBB) characteristics of microvessels in the optic nerve head (ONH). METHODS: Immunohistochemical staining of different regions of the ONH, retro-laminar optic nerve, and retina of human and monkey eyes was carried out, using antibodies against BBB markers (glucose transporter 1, transferrin receptor, and P-glycoprotein), the non-BBB marker PAL-E, and against plasma proteins fibrinogen and IgG, which serve as endogenous markers of nonspecific microvascular permeability. In the ONH of monkey eyes, the number of transport-related endothelial pinocytotic vesicles and their cellular distribution within the microvessels were determined by electron microscopy. RESULTS: In both human and monkey eyes, only microvessels in the prelaminar region of the ONH were positive for the PAL-E antigen. The prelaminar region microvessels showed either no or weak expression of the transferrin receptor and P-glycoprotein but stained positive for glucose transporter 1. In human ONH, fibrinogen and IgG were present around microvessels in the prelaminar region but not in other parts of the optic nerve or retina. By electron microscopy, endothelial cells of prelaminar region microvessels contained a higher number of pinocytotic vesicles, located at the luminal and abluminal side of the endothelial cell membrane, in contrast to a mainly abluminal localization in microvessels of the retina and other parts of the optic nerve. CONCLUSIONS: Microvessels in the prelaminar region of the ONH lack classical BBB characteristics and display nonspecific permeability, possibly mediated by vesicular transport. PMID- 11274062 TI - Inhibition of touch cell fate by egl-44 and egl-46 in C. elegans. AB - In wild-type Caenorhabditis elegans, six cells develop as receptors for gentle touch. In egl-44 and egl-46 mutants, two other neurons, the FLP cells, express touch receptor-like features. egl-44 and egl-46 also affect the differentiation of other neurons including the HSN neurons, two cells needed for egg laying. egl 44 encodes a member of the transcription enhancer factor family. The product of the egl-46 gene, two Drosophila proteins, and two proteins in human and mice define a new family of zinc finger proteins. Both egl-44 and egl-46 are expressed in FLP and HSN neurons (and other cells); expression of egl-46 is dependent on egl-44 in the FLP cells but not in the HSN cells. Wild-type touch cells express egl-46 but not egl-44. Moreover, ectopic expression of egl-44 in the touch cells prevents touch cell differentiation in an egl-46-dependent manner. The sequences of these genes and their nuclear location as seen with GFP fusions indicate that they repress transcription of touch cell characteristics in the FLP cells. PMID- 11274065 TI - Ultrastructure and composition of asteroid bodies. AB - PURPOSE: Asteroid hyalosis is a disease of the vitreous, characterized by brilliant reflecting particles, termed asteroid bodies, which are surrounded by a tightly adhering network of fibrils. The composition and mode of formation of asteroid bodies is not yet understood in detail. The purpose of this study was to investigate the ultrastructure of asteroid bodies and to identify the intrinsic inorganic and organic components that contribute to the nature and development of asteroid bodies. METHODS: Electron energy loss spectroscopy and energy-filtered transmission electron microscopy were used for the elemental analysis of asteroid bodies. The ultrastructural localization of glycosaminoglycans was investigated, using lectin and antibody conjugates in conjunction with transmission electron microscopy and epifluorescence microscopy. Anionic sites of glycosaminoglycans were detected with 15 nm cationic colloidal gold at low pH, applied as a postembedding technique. Ultrastructural details of asteroid bodies were documented using fast Fourier transform analysis of zero-loss filtered images. RESULTS: Element mapping of asteroid bodies by electron spectroscopic imaging revealed a homogeneous distribution of calcium, phosphorus, and oxygen. The electron energy loss spectra of these elements showed details similar to those found for hydroxyapatite. Additionally, high contrast and sensitivity against a calcium-specific chelator highlighted the crystalline, apatite-like nature of asteroid bodies. Immunofluorescence microscopy revealed the presence of chondroitin-6-sulfate at the periphery of asteroid bodies, which is in agreement with the ultrastructural colocalization of anionic sites. Fast Fourier transform analysis revealed that each 7-nm periodicity of asteroid lamellar stacks is divided by a fine, parallel-oriented line, separating each 7-nm layer into two halves of 3.5-nm thickness. Carbohydrates specific for hyaluronic acid were observed by lectin-gold labeling to be part of the inner matrix of asteroid bodies. CONCLUSIONS: The results of this study demonstrate the structural and elemental similarity of asteroid bodies with hydroxyapatite. Proteoglycans and their glycosaminoglycan side chains are implicated in playing a role in regulating the biomineralization process. PMID- 11274066 TI - Blindness in the Indian state of Andhra Pradesh. AB - PURPOSE: To determine the current prevalence and causes of blindness in the Indian state of Andhra Pradesh to assess if blindness has decreased since the last survey of 1986-1989. METHODS: A population-based epidemiology study, using a stratified, random, cluster, systematic sampling strategy, was conducted in the state of Andhra Pradesh in India. Participants of all ages (n = 10,293), 87.3% of the 11,786 eligible, from 94 clusters in one urban and three rural areas representative of the population of Andhra Pradesh, underwent interview and a detailed dilated ocular evaluation by trained professionals. Blindness was defined as presenting distance visual acuity < 6/60 or central visual field < 20(o) in the better eye. RESULTS: Two hundred seventy-five participants were blind, a prevalence of 1.84% (95% confidence interval, 1.49%-2.19%) when adjusted for the age, sex, and urban-rural distribution of the population in 2000. The causes of this blindness were easily treatable in 60.3% (cataract, 44%; refractive error, 16.3%). Preventable corneal disease, glaucoma, complications of cataract surgery, and amblyopia caused another 19% of the blindness. Blindness was more likely with increasing age and decreasing socioeconomic status, and in female subjects and in rural areas. Among the 76 million population of Andhra Pradesh, 714,400 are estimated to have cataract-related blindness (615,600 cataract, 53,200 cataract surgery-related complications, 45,600 aphakia), and 228,000 refractive error-related blindness (159,600 myopia, 22,800 hyperopia, 45,600 refractive error-related amblyopia). If 95% of the cataract and refractive error blindness in Andhra Pradesh had been treated effectively, 3.4 and 7.4 million blind-person-years, respectively, could have been prevented. If 90% of the blindness due to preventable corneal disease and glaucoma had been prevented, another 2.7 million blind-person-years could have been prevented. CONCLUSIONS: The prevalence of blindness in this Indian state has increased from 1.5% in the late 1980s to 1.84% currently, as against the target of the National Program for Control of Blindness to reduce the prevalence to 0.3% by 2000. The number of people with cataract-related blindness has not reduced even with the eye care policy focus on cataract. Reduction of blindness in India will require strategies that are more effective than those that have been pursued so far. PMID- 11274067 TI - Comparison of preschool vision screening methods in a population with a high prevalence of astigmatism. AB - PURPOSE: To compare the effectiveness of four methods of screening 3- to 5-year old children for astigmatism high enough to require spectacle correction. METHODS: Lea Symbols Visual Acuity Screening (LSVAS), MTI Photoscreening (MTIPS), Nidek KM-500 Keratometry Screening (KERS), and Retinomax K-Plus Noncycloplegic Autorefraction Screening (NCARS) were attempted on 379 preschool children who are members of a Native American tribe having a high prevalence of astigmatism that is primarily corneal in origin. The need for spectacle correction was determined by cycloplegic refraction. Receiver Operating Characteristic (ROC) curves were fit, confidence intervals were determined, and area under the curves was compared. RESULTS: Astigmatism > or = 1.00 D was present in the right eye of 47.5% and in the left eye of 48.0% of children. Spectacles were prescribed for children < 48 months of age who had cylinder > or = 2.00 D and children > or = 48 months who had cylinder > or = 1.50 D, with the result that 33% of subjects required spectacles. Area under the ROC curve was 0.98 for NCARS, 0.92 for KERS, 0.78 for MTIPS, and 0.70 for LSVAS, and each of these values differed significantly from the other three (all P < 0.007). Testability was significantly higher for NCARS (99.5%) and KERS (99.7%) than for MTIPS (93.5%) and LSVAS (92.0%). CONCLUSIONS: In a population that included many children with astigmatism, objective, fully automated screening methods (NCARS and KERS) were superior to both visual acuity screening and photoscreening with subjective interpretation in identifying children who had astigmatism requiring spectacle correction. PMID- 11274068 TI - Impaired neurotransmitter release from lacrimal and salivary gland nerves of a murine model of Sjogren's syndrome. AB - PURPOSE: To determine whether lacrimal and salivary gland nerves of an animal model of Sjogren's syndrome, the MRL/lpr mouse, are able to release acetylcholine. The second purpose was to determine whether activation of the lacrimal gland nerves of the MRL/lpr mouse leads to protein secretion. METHODS: Total saliva was collected for 10 minutes from the oral cavity of male and female MRL/lpr and MRL/+ mice, after intraperitoneal stimulation with pilocarpine and isoproterenol. Lacrimal and salivary gland lobules prepared from 18-week-old MRL/lpr and MRL/+ mice were incubated in the presence of depolarizing KCl (75 mM) solution. Acetylcholine release and peroxidase secretion (a protein secreted by the lacrimal gland) were measured using a spectrofluorometric assay. RESULTS: Female, but not male, MRL/lpr mouse salivary glands were hyper-responsive to in vivo injection of secretagogues. These mice produced significantly higher amounts of saliva than did age-matched MRL/+ mice. Lacrimal and salivary gland nerves from 18-week-old MRL/+ mice released acetylcholine in response to a depolarizing KCl solution. In contrast, nerves in glands from 18-week-old MRL/lpr mice did not increase acetylcholine release in response to the depolarizing solution. Moreover, lacrimal glands from 18-week-old MRL/+ mice were able to secrete peroxidase in response to a depolarizing KCl solution, whereas those from 18-week old MRL/lpr could not. This was not due to a defect in the secretory process, because addition of an exogenous secretagogue elicited peroxidase secretion from 18-week-old MRL/lpr as well as MRL/+ mice lacrimal glands. CONCLUSIONS: The results show that activation of nerves of lacrimal and salivary glands infiltrated with lymphocytes does not increase the release of neurotransmitters, which results in impaired secretion from these glands. PMID- 11274069 TI - Regulation of a Rho-associated kinase expression during the corneal epithelial cell cycle. AB - PURPOSE: It has been recognized that an increased expression of the Rho associated kinase (ROCK-I), a downstream target of Rho (a Ras-related small guanosine triphosphatase [GTPase]), is associated with limbal-to-corneal epithelial transition. The purpose of the present study was to determine whether the expression of ROCK-I is regulated during the cell cycle of corneal epithelial cells. METHODS: Rabbit corneal epithelial cells in culture were subjected to different culture conditions to enrich them in the G0, G1, and S phases of the cell cycle. Indirect immunofluorescence staining and western blot techniques were used for analyzing the changes in the relative intracellular concentrations of ROCK-I. Northern blot analysis of the isolated cellular RNA was performed to estimate the relative concentrations of ROCK-I mRNA. RESULTS: Serum deprivation did not cause all the corneal epithelial cells in culture to be arrested in the G0 phase of the cell cycle. However, the cells could be arrested in G0 by treating them with culture medium supplemented with transforming growth factor (TGF)-beta1. The relative concentration of ROCK-I in the G0-arrested cells was higher than in the corresponding control untreated cultures. G0-arrested cells were induced to enter G1, followed by the S phase of the cell cycle, by refeeding them with the medium devoid of TGF-beta1. The total intracellular concentration of ROCK-I significantly decreased during the G1 phase of the cell cycle and increased again during the S phase. The decrease in intracellular ROCK-I during the G1 phase was confirmed by arresting the cells in G1 with isoleucine deprivation and thymidine-mimosine treatments. ROCK-I mRNA levels were also found to be decreased during the G1 phase of the cell cycle. CONCLUSIONS: The levels of ROCK-I in the corneal epithelial cells were significantly lower in the G1 phase than those in the S and G0 phases of the cell cycle. Therefore, a Rho signaling pathway(s) involving ROCK-I may be regulated during the corneal epithelial cell cycle. The downregulation of ROCK-I during the G1 phase, at least in part, is due to the decreased levels of its mRNA. Based on these findings, ROCK-I may have a role in the progression of the cell cycle in the corneal epithelial cells as they migrate centripetally from the limbal to the corneal surface. PMID- 11274070 TI - Role of the small GTP-binding protein rho in epithelial cell migration in the rabbit cornea. AB - PURPOSE: To determine the role of the small guanosine triphosphate (GTP)-binding protein Rho in the migration of corneal epithelial cells. METHODS: The presence of the Rho target proteins Rho-associated coiled coil-containing protein kinase (ROCK)-1 and ROCK-2 in rabbit cornea was examined by immunohistochemical analysis, and that of the corresponding mRNAs in rabbit corneal epithelial cells was determined by reverse transcription-polymerase chain reaction analysis. The effects of various agents on epithelial cell migration were investigated by measuring the length of the migration path in rabbit corneal blocks in culture. RESULTS: Both ROCK-1 and ROCK-2 were detected in the rabbit corneal epithelium at both protein and mRNA levels. The Rho activator lysophosphatidic acid (LPA) stimulated corneal epithelial migration in a dose-dependent manner, whereas exoenzyme C3, a Rho inhibitor, inhibited epithelial migration also in a dose dependent manner. The stimulatory effect of LPA on corneal epithelial migration was prevented by exoenzyme C3. Both cytochalasin B, an inhibitor of actin filament assembly, and ML-7, an inhibitor of myosin light chain kinase, also prevented LPA stimulation of epithelial migration. CONCLUSIONS: These results suggest that Rho mediates corneal epithelial migration in response to external stimuli by regulating the organization of the actin cytoskeleton. PMID- 11274071 TI - Toxicity of natural tear substitutes in a fully defined culture model of human corneal epithelial cells. AB - PURPOSE: Serum and saliva have recently been advocated as natural tear substitutes for intractable aqueous-deficient dry eyes, but the effects of these fluids on corneal epithelium have not been well characterized. A laboratory study was performed in a defined test model to compare the toxicity of natural and pharmaceutical tear substitutes and to identify potentially toxic factors in natural tear substitutes, such as amylase, hypotonicity, and variations in preparation. METHODS: Primary human corneal epithelial cells were cultured with defined keratinocyte serum-free medium. The cells were incubated with hypromellose (hydroxypropylmethylcellulose 0.3%) with and without benzalkonium chloride 0.01%, saliva with differing osmolalities, 100% serum, and 50% serum (1:1 vol/vol with chloramphenicol 0.5%) for varying times and concentrations. Toxicity was examined in four ways. Microvillous density was assessed with scanning electron microscopy. Cell membrane permeability and intracellular esterase activity were analyzed after staining with fluorescent calcein AM/ethidium homodimer and cellular adenosine triphosphate (ATP) was quantified using a luciferin-luciferase-based assay. RESULTS: The toxicity ranking of the tear substitutes correlated in all assays. The ATP assay was the most sensitive, followed by ethidium cell permeability, and finally the esterase activity. Preserved hypromellose was more toxic than the unpreserved preparation. Among natural tear substitutes, natural saliva was most toxic. Isotonic saliva and 50% serum were of similar toxicity, and 100% serum was least toxic. Natural tear substitutes were-except for natural saliva-less toxic than unpreserved hypromellose. Hypotonicity, but not amylase, was the major toxic effect associated with saliva. The dilution of serum with chloramphenicol induced toxicity. CONCLUSIONS: This is the first toxicity study using human primary corneal epithelial cells cultured under fully defined conditions as an in vitro model. Cellular ATP is a sensitive parameter for quantifying toxicity. Isotonic saliva and serum offer greater therapeutic potential for severely aqueous deficient dry eyes than do pharmaceutical tear substitutes. PMID- 11274072 TI - Comparison of pupil perimetry and visual perimetry in normal eyes: decibel sensitivity and variability. AB - PURPOSE: To compare the sensitivity and variability of pupil perimetry with visual perimetry at the same retinal locations in normal subjects. METHODS: Pupil perimetry was performed on the right and left eyes of 10 normal subjects using a computerized infrared pupillometer equipped to present perimetric light stimuli and record pupil light reflexes. Eleven locations were tested at different intensities along the horizontal meridian of each eye, and the decibel sensitivity of the pupil light reflex was compared with the visual threshold at the same location. RESULTS: The shape and height of the hill of vision (retinal sensitivity) was very similar between the right and left eyes of each individual using either pupil perimetry (R2 = 0.69) or standard threshold perimetry (R2 = 0.62) but was less similar between subjects. Comparisons between pupil and visual sensitivity revealed a lack of correlation at the same retinal location in normal eyes (R2 = 0.19). CONCLUSIONS: The high intereye correlation for either pupil or visual sensitivity may provide an important tool for detecting focal or asymmetric visual field damage. Although the basic shape of the sensitivity profile of pupil and visual responses was similar under the conditions of testing, the two did not correlate well within each eye among the normal subjects. This highlights that similarities do exist in the sensitivity profile of the two pathways, but they do not seem to vary in the same proportion between normal individuals. PMID- 11274073 TI - BDNF enhances retinal ganglion cell survival in cats with optic nerve damage. AB - PURPOSE: To determine whether brain-derived neurotrophic factor (BDNF), a neuroprotectant in the small rat eye, might also serve as an effective neuroprotectant in larger vertebrate eyes. METHODS. A cat optic nerve crush model was combined with standard histologic staining and analysis techniques. Twenty nine animals were studied, with the noninjected eye serving as the control eye. RESULTS: No treatment, or intravitreal injection of sterile water, resulted in an approximately 50% loss of ganglion cells 1 week after nerve crush. By contrast, the mean percentages of surviving ganglion cells measured in eyes receiving injections of 15, 30, 60, and 90 microg BDNF at the time of the nerve damage were 52%, 81%, 77%, and 70%, respectively. Similar values were obtained for ganglion cell density. Cell size measurements suggest a complex response among the different classes of cat ganglion cells; 30 microg BDNF treatment retained the highest number of large ganglion cells, whereas 90 microg minimized the loss of medium-sized neurons and retained normal proportions of large, medium, and small ganglion cells. CONCLUSIONS: The data show that BDNF is an effective neuroprotectant in primate-sized eyes after optic nerve injury. Although the amount required to achieve neuroprotection is much greater than that needed for the small rat eye (30 microg versus 0.5 microg), when differences in vitreal volume are considered, the effective dose is similar (0.01 microg BDNF/microl vitreal volume). High doses of BDNF induce inflammation and result in a decrease in total ganglion cell survival but appear necessary to save medium-sized neurons, which are affected most severely by nerve injury. PMID- 11274074 TI - Optic nerve transection in monkeys may result in secondary degeneration of retinal ganglion cells. AB - PURPOSE: Interest in neuroprotection for optic neuropathies is, in part, based on the assumption that retinal ganglion cells (RGCs) die, not only as a result of direct (primary) injury, but also indirectly as a result of negative effects from neighboring dying RGCs (secondary degeneration). This experiment was designed to test whether secondary RGC degeneration occurs after orbital optic nerve injury in monkeys. METHODS: The superior one third of the orbital optic nerve on one side was transected in eight cynomolgus monkeys (Macaca fascicularis). Twelve weeks after the partial transection, the number of RGC bodies in the superior and inferior halves of the retina of the experimental and control eyes and the number and diameter of axons in the optic nerve were compared by detailed histomorphometry. Vitreous was obtained for amino acid analysis. A sham operation was performed in three additional monkeys. RESULTS: Transection caused loss of 55% +/- 13% of RGC bodies in the superior retina of experimental compared with fellow control eyes (mean +/- SD, t-test, P < 0.00,001, n = 7). Inferior RGCs, not directly injured by transection, decreased by 22% +/- 10% (P = 0.002). The loss of superior optic nerve axons was 83% +/- 12% (mean +/- SD, t-test, P = 0.0008, n = 5) whereas, the inferior loss was 34% +/- 20% (P = 0.02, n = 5). Intravitreal levels of glutamate and other amino acids in eyes with transected nerves were not different from levels in control eyes 12 weeks after injury. Fundus examination, fluorescein angiography, and histologic evaluation confirmed that there was no vascular compromise to retinal tissues by the transection procedure. CONCLUSIONS: This experiment suggests that primary RGC death due to optic nerve injury is associated with secondary death of surrounding RGCs that are not directly injured. PMID- 11274075 TI - The presence and properties of myocilin in the aqueous humor. AB - PURPOSE: To determine whether myocilin is present in the aqueous humor (AH) and to examine certain properties of this protein. METHODS: Human AH was obtained at the time of either glaucoma surgery or cataract extraction. Monkey AH was obtained at the time of death, and bovine aqueous was obtained from eyes delivered from an abattoir. Column chromatography was performed on aqueous samples to determine the approximate size of the myocilin present. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blot analysis were performed using antibody prepared against a peptide sequence in myocilin. Analysis of the bovine proteins present in AH that were retained by a microporous filter was also performed using western blot analysis. RESULTS: By western blot analysis, myocilin was present in human, monkey, and bovine AH. The apparent molecular size of the myocilin present in the AH were greater than 250,000 Da, when quantified with a gel filtration column. Myocilin appeared to be hydrophobic and was one of the proteins that was retained on microporous filters that were obstructed by AH. CONCLUSIONS: Myocilin is a constituent in the AH. It appears that myocilin is a hydrophobic protein that may exist in an oligomeric state or in association with other proteins. Myocilin is retained by microporous filters and may be involved in the obstruction of these filters that occurs when AH is perfused through them. PMID- 11274077 TI - The relative contribution of mast cell subsets to conjunctival TH2-like cytokines. AB - PURPOSE: To investigate the distribution of the T-helper (TH)2-like cytokines, interleukin (IL)-4, IL-5, IL-6, and IL-13 between mast cell subsets in conjunctival biopsy specimens from normal subjects and those with seasonal allergic conjunctivitis (SAC) during and outside of the grass pollen season. METHODS: Sequential and double in situ hybridization (ISH) and immunohistochemistry (IHC) were performed on thin sections of human conjunctiva to determine the colocalization of the immunoreactivity of IL-4, IL-5, IL-6, and IL-13 to mast cell subsets in normal subjects and subjects with atopy and to detect IL-4 mRNA in conjunctival mast cells. RESULTS: More than 90% of IL-4+ immunoreactive cells were observed to be mast cells in conjunctival biopsy specimens from all patient groups. The majority of IL-5+, IL-6+, and IL-13+ cells were also noted to be mast cells for each group. IL-4 preferentially colocalized to the tryptase+-chymase+ mast cell phenotype (MC(TC)) with MC(TC) cells comprising 93.3% of cytokine+ mast cells in symptomatic SAC (P = 0.0017), 89.2% in asymptomatic SAC (P = 0.0008), and 77.8% in normal subjects (P = 0.0472). IL 13 appeared to colocalize preferentially to the MC(TC) phenotype and IL-5 and IL 6 to the MC(T) phenotype. ISH showed that 75.8% of mast cells in normal subjects, 78.7% in subjects with symptomatic SAC, and 18.7% in subjects with asymptomatic SAC expressed mRNA for IL-4. CONCLUSIONS: Conjunctival mast cells are an important source of IL-4, IL-5, IL-6, and IL-13 immunoreactivity, with preferential colocalization of IL-4 and IL-13 on the MC(TC) subset and IL-5 and IL-6 to the MC(T) subset. This evidence suggests that differences in protease phenotype may also reflect functional differences evidenced by the different patterns of cytokine distribution. PMID- 11274076 TI - Blockade of CD40-CD154 costimulatory pathway promotes survival of allogeneic corneal transplants. AB - PURPOSE: To determine the effect of systemic anti-CD154 monoclonal antibody on the survival of orthotopic murine corneal transplants. METHODS: BALB/c mice were used as recipients of syngeneic, multiple minor histocompatability (H)-disparate, or major histocompatibility complex MHC-mismatched corneal transplants. Recipient beds were either avascular (normal risk) or neovascularized (high risk). Mice were randomized to receive either anti-CD154 antibody or control immunoglobulin by intraperitoneal injection at surgery and once weekly after surgery. After orthotopic corneal transplantation, all grafts were evaluated for signs of rejection by slit lamp biomicroscopy over 8 weeks. The high-risk transplants were continuously observed until week 18 after the therapy was discontinued at week 8. Allospecific delayed-type hypersensitivity (DTH) was evaluated after transplantation in high-risk graft recipients. Frequency of interferon (IFN) gamma-secreting T cells in the hosts was measured by enzyme-linked immunospot (ELISPOT) assay. RESULTS: In normal-risk transplantation, the 8-week survival rate improved from 25% in control mice to 88% in anti-CD154-treated hosts of minor H-disparate grafts (P = 0.0087) and from 78% in control mice to 100% in anti-CD154-treated recipients of MHC-mismatched transplants (P = 0.177). Of particular significance, in high-risk transplantation, anti-CD154 therapy dramatically enhanced the survival of both minor H- and MHC-disparate corneal transplants to 100% (P = 0.0001) and 92% (P = 0.0002), respectively. In addition, the anti-CD154-treated mice did not exhibit allospecific immunity. However, termination of anti-CD154 led to some loss in graft survival, especially among high-risk minor H-disparate grafts. The frequency of IFN-gamma-producing T cells was significantly reduced in anti-CD154-treated hosts. CONCLUSIONS: Continuous suppression of the CD40-CD154 costimulatory pathway promotes the acceptance of corneal transplants, regardless of the degree of allodisparity or preoperative risk. The beneficial effect of anti-CD154 treatment may be due in part to inhibition of Th1-mediated responses. PMID- 11274078 TI - Regulation of thioltransferase expression in human lens epithelial cells. AB - PURPOSE: To study how the expression of thioltransferase (TTase), a critical thiol repair and dethiolating enzyme, is regulated in human lens epithelial cells under oxidative stress. Also to examine whether depleting the primary cellular antioxidant glutathione (GSH) in these cells has any influence on TTase expression under the same conditions. METHODS: Human lens epithelial cells (B3) were grown to confluence (1.6 million) and gradually weaned from serum in the medium before exposing to 0.1 mM H2O2 for 2 hours. Cells were removed at the time intervals of 0, 5, 10, 15, 30, 60, and 120 minutes for protein measurements of GSH and TTase activity and for reverse transcription-polymerase chain reaction (RT-PCR) or Northern hybridization analysis to quantify TTase mRNA. The effect of GSH depletion on TTase mRNA expression was examined by treating the cells with buthionine S,R-sulfoximine (BSO); 1-chloro, 2,4-dinitrobenzene (CDNB); or 1,3-bis (2-chloroethyl)-1-nitrosourea (BCNU). Lens epithelial cells, depleted of cellular GSH by treatment with BCNU, were subjected to oxidative stress to examine the effect on TTase activity and mRNA level. RESULTS: A transient increase was detected in TTase mRNA after 5 minutes of H2O2 treatment. The upregulation reached a maximum of 80% above the normal level by 10 minutes and gradually decreased as the oxidant was detoxified by the cells. Manipulation of cellular GSH level by treatment with BSO, CDNB, and BCNU resulted in a minimum change in TTase expression. It is noteworthy that when cells depleted of GSH were subjected to oxidative stress, TTase expression was also found to be strongly upregulated. CONCLUSIONS: These observations suggest that the upregulation of TTase expression in the lens epithelial cells could be an adaptive response of the cells to combat oxidative stress to restore the vital functions of the lens proteins and enzymes. Such regulation is independent of cellular GSH concentration. PMID- 11274079 TI - Role of the endoplasmic reticulum in shaping calcium dynamics in human lens cells. AB - PURPOSE: Localized cortical cataracts in the human lens have been shown to involve a selective increase in calcium with no change in sodium content. Recent studies in the rat lens in vitro have shown that the store-operated channel is highly selective for calcium over sodium, and therefore this channel was characterized further in human lens cells. METHODS: Human primary cultures were initiated from epithelial explants and passaged onto coverslips. After incorporating Fura-2, agonist- or thapsigargin-induced changes in cytosolic calcium were monitored and calibrated using fluorometric digital imaging techniques. RESULTS: Histamine and adenosine triphosphate (ATP; 10 microM) induced a large transient increase in cytosolic calcium followed by a maintained lower plateau phase in the continued presence of the calcium-signaling agonist. The second phase was abolished by removing external calcium and represented the contribution from the store-operated influx. The store-operated pathway was blocked by inorganic agents such as zinc and nickel (100 microM) but was insensitive to the voltage-sensitive calcium channel blocker, nifedipine (1 mM). Depolarizing the membrane voltage by raising the external potassium (75 mM) also blocked the influx. Similar results were obtained if the store was first emptied directly using thapsigargin (1 microM), and with this agent it was also possible to observe the very slow activation and inactivation kinetics (>10 seconds) of the channel. Addition of manganese to the bathing medium initiated a quench of Fura-2 isobestic fluorescence that was enhanced 2.9 +/- 0.3-fold after 10 microM ATP addition. There was a delay of 82 +/- 16 seconds between initiation of the calcium spike and the Mn2+ quench rate, indicating the presence of a delayed entry pathway. In the resting state, removal of, or increasing extracellular calcium concentration 10-fold did not perturb the level of cytosolic Ca2+. Similar maneuvers performed after agonist- or thapsigargin-induced store depletion of intracellular stores brought about dramatic changes in cytosolic Ca2+ consistent with the activation of a Ca2+ entry pathway. Lower concentrations of agonist induced oscillations of Ca2+ that continued for a short time in Ca free solution. No increase in Mn2+ quench rate was associated with oscillations. A 100-microM zinc- and KCl-induced blockade of Ca2+ entry had no effect on the form of agonist-induced oscillations. Inhibition of Ca2+ influx by zinc (100 microM) converted a sustained Ca2+ response to a train of repetitive Ca2+ spikes. CONCLUSIONS: Human lens cells normally have very low Ca2+ permeability. Depletion of intracellular stores by agonists or thapsigargin initiates a Ca2+ entry pathway that is not required for the Ca2+ oscillations induced by low concentrations of agonist. This potentially provides a signal transduction mechanism with minimal risk of Ca2+ overload to the lens, whereas overactivation of the store-operated channel is a possible way of increasing calcium in the lens and could explain the distribution found in localized cataracts. PMID- 11274080 TI - The inhibitory influence of endothelin on active sodium-potassium transport in porcine lens. AB - PURPOSE: Endothelin (ET)-1 is known to inhibit active NaK transport by as much as 50% in kidney tubule and other tissues. The presence of low levels of ET-1 in aqueous humor combined with the potential for release of ET-1 from ciliary processes suggests that the lens could be exposed to ET-1 in vivo. In this study, experiments were conducted to examine the influence of ET-1 on active NaK transport in porcine lens. METHODS: The rate of Na,K-adenosine triphosphatase (Na,K-ATPase) dependent potassium transport was determined by measurements of ouabain-sensitive potassium (86Rb) uptake by intact lenses. Lens sodium content was measured by atomic absorption spectrophotometry. Cyclic adenosine monophosphate (cAMP) was measured by radioimmunoassay. Cytoplasmic calcium concentration in cultured porcine lens epithelium was measured by a fluorescence technique using fura-2. RESULTS: In the presence of ET-1 (0.1 nM or higher concentration), the rate of ouabain-sensitive potassium (86Rb) uptake was diminished. The ET receptor antagonist PD145065 (2 microM) suppressed the inhibitory effect of ET-1 (100 nM) on 86Rb uptake. Sodium content was detectably increased in lenses exposed to ET-1 for 24 hours. Forskolin (1 microM) caused an eightfold increase of cAMP in the lens epithelium, but no increase of cAMP was detected in the epithelium of lenses treated with ET-1. Genistein (150 microM), an inhibitor of tyrosine kinases, abolished the inhibitory effects of ET-1 on lens 86Rb uptake. ET-1 caused an increase of cytoplasmic calcium concentration in cultured porcine lens epithelium. The cytoplasmic calcium response to ET-1 was inhibited by PD145065 and genistein. CONCLUSIONS: The results of the present study suggest that ET-1 causes inhibition of lens active Na-K transport by a mechanism that involves activation of ET receptors. Activation of ET receptors also causes an increase of cytoplasmic calcium concentration in cultured lens epithelial cells. Both responses to ET-1 appear to have a tyrosine kinase step, because they could be prevented by genistein. The physiological purpose of an ET 1-induced reduction in the rate of active Na-K transport by the lens is unknown at this time. PMID- 11274081 TI - Intravitreal pharmacokinetics and retinal concentrations of ganciclovir and foscarnet after intravitreal administration in rabbits. AB - PURPOSE: To perform a detailed pharmacokinetic study and to evaluate the drug levels reached in the retina after the intravitreal administration of ganciclovir and foscarnet to rabbits. METHODS: Retinal and vitreal levels of both drugs were measured by high-performance liquid chromatography at 1, 6, 12, 24, 36, 48, 60, and 72 hours after a single intravitreal injection of 196 microg and 800 microg of ganciclovir and 960 microg of foscarnet to three groups of 24 pigmented rabbits. A noncompartmental pharmacokinetic analysis was used. RESULTS: Both drugs incorporated rapidly into the retina, but no equilibrium was observed between the drug levels in the vitreous humor and retina. Mean ganciclovir levels in vitreous and retina were 179.6 microg/g and 131.3 microg/g (dose of 196 microg), 755.7 microg/g and 381.6 microg/g (dose of 800 microg) at 1 hour after administration, decreasing to 0.1 microg/g, 0.6 microg/g, 0.8 microg/g, and 0.7 microg/g, respectively, by 72 hours. Mean foscarnet levels in vitreous and retina were 944 microg/g and 217.1 microg/g at 1 hour after administration, decreasing to 74 microg/g and 17.1 microg/g, respectively, by 72 hours. Whereas both doses of ganciclovir yielded retinal levels above the mean inhibitory concentration (IC50) of most human cytomegalovirus (CMV) isolates for more than 60 hours, foscarnet retinal levels were lower than the CMV IC50 before 36 hours had elapsed after administration. CONCLUSIONS: The results suggest that the intravitreal administration of ganciclovir has a better pharmacokinetic profile than foscarnet for the treatment of retinitis caused by CMV and other herpes viruses and support the administration of intravitreal ganciclovir twice a week as a treatment for CMV retinitis. PMID- 11274082 TI - Modulation of aqueous humor outflow facility by the Rho kinase-specific inhibitor Y-27632. AB - PURPOSE: The goal of this study was to investigate the role of Rho kinase in the modulation of aqueous humor outflow facility. Rho kinase, a critical downstream effector of Rho GTPase is recognized to control the formation of actin stress fibers, focal adhesions, and cellular contraction. METHODS: Expression of Rho GTPase, Rho kinase, and other downstream targets of Rho GTPase were determined in human trabecular meshwork (HTM) and Schlemm's canal (SC) primary cell cultures by Western blot analysis. The Rho kinase-specific inhibitor (Y-27632)-induced changes in actin stress fibers, focal adhesions, and protein phosphotyrosine status were evaluated by staining with rhodamine-phalloidin, anti-paxillin, and anti-phosphotyrosine antibodies, respectively. Myosin light-chain phosphorylation was determined by Western blot analysis. Y-27632-induced changes in SC cell monolayer permeability were quantitated using a colorimetric assay to evaluate horseradish peroxidase diffusion through SC cell monolayers grown in transwell chambers. Aqueous humor outflow facility was measured using enucleated porcine eyes and a constant-pressure perfusion system. RESULTS: Treatment of HTM and SC cells with Y-27632 (10 microM) led to significant but reversible changes in cell shape and decreases in actin stress fibers, focal adhesions, and protein phosphotyrosine staining. SC cell monolayer permeability increased (by 80%) in response to Y-27632 (10 microM) treatment, whereas myosin light-chain phosphorylation was decreased in both HTM and SC cells. Aqueous humor outflow facility increased (40%-80%) in enucleated porcine eyes perfused with Y-27632 (10 100 microM), and this effect was associated with widening of the extracellular spaces, particularly the optically empty area of the juxtacanalicular tissue (JCT). The integrity of inner wall of aqueous plexi, however, was observed to be intact. CONCLUSIONS: Based on the Rho kinase inhibitor-induced changes in myosin light-chain phosphorylation and actomyosin organization, it is reasonable to conclude that cellular relaxation and loss of cell-substratum adhesions in HTM and SC cells could result in either increased paracellular fluid flow across Schlemm's canal or altered flow pathway through the JCT, thereby lowering resistance to outflow. This study also suggests Rho kinase as a potential therapeutic target for the development of drugs to modulate intraocular pressure in glaucoma patients. PMID- 11274083 TI - Ocular blood flow and retinal metabolism in abyssinian cats with hereditary retinal degeneration. AB - PURPOSE: To investigate if retinal blood flow decreases with progression of the disease in Abyssinian cats with progressive retinal atrophy (PRA), to examine if the choroidal blood flow was affected by the disease, and to determine the uptake of glucose and formation of lactate in the outer retina. METHODS: Local blood flow in different parts of the eye was determined with radioactive microspheres, in 9 normal cats and in 10 cats at different stages of PRA. Three blood flow determinations were made in each animal, during control conditions, after IV administration of indomethacin and after subsequent administration of N(omega) nitro-L-arginine (L-NA). Blood samples from a choroidal vein and a femoral artery were collected to determine the retinal formation of lactate and uptake of glucose. RESULTS: In Abyssinian cats with PRA (n = 10), the retinal blood flow was significantly (P < or = 0.01) lower than in normal cats (n = 9) during control conditions, 6.4 +/- 1.7 compared with 14.1 +/- 1.9 g min(-1) x (100 g)( 1). The vascular resistance in the iris and ciliary body was significantly higher in the cats at a late stage of PRA, both compared with normal cats and to cats at an early stage of the disease, whereas the choroidal vascular resistance was not significantly affected. Indomethacin had no effect on ocular blood flows in normal cats, but in cats with PRA, iridal blood flow was more than doubled after indomethacin. The retinal formation of lactate was significantly (P < or = 0.001) lower in cats with PRA than in normal cats, 0.111 +/- 0.035 (n = 8) compared with 0.318 +/- 0.024 (n = 8) micromol x min(-1). The uptake of glucose was not significantly different in cats with PRA. CONCLUSIONS: Retinal blood flow is severely decreased in Abyssinian cats at a late stage of retinal degeneration, whereas the choroidal microcirculation is not significantly affected by the disease. At a late stage of retinal degeneration, vascular resistance in the iris is significantly increased, which at least in part could be caused by cyxlooxygenase products. PMID- 11274084 TI - Substance P and vasoactive intestinal polypeptide in the streptozotocin-induced diabetic rat retina. AB - PURPOSE: Little knowledge exists about how neurotransmitters behave in the diabetic retina. In this study, the authors measured the concentration of two neuropeptides, substance P and vasoactive intestinal polypeptide, in the streptozotocin-induced diabetic rat retina in a time-dependent manner. METHODS: The retinas of 1-, 3-, 5-, 8-, and 12-week diabetic rats were processed using a highly sensitive radioimmunoassay for both substance P and vasoactive intestinal polypeptide. Furthermore, the peptide-immunoreactivities were characterized by high-pressure liquid chromatography. RESULTS: Substance P and vasoactive intestinal polypeptide were found to be significantly reduced with a maximum decrease of 28.6% (+/-6.7) and 64.5% (+/-10.7) after 5 weeks, respectively. The peptide-immunoreactivities were found in a major peak coeluting with the synthetic peptides indicating that the quantitative values measured by radioimmunoassay represent the authentic peptides. CONCLUSIONS: The reduction of substance P and vasoactive intestinal polypeptide is in clear contrast to the amino acid transmitters GABA and glycine, which have been shown to be elevated in this early stage of diabetic retinopathy. This finding is important for three reasons: First, the decrease may result in reduced excitability of inner retinal neurons, as both peptides are known to modulate the excitability of these neurons; second, the decrease may be the consequence of a depressing and/or damaging effect by excitotoxins; and third, it may help explain why neovascularizations do not occur in this animal model, although VEGF is massively upregulated, as substance P is a very potent vascular growth factor. PMID- 11274085 TI - Fundus autofluorescence and development of geographic atrophy in age-related macular degeneration. AB - PURPOSE: To describe the development of new and enlargement of preexisting atrophy confined to areas with abnormally high levels of in vivo autofluorescence in eyes with geographic atrophy (GA) associated with age-related macular degeneration (ARMD). METHODS: The spatial distribution and intensity of fundus autofluorescence as well as the spread of GA and occurrence of new GA was recorded over a period of 3 years in three patients with ARMD using a confocal scanning laser ophthalmoscope. RESULTS: A diffuse irregular increased autofluorescence at the posterior pole was recorded at baseline in the presence of unifocal or multifocal patches of geographic atrophy. Within these areas of elevated autofluorescence, new atrophic areas developed, and existing patches of atrophy enlarged during the review period, whereas this was not observed in areas with normal background autofluorescence. The total area of abnormal autofluorescence also showed enlargement over time. CONCLUSIONS: These preliminary findings suggest that areas of increased autofluorescence precede the development and enlargement of outer retinal atrophy in eyes with ARMD. Because the dominant fluorophores of fundus autofluorescence are part of lipofuscin granules of RPE cells, the observations indicate that excessive RPE lipofuscin accumulation may be of significance in the pathogenesis of GA associated with ARMD. With GA being a major cause for severe visual loss in ARMD, in vivo fundus autofluorescence recording over time may allow identification of prognostic determinants and may give important clues to the understanding of mechanisms of disease. PMID- 11274086 TI - Light-driven retinal ganglion cell responses in blind rd mice after neural retinal transplantation. AB - PURPOSE: Light-elicited retinal ganglion cell (RGC) responses after fetal neural retinal transplantation have not been demonstrated in animal or human subjects blind from outer retinal degeneration, despite apparent morphologic success. This study was designed to test the hypothesis that the functional success of retinal transplantation may be enhanced by using a young host retina (13 days old). METHODS: At postnatal day (P)13 C3H/HeJ (rd/rd) retinal degenerate mice received a subretinal transplant, in one eye only, of neural retinal tissue isolated from newborn normal C57/BL6J mice. Between 33 and 35 days after transplantation, local electroretinograms (ERGs) and ganglion cell responses were recorded directly from the retinal surface using a differential bipolar surface electrode. Measurements were performed both with and without light stimulation. Similar recordings were also performed in age-matched eyes subjected to sham transplantation, in control eyes that were not subjected to surgery, and in animals eyes that underwent transplantation at 8 weeks of age. After the recordings, the eyes were processed for light and transmission electron microscopy. RESULTS: Three of 10 mice showed bursts of ganglion cell action potentials (ON response only) as well as recordable intraocular ERGs over the transplant in response to 1-second and 200 msec light stimuli. Light-driven ganglion cell responses could not be recorded in areas outside the transplant in all transplant-recipient eyes, age-matched control eyes, and sham-transplantation eyes. Light responses also could not be recorded in animal eyes that received transplants at an older age (8 weeks). Electron microscopic examination confirmed the presence of photoreceptor outer segments in the areas affected by transplantation. CONCLUSIONS: This study demonstrates the presence of light-driven ganglion cell responses after subretinal transplantation in a retinal degenerate model. This finding may reflect functional integration of the transplant with the host, but a rescue effect on remaining host photoreceptors cannot be ruled out. The findings suggest, however, that modification of host parameters, such as host age, may be important approaches for improving the functional success of retinal transplantation. PMID- 11274087 TI - Preretinal neovascularization associated with acetazolamide-induced systemic acidosis in the neonatal rat. AB - PURPOSE: NH4Cl gavage in the neonatal rat produces a metabolic acidosis-induced retinopathy which serves as a model for retinopathy of prematurity (ROP). Acetazolamide induces a metabolic acidosis via an alternative biochemical mechanism (bicarbonate loss versus hydrogen ion load). In the present study, the following hypothesis was tested: acetazolamide-induced acidosis is associated with preretinal neovascularization in the neonatal rat. METHODS: All studies used newborn Sprague-Dawley rats raised in expanded litters of 25. Arterial blood pH was measured to determine the level of acidosis induced by intraperitoneal (IP) acetazolamide (50 or 200 mg/kg) or saline. In a separate retinopathy study, newborn rats (n = 75) were randomized to either IP acetazolamide, 50 mg/kg (low dose), or IP saline twice daily from days 2 to 7. After 5 days of recovery, retinal vasculature was assessed using ADPase staining and light microscopy. The presence and severity (clock hours) of neovascularization were assessed by three masked observers. In an additional retinopathy study, newborn rats (n = 100) were randomized to either IP acetazolamide, 200 mg/kg (high-dose), or IP saline twice daily from days 2 to 7. After 5 days of recovery, the retinas were similarly analyzed. RESULTS: Neovascularization occurred in 59% of rats receiving high-dose acetazolamide (200 mg/kg). High-dose acetazolamide produced a severe acidosis (pH 7.13 +/- 0.06) during drug delivery. Low-dose acetazolamide (50 mg/kg) produced a pH (7.22 +/- 0.07) that was intermediate between high-dose (200 mg/kg) acetazolamide (P < 0.001) and saline controls (7.42 +/- 0.06, P < 0.001); however, neither low-dose acetazolamide nor saline induced preretinal neovascularization. CONCLUSIONS: Acidosis induced by high-dose acetazolamide, independent of hyperoxemia or hypoxemia, is associated with preretinal neovascularization in the neonatal rat. Induction of neovascularization appears to depend on a critical threshold of acidosis severity. This study further supports a proposed independent role for acidosis in the pathogenesis of ROP. PMID- 11274088 TI - Electrophysiology of rabbit Muller (glial) cells in experimental retinal detachment and PVR. AB - PURPOSE: To determine the electrophysiological properties of Muller (glial) cells from experimentally detached rabbit retinas. METHODS: A stable local retinal detachment was induced by subretinal injection of a sodium hyaluronate solution. Muller cells were acutely dissociated and studied by the whole-cell voltage-clamp technique. RESULTS: The cell membranes of Muller cells from normal retinas were dominated by a large inwardly rectifying potassium ion (K+) conductance that caused a low-input resistance (<100 M(Omega)) and a high resting membrane potential (-82 +/- 6 mV). During the first week after detachment, the Muller cells became reactive as shown by glial fibrillary acidic protein (GFAP) immunoreactivity, and their inward currents were markedly reduced, accompanied by an increased input resistance (>200 M(Omega)). After 3 weeks of detachment, the input resistance increased further (>300 M(Omega)), and some cells displayed significantly depolarized membrane potentials (mean -69 +/- 18 mV). When PVR developed (in 20% of the cases) the inward K+ currents were virtually completely eliminated. The input resistance increased dramatically (>1000 MOmega), and almost all cells displayed strongly depolarized membrane potentials (-44 +/- 16 mV). CONCLUSIONS: Reactive Muller cells are characterized by a severe reduction of their K+ inward conductance, accompanied by depolarized membrane potentials. These changes must impair physiological glial functions, such as neurotransmitter recycling and K+ ion clearance. Furthermore, the open probability of certain types of voltage-dependent ion channels (e.g., Ca2+-dependent K+ maxi channels) increases that may be a precondition for Muller cell proliferation, particularly in PVR when a dramatic downregulation of both inward current density and resting membrane potential occurs. PMID- 11274089 TI - Induction of adrenomedullin by hypoxia in cultured retinal pigment epithelial cells. AB - PURPOSE: To explore the effects of hypoxia on the production and secretion of adrenomedullin (ADM) and endothelin (ET)-1 in human retinal pigment epithelial (RPE) cells. METHODS: RPE cells were cultured under normoxic or hypoxic (1% O2) conditions. Expression of ADM and ET-1 was examined by Northern blot analysis and radioimmunoassay. Effects of ADM and ET-1 on the number of RPE cells were examined by modified 3-(4,5-dimetylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. RESULTS: ADM mRNA expression levels and immunoreactive ADM levels in the medium were increased by hypoxia in all three human RPE cell lines (ARPE-19, D407, and F-0202). Immunoreactive ET was detected in the cultured media of D407 cells and ARPE-19 cells and identified as ET-1 by reversed-phase high performance liquid chromatography. Hypoxia treatment for 48 hours increased immunoreactive ET levels approximately 1.3-fold in the cultured media of D407, but not ARPE-19 cells. Hypoxia decreased the number of ARPE-19 cells and F-0202 cells, and the treatment with ADM ameliorated the hypoxia-induced decrease in the cell number. In contrast, exogenously added ET-1 had no significant effects on the number of ARPE-19 cells under normoxia and hypoxia. CONCLUSIONS: Hypoxia increased the expression of ADM in all three human RPE cell lines, whereas the induction of ET 1 by hypoxia was found only in D407 cells. ADM induced by hypoxia may have protective roles against hypoxic cell damage in RPE cells. PMID- 11274091 TI - Selective excitotoxic degeneration of adult pig retinal ganglion cells in vitro. AB - PURPOSE: Excitotoxicity is proposed to play a prominent role in retinal ganglion cell (RGC) death ensuing from diseases such as glaucoma and ischemia, but cell culture studies have used tissue from newborn rodents, yielding conflicting data that implicate either N-methyl D-aspartate (NMDA) or non-NMDA glutamate (Glu) receptor-mediated pathways. Excitotoxic RGC death was examined in vitro in this study, using adult pigs, a large-animal model for human retina. METHODS: Adult pig retina (and for comparative purposes young and adult rat retina) were dissociated and maintained in monolayer culture. Medium was supplemented with Glu or pharmacologic agonists or antagonists, and surviving RGCs and other retinal neurons were quantified using specific immunolabeling methods. Electrophysiological responses to externally applied Glu of RGCs in culture were recorded using whole-cell patch-clamp techniques. RESULTS: Application of Glu led to selective, dose-dependent losses in large RGCs (maximal 37% decrease at 1 mM; median effective dose [ED50], approximately 80 microM) and neurite damage in surviving RGCs. Application of Glu agonists and Glu receptor subclass antagonists showed that large RGC death was mediated through both NMDA and non-NMDA receptor pathways. Small RGCs, amacrine cells, and all other retinal neurons were resistant to Glu-induced death. By comparison, rat retinal cultures displayed heightened RGC vulnerability to Glu, mediated exclusively by non-NMDA receptor mediated pathways. Amacrine cells were unaffected by NMDA but were very sensitive to kainate application (>90% loss). Other retinal neurons were unaffected by any treatment. CONCLUSIONS: The molecular pathways underlying excitotoxic RGC death in vitro (non-NMDA or NMDA-preferring Glu receptors) vary among species and developmental stages. The selective elimination of adult pig large RGCs by NMDA receptor-mediated pathways more closely resembles human and animal glaucoma in vivo than other published culture models, providing a simplified experimental system for investigating the pharmacologic and toxicologic bases of glaucoma-like neuronal death. PMID- 11274090 TI - Lens epithelium-derived growth factor promotes photoreceptor survival in light damaged and RCS rats. AB - PURPOSE: To investigate possible protective effects of lens epithelium-derived growth factor (LEDGF) against photoreceptor death in light-damaged, Royal College of Surgeons (RCS) and P23H rhodopsin transgenic rats. METHODS: Twelve-week-old Sprague-Dawley (SD), 6-week-old RCS, and 10-day-old P23H (line 1, heterozygote) rats received an intravitreal injection of LEDGF fused with glutathione-S transferase (GST-LEDGF). Fellow eyes received vehicle and served as control specimens. Two days after the injections, the SD rats were exposed to light of 2000 lux for 48 hours. Corneal Ganzfeld ERGs were recorded 10 days after light damage, at 10 weeks of age in RCS rats, and at 4 weeks of age in P23H rats. The eyes were then processed for histologic analysis. Heat shock protein (hsp) content in the sensory retina was analyzed quantitatively by protein immunoblot. RESULTS: In light-damaged rats, the ERG indicated retinal protection in GST-LEDGF injected eyes, with b-wave and STR thresholds being 1.14 +/- 0.50 (mean +/- SD) and 0.60 +/- 0.26 log candela (cd)/m2 lower, respectively, than in vehicle injected eyes (P < 0.01). The GST-LEDGF-treated eyes had maximum b-wave amplitudes that were significantly larger (P < 0.0005), had more than twice as many remaining photoreceptors, and had better organized outer segments than the control eyes. In RCS rats, the treated eyes had 2.76 +/- 0.73 and 0.83 +/- 0.09 log cd/m(2) lower thresholds for the b-wave and STR, respectively (P < 0.005), and had significantly larger maximum b-wave amplitude (P < 0.0005). GST-LEDGF treated eyes of RCS rats also had more photoreceptors remaining (P < 0.005) and a thinner debris layer than control eyes. In P23H rats, GST-LEDGF treatment did not protect either retinal function or structure. The retinas from GST-LEDGF-treated eyes of SD and RCS rats had higher levels of hsp25 and alphaB-crystallin than vehicle-injected eyes. CONCLUSIONS: GST-LEDGF protects photoreceptor structure and function in both light-damaged and RCS rats. The increased expression of hsp25 and alphaB-crystallin may play a role in this protection. The absence of rescue in P23H raises the possibility that some forms of inherited retinal degeneration may not be amenable to treatment by intraocular injection of LEDGF. PMID- 11274092 TI - Effect of myopia on frequency-doubling perimetry. AB - PURPOSE: To examine the effect of myopia, occasionally associated with glaucomatous eyes, on the results obtained by frequency-doubling perimetry (FDP). METHODS: Sixty emmetropic or myopic normal volunteers (mean age, 26.2 +/- 0.35 years, mean +/- SEM; range, 19-34) with good visual acuity and without glaucoma were divided into three groups. The groups were emmetropia to low-myopia (mean refractive error, -1.16 +/- 0.23 D), intermediate-myopia (-4.95 +/- 0.17 D), and high-myopia (-8.12 +/- 0.36 D; n = 20 each). All subjects were tested on the FDP full-threshold C-20 program and the Humphrey Field Analyzer (HFA; Humphrey, Dublin, CA) full-threshold program on one randomly selected eye. FDP and the HFA test were conducted with the subjects wearing their full distance correction and with their distance correction with appropriate additional correction for near, respectively. The calculated mean sensitivity (MS), mean deviation (MD), pattern standard deviation (PSD), and test durations for FDP and the HFA test for the three groups were compared using one-way analysis of variance. The relationship between the refractive error and MS, MD, or PSD was also analyzed by simple regression analysis. RESULTS: The MS and MD for the fields determined by the HFA decreased significantly as the refractive errors increased, but there were no significant differences in the MS, MD, and PSD for FDP between the three groups. There were no significant differences in the test durations between the three groups for both FDP and HFA testing. The refractive error was correlated with both MS and MD only for the fields determined by the HFA. CONCLUSIONS: The results showed that lens-corrected myopia does not alter the visual fields obtained by FDP, and FDP can therefore be used regardless of the presence of myopia. PMID- 11274093 TI - Graded contribution of retinal maturation to the development of oxygen-induced retinopathy in rats. AB - PURPOSE: Newborn rats exposed to hyperoxia during the first days of life have been shown to exhibit not only vasculopathy but also permanent changes in the structure and function of the retina. Given that the rat retina is immature at birth and that the maturation process continues until the opening of the eyes at 14 days of life, this study was conducted to investigate the susceptibility of the retina to oxygen toxicity as a function of the degree of retinal maturity reached at the time of oxygen exposure. METHODS: Newborn rats were exposed to hyperoxia during selected postnatal day intervals. Scotopic electroretinograms were recorded at 30 and 60 days of age, and retinal histology was obtained at the end of the study. RESULTS: There was a strong correlation between the duration of the hyperoxic event and the structural and functional consequences in the retina. However, the repercussions were significantly more profound when the exposure to oxygen occurred within the second week of life (6-14 days), compared with earlier (0-6 days) or later periods (14-28 days). CONCLUSIONS: The results strongly suggest that the structural and functional retinal changes secondary to postnatal hyperoxia are not only the direct consequence of exposure to high levels of oxygen (i.e., free radicals), but also are determined by the level of retinal maturity reached at the time of oxygen exposure. The results also indicate that the structural anomalies precede the functional impairments. PMID- 11274094 TI - Variation in vernier evoked cortical potential with age. AB - PURPOSE: To investigate the effects of age on transient vernier visual evoked potential (VEP) and vernier acuity estimated by extrapolation. METHODS: Transient vernier VEPs were examined in normal subjects aged 20 to 75 years. Vernier offsets were presented for the first 350 msec of the stimulus period, and the segments were then realigned in the following 400 msec. The six vernier offsets used were 20, 40, 60, 80, 100, and 140 seconds of arc. Averaging for each offset setting produced vernier VEP waveforms, for which amplitude and latency of visual evoked response and background electroencephalographic (EEG) noise level were determined. Extrapolation of the function relating signal-to-noise ratio and log vernier offset to a ratio of 1.0 resulted in an estimate of vernier acuity. RESULTS: Amplitude of vernier VEP waveforms was significantly reduced in subjects more than 60 years of age, and the latency to the first negative peak was progressively prolonged with increasing age. There was no statistically significant change in electroencephalographic (EEG) noise with advancing age. VEP vernier acuity was significantly degraded in the 61- to 75-year age group. These results are parallel to recent psychophysical findings that alignment performance is worse in older persons than in younger ones. CONCLUSIONS: The present findings provide the first electrophysiological evidence of age-related cortical degeneration associated with vernier processing. Reduced neural activity probably contributes to the loss of vernier acuity with advancing age. Also provided are the first normative data for subjects of different ages for vernier VEP and VEP vernier acuity. Moreover, the present study has demonstrated that vernier VEP is sensitive to neural changes and therefore may be applied in clinical situations to evaluate the integrity of the visual system. PMID- 11274096 TI - Molecular sieve mechanism of selective release of cytoplasmic proteins by osmotically shocked Escherichia coli. AB - Escherichia coli cells, the outer membrane of which is permeabilized with EDTA, release a specific subset of cytoplasmic proteins upon a sudden drop in osmolarity in the surrounding medium. This subset includes EF-Tu, thioredoxin, and DnaK among other proteins, and comprises approximately 10% of the total bacterial protein content. As we demonstrate here, the same proteins are released from electroporated E. coli cells pretreated with EDTA. Although known for several decades, the phenomenon of selective release of proteins has received no satisfactory explanation. Here we show that the subset of released proteins is almost identical to the subset of proteins that are able to pass through a 100 kDa-cutoff cellulose membrane upon molecular filtration of an E. coli homogenate. This finding indicates that in osmotically shocked or electroporated bacteria, proteins are strained through a molecular sieve formed by the transiently damaged bacterial envelope. As a result, proteins of small native sizes are selectively released, whereas large proteins and large protein complexes are retained by bacterial cells. PMID- 11274097 TI - Melamine deaminase and atrazine chlorohydrolase: 98 percent identical but functionally different. AB - The gene encoding melamine deaminase (TriA) from Pseudomonas sp. strain NRRL B 12227 was identified, cloned into Escherichia coli, sequenced, and expressed for in vitro study of enzyme activity. Melamine deaminase displaced two of the three amino groups from melamine, producing ammeline and ammelide as sequential products. The first deamination reaction occurred more than 10 times faster than the second. Ammelide did not inhibit the first or second deamination reaction, suggesting that the lower rate of ammeline hydrolysis was due to differential substrate turnover rather than product inhibition. Remarkably, melamine deaminase is 98% identical to the enzyme atrazine chlorohydrolase (AtzA) from Pseudomonas sp. strain ADP. Each enzyme consists of 475 amino acids and differs by only 9 amino acids. AtzA was shown to exclusively catalyze dehalogenation of halo substituted triazine ring compounds and had no activity with melamine and ammeline. Similarly, melamine deaminase had no detectable activity with the halo triazine substrates. Melamine deaminase was active in deamination of a substrate that was structurally identical to atrazine, except for the substitution of an amino group for the chlorine atom. Moreover, melamine deaminase and AtzA are found in bacteria that grow on melamine and atrazine compounds, respectively. These data strongly suggest that the 9 amino acid differences between melamine deaminase and AtzA represent a short evolutionary pathway connecting enzymes catalyzing physiologically relevant deamination and dehalogenation reactions, respectively. PMID- 11274098 TI - GcpE is involved in the 2-C-methyl-D-erythritol 4-phosphate pathway of isoprenoid biosynthesis in Escherichia coli. AB - In a variety of organisms, including plants and several eubacteria, isoprenoids are synthesized by the mevalonate-independent 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway. Although different enzymes of this pathway have been described, the terminal biosynthetic steps of the MEP pathway have not been fully elucidated. In this work, we demonstrate that the gcpE gene of Escherichia coli is involved in this pathway. E. coli cells were genetically engineered to utilize exogenously provided mevalonate for isoprenoid biosynthesis by the mevalonate pathway. These cells were then deleted for the essential gcpE gene and were viable only if the medium was supplemented with mevalonate or the cells were complemented with an episomal copy of gcpE. PMID- 11274099 TI - Recruitment of the mecA gene homologue of Staphylococcus sciuri into a resistance determinant and expression of the resistant phenotype in Staphylococcus aureus. AB - Strains of methicillin-resistant Staphylococcus aureus (MRSA) have become the most important causative agents of hospital-acquired diseases worldwide. The genetic determinant of resistance, mecA, is not a gene native to S. aureus but was acquired from an extraspecies source by an unknown mechanism. We recently identified a close homologue of this gene in isolates of Staphylococcus sciuri, a taxonomically primitive staphylococcal species recovered most frequently from rodents and primitive mammals. In spite of the close sequence similarity between the mecA homologue of S. sciuri and the antibiotic resistance determinant mecA of S. aureus, S. sciuri strains were found to be uniformly susceptible to beta lactam antibiotics. In an attempt to activate the apparently "silent" mecA gene of S. sciuri, a methicillin-resistant derivative, K1M200 (for which the MIC of methicillin is 200 microg/ml), was obtained through stepwise exposure of the parental strain S. sciuri K1 (methicillin MIC of 4 microg/ml) to increasing concentrations of methicillin. DNA sequencing of the mecA homologue from K1M200 revealed the introduction of a point mutation into the -10 consensus of the promoter: the replacement of a thymine residue at nucleotide 1577 in the susceptible strain K1 by adenine in the resistant strain K1M200, which was accompanied by a drastic increase in transcription rate and the appearance of a new protein that reacted with monoclonal antibody prepared against the penicillin binding protein 2A (PBP2A), i.e., the gene product of S. aureus mecA. Transduction of mecA from K1M200 (cloned into a plasmid vector) into a methicillin-susceptible S. aureus mutant resulted in a significant increase of methicillin resistance (from a methicillin MIC of 4 micro/ml to 12 and up to 50 microg/ml), the appearance of a low-affinity PBP detectable by the fluorographic assay, and the production of a protein that reacted in a Western blot with monoclonal antibody to PBP2A. Antibiotic resistance and the protein products disappeared upon removal of the plasmid-borne mecA homologue. The observations support the proposition that the mecA homologue ubiquitous in the antibiotic susceptible animal species S. sciuri may be an evolutionary precursor of the methicillin resistance gene mecA of the pathogenic strains of MRSA. PMID- 11274100 TI - Cloning and characterization of the gene cluster for palatinose metabolism from the phytopathogenic bacterium Erwinia rhapontici. AB - Erwinia rhapontici is able to convert sucrose into isomaltulose (palatinose, 6-O alpha-D-glucopyranosyl-D-fructose) and trehalulose (1-O-alpha-D-glucopyranosyl-D fructose) by the activity of a sucrose isomerase. These sucrose isomers cannot be metabolized by plant cells and most other organisms and therefore are possibly advantageous for the pathogen. This view is supported by the observation that in vitro yeast invertase activity can be inhibited by palatinose, thus preventing sucrose consumption. Due to the lack of genetic information, the role of sucrose isomers in pathogenicity has not been evaluated. Here we describe for the first time the cloning and characterization of the palatinose (pal) genes from Erwinia rhapontici. To this end, a 15-kb chromosomal DNA fragment containing nine complete open reading frames (ORFs) was cloned. The pal gene products of Erwinia rhapontici were shown to be homologous to proteins involved in uptake and metabolism of various sugars from other microorganisms. The palE, palF, palG, palH, palK, palQ, and palZ genes were oriented divergently with respect to the palR and palI genes, and sequence analysis suggested that the first set of genes constitutes an operon. Northern blot analysis of RNA extracted from bacteria grown under various conditions implies that the expression of the palI gene and the palEFGHKQZ genes is oppositely regulated at the transcriptional level. Genes involved in palatinose uptake and metabolism are down regulated by sucrose and activated by palatinose. Palatinose activation is inhibited by sucrose. Functional expression of palI and palQ in Escherichia coli revealed sucrose isomerase and palatinase activity, respectively. PMID- 11274101 TI - Hydrogen peroxide-forming NADH oxidase belonging to the peroxiredoxin oxidoreductase family: existence and physiological role in bacteria. AB - Amphibacillus xylanus and Sporolactobacillus inulinus NADH oxidases belonging to the peroxiredoxin oxidoreductase family show extremely high peroxide reductase activity for hydrogen peroxide and alkyl hydroperoxides in the presence of the small disulfide redox protein, AhpC (peroxiredoxin). In order to investigate the distribution of this enzyme system in bacteria, 15 bacterial strains were selected from typical aerobic, facultatively anaerobic, and anaerobic bacteria. AhpC-linked alkyl hydroperoxide reductase activities were detected in most of the tested strains, and especially high activities were shown in six bacterial species that grow well under aerobic conditions, including aerobic bacteria (Alcaligenes faecalis and Bacillus licheniformis) and facultatively anaerobic bacteria (Amphibacillus xylanus, Sporolactobacillus inulinus, Escherichia coli, and Salmonella enterica serovar Typhimurium). In the absence of AhpC, the purified enzymes from A. xylanus and S. inulinus catalyze the NADH-linked reduction of oxygen to hydrogen peroxide. Similar activities were observed in the cell extracts from each of these six strains. The cell extract of B. licheniformis revealed the highest AhpC-linked alkyl hydroperoxide reductase activity in the four strains, with V(max) values for hydrogen peroxide and alkyl hydroperoxides being similar to those for the enzymes from A. xylanus and S. inulinus. Southern blot analysis of the three strains probed with the A. xylanus peroxiredoxin reductase gene revealed single strong bands, which are presumably derived from the individual peroxiredoxin reductase genes. Single bands were also revealed in other strains which show high AhpC-linked reductase activities, suggesting that the NADH oxidases belonging to the peroxiredoxin oxidoreductase family are widely distributed and possibly play an important role both in the peroxide-scavenging systems and in an effective regeneration system for NAD in aerobically growing bacteria. PMID- 11274102 TI - Independence of circadian timing from cell division in cyanobacteria. AB - In the cyanobacterium Synechococcus elongatus, cell division is regulated by a circadian clock. Deletion of the circadian clock gene, kaiC, abolishes rhythms of gene expression and cell division timing. Overexpression of the ftsZ gene halted cell division but not growth, causing cells to grow as filaments without dividing. The nondividing filamentous cells still exhibited robust circadian rhythms of gene expression. This result indicates that the circadian timing system is independent of rhythmic cell division and, together with other results, suggests that the cyanobacterial circadian system is stable and well sustained under a wide range of intracellular conditions. PMID- 11274103 TI - Adhesion of type 1-fimbriated Escherichia coli to abiotic surfaces leads to altered composition of outer membrane proteins. AB - Phenotypic differences between planktonic bacteria and those attached to abiotic surfaces exist, but the mechanisms involved in the adhesion response of bacteria are not well understood. By the use of two-dimensional (2D) polyacrylamide gel electrophoresis, we have demonstrated that attachment of Escherichia coli to abiotic surfaces leads to alteration in the composition of outer membrane proteins. A major decrease in the abundance of resolved proteins was observed during adhesion of type 1-fimbriated E. coli strains, which was at least partly caused by proteolysis. Moreover, a study of fimbriated and nonfimbriated mutants revealed that these changes were due mainly to type 1 fimbria-mediated surface contact and that only a few changes occurred in the outer membranes of nonfimbriated mutant strains. Protein synthesis and proteolytic degradation were involved to different extents in adhesion of fimbriated and nonfimbriated cells. While protein synthesis appeared to affect adhesion of only the nonfimbriated strain, proteolytic activity mostly seemed to contribute to adhesion of the fimbriated strain. Using matrix-assisted laser desorption ionization-time of flight mass spectrometry, six of the proteins resolved by 2D analysis were identified as BtuB, EF-Tu, OmpA, OmpX, Slp, and TolC. While the first two proteins were unaffected by adhesion, the levels of the last four were moderately to strongly reduced. Based on the present results, it may be suggested that physical interactions between type 1 fimbriae and the surface are part of a surface-sensing mechanism in which protein turnover may contribute to the observed change in composition of outer membrane proteins. This change alters the surface characteristics of the cell envelope and may thus influence adhesion. PMID- 11274104 TI - Interruption of the cydB locus in Brucella abortus attenuates intracellular survival and virulence in the mouse model of infection. AB - Brucellosis is characterized by abortion in ruminants and a protracted undulant fever in humans, which often results in severe pathological manifestations. Scant information exists about the molecular mechanisms employed by Brucella abortus to combat host defenses or to persist and replicate within host cells. Transposon (Tn5) mutagenesis of B. abortus and the subsequent screening of mutants for sensitivity to killing in murine macrophages and in the mouse model led to the identification of mutants which were severely attenuated for intracellular survival. One group of mutants was interrupted in cydB, a gene that is part of the cydAB operon encoding cytochrome bd oxidase, which catalyzes an alternate terminal electron transport step in bacterial respiration. The elevated affinity for molecular oxygen of this enzyme in Escherichia coli has suggested that it is involved in the protection of sensitive enzymatic activities such as those of hydrogenases and nitrogenases from damage. B. abortus cydB::Tn5 strains exhibited heightened sensitivity to the respiratory inhibitors zinc and azide, highly reactive oxygen species such as hydrogen peroxide, low pH, and attenuated virulence in the mouse model of infection. Virulence was restored by an intact copy of cydAB or by B. abortus genes encoding the oxidative radical-scavenging enzyme Cu/Zn superoxide dismutase or catalase. These results suggest a bifunctional role for the products of the cydAB operon, both in preventing the buildup of oxidative free radicals and in detoxifying the intracellular compartment, thus indicating the importance of these products in preventing intracellular destruction. Intracellular conditions that favor expression of the cydAB operon are under investigation and may be linked to the acid sensitivity also observed in this strain. PMID- 11274105 TI - The alternative electron acceptor tetrathionate supports B12-dependent anaerobic growth of Salmonella enterica serovar typhimurium on ethanolamine or 1,2 propanediol. AB - Synthesis of cobalamin de novo by Salmonella enterica serovar Typhimurium strain LT2 and the absence of this ability in Escherichia coli present several problems. This large synthetic pathway is shared by virtually all salmonellae and must be maintained by selection, yet no conditions are known under which growth depends on endogenous B12. The cofactor is required for degradation of 1,2-propanediol and ethanolamine. However, cofactor synthesis occurs only anaerobically, and neither of these carbon sources supports anaerobic growth with any of the alternative electron acceptors tested thus far. This paradox is resolved by the electron acceptor tetrathionate, which allows Salmonella to grow anaerobically on ethanolamine or 1,2-propanediol by using endogenously synthesized B12. Tetrathionate provides the only known conditions under which simple cob mutants (unable to make B12) show a growth defect. Genes involved in this metabolism include the ttr operon, which encodes tetrathionate reductase. This operon is globally regulated by OxrA (Fnr) and induced anaerobically by a two-component system in response to tetrathionate. Salmonella reduces tetrathionate to thiosulfate, which it can further reduce to H2S, by using enzymes encoded by the genes phs and asr. The genes for 1,2-propanediol degradation (pdu) and B12 synthesis (cob), along with the genes for sulfur reduction (ttr, phs, and asr), constitute more than 1% of the Salmonella genome and are all absent from E. coli. In diverging from E. coli, Salmonella acquired some of these genes unilaterally and maintained others that are ancestral but have been lost from the E. coli lineage. PMID- 11274106 TI - Involvement of H-NS in transpositional recombination mediated by IS1. AB - IS1, the smallest active transposable element in bacteria, encodes a transposase that promotes inter- and intramolecular transposition. Host-encoded factors, e.g., histone-like proteins HU and integration host factor (IHF), are involved in the transposition reactions of some bacterial transposable elements. Host factors involved in the IS1 transposition reaction, however, are not known. We show that a plasmid with an IS1 derivative that efficiently produces transposase did not generate miniplasmids, the products of intramolecular transposition, in mutants deficient in a nucleoid-associated DNA-binding protein, H-NS, but did generate them in mutants deficient in histone-like proteins HU, IHF, Fis, and StpA. Nor did IS1 transpose intermolecularly to the target plasmid in the H-NS-deficient mutant. The hns mutation did not affect transcription from the indigenous promoter of IS1 for the expression of the transposase gene. These findings show that transpositional recombination mediated by IS1 requires H-NS but does not require the HU, IHF, Fis, or StpA protein in vivo. Gel retardation assays of restriction fragments of IS1-carrying plasmid DNA showed that no sites were bound preferentially by H-NS within the IS1 sequence. The central domain of H-NS, which is involved in dimerization and/or oligomerization of the H-NS protein, was important for the intramolecular transposition of IS1, but the N- and C-terminal domains, which are involved in the repression of certain genes and DNA binding, respectively, were not. The SOS response induced by the IS1 transposase was absent in the H-NS-deficient mutant strain but was present in the wild-type strain. We discuss the possibility that H-NS promotes the formation of an active IS1 DNA-transposase complex in which the IS1 ends are cleaved to initiate transpositional recombination through interaction with IS1 transposase. PMID- 11274107 TI - Ability for anaerobic growth is not sufficient for development of the petite phenotype in Saccharomyces kluyveri. AB - Saccharomyces cerevisiae is a petite-phenotype-positive ("petite-positive") yeast, which can successfully grow in the absence of oxygen. On the other hand, Kluyveromyces lactis as well as many other yeasts are petite negative and cannot grow anaerobically. In this paper, we show that Saccharomyces kluyveri can grow under anaerobic conditions, but while it can generate respiration-deficient mutants, it cannot generate true petite mutants. From a phylogenetic point of view, S. kluyveri is apparently more closely related to S. cerevisiae than to K. lactis. These observations suggest that the progenitor of the modern Saccharomyces and Kluyveromyces yeasts, as well as other related genera, was a petite-negative and aerobic yeast. Upon separation of the K. lactis and S. kluyveri-S. cerevisiae lineages, the latter developed the ability to grow anaerobically. However, while the S. kluyveri lineage has remained petite negative, the lineage leading to the modern Saccharomyces sensu stricto and sensu lato yeasts has developed the petite-positive characteristic. PMID- 11274108 TI - Topology of OxlT, the oxalate transporter of Oxalobacter formigenes, determined by site-directed fluorescence labeling. AB - The topology of OxlT, the oxalate:formate exchange protein of Oxalobacter formigenes, was established by site-directed fluorescence labeling, a simple strategy that generates topological information in the context of the intact protein. Accessibility of cysteine to the fluorescent thiol-directed probe Oregon green maleimide (OGM) was examined for a panel of 34 single-cysteine variants, each generated in a His(9)-tagged cysteine-less host. The reaction with OGM was readily scored by examining the fluorescence profile after sodium dodecyl sulfate polyacrylamide gel electrophoresis of material purified by Ni2+ linked affinity chromatography. A position was assigned an external location if its single cysteine derivative reacted with OGM added to intact cells; a position was designated internal if OGM labeling required cell lysis. We also showed that labeling of external, but not internal, positions was blocked by prior exposure of cells to the impermeable and nonfluorescent thiol-specific agent ethyltrimethylammonium methanethiosulfonate. Of the 34 positions examined in this way, 29 were assigned unambiguously to either an internal or external location; 5 positions could not be assigned, since the target cysteine failed to react with OGM. There was no evidence of false-positive assignment. Our findings document a simple and rapid method for establishing the topology of a membrane protein and show that OxlT has 12 transmembrane segments, confirming inferences from hydropathy analysis. PMID- 11274110 TI - Bacillus subtilis NhaC, an Na+/H+ antiporter, influences expression of the phoPR operon and production of alkaline phosphatases. AB - When Bacillus subtilis is subjected to phosphate starvation, genes of the Pho regulon are either induced or repressed. Among those induced are genes encoding alkaline phosphatases (APases). A set of isogenic mutants, with a beta galactosidase gene transcriptionally fused to the inactivated target gene, was used to identify genes that influence the operation of the Pho regulon. One such gene was nhaC (previously yheL). In the absence of NhaC, growth and APase production were enhanced, while the production of other non-Pho-regulon secretory proteins (proteases and alpha-amylase) did not change. The influence of NhaC on growth, APase synthesis, and its own expression was dependent on the external Na+ concentration. Other monovalent cations such as Li+ or K+ had no effect. We propose a role for NhaC in the uptake of Na+. nhaC appears to be encoded by a monocistronic operon and, contrary to previous reports, is not in the same transcriptional unit as yheK, the gene immediately upstream. The increase in APase production was dependent on an active PhoR, the sensor kinase of the two component system primarily responsible for controlling the Pho regulon. Transcriptional fusions showed that the phoPR operon and both phoA (encoding APaseA) and phoB (encoding APaseB) were hyperinduced in the absence of NhaC and repressed when this protein was overproduced. This suggests that NhaC effects APase production via phoPR. PMID- 11274109 TI - Regulation of the acetoin catabolic pathway is controlled by sigma L in Bacillus subtilis. AB - Bacillus subtilis grown in media containing amino acids or glucose secretes acetate, pyruvate, and large quantities of acetoin into the growth medium. Acetoin can be reused by the bacteria during stationary phase when other carbon sources have been depleted. The acoABCL operon encodes the E1alpha, E1beta, E2, and E3 subunits of the acetoin dehydrogenase complex in B. subtilis. Expression of this operon is induced by acetoin and repressed by glucose in the growth medium. The acoR gene is located downstream from the acoABCL operon and encodes a positive regulator which stimulates the transcription of the operon. The product of acoR has similarities to transcriptional activators of sigma 54-dependent promoters. The four genes of the operon are transcribed from a -12, -24 promoter, and transcription is abolished in acoR and sigL mutants. Deletion analysis showed that DNA sequences more than 85 bp upstream from the transcriptional start site are necessary for full induction of the operon. These upstream activating sequences are probably the targets of AcoR. Analysis of an acoR'-'lacZ strain of B. subtilis showed that the expression of acoR is not induced by acetoin and is repressed by the presence of glucose in the growth medium. Transcription of acoR is also negatively controlled by CcpA, a global regulator of carbon catabolite repression. A specific interaction of CcpA in the upstream region of acoR was demonstrated by DNase I footprinting experiments, suggesting that repression of transcription of acoR is mediated by the binding of CcpA to the promoter region of acoR. PMID- 11274111 TI - Nitric oxide signaling and transcriptional control of denitrification genes in Pseudomonas stutzeri. AB - The expression of denitrification by a facultatively anaerobic bacterium requires as exogenous signals a low oxygen tension concomitant with an N oxide. We have studied the role of nitric oxide (NO), nitrous oxide (N2O), and nitrite as signal molecules for the expression of the denitrification apparatus of Pseudomonas stutzeri. Transcriptional kinetics of structural genes were monitored by Northern blot analysis in a 60-min time frame after cells were exposed to an N oxide signal. To differentiate the inducer role of NO from that of nitrite, mRNA kinetics were monitored under anoxic conditions in a nirF strain, where NO generation from nitrite is prevented because of a defect in heme D(1) biosynthesis. NO-triggered responses were monitored from the nirSTB operon (encoding cytochrome cd(1) nitrite reductase), the norCB operon (encoding NO reductase), nosZ (encoding nitrous oxide reductase), and nosR (encoding a putative regulator). Transcription of nirSTB and norCB was activated by 5 to 50 nM NO, whereas the nosZ promoter required about 250 nM. Nitrite at 5 to 50 nM elicited no response. At a threshold concentration of 650 nM N2O, we observed in the anoxic cell the transient appearance of nosZ and nosR transcripts. Constant levels of transcripts of both genes were observed in an anoxic cell sparged with N2O. NO at 250 nM stimulated in this cell type the expression of nos genes severalfold. The transcription factor DnrD, a member of the FNR-CRP family, was found to be part of the NO-triggered signal transduction pathway. However, overexpression of dnrD in an engineered strain did not result in NirS synthesis, indicating a need for activation of DnrD. NO modified the transcriptional pattern of the dnrD operon by inducing the transcription of dnrN and dnrO, located upstream of dnrD. Insertional mutagenesis of dnrN altered the kinetic response of the nirSTB operon towards nitrite. Our data establish NO and DnrD as key elements in the regulatory network of denitrification in P. stutzeri. The NO response adds to the previously identified nitrate-nitrite response mediated by the NarXL two component system for the expression of respiratory nitrate reductase encoded by the narGHJI operon. PMID- 11274112 TI - Cytoplasmic RNA Polymerase in Escherichia coli. AB - To obtain an estimate for the concentration of free functional RNA polymerase in the bacterial cytoplasm, the content of RNA polymerase beta and beta' subunits in DNA-free minicells from the minicell-producing Escherichia coli strain chi925 was determined. In bacteria grown in Luria-Bertani medium at 2.5 doublings/h, 1.0% of the total protein was RNA polymerase. The concentration of cytoplasmic RNA polymerase beta and beta' subunits in minicells produced by this strain corresponded to about 17% (or 2.5 microM) of the value found in whole cells. Literature data suggest that a similar portion of cytoplasmic RNA polymerase subunits is in RNA polymerase assembly intermediates and imply that free functional RNA polymerase can form a small percentage of the total functional enzyme in the cell. On infection with bacteriophage T7, 20% of the minicells produced progeny phage, whereas infection in 80% of the cells was abortive. RNA polymerase subunits in lysozyme-freeze-thaw lysates of minicells were associated with minicell envelopes and were without detectable activity in an in vitro transcription assay. Together, these results suggest that most functional RNA polymerase is associated with the DNA and that little if any segregates into DNA free minicells. PMID- 11274113 TI - Efficiency of recombination reactions catalyzed by class 1 integron integrase IntI1. AB - The class 1 integron integrase, IntI1, recognizes two distinct types of recombination sites, attI sites, found in integrons, and members of the 59-be family, found in gene cassettes. The efficiencies of the integrative version of the three possible reactions, i.e., between two 59-be, between attI1 and a 59-be, or between two attI1 sites, were compared. Recombination events involving two attI1 sites were significantly less efficient than the reactions in which a 59-be participated, and the attI1 x 59-be reaction was generally preferred over the 59 be x 59-be reaction. Recombination of attI1 with secondary sites was less efficient than the 59-be x secondary site reaction. PMID- 11274114 TI - Characterization of the sat operon in Streptococcus mutans: evidence for a role of Ffh in acid tolerance. AB - An essential protein translocation pathway in Escherichia coli and Bacillus subtilis involves the signal recognition particle (SRP), of which the 54-kDa homolog (Ffh) is an essential component. In a previous study, we found that a transposon insertion in the ylxM-ffh intergenic region of the designated secretion and acid tolerance (sat) operon of Streptococcus mutans resulted in an acid-sensitive phenotype. In the present study, we further characterized this genomic region in S. mutans after construction of bona fide sat operon mutants and confirmed the role of the SRP pathway in acid resistance. Northern blot and primer extension analyses identified an acid-inducible promoter upstream of ylxM that was responsible for upregulating the coordinate expression of all five genes of the sat operon when cells were grown at acid pH. Two constitutive promoters, one immediately upstream of satD and one just 3' to the acid-inducible promoter, were also identified. Except for Ffh, the functions of the sat operon gene products are unknown. SatC, SatD, and SatE have no homology to proteins with known functions, although YlxM may function as a transcriptional regulator linked to genes encoding SRP pathway proteins. Nonpolar mutations created in each of the five genes of the sat locus resulted in viable mutants. Most striking, however, was the finding that a mutation in ffh did not result in loss of cell viability, as is the case in all other microbial species in which this pathway has been described. This mutant also lacked immunologically detectable Ffh and was severely affected in resistance to acid. Complementation of the mutation resulted in restoration of acid tolerance and reappearance of cytoplasmic Ffh. These data provide evidence that the SRP pathway plays an important role in acid tolerance in S. mutans. PMID- 11274115 TI - Allelic diversity and recombination in Campylobacter jejuni. AB - The allelic diversity and population structure of Campylobacter jejuni were studied by multilocus nucleotide sequence analysis. Sequences from seven housekeeping genes were obtained from 32 C. jejuni isolates isolated from enteritis patients in Germany, Hungary, Thailand, and the United States. Also included was strain NCTC 11168, the complete genomic sequence of which has recently been published. For all loci analyzed, multiple strains carried identical alleles. The frequency of synonymous and nonsynonymous sequence polymorphisms was low. The number of unique alleles per locus ranged from 9 to 15. These alleles occurred in 31 different combinations (sequence types), so that all but two pairs of strains could be distinguished from each other. Sequences were analyzed for evidence of recombination by the homoplasy test and split decomposition. These analyses showed that intraspecific recombination is frequent in C. jejuni and has generated extensive diversity of allelic profiles from a small number of polymorphic nucleotides. PMID- 11274116 TI - Mutually exclusive distribution of IS1548 and GBSi1, an active group II intron identified in human isolates of group B streptococci. AB - The present study shows that active, self-splicing group II intron GBSi1 is located downstream of the C5a-peptidase gene, scpB, in some group B streptococcus (GBS) isolates that lack insertion sequence IS1548. IS1548 was previously reported to be often present at the scpB locus in GBS isolated in association with endocarditis. Since none of 67 GBS isolates examined, 40 of which were of serotype III, harbored both IS1548 and GBSi1, these two elements are suggested to be markers for different genetic lineages in GBS serotype III. The DNA region downstream of scpB in GBS isolates harboring either GBSi1, IS1548, or none of these mobile elements was found to encode the laminin binding protein, Lmb, which shows sequence similarities to a family of streptococcal adhesins. IS1548 is inserted 9 bp upstream of the putative promoter for lmb, while the insertion site for GBSi1 is located 88 bp further upstream. Sequences highly similar to GBSi1 exist also in Streptococcus pneumoniae. An inverted repeat sequence, with features typical of transcription terminators, was identified immediately upstream of the insertion site for the group II intron both in the GBS and S. pneumoniae sequences. This motif is suggested to constitute a target for the GBS intron as well as for rather closely related introns in Bacillus halodurans, Pseudomonas alcaligenes, and Pseudomonas putida. When transcripts containing the GBSi1 intron were incubated at high concentrations of ammonium and magnesium, a major product with the expected length and sequence for the ligated exons was generated. Unlike, however, all members of group II investigated so far, the excised intron was in linear, rather than in a branched (lariat), form. PMID- 11274117 TI - Genetic isolation of meningococci of the electrophoretic type 37 complex. AB - Neisseria meningitidis (the meningococcus) is a naturally competent bacterial species in which intra- and interspecific horizontal gene transfer is a major source of genetic diversity. In strains of the electrophoretic type 37 (ET-37) complex and of the A4 cluster, we identified genomic DNA coding for a novel restriction-modification system and for the tail of a previously unidentified prophage. Furthermore, a novel 7.2-kb DNA segment restricted to clones of the ET 37 complex and the A4 cluster was isolated and shown to occur both as a plasmid (pJS-B) and as a chromosomal integration. Neither the genomic loci nor pJS-B was present in ET-5 complex, lineage 3, or serogroup A meningococci. The differential distribution of the DNA segments described herein, as well as of opcA, porB, nmeAI, nmeBI, and nmeDI described previously, supports the concept of genetic isolation of hypervirulent lineages responsible for most cases of serogroup C disease worldwide. PMID- 11274118 TI - Genetic organization of the region encoding regulation, biosynthesis, and transport of rhizobactin 1021, a siderophore produced by Sinorhizobium meliloti. AB - Eight genes have been identified that function in the regulation, biosynthesis, and transport of rhizobactin 1021, a hydroxamate siderophore produced under iron stress by Sinorhizobium meliloti. The genes were sequenced, and transposon insertion mutants were constructed for phenotypic analysis. Six of the genes, named rhbABCDEF, function in the biosynthesis of the siderophore and were shown to constitute an operon that is repressed under iron-replete conditions. Another gene in the cluster, named rhtA, encodes the outer membrane receptor protein for rhizobactin 1021. It was shown to be regulated by iron and to encode a product having 61% similarity to IutA, the outer membrane receptor for aerobactin. Transcription of both the rhbABCDEF operon and the rhtA gene was found to be positively regulated by the product of the eighth gene in the cluster, named rhrA, which has characteristics of an AraC-type transcriptional activator. The six genes in the rhbABCDEF operon have interesting gene junctions with short base overlaps existing between the genes. Similarities between the protein products of the biosynthesis genes and other proteins suggest that rhizobactin 1021 is synthesized by the formation of a novel siderophore precursor, 1,3 diaminopropane, which is then modified and attached to citrate in steps resembling those of the aerobactin biosynthetic pathway. The cluster of genes is located on the pSyma megaplasmid of S. meliloti 2011. Reverse transcription-PCR with RNA isolated from mature alfalfa nodules yielded no products for rhbF or rhtA at a time when the nifH gene was strongly expressed, indicating that siderophore biosynthesis and transport genes are not strongly expressed when nitrogenase is being formed in root nodules. Mutants having transposon insertions in the biosynthesis or transport genes induced effective nitrogen-fixing nodules on alfalfa plants. PMID- 11274119 TI - Yersinia pestis pFra shows biovar-specific differences and recent common ancestry with a Salmonella enterica serovar Typhi plasmid. AB - Population genetic studies suggest that Yersinia pestis, the cause of plague, is a clonal pathogen that has recently emerged from Yersinia pseudotuberculosis. Plasmid acquisition is likely to have been a key element in this evolutionary leap from an enteric to a flea-transmitted systemic pathogen. However, the origin of Y. pestis-specific plasmids remains obscure. We demonstrate specific plasmid rearrangements in different Y. pestis strains which distinguish Y. pestis bv. Orientalis strains from other biovars. We also present evidence for plasmid associated DNA exchange between Y. pestis and the exclusively human pathogen Salmonella enterica serovar Typhi. PMID- 11274120 TI - A functional myo-inositol dehydrogenase gene is required for efficient nitrogen fixation and competitiveness of Sinorhizobium fredii USDA191 to nodulate soybean (Glycine max [L.] Merr.). AB - Inositol derivative compounds provide a nutrient source for soil bacteria that possess the ability to degrade such compounds. Rhizobium strains that are capable of utilizing certain inositol derivatives are better colonizers of their host plants. We have cloned and determined the nucleotide sequence of the myo-inositol dehydrogenase gene (idhA) of Sinorhizobium fredii USDA191, the first enzyme responsible for inositol catabolism. The deduced IdhA protein has a molecular mass of 34,648 Da and shows significant sequence similarity with protein sequences of Sinorhizobium meliloti IdhA and MocA; Bacillus subtilis IolG, YrbE, and YucG; and Streptomyces griseus StrI. S. fredii USDA191 idhA mutants revealed no detectable myo-inositol dehydrogenase activity and failed to grow on myo inositol as a sole carbon source. Northern blot analysis and idhA-lacZ fusion expression studies indicate that idhA is inducible by myo-inositol. S. fredii USDA191 idhA mutant was drastically affected in its ability to reduce nitrogen and revealed deteriorating bacteroids inside the nodules. The number of bacteria recovered from such nodules was about threefold lower than the number of bacteria isolated from nodules initiated by S. fredii USDA191. In addition, the idhA mutant was also severely affected in its ability to compete with the wild-type strain in nodulating soybean. Under competitive conditions, nodules induced on soybean roots were predominantly occupied by the parent strain, even when the idhA mutant was applied at a 10-fold numerical advantage. Thus, we conclude that a functional idhA gene is required for efficient nitrogen fixation and for competitive nodulation of soybeans by S. fredii USDA191. PMID- 11274121 TI - PatS and products of nitrogen fixation control heterocyst pattern. AB - The filamentous cyanobacterium Anabaena sp. strain PCC 7120 forms a developmental pattern of single heterocysts separated by approximately 10 vegetative cells. Heterocysts differentiate from vegetative cells and are specialized for nitrogen fixation. The patS gene, which encodes a small peptide that inhibits heterocyst differentiation, is expressed in proheterocysts and plays a critical role in establishing the heterocyst pattern. Here we present further analysis of patS expression and heterocyst pattern formation. A patS-gfp reporter strain revealed clusters of patS-expressing cells during the early stage of heterocyst differentiation. PatS signaling is likely to be involved in the resolution of these clusters. Differentiating cells were inhibited by PatS during the time period 6 to 12 h after heterocyst induction, when groups of differentiating cells were being resolved to a single proheterocyst. Increased transcription of patS during development coincided with expression from a new transcription start site. In vegetative cells grown on nitrate, the 5' end of a transcript for patS was localized 314 bases upstream from the first translation initiation codon. After heterocyst induction, a new transcript with a 5' end at -39 bases replaced the vegetative cell transcript. A patS mutant grown for several days under nitrogen fixing conditions showed partial restoration of the normal heterocyst pattern, presumably because of a gradient of nitrogen compounds supplied by the heterocysts. The patS mutant formed heterocysts when grown in the presence of nitrate but showed no nitrogenase activity and no obvious heterocyst pattern. We conclude that PatS and products of nitrogen fixation are the main signals determining the heterocyst pattern. PMID- 11274122 TI - Biochemical analysis of replication factor C from the hyperthermophilic archaeon Pyrococcus furiosus. AB - Replication factor C (RFC) and proliferating cell nuclear antigen (PCNA) are accessory proteins essential for processive DNA synthesis in the domain Eucarya. The function of RFC is to load PCNA, a processivity factor of eukaryotic DNA polymerases delta and epsilon, onto primed DNA templates. RFC-like genes, arranged in tandem in the Pyrococcus furiosus genome, were cloned and expressed individually in Escherichia coli cells to determine their roles in DNA synthesis. The P. furiosus RFC (PfuRFC) consists of a small subunit (RFCS) and a large subunit (RFCL). Highly purified RFCS possesses an ATPase activity, which was stimulated up to twofold in the presence of both single-stranded DNA (ssDNA) and P. furiosus PCNA (PfuPCNA). The ATPase activity of PfuRFC itself was as strong as that of RFCS. However, in the presence of PfuPCNA and ssDNA, PfuRFC exhibited a 10-fold increase in ATPase activity under the same conditions. RFCL formed very large complexes by itself and had an extremely weak ATPase activity, which was not stimulated by PfuPCNA and DNA. The PfuRFC stimulated PfuPCNA-dependent DNA synthesis by both polymerase I and polymerase II from P. furiosus. We propose that PfuRFC is required for efficient loading of PfuPCNA and that the role of RFC in processive DNA synthesis is conserved in Archaea and Eucarya. PMID- 11274123 TI - Biofilm formation by Staphylococcus epidermidis depends on functional RsbU, an activator of the sigB operon: differential activation mechanisms due to ethanol and salt stress. AB - Staphylococcus epidermidis is a common pathogen in medical device-associated infections. Its major pathogenetic factor is the ability to form adherent biofilms. The polysaccharide intercellular adhesin (PIA), which is synthesized by the products of the icaADBC gene cluster, is essential for biofilm accumulation. In the present study, we characterized the gene locus inactivated by Tn917 insertions of two isogenic, icaADBC-independent, biofilm-negative mutants, M15 and M19, of the biofilm-producing bacterium S. epidermidis 1457. The insertion site was the same in both of the mutants and was located in the first gene, rsbU, of an operon highly homologous to the sigB operons of Staphylococcus aureus and Bacillus subtilis. Supplementation of Trypticase soy broth with NaCl (TSB(NaCl)) or ethanol (TSB(EtOH)), both of which are known activators of sigB, led to increased biofilm formation and PIA synthesis by S. epidermidis 1457. Insertion of Tn917 into rsbU, a positive regulator of alternative sigma factor sigma(B), led to a biofilm-negative phenotype and almost undetectable PIA production. Interestingly, in TSB(EtOH), the mutants were enabled to form a biofilm again with phenotypes similar to those of the wild type. In TSB(NaCl), the mutants still displayed a biofilm-negative phenotype. No difference in primary attachment between the mutants and the wild type was observed. Similar phenotypic changes were observed after transfer of the Tn917 insertion of mutant M15 to the independent and biofilm-producing strain S. epidermidis 8400. In 11 clinical S. epidermidis strains, a restriction fragment length polymorphism of the sigB operon was detected which was independent of the presence of the icaADBC locus and a biofilm-positive phenotype. Obviously, different mechanisms are operative in the regulation of PIA expression in stationary phase and under stress induced by salt or ethanol. PMID- 11274124 TI - Endophytic colonization of rice by a diazotrophic strain of Serratia marcescens. AB - Six closely related N2-fixing bacterial strains were isolated from surface sterilized roots and stems of four different rice varieties. The strains were identified as Serratia marcescens by 16S rRNA gene analysis. One strain, IRBG500, chosen for further analysis showed acetylene reduction activity (ARA) only when inoculated into media containing low levels of fixed nitrogen (yeast extract). Diazotrophy of IRBG500 was confirmed by measurement of 15N2 incorporation and by sequence analysis of the PCR-amplified fragment of nifH. To examine its interaction with rice, strain IRBG500 was marked with gusA fused to a constitutive promoter, and the marked strain was inoculated onto rice seedlings under axenic conditions. At 3 days after inoculation, the roots showed blue staining, which was most intense at the points of lateral root emergence and at the root tip. At 6 days, the blue precipitate also appeared in the leaves and stems. More detailed studies using light and transmission electron microscopy combined with immunogold labeling confirmed that IRBG500 was endophytically established within roots, stems, and leaves. Large numbers of bacteria were observed within intercellular spaces, senescing root cortical cells, aerenchyma, and xylem vessels. They were not observed within intact host cells. Inoculation of IRBG500 resulted in a significant increase in root length and root dry weight but not in total N content of rice variety IR72. The inoculated plants showed ARA, but only when external carbon (e.g., malate, succinate, or sucrose) was added to the rooting medium. PMID- 11274126 TI - The hetF gene product is essential to heterocyst differentiation and affects HetR function in the cyanobacterium Nostoc punctiforme. AB - A novel gene, hetF, was identified as essential for heterocyst development in the filamentous cyanobacterium Nostoc punctiforme strain ATCC 29133. In the absence of combined nitrogen, hetF mutants were unable to differentiate heterocysts, whereas extra copies of hetF in trans induced the formation of clusters of heterocysts. Sequences hybridizing to a hetF probe were detected only in heterocyst-forming cyanobacteria. The inactivation and multicopy effects of hetF were similar to those of hetR, which encodes a self-degrading serine protease thought to be a central regulator of heterocyst development. Increased transcription of hetR begins in developing cells 3 to 6 h after deprivation for combined nitrogen (N step-down), and the HetR protein specifically accumulates in heterocysts. In the hetF mutant, this increase in hetR transcription was delayed, and a hetR promoter::green fluorescent protein (GFP) transcriptional reporter indicated that increased transcription of hetR occurred in all cells rather than only in developing heterocysts. When a fully functional HetR-GFP fusion protein was expressed in the hetF mutant from a multicopy plasmid, HetR-GFP accumulated nonspecifically in all cells under nitrogen-replete conditions; when expressed in the wild type, HetR-GFP was observed only in heterocysts after N step-down. HetF therefore appears to cooperate with HetR in a positive regulatory pathway and may be required for the increased transcription of hetR and localization of the HetR protein in differentiating heterocysts. PMID- 11274125 TI - Suppression of hypersensitivity of Escherichia coli acrB mutant to organic solvents by integrational activation of the acrEF operon with the IS1 or IS2 element. AB - The AcrAB-TolC efflux pump plays an intrinsic role in resistance to hydrophobic solvents in Escherichia coli. E. coli OST5500 is hypersensitive to solvents due to inactivation of the acrB gene by insertion of IS30. Suppressor mutants showing high solvent resistance were isolated from OST5500. These mutants produced high levels of AcrE and AcrF proteins, which were not produced in OST5500, and in each mutant an insertion sequence (IS1 or IS2) was found integrated upstream of the acrEF operon, coding for the two proteins. The suppressor mutants lost solvent resistance on inactivation of the acrEF operon. The solvent hypersensitivity of OST5500 was suppressed by introduction of the acrEF operon with IS1 or IS2 integrated upstream but not by introduction of the operon lacking the integrated IS. It was concluded that IS integration activated acrEF, resulting in functional complementation of the acrB mutation. The acrB mutation was also complemented by a plasmid containing acrF or acrEF under the control of Plac. The wild-type tolC gene was found to be essential for complementation of the acrB mutation by acrEF. Thus, it is concluded that in these cells a combination of the proteins AcrA, AcrF, and TolC or the proteins AcrE, AcrF, and TolC is functional in solvent efflux instead of the AcrAB-TolC efflux pump. PMID- 11274127 TI - An alpha/beta-type, small, acid-soluble spore protein which has very high affinity for DNA prevents outgrowth of Bacillus subtilis spores. AB - A derivative of SspC, a minor alpha/beta-type, small, acid-soluble spore protein (SASP) from Bacillus subtilis, was generated that has a very high affinity for DNA. This protein (SspC(Delta11-D13K)) was able to confer UV resistance on spores lacking alpha/beta-type SASP, and spores with SspC(Delta11-D13K) triggered germination normally. However, SspC(Delta11-D13K) blocked outgrowth of > or = 90% of germinated spores, and SspC(Delta11-D13K) persisted in these germinated spores, whereas wild-type SspC was almost completely degraded. The outgrowth phenotype of spores with SspC(Delta11-D13K) is proposed to be due to the high stability of the SspC(Delta11-D13K)-DNA complex, which prevents rapid degradation of this alpha/beta-type SASP early in germination. The persistence of this protein on spore DNA then interferes with transcription during spore outgrowth. PMID- 11274128 TI - Twelve-transmembrane-segment (TMS) version (DeltaTMS VII-VIII) of the 14-TMS Tet(L) antibiotic resistance protein retains monovalent cation transport modes but lacks tetracycline efflux capacity. AB - A "Tet(L)-12" version of Tet(L), a tetracycline efflux protein with 14 transmembrane segments (TMS), was constructed by deletion of two central TMS. Tet(L)-12 catalyzed Na+/H+ antiport and antiport with K+ as a coupling ion as well as or better than wild-type Tet(L) but exhibited no tetracycline-Me2+/H+ antiport in Escherichia coli vesicles. PMID- 11274129 TI - Proteins of Mycobacterium bovis BCG induced in the Wayne dormancy model. AB - Oxygen starvation triggers the shiftdown of the obligate aerobe Mycobacterium bovis BCG to a state of dormancy. Two-dimensional electrophoresis showed a drastic up-regulation of the alpha-crystallin homolog, the putative response regulator Rv3133c, and the two conserved hypothetical proteins Rv2623 and Rv2626c in dormant bacilli. PMID- 11274130 TI - Characterization of Brucella suis clpB and clpAB mutants and participation of the genes in stress responses. AB - Pathogens often encounter stressful conditions inside their hosts. In the attempt to characterize the stress response in Brucella suis, a gene highly homologous to Escherichia coli clpB was isolated from Brucella suis, and the deduced amino acid sequence showed features typical of the ClpB ATPase family of stress response proteins. Under high-temperature stress conditions, ClpB of B. suis was induced, and an isogenic B. suis clpB mutant showed increased sensitivity to high temperature, but also to ethanol stress and acid pH. The effects were reversible by complementation. Simultaneous inactivation of clpA and clpB resulted in a mutant that was sensitive to oxidative stress. In B. suis expressing gfp, ClpA but not ClpB participated in degradation of the green fluorescent protein at 42 degrees C. We concluded that ClpB was responsible for tolerance to several stresses and that the lethality caused by harsh environmental conditions may have similar molecular origins. PMID- 11274131 TI - glnD and mviN are genes of an essential operon in Sinorhizobium meliloti. AB - To evaluate the role of uridylyl-transferase, the Sinorhizobium meliloti glnD gene was isolated by heterologous complementation in Azotobacter vinelandii. The glnD gene is cotranscribed with a gene homologous to Salmonella mviN. glnD1::Omega or mviN1::Omega mutants could not be isolated by a powerful sucrose counterselection procedure unless a complementing cosmid was provided, indicating that glnD and mviN are members of an indispensable operon in S. meliloti. PMID- 11274132 TI - Characterization of outer membrane proteins in Chlamydia trachomatis LGV serovar L2. AB - We used a photoactivatable, lipophilic reagent, 3'-(trifluoromethyl)-3-(m [125I]iodophenyl)diazirine, to label proteins in the outer membrane of elementary bodies of Chlamydia trachomatis LGV serovar L2 and mass spectrometry to identify the labeled proteins. The identified proteins were polymorphic outer membrane proteins E, G, and H, which were made late in the developmental cycle, the major outer membrane protein, and a mixture of 46-kDa proteins consisting of the open reading frame 623 protein and possibly a modified form of the major outer membrane protein. PMID- 11274133 TI - Rapid dephosphorylation of the TorR response regulator by the TorS unorthodox sensor in Escherichia coli. AB - Induction of the torCAD operon, encoding the trimethylamine N-oxide (TMAO) respiratory system, is tightly controlled by the TorS-TorR phosphorelay system in response to TMAO availability. TorS is an unorthodox sensor that contains three phosphorylation sites and transphosphorylates TorR via a four-step phosphorelay, His443-->Asp723-->His850-->Asp(TorR). In this study, we provide genetic evidence that TorS can dephosphorylate phospho-TorR when TMAO is removed. Dephosphorylation probably occurs by a reverse phosphorelay, Asp(TorR)-->His850- >Asp723, since His850 and Asp723 are both essential in this process. By using reverse transcriptase PCR, we also show that TMAO removal results in shutoff of tor operon transcription in less than 2 min. Based on our results and on analogy to other phosphorelay signal transduction systems, we propose that reverse phosphotransfer could be a rapid and efficient mechanism to inactivate response regulators. PMID- 11274134 TI - Development of a new integration site within the Bacillus subtilis chromosome and construction of compatible expression cassettes. AB - The Bacillus subtilis lacA gene, coding for beta-galactosidase, has been explored as a new site able to accept DNA sequences from nonreplicating delivery vectors. Two such delivery expression vectors have been constructed and shown to be useful in obtaining regulated expression from the chromosomal location. In another experiment, it was shown that the integration of a regulatory gene at the lacA locus was able to control the expression of a transcriptional fusion at the amyE locus. These experiments demonstrate that both integration sites can be used simultaneously to obtain regulated expression of desired genes. PMID- 11274135 TI - Purification of the RelB and RelE proteins of Escherichia coli: RelE binds to RelB and to ribosomes. AB - The direct interaction of the Escherichia coli cytotoxin RelE with its specific antidote, RelB, was demonstrated in two ways: (i) copurification of the two proteins and (ii) a positive yeast two-hybrid assay involving the relB and relE genes. In addition, the purified RelE protein exhibited ribosome-binding activity in an in vitro assay, supporting previous observations suggesting that it is an inhibitor of translation. PMID- 11274137 TI - Growth inhibition caused by overexpression of the structural gene for glutamate dehydrogenase (gdhA) from Klebsiella aerogenes. AB - Two linked mutations affecting glutamate dehydrogenase (GDH) formation (gdh-1 and rev-2) had been isolated at a locus near the trp cluster in Klebsiella aerogenes. The properties of these two mutations were consistent with those of a locus containing either a regulatory gene or a structural gene. The gdhA gene from K. aerogenes was cloned and sequenced, and an insertion mutation was generated and shown to be linked to trp. A region of gdhA from a strain bearing gdh-1 was sequenced and shown to have a single-base-pair change, confirming that the locus defined by gdh-1 is the structural gene for GDH. Mutants with the same phenotype as rev-2 were isolated, and their sequences showed that the mutations were located in the promoter region of the gdhA gene. The linkage of gdhA to trp in K. aerogenes was explained by postulating an inversion of the genetic map relative to other enteric bacteria. Strains that bore high-copy-number clones of gdhA displayed an auxotrophy that was interpreted as a limitation for alpha ketoglutarate and consequently for succinyl-coenzyme A (CoA). Three lines of evidence supported this interpretation: high-copy-number clones of the enzymatically inactive gdhA1 allele showed no auxotrophy, repression of GDH expression by the nitrogen assimilation control protein (NAC) relieved the auxotrophy, and addition of compounds that could increase the alpha-ketoglutarate supply or reduce the succinyl-CoA requirement relieved the auxotrophy. PMID- 11274136 TI - Sinorhizobium meliloti plasmid pRm1132f replicates by a rolling-circle mechanism. AB - pRm1132f isolated from Sinorhizobium meliloti is a group III rolling-circle replicating (RCR) plasmid. At least seven of eight open reading frames in the nucleotide sequence represented coding regions. The minimal replicon contained a rep gene and single- and double-stranded origins of replication. Detection of single-stranded plasmid DNA confirmed that pRm1132f replicated via an RCR mechanism. PMID- 11274138 TI - Stable association of hsp90 and p23, but Not hsp70, with active human telomerase. AB - The ribonucleoprotein telomerase holoenzyme is minimally composed of a catalytic subunit, hTERT, and its associated template RNA component, hTR. We have previously found two additional components of the telomerase holoenzyme, the chaperones p23 and heat shock protein (hsp) 90, both of which are required for efficient telomerase assembly in vitro and in vivo. Both hsp90 and p23 bind specifically to hTERT and influence its proper assembly with the template RNA, hTR. We report here that the hsp70 chaperone also associates with hTERT in the absence of hTR and dissociates when telomerase is folded into its active state, similar to what occurs with other chaperone targets. Our data also indicate that hsp90 and p23 remain associated with functional telomerase complexes, which differs from other hsp90-folded enzymes that require only a transient hsp90.p23 binding. Our data suggest that components of the hsp90 chaperone complex, while required for telomerase assembly, remain associated with active enzyme, which may ultimately provide critical insight into the biochemical properties of telomerase assembly. PMID- 11274139 TI - The small heat shock protein alpha B-crystallin negatively regulates cytochrome c and caspase-8-dependent activation of caspase-3 by inhibiting its autoproteolytic maturation. AB - Caspases are universal effectors of apoptosis. The mitochondrial and death receptor pathways activate distinct apical caspases (caspase-9 and -8, respectively) that converge on the proteolytic activation of the downstream executioner caspase-3. Caspase-9 and -8 cleave procaspase-3 to produce a p24 processing intermediate (composed of its prodomain and large subunit), which then undergoes autoproteolytic cleavage to remove the prodomain from the active protease. Recently, several heat shock proteins have been shown to selectively inhibit the mitochondrial apoptotic pathway by disrupting the activation of caspase-9 downstream of cytochrome c release. We report here that the small heat shock protein alphaB-crystallin inhibits both the mitochondrial and death receptor pathways. In S-100 cytosolic extracts treated with cytochrome c/dATP or caspase-8, alphaB-crystallin inhibits the autoproteolytic maturation of the p24 partially processed caspase-3 intermediate. In contrast, neither the closely related small heat shock protein family member Hsp27 nor Hsp70 inhibited the maturation of the p24 intermediate. We also demonstrate that alphaB-crystallin co immunoprecipitates with the p24 partially processed caspase-3 in vivo. Taken together, our results demonstrate that alphaB-crystallin is a novel negative regulator of apoptosis that acts distally in the conserved cell death machinery by inhibiting the autocatalytic maturation of caspase-3. PMID- 11274140 TI - A regulatory light chain of ciliary outer arm dynein in Tetrahymena thermophila. AB - Ciliary beat frequency is primarily regulated by outer arm dyneins (22 S dynein). Chilcote and Johnson (Chilcote, T. J., and Johnson, K. A. (1990) J. Biol. Chem. 256, 17257-17266) previously studied isolated Tetrahymena 22 S dynein, identifying a protein p34, which showed cAMP-dependent phosphorylation. Here, we characterize the molecular biochemistry of p34 further, demonstrating that it is the functional ortholog of the 22 S dynein regulatory light chain, p29, in Paramecium. p34, thiophosphorylated in isolated axonemes in the presence of cAMP, co-purified with 22 S dynein and not with inner arm dynein (14 S dynein). Isolated 22 S dynein containing phosphorylated p34 showed approximately 70% increase in in vitro microtubule translocation velocity compared with its unphosphorylated counterpart. Extracted p34 rebound to isolated 22 S dynein from either Tetrahymena or Paramecium but not to 14 S dynein from either ciliate. Binding of radiolabeled p34 to 22 S dynein was competitive with p29. Phosphorylated p34 was not present in axonemes isolated from a mutant lacking outer arms. Two-dimensional gel electrophoresis followed by phosphorimaging revealed at least five phosphorylated p34-related spots, consistent with multiple phosphorylation sites in p34 or perhaps multiple isoforms of p34. These new features suggest that a class of outer arm dynein light chains including p34 regulates microtubule sliding velocity and consequently ciliary beat frequency through phosphorylation. PMID- 11274141 TI - Nuclear import of the yeast AP-1-like transcription factor Yap1p is mediated by transport receptor Pse1p, and this import step is not affected by oxidative stress. AB - The yeast AP-1-like transcription factor, Yap1p, is essential for the oxidative stress response in budding yeast. Yap1p is located predominantly in the cytoplasm; however, upon imposition of oxidative stress, Yap1p concentrates in the nucleus and activates target genes. Yap1p is constitutively transported in and out of the nucleus. Oxidative stress inhibits the Crm1p/Xpo1p-dependent nuclear export step, resulting in nuclear accumulation of Yap1p. In this study, we examined the mechanism for Yap1p nuclear import, and determined whether the import step is affected by oxidative stress. The nuclear accumulation of Yap1p required the activity of the small GTPase, Ran/Gsp1p. Under conditions in pse1-1 cells carrying a temperature-sensitive mutation of the importin beta family member PSE1/KAP121, nuclear translocation of Yap1p was inhibited dramatically. In an in vitro assay, we showed that Yap1p could directly bind to Pse1p and that this interaction was dissociated by Ran-GTP. These results indicate that Pse1p is the nuclear import receptor for Yap1p. In addition to Pse1p, we suggest that Kap123p, which is homologous to Pse1p, has a minor effect on the nuclear import of Yap1p. Furthermore, we identified the nuclear localization signal of Yap1p and demonstrated that the nuclear import of Yap1p was not affected by oxidative stress. PMID- 11274142 TI - alpha 1(Xx) collagen, a new member of the collagen subfamily, fibril-associated collagens with interrupted triple helices. AB - Chick cDNA clones for a new member of the FACIT (fibril-associated collagens with interrupted triple helices) subfamily have been isolated and sequenced. The collagen chain encoded by these cDNAs was assigned the next consecutive number, making it the alpha1(XX) collagen chain. Assignment of type XX collagen to the FACIT family was based on sequence similarities to types XII and XIV collagen. Type XX collagen mRNA is not abundant in the chick embryo. It is most prevalent in corneal epithelium. It is also detectable by reverse transcription polymerase chain reaction in embryonic skin, sternal cartilage, and tendon, but is barely detectable in calvaria, notochord, or neural retina at select stages of development, suggesting that it is not expressed in these tissues. The cDNA predicts that the alpha1(XX) collagen polypeptide is smaller than the short forms of collagen XII and XIV. A polyclonal antibody against a synthetic alpha1(XX) peptide reacts with polypeptide bands of 185, 170, and 135 kDa by Western blot analysis. From its similarity to types XII and XIV collagen, type XX is expected to bind to collagen fibrils, projecting the amino-terminal domains away from the fibrillar surface. The projecting NC 3 domains are predicted to be about half the length of those of collagen XIV. PMID- 11274143 TI - Receptor activator of NF-kappa B and osteoprotegerin expression by human microvascular endothelial cells, regulation by inflammatory cytokines, and role in human osteoclastogenesis. AB - The receptor activator of NF-kappaB (RANKL) is the essential signal required for full osteoclast (OC) development, activation, and survival. RANKL is highly expressed in areas of trabecular bone remodeling and inflammatory bone loss, is increased on marrow stromal cells or osteoblasts by osteotropic hormones or cytokines, and is neutralized by osteoprotegerin (OPG), a soluble decoy receptor also crucial for preventing arterial calcification. Vascular endothelial cells (VEC) are critically involved in bone development and remodeling and influence OC recruitment, formation, and activity. Although OCs develop and function in close association with bone VEC and sinusoids, signals mediating their interactions are not well known. Here, we show for the first time that human microvascular endothelial cells (HMVEC) express transcripts for both RANKL and OPG; inflammatory cytokines tumor necrosis factor-alpha and interleukin-1alpha elevate RANKL and OPG expression 5-40-fold in HMVEC (with an early OPG peak that declines as RANKL rises), and RANKL protein increases on the surface of tumor necrosis factor-alpha-activated HMVEC. Cytokine-activated HMVEC promoted the formation, fusion, and bone resorption of OCs formed in co-cultures with circulating human monocytic precursors via a RANKL-mediated mechanism fully antagonized by exogenous OPG. Furthermore, paraffin sections of human osteoporotic fractured bone exhibited increased RANKL immunostaining in vivo on VEC located near resorbing OCs in regions undergoing active bone turnover. Therefore, cytokine activated VEC may contribute to inflammatory-mediated bone loss via regulated production of RANKL and OPG. VEC-derived OPG may also serve as an autocrine signal to inhibit blood vessel calcification. PMID- 11274145 TI - The prototypical 4.1R-10-kDa domain and the 4.1g-10-kDa paralog mediate fodrin actin complex formation. AB - A complex family of 4.1R isoforms has been identified in non-erythroid tissues. In this study we characterized the exonic composition of brain 4.1R-10-kDa or spectrin/actin binding (SAB) domain and identified the minimal sequences required to stimulate fodrin/F-actin association. Adult rat brain expresses predominantly 4.1R mRNAs that carry an extended SAB, consisting of the alternative exons 14/15/16 and part of the constitutive exon 17. Exon 16 along with sequences carried by exon 17 is necessary and sufficient to induce formation of fodrin actin-4.1R ternary complexes. The ability of the respective SAB domains of 4.1 homologs to sediment fodrin/actin was also investigated. 4.1G-SAB stimulates association of fodrin/actin, although with an approximately 2-fold reduced efficiency compared with 4.1R-10-kDa, whereas 4.1N and 4.1B do not. Sequencing of the corresponding domains revealed that 4.1G-SAB carries a cassette that shares significant homology with 4.1R exon 16, whereas the respective sequence is divergent in 4.1N and absent from brain 4.1B. An approximately 150-kDa 4.1R and an approximately 160-kDa 4.1G isoforms are present in PC12 lysates that occur in vivo in a supramolecular complex with fodrin and F-actin. Moreover, proteins 4.1R and 4.1G are distributed underneath the plasma membrane in PC12 cells. Collectively, these observations suggest that brain 4.1R and 4.1G may modulate the membrane mechanical properties of neuronal cells by promoting fodrin/actin association. PMID- 11274144 TI - Secretory phospholipase A2 mediates cooperative prostaglandin generation by growth factor and cytokine independently of preceding cytosolic phospholipase A2 expression in rat gastric epithelial cells. AB - Transforming growth factor (TGF)-alpha and interleukin (IL)-1beta are responsible for the healing of gastric lesions through, in part, prostaglandin (PG) generation. We examined the contribution of cytosolic and secretory phospholipase A(2)s (cPLA(2) and sPLA(2)) to the PG generation by rat gastric epithelial cells in response to both stimuli. Stimulation with TGF-alpha for 24 h increased cPLA(2) and cyclooxygenase (COX)-2 markedly, PGE(2) slightly, and type IIA sPLA(2) and COX-1 not at all, whereas IL-1beta increased sPLA(2) only. Both stimuli synergistically increased PGE(2), sPLA(2), and the two COXs but not cPLA(2). The onset of the PGE(2) generation paralleled the sPLA(2) release but was apparently preceded by increases in cPLA(2) and the two COXs. The increase in PGE(2) was impaired by inhibitors for sPLA(2) and COX-2 but not COX-1. cPLA(2) inhibitors suppressed PGE(2) generation by TGF-alpha alone but not augmentation of PGE(2) generation or sPLA(2) release by IL-1beta in combination with TGF alpha. Furthermore, despite an increase in cPLA(2) including its phosphorylated form (phosphoserine), -induced arachidonic acid liberation was impaired in the TGF-alpha/IL-1beta-stimulated cells, in which p11, a putative cPLA(2) inhibitory molecule, was also increased and co-immunoprecipitated with cPLA(2). These results suggest that synergistic stimulation of sPLA(2) and COX-2 expression by TGF-alpha and IL-1beta results in an increase in PGE(2). Presumably, the preceding cPLA(2) expression is not involved in the PGE(2) generation, because of impairment of its hydrolytic activity in the stimulated cells. PMID- 11274146 TI - The adaptor protein BLNK is required for b cell antigen receptor-induced activation of nuclear factor-kappa B and cell cycle entry and survival of B lymphocytes. AB - B lymphocytes lacking the adaptor protein B cell linker (BLNK) do not proliferate in response to B cell antigen receptor (BCR) engagement. We demonstrate here that BCR-activated BLNK(-)/- B cells fail to enter the cell cycle, and this is due to their inability to induce the expression of the cell cycle regulatory proteins such as cyclin D2 and cyclin-dependent kinase 4. BCR-stimulated BLNK(-)/- B cells also do not up-regulate the cell survival protein Bcl-x(L), which may be necessary for the cells to complete the cell cycle. In addition, BLNK(-)/- B cells exhibit a high rate of spontaneous apoptosis in culture. Examination of the various BCR-activated signaling pathways in mouse BLNK(-)/- B cells reveals the intact activation of Akt and mitogen-activated protein kinases but the impaired activation of nuclear factor (NF)-kappaB that is known to regulate genes involved in cell proliferation and survival. The inability to activate NF-kappaB in BCR stimulated BLNK(-)/- B cells is due to a failure to induce the degradation of the inhibitory kappaB protein. In all these aspects, BLNK(-)/- B cells resemble xid B cells that have a mutation in Bruton's tyrosine kinase (Btk). Recently, phospholipase C (PLC)-gamma2 has also been demonstrated to be essential for NF kappaB activation. Since BLNK has been shown separately to interact with both Btk and PLC-gamma2, our finding of normal Btk but impaired PLC-gamma2 activation in BCR-stimulated BLNK(-)/- B cells strongly suggests that BLNK orchestrates the formation of a Btk-PLC-gamma2 signaling axis that regulates NF-kappaB activation. Taken together, the NF-kappaB activation defect may be sufficient to explain the similar defects in BCR-induced B cell proliferation and T cell-independent immune responses in BLNK(-)/-, Btk(-)/-, and PLC-gamma2(-)/- mice. PMID- 11274147 TI - Protein kinase C theta cooperates with Vav1 to induce JNK activity in T-cells. AB - Here we show that in human T-cell leukemia cells Vav1 and protein kinase C theta (PKCtheta) synergize for the activation of c-Jun N-terminal kinase (JNK) but not p38 MAP kinase. Vav1 and PKCtheta also cooperated to induce transcription of reporter genes controlled either by AP-1 binding sites or the CD28RE/AP composite element contained in the IL-2 promoter by stimulating the binding of transcription factors to these two elements. Dominant negative versions of Vav1 and PKCtheta inhibited CD3/CD28-induced activation of JNK, revealing their relative importance for this activation pathway. Gel filtration experiments revealed the existence of constitutively associated Vav1/PKCtheta heterodimers in extracts from unstimulated T-cells, whereas T-cell costimulation induced the recruitment of Vav1 into high molecular weight complexes. Several experimental approaches showed that Vav1 is located upstream from PKCtheta in the control of the pathway leading to synergistic JNK activation. Vav1-derived signals lead to the activation of JNK by at least two different pathways. The major contribution of Vav1 for the activation of JNK relies on the PKCtheta-mediated Ca(2+) independent synergistic activation pathway, whereas JNK is also activated by a separate Ca(2+)-dependent signaling route. PMID- 11274148 TI - Protein-protein interaction of retinoic acid receptor alpha and thyroid transcription factor-1 in respiratory epithelial cells. AB - Surfactant protein B (SP-B) is a 79-amino acid peptide critical to postnatal respiratory adaptation and is developmentally regulated. Previous studies demonstrated that retinoic acid receptors (RARs) and thyroid transcription factor 1 (TTF-1) stimulated SP-B gene expression in respiratory epithelial cells. Clustered retinoic acid-responsive element and TTF-1 binding sites were identified in the enhancer region of the SP-B gene and were required for retinoic acid stimulation of the human SP-B (hSP-B) promoter. In addition, RAR and TTF-1 were colocalized in mouse bronchiolar and alveolar type II epithelial cells, the cellular site of SP-B synthesis. In the present studies, RAR and TTF-1 were colocalized in the nucleus of H441 cells. RAR and TTF-1 synergistically stimulated the hSP-B promoter in H441 cells. Direct protein-protein interactions between RAR and TTF-1 were demonstrated by the glutathione S-transferase pull down assay and the mammalian cell two hybrid assay. Truncation/deletion studies showed that the RAR-TTF-1 interaction was mediated through the RAR DNA binding domain (DBD) and the TTF-1 homeodomain. RAR DBD greatly enhanced TTF-1 homeodomain DNA binding activity to a hSP-B enhancer oligonucleotide, in which retinoic acid-responsive element and TTF-1 DNA binding sites overlap. Chromatin immunoprecipitation assay demonstrated that retinoic acid treatment of H441 cells greatly stimulated both RAR and TTF-1 DNA binding to the hSP-B enhancer region in H441 cells. These findings support a model in which RAR/retinoid X receptor, TTF 1, and coactivators (p160 members and CBP) form an enhanceosome in the enhancer region of the hSP-B gene. PMID- 11274150 TI - Mutual antagonism of calcium entry by capacitative and arachidonic acid-mediated calcium entry pathways. AB - In nonexcitable cells, the predominant mechanism for regulated entry of Ca(2+) is capacitative calcium entry, whereby depletion of intracellular Ca(2+) stores signals the activation of plasma membrane calcium channels. A number of other regulated Ca(2+) entry pathways occur in specific cell types, however, and it is not know to what degree the different pathways interact when present in the same cell. In this study, we have examined the interaction between capacitative calcium entry and arachidonic acid-activated calcium entry, which co-exist in HEK293 cells. These two pathways exhibit mutual antagonism. That is, capacitative calcium entry is potently inhibited by arachidonic acid, and arachidonic acid activated entry is inhibited by the pre-activation of capacitative calcium entry with thapsigargin. In the latter case, the inhibition does not seem to result from a direct action of thapsigargin, inhibition of endoplasmic reticulum Ca(2+) pumps, depletion of Ca(2+) stores, or entry of Ca(2+) through capacitative calcium entry channels. Rather, it seems that a discrete step in the pathway signaling capacitative calcium entry interacts with and inhibits the arachidonic acid pathway. The findings reveal a novel process of mutual antagonism between two distinct calcium entry pathways. This mutual antagonism may provide an important protective mechanism for the cell, guarding against toxic Ca(2+) overload. PMID- 11274149 TI - Interaction of the ring finger-related U-box motif of a nuclear dot protein with ubiquitin-conjugating enzymes. AB - The U-box domain has been suggested to be a modified RING finger motif where the metal-coordinating cysteines and histidines have been replaced with other amino acids. Known U-box-containing proteins have been implicated in the ubiquitin/proteasome system. In a search for proteins interacting with the ubiquitin-conjugating enzyme UbcM4/UbcH7, we have identified a novel U-box containing protein, termed UIP5, that is exclusively found in the nucleus as part of a nuclear dot-like structure. Interaction between UbcM4 and UIP5 was observed in vivo and in vitro with bacterially expressed proteins. In addition to UbcM4, several other ubiquitin-conjugating enzymes (E2s) that share the same sequence within the L1 loop bind to UIP5. Mutational analysis showed that the U-box, like the RING finger in other proteins, forms the physical basis for the interaction with E2 enzymes. Further support for the structural similarity between U-box and RING finger comes from the observation that, in both cases, the same regions within the UbcM4 molecule are required for interaction. Our results establish at the molecular level a link between the U-box and the ubiquitin conjugating system and strongly suggest that proteins containing U-box domains are functionally closely related to RING finger proteins. PMID- 11274151 TI - A point mutation in the juxtamembrane stalk of human angiotensin I-converting enzyme invokes the action of a distinct secretase. AB - Angiotensin I-converting enzyme (ACE) is one of a number of integral membrane proteins that is proteolytically shed from the cell surface by a zinc metallosecretase. Mutagenesis of Asn(631) to Gln in the juxtamembrane stalk region of ACE resulted in more efficient secretion of the mutant protein (ACE(NQ)) as determined by pulse-chase analysis. In contrast to the wild-type ACE, the cleavage of ACE(NQ) was not blocked by the metallosecretase inhibitor batimastat but by the serine protease inhibitor, 1,3-dichloroisocoumarin. Incubation of the cells at 15 degrees C revealed that ACE(NQ) was cleaved in the endoplasmic reticulum, and mass spectrometric analysis of the secreted form of the protein indicated that it had been cleaved at the Asn(635)-Ser(636) bond, three residues N-terminal to the normal secretase cleavage site at Arg(638) Ser(639). These data clearly show that a point mutation in the juxtamembrane region of an integral membrane protein can invoke the action of a mechanistically and spatially distinct secretase. In light of this observation, previous data on the effect of mutations in the juxtamembrane stalk of shed proteins being accommodated by a single secretase having a relaxed specificity need to be re evaluated. PMID- 11274152 TI - Disruption of the interaction of mammalian protein synthesis eukaryotic initiation factor 4B with the poly(A)-binding protein by caspase- and viral protease-mediated cleavages. AB - Eukaryotic initiation factor (eIF) 4B interacts with several components of the initiation pathway and is targeted for cleavage during apoptosis. In a cell-free system, cleavage of eIF4B by caspase-3 coincides with a general inhibition of protein synthetic activity. Affinity chromatography demonstrates that mammalian eIF4B interacts with the poly(A)-binding protein and that a region consisting of the N-terminal 80 amino acids of eIF4B is both necessary and sufficient for such binding. This interaction is lost when eIF4B is cleaved by caspase-3, which removes the N-terminal 45 amino acids. Similarly, the association of eIF4B with the poly(A)-binding protein in vivo is reduced when cells are induced to undergo apoptosis. Cleavage of the poly(A)-binding protein itself, using human rhinovirus 3C protease, also eliminates the interaction with eIF4B. Thus, disruption of the association between mammalian eIF4B and the poly(A)-binding protein can occur during both apoptosis and picornaviral infection and is likely to contribute to the inhibition of translation observed under these conditions. PMID- 11274153 TI - Characterization of the Escherichia coli sigma E regulon. AB - Escherichia coli responds to the accumulation of misfolded proteins by inducing the transcription of heat shock genes. Efinal sigma(E) RNA polymerase controls one of the two heat shock regulons of E. coli. This regulon is activated upon accumulation of misfolded polypeptides in the double membrane envelope of E. coli. final sigma(E) (RpoE) is a member of the extracytoplasmic function subfamily of sigma factors. Here we asked how many genes are activated by Efinal sigma(E) RNA polymerase and what is the identity of these genes. Using two independent genetic approaches, 20 E. coli promoters were identified which activate reporter gene transcription in a final sigma(E)-dependent manner. In all cases examined, a canonical final sigma(E) binding site could be revealed upon mapping transcriptional start sites. 10 identified promoters activated the transcription of previously identified genes with four genes acting directly on the folding of E. coli envelope proteins (dsbC, fkpA, skp, and surA). The remaining promoters transcribed genes that are presumed to encode hitherto unknown extracytoplasmic functions and were named ecf (ecfA-ecfM). Two of these ecf genes were found to be essential for E. coli growth. PMID- 11274154 TI - alpha 1-Antichymotrypsin is the human plasma inhibitor of macrophage ectoenzymes that cleave pro-macrophage stimulating protein. AB - Macrophage stimulating protein (MSP) is secreted as 78-kDa single chain pro-MSP, which is converted to biologically active, disulfide-linked alphabeta chain MSP by cleavage at Arg(483)-Val(484). Murine resident peritoneal macrophages have two cell surface proteolytic activities that cleave pro-MSP. One is a pro-MSP convertase, which cleaves pro-MSP to active MSP; the other degrades pro-MSP. The degrading protease is inhibited by soybean trypsin inhibitor or by low concentrations of blood plasma, which allows the convertase to cleave pro-MSP to MSP. Using pro-MSP cleavage as the assay, we purified the inhibitor from human plasma. The bulk of the plasma protein was removed by salting out and by isoelectric precipitation of albumin. Highly purified inhibitor was then obtained in three steps: dye-ligand binding and elution, ion exchange chromatography, and high performance liquid chromatography gel filtration. After SDS-polyacrylamide gel electrophoresis and transfer to a polyvinylidene membrane, N-terminal sequencing of the product identified it as alpha(1)-antichymotrypsin. The mean concentration of alpha(1)-antichymotrypsin in human plasma is 7 micrometer. At this concentration, alpha(1)-antichymotrypsin inhibits both macrophage enzymes. A concentration of 0.4 micrometer, which is in the expected concentration range in extracellular fluid, preferentially inhibits the degrading enzyme, which allows for cleavage to active MSP by the pro-MSP convertase. PMID- 11274155 TI - Phosphatidylinositol 3-kinase, not extracellular signal-regulated kinase, regulates activation of the antioxidant-responsive element in IMR-32 human neuroblastoma cells. AB - The antioxidant-responsive element (ARE) plays an important role in the induction of phase II detoxifying enzymes including NADPH:quinone oxidoreductase (NQO1). We report herein that activation of the human NQO1-ARE (hNQO1-ARE) by tert butylhydroquinone (tBHQ) is mediated by phosphatidylinositol 3-kinase (PI3 kinase), not extracellular signal-regulated kinase (Erk1/2), in IMR-32 human neuroblastoma cells. Treatment with tBHQ significantly increased NQO1 protein without activation of Erk1/2. In addition, PD 98059 (a selective mitogen activated kinase/Erk kinase inhibitor) did not inhibit hNQO1-ARE-luciferase expression or NQO1 protein induction by tBHQ. Pretreatment with LY 294002 (a selective PI3-kinase inhibitor), however, inhibited both hNQO1-ARE-luciferase expression and endogenous NQO1 protein induction. In support of a role for PI3 kinase in ARE activation we show that: 1) transfection of IMR-32 cells with constitutively active PI3-kinase selectively activated the ARE in a dose dependent manner that was completely inhibited by treatment with LY 294002; 2) pretreatment of cells with the PI3-kinase inhibitors, LY 294002 and wortmannin, significantly decreased NF-E2-related factor 2 (Nrf2) nuclear translocation induced by tBHQ; and 3) ARE activation by constitutively active PI3-kinase was blocked completely by dominant negative Nrf2. Taken together, these data clearly show that ARE activation by tBHQ depends on PI3-kinase, which lies upstream of Nrf2. PMID- 11274156 TI - An array of positioned nucleosomes potentiates thyroid hormone receptor action in vivo. AB - The assembly of the genome into chromatin imposes a poorly understood set of rules and constraints on action by regulatory factors. We investigated the role played by chromatin infrastructure in enabling an acute response of the Xenopus TRbetaA gene to thyroid hormone receptor (TR), an extensively studied member of the nuclear hormone receptor superfamily. We found that in addition to the known TR response element (TRE) in the promoter, full range regulation required an upstream enhancer that contained multiple nonconsensus TREs and augmented ligand action at high receptor levels. An array of translationally positioned nucleosomes formed over the TRbetaA locus in vivo; unliganded TR engaged this array in linker DNA between two nucleosomes and via TREs on the surface of histone octamers. Remarkably, assembly of enhancer DNA into mature chromatin potentiated binding by TR to its target response elements and enabled a greater range of regulation by TR than was observed on immature chromatin templates. Because assembly of enhancer DNA into chromatin increased TR binding to the nonconsensus TREs, we hypothesize that chromatin disruption targeted by liganded TR to the enhancer may lead to receptor release from the template and to an attenuation of response to hormone. PMID- 11274157 TI - Recognition of tRNAs by Methionyl-tRNA transformylase from mammalian mitochondria. AB - Protein synthesis involves two methionine-isoaccepting tRNAs, an initiator and an elongator. In eubacteria, mitochondria, and chloroplasts, the addition of a formyl group gives its full functional identity to initiator Met-tRNA(Met). In Escherichia coli, it has been shown that the specific action of methionyl-tRNA transformylase on Met-tRNA(f)(Met) mainly involves a set of nucleotides in the acceptor stem, particularly a C(1)A(72) mismatch. In animal mitochondria, only one tRNA(Met) species has yet been described. It is admitted that this species can engage itself either in initiation or elongation of translation, depending on the presence or absence of a formyl group. In the present study, we searched for the identity elements of tRNA(Met) that govern its formylation by bovine mitochondrial transformylase. The main conclusion is that the mitochondrial formylase preferentially recognizes the methionyl moiety of its tRNA substrate. Moreover, the relatively small importance of the tRNA acceptor stem in the recognition process accounts for the protection against formylation of the mitochondrial tRNAs that share with tRNA(Met) an A(1)U(72) motif. PMID- 11274158 TI - Human BIN3 complements the F-actin localization defects caused by loss of Hob3p, the fission yeast homolog of Rvs161p. AB - The BAR adaptor proteins encoded by the RVS167 and RVS161 genes from Saccharomyces cerevisiae form a complex that regulates actin, endocytosis, and viability following starvation or osmotic stress. In this study, we identified a human homolog of RVS161, termed BIN3 (bridging integrator-3), and a Schizosaccharomyces pombe homolog of RVS161, termed hob3+ (homolog of Bin3). In human tissues, the BIN3 gene was expressed ubiquitously except for brain. S. pombe cells lacking Hob3p were often multinucleate and characterized by increased amounts of calcofluor-stained material and mislocalized F-actin. For example, while wild-type cells localized F-actin to cell ends during interphase, hob3Delta mutants had F-actin patches distributed randomly around the cell. In addition, medial F-actin rings were rarely found in hob3Delta mutants. Notably, in contrast to S. cerevisiae rvs161Delta mutants, hob3Delta mutants showed no measurable defects in endocytosis or response to osmotic stress, yet hob3+ complemented the osmosensitivity of a rvs161Delta mutant. BIN3 failed to rescue the osmosensitivity of rvs161Delta, but the actin localization defects of hob3Delta mutants were completely rescued by BIN3 and partially rescued by RVS161. These findings suggest that hob3+ and BIN3 regulate F-actin localization, like RVS161, but that other roles for this gene have diverged somewhat during evolution. PMID- 11274159 TI - Dual targeting of spinach protoporphyrinogen oxidase II to mitochondria and chloroplasts by alternative use of two in-frame initiation codons. AB - Protoporphyrinogen oxidase (Protox) is the final enzyme in the common pathway of chlorophyll and heme biosynthesis. Two Protox isoenzymes have been described in tobacco, a plastidic and a mitochondrial form. We isolated and sequenced spinach Protox cDNA, which encodes a homolog of tobacco mitochondrial Protox (Protox II). Alignment of the deduced amino acid sequence between Protox II and other tobacco mitochondrial Protox homologs revealed a 26-amino acid N-terminal extension unique to the spinach enzyme. Immunoblot analysis of spinach leaf extract detected two proteins with apparent molecular masses of 57 and 55 kDa in chloroplasts and mitochondria, respectively. In vitro translation experiments indicated that two translation products (59 and 55 kDa) are produced from Protox II mRNA, using two in-frame initiation codons. Transport experiments using green fluorescent protein-fused Protox II suggested that the larger and smaller translation products (Protox IIL and IIS) target exclusively to chloroplasts and mitochondria, respectively. PMID- 11274160 TI - Caenorhabditis elegans PIAK, a phospholipid-independent kinase that activates the AKT/PKB survival kinase. AB - Phospholipid-dependent kinase 1 (PDK 1) is a 3'-phospholipid-responsive serine/threonine kinase that plays a critical role in cell survival by phosphorylating and activating the anti-apoptotic AKT/PKB kinase. While PDK 1 is clearly an important component of the cell survival machinery, the potential for phospholipid-independent activation of the AKT/PKB survival pathway has not been extensively examined at the molecular level. We have identified a second form of PDK 1 in the nematode Caenorhabditis elegans that we have termed PIAK (phospholipid-independent AKT/PKB kinase). PIAK is highly homologous to C. elegans and mammalian PDK 1 with the exception that the novel kinase lacks a phospholipid binding pleckstrin homology domain. The domain structure of PIAK suggests that it might be a phospholipid-independent kinase, and PIAK phosphorylates mammalian AKT/PKB at the activating Thr(308) residue in the presence of the phosphatidylinositol (PI) 3-kinase inhibitors as well as in the absence of growth factors. In addition, PIAK is capable of inducing the phospholipid-independent, AKT/PKB-induced phosphorylation of the AFX-type forkhead transcription factor, resulting in its cytoplasmic localization. Because the nuclear localization of this transcription factor induces an apoptotic state, this PIAK-mediated cytoplasmic sequestration allows for cell survival. Finally, PIAK activity appears to be induced by various inhibitors of cell cycle G(1) progression. These data suggest an alternate, phosphatidylinositol 3-kinase independent mechanism for the activation of the AKT/PKB survival pathway that may be utilized during periods of cellular quiescence. PMID- 11274161 TI - C-terminal fragments of the alpha 1C (CaV1.2) subunit associate with and regulate L-type calcium channels containing C-terminal-truncated alpha 1C subunits. AB - L-type Ca(2+) channels in native tissues have been found to contain a pore forming alpha(1) subunit that is often truncated at the C terminus. However, the C terminus contains many important domains that regulate channel function. To test the hypothesis that C-terminal fragments may associate with and regulate C terminal-truncated alpha(1C) (Ca(V)1.2) subunits, we performed electrophysiological and biochemical experiments. In tsA201 cells expressing either wild type or C-terminal-truncated alpha(1C) subunits in combination with a beta(2a) subunit, truncation of the alpha(1C) subunit by as little as 147 amino acids led to a 10-15-fold increase in currents compared with those obtained from control, full-length alpha(1C) subunits. Dialysis of cells expressing the truncated alpha(1C) subunits with C-terminal fragments applied through the patch pipette reconstituted the inhibition of the channels seen with full-length alpha(1C) subunits. In addition, C-terminal deletion mutants containing a tethered C terminus also exhibited the C-terminal-induced inhibition. Immunoprecipitation assays demonstrated the association of the C-terminal fragments with truncated alpha(1C) subunits. In addition, glutathione S transferase pull-down assays demonstrated that the C-terminal inhibitory fragment could associate with at least two domains within the C terminus. The results support the hypothesis the C- terminal fragments of the alpha(1C) subunit can associate with C-terminal-truncated alpha(1C) subunits and inhibit the currents through L-type Ca(2+) channels. PMID- 11274162 TI - A family of yeast proteins mediating bidirectional vacuolar amino acid transport. AB - Seven genes in Saccharomyces cerevisiae are predicted to code for membrane spanning proteins (designated AVT1-7) that are related to the neuronal gamma aminobutyric acid-glycine vesicular transporters. We have now demonstrated that four of these proteins mediate amino acid transport in vacuoles. One protein, AVT1, is required for the vacuolar uptake of large neutral amino acids including tyrosine, glutamine, asparagine, isoleucine, and leucine. Three proteins, AVT3, AVT4, and AVT6, are involved in amino acid efflux from the vacuole and, as such, are the first to be shown directly to transport compounds from the lumen of an acidic intracellular organelle. This function is consistent with the role of the vacuole in protein degradation, whereby accumulated amino acids are exported to the cytosol. Protein AVT6 is responsible for the efflux of aspartate and glutamate, an activity that would account for their exclusion from vacuoles in vivo. Transport by AVT1 and AVT6 requires ATP for function and is abolished in the presence of nigericin, indicating that the same pH gradient can drive amino acid transport in opposing directions. Efflux of tyrosine and other large neutral amino acids by the two closely related proteins, AVT3 and AVT4, is similar in terms of substrate specificity to transport system h described in mammalian lysosomes and melanosomes. These findings suggest that yeast AVT transporter function has been conserved to control amino acid flux in vacuolar-like organelles. PMID- 11274163 TI - Subcellular localization of the autoimmune regulator protein. characterization of nuclear targeting and transcriptional activation domain. AB - The autoimmune regulator (AIRE) gene, defective in the hereditary autoimmune disease APECED, encodes a transcriptional regulator protein. AIRE is expressed in the medullary epithelial cells and monocyte-dendritic cells of the thymus with lower expression in the spleen, fetal liver, and lymph nodes. At the cellular level, AIRE is located in microtubular structures of the cytoskeleton and in discrete nuclear dots resembling ND10 nuclear bodies. We studied the determinants of the targeting of AIRE into these structures. We report here that the N terminal HSR domain confers localization to the microtubular network whereas the C-terminal region contains a second nuclear localization signal. We also demonstrate that the consensus nuclear localization signal of AIRE is functional and that the HSR domain harbors a nuclear export signal. Accordingly, the nuclear export inhibitor leptomycin B partially inhibits the nuclear export of AIRE. From a functional standpoint, we show that AIRE can activate the interferon beta minimal promoter in a transfection assay and demonstrate that the transcriptional activating function of AIRE is mediated by its two plant homeodomain (PHD) zinc fingers. PMID- 11274164 TI - Promoter choice influences alternative splicing and determines the balance of isoforms expressed from the mouse bcl-X gene. AB - Differential splicing from the bcl-X gene generates several isoforms with opposite effects on the apoptotic response. To explore the mechanism controlling the balance between the various isoforms, we have characterized the 5' region of the mouse bcl-X gene. We identified three new promoters in addition to the two previously described (Grillot, D. A., M., G.-G., Ekhterae, D., Duan, L., Inohara, N., Ohta, S., Seldin, M. F., and Nunez, G. (1997) J. Immunol. 158, 4750-4757). These five promoters (P1-P5) would give rise to at least five mRNAs with different 5'-untranslated region, all sharing the same translation initiation site. Except for the product of the most proximal promoter (P1), the other mRNAs are generated by alternative splicing of noncoding exons to a common acceptor site located in the first translated exon. Reverse transcriptase-polymerase chain reaction, primer extension, and RNase protection assays demonstrate a tissue specific pattern of promoter usage. P1 and P2 are active in all tissues analyzed, whereas the other three promoter show tissue-specific activities. P3 is active in spleen, liver, and kidney, P4 is active in uterus and spleen, and P5 is active in spleen, liver, brain, and thymus. We present evidence suggesting that promoter selection influences the outcome of the splice process. Transcripts from P1 generate mainly the mRNA for the long isoform Bcl-X(L), whereas transcripts from P2 generate mRNAs for the isoforms Bcl-X(L), Bcl-X(S), and Bcl-X(gamma) and transcripts from P3 yield mainly mRNAs for the isoform Bcl-X(gamma). Our results suggest a key role of promoter choice in determining alternative splicing and, thus, the balance of Bcl-X isoforms. PMID- 11274165 TI - Lipopolysaccharide is in close proximity to each of the proteins in its membrane receptor complex. transfer from CD14 to TLR4 and MD-2. AB - The structural features of some proteins of the innate immune system involved in mediating responses to microbial pathogens are highly conserved throughout evolution. Examples include members of the Drosophila Toll (dToll) and the mammalian Toll-like receptor (TLR) protein families. Activation of Drosophila Toll is believed to occur via an endogenous peptide rather than through direct binding of microbial products to the Toll protein. In mammals there is a growing consensus that lipopolysaccharide (LPS) initiates its biological activities through a heteromeric receptor complex containing CD14, TLR4, and at least one other protein, MD-2. LPS binds directly to CD14 but whether LPS then binds to TLR4 and/or MD-2 is not known. We have used transient transfection to express human TLRs, MD-2, or CD14 alone or in different combinations in HEK 293 cells. Interactions between LPS and these proteins were studied using a chemically modified, radioiodinated LPS containing a covalently linked, UV light-activated cross-linking group ((125)I-ASD-Re595 LPS). Here we show that LPS is cross-linked specifically to TLR4 and MD-2 only when co-expressed with CD14. These data support the contention that LPS is in close proximity to the three known proteins of its membrane receptor complex. Thus, LPS binds directly to each of the members of the tripartite LPS receptor complex. PMID- 11274166 TI - Regulation of human CLC-3 channels by multifunctional Ca2+/calmodulin-dependent protein kinase. AB - The multifunctional calcium/calmodulin-dependent protein kinase II, CaMKII, has been shown to regulate chloride movement and cellular function in both excitable and non-excitable cells. We show that the plasma membrane expression of a member of the ClC family of Cl(-) channels, human CLC-3 (hCLC-3), a 90-kDa protein, is regulated by CaMKII. We cloned the full-length hCLC-3 gene from the human colonic tumor cell line T84, previously shown to express a CaMKII-activated Cl(-) conductance (I(Cl,CaMKII)), and transfected this gene into the mammalian epithelial cell line tsA, which lacks endogenous expression of I(Cl,CaMKII). Biotinylation experiments demonstrated plasma membrane expression of hCLC-3 in the stably transfected cells. In whole cell patch clamp experiments, autonomously active CaMKII was introduced into tsA cells stably transfected with hCLC-3 via the patch pipette. Cells transfected with the hCLC-3 gene showed a 22-fold increase in current density over cells expressing the vector alone. Kinase dependent current expression was abolished in the presence of the autocamtide-2 related inhibitory peptide, a specific inhibitor of CaMKII. A mutation of glycine 280 to glutamic acid in the conserved motif in the putative pore region of the channel changed anion selectivity from I(-) > Cl(-) to Cl(-) > I(-). These results indicate that hCLC-3 encodes a Cl(-) channel that is regulated by CaMKII dependent phosphorylation. PMID- 11274167 TI - Targeted and extended acetylation of histones H4 and H3 at active and inactive genes in chicken embryo erythrocytes. AB - Affinity-purified polyclonal antibodies recognizing the most highly acetylated forms of histones H3 and H4 were used in immunoprecipitation assays with chromatin fragments derived from 15-day chicken embryo erythrocytes by micrococcal nuclease digestion. The distribution of hyperacetylated H4 and H3 was mapped at the housekeeping gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and the tissue-specific gene, carbonic anhydrase (CA). H3 and H4 acetylation was found targeted to the CpG island region at the 5' end of both these genes, falling off in the downstream direction. In contrast, at the beta(A) globin gene, both H3 and H4 are highly acetylated throughout the gene and at the downstream enhancer, with a maximum at the promoter. Low level acetylation was observed at the 5' end of the inactive ovalbumin gene. Run-on assays to measure ongoing transcription showed that the GAPDH and CA genes are transcribed at a much lower rate than the adult beta(A)-globin gene. The extensive high level acetylation at the beta(A)-globin gene correlates most simply with its high rate of transcription. The targeted acetylation of histones H3 and H4 at the GAPDH and CA genes is consistent with a role in transcriptional initiation and implies that transcriptional elongation does not necessarily require hyperacetylation. PMID- 11274168 TI - Ca2+/calmodulin-dependent protein kinase IV stimulates nuclear factor-kappa B transactivation via phosphorylation of the p65 subunit. AB - Calmodulin-dependent protein kinase IV (CaMKIV) is a key mediator of Ca(2+) induced gene expression. In this study, CaMKIV was found to directly associate with and phosphorylate the nuclear factor-kappaB (NFkappaB) component p65 both in vitro and in vivo. The phosphorylation of p65 by CaMKIV resulted in recruitment of transcription coactivator cAMP-response element-binding protein-binding protein and concomitant release of corepressor silencing mediator for retinoid and thyroid hormone receptors, as demonstrated by the glutathione S-transferase pull down and mammalian two hybrid assays. In addition, cotransfection of CaMKIV resulted in cytosolic translocation of the silencing mediator for retinoid and thyroid hormone receptors. Consistent with these results, cotransfected CaMKIV dramatically stimulated the NFkappaB transactivation in mammalian cells. From these results, NFkappaB is suggested to be a novel downstream effector molecule of CaMKIV. PMID- 11274169 TI - AP-1 and Cbfa/runt physically interact and regulate parathyroid hormone-dependent MMP13 expression in osteoblasts through a new osteoblast-specific element 2/AP-1 composite element. AB - The expression of MMP13 (collagenase-3), a member of the matrix metalloproteinase family, is increased in vivo as well as in cultured osteosarcoma cell lines by parathyroid hormone (PTH), a major regulator of calcium homeostasis. Binding sites for AP-1 and Cbfa/Runt transcription factors in close proximity have been identified as cis-acting elements in the murine and rat mmp13 promoter required for PTH-induced expression. The cooperative function of these factors in response to PTH in osteoblastic cells suggests a direct interaction between AP-1 and Cbfa/Runt transcription factors. Here, we demonstrate interaction between c-Jun and c-Fos with Cbfa/Runt proteins. This interaction depends on the leucine zipper of c-Jun or c-Fos and the Runt domain of Cbfa/Runt proteins, respectively. Moreover, c-Fos interacts with the C-terminal part of Cbfa1 and Cbfa2, sharing a conserved transcriptional repression domain. In addition to the distal osteoblast specific element 2 (OSE2) element in the murine and rat mmp13 promoter, we identified a new proximal OSE2 site overlapping with the TRE motif. Both interaction of Cbfa/Runt proteins with AP-1 and the presence of a functional proximal OSE2 site are required for enhanced transcriptional activity of the mmp13 promoter in transient transfected fibroblasts and in PTH-treated osteosarcoma cells. PMID- 11274171 TI - Targeted inactivation of Gh/tissue transglutaminase II. AB - The novel G-protein, G(h)/tissue transglutaminase (TGase II), has both guanosine triphosphatase and Ca(2+)-activated transglutaminase activity and has been implicated in a number of processes including signal transduction, apoptosis, bone ossification, wound healing, and cell adhesion and spreading. To determine the role of G(h) in vivo, the Cre/loxP site-specific recombinase system was used to develop a mouse line in which its expression was ubiquitously inactivated. Despite the absence of G(h) expression and a lack of intracellular TGase activity that was not compensated by other TGases, the Tgm2(-/-) mice were viable, phenotypically normal, and were born with the expected Mendelian frequency. Absence of G(h) coupling to alpha(1)-adrenergic receptor signaling in Tgm2(-/-) mice was demonstrated by the lack of agonist-stimulated [alpha-(32)P]GTP photolabeling of a 74-kDa protein in liver membranes. Annexin-V positivity observed with dexamethasone-induced apoptosis was not different in Tgm2(-/-) thymocytes compared with Tgm2(+/+) thymocytes. However, with this treatment there was a highly significant decrease in the viability (propidium iodide negativity) of Tgm2(-/-) thymocytes. Primary fibroblasts isolated from Tgm2(-/-) mice also showed decreased adherence with culture. These results indicate that G(h) may be importantly involved in stabilizing apoptotic cells before clearance, and in responses such as wound healing that require fibroblast adhesion mediated by extracellular matrix cross-linking. PMID- 11274170 TI - PEA3 is up-regulated in response to Wnt1 and activates the expression of cyclooxygenase-2. AB - The inducible prostaglandin synthase cyclooxygenase-2 (COX-2) is aberrantly expressed in intestinal tumors resulting from APC mutation, and is also transcriptionally up-regulated in mouse mammary epithelial cells in response to Wnt1 expression. beta-Catenin stabilization is a consequence of both APC mutation and Wnt signaling. We have previously observed coordinate regulation of the matrilysin promoter by beta-catenin and Ets family transcription factors of the PEA3 subfamily. Here we show that while beta-catenin only weakly activates the COX-2 promoter, PEA3 family transcription factors are potent activators of COX-2 transcription. Consistent with this, PEA3 is up-regulated in Wnt1-expressing mouse mammary epithelial cells, and PEA3 factors are highly expressed in tumors from Wnt1 transgenic mice, in which Cox-2 is also up-regulated. Promoter mapping experiments suggest that the NF-IL6 site in the COX-2 promoter is important for mediating PEA3 responsiveness. The NF-IL6 site is also important for COX-2 transcription in some colorectal cancer lines (Shao, J., Sheng, H., Inoue, H., Morrow, J. D., and DuBois, R. N. (2000) J. Biol. Chem. 275, 33951-33956), and PEA3 factors are highly expressed in colorectal cancer cell lines. Therefore, we speculate that PEA3 factors may contribute to the up-regulation of COX-2 expression resulting from both APC mutation and Wnt1 expression. PMID- 11274172 TI - A repressor with similarities to prokaryotic and eukaryotic DNA helicases controls the assembly of the CAAT box binding complex at a photosynthesis gene promoter. AB - A single nucleotide exchange in a promoter region located immediately upstream of the CAAT box of the spinach photosynthesis gene AtpC (gene product is subunit gamma of the chloroplast ATP synthase) prevents the formation of a secondary structure and causes an unregulated, constitutive high level of expression (Kusnetsov, V., Landsberger, M., Oelmuller, R. (1999) J. Biol. Chem. 274, 36009 36014). We have isolated cDNAs for ATPC-2, a new polypeptide with homologies to pro- and eukaryotic helicases, which specifically binds to this promoter region. Binding of ATPC-2 competes strongly with that of a CAAT box binding factor (CBF), consistent with the idea that both complexes cannot be formed simultaneously because of sterical reasons. In gel mobility shift assays, high binding activities of ATPC-2 and low binding activities of CBF were observed with nuclear extracts from tissue with low AtpC expression levels, and the opposite was observed with extracts from tissues with high AtpC expression levels. Binding of ATPC-2 to the mutant sequence, which directs a constitutively high level expression in vivo and prevents the formation of a secondary structure in vitro, is significantly weaker than binding to the wild-type sequence. Again, the opposite results were obtained for the CBF. Thus, we conclude that the assembly of the CBF.DNA complex stimulates transcription of AtpC and that CBF binding is prevented if ATPC-2 is bound to the promoter region. The novel mechanism of gene regulation and the role of the helicase-like protein ATPC-2 as a potential transcriptional repressor is discussed in relation to its modular structure. PMID- 11274174 TI - Control of cystic fibrosis transmembrane conductance regulator expression by BAP31. AB - Expression of the cystic fibrosis transmembrane conductance regulator (CFTR) is stringently controlled by molecular chaperones participating in formation of the quality control system. It has been shown that about 75% of all CFTR protein and close to 100% of the [DeltaPhe(508)] CFTR variant are rapidly degraded before leaving the endoplasmic reticulum (ER). B cell antigen receptor-associated proteins (BAPs) are ubiquitously expressed integral membrane proteins that may control association with the cytoskeleton, vesicular transport, or retrograde transport from the cis Golgi to the ER. The present study delivers evidence for cytosolic co-localization of both BAP31 and CFTR and for the control of expression of recombinant CFTR in Chinese hamster ovary (CHO) cells and Xenopus oocytes by BAP31. Antisense inhibition of BAP31 in various cell types increased expression of both wild-type CFTR and [DeltaPhe(508)]CFTR and enabled cAMP activated Cl(-) currents in [DeltaPhe(508)]CFTR-expressing CHO cells. Coexpression of CFTR together with BAP31 attenuated cAMP-activated Cl(-) currents in Xenopus oocytes. These data therefore suggest association of BAP31 with CFTR that may control maturation or trafficking of CFTR and thus expression in the plasma membrane. PMID- 11274175 TI - Prostasin is a glycosylphosphatidylinositol-anchored active serine protease. AB - A recombinant human prostasin serine protease was expressed in several human cell lines. Subcellular fractionation showed that this serine protease is synthesized as a membrane-bound protein while a free-form prostasin is secreted into the culture medium. Prostasin was identified in nuclear and membrane fractions. Membrane-bound prostasin can be released by phosphatidylinositol-specific phospholipase C treatment, or labeled by [(3)H]ethanolamine, indicating a glycosylphosphatidylinositol anchorage. A prostasin-binding protein was identified in mouse and human seminal vesicle fluid. Both the secreted and the membrane-bound prostasin were able to form a covalently linked 82-kDa complex when incubated with seminal vesicle fluid. The complex formation between prostasin and the prostasin-binding protein was inhibited by a prostasin antibody, heparin, and serine protease inhibitors. Prostasin's serine protease activity was inhibited when bound to the prostasin-binding protein in mouse seminal vesicle fluid. This study indicates that prostasin is an active serine protease in its membrane-bound form. PMID- 11274176 TI - Mycobacterium bovis Bacillus Calmette-Guerin and its cell wall complex induce a novel lysosomal membrane protein, SIMPLE, that bridges the missing link between lipopolysaccharide and p53-inducible gene, LITAF(PIG7), and estrogen-inducible gene, EET-1. AB - LITAF and PIG7 encode an identical protein, and they have recently been reported as lipopolysaccharide and p53-inducible genes, respectively. By using the differential display approach, we identified a Mycobacterium bovis BCG cell wall skeleton (BCG-CWS)-inducible gene fragment from human monocytes, showing no homology to any reported gene. Full-length cloning of this fragment reveals the following. 1) The differential display product represents the incomplete 3' untranslated region of LITAF/PIG7. 2) The coding region of the transcript differs from LITAF/PIG7 due to an absence of a single guanine residue, resulting in a potential translational frameshift. 3) The newly coded protein turns out to be 86% identical and 90% similar to an estrogen-inducible rat gene, EET-1. Repeated analysis, expressed sequence tag search, comparison with homologues, and genome sequence analysis confirmed the absence of the single guanine residue. One interesting feature of this protein is that it possesses the RING domain signature and is predicted to be localized in the nucleus. However, detailed analysis together with experimental evidence suggests it is neither a RING family member nor a nuclear protein. Comparison of a total collection of 18 proteins from various species indicates that proteins of this family are small in size and mainly conserved at the C-terminal domain with a unique motif. We characterize this novel protein as an unglycosylated small integral membrane protein of the lysosome/late endosome (SIMPLE) whose expression is elicited in monocytes by live and heat-killed BCG, BCG cell wall complex, lipopolysaccharide, and tumor necrosis factor-alpha. To our knowledge this is the first report of pathogen associated molecular pattern (PAMP)-induced differential expression of a lysosomal membrane protein presumably involved in apoptosis. PMID- 11274177 TI - Characterization of the D-glucuronyl C5-epimerase involved in the biosynthesis of heparin and heparan sulfate. AB - The murine gene for the glucuronyl C5-epimerase involved in heparan sulfate biosynthesis was cloned, using a previously isolated bovine lung cDNA fragment (Li, J.-P., Hagner-McWhirter, A., Kjellen, L., Palgi, J., Jalkanen, M., and Lindahl, U. (1997) J. Biol. Chem. 272, 28158-28163) as probe. The approximately 11-kilobase pair mouse gene contains 3 exons from the first ATG to stop codon and is localized to chromosome 9. Southern analysis of the genomic DNA and chromosome mapping suggested the occurrence of a single epimerase gene. Based on the genomic sequence, a mouse liver cDNA was isolated that encodes a 618-amino acid residue protein, thus extending by 174 N-terminal residues the sequence deduced from the (incomplete) bovine cDNA. Comparison of murine, bovine, and human epimerase cDNA structures indicated 96-99% identity at the amino acid level. A cDNA identical to the mouse liver species was demonstrated in mouse mast cells committed to heparin biosynthesis. These findings suggest that the iduronic acid residues in heparin and heparan sulfate, despite different structural contexts, are generated by the same C5-epimerase enzyme. The catalytic activity of the recombinant full-length mouse liver epimerase, expressed in insect cells, was found to be >2 orders of magnitude higher than that of the previously cloned, smaller bovine recombinant protein. The approximately 52-kDa, similarly highly active, enzyme originally purified from bovine liver (Campbell, P., Hannesson, H. H., Sandback, D., Roden, L., Lindahl, U., and Li, J.-P. (1994) J. Biol. Chem. 269, 26953-26958) was found to be associated with an approximately 22-kDa peptide generated by a single proteolytic cleavage of the full-sized protein. PMID- 11274178 TI - Spatial analysis of key signaling proteins by high-content solid-phase cytometry in Hep3B cells treated with an inhibitor of Cdc25 dual-specificity phosphatases. AB - Protein phosphorylation frequently results in the subcellular redistribution of key signaling molecules, and this spatial change is critical for their activity. Here we have probed the effects of a Cdc25 inhibitor, 2-(2-mercaptoethanol)-3 methyl-1,4-naphthoquinone, or Compound 5, on the spatial regulation and activation kinetics of tyrosine phosphorylation-dependent signaling events using two methods: (i) high-content, automated, fluorescence-based, solid-phase cytometry and (ii) a novel cellular assay for Cdc25A activity in intact cells. Immunofluorescence studies demonstrated that Compound 5 produced a concentration dependent nuclear accumulation of phospho-Erk and phospho-p38, but not nuclear factor kappaB. Immunoblot analysis confirmed Erk phosphorylation and nuclear accumulation, and in vitro kinase assays showed that Compound 5-activated Erk was competent to phosphorylate its physiological substrate, the transcription factor Elk-1. Pretreatment of cells with the MEK inhibitor U-0126 prevented the induction by Compound 5 of phospho-Erk (but not phospho-p38) nuclear accumulation and protected cells from the antiproliferative effects of Compound 5. Overexpression of Cdc25A in whole cells caused dephosphorylation of Erk that was reversed by Compound 5. The data show that an inhibitor of Cdc25 increases Erk phosphorylation and nuclear accumulation and support the hypothesis that Cdc25A regulates Erk phosphorylation status. PMID- 11274179 TI - Microtubule integrity regulates Pak leading to Ras-independent activation of Raf 1. insights into mechanisms of Raf-1 activation. AB - Growth factors activate Raf-1 by engaging a complex program, which requires Ras binding, membrane recruitment, and phosphorylation of Raf-1. The present study employs the microtubule-depolymerizing drug nocodazole as an alternative approach to explore the mechanisms of Raf activation. Incubation of cells with nocodazole leads to activation of Pak1/2, kinases downstream of small GTPases Rac/Cdc42, which have been previously indicated to phosphorylate Raf-1 Ser(338). Nocodazole induced stimulation of Raf-1 is augmented by co-expression of small GTPases Rac/Cdc42 and Pak1/2. Dominant negative mutants of these proteins block activation of Raf-1 by nocodazole, but not by epidermal growth factor (EGF). Thus, our studies define Rac/Cdc42/Pak as a module upstream of Raf-1 during its activation by microtubule disruption. Although it is Ras-independent, nocodazole induced activation of Raf-1 appears to involve the amino-terminal regulatory region in which the integrity of the Ras binding domain is required. Surprisingly, the Raf zinc finger mutation (C165S/C168S) causes a robust activation of Raf-1 by nocodazole, whereas it diminishes Ras-dependent activation of Raf-1. We also show that mutation of residues Ser(338) to Ala or Tyr(340) Tyr(341) to Phe-Phe immediately amino-terminal to the catalytic domain abrogates activation of both the wild type and zinc finger mutant Raf by both EGF/4beta-12 O-tetradecanoylphorbol-13-acetate and nocodazole. Finally, an in vitro kinase assay demonstrates that the zinc finger mutant serves as a better substrate of Pak1 than the wild type Raf-1. Collectively, our results indicate that 1) the zinc finger exerts an inhibitory effect on Raf-1 activation, probably by preventing phosphorylation of (338)SSYY(341); 2) such inhibition is first overcome by an unknown factor binding in place of Ras-GTP to the amino-terminal regulatory region in response to nocodazole; and 3) EGF and nocodazole utilize different kinases to phosphorylate Ser(338), an event crucial for Raf activation. PMID- 11274180 TI - Determination of the complete amino acid sequence for the coat protein of brome mosaic virus by time-of-flight mass spectrometry. Evidence for mutations associated with change of propagation host. AB - Time-of-flight mass spectrometry (TOFMS) has been applied to determine the complete coat protein amino acid sequences of a number of distinct brome mosaic virus (BMV) isolates. Ionization was carried out by both electrospray ionization and matrix-assisted laser desorption/ionization (MALDI). After determining overall coat protein masses, the proteins were digested with trypsin or Lys-C proteinases, and the digestion products were analyzed in a MALDI QqTOF mass spectrometer. The N terminus of the coat protein was found to be acetylated in each BMV isolate analyzed. In one isolate (BMV-Valverde), the amino acid sequence was identical to that predicted from the cDNA sequence of the "type" isolate, but deviations from the predicted amino acid sequence were observed for all the other isolates analyzed. When isolates were propagated in different host taxa, modified coat protein sequences were observed in some cases, along with the original sequence. Sequencing by TOFMS may therefore provide a basis for monitoring the effects of host passaging on a virus at the molecular level. Such TOFMS-based analyses assess the complete profiles of coat protein sequences actually present in infected tissues. They are therefore not subject to the selection biases inherent in deducing such sequences from reverse-transcribed viral RNA and cloning the resulting cDNA. PMID- 11274181 TI - Anions modulate the potency of geranylgeranyl-protein transferase I inhibitors. AB - We have identified and characterized potent and specific inhibitors of geranylgeranyl-protein transferase type I (GGPTase I), as well as dual inhibitors of GGPTase I and farnesyl-protein transferase. Many of these inhibitors require the presence of phosphate anions for maximum activity against GGPTase I in vitro. Inhibitors with a strong anion dependence were competitive with geranylgeranyl pyrophosphate (GGPP), rather than with the peptide substrate, which had served as the original template for inhibitor design. One of the most effective anions was ATP, which at low millimolar concentrations increased the potency of GGPTase I inhibitors up to several hundred-fold. In the case of clinical candidate l 778,123, this increase in potency was shown to result from two major interactions: competitive binding of inhibitor and GGPP, and competitive binding of ATP and GGPP. At 5 mm, ATP caused an increase in the apparent K(d) for the GGPP-GGPTase I interaction from 20 pm to 4 nm, resulting in correspondingly tighter inhibitor binding. A subset of very potent GGPP-competitive inhibitors displayed slow tight binding to GGPTase I with apparent on and off rates on the order of 10(6) m(-)1 s(-)1 and 10(-)3 s(-)1, respectively. Slow binding and the anion requirement suggest that these inhibitors may act as transition state analogs. After accounting for anion requirement, slow binding, and mechanism of competition, the structure-activity relationship determined in vitro correlated well with the inhibition of processing of GGPTase I substrate Rap1a in vivo. PMID- 11274182 TI - Structural compatibility between the putative voltage sensor of voltage-gated K+ channels and the prokaryotic KcsA channel. AB - Sequence similarity among and electrophysiological studies of known potassium channels, along with the three-dimensional structure of the Streptomyces lividans K(+) channel (KcsA), support the tenet that voltage-gated K(+) channels (Kv channels) consist of two distinct modules: the "voltage sensor" module comprising the N-terminal portion of the channel up to and including the S4 transmembrane segment and the "pore" module encompassing the C-terminal portion from the S5 transmembrane segment onward. To substantiate this modular design, we investigated whether the pore module of Kv channels may be replaced with the pore module of the prokaryotic KcsA channel. Biochemical and immunocytochemical studies showed that chimeric channels were expressed on the cell surface of Xenopus oocytes, demonstrating that they were properly synthesized, glycosylated, folded, assembled, and delivered to the plasma membrane. Unexpectedly, surface expressed homomeric chimeras did not exhibit detectable voltage-dependent channel activity upon both hyperpolarization and depolarization regardless of the expression system used. Chimeras were, however, strongly dominant-negative when coexpressed with wild-type Kv channels, as evidenced by the complete suppression of wild-type channel activity. Notably, the dominant-negative phenotype correlated well with the formation of stable, glycosylated, nonfunctional, heteromeric channels. Collectively, these findings imply a structural compatibility between the prokaryotic pore module and the eukaryotic voltage sensor domain that leads to the biogenesis of non-responsive channels. Our results lend support to the notion that voltage-dependent channel gating depends on the precise coupling between both protein domains, probably through a localized interaction surface. PMID- 11274183 TI - G alpha minigenes expressing C-terminal peptides serve as specific inhibitors of thrombin-mediated endothelial activation. AB - The C termini of G protein alpha subunits are critical for binding to their cognate receptors, and peptides corresponding to the C terminus can serve as competitive inhibitors of G protein-coupled receptor-G protein interactions. This interface is quite specific as a single amino acid difference annuls the ability of a G alpha(i) peptide to bind the A(1) adenosine receptor (Gilchrist, A., Mazzoni, M., Dineen, B., Dice, A., Linden, J., Dunwiddie, T., and Hamm, H. E. (1998 ) J. Biol. Chem. 273, 14912--14919). Recently, we demonstrated that a plasmid minigene vector encoding the C-terminal sequence of G alpha(i) could specifically inhibit downstream responses to agonist stimulation of the muscarinic M(2) receptor (Gilchrist, A., Bunemann, M., Li, A., Hosey, M. M., and H. E. Hamm (1999) J. Biol. Chem. 274, 6610--6616). To selectively antagonize G protein signal transduction events and determine which G protein underlies a given thrombin-induced response, we generated minigene vectors that encode the C terminal sequence for each family of G alpha subunits. Minigene vectors expressing G alpha C-terminal peptides (G alpha(i), G alpha(q), G alpha(12), and G alpha(13)) or the control minigene vector, which expresses the G alpha(i) peptide in random order (G(iR)), were systematically introduced into a human microvascular endothelial cell line. The C-terminal peptides serve as competitive inhibitors presumably by blocking the site on the G protein-coupled receptor that normally binds the G protein. Our results not only confirm that each G protein can control certain signaling events, they emphasize the specificity of the G protein-coupled receptor-G protein interface. In addition, the C-terminal G alpha minigenes appear to be a powerful tool for dissecting out the G protein that mediates a given physiological function following thrombin activation. PMID- 11274184 TI - Identification of activating transcription factor 4 (ATF4) as an Nrf2-interacting protein. Implication for heme oxygenase-1 gene regulation. AB - Nrf2 regulates expression of genes encoding enzymes with antioxidant (e.g. heme oxygenase-1 (HO-1)) or xenobiotic detoxification (e.g. NAD(P)H:quinone oxidoreductase, glutathione S-transferase) functions via the stress- or antioxidant-response elements (StRE/ARE). Nrf2 heterodimerizes with small Maf proteins, but the role of such dimers in gene induction is controversial, and other partners may exist. By using the yeast two-hybrid assay, we identified activating transcription factor (ATF) 4 as a potential Nrf2-interacting protein. Association between Nrf2 and ATF4 in mammalian cells was confirmed by co immunoprecipitation and mammalian two-hybrid assays. Furthermore, Nrf2.ATF4 dimers bound to an StRE sequence from the ho-1 gene. CdCl(2), a potent inducer of HO-1, increased expression of ATF4 in mouse hepatoma cells, and detectable induction of ATF4 protein preceded that of HO-1 (30 min versus 2 h). A dominant negative mutant of ATF4 inhibited basal and CdCl(2)-stimulated expression of a StRE-dependent/luciferase fusion construct (pE1-luc) in hepatoma cells but only basal expression in mammary epithelial MCF-7 cells. A dominant mutant of Nrf2 was equally inhibitory in both cell types in the presence or absence of CdCl(2). These results indicate that ATF4 regulates basal and CdCl(2)-induced expression of the ho-1 gene in a cell-specific manner and possibly in a complex with Nrf2. PMID- 11274186 TI - Pivotal role of the P1 N-terminal domain in the assembly of the mammalian ribosomal stalk and in the proteosynthetic activity. AB - In the 60 S ribosomal subunit, the lateral stalk made of the P-proteins plays a major role in translation. It contains P0, an insoluble protein anchoring P1 and P2 to the ribosome. Here, rat recombinant P0 was overproduced in inclusion bodies and solubilized in complex with the other P-proteins. This method of solubilization appeared suitable to show protein complexes and revealed that P1, but not P2, interacted with P0. Furthermore, the use of truncated mutants of P1 and P2 indicated that residues 1-63 in P1 connected P0 to residues 1-65 in P2. Additional experiments resulted in the conclusion that P1 and P2 bound one another, either connected with P0 or free, as found in the cytoplasm. Accordingly, a model of association for the P-proteins in the stalk is proposed. Recombinant P0 in complex with phosphorylated P2 and either P1 or its (1-63) domain efficiently restored the proteosynthetic activity of 60 S subunits deprived of native P-proteins. Therefore, refolded P0 was functional and residues 1-63 only in P1 were essential. Furthermore, our results emphasize that the refolding principle used here is worth considering for solubilizing other insoluble proteins. PMID- 11274185 TI - The bifunctional Entamoeba histolytica alcohol dehydrogenase 2 (EhADH2) protein is necessary for amebic growth and survival and requires an intact C-terminal domain for both alcohol dahydrogenase and acetaldehyde dehydrogenase activity. AB - The intestinal protozoan pathogen Entamoeba histolytica lacks mitochondria and derives energy from the fermentation of glucose to ethanol with pyruvate, acetyl enzyme Co-A, and acetaldehyde as intermediates. A key enzyme in this pathway may be the 97-kDa bifunctional E. histolytica alcohol dehydrogenase 2 (EhADH2), which possesses both alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase activity (ALDH). EhADH2 appears to be a fusion protein, with separate N-terminal ALDH and C-terminal ADH domains. Here, we demonstrate that EhADH2 expression is required for E. histolytica growth and survival. We find that a mutant EhADH2 enzyme containing the C-terminal 453 amino acids of EhADH2 has ADH activity but lacks ALDH activity. However, a mutant consisting of the N-terminal half of EhADH2 possessed no ADH or ALDH activity. Alteration of a single histidine to arginine in the putative active site of the ADH domain eliminates both ADH and ALDH activity, and this mutant EhADH2 can serve as a dominant negative, eliminating both ADH and ALDH activity when co-expressed with wild-type EhADH2 in Escherichia coli. These data indicate that EhADH2 enzyme is required for E. histolytica growth and survival and that the C-terminal ADH domain of the enzyme functions as a separate entity. However, ALDH activity requires residues in both the N- and C-terminal halves of the molecule. PMID- 11274187 TI - Extranuclear lipid bodies, elicited by CCR3-mediated signaling pathways, are the sites of chemokine-enhanced leukotriene C4 production in eosinophils and basophils. AB - Eosinophils and basophils, when activated, become major sources of cysteinyl leukotrienes, eicosanoid mediators pertinent to allergic inflammation. We show that the C-C chemokines, eotaxin and RANTES (regulated upon activation normal T cell expressed and secreted), activate eosinophils and basophils for enhanced leukotriene C(4) (LTC(4)) generation by distinct signaling and compartmentalization mechanisms involving the induced formation of new cytoplasmic lipid body organelles. Chemokine-induced lipid body formation and enhanced LTC(4) release were both mediated by CCR3 receptor G protein-linked downstream signaling involving activation of phosphoinositide 3-kinase, extracellular signal-regulated kinases 1 and 2, and p38 mitogen-activated protein kinases. Chemokine-elicited lipid body numbers correlated with increased calcium ionophore-stimulated LTC(4) production; and as demonstrated by intracellular immunofluorescent localization of newly formed eicosanoid, lipid bodies were the predominant sites of LTC(4) synthesis in both chemokine-stimulated eosinophils and chemokine-primed and ionophore-activated eosinophils. Eotaxin and RANTES initiated signaling via phosphoinositide 3-kinase and mitogen-activated protein kinases both elicits the formation of lipid body domains and promotes LTC(4) formation at these specific extranuclear sites. PMID- 11274189 TI - Identification and characterization of a sac domain-containing phosphoinositide 5 phosphatase. AB - We have characterized a novel Sac domain-containing inositol phosphatase, hSac2. It was ubiquitously expressed but especially abundant in the brain, heart, skeletal muscle, and kidney. Unlike other Sac domain-containing proteins, hSac2 protein exhibited 5-phosphatase activity specific for phosphatidylinositol 4,5 bisphosphate and phosphatidylinositol 3,4,5-trisphosphate. This is the first time that the Sac domain has been reported to possess 5-phosphatase activity. Its 5 phosphatase activity for phosphatidylinositol 4,5-bisphosphate (K(m) = 14.3 microm) was comparable with those of Type II 5-phosphatases. These results imply that hSac2 functions as an inositol polyphosphate 5-phosphatase. PMID- 11274188 TI - Plasma membrane Ca2+-atpase isoforms 2b and 4b interact promiscuously and selectively with members of the membrane-associated guanylate kinase family of PDZ (PSD95/Dlg/ZO-1) domain-containing proteins. AB - Spatial and temporal regulation of intracellular Ca(2+) signaling depends on localized Ca(2+) microdomains containing the requisite molecular components for Ca(2+) influx, efflux, and signal transmission. Plasma membrane Ca(2+)-ATPase (PMCA) isoforms of the "b" splice type contain predicted PDZ (PSD95/Dlg/ZO-1) interaction domains. The COOH-terminal tail of PMCA2b isolated the membrane associated guanylate kinase (MAGUK) protein SAP97/hDlg as a binding partner in a yeast two-hybrid screen. The related MAGUKs SAP90/PSD95, PSD93/chapsyn-110, SAP97, and SAP102 all bound to the COOH-terminal tail of PMCA4b, whereas only the first three bound to the tail of PMCA2b. Coimmunoprecipitations confirmed the interaction selectivity between PMCA4b and SAP102 as opposed to the promiscuity of PMCA2b and 4b in interacting with other SAPs. Confocal immunofluorescence microscopy revealed the exclusive presence and colocalization of PMCA4b and SAP97 in the basolateral membrane of polarized Madin-Darby canine kidney epithelial cells. In hippocampal neurons, PMCA2b was abundant throughout the somatodendritic compartment and often extended into the neck and head of individual spines where it colocalized with SAP90/PSD95. These data show that PMCA "b" splice forms interact promiscuously but also with specificity with different members of the PSD95 family of SAPs. PMCA-SAP interactions may play a role in the recruitment and maintenance of the PMCA at specific membrane domains involved in local Ca(2+) regulation. PMID- 11274190 TI - The apolipoprotein E-dependent low density lipoprotein cholesteryl ester selective uptake pathway in murine adrenocortical cells involves chondroitin sulfate proteoglycans and an alpha 2-macroglobulin receptor. AB - Cells acquire lipoprotein cholesterol by receptor-mediated endocytosis and selective uptake pathways. In the latter case, lipoprotein cholesteryl ester (CE) is transferred to the plasma membrane without endocytosis and degradation of the lipoprotein particle. Previous studies with Y1/E/tet/2/3 murine adrenocortical cells that were engineered to express apolipoprotein (apo) E demonstrated that apoE expression enhances low density lipoprotein (LDL) CE uptake by both selective and endocytic pathways. The present experiments test the hypothesis that apoE-dependent LDL CE selective uptake is mediated by scavenger receptor, class B, type I (SR-BI). Surprisingly, SR-BI expression was not detected in the Y1/E/tet/2/3 clone of Y1 adrenocortical cells, indicating the presence of a distinct apoE-dependent pathway for LDL CE selective uptake. ApoE-dependent LDL CE selective uptake in Y1/E/tet/2/3 cells was inhibited by receptor-associated protein and by activated alpha(2)-macroglobulin (alpha(2)M), suggesting the participation of the LDL receptor-related protein/alpha(2)M receptor. Reagents that inhibited proteoglycan synthesis or removed cell surface chondroitin sulfate proteoglycan completely blocked apoE-dependent LDL CE selective uptake. None of these reagents inhibited SR-BI-mediated LDL CE selective uptake in the Y1-BS1 clone of Y1 cells in which LDL CE selective uptake is mediated by SR-BI. We conclude that LDL CE selective uptake in adrenocortical cells occurs via SR-BI independent and SR-BI-dependent pathways. The SR-BI-independent pathway is an apoE-dependent process that involves both chondroitin sulfate proteoglycans and an alpha(2)M receptor. PMID- 11274191 TI - Two phases of chromatin decondensation during dedifferentiation of plant cells: distinction between competence for cell fate switch and a commitment for S phase. AB - Cellular dedifferentiation is the major process underlying totipotency, regeneration, and formation of new stem cell lineages in multicellular organisms. In animals it is often associated with carcinogenesis. Here, we used tobacco protoplasts (plant cells devoid of cell wall) to study changes in chromatin structure in the course of dedifferentiation of mesophyll cells. Using flow cytometry and micrococcal nuclease analyses, we identified two phases of chromatin decondensation prior to entry of cells into S phase. The first phase takes place in the course of protoplast isolation, following treatment with cell wall degrading enzymes, whereas the second occurs only after protoplasts are induced with phytohormones to re-enter the cell cycle. In the absence of hormonal application, protoplasts undergo cycles of chromatin condensation/decondensation and die. The ubiquitin proteolytic system was found indispensable for protoplast progression into S phase, being required for the second but not the first phase of chromatin decondensation. The emerging model suggests that cellular dedifferentiation proceeds by two functionally distinct phases of chromatin decondensation: the first is a transitory phase that confers competence for cell fate switch, which is followed, under appropriate conditions, by a second proteasome-dependent phase representing a commitment for the mitotic cycle. These findings might have implications for a wide range of dedifferentiation-driven cellular processes in higher eukaryotes. PMID- 11274192 TI - Identification of a novel high affinity copper transport complex in the fission yeast Schizosaccharomyces pombe. AB - Copper is an essential nutrient that serves as a co-factor for enzymes involved in critical cellular processes including energy generation, peptide hormone maturation, oxidative stress protection, and iron homeostasis. Although genes have been identified from yeast and mammals encoding a homologous subunit of a plasma membrane high affinity copper transporter, the presence of additional subunits that function as part of a copper transport complex has not been reported. We observed that ctr4(+), a previously identified copper transport protein from the fission yeast Schizosaccharomyces pombe, fails to complement bakers' yeast cells defective in high affinity copper transport and fails to be targeted to the plasma membrane. However, selection for S. pombe genes, which, when co-expressed with Ctr4, confer high affinity copper transport to S. cerevisiae cells resulted in the identification of ctr5(+). Both Ctr4 and Ctr5 are integral membrane proteins, are co-regulated by copper levels and the copper sensing transcription factor Cuf1, physically associate in vivo, are interdependent for secretion to the plasma membrane, and are each essential for high affinity copper transport. These studies in S. pombe identify Ctr4 and Ctr5 as components of a novel eukaryotic heteromeric plasma membrane complex that is essential for high affinity copper transport. PMID- 11274193 TI - Mg2+ is not catalytically required in the intrinsic and kirromycin-stimulated GTPase action of Thermus thermophilus EF-Tu. AB - The influence of divalent metal ions on the intrinsic and kirromycin-stimulated GTPase activity in the absence of programmed ribosomes and on nucleotide binding affinity of elongation factor Tu (EF-Tu) from Thermus thermophilus prepared as the nucleotide- and Mg(2+)-free protein has been investigated. The intrinsic GTPase activity under single turnover conditions varied according to the series: Mn(2+) (0.069 min(-1)) > Mg(2+) (0.037 min(-1)) approximately no Me(2+) (0.034 min(-1)) > VO(2+) (0.014 min(-1)). The kirromycin-stimulated activity showed a parallel variation. Under multiple turnover conditions (GTP/EF-Tu ratio of 10:1), Mg(2+) retarded the rate of hydrolysis in comparison to that in the absence of divalent metal ions, an effect ascribed to kinetics of nucleotide exchange. In the absence of added divalent metal ions, GDP and GTP were bound with equal affinity (K(d) approximately 10(-7) m). In the presence of added divalent metal ions, GDP affinity increased by up to two orders of magnitude according to the series: no Me(2+) < VO(2+) < Mn(2+) approximately Mg(2+) whereas the binding affinity of GTP increased by one order of magnitude: no Me(2+) < Mg(2+) < VO(2+) < Mn(2+). Estimates of equilibrium (dissociation) binding constants for GDP and GTP by EF-Tu on the basis of Scatchard plot analysis, together with thermodynamic data for hydrolysis of triphosphate nucleotides (Phillips, R. C., George, P., and Rutman, R. J. (1969) J. Biol. Chem. 244, 3330-3342), showed that divalent metal ions stabilize the EF-Tu.Me(2+).GDP complex over the protein-free Me(2+).GDP complex in solution, with the effect greatest in the presence of Mg(2+) by approximately 10 kJ/mol. These combined results show that Mg(2+) is not a catalytically obligatory cofactor in intrinsic and kirromycin-stimulated GTPase action of EF-Tu in the absence of programmed ribosomes, which highlights the differential role of Mg(2+) in EF-Tu function. PMID- 11274194 TI - Identification of oligomerizing peptides. AB - The AraC DNA binding domain is inactive in a monomeric form but can activate transcription from the arabinose operon promoters upon its dimerization. We used this property to identify plasmids encoding peptide additions to the AraC DNA binding domain that could dimerize the domain. We generated a high diversity library of plasmids by inserting 90-base oligonucleotides of random sequence ahead of DNA coding for the AraC DNA binding domain in an expression vector, transforming, and selecting colonies containing functional oligomeric peptide AraC DNA binding domain chimeric proteins by their growth on minimal arabinose medium. Six of seven Ara(+) candidates were partially characterized, and one was purified. Equilibrium analytical centrifugation experiments showed that it dimerizes with a dissociation constant of approximately 2 micrometer. PMID- 11274195 TI - Prp-c and Prp-Sc at the fetal-maternal interface. AB - Scrapie is a naturally occurring prion (PrP) disease causing a fatal neurodegenerative disorder in sheep and goats. Previous studies suggest that scrapie is transmitted naturally through exposure to the scrapie agent in wasted placentas of infected ewes. This study determined the distribution and biochemical properties of PrP cellular (PrP-C) and the distribution of PrP scrapie (PrP-Sc) in reproductive, placental, and selected fetal tissues and fetal fluids in sheep. Glycosylated, N-terminally truncated, proteinase K-sensitive PrP C with apparent molecular masses of 23-37 kDa was present in reproductive, placental, and fetal tissues and fetal fluids. PrP-C was low or undetectable in intercotyledonary chorioallantois, amnion, urachus, amniotic fluid, and fetal urine. In pregnant ewes, cotyledonary chorioallantois, allantoic fluid, and caruncular endometrium contained higher levels of PrP-C than did intercaruncular endometrium, myometrium, oviduct, ovary, fetal bladder, or fetal kidney. Caruncular endometrial PrP-C was up-regulated during pregnancy. Despite the wide distribution of PrP-C in reproductive, placental, and selected fetal tissues and fetal fluid, PrP-Sc was detected only in caruncular endometrium and cotyledonary chorioallantois of pregnant scrapie-infected ewes. The embryo/fetus may not be exposed to scrapie in utero because it is separated physically from PrP-positive allantois and chorioallantois by PrP-negative amnion. PMID- 11274196 TI - Soluble E-selectin induces monocyte chemotaxis through Src family tyrosine kinases. AB - Cellular adhesion molecules such as E-selectin function to recruit leukocytes into the inflammatory lesions of diseases such as rheumatoid arthritis (RA) and atherosclerosis. Monocytes are the key components of the cellular infiltrates present in these disorders. We hypothesized that soluble E-selectin (sE-selectin) might mediate the chemotaxis of monocytes. In this report, we show that sE selectin induced normal human peripheral blood monocyte migration in the nanomolar range in a concentration-dependent manner. Neutralization studies using RA human joint synovial fluids and anti-E-selectin antibody showed a mean 31% reduction in RA synovial fluid-mediated monocyte chemotaxis (p < 0.05), indicating that sE-selectin is a major monocyte recruiter in RA. Next, we investigated the role of tyrosine phosphorylation pathways in sE-selectin-induced monocyte chemotaxis. Human peripheral blood monocytes stimulated with sE-selectin showed a time-dependent increase in the tyrosine phosphorylation of a broad range of cellular proteins, predominantly in the molecular size range of Src family kinases (50-60 kDa) and mitogen-activated protein kinases (MAPKs). Western blot analysis of Src family kinases showed a time-dependent increase in Src, Hck, and Lyn phosphorylation. The pretreatment of monocytes with the Src inhibitor AG1879: 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolol[3,4-d]pyrimidine (PP2) prior to stimulation with sE-selectin markedly inhibited Hck and Lyn phosphorylation, whereas the phosphorylation of Src was partially inhibited. In addition, the sE selectin stimulation of monocytes resulted in the increased phosphorylation of extracellular signal-related kinase (ERK1/2) and p38 MAPK. The pretreatment of monocytes with PP2 showed 89 and 83% inhibition of ERK1/2 and p38 MAPK phosphorylation, respectively. sE-selectin also showed a time-dependent activation of Ras kinase. Furthermore, the pretreatment of monocytes with PP2 completely inhibited sE-selectin-mediated monocyte chemotaxis. Taken together, our data demonstrate a novel function for sE-selectin as a monocyte chemotactic agent and suggest that sE-selectin might be mediating its biological functions through the Src-MAPK pathway. PMID- 11274197 TI - Increased insulin sensitivity in Gsalpha knockout mice. AB - The stimulatory guanine nucleotide-binding protein (G(s)) is required for hormone stimulated cAMP generation. Gnas, the gene encoding the G(s) alpha-subunit, is imprinted, and targeted disruption of this gene in mice leads to distinct phenotypes in heterozygotes depending on whether the maternal (m-/+) or paternal (+/p-) allele is mutated. Notably, m-/+ mice become obese, whereas +/p- mice are thinner than normal. In this study we show that despite these opposite changes in energy metabolism, both m-/+ and +/p- mice have greater sensitivity to insulin, with low to normal fasting glucose levels, low fasting insulin levels, improved glucose tolerance, and exaggerated hypoglycemic response to administered insulin. The combination of increased insulin sensitivity with obesity in m-/+ mice is unusual, because obesity is typically associated with insulin resistance. In skeletal muscles isolated from both m-/+ and +/p- mice, the basal rate of 2 deoxyglucose uptake was normal, whereas the rate of 2-deoxyglucose uptake in response to maximal insulin stimulation was significantly increased. The similar changes in muscle sensitivity to insulin in m-/+ and +/p- mice may reflect the fact that muscle G(s)alpha expression is reduced by approximately 50% in both groups of mice. GLUT4 expression is unaffected in muscles from +/p- mice. Increased responsiveness to insulin is therefore the result of altered insulin signaling and/or GLUT4 translocation. This is the first direct demonstration in a genetically altered in vivo model that G(s)-coupled pathways negatively regulate insulin signaling. PMID- 11274198 TI - Interaction of 11-cis-retinol dehydrogenase with the chromophore of retinal g protein-coupled receptor opsin. AB - Vertebrate opsins in both photoreceptors and the retinal pigment epithelium (RPE) have fundamental roles in the visual process. The visual pigments in photoreceptors are bound to 11-cis-retinal and are responsible for the initiation of visual excitation. Retinochrome-like opsins in the RPE are bound to all-trans retinal and play an important role in chromophore metabolism. The retinal G protein-coupled receptor (RGR) of the RPE and Muller cells is an abundant opsin that generates 11-cis-retinal by stereospecific photoisomerization of its bound all-trans-retinal chromophore. We have analyzed a 32-kDa protein (p32) that co purifies with bovine RGR from RPE microsomes. The co-purified p32 was identified by mass spectrometric analysis as 11-cis-retinol dehydrogenase (cRDH), and enzymatic assays have confirmed the isolation of an active cRDH. The co-purified cRDH showed marked substrate preference to 11-cis-retinal and preferred NADH rather than NADPH as the cofactor in reduction reactions. cRDH did not react with endogenous all-trans-retinal bound to RGR but reacted specifically with 11-cis retinal that was generated by photoisomerization after irradiation of RGR. The reduction of 11-cis-retinal to 11-cis-retinol by cRDH enhanced the net photoisomerization of all-trans-retinal bound to RGR. These results indicate that cRDH is involved in the processing of 11-cis-retinal after irradiation of RGR opsin and suggest that cRDH has a novel role in the visual cycle. PMID- 11274199 TI - Paracrine roles of NAD+ and cyclic ADP-ribose in increasing intracellular calcium and enhancing cell proliferation of 3T3 fibroblasts. AB - CD38 is a bifunctional ectoenzyme synthesizing from NAD(+) (ADP-ribosyl cyclase) and degrading (hydrolase) cyclic ADP-ribose (cADPR), a powerful universal calcium mobilizer from intracellular stores. Recently, hexameric connexin 43 (Cx43) hemichannels have been shown to release cytosolic NAD(+) from isolated murine fibroblasts (Bruzzone, S., Guida, L., Zocchi, E., Franco, L. and De Flora, A. (2001) FASEB J. 15, 10-12), making this dinucleotide available to the ectocellular active site of CD38. Here we investigated transwell co-cultures of CD38(+) (transfected) and CD38(-) 3T3 cells in order to establish the role of extracellular NAD(+) and cADPR on [Ca(2+)](i) levels and on proliferation of the CD38(-) target cells. CD38(+), but not CD38(-), feeder cells induced a [Ca(2+)](i) increase in the CD38(-) target cells which was comparable to that observed with extracellular cADPR alone and inhibitable by NAD(+)-glycohydrolase or by the cADPR antagonist 8-NH(2)-cADPR. Addition of recombinant ADP-ribosyl cyclase to the medium of CD38(-) feeders induced sustained [Ca(2+)](i) increases in CD38(-) target cells. Co-culture on CD38(+) feeders enhanced the proliferation of CD38(-) target cells over control values and significantly shortened the S phase of cell cycle. These results demonstrate a paracrine process based on Cx43 mediated release of NAD(+), its CD38-catalyzed conversion to extracellular cADPR, and influx of this nucleotide into responsive cells to increase [Ca(2+)](i) and stimulate cell proliferation. PMID- 11274200 TI - Identification of the apical membrane-targeting signal of the multidrug resistance-associated protein 2 (MRP2/MOAT). AB - The human canalicular multispecific organic anion transporter (cMOAT), known as the multidrug resistance-associated protein 2 (MRP2), is normally expressed in the liver and to a lesser extent in the kidney proximal tubules. In these tissues MRP2 specifically localizes to the apical membrane. The construction of MRP2 fused to the green fluorescent protein, and subsequent site-directed mutagenesis enabled the identification of a targeting signal in MRP2 that is responsible for its apical localization in polarized cells. The specific apical localization of MRP2 is due to a C-terminal tail that is not present in the basolaterally targeted MRP1. Deletion of three amino acids from the C-terminal of MRP2 (DeltaMRP2) causes the protein to be localized predominantly in the basolateral membrane in polarized Madin-Darby canine kidney cells. Interestingly, MRP2 expressed in a mouse leukemia cell line (L1210 cells) predominantly accumulates intracellularly with minimal cell membrane localization. In contrast, DeltaMRP2 was shown to predominantly localize in the cell membrane in L1210 cells. Increased transport of 2,4-dinitrophenyl glutathione from L1210 cells expressing DeltaMRP2 showed that the re-targeted protein retains its normal function. PMID- 11274201 TI - Development of glucose-induced insulin resistance in muscle requires protein synthesis. AB - Muscles and fat cells develop insulin resistance when exposed to high concentrations of glucose and insulin. We used an isolated muscle preparation incubated with high levels of glucose and insulin to further evaluate how glucose induced insulin resistance (GIIR) is mediated. Incubation with 2 milliunits/ml insulin and 36 mm glucose for 5 h resulted in an approximately 50% decrease in insulin-stimulated muscle glucose transport. The decrease in insulin responsiveness of glucose transport induced by glucose was not due to impaired insulin signaling, as insulin-stimulated phosphatidylinositol 3-kinase activity and protein kinase B phosphorylation were not reduced. It has been hypothesized that entry of glucose into the hexosamine biosynthetic pathway with accumulation of UDP-N-acetylhexosamines (UDP-HexNAcs) mediates GIIR. However, inhibition of the rate-limiting enzyme GFAT (glutamine:fructose-6-phosphate amidotransferase) did not protect against GIIR despite a marked reduction of UDP-HexNAcs. The mRNA synthesis inhibitor actinomycin D and the protein synthesis inhibitor cycloheximide both completely protected against GIIR despite the massive increases in UDP-HexNAcs and glycogen that resulted from increased glucose entry. Activation of AMP-activated protein kinase also protected against GIIR. These results provide evidence that GIIR can occur in muscle without increased accumulation of hexosamine pathway end products, that neither high glycogen concentration nor impaired insulin signaling is responsible for GIIR, and that synthesis of a protein with a short half-life mediates GIIR. They also suggest that dephosphorylation of a transcription factor may be involved in the induction of GIIR. PMID- 11274202 TI - Calmodulin binding and inhibition of cardiac muscle calcium release channel (ryanodine receptor). AB - Metabolically (35)S-labeled calmodulin (CaM) was used to determine the CaM binding properties of the cardiac ryanodine receptor (RyR2) and to identify potential channel domains for CaM binding. In addition, regulation of RyR2 by CaM was assessed in [(3)H]ryanodine binding and single-channel measurements. Cardiac sarcoplasmic reticulum vesicles bound approximately four CaM molecules per RyR2 tetramer in the absence of Ca(2+); in the presence of 100 microm Ca(2+), the vesicles bound 7.5 CaM molecules per tetramer. Purified RyR2 bound approximately four [(35)S]CaM molecules per RyR tetramer, both in the presence and absence of Ca(2+). At least four CaM binding domains were identified in [(35)S]CaM overlays of fusion proteins spanning the full-length RyR2. The affinity (but not the stoichiometry) of CaM binding was altered by redox state as controlled by the presence of either GSH or GSSG. Inhibition of RyR2 activity by CaM was influenced by Ca(2+) concentration, redox state, and other channel modulators. Parallel experiments with the skeletal muscle isoform showed major differences in the CaM binding properties and regulation by CaM of the skeletal and cardiac ryanodine receptors. PMID- 11274203 TI - NatC Nalpha-terminal acetyltransferase of yeast contains three subunits, Mak3p, Mak10p, and Mak31p. AB - The yeast Saccharomyces cerevisiae contains three types of N(alpha)-terminal acetyltransferases, NatA, NatB, and NatC, with each having a different catalytic subunit, Ard1p, Nat3p, and Mak3p, respectively, and each acetylating different sets of proteins with different N(alpha)-terminal regions. We show that the NatC N(alpha)-terminal acetyltransferases contains Mak10p and Mak31p subunits, in addition to Mak3p, and that all three subunits are associated with each other to form the active complex. Genetic deletion of any one of the three subunits results in identical abnormal phenotypes, including the lack of acetylation of a NatC substrate in vivo, diminished growth at 37 degrees C on media containing nonfermentable carbon sources, and the lack of maintenance or assembly of the L-A dsRNA viral particle. PMID- 11274204 TI - Characterization of the Net1 cell cycle-dependent regulator of the Cdc14 phosphatase from budding yeast. AB - In the budding yeast Saccharomyces cerevisiae, the multifunctional protein Net1 is implicated in regulating the cell cycle function of the Cdc14 protein phosphatase. Genetic and cell biological data suggest that during interphase and early mitosis Net1 holds Cdc14 within the nucleolus where its activity is suppressed. Upon its transient release from Net1 at late anaphase, active Cdc14 promotes exit from mitosis by dephosphorylating targets in the nucleus and cytoplasm. In this paper we present evidence supporting the proposed role of Net1 in regulating Cdc14 and exit from mitosis. We show that the NH(2)-terminal fragment Net1(1-600) directly binds Cdc14 in vitro and is a highly specific competitive inhibitor of its activity (K(i) = 3 nm) with five different substrates including the physiologic targets Swi5 and Sic1. An analysis of truncation mutants indicates that the Cdc14 binding site is located within a segment of Net1 containing residues 1-341. We propose that Net1 inhibits by occluding the active site of Cdc14 because it acts as a competitive inhibitor, binds to a site located within the catalytic domain (residues 1-374), binds with reduced affinity to a Cdc14 C283S mutant in which an active site Cys is replaced, and is displaced by tungstate, a transition state analog known to bind in the catalytic site of protein-tyrosine phosphatases. PMID- 11274205 TI - Interdependence of cdk2 activation and interleukin-2Ralpha accumulation in T cells. AB - We have shown previously that serum promotes T cell proliferation by acting with T cell receptor (TCR) agonists to efficiently down-regulate p27(Kip1) and activate cdk2-containing complexes. In the studies described here, the effect of serum on the expression of the alpha subunit of the interleukin-2 receptor (IL 2Ralpha) was examined. We found that serum was required for maximal and sustained IL-2Ralpha protein expression and consequent IL-2 signaling in TCR-activated splenocytes. Serum had no effect on IL-2Ralpha mRNA levels and thus modulates IL 2Ralpha expression post-transcriptionally. Unlike wild-type splenocytes, splenocytes exhibiting serum-independent cdk2 activation due to loss of p27(Kip1) efficiently expressed IL-2Ralpha in serum-deficient medium. Conversely, serum did not promote IL-2Ralpha accumulation in conditions in which cdk2 activity was blocked. These findings demonstrate that cdk2 activation is necessary and sufficient for IL-2Ralpha accumulation in TCR-stimulated splenocytes. On the other hand, IL-2 signaling was required (at least in part) for cdk2 activation in these cells. Thus, cdk2 activation, IL-2Ralpha expression, and IL-2 signaling are interdependent events, and we suggest that this feed-forward regulatory loop plays a key role in T cell mitogenesis. PMID- 11274206 TI - Formation of a stable heterodimer between Smad2 and Smad4. AB - Smad proteins mediate transforming growth factor beta signaling from the cell membrane to the nucleus. Upon phosphorylation by the activated receptor kinases, the receptor-regulated Smad, such as Smad2, forms a heterocomplex with the co mediator Smad, Smad4. This heterocomplex is then translocated into the nucleus, where it associates with other transcription factors and regulates expression of ligand-responsive genes. The stoichiometry between receptor-regulated Smad and co mediator Smad is important for understanding the molecular mechanisms of the signaling process. Using purified recombinant proteins, we demonstrate that Smad2 and Smad4 form a stable heterodimer and that the Smad4 activation domain is important for the formation of this complex. Many tumor-derived missense mutations disrupt the formation of this heterocomplex in in vitro interaction assays. Mapping these mutations onto the structures of Smad4 and Smad2 identifies a symmetric interface between these two Smad proteins. Importantly, two previous models on the formation of a heterocomplex are incompatible with our observations and other reported evidence. PMID- 11274207 TI - Alzheimer's disease amyloid-beta binds copper and zinc to generate an allosterically ordered membrane-penetrating structure containing superoxide dismutase-like subunits. AB - Amyloid beta peptide (Abeta) is the major constituent of extracellular plaques and perivascular amyloid deposits, the pathognomonic neuropathological lesions of Alzheimer's disease. Cu(2+) and Zn(2+) bind Abeta, inducing aggregation and giving rise to reactive oxygen species. These reactions may play a deleterious role in the disease state, because high concentrations of iron, copper, and zinc have been located in amyloid in diseased brains. Here we show that coordination of metal ions to Abeta is the same in both aqueous solution and lipid environments, with His(6), His(13), and His(14) all involved. At Cu(2+)/peptide molar ratios >0.3, Abeta coordinated a second Cu(2+) atom in a highly cooperative manner. This effect was abolished if the histidine residues were methylated at N(epsilon)2, indicating the presence of bridging histidine residues, as found in the active site of superoxide dismutase. Addition of Cu(2+) or Zn(2+) to Abeta in a negatively charged lipid environment caused a conformational change from beta sheet to alpha-helix, accompanied by peptide oligomerization and membrane penetration. These results suggest that metal binding to Abeta generated an allosterically ordered membrane-penetrating oligomer linked by superoxide dismutase-like bridging histidine residues. PMID- 11274208 TI - The carboxyl terminus of type VII collagen mediates antiparallel dimer formation and constitutes a new antigenic epitope for epidermolysis Bullosa acquisita autoantibodies. AB - Type VII collagen, the major component of anchoring fibrils, consists of a central collagenous triple-helical domain flanked by two noncollagenous domains, NC1 and NC2. The NC2 domain has been implicated in catalyzing the antiparallel dimer formation of type VII procollagen. In this study, we produced the entire 161 amino acids of the NC2 domain plus 186 amino acids of adjacent collagenous domain (NC2/COL) and purified large quantities of the recombinant NC2/COL protein. Recombinant NC2/COL readily formed disulfide-bonded hexamers, each representing one antiparallel dimer of collagen VII. Removal of the collagenous helical domain from NC2/COL by collagenase digestion abolished the antiparallel dimer formation. Using site-directed mutagenesis, we found that mutation of either cysteine 2802 or cysteine 2804 alone within the NC2 domain blocked antiparallel dimer formation. In contrast, a single cysteine mutation, 2634, within the collagenous helical domain had no effect. A generated methionine to lysine substitution, M2798K, that is associated with recessive dystrophic epidermolysis bullosa, was unable to form antiparallel dimers. Furthermore, autoantibodies from epidermolysis bullosa acquisita patients also reacted with NC2/COL. We conclude that NC2 and its adjacent collagenous segment mediate antiparallel dimer formation of collagen VII. Epidermolysis bullosa acquisita autoantibodies bound to this domain may destabilize anchoring fibrils by interfering with antiparallel dimer assembly leading to epidermal-dermal disadherence. PMID- 11274209 TI - NF-kappa B activation in tumor necrosis factor alpha-stimulated neutrophils is mediated by protein kinase Cdelta. Correlation to nuclear Ikappa Balpha. AB - The transcription factor NF-kappaB is critical for the expression of multiple genes involved in inflammatory responses and apoptosis. However, the signal transduction pathways regulating NF-kappaB activation in human neutrophils in response to stimulation with tumor necrosis factor-alpha (TNFalpha) are undefined. Since recent studies implicated activation of NF-kappaB as well as protein kinase C-delta (PKCdelta) in neutrophil apoptosis, we investigated involvement of PKCdelta in the activation of NF-kappaB in TNFalpha-stimulated neutrophils. Specific inhibition of PKCdelta by rottlerin prevented IkappaBalpha degradation and NF-kappaB activation in TNFalpha-stimulated neutrophils. This regulation of NF-kappaB activation by PKCdelta was specific only for TNFalpha signaling, since lipopolysaccharide- or interleukin-1beta-induced NF-kappaB activation and IkappaBalpha degradation were not inhibited by rottlerin. In addition, we show that in human neutrophils, but not monocytes, IkappaBalpha localizes in significant amounts in the nucleus of unstimulated cells, and the amount of IkappaBalpha in the nucleus, as well as in the cytoplasm, correlates with the NF-kappaB DNA binding. These results suggest that in human neutrophils, the presence of IkappaBalpha in the nucleus may function as a safeguard against initiation of NF-kappaB dependent transcription of pro-inflammatory and anti apoptotic genes, and represents a distinct and novel mechanism of NF-kappaB regulation. PMID- 11274211 TI - Role of retinoid receptor coactivator pockets in cofactor recruitment and transcriptional regulation. AB - The nuclear receptor for retinoic acid (RAR) forms a heterodimeric complex with the retinoid X receptor (RXR). This RXR/RAR heterodimer binds to the promoter of retinoic acid target genes and recruits coactivators and corepressors to regulate gene expression. Currently, the relative role of each receptor monomer in regulating coactivator and corepressor recruitment remains unclear. Here we show that the receptor-associated coactivator 3 (RAC3) uses two separate LXXLL motifs to bind RAR and RXR. The mutation of the coactivator-binding pockets of RAR and RXR abolishes RAC3 binding. Although the coactivator pocket of RXR is essential for the function of the RXR homodimer, it has a minor role for the recruitment of RAC3 and trans-activation by the RXR/RAR heterodimer. Consistently, deletion of the activation helix of RXR enhances binding of RAC3 to the heterodimer, and mutation of the coactivator pocket of RXR had little effect on RXR/RAR activity. In contrast, the coactivator pocket and the activation helix of RAR are absolutely required. We also show that different residues of the RAR coactivator pocket are used differently for interactions with the corepressor silencing mediator for retinoid and thyroid hormone receptor (SMRT) and coactivator. These results indicate a differential role for each retinoid receptor to the overall binding of cofactors and regulation of transcription by the retinoid receptor heterodimer. PMID- 11274212 TI - Revisiting the specificity of Mamestra brassicae and Antheraea polyphemus pheromone-binding proteins with a fluorescence binding assay. AB - Pheromone-binding proteins (PBPs), located in the sensillum lymph of pheromone responsive antennal hairs, are thought to transport the hydrophobic pheromones to the chemosensory membranes of olfactory neurons. It is currently unclear what role PBPs may play in the recognition and discrimination of species-specific pheromones. We have investigated the binding properties and specificity of PBPs from Mamestra brassicae (MbraPBP1), Antheraea polyphemus (ApolPBP1), Bombyx mori (BmorPBP), and a hexa-mutant of MbraPBP1 (Mbra1-M6), mutated at residues of the internal cavity to mimic that of BmorPBP, using the fluorescence probe 1 aminoanthracene (AMA). AMA binds to MbraPBP1 and ApolPBP1, however, no binding was observed with either BmorPBP or Mbra1-M6. The latter result indicates that relatively limited modifications to the PBP cavity actually interfere with AMA binding, suggesting that AMA binds in the internal cavity. Several pheromones are able to displace AMA from the MbraPBP1- and ApolPBP1-binding sites, without, however, any evidence of specificity for their physiologically relevant pheromones. Moreover, some fatty acids are also able to compete with AMA binding. These findings bring into doubt the currently held belief that all PBPs are specifically tuned to distinct pheromonal compounds. PMID- 11274213 TI - Protein kinase G regulates potassium chloride cotransporter-4 [corrected] expression in primary cultures of rat vascular smooth muscle cells. AB - K-Cl cotransport (KCC) is activated by nitric oxide donors and appears to be regulated by the cGMP signaling pathway. Expression of KCC mRNAs (KCC1-KCC4) in rat vascular smooth muscle cells (VSMCs) is unknown. We have reported the presence of KCC1 and KCC3 mRNAs in primary cultures of VSMCs by specific reverse transcription-polymerase chain reaction. KCC2 mRNA appeared at extremely low levels. KCC4 mRNA was undetectable. Semiquantitative reverse transcription polymerase chain reaction revealed a 2:1 KCC1/KCC3 mRNA ratio in VSMCs. Depletion of protein kinase G (PKG)-1 from VSMCs did not change KCC3 mRNA expression. Analogous results were obtained with PKG-1-catalytic domain- and vector only transfected VSMCs lacking endogenous PKG, suggesting no involvement of PKG-1 in the maintenance of basal KCC3 mRNA expression. However, 8-bromo-cGMP, a PKG stimulator, acutely increased KCC3 mRNA expression in a concentration- and time dependent fashion; this effect was blocked by the PKG inhibitor KT5823 but not by actinomycin D. These findings show that VSMCs express mainly two mRNA isoforms, KCC1 and KCC3, and suggest that PKG participates post-transcriptionally in the acute KCC3 mRNA regulation. The role of KCC3 on cell volume and electrolyte homeostasis in response to PKG modulators remains to be determined. PMID- 11274214 TI - Distinct kinetics of carnitine palmitoyltransferase i in contact sites and outer membranes of rat liver mitochondria. AB - Carnitine palmitoyltransferase I (CPT I) of rat liver mitochondria is an integral, polytopic protein of the outer membrane that is enriched at contact sites. As CPT I kinetics are highly dependent on its membrane environment, we have measured the kinetic parameters of CPT I present in rat liver submitochondrial membrane fractions enriched in either outer membrane or contact sites. The K(m) for palmitoyl-CoA was 2.4-fold higher for CPT I in outer membranes than that for the enzyme in contact sites. In addition, whereas in contact sites malonyl-CoA behaved as a competitive inhibitor of CPT I with respect to palmitoyl-CoA, in outer membranes malonyl-CoA inhibition was non competitive. As a result of the combination of these changes, the IC(50) for malonyl-CoA was severalfold higher for CPT I in contact sites than for the enzyme in bulk outer membrane. The K(i) for malonyl-CoA, the K(m) for carnitine, and the catalytic constant of the enzyme were all unaffected. It is concluded that the different membrane environments in outer membranes and contact sites result in an altered conformation of L-CPT I that specifically affects the long-chain acyl-CoA binding site. The accompanying changes in the kinetics of the enzyme provide an additional potent mechanism for the regulation of L-CPT I activity. PMID- 11274215 TI - Inhibitory effect of selenite on invasion of HT1080 tumor cells. AB - Selenium, an essential biological trace element, has been shown to reduce and prevent the incidence of cancer. Our previous studies have shown that selenite is involved in the chemoprevention of cancer and induction of apoptosis of cancer cells. In this study, we demonstrate that selenite also inhibits the invasion of tumor cells. Cancer cell invasion requires coordinated processes, such as changes in cell-cell and cell-matrix adhesion, degradation of the extracellular matrix, and cell migration. We found that selenite inhibited invasion of HT1080 human fibrosarcoma cells. Adhesion of HT1080 cells to the collagen matrix was also inhibited by treatment with selenite, but cell-cell interaction and cell motility were not affected by selenite. Moreover, selenite reduced expression of matrix metalloproteinase-2 and -9 and urokinase-type plasminogen activator, which are involved in matrix degradation, but increased a tissue inhibitor of metalloproteinase-1. This inhibitory effect of selenite on the protease expressions was mediated by the suppression of transcription factors, NF-kappaB and AP-1. However, selenate showed no remarkable effect on all the steps of cancer cell invasion. PMID- 11274216 TI - Differential activation of protein kinase B and p70(S6)K by glucose and insulin like growth factor 1 in pancreatic beta-cells (INS-1). AB - It has been shown that IGF-1-induced pancreatic beta-cell proliferation is glucose-dependent; however, the mechanisms responsible for this glucose dependence are not known. Adenoviral mediated expression of constitutively active phosphatidylinositol 3-kinase (PI3K) in the pancreatic beta-cells, INS-1, suggested that PI3K was not necessary for glucose-induced beta-cell proliferation but was required for IGF-1-induced mitogenesis. Examination of the signaling components downstream of PI3K, 3-phosphoinositide-dependent kinase 1, protein kinase B (PKB), glycogen synthase kinase-3, and p70-kDa-S6-kinase (p70(S6K)), suggested that a major part of glucose-dependent beta-cell proliferation requires activation of mammalian target of rapamycin/p70(S6K), independent of phosphoinositide-dependent kinase 1/PKB activation. Adenoviral expression of the kinase-dead form of PKB in INS-1 cells decreased IGF-1-induced beta-cell proliferation. However, a surprisingly similar decrease was also observed in adenoviral wild type and constitutively active PKB-infected cells. Upon analysis of extracellular signal-regulated protein kinase 1 and 2 (ERK1/ERK2), an increase in ERK1/ERK2 phosphorylation activation by glucose and IGF-1 was observed in kinase-dead PKB-infected cells, but this phosphorylation activation was inhibited in the constitutively active PKB-infected cells. Hence, there is a requirement for the activation of both ERK1/ERK2 and mammalian target of rapamycin/p70(S6K) signal transduction pathways for a full commitment to glucose-induced pancreatic beta-cell mitogenesis. However, for IGF-1-induced activation, these pathways must be carefully balanced, because chronic activation of one (PI3K/PKB) can lead to dampening of the other (ERK1/2), reducing the mitogenic response. PMID- 11274217 TI - Phospholipase C-gamma mediates the hydrolysis of phosphatidylinositol, but not of phosphatidylinositol 4,5-bisphoshate, in carbamylcholine-stimulated islets of langerhans. AB - In pancreatic islets the activation of phospholipase C (PLC) by the muscarinic receptor agonist carbamyolcholine (carbachol) results in the hydrolysis of both phosphatidylinositol 4,5-bisphosphate (PtdInsP(2)) and phosphatidylinositol (PtdIns). Here we tested the hypothesis that PtdIns hydrolysis is mediated by PLCgamma1, which is known to be regulated by activation of tyrosine kinases and PtdIns 3-kinase. PtdIns breakdown was more sensitive than that of PtdInsP(2) to the tyrosine kinase inhibitor, genistein. Conversely, the tyrosine phosphatase inhibitor, vanadate, alone promoted PtdIns hydrolysis and acted non-additively with carbachol. Vanadate did not stimulate PtdInsP(2) breakdown. Carbachol also stimulated a rapid (maximal at 1-2 min) tyrosine phosphorylation of several islet proteins, although not of PLCgamma1 itself. Two structurally unrelated inhibitors of PtdIns 3-kinase, wortmannin and LY294002, more effectively attenuated the hyrolysis of PtdIns compared with PtdInsP(2). Adenovirally mediated overexpression of PLCgamma1 significantly increased carbachol-stimulated PtdIns hydrolysis without affecting that of PtdInsP(2). Conversely overexpression of PLCbeta1 up-regulated the PtdInsP(2), but not PtdIns, response. These results indicate that the hydrolysis of PtdIns and PtdInsP(2) are independently regulated in pancreatic islets and that PLCgamma1 selectively mediates the breakdown of PtdIns. The activation mechanism of PLCgamma involves tyrosine phosphorylation (but not of PLCgamma directly) and PtdIns 3-kinase. Our findings point to a novel bifurcation of signaling pathways downstream of muscarinic receptors and suggest that hydrolysis of PtdIns and PtdInsP(2) might serve different physiological ends. PMID- 11274218 TI - Split Na+-Ca2+ exchangers. Implications for function and expression. AB - The Na(+)-Ca(2+) exchanger has nine transmembrane segments, with a large cytoplasmic loop between the fifth and sixth transmembrane segments. The protein was split within the cytoplasmic loop into two domains consisting of the first five transmembrane segments and the last four transmembrane segments, respectively. The two domains were either expressed individually or coexpressed. Each of the two domains with different lengths of the cytoplasmic loop was fused to green fluorescent protein. We show that coexpression of both domains is required for proper membrane targeting and for expression of functional exchange activity. Fusion to green fluorescent protein does not alter biophysical properties of the exchange process. In addition, truncation of a large portion of the cytoplasmic loop does not alter important properties of the exchanger such as Na(+)-dependent inactivation, activation by chymotrypsin, or exchanger inhibitory peptide (XIP) sensitivity. PMID- 11274219 TI - Binding of regulator of G protein signaling (RGS) proteins to phospholipid bilayers. Contribution of location and/or orientation to Gtpase-activating protein activity. AB - Regulator of G protein signaling (RGS) proteins must bind membranes in an orientation that permits the protein-protein interactions necessary for regulatory activity. RGS4 binds to phospholipid surfaces in a slow, multistep process that leads to maximal GTPase-activating protein (GAP) activity. When RGS4 is added to phospholipid vesicles that contain m2 or m1 muscarinic receptor and G(i), G(z), or G(q), GAP activity increases approximately 3-fold over 4 h at 30 degrees C and more slowly at 20 degrees C. This increase in GAP activity is preceded by several other events that suggest that, after binding, optimal interaction with G protein and receptor requires reorientation of RGS4 on the membrane surface, a conformational change, or both. Binding of RGS4 is initially reversible but becomes irreversible within 5 min. Onset of irreversibility parallels initial quenching of tryptophan fluorescence (t(12) approximately 30 s). Further quenching occurs after binding has become irreversible (t(12) approximately 6 min) but is complete well before maximal GAP activity is attained. These processes all appear to be energetically driven by the amphipathic N-terminal domain of RGS4 and are accelerated by palmitoylation of cysteine residues in this region. The RGS4 N-terminal domain confers similar membrane binding behavior on the RGS domains of either RGS10 or RGSZ1. PMID- 11274220 TI - Molecular cloning and functional characterization of MCH2, a novel human MCH receptor. AB - Melanin-concentrating hormone (MCH) is involved in the regulation of feeding and energy homeostasis. Recently, a 353-amino acid splice variant form of the human orphan receptor SLC-1 () (hereafter referred to as MCH(1)) was identified as an MCH receptor. This report describes the cloning and functional characterization of a novel second human MCH receptor, which we designate MCH(2), initially identified in a genomic survey sequence as being homologous to MCH(1) receptors. Using this sequence, a full-length cDNA was generated with an open reading frame of 1023 base pairs, encoding a polypeptide of 340 amino acids, with 38% identity to MCH(1) and with many of the structural features conserved in G protein-coupled receptors. This newly discovered receptor belongs to class 1 (rhodopsin-like) of the G protein-coupled receptor superfamily. HEK293 cells transfected with MCH(2) receptors responded to nanomolar concentrations of MCH with an increase in intracellular Ca(2+) levels and increased cellular extrusion of protons. In addition, fluorescently labeled MCH bound with nanomolar affinity to these cells. The tissue localization of MCH(2) receptor mRNA, as determined by quantitative reverse transcription-polymerase chain reaction, was similar to that of MCH(1) in that both receptors are expressed predominantly in the brain. The discovery of a novel MCH receptor represents a new potential drug target and will allow the further elucidation of MCH-mediated responses. PMID- 11274221 TI - Src and Pyk2 mediate G-protein-coupled receptor activation of epidermal growth factor receptor (EGFR) but are not required for coupling to the mitogen-activated protein (MAP) kinase signaling cascade. AB - The epidermal growth factor receptor (EGFR) and the non-receptor protein tyrosine kinases Src and Pyk2 have been implicated in linking a variety of G-protein coupled receptors (GPCR) to the mitogen-activated protein (MAP) kinase signaling cascade. In this report we apply a genetic strategy using cells isolated from Src , Pyk2-, or EGFR-deficient mice to explore the roles played by these protein tyrosine kinases in GPCR-induced activation of EGFR, Pyk2, and MAP kinase. We show that Src kinases are critical for activation of Pyk2 in response to GPCR stimulation and that Pyk2 and Src are essential for GPCR-induced tyrosine phosphorylation of EGFR. By contrast, Pyk2, Src, and EGFR are dispensable for GPCR-induced activation of MAP kinase. Moreover, GPCR-induced MAP kinase activation is normal in fibroblasts deficient in both Src and Pyk2 (Src-/-Pyk2-/- cells) as well as in fibroblasts deficient in all three Src kinases expressed in these cells (Src-/-Yes-/-Fyn-/- cells). Finally, experiments are presented demonstrating that, upon stimulation of GPCR, activated Pyk2 forms a complex with Src, which in turn phosphorylates EGFR directly. These experiments reveal a role for Src kinases in Pyk2 activation and a role for Pyk2 and Src in tyrosine phosphorylation of EGFR following GPCR stimulation. In addition, EGFR, Src family kinases, and Pyk2 are not required for linking GPCRs with the MAP kinase signaling cascade. PMID- 11274222 TI - In vivo effects of uncoupling protein-3 gene disruption on mitochondrial energy metabolism. AB - To clarify the role of uncoupling protein-3 (UCP3) in skeletal muscle, we used NMR and isotopic labeling experiments to evaluate the effect of UCP3 knockout (UCP3KO) in mice on the regulation of energy metabolism in vivo. Whole body energy expenditure was determined from the turnover of doubly labeled body water. Coupling of mitochondrial oxidative phosphorylation in skeletal muscle was evaluated from measurements of rates of ATP synthesis (using (31)P NMR magnetization transfer experiments) and tricarboxylic acid (TCA) cycle flux (calculated from the time course of (13)C enrichment in C-4 and C-2 of glutamate during an infusion of [2-(13)C]acetate). At the whole body level, we observed no change in energy expenditure. However, at the cellular level, skeletal muscle UCP3KO increased the rate of ATP synthesis from P(i) more than 4-fold under fasting conditions (wild type, 2.2 +/- 0.6 versus knockout, 9.1 +/- 1.4 micromol/g of muscle/min, p < 0.001) with no change in TCA cycle flux rate (wild type, 0.74 +/- 0.04 versus knockout, 0.71 +/- 0.03 micromol/g of muscle/min). The increased efficiency of ATP production may account for the significant (p < 0.05) increase in the ratio of ATP to ADP in the muscle of UCP3KO mice (5.9 +/- 0.3) compared with controls (4.5 +/- 0.4). The data presented here provide the first evidence of uncoupling activity by UCP3 in skeletal muscle in vivo. PMID- 11274223 TI - In memoriam Ramzi S. Cotran: December 7, 1932 - October 23, 2000. PMID- 11274224 TI - Transcytosis of retinol-binding protein across renal proximal tubule cells after megalin (gp 330)-mediated endocytosis. AB - Plasma retinol-binding protein (RBP) combined with vitamin A (retinol) is partially filtered through the glomerulus and then absorbed by proximal tubule cells, leading to recycling of retinol to the circulation. Recently, it was shown that reabsorption of RBP-retinol complexes by proximal tubule cells is mediated by megalin (gp 330), an apical endocytic receptor. It was proposed that RBP is transported by megalin to lysosomes, where it is degraded, thus liberating retinol, which then combines with newly synthesized RBP to be secreted into the bloodstream. This study shows that passage of RBP through immortalized rat renal proximal tubule (IRPT) cells occurs by transcytosis after megalin-mediated endocytosis, which provides an alternative pathway for recycling of retinol. IRPT cells cultured as polarized monolayers with tight junctions were used on permeable filters in the upper chamber of dual-chambered devices, with megalin expression exclusively on the upper surface. After addition of RBP to the upper chamber and incubation at 37 degrees C, intact RBP was found in fluids that were collected from the lower chamber. In contrast, control substances (mannitol, lysozyme, albumin, and glutathione-S-transferase) were not appreciably transported across IRPT cells, indicating that passage of RBP was by transcytosis and not by paracellular leakage. Confocal microscopy analysis of IRPT cells after addition of RBP to the upper chamber revealed RBP-containing granules at the apical membrane, subapically, and also at basolateral membranes. When RBP was added to IRPT cells together with megalin competitors, the amount of transcytosed RBP was markedly reduced. We also found that some RBP was internalized and degraded by IRPT cells, but this process was not appreciably affected by megalin competitors, indicating that RBP endocytosed by megalin was not transported to lysosomes and degraded but rather transcytosed across IRPT cells. PMID- 11274225 TI - Induction of cyclooxygenase-2 in thick ascending limb cells by adrenalectomy. AB - Adrenalectomized (ADX) and sham-operated rats received either dexamethasone (DEX) or vehicle. Renal tissue was used for morphologic analysis, assessment of cyclooxygenase-2 (COX-2) protein expression and mRNA accumulation, and quantitation of COX-2 activity. In untreated or shamoperated rats, COX-2 protein was observed in a subset of tubular epithelial cells (<2%), which were located mainly in the cortex. All COX-2-positive cells also expressed Tamm-Horsfall glycoprotein, a highly selective marker for thick ascending limb (TAL) cells. After ADX, >30% of TAL cells expressed COX-2 in a manner consistent with recruitment of COX-2-positive TAL cells toward the medulla. Treatment of ADX rats with DEX reduced the number of COX-2-positive cells to that observed in sham operated or intact rats. COX-2 mRNA accumulation was increased by ADX and partially attenuated by treatment with DEX. Western blot analysis of cortical microsomes revealed a substantial increase in COX-2 expression in ADX rats, compared with ADX/DEX-treated, sham-operated, or intact rats. The increase in COX 2 protein expression was associated with a twofold increase in prostaglandin E(2) formation by cortical microsomes obtained from ADX rats, compared with sham operated rats. It is concluded that ADX induces expression of enzymatically active COX-2, such that expression occurs in the cortical TAL and proceeds in a defined pattern toward the outer medullary TAL. It is suggested that ADX induces expression of TAL cells that, in the basal state, do not express COX-2 protein. PMID- 11274226 TI - Temporary treatment of prepubescent rats with angiotensin inhibitors suppresses the development of hypertensive nephrosclerosis. AB - Hypertensive nephrosclerosis is a leading cause of end-stage renal disease; therefore, strategies to prevent the development of renal disease require close study. Here it is demonstrated that transient treatment of prepubescent rats with angiotensin inhibitors attenuated their susceptibility to the development of hypertensive nephrosclerosis after maturation. Stroke-prone spontaneously hypertensive Izumo strain rats were divided into four groups, treated with vehicle, the angiotensin-converting enzyme inhibitor (ACEI) delapril (40 mg/kg per d), the angiotensin receptor antagonist (AT1R-Ant) candesartan cilexetil (1 mg/kg per d), or the vasodilator hydralazine (25 mg/kg per d) from weaning to puberty (3 to 10 wk of age), and then monitored without treatment for 6 mo. BP in the ACEI- and AT1R-Ant-treated groups remained significantly decreased, compared with the untreated and hydralazine-treated groups. Moreover, marked proteinuria and nephrosclerosis developed in the untreated and hydralazine-treated groups at 30 wk but were suppressed in the ACEI- and AT1R-Ant-treated groups. Of interest, plasma renin activity, plasma angiotensin II concentrations, and renal renin mRNA levels were reduced by >50% in the ACEI- and AT1R-Ant-treated rats, suggesting that the treatments may have attenuated the development of nephrosclerosis by overcoming the susceptibility of stroke-prone spontaneously hypertensive rats to overactivation of the renin-angiotensin system. PMID- 11274227 TI - The membrane-associated guanylate kinase protein MAGI-1 binds megalin and is present in glomerular podocytes. AB - The transmembrane endocytic receptor glycoprotein 330/megalin (hereafter referred to as megalin) is localized to the apical membrane domain of epithelial cells, where it is involved in the uptake of proteins from extracellular sources. The cytoplasmic domain of megalin contains amino acid motifs that have the potential to bind to other proteins, which may influence its localization or function. The yeast two-hybrid system was used to search for proteins that bind to the cytoplasmic tail of megalin, and a protein fragment from a mouse embryonic cDNA library that contained a single PDZ domain was identified. This protein, which was named glycoprotein 330-associated protein (GASP), appears to be a truncated mouse counterpart of the human and rat proteins atrophin-1-interacting protein-1 and synaptic scaffolding molecule, respectively. The interaction of GASP with megalin is mediated by the PDZ domain of GASP binding to the DSDV motif found at the carboxyl-terminus of megalin. A mutant version of megalin that lacks the terminal valine is unable to bind to GASP, illustrating the PDZ domain-dependent interaction between these two proteins. A close homolog of GASP, i.e., membrane associated guanylate kinase with inverted orientation-1 (MAGI-1), is more ubiquitous in its tissue distribution (including kidney) and is also able to specifically bind to megalin via its fifth PDZ domain. Immunofluorescence studies of adult kidney revealed that MAGI-1 is expressed in the glomerulus of the kidney, in a manner that parallels the expression of the podocyte-specific protein glomerular epithelial protein 1. Western analysis of endogenous MAGI-1 from glomerular preparations suggests that it is associated with the cytoskeleton and seems to be expressed in a different form, compared with cell line-derived endogenous MAGI-1. The association of megalin with MAGI-1 may allow the assembly of a multiprotein complex, in which megalin may serve a nonendocytic function in glomerular podocytes. PMID- 11274228 TI - Acetylcholine increases the free intracellular calcium concentration in podocytes in intact rat glomeruli via muscarinic M(5) receptors. AB - The effects of acetylcholine (ACh) on the free intracellular calcium concentration ([Ca2+](i)) of microdissected glomeruli were investigated using fura-2 fluorescence digital imaging and two-photon confocal microscopy. ACh caused a concentration-dependent [Ca2+](i) increases with an initial peak followed by a sustained plateau, which was suppressed by reduced extracellular Ca2+ concentrations. The [Ca2+](i) plateau was not affected by the L-type Ca2+ channel blocker nicardipine, whereas gadolinium and lanthanum (both at 1 microM) blocked the plateau. Diphenylacetoxy-N:-methylpiperidine methiodide (100 nM), an M(3)/M(5) receptor antagonist, and pirenzepine (1 microM), an M(1) receptor antagonist, completely inhibited the effect of ACh. [Ca2+](i) measurements using two-photon excitation of fluo-3 and staining of the cells with calcein/acetoxymethyl ester, for observation of the capillary network together with the glomerular cells, showed that [Ca2+](i) was increased in single podocytes. Immunohistochemical studies did not demonstrate M(3) receptor expression in glomerular cells. M(1) receptors could be detected only in the parietal sheet of Bowman's capsule, whereas M(5) receptors were found only in podocytes. The data show that ACh increases [Ca2+](i) in podocytes of intact glomeruli, most likely via muscarinic M(5) receptors. PMID- 11274229 TI - Suppression of constitutive but not Il-1beta-inducible expression of monocyte chemoattractant protein-1 in mesangial cells by retinoic acids: intervention in the activator protein-1 pathway. AB - Retinoic acid regulates a wide range of biologic processes, including inflammation. This study investigated the effect of all-trans-retinoic acid (t RA) on the constitutive and cytokine-inducible expression of monocyte chemoattractant protein 1 (MCP-1) in rat mesangial cells. Serum-deprived mesangial cells exhibited substantial levels of MCP-1 mRNA, and the expression was markedly upregulated by interleukin-1beta (IL-1beta). Pretreatment with t-RA abrogated the constitutive mRNA expression but did not inhibit the IL-1beta inducible expression. The similar effects were observed by 9-cis-RA. The suppressive effect of t-RA required retinoic acid receptors. t-RA did not affect the stability of MCP-1 mRNA, indicating that its suppressive effect was at the transcriptional level. Experiments that used pharmacologic and genetic inhibitors showed that the IL-1beta-inducible MCP-1 expression was dependent on nuclear factor-kappaB (NF-kappaB) and independent of activator protein 1 (AP-1). In contrast, the constitutive expression of MCP-1 was dependent on both NF-kappaB and AP-1. t-RA substantially inhibited the constitutive activity of AP-1 but did not inhibit NF-kappaB activity in mesangial cells. These data suggested that (1) constitutive and IL-1beta-inducible expression of MCP-1 was differently regulated by AP-1 and NF-kappaB and (2) t-RA inhibited selectively the constitutive expression of MCP-1 via intervention in the AP-1 pathway. PMID- 11274230 TI - CD40 is expressed on human peritoneal mesothelial cells and upregulates the production of interleukin-15 and RANTES. AB - Limited data are available concerning the interaction between lymphocytes and human peritoneal mesothelial cells (HPMC) during peritonitis. CD40 is a member of the tumor necrosis factor (TNF) family of receptors whose ligand (CD154) is mainly expressed on the membrane of activated CD4-positive lymphocytes. CD154 CD40 cross-linking is a central event in antigen presentation, B-cell activation by T cells, and regulation of cytokine secretion from various types of cells. The goal of this study was to demonstrate in vitro the presence of CD40 on HPMC and to test its functionality in inducing interleukin-15 (IL-15) and RANTES. We assayed the levels of CD40 by reverse transcription-PCR and flow cytometry and IL 15 and RANTES by enzyme-linked immunosorbent assay. Genetically modified L cells that express elevated levels of CD154 (CD40L cells) were used to stimulate CD40. HPMC express CD40 mRNA and protein. After stimulation with interferon-gamma (IFNgamma, 5U/ml) or TNFalpha (1 ng/ml), there was a small increase in CD40 mRNA and protein levels; when both cytokines were applied, the increase in CD40 levels was more than threefold. CD40 ligation induced IL-15 production by HPMC and was additive to IFNgamma stimulation. CD40 ligation was strongly synergistic with IFNgamma in induction of RANTES (20-fold as compared with unstimulated HPMC), whereas neither ligation nor IFNgamma alone could induce RANTES. Pretreatment of HPMC with TNFalpha and IFNgamma increased the response to CD40 ligation in magnitudes that correlated with the elevation of CD40 levels induced by the pretreatment. To conclude, the presence of a functional CD40 on HPMC whose ligation induced IL-15 and RANTES production was detected. It is possible that this receptor acts as a major mediator of T-cell-regulated immune and inflammatory response during peritonitis. PMID- 11274231 TI - Thrombospondin-1 is the key activator of TGF-beta1 in human mesangial cells exposed to high glucose. AB - Elevated levels of transforming growth factor-beta1 (TGF-beta1) are synthesized by human mesangial cells that are cultured in medium that contains high concentrations of glucose and mediate increased synthesis of fibronectin (FN), plasminogen activator inhibitor-1 (PAI-1), and changes in the expression of other genes. TGF-beta1 is synthesized as a latent complex. Previous work indicated that high-glucose conditions also upregulate expression of thrombospondin-1 (TSP-1), a potential activator of latent TGF-beta1. With the use of the synthetic peptide GGWSHW, an inhibitor of the TSP-1 activation mechanism, endogenous TSP-1 is shown to be responsible for converting high levels of latent TGF-beta1 to bioactive growth factor over 3 wk of exposure of mesangial cells to 30 mM D-glucose. Peptide inhibition of TGF-beta1 activation by TSP-1 in high-glucose conditions completely suppressed increases in FN and PAI-1 expression. Treating mesangial cells maintained in high glucose with a TSP-1 antisense oligonucleotide reduced TSP-1 expression to levels found in 4 mM D-glucose cultures, prevented TGF-beta1 activation, and normalized expression of FN. PMID- 11274232 TI - Novel nonsense mutation in the Na+/HCO3- cotransporter gene (SLC4A4) in a patient with permanent isolated proximal renal tubular acidosis and bilateral glaucoma. AB - Permanent isolated proximal renal tubular acidosis (pRTA) with ocular abnormalities is a systemic disease involving short stature, isolated pRTA, mental retardation, and ocular abnormalities. Kidney Na+/HCO3- cotransporter (kNBC1) cDNA from peripheral lymphocytes from a patient with permanent isolated pRTA and bilateral glaucoma was screened, and a novel homozygous mutation, namely a cytosine-to-thymine transition at nucleotide 234, which resulted in the formation of a stop codon at codon 29, was identified. This homozygous mutation, Q29X, was identified in the unique 5'-end of the kNBC1 gene (SLC4A4) of the patient. Cosegregation of this Q29X mutation with the disease and heterozygosity in the parents of the affected patient were observed. The absence of this mutation in 156 alleles from 78 Japanese individuals indicates that this mutation is directly related to the disease and is not a common DNA sequence polymorphism. This nonsense mutation predicts a truncated kNBC1 protein that lacks the 1007 amino acids of the carboxyl-terminus, and the effect on kNBC1 cotransport activity is likely to be a loss of function. In contrast, the pancreatic Na+/HCO3 cotransporter of the patient is not likely to be affected by this nonsense mutation. These results have implications for understanding the role of kNBC1 in the pathophysiologic processes of pRTA associated with ocular abnormalities and mental retardation. PMID- 11274233 TI - Role of CFTR in autosomal recessive polycystic kidney disease. AB - An extensive body of in vitro data implicates epithelial chloride secretion, mediated through cystic fibrosis transmembrane conductance regulator (CFTR) protein, in generating or maintaining fluid filled cysts in MDCK cells and in human autosomal dominant polycystic kidney disease (ADPKD). In contrast, few studies have addressed the pathophysiology of fluid secretion in cyst formation and enlargement in autosomal recessive polycystic kidney disease (ARPKD). Murine models of targeted disruptions or deletions of specific genes have created opportunities to examine the role of individual gene products in normal development and/or disease pathophysiology. The creation of a murine model of CF, which lacks functional CFTR protein, provides the opportunity to determine whether CFTR activity is required for renal cyst formation in vivo. Therefore, this study sought to determine whether renal cyst formation could be prevented by genetic complementation of the BPK murine model of ARPKD with the CFTR knockout mouse. The results of this study reveal that in animals that are homozygous for the cystic gene (bpk), the lack of functional CFTR protein on the apical surface of cystic epithelium does not provide protection against cyst growth and subsequent decline in renal function. Double mutant mice (bpk -/-; cftr -/-) developed massively enlarged kidneys and died, on average, 7 d earlier than cystic, non-CF mice (bpk -/-; cftr +/+/-). This suggests fundamental differences in the mechanisms of transtubular fluid secretion in animal models of ARPKD compared with ADPKD. PMID- 11274234 TI - Klf6 is a zinc finger protein expressed in a cell-specific manner during kidney development. AB - Molecular mechanisms that are responsible for the development of the renal collecting duct system during embryogenesis are still poorly understood. A mouse cDNA encoding a zinc finger protein, called Klf6, which is a member of the Kruppel-like family of transcription factors, has been cloned. Northern blot analyses showed that Klf6 was already expressed in 11.5-d postconception mouse embryos and that its expression persisted after birth. They also disclosed that Klf6 had a restricted pattern of expression. In situ hybridization experiments using mouse embryos showed that during kidney development, Klf6 was expressed selectively in the Wolffian duct and in its derivatives. During mesonephros development, it was expressed in the Wolffian duct but not in the mesonephric mesenchyme. Thereafter, Klf6 was expressed in the ureteric bud and its branches and in the collecting ducts, whereas it was not expressed in tubular structures that derive from the metanephric mesenchyme. Glomeruli were not labeled during early stages of differentiation, and it is only at the capillary stage that a staining of the mesangial area was observed, which persisted after birth. This pattern of expression is strikingly similar to the one of GATA-3, which is another zinc finger protein. It suggests that Klf6 may play a role during kidney development and in particular during the development of the renal collecting duct system, possibly in association with GATA-3. PMID- 11274235 TI - TIMP-1 deficiency does not attenuate interstitial fibrosis in obstructive nephropathy. AB - Progressive renal disease as a result of renal fibrosis is caused in part by an impairment of the proteolytic machinery that normally regulates matrix turnover. The goal of the present study was to determine whether genetic deficiency of tissue inhibitor of metalloproteinases-1 (TIMP-1) could attenuate interstitial fibrosis caused by unilateral ureteral obstruction (UUO). Groups of wild-type (Timp-1) mice and TIMP-1-deficient (timp-1) mice were killed after 3 and 14 d of UUO or sham operation. Timp-1 mRNA levels were significantly increased 37- and 19 fold in the wild-type mice 3 and 14 d, respectively, after UUO operation. Matrix metalloproteinase-9 (MMP-9) activity fell in all UUO groups but remained significantly higher in the timp-1 group compared with the Timp-1 group. The degree of interstitial fibrosis (kidney collagen content and percentage of tubulointerstitial area stained with picrosirius red and collagen III) was significantly increased 14 d after UUO operation, but there was no difference between the Timp-1 and timp-1 groups. Many features of the fibrogenic response were similar between the Timp-1 and timp-1 groups, including the number of myofibroblasts and the induction of genes encoding procollagen III, fibronectin, and transforming growth factor-beta. After UUO operation, renal mRNA levels for Timp-3 and plasminogen activator inhibitor-1 were significantly higher in the TIMP-1-deficient mice. The results of this study show that elimination of TIMP-1 alone does not alter the severity of interstitial fibrosis. These findings may be due to compensation by other protease inhibitors such as TIMP-2, TIMP-3, and/or plasminogen activator inhibitor-1 or to the possibility that inhibition of intrinsic MMP activity does not constitute a profibrogenic event in the kidney. PMID- 11274236 TI - Upregulation of ciliary neurotrophic factor (CNTF) and CNTF receptor alpha in rat kidney with ischemia-reperfusion injury. AB - Ciliary neurotrophic factor (CNTF) is presumed to play a role as a survival factor in neuronal cells, but little is known about its role in the kidney. To investigate this, the expression of CNTF and CNTF receptor alpha (CNTFR alpha) was analyzed in the ischemic rat kidney. An ischemia/reperfusion (I/R) injury was induced by clamping both renal arteries for 45 min. Animals were killed at 1, 2, 3, 5, 7, 14, and 28 d after ischemia. The expression of CNTF and CNTFR alpha was monitored by reverse transcription-PCR, in situ hybridization, immunoblotting, immunohistochemistry, and electron microscopy. In sham-operated rat kidneys, CNTF expression was weak and limited to the descending thin limb of the loop of Henle. With I/R injury, CNTF mRNA and protein expressions were strikingly increased as compared with the sham-operated rat kidney, and the immunoreactivity of CNTF was mainly observed in the regenerating proximal tubules. The expression of CNTFR alpha mRNA was also increased after I/R injury, and its location and expression patterns were similar to the expression of CNTF. These findings suggest a possible role of CNTF as a growth factor during renal tubular repair processes after I/R injury and an autocrine or paracrine function of CNTF acting against CNTFR alpha. PMID- 11274237 TI - Puromycin aminonucleoside suppresses integrin expression in cultured glomerular epithelial cells. AB - Puromycin aminonucleoside (PAN)-induced nephrosis is a well-described model of human idiopathic nephrotic syndrome, but the mechanism of PAN's effect is not completely understood. Because PAN injection into rats results in retraction of glomerular epithelial cell foot processes and glomerular epithelial cell detachment, it was hypothesized that PAN might alter the contacts between these cells and the glomerular basement membrane. The major integrin expressed by glomerular epithelial cells is alpha3beta1, which mediates attachment of these cells to extracellular matrix proteins including type IV collagen. T-SV 40 immortalized human glomerular epithelial cells were used to study PAN's effects on alpha3beta1 expression, as well as that of podocalyxin and the slit diaphragm component ZO-1. Glomerular epithelial cells were seeded into plastic flasks and allowed to attach and proliferate for 48 h. The cells were then incubated for another 48 h in media containing 0, 0.5, or 5.0 microg/ml PAN. PAN exposure resulted in dose-dependent decreases in alpha3 and beta1 expression, both at the protein level and at the mRNA level. This was accompanied by a significant decrease in the adhesion of glomerular epithelial cells to type IV collagen. PAN did not affect ZO-1 protein expression. Treatment with PAN increased the expression of podocalyxin at the protein and mRNA levels. Reduced glomerular epithelial cell expression of alpha3beta1 integrins and impaired adhesion to type IV collagen may contribute to the glomerular epithelial cell detachment from glomerular basement membrane seen in the PAN nephrosis model. PMID- 11274238 TI - Verocytotoxin-induced apoptosis of human microvascular endothelial cells. AB - The pathogenesis of the epidemic form of hemolytic uremic syndrome is characterized by endothelial cell damage. In this study, the role of apoptosis in verocytotoxin (VT)-mediated endothelial cell death in human glomerular microvascular endothelial cells (GMVEC), human umbilical vein endothelial cells, and foreskin microvascular endothelial cells (FMVEC) was investigated. VT induced apoptosis in GMVEC and human umbilical vein endothelial cells when the cells were prestimulated with the inflammatory mediator tumor necrosis factor-alpha (TNF alpha). FMVEC displayed strong binding of VT and high susceptibility to VT under basal conditions, which made them suitable for the study of VT-induced apoptosis without TNF-alpha interference. On the basis of functional (flow cytometry and immunofluorescence microscopy using FITC-conjugated annexin V and propidium iodide), morphologic (transmission electron microscopy), and molecular (agarose gel electrophoresis of cellular DNA fragments) criteria, it was documented that VT induced programmed cell death in microvascular endothelial cells in a dose- and time-dependent manner. Furthermore, whereas partial inhibition of protein synthesis by VT was associated with a considerable number of apoptotic cells, comparable inhibition of protein synthesis by cycloheximide was not. This suggests that additional pathways, independent of protein synthesis inhibition, may be involved in VT-mediated apoptosis in microvascular endothelial cells. Specific inhibition of caspases by Ac-Asp-Glu-Val-Asp-CHO, but not by Ac-Tyr-Val Ala-Asp-CHO, was accompanied by inhibition of VT-induced apoptosis in FMVEC and TNF-alpha-treated GMVEC. These data indicate that VT can induce apoptosis in human microvascular endothelial cells. PMID- 11274239 TI - Functional, molecular, and biochemical characterization of streptozotocin-induced diabetes. AB - Altered divalent cation homeostasis with bone mineral loss, hypercalciuria, and hypomagnesemia have been associated consistently with human diabetes mellitus. This study investigated functional, molecular, and biochemical determinants that accompany this condition in chronically (2 wk) streptozotocin (STZ)-diabetic rats. Catheterized, conscious, diabetic rats on servo-controlled fluid replacement exhibited an increased GFR (+70%) and a substantially raised urinary calcium output (+568%) when compared with control rats. In addition, fractional calcium reabsorption was reduced, indicating that the hypercalciuria was not due solely to an osmotic effect but may involve an actual tubular defect. The expression of proteins involved in renal distal Ca2+ and water transport in STZ diabetic rats were then studied by Western analysis and immunofluorescence microscopy to investigate the molecular basis of the hypercalciuria. Extracellular Ca2+-sensing receptor abundance was reduced to 52% of control in STZ-diabetes, whereas thiazide-sensitive NaCl cotransporter expression was increased by 192%. Subcutaneous insulin implant rectified both functional and molecular parameters. The levels of calbindin D(28k), plasma membrane Ca2+ ATPase, and aquaporin 1 in whole kidney and of aquaporin 2 in inner medulla were unchanged in diabetic and/or insulin replacement. Blood levels of 1,25(OH)(2)D(3) were reduced in diabetes as were levels of osteocalcin, a marker of bone formation. It is concluded that diabetic hypercalciuria in rats involves elevated GFR with raised urinary output, reduced Ca2+ reabsorption, and impaired bone deposition. Changes in Ca2+-sensing receptor and NaCl cotransporter protein expression could account for the altered divalent cation homeostasis seen during diabetes mellitus. PMID- 11274240 TI - A randomized trial of high-dose compared with low-dose omega-3 fatty acids in severe IgA nephropathy. AB - Tested was the hypothesis that high-dose omega (omega)-3 fatty acids will be more effective than low-dose omega-3 fatty acids in preserving renal function in patients with severe IgA nephropathy in a randomized, open-label, parallel-group clinical trial. Patients were assigned to receive either high-dose fatty acids (EPA 3.76 g and DHA 2.94 g) or low-dose fatty acids (EPA 1.88 g and DHA 1.47 g), both given daily in a highly purified ethyl ester concentrate (Omacor). Patients were treated for a minimum of 2 yr in the absence of a treatment failure or until study closure (January 2000). Seventy-three patients were enrolled in the trial with two ranges of elevated serum creatinine (SC): 63 patients (86%) with a range of 1.5 to 2.9 mg/dl and 10 patients (14%) with a range of 3.0 to 4.9 mg/dl. The primary end point, within-patient rates of change in SC (2-yr minimum), showed an annualized median increase in SC of 0.08 mg/dl per yr in the low-dose group and 0.10 mg/dl per yr in the high-dose group (P: = 0.51). Patients in the lower entry SC range had lower SC slopes (P: = 0.02) and less end-stage renal disease (ESRD) (P: < 0.001) compared with those in the higher entry SC range. No patient died, and 18 patients developed ESRD: 10 in the low-dose group and 8 in the high-dose group (P: = 0.56). SC slopes were significantly lower, and survival free of ESRD was significantly higher (both, P: = 0.04) in the 63 Omacor-treated patients compared with the 22 placebo-treated patients from our previously reported clinical trial in which both groups had a similar level of renal impairment. Patient compliance was excellent, and no serious adverse events were noted. Low dose and high-dose omega-3 fatty acids were similar in slowing the rate of renal function loss in high-risk patients with IgA nephropathy, particularly those with moderately advanced disease. PMID- 11274241 TI - Detection of verocytotoxin bound to circulating polymorphonuclear leukocytes of patients with hemolytic uremic syndrome. AB - The epidemic form of hemolytic uremic syndrome (HUS) is the most common cause of acute renal failure in children and is characterized by a prodromal phase of sometimes bloody diarrhea. The role of verocytotoxin (VT)-producing Escherichia coli has been strongly implicated. Although antibodies against VT have been detected in the serum of patients with HUS, VT itself has never been detected in circulating blood. In this study, VT-2 was detected in the systemic circulation in 9 of 10 patients with the epidemic form of HUS. In those cases, VT-2 was bound exclusively to polymorphonuclear leukocytes (PMN). The detection of VT-2 bound to PMN was associated with the presence of diarrhea at the time the blood samples were obtained. The one patient for whom VT was not detected presented with atypical HUS. For 5 of the 10 patients with HUS who were studied, the time course of VT binding was analyzed; binding decreased in four patients. The finding of VT bound to PMN in the systemic circulation of patients with HUS is important for a clearer understanding of the pathogenesis of HUS and suggests new approaches for treatment in the future. PMID- 11274242 TI - Improving contact area between the peritoneum and intraperitoneal therapeutic solutions. AB - A general assumption in peritoneal dialysis or intraperitoneal chemotherapy has been that a volume of 2 to 3 L in the human is sufficient to make contact with the entire anatomic peritoneum. On the basis of our previous experimental work and that of others, it was hypothesized that only a fraction of the anatomic peritoneum was in contact with the therapeutic solution in the cavity over a short period of time. It was also hypothesized that use of agitation of the experimental animal or a surfactant in the dialysis fluid would increase the contact area of the intraperitoneal solution. These hypotheses were tested by developing a method to measure the peritoneal contact area simultaneously with the anatomic peritoneal area. Anesthetized mice (25 to 35 g) received an injection of a relatively large volume (10 ml) of isotonic solution containing a radiolabeled protein that adhered to the peritoneum with which it came in contact. After a dwell of 1 to 24 h, the animal was killed and frozen. Cross sections of the abdominal and pelvic cavities were cut and placed against film to develop into autoradiograms, which represent the linear dimension of fluid contact in each sampling plane. The tissue sections that corresponded to the autoradiograms were stained to display the linear dimension of the anatomic peritoneum in the sampling plane. By imaging both the autoradiogram and the corresponding histologic slide, an estimate of the ratio of the contact area to anatomic area in each plane can be calculated (R(mean) = average of all ratios). Applying this method to mice that were dialyzed with an isotonic salt solution under quiescent conditions for 1 h produced R(mean) = 0.43 +/- 0.03. With rapid shaking of the animal, R(mean) = 0.54 +/- 0.03 (P: < 0.05). Addition of the surfactant dioctyl sodium sulfosuccinate (DSS) 0.5% to the solution under quiescent conditions increased R(mean) to 1.07 +/- 0.03 (P: < 0.001). Lengthening the dwell of the isotonic solution to 24 h increased R(mean) to >0.90. In further study of the effect of the concentration of DSS on contact area, there was a direct correlation of R(mean) with concentrations ranging from 0.0005 to 0.05% DSS. It is concluded that less than half of the mouse peritoneum is in contact with a large volume of solution in the peritoneal cavity. Maneuvers such as agitation and use of surfactant in the intraperitoneal solution increase the fraction of contact area. Also demonstrated was a direct dose-response of contact area versus intraperitoneal concentration of DSS, which may be useful in intraperitoneal therapies of peritoneal dialysis or intraperitoneal chemotherapy. PMID- 11274243 TI - Is C-reactive protein a useful predictor of outcome in peritoneal dialysis patients? AB - An elevated C-reactive protein (CRP) has recently been shown to be strongly predictive of mortality in hemodialysis patients. However, its predictive value in peritoneal dialysis (PD) patients has not been assessed. A cohort of 50 PD patients was followed prospectively for a 3-yr period, after initial determination of CRP. Patients with an elevated CRP (>6 mg/L; n = 29) had significantly reduced plasma prealbumin (0.36 +/- 0.02 versus 0.44 +/- 0.03 g/L; P: < 0.05), decreased total weekly creatinine clearance (C(Cr); 52.5 +/- 2.3 versus 63.1 +/- 3.2 L/1.73 m(2); P: < 0.01), and increased left ventricular thickness (1.24 +/- 0.05 versus 1.08 +/- 0.06 cm; P: < 0.05) at baseline compared with those who had a normal CRP (< or =6 mg/L; n = 21). Baseline CRP (log transformed) correlated weakly with baseline Kt/V, C(Cr), and pre-albumin. With the use of a multivariate Cox's proportional hazards model to adjust for potential confounding factors, an elevated CRP was predictive of myocardial infarction (adjusted hazard ratio, 4.8; 95% confidence interval [CI], 1.0 to 23; P: = 0.048) and tended to be predictive of fatal myocardial infarction (adjusted hazard ratio, 6.0; 95% CI, 0.8 to 43; P: = 0.07). However, CRP was not significantly associated with all-cause mortality (adjusted hazard ratio, 2.1; 95% CI,0.8 to 5.4; P: = 0.15). In conclusion, CRP elevation occurs in a substantial proportion of PD patients and is independently predictive of future myocardial infarction. Such patients may warrant closer monitoring and attention to modifiable cardiovascular risk factors. PMID- 11274244 TI - Effects of losartan and amlodipine on intrarenal hemodynamics and TGF-beta(1) plasma levels in a crossover trial in renal transplant recipients. AB - Hypertension and hyperfiltration are two important risk factors for the development of chronic allograft nephropathy. Transforming growth factor-beta(1) (TGF-beta(1)) is the main cytokine involved in the fibrotic process that is involved in chronic rejection. Angiotensin II upregulates TGF-beta(1) production. Angiotensin II receptor antagonists therefore could not only control BP but also reduce TGF-beta(1) production in renal transplant patients. The aim of this study was to compare the effects of losartan and amlodipine on renal hemodynamics, as well as TGF-beta(1) and endothelin-1 (ET-1) plasma levels in a group of renal transplant patients who had normal renal function and who were treated with cyclosporine. Seventeen renal transplant patients who were receiving cyclosporine and who had normal graft function were included in a random 2 x 2 crossover trial with amlodipine and losartan (6 wk with each therapy). Three studies were performed (at baseline and at the end of both treatment periods) to determine renal hemodynamics, TGF-beta(1), and ET-1. Both treatments controlled BP to a similar degree, but only amlodipine increased GFR through an increase in the estimated glomerular hydrostatic pressure and filtration fraction. In contrast, losartan maintained GFR and reduced estimated glomerular hydrostatic pressure and filtration fraction significantly. Losartan and amlodipine had opposite effects on TGF-beta(1). Amlodipine did not affect TGF-beta(1) concentrations. In contrast, losartan reduced the plasma levels of TGF-beta(1) by approximately by 50% (from baseline, 5.2 to 2.6 ng/ml; P: = 0.01); the majority of the patients reached normal levels of TGF-beta(1). ET-1 concentrations were significantly higher during amlodipine compared with losartan treatment. The present study documents that with similar control of BP, losartan and amlodipine have opposite effects on renal hemodynamics and on TGF-beta1 concentrations. These differences could be important for the management of chronic allograft nephropathy. PMID- 11274245 TI - Approaching the therapeutic window for cyclosporine in kidney transplantation: a prospective study. AB - Neoral dosing is traditionally based on cyclosporine (CyA) trough levels (C(0)). Four-h area under the curve (AUC(0-4)) for Neoral in the early posttransplantation period was shown previously to have a better correlation to acute rejection (AR) and CyA nephrotoxicity (CyANT), compared with C(0). An AUC(0 4) range of 4400 to 5500 microg/h per L during the first week was associated with the lowest AR and CyANT. This article describes a prospective study to assess the feasibility, safety, and efficacy of dosing Neoral solely by AUC(0-4) monitoring, regardless of C(0), in the first 3 mo after kidney transplantation. Fifty-nine kidney transplant recipients received Neoral-based triple immunosuppression. AUC(0-4) was measured on days 3, 5, 7, 10, and 14 and weeks 3, 4, 6, and 8, then monthly. Target AUC(0-4) was 4400 to 5500 microg/h per L. Dose was adjusted by percentage difference from target AUC(0-4). Ninety-four percent of AUC were performed on the scheduled day or close to it. No patients had CyANT while AUC(0 4) was in target range. Four patients had reversible CyANT with AUC(0-4) > 5500. Only 1 of 33 patients (3%) who achieved and maintained AUC(0-4) > 4400 by day 3 posttransplantation had AR, whereas 10 of 22 (45%) of those with day 3 to 5 AUC(0 4) < 4400 had AR (P: = 0.0002). In logistic regression analysis, higher early AUC(0-4) was the only significant variable associated with lower serum creatinine at 3 mo. Neoral dose monitoring by AUC(0-4) is a potentially valuable tool for optimizing Neoral immunosuppression. Attainment of a target range of 4400 to 5500 microg/h per L for AUC(0-4) early after transplantation has been demonstrated to reduce significantly the risk of AR and CyANT. PMID- 11274246 TI - Polycystin: new aspects of structure, function, and regulation. AB - Polycystin-1 is a modular membrane protein with a long extracellular N-terminal portion that bears several ligand-binding domains, 11 transmembrane domains, and a > or =200 amino acid intracellular C-terminal portion with several phosphorylation signaling sites. Polycystin-1 is highly expressed in the basal membranes of ureteric bud epithelia during early development of the metanephric kidney, and disruption of the PKD1 gene in mice leads to cystic kidneys and embryonic or perinatal death. It is proposed that polycystin-1 functions as a matrix receptor to link the extracellular matrix to the actin cytoskeleton via focal adhesion proteins. Co-localization, co-sedimentation, and co immunoprecipitation studies show that polycystin-1 forms multiprotein complexes with alpha2beta1-integrin, talin, vinculin, paxillin, p130cas, focal adhesion kinase, and c-src in normal human fetal collecting tubules and sub-confluent epithelial cultures. In normal adult kidneys and confluent epithelial cultures, polycystin-1 is downregulated and forms complexes with the cell-cell adherens junction proteins E-cadherin and beta-, gamma-, and alpha-catenin. Polycystin-1 activation at the cell membrane leads to intracellular signaling via phosphorylation through the c-Jun terminal kinase and wnt pathways leading to activation of AP-1 and TCF/LEF-dependent genes, respectively. The C-terminal of polcystin-1 has been shown to be phosphorylated by c-src at Y4237, by protein kinase A at S4252, and by focal adhesion kinase and protein kinase X at yet-to-be identified residues. Inhibition of tyrosine phosphorylation or increased cellular calcium increases polycystin-1 focal adhesion complexes versus polycystin-1 adherens junction complexes, whereas disruption of the actin cytoskeleton dissociates all polycystin-1 complexes. Genetic evidence suggests that PKD1, PKD2, NPHP1, and tensin are in the same pathway. PMID- 11274247 TI - Treating IgA nephropathy. PMID- 11274248 TI - Cytomegalovirus in renal transplantation. AB - Cytomegalovirus (CMV) was first isolated from the salivary gland and kidney of two dying infants with cytomegalic inclusion bodies and reported in 1956 (1). Two other laboratories isolated CMV at approximately the same time. Thus, CMV was initially called "salivary gland virus" or "salivary gland inclusion disease virus". In 1960, Weller et al. (2) proposed the use of the term cytomegalovirus. Klemola and Kaarianinen (3) first described CMV mononucleosis, the principal presentation of previously healthy individuals, in 1965. CMV was first isolated in a renal transplant patient in 1965, PMID- 11274249 TI - Urea excretion in white rats and kangaroo rats as influenced by excitement and by diet. AB - Reprinted from The American Journal of Physiology, Vol. 181, No.1, April, 1955. PMID- 11274250 TI - Searching for the memory trace in a mini-brain, the honeybee. AB - To determine general or species-specific properties in neural systems, it is necessary to use comparative data in evaluating experimental findings. Presented here are data on associative learning and memory formation in honeybees, emphasizing a comparative approach. We focus on four aspects: (1) the role of an identified neuron, VUM(mx1), as a neural substrate of appetitive reinforcement; (2) the sequences of molecular events as they correlate with five forms of memory stages; (3) the localization of the memory traces following appetitive olfactory learning; and (4) the brief description of several forms of complex learning in bees (configuration in olfactory conditioning, categorization in visual feature learning, delayed matching-to-sample learning, and latent learning in navigation). VUM(mx1) activity following the conditioned stimulus odor is sufficient to replace the unconditioned stimulus, and VUM(mx1) changes its response properties during learning similarly to what is known from dopamine neurons in the basal ganglia of the mammalian brain. The transition from short- to mid- and long-term forms of memory can be related to specific activation of second messenger cascades (involving NOS, PKA, PKC, and PKM) resembling general features of neural plasticity at the cellular level. The particular time course of the various memory traces may be adapted to the behavioral context in which they are used; here, the foraging cycle of the bee. Memory traces for even such a simple form of learning as olfactory conditioning are multiple and distributed, involving first- and second-order sensory neuropils (antennal lobe and mushroom bodies), but with distinctly different properties. The wealth of complex forms of learning in the context of foraging indicates basic cognitive capacities based on rule extraction and context-dependent learning. It is believed that bees might be a useful model for studying cognitive faculties at a middle level of complexity. PMID- 11274251 TI - Analyses of habituation in Caenorhabditis elegans. AB - Although the nonassociative form of learning, habituation, is often described as the simplest form of learning, remarkably little is known about the cellular processes underlying its behavioral expression. Here, we review research on habituation in the nematode Caenorhabditis elegans that addresses habituation at behavioral, neural circuit, and genetic levels. This work highlights the need to understand the dynamics of a behavior before attempting to determine its underlying mechanism. In many cases knowing the characteristics of a behavior can direct or guide a search for underlying cellular mechanisms. We have highlighted the importance of interstimulus interval (ISI) in both short- and long-term habituation and suggested that different cellular mechanisms might underlie habituation at different ISIs. Like other organisms, C. elegans shows both accumulation of habituation with repeated training blocks and long-term retention of spaced or distributed training, but not for massed training. Exposure to heat shock during the interblock intervals eliminates the long-term memory for habituation but not the accumulation of short-term habituation over blocks of training. Analyses using laser ablation of identified neurons, and of identified mutants have shown that there are multiple sites of plasticity for the response and that glutamate plays a role in long-term retention of habituation training. PMID- 11274252 TI - Configural olfactory learning in honeybees: negative and positive patterning discrimination. AB - In an appetitive context, honeybees (Apis mellifera) learn to associate odors with a reward of sucrose solution. If an odor is presented immediately before the sucrose, an elemental association is formed that enables the odor to release the proboscis extension response (PER). Olfactory conditioning of PER was used to study whether, beyond elemental associations, honeybees are able to process configural associations. Bees were trained in a positive and anegative patterning discrimination problem. In the first problem, single odorants were nonreinforced whereas the compound was reinforced. In the second problem, single odorants were reinforced whereas the compound was nonreinforced. We studied whether bees can solve these problems and whether the ratio between the number of presentations of the reinforced stimuli and the number of presentations of the nonreinforced stimuli affects discrimination. Honeybees differentiated reinforced and nonreinforced stimuli in positive and negative patterning discriminations. They thus can process configural associations. The variation of the ratio of reinforced to nonreinforced stimuli modulated the amount of differentiation. The assignment of singular codes to complex odor blends could be implemented at the neural level: When bees are stimulated with odor mixtures, the activation patterns evoked at the primary olfactory neuropile, the antennal lobe, may be combinations of the single odorant responses that are not necessarily fully additive. PMID- 11274253 TI - Differential effects of damage within the hippocampal region on memory for a natural, nonspatial Odor-Odor Association. AB - Debate continues on whether the role of rodent hippocampus in memory is limited to the spatial domain. Recently, this controversy has been addressed with studies on the social transmission of food preference, an odor-odor association task with no spatial requirements. Multiple reports have concluded that damage to the hippocampal region impairs memory in this task, but there remain questions about the extent of damage essential to produce an impairment. Furthermore, a recent study () found no effect of hippocampal lesions on memory in this task. We tested animals with complete lesions of the hippocampus (H) lesions of the hippocampus plus subiculum (HS), and lesions of the adjacent, anatomically related cortices of the parahippocampal region (PHR). H lesions produced an impairment on spatial delayed alternation, but not on memory for the social transmission of food preference, whereas HS and PHR lesions produced severe and equivalent impairments on memory for the socially acquired food preference. We discuss possible explanations for the discrepancy with the results of and conclude that the hippocampus and subiculum together play a critical role in the formation of this form of nonspatial, relational memory. PMID- 11274254 TI - A neural circuit analysis of visual recognition memory: role of perirhinal, medial, and lateral entorhinal cortex. AB - Using a continuous recognition memory procedure for visual object information, we sequentially presented rats with eight novel objects and four repeated objects (chosen from the 8). These were selected from 120 different three-dimensional objects of varying sizes, shapes, textures, and degree of brightness. Repeated objects had lags ranging from 0 to 4 (from 0 to 4 different objects between the first and repeated presentation). An object was presented on one side of a long table divided in half by an opaque Plexiglas guillotine door, and the latency between opening the door and the rat moving the object was measured. The first presentation of an object resulted in reinforcement, but repeated presentations did not result in a reinforcement. After completion of acquisition training (significantly longer latencies for repeated presentation compared with the first presentation of an object), rats received lesions of the perirhinal, medial, or lateral entorhinal cortex or served as sham operated controls. On the basis of postsurgery testing and additional tests, the results indicated that rats with perirhinal cortex lesions had a sustained impairment in performing the task. There were no sustained deficits with medial or lateral entorhinal cortex lesions. The data suggest that recognition memory for visual object information is mediated primarily by the perirhinal cortex but not by the medial or lateral entorhinal cortex. PMID- 11274255 TI - Task-dependent role for dorsal striatum metabotropic glutamate receptors in memory. AB - The effect of post-training intradorsal striatal infusion of metabotropic glutamate receptor (mGluR) drugs on memory consolidation processes in an inhibitory avoidance (IA) task and visible/hidden platform water maze tasks was examined. In the IA task, adult male Long-Evans rats received post-training intracaudate infusions of the broad spectrum mGluR antagonist alpha-methyl-4 carboxyphenylglycine (MCPG; 1.0, 2.0 mM/0.5 microL), the group I/II mGluR agonist 1-aminocyclopentane-1,3-carboxylic acid (ACPD; 0.5 or 1.0 microM/0.5 microL), or saline immediately following footshock training, and retention was tested 24 h later. In the visible- and hidden-platform water maze tasks, rats received post training intracaudate infusions of ACPD (1.0 microM), MCPG (2.0 mM), or saline immediately following an eight-trial training session, followed by a retention test 24 h later. In the IA task, post-training infusion of ACPD (0.5 and 1.0 microM) or MCPG (1.0 and 2.0 mM) impaired retention. In the IA and visible platform water maze tasks, post-training infusion of ACPD (1.0 microM), or MCPG (2.0 mM) impaired retention. In contrast, neither drug affected retention when administered post-training in the hidden-platform task, consistent with the hypothesized role of the dorsal striatum in stimulus-response habit formation. When intradorsal striatal injections were delayed 2 h post-training in the visible-platform water maze task, neither drug affected retention, indicating a time-dependent effect of the immediate post-training injections on memory consolidation. It is hypothesized that MCPG impaired memory via a blockade of postsynaptic dorsal striatal mGluR's, while the impairing effect of ACPD may have been caused by an influence of this agonist on presynaptic "autoreceptor" striatal mGluR populations. PMID- 11274258 TI - The renal tubular Na-Cl co-transporter (NCCT): a potential genetic link between blood pressure and bone density? PMID- 11274257 TI - Emotional memory formation is enhanced across sleep intervals with high amounts of rapid eye movement sleep. AB - Recent studies indicated a selective activation during rapid eye movement (REM) sleep of the amygdala known to play a decisive role in the processing of emotional stimuli. This study compared memory retention of emotional versus neutral text material over intervals covering either early sleep known to be dominated by nonREM slow wave sleep (SWS) or late sleep, in which REM sleep is dominant. Two groups of men were tested across 3-h periods of early and late sleep (sleep group) or corresponding retention intervals filled with wakefulness (wake group). Sleep was recorded polysomnographically. Cortisol concentrations in saliva were monitored at acquisition and retrieval testing. As expected, the amount of REM sleep was about three times greater during late than during early retention sleep, whereas a reversed pattern was observed for SWS distribution (P < 0.001). Sleep improved retention, compared with the effects of wake intervals (P < 0.02). However, this effect was substantial only in the late night (P < 0.005), during which retention was generally worse than during the early night (P < 0.02). Late sleep particularly enhanced memory for emotional texts. This effect was highly significant in comparison with memory for neutral texts (P < 0.01) and in comparison with memory after late and early wake intervals (P < 0.001). Cortisol concentration differed between early and late retention intervals but not between sleep and wake conditions. Results are consonant with a supportive function of REM sleep predominating late sleep for the formation of emotional memory in humans. PMID- 11274260 TI - Diabetic foot ulcers: old problems--new technologies. PMID- 11274261 TI - Haemodialysis renal replacement therapy--do we need more research? PMID- 11274262 TI - Report on training sessions at the Prishtina University Hospital Department of Nephrology and Dialysis Unit, 22-29 July 2000, by the Joint Action Nephrology Eastern Europe of ISN and EDTA/ERA. PMID- 11274256 TI - Chronic, severe hypertension does not impair spatial learning and memory in Sprague-Dawley rats. AB - This study tested the hypothesis that long-term hypertension impairs spatial learning and memory in rats. In 6-wk-old Sprague-Dawley rats, chronic hypertension was induced by placing one of three sizes of stainless steel clips around the descending aorta (above the renal artery), resulting in a 20-80-mm Hg increase of arterial pressure in all arteries above the clip, that is, the upper trunk and head. Ten months later, the rats were tested for 5 d in a repeated acquisition water maze task, and on the fifth day, they were tested in a probe trial; that is, there was no escape platform present. At the end of the testing period, the nonsurgical and sham control groups had similar final escape latencies (16 +/- 4 sec and 23 +/- 9 sec, respectively) that were not significantly different from those of the three hypertensive groups. Rats with mild hypertension (140-160 mm Hg) had a final escape latency of 25 +/- 6 sec, whereas severely hypertensive rats (170-199 mm Hg) had a final escape latency of 21 +/- 7 sec and extremely hypertensive rats (>200 Hg) had a final escape latency of 19 +/- 5 sec. All five groups also displayed a similar preference for the correct quadrant in the probe trial. Together, these data suggest that sustained, severe hypertension for over 10 mo is not sufficient to impair spatial learning and memory deficits in otherwise normal rats. PMID- 11274263 TI - Identification of cellular origin of type I collagen in glomeruli of rats with crescentic glomerulonephritis induced by anti-glomerular basement membrane antibody. AB - BACKGROUND: Type I collagen is an interstitial collagen, which is not present in normal glomeruli. As type I collagen was observed in advanced glomerular lesions, it appears to be associated with deterioration of renal function. However, the origins of cells expressing type I collagen mRNA in glomeruli of diseased kidneys remains controversial. METHODS: We examined the expression of type I collagen in glomeruli at protein and mRNA levels in rat crescentic glomerulonephritis induced by anti-glomerular basement membrane (GBM) antibody. In addition, in situ hybridization and immunohistochemical staining of serial sections were performed to identify the cellular origin of type I collagen in glomeruli. RESULTS: Semi quantitative reverse transcription-polymerase chain reaction (RT-PCR) in isolated glomeruli showed that mRNA expression of type I collagen was remarkably increased on days 7, 14, and 28 after anti-GBM antibody injection (12.2+/-1.4, 20.2+/-2.1 and 14.6+/-1.0-fold over day 0, respectively). Immunofluorescence for type I collagen demonstrated marked staining in the fibrocellular and fibrous crescents, and weak staining within glomerular mesangial areas. In close association with mRNA levels analysed by RT-PCR, in situ hybridization revealed predominant presence of alpha1(I) collagen mRNA in cells within crescentic areas and Bowman's capsules. Serial section analysis for immunostaining and in situ hybridization showed that some alpha1(I) collagen mRNA-positive cells were also positive for cytokeratin. In contrast, no alpha1(I) collagen mRNA-positive cells were stained by ED-1 and podocalyxin. CONCLUSIONS: It appears that increased expression of type I collagen at the protein and mRNA levels in glomeruli is involved in the progression of glomerulonephritis. At least in this crescentic model, parietal epithelial cells (PECs) may partially contribute to the dysregulated production of type I collagen, which leads to glomerulosclerosis. PMID- 11274264 TI - Renal osteopontin protein and mRNA upregulation during acute nephrotoxicity in the rat. AB - BACKGROUND: The effect of segment-specific proximal tubular injury on spatio temporal osteopontin (OPN) distribution was determined in two different nephrotoxic rat models to evaluate its conceivability with a possible role for OPN in acute renal failure (ARF). OPN gene expression was further determined in proximal and distal tubular cells to investigate the origin of increased renal OPN. METHODS: Renal OPN protein and mRNA expression were compared in the rat during mercuric-chloride- vs gentamicin-induced ARF using immunohistochemistry and in situ hybridization. RESULTS: Mercuric chloride primarily induced tubular injury and subsequent cell proliferation in proximal straight tubules (PST), whereas gentamicin predominantly injured proximal convoluted tubules (PCT). In both models, the distribution of OPN protein was associated with increased OPN mRNA levels in proximal as well as distal tubular cells. However, upregulation was delayed in the proximal tubular segment suffering most from injury, i.e. PCT in gentamicin ARF vs PST in mercuric-chloride ARF. OPN immunostaining at the apical cell membrane from distal tubules was in contrast to perinuclear vesicular staining in proximal tubular cells. CONCLUSIONS: OPN gene and protein expression is induced in both proximal and distal tubular cells during rat toxic ARF. The distinct subcellular localization in proximal vs distal tubular cells indicates differences in OPN processing and/or handling. The spatio-temporal distribution is consistent with a possible role in renal injury and regeneration. PMID- 11274265 TI - Selective blockade of vasopressin V2 receptors reveals significant V2-mediated water reabsorption in Brattleboro rats with diabetes insipidus. AB - BACKGROUND: In a previous study we observed that acute administration of the selective antagonist of vasopressin (AVP) V2 receptors, SR 121463A (SR), aggravated the symptoms of diabetes insipidus (DI) in homozygous Brattleboro rats (an AVP-deficient strain). The present study investigates in more details the acute and chronic effects of SR in DI rats. METHODS AND RESULTS: In experiment A, different groups of rats received acute i.p. injections of SR (0.001-10 mg/kg) or vehicle alone, and urine was collected for the next 24 h. SR dose-dependently increased urine flow rate and decreased urine osmolality with no significant change in solute excretion, thus confirming a pure 'aquaretic' effect. In experiments B and C, the chronic effects of orally administered SR were evaluated over 8 days in Brattleboro DI rats (experiment B, 1 mg/kg/day) and in adult Sprague-Dawley rats with normal AVP secretion (experiment C, 3 mg/kg/day). In DI rats, the aquaretic effects of SR persisted with the same intensity over the 8 days. In Sprague-Dawley rats, SR induced a sustained, stable aquaretic effect and also increased non-renal water losses, suggesting an effect of AVP on water conservation in extrarenal sites. Because oxytocin (OT) synthesis is elevated in DI rats and OT is known to bind to V2 receptors, we evaluated the antidiuretic effects of OT in DI rats in experiment D. Chronic infusion of OT (3 microg/kg/h, i.p.) induced a marked antidiuresis, and acute SR (1 mg/kg) in OT-treated DI rats completely abolished this antidiuretic effect, thus indicating that it was due to binding of OT to V2 receptors. CONCLUSION: (i) SR is a potent orally active aquaretic and induces stable effects during 1 week in rats with or without endogenous AVP secretion. (ii) Significant V2 receptor-mediated water reabsorption occurs in collecting ducts of Brattleboro DI rats because their usual urine osmolality is about twofold higher than the minimum observed during SR-induced maximum diuresis. (iii) This V2 agonism could be mediated in part by OT binding to V2 receptors. Small amounts of endogenous AVP, known to be produced by adrenal and testis in DI rats, could also contribute to this V2 agonism, as well as a possible constitutive activation of the V2 receptors. (iv) In normal rats, AVP probably reduces water losses through extrarenal sites, probably the lungs. PMID- 11274266 TI - Protective effect of UR-12670 on chronic nephropathy induced by warm ischaemia in ageing uninephrectomized rats. AB - BACKGROUND: In young animals, renal ischaemia/reperfusion injury and mass reduction are associated with chronic lesions that mimic those found in chronic rejection. We have shown that the phospholipid platelet-activating factor (PAF) participates in young animals in such chronic nephropathy. Here we examine the long-term effects of the orally active PAF antagonist, UR-12670 in ageing uninephrectomized rats exposed to prolonged warm ischaemia. METHODS: Fifteen- to eighteen-month-old uninephrectomized male Sprague-Dawley rats were allocated into three groups and followed for 16 weeks: UNx, rats without ischaemia; UNxISC, ischaemic kidney (60 min), and UNxISC+UR, ischaemic kidney and UR-12670 from day 0 to the 16th week. Serum creatinine and proteinuria were monitored every 4 weeks. At the end of the study, conventional histology was performed and monocyte macrophages were identified with the specific monoclonal antibody ED-1. RESULTS: The UNxISC group had severe acute renal failure with a high mortality rate, which was associated with incomplete restoration of renal function. Renal insufficiency in this group was sustained throughout the follow-up. Both UNx and UNxISC groups developed progressive proteinuria from the 12th week. Though UNxISC+UR group showed similar acute renal failure and mortality rate to the ischaemic non treated group, serum creatinine decreased to levels similar to UNx group, which were maintained until the end of the study. Treatment of ischaemic kidneys with UR-12670 produced a slight decrease in 24-h proteinuria and a reduction in glomerulosclerosis, the mean tubulointerstitial score and number of monocyte macrophages to values similar to UNx group. CONCLUSIONS: The chronic administration of the PAF antagonist UR-12670 attenuates the long-term effects of ischaemia-reperfusion injury in uninephrectomized ageing rats. The beneficial effect of this agent suggests that PAF contributes to the progression to late renal damage in this model. PMID- 11274267 TI - Effect of aluminium load on parathyroid hormone synthesis. AB - BACKGROUND: Aluminium overload leads to parathyroid hormone (PTH) suppression. However, it is unclear whether a decrease in synthesis or release of the hormone is mainly involved. The aim of this study was to assess the effect of an acute administration of aluminium on PTH synthesis and release in rats with chronic renal failure and secondary hyperparathyroidism. METHODS: The study was performed using 100 adult male Wistar rats (body weight 443+/-54 g). 7/8 nephrectomy was performed and the rats were maintained on a high dietary phosphorous intake. Five weeks after surgery, the rats were randomly divided into two groups, one loaded with aluminium (AlCl3) and the other given placebo. Aluminium or placebo were administered i.p. for two consecutive days. The placebo group received saline at the same pH as the aluminium solution. After 2 weeks, serum calcium, phosphorous, creatinine, PTH, and aluminium were measured. The parathyroid glands were removed and PTH messenger RNA (mRNA) was measured by northern blot. Intact PTH was measured by IRMA (Rat PTH, Nichols Institute). RESULTS: No differences in serum PTH levels were found between the two groups after 5 weeks of renal failure. At the end of the study the rats given aluminium had higher aluminium levels than the placebo group and lower PTH levels. No significant differences were found for calcium, phosphorous, renal function, or body weight. PTH mRNA expression was lower in the aluminium group than in the placebo group. CONCLUSION: The administration of aluminium in rats with chronic renal failure resulted in reductions in serum PTH and PTH mRNA. Thus far, previous studies had demonstrated that aluminium suppressed PTH release. The present findings suggest that PTH synthesis is also reduced. PMID- 11274268 TI - Supplementation with a low dose of L-arginine reduces blood pressure and endothelin-1 production in hypertensive uraemic rats. AB - BACKGROUND: We documented recently that increased endothelin-1 (ET-1) production in blood vessels and glomeruli of uraemic rats plays a crucial role in the development of hypertension and the progression of chronic renal failure. Normally, biological effects and local production of ET-1 are attenuated by the immediate release of nitric oxide (NO). Increasing evidence suggest, however, that NO release is impaired in chronic renal failure. We investigated whether supplementation with L-arginine, the natural precursor of NO, improves NO synthesis in uraemic rats with reduced renal mass and modulates vascular and renal ET-1 production as well as blood pressure and renal failure progression. METHODS: One week after surgical renal mass reduction, the uraemic and sham operated animals received either no treatment or 0.1% L-arginine in drinking water for 5 weeks. In another series of experiments, uraemic rats received 1% L arginine for 5 weeks. Immunoreactive-ET-1 (ir-ET-1) levels in plasma, urine, and vascular and renal tissue preparations was measured by radioimmunoassay after sample extraction and purification. RESULTS: Before treatment, systolic blood pressure was significantly elevated in uraemic animals compared to sham-operated controls (156+/-7 vs 111+/-3 mmHg, respectively; P<0.01). Thereafter, systolic blood pressure increased further in uraemic-untreated rats (systolic blood pressure at week 5; 199+/-9 mmHg, P<0.01), whereas it remained similar in uraemic rats supplemented with 0.1% L-arginine (171+/-9 mmHg, NS). At the end of the study, serum creatinine and urea, proteinuria and ir-ET-1 excretion were significantly augmented, while creatinine clearance was reduced in uraemic animals compared to the controls. Ir-ET-1 level was also increased in glomeruli as well as in thoracic aorta, mesenteric arterial bed, and pre-glomerular arteries, and was associated with vascular hypertrophy as assessed by tissue weight. In contrast, ir-ET-1 level was diminished in the renal papilla of uraemic rats. Treatment with 0.1% L-arginine significantly reduced proteinuria and urinary ir-ET-1 excretion (P<0.05) as well as ir-ET-1 level in glomeruli (P<0.01) and in thoracic aorta (P<0.05). These changes were associated with increased plasma NO metabolites NO2/NO3 levels in L-arginine-treated animals (P<0.01) and reduced aortic hypertrophy (P<0.05). In contrast, supplementation with 1% L arginine had no effect on systolic blood pressure in uraemic rats, but exacerbated proteinuria and urinary ir-ET-1 excretion and increased serum urea (P<0.05) were observed. CONCLUSIONS: These results indicate that improvement of NO release with a low dose but not with a high dose of L-arginine significantly attenuates the development of hypertension and the progression of renal insufficiency in rats with reduced renal mass. These protective effects may be mediated in part by the reduction of vascular and renal ET-1 production. PMID- 11274269 TI - Evidence for further genetic heterogeneity in nephronophthisis. AB - BACKGROUND: A new type of nephronophthisis (NPH) has been recently identified in a large Venezuelan kindred: adolescent nephronophthisis (NPH3) causes end-stage renal disease (ESRD) at a median age of 19 years. The responsible gene (NPHP3) maps to 3q21-q22. NPH3 shares with juvenile nephronophthisis (NPH1) the same disease manifestations such as polyuria, polydipsia, and secondary enuresis. Histopathological findings consist of tubular basement membrane changes, cysts at the corticomedullary junction, and a chronic sclerosing tubulointerstitial nephropathy. The only difference is a younger age at ESRD in NPH1 (median age of 13 years) when compared with NPH3. METHODS: In order to evaluate whether there might be a fourth locus of isolated nephronophthisis, we studied eight NPH families without extrarenal disease manifestations and without linkage to the NPH1 locus (NPHP1) on chromosome 2q12-q13. ESRD was reached at ages ranging from 7 to 33 years. Individuals were haplotyped with microsatellites covering the genetic locus of NPHP3. Infantile NPH (NPH2) was excluded in all families by the clinical history and histological findings. RESULTS: In four of the examined families haplotype analysis was compatible with linkage to the NPHP3 locus. In one of these families identity by descent was observed. In contrast, in another four families linkage was excluded for NPHP3. CONCLUSION: Four NPH-families were neither linked to NPHP1 nor to NPHP3, indicating further genetic heterogeneity within the group of nephronophthisis. The finding of further genetic heterogeneity in NPH has important implications for genetic counselling. PMID- 11274270 TI - Polymorphisms in the promoter region and at codon 54 of the MBL2 gene are not associated with IgA nephropathy. AB - BACKGROUND: IgA nephropathy (IgAN) occurs sporadically in unrelated individuals. Several different polymorphic genes have been investigated in recent years in order to demonstrate their possible association with IgAN. Three recent, different studies with conflicting conclusions have discussed the role of the mannose binding lectin (MBL), a serum lectin involved in natural immunity, in the IgAN pathogenesis by examination of MBL deposits in biopsies. In the present study we investigated several polymorphisms of the MBL gene located in the promoter region and in the first exon. METHODS: MBL polymorphism detection was performed in 22 Italian patients with familial IgA nephropathy and in 138 Italian patients with the sporadic form of the disease. The polymorphisms in the MBL2 promoter region and in the exon 1 were investigated, respectively, by direct sequencing and by amplification refractory mutation system-polymerase chain reaction on genomic DNA collected from peripheral blood. Seventy-four unrelated healthy subjects matched for ethnic origin were used as controls. RESULTS: Allelic and genotypic frequencies of the polymorphisms at position -550, -328, 221 and at codon 54 did not show any differences between patients and controls. Similar frequency distributions of these polymorphisms were also found in the subgroups of IgAN patients subdivided according to the clinical manifestations and the progression of the disease. CONCLUSIONS: This study indicates that the analysed polymorphisms of the MBL gene do not appear to be primarily involved in the susceptibility and severity of IgAN. PMID- 11274271 TI - Clinicopathological correlation in biopsy-proven atherosclerotic nephropathy: implications for renal functional outcome in atherosclerotic renovascular disease. AB - BACKGROUND: Atherosclerotic renovascular disease (ARVD) is commonly associated with renal failure. It is now recognized that intrarenal damage, (ischaemic or atherosclerotic nephropathy) is a major contributor to the renal impairment in these patients. In this study the impact of histological changes upon renal functional outcome was investigated in patients with atherosclerotic nephropathy. METHODS: The Hope Hospital renal biopsy database (1985-1998) was interrogated for patients with histology compatible with atherosclerotic nephropathy. Case-note review enabled the assessment of several clinical parameters and outcomes, including change in creatinine clearance per year (DeltaCrCl (ml/min/year)), blood pressure control, dialysis need, and death. Renal parenchymal damage was analysed by morphometric analysis (of interstitial fibrosis and glomerulosclerosis) and a semi-quantitative chronic damage score (score 0-3 (normal-severe) for each of glomerulosclerosis, interstitial fibrosis, tubular atrophy, and arteriolar hyalinosis; maximum=12). Patients were stratified into two groups who had either deteriorating (group 1) or stable (group 2) renal function during follow-up. RESULTS: Twenty-five patients (age 64.7+/-10.5, range 43-83 years; 17 male, eight female) were identified. Sixteen patients had undergone angiography; two had significant (>50%) renal artery stenosis. Mean follow-up was 25.6+/-14.8 (range 5-50) months. Group 1 patients had DeltaCrCl 7.4+/-6.8 ml/min/year, n=14 and group 2 patients had DeltaCrCl 4.8+/-7.0 ml/min/year, n=11. Four patients in group 1 developed end-stage renal disease and five patients died (three in group 1 and two in group 2). At study entry, group 1 patients had worse renal function (CrCl 27.6+/-17.6 vs 36.0+/-33.9, NS), greater proteinuria (1.2 vs 0.5 g/24 h, NS), and higher systolic blood pressure (167.1+/ 30.8 mmHg vs 150.6+/-37.8, NS) compared with group 2 patients. Group 1 patients showed more glomerulosclerosis (51.6 vs 24.9%, P:<0.01), greater proportional interstitial volume (44.9 vs 33.9%, P:<0.02), and higher overall chronic damage score (P:<0.05) than those in group 2. There was a significant correlation between renal functional outcome and chronic damage score, glomerulosclerosis and proportional interstitial volume for the entire patient cohort. CONCLUSION: In patients with atherosclerotic nephropathy the severity of histopathological damage is an important determinant and predictor of renal functional outcome. PMID- 11274273 TI - Uraemic symptoms, nutritional status and renal function in pre-dialysis end-stage renal failure patients. AB - BACKGROUND: Deciding on the right moment to initiate dialysis and finding the best method to establish this critical stage of chronic renal failure are both controversial issues. This study attempted to address this subject by correlating a uraemic score with the most common clinical methods for assessing renal function in pre-dialysis chronic renal failure (end-stage renal disease, ESRD) patients. METHODS: The study group consisted of 201 non-selected ESRD patients. A uraemic score, composed of the uraemic symptoms, the subjective global assessment of nutritional status, serum albumin concentration, and protein catabolic rate normalized for ideal body weight, was taken as a clinical marker of uraemic toxicity. Correlations that best fit this uraemic score with creatinine clearance (Ccr), the arithmetic mean of Ccr, urea clearance (Ccr-Cu) and Kt/V urea were then investigated. RESULTS: Thirty-six per cent of patients had malnutrition. By multiple logistic regression analysis, the presence of comorbidity, Ccr-Cu and haematocrit were the best determinants of malnutrition. The correlation that best fit Ccr or Ccr-Cu with the uraemic score was a cubic curve (r=0.38, P<0.0001, and r=0.42, P<0.0001, respectively), in which an ascending inflection was observed when Ccr and Ccr-Cu fell below 12-13 and 10 ml/min, respectively. However, the relationship between Kt/V urea and the uraemic score was less predictable, especially in male patients. CONCLUSION: Ccr or Ccr-Cu are reliable methods for establishing the degree of severity of chronic renal failure below which the development of symptoms and malnutrition are highly prevalent. In contrast, Kt/V urea may be a less sensitive and specific method for assessing the severity of uraemia in ESRD patients. PMID- 11274272 TI - Time course of the antiproteinuric and renal haemodynamic responses to losartan in microalbuminuric IDDM. AB - BACKGROUND: Interference in the renin-angiotensin system with angiotensin converting enzyme (ACE) inhibitors has proven to be effective in lowering albuminuria in patients with insulin-dependent diabetes mellitus (IDDM). We studied whether angiotensin II receptor antagonism reduces urinary albumin excretion (UAE) in IDDM patients, and the relationship between the antiproteinuric effect and changes in systemic and renal haemodynamics. METHODS: Nine IDDM patients with microalbuminuria (30-300 mg/24 h) were studied. Patients were studied after a 4 week placebo period, on days 3, 7 and 28 of treatment with losartan 50 mg once daily, and after a 4 week placebo-controlled recovery period. RESULTS: Mean arterial pressure (MAP) was only slightly lowered during losartan treatment. Effective renal plasma flow (ERPF) was significantly increased on the third day of treatment and remained stable throughout the treatment period. Glomerular filtration rate (GFR) did not change throughout the study. Filtration fraction (FF) was maximally lowered on the third day of treatment and remained stable during treatment. UAE was already significantly lowered after 2 days of treatment, during both the day and night, and remained stable throughout the treatment period. The time course of the changes in UAE paralleled that of the changes in MAP, ERPF and FF. CONCLUSIONS: The angiotensin receptor antagonist losartan effectively lowers UAE in microalbuminuric IDDM patients. The changes observed in renal haemodynamics and UAE are concordant in time and maximal within only a few days of treatment. These results support the importance of the specific effects of interference in the renin angiotensin system (RAS) in microalbuminuric IDDM on blood pressure and renal haemodynamics in reducing urinary protein leakage, rather than non-haemodynamic, structural changes of the glomerular basement membrane. PMID- 11274274 TI - Plasma levels of mature form of adrenomedullin in patients with haemodialysis. AB - BACKGROUND: Adrenomedullin (AM) is a potent vasodilator and natriuretic peptide with hypotensive effects. Immunoreactive AM in human plasma consists of the biologically active mature form, AM (1-52)-CONH2 (mAM) and the intermediate form, AM-gly-COOH (iAM). However, the different effects of mAM and iAM in patients on haemodialysis (HD) have remained unclear. METHODS: Thirty-nine patients on HD and 10 controls were included in this study. We determined plasma levels of mAM and iAM using an immunoradiometric assay that recognizes total AM (tAM) and another that is specific for only mAM. RESULTS: The plasma concentrations of mAM and iAM in patients before HD were significantly higher than those in the controls (n=10) (4.76+/-0.28 vs 1.28+/-0.22 fmol/ml, P<0.001, 25.99+/-1.47 vs 8.52+/- 0.91 fmol/ml, P<0.001 respectively). The plasma levels of mAM and iAM before HD significantly and negatively correlated with systolic blood pressure (SBP) (r= 0.46, P<0.01, and r=-0.32, P<0.05 respectively) and diastolic blood pressure (DBP) (r=-0.32, P<0.05, and r=-0.35, P<0.05 respectively). After HD, plasma mAM and iAM levels as well as SBP and DBP were significantly lower than before HD. Plasma levels of mAM and iAM correlated significantly (r=0.73, P<0.001). CONCLUSIONS: These data suggest that mAM and/or iAM are involved in blood pressure regulation in patients undergoing HD, and further work is needed to understand the precise role of adrenomedullin in this regulation. PMID- 11274275 TI - Soluble thrombomodulin is associated with viral hepatitis, blood pressure, and medications in haemodialysis patients. AB - BACKGROUND: The level of soluble thrombomodulin (sTM), a traditional marker of endothelial injury, is also dependent on renal excretory function. We studied serum sTM in chronic haemodialysis (HD) patients to determine which factors are predictive of its levels in this population. METHODS AND RESULTS: sTM levels of 10.7 (5.72-30.7) ng/ml in 100 HD patients were higher than in 30 controls (P<0.0001). In a bivariate regression analysis, immunoreactive sTM was positively associated with the presence of hepatitis B virus surface antigen and/or anti hepatitis C virus antibodies measured by third generation ELISAs (P<0.0001), and was related to certain markers of liver injury and biosynthetic dysfunction. sTM was also directly associated with time on dialysis (P=0.001), or use of unfractionated heparin (UFH) (vs enoxaparin) (P=0.0007), erythropoietin (P=0.008), ACE-inhibitors (P=0.034), acetate-buffered dialysate (vs bicarbonate) (P=0.040), pre-dialysis systolic (P=0.012), and diastolic blood pressure (P=0.043). It was negatively associated with lipoprotein(a) (P=0.029). sTM was not related to age, sex, smoking, cause of renal failure, prevalence of cardiovascular disease, amount of HD delivered, preserved residual renal function, ferritin, C-reactive protein, and other vasoactive medications used. In a multivariable analysis, a positive hepatitis marker (P=0.0002), the use of UFH (P=0.030) and erythropoietin (P=0.019), and raised pre-dialysis blood pressure (P=0.024) were positive independent predictors of high sTM level. CONCLUSION: These data indicate that, in addition to endothelial activation, elevated sTM levels in HD patients may be related to viral infection and/or liver dysfunction, and influenced by modifiable factors such as increased blood pressure, and the type of heparin and erythropoietin treatment used. PMID- 11274276 TI - Baseline blood pressure and other variables influencing survival on haemodialysis of patients without overt cardiovascular disease. AB - BACKGROUND: Age, diabetes and concomitant cardiovascular disease, recorded at the initiation of dialysis, allows the identification of patients with a high probability of early mortality. When all of these factors are taken into account the mortality rate of dialysis patients is still 3.5 times higher than for the general population. Information on the factors that increase the mortality of patients lacking the major cardiovascular risk factors is important because these are likely to be correctable, especially if detected early. METHODS: We investigated prospectively the relevance of blood pressure and other variables recorded at the initiation of dialysis treatment on the survival of a group of 103 relatively young adult haemodialysis patients (mean age 44.3 years +/-13 SD), with a low prevalence of comorbidity and a median follow-up period of 79 months. Data were analysed by the Cox proportional regression model and survival curves were constructed by the Kaplan-Meier method. RESULTS: Forty-four patients died, 20 (46%) of whom as a result of cardiovascular causes. Multivariate analysis showed that mortality was associated with age (P=0.0001), serum creatinine (P=0.005, negative association), left ventricular (LV) mass (P=0.003) and hypertension (P=0.03). Mortality was increased by 7% for each additional year of age, by 0.7% for each 1 g increase in LV mass, and was reduced by 23% for each additional mg/dl of serum creatinine. Hypertensive patients had a higher probability (x2.2) of dying compared with normotensive patients. CONCLUSIONS: In addition to age and conditions of occult malnutrition, hypertension and LV hypertrophy, when present at the initiation of dialysis, play a major role in the mortality of low risk, relatively young dialysis patients. These potentially correctable factors should be actively sought and treated during the early stage of renal insufficiency to improve prognosis. PMID- 11274277 TI - Changes in major blood components after adopting the supine position during haemodialysis. AB - BACKGROUND: In Japan, haemodialysis (HD) is usually performed with patients in the supine position. However, the effects of changing posture on major blood components have not been investigated in HD patients. It is possible that several fluid components change rapidly when patients change from the upright to the supine position. We therefore investigated the effects of posture on blood component analysis. METHODS: A first blood sample was taken from 10 HD patients 5 min after they adopted a supine position; HD was begun immediately after sampling. Additional blood samples were collected 15 and 30 min later while patients remained in the supine position. On an alternate day, blood samples were taken from these same patients in the supine position, but not during HD. The same procedure was performed in 10 healthy volunteers. RESULTS: Haematocrit significantly decreased in patients undergoing HD at 15 and 30 min into the HD session. Similar decreases were observed in HD patients not undergoing HD and in normal control subjects. Haematocrit changes at 15 min were not significantly different between the three groups. Serum albumin concentrations decreased in the same way as haematocrit. Consequently, the reductions in haematocrit and albumin concentrations in HD patients during the HD session were not attributable to the HD procedure or to end-stage renal disease, but rather were due to the supine position and consequent haemodilution caused by redistribution of water from the extra- to the intravascular space. Finally, WBC counts decreased significantly at 15 min in both HD patient groups and in normal controls. The relative decrease at 15 min was significantly greater in HD patients undergoing HD (61.4% of baseline) than in those not undergoing HD (88.0%) or in normal controls (94.7%). These differences were probably due to previously reported WBC sequestration in the lungs during the early phase of HD. CONCLUSIONS: This study suggests that the change from the upright to the supine positions during HD causes changes in blood components that are critical for quality control determinations. PMID- 11274278 TI - Prostaglandin inhibition by intraperitoneal indomethacin has no effect on peritoneal permeability during stable CAPD. AB - BACKGROUND: Prostaglandins can affect the vascular response and are locally produced in the peritoneal cavity. Prostaglandin inhibition in continuous ambulatory peritoneal dialysis (CAPD) patients during peritonitis using indomethacin intraperitoneally was found to decrease the intrinsic permeability to macromolecules. METHODS: In the present study the effects of prostaglandin inhibition were studied during stable, uninfected CAPD. Two standard peritoneal permeability analyses (1.36% glucose) were performed in 10 stable CAPD patients within 1 week with and without addition of 12.5 mg/l indomethacin. Furthermore, possible effects on the parameters of nitric oxide synthesis were determined. In five other patients a high dose of indomethacin was tested. The night before the indomethacin test, 12.5 mg/l indomethacin was added to the nightdwell and the test was performed with 25 mg/l indomethacin. RESULTS: In the normal dose indomethacin group, the dialysate concentrations of prostaglandin (PG) 6-keto PGF1alpha and thromboxane (Tx) TxB2 were significantly lower with indomethacin (IND) compared with the control dwell (C): 6-keto-PGF1alpha median 93 (C) vs 7.5 (IND) ng/l, P=0.006 and TxB2 12.3 (C) vs 9.0 (IND) ng/l, P=0.04. The dialysate concentration of PGE2 was not different during the control dwell (68.5 ng/l) compared with the indomethacin experiment (50.3 ng/l, P=0.5). The mass transfer area coefficients (MTAC) of nitrate and cGMP, and parameters of nitric oxide synthesis, were similar during both experiments. The MTAC of creatinine and urate were not different with indomethacin: creatinine median 9.5 (C) vs 10.2 ml/min (IND), P=0.2 and urate 7.2 (C) vs 7.3 ml/min (IND), P=0.3. Only the MTAC of urea was marginally higher with indomethacin: 16.0 (C) vs 16.6 ml/min (IND), P=0.04. No differences were found in the clearances of the macromolecules beta2 microglobulin, albumin, IgG and alpha2-microglobulin. With the high indomethacin dose no inhibition of PGE2 was found: 69 (C) vs 63 ng/l(IND), not significant. Furthermore, no differences were found in the transport rates of small solutes or proteins. This indicates no effect of indomethacin on the peritoneal surface area and the size-selective permeability to macromolecules. In both groups no effect was found on the transcapillary ultrafiltration and the effective lymphatic absorption rate during the 4-h dwell. Consequently, the net ultrafiltration, the difference between these, did not change. CONCLUSIONS: The indomethacin induced inhibition of the synthesis of 6-keto-PGF1alpha and TxB2 did not lead to alterations in functional parameters of the peritoneal surface area, the intrinsic permeability to macromolecules and fluid kinetics. Therefore, these prostaglandins are not likely to be involved in the regulation of peritoneal transport during stable CAPD. PMID- 11274279 TI - C-reactive protein and chronic Chlamydia pneumoniae infection--long-term predictors for cardiovascular disease and survival in patients on peritoneal dialysis. AB - BACKGROUND: Accelerated arteriosclerosis with cardiovascular disease is the main cause of death in end-stage renal disease patients. Increased, levels of C reactive protein (CRP) and evidence of chronic Chlamydia pneumoniae infection have been identified as risk factors for cardiovascular disease in the general population. We tested the hypothesis that elevation of CRP, indicating chronic inflammation, and positive serum antibody titres for C. pneumoniae are associated with an increased cardiovascular mortality in patients on chronic peritoneal dialysis. METHODS: We measured CRP and antibodies to C. pneumoniae in 34 patients on peritoneal dialysis. CRP was measured by a sensitive ELISA and C. pneumoniae antibodies by microimmunofluorescence. In addition, risk factors such as lipids, smoking status and hypertension were assessed. Coronary artery disease (CAD) was defined by cardiac stress testing and/or angiography. Patients showing clinical evidence of systemic or peritoneal dialysis-associated infection during the investigation period of 6 months (between 1990 and 1991) were excluded. RESULTS: The incidence of CAD was significantly increased in patients with CRP values >1.5 mg/l (odds ratio 7.0, P<0.022) during 72 months of follow-up. In addition, in patients seropositive for IgA C. pneumoniae antibodies, the incidence of CAD was significantly increased (odds ratio 7.2, P<0.014). These findings resulted in an increased risk of death in patients with mean CRP values >1.5 mg/l at the start of the study (odds ratio 20.0, P<0.001). Furthermore, in patients seropositive for IgA C. pneumoniae antibodies, the risk of death (odds ratio 10.2, P<0.005) was significantly increased. There was a highly significant correlation between CRP and seropositivity for IgA C. pneumoniae antibodies (r=0.445, P<0.01). CONCLUSIONS: Increased circulating CRP and seropositivity for C. pneumoniae in patients on chronic peritoneal dialysis are associated with reduced survival due to cardiovascular complications. CRP and C. pneumoniae antibodies may indicate a chronic inflammatory process as an underlying cause and/or result of arteriosclerosis. PMID- 11274280 TI - The effect of altering eligibility criteria for entry onto a kidney transplant waiting list. AB - BACKGROUND: This paper concerns the allocation of kidneys from cadaveric donors to patients with end-stage renal disease (ESRD). Currently, the decision as to whether or not a particular patient should go onto the renal transplant waiting list is left to the discretion of the local dialysis centre, and is usually based almost entirely upon consideration of each case on its individual merits. Would this person like to have a renal transplant, is this possible, and would it seem reasonable to give them a chance? It could be argued that such an approach may not make best use of a scarce national resource. In this study we explore the effects of altering the eligibility criteria for transplantation to take explicit and quantitative account of the fact that some patients are more likely to die than others. METHODS: We performed a survey of one unit's dialysis patients to ascertain the characteristics used in practice to determine who should go onto the transplant waiting list and who should not. We then created a computer model to simulate a cohort of ESRD patients, initially of the same size and characteristics as that in the unit surveyed, receiving renal replacement therapy over a period of 10 years. Using this model, we compared four strategies for defining eligibility for transplantation: (1) all patients eligible; (2) standard and medium risk patients eligible; (3) only standard risk patients eligible; and (4) no regrafts performed (standard and medium risk according to definitions in the Renal Association Standards Document). RESULTS: Strategies of allowing only standard or standard and medium risk patients onto the waiting list most closely reflected the current decisions made regarding eligibility. The different strategies considered in the models necessarily gave rise to very considerable variation in the size of the waiting list at the end of the 10 year period (range 98-368), which would have important practical implications. The predicted mean time of kidney function varied from 9.8 years for strategy 4 (no regrafts) to 10.8 years for strategy 3 (only standard risk patients eligible). However, the different strategies had very little effect on other parameters, such as numbers of deaths and the size of the dialysis population. CONCLUSIONS: Variation in decision making from centre to centre regarding access to renal transplantation could make up to a 10% (1 year) difference in the expected half-life of renal transplants performed. Information about recipient characteristics is therefore required when making comparisons between outcome in one transplant unit with that in another, or when comparing one immunosuppressive regime with another. PMID- 11274281 TI - Renal transplantation in the elderly: a long-term, single-centre experience. AB - BACKGROUND: End-stage renal failure increases with advancing age and renal transplantation should be considered in end-stage renal failure patients older than 60 years. However, there is a paucity of data on long-term patient and graft survival in this population. METHODS: From October 1983 to March 1999, 310 renal transplantations were performed at Geneva University Hospital in 283 patients, of which 49 were done in 48 patients older than 60 years (mean age 65.6+/-4.1 years). The following data were analysed at 1, 5, and 10 years, and compared between the patients >60 years and <60 years old: actuarial patient and graft survival, serum creatinine, causes of graft loss, and patient death. RESULTS: Patient survival at 10 years was 81% for patients <60 years and 44% for patients >60 years. Graft survival at 10 years was 59% for patients <60 years and 32% for patients >60 years. Graft survival at 10 years censored for death with functioning graft was 65% for patients <60 years and 81% for patients >60 years. Main causes of mortality in the older patients were related to cardiovascular events (47%), neoplasia (41%), and sepsis (18%). Overall, recipient and donor age were not predictive factors for graft survival, as shown by multiple logistic regression. CONCLUSIONS: Renal transplantation should be considered in patients older than 60 years, since graft survival is excellent in this population. Although these patients have a shorter life expectancy, they benefit from renal transplantation similarly to younger kidney transplant recipients. PMID- 11274282 TI - Tapering off prednisolone and cyclosporin the first year after renal transplantation: the effect on glucose tolerance. AB - BACKGROUND: Glucose intolerance is an untoward side effect of some immunosuppressive and anti-hypertensive drugs. The primary aim of the present prospective observational study was to test the hypothesis that tapering off prednisolone and cyclosporin (CsA) the first year after transplantation may have beneficial effects on glucose tolerance in renal transplant recipients. METHODS: Ninety-one non-diabetic recipients were included, and 87 patients underwent a 75 g oral glucose tolerance test both 10 weeks and 1 year after renal transplantation. The change over time in 2-h blood glucose was compared with a number of variables potentially influencing glucose tolerance. RESULTS: The proportion of glucose intolerant recipients was reduced from 55 to 34% during the study. Univariate linear regression analysis showed a significant association between the reduction in daily prednisolone dose down to 5 mg and decline in blood glucose (P=0.001), whereas weight gain was associated with increasing blood glucose (P=0.031). Each 1-mg reduction of prednisolone dose leads to an estimated decline in 2-h blood glucose of 0.12 mmol/l based on the multiple linear regression model (P=0.003). Twelve out of 22 patients with post-transplant diabetes mellitus (PTDM) at baseline improved to normal or impaired glucose tolerance. Ten PTDM-subjects who remained diabetic 1 year after transplantation had lower serum insulin levels during the oral glucose challenge, and five patients treated with anti-diabetic drugs at baseline required hypoglycaemic drugs also at follow up. The decline in CsA level of 100 microg/l and the lower number of patients treated with beta-blockers at follow-up, did not alter glucose tolerance significantly. CONCLUSIONS: Tapering off prednisolone, but not CsA, significantly improves glucose tolerance during the first year after renal transplantation. PMID- 11274283 TI - A new 'online' method to measure increased exhaled isoprene in end-stage renal failure. AB - BACKGROUND: Isoprene is the most abundant hydrocarbon present in breath, and recent reports indicate that breath concentrations increase following haemodialysis. The purpose of this study was to establish whether selected ion flow tube mass spectrometry (SIFT-MS), a newly established technique in breath analysis, may be used to quantify breath isoprene in haemodialysis patients in the clinical setting. SIFT-MS is compared and contrasted with the established gas chromatography mass spectrometric technique for this purpose. METHODS: Three consecutive exhalations from 19 haemodialysis patients (12 males, seven females) undergoing a morning dialysis shift were analysed just prior to commencing treatment. Within 5 min of completing their usual dialysis regimen, using polysulphone membranes, the breath of each patient was analysed again. Additional contemporary samples were obtained from 17 normal controls. Breath isoprene was quantified using SIFT-MS, a method previously validated quantitatively using neat isoprene. RESULTS: Successful measurements of breath isoprene were obtained for each subject within 2 min, with minimum disruption to a busy dialysis environment. The coefficient of variation of triplicate measurements of breath isoprene was <10%. Prior to dialysis, the mean (+/-SD) breath isoprene concentration (138+/-63 parts per billion (ppb)) was significantly greater than for normal controls (89+/-36 ppb; P=0.016). Immediately following treatment, breath isoprene increased significantly to 184+/-95 ppb (P=0.023). CONCLUSIONS: SIFT-MS permits the accurate and rapid measurement of breath isoprene in haemodialysis patients in the clinical setting. The previously reported increase in breath isoprene following dialysis treatment is confirmed. SIFT-MS is the ideal analytical tool to investigate this phenomenon further. PMID- 11274284 TI - Guillain-Barre syndrome as presenting feature in a patient with lupus nephritis, with complete resolution after cyclophosphamide treatment. PMID- 11274285 TI - Life-threatening thrombosis 18 years after first presentation of primary antiphospholipid antibody syndrome. PMID- 11274286 TI - Renal thrombotic microangiopathy induced by interferon-alpha. PMID- 11274287 TI - Late recurrence of lupus nephritis after long-term clinical remission. PMID- 11274288 TI - A young woman with intermittent macroscopic haematuria. PMID- 11274289 TI - Peritonitis in CAPD patients--do not always use antibiotics! PMID- 11274290 TI - Cystic infection of the liver in a maintenance haemodialysis patient. PMID- 11274291 TI - Acute allograft glomerulopathy associated with CMV viraemia. PMID- 11274292 TI - The clinical role of endothelin antagonists. PMID- 11274293 TI - Hyponatraemia in a neurosurgical patient. PMID- 11274294 TI - Chronic tubulointerstitial nephritis and distal renal tubular acidosis in a patient with frusemide abuse. PMID- 11274295 TI - Hepatitis C-associated glomerulonephritis--a novel therapeutic approach. PMID- 11274296 TI - Prolonged hypocomplementaemia after post-streptococcal glomerulonephritis. PMID- 11274297 TI - Magnetic resonance angiographic patterns of renal artery stenosis in patients with chronic uraemia: differences between type 2 diabetic and non-diabetic patients. PMID- 11274298 TI - Small dose oral corticosteroid treatment rapidly improved renal function in a patient with an acute aggravation of chronic renal failure due to cholesterol embolism. PMID- 11274299 TI - Rhabdomyolysis due to simvastatin in a transplant patient: Are some statins safer than others? PMID- 11274301 TI - Homocarnosine elevations: a cause or a sign of seizure control? PMID- 11274302 TI - A letter is a letter is a letter: pure alexia for kana. PMID- 11274303 TI - Homocarnosine and seizure control in juvenile myoclonic epilepsy and complex partial seizures. AB - OBJECTIVE: To assess the relationship between seizure control and gamma aminobutyric acid (GABA), homocarnosine, and pyrrolidinone levels in the visual cortex of patients with epilepsy taking valproate or lamotrigine. Previous studies suggested that poor seizure control was associated with low GABA and homocarnosine levels. METHODS: In vivo measurements of GABA, homocarnosine, and pyrrolidinone were made in a 14-cm(3) volume of the occipital cortex using (1)H spectroscopy with a 2.1-Tesla MR spectrometer and an 8-cm surface coil. Twenty six adults (eight men) taking valproate or lamotrigine were recruited; 12 had complex partial seizures (CPS) and 14 had juvenile myoclonic epilepsy (JME). RESULTS: Median homocarnosine levels were normal for patients with JME and below normal for patients with CPS. Better seizure control was associated with higher homocarnosine levels for both groups. Median GABA was below normal for patients with JME, lower than for patients with CPS. Brain GABA was lowest in patients with JME even when seizure control was excellent. Pyrrolidinone levels were above normal in almost all patients with JME. CONCLUSIONS: Low GABA levels are associated with poor seizure control in patients with CPS, but not in JME. Higher homocarnosine levels are associated with better seizure control in both types of epilepsy. PMID- 11274304 TI - Effects of internal globus pallidus stimulation on motor cortex excitability. AB - BACKGROUND: Deep brain stimulation is a promising treatment for PD, but its physiologic effects and mechanisms of action remain poorly understood. Magnetic stimulation studies have revealed abnormalities in several different excitatory and inhibitory circuits in the motor cortex in PD. METHODS: The physiologic effects of internal globus pallidus (GPi) stimulation in seven patients with PD and seven age-matched healthy volunteers were studied. The stimulators were set at the optimal parameters (ON), at half the optimal amplitude (Half-Amp), or switched off (OFF) in random order. Patients were taking their usual medications. Magnetic stimulation was applied to the motor cortex, and motor evoked potentials (MEP) were recorded from the contralateral first dorsal interosseous muscle. Several excitatory and inhibitory pathways that have been found to be abnormal in PD were tested. RESULTS: The motor threshold (MT), MEP recruitment curve (stimulus intensities from 100 to 150% of MT), short and long interval intracortical inhibition, and intracortical facilitation were similar in the three stimulator conditions tested both at rest and during voluntary contraction. The silent period (SP) was longer in the OFF and Half-Amp conditions than in normal control subjects. In the stimulator ON condition, the SP was significantly reduced compared with the OFF condition and became similar to that in normal control subjects. CONCLUSIONS: GPi stimulation while on dopaminergic medications reduced the SP following magnetic stimulation but did not change corticospinal excitability or other measures of intracortical inhibition and facilitation. The reduction of SP may be related to the antidyskinetic and levodopa-blocking effects of ventral GPi stimulation. PMID- 11274305 TI - Seeing trees but not the forest: limited perception of large configurations in PD. AB - OBJECTIVE: To learn if Parkinson's disease (PD) is associated with a restricted attentional "floodlight." BACKGROUND: Different visual tasks may have different attentional requirements. Focused attention may be needed for some tasks; other tasks demand spatially distributed attention. Neglect after right cortical injury and dopamine depletion may limit the area over which attention can be spread. Although subjects with PD have dopamine depletion and can perform poorly on tests of visuospatial function, it is unclear if their attentional floodlight is restricted. METHODS: Eleven subjects with PD and 11 control subjects viewed different-sized letters on five printed stimulus sheets, 43 x 56 cm. On each sheet, four different large letters (14 cm2) were composed of four different medium-sized letters (2.5 cm2), which in turn were composed of four different small letters (0.4 cm2). Stimulus sheets were presented at 30- and 75-cm viewing distances. Subjects named "all the letters they could see." RESULTS: Subjects with PD named small- and medium-sized letters comparably to control subjects, but PD subjects named fewer large letters than control subjects (control = 65.68%, PD = 24.55%; group-by-letter-size interaction, p < 0.05). Subjects with PD who had undergone stereotactic pallidotomy named more letters than prepallidotomy PD subjects (p = 0.05). CONCLUSIONS: PD may affect the patient's ability to perceive large spatial configurations. As global configurations in subjects may be perceived preferentially over local patterns, it is possible that DA depletion induces an aberrant perceptual-attentional bias, such that patients have a narrowed attentional floodlight. PMID- 11274306 TI - Risk of dementia in Parkinson's disease: a community-based, prospective study. AB - OBJECTIVE: To calculate the incidence of and determine possible risk factors for dementia in PD. BACKGROUND: Dementia has important clinical consequences for patients with PD and their caregivers, but the incidence is unknown. METHODS: A population-based cohort of nondemented patients with PD (n = 171) from the county of Rogaland, Norway, was assessed at baseline and 4.2 years later with a comprehensive evaluation of motor, cognitive, and neuropsychiatric symptoms. The diagnosis of dementia was made according to the Diagnostic and Statistical Manual of Mental Disorders, 3rd edition, revised (DSM-III-R) criteria, based on interview of the patient and a caregiver, cognitive rating scales, and neuropsychologic tests. A representative sample of 3,062 nondemented elderly subjects without PD served as control group. RESULTS: Forty-three patients with PD were demented at follow-up evaluation, equivalent to an incidence rate of 95.3 per 1,000 person-years (95% CI, 68.2 to 122.0). The risk for the development of dementia in patients with PD relative to the control subjects after adjusting for age, sex, and education was 5.9 (95% CI, 3.9 to 9.1). Predictive factors at baseline for dementia in PD in addition to age were Hoehn & Yahr score >2 (OR, 3.4; 95% CI, 1.3 to 8.6) and Mini-Mental State Examination score < 29 (OR, 3.3; 95% CI, 1.3 to 8.2). CONCLUSIONS: Patients with PD have an almost sixfold increased risk for becoming demented compared with subjects without PD. PMID- 11274307 TI - Decreased glutamate + glutamine in Alzheimer's disease detected in vivo with (1)H MRS at 0.5 T. AB - OBJECTIVE: To determine whether glutamate + glutamine (GLX) levels in the brain as measured in vivo with proton MRS at 0.5 tesla (T) distinguish between probable Alzheimer's disease and normal aging. BACKGROUND: Glutamatergic markers had been measured previously in postmortem brain tissue. Conventional proton MRS at 1.5 T cannot reliably detect the GLX resonance in vivo. The authors developed a technique at 0.5 T that is sensitive to the GLX resonance. METHODS: Metabolite ratios using creatine and phosphocreatine resonance as an internal standard were acquired from the cingulate region of 18 patients with AD and 12 healthy controls. The major resonances in the spectrum were examined: N-acetylaspartate (NAA), choline-containing compounds, myo-inositol, and GLX. The Mini-Mental State Examination (MMSE) was used to assess cognitive status. The Instrumental Activities of Daily Living Scale (Instrumental ADL) was used to assess functional status. RESULTS: Reduced ratios of GLX (-10%, p = 0.001) and NAA (-12%, p = 0.000) were found in patients with AD. Increased ratios of myo-inositol in patients with AD approached significance (+14%). GLX ratios of patients with AD were correlated with MMSE (r = 0.61, p = 0.007) and Instrumental ADL (r = 0.59, p = 0.01) scores. The combined sensitivity of NAA and myo-inositol in correctly diagnosing AD was 78%. The addition of GLX to NAA and myo-inositol increased the sensitivity to 89%. Overall diagnostic accuracy improved from 80 to 83% with the addition of GLX. CONCLUSIONS: Glutamate + glutamine reduction may be a biologic marker for AD and may be a potential aid in the early clinical diagnosis of AD. PMID- 11274308 TI - Rapid initiation of gabapentin: a randomized, controlled trial. AB - OBJECTIVE: To compare the tolerability of two different dose-initiation regimens of gabapentin for the adjunctive treatment of partial seizures. BACKGROUND: Patient compliance is a key feature of successful outpatient pharmacologic therapy for epilepsy, and one aspect of compliance is simplicity of initiation. By using a rapid titration rate, leading to a rapid therapeutic gabapentin dose, perhaps there could be an improvement with compliance. METHODS: Male or female patients, at least 12 years old, with a recent history of partial seizures with or without secondary generalization, were randomized to receive gabapentin (following a blinded placebo period of an undisclosed number of days) as either a Slow initiation (300 mg day 1, 600 mg day 2, then 900 mg/day) or a Rapid initiation (900 mg/day immediately following the placebo lead-in). RESULTS: Starting gabapentin therapy at an initial therapeutic dosage of 900 mg/day is well tolerated by patients with epilepsy and is as safe as initiating with a titration schedule over 3 days. Of the four most common adverse events (somnolence, dizziness, ataxia, fatigue), only one, dizziness, occurred more often in the nontitrated (Rapid initiation) group than in the titrated (Slow initiation) group. CONCLUSION: Initiation of gabapentin at 900 mg/day is as well tolerated as is a 3-day titration, except for a higher incidence of dizziness. PMID- 11274309 TI - Homozygous deletion of the survival motor neuron 2 gene is a prognostic factor in sporadic ALS. AB - BACKGROUND: Spinal muscular atrophy (SMA) results from mutations of the survival motor neuron (SMN) gene on chromosome 5. The SMN gene exists in two highly homologous copies, telomeric (SMN1) and centromeric (SMN2). SMA is caused by mutations in SMN1 but not SMN2. The clinical phenotype of SMA appears to be related to the expression of SMN2. Patients suffering from the milder forms of SMA carry more copies of the SMN2 gene compared with patients with more severe SMA. It is suggested that the SMN2 gene is translated into an at least partially functional protein that protects against loss of motor neurons. OBJECTIVE: To investigate whether genetic mechanisms implicated in motor neuron death in SMA have a role in ALS. METHODS: The presence of deletions of exons 7 and 8 of SMN1 and SMN2 was determined in 110 patients with sporadic ALS and compared with 100 unaffected controls. RESULTS: The presence of a homozygous SMN2 deletion was overrepresented in patients with ALS compared with controls (16% versus 4%; OR, 4.4; 95% CI, 1.4 to 13.5). Patients with a homozygous SMN2 deletion had a shorter median time of survival (p < 0.009). Furthermore, multivariate regression analysis showed that the presence of an SMN2 deletion was independently associated with survival time (p < 0.02). No homozygous deletions in SMN1 were found. Carrier status of SMA appeared to be equally present in patients and controls (1 in 20). CONCLUSION: These results indicate that, similar to SMA, the SMN2 gene can act as a prognostic factor and may therefore be a phenotypic modifier in sporadic ALS. Increasing the expression of the SMN2 gene may provide a strategy for treatment of motor neuron disease. PMID- 11274310 TI - Hospitalization in amyotrophic lateral sclerosis: causes, costs, and outcomes. AB - OBJECTIVE: As ALS progresses, extensive supportive care is required, including multidisciplinary outpatient care and hospitalization. The authors studied the causes, health care utilization, and outcomes for hospitalized patients with ALS. METHODS: With use of the 1996 Nationwide Inpatient Sample, an administrative database representing 20% of U.S. hospitals, 1,600 hospitalizations in patients with ALS were identified and compared with 5,364,728 non-ALS hospitalizations. RESULTS: The most common concurrent diagnoses in patients with ALS were dehydration and malnutrition (574 patients, 36%), pneumonia (507 patients, 32%), and respiratory failure (398 patients, 25%). Only 38% of patients with ALS were discharged to home without home health care compared with 73% of patients with non-ALS. Fifteen percent of patients with ALS died in the hospital compared with 3% of non-ALS patients. The average length of hospital stay and charges were greater for patients with ALS than for non-ALS patients (8.4 days and $19,810 for ALS patients and 5.4 days and $11,924 for non-ALS patients). Mortality was significantly associated with emergency room admission (versus nonemergency admission; OR = 1.60), increasing age (per year; OR = 1.03), respiratory failure (OR = 3.37), and pneumonia (OR = 2.02) (p < 0.01 for all comparisons). CONCLUSIONS: Patients with ALS have lengthy and costly hospital admissions, a high in-hospital mortality rate, and few routine discharges. Recognition of the issues that precipitate hospitalization may allow development of preventive strategies. PMID- 11274311 TI - Preceding infections, immune factors, and outcome in Guillain-Barre syndrome. AB - OBJECTIVE: To test the hypothesis that different preceding infections influence the neurophysiologic classification and clinical features of Guillain-Barre syndrome (GBS). METHODS: We tested pretreatment sera, 7 +/- 3 (mean +/- SD) days from onset, from 229 patients with GBS in a multicenter trial of plasma exchange and immunoglobulin, for serological markers of infection, adhesion molecules, and cytokine receptors, and compared these with neurophysiologic and clinical features. RESULTS: Recent infection by Campylobacter jejuni was found in 53 patients (23%), cytomegalovirus in 19 (8%), and Epstein-Barr virus in four (2%). Patients with C. jejuni infection were more likely than others to have neurophysiologic criteria of axonal neuropathy or inexcitable nerves, antiganglioside GM(1) antibodies, pure motor GBS, lower CSF protein, and worse outcome. Patients with cytomegalovirus infection were younger and more likely than others to have raised serum concentrations of molecules important in T lymphocyte activation and migration, soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular cell adhesion molecule-1 (sVCAM-1), soluble leukocyte selectin, and soluble interleukin-2 receptor (sIL-2R). Concentrations of sICAM-1 and soluble tumor necrosis factor receptor were higher in patients with inexcitable nerves than those with demyelinating neurophysiology. Logistic regression analysis showed death or inability to walk unaided at 48 weeks were associated with diarrhea, inexcitable nerves, severe arm weakness, age over 50, raised sIL-2R concentration and absence of immunoglobulin (Ig) M antiganglioside GM(1) antibodies. CONCLUSIONS: Subtypes of GBS defined by preceding infections were only approximately associated with different patterns of clinical, neurophysiologic, and immunologic features. A single infectious agent caused more than one type of pathology in GBS, implying interaction with additional host factors. Most patients had no identified infection. PMID- 11274312 TI - Withdrawal of support in intracerebral hemorrhage may lead to self-fulfilling prophecies. AB - BACKGROUND: Withdrawal of support in patients with severe brain injury invariably leads to death. Preconceived notions about futility of care in patients with intracerebral hemorrhage (ICH) may prompt withdrawal of support, and modeling outcome in patient populations in whom withdrawal of support occurs may lead to self-fulfilling prophecies. METHODS: Subjects included consecutive patients with supratentorial ICH. Radiographic characteristics of the hemorrhage, clinical variables, and neurologic outcome were assessed. Attitudes about futility of care were examined among members of the departments of neurology and neurologic surgery through a written survey and case presentations. RESULTS: There were 87 patients with supratentorial ICH; overall mortality was 34.5% (30/87). Mortality was 66.7% (18/27) in patients with Glasgow Coma Score < or = 8 and ICH volume > 60 cm(3). Medical support was withdrawn in 76.7% (23/30) of patients who died. Inclusion of a variable to account for the withdrawal of support in a model predicting outcome negated the predictive value of all other variables. Patients undergoing surgical decompression were unlikely to have support withdrawn, and surgery was less likely to be performed in older patients (p < 0.01) and patients with left hemispheric hemorrhage (p = 0.04). Survey results suggested that practitioners tend to be overly pessimistic in prognosticating outcome based upon data available at the time of presentation. CONCLUSIONS: The most important prognostic variable in determining outcome after ICH is the level of medical support provided. Withdrawal of support in patients felt likely to have a "poor outcome" biases predictive models and leads to self-fulfilling prophecies. Our data show that individual patients in traditionally "poor outcome" categories can have a reasonable neurologic outcome when treated aggressively. PMID- 11274313 TI - Recurrent brain hemorrhage is more frequent than ischemic stroke after intracranial hemorrhage. AB - OBJECTIVE: To characterize the rates of recurrent intracranial hemorrhage (ICH), ischemic stroke, and death in survivors of primary ICH. METHODS: Systematic review of studies reporting recurrent stroke in survivors of primary ICH, identified at index ICH and followed forward. Studies were identified by computerized search of the literature and review of reference lists. RESULTS: Ten studies published between 1982 and 2000 reporting 1,880 survivors of ICH, followed for a total of 6,326 patient-years (mean follow-up, 3.4 patient-years), were included. The aggregate rate of all stroke from five studies was 4.3% per patient-year (95% CI, 3.5% to 5.4%). The rate in the three population-based studies was higher than in the two hospital-based studies, 6.2% versus 4.0% per patient-year (p = 0.04). About three fourths of recurrent strokes were ICH. Considering all 10 studies, a total of 147 patients had a recurrent ICH, an aggregate rate of 2.3% per patient-year (95% CI, 1.9% to 2.7%). Based on data from four studies, patients with a primary lobar ICH had a higher rate of recurrent ICH than those with a deep, hemispheric ICH (4.4% versus 2.1% per patient-year; p = 0.002). The aggregate rates of subsequent ischemic stroke and mortality were 1.1% per patient-year (95% CI, 0.8% to 1.7%) and 8.8% per patient year (95% CI, 5.2% to 11.0%). CONCLUSIONS: Recurrent stroke among survivors of primary ICH occurs at a rate of about 4% per patient-year, and most are recurrent ICH. Survivors of ICH have a higher risk of recurrent ICH than of ischemic stroke, and this has implications for the use of antithrombotic agents in these patients. PMID- 11274314 TI - Primary cerebral amyloidoma. PMID- 11274315 TI - Pure alexia from a posterior occipital lesion. AB - The authors report a patient with pure alexia (letter-by-letter reading) selectively impaired for kana (Japanese phonograms), cerebral achromatopsia, and right lower quadrantanopsia after hemorrhage in the left posterior occipital lobe, mainly under the lateral occipital gyri. The patient also could not recognize some single-character kana, nor could he discriminate between two shapes of a similar size. The authors believe that the posterior occipital lobe, including the lateral occipital gyri, is specialized to recognize kana characters in this patient. PMID- 11274316 TI - Preliminary evidence of widespread morphological variations of the brain in dyslexia. AB - The MR images of 16 men with dyslexia and 14 control subjects were compared using a voxel-based analysis. Evidence of decreases in gray matter in dyslexic subjects, most notably in the left temporal lobe and bilaterally in the temporoparietooccipital juncture, but also in the frontal lobe, caudate, thalamus, and cerebellum, was found. Widely distributed morphologic differences affecting several brain regions may contribute to the deficits associated with dyslexia. PMID- 11274317 TI - Recurrence risks to sibs of MS index cases: impact of consanguineous matings. AB - Using population-based data, the authors identified 24 MS index cases whose parents were related. Twenty-two had 67 sibs of whom 6 also had MS, yielding a recurrence risk of approximately 9%, significantly higher than for sibs of MS index cases from nonconsanguineous parents. These findings support the concept that multiple interacting genes increase the risk of MS. PMID- 11274319 TI - How do AD patients and their caregivers decide whether to enroll in a clinical trial? AB - To examine how patients and caregivers decide whether to enroll in a clinical trial, the authors conducted semi-structured interviews with 22 family caregivers of patients with mild to moderate AD who were recruited for a clinical trial. They found that a caregiver who enrolls a patient in research generally involves the patient in the decision-making process, reports that the patient shares in the decision, and regards the risks and benefits to the patient and to the caregiver as interdependent. PMID- 11274318 TI - Evidence for neuroaxonal injury in patients with proteolipid protein gene mutations. AB - The authors used proton MRS to investigate neuropathologic correlates in nine patients with proteolipid protein (PLP) gene mutations who did not show cerebral atrophy on cranial MRI. When compared with 16 age-matched control participants, patients with PLP mutations had significant and widespread decreased brain N acetyl aspartate, a neuronal marker. The authors conclude that PLP mutations cause neuroaxonal injury, which in turn contributes to the neurologic deficit observed in these patients. PMID- 11274320 TI - Cerebral air embolism complicating cardiac ablation procedures. AB - Two patients with similar courses of neurologic impairment and subsequent recovery after cerebral air embolism complicating cardiac ablation procedures are described. Hyperbaric oxygen therapy, combined with aggressive resuscitative efforts, appears to have contributed to each patient's recovery. PMID- 11274321 TI - Modafinil for excessive daytime sleepiness in myotonic dystrophy. AB - The authors conducted an open-label trial of modafinil for excessive daytime sleepiness in myotonic dystrophy. Eleven patients were evaluated: two were not treated because of obstructive sleep apnea, and nine received 200 to 400 mg modafinil/day for an average of 16.4 weeks. There were no major side effects. Average sleep latency as measured by the Multiple Sleep Latency Test increased from 7.3 to 22.7 minutes ( p = 0.00013), and average Epworth Sleepiness Scale score decreased from 13.25 to 7.75 (p = 0.01028). Modafinil shows evidence of effectiveness for excessive daytime somnolence in myotonic dystrophy and should be investigated further. PMID- 11274322 TI - Relative pupil-sparing third nerve palsy: etiology and clinical variables predictive of a mass. AB - The causes of ophthalmoplegia in 24 consecutive patients with neurologically isolated, relative pupil-sparing third nerve palsy included infarction in 10 patients, compression by tumors or aneurysms in 10 patients, and miscellaneous disorders in four patients. There was no significant difference in the proportion of patients with pain, degree of external ophthalmoplegia, or degree of internal ophthalmoplegia between the groups with infarction or mass lesions. Therefore, screening for mass lesions using neuroimaging is indicated in patients with this presentation of third nerve palsy. PMID- 11274323 TI - Outcomes following staged bilateral pallidotomy in advanced Parkinson's disease. AB - The authors assessed clinical outcome for up to one year after staged bilateral pallidotomy in 14 patients with advanced PD. One year after surgery, dyskinesias were virtually abolished and there were significant reductions in "off" time (67%) and activities of daily living "off" scores (24%), as well as nonsignificant reduction in "off" motor score (39%); "on" scores were unchanged. One patient developed a visual field deficit; two had transient confusion. Staged bilateral pallidotomy improves motor function in selected patients with advanced PD. PMID- 11274324 TI - Mitochondrial myopathy, parkinsonism, and multiple mtDNA deletions in a Sephardic Jewish family. AB - The authors describe a family of Sephardic Jews with progressive external ophthalmoparesis, skeletal muscle weakness, and parkinsonism. Autosomal recessive inheritance was suggested by many consanguineous marriages, although a dominant disorder could not be excluded. No linkage to known progressive external ophthalmoparesis locus was found. The presence of cytochrome c oxidase-negative ragged-red fibers, biochemically reduced respiratory chain complexes, and multiple mitochondrial DNA deletions in muscle biopsies from four patients suggested a new mitochondrial disorder of intergenomic communication. PMID- 11274325 TI - The course of tardive dyskinesia and parkinsonism in psychiatric inpatients: 14 year follow-up. AB - Tardive dyskinesia and parkinsonism were assessed in 53 patients residing in a state psychiatric hospital in 1984 and 1998. A 4.0-point decrease in the mean Abnormal Involuntary Movement Scale score (6.0 versus 2.0; p < 0.001) and a 3.5 point increase in the Rating Scale for Extrapyramidal Signs score (2.8 versus 6.3; p < 0.001) were noted between 1984 and 1998. Over a 14-year period, tardive dyskinesia improved and parkinsonism worsened in patients who continued to receive neuroleptic drugs. PMID- 11274326 TI - Circulating levels of MMP-1, -2, -3, -9, and TIMP-1 are increased in POEMS syndrome. AB - The authors quantitatively measured levels of matrix metalloproteinases (MMP), tissue inhibitor of metalloproteinases (TIMP), and vascular endothelial growth factor (VEGF) in blood samples of POEMS syndrome. Circulating levels of MMP-1, 2, -3, -9, and TIMP-1 were more increased in patients with POEMS syndrome than in patients with other neurologic disorders or in healthy controls. Serum levels of VEGF and TIMP-1 were strongly correlated with each other. Increased circulating levels of MMP-1, -2, -3, -9, and TIMP-1 may lead to a better understanding the pathogenesis of POEMS syndrome. PMID- 11274327 TI - Neurologic manifestations of disseminated infection with Mycobacterium abscessus. AB - Mycobacterium abscessus is a ubiquitous, saprophytic organism with low pathogenic potential. The authors describe the previously unreported clinical features of meningitis and native valve endocarditis caused by this rapidly growing atypical mycobacterium. The fatal outcome of this unusual case coincides with the grim prognosis of this disseminated infection and the significant mortality rate associated with neurologic complications of infective endocarditis. PMID- 11274328 TI - Rheumatoid disease of the CNS with meningeal vasculitis presenting with a seizure. PMID- 11274329 TI - Encephalitis associated with glutamic acid decarboxylase autoantibodies. PMID- 11274330 TI - Serial MR angiography and contrast-enhanced MRI in chickenpox-associated stroke. PMID- 11274331 TI - Insulinoma presenting as seizure disorder. PMID- 11274332 TI - Solitary plasmacytoma with VEGF overproduction: report of a patient with polyneuropathy. PMID- 11274333 TI - Transplantation of cultured human neuronal cells for patients with stroke. PMID- 11274334 TI - MRI evidence of mesial temporal sclerosis in patients with psychogenic nonepileptic seizures. PMID- 11274335 TI - Usefulness of MRI measures of entorhinal cortex versus hippocampus in AD. PMID- 11274338 TI - Membrane transport in the malaria-infected erythrocyte. AB - The malaria parasite is a unicellular eukaryotic organism which, during the course of its complex life cycle, invades the red blood cells of its vertebrate host. As it grows and multiplies within its host blood cell, the parasite modifies the membrane permeability and cytosolic composition of the host cell. The intracellular parasite is enclosed within a so-called parasitophorous vacuolar membrane, tubular extensions of which radiate out into the host cell compartment. Like all eukaryote cells, the parasite has at its surface a plasma membrane, as well as having a variety of internal membrane-bound organelles that perform a range of functions. This review focuses on the transport properties of the different membranes of the malaria-infected erythrocyte, as well as on the role played by the various membrane transport systems in the uptake of solutes from the extracellular medium, the disposal of metabolic wastes, and the origin and maintenance of electrochemical ion gradients. Such systems are of considerable interest from the point of view of antimalarial chemotherapy, both as drug targets in their own right and as routes for targeting cytotoxic agents into the intracellular parasite. PMID- 11274339 TI - Proprioception from a spinocerebellar perspective. AB - This review explores how proprioceptive sensory information is organized at spinal cord levels as it relates to a sense of body position and movement. The topic is considered in an historical context and develops a different framework that may be more in tune with current views of sensorimotor processing in other central nervous system structures. The dorsal spinocerebellar tract (DSCT) system is considered in detail as a model system that may be considered as an end point for the processing of proprioceptive sensory information in the spinal cord. An analysis of this system examines sensory processing at the lowest levels of synaptic connectivity with central neurons in the nervous system. The analysis leads to a framework for proprioception that involves a highly flexible network organization based in some way on whole limb kinematics. The functional organization underlying this framework originates with the biomechanical linkages in the limb that establish functional relationships among the limb segments. Afferent information from limb receptors is processed further through a distributed neural network in the spinal cord. The result is a global representation of hindlimb parameters rather than a muscle-by-muscle or joint-by joint representation. PMID- 11274340 TI - Nonvertebrate hemoglobins: functions and molecular adaptations. AB - Hemoglobin (Hb) occurs in all the kingdoms of living organisms. Its distribution is episodic among the nonvertebrate groups in contrast to vertebrates. Nonvertebrate Hbs range from single-chain globins found in bacteria, algae, protozoa, and plants to large, multisubunit, multidomain Hbs found in nematodes, molluscs and crustaceans, and the giant annelid and vestimentiferan Hbs comprised of globin and nonglobin subunits. Chimeric hemoglobins have been found recently in bacteria and fungi. Hb occurs intracellularly in specific tissues and in circulating red blood cells (RBCs) and freely dissolved in various body fluids. In addition to transporting and storing O(2) and facilitating its diffusion, several novel Hb functions have emerged, including control of nitric oxide (NO) levels in microorganisms, use of NO to control the level of O(2) in nematodes, binding and transport of sulfide in endosymbiont-harboring species and protection against sulfide, scavenging of O(2 )in symbiotic leguminous plants, O(2 )sensing in bacteria and archaebacteria, and dehaloperoxidase activity useful in detoxification of chlorinated materials. This review focuses on the extensive variation in the functional properties of nonvertebrate Hbs, their O(2 )binding affinities, their homotropic interactions (cooperativity), and the sensitivities of these parameters to temperature and heterotropic effectors such as protons and cations. Whenever possible, it attempts to relate the ligand binding properties to the known molecular structures. The divergent and convergent evolutionary trends evident in the structures and functions of nonvertebrate Hbs appear to be adaptive in extending the inhabitable environment available to Hb-containing organisms. PMID- 11274341 TI - The oxytocin receptor system: structure, function, and regulation. AB - The neurohypophysial peptide oxytocin (OT) and OT-like hormones facilitate reproduction in all vertebrates at several levels. The major site of OT gene expression is the magnocellular neurons of the hypothalamic paraventricular and supraoptic nuclei. In response to a variety of stimuli such as suckling, parturition, or certain kinds of stress, the processed OT peptide is released from the posterior pituitary into the systemic circulation. Such stimuli also lead to an intranuclear release of OT. Moreover, oxytocinergic neurons display widespread projections throughout the central nervous system. However, OT is also synthesized in peripheral tissues, e.g., uterus, placenta, amnion, corpus luteum, testis, and heart. The OT receptor is a typical class I G protein-coupled receptor that is primarily coupled via G(q) proteins to phospholipase C-beta. The high-affinity receptor state requires both Mg(2+) and cholesterol, which probably function as allosteric modulators. The agonist-binding region of the receptor has been characterized by mutagenesis and molecular modeling and is different from the antagonist binding site. The function and physiological regulation of the OT system is strongly steroid dependent. However, this is, unexpectedly, only partially reflected by the promoter sequences in the OT receptor gene. The classical actions of OT are stimulation of uterine smooth muscle contraction during labor and milk ejection during lactation. While the essential role of OT for the milk let-down reflex has been confirmed in OT-deficient mice, OT's role in parturition is obviously more complex. Before the onset of labor, uterine sensitivity to OT markedly increases concomitant with a strong upregulation of OT receptors in the myometrium and, to a lesser extent, in the decidua where OT stimulates the release of PGF(2 alpha). Experiments with transgenic mice suggest that OT acts as a luteotrophic hormone opposing the luteolytic action of PGF(2 alpha). Thus, to initiate labor, it might be essential to generate sufficient PGF(2 alpha) to overcome the luteotrophic action of OT in late gestation. OT also plays an important role in many other reproduction-related functions, such as control of the estrous cycle length, follicle luteinization in the ovary, and ovarian steroidogenesis. In the male, OT is a potent stimulator of spontaneous erections in rats and is involved in ejaculation. OT receptors have also been identified in other tissues, including the kidney, heart, thymus, pancreas, and adipocytes. For example, in the rat, OT is a cardiovascular hormone acting in concert with atrial natriuretic peptide to induce natriuresis and kaliuresis. The central actions of OT range from the modulation of the neuroendocrine reflexes to the establishment of complex social and bonding behaviors related to the reproduction and care of the offspring. OT exerts potent antistress effects that may facilitate pair bonds. Overall, the regulation by gonadal and adrenal steroids is one of the most remarkable features of the OT system and is, unfortunately, the least understood. One has to conclude that the physiological regulation of the OT system will remain puzzling as long as the molecular mechanisms of genomic and nongenomic actions of steroids have not been clarified. PMID- 11274342 TI - Molecular basis of mechanotransduction in living cells. AB - The simplest cell-like structure, the lipid bilayer vesicle, can respond to mechanical deformation by elastic membrane dilation/thinning and curvature changes. When a protein is inserted in the lipid bilayer, an energetic cost may arise because of hydrophobic mismatch between the protein and bilayer. Localized changes in bilayer thickness and curvature may compensate for this mismatch. The peptides alamethicin and gramicidin and the bacterial membrane protein MscL form mechanically gated (MG) channels when inserted in lipid bilayers. Their mechanosensitivity may arise because channel opening is associated with a change in the protein's membrane-occupied area, its hydrophobic mismatch with the bilayer, excluded water volume, or a combination of these effects. As a consequence, bilayer dilation/thinning or changes in local membrane curvature may shift the equilibrium between channel conformations. Recent evidence indicates that MG channels in specific animal cell types (e.g., Xenopus oocytes) are also gated directly by bilayer tension. However, animal cells lack the rigid cell wall that protects bacteria and plants cells from excessive expansion of their bilayer. Instead, a cortical cytoskeleton (CSK) provides a structural framework that allows the animal cell to maintain a stable excess membrane area (i.e., for its volume occupied by a sphere) in the form of membrane folds, ruffles, and microvilli. This excess membrane provides an immediate membrane reserve that may protect the bilayer from sudden changes in bilayer tension. Contractile elements within the CSK may locally slacken or tighten bilayer tension to regulate mechanosensitivity, whereas membrane blebbing and tight seal patch formation, by using up membrane reserves, may increase membrane mechanosensitivity. In specific cases, extracellular and/or CSK proteins (i.e., tethers) may transmit mechanical forces to the process (e.g., hair cell MG channels, MS intracellular Ca(2+) release, and transmitter release) without increasing tension in the lipid bilayer. PMID- 11274343 TI - Alzheimer's disease: genes, proteins, and therapy. AB - Rapid progress in deciphering the biological mechanism of Alzheimer's disease (AD) has arisen from the application of molecular and cell biology to this complex disorder of the limbic and association cortices. In turn, new insights into fundamental aspects of protein biology have resulted from research on the disease. This beneficial interplay between basic and applied cell biology is well illustrated by advances in understanding the genotype-to-phenotype relationships of familial Alzheimer's disease. All four genes definitively linked to inherited forms of the disease to date have been shown to increase the production and/or deposition of amyloid beta-protein in the brain. In particular, evidence that the presenilin proteins, mutations in which cause the most aggressive form of inherited AD, lead to altered intramembranous cleavage of the beta-amyloid precursor protein by the protease called gamma-secretase has spurred progress toward novel therapeutics. The finding that presenilin itself may be the long sought gamma-secretase, coupled with the recent identification of beta-secretase, has provided discrete biochemical targets for drug screening and development. Alternate and novel strategies for inhibiting the early mechanism of the disease are also emerging. The progress reviewed here, coupled with better ability to diagnose the disease early, bode well for the successful development of therapeutic and preventative drugs for this major public health problem. PMID- 11274344 TI - Adenosine 5'-triphosphate: a P2-purinergic agonist in the myocardium. AB - ATP, besides an intracellular energy source, is an agonist when applied to a variety of different cells including cardiomyocytes. Sources of ATP in the extracellular milieu are multiple. Extracellular ATP is rapidly degraded by ectonucleotidases. Today ionotropic P2X(1--7) receptors and metabotropic P2Y(1,2,4,6,11) receptors have been cloned and their mRNA found in cardiomyocytes. On a single cardiomyocyte, micromolar ATP induces nonspecific cationic and Cl(-) currents that depolarize the cells. ATP both increases directly via a G(s) protein and decreases Ca(2+) current. ATP activates the inward-rectifying currents (ACh- and ATP-activated K(+) currents) and outward K(+) currents. P2-purinergic stimulation increases cAMP by activating adenylyl cyclase isoform V. It also involves tyrosine kinases to activate phospholipase C gamma to produce inositol 1,4,5-trisphosphate and Cl(-)/HCO(3)(-) exchange to induce a large transient acidosis. No clear correlation is presently possible between an effect and the activation of a given P2-receptor subtype in cardiomyocytes. ATP itself is generally a positive inotropic agent. Upon rapid application to cells, ATP induces various forms of arrhythmia. At the tissue level, arrhythmia could be due to slowing of electrical spread after both Na(+) current decrease and cell-to-cell uncoupling as well as cell depolarization and Ca(2+) current increase. In as much as the information is available, this review also reports analog effects of UTP and diadenosine polyphosphates. PMID- 11274345 TI - Mammalian mitogen-activated protein kinase signal transduction pathways activated by stress and inflammation. AB - The molecular details of mammalian stress-activated signal transduction pathways have only begun to be dissected. This, despite the fact that the impact of these pathways on the pathology of chronic inflammation, heart disease, stroke, the debilitating effects of diabetes mellitus, and the side effects of cancer therapy, not to mention embryonic development, innate and acquired immunity, is profound. Cardiovascular disease and diabetes alone represent the most significant health care problems in the developed world. Thus it is not surprising that understanding these pathways has attracted wide interest, and in the past 10 years, dramatic progress has been made. Accordingly, it is now becoming possible to envisage the transition of these findings to the development of novel treatment strategies. This review focuses on the biochemical components and regulation of mammalian stress-regulated mitogen-activated protein kinase (MAPK) pathways. The nuclear factor-kappa B pathway, a second stress signaling paradigm, has been the subject of several excellent recent reviews (258, 260). PMID- 11274346 TI - Biology of oligodendrocyte and myelin in the mammalian central nervous system. AB - Oligodendrocytes, the myelin-forming cells of the central nervous system (CNS), and astrocytes constitute macroglia. This review deals with the recent progress related to the origin and differentiation of the oligodendrocytes, their relationships to other neural cells, and functional neuroglial interactions under physiological conditions and in demyelinating diseases. One of the problems in studies of the CNS is to find components, i.e., markers, for the identification of the different cells, in intact tissues or cultures. In recent years, specific biochemical, immunological, and molecular markers have been identified. Many components specific to differentiating oligodendrocytes and to myelin are now available to aid their study. Transgenic mice and spontaneous mutants have led to a better understanding of the targets of specific dys- or demyelinating diseases. The best examples are the studies concerning the effects of the mutations affecting the most abundant protein in the central nervous myelin, the proteolipid protein, which lead to dysmyelinating diseases in animals and human (jimpy mutation and Pelizaeus-Merzbacher disease or spastic paraplegia, respectively). Oligodendrocytes, as astrocytes, are able to respond to changes in the cellular and extracellular environment, possibly in relation to a glial network. There is also a remarkable plasticity of the oligodendrocyte lineage, even in the adult with a certain potentiality for myelin repair after experimental demyelination or human diseases. PMID- 11274348 TI - Behavior of several two-dimensional fluid equations in singular scenarios. AB - We give conditions that rule out formation of sharp fronts for certain two dimensional incompressible flows. We show that a necessary condition of having a sharp front is that the flow has to have uncontrolled velocity growth. In the case of the quasi-geostrophic equation and two-dimensional Euler equation, we obtain estimates on the formation of semi-uniform fronts. PMID- 11274347 TI - Brain stem control of swallowing: neuronal network and cellular mechanisms. AB - Swallowing movements are produced by a central pattern generator located in the medulla oblongata. It has been established on the basis of microelectrode recordings that the swallowing network includes two main groups of neurons. One group is located within the dorsal medulla and contains the generator neurons involved in triggering, shaping, and timing the sequential or rhythmic swallowing pattern. Interestingly, these generator neurons are situated within a primary sensory relay, that is, the nucleus tractus solitarii. The second group is located in the ventrolateral medulla and contains switching neurons, which distribute the swallowing drive to the various pools of motoneurons involved in swallowing. This review focuses on the brain stem mechanisms underlying the generation of sequential and rhythmic swallowing movements. It analyzes the neuronal circuitry, the cellular properties of neurons, and the neurotransmitters possibly involved, as well as the peripheral and central inputs which shape the output of the network appropriately so that the swallowing movements correspond to the bolus to be swallowed. The mechanisms possibly involved in pattern generation and the possible flexibility of the swallowing central pattern generator are discussed. PMID- 11274349 TI - Effect of nicotine on brain activation during performance of a working memory task. AB - Nicotine influences cognition and behavior, but the mechanisms by which these effects occur are unclear. By using positron emission tomography, we measured cognitive activation (increases in relative regional cerebral blood flow) during a working memory task [2-back task (2BT)] in 11 abstinent smokers and 11 ex smokers. Assays were performed both after administration of placebo gum and 4-mg nicotine gum. Performance on the 2BT did not differ between groups in either condition, and the pattern of brain activation by the 2BT was consistent with reports in the literature. However, in the placebo condition, activation in ex smokers predominated in the left hemisphere, whereas in smokers, it occurred in the right hemisphere. When nicotine was administered, activation was reduced in smokers but enhanced in ex-smokers. The lateralization of activation as a function of nicotine dependence suggests that chronic exposure to nicotine or withdrawal from nicotine affects cognitive strategies used to perform the memory task. Furthermore, the lack of enhancement of activation after nicotine administration in smokers likely reflects tolerance. PMID- 11274351 TI - A positivity result in the theory of Macdonald polynomials. AB - We outline here a proof that a certain rational function C(n)(q, t), which has come to be known as the "q, t-Catalan," is in fact a polynomial with positive integer coefficients. This has been an open problem since 1994. Because C(n)(q, t) evaluates to the Catalan number at t = q = 1, it has also been an open problem to find a pair of statistics a, b on the collection (n) of Dyck paths Pi of length 2n yielding C(n)(q, t) = summation operator(pi) t(a(Pi))q(b(Pi)). Our proof is based on a recursion for C(n)(q, t) suggested by a pair of statistics recently proposed by J. Haglund. One of the byproducts of our results is a proof of the validity of Haglund's conjecture. PMID- 11274350 TI - A strategy for the identification of proteins targeted by thioredoxin. AB - Thioredoxins are 12-kDa proteins functional in the regulation of cellular processes throughout the animal, plant, and microbial kingdoms. Growing evidence with seeds suggests that an h-type of thioredoxin, reduced by NADPH via NADP thioredoxin reductase, reduces disulfide bonds of target proteins and thereby acts as a wakeup call in germination. A better understanding of the role of thioredoxin in seeds as well as other systems could be achieved if more were known about the target proteins. To this end, we have devised a strategy for the comprehensive identification of proteins targeted by thioredoxin. Tissue extracts incubated with reduced thioredoxin are treated with a fluorescent probe (monobromobimane) to label sulfhydryl groups. The newly labeled proteins are isolated by conventional two-dimensional electrophoresis: (i) nonreducing/reducing or (ii) isoelectric focusing/reducing SDS/PAGE. The isolated proteins are identified by amino acid sequencing. Each electrophoresis system offers an advantage: the first method reveals the specificity of thioredoxin in the reduction of intramolecular vs. intermolecular disulfide bonds, whereas the second method improves the separation of the labeled proteins. By application of both methods to peanut seed extracts, we isolated at least 20 thioredoxin targets and identified 5-three allergens (Ara h2, Ara h3, and Ara h6) and two proteins not known to occur in peanut (desiccation-related and seed maturation protein). These findings open the door to the identification of proteins targeted by thioredoxin in a wide range of systems, thereby enhancing our understanding of its function and extending its technological and medical applications. PMID- 11274352 TI - Directed evolution of polymerase function by compartmentalized self-replication. AB - We describe compartmentalized self-replication (CSR), a strategy for the directed evolution of enzymes, especially polymerases. CSR is based on a simple feedback loop consisting of a polymerase that replicates only its own encoding gene. Compartmentalization serves to isolate individual self-replication reactions from each other. In such a system, adaptive gains directly (and proportionally) translate into genetic amplification of the encoding gene. CSR has applications in the evolution of polymerases with novel and useful properties. By using three cycles of CSR, we obtained variants of Taq DNA polymerase with 11-fold higher thermostability than the wild-type enzyme or with a >130-fold increased resistance to the potent inhibitor heparin. Insertion of an extra stage into the CSR cycle before the polymerase reaction allows its application to enzymes other than polymerases. We show that nucleoside diphosphate kinase and Taq polymerase can form such a cooperative CSR cycle based on reciprocal catalysis, whereby nucleoside diphosphate kinase produces the substrates required for the replication of its own gene. We also find that in CSR the polymerase genes themselves evolve toward more efficient replication. Thus, polymerase genes and their encoded polypeptides cooperate to maximize postselection copy number. CSR should prove useful for the directed evolution of enzymes, particularly DNA or RNA polymerases, as well as for the design and study of in vitro self-replicating systems mimicking prebiotic evolution and viral replication. PMID- 11274353 TI - Protein folding from a highly disordered denatured state: the folding pathway of chymotrypsin inhibitor 2 at atomic resolution. AB - Previous experimental and theoretical studies have produced high-resolution descriptions of the native and folding transition states of chymotrypsin inhibitor 2 (CI2). In similar fashion, here we use a combination of NMR experiments and molecular dynamics simulations to examine the conformations populated by CI2 in the denatured state. The denatured state is highly unfolded, but there is some residual native helical structure along with hydrophobic clustering in the center of the chain. The lack of persistent nonnative structure in the denatured state reduces barriers that must be overcome, leading to fast folding through a nucleation-condensation mechanism. With the characterization of the denatured state, we have now completed our description of the folding/unfolding pathway of CI2 at atomic resolution. PMID- 11274354 TI - A role for intermolecular disulfide bonds in prion diseases? AB - The key event in prion diseases seems to be the conversion of the prion protein PrP from its normal cellular isoform (PrP(C)) to an aberrant "scrapie" isoform (PrP(Sc)). Earlier studies have detected no covalent modification in the scrapie isoform and have concluded that the PrP(C) --> PrP(Sc) conversion is a purely conformational transition involving no chemical reactions. However, a reexamination of the available biochemical data suggests that the PrP(C) --> PrP(Sc) conversion also involves a covalent reaction of the (sole) intramolecular disulfide bond of PrP(C). Specifically, the data are consistent with the hypothesis that infectious prions are composed of PrP(Sc) polymers linked by intermolecular disulfide bonds. Thus, the PrP(C) --> PrP(Sc) conversion may involve not only a conformational transition but also a thiol/disulfide exchange reaction between the terminal thiolate of such a PrP(Sc) polymer and the disulfide bond of a PrP(C) monomer. This hypothesis seems to account for several unusual features of prion diseases. PMID- 11274355 TI - Galactosides in the rhizosphere: utilization by Sinorhizobium meliloti and development of a biosensor. AB - Identifying the types and distributions of organic substrates that support microbial activities around plant roots is essential for a full understanding of plant-microbe interactions and rhizosphere ecology. We have constructed a strain of the soil bacterium Sinorhizobium meliloti containing a gfp gene fused to the melA promoter which is induced on exposure to galactose and galactosides. We used the fusion strain as a biosensor to determine that galactosides are released from the seeds of several different legume species during germination and are also released from roots of alfalfa seedlings growing on artificial medium. Galactoside presence in seed wash and sterile root washes was confirmed by HPLC. Experiments examining microbial growth on alpha-galactosides in seed wash suggested that alpha-galactoside utilization could play an important role in supporting growth of S. meliloti near germinating seeds of alfalfa. When inoculated into microcosms containing legumes or grasses, the biosensor allowed us to visualize the localized presence of galactosides on and around roots in unsterilized soil, as well as the grazing of fluorescent bacteria by protozoa. Galactosides were present in patches around zones of lateral root initiation and around roots hairs, but not around root tips. Such biosensors can reveal intriguing aspects of the environment and the physiology of the free-living soil S. meliloti before and during the establishment of nodulation, and they provide a nondestructive, spatially explicit method for examining rhizosphere soil chemical composition. PMID- 11274356 TI - Purification and characterization of an autoregulatory substance capable of regulating the morphological transition in Candida albicans. AB - The yeast Candida albicans has a distinguishing feature, dimorphism, which is the ability to switch between two morphological forms: a budding yeast form and a multicellular invasive filamentous form. This ability has been postulated to contribute to the virulence of this organism. Studies on the morphological transition from a filamentous to a budding yeast form in C. albicans have shown that this organism excretes an autoregulatory substance into the culture medium. This substance was extracted and purified by normal-phase and reversed-phase HPLC. The autoregulatory substance was structurally identified as 3,7,11 trimethyl-2,6,10-dodecatrienoate (farnesoic acid) by NMR and mass spectrometry. Growth experiments suggest that this substance does not inhibit yeast cell growth but inhibits filamentous growth. These findings have implications for developmental signaling by the fungus and might have medicinal value in the development of antifungal therapies. PMID- 11274358 TI - Recovery of Diadema antillarum reduces macroalgal cover and increases abundance of juvenile corals on a Caribbean reef. AB - The transition of many Caribbean reefs from coral to macroalgal dominance has been a prominent issue in coral reef ecology for more than 20 years. Alternative stable state theory predicts that these changes are reversible but, to date, there is little indication of this having occurred. Here we present evidence of the initiation of such a reversal in Jamaica, where shallow reefs at five sites along 8 km of coastline now are characterized by a sea urchin-grazed zone with a mean width of 60 m. In comparison to the seaward algal zone, macroalgae are rare in the urchin zone, where the density of Diadema antillarum is 10 times higher and the density of juvenile corals is up to 11 times higher. These densities are close to those recorded in the late 1970s and early 1980s and are in striking contrast to the decade-long recruitment failure for both Diadema and scleractinians. If these trends continue and expand spatially, reefs throughout the Caribbean may again become dominated by corals and algal turf. PMID- 11274357 TI - Tumor metastasis suppressor nm23H1 regulates Rac1 GTPase by interaction with Tiam1. AB - The putative tumor metastasis suppressor nm23H1 was originally identified in murine melanomas by subtraction cloning. It displays nucleoside diphosphate kinase activity and regulates cellular events, including growth and development. Recently nm23H1 has been reported to also act as a GTPase-activating protein of the Ras-related GTPase Rad. We attempted to determine whether nm23H1 also regulates Rho-family GTPases. Although we were unable to detect a direct association between nm23H1 and Rho-family GTPases, nm23H1 was shown to be associated with a Rac1-specific nucleotide exchange factor, Tiam1, by interaction with its amino-terminal region in extracts from the cells expressing exogenous Tiam1 and from native tissue. Overexpression of nm23H1 inhibited the Tiam1 induced production of GTP-bound Rac1 and activation of c-Jun kinase. On the other hand, forced overexpression of the wild type, but not the kinase-inactivated mutant of nm23H1, converted the GDP-bound forms of Rac1, Cdc42, and RhoA to their GTP-bound forms in vitro by its nucleoside diphosphate kinase activity, but nm23H1 alone apparently did not produce the GTP-bound form of these GTPases in vivo. These results suggest that nm23H1 negatively regulates Tiam1 and inhibits Rac1 activation in vivo. Moreover, adhesion-stimulated membrane ruffles of Rat1 fibroblasts were reduced by overexpression of nm23H1. Based on these observations, we concluded that we had identified a function of nm23H1 as a regulator of Rac1 and that it may be related to the effect of nm23H1 as a tumor metastasis suppressor. PMID- 11274360 TI - Rhesus monkeys know when they remember. AB - Humans are consciously aware of some memories and can make verbal reports about these memories. Other memories cannot be brought to consciousness, even though they influence behavior. This conspicuous difference in access to memories is central in taxonomies of human memory systems but has been difficult to document in animal studies, suggesting that some forms of memory may be unique to humans. Here I show that rhesus macaque monkeys can report the presence or absence of memory. Although it is probably impossible to document subjective, conscious properties of memory in nonverbal animals, this result objectively demonstrates an important functional parallel with human conscious memory. Animals able to discern the presence and absence of memory should improve accuracy if allowed to decline memory tests when they have forgotten, and should decline tests most frequently when memory is attenuated experimentally. One of two monkeys examined unequivocally met these criteria under all test conditions, whereas the second monkey met them in all but one case. Probe tests were used to rule out "cueing" by a wide variety of environmental and behavioral stimuli, leaving detection of the absence of memory per se as the most likely mechanism underlying the monkeys' abilities to selectively decline memory tests when they had forgotten. PMID- 11274359 TI - Lack of tissue glucocorticoid reactivation in 11beta -hydroxysteroid dehydrogenase type 1 knockout mice ameliorates age-related learning impairments. AB - 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) intracellularly regenerates active corticosterone from circulating inert 11-dehydrocorticosterone (11-DHC) in specific tissues. The hippocampus is a brain structure particularly vulnerable to glucocorticoid neurotoxicity with aging. In intact hippocampal cells in culture, 11beta-HSD-1 acts as a functional 11beta-reductase reactivating inert 11-DHC to corticosterone, thereby potentiating kainate neurotoxicity. We examined the functional significance of 11beta-HSD-1 in the central nervous system by using knockout mice. Aged wild-type mice developed elevated plasma corticosterone levels that correlated with learning deficits in the watermaze. In contrast, despite elevated plasma corticosterone levels throughout life, this glucocorticoid-associated learning deficit was ameliorated in aged 11beta-HSD-1 knockout mice, implicating lower intraneuronal corticosterone levels through lack of 11-DHC reactivation. Indeed, aged knockout mice showed significantly lower hippocampal tissue corticosterone levels than wild-type controls. These findings demonstrate that tissue corticosterone levels do not merely reflect plasma levels and appear to play a more important role in hippocampal functions than circulating blood levels. The data emphasize the crucial importance of local enzymes in determining intracellular glucocorticoid activity. Selective 11beta HSD-1 inhibitors may protect against hippocampal function decline with age. PMID- 11274361 TI - Tissue spreading on implantable substrates is a competitive outcome of cell-cell vs. cell-substratum adhesivity. AB - While the interactions of cells with polymeric substrata are widely studied, the influence of cell-cell cohesiveness on tissue spreading has not been rigorously investigated. Here we demonstrate that the rate of tissue spreading over a two dimensional substratum reflects a competition or "tug-of-war" between cell-cell and cell-substratum adhesions. We have generated both a "library" of structurally related copolymeric substrata varying in their adhesivity to cells and a library of genetically engineered cell populations varying only in cohesiveness. Cell substratum adhesivity was varied through the poly(ethylene glycol) content of a series of copolymeric substrata, whereas cell-cell cohesiveness was varied through the expression of the homophilic cohesion molecules N- and R-cadherin by otherwise noncohesive L929 cells. In the key experiment, multicellular aggregates containing about 600 cells were allowed to spread onto copolymeric surfaces. We compared the spreading behavior of aggregates having different levels of cell cell cohesiveness in a series of copolymeric substrata having different levels of cell-substratum adhesivity. In these experiments, cell-cell cohesiveness was measured by tissue surface tensiometry, and cell-substratum adhesivity was assessed by a distractive method. Tissue spreading was assayed by confocal microscopy as the rate of cell emigration from similar-sized, fluorescence labeled, multicellular aggregates deposited on each of the substrata. We demonstrate that either decreasing substratum adhesivity or increasing cell-cell cohesiveness dramatically slowed the spreading rate of cell aggregates. PMID- 11274362 TI - Migration of keratinocytes through tunnels of digested fibrin. AB - We report here a hitherto undescribed form of cell migration. When a suspension of human keratinocytes is plated on a fibrin matrix, single cells invade the matrix and progress through it as rounded cells by dissolving the fibrin and thereby creating tunnels. These tunnels are cylindrical or helical, the latter being the result of constant change in the path of cellular advance around the helical axis. Helical tunnel formation is strongly promoted by epidermal growth factor. The rate of migration of the cell through the track of a helical tunnel (up to 2.1 mm per day) is about 7-fold greater than through a cylindrical tunnel. Pericellular fibrinolysis leading to tunnel formation depends on the presence of plasminogen in the medium and its conversion to plasmin by a cellular activator. Formation of tunnels requires that plasminogen activator be localized on the advancing surface of the keratinocyte; we propose that the tunnel is cylindrical when the site of release of plasmin is located at a fixed point on the cell surface and helical when the site of release precesses. PMID- 11274363 TI - Role of MEKK2-MEK5 in the regulation of TNF-alpha gene expression and MEKK2-MKK7 in the activation of c-Jun N-terminal kinase in mast cells. AB - Cross-linking of the high-affinity IgE receptor (FcepsilonRI) on mast cells with IgE and multivalent antigen triggers mitogen-activated protein (MAP) kinase activation and cytokine gene expression. We report here that MAP kinase kinase 4 (MKK4) gene disruption does not affect either MAP kinase activation or cytokine gene expression in response to cross-linking of FcepsilonRI in embryonic stem cell-derived mast cells. MKK7 is activated in response to cross-linking of FcepsilonRI, and this activation is inhibited by MAP/ERK kinase (MEK) kinase 2 (MEKK2) gene disruption. In addition, expression of kinase-inactive MKK7 in the murine mast cell line MC/9 inhibits c-Jun NH(2)-terminal kinase (JNK) activation in response to cross-linking of FcepsilonRI, whereas expression of kinase inactive MKK4 does not affect JNK activation by this stimulus. However, FcepsilonRI-induced activation of the tumor necrosis factor-alpha (TNF-alpha) gene promoter is not affected by expression of kinase-inactive MKK7. We describe an alternative pathway by which MEKK2 activates MEK5 and big MAP kinase1/extracellular signal-regulated kinase 5 in addition to MKK7 and JNK, and interruption of this pathway inhibits TNF-alpha promoter activation. These findings suggest that JNK activation by antigen cross-linking is dependent on the MEKK2-MKK7 pathway, and cytokine production in mast cells is regulated in part by the signaling complex MEKK2-MEK5-ERK5. PMID- 11274364 TI - p19ARF targets certain E2F species for degradation. AB - p19ARF suppresses the growth of cells lacking p53 through an unknown mechanism. p19ARF was found to complex with transcription factors E2F1, -2, and -3. Levels of endogenous or ectopically expressed E2F1, -2, and -3, but not E2F6, were reduced after synthesis of p19ARF, through a mechanism involving increased turnover. p19ARF-induced degradation of E2F1 depended on a functional proteasome, and E2F1 was relocalized to nucleoli when coexpressed with p19ARF. Consistent with reduced levels of E2F1 and E2F3, the proliferation of cells defective for p53 function was suppressed by p19ARF, and the effect was partially reversed by ectopic overexpression of E2F1. These results suggest a broader role for p19ARF as a tumor suppressor, in which targeting of certain E2F species may cooperate with stimulation of the p53 pathway to counteract oncogenic growth signals. PMID- 11274365 TI - PTEN controls tumor-induced angiogenesis. AB - Mutations of the tumor suppressor PTEN, a phosphatase with specificity for 3 phosphorylated inositol phospholipids, accompany progression of brain tumors from benign to the most malignant forms. Tumor progression, particularly in aggressive and malignant tumors, is associated with the induction of angiogenesis, a process termed the angiogenic switch. Therefore, we tested whether PTEN regulates tumor progression by modulating angiogenesis. U87MG glioma cells stably reconstituted with PTEN cDNA were tested for growth in a nude mouse orthotopic brain tumor model. We observed that the reconstitution of wild-type PTEN had no effect on in vitro proliferation but dramatically decreased tumor growth in vivo and prolonged survival in mice implanted intracranially with these tumor cells. PTEN reconstitution diminished phosphorylation of AKT within the PTEN-reconstituted tumor, induced thrombospondin 1 expression, and suppressed angiogenic activity. These effects were not observed in tumors reconstituted with a lipid phosphatase inactive G129E mutant of PTEN, a result that provides evidence that the lipid phosphatase activity of PTEN regulates the angiogenic response in vivo. These data provide evidence that PTEN regulates tumor-induced angiogenesis and the progression of gliomas to a malignant phenotype via the regulation of phosphoinositide-dependent signals. PMID- 11274366 TI - The many ways to cross the plasma membrane. PMID- 11274367 TI - Ca2+-binding activity of a COOH-terminal fragment of the Drosophila BK channel involved in Ca2+-dependent activation. AB - Mutational and biophysical analysis suggests that an intracellular COOH-terminal domain of the large conductance Ca(2+)-activated K(+) channel (BK channel) contains Ca(2+)-binding site(s) that are allosterically coupled to channel opening. However the structural basis of Ca(2+) binding to BK channels is unknown. To pursue this question, we overexpressed the COOH-terminal 280 residues of the Drosophila slowpoke BK channel (Dslo-C280) as a FLAG- and His(6)-tagged protein in Escherichia coli. We purified Dslo-C280 in soluble form and used a (45)Ca(2+)-overlay protein blot assay to detect Ca(2+) binding. Dslo-C280 exhibits specific binding of (45)Ca(2+) in comparison with various control proteins and known EF-hand Ca(2+)-binding proteins. A mutation (D5N5) of Dslo C280, in which five consecutive Asp residues of the "Ca-bowl" motif are changed to Asn, reduces (45)Ca(2+)-binding activity by 56%. By electrophysiological assay, the corresponding D5N5 mutant of the Drosophila BK channel expressed in HEK293 cells exhibits lower Ca(2+) sensitivity for activation and a shift of approximately +80 mV in the midpoint voltage for activation. This effect is associated with a decrease in the Hill coefficient (N) for activation by Ca(2+) and a reduction in apparent Ca(2+) affinity, suggesting the loss of one Ca(2+) binding site per monomer. These results demonstrate a functional correlation between Ca(2+) binding to a specific region of the BK protein and Ca(2+) dependent activation, thus providing a biochemical approach to study this process. PMID- 11274368 TI - Myc represses the p21(WAF1/CIP1) promoter and interacts with Sp1/Sp3. AB - The cyclin-dependent kinase inhibitor p21((WAF1/CIP1)) inhibits proliferation both in vitro and in vivo, and overexpression of p21 in normal and tumor cell lines results in cell cycle arrest. In contrast, ectopic expression of Myc alleviates G(1) cell cycle arrest. Recent studies showed that Myc can repress p21 transcription, thereby overriding a p21-mediated cell cycle checkpoint. We found that activation of a Myc-estrogen receptor fusion protein by 4-hydroxytamoxifen in mouse cells resulted in suppression of endogenous p21 transcription. This effect was observed in the absence of de novo protein synthesis and was independent of histone deacetylase activity. In transient transfection studies, Myc effectively repressed p21 promoter constructs containing only 119 bp of sequence upstream of the transcription start site. This region contains multiple Sp1-binding sites and a potential initiator element, but no canonical Myc DNA binding sites. Deletion of the potential initiator element does not affect repression of the p21 promoter by c-Myc. Coimmunoprecipitation and glutathione S transferase pull-down experiments demonstrate that c-Myc may form complexes with Sp1/Sp3. We found that the central region of c-Myc interacts with the zinc finger domain of Sp1. Because Sp1 is required for p21 transcription, it is possible that Myc may down-regulate p21 transcription, at least in part, by sequestering Sp1. Repression of the p21 promoter may contribute to the ability of c-Myc to promote cell proliferation. PMID- 11274369 TI - Multicopy plasmids are clustered and localized in Escherichia coli. AB - We localized the multicopy plasmid RK2 in Escherichia coli and found that the number of fluorescent foci observed in each cell was substantially less than the copy number of the plasmid, suggesting that many copies of RK2 are grouped into a few multiplasmid clusters. In minimal glucose media, the majority of cells had one or two foci, with a single focus localized near midcell, and two foci near the 1/4 and 3/4 cell positions. The number of foci per cell increased with cell length and with growth rate, and decreased upon entering stationary phase, suggesting a coordination of RK2 replication or segregation with the bacterial cell cycle. Time-lapse microscopy demonstrated that partitioning of RK2 foci is achieved by the splitting of a single focus into two or three smaller foci, which are capable of separating with rapid kinetics. A derivative of the high-copy number plasmid pUC19 containing the lacO array was also localized by tagging with GFP-LacI. Whereas many of the cells contained numerous, randomly diffusing foci, most cells exhibited one or two plasmid clusters located at midcell or the cell quarter positions. Our results suggest a model in which multicopy plasmids are not always randomly diffusing throughout the cell as previously thought, but can be replicated and partitioned in clusters targeted to specific locations. PMID- 11274370 TI - Mammalian mad2 and bub1/bubR1 recognize distinct spindle-attachment and kinetochore-tension checkpoints. AB - Metaphase checkpoint controls sense abnormalities of chromosome alignment during mitosis and prevent progression to anaphase until proper alignment has been attained. A number of proteins, including mad2, bub1, and bubR1, have been implicated in the metaphase checkpoint control in mammalian cells. Metaphase checkpoints have been shown, in various systems, to read loss of either spindle tension or microtubule attachment at the kinetochore. Characteristically, HeLa cells arrest in metaphase in response to low levels of microtubule inhibitors that leave an intact spindle and a metaphase plate. Here we show that the arrest induced by nanomolar vinblastine correlates with loss of tension at the kinetochore, and that in response the checkpoint proteins bub1 and bubR1 are recruited to the kinetochore but mad2 is not. mad2 remains competent to respond and is recruited at higher drug doses that disrupt spindle association with the kinetochores. Further, although mad2 forms a complex with cdc20, it does not associate with bub1 or bubR1. We conclude that mammalian bub1/bubR1 and mad2 operate as elements of distinct pathways sensing tension and attachment, respectively. PMID- 11274371 TI - Evidence for two active branches for electron transfer in photosystem I. AB - All photosynthetic reaction centers share a common structural theme. Two related, integral membrane polypeptides sequester electron transfer cofactors into two quasi-symmetrical branches, each of which incorporates a quinone. In type II reaction centers [photosystem (PS) II and proteobacterial reaction centers], electron transfer proceeds down only one of the branches, and the mobile quinone on the other branch is used as a terminal acceptor. PS I uses iron-sulfur clusters as terminal acceptors, and the quinone serves only as an intermediary in electron transfer. Much effort has been devoted to understanding the unidirectionality of electron transport in type II reaction centers, and it was widely thought that PS I would share this feature. We have tested this idea by examining in vivo kinetics of electron transfer from the quinone in mutant PS I reaction centers. This transfer is associated with two kinetic components, and we show that mutation of a residue near the quinone in one branch specifically affects the faster component, while the corresponding mutation in the other branch specifically affects the slower component. We conclude that both electron transfer branches in PS I are active. PMID- 11274372 TI - Roles for genomic imprinting and the zygotic genome in placental development. AB - The placenta contains several types of feto-maternal interfaces where zygote derived cells interact with maternal cells or maternal blood for the promotion of fetal growth and viability. The genetic factors regulating the interactions between different cell types within feto-maternal interfaces and the relative contributions of the maternal and zygotic genomes are poorly understood. Genomic imprinting, the epigenetic process responsible for parental origin-dependent functional differences between homologous chromosomes, has been proposed to contribute to these events. Previous studies showed that mouse conceptuses with an absence of imprinted differences between the two copies of chromosome 12 (upon paternal inheritance of both copies) die late in gestation and have a variety of defects, including placentomegaly. Here we examined the role of chromosome 12 imprinting in these placentae in more detail. We show that the spatial interactions between different cell types within feto-maternal interfaces are defective and identify abnormal behaviors in both zygote-derived and maternal cells that are attributed to the genome of the zygote but not the mother. These include compromised invasion of the maternal decidualized endometrium and the central maternal artery situated within it by zygote-derived trophoblast, abnormalities in the wall of the central maternal artery, and defects within the zygote-derived cellular layer of the labyrinth, which is in direct contact with maternal blood. These findings demonstrate multiple roles for chromosome 12 imprinting in the placenta that have not previously been associated with imprinting effects. They provide insights into the function of imprinting in placental development and have evolutionary and clinical implications. PMID- 11274373 TI - beta -Amyloid peptide blocks the response of alpha 7-containing nicotinic receptors on hippocampal neurons. AB - Alzheimer's disease produces a devastating decline in mental function, with profound effects on learning and memory. Early consequences of the disease include the specific loss of cholinergic neurons in brain, diminished cholinergic signaling, and the accumulation of beta-amyloid peptide in neuritic plaques. Of the nicotinic acetylcholine receptors at risk, the most critical may be those containing the alpha7 gene product (alpha7-nAChRs), because they are widespread, have a high relative permeability to calcium, and regulate numerous cellular events in the nervous system. With the use of whole-cell patch-clamp recording we show here that nanomolar concentrations of beta-amyloid peptides specifically and reversibly block alpha7-nAChRs on rat hippocampal neurons in culture. The block is noncompetitive, voltage-independent, and use-independent and is mediated through the N-terminal extracellular domain of the receptor. It does not appear to require either calcium influx or G protein activation. beta-Amyloid blockade is likely to be a common feature of alpha7-nAChRs because it applies to the receptors at both somato-dendritic and presynaptic locations on rat hippocampal neurons and extends to homologous receptors on chick ciliary ganglion neurons as well. Because alpha7-nAChRs in the central nervous system are thought to have numerous functions and recently have been implicated in learning and memory, impaired receptor function in this case may contribute to cognitive deficits associated with Alzheimer's disease. PMID- 11274374 TI - Comparative evaluation of the antitumor activity of antiangiogenic proteins delivered by gene transfer. AB - Although the systemic administration of a number of different gene products has been shown to result in the inhibition of angiogenesis and tumor growth in different animal tumor models, the relative potency of those gene products has not been studied rigorously. To address this issue, recombinant adenoviruses encoding angiostatin, endostatin, and the ligand-binding ectodomains of the vascular endothelial growth factor receptors Flk1, Flt1, and neuropilin were generated and used to systemically deliver the different gene products in several different preexisting murine tumor models. Single i.v. injections of viruses encoding soluble forms of Flk1 or Flt1 resulted in approximately 80% inhibition of preexisting tumor growth in murine models involving both murine (Lewis lung carcinoma, T241 fibrosarcoma) and human (BxPC3 pancreatic carcinoma) tumors. In contrast, adenoviruses encoding angiostatin, endostatin, or neuropilin were significantly less effective. A strong correlation was observed between the effects of the different viruses on tumor growth and the activity of the viruses in the inhibition of corneal micropocket angiogenesis. These data underscore the need for comparative analyses of different therapeutic approaches that target tumor angiogenesis and provide a rationale for the selection of specific antiangiogenic gene products as lead candidates for use in gene therapy approaches aimed at the treatment of malignant and ocular disorders. PMID- 11274375 TI - Role of tumor-host interactions in interstitial diffusion of macromolecules: cranial vs. subcutaneous tumors. AB - The large size of many novel therapeutics impairs their transport through the tumor extracellular matrix and thus limits their therapeutic effectiveness. We propose that extracellular matrix composition, structure, and distribution determine the transport properties in tumors. Furthermore, because the characteristics of the extracellular matrix largely depend on the tumor-host interactions, we postulate that diffusion of macromolecules will vary with tumor type as well as anatomical location. Diffusion coefficients of macromolecules and liposomes in tumors growing in cranial windows (CWs) and dorsal chambers (DCs) were measured by fluorescence recovery after photobleaching. For the same tumor types, diffusion of large molecules was significantly faster in CW than in DC tumors. The greater diffusional hindrance in DC tumors was correlated with higher levels of collagen type I and its organization into fibrils. For molecules with diameters comparable to the interfibrillar space the diffusion was 5- to 10-fold slower in DC than in CW tumors. The slower diffusion in DC tumors was associated with a higher density of host stromal cells that synthesize and organize collagen type I. Our results point to the necessity of developing site-specific drug carriers to improve the delivery of molecular medicine to solid tumors. PMID- 11274376 TI - Epithelial water absorption: osmosis or cotransport? PMID- 11274377 TI - New functions for glia in the brain. PMID- 11274378 TI - Discovery of a protein required for photosynthetic membrane assembly. PMID- 11274379 TI - How does a system respond when driven away from thermal equilibrium? PMID- 11274380 TI - Computing an organism. PMID- 11274382 TI - Brownian flocculation of polymer colloids in the presence of a secondary minimum. AB - Most analyses of Brownian flocculation apply to conditions where London-van der Waals attractive forces cause particles to be strongly bound in a deep interparticle potential well. In this paper, results are reported that show the interaction between primary- and secondary-minimum flocculation when the interparticle potential curve reflects both attractive and electrostatic repulsive forces. The process is highly time-dependent because of transfer of particles from secondary- to primary-minimum flocculation. Essential features of the analysis are corroborated by experiments with 0.80-microm polystyrene spheres suspended in aqueous solutions of NaCl over a range of ionic strengths. In all cases, experiments were restricted to the initial stage of coagulation, where singlets and doublets predominate. PMID- 11274381 TI - Uncoupling fibroblast growth factor receptor 2 ligand binding specificity leads to Apert syndrome-like phenotypes. PMID- 11274383 TI - Self-assembled monolayers from a designed combinatorial library of de novo beta sheet proteins. AB - A variety of naturally occurring biomaterials owe their unusual structural and mechanical properties to layers of beta-sheet proteins laminated between layers of inorganic mineral. To explore the possibility of fabricating novel two dimensional protein layers, we studied the self-assembly properties of de novo proteins from a designed combinatorial library. Each protein in the library has a distinct 63 amino acid sequence, yet they all share an identical binary pattern of polar and nonpolar residues, which was designed to favor the formation of six stranded amphiphilic beta-sheets. Characterization of proteins isolated from the library demonstrates that (i) they self assemble into monolayers at an air/water interface; (ii) the monolayers are dominated by beta-sheet secondary structure, as shown by both circular dichroism and infrared spectroscopies; and (iii) the measured areas (500- 600 A(2)) of individual protein molecules in the monolayers match those expected for proteins folded into amphiphilic beta-sheets. The finding that similar structures are formed by distinctly different protein sequences suggests that assembly into beta-sheet monolayers can be encoded by binary patterning of polar and nonpolar amino acids. Moreover, because the designed binary pattern is compatible with a wide variety of different sequences, it may be possible to fabricate beta-sheet monolayers by using combinations of side chains that are explicitly designed to favor particular applications of novel biomaterials. PMID- 11274384 TI - Free energy reconstruction from nonequilibrium single-molecule pulling experiments. AB - Laser tweezers and atomic force microscopes are increasingly used to probe the interactions and mechanical properties of individual molecules. Unfortunately, using such time-dependent perturbations to force rare molecular events also drives the system away from equilibrium. Nevertheless, we show how equilibrium free energy profiles can be extracted rigorously from repeated nonequilibrium force measurements on the basis of an extension of Jarzynski's remarkable identity between free energies and the irreversible work. PMID- 11274385 TI - Covalent intermediate trapped in 2-keto-3-deoxy-6- phosphogluconate (KDPG) aldolase structure at 1.95-A resolution. AB - 2-Keto-3-deoxy-6-phosphogluconate (KDPG) aldolase catalyzes the reversible cleavage of KDPG to pyruvate and glyceraldehyde-3-phosphate. The enzyme is a class I aldolase whose reaction mechanism involves formation of Schiff base intermediates between Lys-133 and a keto substrate. A covalent adduct was trapped by flash freezing KDPG aldolase crystals soaked with 10 mM pyruvate in acidic conditions at pH 4.6. Structure determination to 1.95-A resolution showed that pyruvate had undergone nucleophilic attack with Lys-133, forming a protonated carbinolamine intermediate, a functional Schiff base precursor, which was stabilized by hydrogen bonding with active site residues. Carbinolamine interaction with Glu-45 indicates general base catalysis of several rate steps. Stereospecific addition is ensured by aromatic interaction of Phe-135 with the pyruvate methyl group. In the native structure, Lys-133 donates all of its hydrogen bonds, indicating the presence of an epsilon-ammonium salt group. Nucleophilic activation is postulated to occur by proton transfer in the monoprotonated zwitterionic pair (Glu-45/Lys-133). Formation of the zwitterionic pair requires prior side chain rearrangement by protonated Lys-133 to displace a water molecule, hydrogen bonded to the zwitterionic residues. PMID- 11274386 TI - Akt phosphorylates and regulates the orphan nuclear receptor Nur77. AB - The immediate early gene NUR77 (also called NGFI-B) is required for T cell antigen receptor-mediated cell death and is induced to very high levels in immature thymocytes and T cell hybridomas undergoing apoptosis. The Akt (PKB) kinase is a key player in transduction of anti-apoptotic and proliferative signals in T cells. Because Nur77 has a putative Akt phosphorylation site at Ser 350, and phosphorylation of this residue is critical for the transactivation activity of Nur77, we investigated whether Akt regulates Nur77. Coimmunoprecipitation experiments showed the detection of Nur77 in Akt immune complexes, suggesting that Nur77 and Akt physically interact. We further show that Akt specifically phosphorylates Ser-350 of the Nur77 protein within its DNA binding domain in vitro and in vivo in 293 and NIH 3T3 cells. Because phosphorylation of Ser-350 of Nur77 is critical for its function as a transcription factor, we examined the effect of Akt on this function. By using luciferase assay experiments, we showed that phosphorylation of Nur77 by Akt decreased the transcriptional activity of Nur77 by 50--85%. Thus, we show that Akt interacts with Nur77 and inactivates Nur77 by phosphorylation at Ser-350 in a phosphatidylinositol 3-kinase-dependent manner, connecting the phosphatidylinositol 3-kinase-dependent Akt pathway and a nuclear receptor pathway. PMID- 11274387 TI - Formation of a GNRA tetraloop in P5abc can disrupt an interdomain interaction in the Tetrahymena group I ribozyme. AB - The secondary structure of a truncated P5abc subdomain (tP5abc, a 56-nucleotide RNA) of the Tetrahymena thermophila group I intron ribozyme changes when its tertiary structure forms. We have now used heteronuclear NMR spectroscopy to determine its conformation in solution. The tP5abc RNA that contains only secondary structure is extended compared with the tertiary folded form; both forms coexist in slow chemical exchange (the interconversion rate constant is slower than 1 s(-1)) in the presence of magnesium. Kinetic experiments have shown that tertiary folding of the P5abc subdomain is one of the earliest folding transitions in the group I intron ribozyme, and that it leads to a metastable misfolded intermediate. Previous mutagenesis studies suggest that formation of the extended P5abc structure described here destabilize a misfolded intermediate. This study shows that the P5abc RNA subdomain containing a GNRA tetraloop in P5c (in contrast to the five-nucleotide loop P5c in the tertiary folded ribozyme) can disrupt the base-paired interdomain (P14) interaction between P5c and P2. PMID- 11274388 TI - A human protein containing multiple types of protease-inhibitory modules. AB - By using sensitive homology-search and gene-finding programs, we have found that a genomic region from the tip of the short arm of human chromosome 16 (16p13.3) encodes a putative secreted protein consisting of a domain related to the whey acidic protein (WAP) domain, a domain homologous with follistatin modules of the Kazal-domain family (FS module), an immunoglobulin-related domain (Ig domain), two tandem domains related to Kunitz-type protease inhibitor modules (KU domains), and a domain belonging to the recently defined NTR-module family (NTR domain). The gene encoding these WAP, FS, Ig, KU, and NTR modules (hereafter referred to as the WFIKKN gene) is intron-depleted--its single 1,157-bp intron splits the WAP module. The validity of our gene prediction was confirmed by sequencing a WFIKKN cDNA cloned from a lung cDNA library. Studies on the tissue expression pattern of the WFIKKN gene have shown that the gene is expressed primarily in pancreas, kidney, liver, placenta, and lung. As to the function of the WFIKKN protein, it is noteworthy that it contains FS, WAP, and KU modules, i.e., three different module types homologous with domains frequently involved in inhibition of serine proteases. The protein also contains an NTR module, a domain type implicated in inhibition of zinc metalloproteinases of the metzincin family. On the basis of its intriguing homologies, we suggest that the WFIKKN protein is a multivalent protease inhibitor that may control the action of multiple types of serine proteases as well as metalloproteinase(s). PMID- 11274389 TI - Identification, retinoid binding, and x-ray analysis of a human retinol-binding protein. AB - Two cellular retinol-binding proteins (CRBP I and II) with distinct tissue distributions and retinoid-binding properties have been recognized thus far in mammals. Here, we report the identification of a human retinol-binding protein resembling type I (55.6% identity) and type II (49.6% identity) CRBPs, but with a unique H residue in the retinoid-binding site and a distinctively different tissue distribution. Additionally, this binding protein (CRBP III) exhibits a remarkable sequence identity (62.2%) with the recently identified iota crystallin/CRBP of the diurnal gecko Lygodactylus picturatus [Werten, P. J. L., Roll, B., van Alten, D. M. F. & de Jong, W. W. (2000) Proc. Natl. Acad. Sci. USA 97, 3282-3287 (First Published March 21, 2000; 10.1073/pnas.050500597)]. CRBP III and all-trans-retinol form a complex (K(d) approximately 60 nM), the absorption spectrum of which is characterized by the peculiar fine structure typical of the spectra of holo-CRBP I and II. As revealed by a 2.3-A x-ray molecular model of apo-CRBP III, the amino acid residues that line the retinol-binding site in CRBP I and II are positioned nearly identically in the structure of CRBP III. At variance with the human CRBP I and II mRNAs, which are most abundant in ovary and intestine, respectively, the CRBP III mRNA is expressed at the highest levels in kidney and liver thus suggesting a prominent role for human CRBP III as an intracellular mediator of retinol metabolism in these tissues. PMID- 11274390 TI - Allosteric crosstalk between peptide-binding, transport, and ATP hydrolysis of the ABC transporter TAP. AB - The transporter associated with antigen processing (TAP) is essential for intracellular transport of protein fragments into the endoplasmic reticulum for loading of major histocompatibility complex (MHC) class I molecules. On the cell surface, these peptide-MHC complexes are monitored by cytotoxic T lymphocytes. To study the ATP hydrolysis of TAP, we developed an enrichment and reconstitution procedure, by which we fully restored TAP function in proteoliposomes. A TAP specific ATPase activity was identified that could be stimulated by peptides and blocked by the herpes simplex virus protein ICP47. Strikingly, the peptide binding motif of TAP directly correlates with the stimulation of the ATPase activity, demonstrating that the initial peptide-binding step is responsible for TAP selectivity. ATP hydrolysis follows Michaelis-Menten kinetics with a maximal velocity V(max) of 2 micromol/min per mg TAP, corresponding to a turnover number of approximately 5 ATP per second. This turnover rate is sufficient to account for the role of TAP in peptide loading of MHC molecules and the overall process of antigen presentation. Interestingly, sterically restricted peptides that bind but are not transported by TAP do not stimulate ATPase activity. These results point to coordinated dialogue between the peptide-binding site, the nucleotide binding domain, and the translocation site via conformational changes within the TAP complex. PMID- 11274392 TI - The use of mRNA display to select high-affinity protein-binding peptides. AB - We report the use of "mRNA display," an in vitro selection technique, to identify peptide aptamers to a protein target. mRNA display allows for the preparation of polypeptide libraries with far greater complexity than is possible with phage display. Starting with a library of approximately 10(13) random peptides, 20 different aptamers to streptavidin were obtained, with dissociation constants as low as 5 nM. These aptamers function without the aid of disulfide bridges or engineered scaffolds, yet possess affinities comparable to those for monoclonal antibody-antigen complexes. The aptamers bind streptavidin with three to four orders of magnitude higher affinity than those isolated previously by phage display from lower complexity libraries of shorter random peptides. Like previously isolated peptides, they contain an HPQ consensus motif. This study shows that, given sufficient length and diversity, high-affinity aptamers can be obtained even from random nonconstrained peptide libraries. By engineering structural constraints into these ultrahigh complexity peptide libraries, it may be possible to produce binding agents with subnanomolar binding constants. PMID- 11274391 TI - Crystal structure of the ectodomain of Methuselah, a Drosophila G protein-coupled receptor associated with extended lifespan. AB - The Drosophila mutant methuselah (mth) was identified from a screen for single gene mutations that extended average lifespan. Mth mutants have a 35% increase in average lifespan and increased resistance to several forms of stress, including heat, starvation, and oxidative damage. The protein affected by this mutation is related to G protein-coupled receptors of the secretin receptor family. Mth, like secretin receptor family members, has a large N-terminal ectodomain, which may constitute the ligand binding site. Here we report the 2.3-A resolution crystal structure of the Mth extracellular region, revealing a folding topology in which three primarily beta-structure-containing domains meet to form a shallow interdomain groove containing a solvent-exposed tryptophan that may represent a ligand binding site. The Mth structure is analyzed in relation to predicted Mth homologs and potential ligand binding features. PMID- 11274394 TI - Computational method to reduce the search space for directed protein evolution. AB - We introduce a computational method to optimize the in vitro evolution of proteins. Simulating evolution with a simple model that statistically describes the fitness landscape, we find that beneficial mutations tend to occur at amino acid positions that are tolerant to substitutions, in the limit of small libraries and low mutation rates. We transform this observation into a design strategy by applying mean-field theory to a structure-based computational model to calculate each residue's structural tolerance. Thermostabilizing and activity increasing mutations accumulated during the experimental directed evolution of subtilisin E and T4 lysozyme are strongly directed to sites identified by using this computational approach. This method can be used to predict positions where mutations are likely to lead to improvement of specific protein properties. PMID- 11274393 TI - RAC, a stable ribosome-associated complex in yeast formed by the DnaK-DnaJ homologs Ssz1p and zuotin. AB - The yeast cytosol contains multiple homologs of the DnaK and DnaJ chaperone family. Our current understanding of which homologs functionally interact is incomplete. Zuotin is a DnaJ homolog bound to the yeast ribosome. We have now identified the DnaK homolog Ssz1p/Pdr13p as zuotin's partner chaperone. Zuotin and Ssz1p form a ribosome-associated complex (RAC) that is bound to the ribosome via the zuotin subunit. RAC is unique among the eukaryotic DnaK-DnaJ systems, as the 1:1 complex is stable, even in the presence of ATP or ADP. In vitro, RAC stimulates the translocation of a ribosome-bound mitochondrial precursor protein into mitochondria, providing evidence for its chaperone-like effect on nascent chains. In agreement with the existence of a functional complex, deletion of each RAC subunit resulted in a similar phenotype in vivo. However, overexpression of zuotin partly rescued the growth defect of the Delta ssz1 strain, whereas overexpression of Ssz1p did not affect the Delta zuo1 strain, suggesting a pivotal function for the DnaJ homolog. PMID- 11274395 TI - Chemically distinct transition states govern rapid dissociation of single L selectin bonds under force. AB - Carbohydrate--protein bonds interrupt the rapid flow of leukocytes in the circulation by initiation of rolling and tethering at vessel walls. The cell surface carbohydrate ligands are glycosylated proteins like the mucin P-selectin glycoprotein ligand-1 (PSGL-1), which bind ubiquitously to the family of E-, P-, and L-selectin proteins in membranes of leukocytes and endothelium. The current view is that carbohydrate-selectin bonds dissociate a few times per second, and the unbinding rate increases weakly with force. However, such studies have provided little insight into how numerous hydrogen bonds, a Ca(2+) metal ion bond, and other interactions contribute to the mechanical strength of these attachments. Decorating a force probe with very dilute ligands and controlling touch to achieve rare single-bond events, we have varied the unbinding rates of carbohydrate--selectin bonds by detachment with ramps of force/time from 10 to 100,000 pN/sec. Testing PSGL-1, its outer 19 aa (19FT), and sialyl Lewis(X) (sLe(X)) against L-selectin in vitro on glass microspheres and in situ on neutrophils, we found that the unbinding rates followed the same dependence on force and increased by nearly 1,000-fold as rupture forces rose from a few to approximately 200 pN. Plotted on a logarithmic scale of loading rate, the rupture forces reveal two prominent energy barriers along the unbinding pathway. Strengths above 75 pN arise from rapid detachment (<0.01 sec) impeded by an inner barrier that requires a Ca(2+) bond between a single sLe(X) and the lectin domain. Strengths below 75 pN occur under slow detachment (>0.01 sec) impeded by the outer barrier, which appears to involve an array of weak (putatively hydrogen) bonds. PMID- 11274396 TI - Computational adaptive optics for live three-dimensional biological imaging. AB - Light microscopy of thick biological samples, such as tissues, is often limited by aberrations caused by refractive index variations within the sample itself. This problem is particularly severe for live imaging, a field of great current excitement due to the development of inherently fluorescent proteins. We describe a method of removing such aberrations computationally by mapping the refractive index of the sample using differential interference contrast microscopy, modeling the aberrations by ray tracing through this index map, and using space-variant deconvolution to remove aberrations. This approach will open possibilities to study weakly labeled molecules in difficult-to-image live specimens. PMID- 11274397 TI - Local osmotic gradients drive the water flux associated with Na(+)/glucose cotransport. AB - It recently was proposed [Loo, D. D. F., Zeuthen, T., Chandy, G. & Wright, E. M. (1996) Proc. Natl. Acad. Sci. USA 93, 13367--13370] that SGLT1, the high affinity intestinal and renal sodium/glucose cotransporter carries water molecules along with the cosubstrates with a strict stoichiometry of two Na(+), one glucose, and approximately 220 water molecules per transport cycle. Using electrophysiology together with sensitive volumetric measurements, we investigated the nature of the driving force behind the cotransporter-mediated water flux. The osmotic water permeability of oocytes expressing human SGLT1 (L(p) +/- SE) averaged 3.8 +/- 0.3 x 10(-4) cm x s(-1) (n = 15) and addition of 100 microM phlorizin (a specific SGLT1 inhibitor) reduced the permeability to 2.2 +/- 0.2 x 10(-4) cm x s(-1) (n = 15), confirming the presence of a significant water permeability closely associated with the cotransporter. Addition of 5 mM alpha-methyl-glucose (alpha MG) induced an average inward current of 800 +/- 10 nA at -50 mV and a water influx reaching 120 +/- 20 pL cm(-2) x s(-1) within 5-8 min. After rapidly inhibiting the Na(+)/glucose cotransport with phlorizin, the water flux remained significantly elevated, clearly indicating the presence of a local osmotic gradient (Delta pi) estimated at 16 +/- 2 mOsm. In short-term experiments, a rapid depolarization from -100 to 0 mV in the presence of alpha MG decreased the cotransport current by 94% but failed to produce a comparable reduction in the swelling rate. A mathematical model depicting the intracellular accumulation of transported osmolytes can accurately account for these observations. It is concluded that, in SGLT1-expressing oocytes, alpha MG-dependent water influx is induced by a local osmotic gradient by using both endogenous and SGLT1-dependent water permeability. PMID- 11274398 TI - Cell autonomous regulation of multiple Dishevelled-dependent pathways by mammalian Nkd. AB - Genetic studies have identified Drosophila Naked Cuticle (Nkd) as an antagonist of the canonical Wnt/beta-catenin signaling pathway, but its mechanism of action remains obscure [Zeng, W., Wharton, K. A., Jr., Mack, J. A., Wang, K., Gadbaw, M., et al. (2000) Nature (London) 403, 789--795]. Here we have cloned a cDNA encoding a mammalian homolog of Drosophila Nkd, mNkd, and demonstrated that mNkd interacts directly with Dishevelled. Dishevelled is an intracellular mediator of both the canonical Wnt pathway and planar cell polarity (PCP) pathway. Activation of the c-Jun-N-terminal kinase has been implicated in the PCP pathway. We showed that mNkd acts in a cell-autonomous manner not only to inhibit the canonical Wnt pathway but also to stimulate c-Jun-N-terminal kinase activity. Expression of mNkd disrupted convergent extension in Xenopus, consistent with a role for mNkd in the PCP pathway. These data suggest that mNkd may act as a switch to direct Dishevelled activity toward the PCP pathway, and away from the canonical Wnt pathway. PMID- 11274399 TI - Restoration of insulin-sensitive glucose transporter (GLUT4) gene expression in muscle cells by the transcriptional coactivator PGC-1. AB - Muscle tissue is the major site for insulin-stimulated glucose uptake in vivo, due primarily to the recruitment of the insulin-sensitive glucose transporter (GLUT4) to the plasma membrane. Surprisingly, virtually all cultured muscle cells express little or no GLUT4. We show here that adenovirus-mediated expression of the transcriptional coactivator PGC-1, which is expressed in muscle in vivo but is also deficient in cultured muscle cells, causes the total restoration of GLUT4 mRNA levels to those observed in vivo. This increased GLUT4 expression correlates with a 3-fold increase in glucose transport, although much of this protein is transported to the plasma membrane even in the absence of insulin. PGC-1 mediates this increased GLUT4 expression, in large part, by binding to and coactivating the muscle-selective transcription factor MEF2C. These data indicate that PGC-1 is a coactivator of MEF2C and can control the level of endogenous GLUT4 gene expression in muscle. PMID- 11274400 TI - Switch from Myc/Max to Mad1/Max binding and decrease in histone acetylation at the telomerase reverse transcriptase promoter during differentiation of HL60 cells. AB - Recent evidence suggests that the Myc and Mad1 proteins are implicated in the regulation of the gene encoding the human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase. We have analyzed the in vivo interaction between endogenous c-Myc and Mad1 proteins and the hTERT promoter in HL60 cells with the use of the chromatin immunoprecipitation assay. The E-boxes at the hTERT proximal promoter were occupied in vivo by c-Myc in exponentially proliferating HL60 cells but not in cells induced to differentiate by DMSO. In contrast, Mad1 protein was induced and bound to the hTERT promoter in differentiated HL60 cells. Concomitantly, the acetylation of the histones at the promoter was significantly reduced. These data suggest that the reciprocal E-box occupancy by c-Myc and Mad1 is responsible for activation and repression of the hTERT gene in proliferating and differentiated HL60 cells, respectively. Furthermore, the histone deacetylase inhibitor trichostatin A inhibited deacetylation of histones at the hTERT promoter and attenuated the repression of hTERT transcription during HL60 cell differentiation. In addition, trichostatin A treatment activated hTERT transcription in resting human lymphocytes and fibroblasts. Taken together, these results indicate that acetylation/deacetylation of histones is operative in the regulation of hTERT expression. PMID- 11274401 TI - Profilin I is essential for cell survival and cell division in early mouse development. AB - Profilins are thought to play a central role in the regulation of de novo actin assembly by preventing spontaneous actin polymerization through the binding of actin monomers, and the adding of monomeric actin to the barbed actin-filament ends. Other cellular functions of profilin in membrane trafficking and lipid based signaling are also likely. Binding of profilins to signaling molecules such as Arp2/3 complex, Mena, VASP, N-WASP, dynamin I, and others, further implicates profilin and actin as regulators of diverse motile activities. In mouse, two profilins are expressed from two distinct genes. Profilin I is expressed at high levels in all tissues and throughout development, whereas profilin II is expressed in neuronal cells. To examine the function of profilin I in vivo, we generated a null profilin I (pfn1(ko)) allele in mice. Homozygous pfn1(ko/ko) mice are not viable. Pfn1(ko/ko) embryos died as early as the two-cell stage, and no pfn1(ko/ko) blastocysts were detectable. Adult pfn1(ko/wt) mice show a 50% reduction in profilin I expression with no apparent impairment of cell function. However, pfn1(ko/wt) embryos have reduced survival during embryogenesis compared with wild type. Although weakly expressed in early embryos, profilin II cannot compensate for lack of profilin I. Our results indicate that mouse profilin I is an essential protein that has dosage-dependent effects on cell division and survival during embryogenesis. PMID- 11274402 TI - Inactivation of menin, a Smad3-interacting protein, blocks transforming growth factor type beta signaling. AB - Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder characterized by endocrine tumors of parathyroids, pancreatic islets, and anterior pituitary. The MEN1 gene encodes a nuclear protein called menin. In MEN1 carriers inactivating mutations give rise to a truncated product consistent with menin acting as a tumor suppressor gene. However, the role of menin in tumorigenesis and its physiological functions are not known. Here, we show that menin inactivation by antisense RNA antagonizes transforming growth factor type beta-mediated cell growth inhibition. Menin interacts with Smad3, and antisense menin suppresses transforming growth factor type beta-induced and Smad3-induced transcriptional activity by inhibiting Smad3/4-DNA binding at specific transcriptional regulatory sites. These results implicate a mechanism of tumorigenesis by menin inactivation. PMID- 11274403 TI - Analysis of transforming activity of human synovial sarcoma-associated chimeric protein SYT-SSX1 bound to chromatin remodeling factor hBRM/hSNF2 alpha. AB - Human synovial sarcoma has been shown to exclusively harbor the chromosomal translocation t(X;18) that produces the chimeric gene SYT-SSX. However, the role of SYT-SSX in cellular transformation remains unclear. In this study, we have established 3Y1 rat fibroblast cell lines that constitutively express SYT, SSX1, and SYT-SSX1 and found that SYT-SSX1 promoted growth rate in culture, anchorage independent growth in soft agar, and tumor formation in nude mice. Deletion of the N-terminal 181 amino acids of SYT-SSX1 caused loss of its transforming activity. Furthermore, association of SYT-SSX1 with the chromatin remodeling factor hBRM/hSNF2 alpha, which regulates transcription, was demonstrated in both SYT-SSX1-expressing 3Y1 cells and in the human synovial sarcoma cell line HS-SY II. The binding region between the two molecules was shown to reside within the N terminal 181 amino acids stretch (aa 1--181) of SYT-SSX1 and 50 amino acids (aa 156--205) of hBRM/hSNF2 alpha and we found that the overexpression of this binding region of hBRM/hSNF2 alpha significantly suppressed the anchorage independent growth of SYT-SSX1-expressing 3Y1 cells. To analyze the transcriptional regulation by SYT-SSX1, we established conditional expression system of SYT-SSX1 and examined the gene expression profiles. The down-regulation of potential tumor suppressor DCC was observed among 1,176 genes analyzed by microarray analysis, and semi-quantitative reverse transcription--PCR confirmed this finding. These data clearly demonstrate transforming activity of human oncogene SYT-SSX1 and also involvement of chromatin remodeling factor hBRM/hSNF2 alpha in human cancer. PMID- 11274404 TI - Identification of a sequence element directing a protein to nuclear speckles. AB - SF3b(155) is an essential spliceosomal protein, highly conserved during evolution. It has been identified as a subunit of splicing factor SF3b, which, together with a second multimeric complex termed SF3a, interacts specifically with the 12S U2 snRNP and converts it into the active 17S form. The protein displays a characteristic intranuclear localization. It is diffusely distributed in the nucleoplasm but highly concentrated in defined intranuclear structures termed "speckles," a subnuclear compartment enriched in small ribonucleoprotein particles and various splicing factors. The primary sequence of SF3b(155) suggests a multidomain structure, different from those of other nuclear speckles components. To identify which part of SF3b(155) determines its specific intranuclear localization, we have constructed expression vectors encoding a series of epitope-tagged SF3b(155) deletion mutants as well as chimeric combinations of SF3b(155) sequences with the soluble cytoplasmic protein pyruvate kinase. Following transfection of cultured mammalian cells, we have identified (i) a functional nuclear localization signal of the monopartite type (KRKRR, amino acids 196--200) and (ii) a molecular segment with multiple threonine proline repeats (amino acids 208--513), which is essential and sufficient to confer a specific accumulation in nuclear speckles. This latter sequence element, in particular amino acids 208--440, is required for correct subcellular localization of SF3b(155) and is also sufficient to target a reporter protein to nuclear speckles. Moreover, this "speckle-targeting sequence" transfers the capacity for interaction with other U2 snRNP components. PMID- 11274405 TI - A splicing switch and gain-of-function mutation in FgfR2-IIIc hemizygotes causes Apert/Pfeiffer-syndrome-like phenotypes. AB - Intercellular signaling by fibroblast growth factors plays vital roles during embryogenesis. Mice deficient for fibroblast growth factor receptors (FgfRs) show abnormalities in early gastrulation and implantation, disruptions in epithelial mesenchymal interactions, as well as profound defects in membranous and endochondrial bone formation. Activating FGFR mutations are the underlying cause of several craniosynostoses and dwarfism syndromes in humans. Here we show that a heterozygotic abrogation of FgfR2-exon 9 (IIIc) in mice causes a splicing switch, resulting in a gain-of-function mutation. The consequences are neonatal growth retardation and death, coronal synostosis, ocular proptosis, precocious sternal fusion, and abnormalities in secondary branching in several organs that undergo branching morphogenesis. This phenotype has strong parallels to some Apert's and Pfeiffer's syndrome patients. PMID- 11274406 TI - Regulation of eye development by frizzled signaling in Xenopus. AB - Eye development in both invertebrates and vertebrates is regulated by a network of highly conserved transcription factors. However, it is not known what controls the expression of these factors to regulate early eye formation and whether transmembrane signaling events are involved. Here we establish a role for signaling via a member of the frizzled family of receptors in regulating early eye development. We show that overexpression of Xenopus frizzled 3 (Xfz3), a receptor expressed during normal eye development, functions cell autonomously to promote ectopic eye formation and can perturb endogenous eye development. Ectopic eyes obtained with Xfz3 overexpression have a laminar organization similar to that of endogenous eyes and contain differentiated retinal cell types. Ectopic eye formation is preceded by ectopic expression of transcription factors involved in early eye development, including Pax6, Rx, and Otx2. Conversely, targeted overexpression of a dominant-negative form of Xfz3 (Nxfz3), consisting of the soluble extracellular domain of the receptor, results in suppression of endogenous Pax6, Rx, and Otx2 expression and suppression of endogenous eye development. This effect can be rescued by coexpression of Xfz3. Finally, overexpression of Kermit, a protein that interacts with the C-terminal intracellular domain of Xfz3, also blocks endogenous eye development, suggesting that signaling through Xfz3 or a related receptor is required for normal eye development. In summary, we show that frizzled signaling is both necessary and sufficient to regulate eye development in Xenopus. PMID- 11274408 TI - How amoeboids self-organize into a fruiting body: multicellular coordination in Dictyostelium discoideum. AB - When individual amoebae of the cellular slime mold Dictyostelium discoideum are starving, they aggregate to form a multicellular migrating slug, which moves toward a region suitable for culmination. The culmination of the morphogenesis involves complex cell movements that transform a mound of cells into a globule of spores on a slender stalk. The movement has been likened to a "reverse fountain," whereby prestalk cells in the upper part form a stalk that moves downwards and anchors to the substratum, while prespore cells in the lower part move upwards to form the spore head. So far, however, no satisfactory explanation has been produced for this process. Using a computer simulation that we developed, we now demonstrate that the processes that are essential during the earlier stages of the morphogenesis are in fact sufficient to produce the dynamics of the culmination stage. These processes are cAMP signaling, differential adhesion, cell differentiation, and production of extracellular matrix. Our model clarifies the processes that generate the observed cell movements. More specifically, we show that periodic upward movements, caused by chemotactic motion, are essential for successful culmination, because the pressure waves they induce squeeze the stalk downwards through the cell mass. The mechanisms revealed by our model have a number of self-organizing and self-correcting properties and can account for many previously unconnected and unexplained experimental observations. PMID- 11274407 TI - The dual role of ultraspiracle, the Drosophila retinoid X receptor, in the ecdysone response. AB - The Drosophila homolog of the retinoid X receptor, ultraspiracle (USP), heterodimerizes with the ecdysone receptor (EcR) to form a functional complex that mediates the effects of the steroid molting hormone ecdysone by activating and repressing expression of ecdysone response genes. As with other retinoid X receptor heterodimers, EcR/USP affects gene transcription in a ligand-modulated manner. We used in vivo, cell culture, and biochemical approaches to analyze the functions of two usp alleles, usp(3) and usp(4), which encode stable proteins with defective DNA-binding domains. We observed that USP is able to activate as well as repress the Z1 isoform of the ecdysone-responsive broad complex (BrC-Z1). Activation of BrC-Z1 as well as EcR, itself an ecdysone response gene, can be mediated by both the USP3 and USP4 mutant proteins. USP3 and USP4 also activate an ecdysone-responsive element, hsp27EcRE, in cultured cells. These results differ from the protein null allele, usp(2), which is unable to mediate activation [Schubiger, M. & Truman, J. W. (2000) Development 127, 1151--1159]. BrC-Z1 repression is compromised in all three usp alleles, suggesting that repression involves the association of USP with DNA. Our results distinguish two mechanisms by which USP modulates the properties of EcR: one that involves the USP DNA-binding domain and one that can be achieved solely through the ligand binding domain. These newly revealed properties of USP might implicate similar properties for retinoid X receptor. PMID- 11274410 TI - Microbial phyllosphere populations are more complex than previously realized. AB - Phyllosphere microbial communities were evaluated on leaves of field-grown plant species by culture-dependent and -independent methods. Denaturing gradient gel electrophoresis (DGGE) with 16S rDNA primers generally indicated that microbial community structures were similar on different individuals of the same plant species, but unique on different plant species. Phyllosphere bacteria were identified from Citrus sinesis (cv. Valencia) by using DGGE analysis followed by cloning and sequencing of the dominant rDNA bands. Of the 17 unique sequences obtained, database queries showed only four strains that had been described previously as phyllosphere bacteria. Five of the 17 sequences had 16S similarities lower than 90% to database entries, suggesting that they represent previously undescribed species. In addition, three fungal species were also identified. Very different 16S rDNA DGGE banding profiles were obtained when replicate cv. Valencia leaf samples were cultured in BIOLOG EcoPlates for 4.5 days. All of these rDNA sequences had 97--100% similarity to those of known phyllosphere bacteria, but only two of them matched those identified by the culture independent DGGE analysis. Like other studied ecosystems, microbial phyllosphere communities therefore are more complex than previously thought, based on conventional culture-based methods. PMID- 11274409 TI - The GATA factor Serpent is required for the onset of the humoral immune response in Drosophila embryos. AB - Innate immunity in Drosophila is characterized by the inducible expression of antimicrobial peptides. We have investigated the development and regulation of immune responsiveness in Drosophila embryos after infection. Immune competence, as monitored by the induction of Cecropin A1-lacZ constructs, was observed first in the embryonic yolk. This observation suggests that the yolk plays an important role in the humoral immune response of the developing embryo by synthesizing antimicrobial peptides. Around midembryogenesis, the response in the yolk was diminished. Simultaneously, Cecropin expression became inducible in a large number of cells in the epidermis, demonstrating that late-stage embryos can synthesize their own antibiotics in the epidermis. This production likely serves to provide the hatching larva with an active antimicrobial barrier and protection against systemic infections. Cecropin expression in the yolk required the presence of a GATA site in the promoter as well as the involvement of the GATA binding transcription factor Serpent (dGATAb). In contrast, neither the GATA site nor Serpent were necessary for Cecropin expression in the epidermis. Thus, the inducible immune responses in the yolk and in the epidermis can be uncoupled and call for distinct sets of transcription factors. Our data suggest that Serpent is involved in the distinction between a systemic response in the yolk/fat body and a local immune response in epithelial cells. In addition, the present study shows that signal transduction pathways controlling innate and epithelial defense reactions can be dissected genetically in Drosophila embryos. PMID- 11274411 TI - Gregarious behavior in desert locusts is evoked by touching their back legs. AB - Desert locusts in the solitarious phase were repeatedly touched on various body regions to identify the site of mechanosensory input that elicits the transition to gregarious phase behavior. The phase state of individual insects was measured after a 4-h period of localized mechanical stimulation, by using a behavioral assay based on multiple logistic regression analysis. A significant switch from solitarious to gregarious behavior occurred when the outer face of a hind femur had been stimulated, but mechanical stimulation of 10 other body regions did not result in significant behavioral change. We conclude that a primary cause of the switch in behavior that seeds the formation of locust swarms is individuals regularly touching others on the hind legs within populations that have become concentrated by the environment. PMID- 11274412 TI - A nonspecific fatty acid within the bumblebee mating plug prevents females from remating. AB - The best mating strategy for males differs from that of females, because females gain from mating with several males (polyandry), but males gain from monopolizing the females. As a consequence, males have evolved a variety of methods, such as the transfer of inhibitory substances from their accessory glands, to ensure exclusive paternity of the female's offspring, generally with detrimental effects on female fitness. Inhibitory substances have been identified as peptides or other specific molecules. Unfortunately, in social insects male-mating traits are investigated only poorly, although male social insects might have the same fundamental influence on female-mating behavior as found in other species. A recently developed technique for the artificial insemination of bumblebee queens allowed us to investigate which chemical compound in the mating plug of male bumblebees, Bombus terrestris L., prevents females (queens) from further mating. Surprisingly, we found that the active substance is linoleic acid, a ubiquitous and rather unspecific fatty acid. Contrary to mating plugs in other insect species, the bumblebee mating plug is highly efficient and allows the males to determine queen-mating frequencies. PMID- 11274413 TI - Deletion of cytosolic phospholipase A(2) suppresses Apc(Min)-induced tumorigenesis. AB - Although nonsteroidal antiinflammatory drugs (NSAIDs) show great promise as therapies for colon cancer, a dispute remains regarding their mechanism of action. NSAIDs are known to inhibit cyclooxygenase (COX) enzymes, which convert arachidonic acid (AA) to prostaglandins (PGs). Therefore, NSAIDs may suppress tumorigenesis by inhibiting PG synthesis. However, various experimental studies have suggested the possibility of PG-independent mechanisms. Notably, disruption of the mouse group IIA secretory phospholipase A(2) locus (Pla2g2a), a potential source of AA for COX-2, increases tumor number despite the fact that the mutation has been predicted to decrease PG production. Some authors have attempted to reconcile the results by suggesting that the level of the precursor (AA), not the products (PGs), is the critical factor. To clarify the role of AA in tumorigenesis, we have examined the effect of deleting the group IV cytosolic phospholipase A(2) (cPLA(2)) locus (Pla2g4). We report that Apc(Min/+), cPLA(2)( /-) mice show an 83% reduction in tumor number in the small intestine compared with littermates with genotypes Apc(Min/+), cPLA(2)(+/-) and Apc(Min/+), cPLA(2)(+/+). This tumor phenotype parallels that of COX-2 knockout mice, suggesting that cPLA(2) is the predominant source of AA for COX-2 in the intestine. The protective effect of cPLA(2) deletion is thus most likely attributed to a decrease in the AA supply to COX-2 and a resultant decrease in PG synthesis. The tumorigenic effect of sPLA(2) mutations is likely to be through a completely different pathway. PMID- 11274414 TI - Visualization of oligonucleotide probes and point mutations in interphase nuclei and DNA fibers using rolling circle DNA amplification. AB - Rolling circle amplification (RCA) is a surface-anchored DNA replication reaction that can be exploited to visualize single molecular recognition events. Here we report the use of RCA to visualize target DNA sequences as small as 50 nts in peripheral blood lymphocytes or in stretched DNA fibers. Three unique target sequences within the cystic fibrosis transmembrane conductance regulator gene could be detected simultaneously in interphase nuclei, and could be ordered in a linear map in stretched DNA. Allele-discriminating oligonucleotide probes in conjunction with RCA also were used to discriminate wild-type and mutant alleles in the cystic fibrosis transmembrane conductance regulator, p53, BRCA-1, and Gorlin syndrome genes in the nuclei of cultured cells or in DNA fibers. These observations demonstrate that signal amplification by RCA can be coupled to nucleic acid hybridization and multicolor fluorescence imaging to detect single nucleotide changes in DNA within a cytological context or in single DNA molecules. This provides a means for direct physical haplotyping and the analysis of somatic mutations on a cell-by-cell basis. PMID- 11274415 TI - A large-scale overexpression screen in Saccharomyces cerevisiae identifies previously uncharacterized cell cycle genes. AB - We have undertaken an extensive screen to identify Saccharomyces cerevisiae genes whose products are involved in cell cycle progression. We report the identification of 113 genes, including 19 hypothetical ORFs, which confer arrest or delay in specific compartments of the cell cycle when overexpressed. The collection of genes identified by this screen overlaps with those identified in loss-of-function cdc screens but also includes genes whose products have not previously been implicated in cell cycle control. Through analysis of strains lacking these hypothetical ORFs, we have identified a variety of new CDC and checkpoint genes. PMID- 11274416 TI - Etoposide induces heritable chromosomal aberrations and aneuploidy during male meiosis in the mouse. AB - Etoposide, a topoisomerase II inhibitor widely used in cancer therapy, is suspected of inducing secondary tumors and affecting the genetic constitution of germ cells. A better understanding of the potential heritable risk of etoposide is needed to provide sound genetic counseling to cancer patients treated with this drug in their reproductive years. We used a mouse model to investigate the effects of clinical doses of etoposide on the induction of chromosomal abnormalities in spermatocytes and their transmission to zygotes by using a combination of chromosome painting and 4',6-diamidino-2-phenylindole staining. High frequencies of chromosomal aberrations were detected in spermatocytes within 64 h after treatment when over 30% of the metaphases analyzed had structural aberrations (P < 0.01). Significant increases in the percentages of zygotic metaphases with structural aberrations were found only for matings that sampled treated pachytene (28-fold, P < 0.0001) and preleptotene spermatocytes (13-fold, P < 0.001). Etoposide induced mostly acentric fragments and deletions, types of aberrations expected to result in embryonic lethality, because they represent loss of genetic material. Chromosomal exchanges were rare. Etoposide treatment of pachytene cells induced aneuploidy in both spermatocytes (18-fold, P < 0.01) and zygotes (8-fold, P < 0.05). We know of no other report of an agent for which paternal exposure leads to an increased incidence of aneuploidy in the offspring. Thus, we found that therapeutic doses of etoposide affect primarily meiotic germ cells, producing unstable structural aberrations and aneuploidy, effects that are transmitted to the progeny. This finding suggests that individuals who undergo chemotherapy with etoposide may be at a higher risk for abnormal reproductive outcomes especially within the 2 months after chemotherapy. PMID- 11274417 TI - Mutations in Drosophila heat shock cognate 4 are enhancers of Polycomb. AB - The homeotic genes controlling segment identity in Drosophila are repressed by the Polycomb group of genes (PcG) and are activated by genes of the trithorax group (trxG). An F(1) screen for dominant enhancers of Polycomb yielded a point mutation in the heat shock cognate gene, hsc4, along with mutations corresponding to several known PcG loci. The new mutation is a more potent enhancer of Polycomb phenotypes than an apparent null allele of hsc4 is, although even the null allele occasionally displays homeotic phenotypes associated with the PcG. Previous biochemical results had suggested that HSC4 might interact with BRAHMA, a trxG member. Further analyses now show that there is no physical or genetic interaction between HSC4 and the Brahma complex. HSC4 might be needed for the proper folding of a component of the Polycomb repression complex, or it may be a functional member of that complex. PMID- 11274418 TI - Substance P antagonist (CP-96,345) inhibits HIV-1 replication in human mononuclear phagocytes. AB - Substance P (SP) is a potent modulator of neuroimmunoregulation. We recently reported that human immune cells express SP and its receptor. We have now investigated the possible role that SP and its receptor plays in HIV infection of human mononuclear phagocytes. SP enhanced HIV replication in human blood-isolated mononuclear phagocytes, whereas the nonpeptide SP antagonist (CP-96,345) potently inhibited HIV infectivity of these cells in a concentration-dependent fashion. CP 96,345 prevented the formation of typical giant syncytia induced by HIV Bal strain replication in these cells. This inhibitory effect of CP-96,345 was because of the antagonism of neurokinin-1 receptor, a primary SP receptor. Both CP-96,345 and anti-SP antibody inhibited SP-enhanced HIV replication in monocyte derived macrophages (MDM). Among HIV strains tested (both prototype and primary isolates), only the R5 strains (Bal, ADA, BL-6, and CSF-6) that use the CCR5 coreceptor for entry into MDM were significantly inhibited by CP-96,345; in contrast, the X4 strain (UG024), which uses CXCR4 as its coreceptor, was not inhibited. In addition, the M-tropic ADA (CCR5-dependent)-pseudotyped HIV infection of MDM was markedly inhibited by CP-96,345, whereas murine leukemia virus-pseudotyped HIV was not affected, indicating that the major effect of CP 96,345 is regulated by Env-determined early events in HIV infection of MDM. CP 96,345 significantly down-regulated CCR5 expression in MDM at both protein and mRNA levels. Thus, SP-neurokinin-1 receptor interaction may play an important role in the regulation of CCR5 expression in MDM, affecting the R5 HIV strain infection of MDM. PMID- 11274419 TI - L-selectin can facilitate metastasis to lymph nodes in a transgenic mouse model of carcinogenesis. AB - L-selectin mediates homing of lymphocytes to lymph nodes (LN). Transgenic mice that express rat insulin promoter regulated simian virus 40 Tag (RIP-Tag) develop large, local cancers that metastasize to liver but not LN. To test whether this lack of LN metastases reflects their absence from the circulation, transgenic mice were produced that express Tag (T), L-selectin (L), and Escherichia coli LacZ (Z), in pancreatic beta cells. LTZ mice developed insulinomas that specifically had LN metastases; metastasis was blocked by an anti L-selectin mAb. LacZ(+) tumor cells from these LN homed to secondary LN upon transfer. These results suggest that the highly vascularized islet carcinomas are shedding tumor cells into the bloodstream, which is a necessary but insufficient condition for metastasis to occur; L-selectin can facilitate homing of such tumor cells to LN, resulting in metastasis. PMID- 11274421 TI - Cytomegalovirus in autoimmunity: T cell crossreactivity to viral antigen and autoantigen glutamic acid decarboxylase. AB - Antigens of pathogenic microbes that mimic autoantigens are thought to be responsible for the activation of autoreactive T cells. Viral infections have been associated with the development of the neuroendocrine autoimmune diseases type 1 diabetes and stiff-man syndrome, but the mechanism is unknown. These diseases share glutamic acid decarboxylase (GAD65) as a major autoantigen. We screened synthetic peptide libraries dedicated to bind to HLA-DR3, which predisposes to both diseases, using clonal CD4(+) T cells reactive to GAD65 isolated from a prediabetic stiff-man syndrome patient. Here we show that these GAD65-specific T cells crossreact with a peptide of the human cytomegalovirus (hCMV) major DNA-binding protein. This peptide was identified after database searching with a recognition pattern that had been deduced from the library studies. Furthermore, we showed that hCMV-derived epitope can be naturally processed by dendritic cells and recognized by GAD65 reactive T cells. Thus, hCMV may be involved in the loss of T cell tolerance to autoantigen GAD65 by a mechanism of molecular mimicry leading to autoimmunity. PMID- 11274420 TI - Mobilization of MHC class I molecules from late endosomes to the cell surface following activation of CD34-derived human Langerhans cells. AB - Langerhans cells are a subset of dendritic cells (DCs) found in the human epidermis with unique morphological and molecular properties that enable their function as "sentinels" of the immune system. DCs are pivotal in the initiation and regulation of primary MHC class I restricted T lymphocyte immune responses and are able to present both endogenous and exogenous antigen onto class I molecules. Here, we study the MHC class I presentation pathway following activation of immature, CD34-derived human Langerhans cells by lipopolysaccharide (LPS). LPS induces an increase in all components of the MHC class I pathway including the transporter for antigen presentation (TAP), tapasin and ERp57, and the immunoproteasome subunits LMP2 and LMP7. Moreover, in CD34-derived Langerhans cells, the rapid increase in expression of MHC class I molecules seen at the cell surface following LPS activation is because of mobilization of MHC class I molecules from HLA-DM positive endosomal compartments, a pathway not seen in monocyte-derived DCs. Mobilization of class I from this compartment is primaquine sensitive and brefeldin A insensitive. These data demonstrate the regulation of the class I pathway in concert with the maturation of the CD34-derived Langerhans cells and suggest potential sites for antigen loading of class I proteins. PMID- 11274422 TI - The immunoprotective MHC II epitope of a chemically induced tumor harbors a unique mutation in a ribosomal protein. AB - CD4(+) T lymphocyte clones, generated from mice immunized with the methylcholanthrene-induced fibrosarcoma Meth A (H-2(d)), are restricted by I-E(d) and recognize a unique antigen on Meth A. The antigen has been purified and characterized as the ribosomal protein L11. The antigenic epitope is contained within the sequence EYELRKHNFSDTG and is generated by substitution of Asn by His (italic) caused by a single point mutation. The tumor contains the wild-type and the mutated alleles. Immunization of BALB/cJ mice with the mutated epitope but not with the wild-type epitope protects mice against a subsequent challenge with the Meth A sarcoma. Adoptive transfer of CD4(+) clones into BALB/c mice renders the mice specifically resistant to Meth A sarcoma. The mutated L11 epitope is thus shown to be an immunoprotective epitope in vivo by several criteria. PMID- 11274423 TI - An unliganded thyroid hormone receptor causes severe neurological dysfunction. AB - Congenital hypothyroidism and the thyroid hormone (T(3)) resistance syndrome are associated with severe central nervous system (CNS) dysfunction. Because thyroid hormones are thought to act principally by binding to their nuclear receptors (TRs), it is unexplained why TR knock-out animals are reported to have normal CNS structure and function. To investigate this discrepancy further, a T(3) binding mutation was introduced into the mouse TR-beta locus by homologous recombination. Because of this T(3) binding defect, the mutant TR constitutively interacts with corepressor proteins and mimics the hypothyroid state, regardless of the circulating thyroid hormone concentrations. Severe abnormalities in cerebellar development and function and abnormal hippocampal gene expression and learning were found. These findings demonstrate the specific and deleterious action of unliganded TR in the brain and suggest the importance of corepressors bound to TR in the pathogenesis of hypothyroidism. PMID- 11274424 TI - Methylenetetrahydrofolate reductase (MTHFR) polymorphisms and risk of molecularly defined subtypes of childhood acute leukemia. AB - Low folate intake as well as alterations in folate metabolism as a result of polymorphisms in the enzyme methylenetetrahydrofolate reductase (MTHFR) have been associated with an increased incidence of neural tube defects, vascular disease, and some cancers. Polymorphic variants of MTHFR lead to enhanced thymidine pools and better quality DNA synthesis that could afford some protection from the development of leukemias, particularly those with translocations. We now report associations of MTHFR polymorphisms in three subgroups of pediatric leukemias: infant lymphoblastic or myeloblastic leukemias with MLL rearrangements and childhood lymphoblastic leukemias with either TEL-AML1 fusions or hyperdiploid karyotypes. Pediatric leukemia patients (n = 253 total) and healthy newborn controls (n = 200) were genotyped for MTHFR polymorphisms at nucleotides 677 (C- >T) and 1,298 (A-->C). A significant association for carriers of C677T was demonstrated for leukemias with MLL translocations (MLL+, n = 37) when compared with controls [adjusted odd ratios (OR) = 0.36 with a 95% confidence interval (CI) of 0.15-0.85; P = 0.017]. This protective effect was not evident for A1298C alleles (OR = 1.14). In contrast, associations for A1298C homozygotes (CC; OR = 0.26 with a 95% CI of 0.07--0.81) and C677T homozygotes (TT; OR = 0.49 with a 95% CI of 0.20--1.17) were observed for hyperdiploid leukemias (n = 138). No significant associations were evident for either polymorphism with TEL-AML1+ leukemias (n = 78). These differences in allelic associations may point to discrete attributes of the two alleles in their ability to alter folate and one carbon metabolite pools and impact after DNA synthesis and methylation pathways, but should be viewed cautiously pending larger follow-up studies. The data provide evidence that molecularly defined subgroups of pediatric leukemias have different etiologies and also suggest a role of folate in the development of childhood leukemia. PMID- 11274425 TI - Mammalian Scratch: a neural-specific Snail family transcriptional repressor. AB - Members of the Snail family of zinc finger transcription factors are known to play critical roles in neurogenesis in invertebrates, but none of these factors has been linked to vertebrate neuronal differentiation. We report the isolation of a gene encoding a mammalian Snail family member that is restricted to the nervous system. Human and murine Scratch (Scrt) share 81% and 69% identity to Drosophila Scrt and the Caenorhabditis elegans neuronal antiapoptotic protein, CES-1, respectively, across the five zinc finger domain. Expression of mammalian Scrt is predominantly confined to the brain and spinal cord, appearing in newly differentiating, postmitotic neurons and persisting into postnatal life. Additional expression is seen in the retina and, significantly, in neuroendocrine (NE) cells of the lung. In a parallel fashion, we detect hScrt expression in lung cancers with NE features, especially small cell lung cancer. hScrt shares the capacity of other Snail family members to bind to E-box enhancer motifs, which are targets of basic helix--loop--helix (bHLH) transcription factors. We show that hScrt directly antagonizes the function of heterodimers of the proneural bHLH protein achaete-scute homolog-1 and E12, leading to active transcriptional repression at E-box motifs. Thus, Scrt has the potential to function in newly differentiating, postmitotic neurons and in cancers with NE features by modulating the action of bHLH transcription factors critical for neuronal differentiation. PMID- 11274426 TI - Muscle-specific mutations accumulate with aging in critical human mtDNA control sites for replication. AB - The recently discovered aging-dependent large accumulation of point mutations in the human fibroblast mtDNA control region raised the question of their occurrence in postmitotic tissues. In the present work, analysis of biopsied or autopsied human skeletal muscle revealed the absence or only minimal presence of those mutations. By contrast, surprisingly, most of 26 individuals 53 to 92 years old, without a known history of neuromuscular disease, exhibited at mtDNA replication control sites in muscle an accumulation of two new point mutations, i.e., A189G and T408A, which were absent or marginally present in 19 individuals younger than 34 years. These two mutations were not found in fibroblasts from 22 subjects 64 to 101 years of age (T408A), or were present only in three subjects in very low amounts (A189G). Furthermore, in several older individuals exhibiting an accumulation in muscle of one or both of these mutations, they were nearly absent in other tissues, whereas the most frequent fibroblast-specific mutation (T414G) was present in skin, but not in muscle. Among eight additional individuals exhibiting partial denervation of their biopsied muscle, four subjects >80 years old had accumulated the two muscle-specific point mutations, which were, conversely, present at only very low levels in four subjects < or =40 years old. The striking tissue specificity of the muscle mtDNA mutations detected here and their mapping at critical sites for mtDNA replication strongly point to the involvement of a specific mutagenic machinery and to the functional relevance of these mutations. PMID- 11274427 TI - A feedback-controlled ensemble model of the stress-responsive hypothalamo pituitary-adrenal axis. AB - The present work develops and implements a biomathematical statement of how reciprocal connectivity drives stress-adaptive homeostasis in the corticotropic (hypothalamo-pituitary-adrenal) axis. In initial analyses with this interactive construct, we test six specific a priori hypotheses of mechanisms linking circadian (24-h) rhythmicity to pulsatile secretory output. This formulation offers a dynamic framework for later statistical estimation of unobserved in vivo neurohormone secretion and within-axis, dose-responsive interfaces in health and disease. Explication of the core dynamics of the stress-responsive corticotropic axis based on secure physiological precepts should help to unveil new biomedical hypotheses of stressor-specific system failure. PMID- 11274428 TI - B lymphocyte-restricted expression of prion protein does not enable prion replication in prion protein knockout mice. AB - Prion replication in spleen and neuroinvasion after i.p. inoculation of mice is impaired in forms of immunodeficiency where mature B lymphocytes are lacking. In spleens of wild-type mice, infectivity is associated with B and T lymphocytes and stroma but not with circulating lymphocytes. We generated transgenic prion protein knockout mice overexpressing prion protein in B lymphocytes and found that they failed to accumulate prions in spleen after i.p. inoculation. We conclude that splenic B lymphocytes are not prion-replication competent and that they acquire prions from other cells, most likely follicular dendritic cells with which they closely associate and whose maturation depends on them. PMID- 11274429 TI - A serologically identified tumor antigen encoded by a homeobox gene promotes growth of ovarian epithelial cells. AB - Ovarian carcinomas are thought to arise from cells of the ovarian surface epithelium by mechanisms that are poorly understood. Molecules associated with neoplasia are potentially immunogenic, but few ovarian tumor antigens have been identified. Because ovarian carcinomas can elicit humoral responses in patients, we searched for novel tumor antigens by immunoscreening a cDNA expression library with ovarian cancer patient serum. Seven clones corresponding to the homeobox gene HOXB7 were isolated. ELISAs using purified recombinant HOXB7 protein revealed significant serologic reactivity to HOXB7 in 13 of 39 ovarian cancer patients and in only one of 29 healthy women (P < 0.0001). Ovarian carcinomas were found to express HOXB7 at markedly higher levels than normal ovarian surface epithelium, suggesting that immunogenicity of HOXB7 in patients could be associated with its elevated expression in ovarian carcinomas. Overexpression of HOXB7 in immortalized normal ovarian surface epithelial cells dramatically enhanced cellular proliferation. Furthermore, HOXB7 overexpression increased intracellular accumulation and secretion of basic fibroblast growth factor, a potent angiogenic and mitogenic factor. These results reveal HOXB7 as a tumor antigen whose up-regulated expression could play a significant role in promoting growth and development of ovarian carcinomas. PMID- 11274430 TI - Increased CNS levels of apolipoprotein D in schizophrenic and bipolar subjects: implications for the pathophysiology of psychiatric disorders. AB - Chronic administration of the atypical antipsychotic drug, clozapine, to rodents has been shown to increase the concentration of apolipoprotein D (apoD) in several area of the brain, suggesting that apoD could be involved in the therapeutic effects of antipsychotic drugs and/or the pathology of psychotic illnesses. Here, we measured a significant decrease in the concentration of apoD in serum samples from schizophrenic patients. In contrast, apoD levels were significantly increased (92--287%) in dorsolateral prefrontal cortex (Brodmann's area 9) of schizophrenic and bipolar subjects. Elevated levels of apoD expression were also observed in the caudate of schizophrenic and bipolar subjects (68- 89%). No differences in apoD immunoreactivity were detected in occipital cortex (Brodmann's area 18) in either group, or in the hippocampus, substantia nigra, or cerebellum of the schizophrenic group. The low serum concentrations of apoD observed in these patients supports recent hypotheses involving systemic insufficiencies in lipid metabolism/signaling in schizophrenia. Elevation of apoD expression selectively within central nervous system regions implicated in the pathology of these neuropsychiatric disorders suggests a focal compensatory response that neuroleptic drug regimens may augment. PMID- 11274431 TI - Defect in regulated secretion of P-selectin affects leukocyte recruitment in von Willebrand factor-deficient mice. AB - Stimulation of endothelial cells by various inflammatory mediators leads to release of Weibel--Palade bodies and therefore to exocytosis of both P-selectin (adhesion receptor for leukocytes) and von Willebrand factor (vWf) (platelet ligand). The potential role of vWf in leukocyte recruitment was investigated with the use of vWf-deficient mice. We report a strong reduction of leukocyte rolling in venules of vWf-deficient mice. Similarly, vWf deficiency led to a decrease in neutrophil recruitment in a cytokine-induced meningitis model as well as in early skin wounds. In all instances with an antibody that preferentially recognizes plasma membrane P-selectin, we observed a dramatic reduction in P-selectin expression at the cell surface of vWf-deficient endothelium. With confocal microscopy, we found that the typical rodlike shape of the Weibel--Palade body is missing in vWf -/- endothelial cells and that part of the P-selectin content in the vWf -/- cells colocalized with LAMP-1, a lysosomal marker. However, intracellular P-selectin levels were similar in tumor necrosis factor alpha- and lipopolysaccharide-activated cells of both genotypes. We conclude that the absence of vWf, as found in severe von Willebrand disease, leads to a defect in Weibel--Palade body formation. This defect results in decreased P-selectin translocation to the cell surface and reduced leukocyte recruitment in early phases of inflammation. PMID- 11274432 TI - Local gene transfer of tissue factor pathway inhibitor regulates intimal hyperplasia in atherosclerotic arteries. AB - Tissue factor (TF), the initiator of blood coagulation and thrombosis, is up regulated after vascular injury and in atherosclerotic states. Systemic administration of recombinant TF pathway inhibitor (TFPI) has been reported to decrease intimal hyperplasia after vascular injury and also to suppress systemic mechanisms of blood coagulation and thrombosis. Here we report that, in heritable hyperlipidemic Watanabe rabbits, adenoviral gene transfer of TFPI to balloon injured atherosclerotic arteries reduced the extent of intimal hyperplasia by 43% (P < 0.05) compared with a control vector used at identical titer (1 x 10(10) plaque-forming units/ml). Platelet aggregation and coagulation studies performed 7 days after local gene transfer of TFPI failed to show any impairment in systemic hemostasis. At time of sacrifice, 4 weeks after vascular injury, the 10 Ad-TFPI treated carotid arteries were free of thrombi, whereas two control treated arteries were occluded (P, not significant). These findings suggest that TFPI overexpressed in atherosclerotic arteries can regulate hyperplastic response to injury in the absence of changes in the hemostatic system, establishing a role for local TF regulation as target for gene transfer-based antirestenosis therapies. PMID- 11274433 TI - Effect of DNA-dependent protein kinase on the molecular fate of the rAAV2 genome in skeletal muscle. AB - We report here that the DNA-dependent protein kinase (DNA-PK) affects the molecular fate of the recombinant adeno-associated virus (rAAV) genome in skeletal muscle. rAAV-human alpha1-antitrypsin (rAAV-hAAT) vectors were delivered by intramuscular injection to either C57BL/6 (DNA-PKcs(+)) or C57BL/6-SCID [severe combined immunodeficient (SCID), DNA-PKcs(-)] mice. In both strains, high levels of transgene expression were sustained for up to 1 year after a single injection. Southern blot analysis showed that rAAV genomes persisted as linear episomes for more than 1 year in SCID mice, whereas only circular episomal forms were observed in the C57BL/6 strain. These results indicate that DNA-PK is involved in the formation of circular rAAV episomes. PMID- 11274434 TI - Dynamic localization of a cytoplasmic signal transduction response regulator controls morphogenesis during the Caulobacter cell cycle. AB - We present evidence that a bacterial signal transduction cascade that couples morphogenesis with cell cycle progression is regulated by dynamic localization of its components. Previous studies have implicated two histidine kinases, DivJ and PleC, and the response regulator, DivK, in the regulation of morphogenesis in the dimorphic bacterium Caulobacter crescentus. Here, we show that the cytoplasmic response regulator, DivK, exhibits a dynamic, cyclical localization that culminates in asymmetric distribution of DivK within the two cell types that are characteristic of the Caulobacter cell cycle; DivK is dispersed throughout the cytoplasm of the progeny swarmer cell and is localized to the pole of the stalked cell. The membrane-bound DivJ and PleC histidine kinases, which are asymmetrically localized at the opposite poles of the predivisional cell, control the temporal and spatial localization of DivK. DivJ mediates DivK targeting to the poles whereas PleC controls its release from one of the poles at times and places that are consistent with the activities and location of DivJ and PleC in the late predivisional cell. Thus, dynamic changes in subcellular location of multiple components of a signal transduction cascade may constitute a novel mode of prokaryotic regulation to generate and maintain cellular asymmetry. PMID- 11274435 TI - Ribozymes that cleave reovirus genome segment S1 also protect cells from pathogenesis caused by reovirus infection. AB - Reovirus genome segment S1 encodes protein final sigma1, which is the receptor binding protein, modulates tissue tropism, and specifies the nature of the antiviral immune response. It makes up less than 2% of reovirus particles and is synthesized in very small amounts in infected cells. Any antiviral strategy aimed at reducing specifically the expression of this genome segment should, in principle, reduce the infectivity of the virus. To test this hypothesis, we have assembled two hammer-head motif-containing ribozymes (Rzs) targeted to cleave at the conserved B and C domains of the reovirus s1 RNA. Protein-independent but Mg(2+)-dependent sequence-specific cleavage of s1 RNA was achieved by both the Rzs in trans. Cells that transiently express these Rzs, when challenged with reovirus, were protected against the cytopathic effects caused by the virus. This protection correlated with the specific intracellular reduction of s1 transcripts that was due to their cleavage by the Rzs. Rz-treated cells that were challenged with reovirus showed almost complete disappearance of protein final sigma1 without significantly altering the levels of the other reovirus structural proteins. Thus, Rzs, besides acting as antiviral agents, could be exploited as biological tools to delineate specific functions of target genes. PMID- 11274436 TI - Modular organization of the Friend murine leukemia virus envelope protein underlies the mechanism of infection. AB - Retrovirus infection is initiated by receptor-dependent fusion of the envelope to the cell membrane. The modular organization of the envelope protein of C type retroviruses has been exploited to investigate how binding of the surface subunit (SU) to receptor triggers fusion mediated by the transmembrane (TM) subunit. We show that deletion of the receptor-binding domain (RBD) from SU of Friend murine leukemia virus (Fr-MLV) abolishes infection that is restored by supplying RBD as a soluble protein. Infection by this mechanism remains dependent on receptor expression. When membrane attachment of the virus lacking RBD is reestablished by inserting the hormone erythropoietin, infection remains dependent on the RBD/receptor complex. However, infection increases 50-fold to 5 x 10(5) units/ml on cells that also express the erythropoietin receptor. Soluble RBD from Fr-MLV also restores infection by amphotropic and xenotropic MLVs in which RBD is deleted. These experiments demonstrate that RBD has two functions: mediating virus attachment and activating the fusion mechanism. In addition, they indicate that receptor engagement triggers fusion by promoting a subgroup-independent functional interaction between RBD and the remainder of SU and/or TM. PMID- 11274437 TI - Activation of latent Kaposi's sarcoma-associated herpesvirus by demethylation of the promoter of the lytic transactivator. AB - Kaposi's sarcoma-associated herpesvirus (KSHV) is strongly linked to Kaposi's sarcoma, primary effusion lymphomas, and a subset of multicentric Castleman's disease. The mechanism by which this virus establishes latency and reactivation is unknown. KSHV Lyta (lytic transactivator, also named KSHV/Rta), mainly encoded by the ORF 50 gene, is a lytic switch gene for viral reactivation from latency, inasmuch as it is both essential and sufficient to drive the entire viral lytic cycle. Here we show that the Lyta promoter region was heavily methylated in latently infected cells. Treatment of primary effusion lymphoma-delivered cell lines with tetradecanoylphorbol acetate caused demethylation of the Lyta promoter and induced KSHV lytic phase in vitro. Methylation cassette assay shows demethylation of the Lyta promoter region was essential for the expression of Lyta. In vivo, biopsy samples obtained from patients with KSHV-related diseases show the most demethylation in the Lyta promoter region, whereas samples from a latently infected KSHV carrier remained in a methylated status. These results suggest a relationship among a demethylation status in the Lyta promoter, the reactivation of KSHV, and the development of KSHV-associated diseases. PMID- 11274438 TI - Efficient use of a small genome to generate antigenic diversity in tick-borne ehrlichial pathogens. AB - Ehrlichiae are responsible for important tick-transmitted diseases, including anaplasmosis, the most prevalent tick-borne infection of livestock worldwide, and the emerging human diseases monocytic and granulocytic ehrlichiosis. Antigenic variation of major surface proteins is a key feature of these pathogens that allows persistence in the mammalian host, a requisite for subsequent tick transmission. In Anaplasma marginale pseudogenes for two antigenically variable gene families, msp2 and msp3, appear in concert. These pseudogenes can be recombined into the functional expression site to generate new antigenic variants. Coordinated control of the recombination of these genes would allow these two gene families to act synergistically to evade the host immune response. PMID- 11274439 TI - Formation of temporal-feature maps by axonal propagation of synaptic learning. AB - Computational maps are of central importance to a neuronal representation of the outside world. In a map, neighboring neurons respond to similar sensory features. A well studied example is the computational map of interaural time differences (ITDs), which is essential to sound localization in a variety of species and allows resolution of ITDs of the order of 10 micros. Nevertheless, it is unclear how such an orderly representation of temporal features arises. We address this problem by modeling the ontogenetic development of an ITD map in the laminar nucleus of the barn owl. We show how the owl's ITD map can emerge from a combined action of homosynaptic spike-based Hebbian learning and its propagation along the presynaptic axon. In spike-based Hebbian learning, synaptic strengths are modified according to the timing of pre- and postsynaptic action potentials. In unspecific axonal learning, a synapse's modification gives rise to a factor that propagates along the presynaptic axon and affects the properties of synapses at neighboring neurons. Our results indicate that both Hebbian learning and its presynaptic propagation are necessary for map formation in the laminar nucleus, but the latter can be orders of magnitude weaker than the former. We argue that the algorithm is important for the formation of computational maps, when, in particular, time plays a key role. PMID- 11274440 TI - Differences in quantal amplitude reflect GluR4- subunit number at corticothalamic synapses on two populations of thalamic neurons. AB - Low-frequency thalamocortical oscillations that underlie drowsiness and slow-wave sleep depend on rhythmic inhibition of relay cells by neurons in the reticular nucleus (RTN) under the influence of corticothalamic fibers that branch to innervate RTN neurons and relay neurons. To generate oscillations, input to RTN predictably should be stronger so disynaptic inhibition of relay cells overcomes direct corticothalamic excitation. Amplitudes of excitatory postsynaptic conductances (EPSCs) evoked in RTN neurons by minimal stimulation of corticothalamic fibers were 2.4 times larger than in relay neurons, and quantal size of RTN EPSCs was 2.6 times greater. GluR4-receptor subunits labeled at corticothalamic synapses on RTN neurons outnumbered those on relay cells by 3.7 times, providing a basis for differences in synaptic strength. PMID- 11274441 TI - Reciprocal electromechanical properties of rat prestin: the motor molecule from rat outer hair cells. AB - Cochlear outer hair cells (OHCs) are responsible for the exquisite sensitivity, dynamic range, and frequency-resolving capacity of the mammalian hearing organ. These unique cells respond to an electrical stimulus with a cycle-by-cycle change in cell length that is mediated by molecular motors in the cells' basolateral membrane. Recent work identified prestin, a protein with similarity to pendrin related anion transporters, as the OHC motor molecule. Here we show that heterologously expressed prestin from rat OHCs (rprestin) exhibits reciprocal electromechanical properties as known for the OHC motor protein. Upon electrical stimulation in the microchamber configuration, rprestin generates mechanical force with constant amplitude and phase up to a stimulus frequency of at least 20 kHz. Mechanical stimulation of rprestin in excised outside-out patches shifts the voltage dependence of the nonlinear capacitance characterizing the electrical properties of the molecule. The results indicate that rprestin is a molecular motor that displays reciprocal electromechanical properties over the entire frequency range relevant for mammalian hearing. PMID- 11274442 TI - Visual habit formation in monkeys with neurotoxic lesions of the ventrocaudal neostriatum. AB - Visual habit formation in monkeys, assessed by concurrent visual discrimination learning with 24-h intertrial intervals (ITI), was found earlier to be impaired by removal of the inferior temporal visual area (TE) but not by removal of either the medial temporal lobe or inferior prefrontal convexity, two of TE's major projection targets. To assess the role in this form of learning of another pair of structures to which TE projects, namely the rostral portion of the tail of the caudate nucleus and the overlying ventrocaudal putamen, we injected a neurotoxin into this neostriatal region of several monkeys and tested them on the 24-h ITI task as well as on a test of visual recognition memory. Compared with unoperated monkeys, the experimental animals were unaffected on the recognition test but showed an impairment on the 24-h ITI task that was highly correlated with the extent of their neostriatal damage. The findings suggest that TE and its projection areas in the ventrocaudal neostriatum form part of a circuit that selectively mediates visual habit formation. PMID- 11274443 TI - Chronic morphine induces the concomitant phosphorylation and altered association of multiple signaling proteins: a novel mechanism for modulating cell signaling. AB - Traditional mechanisms thought to underlie opioid tolerance include receptor phosphorylation/down-regulation, G-protein uncoupling, and adenylyl cyclase superactivation. A parallel line of investigation also indicates that opioid tolerance development results from a switch from predominantly opioid receptor G(i alpha) inhibitory to G(beta gamma) stimulatory signaling. As described previously, this results, in part, from the increased relative abundance of G(beta gamma)-stimulated adenylyl cyclase isoforms as well as from a profound increase in their phosphorylation [Chakrabarti, S., Rivera, M., Yan, S.-Z., Tang, W.-J. & Gintzler, A. R. (1998) Mol. Pharmacol. 54, 655-662; Chakrabarti, S., Wang, L., Tang, W.-J. & Gintzler, A. R. (1998) Mol. Pharmacol. 54, 949--953]. The present study demonstrates that chronic morphine administration results in the concomitant phosphorylation of three key signaling proteins, G protein receptor kinase (GRK) 2/3, beta-arrestin, and G(beta), in the guinea pig longitudinal muscle myenteric plexus tissue. Augmented phosphorylation of all three proteins is evident in immunoprecipitate obtained by using either anti-GRK2/3 or G(beta) antibodies, but the phosphorylation increment is greater in immunoprecipitate obtained with G(beta) antibodies. Analyses of coimmunoprecipitated proteins indicate that phosphorylation of GRK2/3, beta-arrestin, and G(beta) has varying consequences on their ability to associate. As a result, increased availability of and signaling via G(beta gamma) could occur without compromising the membrane content (and presumably activity) of GRK2/3. Induction of the concomitant phosphorylation of multiple proteins in a multimolecular complex with attendant modulation of their association represents a novel mechanism for increasing G(beta gamma) signaling and opioid tolerance formation. PMID- 11274444 TI - Excitation--contraction uncoupling by a human central core disease mutation in the ryanodine receptor. AB - Central core disease (CCD) is a human congenital myopathy characterized by fetal hypotonia and proximal muscle weakness that is linked to mutations in the gene encoding the type-1 ryanodine receptor (RyR1). CCD is thought to arise from Ca(2+)-induced damage stemming from mutant RyR1 proteins forming "leaky" sarcoplasmic reticulum (SR) Ca(2+) release channels. A novel mutation in the C terminal region of RyR1 (I4898T) accounts for an unusually severe and highly penetrant form of CCD in humans [Lynch, P. J., Tong, J., Lehane, M., Mallet, A., Giblin, L., Heffron, J. J., Vaughan, P., Zafra, G., MacLennan, D. H. & McCarthy, T. V. (1999) Proc. Natl. Acad. Sci. USA 96, 4164--4169]. We expressed in skeletal myotubes derived from RyR1-knockout (dyspedic) mice the analogous mutation engineered into a rabbit RyR1 cDNA (I4897T). Here we show that homozygous expression of I4897T in dyspedic myotubes results in a complete uncoupling of sarcolemmal excitation from voltage-gated SR Ca(2+) release without significantly altering resting cytosolic Ca(2+) levels, SR Ca(2+) content, or RyR1-mediated enhancement of dihydropyridine receptor (DHPR) channel activity. Coexpression of both I4897T and wild-type RyR1 resulted in a 60% reduction in voltage-gated SR Ca(2+) release, again without altering resting cytosolic Ca(2+) levels, SR Ca(2+) content, or DHPR channel activity. These findings indicate that muscle weakness suffered by individuals possessing the I4898T mutation involves a functional uncoupling of sarcolemmal excitation from SR Ca(2+) release, rather than the expression of overactive or leaky SR Ca(2+) release channels. PMID- 11274446 TI - Flexibility of the Kir6.2 inward rectifier K(+) channel pore. AB - Interactions of sulfhydryl reagents with introduced cysteines in the pore-forming (Kir6.2) subunits of the K(ATP) channel were examined. 2-Aminoethyl methanethiosulfonate (MTSEA(+)) failed to modify Cd(2+)-insensitive control Kir6.2 channels, but rapidly and irreversibly modified Kir6.2[L164C] (L164C) channels. Although a single Cd(2+) ion is coordinated by L164C, four MTSEA(+) "hits" can occur, each sequentially reducing the single-channel current. A dimeric fusion of control-Kir6.2 and L164C subunits generates Cd(2+)-insensitive channels, confirming that at least three cysteines are required for coordination, but MTSEA(+) modification of the dimer occurs in two hits. L164C channels were not modified by bromotrimethyl ammoniumbimane (qBBr(+)), even though qBBr(+) caused voltage-dependent block (as opposed to modification) that was comparable to that of MTSEA(+) or 3-(triethylammonium)propyl methanethiosulfonate (MTSPTrEA(+)), implying that qBBr(+) can also enter the inner cavity but does not modify L164C residues. The Kir channel pore structure was modeled by homology with the KcsA crystal structure. A stable conformation optimally places the four L164C side chains for coordination of a single Cd(2+) ion. Modification of these cysteines by up to four MTSEA(+) (or three MTSPTrEA(+), or two qBBr(+)) does not require widening of the cavity to accommodate the derivatives within it. However, like the KcsA crystal structure, the energy-minimized model shows a narrowing at the inner entrance, and in the Kir6.2 model this narrowing excludes all ions. To allow entry of ions as large as MTSPTrEA(+) or qBBr(+), the entrance must widen to >8 A, but this widening is readily accomplished by minimal M2 helix motion and side-chain rearrangement. PMID- 11274447 TI - VIPP1, a nuclear gene of Arabidopsis thaliana essential for thylakoid membrane formation. AB - The conversion of light to chemical energy by the process of photosynthesis is localized to the thylakoid membrane network in plant chloroplasts. Although several pathways have been described that target proteins into and across the thylakoids, little is known about the origin of this membrane system or how the lipid backbone of the thylakoids is transported and fused with the target membrane. Thylakoid biogenesis and maintenance seem to involve the flow of membrane elements via vesicular transport. Here we show by mutational analysis that deletion of a single gene called VIPP1 (vesicle-inducing protein in plastids 1) is deleterious to thylakoid membrane formation. Although VIPP1 is a hydrophilic protein it is found in both the inner envelope and the thylakoid membranes. In VIPP1 deletion mutants vesicle formation is abolished. We propose that VIPP1 is essential for the maintenance of thylakoids by a transport pathway not previously recognized. PMID- 11274445 TI - Pendrin, encoded by the Pendred syndrome gene, resides in the apical region of renal intercalated cells and mediates bicarbonate secretion. AB - Pendrin is an anion transporter encoded by the PDS/Pds gene. In humans, mutations in PDS cause the genetic disorder Pendred syndrome, which is associated with deafness and goiter. Previous studies have shown that this gene has a relatively restricted pattern of expression, with PDS/Pds mRNA detected only in the thyroid, inner ear, and kidney. The present study examined the distribution and function of pendrin in the mammalian kidney. Immunolocalization studies were performed using anti-pendrin polyclonal and monoclonal antibodies. Labeling was detected on the apical surface of a subpopulation of cells within the cortical collecting ducts (CCDs) that also express the H(+)-ATPase but not aquaporin-2, indicating that pendrin is present in intercalated cells of the CCD. Furthermore, pendrin was detected exclusively within the subpopulation of intercalated cells that express the H(+)-ATPase but not the anion exchanger 1 (AE1) and that are thought to mediate bicarbonate secretion. The same distribution of pendrin was observed in mouse, rat, and human kidney. However, pendrin was not detected in kidneys from a Pds-knockout mouse. Perfused CCD tubules isolated from alkali-loaded wild type mice secreted bicarbonate, whereas tubules from alkali-loaded Pds-knockout mice failed to secrete bicarbonate. Together, these studies indicate that pendrin is an apical anion transporter in intercalated cells of CCDs and has an essential role in renal bicarbonate secretion. PMID- 11274448 TI - Vipp1 deletion mutant of Synechocystis: a connection between bacterial phage shock and thylakoid biogenesis? AB - Plant chloroplasts originated from an endosymbiotic event by which an ancestor of contemporary cyanobacteria was engulfed by an early eukaryotic cell and then transformed into an organelle. Oxygenic photosynthesis is the specific feature of cyanobacteria and chloroplasts, and the photosynthetic machinery resides in an internal membrane system, the thylakoids. The origin and genesis of thylakoid membranes, which are essential for oxygenic photosynthesis, are still an enigma. Vipp1 (vesicle-inducing protein in plastids 1) is a protein located in both the inner envelope and the thylakoids of Pisum sativum and Arabidopsis thaliana. In Arabidopsis disruption of the VIPP1 gene severely affects the plant's ability to form properly structured thylakoids and as a consequence to carry out photosynthesis. In contrast, Vipp1 in Synechocystis appears to be located exclusively in the plasma membrane. Yet, as in higher plants, disruption of the VIPP1 gene locus leads to the complete loss of thylakoid formation. So far VIPP1 genes are found only in organisms carrying out oxygenic photosynthesis. They share sequence homology with a subunit encoded by the bacterial phage shock operon (PspA) but differ from PspA by a C-terminal extension of about 30 amino acids. In two cyanobacteria, Synechocystis and Anabaena, both a VIPP1 and a pspA gene are present, and phylogenetic analysis indicates that VIPP1 originated from a gene duplication of the latter and thereafter acquired its new function. It also appears that the C-terminal extension that discriminates VIPP1 proteins from PspA is important for its function in thylakoid formation. PMID- 11274449 TI - 14-3-3 protein is a regulator of the mitochondrial and chloroplast ATP synthase. AB - Mitochondrial and chloroplast ATP synthases are key enzymes in plant metabolism, providing cells with ATP, the universal energy currency. ATP synthases use a transmembrane electrochemical proton gradient to drive synthesis of ATP. The enzyme complexes function as miniature rotary engines, ensuring energy coupling with very high efficiency. Although our understanding of the structure and functioning of the synthase has made enormous progress in recent years, our understanding of regulatory mechanisms is still rather preliminary. Here we report a role for 14-3-3 proteins in the regulation of ATP synthases. These 14-3 3 proteins are highly conserved phosphoserine/phosphothreonine-binding proteins that regulate a wide range of enzymes in plants, animals, and yeast. Recently, the presence of 14-3-3 proteins in chloroplasts was illustrated, and we show here that plant mitochondria harbor 14-3-3s within the inner mitochondrial-membrane compartment. There, the 14-3-3 proteins were found to be associated with the ATP synthases, in a phosphorylation-dependent manner, through direct interaction with the F(1) beta-subunit. The activity of the ATP synthases in both organelles is drastically reduced by recombinant 14-3-3. The rapid reduction in chloroplast ATPase activity during dark adaptation was prevented by a phosphopeptide containing the 14-3-3 interaction motif, demonstrating a role for endogenous 14-3 3 in the down-regulation of the CF(o)F(1) activity. We conclude that regulation of the ATP synthases by 14-3-3 represents a mechanism for plant adaptation to environmental changes such as light/dark transitions, anoxia in roots, and fluctuations in nutrient supply. PMID- 11274450 TI - Futile transmembrane NH4(+) cycling: a cellular hypothesis to explain ammonium toxicity in plants. AB - Most higher plants develop severe toxicity symptoms when grown on ammonium (NH(4)(+)) as the sole nitrogen source. Recently, NH(4)(+) toxicity has been implicated as a cause of forest decline and even species extinction. Although mechanisms underlying NH(4)(+) toxicity have been extensively sought, the primary events conferring it at the cellular level are not understood. Using a high precision positron tracing technique, we here present a cell-physiological characterization of NH(4)(+) acquisition in two major cereals, barley (Hordeum vulgare), known to be susceptible to toxicity, and rice (Oryza sativa), known for its exceptional tolerance to even high levels of NH(4)(+). We show that, at high external NH(4)(+) concentration ([NH(4)(+)](o)), barley root cells experience a breakdown in the regulation of NH(4)(+) influx, leading to the accumulation of excessive amounts of NH(4)(+) in the cytosol. Measurements of NH(4)(+) efflux, combined with a thermodynamic analysis of the transmembrane electrochemical potential for NH(4)(+), reveal that, at elevated [NH(4)(+)](o), barley cells engage a high-capacity NH(4)(+)-efflux system that supports outward NH(4)(+) fluxes against a sizable gradient. Ammonium efflux is shown to constitute as much as 80% of primary influx, resulting in a never-before-documented futile cycling of nitrogen across the plasma membrane of root cells. This futile cycling carries a high energetic cost (we record a 40% increase in root respiration) that is independent of N metabolism and is accompanied by a decline in growth. In rice, by contrast, a cellular defense strategy has evolved that is characterized by an energetically neutral, near-Nernstian, equilibration of NH(4)(+) at high [NH(4)(+)](o). Thus our study has characterized the primary events in NH(4)(+) nutrition at the cellular level that may constitute the fundamental cause of NH(4)(+) toxicity in plants. PMID- 11274451 TI - Seeing properties of an invisible object: feature inheritance and shine-through. AB - We characterize a class of spatio-temporal illusions with two complementary properties. Firstly, if a vernier stimulus is flashed for a short time on a monitor and is followed immediately by a grating, the latter can express features of the vernier, such as its offset, its orientation, or its motion (feature inheritance). Yet the vernier stimulus itself remains perceptually invisible. Secondly, the vernier can be rendered visible by presenting gratings with a larger number of elements (shine-through). Under these conditions, subjects perceive two independent "objects" each carrying their own features. Transition between these two domains can be effected by subtle changes in the spatio temporal layout of the grating. This should allow psychophysicists and electrophysiologists to investigate feature binding in a precise and quantitative manner. PMID- 11274459 TI - Analysis of inhibitor binding in influenza virus neuraminidase. AB - 2,3-didehydro-2-deoxy-N:-acetylneuraminic acid (DANA) is a transition state analog inhibitor of influenza virus neuraminidase (NA). Replacement of the hydroxyl at the C9 position in DANA and 4-amino-DANA with an amine group, with the intention of taking advantage of an increased electrostatic interaction with a conserved acidic group in the active site to improve inhibitor binding, significantly reduces the inhibitor activity of both compounds. The three dimensional X-ray structure of the complexes of these ligands and NA was obtained to 1.4 A resolution and showed that both ligands bind isosterically to DANA. Analysis of the geometry of the ammonium at the C4 position indicates that Glu119 may be neutral when these ligands bind. A computational analysis of the binding energies indicates that the substitution is successful in increasing the energy of interaction; however, the gains that are made are not sufficient to overcome the energy that is required to desolvate that part of the ligand that comes in contact with the protein. PMID- 11274458 TI - A refined solution structure of hen lysozyme determined using residual dipolar coupling data. AB - A high resolution NMR structure of hen lysozyme has been determined using 209 residual 1H-15N dipolar coupling restraints from measurements made in two different dilute liquid crystalline phases (bicelles) in conjunction with a data set of 1632 NOE distance restraints, 110 torsion angle restraints, and 60 hydrogen bond restraints. The ensemble of 50 low-energy calculated structures has an average backbone RMSD of 0.50+/-0.13A to the mean structure and of 1.49+/ 0.10A to the crystal structure of hen lysozyme. To assess the importance of the dipolar coupling data in the structure determination, the final structures are compared with an ensemble calculated using an identical protocol but excluding the dipolar coupling restraints. The comparison shows that structures calculated with the dipolar coupling data are more similar to the crystal structure than those calculated without, and have better stereochemical quality. The structures also show improved quality factors when compared with additional dipolar coupling data that were not included in the structure calculations, with orientation dependent 15N chemical shift changes measured in the bicelle solutions, and with T1/T2 values obtained from 15N relaxation measurements. Analysis of the ensemble of NMR structures and comparisons with crystal structures, 15N relaxation data, and molecular dynamics simulations of hen lysozyme provides a detailed description of the solution structure of this protein and insights into its dynamical behavior. PMID- 11274460 TI - Three-dimensional structures of the three human class I alcohol dehydrogenases. AB - In contrast with other animal species, humans possess three distinct genes for class I alcohol dehydrogenase and show polymorphic variation in the ADH1B and ADH1C genes. The three class I alcohol dehydrogenase isoenzymes share approximately 93% sequence identity but differ in their substrate specificity and their developmental expression. We report here the first three-dimensional structures for the ADH1A and ADH1C*2 gene products at 2.5 and 2.0 A, respectively, and the structure of the ADH1B*1 gene product in a binary complex with cofactor at 2.2 A. Not surprisingly, the overall structure of each isoenzyme is highly similar to the others. However, the substitution of Gly for Arg at position 47 in the ADH1A isoenzyme promotes a greater extent of domain closure in the ADH1A isoenzyme, whereas substitution at position 271 may account for the lower turnover rate for the ADH1C*2 isoenzyme relative to its polymorphic variant, ADH1C*1. The substrate-binding pockets of each isoenzyme possess a unique topology that dictates each isoenzyme's distinct but overlapping substrate preferences. ADH1*B1 has the most restrictive substrate-binding site near the catalytic zinc atom, whereas both ADH1A and ADH1C*2 possess amino acid substitutions that correlate with their better efficiency for the oxidation of secondary alcohols. These structures describe the nature of their individual substrate-binding pockets and will improve our understanding of how the metabolism of beverage ethanol affects the normal metabolic processes performed by these isoenzymes. PMID- 11274461 TI - Catalytic center of an archaeal type 2 ribonuclease H as revealed by X-ray crystallographic and mutational analyses. AB - The catalytic center of an archaeal Type 2 RNase H has been identified by a combination of X-ray crystallographic and mutational analyses. The crystal structure of the Type 2 RNase H from Thermococcus kodakaraensis KOD1 has revealed that the N-terminal major domain adopts the RNase H fold, despite the poor sequence similarity to the Type 1 RNase H. Mutational analyses showed that the catalytic reaction requires four acidic residues, which are well conserved in the Type 1 RNase H and the members of the polynucleotidyl transferase family. Thus, the Type 1 and Type 2 RNases H seem to share a common catalytic mechanism, except for the requirement of histidine as a general base in the former enzyme. Combined with the results from deletion mutant analyses, the structure suggests that the C terminal domain of the Type 2 RNase H is involved in the interaction with the DNA/RNA hybrid. PMID- 11274462 TI - Amino-acid substitutions at the fully exposed P1 site of bovine pancreatic trypsin inhibitor affect its stability. AB - It is widely accepted that solvent-exposed sites in proteins play only a negligible role in determining protein energetics. In this paper we show that amino acid substitutions at the fully exposed Lys15 in bovine pancreatic trypsin inhibitor (BPTI) influenced the CD- and DSC-monitored stability: The T(den) difference between the least (P1 Trp) and the most stable (P1 His) mutant is 11.2 degrees C at pH 2.0. The DeltaH(den) versus T(den) plot for all the variants at three pH values (2.0, 2.5, 3.0) is linear (DeltaC(p,den) = 0.41 kcal* mole(-1) * K(-1); 1 cal = 4.18 J) leading to a DeltaG(den) difference of 2.1 kcal*mole(-1). Thermal denaturation of the variants monitored by CD signal at pH 2.0 in the presence of 6 M GdmCl again showed differences in their stability, albeit somewhat smaller (DeltaT(den) =7.1 degrees C). Selective reduction of the Cys14 Cys 38 disulfide bond, which is located in the vicinity of the P1 position did not eliminate the stability differences. A correlation analysis of the P1 stability with different properties of amino acids suggests that two mechanisms may be responsible for the observed stability differences: the reverse hydrophobic effect and amino acid propensities to occur in nonoptimal dihedral angles adopted by the P1 position. The former effect operates at the denatured state level and causes a drop in protein stability for hydrophobic side chains, due to their decreased exposure upon denaturation. The latter factor influences the native state energetics and results from intrinsic properties of amino acids in a way similar to those observed for secondary structure propensities. In conclusion, our results suggest that the protein-stability-derived secondary structure propensity scales should be taken with more caution. PMID- 11274463 TI - Pressure versus temperature unfolding of ribonuclease A: an FTIR spectroscopic characterization of 10 variants at the carboxy-terminal site. AB - FTIR spectroscopy was used to characterize and compare the temperature- and pressure-induced unfolding of ribonuclease A and a set of its variants engineered in a hydrophobic region of the C-terminal part of the molecule postulated as a CFIS. The results show for all the ribonucleases investigated, a cooperative, two state, reversible unfolding transition using both pressure and temperature. The relative stabilities, among the different sites and different variants at the same site, monitored either through the changes in the position of the maximum of the amide I' band and the tyrosine band, or the maximum of the band assigned to the beta-sheet structure, corroborate the results of a previous study using fourth-derivative UV absorbance spectroscopy. In addition, variants at position 108 are the most critical for ribonuclease structure and stability. The V108G variant seems to present a greater conformational flexibility than the other variants. The pressure- and temperature-denaturated states of all the ribonucleases characterized retained some secondary structure. However, their spectral maxima were centered at different wavenumbers, which suggests that pressure- and temperature-denaturated states do not have the same structural characteristics. Nevertheless, there was close correlation between the pressure and temperature midpoint transition values for the whole series of protein variants, which indicated a common tendency of stability toward pressure and heat. PMID- 11274464 TI - Reactivity of peptidyl-tyrosine to hydroxylation and cross-linking. AB - Tyrosine residues of neuroendocrine peptides are frequently the targets of oxidation reactions, one of which involves hydroxylation to peptidyl-3, 4 dihydroxy-phenyl-L-alanine (DOPA). The reactivity in vitro of peptidyl-DOPA in two neuroendocrine peptides, a neurotensin fragment (pELYENK) and proctolin (RYLPT), was investigated using ultraviolet-visible scanning spectrophotometry and matrix-assisted laser desorption ionization mass spectrometry following oxidation by tyrosinase and periodate. The peptides form covalently coupled dimers and trimers, and their masses are consistent with the presence of diDOPA cross-links. Lysine does not appear to participate in multimer formation because it is efficiently recovered in fragmentation ladders using subtilisin. While multimer formation in the neurotensin-derived peptide can be blocked effectively by adding N-acetyl-DOPA-ethylester to the reaction medium, the DOPA ethylester couples itself four to five times to each peptide. PMID- 11274465 TI - Roles of dimerization in folding and stability of ketosteroid isomerase from Pseudomonas putida biotype B. AB - Equilibrium and kinetic analyses have been performed to elucidate the roles of dimerization in folding and stability of KSI from Pseudomonas putida biotype B. Folding was reversible in secondary and tertiary structures as well as in activity. Equilibrium unfolding transition, as monitored by fluorescence and ellipticity measurements, could be modeled by a two-state mechanism without thermodynamically stable intermediates. Consistent with the two-state model, one dimensional (1D) NMR spectra and gel-filtration chromatography analysis did not show any evidence for a folded monomeric intermediate. Interestingly enough, Cys 81 located at the dimeric interface was modified by DTNB before unfolding. This inconsistent result might be explained by increased dynamic motion of the interface residues in the presence of urea to expose Cys 81 more frequently without the dimer dissociation. The refolding process, as monitored by fluorescence change, could best be described by five kinetic phases, in which the second phase was a bimolecular step. Because <30% of the total fluorescence change occurred during the first step, most of the native tertiary structure may be driven to form by the bimolecular step. During the refolding process, negative ellipticity at 225 nm increased very fast within 80 msec to account for >80% of the total amplitude. This result suggests that the protein folds into a monomer containing most of the alpha-helical structures before dimerization. Monitoring the enzyme activity during the refolding process could estimate the activity of the monomer that is not fully active. Together, these results stress the importance of dimerization in the formation and maintenance of the functional native tertiary structure. PMID- 11274466 TI - Chemical characteristics of dimer interfaces in the legume lectin family. AB - The Erythrina corallodendron lectin (EcorL) crystallizes in monoclinic and hexagonal crystal forms. Comparison of the newly determined hexagonal form (PDB code 1fyu) with the monoclinic form shows that the dimeric structure of EcorL reflects the inherent biological structure of the protein and is not an artifact of the crystal packing. To further understand the factors determining the dimerization modes of legume lectins, EcorL, concanavalin A (ConA), and Griffonia simplicifolia (GS4) were taken as representatives of the three unique dimers found in the family. Six virtual homodimers were generated. The hydropathy, amino acid composition, and solvation energy were calculated for all nine homodimers. Each of the three native dimers has a distinct chemical composition. EcorL has a dominant hydrophobic component, and ConA has a strong polar component, but in GS4 the three components contribute equally to the interface. This distribution pattern at the interface is unique to the native dimers and distinct from the partition observed in the virtual dimers. Amino acid composition of other members of the family that dimerize like EcorL or ConA maintain the same pattern of amino acids distribution observed in EcorL and ConA. However, lectins that dimerize like GS4 do not show a particularly distinct distribution. In all cases, the calculated solvation energy of the native dimer was lower than that of the virtual dimers, suggesting that the observed mode of dimerization is the most stable organization for the given sequence and tertiary structure. The dimerization type cannot be predicted by sequence analysis. PMID- 11274467 TI - Magnetization transfer from laser-polarized xenon to protons located in the hydrophobic cavity of the wheat nonspecific lipid transfer protein. AB - Nonspecific lipid transfer protein from wheat is studied by liquid-state NMR in the presence of xenon. The gas-protein interaction is indicated by the dependence of the protein proton chemical shifts on the xenon pressure and formally confirmed by the first observation of magnetization transfer from laser-polarized xenon to the protein protons. Twenty-six heteronuclear nOes have allowed the characterization of four interaction sites inside the wheat ns-LTP cavity. Their locations are in agreement with the variations of the chemical shifts under xenon pressure and with solvation simulations. The richness of the information obtained by the noble gas with a nuclear polarization multiplied by approximately 12,000 makes this approach based on dipolar cross-relaxation with laser-polarized xenon promising for probing protein hydrophobic pockets at ambient pressure. PMID- 11274468 TI - Folding units in calcium vector protein of amphioxus: Structural and functional properties of its amino- and carboxy-terminal halves. AB - Muscle of amphioxus contains large amounts of a four EF-hand Ca2+-binding protein, CaVP, and its target, CaVPT. To study the domain structure of CaVP and assess the structurally important determinants for its interaction with CaVPT, we expressed CaVP and its amino (N-CaVP) and carboxy-terminal halves (C-CaVP). The interactive properties of recombinant and wild-type CaVP are very similar, despite three post-translational modifications in the wild-type protein. N-CaVP does not bind Ca2+, shows a well-formed hydrophobic core, and melts at 44 degrees C. C-CaVP binds two Ca2+ with intrinsic dissociation constants of 0.22 and 140 microM (i.e., very similar to the entire CaVP). The metal-free domain in CaVP and C-CaVP shows no distinct melting transition, whereas its 1Ca2+ and 2Ca2+) forms melt in the 111 degrees -123 degrees C range, suggesting that C-CaVP and the carboxy- domain of CaVP are natively unfolded in the metal-free state and progressively gain structure upon binding of 1Ca2+ and 2Ca2+. Thermal denaturation studies provide evidence for interdomain interaction: the apo, 1Ca2+ and 2Ca2+ states of the carboxy-domain destabilize to different degrees the amino domain. Only C-CaVP forms a Ca2+-dependent 1:1 complex with CaVPT. Our results suggest that the carboxy-terminal domain of CaVP interacts with CaVPT and that the amino-terminal lobe modulates this interaction. PMID- 11274469 TI - Prediction of the transmembrane regions of beta-barrel membrane proteins with a neural network-based predictor. AB - A method based on neural networks is trained and tested on a nonredundant set of beta-barrel membrane proteins known at atomic resolution with a jackknife procedure. The method predicts the topography of transmembrane beta strands with residue accuracy as high as 78% when evolutionary information is used as input to the network. Of the transmembrane beta-strands included in the training set, 93% are correctly assigned. The predictor includes an algorithm of model optimization, based on dynamic programming, that correctly models eight out of the 11 proteins present in the training/testing set. In addition, protein topology is assigned on the basis of the location of the longest loops in the models. We propose this as a general method to fill the gap of the prediction of beta-barrel membrane proteins. PMID- 11274470 TI - Identification of related proteins with weak sequence identity using secondary structure information. AB - Molecular modeling of proteins is confronted with the problem of finding homologous proteins, especially when few identities remain after the process of molecular evolution. Using even the most recent methods based on sequence identity detection, structural relationships are still difficult to establish with high reliability. As protein structures are more conserved than sequences, we investigated the possibility of using protein secondary structure comparison (observed or predicted structures) to discriminate between related and unrelated proteins sequences in the range of 10%-30% sequence identity. Pairwise comparison of secondary structures have been measured using the structural overlap (Sov) parameter. In this article, we show that if the secondary structures likeness is >50%, most of the pairs are structurally related. Taking into account the secondary structures of proteins that have been detected by BLAST, FASTA, or SSEARCH in the noisy region (with high E: value), we show that distantly related protein sequences (even with <20% identity) can be still identified. This strategy can be used to identify three-dimensional templates in homology modeling by finding unexpected related proteins and to select proteins for experimental investigation in a structural genomic approach, as well as for genome annotation. PMID- 11274471 TI - Extending the capabilities of targeted molecular dynamics: simulation of a large conformational transition in plasminogen activator inhibitor 1. AB - Plasminogen activator inhibitor type 1 (PAI-1) is an inhibitor of plasminogen activators such as tissue-type plasminogen activator or urokinase-type plasminogen activator. For this molecule, different conformations are known. The inhibiting form that interacts with the proteinases is called the active form. The noninhibitory, noncleavable form is called the latent form. X-ray and modeling studies have revealed a large change in position of the reactive center loop (RCL), responsible for the interaction with the proteinases, that is inserted into a beta-sheet (s4A) in the latent form. The mechanism underlying this spontaneous conformational change (half-life = 2 h at 37 degrees C) is not known in detail. This investigation attempts to predict a transition path from the active to the latent structure at the atomic level, by using simulation techniques. Together with targeted molecular dynamics (TMD), a plausible assumption on a rigid body movement of the RCL was applied to define an initial guess for an intermediate. Different pathways were simulated, from the active to the intermediate, from the intermediate to the latent structure and vice versa under different conditions. Equilibrium simulations at different steps of the path also were performed. The results show that a continuous pathway from the active to the latent structure can be modeled. This study also shows that this approach may be applied in general to model large conformational changes in any kind of protein for which the initial and final three-dimensional structure is known. PMID- 11274472 TI - Computational estimation of specific side chain interaction energies in alpha helices. AB - We have used a structure energy-based computer program developed for protein design, Perla, to provide theoretical estimates of all specific side chain-side chain interaction energies occurring in alpha helices. The computed side chain side chain interaction energies were used as substitutes for the corresponding values used by the helix/coil transition algorithm, AGADIR. Predictions of peptide helical contents were nearly as successful as those obtained with the originally calibrated set of parameters; a correlation to experimentally observed alpha-helical populations of 0.91 proved that our theoretical estimates are reasonably correct for amino acid pairs that are frequent in our database of peptides. Furthermore, we have determined experimentally the previously uncharacterized interaction energies for Lys-Ile, Thr-Ile, and Phe-Ile amino acid pairs at i,i + 4 positions. The experimental values compare favorably with the computed theoretical estimates. Importantly, the computed values for Thr-Ile and Phe-Ile interactions are better than the energies based on chemical similarity, whereas for Lys-Ile they are similar. Thus, computational techniques can be used to provide precise energies for amino acid pairwise interactions, a fact that supports the development of structure energy-based computational tools for structure predictions and sequence design. PMID- 11274473 TI - Conformational propagation with prion-like characteristics in a simple model of protein folding. AB - Protein refolding/misfolding to an alternative form plays an aetiologic role in many diseases in humans, including Alzheimer's disease, the systemic amyloidoses, and the prion diseases. Here we have discovered that such refolding can occur readily for a simple lattice model of proteins in a propagatable manner without designing for any particular alternative native state. The model uses a simple contact energy function for interactions between residues and does not consider the peculiarities of polypeptide geometry. In this model, under conditions where the normal (N) native state is marginally stable or unstable, two chains refold from the N native state to an alternative multimeric energetic minimum comprising a single refolded conformation that can then propagate itself to other protein chains. The only requirement for efficient propagation is that a two-faced mode of packing must be in the ground state as a dimer (a higher-energy state for this packing leads to less efficient propagation). For random sequences, these ground state dimeric configurations tend to have more beta-sheet-like extended structure than almost any other sort of dimeric ground-state assembly. This implies that propagating states (such as for prions) are beta-sheet rich because the only likely propagating forms are beta-sheet rich. We examine the details of our simulations to see to what extent the observed properties of prion propagation can be predicted by a simple protein folding model. The formation of the alternative state in the present model shows several distinct features of amyloidogenesis and of prion propagation. For example, an analog of the phenomenon of conformationally distinct strains in prions is observed. We find a parallel between 'glassy' behavior in liquids and the formation of a propagatable state in proteins. This is the first report of simulation of conformational propagation using any heteropolymer model. The results imply that some (but not most) small protein sequences must maintain a sequence signal that resists refolding to propagatable alternative native states and that the ability to form such states is not limited to polypeptides (or reliant on regular hydrogen bonding per se) but can occur for other protein-like heteropolymers. PMID- 11274474 TI - Sequencing of the ddl gene and modeling of the mutated D-alanine:D-alanine ligase in glycopeptide-dependent strains of Enterococcus faecium. AB - Glycopeptide dependence for growth in enterococci results from mutations in the ddl gene that inactivate the host D-Ala:D-Ala ligase. The strains require glycopeptides as inducers for synthesis of resistance proteins, which allows for the production of peptidoglycan precursors ending in D-Ala-D-Lac instead of D-Ala D-Ala. The sequences of the ddl gene from nine glycopeptide-dependent Enterococcus faecium clinical isolates were determined. Each one had a mutation consisting either in a 5-bp insertion at position 41 leading to an early stop codon, an in-frame 6-bp deletion causing the loss of two residues (KDVA243-246 to KA), or single base-pair changes resulting in an amino acid substitution (E13 --> G, G99 --> R, V241 --> D, D295 --> G, P313 --> L). The potential consequences of the deletion and point mutations on the 3-D structure of the enzyme were evaluated by comparative molecular modeling of the E. faecium enzyme, using the X ray structure of the homologous Escherichia coli D-Ala:D-Ala ligase DdlB as a template. All mutated residues were found either to interact directly with one of the substrates of the enzymatic reaction (E13 and D295) or to stabilize the position of critical residues in the active site. Maintenance of the 3-D structure in the vicinity of these mutations in the active site appears critical for D-Ala:D-Ala ligase activity. PMID- 11274475 TI - Thermal and urea-induced unfolding in T7 RNA polymerase: calorimetry, circular dichroism and fluorescence study. AB - Structural changes in T7 RNA polymerase (T7RNAP) induced by temperature and urea have been studied over a wide range of conditions to obtain information about the structural organization and the stability of the enzyme. T7RNAP is a large monomeric enzyme (99 kD). Calorimetric studies of the thermal transitions in T7RNAP show that the enzyme consists of three cooperative units that may be regarded as structural domains. Interactions between these structural domains and their stability strongly depend on solvent conditions. The unfolding of T7RNAP under different solvent conditions induces a highly stable intermediate state that lacks specific tertiary interactions, contains a significant amount of residual secondary structure, and undergoes further cooperative unfolding at high urea concentrations. Circular dichroism (CD) studies show that thermal unfolding leads to an intermediate state that has increased beta-sheet and reduced alpha helix content relative to the native state. Urea-induced unfolding at 25 degrees C reveals a two-step process. The first transition centered near 3 M urea leads to a plateau from 3.5 to 5.0 M urea, followed by a second transition centered near 6.5 M urea. The CD spectrum of the enzyme in the plateau region, which is similar to that of the enzyme thermally unfolded in the absence of urea, shows little temperature dependence from 15 degrees to 60 degrees C. The second transition leads to a mixture of poly(Pro)II and unordered conformations. As the temperature increases, the ellipticity at 222 nm becomes more negative because of conversion of poly(Pro)II to the unordered conformation. Near-ultraviolet CD spectra at 25 degrees C at varying concentrations of urea are consistent with this picture. Both thermal and urea denaturation are irreversible, presumably because of processes that follow unfolding. PMID- 11274477 TI - Total chemical synthesis of human matrix Gla protein. AB - Human matrix Gla protein (MGP) is a vitamin K-dependent extracellular matrix protein that binds Ca2+ ions and that is involved in the prevention of vascular calcification. MGP is a 10.6-kD protein (84 amino acids) containing five gamma carboxyglutamic acid (Gla) residues and one disulfide bond. Studies of the mechanism by which MGP prevents calcification of the arterial media are hampered by the low solubility of the protein (<10 microg/mL). Because of solubility problems, processing of a recombinantly expressed MGP-fusion protein chimera to obtain MGP was unsuccessful. Here we describe the total chemical synthesis of MGP by tBoc solid-phase peptide synthesis (SPPS) and native chemical ligation. Peptide Tyr1-Ala53 was synthesized on a derivatized resin yielding a C-terminal thioester group. Peptide Cys54-Lys84 was synthesized on Lys-PAM resin yielding a C-terminal carboxylic acid. Subsequent native chemical ligation of the two peptides resulted in the formation of a native peptide bond between Ala53 and Cys54. Folding of the 1-84-polypeptide chain in 3 M guanidine (pH 8) resulted in a decrease of molecular mass from 10,605 to 10,603 (ESI-MS), representing the loss of two protons because of the formation of the Cys54-Cys60 internal disulfide bond. Like native MGP, synthetic MGP had the same low solubility when brought into aqueous buffer solutions with physiological salt concentrations, confirming its native like structure. However, the solubility of MGP markedly increased in borate buffer at pH 7.4 in the absence of sodium chloride. Ca2+ binding to MGP was confirmed by analytical HPLC, on which the retention time of MGP was reduced in the presence of CaCl2. Circular dichroism studies revealed a sharp increase in alpha-helicity at 0.2 mM CaCl2 that may explain the Ca2+ dependent shift in high-pressure liquid chromatography (HPLC)-retention time of MGP. In conclusion, facile and efficient chemical synthesis in combination with native chemical ligation yielded MGP preparations that can aid in unraveling the mechanism by which MGP prevents vascular calcification. PMID- 11274476 TI - Strain-specified relative conformational stability of the scrapie prion protein. AB - Studies of prion biology and diseases have elucidated several new concepts, but none was more heretical than the proposal that the biological properties that distinguish different prion strains are enciphered in the disease-causing prion protein (PrP(Sc)). To explore this postulate, we examined the properties of PrP(Sc) from eight prion isolates that propagate in Syrian hamster (SHa). Using resistance to protease digestion as a marker for the undenatured protein, we examined the conformational stabilities of these PrP(Sc) molecules. All eight isolates showed sigmoidal patterns of transition from native to denatured PrP(Sc) as a function of increasing guanidine hydrochloride (GdnHCl) concentration. Half maximal denaturation occurred at a mean value of 1.48 M GdnHCl for the Sc237, HY, SHa(Me7), and MT-C5 isolates, all of which have approximately 75-d incubation periods; a concentration of 1.08 M was found for the DY strain with a approximately 170-d incubation period and approximately 1.25 M for the SHa(RML) and 139H isolates with approximately 180-d incubation periods. A mean value of 1.39 M GdnHCl for the Me7-H strain with a approximately 320-d incubation period was found. Based on these results, the eight prion strains segregated into four distinct groups. Our results support the unorthodox proposal that distinct PrP(Sc) conformers encipher the biological properties of prion strains. PMID- 11274478 TI - Ligand binding to the inhibitory and stimulatory GTP cyclohydrolase I/GTP cyclohydrolase I feedback regulatory protein complexes. AB - GTP cyclohydrolase I feedback regulatory protein (GFRP) mediates feedback inhibition of GTP cyclohydrolase I activity by 6R-L-erythro-5,6,7,8 tetrahydrobiopterin (BH4), which is an essential cofactor for key enzymes producing catecholamines, serotonin, and nitric oxide as well as phenylalanine hydroxylase. GFRP also mediates feed-forward stimulation of GTP cyclohydrolase I activity by phenylalanine at subsaturating GTP levels. These ligands, BH4 and phenylalanine, induce complex formation between one molecule of GTP cyclohydrolase I and two molecules of GFRP. Here, we report the analysis of ligand binding using the gel filtration method of Hummel and Dreyer. BH4 binds to the GTP cyclohydrolase I/GFRP complex with a Kd of 4 microM, and phenylalanine binds to the protein complex with a Kd of 94 microM. The binding of BH4 is enhanced by dGTP. The binding stoichiometrics of BH4 and phenylalanine were estimated to be 10 molecules of each per protein complex, in other words, one molecule per subunit of protein, because GTP cyclohydrolase I is a decamer and GFRP is a pentamer. These findings were corroborated by data from equilibrium dialysis experiments. Regarding ligand binding to free proteins, BH4 binds weakly to GTP cyclohydrolase I but not to GFRP, and phenylalanine binds weakly to GFRP but not to GTP cyclohydrolase I. These results suggest that the overall structure of the protein complex contributes to binding of BH4 and phenylalanine but also that each binding site of BH4 and phenylalanine may be primarily composed of residues of GTP cyclohydrolase I and GFRP, respectively. PMID- 11274480 TI - Solubilization and disaggregation of polyglutamine peptides. AB - A method is described for dissolving and disaggregating chemically synthesized polyglutamine peptides. Polyglutamine peptides longer than about Q20 have been reported to be insoluble in water, but dissolution in--and evaporation from--a mixture of trifluoroacetic acid and hexafluoroisopropanol converts polyglutamine peptides up to at least Q44 to a form readily soluble in aqueous buffers. This procedure also has a dramatic effect on peptides which appear to be completely soluble in water, by removing traces of aggregate that seed aggregation. The protocol makes possible solution studies-including in vitro aggregation experiments--on polyglutamine peptides with repeat lengths associated with increased risk of Huntington's Disease and other expanded CAG repeat diseases. It may also be useful in conducting reproducible, quantitative aggregation studies on other polypeptides. PMID- 11274479 TI - Reduction of the amyloidogenicity of a protein by specific binding of ligands to the native conformation. AB - It is known that human muscle acylphosphatase (AcP) is able, under appropriate conditions in vitro, to aggregate and form amyloid fibrils of the type associated with human diseases. A number of compounds were tested for their ability to bind specifically to the native conformation of AcP under conditions favoring denaturation and subsequent aggregation and fibril formation. Compounds displaying different binding affinities for AcP were selected and their ability to inhibit protein fibrillization in vitro was evaluated. We found that compounds displaying a relatively high affinity for AcP are able to significantly delay protein fibrillization, mimicking the effect of stabilizing mutations; in addition, the effectiveness of such outcome correlates positively to both ligand concentration and affinity to the native state of AcP. By contrast, the inhibitory effect of ligands on AcP aggregation disappears in a mutant protein in which such binding affinity is lost. These results indicate that the stabilization of the native conformation of amyloidogenic proteins by specific ligand binding can be a strategy of general interest to inhibit amyloid formation in vivo. PMID- 11274481 TI - From proteases to proteomics. AB - This personal and professional autobiography covers the 50-yr period of 1950-2000 and includes the following topics: History of the University of Washington School of Medicine and its Department of Biochemistry (Mount Rainier and the University of Washington, recruiting faculty, biology, research programs); scientific editing (publication, Biochemistry, Protein Science, electronic publication); Europe revisited (Heidelberg, approaching retirement, the German Research Center, reunion in Vienna); and 50 yr of research on proteolytic enzymes (trypsin, carboxypeptidases, mast cell proteases, future developments). PMID- 11274482 TI - Fidelity, adherence, and robustness of interventions. PMID- 11274484 TI - Consumer & family information: substance abuse and addiction. PMID- 11274485 TI - Economic grand rounds: the costs of parity mandates for mental health and substance abuse insurance benefits. PMID- 11274487 TI - State health care reform: behavioral health services under Medicaid managed care: the uncertain implications of state variation. PMID- 11274486 TI - Datapoints: the role of computer use in different medical specialties. PMID- 11274488 TI - Law & psychiatry: mental health courts: their promise and unanswered questions. PMID- 11274489 TI - Managed care: strengthening the consumer voice in managed care: I. Can the private sector meet the public-sector standard? PMID- 11274490 TI - Personal accounts: luck of the draw. PMID- 11274491 TI - Implementing dual diagnosis services for clients with severe mental illness. AB - After 20 years of development and research, dual diagnosis services for clients with severe mental illness are emerging as an evidence-based practice. Effective dual diagnosis programs combine mental health and substance abuse interventions that are tailored for the complex needs of clients with comorbid disorders. The authors describe the critical components of effective programs, which include a comprehensive, long-term, staged approach to recovery; assertive outreach; motivational interventions; provision of help to clients in acquiring skills and supports to manage both illnesses and to pursue functional goals; and cultural sensitivity and competence. Many state mental health systems are implementing dual diagnosis services, but high-quality services are rare. The authors provide an overview of the numerous barriers to implementation and describe implementation strategies to overcome the barriers. Current approaches to implementing dual diagnosis programs involve organizational and financing changes at the policy level, clarity of program mission with structural changes to support dual diagnosis services, training and supervision for clinicians, and dissemination of accurate information to consumers and families to support understanding, demand, and advocacy. PMID- 11274492 TI - Mental health courts and the complex issue of mentally ill offenders. AB - Mental health courts are emerging in communities across the country to address the growing number of individuals with serious mental illness in jails and the complex issues they present to the courts. Based on concepts of therapeutic jurisprudence and patterned after drug courts, mental health courts attempt to prevent criminalization and recidivism by providing critical mental health services. The authors describe mental health courts in Broward County, Florida; King County, Washington; Anchorage, Alaska; and Marion County, Indiana. Each of these courts is designed to meet the specific needs and resources of its jurisdiction. The courts' experiences suggest that involving all players from the beginning is essential. The authors discuss the issues of due process, availability of services, and control of resources, which must be addressed before mental health courts are widely implemented. PMID- 11274493 TI - What is recovery? A conceptual model and explication. AB - This paper describes a conceptual model of recovery from mental illness developed to aid the state of Wisconsin in moving toward its goal of developing a "recovery oriented" mental health system. In the model, recovery refers to both internal conditions experienced by persons who describe themselves as being in recovery- hope, healing, empowerment, and connection--and external conditions that facilitate recovery--implementation of the principle of human rights, a positive culture of healing, and recovery-oriented services. The aim of the model is to link the abstract concepts that define recovery with specific strategies that systems, agencies, and individuals can use to facilitate it. PMID- 11274494 TI - What is recovery? A commentary. PMID- 11274495 TI - Insight into mental illness and child maltreatment risk among mothers with major psychiatric disorders. AB - OBJECTIVES: This study examined the relationship between insight into mental illness and current child maltreatment risk among mothers who had a major psychiatric disorder and who had lost custody of a child because of abuse, neglect, or having placed the child at risk of harm. Specifically, a measure of insight was examined in relation to systematically observed parenting behaviors known to be correlated with past child maltreatment and in relation to a comprehensive clinical determination of risk. METHODS: Forty-four mothers who had a major psychiatric disorder were independently rated for their insight into their illness, the quality of mother-child interaction, and the overall clinical risk of maltreatment. RESULTS: Better insight into mental illness was associated with more sensitive mothering behavior and with lower assessed clinical risk of maltreatment. The association remained when mothers with current psychotic symptoms were excluded from the analyses. Better insight did not appear to be associated with past psychotic symptoms, maternal psychiatric diagnosis, or the mother's level of education. CONCLUSIONS: Insight into mental illness may function as a protective factor that influences the risk of child maltreatment in mothers with mental illness. Measures of insight could be usefully incorporated into comprehensive parenting assessments for mothers with psychiatric disorders. PMID- 11274496 TI - Consumer-run service participation, recovery of social functioning, and the mediating role of psychological factors. AB - OBJECTIVE: This study examined the relationship between participation in consumer run services and recovery of social functioning among persons diagnosed as having serious mental illness. It also assessed the role of psychological factors in mediating this relationship. METHODS: Research questions investigated were whether involvement in consumer-run services is positively associated with recovery when premorbid and demographic factors are controlled for, whether psychological factors are positively associated with recovery irrespective of involvement in consumer-run services, and whether the relationship between involvement in consumer-run services and recovery is mediated by the psychological factors. The factors examined were self-efficacy, hopefulness, and active coping strategies. Sixty participants with a past or present diagnosis of schizophrenia, schizoaffective disorder, or bipolar disorder and at least one past psychiatric hospitalization were recruited from a community mental health center and two consumer-run programs. Data were collected on hopefulness, self efficacy, coping strategies, social functioning, and premorbid and demographic characteristics. RESULTS: Findings indicated that participants involved in consumer-run services had better social functioning than those involved only in traditional mental health services, that psychological variables were significantly associated with social functioning, and that the relationship between involvement in consumer-run services and social functioning was partly mediated by the use of more problem-centered coping strategies. Premorbid and demographic factors did not account for the relationship between psychosocial variables and social functioning, although education was a significant predictor of social functioning. CONCLUSIONS: The findings support the view that psychosocial factors may play a role in facilitating good community adjustment for individuals diagnosed as having serious mental illness. PMID- 11274497 TI - Six-month outcomes for patients who switched to olanzapine treatment. AB - OBJECTIVE: This study evaluated the outcomes of patients in a community mental health center who switched from treatment with another antipsychotic to olanzapine treatment. It also sought to determine whether simultaneous access to case management and psychosocial rehabilitation and olanzapine leads to enhanced functional improvement. METHODS: Six-month outcomes for a consecutive series of 104 patients who switched from a conventional antipsychotic medication to olanzapine were evaluated. Forty-nine patients in the same treatment program who continued to take conventional antipsychotics were also monitored as a reference group. Outcomes of the group receiving olanzapine were compared with their own baseline status and with outcomes of the reference group. RESULTS: At six months, patients in the olanzapine group demonstrated significant improvement over baseline across multiple measures of symptoms and psychosocial function. Compared with the reference group, the olanzapine group was more symptomatic at baseline and demonstrated significantly greater improvement at follow-up on the Brief Psychiatric Rating Scale and all subscales; Mini Psychiatric Rating Scale negative symptom, disorganization, anxiety, depression, and medication side effects items; and Clinical Global Improvement scale and Case Manager's Rating Scale-Plus illness factors. There was a trend toward superior improvement in psychosocial functioning among patients in the olanzapine group that achieved significance when patients in acute relapse at baseline were excluded. CONCLUSIONS: Olanzapine is effective in managing markedly to severely ill patients with psychotic disorders in a community mental health center. Simultaneous treatment with olanzapine, case management, and psychosocial rehabilitation leads to enhanced functional improvement among nonrelapsing patients. PMID- 11274498 TI - HIV risk behaviors and their relationship to posttraumatic stress disorder among women prisoners. AB - OBJECTIVE: This study assessed HIV risk behaviors and their association with psychiatric disorders among women prisoners. METHODS: HIV risk behaviors practiced in the five years before incarceration were ascertained with the Risk Behavior Assessment interview for 177 inmates at the Maryland Correctional Institution for Women. The Structured Clinical Interview for the DSM-IV was used to determine the occurrence of posttraumatic stress disorder (PTSD), major depression, and dysthymic disorder among the women. Regression models were used to determine the association between HIV risk behavior and psychiatric disorders. RESULTS: HIV risk behaviors in the five years before incarceration included never or rarely having used condoms (56 percent of the women), injection drug use (42 percent), sexual intercourse with a partner who used injection drugs (42 percent), prostitution (30 percent), needle sharing (30 percent), receptive anal sex (19 percent), and having more than 100 sex partners (7 percent). After the analysis adjusted for age, education, race, HIV status, and addictive disorders, a lifetime occurrence of PTSD was associated with the practice of anal sex (odds ratio=1.7; 95 percent confidence interval=1.26 to 2.16; p<.02) and prostitution (OR=1.56; 95% CI=1.17 to 1.95; p<.03). CONCLUSIONS: HIV risk behaviors before incarceration were highly prevalent among the women in this study. Rates of PTSD, depression, and dysthymic disorder were also high. PTSD was associated with prostitution and receptive anal sex, and the disorder may contribute to high rates of risky sexual behavior. Targeted HIV risk reduction efforts among women prisoners should include evaluation for PTSD; conversely, women prisoners with a diagnosis of PTSD should be evaluated for prior HIV sexual risk behaviors. PMID- 11274499 TI - Factors in the use of coercive retention in civil commitment evaluations in psychiatric emergency services. AB - OBJECTIVE: The authors examined whether factors other than civil commitment criteria influence the involuntary retention of patients who are evaluated for civil commitment in psychiatric emergency services in California general hospitals. METHODS: Logistic regression analysis was used to determine whether admission criteria, institutional constraints, social biases, and procedural justice indicators contributed to the use of coercive retention in the evaluations of 583 patients in the psychiatric emergency services of nine California county general hospitals. RESULTS: Of the 583 patients, 109 (18.7 percent) were retained against their wishes. Clinicians relied primarily on admission criteria in making the decision to retain a patient, which suggests that patients were generally afforded procedural due process during the evaluation in the psychiatric emergency service. Staff workload was a possible factor in violations of due process. CONCLUSIONS: Psychiatric emergency services need additional resources to ensure procedural due process protection for patients who are being evaluated for civil commitment. PMID- 11274500 TI - Ward crowding and incidents of violence on an acute psychiatric inpatient unit. AB - OBJECTIVE: Violence in psychiatric wards is common, and it is on the rise. This study examined the relationship between ward occupancy level and staff-to-patient ratio and incidents of aggressive behavior, both physical and verbal, on an acute inpatient unit in rural New Zealand. METHODS: Logistic regression was used to analyze data collected from the ward's log of adverse incidents and the ward census over a 12-month period. A physical incident was defined as an unwelcome physical contact or willful damage to property. Incidents of self-harm were excluded. Verbal aggression was any threat of physical or sexual harm. RESULTS: Among 381 admissions during the study period, 58 incidents were recorded--25 incidents of verbal aggression and 33 incidents of physical violence. Logistic regression demonstrated that the occupancy level was positively associated with the occurrence of any type of violent incident. The average occupancy level when an incident occurred was 77 percent, compared with 69 percent when no incidents occurred. The average occupancy level was significantly higher when verbal incidents occurred (80 percent) than when physical incidents occurred (70 percent). No association was found between violence and staff-to-patient ratio. Incidents were significantly more likely to occur during the afternoon shifts (3 p.m. to 11 p.m.). CONCLUSIONS: Crowding was found to be significantly associated with aggressive incidents, and in particular with verbal aggression. PMID- 11274501 TI - A comparison of long-term and short-term residential treatment programs for dual diagnosis patients. AB - The authors compared measures of process and six-month outcomes for 45 individuals who were treated in a long-term residential treatment program for patients with dual diagnoses with measures for 39 individuals who were treated in a short-term program. They also compared outcomes for individuals within each group. Those who received long-term treatment experienced improvements between entry into the program and six-month follow-up, and they were more likely to have engaged in treatment than individuals in the short-term group. At follow-up, individuals in the long-term residential treatment group were more likely to have maintained abstinence and less likely to have experienced homelessness than those in the short-term group. PMID- 11274502 TI - Fish consumption and depressive symptoms in the general population in Finland. AB - Fish contains high concentrations of omega-3 polyunsaturated fatty acids. Several studies have reported depletions of omega-3 fats among depressed patients, and a cross-national comparison has revealed a significant inverse correlation between annual prevalence of major depression and fish consumption. In a sample of 3,204 Finnish adults, depressive symptoms were estimated with the Beck Depression Inventory. A frequency question was used to measure fish consumption. Multiple logistic regression analysis was conducted to assess the association between depression and fish consumption. After the analysis adjusted for potential confounders, the likelihood of having depressive symptoms was significantly higher among infrequent fish consumers than among frequent consumers. PMID- 11274503 TI - An intensive outpatient program for patients with borderline personality disorder. AB - Intensive outpatient programs are designed to promote patients' functioning in the community by offering a more intensive level of structure and support than was previously available for outpatients. This paper describes the intensive outpatient program at McLean Hospital in Belmont, Massachusetts, which is tailored for patients with borderline personality disorder. These patients are susceptible to control struggles and regressive behaviors in more restrictive treatment settings. Through frequent contact with clinicians and other patients in this group-oriented program, patients with borderline personality disorders appear to feel sufficiently "held" and understood to develop their functional capacities as outpatients. PMID- 11274504 TI - Validity of self-reports about quality of life among patients with schizophrenia. AB - Lehman's Quality of Life Interview was administered to 22 patients with schizophrenia and their proxies and to 15 patients with cancer and their proxies. The results indicated that there was a discrepancy between responses on global objective and subjective measures for patients with schizophrenia but not for patients with cancer. A discrepancy was also found for the proxies of the patients with schizophrenia but not for the proxies of the patients with cancer. These findings suggest that the discrepancy between subjective and objective indicators of quality of life of patients with schizophrenia signifies a genuine difference rather than an anomaly related to the patient's psychiatric condition. PMID- 11274505 TI - Roles of psychiatrists on multidisciplinary mental health disaster teams. PMID- 11274506 TI - A psychiatric primary care clinic for chronically ill veterans. PMID- 11274507 TI - Managed care in workers' compensation plans. AB - Workers' compensation plans have lagged behind most public and private health care plans in the adoption of managed care techniques. This is largely attributable to the underlying differences between workers' compensation and group health plans. Managed care techniques were developed within group health plans with the objective of health at the lowest cost. In workers' compensation, managed care must address a different objective-restoring a worker to health and productivity at the lowest cost. It is this fundamental difference that makes the application of managed care techniques to workers' compensation plans contentious and at times inappropriate. Research on the impact of managed care on the health and welfare of injured workers is sparse, and important questions remain about the appropriateness of care delivered under workers' compensation managed care plans. In this paper, we discuss the application of managed care to workers' compensation, and highlight the barriers to effective implementation. PMID- 11274509 TI - Graduate medical education: the policy debate. AB - The cost of providing graduate medical education to the approximately 100,000 medical residents in the United States is approximately $18 billion. The government, primarily through the Medicare program, funds almost two thirds of the cost. Unfortunately, the federal government lacks a coherent policy with respect to what objectives it wants to achieve for this expenditure. This article traces (a) the evolution of graduate medical education funding; (b) current proposals to reform the funding mechanism; (c) how the Medicare program currently funds graduate medical education; (d) how funds are allocated to specific institutions; and (e) specific policy objectives that academic medical centers should be held accountable for achieving in return for receiving public funds. PMID- 11274508 TI - U-shaped dose-responses in biology, toxicology, and public health. AB - The occurrence of U-shaped dose-response relationships (often termed hormesis) has been documented in numerous biological, toxicological, and pharmacological investigations. Many of the endpoints studied are of considerable significance to public health (e.g. body weight, cholesterol levels, ethanol consumption, longevity, cancer incidence, etc). Despite the fact that U-shaped dose-responses are widely and independently observed, little attempt has been made to assess this phenomenon in an integrative manner. This review provides an overview of the historical foundations of hormesis and a discussion of its definition within a mechanistic framework. The occurrence, generalizability, and biological significance of U-shaped dose-response relationships along with the concept of biological optimality are addressed. PMID- 11274510 TI - The case for a medicare drug coverage benefit: a critical review of the empirical evidence. AB - The lack of an outpatient prescription drug benefit under Medicare has become a conspicuous omission in the face of accelerated growth in prescription drug expenditures and increased availability of highly effective medications. This article provides a critical review of the empirical evidence on the effect of drug coverage on the use of prescription drugs, health care outcomes, and health care costs among Medicare beneficiaries. The existing literature provides considerable evidence that drug coverage is associated with greater use of all drugs and clinically essential medications and that not all forms of coverage provide the same protection. Longitudinal evidence from elderly and disabled persons in Medicaid indicates that restricting coverage has serious adverse health outcomes for sick and low-income beneficiaries that actually lead to increased health care costs. PMID- 11274511 TI - Hormesis: implications for public policy regarding toxicants. AB - Protecting workers and the public from toxic chemicals, particularly carcinogens, has been a principal goal of public policy. In the absence of knowing by what mechanism of action a toxicant harms people, regulatory toxicology assumes that even tiny doses can cause harm. Risk aversion has led to legislation and regulation that seek to ban toxic chemicals or lower exposure to trivial levels. Contradicting this policy, many studies show health benefits from low-level exposure to toxicants, including some carcinogens. This is known as hormesis. Thus, hormesis could lead to a fundamental change in the policy for regulating toxic substances. In particular, all toxicants that benefit health at low-level exposures should face similar change in regulations for low-dose exposure. The result would be the dissolving of the source of differences in policy for carcinogens and noncarcinogens at low doses. Two questions must be answered before hormesis can be incorporated into regulatory policy. (a) Are there sensitive individuals who would be harmed at doses that would help most people? (b) Is the hormetic effect toxicant specific or would exposure to just a few toxicants achieve the full benefit from hormesis? PMID- 11274512 TI - Consumer reports in health care: do they make a difference? AB - The public release of health care-quality data into more formalized consumer health report cards is intended to educate consumers, improve quality of care, and increase competition in the marketplace The purpose of this review is to evaluate the evidence on the impact of consumer report cards on the behavior of consumers, providers, and purchasers. Studies were selected by conducting database searches in Medline and Healthstar to identify papers published since 1995 in peer-review journals pertaining to consumer report cards on health care. The evidence indicates that consumer report cards do not make a difference in decision making, improvement of quality, or competition. The research to date suggests that perhaps we need to rethink the entire endeavor of consumer report cards. Consumers desire information that is provider specific and may be more likely to use information on rates of errors and adverse outcomes. Purchasers may be in a better position to understand and use information about health plan quality to select high-quality plans to offer consumers and to design premium contributions to steer consumers, through price, to the highest-quality plans. PMID- 11274513 TI - The burden of illness of cancer: economic cost and quality of life. AB - Cancer is a major public health issue and represents a significant burden of disease. In this chapter, we analyze the main measures of burden of disease as relate to cancer. Specifically, we review incidence and mortality, years of life lost from cancer, and cancer prevalence. We also discuss the economic burden of cancer, including cost of illness, phase-specific and long-term costs, and indirect costs. We then examine the impact of cancer on health-related quality of life as measured in global terms (disability-adjusted life years and quality adjusted life years) and using evaluation-oriented applications of health-related quality of life scales. Throughout, we note the relative strengths and weaknesses of the various approaches to measuring the burden of cancer as well as the methodologic challenges that persist in burden-of-illness research. We conclude with a discussion of the research agenda to improve our understanding of the burden of cancer and of illness more generally. PMID- 11274514 TI - Assessing change with longitudinal and clustered binary data. AB - Investigators often gather repeated measures on study subjects to directly measure how a subject's response changes with changes in explanatory variables. This paper focuses on several statistical issues related to assessing change with longitudinal and clustered binary data. Many popular approaches for analyzing repeated binary outcomes measure cross-sectional or between-subject, rather than within-subject, effects of covariates. The class of models known as cluster specific measures within-subject effects of covariates on responses but are subject to additional statistical complications. It is useful to decompose covariates into between- and within-cluster components. This paper describes several approaches that yield consistent estimates of the within-subject covariate effects of interest. Example data from three studies illustrate the results. PMID- 11274515 TI - Design issues for conducting cost-effectiveness analyses alongside clinical trials. AB - In response to rising demands for timely economic data on new medical technologies, cost-effectiveness studies are increasingly being conducted alongside clinical trials. Because of the historical differences in perspective and methods between cost-effectiveness studies and clinical trials, the design phase of these hybrid trials requires special consideration. Cost-effectiveness studies require more comprehensive evaluations of outcomes than the endpoints typically measured in clinical trials. Often, these comprehensive outcome measures (such as quality of life) prove useful for interpreting the other endpoints measured in the trial, as well as for estimating the cost-effectiveness of the intervention. In this manuscript, we discuss several aspects related to the design of joint clinical/economic trials, including study perspective, hypothesis testing, sample size estimation, and methods for collecting cost and outcome data. We also discuss issues that may limit the external validity of the cost-effectiveness results of these trials. Many potential threats to external validity can be successfully addressed if they are identified and accounted for in the design phase of the study. PMID- 11274516 TI - The social ecology of child health and well-being. AB - The term social ecology refers to the nested arrangement of family, school, neighborhood, and community contexts in which children grow up. In this chapter, new directions in public health science as reflected in the theoretical and methodological implications of the concept are explored. The contributions of this ecologically oriented approach to child health practice, designed as it is from a health promotions perspective, are considered. A critique of the term social capital is also presented because of its growing popularity in matters of child health. The point is made that application of this vague term carries the serious risk of misspecifying social phenomena. Future trends in the promotion of child well-being are in a position to flourish given the confluence of advances in theory, methods, and analytical capacity. The capacity to benefit children is also enhanced as public health science aims to translate the principles of child rights into health practice and policy. PMID- 11274517 TI - Selected statistical issues in group randomized trials. AB - Group randomized trials (GRTs) in public health research typically use a small number of randomized groups with a relatively large number of participants per group. Two fundamental features characterize GRTs: a positive correlation of outcomes within a group, and the small number of groups. Appropriate consideration of these fundamental features is essential for design and analysis. This paper presents the fundamental features of GRTs and the importance of considering these features in design and analysis. It also reviews and contrasts the main analytic methods proposed for GRTs, emphasizing the assumptions required to make these methods valid and efficient. Also discussed are various design issues, along with guidelines for choosing among them. A real data example illustrates these issues and methods. PMID- 11274518 TI - Confounding in health research. AB - Consideration of confounding is fundamental to the design, analysis, and interpretation of studies intended to estimate causal effects. Unfortunately, the word confounding has been used synonymously with several other terms, and it has been used to refer to at least four distinct concepts. This paper provides an overview of confounding and related concepts based on a counterfactual model of causation. In this context, which predominates in nonexperimental research, confounding is a source of bias in the estimation of causal effects. Special attention is given to the history of definitions of confounding, the distinction between confounding and confounders, problems in the control of confounding, the relations of confounding to exchangeability and collapsibility, and confounding in randomized trials. PMID- 11274519 TI - Administrative data for public health surveillance and planning. AB - Electronically available administrative data are increasingly used by public health researchers and planners. The validity of the data source has been established, and its strengths and weaknesses relative to data abstracted from medical records and obtained via survey are documented. Administrative data are available from a variety of state, federal, and private sources and can, in many cases, be combined. As a tool for planning and surveillance, administrative data show great promise: They contain consistent elements, are available in a timely manner, and provide information about large numbers of individuals. Because they are available in an electronic format, they are relatively inexpensive to obtain and use. In the United States, however, there is no administrative data set covering the entire population. Although Medicare provides health care for an estimated 96% of the elderly, age 65 years and older, there is no comparable source for those under 65. PMID- 11274520 TI - Small-community-based surveys. AB - Rapid, small surveys are routinely done in much of the developing world but are less common in the United States. We present as an example a rapid survey of immunization status and other factors in a predominantly Hispanic region in Los Angeles. The survey united county employees, students, and community volunteers, first to enumerate the eligible population and then to conduct in-person interviews. Sampling was done in two stages in a downtown region of Los Angeles. Over the course of two weekends and during clean-up the following week, volunteers and others enumerated 718 eligible children in 30 clusters (i.e. groups of blocks). At the second stage, also in two weekends with midweek clean up, we selected by simple random sample 10 children per cluster. The parents or legal guardians of 270 children were interviewed about vaccination issues, including home presence of an immunization card. Nearly one fourth of the respondents did not have a home telephone number and thus would have been underrepresented in a telephone survey. Information from such rapid surveys is important for local program planning and evaluation. PMID- 11274521 TI - Innovations in treatment for drug abuse: solutions to a public health problem. AB - Illicit drug use is an important public health problem with broad social costs. The low effectiveness of prevention efforts leaves treatment of drug dependence as one of the most powerful means of fighting illicit drug use. Treatment reduces drug use and crime and increases individuals' functioning. However, programs that treat drug dependence have high dropout rates and low completion rates. In addition, some individuals continue to use drugs while in treatment, and relapse is common. Furthermore, only a fraction of those who need treatment receive it. Recently, there have been important innovations that reduce barriers and increase effectiveness of treatment. These innovations include new pharmacological agents, novel counseling strategies, promising ways to motivate, and treatment in new settings. This paper describes standard treatments and recent innovations designed to increase (a) effectiveness of treatment, (b) motivation to seek care, (c) access, (d) retention, and (e) cost-effectiveness. We provide criteria on how these innovations should be evaluated in order to determine which should be adopted, funded, and transferred to existing and future treatment programs. PMID- 11274522 TI - Managed care: a view from Europe. AB - This article summarizes recent developments in the United Kingdom, the Netherlands, Switzerland, and the new Baltic states that reflect the influence of US managed care concepts and practices. We emphasize (a) developments in restructuring traditional health insurance mechanisms by shifting premium and out of-pocket burdens to consumers so as to constrain demand and costs and (b) reliance on prospective hospital budgets and case management by primary physicians. Social insurance mechanisms and universal coverage remain national tasks as well as basic components of the social structure of most European countries. Full open-market competition between traditional sick funds and private insurance companies and the introduction of for-profit MCOs beholden to their shareholders appears unlikely on other than an experimental basis. Increased competition between providers may well result from the new right of insurers and payers to contract with medical care providers of their choice. It remains to be shown how far these experiments, which differ substantially between the countries examined, will succeed in their objectives and become permanent features of their national systems. PMID- 11274523 TI - Minisymposium on obesity: overview and some strategic considerations. AB - The high and still increasing prevalence of obesity in US children, adolescents, and adults poses a major economic and health threat to our society. The three reviews in this minisymposium on obesity explore the health issues by: 1) describing the public health impact of obesity; 2) examining the multiple and complex environmental influences on eating and physical activity patterns; and 3) considering how the development of obesity during childhood and adolescence can be prevented through interventions in school, family, and primary care settings. This overview explains the importance, for effective long-term obesity prevention and control, of intersectoral policy and environmental initiatives-in addition to behavior change approaches aimed at individuals. The need for public health professionals to influence and operate within a variety of non-health sectors such as transportation, education, urban planning, and commerce may be seen as the greatest barrier but may also be the greatest opportunity. PMID- 11274524 TI - Environmental influences on eating and physical activity. AB - Obesity has increased dramatically over the past two decades and currently about 50% of US adults and 25% of US children are overweight. The current epidemic of obesity is caused largely by an environment that promotes excessive food intake and discourages physical activity. This chapter reviews what is known about environmental influences on physical activity and eating behaviors. Recent trends in food supply, eating out, physical activity, and inactivity are reviewed, as are the effects of advertising, promotion, and pricing on eating and physical activity. Public health interventions, opportunities, and potential strategies to combat the obesity epidemic by promoting an environment that supports healthy eating and physical activity are discussed. PMID- 11274525 TI - Preventing obesity in children and adolescents. AB - In this review, we address the natural history of obesity in children, the most promising family- and school-based approaches to the prevention of obesity, and the barriers and opportunities associated with secondary prevention. In childhood, the most important periods of risk appear to be the periods of adiposity rebound and adolescence. Caution regarding the period of adiposity rebound is still warranted, because it is not yet clear that early rebound is attributable to changes in body fat. Families and schools represent the most important foci for preventive efforts in children and adolescents. One productive approach is to proceed from an examination of factors that affect energy balance to the identification of more proximal influences on those factors. This approach may help to narrow the strategies necessary to prevent or treat childhood obesity. For example, television viewing affects both energy intake and energy expenditure, and therefore represents a logical target for interventions. Anticipatory guidance by pediatricians may offer an effective mechanism by which to change parental attitudes and practices regarding television viewing. A similar process is used to emphasize the potential influence of school-based interventions directed at changes in food choices and sedentary behavior. PMID- 11274526 TI - The public health impact of obesity. AB - The increase in obesity worldwide will have an important impact on the global incidence of cardiovascular disease, type 2 diabetes mellitus, cancer, osteoarthritis, work disability, and sleep apnea. Obesity has a more pronounced impact on morbidity than on mortality. Disability due to obesity-related cardiovascular diseases will increase particularly in industrialized countries, as patients survive cardiovascular diseases in these countries more often than in nonindustrialized countries. Disability due to obesity-related type 2 diabetes will increase particularly in industrializing countries, as insulin supply is usually insufficient in these countries. As a result, in these countries, an increase in disabling nephropathy, arteriosclerosis, neuropathy, and retinopathy is expected. Increases in the prevalence of obesity will potentially lead to an increase in the number of years that subjects suffer from obesity-related morbidity and disability. A 1% increase in the prevalence of obesity in such countries as India and China leads to 20 million additional cases of obesity. Prevention programs will stem the obesity epidemic more efficiently than weight loss programs. However, only a few prevention programs have been developed or implemented, and the success rates reported to date have been low. Obesity prevention programs should be high on the scientific and political agenda in both industrialized and industrializing countries. PMID- 11274527 TI - Radiation oncology: contributions of the United States in the last years of the 20th century. AB - The advancements in radiation oncology in the past 50 years in the United States were probably more dramatic than those in the first half of the 20th century. Not only were there major technical achievements, but there was also an associated increase in the overall cure rates of cancer, from 20% at 5 years 50 years ago to now nearly 60% at 5 years. The cure rates in selected tumor sites at 5 years in 1950 and in 2000, respectively, were as follows: breast, 50% and 80%; colon and rectum, 40% and 85%; lung, 5% and 15%-20%; prostate, 40% and 80%; Hodgkin disease, 50% and more than 90%; cervix, 40% and 70%-80%; uterus (endometrium), 80% and more than 90%; bladder, 30% and 50%; head and neck, 30% and 60%; and esophagus, 2% and 15%. Much of this has been due to a broader array of techniques in radiation therapy available for treatment but also because of new emphasis on combined integrated modalitities (surgery, radiation therapy, and chemotherapy). New imaging techniques have contributed substantially, allowing better selection of patients for treatment and better selections of treatment modalities. PMID- 11274528 TI - The medical profession: a satisfying career. PMID- 11274529 TI - Apparent diffusion coefficient mapping in patients with Alzheimer disease or mild cognitive impairment and in normally aging control subjects: present and future. PMID- 11274530 TI - Wrist fractures: what the clinician wants to know. AB - With the recent improvements in diagnosis and treatment of distal radius and carpal injuries, the hand surgeons' expectations of relevant radiologic interpretation of imaging studies are heightened. Conventional radiographic examinations, as well as more sophisticated and invasive studies, have important roles in the evaluation of wrist fractures and dislocations. On the basis of physical examination results and the mechanism of injury, the onus is on the examining surgeon to pinpoint potential sites of bone or ligament disruption. After this evaluation, appropriate imaging studies appropriately performed and interpreted will help direct treatment and improve outcome with greater clarity and certainty. PMID- 11274531 TI - Role of us in the preoperative evaluation of patients with anterior shoulder instability. AB - PURPOSE: To assess the value of ultrasonography (US) in the preoperative evaluation of patients with anterior shoulder instability. MATERIALS AND METHODS: Twenty-two patients with one-sided anterior shoulder instability were examined with US by using three dynamic scanning approaches: two frontal and one axillary. The anterior labrum, the anterior ligamental-capsular complex, and the presence of humeral head and glenoid rim fractures were evaluated. Arthroscopy or arthrotomy was subsequently performed in all patients and was considered the standard. RESULTS: US correctly depicted the presence (n = 20) or absence (n = 1) of humeral head fractures and the presence (n = 10) or absence (n = 9) of glenoid rim fractures. All 22 patients had anterior labral tears; 21 tears were correctly depicted with US. The labral tear was seen as a hypoechoic zone larger than 2 mm (n = 15), labral movement (n = 10), a degenerated labrum (n = 6), or a vacuum phenomenon (n = 3). The anterior ligamental-capsular complex was correctly evaluated in 14 patients. The use of multiple approaches helped to prevent misinterpretation, but there were no substantial differences among the approaches in the depiction of the anterior shoulder structures. CONCLUSION: The high accuracy in the depiction of labral tears and associated fractures indicates that US can provide useful preoperative information in patients with anterior shoulder instability. PMID- 11274532 TI - Shell osteochondral allografts of the knee: comparison of mr imaging findings and immunologic responses. AB - PURPOSE: To define the magnetic resonance (MR) imaging appearance of shell osteochondral allografts of the knee and compare the MR findings with antibody responses. MATERIALS AND METHODS: Thirty-six grafts were evaluated with a 1.5-T unit with T1-, intermediate-, and T2-weighted, and three-dimensional spoiled gradient-recalled MR imaging at 3, 6, 12, 24, and/or 36 months after surgery. Nineteen patients underwent imaging serially. Two osteoradiologists scored by consensus host marrow edema, thickness of graft-host interface, signal intensity of graft marrow, cyst formation, joint effusion, articular cartilage defects, and surface collapse. Patients were divided into antibody-positive (AP) (n = 11) and antibody-negative (AN) (n = 25) groups evenly distributed across the different time points on the basis of results of anti-human leukocyte antigen antibody screening. MR findings for the two groups were compared. RESULTS: AP patients demonstrated greater mean edema (P<.002), thicker interface (P<.03), and more abnormal graft marrow (P<.04) than AN patients, and they had a higher proportion of surface collapse (P<.03). CONCLUSION: Humoral immune responses were associated with more inflammation and less complete incorporation after allograft placement. MR imaging shows promise as a surrogate biomarker for success of shell osteochondral allograft implantation. PMID- 11274533 TI - Cost-effectiveness of colorectal cancer screening. AB - PURPOSE: To determine the most cost-effective colorectal cancer screening strategy costing less than $100,000 per life-year saved and to determine how available strategies compare with each other. MATERIALS AND METHODS: Standardized methods were used to calculate incremental cost-effectiveness ratios (ICERs) from published estimates of cost and effectiveness of colorectal cancer screening strategies, and the direction and magnitude of any effect on the ratio from parameter estimate adjustments based on literature values were estimated. RESULTS: Strategies in which double-contrast barium enema examination was performed emerged as optimal from all studies included. In average-risk individuals, screening with double-contrast barium enema examination every 3 years, or every 5 years with annual fecal occult blood testing, had an ICER of less than $55,600 per life-year saved. However, double-contrast barium enema examination screening every 3 years plus annual fecal occult blood testing had an ICER of more than $100,000 per life-year saved. Colonoscopic screening had an ICER of more than $100,000 per life-year saved, was dominated by other screening strategies, and offered less benefit than did double-contrast barium enema examination screening. CONCLUSION: Double-contrast barium enema examination can be a cost-effective component of colorectal cancer screening, but further modeling efforts are necessary. PMID- 11274534 TI - Automated polyp detection at CT colonography: feasibility assessment in a human population. AB - PURPOSE: To test the feasibility of and improve a computer algorithm to automatically detect colonic polyps in real human computed tomographic (CT) colonographic data sets. MATERIALS AND METHODS: Twenty patients with known polyps underwent CT colonography in the supine position. CT colonographic data were processed by using a shape-based algorithm that depicts masses that protrude into the lumen. We studied nine shape criteria and three isosurface threshold settings. Results were compared with those of conventional colonoscopy performed the same day. RESULTS: There were 50 polyps (28 were > or =10 mm in size; 12, 5-9 mm; 10, <5 mm). The sensitivity with optimal settings for detecting polyps 10 mm or greater was 64% (18 of 28). Sensitivity improved to 71% (10 of 14) for polyps 10 mm or greater in well-distended colonic segments. Performance decreased for polyps less than 10 mm, poorly distended colonic segments, and other shape algorithms. There was a mean of six false-positive lesion sites per colon. These sites were reduced 39% to 3.5 per colon by sampling CT attenuation at the lesion site and discarding sites having attenuation less than a threshold. CONCLUSION: Automated detection of colonic polyps, especially clinically important large polyps, is feasible. Colonic distention is an important determinant of sensitivity. Further increases in sensitivity may be achieved by adding prone CT colonography. PMID- 11274535 TI - Focal nodular hyperplasia: CT findings with emphasis on multiphasic helical CT in 78 patients. AB - PURPOSE: To evaluate features of focal nodular hyperplasia (FNH) at multiphasic helical computed tomography (CT). MATERIALS AND METHODS: Clinical, pathologic, and preoperative imaging findings were retrospectively reviewed in 78 patients. Conventional liver CT was performed in nine patients; helical multiphasic CT, in 69. Diagnosis was based on complete resection (n = 20), biopsy (n = 42), or clinical and imaging follow-up for a minimum of 6 months (n = 16). Number, size, location, margins, surface, homogeneity of enhancement, and presence of a central scar, mass effect, exophytic growth, calcification, pseudocapsule, or vessels feeding or draining the lesion were evaluated. RESULTS: CT depicted 124 tumors (mean diameter, 4.1 cm; range, 1-11 cm); 62 were small (< or =3 cm). FNHs were hypervascular and hyperattenuating to liver on 106 of 106 arterial phase scans and were isoattenuating to liver on 82 of 89 delayed scans. Of the 124 tumors, 111 enhanced homogeneously, 109 had a smooth surface, 101 were subcapsular, 89 had ill-defined margins, and 62 had a central scar that was observed more often in large lesions (40 of 62 lesions) than in small lesions (22 of 62 lesions). FNHs less frequently exerted a mass effect (43 lesions), had vessels around or within the lesion (42 lesions), demonstrated exophytic growth (40 lesions), or showed a pseudocapsule (10 lesions). Only one FNH had calcification. CONCLUSION: Helical CT demonstrates characteristic features that may allow confident diagnosis of FNH. In typical cases, neither biopsy nor further imaging is necessary. PMID- 11274536 TI - Hemangioma in the cirrhotic liver: diagnosis and natural history. AB - PURPOSE: To investigate the natural history and diagnosis of cavernous hemangioma in the cirrhotic liver with computed tomography (CT) and magnetic resonance (MR) imaging. MATERIALS AND METHODS: Imaging and pathologic findings of 21 hemangiomas in 17 patients were retrospectively reviewed. CT of the liver was performed in all patients; MR imaging, in four. Cirrhosis was confirmed histologically in all patients, and the diagnosis of hemangioma was based on histopathologic findings (15 patients, 18 hemangiomas) or strict imaging criteria (two patients, three hemangiomas). Ten patients underwent imaging follow-up. The number, sizes, location, attenuation, pattern of enhancement, exophytic growth, presence of capsular retraction, and size stability were evaluated. RESULTS: Of the 21 hemangiomas, five were not detected at CT or MR imaging. Twelve (75%) of 16 hemangiomas were subcapsular, two (12%) of 16 demonstrated exophytic growth, 14 (87%) of 16 demonstrated nodular peripheral enhancement, and 16 (100%) of 16 were isoattenuating to blood vessels. At MR imaging, all five hemangiomas demonstrated nodular peripheral enhancement and hyperintensity on T2-weighted images. Seven lesions were smaller at follow-up, and five lesions developed retraction of the hepatic capsule. CONCLUSION: Even within the cirrhotic liver, larger hemangiomas can usually be diagnosed confidently with CT or MR imaging. With progressive cirrhosis, however, hemangiomas are likely to decrease in size, become more fibrotic, and are difficult to diagnose radiologically and pathologically. PMID- 11274537 TI - Breath-hold three-dimensional CT of the liver with multi-detector row helical CT. AB - PURPOSE: To compare image quality on transverse source images and coronal and sagittal reformations to determine the feasibility of using single-breath-hold three-dimensional liver computed tomography (CT) with multi-detector row helical CT in patients suspected of having hepatic metastases. MATERIALS AND METHODS: Fifty-three patients underwent the protocol. Coronal and sagittal reformations were constructed. Images were reviewed for duration of scan acquisition and length and adequacy of z-axis coverage. Reformations were scored for visualization of portal and hepatic vein branches, liver edge sharpness, cardiac pulsation and respiratory motion artifacts, noise due to mottle, and overall impression. RESULTS: Mean z-axis coverage was 207 mm +/- 33 (SD) (range, 145-280 mm), with a mean acquisition time of 10.96 seconds +/- 1.78 (range, 7.73-14.93 seconds). In 44 (83%) patients, the entire liver was imaged on a single helical scan. Artifact from cardiac motion was not identified on the transverse source images in any patient but was identified on coronal images in eight (15%) and on sagittal images in seven (13%). Similarly, noise due to mottle was not identified on the transverse source images but was identified on coronal images in seven (13%) patients and on sagittal images in six (11%). CONCLUSION: It is feasible to perform single-breath-hold three-dimensional liver CT with multi-detector row helical CT technology. Reformations provide a unique perspective with which to view the liver and may improve diagnostic capacity. PMID- 11274539 TI - Case 33: sinus of valsalva aneurysm. PMID- 11274540 TI - Protein-losing enteropathy: diagnosis with (99m)Tc-labeled human serum albumin scintigraphy. AB - PURPOSE: To investigate the diagnostic value of technetium 99m-labeled human serum albumin (HSA) scintigraphy in a group of patients suspected of having protein-losing enteropathy (PLE). MATERIALS AND METHODS: After intravenous injection of 740 MBq of freshly prepared (99m)Tc HSA, serial images of the abdomen were obtained from 10 minutes until 24 hours after injection. A (99m)Tc HSA scan was considered positive for PLE if there was visible tracer exudation in the gut. The diagnosis was established on the basis of standard clinical and biopsy findings. RESULTS: Thirty-nine scans were obtained: 27 scans in 26 suspected cases of PLE and 12 scans in control subjects with no known gastrointestinal disease. Twenty-five of the 26 studies in patients suspected of having PLE showed (99m)Tc HSA activity in the bowel. Among the 25 studies with positive findings, seven demonstrated PLE only on images obtained 24 hours after injection. In the control subjects, no activity was seen in the bowel. CONCLUSION: (99m)Tc HSA with serial scanning for up to 24 hours is reliable and useful for imaging PLE. Sites of protein loss may also be demonstrated. This imaging method is convenient, easy to perform, and yields results within 24 hours. PMID- 11274541 TI - Frequency of right lower quadrant position of the sigmoid colon in infants and young children. AB - PURPOSE: To evaluate the frequency of right lower quadrant positioning of the sigmoid colon in infants and young children. MATERIALS AND METHODS: Findings in 169 patients who underwent enema examination were retrospectively reviewed. Sigmoid colon position was categorized as in the left or right lower quadrant or midline. Patients who had an anatomic abnormality that affected colonic position (eg, malrotation or abdominal mass) or had previously undergone abdominal surgery were excluded. The frequency of right lower quadrant sigmoid position was evaluated for a relationship with patient age (analysis of variance) and sex (chi(2) test). RESULTS: Patient ages were 1 day to 5 years (mean age, 13 months). The sigmoid colon was in the right lower quadrant in 74 (44%), in the left lower quadrant in 73 (43%), and in the midline in 18 (11%). The position was variable in one patient and indeterminate in three. When the sigmoid colon was within the right lower quadrant, it often extended laterally, overlying the position of the cecum and ascending colon. There were no significant correlations between right lower quadrant position and patient age (P =.262) and sex (P =.162). CONCLUSION: In children, the sigmoid colon is often within the right lower quadrant. Knowledge of this high frequency should reduce the likelihood of misinterpreting air within a redundant right-sided sigmoid colon as air within the cecum in children suspected of having abnormalities such as intussusception. PMID- 11274542 TI - Early prediction of irreversible brain damage after ischemic stroke at CT. AB - PURPOSE: To assess the capability of computed tomography (CT) in the prediction of irreversible ischemic brain damage and its association with the clinical course within 6 hours of stroke onset. MATERIALS AND METHODS: Serial CT scans obtained within 6 hours of stroke onset, at 22-96 hours (median, 1 day), and at 2 36 days (median, 7 days) after symptom onset in 786 patients with ischemic stroke were prospectively studied, and follow-up CT scans were used as the reference. Clinical variables were assessed prospectively and independently of CT evaluation. RESULTS: The specificity and positive predictive value of ischemic edema at baseline CT for brain infarcts were 85% (95% CI: 77%, 91%) and 96% (95% CI: 94%, 98%), respectively. Sensitivity and negative predictive values were 64% (95% CI: 60%, 67%) and 27% (95% CI: 23%, 32%), respectively. Patients without early CT findings were less severely affected (P<.001), developed smaller infarcts (P<.001), had fewer intracranial bleeding events (P<.001), and had a better clinical outcome at 90 days (P<.001) compared with patients with hypoattenuating brain tissue at early CT. CONCLUSION: After ischemic stroke, x ray hypoattenuation at CT is highly specific for irreversible ischemic brain damage if detection occurs within the first 6 hours. Patients without hypoattenuating brain tissue have a more favorable clinical course. PMID- 11274543 TI - Mild cognitive impairment and Alzheimer disease: regional diffusivity of water. AB - PURPOSE: To compare the regional diffusivity of water in the brains of normally aging elderly people and patients with mild cognitive impairment (MCI) or Alzheimer disease. MATERIALS AND METHODS: Magnetic resonance images were obtained in 21 patients with Alzheimer disease, 19 patients with MCI, and 55 normally aging elderly control subjects without evidence of cognitive impairment. Regions of interest were drawn to compare the apparent diffusion coefficient (ADC) and the anisotropy index (AI) in frontal, parietal, temporal, occipital, anterior, and posterior cingulate white matter (WM), and the thalami and hippocampi. RESULTS: Hippocampal ADC was higher in MCI and Alzheimer disease patients than in control subjects. ADC of the temporal stem and posterior cingulate, occipital, and parietal WM was higher in Alzheimer disease patients than in control subjects. Except for occipital AI, which was lower in MCI patients than in control subjects, there were no differences in AI among the three groups for any of the regions. CONCLUSION: Hippocampal ADC was significantly different between control subjects and MCI patients, many of whom likely have preclinical Alzheimer disease. Elevation in hippocampal ADC may reflect early ultrastructural changes in the progression of Alzheimer disease. PMID- 11274544 TI - Follow-up of intracranial aneurysms treated with detachable coils: comparison of gadolinium-enhanced 3D time-of-flight MR angiography and digital subtraction angiography. AB - PURPOSE: To compare three-dimensional (3D) time-of-flight magnetic resonance (MR) angiography with digital subtraction angiography (DSA) in the follow-up of intracranial aneurysms treated with selective endovascular placement of detachable coils. MATERIALS AND METHODS: Sixty-eight consecutive patients with intracranial aneurysms were included in the prospective study. The goal was to evaluate 3D time-of-flight MR angiography versus DSA for the detection of a residual aneurysm neck or residual flow inside the coil mesh. RESULTS: Eighty-one MR angiographic and 83 DSA examinations were performed; 15 patients were examined with both modalities twice. MR angiography was not possible in two patients. In another patient, the quality of MR angiography was not sufficient to assess the treated aneurysm. In 72 of the remaining 80 MR angiographic and DSA examinations, there was good correlation between the two modalities. In 54 cases, neither image type showed remnants or recurrence, but in 18, both showed residual aneurysm. In eight cases, the MR angiographic and DSA results differed. In one of these cases, MR angiography depicted residual aneurysm but DSA depicted an arterial loop. In seven cases, a small (<3-mm) remnant was not detected at MR angiography. CONCLUSION: Because very small aneurysm remnants or recurrences probably are not clinically important, MR angiography is an option for following up intracranial aneurysms treated with detachable coils and may partly replace DSA. PMID- 11274545 TI - Carotid MR angiography: phase II study of safety and efficacy for MS-325. AB - PURPOSE: To evaluate the safety and efficacy of MS-325 in patients suspected of having carotid arterial disease. MATERIALS AND METHODS: Fifty carotid arteries in 26 patients were imaged with three-dimensional spoiled gradient-recalled-echo magnetic resonance (MR) angiography at 5 and 50 minutes after injection of MS 325. MS-325 was administered intravenously as a single dose of 0.01, 0.03, or 0.05 mmol per kilogram of body weight as determined with a dose randomization scheme for four, nine, and 13 patients, respectively. Safety, including clinical laboratory changes and electrocardiographic monitoring, was assessed until approximately 3 days after injection. Conventional contrast agent-enhanced angiography was used as the standard of reference. Independent readers blinded to the dose interpreted the MR angiographic and conventional images. Images were assessed for location and extent of carotid arterial stenosis. RESULTS: There were no severe or serious adverse events. For the determination of clinically significant stenosis (>70%) on the 5-minute images, sensitivity, specificity, and accuracy (P =.07, three-way comparison) were 100%, 100%, and 100%; 63%, 100%, and 88%; and 40%, 75%, and 55% at 0.01, 0.03, and 0.05 mmol/kg, respectively. Sensitivity and specificity for images at 50 minutes after MS-325 administration showed the same trends as the 5-minute images. CONCLUSION: Overall accuracy for MS-325-enhanced carotid MR angiography performed during steady-state conditions of circulating contrast agent approximately 5 minutes after injection was high (88%-100%) at 0.03 and 0.01 mmol/kg. MS-325 was well tolerated at all evaluated doses. PMID- 11274546 TI - Treatment of iatrogenic femoral pseudoaneurysms with percutaneous thrombin injection: experience in 54 patients. AB - PURPOSE: To assess the clinical success of ultrasonography (US)-guided thrombin injection for the treatment of iatrogenic femoral pseudoaneurysms and to identify criteria that may predispose to treatment failure. MATERIALS AND METHODS: Fifty four iatrogenic femoral pseudoaneurysms were treated with US-guided thrombin injection. Forty-five were classified as simple (single lobe) and nine, as complex (at least two lobes and a single neck to the native vessel). Pseudoaneurysm volume, classification, thrombin dose, anticoagulation therapy status, and sheath size were compared between failed and successful cases. Seven- to 10-day follow-up US and a minimum 4-month clinical follow-up were also performed to evaluate success. RESULTS: Fifty of 54 pseudoaneurysms were successfully treated with topical thrombin without complication and included all 45 simple and five of nine complex pseudoaneurysms. US follow-up in all 50 successful cases and clinical follow-up in 37 of these revealed no recurrence. Only a complex pseudoaneurysm classification was significantly associated with failure (P<.01). Among the complex pseudoaneurysms, successful cases involved two injections and a total thrombin dose of at least 1,500 units. In failed cases, pseudoaneurysms were treated with a single injection of 1,000 units, initially thrombosed, and recurred. CONCLUSION: Simple iatrogenic femoral pseudoaneurysms, regardless of size or concomitant anticoagulation therapy, can be treated with a single injection of up to 1,000 units of topical thrombin and require no follow up. Complex pseudoaneurysms will likely require a second injection (total thrombin dose of at least 1,500 units) and short-term clinical and US follow-up to ensure successful treatment. PMID- 11274547 TI - Iliac arterial injuries after endovascular repair of abdominal aortic aneurysms: correlation with iliac curvature and diameter. AB - PURPOSE: To determine the relationship between iliac arterial tortuosity and cross-sectional area and the occurrence of iliac arterial injuries following transfemoral delivery of endovascular prostheses for repair of abdominal aortic aneurysms. MATERIALS AND METHODS: Iliac arterial curvature values and orthogonal cross-sectional areas were determined from helical computed tomographic (CT) data acquired in 42 patients prior to transfemoral delivery of aortic stent-grafts. The curvature and luminal cross-sectional area orthogonal to the median centerline were quantified every millimeter along the median centerline of the iliac arteries. An indicator of global iliac tortuosity, the iliac tortuosity index, was defined as the sum of the curvature values for all points with a curvature of 0.3 cm(-1) or greater, and cross-sectional area (CSA) was indexed for all points as the mean cross-sectional diameter (D = 2 radical[CSA/pi]). Following stent-graft deployment, helical CT data were analyzed for the presence of iliac arterial dissections independently by two reviewers. RESULTS: Eighteen dissections were detected in 16 patients. The iliac tortuosity index was significantly larger in iliac arteries with dissections (35.5 +/- 20.8 [mean +/- SD]) when compared with both nondissected contralateral iliac arteries in the same patients (26.1 +/- 21.0, P =.001) and iliac arteries in patients without any iliac arterial injury (20 +/- 9, P =.009). The tortuosity index was higher ipsilateral to the primary component delivery in 10 of 11 iliac dissections that developed along the primary component delivery route. CONCLUSION: A high degree of iliac arterial tortuosity appears to impart greater risk for the development of iliac arterial injuries in patients undergoing transfemoral delivery of endovascular devices. PMID- 11274548 TI - Safety and effectiveness of single- versus triple-dose gadodiamide injection- enhanced MR angiography of the abdomen: a phase III double-blind multicenter study. AB - PURPOSE: To evaluate the safety and effectiveness of gadodiamide-enhanced magnetic resonance (MR) angiography with single and triple doses in the assessment of abdominal arterial stenoses. MATERIALS AND METHODS: One hundred five patients were included in the randomized, double-blind, phase III multicenter trial. Results of MR angiography with 0.1 mmol/kg and 0.3 mmol/kg doses of gadodiamide were compared with those of digital subtraction angiography (DSA) and according to dose. RESULTS: No serious adverse events were observed. The mean contrast index at the region proximal to the primary stenosis was significantly higher in the triple-dose group (P =.03). Mean 95% CI values for the difference in depicted degree of stenosis between DSA and postcontrast MR angiography improved from -3.4% +/- 4.7 (SD) in the single-dose group to -1.2% +/ 4.7 in the triple-dose group. Mean values for overall image quality on the visual analogue scale improved with the triple dose (P =.02). Confidence in diagnosis was high at postcontrast MR angiography in 88% and 96% of cases in the single- and triple-dose groups, respectively. CONCLUSION: Gadodiamide-enhanced MR angiography performed with single and triple doses is safe and effective for assessing major abdominal arterial stenoses. Although high agreement between MR angiography and DSA was achieved with both doses, triple-dose MR angiography was superior in the evaluations of image quality, degree of arterial stenoses, and confidence in diagnosis. PMID- 11274549 TI - Hepatocellular carcinoma: involvement of the internal mammary artery. AB - PURPOSE: To investigate factors related to the development of internal mammary arteries (IMAs) as feeding arteries of hepatocellular carcinomas (HCCs). MATERIALS AND METHODS: In 30 patients with HCC located in ventral hepatic areas directly beneath the diaphragm, bilateral internal mammary arteriography was performed to explore involvement of the IMA with HCC. The number of previous transcatheter arterial embolizations (TAEs), tumor size, time from initial TAE to IMA angiography, inferior phrenic artery (IPA) involvement with tumor, presence of hepatic artery occlusion, and use of other treatments were compared in groups with and without involvement of the IMA. RESULTS: The group with IMA involvement included 10 patients; the group without involvement, 20 patients. TAE had been performed two to 12 times in the group with involvement and zero to six times in the group without involvement (P =.01). Mean tumor sizes in these two groups were 5.1 and 6.0 cm, respectively; hepatic artery occlusion was noted in nine and zero patients (P =.01) in the two groups. The time from initial TAE to IMA angiography ranged from 3 to 53 months (median, 31.5 months) and from zero to 89 months (median, 0 months) (P =.01). IPA involvement was observed in seven and four patients (P =.015). CONCLUSION: These results strongly suggest that, regardless of tumor size, when HCCs are located in the ventral hepatic areas directly beneath the diaphragm, the IMAs serve as feeding arteries in patients with hepatic artery occlusion caused by repeated TAE. PMID- 11274550 TI - Comparison of three mechanical thrombus removal devices in thrombosed canine iliac arteries. AB - PURPOSE: To assess and compare intimal and medial vascular damage caused by three mechanical wall-contact thrombectomy devices: Fogarty embolectomy catheter, Arrow Trerotola peripheral thrombectomy device, and MTI-Castaneda over-the-wire brush. MATERIALS AND METHODS: Bilateral external iliac arteries of 15 canines were thrombosed before mechanical thrombolysis. Ten thrombosed arteries were randomly assigned to receive each device. Animals were sacrificed immediately, and histologic assessment of endothelial and medial damage in the vessels was performed. RESULTS: The vascular damage found with all devices extended into the tunica media. The Fogarty embolectomy catheter and the Arrow-Trerotola device caused significantly more damage than the Castaneda brush. CONCLUSION: All devices caused lesions extending into the media. Previous research has shown that the extent and depth of the vascular lesion may be contributing factors in promoting early atherosclerotic and accelerated hyperplastic intimal and medial changes. These findings warrant further study of these devices in an atherosclerotic model with longer follow-up. PMID- 11274551 TI - Radio-frequency thermal ablation with NaCl solution injection: effect of electrical conductivity on tissue heating and coagulation-phantom and porcine liver study. AB - PURPOSE: To characterize the effects of NaCl concentration on tissue electrical conductivity, radio-frequency (RF) deposition, and heating in phantoms and optimize adjunctive NaCl solution injection for RF ablation in an in vivo model. MATERIALS AND METHODS: RF was applied for 12-15 minutes with internally cooled electrodes. For phantom experiments (n = 51), the NaCl concentration in standardized 5% agar was varied (0%-25.0%). A nonlinear simplex optimization strategy was then used in normal porcine liver (n = 44) to determine optimal pre RF NaCl solution injection parameters (concentration, 0%-38.5%; volume, 0-25 mL). NaCl concentration and tissue conductivity were correlated with RF energy deposition, tissue heating, and induced coagulation. RESULTS: NaCl concentration had significant but nonlinear effects on electrical conductivity, RF deposition, and heating of agar phantoms (P<.01). Progressively greater heating was observed to 5.0% NaCl, with reduced temperatures at higher concentrations. For in vivo liver, NaCl solution volume and concentration significantly influenced both tissue heating and coagulation (P<.001). Maximum heating 20 mm from the electrode (102.9 degrees C +/- 4.3 [SD]) and coagulation (7.1 cm +/- 1.1) occurred with injection of 6 mL of 38.5% (saturated) NaCl solution. CONCLUSION: Injection of NaCl solution before RF ablation can increase energy deposition, tissue heating, and induced coagulation, which will likely benefit clinical RF ablation. In normal well-perfused liver, maximum coagulation (7.0 cm) occurs with injection of small volumes of saturated NaCl solution. PMID- 11274552 TI - Quantified power Doppler US of tumor blood flow correlates with microscopic quantification of tumor blood vessels. AB - PURPOSE: To evaluate the ability of a quantified power Doppler ultrasonography (US) system to help quantitate differences in tumor vascularity after radiation therapy and administration of tumor necrosis factor (TNF). MATERIALS AND METHODS: Murine glioblastoma tumors were grown in the thighs of two sets of 25 mice each. Each mouse was assigned to one of four treatment groups: control (no treatment), radiation therapy, TNF therapy, or combination therapy (both radiation and TNF therapies). Mice were then evaluated with quantified power Doppler US, and a vascularity index (color area) was calculated for different tumor regions in each group. The tumors were then excised, and histologic evaluation was performed by using an immunofluorescence-tagged monoclonal antibody against blood vessel endothelium. The number of stained blood vessels per high-power field was correlated with the sonographically determined vascularity index. RESULTS: The color area of the total tumor decreased to 37% of that in the control group in mice treated with radiation therapy alone (P: =.02), 26% of that in the control group in mice treated with TNF alone (P: =.05), and 8% of that in the control group in those treated with both TNF and radiation (P: =.006). These results correlated well with the quantified results from immunofluorescent staining (r = 0.98). CONCLUSION: Quantified power Doppler US is a noninvasive method for the evaluation of tumor vascularity and blood flow. PMID- 11274553 TI - Digital optical imaging of green fluorescent proteins for tracking vascular gene expression: feasibility study in rabbit and human cell models. AB - PURPOSE: To investigate the feasibility of using a sensitive digital optical imaging technique to detect green fluorescent protein (GFP) expressed in rabbit vasculature and human arterial smooth muscle cells. MATERIALS AND METHODS: A GFP plasmid was transfected into human arterial smooth muscle cells to obtain a GFP smooth muscle cell solution. This solution was imaged in cell phantoms by using a prototype digital optical imaging system. For in vivo validation, a GFP lentivirus vector was transfected during surgery into the carotid arteries of two rabbits, and GFP-targeted vessels were harvested for digital optical imaging ex vivo. RESULTS: Optical imaging of cell phantoms resulted in a spatial resolution of 25 microm/pixel. Fluorescent signals were detected as diffusely distributed bright spots. At ex vivo optical imaging of arterial tissues, the average fluorescent signal was significantly higher (P <.05) in GFP-targeted tissues (mean +/- SD, 9,357.3 absolute units of density +/- 1,001.3) than in control tissues (5,633.7 absolute units of density +/- 985.2). Both fluorescence microscopic and immunohistochemical findings confirmed these differences between GFP-targeted and control vessels. CONCLUSION: The digital optical imaging system was sensitive to GFPs and may potentially provide an in vivo imaging tool to monitor and track vascular gene transfer and expression in experimental investigations. PMID- 11274555 TI - Breast biopsy avoidance: the value of normal mammograms and normal sonograms in the setting of a palpable lump. AB - PURPOSE: To review the authors' experience with patients who presented with breast lumps and had normal mammograms and normal sonograms. MATERIALS AND METHODS: The findings from 600 lumps in 486 women with no focal ultrasonographic (US) mass or mammographic finding in the area of clinical concern were retrospectively studied. Evaluated parameters included the individual reporting the lump, qualitative descriptors for the physical finding, mammographic density, US characteristics in the area of concern, whether there was a change in imaging and/or physical examination results, and whether there were diagnostic biopsy findings at follow-up. The study group included 540 lumps in 435 women who had a minimum mammographic and clinical follow-up of 2 years, as well as 60 additional lumps in 51 patients who underwent biopsy. RESULTS: No patient in the nonbiopsy group developed carcinoma at the initial site of concern during a mean mammographic and clinical follow-up period of 43 months, and all biopsy specimens were benign (negative predictive value, 100%). CONCLUSION: Results of this retrospective study suggest that breast biopsy may be avoided in women with palpable abnormalities when both US and mammography depict normal tissue at the lump site. PMID- 11274554 TI - MR imaging-guided focused ultrasound surgery of fibroadenomas in the breast: a feasibility study. AB - PURPOSE: To test the feasibility of noninvasive magnetic resonance (MR) imaging guided focused ultrasound surgery (FUS) of benign fibroadenomas in the breast. MATERIALS AND METHODS: Eleven fibroadenomas in nine patients under local anesthesia were treated with MR imaging-guided FUS. Based on a T2-weighted definition of target volumes, sequential sonications were delivered to treat the entire target. Temperature-sensitive phase-difference-based MR imaging was performed during each sonication to monitor focus localization and tissue temperature changes. After the procedure, T2-weighted and contrast material enhanced T1-weighted MR imaging were performed to evaluate immediate and long term effects. RESULTS: Thermal imaging sequences were improved over the treatment period, with 82% (279 of 342) of the hot spots visible in the last seven treatments. The MR imager was used to measure temperature elevation (12.8 degrees -49.9 degrees C) from these treatments. Eight of the 11 lesions treated demonstrated complete or partial lack of contrast material uptake on posttherapy T1-weighted images. Three lesions showed no marked decrease of contrast material uptake. This lack of effective treatment was most likely due to a lower acoustic power and/or patient movement that caused misregistration. No adverse effects were detected, except for one case of transient edema in the pectoralis muscle 2 days after therapy. CONCLUSION: MR imaging-guided FUS can be performed to noninvasively coagulate benign breast fibroadenomas. PMID- 11274556 TI - Mammographic characteristics of 115 missed cancers later detected with screening mammography and the potential utility of computer-aided detection. AB - PURPOSE: To retrospectively determine the mammographic characteristics of cancers missed at screening mammography and assess the ability of computer-aided detection (CAD) to mark the missed cancers. MATERIALS AND METHODS: A multicenter retrospective study accrued 1,083 consecutive cases of breast cancer detected at screening mammography. Prior mammograms were available in 427 cases. Of these, 286 had lesions visible in retrospect. The 286 cases underwent blinded review by panels of radiologists; a majority recommended recall for 112 cases. Two experienced radiologists compared prior mammograms in 110 of these cases with the subsequent screening mammograms (when cancer was detected), noting mammographic characteristics of breast density, lesion type, size, morphology, and subjective reasons for possible miss. The prior mammograms were then analyzed with a CAD program. RESULTS: There were 110 patients with 115 cancers. On the prior mammograms with missed cancers, 35 (30%) of the 115 lesions were calcifications, with 17 of 35 (49%) clustered or pleomorphic. Eighty of the 115 (70%) were mass lesions, with 32 of 80 (40%) spiculated or irregular. For calcifications and masses, the most frequently suggested reasons for possible miss were dense breasts (12 of 35; 34%) and distracting lesions (35 of 80; 44%), respectively. CAD marked 30 (86%) of 35 missed calcifications and 58 (73%) of 80 missed masses. CONCLUSION: Detection errors affected cases with calcifications and masses. CAD marked most (77%; 88 of 115) cancers missed at screening mammography that radiologists retrospectively judged to merit recall. PMID- 11274557 TI - Breast electron boost planning: comparison of CT and US. AB - PURPOSE: To compare computed tomography (CT) with ultrasonography (US) for depiction of the biopsy cavity. MATERIALS AND METHODS: Thirty-two consecutive patients who underwent radiation therapy following lumpectomy with a planned electron boost were examined. At the time of simulation for whole-breast radiation therapy, all patients underwent planning CT (CT 1) at 3-mm section intervals. At the time of electron boost simulation, US was performed to define the biopsy cavity. In 17 cases, a second CT examination (CT 2) was performed at the time of electron boost simulation. CT and US studies were reviewed jointly and assigned a cavity visualization score (CVS) of 1 (cavity not visualized) to 5 (all cavity margins clearly defined). RESULTS: The median CVS at CT 1 was 5; at CT 2, 4; and at US, 4. For patients who underwent all three studies, the median CVS at CT 1 was 5; at CT 2, 4; and at US, 4. Factors related to CVS at CT 1 were homogeneous versus heterogeneous appearance (score, 5 vs 4), surgery-to-CT interval (< or =30 days, 5; 31-60 days, 4; >60 days, 4), and cavity size (>15 cm(3), 5; <15 cm(3), 4). In all cases, cavity volume decreased somewhat during the CT 1-to-CT 2 interval. CONCLUSION: CT performed at the time of whole-breast simulation can be used to plan electron boost fields, with cavity visualization similar to that at US. PMID- 11274558 TI - Carcinoma of the uterine cervix: twice- versus once-weekly high-dose-rate brachytherapy. AB - PURPOSE: To compare the effectiveness and safety of once- versus twice-weekly high-dose-rate (HDR) brachytherapy for cervical cancer. MATERIALS AND METHODS: From 1980 to 1997, 124 consecutive previously untreated patients with cervical cancer were treated with external-beam irradiation (50 Gy) and HDR brachytherapy. Clinical stages were I, 4 (3%) patients; II, 51 (41%); III, 64 (52%); and IV, 5 (4%). From 1980 to 1992, 74 patients (group A) were treated with HDR brachytherapy once weekly (about three fractions of 7 Gy each to point A [2 cm superior and 2 cm lateral to the inferior end of the intrauterine radioactive source]), while from 1992 to 1997, 50 patients (group B) were treated twice weekly (about six fractions of 4.5 Gy each to point A). RESULTS: Overall survival rate at 5 years was 65.2% in group A and 65.3% in group B (P=.96). Local recurrence-free survival rate at 5 years was 69% (51 of 74 patients) in group A and 90% (45 of 50 patients) in group B (P<.001). The rate of grade 2 (moderate) and grade 3 (severe) complications was significantly lower in group B (6% vs. 32% in group A, P<.001). At multivariate analysis, the variables significantly associated with increased local-regional recurrence rates were having stage III IV lesions (P=.04) and with fewer than six sessions of HDR brachytherapy (P=.02). CONCLUSION: The twice-weekly HDR regimen may improve the local control rate with fewer complications. PMID- 11274559 TI - Primary versus secondary ovarian malignancy: imaging findings of adnexal masses in the Radiology Diagnostic Oncology Group Study. AB - PURPOSE: To analyze ultrasonographic (US), computed tomographic (CT), and magnetic resonance (MR) imaging features of primary and secondary ovarian malignant neoplasms to determine if there is any significant difference in their appearance. MATERIALS AND METHODS: Analysis of the multi-institutional Radiology Diagnostic Oncology Group data revealed 86 patients with primary ovarian carcinoma and 24 patients with a secondary ovarian neoplasm. Numerous imaging features that had been recorded for the adnexal masses with each imaging modality were reviewed and compared between primary and secondary malignant ovarian neoplasms. RESULTS: Of the imaging features assessed with all three modalities, multilocularity as determined at US (P =.02) or MR imaging (P: =.01) was the only significant feature. At US, 30 (37%) of 81 primary ovarian cancers were multilocular, whereas only three (12%) of 24 metastatic neoplasms were multilocular. At MR imaging, 40 (74%) of 54 primary ovarian cancers were multilocular, whereas only five (36%) of 14 metastatic neoplasms were multilocular. Neither a predominately solid appearance nor bilaterality was significantly different between primary and secondary neoplasms. CONCLUSION: For malignant ovarian masses, multilocularity at MR imaging or US favors the diagnosis of primary ovarian malignancy rather than secondary neoplasm, but it is difficult to accurately distinguish between primary and secondary ovarian malignancies. PMID- 11274561 TI - The drooping lily sign. PMID- 11274560 TI - Prostate cancer: contrast-enhanced us for detection. AB - PURPOSE: To assess the detection of prostate cancer with contrast material enhanced transrectal sonography. MATERIALS AND METHODS: Sixty subjects were examined with conventional gray-scale, harmonic gray-scale, and power Doppler sonography. Evaluation was repeated during intravenous infusion of contrast agent. Gray-scale imaging was performed in continuous mode and with intermittent imaging by using interscan delay times of 0.5, 1.0, 2.0, and 5.0 seconds. Sextant biopsy sites were scored prospectively as benign or malignant at baseline imaging and again during enhanced transrectal sonography. RESULTS: Prostate cancer was present in 37 biopsy sites from 20 subjects. Baseline imaging demonstrated prostate cancer in 14 sites in 11 subjects. Enhanced transrectal sonography depicted prostate cancer in 24 sites in 15 subjects. Each of the five subjects in whom prostate cancer was missed had only a single biopsy core with positive findings (Gleason score < or = 6). In three of these five subjects, prostate cancer made up less than 10% of the core. The improvement in sensitivity from 38% (14 of 37 malignant foci) at baseline to 65% (24 of 37 malignant foci) with contrast enhancement was significant (P<.004, McNemar chi(2) test). Specificity was similar at baseline (267 [83%] of 323 malignant foci) and during enhanced transrectal sonography (257 [80%] of 323 malignant foci). Clustered receiver operating characteristic analysis demonstrated significant improvement in diagnostic accuracy during enhanced transrectal sonography (P =.027). CONCLUSION: Enhanced transrectal sonography improves sensitivity for the detection of malignant foci within the prostate without substantial loss of specificity. Low volume tumors with a Gleason score of 6 or less may not be detected with enhanced transrectal sonography. PMID- 11274562 TI - Us of blunt abdominal trauma: importance of free pelvic fluid in women of reproductive age. AB - PURPOSE: To assess the importance of free fluid and to determine the accuracy of screening ultrasonography (US) in female patients of reproductive age with trauma. MATERIALS AND METHODS: US was performed in 1,047 patients, aged 10-60 years, to evaluate blunt trauma. Patients were retrospectively assigned to groups on the basis of presence and location of intraperitoneal free fluid. Injury and surgical injury rates were assessed by comparing US results with computed tomographic, repeat US, cystographic, peritoneal lavage, surgical, and/or autopsy findings in 144 patients and with final clinical outcome in 903. US scans were positive if fluid was outside the cul-de-sac or if suspicious parenchymal abnormalities were present. RESULTS: In 939 patients, no fluid was seen: Eight had injuries; three were surgical. In 56, anechoic fluid was isolated to the cul de-sac: Two had injuries; one was surgical. In 26, fluid was isolated to the upper abdomen: Fifteen had injuries; five were surgical. In 22, fluid involved the pelvis and abdomen: Nineteen had injuries; 14 were surgical. In four, questionable fluid was isolated to the supravesical space. Patients with fluid in the cul-de-sac had similar injury and surgical injury rates as those with no fluid but had lower rates than those of patients with fluid elsewhere (P<.02 to P<.001). US screening had 89% sensitivity, 98% specificity, 97% accuracy, a 61% positive predictive value, and a 99% negative predictive value. CONCLUSION: In female patients of reproductive age with trauma, free fluid isolated to the cul de-sac is likely physiologic; clinical follow-up should suffice. Patients with fluid elsewhere usually have clinically important injury and require further evaluation. PMID- 11274564 TI - Pulmonary arteriovenous malformations: three-dimensional gadolinium-enhanced MR angiography-initial experience. AB - PURPOSE: To determine whether three-dimensional gadolinium-enhanced magnetic resonance (MR) angiography could be used to identify pulmonary arteriovenous malformations (PAVMs) and to accurately identify the size and number of feeding arteries. MATERIALS AND METHODS: Eight patients suspected of having PAVM were examined with three-dimensional MR angiography at 1.5 T. Images were reviewed by a single radiologist blinded to conventional angiographic findings who evaluated each image for the size, number, and location of PAVMs, as well as for the size and number of feeding arteries. Five patients underwent conventional angiography with embolization therapy, and one patient underwent lobectomy. Two patients did not undergo either surgery or angiography. RESULTS: Three-dimensional MR angiography revealed nine (90%) of 10 PAVMs that were confirmed at conventional angiography (n = 9) or examination of a surgical specimen (n = 1). The single PAVM that was not identified prospectively at MR angiography was small (3-4 mm) and peripheral. Two additional PAVMs were identified in the two patients who did not undergo surgery or angiography. CONCLUSION: Three-dimensional MR angiography is a promising technique for use in the diagnosis of PAVM, although small (<5-mm) PAVMs may be more difficult to identify with the technique. The technique is a particularly useful means of noninvasively demonstrating the size and number of feeding arteries prior to treatment. PMID- 11274563 TI - Fetal lung volume: estimation at MR imaging-initial results. AB - PURPOSE: To plot normal fetal lung volume (FLV) obtained with fast spin-echo magnetic resonance (MR) images against gestational age; to investigate the correlation between lung growth and fetal presentation, sex, and ultrasonographic (US) biometric measurements; and to investigate its potential application in fetuses with thoracoabdominal malformations. MATERIALS AND METHODS: In a prospective multicenter study, 336 fetuses suspected of having central nervous system disorders underwent fast spin-echo T2-weighted lung MR imaging. Data obtained at 21-38 weeks gestation in 215 fetuses without thoracoabdominal malformations and with normal US biometric findings were selected for an FLV normative curve. FLV measurements obtained at pathologic examination with an immersion method were compared with MR FLV measurements in 11 fetuses. MR FLV values in 16 fetuses with thoracoabdominal malformations were compared with the normative curve. RESULTS: Normal FLV increased with gestational age as a power curve; the spread of values increased with age. Interobserver correlation was excellent (R(2) = 0.96). FLV measurements at MR imaging were 0.90 times those at pathologic examination. A constant ratio (0.78) between FLV on the left and right sides was observed. No significant difference in FLV was observed between fetal presentations. Normal FLV was observed in all fetuses with cystic adenomatoid malformations and in four of six with oligohydramnios. Lowest FLV values were observed in fetuses with diaphragmatic hernia. CONCLUSION: In fetuses with normal lungs, FLV distribution against gestational age is easily assessed in utero with fast spin-echo T2-weighted MR imaging. These preliminary findings illustrate the potential for comparing FLV measurements in fetuses at risk of lung hypoplasia with normative values. PMID- 11274565 TI - Transthoracic needle biopsy of the lung: results of early discharge in 506 outpatients. AB - PURPOSE: To determine the safety of early discharge (30 minutes) after transthoracic needle biopsy (TTNB) of the lung. MATERIALS AND METHODS: In a prospective study of 506 consecutive outpatients who underwent TTNB of the lung, 440 patients underwent fine-needle aspiration biopsy (FNAB) only, and 66 underwent FNAB and core biopsy. Patients were discharged after 30-minute postbiopsy chest radiography if there was no pneumothorax. Patients were discharged after 60-minute chest radiography if they had a stable asymptomatic pneumothorax. These patients were followed up 1 day and/or 1 week after biopsy to identify delayed complications. Patients with a symptomatic or enlarging pneumothorax were treated with an 8-F pigtail catheter attached to a Heimlich valve, discharged, and followed up 24 hours later for chest tube removal. RESULTS: The pneumothorax rate was 22.9% (116 patients). Eighty-one patients (16.0%) had an asymptomatic pneumothorax, and 33 (6.5%) had a pigtail catheter in place. Seven (1.4%) patients developed a symptomatic pneumothorax after discharge; two of them (0.4%) underwent large-bore chest tube insertion. The other five (1.0%) underwent delayed pigtail catheter insertion. There were no deaths or other major complications. CONCLUSION: Early discharge after outpatient TTNB of the lung is associated with little morbidity and no mortality. PMID- 11274566 TI - Evaluation of mediastinal lymphadenopathy with endoscopic US-guided fine-needle aspiration biopsy. AB - PURPOSE: To evaluate the safety and accuracy of endoscopic ultrasonography (US) guided fine-needle aspiration biopsy (FNAB) of lymph nodes in the paratracheal, aortopulmonic, subcarinal, and posterior mediastinal regions. MATERIALS AND METHODS: Eighty-six consecutive patients with mediastinal lymphadenopathy who did not have a primary gastrointestinal neoplasm were examined. In 29 patients, endoscopic US-guided FNAB of mediastinal lymphadenopathy was performed as a component of staging non-small cell lung cancer (NSCLC); in the remaining 57 patients, it was performed to obtain a primary diagnosis. Final diagnosis was based on clinical follow-up, cytologic, and/or surgical results. RESULTS: In 82 patients in whom a final diagnosis was available, the sensitivity, specificity, accuracy, negative predictive value, and positive predictive value of endoscopic US-guided FNAB in distinguishing benign from malignant mediastinal lymph nodes were 96%, 100%, 98%, 94%, and 100%, respectively. In those patients who underwent staging of NSCLC, endoscopic US-guided FNAB had superior mediastinal lymph node staging accuracy compared with endoscopic US alone (79%) and CT alone (79%) (P =.01). The results of endoscopic US-guided FNAB prompted a change to nonsurgical management in 66 (80%) of 82 patients who underwent the procedure. One minor complication, postprocedural fever that resolved with oral antibiotics, occurred (1%; 95% CI: 0%, 6%). CONCLUSION: Endoscopic US-guided FNAB is accurate and safe for biopsy of mediastinal lymph nodes to stage NSCLC, establish a primary diagnosis, or examine patients with prior inconclusive biopsy results. PMID- 11274567 TI - First-pass myocardial perfusion MR imaging with interleaved notched saturation: feasibility study. AB - The authors evaluated a magnetization preparation scheme with a "notched" section profile for T1-weighted first-pass myocardial perfusion magnetic resonance (MR) imaging at 1.5 T. The pulse sequence consisted of a preparation sequence followed by an interleaved gradient-echo echo-planar sequence. Image contrast was evaluated in a feasibility study in 12 adult patients. The notched saturation pulse allowed long magnetization recovery times without sacrificing section coverage. Image contrast between normal and ischemic myocardium was excellent. PMID- 11274568 TI - MR evaluation of ventricular function: true fast imaging with steady-state precession versus fast low-angle shot cine MR imaging: feasibility study. AB - Short- and long-axis cine magnetic resonance (MR) images were obtained with a standard fast low-angle shot, or FLASH, sequence and a first-generation true fast imaging with steady-state precession (FISP) sequence on a 1.5-T MR imager. Contrast-to-noise ratios and volumetric left ventricular measurements were compared for manual and automatic segmentation. True FISP images were associated with significantly (P<.01) higher contrast-to-noise ratios and allowed better detection of the endocardial border. True FISP images were provided with short acquisition times and excellent contrast between the myocardium and the ventricular lumen. PMID- 11274570 TI - Free-breathing black-blood coronary MR angiography: initial results. AB - The authors developed a free-breathing black-blood coronary magnetic resonance (MR) angiographic technique with a potential for exclusive visualization of the coronary blood pool. Results with the MR angiographic technique were evaluated in eight healthy subjects and four patients with coronary disease identified at conventional angiography. This MR angiographic technique accurately depicted luminal disease in the patients and permitted visualization of extensive continuous segments of the native coronary tree in both the healthy subjects and the patients. Black-blood coronary MR angiography provides an alternative source of contrast enhancement. PMID- 11274571 TI - A precious metal alloy for construction of MR imaging-compatible balloon expandable vascular stents. AB - The authors developed ABI alloy, which mechanically resembles stainless steel 316. The main elements of ABI alloy are palladium and silver. Magnetic resonance (MR) images and radiographs of ABI alloy and stainless steel 316 stent models and of nitinol, tantalum, and Elgiloy stents were compared. ABI alloy showed the least MR imaging artifacts and was more radiopaque than stainless steel 316. ABI alloy has the potential to replace stainless steel 316 for construction of balloon-expandable MR imaging-compatible stents. PMID- 11274569 TI - Coronary arteries: magnetization-prepared contrast-enhanced three-dimensional volume-targeted breath-hold MR angiography. AB - A volume-targeted contrast agent-enhanced breath-hold coronary magnetic resonance angiographic technique was optimized and evaluated in 16 volunteers. Substantial increases in coronary signal-to-noise ratio, contrast-to-noise ratio, lengths of depiction, and vessel sharpness were observed on enhanced images. The imaging approach with two 20-mL injections of contrast agent covers the left and right coronary arteries in two breath holds and is a promising method for coronary imaging. PMID- 11274572 TI - Reduction of patient motion artifacts in digital subtraction angiography: evaluation of a fast and fully automatic technique. AB - The performance of an automatic technique for the reduction of patient motion artifacts in digital subtraction angiography was evaluated. Four observers assessed the quality of 104 cerebral digital subtraction angiographic images that were processed by means of both the automatic technique and manual pixel shifting. The automatic technique resulted in better image quality and was considerably less time-consuming. PMID- 11274573 TI - MR imaging and electronically activated devices. PMID- 11274574 TI - Lack of effect of position restriction after transthoracic biopsy. PMID- 11274575 TI - Is excision of all lobular carcinoma in situ really necessary? PMID- 11274576 TI - Diagnosis of acute appendicitis. PMID- 11274577 TI - Metastatic liver tumor: circumferential versus wedge-shaped perilesional enhancement and quantitative image and pathologic correlation. PMID- 11274580 TI - Measuring the volume of packed RBCs in donors. PMID- 11274581 TI - Transfusion-related acute lung injury: femme fatale? PMID- 11274584 TI - Hyperhemolytic transfusion reaction in sickle cell disease. AB - BACKGROUND: An atypical form of life-threatening hemolytic transfusion reaction (HTR) in patients with sickle cell disease (SCD) has been well described in the literature. Continuation of blood transfusion may be lethal, as it can further exacerbate hemolysis. The pathophysiologic mechanism of HTR is not well understood. CASE REPORTS: Two cases of severe HTR in SCD after the transfusion of compatible RBC units are reported. Hemolysis of both autologous and transfused cells was documented in Case 1 by urine Hb high-performance liquid chromotography. Multispecific HLA antibodies were identified in Case 1. Reticulocytopenia was noted in both cases during the acute hemolytic process. This was followed by a rise in reticulocyte count during receipt of IVIG and steroid therapy. Bone marrow examination during reticulocytopenia in Case 2 showed erythroid hyperplasia. CONCLUSION: In SCD, both mature sickle cells and sickle reticulocytes adhere more readily to macrophages. In view of the bone marrow aspiration results, it appears that the recipients' HbS cells are destroyed by hyperactive macrophages and that the reticulocytopenia observed during HTR is likely to be due to peripheral consumption (i.e., destruction by macrophages), rather than suppression of erythropoiesis. Cessation of hemolysis during IVIG and steroid treatment may be due to IVIG's blocking of the adhesion of sickle cells and reticulocytes to macrophages, together with steroid suppression of macrophage activity. PMID- 11274583 TI - A randomized controlled trial oftransfusion-related acute lung injury: is plasma from multiparous blood donors dangerous? AB - BACKGROUND: Transfusion-related acute lung injury (TRALI) and other posttransfusion reactions may be caused by granulocyte and/or HLA antibodies, which are often present in blood from multiparous donors. The purpose of this study was to compare the effects of plasma from multiparous donors with those of plasma from donors with no history of transfusion or pregnancy (control plasma) in a prospective, randomized, double-blind, crossover study. STUDY DESIGN AND METHODS: Intensive care patients, judged to need at least 2 units of plasma, were randomly assigned to receive a unit of control plasma and, 4 hours later, a plasma unit from a multiparous donor (> or = 3 live births) or to receive the plasma units in opposite order. The patients were closely monitored, and body temperature, blood pressure, and heart rate were recorded. Blood samples for analysis of blood gases, TNFalpha, IL-1 receptor antagonist, soluble E selectin, and C3d complement factor were collected at least on four occasions (before and after the transfusion of each unit). RESULTS: Transfusion of plasma from multiparous donors was associated with significantly lower oxygen saturation and higher TNFalpha concentrations than transfusion of control plasma. The mean arterial pressure increased significantly after the transfusion of control plasma, whereas plasma from multiparous donors had no effect on it. Five posttransfusion reactions were observed in 100 patients, in four cases after the transfusion of plasma from multiparous donors. CONCLUSION: Plasma from multiparous blood donors may impair pulmonary function in intensive care unit patients. PMID- 11274585 TI - Infectivity of blood from PCR-positive, HBsAg-negative, anti-HBs-positive cases of resolved hepatitis B infection. AB - BACKGROUND: Numerous reports have noted the existence of sera, particularly from resolving cases of HBV infection, that are positive for HBV DNA by PCR, despite being negative for HBsAg and IgM anti-HBc. If such blood is infective and detectable by HBV NAT screening, it seems desirable to introduce such screening for transfused blood. STUDY DESIGN AND METHODS: Three chimpanzees were inoculated with serum and lymphocytes from three patients who were HBV DNA PCR positive, but HBsAg negative. The animals were tested over a period of 15 months for HBsAg, anti-HBs, anti-HBc, and HBV DNA by PCR. RESULTS: All animals remained uninfected. CONCLUSION: Small amounts of plasma and MNCs from HBV DNA-positive HBsAg-negative blood do not appear to be infectious; however, further studies with larger volumes of inoculum should be conducted. PMID- 11274586 TI - Routine HCV PCR screening of blood donations to identify early HCV infection in blood donors lacking antibodies to HCV. AB - BACKGROUND: Detection of early hepatitis C infection of blood donors is still a major problem for blood transfusion. Common anti-HCV screening assays show differences in sensitivity and specificity. The often mild symptoms of acute hepatitis C also cause difficulties in the identification of early HCV infection. The feasibility and efficacy of routine screening of blood donations for HCV RNA were investigated. STUDY DESIGN AND METHODS: Blood donations (n = 251,737) were screened for HCV RNA over 4 years. RNA extraction, amplification, and detection were done by two commercial HCV PCR kits (HCV Cobas Amplicor and HCV Cobas Amplicor 2.0, Roche Diagnostics). Screening was done by pool testing with a maximum pool size of 40 serum samples. RESULTS: Three donations out of 251,737 were HCV RNA positive and anti-HCV negative. ALT levels of these donations were 271, 32, and 10 U per L. The HCV infection of a fourth HCV RNA-positive donor could not be identified by routine, second-generation HCV EIA (Abbott Diagnostika). In this case, two previous donations were also HCV RNA positive, and three second-generation test systems (Abbott) could not detect anti-HCV, whereas third-generation anti-HCV screening assays detected antibody with different sensitivity. The first HCV RNA-positive donation was identified only by the HCV ELISA 3.0 (Ortho Diagnostic Systems). The results of confirmatory assays like RIBA HCV 3.0 (Ortho) and Matrix (Abbott) indicate a restricted immune response to NS3 only. CONCLUSION: HCV RNA detection by PCR can be carried out routinely in blood donor screening without significant delay of release of the components. The residual risk of transmission can be reduced by identification of early infection, which can lead to an improved safety of blood components. RNA screening can also be advantageous in cases of incomplete or lack of antibody response to HCV. PMID- 11274587 TI - Risk factors for infection with HBV and HCV in a largecohort of hemophiliac males. AB - BACKGROUND: Before the implementation of donor screening and the development of effective virus-inactivation procedures, persons with hemophilia (PWHs) were at risk of infection with HBV and HCV transmitted through clotting factor concentrates. STUDY DESIGN AND METHODS: Data collected from the medical records of a cohort of 2,772 males with hemophilia who resided in six states of the United States were used to examine relations between demographic and clinical characteristics and laboratory markers of past or present infection with HBV and HCV using logistic regression. RESULTS: Test results were available for 60 percent of the cohort. Among those tested, 30 percent were positive for markers of HBV infection and 64 percent for HCV infection. Factors associated with increased odds of positive HBV markers and HCV infection were greater severity of hemophilia, larger amounts of factor use, and HIV infection. Markers of HBV infection persisted in birth cohorts as late as 1992 and those of HCV infections in birth cohorts through 1991. Compared to same-age US males, PWHs born between 1987 and 1989 were more likely to have markers of HBV and HCV infection. CONCLUSION: PWHs who received clotting factor concentrates before 1990 may be at risk for infection with hepatitis B or hepatitis C and should be tested. PMID- 11274589 TI - Accuracy of predonation Hct sampling affects donor safety, eligibility, and deferral rates. AB - BACKGROUND: Safe blood donation depends upon reliable predonation Hct screening. Earstick (ES) capillary samples are frequently used, but they may not be accurate. STUDY DESIGN AND METHODS: Predonation ES and fingerstick (FS) and postdonation venous Hct results were compared in 1960 whole-blood and 210 apheresis donors. The validity of using postdonation venous samples to evaluate predonation ES and FS Hct was assessed in 20 whole-blood donors. The impact of Hct screening method on donor Hct deferrals was examined during periods when either ES or FS sampling was used exclusively. RESULTS: All donors were eligible to donate on the basis of a predonation capillary Hct of > or = 38 percent. In venous samples obtained immediately after donation, 36 percent of whole-blood donors had a Hct <38 percent. With correction for a decrease of approximately 2 Hct units during donation, 20 percent of these donors had a predonation Hct <38 percent. The lowest venous Hct was 23.1 percent. FS samples showed better correlation with venous Hct. Hct discrepancies were similar for apheresis donors. Hct deferrals were significantly higher with FS sampling, especially among women. CONCLUSION: Hct determinations from ES samples overestimate venous Hct. Fingerstick samples are more sensitive in detecting anemia. The accuracy of predonation Hct sampling has implications for donor safety, eligibility, and deferral rates. PMID- 11274588 TI - Dextran sedimentation in a semi-closed system for the clinical banking of umbilical cord blood. AB - BACKGROUND: The results of current processing procedures for reducing volume and recovering HPCs from umbilical cord blood (UCB) before cryopreservation vary. STUDY DESIGN AND METHODS: Dextran was added to bags containing UCB, followed by sedimentation for 30 minutes. The processed UCB was then frozen. RBCs, nucleated cells, MNCs, CD34+ cells, CFUs and long-term culture-initiating cells (LTC-ICs), viability, and sterility were evaluated. Fractionations in ficoll-hypaque and hydroxyethyl starch (HES) were also run in parallel for comparison. RESULTS: The nucleated cell (NC) recovery and RBC depletion were 86.1 percent and 94.3 percent, respectively (n = 50). Sedimentation with dextran also enabled the recovery of 80.7 percent MNCs and 82.6 percent CD34+ cells (n = 30). Postsedimentation samples displayed no impairment of CFU growth (n = 42, 108.7% CFU-C, 104.6% CFU-GEMM, 107% CFU-GM, and 95.7% BFU-E). Long-term cultures on five paired samples before and after sedimentation generated similar numbers of CFU-C each week (p = 0.88). Limiting dilution analysis of 12 paired pre/postsedimentation samples showed comparable median proportions of LTC-ICs (1/6494 vs. 1/5236; p = 0.18). The cell viability of 24 samples of thawed UCB after sedimentation was 90.3 percent (77.5-96%) and the recovery of CFU-C, CFU GEMM, CFU-GM, and BFU-E of 11 postsedimentation samples was 93.4 percent, 84.9 percent, 92.3 percent, and 83.4 percent, respectively. NC recovery was significantly higher after treatment with dextran than with ficoll-hypaque (n = 30; 88.5% vs. 29.1%; p<0.005) and HES treatment (n = 21; 88.5% vs. 76.4%; p = 0.004). However, MNCs, CD34+ cells, CFUs, LTC-ICs, and RBCs were comparable. Two cycles of dextran sedimentation recovered 93.9 percent of NCs with cell viability of 98.6 percent (96.5-100%), whereas 11.7 percent of RBCs were retained (n = 20). The final yield volume was 33.5 (28-41) mL. CONCLUSION: In a semi-closed system, dextran sedimentation enabled volume reduction of UCB without significant quantitative and qualitative losses of HPCs. PMID- 11274590 TI - First-time blood donors: demographic trends. AB - BACKGROUND: With changing demographics of the United States population and the continuous need to recruit new donors, it is important to monitor the demographic profile of first-time donors and to evaluate changes in the donor pool to improve recruitment targeting. STUDY DESIGN AND METHODS: First-time whole blood (n = 901,862) donors at five United States blood centers between 1991 and 1996 were analyzed. RESULTS: The total number of first-time donors appears to be decreasing. Over the 6-year period, there was an overall increase in the proportion of Hispanic and other minority first-time donors and a concurrent decrease in the proportion of white donors at Retrovirus Epidemiology Donor Study centers. Other variables, including age, sex, and education, did not show a consistent trend. CONCLUSION: The demographic profile of first-time donors is changing. These data highlight the importance for blood centers to continuously monitor the donor population. A better understanding of the donor population may help blood centers adjust their donor outreach, recruitment, and retention programs. New recruitment efforts appear needed to counter general apathy toward donating blood, and minority groups appear to be receptive to becoming blood donors. PMID- 11274591 TI - Donor-derived alloantibodies and passenger lymphocyte syndrome in two of four patients who received different organs from the same donor. AB - BACKGROUND: Reported here is the occurrence of RBC alloimmunization in two of four patients who received different organs from an immunized donor. STUDY DESIGN AND METHODS: The donor, a 58-year-old woman, was group O D+, K-, and Fy(a-). Initially, her serum contained only a K antibody. After blood transfusion, a second antibody (anti-Fy(a)) could also be identified. The liver was given to a group O D+, K-, Fy(a+) patient; the pancreas and one kidney to a group O D+, K-, Fy(a+) patient; the heart to a group A D+, K-, Fy(a-) patient; and the other kidney to a group B D+, K-, Fy(a+) patient. RBC grouping and antibody screening were performed by standard techniques. Lymphoid microchimerism in the peripheral blood of the recipients was analyzed by flow cytometry and nested PCR. RESULTS: None of the recipients had irregular RBC alloantibodies at the time of transplantation. After the transplant, anti-K became detectable in the serum of the liver recipient, and anti-Fy(a) could be eluted from the RBCs of the liver recipient and the pancreas-kidney recipient. The latter patient also developed mild hemolysis, and his Hb dropped to 8 g per dL on posttransplant Day 9. Donor derived lymphocytes were detectable by flow cytometry in the peripheral blood of the liver recipient and the pancreas-kidney recipient until Days 8 and 63, respectively, whereas no lymphoid chimerism could be demonstrated in the heart recipient. PCR chimerism analyses were positive in all three recipients over the whole observation period of 97 postoperative days. CONCLUSION: The amount of cotransplanted lymphoid tissue may correlate with the extent of peripheral lymphoid microchimerism and the antibody-formation capacity in solid organ transplantation. PMID- 11274592 TI - The first example of a paraben-dependent antibody to an Rh protein. AB - BACKGROUND: Parabens are added to a commercial LISS (C-LISS) to retard microbial growth. Paraben-dependent anti-Jk(a) has been detected by the use of C-LISS. CASE REPORT: Serum from a D+ woman reacted in antiglobulin tests with RBCs stored (2-4 hours, 22-25 degrees C) in C-LISS (Low and Messeter formulation, Immucor). Freshly prepared C-LISS-suspended RBCs did not react; nor did RBCs stored in LISS additive reagents, PEG, saline, or homemade LISS. RESULTS: Studies using C-LISS stored RBCs revealed an antibody that reacted with D+ and rrV+ RBCs, but not with r'r, r"r, or rrV-VS- RBCs. All partial D RBC phenotypes tested reacted, as did D+LW-, rGr, r"Gr, r(y)r, r'(s)rV+VS+, and r'(s)rV-VS+ RBCs. The active ingredient in C-LISS was propylparaben. Other LISS ingredients were not required; saline solutions of propylparaben, ethylparaben, methyl salicylate, 2-phenoxyethanol, and butylparaben were active. Methylparaben and methyl-m-hydroxybenzoate were inactive. Reactivity to C-LISS-stored RBCs could not be inhibited by propylparaben. Reactivity with D+V- and D-V+VS+ RBCs was not separable by adsorption-elution. CONCLUSIONS: This antibody likely detects a neoantigen formed between active compounds and RBC membranes. Review of the structure of active compounds suggests that proximity between methyl and hydroxyl groups is important for binding with RBC membranes. The role of RhD is unclear; no single portion of RhD protein appears to be implicated. PMID- 11274593 TI - Neonatal alloimmune thrombocytopenia due to anti-Nak(a). AB - BACKGROUND: The accurate diagnosis of neonatal alloimmune thrombocytopenia is essential in the effective treatment of potentially serious bleeding in neonates. CASE REPORT: Reported here is a case of a full-term female baby who was delivered by vacuum extraction from a gravida 1 para 1 healthy mother. She presented with generalized petechiae and bilateral cephalhematoma, which she had had since birth. At 7 hours of life, she had an upper gastrointestinal hemorrhage and was found to have severe anemia and marked thrombo-cytopenia. Coagulation screening tests were normal. The diagnosis of neonatal alloimmune thrombocytopenia was suspected, and maternal serum was collected for further study. The baby was treated with a single dose of hydrocortisone (10 mg/kg) and IVIG (400 mg/kg) while waiting for irradiated platelets from her mother. After 30 mL of a transfusion of maternal platelets, the baby's platelet count rose dramatically, from 15,000 to 162,000 per microL, and it remained stable at that level. She was discharged on the 10th hospital day in good condition. During the follow-up period of 8 months, her growth and development were satisfactorily normal, as well as her platelet count. A high-titered platelet antibody was detected in the maternal serum by use of a solid phase platelet adherence technique. RESULTS: The specificity of the platelet antibody was identified as anti-Nak(a) by the mixed passive hemagglutination test method. CONCLUSION: These findings suggested a diagnosis of NAIT caused by anti-Nak(a). PMID- 11274594 TI - Fy phenotype and gender determine plasma levels of monocyte chemotactic protein. AB - BACKGROUND: In vitro studies indicate that the Fy blood group system antigens serve as receptors for chemokines such as monocyte chemotactic protein-1 (MCP-1) and RANTES. However, it is unclear whether subjects with the Fy(a-b-) phenotype exhibit altered clearance and hence altered plasma levels of chemo-kines, because they still express Fy on endothelial cells. STUDY DESIGN AND METHODS: To clarify a possible in vivo role of Fy on RBCs in the regulation of chemo-kine levels, healthy young volunteers of common Fy phenotypes were compared in a cross sectional study. RESULTS: More than 90 percent of the 34 subjects of African origin were Fy(a-b-), one black volunteer was Fy(a+b-), and two were Fy(a-b+). As expected, all 65 white volunteers were positive for either Fy(a) and/or Fy(b). Unexpectedly, persons expressing either Fy(a) and/or Fy(b) had significantly higher plasma levels of MCP-1 than Fy(a-b-) volunteers (women: 154 vs. 110 ng/L, p<0.01; men: 179 vs. 169 ng/L, p = 0.03). Surprisingly, plasma levels of MCP-1 were found to be sex-dependent: median MCP-1 levels averaged 180 ng per L in men but only 139 ng per L in women (p<0.001). Further, MCP-1 levels decreased significantly throughout the menstrual cycle of 18 women studied longitudinally. CONCLUSION: MCP-1 levels are about 30 percent higher in men than in premenopausal women, and MCP-1 levels are also higher in persons with RBCs expressing Fy antigens than in Fy(a-b-) persons. These findings have direct implications for the concept and interpretation of clinical studies measuring MCP-1 levels; the role of the observed differences in MCP-1 levels for the pathogenesis of MCP-1 dependent diseases, such as atherosclerosis, merits further investigation. PMID- 11274595 TI - Virus inactivation of plasma-derived proteins by pasteurization in the presence of guanidine hydrochloride. AB - BACKGROUND: Viruses, among them parvovirus B19 and other small, nonenveloped viruses, may be present in human blood and may contaminate plasma-derived therapeutics. Efficient inactivation or removal of such viruses, especially parvoviruses, represents a current problem and corresponding technologies are under investigation. In this report, such a technology is described. STUDY DESIGN AND METHODS: A recently developed pasteurization of human apolipoprotein A-I (apoA-I), which is performed at 60 degrees C for 10 hours in the presence of guanidine hydrochloride (GdnHCl), was validated by using a series of model viruses, including members of the families parvoviridae and picornaviridae. The model viruses were spiked into the apoA-I- and GdnHCl-containing solutions, and virus inactivation was evaluated by infectivity assays in cell cultures. The mechanism of virus inactivation was studied by virus sedimentation analysis using the picornavirus model. RESULTS: All viruses tested were inactivated to levels below the limit of detection, although different inactivation kinetics were obtained for the different viruses. The mechanism of virus inactivation by this pasteurization was disassembly of the virus particles into single proteins or small noninfectious viral subunits. CONCLUSION: The pasteurization validated in this report has the potential to inactivate a wide range of transfusion-relevant viruses including parvoviruses and picornaviruses. PMID- 11274596 TI - Serial granulocytapheresisunder daily administration of rHuG-CSF: effects on peripheral blood counts, collection efficiency, and yield. AB - BACKGROUND: An option for treatment of severe infections in neutropenic patients is the transfusion of granulocytes from donors stimulated with rHuG-CSF. The schedule of rHuG-CSF-stimulated granulocyte donations and the quality of the components remain controversial. This study was done with the intention of ensuring daily granulocyte support with therapeutic cell numbers, while keeping the patients' allogeneic exposure as low as possible. STUDY DESIGN AND METHODS: Granulocyte collection with multiple consecutive leukapheresis procedures under daily rHuG-CSF administration and with hydroxyethyl starch as sedimenting agent were prospectively studied. Complete blood counts of the donors, collection yield, and efficiency were analyzed. RESULTS: Products (n = 259) from 76 donors were examined. The median peripheral blood WBC and neutrophil counts were 28.1 g per L and 24.1 g per L, respectively, and they were significantly higher on Day 5 of collections than on Days 1 to 3. Platelet counts and Hb levels decreased steadily. Collection yields increased over time from 4.9 to 6.7 x 10(10) neutrophils. Side effects of cytokines and aphereses did not exceed World Health Organization grade II status. CONCLUSION: Repetitive daily rHuG-CSF administration-even under daily leukapheresis procedures-results in a continuing increase in WBC and neutrophil levels and thus leads to increased collection yields. Side effects are tolerable, although Hb and platelet levels should be monitored closely. PMID- 11274597 TI - Comparison of a new WBC-reduction system and the standard plateletpheresis protocol in the same donors. AB - BACKGROUND: A cell separator (Spectra, Gambro BCT) with an integrated leukoreduction system (LRS) for producing WBC-reduced single-donor platelet concentrates has been shown to result in a slightly reduced collection efficiency as compared to the former Spectra system without LRS. A novel modified system for improved collection efficiencies (LRS Turbo, Gambro BCT) was evaluated. STUDY DESIGN AND METHODS: Each of 37 donors underwent plateletpheresis using the LRS Turbo (LRS-T) and the standard LRS (LRS) of the Spectra cell separator. The collection efficiency and WBC contamination of the different techniques were compared. Platelets were counted automatically and WBCs were counted by using one or two full grids of a Nageotte chamber. RESULTS: The preseparation and postseparation numbers of RBCs, WBCs, and platelets, as well as the number of collected platelets, did not differ for the two techniques. In the LRS-T separations, the collection efficiency was 112 percent of that in the LRS procedures. Median residual WBCs in the platelet components were 0.0256 x 10(6) per LRS-T procedure and 0.0253 x 10(6) per LRS procedure. The purity of the LRS-T components was not less than that of the standard LRS components, whereas the collection efficiency of the LRS-T was significantly greater, 44.9 percent versus 40.7 percent. CONCLUSIONS: The LRS-T procedures produced platelet concentrates with WBC-reduction capacity that is comparable to that obtained with the standard LRS procedures, which have previously been described as satisfying the most stringent criteria for WBC-reduced platelets. The new technique significantly improved the collection efficiency of the plateletpheresis procedure. PMID- 11274598 TI - In vitro and in vivo measurements of human RBCs frozen with glycerol and subjected to various storage temperatures before deglycerolization and storage at 4 degrees C for 3 days. AB - BACKGROUND: This study was designed to assess the effects of changes in storage temperature of frozen RBCs such as might occur during a malfunction of the -80 degrees C mechanical freezer or during shipment. STUDY DESIGN AND METHODS: Fifteen participants donated blood for autologous transfusion of RBCs; all RBCs were frozen with 40-percent (wt/vol) glycerol. Five subjects received RBCs that were stored at -80 degrees C alone before transfusion. Five subjects received RBCs that were stored initially at -80 degrees C, then at -40 degrees C for 4 weeks, and finally at -80 degrees C before transfusion. Five subjects received RBCs that were stored at -80 degrees C, then at -20 degrees C for 2 weeks, and finally at -80 degrees C before transfusion. After deglycerolization, the RBCs were stored at 4 degrees C in a sodium chloride-glucose solution for 3 days before transfusion. RESULTS: No significant differences were observed in freeze thaw recovery, freeze-thaw-wash recovery, 24-hour posttransfusion survival, index of therapeutic effectiveness, or RBC ATP levels. Greater hemolysis and reduced RBC K+ levels were observed in the units stored at -80 degrees C/-40 degrees C/ 80 degrees C and in those stored at -80 degrees C/ -20 degrees C/-80 degrees C compared with the units stored at -80 degrees C alone, but these differences did not affect the 24-hour posttransfusion survival. CONCLUSIONS: The results of this study indicated that RBCs frozen with 40-percent (wt/vol) glycerol can be stored at -40 degrees C for 4 weeks or at -20 degrees C for 2 weeks between periods of frozen storage at -80 degrees C with satisfactory results. PMID- 11274599 TI - Meta-analysis of controlled clinical trials studying the efficacy of rHuEPO in reducing blood transfusions in the anemia of prematurity. AB - BACKGROUND: Recombinant human EPO (rHuEPO) has not gained broad acceptance in the treatment of the anemia of prematurity, because its efficacy in diminishing RBC transfusions is questionable. Meta-analysis was used to investigate the extent and reasons for variation in the results of published clinical trials. STUDY DESIGN AND METHODS: Prospective controlled trials published from 1990 through 1999 were retrieved; 21 met the criteria for meta-analysis. Calculated across these studies were the summary OR of RBC transfusion in treated neonates as compared with controls and the summary mean difference between controls and treated neonates in the volume of RBCs transfused and the number of RBC transfusions per infant. Twelve study descriptors were examined as possible reasons for the variation in results. RESULTS: Results of 21 eligible studies varied widely (p<0.001 for the Q test statistic), and this variation persisted in most analyses when studies were stratified by individual study descriptors. When the difference in volume of RBCs transfused was the outcome measure, variation was modest across the four studies with highly desired characteristics (i.e., high blindness and design quality scores, "conservative" transfusion criteria, and the majority of neonates weighing <1 kg at birth), and treatment with rHuEPO reduced RBC transfusions by an average of 11.0 mL per kg (p<0.001). CONCLUSION: Benefit from rHuEPO is detected across high-quality studies using conservative RBC transfusion criteria. However, there is extreme variation overall in the findings of available trials, and-until this variation is accounted for-it is premature to recommend rHuEPO as standard treatment for the anemia of prematurity. PMID- 11274600 TI - Fas, Fas ligand,and transfusion immunomodulation. PMID- 11274601 TI - Microchimerism, GVHD, and tolerance in solid organ transplantation. AB - The phenomenon of microchimerism and its relationship to long-term graft tolerance is an area of active study. The ability to establish a tolerant state has been enhanced with current immunosuppressive drugs and emerging therapies such as donor HPC infusions. An undesirable outcome of host-donor WBC interaction is GVHD. GVHD is a rare complication reported most frequently in liver transplantation. Two cases of GVHD reported in recipients of organs from donors homozygous for a shared HLA haplotype would support a policy of avoiding the use of these donors. TA-GVHD is very rare in solid organ transplant recipients, with only four published cases; only two had convincing supportive evidence and one of these had an underlying hematologic abnormality. These few cases do not support a policy of routine irradiation of cellular blood components for all solid organ transplant recipients. The use of donor HPC infusions to enhance chimerism and graft tolerance has increased the number of GVHD cases observed (usually mild) and decreased the severity and number of rejection episodes. The long-term effects of donor HPC infusions on graft survival is under investigation. PMID- 11274602 TI - A second example of a transfusion-associated septic reaction associated with Clostridium perfringens. PMID- 11274603 TI - Long-term hepatitis C seroconversion in a blood donor. PMID- 11274604 TI - Erroneous HCV genotype assignment by a hybridization typing assay in a case of posttransfusion HCV infection. PMID- 11274605 TI - Dispersive Pad Injuries Associated with Hysteroscopic Surgery. PMID- 11274606 TI - GnRH Analogs and Uterine Artery Embolization. PMID- 11274608 TI - Hysteroscopic Fluid Monitoring Guidelines. PMID- 11274609 TI - Development of Flexible Culdoscopy. PMID- 11274607 TI - An Open Letter to the American Association of Gynecologic Laparoscopists. PMID- 11274610 TI - Laparoscopic Suturing. PMID- 11274611 TI - Genital Tract Burns. PMID- 11274612 TI - Presidential Report. PMID- 11274613 TI - Tribute to Jordan M. Phillips, M.D. PMID- 11274614 TI - Laparoscopic complications. PMID- 11274615 TI - Laparoscopic closure of patent canal of Nuck for female indirect inguinal hernia. PMID- 11274616 TI - Retrocervical, retrovaginal pouch, and rectovaginal septum endometriosis. PMID- 11274618 TI - Laparoscopic-assisted vaginal hysterectomy--experience of an Italian university tertiary hospital. PMID- 11274617 TI - Abnormal uterine bleeding: surgical management--part III. PMID- 11274619 TI - Laparoscopic-Assisted Vaginal Hysterectomy-Experience of an Italian University Tertiary Hospital. PMID- 11274620 TI - Noninvasive diagnosis of renal-allograft rejection by measurement of messenger RNA for perforin and granzyme B in urine. AB - BACKGROUND: Acute rejection is a serious and frequent complication of renal transplantation, and its diagnosis is contingent on the invasive procedure of allograft biopsy. A noninvasive diagnostic test for rejection could improve the outcome of transplantation. METHODS: We obtained 24 urine specimens from 22 renal allograft recipients with a biopsy-confirmed episode of acute rejection and 127 samples from 63 recipients without evidence of acute rejection. RNA was isolated from the urinary cells. Messenger RNA (mRNA) encoding the cytotoxic proteins perforin and granzyme B and a constitutively expressed cyclophilin B gene were measured with the use of a competitive, quantitative polymerase chain reaction, and the level of expression was correlated with allograft status. RESULTS: The log-transformed mean (+/-SE) levels of perforin mRNA and granzyme B mRNA, which encode cytotoxic proteins, but not the levels of constitutively expressed cyclophiiin B mRNA, were higher in the urinary cells from the 22 patients with a biopsy-confirmed episode of acute rejection than in the 63 recipients without an episode of acute rejection (perforin, 1.4+/-0.3 vs. -0.6+/-0.2 fg per microgram of total RNA; P<0.001; and granzyme B, 1.2+/-0.3 vs. -0.9+/-0.2 fg per microgram of total RNA; P<0.001). Analysis involving the receiver-operating-characteristic curve demonstrated that acute rejection can be predicted with a sensitivity of 83 percent and a specificity of 83 percent with the use of a cutoff value of 0.9 fg of perforin mRNA per microgram of total RNA, and with a sensitivity of 79 percent and a specificity of 77 percent with the use of a cutoff value of 0.4 fg of granzyme B mRNA per microgram of total RNA. Sequential urine samples were obtained from 37 patients during the first nine days after transplantation; and measurements of the levels of mRNA that encoded cytotoxic proteins identified those in whom acute rejection developed. CONCLUSIONS: Measurement of mRNA encoding cytotoxic proteins in urinary cells offers a noninvasive means of diagnosing acute rejection of renal allografts. PMID- 11274621 TI - The effectiveness of the varicella vaccine in clinical practice. AB - BACKGROUND: A live attenuated varicella vaccine was approved for use in the United States in March 1995 and is recommended for all susceptible persons 12 months of age or older. METHODS: To assess the effectiveness of the varicella vaccine, we conducted a case-control study with two controls per child with chickenpox, matched according to both age and pediatric practice. Children with potential cases of chickenpox were identified by active surveillance of pediatric practices in the New Haven, Connecticut, area. Research assistants visited the children on day 3, 4, or 5 of the illness, assessed the severity of the illness, and collected samples from lesions to test for varicella-zoster virus by polymerase chain reaction (PCR). RESULTS: From March 1997 through November 2000, data collection was completed for 330 potential cases, of which 243 (74 percent) were in children who had positive PCR tests for varicella-zoster virus. Of the 56 vaccinated children with chickenpox, 86 percent had mild disease, whereas only 48 percent of the 187 unvaccinated children with chickenpox had mild disease (P<0.001). Among the 202 children with PCR-confirmed varicella-zoster virus and their 389 matched controls, 23 percent of the children with chickenpox and 61 percent of the matched controls had received the vaccine (vaccine effectiveness, 85 percent; 95 percent confidence interval, 78 to 90 percent; P<0.001). Against moderately severe and severe disease the vaccine was 97 percent effective (95 percent confidence interval, 93 to 99 percent). The effectiveness of the vaccine was virtually unchanged (87 percent) after adjustment for potential confounders by means of conditional logistic regression. CONCLUSIONS: Varicella vaccine is highly effective as used in clinical practice. PMID- 11274622 TI - Paroxetine for the prevention of depression induced by high-dose interferon alfa. AB - BACKGROUND: Depression commonly complicates treatment with the cytokine interferon alfa-2b. Laboratory animals pretreated with antidepressants have less severe depression-like symptoms after the administration of a cytokine. We sought to determine whether a similar strategy would be effective in humans. METHODS: In a double-blind study of 40 patients with malignant melanoma who were eligible for high-dose interferon alfa therapy, we randomly assigned 20 patients to receive the antidepressant paroxetine and 20 to receive placebo. The treatment was begun 2 weeks before the initiation of interferon alfa and continued for the first 12 weeks of interferon alfa therapy. RESULTS: During the first 12 weeks of interferon alfa therapy, symptoms consistent with a diagnosis of major depression developed in 2 of 18 patients in the paroxetine group (11 percent) and 9 of 20 patients in the placebo group (45 percent) (relative risk, 0.24; 95 percent confidence interval, 0.08 to 0.93). Severe depression necessitated the discontinuation of interferon alfa before 12 weeks in 1 of the 20 patients in the paroxetine group (5 percent), as compared with 7 patients in the placebo group (35 percent) (relative risk, 0.14; 95 percent confidence interval, 0.05 to 0.85). The incidence of adverse events was similar in the two groups. CONCLUSIONS: In patients with malignant melanoma, pretreatment with paroxetine appears to be an effective strategy for minimizing depression induced by interferon alfa. PMID- 11274623 TI - Preventing recurrent upper gastrointestinal bleeding in patients with Helicobacter pylori infection who are taking low-dose aspirin or naproxen. AB - BACKGROUND: Many patients who have had upper gastrointestinal bleeding continue to take low-dose aspirin for cardiovascular prophylaxis or other non-steroidal antiinflammatory drugs (NSAIDs) for musculoskeletal pain. It is uncertain whether infection with Helicobacter pylori is a risk factor for bleeding in such patients. METHODS: We studied patients with a history of upper gastrointestinal bleeding who were infected with H. pylori and who were taking low-dose aspirin or other NSAIDs. We evaluated whether eradication of the infection or omeprazole treatment was more effective in preventing recurrent bleeding. We recruited patients who presented with upper gastrointestinal bleeding that was confirmed by endoscopy. Their ulcers were healed by daily treatment with 20 mg of omeprazole for eight weeks or longer. Then, those who had been taking aspirin were given 80 mg of aspirin daily, and those who had been taking other NSAIDs were given 500 mg of naproxen twice daily for six months. The patients in each group were then randomly assigned separately to receive 20 mg of omeprazole daily for six months or one week of eradication therapy, consisting of 120 mg of bismuth subcitrate, 500 mg of tetracycline, and 400 mg of metronidazole, all given four times daily, followed by placebo for six months. RESULTS: We enrolled 400 patients (250 of whom were taking aspirin and 150 of whom were taking other NSAIDs). Among those taking aspirin, the probability of recurrent bleeding during the six-month period was 1.9 percent for patients who received eradication therapy and 0.9 percent for patients who received omeprazole (absolute difference, 1.0 percent; 95 percent confidence interval for the difference, -1.9 to 3.9 percent). Among users of other NSAIDs, the probability of recurrent bleeding was 18.8 percent for patients receiving eradication therapy and 4.4 percent for those treated with omeprazole (absolute difference, 14.4 percent; 95 percent confidence interval for the difference, 4.4 to 24.4 percent; P=0.005). CONCLUSIONS: Among patients with H. pylori infection and a history of upper gastrointestinal bleeding who are taking low-dose aspirin, the eradication of H. pylori is equivalent to treatment with omeprazole in preventing recurrent bleeding. Omeprazole is superior to the eradication of H. pylori in preventing recurrent bleeding in patients who are taking other NSAIDs. PMID- 11274624 TI - Images in clinical medicine. Complication of Cantor-tube insertion. PMID- 11274625 TI - Cutaneous squamous-cell carcinoma. PMID- 11274626 TI - Interactions among drugs for HIV and opportunistic infections. PMID- 11274627 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 10-2001. A 53-year-old woman with arthritis and pulmonary nodules. PMID- 11274628 TI - Immune monitoring for rejection of kidney transplants. PMID- 11274629 TI - Varicella vaccine--the first six years. PMID- 11274630 TI - Medicare and prescription drugs. PMID- 11274631 TI - Understanding Helicobacter pylori. PMID- 11274632 TI - Expression of Fas and Fas-related molecules in human hepatocellular carcinoma. AB - Many tumor cells, including hepatocellular carcinoma (HCC), express both Fas and its ligand on their surfaces, and it has remained a mystery why such cells do not spontaneously become apoptotic. In the current study, we analyzed the alterations of Fas structure and the expression of Fas and Fas ligand (FasL) and of Fas pathway inhibitors, including soluble Fas (sFas), Fas-associated phosphatase-1 (FAP-1), and bcl-2, in 50 cases of human HCC. Monoallelic loss of the Fas gene, as determined by loss of heterozygosity with intragenic polymorphisms, was observed in 5 of the 34 informative cases (15%), but none of the 50 cases showed Fas gene mutation. Expression of Fas and FasL was detected in 44 (88%) and 50 (100%) cases, respectively. sFas messenger RNA, as analyzed by in situ reverse transcription polymerase chain reaction was expressed in 42 of the 50 cases (84%), and FAP-1 expression was observed in 40 of the 50 cases (80%). In contrast, none of the 50 cases showed bcl-2 expression. Our results showed that the majority of the HCCs (88%) coexpressed a death receptor, Fas and its cognate ligand, FasL, but all HCCs showed one or more alterations of the Fas pathway molecules known to inhibit Fas-mediated apoptosis. These findings suggest that the expression of sFas and FAP-1 and, in part, loss of Fas expression, rather than Fas gene alteration or bcl-2 expression, may be involved in the Fas resistance of HCC in vivo and that these mechanisms may play important roles in the pathogenesis of human HCC. HUM PATHOL 32:250-256. PMID- 11274633 TI - Prognostic significance of histologic grade and nuclear expression of beta catenin in synovial sarcoma. AB - Synovial sarcoma, which has a wide spectrum of biologic behavior, warrants accurate grading to assess the patient's prognosis. We studied the clinicopathologic and immunohistochemical features of 44 cases of synovial sarcoma in patients treated primarily or secondarily at the National Cancer Center, Tokyo, to identify independent prognostic factors. There were local recurrences in 16 patients (36%), and 25 (57%) developed metastases, primarily to the lungs. The estimated cumulative 5-year and 10-year survival rates were 68% and 41%, respectively. Variables associated with an adverse outcome included tumor size > 6.7 cm; initial treatment outside the National Cancer Center; poorly differentiated subtype; high nuclear atypia; mitosis count > 27/10 high-power fields; tumor necrosis; absence of stromal calcification; nuclear expression of beta-catenin, which was found in 25 cases (57%); Ki-67 (MIB-1) index > 27%; and histologic grade 3. Nuclear accumulation of beta-catenin as a cell-signaling event may play an important role in the progression of synovial sarcoma and therefore might be predictive of short survival. However, multivariate analysis clearly showed that only histologic grade, as defined by using categorized variables for the MIB-1 index and tumor necrosis, was an independent prognostic factor. Most variables were correlated with lung metastasis and histologic grade. High-grade synovial sarcoma assessed by a histologic grading system based on the proliferative activity of the neoplastic cells can be viewed as high risk with the patients most likely to die of disease within 10 years after surgery and in need of improved chemotherapy. HUM PATHOL 32:257-263. PMID- 11274634 TI - Helicobacter pylori genotypes, host factors, and gastric mucosal histopathology in peptic ulcer disease. AB - From 183 patients undergoing upper gastrointestinal endoscopy, we used antral and corpus gastric biopsies for bacterial culture and histopathologic examination, blood samples to detect immunoglobulin G antibodies against Helicobacter pylori, and H pylori genomic DNA to analyze cytotoxin-associated gene A (cagA) and vacuolating cytotoxin (vacA) genotypes. As expected, among H pylori biopsy positive patients, those with duodenal ulcer (DU) (n = 34) had significantly more severe chronic and acute inflammation (P <.001) and epithelial degeneration (P =.004) in the gastric antrum than in the gastric corpus. Each of those 3 parameters and H pylori density were significantly higher in the antrum of patients with DU than in patients with gastric ulcer (GU) or no ulcer. Colonization with vacA s1/cagA-positive strains of H pylori was associated with inflammation and epithelial degeneration in gastric mucosa and increased risk for peptic ulcer disease (PUD), whereas colonization with vacA s2m2/cagA-negative strains was associated with mild gastric histopathology and was not associated with any significant risk for PUD. The predominant H pylori strains in African Americans were vacA s1bm1/cagA-positive, whereas all genotypes were well represented in non-Hispanic-Caucasians. By multivariate analysis, H pylori colonization was significantly associated with DU (Adjusted odds ratio [AdjOR] = 3.2 [1.4-7.2]) and nonsteroidal anti-inflammatory drugs (NSAID) use was inversely associated (AdjOR = 0.3 [0.2-0.7]). NSAID use (AdjOR = 4.3 [1.02-18.5]) and African-American ethnicity (AdjOR = 10.9 [2.6-50]) were significantly associated with GU. Smoking and age were not significantly associated with either DU or GU. These data indicate that DU is associated with an antral-dominant gastritis, and H pylori genotype and NSAID use independently contribute to the pathogenesis of PUD. HUM PATHOL 32:264-273. This is a US Government work. There are no restrictions on its use. PMID- 11274635 TI - Mucinous bronchioloalveolar carcinomas display a specific pattern of mucin gene expression among primary lung adenocarcinomas. AB - Lung adenocarcinomas are heterogeneous clinically and histologically. Expression of the mucin genes was analyzed as a molecular marker of glandular cytodifferentiation in primary lung adenocarcinomas. Expression was correlated with histopathologic subtypes of World Health Organization classification with the aim of investigating the histogenesis of primary lung adenocarcinomas. Thirty four primary lung adenocarcinomas were examined by in situ hybridization for mucin gene expression (MUC1-4, MUC5AC, MUC5B, MUC6-7) and by immunohistochemistry for MUC5AC and MUC5B apomucin expression. Mucinous bronchioloalveolar carcinoma (BAC) had a homogeneous pattern of mucin gene expression different from those of other types of lung adenocarcinoma, involving secreted mucins (MUC5AC, MUC5B, and MUC6) and membrane-bound mucins (MUC1, MUC3, and MUC4). Non-BAC adenocarcinoma and mucinous BAC aberrantly expressed mucin genes MUC3, and MUC3 and MUC6, respectively, which are undetectable in normal fetal and adult lung. Our results show the particular phenotype of mucin gene expression in mucinous type of BACs and the heterogeneous expression of respiratory and nonrespiratory mucins in the other types. This finding supports the theory of a common progenitor cell with the potential of multicellular differentiation. From a practical point of view, the aberrant expression of MUC3 and MUC6 could serve as a diagnostic marker in the management of the mucinous type of bronchioloalveolar carcinomas. HUM PATHOL 32:274-281. PMID- 11274636 TI - Pax-2 expression in adult renal tumors. AB - To assess the expression of the homeogene Pax-2 in adult renal cell carcinomas, we did a retrospective immunohistochemical analysis of 56 frozen tumor samples representing all major histologic subtypes of renal tumors. There were 33 conventional renal cell carcinomas (58.9%), 12 papillary renal cell carcinomas (21.4%), 4 chromophobe cell renal carcinomas, 4 urothelial cell renal carcinomas, and 3 oncocytomas. Forty-five tumors (62.5%) were localized, and 21 tumors had extrarenal involvement. Eight patients (14%) had metastatic disease at the end of the follow-up. We searched for relationships between Pax-2 expression and nuclear grading, TNM staging, Ki-67 proliferation index, expression of transforming growth factor-beta1 (TGF-beta 1), an in vitro down-regulator of Pax-2 expression, and finally cytogenetic abnormalities. All histologic subtypes expressed Pax-2 protein, except urothelial renal carcinomas. The highest expression was in papillary renal cell carcinomas. In this subtype, all tumors and 83.3% +/- 12.3% of tumor cells were immunoreactive for Pax-2. All but 2 conventional renal cell carcinomas expressed Pax-2, but with 26.3% +/- 29.6% of immunoreactive cells (P <.001). Pax-2 expression was not correlated with nuclear grading (P =.6), tumor size (P =.3), and TGF-beta 1 expression (P =.1). Nevertheless, Pax-2 expression correlated with the Ki-67 proliferation index only for the conventional histologic subtype (P =.03). In this histologic subtype, Pax-2 expression was higher in patients with metastatic disease than in those without (P =.02). Pax-2 expression was not associated with specific cytogenetic abnormalities like trisomy 7 (P =.1), 3p deletion (P =.5), and hyperdiploidy (P =.2). TGF-beta 1 expression, positive in 33 tumors (59%), was not correlated with either Pax-2 expression (P =.1) or current prognostic factors such as nuclear grading (P =.2). Interestingly, we also observed an expression of TGF-beta RI and TGF-beta RII in the tumors with high nuclear grading (P =.005). We conclude that Pax-2 protein is expressed in all major histologic subtypes of renal cell carcinomas. The pattern of expression differs between these subtypes. Pax-2 expression in conventional renal cell carcinomas is correlated with the proliferation index and is significantly higher in patients with metastatic disease. HUM PATHOL 32:282-287. PMID- 11274637 TI - Detection and characterization of human herpesvirus-8-infected cells in bone marrow biopsies of human immunodeficiency virus-positive patients. AB - We studied 15 bone marrow biopsy specimens from patients with human immunodeficiency virus infection for detection of Kaposi sarcoma herpesvirus (KSHV/HHV-8) DNA sequences by a very sensitive and specific polymerase chain reaction (PCR) assay (with 3 different sets of primers). In addition, we used immunohistochemistry with antiviral interleukin-6 (vIL-6) and anti-latent nuclear antigen-1 (LNA-1) antibodies to localize the infected cells on tissue sections. Among the 15 samples, 6 had positive PCR results with the 3 sets of primers (orf26, orf72, orf75). Interestingly, in 2 of these 6 patients (both with Kaposi sarcoma) vIL-6 and LNA-1 were detected in mononuclear lymphoid cells but not in stromal cells of the bone marrow. The detection of vIL-6--positive lymphoid cells in bone marrow suggests a homing for HHV-8--infected elements in this tissue. The local release of vIL-6 may play some role in the plasmacytosis observed in bone marrow in the acquired immunodeficiency syndrome. HUM PATHOL 32:288-291. PMID- 11274638 TI - Loss of chromosome 16q in lobular carcinoma in situ. AB - Lobular carcinoma in situ (LCIS) and infiltrating lobular carcinoma may represent different forms of the same disease based on their frequent clinical association and similar histologic features. Patients with LCIS are at increased risk of multicentric and bilateral disease. Thus, LCIS may represent both a precursor to infiltrating lobular carcinoma and a marker of risk for breast cancer. To identify genomic alterations in LCIS, comparative genomic hybridization was performed on 17 cases without concurrent invasive carcinoma. Loss involving chromosome 16q was present in 88% of cases and was the sole detected alteration in 29%. Gain involving 1q was second in frequency, occurring in 41% of tumors, and in all cases was associated with loss of 16q. Other recurrent changes were loss involving 17p (18%), 8p (12%), and 12q24 (12%). E-cadherin immunohistochemistry was performed on all LCIS cases to evaluate the correlation of loss involving 16q22, the site of the E-cadherin gene, and altered protein expression. Most cases with 16q22 loss showed altered E-cadherin expression (12 of 13). These results in LCIS are similar to changes reported in infiltrating lobular cancer, confirming a genetic relationship between them. HUM PATHOL 32:292 296. PMID- 11274639 TI - In situ hybridization of Epstein-Barr virus in tumor cells and tumor-infiltrating lymphocytes of the gastrointestinal tract. AB - Unlike gastric carcinoma, associations of the Epstein-Barr virus (EBV) with carcinomas of other sites in the gastrointestinal tract have not yet been clarified. To elucidate these associations, we investigated the presence of EBV in 142 cases of esophageal carcinoma, 107 cases of ampulla of Vater carcinoma, and 274 cases of colorectal carcinoma in Korean patients using EBV-encoded small RNAs (EBER)-in situ hybridization (ISH). In all cases, none of the tumor cells showed a positive signal, indicating that EBV is not generally related to the carcinogenesis of these cancers. Some EBV-positive tumor-infiltrating lymphocytes (TILs) were found in 8 of 142 cases (5.6%) of esophageal carcinoma, 8 of 107 cases (7.5%) of ampulla of Vater cancer, and 35 of 274 cases (12.8%) of colorectal carcinoma. For comparison, EBER-ISH was performed in consecutive gastric carcinomas; the EBER signal on tumor cells was observed in 17 of 306 cases (5.6%), and EBV-positive TILs were seen in 31 of the 289 cases (10.7%). There was no statistically significant difference in the frequencies of cases with EBV-positive TILs among the gastrointestinal tract cancers. We suggest that the reservoir lymphocytes carrying EBV, like other inflammatory cells, are able to reach anywhere, and that the chance for an epithelial cell to be exposed to EBV is similar at different sites of the gastrointestinal tract, regardless of its carcinogenic effect on the epithelial cell. HUM PATHOL 32:297-301. PMID- 11274640 TI - Expression of endogenous galectin-1 and galectin-3 in intrahepatic cholangiocarcinoma. AB - Galectins, a family of beta-galactoside-binding animal lectins, might be involved in tumor progression. In this study, the expression patterns of galectin-1 and -3 were examined immunohistochemically in intrahepatic cholangiocarcinoma (ICC), with emphasis on its development and progression as well as its histopathologic features, by use of samples of normal intrahepatic bile duct (n = 20), biliary epithelial dysplasia (n = 15), ICC (n = 40), and a cholangiocarcinoma cell line, CCKS1. In normal intrahepatic bile ducts, galectin-3 was constitutively but weakly expressed, whereas galectin-1 was not expressed. In hepatolithiasis, biliary epithelial dysplasia was strongly positive for galectin-3 but negative for galectin-1. Galectin-3 was frequently and strongly expressed in the cytoplasm of well-differentiated ICCs, and its expression was significantly decreased and less intense or even absent in poorly differentiated ICCs. Galectin-1 was expressed in carcinoma cells in ICC, and its incidence and extent were correlated with histologic dedifferentiation of ICC. Proliferative cell nuclear antigen (PCNA) labeling index (LI) was higher in ICC cases positive for galectin-1 than in those that were negative. Galectin-1 was strongly expressed in cancerous stroma of ICC, and this stromal expression was related to histologic dedifferentiation of ICC. In the carcinoma cell line CCKS1, galectin-1 and -3 were expressed in the cytoplasm of carcinoma cells, and galectin-1 was additionally detected in the culture medium. These results suggest that galectin 1 was newly expressed on carcinoma cells of ICC, and its overexpression seems to be associated with neoplastic progression and proliferative activities, and the expression of galectin-1 in cancerous stroma may also be related to the progression of ICC. Galectin-3 expression in epithelial cells is up-regulated in the preneoplastic and early neoplastic stages of ICC, although galectin-3 tends to disappear at later stages of ICC. HUM PATHOL 32:302-310. PMID- 11274641 TI - Concurrent overexpression of p53 and c-erbB-2 correlates with accelerated cycling and concomitant poor prognosis in node-negative breast cancer. AB - Simultaneous overexpression of c-erbB-2 and p53 has been reported to be prognostically unfavorable in breast cancer. Herein, we show that concurrent overexpression of these 2 proteins is associated with a marked reduction in the relative fraction of cells in G(1) phase of the cell cycle, indicating an accelerated cell cycle progression. Using an immunohistochemical approach, we examined 261 cases of node-negative infiltrating ductal carcinomas of the breast with respect to c-erbB-2 and p53 expression and to the proliferative activity measured by the Ki-67 index. By means of a novel monoclonal antibody, Ki-S2, which exclusively recognizes proliferating cells in the S, G(2), and M phases of the reproductive cycle, we were further able to calculate the relative fraction of the cells having passed the restriction point at the G(1)/S boundary, thus defining a cycling ratio (CR). The results were correlated with clinical outcome; median follow-up time was 96 months. Tumors that simultaneously overexpressed c erbB-2 and p53 had a high median CR and followed an unfavorable course. However, increased CRs were also observed independently of c-erbB-2 and p53 overexpression, suggesting that other molecular mechanisms may contribute to acceleration of cell cycle progression. In a multivariate analysis that included patient age, tumor size, hormone receptor status, c-erbB-2 and p53 expression, and the Ki-67 index, CR emerged as the most significant independent predictor of overall and disease-free survival (P <.0001). It is concluded that the CR is a gauge of cell cycle deregulation and therefore may be a powerful indicator of the biologic behavior of cancers. HUM PATHOL 32:311-319. PMID- 11274643 TI - Replicative senescence in normal liver, chronic hepatitis C, and hepatocellular carcinomas. AB - There is growing evidence that senescent cells accumulate in vivo and are associated with the aging process in parallel with the progressive erosion of telomeres. Because recent data show that telomere shortening is involved in the pathogenesis of liver cirrhosis, we looked for replicative senescence cells in normal livers, chronic hepatitis C, and hepatocellular carcinoma (HCC). Replicative senescent cells were detected on liver tissue cryosections using expression of a specific marker, senescence-associated beta-galactosidase, a cytoplasmic enzyme detected at pH 6. A total of 57 frozen liver samples (15 normal liver, 32 chronic hepatitis C, and 10 HCCs) were studied. Replicative senescence was graded as absent in 56% of cases (32 of 57) and present in 44% (25 of 57). Replicative senescence was considered present in 3 of 15 normal livers (20%), 16 of 32 chronic hepatitis cases (50%), and 6 of 10 HCCs (60%). In the group of nontumoral livers, the presence of senescent cells in liver was associated with older age (P =.03). In the group with chronic hepatitis C, fibrosis stage, but not activity grade, was significantly correlated with the accumulation of replicative senescent cells (P <.001). Finally, beta-Gal staining in nontumoral tissue was strongly correlated with the presence of HCC in the surrounding liver (P <.001). These results suggest that chronic hepatitis C represents a relevant model of accelerated replicative senescence and that accumulation of replicative senescent cells predispose to HCC development. Detection of replicative senescent cells may then serve as a predictive marker of a hepatocellular carcinoma in the surrounding tissue. HUM PATHOL 32:327-332. PMID- 11274644 TI - Loss of heterozygosity on chromosome arm 11q in lung carcinoids. AB - Neuroendocrine lung tumors such as typical carcinoid, atypical carcinoid, small cell lung carcinoma, and large-cell neuroendocrine carcinoma represent a variable group with different biologic characteristics and unclear genetical relationships. We investigated the pattern of allelic loss on chromosome arm 11q in 20 sporadic carcinoid tumors of the lung using 10 microsatellite markers. Loss of heterozygosity was found in 13 of 20 tumors. In 5 of 9 typical carcinoids, 3 distinct regions of allelic loss were identified: 11q13.1 (D11S1883), 11q14.3 11q21 (D11S906), and 11q25 (D11S910). Atypical carcinoids showed loss of heterozygosity at 4 different regions: the first, most proximal region at 11q13 between markers PYGM and D11S937; the second at 11q14.3-11q21 (D11S906); and the third and fourth defined by markers D11S939 (11q23.2-23.3) and D11S910 (11q25). However, the region 11q13 harboring the MEN1 gene was more frequently affected in atypical carcinoids (7 of 11) than in typical carcinoids (2 of 9). The high rate of allelic losses within chromosomal region 11q13 in atypical carcinoids emphasizes the importance of this region for tumor development. We also recognized that more aggressive atypical carcinoids defined by high mitotic counts, vascular invasion, and/or organ metastasis are combined with increased allelic losses. HUM PATHOL 32:333-338. PMID- 11274642 TI - Expression of the chemokine RANTES in pulmonary Wegener's granulomatosis. AB - Wegener's granulomatosis (WG) is an inflammatory, destructive, angiotropic lesion. The inflammatory process involves accumulation of macrophages, lymphocytes, and polymorphonuclear neutrophils. We studied 6 lung biopsy specimens from patients with WG to characterize the cellular infiltrate and to analyze the mechanism of immune cell recruitment. We show that lymphocytes accumulating in WG lesions are mostly memory CD4(+)CD45RO(+) T lymphocytes and, although less numerous, CD8(+)CD45RO(+) T lymphocytes. Few if any B lymphocytes or natural killer cells are present within lesions. The chemokine RANTES (regulated upon activation in normal T cells, expressed and secreted) has been reported to recruit memory T lymphocytes and macrophages selectively. We used reverse-transcription polymerase chain reaction, in situ hybridization, and immunohistochemistry to study its production in WG. RANTES was expressed at a higher level in WG lungs than in normal controls, especially around microabscesses. As visualized immunohistochemically in serial sections with anti RANTES monoclonal antibody, RANTES production was produced mainly by macrophages. Expression of the gene coding for interferon-gamma (IFN-gamma), a potent RANTES inducer, was also studied. Its expression was also much stronger in WG than in controls. Our observations are consistent with a cascade of events leading to the recruitment of immune cells in WG, sequentially involving production of IFN-gamma by T lymphocytes and RANTES production by macrophages, leading to the homing of memory T-helper lymphocytes and macrophages. HUM PATHOL 32:320-326. PMID- 11274645 TI - An angiocentric orbital lesion with an immature natural killer cell immunophenotype. AB - In this report, a unique case of a large unilateral orbital tumor occurring in a 5-year-old white girl is described. The lesion, which was associated with no systemic clinical symptoms, waxed and waned in size over 12 months and eventually spontaneously resolved. Multiple biopsies were performed, which showed an angiocentric and angioinvasive infiltrate composed of a monotonous population of atypical, immature-appearing, large lymphocytes. Extensive molecular and immunophenotypic studies led to the diagnosis of an Epstein-Barr virus-negative angiocentric lymphoproliferative disorder with an immature natural killer (NK) cell immunophenotype (CD16+CD56-), specifically an immunophenotype expressed by normal cord blood NK cells. HUM PATHOL 32:339-342. PMID- 11274646 TI - Fatal parvovirus myocarditis in a 5-year-old girl. AB - Infection with parvovirus B19 is common in children and typically causes mild illness. We report here the case of a 5-year-old girl who died suddenly, 2 weeks after the clinical diagnosis of a parvoviral infection (erythema infectiosum). Microscopic examination of the heart showed severe myocarditis with extensive T cell and macrophage infiltration. Cultures, serology, and molecular analyses of serum for enteroviridae, adenovirus, influenza, varicella zoster, cytomegalovirus, and herpes simplex viruses were negative. Quantitative polymerase chain reaction (PCR) analysis for parvovirus B19 in peripheral blood, however, showed active infection (91,000 genomes/mL serum; 2.4 genomes/mononuclear cell). Despite the presence of myocarditis, immunostaining for parvoviral surface antigens was negative in the heart. Quantitative PCR analysis of paraffin sections showed that myocardial parvoviral content was significantly less than that of the normal appearing kidney and within the range predicted simply by tissue blood content. Thus, parvovirus B19 infection can be complicated by fatal myocarditis. Because the virus does not appear to have infected the heart, per se, we speculate that myocarditis arose from immunological cross-reaction to epitopes shared between the virus and the myocardium. HUM PATHOL 32:342-345. PMID- 11274647 TI - Subcutaneous nodular amyloidosis: a case report and review of the literature. AB - Amyloidosis typically manifests with disseminated infiltration of multiple organ systems. Rarely, amyloidosis may be localized. We report a patient with localized subcutaneous nodular amyloidosis, without systemic amyloid involvement or myeloma, whose presenting symptom was multiple discrete neck nodules. Immunohistochemical analysis showed the amyloid deposits to be derived from lambda light chains. Twenty-four month follow-up showed minimal disease progression. A literature review showed only 5 reported cases of subcutaneous nodular amyloidosis. This is the first description of a patient with subcutaneous nodular amyloidosis derived from lambda light chains. HUM PATHOL 32:346-348. PMID- 11274648 TI - Breast carcinomas with immunocytochemical detection of aromatase in fine-needle aspirates: report of three cases. AB - Three postmenopausal women with breast carcinoma underwent the fine-needle aspiration (FNA) preoperatively, and these specimens were stained by the antiaromatase antibody. We evaluated the identification of the aromatase immunoreactivity in breast carcinoma specimens obtained from both FNA and surgery. FNA specimens showed positive intracellular immunoreactivity of aromatase in these cases. The presence for aromatase in FNA specimens was identified with that in the surgical specimens. To our knowledge, the present cases are the first to report the aromatase staining of FNA specimens. The immunoreactivity of aromatase in FNA specimen may be useful to estimate the effectiveness of new aromatase inhibitors in patients with breast carcinoma. HUM PATHOL 32:348-351. PMID- 11274649 TI - Warthin's tumors. PMID- 11274650 TI - Glucocorticoid receptors in major depression: relevance to pathophysiology and treatment. AB - Hyperactivity of the hypothalamic--pituitary--adrenal (HPA) axis has been reliably observed in patients with major depression. One of the primary features of this HPA axis hyperactivity is reduced sensitivity to the inhibitory effects of the glucocorticoid dexamethasone on the production of adrenocorticotropic hormone and cortisol during the dexamethasone suppression test and, more recently, the dexamethasone--corticotropin-releasing hormone test. Because the effects of glucocorticoids are mediated by intracellular receptors including, most notably, the glucocorticoid receptor (GR), a number of studies have considered the possibility that the number and/or function of GRs are reduced in depressed patients. Moreover, whether antidepressants act by reversing these putative GR changes has been examined. The extant literature on GR receptors in major depression was reviewed along with studies examining the impact of antidepressants on the GR. The data support the hypothesis that the function of the GR is reduced in major depression in the absence of clear evidence of decreased GR expression. The data also indicate that some antidepressants have direct effects on the GR, leading to enhanced GR function and increased GR expression. Hypotheses regarding the mechanism of these receptor changes involve relevant second messenger pathways that regulate GR function. The findings indicate that the GR is an important molecular target in major depression. Further elucidation of the biochemical and molecular mechanisms involved in GR changes in major depression is an exciting frontier that will no doubt lead to new insights into the pathophysiology and treatment of affective disorders. PMID- 11274652 TI - High and low neuroticism predict different cortisol responses to the combined dexamethasone--CRH test. AB - BACKGROUND: Depression and posttraumatic stress disorder are both associated with altered function of the hypothalamic--pituitary--adrenal axis. Neuroticism is a strong predisposing factor for depression and probably also a risk factor for posttraumatic stress disorder. This study investigated whether young adults with high and low neuroticism scores show differences in hypothalamic-pituitary adrenal axis regulation that might relate to their differential vulnerability to psychopathology. METHODS: Neuroticism was measured with the Eysenck Personality Questionnaire in 258 students aged 18--25. Fourteen scoring in each of the upper and lower quartiles of the neuroticism distribution according to gender participated in a combined dexamethasone-corticotropin-releasing hormone test. RESULTS: Low-neuroticism individuals showed a significantly greater cortisol response than high-neuroticism individuals. CONCLUSIONS: The mechanism of this effect remains to be elucidated. High-neuroticism subjects may have a downregulated hypothalamic--pituitary--adrenal axis to prevent harmful overactivation. This is the first demonstration of a difference in hypothalamic- pituitary--adrenal axis regulation associated with neuroticism. PMID- 11274651 TI - Tryptophan hydroxylase polymorphism and suicidality in unipolar and bipolar affective disorders: a multicenter association study. AB - BACKGROUND: Being the rate-limiting enzyme in the biosynthesis of serotonin, the tryptophan hydroxylase gene (TPH) has been considered a possible candidate gene in bipolar and unipolar affective disorders (BPAD and UPAD). Several studies have investigated the possible role of TPH polymorphisms in affective disorders and suicidal behavior. METHODS: The TPH A218C polymorphism has been investigated in 927 patients (527 BPAD and 400 UPAD) and their matched healthy control subjects collected within the European Collaborative Project on Affective Disorders. RESULTS: No difference of genotype distribution or allele distribution was found in BPAD or UPAD. No statistically significant difference was observed for allele frequency and genotypes counts. In a genotype per genotype analysis in UPAD patients with a personal history of suicide attempt, the frequency of the C-C genotype (homozygosity for the short allele) was lower in UPAD patients (24%) than in control subjects (43%) (chi(2) = 4.67, p =.03). There was no difference in allele or genotype frequency between patients presenting violent suicidal behavior (n = 48) and their matched control subjects. CONCLUSIONS: We failed to detect an association between the A218C polymorphism of the TPH gene and BPAD and UPAD in a large European sample. Homozygosity for the short allele is significantly less frequent in a subgroup of UPAD patients with a history of suicide attempt than in control subjects. PMID- 11274653 TI - Electroencephalographic and perceptual asymmetry differences between responders and nonresponders to an SSRI antidepressant. AB - BACKGROUND: Recent reports suggest the value of electroencephalographic and dichotic listening measures as predictors of response to antidepressants. This study examines the potential of electroencephalographic alpha asymmetry and dichotic measures of perceptual asymmetry as predictors of clinical response to 12 weeks of treatment with fluoxetine (Prozac). METHODS: Resting electroencephalography (eyes open and eyes closed) and dichotic listening with word or complex tone stimuli were assessed in depressed outpatients during a pretreatment period. RESULTS: Fluoxetine responders (n = 34) differed from nonresponders (n = 19) in favoring left over right hemisphere processing of dichotic stimuli. They also differed in their resting electroencephalographic alpha asymmetry, particularly in the eyes open condition. Nonresponders showed an alpha asymmetry indicative of overall greater activation of the right hemisphere than the left, whereas responders did not. The relationship between hemispheric asymmetry and treatment response interacted with gender, being evident among depressed women but not men. CONCLUSIONS: The results are consistent with the hypothesis that a characteristic tendency toward greater left than right hemisphere activation is associated with favorable response to fluoxetine, whereas the opposite hemispheric asymmetry predicts poor response. PMID- 11274654 TI - Patterns of cortical activity and memory performance in Alzheimer's disease. AB - BACKGROUND: Declarative memory changes are the hallmark of Alzheimer's disease, although their functional neuroanatomy is not restricted to a single structure. Factor analysis provides statistical methods for evaluating patterns of cerebral changes in regional glucose uptake. METHODS: Thirty-three Alzheimer's patients and 33 age- and gender-matched control subjects were studied with magnetic resonance imaging and positron emission tomography with [(18)F] deoxyglucose. During the tracer-uptake period, subjects performed a serial verbal learning task. Cortical activity was measured in 32 regions of interest, four in each lobe on both hemispheres. RESULTS: Factor analysis with varimax rotation identified seven factors explaining 80% of the variance ("parietal cortex," "occipital cortex," "right temporo-prefrontal areas," "frontal cortex," "motor strip," "left temporal cortex," and "posterior temporal cortex"). Relative to control subjects, Alzheimer's patients showed significantly reduced values on the factors occipital cortex, right temporo-prefrontal areas, frontal cortex, and left temporal cortex. The factor temporo-prefrontal areas showed large differences between patients with good and poor performance, but little difference when control subjects were similarly divided. CONCLUSIONS: Findings suggest that Alzheimer's disease is characterized by altered patterns of cortical activity, rather than deficits in a single location, and emphasize the importance of right temporo-prefrontal circuitry for understanding memory deficits. PMID- 11274656 TI - Chronic myo-inositol increases rat brain phosphatidylethanolamine plasmalogen. AB - BACKGROUND: Oral myo-inositol (12--18 g/day) has shown beneficial effect in placebo-controlled studies of major depression, panic disorder, and obsessive compulsive disorder, and preliminary data suggest it also may be effective in bipolar depression. Evidence linking antidepressant activity to membrane phospholipid alterations suggested the examination of acute and chronic myo inositol effects on rat brain membrane phospholipid metabolism. METHODS: With both (31)P nuclear magnetic resonance (NMR) and quantitative high-performance thin-layer chromatography (HPTLC; hydrolysis) methods, rat brain phospholipid levels were measured after acute (n = 20, each group) and chronic myo-inositol administration (n = 10, each group). With (31)P NMR, we measured myo-inositol rat brain levels after acute and chronic myo-inositol administration. RESULTS: Brain myo-inositol increased by 17% after acute myo-inositol administration and by 5% after chronic administration, as compared with the control groups. Chronic myo inositol administration increased brain phosphatidylethanolamine (PtdEtn) plasmalogen by 10% and decreased brain PtdEtn by 5%, thus increasing the ratio PtdEtn plasmalogen (PtdEtn-Plas)/PtdEtn by 15%. Phosphatidylethanolamine plasmalogen levels quantified by (31)P NMR and HPTLC were highly correlated. The validity and reliability of the (31)P NMR method for phospholipid analysis were demonstrated with phospholipid standards. CONCLUSIONS: The observed alteration in the PtdEtn-Plas/PtdEtn ratio could provide insights into the therapeutic effect of myo-inositol in affective disorders. PMID- 11274655 TI - Brain lithium concentrations in bipolar disorder patients: preliminary (7)Li magnetic resonance studies at 3 T. AB - BACKGROUND: This study was conducted to investigate the feasibility of human brain (7)Li MRS investigations at a high magnetic field (3 T), and to further explore the relationship between brain and serum lithium measures in lithium treated bipolar patients. METHODS: Eight bipolar disorder type I patients (5 males, 3 females; mean age +/- SD = 33 +/- 9 years) were studied. A 3-T scanner, using a dual-tuned ((1)H and (7)Li) echoplanar imaging (EPI) compatible radiofrequency (RF) birdcage coil was used. (7)Li magnetic resonance spectroscopy (MRS) signal was acquired at the frequency of 49.64 MHz using an imaging selective in vivo spectroscopy (ISIS) sequence (TR = 15 sec, 128 averages), and quantitation was obtained in reference to an external standard. RESULTS: The mean +/- SD oral lithium dose was 1265 +/- 442 mg/day, and the mean +/- SD 12-hour serum level was 0.69 +/- 0.19 mEq/L. The measured brain lithium concentrations varied from 0.23 to 0.55 mEq/L (mean +/- SD = 0.35 +/- 0.11 mEq/L). The brain serum ratios varied from 0.30 to 0.80 (mean +/- SD = 0.52 +/- 0.16). Subjects on single daily doses of lithium at bedtime (n = 5) had higher brain-serum lithium ratios compared with those on twice-a-day schedules (n = 3) (0.61 +/- 0.12 and 0.37 +/- 0.07, respectively; Mann--Whitney U test, Z = -2.24, p =.03). CONCLUSIONS: This study demonstrated for the first time the feasibility of (7)Li MRS human studies at 3 T. Future studies should examine a possible role for this methodology in investigations of lithium refractoriness and prediction of treatment outcome in bipolar patients. PMID- 11274657 TI - The transcranial magnetic stimulation motor threshold depends on the distance from coil to underlying cortex: a replication in healthy adults comparing two methods of assessing the distance to cortex. AB - Using transcranial magnetic stimulation (TMS), a handheld electrified copper coil against the scalp produces a powerful and rapidly oscillating magnetic field, which in turn induces electrical currents in the brain. The amount of electrical energy needed for TMS to induce motor movement (called the motor threshold [MT]), varies widely across individuals. The intensity of TMS is dosed relative to the MT. Kozel et al observed in a depressed cohort that MT increases as a function of distance from coil to cortex. This article examines this relationship in a healthy cohort and compares the two methods of assessing distance to cortex. Seventeen healthy adults had their TMS MT determined and marked with a fiducial. Magnetic resonance images showed the fiducials marking motor cortex, allowing researchers to measure distance from scalp to motor and prefontal cortex using two methods: 1) measuring a line from scalp to the nearest cortex and 2) sampling the distance from scalp to cortex of two 18-mm-square areas. Confirming Kozel's previous finding, we observe that motor threshold increases as distance to motor cortex increased for both methods of measuring distance and that no significant correlation exists between MT and prefontal cortex distance. Distance from TMS coil to motor cortex is an important determinant of MT in healthy and depressed adults. Distance to prefontal cortex is not correlated with MT, raising questions about the common practice of dosing prefontal stimulation using MT determined over motor cortex. PMID- 11274658 TI - Sham TMS: intracerebral measurement of the induced electrical field and the induction of motor-evoked potentials. AB - Testing the therapeutic potential of transcranial magnetic stimulation (TMS) in controlled trials requires a valid sham condition. Sham TMS is typically administered by tilting the coil 45--90 degrees off the scalp, with one or two wings of the coil touching the scalp. Lack of cortical effects has not been verified. We compared sham manipulations in their thresholds for eliciting motor evoked potentials (MEPs) in human volunteers and in intracerebral measurements of voltage induced in the prefrontal cortex of a rhesus monkey. Three types of sham (one-wing 45 degrees and 90 degrees and two-wing 90 degrees tilt) induced much lower voltage in the brain than active TMS (67--73% reductions). However, the two wing 45 degrees sham induced values just 24% below active TMS. This sham was about half as potent in inducing MEPs over the motor cortex as active TMS. Some sham TMS conditions produce substantial cortical stimulation, making it critical to carefully select the sham manipulation for clinical trials. PMID- 11274659 TI - Stimulation of reactive oxygen species production by an antidepressant visible light source. AB - BACKGROUND: The mechanism by which visible light stimulates chronobiological phase-shifting or antidepressant effects in humans is unknown. METHODS: Normal human NIH/3T3 nonpigmented fibroblasts were irradiated with a visible light source (SunRay) used in the treatment of winter seasonal depression. Electron spin resonance was assessed before and after 10 min of illumination at 2 mW/cm(2) (illuminance of 3700 lux), with and without the presence of 5 microL of 0.0214 mg/mL vitamin C. RESULTS: The fibroblasts showed evidence of production of reactive oxygen species after 10 min of irradiation. CONCLUSIONS: These in vitro data establish that an antidepressant source of visible light is capable of inducing the production of reactive oxygen species in skin. Such species may participate in signal transduction pathways leading to mood changes. PMID- 11274660 TI - Transcranial magnetic stimulation-induced switch into mania: a report of two cases. AB - BACKGROUND: Transcranial magnetic stimulation is a novel, experimental procedure in the treatment of psychiatric disorders, most notably mood disorders. Transcranial magnetic stimulation is currently being widely studied in other applications, and its efficacies and potential side effects are being investigated. METHODS: Transcranial magnetic stimulation was administered five times a week for 4 weeks. RESULTS: In this report, a manic episode followed treatment with transcranial magnetic stimulation in two patients. CONCLUSIONS: Clinicians should be aware that, like with other antidepressive treatments, a switch into mania might complicate treatment with transcranial magnetic stimulation in bipolar patients. PMID- 11274661 TI - Job control, perceptions of control, and cardiovascular activity: an analysis of ambulatory measures collected over the working day. AB - OBJECTIVE: To assess the influence of job control as a personal characteristic, and momentary perceptions of lack of control, on blood pressure and heart rate monitored over the working day. METHODS: The study was carried out with a sample of 122 school teachers (45 men, 77 women), divided into high and low job control groups on a standard questionnaire. Blood pressure and heart rate were measured every 20 min using ambulatory techniques, and ratings of concurrent perceived control were also obtained. Energy expenditure was assessed using accelerometers. Cardiovascular measures associated with low perceived control were averaged for each individual, as were readings associated with high perceived control. RESULTS: Low perceived control ratings were infrequent overall. However, in the participants who reported episodes of both high and low perceived control, systolic and diastolic blood pressure and heart rate were significantly greater during episodes of low control. These effects were independent of concurrent energy expenditure, gender, and time of day. High and low job control groups did not differ in cardiovascular responses to episodes of low perceived control. But low job control was associated with more frequent episodes of low perceived control and fewer periods of high perceived control. CONCLUSION: The results suggest that laboratory observations concerning the impact of lack of control on cardiovascular activity are corroborated by naturalistic measures in everyday life. Differences in exposure to periods of low perceived control rather than differences in reactivity to uncontrollable situations may contribute to the elevation in cardiovascular risk associated with low job control. PMID- 11274662 TI - Illness perceptions and mood in chronic fatigue syndrome. AB - BACKGROUND: Individual beliefs and cognitions may affect adjustment to chronic fatigue syndrome (CFS) and illness perceptions, in particular, have been reported to correlate with both disability and psychological adjustment to CFS in self diagnosed cases. OBJECTIVES: The aim of the present study was to examine these relationships in a clinic sample of CFS patients assessed by both a physician and psychiatrist. METHOD: A sample of 173 patients referred to a multidisciplinary CFS clinic and fulfilling current operational criteria for CFS [Ann Intern Med 121 (1994) 953; J R Soc Med 84 (1991) 118.] were randomly selected from the clinic database and surveyed with the Hospital Anxiety and Depression scale, Fatigue Questionnaire and Illness Perceptions Questionnaire [J Psychosom Res 37 (1993) 147; Psychol Health 11 (1996) 431; Acta Psychiatr Scand 67 (1983) 361.]. RESULTS: A total of 126 patients responded (73% response rate). The illness perception components studied were consequences (of illness), illness identity, causes (of illness), the ability to control/cure the illness and (expected) timeline of the illness. These components accounted for 15%, 28% and 30% of the variance in levels of fatigue, depression and anxiety, respectively. Two of the illness perception components (consequences and illness identity) were stronger predictors of fatigue score than mood scores. CONCLUSIONS: These findings confirmed in a clinical sample that illness perceptions are associated with variation in both disability and psychological adjustment in CFS. Illness perceptions may have an important and long-lasting effect on adaptation to CFS, and it is necessary to have a greater understanding of their role in order to tailor effective interventions for the condition. PMID- 11274663 TI - Defending against patients' pain: a qualitative analysis of nurses' responses to children's postoperative pain. AB - OBJECTIVE: Cognitive approaches to clinical communication attribute deficits in communication to lack of skill. We examined, instead, emotional influences on communication by finding out how nurses construed patients who were in pain, and how these constructions were related to the emotional challenge of patients' pain and to deficits in clinical communication. METHODS: Data, analyzed qualitatively, included: (i) direct observations of verbal interactions of 13 nurses with 16 children after orthopedic surgery; (ii) standardized open-ended interviews with the nurses, patients and parents. RESULTS: Nurses tried to prevent children from displaying pain behavior. When pain behavior did occur, they construed pain as unreal, unwarranted or not deserving help. These findings were apparent in observations of, and interviews with, nurses and also in parents' and children's accounts of nurses' behavior. CONCLUSION: We identified behavioral and cognitive strategies whereby clinicians defend themselves emotionally against patients' pain, and which compromise communication with patients in pain. PMID- 11274664 TI - Sense of coherence as a predictor of subjective state of health: results of 4 years of follow-up of adults. AB - A number of cross-sectional population studies have shown that a strong sense of coherence (SOC) is associated with various aspects of good perceived health. The association does not seem to be entirely attributable to underlying associations of SOC with other variables, such as age or level of education. OBJECTIVE: The aim of the study reported here was to determine whether SOC predicted subjective state of health. METHODS: The study was carried out as a two-way panel mail survey of 1976 individuals with 4 years interval for two collections of data. The statistical method used was multivariate cumulative logistic modeling. Age, initial subjective state of health, initial occupational training level, and initial degree of social integration were included as potential explanatory variables. RESULTS: A strong SOC predicted good health in women and men. CONCLUSIONS: SOC can be interpreted as an autonomous internal resource contributing to a favorable development of subjective state of health. SOC data should, however, be regarded as complementary to and not a substitute for information already known to be associated with increased risk of future ill health. PMID- 11274665 TI - Deliberate self-harm patients who leave the accident and emergency department without a psychiatric assessment: a neglected population at risk of suicide. AB - OBJECTIVES: Deliberate self-harm (DSH) patients, despite their risk of suicide, are often discharged directly from accident and emergency (A&E) departments without undergoing a psychiatric assessment. The aims of this study were to determine the characteristics and outcome of these patients. METHODS: The characteristics of DSH patients who were discharged directly from an A&E department over a 2-year period were investigated, comparing those who had a psychiatric assessment with those who did not. In a matched control design, the outcome of a group of patients who did not receive a psychiatric assessment was compared with that of a group of patients who were assessed. RESULTS: Of DSH patients who were discharged directly from the A&E department 58.9% (145/246) did not have a psychiatric assessment. Nonassessed patients were more likely to have a past history of DSH, to be in the 20-34 year age group, and to have exhibited difficult behaviour in the A&E department. Patients presenting between 5 p.m. and 9 a.m. were less likely to be assessed than those attending between 9 a.m. and 5 p.m. Further DSH during the subsequent year occurred in 37.5% of the nonassessed patients compared with 18.2% of matched assessed patients. They were also more likely to have psychiatric treatment. CONCLUSION: A substantial proportion of DSH patients discharged directly from A&E departments do not receive a psychiatric assessment. Nonassessed patients may be at greater risk of further DSH and completed suicide than those who are assessed. Hospital services need to be organised such that DSH patients managed in A&E departments can receive an assessment of psychosocial problems and risk. PMID- 11274666 TI - Psychosocial patient characteristics and GP-registered chronic morbidity: a prospective study. AB - OBJECTIVE: The aim of this study was to get a profile of patients who are vulnerable to get multiple chronic, recurrent or high-impact diseases in a limited time period. We studied the incidence rates of morbidity and multimorbidity, and the influence of psychosocial characteristics on their occurrences. METHOD: Cohort study with 3551 subjects. Baseline measurement of psychosocial characteristics and a 2-year follow-up period for morbidity. The relations were evaluated using multiple logistic regression analysis. RESULTS: After adjustment for basic socio-demographic variables, a high internal locus of control belief was found to be protective [odds ratio (OR)=0.82] for the occurrence of morbidity, negative life events increased the risk (OR=1.22). Characteristics specifically protective for the occurrence of multimorbidity as compared to monomorbidity were: a high internal locus of control belief (OR=0.73), living as a couple or in a family as compared to living alone (OR=0.68) and a large social network (OR=0.41). CONCLUSION: It appears that certain patient characteristics are specifically related to the occurrence of multimorbidity. This provides opportunities for the future development of preventive interventions. PMID- 11274667 TI - Psychiatric morbidity in patients undergoing heart, heart and lung, or lung transplantation. AB - OBJECTIVES: To determine the rate of psychiatric disorder in people undergoing heart and/or lung transplantation; to identify the associations of psychiatric disorder in this group. METHOD: Preoperative assessments were carried out on an 18-month sample of consecutive admissions to a regional unit for heart and lung transplantation in the UK. Assessment included psychiatric morbidity, sexual dysfunction, quality of life, and demographic and clinical characteristics. RESULTS: Seventy-six of 79 eligible subjects took part in the assessment. Thirty (39%) were suffering from a psychiatric disorder, the most common being major depressive disorder. Forty-four (58%) reported sexual dysfunction. Clinically significant psychiatric morbidity was associated with a history of treatment for mental disorder, unemployment, and length of physical illness. Patients with psychiatric disorder reported poorer quality of life on the SF-36, with lower scores on subscales for general health perception, social functioning, and energy/vitality. CONCLUSION: There is a substantial rate of psychiatric disorder in people undergoing heart and/or lung transplantation. Risk is higher in people with a history of psychiatric vulnerability and current illness-related factors. Preoperative psychiatric assessment and intervention in some patients may be a valuable part of their clinical care. PMID- 11274668 TI - A survey of frequent attenders at a gastroenterology clinic. AB - OBJECTIVE: To examine a group of patients satisfying criteria for "frequent attending" as part of an audit of an outpatient gastroenterology service, and to note the prevalence of those with no conspicuous organic disease to account for their symptomatology. METHODS: We used the hospital computer (Oxford Patient Administration System, OXPAS) to identify 2530 consecutive patients who were given an appointment to attend the gastroenterology clinic during an 11-month period. Patients designated "frequent attenders" had their notes flagged before the clinic attendance and were examined in more detail. A frequent attender was defined as a patient who had attended any hospital outpatient clinic in the three Oxford general hospitals on four or more occasions in the previous 12 months. The gastroenterologist then interviewed the patients satisfying these criteria and indicated whether he/she was satisfied that there was no relevant organic disease to account for the symptoms. RESULTS: Of the total 2530 patients, 762 (30%) satisfied our criteria for frequent attendance (FA). Of these, 452 (59%) had organic disease, 128 (17%) either did not attend or cancelled and 159 (21%) had no relevant organic disease. The diagnosis was uncertain in 23 patients (3%). Of patients satisfying our criteria for frequent attending, approximately 20-25% had no established gastroenterological disease. CONCLUSIONS: Frequent attenders present formidable management problems for the gastroenterologist. If they can be identified by computer before the outpatient visit then assessment and management might be more appropriately supervised in designated clinics by more experienced gastroenterology staff. PMID- 11274669 TI - Acute anosmia in the mouse: behavioral discrimination among the four basic taste substances. AB - The importance of taste and smell in discrimination of tastes was examined in normal and anosmic mice. We studied the influence of olfaction on taste sensation using behavioral and electrophysiological methods in both normal animals and animals made anosmic mice by destroying their olfactory receptor cells with zinc sulfate (ZnSO(4)) solution. Electrophysiological responses from chorda tympani nerves showed that peripheral taste receptor cells transmitted taste signals normally to the central nervous system, even when the olfactory senses were abnormal. Behavioral observations showed that mice with abnormal olfaction could not differentiate tastes. PMID- 11274670 TI - Basal midbrain modulation of tonic immobility in the toad Bufo paracnemis. AB - Tonic immobility (TI) is considered to be a final stage in a sequence of defensive responses occurring in the prey/predator encounter. It is known that the basal midbrain of toads is involved in the organization of defensive behavior and analgesia. This study investigated the effect of electrolytic or neurotoxic lesions of two mesencephalic regions [tegmentum (TEG) and interpeduncular nucleus (IPN)] on the latency and duration of TI (induced by postural inversion and by movement restriction) and on the latency of the motor response to a nociceptive stimulus (hot plate) in toads. Electrolytic lesions of TEG and IPN promoted an increase in the duration of TI episodes. Neurotoxic lesion of these two regions also caused an increase in the duration of TI episodes. The effect was more intense in the animals with electrolytic lesion, possibly due to more extensive damage associated with this procedure or to damage of passage fibers. The results suggest that lesions of the midbrain TEG liberate basic circuits placed caudally and are involved in the organization of the TI response. It remains to be determined if the IPN exerts its effect directly on the caudal levels or by acting via the mesencephalic TEG. Lesions do not interfere with the latency of the motor response to a thermal noxious stimulus, indicating that the lesioned regions do not affect the reflexive response and are not essential for the perception of the noxious stimulus. PMID- 11274671 TI - Social memory in the rat: circadian variation and effect of circadian rhythm disruption. AB - Disruption of circadian rhythm can impair long-term passive avoidance memory of rats and mice. The present study investigated whether disruption of circadian rhythm can also impair social memory of male rats. Social memory was assessed using the social discrimination test, in which a short-term olfactory memory is formed by social interaction with a juvenile rat during a learning trial. After an intertrial interval, a retrieval trial is performed, in which social memory is expressed as a decreased attention paid to the same juvenile as compared to a new juvenile. First, the social memory at four different time points across the light dark cycle was measured with an intertrial interval of 10 or 25 min. There was no significant circadian variation of social memory across the light-dark cycle. Subsequently, the effect of a -6 or 12-h phase shift on social memory was studied. These phase shifts were previously found to impair long-term passive avoidance memory. However, no effect of either phase shift was observed in the social discrimination test. It is concluded that the disruption of circadian rhythm had no effect on the social memory of rats. Differences between short-term social memory and long-term passive avoidance memory are discussed in relation to their apparent differential susceptibility to the effects of circadian rhythm disruption. PMID- 11274672 TI - Tyrosine improves behavioral and neurochemical deficits caused by cold exposure. AB - The effects of acute cold stress were assessed behaviorally and neurochemically. The norepinephrine (NE) precursor, tyrosine (TYR), the catecholamine-releasing compound, amphetamine (AMPH), and the adrenoceptor agonist, phenylpropanolamine (PPA), were administered systemically either alone or in conjunction with TYR 30 min prior to cold exposure. All three sympathomimetic treatments dose-dependently improved performance in a forced swim test following hypothermia (T(c)=30 degrees C). AMPH/TYR or PPA/TYR combinations further improved performance vs. either agent given alone. Microdialysis showed elevated hippocampal NE concentrations in response to hypothermia. TYR further elevated NE concentration in cold/restrained rats vs. saline (SAL)-treated controls. These results suggest that sympathomimetic agents, including the nutrient TYR, which enhance noradrenergic function, improve performance in animals acutely stressed by hypothermia. PMID- 11274673 TI - Oral irritation by sodium chloride: sensitization, self-desensitization, and cross-sensitization to capsaicin. AB - Psychophysical methods were used to investigate the irritant sensory properties of concentrated NaCl. The first experiment investigated potential sensitization and desensitization properties. Subjects rated the intensity of the irritation elicited by 10 successive applications of 5 M NaCl on one side of the dorsal surface of the tongue. The mean irritant sensation increased significantly across trials, consistent with sensitization. To test for self- and cross desensitization effects of unilateral sequential stimulation with NaCl followed by a 10-min rest period, either 5 M NaCl or 10 microM capsaicin was applied bilaterally. In a two-alternative forced-choice (2-AFC) test, subjects indicated which side of the tongue had a stronger irritant sensation. They also rated the intensity of irritation on each side separately. When NaCl was applied bilaterally, the side not previously receiving NaCl was chosen as stronger by a significant majority of subjects and was given significantly higher intensity ratings, consistent with self-desensitization. In contrast, when capsaicin was applied bilaterally, the side that had previously received sequential NaCl was perceived as having a significantly more intense irritation, consistent with cross-sensitization. In a second experiment, the effect of amiloride on NaCl evoked irritation was studied. One side of the tongue was treated with 1 mM amiloride, after which 5 M NaCl was applied bilaterally and subjects performed the same 2-AFC and rating procedures. Since amiloride significantly reduced the intensity of the irritant sensation, the contribution of amiloride-sensitive ionic currents or the Na+/H+ exchange pump (NHE) are suggested as possible transduction mechanisms in lingual nociceptors mediating NaCl-evoked oral irritation. PMID- 11274674 TI - Dilutions of low- vs. high-fat diets on intakes and gastric volumes in rats. AB - Rats adapt to changes in dietary energy to maintain nearly constant energy intakes. This regulation indicates that animals sense and respond to nutrient content. We sought to determine whether this response was affected by the fat content of the diet. Our second goal was to determine how energy dilution affected intragastric volumes. Rats were randomized to high (18% w/w) and low fat (4.5% w/w) as the energy density of the diet was altered from 2.0 to 3.5 kcal/g. Average energy intake during 7-h feeds rose steeply (P<.01) when density was increased from 2.0 to 3.0 kcal/g, but modestly as density increased from 3.0 to 3.5 kcal/g. In other rats on 18% vs. 32% fat diets, energy intakes increased significantly (P<.01) as density of the diet was raised from 3.5 to 4.5 kcal/g. During diets at 2.0 and 2.5 kcal/g, animals on 18% fat ate fewer kilocalories than those on 4.5% fat; but over 3.0-4.5 kcal/g, energy intake was similar regardless of fat concentration (4.5-32%). Gastric contents after 7-h feeds increased with grams of food ingested similarly for high- and low-fat diets. We concluded that in rats: (a) compensation to energy dilution or concentration was inexact but (b) was about equal for high- vs. low-fat diets; thus, high fat was as well sensed as high carbohydrate; (c) compensations for energy densities were made despite varied gastric volumes; thus, rats learned to override the stimulus of gastric stretch and to sense energy via extra gastric mechanisms. PMID- 11274675 TI - Age-dependent influence of dietary zinc restriction on short-term memory in male rats. AB - Zinc is an essential micro-nutrient involved in numerous physiological functions. The high content of zinc in the hippocampus, coupled with the integral involvement of the hippocampus in memory, strongly implicates zinc in memory processing. The hypothesis of the current study was that dietary zinc restriction influenced short-term memory in postweaned rats, and this influence was age dependent. Male rats (43 days to 18 months old) were divided into five experimental groups based on age, and fed zinc-adequate (zinc at 20 mg/kg as zinc chloride) or zinc-deficient (zinc less than 1-2 mg/kg) diets for a minimum of 3 weeks. Short-term memory was assessed using the distal-cue version of the Morris water maze (MWM). All rats fed the zinc-restricted diet exhibited cyclic anorexia, decreased weight gain, and significantly lower liver and femur zinc concentrations compared to age-matched controls. Further, whole brain, hippocampal, and cerebral wet weights were significantly reduced in the zinc restricted treatment groups of all the age groups. Only zinc-restricted rats that were less than 62 days of age at the start of zinc restriction demonstrated significantly prolonged escape latencies in the water maze, indicating deficits in short-term memory. Regression analyses confirmed that the short-term memory deficits were correlated with significantly lower hippocampal and cerebral zinc concentrations compared to age-matched control and pair-fed rats. These results emphasize the significance of a critical age of influence for dietary zinc in memory processing, and the importance of considering age when studying zinc nutriture and CNS function. PMID- 11274677 TI - Naloxone attenuates the conditioned place preference induced by wheel running in rats. AB - Pairings, during which an episode of wheel running is followed by confinement in a distinctive place, produce conditioned place preference (CPP) in rats. This finding indicates that wheel running has a rewarding effect that outlasts the activity itself. In two similar experiments, we tested the hypothesis that this rewarding effect of wheel running is mediated by endogenous opioids. During a paired trial, the rats in the naloxone group were first allowed to wheel run for 2 h, then injected with naloxone (0.5 or 0.1 mg/kg in Experiments 1 and 2, respectively), and 10 min later placed in a distinctive chamber. During an unpaired trial, these rats were confined in an adjoining chamber without wheel running. Naloxone was injected before placement in both chambers, so that if naloxone-induced conditioned place aversion occurred, it would have counteracting effects on performance during the preference test. The rats in the saline group were similarly treated, except that saline was injected instead of naloxone. CPP occurred in the saline group, but not in the naloxone group. Thus, naloxone attenuated the CPP induced by wheel running. This finding supports the hypothesis that the rewarding effect of wheel running is mediated by endogenous opioids. PMID- 11274676 TI - Temporal and spatial dynamics of corticosteroid receptor down-regulation in rat brain following social defeat. AB - The experiments explored the nature and time course of changes in glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) binding in homogenates of various brain regions and pituitary of male Wistar rats following social defeat stress. One week after defeat, the binding capacity of GRs was decreased in the hippocampus and the hypothalamus while no changes were observed in the parietal cortex and the pituitary. The number of MRs remained at the same level as in undefeated rats. Three weeks postdefeat, the initially down-regulated GR returned to baseline level in the hippocampus and the hypothalamus. However, GR binding was now decreased in the parietal cortex. Severe down-regulation of MRs was detected in the hippocampal and septal tissue. The results show that brief but intense stress like social defeat induces a long-lasting down-regulation of corticosteroid receptors and that the temporal dynamics of these changes are not only differential for GRs and MRs but also for brain sites. PMID- 11274678 TI - Social dominance rank and accessory sex glands in wild adult male house mice born to food-deprived mothers. AB - Food deprivation after weaning often has greater effects on the reproduction of females than males. However, if animals are deprived prenatally (i.e., through deprivation of the mother during gestation), the reproduction of males may be more negatively impacted because it may decrease their ability to compete with other males and their attractiveness to females. We tested the predictions that adult sons of females that are food-deprived during gestation would tend to lose agonistic encounters with sons of well-nourished (control) females and would have smaller accessory sex glands as well. Sons of control mothers were more frequently dominant to sons of deprived mothers. They also had heavier vesicular coagulating gland complexes and tended to have heavier preputial glands. However, among males that had not been tested for social dominance rank, there were no such differences in accessory gland weights. These data indicate that maternal food deprivation affects sons only if they engage in agonistic encounters. These effects may be due to a disruption of the organizational effects of testosterone that occur in neonatal male mice and they are likely to have a strong negative impact on the reproduction of the sons of deprived mothers. PMID- 11274679 TI - Changes in the auditory-evoked potentials induced by fear-evoking stimulations. AB - It is long established that the inferior colliculus is involved in conveying all kinds of auditory information to higher cortical structures. Moreover, gradual increases in the electrical stimulation of this structure produces progressive aversive responses from vigilance, through freezing, until escape. Recently, we have shown that microinjections of the excitatory amino acids, N-methyl-D aspartate (NMDA) and glutamate, into the inferior colliculus mimic these aversive effects. In the present study, we extend these observations showing that unilateral microinjections of 5 nmol of glutamate into the inferior colliculus--a dose that causes freezing behavior--in rats with bilateral recording electrodes into this structure produce an increase in the magnitude of the collicular-evoked potential in the ipsilateral side of the injection in relation to saline-injected animals. Besides, the application of two kinds of fear-evoking stimulations- light as a conditioned stimuli (CS) and ultrasound signals at the frequency of 22 kHz--also produced an increase in the amplitude of the evoked potentials recorded from the inferior colliculus in comparison to control situations without aversive stimuli presentations. These data support previous reports showing that fast acting excitatory amino acid receptors in this midbrain region are involved in the processing of auditory information. Moreover, fear-eliciting stimulations, such as light-CS and ultrasound signals, increase acoustically evoked firing of neurons in the central nucleus of the inferior colliculus of rats. PMID- 11274680 TI - NaCl thresholds: relationship to anterior tongue locus, area of stimulation, and number of fungiform papillae. AB - NaCl detection thresholds were determined for 12.5- and 50-mm(2) lingual areas at four anterior tongue locations in eight subjects using a device that allowed for accurate temporal and spatial presentation of tastants to small regions of the anterior tongue. The locations, all on the right side of the tongue, were the tongue tip, an area 1.7 cm posterior to the tongue tip, and regions 1.7 and 3.4 cm posterior to the tip along the tongue's lateral margin. Stimulus duration was 0.75 s. Thresholds were established using a two-alternative forced-choice single staircase procedure, and the number of fungiform papillae at each stimulation site was counted with the aid of videomicroscopy. NaCl thresholds were lower for the 50-mm(2) than the 12.5-mm(2) stimulation area at all target sites, and were directly related to papillary number among and within the stimulated regions. For a given number of papillae, thresholds were lower within the 12.5-mm(2) than within the 50-mm(2) stimulation region, likely reflecting taste bud density and activation of common afferent pathways. The tongue tip was more sensitive than any other tongue region, and the lateral margins were seemingly more sensitive than the lingual centrum. Large individual differences in taste sensitivity and tongue papilla numbers were noted, and some subjects were insensitive to the highest tastant concentrations at the nontip loci. This study empirically demonstrates that NaCl detection sensitivity varies across discrete regions of the anterior tongue and is related to the relative number and density of fungiform papillae. PMID- 11274682 TI - Ambient temperatures preferred by humans acclimated to heat given at a fixed daily time. AB - We investigated preferred ambient temperatures (T(pref)) of heat-acclimated humans to assess their behavioral thermoregulation. Seven male volunteers were exposed to an ambient temperature (T(a)) of 42 degrees C and relative humidity (RH) of 40% for 4 h (14:00-18:00 h)/day for 9-10 consecutive days. Rectal temperature (T(re)) was measured, and T(pref) was determined at two distinct times of day, 09:00-11:00 h (AM test) and 14:00-16:00 h (PM test), in both heat- and nonheat-acclimated (control) conditions. Heat acclimation significantly decreased T(re) only in the PM test. There was no difference in the T(pref) between the two tests in the control condition. However, T(pref) in the PM test was significantly lower than that of the AM test in the heat-acclimated condition. The findings suggest that repeated heat exposure in humans for 4 h at a fixed time daily alters the core temperature level and behavioral thermoregulatory function, particularly during the period when the subjects had previously been exposed to heat. PMID- 11274681 TI - Olfactory discrimination deficits in n-3 fatty acid-deficient rats. AB - Docosahexaenoic acid (DHA), a long chain n-3 fatty acid, is present in high concentrations in the central nervous system. Although the role that DHA may play in neural function is not well understood, infants fed formulas containing low levels of n-3 fatty acids have decreased visual acuity and neurodevelopmental test scores. The present experiment assessed whether dietary manipulations that decrease the concentration of DHA in the brain interfered with olfactory-based learning. We fed rats a diet that provided adequate n-3 fatty acids or a diet that was deficient in n-3 fatty acids for two generations. The second generation n-3-deficient group had 81% less brain DHA (82% less in olfactory bulb) compared to the n-3-adequate group and made significantly more errors in a series of olfactory-cued, 2-odor discrimination tasks compared to the adequate group. These results suggest that lower levels of central nervous system DHA lead to poorer performance in a series of simple odor discrimination tasks. PMID- 11274683 TI - The impact of sucrose-derived unconditioned and conditioned negative feedback on the microstructure of ingestive behavior. AB - We describe at the microstructural level the impact of unconditioned and conditioned negative feedback on the licking behavior of the rat. Seven groups of rats were trained to ingest one of seven different concentrations of sucrose (0.025, 0.05, 0.1, 0.2, 0.4, 0.6, and 0.8 M sucrose) under real-feeding conditions until intake was stable. They were then given three sham-feeding tests with the same solution. We compared the size of the clusters (SC) and number of clusters (NC) during the 17-min period when intake rate was declining in the real feeding test with SC and NC during the corresponding 17-min period in the following sham-feeding test. Intake increased significantly over the three sham feeding tests with the three highest concentrations, indicating the extinction of conditioned negative feedback. With these three solutions, we compared the microstructure of licking behavior in the first with that in the third sham feeding test to determine if conditioned negative feedback affected SC or NC or both. The effect of both unconditioned and conditioned negative feedback on licking behavior was to decrease the NC without significantly affecting their size. We conclude that negative feedback derived from the accumulation of sucrose in the gastrointestinal tract decreases the probability of initiating a bout of licking during a pause. It has no effect on the ability to continue a bout of licking once it has begun. PMID- 11274684 TI - Exposure to a novel stimulus reduces anxiety level in adult and aging rats. AB - Male Wistar rats aged 3, 15 and 24 months were isolated and housed individually for 72 h prior to being subjected to inanimate objects (two objects per rat, each 1.5 cm in diameter and 4 cm in length, made of surgical gauze). Following the exposure to the objects, rats were subsequently tested in an elevated plus-maze. The inanimate objects induced locomotor activity, chewing and transportation of the object. This changed to social interaction and play-like behavioral activity in rats of all ages that were kept in small groups with a latency of 1 to 2 min. One hour after the start of exposure, the animals were tested in the elevated plus-maze to measure anxiety behavior. It was found that all age groups spent significantly more time in the open arm of the elevated plus-maze than the corresponding controls. Latencies to first entry into open arms were also significantly lowered. The number of entries to the open or to the dark arm, however, did not show statistical difference, indicating that the novel object induced activity failed to exert influence on gross motor activity in the elevated plus-maze. In conclusion, the stimulation elicited by the exposure to novel stimulus (inanimate objects) resulted in a significant reduction of anxiety level both in adult and in aging rats. PMID- 11274685 TI - Age specificity of the relationship between serum cholesterol and mood in obese women. AB - Despite increasing evidence of an association between lower cholesterol levels and negative mood, no study has specifically investigated this relationship in obese people, a population at high risk for both dyslipidaemia and depression. Data on serum cholesterol and mood were collected in a group of 73 healthy women, aged 16 to 76 years, with different degrees of obesity and widely varying total cholesterol concentrations. Mood was assessed using three self-rated scales: the Toronto Alexithymia Scale (TAS-20), the State-Trait Anger Scale (STAS), and the Beck Depression Inventory (BDI). The association between lower total cholesterol levels and negative mood was age-dependent. No significant association was found in the younger age group (<50 years). In contrast, in the subgroup of older women, serum cholesterol was negatively and significantly correlated with the TAS 20 and the STAS. The negative correlation between serum cholesterol and the BDI was nearly statistically significant. Restricting analysis to the subjects in the highest quartile of the age distribution (>60 years) yielded stronger correlations between cholesterol and mood. In this sample of obese women, the relationship between lower cholesterol levels and negative mood was age-specific and limited to the older age group. The results of this study suggest that preventive programs or drug treatments for reducing cholesterol levels in elderly obese women should include a careful evaluation of mood state. PMID- 11274686 TI - Stress-induced increment on total bronchoalveolar cell count in OVA-sensitized rats. AB - The influence of stress on total leukocyte count from bronchoalveolar lavage (BAL) was investigated in rats sensitized and challenged with ovalbumin (OVA). The animals were injected intraperitoneally with a suspension of OVA plus aluminum hydroxide in 0.9% NaCl (Day 0) and boosted at Day 7 with an identical OVA solution, administered subcutaneously. From the first to the 13th day after sensitization, rats were placed individually in a shuttle box where they received 50 escapable footshocks per day, always preceded by a sound signal (S); the responses that occurred during both S and shocks canceled the stressful stimulation. On Day 14, animals were submitted to a single session of 50 inescapable footshocks, preceded by the same S; immediately after, the animals were submitted to a 1% OVA-inhalation challenge. Results showed high levels of stress in the shocked animals as detected through both ultrasonic vocalizations (UVs) and social interaction test in an open field. Total leukocyte count in BAL from stressed animals (24 h post-OVA challenge) revealed a significant increase in the number of inflammatory cells in comparison to that measured in sensitized, nonstressed challenged rats. These data demonstrate that stress plays a relevant and important role on total bronchoalveolar cell count in OVA-sensitized rats. PMID- 11274687 TI - PTZ-induced seizures in rats: effects of age and strain. AB - The susceptibility to pentylenetetrazol (PTZ)-induced seizures during postnatal ontogeny [postnatal day (PN) 10-220] was investigated in two rat strains. The WAG/Rij strain, genetically prone for developing generalized absence epilepsy, and Wistar rats were tested and compared at PN 10, 26, 30, 70, 90, 125, and 220 on the PTZ-convulsive threshold. A subconvulsive dose of 25-mg/kg PTZ was administered every 15 min, and the occurrence of clonic and tonic-clonic seizures was scored. The 10-day-old pups were quite sensitive to PTZ and showed mainly clonic seizures. The highest threshold and latency of PTZ-induced clonic and tonic-clonic convulsions were observed at PN 26 in both strains. From that age onwards, the seizure threshold significantly decreased and reached a minimum at PN 220. Between strain comparisons showed that WAG/Rij rats have a lower tonic clonic seizure threshold than Wistar rats. The data indicate that changes in susceptibility first quickly decreases until PN 26-30 and then tend to monotonically increase with age, and that genetically prone nonconvulsive WAG/Rij rats are more vulnerable to convulsive seizures induced by PTZ than Wistar rats. PMID- 11274688 TI - Relation between PROP taster status and fat perception, touch, and olfaction. AB - We tested the hypothesis that fat perception (sensitivity to and preferences for fat) may be linked to 6-n-propylthiouracil (PROP) taster status as a result of differences in trigeminal innervation of the oral cavity. In addition, we examined the relationship between taster status and sensitivity to other taste attributes, as well as tactile and olfactory sensitivities. Subjects (40 nontasters, 67 medium tasters, and 40 supertasters of PROP) rated samples (potato chips, chocolate drink, mashed potatoes, and vanilla pudding) varying in fat and flavor concentrations for the intensity of fattiness, saltiness, and sweetness, first without and then with nose clips, and for liking. Tactile sensitivity of the tongue was assessed according to responses to stimulation with Von Frey filaments (2.36, 2.44). Olfactory thresholds were determined for two odors (diacetyl and phenylethyl methyl ethyl carbinol). In general, taster status was not related to the perceptions of fat, saltiness, and sweetness. Subjects were able to accurately assess the fat content of the samples. Increasing the flavor levels in the potato chips and mashed potatoes enhanced the perception of fattiness for these systems. Supertasters were more sensitive to stimulation on the median of the tongue with the no. 2.36 Von Frey filament, and the olfactory thresholds for diacetyl were lower for PROP tasters and supertasters than for nontasters. PMID- 11274689 TI - Sex differences in relation to conditioned fear-induced enhancement of morphine analgesia. AB - A number of studies have reported that both the immediate and proactive effects of exposure to a shock stressor are less pronounced in female than in male rats. A separate area of research has demonstrated that female rats are less sensitive to the analgesic effects of morphine than males. Experiments from our laboratory, as well as others, have found that exposure to a context associated with shock (i.e., conditioned fear context) at the time of morphine administration, enhances the analgesic effects of morphine. Since previous studies have exclusively employed male rats, the purpose of Experiment 1 was to determine if a sex difference exists to this context conditioned fear-induced enhancement of morphine-induced analgesia. The findings of Experiment 1 showed that females do not appear to exhibit conditioned fear-induced enhancement of morphine analgesia as compared to males. Experiment 2 demonstrated that females exhibited higher levels of conditioned fear-induced enhancement of morphine analgesia during diestrus I than estrous. Experiment 3 demonstrated that females exhibited lower levels of conditioned analgesia compared to males, while both groups exhibited similar freezing levels. The findings of the present experiments suggest that the sex difference observed in Experiment 1 may be due to differences in conditioned analgesia. PMID- 11274691 TI - Ocular nocardia infections with special emphasis on the cornea. AB - Nocardia are aerobic, gram-positive, nonmotile and branching filamentous bacteria. Corneal infection by Nocardia is rare. Trauma is the most common predisposing factor. Isolated case reports of nocardial infection associated with contact lens wear and laser in situ keratomileusis (LASIK) have been reported. The clinical picture usually consists of superficial patchy infiltrates, which may be arranged in a wreath pattern. Presence of gram-positive, branching, beaded filaments that stain with 1% acid-fast stain (using 1% sulfuric acid, modified Kinyoun's method) in smears of corneal scrapings is suggestive of nocardial infection. Nocardia grow on commonly used media as tiny, white, dry colonies. Available knowledge and clinical experience suggest that although sulfacetamide eyedrops can be tried as the initial drug, trimethoprim-sulfamethoxazole and amikacin are effective drugs. Once therapy is initiated, the infiltrate responds promptly and resolves, forming a corneal scar with or without vascularization, and good visual recovery can be expected. PMID- 11274692 TI - Central disorders of vision in humans. AB - Over the past 20 years, researchers have discovered over 30 separate visual areas in the cortex of the macaque monkey that exhibit specific responses to visual and environmental stimuli. Many of these areas are homologous to regions of the human visual cortex, and numerous syndromes involving these areas are described in the neurologic and ophthalmic literature. The focus of this review is the anatomy and physiology of these higher cortical visual areas, with special emphasis on their relevance to syndromes in humans. The early visual system processes information primarily by way of two separate systems: parvocellular and magnocellular. Thus, even at this early stage, visual information is functionally segregated. We will trace this segregation to downstream areas involved in increasingly complex visual processing and discuss the results of lesions in these areas in humans. An understanding of these areas is important, as many of these patients will first seek the attention of the ophthalmologist, often with vague, poorly defined complaints that may be difficult to specifically define. PMID- 11274694 TI - The dilemma of color deficiency and art. AB - No "major" painter is known to be color deficient. Are there truly no color deficient artists, or have they not been recognized? The historical literature cites criteria for recognizing color deficiency in artists, but they are hard to apply without knowing the intentions of an artist. The work and commentary of a color-deficient artist who works currently in Paris are presented as an example. He uses a limited palette of colors, based on advice from colleagues as much as his own perceptions, and he uses colors in ways that do not always fit with expectations for color deficiency. Biographies of earlier painters suggest that there were a few whose color sense was poor, but these painters used assistants to help. The color sense of others, such as the English landscape painter John Constable (1776-1837), has been questioned because of a preponderance of suspicious color, such as murky green. However, there are good reasons to doubt that Constable was color deficient. It is instructive to know how proven color deficiency has influenced an artist's style. When medical information is unavailable, the best advice for the diagnostically-inclined observer is just to enjoy the art. PMID- 11274695 TI - Chemotherapy for eye cancer. AB - Chemotherapy has been used to treat a multitude of eye cancers. We attempted to review the role of chemotherapy in the treatment of ocular, adnexal, and orbital malignancies by conducting an extensive search of the medical literature. Unfortunately, the published reports typically contain few patients with limited follow-up, precluding definitive recommendations. For most eye cancers, multicenter trials will offer the potential to gather the numbers of patients required to determine the clinical utility of chemotherapy. PMID- 11274696 TI - Retinal findings in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (cadasil). AB - We describe a 45-year-old man with biopsy proven cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL). This patient demonstrated unique retinal findings, including arteriole narrowing and sheathing, irregular choroidal filling on fluorescein angiography, and patchy visual field loss. CADASIL is a hereditary, nonamyloid, nonathersclerotic microangiopathy. This disorder has been mapped to chromosome 19 with mutations in the Notch 3 gene. Deposits of granular osmiophilic material in the basal lamina of the smooth muscle cells of small vessels are considered pathognomonic for CADASIL and are typically seen only on electron microscopy. Although CADASIL is a systemic vascular disease affecting the entire arteriole tree, we are unaware of other reports describing the retinal findings observed in our patient. PMID- 11274697 TI - Management of glaucoma in pregnancy and lactation. AB - A 30-year-old pregnant woman with glaucoma is presented. The management of her case is used as a basis for a discussion of the use of glaucoma medications, including newer formulations, during pregnancy and lactation. PMID- 11274698 TI - Ocular microtremor (OMT): a new neurophysiological approach to multiple sclerosis, by C. Bolger, S. Bojanic, N. Sheahan, J. Malone, and D. Coakley. J Neurol Neurosurg Psychiatry 68:639-42, 2000. PMID- 11274699 TI - Objective evaluation of improvement in optic neuropathy following radiation therapy for thyroid eye disease, by S. Rush, J. M. Winterkorn, and R. Zak. Int J. Radiat Oncol Biol. Phys 47:191-4, 2000. PMID- 11274700 TI - Retinopathy and nephropathy in patients with type 1 diabetes four years after a trial of intensive insulin therapy, by The Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications Research Group. N. Engl. J. Med 342:381-9, 2000. PMID- 11274702 TI - Effect of immobilization stress on anticonvulsant actions and pharmacokinetics of zonisamide in mice. AB - The effects of immobilization stress on anticonvulsant actions and pharmacokinetics of zonisamide were investigated in mice. Oral administrations of zonisamide (10, 20, and 50 mg/kg) dose-dependently reduced incidence of tonic extension (TE) induced by maximal electroshock seizure (MES). Immobilization stress for 2 h immediately after the administration of zonisamide further enhanced the anticonvulsive actions of it. On the other hand, the serum zonisamide concentrations in stressed group were lower during the first 30 min after the administration compared with that in nonstressed control group. Thereafter, there were no significant differences in the serum concentrations between two groups. The brain zonisamide concentration and the concentration ratio of brain/serum at 2 h after administration of zonisamide (50 mg/kg) were significantly higher in stressed group, rather than that in the nonstressed control group without changing the serum concentration. These results suggest that immobilization stress enhances anticonvulsant actions of zonisamide, and that increases of brain zonisamide concentration by immobilization stress may be related with this phenomenon. PMID- 11274701 TI - Quantifiable dose-dependent withdrawal after morphine discontinuation in a rat model. AB - We evaluated the intensity of the withdrawal symptoms after the discontinuation of the morphine infusion in rats. Opiate addiction was induced by progressively increasing intraperitoneal morphine infusion rates. The control group (Group 1) received normal saline. The initial morphine rates were 1, 4, and 16 mg kg(-1) h for Groups 2, 3, and 4, respectively. Infusion rates were gradually increased by a factor of 1.4, 2, 2.8, and 4 on the second, third, fourth, and fifth days, respectively. The last rate was used for 48 h and then infusions were disconnected. Weight reduction, food consumption, and water intake were used for evaluation of withdrawal. All morphine groups showed a significant reduction of body weight during the 4 postdiscontinuation days and a decline in food and water intake on the first postdiscontinuation day. All changes were dependent on the morphine infusion concentration. No changes were observed in the control group. We suggest that the rat model used in this study may be utilized for quantification of spontaneous withdrawal. PMID- 11274703 TI - Effects of sweetened ethanol solutions on ethanol self-administration and blood ethanol levels. AB - The enhancement of voluntary self-administration of ethanol by sucrose or saccharin was tested in conjunction with measurements of blood ethanol levels. Adult male rats were given access to both tap water and one of five solutions: 0.125% saccharin, 10% sucrose, ethanol, saccharin+ethanol, or sucrose+ethanol. The rats receiving the sucrose+ethanol solution drank consistently more ethanol (>5 g/kg/day) than did the rats receiving the saccharin+ethanol solution (<3 g/kg/day) or ethanol only (<2 g/kg/day). Both sweetened solutions produced higher ethanol consumption during these periods than ethanol alone. However, no significant differences in blood ethanol levels were found between the sucrose+ethanol and saccharin+ethanol conditions, when tested at different intervals on Day 44 or Day 45 of ethanol consumption. Following 45 days of consumption, no change in the bicuculline seizure threshold was observed in the ethanol-consuming rats compared to the controls. In a separate study using 90 naive rats, rats were gavaged with ethanol (1, 2, or 3 g/kg) containing either 10% sucrose (n=10 for each dose of ethanol), 0.125% saccharin (n=10 for each dose of ethanol), or ethanol alone (n=10 for each dose of ethanol), and blood was collected from the tip of the tail 30, 60, 180, 300, and 540 min later and analyzed for ethanol concentrations. Sucrose significantly decreased the resultant blood ethanol levels at several time points following gavage. These results indicate that sucrose can significantly alter blood ethanol levels and that chronic self-administration of a sweetened ethanol solution for 6 weeks does not produce ethanol dependence. PMID- 11274704 TI - Anxiolytic effects of valproate and diazepam in mice are differentially sensitive to picrotoxin antagonism. AB - Although it is widely believed that the anxiolytic effects of benzodiazepines are mediated through facilitation of GABA(A) receptor function, behavioural studies have to date provided rather weak support for this hypothesis. In particular, considerable inconsistency has been noted both for the effects of GABAergic manipulations in animal models of anxiety and the ability of GABA(A) receptor antagonists to block the anxiolytic effects of diazepam (DZ) and chlordiazepoxide. In view of the sensitivity of the murine plus-maze to the anxiety-modulating effects of GABAergic agents as well as classical benzodiazepines, the current study examined the extent to which the anxiolytic actions of valproic acid (VPA) and DZ in this test involve picrotoxin (PX) sensitive receptor mechanisms. Subjects were male DBA/2 mice, test duration was 5 min, and ethological scoring methods were employed. Our results show that, while devoid of intrinsic behavioural effects under present test conditions, PX (0.25 0.5 mg/kg) selectively antagonised the anxiolytic-like (but not other) effects of VPA (400 mg/kg). In contrast, the same doses of PX failed to block any of the behavioural changes induced by DZ (1.5 mg/kg), including disinhibition of open arm exploration. These data suggest that the plus-maze anxiolytic effects of DZ in DBA/2 mice are not mediated through PX-sensitive GABA(A) receptors. Further studies will be required to assess the generality of present findings to other mouse strains, species and behavioural paradigms. PMID- 11274705 TI - Use of negatively reinforcing electrical brain stimulation to detect conventional and nonconventional anxiolytics as well as an anxiogenic drug. AB - The present study determined whether anxiolytics such as diazepam (DZP), the benzodiazepine (BZD) receptor-selective agonist abecarnil (ABC), or the 5-HT1(A) agent buspirone (BUS) would increase the response latency of rats to switch-off electrical brain stimulation (EBS) of the periaqueductal gray (PAG). We also investigated the effects of pentylenetetrazole (PTZ), a purported anxiogenic. Given acutely, DZP (2.5 and 5 mg/kg, ip) and ABC (0.5 and 1 mg/kg, ip) increased response latency. The BZD receptor antagonist flumazenil (10.0 mg/kg, ip) blocked these effects. Increasing the frequency of EBS reversed the effects of DZP and ABC, suggesting that motor disruption did not account for the increase in latency seen with these drugs. Given acutely, BUS (10.0 mg/kg, ip) also increased response latency, which was likely due to motor disruption because it was not reversed by increasing the frequency of EBS. When BUS (2.5 mg/kg, ip) was given every 8 h for 3 days, an increase in latency was also obtained, which was reversible by increasing the frequency of EBS. Finally, PTZ (10 and 20 mg/kg, ip) shortened the latency to respond. These results (1) suggest that DZP, ABC, and chronic BUS attenuate, whereas PTZ potentiates, the negative reinforcing stimulus (NRS) induced by PAG stimulation, and (2) support the hypothesis that the switch off procedure accurately detects anxiolytic and anxiogenic drugs. PMID- 11274706 TI - Ventromedial hypothalamic mediation of sucrose feeding induced pain modulation. AB - Electrophysiological and behavioural studies suggest a modulatory role of ventromedial nucleus of the hypothalamus (VMH) in nociceptive behaviour. Lesion of the VMH produces hyperalgesia and a greater preference for sucrose solution. Hyperalgesia is also produced by sucrose feeding. To explore specifically the contribution of glucoreceptor neurons of the VMH in the mediation of sucrose-fed hyperalgesia, 2-deoxy-D-glucose (2-DG, antimetabolite of glucose) was slowly albeit continuously infused (1 microl/h for 7 days by microinfusion pumps) into the VMH of adult male rats. Simultaneously, the rats underwent tests for their nociceptive responses in control and sucrose-fed states. The tests for nociception, namely, tail flick latency (TFL), thresholds of tail flick (TF), vocalization during stimulus (SV), vocalization after discharge (VA) were recorded at 0500 h. The tests were repeated after 6, 12, and 48 h in 1 M saline (control group) and 2-DG (experimental group) microinfused rats. Rats were presented with sucrose (20%) solution for 48 h at 0500 h ad libitum in addition to food pellets and tap water. Infusion of 2-DG per se in the VMH led to hypoalgesia (in threshold of TF, SV, VA) while feeding sucrose for 6-12 h per se led to hyperalgesia (in TFL, threshold of SV and VA). Sucrose feeding to 2-DG rats, however, attenuated the hypoalgesia of 2-DG as well as the hyperalgesia of sucrose feeding. The results suggest that the VMH glucoreceptor neurons probably modulate sucrose mediated phasic pain responses. PMID- 11274708 TI - Schedule-dependent effects of haloperidol and amphetamine: multiple-schedule task shows within-subject effects. AB - A two-lever, multiple-schedule task was used to evaluate the effects of haloperidol (HA) and amphetamine (AM) on responding controlled by continuous reinforcement (CRF) and progressive ratio (PR) schedules of reinforcement. Rats were trained to press one lever for food delivered on a CRF schedule and the other lever for food delivered on a PR schedule. The operative schedule was signaled by the illumination of a cuelight mounted above the appropriate lever. Following 30 sessions of training, dose-response functions were determined for HA (0.0075, 0.015, 0.03, and 0.06 mg/kg) and AM (0.0625, 0.125, 0.25, 0.50, 0.75, and 1.00 mg/kg). Both drugs produced dose- and schedule-dependent effects. For example, administration of 0.03 mg/kg HA did not affect responding under the CRF schedule but did reduce responding during PR components, whereas administration of 0.06 mg/kg reduced responding under both schedules of reinforcement. Some doses of AM produced increased responding under the CRF schedule and, within the same session, decreased responding under the PR schedule. The results with HA are consistent with the view that interfering with dopaminergic function affects the allocation and maintenance of responding and that this effect depends on properties of the schedule of reinforcement. The results with AM emphasize that statements about the effects of the drug on positively reinforced behavior cannot be made without reference to specific schedules of reinforcement. PMID- 11274707 TI - The effect of the NMDA receptor blocker, dextromethorphan, on cribbing in horses. AB - Stereotypic cribbing in horses is thought to involve excess dopaminergic activity within the striatum. Various models of stress-induced stereotypies including cribbing in horses postulate that stress stimulates the release of endorphins, triggering the release of striatal dopamine. Dopamine in turn activates basal ganglia motor programs, reinforcing behavior via a reward mechanism. Furthermore, the release of dopamine by endorphins has been shown to depend on activation of NMDA receptors. In the present study, horses identified as cribbers and volunteered by their owners were treated with the NMDA receptor antagonist dextromethorphan (DM). When DM was administered via jugular injection (1 mg/kg), eight of nine horses responded with reductions in cribbing rate (CR) compared to baseline, and cribbing was suppressed completely for a period of time in almost half of the horses tested. PMID- 11274710 TI - Cocaine induces conditioned place preference and increases locomotor activity in male Japanese quail. AB - The conditioned place preference (CPP) procedure is a popular method used for testing the rewarding properties of human drugs of abuse. Most CPP studies utilize mammalian models. However, avian species have better visual systems than rodent species, and because the cues that become associated with human drug taking behavior are often visual, Aves might serve as an alternative animal model for investigating drugs of abuse. In three experiments, we examined the locomotor stimulant and rewarding effects of cocaine in adult male Japanese quail. In Experiment 1, cocaine increased locomotor activity relative to saline. In addition, behavioral sensitization was evident across repeated injections. In Experiment 2, CPP was established after six pairings of cocaine. Finally, the dopamine D(2) receptor subtype antagonist eticlopride did not attenuate acquisition of cocaine CPP in Experiment 3. Rather, subjects receiving pretreatment of eticlopride demonstrated a place preference for the cocaine paired context. In contrast, pretreatment of eticlopride reduced cocaine-induced locomotor activity. The findings suggest that drug-reward processes may be highly conserved across species and that birds may serve as a viable model for investigating drug-reward processes especially with regard to the ability of cocaine to become associated with visual cues. PMID- 11274709 TI - Aniracetam restores motivation reduced by satiation in a choice reaction task in aged rats. AB - This study aims to examine the effects of aniracetam on satiation-induced poor performance in a choice reaction task. Aged rats that mastered the task under food restriction stably maintained the task performance for a long period. Satiation by successive free feeding greatly diminished the performance. Satiation resulted in a decreased % correct, increased % omission and prolonged choice reaction time, indicating a reduction in lever response with low choice accuracy and slow responding speed. Repeated administration of aniracetam (30 mg/kg, po, for 14 days) partially recovered the choice accuracy and lever response, but not the responding speed, task-associated motor activity or impulsivity. In addition, aniracetam did not affect the animals' weights. These results indicate that satiation reduces motivation to perform and attain the task. Aniracetam may restore motivation, probably by improving poor behavioral states (daily attentional and vigilance failures), thereby creating the driving force. PMID- 11274711 TI - The effects of pyridostigmine bromide on progressive ratio performance in male and female rats. AB - Small doses of pyridostigmine bromide (PB) affect the acquisition and maintenance of food-maintained behavior in laboratory animals. The present experiment was designed to investigate the effects of this drug on food motivation. Male and female rats were trained to respond on a progressive-ratio schedule of reinforcement and treated with different doses of PB. PB dose-dependently decreased breaking points and response rates in male and female rats. Gender differences were not observed. The results indicate that decreased food motivation may be a factor that contributes to the behavioral effects of PB administration. PMID- 11274712 TI - Passive immunization against nicotine prevents nicotine alleviation of nicotine abstinence syndrome. AB - Passive immunization against nicotine interferes with its locomotor and pressor effects. The current study determined whether immunization could prevent another nicotine action: the reversal of nicotine abstinence syndrome. IgG containing 4.4 5.6% nicotine-specific antibody was isolated from rabbits immunized with 3'-amino methyl-nicotine conjugated to a carrier protein. Twenty rats were rendered dependent by 7 days of subcutaneous infusion of 3.15 mg/kg/day nicotine (expressed as the base). Upon termination of nicotine infusion, each rat was injected intraperitoneally with 150 mg of IgG from normal serum (n=13) or from nicotine antiserum (n=7). Twenty-two and one-half hours later, all rats were observed over 15 min for baseline nicotine abstinence signs. Two and one-half hours after baseline observations, seven of the 13 rats pretreated with control IgG and all seven rats pretreated with nicotine-specific IgG were then challenged by 0.12 mg/kg (sc) nicotine. The remaining six rats pretreated with control IgG were challenged with saline alone. All rats were then observed again for abstinence signs. Nicotine injection caused significantly less reduction of abstinence signs in the immunized rats. The nicotine effect in immunized rats was comparable to the saline effect in nonimmunized rats. Immunization also significantly reduced free serum nicotine concentration and nicotine distribution to the brain. These results raise the possibility that immunization might prevent nicotine consumption from relieving the discomforts of smoking cessation. PMID- 11274713 TI - Morphine withdrawal-facilitated aggression is attenuated by morphine-conditioned stimuli. AB - There is a considerable body of evidence indicating that stimuli associated with drug administration may become conditioned and evoke drug-like effects. The purpose of this study was to evaluate the ability of morphine-paired stimuli to affect an expression of morphine withdrawal-facilitated aggression. Individually housed aggressive adult mice were subjected to the repeated subcutaneous administration of morphine (twice a day, 8 days, increasing doses 10-80 mg/kg). Morphine treatment cessation facilitated an aggressive behaviour of animals during the second day of withdrawal. Subcutaneous but not intraperitoneal injection of saline attenuated the aggressive behaviour in morphine-withdrawn mice. These results suggest that the site of drug injection may serve as a conditioned stimulus. PMID- 11274714 TI - Gender and the behavioral manifestations of neuropathic pain. AB - A model of peripheral nerve injury was used to study gender differences in the development and progression of chronic constriction injury (CCI)-induced hyperalgesia and allodynia in male and female Fischer 344 FBNF1 hybrid rats. Rats were randomly assigned to one of the following treatment groups: (1) gonadally intact unligated males (male); (2) gonadally intact ligated males (male (CCI)); (3) castrated ligated males (male (CAS/CCI)); (4) gonadally intact unligated females (female); (5) gonadally intact ligated females (female (CCI)); and (6) ovariectomized ligated females (female (OVX/CCI)). A plantar analgesia meter and calibrated von Frey pressure filaments were used as the analgesiometric assays. In the absence of nerve injury, gonadally intact males responded significantly faster than females to a thermal nociceptive stimulus. The onset of the behavioral manifestations of unilateral ligation of the sciatic nerve did not differ as a function of sex or hormonal status (e.g., gonadally intact and gonadectomized male and female rats developed thermal hyperalgesia within 14 days post-CCI). Paw withdrawal latency (PWL) values of gonadally intact males returned to baseline control values after postligation day 14, whereas gonadally intact females, ovariectomized females and castrated males continued to elicit robust thermal hyperalgesic symptoms throughout the 35-day duration of the experiment. Allodynic responses to peripheral nerve injury were less variable across genders. These data suggest that the mechanisms underlying chronic nociceptive processing differ as a function of gender and gonadal hormone status. PMID- 11274715 TI - Involvement of spinal NK2 and NMDA receptors in aversive behavior induced by intra-arterial injection of capsaicin. AB - The spinal processing by which intra-arterial injection of capsaicin (CAP) induces vocalization response (VOR) was investigated in guinea pigs. Intrathecal pre-treatment with CP-96,345 (a selective NK(1) receptor antagonist, 50 nmol) did not affect the CAP-induced VOR. However, significant attenuation of the VOR was observed by intrathecal pre-treatment with a selective NK(2) receptor antagonist MEN-10,376 (40 nmol) accompanied with a significant change in the response modality. MK-801 [an N-methyl-D-aspartate (NMDA) receptor antagonist, 20 and 40 nmol] inhibited the CAP-induced VOR dose-dependently without affecting the response modalities. Furthermore, intrathecal co-treatment with 40-nmol MEN 10,376 and 40-nmol MK-801 resulted in a marked inhibitory effect on the VOR followed by a significant alteration of response modalities. Intrathecal pre treatment with neurokinin A (NKA; a tachykinin NK(2) receptor agonist, 1 nmol) enhanced the CAP-induced VOR. These behavioral results suggested that spinal NK(2) and NMDA receptors might have priority over NK(1) receptors in the spinal processing of nociceptive information from the CAP-sensitive nociceptor. PMID- 11274716 TI - Striatal dopamine sensitization to D-amphetamine in periadolescent but not in adult rats. AB - The neurobiological and behavioral facets of adolescence have been poorly investigated in relation to the vulnerability to psychostimulants. Periadolescent (33-43 days) and adult (>70 days) Sprague-Dawley rats underwent a 3-day treatment history with D-amphetamine (AMPH) at 0, 2, or 10 mg/kg (once a day). After a short 5-day-long withdrawal interval, freely moving animals were challenged with a 2-mg/kg AMPH dose and their behavior as well as in vivo intrastriatum dopamine (DA) release in the CNS were assessed. Microdialysis data indicated that AMPH history periadolescent rats showed a prominent sensitization of AMPH-stimulated DA release, whereas no such change was found in adult subjects. As expected, acute AMPH administration strongly reduced time spent lying still and increased levels of cage exploration in animals of both ages. A treatment history of high AMPH dosage was associated with a marked sensitization of the exploratory behavior in adults, whereas it induced a quite opposite profile in periadolescents. The latter group only was also characterized by a compulsive involvement in the stereotyped head-bobbing response. These results indicate that differently from adults, marked alterations in neurobiological target mechanisms are observed in rats around periadolescence as a consequence of a quite mild regimen of intermittent AMPH exposure. Thus, a neurobiological substrate for an age-related increased vulnerability towards the addictive risks of these drugs is suggested. PMID- 11274717 TI - Salmon calcitonin potentiates the analgesia induced by antidepressants. AB - Antidepressants are used in the treatment of a variety of pain syndromes. Most of them act by blocking noradrenaline (NA) and serotonin (5-HT) reuptake. It is also well known that the serotonergic system is also involved in calcitonin (CT) analgesia. Taking these two evidences into account, the modification of the analgesic effect of nortriptyline, amitriptyline, and paroxetine in the presence of salmon CT (s-CT) was examined in mice. The forced-swimming test was carried out in order to choose doses of each drug that did not induce an antidepressant effect under our experimental conditions (nortriptyline: 0.2-5 mg/kg ip, amitriptyline: 2.5-20 mg/kg ip, and paroxetine: 5-30 mg/kg ip). The analgesic effect of each antidepressant was then evaluated using the acetic acid test. At the doses tested, the antidepressants induced a dose-dependent analgesic effect. When mice were pre-treated with a subanalgesic dose of s-CT (2.5 IU/kg), the analgesic effect of amitriptyline and paroxetine was significantly increased though no modification was found for nortriptyline. In summary, s-CT was able to increase the analgesic effect of the antidepressant drugs that reduce the uptake of 5-HT, suggesting that the joint administration of antidepressants and CT may be an interesting alternative in pain management. PMID- 11274719 TI - Effects of the dopamine D2 agonist, quinpirole, on time and number processing in rats. AB - In Experiment 1, rats were trained to discriminate discrete sound sequences that varied in time or number. On time trials, the number of sounds was held constant at 4 and the duration of the sound sequence was either 2 or 8 s. On number trials, the duration of the sound sequence was held constant at 4 s, and the number of sounds was either 2 or 8. Psychophysical functions for time and number were obtained by presenting unreinforced sequences of intermediate duration or number. In agreement with previous research, sensitivity to variation in time was greater than variation in number. Rats received injections of the specific D2 agonist, quinpirole (0.08 mg/kg), or saline. Quinpirole significantly attenuated control by both time and number, but it did not increase behavioral estimates of time or number. In Experiment 2, rats were given different dosages of quinpirole (0.02, 0.04 or 0.06 mg/kg). The steepness of the psychophysical functions for both time and number was reduced in a dose-related fashion without any evidence of an increase in the estimation of time or number. The similarity of the effect of quinpirole on both time and number processing is consistent with the idea that the same internal mechanism is used for timing and counting. However, it is not consistent with the idea that D2 dopamine agonists selectively increase the rate of the internal clock, which is assumed to underlie performance in a temporal bisection procedure. Quinpirole (at doses between 0.02 and 0.08 mg/kg) reduces sensitivity to time and number in a bisection procedure without altering the speed of the internal clock. PMID- 11274718 TI - Nicotine-conditioned locomotor activity in rats: dopaminergic and GABAergic influences on conditioned expression. AB - Little is known about the processes that mediate acquisition and expression of conditioned associations between contextual cues and psychomotor effects of nicotine. In four separate experiments using rats, an environment repeatedly paired with nicotine acquired the ability to elicit increases in activity even in the absence of drug. This conditioned effect was sensitive to nicotine dose. Rats that had 0.6 or 1.2 mg/kg nicotine, but not 0.3 mg/kg, paired with the environment were more active than an unpaired control group (Experiment 1). In Experiment 2, control groups eliminated accounts based on nonspecific effects of nicotine and inhibitory conditioning decreasing activity in the unpaired controls of Experiment 1. Pretreatment on the test day with 100 mg/kg of gamma vinyl-GABA (GVG), a compound that inhibits the enzyme required to breakdown GABA, partially blocked the expression of locomotor conditioning without impairing activity in controls (Experiment 3). In Experiment 4, pretreatment on the test day with the dopamine D(1) receptor antagonist SCH-23390 (0.03 mg/kg) blocked expression of nicotine-conditioned locomotor activity; the D(2)/D(3) receptor antagonist eticlopride did not. Thus, the dopamine D(1) receptor subtype appears to play a role in context-elicited increases in activity conditioned by nicotine; GABA may also modulate the expression of this conditioned effect. PMID- 11274720 TI - Attenuation of morphine dependence and withdrawal by glycine B site antagonists in rats. AB - Numerous data indicate that noncompetitive and competitive N-methyl-D-aspartate (NMDA) receptor antagonists inhibit the development of physical dependence on opioids when these substances are administered together, and NMDA receptor antagonists are used at lower range of doses. Higher doses of these antagonists can enhance some opioid-induced effects. The present study extends these findings to the effects of NMDA/glycine (glycine(B)) site antagonists. Wistar rats were rendered dependent on morphine by implantation of morphine pellets. Both of the glycine(B) site antagonists used, 7-chloro-4-hydroxy-3-(3-phenoxy)-phenyl-2(H) quinolone (L-701,324; 2.5 and 5.0 mg/kg) and 5,7-dichlorokynurenic acid (5,7 DCKA; 25, 50, and 100 mg/kg), suppressed the expression of morphine withdrawal syndrome estimated as wet dog shakes. Furthermore, L-701,324 (2.5 and 5 mg/kg), given twice a day during the development of morphine dependence, attenuated the development of morphine dependence, and the results were comparable to those obtained after administration of noncompetitive NMDA receptor antagonist - MK801 (0.1 mg/kg). Our data suggest that glycine(B) site antagonists may attenuate wet dog shakes (withdrawal) and the development of dependence, both being induced by chronic morphine administration in rats. PMID- 11274721 TI - 5-HT 3 receptor antagonist ICS 205-930 alters the discriminative effects of ethanol. AB - The ability of a selective 5-hydroxytryptamine (5-HT(3)) receptor antagonist, ICS 205-930 (3-tropanyl-indole-1-carboxylate, tropisetron), to block the discriminative stimulus effects of ethanol was investigated in rats that were trained to discriminate ethanol (1.25 g/kg ip) from saline with food as the reinforcement. Prior administration of ICS 205-930, at the dose of 0.01 mg/kg, significantly decreased ethanol's discriminative stimulus effect at ED(75) dose of ethanol, while higher doses of ICS 205-930 (10 and 17 mg/kg) showed enhancement of ethanol's discriminative effects at ED(0), ED(25), and ED(50) doses of ethanol. Under conditions where ICS 205-930 (10, 17 mg/kg) was tested alone, rats responded exclusively on the saline-appropriate lever. These effects occurred without significantly altering response rates or blood ethanol concentrations. The results suggest that the 5-HT(3) antagonist ICS 205-930 at lower concentration decreases, and at higher concentration enhances the discriminative stimulus effects associated with a lower to moderate dose of ethanol. PMID- 11274722 TI - Novel monoamine transporter ligands reduce cocaine-induced enhancement of brain stimulation reward. AB - Six novel monoamine reuptake inhibitors were screened for their intrinsic effects on brain stimulation reward (BSR), as well as for their potential to reduce cocaine-induced reward-enhancement in that paradigm. Two of the compounds, nocaine-3B and 5-ara-74A (disubstituted piperidines) significantly reduced locus of rise (LOR), threshold measure of reward, at some doses. One compound, 1-RV-96A (a hybrid of the GBR and WIN-like agents) significantly reduced reward (increased LOR), but only at the highest dose tested. No effect of dose was found for MC9-20 (a GBR-like acyclic analogue of the N-bisarylmethoxyethyl-N'-phenylpropyl piperazine), nocaine-250B or 4-ara-42C (disubstituted piperidines). When cocaine (10 mg/kg, ip) and selected, hedonically neutral doses of novel compounds were combined, the following findings were obtained: MC9-20 (2.5 mg/kg, ip) showed a significant increase in cocaine-induced reward enhancement (0.2 log units or 53%). In contrast, nocaine-250B and 1-RV-96A (both at 10 mg/kg, ip) demonstrated a significant reduction (0.13 log units or 41%) in cocaine-induced reward enhancement (P<.01 and P<.05, respectively), as measured by changes in LOR. There were no differences in the maximum behavioral output (MAX) at either dose of each of the six drugs, or when selected doses were combined with cocaine. These results indicate that nocaine-250B and 1-RV-96A constitute two potential anticocaine compounds worthy of further behavioral and biochemical evaluation. PMID- 11274723 TI - Association of immune complexes and plasma viral load with CD4+ cell depletion, CD8+ DR+ and CD16+ cell counts in HIV+ hemophilia patients. Implications for the immunopathogenesis of HIV-induced CD4+ lymphocyte depletion. AB - OBJECTIVE: There is evidence that HIV induces CD4+ depletion in part by the formation of immune complexes (IC) that attach to CD4+ blood lymphocytes. In the present study we examined the relationship of IC-coated CD4+ blood cells with retroviral replication in HAART-treated patients. PATIENTS AND METHODS: 52 hemophilia patients were studied from 1997 to 1999. Lymphocyte subsets, IgM, IgG and gp120 on CD4+ blood cells, in vitro responses of lymphocytes to mitogens, plasma neopterin and plasma viral load were measured. RESULTS: Patients with detectable viral replication and without ICs on CD4+ blood lymphocytes had a lower viral load (4100 versus 21000 HIV-1 mRNA copies/ml; P = 0.079) and higher CD4+ cell counts (310/microl versus 161/microl; P = 0.035) than patients with ICs on circulating CD4+ lymphocytes. Among patients with < 80 HIV-1 mRNA copies/ml, IC- individuals had slightly higher CD4+ lymphocyte counts than IC+ patients (384/microl versus 316/microl; n.s.). Further evidence for the clinical relevance of the ICs was obtained when 18 patients who had an undetectable viral load at previous investigations were analyzed. Among patients with a stable undetectable viral load, CD4+ counts increased in 6 of 8 IC- but in none of 2 IC+ individuals. In patients whose viral load increased during the observation period, 5 of 6 IC- but none of 2 IC+ individuals showed higher CD4+ cell counts. Impaired virus killing is suggested by lower CD16+ (35/microl versus 107/microl; P = 0.016), higher CD3+ DR+ (178/microl versus 66/microl; P = 0.006), and higher CD8+ DR+ (142/microl versus 34/microl; P = 0.017) cell counts in IC(-) patients compared to IC- patients without detectable viral load. Strong retroviral replication induced strong T cell dysfunctions. Fewer CD3+ 25+ blood lymphocytes (19/microl versus 47/microl; P = 0.006) and a lower in vitro response of T lymphocytes to the mitogens Con A (RR: 0.3 versus 1.2; P=0.023) and CD3 mab (RR: 0.5 versus 2.4; P = 0.012) was observed in IC+ patients with detectable versus undetectable viral load. CONCLUSION: Our data suggest that ICs on circulating CD4+ blood lymphocytes are primarily associated with CD4+ lymphocyte depletion whereas the plasma viral load is primarily associated with decreased T lymphocyte activation, lower CD16+ counts, and higher CD8+ DR+ lymphocytes which might be the effector cells for virus elimination. PMID- 11274724 TI - The response of human dendritic cells to recombinant adenovirus, recombinant Mycobacterium bovis Bacillus Calmette Guerin and biolistic methods of antigen delivery: different induction of contact-dependant and soluble signals. AB - Dendritic cells (DCs) are the most potent antigen presenting cells for inducing T cell immune responses. The ability to grow human DCs from monocyte precursors provides an abundant source of these cells, which can be modified in vitro to present antigens. Re-administration of modified DCs to patients as vaccines has been shown in some cases to induce immune responses against cancer and infectious disease. Gene delivery to DCs provides an intracellular source of antigen for efficient and persistent loading of major histocompatibility complex (MHC) class I molecules. The aim of this study was to use monocyte-derived DCs (MD-DCs) from healthy donors to compare in vitro gene transfer, mediated by adenovirus, M. bovis Bacillus Calmette Guerin (BCG) and biolistic delivery. Efficiency of transfection and effect on DC phenotype, allostimulatory capacity and cytokine secretion was investigated. Adenovirus and BCG both showed a comparable ability to transfect MD-DCs, whereas the biolistic delivery by gene gun was unsuccessful in the reporter gene delivery. BCG transfection promoted MD-DC maturation as is apparent in the surface phenotype, allostimulatory capacity and cytokine secretion from cells. In comparison, adenovirus and biolistic delivery had a reduced effect on MD-DCs although enhancement of co-stimulatory and MHC molecule expression occurred in the cells of some donors. Both BCG and adenovirus represent useful vectors for gene transfer to human DCs. The effect of BCG on DC maturation may provide additional signals for the induction of antigen-specific T cell responses. PMID- 11274725 TI - Ontogenesis of protein kinase C betaII and its anchoring protein RACK1 in the maturation of alveolar macrophage functional responses. AB - Alveolar macrophages are the resident airway cells primarily responsible for the protection of the lungs against inhaled toxins and other biologically active materials. The purpose of this study was to investigate the maturation with age of alveolar macrophage functional responses. We characterised the ontogenesis of PKC betaII and its anchoring protein RACK1 in correlation with PKC-dependent immune functions, such as TNF-alpha, hydrogen peroxide production and lysozyme release in resident alveolar macrophages obtained from rats 2, 4 and 12 weeks old. Our results show an age-associated increase in the expression of PKC betaII and RACK1, which correlated with a maturation of alveolar macrophage functional responses. PMID- 11274727 TI - Analysis of the adjuvant effect of recombinant Leishmania infantum Hsp83 protein as a tool for vaccination. AB - The properties of Leishmania infantum hsp83 (LiHsp83) to elicit an immune response against a fused reporter antigen, maltose binding protein (MBP), was studied. CF1 mice were immunized with different purified recombinant proteins: MBP, LiHsp83 and MBP fused to LiHsp83 (MBP-LiHsp83). Serum samples were obtained at days 0, 21, 28, 60, 90, 120 and 150 post-immunization. MBP-LiHsp83 fusion protein elicited a strong humoral response against MBP, higher than that one obtained in mice immunized with MBP alone or MBP mixed with LiHsp83, showing the secretion of both anti-MBP IgG2a and IgG1 isotypes (IgG2a/IgG1 ratio: 2:1). This response was specific for recombinant proteins and was maintained for at least 150 days, whereas the reactivity in mice immunized with MBP alone dissapeared at day 90. After in vitro stimulation with MBP, spleen cells from MBP-LiHsp83 immunized mice showed higher proliferation indices and produced higher secretion of IFN-gamma than spleen cells from either control or MBP-immunized mice. In all groups of mice IL-4 was undetectable. Thus we consider that LiHsp83 may be a promising candidate to be used as carrier of fused antigens for adjuvant-free vaccination. PMID- 11274726 TI - CD40 expression is induced by the introduction of IgM receptor on the surface of pro-B cell line NFS70. AB - To examine the molecular mechanism of B cell differentiation, we introduced rearranged immunoglobulin (Ig) mu- and kappa-chain genes into the NFS70 pro-B cell line and observed their maturation. The IgR(+)-transfectants had characteristics of mature surface IgM (sIgM)+ B220high CD40+ CD38+ CD25+ B cells. CD40 expression levels were regulated by stimulation via the IgR. In comparison to wild type NFS70 cells, NF-kappaB activity was up-regulated in the IgR transfectants. Anti-IgR crosslinking of IgR+ cells induced down-regulation of CD40 expression that correlated with down-regulation of NF-kappaB activity in the IgR(+)-transfectants. Immature CD19+ sIgD- B cells from bone marrow also showed an alteration of CD40 expression in response to anti-IgR crosslinking. The results suggest that expression of IgR on the surface is one of the factors responsible for further maturation of B cells. PMID- 11274729 TI - Frequency of CCR5 gene 32-basepair deletion in Chilean HIV-1 infected and non infected individuals. AB - A 32-basepair deletion polymorphism in the CCR5 chemokine receptor gene (CCR5Delta32) has been identified and shown to have functional significance in determining susceptibility to infection by human immunodeficiency virus type 1 (HIV-1) and possibly in influencing disease progression in HIV-1 positive individuals. These findings led to an interest in studies of DeltaCCR5 allele geographical distribution in human population, for complete understanding of the role of CCR5 in HIV-1 epidemiology. Inter-population variation in CCR5Delta32 frequency may be a significant factor in the prediction of AIDS endemicity. In this report we assessed the frequency of DeltaCCR5 in a Chilean population (63 HIV-1 infected and 62 non-infected individuals). No homozygous CCR5Delta32 individual was identified, and no significant difference was observed between HIV 1 infected (3/63) and non-infected (3/62) individuals for the heterozygote CCR5Delta32 state. This is the first evidence of the contribution of DeltaCCR5 allele to the genetic background of the Chilean population, which is characterized by intense ethnic admixture and by gene flow from the European Spanish gene pool. PMID- 11274728 TI - Constitutive expression of MCP-1 and RANTES in the human histiocytic lymphoma cell line U-937. PMID- 11274730 TI - Modulation of C3 gene expression in HepG2 human hepatoma cells. AB - Inflammation elicits an acute phase response, which includes changes in plasma concentrations of a number of cytokines, reflecting changes in their gene transcription in the liver. In this study, the induction of complement factor 3 (C3) was investigated in HepG2 cells, a human hepatoma cell line often used as a model system for cytokine-dependent expression of acute phase proteins of the liver. By using a very sensitive RT-PCR assay, the amount of mRNA for C3 was measured after induction with lipopolysaccharide (LPS) and interleukin-6 (IL-6). Both substances were found to up-regulate C3 gene expression. C3 mRNA level was lower in LPS-treated cells compared to IL-6 induction and also reached maximum expression at an earlier time point. These findings suggest a coordinate stimulation of C3 expression in the hepatocytes, which then maintains the host response to infectious agents. PMID- 11274731 TI - Tolerance to lipopolysaccharide (LPS) regulates the endotoxin effects on Shiga toxin-2 lethality. AB - It has been suggested that Shiga toxin (Stx) is necessary but not sufficient for hemolytic uremic syndrome (HUS) development, and pro-inflammatory stimuli such as lipopolysaccharide (LPS) from Gram negative bacteria are needed. Taking into account that LPS is present in the natural infection during HUS development, detoxification or regulation of LPS activity could be crucial to define the course of the disease. The objective of the present study was to investigate whether tolerance to LPS and/or antibodies to LPS, are able to modify the LPS induced modulation of Stx type-2 (Stx2) lethality in a mouse model. Our results demonstrate that the high levels of IgG anti-LPS antibodies in immunized mice did not modify the dual effects of LPS (enhancement or protection) on Stx2 action. This could be attributed to the fact that antibodies do not recognize the active portion of LPS molecule (lipid A). However, the enhancement of Stx2 toxicity exerted by LPS was inhibited in tolerant mice. This effect could be ascribed to the inhibition of LPS-induced TNF-alpha and IL-1beta secretion in tolerant animals, two cytokines known to be involved in the overexpression of Stx receptors. The phenomenon of LPS-induced protection on Stx2 toxicity was also inhibited in tolerant animals, although the mechanism involved in this effect is not clear. This is the first description which shows the influence of endotoxin tolerance on the evolution of experimental HUS. However, like in Gram negative infections, further knowledge on tolerance mechanism is necessary in order to achieve a comprehensive view of this phenomenon. PMID- 11274732 TI - A pivotal role of cysteine 3 of Lck tyrosine kinase for localization to glycolipid-enriched microdomains and T cell activation. AB - Lck, a Src family protein tyrosine kinase (PTKs), is post-translationally modified by palmitoylation, a process thought to regulate the biological function, membrane affinity and glycolipid-enriched microdomain (GEM) localization of this molecule. To examine the importance of palmitoylation sites Cys3 and Cys5 in Lck, one or both of these residues was mutated to serine to create mutants S3, S5, and S3,5, respectively. Immunofluorescence and confocal microscopy of COS-7 cells transfected with these constructs showed that while S5 and S3 localized to the plasma membrane, S3,5 was localized to the cytoplasm, suggesting that palmitoylation at at least one site is essential for membrane localization. Sucrose gradient based fractionation of these mutants expressed in COS-7 cells showed that while S5 localized to GEMs in similar fashion to the wild type, GEM localization of S3 was severely inhibited. Expression of these mutants in Lck-negative JCaM1 cells showed that although S5 reconstituted activation of nuclear factor NFAT as per the wild type, S3 expression failed to do so. These results suggest that Cys3 of Lck plays a more important role than Cys5 in GEM localization and T cell activation. Additionally, it was found that the degree of T cell function recovery is positively correlated with the degree of Lck expression in GEMs. PMID- 11274733 TI - Leukocyte-endothelial adhesion is impaired in the cremaster muscle microcirculation of the copper-deficient rat. AB - Dietary copper deficiency impairs the function of both the vascular endothelium and circulating leukocytes. In the current study, leukocyte-endothelium adhesion was observed in the in vivo cremaster muscle microcirculation of copper-adequate and copper-deficient rats. Male, weanling Sprague-Dawley rats were fed purified diets that were either adequate (5.6 microg/g) or deficient (0.3 microg/g) in copper. Adhesion was stimulated with the inflammatory mediators tumor necrosis factor-alpha and bradykinin, and the chemotactic peptide N-formyl-methionyl leucyl-phenylalanine. Intravascular adhesion of leukocytes to the vascular endothelium was significantly attenuated in the copper-deficient group in response to all three agonists. These results occurred without any difference in intravascular wall shear rate between the dietary groups. Based on previous work, we propose that the attenuated response is caused by either decreased expression of adhesion molecules on leukocytes and endothelial cells or by inhibition of the endothelial cell calcium signaling associated with copper deficiency. PMID- 11274735 TI - Current topics in comparative developmental biology of vertebrate brains. AB - Little is known about how unique features in a species can emerge along the conserved body plan beyond species. For example, mammals, including human beings, have acquired the neocortex with distinct function and morphology. Here we review current topics in comparative developmental biology of vertebrate brains, especially focusing on the cerebral neocortex as a suitable model for considering species-specific aspects. PMID- 11274736 TI - Ultrastructural localization of brain-derived neurotrophic factor in rat primary sensory neurons. AB - In a previous study we have shown that brain-derived neurotrophic factor (BDNF) is present in a subpopulation of small- to medium-sized sensory neurons in the dorsal root ganglia (DRG) and is anterogradely transported in both the peripheral and central processes. Within the spinal cord, BDNF is localized to varicosities of sensory nerve terminals in laminae I and II of the dorsal horn. This study raised the question of whether BDNF is localized in synaptic vesicles of the afferent nerve terminals. Using immunohistochemical and immunocytochemical techniques we have now investigated the ultrastructural localization of BDNF in the spinal cord of the rat. In addition, its colocalization with the low affinity neurotrophin receptor, p75, and calcitonin gene related peptide (CGRP) was also investigated. In lamina II of the spinal cord, BDNF immunoreactivity was restricted to nerve terminals. The reaction product appeared associated with dense-cored and clear vesicles of terminals superficial laminae. Double labelling experiments at the light microscopic level showed that 55% of BDNF immunoreactive neurons in DRG are colocalized with CGRP and many nerve terminals in laminae I and II of the spinal cord contained both BDNF and CGRP immunoreactivities. The results of double labelling at the ultrastructural level showed that most BDNF-ir (immunoreactive) nerve terminals contained CGRP or the low affinity neurotrophin receptor, p75, but not vice versa. These results point to the conclusion that BDNF may be released in parallel with neurotransmitters from nerve terminals in the spinal cord from a subpopulation of nociceptive primary afferents. PMID- 11274737 TI - Non-opioid actions of lamotrigine within the rat dorsal horn after inflammation and neuropathic nerve damage. AB - Some opioid-resistant pain conditions can be alleviated by voltage-dependent Na(+) channel blockers such as lamotrigine. The mu-opioid-receptor agonist morphine can modulate cation entry into cells to affect overall cellular excitability, an effect which can in turn be endogenously antagonised by the neuropeptide cholecystokinin (CCK). However, lamotrigine may also modulate cellular excitability by non-specifically blocking voltage-dependent ion channels. We have looked for interactions of lamotrigine with the opioid/CCK pathway within the spinal dorsal horn, to rule out the possibility that lamotrigine may attenuate nociceptive responses via actions on this pathway. Both lamotrigine and the mu-opioid agonist DAMGO inhibited mustard oil-evoked cell firing by approximately 50% compared with control levels. Co-application of CCK8S reversed DAMGO-, but not lamotrigine-induced inhibition of cell firing and this reversal was prevented with the selective CCK(B) receptor antagonist PD 135158. Although lamotrigine inhibited both brush- and cold-evoked cell firing in neuropathic animals, lamotrigine inhibition of mustard oil-evoked cell firing in the same animals was not significantly greater than that observed in controls. These results suggest that the antinociceptive properties of lamotrigine within the spinal dorsal horn are unlikely to be mediated via interactions with the opioid/CCK pathway. PMID- 11274738 TI - Expression of JNK cascade scaffold protein JSAP1 in the mouse nervous system. AB - The mitogen-activated protein kinase (MAPK) cascades consist of MAPK, MAPK kinase (MAPKK), and MAPKK kinase (MAPKKK). The specificity of activation of MAPK cascades may be determined, in part, by scaffold proteins that organize multi enzyme complexes. We have earlier reported a scaffold protein JSAP1 (also known as JIP3) in the JNK MAPK cascade. We also showed that, of the adult mouse tissues tested, JSAP1 mRNA was predominantly expressed in brain. Here we report the localization of JSAP1 protein in mouse embryos and adult brain by immunohistochemical analysis. In embryos (E11-16), JSAP1 immunoreactivity was mainly found in the central and peripheral nervous systems, where it was localized to the cell bodies and/or axons of developing neurons, but not neural precursor cells. In the adult brain, immunoreactive JSAP1 was localized mostly to cell bodies in almost all neurons. We also showed that the expression of JSAP1 transcripts and proteins gradually increased during the neural differentiation of mouse P19 embryonal carcinoma (EC) cells. Furthermore, we showed that overexpressed JSAP1 facilitated the efficient activation of JNK by MEKK1 in P19 cells. These results suggest that JSAP1 may function as a scaffold protein for the JNK signaling module in neuronal cells. PMID- 11274739 TI - Oligodendroglial cell death with DNA fragmentation in the white matter under chronic cerebral hypoperfusion: comparison between normotensive and spontaneously hypertensive rats. AB - We investigated the neuropathological and biochemical changes in the white matter of normotensive Wistar Kyoto (WKY) and spontaneously hypertensive rats (SHR) after bilateral carotid artery ligation (BCAL). One week after BCAL, both WKY and SHR showed white matter rarefaction and vacuolation with reduced oligodendrocytes, but there was no difference between WKY and SHR. On the other hand, vacuoles formed by oligodendroglial cell death were increased significantly from 2 to 4 weeks in the optic tract and fimbria fornix of hypoperfused SHR. Furthermore, terminal deoxynucleotidyl transferase-mediated dUTP in situ nick end labeling (TUNEL)-positive cells and lectin-positive microglia increased in number and intensities of staining more markedly in SHR than in WKY. In situ cell death detection ELISA supported these results quantitatively. RT-PCR represented the expression of TNF-alpha, TNF receptor 1 (p55), caspase-2 (Ich-1) and -3 (CPP32) mRNAs in both WKY and SHR brains after BCAL. Immunohistochemical analyses revealed that TNF-alpha, TNF receptor 1 (p55), Ich-1 and CPP32 immunoreactive cells could also be detected in the white matter regions of hypoperfused SHR. These results suggested that local production of TNF-alpha by the activated microglia might selectively induce oligodendroglial cell death through the death domain-containing TNF receptor 1 (p55), caspase-2 or -3 activation, resulting in white matter changes as a primary pathological feature. PMID- 11274740 TI - Tetrodotoxin-resistant conductivity and spinal effects of cutaneous C-fibre afferents in the rat. AB - The effect of the sodium channel blocking agent tetrodotoxin (TTX) on signal processing in afferent fibres of dorsal roots was tested in Sprague-Dawley rats. TTX applied to the dorsal roots L4-L6 blocked the fast afferent volleys from the sural nerve, which was stimulated electrically with supramaximal strength for A fibres. Afferent C-fibre compound action potentials (CAPs) elicited by electrical stimulation of the dorsal root L5 peripherally from the TTX block or by electrical stimulation of the sural nerve likewise disappeared from the recording. Cord dorsum potentials (CDPs) recorded at the dorsal root entry zone of L4 were blocked completely if elicited by A-fibre volleys. In contrast, CDPs elicited by C-fibre stimulation persisted with longer latency and reduced amplitude in the first part of the CDP. During TTX block, C-fibre potentials could also be recorded from dorsal root filaments after stimulation of the sural nerve or the dorsal root L5 peripherally of the TTX-block. The results suggest that in the axonal membrane of cutaneous C-afferents, both TTX sensitive and TTX resistant voltage gated sodium channels exist, the latter being responsible for the propagation of signals in a portion of C-fibres after TTX application. The TTX-resistant portion of the afferent potential does not seem to contribute much to the afferent C-fibre CAP before TTX application, but its central effects seem to be overproportionally strong. PMID- 11274741 TI - Anterior cingulate activity during pain-avoidance and reward tasks in monkeys. AB - We recorded single neuronal activities in the anterior cingulate cortex of monkeys while they were performing discriminative pain-avoidance and reward tasks: a prediction cue was presented for 0.5-1.5 s, followed by a red or green discrimination cue (1:1, random) for 1.0 s; painful stimuli were presented if the monkey failed to respond during the red cue; a reward was given, subsequent to a cued 1 s delay, if the monkey responded after the green cue. Among 775 neurons recorded, 196 neurons showed significant activity during one or more observation periods; 36 during the prediction period; 77 during the discrimination period; 41 during the delay period; 85 during the response period; 40 during the pre-reward period; and 15 during the reward-ingestion period. Of 77 neurons activated during the discrimination period, 47 showed exclusive activity either during the red (34) or during the green (13) cue: of 85 neurons activated during the response period, 64 showed exclusive activity either for pain-avoidance (37) or obtaining a reward (27). Control experiments confirmed that the neuronal activity could not be attributed to simple visual or motor processes. The results suggest that some anterior cingulate neurons are involved with anticipation of, and response selection for, imminent events. PMID- 11274742 TI - Imaging stochastic spatial variability of active channel clusters during excitation of single neurons. AB - Topographical maps of membrane voltages were obtained during action potentials by imaging, at 1 microm resolution, live dissociated neurons stained with the voltage sensitive dye RH237. We demonstrate with a theoretical approach that the spatial patterns in the images result from the distribution of net positive charges condensed in the inner sites of the membrane where clusters of open ionic channels are located. We observed that, in our biological images, this spatial distribution of open channels varies randomly from trial to trial while the action potentials recorded by the microelectrode display similar amplitudes and time-courses. The random differences in size and intensity of the spatial patterns in the images are best evidenced when the time of observation coincides with the duration of single action potentials. This spatial variability is explained by the fact that only part of the channel population generates an action potential and that different channels open in turn in different trials due to their stochastic operation. Such spatial flicker modifies the direction of lateral current along the neuronal membrane and may have important consequences on the intrinsic processing capabilities of the neuron. PMID- 11274743 TI - Site-specific calcium-dependent proteolysis of neuronal protein GAP-43. AB - GAP-43 is a presynaptic protein participating in signal transduction processes in nerve terminals. GAP-43 exists in neurons along with two truncated forms devoid of 4 and 40 N-terminal residues. In this report, we show that these forms of GAP 43 are proteolytic fragments derived from calcium-dependent cleavage of GAP-43 molecule at 5th and 41st residues. GAP-43 site-specific proteolysis in synaptosome and cytosol fractions proved to be dependent on the addition of millimolar amounts of calcium. This fact together with inhibition of GAP-43 proteolysis by calpain inhibitors as well as local composition of the cleavage sites indicates to the participation of calpain in this process. The proteolysis disturbs some properties characteristic for whole GAP-43 molecules, in particular, calmodulin binding and Ser-41 phosphorylation, when the cleavage occurs at 41st residue. Some other GAP-43 properties (G(o) protein activation and membrane attachment) are retained by separate fragments. Therefore, calcium controlled site-specific proteolysis of GAP-43 can be of great physiological significance. PMID- 11274744 TI - Expression of neuropsin in oligodendrocytes after injury to the CNS. AB - Proteases are involved in a variety of processes including demyelination after injury to the central nervous system. Neuropsin is a serine protease, which is constitutively expressed in the neurons of the limbic system. In the present study, intrahippocampal kainate injection and enucleation were performed on adult mice. Neuropsin mRNA and protein expression was detected by in situ hybridization and immunohistochemistry. Double in situ hybridization confirmed that the mRNA expression was induced in oligodendrocytes. One day after kainate injection to the hippocampus, neuropsin mRNA was expressed, peaking 4-8 days postoperatively and disappearing at 14 days. Immunohistochemistry and immunoelectron microscopy revealed that neuropsin was expressed in the cell body of oligodendrocytes and myelin. To see if neuropsin degrades myelin protein, purified myelin was incubated with recombinant neuropsin. A decrease in the intensity of the bands of myelin basic protein was observed. These results indicate that neuropsin is involved in demyelination. PMID- 11274745 TI - Long-term enhancement of synaptic transmission induced by veratridine in rat CA3 hippocampal neurons. AB - Veratridine is a neurotoxin that induces persistent activation of sodium channels in excitable cells. We investigated the effects of this toxin on excitatory synaptic transmission in CA3 neurons of juvenile rat hippocampus using whole-cell patch-clamp and field-potential recordings. The population spikes evoked by electrical stimulation of the mossy fiber were gradually enhanced after washout of veratridine (0.3 microM), but they were not enhanced by the co-application of veratridine and an N-methyl-D-aspartate (NMDA) receptor antagonist (D-APV, 30 microM). When a pipette solution contained QX-314 that antagonized the effect of veratridine in the recorded neuron, oscillatory membrane depolarization appeared in the early stage during bath-application of veratridine and gradually decreased in the late stage. After washout of veratridine, however, the oscillatory depolarization was gradually restored and maintained for at least 3 h. This oscillatory depolarization was also abolished by D-APV. We suggest that the activation of NMDA receptors is involved in the veratridine-induced long-lasting enhancement in the excitatory synaptic transmission in rat CA3 hippocampal neurons. PMID- 11274746 TI - Attentional modulation of neural activity in the macaque inferior temporal cortex during global and local processing. AB - To examine whether visual attention to global and local features of visual stimuli modulates neural activity in the monkey visual cortex, we applied positron emission tomography techniques to monkeys while they were discriminating either global or local features of visual stimuli. The posterior inferior temporal cortex was more activated in discriminating global features than in discriminating local ones, whereas the anterior inferior temporal cortex was more activated in discriminating local features than in discriminating global ones. The results suggest that a functional difference exists in terms of processing of global and local features within the inferior temporal cortex. PMID- 11274747 TI - Marked hypotensive and blood flow-increasing effects of a new lipo-PGE(1) (lipo AS013) due to vascular wall targeting. AB - Lipo-AS013 is being developed as an improved formulation of lipo-PGE(1), which is widely used in clinical practice in Japan and some Asian countries. We have previously reported that lipo-AS013, which is a lipid microsphere (LM) preparation of a chemically stable and lipophilic PGE(1) prodrug (AS013, Fig. 1), slowly releases small amounts of the active ingredient (AS013) in human plasma. In the present study, to estimate the vascular wall targeting ability and efficacy of lipo-AS013, we determined the hypotensive and blood flow-increasing effects of lipo-AS013, lipo-PGE(1), PGE(1)CD, and AS013. Lipo-AS013 was found to have longer-lasting hypotensive and blood flow-increasing effects than the other agents. The two LM preparations, lipo-PGE(1) and lipo-AS013, had a markedly stronger effect than PGE(1)CD and AS013 alone, demonstrating the benefit of drug delivery using LM. In spontaneously hypertensive rats (SHR), lipo-AS013 also had a significant hypotensive effect. To confirm vascular wall targeting by lipo AS013, the localization of PGE(1) in the aorta and neovascular capillaries of rat was investigated by immunostaining. The results indicated that lipo-AS013 was more efficient at delivering the active ingredient (AS013) to the vessel wall. In conclusion, lipo-AS013 could supersede lipo-PGE(1) and PGE(1)CD in clinical use. PMID- 11274748 TI - In vitro degradation and release profiles for poly-dl-lactide-poly(ethylene glycol) microspheres containing human serum albumin. AB - Poly-dl-lactide-poly(ethylene glycol) (PELA) block copolymers containing same the content (10%) of polyethylene glycol (PEG) were synthesized with five different molecular weight of PEG by ring-opening polymerization. PELA microspheres containing human serum albumin (HSA) were elaborated by solvent extraction method based on the formation of double w/o/w emulsion. In vitro matrix degradation and protein release of these microspheres were performed in phosphate-buffered saline (PBS) (154 mM, pH 7.43). The degradation profiles were characterized by measuring the loss of microspheres mass, the decrease of polymer intrinsic viscosity, the decrease of pH value of degradation medium, the reduction of polymer number average molecular weight (M(n)) and the change of molecular weight polydispersity (M(w)/M(n)). The release profiles were investigated from the measurement of protein presented in the release medium at various intervals. It showed that the matrix degradation and protein release profiles were highly polymer-dependent. The extent of burst release in the initial protein release increased with the decrease of molecular weight of PELA copolymer. It is suggested that these matrix polymers may be optimized as carriers in protein (antigen) delivery system for different purposes. PMID- 11274749 TI - Evaluation of an intestinal pressure-controlled colon delivery capsules prepared by a dipping method. AB - A new method for preparation of large amounts of empty pressure-controlled colon delivery capsules (PCDCs) by a dipping method has been developed. Empty PCDCs are composed of two polymer membranes. The inner one was a water-insoluble polymer membrane, ethylcellulose (EC). The outer one was an enteric polymer membrane, hydroxypropylmethylcellulose phthalate (HPMCP) or hydroxypropylmethylcellulose acetate succinate (HPMCAS). By consequently dipping into an ethanolic EC solution and an alkalized enteric polymer solution, empty PCDCs were obtained after both the capsule body and cap were adjusted to the size of #2 capsules. With each enteric polymer, two types of empty PCDCs of different thickness were prepared. Fluorescein (FL) was formulated with suppository base, PEG1000, and used as a model drug. FL/PEG1000 suspension was introduced into empty PCDCs which were then sealed with enteric polymer solution. The PCDCs were evaluated by an in vivo experiment using beagle dogs. After oral administration of the test PCDC preparations containing 30 mg of FL, blood samples were obtained from the jugular vein and serum FL levels were measured. The thickness of the EC membrane layer varied in both the capsule body and cap. HPMCAS PCDCs had 62.1+/-5.0 (S.E.) microm (body) and 49.7+/-3.3 microm (cap) with thicker ones and 55.7+/-6.6 microm (body) and 46.8+/-6.2 microm (cap) with thinner ones. HPMCP PCDCs had 28.1+/-3.3 microm (body), 30.9+/-1.0 microm (cap) with thinner ones and 43.1+/-9.8 microm (body), 42.4+/-8.2 microm (cap) with thicker ones. The mean T(i) values, the first appearance time, of FL in the serum of HPMCAS PCDCs were 2.0+/-0.7 h for thicker ones and 3.8+/-0.5 h for thinner ones, while the mean T(i) values of HPMCP PCDCs were 2.0+/-0.0 h for thinner ones and 3.5+/-0.7 h for thicker ones. Since the colon arrival time in beagle dogs was 3.5+/-0.3 h as determined by a sulfasalazine test, thinner HPMCAS PCDCs and thicker HPMCP PCDCs were thought to deliver FL to the colon. PMID- 11274750 TI - Preparation of controlled release systems by free-radical UV polymerizations in the presence of a drug. AB - UV free-radical polymerization techniques are often used to synthesize hydrogels for controlled release applications. Numerous techniques exist for immobilizing drugs or solutes in the gel. This work focuses on the entrapment of solute in a hydrogel by conducting a photopolymerization in the presence of the monomer and the solute. A kinetic gelation model has been developed to examine the effect of the solute material on the polymerization process and the ensuing network structure. Kinetic experiments have also been conducted of the polymerization of poly(ethylene glycol) methacrylate in the presence of theophylline. It was found that the presence of the solute led to a more heterogeneous network with numerous microgel regions present. The effect of the size of the solute on the polymerization was also investigated. PMID- 11274751 TI - Regional drug delivery with radiation for the treatment of Ewing's sarcoma. In vitro development of a taxol release system. AB - Recently, several studies have suggested the radiosensitizing effect of taxol, a microtubular inhibitor. Our overall hypothesis is that a combination of radiation and taxol may demonstrate therapeutic efficacy over doses of either individually. Studies examining taxol use have mostly focused on systemic administration, which can lead to undesired effects. To circumvent these side effects, we propose a locally administered polymeric microsphere delivery system combined with radiation therapy for the treatment of Ewing's sarcoma. The present study focuses on the in vitro ability of taxol when present as a microencapsulated drug delivery system, and delivered locally at the site of the sarcoma/tumor, to block cells in the G2/M phase of the cell cycle and potentially enhance the radiation sensitivity of cells. Using the bioresorbable poly(anhydride-co-imide), poly[pyromellityl-imidoalanine-1,6-bis(carboxy-phenoxy)hexane] (PMA-CPH), and the radiosensitizing agent taxol, a microsphere based delivery system was fabricated. A solvent evaporation technique was used to encapsulate taxol at doses of 1%, 5%, and 10% in PMA-CPH microspheres. Release kinetics studies demonstrated that the total amount of taxol released and the release rate were directly dependent on loading percentage. Taxol's bioactivity and radiosensitizing ability were measured using flow cytometry. Co-culture of Ewing's sarcoma cells with and without taxol-loaded microspheres demonstrated that released taxol retained its bioactivity and effectively blocked cells in the radiosensitive G2/M phase of mitosis. The taxol-radiation delivery system studied achieved an 83% decrease in tumor cell count compared to control. Taxol effectively sensitized Ewing's sarcoma cells to radiation with radiosensitivity shown to be independent of radiation dose at levels of dosages studied. This work has demonstrated that taxol can be effectively released from a biodegradable PMA-CPH microsphere delivery system while maintaining potent combined cytotoxic and radiosensitizing abilities. PMID- 11274752 TI - PEGylated PLGA nanoparticles as protein carriers: synthesis, preparation and biodistribution in rats. AB - The aim of the present work was to assess the merits of PEGylated poly(lactic-co glycolic acid) (PEG-PLGA) nanoparticles as protein and peptide drugs (PPD) carriers. PEG-PLGA copolymer, which could be used to prepare the stealth nanoparticles or long-circulating nanoparticles, was synthesized with methoxypolyethyleneglycol (MePEG) and PLGA. The structure of PEG-PLGA was confirmed with (1)H NMR and Fourier transform infrared (FTIR) spectrum, and molecular weight was determined by gel permeation chromatography (GPC). Bovine serum albumin (BSA), chosen as model protein, was encapsulated within the stealth nanoparticles with the double emulsion method. The particles were characterized in terms of size, zeta potential and in vitro release of the protein. The biological fate of the BSA-loaded nanoparticles following intravenous administration was determined over 24 h in rats. The experimental results showed that PEG-PLGA could be obtained by ring-opening polymerization of lactide and glycolide in the presence of MePEG. (1)H NMR and FTIR spectrum were consistent with the structure of PEG-PLGA copolymer. Molecular weight determined by GPC was 50800. The stealth nanoparticles loading BSA could be prepared by the double emulsion technique. The entrapment efficiency was 48.6%, particle size about 200 nm and zeta potential -16.1 mV. BSA release from the stealth nanoparticles showed an initial burst release and then sustained release. PEG-PLGA nanoparticles could extend half-life of BSA from 13.6 min of loaded in PLGA nanoparticles to 4.5 h and obviously change the protein biodistribution in rats compared with that of PLGA nanoparticles. Thus, PEG-PLGA nanoparticles could be an effective carrier for PPD delivery. PMID- 11274754 TI - Mechanism of action of benzene toxicity: cell cycle suppression in hemopoietic progenitor cells (CFU-GM). AB - The aim of this study was to clarify previously reported controversial data and hypotheses concerning the effect of benzene on the cell cycle of hemopoietic stem cells. In this study, the bromodeoxyuridine UV (BUUV) suicide assay was performed in normal C57BL/6 and p53 knockout (KO) C57BL/6 mice during and after exposure to 300 ppm of benzene for 2 weeks. Our kinetic studies revealed that the cell cycle of hemopoietic myeloid progenitor cells (colony-forming unit granulocyte macrophage [CFU-GM]), rather than being stimulated, was suppressed by exposure to benzene. The fraction of CFU-GM in S phase was significantly depressed, from 37.1% in controls to 16.3% in normal mice. BrdUrd incorporation in both groups revealed significantly different slopes for untreated and benzene-exposed normal C57BL/6 mice. p53 appeared to induce suppression of both the number and the cycling fraction of hemopoietic progenitor cells, as demonstrated by the lack of benzene-induced suppression of these parameters in p53 KO mice. The likelihood that suppression of bone marrow cellularity and cell cycling is mediated by p53 was supported by the upregulation of p21, a cyclin-dependent kinase inhibitor. Our present study revealed the mechanism of action of benzene hematotoxicity. Benzene suppresses the cell cycle by p53-mediated overexpression of p21, a cyclin dependent kinase inhibitor, resulting not simply in suppression of hemopoiesis but rather in a dynamic change of hemopoiesis during and after benzene exposure. Thus, the controversies raised by previously reported data are resolved by our present findings of hemopoietic stem cell kinetics. PMID- 11274753 TI - Acute graft-vs-host disease: pathobiology and management. AB - Acute graft-vs-host disease (GVHD) is a major obstacle to safe allogeneic hematopoietic stem cell transplantation (HSCT), leading to a significant morbidity and mortality. GVHD occurs when transplanted donor T lymphocytes react to foreign host cells. It causes a wide variety of host tissue injuries. This review focuses on the pathobiological basis, clinical aspects, and current management strategies of acute GVHD. Afferent phase of acute GVHD starts with myeloablative conditioning, i.e., before the infusion of the graft. Total-body irradiation (TBI) or high-dose chemotherapy regimens cause extensive damage and activation in host tissues, which release inflammatory cytokines and enhance recipient major histocompatibility complex (MHC) antigens. Recognition of the foreign host antigens by donor T cells and activation, stimulation, and proliferation of T cells is crucial in the afferent phase. Effector phase of acute GVHD results in direct and indirect damage to host cells. The skin, gastrointestinal tract, and liver are major target organs of acute GVHD. Combination drug prophylaxis in GVHD is essential in all patients undergoing allogeneic HSCT. Steroids have remained the standard for the treatment of acute GVHD. Several clinical trials have evaluated monoclonal antibodies or receptor antagonist therapy for steroid-resistant acute GVHD, with different successes in a variety of settings. There are some newer promising agents like mycophenolate mofetil, glutamic acid-lysine-alanine-tyrosine (GLAT), rapamycin, and trimetrexate currently entering in the clinical studies, and other agents are in development. Future experimental and clinical studies on GVHD will shed further light on the better understanding of the disease pathobiology and generate the tools to treat malignant disorders with allogeneic HSCT with specific graft-vs tumor effects devoid of GVHD. PMID- 11274755 TI - Latent hematopoietic stem cell toxicity associated with protracted drug administration. AB - OBJECTIVE: The protracted administration of near-conventional daily doses of chemotherapeutic agents is a strategy to increase dose intensity and, potentially, efficacy as well. However, protracted therapy carries the risk of damage to stem cells in proliferative tissues that are not targeted by intermittent schedules. Therefore, we have investigated the effects produced by the protracted administration of two anticancer drugs on hematopoietic stem cell function. MATERIALS AND METHODS: We used the competitive repopulating assay to assess stem cell damage caused by protracted daily drug treatment of mice. RESULTS: Treatment with acetyldinaline for 10 consecutive days mediated a modest effect on the short-term repopulating cells (STRCs) but spared the long-term repopulating cells (LTRCs). Gemcitabine for 10 days led to a modest decline in both the STRCs and LTRCs. Extending treatment with gemcitabine for 28 days resulted in more severe repopulating cell (RC) damage, which was much worse than in acetyldinaline-treated mice. As expected, melphalan for 10 or 28 days mediated a marked reduction in all of the RCs of treated mice. The analysis of the RCs from mice that were allowed a 1-year recovery period after completing the 28-day treatment with either acetyldinaline or gemcitabine showed normal levels of neutrophils and bone marrow (BM) progenitors. However, a reduction in the RCs was observed in both groups, with larger reductions in gemcitabine-treated mice. CONCLUSIONS: Our data show that protracted treatment with gemcitabine, but not acetyldinaline, of mice caused severe permanent damage to the stem cell components. Therefore, although 28-day therapy with acetyldinaline or gemcitabine appeared to be well tolerated at the level of peripheral blood and bone marrow progenitors, gemcitabine produces permanent stem cell damage when used in long term administration regimens that should perhaps only be explored clinically with stem cell support available. PMID- 11274756 TI - Improvement of mouse beta-thalassemia by electrotransfer of erythropoietin cDNA. AB - OBJECTIVE: A new intramuscular DNA electrotransfer method for erythropoietin (EPO) expression was evaluated in the natural mouse model of human beta thalassemia (Hbb-thal1) in terms of its ability to reverse the anemia and improve the thalassemic features of erythrocytes. MATERIALS AND METHODS: Intramuscular injection of small amounts of a plasmid encoding mouse EPO, immediately followed by controlled electric pulses, was used. RESULTS: This procedure induced very high hematocrit levels in beta-thalassemic mice compared to nonelectrotransferred mice. The hematocrit increase was dose dependent, still increased 4 months after injection of plasmid DNA, and associated with a high transgenic EPO blood level in all mice (up to 2500 mU/mL of plasma). EPO gene electrotransfer not only led to a long-lasting and dose-dependent increase in the hematocrit but also to a 100% increase in the lifespan of erythrocytes of thalassemic mice. This was related to a nearly complete reestablishment of alpha/beta globin chain balance, as demonstrated by a marked decrease in unpaired alpha globin chain. Eight months after the first electrotransfer of pCMV-mEPO plasmid, reinjection of the same construct raised the hematocrit to a level close to that observed following the first electrotransfer. CONCLUSION: This is the first description of the use of plasmid DNA to achieve long-term improvement in a mouse model of a human genetic disorder. PMID- 11274757 TI - Expression of FLRG, a novel activin A ligand, is regulated by TGF-beta and during hematopoiesis [corrected]. AB - OBJECTIVE: The human gene FLRG, identified from a B-cell chronic lymphocytic leukemia bearing a t(11;19) translocation, encodes a secreted glycoprotein highly homologous with follistatin. Activin A is a TGF-beta family member involved in the regulation of growth and differentiation of various types of cells, such as those of the hematopoietic system. Its biological activity is antagonized by binding with follistatin. We investigated the binding of FLRG to activin A and the expression pattern of FLRG, follistatin, and activin A during hematopoiesis. MATERIALS AND METHODS: The binding of FLRG with activin A was investigated by immunoprecipitation and Far-Western blot analysis. Gene expression was analyzed by reverse transcriptase polymerase chain reaction (RT-PCR) and Northern Blot in purified hematopoietic populations. RESULTS: We demonstrate that FLRG, like follistatin, is able to bind to activin A. In bone marrow stromal cells, both mRNA and protein FLRG levels were found to be dramatically increased by TGF-beta. FLRG and activin A are expressed in the same cells, with a higher level of expression in the myeloid cells compared with the erythroid and megakaryocytic cells. FLRG and follistatin expression were different in the hematopoietic subpopulations tested. Moreover, we observed that FLRG and activin A expression was up-regulated during hematopoiesis. CONCLUSION: FLRG and activin A are expressed in the same hematopoietic cells and regulated by TGF-beta. Moreover, FLRG interacts with activin A, suggesting that FLRG, like follistatin, participates in the diverse regulatory functions of activin A, such as those in hematopoiesis. PMID- 11274758 TI - Modified in vitro conditions for cord blood-derived long-term culture-initiating cells. AB - OBJECTIVE: The aim of this study was to compare the in vitro growth of cord blood derived progenitors with that of bone marrow and peripheral blood. MATERIALS AND METHODS: We analyzed 192 umbilical cord blood (UCB), 35 normal bone marrow (NBM), and 35 granulocyte colony-stimulating factor (G-CSF)-primed normal peripheral blood (NPB) samples. Standard clonogenic assays (colony-forming unit granulocyte macrophage [CFU-GM], burst-forming unit erythroid [BFU-E], CFU-granulocyte erythroid megakaryocyte macrophage [GEMM]) and standard long-term culture initiating cell (LTC-IC) assay were performed. LTC-IC frequency also was tested under modified culture conditions. The variables tested were incubation temperature (37 degrees C and 33 degrees C) and supportive stromal cell lines (NIH3T3 and M210-B4). RESULTS: The CFU-GM and CFU-GEMM frequencies of UCB samples were similar to NPB and higher compared to NBM samples (p < 10(-4) and p < 0.007 respectively). On the other hand, the BFU-E frequency was lower in cord blood samples (5.2 +/- 5.6/10(4) MNC) compared to bone marrow (7 +/- 3.8/10(4) MNC; p < 0.005) and peripheral blood (15.2 +/- 11.1/10(4) MNC; p < 10(-4)). All colony types (CFU-GM, BFU-E, CFU-GEMM) generated from cord blood progenitors were larger with respect to the other tissues. The LTC-IC frequency was markedly decreased (8.8 +/- 3.8/10(6) MNC) in cord blood with respect to bone marrow (40.7 +/- 7.4/10(6) MNC; p < 10(-4)) and peripheral blood (28.8 +/- 3.8/10(6) MNC; p < 0.04). However, when culture conditions (temperature, stromal layers) were modified, UCB-LTC-IC frequency significantly increased, while the growth of early progenitors derived from adult tissues (BM and PB) did not show any variation. Whatever culture conditions were used, the proliferative potential of UCB LTC-IC was significantly higher with respect to bone marrow and G-CSF-primed PB (10.6 +/ 7.7 colonies vs. 5.9 +/- 5 vs 3.2 +/- 2.2 colonies; p < 0.02 and p < 0.001 respectively). CONCLUSIONS: Optimal conditions for estimation of the LTC-IC frequency in cord blood samples seem to be different from those usually applied to PB and BM progenitors. Although UCB hemopoietic progenitors have a higher proliferative potential than those from bone marrow and G-CSF-primed peripheral blood, their quantitation depends on the culture conditions, which makes it difficult to establish their exact number. This problem and the fact that a significant proportion of UCB samples grew poorly in culture make it necessary to develop suitable and standardized functional assays to test UCB progenitor content before the transplantation procedure. PMID- 11274760 TI - Monitoring of engraftment and progression of acute lymphoblastic leukemia in individual NOD/SCID mice. AB - OBJECTIVE: The aim of this study was to develop an animal model for human acute lymphoblastic leukemia (ALL) in which the kinetics and characteristics of leukemia can be sequentially monitored in individual mice. MATERIALS AND METHODS: NOD/SCID mice were inoculated intravenously with primary ALL. Progression of leukemia was monitored throughout the development of disease by determination of absolute leukemic cell counts (LCC) in peripheral blood. RESULTS: LCC as low as 10(4) leukemic cells/mL blood could be detected. ALL cells from 5 of 5 patients engrafted, and after identification of the first leukemic cells in peripheral blood, LCC increased exponentially. Leukemic cells showed specificity of homing to spleen and bone marrow, and LCC strongly correlated with the level of leukemic engraftment in these organs throughout disease progression, demonstrating that LCC are representative for overall leukemic burden. Cytogenetic analysis of leukemic cells recovered after six successive in vivo transfers revealed no major karyotypic changes as compared to primary cells, and selection of the dominant clones was observed. This selection process was reflected by an increase in the rate of leukemic progression as compared to the first inoculation, demonstrating the accuracy with which kinetics of leukemic progression can be studied by determination of LCC. CONCLUSIONS: This model is suitable for detailed studies of kinetics and characteristics of ALL in vivo, and it may be useful for monitoring effects of novel therapeutic regimens. PMID- 11274759 TI - Purging of myeloma cells using all-trans retinoic acid in a mouse model. AB - OBJECTIVE: The 5T33 murine model of multiple myeloma was used to investigate the potential of all-trans retinoic acid (ATRA) to purge clonogenic myeloma cells from autologous hemopoietic stem-cell harvests by differentiating immature 5T33 cells into terminal-stage plasma cells with limited repopulation capacity. MATERIALS AND METHODS: 5T33 cells were treated with 10 microM ATRA and the effect on cell clonogenicity was determined by measuring the time to paraprotein detection in C57Bl/KaLwRij mice compared to control animals. Cell differentiation and apoptosis following ATRA treatment were investigated using flow cytometry and caspase-3 assay. Treatment with ATRA resulted in a 33% reduction in the in vitro cloning efficiency of 5T33 cells. Reduced in vitro clonogenicity of 5T33 cells following ATRA treatment was supported by a 16-49% increase in the time taken for C57Bl/KaLwRij mice to develop paraprotein following injection of 5T33 cells pretreated with ATRA for 8 days. Although ATRA was shown not to alter the in vitro growth characteristics of 5T33 cells, significant inhibition of apoptosis was observed. RESULTS: Treatment with ATRA also resulted in an increase in the proportion of 5T33 cells expressing the CD54 adhesion molecule, which is known to be highly expressed on mature myeloma cells. CONCLUSION: The ability of ATRA to decrease the clonogenicity of 5T33 cells in vitro and increase the time to disease development in vivo suggests that this drug may be useful for purging autologous stem cell harvests in the clinical setting. PMID- 11274761 TI - Higher LPS-stimulated TNF-alpha mRNA levels in peripheral blood mononuclear cells from non-Hodgkin's lymphoma patients. AB - OBJECTIVE: The aim of the present study was to investigate the capacity of normal immune blood cells from non-Hodgkin's lymphoma patients to produce tumor necrosis factor (TNF) after lipopolysaccharide (LPS) stimulation and the influence of the TNF (-308) polymorphism in this production. MATERIALS AND METHODS: A whole peripheral blood cell assay was utilized following LPS stimulation. At selected incubation times, supernatants were harvested for protein dosage, while mRNA was extracted and reverse-transcribed. The amount of TNF mRNA was quantified using real-time quantitative polymerase chain reaction (PCR) and genomic DNA was typed for TNF (-308) polymorphism. RESULTS: Upon LPS stimulation, TNF-secreted protein was slightly but not significantly increased in lymphoma patients when compared to controls. In contrast, the relative TNF mRNA amounts were significantly higher in lymphoma patients at 30 minutes (median 27.75 vs. 16.00; Mann-Whitney U-test p < 0.05), at 4 hours (52.00 vs. 31.00; p < 0.05), and at 24 hours (19.50 vs. 9.00; p < 0.05). In addition, patients carrying the variant TNF2 allele had higher relative TNF mRNA levels than TNF1 homozygotes (p = 0.02). CONCLUSION: The LPS induced TNF mRNA levels are higher in peripheral blood cells (PBC) from lymphoma patients than from controls, while TNF protein secretion is not strikingly different. Altered regulation of TNF mRNA translation or TNF protein secretion may contribute to these observations. Taken together, an increased susceptibility for TNF gene transcription after LPS stimulation was observed in PBC (mainly in monocytes) from lymphoma patients, and especially those carrying the TNF2 allele. PMID- 11274762 TI - Bone marrow aplasia induced by passenger leukocytes from heart allografts. AB - OBJECTIVE: Organ allografts contain passenger leukocytes that are transferred to the recipient with the transplantation, but their functional relevance to the recipient's immune system is still controversial. MATERIALS AND METHODS: To clarify the functional capacity of passenger leukocytes, we attempted to enhance their effect in rat heart allograft recipients by selective depletion of recipient leukocytes using a monoclonal antibody (mAb) against a recipient specific allotype of CD45 (RT7(a)). RESULTS: Although antibody treatment of the recipient alone led to profound lymphopenia and reversible myelosuppression, additional transplantation of an major histocompatibility complex-incompatible heart graft from an RT7(b) donor led to lethal aplastic anemia in the recipients. This lethal effect was completely abrogated by postoperative anti-CD3 treatment of the recipient and was partially abrogated or delayed by depletion of passenger leukocytes through additional anti-RT7(b) antibody treatment of the recipient or gamma-irradiation of the graft. CONCLUSIONS: The results suggest a role for both donor and recipient-type T cells for the induction of aplastic anemia in this model. The study shows that, under defined conditions, allogeneic passenger leukocytes in a heart graft can have a profound effect on the recipient's immune system and bone marrow. PMID- 11274763 TI - Chemokine stromal cell-derived factor-1alpha modulates VLA-4 integrin-dependent adhesion to fibronectin and VCAM-1 on bone marrow hematopoietic progenitor cells. AB - Stromal cell-derived factor-1alpha (SDF-1alpha) is a potent chemoattractant for hematopoietic progenitor cells (HPC), suggesting that it could play an important role during their migration within or to the bone marrow (BM). The integrin VLA-4 mediates HPC adhesion to BM stroma by interacting with CS-1/fibronectin and VCAM 1. It is required during hematopoiesis and homing of HPC to the BM. As HPC migration in response to SDF-1alpha might require dynamic regulation of integrin function, we investigated if SDF-1alpha could modulate VLA-4 function on BM CD34(hi) cells.CD34(hi) BM cells and hematopoietic cell lines were tested for the effect of SDF-1alpha on VLA-4-dependent adhesion to CS-1/fibronectin and VCAM-1, as well as to BM stroma. CD34(hi) BM cells that adhered to VLA-4 ligands after SDF-1alpha treatment were characterized in colony-forming and long-term culture initiating cell (LTC-IC) assays.SDF-1alpha rapidly (1 minute) and transiently upregulated the adhesion of CD34(hi) BM cells and hematopoietic cell lines to both CS-1/fibronectin and VCAM-1, and to BM stromal cells. The upregulation of VLA-4-dependent cell adhesion by SDF-1alpha targeted primitive LTC-IC as well as committed CD34(hi) cells. SDF-1alpha-triggered enhancement in VLA-4 function was inhibited by pertussis toxin (PTx) and cytochalasin D, indicating the involvement of G(i) protein downstream signaling and an intact cytoskeleton. Instead, activation of p44/42 MAP kinases by SDF-1alpha did not functionally correlate with enhancement of VLA-4-dependent cell adhesion. Modulation of VLA-4-mediated CD34(hi) BM cell adhesion by SDF-1alpha could play a key role in their migration within and to the BM and therefore influence their proliferation and differentiation. PMID- 11274765 TI - Low-intensity conditioning is sufficient to ensure engraftment in matched unrelated bone marrow transplantation. AB - OBJECTIVE: Matched unrelated bone marrow transplantation (BMT) for patients with hematological malignancies is associated with a high incidence of transplant related complications due to high doses of chemoradiotherapy administered pre-BMT to ensure engraftment. The aim of this study was to investigate the feasibility of low-intensity conditioning for BMT from matched unrelated donors. MATERIALS AND METHODS: Sixteen patients with hematologic malignancies underwent non-T-cell depleted BMT following a low-intensity conditioning regimen consisting of fludarabine monophosphate 30 mg/m(2)/day for 6 days, busulfan 4 mg/kg/day for 2 days, anti-T lymphocyte globulin 10 mg/kg/day for 4 days. Seven of the patients suffered from chronic myelogenous leukemia, four from acute lymphoblastic leukemia, four from acute myelogenous leukemia, and one from Ki-1 non-Hodgkin's lymphoma. Three of the patients had secondary leukemia and two were post autologous BMT (ABMT). All patients were transplanted from fully matched unrelated donors. RESULTS: Fifteen of the 16 patients had 100% donor chimerism; no graft rejection was observed. None of the patients developed >Grade II veno occlusive disease, sepsis, multiorgan failure, or renal or pulmonary toxicity. Four patients died posttransplant; one of thrombocytopenia and severe hemorrhagic cystitis, one of central nervous system toxicity, one of Grade IV graft-vs-host disease, and one following relapse (9 months post-BMT). Survival and disease-free survival at 36 months are 75% (95% confidence interval 46-90%) and 60% (95% confidence interval 30-80%), respectively. CONCLUSION: These results indicate that low-intensity conditioning is sufficient to ensure stable engraftment of bone marrow grafts in a matched unrelated setting. PMID- 11274764 TI - CIS1, a cytokine-inducible SH2 protein, suppresses BCR/ABL-mediated transformation. Involvement of the ubiquitin proteasome pathway. AB - OBJECTIVE: BCR/ABL is a chimeric oncoprotein that exhibits deregulated tyrosine kinase activity and is implicated in the pathogenesis of Philadelphia chromosome (Ph)-positive leukemia. A general understanding of BCR/ABL signaling events is emerging, but little is known about the endogenous inhibitors of p210 BCR/ABL. The present study focused attention on CIS1, a cytokine-inducible SH2 protein, as a potential physiologic antagonist for BCR/ABL. MATERIALS AND METHODS: The murine hematopoietic cell line NSF/N1.H7 stably transfected with BCR/ABL was compared to the parental counterparts for induction of CIS1 by immunoblotting and immunoprecipitation. Cells were treated with a proteasome inhibitor to examine the effect of a proteasome inhibitor on CIS1 protein expression. To determine the effect of CIS1 on BCR/ABL-mediated transformation, we generated Rat-1 fibroblasts transfected with either a control vector, CIS1, BCR/ABL p210, or CIS1 plus BCR/ABL p210. RESULTS: Three forms of CIS1 with molecular masses of 32, 37, and 47 kDa were detected in BCR/ABL-transformed cells. The 47-kDa protein was a ubiquitinated protein. The proteasome inhibitor increased the formation of complexes between CIS1 and BCR/ABL. Transformation of p210 BCR/ABL was significantly suppressed in cells overexpressing CIS1. CONCLUSION: The results suggest that CIS1 is an endogenous inhibitor of p210 BCR/ABL and is likely to be important in the pathogenesis of Ph-positive leukemia. PMID- 11274766 TI - Analysis of immune reconstitution in children undergoing cord blood transplantation. AB - OBJECTIVE: The aim of this study was to investigate and compare immune reconstitution in allogeneic cord blood transplantation (CBT) and bone marrow transplantation (BMT) recipients. MATERIALS AND METHODS: Twenty-three children underwent CBT from either human leukocyte antigen-identical siblings (11 cases) or unrelated donors (12 cases) were enrolled in the study, together with 23 matched children receiving BMT. Patients were analyzed 2-3 and 12-15 months after transplant. Recovery of T-, B-, and NK-lymphocyte subsets, proliferative in vitro response to mitogens, as well as cytotoxic activities, were investigated. RESULTS: CBT recipients showed a marked increase in the number of B lymphocytes as compared with patients who underwent BMT (p < 0.001). The absolute number of CD3(+) and CD8(+) T cells, as well as the proliferative response to T-cell mitogens, recovered with time after transplantation, irrespective of the source of stem cells used. Recipients of unrelated CBT had a better recovery of CD4(+) T lymphocytes (p < 0.01). Among patients experiencing acute graft-versus-host disease (GVHD), children given CBT had a much greater production of CD4(+) CD45RA(+) T cells than BMT recipients (p < 0.005). Recovery of NK cell number and innate cytotoxic activities was fast, irrespective of the source of stem cells used. CONCLUSIONS: Despite the much lower number of lymphocytes transferred with the graft, recovery of lymphocyte number and function toward normal in CBT recipients was rapid and comparable to that observed after transplantation of bone marrow progenitors. This prompt immune recovery possibly was favored by the reduced incidence and severity of GVHD observed in children who underwent CBT. PMID- 11274768 TI - Effect of a chronic and moderate ozone pollution on the phenolic pattern of bean leaves (Phaseolus vulgaris L. cv Nerina): relations with visible injury and biomass production. AB - From sowing, bean (Phaseolus vulgaris L. cv Nerina) plants were exposed to three chronic doses of ozone for 7h.day(-1): non-filtered air (NF), non-filtered air supplied with 40nl.l(-1) ozone (NF+40) and non-filtered air supplied with 60nll( 1) ozone (NF+60). Four harvests were carried out 6, 13, 20 and 27 days after emergence. Either primary leaves, or first trifoliate leaves, or both were sampled as far as possible. For each sampled leaf, visible ozone injuries were registered, the free polyphenolic pool was analysed using HPLC and the dry matter was weighed. Visible damage on leaves was related to both exposure time and ozone concentration added. There were no adverse effects of added ozone on the biomass of primary leaves while a significant reduction of first trifoliates dry matter could be observed (NF+60 atmosphere, third and fourth harvest). Among the normally occurring phenolics, we detected a significant decrease in the accumulation of a hydroxycinnamic acid derivative as the ozone concentration increased. Nevertheless, we demonstrated that this ozone-induced modification could be sometimes distinguishable with difficulties from changes expected to be of development relevance. Beside this phenolic disbalance, we detected a de novo biosynthesis of compounds that closely depended on the level of visible ozone injury. Since their accumulation increased with leaf damage, these ozone-induced phenolics could be used to detect phytotoxic ambient levels of tropospheric ozone. PMID- 11274769 TI - Exudation of low molecular weight compounds (thiobismethane, methyl isocyanide, and methyl isothiocyanate) as a possible chemical defense mechanism in the marine sponge Ircinia felix. AB - The volatile constituents of the marine sponge Ircinia felix were obtained by dynamic headspace extraction and analyzed by HRGC, HRGC-MS and HRGC-Odor at sniffing port. Fifty-nine volatiles were identified for the first time in the odor of this sponge. Hydrocarbons (32.9%), alcohols (17.8%) and carbonyl compounds (16.0%) predominated in the sponge volatile profile, followed by esters (11.6%), halogen compounds (8.6%), ethers (7.7%), nitrogen and/or sulfur compounds (4.6%) and carboxylic acids (0.8%). Among the identified volatiles, thiobismethane (commonly known as dimethylsulfide), methyl isocyanide and methyl isothiocyanate were found to be responsible for the nauseating and toxic smell emitted by the sponge and for the antimicrobial activity detected in the volatile extract. Exudation experiments in aquarium and in situ conditions revealed that thiobismethane, methyl isocyanide and methyl isothiocyanate are continuously released by the sponge. Upon injury, the concentration of these volatiles increased strongly. Hence, these substances form a chemical protective barrier which may help these sponges avoid fouling, compete for space, prevent infection in the short term, and/or signal generalist predators regarding the existence of other toxic substances in the internal tissues. PMID- 11274767 TI - Combining G-CSF with a blockade of adhesion strongly improves the reconstitutive capacity of mobilized hematopoietic progenitor cells. AB - OBJECTIVE: Mobilization of hematopoietic progenitor cells is achieved mainly by application of growth factors and, more recently, by blockade of adhesion. In this report, we describe the advantages of a combined treatment with granulocyte colony-stimulating factor (G-CSF) and anti-VLA4 (CD49d)/anti-CD44 as compared to treatment with the individual components. MATERIALS AND METHODS: Mobilization by intravenous injection of anti-CD44, anti-VLA4, or G-CSF was controlled in spleen and bone marrow with regard to frequencies of multipotential colony-forming unit (C-CFU), marrow repopulating ability, long-term reconstitution, recovery of myelopoiesis, and regain of immunocompetence. RESULTS: Mobilization by anti-CD44 had a strong effect on expansion of early progenitor cells in the bone marrow, while the recovery in the spleen was poor. In anti-CD49d-mobilized noncommitted and committed progenitors, progenitor expansion was less pronounced, but settlement in the spleen was quite efficient. Thus, anti-CD44 and anti-CD49d differently influenced mobilization. Accordingly, mobilization and recovery after transfer were improved by combining anti-CD44 with anti-CD49d treatment. Mobilization by G-CSF was most efficient with respect to recovery of progenitor cells in the spleen. However, when transferring G-CSF-mobilized cells, regain of immunocompetence was strongly delayed. This disadvantage could be overridden when progenitor cells were mobilized via blockade of adhesion and when expansion of these mobilized progenitor cells was supported by low-dose G-CSF only during the last 24 hours before transfer. CONCLUSION: Mobilization of pluripotent progenitor cells via antibody blockade of CD44 or CD49d or via G-CSF relies on distinct mechanisms. Therefore, the reconstitutive capacity of a transplant can be significantly improved by mobilization regimens combining antibody with low-dose G-CSF treatment. PMID- 11274771 TI - Allozyme variation in four populations of African whitebacked vultures (Gyps africanus) and phylogenetic relationships between four vulture species from southern Africa. AB - Genetic variation detected by protein electrophoresis at 41 presumptive gene loci was assayed in four populations of Gyps africanus and compared to values previously obtained for Gyps coprotheres. Values calculated for percentage of polymorphic loci (P=34.15%, 0.99 criterion) and average heterozygosity (&Hmacr;=0.108, +/-0.032) in G. africanus, confirm low levels of genetic variation as reported for G. coprotheres. Allele frequency data, assessed at 19 loci, were obtained to evaluate genetic differentiation among four vulture species. Six (31.58%) of the 19 shared loci were polymorphic. Values of 1.26 (+/ 0.1), 26.32% and 0.076 (+/-0.047) for G. africanus, 1.21 (+/-0.1), 21.05% and 0.097 (+/-0.045) for Torgos tracheliotus, 1.11 (+/-0.7), 21.05% and 0.053 (+/ 0.053) for Neophron percnopterus and 1.05 (+/-0.5), 5.26% and 0.044 (+/-0.047) for G. coprotheres were obtained for the mean number of alleles per locus, P and &Hmacr;, respectively. An average between-population fixation index (F(ST)) value of 0.322 was obtained, which is indicative of significant (P<0.01) differentiation between the four accipitrid species studied. Considerable concordance was obtained between dendograms produced from different analyses, pointing to the distinctiveness of N. percnopterus, which has evolved along a separate lineage as G. africanus, G. coprotheres and T. tracheliotus. Along the latter lineage G. africanus is clustered together with G. coprotheres which is consistent with the morphological similarities of these species. PMID- 11274770 TI - A review of the use of allozyme electrophoresis in plant systematics. AB - The role of electrophoretic data is discussed as it applies to plant taxonomy and systematic studies. Nei's (Am. Nat. 106 (1972) 283-292; Genetics 89 (1978) 583 590) genetic distances calculated for a large number of populations, species and genera were taken from published data. The relation between Nei's genetic identity measures and taxonomic rank (populations, species and genera) are shown graphically. The graphs obtained in this way (from 3021 pairs of plant taxa) differ substantially from previous graphs published by Thorpe (Ann. Rev. Ecol. Syst. 13 (1982) 139-168; in: G.S. Oxford, D. Rollinson (Eds.), Protein Polymorphism: Adaptive and Taxonomic Significance, Academic Press, London, 1983, pp. 131-152) and Thorpe and Sole-Cava (Zool. Scripta 23 (1994) 3-18). These authors suggested that the divergence between the different taxonomic ranks is roughly similar across a wide range of taxa. The latter was based on values for 2664 (Thorpe, 1982) and 8060 (Thorpe, 1983) pairs of animal and plant taxa, but the plant data contributed little to the total. For any given taxonomic rank, we found that plants are genetically more closely related than animals (possibly with the exception of birds). This result is important because the empirical relationships of genetic distance measures, to different levels of taxonomic separation, is often used for distinguishing and identifying cryptic or sibling species where conventional methods are unable to resolve systematic problems. PMID- 11274772 TI - The domestication syndrome within Hybrid Perpetuals roses: the effect of unconscious selection on flavonoids. AB - Cultivated GallicanaexChinenses hybrids of roses, namely Hybrid Perpetuals, were compared with their parents as to morphology, petal colour, flavonol and anthocyanin metabolism. Morphology exhibited clear patterns of hybridity. An objective measure of petal lightness (L) indicated that Hybrid Perpetuals were submitted to a selection pressure favouring dark-flowered cultivars. When compared to the parental flavonoid metabolisms, Hybrid Perpetuals exhibited increased synthesis of anthocyanin and quercetin. High amounts of anthocyanin in Hybrid Perpetuals resulted from the selection of deeper-coloured flowers. High amounts of quercetin were correlated with enhanced anthocyanin synthesis, so that this originality of the flavonol metabolism was interpreted on biogenetic ground as a repercussion of this same selection pressure. Finally, the patterns of variation of flavonol glycosides within the Hybrid Perpetuals reflected the indirect selection pressure for the quercetin end-products, and with the ancestral hybridizations for the kaempferol derivatives. PMID- 11274773 TI - 1-Octen-3-ol, a banana slug antifeedant from mushrooms. PMID- 11274774 TI - Etioline, a steroidal alkaloid from Lilium candidum L. PMID- 11274775 TI - Tetrahydropiperine, the first natural aryl pentanamide from Piper longum. PMID- 11274777 TI - Homohesperetin and phaseollidin from Erythrina velutina. PMID- 11274776 TI - Scillascillin-type homoisoflavanones from Drimiopsis maculata (Hyacinthaceae). PMID- 11274778 TI - On the occurrence of glucozaluzanin C in Leontodon cichoraceus and its chemotaxonomic significance. PMID- 11274779 TI - "Constituents of Rhamnus virgatus (Rhamnaceae)" by D. Prasad, G. Pant, M.S.M. Rawat, A. Nagatsu. Biochemical Systematics and Ecology 28(10) pp. 1027-1030. PMID- 11274780 TI - Low-resolution brain electromagnetic tomography revealed simultaneously active frontal and parietal sleep spindle sources in the human cortex. AB - Analyses of scalp-recorded sleep spindles have demonstrated topographically distinct slow and fast spindle waves. In the present paper, the electrical activity in the brain corresponding to different types of sleep spindles was estimated by means of low-resolution electromagnetic tomography. In its new implementation, this method is based on realistic head geometry and solution space is restricted to the cortical gray matter and hippocampus. In multichannel all-night electroencephalographic recordings, 10-20 artifact-free 1.25-s epochs with frontally, parietally and approximately equally distributed spindles were marked visually in 10 normal healthy subjects aged 20-35years. As a control condition, artifact-free non-spindle epochs 1-3s before or after the corresponding spindle episodes were marked. Low-resolution electromagnetic tomography demonstrated, independent of the scalp distribution, a distributed spindle source in the prefrontal cortex (Brodmann areas 9 and 10), oscillating with a frequency below 13Hz, and in the precuneus (Brodmann area 7), oscillating with a frequency above 13Hz. In extremely rare cases only the prefrontal or the parietal source was active. Brodmann areas 9 and 10 have principal connections to the dorsomedial thalamic nucleus; Brodmann area 7 is connected to the lateroposterior, laterodorsal and rostral intralaminar centrolateral thalamic nuclei. Thus, the localized cortical brain regions are directly connected with adjacent parts of the dorsal thalamus, where sleep spindles are generated. The results demonstrated simultaneously active cortical spindle sources which differed in frequency by approximately 2Hz and were located in brain regions known to be critically involved in the processing of sensory input, which is in line with the assumed functional role of sleep spindles. PMID- 11274781 TI - Neural interaction between the basal forebrain and functionally distinct prefrontal cortices in the rhesus monkey. AB - The prefrontal cortex in rhesus monkeys is a heterogeneous region by structure, connections and function. Caudal medial and orbitofrontal cortices receive input from cortical and subcortical structures associated with emotions, autonomic function and long-term memory, while lateral prefrontal cortices are linked with structures associated with working memory. With the aid of neural tracers we investigated whether functionally distinct orbitofrontal, medial and lateral prefrontal cortices have specific or common connections with an ascending modulatory system, the basal forebrain. Ascending projections originated in the diagonal band and the basalis nuclei of the basal forebrain in regions demarcated by choline acetyltransferase. Although the origin of projections from the basal forebrain to lateral, medial and orbitofrontal cortices partially overlapped, projections showed a general topography. The posterior part of the nucleus basalis projected preferentially to lateral prefrontal areas while its rostrally adjacent sectors projected to medial and orbitofrontal cortices. The diagonal band nuclei projected to orbitofrontal and medial prefrontal areas. Cortical and subcortical structures that are interconnected appear to have a similar pattern of connections with the basal forebrain. In comparison to the ascending projections, the descending projections were specific, originating mostly in the posterior (limbic) component of medial and orbitofrontal cortices and terminating in the diagonal band nuclei and in the anterior part of the nucleus basalis. In addition, prefrontal limbic areas projected to two other systems of the basal forebrain, the ventral pallidum and the extended amygdala, delineated with the striatal-related markers dopamine, adenosine 3':5'-monophosphate regulated phosphoprotein of M(r) 32kDa, and the related phosphoprotein Inhibitor-1. These basal forebrain systems project to autonomic nuclei in the hypothalamus and brainstem. We interpret these results to indicate that lateral prefrontal areas, which have a role in working memory, receive input from, but do not issue feedback projections to the basal forebrain. In contrast, orbitofrontal and medial prefrontal areas, which have a role in emotions and long-term memory, have robust bidirectional connections with the basal forebrain. Moreover, orbitofrontal and medial prefrontal cortices target the ventral pallidum and the extended amygdala, through which high-order association areas may activate motor autonomic structures for the expression of emotions. PMID- 11274782 TI - Neurons in the lateral agranular frontal cortex have divided attention correlates in a simultaneous temporal processing task. AB - The frontal cortex is an important brain area for divided attention. Lesions of the lateral agranular frontal cortex in rats disrupt divided attention in a simultaneous temporal processing task. In the present study, the activity of lateral agranular neurons was examined while rats performed a simultaneous temporal processing procedure. Rats were trained to time two stimuli (a light and a tone), each associated with a different fixed interval. Simple trials, in which a single stimulus was presented, and compound trials, in which both stimuli were presented simultaneously, occurred randomly in a session. Rats were able to divide attention between the two stimuli, as assessed by the pattern of lever presses. Approximately 50% of lateral agranular neurons responded to at least one phase of the task with four response patterns observed. The activity of type 1 cells (60%) was altered to compound, but not simple, stimuli. Type 2 cells (10%) responded to both types of simple stimuli and to compound stimuli. Type 3 cells (27%) had changes in firing rate to one type of simple stimulus and to compound stimuli. Type 4 cells (3%) responded to one type of simple stimulus, but were unresponsive to all other stimuli. The large proportion of type 1 cells supports the hypothesis that the lateral agranular cortex is important in divided attention. Previous studies have suggested that the lateral agranular cortex in rats is equivalent to the primary motor cortex. If so, the results from the present study provide evidence that the lateral agranular cortex may have some cognitive functions, in addition to being part of the motor system. PMID- 11274783 TI - Ovariectomy up-regulates neuronal neurofilament light chain mRNA expression with regional and temporal specificity. AB - Estrogens can influence the survival, plasticity and function of many adult neurons. Many of these effects, such as neurite outgrowth and increased dendritic spine density, are mediated by changes in neuronal cytoskeletal architecture. Since neurofilament proteins play a key role in the maintenance and remodeling of the neuronal cytoskeleton, we postulated that changes in neurofilament light chain mRNA may parallel some of the alterations in neuronal architecture which follow bilateral ovariectomy. We measured neurofilament light chain mRNA levels using a ribonuclease protection assay at two time-points after ovariectomy in mature female rats. One week after ovariectomy, neurofilament light chain mRNA levels (corrected for glucose-6-phosphate dehydrogenase mRNA) did not differ from sham-operated animals in the five brain regions examined (hypothalamus, striatum, hippocampus, frontal cortex and occipital cortex). Four months after ovariectomy, neurofilament light chain mRNA levels were similarly unchanged in the hypothalamus and striatum. In contrast, statistically significant increases in neurofilament light chain mRNA expression were observed in the three regions receiving basal forebrain projections (hippocampus, frontal cortex and occipital cortex). In situ hybridization demonstrated increases in neurofilament light chain mRNA expression involving subpopulations of smaller medial septal neurons. There also appeared to be an increased number of larger septal neurons following long-term ovariectomy. We propose that atrophic changes involving basal forebrain projection fibers are followed by compensatory axonal growth by other 'intact' basal forebrain neurons. Increased neurofilament light chain mRNA expression and somatic hypertrophy in medial septal neurons may both be reflective of the need to sustain an axonal network which is larger and more complex. In contrast, increased neurofilament light chain mRNA expression observed in basal forebrain targets following long-term ovariectomy may be reflective of compensatory changes taking place in local neurons. PMID- 11274784 TI - Mechanisms of the effects of exogenous levodopa on the dopamine-denervated striatum. AB - The efficacy of exogenous levodopa (L-DOPA) is attributed to its conversion to dopamine by the enzyme aromatic L-amino-acid decarboxylase in striatal dopaminergic terminals. However, there is controversy about the mechanisms underlying the therapeutic and adverse effects of L-DOPA after almost all striatal dopaminergic afferents have disappeared (i.e. in the later stages of Parkinson's disease). After administration of 30mg/kg or 100mg/kg of L-DOPA, rats subjected to unilateral dopaminergic denervation showed intense contraversive rotation and a high density of Fos-immunoreactive nuclei throughout the denervated striatum, with no significant induction of Fos in the intact striatum. Injection of the central aromatic L-amino-acid decarboxylase inhibitor NSD-1015 30min before and 15min after the injection of L-DOPA suppressed the rotational behavior and the striatal induction of Fos. Comparison of results obtained in rats subjected to unilateral and bilateral dopaminergic denervation indicated that the presence of contralateral dopaminergic innervation does not significantly modulate the effects of L-DOPA on the denervated striatum. Serotonergic denervation led to slight and statistically non-significant decrease in the rotational behavior and Fos expression induced by high doses of L-DOPA (100mg/kg) in the dopamine-denervated striatum, but totally suppressed the rotational behavior and Fos expression induced by low doses of L-DOPA (30mg/kg). The present data indicate that the major effects observed after administration of exogenous L-DOPA are not due to a direct action of L-DOPA on dopamine receptors, or to extrastriatal release of dopamine, but to conversion of L-DOPA to dopamine by serotonergic terminals and probably some intrastriatal cells. Given that serotonergic neurons appear to play an important role in the action of L-DOPA in the later stages of Parkinson's disease, strategies targeting the serotonergic system should be considered for the treatment of Parkinson's disease and for combating undesirable side effects of L-DOPA therapy. PMID- 11274785 TI - Parafascicular thalamic nucleus deafferentation reduces c-fos expression induced by dopamine D-1 receptor stimulation in rat striatum. AB - We investigated the role played by the parafascicular thalamostriatal pathway, one of the major excitatory inputs to the striatum, in the D-1 receptor induction of c-fos messenger RNA expression in the rat striatum. The full D-1 receptor agonist, SKF-82958 (0.05, 0.1, 0.5 and 1 mg/kg, s.c., 30 min), dose-dependently induced c-fos messenger RNA in naive rat striatum as determined by northern blot analysis. One day following electrolytic lesion of the parafascicular thalamostriatal nucleus, striatal c-fos signal by itself was not altered but the stimulated expression of c-fos was strongly decreased. Sections of sham-operated and acute-lesioned dorsal striata of vehicle- or SKF-82958-treated rats were processed for in situ hybridization histochemistry at the single-cell level with an RNA probe for c-fos. The basal expression of striatal c-fos was poorly detectable in sham and lesioned groups. Sections of dorsal striata from sham operated rats treated with SKF-82958 showed two types of labeled neurons for c fos: the lightly and the very densely labeled neurons which are few in number. Thalamic lesion strongly reduced SKF-82958 stimulated expression of c-fos RNA in both types of labeled cells. The frequency distribution of c-fos labeling per neuron in dorsal striata of lesioned rats treated with SKF-82958 was shifted to the left, and its median was lower than in the sham-operated striata treated with the D-1 receptor agonist. The results provide evidence that the excitatory projections from the parafascicular nucleus of the thalamus, thought to operate primarily through the N-methyl-D-aspartate receptors, exert a facilitatory control over D-1 receptor-induced c-fos gene expression specifically in the dorsal striatum. PMID- 11274786 TI - Effects of neurotensin on discharge rates of rat suprachiasmatic nucleus neurons in vitro. AB - The neuropeptide neurotensin and two classes of its receptors, the neurotensin receptor-1 and 2, are present in the suprachiasmatic nucleus of the mammalian hypothalamus. The suprachiasmatic nucleus houses the mammalian central circadian pacemaker, but the effects of neurotensin on cellular activity in this circadian pacemaker are unknown. In this study, we examined the effects of neurotensin on the spontaneous discharge rate of rat SCN cells in an in vitro slice preparation. Neurotensin (1-10 microM) increased cell firing rate in approximately 50% of cells tested, while approximately 10% of suprachiasmatic cells tested showed a decrease in firing rate in response to neurotensin. These effects of neurotensin were not altered by the GABA receptor antagonist bicuculline (20 microM) or the glutamate receptor antagonists, D-aminophosphopentanoic acid (50 microM) and 6 cyano-7-nitroquinoxaline-2,3-dione (20 microM). The neurotensin receptor selective antagonists SR48692 and SR142948a (10 microM) failed to antagonise neurotensin responses in the majority of cells examined. Compounds that function as agonists selective for the neurotensin-receptor subtypes 1 and 2, JMV-510 and JMV-431 respectively, elicited neurotensin-like responses in approximately 90% of cells tested. Six out of seven cells tested responded to both JMV-510 and JMV 431. Neuropeptide Y (100nM) treatment of suprachiasmatic nucleus slices was found to elicit profound suppression of neuronal firing rate. Co-application of neurotensin with neuropeptide Y significantly (P<0.05) reduced the duration of the response, as compared to that elicited with neuropeptide Y alone. Together, these results demonstrate for the first time the actions of neurotensin in the suprachiasmatic nucleus and raise the possibility that this neuropeptide may play a role in modulating circadian pacemaker function. PMID- 11274787 TI - Honeycomb-like structure of the intermediate layers of the rat superior colliculus: afferent and efferent connections. AB - There is increasing evidence that acetylcholinesterase is organised in a lattice like fashion in the intermediate layers of the mammalian superior colliculus. In a recent study, we described this organisation in rat by showing that it comprises a well formed honeycomb-like lattice with about 100 cylindrical compartments or modules occupying both the intermediate collicular layers. Considering this enzyme domain as a reference marker for comparing the organisation of collicular input-output systems, the present study investigates whether the principal sensori-motor systems in intermediate layers also have honeycomb-like arrangements. In 33 animals, the distributions of afferents (visual from extrastriate cortex; somatic from the primary somatosensory cortex, the trigeminal nucleus and the cervical spinal cord) and efferents (cells of origin of the crossed descending bulbospinal tract and uncrossed pathway to the pontine gray, the ascending system to the medial dorsal thalamus) were examined in a tangential plane following applications of horseradish peroxidase-wheatgerm agglutinin conjugate (used as an anterograde and retrograde tracer). In 22 of the 33 rats, axonal tracing was made within single tangential sections also stained for cholinesterasic activity in order to compare the neuron profiles with the cholinesterasic lattice.The results show that these afferent and efferent systems are also organised in honeycomb-like networks. Moreover, those related to the cortical, trigeminal and some of the spinal afferents are aligned with the cholinesterasic lattice. Likewise most of colliculo-pontine, colliculo bulbospinal and half of colliculo-diencephalic projecting cells also tend to be in spatial register with the enzyme lattice. This indicates that the honeycomb like arrangement is a basic architectural plan in the superior colliculus for the organisation of both acetylcholinesterase and major sensori-motor systems for orientation. PMID- 11274789 TI - Gene expression of catecholamine biosynthetic enzymes following exercise: modulation by age. AB - Both age and exercise training are associated with tissue specific alterations in the catecholaminergic system. We examined the effect of short-term exercise training on tyrosine hydroxylase and dopamine beta-hydroxylase gene expression in adrenals and specific brain regions with aging. In addition, we examined activator protein-1 and cyclic AMP response element transcription factor binding activity in the adrenal medulla. Male, six- and 24-month-old F-344 rats were exercised by treadmill running for five consecutive days. One group was killed immediately and a second group was killed 2h after the last training session. Exercise significantly elevated tyrosine hydroxylase messenger RNA equally in adrenals of both young and old rats. Training had no effect on dopamine beta hydroxylase messenger RNA in adrenals of young, but levels were elevated in old rats. Binding activities of both activator protein-1 and cyclic AMP response element binding protein were diminished with age in the adrenal medulla. Exercise training had no significant effect on the binding activity of cyclic AMP response element binding protein in either young or old animals, whereas activator protein 1 binding activity increased equally in young and old animals. Exercise training revealed divergent changes in tyrosine hydroxylase messenger RNA in brain catecholaminergic neurons. In the locus coeruleus and the ventral tegmental areas, training elevated tyrosine hydroxylase messenger RNA levels only in young rats. In the substantia nigra, there was no change in young, but a 45% increase in tyrosine hydroxylase messenger RNA in old rats. In the ventral tegmental area, training increased tyrosine hydroxylase gene expression 80% in young but not in old rats. These results indicate that short-term exercise training increases tyrosine hydroxylase messenger RNA levels in young animals in the adrenals, the locus coeruleus and the ventral tegmental area. The responses for exercise training of aged animals differed from the young in brain noradrenergic and dopaminergic nuclei, especially in the substantia nigra, and to some extent in the locus coeruleus and the ventral tegmental area. PMID- 11274788 TI - Age-related changes in levels of tyrosine kinase B receptor and fibroblast growth factor receptor 2 in the rat inferior colliculus: implications for neural senescence. AB - Brain-derived neurotrophic factor and fibroblast growth factor 2, and their respective binding sites, tyrosine kinase B receptor and fibroblast growth factor receptor 2, are known to regulate neurite outgrowth and antioxidant enzyme activity. Several studies suggest that brain-derived neurotrophic factor and fibroblast growth factor are contained in the inferior colliculus. Previous work in our laboratories revealed dendritic and synaptic losses in the inferior colliculus of aged Fischer-344 rats, along with coincident increases in lipid peroxidation possibly linked to a decrease in activity of antioxidant enzymes. In an effort to identify potential causal mechanisms underlying age-related synaptic and dendritic losses that occur in the inferior colliculus, the present study attempted to determine if inferior colliculus levels of tyrosine kinase B receptor and fibroblast growth factor receptor 2 expression are altered with age. Immunocytochemistry was performed in the inferior colliculus, hippocampus and cerebellum of 3-month-old F344 rats to study distributions of the full-length and truncated isoforms of tyrosine kinase B receptor, and fibroblast growth factor receptor 2. The latter two brain regions served as positive controls. For all three antigens, immunolabeling was localized primarily in somata and proximal dendrites in all subdivisions of the inferior colliculus, and in the dentate gyrus and Ammon's horn of the hippocampus. In the cerebellum, the somata and dendrites of the Purkinje cells were also immunolabeled.A significant reduction in levels of the full-length form of tyrosine kinase B receptor in 18- and 25 month-old rats (respectively, approximately 20% and 30% relative to 3-month-olds) was revealed using western blot analyses. Inferior colliculus and hippocampal levels of the truncated form were modestly decreased ( approximately 7%) as well in the two older age groups. In contrast, levels of fibroblast growth factor receptor 2 in the inferior colliculus and hippocampus were elevated by approximately 35% in the two older age groups when compared to 3-month-olds. Changes in cerebellar levels of tyrosine kinase B receptor and fibroblast growth factor receptor 2, while similar to those in the inferior colliculus and hippocampus among the age groups, did not achieve statistical significance in this study. These findings give rise to the possibility that age-related reductions in tyrosine kinase B receptor levels could be a causal factor in the degenerative changes observed in the inferior colliculus of aged animals, including mitochondrial damage and dendritic regression. The observed increases in fibroblast growth factor receptor 2 levels may be compensatory to the increased oxidative stress. The effectiveness of the fibroblast growth factor receptor 2 response is questionable given the damage that occurs in the inferior colliculus and hippocampus of aged animals. However, the deficits could worsen in the absence of an increase in fibroblast growth factor receptor 2. PMID- 11274790 TI - Creatine-supplemented diet extends Purkinje cell survival in spinocerebellar ataxia type 1 transgenic mice but does not prevent the ataxic phenotype. AB - It is not known why expression of a protein with an expanded polyglutamine region is pathogenic in spinocerebellar ataxia, Huntington's disease and several other neurodegenerative diseases. Dietary supplementation with creatine improves survival and motor performance and delays neuronal atrophy in the R6/2 transgenic mouse model of Huntington's disease. These effects may be due to improved energy and calcium homeostasis, enhanced presynaptic glutamate uptake, or protection of mitochondria from the mitochondrial permeability transition. We tested the effects of a 2% creatine-supplemented diet and treatment with taurine-conjugated ursodeoxycholic acid, a bile constituent that can inhibit the mitochondrial permeability transition, on ataxia and Purkinje cell survival in a transgenic model of spinocerebellar ataxia type 1. After 24 weeks, transgenic mice on the 2% creatine diet had cerebellar phosphocreatine levels that were 72.5% of wildtype controls, compared to 26.8% in transgenic mice fed a control diet. The creatine diet resulted in maintenance of Purkinje cell numbers in these transgenic mice at levels comparable to wildtype controls, while transgenic mice fed a control diet lost over 25% of their Purkinje cell population. Nevertheless, the ataxic phenotype was neither improved nor delayed. Repeated s.c. ursodeoxycholic acid injections markedly elevated ursodeoxycholic acid levels in the brain without adverse effects, but provided no improvement in phenotype or cell survival in spinocerebellar ataxia type 1 mice. These results demonstrate that preserving neurons from degeneration is insufficient to prevent a behavioral phenotype in this transgenic model of polyglutamine disease. In addition, we suggest that the means by which creatine mitigates against the neurodegenerative effects of an ataxin-1 protein containing an expanded polyglutamine region is through mechanisms other than stabilization of mitochondrial membranes. PMID- 11274791 TI - Presynaptic regulation of spinal cord tachykinin signaling via GABA(B) but not GABA(A) receptor activation. AB - Internalization of spinal cord neurokinin-1 receptors following noxious stimulation provides a reliable measure of tachykinin signaling. In the present study, we examined the contribution of GABAergic mechanisms to the control of nociceptor processing involving tachykinins. Spinal administration of the GABA(B) receptor agonist R(+)-baclofen in the rat, at antinociceptive doses, significantly reduced the magnitude of neurokinin-1 receptor internalization in neurons of lamina I in response to acute noxious mechanical or thermal stimulation. By contrast, administration of even high doses of the GABA(A) receptor agonists, muscimol or isoguvacine, were without effect. CGP55845, a selective GABA(B) receptor antagonist, completely blocked the effects of baclofen, but failed to increase the incidence of internalization when administered alone. These results provide evidence for a presynaptic control of nociceptive primary afferent neurons by GABA(B) but not GABA(A) receptors in the superficial laminae of the spinal cord, limiting tachykinin release. Because CGP5584 alone did not increase the magnitude of neurokinin-1 receptor internalization observed following noxious stimulation, there appears to be little endogenous activation of GABA(B) receptors on tachykinin-releasing nociceptors under acute stimulus conditions. The contribution of pre- and postsynaptic regulatory mechanisms to GABA(B) receptor-mediated antinociception was also investigated by comparing the effect of baclofen on Fos expression evoked by noxious stimulation to that induced by intrathecal injection of substance P. In both instances, baclofen reduced Fos expression not only in neurons that express the neurokinin-1 receptor, but also in neurons that do not. We conclude that baclofen acts at presynaptic sites to reduce transmitter release from small-diameter nociceptive afferents. Presynaptic actions on non-tachykinin containing nociceptors could similarly account for the reduction by baclofen of noxious stimulus-induced Fos expression in neurokinin-1 receptor-negative neurons. However, the inhibition of Fos expression induced by exogenous substance P indicates that actions at sites postsynaptic to tachykinin- and/or non tachykinin-containing primary afferent terminals must also contribute to the antinociceptive actions of GABA(B) receptor agonists. PMID- 11274793 TI - Prostaglandin EP3 receptor protein in serotonin and catecholamine cell groups: a double immunofluorescence study in the rat brain. AB - Prostaglandin E(2) exerts diverse physiological actions in the central nervous system with unknown mechanisms. We have reported the immunohistochemical localization of the EP3 receptor, one of the prostaglandin E receptor subtypes, in various brain regions including many monoaminergic nuclei. In the present study, a double immunofluorescence technique with an antibody to EP3 receptor and antibodies to markers for monoamine neurons was employed to examine the expression of the receptor in serotonin and catecholamine neurons, and to reveal the distribution of the receptor-expressing monoamine neurons in the rat brain. Almost all serotonergic cells in the medulla oblongata (B1-B4) exhibited EP3 receptor-like immunoreactivity, whereas mesencephalic and pontine serotonergic cell groups (B5-B9) contained relatively small populations of EP3 receptor immunoreactive cells. In the catecholaminergic cell groups, many of the noradrenergic A7 cells in the subcoeruleus nucleus showed immunoreactivity for the receptor. The locus coeruleus exhibited EP3 receptor-like immunoreactivity densely in the neuropil and occasionally in neuronal cell bodies, all of which were immunopositive for dopamine beta-hydroxylase, as observed by confocal laser microscopy. Many of the other noradrenergic and adrenergic cell groups contained small populations of EP3 receptor-like immunoreactive cells. In contrast, no EP3 receptor-like immunoreactivity was detected in the noradrenergic A2 and A4, the adrenergic C2, and all the dopaminergic cell groups. The expression of EP3 receptor by most of the serotonergic, noradrenergic and adrenergic cell groups suggests that prostaglandin E(2) modulates many physiological processes mediated by widely distributed monoaminergic projections through activation of the EP3 receptor on the monoaminergic neurons; for instance, it may modulate nociceptive and autonomic processes by affecting the descending serotonergic pathway from the raphe magnus nucleus to the spinal cord. PMID- 11274792 TI - Differential patterns of nerve growth factor, brain-derived neurotrophic factor and neurotrophin-3 mRNA and protein levels in developing regions of rat brain. AB - The present studies were undertaken to characterize the regional and temporal patterns of neurotrophin messenger RNA and protein levels for beta-nerve growth factor, brain-derived neurotrophic factor and neurotrophin-3 in the developing CNS. We have examined the levels of these neurotrophin messenger RNAs with ribonuclease protection assays and corresponding protein levels with enzyme linked immunosorbent assays in the developing Long-Evans rat hippocampus, neocortex and cerebellum on postnatal days 1, 7, 14, 21, and 92. In addition, immunohistochemistry was used to localize the neurotrophins in these developing brain regions. Results indicated that in neocortex and hippocampus, messenger RNA for both nerve growth factor and brain-derived neurotrophic factor increased in an age-dependent manner, reaching a plateau by postnatal day 14. In the neocortex, nerve growth factor and brain-derived neurotrophic factor protein levels both peaked at postnatal day 14. In hippocampus, nerve growth factor protein peaked at postnatal day 7 while brain-derived neurotrophic factor peaked at postnatal day 14. In cerebellum, nerve growth factor messenger RNA levels were flat, while nerve growth factor protein peaked at postnatal day 7. Brain-derived neurotrophic factor messenger RNA increased in an age-dependent manner while the pattern for its protein levels was mixed. Neurotrophin-3 messeger RNA levels increased in an age-dependent manner in hippocampus, peaked at postnatal day14 in cerebellum, and no changes occurred in neocortex. Neurotrophin-3 protein was at its peak at postnatal day 1 and thereafter decreased at other postnatal days in all three brain regions. Results of neurotrophin immunohistochemistry often paralleled and complemented enzyme-linked immunosorbent assay data, demonstrating specific cell groups containing neurotrophin proteins in these regions. Within each region, patterns with regard to messenger RNA and respective protein levels for each neurotrophin were unique. No consistent relationship between patterns of neurotrophin messenger RNAs and their cognate proteins was observed between regions. The different regional patterns for neurotrophin messengerRNA and protein levels in each brain region indicate that messenger RNA studies of neurotrophin messenger RNA must be augmented by protein determination to fully characterize spatial and temporal neurotrophin distribution. PMID- 11274794 TI - Gene expression and protein distribution of the orexin-1 receptor in the rat brain and spinal cord. AB - Orexins-A and -B are neuropeptides derived from a single precursor prepro-orexin. The mature peptides are mainly expressed in the lateral hypothalamic and perifornical areas. The orexins have been implicated in the control of arousal and appear to be important messengers in the regulation of food intake. Two receptors for orexins have been characterised so far: orexin-1 and -2 receptors. To gain a further understanding of the biology of orexins, we studied the distribution of the orexin-1 receptor messenger RNA and protein in the rat nervous system. We first assessed the expression profile of the orexin-1 receptor gene (ox-r1) in different regions by using quantitative reverse transcription followed by polymerase chain reaction. Using immunohistochemical techniques, we investigated the distribution of orexin-1 receptor protein in the rat brain using a rabbit affinity-purified polyclonal antiserum raised against an N-terminal peptide. The orexin-1 receptor was widely and strongly expressed in the brain. Thus, immunosignals were observed in the cerebral cortex, basal ganglia, hippocampal formation, and various other subcortical nuclei in the hypothalamus, thalamus, midbrain and reticular formation. In particular, robust immunosignals were present in many hypothalamic and thalamic nuclei, as well as in the locus coeruleus. The distribution of the receptor protein was generally in agreement with the distribution of the receptor messenger RNA in the brain as reported previously by others and confirmed in the present study. In addition, we present in situ hybridisation and immunohistochemical data showing the presence of orexin 1 receptor messenger RNA and protein in the spinal cord and the dorsal root ganglia. Finally, due to the shared anatomical and functional similarities between orexins and melanin-concentrating hormone, we present a comparison between the neuroanatomical distribution of the orexin-1 receptor and melanin concentrating hormone receptor protein-like immunoreactivities in the rat central nervous system, and discuss some functional implications. In conclusion, our neuroanatomical data are consistent with the biological effects of orexins on food intake and regulation of arousal. In addition, the data suggest other physiological roles for orexins mediated through the orexin-1 receptor. PMID- 11274795 TI - Isolation and characterization of an alpha 2-type zebrafish glycine receptor subunit. AB - The complementary DNA for a novel alpha subunit of the glycine receptor, alphaZ2, was isolated from a zebrafish adult brain library. The molecular characteristics, phylogenetic relationships and messenger RNA length of this alphaZ2 subunit show it to be an alpha2-type glycine receptor subunit isoform. The leader peptide however, diverges from those of known glycine receptor alpha isoforms. Recombinantly expressed in Xenopus oocytes, alphaZ2 formed functional glycine receptor channels. These homomeric channels were activated by glycine and taurine, with apparent affinities similar to those reported for zebrafish alphaZ1 glycine receptor, and were also effectively antagonized by nanomolar concentrations of strychnine. However, during prolonged applications of agonists, ionic currents of alphaZ2 receptor channels declined to a much lower steady-state level than those of alphaZ1, indicating different desensitization properties. Analysis of messenger RNA revealed that alphaZ2 is specifically expressed in adult brain tissue and present in both adult and embryonic zebrafish. This report contributes to the characterization of the diversity of glycine receptor isoforms in vertebrates. PMID- 11274797 TI - Defining the concentration gradient of nerve growth factor for guided neurite outgrowth. AB - The developing axon is believed to navigate towards its target tissue in response to a concentration gradient of neurotrophic factors, among other diffusible and surface-bound stimuli. However, the minimum concentration gradient required for guidance over the maximum distance is still unknown, largely because well-defined systems have not been utilized to address this question. In this study, a linear concentration gradient of nerve growth factor was achieved across a 5-mm agarose membrane that separated a nerve growth factor source compartment from a sink compartment. The concentrations in both compartments were maintained constant (and different). Both concentration and concentration gradient were well defined across the membrane, allowing us to study the relative importance of concentration gradient vs concentration for neurite guidance. The orientation of PC12 cell neurites was studied in response to a series of nerve growth factor concentration gradients in vitro. For effective guidance of PC12 cell neurite outgrowth, a minimum concentration gradient of 133ng/ml per mm was required, below which guidance was ineffective. Higher gradients were effective for guidance yet were limited by the concentration of nerve growth factor in the source compartment. At a nerve growth factor concentration of 995ng/ml, the PC12 cells' receptors were saturated, thereby limiting the maximum effective distance for guidance to less than 7.5mm in response to a diffusible nerve growth factor cue. This distance exceeds the 0.5-2mm distance observed by others for effective neurite guidance. Using this model system, we propose that the minimum concentration gradient can be defined for other cells and growth factors. Ultimately, it is anticipated that such concentration gradients could be included in a device to promote regeneration. PMID- 11274796 TI - Regional distribution of glycine receptor messenger RNA in the central nervous system of zebrafish. AB - We report the cloning of the zebrafish beta subunit of the glycine receptor and compare the anatomical distribution of three glycine receptor subunit constituents in adult zebrafish brain (alphaZ1, alphaZ2 and betaZ) to the expression pattern of homologous receptor subunits (alpha1, alpha2 and beta) in the mammalian adult CNS. Non-radioactive hybridization was used to map the distribution of the alphaZ1, alphaZ2 and betaZ glycine receptor subunit messenger RNAs in the adult zebrafish brain. The anterior-posterior expression gradient found in adult zebrafish brain was similar to that reported in mammalian CNS. However, the glycine receptor transcripts, notably the alphaZ1 subunit, were more widely distributed in the anterior regions of the zebrafish than in the adult mammalian brain. The isoform-specific distribution pattern was less regionalized in zebrafish than in the rat mammalian CNS. Nevertheless, there was some regionalization of alphaZ1, alphaZ2 and betaZ transcripts in the diencephalic and mesencephalic nuclei where different sensory and motor centers express either alphaZ1/betaZ or alphaZ2 subunits. In contrast to the widespread distribution of the beta subunit in adult mammalian brain, alphaZ2 messenger RNA presented the widest expression territory of all three glycine receptor subunits tested. alphaZ2 messenger RNA was expressed in the absence of alphaZ1 and betaZ messenger RNA in the outer nuclear layer of the retina, the inferior olive and the raphe of the medulla oblongata, as well as in the nucleus of Cajal of the medulla spinalis. In contrast, an identified central neuron of the reticular formation, the Mauthner cell, expresses all three glycine receptor subunits (alphaZ1, alphaZ2 and betaZ). This report extends the already described glycine receptor expression in the vertebrate CNS and confirms the importance of glycine-mediated inhibition in spinal cord and brainstem. PMID- 11274798 TI - Artificial electrotonic coupling affects neuronal firing patterns depending upon cellular characteristics. AB - While there have been numerous theoretical studies indicating that electrotonic coupling via gap junctions interacts with the intrinsic characteristics of the coupled neurons to modify their electrical behaviour, little experimental evidence has been provided in coupled mammalian neurons. Using an artificial electrotonic junction, two distant uncoupled neurons were coupled through the computer, and the coupling conductance was varied. Tonically firing CA1 hippocampal pyramidal neurons reduced their spike firing frequency when coupled to thalamic or pyramidal cells, showing that the electrical coupling can be considered as a low-pass filter. The strength of coupling needed to entrain spike bursts of pyramidal neurons was considerably lower than the coupling needed to synchronize two neurons with different cellular characteristics (thalamic and pyramidal cells). Coupling promoted burst firing in a non-bursting cell if it was coupled to a spontaneously bursting neuron. These results support modelling studies that indicate a role for gap-junctional coupling in the synchronization of neuronal firing and the expression of low-frequency bursting. PMID- 11274800 TI - Use of ultrasound to prepare lipid emulsions of lorazepam for intravenous injection. AB - Lipid emulsions can be used as a vehicle for the production of low-volume injectable preparations with minimally water-soluble active ingredients. First, we focus on the galenic and technological conditions established by ultrasound techniques. A 2(5) factorial design was used to optimize the carrier emulsion. The study then deals with the development of a parenteral emulsion formulation for lorazepam (1 mg/ml), which is compared with the highest concentration (0.05 mg/ml) achieved in the optimal aqueous diluent for lorazepam (dextrose 5% in water). The physical and chemical stability of lorazepam in the emulsion was examined for 7 months. PMID- 11274801 TI - A multiparticulate drug-delivery system based on pellets incorporated into congealable polyethylene glycol carrier materials. AB - As a novel alternative to the incorporation into hard gelatin capsules or tablets, extended-release (Aquacoat- or Eudragit RS-coated) or enteric (Eudragit L-coated) pellets were embedded into congealed tablet-shaped PEG-plugs of different molecular weights, which rapidly released the pellets upon contact with aqueous fluids. The lower-molecular-weight PEGs (600 and 1000) were not suitable carrier materials: they dissolved the coatings or significantly increased their permeability. The release characteristics of the original pellets were maintained after embedding the pellets into the higher-molecular-weight PEGs 4000 or 10000. The shelf-life stability was a function of storage temperature and coating material. Stored at 40 degrees C, Aquacoat-coated pellets embedded in PEG 4000 exhibited a decreased drug release because of curing effects, while storage at 20 degrees C or below resulted in stable release profiles over a 3 month period. Eudragit RS-coated pellets, stored at room temperature or above, showed an increased release, and the carrier material possibly migrated into the film, thus increasing its permeability. At 4 degrees C, the release was stable over a 6 month period. PMID- 11274802 TI - Pharmacokinetic behavior of cyclosporin A in rabbits by oral administration of lecithin vesicle and Sandimmun Neoral. AB - The present study was undertaken to investigate the incorporation of lipophilic polypeptide, cyclosporin A (CsA) into lecithin vesicular system and to compare its pharmacokinetics behavior with Sandimmun Neoral (CsA-NEO). Lecithin vesicles of cyclosporin A (CsA-VES) were prepared by the rotary evaporation method, treated further with sonication. Studies were carried out to characterize the vesicles on physical properties, content, entrapment efficiency, particle size, polydispersity and Zeta potential. Pharmacokinetic behaviors were studied in rabbits at dose of 30 mg/kg. Results showed CyA vesicles were spherical particles, with content of 3.137+/-0.060% mg/ml, entrapment efficiency of 98.91+/ 0.80%, particle size of 63.89+/-4.75 nm, polydispersity of 43.2+/-6.1% and Zeta potential of -13 mV. The best model fitting experimental data was a two compartment open model with first-order kinetics. The relative bioavailability of CsA-VES versus CsA-NEO was 105+/-21% (n=6) and statistical analysis demonstrated both preparations were bioequivalent. In conclusion, lecithin vesicles are promising carriers in the oral delivery of CsA, considering their absorption enhancement effect and low-toxic property. PMID- 11274803 TI - Stabilisation of natural anthocyanins by micellar systems. AB - In this paper, we discuss the influence of different micellar systems on the degradation of natural anthocyans, either glycosides and aglycones, at pH values ranging from 2.8 to 6.0. The interaction of anthocyanins, in suitable dispersed systems such as negative micelles of sodium dodecylsulphate (SDS), consistently increased their chemical stability in aqueous solutions. The results of these experiments point out how both the number of available negative charges and the presence of an organised distribution of the negative charges on the micellar surface appear to be necessary conditions to achieve the anthocyanins' stability and colour retention. The sodium dodecylbenzensulphonate (SDBS), containing an aromatic ring near the negative surface of the micelle, seems to increase the rate of decomposition. Preliminary findings of circular dicroism (CD) investigation allowed us to hypothesise that these pigments undergo an intermolecular self-association process induced by the SDS micelles and this phenomenon presumably contribute to increase stability. PMID- 11274804 TI - Solubility prediction of salmeterol xinafoate in water--dioxane mixtures. AB - The mole fraction solubility of salmeterol xinafoate was determined in various concentrations of dioxane in aqueous binary mixture. Maximum solubility was observed in 90% v/v dioxane and solubility parameter of the solute was estimated from solubility peak equal to 24.99 MPa(0.5). The predicting capability of four different cosolvency models was also evaluated employing a five data point training set. The solubility data at other cosolvent concentrations were predicted using the trained models, with percentage average errors for 28 drug solubility data sets in water-cosolvent mixtures lying between 12.5 and 15.0%. Further predictive model is proposed for accurate solubility predictions based on a minimum number of experiments. The percentage average error where tested was 10.6%. PMID- 11274806 TI - ATR-FTIR spectroscopic investigations on the effect of solvents on the permeation of benzoic acid and salicylic acid through silicone membranes. AB - The effect of a series of alcohols on the permeation of salicylic acid (SA) and benzoic acid (BA) through silicone membrane was evaluated, using Franz-type diffusion cells. Although permeants were applied at the same thermodynamic activity in all vehicles, the resulting fluxes were found to differ significantly. This was a consequence of the interactions between the vehicles and the membrane. The interactions between the vehicles and the membrane were further investigated using ATR-FTIR spectroscopy. With this technique, it was possible to identify two different diffusion processes when the membrane was pre treated with buffer, whereas one single diffusion process was observed when the membrane was pre-soaked with the vehicle. The technique was successfully used to deconvolute the relative magnitude of partition and diffusion in the permeation process. It was shown that the permeation of both acids was affected by the effect of the vehicles on the diffusion coefficient and the partition coefficient in the silicone membrane. The solubility of the drug in the impregnated membrane was found to be proportional to the saturated solubility in the vehicle used to treat the membrane. The solubility of BA in the impregnated silicone membrane was twice that of SA. PMID- 11274805 TI - The effect of co-spray drying with polyethylene glycol 4000 on the crystallinity and physical form of lactose. AB - The effect of spray drying lactose alone and in the presence of polyethylene glycol 4000 was investigated. Lactose was added to distilled water to give concentrations of 10, 20, 30 and 40g/100ml at room temperature and each spray dried in turn. Identical samples were prepared to which polyethylene glycol (PEG) 4000 was added (12% by weight of lactose) prior to spray drying. Microcalorimetric and X-ray diffraction studies showed that spray drying lactose solutions produced completely amorphous material due to rapid solidification during the spray drying process, whereas lactose suspensions yielded partially crystalline products due to crystalline material that remained in suspension. However, all the PEG/lactose (12%w/w) co-spray dried products were found to be crystalline. It can be inferred that the solidification rates of the lactose in the presence of PEG must have been slower than that of lactose alone which allowed PEG and lactose to crystallize. The PEG/lactose products that were spray dried from solution consisted of alpha-anhydrous, alpha-monohydrate, beta-lactose and PEG extended chain polymorph, whereas those formed from suspension PEG/lactose samples consisted of only alpha-anhydrous, alpha-monohydrate and extended chain PEG crystals. PEG probably caused the more concentrated lactose suspensions to crystallize slowly due to the strong hydrogen bonding between PEG and water, which allowed growth on the alpha-lactose seed crystals. PMID- 11274807 TI - Pharmacokinetics of tetramethylpyrazine in rat blood and brain using microdialysis. AB - Since the central nervous acting agent, tetramethylpyrazine, is reported to have appreciable blood-brain barrier penetrability, a design allowing simultaneous and continual monitoring of drug concentrations in blood and brain was employed to study the distribution of intravenously administered tetramethylpyrazine (10 mg kg(-1)). The system consisted of two microdialysis probes, each optimally constructed for sampling of the respective body fluids, inserted into the right jugular vein and striatum of male Sprague--Dawley rats. The probes were perfused with appropriate media at rates optimized for recovery. Dialysates were automatically collected using a microfraction collector and drugs were analyzed by high performance liquid chromatography (HPLC) with ultra violet (UV) detection. Results indicate that both blood and brain pharmacokinetics of unbound tetramethylpyrazine fit best to a two-compartment model. The elimination half life of tetramethylpyrazine in rat blood and brain were 82.1 and 184.6 min, respectively. Increasing brain/blood concentration ratios suggested that tetramethylpyrazine effectively penetrated the blood--brain barrier. PMID- 11274808 TI - Use of ion-exchange resins to prepare 100 microm-sized microcapsules with prolonged drug-release by the Wurster process. AB - Ion-exchange resin (IER)--drug complexes were used as core materials to explore their capability to prepare a 100 microm-sized, highly drug-incorporated microcapsule with a prolonged drug release by the Wurster process. Diclofenac sodium was loaded into Dowex 1-X2 fractionated into 200--400 mesh and subsequently microencapsulated with two types of aqueous colloidal polymer dispersion, Aquacoator Eudragit RS30D. The mass median diameter and drug content of the microcapsules thus obtained were 98 microm and 46% with Aquacoat, and 95 microm and 50% with Eudragit RS30D, respectively. Each microcapsule was obtained at a product yield of 94%. The rate of drug release from the microcapsules was highly dependent on the encapsulating materials. For the microcapsules coated with Aquacoat, diclofenac sodium was found to be rapidly released over 4 h, even at a 25 wt% coating level because of cracks on the microcapsule surfaces resulting from the swelling stress of the drug-loaded IER cores. In contrast, significantly prolonged drug-release was achieved in the microcapsules prepared with Eudragit RS30D: even such a very low coating level as 3 wt% provided an exceptionally prolonged drug-release over 24 h. The results indicated that the use of IER along with a flexible coating material would be a feasible way to prepare a prolonged release type of microcapsules with a diameter of 100 microm and a drug content of more than 50% by the Wurster process. PMID- 11274809 TI - Activity of pancreatic endopeptidases towards luteinizing hormone-releasing hormones. AB - LHRH and its analogues have low oral bioavailability; this is in part due to their degradation by peptidases present in the intestinal lumen. To determine the appropriate inhibitors to co-administer with LHRH oral formulations, the peptidases involved in their digestion have to be identified. Human (hLHRH) and salmon (sLHRH) LHRH analogues contain a number of potential cleavage sites for the lumenal pancreatic secreted serine endopeptidases: chymotrypsin, trypsin and elastase. The rate of LHRH degradation by equimolar concentrations of chymotrypsin, trypsin and elastase were examined separately in vitro, at pH 8.0, 15 degrees C. At a molar ratio of 1:1000 (enzyme:LHRH), both LHRH analogues were rapidly hydrolysed by alpha-chymotrypsin with half-lives of 2.5+/-0.3 and 2.7+/ 0.4 min (mean+/-S.D., n=3), respectively, whereas in the presence of elastase both LHRH analogues were slowly hydrolysed with half-lives of 90+/-15 and 114+/ 21 min (mean+/-S.D., n=3), respectively. Trypsin had no activity towards either LHRH analogues after 2 h incubation. The degradation of the LHRH analogues by elastase is likely to be a property of the chymotrypsin impurity. It is concluded that protection of the LHRH analogues from alpha-chymotrypsin is a requirement for the development of oral absorbable product. PMID- 11274810 TI - Effect of drug lipophilicity on in vitro release rate from oil vehicles using nicotinic acid esters as model prodrug derivatives. AB - The rate constants for transfer of a homologous series of nicotinic acid esters from oil vehicles to aqueous buffer phases were determined using a rotating dialysis cell. The chemical stability of butyl nicotinate has been investigated at 60 degrees C over pH range 0.5--10. Maximum stability occurs at pH 4--5 and an inflection point was seen around the pK(a). For the nicotinic acid esters, a linear correlation was established between the first-order rate constant related to attainment of equilibrium, k(obs) and the apparent partition coefficient, P(app): log k(obs)=-0.83log P(app)+0.26 (k(obs) in h(-1), n=9). For hexyl nicotinate with a true partition coefficient of 4 it was possible to determine k(obs) by decreasing pH in the aqueous release medium to 2.05. Thus, under the latter experimental conditions estimation of the relative release rates for the esters were performed. The ratio between the specific rate constant k(ow), related to the transport from oil vehicle to aqueous phase, for ethyl and hexyl nicotinate was 139. The hydrophobic substituent constant for a methylene group, pi(CH(2)), was determined for nicotinic acid esters in different oil/buffer partitioning systems to 0.54--0.58. Addition of hydroxypropyl-beta-cyclodextrin to the aqueous release medium did not enhance the transport rate of the esters from the oil phase. PMID- 11274811 TI - Flow-through UV spectrophotometric sensor for determination of (acetyl)salicylic acid in pharmaceutical preparations. AB - The solid phase spectrophotometry technique, in which the absorbance of the species of interest sorbed on a solid support is measured directly, was applied to the determination of salicylic acid using flow injection-analysis. Salicylic acid was determined by monitoring of its intrinsic absorbance at 297 nm sorbed on Sephadex QAE A-25 resin placed in an appropriate flow-through cell. The method proposed improves the selectivity compared with the corresponding solution-phase method and the sensitivity is increased by a factor of 30 or more. The flow through sensor proposed allows working with several calibration lines simply by varying the sample volume injected. Thus, linear dynamic ranges from 1 to 20 and from 2 to 40 microg ml(-1) can be obtained by using 1000 and 300 microl, respectively, with detection limits being 0.064 and 0.135 microg ml(-1). Relative Standard Deviations (RSDs) of 0.52 and 0.38%, and sampling frequencies of 18 and 25 h(-1), respectively, were also achieved. The sensor also allows the indirect determination of acetylsalicylic acid previous hydrolysis on-line to salicylic acid. For acetylsalicylic acid, a linear dynamic range from 5 to 120 microg ml( 1) and 25 h(-1) of sampling frequency (300 microl of sample volume) were obtained. The proposed flow-through sensor has been successfully applied to the determination of both analytes in pharmaceutical preparations. PMID- 11274812 TI - Liposomes with phosphatidylethanol as a carrier for oral delivery of insulin: studies in the rat. AB - In this study, phosphatidylethanol formed by phospholipase D catalysed transphosphatidylation of phosphatidylcholine was employed as a component for preparation of liposomal carrier for oral delivery of insulin. Thermotropic behaviour of liposomes from mixtures of dipalmitoyl phosphatidylcholine and dipalmitoyl phosphatidylethanol, and their resistance to pancreatic phospholipase A(2) catalysed hydrolysis were studied. Three kinds of liposomes with insulin were prepared to examine the pharmacological availability of liposomes with phosphatidylethanol: (i) dipalmitoyl phosphatidylcholine/dipalmitoyl phosphatidylethanol (1:1 w/w) liposomes; (ii) dipalmitoyl phosphatidylcholine/dipalmitoyl phosphatidylethanol/palmitoyl-stearoyl sucrose (1:1:0.2) liposomes; and (iii) liposomes composed of natural phosphatidylcholine and phosphatidylinositol (1:1). The resultant liposomes were orally administrated to rats with blood glucose concentration of 270 mg/100 ml in a dose of 12 IU/kg body weight. Blood samples were collected 0.5, 1.5, 3, 5, and 24 h after treatment. Oral administration of all liposomal species resulted in hyperinsulinemia. Hyperinsulinemia induced by liposomes containing dipalmitoyl phosphatidylethanol was attended by a decrease of blood glucose concentration. No correlation between insulin level and glucose concentration in the rat blood after oral administration of phosphatidylinositol-containing liposomes was observed. PMID- 11274813 TI - Influence of admixed carboxymethylcellulose on release of 4-aminopyridine from hydroxypropyl methylcellulose matrix tablets. AB - Among different technological variables that influence drug release from hydrophilic matrices, the use of mixtures of polymers represents a potential way of achieving a variety of release properties. Tablets of the model drug 4 aminopyridine with hydroxypropyl methylcellulose were prepared with different proportions of polymer content as well as with different proportions of admixed carboxymethylcellulose (CMC) in the range up to 35% (based on the total polymer content). The matrices release behavior was examined by absorption of samples at 261 nm (USP 23 apparatus 2, paddle, at 50 rpm) using 0.1 N HCl and 0.2 M phosphate buffer as dissolution media. Increasing proportions of CMC in the polymer mixture lead to decreasing dissolution rates, in a range of k=0.094-0.036 for HCl and k=0.044--0.009 for phosphate buffer. The release mechanism in HCl is predominantly controlled by diffusion (n=0.46--0.62), while in phosphate buffer it is controlled, as reported previously, by diffusion/relaxation (n=0.58--0.85) and near zero order release at high CMC concentrations. Approximately doubling the total polymer content gives lower release rates for HCl in the range k=0.038- 0.015 and for phosphate buffer k=0.0099--0.0034. Near zero order release is observed only at pH 7.4 (n=0.79--0.96). Decreasing release constant values show a logarithmic relationship with increasing values of the exponent n. This indicates that zero-order release occurs with sufficiently reduced release rate. PMID- 11274814 TI - Effect of disintegrants with different hygroscopicity on dissolution of Norfloxacin/Pharmatose DCL 11 tablets. AB - This paper reports the effect of disintegrant hygroscopicity on dissolution of tablets obtained by compression at 85 MPa of mixtures of Norfloxacin and different proportions of a disintegrant (Starch 1500, PVP XL 10 or Croscarmellose sodium) and a diluent (Pharmatose DCL 11). Dissolution behavior was evaluated according to USP 23, apparatus 2 (paddle) at 50 rpm and using 750 ml acetate buffer solution of pH 4, at 37 degrees C, as medium. Norfloxacin added of increasing proportions, in a given range, of each disintegrant or the diluent increased the drug dissolved. Addition of increasing proportions of Pharmatose DCL 11 to Norfloxacin with 5% of the high hygroscopic Starch 1500 reduced the dissolution improvement effect of Pharmatose DCL 11. Addition of 5% Pharmatose DCL 11 to tablets of the middle hygroscopic Croscarmellose sodium and Norfloxacin slightly reduced the Croscarmellose sodium dissolution promoting effect, while addition of 15% Pharmatose DCL 11 to tablets of the low hygroscopic PVP XL 10 and Norfloxacin showed no inhibition but potentiated substantially the dissolution of Norfloxacin. These effects were attributed to competition for the available water in the tablet and to different water consume, for dissolution or hydration, by the diluent and the disintegrants. PMID- 11274815 TI - Effect of magnesium stearate or calcium stearate as additives on dissolution profiles of diltiazem hydrochloride from press-coated tablets with hydroxypropylmethylcellulose acetate succinate in the outer shell. AB - Effect of magnesium stearate (MgSt) or calcium stearate (CaSt) on the dissolution profiles of diltiazem hydrochloride in the core of press-coated (PC) tablets with an outer shell composed of hydroxypropylmethylcellulose acetate succinate (HPMCAS) was evaluated by porosity and changes in IR spectra of tablets. In JP first fluid (pH 1.2), the lag time increased with decreasing porosity and was greatest by the addition of MgSt to HPMCAS. While, in JP second fluid (pH 6.8), it increased with decreasing porosity by the addition of CaSt, but hardly changed by the addition of MgSt. Thus, using tablets prepared with the same composition as the outer shell, the changes in IR spectra and uptake amount of the dissolution media after immersion in first fluid and second fluid were determined. The results suggested that some physicochemical interaction occur between MgSt and HPMCAS in tablets with HPMCAS and MgSt and the uptake increased markedly in each dissolution medium. These phenomena seem to cause a prolongation of lag time in first fluid but a shortening of it in second fluid in PC tablets with HPMCAS and MgSt. In contrast, CaSt and HPMCAS did not show such interactions and increased the hydrophobic properties of the outer shell. Consequently, the lag time was only slightly prolonged in first fluid, however, markedly prolonged in second fluid due to suppression of second fluid penetration into micro pores in the outer shell and HPMCAS gel formation on the surface in PC tablets with HPMCAS and CaSt. PMID- 11274816 TI - Use of a capillary rheometer to evaluate the rheological properties of microcrystalline cellulose and silicified microcrystalline cellulose wet masses. AB - The influence of microcrystalline cellulose (MCC) type and water content on the rheological properties of the wet powder masses were studied using two different MCC grades (Avicel and Emcocel) and silicified microcrystalline cellulose (SMCC, Prosolv). A ram extruder was used as a capillary rheometer and unique flow curves for each cellulose grade and moisture content were derived. In addition, the elastic parameters of recoverable shear and compliance were determined. From different flow curve models evaluated, it was not possible to obtain clear evidence, which model best described the rheological properties of each cellulose grade at each water level. Furthermore, the residuals were shear rate dependent, which indicates that the models do not perfectly agree with physical properties of the wet masses. The elastic properties of wet masses increased with increasing water content and decreased with increasing shear stresses. SMCC grade proved to be more elastic than the simple MCC grades at each moisture content. Thus, the rheological properties of MCC and SMCC wet masses were different and changed with water content. Consequently, it was not possible to achieve similar rheological properties between different grades of cellulose by altering the water content of the wet mass. PMID- 11274817 TI - Correlation between loose density and compactibility of granules prepared by various granulation methods. AB - The objectives of this study were to prepare the lactose granules by various granulation methods using polyethylene glycol 6000 (PEG 6000) as a binder and to evaluate the effects of granulation methods on the compressibility and compactibility of granules in tabletting. Lactose was granulated by seven granulation methods -- four wet granulations including wet massing granulation, wet high-speed mixer granulation, wet fluidized bed granulation and wet tumbling fluidized bed granulation; and three melt granulations including melt high-speed mixer granulation, melt fluidized bed granulation and melt tumbling fluidized bed granulation. The loose density, angle of repose, granule size distribution, mean diameter of granules, and the tensile strength and porosity of tablets were evaluated. The compactibilities of granules were varied by the granulation methods. However, the difference in compactibility of granules could not be explained due to the difference in compressibility, since there was no difference in Heckel plots due to granulation methods. Among their granule properties, the loose density of granules seemed to have a correlation with the tablet strength regardless of the granulation methods. PMID- 11274818 TI - An easy producible new oral hydrocolloid drug delivery system with a late burst in the release profile. AB - One of the main drawbacks of hydrocolloid matrices as oral controlled drug delivery systems is the often observed decreasing rate of drug release at the end of the release process. This study describes a new pH-controlled hydrocolloid drug delivery system consisting of a neutral cellulose ether as basis polymer and enteric coating materials as additives. The new dosage form is able to accelerate the drug release at a predetermined pH. In a typical example, methylhydroxy ethylcellulose, MHEC 10000 B, was used as the basis polymer and hydroxypropyl methylcellulose acetate succinate, HPMCAS HF, as release modifier. The new delivery system is characterized by its homogeneous structure and easy production by direct compression of the components. The acceleration is well reproducible. Furthermore the new formulation shows high stability against hydrodynamic stress and tolerates ionic strengths up to 0.25 without any significant changes in the release profile. As mechanism of the final burst at pH values >5.7, enforced erosion of the gel layer surrounding the tablet core, could be identified. PMID- 11274819 TI - Neurotropic, immunological and gastric effects of low doses of Atropa belladonna L., Gelsemium sempervirens L. and Poumon histamine in stressed mice. AB - Previous studies realized in the laboratory have indicated that application of experimental stress (such as unavoidable footshock) induced significant behavioral, gastric and immunological alterations in mice. The aim of this study was to evaluate effects of low doses of Atropa belladonna L., Gelsemium sempervirens L. and Poumon histamine on stress-induced behavioral, immunological and gastric alterations. Locomotor, postural and exploratory activities have been evaluated by two behavioral tests: light/dark box and staircase tests. Immunological studies were investigated to count white blood cells subpopulations (lymphocytes, neutrophils, monocytes and basophils) by coulter counter. The severity of gastric erosions was evaluated by microscopic technique in mice after experimental stress. The results have demonstrated that low doses of G. sempervirens L. and A. belladonna L. had a significant neurotropic and protective effects on behavioral and gastric alterations induced by experimental stress. The immunological protective effects observed were probably induced via their neurotropic effects. The P. histamine showed a significant immunoprotective and gastroprotective effect in mice exposed to experimental stress. PMID- 11274820 TI - Antimicrobial activity of certain Indian medicinal plants used in folkloric medicine. AB - Fifty medicinal plants belonging to 26 families were studied for their antimicrobial activity. Among 50 plants tested, 72% showed antimicrobial activity. About 22 plant extracts from 15 families exhibited activity against both Gram-positive and Gram-negative bacteria. Fourteen plants belonging to 11 families did not show activity against any of the bacteria tested. Only nine plant extracts showed antifungal activity. The bulb extracts of A. cepa and A. sativum exhibited activity against both filamentous and non-filamentous fungus. Eight plant extracts belonging to seven families exhibited both antibacterial and antifungal activity. PMID- 11274821 TI - Toxicity studies of Rhizoma Polygonati Odorati. AB - The toxicity of the water-soluble extract of Rhizoma Polygonati Odorati was studied in the present investigation, which included its acute toxicity, chronic toxicity, and genetic toxicity. The aqueous extract of Rhizoma Polygonati Odorati did not cause seriously abnormal signs or death to animals in the acute toxicity test and in the 6-month chronic toxicity test. Neither was genetic toxicity found in the Ames test, the micronucleus test of bone marrow and the sperm malformation test in mice. PMID- 11274822 TI - Hypoglycemic effects of aqueous extract of Rhizoma Polygonati Odorati in mice and rats. AB - Water soluble extract of Rhizoma Polygonati Odorati (RPO) was studied for its hypoglycemic effect in diabetic mice and rats. Results showed that RPO significantly lowered hyperglycemia caused by starch loading in both normal and diabetic mice. Four week's administration with RPO reduced fasting blood glucose, decreased glycosylated hemoglobin (GHb), and improved the glucose tolerance in diabetic mice. In diabetic rats complicated with hyperlipemia, RPO prevented and reduced both hyperglycemia and hypertriglyceridemia. The results support the view that RPO may influence glucose or carbohydrate metabolism of diabetic animals in many ways including inhibiting the activity of alpha-glucosidase in digestive canal, and improving the metabolism of glucose and triglyceride. PMID- 11274824 TI - In vitro hemolysis of human erythrocytes -- by plant extracts with antiplasmodial activity. AB - Human erythrocytes were exposed in a dose dependent manner to various ethanolic plant extracts, and fractions obtained from plant parts of Calotropis procera (Ait.) R. Br. and the gum--oleo resin of Commiphora wightii (Arnott.) Bhand. These have been screened for in vitro schizontocidal activity and graded with respect to their 50% inhibitory concentration (IC(50)) derived from the twofold serial dilution of the dose range 0.0625--2 mg/ml. An attempt had been made to relate their antiplasmodial activity with their cytotoxicity as represented by the in vitro rate of hemolysis. Intact erythrocytes were found to respond with a dose--time-integral and fitted to models of pseudo first-order reaction, Michaelis--Menten equation and Hill equation with k(1), k(2) and k(3) as their rate constants, respectively. Hemolysis isotherms of flower and root of C. procera and gum--oleo resin of C. wightii extracts were representative. Erythrocytic membrane instability is possibly a major factor as has been earlier reported with ethanol and chloroquine for the cytotoxicity of these plant extracts. PMID- 11274823 TI - Cyclooxygenase inhibiting and anti-bacterial activities of South African Erythrina species. AB - Aqueous, ethanolic and ethyl acetate extracts of the bark and leaves of five South African Erythrina species Erythrina caffra, Erythrina humeana, Erythrina latissima, Erythrina lysistemon and Erythrina zeyheri were screened for prostaglandin synthesis-inhibitory and anti-bacterial activity. The bark generally displayed higher activity than the leaves in both bioassays. The highest cyclooxygenase inhibiting activity and anti-bacterial activity was recorded for the ethanol and ethyl acetate bark extracts of E. caffra, E. latissima and E. lysistemon. An anti-bacterial compound, 4',5,7-trihydroxy-6 prenylisoflavone, was isolated by bioassay-guided fractionation from bark of E. lysistemon. PMID- 11274825 TI - Antiimplantation and pregnancy interruption efficacy of Rivea hypocrateriformis in the rat. AB - Petroleum ether, chloroform, ethanol and distilled water extracts of the aerial parts of the plant Rivea hypocrateriformis (Convolvulaceae) were tested for antiimplantation and pregnancy interruption properties in female albino rats. Among these, the ethanol extract was found to be most effective in causing significant antiimplantation and interruption of early pregnancy. The antifertility activity of ethanol extract was reversible on exogenous administration of hydroxy progesterone. However, the same ethanol extract was found to be ineffective in interruption of late pregnancy. Among the four extracts subjected to preliminary phytochemical screening, the active ethanol extract showed positive tests for alkaloids, glycosides, saponins, tannins and phenolic compounds. PMID- 11274826 TI - Screening of Nepalese medicinal plants for antiviral activity. AB - In an ethnopharmacological screening, plants used in Nepalese traditional medicine were evaluated for antiviral activity. Methanolic and methanolic-aqueous extracts derived of 23 species were assayed in two in vitro viral systems, influenza virus/MDCK cells and herpes simplex virus/Vero cells. Two species, Bergenia ligulata and Nerium indicum showed the highest antiinfluenzaviral activity with 50% inhibitory dose of 10 microg/ml. Holoptelia integrifolia and N. indicum exhibited considerable antiviral activity against herpes simplex virus. None of these extracts showed cytotoxic effects. Additionally for B. ligulata and H. integrifolia partial protease inhibitory activity was estimated. PMID- 11274827 TI - Anthelmintic screening of Zimbabwean plants traditionally used against schistosomiasis. AB - Extracts of 23 plant species used popularly against schistosomiasis in Zimbabwe were screened for their anthelmintic effect. Schistosomules of the trematode Schistosoma mansoni and cysticercoids of the cestode Hymenolepis diminuta were studied in vitro. The material consisted of 58 plant extracts, of which 37 killed the newly excysted cysticercoids within an hour, when incubated in a culture medium. Lethal concentrations varied from 0.8 to 103 mg/ml. All plant extracts showed activity against the tapeworms after 24 h. Ten of the best extracts were also tested against schistosomules. Five of these extracts showed activity. Lethal concentrations varied from 0.6 to 33.8 mg/ml of dry plant material. Extracts of stem and root from Abrus precatorius (Fabaceae), of root bark and leaves from Ozoroa insignis (Anacardiaceae) and of root bark from Zizyphus mucronata (Rhamnaceae) gave the best results against tapeworms. The best results against schistosomules were obtained with stem and root extracts from Abrus precatorius (Fabaceae) and stem bark from Elephantorrhiza goetzei (Mimosaceae). Although the activity of root and root bark extracts commonly used in traditional medicine was verified in this study, our results showed that also extracts from leaf and stem can be effective anthelmintics. PMID- 11274829 TI - Bioactive chemical constituents from Alchornea laxiflora (benth) pax and hoffman. AB - Quercetin-7,4'-disulphate (1) quercetin (2) quercetin-3',4'-disulphate (3) quercetin-3,4'-diacetate (4) rutin (5) and quercetrin (6) were isolated from the ethyl acetate soluble fraction (ESM) of the crude methanolic leaf extract of Alchornea laxiflora. Purification of these compounds was carried out by column chromatography utilising sephadex LH 20 and various mixtures of water, methanol, ethanol and toluene as eluents. Structural elucidation was by UV, IR, (1)HNMR and (13)CNMR spectroscopy as well as by FAB-MS. Antimicrobial activity of isolated compounds was detected in Gram-positive, Gram-negative and fungal organisms. PMID- 11274830 TI - Intellectual property rights and indigenous knowledge: credit where credit is due. PMID- 11274828 TI - Behavioral and pharmaco-toxicological study of Papaver rhoeas L. in mice. AB - A lyophilized ethanolic aqueous extract of Papaver rhoeas petals was evaluated for its behavioral and pharmaco-toxicological effects in mice and its chemical composition was studied using thin layer chromatography (TLC). In this study, chemical analysis by TLC showed that the petals contain some anthocyanins, whereas no alkaloids were detected. The toxicological effect of alcoholic and aqueous plant extract administered intraperitoneally was determined in mice. The toxicological results obtained indicated that 2000 mg/kg is LD10 and 4000 mg/kg is LD50. Behavioral and pharmacological studies of ethanolic and aqueous extract showed that the plant extract reduced locomotory, exploratory and postural behavior of mice. This was evaluated through two specific behavioral tests; a non familiar environment test (the Staircase test) and a familiar environment test (Free exploratory test). These behavioral and pharmacological effects are more pronounced when the solvent used for extraction is 10% ethanol and is not antagonized by benzodiazepines, opioids, dopaminergic and cholinergic compounds (flumazenil, naloxone, sulpuride and atropine). The plant extract did not induce sleep in mice after treatment with an infrahypnotic dose of pentobarbital. This finding shows that the plant extract has a sedative effect at a 400 mg/kg dosage. PMID- 11274831 TI - Odontogenic myxoma of maxillary sinus: CT and MR-pathologic correlation. AB - We showed the characteristic features of odontogenic myxoma in the maxillary sinus with computed tomography (CT), magnetic resonance imaging (MRI), and histopathological findings. CT images showed a multilocular soft tissue mass with bone destruction and thinning, and the characteristic finding of this lesion as strands of fine lacelike density. MRI revealed intermediate signal intensity on T1-weighted image and high signal intensity on T2-weighted image. MRI showed the erosive extent of the lesion into the adjacent structures. In contrast T1 weighted image, the peripheral portion of the lesion with a relatively large quantity of collagen bundles was enhanced, while the central portion with only mucoid component was not. The CT and MRI appearances correlated well with the histologic features and therefore were considered to be a useful tool for diagnosis of myxoma. PMID- 11274832 TI - Primary multifocal leptomeningeal gliomatosis. AB - A 23-year-old female university student was presented with recent onset of non specific headache and dizziness. She had no neurological deficit on neurological examination and magnetic resonance imaging of the brain revealed diffuse enhancement in the basal cisterns and cerebral sulci. She was treated as tuberculous meningitis but she did not improve and developed respiratory arrest. Autopsy showed primary multifocal leptomeningeal gliomatosis. PMID- 11274833 TI - Standardisation of imaging in neuroendocrine tumours: results of a European delphi process. AB - In 1998 and 1999, a delphi consensus procedure was performed to establish guidelines for standardised diagnostic imaging of neuroendocrine tumours. The procedure included four consecutive workshops of a European group of experts in neuroendocrine tumours as well as feedback given by specialists from the departments of radiology, nuclear medicine, surgery and internal medicine of the according home institutions. Diverging approaches among the centres, which became apparent during the discussion, reflect a lack of controlled studies specifically for rare subgroups of neuroendocrine tumours. This paper summarises the standards for diagnostic imaging as developed during the delphi process. In particular, the diagnostic workflows as well as the technical properties of different imaging modalities are described in detail. PMID- 11274834 TI - Comparison of 2-D turbo spin echo and 3-D gradient echo sequences for the detection of the trigeminal nerve and branches anatomy. AB - The aim of this study was to assess the detectability of the trigeminal nerve and its branches using T1 weighted (w.) 3-D magnetization prepared rapid gradient echo (MP-RAGE), T2* w. 3D CISS and T2 w. 2-D turbo spin echo MR sequences. Thirty healthy volunteers were examined for this purpose using a 1.5 Tesla MR unit. The detectability of the trigeminal nerve and Gasser's Ganglion, i.e. structures that are surrounded by liquor was best using 3-D CISS. In the case of the ophthalmic, maxillary and mandibular nerves, the T1 w. 3-D MPRAGE was significantly better than T2* w. CISS and T2 w. 2-D turbo spin echo. The latter yielded the poorest results. We conclude that both high resolution T2* w. and T1 w. 3-D sequences are necessary in order to detect the liquor-surrounded trigeminal nerve and its soft tissue-surrounded branches. We would therefore recommend the inclusion of constructive interference in steady state (CISS) and MP-RAGE in a MR imaging protocol of the trigeminal nerve and its branches. PMID- 11274835 TI - Atlantoaxial joints: patterns of gadolinium enhancement with MR imaging in normal subjects. AB - OBJECTIVE: To evaluate normal patterns of enhancement of signal intensity in and about the atlantoaxial joints following intravenous administration of a gadolinium-containing contrast agent. METHODS AND MATERIAL: Fat-suppressed axial T1 weighted SE images were obtained in 12 patients without evidence of inflammmatory arthritis before and immediately after intravenous administration of a gadolinium-containing contrast agent. Patterns of enhancement of signal intensity in and about the atlantoaxial joints were evaluated qualitatively. RESULTS: Four different MR imaging patterns were observed: no enhancement of signal intensity, enhancement in the form of a single punctate region, enhancement in a confluent pattern, and bandlike enhancement. CONCLUSION: Enhancement occurs around the odontoid process in patients without evidence of rheumatoid arthritis (RA). The dividing line between early synovitis and normal enhancement seems broader than expected. The reasons remain unclear and warant further studies. PMID- 11274836 TI - Morphology and patency of Gore-Tex wrapped internal mammary artery bypass evaluation with helical CT. AB - OBJECTIVE: To determine the patency of coronary internal mammary artery bypass (IMAB) with CT-angiography (CTA) and to evaluate the morphology of a covering Gore-tex IMAB-sleeve (PIMAS) used to protect the bypass at possible reoperation. MATERIALS AND METHODS: Sixty-five patients with IMAB wrapped with PIMAS (67 grafts) were prospectively investigated by CTA for bypass patency and sleeve morphology 6 months postoperatively with a standardised radiological and clinical protocol. RESULTS: All patent bypass arteries (62/62) were identified by CTA as open. In the remaining five cases, CTA revealed a bypass occlusion, which could be proven by coronary angiography in two cases (two patients refused angiography, one bypass was open angiographically). Morphology of the PIMAS could be imaged exactly in all cases. Sleeve implantation did not lead to adverse effects in terms of bypass occlusion or compression. In four patients, additional clinically relevant information were achieved. CONCLUSION: PIMAS implantation proved to be a safe procedure with good short-term results. CTA is a valuable method to exclude occlusion of sleeved IMA bypasses. Depiction of the wrapped IMAB by CTA supplies important information for preparing strategy in case of reoperation. PMID- 11274837 TI - MR imaging of primary leiomyosarcoma of the thyroid gland. AB - Primary leiomyosarcoma of the thyroid gland is extremely rare and radiological information on this tumor is scant. We presented radiological findings on primary thyroid leiomyosarcoma in a 66-year-old woman in which anaplastic carcinoma was suspected based on clinical and cytological features and discussed the radiological clues to distinguish between the two diseases. Ultrasonography showed an ill-defined hypoechoic mass without halo in the left lobe and the isthmus of the thyroid gland. Computed tomography depicted a low-density mass with calcification and necrosis, which invaded the thyroid cartilage. No lymphadenopathy was seen. The tumor was demonstrated as an isointense mass on T1 weighted MR images and a mass of intermediate signal on T2-weighted images. The tumor showed a fair enhancement on gadolinium-enhanced T1-weighted images. Although the radiological picture was nonspecific, primary thyroid leiomyosarcoma appeared less invasive and far less frequent in developing nodal metastasis than anaplastic carcinoma in light of the literature. PMID- 11274838 TI - Pulmonary mycosis in AIDS. AB - We retrospectively reviewed our series of 35 pulmonary mycosis in patients with AIDS, observed from 1987 to 1999, to correlate the imaging and pathologic findings. We further evaluated the frequency of fungal pneumonia before and after the use of a highly active antiretroviral therapy (HAART). Early recognition of pulmonary mycosis is imperative in these patients and improved survival can be achieved with early CT detection and prompt institution of high-dose antifungal therapy. PMID- 11274839 TI - General practitioners' willingness to request plain lumbar spine radiographic examinations. AB - OBJECTIVES: To examine general practitioners' attitudes to plain lumbar spine radiographic examinations. DESIGN: A postal questionnaire consisting of questions on background data and doctors' opinions about plain lumbar spine radiographic examinations, as well as eight vignettes (imaginary patient cases) presenting indications for lumbar radiography, and five vignettes focusing on the doctors' willingness to request lumbar radiography on the basis of patients' age and duration of symptoms. The data were analysed according to the doctor's age, sex, workplace and the medical school of graduation. SETTING: Finland. SUBJECTS: Six hundred and fifteen randomly selected physicians working in primary health care (64% of original target group). RESULTS: The vignettes revealed that the use of plain lumbar radiographic examination varied between 26 and 88%. Patient's age and radiation protection were the most prominent factors influencing doctors' decisions to request lumbar radiographies. Only slight differences were observed between the attitudes of male and female doctors, as well as between young and older doctors. Doctors' willingness to request lumbar radiographies increased with the patient's age in most vignettes. The duration of patients' symptoms had a dramatic effect on the doctor's decision: in all vignettes, doctors were more likely to request lumbar radiography when patient's symptoms had exceeded 4 weeks. CONCLUSIONS: General practitioners commonly use plain lumbar spine radiographic examinations, despite its limited value in the diagnosis of low back pain. Further consensus and medical education is needed to clarify the indications for plain lumbar radiographic examination. PMID- 11274840 TI - High resolution CT in children with cystic fibrosis: correlation with pulmonary functions and radiographic scores. AB - OBJECTIVE: To compare the high resolution CT (HRCT) scores of the Bhalla system with pulmonary function tests and radiographic and clinical points of the Shwachman-Kulczycki clinical scoring system. METHODS: HRCT of the chest was obtained in 40 children to assess the role of HRCT in evaluating bronchopulmonary pathology in children with cystic fibrosis (CF). The HRCT severity scores of the Bhalla system were compared with chest radiographic and clinical points of the Shwachman-Kulczycki scoring system and pulmonary function tests. Only 14 of the patients older than 6 years cooperated with spirometry. RESULTS: HRCT scores correlated well with radiographic points (r = 0.80, P<0.0001) and clinical points (r=0.67. P<0.0001) of the Shwachman-Kulczycki system, FVC (r = 0.71 P = 0.004) and FEV1 (r = 0.66, P = 0.01). Although radiographic points correlated significantly with FVC (r = 0.61, P = 0.02) and FEV1 (r = 0.56, P = 0.04), HRCT provides a more precise scoring than the chest X-ray. CONCLUSION: The HRCT scoring system may provide a sensitive method of monitoring pulmonary disease status and may replace the radiographic scoring in the Shwachman-Kulczycki system. It may be helpful especially in follow-up of small children too young to cooperate with spirometry. PMID- 11274841 TI - Quiz case. Symptomatic type II accessory navicular. AB - A 17 year old boy, who twisted his left foot while playing soccer a few days earlier, presented with pain and swelling along the medial aspect of the left foot. Clinical examination revealed an area of swelling and tenderness anteroinferiorly to the medial malleolus. Radiographs were considered normal and the patient was treated with a topical anti-inflammatory agent. During the following seven months the patient continued to experience pain on the medial side of the left foot, especially after a prolonged activity and when putting on shoes. Focal redness and tenderness was also evident. PMID- 11274842 TI - Structural basis for pulmonary functional imaging. AB - An understanding of fine normal lung morphology is important for effective pulmonary functional imaging. The lung specimens must be inflated. These include (a) unfixed, inflated lung specimen, (b) formaldehyde fixed lung specimen, (c) fixed, inflated dry lung specimen, and (d) histology specimen. Photography, magnified view, radiograph, computed tomography, and histology of these specimens are demonstrated. From a standpoint of diagnostic imaging, the main normal lung structures consist of airways (bronchi and bronchioles), alveoli, pulmonary vessels, secondary pulmonary lobules, and subpleural pulmonary lymphatic channels. This review summarizes fine radiologic normal lung morphology as an aid to effective pulmonary functional imaging. PMID- 11274843 TI - Quantification of pulmonary perfusion with MR imaging: recent advances. AB - Recent advances in magnetic resonance pulmonary perfusion imaging are reviewed, focusing on magnetic resonance perfusion imaging using gadolinium contrasts agents or spin labeling of blood using naturally flowing spins as the source of intravascular signal. These recent developments in magnetic resonance imaging have made it possible to analyze data quantitatively which holds significant potential for clinical imaging of lung perfusion and opens windows to functional MR imaging of the lung. We believe that fast magnetic resonance functional imaging will play an important role in the assessment of pulmonary function and the pulmonary disease process. PMID- 11274844 TI - Oxygen-enhanced magnetic resonance ventilation imaging of lung. AB - The oxygen-enhanced magnetic resonance (MR) ventilation imaging is a new technique, and the full extent of its physiological significance has not been elucidated. This review article includes background on (1) respiratory physiology; (2) mechanism and optimization of oxygen-enhanced MR imaging technique; (3) recent applications in animal and human models; and (4) merits and demerits of the technique in comparison with hyperpolarized noble gas MR ventilation imaging. Application of oxygen-enhanced MR ventilation imaging to patients with pulmonary diseases has been very limited. However, we believe that further basic studies, as well as clinical applications of this new technique will define the real significance of oxygen-enhanced MR ventilation imaging in the future of pulmonary functional imaging and its usefulness for diagnostic radiology. PMID- 11274845 TI - Pulmonary ventilation: dynamic MRI with inhalation of molecular oxygen. AB - We have recently demonstrated a non-invasive technique to visualize pulmonary ventilation in humans with inhalation of molecular oxygen as a paramagnetic contrast agent. In the current study, T1 shortening of lung tissue by inhalation of oxygen was observed (P<0.001). The T1 values of lung tissue were also correlated with arterial blood oxygen pressure (PaO(2)) in a pig, resulting in excellent correlation (r(2)=0.997). Dynamic wash-in and wash-out MR ventilation images as well as dynamic wash-in wash-out signal intensity versus time curves were obtained. The mean wash-in decay constants were 26.8+/-10.5 s in the right lung, and 26.3+/-9.5 s in the left lung. The mean wash-out decay constants were 23.3+/-11.3 s in the right lung, and 20.8+/-10.5 s in the left lung. Dynamic assessment of pulmonary ventilation is feasible using oxygen-enhanced MR imaging, which could provide dynamic MR ventilation-perfusion imaging in combination with recently developed MR perfusion imaging technique, and thus a robust tool for the study of pulmonary physiology and pathophysiology. PMID- 11274846 TI - Assessment of pulmonary perfusion using a subtracted HASTE image between diastole and systole. AB - The MR signal intensity change in the pulmonary parenchyma during the cardiac cycle was studied using HASTE sequence in volunteers. In addition, the potential to assess pulmonary perfusion abnormality by subtraction between diastolic and systolic HASTE images was tested in a pig model of pulmonary embolism. Signal intensity decreased in systole while it increased gradually in diastole. In a pig model with pulmonary embolism, subtracted images could identify the perfusion abnormality. Thus, subtraction of diastolic and systolic HASTE images has the potential to detect pulmonary perfusion abnormality. The technique may provide a new simple method for evaluating pulmonary perfusion. PMID- 11274847 TI - Computerized classification of interstitial lung abnormalities on chest radiographs with normalized radiographic index and normalized fractal dimension. AB - OBJECTIVE: To evaluate the performance of two kinds of physical measures, the normalized radiographic index (R) and the normalized fractal dimension (F), for computerized classification of interstitial lung abnormalities on chest radiographs. METHODS AND MATERIAL: The values of R were obtained as the normalized percent area of extracted opacities in selected regions of interest (ROIs). The values of F were calculated with a box-counting algorithm and then normalized. To extract linear opacities on chest radiographs selectively, we processed ROIs by four-directional Laplacian-Gaussian filtering and binarization (4LG/B), linear opacity judgment (LOJ), and linear opacity subtraction (LOS). We used the ROIs of 50 mild and 50 severe interstitial lung abnormalities. In both groups, all cases were divided into H (n=21, honeycombing opacities were found to be dominant) and Non-H (n=79, abnormal opacities were found, but these were excluded from H). We obtained three types of normalized physical measures of R and F in one ROI from 4LG/B, LOJ, and LOS images, and the combined indices, R(COM) and F(COM) were calculated. RESULTS: The values of F(LOJ) could differentiate H from Non-H in the mild-and the severe-abnormality group. However, all Rs could not differentiate H from Non-H in the severe-abnormality group. The combined indices of both R and F could differentiate H from Non-H in the mild abnormality group; however, these could not differentiate H from Non-H in the severe-abnormality group. CONCLUSION: The values of F(LOJ) seem to be useful in the classification of interstitial lung abnormalities on chest radiographs. PMID- 11274848 TI - Spiral CT findings in septic pulmonary emboli. AB - OBJECTIVES: The aim of the study was to determine the characteristics of septic pulmonary emboli and their prevalence on spiral computed tomographic (CT) scans. METHODS AND MATERIALS: We evaluated 65 lesions on spiral CT scans in ten patients with septic pulmonary emboli. Spiral CT scans (10-mm collimation) were obtained at 10-mm intervals from the lung apex to the diaphragm and were compared with posteroanterior chest radiographs obtained within 24 h after CT scanning. RESULTS: Only 21 (32%) of the 65 lesions detected on CT scans were also detected on chest radiographs. Peripheral nodules (39 lesions (60%)) were seen in all ten patients, wedge-shaped peripheral lesions (15 lesions (23%)) in nine patients, and infiltrates (11 lesions (17%)) in four patients. Subpleural lesions (45 lesions (69%)) and feeding vessels (35 (54%)) were found in all patients, and cavitary lesions (seven lesions (11%)) were seen in four patients. Subpleural peripheral nodules and wedge-shaped peripheral lesions were seen in nine patients. Thirty-two lesions (49%) ranged in diameter from 10 to 19 mm, and 59 lesions (91%) were less than 30 mm. CONCLUSIONS: Spiral CT is useful in detecting septic pulmonary emboli. On spiral CT subpleural peripheral nodules and wedge shaped peripheral lesions less than 30 mm in diameter are often found in patients with septic pulmonary emboli. PMID- 11274849 TI - Pulmonary actinomycosis with thoracic soft tissue mass: a rare onset form. AB - Actinomycosis is unusual, and rare especially when the lung and the thoracic wall are involved. It is more frequent in immunocompromised patient. US, CT, or MRI are imaging methods of diagnosis with high sensibility to recognise the disease and are able to the management. We point out a rare case in a normal teenager with thoracic abscess. PMID- 11274850 TI - Fine needle aspiration of solitary pulmonary lesions. AB - A 2-year experience using 25G sized needles for transthoracic fine needle aspiration (FNA) for solitary lung lesions under computed tomography (CT) guidance is documented with the main objective of reducing the postoperative complications through the use of a needle smaller than that previously routinely used. This study, in the hospital experience, demonstrated that FNAs became complication-free with the use of a smaller needle; whilst the sensitivity of the procedure was not much compromised. The duration of the hospitalization for the patients was reduced to under 24 h. The mandatory postoperative chest radiography can therefore be eliminated, provided no adverse clinical signs and symptoms are noted during the postoperative period. This is one of the first documented reports using 25G needles for FNAs. Further studies are needed on a wider scale to confirm the findings. PMID- 11274851 TI - Dose constraints to the individual annual doses of exposed workers in the medical sector. AB - The study is an attempt, within the process of the optimization of radiation protection, to propose constraints to the individual annual doses of classified workers employed in the medical sector of ionizing radiation applications in Greece. These exposed workers were grouped according to their specialties, i.e. medical doctors, technicians and nurses and their occupational category with common or similar tasks, such as diagnostic radiology, interventional radiology, nuclear medicine and radiotherapy. The last 5 years' annual dose distributions of these occupational groups, coming from the National Dose Registry Information System (NDRIS) of the Greek Atomic Energy Commission (GAEC) were analyzed. The proposed dose constraints (DCs) were set at levels, below which the annual doses of the 70 or 75% of the exposed workers per category are expected to be included. At the present stage the derived values may be considered achievable ceiling values referring to acceptably applied practices rather than to optimized ones, taking into account social and economic criteria. PMID- 11274852 TI - Indirect atomic absorption determination of atropine, diphenhydramine, tolazoline, and levamisole based on formation of ion-associates with potassium tetraiodometrcurate. AB - Ion-associate complexes of atropine sulphate (I), diphenhydramine HCl (II), tolazoline HCl (III) and levamisole HCl (IV) with potassium tetraiodomercurate were precipitated and their solubilities were studied as a function of pH, ionic strength and temperature. Saturated solutions of each ion-associate under the optimum precipitation conditions were prepared and the metal ion-content in the supernatant was determined. The solubility products were thus calculated at different temperatures. A new accurate and precise method using atomic absorption spectrometry for the determination of the investigated drugs in pure solutions and in pharmaceutical preparations is described. The drugs can be determined by the present method in the ranges 13.6--138.8, 5.6-58, 3.6--39.6 and 4.8--48 microg/ml solutions of I--IV, respectively. PMID- 11274853 TI - Development and validation of a stability indicating HPLC assay method for cyclosporine in cyclosporine oral solution USP. AB - The suitability of the United States Pharmacopeia (USP) assay method for analysing stressed samples of cyclosporine oral solution USP was evaluated for stability samples by analyzing cyclosporine oral solution after acid, alkali, hydrogen peroxide, heat and light treatment. Some of the degradants generated during stress testing, as well as dihydrocyclosporine A, which is a known degradant of cyclosporine A, were not adequately resolved from the cyclosporine peak and mobile phase adjustments did not improve the resolution. In addition, isocyclosporine A, another known degradant of cyclosporine, could not be quantitated as it was eluting too early with the system peaks. Therefore, a binary gradient, reverse phase, stability indicating, HPLC method for the assay of cyclosporine in cyclosporine oral solution USP has been developed and validated. Analysis of degraded samples showed that the cyclosporine A eluted as a spectrally pure peak resolved from its degradation products. PMID- 11274854 TI - Stability indicating HPTLC determination of Trimetazidine as bulk drug and in pharmaceutical formulations. AB - A simple, selective, precise and stability-indicating high-performance thin-layer chromatographic method of analysis of trimetazidine hydrochloride both as a bulk drug and in formulations is reported. The mobile phase composition was n-butanol water-methanol-ammonia (20%) (14:0.2:0.2:2, v/v/v/v). Densitometric analysis of trimetazidine hydrochloride was carried out in the absorbance mode at 254 nm. the calibration curve of trimetazidine hydrochloride in methanol was linear in the range 400 -- 2400 ng. The mean value of correlation coefficient, slope and intercept were 0.99815 and #61617;0.001, 0.4849 and #61617;0.001 and 31.633 and #61617;5.996 respectively. The limits of detection and quantitation were 50 and 80 ng respectively. The recovery of trimetazidine hydrochloride was about 98 -- 100%. This method was utilized to analyze trimetazidine hydrochloride from conventional tablets and controlled release pellets in the presence if commonly used excipients. PMID- 11274855 TI - LC separation of ortho and meta isomers of celecoxib in bulk and formulations using a chiral column. AB - A normal phase, isocratic LC method was developed for the separation of positional isomers of celecoxib (I) using a chiral column, Chiralpak-AD. The method is useful for the quantification of ortho (II) and meta (III) forms in bulk drugs and formulation samples of celecoxib. The method has been completely validated and proven to be rugged. The limit of detection (LOD) and limit of quantitation (LOQ) of ortho and meta forms were found to be 38 ng and 116 ng respectively. The active pharmaceutical ingredient was extracted from its finished dosage form (capsule) using ethanol. The percentage recoveries of ortho isomer was found to be 99.8--102.7 and 97.8--103.2 and the percentage recoveries of meta isomer was found to be 99.3--102.6 and 99.7--104 in spiked bulk and formulation samples of celecoxib respectively. PMID- 11274856 TI - Cathodic adsorptive stripping voltammetric determination of muscle relaxant: gallamine triethiodide (flaxedil). AB - A sensitive and simple voltammetric method of analysis is developed for the determination of trace amounts of gallamine triethiode in phosphate media. This method is based on controlled adsorptive preconcentration of the relaxant onto a Hanging Mercury Drop Electrode (HMDE) whereby mercurous iodide salt(s) are formed. The technique used is Cathodic Linear Sweep Stripping Voltammetry (CLSSV). The adsorptive response was evaluated with respect to preconcentration time and potential. As little as 3 x 10(-9) mol dm(-3) i.e. 2.7 ppb flaxedil (proconcentration time 300 seconds) can be determined successfully. The application of this method was tested in the determination of flaxedil in pharmaceutical preparation (ampoules). PMID- 11274857 TI - Determination of captopril in pharmaceutical tablets by anion-exchange HPLC using indirect photometric detection; a study in systematic method development. AB - The development and validation of a significantly cost effective and simpler anion-exchange high performance liquid chromatorgaphic (HPLC) procedure than the compendial methods for the analysis of captopril in tablet dosage forms using indirect photometric detection is described. A low capacity anion-exchange column was used with potassium phthalate as the mobile phase marker and indirect detection at 280 nm. The chromatographic conditions were optimized using the Box and Behnken factorial experimental design. The method was precise and accurate with percent recovery (+/-%R.S.D.) of 99.8+/-0.7% (n=12) with the spiked concentrations ranging between +/-30% of the assay level. Youden and Steiner's robustness test, involving seven chosen variables, showed that the method is robust. Using the developed method, commercially available captopril tablets were assayed and the results were found to be comparable with those obtained by the compendial method. PMID- 11274858 TI - Determination of amphotericin B in human plasma using solid-phase extraction and high-performance liquid chromatography. AB - A rapid and selective HPLC method is described and validated for measuring amphotericin B (AB) in plasma. The procedure involves the solid phase extraction of AB from plasma by incorporating 1-amino-4-nitronaphthalene as an internal standard during the last elution step in extraction followed by HPLC analysis with UV detection at 407 nm. The chromatographic separation is achieved in less than 10 min on a reversed-phase C-18 column using acetonitrile-disodium edetate (20 mM) (45:55, v/v) at pH 5.0 as eluent. A linear response over the concentration range of 0.0100--2.00 microg ml(-1) is obtained having a detection limit of 0.00500 microg ml(-1) for AB. The mean extraction recovery is found to be 98.1+/-1.1% (n=15). The within-day and day-to-day R.S.D. were less than 2% (n=15) and 6.54% (n=45) respectively. This method is applied for quantifying AB trough levels in the plasma of cancer patients who have been on antifungal therapy with AmBisome. It can further be applied either for AB therapeutic monitoring or single/multiple pharmacokinetic analysis of AB in plasma. PMID- 11274859 TI - Determination of UDP-glucuronosyltransferase UGT1A6 activity in human and rat liver microsomes by HPLC with UV detection. AB - A simple and sensitive method for the determination of UDP glucuronosyltransferase UGT1A6 activity using 4-methylumbelliferone (4-MU) and 4 nitrophenol (4-NP) as substrates in human and rat liver microsomes by high performance liquid chromatography (HPLC) with uv detection is reported. The method was validated for the determination of 4-methylumbelliferyl beta-D glucuronide (4-MUG) and 4-nitrophenyl beta-D-glucuronide (4-NPG) with respect to specificity, linearity, detection limit, recovery, stability, precision and accuracy. There was no interference from matrix and non-enzymatic reactions. Calibration curves for 4-MUG and 4-NPG are linear from 0.5 to 500 microM. Average recoveries ranged from 98 to 100% in spiked liver microsomes samples. 4-MUG and 4 NPG were stable at 4 degrees C for at least 72 h in spiked liver microsomes samples. The method was found to be more sensitive than previous methods using a spectrophotometer, a spectrofluorometer and HPLC. The detection limit for 4-MUG and 4-NPG (signal-to-noise ratio of 3) was 14 and 23 nM, respectively. The intra- and inter-day precision (relative S.D. (RSD)) and accuracy (relative mean error (RME)) was <5 and 9%, respectively. The intra- and inter-day reproducibility (RSD) of UGT1A6 enzyme assay in liver microsomes was <6%. With this improved sensitivity, the kinetics of UGT activities toward 4-MU and 4-NP in human and rat liver microsomes could be determined more precisely. In addition, the method could determine the non-inducible, and 3-methylcholanthrene- and phenobarbital inducible activities of UGT1A6 in rat liver microsomes under the same assay conditions. Therefore, this method is applicable to in vivo and in vitro studies on the interaction of xenobiotic chemicals with UGT1A6 isoform in mammals using small amounts of biological samples. PMID- 11274860 TI - ION-pair liquid chromatography technique for the estimation of metformin in its multicomponent dosage forms. AB - A simple, precise and accurate high performance liquid chromatography (HPLC) method was developed for the simultaneous estimation of metformin with gliclazide and glipizide present in multicomponent dosage forms. The method was carried out on Inertsil C(18) column. A mobile phase composed of acetonitrile-water containing camphor sulphonic acid (adjusted to pH 7 using 0.1 N sodium hydroxide; 75 mM) at a flow rate of 1 ml min(-1) was used for the separation. Detection was carried out at 225 nm. Tolbutamide was used as internal standard. Validation of the developed HPLC method was carried out. PMID- 11274861 TI - The rapid quantitative analysis of phenprobamate and acetaminophen by RP-LC and compensation technique. AB - For the analysis of phenprobamate and acetaminophen in combination, the main analytical methods used were spectrophotometric compensation technique and Vierordt's method with high performance liquid chromatography, used as an analytical reference method. The first procedure for the simultaneous quantitative determination phenprobamate and acetaminophen by high performance liquid chromatographic (HPLC) method was proposed. The method was standardized using a LiChrosorb RP18-5 column, methanol-water-formic acid (120:80:1 v/v), apparent pH 4.25 with triethylamine, as mobil phase and UV detection at 254 nm. The peak area response versus concentration was linear in a concentration range from 4 to 28 microg ml(-1) of phenprobamate and from 4 to 30 microg ml(-1) for acetaminophen. The correlation coefficients were 0.9999 for phenprobamate and 0.9987 for acetaminophen. The second procedure, based on the compensation technique, is presented for the derivative spectrophotometric determination of binary mixtures with overlapping spectra. The proposed methods, which give thoroughly comparable data, are simple and rapid and allow precise and accurate results. PMID- 11274863 TI - Determination of iron and molybdenum in a dietetic preparation by flame AAS after dry ashing. AB - Methods for the determination of iron and molybdenum in a dietetic pharmaceutical preparation by flame atomic absorption spectrometry (FAAS) after dry ashing at 600 degrees C have been validated. Linearity, precision, accuracy, detection and quantification limits, specificity and robustness have been determined. Linearity of response was verified for concentrations ranging from 0.50 to 4.00 mg l(-1) of iron and 1.00 to 6.00 mg l(-1) of molybdenum. Precision of the methods, performed under conditions of repeatability and reproducibility, gave relative standard deviations of 0.4 and 1.1%, respectively, for the iron determination and of 1.0 and 6.5%, respectively, for the molybdenum determination. Mean recoveries determined after spiking dietetic preparation placebos ranged from 97.1 to 102.6% for iron and 95.2 to 102.9% for molybdenum. The limit of detection for iron was 126 microg g(-1) and for molybdenum 129 microg l(-1). Quantification limits were 420 and 433microg l(-1) for iron and molybdenum, respectively. No interference in the iron and molybdenum determination due to other components present in the dietetic capsules was found. Day-to-day and analyst-to-analyst variability was less than 1.1% for iron and 4.5% for molybdenum. Results show the suitability of the method for measurement of iron and molybdenum in a complex matrix sample such as a dietetic pharmaceutical preparation. PMID- 11274862 TI - LC method for the analysis of acetylsalicylic acid, caffeine and codeine phosphate in pharmaceutical preparations. AB - An accurate, simple, reproducible and sensitive method for the determination of acetylsalicylic acid, caffeine and codeine phosphate has been developed and validated. Acetylsalicylic acid, caffeine and codeine phosphate were separated using a microBondapack C(8) column by isocratic elution with flow rate 1.0 ml/min. The mobile phase composition was 125/125/250/0.5 (v/v) isopropyl alcohol, acetonitrile, water and o-phosphoric acid. The samples were detected at 215 nm using photo-diode array detector. The linear range of detection for acetylsalicylic acid, caffeine and codeine phosphate were between 0.40 and 1000, 0.25 and 250, and 0.48 and 96 microg/ml, respectively. The linearity, range, selectivity, system performance parameters, precision, accuracy, and ruggedness for acetylsalicylic acid, caffeine and codeine phosphate were also shown to have acceptable values. PMID- 11274865 TI - Spectrophotometric resolution of metronidazole and miconazole nitrate in ovules using ratio spectra derivative spectrophotometry and RP-LC. AB - Metronidazole and miconazole nitrate in ovules was determined by ratio spectra derivative spectrophotometry and by high-performance liquid chromatography (HPLC). The first method depends on ratio spectra first derivative spectrophotometry, by utilizing the linear relationship between substances concentration and ratio spectra first derivative peak amplitude. The ratio first derivative amplitudes at 242.6 [(1)DD(242.6)], 274.2 [(1)DD(274.2))] 261.8 [(1)DD(261.8))] 273.5 [(1)DD(273.5))]and 281.5 [(1)DD(281.5)] nm were selected for the assay of metronidazole and miconazole nitrate, respectively. The second method is based on high-performance liquid chromatography on a reversed-phase column using a mobile phase of methanol-water-phosphoric acid (30:70:0.20 v/v) (pH 2.8) with programmable detection at 220.0 nm. The minimum concentration detectable by HPLC was 0.9 microg ml(-1) for metronidazole and 0.3 microg ml(-1) for miconazole nitrate and by ratio derivative spectrophotometry 4.0 microg ml( 1) for metronidazole and 0.5 microg ml(-1) for miconazole nitrate. The proposed procedures were successfully applied to the simultaneous determination of metronidazole and miconazole nitrate in ovules with a high percentage of recovery, good accuracy and precision. PMID- 11274864 TI - Determination of mangafodipir trisodium and related impurities in bulk substance and pharmaceutical formulation by ion-pair high-performance liquid chromatography. AB - The development of an ion-pair liquid chromatographic method for determination of mangafodipir trisodium and related impurities is described. Good resolution was obtained when using a polymeric reverse-phase column and a mobile phase of pH* 10.5 composed by borate buffer, acetonitrile, and tetrabutylammonium hydrogensulphate as ion pair agent. Validation of the method showed good selectivity, precision, accuracy and linearity, and detection limits of 0.1--0.2 microg/ml. PMID- 11274866 TI - Immunoassays for the detection of nicergoline and its metabolites in human plasma. AB - In order to determine nicergoline pharmacokinetics after oral administration to humans, we have developed two radioimmunoassays, one directed against nicergoline and the other directed against known nicergoline metabolites. The assays were validated according to the recommendations of international regulatory agencies and their limits of quantification were 40 and 10 pg/ml, respectively. In order to further validate the methods, a chromatographic separation of immunoreactive entities was performed with samples from healthy volunteers who were given 15 mg of Sermion (nicergoline orally administered). Chromatographic determination of assay specificity showed that the metabolite radioimmunoassay recognised known nicergoline metabolites but also a new metabolite. Using the antibodies directed against nicergoline, we were unable to detect nicergoline in the human plasma. This suggests that nicergoline is absent in the circulation because of complete metabolism through its first-pass effect. PMID- 11274867 TI - Screening of an enterovirus specific RT-PCR ELISA method for the quantification of enterovirus genomes in human body fluids by means of a three-level experimental design. AB - In order to obtain a detection limit as low as possible for a quantitative enterovirus specific RT-PCR ELISA assay, optimal reaction conditions, which give rise to the highest response, need to be determined. This was done by investigating the influence of 13 factors, selected from RT and PCR, in a multivariate approach by means of a well-balanced three-level screening design, derived from a three-level Plackett--Burman design. Optimal reaction conditions could be determined by calculation and evaluation of the effects of the different factors on the response, i.e. the measured absorbance of the ELISA detection. The method will be used to study a possible longitudinal relationship between enteroviruses and the development of multiple sclerosis and juvenile diabetes. PMID- 11274868 TI - Stability indicating method for determination of nortriptyline hydrochloride using 3-methyl-2-benzothiazolinone hydrazone (MBTH). AB - A spectrophotometric procedure is described for determination of nortriptyline hydrochloride in pure and dosage form as well as in the presence of its degradate. 3-Methyl-2-benzothiazolinone hydrazone (MBTH) has been used as the chromogenic reagent, where aqueous solutions of the drug and reagent are treated with cerium(IV) ammonium sulphate in an acidic medium. Nortriptyline hydrochloride reacts to give a blue coloured product having two absorption maxima at 619 and 655 nm. Various parameters affecting the reaction have been studied. Beer's law is obeyed in the concentration range of 24-216 microg ml(-1) of nortriptyline hydrochloride, with mean percentage recoveries of 100.22+/-0.870 and 100.66+/-0.642% for both maxima, 619 and 655 nm, respectively. Results were statistically analyzed and compared with those obtained by applying the British Pharmacopoeia (1993) method. PMID- 11274869 TI - Analysis of diflunisal by electrochemical methods. AB - A new differential pulse polarographic (DPP) and differential pulse adsorptive stripping voltammetric (DPAdSV) methods for the electrochemical behavior and quantitative determination of diflunisal were described. In these voltammetric methods, the peak potential of diflunisal was found as -0.31 V (vs. Ag/AgCI) with selected Britton--Robinson buffer (BR, pH 7.8) as a supporting electrolyte. The variation of the peak current with the concentration of diflunisal were linear in the 9.0--40.0 and 4.0--30.0 microg ml(-1) concentration ranges for DPP and DPAdSV methods, respectively. The limits of detection (LOD) were found as 5.0 and 0.1 microg ml(-1) for DPP and DPAdSV methods, respectively. The developed methods were validated by evaluation of the validation parameters. The characteristics of the peak current of diflunisal were examined in detail and the results proved that the peak current has an adsorption characteristic. The developed methods were proposed for rapid determination of diflunisal in commercial tablets. The recovery studies showed that developed assays had a good accuracy and precision with mean recoveries 99.92 and 100.02% and mean variation coefficients 0.29 and 0.24% in DPP and DPAdSV methods, respectively. PMID- 11274870 TI - Direct determination of s-carboxymethyl-l-cysteine in syrups by reversed-phase high-performance liquid chromatography. AB - A simple reversed-phase high-performance liquid chromatography method was developed for the determination of s-carboxymethyl-l-cysteine in syrup preparations. The experiments were performed without specific sample pre treatment. The LC conditions used were acetonitrile-10 mM sodium dihydrogenphosphate buffer, pH 2.0 (1:99, v/v) on a C(18) Inersil column with a flow rate of 1.5 ml/min. Ultraviolet detection was carried out at 240 nm. The method showed excellent linearity (r(2)>0.9998) over the concentration range tested (0.8-25.6 mg/ml) with good precision and accuracy (%R.S.D. 0.7%). Recoveries were good (>99%) with a limit of detection and limit of quantitation of 0.1 and 0.8 mg/ml. Other compositions in the syrup vehicle did not interfere the analysis of s-carboxymethyl-l-cysteine. PMID- 11274871 TI - A literature review of the consequences of prenatal marihuana exposure. An emerging theme of a deficiency in aspects of executive function. AB - In spite of marihuana being the most widely used illegal drug among women of reproductive age, there is a relative paucity of literature dealing with the neurobehavioral consequences in offspring--particularly the longer-term effects. However, there is a degree of consistency in the limited data, both across cross sectional reports and longitudinally, where offspring have been followed for a number of years. Two cohort studies fall into the latter category; one involving a low-risk sample and, the other, a high-risk sample. Global IQ is not impacted by prenatal marihuana exposure but aspects of executive function (EF)--in particular, attentional behavior and visual analysis/hypothesis testing--appear to be negatively associated with in utero cannabis exposure in children beyond the toddler stage. This hypothesized influence of prenatal marihuana on EF is examined and discussed relative to effects (or lack of effects) across different ages in the offspring, cannabinoid receptors, and the extant general marihuana and prefrontal literature. PMID- 11274872 TI - Contribution of maternal smoking during pregnancy and lead exposure to early child behavior problems. AB - Maternal smoking during pregnancy elevates risk for later child behavior problems. Because prior studies considered only Western settings, where smoking co-occurs with social disadvantage, we examined this association in Yugoslavia, a different cultural setting. Mothers enrolled in pregnancy as the low-exposure group in a prospective study of lead exposure were interviewed about health, including smoking history. A total of 199 children were assessed on the Child Behavior Checklist (CBCL) at ages 4, 4 1/2, and 5 years. Average cumulative blood lead (BPb) was determined from serial samples taken biannually since delivery. Longitudinal analyses were derived from 191 children with available data on behavior and covariates. Smoking was unrelated to social adversity. Controlling for age, gender, birthweight, ethnicity, maternal education, and Home Observation for Measurement of the Environment (HOME) Acceptance, smoking was associated with worse scores on almost all subscales; BPb concentration was related to small increases in the Delinquency subscale. Daughters of smokers received significantly higher scores on Somatic Complaints compared to daughters of nonsmokers, consistent with other work relating biological factors and internalizing problems in young girls. Because the present smoking/child behavior associations persist after control for individual and social factors also related to behavior problems, possible biological mediators are considered. PMID- 11274873 TI - Elevations in plasmatic titers of corticosterone and aldosterone, in the absence of changes in ACTH, testosterone, or glial fibrillary acidic protein, 72 h following D,L-fenfluramine or D-fenfluramine administration to rats. AB - Studies in both humans and animals demonstrate that D,L- and D-fenfluramine (D,L FEN and D-FEN, respectively) can activate the hypothalamic-pituitary-adrenal axis following an acute dose. No data exist showing a prolonged effect of either drug, although two studies have hinted at increased adrenal activity. There are also considerable differences in the literature pertaining to the neurotoxic effects of D,L- and D-FEN. Some possible explanations for these differences include: activation of different neurotransmitter systems, the temperature at which the animals were maintained during exposure, or the substance sampled in each study. We investigated the effects of either D,L-FEN or D-FEN on pituitary, adrenal, and gonadal hormones 72 h after drug exposure. Furthermore, using a dosing regimen adapted from studies on methamphetamine (e.g., four times every 2 h in a single day) known to produce elevations in glial fibrillary acidic protein (GFAP) under hyperthermic conditions, we examined the effects of D- and D,L-FEN (15 mg/kg, four times) on GFAP content when the animals were dosed at ambient temperatures of 21 or 32 degrees C. Approximately fivefold increases of corticosterone and threefold increases of aldosterone were found 72 h later under resting conditions following both D- and D,L-FEN. Nonetheless, when animals were dosed with D-FEN at 32 degrees C, no significant elevation in corticosterone was detected. No effect was observed for ACTH, testosterone, or GFAP following D- or D,L-FEN treatment. These data suggest that: (1) FEN treatment causes prolonged elevations in adrenal cortical hormones; (2) FEN-treated animals displayed hormonal characteristics similar to animals undergoing a chronic stressor as suggested by no difference in ACTH titers; (3) D,L-FEN treatment or D-FEN treatment (as reported previously) is not similar to other substituted amphetamines in that it does not increase GFAP, even under hyperthermic conditions. PMID- 11274874 TI - Comparing cognitive and screening tests for neurotoxicity. Effects of acute chlorpyrifos on visual signal detection and a neurobehavioral test battery in rats. AB - It is often assumed that cognitive function is more sensitive to neurotoxic chemicals than are the unconditioned behaviors employed in neurobehavioral screens; however, direct comparisons of the sensitivity of these test methods are lacking. The present studies were conducted to compare the effects of the widely used cholinesterase-inhibiting insecticide, chlorpyrifos (O,O'-diethyl O-3,5,6 trichloro-2-pyridyl phosphorothionate, CPF), on a visual signal detection task (SDT) with its effects on a neurobehavioral test battery. Adult male Long-Evans rats were trained to perform the SDT, dosed with CPF, and then assessed with both test instruments. Oral CPF (50 mg/kg) impaired signal detection for 8 days, and subcutaneous CPF (250 mg/kg) did so for 4 weeks. CPF (30 and 50 mg/kg po and 250 mg/kg sc) also lowered activity in the test battery for up to 18 days. Thus, CPF impaired attention and altered behavior in the test battery in the same dose ranges under two very different dosing scenarios. PMID- 11274875 TI - Developmental exposure to methylmercury alters behavioral sensitivity to D amphetamine and pentobarbital in adult rats. AB - Female rats were exposed to 0, 0.5, or 6.4 ppm methylmercury in their drinking water before mating, and throughout gestation and lactation. When the female offspring were 4-6 months old, they were trained to respond under a multiple differential reinforcement of high rate (DRH) 9:4-- Extinction schedule of reinforcement. No differences among exposure groups were apparent in steady-state behavior. Drug challenges were conducted with multiple doses of D-amphetamine, scopolamine, pentobarbital, haloperidol, and dizocilpine, drugs selected for their different pharmacological effects. The ED(50) values for amphetamine's reinforcement rate-reducing effects for the control, 0.5-, and 6.4-ppm groups were 3.1, 1.9, and 0.9 mg amphetamine/kg body weight, respectively, demonstrating an increased sensitivity to D-amphetamine in methylmercury-exposed rats. Rats in the 6.4-ppm group also demonstrated a relative insensitivity to pentobarbital. Further, these exposed rats exhibited an inverted U-shaped dose-effect curve under the pentobarbital dose-effect determination, while controls showed only a declining curve. Exposed rats did not respond differentially to haloperidol, scopolamine, or dizocilpine, suggesting specificity. The present data suggest an involvement of catecholaminergic and GABAergic activity in methylmercury's neurotoxicity. PMID- 11274876 TI - Neonatal dexamethasone on day 7 in rats causes behavioral alterations reflective of hippocampal, but not cerebellar, deficits. AB - Developmental glucocorticoid treatment in rats has been shown to cause body and brain weight decrements concurrent with behavioral alterations. Here, Sprague Dawley rats were treated with the synthetic glucocorticoid, dexamethasone (DEX), on postnatal day (PND) 7 (1.5 mg/kg, sc, injected in the morning and afternoon). Behavioral assessments of negative geotaxis, locomotor activity (open field, maze exploration, residential running wheel, residential figure 8 maze), open-field activity response to amphetamine, acoustic startle, prepulse inhibition (PPI) of acoustic startle, juvenile play behavior, anxiety (emergence tests), motor coordination (rotarod performance), spatial learning (Morris water maze and food reinforced complex maze), and operant performance (time estimation and response inhibition) were assessed in male rats. Body weight was decreased beginning at PND 43 until sacrifice on PND 127. Whole and regional brain weights were less, especially hippocampus, cerebellum, brainstem, and cortical remnant. Indications of delayed development were apparent; specifically, DEX-treated rats took significantly longer to turn on PND 8, but not PND 9, in the negative geotaxis test. DEX treatment induced deficits in the Morris water maze that were similar to hippocampal deficits. Open-field activity changes were inconsistent; however, DEX-treated rats were hyperactive during the dark period in running wheel tests. There were no indications of changes in reactivity or emotionality. PMID- 11274877 TI - Cochlear pathology induced by styrene. AB - Hair cells, spiral fibers and spiral ganglion cells (SGCs) coming from cochleae of styrene-treated Long-Evans rats were counted in order to assess the extent and location of the cochlear injury after the solvent inhalation. If the hair cells, and more specifically the outer hair cells (OHCs), were undoubtedly the first targets of inhaled styrene, the histological results of the present study would seem to indicate that neurons of the spiral ganglion were also injured with increasing styrene doses. The degenerative process of SGCs and spiral fibers within the osseous lamina was predominant in the middle and mid-basal turn. The electrophysiological data, obtained by recording near-field potentials from the inferior colliculus, reflected the damages of the SGCs and fibers but were not consistent with the histopathological data of the organ of Corti. Because of the weak correlation between the styrene-induced injury at the level of the organ of Corti and that induced at the level of the spiral ganglion, it is likely that two different intoxication routes exist within the cochlea. Such an assumption is discussed in the present paper. PMID- 11274878 TI - Plasma and brain methamphetamine concentrations in neonatal rats. AB - D-Methamphetamine (D-MA) treatment during the neonatal period has been shown to induce acoustic startle hyperreactivity and Morris maze spatial learning deficits, but not to significantly affect Cincinnati maze sequential learning. In order to characterize the internal dose in these experiments, MA was measured in plasma and brain of neonatal rats at one of two ages, and using one of three dose schedules, two of which were selected to be representative of those used in previously published neurobehavioral studies. Plasma parameters showed few age and dose-frequency effects; however, brain concentrations showed more consistent age-dependent effects. Brain area under the concentration (AUC) values were consistently higher, regardless of dosing schedule, in offspring treated on postnatal day (P) 1 compared to those treated on P11. Previous results with the multiple-dose schedules have shown that Morris maze spatial learning deficits only occur in those exposed beginning on P11, whereas acoustic startle hyperreactivity is associated with exposure beginning on either P1 or P11. The pharmacokinetic parameters did not predict the long-term spatial learning and memory effects of neonatal MA administration, nor are they well correlated to the acoustic startle effects. The plasma concentrations obtained in rats are within the range for human MA abusers based on extrapolations from human low-dose values to those expected for heavy users. PMID- 11274879 TI - Diffusion-weighted echo-planar MR imaging: clinical applications and pitfalls -- a pictorial essay. AB - Diffusion-weighted imaging (DWI) provides unique information about various pathological changes of the brain. DWI is sensitive for the detection of hyperacute infarcts, and useful in distinguishing acute or subacute infarcts from chronic infarcts. DWI is useful in differentiating cytotoxic edema from vasogenic or interstitial edema, which may help to determine prognosis. DWI is useful in differentiating cystic or necrotic tumors from abscesses or epidermoids. DWI can discriminate nonenhanced tumor infiltration from vasogenic edema, and differentiate dysmyelination from demyelination. PMID- 11274880 TI - Proptosis with orbital soft tissue and bone changes and unilateral papilloedema: unusual presentation of POEMS syndrome. AB - POEMS syndrome is a rare manifestation comprising of polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin changes. Orbital involvement in this syndrome is rare and manifestation with proptosis, even rarer. The imaging features in Poems syndrome and rarity of various features in our case are being highlighted here. PMID- 11274882 TI - Gastric carcinoid tumors as a consequence of chronic hypergastrinemia: spiral CT findings. AB - The development of gastric carcinoid tumors is a rare but recognized complication of prolonged, severe hypergastrinemia. We present 2 patients with elevated gastrin levels who developed gastric carcinoid tumors and the CT findings are reviewed. PMID- 11274881 TI - Lemierre's syndrome presenting as multiple lung abscesses. AB - Lemierre's syndrome is thrombophlebitis of the internal jugular vein (IJV), complicating an oropharingeal infection. The causative organism is Fusobacterium, an anaerobic bacillus, and the syndrome typically occurs in previously healthy teenagers and young adults. Thromboembolic metastases are a common sequela, and the lungs are most frequently affected. We present a case of a 25-year-old woman, who presented with multiple lung abscesses, in whom IJV thrombophlebitis was subsequently noted. PMID- 11274883 TI - Perforation of the small bowel as a complication of laparoscopic cholecystectomy: CT findings. AB - Despite the widespread use of laparoscopic cholecystectomy, technical complications unique to the laparoscopic approach may lead to significant postoperative morbidity and mortality. We report a rare case of small bowel perforation due to trocar injury that led to extensive pneumoperitoneum and pneumomediastinum in a patient who underwent laparoscopic cholecystectomy. Small bowel injuries should be suspected when a large or an increasing amount of free air is detected following this procedure. PMID- 11274884 TI - Primary epiploic appendagitis: a report of two cases. AB - Primary epiploic appendagitis (PEA) is a rare benign self-limiting inflammatory process of the colonic epiploic appendices. Patients present with acute abdominal pain, often misdiagnosed clinically as acute appendicitis or diverticulitis. Computed tomography (CT) scan findings of this condition are characteristic and can confidently suggest the diagnosis avoiding unnecessary barium enemas and colonoscopy, biopsy, or surgery. PMID- 11274885 TI - Evaluation of therapeutic effectiveness of transarterial chemoembolization for hepatocellular carcinoma: correlation of dynamic susceptibility contrast-enhanced echoplanar imaging and hepatic angiography. AB - The objective of this study was to evaluate the therapeutic effectiveness of transarterial chemoembolization (TACE) for hepatocellular carcinoma (HCC) with dynamic susceptibility contrast-enhanced magnetic resonance imaging (DSC-MRI). Seventeen patients with histopathologically proven HCC were included in this study. All patients underwent MR examinations with conventional T1- and T2 weighted images, gadolinium-enhanced images, and DSC-MRI before TACE treatment. Hepatic blood volume (HBV) maps were reconstructed from the time-intensity curves. The same MRI sequences and techniques were repeated 24 h and 6 weeks after TACE. Serial changes in tumor perfusion on HBV maps were correlated with vascularity in hepatic angiography. All tumors were hypointense on T1-weighted images and hyperintense on T2-weighted images. Heterogeneous enhancement was observed in all tumors before and immediately after TACE. Hyperperfusion was noted in most of the tumors on HBV map before TACE and moderate to marked hypoperfusion following TACE. The degree of tumor perfusion on HBV map correlated well with the vascularity in angiography. In conclusion, the noninvasive nature of DSC-MRI is useful to evaluate the effectiveness of TACE. Invasive procedures, such as angiography, are seldom necessary. PMID- 11274887 TI - Multiple renal artery aneurysms diagnosed by three-dimensional CT angiography. AB - Renal artery aneurysm is a relatively rare disease that is found in 0.3--1.0% of patients examined by renal angiography. We report a case of multiple renal artery aneurysms that were accurately diagnosed by three-dimensional CT angiography (3D CTA) using volumetric data sets of spiral CT. PMID- 11274888 TI - Extraperitoneal pelvic leiomyosarcoma. MR findings in a case. AB - The magnetic resonance imaging (MRI) of a patient with nongynecologic pelvic leiomyosarcoma is presented. A retroperitoneal mass appeared under the broad ligaments, in the right paravesical and parametrial, lateral pararectal site. The mass leaned on the uterus and vagina. On MRI, the mass had solid structure, isointense on T1-weighted images, inhomogeneously iperintense on T2-weighted images with central areas of increased intensity. Late after contrast agent administration, the mass appeared inhomogeneously ipointense with areas of fair late contrast enhancement. The morphology of the tumor, the retroperitoneal site, and MRI sequences make the differential diagnosis. These evidences were useful in treatment planning. PMID- 11274886 TI - Intrarenal pseudoaneurysms complicating renal choriocarcinoma metastases: treatment with coil embolization. AB - Most intrarenal pseudoaneurysms result from a laceration of the renal artery or its branches. However, tumor-induced renal pseudoaneurysm is very rare. We report a case in which embolization of an intrarenal pseudoaneurysm complicating renal metastases resulting from a choriocarcinoma was successful. PMID- 11274889 TI - Radiologically identified molar invasion into pelvic arteriovenous shunts. AB - A case of radiologically identified molar invasion into extensive arteriovenous shunts (AVSs) is described. CT and MRI revealed a large uterine mass, accompanied by multiple AVSs. Dynamic MRI and pelvic angiography demonstrated multiple trophoblastic cysts invading into the AVSs. Resected specimen confirmed the diagnosis of invasive mole. Dynamic MRI was very useful in determining the etiology of AVS. PMID- 11274890 TI - A revisit of MRI analysis for synovial sarcoma. AB - We evaluated magnetic resonance imaging (MRI) findings of synovial sarcomas in 22 patients, and the most common MRI findings were oval and well-defined nodular masses with heterogeneous intermediate signal intensity (SI) on T1 weighted images (WI), high SI on T2-WI and heterogeneous contrast enhancement. A cystic component was seen in 77%, intratumoral hemorrhage in 73%, and calcification in three monophasic sarcomas. Metastases were noted in lung (mostly biphasic type), lymph node, and bone. Posttreatment changes revealed diffusely increased S1 on T2 W1 and slightly diffuse contrast enhancement with feathery appearance. Morphology and MR signal characteristics assist in synovial sarcoma management. PMID- 11274891 TI - Magnetic resonance imaging of septic arthritis. AB - Septic arthritis is a disabling and life-threatening disease that requires early diagnosis for optimal outcome. Although traditionally a clinical and laboratory diagnosis, some patients may be misdiagnosed and referred for magnetic resonance (MR) imaging. Therefore, radiologists need to be aware of the MR imaging findings of septic arthritis, its complications, and diagnostic pitfalls. PMID- 11274892 TI - MR evaluation of talonavicular angle in congenital talipes equinovarus. AB - The talonavicular relationship in 14 patients with talipes equinovarus was quantified using gradient echo sequences. The angle formed between the short axis of the navicular and the long axis of the talus was measured. There is a significant difference in the talonavicular angle between patients who had posteromedial release (PMR) and those who had posterior release (PR) or casting only (P=.0004). This method provides an objective assessment of residual deformity following surgical and conservative management of talipes equinovarus. PMID- 11274893 TI - Drug-like properties and the causes of poor solubility and poor permeability. AB - There are currently about 10000 drug-like compounds. These are sparsely, rather than uniformly, distributed through chemistry space. True diversity does not exist in experimental combinatorial chemistry screening libraries. Absorption, distribution, metabolism, and excretion (ADME) and chemical reactivity-related toxicity is low, while biological receptor activity is higher dimensional in chemistry space, and this is partly explainable by evolutionary pressures on ADME to deal with endobiotics and exobiotics. ADME is hard to predict for large data sets because current ADME experimental screens are multi-mechanisms, and predictions get worse as more data accumulates. Currently, screening for biological receptor activity precedes or is concurrent with screening for properties related to "drugability." In the future, "drugability" screening may precede biological receptor activity screening. The level of permeability or solubility needed for oral absorption is related to potency. The relative importance of poor solubility and poor permeability towards the problem of poor oral absorption depends on the research approach used for lead generation. A "rational drug design" approach as exemplified by Merck advanced clinical candidates leads to time-dependent higher molecular weight, higher H-bonding properties, unchanged lipophilicity, and, hence, poorer permeability. A high throughput screening (HTS)-based approach as exemplified by unpublished data on Pfizer (Groton, CT) early candidates leads to higher molecular weight, unchanged H-bonding properties, higher lipophilicity, and, hence, poorer aqueous solubility. PMID- 11274894 TI - Progress in predicting human ADME parameters in silico. AB - Understanding the development of a scientific approach is a valuable exercise in gauging the potential directions the process could take in the future. The relatively short history of applying computational methods to absorption, distribution, metabolism and excretion (ADME) can be split into defined periods. The first began in the 1960s and continued through the 1970s with the work of Corwin Hansch et al. Their models utilized small sets of in vivo ADME data. The second era from the 1980s through 1990s witnessed the widespread incorporation of in vitro approaches as surrogates of in vivo ADME studies. These approaches fostered the initiation and increase in interpretable computational ADME models available in the literature. The third era is the present were there are many literature data sets derived from in vitro data for absorption, drug-drug interactions (DDI), drug transporters and efflux pumps [P-glycoprotein (P-gp), MRP], intrinsic clearance and brain penetration, which can theoretically be used to predict the situation in vivo in humans. Combinatorial synthesis, high throughput screening and computational approaches have emerged as a result of continual pressure on pharmaceutical companies to accelerate drug discovery while decreasing drug development costs. The goal has become to reduce the drop-out rate of drug candidates in the latter, most expensive stages of drug development. This is accomplished by increasing the failure rate of candidate compounds in the preclinical stages and increasing the speed of nomination of likely clinical candidates. The industry now understands the reasons for clinical failure other than efficacy are mainly related to pharmacokinetics and toxicity. The late 1990s saw significant company investment in ADME and drug safety departments to assess properties such as metabolic stability, cytochrome P-450 inhibition, absorption and genotoxicity earlier in the drug discovery paradigm. The next logical step in this process is the evaluation of higher throughput data to determine if computational (in silico) models can be constructed and validated from it. Such models would allow an exponential increase in the number of compounds screened virtually for ADME parameters. A number of researchers have started to utilize in silico, in vitro and in vivo approaches in parallel to address intestinal permeability and cytochrome P-450-mediated DDI. This review will assess how computational approaches for ADME parameters have evolved and how they are likely to progress. PMID- 11274895 TI - HTS in the new millennium: the role of pharmacology and flexibility. AB - Over the past decade, high throughput screening (HTS) has become the focal point for discovery programs within the pharmaceutical industry. The role of this discipline has been and remains the rapid and efficient identification of lead chemical matter within chemical libraries for therapeutics development. Recent advances in molecular and computational biology, i.e., genomic sequencing and bioinformatics, have resulted in the announcement of publication of the first draft of the human genome. While much work remains before a complete and accurate genomic map will be available, there can be no doubt that the number of potential therapeutic intervention points will increase dramatically, thereby increasing the workload of early discovery groups. One current drug discovery paradigm integrates genomics, protein biosciences and HTS in establishing what the authors refer to as the "gene-to-screen" process. Adoption of the "gene-to-screen" paradigm results in a dramatic increase in the efficiency of the process of converting a novel gene coding for a putative enzymatic or receptor function into a robust and pharmacologically relevant high throughput screen. This article details aspects of the identification of lead chemical matter from HTS. Topics discussed include portfolio composition (molecular targets amenable to small molecule drug discovery), screening file content, assay formats and plating densities, and the impact of instrumentation on the ability of HTS to identify lead chemical matter. PMID- 11274896 TI - Genomics and proteomics: the new millennium of drug discovery and development. AB - One of the most pressing issues facing the pharmaceutical and biotechnology industry is the tremendous dropout rate of lead drug candidates. Over the last two decades, several new genomic technologies have been developed in hopes of addressing the issues of target identification and lead candidate optimization. Gene expression microarray is one of these technologies and this review describes the four main formats, which are currently available: (a) cDNA; (b) oligonucleotide; (c) electrokinetic; and (d) fiberoptic. Many of these formats have been developed with the goal of screening large numbers of genes. Recently, a high-throughput array format has been developed where a large number of samples can be assayed using arrays in parallel. In addition, focusing on gene expression may be only one avenue in preventing lead candidate failure. Proteomics or the study of protein expression may also play a role. Two-dimensional polyacrylamide gel electrophoresis (2-DE) coupled with mass spectroscopy has been the most widely accepted format to study protein expression. However, protein microarrays are now being developed and modified to a high-throughput screening format. Examples of several gene and protein expression studies as they apply to drug discovery and development are reviewed. These studies often result in large data sets. Examples of how several statistical methods (principal components analysis [PCA], clustering methods, Shannon entropy, etc.) have been applied to these data sets are also described. These newer genomic and proteomic technologies and their analysis and visualization methods have the potential to make the drug discovery and development process less costly and more efficient by aiding to select better target and lead candidates. PMID- 11274897 TI - Current methodologies used for evaluation of intestinal permeability and absorption. AB - This review article will focus on the various techniques that are currently employed by drug discovery scientists in evaluating permeability/absorption of drug candidates during the drug candidate selection process. Various preclinical methodologies are available; each having advantages and disadvantages, but it is the judicious use of these techniques that can help identify drug candidates that will be well absorbed in humans. It is well recognized that the human intestinal permeability cannot be accurately predicted based on a single methodology (in vitro: tissue/cell culture, in situ, or in vivo). PMID- 11274898 TI - Present and future in vitro approaches for drug metabolism. AB - The 1980s through 1990s witnessed the widespread incorporation of in vitro absorption, distribution, metabolism, and excretion (ADME) approaches into drug development by drug companies. This has been exemplified by the integration of the basic science of cytochrome P450s (CYPs) into most drug metabolism departments so that information on the metabolic pathways of drugs and drug-drug interactions (DDIs) is no longer an academic exercise, but essential for regulatory submission. This has come about due to the application of a variety of new technologies and in vitro models. For example, subcellular fractions have been widely used in metabolism studies since the 1960s. The last two decades has seen the increased use of hepatocytes as the reproducibility of cell isolations improved. The 1990s saw the rejuvenation of liver slices (as new slicers were developed) and the utilization of cDNA expressed enzymes as these technologies matured. In addition, there has been considerable interest in extrapolating in vitro data to in vivo for parameters such as absorption, clearance and DDIs. The current philosophy of drug development is moving to a 'fail early--fail cheaply' paradigm. Therefore, in vitro ADME approaches are being applied to drug candidates earlier in development since they are essential for identifying compounds likely to present ADME challenges in the latter stages of drug development. These in vitro tools are also being used earlier in lead optimization biology, in parallel with approaches for optimizing target structure activity relationships, as well as identification of DDI and the involvement of metabolic pathways that demonstrate genetic polymorphisms. This would suggest that the line between discovery and development drug metabolism has blurred. In vitro approaches to ADME are increasingly being linked with high-throughput automation and analysis. Further, if we think of perhaps the fastest available way to screen for successful drugs with optimal ADME characteristics, then we arrive at predictive computational algorithms, which are only now being generated and validated in parallel with in vitro and in vivo methods. In addition, as we increase the number of ADME parameters determined early, the overall amount of data generated for both discovery and development will increase. This will present challenges for the efficient and fast interpretation of such data, as well as incorporation and communication to chemistry, biology, and clinical colleagues. This review will focus on and assess the nature of present in vitro metabolism approaches and indicate how they are likely to develop in the future. PMID- 11274899 TI - Fluorometric screening for metabolism-based drug--drug interactions. AB - Inhibition of cytochromes P-450 (CYP) is a principal mechanism for metabolism based drug interactions. In vitro methods for quantitatively measuring the extent of CYP inhibition are well-established. Classical methods use drug molecules as substrates and HPLC-based analysis. However, methodologies, which do not require HPLC separations for data acquisition generally offer higher throughputs and lower costs. Multiwell plate-based, direct, fluorometric assays for the activities of the five principal drug-metabolizing enzymes are available and parameters for the use of these substrates to measure CYP inhibition have been established. This methodology is quantitative, rapid, reproducible, and compatible with common high throughput screening instrumentation. This article describes approaches to establishing this methodology in a drug-discovery support program. PMID- 11274900 TI - Relationship of leptin level with metabolic disorders and hypertension in Japanese type 2 diabetes mellitus patients. AB - Leptin is considered to play an important role in the regulation of body weight and metabolism in obese individuals. However, the relationship of leptin with metabolic disorders or vascular complications in type 2 diabetic patients has yet to be elucidated. In this study, we investigated the association of leptin levels with clinical parameters (glycemic control, lipid levels, abdominal fat distribution) and investigated the leptin levels of diabetic patients with and without vascular complications in Japanese diabetic patients. In male and female patients, leptin levels were significantly associated with body mass index (BMI), percent body fat, insulin level, triglyceride (TG) level, total abdominal fat area (TFA), visceral fat area (VFS), and subcutaneous fat area (SFA). Only in male patients, leptin levels were inversely correlated with HDL-cholesterol, fasting plasma glucose (FPG), and HbA(1C). Leptin levels in male and female patients with hypertension were higher than in those without hypertension. Leptin levels of both males and females with angiopathy were not statistically different from those without angiopathy. In conclusion, leptin is involved in various metabolic disorders and hypertension, and we speculate that it may not be strongly associated with vascular complications in Japanese diabetic individuals. PMID- 11274901 TI - Identification of patients at risk for diabetic foot: a comparison of standardized noninvasive testing with routine practice at community diabetes clinics. AB - The aim of the study was the comparison of a simple standardized noninvasive examination of neuropathy and angiopathy with routine diagnostic practice in community diabetes clinics for the identification of patients at risk of foot ulceration. Consecutive patients (n=322), aged 30 years and more, with a diabetes duration of more than 5 years, were examined by trained podiatric nurses in six diabetes clinics over a 1-year period; 44 of these patients had active or previous foot ulcerations. We evaluated the differences between the routine diagnostic practice (based on the patient's medical history and a physical examination) and noninvasive testing of peripheral neuropathy [vibration perception threshold (VPT) and the Semmes-Weinstein 10-g monofilament wire system] and angiopathy [Doppler ankle/brachial index (ABI)]. Using receiver operating characteristic (ROC) analysis, we evaluated the sensitivity and specificity of noninvasive testing methods for identifying patients at risk and selecting the optimal diagnostic cutoff points. Patients with severe neuropathy, as determined by noninvasive testing (VPT > or =30 V and/or insensitivity to 10 g monofilament), had been diagnosed to have neuropathy in diabetes clinics in 54% of cases. Patients with angiopathy at risk of developing diabetic foot ulcers (ABI < or =0.8) had been diagnosed, in diabetes clinics, to have peripheral arterial disease in 50% (they reported claudications in 41%, had femoral artery bruits detected in 29% and nonpalpable peripheral pulsations in 12%). Our findings stress the importance of using standardized simple noninvasive testing methods to increase the accuracy of identifying patients at risk for the diabetic foot at the community level. PMID- 11274902 TI - The effect of insulin and sulodexide (Vessel Due F) on diabetic foot syndrome: pilot study in elderly patients. AB - OBJECTIVE: To assess the efficacy of insulin plus sulodexide (a mixture of 80% heparin-like substances and 20% dermatan sulphate) on diabetic ulcers, and its influence on foot skin microcirculation and diabetic neuropathy. RESEARCH DESIGN AND METHODS: Two groups of diabetic patients, suffering from severe neuropathy and ulceration, were randomly assigned to insulin (I) plus sulodexide (S) (n=12) or insulin plus placebo (P) (n=6) therapy, for 10 weeks. Laser Doppler assessment of foot skin flow (LDF), at rest and 30 or 60 s after arterial occlusion, and nerve conduction tests (sensorial evoked and motoric conduction potentials) have been evaluated in both groups. RESULTS: Postischaemic flow was 2.5 times shorter in ulcerated vs. non-ulcerated feet in diabetic patients. A significant increase in flows after 30 and 60 s ischaemia was detected in both groups at the end of therapy (IS group, ulcerated foot, LDF=60 s: from 99.1+/-14.3 to 218.6+/-28.6 PU, P<.001. IP group=from 110.5+/-13.0 to 164.8+/-15.4 PU, P<.05). The length of reactive hyperaemia was higher in IS vs. IP group (IS: from 30.3+/-2.9 to 43.9+/ 2.2 s, P<.001; IP: from 28.7+/-3.0 to 33.3+/-3.3 s, ns). Ninety-two percent of ulcers heals in a mean time of 46.4 days (IS group) vs. 83% and 63.0 days, respectively, in IP group. Nerve conduction studies have not demonstrated within- and between-group differences. CONCLUSIONS: Sulodexide and insulin improve the postischaemic skin flow in ulcerated feet, without affecting nerve conduction tests. The effect of sulodexide results additive to insulin; it is clinically relevant, in the view of the possibility of reducing the time needed to completely heal ulcers. The ultimate validation of these preliminary results requires extensive trials. PMID- 11274903 TI - Trandolapril restores circadian blood pressure variation in normoalbuminuric normotensive Type 1 diabetic patients. AB - A large proportion, from 30% to 50%, of diabetic patients frequently manifests loss of the normal diurnal variation of blood pressure, i.e. their blood pressure does not show at least 10% fall at night (non-dippers). It has been demonstrated that non-dippers are at increased risk of end-organ damage, in particular, renal and cardiovascular complications. As no reliable means of reversing impaired blood pressure variation has been established so far, we aimed at assessing the effect of a long-acting angiotensin-converting enzyme (ACE) inhibitor trandolapril on the disturbed circadian blood pressure rhythm in diabetics without hypertension or nephropathy. A total of 18 type 1 diabetes patients (8 male, 10 female), aged 33.5+/-4.8, with duration of diabetes 5.8+/-2.8 years and HbA(1c) 6.6+/-0.4% (range 5.8--7.1%) were enrolled into the study. Ten well matched type 1 diabetes patients served as an untreated control group. Twenty four-hour ambulatory blood pressure measurements (ABPM) were performed thrice in each subject: before trandolapril, 2 mg once daily in the morning, was started; after the first dose of the drug; and after 2 weeks of the treatment. Mean (+/ S.D.) values of systolic, diastolic blood pressure, night fall in systolic, and diastolic blood pressure in the treatment group were (1) at baseline: 124.0+/-5.8 mm Hg, 89.3+/-4.2 mm Hg, 3.0+/-2.2%, 5.1+/-3.8%; (2) after the first dose: 116.1+/-7.6 mm Hg (P<.01 vs. baseline), 82.6+/-6.7 mm Hg (P<.01 vs. baseline), 4.2+/-2.6%, 4.7+/-2.5%; and (3) after 14-day treatment: 116.6+/-8.1 mm Hg (P<.01 vs. baseline), 76.9+/-9.6 mm Hg (P<.01 vs. baseline), 17.6+/-6.9% (P<.01 vs. baseline and after 24-h values), 19.4+/-8.0% (P<.01 vs. baseline and after 24-h values), respectively. ABPM results for the untreated controls were similar in all three measurements. In conclusion, within 14 days of trandolapril treatment, circadian blood pressure variation was successfully restored in normoalbuminuric normotensive insulin-dependent diabetic patients. PMID- 11274904 TI - The control of blood glucose in the critical diabetic patient: a neuro-fuzzy method. AB - Conventional algorithms for regulating insulin infusion rates in those critical diabetic patients submitted to parenteral glucose and insulin infusions do not allow to approach near normal blood glucose (BG) levels since traditional control systems are not fully effective in complex nonlinear systems as BG control is. Thus, we applied fuzzy logic principles and neural network techniques to modify intravenous insulin administration rates during glucose infusion. Forty critically ill, fasted diabetic subjects submitted to glucose and potassium infusion entered the study. They were randomly assigned to two treatment regimes: in group A, insulin infusion rates were adjusted, every 4 h at any step between 1.5 and +1.5 U/h, according to a neuro-fuzzy nomogram; in control group B, insulin infusion rates were modified according to a conventional algorithm. In group A, BG was lowered below 10 mmol/l faster than in group B (8.2+/-0.7 vs. 13+/-1.8 h, P<.02). Mean BG was 7.8+/-0.2 in group A and 10.6+/-0.3 mmol/l in group B (P<.00001). BG values below 4.4 mmol/l were: A=5.8% and B=10.2%. BG values lower than 2.5 mmol/l had never been observed. In conclusion, the neuro fuzzy control system is effective in improving the BG control in critical diabetic patients without increasing either the number of BG determinations or the risk of hypoglycemia. PMID- 11274905 TI - Glucose-induced insulin resistance of phosphatidylinositol 3'-OH kinase and AKT/PKB is mediated by the hexosamine biosynthesis pathway. AB - Hyperglycemia is responsible for many of the vascular complications and metabolic derangements seen in diabetes. One potential regulator of the effects of glucose is the hexosamine biosynthesis pathway (HBP). Glutamine: fructose-6-phosphate amidotransferase (GFA), the first and rate-limiting enzyme in this pathway, catalyzes the transfer of an amino group from glutamine to fructose-6-phosphate to form glucosamine-6-phosphate. Overexpression of GFA in rat-1 fibroblasts results in insulin resistance for glycogen synthase (GS) activity, and renders these cells more sensitive to the effects of glucose. Using rat-1 cells, we examine further the mechanisms whereby hexosamines lead to insulin resistance. Insulin stimulated GS activity was found to occur via a PI-3 kinase (PI-3K) dependent pathway as wortmannin, an inhibitor of PI-3K, blocked insulin's ability to stimulate GS activity. Subsequently, we examined the effects of hexosamines on PI-3K and Akt/PKB activity. Cells were cultured in 1 mM glucose (low glucose, LG), 20 mM glucose (high glucose, HG), or 1 mM glucose plus 3 mM glucosamine (GlcN) for 16--20 h. After treatment with insulin (100 nM) for 5 min, cell extracts were assayed for IRS-1 associated and total PI-3K activity. At LG, insulin increased PI-3K activity by 43%. There was no insulin stimulation of PI 3K activity in cells cultured in HG or GlcN. There was a trend for IRS-1 protein levels to decrease in HG but not GlcN. PI-3K protein levels were not altered by HG or GlcN. Finally PKB activity was assayed. At LG, insulin stimulated PKB activity. Again, both HG and GlcN significantly reduced insulin's ability to stimulate PKB activity. We conclude that the hexosamine-mediated insulin resistance of GS activity seen in rat-1 cells is mediated by hexosamine regulation of PI-3K and PKB. PMID- 11274906 TI - Acute biochemical variations induced by calcium citrate and calcium carbonate in Type 2 diabetic patients: impaired calcium absorption in Type 2 diabetic patients with prolonged gastric emptying time. AB - Calcium supplementation is important in the treatment of osteoporosis, a disease that may also occur in diabetic patients. The acute effects of calcium supplementation and their relationship to gastric emptying time, however, have rarely been studied in type 2 diabetic patients. We evaluated the acute biochemical variations induced by the administration of two different calcium preparations, calcium citrate and calcium carbonate, in 16 (male/female: 13/3) Chinese diabetic patients. Serum free calcium, intact parathyroid hormone (i PTH), and amount of urinary excretion of calcium (uCal/uCr) were evaluated after a single dose of 1200 mg of elemental calcium in each preparation. The free calcium levels did not change significantly in either group. However, significant suppression of i-PTH after calcium citrate administration at 1 h (17.1+/-2.0 pg/ml, P=.023), and after calcium carbonate administration at 2 h (14.2+/-2.5 pg/ml, P=.000), was noted when compared with individual basal level (21.2+/-2.5 and 19.3+/-2.4 pg/ml, respectively). The suppressive effect on i-PTH lasted for 6 h after calcium citrate and 5 h after calcium carbonate preparation of the 6-h study period. After administration of calcium citrate, the uCal/uCr of 2-to-4-h collection was significantly higher than that of the basal and 0-to-2-h collections: 0.25+/-0.04 vs. 0.19+/-0.03, P=.025; and 0.25+/-0.04 vs. 0.19+/ 0.02, P=.014, respectively. A similar finding was observed for calcium carbonate: 0.23+/-0.03 vs. 0.18+/-0.02, P=.019; and 0.23+/-0.03 vs. 0.18+/-0.02, P=.011, respectively. We conclude that, in this group of Chinese type 2 diabetic patients in our study, the oral administration of 1200 mg elemental calcium in either calcium citrate or calcium carbonate preparation can induce a significant suppression of i-PTH. This may be helpful in preventing or treating osteoporosis. A prolonged gastric emptying time in these diabetic subjects may contribute to the non-significant alteration in free calcium levels after the administration of either calcium preparation. PMID- 11274907 TI - Differential absorption and distribution of epidermal growth factor and insulin like growth factor in diabetic NOD mice. AB - Previous studies have shown that absorption of growth factors occurs through the gastrointestinal tract and the oral cavity. The non-obese diabetic (NOD) mouse, a model for spontaneous development of type 1 insulin-dependent diabetes (IDDM), was evaluated for the absorption and systemic distribution of growth factors. Radiolabeled epidermal growth factor (EGF) and insulin-like growth factor, type I (IGF-I), were administered by gavage into the stomach or by lozenge into the sublingual vasculature of either diabetic or nondiabetic mice. After a time dependent uptake, the levels of absorption and distribution through the tissues were measured. A similar time course of EGF absorption following gavage administration was determined for NOD and C57BL/6 mice, with a maximum tissue distribution by 30-min post infusion. Diabetic NOD mice showed similar levels of IGF uptake and tissue distribution compared with nondiabetic NOD and normal healthy C57BL/6 mice, whether administered by gavage or sublingual lozenge. On the other hand, gavage uptake and tissue distribution of EGF was significantly higher in diabetic mice when compared to sublingual administration in nondiabetic NOD or C57BL/6 healthy control mice. These findings suggest that the overall potential uptake and distribution of saliva-derived growth factors in systemic wound-healing processes is retained with diabetes onset, and may offer a new avenue to treating this complication of diabetes. PMID- 11274908 TI - Immunology, climate change and vector-borne diseases. AB - Global climate change might expand the distribution of vector-borne pathogens in both time and space, thereby exposing host populations to longer transmission seasons, and immunologically naive populations to newly introduced pathogens. In the African highlands, where cool temperatures limit malaria parasite development, increases in temperature might enhance malaria transmission. St Louis encephalitis viral replication and the length of the transmission season depend upon ambient temperature. Warming temperatures in the American southwest might place at risk migratory, non-immune elderly persons that arrive in early fall to spend the winter. Warm temperatures might intensify or extend the transmission season for dengue fever. Immunologists should examine this interplay between human immunocompetence and vector-borne disease risks in a warmer world. PMID- 11274909 TI - Dendritic cells and transmission of HIV-1. AB - Sexual transmission of HIV-1 requires that small amounts of virus at mucosal sites of inoculation gain access to replication-permissive cells. Recent observations have increased our understanding of the mechanisms by which this relatively inefficient virus can exploit the migratory nature of dendritic cells to establish infection within the host. PMID- 11274910 TI - Bypassing IgE and targeting T cells for specific immunotherapy of allergy. AB - Specific immunotherapy (SIT) is a common treatment for allergic diseases. Despite its usage in clinical practice for nearly a century, more-rational and safer allergen preparations are required. Here, the underlying mechanisms and principles of allergen modification for the future use of SIT in the treatment of allergy are discussed. PMID- 11274911 TI - A new T-helper cell subset? PMID- 11274912 TI - A roll-call of monocytic gene induction. PMID- 11274913 TI - Co-stimulating allergy. PMID- 11274914 TI - What makes a bee sting deadly? PMID- 11274915 TI - 20% of sufferers could outgrow peanut allergy. PMID- 11274918 TI - New model of asthma proposed. PMID- 11274916 TI - Anti-CD20 antibody therapy is highly effective in the treatment of follicular lymphoma. PMID- 11274919 TI - MS therapy works in monkeys. PMID- 11274920 TI - RA drug receives EU approval. PMID- 11274921 TI - New study fails to find link between MMR and autism. PMID- 11274922 TI - Interaction of T cells with APCs: the serial encounter model. AB - Primary immune responses are initiated by specific physical interaction of antigen-specific T cells and professional antigen-presenting cells (APCs). Productive interactions can be a dynamic process that combines physical T-cell binding to APCs with vigorous crawling across and scanning of the APC surface, resulting in signal induction. After T-cell detachment, subsequent migratory contacts to the same or neighboring dendritic cells (DCs) allow the accumulation of sequential signals and interaction time. Here, we develop a serial encounter model of T-cell activation and discuss how the summation of multiple signals provides an efficient strategy to control an ongoing immune response. PMID- 11274923 TI - Environmental control of immunological synapse formation and duration. AB - The coordination of T-cell migration and antigen recognition is crucial for an effective immune response. We have proposed that this coordination is achieved by formation of an immunological synapse between the T cell and the antigen presenting cell (APC). Our view contrasts with the serial encounter model also proposed in this issue of Trends in Immunology, which is based on transient T cell-APC interactions when surrounded by collagen. Here, we propose a model that reconciles immunological synapse formation and serial encounters based on environmental control of immunological synapse formation. PMID- 11274924 TI - Tapasin: an ER chaperone that controls MHC class I assembly with peptide. AB - The stable assembly of MHC class I molecules with peptides in the endoplasmic reticulum (ER) involves several accessory molecules. One of these accessory molecules is tapasin, a transmembrane protein that tethers empty class I molecules to the peptide transporter associated with antigen processing (TAP). Here, evidence is presented that tapasin retains class I molecules in the ER until they acquire high-affinity peptides. PMID- 11274925 TI - Fibroblasts regulate the switch from acute resolving to chronic persistent inflammation. AB - Fibroblasts are important sentinel cells in the immune system and, here, it is proposed that these cells play a critical role in the switch from acute inflammation to adaptive immunity and tissue repair. It is suggested that chronic inflammation occurs because of disordered fibroblast behaviour in which failure to switch off their inflammatory programme leads to the inappropriate survival and retention of leukocytes within inflamed tissue. PMID- 11274926 TI - IgA and the IgA Fc receptor. AB - IgA has traditionally been regarded a non-inflammatory antibody. This might indeed be true for secretory IgA (SIgA), which exerts its function at mucosal surfaces where commensal microorganisms and dietary antigens prevail. Serum IgA, however, potently triggers (pro)-inflammatory activity upon binding to the myeloid IgA receptor, FcalphaRI. Here, new insights in the roles of IgA and FcalphaRI are addressed and a model integrating the various functions of IgA in immunity is discussed. PMID- 11274927 TI - VAP-1: an adhesin and an enzyme. AB - Leukocyte extravasation from the blood into tissues is of paramount importance for normal immunosurveillance and in mounting adequate inflammatory responses. Multiple traditional adhesion molecules and chemoattractants on leukocytes and endothelial cells are involved in the emigration process. Vascular adhesion protein 1 (VAP-1) is a nonclassical inflammation-inducible endothelial molecule involved in leukocyte-subtype-specific rolling under physiological shear. Molecularly, VAP-1 belongs to a special class of cell surface amino oxidases. The enzymatic reaction itself and the biologically active end products can potentially regulate the adhesive status of the vessel wall. Thus, VAP-1 is an ectoenzyme that has inter-related adhesive and enzymatic functions in regulating physiological trafficking and inflammation. PMID- 11274933 TI - Improved treatment of coronary heart disease by implementation of a Cardiac Hospitalization Atherosclerosis Management Program (CHAMP). AB - Despite scientific evidence that secondary prevention medical therapies reduce mortality in patients with established coronary artery disease, these therapies continue to be underutilized in patients receiving conventional care. To address this issue, a Cardiac Hospital Atherosclerosis Management Program (CHAMP) focused on initiation of aspirin, cholesterol-lowering medication (hydroxymethylglutaryl coenzyme A [HMG CoA] reductase inhibitor titrated to achieve low-density lipoprotein [LDL] cholesterol < or =100 mg/dl), beta blocker, and angiotensin converting enzyme (ACE) inhibitor therapy in conjunction with diet and exercise counseling before hospital discharge in patients with established coronary artery disease. Treatment rates and clinical outcome were compared in patients discharged after myocardial infarction in the 2-year period before (1992 to 1993) and the 2-year period after (1994 to 1995) CHAMP was implemented. In the pre- and post-CHAMP patient groups, aspirin use at discharge improved from 68% to 92% (p <0.01), beta blocker use improved from 12% to 62% (p <0.01), ACE inhibitor use increased from 6% to 58% (p <0.01), and statin use increased from 6% to 86% (p <0.01). This increased use of treatment persisted during subsequent follow-up. There was also a significant increase in patients achieving a LDL cholesterol < or =100 mg/dl (6% vs 58%, p <0.001) and a reduction in recurrent myocardial infarction and 1-year mortality. Compared with conventional guidelines and care, CHAMP was associated with a significant increase in use of medications that have been previously demonstrated to reduce mortality; more patients achieved an LDL cholesterol < or =100 mg/dl, and there were improved clinical outcomes in patients after hospitalization for acute myocardial infarction. PMID- 11274928 TI - The expanding world of co-stimulation: the two-signal model revisited. AB - The crucial role for CD28, its homolog CTLA-4 and their binding partners B7-1 and B7-2 in the generation of effective T-cell responses has been well documented. Recently, two new pairs of the CD28/B7 families were identified. The ability of these molecules to regulate T-cell expansion and effector function and the dynamic integration of the co-stimulatory and T-cell receptor signals are just beginning to be explored. Understanding these processes will be crucial for designing clinically relevant approaches to manipulate the adaptive immune system. PMID- 11274934 TI - Comparative effects of three beta blockers (atenolol, metoprolol, and propranolol) on survival after acute myocardial infarction. AB - The beneficial impact of beta blockade after an acute myocardial infarction (AMI) is clear, but beta-adrenergic blockers differ in multiple characteristics, including lipophilicity and selectivity. The impact of these factors on the effects of beta blockade is unknown. We therefore compared the effects of different beta blockers on mortality after AMI. Charts of 201,752 patients with AMI were abstracted by the Cooperative Cardiovascular Project, a quality assurance program sponsored by the Health Care Financing Administration. Of the 69,338 patients prescribed beta blockers, we compared mortality of patients receiving different beta-adrenergic blockers using the Cox proportional-hazards model accounting for multiple factors that might influence survival. The mortality rates of the 2 selective agents, metoprolol and atenolol, were virtually identical (13.5% and 13.4% 2-year mortality, respectively). Compared with metoprolol, patients discharged on propranolol had a slightly increased mortality (15.9% 2-year mortality), which may be related to undetected differences at baseline. Survival with all of the drugs was superior to the 23.9% 2-year mortality seen in patients not receiving beta blockers. Beta blockade overall was associated with a 40% improvement in survival. Although the use of beta blockade after AMI has major prognostic importance, the present study suggests that the specific beta blocker chosen will have little influence on mortality. PMID- 11274935 TI - Effect of orlistat-assisted weight loss in decreasing coronary heart disease risk in patients with syndrome X. AB - This study describes the changes in risk factors for coronary heart disease in obese persons with syndrome X after orlistat-assisted weight loss. Data were available for 1,700 patients who completed 52 weeks of weight loss; 128 were defined as having syndrome X by being in the quintile with the highest plasma triglyceride levels (>2.2 mM/L) and the lowest high-density lipoprotein cholesterol (HDL, <1.0 mM/L) concentrations. Initial characteristics of those with syndrome X were similar to the 119 subjects (non-syndrome X) in the lowest quintile of plasma triglyceride (<0.975 mM/L) and highest quintile of HDL cholesterol (>1.5 mM/L). Subjects were placed on a calorie-restricted diet, and randomized to receive orlistat or placebo. Initial values were higher in those with syndrome X for diastolic blood pressure (p = 0.03), plasma insulin (p = 0.0001), triglyceride (p = 0.0001) concentrations, and ratio of low-density lipoprotein cholesterol to HDL cholesterol (p = 0.0001), and were lower for HDL cholesterol (p = 0.001) concentrations. Weight loss was greater in both groups of orlistat-treated patients (p = 0.026); in those with syndrome X, it was associated with a significant reduction in plasma insulin (p = 0.019) and triglyceride (p = 0.0001) concentrations, an increase in HDL cholesterol concentration, and a decrease in low-density lipoprotein/HDL cholesterol ratio (p = 0.0001). There were no significant changes in plasma insulin, triglycerides, or HDL cholesterol concentration in the non-syndrome X group. In conclusion, weight loss attenuates coronary heart disease risk factors in obese persons with syndrome X, and the risk factor reduction is enhanced with administration of orlistat. PMID- 11274936 TI - Role of coronary interventional procedures in improved postinfarction survival in the 1990s. AB - The contribution of increased use of same-admission percutaneous coronary interventional procedures to recent improvements in hospital survival of patients with acute myocardial infarction (AMI) remains unclear. Patients with International Classification of Diseases codes for AMI (code 410), who were admitted to the emergency coronary care unit and underwent an initial episode of treatment, were studied over the 9-year period 1990 to 1998 (n = 2,628). Three triennia between 1990 and 1998 were compared. Trends in risk, the use of procedures, and hospital outcomes were analyzed. Hospital mortality was 33% lower (p <0.02) in the third triennium (5.8%) than in the earlier 2 triennia (8.7%), equivalent to an absolute reduction of 29 hospital deaths/1,000 patients treated. The lower hospital mortality was not due to: (1) shorter hospital stays (reduction in mortality was primarily in the first 3 hospital days), (2) treatment of lower risk subjects (a risk score based on age, gender, and presence of diabetes increased between the first and third triennia), or (3) use of in hospital interventional procedures (although the use of percutaneous coronary intervention more than doubled in the third triennium, most procedures were performed in patients with a 1% risk of hospital death). We conclude from this study that there has been a substantial improvement over a 9-year period in early case fatality after AMI, but that this cannot be attributed to the increased use of in-hospital coronary interventions, which were largely performed on low-risk patients. PMID- 11274937 TI - Initial and long-term results of directional coronary atherectomy in unprotected left main coronary artery. AB - Angioplasty in the unprotected left main coronary artery (LMCA) has been controversial. Recently, several studies have suggested that new procedures and devices such as directional coronary atherectomy (DCA) and stents may change this situation. Although there are many reports of unprotected LMCA stenting, there are few reports of DCA of this lesion. Therefore, initial and long-term results were evaluated in 101 patients who underwent DCA for unprotected LMCA in our hospital. Emergency procedures were performed in 15 patients and electively in 86 patients. Scheduled angiographic follow-up was routinely performed, and all patients were clinically followed for >4 months after DCA. Technical success was achieved in 99%, and in-hospital outcomes were cardiac death (2%), noncardiac death (4%), Q-wave myocardial infarction (1%), non-Q-wave myocardial infarction (8.9%), coronary artery bypass grafting (0%), and repeat angioplasty (4%). In hospital results varied considerably, depending on presentation. In-hospital mortality was significantly higher in the emergency, left ventricular ejection fraction < or =35%, and high-risk surgical subgroups. The angiographic restenosis rate was 20.4% at follow-up, and its predictor was postminimal lumen diameter by multivariate analysis. Mean clinical follow-up was 2.8 years; estimated 1- and 3 year survival rates were 87% and 80.7%, respectively. The cardiac survival rate of the low-risk surgical subgroup was significantly higher than that of the high risk surgical subgroup (p <0.05). Thus, our data show that DCA can be performed safely and effectively in unprotected LMCA with an acceptable low restenosis rate and high survival rate. PMID- 11274938 TI - Outcomes and early revascularization for patients > or = 65 years of age with cardiogenic shock. AB - Hospital survival of patients with acute myocardial infarction (AMI) complicated by cardiogenic shock has improved during recent years. It is unclear whether this mortality benefit also applies to elderly patients with cardiogenic shock. Elderly residents (age > or = 65 years) of the Worcester, Massachusetts metropolitan area (1990 census population = 437,000) hospitalized with confirmed AMI and cardiogenic shock in all metropolitan Worcester, Massachusetts hospitals between 1986 and 1997 constituted the sample of interest. We examined the use of coronary reperfusion strategies, adjunctive therapy, and hospital mortality in a cohort of 166 cardiogenic patients treated early in the reperfusion era (1986 to 1991) compared with 144 patients with AMI treated approximately 1 decade later (1993 to 1997). There was a significant increase in the use of an early revascularization strategy over time (2% vs 16%, p <0.001). Marked increases in use of antiplatelet therapy, beta blockers, and angiotensin-converting enzyme inhibitors were also observed over the decade-long experience. In-hospital case fatality declined significantly over time, from 80% (1986 to 1991) to 69% (1993 to 1997) in elderly patients who developed cardiogenic shock (p = 0.03). After adjusting for differences in potentially confounding prognostic characteristics between patients hospitalized in the 2 study periods, an even more pronounced reduction in hospital mortality (42%) was observed for the most recently hospitalized cohort. The most powerful predictor of in-hospital survival was use of an early revascularization approach to treatment. Thus, hospital mortality has declined for patients > or = 65 years of age with AMI complicated by cardiogenic shock, and this decline has occurred in the setting of broader use of early revascularization and adjunctive medical therapy for this high-risk population. PMID- 11274939 TI - Effectiveness of excimer laser coronary angioplasty in acute myocardial infarction or in unstable angina pectoris. AB - This study was conducted to evaluate the feasibility, safety, and acute results of percutaneous excimer laser coronary angioplasty (ELCA) in acute coronary syndromes. Fifty-nine patients were treated with ELCA (308 nm), including 33 patients with unstable angina pectoris (UAP) (35 vessels with 39 lesions) and 26 patients with acute myocardial infarction (AMI) (26 vessels with 29 lesions). In each patient the target lesion had a complex morphology. Overall, 71% of the patients had contraindications for pharmacologic thrombolytic agents or glycoprotein IIb/IIIa receptor antagonists. All patients received adjunct balloon dilation followed by stent implantation in 88% of patients with AMI versus 76% of patients with UAP (p = NS). Quantitative angiography was performed at an independent core laboratory; 86% laser success and 100% procedural success was achieved in the AMI group versus 87% laser success and 97% procedural success in the UAP group (p = NS). In the AMI group, the minimal luminal diameter increased from 0.77 +/- 0.56 to 1.44 +/- 0.47 mm after lasing to a final 2.65 +/- 0.47 mm versus 0.77 +/- 0.38 to 1.35 +/- 0.4 mm after lasing to 2.66 +/- 0.5 mm final in the UAP group. A prelaser percent stenosis of 76 +/- 17% for the AMI group versus 70 +/- 16% for the UAP group (p = NS) was decreased after lasing to 52 +/- 16% for the AMI group versus 51 +/- 14% for the UAP group (p = NS) and to a final stenosis of 15 +/- 17% for the AMI group versus 12 +/- 15% for the UAP group (p = NS). A 96% laser-induced reduction of thrombus burden area was achieved in the AMI group versus 97% in the UAP group (p = NS). Preprocedure Thrombolysis In Myocardial Infarction flow of 1.3 +/- 0.9 in the AMI group versus 2.3 +/- 1.2 for the UAP group (p = 0.01) increased to a final flow of 3.0 +/- 0 for the AMI group versus 3.0 +/- 0 for the UAP group (p = NS). There were no deaths, cerebrovascular accident, emergency bypass surgery, acute closure, major perforation or major dissection, distal embolization, or bleeding complications in either group. One patient with AMI had localized perforation (caused by guidewire) without sequelae and 1 patient with UAP had an abnormal increase in creatine kinase levels. All 59 patients survived the laser procedure, improved clinically, and were discharged. Thus, early experience in patients with acute coronary syndromes suggest that percutaneous ELCA is feasible and safe. PMID- 11274940 TI - Effectiveness of and adverse events after percutaneous coronary intervention in patients with mild versus severe renal failure. AB - Patients with renal failure undergoing percutaneous coronary intervention (PCI) experience reduced procedural success rates and increased in-hospital and long term follow-up major adverse cardiac events. This study was designed to determine whether the severity of preprocedural renal failure influences the outcomes of patients with renal failure undergoing PCI. We compared the immediate and long term outcomes of 192 patients with mild renal failure (creatinine 1.6 to 2.0 mg/dl, mean 1.76) with those of 131 patients with severe renal failure (creatinine >2.0 mg/dl, mean 2.90), selected from 3,334 consecutive patients undergoing PCI between 1994 and 1997. Although the overall population with renal failure represents a high-risk group, the severe renal failure cohort had a higher incidence of hypertension, multivessel disease, prior coronary bypass surgery, vascular disease, and congestive heart failure (all p values <0.05), yet had similar angiographic characteristics. Procedural success was higher in the group with severe renal failure (93.7% vs 87.7%, p = 0.04). There were no statistically significant differences in in-hospital mortality (11.5% vs 9.9%, p = 0.7), Q-wave myocardial infarction (0.5% vs 0%, p = 0.4), emergent bypass surgery (0% vs 0%, p = 1.0), and in-hospital major adverse cardiac events (11.5% vs 9.9%, p = 0.7) between the mild and severe renal groups, respectively. Kaplan Meier analyses showed no statistically significant difference in long-term survival (log rank test, p = 0.1) or event-free survival (log rank test, p = 0.3) between the 2 groups. Finally, creatinine was not identified as an independent predictor of in-hospital or long-term follow-up major adverse cardiac events. In our high-risk population, patients with mild renal insufficiency undergoing PCI experience major adverse outcomes in the hospital and at long-term follow-up similar to those of patients with severe renal failure. PMID- 11274941 TI - Quantitative analysis of myocardial perfusion changes with transmyocardial laser revascularization. AB - Transmyocardial laser revascularization (TLR) is a technique of creating left ventricular transmural channels in patients with refractory angina. We aimed to measure perfusion changes quantitatively using technetium-99m methoxyisobutyl isonitrile. Perfusion scans were performed on 94 TLRs and in 94 control patients at rest and during exercise at assessment, and 3-, 6-, and 12-month follow-up. A serial set of scans allowed direct comparison of each patient over all visits. Bull's-eyes were divided into 5 anatomic regions and a 20-region model. Severity values were calculated for rest, stress, and each cardiac region using a threshold of 1 for analysis. Higher scores indicated greater severity of ischemia and lower perfusion. At 3-month follow-up, the severity was significantly worse during TLR than in control patients both during stress (0.172 +/- 0.003 and 0.161 +/- 0.003, respectively, p = 0.007) and at rest (0.170 +/- 0.003 and 0.158 +/- 0.003, respectively, p = 0.002). At 6 months, severity during stress was 0.176 +/ 0.003 with TLR and 0.162 +/- 0.003 in controls (p = 0.001), with no significant difference at rest. At 12 months, there was no significant difference between TLR and control groups at stress and rest. Regional severity deteriorates during TLR compared with control patients anteriorly (p = 0.001, p = 0.0016, p = 0.005 at 3, 6, and 12 months), apically (p = 0.005, p = 0.0046, p = 0.032, respectively), and laterally (p <0.0001, p = 0.001, p = 0.002, respectively). An apparent improvement is observed in the inferoseptal region at 6- and 12-month follow-up an area not lasered. Thus, TLR appears to produce deterioration in resting myocardial perfusion in lasered regions, and improvement in nonlasered regions, with no difference in exercise-induced myocardial ischemia compared with that in control patients. PMID- 11274942 TI - Assessment of the exercise electrocardiogram in women versus men using tomographic myocardial perfusion imaging as the reference standard. AB - The exercise electrocardiogram (ECG) is widely believed to be less accurate in women, primarily due to a high prevalence of false-positive tests. The purpose of this study was to examine the relative accuracy of the exercise ECG in women versus men in 8,671 patients (3,213 women, 5,458 men) using myocardial perfusion imaging as the reference standard. More women (14%) than men (10%) had a false positive ECG (p <0.001), but the absolute difference was relatively small. The false-negative rate was considerably lower in women (17% vs 32%, p <0.001). Compared with men, women had lower test sensitivity (30% vs 42%, p <0.001) and positive predictive value (34% vs 70%, p <0.001) but higher specificity (82% vs 78%, p = 0.002), negative predictive value (78% vs 52%, p <0.001), and accuracy (69% vs 58%, p <0.001). In patients with a false-negative exercise ECG, "high risk" scans were less prevalent in women (12% vs 19%, p <0.001). In the smaller subset of patients referred for coronary angiography (205 women, 838 men), the false-positive electrocardiographic rate was again higher in women (13% vs 7%, p = 0.003), but neither specificity (69% vs 74%, p = NS) nor accuracy (60% vs 66%, p = NS) was different between the sexes. Thus, the percentage of patients with a false-positive exercise ECG was higher in women than men but low in absolute terms (<15%) for both sexes. Test specificity was not lower in women. These results suggest that gender should not be a major determinant for selecting stress imaging over standard treadmill testing. PMID- 11274943 TI - Comparison of endocardial electromechanical mapping with radionuclide perfusion imaging to assess myocardial viability and severity of myocardial ischemia in angina pectoris. AB - The assessment of left ventricular electromechanical activity using a novel, nonfluoroscopic 3-dimensional mapping system demonstrates considerable differences in electrical and mechanical activities within regions of myocardial infarction or ischemia. We sought to determine whether these changes correlate with indexes of myocardial perfusion, viability, or ischemia. A 12-segment comparative analysis was performed in 61 patients (45 men, 61 +/- 12 years old) with class III to IV angina, having reversible and/or fixed myocardial perfusion defects on single-photon emission computed tomographic perfusion imaging. A dual isotope protocol was used, consisting of rest and 4-hour redistribution thallium images followed by adenosine technetium-99m sestamibi imaging. Average rest endocardial unipolar voltage (UpV) and local shortening (LS) mapping values were compared with visually derived perfusion scores. There was gradual and proportional reduction in regional UpV and LS in relation to thallium-201 uptake score at rest (p = 0.0001 and p = 0.0002, respectively) and redistribution studies (p = 0.0001 and p = 0.003, respectively). UpV > or = 7.4 mV and LS > or = 5.0% had a sensitivity of 78% and 65%, respectively, with a specificity of 68% and 67% for detecting viable myocardium. UpV values of 12.3 and 5.4 mV had 90% specificity and sensitivity, respectively, to predict viable tissue. UpV, but not LS, values differentiated between normal segments and those with adenosine induced severe perfusion defects (11.8 +/- 5.3 vs 8.8 +/- 4.1 mV, p = 0.005). Catheter-based left ventricular assessment of electromechanical activity correlates with the degree of single-photon emission computed tomographic perfusion abnormality and can identify myocardial viability with a greater accuracy than myocardial ischemia. PMID- 11274944 TI - Effect of postoperative atrial fibrillation on length of stay after cardiac surgery (The Postoperative Atrial Fibrillation in Cardiac Surgery study [PACS(2)]. AB - Atrial fibrillation (AF) after cardiac surgery is thought to increase length of stay (LOS). A clinical pathway focused on the management of postoperative AF, including prophylaxis with beta blockers, was implemented to assess the effect of AF on LOS after cardiac surgery. Data were obtained on consecutive cardiac surgery patients in preoperative normal sinus rhythm, no prior history of AF, and no chronic antiarrhythmic therapy from January to May 1995 (control) and November 1996 to June 1997 (pathway). Statistical analysis was performed to assess the effect of postoperative AF on the LOS, clinical outcomes, and cost after cardiac surgery. Despite the clinical pathway, the LOS (7 days for both periods; p = 0.12) and incidence of AF (28.9% vs 28.4%; p = 0.92) remained unchanged. Unadjusted direct costs were 15% higher in the pathway period (p <0.001). Increased rates of beta-blocker therapy had a marginal effect on the incidence of postoperative AF, except in the group who only underwent primary coronary artery bypass graft surgery (31.2% vs 25.3%; p = 0.31). Multivariate analysis revealed that AF contributed only 1 to 1.5 days to the LOS. Thus, this investigation represents the most recent analysis of the effects of postoperative AF on LOS, clinical outcomes, and cost after cardiac surgery. Unlike prior studies, the impact of postoperative AF is less prominent in the current era of cardiac surgical care regardless of the presence of a clinical pathway addressing AF. PMID- 11274945 TI - Outcomes after radiofrequency catheter ablation of atrial tachycardia. AB - The purpose of this study was to evaluate the efficacy, safety, and clinical benefit of radiofrequency catheter ablation (RFCA) in a large series of patients with atrial tachycardia (AT). The determinants of success or failure of RFCA in AT remain unclear. We evaluated the results of radiofrequency ablation in 73 women and 32 men (mean age 48 +/- 19 years) with AT. Mapping techniques were based on identification of the earliest endocardial atrial electrogram recorded during AT. AT originated from the right atrium in 91 patients and from the left atrium in 14. The cardiac ventricles were dilated in 12 patients. AT ablation was successful in 80 patients (77%) regardless of the site of origin. Age, gender, rate of tachycardia, temperature achieved during application, or presence of tachycardiomyopathy were not significant determinants of acute success by univariate analysis. There was a significantly higher acute success rate of ablation in patients with paroxysmal (88%, 45 of 51) and permanent (71%, 30 of 42) forms than in patients with repetitive forms of AT (41%, 5 of 12) (p <0.005). The mean local endocardial electrogram time (relative-to-surface P-wave onset) was -47 +/- 17 ms at successful ablation sites and -29 +/- 21 ms at unsuccessful sites (p <0.03). Ablation was unsuccessful in 25 cases. Thus, RFCA of AT can be performed with a high acute success rate. Patients with repetitive forms and those with multifocal origin had a lower acute success rate. The highest incidence of recurrences was found in anterior right atrial foci. PMID- 11274946 TI - Usefulness of ST depression in ventricular premature complexes to predict myocardial ischemia. PMID- 11274947 TI - Comparison of results of rotational atherectomy for diffuse coronary artery disease in diabetics versus nondiabetics. PMID- 11274948 TI - Quantitative evaluation of regional left ventricular wall motion using color kinesis after percutaneous transluminal myocardial revascularization. PMID- 11274949 TI - Comparison of functional testing patterns after coronary artery bypass grafting in Canada and in the United States. PMID- 11274950 TI - Predictors and mode of detection of transvenous lead malfunction in implantable defibrillators. PMID- 11274951 TI - Comparability of nonlinear measures of heart rate variability between long- and short-term electrocardiographic recordings. PMID- 11274953 TI - Usefulness of short-term symptomatic status as a predictor of mid- and long-term outcome after balloon mitral valvuloplasty. PMID- 11274952 TI - Effect of the antimalarial drug halofantrine in the long QT syndrome due to a mutation of the cardiac sodium channel gene SCN5A. PMID- 11274954 TI - Effect of balloon aortic valvuloplasty of congenital aortic stenosis in children in regression of left ventricular mass. PMID- 11274955 TI - Aortic root dilation in Kawasaki disease. PMID- 11274957 TI - Comparison of serous and bloody pericardial effusion as an ominous prognostic sign. PMID- 11274956 TI - Circadian and circannual rhythm of nonfatal pulmonary embolism. PMID- 11274959 TI - Significance of recurrent pain in acute type B aortic dissection. PMID- 11274958 TI - Effect of arginine on cyclosporine-induced systemic hypertension after cardiac transplantation in the young. PMID- 11274960 TI - Mitral annulus velocity in the noninvasive estimation of left ventricular peak dP/dt. PMID- 11274961 TI - Molecular targets for pharmacological cytoprotection. AB - Cell death is common to many pathological conditions. In the past two decades, research into the mechanism of cell death has characterized the cardinal features of apoptosis and necrosis, the two distinct forms of cell death. Studies using in vivo disease models have provided evidence that apoptosis is induced by an array of pathological stimuli. Thus, molecular components of the machinery of apoptosis are potential pharmacological targets. The mechanism of apoptosis can be dissected into: (i) the initiation and signaling phase, (ii) the signal amplification phase, and (iii) the execution phase. Reflecting on the diversity of apoptotic stimuli, the initiation and signaling phase utilizes a variety of molecules: free radicals, ions, plasma membrane receptors, members of the signaling kinase cascades, transcription factors, and signaling caspases. In most of the apoptotic scenarios, impairment of mitochondrial function is an early event. Dysfunctioning mitochondria release more free radicals and hydrolytic enzymes (proteases and nucleases), amplifying the primary death signal. In the final phase of apoptosis, executioner caspases are activated. Substrates of the executioner caspases include nucleases, members of the cellular repair apparatus, and cytoskeletal proteins. Partial proteolysis of these substrates leads to distinctive morphological and biochemical changes, the hallmarks of apoptosis. The first steps toward pharmacological utilization of specific modifiers of apoptosis have been promising. However, since the potential molecular targets of cytoprotective therapy play important roles in the maintenance of cellular homeostasis, specificity (diseased versus healthy tissue) of pharmacological modulation is the key to success. PMID- 11274962 TI - Protective effects of morphine in peroxynitrite-induced apoptosis of primary rat neonatal astrocytes: potential involvement of G protein and phosphatidylinositol 3-kinase (PI3 kinase). AB - Opiates, such as morphine, have been used extensively in the clinical management of pain due to their potent analgesic effect. Astrocytes, representing a major non-neuronal cell population in the CNS, contain opioid receptors that are actively involved in several brain functions. This study was designed to evaluate the effects by which morphine, a preferential mu-opioid receptor agonist, contributes to cytotoxicity of nitric oxide (NO) species, including NO and peroxynitrite (ONOO-), in primary rat neonatal astrocytes. Primary astrocytes isolated from the cerebral cortex of 1- to 2-day-old Sprague-Dawley rats were treated with morphine, naloxone, and 3-morpholinosydnonimine (SIN-1), a donor of peroxynitrite. Morphine significantly protected primary rat astrocytes from apoptosis mediated by sodium nitroprusside, an NO donor, and SIN-1 in a dose dependent manner, whereas it did not in other types of cells including C6 glioma, RAW 264.7, and HL-60 cells. Moreover, naloxone antagonized the protective effects of morphine on SIN-1-induced apoptosis. Morphine also inhibited the nuclear condensation and fragmentation of SIN-1-treated cells that was antagonized by naloxone pretreatment. The protective role of morphine in SIN-1-induced apoptosis was dependent on an intracellular antioxidant system such as GSH. Furthermore, the effects of morphine on SIN-1-induced cytotoxicity were prohibited by pretreatment with the G(i) protein inhibitor, pertussis toxin, and the phosphatidylinositol 3-kinase (PI3 kinase) inhibitors, wortmannin and LY294002. Taken together, these results suggest that morphine may protect primary rat astrocytes from apoptosis by NO species via the signaling cascades that involve both G protein and PI3 kinase. PMID- 11274963 TI - Noncompetitive inhibition by camphor of nicotinic acetylcholine receptors. AB - The effect of camphor, a monoterpenoid, on catecholamine secretion was investigated in bovine adrenal chromaffin cells. Camphor inhibited [3H]norepinephrine ([3H]NE) secretion induced by a nicotinic acetylcholine receptor (nAChR) agonist, 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), with a half-maximal inhibitory concentration (IC50) of 70 +/- 12 microM. In addition, camphor inhibited the rise in cytosolic calcium ([Ca2+]i) and sodium ([Na+]i) induced by DMPP with IC50 values of 88 +/- 32 and 19 +/- 2 microM, respectively, suggesting that the activity of nAChRs is also inhibited by camphor. On the other hand, binding of [3H]nicotine to nAChRs was not affected by camphor. [Ca2+]i increases induced by high K+, veratridine, and bradykinin were not affected by camphor. The data suggest that camphor specifically inhibits catecholamine secretion by blocking nAChRs without affecting agonist binding. PMID- 11274964 TI - Role of proteasomal degradation in the cell cycle-dependent regulation of DNA topoisomerase IIalpha expression. AB - 1DNA topoisomerase II (topo II) is a nuclear enzyme that modifies DNA topology and also serves as a target to mediate the cytotoxicity of several antineoplastic agents. Several reports have demonstrated that a reduction of topo II is associated with reduced sensitivity to these agents. Topo II exists as two isoforms in mammalian cells: topo IIalpha and topo IIbeta. In MCF-7 cells, the half-life (mean +/- SEM) values of topo IIalpha and topo IIbeta in situ were 6.6 +/- 0.3 and 17.6 +/- 2.3 hr, respectively, as determined by [(35)S]methionine/cysteine pulse-chase analysis. Degradation of topo IIalpha in situ was abrogated by the presence of proteasome inhibitors, and the relative activities were carbobenzoxy-leucyl-leucyl-leucinal (MG132) > carbobenzoxy-leucyl leucyl-norvalinal (MG115) > ALLN congruent with lactacystin. ATP-dependent degradation of topo IIalpha, but not topo IIbeta, was observed in extracts of asynchronously dividing HeLa and MCF-7 cells. Furthermore, degradation of topo IIalpha was abrogated by the proteasome inhibitors MG132 and MG115, but not by lactacystin, in extracts of asynchronously dividing MCF-7 cells. Finally, degradation of topo IIalpha, but not topo IIbeta, was observed to occur in a cell cycle-dependent fashion, in extracts of synchronized HeLa cells, with maximal loss of the alpha isoform occurring 2 hr after release from mitotic arrest. This degradation of topo IIalpha appeared to be facilitated by an ATP-dependent activity. Furthermore, high molecular weight bands (>200 kDa), which may represent polyubiquitinated-topo IIalpha conjugates, were also detected in extracts of synchronized HeLa cells. This study provides evidence for a role of the ubiquitin-proteasome pathway in the cell cycle-dependent regulation of topo IIalpha expression. PMID- 11274965 TI - Modulation of inositol 1,4,5-trisphosphate binding to the various inositol 1,4,5 trisphosphate receptor isoforms by thimerosal and cyclic ADP-ribose. AB - Three different genes encode the inositol 1,4,5-trisphosphate (IP3) receptor (IP3R), an intracellular Ca2+ channel involved in cellular Ca2+ signaling. The IP3-binding characteristics of the various IP3R isoforms differ, but until now no specific activators or inhibitors of IP3 binding have been described. We compared the effects of oxidizing reagents, in particular thimerosal, and of cyclic ADP ribose (cADPR) on IP3 binding to the various IP3R isoforms. We therefore expressed the N-terminal 581 amino acids of the three IP(3)R isoforms as recombinant proteins in the soluble fraction of Escherichia coli (ligand-binding sites [lbs] 1, 2, and 3) as well as the full-length IP3R1 and IP3R3 in Spodoptera frugiperda (Sf9) insect cells. Thimerosal (100 microM) stimulated IP3 binding to lbs-1 (1.4-fold) and lbs-3 (2.5-fold), but had no effect on lbs-2. Thimerosal acted on lbs-1 and lbs-3 by decreasing the Kd for IP3 binding (from 46 +/- 4 nM to 20 +/- 2 nM and from 54 +/- 21 nM to 19 +/- 7 nM for lbs-1 and -3, respectively) without modifying the Bmax. Similarly, IP3 binding to microsomes of Sf9 insect cells overexpressing the full-length IP3R1 was 1.2-fold stimulated by thimerosal. Thimerosal, however, did not affect IP3 binding to Sf9-IP3R3 microsomes, suggesting that in situ thimerosal will only directly affect ligand binding to the type 1 isoform. cADPR (50 microM) stimulated IP3 binding to Sf9 IP3R1 microsomes (1.5-fold), but not to Sf9-IP3R3 microsomes. In addition, cADPR inhibited IP3 binding to lbs-1 and lbs-2 by decreasing the affinity for IP3 1.8- and 2.8-fold, respectively, while IP3 binding to lbs-3 was not affected. These results suggest that a regulatory site for cADPR is present in the ligand-binding domain of IP3R1 and 2, but not of IP3R3. PMID- 11274966 TI - The effect of high molecular phospholipase A2 inhibitors on 3T6 fibroblast proliferation. AB - Recently, we suggested that arachidonic acid and/or its cyclooxygenase pathway metabolites may be involved in regulating 3T6 fibroblast proliferation. In the present study we evaluate the role of high-molecular phospholipase A2 (PLA2) enzymes in the 3T6 fibroblast growth. Our results demonstrate that the cytosolic PLA2 inhibitor, arachidonyl trifluoromethylketone and the cytosolic calcium independent PLA2 (iPLA2) inhibitor, bromoenol lactone, decrease arachidonic acid release and prostaglandin E2 production in 3T6 fibroblast cultures stimulated by fetal calf serum. These effects were correlated with the impairment of 3T6 fibroblast proliferation and DNA synthesis at the S/G2 boundary, which prolongs the S phase. These data suggest a role of iPLA2 in the control of 3T6 fibroblast growth. PMID- 11274967 TI - Chlorpromazine-induced increase in dipalmitoylphosphatidylserine surface area in monolayers at room temperature. AB - The Langmuir technique revealed that the surface area of acidic glycerophospholipids (dipalmitoylphosphatidylserine, -glycerol, and dipalmitoylphosphatidic acid) in monolayers increased dramatically when micromolar concentrations of the antipsychotic drug chlorpromazine (CPZ) were present in the subphase. Monolayers of neutral glycerophospholipids (dipalmitoylphosphatidylcholine and -ethanolamine) did not show such a large effect with CPZ. Compared to CPZ, millimolar concentrations of the monovalent cations Li+, K+, Na+, Rb+, and Cs+ did not appear to influence the dipalmitoylphosphatidylserine monolayer, suggesting that the effect of CPZ, a monovalent cationic amphophile, was due to an interaction with the acyl chains of the lipids. In addition, the effect of CPZ was reduced by 150 mM Na+, suggesting that the sodium cations might screen the negatively charged headgroups from an electrostatic interaction with the positively charged drug molecule. Two CPZ analogs, chlorpromazine sulfoxide and CPZ with 2 carbons in the side chain, were also studied. These observations suggest that part of the biological effects of CPZ, being antipsychotic and/or side effects, may be due to CPZ's action on the acidic glycerophospholipids in nerve cell membranes. PMID- 11274968 TI - Modulation of cytosolic calcium levels of human lymphocytes by yessotoxin, a novel marine phycotoxin. AB - Yessotoxin (YTX) is a polyether toxin of marine origin that has been classified among the diarrheic shellfish poisoning (DSP) toxins group due to its lipophilic nature. However, unlike other DSP toxins, YTX does not produce diarrhea and its mechanisms of action are unknown. We studied the effect of YTX on the cytosolic calcium levels of freshly isolated human lymphocytes by means of fluorescence imaging microscopy. We showed that YTX produced a calcium influx through nifedipine and SKF 96365 (1-[beta-[3-(4-methoxyphenyl)propoxyl]-4-methoxyphenyl] 1H-imidazole hydrochloride)-sensitive channels. This Ca2+ entry was not affected by the DSP toxin okadaic acid, which inhibits protein phosphatases. In addition, YTX also produced an inhibition of capacitative calcium entry activated by thapsigargin or by preincubation in a Ca2+-free medium. This capacitative calcium entry was not sensitive to nifedipine. Furthermore, the inhibitory effect of YTX was dependent on the time of addition of the toxin. We suggest that YTX may interact with calcium channels in a way similar to that described for other polyether marine compounds such as brevetoxins and maitotoxin, although an involvement of other second messengers is also likely. PMID- 11274969 TI - Potency and selectivity of RXP407 on human, rat, and mouse angiotensin-converting enzyme. AB - By screening phosphinic peptide libraries, we recently reported the discovery of RXP407 (Ac-Asp-PheY(PO2-CH2)LAla-Ala-NH2), a potent N-domain-selective inhibitor of recombinant human angiotensin-converting enzyme (ACE). Preliminary studies to evaluate the in vivo activity of RXP407 in rat led us to suspect possible differences in the binding property of RXP407 between human and rat ACE. The aim of the present study was thus to determine the potency of RXP407 toward rat and mouse ACEs, as compared to non-recombinant human ACE, and to assess the efficacy of this inhibitor in discriminating between the N- and C-domains of these ACE enzymes. By comparing the ability of RXP407 to block purified somatic and germinal ACE from mice, RXP407 was shown to be a potent N-domain-selective inhibitor of mouse somatic ACE, a behavior similar to that observed with human somatic ACE. In contrast, RXP407 appeared less potent toward purified ACE from rat and furthermore was unable to block ACE activity present in crude rat plasma. This study demonstrated that for further evaluation of the in vivo efficacy of RXP407, mice rather than rats should be used as the animal model. Thus, following the change in the Ac-S-D-K-P plasmatic levels, after i.v. injection of RXP407 to mice, will permit the potency and selectivity of this novel ACE inhibitor to be assessed. PMID- 11274970 TI - Role of cytochrome P450 1A2 in bilirubin degradation Studies in Cyp1a2 (-/-) mutant mice. AB - In congenital jaundice, which is due to defects of bilirubin gluruconidation, bilirubin is degraded by an alternative pathway into unidentified products. Previously, it was shown that plasma bilirubin levels can be decreased in rats with this defect by inducers of CYP1A enzymes. Here, liver microsomes from rats or mice treated with beta-naphthoflavone (BNF) or 3-methylcholanthrene (3 MC) had increased activity for bilirubin degradation. The activity was further stimulated by addition of the coplanar molecule 3,4,3',4'-tetrachlorobiphenyl (TCB). There was more stimulation of bilirubin degradation by TCB in microsomes from BNF treated rats than in microsomes from BNF-treated mice. CYP1A1 to CYP1A2 ratios were greater in rats treated with BNF. In Cyp1a2 (-/-) mutant mice, 3-MC treatment did not increase the rate of bilirubin degradation, but TCB increased this degradation severalfold. Between SWR and C57BL/6 inbred mouse strains that have a 2-fold difference in hepatic constitutive CYP1A2 levels, there was also a 2-fold difference in bilirubin degradation; TCB did not stimulate in either strain. We conclude that CYP1A2 is responsible for microsomal bilirubin degradation in the absence of TCB. TCB was required for bilirubin degradation by CYP1A1. Manipulation of CYP1A2 may be of therapeutic benefit in patients with these diseases of bilirubin conjugation. PMID- 11274971 TI - Potentiation of okadaic acid-induced ceramide elevation but not apoptosis by inhibition of glucosylceramide synthase in human neuroepithelioma cells. AB - Caspase-dependent apoptosis induced by okadaic acid (OA) in CHP-100 neuroepithelioma cells has previously been shown to associate with a rapid and sustained elevation in intracellular ceramide concentration. We now report that treatment of CHP-100 cells with OA also evoked a rapid elevation in glucosylceramide levels that was maintained at steady state as cells underwent apoptosis; moreover, as observed for ceramide, OA-induced glucosylceramide accumulation was not blocked by fumonisin B1. Remarkably, when cell death was prevented by caspase inhibition, glucosylceramide accumulation was potentiated and ceramide elevation reduced, thus suggesting that, during apoptosis completion, accumulation of ceramide was partly driven by impairment of its glucosylation through a caspase-dependent mechanism. We studied whether ceramide glucosylation provided a mechanism for negative modulation of OA-induced apoptosis. We observed that the blocking of glucosylceramide synthesis markedly potentiated OA-induced ceramide elevation, but neither accelerated apoptosis onset nor potentiated the apoptotic response. These results indicate that modulation of ceramide glucosylation does not affect the apoptotic response to okadaic acid and suggest that caution must be exercised concerning the possibility that ceramide plays a key role in apoptosis induction. PMID- 11274972 TI - Marked suppression of the activity of some, but not all, antifolate compounds by augmentation of folate cofactor pools within tumor cells. AB - Folates have been co-administered with some antifolates to diminish host toxicity; however, the extent to which this will reduce antitumor activity is not known. To further clarify this issue, studies were undertaken to characterize and quantitate the impact of alterations in intracellular folate levels on the activities of a variety of antifolates in L1210 murine leukemia cells. Intracellular folate cofactor levels increased almost in proportion to the increase in extracellular 5-formyltetrahydrofolate (5-CHO-THF) over a concentration range that encompassed physiological levels of 5 methyltetrahydrofolate. This resulted in a spectrum of increases in the ic50 values of antifolates upon continuous exposure to drugs [Lometrexol (DDATHF) (70x) > trimetrexate (TMQ) (30x), multitargeted antifolate, LY231514 (ALIMTA) (30x) > Raltitrexed, Tomudex (ZD1694) (10x), 6R-2',5'-thienyl-5,10 dideazatetrahydrofolic acid (LY309887) (10x) > methotrexate (MTX) (6x) > (2S)-2 [o-fluoro-p-[N-(2,7-dimethyl-4-oxo-3,4-dihydroquinazolin-6-ylmethyl)-N-(prop-2 ynyl)amino]benzamido]-4-(tetrazol-5-yl) butyric acid (ZD9331) (3x), N(alpha)-(4 amino-4-deoxypteroyl)-N(delta)-hemiphthaloyl-l-ornithine (PT523) (3x)]. Upon a 4 hr pulse exposure to drug, the ic50 values for DDATHF and ALIMTA were increased > 180- and 5-fold, respectively, with only a 2.5-fold increase in the extracellular 5-CHO-THF level within the physiological range. The reductions in drug sensitivities could be attributed to decreases in accumulation of polyglutamate derivatives of ALIMTA and DDATHF. Hence, in these studies, natural folates diminished the activity of agents that undergo polyglutamation by suppression of the formation of these active congeners at the level of folylpolyglutamate synthetase. For inhibitors of dihydrofolate reductase, the suppressive effect of endogenous folates appears to be due to competition between the antifolate and dihydrofolate at the level of the target enzyme. These data should be carefully considered in the design of regimens with antifolates, which incorporate co administration of folates. PMID- 11274973 TI - Cytotoxic synergism of methioninase in combination with 5-fluorouracil and folinic acid. AB - Potentiation of the cytotoxic activity of 5-fluorouracil (FUra) by folinic acid (5-HCO-H4folate) is due to elevation of the methylene tetrahydrofolate (CH2 H4folate) level, which increases the stability of the ternary complex of thymidylate synthase (TS), fluorodeoxyuridine monophosphate, and CH2-H4folate that inactivates the TS. Methionine deprivation results in the production of tetrahydrofolate (H4folate) and, subsequently, CH2-H4folate from methyl tetrahydrofolate, as a consequence of the induction of methionine synthesis. We hypothesized that the efficacy of FUra could be augmented by the combination of high-concentration 5-HCO-H4folate and recombinant methioninase (rMETase), a methionine-cleaving enzyme. Studies in vitro were performed with the cell line CCRF-CEM. Cytotoxic synergism of FUra + rMETase and FUra + 5-HCO-H4folate + rMETase was demonstrated with the combination index throughout a broad concentration range of FUra and rMETase. A subcytotoxic concentration of rMETase reduced the IC50 of FUra by a factor of 3.6, and by a factor of 7.5, in the absence and in the presence of 5-HCO-H4folate, respectively. 5-HCO-H4folate increased the intracellular concentrations of CH2-H4folate and H4folate from their baseline levels. Concentrations of folates were not changed by exposure to rMETase. Levels of free TS in cells treated with FUra + 5-HCO-H4folate and with FUra + rMETase were lower than those in cells exposed to FUra alone. The decrease of TS was still more pronounced in cells treated with FUra + 5-HCO-H4folate + rMETase. The synergism described in this study will be a basis for further exploration of combinations of fluoropyrimidines, folates, and rMETase. PMID- 11274974 TI - Vinpocetine-induced stimulation of calcium-activated potassium currents in rat pituitary GH3 cells. AB - The effects of vinpocetine, an inhibitor of cyclic GMP phosphodiesterase, on ionic currents were examined in rat pituitary GH3 lactotrophs with the aid of the patch-clamp technique. In GH3 cells bathed in normal Tyrode's solution, vinpocetine (10 microM) reversibly increased the amplitude of Ca2+-activated K+ current (I(K)Ca) with an EC50 value of 4 microM. When the recording pipettes were filled with 10 mM EGTA, vinpocetine also stimulated I(K)Ca. In the cell-attached configuration, application of vinpocetine to the bath increased the activity of large-conductance Ca2+-activated K+ (BK(Ca)) channels. In excised membrane patches, application of vinpocetine (10 microM) to the bath did not change the single-channel conductance of BK(Ca) channels; however, it did increase channel activity. In the inside-out configuration, neither 8-bromo cyclic GMP nor YC-1 applied intracellularly affected BK(Ca) channel activity. The vinpocetine-induced change in the kinetic behavior of BK(Ca) channels was due to an increase in mean open time and a decrease in mean closed time. Vinpocetine (10 microM) caused a leftward shift in the midpoint for the voltage-dependent opening. Under the current-clamp mode, vinpocetine (10 microM) decreased the firing rate of spontaneous action potentials induced by thyrotropin-releasing hormone (10 microM) in GH3 cells. In pheochromocytoma PC12 cells, vinpocetine (10 microM) applied intracellularly also enhanced the activity of BK(Ca) channels without altering single-channel conductance. Thus, the present study suggests that vinpocetine-mediated stimulation of I(K)Ca may result from the direct activation of BK(Ca) channels and indirectly from elevated cytosolic Ca2+. PMID- 11274975 TI - Metabolism of a 20-methyl substituted series of vitamin D analogs by cultured human cells: apparent reduction of 23-hydroxylation of the side chain by the 20 methyl group. AB - We describe here for the first time the effect of introducing a 20-methyl group on the side-chain metabolism of the vitamin D molecule. Using a series of 20 methyl-derivatives of 1alpha,25-(OH)2D3 incubated with two different cultured human cell lines, HPK1A-ras and HepG2, previously shown to metabolize vitamin D compounds, we obtained a series of metabolic products that were identified by comparison to chemically synthesized standards on HPLC and GC-MS. 24-Hydroxylated , 24-oxo-hydroxylated-, and 24-oxo-23-hydroxylated products of 20-methyl 1alpha,25-(OH)2D3 were observed, but the efficiency of 23-hydroxylation was low as compared with that of the natural hormone and, in contrast to 1alpha,25 (OH)2D3, no truncated 23-alcohol was formed from the 20-methyl analog. These data, taken together with results from other analogs with changes in the vicinity of the C17-C20 positions, lead us to speculate that such changes must alter the accessibility of the C-23 position to the cytochrome P450 involved. Using the HepG2 cell line, we found evidence that the 24S-hydroxylated product of 20-methyl 1alpha,25-(OH)2D3 predominates, implying that the liver cytochrome involved in metabolism is a different isoform. Studies with a more metabolically resistant analog of the series, 20-methyl-Delta(23)-1alpha,25-(OH)2D3, gave the expected block in 23- and 24-hydroxylation, and evidence of an alternative pathway, namely 26-hydroxylation. 20-Methyl-Delta(23)-1alpha,25-(OH)2D3 was also more potent in biological assays, and the metabolic studies reported here help us to suggest explanations for this increased potency. We conclude that the 20-methyl series of vitamin D analogs offers new perspectives into vitamin D analog action, as well as insights into the substrate preferences of the cytochrome(s) P450 involved in vitamin D catabolism. PMID- 11274976 TI - Suppression by a sesquiterpene lactone from Carpesium divaricatum of inducible nitric oxide synthase by inhibiting nuclear factor-kappaB activation. AB - Excessive nitric oxide (NO) produced by inducible NO synthase (iNOS) acts as a causative regulator in various inflammatory disease states. Carpesium divaricatum has been used in Korean traditional herbal medicine for its antipyretic, analgesic, vermifugic, and anti-inflammatory properties. We investigated the molecular mechanism for the suppression of lipopolysaccharide/interferon-gamma (LPS/IFN-gamma)-induced NO production in RAW 264.7 macrophages by the sesquiterpene lactone 2beta,5-epoxy-5,10-dihydroxy-6alpha-angeloyloxy-9beta isobutyloxy-germacran-8alpha,12-olide (C-1), which has been identified recently as a new compound from C. divaricatum. C-1 decreased NO production in LPS/IFN gamma-stimulated RAW 264.7 cells in a concentration-dependent manner, with an IC50 of approximately 2.16 microM; however, it had no direct effect on the iNOS activity of fully LPS/IFN-gamma-stimulated RAW 264.7 cells. Furthermore, treatment with C-1 led to a decrease in iNOS protein and mRNA. These effects appear to be due to inhibition of nuclear factor-kappaB (NF-kappaB) activation through a mechanism involving stabilization of the NF-kappaB/inhibitor of the kappaB (I-kappaB) complex, since inhibition of NF-kappaB DNA binding activity by C-1 was accompanied by a parallel reduction of nuclear translocation of subunit p65 of NF-kappaB and I-kappaBalpha degradation. Taken together, the results suggest that the ability of C-1 to inhibit iNOS gene expression may be responsible, in part, for its anti-inflammatory effects. PMID- 11274977 TI - Evidence for bradykinin mediation of carrageenin-induced inflammatory pain: a study using kininogen-deficient Brown Norway Katholiek rats. AB - Inflammatory pain was induced following an intradermal injection of carrageenin into rat paws, and the hyperalgesia was measured in terms of withdrawal time following thermal pain stimulation of the inflamed paw. This hyperalgesia was significantly less in kininogen-deficient Brown Norway (B/N)-Katholiek rats, which also showed less swelling in carrageenin-induced paw edema, than in normal B/N-Kitasato rats at 1 approximately 4 hr after the carrageenin injection (at the early phase). However, 24 hr after the injection, hyperalgesia and the swelling volume of the kininogen-deficient rats were almost the same as those in normal rats. The bradykinin B2 receptor antagonist FR173657, (E)-3-(6-acetamido-3 pyridyl)-N-[N-[2,4-dichloro-3-[(2-methyl-8-quinolinyl)oxymethyl]phenyl]-N methylaminocarbonylmethyl]acrylamide, attenuated the carrageenin-induced swelling and hyperalgesia of the normal rats at the early phase to almost the levels of the B/N-Katholiek rats. Pretreatment with indomethacin, a cyclooxygenase inhibitor, also inhibited the carrageenin-induced responses significantly in normal rats. These results indicate that bradykinin, acting on the B2 receptor, is the main mediator at the early phase of inflammatory pain of carrageenin edema and that prostaglandins, produced by cyclooxygenase, potentiate the effects of bradykinin. PMID- 11274978 TI - Proteolytic activation of membrane-bound guanylate cyclase. AB - Membrane-bound guanylate cyclase-A (GC-A), the receptor for atrial natriuretic factor (ANF), has been shown to be regulated by its kinase-like domain. To resolve the nature of this regulation, we measured the effects of various proteases on the activity of guanylate cyclase in rat lung membranes, and on the activity of the bacterial-expressed catalytic domain (GC-c) and on a recombinant peptide composed of both the kinase-like and catalytic domain (GC-kc) of guanylate cyclase. Pronase increased rat guanylate cyclase activity in a biphasic manner with a maximal effect at about 10-20 microg per assay tube. Thermolysin had effects similar to those of pronase on the activity of guanylate cyclase in rat lung membranes. In the case of bacterial-expressed proteins, pronase increased the activity of GC-kc, but not GC-c. These results indicate that GC-A contains an autoinhibitory site on its kinase-like domain, and that removal of the autoinhibitory site by limited proteolysis leads to enzyme activation. GC-A was poorly activated by ANF and ATP after the rat lung membrane was pretreated with pronase, suggesting that ANF/ATP and pronase activate guanylate cyclase through the same mechanism. It is suggested that the binding of ANF and ATP to GC A may induce a conformational change of the receptor that releases the inhibitory constraint on enzyme activity leading to enzyme activation. PMID- 11274979 TI - Possible involvement of G(i3) protein in augmented contraction of bronchial smooth muscle from antigen-induced airway hyperresponsive rats. AB - To investigate a possible involvement of pertussis toxin (PTX)-sensitive heterotrimeric G proteins in the pathogenesis of airway hyperresponsiveness, the effect of PTX treatment on the augmented contractile response to acetylcholine (ACh) in bronchial smooth muscle of antigen-induced airway hyperresponsive rats was determined. In bronchial smooth muscle of airway hyperresponsive rats that were actively sensitized and repeatedly challenged with 2,4-dinitrophenylated Ascaris suum antigen, ACh-induced contractions were markedly augmented. The augmented contractile responses in the airway hyperresponsive group were significantly inhibited after treatment with PTX (1 microg/mL for 6 hr, 37 degrees ), whereas only a slight attenuation was observed in the normal control group. The level of G(alpha)i3 (measured by immunoblotting), but not other alpha subunits of G(i/o) family proteins, in bronchial smooth muscle of the airway hyperresponsive rats was significantly increased as compared with that of control animals. It is concluded that PTX-sensitive muscarinic contractile responses of bronchial smooth muscle might be augmented upon antigen-induced airway hyperresponsiveness in rats, probably due to an up-regulation of G(alpha)i3 protein of bronchial smooth muscle. PMID- 11274981 TI - Apoptosis. PMID- 11274980 TI - Effects of 2(RS)-n-propylthiazolidine-4(R)-carboxylic acid on extrahepatic sulfhydryl levels in mice treated with acetaminophen. AB - The cysteine (Cys) precursor 2(RS)-n-propylthiazolidine-4(R)-carboxylic acid (PTCA) has been shown to protect against acetaminophen (APAP)-induced hepatic GSH, GSSG, and Cys depletion and hepatic necrosis. The aim of this study was to determine the effects of PTCA on the concentrations of sulfhydryl compounds in extrahepatic tissues, including renal cortex, whole blood, and brain, in C57BL/6 mice treated with hepatotoxic doses of APAP. PTCA (1-5 mmol/kg, i.p.) was administered 30 min after the administration of APAP at a dose (800 mg/kg; 5.29 mmol/kg, i.p.) that depleted hepatic GSH and Cys at 4 hr by 95 and 86%, respectively. Tissue concentrations of GSH and Cys were determined by HPLC. At 4 hr following APAP administration, renal cortical GSH and Cys concentrations were decreased to 64 and 39%, respectively, of vehicle-treated control values, and blood concentrations were decreased to 87 and 30%, respectively, of vehicle controls. Brain GSH and Cys were not depleted by APAP. PTCA at 5 mmol/kg (i) attenuated the APAP-induced depletion of GSH and Cys at 4 hr in renal cortex (78 and 65%, respectively, of vehicle controls), (ii) prevented APAP-induced Cys depletion in blood (670% of vehicle controls) with no effect on GSH concentration (94% of vehicle controls), and (iii) increased GSH and Cys concentrations in brain (119 and 411%, respectively, of vehicle controls). The results demonstrate a high degree of tissue selectivity in the APAP-induced depletion of GSH and Cys, and in the effectiveness of PTCA in maintaining and even elevating sulfhydryl levels in extrahepatic tissues of APAP-treated mice. PMID- 11274983 TI - Unconscious semantic priming extends to novel unseen stimuli. AB - Many subliminal priming experiments are thought to demonstrate unconscious access to semantics. However, most of them can be reinterpreted in a non-semantic framework that supposes only that subjects learn to map non-semantic visual features of the subliminal stimuli onto motor responses. In order to clarify this issue, we engaged subjects in a number comparison task in which the target number was preceded by another invisible masked number. We show that unconscious semantic priming occurs even for prime stimuli that are never presented as target stimuli, and for which no stimulus-response learning could conceivably occur. We also report analyses of the impact of the numerical relation between prime and target, and of the impact of learning on priming, all of which confirm that unconscious utilization of semantic information is indeed possible. PMID- 11274982 TI - Set size and repetition in the picture--word interference paradigm: implications for models of naming. AB - Caramazza and Costa (Cognition 75 (2000) B51) reported results which demonstrate that a semantically related word distractor interferes in picture naming even when it is not in the response set and there is no possibility for mediated interference. They interpreted the results to be problematic for the model of lexical access proposed by Levelt, Roelofs, and Meyer (Behavioral and Brain Sciences 22 (1999) 1). Roelofs (Cognition 80 (2001, this issue 283--90)) argues that those results are not inconsistent with Levelt et al.'s model when certain new assumptions about the mechanism of lexical selection are considered. Here we show that even with these assumptions the model still makes the wrong predictions. We report new results which demonstrate that the semantic interference and facilitation effects that are obtained respectively in the basic level and category-level naming variants of the picture-word interference paradigm are not the result of response set size and response repetitions. PMID- 11274984 TI - The effect of oral vocabulary on reading visually novel words: a comparison of the dual-route-cascaded and triangle frameworks. AB - Dual-route-cascaded (DRC) (e.g. Coltheart & Rastle, Journal of Experimental Psychology: Human Perception and Performance 20 (1994) 1197) and triangle framework (e.g. Seidenberg & McClelland, Psychological Review 96 (1989) 523) predictions were tested regarding the effect of having a word in oral vocabulary prior to reading that same word. Over two sessions, at intervals of 2--3 days, 44 Grade 1 (6--7-year-old) children were aurally familiarized with the sound and meaning of ten novel words (semantic oral instantiation), and with just the sound of another ten novel words (non-semantic oral instantiation). Two to three days later non-word naming performance was significantly more accurate for aurally trained novel words compared to pseudohomophones, which were in turn advantaged over untrained non-words. The semantic manipulation had no effect. Experiment 2 manipulated articulation during (non-semantic) training. Forty Grade 1 children participated. Again, aurally trained items were named more accurately and quickly than equivalently trained pseudohomophones, which were in turn advantaged over untrained non-words. The articulation manipulation had no effect. The results suggest that word-specific phonological information is represented in the reading system independently of semantic or articulatory influences. The results are interpreted as being problematic for both the DRC and triangle frameworks, but more so for the latter. PMID- 11274985 TI - Indexical and symbolic referencing: what role do they play in children's success on theory of mind tasks? AB - Numerous measures have been employed in the last 17 years to assess theory of mind (ToM). The literature reports marked variability in the age at which children succeed on these measures. To account for this variability, researchers have provided explanations ranging from cognitive shifts and voids to the inability to understand the language of the tasks or to social/pragmatic considerations, all of which tell us little if anything about the internal mechanism underlying ToM. The main purpose of this paper is to provide a comprehensive theoretical account of children's success and the discrepancies found across different ToM tasks. We test the hypothesis that children's understanding of ToM is sensitive to the basic elements of language, that is, to whether the language is indexical or symbolic. Support for this account was found in the analysis of selected test protocols in four published studies of ToM, and new data collected from 53 children (4--6 years) which showed that a higher percentage of children succeeded on tasks with a high ratio of indexical to symbolic references than on tasks with a high ratio of symbolic to indexical references. There was also a main effect of age with older children succeeding at higher rates on both tasks than younger children. Our findings suggest that indexical representation can afford ToM understanding in 4-year-olds, but is not sufficient for a more mature ToM. The latter requires symbolic representation that was demonstrated by the majority of 5--6-year-olds. PMID- 11274986 TI - A developmental study of the affective value of tempo and mode in music. AB - Do children use the same properties as adults in determining whether music sounds happy or sad? We addressed this question with a set of 32 excerpts (16 happy and 16 sad) taken from pre-existing music. The tempo (i.e. the number of beats per minute) and the mode (i.e. the specific subset of pitches used to write a given musical excerpt) of these excerpts were modified independently and jointly in order to measure their effects on happy-sad judgments. Adults and children from 3 to 8 years old were required to judge whether the excerpts were happy or sad. The results show that as adults, 6--8-year-old children are affected by mode and tempo manipulations. In contrast, 5-year-olds' responses are only affected by a change of tempo. The youngest children (3--4-year-olds) failed to distinguish the happy from the sad tone of the music above chance. The results indicate that tempo is mastered earlier than mode to infer the emotional tone conveyed by music. PMID- 11274987 TI - Linguistic and cognitive abilities in infancy: when does language become a tool for categorization? AB - Infants' ability to form new object categories based on either visual or naming information alone was evaluated at two different ages (16 and 20 months) using an object manipulation task. Estimates of productive vocabulary size were also collected. Infants at both ages showed evidence of using visual information to categorize the objects, while only the older ones used naming information. Moreover, there was a correlation between vocabulary size and name-based categorization among the 20-month-olds. The present results establish that infants as young as 20 months can use the non-obvious cue of naming to categorize objects. The possibility of a link between this ability and lexical development is discussed. PMID- 11274988 TI - Strategy choice for arithmetic verification: effects of numerical surface form. AB - Canadian university students (n=48) solved simple addition problems in a true/false verification task with equations in digit format (3+4=8) or written English format (three+four=eight). Participants reported their solution strategy (e.g. retrieval or calculation) after each trial. Reported use of calculation strategies was much greater with word (41%) than digit stimuli (26%), and this difference was exaggerated for numerically larger problems. Word-format costs on reaction time (RT) were correspondingly greater for large than for small problems, but this Format x Size RT effect was bigger for true than for false equations. The results demonstrate that surface format affects central, rather than only peripheral, stages of cognitive arithmetic. PMID- 11274989 TI - Imidazoline I(1) receptor-induced activation of phosphatidylcholine-specific phospholipase C elicits mitogen-activated protein kinase phosphorylation in PC12 cells. AB - In the present study, we tested the hypothesis that the activation of imidazoline I(1)-receptor, which is coupled to phosphatidylcholine-specific phospholipase C, results in downstream activation of mitogen-activated protein kinase (p42(mapk) and p44(mapk) isoforms) in PC12 cells. PC12 cells pretreated with nerve growth factor (50 ng/ml, 48 h) to initiate neuronal differentiation were incubated with [methyl-3H]choline and [3H]myristate. Activation of imidazoline I(1) receptor by rilmenidine (10 microM) caused time-dependent increases in diacylglycerol accumulation and phosphocholine release. The Western blotting analysis showed that rilmenidine (10 microM) produced a time-dependent activation of p42(mapk) and p44(mapk) that reached its maximum at 15 min and returned to control levels after 30 min. This finding was confirmed by immunofluorescence labeling of activated mitogen-activated protein kinase in the same model system. Efaroxan (imidazoline I(1)-receptor antagonist) or tricyclodecan-9-yl-xanthogenate (D609, phosphatidylcholine-specific phospholipase C inhibitor) attenuated the phosphorylation of p42(mapk) and p44(mapk) induced by rilmenidine. Nerve growth factor-induced phosphorylation of both mitogen-activated protein kinase isoforms was not affected by D609. These results support the hypothesis that the activation of the imidazoline I(1) receptor coupled phosphatidylcholine-specific phospholipase C results in the downstream activation of mitogen-activated protein kinase. PMID- 11274991 TI - Interaction of the neuroprotective drug riluzole with GABA(A) and glycine receptor channels. AB - Riluzole is used as therapeutic agent in amyotrophic lateral sclerosis. We investigated the interaction of riluzole with recombinant GABA (gamma aminobutyric acid)(A) receptor channels (alpha(1)beta(2)gamma(2)-subunits) and glycine receptor channels (alpha(1)beta-subunits) transiently expressed in HEK293 cells. For electrophysiological experiments, the patch-clamp technique in combination with tools for ultrafast solution exchange was used. Saturating concentrations of GABA or glycine were applied with different concentrations of riluzole to outside-out patches containing alpha(1)beta(2)gamma(2) GABA(A) receptor channels or alpha(1)beta-glycine receptor channels on their surface, respectively. The current declined after application of GABA or glycine with three time constants of desensitization to a steady-state current amplitude. Application of riluzole resulted in a shift to fast desensitized states at both receptors. The proportion of the time constants of fast desensitization increased and the time constants of slow desensitization and the steady-state current decreased whereas the maximal current amplitudes were not affected by riluzole. The data of the study demonstrate for the first time interaction of GABAergic and glycinergic currents with riluzole under physiological conditions. PMID- 11274990 TI - Pharmacological characterisation of pyrimidinoceptor responses in NG108-15 cells. AB - In the present study, the P2Y receptor(s) mediating the effects of the pyrimidines UTP and UDP on phospholipase C activation in the mouse neuroblastoma x rat glioma hybrid cell line NG108-15 was investigated. Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) analysis detected transcripts for the P2Y(6) and P2Y(2) receptors, but not for P2Y(1) and P2Y(4.) UTP and UDP were equipotent agonists and their effects were partially additive. Suramin, reactive blue 2 and pyridoxal phosphate-6-azophenyl-2',4'disulfonic acid (PPADS) antagonised the phospholipase C response to both UTP and UDP. High micromolar concentrations of adenosine, 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine (CGS 21680), 2',3'-O-isopropylideneadenosine (iPAdo) and adenosine 3':5'-cyclic monophosphate (3',5'-cAMP) were able to antagonise the effect of UTP on phospholipase C but not that of UDP. The additivity of the UTP and UDP responses, novel P2 receptor antagonist profile and the distinguishing action of adenosine may indicate the expression of a pyrimidine selective P2Y receptor in addition to the P2Y(6) type in these cells. PMID- 11274992 TI - Effects of ethosuximide, a T-type Ca(2+) channel blocker, on dorsal horn neuronal responses in rats. AB - Plasticity in transmission and modulatory systems are implicated in mechanisms of neuropathic pain. Studies demonstrate the importance of high voltage-activated Ca(2+) channels in pain transmission, but the role of low voltage-activated, T type Ca(2+) channels in nociception has not been investigated. The Kim and Chung rodent model of neuropathy [Pain 50 (1992) 355] was used to induce mechanical and cold allodynia in the ipsilateral hindpaw. In vivo electrophysiological techniques were used to record the response of dorsal horn neurones to innocuous and noxious electrical and natural (mechanical and thermal) stimuli after spinal nerve ligation. Spinal ethosuximide (5-1055 microg) exerted dose-related inhibitions of both the electrically and low- and high-intensity mechanical and thermal evoked neuronal responses and its profile remained unaltered after neuropathy. Measures of spinal cord hyperexcitability were most susceptible to ethosuximide. This study, for the first time, indicates a possible role for low voltage-activated Ca(2+) channels in sensory transmission. PMID- 11274993 TI - Characterisation of P2Y(1)-like receptor in cultured rat pineal glands. AB - The rat pineal gland possesses P2 receptors which potentiate the effect of noradrenaline-induced N'-acetyl-5-hydroxytryptamine (N'-acetyl-5-HT) production. In the current study, this receptor was characterised according to agonist selectivity and signal transduction mechanisms. 2-MethylthioATP (2MeSATP), 2 chloroATP (2-ClATP), adenosine 5'-O-2-thiodiphosphate, (ADPbetaS), ATP and ADP, but not UTP, potentiated noradrenaline-induced N'-acetyl-5-HT production in a concentration-dependent manner. 2MeSATP neither induced the production of adenosine 3':5'-cyclic monophosphate (cyclic AMP), nor inhibited its formation when the glands were stimulated by forskolin. The phospholipase C inhibitor 1-[6 [[(17beta)-3-Methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione (U73122), but not the inactive analogue, 1-[6-[[(17beta)-3-Methoxyestra-1,3,5(10) trien-17-yl]amino]hexyl]-2,5-pyrrolidinedione (U73343), blocked the 2MeSATP effect. The P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2',4' dissulphonic acid (PPADS), which inhibits phospholipase C-coupled P2Y(1) receptors, blocked the 2MeSATP effect. In conclusion, our data strongly suggest that the P2-like receptor that is present in rat pinealocytes and which is responsible for the potentiation of noradrenaline-induced N'-acetyl-5-HT production is a P2Y(1)-like receptor, coupled to a G protein which stimulates phospholipase C. PMID- 11274994 TI - Central and peripheral activity of cholinesterase inhibitors as revealed by yawning and fasciculation in rats. AB - This study was designed to investigate the central and peripheral activity profile of cholinesterase inhibitors in rats. Intravenous injection of cholinesterase inhibitors caused fasciculation, a fine involuntary muscular movement. This peripheral cholinergic sign was tightly correlated with in vitro anti-acetylcholinesterase activity by cholinesterase inhibitors, suggesting that fasciculation is a valid index of peripheral cholinergic activation. Yawning, used as a marker of central cholinergic activation, was also monitored. E2030 (3 (2-(1-(1,3-dioxolan-2-ylmethyl)-4-piperidyl)ethyl)-2H-3,4-dihydro-1,3-benzoxazin 2,4-dione hydrochloride) elicited yawning at more than 4 mg/kg, while fasciculation was significantly intensified only at a dose of 16 mg/kg. Donepezil and tacrine induced both yawning and fasciculation at doses greater than 4 mg/kg, whereas physostigmine induced both behaviors at a dose of 8 mg/kg and above. Finally, ipidacrine elicited yawning at a dose of 16 mg/kg and fasciculation at doses greater than 8 mg/kg. Thus, all putative centrally acting cholinesterase inhibitors elicited yawning. TAK-147 (3-[1-(phenylmethyl)-4-piperidinyl]-1 (2,3,4,5-tetrahydro-1H-benzazepin-8-yl)-1-propanone fumarate) did not significantly elicit yawning at doses under 16 mg/kg, but elicited fasciculation at a dose of more than 4 mg/kg. Distigmine, a peripherally acting cholinesterase inhibitor, evoked fasciculations, but not yawning. When mild to moderate fasciculation was evoked, donepezil and E2030 elicited more than nine yawns over 30 min, while the other cholinesterase inhibitors elicited approximately five yawns at most during this period. These results indicated that E2030 and donepezil exhibited the most marked preferential central cholinergic activity, relative to peripheral activity, among cholinesterase inhibitors tested. Scopolamine, a centrally acting antimuscarinic drug, completely inhibited E2030 induced yawning, while peripherally acting methylscopolamine did not. Haloperidol, a dopamine receptor antagonist, partially blocked E2030-induced yawning, but did not block donepezil-induced yawning. These results suggest that central cholinergic and, in part, dopaminergic mechanisms are involved in E2030 induced yawning. PMID- 11274995 TI - Ultrasonic vocalizations of preweanling rats: involvement of both alpha(2) adrenoceptor and kappa-opioid receptor systems. AB - Stimulation of alpha(2)-adrenoceptors and kappa-opioid receptors increases the ultrasonic vocalizations of preweanling rats. The purpose of the present study was to determine whether alpha(2)-adrenoceptors and kappa-opioid receptors modulate ultrasonic vocalization production via a common mechanism. To that end, 11-day-old rats were injected with the alpha(2)-adrenoceptor antagonist yohimbine (0, 0.5, or 1.0 mg/kg, i.p.) or the kappa-opioid receptor antagonist nor binaltorphimine (0, 5, or 10 mg/kg, i.p.). After 15 min, the same rats were injected with saline, the alpha(2)-adrenoceptor agonist clonidine (0.25 mg/kg, i.p.), or the kappa-opioid receptor agonist trans-(+/-)-3,4-dichloro-N-methyl-N [2-(1-pyrrolidinyl)-cyclohexyl]-benzeneacetamide methanesulfonate (U-50,488; 2.5 mg/kg, i.p.). Results showed that both clonidine and U-50,488 increased the ultrasonic vocalizations of preweanling rats. Not surprisingly, clonidine-induced ultrasonic vocalizations were blocked by yohimbine, while U-50,488-induced vocalizations were blocked by nor-binaltorphimine. Importantly, yohimbine also attenuated the vocalizations produced by U-50,488, whereas nor-binaltorphimine did not alter clonidine-induced ultrasonic vocalizations. Thus, it appears that alpha(2)-adrenoceptor and kappa-opioid receptor stimulation increases ultrasonic vocalization production via a common mechanism. It is likely that the kappa opioid receptors responsible for modulating ultrasonic vocalizations are located "upstream" from the alpha(2)-adrenoceptors. PMID- 11274996 TI - Failure of GPI compounds to display neurotrophic activity in vitro and in vivo. AB - The aim of this study was to evaluate the neurotrophic and neuroprotective properties of a series of immunophilin ligands and to assess the potential involvement of FK506 Binding Protein 12 kDa (FKBP12) rotamase inhibition in this activity. Both FK506 and rapamycin induced a potent inhibition of the FKBP12 rotamase activity (pIC(50) values of 7.3 and 7.4, respectively) but only a modest inhibition was observed with 1-(3,3-dimethyl-2-oxo-pentanoyl)-pyrrolidine-2 carboxylic acid S-3-pyridin-3-yl-propyl ester (GPI 1046) (5.8), its N-oxide (5.4) and thioester (6.3) analogues. Compared to nerve growth factor, all these immunophilin ligands only induced marginal increases in neurite outgrowth of rat dissociated newborn dorsal root ganglia cells. Furthermore, systemic administration of GPI 1046 and its N-oxide and thioester analogues failed to prevent striatal dopamine depletion induced by acute or chronic i.p. treatment with 1-methyl-4-phenyl 1,2,3,6 tetrahydropyridine (MPTP). These results suggest that inhibition of FKBP12 rotamase activity is not predictive for neurotrophic and neuroprotective properties of immunophilin ligands and question their therapeutic utility in neurodegenerative diseases like Parkinson's disease. PMID- 11274997 TI - Sedative and anxiolytic effects of zopiclone's enantiomers and metabolite. AB - We evaluated racemic zopiclone, its (S)- and (R)-enantiomers and a metabolite, (S)-desmethylzopiclone, for their actions on locomotor activity, rotarod performance, the elevated plus maze and the Vogel conflict test of anxiety, and electroconvulsive shock-induced seizures duration. Zopiclone and its (R)- and (S) enantiomers reduced locomotor activity, and zopiclone and its (S)-enantiomer disrupted rotarod performance at 10 mg/kg. (S)-desmethylzopiclone did not alter these measures at doses of less than 200 mg/kg. (S)-desmethylzopiclone altered plus maze performance at the lowest dose of all the zopiclone derivatives tested, caused a dose-related effect on the Vogel conflict test and caused a dose-related reduction of electroconvulsive shock-induced seizure durations. The data indicate that (S)-desmethylzopiclone can bring about an anxiolytic effect without a substantial degree of central nervous system depression, and suggest that the agent may be particularly useful clinically in the treatment of anxiety. PMID- 11274998 TI - CGP 36216 is a selective antagonist at GABA(B) presynaptic receptors in rat brain. AB - In rat neocortical preparations maintained in Mg(2+)-free Krebs medium, baclofen depressed the frequency of spontaneous discharges in a concentration-dependent manner (EC(50) = 6 microM), sensitive to (3-aminopropyl)ethylphosphinic acid (CGP 36216) (100, 300 and 500 microM) (pA(2) = 3.9 +/- 0.1). By contrast, CGP 36216, up to 1 mM, was ineffective in antagonising baclofen-induced hyperpolarisations, mediated through gamma-aminobutyric acid(B) (GABA(B)) postsynaptic receptors. In electrically stimulated brain slices preloaded with [3H]GABA, CGP 36216 increased [3H]GABA release (IC(50) = 43 microM), which was reversed by baclofen (20 microM). While CGP 36216 is ineffective at GABA(B) postsynaptic receptors, it is appreciably more active at presynaptic receptors. PMID- 11274999 TI - Group housing of mice increases immobility and antidepressant sensitivity in the forced swim and tail suspension tests. AB - The forced swim test and tail suspension test are often used in laboratory practice to identify compounds that possess antidepressant-like activity. This experiment was conducted to determine whether housing conditions per se influence the response of mice in these antidepressant screening procedures. Male NIH Swiss mice were housed individually or in groups (five per cage) for 8 weeks prior to testing. After 8 weeks, the animals were exposed to the forced swim and tail suspension tests. Group housed mice displayed high levels of immobility in the forced swim and tail suspension tests. Desipramine injection 60 min prior testing, in doses 7.5 and 15 mg/kg, produced significant reductions in the immobility time in forced swimming and tail suspension tests. Individually housed mice, when exposed to these tests, displayed lower levels of immobility with a magnitude comparable to the effect of desipramine in group housed mice. Desipramine given to individually housed mice did not reduce the duration of immobility either in the forced swim test or in the tail suspension test. These results indicate that both tests are sensitive to housing conditions. This observation suggests that long lasting group housing may be critical to the behavioral response in these preclinical screening procedures in mice. PMID- 11275000 TI - Corticotropin-releasing factor receptor type 1 mediates stress-induced relapse to cocaine-conditioned place preference in rats. AB - Corticotropin-releasing factor (CRF) has been suggested to play an important role in the development of drug dependence and withdrawal. Based on the recent finding that CRF receptor antagonists inhibit the stress-induced relapse to opiate dependence and attenuate anxiety-like responses related to cocaine withdrawal, the present experiment was performed to examine the possible effect of different CRF receptor antagonists on reactivation of cocaine-conditioned place preference induced by cocaine and stress in rats. The results show that a single injection of cocaine (10 mg/kg, i.p.) could reactivate cocaine-conditioned place preference following a 28-day extinction, and pretreatment with i.c.v. 10 microg alpha helical CRF, a nonspecific CRF receptor antagonist, significantly attenuated this reactivation of conditioned place preference. However, pretreatment with i.p. 1 or 10 mg/kg CP-154,526 (butyl-[2,5-dimethyl-7-(2,4,6-trimethylphenyl)-7H pyrrolo[2,3-d]pyrimidin-4-yl]-ethylamine), a specific CRF receptor subtype 1 antagonist, and i.c.v. 1 or 10 microg AS-30 ([D-Phe(11),His(12)]Svg-(11-40)), a specific CRF receptor subtype 2 antagonist, failed to show the same effects. In addition, a single footshock stress also elicited the reactivation of cocaine conditioned place preference following a 28-day extinction and pretreatment with alpha-helical CRF (10 microg, i.c.v.) and CP-154,526 (1 or 10 mg/kg, i.p.) significantly blocked this effect. In contrast, pretreatment with AS-30 at a dose of 1 or 10 microg (i.c.v.) did not affect the stress-induced reactivation of cocaine-conditioned place preference. The present study demonstrated that CRF receptor type 1, but not CRF receptor type 2, mediates the stress-induced reactivation of cocaine-conditioned place preference. These findings suggest that CRF receptor subtype 1 antagonists might be of some value in the treatment and prevention of stress-induced relapse to drug dependence long after detoxification. PMID- 11275001 TI - Release of endothelial nitric oxide in coronary arteries by celiprolol, a beta(1) adrenoceptor antagonist: possible clinical relevance. AB - Mechanisms underlying celiprolol-induced vasodilatation were analyzed in isolated porcine coronary arteries. Celiprolol induced dose-related relaxation of the artery rings with endothelium, an effect which was suppressed by N(G)-nitro-L arginine methylester (L-NAME), nitric oxide (NO) scavenger, guanylate cyclase inhibitor, endothelium denudation, and removal of Ca(2+). L-NAME contracted, and superoxide dismutase relaxed, the arteries only when the endothelium was preserved. Neither superoxide dismutase nor beta-adrenoceptor antagonists changed celiprolol-induced relaxations. Celiprolol increased the cyclic GMP content in the tissue. The release of NO from endothelium, estimated by the extracellular production of cyclic GMP in arteries incubated in medium containing guanylate cyclase and GTP, was augmented by celiprolol, and L-NAME abolished this action of celiprolol. It is concluded that celiprolol elicits relaxation by acting on sites other than beta-adrenoceptors in the endothelium and by releasing NO, which activates soluble guanylate cyclase in smooth muscle and produces cyclic GMP. Scavenging of superoxide anions from the endothelium does not seem to account for the induced relaxation. PMID- 11275002 TI - Effect of simvastatin on vascular smooth muscle responsiveness: involvement of Ca(2+) homeostasis. AB - This report is focused on the study of simvastatin-induced relaxation of rat aorta through its effects on vascular smooth muscle and Ca(2+) signalling. The presence of endothelium affected only the simvastatin-induced relaxation of aortic rings precontracted with noradrenaline, but not by depolarization with KCl 80 mM. Blockade of Ca(2+) entry through voltage-operated Ca(2+) channels (VOCCs) by diltiazem abolished the endothelium-dependent and direct relaxation, whereas Ca(2+)-ATPase inhibition by cyclopiazonic acid (3 x 10(-5) M) only affected the endothelium-dependent relaxation. In KCl-depolarised arteries concentration response curves for CaCl(2) were shifted to the right in the presence of simvastatin (3 x 10(-6) and 3 x 10(-5) M) or diltiazem (10(-6) and 10(-7) M). The transient contraction caused by noradrenaline in Ca(2+)-free medium, which is mainly due to intracellular Ca(2+) release, was inhibited by simvastatin (3 x 10( 5) M) or cyclopiazonic acid (3 x 10(-5) M) and the contraction induced by CaCl(2) (2 x 10(-3) M) added after noradrenaline was inhibited by diltiazem and simvastatin. All the reported effects of simvastatin were inhibited by the product of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, mevalonate (10(-3) M). These findings demonstrate that the vascular effects of simvastatin may involve both Ca(2+) release from intracellular stores, which could promote activation of endothelial factors, and blockade of extracellular Ca(2+) entry, which promote relaxations independent of the presence of endothelium. This action on Ca(2+) could be related to the inhibition of isoprenoid synthesis, which subsequently affects the function of G-proteins involved in communication among intracellular Ca(2+) pools and capacitative Ca(2+) entry. PMID- 11275003 TI - Diabetes potentiates acetylcholine-induced relaxation in rabbit renal arteries. AB - The response of rabbit renal arteries to acetylcholine and its endothelial modulation in diabetes were investigated. Acetylcholine induced concentration related endothelium-dependent relaxation of renal arteries that was significantly more potent in diabetic rabbits than in control rabbits. Pretreatment with N(G) nitro-L-arginine (L-NOArg), indomethacin, or L-NOArg plus indomethacin induced partial inhibition of acetylcholine-induced relaxation. Inhibition induced by L NOArg plus indomethacin was significantly higher in arteries from diabetic rabbits than in arteries from control rabbits. In renal arteries depolarised with KCl 30 mM and incubated with L-NOArg plus indomethacin, acetylcholine-induced relaxation was almost abolished in both groups of rabbits and this response was not different from that obtained in arteries without endothelium. Sodium nitroprusside induced concentration-dependent relaxation of renal arteries from control and diabetic rabbits without significant differences between the two groups of animals. These results suggest that diabetes potentiates the acetylcholine-induced relaxation in rabbit renal arteries. Increased release of nitric oxide and prostacyclin could be responsible for the enhanced relaxant potency of acetylcholine in diabetes. PMID- 11275004 TI - 2-Arachidonoylglycerol, a candidate of endothelium-derived hyperpolarizing factor. AB - We investigated whether 2-arachidonoylglycerol, an endogenous cannabinoid receptor ligand, is involved in acetylcholine- and calcium ionophore A23187 induced relaxations in the presence of N(G)-nitro-L-arginine methyl ester (L NAME) and indomethacin, which is considered to be mediated by endothelium-derived hyperpolarizing factor (EDHF). In rabbit mesenteric arterial rings pre constricted with noradrenaline, 2-arachidonoylglycerol caused concentration dependent relaxation. The 2-arachidonoylglycerol-induced relaxations were not affected by endothelium removal. N-piperidino-5-(4-chlorophenyl)-1-(2,4 dichlorophenyl)-4-methyl-3-pyrazole-caroxamide (SR141716A) and 1-(2,4 dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-4-morholinyl-1H-pyrazole-3 carboxamide (AM281), cannabinoid CB(1) receptor antagonists, significantly attenuated 2-arachidonoylglycerol-induced relaxation and the acetylcholine induced relaxation only slightly, but not the calcium ionophore A23187-induced relaxation. On the other hand, charybdotoxin plus apamin, K(+) channel blockers, significantly attenuated acetylcholine and calcium ionohore A23187-induced relaxations but not 2-arachidonoylglycerol-induced relaxations. These results suggest that 2-arachidonoylglycerol can cause relaxations via cannabinoid CB(1) receptors, but is not involved in EDHF-mediated relaxations. PMID- 11275005 TI - Contribution of peroxynitrite to the beneficial effects of preconditioning on ischaemia-induced arrhythmias in rat isolated hearts. AB - We studied the effects of urate, a peroxynitrite scavenger, on ischaemia- and peroxynitrite-induced preconditioning in rat isolated hearts. Isolated hearts perfused with Krebs-Henseleit solution were preconditioned either by 3 min of coronary artery occlusion or by peroxynitrite administration (1 microM) for 3 min, followed by 10 min of reperfusion and 30 min of coronary artery occlusion. Both ischaemia and peroxynitrite produced a marked reduction in arrhythmias. Urate (1 mM) added to the perfusate 10 min prior to ischaemic preconditioning or peroxynitrite infusion and maintained until coronary artery occlusion, markedly reversed the beneficial effects in the ischaemic and peroxynitrite-treated groups. Urate administration in the peroxynitrite-treated group increased the incidence of ventricular tachycardia from 57% (n = 11) to 100% (n = 6) and total ventricular fibrillation from 0% (n=0) to 44% (n=4). Similarly, urate augmented the incidence of ventricular tachycardia from 47% (n=8) to 85% (n = 6) in the ischaemic preconditioning group. On its own, urate did not affect the severity of cardiac arrhythmias. Peroxynitrite infusion caused a marked increase in the effluent nitrate levels, from 0.05 +/- 0.1 microM (n = 5) to 0.4 +/- 0.2 microM (n = 6), and urate significantly decreased these levels to 0.08 +/- 0.03 microM (n = 9). These results suggest that peroxynitrite at low concentrations contributes to the beneficial effects of preconditioning on ischaemia-induced arrhythmias in rat isolated hearts. PMID- 11275006 TI - Melatonin inhibits calcitonin gene-related peptide-induced vasodilation and increase in cAMP in rat middle cerebral arteries. AB - The action of melatonin to alter calcitonin gene-related peptide (CGRP)-mediated vasodilation and stimulation of adenylate cyclase activity in middle cerebral arteries of rats was investigated. Concentration-dependent dilation of the rat middle cerebral artery produced by CGRP (EC(50) of 9.4 x 10(-10) M) was significantly inhibited in the presence of 10(-8) M melatonin (EC(50) of 3.4 x 10(-9) M). In addition, CGRP (10(-7) M)-mediated increase in adenylate cyclase activity was also significantly attenuated by the receptor mediated action of melatonin. These results indicate that melatonin may interact with CGRP to regulate cerebral arterial tone. PMID- 11275007 TI - Relaxation of mouse isolated aorta to adenosine and its analogues does not involve adenosine A(1), A(2) or A(3) receptors. AB - Relaxations to adenosine and analogues were investigated in the mouse aorta in the presence of the adenosine A(1) receptor-selective antagonist 1,3-dipropyl-8 cyclopentylxanthine (DPCPX, 30 nM), which did not affect relaxations to adenosine or its analogue N(6)-R-phenylisopropyladenosine (R-PIA) but abolished contractile adenosine A(1) receptor-mediated responses to these agonists. Relaxations to adenosine, 5'-N-ethylcarboxamidoadenosine, R-PIA, 2-[p-(2-carbonylethyl) phenylethylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680), and N(6)-(3 iodobenzyl)-adenosine-5'-N-methyluronamide (IB-MECA) were unaffected by the adenosine A(1)/A(2) receptor antagonist 8-sulphophenyltheophylline (100 microM). IB-MECA relaxations were unaffected by the adenosine A(3) receptor-selective antagonist 3-ethyl-5-benzyl-2-methyl-6-phenyl-4-phenylethynyl-1,4-(+/-) dihydropyridine-3,5-dicarboxylate (MRS1191, 30 microM) and R-PIA relaxations were unaffected by N(G)-nitro-L-arginine methyl ester (100 microM) and endothelium removal. In conclusion, relaxant responses to adenosine and analogues do not involve adenosine A(1), A(2) or A(3) receptors and are endothelium- and nitric oxide-independent. PMID- 11275008 TI - beta(1)- and beta(3)-adrenoceptor mediated smooth muscle relaxation in hypothyroid rat ileum. AB - The effect of hypothyroidism on gastrointestinal beta(1)- and beta(3) adrenoceptor function and expression was examined in rat ileal smooth muscle preparations. (-)-Isoprenaline and the selective beta(3) agonist disodium (R,R)-5 [2-[[2-3-chlorophenyl)-2-hydroxyethyl]-amino]propyl]-1,3-benzodioxole-2,2 dicarboxylate (CL 316234) relaxed both control and hypothyroid tissues in a dose dependent manner. Responses to isoprenaline were reduced in tissues from hypothyroid rats, as was the shift produced with the beta(3)-adrenoceptor antagonist, 3-(2-ethylphenoxy)-1-[(1S)-1,2,3,4-tetrahydronaphth-1-ylamino]-(2S)-2 propanol oxalate (SR 59230A). No change was seen in responses to CL 316243. Experiments with a selective beta(1)-adrenoceptor antagonist produced results suggesting that isoprenaline did not act at this receptor. Messenger RNA levels for both beta(1)- and beta(2)-adrenoceptors were not affected by hypothyroidism. These results show that, unlike in adipose tissues, ileal beta(1)- and beta(3) adrenoceptors are not directly regulated by thyroid hormone and that beta(3) adrenoceptor coupling to the relaxation response is reduced in a rat model of hypothyroidism. PMID- 11275009 TI - Failure of AH11110A to functionally discriminate between alpha(1)-adrenoceptor subtypes A, B and D or between alpha(1)- and alpha(2)-adrenoceptors. AB - The potency of the putatively alpha(1B)-adrenoceptor selective drug, 1-[biphenyl 2-yloxy]-4-imino-4-piperidin-1-yl-butan-2-ol (AH11110A), to antagonize contraction upon stimulation of alpha(1A)-adrenoceptors in rat vas deferens and rat perfused kidney, alpha(1B)-adrenoceptors in guinea-pig spleen, mouse spleen and rabbit aorta, and alpha(1D)-adrenoceptors in rat aorta and pulmonary artery was evaluated and compared to that of a number of subtype-discriminating antagonists. N-[3-[4-(2-Methoxyphenyl)-1-piperazinyl]propyl]-3-methyl-4-oxo-2 phenyl-4H-1-benzopyran-8-carboxamide (Rec 15/2739) and (+/-)-1,3,5-trimethyl-6 [[3-[4-((2,3-dihydro-2-hydroxymethyl)-1,4-benzodioxin-5-yl)-1 piperazinyl]propyl]amino]-2,4(1H,3H)-pyrimidinedione (B8805-033) were confirmed as selective for alpha(1A)-adrenoceptors, 8-[2-[4-(2-methoxyphenyl)-1 piperazinyl]ethyl]-8-azaspiro[4.5]decane-7,9-dione (BMY 7378), 8-[2-(1,4 benzodioxan-2-ylmethylamino)ethyl]-8-azaspiro[4.5]decane-7,9-dione (MDL 73005EF), and cystazosin were found to be selective for alpha(1D)-adrenoceptors, whereas spiperone was weakly selective for alpha(1B)-over alpha(1A)-adrenoceptors. However, from the functional affinity profile obtained for AH11110A at alpha(1A) adrenoceptors (pA(2)=6.41 in rat vas deferens), alpha(1B)-adrenoceptors (pA(2)=5.40-6.54) and alpha(1D)-adrenoceptors (pA(2)=5.47-5.48), the affinity and presumed selectivity previously obtained for AH11110A in radioligand binding studies at native alpha(1B)- and cloned alpha(1b)-adrenoceptors (pK(i)=7.10-7.73) could not be confirmed. Additionally, AH11110A enhanced the general contractility of rat vas deferens, produced a bell-shaped dose-response curve of vasodilation in perfused rat kidney, and its antagonism in most other tissues was not simply competitive. The affinity of AH11110A for prejunctional alpha(2)-adrenoceptors in rabbit vas deferens (pA(2)=5.44) was not much lower than that displayed for alpha(1)-adrenoceptor subtypes, revealing that AH11110A, besides alpha(1) adrenoceptors, also interacts with alpha(2)-adrenoceptors, and thus may be unsuitable for alpha-adrenoceptor subtype characterization, at least in smooth muscle containing functional studies. PMID- 11275010 TI - Tachykinin NK(2) receptor mediates contraction and ion transport in rat colon by different mechanisms. AB - We have characterized the tachykinin NK(2) receptor-mediated contraction and vectorial ion transport responses in the muscularis mucosae and mucosa of the rat isolated distal colon, respectively. The tachykinin NK(2) receptor-selective antagonist nepadutant (c([(beta-D-GlcNAc)Asn-Asp-Trp-Phe-Dpr-Leu]c(2beta-5beta))) produced competitive antagonism of [betaAla(8)]neurokinin A-(4-10)-induced contraction (pK(B) = 9.3) in the muscularis mucosae, and insurmountable blockade of increases in short-circuit current (I(sc)) responses (pK(B) = 8.6) in the mucosa. However, this latter effect was completely reversed by washout of the antagonist. [betaAla(8)]Neurokinin A-(4-10)-induced contractions were unaffected by indomethacin (3 microM). In sharp contrast, I(sc) responses induced by [betaAla(8)]neurokinin A-(4-10) (100 nM) were inhibited (>70%) by indomethacin (3 microM), while I(sc) responses to substance P (3 microM) were unchanged. Our study provides the first evidence that in the same organ stimulation of tachykinin NK(2) receptors leads to two independent responses mediated by different effector mechanisms both of which are blocked (albeit with different kinetics) by the potent and selective tachykinin NK(2) receptor antagonist, nepadutant. PMID- 11275011 TI - Dysidotronic acid, a new sesquiterpenoid, inhibits cytokine production and the expression of nitric oxide synthase. AB - In a previous study, we reported a new bioactive sesquiterpenoid, named dysidotronic acid, to be a potent, selective human synovial phospholipase A(2) inhibitor. Dysidotronic acid is a novel, non-complex manoalide analogue lacking the pyranofuranone ring. We now investigate the effect of this compound on cytokine, nitric oxide and prostanoid generation on the mouse macrophage cell line RAW 264.7, where it showed a dose-dependent inhibition with inhibitory concentration 50% values in the micromolar range. This effect was also confirmed in the mouse air pouch injected with zymosan. Dysidotronic acid inhibited the production of tumor necrosis factor alpha and interleukin-1 beta as well as the production of nitric oxide, prostaglandin E(2) and leukotriene B(4). Decreased nitric oxide generation was the consequence of inhibition of the expression of nitric oxide synthase, whereas PGE(2) and LTB(4) reduction was due to inhibition of arachidonic acid bioavailability through a direct inhibitory effect of dysodotronic acid on secretory phospholipase A(2). PMID- 11275012 TI - Effects of F2833 on cholesterol metabolism in the genetically hyperlipidemic rat. AB - The effects of the new hypolipidemic agent, F2833 or (chloro 2' (1-1') biphenyl 4)-2 propanol-2, on cholesterol metabolism were studied in genetically hyperlipidemic rats (RICO). Cholesterolemia decreased after 2 days of treatment to 60% of its initial value (1.20+/-0.10 g/l vs. 1.99+/-0.08, P < 0.001) and then stabilised within 10 days. This hypocholesterolemic action was effective for as long as 3 months. Concerning the different classes of lipoproteins, a significant drop was observed in HDL (high density lipoproteins) (25%, 0.49 +/- 0.02 g/l vs. 0.66 +/- 0.007, P < 0.01) and particularly in LDL (low density lipoproteins) (70%, 0.30 +/- 0.04 g/l vs. 0.92 +/- 0.05, P < 0.001). Whole body cholesterol showed a higher fractional catabolic rate (0.25 +/- 0.02 vs. 0.17 +/- 0.005 day( 1), P < 0.01) together with an increased cholesterol synthesis (60 +/- 5 vs. 36 +/- 4 mg/day, P < 0.01). LDL kinetics showed that the decrease in these lipoproteins is essentially caused by an increase in the fractional catabolic rate (10.6 +/-0.1%/h vs. 5.2 +/- 0.1%/h, P < 0.001) and by a lesser decrease in the LDL production rate. This cholesterol metabolic profile created by treatment suggests an effect through stimulation of cholesterol output (biliary cholesterol elimination or cholesterol transformation into bile acids). PMID- 11275014 TI - Antenatal dexamethasone and decreased birth weight. AB - OBJECTIVE: To test the hypothesis that antenatal dexamethasone treatment to promote fetal lung maturation results in decreased birth weight corrected for gestational age. METHODS: The birth weights of all dexamethasone-treated, singleton, live-born infants delivered at our hospital were compared with our overall obstetric population; a group of untreated infants frequency matched approximately 3:1 according to maternal race, infant sex, and gestational age at delivery; and an historical cohort of infants with an indication for dexamethasone but delivered in the 12 months before the introduction of corticosteroid therapy at our hospital. RESULTS: Dexamethasone-treated infants (n = 961), when compared with either the overall population (n = 122,629) or matched controls (n = 2808), had significantly lower birth weights after adjustment for week of gestation (P <.001). Compared with the historical cohort of infants, the average birth weight of dexamethasone-treated infants was smaller by 12 g at 24 26 weeks, 63 g at 27-29 weeks, 161 g at 30-32 weeks, and 80 g at 33-34 weeks' gestation. CONCLUSION: Antenatal dexamethasone administered to promote fetal maturation is associated with diminished birth weight. PMID- 11275015 TI - Is betamethasone effective longer than 7 days after treatment? AB - OBJECTIVE: To determine whether perinatal outcomes are influenced by the interval between antenatal betamethasone administration and delivery. METHODS: We did a retrospective cohort analysis of live-born singleton neonates born between 28 and 34 weeks' gestation after a single course of betamethasone, defined as two 12-mg doses over 24 hours. Subjects were grouped according to length of interval between initial betamethasone dose and delivery (1-2 days, 3-7 days, and 8-14 days). We excluded women who had membranes ruptured for longer than 24 hours before delivery, delivery before the second dose of betamethasone, or more than two doses of betamethasone. Data were analyzed by Student t test, chi(2) test, or Fisher exact test. Multiple logistic regression analyses were done using suspected risk factors for respiratory distress syndrome (RDS) and intraventricular hemorrhage (IVH). We calculated that a sample of 200 women would provide more than 80% power to detect a 50% reduction in incidence of RDS for a two-sided test of significance at a critical level of.05. RESULTS: Among 216 women, 97 delivered in 1-2 days, 78 in 3-7 days, and 41 in 8-14 days after a single course of betamethasone. Groups were similar in selected demographics, tocolytic exposure, gestational age at delivery, modes of delivery, and mean birth weights. There were no significant differences in frequencies of RDS (39.2%, 41.1%, and 36.6%, respectively) or grades 3-4 IVH (1.1%, 1.3%, and 0%, respectively) between groups. Frequencies of selected perinatal infectious outcomes also were similar between groups. Multiple logistic regression analyses found no association between RDS or IVH and delivery more than 7 days from betamethasone therapy. CONCLUSION: There were no differences in perinatal outcomes in pregnancies delivered 8-14 days after antenatal exposure to betamethasone compared with those delivered within 7 days of exposure. PMID- 11275017 TI - Interval between fetal measurements in predicting growth restriction. AB - OBJECTIVE: To determine the influence of the interval between fetal measurements on performance of fetal growth velocity for predicting infants with anthropometric features of fetal growth restriction (FGR). METHODS: Two hundred seventy-four low-risk women had serial fetal biometry at scheduled intervals. Growth velocity of the fetal abdominal area for each was calculated with 2-, 4-, and 6-week scan intervals in which the second measurement was the last scan before delivery. Fetal abdominal area velocity over a 4-week interval in the early third trimester also was included. Fetal growth restriction was defined as skinfold thickness under the tenth percentile, ponderal index under the 25th percentile, midarm circumference-to-occipitofrontal circumference ratio of under 1 standard deviation (SD). Test performance was expressed as likelihood ratios with 95% confidence intervals (CI). RESULTS: Fetal abdominal area velocity calculated over a 4-week interval predicted FGR with a likelihood ratio of 10.4 (95% CI 3.9, 26) for skinfold thickness; 9.5 (95% CI 4.6, 19) for ponderal index; and 4.7 (2.3, 8.4) for midarm circumference-to-occipitofrontal circumference ratio. Intermeasurement intervals of 6 weeks had a likelihood ratio of 8.5 (95% CI 4, 17) for skinfold thickness; 7.5 (95% CI 3.4, 16.1) for ponderal index; and 14 (6.7, 28) for midarm circumference-to-occipitofrontal circumference ratio. The likelihood ratios for the 2-week interval and the early third trimester 4-week interval were all less than 5. CONCLUSION: Four- and 6-week measurement intervals were useful for predicting infants with FGR and were superior to a 2-week interval. Fetal growth velocity is influenced by proximity of the last fetal measurement to date of delivery, which adversely affects clinical use of growth velocity for predicting FGR. PMID- 11275016 TI - Effect of labor on infant morbidity and mortality with preterm premature rupture of membranes: United States population-based study. AB - OBJECTIVE: To evaluate whether labor, in the setting of premature rupture of membranes (PROM), affects infant morbidity and mortality rates. METHODS: We derived data for this population-based cohort study from the United States national linked birth infant death data sets, comprised of singleton live births delivered between 1995 and 1997. We included women (n = 34,594) who had preterm PROM more than 12 hours and delivered between 23 and 32 weeks' gestation. Birth records were used to determine whether delivery occurred with or without labor. Infants with birth weights below the tenth percentile for gestational age were classified as small for gestational age (SGA) on the basis of a nomogram of all singleton births in the United States between 1995 and 1997. Primary outcomes were early neonatal (0-6 days), late neonatal (7-27 days), postneonatal (28-365 days), and infant death (0-365 days). Secondary outcomes included respiratory distress syndrome (RDS), assisted ventilation, and neonatal seizures. Risks of infant mortality and morbidity from labor were examined separately for SGA and non-SGA infants. RESULTS: Overall rates were infant death 11.6%, RDS 15.1%, assisted ventilation 25.9%, and neonatal seizure 0.2%. Labor was associated with higher incidence of early neonatal death in SGA infants (adjusted relative risk [RR] 1.24, 95% confidence interval [CI] 1.11, 1.38) but had no effect on other outcomes. Among non-SGA infants, labor had no effect on infant death but was associated with higher rates of RDS (RR 1.15, 95% CI 1.08, 1.22) and assisted ventilation (RR 1.16, 95% CI 1.08, 1.24). CONCLUSION: Although labor was associated with a slightly higher mortality rate in SGA infants and slightly more respiratory morbidity in non-SGA infants, recommendations regarding clinical treatment should await future clinical trials. PMID- 11275018 TI - Rescue by birth: defective placental maturation and late fetal mortality. AB - OBJECTIVE: To estimate the incidence and lethality of placental maturation defect, and to determine the impact of the pattern of placental dysfunction on the risk of recurrent stillbirth or maternal disease in later life. METHODS: Questionnaire and archival analysis of fetal deaths from placental dysfunction at 32-42 weeks (1975-1995 in Zurich), classified as chronic (parenchyma loss) or acute (maturation defect of the terminal chorionic villi). Population survey of 17,415 consecutive unselected singleton placentas (1994-1998 in Berlin). RESULTS: Of the 71 stillbirths, 34 were due to parenchyma loss and 37 to maturation defect. Parenchyma loss predominated in the first pregnancy (73.5% compared with 43.2%; P <.05). The risks of recurrent stillbirth and subsequent childlessness did not differ between the two groups. Eleven percent of mothers whose placenta had maturation defect had diabetes in the index pregnancy; none of the other women in the group developed diabetes over the 5-20-year observation period. In the population survey, incidence of maturation defect was 5.7%, and was associated with fetal death in 2.3% of cases. Normal placentas were associated with fetal death in 0.033%. CONCLUSION: Placental maturation defect can be a cause of fetal hypoxia. Although the risk of stillbirth is 70-fold that of a normal placenta, few affected fetuses actually die. The risk of recurrent stillbirth is tenfold above baseline and occurs mostly after 35 weeks' gestation. PMID- 11275019 TI - Immunologic studies in presumed amniotic fluid embolism. AB - OBJECTIVE: To evaluate the potential role of immunologic mechanisms that involve mast cell degranulation (anaphylaxis) or complement activation in the mechanism of amniotic fluid embolism. METHODS: This study was a case series of nine women with presumed amniotic fluid embolism and a control group of 22 women who had normal labor. Women were from community and tertiary referral hospitals in Japan and the United States. Main outcome measures were maternal peripartum complement levels (C3 and C4), serum levels of tryptase, urinary histamine concentrations, and serum levels of a fetal antigen (sialyl Tn). RESULTS: Serum tryptase and urinary histamine measurements were negative in women with amniotic fluid embolism; seven of nine had elevated levels of fetal antigen. All eight who had serum available for testing had abnormally low levels of complement. Mean C3 level of 44.0 mg/dL and C4 level of 10.7 mg/dL were significantly lower than corresponding postpartum control values of 117.3 mg/dL and 29.4 mg/dL (P =.018 for C3, P =.012 for C4). Postpartum C3 and C4 levels decreased by 8% and 5%, respectively, compared with intrapartum values (P =.003 for C3, P =.021 for C4) but were still within normal range. CONCLUSION: Serologic findings suggest a role for complement activation in the mechanism of amniotic fluid embolism. Laboratory data from this series did not implicate mast cell degranulation (anaphylaxis) in the pathophysiology of the disease. PMID- 11275020 TI - Pulse pressure and risk of preeclampsia: a prospective study. AB - OBJECTIVE: To find whether pulse pressure, a measure of arterial compliance, is associated early in pregnancy with increased risk of developing preeclampsia. METHODS: In a prospective cohort of 576 nulliparas, we examined blood pressures throughout pregnancy and at 6-8 weeks postpartum. Measurements during weeks 7-15, 16-24, and 25-38 of gestation were pooled to find averages for each period. Outcomes assessed were gestational hypertension and preeclampsia. Logistic regression analysis was used to develop relative risks and 95% confidence intervals. RESULTS: We confirmed 34 (5.9%) cases of preeclampsia, 32 (5.6%) cases of gestational hypertension, and 510 normotensive women. Mean systolic and diastolic blood pressures and mean arterial pressures were elevated throughout pregnancy in women who developed hypertensive disorders of pregnancy compared with normotensive women. Pulse pressure at 7-15 weeks was significantly higher in women who developed preeclampsia (45 +/- 6 mmHg) than in those who developed gestational hypertension (41 +/- 7 mmHg, P =.03) and normotensive women (41 +/- 8 mmHg, P =.01). Examined in tertiles, increasing pulse pressure was associated with increasing risk of developing preeclampsia (P for trend =.01) but not gestational hypertension (P for trend =.95). After adjustment for potential confounders, a 1-mmHg rise in early pregnancy pulse pressure was associated with a 6% (95% confidence interval: 1, 10) increase in risk for developing preeclampsia but not gestational hypertension (relative risk: 1%; 95% confidence interval: -1, 6). Beyond 15 weeks' gestation, differences between groups diminished, but women with any hypertensive disorder had higher pulse pressures than women with uncomplicated pregnancies. CONCLUSION: Elevated pulse pressure, indicating poor arterial compliance, was evident early in pregnancies of women who subsequently developed preeclampsia. PMID- 11275021 TI - Serum insulin, insulin-like growth factor-I, and insulin-like growth factor binding protein-1 in women who develop preeclampsia. AB - OBJECTIVE: To determine whether second-trimester serum concentrations of insulin, insulin-like growth factor-I (IGF-I), and insulin-like growth factor binding protein-1 (IGFBP-1) were altered in women before they developed clinical signs of preeclampsia. METHODS: A nested case-control study used serum obtained during second-trimester pregnancies from 12 women who developed preeclampsia matched with 24 controls who remained normotensive. Nine preeclamptic subjects and 18 controls were necessary to have 80% power to discern a 20% difference between groups with regard to the analytes under consideration. RESULTS: There were no significant differences between cases and controls with respect to many demographic factors. Women who developed preeclampsia had insulin concentrations that were not significantly different from controls, but serum concentrations of IGF-I were significantly higher and IGFBP-1 were significantly lower than those of the controls. The IGF-I/IGFBP-1 ratio helped to identify those at risk for developing preeclampsia. CONCLUSIONS: Serum concentrations of IGF-I and IGFBP-1 were abnormal long before women manifested clinical evidence of preeclampsia in this study. These alterations might be related to abnormalities in trophoblastic invasion and prove useful as potential markers for the identification of women who are at high risk of developing preeclampsia. PMID- 11275022 TI - Serum homocysteine at 16 weeks and subsequent preeclampsia. AB - OBJECTIVE: To determine whether elevated homocysteine levels precede the development of preeclampsia. METHODS: Study subjects were selected from a population-based cohort of 1049 nulliparous women from whom serum was collected for Down syndrome screening at 16 weeks' gestation. For 34 women who developed preeclampsia, 68 control women were chosen who remained normotensive. Homocysteine was analyzed by high-performance liquid chromatography and fluorescence detection. The sample size allowed detection of a 1.25-micromol/L difference at a significance level of 0.05 and the power of 0.81. RESULTS: At 16 weeks' gestation, concentrations (mean, 95% confidence interval) of homocysteine in women who developed preeclampsia, 6.99 (6.42, 7.55) micromol/L, were similar to those who remained normotensive, 6.91 (6.45, 7.34) micromol/L. CONCLUSION: Significant changes in homocysteine metabolism did not predate the appearance of clinical preeclampsia. PMID- 11275023 TI - Serum soluble Fas levels in preeclampsia. AB - OBJECTIVE: To determine if serum soluble Fas levels are altered in women with preeclampsia. METHODS: Thirty-four pregnant women with preeclampsia and 34 normotensive pregnant women were studied. Subjects were matched as much as possible for demographics. Preeclampsia was defined as proteinuric hypertension. Serum soluble Fas levels were measured by enzyme-linked immunoassay. Two-tailed Student t test, chi(2) test, Pearson correlation coefficients, and analysis of variance with post hoc test were used for statistical analyses. RESULTS: Mean serum soluble Fas levels were significantly higher in preeclamptic than normotensive women (10.59 +/- 0.68 U/mL versus 5.65 +/- 0.35 U/mL, P <.001). CONCLUSION: Elevated serum soluble Fas is associated with preeclampsia. Such elevation might indicate protection of maternal T-lymphocyte apoptosis and consequently lead to the maternal immune intolerance noted in preeclampsia. PMID- 11275024 TI - Pregnancy-related mortality from preeclampsia and eclampsia. AB - OBJECTIVE: To examine the role of preeclampsia and eclampsia in pregnancy-related mortality. METHODS: We used data from the Centers for Disease Control and Prevention's Pregnancy Mortality Surveillance System to examine pregnancy-related deaths from preeclampsia and eclampsia from 1979 to 1992. The pregnancy-related mortality ratio for preeclampsia-eclampsia was defined as the number of deaths from preeclampsia and eclampsia per 100,000 live births. Case-fatality rates for 1988-1992 were calculated for preeclampsia and eclampsia deaths per 10,000 cases during the delivery hospitalization, using the National Hospital Discharge Survey. RESULTS: Of 4024 pregnancy-related deaths at 20 weeks' or more gestation in 1979-1992, 790 were due to preeclampsia or eclampsia (1.5 deaths/100,000 live births). Mortality from preeclampsia and eclampsia increased with increasing maternal age. The highest risk of death was at gestational age 20-28 weeks and after the first live birth. Black women were 3.1 times more likely to die from preeclampsia or eclampsia as white women. Women who had received no prenatal care had a higher risk of death from preeclampsia or eclampsia than women who had received any level of prenatal care. The overall preeclampsia-eclampsia case fatality rate was 6.4 per 10,000 cases at delivery, and was twice as high for black women as for white women. CONCLUSION: The continuing racial disparity in mortality from preeclampsia and eclampsia emphasizes the need to identify those differences that contribute to excess mortality among black women, and to develop specific interventions to reduce mortality from preeclampsia and eclampsia among all women. PMID- 11275025 TI - Safety and efficacy of levonorgestrel implant, intrauterine device, and sterilization. AB - OBJECTIVE: To evaluate safety and efficacy of levonorgestrel-releasing contraceptive implants (Norplant; Leiras Oy, Turku, Finland) in developing countries. METHODS: We used controlled cohort methodology. Women attending family planning clinics in eight developing countries selecting Norplant were enrolled, together with women of similar age choosing intrauterine devices (IUDs) or surgical sterilization. Participants were interviewed and examined at semi-annual visits and followed-up for 5 years regardless of change of contraceptive methods. Incidence rate ratios of health events were estimated for initial and current method use. RESULTS: Altogether, 7977 women initiated Norplant, 6625 IUD, and 1419 sterilization. The overall follow-up rate was 94.6% and 78,323 woman-years of observation were accumulated. Pregnancy rates for Norplant, copper IUDs, and sterilization each averaged less than 1 per 100 woman-years. With two exceptions, no significant excess risk of serious morbidity was detected for Norplant users compared with controls. The incidence of gallbladder disease was higher in women who initiated Norplant use than in controls (rate ratio 1.52, 95% confidence interval [CI] 1.02, 2.27), as was the incidence of hypertension and borderline hypertension in current implant users (rate ratio 1.81; CI 1.12, 2.92). Other new findings were increased risks of respiratory diseases and decreased risks of inflammatory disease of the genital tract in Norplant users compared with IUD users and sterilized women. CONCLUSION: The study confirms the safety with respect to serious disease and the high contraceptive efficacy of Norplant, copper IUDs, and sterilization. PMID- 11275026 TI - Estrogen replacement therapy and nocturnal periodic limb movements: a randomized controlled trial. AB - OBJECTIVE: To evaluate the effect of estrogen replacement therapy on nocturnal periodic limb movements in a randomized, double-masked, placebo-controlled, crossover trial. METHODS: Seventy-one healthy postmenopausal women volunteered in answer to a newspaper announcement; 62 women completed the follow-up. Frequency of nocturnal body movements was measured with the static-charge-sensitive bed and all-night polysomnographic recordings. Serum estradiol (E2) and FSH concentrations were also measured at baseline and after each treatment period. The power of the study setup was 94%. RESULTS: Nearly half the women presented with episodes of periodic limb movements (30 of 62 women, or 48%, during placebo and 27, or 44%, during estrogen therapy). In 17 (27%) during placebo and 19 (31%) during estrogen therapy, frequency of periodic limb movements exceeded index level 5 per hour while subjects were in bed. Incidence or intensity of movements, movement durations, and movement intervals did not change with estrogen therapy. The arousal index was similar during the two treatments (medians = 1.7 for placebo and 1.3 for estrogen, P =.758). Variations in serum E2 concentration, age, and body mass index did not explain variations in movement activity. CONCLUSION: Estrogen replacement therapy in doses used to control climacteric symptoms does not alter the incidence or intensity of nocturnal periodic limb movements. PMID- 11275027 TI - Estrogen replacement therapy in endometrial cancer patients: a matched control study. AB - OBJECTIVE: To determine if estrogen replacement therapy, in women with a history of endometrial cancer, increases the risk of recurrence or death from that disease. METHODS: Two hundred forty-nine women with surgical stage I, II, and III endometrial cancer were treated between 1984 and 1998; 130 received estrogen replacement after their primary cancer treatments and 49% received progesterone in addition to estrogen. Among this cohort, 75 matched treatment-control pairs were identified. The two groups were matched by using decade of age at diagnosis and stage of disease. Both groups were comparable in terms of parity, grade of tumor, depth of invasion, histology, surgical treatment, lymph node status, postoperative radiation, and concurrent diseases. The outcome events included the number of recurrences and deaths from disease. RESULTS: The hormone users were followed for a mean interval of 83 months (95% confidence interval [CI] 71.0, 91.4) and the nonhormone users were followed for a comparable mean interval of 69 months (CI 59.1, 78.7). There were two recurrences (1%) among the 75 estrogen users compared with 11 (14%) recurrences in the 75 nonhormone users. Hormone users had a statistically significant longer disease-free interval than nonestrogen users (P =.006). CONCLUSION: Estrogen replacement therapy with or without progestins does not appear to increase the rate of recurrence and death among endometrial cancer survivors. PMID- 11275028 TI - Secretory leukocyte protease inhibitor in ovarian endometriomas following GnRH agonist therapy. AB - OBJECTIVE: To determine whether expression of secretory leukocyte protease inhibitor is affected in tissue and peritoneal fluid of women with ovarian endometriomas treated with GnRH analogues. METHODS: In 32 women with endometriomas (17 untreated and 15 treated with GnRH analogue) and 21 with ovarian cystadenomas, we examined the expression of secretory leukocyte protease inhibitor messenger RNA (mRNA) by Northern blot analysis; protein distribution was measured immunohistochemically. Concentrations of secretory leukocyte protease inhibitor in peritoneal fluid were measured by enzyme-linked immunosorbent assay. Expression of secretory leukocyte protease inhibitor in endometrioma explants in vitro was also studied with and without the GnRH analogue treatment. RESULTS: Secretory leukocyte protease inhibitor mRNA expression was identified only in untreated endometriomas. In the GnRH agonist treated endometriomas, the semiquantitative H-score for secretory leukocyte protease inhibitor immunostaining was significantly lower than that for untreated endometriomas (P <.001). The peritoneal fluid of the GnRH agonist-treated women also contained significantly lower concentrations of secretory leukocyte protease inhibitor (median 76 ng/mL, interquartile range 51-131 ng/mL; P <.001) than untreated women (124 ng/mL, 70-186 ng/mL). Secretory leukocyte protease inhibitor in endometrioma explants in vitro was significantly inhibited by the GnRH analogue (P <.05). CONCLUSION: Expression of secretory leukocyte protease inhibitor in tissue and peritoneal fluid of women with ovarian endometriomas was decreased by GnRH agonist treatment. PMID- 11275029 TI - Effect of academic affiliation and obstetric volume on clinical outcome and cost of childbirth. AB - OBJECTIVE: To determine whether the academic affiliation and obstetric volume of the delivering hospital has an impact on clinical and economic outcomes. METHODS: We performed a cross-sectional analysis of data for all births in the State of Maryland during 1996. Acute hospital discharge data were obtained from the publicly available Maryland Health Services Cost Review Commission database. Institutions were classified as community hospitals, community teaching hospitals, and academic medical centers. Principal outcome variables included cesarean birth and complication rates, total hospital charges, and length of stay. RESULTS: A total of 63,143 cases were identified for analysis. The cesarean delivery rate was lower among academic medical centers, compared with community teaching hospitals and community hospitals (18.4% compared with 24.3% and 21.2%, respectively). After adjustment for patient case-mix, the adjusted odds ratio (OR) for cesarean birth was 0.66 at academic medical centers and 1.23 at community teaching hospitals compared with community hospitals (P <.01). Rates of episiotomy and serious complications were lower at academic medical centers compared with community hospitals. Adjusted total hospital charges were lower and length of stay was shorter for community hospitals compared with academic medical centers ($2937 compared with $3564 and 2.2 days compared with 2.5 days, respectively). CONCLUSION: Hospital academic affiliation was an important predictor of clinical outcomes. Better clinical outcomes were found primarily among patients at academic medical centers, although these institutions demonstrated moderately higher resource utilization, compared with community hospitals. PMID- 11275030 TI - Ginger for nausea and vomiting in pregnancy: randomized, double-masked, placebo controlled trial. AB - OBJECTIVE: To determine the effectiveness of ginger for the treatment of nausea and vomiting of pregnancy. METHODS: Women with nausea and vomiting of pregnancy, who first attended an antenatal clinic at or before 17 weeks' gestation, were invited to participate in the study. During a 5-month period, 70 eligible women gave consent and were randomized in a double-masked design to receive either oral ginger 1 g per day or an identical placebo for 4 days. Subjects graded the severity of their nausea using visual analog scales and recorded the number of vomiting episodes in the previous 24 hours before treatment, and again during 4 consecutive days while taking treatment. At a follow-up visit 7 days later, five item Likert scales were used to assess the severity of their symptoms. RESULTS: All participants except three in the placebo group remained in the study. The visual analog scores of posttherapy minus baseline nausea decreased significantly in the ginger group (2.1 +/- 1.9) compared with the placebo group (0.9 +/- 2.2, P =.014). The number of vomiting episodes also decreased significantly in the ginger group (1.4 +/- 1.3) compared with the placebo group (0.3 +/- 1.1, P <.001). Likert scales showed that 28 of 32 in the ginger group had improvement in nausea symptoms compared with 10 of 35 in the placebo group (P <.001). No adverse effect of ginger on pregnancy outcome was detected. CONCLUSION: Ginger is effective for relieving the severity of nausea and vomiting of pregnancy. PMID- 11275031 TI - Urinary interleukin-8 with asymptomatic bacteriuria in pregnancy. AB - OBJECTIVE: To evaluate urinary interleukin-8 (IL-8), an inflammatory cytokine, as a screening method for detecting asymptomatic bacteriuria in pregnancy. METHODS: Clean-catch urine samples from 200 pregnant women undergoing screening for asymptomatic bacteriuria were evaluated by urine culture, urine dipstick analysis, and measurement of IL-8. Interleukin-8 levels were measured by a chemiluminescent immunoassay (Immulite IL-8, Diagnostic Products Corp., Los Angeles, CA), and a receiver operating characteristic curve was used to determine the optimal cutoff point. Asymptomatic bacteriuria was defined as at least 100,000 colony-forming units of a single organism per mL. Dipstick testing included nitrite assessment as positive or negative and leukocyte esterase as negative, trace, 1+, 2+, or 3+. Dipstick testing was considered positive if nitrite was positive or leukocyte esterase was trace or greater. Sensitivities, specificities, positive and negative predictive values were determined for urinary leukocyte esterase and nitrite and compared with those of IL-8. chi(2) and Mann-Whitney U tests were used for statistical analyses. RESULTS: Twenty women were identified with asymptomatic bacteriuria by urine culture. The median urinary IL-8 levels for women with and without asymptomatic bacteriuria were 356 pg/mL and 125 pg/mL, respectively (P <.01, Mann-Whitney U test). Using an optimal cutoff point of 264 pg/mL, IL-8 had a sensitivity, specificity, positive and negative predictive value of 70%, 67%, 19%, and 95% for predicting asymptomatic bacteriuria. Urine dipstick analysis with either a positive leukocyte esterase or nitrite had a sensitivity, specificity, positive and negative predictive value of 45%, 62%, 12%, and 91%, respectively, for detecting asymptomatic bacteriuria. The differences between these testing methods were not statistically significant. CONCLUSION: Urinary interleukin-8 is not an acceptable screening method for asymptomatic bacteriuria in pregnancy because it fails to detect 30% of women with this condition. PMID- 11275032 TI - Fetal weight and progression of diabetic retinopathy. AB - OBJECTIVE: To test the hypothesis that progression of diabetic retinopathy in pregnancy is associated with reduced fetal growth and related neonatal morbidity. METHODS: Women with type 1 diabetes (n = 205) were enrolled before 14 weeks' gestation in a prospective study of diabetes in pregnancy and treated with intensive insulin therapy. They had serial ophthalmologic evaluations before 20 weeks' gestation and in late gestation or postpartum. Subjects were divided into two groups based on whether retinopathy progressed (progression group) or remained unchanged (no progression group). RESULTS: Retinopathy progressed in 59 of 205 women (29%) and was associated with advanced White classification (P =.001): three (5%) were class B, 14 (23%) class C, 24 (41%) class D, and 18 (30%) class F-RF. Reduced fetal growth was associated with progression of retinopathy. Mean birth weight was lower (P =.02), and more infants were small for gestational age (P =.02) and had low birth weights (P =.02) in the progression group. More large-for-gestational-age infants were noted in the no-progression group (P =.04). Birth weight percentile distributions showed a shift of the curve to the left in the progression group (P =.03). There were no differences in gestational age at delivery, macrosomia, preterm delivery, respiratory distress syndrome, neonatal hypoglycemia, or neonatal death. Small for gestational age was associated with chronic hypertension (odds ratio [OR] 6.4; 95% confidence interval [CI] 1.5, 27.9) and retinopathy progression (OR 4.7; 95% CI 1.2, 23.8). CONCLUSION: Development and progression of diabetic retinopathy during pregnancy were associated with reduced fetal growth manifested as increased rate of small for-gestational-age and low-birth-weight infants. PMID- 11275033 TI - Nucleated red blood cells in meconium aspiration syndrome. AB - OBJECTIVE: To evaluate whether the absolute nucleated red blood cell (RBC) count is higher in infants who had meconium aspiration with respiratory symptoms compared with infants with asymptomatic meconium aspiration and controls. METHODS: We compared the absolute nucleated RBC counts during the first 12 hours of life in three groups of term, vaginally delivered infants, including those who had meconium aspiration with respiratory symptoms (n = 11), asymptomatic meconium aspiration (n = 45), and control healthy infants (n = 32). We excluded infants of women with diabetes in pregnancy; hypertension; alcohol, tobacco, or drug abuse; and those with hemolysis, blood loss, or chromosomal anomalies. RESULTS: There were no significant differences among groups in gestational age; gravidity; parity; maternal analgesia; lymphocyte, platelet, and granulocyte counts; and hematocrit. The median nucleated RBC count was significantly higher in the meconium aspiration group with respiratory symptoms (0.007 x 10(9)/L) than the asymptomatic meconium aspiration group (0.004 x 10(9)/L) or controls (0.003 x 10(9)/L). CONCLUSION: At birth, infants with meconium aspiration syndrome had higher absolute nucleated RBC counts compared with infants with asymptomatic meconium aspiration and normal infants. PMID- 11275034 TI - Psychologic and obstetric predictors of couples' grief during pregnancy after miscarriage or perinatal death. AB - OBJECTIVE: To determine if the psychologic constructs of self-criticism and marital adjustment, considered jointly with obstetric and demographic factors, are significant predictors of grief during a pregnancy after a miscarriage or perinatal death. METHODS: Participants included 60 pregnant women with previous miscarriages or perinatal deaths, and 50 of their partners. Participants completed a package of psychometric instruments between the tenth and 19th week of gestation. Predictors of grief (active grief, difficulty coping, despair) included (1) psychologic factors: marital adjustment and self-criticism; (2) demographic factors: age and number of living children; and (3) obstetric factors: gestational age at time of loss, number of losses, and time between loss and subsequent conception. RESULTS: Stepwise regression analyses were conducted for each grief component for women and men. For women, active grief was significantly associated with high self-criticism and later losses (R(2) = 0.31). Later losses and longer time between loss and conception were significantly associated with difficulty coping (R(2) = 0.55) and despair (R(2) = 0.44). In men, active grief was associated with high self-criticism and later losses (R(2) = 0.28), difficulty coping (R(2) = 0.18), and despair (R(2) = 0.25) with high self-criticism. A trend was found for poor marital adjustment to be associated with higher levels of difficulty coping and despair in men. CONCLUSION: High levels of self-criticism and later gestational age at time of loss are predictors of increased grief during a pregnancy after a miscarriage or perinatal death. Increased time between loss and subsequent conception is also predictive of increased grief for women. For men, low levels of marital adjustment are predictive of increased grief. These results may be helpful in counselling couples considering pregnancy after a loss. PMID- 11275035 TI - A randomized comparison of transcervical Foley catheter to intravaginal misoprostol for preinduction cervical ripening. AB - OBJECTIVE: To compare the efficacy of intravaginal misoprostol tablets with transcervical Foley catheter for preinduction cervical ripening. METHODS: Pregnant women who presented for induction of labor with unfavorable cervices (Bishop score less than 6) were assigned randomly to intravaginal misoprostol (50 microg tablet every 4 hours for a maximum of six doses) or 30-mL Foley catheter placed transcervically with maintenance of traction. RESULTS: Among 111 women, 53 were allocated to misoprostol and 58 to Foley bulb. Contractile abnormalities were more frequent in the misoprostol group (20.4%) than the Foley group (0%) (P <.001). No statistically significant differences were noted between groups in change in Bishop score, preinduction cervical ripening times, and total induction times. There were no statistically significant differences in mode of delivery or adverse neonatal outcomes. Uterine rupture occurred in one woman with two previous cesarean deliveries in the misoprostol group. CONCLUSION: Intravaginal misoprostol and transcervical Foley catheter are equivalent for cervical ripening. Uterine contractile abnormalities and meconium passage are more common with misoprostol. PMID- 11275036 TI - Epidural analgesia and fetal head malposition at vaginal delivery. AB - OBJECTIVE: To determine if nulliparas who delivered with on-demand epidural analgesia are more likely to have malpositioning of the fetal vertex at delivery than women delivered during a period of restricted epidural use. METHODS: A retrospective cohort of nulliparous women with spontaneous labor delivered during a 12-month period immediately before the availability of on-demand labor epidural analgesia was compared with a similar group of nulliparas delivered after labor epidural analgesia was available on request. The primary outcome variable was a non-occiput anterior position or malpositioned fetal head at vaginal delivery. RESULTS: The frequency of epidural use increased from 0.9% before epidural analgesia became available on demand to 82.9% afterward. Fetal head malpositioning at vaginal delivery occurred in 26 of 434 (6.0%) women delivered in the before period compared with 29 of 511 (5.7%) in the after period (relative risk 0.95, 95% confidence interval 0.6, 1.6). No statistically significant difference in the incidence of fetal head malpositioning was present after patients were stratified by mode of delivery (Mantel-Haenszel weighted relative risk 0.94, 95% confidence interval 0.6, 1.4). The study sample size provided 85% power to detect a two-fold increase in the incidence of fetal malpositioning from a baseline rate of 6% associated with on-demand epidural use. CONCLUSION: Providing on-request labor epidural analgesia to nulliparas in spontaneous labor did not result in a clinically significant increase in the frequency of fetal head malpositioning at vaginal delivery. PMID- 11275037 TI - Vaginal route as the norm when planning hysterectomy for benign conditions: change in practice. AB - OBJECTIVE: To investigate if a deliberate decision to carry out as many hysterectomies as possible by the vaginal route can be effective in increasing the proportion of vaginal hysterectomies for benign conditions in the absence of prolapse. METHODS: Practice over 5 years at a district general hospital in the United Kingdom was studied. Patients with prolapse, adnexal disease, leiomyoma larger than 16 weeks, and malignancy were excluded, leaving 272 hysterectomies of 553 originally. Change in the route of hysterectomy, the main endpoint, was observed at yearly intervals. RESULTS: At the start of the study, the route of surgery was 68% abdominal and 32% vaginal. By the end of the fifth year the pattern was 5% abdominal 95% vaginal. The conversion from vaginal to abdominal hysterectomy occurred in only two cases during the study period. There was no change in the case mix during this period. In the fifth year of study most associated oophorectomies were also performed vaginally. There was no increase in patient morbidity. CONCLUSION: A major determinant of the route of hysterectomy is not the clinical situation but the attitude of the surgeon. There is no need for extra training and special skills or complicated equipment for vaginal hysterectomy. PMID- 11275038 TI - Effects of ball cauterization following loop excision and follow-up colposcopy. AB - OBJECTIVE: To investigate whether central diathermy ball cauterization after loop excision affects satisfactory colposcopy at follow-up. METHODS: One hundred one consecutive women with the squamocolumnar junction visible at the ectocervix scheduled for loop excision were assigned alternately into two groups. In group A, diathermy ball cauterization was applied to the entire crater following excision. In group B, cauterization was avoided in a 2-3-mm zone around the new os. The women were re-examined 4 months postoperatively by colposcopy and microcolpohysteroscopy with specific intention to identify the location of the squamocolumnar junction. The examiners performing colposcopy and microcolpohysteroscopy were not aware of each other's interpretation, or of the method of cauterization used. RESULTS: Follow-up colposcopy was satisfactory in 12 women in group A (24%) and 47 women in group B (92.2%) (P <.001). Forty-three women (86%) in group A and ten in group B (19.6%) had the squamocolumnar junction partly or fully located within the cervical canal (P <.001). Microcolpohysteroscopy located the squamocolumnar junction at a mean depth of 4.5 +/- 2.4 mm (+/- standard deviation [SD]) in the women in group A and 1 +/- 0.9 mm in group B (P <.001). Microcolpohysteroscopy could not be performed in 13 women in group A (26%) and one woman in group B (2%) (P <.001). CONCLUSION: Diathermy ball cauterization at the new cervical os after loop excision results in a shift of the squamocolumnar junction toward the endocervical canal, and predisposes to cervical stenosis, thereby decreasing satisfactory colposcopy rates. PMID- 11275039 TI - Randomized placebo-controlled evaluation of intramuscular interferon beta treatment of recurrent human papillomavirus. AB - OBJECTIVE: To evaluate the effectiveness and safety of interferon beta in women with recurrent cervical human papillomavirus (HPV) lesions. METHODS: Women with recurrent HPV of the cervix were assigned randomly to received either 3 million IU of interferon beta daily for 5 days, followed by 2 days of rest for 3 weeks, or placebo on the same schedule (N = 61 in each group). They were evaluated at 6 and 12 months after cytology, colposcopy, and directed punch biopsy. Comparison between groups was carried out by chi(2), Fisher exact test, and Student t test, depending on the variable. Multivariable logistic regression was used to evaluate influence of variables to treatment and categorical and continuous variables were compared by Mantel-Haenszel and Wilcoxon tests. RESULTS: When treatment success rates for all patients at 6 and 12 months were compared, a highly significant statistical difference was found in the treated group compared with the placebo group [48 of 61 (79%) versus 33 of 61 (54%), P =.001, and 43 of 61 (70%) versus 26 of 61 (43%), P =.002, respectively]. Multivariable analysis showed treatment success rates with interferon beta were higher between the group with initial histopathology of cervical intraepithelial neoplasia (CIN) (odds ratio 4.86; 95% confidence interval 1.75, 13.49), and the group receiving placebo (P =.002). Side effects treatments were minimal in 70% of women; the most severe events were headaches and flulike symptoms that did not interfere with the treatment. No clinically significant changes were found in laboratory measurements of glucose or transaminases during treatment or follow-up. CONCLUSIONS: Intramuscular injections of interferon beta were effective for treating recurrent HPV lesions, particularly when associated with CIN. The only side effects were mild and controllable. PMID- 11275040 TI - Galactography and exfoliative cytology in women with abnormal nipple discharge. AB - OBJECTIVE: To evaluate galactography and cytology in women with nipple discharge without clinical or mammographic evidence of cancer. METHODS: During a 12.5-year period, 384 women (15-85 years, mean age 47.5 +/- 14 years) were referred for galactography and smear cytology for recent onset of spontaneous, non-milky nipple discharge. Patients with clinical or mammographic evidence of tumor underwent excisional biopsy directly. Among 314 galactograms, 189 [60.2%; 95% confidence interval (CI) 54.5%, 65.6%] biopsies were recommended. A further 11 patients were scheduled for biopsy because of mammography or cytology. RESULTS: Sixteen of 182 biopsied patients had malignancies (8.8%; CI 5.3%, 14.1%). Combined rate of papillomas, papillomatous proliferation, and malignant tumors was 59.9% (109 of 182; CI 52.4%, 67.0%). Biopsy was malignant in three of 56 women (5%) with nonhemorrhagic discharge and in 13 of 97 (13%) with hemorrhagic discharge (P =.26). Exfoliative cytology revealed 11 false-negatives, four false positives, five true-positives, and 153 true-negatives (sensitivity 31.2%, CI 11%, 58%; specificity 97.4%, CI 93%, 99%). In ten of 158 patients (6.3%) with suspicious galactography, cancer was found by biopsy. Sensitivity of galactography for malignancy was 83% (CI 51.6%, 97.9%) and specificity was 41% (CI 35.2%, 46.5%). Galactographic sensitivity for any (benign or malignant) neoplasm was 94% (93 of 99; CI 87%, 98%) and specificity was 55% (119 of 215; CI 48%, 62%). Half of the cancers were detected exclusively by galactography. CONCLUSION: Cytology is helpful when positive and galactography localizes the source of discharge. Biopsy is indicated when palpation, mammography, cytology, or galactography is suspicious. PMID- 11275219 TI - Involvement of antioxidants and lipid peroxidation in the adaptation of two cool season grasses to localized drought stress. AB - In natural environments, drought often occurs in surface soil while water is available for plant uptake deeper in the soil profile. The objective of the study was to examine the involvement of antioxidant metabolism and lipid peroxidation in the responses of two cool-season grasses to surface soil drying. Kentucky bluegrass (Poa pratensis L) and tall fescue (Festuca arundinacea Schreb.) were grown in split tubes, consisting of two sections (each 10 cm in diameter and 20 cm long). Grasses were subjected to three soil moisture regimes: (a) well-watered control: whole soil profile was watered; (b) surface drying: surface 20 cm of soil was dried by withholding irrigation and the lower 20 cm of soil was watered; (c) full drying: whole soil profile was dried. Surface drying had no effects on relative water content (RWC) and chlorophyll content (Chl) for both grasses and only slightly reduced shoot growth for tall fescue. Superoxide dismutase (SOD) activity increased, while catalase (CAT) and peroxidase (POD) activities remained unchanged during most periods of surface drying. Malondialdehyde (MDA) content was unaffected by surface drying for tall fescue, but increased initially and then decreased to the control level for Kentucky bluegrass. Under full drying, RWC, Chl content, and shoot dry weight decreased, but MDA content increased in both grasses; SOD and POD activities initially increased transiently and then decreased; CAT remained unchanged for 25 days and then decreased. These results suggested that both Kentucky bluegrass and tall fescue were capable of surviving surface soil drying. This capability could be related to increases in antioxidant activities, particularly SOD and CAT. However, full drying suppressed antioxidant activities and induced lipid peroxidation. PMID- 11275041 TI - von Willebrand disease and other inherited bleeding disorders in women with diagnosed menorrhagia. AB - OBJECTIVE: To estimate the prevalence of von Willebrand disease and other bleeding disorders in women with and without diagnosed menorrhagia. METHODS: Women with menorrhagia were identified among members of a health maintenance organization in the southeastern United States through a computer search for appropriate International Classification of Diseases, 9th Revision codes. A random sample of members with no such code was selected as controls. The study included 121 women with menorrhagia and 123 controls. Subjects were interviewed in person, and blood was drawn for coagulation testing. Laboratory results for menorrhagia patients were compared with those in controls using race and blood type specific ranges developed from the control group. A test was considered abnormal if it exceeded two standard deviations below the control mean. RESULTS: Bleeding disorders (von Willebrand disease, factor deficiency, or a platelet abnormality) were diagnosed in 10.7% of menorrhagia patients and 3.2% of controls (P =.02). von Willebrand disease was present in eight menorrhagia patients (6.6%) and in one control (0.8%) (P =.02); separate analyses by race revealed a von Willebrand disease prevalence of 15.9% among white and 1.4% among black menorrhagia patients (P =.01). Women with bleeding disorders did not differ significantly from controls in other symptoms of bleeding. CONCLUSION: The prevalence of inherited bleeding disorders among white women with menorrhagia was substantial, consistent with European data published recently. For unknown reasons, the prevalence of von Willebrand disease was lower among black women. These findings indicate the importance of considering inherited bleeding disorders as a cause of menorrhagia. PMID- 11275220 TI - Morphological and stomatal responses of Norway spruce foliage to irradiance within a canopy depending on shoot age. AB - Morphological and stomatal responses of Norway spruce (Picea abies) foliage to light availability were studied in respect to shoot age. Needle minor diameter (D(1), anatomical width), major diameter (D(2), anatomical thickness), dry weight (M), and tissue density index (I(D)) increased, and needle flatness (Fl) and specific leaf area (SLA) decreased with foliage age, while shade foliage demonstrated higher morphological plasticity as compared to sun foliage. Needle minor diameter, dry weight, and the ratio of total to projected leaf area increased, and needle flatness and specific leaf area decreased with daily average photosynthetic photon flux density (Q(D)). The current-year foliage exhibited the highest variation with irradiance, while the morphological plasticity decreased with needle ageing. The morphological characteristics of needles were independent of irradiance if Q(D) was above 300 umol m(-2) s(-1). D(1) was the only linear needle characteristic which significantly changed with light availability within a canopy, and thus determined needle flatness, SLA, as well as the ratio of total to projected leaf area (TLA/PLA). Needle flatness was a characteristic responding most sensitively to the photosynthetic photon flux density, R(2) was 0.68, 0.44, and 0.49 for the current-year, 1-year-old, and 2 year-old foliage, respectively. TLA/PLA ranged from 2.2 to 4.0 depending on D(1). Variation in SLA in response to light availability can be attributed to changes both in needle shape and tissue density. Stomatal responses to photosynthetic photon flux density (Q(P)) depended on foliage type (sun or shade) and age. Sun needles demonstrated higher daily maximum leaf conductances to water vapour compared to shade needles. The shade needles responded more sensitively to changes in Q(P) at dawn and sunset than the sun needles, while older needles of both foliage types exhibited faster stomatal responses. The light-saturation of leaf conductance (g(L)) was achieved by 20 umol m(-2) s(-1) for shade foliage, and approximately by 50 umol m(-2) s(-1) for sun foliage. As a rule, g(L) changed in response to irradiance faster in the evening, i.e. at decreasing irradiance. Stomata were not usually completely closed in the dark before sunrise and after sunset, the phenomenon being more pronounced in older shoots and sun needles. Nightly water losses from spruce foliage are attributable primarily to older shoots, and are related to age-dependent changes in stomatal responsiveness. PMID- 11275221 TI - Impact of elevated ozone on chlorophyll a fluorescence in field-grown oat (Avena sativa). AB - Oat (Avena sativa) plants were grown in the field near the urban area of Valencia, Eastern Spain. The data on air quality showed that ozone was the main phytotoxic pollutant present in ambient air reaching a 7-h mean of 46 nl l(-1) and a maximum hourly peak of 322 nl l(-1). The effect of ambient ozone on PSII activity was examined by measurements of chlorophyll (Chl) a fluorescence. In leaves with visible symptoms, the function of PSII was changed at high actinic irradiances. Nonphotochemical quenching (NPQ) was higher and quantum efficiency of PSII (Phi(PSII)), photochemical quenching (q(p)), quantum efficiency of excitation capture and PSII electron flow (F(v)'/F(m)') were lower. An enhanced susceptibility to photoinhibition was observed for symptom-exhibiting leaves compared to leaves that remain free of visible symptoms. Both the lowering of photosynthesis efficiency and the increased sensitivity to photoinhibition probably contribute to reduced crop yield in the field, to different extents, depending on growth conditions. To our knowledge, this is the first report that demonstrates that quantum efficiency of exciton trapping in PSII is associated with foliar injury in oat leaves in response to ambient concentration of ozone. PMID- 11275222 TI - Size and longevity of seed banks in Antarctica and the influence of ultraviolet-B radiation on survivorship, growth and pigment concentrations of Colobanthus quitensis seedlings. AB - Populations of Colobanthus quitensis and Deschampsia antarctica, the only two vascular plant species native to Antarctica, are increasing. We performed a seed bank assay to determine the persistence of seeds from intact vegetation/soil cores collected near Palmer Station on the west coast of the Antarctic Peninsula. Vegetation/soil cores were cold stratified at 3 degrees C for >4 years. Subsequent seed bank densities, estimated from seedlings germinated, averaged 847 and 5645 seedlings m(-2) for C. quitensis and D. antarctica, respectively. We also conducted germination trials on C. quitensis seeds collected at our field site and stored for either 120 days or >4 years at 3 degrees C. Germination rates ranged from 6% after 120 days of cold storage to 38% after >4 years of cold storage. These findings show that previous estimates of seed bank densities and germination rates in these species, based on short-term laboratory stratification experiments, may underestimate those found in the field. Stratospheric ozone depletion has lead to increases in ultraviolet-B radiation (UV-B; 280-320 nm) along the Antarctic Peninsula during the austral spring. In a separate experiment we manipulated levels of biologically effective UV-B (UV-B(BE)), over current year C. quitensis seedlings near Palmer Station on the west coast of the Antarctic Peninsula by placing frames over them that either held filters that absorbed most UV-B(BE) ('reduced UV-B(BE)'), transmitted most UV-B(BE) ('near ambient UV-B(BE)') or had no filters ('ambient UV-B(BE)'). We monitored seedling survivorship over the course of the growing season (January-March) and growth and pigment concentrations at the end of the season. There were no UV-B(BE) treatment effects on seedling survivorship over the course of the season and overwinter survivorship averaged 12%. However, seedlings growing under near-ambient and ambient UV-B(BE) had 25 and 48% smaller total leaf areas, 7 and 16% fewer leaves and 65 and 82% fewer branches, respectively, than those growing under reduced UV B(BE). In addition, concentrations of methanol-soluble UV-B-absorbing compounds were 26% higher and concentrations of chlorophyll b were 26% lower in leaves of seedlings growing under ambient UV-B(BE) compared with those under reduced UV B(BE). PMID- 11275223 TI - Relationships between growth and gas exchange characteristics in some salt tolerant amphidiploid Brassica species in relation to their diploid parents. AB - Relationships between growth and different gas exchange characteristics of two amphidiploid salt tolerant species, Brassica napus, and B. carinata with respect to their salt sensitive parents, B. oleracea, and B. nigra were investigated. Twenty three-day old plants of these four species along with those of another amphidiploid moderately salt tolerant B. juncea (developed by hybridization of diploids, B. campestris and B. nigra), and a diploid moderately salt tolerant, B. campestris, were subjected for 28 days to salinized sand culture containing 0, 100 or 200 mol NaCl m(-3) in Hoagland's nutrient solution. The species B. napus and B. carinata produced significantly greater shoot fresh and dry matters than their parents under saline conditions. A close association was found between growth, and assimilation rate for all species differing in degree of salt tolerance. Stomatal conductance (g(s)) was reduced due to salt stress in all species but this variable had no significant correlation with assimilation rate (A). However, the amphidiploid salt tolerant species, B. napus and B. carinata had significantly greater photosynthetic rate, water use efficiency (A/E), intrinsic water use efficiency (A/g(s)) than those of their diploid parents. In conclusion, high salt tolerance of the two amphidiploid species, B. napus and B. carinata was associated with a high assimilation rate, water use efficiency and intrinsic water use efficiency but there was little association of the tolerance of these species with stomatal conductance, leaf water potential or transpiration rate (E). PMID- 11275224 TI - Acclimation of Myrtus communis to contrasting Mediterranean light environments - effects on structure and chemical composition of foliage and plant water relations. AB - Leaf anatomical and chemical characteristics, water relations and stomatal regulation were studied in the shrub Myrtus communis growing under two contrasting Mediterranean light environments (full light versus 30% of full light) during the spring-summer period. These studies aimed to assess plant response to the combined effects of light and water availability. Foliar morphology, anatomy and chemistry composition acclimated positively to light conditions. Leaves of sun-exposed plants were thicker (38.7%) than those of shaded plants, mainly due to increased palisade parenchyma thickness, had a higher nitrogen concentration and stomatal density than the shade ones, which maximized foliar area (>SLA) and Chl/N molar ratio to improve light interception. Chlorophyll concentration per leaf area (Chl(a)) was always higher in sun leaves while, as expressed on dry mass (Chl(m)), significant differences were only apparent in September, shade leaves presenting higher values. During the summer period Chl(a) and Chl(m) markedly declined in sun leaves and remained unchanged in shade ones. The ratio of chlorophyll a/b was not affected either by the light intensity or by the season. Shade leaves presented generally a higher concentration of soluble carbohydrates per dry mass. No significant differences in starch concentration were apparent between sun and shade leaves and a gradual depletion occurred during the water stress period. Maximum stomatal conductances correlated positively with predawn water potential. Throughout the season, sun plants always presented higher leaf conductance to water vapour and lower minimum leaf water potentials, indicating an interaction of light-environment on these water relation parameters. Stomatal closure constitutes a mechanism to cope with diurnal and seasonal water deficits, sun plants presenting a more efficient control of water losses during water deficiency period. In addition, both sun and shade plants evidenced leaf osmotic adjustment ability in response to water stress, which was greater in sun ones. PMID- 11275225 TI - Effects of atmospheric CO(2) enrichment on plant constituents related to animal and human health. AB - Atmospheric CO(2) enrichment is known to significantly enhance the growth and development of nearly all plants, implying a potential for elevated levels of CO(2) to alter the concentrations of plant constituents related to animal and human health. Our review of this subject indicates that increases in the air's CO(2) content typically lead to reductions in the nitrogen and protein concentrations of animal-sustaining forage and human-sustaining cereal grains when soil nitrogen levels are sub-optimal. When plants are supplied with all the nitrogen they can use, however, no such reductions are observed. CO(2)-enriched plants growing in the natural environment also tend to overcome initial reductions in plant mineral concentrations as time progresses, possibly due to development of larger root systems and consequent enhanced abilities to locate and absorb mineral nutrients. Atmospheric CO(2) enrichment additionally appears to reduce oxidative stresses in plants; and it has been shown to increase the concentration of vitamin C in certain fruits and vegetables. Elevated CO(2) has also been demonstrated to increase the biomass of plants grown for medicinal purposes while simultaneously increasing the concentrations of the disease fighting substances produced within them. It is likely, therefore, that the ongoing rise in the air's CO(2) content will continue to increase food production around the world, while maintaining the nutritive quality of that food and enhancing the production of certain disease-inhibiting plant compounds. PMID- 11275226 TI - The evolution of medicine: creative or emergent? PMID- 11275227 TI - Endoscopic retrograde cholangiopancreatography and gallstone pancreatitis. PMID- 11275229 TI - Imaging and staging in the management of rectal cancer. PMID- 11275228 TI - Treating liver metastases: let us count the ways. PMID- 11275230 TI - Medullary thyroid cancer: how is it different? PMID- 11275231 TI - Preoperative evaluation. PMID- 11275233 TI - Breast reconstruction after breast conservation therapy: part 2 of the 6-part series on current concepts in breast reconstruction. PMID- 11275232 TI - Wound care. PMID- 11275234 TI - Hepatic arterial infusion chemotherapy. PMID- 11275235 TI - PET and head and neck cancer. PMID- 11275237 TI - Cardiac trauma. PMID- 11275236 TI - Pancreatic transplantation. PMID- 11275238 TI - Internet and e-mail security: for sale, your vital statistics (part 2). PMID- 11275239 TI - The renaissance surgeon: an educational perspective. PMID- 11275240 TI - Pathophysiology. PMID- 11275241 TI - Clinical significance and approach. PMID- 11275242 TI - Editorial comment. PMID- 11275244 TI - Ulcerative colitis: natural history and medical management. PMID- 11275243 TI - Ischemic colitis: a brief review. PMID- 11275245 TI - Ulcerative colitis: surgical intervention. PMID- 11275246 TI - Gastric trichobezoar as a manifestation of child abuse(1). PMID- 11275247 TI - Utility of computed tomographic enteroclysis for the general surgeon(1). AB - Enteroclysis uses contrast fluid distention of the small bowel through a jejunal catheter with flouroscopic imaging to identify abnormalities. Computed tomograpic enteroclysis (CT-E) adds cross-sectional imaging to identify small bowel pathology to include masses, gastrointestinal bleeding of unknown origin, and partial obstruction. Computed tomography-enteroclysis is being used more frequently in the assessment of patients with possible small bowel pathology. This study examines the applicability of CT-E and its superiority over conventional enteroclysis.A retrospective chart review was used to examine all CT E and enteroclysis studies performed at our institution during a 24-month period (August 1997 to August 1999). All patients that had received CT-E or enteroclysis were divided into 3 categories; group I: small bowel mass, group II: gastrointestinal bleeding, and group III: partial small bowel obstruction (pSBO). All patients included had received other radiological procedures based on the indication for examination to include esophagogastroduodenoscopy, colonoscopy, CT, abdominal x-rays, barium enema, and upper gastrointestinal with small bowel follow-through.Forty-nine studies were performed, with enteroclysis or CT-E, used in 46 patients. Median age was 62 years (M:F, 1:1). In group 1 (n = 10), no masses were noted, but all patients identified as having a mass on previous studies (n = 6) were determined not to have a mass by CT-E (n = 1) and enteroclysis (n = 5). In group II (n = 19), 1 small bowel source (jejujunal arteriovenous malformation) was identified through CT-E, and all other studies in both categories were negative/normal. In group III (n = 20), 5 pSBO were identified through CT-E that had not been previously described.Enteroclysis and CT-E are both effective at disproving the presence of small bowel masses discovered through less-specific radiological methods. In terms of gastrointestinal bleeding, CT-E is as effective as enteroclysis at identifying source of bleeding and may have an added role through its ability to better identify anatomic relationships. Computed tomography-enteroclysis was able to determine the presence of pSBO in 5 patients that previously had been undiagnosed. In conclusion, enteroclysis remains an effective radiological study for examination of the small bowel. Computed tomography-enteroclysis matches that effectiveness with the added benefit on high-resolution anatomic images that serve it well as an additional diagnostic tool for the General Surgeon in patients with difficult to diagnose small bowel pathology. PMID- 11275248 TI - Gallbladder mass in a sexagenarian. PMID- 11275249 TI - Improved reduction in pain in chronic pancreatitis with combined intraoperative celiac axis plexus block and lateral pancreaticojejunostomy. AB - PURPOSE:Severe abdominal pain secondary to chronic pancreatitis is often multifactorial in origin. Lateral pancreaticojejunostomy (LPJ) is currently the accepted surgical treatment of choice when the main pancreatic duct is dilated. Chemical ablation of the celiac plexus for the treatment of intractable pain in chronic pancreatitis has been used without clear benefit. The aim of this study is to compare treatment outcomes of 2 groups of patients with the diagnosis of chronic pancreatitis and intractable abdominal pain (LPJ alone versus LPJ with intraoperative alcohol celiac ablation).Between 1994 and 1997, 34 patients underwent LPJ to control intractable pain secondary to chronic pancreatitis. These patients were divided into 2 groups, group 1 was LPJ only (16 patients) and group 2 was LPJ and intraoperative celiac ablation with 50% absolute alcohol (18 patients). Preoperative diagnosis and treatment criteria were similar for both groups. The clinical characteristics and outcome of both groups were retrospectively analyzed. Fisher exact test was used for statistical analysis.Demographic characteristics were similar in both groups. Pain control at short- and long-term follow-up was significantly improved in group 2 compared with group 1 (p < 0.035).Intraoperative celiac ablation in addition to LPJ appears to have a better response than does LPJ alone. Even though the number of patients is small, these results provide a basis for pursuing a prospective, randomized study to definitively answer this question. PMID- 11275250 TI - Resident and faculty perceptions of a surgical residency program merger. AB - To evaluate resident and faculty perceptions of a residency merger process.Survey of faculty and residents of a recently merged general surgical residency. Nineteen separate program characteristics were evaluated via a numerical scoring system, and additional written commentary regarding dominant perceived benefits and detriments of the merger was solicited. Statistical significance was evaluated on numerically scored items by applying the Mann-Whitney U test to median values expressed with interquartile ranges, comparing resident and faculty responses.Scoring system responses from faculty and residents were generally similar. The merger was seen as neutral to positive in its impact on academic issues, but it had more negative effects on issues related to overall program atmosphere and morale. Statistically significant differences between resident and faculty responses were noted in 2 areas: teaching conference timing and overall program effectiveness in preparing for practice. Both of these areas were more favorably impacted by the merger from the residents' perspective, and more negatively as judged by the faculty (p < 0.05). Written commentary by both groups similarly emphasized areas of academic strengthening as a positive effect of the merger, and relationship and morale issues as being more negatively impacted.As reflected by resident and faculty perceptions, program mergers may provide opportunities to strengthen and enhance the academic and clinical foundation of residency. This may, however, occur at the expense of morale and relational issues, which may be negatively impacted by program administrative and geographic expansion. PMID- 11275251 TI - Risk factors predictive of positive findings at colonoscopy(1). AB - PURPOSE:The surgery department at our institution has become the primary provider of colonoscopy. We sought to determine which risk factors, if any, were most predictive of positive findings on colonoscopy.Between March and December 1999, 202 consecutive patients referred for colonoscopy were identified. Each patient was interviewed and a standard questionnaire completed before colonoscopy to establish possible risk factors for the presence of colorectal cancer or polyps. The colonoscopy findings, including pathology reports, were correlated with the questionnaire and subjected to chi-square analysis to determine statistical significance.The risk factors most likely to be associated with a finding of colorectal cancer or polyp were family history of colorectal cancer (65%), bleeding (65%), fecal occult blood positive (64%), abdominal pain (60%), and alteration of bowel habits (53%).No risk factor by history or presenting symptoms reached statistical significance as an independent predictor of a positive colonoscopy finding. However, most frequently associated with positive colonoscopy findings were a family history of colorectal cancer, bleeding, positive fecal occult blood test, presence of abdominal pain, and alteration of bowel habits. A history to include these risk factors can serve to prioritize the need for a colonoscopic examination. PMID- 11275253 TI - The tortoise and the air. PMID- 11275252 TI - The effects of practice and instruction on speed and accuracy during resident acquisition of simulated laparoscopic skills. AB - To assess the effects of practice and dynamic instruction on changes in speed and accuracy during acquisition of simulated laparoscopic surgical skills.Fourteen PGY-1 general surgery residents were randomly assigned to 1 of 2 experimental conditions (n = 7 per group), either practice only or practice with instruction, and required to perform 10 trials of each of 2 laparoscopic surgical skills cannulation and object passing. Practice only subjects were given verbal instructions for each task, and corrective feedback only after trial 1. Practice with instruction subjects were treated the same, but also saw a videotaped demonstration and received dynamic feedback during and between each trial. Performance speed was recorded for each trial and number of errors was recorded for trials 8 to 10 by videotape review.Mean speed for subjects in both groups increased significantly for both tasks (p < 0.01). Practice with instruction subjects committed significantly fewer errors on object passing (p < 0.04) and were less variable in the number of errors committed during the cannulation task (p < 0.01).Practice, with or without dynamic instruction, results in significant improvement in the speed of performance of simulated laparoscopic surgical skills. The addition of dynamic instruction to simulator-based practice improves the quality and consistency of resident acquisition of laparoscopic surgical skills. PMID- 11275254 TI - Chimeric CD46/DAF molecules reveal a cryptic functional role for SCR1 of DAF in regulating complement activation. AB - Chimeric proteins using membrane cofactor (CD46) and decay accelerating factor (DAF or CD55) were generated to further investigate the functional domains involved in the regulation of human serum complement. Following activation of the classical pathway, the isolated substitution of CD46 SCR III (x3DAF) exhibited a modest regulatory activity comparable to that of CD46. The isolated substitution of CD46 SCR IV (x4DAF), and the combined CD46 SCR III+IV substitutions (x3/4DAF) were essentially as efficient as DAF. No regulation of C3b deposition was observed with the combined CD46 SCR I+II substitutions (x1/2DAF). When tested after activation of the alternative pathway, both the x3DAF and x3/4DAF chimeras failed to regulate C3b deposition, while the x4DAF chimera still displayed some activity. In contrast to that observed following classical pathway activation, the x1/2DAF chimera exhibited a similar efficiency to wild type CD46 and DAF in controlling C3b deposition. Using SCR specific antibodies, the regulatory activity of the x1/2DAF chimera against the alternative pathway was mapped to the first three distal SCR (i.e. DAF 1, DAF 2 and CD46 III). These data demonstrate that several combinations of SCR domains from two related complement regulators can result in functional molecules, and reveal a novel and cryptic functional role for DAF SCR1. PMID- 11275256 TI - Cholinesterase-like catalytic antibodies: reaction with substrates and inhibitors. AB - We have previously described a catalytic monoclonal antibody, raised against acetylcholinesterase (AChE) and capable of hydrolysing acetylthiocholine. Here, we describe two more such antibodies. All three antibodies were raised against the same antigen, human erythrocyte AChE, a commercial product purified using the cholinesterase anionic site inhibitor, tetramethylammonium. IgG was purified on Protein A-Sepharose, and lack of contamination with AChE or butyrylcholinesterase (BChE) was demonstrated on sucrose density gradients and immunoassay of the fractions. The antibodies recognised AchE and were capable of hydrolysing acetylthiocholine and the larger butyrylthiocholine substrate, and were inactivated by phenylmethylsulphonyl fluoride (PMSF), indicating a serine residue in the active site. K(m), K(cat), K(cat)/K(uncat) and K(cat)/K(m) values were obtained for both substrates. The active sites of the antibodies were probed with anti-cholinesterases known to react with the active and anionic sites of acetyl- and BChE, and the peripheral anionic site of AChE. The antibodies were inactivated to varying degrees by the BChE inhibitors iso-OMPA, ethopropazine and tetracaine, indicating a less sterically constrained site than AChE and the lack of an acyl-binding pocket. They were also partially inhibited by the AChE specific inhibitors, BW284c51 and propidium. No peripheral anionic site, as seen in AChE, was observed, shown by the almost complete lack of reaction with fasciculin. All three antibodies appear to have structures resembling the anionic sites of the cholinesterases, seen by their inhibition by quaternary and tricyclic compounds. Further work is required to determine whether the catalytic activity shown by these antibodies is germline-encoded, or is the result of complexation of the antigen with an inhibitor at a peripheral site. PMID- 11275255 TI - The influence of glycosylation on the thermal stability and effector function expression of human IgG1-Fc: properties of a series of truncated glycoforms. AB - Antibodies are multifunctional molecules that following the formation of antibody antigen complexes, may activate mechanisms to effect the clearance and destruction of the antigen (pathogen). The IgG molecule is comprised of three globular protein moieties (2Fab+Fc) linked through a flexible hinge region. While the Fabs bind antigens, the Fc triggers effector mechanisms through interactions with specific ligands, e.g. cellular receptors (FcgammaR), and the C1 component of complement. Glycosylation of IgG-Fc has been shown to be essential for efficient activation of FcgammaR and C1. We report the generation of a series of truncated glycoforms of IgG-Fc, and the analysis of the contribution of the residual oligosaccharide to IgG-Fc function and thermal stability. Differential scanning microcalorimetry has been used to compare the stabilities of the homogeneous glycoforms of IgG1-Fc. The results show that all truncated oligosaccharides confer a degree of functional activity, and thermodynamic stability to the IgG1-Fc, in comparison with deglycosylated IgG1-Fc. The same truncated glycoforms of an intact IgG1 anti-MHC Class II antibody are shown to exhibit differential functional activity for FcgammaRI and C1 ligands, relative to deglycosylated IgG1. The minimal glycoform investigated had a trisaccharide attached to each heavy chain and can be expected to influence protein structure primarily in the proximity of the N-terminal region of the C(H)2 domain, implicated as a binding site for multiple effector ligands. These data provide a thermodynamic rationale for the modulation of antibody effector functions by different glycoforms. PMID- 11275257 TI - Complete structural characterisation of the mammalian and Drosophila TRAF genes: implications for TRAF evolution and the role of RING finger splice variants. AB - The complete murine TRAF2 gene was obtained using a lambda phage and PCR cloning strategy. The gene was found to consist of ten coding and one 5' non-coding exon spread over 28 kbp of DNA. We also report the basic structure of the human TRAF5 and TRAF6 genes obtained by analysis of the genomic DNA database. Comparison of these three gene structures, along with those previously described for TRAF1, TRAF3 and TRAF4, revealed the evolutionary relationship between the six known mammalian TRAFs. The TRAF1/TRAF2 and TRAF3/TRAF5 gene pairs were found to have arisen from recent independent gene duplications and to share a common ancestral gene. Specific TRAF4 and TRAF6 precursor genes were found to have arisen earlier during evolution, with the divergence of the TRAF6 precursor occuring earliest of all. The Drosophila genome was found to contain three TRAF family genes: dTRAF1, dTRAF6 (dTRAF2) and a previously undescribed member we have designated dTRAF3. TRAF-C domain homology indicated that dTRAF3 is likely to have derived from the common precursor for the TRAF 1, 2, 3 and 5 genes, whilst dTRAF1 and dTRAF6 have derived from the TRAF4 and TRAF6 precursor genes, respectively. The implication of these results for the functional evolution of TRAFs is discussed. Analysis is also presented of the conservation of the TRAF2A molecule, a TRAF2 alternate splice isoform with an extended RING finger domain previously described in mice. TRAF2A was not found to be encoded by the human or rat TRAF2 genes and no other murine TRAF gene was found to produce a similar alternate splice product. We also report that the sequence of murine C57BL/6 TRAF4 differs significantly from the published murine TRAF4 sequence, but appears to represent the actual TRAF4 sequence expressed in many mouse strains. PMID- 11275258 TI - Molecular characterization of the rat NK cell receptor 2B4. AB - 2B4 (CD244) is a cell surface glycoprotein of the immunoglobulin superfamily involved in the regulation of natural killer and T lymphocyte function. It is the high affinity counter-receptor for CD48. In mouse and human NK cells, crosslinking of 2B4 with a specific monoclonal antibody or with CD48 can trigger cell-mediated cytotoxicity, IFN-gamma secretion, phosphoinositol turnover and NK cell invasiveness. Recent reports of defective 2B4 signaling and NK cell function in X-linked lymphoproliferative syndrome suggest that this may contribute to the progression of this human disease. Here we describe the molecular characterization of the rat 2B4 gene. The cDNA encodes a protein of 395 amino acid residues that contain two Ig domains in the extracellular region and three unique tyrosine motifs (TxYxxV/I/A) in the cytoplasmic region. The predicted protein has 81 and 68% similarity with mouse 2B4 and human 2B4, respectively. Additionally, it has 94 and 89% similarity at the protein level with the recently reported rat 2B4 related genes, r2B4R-tm and r2B4R-se respectively. Northern blot analysis indicated the presence of multiple transcripts in rat LAK cells and RNK 16 cells. Immunoprecipitation and deglycosylation studies showed that rat 2B4 is glycosylated to similar extent as that of mouse and human 2B4. The cloning of r2B4 in the light of the availability of rat NK cell lines should facilitate in vitro and in vivo experiments to decipher the functional role of 2B4 in NK cell biology. PMID- 11275259 TI - Investigation on interaction of Achatinin, a 9-O-acetyl sialic acid-binding lectin, with lipopolysaccharide in the innate immunity of Achatina fulica snails. AB - Achatinin, a 9-O-acetyl sialic acid (9-O-AcSA) binding lectin, has been demonstrated to be synthesized in amoebocytes of Achatina fulica snails. This lectin was affinity-purified from Achatina amoebocytes lysate (AAL); it appeared as a single band on native polyacrylamide gel electrophoresis (PAGE) and showed 16 identical subunits of M.W. 15 kDa on sodium dodecyl sulphate (SDS)-PAGE. It was found to be homologous with an earlier reported lectin, Achatinin-H, derived from hemolymph of A. fulica snails (Sen, G., Mandal, C., 1995. The specificity of the binding site of Achatinin-H, a sialic-acid binding lectin from Achantia fulica. Carbohydr. Res., 268, 115-125). Homology between both lectins was confirmed by their similar electrophoretic mobilities, carbohydrate specificity and cross reactivity on immunodiffusion. Achatinin showed in vitro calcium dependent binding to two 9-O-acetylated sialoglyoconjugates (9-O-AcSG) on lipopolysaccharide (LPS) (Escherichia coli 055: B5) of M.W. 40 kDa and 27.5 kDa, which was abolished following de-O-acetylation. Based on the previously defined narrow sugar specificity of Achatinin towards 9-O-AcSAalpha2-->6GalNAc [Sen, G., Mandal, C., 1995. The specificity of the binding site of Achatinin-H, a sialic acid binding lectin from Achatina fulica. Carbohydr. Res., 268, 115-125], we conclude that LPS contains this lectinogenic epitope at the terminal sugar moiety. The Achatinin-mediated hemagglutination inhibition of rabbit erythrocytes by LPS further confirmed it. The lectin exhibited bacteriostatic effect on Gram negative bacteria E. coli, DH5alpha and C600. AAL was earlier reported to undergo coagulation in presence of pg level of LPS (Biswas, C., Mandal, C., 1999. The role of amoebocytes in the endotoxin-mediated coagulation in the innate immunity of Achatina fulica snail, Scand. J. Immunol. 49, 131-138). We now demonstrate that Achatinin participates in LPS-mediated coagulation of AAL as indicated by enhanced release of Achatinin from the LPS stimulated amoebocytes and most importantly, by exhibiting a 77% decline in the coagulation of AAL when depleted of Achatinin. Level of Achatinin sharply declined (17-fold) following injection of LPS (20 microg per snail) to the snails, which was reversible by simultaneous injection of LPS and leupeptin implying the presence of LPS-mediated serine protease activity in Achatinin. This was substantiated when purified Achatinin in vitro showed serine protease activity in the presence of LPS followed by its complete blockage in the presence of leupeptin and phenyl methyl sulphonyl fluoride. Therefore, Achatinin, an abundantly available lectin at multiple sites of A. fulica, by virtue of its interaction with LPS, essentially plays a crucial role in the innate immune protection of A. fulica snails. PMID- 11275260 TI - Isolation and structure-functional characterization of phage display library derived mimotopes of noxiustoxin, a neurotoxin of the scorpion Centruroides noxius Hoffmann. AB - Noxiustoxin (NTX) is a short-chain toxin from the venom of the scorpion Centruroides noxius Hoffmann, whose molecular structure and physiological effects have been characterized in detail, whereas the antigenic properties of this and other K(+) channel-blocking toxins are poorly studied. A monoclonal antibody against NTX, BNTX18, able to inhibit the binding of NTX to rat brain synaptosomes, was used in the present study for selecting immunoreactive peptides, mimotopes, from a 12mer and a 7mer phage library. The peptides were characterized immunologically and used for mapping the epitope on NTX. In total, 75 phage clones carrying 43 different peptides were analyzed of which 42 clones carrying 17 different peptides, twelve 12mer and five 7mer peptides, presented a single consensus motif: Leu(Ile, Val)-Tyr(Phe, Trp, Leu)-Gly-Met(Ala). All but three of the peptides containing this motif were reactive with selected mAb BNTX18 in a dot-blot assay of which eight were clearly positive in ELISA and exhibited in competition-inhibition assay the antibody binding specificity of the NTX epitope recognized by BNTX18. The two most reactive mimotopes injected into mice showed the ability to induce antibodies reacting with NTX, thus, to mimic the epitope of NTX antigenically. Sequence comparison and the analysis of the three-dimensional structure of NTX led to the proposal that residues Glu19-Leu20 Tyr21-Gly22 and the hydrophobic part of the side chain of Lys18 form the C terminal part of the epitope. Due to the frequent presence of residues Pro, Leu, Thr, Arg, and Gln in the N-terminal part of the mimotopes, corresponding homologous residues in the N-terminal proximity of the partial epitope may be part of an additional more hydrophilic epitope element. PMID- 11275261 TI - Integration of TCR and IL-4 signals through STAT6 and the regulation of IL-4 gene expression. AB - Signals generated by both antigen and cytokines binding to CD4(+) T cells synergize to promote helper T cell subset differentiation. For Th2 cell commitment neither IL-4 nor T cell receptor (TCR) stimulation are sufficient to drive differentiation. Th2 differentiation requires IL-4-receptor mediated signal transducers and activators of transcription 6 (STAT6) activation, but the possibility that IL-4 can directly enhance IL-4 production by T cells has remained unclear. In this report, IL-4 is shown to increase anti-CD3 or ionomycin induced IL-4 mRNA in differentiated murine Th2 cells. Anti-CD3 or ionomycin also enhances IL-4 induction of STAT6 activation, assayed by gel shift analysis of nuclear lysates. Surprisingly, cyclosporin A (CsA) also enhances STAT6 induction. Both ionomycin and CsA also enhance IL-4-induced transcriptional activity of a STAT6-linked promoter-reporter construct. In vitro calcineurin activity reduces STAT6 interactions with DNA, and CsA or FK506 can block this effect of calcineurin. These results indicate that IL-4 and TCR-generated calcium signals interact to maximize IL-4 gene expression and STAT6 activity. Calcineurin mediated serine dephosphorylation of STAT6 and STAT6 serine phosphorylation may counter-regulate transcriptional activity of STAT6. PMID- 11275262 TI - Double producers of kappa and lambda define a subset of B cells in mouse plasmacytomas. AB - Rearrangement of the light chain locus is believed to be an ordered process in which Iglambda rearrangements only occur if Igkappa rearrangements are found to be non-productive or self-reactive. Secondary rearrangements of the B-cell receptor (BCR) have shown, however, that rescue of abortive Igkappa rearrangements or autoreactive B cells can be achieved through receptor editing using upstream V-regions as the template sequences. Since secondary rearrangement can occur in the periphery, possibly in a subset of B cells maintaining constitutive Rag activity, it is conceivable that two light chains (kappa:kappa or kappa:lambda) could be expressed in these cells, apparently in violation of allelic exclusion. Previously, we have reported that silicone-induced plasmacytomas (SIPCs) exhibit dual expression and ongoing rearrangements of Igkappa and Iglambda. In this paper, we show by ELISA that both Igkappa and Iglambda are found at the protein level, but are secreted in different amounts. Furthermore, we demonstrate by micro-manipulation and RT-PCR amplification that Igkappa and Iglambda are simultaneously expressed in a single SIPC cell. We propose that these dual-expressing cells, found intermittently in cases of plasmacytomas (PCs), may have originally been immature B cells when transformed but now are maintained as a long-lived mature B cell found infrequently in the tumor population. PMID- 11275263 TI - Transcription factor NF-kappaB regulates inducible CD83 gene expression in activated T lymphocytes. AB - The immunoglobulin superfamily member CD83 is expressed on the surface of mature dendritic cells that present processed antigens to T lymphocytes. In addition, T cells acquire CD83 expression following mitogenic stimulation in vitro. Here we report two lines of evidence demonstrating that this inducible lymphocyte response is genetically programmed by transcription factor NF-kappaB and contingent upon proteolytic breakdown of its cytoplasmic inhibitor IkappaBalpha. First, signal-dependent induction of CD83 mRNA expression is blocked in both transformed and primary T cells harboring a degradation-resistant mutant of IkappaBalpha that constitutively represses NF-kappaB. Second, as revealed in gel retardation assays, the IkappaBalpha constitutive repressor prevents the inducible interaction of NF-kappaB with consensus recognition sites identified in the CD83 promoter. Given that IkappaBalpha is functionally coupled to the T-cell antigen receptor, these findings suggest that the downstream transcription unit for CD83 is triggered by NF-kappaB during an adaptive immune response. PMID- 11275264 TI - Unique and shared IgE epitopes of Hev b 1 and Hev b 3 in latex allergy. AB - Of the several latex proteins cloned and expressed, the rubber elongation factor, Hev b 1, and the closely related Hev b 3, represent two major allergens associated with latex allergy. Although both allergens demonstrated IgE binding with sera from latex allergic patients, it was not known whether these two molecules shared any epitopes. Hence, in the present study using health care workers (HCW) and spina bifida (SB) patients with latex allergy, we investigated the IgE binding epitopes in Hev b 1 and Hev b 3. Recombinant Hev b 1 and Hev b 3 were expressed in a prokaryotic expression system, while overlapping decapeptides of Hev b 1 and Hev b 3 were synthesized on derivatized cellulose membrane. Eight IgE binding epitopes for Hev b 1 and eleven for Hev b 3 were identified using sera from latex allergic patients with SB. On further analysis of synthetic peptides encompassing these epitopes, similar IgE antibody reactivity was demonstrated with three Hev b 1 epitopes b1E3, b1E5, b1E6 and two Hev b 3 epitopes; b3E10 and b3E 11. For Hev b 1, a unique IgE binding epitope was identified in the region of amino acid residues 16-25. In competitive ELISA, peptides bIE2 and bIE4 together inhibited 58% of IgE binding of Hev b 1, while b3E5 showed 22% inhibition in the IgE binding of Hev b 3. The results of the present study suggest that the understanding of linear and conformational IgE epitopes in the major latex allergens may provide better insight into the structure-function relationship of the allergens, and may lead to the development of better patient care and management strategies in latex allergy. PMID- 11275265 TI - The role of hepatocytes in the clearance of liposomes from the blood circulation. AB - In this chapter we summarize literature and describe in more detail our own observations over a period of nearly two decennia on the role of hepatocytes in the hepatic clearance of intravenously administered liposomes. Evidence is presented indicating that, although size is an important parameter, it is not decisive in determining access of liposomes to the hepatocytes. Also lipid composition is an important parameter, including charge, rigidity and headgroup composition. The role of the fenestrated sinusoidal endothelial cells in determining liposome accessibility of hepatocytes is discussed as well as the involvement of opsonizing plasma proteins such as apolipoprotein E. Our observations led us to postulate the existence of at least four different mechanisms of interaction of liposomes with hepatocytes, i.e. an endocytic and a non-endocytic one for both neutral and negatively charged vesicles PMID- 11275266 TI - Mammalian phospholipases A(2): mediators of inflammation, proliferation and apoptosis. PMID- 11275267 TI - Plasmalogens: biosynthesis and functions. PMID- 11275268 TI - Cytokine production consequent to T cell--microglia interaction: the PMA/IFN gamma-treated U937 cells display similarities to human microglia. AB - Cognate interactions between human adult microglia and activated T lymphocytes induce the production of inflammatory cytokines. Since this interaction can occur in a non-antigen-dependent manner, it is relevant to a variety of CNS diseases where activated T cells, regardless of specificities, come into contact with microglia; these disorders include multiple sclerosis, trauma, stroke and Alzheimer's disease. A model cell line would facilitate studies of the engagement between T cells and human adult microglia, since the latter are difficult to obtain in substantial quantity or frequency. This study shows that the PMA/IFN gamma-treated U937 cell line shows similarities to microglia in its interaction with activated T lymphocytes, in that the production of tumor necrosis factor (TNF)-alpha, interleukin (IL)-4, IL-10 and IL-12 is induced. Morphological features and mechanisms of cytokine production resemble those observed in microglia--T cell co-cultures since CTLA-4 and CD40--CD40L blockades reduce TNF alpha and IL-10 levels, while anti-CD23 inhibits IL-10 only in U937--T cell interactions. We propose that PMA/IFN gamma-treated U937 cells can serve as a model of human adult microglia to study cytokine generation in response to interactions with activated T cells. PMID- 11275269 TI - Quantitative estimation of 3-D fiber course in gross histological sections of the human brain using polarized light. AB - Series of polarized light images can be used to achieve quantitative estimates of the angles of inclination (z-direction) and direction (in xy-plane) of central nervous fibers in histological sections of the human brain. (1) The corpus callosum of a formalin-fixed human brain was sectioned at different angles of inclination of nerve fibers and at different thicknesses of the samples. The minimum, and maximum intensities, and their differences revealed a linear relationship to the angle of inclination of fibers. It was demonstrated that sections with a thickness of 80--120 microm are best suited for estimating the angle of inclination. (2) Afterwards the optic tracts of eight formalin-fixed human brains were sliced at different angles of fiber inclination at 100 microm. Measurements of intensity in 30 pixels in each section were used to calculate a linear function of calibration. The maximum intensities and the differences between maximum and minimum values measured with two polars only were best suited for estimation of fiber inclination. (3) Gross histological brain slices of formalin-fixed human brains were digitized under azimuths from 0 to 80 degrees using two polars only. These sequences were used to estimate the inclination of fibers (in z-direction). The same slices were digitized under azimuths from 0 to 160 degrees in steps of 20 degrees using a quarter wave plate additionally. These sequences were used to estimate the direction of the fibers in xy-direction. The method can be used to produce maps of fiber orientation in gross histological sections of the human brain similar to the fiber orientation maps derived by diffusion weighted magnetic resonance imaging. PMID- 11275270 TI - Time course and effective spread of lidocaine and tetrodotoxin delivered via microdialysis: an electrophysiological study in cerebral cortex. AB - Microdialysis is a useful tool for administering drugs into localized regions of brain tissue, but the diffusion of drugs from the probe has not been systematically examined. Lidocaine (10%) and tetrodotoxin (TTX, 10 microM), drugs typically used in neural inactivation studies, were infused through a microdialysis probe into raccoon somatosensory cortex while evoked responses were recorded at four electrodes equally spaced 0.5--2.0 mm from the probe. The decreases in evoked response amplitude as a function of time and distance from the probe were used as functional measures to describe the time course and spread of the drugs. TTX inactivated distant sites more quickly and to a greater extent than lidocaine. Responses recovered within approximately 40 min after termination of lidocaine, but did not recover for at least 2 h after TTX. Based on these measurements, we estimated that, at the concentrations used, lidocaine has a maximal spread of 2.1 mm, while TTX could spread as far as 4.8 mm from the microdialysis probe. However, in terms of significant inactivation of neuronal activity, lidocaine and TTX have an effective spread of 1 and 2 mm, respectively. PMID- 11275271 TI - Instantaneous multivariate EEG coherence analysis by means of adaptive high dimensional autoregressive models. AB - This study presents an efficient algorithm for the fitting of multivariate autoregressive models (MVAR) with time-dependent parameters to multidimensional signals. Thereby, the dimension of the model may be chosen to equal the number of signal channels. The autoregressive (AR) parameter matrices are estimated by an extension of the recursive least squares (RLS) algorithm with forgetting factor. The estimation procedure includes a single trial as well as an ensemble mean approach. The latter approach allows the simultaneous fit of one mean MVAR model to a set of single trials, each of them representing the measurement of the same task. A particular advantage of this ensemble mean approach is that it requires only a low computation effort in comparison to well known procedures applied to single trials. Furthermore, the ensemble mean approach is linked with a high adaptation capability. The properties of the estimator are investigated using simulated time series. It can be demonstrated that the adaptation capability of the estimation (measured by its adaptation speed and variance) does not depend on the model dimension. The mean MVAR fit is applied to 19-dimensional EEG data, recorded during an elementary comparison procedure. The calculation of ordinary and multiple coherence is discussed. The sensitivity of the multiple instantaneous EEG coherence will be demonstrated. PMID- 11275272 TI - A robust GTP-induced shift in alpha(2)-adrenoceptor agonist affinity in tissue sections from rat brain. AB - A method is presented for monitoring the coupling of the alpha(2)-adrenoceptor, as well as other receptors, to their G proteins using the GTP-induced shift in agonist affinity states. In tissue sections GTP, but not ATP, induces a robust decrease in agonist affinity of greater than 100-fold, which is much larger than previously found in membrane preparations. A sensitive and easy procedure to monitor the extent of coupling is to compare the amount of [(3)H]RX821002 binding remaining in the presence of 100 nM brimonidine in the absence and presence of 100 microM GTP. This method should be especially applicable for determining the extent of coupling of receptors to their G proteins in multiple brain regions using autoradiographic procedures. PMID- 11275273 TI - An in vivo eyecup preparation for the rat. AB - A method for the preparation of an in vivo eyecup and a complex stimulating sampling device are described; these are suitable for long-term parallel neurochemical and electrophysiological experiments on the rat retina without any additives into the eyecup. In this in vivo eyecup the extracellular microenvironment is under the normal homeostatic control of the vascular system; no continuous exchange of the eyecup fluid and no addition of glutamate is necessary to maintain stable retinal electric responses and amino acid concentrations. The eyecup viability was tested by monitoring the electroretinogram (ERG) and the amino acid contents of the eyecup fluid sampled from the preretinal space by means of microdialysis. After the initial increase the b-wave of the ERG changed by less than 10% in maximal amplitude during experiments lasting 5 h. The glutamate, glutamine, and glycine levels proved comparatively, whereas the taurine level rose continuously throughout the experimental protocol. Recovery of ERG was achieved following exposure to bright background illumination. Total exchange of the eyecup volume requires 20 min at a flow rate of 1 microl/min. The effect of L-AP4 on the ERG was successfully reproduced, which suggests the applicability of this in vivo eyecup for pharmacological experiments on the rat retina. PMID- 11275274 TI - A two-compartment in vitro model for studies of modulation of nociceptive transmission. AB - Here we present a two-compartment in vitro model in which embryonic rat dorsal root ganglia (DRG) neurons are cultured separately from their target dorsal horn neurons. Although separated, synaptic contact can be established between the peripheral and central neurons since the system allows the DRG axons to project into the other compartment, which contains a network of dorsal horn neurons. The efficacy of the model was evaluated by immunocytochemical, calcium imaging and electrophysiological experiments. The results showed that a subpopulation of the DRG neurons had nociceptor characteristics and that these made synaptic contact with the dorsal horn network. Application of current pulses, according to the stimulus paradigm used, evoked action potentials in DRG axons selectively. This in turn gave rise to increased postsynaptic activity in the network of dorsal horn neurons. The model offers a high degree of efficiency since large numbers of DRG axons can be stimulated simultaneously, thus permitting recording of strong output responses from the dorsal horn neurons. This in vitro model provides a means for studying the mechanisms by which modulatory factors, such as immunoregulatory molecules, applied at either the PNS or the CNS level, can affect synaptic activity and nociceptive transmission in single neurons or network of neurons in the dorsal horn. PMID- 11275275 TI - Sleep estimation from wrist movement quantified by different actigraphic modalities. AB - Progress in transducer design and empirical characterization of wrist movement has led to diverse wrist activity monitors, each with its unique features and modality of operation. This study compared sleep--wake estimates from nocturnal wrist activity quantified by different motion-quantifying algorithms. Healthy young adults wore an Actillume and a Mini Motionlogger on the same wrist while nocturnal polysomnography data were recorded simultaneously in the laboratory. Activity data were analyzed with ACTION3 using scoring algorithms independently calibrated for each measurement modality. Overall, each modality yielded accurate and reliable sleep estimates relative to polysomnographic estimates (agreement rates: 91.4--96.5%, correlations for sleep duration: 0.79--0.94). Estimates derived from Actillume modalities were comparable to those of Mini Motionloggers, suggesting that the transducers of these two devices performed comparably for monitoring sleep and wakefulness. Wrist movement quantified by the Mini Motionlogger proportional-integrating mode yielded the best accuracy for detection of sleep--wake states. PMID- 11275276 TI - The quantitative distribution of a putative PKC epsilon mRNA in Limulus central nervous system by modified competitive RT-PCR. AB - Recently, a full length cDNA for the epsilon (epsilon) isoform of protein kinase C (PKC) was cloned and sequenced from a cDNA library for the horseshoe crab, Limulus polyphemus. This multifunctional enzyme has been implicated in the modulation of the choline cotransporter in Limulus and the epsilon isoform has been identified in homogenates from its central nervous system (CNS). RT-PCR has proven to be a very useful method for quantifying even a few molecules of mRNA in tissue samples. A modified competitive RT-PCR was used here to quantify a putative PKC epsilon mRNA in Limulus CNS preparations. First, we replaced normally used oligo dT and random primers generated from mRNA with a PKC epsilon specific (3' end) primer P4. Then we used modified nucleotides to extend sample life in storage and finally, we used only annealing and denaturing temperatures during PCR to reduce background. The modified method was used for the first time to quantify PKC epsilon mRNA from three distinct areas of the CNS in Limulus. Results revealed high levels of PKC epsilon mRNA in the corpora pedunculata, in the abdominal ganglia and in the brain ring. These results indicate that PKC epsilon mRNA is broadly distributed throughout the Limulus CNS. Importantly, this modified competitive RT-PCR technique was successfully applied to the quantitation of specific mRNA from Limulus nervous tissue for which no internal standard is available commercially. PMID- 11275277 TI - A method for standardizing jaw displacements in the horizontal plane while recording single motor unit activity in the human lateral pterygoid muscle. AB - The normal function of the lateral pterygoid muscle is not well understood although this muscle is thought to play an important role in the control of jaw and jaw-joint function and is implicated in temporomandibular disorders (TMD). The lack of a validated method for standardization of jaw movement in studies of lateral pterygoid function has contributed to the lack of understanding of the normal function of this muscle. An improved understanding of normal function will allow valid comparisons to be made with TMD patients in order to identify whether purported differences in activity actually exist. This paper describes a methodology for standardizing command jaw movements in the horizontal plane, together with reliable recordings of single-motor-unit (SMU) activity. In six human participants, jaw movements were standardized by having participants track a linear bank of light-emitting diodes (LEDs) aligned on a monitor displaying the mid-incisor point (MIPT). In all participants, the MIPT target (i.e. an illuminated LED) could be tracked, according to a pre-determined criterion, during single- and multiple-step displacements at different rates (1.3--6.5 mm/s at MIPT) and magnitudes (0.65--12 mm) of movement. SMU activity from the superior (SHLP) or inferior (IHLP) head of the lateral pterygoid muscle could be reliably discriminated during repeated trials of these defined tasks. This methodology establishes a reliable technique for characterizing the firing properties of SMUs within the lateral pterygoid, and has implications for analogous studies in other jaw muscles. PMID- 11275278 TI - Analysis of extracellular gamma-aminobutyric acid, glutamate and aspartate in cat visual cortex by in vivo microdialysis and capillary electrophoresis-laser induced fluorescence detection. AB - To investigate the influence of a partial sensory deprivation on the extracellular concentration of amino acid neurotransmitters in cat visual cortex, a capillary electrophoresis method was developed for the quantification of gamma aminobutyric acid (GABA), glutamate (Glu) and aspartate (Asp) in in vivo microdialysis samples of cat brain. Microdialysis samples from different regions of area 17 were obtained every 15-min using CMA 12 2-mm probes perfused with synthetic cerebrospinal fluid and derivatized using fluorescein isothiocyanate (FITC). Laser-induced fluorescence (LIF) detection was employed. Good selectivity was obtained with a borate buffer (20 mM, pH 10.25). The whole procedure, including the washing step takes only 15 min. The conditions for derivatization and separation were optimized. The parameters for validation such as linearity, precision and detection limit are also reported. The results are consistent with those of HPLC but, as the sample volumes needed are only 1--5 nl, a much better time resolution can be obtained. PMID- 11275279 TI - Crossed unilateral lesions of medial forebrain bundle and either inferior temporal or frontal cortex impair object recognition memory in Rhesus monkeys. AB - In monkeys, section of the fornix, amygdala and anterior temporal stem results in a severe anterograde amnesia. Immunolesions of the cholinergic cells of the basal forebrain suggest that this amnesia is a result of isolating the inferior temporal cortex and medial temporal lobe from their cholinergic basal forebrain afferents. In this experiment, six monkeys were trained in a delayed match-to sample task and then received a section of the medial forebrain bundle in one hemisphere and an ablation of either the frontal or inferior temporal cortex in the opposite hemisphere. All the animals were severely impaired in the performance of this task following this surgery, and the severity of the impairment was independent of the cortical area from which the medial forebrain bundle was disconnected. These results support a model of fronto-temporal interaction via the basal forebrain in new learning, in which midbrain sites related to reward modulate the cholinergic basal forebrain activity. PMID- 11275280 TI - Enrichment-dependent differences in novelty exploration in rats can be explained by habituation. AB - In rats, exploratory activity and emotional reactivity towards novel stimuli reflect independent biological functions that are modulated differently by rearing experiences. Environmental enrichment is known to improve performance in exploratory tasks, while having inconsistent effects on emotionality. This study examined the effect of environmental enrichment on the behaviour of rats in two exploratory tasks. Male rats were reared under one of four conditions, differing in social and non-social complexity. At 9 weeks of age, exploration of a novel open field, and exploration of novel objects in the same open field following 24 h habituation, was assessed. Differences in social and non-social complexity of the rearing environment had inconsistent effects on exploration in the novel open field. In contrast, when rats were faced with novel objects in an otherwise familiar environment, exploration habituated faster with increasing stimulus complexity of the non-social environment. The social environment had no effect on this latter test. These findings indicate that environmental enrichment affects exploratory activity primarily through its effect on habituation to novelty. This effect depends on relative stimulus complexity of the rearing environment, but is independent of social factors. The present results further suggest that aversive tasks can obscure the expression of enrichment-dependent differences in habituation to novelty. PMID- 11275281 TI - Role of 5-HT(1A) and 5-HT(7) receptors in the facilitatory response induced by 8 OH-DPAT on learning consolidation. AB - The present study further explored the mechanisms involved in the facilitatory effect induced by (+/-)-8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) on learning consolidation. For this purpose, we analyzed in parallel the effects of LY215840 and ritanserin, two 5-HT(2) receptor antagonists with high affinity for the 5-HT(7) receptor, and WAY100635, a selective 5-HT(1A) receptor antagonist, on the facilitatory effect induced by 8-OH-DPAT on learning consolidation. We also determined whether LY215840 and/or ritanserin could be beneficial in restoring a deficient learning condition. Using the model of autoshaping task, post-training injection of LY215840 or WAY100635 had no effect on learning consolidation. However, both drugs abolished the enhancing effect of 8-OH-DPAT, with LY215840 being slightly more effective than WAY100635 in this respect. Ritanserin produced an increase in performance by itself and also abolished the effect of 8-OH-DPAT. Remarkably, selective blockade of 5-HT(2A) and 5-HT(2B/2C) receptors with MDL100907 and SB200646, respectively, failed to alter the 8-OH-DPAT effect. LY215840 and ritanserin, at the doses that inhibited the 8-OH-DPAT-induced response, reversed the learning deficits induced by scopolamine and dizocilpine. The present results suggest that the enhancing effect produced by 8-OH-DPAT on learning consolidation involves activation of 5-HT(1A) receptors and an additional mechanism, probably related to the 5-HT(7) receptor. Blockade of 5 HT(2) receptors, and perhaps of 5-HT(7) receptors as well, may provide some benefit in reversing learning deficits associated with decreased cholinergic and/or glutamatergic neurotransmission. PMID- 11275282 TI - Involvement of adrenal medulla grafts in the open field behavior. AB - Immunohistochemical and behavioral techniques were used to study the effects of adrenal medulla grafts, implanted in striatum after bilateral kainic acid (KA) lesions of this structure, on the open field behavior of mice. KA-induced behavioral changes in leaning, grooming and locomotor activity of the open field test were significantly improved after grafting of the adrenal medulla, and in some respects, fully restored. Immunohistochemical identification showed that grafts contained neuron-like cells with a tyrosine hydroxylase (TH), phenylethanolamine N-methyltransferase, gamma-aminobutyric acid (GABA), choline acetyltransferase (ChAT), and enkephalin-like immunostainings. A likely interpretation of this complex pattern of results is that adrenal medullary grafts may restore the deficits of GABAergic neurons which in turn reverse the abnormalities in emotionality and locomotion. Neurobiologically, these behavioral improvements probably involve GABAergic and catecholaminergic factors of adrenal medulla grafts, although other neuroactive substances, such as acetylcholine and enkephalins, cannot be excluded. PMID- 11275283 TI - Effects of cholinergic manipulation on operant delayed non-matching to position performance in two inbred strains of mice. AB - With the increasing demand on phenotyping of mouse mutants there is a clear need to develop novel paradigms for testing mice. Mice are able to learn a non matching to position rule to high accuracy in a variety of maze paradigms, but an operant version of this task is desirable. In the present study, mice of the C57BL/6 and DBA/2 strains were trained and tested on an operant delayed non matching to position (DNMTP) paradigm. Data were analysed according to the methods of signal detection theory (SDT), which allows conclusions as to whether strain differences in DNMTP performance are more related to changes in accuracy or in motivational factors. Mice can learn to respond on an operant DNMTP paradigm with high accuracy, and accurate performance depends on the duration of the delay-period, i.e. forgetting curves can be generated. Comparison between the two strains of mice revealed that DBA/2 mice learned faster than C57BL/6 mice to associate the lever press with food during initial shaping, but no further strain differences were observed in accurate responding during later stages of the experiment. However, differences in biased responding and, in particular, responsivity were observed between the two strains. Muscarinic blockade with scopolamine (0.1--1.0 mg/kg) failed to affect accuracy in the two strains, but altered responsivity. This task should be of great value for a more in-depth analysis of cognitive function in mutant mice as it allows a better dissociation between mnemonic and non-mnemonic factors. In particular, such paradigm may be of interest for testing conditional mutants, which allow time-sensitive induction or inhibition of gene expression, i.e. where animals can be trained while non impaired to stable baseline and then tested once the gene is activated or inhibited. PMID- 11275284 TI - Increased sensitization of acoustic startle response in spasmodic mice with a mutation of the glycine receptor alpha1-subunit gene. AB - The spontaneous mutant mouse spasmodic (spd) carries a missense mutation affecting the glycine receptor alpha1-subunit gene. This results in a decreased binding affinity to glycine. Spd mutants show exaggerated acoustic startle responses (ASR). The present study sought to elucidate whether this increased ASR is due to a changed auditory processing or to stronger motor output resulting from a disinhibited motor system or, alternatively, to changes in modulatory influences on the startle pathway, namely in the mechanisms underlying habituation and sensitization. We found that in homozygous spd/spd mutants the startle threshold was lower, and the recorded slope of input/output (i/o) function, which reflects the relation between sensory input and motor output, was steeper. During repetitive presentation of high sound pressure level (SPL) startle stimuli (25 dB above startle threshold), ASR amplitudes did not decrease in spd/spd mutants as they do in the wildtype. In contrast, ASR amplitudes decreased when low SPL startle stimuli were presented. Footshocks presented after high SPL startle stimuli did not cause a further increase in ASR amplitudes of spd/spd mutants as in the wildtype. In heterozygous spd/+ mutants all these parameters were between those of spd/spd mutants and wildtype mice but closer to those of the wildtype. The steeper slope of i/o function in spd/spd mutants may be caused by both an increased sensory input and an increased motor output. The altered course of ASR amplitudes during repetitive stimulation and the deficit in additional footshock sensitization, however, can only be explained by an increased sensitization level in the spd/spd mutants. In accordance with the "dual process theory" strong sensitization evoked by high SPL startle stimuli supposedly counteracts habituation, leading to a constant high ASR amplitude. Furthermore, additional footshock sensitization is prevented. The increased sensitization level may be due to a change in auditory processing leading to a stronger sensitizing effect of the startle stimuli with high SPL. Alternatively, glycinergic tonic inhibition of sensitizing structures (e.g. the amygdala) in the wildtype may be diminished in spd/spd mutants, thus leading to a high sensitization level. PMID- 11275285 TI - Spatio-temporal constraints for auditory--visual integration. AB - The perceptual coherence of auditory and visual information is achieved by integrative brain processes. Specialized single neurons with spatial and temporal interactions of auditory and visual stimuli have been demonstrated by several neurophysiological studies. The present, psychophysical, study investigates possible perceptual correlates of these neuronal features. Subjects had to indicate the point of subjective spatial alignment (PSSA) for a horizontally moving visual stimulus that crossed the position of a stationary sound source. Auditory and visual stimuli consisted of periodic pulses that were systematically varied in their phase relationship or repetition rate. PSSAs obtained for continuous visual stimuli served as a reference. When sound and light pulses were coincident in phase at a repetition rate of 2 Hz, PSSAs were shifted by approximately 3 degrees in a direction opposite to the movement of the visual stimulus (with respect to the reference condition). This shift markedly decreased when the temporal disparity exceeded approximately 100 ms and disappeared near phase opposition (250 ms disparity). With 4 Hz repetition rate (temporal disparity < or =125 ms), there was no significant effect of phase relationship on PSSAs, but still an approximately constant shift with respect to the reference value. Variation of the repetition rate resulted in almost constant shifts in PSSA of approximately 3 degrees between 1 and 4 Hz and a linear decrease (slope 0.27 degrees /Hz) with higher repetition rates. These results suggest a spatio temporal 'window' for auditory-visual integration, that extends over approximately 100 ms and approximately 3 degrees : when auditory and visual stimuli are within this window, they are always perceived as spatially coincident. These psychophysical findings may be related to properties of bimodal neurons such as have been demonstrated by neurophysiological recordings in midbrain and cortex. PMID- 11275286 TI - Apamin improves reference memory but not procedural memory in rats by blocking small conductance Ca(2+)-activated K(+) channels in an olfactory discrimination task. AB - Apamin blocks SK channels responsible for long-lasting hyperpolarization following the action potential. Using an olfactory associative task, the effect of an intracerebroventricular 0.3 ng apamin injection was tested on learning and memory. Apamin did not modify the learning of the procedure side of the task or the learning of the odor-reward association. To test reference memory specifically, the rats were trained on a new odor-association problem using the same procedure (acquisition session), and they were tested for retention 24 h later. Apamin injected before or after the acquisition session improved retention of the valence of a new odor pair. Apamin injected before the retention session did not affect the retrieval of the new valence. Thus, the results indicate that the blockage of apamin-sensitive SK channels facilitate consolidation on new-odor reward association. PMID- 11275287 TI - Serotonin reverses dominant social status. AB - Social stress from aggressive interaction is expressed differently in specific brain regions of dominant and subordinate male Anolis carolinensis. Prior to aggressive behavior, the outcome is predictable via the celerity of postorbital coloration: Dominant males exhibit more rapid eyespot darkening. Serotonergic activation is manifest rapidly (1 h) in hippocampus, nucleus accumbens and brainstem of subordinate males, and is expressed more rapidly in dominant males. Amygdalar serotonergic activation responds rapidly (1 h) in dominant males, but is expressed slowly (1 w) and chronically in subordinate males. We hypothesized that chronic (1 w) serotonin elevation, manipulated by the selective serotonin reuptake inhibitor sertraline, would decrease aggressiveness and result in subordinate status. Dominant status was established in pairs of male A. carolinensis. The pairs were separated and treated with sertraline or vehicle. Sertraline was given in food to either the dominant or the subordinate male, both males or neither male for 1 week. Pairs were reintroduced, and behavior and social status recorded. When both dominant and subordinate males were treated with sertraline (or vehicle), or when subordinate males alone were treated with sertraline, previously established social relationships remained unchanged or became associative. However, when dominant males alone were treated with sertraline, their social status was reversed (43%) or negated (57%). Latency to eyespot darkening was significantly retarded in dominant males treated with sertraline, and aggressive displays and attacks were reduced. Chronic 5-HT elevation is consistent with subordinate status. Social status and aggressive disposition do not appear to be immutable, but may be changed by neuroendocrine mechanisms that mediate adaptation to environmental conditions like stress. PMID- 11275288 TI - Rats' processing of visual scenes: effects of lesions to fornix, anterior thalamus, mamillary nuclei or the retrohippocampal region. AB - We analysed the effects of lesions of hippocampal-diencephalic projections -- fornix (FX) mamillary bodies (MB) and anterior thalamic nuclei (AT) -- and retrohippocampal (RH) lesions including entorhinal cortex and ventral subiculum, upon scene processing. All lesions except FX were neurotoxic. Rats learned to discriminate among computer-generated visual displays ("scenes") each comprising three different shapes ("objects"). The paradigm was constant-negative; one constant scene (unrewarded) appeared on every trial together with a trial-unique variable scene (rewarded). Four types of variable scene were intermingled: (1) unfamiliar objects in different positions from those of the constant (type O+P), (2) unfamiliar objects in same positions as in the constant (type O), (3) same objects as the constant in different positions (type P), (4) same objects and positions as the constant but recombined (type X). Group RH performed like controls while groups FX, AT and MB showed (surprisingly) enhanced performance on types X and O. One explanation is that normal rats attempt to process all objects in a scene concurrently, while hippocampal-projection lesions disrupt this tendency, producing a narrower attention, which paradoxically aids performance with some variable types. The results confirm that the entorhinal cortex has a different function from other components of the hippocampal system. PMID- 11275289 TI - Role of the striatum and nucleus accumbens in paced copulatory behavior in the female rat. AB - Female rats engage in a series of approach and avoidance behaviors (pacing behavior) directed at the male in order to achieve a preferred rate of intromissions that make pregnancy more likely to occur with insemination. The striatum and nucleus accumbens have been implicated in the modulation of pacing behavior. It is unclear, however, whether these areas of the brain are necessary for the display of pacing behavior. To address this question, ovariectomized female rats received either bilateral quinolinic acid lesions of the striatum or nucleus accumbens or sham surgeries. After hormone priming, rats were allowed to engage in mating behaviors in an apparatus in which they could pace the rate of the copulatory bout. There was a significant reduction in pacing efficiency after striatal lesions, in that females were less likely to leave the male's side of the chamber after a contact. Animals with lesions of the nucleus accumbens that included the shell were more likely to avoid sexual interaction altogether than animals with control lesions. Therefore, it is concluded that the striatum and nucleus accumbens modulate specific aspects of pacing behavior in the female rat. PMID- 11275290 TI - Opioidergic contribution to conditioned place preference induced by corn oil in mice. AB - We previously reported that voluntary intake of corn oil in the light box showed place preference in the conditioned place preference (CPP) test in mice. In the present study, we investigated the contribution of opioidergic systems to the corn oil-induced CPP in mice. Acquisition of the place preference by corn oil intake was blocked by i.p. injections of an opioid mu antagonist, naloxone (0.1 and 0.3 mg/kg), and delta antagonists, 7-benzylidenenaltrexone (0.5 mg/kg) and naltriben (0.5 mg/kg) 15 min before conditioning. The opioid kappa agonist U 50488H (1 and 3 mg/kg i.p.) also blocked corn oil-induced CPP. Naloxone (1 mg/kg, i.p.) and naltriben (0.5 mg/kg, i.p.) did not affect corn oil intake in the home cage. However, 7-benzylidenenaltrexone (0.5 mg/kg, i.p.) and U-50488H (1 mg/kg i.p.) decreased and increased the corn oil intake, respectively. These results suggested that the rewarding effects of corn oil in the CPP test are at least partially mediated via opioidergic systems through mu and delta receptors. Further, we showed that an opioid kappa agonist reduced the rewarding effects of corn oil in the CPP test in mice, although it increased corn oil intake. PMID- 11275291 TI - Nicotine-induced behavioral sensitization is associated with extracellular dopamine release and expression of c-Fos in the striatum and nucleus accumbens of the rat. AB - It is well known that repeated injections of nicotine produce progressively larger increases in locomotor activity, an effect referred to as behavioral sensitization. This study was carried out to investigate the neural mechanisms underlying nicotine-induced behavioral sensitization using in vivo microdialysis and Fos-like immunohistochemistry (FLI). Rats were given repeated injections of saline or nicotine (0.4 mg/kg s.c., twice daily for 7 days) followed by one challenge injection on the 4th day after the last daily injection. Systemic challenge with nicotine produced a much larger increase in locomotor activity in nicotine-pretreated rats (659.1+/-94.9 counts/2 h) than in saline-pretreated rats (218.1+/-61 counts/2 h). A direct local challenge of nicotine (1 or 5 mM) via a microdialysis probe in the nucleus accumbens or striatum induced a much greater dose-dependent increase of dopamine (DA) output in nicotine-pretreated rats than in saline-pretreated rats. Furthermore, in parallel with the behavioral and biochemical data, systemic challenge with nicotine produced marked Fos-like immunohistochemistry in the nucleus accumbens and the striatum in the nicotine pretreated rats. Taken together, this study demonstrates that behavioral sensitization is clearly associated with an increase in DA release and activation of Fos-like immunoreactive cells in the striatum and the nucleus accumbens produced by repeated nicotine treatment. Our results strongly suggest that the striatum and the nucleus accumbens may play a major role in nicotine-induced behavioral sensitization. The present results are discussed in terms of the development and expression of nicotine-induced behavioral sensitization. PMID- 11275292 TI - Context-specific interference on reversal learning of a stimulus-response habit. AB - Learning occurs in a particular place and time. In most learning situations, information about the training context is encoded along with the task demands and solution. However, the extent to which context contributes to the acquisition and expression of a particular learned response is unclear. In the present paper we examined two fundamental issues underlying the importance of context information and its role in expression of discrimination learning and reversal learning. Rats were trained on a stimulus-response (S-R) habit task designed for the eight-arm radial maze and after reaching a set criterion different context manipulations were performed. Results from Section 2.2.1 revealed that although rats detected a change in context, the learning was not context specific. Results from Section 2.2.2 showed that S-R reversal learning was enhanced when animals were reversed in a context that was different from the one used during original training. Animals that were reversed in a different context showed a renewal effect to the initial S-R when brought back to the original training context. PMID- 11275293 TI - Early morning melatonin administration impairs psychomotor vigilance. AB - The acute soporific effect of melatonin in humans has been demonstrated in a range of studies. How alertness and performance are changed after melatonin given in the morning is not yet known. In a double-blind, placebo-controlled study of nine healthy young men, melatonin was given at 0700 h under controlled conditions of a modified constant routine protocol lasting 56 h (2 days, 3 nights with sleep). A clear decrement in neurobehavioral functions as measured by the Psychomotor Vigilance Test lasted for 6 h after melatonin administration (particularly in the lapse domain and median of the reaction time) without any effect on a letter cancellation task. A subjective soporific effect was present but less pronounced. Thus, melatonin taken in the morning requires caution in situations where high attention is needed. PMID- 11275294 TI - Dose- and time-dependent scopolamine-induced recovery of an inhibitory avoidance response after its extinction in rats. AB - The present investigation was aimed at elucidating the dose and time dependency of scopolamine-induced recovery of inhibitory avoidance after its extinction. Two experiments were conducted: in the first, we analyzed the effects of four doses (1, 2, 4, and 8 mg/kg) of the musacrinic receptor antagonist scopolamine, on the expression of this conditioned response once it had been extinguished. Independent groups of rats were trained in a one-trial, step-through inhibitory avoidance task and submitted to daily retention (extinction) tests. After extinction had occurred, animals were injected intraperitoneally 10 min before retention testing, either with saline or scopolamine. Results show that scopolamine produced a dose-dependent recovery of the avoidance response. The second experiment was carried out in the same animals, which were now tested for retention of inhibitory avoidance at 1, 2, 3, 6, and 9 months after completion of the first experiment. All rats received counterbalanced injections of saline or scopolamine 10 min before testing at each time interval. Reliable recovery of the avoidance response was observed at the 1-month interval with a clear dose dependency while, after the second month, only the groups treated with the two higher doses continued responding. The results indicate that recovery of the extinguished response produced by muscarinic blockade follows dose- and time dependent curves, and can be achieved long after a single training session. These data suggest that the inhibitory avoidance memory trace is retained in the brain after behavioural extinction of this response, thus supporting the view of extinction as new learning that affects the retrieval of the original memory, but does not modify its storage. PMID- 11275295 TI - The selective 5-HT(1A) receptor antagonist p-MPPI antagonizes sleep--waking and behavioural effects of 8-OH-DPAT in rats. AB - Systemic administration of the selective 5-HT(1A) receptor agonist 8-hydroxy-2 (di-n-propylamino)tetralin HBr (8-OH-DPAT) increases waking and reduces slow wave sleep (SWS) and rapid eye movement (REM) sleep in the freely moving rat. The selective 5-HT(1A) antagonist 4-(2'-methoxy-phenyl)-1-[2'-(n-2"-pyridinyl)-p iodobenzamido]-ethyl-piperazine (p-MPPI) induces a dose-related decrease in REM sleep. The present study examined p-MPPI's potential as an antagonist of the sleep and waking responses elicited by 8-OH-DPAT. Also, the experiments explored the ability of p-MPPI to block behavioural reactions of the 5-HT syndrome induced by 8-OH-DPAT, and whether p-MPPI induced any behavioural effects of its own. This study demonstrated that pre-treatment with p-MPPI (5 mg/kg intraperitoneal (i.p.)) 30 min before 8-OH-DPAT (0.375 mg/kg subcutaneously (s.c.)) reduced the effect of 8-OH-DPAT on waking and REM sleep. Also, p-MPPI (5 and 10 mg/kg i.p.) reduced the effect of 8-OH-DPAT on locomotion and partially or completely antagonized hindlimb abduction and flat body posture. No overt behavioural change was produced by p-MPPI alone. Thus, p-MPPI behaved as a true 5-HT(1A) antagonist. PMID- 11275296 TI - Effects of dopamine antagonists with different receptor blockade profiles on morphine-induced place preference in male mice. AB - The effects of dopamine (DA) antagonists with different selectivity for the DA receptors (SCH 23390, 0.5, 0.25, 0.125 mg/kg; haloperidol, 0.2, 0.1 mg/kg; raclopride, 1.2, 0.6, 0.3 mg/kg; risperidone, 0.4, 0.2, 0.1 mg/kg; U-99194A maleate, 40, 20 mg/kg; clozapine, 2.5, 1.25, 0.625 mg/kg) on the acquisition of place conditioning and morphine-induced conditioned place preference (CPP) were explored in male mice. Morphine (40 mg/kg) produced CPP while SCH 23390, haloperidol and clozapine (highest dose) and risperidone (lowest dose) produced conditioned place aversion (CPA). Raclopride and U-99194A maleate did not produce CPP or CPA. Morphine-induced CPP was reversed by the administration of SCH 23390 and risperidone (all doses), haloperidol (highest dose) and raclopride and clozapine (intermediate and lowest doses). U-99194A maleate did not reverse morphine-induced CPP. These results suggest that the conditioned rewarding effects of morphine are mediated by the different subtypes of DA receptors. PMID- 11275297 TI - Nitric oxide involvement in the anxiogenic-like effect of substance P. AB - This study investigates whether nitric oxide (NO) is involved in the anxiogenic profile of action of substance P (SP) in mice in the elevated plus-maze (EPM). Adult Swiss mice were injected with NOS inhibitors such as L-NOARG (20 nmol/kg) i.p., L-NAME (3 nmol per site), 7-NI (0.25 nmol per site) i.c.v. or vehicle (NaCl 0.9% i.p. or PBS i.c.v.). About 30 min (i.p. pretreatment) or 5 min later (i.c.v. pretreatment), the animals received i.c.v. injections of SP (10 pmol) or phosphate buffered saline (PBS) (2 microl). Afterwards, they were observed in the EPM. SP per se reduced the time spent on open arms, an anxiogenic-like effect. This effect was reverted by different NOS inhibitors and the NO donor. NOS inhibitors had no influence on the EPM parameters but the NO-releasing compound SNAP, as well as its parent thiol NAP, increased the animals' locomotor activity. 8-Br-cGMP (20 nmol), a permeable cGMP analog, promoted an anxiogenic-like effect per se and enhanced the SP effect on the EPM. Altogether, these results suggest a putative NO role in the mediation of the anxiogenic-like effect of SP. PMID- 11275298 TI - Dissociation between the effects of pre-weaning and/or post-weaning social isolation on prepulse inhibition and latent inhibition in adult Sprague--Dawley rats. AB - Human attentional impairments can be modelled in the rat using the prepulse inhibition (PPI) or the latent inhibition (LI) paradigm. The present study investigated the consequences of a combination of pre-weaning maternal separation (MS) and post-weaning social isolation (SI) on both PPI and LI in male and female Sprague--Dawley rats tested as adults. We report here a double dissociation between the effects of MS (repeated 4 h daily separations) and SI on PPI and LI: MS did not modify PPI, but enhanced LI. In contrast, SI disrupted PPI, the deficits being restricted to male rats, but left LI intact. There were no additive effects of MS and SI on PPI or LI. While MS improved avoidance learning, SI impaired it. Although both PPI and LI assess processes of selective attention, our results support the contention, already stated in the literature, that they involve differing neuro-psychological mechanisms. Furthermore, the fact that only males exhibited PPI deficits following SI has implications for the well-known differential vulnerability of human males to certain psychiatric disorders (e.g. schizophrenia). Finally, the combination of MS and SI could represent a relevant animal model for some aspects of schizophrenia, since both PPI and LI were altered. PMID- 11275299 TI - Health care under transformation in Poland. AB - The general health insurance introduced in Poland in 1999 is essentially a social insurance. In this article, the main features of the present health care system are discussed, i.e. the sources and principles of financing, ownership relations, structures, entitlements to obtain medical services and the rules of access to services. Emphasis has been put on the operations of various entities operating within the health care sector, including opportunistic conduct of the providers of services financed from public sources, cost dumping, establishing provider alliances, methods of cost control, and the fact that some patients leave the publicly financed system. In Poland, a parallel private system has been developing for many years. Systemic transformations have not changed that direction, but increased considerably the significance of household income and education as the factors that differentiate patient in equality. This article is concluded with the note on the opportunities for the development of supplementary private insurance. PMID- 11275300 TI - Legislation analysis according to WHO and INCB criteria on opioid availability: a comparative study of 5 countries and the state of Texas. AB - Opioids are not always available in many developing countries, including those in Latin America. In this study we analyzed the national laws on opioids and other controlled substances from Argentina, Colombia, Costa Rica, Peru, Mexico, and the state of Texas, according to the principles set by the World Health Organization (WHO) and the International Narcotics Control Board (INCB), as well as to the presence of over-regulations regarding their medical and scientific use. The six main principles outlined by WHO and INCB for opioid availability were analyzed by using a total of 17 criteria as shown in Table 3. The result scores ranged from 17/17 (full compliance with all criteria) to 0/17 (non-compliance). Results showed that with the exception of the state of Texas 16/17 (94%), the countries failed to adequately meet the INCB and WHO criteria: Argentina: 7/17 (41%); Colombia: 9 /17 (53%); Costa Rica: 9/17 (53%); Mexico: 4/17 (24%); and Peru: 7/17 (41%). In all 5 Latin American countries, national laws and regulations imposed limits on the number of days allowed for prescription, the potency of the dosage, and the number of doses allowed per day. In all cases, including Texas, there was confusion on the meaning and utilization of the terms physical dependence, psychological dependence, addiction, tolerance and abuse. In total, combining all cases, only 51% of the criteria were met. Additionally, all laws and regulations, especially in Argentina, include over regulations and statements that may further interfere with patient access to opioids. The prescription criteria were fully met by the state of Texas and all five countries. These results indicate that there is need to revise the existing laws and regulations in countries with opioid availability problems, and identify the potential barriers, which may be playing a significant role in the access to adequate treatment. Such review seeks to carefully consider all possible criteria, since partial resolution of legislative articles will not result in increased opioid availability. PMID- 11275301 TI - Maternal serum screening, political decision-making and social learning. AB - OBJECTIVE: Analysis and evaluation of the process by which serum screening has been introduced in Dutch maternity care. In our analysis of Dutch medical journals, reports of the government and the political and cultural debate, we use a theoretical framework in which the process of introduction of new technologies is described in terms of different social learning processes. RESULTS: The analysis shows a dual-track pattern: promotion by the medical community, and control and regulation of serum screening by the political decision-makers. This process left little room for social learning processes. This applied in particular to learning processes about the social and cultural meaning and acceptability of serum screening. Acceptability of the new serum screening was framed nearly exclusively in terms of freedom of choice. CONCLUSIONS: A too limited framework of assessment was used. Health technology assessment (HTA) should incorporate in the evaluation process the ethical, social, cultural and political dimensions of health technology, and stimulate the interplay between different perspectives and preferences among parties involved. As a new promising approach, we suggest the use of pilot experiments that are designed as social experiments involving different parties in a common learning process. PMID- 11275302 TI - The German strategy for quality improvement in health care: still to improve. AB - One basic aim of German Health Policy is to secure a high quality of medical services financed by Statutory Health Insurance (SHI). Due to the specific relationships among patients, insurance funds and medical providers this aim is in a potential conflict with various cost-containment efforts introduced within the last 10 years. This article outlines a general theoretical framework to analyze the possible effects of these efforts on the quality of health care. It then analyzes the actual regulation approach strategy taken by German Health Policy for quality assurance in health care. Finally, it outlines a two-tier strategy based on regulation and competition as a means to ensure a high quality of care. PMID- 11275303 TI - Using computerised patient-level costing data for setting DRG weights: the Victorian (Australia) cost weight studies. AB - Casemix-funding systems for hospital inpatient care require a set of resource weights which will not inadvertently distort patterns of patient care. Few health systems have very good sources of cost information, and specific studies to derive empirical cost relativities are themselves costly. This paper reports a 5 year program of research into the use of data from hospital management information systems (clinical costing systems) to estimate resource relativities for inpatient hospital care used in Victoria's DRG-based payment system. The paper briefly describes international approaches to cost weight estimation. It describes the architecture of clinical costing systems, and contrasts process and job costing approaches to cost estimation. Techniques of data validation and reliability testing developed in the conduct of four of the first five of the Victorian Cost Weight Studies (1993-1998) are described. Improvement in sampling, data validity and reliability are documented over the course of the research program, the advantages of patient-level data are highlighted. The usefulness of these byproduct data for estimation of relative resource weights and other policy applications may be an important factor in hospital and health system decisions to invest in clinical costing technology. PMID- 11275304 TI - Yeast prions and evolvability. PMID- 11275305 TI - DNA methylation learns to fly. PMID- 11275306 TI - Why do genes have introns? Recombination might add a new piece to the puzzle. PMID- 11275307 TI - Transcription unit conservation in the three domains of life: a perspective from Escherichia coli. AB - Here we address the question of the degree to which genes within experimentally characterized operons in one organism (Escherichia coli) are conserved in other genomes. We found that two genes adjacent within an operon are more likely both to have an ortholog in other organisms, regardless of relative position, than genes adjacent on the same strand but in two different transcription units. They are also more likely to occur next to, or fused to, one another in other genomes. Genes frequently conserved adjacent to each other, especially among evolutionarily distant species, must be part of the same transcription unit in most of them. PMID- 11275308 TI - The increasing complexity of the Snail gene superfamily in metazoan evolution. AB - The Snail family of zinc-finger transcription factors is involved not only in the development of vertebrate and invertebrate embryos, but also in tumour progression. Following the identification of eight new members, we have analysed the evolutionary history of these genes and found that they constitute a superfamily that groups two independent families, Snail and Scratch. We propose that the duplication of an ancestral gene at the time of the metazoan radiation (1000-500 Myr ago) gave rise to Snail and Scratch, and that independent duplications in protostomes and deuterostomes led to the present situation. We discuss the implications of the distinct duplication events on the acquisition of new functions. PMID- 11275324 TI - How Bacillus thuringiensis has evolved specific toxins to colonize the insect world. AB - Bacillus thuringiensis is a bacterium of great agronomic and scientific interest. Together the subspecies of this bacterium colonize and kill a large variety of host insects and even nematodes, but each strain does so with a high degree of specificity. This is mainly determined by the arsenal of crystal proteins that the bacterium produces during sporulation. Here we describe the properties of these toxin proteins and the current knowledge of the basis for their specificity. Assessment of phylogenetic relationships of the three domains of the active toxin and experimental results indicate how sequence divergence in combination with domain swapping by homologous recombination might have caused this extensive range of specificities. PMID- 11275325 TI - What regulates mitochondrial DNA copy number in animal cells? AB - The study of the control of mitochondrial DNA copy number spans several decades and has identified many factors involved in the replication of the mitochondrial genome. However, the mechanisms involved in the regulation of this process are still obscure, particularly in animal cells. During the past decade, however, the identification of human diseases associated with drastically reduced levels of mtDNA caused renewed interest in this topic. Here, I will discuss recent work that sheds some light on how animal cells might maintain and control mtDNA levels. PMID- 11275326 TI - Nuclear receptors in nematodes: themes and variations. AB - Large-scale sequencing efforts are providing new perspectives on similarities and differences among species. Sequences encoding nuclear receptor (NR) transcription factors furnish one striking example of this. The three complete or nearly complete metazoan genome sequences - those of the nematode Caenorhabditis elegans, the fruit fly (Drosophila melanogaster) and the human - reveal dramatically different numbers of predicted NR genes: 270 for the nematode, 21 for the fruit fly and approximately 50 for the human. Although some classes of NRs present in insects and mammals are also represented among the nematode genes, most of the C. elegans NR sequences are distinct from those known in other phyla. Questions regarding the evolution and function of NR genes in nematodes, framed by t00e abundance and diversity of these genes in the C. elegans genome, are the focus of this article. PMID- 11275327 TI - Endogenous DNA damage and mutation. AB - In humans, approximately 10(7) cells divide per second. Estimates suggest that spontaneous mutations arise in about a third of those cells. These mutations arise as mistakes in DNA replication and when DNA polymerases copy damaged templates. The latter result from chemical hydrolysis of nucleoside bases or by reaction of DNA with electrophiles or reactive free radicals generated during metabolism (endogenous DNA damaging agents). This article highlights recent discoveries and emerging opportunities in the study of endogenous DNA damage and mutation. PMID- 11275328 TI - The myotubularin family: from genetic disease to phosphoinositide metabolism. AB - The myotubularin-related genes define a large family of eukaryotic proteins, most of them initially characterized by the presence of a ten-amino acid consensus sequence related to the active sites of tyrosine phosphatases, dual-specificity protein phosphatases and the lipid phosphatase PTEN. Myotubularin (hMTM1), the founder member, is mutated in myotubular myopathy, and a close homolog (hMTMR2) was recently found mutated in a recessive form of Charcot-Marie-Tooth neuropathy. Although myotubularin was thought to be a dual-specificity protein phosphatase, recent results indicate that it is primarily a lipid phosphatase, acting on phosphatidylinositol 3-monophosphate, and might be involved in the regulation of phosphatidylinositol 3-kinase (PI 3-kinase) pathway and membrane trafficking. PMID- 11275330 TI - Visual tests for measuring the picture quality of teleconsultations for medical purposes. AB - Telemedicine provides a new way of delivering medical services. The good quality of the picture is, however, essential for a proper and reliable teleconsultation, especially in the cases when visual information about the patient's physical condition is of great importance as in ophthalmology and dermatology. Therefore real-time telemedical applications need standards for these procedures, e.g. minimum requirements for resolution, as well as contrast and color discrimination. The present study was carried out to test these parameters in a real environment. A panel of different resolution, contrast sensitivity and color discrimination tests was carried out for five test persons via PictureTel videoconference system by ISDN 128 kbit/s line speed and in a normal way. The good color discrimination of the teleconsultation system makes it possible to use the system in the evaluation of different skin lesions and exemas. However, the poor resolution and especially poor contrast sensitivity makes the use of the equipment valueless in the evaluation of diseases where the diagnoses are based on the discrimination of small details, like the biomicroscopical analysis of inflammatory cells in the aqua's humor in cases of intraocular inflammation. The authors are suggesting a simple panel of tests to optimize and standardize these parameters. PMID- 11275331 TI - Presentation of dermatological images on the Internet. AB - In this paper, we focused on selected problems of integrating and presenting medical images organised in a World Wide Web (WWW) database. To solve these problems we developed a prototype of a bilingual (Slovenian and English) WWW database of medical images for the field of dermatology. This dermatology database includes a graphic interface with four modes of access: (1) browsing, (2) searching, (3) comparison of images, and (4) self-testing. The quantity and quality of requests to this WWW database was estimated with log file analysis. There was a steady increase in the number of users and volume of data transferred from the dermatology WWW database. PMID- 11275332 TI - Solving a linear model of nonfatal risk behavior and injuries in school children. AB - A calculating method and computer program have been developed for solving an overdetermined system of linear equations. It was applied for calculation of group risk exposure of school children (10-15 years) to nonfatal injuries in sport recreation and playing (SRP). Interviewed non-injured school children were separated into non-overlapping groups of risky behavior. Data on interviewed non injured and non-interviewed injured children were put into a linear system of equations consisting of a 4 x 3 matrix scheme. Each equation represents children of the same age and sex and consists of the percentage of interviewed children grouped into high medium or low risk behavior and of the percentage of injured children. Four methods of calculating the matrix system were evaluated and the best implemented as a new developed computer program. The resulting linear group risk factors enable the prediction of behavior outcome. Data on interviewed injured children were compared with the results obtained. The validation procedure is proposed by testing the sensitivity and robustness of the method. PMID- 11275333 TI - A simple computerized program for the calculation of the required sample size necessary to ensure statistical accuracy in medical experiments. AB - We developed a sample size estimation program (SSEP) with which medical researchers can easily estimate the appropriate sample size for a specific significance level and statistical power using their favorite WWW browsers. SSEP can estimate the sample sizes for six statistical methods by Monte-Carlo simulation: Student's t-test, Welch's t-test, Analysis of variance, Wilcoxon's rank sum test, Kruskal-Wallis test, and the Cochran-Armitage test for linear trends. The SSEP simulation programs were created using the SAS software macro language. Medical researchers can interactively use this program and determine reliable sample sizes when planning new prospective clinical studies and animal experiments. PMID- 11275334 TI - Development and implementation of the population Fisher information matrix for the evaluation of population pharmacokinetic designs. AB - In population pharmacokinetic studies, the precision of parameter estimates is dependent on the population design. Methods based on the Fisher information matrix have been developed and extended to population studies to evaluate and optimize designs. In this paper we propose simple programming tools to evaluate population pharmacokinetic designs. This involved the development of an expression for the Fisher information matrix for nonlinear mixed-effects models, including estimation of the variance of the residual error. We implemented this expression as a generic function for two software applications: S-PLUS and MATLAB. The evaluation of population designs based on two pharmacokinetic examples from the literature is shown to illustrate the efficiency and the simplicity of this theoretic approach. Although no optimization method of the design is provided, these functions can be used to select and compare population designs among a large set of possible designs, avoiding a lot of simulations. PMID- 11275336 TI - Proceedings of the 1st International Workshop Modelling of Spinal Loads Associated with Vibration and Shock. October 1999, Berlin, Germany. PMID- 11275335 TI - Volumetric quantification of the gastric emptying: computer-based method for generation of volumetric index from fluoroscopic images. AB - We developed an automated software-based procedure for estimation of the volume variation of the stomach using videofluoroscopic analysis of the gastric emptying. We used radiological images with postero-anterior incidence of eight healthy volunteers and in vitro experimental tests, with different volumes and concentrations of the contrast medium. This computational method generates an index that measures, in the three dimensions, the dynamic behaviour of gastric emptying. Using adequate contrast concentration (barium sulphate solution), it is possible to determine volume behaviour from density variations. This software can automate this computation, facilitating the amount of work, avoiding mistakes and improving reproducibility. PMID- 11275337 TI - Modelling the response of the spinal system to whole-body vibration and repeated shock. AB - Back problems are very common and contribute to discomfort and days off work. Some back disorders are attributed to inappropriate loading of the spine that can be combined with other influential factors such a body posture, whole-body vibration and shock. Many models have been developed to predict the forces in the spine associated with vibration and shock. However, the problem is complex due to the incompletely understood dynamic responses of the body, the influence of many variables and the effect of muscle forces. This paper summarises the current state of knowledge relevant to the prediction of forces in the spine associated with whole-body vibration and shock. The paper also introduces presentations at a workshop on the modelling of spinal loads associated with vibration and shock held in Berlin in October 1999. RelevanceBiodynamic models may be used to estimate risks of injury or disease and optimise environments so as to minimise risks. However, the development of useful models requires an understanding of the complex interaction between mechanical forces and the living body. The application of models requires knowledge of their accuracy and limitations. PMID- 11275338 TI - Recent advances in lumbar spinal mechanics and their significance for modelling. AB - Mathematical models are often used to quantify the overall forces and moments acting on the lumbar spine. However, if the purpose of the research is to explain how spinal tissues can be injured, it is necessary to distribute the overall forces and moments between (and within) different spinal structures, because it is the concentration of force which causes injury, and elicits pain. This paper reviews recent experimental evidence concerning the distribution of forces and moments acting on the lumbar spine. Lordotic postures increase loading of the posterior annulus and apophyseal joints, whereas moderately flexed postures tend to equalise compressive stress across the disc, and unload the apophyseal joints. Sustained compression reduces the volume and pressure of the nucleus pulposus, while increasing compressive stresses in the annulus and neural arch. Sustained compression also reduces disc height, giving some slack to collagen fibres in the intervertebral disc and ligaments, and causing them to resist bending less. Disc degeneration has a similar effect on disc height, and stress distributions. On the other hand, discs and ligaments can be subjected to greater bending moments following a period of sustained or repetitive bending, because sustained bending impairs the normal protective reflex from the back muscles, and repetitive bending fatigues the back muscles, reducing their ability to protect the spine. Incorporating this information into mathematical models will make them better able to identify which activities are most likely to injure the lumbar spine in life. PMID- 11275339 TI - An EMG technique for measuring spinal loading during asymmetric lifting. AB - OBJECTIVES: To compare two methods of calibrating the erector spinae electromyographic signal against moment generation in order to predict extensor moments during asymmetric lifting tasks, and to compare the predicted moments with those obtained using a linked-segment model. METHODS: Eight men lifted loads of 6.7 and 15.7 kg at two speeds, in varying amounts of trunk rotation. For each lift, the following were recorded at 60 Hz; the rectified and averaged surface electromyographic signal, bilaterally at T10 and L3, lumbar curvature using the 3 Space Isotrak, movement of body segments using a 4-camera Vicon system, and ground reaction forces using a Kistler force-plate. Electromyographic (EMG) and Isotrak data were used to calculate lumbosacral extensor moments using the electromyographic model, whereas movement analysis data and ground reaction forces were used to estimate net moments using the linked-segment model. For the electromyographic technique, predictions of extensor moment were based on two different sets of EMG-extensor moment calibrations: one performed in pure sagittal flexion and the other in flexion combined with 45 degrees of trunk rotation. RESULTS: Extensor moments predicted by the electromyographic technique increased significantly with load and speed of lifting but were not influenced by the method of calibration. These moments were 7-40%greater than the net moments obtained with the linked-segment model, the difference increasing with load and speed. CONCLUSIONS: The calibration method does not influence extensor moments predicted by the electromyographic technique in asymmetric lifting, suggesting that simple, sagittal-plane calibrations are adequate for this purpose. Differences in predicted moments between the electromyographic technique and linked-segment model may be partly due to different anthropometric assumptions and different amounts of smoothing and filtering in the two models, and partly due to antagonistic muscle forces, the effects of which cannot be measured by linked-segment models. RelevanceAsymmetric lifting is a significant risk factor for occupationally-related low back pain. Improved techniques for measuring spinal loading during such complex lifting tasks may help to identify work practices which place the spine at risk of injury. PMID- 11275340 TI - Examination of the myoelectric activity of back muscles during random vibration- methodical approach and first results. AB - OBJECTIVE: To elaborate methods for an elimination of artefacts and the analysis of the relationship between random whole-body vibration and electromyographic responses of back muscles. DESIGN: A procedure involving wavelets and digital filtering has been used for the removal of artefacts from the electromyogram during whole-body vibration. BACKGROUND: Back muscle forces contribute essentially to the whole-body vibration-induced spinal load. The electromyogram can help to estimate these forces during whole-body vibration. METHODS: 38 subjects were exposed to identical random low-frequency whole-body vibration. Artefacts caused by the electrocardiogram in the electromyogram were identified by appropriate wavelets and eliminated in the time-domain. After averaging the individual high-pass filtered and rectified undistorted electromyograms across subjects, the transfer function from seat acceleration to the average electromyogram was determined and used for the prediction of the electromyogram. RESULTS: A sufficient procedure involving wavelets and digital filtering has been elaborated for the removal of artefacts from the electromyogram of back muscles during whole-body vibration. A systematic relationship between random vibration and back muscle-response was obtained and described. The transfer function suggests two different reflex-mechanisms - one elicited below, the other above 4 Hz. CONCLUSIONS: The approach of analysing and predicting the muscle-response to random vibration by using the transfer function seems to be promising and could be a valuable tool for the future calculation of muscle forces as an input to active models. RELEVANCE: The knowledge of the extent and timing of the back muscle-response to random whole-body vibration is relevant for an improved evaluation of whole-body vibration with respect to health. PMID- 11275341 TI - Modelling the dynamic mechanisms associated with the principal resonance of the seated human body. AB - OBJECTIVES: Simple mathematical models have been developed to obtain insights into resonance phenomena observed at about 5 Hz in the dynamic responses of the seated human body exposed to vertical whole-body vibration. DESIGN: Alternative lumped parameter models with a few degrees-of-freedom have been investigated. Rotational degrees-of-freedom, with eccentricity of the centre of gravity of the mass elements, represented responses in the fore-and-aft and pitch axes caused by vertical vibration. BACKGROUND: The causes of body resonance are not fully understood, but this information is required to develop cause-effect relationships between vibration exposures and effects on human health, comfort and performance.Method. The inertial and geometric parameters for models were based on published anatomical data. Other mechanical parameters were determined by comparing model responses to experimental data. RESULTS: Two models, with four and five degrees-of-freedom, gave more reasonable representations than other models. Mechanical parameters obtained with median and individual experimental data were consistent for vertical degrees-of-freedom but varied for rotational degrees-of-freedom. CONCLUSIONS: The resonance of the apparent mass at about 5 Hz may be attributed to a vibration mode consisting of vertical motion of the pelvis and legs and a pitch motion of the pelvis, both of which cause vertical motion of the upper-body above the pelvis, a bending motion of the spine, and vertical motion of the viscera. RELEVANCE: The mathematical models developed in this study may assist understanding of the dynamic mechanisms responsible for resonances in the seated human body. The information is required to represent mechanical responses of the body and assist the development of models for specific effects of vibration. PMID- 11275342 TI - Determination of vibration-related spinal loads by numerical simulation. AB - OBJECTIVE: Dynamic spinal loads due to human whole body vibrations are extremely difficult to determine experimentally. However, they can be predicted by numerical simulation. This paper presents an approach for the prediction of dynamic spinal loads caused by whole body vibrations, as well as some basic considerations concerning the process of numerical simulation. BACKGROUND: Long term whole body vibrations have been found to cause health risks for the lumbar spine. As an increasing percentage of the population is exposed to whole body vibrations at work, more and more people have to face the risk of whole body vibrations-related injury. Knowledge about the actual loads in the lumbar spine is essential when spinal loads are to be compared with spinal strength in order to assess the possible health risks caused by whole body vibrations. METHODS: Since an extrapolation of results to unknown data such as spinal loads can only be done using anatomical models of the human body, a simplified finite-element model is presented which is adaptable to body height, body mass, and posture of any specific subject under investigation. The model has been built by reducing a very detailed, nonlinear finite-element model of seated man in its complexity (number of degrees of freedom). Furthermore, the simplified model has been linearised to avoid nonlinear solution procedures. RESULTS: The model has been verified for vertical and horizontal excitation at the seat. Model results have been compared to measurements on subjects. Individual exposure-effect relationships may be predicted by this model, due to the adaptability to a specific subject. Additionally, a new phenomenological method of eliminating the influence of local skin-accelerometer vibrations on vibration measurements on the skin surface is discussed. This method may provide data about bone acceleration that can be used in the process of model verification. CONCLUSIONS: Integral loading measures, such as spinal loads, may be predicted with simplified finite element models. Quantitative judgements of these loads may be performed for individual conditions. Linearised models may be used for limited ranges of excitation intensities. Energy dissipation should be modeled by discrete dashpot elements instead of proportional damping. RELEVANCE: In order to assess the risk of an injury to the lumbar spine due to whole body vibrations, spinal loads have to be compared with spinal strength. This paper presents the development and verification of a simplified finite-element model of the human body which is based on human anatomy and therefore well-suited to occupational/clinical biomechanics for the prediction of spinal loads. PMID- 11275343 TI - Application of finite-element models to predict forces acting on the lumbar spine during whole-body vibration. AB - OBJECTIVE: To predict forces acting on the spine during whole-body vibration for a variety of boundary conditions - body mass, height and posture.Design. Representative anthropometric data and models for an upright, relaxed and bent forward sitting posture were used to derive model families with 30 variants of a finite-element model. BACKGROUND: A given exposure to whole-body vibration can cause a variable health risk depending on the concomitant conditions. The latter could contribute to the considerable uncertainty of the current evaluation of whole-body vibration. METHODS: Plane symmetric linear finite-element models were used for the prediction of static and dynamic compression and shear forces acting on the lumbar discs during whole-body vibration. Transfer functions from seat acceleration to forces were determined. RESULTS: A bent forward posture augments essentially the compressive and shear stress, predicted for erect and relaxed sitting postures. The normal variation of body mass and height causes a considerable variation of static internal shear stress, but a minor variation of compressive pressure. The dynamic internal stress varies nearly proportionally to the body mass. The transfer functions from seat acceleration to compressive force depend significantly on the posture. CONCLUSIONS: The variability of the spinal loads for a given whole-body vibration and associated with a normal range of several biological factors suggests a ratio between the minimum and maximum internal loads of about 1:2. RELEVANCE: Finite-element models can be used to compare the health risk arising from different whole-body vibration exposures and individual conditions. These results help to prevent work-related disorders of the lumbar spine. PMID- 11275344 TI - Design of an active vibration dummy of sitting man. AB - OBJECTIVE: The determination of vibration transmission through vehicle seats today is still performed with small groups of test subjects. This method suffers from several severe disadvantages, in particular poor repeatability and objectivity due to varying test conditions and limited group sizes. Replacing test persons with a vibration dummy helps to solve this problem. DESIGN: The active vibration dummy simulates the dynamic behaviour of sitting man expressed in terms of the driving point impedance for arbitrary body masses and excitation signals. METHODS: The dummy is realized as a mechatronic system basing on a single degree of freedom setup. A real-time control loop of mass accelerations (and thus acting forces) fits the active dummy to the desired driving point impedance data set. Model and controller parameters are determined by a parameter identification technique giving meaningful results for arbitrary impedance data sets. RESULTS: The prototype shows excellent agreement with the target data under laboratory conditions. Body mass and excitation level can be varied over the full range of car seat test requirements. RELEVANCE: The determination of vibration transmission through vehicle seats should be possible without human experiments. An active vibration dummy with adjustable vibration behaviour expressed by the vertical driving point impedance covering the entire scope of car seat tests (masses/excitation intensities) is presented. With the dummy, improved seat test procedures could be established, leading to design improvements and therefore to prevention of whole-body vibration injuries. PMID- 11275345 TI - Estimation of muscle forces in the lumbar spine during upper-body inclination. AB - OBJECTIVE: To estimate the muscle forces during upper-body inclination and to determine their influence on stress distribution in the annulus fibrosus of the lumbar spine discs. DESIGN: The muscle forces and stresses were calculated using a non-linear finite element model of the lumbar spine. BACKGROUND: Little is known about the influence of muscle forces on the deformation of, and stresses in, the lumbar spine. In most studies, muscle forces are neglected. METHODS: Three-dimensional non-linear finite element models of the ligamentous lumbar spine, with and without internal spinal fixators, were created. They were validated by use of experimental data from in vitro measurements on cadaver specimens. In a second step, the influence of muscle forces on stresses in the annulus fibrosus of the lumbar spine discs was investigated in a parameter study. This was done for different inclination angles of the upper-body. RESULTS: Good agreement between analytical and experimental results proved achievable when loading with pure moments in the three main planes of the lumbar spine. For inclination of the upper-body, the flexion angle clearly has a strong influence on the stresses in the lumbar spine while the influence of local muscles was small. The stress distribution in the discs differed considerably when the muscle forces are neglected and only a pure moment is applied. CONCLUSIONS: This study confirmed earlier ones that have shown that muscle forces should not be neglected when studying the stresses in the lumbar spine. The local dorsal muscles, however, have only a small influence on the stresses in the discs. RELEVANCE: For investigations of the biomechanical effects of spinal implants and surgical procedures, experimental or analytical methods are used. Due to the complexity involved, as well as to a lack of information, muscle forces are often neglected. Our study showed that muscles do in fact have a major influence on the mechanical behaviour of the spine and should always be taken into account. PMID- 11275346 TI - The validation of biodynamic models. AB - Biodynamic models may: (i) represent understanding of how the body moves (i.e., 'mechanistic models'), (ii) summarise biodynamic measurements (i.e., 'quantitative models'), and (iii) provide predictions of the effects of motion on human health, comfort or performance (i.e., 'effects models'). Model validation may involve consideration of evidence used to derive a model, comparison of the model with alternatives, and a comparison between model predictions and independent observations of the predicted qualities or quantities. Models should be associated with a specified range of independent and dependent variables and indicate how intra-subject variability and inter-subject variability are accommodated. Models of the mechanisms of body movement may be validated by demonstrations that the mechanisms are well represented. Models giving numerical predictions ('quantitative models' and 'effects models') should specify the expected accuracy of predictions. 'Effects models' advocated for predicting health, comfort or performance require that: (i) vibration or shock is a proven cause of the specified effect, (ii) within all reasonable ranges of model inputs, there must be reason to expect a positive correlation and acceptable error between the model predictions and the effect, (iii) other variables having a large influence on the effect must be taken into consideration. It is more useful to report the accuracy of 'quantitative models' and 'effects models' models than to state that they are 'validated' or 'un-validated'. Checklists for assessing the quality of a biodynamic model are proposed, taking into account the type of model and the model assertions, the evidence, the assumptions, the accuracy, and the appropriateness of the model. RELEVANCE: Biodynamic models can be used to predict risks of injury or disease. Models can be used to optimise designs in order to minimise predicted risks. However, models can be promulgated and used without knowledge of their accuracy or usefulness. PMID- 11275347 TI - Transfer functions as a basis for the verification of models--variability and restraints. AB - OBJECTIVE: The seat-to-head transfer function of the human body reflects the biodynamic response. Based on measured data, biodynamic models have been proposed to reflect this response. They must satisfy usually the international published mean values of the seat-to-head transfer function. The question arises to what extent mean values reflect individual pattern of biodynamics. METHODS: An experimental study was performed with 39 male subjects sitting on a hard seat without back rest and with supported feet. They were exposed to random whole-body vibration at three intensities with a relaxed and an erect posture. The accelerations in the z-direction were measured at the seat and head. The seat-to head transfer functions with the associated coherence functions were calculated. RESULTS: The biodynamic response characterised by the maximum of the seat-to-head transmissibility and the frequency of its occurrence is influenced by the posture of the subjects in a dominant way and shows an individual variability of considerable extent. The mean responses suggest a missing effect of vibration intensity, but individually different effects of the intensity were found. Repeated measurements confirmed this result. CONCLUSIONS: The application of a model validated by the comparison with mean values of the transmissibility could cause misleading conclusions, if it is used for the prediction of individual spinal loads. Models prepared for the calculation of individual loads should be validated by a mean individual transmissibility derived from repeated measurements. RELEVANCE: The results illustrate the loss of information by averaging individual transfer functions and the consequence of a limited validity and applicability in occupational health, ergonomics, and design. PMID- 11275348 TI - Mechanical impedance of the sitting human body in single-axis compared to multi axis whole-body vibration exposure. AB - OBJECTIVE: The study was aimed to investigate the mechanical impedance of the sitting human body and to compare data obtained in laboratory single-axis investigations with multi-axis data from in vehicle measurements. DESIGN: The experiments were performed in a laboratory for single-axis measurements. The multi-axis exposure was generated with an eight-seat minibus where the rear seats had been replaced with a rigid one. The subjects in the multi-axis experiment all participated in the single-axis experiments. BACKGROUND: There are quite a few investigations in the literature describing the human response to single-axis exposure. The response from the human body can be expected to be affected by multi-axis input in a different way than from a single-axis exposure. The present knowledge of the effect of multiple axis exposure is very limited. METHODS: The measurements were performed using a specially designed force and accelerometer plate. This plate was placed between the subject and the hard seat. RESULTS: Outcome shows a clear difference between mechanical impedance for multi-axis exposure compared to single-axis. This is especially clear in the x-direction where the difference is very large. CONCLUSIONS: The conclusion is that it seems unlikely that single-axis mechanical impedance data can be directly transferred to a multi-axis environment. This is due to the force cross-talk between different directions. PMID- 11275349 TI - Intradiscal pressure together with anthropometric data--a data set for the validation of models. AB - OBJECTIVE: To provide a database of intradiscal pressure measurements together with anthropometric data as basis for the validation of models that predict spinal loads. DESIGN: Intradiscal pressure was measured in a non-degenerated L4-5 disc of a volunteer. The anthropometric characteristics of this subject were extensively determined. BACKGROUND: Since it is usually impossible to quantify the load in the spine directly, it is predicted by various biomechanical models. However, they often cannot be validated because of the few in vivo data and missing anthropometric characteristics pertaining to them. METHODS: A pressure transducer (diameter 1.5 mm) was implanted in the nucleus pulposus of a non degenerated L4-5 disc of a volunteer. Pressure was determined during exercises while standing, lifting activities, sitting unsupported on a stool or an ergonomic sitting ball, sitting in different postures and others. The anthropometric characteristics were determined using different tools. RESULTS: Pressure values: relaxed standing 0.5 MPa; standing flexed forward 1.1 MPa; standing extended backward 0.6 MPa; sitting unsupported 0.46 MPa; maximum values during lateral bending 0.6 MPa, during axial rotation 0.7 MPa, lifting a 20 kg weight with a round flexed back 2.3 MPa, with flexed knees 1.7 MPa, close to the body 1.1 MPa; sitting unsupported relaxed 0.45 MPa, actively straightening the back 0.55 MPa, with flexion 0.9 MPa; non-chalant sitting 0.3 MPa and others. Anthropometric characteristics with emphasis on data for the trunk are provided in tables.Conclusions. Intradiscal pressure depends on the kind of preceding activity, posture, external loads and muscle activity. RELEVANCE: The data set can be used to verify a biomechanical model adjusted to the individual characteristics by a comparison of measured and predicted intradiscal pressures. PMID- 11275350 TI - Lumbar intradiscal pressure and whole-body vibration--first results. AB - OBJECTIVE: To contribute to the scientific background for the assessment of the health risk at the lumbar spine from whole-body vibration. DESIGN: Experimental study. BACKGROUND: Many workers have monitored the vibration at spinal locations on the skin or using skeleton mounted devices in order to assess the vertebral response when sitting. They have shown that resonance occurred in the 4-7 Hz range. Considering the different structures of the intervertebral joint, it seemed interesting to assess their behaviour separately, using intranuclear pressures at the lumbar discs monitored simultaneously with the vertebral accelerations. METHODS: Seven unembalmed cadavers were submitted to 5 min. whole body random vibration, in four seated postures (erect or as in a car, both with or without a lumbar support). Power spectral density functions were estimated at each lumbar level and each posture and the dominant frequency identified. The energy of the pressure signal was also estimated in the 0-25 Hz band. Analysis of variance was then performed to study the effects of subject, disc level, and posture. RESULTS: Energy of the intranuclear pressure variation decreased when leaning the backrest backwards. The effect of the lumbar support depended on the discal level and on posture. The shape of the power spectral density function suggested the existence of a cyclic loading of the nucleus pulposus, while more complex phenomena were observed at the vertebral body. CONCLUSIONS: The response of the lumbar spine cannot be assessed by examining only vertebral acceleration at one level. RELEVANCE: To understand how posture affects lumbar behaviour to minimize low back disorders. PMID- 11275351 TI - Stress distribution changes in bovine vertebrae just below the endplate after sustained loading. AB - OBJECTIVE: To describe the pattern of stress distribution in the vertebral body just behind the endplate, and to document its changes due to sustained loading. METHODS: Twelve fresh bovine coccygeal motion segments were dissected and tested. Each specimen was axially loaded with a sustained compressive force of 50% of its estimated compressive strength. Before loading, after 1.5 h and after 3 h of loading, the distribution of the axial pressure under the bottom vertebra (i.e., just below its top endplate) was recorded at three force levels (25%, 37.5% and 50% of the estimated compressive strength), using pressure-sensitive film. RESULTS: Stress distribution over the endplate was found to be fairly uniform. At low compression forces, the stress was the highest centrally. With increased compression and after sustained compression the uniformity improved through a significant redistribution of stress to the periphery. No stress peaks were found to occur after sustained loading. CONCLUSION: Stress peaks after sustained loading cannot explain the occurrence of endplate fractures in sustained cyclic compression in non-degenerated discs. Competing explanations, such as creep, and fatigue failure, would appear more likely candidates. RELEVANCE: It has been hypothesised that compression induced fractures of the lumbar vertebral endplate constitute an important etiological factor for low back pain. Competing theories exist on the fracture mechanism in sustained loading and these would have different implications with respect to prevention. The present study evaluated one of these theories. PMID- 11275352 TI - A review of mechanisms controlling ovulation with implications for the anovulatory effects of polychlorinated dibenzo-p-dioxins in rodents. AB - Polychlorinated dibenzo-p-dioxins (PCDDs) can impinge on female fertility by preventing ovulation. In this review, the aspects of normal ovulatory physiology most relevant to our current understanding of PCDD action on the ovary are briefly reviewed. This is followed by a comprehensive assessment of data relevant to the effects of PCDDs during ovulation in the rat. PCDDs interrupt ovulation through direct effects on the ovary in combination with dysfunction of the hypothalamo-hypophyseal axis. PMID- 11275353 TI - Methyl methacrylate toxicity in rat nasal epithelium: studies of the mechanism of action and comparisons between species. AB - Female F344 rats exposed to 200 ppm methyl methacrylate for 6 h developed a lesion in the nasal olfactory epithelium which was characterised by degeneration and atrophy. The severity of the lesion was markedly reduced by pre-treatment of the rats with an intraperitoneal dose of 100 mg/kg bis-(p-nitrophenyl)phosphate, an inhibitor of carboxylesterase enzymes, thus demonstrating that the lesion is caused by the carboxylesterase mediated metabolism of methyl methacrylate to methacrylic acid, an irritant and corrosive metabolite. The distribution of the carboxylesterases in nasal tissues has been investigated and the metabolism of methyl methacrylate to methacrylic acid has been compared in rat, hamster and human nasal tissue fractions in vitro. Histocytochemistry showed that the carboxylesterases are heavily localised in the sustentacular cells and Bowman's glands of the rat olfactory region, but are more generally distributed in human olfactory epithelium. Consistent with this, the enzyme activity in all three species was higher in fractions prepared from olfactory tissue than from respiratory tissue, 3-fold in rat and human and 12-fold in the hamster. The maximum rates (V(max)) of metabolism in rat and hamster olfactory tissue fractions were comparable, whereas those in human olfactory tissue fractions were at least 13-fold lower. The rate of metabolism in rat olfactory tissue was also comparable to that in rat liver whereas in humans, the rate in olfactory tissue was 500-fold lower than that in the liver. In respiratory tissues, the rate in humans was at least 6-fold lower than that in the rat. These results suggest that humans are significantly less sensitive than rodents to the nasal toxicity of methyl methacrylate. PMID- 11275354 TI - Studies of early hepatocellular proliferation and peroxisomal proliferation in Wistar rats treated with herbicide diclofop. AB - A study was performed to determine whether diclofop (2-(4-(2,4-dichlorophenoxy) phenoxy)propionic acid), introduced as a herbicide, exhibits the properties of peroxisome proliferators (PPs). Diclofop was administered orally at 7-56 mg/kg body weight per day to male Wistar rats for 2, 4, 7 or 14 consecutive days and some effects regarded as early hepatic markers of PPs were studied. The early changes in rat liver, produced by short-term treatment with diclofop consisted of mitogenesis and, time- and dose-related increase in liver weight. Hepatomegaly was typically associated with proliferation of smooth endoplasmic reticulum (SER) and peroxisomes. The parallel biochemical measurements showed that there was a dose-dependent increase in peroxisomal palmitoyl-CoA oxidation and catalase activity in treated rats. Markers of hepatocellular proliferation (S- and M phase) indicated that mitogenesis was transient and declined despite continuation of diclofop treatment. The threshold exposure level for the palmitoyl-CoA oxidation (one of the peroxisome proliferation markers) was approximately the same (14 mg/kg body weightxper day) as for the stimulation of mitogenesis in Wistar rats. However, for hepatomegaly and catalase activity the threshold exposure level was 7 mg/kg body weightxper day. The results presented here demonstrate clearly that diclofop belongs to a class of rodent PPs. PMID- 11275355 TI - Uridine uptake inhibition as a cytotoxicity test for a human hepatoma cell line (HepG2 cells): comparison with the neutral red assay. AB - This study describes a sensitive microassay for measuring cytotoxicity based on the degree of inhibition of RNA synthesis in HepG2 cells. RNA synthesis is measured by the kinetic uptake of radiolabeled uridine. A large number of compounds were tested in a wide range of concentrations. The concentration required to induce 50% inhibition of HepG2 uridine uptake rates (IC(50)) was determined for each compound and used to rank its potency. These IC(50)s were compared with IC(50)s measured with the neutral red assay. 2-acetylaminofluorene, benzo[a]pyrene and methylnitrosourea were not cytotoxic in the neutral red assay. Uridine uptake was always inhibited at lower concentrations than those required in the neutral red assay, suggesting that the uridine uptake assay is a more sensitive indicator of toxic action than the neutral red inclusion. Uridine uptake assay provides a rapid and quantitative method for assessing toxicity in a human cell line. Application of this method to bottled spring waters are described. Due to its high sensitivity and reproducibility, this method provides a suitable tool for screening a great number of samples and will be a helpful test for evaluating food safety and controlling the recycling process of wrapping materials. PMID- 11275356 TI - Effect of highly bioaccumulated polychlorinated biphenyl congeners on estrogen and androgen receptor activity. AB - Polychlorinated biphenyls (PCBs) are ubiquitous environmental persistent contaminants giving rise to potential health hazard. Some PCBs exert dioxin-like activities mediated through the aryl hydrocarbon receptor. Although reports on interaction with other nuclear receptors are sparce, some congeners are hypothesized to possess endocrine disruptive potential. Here we present evidence that the three PCBs most abundant in biological extracts, 2,2',3'4,4',5 hexachlorobiphenyl (PCB#138), 2,2',4,4',5,5'-hexachlorobiphenyl (PCB#153), and 2,2',3,4,4',5,5'-heptachlorobiphenyl (PCB#180) have pleiotropic effects on the estrogen- and androgen-receptor. In MCF-7 cells a slightly increased cell proliferation was observed at low concentrations (1-10 nM) in cells co-treated with 0.01 nM 17beta-Estradiol, whereas the compounds inhibited cell growth significantly at 1 and 10 microM. In reporter gene (ERE-tk-CAT) analysis the three congeners exhibited a significantly estrogen receptor-ligand mediated decrease of the chloramphenicol transferase activity in both control and 10 nM 17beta-estradiol induced MCF-7 cells. In addition, PCB#138 elicited a dose dependent antagonistic effect on androgen receptor activity in transiently co transfected Chinese Hamster Ovary cells with an IC(50), of 6.2 microM. In summary, this study indicate that the di-ortho, multiple-chloro substituted biphenyls, PCB#138, PCB#153 and PCB#180, can compete with the binding of the natural ligand to two nuclear receptors and thus possess the ability to interfere with sexual hormone regulated processes. PMID- 11275357 TI - Prostaglandins and cyclooxygenases [correction of cycloxygenases] in the spinal cord. AB - The spinal cord is one of the sites where non-steroidal anti-inflammatory drugs (NSAIDs) act to produce analgesia and antinociception. Expression of cyclooxygenase(COX)-1 and COX-2 in the spinal cord and primary afferents suggests that NSAIDs act here by inhibiting the synthesis of prostaglandins (PGs). Basal release of PGD(2), PGE(2), PGF(2alpha) and PGI(2) occurs in the spinal cord and dorsal root ganglia. Prostaglandins then bind to G-protein-coupled receptors located in intrinsic spinal neurons (receptor types DP and EP2) and primary afferent neurons (EP1, EP3, EP4 and IP). Acute and chronic peripheral inflammation, interleukins and spinal cord injury increase the expression of COX 2 and release of PGE(2) and PGI(2). By activating the cAMP and protein kinase A pathway, PGs enhance tetrodotoxin-resistant sodium currents, inhibit voltage dependent potassium currents and increase voltage-dependent calcium inflow in nociceptive afferents. This decreases firing threshold, increases firing rate and induces release of excitatory amino acids, substance P, calcitonin gene-related peptide (CGRP) and nitric oxide. Conversely, glutamate, substance P and CGRP increase PG release. Prostaglandins also facilitate membrane currents and release of substance P and CGRP induced by low pH, bradykinin and capsaicin. All this should enhance elicitation and synaptic transfer of pain signals in the spinal cord. Direct administration of PGs to the spinal cord causes hyperalgesia and allodynia, and some studies have shown an association between induction of COX-2, increased PG release and enhanced nociception. NSAIDs diminish both basal and enhanced PG release in the spinal cord. Correspondingly, spinal application of NSAIDs generally diminishes neuronal and behavioral responses to acute nociceptive stimulation, and always attenuates behavioral responses to persistent nociception. Spinal application of specific COX-2 inhibitors sometimes diminishes behavioral responses to persistent nociception. PMID- 11275358 TI - Arginine metabolism and the synthesis of nitric oxide in the nervous system. AB - The biochemistry and physiology of L-arginine have to be reconsidered in the light of the recent discovery that the amino acid is the only substrate of all isoforms of nitric oxide synthase (NOS). Generation of nitric oxide, NO, a versatile molecule in signaling processes and unspecific immune defense, is intertwined with synthesis, catabolism and transport of arginine which thus ultimately participates in the regulation of a fine-tuned balance between normal and pathophysiological consequences of NO production. The complex composition of the brain at the cellular level is reflected in a complex differential distribution of the enzymes of arginine metabolism. Argininosuccinate synthetase (ASS) and argininosuccinate lyase which together can recycle the NOS coproduct L citrulline to L-arginine are expressed constitutively in neurons, but hardly colocalize with each other or with NOS in the same neuron. Therefore, trafficking of citrulline and arginine between neurons necessitates transport capacities in these cells which are fulfilled by well-described carriers for cationic and neutral amino acids. The mechanism of intercellular exchange of argininosuccinate, a prerequisite also for its proposed function as a neuromodulator, remains to be elucidated. In cultured astrocytes transcription and protein expression of arginine transport system y(+) and of ASS are upregulated concomittantly with immunostimulant-mediated induction of NOS-2. In vivo ASS-immunoreactivity was found in microglial cells in a rat model of brain inflammation and in neurons and glial cells in the brains of Alzheimer patients. Any attempt to estimate the contributions of arginine transport and synthesis to substrate supply for NOS has to consider competition for arginine between NOS and arginase, the latter enzyme being expressed as mitochondrial isoform II in nervous tissue. Generation of NOS inhibitors agmatine and methylarginines is documented for the nervous system. Suboptimal supply of NOS with arginine leads to production of detrimental peroxynitrite which may result in neuronal cell death. Data have been gathered recently which point to a particular role of astrocytes in neural arginine metabolism. Arginine appears to be accumulated in astroglial cells and can be released after stimulation with a variety of signals. It is proposed that an intercellular citrulline-NO cycle is operating in brain with astrocytes storing arginine for the benefit of neighbouring cells in need of the amino acid for a proper synthesis of NO. PMID- 11275359 TI - Safety factor at the neuromuscular junction. AB - Reliable transmission of activity from nerve to muscle is necessary for the normal function of the body. The term 'safety factor' refers to the ability of neuromuscular transmission to remain effective under various physiological conditions and stresses. This is a result of the amount of transmitter released per nerve impulse being greater than that required to trigger an action potential in the muscle fibre. The safety factor is a measure of this excess of released transmitter. In this review we discuss the practical difficulties involved in estimating the safety factor in vitro. We then consider the factors that influence the safety factor in vivo. While presynaptic transmitter release may be modulated on a moment to moment basis, the postsynaptic features that determine the effect of released transmitter are not so readily altered to meet changing demands. Different strategies are used by different species to ensure reliable neuromuscular transmission. Some, like frogs, rely on releasing a large amount of transmitter while others, like man, rely on elaborate postsynaptic specialisations to enhance the response to transmitter. In normal adult mammals, the safety factor is generally 3-5. Both pre- and postsynaptic components change during development and may show plasticity in response to injury or disease. Thus, both acquired autoimmune and inherited congenital diseases of the neuromuscular junction (NMJ) can significantly reduce, or even transiently increase, safety factor. PMID- 11275360 TI - Time-dependent promotion activity of 17beta-estradiol on uterine carcinogenesis in mice initiated with N-ethyl-N-nitrosourea. AB - The time-dependent promotion activity of 17beta-estradiol (E2) by initiation with N-ethyl-N-nitrosourea (ENU) on induction of mouse uterine endometrial proliferative lesions was examined. Illumination-induced persistent estrous female CD-1 mice were divided into five groups at 9 weeks of age. At 10 weeks of age, mice in all groups (n=25) were given a single intra-uterine administration of ENU (50 mg/kg), dissolved in polyethylene glycol. Animals in Groups 2 to 5 were then implanted s.c. with an E2 pellet at 9, 11, 14 and 17 weeks of age. The implants were left in place for 8 weeks and then taken out. At the termination of the experiment (week 15 after the ENU-treatment), all surviving mice were killed and the development of uterine proliferative lesions were assessed. All groups demonstrated endometrial hyperplasias and adenocarcinomas and the incidences of the latter in ENU plus E2 treated animals (Groups 2 to 5; 36, 48, 35 and 36%, respectively) were significantly higher compared to 8% for Group 1, without any variation with the age at E2 treatment. However, the incidences of adenocarcinomas plus severe hyperplasias increased from Groups 1 to 5 (28, 40, 56; P<0.05, 61; P<0.05 and 80%; P<0.01, respectively), indicating that promotion effects of E2 on induction of uterine proliferative lesions in the uterine endometrium become more pronounced with the interval after ENU initiation. PMID- 11275361 TI - Effect of ethanol on proliferation and estrogen receptor-alpha expression in human breast cancer cells. AB - There is substantial epidemiological evidence suggesting that alcohol consumption is associated with increased risk for breast cancer. However, possible biological mechanisms have not been clearly established. In the present studies, a direct effect of ethanol on the proliferation and intracellular content of cyclic AMP (cAMP) in two estrogen receptor-positive (ER+) and two estrogen receptor-negative (ER-) human breast cancer cell lines was examined. Treatment of ER+ human breast cancer cells (MCF-7 and ZR75.1) with ethanol at concentrations between 10 and 100 mM was associated with increased cell numbers compared to controls. The ERalpha content and the amount of intracellular cAMP also increased in ER+ cells exposed to ethanol, compared to controls. On the other hand, ethanol treatment did not increase cell proliferation or cAMP levels in the ER- (BT-20 and MDA-MB-231) human breast cancer cells. Therefore, ethanol added at physiologically relevant concentrations to ER+ human breast cancer cell cultures can enhance cell proliferation and increase the content of ERalpha. PMID- 11275362 TI - Capsaicin-induced apoptosis in SK-Hep-1 hepatocarcinoma cells involves Bcl-2 downregulation and caspase-3 activation. AB - Hepatocellular carcinoma is one of the most lethal malignancies and there is no effective preventive measure in this highly malignant disease to date. In the present study, we investigated the chemopreventive potential of capsaicin (8 methyl-N- vanillyl-6-nonenamide), the principal pungent ingredient found in hot red pepper, in SK-Hep-1 hepatocellular carcinoma cells. Treatment of capsaicin inhibited growth of SK-Hep-1 cells in a concentration-dependent manner while 4 methoxy capsaicin (Met-capsaicin) was less potent. This inhibitory effect of capsaicin on SK-Hep-1 cell growth was mainly due to the induction of apoptosis as evidenced by DNA fragmentation and nuclear condensation. Furthermore, capsaicin prominently reduced the ratio of anti-apoptotic Bcl-2 to pro-apoptotic Bax and consequently increased caspase-3 activity. These results demonstrate that capsaicin efficiently induced apoptosis in SK-Hep-1 cells through a caspase-3 dependent mechanism, which may contribute to its chemopreventive function. PMID- 11275363 TI - Taxol-induced cell cycle arrest and apoptosis: dose-response relationship in lung cancer cells of different wild-type p53 status and under isogenic condition. AB - The effective dose, schedule, molecular basis of the cytotoxicity of taxol and their dependence on the genetic background in tumor cells are still not well understood. Here, we examined how the dose-response relationship for taxol varies in lung cancer cells with different p53 status and under isogenic conditions. DNA content analyses in A 549 (p53, +/+) and H 1299 (p53, -/-) cells, showed that taxol progressively induced G2/M arrest in both cell lines in a concentration dependent manner, which was accompanied by a parallel decrease in the G1 population. G2/M arrest, however, occurred at a lower concentration in A 549 cell lines than in H 1299 cells. The S-phase population in A 549 cells was not significantly changed up to 0.025 microM, but dropped by six-fold at 1.0 microM taxol, in contrast to that in H 1299 cells. A sub-G1 apoptotic population was present at 24 h, even at 0.002 microM taxol, when G2/M arrest was not appreciably detected. In both cell lines, the maximum apoptosis of about 28% was achieved at 0.025 microM taxol, implicating that wild-type p53 does not modulate the level of taxol-induced apoptosis. When we examined the role of the wild-type p53 in isogenic cell lines developed in a H 1299 background, the maximum level of apoptosis was in the range of 28-34% at a drug concentration around 0.03 microM, not significantly different from that observed in parental H 1299 cells. We conclude that taxol is effective in inducing apoptosis at very low doses (0.020 0.035 microM), and that the presence or absence of the wild-type p53 does not make a statistically significant difference in the level of apoptotic cell death in these lung cancer cell lines, but the maximum is attained at a lower drug concentration in the presence of p53. PMID- 11275364 TI - Detection of early gastric cancer by a real-time autofluorescence imaging system. AB - A light-induced fluorescence endoscopy in the gastrointestinal tract system was used in 52 patients with 54 lesions (33 early gastric cancers, 21 benign lesions) to assess its ability to detect early gastric cancer. Comparing the images with the histological findings, 21 of the 33 carcinomas appeared dark red, ten had a mixed pattern of dark red and white, and two could not be detected. Of the 21 benign lesions, 18 appeared light blue, as do normal mucosa, with this system. In 85% of the cancer lesions (28/33), cancer extension was correctly detected. The sensitivity and specificity were 94 and 86%, respectively. Real-time autofluorescence endoscopy is a useful adjunct to conventional white-light endoscopy for detecting early gastric cancer. PMID- 11275365 TI - Inhibitory effect of a matrix metalloproteinase inhibitor on growth and spread of human pancreatic ductal adenocarcinoma evaluated in an orthotopic severe combined immunodeficient (SCID) mouse model. AB - The present study was aimed at evaluating the effect of the matrix metalloproteinase (MMP) inhibitor prinomastat (AG3340) on tumor progression using an orthotopic pancreatic carcinoma model in severe combined immunodeficient mice. In controls, receiving vehicle only, the poorly differentiated ductal adenocarcinoma invaded into adjacent organs and metastasized to different sites in the abdomen and to the lungs. Treatment with prinomastat, intraperitoneally twice daily for 21 days, reduced primary tumor volume significantly to 19.0 (+/ 7.7)% of control, with induction of necrosis, differentiation, and fibrotic tissue in the pancreatic tumors. Invasion was not observed in 63% of prinomastat treated mice, and metastases were reduced markedly. Surprisingly, prinomastat treated tumors had on average higher microvessel densities as a consequence of an increased angiogenesis in perinecrotic tumor areas. We conclude that prinomastat is highly effective in inhibiting pancreatic carcinoma growth and progression in an orthotopic cancer model. This model appears to be a valuable tool to investigate the potency of novel antimetastatic strategies in pancreatic ductal adenocarcinoma by specifically targeting certain MMPs. PMID- 11275366 TI - Genetic polymorphisms in cytochrome P450 (CYP) 1A1, CYP1A2, CYP2E1, glutathione S transferase (GST) M1 and GSTT1 and susceptibility to prostate cancer in the Japanese population. AB - Associations between genetic polymorphisms of CYP1A1, CYP1A2, CYP2E1, GSTM1 and GSTT1 and prostate cancer (PCa) were analyzed in a case-control study of 315 individuals. The frequency of valine (Val)/valine (Val) genotypes for CYP1A1 was 11.3% in cases compared with 5.5% in controls, this polymorphism thus being associated with a significantly increased risk of PCa (odds ratio=2.4, 95% confidence interval (CI)=1.01-5.57). No links were detected between PCa and polymorphisms in other enzymes. However, the combination of CYP1A1 (Ile/Val and/or Val/Val) polymorphisms with the GSTM1 null type resulted in an OR of 2.2 (CI=1.10-4.57, 1.12-4.20, respectively). This study suggests that the CYP1A1 polymorphism and its combination with GSTM1 may be associated with PCa susceptibility in the Japanese population. PMID- 11275368 TI - Id1 expression is associated with histological grade and invasive behavior in endometrial carcinoma. AB - Basic helix-loop-helix (bHLH) DNA-binding proteins have been reported to regulate tissue-specific transcription of cellular differentiation within multiple cell lineages. The Id family of helix-loop-helix proteins does not possess a basic DNA binding domain and functions as a negative regulator of bHLH proteins by forming high-affinity heterodimers with bHLH proteins. Id proteins were originally characterized as inhibitors of DNA binding and cell differentiation. Thus, overexpression of Id proteins correlates with cell proliferation and arrested differentiation in many cell lineages. To elucidate the involvement of Id1 in endometrial carcinogenesis, we analyzed serial frozen sections for Id1 protein expression in 20 cases of endometrial carcinoma and 20 cases of normal endometria by fluorescent immunohistochemistry. We analyzed the relationship between the percentages of Id1-stained cells and the patient's characteristics, including histological grade, clinical stage, presence of invasion to >1/2 myometrium, and clinical outcome. In normal endometria, Id1 was not detected in either the proliferative or the secretory phase. There was, however, abundant Id1 immunoreactivity in the endometrial carcinoma cells. Moreover, Id1 was strongly expressed in the inflammatory cells. Scoring on the basis of the percentage of positive cells indicated that Id1 expression is significantly associated with histological grade (P<0.05) and the presence of invasion to >1/2 myometrium (P<0.05). Our results demonstrate that increased Id1 expression in endometrial carcinoma correlates with the malignant potential of this tumor. PMID- 11275367 TI - Humanized anti-ganglioside GM2 antibody is effective to induce antibody-dependent cell-mediated cytotoxicity in mononuclear cells from lung cancer patients. AB - Ganglioside GM2 is one of the major gangliosides expressed on the cell surface of human tumors including lung cancer. We have previously reported that a mouse human chimeric monoclonal antibody (mAb), KM966, against GM2 promotes the lysis of lung cancer cells by human blood mononuclear cells (MNC) of healthy donors. In this study, we examined antibody-dependent cell-mediated cytotoxicity (ADCC) of MNC, using KM966 mAb and its humanized counterpart, KM8969, in 16 lung cancer patients and 18 control patients. The ADCC activity was assessed by 4-h (51)Cr release from GM2 positive SBC-3 small cell lung cancer cells. MNC from lung cancer patients exhibited similar ADCC activity to those from control patients when KM966 and KM8969 were used as mAb. Moreover, effective ADCC activity was observed even in MNC from advanced lung cancer patients. These observations suggest the potential activity of humanized anti-GM2 mAb (KM8969), as well as chimeric KM966, in biological therapy for lung cancer patients. PMID- 11275369 TI - Deoxynucleoside anabolic enzyme levels in acute myelocytic leukemia and chronic lymphocytic leukemia cells. AB - The deoxynucleoside kinase reaction is often rate-limiting in the anabolism of pharmacologically active anti-cancer nucleosides. The levels of thymidine kinase (TK), deoxycytidine kinase, deoxyguanosine kinase (dGK), and thymidylate kinase were determined in leukocyte extracts from patients with chronic lymphocytic leukemia (CLL) and acute myelocytic leukemia (AML). The extracts from AML patients showed significantly higher TK activity than the ones from CLL patients. There were no differences in the levels of the other three kinases. In the case of dGK, the determinations were carried out with both an immunoblotting assay and selective enzyme activity measurements. PMID- 11275370 TI - Oligonucleotide microarray expression analysis of genes whose expression is correlated with tumorigenic and non-tumorigenic phenotype of HeLa x human fibroblast hybrid cells. AB - In order to understand the differences and similarities between tumorigenic and non-tumorigenic HeLaxhuman fibroblast hybrids, gene expression profiles were examined with synthetic oligonucleotide arrays containing nearly 7000 gene probe sets. We used two pairs of genetically related hybrids, each pair representing individual clones of non-tumorigenic and tumorigenic segregant hybrids, respectively. Analysis of six possible comparisons, utilizing two algorithms, identified 204 genes with differential expression. The greater number of differentially expressed genes was observed when non-tumorigenic hybrids were compared with tumorigenic segregants. Fifteen and 14 genes, respectively, were consistently found to be differentially expressed in non-tumorigenic and tumorigenic cells. Among those 29 differentially expressed genes, three (intestinal alkaline phosphatase, caveolin-1, and solute carrier family2, member3) have been reported previously to be associated with the tumorigenic phenotype, using the same hybrid pairs. In addition, among the genes previously detected by differential display, 78% of them exhibited more than 5-fold change, demonstrating a high consistency between the two methods of differential gene expression. These findings suggest that synthetic oligonucleotide arrays are a powerful and highly reproducible tool to identify those genes whose expression is associated with certain phenotypes. PMID- 11275371 TI - Key peptide processing enzymes are expressed by breast cancer cells. AB - The expression of the three key peptide processing enzyme families, represented by CPE, PAM, and PC1/3 plus PC2, were examined in MCF-7 and ZR-75-1 breast cancer cell lines. Both of these cell lines express vasopressin receptors as well as the vasopressin gene, but the processing of vasopressin gene-related proteins appears to be limited. Products of the expected size for, CPE, PAM and PC1/PC3 could be amplified by reverse transcription-polymerase chain reaction (RT-PCR) from both cell lines. Cloning and sequencing of these RT-PCR products revealed that each enzyme mRNA had a structure identical to that published for the human form of the respective enzyme. Western analysis provided evidence that mRNAs for these enzymes are translated into proteins. Alternatively, PC2 mRNA was identified to be present in MCF-7 cells both by RT-PCR and Western blot analysis, but could not be demonstrated for ZR-75-1 cells. Our findings suggest that the key processing enzymes needed to generate active vasopressin and other neuropeptide growth factors are present in breast cancer cells. PMID- 11275372 TI - Different kinds of reactive oxygen and nitrogen species were detected in colon and breast tumors. AB - Several studies have shown the involvement of reactive oxygen species (ROS; O2*-, hypochlorite, hydroxyl radical, hydrogen peroxide) in carcinogenesis. With certain pathologies, nitric oxide (NO) is formed and can interact with superoxide radical (O2*-) resulting in the propagation of the highly reactive species, peroxynitrite. In order to study the molecular mechanisms underlying the ability of reactive oxygen and nitrogen species (RONS) to mediate carcinogenesis, we have measured ROS, NO, and peroxynitrite content of cancerous tissues obtained from colon and breast carcinoma cases by chemiluminescence technique. All ROS were significantly increased in cancerous colon tissues with hypochlorite making the most important contribution and suggesting the role of inflammatory cells. NO was also increased and the peroxynitrite concentration was higher in cancerous samples. For breast carcinoma cases, only O2*- was significantly increased. Hypochlorite was not detected excluding the contribution of inflammatory cells. NO concentrations were not significantly different, therefore, ROS might originate by change in the redox state of the tissue. PMID- 11275374 TI - Botulinum toxin A in the treatment of headache syndromes and pericranial pain syndromes. PMID- 11275375 TI - Mrz 2/579, a fast kinetic NMDA channel blocker, reduces the development of morphine tolerance in awake rats. AB - The purpose of the present study was to investigate whether uncompetitive NMDA antagonists with fast channel blocking kinetics, which show fewer side effects in man than compounds such as ketamine, affect the development of tolerance to continuous exposure to morphine. Rats were trained on the Randall--Selitto apparatus before being implanted, under halothane anaesthesia, with primed mini osmotic pumps (240 microl/day). Six rats were implanted with a vehicle filled pump, seven with a morphine filled pump (28.8 mg/kg/day), and eight with a pair of pumps, one containing morphine and the other Mrz 2/579, a new NMDA antagonist (40 mg/kg/day). A fourth group was implanted with a morphine filled pump followed 25 h later by a Mrz 2/579 filled pump. Paw withdrawal tests were undertaken immediately before, and at 2, 4, 6, 8, 10, 12, 24, 48 and 72 h after the first pump was implanted. Before pump implantation, withdrawal thresholds were 120+/-7 g (mean+/-SEM, n=30). Vehicle infusion had no effect on withdrawal thresholds, whereas morphine infusion increased them significantly at 2 and 4 h after pump implantation (+2 h: 208+/-14 g; P<0.001 vs. control). From 6 h the antinociception elicited by morphine declined progressively; at 10 h withdrawal thresholds were significantly lower than the 2 h post-treatment value (P<0.001). In rats treated with morphine plus Mrz 2/579, thresholds remained significantly higher between 10--72 h post-implantation than with morphine alone (P<0.05). In contrast, infusion of the same level of Mrz 2/579 once tolerance had developed did not reverse tolerance. These results indicate that fast NMDA channel blockers such as Mrz 2/579 may prove to be useful in enhancing analgesia to continuous morphine administration. PMID- 11275376 TI - NK-1 receptor gene expression is related to pain in chronic pancreatitis. AB - Recent theories of pathogenesis of pain in chronic pancreatitis (CP) are neuroimmune interactions of intrapancreatic nerves and inflammatory cells and increase in levels of pain neurotransmitters such as substance P (SP). This study analyzed the expression and localization of neurokinin 1 receptor (NK-1R), which binds SP, and its association with pain and inflammation in CP. Pancreatic tissues from 31 patients (22 males, nine females; mean age 45.9+/-9.4 years) with CP were evaluated. Nine normal pancreases (five males, four females; mean age 42.9+/-9.5 years) served as controls. Quantitative PCR was used to determine the NK-1R mRNA expression levels and in situ hybridization and immunohistochemistry were used to localize expression sites of NK-1R mRNA and protein, respectively. We also analyzed whether an association exists between NK-1R mRNA expression and pain and inflammation. In CP samples, in situ hybridization and immunohistochemistry localized NK-1R mRNA expression and protein mainly in the nerves, ganglia, blood vessels, inflammatory cells and occasionally in fibroblasts. In patients with mild to moderate and strong intensity of pain, NK 1R mRNA levels were increased 14- and 30-fold over controls, respectively. There was a significant relationship between NK-1R mRNA levels and intensity of pain (r=0.46, P=0.03), NK-1R mRNA and the frequency of pain (r=0.51, P=0.04), and NK-1 mRNA and duration of pain (r=0.46, P=0.01) in CP patients, but not with the degree of tissue inflammation. NK-1R signaling may be involved in the pain syndrome of CP. The expression of NK-1R in inflammatory cells and blood vessels also points to an interaction of immunoreactive substance P nerves, inflammatory cells and blood vessels, and further supports the existence of a neuroimmune interaction that probably influences the pain syndrome and chronic inflammatory changes so characteristic of CP. PMID- 11275377 TI - Capsaicin-evoked release of immunoreactive calcitonin gene-related peptide from rat trigeminal ganglion: evidence for intraganglionic neurotransmission. AB - Chemically-mediated cross-excitation has been described between neurons within sensory ganglia. However, the identity and source of the chemical mediators is not known. Ca(2+)-dependent release of neurotransmitters from cultured sensory neurons in vitro has been observed, although neurite outgrowth may confound the ability to extrapolate findings from culture systems to in vivo conditions. Thus, the present studies evaluate the hypothesis of capsaicin-sensitive intraganglionic neuropeptide release from freshly prepared slices of rat sensory ganglia. The ganglionic slice preparation provides an advantage over neuronal cultures, because release may be assessed within minutes after tissue collection (minimizing phenotypic changes) and while maintaining gross anatomical relationships. Trigeminal ganglia (TGG) were quickly removed from male, Sprague- Dawley rats (175--200 g), chopped into 200 microm slices and placed into chambers within 3 min of collection. Chambers were perfused with buffer, and superfusates were collected and assayed for immunoreactive calcitonin gene-related peptide (iCGRP) release via radioimmunoassay. After about 90 min of baseline collection, tissue was treated with capsaicin followed by a washout period. Capsaicin (1--100 microM) evoked concentration-dependent increases in iCGRP release. A competitive capsaicin receptor antagonist, capsazepine, significantly inhibited capsaicin evoked release of iCGRP. In addition, capsaicin-evoked release of iCGRP was dependent on the presence of extracellular calcium. Furthermore, capsaicin-evoked release from TGG slices was significantly greater than that from slices of equivalent weights of adjacent trigeminal nerve shown histologically to be free of neuronal somata. These data support the hypothesis that Ca(2+)-dependent exocytosis of neuropeptides may occur within the TGG in vivo and that the majority of this release derives from neuronal somata. PMID- 11275378 TI - Alternative diagnostic criteria for major depressive disorder in patients with chronic pain. AB - Chronic pain is associated with high rates of major depressive disorder (MDD), but somatic symptoms caused by pain may complicate the diagnosis of MDD. Different methods to address this issue include the adoption of an inclusive approach to diagnosis (i.e. including all symptoms when assessing MDD, regardless of their presumed cause), an etiologic approach (i.e. disregarding symptoms that are caused by medical problems), and a substitutive approach (i.e. replacing somatic symptoms with non-somatic alternatives). In this study, 129 patients with chronic pain (56 men and 73 women) underwent semi-structured interviews addressing 23 individual symptoms of MDD. Detailed probing was undertaken into patients' perceptions of the causes of those symptoms that could potentially be brought on by pain. We found that the prevalence of MDD was highest with the inclusive diagnostic method (35.7%), lowest with an etiologic approach that discounted symptoms based on patient attributions (19.4%), and intermediate with the substitutive method (30.3%). Although some symptoms, such as insomnia, fatigue, and difficulty concentrating, were reported by 34--53% of the patients who did not meet criteria for MDD, they were still more common among those who did (85--94%, P<0.001). Patients who met criteria for MDD with the inclusive method, but who did not meet criteria using the etiologic method, had Beck Depression Inventory scores (M=24.5) that were comparable to those of patients who were consistently classified with MDD across methods (M=25.6). These scores were much higher than those of patients who were consistently classified without MDD (M=13.8, P<0.001). In conclusion, excluding criterion symptoms that patients attribute to pain can reduce the observed prevalence of MDD by about 45%. However, this method introduces a problem of false-negative diagnoses that appears to be more significant than the problem of false positives associated with the inappropriate inclusion of somatic symptoms. PMID- 11275379 TI - The effect of conduction block on the spinal release of immunoreactive neuropeptide Y (ir-NPY) in the neuropathic rat. AB - Peripheral nerve injury may result in significant changes in neuropeptide production and the development of neuropathic pain behaviour. Rats with a chronic constriction injury of one sciatic nerve were used to study the spinal release of immunoreactive neuropeptide Y (ir-NPY), using the antibody-coated microprobe technique. Previous work has shown an increase in NPY synthesis by large to medium-sized primary afferent neurones, as well as a new area of ir-NPY release in the deep dorsal horn on the side of nerve injury, when compared to uninjured rats. The stimulus for spontaneous ir-NPY release was unclear, but may have been due to ectopic neuronal discharges developing after nerve injury. This study used local anaesthetic to block all electrical input from the injured nerve. No change was found in the new zone of spontaneous release of ir-NPY in the deep dorsal horn ipsilateral to nerve injury. It appears therefore, that ir-NPY is released from the central termination of primary afferent neurones, without regulation from neuronal activity in the primary afferent neurones themselves. PMID- 11275380 TI - Influence of thermode size for detecting heat pain dysfunction in a capsaicin model of epidermal nerve fiber loss. AB - Quantitative sensory testing of heat pain sensation has become an important tool to evaluate small caliber afferent nerve function in peripheral neuropathy. In earlier studies, we found that topical application of capsaicin in humans results in the loss of epidermal nerve fibers (ENFs) with a corresponding decrease in detection of heat pain sensation. Capsaicin may therefore be a useful model for developing optimal psychophysical testing procedures for detection of neuropathy in its early stages. Here we determined the influence of thermal probe (thermode) size in detecting the diminished heat pain sensation following capsaicin application. Twelve healthy volunteers applied 0.075% capsaicin topically to the volar forearm four times daily for 7 days. Psychophysical measures of heat pain, mechanical (sharp) pain, and tactile threshold were obtained daily from untreated control skin and from capsaicin-treated skin during capsaicin application, and once weekly for 5 weeks following discontinuation of capsaicin. Heat pain sensation was assessed using a large (30 x 30 mm) and small (3 x 3 mm) thermode and different algorithms to assess pain threshold and suprathreshold heat pain. Skin biopsies were obtained and were processed for immunohistochemical localization of (ENFs) using the pan neuronal marker protein gene product 9.5. Capsaicin produced a rapid decrease in the number of ENFs, with nearly complete disappearance after 3 days of treatment. Heat pain evoked by the small, but not the large, thermode decreased dramatically after capsaicin treatment. The sensation of heat pain returned toward normal after 2--3 weeks following discontinuation of capsaicin treatment concordant with gradual reinnervation of the epidermis. Regression analysis indicated that the sensation of heat pain evoked by the small thermode correlated much better with the number of ENFs than heat pain evoked by the large thermode. The detection of sharp pain decreased moderately after capsaicin treatment. Assessment of heat pain sensation using small thermodes has potential for detecting sensory deficits in early stages of small fiber neuropathy. PMID- 11275381 TI - Facilitated neurogenic inflammation in complex regional pain syndrome. AB - Complex regional pain syndrome (CRPS) is characterized by a variety of clinical features including spontaneous pain and hyperalgesia. Increased neuropeptide release from peripheral nociceptors has been suggested as a possible pathophysiologic mechanism triggering the combination of trophic changes, edema, vasodilatation and pain. In order to verify the increased neuropeptide release in CRPS, electrically induced neurogenic vasodilatation and protein extravasation were evaluated in patients and controls. We performed a prospective study on 10 patients with acute and untreated CRPS and 10 matched healthy controls. Neurogenic inflammation was elicited by strong transcutaneous electrical stimulation via intradermal microdialysis capillaries which simultaneously enabled measurement of protein extravasation. Laser-Doppler scanning was used to assess axon reflex vasodilatation. Axon reflex vasodilatation was significantly increased in CRPS patients (438 +/- 68% of baseline vs. 306 +/- 52%; P < 0.05) and transcutaneous electrical stimulation provoked protein extravasation only in the patients (before, 0.28 +/- 0.03 mg/ml; during stimulation, 0.34 +/- 0.04 mg/ml), whereas protein concentration steadily declined during stimulation in the healthy controls (before, 0.23 +/- 0.04 mg/ml; during stimulation, 0.18 +/- 0.04; P < 0.001). The time course of electrically induced protein extravasation in the patients resembled the one observed following application of exogenous substance P (SP). We conclude that neurogenic inflammation is facilitated in CRPS. Our results suggest an increased releasability of neuropeptides in CRPS. PMID- 11275382 TI - Innate cytokine profile in patients with complex regional pain syndrome is normal. AB - Complex regional pain syndrome (CRPS) is a disabling disease characterized by the classic symptoms and signs of inflammation. In this study we investigated the innate cytokine profile in patients with CRPS to determine a possible role of the immune system in the pathophysiology of CRPS. The cytokine profile before and after lipopolysaccharide and thrombin stimulation was determined in 26 severely affected CRPS patients and 20 healthy controls. No difference in the production of pro- and anti- inflammatory cytokines between patients and controls was found. Hence, our results do not support a role of genetic factors responsible for the cytokine profile in the pathophysiology of CRPS. These findings encourage further investigations of mechanisms responsible for neurogenic-induced inflammation. PMID- 11275383 TI - Induction of long-term potentiation of single wide dynamic range neurones in the dorsal horn is inhibited by descending pathways. AB - Previous studies have shown that long-term potentiation (LTP) in the dorsal horn may be induced by noxious stimuli. In this study it is investigated whether induction of LTP in the dorsal horn may be affected by the descending pathways. Extracellular recordings of wide dynamic range (WDR) neurones in the lumbar dorsal horn in intact urethane-anaesthetized Sprague--Dawley rats were performed, and the electrically evoked neuronal responses in these neurones were defined as A-fibre and C-fibre responses according to latencies. Using a short-term cold block of the thoracic spinal cord, which produced a completely reversible increase of the A-fibre and C-fibre responses, the influence of the descending inhibitory system on the induction of LTP by electrical high-frequency conditioning applied to the sciatic nerve was examined. As previously shown the A fibre responses were almost unchanged following the conditioning. In contrast, the C-fibre responses following the same conditioning were strongly increased. Thus, a clear LTP of the nociceptive transmission in the dorsal horn was observed following electrical high-frequency conditioning. Interestingly, we found that the LTP was more powerful when the effects of the descending pathways were temporarily eliminated during conditioning. It is concluded that induction of LTP by electrical high-frequency conditioning stimulation, which may be part of the wider term central sensitization, is inhibited by descending pathways. PMID- 11275384 TI - Analgesic effect of bisphosphonates in mice. AB - Bisphosphonates are analogues of inorganic pyrophosphate and are inhibitors of bone resorption. Many derivatives have been developed for the treatment of enhanced bone resorption; several reports reveal that treatment with bisphosphonates is able to reduce the pain associated with different painful diseases. This study tested the antinociceptive action of four bisphosphonates, clodronate, alendronate, pamidronate and etidronate, in comparison with that of morphine and acetylsalicylic acid using two algesimetric tests in mice, tail flick and writhing tests. In the tail-flick test, after intravenous (i.v.) injection, a dose-dependent antinociception was present after pamidronate, clodronate and acetylsalicylic acid whereas etidronate and alendronate produced an analgesic effect only with the highest dose tested. We also studied the central effect of clodronate and pamidronate and, after intracerebroventricular injection, both bisphosphonates showed a dose-dependent antinociceptive effect. In the writhing test clodronate and pamidronate showed a statistically significant antinociceptive action after i.v. and intramuscular administration. To verify if clodronate and pamidronate could modulate the peripheral opioid receptors we evaluated the gastrointestinal transit time in mice, but we did not find any effect on the gastrointestinal motility. These data indicate that clodronate and pamidronate present a central and peripheral antinociceptive effect; however, the main mechanism cannot be determined from the present data. We discuss the possible pharmacological hypothesis to interpret the present results. The findings suggest a pharmacological role of the bisphosphonates in the modulation of antinociception even in acute conditions not related to accelerated osteolytic and inflammatory response, with a possible clinical application to control pain. PMID- 11275385 TI - Electronic diaries for monitoring chronic pain: 1-year validation study. AB - Electronic data collection for monitoring pain has become increasingly popular in clinical research. However, no direct comparison has been made between electronic diaries and self-report paper diaries or phone interviews. We asked 36 patients with chronic low back pain to monitor their pain for 1 year; 20 of them used both a palmtop computer and paper diaries, and 16 used paper diaries alone. All patients were called once a week and asked to rate their pain. Regression analyses with a measurement error model were run on hourly pain scores recorded by both palmtop computer and paper diaries. Ratings of pain intensity were highly reliable between data recorded with a palmtop computer and with data from paper diaries. Patients who monitored their pain with the palmtop computer entered data on average 6.75 times a week and were 89.9% compliant with daily monitoring throughout the year. Two-way messaging available through the palmtop computer seemed to encourage continued use of the device. Internal consistency of reporting and correlations with phone reports and standardized measures were highly significant, suggesting that data from electronic diaries are both reliable and valid. Patients using electronic diaries preferred them to paper diaries and showed much higher rates of compliance and satisfaction over the 1 year trial. PMID- 11275386 TI - Activation of diffuse noxious inhibitory controls (DNIC) in rats with an experimental peripheral mononeuropathy. AB - Diffuse noxious inhibitory controls (DNIC), which involve supraspinal structures and modulate the transmission of nociceptive signals, were investigated in rats with chronic constriction injury of the sciatic nerve. Nerve-injured rats with increased sensitivity to mechanical and thermal stimulation on the operated side were anesthetized and recordings were made from trigeminal convergent neurons. Inhibitions of C-fiber-evoked neuronal responses during and after the application of nociceptive conditioning stimuli to the hindpaw, were measured to evaluate DNIC. The conditioning stimuli consisted of graded natural (pressure and heat) and electrical stimuli and were applied alternately to non-operated and operated hindpaws. Compared with the non-operated paw, inhibitions elicited by pressure on the operated hindpaw were increased significantly at all stimulus intensities. Albeit to a lesser extent, inhibitions elicited by thermal stimulation of the operated paw were also increased in the nerve-injured animals. Such exacerbation of DNIC-induced inhibitions produced by mechanical and thermal stimulation of the operated paw can be explained by an increase in the afferent input to the spinal cord. In contrast to the results obtained with natural stimulations, inhibitions evoked from the operated and non-operated paws were similar when graded electrical stimulation was used as the conditioning stimulus. This was true regardless of the intensity and frequency of stimulation and regardless of whether the stimuli were applied transcutaneously or directly to the sciatic nerve. The clear-cut difference between the results obtained with natural and electrical conditioning stimuli suggests that the nociceptive neurons involved in the triggering of DNIC may not be sensitized at the central level. Peripheral mechanisms such as the sensitization of nerve injured fibers and/or sprouting of nerve terminals may thus be the main causes of DNIC increase in this model of neuropathic pain. PMID- 11275387 TI - A randomized comparison of group cognitive--behavioral therapy, surface electromyographic biofeedback, and vestibulectomy in the treatment of dyspareunia resulting from vulvar vestibulitis. AB - This study compared group cognitive-behavioral therapy (12-week trial), surface electromyographic biofeedback (12-week trial), and vestibulectomy in the treatment of dyspareunia resulting from vulvar vestibulitis. Subjects were 78 women randomly assigned to one of three treatment conditions and assessed at pretreatment, posttreatment and 6-month follow-up via gynecological examinations, structured interviews and standard questionnaires pertaining to pain (Pain Rating Index and Sensory scale of the McGill Pain Questionnaire, vestibular pain index, pain during intercourse), sexual function (Sexual History Form, frequency of intercourse, Information subscale of the Derogatis Sexual Functioning Inventory), and psychological adjustment (Brief Symptom Inventory). As compared with pretreatment, study completers of all treatment groups reported statistically significant reductions on pain measures at posttreatment and 6-month follow-up, although the vestibulectomy group was significantly more successful than the two other groups. However, the apparent superiority of vestibulectomy needs to be interpreted with caution since seven women who had been assigned to this condition did not go ahead with the intervention. All three groups significantly improved on measures of psychological adjustment and sexual function from pretreatment to 6-month follow-up. Intent-to-treat analysis supported the general pattern of results of analysis by-treatment-received. Findings suggest that women with dyspareunia can benefit from both medical and behavioral interventions. PMID- 11275388 TI - Encoding of noxious stimulus intensity by putative pain modulating neurons in the rostral ventromedial medulla and by simultaneously recorded nociceptive neurons in the spinal dorsal horn of rats. AB - Neurons in the nucleus raphe magnus and adjacent structures of the rostral ventromedial medulla (RVM) are involved in the control of nociceptive transmission. In the RVM the so-called on-cells are excited, and the so-called off-cells are inhibited, by noxious stimuli applied almost anywhere on the body surface, thus showing that they receive information from spinal and trigeminal nociceptive neurons. In deeply anesthetized rats, recordings were made from RVM neurons that resembled on- and off-cells (herein called putative on- and off cells) in order to investigate (1) how they encode the intensity of thermal noxious stimuli (46--56 degrees C) applied to a hindpaw, and (2) how their encoding properties relate to those of simultaneously recorded spinal neurons. In 49 of 98 cases, a graded increase in the stimulus temperature caused a monotonic decrease in the response latency of putative on-cells, putative off-cells and spinal neurons, while the response discharge rate monotonically increased for putative on-cells and spinal neurons and decreased for putative off-cells. In the majority of simultaneous recordings of RVM and spinal neurons, the latency and discharge rate of the putative on- or off-cell were highly correlated with the latency and discharge rate of the spinal neuron, and the stimulus/response slopes were similar. These results show that putative on- and off-cells can encode the stimulus intensity in terms of response latency and discharge rate, and suggest that such encoding closely reflects spinal neuronal encoding. This may be relevant for the transmission and modulation of pain information by RVM neurons. PMID- 11275389 TI - Pain site and the effects of amputation pain: further clarification of the meaning of mild, moderate, and severe pain. AB - Research among persons with cancer pain suggests that the association between pain intensity and pain interference is non-linear. That is, pain begins to have a serious impact on functioning when it reaches a certain threshold level (about 5 on 0--10 scales). Often, a second pain threshold can be identified which, once reached, shows an even greater proportional negative impact on functioning. This finding supports the potential clinical utility of classifying pain as mild, moderate, and severe based on the impact of pain on quality of life, and research among persons with cancer pain supports specific cutoffs (mild: 1--4, moderate: 5 -6, severe: 7--10, see Pain 61 (1995) 277) for this classification. The current study sought to replicate the non-linear association between pain and pain interference in a non-cancer pain sample, determine whether the cutoffs that have been identified as optimal for cancer patients are also optimal for persons with pain associated with amputation, and determine whether the optimal cutoffs replicate across pain types (in this case, phantom limb, back, and general pain) within a single sample. Two-hundred and five persons with acquired amputation and phantom limb pain, back pain, or both, rated their average pain intensity and degree of pain interference for each type of pain. The results support a non linear association between pain intensity and pain interference. However, the optimal cutoffs for classifying mild, moderate and severe pain in the present sample replicated the findings for persons with cancer pain only for back pain -- different optimal cutoffs were found for phantom limb and general pain. Moreover, the degree of pain interference appeared to vary as a function of pain type. The same level of back pain interfered more significantly with daily function than phantom limb pain did after pain levels reached five or more (on a 0--10 scale). These findings have implications for understanding the meaning of pain intensity levels, as well as for the assessment of pain intensity in persons with amputation-related pain. PMID- 11275390 TI - TMJ disorders and myogenic facial pain: a discriminative analysis using the McGill Pain Questionnaire. AB - Our aim was to assess the discriminative capacity of the McGill Pain Questionnaire (MPQ) in patients with temporomandibular joint (TMJ) disorders or with myogenous facial pain (MP). The MPQ was administered to 57 TMJ and 28 MP patients who were also asked to assess the level of pain using the Visual Analog Scale (VAS). Weighted MPQ item scores, subscale Pain Rating Indexes (PRI), total PRI and the number of words chosen were calculated. Mean scores were tested for significant differences (Student's t-test), and the frequency with which each descriptor was chosen by the patients in both groups was analyzed. Data were also processed through two systems based on a counter-propagation neural network: the Self-Organizing Map (SOM) system, and a cluster-like analysis. In the MP group, 16 out of 20 mean MPQ item scores and all mean PRI and VAS scores were significantly higher than those in the TMJ group. There was a marked difference in descriptor choice. In the TMJ group the following descriptors were chosen by 25% or more of the patients: tiring, troublesome, nagging, sore, tender, and aching. In the MP group the descriptors most frequently chosen were: 'exhausting' (57%), 'punishing' (50%), and pulling (47%). SOM analysis distributed the two groups in the two halves of the map: only two out of 28 MP cases (7%) and 12 out of 57 TMJ cases (21%) were misplaced. The cluster-like analysis based on the 20 MPQ item scores correctly recognized 94.73% of TMJ patients and 89.28% of MP patients. In conclusion, the MPQ consistently discriminated between TMJ and MP patients. Although the higher affective scores in the MP patients may be partly induced by higher levels of anxiety in these patients, the data convincingly show that the system's discriminative capacity relates to all MPQ subscores and to the majority of the MPQ items. Moreover, within the same item, the choice of verbal descriptors varies consistently between the two groups of patients. PMID- 11275391 TI - The importance of stimulus site and intensity in differences of pain-induced vascular reflexes in human orofacial regions. AB - Studies in anaesthetized animals have indicated that noxious stimulation may produce marked blood flow changes in various orofacial structures, but the influence of painful stimulation on the blood flow regulation of the orofacial area of humans has been studied only to a limited extent. The purpose of this investigation was to study whether there are differences in temporal and spatial patterns of pain-induced vasoactive reflexes between various orofacial regions and hand in healthy human volunteers. Dynamic changes in blood flow in various orofacial regions elicited by painful stimulation of the tooth and finger were measured by means of Laser Doppler imaging (LDI) and computer-assisted infrared thermography (IRT). Blood flow of the finger was recorded by laser Doppler flowmetry (LDF) and plethysmography (PLET). During both stimulus paradigms there was a transient elevation in heart rate (HR) and blood pressure (BP). At the same time there was a significant blood flow decrease in the finger (LDF, PLET) and in the nose (LDI, IRT). In contrast to tooth stimulation, finger stimulation caused a more marked blood flow reduction in the finger. Only high intensity tooth stimulation, but not finger stimulation, caused a long-lasting vasodilatation both in lower and upper lip. The blood flow changes in the lips were not correlated with changes in systemic blood pressure or heart rate. In the cheek, there were no marked flow changes during either finger or tooth stimulation. These data indicate that painful tooth (regional) stimulation, but not finger (remote) stimulation, can induce a long-lasting vasodilatation in parts of orofacial tissues which cannot be explained by changes in central cardiovascular parameters. This tooth-stimulation-induced blood flow increase supports the hypothesis of a special vasodilator reflex mechanism in the orofacial area. Furthermore, tooth-stimulation-induced vasoconstriction in the nose and dilatation in the lips indicate that separate vasoactive reflex mechanisms may exist for different orofacial regions. PMID- 11275392 TI - The Amsterdam Pain Management Index compared to eight frequently used outcome measures to evaluate the adequacy of pain treatment in cancer patients with chronic pain. AB - There is no 'gold standard' to assess the adequacy of pain treatment in cancer patients. The purpose of the study is to explore the Amsterdam Pain Management Index, a newly designed measure to evaluate the adequacy of cancer pain treatment, and to compare it with eight frequently used outcome measures. The Amsterdam Pain Management Index compares patients' Present Pain Intensity, Average Pain Intensity, and Worst Pain Intensity with a composite score of analgesics used, while correcting for what a patient considers as a tolerable level of pain. The eight frequently used outcome measure consisted of three Pain Intensity Markers, the Pain Relief Scale, the Patient Satisfaction Scale, and three Pain Management Indexes. In a randomized controlled trial, 313 cancer patients with a pain duration of at least 1 month were included and followed-up three times until 2 months postdischarge at home. The experimental group received a Pain Education Program, consisting of tailored pain information and instruction. Results showed that, except for the three Pain Management Indexes, the agreement between the measures was very low to moderate. The test of known groups comparisons and equivalence between groups indicated that the Amsterdam Pain Management Index showed promising results. The Pain Intensity Markers and the Pain Relief Scale were limited in discriminating between groups, while the Patient Satisfaction Scale showed no differences between patient groups. Although it was possible for the Pain Management Indexes to distinguish between patient groups, the differences were not in the expected direction. The ability of the outcome measures to detect changes over time was clearly demonstrated by all outcome measures. Effects of the intervention were only found for the Amsterdam Pain Management Index and patients' Substantial Worst Pain score. Although support was provided for the use of the Amsterdam Pain Management Index, more research is warranted. PMID- 11275393 TI - Differential regulation of P2X(3) mRNA expression by peripheral nerve injury in intact and injured neurons in the rat sensory ganglia. AB - The P2X(3) receptor is a ligand-gated cation channel activated by the binding of extracellular adenosine 5'-triphosphate (ATP), an agent that has been suggested to have a role in the nociceptive pathway after tissue and nerve injury. After peripheral nerve injury, both down regulation and up regulation of the P2X(3) receptor in sensory ganglion neurons have been observed. The purpose of this study was to examine the precise regulation of P2X(3) mRNA expression in primary sensory neurons after nerve injury. We used two nerve injury models in the rat, the transection of the tibial and common peroneal nerves and the transection of the infraorbital nerve, and observed dorsal root ganglion (DRG) and trigeminal ganglion neurons, respectively. P2X(3) mRNA in both neuron populations was detected by in situ hybridization with an oligonucleotide probe that was confirmed by Northern blot analysis. To identify axotomized neurons, we examined the expression of activating transcription factor 3 (ATF3), which is regarded as a neuronal-injury marker, using immunohistochemistry. In the DRG, the mean percentage of P2X(3) mRNA-labeled neurons relative to the total number of neurons increased from 32.7% in the naive rats to 42.7% at 3 days after injury. The mean percentage of P2X(3) mRNA-labeled neurons in ATF3 immunoreactive (ir) neurons was 29.5% at 3 postoperative days, which gradually decreased to 11.2% at 28 days after injury. In the trigeminal ganglion, the mean percentage of P2X(3) mRNA labeled neurons was 36.9% at 3 days after injury, versus 26.0% in the naive rats. In the ATF3-ir neurons, the mean percentage of P2X(3) mRNA-labeled neurons was 25.3% at 1 postoperative day and was reduced to 6.1% at 28 postoperative days. The finding that P2X(3) mRNA in ATF3-ir neurons decreased significantly after injury indicates that axotomized neurons decreased the expression of P2X(3) mRNA, despite the increase in P2X(3) mRNA relative to the total number of sensory ganglion neurons. These data strongly suggest that P2X(3) mRNA expression increases in intact neurons and that P2X(3) mRNA in intact neurons may play a role in the pathomechanism of post-nerve injury in primary sensory neurons. PMID- 11275394 TI - St. John's wort has no effect on pain in polyneuropathy. AB - Tricyclic antidepressants are the mainstay of treatment of painful polyneuropathy but cannot be used in a substantial number of patients. St. John's wort is a herbal antidepressant, which may act via mechanisms similar to the tricyclics. The aim of this study was to test if St. John's wort would relieve painful polyneuropathy. The study design was randomized, double-blind, placebo-controlled and cross-over. Fifty-four patients were assigned to one of the two treatment sequences. The daily dose of St. John's wort was three tablets each containing 900 microg totalhypericin. During the two treatment periods of 5 weeks duration, patients rated constant pain, lancinating pain paroxysms, touch-evoked pain and pain on pressure by use of 0--10 point numeric rating scales. Forty-seven patients -- 18 diabetics and 29 non-diabetics -- completed the study. There was a trend of lower total pain score (sum of the individual pain scores) on St. John's wort than on placebo (median 14 vs. 15, P=0.05). None of the individual pain ratings were significantly changed by St. John's wort as compared to placebo (P=0.09--0.33). Complete, good or moderate pain relief was experienced by nine patients with St. John's wort and two with placebo (P=0.07). In conclusion, St. John's wort has no significant effect on pain in polyneuropathy. PMID- 11275395 TI - Pivotal role of capsaicin-sensitive primary afferents in development of both heat and mechanical hyperalgesia induced by intraplantar bee venom injection. AB - To investigate the roles of primary afferent fibers in development of the bee venom (BV)-induced persistent spontaneous nociception (PSN) and hyperalgesia (HA), the sciatic nerve or both the sciatic and saphenous nerves of rats were topically treated with capsaicin respectively under pentobarbital anesthesia to destroy the capsaicin-sensitive primary afferent (CSPA) fibers. Effect of the sciatic nerve capsaicin on the formalin-induced PSN was also evaluated. Destruction of the CSPA fibers of the sciatic nerve or both the sciatic and saphenous nerves only produced 34 or 69% inhibition of the mean total number of 1 h BV-induced paw flinches. However, the total number of 1 h formalin-induced paw flinches was inhibited by 90% (85% for phase 1 and 91% for phase 2). In naive rats, destruction of the CSPA fibers of the sciatic nerve caused 237 and 60% increase in paw withdrawal thermal latency (PWTL) to radiant heat in the injection site (paw pad) and at the heel of the treated hind paw compared to the baseline values. However, it was without significant influence upon the PWTL in the non-treated side or the paw withdrawal mechanical threshold (PWMT) to von Frey filament stimuli in both hind paws. In the BV-treated rats, the CSPA fiber destruction of the sciatic nerve completely blocked development of the heat and mechanical HA in the BV injection site. However, the reduction in either PWTL (drop to baseline level) or PWMT (drop by 56% from the baseline level) at the heel of the BV-treated side was not affected by this treatment. However, destruction of the CSPA fibers of both the sciatic and saphenous nerves was able to block development of both heat and mechanical HA in the whole BV-treated hind paw and heat hyperalgesia in the non-injected hind paw. Taken together, we conclude that: (1) the CSPA (C- and A delta-) fibers play a pivotal role in mediation of either the heat or the mechanical hyperalgesia induced by s.c. BV; (2) the CSPA fibers may play a crucial role in mediation of the formalin-induced PSN, but play a partial role in the BV-induced nociceptive process; (3) in addition to the sciatic nerve, the saphenous nerve is also involved in mediation of the BV-induced PSN as well as heat and mechanical hyperalgesia, while it is not likely to be involved in the formalin-induced nociception. PMID- 11275396 TI - A cellular mechanism for the antinociceptive effect of a kappa opioid receptor agonist. AB - This study used concordant behavioral and electrophysiological approaches to examine the actions of the prototypic kappa opioid receptor agonist U69593 in the rostral ventromedial medulla (RVM). In vitro whole-cell voltage clamp recordings indicated that bath application of U69593 produced outward currents in primary cells in the RVM. In secondary cells, which comprised 80% of the population, U69593 produced a concentration-dependent and norbinaltorphimine (norBNI) reversible inhibition of evoked excitatory postsynaptic currents (EPSCs) in the absence of any postsynaptic effect. U69593 also decreased the frequency, but not the amplitude of spontaneous miniature excitatory postsynaptic currents (mEPSCs) in secondary cells. The inhibition of excitatory inputs to secondary cells would be consonant with disinhibition of primary cells and the production of antinociception. Consistent with this expectation, the activation of kappa opioid receptors in the RVM by microinjection of U69593 produced a dose-dependent increase in paw-withdrawal latency that was antagonized by norBNI. Furthermore, microinjection of norBNI in the RVM antagonized the increases in paw-withdrawal latency and hot-plate latency produced by systemically-administered U69593. In contrast, microinjection of norBNI in the RVM did not antagonize the increase in tail-flick latency produced by systemically-administered U69593. Also, microinjection of U69593 in the RVM did not increase tail-flick latency. The highly test-dependent nature of U69593's effects suggests that the mechanisms by which neurons in the RVM modulate thermal nociceptive responses evoked from the tail and hindpaw are not uniform. Collectively, these data suggest that the RVM is a primary site of action for the antinociceptive actions of kappa opioid receptor agonists and that the mechanism most likely involves a presynaptic inhibition of excitatory inputs to secondary cells. Thus, disinhibition of pain inhibitory neurons in the RVM is likely to be a common mechanism by which opioid receptor agonists produce antinociception, whether by the direct inhibition of inhibitory secondary cells, as in the case of mu opioid receptor agonists, or by a reduction in the excitatory drive to these neurons, as in the case of kappa opioid receptor agonists. PMID- 11275397 TI - Is the R3 component of the human blink reflex nociceptive in origin? AB - The R3 component of the blink reflex can reproducibly be evoked by noxious stimulation but can probably also be elicited by innocuous stimuli. This study was conducted to investigate the contribution of nociceptive A delta and C fibers to the generation of the electrically evoked R3 blink reflex. Electrical thresholds for detection, pain and all blink reflex components were determined and the modulatory effects of local anesthesia were investigated. The electrical R3 threshold of 4.6 +/- 0.5 mA (mean +/- SE) corresponded to 2.9 times the detection threshold and to 0.35 times the pain threshold. The R3 threshold was significantly below the pain threshold. Under local anesthesia of the supraorbital skin with a complete loss of warm and cold sensation, a loss of pinprick sensation, but a normal detection of tactile stimuli, the electrical pain threshold increased, all other thresholds remained unchanged. Under local anesthesia none of the reflex components were significantly reduced. Cutaneous A beta fibers and nociceptive A delta fibers, but not unmyelinated C fibers, contribute to the generation of the electrically evoked R3 component. According to the recruitment order in peripheral sensory nerves the electrical threshold of the R3 is mainly determined by activation of A beta fibers. Thus, it can not be assumed that the electrically evoked R3 is an adequate model to investigate nociceptive processing. PMID- 11275398 TI - A cervical anterior spinal artery syndrome after diagnostic blockade of the right C6-nerve root. AB - A 48-year-old man suffered from intractable neck pain irradiating to his right arm. Magnetic resonance imaging (MRI) of the cervical spine was unremarkable. A right-sided diagnostic C6-nerve root blockade was performed. Immediately following this seemingly uneventful procedure he developed a MRI-proven fatal cervical spinal cord infarction. We describe the blood supply of the cervical spinal cord and suggest that this infarction resulted from an impaired perfusion of the major feeding anterior radicular artery of the spinal cord, after local injection of iotrolan, bupivacaine, and triamcinolon-hexacetonide around the C6 nerve root on the right side. PMID- 11275400 TI - Effects of stimulus presentation rate on the activity of primary somatosensory cortex: a functional magnetic resonance imaging study in humans. AB - To investigate the effect of stimulus presentation rate on the activity of primary somatosensory cortex, we performed echo-planar functional magnetic resonance imaging using a 1.5-tesla magnetic resonance system. Eight right-handed normal volunteers underwent functional magnetic resonance imaging during somatosensory stimulation with a 0.2 ms electrical square wave to the left index finger at 1, 4, 8, 16, and 32 Hz at constant intensity. Activated areas were located mainly around 'the hand area' of the right postcentral gyrus. Between 4 and 16 Hz, almost all subjects showed significant activation, but at 1 Hz and 32 Hz, five of eight subjects showed no activation. The average number of activated pixels in this area between 4 and 16 Hz were significantly greater than those at 1 Hz and 32 Hz, and the average percent signal increase had its activation peak at 8 Hz. Our results suggest that the existence of the optimal stimulation rate range may be a common phenomenon to a variety of sensory modalities. The electrical somatosensory stimulation rates from 4 Hz to 16 Hz are advisable to investigate the activation of the primary somatosensory cortex in human subjects. PMID- 11275401 TI - Ultrastructural evidence for presynaptic mu opioid receptor modulation of synaptic plasticity in NMDA-receptor-containing dendrites in the dentate gyrus. AB - Physiological studies have demonstrated that long-term potentiation (LTP) induction in N-methyl-D-aspartate (NMDA) receptor containing dentate granule cells following lateral perforant path stimulation is opioid dependent, involving mu-opioid receptors (MORs) on gamma-aminobutyric acid (GABA)-ergic neurons. To determine the cellular relationships of MORs to postsynaptic NMDA receptor containing dendrites, immunoreactivity (-I) against MOR and the NMDA receptor subunit 1 (NMDAR1) was examined in the outer molecular layer of the dentate gyrus using electron microscopy. MOR-I was predominantly in axons and axon terminals. NMDAR1-I was almost exclusively in spiny dendrites, but was also in a few terminals. Using immunogold particles to localize precisely NMDAR1, one-third of the NMDAR1-I was detected on the dendritic plasmalemma; in dendritic spines plasmalemmal immunogold particles were near synaptic densities. Many MOR-labeled axons and terminals contacted NMDAR1-labeled dendrites. MOR-labeled terminals formed symmetric (inhibitory-type) synapses on NMDAR1-labeled dendritic shafts or nonsynaptically contacted NMDAR1-labeled shafts and spines. MOR-labeled axons often abutted NMDAR1-containing dendritic spines which received asymmetric (excitatory-type) synapses from unlabeled terminals. Occasionally, MOR-labeled terminals and dendrites were apposed to NMDAR1-containing terminals. These results provide anatomical evidence that endogenous enkephalins or exogenous opioid agonists could inhibit GABAergic terminals that modulate granule cell dendrites, thus boosting depolarizing events in granule cells and facilitating the activation of NMDA receptors located on their dendrites. PMID- 11275402 TI - Alteration of spontaneous firing rate of primary myelinated afferents by ATP in adjuvant-induced inflamed rats. AB - The action of adenosine 5'-triphosphate (ATP) on primary myelinated mechanosensitive afferents was investigated in adjuvant-induced inflamed rats. ATP injected intravenously increased the spontaneous firing rate in most of the A fibers from sciatic nerve innervating inflamed hindpaw, whereas it affected only a few units in normal rats. The effects of ATP were blocked or attenuated by competitive P2 receptor antagonist suramin or reactive blue 2 in most of the fibers tested. Degranulation of mast cells with compound 48/80 did not change the response rate of afferents to ATP in inflamed rats while chemical sympathectomy with 6-hydroxydopamine decreased it to some extent, suggesting an involvement of sympathetic efferents in mediating ATP effects. The results support the view that ATP could change the activity of large mechanosensitive afferents via P2 receptors under state of inflammation, which may be related to touch-evoked pain. PMID- 11275403 TI - Neural correlates of sensory gating in the rat: decreased Fos induction in the lateral septum. AB - In the P(50) gating or conditioning-testing paradigm in the rat, two identical click stimuli are presented with an inter-click interval of 500 ms. The reaction towards the second click, as measured with evoked potentials, is reduced in respect to that towards the first click; this phenomenon is called sensory gating. In the present experiments, the inter-click interval was varied systematically and auditory evoked potentials were measured. Sensory gating was found to occur only at intervals between 500 and 1000 ms, but not at longer intervals. Fos immunohistochemistry was then performed using two groups of rats exposed to double clicks: the inter-click interval was 500 ms in the experimental group and 2500 ms in the control group. Fos induction was analyzed in selected brain structures. In the auditory pathways, Fos-immunoreactive neurons were found in both groups of rats in the inferior colliculus and medial geniculate body. Fos immunoreactive cells were also examined in the septum and hippocampus. In the ventral part of the lateral septal nucleus, the labeled neurons were significantly fewer in the experimental animals compared to the control group. Smaller and non-significant quantitative differences of Fos-positive neurons were documented in the medial septum and hippocampal CA1 region. These data point out a selective decrease in the lateral septum of Fos induced by auditory sensory gating, and suggest an involvement of this structure, and possibly of other parts of the septo-hippocampal system, in sensory gating mechanisms. The results might be relevant for theories on sensory gating deficits in schizophrenia. PMID- 11275404 TI - Involvement of NMDA and AMPA/kainate receptors in the effects of endogenous glutamate on extracellular concentrations of dopamine and GABA in the nucleus accumbens of the awake rat. AB - We have investigated the effects of perfusion of the N-methyl-D-aspartate (NMDA) receptor antagonist 3-((R)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP) and the alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA)/kainate receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX) on the endogenous glutamate-evoked changes of extracellular dopamine and alpha aminobutyric acid (GABA) in the nucleus accumbens of the awake rat. Local infusion of the glutamate uptake inhibitor L-trans-pyrrolidine-2,4-dicarboxilic acid in the nucleus accumbens produced an increase in extracellular concentrations of glutamate, dopamine, and GABA. At the dose of 4 mM, the increase of extracellular glutamate, dopamine, and GABA were 3.73 +/- 0.46 microM (n = 8; p < 0.001), 4.70 +/- 0.92 nM (n = 6; p < 0.001) and 0.36 +/- 0.08 microM (n = 8; p < 0.001), respectively. Perfusion of the NMDA-receptor antagonist CPP attenuated the increases of dopamine by 90% (n = 5; p < 0.001), but enhanced the increases of GABA by 70% (n = 7; p < 0.01). Perfusion of the AMPA-receptor antagonist DNQX did not attenuate the increases of GABA. These results suggest a differential mediation of ionotropic glutamatergic receptors in the actions of endogenous glutamate on extracellular concentration of dopamine and GABA. PMID- 11275405 TI - Laterality of the spinocerebellar axons and location of cells projecting to anterior or posterior cerebellum in the chicken spinal cord. AB - In the cervical and lumbosacral enlargements of the chicken, there are seven spinocerebellar nuclei, the Clarke's column, the spinal border cells, the ventral margin of the ventral horn of both enlargements, and the ventral marginal nucleus in the lumbosacral enlargement. In the present study, we investigated the laterality of spinocerebellar tract axons and the distribution of the spinocerebellar tract neurons projecting into the anterior or posterior part of the cerebellum in these seven nuclei by retrograde transport of wheat germ agglutinin-horseradish peroxidase. The spinocerebellar tract neurons with uncrossed axons were found in the cervical Clarke's column and the cervical spinal border cells, and with crossed ones in the lumbar Clarke's column, lumbar spinal border cells, lumbar lamina IX included in the ventral margin of the ventral horn of the lumbosacral enlargement, and the ventral marginal nucleus. The ventral margin of the ventral horn of the cervical enlargement and lumbar lamina VIII included in the ventral margin of the ventral horn of the lumbosacral enlargement issued spinocerebellar tract axons bilaterally. The spinocerebellar tract neurons of the lumbar spinal border cells and lumbar lamina IX projected to the anterior part of the cerebellum only. And those of the other nuclei projected to both the anterior and posterior parts. PMID- 11275406 TI - Lamina I-periaqueductal gray (PAG) projections represent only a limited part of the total spinal and caudal medullary input to the PAG in the cat. AB - The periaqueductal gray is well known for its involvement in nociception control, but it also plays an important role in the emotional motor system. To accomplish these functions the periaqueductal gray receives input from the limbic system and from the caudal brainstem and spinal cord. Earlier studies gave the impression that the majority of the periaqueductal gray projecting cells in caudal brainstem and spinal cord are located in the contralateral lamina I, which is involved in nociception. The present study in the cat, however, demonstrates that of all periaqueductal gray projecting neurons in the contralateral caudal medulla less than 7% was located in lamina I. Of the spinal periaqueductal gray projecting neurons less than 29% was located in lamina I. However, within the spinal cord large segmental differences exist: in few segments of the enlargements the lamina I-periaqueductal gray projecting neurons represent a majority. In conclusion, although the lamina I-periaqueductal gray projection is a very important nociceptive pathway, it constitutes only a limited part of the total projection from the caudal medulla and spinal cord to the periaqueductal gray. These results suggest that a large portion of the medullo- and spino-periaqueductal gray pathways conveys information other than nociception. PMID- 11275407 TI - The efferent connections to the thalamus and brainstem of the physiologically defined eye field in the rat medial frontal cortex. AB - The anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) was injected into sites of the rat frontal eye field (FEF) located in the medial frontal cortex. After a single iontophoretic injection of PHA-L into a FEF site where intracortical microstimulation elicited eye movements, anterogradely labelled fibres and terminal-like elements were found in the thalamus in the anterior nuclei, intralaminar nuclei, lateral portion of the mediodorsal nucleus and posterior nuclear group. In the midbrain and pons, labelled fibres were located in the anterior pretectal area, Darkschewitsch nucleus, superior colliculus and dorsolateral portion of the central gray. When the tracer was injected at the FEF periphery, at a site the stimulation of which evoked both eye and whisker movements, labelling distribution in the thalamus differed from that observed after FEF injections, while a similar distribution was observed in the brainstem. In the thalamus, anterograde labelling was observed in these latter cases in the anterior nuclei, ventral nuclei, medial portion of the laterodorsal nucleus. The present findings point out that the FEF and FEF periphery are connected with numerous subcortical structures of the thalamus and brainstem. In addition, the connections of FEF and FEF periphery with the thalamus differ, whereas the midbrain and pons connections of the two subdivisions share common targets. PMID- 11275408 TI - Expression of the E2F and retinoblastoma families of proteins during neural differentiation. AB - Development of the brain is determined by a strictly orchestrated program of proliferation, migration, apoptosis, differentiation, synaptogenesis, tract formation, and myelination. The E2F family of transcription factors, whose activity and functions are regulated in large part through interactions with the retinoblastoma (Rb) family of tumor suppressor proteins, has been implicated as a key regulator of proliferation, differentiation, and apoptosis in a variety of tissues. We have examined levels of the E2F and Rb families of proteins during both brain development and neural differentiation of P19 cells, and found the expression profiles during these two processes of neural development and maturation to be quite similar, i.e., strong up-regulation of p130, pronounced down-regulation of p107, moderate up-regulation of pRb, and significant down regulation of most species of E2F and dimerization protein (DP). However, several specific isoforms, namely a 30 kDa form of DP-2, a 57 kDa species of E2F-3, a 59 kDa form of E2F-5 and the isoforms of E2F-1 recognized by the E2F-1 (KH-95) antibody were up-regulated suggesting that these particular isoforms of E2F and DP play a tissue-specific function in differentiation and maturation of nervous tissue. The potential role of the E2F/DP family of transcription factors in aspects of neural development and differentiation are considered. PMID- 11275409 TI - Increased nitric oxide synthase activity in a model of serotonin depletion. AB - Serotonin (5HT) containing cell bodies are localized in mesencephalic and rhombencephalic raphe nuclei. It has been proposed that 5HT could be involved in neuronal development and plasticity. In the central nervous system, nitric oxide (NO) has been postulated as a neurotransmitter and neuromodulator, and has been implicated in neurotoxicity as well as in neuroprotection. Using the nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) technique, NO synthesizing neurons were described in raphe nuclei. By immunohistochemistry, nitric oxide synthase (NOS) was found colocalized with 5HT in some dorsal raphe nucleus (DRN) neurons. In a model of inhibition of 5HT synthesis produced by daily administration of parachlorophenilalanine during 14 days, we have studied the relationship between 5HT and NO systems after 5HT depletion by histochemical and immunocytochemical methods. After the treatment, we observed an important reduction of 5HT immunostaining in the DRN and enhanced NOS activity demonstrated by NADPH-d technique, especially in the dorsomedial and ventromedial subgroups. In spite of the increased NOS activity, we could not observe significant changes in the NOS-immunoreactivity in the DRN after 5HT depletion. These results could indicate that 5HT depletion is concomitant with changes in NOS activity without affecting NOS expression in the DRN. PMID- 11275410 TI - Involvement of nitric oxide in morphine-induced c-Fos expression in the rat striatum. AB - Induction of expression of immediate-early gene c-Fos in the striatum is a common effect of many drugs of abuse, including morphine. Previous studies have shown that the morphine-mediated c-Fos response is attenuated by antagonists of the N methyl-D-aspartate (NMDA) subtype of glutamate receptor. Other evidence suggests that the NDMA receptor may be coupled to the enzyme neuronal nitric oxide synthase (nNOS). NMDA receptor-mediated increases in intracellular calcium can activate nNOS, which catalyzes the formation of the signaling molecule nitric oxide. Because activation of NMDA receptors mediates morphine-induced c-Fos expression, we tested the hypothesis that activation of nNOS is involved in this cascade. Male rats were injected with the nNOS-selective inhibitor 7 nitroindazole (7-NI) or vehicle 30 min prior to injection of morphine sulfate or vehicle. Two hours later they were perfused with fixative and the brains removed for immunocytochemical analysis for c-Fos. Morphine induced c-Fos expression in the striatum, cerebral cortex, and midline/intralaminar nuclei of thalamus. Expression in the striatum, but not thalamus or cortex, was significantly blocked by 7-NI. Double-label immunocytochemistry revealed no co-localization of c-Fos and nNOS in any brain region. These results support a role for nNOS in the neural circuits activated by morphine. PMID- 11275411 TI - Effects of a kappa agonist, spiradoline mesylate (U62,066E), on activation and vaginocervical-stimulation produced analgesia in rats. AB - Previous research has demonstrated increased pain threshold during copulation, gestation, and parturition in animals. In the laboratory, mechanostimulation of the vaginocervical region in many animals, as well as humans, can increase responsiveness to noxious but not to innocuous stimuli. This increased pain inhibition to vaginocervical stimulation, which mimics natural parturition, is mediated by spinal and supraspinal neuropeptides, including the opiates. The present research was designed to ascertain the possible effects of a kappa opioid agonist on vaginocervical-stimulated analgesia in rats. Initially, the novel kappa-selective agonist, spiradoline mesylate (U62,066E; 0, 0.1, 1.0, 10.0 mg/kg, i.p.), was injected intraperitoneally and general behavioral arousal in an open field apparatus was recorded. Results from this experiment indicate that stimulation with the kappa-selective drug caused significant decreases in behavioral activity at the high dose as compared to saline and the medium and low doses. Next, the effects of U62,066E (0, 0.1, 1.0, 10.0 mg/kg, i.p.) on the analgesia associated with vaginocervical stimulation were determined in a tail flick apparatus. The kappa drug significantly increased antinociceptive thresholds prior to and during vaginocervical stimulation at the 0.1 and 1.0 mg/kg doses. By contrast, the high dose (10.0 mg/kg) of U62,066E decreased vaginocervical stimulation-produced analgesia. Results are discussed in terms of the potential of nonaddictive kappa-selective opioid compounds being utilized in reproductive pain. PMID- 11275412 TI - An overview of the central nervous system of the elephant through a critical appraisal of the literature published in the XIX and XX centuries. AB - The two species of elephants (Indian: Elephas maximus and African: Loxodonta africana) possess the largest brain among land mammals. Due to its size, the elephant brain is discussed in virtually every paper dealing with the evolution of the central nervous system of mammals and comparative brain size. Studies on the social habits of elephants also deal with the skills and the "intelligence" and brain size of these species. Yet most of the descriptions and conclusions reported in comparative studies rely on second-hand data derived from investigations performed several decades before, often dating as far back as the XIX century. Furthermore, many of the original papers actually describing gross and detailed features of the brain of elephants are either no longer available, are written in languages other than English, or are difficult to trace. The present study gives a short description of the anatomy of the central nervous system of elephants, with special attention to its distinctive features, reports all available literature on the subject, and briefly discusses its origins and rationale. PMID- 11275413 TI - Agonists determine the pattern of G-protein activation in mu-opioid receptor mediated supraspinal analgesia. AB - The opioids heroin, methadone, buprenorphine, and morphine produce supraspinal antinociception in CD-1 mice that is antagonized by Cys(2), Tyr(3), Orn(5), Pen(7)-amide but not by naltrindole or nor-binaltorphimine. The patterns of GTP binding regulatory proteins (G-proteins) activation exhibited by these agonists at mu-opioid receptors were characterized. The expression of alpha-subunits of Gi protein classes, Gi1, Gi2, Gi3, Go1, Go2 and Gz, and those of the Gq-protein family, Gq and G11, was reduced by administration of antisense oligodeoxynucleotides (ODNs) complementary to sequences in their respective mRNAs. The ODN treatments demonstrated differences in the analgesic profiles of these opioids. Though the knock-down of G(i2)alpha or G(z)alpha subunits diminished the analgesic effects of the four opioids, impairment of G(i3)alpha did not modify the potency of morphine. In mice with reduced G(i1)alpha, G(o1)alpha or G(11)alpha levels, antinociception induced by heroin and methadone was diminished, but buprenorphine and morphine showed no change in their effects. Also, antinociception induced by heroin and buprenorphine, but neither morphine nor methadone, required intact G(o2)alpha or G(q)alpha levels. Thus, morphine, heroin, methadone, and buprenorphine showed different patterns of G-protein activation in evoking mu-opioid receptor-mediated supraspinal antinociception. Therefore, after binding identical receptors, each agonist determines the classes of GTP-binding regulatory transducer proteins to be activated. PMID- 11275414 TI - A grasp-related deficit in tactile discrimination following dorsal column lesion in the rat. AB - The dorsal columns of the spinal cord are a major source of haptic (sense of active touch) and proprioceptive input to the brainstem and sensory-motor cortex. Following injury in primates, there are impairments in two-point discrimination, direction of movement across the skin, and frequency of vibration, and qualitative control of the digits, but simple spatial discriminations recover. In the rat there are qualitative deficits in paw control in skilled reaching, but no sensory deficits have been reported. Because recent investigations of sensory control suggest that sensory functions may be related to specific actions, the present study investigated whether the dorsal columns contribute to hapsis during food grasping in the rat. Adult female Long-Evans rats were trained to reach with a single forepaw for a piece of uncooked pasta or for equivalent sized but tactually different nonfood items. One group was given lesions of the dorsal column ipsilateral to their preferred paw, while the second group served as a control. Postlesion, both groups were tested for skilled reaching success and force application as well as adhesive dot removal and forepaw placing. Performance levels on these tests were normal. Nevertheless, the rats with dorsal column lesions were unable to discriminate a food item from a tactually distinctive nonfood item as part of the reaching act, suggesting that the dorsal columns are important for on-line tactile discriminations, or "haptic actions," which contribute to the normal performance of grasping actions. PMID- 11275415 TI - The staircase test of skilled reaching in mice. AB - The "staircase" test has become established for measurement of side-specific deficits in coordinated paw reaching in rats, and has been shown to reveal impairments on the contralateral side following unilateral lesions in a wide range of motor structures of the brain. As mice become more widely used in behavioural neuroscience, we have scaled down the staircase reaching test for application to this latter species. We here validate the test in C57BL/6J mice by (a) establishing the optimal dimensions of the apparatus, (b) comparing the effects of test parameters including sex, test duration, levels of deprivation and alternative reward pellets, and (c) demonstrating contralateral deficits after aspirative lesions of the motor cortex. Differences between mice and rats in normal performance of the task are noted. The staircase test provides a simple objective test of skilled motor function that allows measurement of lateralised effects without unduly constraining the animal, and which may prove as useful for mice as has previously been demonstrated in rats. PMID- 11275416 TI - Effects of an aqueous extract of processed bidi tobacco on the growth of hamster tracheal epithelial cells. AB - Inhalation of tobacco dust is responsible for elevated genotoxicity and pulmonary ailments in workers engaged in processing tobacco for the manufacture of bidis, the Indian version of cigarettes. Tracheal tissue being the major site of interaction with tobacco dust, the effects of different concentrations of an aqueous extract of bidi tobacco (ATE) on the growth of a hamster tracheal epithelial cell line (HTE) were investigated. Colony forming efficiency assay revealed that ATE was cytotoxic only at the highest concentration of 5.0 mg/ml. In cultures treated with 1.25 mg/ml ATE, the cell doubling time and growth rate were similar to that of the controls, while a significant increase in cell doubling time (29.4+/-0.3 h vs 14.0+/-3.75 h, P<0.001) was observed at 2.5 mg/ml ATE concentration. Exposure of HTE cells to the non-toxic ATE concentration of 2.5 mg/ml was found to stimulate ornithine decarboxylase (ODC) activity, incorporation of [3H] methyl thymidine into DNA and increase in the S phase fraction was seen by flow cytometry. However, a 56% reduction in the growth rate of cultures treated with 2.5 mg/ml ATE was related to the prolongation of the traverse of cells through S phase. ATE-induced growth suppression was reversed when cultures were grown in ATE-free medium or upon repeated exposure to ATE. The findings suggest that increased tracheal cell proliferation induced by chronic inhalation of tobacco dust may contribute to the development of pulmonary disorders and possibly neoplasia in exposed individuals. PMID- 11275417 TI - Significant differences in the cellular and molecular reactions of rat and hamster lung after quartz exposure. AB - Exposure of rats to high doses of quartz and other insoluble isometric particles can produce lung tumors. In contrast, after exposure of such particles in hamsters no tumor outcome has been observed. Recent studies have demonstrated that the tumorigenic effect of particles is closely linked to the induction of inflammatory processes and the subsequent formation and persistence of mutagenic oxidative DNA-modifications. Species-specific differences in sensitivity to particles should therefore be reflected in the molecular reaction of the lung cells. We exposed rats and hamsters to two different doses of quartz (0.3 mg, 1.2 mg/100 g body weight) by intratracheal instillation and characterized the dose related pattern of pulmonary inflammation (neutrophil recruitment, TNF), toxicity (protein content, surfactant phospholipids), antioxidant defence (glutathione content), mutagenicity (8-oxoguanine, p53) and proliferation. Our results clearly demonstrate a significantly higher response of the rat to quartz exposure for all determined molecular and cellular parameters. Therefore the examination of these parameters in humans would contribute to the evaluation of the relevance of rats or hamsters as models to predict particle-induced human lung cancer risk. PMID- 11275418 TI - Diethylphosphorylation of rat cardiac M2 muscarinic receptor by chlorpyrifos oxon in vitro. AB - The acute toxicity of chlorpyrifos oxon (CPO), the metabolically-activated form of the major organophosphorus insecticide chlorpyrifos, is attributable to diethylphosphorylation of acetylcholinesterase at its esteratic site. As a secondary effect, CPO is known to compete with agonist binding to the M2 muscarinic acetylcholine receptor (mAChR). This study tests the hypothesis that [ethyl-1,2-(3)H]CPO labels the M2 mAChR in rat cardiac membrane proteins. Of four labeled protein regions observed, only one had an apparent molecular mass (70-75 kDa) consistent with that of glycosylated M2 mAChR. It was identified as M2 muscarinic receptor by Western blotting and immunoprecipitation using a cardiac specific M2 mAChR monoclonal antibody, providing the first direct evidence for diethylphosphorylation of a muscarinic receptor. This may be a functionally important M2 mAChR site, but the toxicological relevance and species and organ specificity of diethylphosphorylation are unknown. PMID- 11275419 TI - Selenoxides inhibit delta-aminolevulinic acid dehydratase. AB - The effect of two selenides and their selenoxides on delta-aminolevulinic acid dehydratase (delta-ALA-D) from liver of adult rats was investigated. In vivo, selenides can be oxidized to selenoxides by flavin-containing monooxygenases (FMO) and selenoxides can regenerate selenides by thiol oxidation. Phenyl methyl selenide (PhSeCH3) and 1-hexynyl methyl selenide (C4H9Ctriple bondCSeCH3) were converted to selenoxides by reaction with H2O2. PhSeCH3 and C4H9Ctriple bondCSeCH3 had no effect on delta-ALA-D up to 400 microM. Conversely, their selenoxides inhibited delta-ALA-D, and the IC(50) for enzyme inhibition was about 100 and 70 microM, respectively. Partially purified delta-ALA-D (P(55)) from swine liver was also inhibited by these selenoxides. The inhibitory action of selenoxides was antagonized by dithiotreitol (DTT). Moreover, delta-ALA-D from a plant source was inhibited by the selenoxides, suggesting a possible involvement of SH groups in a distinct site of the homologous region implicated in Zn2+ binding in mammalian delta-ALA-D. After exposure to PhSeCH3 (500 micromol/kg/day) for 45 or 30 days, the activity of delta-ALA-D from liver of mice decreased to about 50% of the control group. The in vivo inhibitory action of this compound was not antagonized by DTT. PhSeCH3 and C4H9Ctriple bondCSeCH3 had no effect on the rate of DTT oxidation, but their selenoxides oxidized DTT. The results of the present study suggest that hepatic delta-ALA-D of rodents is a potential molecular target for selenides as a consequence of their metabolism to selenoxides by FMO. PMID- 11275420 TI - Interaction of zinc on biomarker responses in rats exposed to 2,5-hexanedione by two routes of exposure. AB - The interaction of zinc(II) on the toxicokinetics of 2,5-hexanedione (2,5-HD), the ultimate toxic metabolite of n-hexane, was performed by quantifying the changes of two urinary biomarkers, free 2,5-HD and pyrrole derivatives, in rats exposed to 2,5-HD and to 2,5-HD plus zinc acetate. Eight groups of Wistar rats were exposed for 4 days (dietary and intraperitoneally) to 2,5-HD, zinc acetate and 2,5-HD plus zinc acetate and the 24 h urine was used to determine the excretion of these biomarkers. On comparing the results obtained by the two routes of exposure with different doses of 2,5-HD and zinc acetate, it was observed that there was a significant decrease (P<0.05) in the excretion of free 2,5-HD and pyrroles derivatives in rats exposed to the chemical mixture, when compared with the excretion of these biomarkers in rats exposed to 2,5-HD alone. To evaluate the mechanism of this interaction, further experiments were performed using one group of rat dietary pre-exposed to zinc acetate followed by 2,5-HD exposure. The results of our experiment suggest that zinc protect proteins of pyrrolization by coordination to amino groups, with the subsequent inhibition of protein cross-linking responsible by 2,5-HD neurotoxicity. PMID- 11275421 TI - Induction of human NAD(P)H:quinone oxidoreductase (NQO1) gene expression by the flavonol quercetin. AB - Flavonoids are plant polyphenolic compounds ubiquitous in fruits, vegetables and herbs. The flavonol quercetin is one of the most abundant dietary flavonoids. It has diverse biological properties in cultured cells, including cytoprotection, and exhibits antitumorigenic effects in animal models. The mechanism(s) for the protective properties of flavonoids are currently unknown but may involve modulation of phase II detoxifying enzymes. We have investigated the effect of quercetin on expression and enzymatic activity of one of the major phase II detoxification systems, NAD(P)H:quinone oxidoreductase (NQO1) in the MCF-7 human breast carcinoma cell line. We show that treatment of MCF-7 cells for 24 h with 15 microM quercetin results in a twofold increase in NQO1 protein levels and enzyme activity, and a three- to fourfold increase in NQO1 mRNA expression. We found that when these cells were transiently transfected with a luciferase (Luc) reporter plasmid containing two copies of the antioxidant response element (ARE) of the human NQO1 gene linked to a minimal viral promoter, quercetin caused an approximately twofold increase in Luc activity. Quercetin failed to increase Luc activity in cells transfected with a reporter vector containing a mutated ARE. The increase in NQO1 transcription in response to quercetin suggests that dietary plant polyphenols can stimulate transcription of phase II detoxifying systems, potentially through an ARE-dependent mechanism. Induction of the human NQO1 gene by dietary polyphenolics could afford protection against carcinogenic chemicals in molecular pathways utilizing the ARE. PMID- 11275423 TI - 5-Formyluracil and its nucleoside derivatives confer toxicity and mutagenicity to mammalian cells by interfering with normal RNA and DNA metabolism. AB - Oxidation of the methyl group of thymine yields 5-(hydroxymethyl)uracil (5-hmU) and 5-formyluracil (5-foU) as major products. Whereas 5-hmU appears to have normal base pairing properties, the biological effects of 5-foU are rather poorly characterised. Here, we show that the colony forming ability of Chinese hamster fibroblast (CHF) cells is greatly reduced by addition of 5-foU, 5-formyluridine (5-foUrd) and 5-formyl-2'-deoxyuridine (5-fodUrd) to the growth medium. There are no toxic effects of 5-fodUrd on cells defective in thymidine kinase or thymidylate synthetase, suggesting that the toxicity may be caused by 5-fodUrd phosphorylation and subsequent inhibition of thymidylate synthetase. Whereas 5 fodUrd was the most effective 5-foU derivative causing cell growth inhibition, the corresponding ribonucleoside 5-foUrd was more effective in inhibiting [3H]uridine incorporation in non-dividing rat nerve cells in culture, suggesting that 5-foUrd exerts its toxicity through interference with RNA rather than DNA synthesis. Addition of 5-foU and 5-fodUrd was also found to promote mutagenicity at the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus of CHF cells; 5-fodUrd being three orders of magnitude more potent than 5-foU. In contrast, neither 5-hmU nor 5-(hydroxymethyl)-2'-deoxyuridine induced HPRT mutations. The mutation induction indicates that 5-foU will be incorporated into DNA and has base pairing properties different from that of thymine. These results suggest that 5-foU residues, originating from incorporation of oxidised bases, nucleosides or nucleotides or by oxidation of DNA, may contribute significantly to the damaging effects of oxygen radical species in mammalian cells. PMID- 11275422 TI - Pharmacological interventions of cyanide-induced cytotoxicity and DNA damage in isolated rat thymocytes and their protective efficacy in vivo. AB - Cyanide inhibits the mitochondrial respiratory chain enzyme cytochrome oxidase causing histotoxic hypoxia. It is primarily considered as a neurotoxin but its other toxic manifestations are also well documented. Cyanide-induced apoptosis in neuronal cells has also been demonstrated recently. At the same time we also reported that potassium cyanide (KCN) produces extensive cytotoxicity and DNA fragmentation in rat thymocytes. The DNA damage was sensitive to elevated levels of extracellular Ca2+ and was attenuated by Zn2+ (modulator of Ca2+ dependent endonuclease), N-acetylcysteine (free radical scavenger) and diltiazem (Ca2+ channel blocker). In a continuation of this work, in the present study we have shown that the cytotoxicity and DNA fragmentation induced by 5 mM KCN was preceded by loss of mitochondrial integrity (MTT assay and rhodamine-123 staining) and nuclear viability (propidium iodide uptake) which were mediated by generation of reactive oxygen species (DCHF-DA staining). The DNA damage was also accompanied by nuclear fragmentation (Hoechst 33342 staining), a phenomenon that characterises the 'apoptotic' type of cell death. The in vitro toxic insult of KCN was challenged by pre-treatment (0.5 h), simultaneous treatment or post treatment (0.5-3 h) of various pharmacological agents viz., Trolox (antioxidant), EGTA (Ca2+ modulator) and aurintricarboxylic acid (ATA; Ca2+/Mg2+ dependent endonuclease inhibitor). In addition, Quercetin (antioxidant) was tested as simultaneous treatment alone and was found to be ineffective. On the basis of various biochemical indices and DNA fragmentation (quantitative and qualitative), simultaneous treatment of Trolox was found to be the most effective in attenuating cyanide toxicity in vitro. This protection can be attributed to interventions in oxidative stress-mediated cell injury which is an early event preceding DNA damage. Both EGTA and ATA could not prevent this damage. Trolox also increased the LD(50) of KCN in mice 2.5-fold as compared to 1.8- and 1.6 fold for EGTA and ATA, respectively. PMID- 11275425 TI - Application of LC and HPTLC-densitometry for the simultaneous determination of benazepril hydrochloride and hydrochlorothiazide. AB - Two methods are described for the simultaneous determination of benazepril HCl and hydrochlorothiazide in binary mixture. The first method was based on HPTLC separation of the two drugs followed by densitometric measurements of their spots at 238 and 275 nm for benazepril HCl and hydrochlorothiazide, respectively. The separation was carried out on Merck HPTLC aluminum sheets of silica gel 60 F(254,) using ethyl acetate-methanol-chloroform (10:3:2 v/v) as mobile phase. Second order polynomial equation was used for the regression line in the range 2 20 and 2.5-25 microg/spot for benazepril HCl and hydrochlorothiazide, respectively. The second method was based on HPLC separation of the two drugs on reversed phase, ODS column at ambient temperature using a mobile phase consisting of acetonitrile and water (35:65 v/v) and adjusting to pH 3.3 with acetic acid. Quantitation was achieved with UV detection at 240 nm based on peak area with linear calibration curves at concentration ranges 10-60 and 12.5-75 microg ml(-1) for benazepril HCl and hydrochlorothiazide, respectively. The two proposed methods were successfully applied to the determination of both drugs in laboratory prepared mixtures and in commercial tablets. No chromatographic interference from the tablets excipients was found. PMID- 11275424 TI - Effects of 4-tert-octylphenol on initiation and maintenance of pregnancy following oral administration during early pregnancy in rats. AB - 4-tert-Octylphenol (OP) is an alkylphenol that is an intermediate in the production of alkylphenol ethoxylates. OP has been reported to be the most potent estrogenic alkylphenol in vitro. In the present study, the effects of OP on initiation and maintenance of pregnancy were investigated in rats. Inseminated female rats were orally given OP at 0,15.6,31.3,62.5 and 125 mg/kg on day 0 through day 8 of pregnancy. Female rats were sacrificed on day 20 of pregnancy, and pregnancy outcome was determined. Decreases in body weight gain and food consumption on days 0-9 were found at 31.3 mg/kg and above, and at 15.6 mg/kg and above, respectively. The pregnancy rate was not adversely affected by OP administration during early pregnancy even at 125 mg/kg. The incidence of post implantation loss per litter at 31.3 mg/kg and above was significantly higher than that in the control group. The body weights of live fetuses in the OP treated groups were not significantly different from those in the control group. No increase in the incidence of fetuses with external malformations was found in any OP-treated group. We concluded that OP during early pregnancy caused post implantation embryonic loss at doses that showed maternal toxicity. PMID- 11275426 TI - Chromatographic separation and spectrometric identification of the oxidation products from a tetrahydro-isoquinoline alkaloid. AB - The oxidation chemistry of 3',4'-deoxynorlaudanosoline carboxylic acid, a tetrahydroisoquinoline alkaloid, has been studied by electrochemical approaches. Four reaction products were isolated by semi-preparative high performance liquid chromatography and identified structurally by nuclear magnetic resonance, mass spectrometry, ultraviolet-visible spectrophotometry and electrochemistry studies. An oxidation mechanism was proposed. PMID- 11275427 TI - Trimetazidine: stability indicating RPLC assay method. AB - An accurate, specific and reproducible reversed phase liquid chromatographic method for the determination of trimetazidine hydrochloride in presence of its degradation products is reported. The mobile phase consisted of water acetonitrile-triethylamine (90:10:0.1, v/v/v) adjusted with o-phosphoric acid to a pH of 3.3. Chromatography was performed using C-18 column at a flow rate of 1.0 ml/min and the drug along with its degraded products was detected at 270 nm. The calibration curve of trimetazidine hydrochloride in methanol was linear in the range 500-3000 ng. The mean value of correlation coefficient, slope and intercept were 0.99859 &# 0.001, 17.7986 &# 0.0709 and 482.56 &# 147.03, respectively. The limits of detection and quantitation were 5 and 20 ng, respectively. The recovery of trimetazidine hydrochloride was about 99-100%. This method was utilized to analyze trimetazidine hydrochloride from conventional tablets and controlled release pellets in the presence of commonly used excipients. PMID- 11275428 TI - An HPLC assay and a microbiological assay to determine levofloxacin in soft tissue, bone, bile and serum. AB - A simple, specific and sensitive HPLC assay for levofloxacin in serum, bile, soft tissue and bone was evaluated and validated. The samples were prepared by protein precipitation with acids and methanol, which yielded high recoveries (for serum and bile>98% and for bone and soft tissue>90%). The compounds were separated on a reversed phase column with an acidic mobile phase containing triethylamine. The eluate was monitored by fluorescence detection. The HPLC assay is linear over the usable concentration range (0.1-40 microg/ml) and it provides good validation data for accuracy and precision. Although comparison of HPLC results to the results of a microbiological assay showed congruent results (regression coefficients>0.967). HPLC should be the method of choice for determination of levofloxacin in biological matrices. PMID- 11275429 TI - Determination of Norfloxacin by square-wave adsorptive voltammetry on a glassy carbon electrode. AB - The adsorptive and electrochemical behavior of norfloxacin on a glassy carbon electrode were investigated by cyclic and square-wave voltammetry. Cyclic voltammetric studies indicated that the process was irreversible and fundamentally controlled by adsorption. To obtain a good sensitivity, the solution conditions and instrumental parameters were studied using square-wave voltammetry. In acetate buffer of pH 5.0, norfloxacin gave a sensitive adsorptive oxidative peak at 0.920 V (versus Ag-AgCl). Applicability to measurement of norfloxacin at submicromolar levels in urine samples was illustrated. The peak current was linear with the norfloxacin concentration in the range 5-50 microg ml(-1) urine. The detection limit was 1.1 microg ml(-1) urine. PMID- 11275430 TI - Synthesis and LC characterization of clenbuterol molecularly imprinted polymers. AB - Highly selective non-covalent clenbuterol (CL) imprinted polymers were prepared using methacrylic acid as monomer and ethylene glycol dimethacrylate as cross linking agent. HPLC experiments with columns packed with this material showed that CL are selectively recognised with respect to all other adrenergic substances studied using a phosphate buffer/acetonitrile eluent. The separation was strongly dependent on pH and the organic/aqueous phase ratio. An important contribution to the recognition mechanism from hydrophobic interactions was found at higher water content. These results demonstrate that a novel family of absorbents with high selectivity for CL was obtained which can be exploited in solid phase extractions or as recognition elements for selective sensors. PMID- 11275431 TI - Development and validation of a sensitive and robust LC-tandem MS method for the analysis of warfarin enantiomers in human plasma. AB - A liquid chromatography-tandem mass spectrometry method (LC-MS-MS) was developed and validated for measuring warfarin (WAR) enantiomers (R-WAR and S-WAR) in human EDTA plasma. Liquid-liquid extraction using ethyl ether was used to extract the analytes from the plasma. Baseline resolution of S- and R-WAR as well as the internal standard enantiomers (S- and R-p-ClWAR, S-IS and R-IS) was achieved on a beta-cyclodextrin column with a mobile phase of acetonitrile-acetic acid triethylamine (1000:3:2.5, v/v/v). The retention times are 6.9, 8.0, 7.0, and 7.9 min for S-WAR, R-WAR, S-IS and R-IS, respectively. The detection was by monitoring S- and R-WAR at m/z 307-->161 and S- and R-IS at m/z 341-->161 using ( ) ESI. The standard curve range was 1-100 ng ml(-1) for both S- and R-WAR. The inter-day precision and accuracy of the quality control (QC) samples were <7.3% relative standard deviation (RSD) and <7.3% bias for S-WAR, and <6.5% RSD and <5.8% bias for R-WAR, respectively. Analyte stability during sample processing and storage were established. Method ruggedness was demonstrated by the reproducible performance from analysis of clinical samples. PMID- 11275432 TI - Theoretically-derived molecular descriptors important in human intestinal absorption. AB - A quantitative structure-human intestinal absorption relationship was developed using artificial neural network (ANN) modeling. A set of 86 drug compounds and their experimentally-derived intestinal absorption values used in this study was gathered from the literature and a total of 57 global molecular descriptors, including constitutional, topological, chemical, geometrical and quantum chemical descriptors, calculated for each compound. A supervised network with radial basis transfer function was used to correlate calculated molecular descriptors with experimentally-derived measures of human intestinal absorption. A genetic algorithm was then used to select important molecular descriptors. Intestinal absorption values (IA%) were used as the ANN's output and calculated molecular descriptors as the inputs. The best genetic neural network (GNN) model with 15 input descriptors was chosen, and the significance of the selected descriptors for intestinal absorption examined. Results obtained with the model that was developed indicate that lipophilicity, conformational stability and inter molecular interactions (polarity, and hydrogen bonding) have the largest impact on intestinal absorption. PMID- 11275434 TI - Determination of 24(R)-pseudoginsenoside F(11) in North American ginseng using high performance liquid chromatography with evaporative light scattering detection. AB - A gradient liquid chromatographic method with evaporative light scattering detection (ELSD) for the determination of 24(R)-pseudoginsenoside F(11) in North American ginseng is described. Samples are analyzed by means of a reverse-phase column (Waters Spherisorb ODS-2, C(18)) using acetonitrile and water under gradient conditions as the mobile phase over 20 min. The evaporative light scattering detector (ELSD) used, was set at an evaporating temperature of 35 degrees C and nitrogen gas pressure of 3.4 bar. The detection limit (S/N>5) of 24(R)-pseudoginsenoside F(11) is 53 ng on column. PMID- 11275433 TI - Measurement of potency and lipids in monensin fermentation broth by near-infrared spectroscopy. AB - A transmission near-infrared (NIR) spectroscopic method for quantification of potency and lipids in monensin fermentation broth was developed and validated. Two multiple linear regression calibration curves were established for a set of 100 fermentation samples, correlating the appropriate absorption bands in the NIR spectrum to the laboratory reference methods; high-performance liquid chromatography for potency, and chloroform extraction for lipids. During method development, potency was found to be well correlated to NIR absorbances specific for monensin. While acceptable, correlation of NIR absorbances characteristic of oil to the chloroform lipid method was weaker due to a greater amount of relative variation in the lipid measurements. Following establishment of the optimal calibration curves, the NIR method for potency and lipids was validated for selectivity, accuracy, precision, and robustness. In order to investigate long term drift in the measurement system, samples were tested both by the NIR and the reference methods over a 7-month period. The differences between results from the two measurements were calculated and statistically analyzed. PMID- 11275435 TI - Development and validation of a reverse-phase HPLC method for analysis of efavirenz and its related substances in the drug substance and in a capsule formulation. AB - A stability-indicating high performance liquid chromatographic (HPLC) method was developed for the assay of efavirenz, a non-nucleoside reverse transcriptase inhibitor used in the treatment of AIDS. The HPLC method, which is used to determine potency in efavirenz capsules and related substances in efavirenz drug substance and capsules, was validated per ICH guidelines. This method, which uses a cyano column, is capable of separating efavirenz from its trans-alkene reduction product. This paper will discuss development and validation of this method, which, to the best of our knowledge, is the first known separation of homologs containing double and triple bonds using reverse-phase HPLC. PMID- 11275436 TI - Evaluation of a fluorogenic derivatization method for the reversed-phase HPLC analysis of 2'-beta-fluoro-2',3'-dideoxyadenosine, a new anti-AIDS drug. AB - High sensitivity (10(-7) to 10(-9) M) reversed-phase high-performance liquid chromatography (HPLC) analysis of adenine nucleosides and nucleotides, especially in a biological matrix, is difficult using only ultraviolet detection. Derivatization coupled with fluorescence detection has been investigated as a means of enhancing sensitivity for the reversed-phase HPLC analysis of 2'-beta fluoro-2',3'-dideoxyadenosine (F-ddA), an experimental, acid-stable, anti-AIDS drug. The reaction of chloroacetaldehyde with the adenine base has been employed to form fluorescent 1,N(6)-etheno derivatives of F-ddA and 5'-deoxyadenosine, which is used as an internal standard. These derivatives give an analytically useful fluorescence emission at 416 nm after excitation at 230, 265, or 275 nm. Derivatization, fluorescence detection and reversed-phase chromatography have been optimized for the analysis of nanomolar concentrations of F-ddA in human plasma. This method has potential for the measurement of F-ddA at low concentration and in limited volume samples from in vivo biological studies. PMID- 11275437 TI - Spectrophotometric determination of benazepril hydrochloride and hydrochlorothiazide in binary mixture using second derivative, second derivative of the ratio spectra and chemometric methods. AB - Different spectrophotometric methods are presented for the simultaneous determination of benazepril hydrochloride and hydrochlorothiazide in pharmaceutical tablets. The first method depends on second derivative (2D) ultraviolet spectrophotometry, with zero crossing and peak to base measurement. The second derivative amplitudes at 214.8 and 227.4 nm were selected for the assay of benazepril hydrochloride and hydrochlorothiazide, respectively. The second method depends on second derivative of the ratio spectra by measurement of the amplitudes at 241.2 and 273.2 nm for benazepril hydrochloride and hydrochlorothiazide, respectively. Chemometric methods, classical least squares and principal component regression, were applied to analyze the mixture. Both the chemometric methods were applied to the zero and first order spectra of the mixture. The proposed methods were successfully applied for the determination of the two drugs in laboratory prepared mixtures and in commercial tablets. PMID- 11275438 TI - An LC-MS-MS method for the determination of indinavir, an HIV-1 protease inhibitor, in human plasma. AB - A method for the determination of indinavir (IDV) (L-735 524) in human plasma by LC-MS-MS is discussed, and the validation data is presented. The analyte and internal standard are isolated from plasma by a simple acetonitrile precipitation of plasma proteins followed by centrifugation. LC-tandem mass spectrometry in positive ion, multiple reaction monitoring mode used pairs of ions at m/z of 614/421 for indinavir and 628/421 for internal standard, respectively. The calibration curve had a linear range from 3.0 to 12320 ng/ml when linear least square regression weighing 1/x was applied to the concentration versus peak area plot. The advantages of this method are the fast sample preparation, wide dynamic assay range and quick analysis taking only 5 min for each sample run. The robust nature of this assay has been further verified during routine use over several months involving multiple analysts. PMID- 11275439 TI - Determination of partition coefficients of diazepam and flurazepam between phosphatidylcholine bilayer vesicles and water by second derivative spectrophotometric method. AB - Second derivative spectrophotometry allowed the establishment of a simple and accurate method for the determination of partition coefficients of benzodiazepine drugs in a liposome/water system. The absorption spectra of diazepam (DZ) and flurazepam (FZ) in phosphatidylcholine (egg yolk) bilayer vesicle suspensions showed small spectral changes depending on the concentration of phosphatidylcholine vesicles. However, the intense background signals caused by the light scattering of the phosphatidylcholine vesicles made it difficult to yield a correct base line, thus the quantitative spectral data could not be obtained. In the second derivative spectra, the spectral changes were enhanced and three derivative isosbestic points were observed for each drug indicating the entire elimination of the residual background signal effects. The derivative intensity change of each drug (DeltaD) induced by its interaction with phosphatidylcholine bilayers was measured at a specific wavelength. From the relationship between the DeltaD value and the lipid concentration, the molar partition coefficients (K(p)s) of DZ and FZ were calculated and obtained with a good precision of R.S.D below 10%. The fractions of the partitioned DZ and FZ calculated by using the obtained K(p) values agreed well with the experimental values. The results prove that the derivative method can be usefully and easily applied to the determination of partition coefficients of benzodiazepines in the liposomes/water system without any separation procedures. PMID- 11275440 TI - Simultaneous determination of amoxycillin and clavulanic acid in pharmaceutical dosage forms by LC with amperometric detection. AB - A simple and sensitive liquid chromatographic method with electrochemical (EC) detection is described for the quantitative determination of amoxycillin and clavulanic acid in pharmaceutical dosage forms. Sample components were separated by a reversed phase C18 column, using a mixture of methanol-phosphate buffer (pH 3.2-3.4) (5:95, v/v) as the mobile phase. Detection of antibiotics was performed amperometrically by applying a potential of +1.25 V. High linearity over a concentration range of 15.625-500 ng was demonstrated for amoxycillin (r=0.9999) and clavulanic acid (r=0.9979). Detection limits were 0.8 ng ml(-1) for amoxycillin and 15 ng ml(-1) for clavulanic acid. This method was found to be convenient and reproducible for analysis of these two components in oral suspensions and tablets and might be useful for other pharmaceutical dosage forms. PMID- 11275441 TI - Comparison of manual protein precipitation (PPT) versus a new small volume PPT 96 well filter plate to decrease sample preparation time. PMID- 11275442 TI - FIA of sildenafil citrate using UV-detection. AB - A flow injection analysis (FIA) of sildenafil citrate (SLD) using UV detection is described in this study. The best solvent system was found to be consisting of 0.2 M phosphate buffer at pH 8 having 10% MeOH. A flow rate of 1 ml. min(-1) was pumped and active material was detected at 292 nm. The calibration equation was linear in the range of 1x10(-6)-5x10(-6) M. Limit of detection (LOD) and limit of quantitation (LOQ) were calculated to be 3x10(-7) and 8.9x10(-7) M with a R.S.D. 1.9 and 0.6% (n=7), respectively. The proposed method was applied to the determination of SLD in VIAGRA tablet, containing 50 mg active material. The results were compared with those obtained from UV-Spectrophotometry. The results showed that there is a good agreement between FIA method and the UV Spectrophotometry. The validation studies were realised by the related applications and the results were evaluated statistically. According to the results, insignificant difference was observed between the methods. PMID- 11275443 TI - Enhancing primary care HIV prevention: a comprehensive clinical intervention. AB - CONTEXT: Human immunodeficiency virus (HIV) and sexually transmitted disease (STD) risk assessment and counseling are recommended for a large proportion of the population, yet measured rates of such counseling remain low. OBJECTIVE: Use a comprehensive intervention to improve and sustain rates of HIV/STD risk assessment and counseling by providers. DESIGN: Patient telephone survey using a one-group pre- and post-intervention design with measurements over a 62-week period. SETTING AND PARTICIPANTS: Patients (N=1042) from two outpatient clinics at a health maintenance organization (HMO) presenting for either of two types of index visit: symptomatic (n=210), or routine physical examination or birth control (n=832) visits. MAIN OUTCOME MEASURES: Telephone survey performed within 3 weeks of the index visit. Patients' recall of a general discussion of HIV/STDs and specific discussion of sexual behaviors/risk factors. RESULTS: The intervention was associated with increased patient recall of providers: discussing HIV/STD in general (OR 1.6; 95% CI, 1.12-2.22), asking about sexual behaviors/risk factors (OR 1.7; 95% CI, 1.2-2.6), discussing HIV prevention generally (OR 2.4; 95% CI, 1.4-4.0), and discussing personal risk reduction (OR 2.6; 95% CI, 1.6-4.3). Provision of written materials concerning HIV/STD also increased significantly (OR 2.8; 95% CI, 1.3-4.3). A clear-cut pattern of improved provider effort was seen, with the most pronounced improvements in high risk patients. Results were stable over a 38-week follow-up period. CONCLUSION: A sustained improvement in HIV/STD risk assessment and counseling can be achieved in an outpatient HMO setting using a relatively non-intensive systematized intervention. PMID- 11275444 TI - Maintaining prevention in practice: survival of PPIP in primary care settings. Put Prevention Into Practice. AB - INTRODUCTION: Put Prevention Into Practice (PPIP) consists of a kit of office based tools intended to support the provision of preventive services by primary care providers. The purpose of this study was to examine the institutionalization of PPIP within five primary care clinics funded by the Texas Department of Health to implement PPIP, and to examine the organizational determinants of program institutionalization. METHODS: We utilized an adaptation of the Level of Institutionalizaton (LoIn) scales for qualitative data collection and for development of an institutionalization score for each site. The determinants of institutionalization were derived from the organizational behavior and health promotion literatures and used as categories for analysis. In addition, for purposes of triangulation, chart audit data for three documentation behaviors were also collected. RESULTS: PPIP has been maintained--at varying degrees of integration--in four of the five sites studied, for 6 years after adoption. Organizational factors that facilitated the institutionalization process were the site's institutional strength, the integration of PIPP within extant programs and services, visibility of the program within and outside the site, planning for the termination of grant funding, and presence of a program champion with mid- to upper-level managerial authority. Successful initiation of the program was not a predictor of institutionalization outcomes. CONCLUSIONS: We have highlighted the need to consider organizational determinants of institutionalization in relation to their specific sociopolitical contexts, and in relation to each other, not in isolation. PMID- 11275445 TI - Evaluating the teaching of clinical preventive medicine: a multidimensional approach. AB - BACKGROUND: A study was undertaken to determine the amount, methods, and adequacy of instruction in clinical preventive medicine topics in the medical school curriculum at the University of California, San Francisco (UCSF) in the 1996-1997 academic year. METHODS: A protocol of 35 clinical preventive medicine topics was developed. The preclinical (Years 1 and 2) curriculum was evaluated by reviewing all syllabi and other printed materials for the presence and quantity of instruction in the specific clinical preventive medicine topics. The clinical curriculum (Years 3 and 4) was evaluated by asking students on completion of eight clinical clerkships to answer a questionnaire. Clerkship directors were also asked to answer the same questionnaire. RESULTS: In the preclinical curriculum, clinical preventive medicine topics were found to receive 63.3 hours of instruction (4.2% of total instruction hours). Counseling and screening topics received the most hours (31.3 and 20.5, respectively) with immunization/prophylaxis and prenatal care receiving considerably less (4.0 and 2.4 hours, respectively). In the clinical curriculum, students reported receiving an average of 118.5 hours of instruction in preventive medicine (5.9% of total instruction hours). Clerkship directors reported more than twice as many hours of instruction (330.8) as students. Overall, only 50% of students reported that a topic had been covered in a clerkship when the clerkship director reported that it had been covered. Both students and clerkship directors reported that exposure to clinical preventive medicine topics was in general inadequate. CONCLUSIONS: Instruction in clinical preventive medicine constituted a relatively modest percentage of the total instruction time in both the preclinical and clinical curricula at UCSF. Some topics were only minimally covered in the curriculum, and instruction during the clinical years was variable across students and clerkships. The disparity in the amount of instruction in clinical preventive medicine reported by students and faculty illustrates the importance of using multiple methods, including student input, to evaluate curriculum content. PMID- 11275446 TI - Cardiovascular disease and risk factors in Montana American Indians and non Indians. AB - BACKGROUND: Cardiovascular disease (CVD) is the leading cause of death for both American Indian and non-Indian adults. Few published studies have compared the prevalence of CVD and related risk factors in Indians to that in non-Indians in the same geographic area. OBJECTIVE: To compare CVD and risk factors in American Indian and non-Indian populations in Montana. METHODS: Adult American Indians (n=1000) living on or near Montana's seven reservations and non-Indian (n=905) Montanans statewide were interviewed through the 1999 Behavioral Risk Factor Surveillance Survey (BRFSS). RESULTS: Indians aged > or =45 years reported a significantly higher prevalence of CVD compared to non-Indians (18% vs 10%). In persons aged 18-44 years, Indians were more likely to report hypertension (15% vs 10%), obesity (29% vs 12%), and smoking (42% vs 24%) compared to non-Indians. For persons aged > or =45 years, Indians reported higher rates of diabetes (24% vs 9%), obesity (38% vs 16%), and smoking (32% vs 13%) compared to non-Indians. Non Indians aged > or =45 years reported having been diagnosed with high cholesterol more frequently than did Indians (32% vs 24%). CONCLUSIONS: Both Indians and non Indians in Montana reported a substantial burden of CVD. The CVD risk patterns differ in the two populations. Prevention programs should be tailored to the risk burdens in these communities with particular emphasis on smoking cessation and the prevention of obesity. PMID- 11275447 TI - Cigarette use by college students in smoke-free housing: results of a national study. AB - BACKGROUND: Cigarette-smoking rates have increased in recent years among college students. Smoke-free residences offer a possible means of reducing or preventing smoking. However, their use has as yet not been evaluated. This paper examines whether students residing in smoke-free residences are less likely to smoke cigarettes than students in other campus residences, and if such lower rates apply to all types of students and colleges. METHODS: The Harvard School of Public Health College Alcohol Study surveyed a nationally representative sample of college students at 128 U.S. 4-year colleges regarding tobacco use and related behaviors in the spring of 1999. The responses of students living in smoke-free and unrestricted residences at 101 campuses were compared. RESULTS: Current smoking prevalence was significantly lower among residents of smoke-free housing (21.0%) as compared with residents of unrestricted housing (30.6%, p<0.0001). The lower rate of current cigarette use was consistent with all types of student and college characteristics with few exceptions. Current cigarette use was significantly lower for those living in smoke-free housing than for residents of unrestricted housing among students who were not regular smokers before age 19 (10% vs 16.9%, p<0.0001), but not among students who smoked regularly before age 19. CONCLUSIONS: Smoke-free residences may help protect those students who were not regular smokers in high school from smoking in college. However, the difference in smoking rates may be due to self-selection of students into smoke free residences. Since smoke-free options also protect students from second-hand smoke and dormitory fires, colleges should provide these types of residences for all students who request them, and should also encourage others to choose them. PMID- 11275448 TI - A review of state legislation related to immunization registries. AB - BACKGROUND: Since the early 1990s, a concerted effort has been made to develop community- and state-based immunization registries. A 1995 survey showed that nine states had laws specifically authorizing immunization registries. This survey was conducted to describe the current status of legislation and policies addressing immunization registries and the sharing of immunization information. METHODS: A telephone survey was administered from September 1997 to February 1998 to immunization program managers and/or their designees within the state health department of each of the 50 states and the District of Columbia. Some of the survey items were later updated through follow-up interviews and informal communications. Copies of legislation, administrative rules and regulations, and immunization registry policies were collected for review. RESULTS: As of October 2000, 24 of 51 states (47%) had laws (21) or rules (3) specifically authorizing an immunization registry. Nine additional states (18%) have laws specifically addressing the sharing of immunization information. CONCLUSIONS: Over half of the states have enacted legislation or rules addressing registries or the sharing of immunization information. Further research should be conducted to assess the impact of this legislation on immunization registries. PMID- 11275449 TI - The risk of developing breast cancer within the next 5, 10, or 20 years of a woman's life. AB - BACKGROUND: The lifetime risk of developing breast cancer is a frequently misinterpreted statistic. Risk projections over a shorter time period, conditioned on current age, may be less prone to misconceptions and more relevant to populations at different ages. The purpose of this study was to estimate the risk of developing breast cancer within the next 5, 10, or 20 years for women currently aged 30 to 70 years in California's four major race/ethnic groups. METHODS: Life tables were used to obtain risk estimates based on 1993-1997 breast cancer incidence rates from the California Cancer Registry and statewide mortality rates. RESULTS: For women currently aged 50, the estimated risk of developing invasive breast cancer within 5 years varied from 0.8% (1 in 133) among Hispanics to 1.3% (1 in 75) among Caucasians. Risk estimates within 10 years were 2.9% (1 in 34) among Caucasians, 2.3% (1 in 43) among African Americans, 2.0% (1 in 51) among Asian/Pacific Islanders, and 1.6% (1 in 63) among Hispanics. Within 20 years, estimated risks increased to 6.6% (1 in 15) among Caucasians, 5.0% (1 in 20) among African Americans, 3.9% (1 in 26) among Asian/Pacific Islanders, and 3.7% (1 in 27) among Hispanics. Risk estimates were 8% to 20% higher when in situ tumors were included in the calculations. CONCLUSIONS: Based on these estimates, the baseline risk of developing breast cancer in the next 1 or 2 decades of life varies by race/ethnicity and current age, but may be lower than the risk perceived by most women. PMID- 11275451 TI - Trauma experience of North Korean refugees in China. AB - OBJECTIVES: We examined trauma experience and mental health conditions among North Korean migrants in China. METHODS: Personal interviews of 170 North Korean "food refugees" in China were conducted in 1999. Structured questionnaires were used to identify specific trauma experience and symptoms of post-traumatic stress, anxiety, and depression. RESULTS: All participants surveyed reported trauma exposure, with food and water shortage (93%) and illness without access to medical care (89%) being the most frequently cited events. Post-traumatic stress disorder was suspected in 56% of the respondents. Above-threshold scores on anxiety and depression were found in 90% and 81% of the cases, respectively. CONCLUSIONS: The burden on the part of North Korean migrants in China who experience trauma appears to be very high. An international call for action is warranted to monitor and safeguard the mental health status of this vulnerable segment of people and respond to their urgent psychiatric and medical care needs. PMID- 11275450 TI - Lyme disease and preventive behaviors in residents of Nantucket Island, Massachusetts. AB - BACKGROUND: To determine the age-specific prevalence of Lyme disease and whether preventive behaviors on Nantucket Island correlate with Lyme disease, we surveyed island residents. METHODS: A survey with questions on Lyme disease symptoms, history, and preventive behaviors was mailed to all residents. Respondents were stratified by likelihood of having had Lyme disease. A subsample was selected for examination, and then classified according to the Lyme disease national surveillance case definition. RESULTS: The overall lifetime prevalence of Lyme disease for Nantucket residents was 15% (CI, 10%-19.8%): 19% among females, and 11% among males. The prevalence was highest among age groups 0-16 and 30-49 years. Overall, 86% of the population practiced at least one behavior. The most frequently reported preventive behavior was checking oneself for ticks (80%), followed by wearing protective clothing (53%), avoiding tick areas (34%), and using tick repellent (11%). Younger individuals practiced fewer preventive behaviors than older individuals (p=0.001). Although males reported greater tick exposure than females, females uniformly practiced preventive behaviors more frequently (p=0.001). The practice of preventive behaviors was not associated with a history of Lyme disease, but finding more than 5 ticks per year on oneself was (p=0.001). CONCLUSION: Lyme disease is highly prevalent on Nantucket Island. Young people are particularly at risk and health education should emphasize preventive behaviors less frequently practiced: using tick repellent, avoiding tick areas, and wearing protective clothing. PMID- 11275452 TI - Risk factors for injury in rural Iowa: round one of the Keokuk County Rural Health Study. AB - BACKGROUND: Unintentional injuries represent a major cause of morbidity and mortality in rural communities. This study aimed to determine the distribution of injury risk factors in a rural Iowa community and to identify the rural subgroups at highest risk for injury. METHODS: We reported on 1583 participants, aged > or =25 years, from Round One of the Keokuk County Rural Health Study, a longitudinal panel study of a rural community. The self-reported data were collected during face-to-face interviews. RESULTS: Our data suggested that several risk factors for injury are not uniformly distributed among rural populations. Male farmers were significantly less likely to wear their seatbelts than townspeople or rural nonfarmers. However, farm women were as likely to wear seatbelts as other women. Both male and female farmers were more likely to use all-terrain vehicles than townspeople or rural nonfarmers. In contrast, townspeople were more likely to ride bicycles than either farmers or rural nonfarmers. Townspeople were less likely to have firearms in their homes than either farmers or rural nonfarmers. Farmers were most likely to have fired a gun in the last year. Male farmers aged <65 years were less than half as likely as other men the same age to report a history of alcohol abuse. Binge drinking was equally frequent among farmers, rural nonfarmers, and townspeople. CONCLUSIONS: These differences in risk behavior in a rural county suggest the possibility of targeting specific rural injury prevention interventions at those with the highest risk for dangerous behavior. PMID- 11275453 TI - Beyond toxicity: human health and the natural environment. AB - Research and teaching in environmental health have centered on the hazardous effects of various environmental exposures, such as toxic chemicals, radiation, and biological and physical agents. However, some kinds of environmental exposures may have positive health effects. According to E.O. Wilson's "biophilia" hypothesis, humans are innately attracted to other living organisms. Later authors have expanded this concept to suggest that humans have an innate bond with nature more generally. This implies that certain kinds of contact with the natural world may benefit health. Evidence supporting this hypothesis is presented from four aspects of the natural world: animals, plants, landscapes, and wilderness. Finally, the implications of this hypothesis for a broader agenda for environmental health, encompassing not only toxic outcomes but also salutary ones, are discussed. This agenda implies research on a range of potentially healthful environmental exposures, collaboration among professionals in a range of disciplines from public health to landscape architecture to city planning, and interventions based on research outcomes. PMID- 11275454 TI - Nature matters. PMID- 11275455 TI - Gone barefoot lately? PMID- 11275456 TI - Infantile spasms. AB - Infantile spasms constitute both a distinctive seizure type and an age-specific epilepsy syndrome that have been extensively described for over a century. Standardization of the classification of infantile spasms has evolved, culminating in recent recommendations for separately recognizing and distinguishing the seizure type (spasms or epileptic spasms) and the epilepsy syndrome of infantile spasms (West syndrome). More-detailed descriptions of the clinical and electrographic features of epileptic spasms and hypsarrhythmia have emerged. Advances in neuroimaging techniques have revealed clues about pathophysiology and increased the etiologic yield of the diagnostic evaluation of patients with infantile spasms. Adrenocorticotrophic hormone remains the treatment of choice for many neurologists. Recent controlled studies support vigabatrin as first-line therapy, and open-label studies suggest that topiramate, lamotrigine, and zonisamide may be useful in treating spasms. Recent reports of visual-field constriction with vigabatrin may limit its use. Surgical treatment has been used successfully in a select subgroup of patients with secondarily generalized spasms from a single epileptogenic zone. Although the prognosis for most patients with infantile spasms remains poor, further studies identifying predictors of favorable prognosis and recent advances in understanding the pathophysiology of infantile spasms offer hope of safer and more-effective therapies that improve long-term outcome. PMID- 11275457 TI - Stimulant therapy and seizure risk in children with ADHD. AB - Stimulants are an effective treatment frequently prescribed for attention-deficit hyperactivity disorder (ADHD), but they commonly are believed to lower the threshold for seizures. Although several studies have revealed that stimulants do not exacerbate well-controlled epilepsy, there is a paucity of data about seizure risk in nonepileptic children treated with stimulants. Two hundred thirty-four children (179 males, 9.1 +/- 3.6 years of age; 55 females, 9.6 +/- 3.9 years of age) with uncomplicated ADHD received electroencephalograms (EEGs) performed in our institution. Thirty-six patients (15.4%) demonstrated epileptiform abnormalities, and 198 (84.6%) demonstrated normal or nonepileptiform EEGs. Rolandic spikes accounted for 40% of the abnormal EEGs and 60% of those with focal abnormalities. Stimulant therapy was elected by 205 of 234 patients (87.6%). Seizures occurred only in the treated group, in one of 175 patients with a normal EEG (incidence 0.6%, 95% confidence intervals 0%-1.7%) and three of 30 treated patients with epileptiform EEGs (incidence 10%, 95% confidence interval 0%-20.7%). Seizures occurred in two of 12 children (16.7%) with rolandic spikes. These data suggest that a normal EEG can be used to assign children with ADHD to a category of minimal risk for seizure. In contrast, an epileptiform EEG in neurologically normal children with ADHD predicts considerable risk for the eventual occurrence of seizure. The risk, however, is not necessarily attributable to stimulant use. PMID- 11275458 TI - Postnatal adaptation of brain circulation in preterm infants. AB - Global and regional postnatal cerebral circulatory changes in stable preterm infants were studied, and their relation to brain injury was assessed. Thirty five preterm infants were studied on the first and second days of age. Cerebral blood flow (CBF) (mL/hg per min) and cerebral blood volume (CBV) (mL/hg) were measured using near-infrared spectroscopy. The cerebral blood flow velocity (cm/second) (peak systolic, diastolic flow, mean flow) and resistance index (RI) were determined in the internal carotid, anterior cerebral, and striate arteries by color Doppler flow imaging. Serial cerebral ultrasound studies were performed to detect changes in brain parenchymal echogenicity or intraventricular hemorrhage (IVH); the maximum severity of these findings was considered. CBF and cerebral blood flow velocity increased significantly with time, and such changes were independent of mean blood pressure, PO(2), PCO(2), hematocrit, or glycemia. In contrast, CBV and RI remained unchanged. According to the results of sonograms, no differences were found in postnatal CBF and cerebral blood flow velocity changes, regardless of whether patients had or did not have parenchymal lesions or IVH. However, higher CBV values were found on the second day in infants with IVH compared with infants without IVH. Early coupling of CBF and metabolic demands is independent of blood pressure. Improved venous return, instead of vasodilation, could be important in this adaptation. PMID- 11275459 TI - Relationship between clinical and genetic features in "inverted duplicated chromosome 15" patients. AB - Inverted duplicated chromosome 15 (Inv dup [15]) syndrome is a genetic disorder characterized by psychologic or intellectual language delay; neurologic signs, such as hypotonia, ataxia, and epilepsy; mental retardation ranging from mild to severe; and facial dysmorphisms. All patients present with a psychopathologic impairment that is highly variable in severity but always classifiable as pervasive developmental disorder (PDD). Many genetic mechanisms have been hypothesized to explain the clinical variability. This article describes the neurologic and psychopathologic features of six Inv dup(15) patients, one male and five females, between 8 and 14 years of age, all with a maternal marker chromosome. Four patients were diagnosed with PDD not otherwise specified, whereas two patients received a diagnosis of autism. Epilepsy was present in three patients (two generalized symptomatic and one focal symptomatic), and a correlation between the severity of the disease and its outcome was not always observed. Nevertheless, the influence of gene content of the marker chromosome, particularly the three gamma-aminobutyric acid-A receptor subunit genes, may represent the link between epilepsy, mental retardation, and PDD. PMID- 11275460 TI - Recurrent intracranial ependymoma in children: salvage therapy with oral etoposide. AB - Chronic oral VP-16 (etoposide) is a chemotherapy regimen with a wide application in oncology and documented efficacy against germ cell tumors, lymphomas, Kaposi's sarcoma, and primary brain tumors. This study was performed to assess the toxicity and activity of chronic oral etoposide in the management of children with recurrent intracranial nondisseminated ependymoma. Twelve children (median age of 8 years) with recurrent ependymoma who were refractory to surgery, radiotherapy, and chemotherapy (carboplatinum or the combination of procarbazine, lomustine, and vincristine) were treated with chronic oral etoposide (50 mg/m(2)/day). Treatment-related complications included the following: alopecia (10 children), diarrhea (6), weight loss (5), anemia (4), neutropenia (3), and thrombocytopenia (3). Three children required transfusion (two with packed red blood cells; two with platelets), and two children developed neutropenic fever. No treatment-related deaths occurred. Six children (50%) demonstrated either a radiographic response (two children, both with partial response) or stable disease (four children) with a median duration of response or stable disease of 7 months. In this small cohort of children with recurrent intracranial ependymoma, oral etoposide was well tolerated, produced modest toxicity, and had apparent activity. PMID- 11275461 TI - Cavernous sinus thrombophlebitis in children. AB - Eight Thai patients less than 15 years of age who were diagnosed with cavernous sinus thrombophlebitis at Ramathibodi Hospital, Bangkok, Thailand over the past 30 years were reviewed retrospectively. The predisposing infections and causative microorganisms were similar to previous reports in children and adults. Despite severe neurologic dysfunction during admission, including blindness, there was neither death nor severe permanent deficit found in the majority of the patients. Only one patient experienced mild hemiparesis caused by cerebral infarction, which was secondary to this condition. Early recognition of this condition, the appropriate selection of empirical antibiotic therapy, and the awareness of associated complication were the key factors leading to excellent outcome. PMID- 11275462 TI - Long-term sleep disturbances in adolescents after minor head injury. AB - It has been demonstrated that patients in the acute phase after minor head injury (MHI) complain of sleep disturbances. The purpose of the present study was to characterize the long-term effects of MHI on sleep in adolescents. Nineteen adolescents who had suffered MHI 3 years before the study and had complained of sleep disturbances completed a sleep questionnaire and were investigated in the sleep laboratory by whole-night polysomnographic recordings and were actigraphically monitored for 5 days at home. Questionnaire results revealed severe complaints regarding sleep behavior. Polysomnographic recordings revealed that in comparison with controls, MHI was associated with lower sleep efficiency (79.8 +/- [9.8]% vs 87.7 +/- [6.8]%; P < 0.005), with more wake time (10.6 +/- [9.0]% vs 3.4 +/- [4.4]%; P < 0.005), and with more awakenings lasting more than 3 minutes (2.1 +/- [1.5] vs 0.6 +/- [0.8]; P < 0.005). These findings were confirmed by actigraphic monitoring that revealed lower sleep efficiency (90 +/- [5]% vs 94 +/- [3]%; P < 0.05), more minutes of wake time (49 +/- [21] min vs 28 +/- [15] min; P < 0.05), and a trend toward more awakenings longer than 5 minutes (1.8 +/- [0.8] vs 1.2 +/- [0.8]; P = 0.063). Our data demonstrated that 3 years after MHI without any discernible clinical sequel, adolescents still complain of sleep disturbances that could be confirmed by both polysomnographic and actigraphic monitoring. PMID- 11275463 TI - A treatable cause of ataxia in children. AB - An 11-year-old black male presenting with severe subacute sensory ataxia, unusual skin hyperpigmentation, megaloblastic anemia, low serum B12 levels, and an abnormal part I Schilling test was diagnosed with pernicious anemia in the context of a polyglandular syndrome. Intrinsic factor and thyroid microsomal antibodies were positive, and thyroid-stimulating hormone levels were undetectable. There was a strong familial aggregation because the mother, a maternal aunt, the maternal grandfather, and the maternal great-grandmother had been diagnosed with pernicious anemia, albeit of unspecified etiology. Spinal magnetic resonance imaging (MRI) demonstrated extensive demyelination of the posterior columns along the entire length of the cord, as well as areas of contrast enhancement. Treatment with cobalamin produced complete remission of the neurologic deficits and normalization of the MRI findings in the short space of 2 months. Although rare, childhood pernicious anemia is a treatable disease that should be included in the differential diagnosis of the sensory ataxias in children. In this article, we review the causes of pernicious anemia in children and discuss the MRI findings. PMID- 11275464 TI - Improvement of atypical acute disseminated encephalomyelitis with steroids and intravenous immunoglobulins. AB - Acute disseminated encephalomyelitis is a demyelinating syndrome that occurs infrequently in children. Various treatment modalities, such as plasmapheresis or steroids or intravenous immunoglobulins (IVIG), have been prescribed. The article describes the results of combined IVIG and high-dose steroids given for 3 days in the treatment of a patient with atypical encephalomyelitis. The results suggest that this approach may be more beneficial than the application of either drug alone. PMID- 11275465 TI - Transient posterior encephalopathy induced by chemotherapy in children. AB - The cases of three children, 16, 12, and 12 years of age, who suffered sudden confusional state and cortical blindness lasting 12 to 30 minutes while under treatment with high-dose methotrexate, cyclophosphamide, and dactinomycin for a lower limb osteosarcoma are reported. Transient neuropsychologic deficits arose after the acute phase of treatment: left hemispatial neglect and constructive apraxia (Patient 1); constructive apraxia (Patient 2); and constructive apraxia and alexia without aphasia (Patient 3). The three patients recovered completely from all their deficits within the time frame of 3 hours to 2 weeks. Arterial hypertension and hypomagnesemia were found during the acute phase in all patients. In Patients 2 and 3, magnetic resonance imaging revealed increased parieto-occipital T(2) signal involving gray and white matter. In Patients 1 and 2, HmPAO-SPECT revealed parieto-occipital hypoperfusion that resolved a few days later. The alterations detected by neuroimaging were concurrent with the appearance and disappearance of the clinical symptoms. Such transient acute episodes have been named occipital-parietal encephalopathy. On the basis of our clinical, laboratory, and neuroimaging findings, an explanation for the origin of this syndrome, a migrainelike mechanism, triggered by chemotherapy-induced hypomagnesemia, is proposed. PMID- 11275466 TI - Multilocular hydrocephalus: ultrasound studies of origin and development. AB - Multilocular hydrocephalus is a complication of neonatal hydrocephalus. Its main feature is the presence of multiple cysts inside the ventricles, which requires a specific therapeutic approach. The case of a preterm infant with intracranial hemorrhage grade II-III and central nervous system infection is reported. The cysts developed at the subependymal layer in the posterior area of the patient's thalamus. Their growth and development were charted by ultrasound imaging for several weeks. These types of cysts may grow to occupy the totality of the lateral ventricles, isolating the temporal horns. Of all the reviewed pathogenic mechanisms, we support the hypothesis of an inflammatory vasculitis at the subependymal level, with the subsequent infarct giving rise to the cysts. The osmotic pressure within the cavities, rather than intraventricular fluid, would account for the enlargement of the cysts. PMID- 11275467 TI - Worster-Drought and congenital perisylvian syndromes-a continuum? AB - A 5-year-old female was evaluated because of severe speech and expressive language delay. On examination, she could hardly speak and communicated through gestures. She manifested severe dysarthria and difficulty in protruding and moving her tongue laterally. She lacked coordination of the swallowing process, with drooling and an increased mental reflex. Her cognitive development was normal, and no associated neurologic dysfunction of the limbs was noted. On follow-up, the child experienced two episodes of seizures at 6 years of age. Magnetic resonance imaging of the brain demonstrated perisylvian and frontal polymicrogyria. Clinical and radiologic findings demonstrated a similarity and continuum between congenital suprabulbar paresis (Worster-Drought syndrome) and perisylvian syndrome. PMID- 11275468 TI - Intrathecal baclofen in X-linked adrenoleukodystrophy. AB - X-linked adrenoleukodystrophy is a progressive neurodegenerative disorder involving the destruction of white matter in the brain and adrenocortical hormone deficiency. Clinical symptoms first appear between 4 and 8 years of age and include spasticity, visual loss, dysphagia, and seizures. In this report, continuous infusion of intrathecal baclofen was used to treat the severe spasticity of an 8-year-old patient with X-linked adrenoleukodystrophy. The improvement in this patient's quality of life, including the elimination of pain and the increased ease of care, suggests that intrathecal baclofen should be considered as part of the treatment strategy for spasticity associated with X linked adrenoleukodystrophy and other neurodegenerative disorders in children and adults. PMID- 11275469 TI - Functional neuroradiologic investigations in band heterotopia. AB - Band heterotopias are an example of genetic generalized neuronal migration disorders that may be present in patients with mild epilepsy and normal or slightly impaired intellect, as well as in patients with intractable epilepsy and mental retardation. The case of a 17-year-old left-handed female patient with epilepsy and normal cognitive development is reported in whom single-photon emission computed tomography (SPECT), proton magnetic resonance spectroscopy, and functional magnetic resonance imaging (fMRI) were performed. MRI revealed the presence of bilateral asymmetric band heterotopia. SPECT revealed a left frontoparietal and occipital hypoperfusion, demonstrating a good correlation with the electroencephalogram abnormalities. Because of the appearance of new types of seizures, the patient underwent a second MRI investigation together with a proton magnetic resonance spectroscopy (MRS) study. MRI confirmed bilateral band heterotopia characterized by greater thickness in the left hemisphere at the frontal and occipital level. MRI and SPECT findings were in agreement with left occipital electroencephalogram abnormalities and with occipital seizure type. Qualitative results of proton MRS revealed normal spectra profiles in the examined left frontal and occipital heterotopic area and in the normal overlying cortex. Later, fMRI was performed. The finger-tapping test of the right hand yielded the activation of both normal left sensory-motor cortex and the facing band heterotopia. In the right hemisphere, only the activation of the sensory motor neocortex was observed; no involvement of the right misplaced brain tissue was present. This functional behavior could be considered the consequence of poor neuronal representation. On the contrary, the involvement of both band heterotopia and normal cortex observed in the left hemisphere could be the result of many synaptic interconnections. Functional investigations may have an important role in defining the activity of band heterotopia per se and in relation to the overlying neocortex. PMID- 11275470 TI - Modulation of catalase peroxidatic and catalatic activity by nitric oxide. AB - Previously, we found that catalase enhanced the protection afforded by superoxide dismutase to Escherichia coli against the simultaneous generation of superoxide and nitric oxide (Brunelli et al., Arch. Biochem. Biophys. 316:327-334, 1995). Hydrogen peroxide itself was not toxic in this system in the presence or absence of superoxide dismutase. We therefore investigated whether catalase might consume nitric oxide in addition to hydrogen peroxide. Catalase rapidly formed a reversible complex stoichiometrically with nitric oxide with the Soret band shifting from 406 to 426 nm and two new peaks appeared at 540 and at 575 nm, consistent with the formation of a ferrous-nitrosyl complex. Catalase consumed more nitric oxide upon the addition of hydrogen peroxide. Conversely, micromolar concentrations of nitric oxide slowed the catalase-mediated decomposition of hydrogen peroxide. Catalase pretreated with nitric oxide and hydrogen peroxide regained full activity after dialysis. Our results suggest that catalase can slowly consume nitric oxide while nitric oxide modestly inhibits catalase dependent scavenging of hydrogen peroxide. The protective effects of catalase in combination with superoxide dismutase may result from two actions; reducing peroxynitrite formation by scavenging nitric oxide and by scavenging hydrogen peroxide before it reacts with superoxide dismutase to form additional superoxide. PMID- 11275471 TI - Blood glutathione redox status in gestational hypertension. AB - Gestational hypertension during the third trimester reflects an exaggerated maternal inflammatory response to pregnancy. We hypothesized that oxidative stress present even in normal pregnancy becomes uncompensated in hypertensive patients. A glucose-6-phosphate dehydrogenase (G6PD) activity sufficient to meet the increased reductive equivalent need of the cells is indispensable for defense against oxidative stress. The erythrocyte glutathione redox system was studied, where G6PD is the only NADPH source. The glutathione (GSH) redox status was measured both in vivo and after an in vitro oxidative challenge in pregnant women with gestational hypertension (n = 19) vs. normotensive pregnant subjects (n = 18) and controls (n = 20). An erythrocyte GSH depletion with an increase in the oxidized form (GSSG) resulted in an elevated ratio GSSG/GSH (0.305 +/- 0.057; mean +/- SD) in hypertensive pregnant women vs. normotensive pregnant or control subjects (0.154 +/- 0.025; 0.168 +/- 0.073; p <.001). In hypertensive pregnant patients, a "GSH stability" decrease after an in vitro oxidative challenge suggested a reduced GSH recycling capacity resulting from an insufficient NADPH supply. The erythrocyte GSSG/GSH ratio may serve as an early and sensitive parameter of the oxidative imbalance and a relevant target for future clinical trials to control the effects of antioxidant treatment in women at increased risk of the pre-eclampsia syndrome. PMID- 11275472 TI - Modulation of ceramide-induced NF-kappaB binding activity and apoptotic response by caffeic acid in U937 cells: comparison with other antioxidants. AB - Ceramide acts as second messenger in the signal transduction triggered by a variety of stress stimuli and extracellular agents. Stress response through ceramide is involved in the development of many human diseases, such as atherosclerosis, inflammation, neurodegenerative disorders, and acquired immunodeficiency syndrome. Dietary polyphenols have been reported to exert a beneficial effect on the onset and development of most of these human chronic degenerative pathologies. However, the mechanisms underlying this beneficial effect are mostly not understood at the present. To investigate the ability of polyphenols in modulating fundamental cellular functions, we studied the effect of caffeic acid, a widespread phenolic acid largely present in human diet, in the modulation of ceramide-induced signal transduction pathway leading to apoptosis in U937 cells, in comparison with other established antioxidants of nutritional interest (N-acetylcysteine, d-alpha-tocopherol acetate and ascorbic acid). Our results indicate that caffeic acid efficiently inhibits both ceramide-induced NF kappaB binding activity and apoptosis at micromolar concentration. Other antioxidants tested are totally ineffective in inhibiting apoptosis, although affecting NF-kappaB activation. Caffeic acid was found to inhibit protein tyrosine kinase activity, suggesting that this mechanism can be on the basis of the inhibition of apoptosis. Our results suggest that dietary caffeic acid might modulate ceramide-induced signal transduction pathway and NF-kappaB activation through either antioxidant and nonantioxidant mechanisms. PMID- 11275473 TI - Spin trapping agents (Tempol and POBN) protect HepG2 cells overexpressing CYP2E1 against arachidonic acid toxicity. AB - Polyunsaturated fatty acids such as arachidonic acid were previously shown to be toxic to HepG2 cells expressing CYP2E1 by a mechanism involving oxidative stress and lipid peroxidation. This study investigated the effects of the spin trapping agents Tempol and POBN on the arachidonic acid toxicity. Arachidonic acid caused toxicity and induced lipid peroxidation and mitochondrial membrane damage in cells overexpressing CYP2E1 but had little or no effect in control cells not expressing CYP2E1. The toxicity appeared to be both apoptotic and necrotic in nature. 4-Hydroxy-[2,2,6,6-tetramethylpiperidine-1-oxyl] (Tempol) and alpha-(4 pyridyl-1-oxide)-N-tert-butyl nitrone (POBN) protected against the decrease in cell viability and the apoptosis and necrosis. These spin traps prevented the enhanced lipid peroxidation and the loss of mitochondrial membrane potential. Tempol and POBN had little or no effect on cellular viability or on CYP2E1 activity at concentrations which were protective. It is proposed that elevated production of reactive oxygen intermediates by cells expressing CYP2E1 can cause lipid peroxidation, which subsequently damages the mitochondrial membrane leading to a loss in cell viability when the cells are enriched with arachidonic acid. Tempol and POBN, which scavenge various radical intermediates, prevent in this way the enhanced lipid peroxidation, mitochondrial dysfunction, and the cell toxicity. Since oxidative stress appears to play a key role in ethanol hepatotoxicity, it may be of interest to evaluate whether such spin trapping agents are useful candidates for the prevention or improvement of ethanol-induced liver injury. PMID- 11275475 TI - Measurement of 8-oxo-2'-deoxyguanosine and 8-oxo-2'-deoxyadenosine in DNA and human urine by high performance liquid chromatography-electrospray tandem mass spectrometry. AB - A method for the determination of 8-oxo-2'-deoxyguanosine and 8-oxo-2' deoxyadenosine in DNA and urine by High Performance Liquid Chromatography (HPLC) Tandem Mass Spectrometry is described. For the urine samples there is no sample preparation except for addition of buffer and internal standards followed by redissolvation of precipitate containing 8-oxo-2'-deoxyguanosine and a centrifugation step before the samples are injected onto the HPLC column. The detection limit for 8-oxo-2'-deoxyguanosine and 8-oxo-2'-deoxyadenosine is approximately 0.3 nM corresponding to 7.5 fmol injected. Long runs, that is, > 50 samples, can be analyzed with only minimal loss of sensitivity. The concentrations excreted into urine samples from humans are between 1 and 100 nM for 8-oxo-2'-deoxyguanosine and below 0.3 nM for 8-oxo-2'-deoxyadenosine. In calf thymus DNA levels down to about 1 oxidized guanosine and adenosine per 10(6) unmodified bases can be detected. High levels of 8-oxo-2'-deoxyguanosine were found, 30 per 10(6) 2'-deoxyguanosine, levels of 8-oxo-2'-deoxyadenosine are at or below the detection limit. These findings indicate that High Performance Liquid Chromatography-Tandem Mass Spectrometry is a highly sensitive and specific method for analysis of oxidative DNA modifications in tissue as well as for analysis of excretion of oxidized nucleotides into urine that ensures a minimum artifact formation. PMID- 11275474 TI - Differential regulation of NO availability from macrophages and endothelial cells by the garlic component S-allyl cysteine. AB - Garlic has been used as a traditional medicine for prevention and treatment of cardiovascular diseases. However, the molecular mechanism of garlic's pharmacological action has not been clearly elucidated. We examined here the effect of garlic extract and its major component, S-allyl cysteine (SAC), on nitric oxide (NO) production by macrophages and endothelial cells. The present study demonstrates that these reagents inhibited NO production through the suppression of iNOS mRNA and protein expression in the murine macrophage cell line RAW264.7, which had been stimulated with LPS and IFNgamma. The garlic extract also inhibited NO production in peritoneal macrophages, rat hepatocytes, and rat aortic smooth muscle cells stimulated with LPS plus cytokines, but it did not inhibit NO production in iNOS-transfected AKN-1 cells or iNOS enzyme activity. These reagents suppressed NF-kappaB activation and murine iNOS promoter activity in LPS and IFNgamma-stimulated RAW264.7 cells. In contrast, these reagents significantly increased cGMP production by eNOS in HUVEC without changes in activity, protein levels, and cellular distribution of eNOS. Finally, garlic extract and SAC both suppressed the production of hydroxyl radical, confirming their antioxidant activity. These data demonstrate that garlic extract and SAC, due to their antioxidant activity, differentially regulate NO production by inhibiting iNOS expression in macrophages while increasing NO in endothelial cells. Thus, this selective regulation may contribute to the anti-inflammatory effect and prevention of atherosclerosis by these reagents. PMID- 11275477 TI - Identification and quantification of 8,5'-cyclo-2'-deoxy-adenosine in DNA by liquid chromatography/ mass spectrometry. AB - Recent studies suggested that 8,5'-cyclo-2'-deoxyadenosine may play a role in diseases with defective nucleotide-excision repair. This compound is one of the major lesions, which is formed in DNA by hydroxyl radical attack on the sugar moiety of 2'-deoxyadenosine. It is likely to be repaired by nucleotide-excision repair rather than by base-excision repair because of a covalent bond between the sugar and base moieties. We studied the measurement of 8,5'-cyclo-2' deoxyadenosine in DNA by liquid chromatography/isotope-dilution mass spectrometry. A methodology was developed for the analysis of 8,5'-cyclo-2' deoxyadenosine by liquid chromatography in DNA hydrolyzed to nucleosides by a combination of four enzymes, i.e., DNase I, phosphodiesterases I and II, and alkaline phosphatase. Detection by mass spectrometry was performed using atmospheric pressure ionization-electrospray process in the positive ionization mode. Results showed that liquid chromatography/isotope-dilution mass spectrometry is well suited for identification and quantification of 8,5'-cyclo 2'-deoxyadenosine in DNA. Both (5'R)- and (5'S)-diastereomers of 8,5'-cyclo-2' deoxyadenosine were detected. The level of sensitivity of liquid chromatography/mass spectrometry with selected-ion monitoring amounted to 2 fmol of this compound on the column. The yield of 8,5'-cyclo-2'-deoxyadenosine was measured in DNA in aqueous solution exposed to ionizing radiation at doses from 2.5 to 80 Gray. Gas chromatography/mass spectrometry was also used to measure this compound in DNA. Both techniques yielded similar results. The yield of 8,5' cyclo-2'-deoxyadenosine was comparable to the yields of some of the other major modified bases in DNA, which were measured using gas chromatography/mass spectrometry. The measurement of 8,5'-cyclo-2'-deoxyadenosine by liquid chromatography/mass spectrometry may contribute to the understanding of its biological properties and its role in diseases with defective nucleotide-excision repair. PMID- 11275476 TI - Mechanism of oxidative DNA damage induced by carcinogenic 4-aminobiphenyl. AB - DNA adduct formation is thought to be a major cause of DNA damage by carcinogenic aromatic amines. We investigated the ability of an aromatic amine, 4 aminobiphenyl (4-ABP) and its N-hydroxy metabolite (4-ABP(NHOH)) to cause oxidative DNA damage, using (32)P-labeled human DNA fragments from the p53 tumor suppressor gene and the c-Ha-ras-1 protooncogene. 4-ABP(NHOH) was found to cause Cu(II)-mediated DNA damage, especially at thymine residues. Addition of the endogenous reductant NADH led to dramatic enhancement of this process. Catalase and bathocuproine, a Cu(I)-specific chelator, reduced the amount of DNA damage, suggesting the involvement of H(2)O(2) and Cu(I). 4-ABP(NHOH) dose-dependently induced 8-hydroxy-2'-deoxyguanosine (8-OHdG) formation in the presence of Cu(ll) and NADH. 4-ABP(NHOH) conversion to nitrosobiphenyl, as measured by UV-visible spectroscopy, occurred rapidly in the presence of Cu(II), suggesting Cu(II) mediated autoxidation. Increased amounts of 8-OHdG were found in HL-60 cells compared to the H(2)O(2)-resistant clone HP100 following 4-ABP(NHOH) treatment, further supporting the involvement of H(2)O(2). The present study demonstrates that an N-hydroxy derivative of 4-ABP induces oxidative DNA damage through H(2)O(2) in both a cell-free system and in cultured human cells. We conclude that, in addition to DNA adduct formation, oxidative DNA damage may play an important role in the carcinogenic process of 4-ABP. PMID- 11275478 TI - Physiological oxidative stress model: Syrian hamster Harderian gland-sex differences in antioxidant enzymes. AB - The Syrian hamster Harderian gland, a juxtaorbital organ exhibiting marked gender associated differences in contents of porphyrins and melatonin, was used as a model system for comparing strong (in females) and moderate (in males) physiological oxidative stress. Histological differences showing much higher cell damage in females were studied in conjunction with lipid peroxidation and activities of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase. Lipid peroxidation and enzyme activities were measured throughout the circadian cycle, revealing the importance of dynamical processes in oxidative stress. Especially in lipid peroxidation and in catalase, short lasting rises exhibited strongest gender differences. Peaks of lipid peroxidation were about three times higher in females, compared to males. Catalase peaks of females exceeded those in males by several hundred-fold. Average levels of superoxide dismutase and glutathione peroxidase were about three or two times higher in females, respectively. A clear-cut diurnally peaking rhythm was found in glutathione peroxidase of females, which was not apparent in males. Glutathione reductase showed differences in time patterns, but less in average activities. The time courses of lipid peroxidation and of protective enzymes are not explained by circulating melatonin, whereas melatonin formed in the Harderian gland should contribute to differences in average levels. Neither damage nor antioxidative defense simply reflect the illumination cycle and are, therefore, not only a consequence of photoreactions. PMID- 11275479 TI - Programmed cell death induced by glutathione depletion in PC 12 cells is blocked by inhibitors of 12 lipoxygenase, but does not appear to be mediated through the formation of 12 HETE derivatives. AB - Lipoxygenase metabolites have been postulated to be involved in the degenerative events provoked by oxidative stress in neuronal and nonneuronal targets, but their roles remain controversial. In the present work, we investigated the putative role of 12 lipoxygenase metabolites in the programmed cell death induced by glutathione depletion in PC 12 cells. Determinations of 12 lipoxygenase expression and activity reveal the presence of the enzyme in PC 12 cells, but the formation of arachidonate metabolites appears rather low and is not influenced by glutathione depletion. In addition, although the death induced by buthionine sulfoximine (BSO) treatment is abolished by known inhibitors of lipoxygenase enzymes, dexamethasone, a potent steroidal inhibitor of both cyclooxygenase and lipoxygenase pathways, fails to protect the cells from BSO-induced degeneration. Finally, incubation of the cells for 24 h in the presence of exogenous 12 HETE did not induce any significant decrease in cell viability. Our results indicate that 12 lipoxygenase is unlikely to play a major role in the process of cell degeneration provoked by glutathione depletion. PMID- 11275480 TI - Nitroxyl oxidizes NADPH in a superoxide dismutase inhibitable manner. AB - Nitric oxide synthases (NOS) convert L-arginine and N(omega)-hydroxy-L-arginine to nitric oxide (*NO) and/or nitroxyl (NO(-)) in a NADPH-dependent fashion. Subsequently, *NO/superoxide (O(2-)-derived peroxynitrite (ONOO(-)) consumes one additional mol NADPH. The related stoichiometry of NO(-) and NADPH is unclear. We here describe that NO(-) also oxidizes NADPH in a concentration-dependent manner. In the presence of superoxide dismutase (SOD), which also converts NO(-) to *NO, nitrite accumulation was almost doubled and no oxidation of NADPH was observed. Nitrate yield from NO(-) was low, arguing against intermediate ONOO(-) formation. Thus, biologically formed NO(-) may function as an effective pro-oxidant unless scavenged by SOD and affect the apparent NADPH stoichiometry of the NOS reaction. PMID- 11275481 TI - Chemical network of the living human brain. Evidence of reorganization with aging. AB - We recently described the chemical network properties of the human brain using in vivo proton magnetic resonance spectroscopy ((1)H MRS). In a separate study of aging we found increased concentration of chemicals in the prefrontal and sensorimotor cortices up to the third decade of life, and subsequent decrease of chemical concentrations in the same brain regions after the third decade between young and middle age. We anticipated that these age-dependent differences in chemical concentrations might be a reflection of the chemical network reorganization of the brain during aging. The pattern of chemical connectivity within and across brain regions for all regional chemicals, and specific patterns of chemical connectivity for each chemical type were examined for young and middle age groups using (1)H MRS and correlation analysis. For all studied ages, the dominant positive correlations occurred within brain regions and negative correlations were seen across brain regions. However, the pattern of negative chemical connectivity across brain regions was weaker in middle-aged group (F = 40.4, P < 10(-7) comparing r-values between the two age groups, ANOVA). Within brain regions, the age effects on chemical correlations were seen in the cingulate cortex (46% decrease in the middle-aged group, F = 7.2, P < 0.007) and sensorimotor cortex (SMC) (27% decrease, F = 8.9, P<0.003). Between brain regions, the age effects on chemical correlations were seen in the chemical interactions between the thalamus (433.3% increase in the middle-aged group, F = 11.7, P < 0.003), SMC (280% increase, F=20.1, P < 10(-5)), cingulate cortex (100.7% increase, F = 21.3, P < 10(-7)), and other brain regions. We found also age-differential patterns of chemical connectivity across the studied brain regions for most chemical types. The results provide evidence that normal aging is associated with reorganization of chemical network of the human brain. PMID- 11275482 TI - What's left if the Jabberwock gets the semantics? An ERP investigation into semantic and syntactic processes during auditory sentence comprehension. AB - This study examined auditory ERP responses to syntactic phrase structure violations occurring either in sentences containing regular words or in sentences in which content words had been replaced by pseudowords while retaining morphological markers (so-called jabberwocky sentences). Syntactic violations were found to elicit an early anterior negativity followed by a P600 for both types of sentences, suggesting that the syntactic processes in question are independent of the presence of lexical-semantic information. In syntactically correct sentences, content words in regular sentences elicited an N400 component while their pseudoword place-holders in jabberwocky sentences did not. By contrast, in syntactically incorrect sentences neither sentence type showed an N400 for the word creating the syntactic violation, indicating that the detection of a syntactic error at an early stage blocks semantic integration processes in regular sentences. We discuss these results and findings from related studies in the light of a timing hypothesis of syntactic and semantic information processing and propose that syntactic information extracted particularly early can affect semantic processes while syntactic information extracted relatively late cannot. PMID- 11275484 TI - Intracranial identification of an electric frontal-cortex response to auditory stimulus change: a case study. AB - The aim of the present study was to clarify whether ERPs recorded directly from the human frontal cortex contributed to the auditory N1 and mismatch negativity (MMN) elicited by changes in non-phonetic and phonetic sounds. We examined the role of prefrontal cortex in the processing of stimulus repetition and change in a 6-year-old child undergoing presurgical evaluation for epilepsy. EEG was recorded from three bilateral sub-dural electrode strips located over lateral prefrontal areas during unattended auditory stimulation. EEG epochs were averaged to obtain event-related potentials (ERPs) to repeating (standard) tones and to infrequent (deviant) shorter duration tones and complex sounds (telephone buzz). In another condition, ERPs were recorded to standard and deviant syllables, /ba/ and /da/, respectively. ERPs to vibration stimuli delivered to the fingertips were not observed at any of the sub-dural electrodes, confirming modality specificity of the auditory responses. Focal auditory ERPs consisting of P100 and N150 deflections were recorded to both tones and phonemes over the right lateral prefrontal cortex. These responses were insensitive to the serial position of the repeating sound in the stimulus train. Deviant tones evoked an MMN peaking at around 128 ms. Deviant complex sounds evoked ERPs with a similar onset latency and morphology but with an approximately two-fold increase in peak-to-peak amplitude. We conclude that right lateral prefrontal cortex (Brodmann's area 45) is involved in early stages of processing repeating sounds and sound changes. PMID- 11275483 TI - Neural substrates of facial emotion processing using fMRI. AB - We identified human brain regions involved in the perception of sad, frightened, happy, angry, and neutral facial expressions using functional magnetic resonance imaging (fMRI). Twenty-one healthy right-handed adult volunteers (11 men, 10 women; aged 18-45; mean age 21.6 years) participated in four separate runs, one for each of the four emotions. Participants viewed blocks of emotionally expressive faces alternating with blocks of neutral faces and scrambled images. In comparison with scrambled images, neutral faces activated the fusiform gyri, the right lateral occipital gyrus, the right superior temporal sulcus, the inferior frontal gyri, and the amygdala/entorhinal cortex. In comparisons of emotional and neutral faces, we found that (1) emotional faces elicit increased activation in a subset of cortical regions involved in neutral face processing and in areas not activated by neutral faces; (2) differences in activation as a function of emotion category were most evident in the frontal lobes; (3) men showed a differential neural response depending upon the emotion expressed but women did not. PMID- 11275485 TI - An ERD mapping study of the neurocognitive processes involved in the perceptual and semantic analysis of environmental sounds and words. AB - The aim of this paper was to investigate and compare the EEG mechanisms underlying the perceptual and semantic processes involved in environmental and language sounds perception by manipulating the degree of identification of sounds and using the ERD (event-related desynchronization) method in healthy subjects. Four types of stimuli were analyzed: meaningful environmental sounds, meaningless sounds, words and non-words. We report many similarities in the ERDs and ERSs (event-related synchronizations) patterns among all stimuli, with: (i) similar time-course of ERDs and ERSs between meaningful environmental sounds and words, and between meaningless sounds and non-words; (ii) similar topography of the maximal ERDs for meaningful environmental sounds, words and non-words; and (iii) same right posterior ERSs for all four stimuli. However, differences were also observed: (i) in time-course, with earlier ERSs for meaningless than meaningful stimuli, whether environmental or verbal; and (ii) in topography, with ERDs predominating in left and right hemisphere channels for meaningful and meaningless environmental sounds, respectively; (iii) ERSs predominating in the left temporal channel for non-words and in the left posterior and right frontal channels for meaningless sounds. The results of this study suggest that meaningful stimuli involve greater and longer-lasting semantic processes than meaningless stimuli, while the occurrence of ERSs for the latter points to the possible involvement of an inhibitory processing of semantic representations. Finally, the findings concerning the comparison between verbal and non verbal stimuli suggest the involvement of left-lateralized phonological and semantic processes for the former, and more distributed neurocognitive processes in both hemispheres for the latter although with predominant left laterality for their semantic processing. PMID- 11275486 TI - False recognition depends on depth of prior word processing: a magnetoencephalographic (MEG) study. AB - Brain activity was measured with a whole head magnetoencephalograph (MEG) during the test phases of word recognition experiments. Healthy young subjects had to discriminate between previously presented and new words. During prior study phases two different levels of word processing were provided according to two different kinds of instructions (shallow and deep encoding). Event-related fields (ERFs) associated with falsely recognized words (false alarms) were found to depend on the depth of processing during the prior study phase. False alarms elicited higher brain activity (as reflected by dipole strength) in case of prior deep encoding as compared to shallow encoding between 300 and 500 ms after stimulus onset at temporal brain areas. Between 500 and 700 ms we found evidence for differences in the involvement of neural structures related to both conditions of false alarms. Furthermore, the number of false alarms was found to depend on depth of processing. Shallow encoding led to a higher number of false alarms than deep encoding. All data are discussed as strong support for the ideas that a certain level of word processing is performed by a distinct set of neural systems and that the same neural systems which encode information are reactivated during the retrieval. PMID- 11275487 TI - Influence of action and expectation on visual control of posture. AB - Previous studies have shown that human subjects presented with a moving visual environment initiate a postural re-adjustment in the direction of motion. The present study investigated how active control or expectation of the displacement of a visual scene affects this postural response. Center of foot pressure (COP) and head displacement were recorded using a sway platform and a tracking system, respectively. The subjects faced a visual scene (1 x 1 m, at a distance of 45 cm) which moved transiently (with a velocity of 1 cm/s) in a direction parallel to the interaural axis. When the displacement of the visual scene was under the active control of the subjects, visually induced body sway was strongly inhibited, in comparison with the response to unexpected stimuli. Prior knowledge of the characteristics of the forthcoming displacement was sufficient, in most subjects, to reduce postural re-adjustment, even when subjects did not exert active control. Finally, the visually induced postural response was strongly reduced even when subjects only triggered the stimulus, without any knowledge about the direction of motion. In conclusion, it appears that although vision is of primary importance in the control of postural orientation, high level processes such as expectation can modulate its impact by providing cues as to whether forthcoming visual flow is the consequence of self-motion or object motion. PMID- 11275488 TI - Bisensory stimulation increases gamma-responses over multiple cortical regions. AB - In the framework of the discussion about gamma (approx. 40 Hz) oscillations as information carriers in the brain, we investigated the relationship between gamma responses in the EEG and intersensory association. Auditory evoked potentials (AEPs) and visual evoked potentials (VEPs) were compared with bisensory evoked potentials (BEPs; simultaneous auditory and visual stimulation) in 15 subjects. Gamma responses in AEPs, VEPs and BEPs were assessed by means of wavelet decomposition. Overall maximum gamma-components post-stimulus were highest in BEPs (P < 0.01). Bisensory evoked gamma-responses also showed significant central, parietal and occipital amplitude-increases (P < 0.001, P < 0.01, P < 0.05, respectively; prestimulus interval as baseline). These were of greater magnitude when compared with the unisensory responses. As a correlate of the marked gamma responses to bimodal stimulation we suggest a process of 'intersensory association', i.e. one of the steps between sensory transmission and perception. Our data may be interpreted as a further example of function related gamma responses in the EEG. PMID- 11275489 TI - Frontal midline theta rhythm is correlated with cardiac autonomic activities during the performance of an attention demanding meditation procedure. AB - Frontal midline theta rhythm (Fm theta), recognized as distinct theta activity on EEG in the frontal midline area, reflects mental concentration as well as meditative state or relief from anxiety. Attentional network in anterior frontal lobes including anterior cingulate cortex is suspected to be the generator of this activity, and the regulative function of the frontal neural network over autonomic nervous system (ANS) during cognitive process is suggested. However no studies have examined peripheral autonomic activities during Fm theta induction, and interaction of central and peripheral mechanism associated with Fm theta remains unclear. In the present study, a standard procedure of Zen meditation requiring sustained attention and breath control was employed as the task to provoke Fm theta, and simultaneous EEG and ECG recordings were performed. For the subjects in which Fm theta activities were provoked (six men, six women, 48% of the total subjects), peripheral autonomic activities were evaluated during the appearance of Fm theta as well as during control periods. Successive inter-beat intervals were measured from the ECG, and a recently developed method of analysis by Toichi et al. (J. Auton. Nerv. Syst. 62 (1997) 79-84) based on heart rate variability was used to assess cardiac sympathetic and parasympathetic functions separately. Both sympathetic and parasympathetic indices were increased during the appearance of Fm theta compared with control periods. Theta band activities in the frontal area were correlated negatively with sympathetic activation. The results suggest a close relationship between cardiac autonomic function and activity of medial frontal neural circuitry. PMID- 11275490 TI - Auditory memory in congenitally blind adults: a behavioral-electrophysiological investigation. AB - Blind people must rely more than sighted people on auditory input in order to acquire information about the world. The present study was designed to test the hypothesis that blind people have better memory than sighted individuals for auditory verbal material and specifically to determine whether memory encoding and/or retrieval are improved in blind adults. An incidental memory paradigm was employed in which 11 congenitally blind people and 11 matched sighted controls first listened to 80 sentences which ended either with a semantically appropriate or inappropriate word. Immediately following, the recognition phase occurred, in which all sentence terminal words were presented again randomly intermixed with the same number of new words. Participants indicated whether or not they had heard the word in the initial study phase. Event-related brain potentials (ERPs) were recorded from 28 electrode positions during both the encoding and the retrieval phase. Blind participants' memory performance was superior to that of sighted controls. In addition, during the recognition phase, previously presented words elicited ERPs with larger positive amplitudes than new words, particularly over the right hemisphere. During the study phase, words that would subsequently be recognized elicited a more pronounced late positive potential than words that were not subsequently recognized. These effects were reliable in the congenitally blind participants but could only be obtained in the subgroup of sighted participants who had the highest memory performance. These results imply that blind people encode auditory verbal material more efficiently than matched sighted controls and that this in turn allows them to recognize these items with a higher probability. PMID- 11275491 TI - Syntactic parsing preferences and their on-line revisions: a spatio-temporal analysis of event-related brain potentials. AB - The present study investigates the processes involved in the recovery from temporarily ambiguous garden-path sentences. Event-related brain potentials (ERP) were recorded while subjects read German subject-object ambiguous relative and complement clauses. As both clause types are initially analyzed as subject-first structures, object-first structures require a revision which is more difficult for complement than for relative clauses. The hypothesis is tested that the revision process consists of two sub-processes, namely diagnosis and actual reanalysis. Applying a spatio-temporal principal component analysis to the ERP data, distinct positive sub-components presumably reflecting different sub processes could be identified in the time range of the P300 and P600. It will be argued that the P600 is not a monolithic component, and that different sub processes may be involved at varying time points depending on the type of garden path sentence. PMID- 11275493 TI - Promoting rational drug policy. PMID- 11275494 TI - Harm reduction - a historical view from the left. AB - The harm reduction movement formed during a period in which social movements of the working class and the excluded were weak, neo-liberalism ideologically triumphant, and potential opposition movements were viewed as offering "tinkering" with the system rather than a total social alternative. This climate shaped and limited the perspectives, strategies, and tactics of harm reductionists almost everywhere. In many countries, this period was also marked by a "political economy of scapegoating" that often targeted drug users as the cause of social woes. This scapegoating took the form of "divide and rule" political initiatives by business and political leaderships to prevent social unrest in a long period of worldwide economic trends toward lowered profit rates and toward increasing income inequality. However, times have changed. Mass strikes and other labor struggles, opposition to the World Trade Organisation and other agencies of neo-liberalism, community-based protests against belt tightening, and other forms of social unrest have been increasing in many countries. This opens up the possibility of new allies for the harm reduction movement, but also poses difficult problems for which we need to develop answers. On-the-ground experience in alliance formation needs to be combined with careful discussion of and research about what approaches work to convince other movements to work for and with harm reduction, and which approaches do not. Class differences within the harm reduction movement are likely to become more salient in terms of (a) creating internal tensions, (b) increasingly, opening up new ways in which working class harm reductionists can organize within their own communities and workplaces, and (c) producing different strategic orientations that will need to be discussed and debated. As a movement, we will need to find ways to accommodate and discuss differing perspectives, needs, and assessments of opportunities and threats without paralyzing harm reduction activities. PMID- 11275492 TI - 3rd Congress of Pharmaceutical Sciences. CIFARP 2001. April 8-11, 2001, Aguas de Lindoia, Sao Paulo, Brazil. Abstracts. PMID- 11275495 TI - Harm reduction from a broader political-economic perspective. PMID- 11275496 TI - Capitalism, prohibitionism, and the drug policy reform movement: theses on Friedman et al. PMID- 11275497 TI - A view from outside. PMID- 11275498 TI - Notes from South America. PMID- 11275500 TI - The importance of clearly communicating the essence of harm reduction. PMID- 11275501 TI - Class, alliances and harm reduction: a personal view. PMID- 11275499 TI - Harm reduction alliances in an Argentinean context. PMID- 11275502 TI - Harm reduction - a historical view from the trenches. PMID- 11275503 TI - Point Defiance: a case study of the United States' first public needle exchange in Tacoma, Washington. AB - The first publicly funded needle exchange program in the United States began in Tacoma, Washington, in August 1988. The exchange's history is characterized by a series of firsts: the first American publicly funded exchange; the first pharmacy exchange; the first American needle delivery program; and the first state Supreme Court ruling not only supporting the existence of a needle exchange program but superseding existing drug paraphernalia laws. It is also unique because it began outside of the public health system and was not compromised by political feasibility. This article documents the events and personalities which led to the exchange's establishment and its expansion over time; the program's local, state and legal challenges and advocates; its portrayal in the local and national media; the research that documented its successes; and its important contribution to the fight for drug users' unencumbered access to sterile needles. PMID- 11275504 TI - A design for strife: alcopops, licit drug - familiar scare story. AB - In Scotland during the mid 1990s the news media began reporting growing concern about a new form of alcoholic beverage known as 'alcopops'. Fears raised by the press centred on the claim that these drinks were being marketed towards young children and as such were responsible for the rising levels of adolescent drunkenness. In focusing on underage consumption and alleged marketing aimed at children, the press portrayed alcopops use as illicit, allowing these drinks to be reported as if they were an illegal drug. This 'negative' publicity was in contrast to the 'positive' stories and advertising space given over to other alcoholic beverages elsewhere in their pages. This manuscript quantifies the volume of reporting of these drinks in the Scottish press over the life-span of their news-worthiness. It argues that the press's claims could not be supported by the realities of underage drinking at the time. When this became apparent the scare story ended as rapidly as it had begun. Though concerning a licit substance these findings were found to have many illuminating parallels with illegal drug scare stories. PMID- 11275505 TI - General practice or drug clinic for methadone maintenance? A controlled comparison of treatment outcomes. AB - The model for management of opiate dependence in the United Kingdom includes long term methadone maintenance. A consequence is either long waiting lists for treatment or that treatment capacity is expanded. General practitioners are encouraged to prescribe methadone for opiate dependent patients, but little is known about the differences between patients or outcomes in primary or secondary care settings. This paper compares patients' characteristics and treatment outcomes in a specialist drug clinic and a general practice operating a shared care policy (with the specialist clinic). We undertook a retrospective review of patient records. All patients prescribed methadone maintenance during a 2 year period in one general practice were compared with one in three patients treated at a drug clinic during the same period. Outcome was determined at the end of a treatment episode or on 30 June 1997 (whichever was sooner). Eighty-nine drug clinic and 36 general practice patients were followed up for a minimum of nine months each. Patient characteristics were similar at the start of treatment. A 'good' outcome (remaining in treatment or becoming drug free) was equally likely in either setting. Patients treated in the general practice setting were significantly more likely to be immunised (or have known natural immunity) against hepatitis B (adjusted odds ratio 6.0). Our findings suggest that with similar patient groups this model of care in general practice can produce results at least as good as those of a drug clinic. PMID- 11275506 TI - Survey of injectable methadone prescribing in general practice in England and Wales. AB - Little is known about the extent and conduct of general practice prescribing of injectable methadone to opiate users in England and Wales. A postal questionnaire survey of general practitioners (GPs) was conducted in 1999 to ascertain how many GPs were prescribing injectable methadone, to describe their prescribing practices and to explore their perceptions of the service they provided. Four hundred and seven GP practices in 77 out of 105 health authorities were apparently prescribing injectable methadone. The difficulties in identifying, and obtaining a response from, GPs prescribing injectable methadone are discussed. Analysis of 93 usable returned questionnaires showed a range of GP characteristics and experience. The GPs were treating 211 patients with injectable methadone and a further 2003 patients with oral methadone. Prescribing practice and monitoring arrangements did not always follow national guidelines. A minority of GPs had received training in the management of drug dependency. They were most likely to decide to prescribe injectable treatment on the recommendation of a specialist drug agency, and to discontinue prescribing if there was a suspicion of diversion of prescribed ampoules. Although significant numbers of GPs felt unsure about their skills and the support available to them, most appeared to be managing their patients thoughtfully and with appropriate outcomes in mind. Recent policy documents from the Department of Health and the Home Office have questioned the place of injectable methadone in the treatment of opiate misuse, particularly in a general practice setting. A Home Office licence will shortly be required to prescribe injectable methadone. This survey suggests a minority of GPs would apply for such a licence. More research into the effectiveness of injectable methadone treatment in a range of settings is needed before conclusions can be drawn about the appropriateness of providing this treatment in general practice. PMID- 11275507 TI - Opium past, opioid futures: imperialism, insurgency and pacification in a global commodity market. AB - This is a lightly edited version of Roger Lewis' notes for the speech he gave at the 11th International Conference on the Reduction of Drug Related Harm in Jersey in March 2000, shortly before his death. In this paper Roger Lewis argues for the need to take global and historical view of both drug markets and ways that nations try to control them. Drug markets have become globalised, influenced by changes in communication, finance, commodity and labour markets. Drug control activities are used to defend global strategic interests, amd foreign policy imperatives usually take precendence over drug policy. PMID- 11275508 TI - BRCA1 screening in a woman with breast cancer: a patient's perspective with commentary. AB - Genetic screening for breast cancer and other diseases poses major personal and psychological challenges for persons facing the prospect of testing. The personal report of those dilemmas by a woman considering BRCA1 screening who had had a lumpectomy for breast cancer and whose mother had died from that disease is presented here. Commentaries by a bioethicist and a genetics counselor show further complexities in the testing experience which persons considering testing and those providing them will need to confront. PMID- 11275509 TI - Abortion in adolescence: a four-country comparison. AB - The purpose of this study was to conduct a comparison, using qualitative analytic methodology, of perceptions concerning abortion among health care providers and administrators, along with politicians and anti-abortion activists (total n = 75) in Great Britain, Sweden, The Netherlands, and the United States. In none of these countries was there consensus about abortion prior to legalization, and, in all countries, public discussion continues to be present. In general, after legalization of abortion has no longer made it a volatile issue European countries have refocused their energy into providing family planning services, education, and more straightforward access to abortion compared with similar activities in the United States. PMID- 11275510 TI - Bringing emergency contraception to American women: the history and remaining challenges. AB - Emergency contraception has been called "America's best-kept secret." This article chronicles what it took to move it from secret to the pharmacy shelf. The fact that an emergency contraception product is available today in many pharmacies is indeed a major accomplishment. However, the job is not yet done. The shelf it needs to be found on is not just the pharmacists' shelf, behind the counter-but the shelf in the medicine cabinet in millions of homes everywhere, like burn medicine, "just in case." PMID- 11275511 TI - Improved use of contraceptives, attitudes toward pornography, and sexual harassment among female university students. AB - This study describes sexual behavior over a 10-year period in a female student population. The use of condoms at first coitus increased from 40% to 77%. Sexually transmitted diseases decreased from 26% to 14%, and abortions from 11% to 5.5%. One-fourth of students had had anal intercourse, and 86% had performed oral sex. Half of the women had read pornography. The majority of women with experience of oral sex graded it as positive, whereas they graded anal sex as mostly negative. Twelve percent of the women had been sexually harassed, mainly by their male peers (80%). PMID- 11275512 TI - Effects of SERMs on important indicators of cardiovascular health: lipoproteins, hemostatic factors, and endothelial function. PMID- 11275513 TI - Cigarette smoking in veteran women: the impact of job strain. AB - To evaluate the health effects of role overload, the relationship between multiple role (i.e., worker, spouse, caretaker) strain and current cigarette smoking was examined. A cross-sectional survey of women veterans, aged 36-85 years, was performed measuring home and job strain and health behaviors. Of the 275 women who rated both their work and home strains, 25% (n = 69) currently smoke cigarettes. Higher work strain, but not higher home strain, was associated with smoking adjusting for age, education, income, weight, and marital status. A stressful work environment may trigger persistent smoking and should be addressed during smoking cessation counseling. PMID- 11275514 TI - Mammography-related anxiety: effect of preprocedural patient education. AB - OBJECTIVES: To determine the effect of preprocedural education on mammography related anxiety. MATERIALS AND METHODS: A total of 613 women undergoing mammography were surveyed regarding anxiety about the procedure and expected results. Half the study population watched an educational videotape and half watched an entertaining movie in the waiting room. RESULTS: Anxiety levels about results were significantly higher than anxiety levels about the procedure (P <.001). There was no difference in procedural or cancer anxiety levels among women shown the educational tape and those shown the entertaining movie. CONCLUSION: The fear of discovering breast cancer generates most of mammography related anxiety. Preprocedural education did not affect procedural or cancer related anxiety. PMID- 11275515 TI - Partner violence: implications for health and community settings. AB - OBJECTIVE: To assist in the design and implementation of strategies to address partner violence, the objective of this study was to evaluate differences in mental health, health behaviors, and use of health care and specific community services between women who do or do not report experiences of partner violence as an adult. METHODS: During interviews with 392 women enrolled in a Medicaid managed care organization, measures of mental health status, health behaviors, use of health care and community services, and experiences of partner violence were collected. Using bivariate statistical analyses, characteristics between women reporting or not reporting partner violence were compared. Chi-square tests were used to assess significant differences between the groups. The relationships between outcomes of interest and violence were estimated with logistic regression models adjusting for significant demographic and health characteristics. RESULTS: Overall, 28% of women reported experiences of partner violence. Women reporting partner violence had twice the adjusted odds of depression and three times the adjusted odds of negative self-esteem compared with women not reporting experiences of partner violence. Women reporting partner violence, compared with those who did not, indicated higher use of specific types of health and community services such as mental health services [odds ratio (OR) 2.9; confidence interval (CI) 1.5-5.6] and individual counseling (OR 3.6; CI 2.2-6.1). CONCLUSIONS: A communitywide effort that establishes linkages between health care settings and community services may be important in addressing the needs of women who are experiencing partner violence. PMID- 11275516 TI - Improving the accuracy of identifying lesbians for telephone surveys about health. AB - Knowledge about the health status and health care needs of lesbians is limited by the lack of population-based studies, although recent survey methods research offers suggestions that may be relevant to involving lesbians in more rigorous studies. To explore the transferability of findings about the general population to research on lesbian health, focus groups were conducted in 1997-1998 with self identified lesbians in five U.S. urban areas. Videotaped telephone interviews stimulated discussion about methods for enhancing participation of lesbians in random digit dial telephone surveys. Results are useful for developing improved practices for conducting health surveys with lesbians. PMID- 11275517 TI - Planning a successful women's health and wellness conference in your community. AB - Planning a woman's health conference in your community requires knowledge of the audience's interests and the compilation of a diversified planning committee. PMID- 11275518 TI - Methods for inducing neuronal loss in preweanling rats using intracerebroventricular infusion of kainic acid. AB - Excitotoxins, such as kainic acid (KA), have been shown to produce neuronal degeneration in the adult rat brain. While preweanling rats have been shown to be relatively resistant to the neurotoxicity of lower doses of KA, the presence of neuronal loss at higher doses (of KA) has only begun to be investigated in such animals. A reliable method of producing neuronal loss in preweanling rats is to administer nmol concentrations of KA via intracerebroventricular (i.c.v.) injections on postnatal day 7 (P7). Using a three-dimensional, non-biased cell counting technique, we have shown that neuronal loss is observed in the CA3 subfield of the hippocampal formation at P45 and P75. Further, immunohistochemical studies of markers for cell death may be useful to examine the types of cellular processes associated with such neuronal loss. Data from our own experiments suggest the activation of immediate-early genes in the neuronal loss produced by KA administration at P7. This developmental animal model of neuronal loss may be useful in studying neurodevelopmental disorders where the onset of symptoms or cognitive deficits is thought to follow an early developmental insult. PMID- 11275519 TI - Animal models for the study of antidepressant activity. AB - Three behavioral paradigms are presented for the study of the mechanism of action of antidepressant treatments and for the screening of new antidepressant drugs. The first model (acute escape deficit) exploits the decreased ability of a rat exposed to an unavoidable stress to avoid a noxious stimulus, and it allows us to evaluate the preventive activity of a treatment on the development of escape deficit. The second paradigm (chronic escape deficit) begins as acute escape deficit, that is then indefinitely sustained by the repeated administration of mild stressors; this model allows us to evaluate the efficacy of a treatment to revert the escape deficit. The third is a model of anhedonia based on the finding that exposure to repeated unavoidable stress prevents the acquisition of an appetitive behavior induced and maintained by a highly palatable food (vanilla sugar) in rats fed ad libitum; this paradigm assesses the efficacy of a treatment to restore an animal's motivation. A long-term (2 to 3 week) treatment with classical antidepressants, such as imipramine or fluoxetine, resulted in a clear cut preventive and/or revertant activity in the three models. PMID- 11275520 TI - Synaptic interactions in neurones of the rat rostral ventrolateral medulla oblongata. AB - Increasing number of studies indicate that stimulation of peripheral nerves elicits complex postsynaptic responses in neurones of the ventral medulla oblongata. The present study describes a recommended protocol for intracellular recording of complex postsynaptic events in neurones of the ventral medulla oblongata. The aim was to provide surgical and experimental details that will enable successful recording of slow synaptic responses in vivo, in addition to the recording of fast evoked responses. The existence of slow inhibitory responses to stimulation of the cervical vagus and sciatic nerves have already been demonstrated together with the existence of convergent visceral and viscero somatic inputs to neurones here. The data collected in over 200 neurones so far indicated that neurones of the rostral ventrolateral medulla oblongata play an important and versatile role in the integration of somato-visceral sensory inputs. PMID- 11275521 TI - A modified roller method for organotypic brain cultures: free-floating slices of postnatal rat hippocampus. AB - We describe a novel procedure for organotypic cultivation of free-floating brain sections of postnatal rats with a modified roller technique. Three hundred to 350 microm-thick sections of hippocampus are cultured for 13-15 days at 35.5 degrees C in 10-15 ml of feeding medium in 50-100 ml bottles under constant rotation on a horizontal high-speed mini-roller (60 rpm). Histological analysis (paraffin sections, Nissl Cresyl Violet and Hematoxylin/Eosin staining) demonstrates good survival of neuronal and glial cells and complete preservation of the neuronal organization of cultivated hippocampus with minimal central necrosis. This novel protocol permits not only survival and development of long-term three-dimensional organotypic postnatal brain tissue but also allows simultaneous cultivation of any number of brain sections in one bottle (up to 50 and even more) and therefore is useful for high throughput study of neurocytotoxic and hypoxic/ischemic neuronal damage with subsequent histological, immunocytochemical, biochemical, and molecular analysis. PMID- 11275522 TI - In vivo microdialysis in the visual cortex of awake cat. I: surgery, animal training and sampling. AB - Sampling and monitoring release of excitatory and inhibitory amino acids in the striate cortex of mammals will provide important information for visual system research. A method allowing repeated microdialysis in the cortical layers of area 17 of the awake cat is described. Under visual control through a surgical microscope and using a stereotactic instrument, four probe guides are permanently implanted in area 17 of one hemisphere of the anesthetized animal and two fixation bars are mounted on the skull to allow fixation of the cat in a stereotactic frame. The implantation of four probe guides in the same hemisphere allows simultaneous sampling from different cortical regions subserving different parts of the visual field. A removable transparent cover protects the probe guides. After recovery from surgery the awake cats are trained to adapt to a fixation in a stereotaxic apparatus. Once adapted to that situation, the cats are used for 5 h in vivo microdialysis experiments without anesthesia. PMID- 11275523 TI - In vivo microdialysis in the visual cortex of awake cat. II: sample analysis by microbore HPLC-electrochemical detection and capillary electrophoresis-laser induced fluorescence detection. AB - Sampling and monitoring release of excitatory and inhibitory amino acids in the striate cortex of mammals will provide important information for visual system research. Two microbore high performance liquid chromatography-electrochemical detection methods and a capillary electrophoresis-laser induced fluorescence detection were developed to determine the inhibitory amino acid, gamma aminobutyric acid and the excitatory amino acids, glutamate and aspartate in microdialysates of cat striate cortex. In the liquid chromatography method, samples were derivatized using OPA-TBT. Ten microliters of derivatized product was injected onto the microbore column (100 x 1 mm i.d., C8) for quantitative analysis. Electrochemical detection was employed. In the capillary electrophoresis method, samples were derivatized using fluorescein isothiocyanate and separated in borate buffer within 15 min, then detected by a laser-induced fluorescence detector. PMID- 11275525 TI - A single motor unit recording technique for studying the differential activation of corticospinal volleys by transcranial magnetic stimulation. AB - The purpose of this method is to establish a single motor unit recording technique to study the differential activation of corticospinal volleys by various types of transcranial magnetic stimulation (TMS). TMS is performed with various coil orientations over the hand or leg motor areas and surface EMG, and single motor unit recordings are made either from the studied hand or leg muscle. Transcranial electrical stimulation (TES) is also performed over the motor cortex as well as at the foramen magnum level to determine the latency of D waves. The intensity of stimulation is set just above the motor threshold for each type of stimulation. This method makes it possible to activate some I volleys (especially I1 and I3 waves) preferentially, if not selectively, from the hand and leg motor areas. The obtained results accord well with recent epidural recording studies, which lends support to the validity of this method. PMID- 11275524 TI - In vivo microdialysis in the visual cortex of awake cat. III: histological verification. AB - In vivo microdialysis sampling extracellular excitatory and inhibitory amino acids from the striate cortex of mammals will provide important information for visual system research. To facilitate the interpretation of microdialysis results, this protocol critically examines: (1) the location of probe implantation in the visual cortex using Nissl staining; (2) the morphological changes after probe implantation by visualization of neurons containing glutamate; (3) the morphological changes after probe implantation by visualization of gliosis using glial fibrillary acidic protein (GFAP) immunocytochemistry; (4) the implantation of the probe in sensory-deprived versus non-deprived cortical regions by visualization of neurons containing c-Fos protein after limited retinal lesion. The histochemical and immunocytochemical methods of Glu, GFAP and c-Fos used are described. PMID- 11275526 TI - An improved cell isolation technique for studying intracellular Ca(2+) homeostasis in neurones of the cochlear nucleus. AB - Neurones isolated from various parts of the brain are used extensively for electrophysiological and immuncytochemical studies, as well as to investigate their Ca(2+) homeostasis. In this work we report on an isolation technique that yielded neurones suitable for functional studies targeting the investigation of their Ca(2+) handling mechanisms. The cell isolation involved enzymatic dissociation with combined collagenase/pronase treatment and gentle mechanical trituration. At the end of the isolation the cells were incubated in a cell culture incubator (CO2 concentration = 5.1%) at 37 degrees C in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% heat-inactivated horse serum. The vitality of the isolated cells was indicated by their low intracellular Ca(2+) concentrations (17.2 +/- 0.5 nM; n = 38) and by their ability to produce large Ca(2+) transients on depolarization. These Ca(2+) transients were rapidly terminated and the resting intracellular Ca(2+) concentration was quickly restored proving that isolation did not compromise the Ca(2+) homeostatic mechanisms of the nerve cells. The technique allowed reliable, long (45-60 min) and reproducible measurements of Ca(2+) currents on these neurones as well as the recording of their intracellular Ca(2+) concentration. Our results indicate that incubation in DMEM with horse serum markedly increases the number of surviving neurones after the enzyme treatment, and their Ca(2+) homeostasis can be studied for significantly longer periods of time. PMID- 11275527 TI - Progressive and reproducible focal cortical ischemia with or without late spontaneous reperfusion generated by a ring-shaped, laser-driven photothrombotic lesion in rats. AB - Clinical stroke is mostly of thromboembolic origin, in which the magnitude of brain damage resulting from arterial occlusions depends on the degree and duration of the concomitant ischemia. To facilitate more controllable and reproducible study of stroke-related pathophysiological mechanisms, a photothrombotic ring stroke model was initially developed in adult rats. The ring interior zone comprises an anatomically well confined cortical region-at-risk which is gradually encroached by progressive hypoperfusion, thus mimicking the situation (albeit in inverse fashion) of an ischemic penumbra or stroke-in evolution. Modification of this model using a thinner ring irradiation beam resulted in late spontaneous reperfusion in the cortical region-at-risk and a remarkable morphological tissue recovery in this ostensibly critically injured region. On the other hand, doubling the thin irradiating beam intensity facilitates a complementary situation in which lack of reperfusion in the region at-risk after stroke induction leads to tissue pannecrosis. The dual photothrombotic ring stroke model, effectuated either with or without reperfusion and thereby tissue recovery or pannecrosis, may be well suited for the study of events related to postischemic survival or cell death in the penumbra region. To popularize the photothrombotic ring stroke model, we present a detailed protocol of how this model is induced in either version as well as protocols for transcardial carbon black perfusion and laser-Doppler flowmetry experiments. PMID- 11275528 TI - Induced electrocorticographic gamma activity during auditory perception. Brazier Award-winning article, 2001. AB - OBJECTIVE: To define the spatial, temporal, and functional characteristics of induced gamma (>30 Hz) activity during functional activation of the left superior temporal gyrus. METHODS: Electrocorticographic (ECoG) recordings were made in 4 clinical subjects during auditory tone and phoneme discrimination tasks, and event-related changes in the ECoG band power were calculated. The topography and temporal sequence of event-related power changes in different gamma bands were contrasted with those of auditory evoked potentials (AEPs), and with those of event-related power changes in the alpha band (8-12 Hz). RESULTS: Auditory stimuli induced a broadband power augmentation that included 40 Hz, as well as higher (80-100 Hz) gamma frequencies. The topography of gamma augmentation was similar, but not identical, to that of the AEP, and was more focused than that of alpha power suppression. Its temporal onset coincided with the N100, but outlasted it. Phonemes produced greater gamma augmentation than tones, while a similar difference was not observed in the N100. CONCLUSIONS: Auditory perception induces ECoG gamma activity not only at 40 Hz, but also in higher gamma frequencies. This activity appears to be an index of cortical activation that reflects task-specific processing in the human auditory cortex more closely than the AEP or alpha power suppression. PMID- 11275529 TI - A convenient method to reduce crosstalk in surface EMG. Cobb Award-winning article, 2001. AB - OBJECTIVES: Crosstalk in surface EMG can be reduced by the use of spatial filters. We compared a variety of spatial filters to establish the most effective and the least complex method to reduce crosstalk. METHODS: Six different spatial filters described in the literature were tested in 8 healthy volunteers. Electrode arrays were placed over the anterior tibial and triceps surae muscles. Selective muscle activation was achieved both by supramaximal nerve stimulation and by maximal voluntary contraction. Selectivity of activation was guaranteed by using intramuscular wire electrodes. Crosstalk was quantified by dividing the amount of EMG activity recorded during pure agonist activation (i.e. the muscle directly under the electrode array) by the EMG activity recorded during pure antagonist activation. This was done for both compound muscle action potentials and voluntary muscle activation. The amount of crosstalk recorded with the different spatial filters was compared with that recorded with a standard bipolar lead. RESULTS: Crosstalk was most reduced by the "double-differential" (DD) filter, yielding an up to 6-fold improvement of EMG selectivity. We then compared signals recorded with this DD filter with those recorded with the less complex "branched electrode". As expected on theoretical grounds, signals from both filter types were identical. CONCLUSIONS: Crosstalk is best reduced using a "double-differentiating" recording technique, which can be achieved easily using a branched electrode instead of a standard bipolar lead. This technique can be used with all conventional EMG equipment. PMID- 11275530 TI - Optimization of facilitation related to threshold in transcranial magnetic stimulation. AB - OBJECTIVES: To attain the standardized procedure for optimal facilitation, we analyzed motor-evoked potential (MEP) responses to the degree of voluntary contraction and stimulus intensity. METHODS: Fifteen normal subjects were included. MEPs were elicited at thenar muscles during rest and at gradual voluntary contraction (MVC), using 10, 30, and 50% of MVC. During rest and each contraction, the excitability threshold at rest (RET) and at contraction (CET) were determined. Consecutive stimuli were applied, with the intensity of ratio increments (110-150% of ET). RESULTS: The RET showed a remarkable decrease after contraction. Shortening of latency reached a saturation level at 10% of MVC. Amplitude reached a saturation level at 30% of MVC with 62.7+/-8.5% of the maximum output, which is equal to 140% intensity of CET, and 110% of RET. The MEP amplitudes at rest and at 10% MVC were influenced by their ETs, but those measured above 30% of MVC were not related. CONCLUSIONS: The procedure recommended for optimal facilitation is as follows: to achieve minimal latency of MEPs, a minimal contraction (10% of MVC) with RET intensity is sufficient and for maximal amplitude, a moderate contraction (30% of MVC) with 110% of RET intensity is adequate. PMID- 11275531 TI - Impairment of motor cortex activation and deactivation in Parkinson's disease. AB - OBJECTIVES: To examine the time course of corticospinal excitability before and after voluntary movement in Parkinson's disease (PD). METHODS: We studied 9 mild PD patients at least 12 h off medications and 11 healthy volunteers in a simple reaction time (RT) paradigm. Suprathreshold transcranial magnetic stimulation was delivered to the left motor cortex at intervals covering the periods before and after movement. RESULTS: PD patients (284+/-90 ms) and normal subjects (282+/-56 ms) had similar median RT. The time courses of both the premovement increase and the postmovement decrease in corticospinal excitability were significantly different between PD patients and normal subjects. The increase in motor-evoked potential (MEP) amplitudes began earlier for PD patients (200 ms before electromyographic (EMG) onset) than for normal subjects (150 ms before EMG onset), but the rate of increase was slower in PD patients than controls. After EMG offset, MEP amplitudes were increased for about 150 ms in normal subjects, but in PD patients this period was prolonged to about 350 ms. CONCLUSIONS: Impairment of motor cortex activation and deactivation is an early feature of PD and may be a physiological correlate of bradykinesia. The normal RT in our patients may be related to the earlier occurrence of the premovement increase in corticospinal excitability compensating for the slower rate of rise. PMID- 11275532 TI - A medial to lateral shift in pre-movement cortical activity in hemi-Parkinson's disease. AB - OBJECTIVE: Recent evidence suggests that cortical activity associated with voluntary movement is relatively shifted from medial to lateral premotor areas in Parkinson's disease. This shift occurs bilaterally even for unilateral responses. It is not clear whether the shift in processing reflects an overall change in movement strategy, thereby involving alternate cortical areas, or reflects a compensatory change whereby, given the appropriate conditions, less impaired cortical areas are able to provide a similar function in compensation for those areas which are more impaired. This issue was examined in patients with hemi Parkinson's disease, in whom basal ganglia impairment is most pronounced in one hemisphere. METHODS: Fourteen patients with hemi-Parkinson's disease and 15 age matched control subjects performed a Go/NoGo finger movement task and the contingent negative variation (CNV) was recorded from 21 scalp positions. RESULTS AND CONCLUSIONS: Maximal CNV amplitudes were found over central medial regions for control subjects, but were shifted more frontally for Parkinson's disease patients, reduced in amplitude over the midline and lateralized towards the side ipsilateral to the greatest basal ganglia impairment. This shift in cortical activity from medial to lateral areas in Parkinson's disease patients appears to reflect a compensatory mechanism operating predominantly on the side of greatest basal ganglia impairment. PMID- 11275533 TI - Cerebellar stimulation in acute cerebellar ataxia. AB - OBJECTIVES: To report follow-up studies of cerebellar stimulation in patients with acute cerebellar ataxia (ACA). METHODS: We studied two patients with ACA. One patient also had decreased deep sensations in the feet due to combined diseases such as diabetic polyneuropathy and lumbosacral radiculopathies. We applied the technique of electrical stimulation over the cerebellum which was reported previously (Ugawa et al., J Physiol 441 (1991a) 57). RESULTS: Conditioning stimulation over the cerebellum did not reduce the size of motor evoked potentials to test magnetic stimulation of the motor cortex at conditioning-test intervals of 5, 6, and 7 ms in the acute stage in both patients. However, normal suppression was recognized in the recovery stage in both patients. CONCLUSIONS: This technique was useful for follow-up evaluation of cerebellar function in patients with ACA and was also useful for distinguishing cerebellar ataxia from sensory ataxia in a patient with combined diseases. PMID- 11275534 TI - Interhemispheric interaction between the hand motor areas in patients with cortical myoclonus. AB - OBJECTIVE: To study interhemispheric interaction between the hand motor areas of both hemispheres through the corpus callosum in myoclonus epilepsy. SUBJECTS: Five patients with benign myoclonus epilepsy and ten age matched normal volunteers. METHODS: We studied effects of a medially directed conditioning stimulus over the right hand motor area on responses in the right first dorsal interosseous muscle to a posteriorly directed test stimulus over the left hand motor area. RESULTS: In normal subjects, inhibition was evoked at interstimulus intervals (ISIs) of 8-20ms (late inhibition). In contrast, facilitation occurred in patients at ISIs of 4-6ms (early facilitation) with no late inhibition. CONCLUSIONS: The lack of late inhibition in the patients is consistent with the idea that cortical inhibitory interneurones are affected in myoclonus epilepsy. We propose that this releases interhemispheric facilitation from powerful surround inhibition. The consequence is a predominant early facilitation between the hemispheres in patients with myoclonus epilepsy. PMID- 11275536 TI - Remote F-wave changes after local botulinum toxin application. AB - OBJECTIVE: Although the therapeutic effects of botulinum toxin A can be explained by its action at the neuromuscular junction, central or more proximal effects have also been discussed. METHODS: Eleven patients with torticollis spasmodicus and 3 patients with writer's cramp were studied before and 1 and 5 weeks after the first treatment with botulinum toxin. We measured compound muscle action potentials (CMAPs), motor conduction velocities (MCVs), the shortest (SFL) and the mean F-wave latencies (MFL) and F-wave persistence (30 trials) of untreated muscles for each side (ulnar nerve-abductor digiti minimi muscle, peroneal nerve tibialis anterior muscle). RESULTS: CMAPs and MCVs showed no significant changes. For both nerves, however, SFL and MFL were prolonged slightly 1 week after treatment and returned to about baseline after 5 weeks (t test). The F-wave persistence was reduced 1 week after treatment for the right ulnar and both peroneal nerves (t test). CONCLUSIONS: These results are not likely due to an impairment of neuromuscular transmission. Instead, we propose a decreased excitability of alpha-motoneurons supplying non-treated muscles. A reduction of muscle spindle activity or changes of the recurrent inhibition are discussed as possible causes. PMID- 11275535 TI - Assessment of reorganization in the sensorimotor cortex after upper limb amputation. AB - OBJECTIVE: We wanted to investigate plastic changes occurring in the motor and somatosensory cortex after upper limb amputation, and their possible relationship to phantom pain. METHOD: To assess these plastic changes, we used transcranial magnetic stimulation (TMS) and source localization of somatosensory evoked potentials (SEP). Eleven patients with upper limb amputation were investigated. The phantom pain intensity was assessed by visual analogue scaling (VAS). RESULTS: Using TMS mapping, we found a significant lateralization of the amplitude-weighted centre of gravity (P<0.01) and an enlargement of the excitable area (P<0.05) on the hemisphere contralateral to the amputation. SEP mapping showed a significant medialization of the N20 dipole (P<0.05) on this side. None of these changes correlated with the phantom pain intensity. CONCLUSIONS: We conclude that after limb amputation, the relationship between plastic changes occurring in the sensorimotor cortex and phantom pain seems to be more complex than previously believed. PMID- 11275537 TI - Reflex receptive fields for human withdrawal reflexes elicited by non-painful and painful electrical stimulation of the foot sole. AB - OBJECTIVES: Human withdrawal reflex receptive fields (RRFs) were assessed for 4 different electrical stimulus intensities, ranging from below the pain threshold (PTh) to up to two times the PTh intensity (0.8x, 1.2x, 1.6x, and 2.0xPTh). METHODS: Thirteen subjects participated, and the reflexes were recorded in a sitting position. The stimuli were delivered in random order to 12 positions distributed over the foot sole. Tibialis anterior (TA), gastrocnemius medialis (GM), vastus lateralis (VL), and biceps femoris (BF) reflexes were recorded. Further, knee and ankle joint angle changes were recorded. RESULTS: The strongest reflexes were seen in the TA compared with the other 3 muscles. Dorsi-flexion dominated distal to the talocrural joint corresponding to the TA receptive field area. An expansion of the RRF for the TA and GM was seen when increasing the stimulus intensity from 0.8xPTh to 1.2xPTh and from 1.2xPTh to 1.6xPTh, indicating a gradually increasing reflex threshold towards the border, where TA contraction is inappropriate in a withdrawal reaction. For the BF and VL, the borders of the RRF areas were not detected. By integrating the reflex size within the RRF (i.e. the reflex volume), gradually increasing reflexes for increasing stimulus intensity were seen in all 4 muscles tested, most clearly in the TA and GM. The subjective pain intensity correlated to the reflex volume for the TA, GM, and BF. CONCLUSIONS: In conclusion, the highest reflex sensitivity was seen in the centre of the RRF, while the stimulus intensity needed for eliciting a reflex increased towards the receptive field border. Within the RRF, stronger reflexes were evoked for increasing stimulus intensity. The limit in the size of the receptive field size for the TA and GM supports a modular withdrawal reflex organisation. PMID- 11275538 TI - Intracerebral event-related potentials to subthreshold target stimuli. AB - OBJECTIVES: Event-related potentials (ERPs) elicited by subthreshold visual stimuli were recorded directly from human frontal and temporal lobe structures to study unconscious perception. METHODS: Thirteen intractable epileptic patients undergoing depth electrode recordings prior to their surgical treatment participated in the study. An original method of modified visual oddball paradigm with supraliminal and subliminal stimuli was applied, and the averaged responses to both kinds of stimuli were subsequently compared. RESULTS: The results clearly prove that, at least from an electrophysiological viewpoint, the mechanism of unaware processing of visual stimuli in the human brain does not differ substantially from the aware processing. Finding the subliminal P3 waveform in a number of cortical structures (hippocampus and parahippocampal gyrus bilaterally, and left-sided mesiofrontal, orbitofrontal and lateral temporal cortex) indicates their involvement in unconscious processing, in spite of the fact that typical large-scale neurocognitive networks are not completely activated. The absence of activation consistently observed bilaterally in dorsolateral prefrontal cortices, in connection with right-sided cortical frontal lobe structures and right-sided lateral temporal neocortex in unconscious perception, supports the importance of these structures for the awareness of visual stimuli. The proof of the significantly faster unaware information processing represents another distinctive feature of implicit visual perception. CONCLUSIONS: Based on the presented findings and comparisons with the results of previous ERP, functional magnetic resonance imaging, positron emission tomography, and clinical neuropsychological studies, a crucial role of the large-scale neural system for conscious experience of perception is suggested, which is distributed extensively among the dorsal posterior association areas and the prefrontal cortex, with the dominant part being that of the right hemisphere. PMID- 11275540 TI - The heartbeat-evoked brain potential in patients suffering from diabetic neuropathy and in healthy control persons. AB - OBJECTIVES: Neurotransmission from the heart to the brain results in a heartbeat evoked potential (HEP). This potential appears as a positive waveform ranging from 250 to 600 ms after the onset of ventricular contraction. Only limited information exists as to what extent the HEP is sensitive to a dysfunction in cardio-afferent pathways. Thus, the HEP was studied in patients with autonomic diabetic neuropathy. METHODS: Twenty-five patients and a healthy control group of equal size participated. The HEP was obtained as the average over 1200 EEG sweeps (18 channels) sampled contingent upon the onset of ventricular contraction. A heartbeat attention task and a distraction task were employed. Patients answered a questionnaire pertaining to the frequency of subjective symptoms related to diabetic neuropathy. RESULTS: The HEP amplitude at frontal, central and temporal locations was significantly diminished in patients in the latency range of 280 330 ms. A significant correlation was found between the questionnaire score of subjective autonomic symptoms and the reduction in the HEP. CONCLUSIONS: We conclude that the HEP is sensitive to a comparably moderate abnormality in nerve function. Furthermore, we assume that the processing of subjective symptoms of the disease and the generation of the HEP share some common neuronal pathways. PMID- 11275539 TI - Impairment of an event-related potential correlate of memory in schizophrenia: effects of immediate and delayed word repetition. AB - OBJECTIVE: We investigated the nature of the memory impairment in schizophrenia using an event-related potential (ERP). METHODS: Visual ERPs were recorded while 20 schizophrenics and 20 controls performed semantic categorization tasks with incidental word repetitions. Participants responded to occasional target words. Half of the non-target words were repeated immediately after initial presentation (lag 0) or after 5 intervening words (lag 5). RESULTS: In both groups, ERPs to words at lag 0 were more positive than those to non-repeated words, though this positive-going effect was attenuated in the schizophrenics, especially around 400 500 ms. The effect at lag 5 was smaller and shorter than that at lag 0 but was comparable between groups. Attenuation of the N400 peak occurred for word repetition at lag 0 in controls but not in schizophrenics, whereas a peak increment in the late positive component induced by word repetition at both lags was observed in both groups. CONCLUSIONS: Findings indicate that patients with schizophrenia have a deficit in a brain process modulating ERP correlates of memory, when words are repeated immediately. This deficit might be related to an abnormal N400 priming effect in schizophrenia. PMID- 11275541 TI - Melatonin as a sleep inductor for electroencephalogram recordings in children. AB - OBJECTIVE: To determine the efficacy of melatonin in obtaining sleep electroencephalograms in children. METHODS: Melatonin was used in 68 unselected children referred to the neurophysiology department for sleep electroencephalogram (EEG). A group of 68 children matched for age and sex who underwent EEGs after sleep deprivation in the same period was used as control. Sleep latency, as well as latency from stage 1 to stages 2, 3 and 4 were measured. RESULTS: No difference in the number of children who went to sleep was seen. No significant difference is the macrostructure of sleep was seen, other than a reduced sleep latency for the melatonin group (P<0.01). CONCLUSION: The study suggests that melatonin can reliably be used for obtaining sleep EEGs in children. Its use seems to provide a good alternative to pharmacological sedation and a complementary method to sleep deprivation. PMID- 11275542 TI - Psychomotor EEG variant of Gibbs: an association with underlying structural pathology. AB - OBJECTIVE: To describe the association between a unilateral mid-temporal rhythmic theta discharge ("psychomotor variant of Gibbs") with neuroimaging-demonstrated underlying mass lesion. METHODS: Standard routine awake and scalp electroencephalography, continuous video-EEG monitoring and magnetic resonance brain imaging were employed in the diagnostic work-up of a 9-year-old boy with a severe behavioural disturbance and episodic outbursts of aggression. RESULTS: EEG showed a unilateral mid-temporal rhythmic discharge which was continuous in drowsiness and which remained confined to the right hemisphere. MRI showed a lesion in the temporal horn of the right lateral ventricle displacing superiorly the white matter stem of the right temporal lobe. CONCLUSION: A rhythmic mid temporal theta discharge, commonly regarded as a benign EEG variant, may, in some patients, reflect underlying structural pathology. Neuroimaging should be considered particularly when this EEG pattern remains confined to one hemisphere. PMID- 11275543 TI - Space-oriented segmentation and 3-dimensional source reconstruction of ictal EEG patterns. AB - OBJECTIVES: Characterization of the EEG pattern during the early phase of a seizure is crucial for identifying the epileptic focus. The purpose of the present investigation was to evaluate a method that divides ictal EEG activity into segments of relatively constant surface voltage distribution, and to provide a 3-dimensional localization of the activity during the different segments. METHODS: For each timepoint the electrical voltage distribution on the scalp (the voltage map) was determined from the digitized EEG recording. Through a spatial cluster analysis time sequences where the maps did not change much (segments) were identified, and a 3-dimensional source reconstruction of the activity corresponding to the different mean maps was performed using a distributed linear inverse solution algorithm. RESULTS: Segments dominating early in seizure development were identified, and source reconstruction of the EEG activity corresponding to the maps of these segments yielded results which were consistent with the results from invasive recordings. In some cases a sequence of consecutive segments was obtained, which might reflect ictal propagation. CONCLUSIONS: Segmentation of ictal EEG with subsequent 3-dimensional source reconstruction is a useful method to non-invasively determine the initiation and perhaps also the spread of epileptiform activity in patients with epileptic seizures. PMID- 11275544 TI - Exploratory data analysis of evoked response single trials based on minimal spanning tree. AB - OBJECTIVE: An exploratory data analysis framework, based on minimal spanning tree, is proposed as a means to support the analysis of single trial (ST) electrophysiological signals. The core of this framework is the compact description of the input ST sample in a form of content-dependent ordered lists. Based on the established hierarchies, efficient ways to increase the SNR, extract prototypical responses, visualize possible self-organization trends in the sample and track the course of evoked response along the trial-to-trial dimension, are proposed. METHOD: Magnetoencephalographic auditory evoked responses were used for demonstrating and validating the introduced framework. RESULTS AND CONCLUSION: The results demonstrate the benefits, from this intelligent manipulation of STs, in understanding and enhancing the actual evoked signal. Specifically we find support for stimulus-induced phase-resetting hypothesis in the 3-20 Hz band, the existence of trials void of the prototypical evoked response, and an order across the single trial set hinting at an underlying process with long time scale. PMID- 11275545 TI - The five percent electrode system for high-resolution EEG and ERP measurements. AB - OBJECTIVE: A system for electrode placement is described. It is designed for studies on topography and source analysis of spontaneous and evoked EEG activity. METHOD: The proposed system is based on the extended International 10-20 system which contains 74 electrodes, and extends this system up to 345 electrode locations. RESULTS: The positioning and nomenclature of the electrode system is described, and a subset of locations is proposed as especially useful for modern EEG/ERP systems, often having 128 channels available. CONCLUSION: Similar to the extension of the 10-20 system to the 10-10 system ("10% system"), proposed in 1985, the goal of this new extension to a 10-5 system is to further promote standardization in high-resolution EEG studies. PMID- 11275546 TI - Participation of histones and histone-modifying enzymes in cell functions through alterations in chromatin structure. AB - Alterations in the chromatin structure are preferentially involved in the regulation of cell functions, including gene expression, in eukaryotes. Three types of mechanisms, by which the alterations are caused have been reported: (i) variants of histone subtypes, (ii) chromatin remodeling, and (iii) post translational modification. This review focuses mainly on the first and third mechanisms, especially on the acetylation of core histones, one of the third mechanisms. Using the gene targeting technique for the DT40 chicken B cell line, we systematically generated a number of mutants, respectively, devoid of particular genes encoding histones and histone deacetylase(s) (HDACs). Most of the H1 and core histone variants should be involved positively or negatively in the transcription regulation of particular genes. Of the chicken HDACs (chHDACs), chHDAC-2 controls the amount of the IgM H-chain at the steps of both transcription and alternative pre-mRNA processing, and chHDAC-3 is essential for cell viability, whereas chHDAC-1 merely affects gene expression in DT40 cells. These results indicate that HDAC family members should participate, in combination with one another, and/or histone acetyltransferase(s) (HATs), in the acetylation of core histones that regulates gene expression through alterations in the chromatin structure. PMID- 11275547 TI - Functions of RecQ family helicases: possible involvement of Bloom's and Werner's syndrome gene products in guarding genome integrity during DNA replication. AB - Escherichia coli RecQ helicase is a component of the RecF pathway of recombination whose components are required to reassemble a replisome complex at the site of the replication fork after the removal of a lesion. There are at least five RecQ homologues in human cells, including BLM and WRN. The genes encoding BLM and WRN are mutated in the cancer-prone disorder Bloom's syndrome (BS) and the plogeroid disorder Werner's syndrome (WS), respectively. These syndromes are characterized by a high degree of genomic instability, including chromosomal breaks, multiple large deletions, and translocations, and cells derived from BS and WS patients show defects in DNA replication. Recently, it has become clear that a Holliday junction-like structure is formed at stalled replication forks to result in the formation of double-stranded breaks, and recombination plays an important role in the repair of stalled or broken replication forks, leading to the reinitiation of replication. Defects in the processing of stalled replication forks could lead to aberrant recombination events resulting in genetic instability. Recent studies on BLM, WRN, and the RecQ homologue of Saccharomyces cerevisiae, Sgs1, indicate that these RecQ homologues interact with proteins involved in DNA replication, and function in a pathway from the DNA replication check point to homologous recombination. PMID- 11275548 TI - The mouse putative pheromone receptor was specifically activated by stimulation with male mouse urine. AB - To detect the biological activity of mammalian putative pheromone receptors (V1Rs and V2Rs), the mouse V1R gene was introduced into a primary culture of vomeronasal cells using the adenovirus expression system, and the response of these cells to mouse urine was analyzed by calcium imaging. These cells specifically responded to male but not female mouse urine. This response was attenuated by pertussis toxin, a specific inhibitor of G-protein G(ialpha)/G(oalpha) coupling from receptors. Our findings indicate that a putative pheromone receptor was specifically activated by mouse urine, a major source of mouse pheromones, and suggest that G(i)/G(o) are functionally coupled with the receptor. PMID- 11275549 TI - Characterization of the flavoprotein domain of gp91phox which has NADPH diaphorase activity. AB - A series of truncated forms of gp91phox were expressed in Escherichia coli in which the N-terminal hydrophobic transmembrane region was replaced with a portion of the highly soluble bacterial protein thioredoxin. TRX-gp91phox (306-569), which contains the putative FAD and NADPH binding sites, showed weak NADPH dependent NBT (nitroblue tetrazolium) reductase activity, whereas TRX-gp91phox (304-423) and TRX-gp91phox (424-569) were inactive. Activity saturated at about a 1:1 molar ratio of FAD to TRX-gp91phox (306-569), and showed the same K(m) for NADPH as that for superoxide generating activity by the intact enzyme. Activity was not inhibited by superoxide dismutase, indicating that it was not mediated by superoxide, but was blocked by an inhibitor of the respiratory burst oxidase, diphenylene iodonium. In the presence of Rac1, the cytosolic regulatory protein p67phox stimulated the NBT reductase activity, but p47phox had no effect. Truncated p67phox containing the activation domain (residues 199-210) [C.-H. Han, J.R. Freeman, T. Lee, S.A. Motalebi, and J.D. Lambeth (1998) J. Biol. Chem. 273, 16663-16668] stimulated activity approximately 2-fold, whereas forms mutated or lacking this region failed to stimulate the activity. Our data indicate that: (i) TRX-gp91phox (306-569) contains binding sites for both pyridine and flavin nucleotides; (ii) this flavoprotein domain shows weak diaphorase activity; and (iii) the flavin-binding domain of gp91phox is the target of regulation by the activation domain of p67phox. PMID- 11275550 TI - Apoptosis of CTLL-2 cells induced by an immunosuppressant, ISP-I, is caspase-3 like protease-independent. AB - In our previous study, the sphingosine-like immunosuppressant ISP-1 was shown to induce apoptosis in the mouse cytotoxic T cell line CTLL-2. In this study, we characterized the ISP-1-induced apoptotic pathway. Although caspase-3-like protease activity increases concomitantly with ISP-1-induced apoptosis in CTLL-2 cells, the apoptosis is not inhibited by caspase-3-like protease inhibitors, i.e. DEVD-cho and z-DEVD-fmk. In contrast, sphingosine-induced apoptosis in CTLL-2 cells is caspase-3-like protease-dependent. A caspase inhibitor with broad specificity, z-VAD-fmk, protects cells from apoptosis induced by ISP-1, indicating that ISP-1-induced apoptosis is dependent on caspase(s) other than caspase-3. Overexpression of Bcl-2 or Bcl-xL suppresses the apoptosis induced by ISP-1, although sphingosine-induced apoptosis is not efficiently inhibited by Bcl 2. Finally, ISP-1-induced mitochondrial depolarization, which is thought to be a checkpoint dividing the apoptotic pathway into upstream and downstream stages, is not inhibited by DEVD-cho, but is inhibited by z-VAD-fmk. These data suggest that a pathway dependent on caspase(s) other than caspase-3 is involved upstream of mitochondrial depolarization in ISP-1-induced apoptosis. PMID- 11275551 TI - Identification of functionally important cysteine residues of the human renin binding protein as the enzyme N-acetyl-D-glucosamine 2-epimerase. AB - Renin-binding protein (RnBP) is an endogenous renin inhibitor originally isolated from porcine kidney. It was recently identified as the enzyme N-acetyl-D glucosamine (GlcNAc) 2-epimerase [Takahashi, S. et al. (1999) J. Biochem. 125, 348-353] and its active site residue was determined to be cysteine 380 by site directed mutagenesis [Takahashi, S. et al. (1999) J. Biochem. 126, 639-642]. To further investigate the relationship between structure and function of recombinant human (rh) RnBP as a GlcNAc 2-epimerase, we have constructed several C-terminal deletion and multi-cysteine/serine mutants of rhGlcNAc 2-epimerase and expressed them in Escherichia coli cells. The expression was detected by Western blotting using anti-rhRnBP antiserum. The C-terminal deletion mutant, Delta400 417, had approximately 50% activity relative to the wild-type enzyme, but other C terminal deletion mutants, Delta380-417, Delta386-417, and Delta390-417, had no enzymatic activity. Mutational analysis of multi-cysteine/serine mutants revealed that cysteines 41 and 390 were critical for the activity or stabilization of the enzyme, while cysteine residues in the middle of the enzyme, cysteines 125, 210, 239, and 302, had no essential function in relation to the activity. PMID- 11275552 TI - Isolation and characterization of an N-linked oligosaccharide that is significantly increased in sera from patients with non-small cell lung cancer. AB - The structures of N-linked oligosaccharides present in human sera from 12 healthy volunteers and from 14 patients with non-small cell lung cancer (NSCLC) were analyzed by our recently developed partially automated systematic method. Thirty different structures of oligosaccharides were deduced, and these accounted for 84.1% of the total N-linked oligosaccharides present in human sera. All of the quantified oligosaccharide levels in healthy human sera were within twice the standard deviation. The amount of a triantennary trigalactosylated structure with one outer arm fucosylation (A3G3Fo) was found to be markedly increased in NSCLC patients in comparison to that in healthy volunteers (p < 0.01). No significant positive correlation with other clinical data was found. Serum A3G3Fo levels can thus be a novel marker for the diagnosis of NSCLC. PMID- 11275553 TI - Suppression of the accumulation of triosephosphates and increased formation of methylglyoxal in human red blood cells during hyperglycaemia by thiamine in vitro. AB - The accumulation of triosephosphates and the increased formation of the potent glycating agent methylglyoxal in intracellular hyperglycaemia are implicated in the development of diabetic complications. A strategy to counter this is to stimulate the anaerobic pentosephosphate pathway of glycolysis by maximizing transketolase activity by thiamine supplementation, with the consequent consumption of glyceraldehyde-3-phosphate and increased formation of ribose-5 phosphate. To assess the effect of thiamine supplementation on the accumulation of triosephosphates and methylglyoxal formation in cellular hyperglycaemia, we incubated human red blood cell suspensions (50% v/v) in short-term culture with 5 mM glucose and 50 mM glucose in Krebs-Ringer phosphate buffer at 37 degrees C as models of cellular metabolism under normoglycaemic and hyperglycaemic conditions. In hyperglycaemia, there is a characteristic increase in the concentration of the triosephosphate pool of glycolytic intermediates and a consequent increase in the concentration and metabolic flux of the formation of methylglyoxal. The addition of thiamine (50-500 microM) increased the activity of transketolase, decreased the concentration of the triosephosphate pool, decreased the concentration and metabolic flux of the formation of methylglyoxal, and increased the concentration of total sedoheptulose-7-phosphate and ribose-5-phosphate. Biochemical changes implicated in the development of diabetic complications were thereby prevented. This provides a biochemical basis for high dose thiamine therapy for the prevention of diabetic complications. PMID- 11275554 TI - Cloning and differential expression of new calcium, calmodulin-dependent protein kinase II isoforms in Xenopus laevis oocytes and several adult tissues. AB - The different oligomers composing the high molecular weight calcium/calmodulin dependent protein kinase II (CaMKII) holoenzyme, previously shown to be transiently activated during Xenopus oocyte maturation, migrate on SDS-PAGE as proteins of 83, 72, 62, 56, and 52 kDa and have all been cloned. The holoenzyme consists of the CaMKII isoforms gammaB, gammaC, and delta12, already described in other species, while gammaJ, gammaK, gammaL, gammaM, and gammaN are now described for the first time. The gamma-isoforms are splice variants of the gamma-gene, containing in their variable region different combinations of known exons and one, two or three novel exons. Semi-quantitative RT-PCR revealed that all isoforms identified in prophase oocytes are also expressed in adult tissues with a tissue-specific expression pattern. At least thirty different CaMKII isoforms could be identified in different Xenopus adult tissues, most of which are described here for the first time. PMID- 11275555 TI - Identification and cloning of genes associated with the guinea pig skin delayed type hypersensitivity reaction. AB - Although the cellular and molecular mechanisms underlying the delayed-type hypersensitivity (DTH) reaction have been investigated, the functions of infiltrating leukocytes and skin resident cells in the elicitation phase of the DTH reaction are not completely understood. To gain more insight into the role of these cells in the DTH reaction, we identified about 250 cDNA fragments showing elevated expression during the DNCB-induced guinea pig skin DTH reaction by differential display analysis. Characterization of 50 of them led to the identification of 28 genes whose expression was elevated in the DNCB-induced DTH reactive tissue. Sequencing of the 28 cDNA fragments and homology search analysis demonstrated that 10 of them represented known genes, some of which, in particular elafin (an elastase inhibitor) and ferritin, are considered to play roles in the DTH reaction. The other 18 fragments are probably derived from unknown genes. Cloning of the cDNAs of one of these genes indicated that it is that for guinea pig tryptophanyl-tRNA synthetase (WRS), a protein found to be induced by interferon-gamma and upregulated during the late stages of mononuclear phagocyte maturation in vitro. Strong induction of the WRS gene during the DTH reaction suggests its involvement in the in vivo immune response. PMID- 11275556 TI - Chimeric Na(+)/H(+) antiporters constructed from NhaA of Helicobacter pylori and Escherichia coli: implications for domains of NhaA for pH sensing. AB - In order to delineate regions which play a role in the regulation of Na(+)/H(+) antiporter NhaA activity by pH, we analyzed the antiporter activities of various chimeric mutants constructed from specific portions of NhaA from Escherichia coli and Helicobacter pylori (EC and HP NhaA). HP NhaA contains 10 residues at the amino-terminus, and 38 residues in a loop region between the eighth and ninth transmembrane spans (loop 8), which are absent in EC NhaA. Deletion from HP NhaA or insertion into EC NhaA of the sequences caused almost no change in pH dependent antiport activities relative to in the case of the wild-type parent molecules. Chimeras consisting of various combinations of the amino-terminal (amino terminus to sixth or eighth transmembrane span) and carboxy-terminal (seventh or ninth transmembrane span to the carboxy-terminus) regions of EC and HP NhaA showed antiporter activity profiles intermediate between those of the parent molecules. These results show that the two HP-specific sequences are not directly involved in the mechanism of pH sensing by HP NhaA and that the pH sensitivity of NhaA activity is not determined by the amino- or carboxy-terminal regions of NhaA alone, but may be due to interaction between unconserved residues in the two domains. In addition, it was suggested that loop 8 functions primarily as a hinge in both NhaA molecules. PMID- 11275557 TI - Peptide mimics of monocyte chemoattractant protein-1 (MCP-1) with an antagonistic activity. AB - In this study, we attempted to analyze the peptide motifs recognized by 24822.111 and F9, monoclonal antibodies (mAbs) that inhibit the chemotactic activity of monocyte chemoattractant protein-1 (MCP-1), a member of the CC subfamily of chemokines. We isolated phage clones from a phage display library and identified six peptide motifs. One of these clones, C27, was strongly and specifically recognized by 24822.111 mAb, while another, G25, was similarly recognized by F9 mAb. Both the C27 motif and the G25 motif contain two cysteines in their sequences and have little homology to the primary amino acid sequence of MCP-1. These clones, however, bound to THP-1 cells, and the binding was competitively inhibited by MCP-1. The clones strongly inhibited the MCP-1-induced chemotaxis of human monocytes. The synthetic and intramolecularly disulfide-linked peptides of C27 and G25 (sC27 and sG25) also inhibited the chemotaxis induced by MCP-1, while their derivatives with serine in place of cysteine did not, suggesting the importance of the loop structure for the inhibition. These results suggest that sC27 and sG25 may mimic the MCP-1-binding domain to the MCP-1 receptor. PMID- 11275558 TI - A pyrrolidinone derivative inhibits cytokine-induced iNOS expression and NF kappaB activation by preventing phosphorylation and degradation of IkappaB-alpha. AB - We previously showed that 1-[3-(3-pyridyl)-acryloyl]-2-pyrrolidinone hydrochloride (N2733) inhibits lipopolysaccharide (LPS)-induced tumour necrosis factor (TNF)-alpha secretion and improves the survival of endotoxemic mice. Since overproduction of nitric oxide (NO) by inducible NO synthase (iNOS) in vascular smooth muscle cells (VSMCs) is largely responsible for the development of endotoxemic shock, and iNOS gene expression is mainly regulated by LPS and inflammatory cytokines, we studied whether or not N2733 affects interleukin (IL) 1beta-induced iNOS gene expression, NF-kappaB activation, and NF-kappaB inhibitor (IkappaB)-alpha degradation in cultured rat VSMCs. N2733 dose-dependently (10-100 microM) inhibited IL-1beta-stimulated NO production, and decreased IL-1beta induced iNOS mRNA and protein expression, as found on Northern and Western blot analyses, respectively. Gel shift assay and an immunocytochemical study showed that N2733 inhibited IL-1beta-induced NF-kappaB activation and its nuclear translocation. Western blot analyses involving anti-IkappaB-alpha and anti phospho IkappaB-alpha antibodies showed that IL-1beta induced transient degradation of IkappaB-alpha preceded by the rapid appearance of phosphorylated IkappaB-alpha, both of which were markedly blocked by N2733. N2733 blocked IL 1beta-induced phosphorylated IkappaB-alpha even in the presence of a proteasome inhibitor (MG115). Immunoblot analysis involving anti-IkappaB kinase (IKK)-alpha and anti-phosphoserine antibodies revealed that N2733 inhibited IL-1beta-induced IKK-alpha phosphorylation, whereas N2733 had no inhibitory effect on IL-1beta stimulated p42/p44 MAP kinase or p38 MAP kinase activity. Our results suggest that the inhibitory action of N2733 toward IL-1beta-induced NF-kappaB activation and iNOS expression is due to its blockade of the upstream signal(s) leading to IKK-alpha activation, and subsequent phosphorylation and degradation of IkappaB alpha in rat VSMCs. PMID- 11275559 TI - Crystal structure of cyclodextrin glucanotransferase from alkalophilic Bacillus sp. 1011 complexed with 1-deoxynojirimycin at 2.0 A resolution. AB - 1-Deoxynojirimycin, a pseudo-monosaccharide, is a strong inhibitor of glucoamylase but a relatively weak inhibitor of cyclodextrin glucanotransferase (CGTase). To elucidate this difference, the crystal structure of the CGTase from alkalophilic Bacillus sp. 1011 complexed with 1-deoxynojirimycin was determined at 2.0 A resolution with the crystallographic R value of 0.154 (R(free) = 0.214). The asymmetric unit of the crystal contains two CGTase molecules and each molecule binds two 1-deoxynojirimycins. One 1-deoxynojirimycin molecule is bound to the active center by hydrogen bonds with catalytic residues and water molecules, but its binding mode differs from that expected in the substrate binding. Another 1-deoxynojirimycin found at the maltose-binding site 1 is bound to Asn-667 with a hydrogen bond and by stacking interaction with the indole moiety of Trp-662 of molecule 1 or Trp-616 of molecule 2. Comparison of this structure with that of the acarbose-CGTase complex suggested that the lack of stacking interaction with the aromatic side chain of Tyr-100 is responsible for the weak inhibition by 1-deoxynojirimycin of the enzymatic action of CGTase. PMID- 11275560 TI - A splicing factor, Prp8: preferential localization in the testis and ovary in adult mice. AB - Prp8 is a splicing factor of 220 kDa originally identified in yeast and is a component of the U5 small nuclear ribonucleoprotein particle. Mouse Prp8 cDNA was cloned and shown to share 62.6 and 68.2% sequence identity with the yeast homologue at the amino acid and nucleotide level, respectively, while it differs by only 3 amino acid residues from the human homologue. During mouse embryogenesis, Prp8 is expressed intensely at day 9.5 of gestation, and its expression decreases progressively during embryogenesis. In adult mice, Prp8 is expressed strongly in the testis and moderately in the ovary. in situ hybridization analysis revealed that Prp8 is preferentially expressed in the outer cell layer in the testis, probably in the spermatogonia and primary spermatocytes, and in granulosa cells in the ovary. In Caenorhabditis elegans, microinjection of a double stranded RNA corresponding to a portion of the Prp8 sequence results in the arrest of embryogenesis at the late-gastrulation stage. These results suggest that Prp8 plays an important role in reproduction and development. Prp8 was shown to bind to midkine (MK), a heparin-binding growth factor. Since Prp8 expression partially overlaps with the sites of action of MK, it is possible that binding to Prp8 is involved in part of MK signaling. PMID- 11275561 TI - A cryptic melibiose transporter gene possessing a frameshift from Citrobacter freundii. AB - Wild-type Citrobacter freundii cannot grow on melibiose as a sole source of carbon. The melibiose transporter gene melB was cloned from a C. freundii mutant M4 that could utilize melibiose as a sole carbon source. Although the cloned melB gene is closely similar to the melB genes of other bacteria, it is cryptic because of a frameshift mutation. Site-directed mutagenesis was used to construct a functional melB gene by deleting one nucleotide, resulting in the production of an active melibiose transporter. The active MelB transporter could utilize Na(+) and H(+) as coupling cations to melibiose transport. The amino acid sequence of the C. freundii MelB was found to be most similar to those of Salmonella typhimurium and Escherichia coli MelB. These facts are consistent with the phylogenetic relationship of bacteria and the cation coupling properties of the melibiose transporters. PMID- 11275562 TI - Comparative biochemistry of disintegrins isolated from snake venom: consideration of the taxonomy and geographical distribution of snakes in the genus Echis. AB - Species in the genus Echis have been classified mainly based on their morphological appearance and the analytical patterns of their serum. However, re classification of the genus Echis has recently been suggested by taxonomists, toxicologists, and clinicians, since there have been problems with the current classification, such as the efficacy of antivenoms used for treating bites and the broad geographical distribution of Echis snakes. In this study, we purified five novel disintegrins, the platelet aggregation inhibitors pyramidin A and B from the venom of Echis pyramidum, ocellatin from the venom of Echis ocellatus, and leucogastin A and B from the venom of Echis leucogaster, to compare their sequences and allow us to re-evaluate the classification of various species in the genus Echis. Comparison of the amino acid sequences of five new and four known isolated disintegrins from snake venoms of six Echis species and their distribution strongly support the recent re-classification of the genus Echis. PMID- 11275563 TI - Distribution of the intracellular Ca(2+)-ATPase isoform 2b in pig brain subcellular fractions and cross-reaction with a monoclonal antibody raised against the enzyme isoform. AB - The presence and distribution of sarco-endoplasmic reticulum Ca(2+)-ATPase (SERCA) isoform 2b in microsomes and other subcellular fractions isolated from pig brain has been demonstrated by the combined use of a specific antibody raised against the SERCA2b isoform and ATP phosphorylation experiments. All subcellular fractions show an approximately 110 kDa phosphorylated protein, the band intensity being stronger in microsomes. Preliminary treatment of the samples with trypsin generates two phosphorylated fragments of about 57 and 33 kDa in the presence of Ca(2+). The observed fragments are typical trypsinized products of the SERCA2b isoform. The monoclonal antibody Y/1F4 raised against the sarcoplasmic reticulum Ca(2+)-ATPase (isoform 1) binds to the 110 kDa band in membranes isolated from brain. The binding was stronger in microsomes than in other fractions. Furthermore, this antibody also recognizes a clear band at around 115 kDa. This band is always stronger in plasma membrane than in synaptosomes or microsomes and is unaffected by trypsin. Phosphorylation studies in the absence of Ca(2+) suggest that the 115 kDa protein is not a Ca(2+)-ATPase. PMID- 11275564 TI - Production of a biologically active epidermal growth factor fusion protein with high collagen affinity. AB - Collagen is generally incapable of capturing polypeptides such as growth factors in a specific manner. In this study, we established a collagen-binding growth factor (FNCBD-EGF) consisting of epidermal growth factor (EGF) and the fibronectin collagen-binding domain. A typical yield of FNCBD-EGF was approximately 200 microg/ml culture in an Escherichia coli expression system. This fusion protein bound to gelatin and fibrillar collagen sponges, and the bound protein was not effectively eluted even with 2 M NaCl. In addition, FNCBD EGF bound to type I, II, III, or IV collagen-coated plates, and the specificity of binding was confirmed by competitive inhibition using fibronectin. FNCBD-EGF substantially stimulated cell growth after binding to collagen-coated culture plates, whereas EGF had no effect, indicating that this fusion protein acted as a collagen-associated growth factor. In an animal model of impaired wound healing, FNCBD-EGF, but not EGF, was retained with collagen sponges at wound sites 4 d after implantation, and repair of epidermis was observed underneath the sponges. These results suggested that our fusion protein with high collagen affinity would be useful for wound healing. PMID- 11275565 TI - Tip60 is a cell-type-specific transcriptional regulator. AB - Tip60 was originally identified as cellular HIV-Tat interacting protein and has been shown to augment Tat-dependent transcription. It has also been shown to interact with various cellular transcription factors and to belong to the nuclear histone acetyltransferase (HAT) family. To further elucidate the function of Tip60 and its HAT domain in transcription regulation, we compared Tip60 activity in HeLa and Jurkat T lymphoma cells. Here we show that Tip60 augments the HIV-1 Tat activity at the HIV-LTR promoter in HeLa but inhibits it in Jurkat cells. Moreover, we isolated two new variants of the Tip60 protein (Tip60Delta1, Tip60Delta2) from Jurkat cells. The Tip60Delta2 variant lacks the entire HAT domain but modulates HIV-1 Tat activity like full-length Tip60. In addition, Tip60 and the transcriptional repressor ZEB (zinc finger E box binding protein) interact specifically in the yeast two-hybrid system and additively inhibit the CD4 enhancer/promoter activity in Jurkat cells. Thus, Tip60 may function as corepressor of the ZEB protein. In summary, these data show that Tip60 functions as a cell-type-specific transcriptional regulator and that the HAT domain is not required for either transcriptional activation or inhibition. This indicates that Tip60 may function by recruiting additional cell-type-specific cofactors. PMID- 11275566 TI - HMG box A in HMG2 protein functions as a mediator of DNA structural alteration together with box B. AB - Nonhistone protein HMG2, like HMG1, binds with B-DNA in a sequence-nonspecific manner and causes structural alterations in DNA such as bending, kinking and unwinding. Here, we studied the functions of HMG2 domains in the DNA structural alteration and modulation by using various HMG2 peptides, and we demonstrated several new findings. The HMG box itself as a DNA-binding motif may have the basic function of inducing curvature, resulting in the apparent DNA bending in the DNA cyclization assay, but not of abruptly kinking DNA. The DNA-binding activity of HMG box B, which is enhanced by the presence of box A, together with the flanking regions of box B, causes DNA bending accompanying the kinking of the DNA main chain. The DNA unwinding accompanied by DNA kinking diminishes cruciform structures in supercoiled DNA. Analysis using mutant peptides for box A confirmed that box A in HMG2 functions as a mediator of DNA structural alteration together with box B. The present studies on the functional properties of the respective regions of HMG2 may help to elucidate the protein function. PMID- 11275567 TI - An examination of intimate partner violence in rural communities: results from a hospital emergency department study from Southwest United States. AB - Intimate partner violence (IPV) affects women disproportionately, women being 5-8 times more likely than men to be victimized by their intimates. Women in abusive relationships use a disproportionate share of health care services, including more visits to the emergency departments. Female clients from two hospital emergency departments in the rural Southwest were recruited and administered semistructured interviews. Study findings indicate the prevalence and pervasiveness of IPV among study participants and illustrate the significant effects of and predictive nature of three factors associated with abuse in the past to current IPV. PMID- 11275568 TI - Community interdisciplinary education to promote partnerships in family violence prevention. AB - Family violence is a major social and health problem in the United States. Educational approaches are needed that help professionals and communities develop more effective skills to work with families and communities. This article describes a statewide, interdisciplinary, community-based educational program for professionals and paraprofessionals and a 6-month post-evaluation. Participants reported knowledge and skill development in assessment and interventions, improved use of violence prevention data for planning and interventions, and increased community partnerships and collaborations. Recommendations address violence prevention leadership, funding, infrastructure, interdisciplinary professional education, greater community awareness, and policy development. PMID- 11275569 TI - Domestic violence and help-seeking behaviors among rural women: results from a shelter-based study. AB - Research on domestic violence and help-seeking behaviors of women living in rural communities has been limited. This study adds to existing knowledge by examining this type of violence along with mental health characteristics and related help seeking behaviors of a sample of predominantly Hispanic women seeking shelter at a rural domestic violence shelter. Study participants experienced physical, verbal, emotional, and sexual abuse, harassment, stalking, and abuse with a weapon in their current intimate relationship. Twenty-four percent of study participants of Hispanic backgrounds and 10% of participants from all other racial/ethnic groups reported experiencing all types of abuse listed above. When compared with other study participants, a greater percentage of Hispanic participants indicated that they had thought of and/or attempted suicide. Participants' help-seeking behaviors from formal support systems suggest a mismatch between the types of abuse experienced and the resultant help-seeking behaviors they used. These help-seeking behaviors also indicate the relevance of mental health characteristics (e.g., suicide ideation) in these behaviors. These and findings from other studies may provide the impetus for a systematic documentation of domestic violence and help-seeking behaviors of women living in rural communities. PMID- 11275570 TI - Health care professionals' skills, beliefs, and expectations about screening for domestic violence in a border community. AB - This article describes domestic violence education of health professionals and determines association between screening behavior and preparedness, outcome expectations, and beliefs about how and when to screen. A survey was mailed to all primary care physicians, dentists, and nurse practitioners in El Paso, Texas (n = 561). Return rate was 34.4%. Using linear regression, differences (p < 0.05) were found between dentists and others in percentage of patients screened, education, preparedness, and beliefs. Education had a positive association (p < 0.001) with preparedness, beliefs about when to screen, and outcome expectations, and a negative association with beliefs about how to screen. Preparedness, beliefs, and realistic outcome expectations had a positive association (p < 0.02) with percentage of female patients screened. Education about domestic violence is important in increasing preparedness and influencing beliefs about when to screen and what outcomes can be expected. Educational programs should include not only information, but also skills training to increase perceptions of preparedness. PMID- 11275571 TI - Disagreement about the occurrence of male-to-female intimate partner violence: a qualitative study. AB - This work explored the reasons underlying inter-partner disagreement about the occurrence of intimate partner violence (IPV). Research indicates that partners often do not agree about episodes of conflict in their relationship. We conducted interviews with 48 women and men with and without histories of IPV to investigate this lack of agreement. Participant responses were analyzed and themes were identified about why men and women disagree about episodes of conflict. The main results indicate that participants think women and men remember differently; women remember more than men, both choose what they want to remember, and both remember that they were right in the conflict. This work contributes to understanding the disagreement that occurs between partners. Many of these findings have never been suggested by other IPV researchers. The broad-reaching implications of this study include improvement in the accuracy of measuring IPV. PMID- 11275574 TI - Quantitative magnetic resonance imaging in traumatic brain injury. AB - Quantitative neuroimaging has now become a well-established method for analyzing magnetic resonance imaging in traumatic brain injury (TBI). A general review of studies that have examined quantitative changes following TBI is presented. The consensus of quantitative neuroimaging studies is that most brain structures demonstrate changes in volume or surface area after injury. The patterns of atrophy are consistent with the generalized nature of brain injury and diffuse axonal injury. Various clinical caveats are provided including how quantitative neuroimaging findings can be used clinically and in predicting rehabilitation outcome. The future of quantitative neuroimaging also is discussed. PMID- 11275575 TI - Metabolic recovery following human traumatic brain injury based on FDG-PET: time course and relationship to neurological disability. AB - OBJECTIVE: Utilizing [(18)F]fluorodeoxyglucose positron emission tomography (FDG PET), we assessed the temporal pattern and the correlation of functional and metabolic recovery following human traumatic brain injury. DESIGN AND SUBJECTS: Fifty-four patients with injury severity ranging from mild to severe were studied. Thirteen of these patients underwent both an acute and delayed FDG-PET study. RESULTS: Analysis of the pooled global cerebral metabolic rate of glucose (CMRglc) values revealed that the intermediate metabolic reduction phase begins to resolve approximately one month following injury, regardless of injury severity. The correlation, in the 13 patients studied twice, between the extent of change in neurologic disability, assessed by the Disability Rating Scale (DRS), and the change in CMRglc from the early to late period was modest (r = 0.42). Potential explanations for this rather poor correlation are discussed. A review of the pertinent literature regarding the use of PET and related imaging modalities, including single photon emission tomography (SPECT) for the assessment of patients following traumatic brain injury is given. CONCLUSION: The dynamic profile of CMRglc that changes following traumatic brain injury is seemingly stereotypic across a broad range and severity of injury types. Quantitative FDG-PET cannot be used as a surrogate technique for estimating degree of global functional recovery following traumatic brain injury. PMID- 11275576 TI - Magnetic resonance spectroscopy in traumatic brain injury. AB - Magnetic resonance spectroscopy (MRS) offers a unique non-invasive approach for assessing the metabolic status of the brain in vivo and is particularly suited to studying traumatic brain injury (TBI). In particular, MRS provides a noninvasive means for quantifying such neurochemicals as N-acetylaspartate (NAA), creatine, phosphocreatine, choline, lactate, myo-inositol, glutamine, glutamate, adenosine triphosphate (ATP), and inorganic phosphate in humans following TBI and in animal models. Many of these chemicals have been shown to be perturbed following TBI. NAA, a marker of neuronal integrity, has been shown to be reduced following TBI, reflecting diffuse axonal injury or metabolic depression, and concentrations of NAA predict cognitive outcome. Elevation of choline-containing compounds indicates membrane breakdown or inflammation or both. MRS can also detect alterations in high energy phosphates reflecting the energetic abnormalities seen after TBI. Accordingly, MRS may be useful to monitor cellular response to therapeutic interventions in TBI. PMID- 11275577 TI - A history and review of quantitative electroencephalography in traumatic brain injury. AB - The electroencephalogram (EEG) is a physiologic measure of cerebral function that has been used by some to assess coma and prognosticate survival and global outcome after traumatic brain injury (TBI). Surface recordings of the brain's electrical activity reveal distinct patterns that indicate injury severity, depth of unconsciousness, and patient survival. The data produced with traditional qualitative studies, however, does not allow resolution and quantification of the wave frequency spectrum present in the brain. As a result, conventional EEG typically has only been used for gross and qualitative analyses and is not practical for use in long-term patient monitoring or as a sophisticated prognostic tool. One area of investigation that is working to address the limitations of conventional EEG has been the development and implementation of Fourier Transform (FT) EEG which resolves and quantifies frequency bands present in the brain. When FT analysis is applied to EEG, it provides concurrent and continuous monitoring, resolution, and quantification of all frequencies emitted. This review discusses the history and significance of conventional EEG and provides a review of how FT-EEG, commonly referred to as Quantitative EEG (QEEG), is being used in the clinical setting. The specific applications and significance of QEEG methods regarding treatment of patients with TBI are discussed in detail. The advantages, disadvantages, and future directions of QEEG in TBI are also discussed. PMID- 11275578 TI - Functionally activated brain imaging (O-15 PET and fMRI) in the study of learning and memory after traumatic brain injury. AB - Advances in functional imaging technology and cognitive neuropsychology have resulted in paradigms in which participants can perform cognitive tasks during functional image acquisition. We will discuss the application of two approaches (oxygen-15 positron emission tomography and functional magnetic resonance imaging) that have recently been used to examine components of learning and memory following traumatic brain injury (TBI). Activated functional brain imaging findings that we will discuss may suggest possible functional reallocation and reorganization of brain substrates involved in verbal learning and memory following brain injury. The findings also are clearly in line with other research that indicates a prominent role for the frontal lobes in learning and memory functioning, and support the concept of distributed neural networks for memory related functions, cognitive load, and the potential for examining brain re organization after injury. PMID- 11275579 TI - "Low-tech" neuroprotection for brain injury. PMID- 11275580 TI - The Traumatic Brain Injury Act Amendments of 2000. PMID- 11275582 TI - [Why do autosomal dominant genodermatoses have two different segmentary expressions?]. PMID- 11275583 TI - [What is Verneuil's disease?]. PMID- 11275584 TI - [Sensitization to cockroach allergens evaluated by skin tests in children with atopic dermatitis]. AB - INTRODUCTION: Cockroach and house dust mites (Dermatophagoides pteronyssinus, Dp and farinae, Df) are the most often implicated aeroallergens in severe asthma, hay fever and conjunctivitis. Cockroach allergy is still unknown in atopic dermatitis. PATIENTS AND METHOD: 146 children with atopic dermatitis-aged 6 months to 15 years- have been patch tested with the European standard series and some aeroallergens. We have studied the sensitisation to cockroach allergens and compared to Dp and Df. RESULTS: 113 children reacted positively at least to one of the 3 aeroallergens (77 p. 100), 61 children had a positive reaction to cockroach (42 p. 100) and 29 simultaneously to the 3 allergens. DISCUSSION: Delayed hypersensitivity to house dust mites in young children with atopic dermatitis suggests early epicutaneous sensitization due to an altered epidermal barrier. For us, cockroach could also be implicated in some flare-ups of atopic dermatitis. Eviction of cockroach and house dust mite should be proposed for children with a positive patch test to cockroach. PMID- 11275585 TI - [Liminal perception threshold of cutaneous distension]. AB - BACKGROUND: Cutaneous sensorial perception is complex and proves to be difficult to quantify. Skin sensitivity to mechanical distension is largely unexplored. We have developed an original method aimed at quantifiying these physiological parameters. VOLUNTEERS AND METHODS: The study was performed on 24 adult volunteers aged 20 to 50 years. A Cutometer SEM 474 equipped with 2 and 4 mm diameter probes was used to exert a progressive or a steep suction on the cheeks. The intensities of both the suction and the skin deformation observed at the earliest moment the traction was perceived by the volunteers were recorded. RESULTS: The liminar sensorial detection under progressive suction force was 1.5 times lower when using the 4 mm probe than the 2 mm probe. Suction necessary for liminar sensation was 7 times lower under steep suction than in response to the progressive mode. The skin deformation corresponding to the liminar sensorial perception was almost constant (C=0.5 p. 100) for each subject. By contrast, a large inter individual heterogeneity unrelated to age was present. COMMENTS: Sensitivity of facial skin to stretching can be explored using a precise suction method. Perception is more related to the rate of traction than to the surface area submitted to suction. Cutaneous deformation, nearly constant for each individual, appears to be the main parameter. Steep and progressive suctions appear to stimulate distinct types of mechanoreceptors. The presently described method can be applied in cosmetology and in functional investigation of various diseases such as diabetes, acromegaly and atopic dermatitis among others. PMID- 11275586 TI - [Allergic contact dermatitis to cosmetics containing Melaleuca alternifolia (tea tree oil)]. AB - INTRODUCTION: Melaleuca alternifolia is a coniferous tree found in tropical regions, the needles contain an essential oil that is used in medical and cosmetic products. The essential oil contains turpentines (limonene, alpha pinene, phellandrene) that are potentially allergenic. PATIENTS AND METHODS: In 1997, 1216 patients were patch tested in our dermatoligic unit. Fourteen of them tested because of eczema used products containing tea tree oil. The patients used creams, hair products and essential oils containing Melaleuca alternifolia for cosmetic reasons and to treat skin affections. They were patch tested for a standard panel of allergens, topical emulgators, perfumes, plants, topical medications, metal, gloves, topical disinfectants and preservatives, dental products and rubber derivatives. Products containing Melaleuca alternifolia were tested concentrated or diluted. RESULTS: We report on 7 cases of patients with an allergic contact dermatitis due to tea tree oil. Two of them also exhibited from a delayed type IV hypersensitivity towards fragrance-mix or colophony suggesting the possibility of cross reaction or an allergic group reaction caused by contamination of the colophony with the volatile fraction of turpentines. DISCUSSION: The allergic potential of low concentrations of Melaleuca alternifolia is presumed to be low on healthy skin. Photoaged Melaleuca alternifolia must be considered to be a stronger sensitizer. PMID- 11275587 TI - [Surgical treatment of Verneuil's disease (hidradenitis suppurativa): 15 cases]. AB - INTRODUCTION: Verneuil's disease (hidradenitis suppurativa) is a chronic inflammatory, suppurating and fistulizing disease of apocrine sweat gland-bearing skin. The aim of this study was to describe the surgical treatment, conducted in 15 patients suffering from this disease. PATIENTS AND METHODS: We retrospectively analyzed 15 observations (9 men, 6 women, mean age 38.6). The mean delay between beginning of the symptoms and diagnosis was 55.5 months. Surgery was conducted at the stage of abscesses, fistulization and keloids in all patients. The first surgical step was wide and deep excision of affected skin and subcutaneous fat. The second step was secondary intention healing, or ideal suture, Z plasty or dorsalis major flap. RESULTS: Only four patients had complications: two axillary strictures, one anal margin stenosis and one hypertrophic scarring. Three relapses occurred, treated by excision under local anesthesia. CONCLUSION: Surgical treatment seems to be the only definitive treatment of Verneuil's disease. PMID- 11275588 TI - [Subcutaneous mycosis due to Scopulariopsis brevicaulis in an immunocompromised patient]. AB - BACKGROUND: Scopulariopsis brevicaulis is a causal agent of onchomycosis. We report the unusual clinical manifestations caused by this opportunist fungus. CASE REPORT: A 61-year-old man consulted in February 1997 for a budding lesion located on the right medial malleolus. This patient had had a liver transplantation for primary biliary cirrhosis in 1990 and had been taking prednisone and cyclosporine since this time. Cyclosporine had been recently replaced by tacrolimus. The histology examination of a lesion specimen taken from the ankle evidenced a dermal mycosis due to opportunist filamentous fungus. Total excision was performed. The patient then developed nodular lesions of the left elbow during the summer of 1997. Mycology culture of a skin biopsy grew numerous colonies of Scopulariopsis brevicaulis. Excision of the elbow lesion was delayed due to hospitalization for suspected graft rejection and development of insulin dependent diabetes. The elbow lesion was then resected followed by a skin graft. The mycology examination identified the same causal agent. DISCUSSION: This liver transplant recipient developed two unusual extra-ungual localizations (ankle and elbow) of a Scopulariopsis brevicaulis infection. Chronic immunosuppression favored development of the infection with a pseudo-epitheliomatous presentation. The histology and mycology examinations were necessary for positive diagnosis. PMID- 11275589 TI - [Sodium valproate-induced kinky hair]. AB - BACKGROUND: We report a case of acquired progressive kinking hair, a rare pilar dysplasia. This is the first case induced by sodium valproate. CASE REPORT: A 6 year-old girl, treated with valproate, observed a progressive change in the texture of her hair. They were kinky, lusterless, dry and, under light microscopy, exhibited an aspect of pseudo-pili torti. DISCUSSION: This child had acquired progressive kinking hair syndrome most likely induced by valproate. Five other cases of valproate-induced pilar dysplasia have been reported but without clinical descriptions. CONCLUSION: We propose adding sodium valproate to the list of drugs susceptible of inducing kinky hair syndrome. PMID- 11275591 TI - [Mycobacterium malmoense cutaneous infection in an immunocompetent patient]. AB - BACKGROUND: Mycobacterium malmoense is a mycobacterium rarely described as a human pathogen. We report the first case of cutaneous infection in an immunocompetent patient. CASE REPORT: A 75-year-old woman presented with a cutaneous nodula on the back of her left hand. Histology of the node biopsy showed non caseating granuloma with giant and epithelioid cells. Acid-fast bacilli were isolated at direct smear and after culture. Mycobacterium malmoense was identified. The lesion has healed after surgery. DISCUSSION: Mycobacterium malmoense is an environmental mycobacterium isolated from soil and water. Typically, most patients with Mycobacterium malmoense infections are old people with previously lung disease. A few cervical lymphadenitis in children and rare cases of tenosynovitis of the hand have been reported. Disseminated infections with cutaneous lesions have been exceptionally described in immunocompromised patients. Mycobacterium malmoense is slow growing and its identification is hard, sometimes requiring molecular analysis. Probably it's the reason why we present the first description of a cutaneous infection in an immunocompetent patient. This case indicates that Mycobacterium malmoense is also a cutaneous pathogen. PMID- 11275590 TI - [Dyschromatosis universalis: two cases]. AB - BACKGROUND: Universalis dyschromatosis is a rare genodermatosis. Melanogenesis dysfunction appears to be the main etiology. We report two cases, discussing the clinical features, diagnosis and etiology of this disease. CASE REPORTS: A 21 year-old man was referred for a mixture of achromatic and hyperchromatic lesions that had progressed on sun-exposed skin areas since birth. The histopathologic study evidenced increased melanin content in the basal cell zone but no changes in the aspect or the number of melanocytes. The second case was a 20-year-old man who presented the same clinical features. His brother also had these lesions. A scotch test and a Wood light test were negative. DISCUSSION: Universalis dyschromatosis is a generalized leukomelanoderma. The familial nature of the disease in our second case points out the genetic component. Recent ultrastructure studies have demonstrated the melanogenesis dysfunction involved. The location of the lesions on sun exposed areas points out to the role of ultraviolet light. In Tunisia, Xeroderma pigmentosum is the main differential diagnosis of universalis dyschromatosis. PMID- 11275592 TI - [Cutaneous metastases revealing gastric linitis]. AB - BACKGROUND: Cutaneous metastases from gastric carcinoma are uncommon and are exceptionally the first sign of disease. CASE REPORT: A 33-year-old man presented with nodular lesions that had developed three months earlier on the trunk, scalp and limbs. A skin biopsy suggested a metastatic origin. The upper digestive fibroscopy with biopsy demonstrated gastric linitis. Due to the presence of bone metastasis, treatment was not initiated, the patient died four months later. DISCUSSION: Metastatic dissemination of gastric carcinoma to the skin usually occurs in advanced stage disease. Generally, the skin lesions are painless hard nodules on the anterior abdominal wall. Our patient developed unusual inaugural skin metastases with an exceptional localization. PMID- 11275593 TI - [Complete supernumerary breast on the thigh in a male patient]. AB - BACKGROUND: Supernumerary nipples are not rare but the developement of a complete supernumerary breast is exceptional. CASE REPORT: A 59-year-old man presented a progressively increasing ancient right- groin-masse. The histopathologic examination of the lesion confirmed the diagnostic of polymastia. DISCUSSION: The interest of this observation results from the very unusual occurence of polymastia (less than 1 p. 100 of supernumerary nipples), especially for a caucasian man. The prevalence appears to be higher in women and oriental people. Diagnostic of supernumerary nipple is difficult because of its atypical appearance and ectopic location. However, this diagnostic is important because ectopic breast tissue is subject to the same pathologic changes that occur in normally positioned breasts and it can be a marker for associated diseases such as urologic malformations or urogenital malignancies. PMID- 11275594 TI - [Night flight. Aviation dermatology: anecdote or trend?]. PMID- 11275595 TI - [Deep vein thrombosis and pulmonary embolism in a patient with aphtosis treated with thalidomide]. PMID- 11275596 TI - [Neonatal curettage of giant congenital naevi]. PMID- 11275598 TI - [Black tumor]. PMID- 11275599 TI - [Palatine lesion]. PMID- 11275600 TI - [Hidradenitis suppurativa (Verneuil's disease)]. PMID- 11275601 TI - [Cutis marmorata telangiectatica congenita]. PMID- 11275602 TI - [Erythema nodosum]. PMID- 11275603 TI - [Dubreuilh melanoma(lentigo maligna melanoma)]. PMID- 11275604 TI - [Alopecia areata: management strategy]. PMID- 11275605 TI - [Pigmented lesion]. PMID- 11275606 TI - [Is there a contraindication to copper intra-uterine devices in patients with hand hyperhidrosis?]. PMID- 11275607 TI - [Cutaneous hyperergia after an insect bite]. PMID- 11275608 TI - [Photoallergy to Zovirax cream]. PMID- 11275609 TI - [Acute porphyria and indinavir]. PMID- 11275610 TI - [Angular cheilitis: how do you manage?]. PMID- 11275612 TI - [Gastroenterologie Clinique et Biologique: the team moves on]. PMID- 11275611 TI - [What is bed rest good for?]. PMID- 11275613 TI - [Evidence-based digestive surgery]. PMID- 11275614 TI - [Comparative effects of oral rehydratation solutions in experimental cholera in the rat]. AB - The composition of the World Health Organisation (WHO) solution in oral rehydration therapy has remained controversial because of its total osmolarity (303 mosm/L) and higher sodium concentration (90 mEq/L), increasing the risk of hypernatraemia. AIM OF THE STUDY: To compare the efficacy of two reduced osmolarity oral rehydration solutions (S1: 268 mosm/L and 50 mEq/L Na(+); S2: 240 mosm/L and 60 mEq/L Na(+) ) with the WHO recommended formula taken as the reference solution. Water, electrolytes and glucose fluxes were directly measured in vivo, in isolated ligated loops of rat jejunum (n=12). Intestinal secretion was induced by exposing jejunum to cholera toxin (CT=20 microg/loop). RESULTS: All three test solutions similarly reversed cholera toxin-induced net water absorption (3.37 +/- 1.35; 3.31 +/- 0.43 and 3.13 +/- 0.66 microL/min.cm(2) for S1, S2 and WHO solutions respectively). However, net Na secretion induced by cholera toxin was observed with S1 and S2 while Na absorption occurred with the WHO solution. CONCLUSION: For a same amount of water absorbed, Na absorption from reduced - osmolarity rehydration solutions is lower than with the WHO solution. Our data may contribute to a better rationale for the use of orally administered hydration solutions in man. PMID- 11275615 TI - [Contribution of endorectal ultrasonography in preoperative evaluation for very low rectal cancer]. AB - Abdominoperineal resection is the standard treatment of very low rectal carcinoma. Pretherapeutic evaluation of locoregional extension relies mainly on digital rectal examination. The interest of endorectal ultrasonography to assess lateral and inferior margins is still to be determined. AIM OF THE STUDY: To assess the ability of endorectal ultrasonography to evaluate the possibility of conservative anal sphincter surgery. PATIENTS AND METHODS: Between April 1996 and June 1998, 34 patients (20 men, 14 women, mean age: 61 years, range: 43-80) have been treated for rectal adenocarcinoma. Endorectal ultrasonography was made with a linear probe (EUP-U33). Before treatment, the mean distance between the lower pole of the tumor and the anal verge was 3.9 cm (range: 2-5), and between the lower pole and the puborectalis sling 2.3 mm (range: 0-7). A uTN classification was made. Preoperative treatment was radiotherapy (40 Gy in 4 patients, 60 Gy in 24 patients), or radiochemotherapy (6 patients). Pre- and post-radiotherapy endorectal ultrasonography results were compared to the patholocical analysis of operative specimen. RESULTS: Wall infiltration was correctly evaluated in 57% of patients after radiotherapy. In 26/34 cases, a safe plane existed before and after radiotherapy, and correlation of endorectal ultrasonography with histology was 96%. For patients without safe plane, correlation with histology was 75%. CONCLUSION: For very low rectal tumors, with an aggressive sphincter conservation approach, endorectal ultrasonography allows to assess sphincter invasion with 96% fiability when safe plane exists. PMID- 11275616 TI - [Prospective randomized single-blind trial comparing oral sodium phosphate with polyethylene glycol for colonoscopy preparation]. AB - AIM AND METHODS: The aim of this prospective, randomized, study performed in 60 outpatients was to compare 2 precolonoscopy bowel preparations: oral sodium phosphate (NaP) versus standard polyethylene glycol-based lavage solution (PEG). None of the patients met any of NaP exclusion criteria. All patients were prepared on the day prior to colonoscopy. A patient-questionnaire and measure of serum electrolytes (calcium, phosphate, sodium, potassium), pulse and blood pressure were used to assess tolerance and acceptability of the preparation. The quality of colon cleansing was judged by blinded endoscopists. RESULTS: Patient's tolerance to NaP was superior to PEG: NaP preparation was easier to drink and feelings of abdominal plenitude occurred in a smaller proportion of patients. A potassium decrease, a sodium increase and hyperphosphatemia were observed in the NaP group but without clinical events. PEG preparation seemed to allow a better cleansing ability compared with NaP but this difference was not statistically significant. CONCLUSIONS: NaP solution was better tolerated and accepted by patients. Colonic preparation quality compared to PEG is still to be discussed depending on the intake schedule. A biochemical data check seems necessary on account of significant serum electrolytes changes induced by NaP preparation. PMID- 11275617 TI - [Contribution of endo-anal sonography in non-tumoral anus diseases]. PMID- 11275618 TI - [Can liver needle biopsy be standardized?]. PMID- 11275619 TI - [Liver transplantation in an era of organ shortage]. PMID- 11275620 TI - [Practices of transcutaneous liver biopsies in France. Results of a retrospective nationwide study]. AB - OBJECTIVES: Few nationwide studies have evaluated the number of transcutaneous liver biopsies performed for diffuse parenchymal liver diseases and the practices of this procedure. The aims of this retrospective nationwide survey were to precise these data. METHODS: In 1997, a confidential questionnaire was mailed to all AFEF and ANGH members. Parameters studied were annual number of transcutaneous liver biopsies performed by center for diffuse parenchymal liver diseases, sedation and/or premedication, haemostasis parameters required for choosing transcutaneous liver biopsy route, fasting liver biopsy, use of venous access, ultrasonography use during liver biopsy (determination of puncture site), modalities of follow-up after liver biopsy, number of biopsies performed as day care procedure. RESULTS: Sixty seven centers were involved in the study. About 12 000 transcutaneous liver biopsies are performed each year in France for diffuse liver parenchymal diseases. Mean number of biopsies per center is 130 (median 70, ranges 5-600). Sedation is routinely used before liver biopsy in 31% of centers; APTT is not measured in 20% of centers and bleeding time is measured in 30% of centers before liver biopsy. Ultrasonography for determination of puncture site is used in 41% of centers. Venous access is implemented in 36% of centers. Outpatient liver biopsies are performed in less than 15% of cases by 64% of centers whereas 30% of centers practice outpatient liver biopsy of more than 50% of cases. Heterogeneity of biopsy practices are related to individual choices rather than the type or location of medical practice. CONCLUSIONS: Many transcutaneous liver biopsies are performed each year in France for diffuse parenchymal liver diseases, and practices vary greatly. Ultrasonography use and outpatient liver biopsy should be developed. PMID- 11275621 TI - [New bile acid biosynthesis pathways]. PMID- 11275622 TI - [HIV and the liver]. PMID- 11275623 TI - [Thrombosed aneurysm of the femoral vein mimicking a strangulated crural hernia]. AB - Venous aneurysms are usually difficult to diagnose, especially when they are located in the inguinal area where they can be misinterpreted as inguinal or femoral hernias. We report the case of a 70-year-old man with a painful thrombosed aneurysm of the femoral vein that was clinically undistinguishable from a strangulated hernia. The ultrasonogram, performed 2 months before the acute pain when the femoral swelling was painless, seemed to confirm the incorrect diagnosis of femoral hernia, and the aneurysm was finally labeled as such during surgical exploration. Massive embolism to the lungs and heart occurred peroperatively and resulted in death. PMID- 11275624 TI - [Acute intracranial hypertension and anicteric cholestasis revealing Whipple's disease without digestive involvement]. AB - Whipple's disease is a rare infectious disease with potential central nervous system manifestations and a poor prognosis. We report the case of a young woman who presented with acute intracranial hypertension associated with cholestasis which revealed Whipple's disease without digestive involvement. The diagnosis was supported by the presence of PAS-diastase positive hepatic granulomas. A long course of antibiotics resulted in complete remission of the disease without relapse. An acute neurologic syndrome associated with cholestasis should suggest Whipple's disease. PMID- 11275625 TI - [Anesthesia for digestive endoscopy]. PMID- 11275626 TI - Renal effects of COX-2-selective inhibitors. AB - Although nonsteroidal anti-inflammatory drugs (NSAIDs) effectively treat a variety of inflammatory diseases, these agents may cause deleterious effects on kidney function, especially with respect to solute homeostasis and maintenance of renal perfusion and glomerular filtration. NSAIDs act by reducing prostaglandin biosynthesis through inhibition of cyclooxygenase (COX) which exists as two isoforms (COX-1 and COX-2). NSAID-induced gastrointestinal toxicity is generally believed to occur through blockade of COX-1 activity, whereas the anti inflammatory effects of NSAIDs are thought to occur primarily through inhibition of the inducible isoform, COX-2. However, the situation in the kidney may be somewhat different. Recent studies have demonstrated that COX-2 is constitutively expressed in renal tissues of all species; this isoform may, therefore, be intimately involved in prostaglandin-dependent renal homeostatic processes. Drugs that selectively inhibit COX-2 might, therefore, be expected to produce effects on renal function similar to nonselective NSAIDs which inhibit both COX-1 and COX 2. This assertion is borne out by recent clinical studies showing that the COX-2 inhibitors rofecoxib and celecoxib procedure qualitative changes in urinary prostaglandin excretion, glomerular filtration rate, sodium retention, and their consequences similar to nonselective NSAIDs. It, therefore, seems unlikely that these COX-2 inhibitors (and perhaps their successors) will offer renal safety benefits over nonselective NSAID therapies, and, at this juncture, it is reasonable to assume that all NSAIDs, including COX-2-selective inhibitors, share a similar risk for adverse renal effects. PMID- 11275628 TI - Augmented interleukin-18 production by peripheral blood monocytes in patients with minimal-change nephrotic syndrome. AB - BACKGROUND/AIM: The etiology of minimal-change nephrotic syndrome (MCNS) is poorly understood. It has been proposed that cell-mediated immunity and T-cell activation are key features of this glomerular disease. Interleukin (IL)-18, a novel interferon-gamma-stimulating factor, may act as an important effector molecule involved in various immune responses. To our knowledge, very little is known about the involvement of IL-18 in NCNS. The aim here was to define further the involvement of IL-18 in MCNS. METHODS: To understand the role of this cytokine, in vitro IL-18 levels were analyzed by a sensitive enzyme-linked immunosorbent assay (ELISA) method in 16 patients with MCNS who were either in a stable or active condition. The disease controls included 16 patients with IgA nephropathy (IgAN). The IL-18 levels were compared with values in healthy controls. RESULTS: Significantly increased spontaneous and lipopolysaccharide (LPS)-stimulated production of IL-18 was detected in peripheral blood monocyte (PBM) cultures of MCNS patients with the nephrotic syndrome (NS) as compared with those of normal controls. Moreover, when individual MCNS patients were followed through their clinical illness, IL-18 levels were increased during the active phase and normalized as the patients went into remission. The amounts of IL-18 are significantly correlated with the levels of vascular permeability factor (VPF) in MCNS patients. CONCLUSIONS: Thus, in MCNS patients, the level of IL-18 was increased and this increase was related to the activity of this disease. The data provide circumstantial evidence for a role of IL-18 in MCNS. PMID- 11275627 TI - Comparison of blood loss with different high-flux and high-efficiency hemodialysis membranes. AB - Iron deficiency is a common problem in patients on chronic HD. Earlier studies have shown significant blood loss per HD session. To identify whether the new more biocompatible high-flux or high-efficiency membranes are also responsible for significant blood loss during HD, we quantitated the amount of blood loss associated with 4 commonly used membranes (F-50, F-80, CA-210, and CT-190). The residual blood in each compartment of extracorporeal circuit was quantitated after total lysis of the red blood cells (RBC), hemoglobin assay, and calculation of the RBC volume using the patient's hemoglobin and hematocrit concentrations just prior to the study. The average residual RBC volume in different membranes was 0.2-0.3 ml. The residual RBC volume in the dialysis lines (arterial or venous) was 0.1-0.2 ml and did not correlate with the residual RBC volume in the dialysis membranes. The residual RBC volume in the whole extracorporeal circuit (HD membrane, arterial and venous lines) ranged from 0.5 to 0.6 ml. It was significantly higher with F-50 vs. CA-210. The residual RBC volume in the dialysis membrane was significantly higher in the F-80 vs. CA-210 and CT-190 dialyzers. There was also significant difference in the residual RBC volume in the arterial lines of F-50 vs. CT-190, and F-50 vs. F-80 dialyzers. CONCLUSION: Our results demonstrate for the first time that the total RBC loss per HD session is minimal in chronic HD patients. PMID- 11275629 TI - Adult-onset minimal change disease among Taiwanese: clinical features, therapeutic response, and prognosis. AB - There are some racial differences in the prevalence and prognosis of idiopathic nephrotic syndrome; however, reports about minimal change disease (MCD) in Chinese were rare. We retrospectively analyzed 123 Chinese adults with idiopathic nephrotic syndrome, who received percutaneous renal biopsy in our institution within the last 10 years. In total, 46 patients (37.4%) were compatible with the pathological diagnosis of MCD. The male to female ratio was 1.2:1. The mean age of onset was 30.9 years, and 80% of the patients with MCD were less than 40 years. The mean daily proteinuria was 10.2 g, and serum albumin was 1.8 mg/dl. Azotemia occurred in 16 (35%) of 46 cases; hypertension, 13%; and microscopic hematuria, 13%. High selectivity index for proteinuria (SI <0.1) was noted in 12 (39%) of 31 cases; and high IgE level was found in 83.7% of the study subjects, although only one case had allergic history. Complete remission in 36 MCD patients treated with corticosteroid was achieved by 42% (15/36), 80% (29/36), and 94% (34/36) within 4, 8, and 12 weeks, respectively. The time interval to remission was similar between the younger group (<40 years old, 1.7 months) and older group (>40 years old, 1.6 months). Nineteen (56%) of 34 cases with steroid response did not relapse, and the other cases (44%) had a mean relapse rate of 1.5 times per patient within a period of 45 months. The age of onset in MCD cases was not significantly correlated with steroid-responsive rate, and the time interval to remission. However, a tendency existed between the onset in the young age and the sequentially relapsing rate (p = 0.06). Two cases with primary steroid resistance and 5 cases with frequent relapse or steroid dependence responded well to intravenous pulse therapy of cyclophosphamide, except one refractory case. No thrombotic episode was ever noted in our group. Regarding infectious complications, primary peritonitis occurred in one, pneumonia in one, and cellulitis in 6 cases during active nephrotic stage. Two mortality cases, one with E. coli-related necrotizing fasciitis and one from pneumonia, were noted. In brief, compared with children, adult patients with MCD had lesser high selectivity index for proteinuria, the same steroid-responsive rate (94%), but slower response, and significantly lesser relapsing rate. The intravenous pulse therapy of cyclophosphamide may be an alternative regimen for adult patients with steroid resistance or dependency. In addition, the Asian adult-onset MCD had younger age, male predominance, and lesser relapsing rate in comparison to those of the Western population. PMID- 11275630 TI - Incidence of stroke among chronic hemodialysis patients with nonrheumatic atrial fibrillation. AB - In general, nonrheumatic atrial fibrillation is associated with a high risk of stroke. However, its impact on stroke in the setting of chronic hemodialysis treatment is insufficiently addressed in the literature. We assessed the incidence of stroke among 430 chronic hemodialysis patients and the impact of atrial fibrillation and various other potential risk factors on stroke in a retrospective study covering 1,111.16 patient-years. The overall incidence of stroke was 3.78/100 patient-years. Among patients with chronic atrial fibrillation without any antithrombotic therapy besides regular dialysis anticoagulation, the stroke incidence was 1.0/100 patient-years and did not differ statistically significantly from the rate among patients without this arrhythmia, in whom the incidence was 2.8/100 patient-years (p = 0.220). Conversely, the overall rate of stroke incidence per 100 patient-years was statistically significantly higher in patients with diabetic nephropathy (6.46, p = 0.0036), age > 65 years (5.90, p = 0.0001), moderate to severe hypertension (6.8, p = 0.0017), weight gain of > 2 kg between dialyses as a marker of poor patient compliance (6.47, p = 0.0433), and antithrombotic therapy with salicylates or warfarin (8.33, p = 0.0002), as compared with corresponding groups without these risk factors. Our data suggest that in contrast to other risk factors nonrheumatic atrial fibrillation in itself is not associated with an increased risk of stroke in patients on maintenance hemodialysis treatment. PMID- 11275631 TI - Monocyte-related determinants of inflammation in patients on peritoneal dialysis. AB - AIMS: We studied markers of monocyte activation, i.e., the cell surface expression of CD11b and CD62L, and the serum concentrations of monocyte chemotactic protein 1 (MCP-1; a monocyte-specific chemoattractant) and soluble vascular cell adhesion molecule 1 (sVCAM-1; an adhesion molecule involved in monocyte recruitment) in 20 patients on peritoneal dialysis (PD), in 25 patients with chronic renal insufficiency, and in 27 healthy subjects. RESULTS: Monocytes obtained from the peripheral blood of PD patients had a significantly higher expression of CD62L (p = 0.02) as compared with monocytes from healthy subjects and a lower CD11b/CD18 expression as compared with monocytes collected from healthy subjects (p < 0.001) and from patients with renal insufficiency (p < 0.001). Monocytes from PD patients had, however, the capacity to increase the expression of CD11b following stimulation with a potent chemotactic factor. The serum concentrations of MCP-1 and sVCAM-1 were higher in PD patients (575 +/- 51 and 1,517 +/- 89 ng/ml) than in healthy subjects (225 +/- 17 and 668 +/- 64 ng/ml, respectively; p < 0.001 for both comparisons). There was a correlation between the levels of sVCAM-1 and MCP-1 (r = 0.48, p < 0.05) in patients on PD, but neither correlated with the monocyte expression of CD11b/CD18 or CD62L. The concentration of C-reactive protein was higher in patients on PD as compared with healthy subjects and correlated significantly with the concentration of sVCAM-1 (r = 0.63, p < 0.01). CONCLUSIONS: Monocytes in the peripheral circulation of patients on PD have a CD62L(high)/CD11b(low) phenotype, indicating that they have not undergone complete differentiation. Patients also have an increase in the systemic chemotactic activity for monocytes in combination with increased levels of sVCAM-1 and C-reactive protein. These inflammatory aberrations may play a pathophysiological role in the response to inflammatory and infectious diseases in patients on PD. PMID- 11275632 TI - Kimura's disease in a chronic hemodialysis patient. AB - Eosinophilia is not an uncommon finding in chronic dialysis patients. It is usually benign in nature although definite pathogenesis is unknown. We have encountered a young uremic Chinese adult who developed Kimura's disease after being on maintenance hemodialysis for about 3 years. Asymptomatic eosinophilia had been noted for 1 year and 8 months prior to the development of progressively enlarged neck masses, which leads to the diagnosis of Kimura's disease. In contrast to most cases, eosinophilia was first noticed before the neck masses appeared. There is often a close correlation between Kimura's disease and glomerular disease, where renal involvement is considered as a systemic manifestation. However, we do not have strong evidence to support this relationship between terminal renal failure and Kimura's disease in this patient. To our knowledge, our patient is the first reported case of Kimura's disease occurring in chronic hemodialysis patients. Eosinophilia persisted for nearly 2 years before the neck mass developed and recurred after the excision. Besides, our patient also demonstrated a chronic and recurrent course typical of Kimura's disease. PMID- 11275633 TI - Membranoproliferative glomerulonephritis associated with a mixed-cell germinal ovary tumor. AB - We describe a patient with membranoproliferative glomerulopathy associated with a mixed-cell germinal ovary tumor (embryonal and dysgerminoma components). Advanced renal failure ensued without remission of nephrotic syndrome after surgery. Five other cases of ovary tumor associated with glomerulopathy and reported in the literature are reviewed. The association between membranoproliferative glomerulonephritis and mixed-cell germinal ovary tumor has not been previously reported. PMID- 11275635 TI - Iga nephropathy, antineutrophil cytoplasmic antibodies and crescentic glomerulonephritis in a patient with the Hermansky-Pudlak syndrome. AB - Hermansky-Pudlak syndrome is an uncommon cause of renal dysfunction. Because of the risk of bleeding in this condition, few patients have undergone a renal biopsy. Renal dysfunction has been attributed to the deposition of ceroid pigment in the tubules and interstitial fibrosis. We report a case with renal biopsy findings of ceroid deposition and interstitial fibrosis, but also of mesangial IgA deposition, crescentic glomerulonephritis, and an interstitial lymphocytic infiltrate. Furthermore, perinuclear antineutrophil cytoplasmic antibodies of the IgG subclass were detected in a blood sample. It is well known that ceroid pigment in this syndrome accumulates in monocytes, macrophages and T lymphocytes and it has been suggested that this may affect their function. We suggest that this novel combination of renal changes might be explained on the basis of alterations in immune mechanisms in the Hermansky-Pudlak syndrome. PMID- 11275636 TI - A case of partial carnitine palmitoyltransferase deficiency in a patient undergoing chronic hemodialysis. AB - A 71-year-old male undergoing hemodialysis for chronic renal failure presented with proximal muscle weakness. He had normal levels of serum creatine phosphokinase. The results of nerve conduction velocity studies and a needle exploration electromyogram were normal. Ultrasonography revealed adenomatous enlargement of the parathyroid glands, and he had marked elevation of the serum parathormone level. The level of serum free carnitine before hemodialysis was significantly lower than normal, while the acyl/free ratio was high. However, the muscle carnitine content was within the normal range. Interestingly, partial inactivation of carnitine palmitoyltransferase activity in the muscle was observed in association with the elevation of the serum parathormone level. Uremic myopathy in this case may be caused not only by abnormal carnitine metabolism but could also be attributable to the partial carnitine palmitoyltransferase deficiency associated with secondary hyperparathyroidism. PMID- 11275637 TI - Neurobrucellosis--a rare complication of renal transplantation. AB - Brucellosis is an intracellular bacterial infection contracted by consuming raw milk or by contact with infected cattle. Neurobrucellosis is a rather rare manifestation of brucellosis and has protean clinical presentations characterized by meningoencephalitis, myelitis, myelopathies, subarachnoid hemorrhage and psychiatric manifestations. A depressed immune status is believed to be a risk factor for developing neurobrucellosis. We report a case of neurobrucellosis in a patient 13 years after a cadaveric renal transplantation. Even though a Brucella organism was not isolated from body fluids she satisfied other criteria for establishing the diagnosis. Treatment with doxycycline and rifampin led to a clinical cure as well as to marked improvement in the Brucella titer. PMID- 11275634 TI - Acute renal failure in a renal transplant donor due to primary antiphospholipid syndrome. AB - Primary antiphospholipid antibody (APA) syndrome, a common prothrombotic disorder, has been known in dialysis patients and renal transplant recipients. We report a case of primary APA syndrome presenting as a posttransplant complication in a renal transplant donor. A renal donor presented with acute, painless anuria due to renal artery thrombosis 6 years following renal transplant surgery, subsequent thrombosis of jugular catheter and arteriovenous fistula occurred, despite anticoagulation treatment, due to primary APA syndrome. This incident represents the most catastrophic complication reported in a renal donor due to primary APA syndrome. The validity of a prothrombotic assay in an organ donor workup to detect predilection to hypercoagulable disorders and to prevent such complications is open to question. The actual significance of APA in the blood is unclear; hence, the presence of APA in a potential renal donor would pose an ethical and practical dilemma. PMID- 11275638 TI - Oxalate-induced redistribution of phosphatidylserine in renal epithelial cells: implications for kidney stone disease. AB - AIMS: The present studies assessed the possibility that exposure to oxalate leads to alterations in membrane structure that promote crystal binding to renal epithelial cells. Specifically, we determined whether oxalate exposure produces a redistribution of membrane phosphatidylserine (PS) and an increase in the binding of (14)C-oxalate crystals to renal epithelial cells. METHODS: PS distribution was monitored in MDCK cells and in phospholipid-containing vesicles using NBD-PS, a fluorescent derivative of PS. Superficial PS was also detected by monitoring the binding of annexin V to MDCK cells. RESULTS: Oxalate exposure rapidly increased the abundance of superficial NBD-PS and increased the binding of annexin V to MDCK cells. Oxalate exposure also increased PS at the surface of phospholipid vesicles, suggesting that oxalate may interact directly with PS. The oxalate concentrations that increased superficial PS also increased binding of (14)C oxalate crystals to MDCK cells, and the increased crystal binding was blocked by annexin V. CONCLUSIONS: These findings provide direct evidence that oxalate exposure promotes both a redistribution of PS and an increase in crystal binding in renal epithelial cells and support the notion that oxalate toxicity may contribute to the development of stone disease by altering the properties of the renal epithelial cell membrane. PMID- 11275639 TI - Renoprotective effect of angiotensin II receptor antagonists in experimental chronic renal failure. AB - We compared the antihypertensive and renoprotective effects of the angiotensin II receptor antagonist losartan and the calcium channel blocker verapamil in the rat with chronic renal failure. One month after five-sixths nephrectomy, male WKY rats were treated for 2 months with either losartan or verapamil. Both resulted in a similar reduction in blood pressure: from 147.1 to 112 mm Hg in losartan treated and from 155 to 118 mm Hg (p = NS) in verapamil-treated rats. Losartan improved the creatinine clearance (difference + 17.1% as compared with + 6.6% for verapamil, p = 0.039) and prevented the increase in proteinuria: 12.26 +/- (SE) 2.33 mg/day before and 18.48 +/- 2.19 mg/day (p = NS) after therapy in the losartan-treated and 17.27 +/- 2.73 mg/day before and 32.27 +/- 10.29 mg/day after therapy (p = 0.0484) in the verapamil-treated group. In addition, losartan resulted in minimal mesangial proliferation and significantly less glomerular sclerosis and thickening of the small arterial and arteriolar walls. The changes in interstitial fibrosis and tubular hypertrophy, however, were similar in both the verapamil- and losartan-treated groups. Treatment with losartan 1 month after five-sixths nephrectomy in male WKY rats resulted in reduced blood pressure, similar to that of the verapamil-treated group. However, despite similar antihypertensive properties, losartan improved creatinine clearance and reduced proteinuria. The losartan-treated group also had a marked reduction in mesangial proliferation and less glomerular sclerosis and less reduced vascular wall thickness in renal small arteries and arterioles. However, losartan did not totally eliminate nephrosclerosis. The tubular and interstitial changes were fewer in the losartan-treated group. Thus losartan has an additional, although only partial, renoprotective effect when compared with verapamil. PMID- 11275640 TI - Fulminant acute nephrotic syndrome in a patient with idiopathic collapsing focal segmental glomerulosclerosis after a febrile illness. PMID- 11275641 TI - Cerivastatin-induced rhabdomyolisis in a patient with diabetic nephropathy. PMID- 11275643 TI - Human placenta as a source of neuroendocrine factors. AB - Progress in the understanding of the physiological and pathological functions of the placenta introduced the concept that the placenta is a neuroendocrine organ, since it shows local production and release of substances analog to neurohormones. These products act as endocrine, paracrine and autocrine factors to control the secretion of other regulatory molecules, including the pituitary hormones of both mother and fetus and their placental counterparts. Furthermore, they may play a role in the regulation of maternal and fetal physiology during pregnancy, ranging from the control of placental anchoring to fetal growth and maturation, fine regulation of uterine blood flow and/or initiation of labor. All this evidence underlines the decisive contribution of the placenta to all phases of gestation, through a range of substances largely exceeding the classically known sex steroids and chorionic gonadotropin, throughout normal pregnancy as well as in the presence of gestational diseases. PMID- 11275644 TI - Maternal risk factors for fetal and neonatal brain damage. AB - Prematurity is probably the major factor associated with brain damage in newborns. Our growing knowledge of the biochemical mechanisms leading to the onset of labour at term allows the biochemical correlates of the epidemiological risk factors for prematurity to be understood. Infection is the major cause of early preterm labour and is now recognised to be a major cause of fetal cerebral damage leading to cerebral palsy. Only some 5% of cerebral palsy is due to intrapartum asphyxia at term. This may occur due to an obstetric catastrophe or through inadequate placental function leading to chronic intrapartum asphyxia. PMID- 11275645 TI - Fetal endocrine signals and preterm labor. AB - Increased uterine contractility at term and preterm results from activation and then stimulation of the myometrium. Activation can be provoked by mechanical stretch of the uterus and by an endocrine pathway resulting from increased activity of the fetal hypothalamic-pituitary-adrenal axis. Cortisol, derived from the fetal adrenal in cases of intrauterine compromise or from the maternal adrenal in response to stress, or generated locally from cortisone in choriodecidual trophoblasts, provides a crucial link to uterine stimulation. Cortisol contributes to the increased production of prostaglandins (PGs) by fetal membranes and the decidua through the upregulation of PG synthase and the downregulation of PG dehydrogenase enzymes. Cortisol also stimulates placental corticotropin-releasing hormone (CRH) output, although CRH may both relax and stimulate uterine activity depending on the distribution and affinity of its receptor subtypes. Other agents such as cytokines may intercede in this sequence to stimulate PGs and/or CRH, giving rise to a cascade phenomenon that results in preterm birth. PMID- 11275646 TI - Influence of maternal stress on fetal behavior and brain development. AB - The very early establishment of certain sensory faculties turns the fetus into a being capable of perceiving multiple stimuli. This perceptive capability forms part of many interchanges between the mother and her developing child. These interchanges are doubtless not only biological and metabolic in nature, but also sensorial and sensitive. The importance of a good quality of psychoaffective communication between mother and child during pregnancy has been shown to be decisive for fetal growth and also for the perinatal period and further development of the child. Maternal psychological stress leads to adverse pregnancy outcome. Chronic anxiety causes an increased stillbirth rate, fetal growth retardation and altered placental morphology. Experimental studies have demonstrated a relationship between specific episodes of maternal psychological stress and exacerbation of fetal asphyxia in utero. It is concluded that all the psychoaffective interchanges between the mother and child are decisive for harmonious fetal growth and brain development. PMID- 11275647 TI - Caspase-3 activation after neonatal rat cerebral hypoxia-ischemia. AB - Caspase-3 is a major effector protease in several apoptotic pathways, but its role in hypoxic-ischemic (HI) brain injury is incompletely understood. Cerebral HI was induced in 7-day-old rats by unilateral carotid artery ligation and exposure to 7.7% oxygen for 55 min. Caspase-3-like activity was significantly increased at 1 h (208%), peaked at 24 h (2,563%) and was still increased 6 days after HI (169%) in the ipsilateral cerebral cortex. Concomitantly, cleavage of the caspase-3 proform (31/33 kD) was detected on immunoblots, producing 29- and 17-kD fragments. Furthermore, significant degradation of the endogenous caspase-3 substrates inhibitor of caspase-activated DNase (DNA fragmentation factor 45), poly(ADP-ribose) polymerase and fodrin occurred. In conclusion, caspase-3 is activated extensively in the immature brain after HI. The subsequent cleavage of proteins involved in cellular homeostasis and repair may contribute to the process of brain injury. PMID- 11275648 TI - Free radicals and brain damage in the newborn. AB - Newborns and particularly preterm infants are at high risk of oxidative stress and they are very susceptible to free radical oxidative damage. Indeed, there is evidence of an imbalance between antioxidant- and oxidant-generating systems which causes oxidative damage. The brain may be especially at risk of free radical-mediated injury because neuronal membranes are rich in polyunsaturated fatty acids and because the human newborn has a relative deficiency of brain superoxide dismutase and glutathione peroxidase. The brain of the term fetus is at higher risk of oxidative stress than that of the preterm fetus, as a consequence of its higher concentration of polyunsaturated fatty acids and the maturity of the N-methyl-D-aspartate receptor system at term. There seems to be a maturation-dependent window of vulnerability to free radical attack during oligodendrocyte development. Early in its differentiation, the oligodendrocyte may be vulnerable because of active acquisition of iron for differentiation at a time of relative delay in the development of certain key antioxidant defenses in the brain. Excess free iron and deficient iron-binding and -metabolizing capacity are additional features favoring oxidant stress in premature infants. Free radicals may be generated by different mechanisms, such as ischemia-reperfusion, neutrophil and macrophage activation, Fenton chemistry, endothelial cell xanthine oxidase, free fatty acid and prostaglandin metabolism and hypoxia. Reactive oxidant production by these different mechanisms contributes in a piecewise manner to the pathogenesis of perinatal brain injury. PMID- 11275650 TI - Is periventricular leucomalacia a result of hypoxic-ischaemic injury? Hypocapnia and the preterm brain. AB - Decrease in the arterial partial pressure of carbon dioxide (PaCO(2)) causes a reduction in cerebral blood flow in humans and in most animal species; in adults as well as in newborns and even in fetal life. Severely decreased PaCO(2) increases cerebral lactate production, modifies spontaneous electric brain activity, and may decrease the metabolic rate of oxygen. A relation between very low PaCO(2) and brain injury, however, has not been shown in adult humans or full term newborn infants, nor in perinatal animals. In contrast, an association between low PaCO(2) and cerebral palsy and white matter injury in preterm infants has been reported repeatedly. A cause-and-effect relation is suggested by data from the immature rat: brain damage induced by ligation of a carotid artery can be reduced by adding CO(2) to the inspired gas and hence avoiding the consequences of spontaneous hyperventilation. This may be relevant for the clinical care of preterm infants, since PaCO(2) to a large extent is a function of respiratory management. The questions to be addressed are whether hypocapnia sensitizes the brain to hypoxaemia, and also whether the escape mechanisms are less effective in the preterm human brain. PMID- 11275649 TI - Effect of graded hypoxia on cerebral cortical genomic DNA fragmentation in newborn piglets. AB - Previous studies have shown that hypoxia is associated with modification of the cerebral cortical nuclear membrane, leading to increased intranuclear calcium. The increased intranuclear calcium activates calcium-dependent endonucleases, resulting in DNA fragmentation. The present study tests the hypothesis that the fragmentation of neuronal genomic DNA increases with an increase in the degree of cerebral tissue hypoxia. Sixteen newborn piglets were anesthetized, ventilated and divided into normoxic and hypoxic groups with varying degrees of hypoxia. Cerebral hypoxia was documented biochemically by measuring tissue levels of ATP and phosphocreatine. Isolation of cerebral cortical neuronal nuclei and DNA and their purity was confirmed by standard techniques. DNA samples were separated by electrophoresis on 1% agarose gel and stained with ethidium bromide. In the hypoxic samples, multiple low-molecular-weight DNA fragments were present as a smear pattern from 200 to 2,000 base pairs. Levels of high-energy phosphates were compared to the area of each smear for each animal to correlate the degree of hypoxia with the degree of DNA fragmentation. DNA fragmentation increased when high-energy phosphate levels decreased. We conclude that there is a critical threshold value of oxidative metabolism beyond which there are progressive changes in the cortical neuronal cells, leading to DNA fragmentation. PMID- 11275651 TI - Chorioamnionitis and fetal/neonatal brain injury. AB - Chorioamnionitis (CA) is the leading cause of preterm birth and neonatal complications. Even in the absence of a proven infection, fetuses and neonates present a systemic inflammatory response which can be identified by radiological and morphological examination of the thymus. The frequent occurrence of brain injury in neonates with CA is probably linked to systemic, unspecific mechanisms which have not yet been completely clarified. Only by relating placental pathology to clinical evaluation of the newborn will it be possible to achieve a better understanding of these infections and to reduce long-term morbidity and mortality. PMID- 11275652 TI - New insights into the pathogenesis of pulmonary inflammation in preterm infants. AB - Chronic lung disease (CLD) and bronchopulmonary dysplasia are associated with a significant inflammatory response of the airways and the interstitium of the lungs. Besides inflammatory cells, various cytokines, lipid mediators, proteolytic enzymes and toxic oxygen radicals may play an essential role in the pathogenesis of this disease. Intrauterine exposure to chorioamnionitis or proinflammatory cytokines has been shown to induce a pulmonary and systemic inflammatory response in the fetus. In this subgroup, antenatal infection may prime the lung such that minimally injurious postnatal events provoke an excessive inflammatory response in the airways and the pulmonary tissue. Inflammation and lung injury most certainly affect normal alveolization and pulmonary vascular development in preterm infants with CLD. PMID- 11275653 TI - Red blood cell involvement in fetal/neonatal hypoxia. AB - Free radical release plays an important role in the development of brain injury following hypoxic-ischemic encephalopathy. It causes endothelial cell damage and anomalies in NMDA receptors, synaptosome structure and astrocyte function. Mitochondrial dysfunctions caused by asphyxia, reperfusion after ischemia, arachidonic acid cascade, catecholamine metabolism and phagocyte activation are known sources of reactive oxygen species, particularly the superoxide anion (O2( )). O2(-) mainly induces peroxidation by the Fenton/Haber Weiss reaction or via iron-oxygen complexes. Since both reactions require reactive heavy metals, non protein-bound iron (NPBI) is essential for the induction of lipid peroxidation. Experimental studies have demonstrated the neurotoxicity of iron in ischemia reperfusion. Normal axonal transport of brain iron is also reported to be disrupted in hypoxia-ischemia, leading to a buildup of iron in the white matter. The free iron content of erythrocytes (ICRBC) is considered a marker of oxidative stress. Free iron release is accompanied by the oxidation of membrane proteins and the appearance of senescent antigen, as measured by autologous IgG binding. Our preliminary results suggest a significant positive correlation between plasma free iron and the number of nucleated red cells in cord blood, currently considered a reliable index of lasting intrauterine asphyxia but also possessing a high predictive value for poor neurodevelopmental outcome. The rate of erythropoiesis and the entity of ICRBC are related to the degree of asphyxia and the probability of neurological impairment. Since even an increase in NPBI during asphyxia is related to a poor outcome, iron released by red cells could possibly also contribute to NPBI levels. PMID- 11275654 TI - Early markers of brain damage in premature low-birth-weight neonates who suffered from perinatal asphyxia and/or infection. AB - We studied 57 low-birth-weight premature neonates, of whom 29 suffered from perinatal asphyxia and/or infection, while the remaining 28 did not and served as controls. We measured peripheral nucleated red blood cell (NRBC) absolute numbers as well as interleukin (IL)-1beta, IL-6 and tumour necrosis factor (TNF)-alpha cytokine serum levels at 24 h postnatally and on days 3 and 7 following birth. Fourteen of the asphyxiated/infected neonates and 12 controls had neurologic assessments at the corrected postnatal age of 18 months. We found NRBC absolute numbers and serum IL-1beta and IL-6 cytokine levels at 24 h postnatally to be significantly higher in neonates with perinatal asphyxia/infection than in the controls (p = 0.022, p = 0.036 and p = 0.037, respectively). TNF-alpha levels did not differ. Neurologic examination at the corrected postnatal age of 18 months showed 8 out of the 14 children who had been asphyxiated/infected as neonates to have abnormal findings, while 12 children who were used as controls during their neonatal period were normal. Abnormal neurologic findings correlated with high NRBC counts and IL-1beta and IL-6 levels at 24 h postnatally. In conclusion, increased NRBC counts and proinflammatory cytokine levels in asphyxiated/infected neonates represent early markers for subsequent neurologic impairment. PMID- 11275655 TI - Prevention of bilirubin encephalopathy. AB - Prevention of bilirubin encephalopathy is based on the detection of infants at risk of developing a significant hyperbilirubinemia. This task can be accomplished by performing a simple umbilical cord blood test, such as blood group, Rh, Coombs' test and glucose-6-phosphate dehydrogenase, in order to detect hemolytic diseases. In preterm infants, the prevention of hyperbilirubinemia with phototherapy is a relatively simple task, since these infants are cared for in hospital. Early hospital discharge of full-term neonates represents a major concern. The management of neonatal jaundice requires that therapy begins when total serum bilirubin levels are significantly below the levels at which kernicterus is considered an immediate threat. Unfortunately, determination of serum bilirubin is a painful procedure, and is not very accurate since there is a high variability in laboratory measurements. The accuracy and precision of a new transcutaneous bilirubin measurement, comparable to the standard of care laboratory test, makes the daily evaluation of transcutaneous bilirubin measurement a useful tool in distinguishing physiological from nonphysiological hyperbilirubinemia, and determining the bilirubin increment in the first days of life. Full-term neonates who lose a significant amount of weight are especially at risk of significant hyperbilirubinemia and must be treated with ad libitum feeding and intensive phototherapy. PMID- 11275656 TI - Inflammatory mediators and neonatal brain damage. AB - Inflammatory mediators are multifunctional cytokines that play important roles both in normal central nervous system (CNS) development and in the response of the brain to diverse forms of injury. Interleukin (IL)-1beta, tumor necrosis factor-alpha and IL-6 are among the best-characterized early-response cytokines. Recent data suggest that they may be synthesized and secreted by several CNS cell types, including microglia, astrocytes and neurons. Biological effects of these cytokines that could influence the progression of injury in the brain include stimulating the synthesis of other cytokines and neuronal injury mediators such as nitric oxide synthase, inducing leukocyte infiltration and the expression of adhesion molecules, influencing glial gene expression and damaging oligodendrocytes. In the immature brain, proinflammatory cytokines might lead to white matter damage during prenatal intrauterine infection and contribute to progressive neuronal damage in acute brain injury evoked by cerebral hypoxia ischemia. Interrupting the proinflammatory cascade might limit the extent of irreversible injury. PMID- 11275657 TI - The biology of erythropoietin in the central nervous system and its neurotrophic and neuroprotective potential. AB - This review summarizes published as well as preliminary data on the biology of erythropoietin (Epo) in the developing and mature human central nervous system (CNS). Both Epo receptor (Epo-R) and Epo gene expression underlie developmental changes and a brain-specific regulation. These features suggest a different role of Epo in normal brain development than in neuroprotection and neuronal tissue repair after brain injury. Epo concentrations in the cerebrospinal fluid may have primary paracrine effects. While the transport of Epo across the intact blood brain barrier (BBB) is generally limited in humans, systemically produced or administrated Epo may cross during BBB dysfunction. Summarized data of the in vivo and in vitro effects of Epo in the CNS show significant neuroprotective and neurotrophic effects of this molecule. These effects are mediated by several mechanisms, including the activation of a variety of genes and their consecutive protein production. Therapeutic strategies involving activation of the CNS Epo-R are discussed, including the potential use of Epo mimetic peptides. PMID- 11275658 TI - Fetal and neonatal cerebral infarcts. AB - Focal arterial infarction in the full-term newborn is an important cause of acquired cerebral lesions in the perinatal period. Clinical motor seizures, most often unifocal, are the nearly constant disclosing symptom confirmed by focal EEG abnormalities. A multifactorial physiopathology is usual, including genetic and perinatal environmental factors. In the past decade, various acquired or genetic thrombophilias have been discussed as risk factors. For several of the involved mechanisms, the excitotoxic cascade could represent a common final pathway leading to neuronal cell death. Early magnetic resonance imaging studies and EEG help to identify the newborns with strokes who are likely to develop hemiplegia and disabilities at school. Protection of the human fetal brain remains difficult, since the triggering factor initiating the excitotoxic cascade is rarely observed. Treatment of seizures is nevertheless necessary, because it seems that they accelerate anoxia-induced neuronal death in animal models of focal hypoxic ischemia. PMID- 11275660 TI - Monitoring of antepartum and intrapartum fetal hypoxemia: pathophysiological basis and available techniques. AB - The challenge of obstetric surveillance is to identify those fetuses whose physiological defence mechanisms are compromised, in order to be able to act before decompensation has occurred. During the antenatal period, the evaluation of fetal hemodynamic adaptation to hypoxemia and the assessment of its chronological evolution by Doppler technology are crucial. During the intrapartum period, the relative inaccessibility of the fetus and the complexity of the pathophysiology of fetal oxygenation make it difficult to obtain and interpret information on the fetal response to labor stress. Due to the limitations of cardiotocography, additional information is required for appropriate decision making during labor. Current evidence suggests that modern technology applied to fetal surveillance can provide useful additional information that can improve our capacity to interpret fetal reactions to labor events. PMID- 11275659 TI - Blood pressure and tissue oxygenation in the newborn baby at risk of brain damage. AB - A cardinal aim of neonatal intensive care is the maintenance of an adequate oxygen supply to the tissues, particularly the brain. This process depends on several factors. These include an adequate blood oxygen content, blood flow to the tissues and the ability of cells to extract and utilise oxygen. Oxygen carriage depends on ventilation and haemoglobin concentration and type. Blood flow depends on cardiac output (in turn dependent on cardiac contractility, heart rate, blood pressure and vascular resistance). Different tissues also have different oxygen demands depending on their oxygen consumption, which are likely to vary within the tissue itself and with the activity of the infant. This paper discusses evidence that suggests that even in preterm neonates, cerebral blood flow may be independent of blood pressure, and that even very low cerebral blood flow seems to be consistent with healthy survival. Evidence is considered that cardiac output rather than blood pressure may be more important in determining brain tissue oxygenation. We have found a negative correlation between cardiac output and cerebral oxygen extraction in preterm infants, but no relationship between mean arterial blood pressure and cerebral oxygen extraction. PMID- 11275661 TI - Glutamate in cerebral tissue of asphyxiated neonates during the first week of life demonstrated in vivo using proton magnetic resonance spectroscopy. AB - We tested the hypothesis that glutamate (Glx) levels as demonstrated by proton magnetic resonance spectroscopy ((1)H-MRS) are elevated in brain tissue of neonates with severe hypoxic-ischemic encephalopathy (HIE). Studies were performed in 26 neonates (median gestational age 40.5 weeks, range 36.7-42.4 weeks; median birth weight 3,360 g, range 2,180-4,200 g). The median postnatal age at the time of testing was 2.5 days (range 1-7 days). HIE was scored according to Sarnat as grade I (n = 4), grade II (n = 15) or grade III (n = 7). Results for neonates with mild to moderate HIE (group 1) were compared to those with severe HIE (group 2). After magnetic resonance imaging, (1)H-MRS was performed in a single volume of interest including the basal ganglia. An echo time of 31 ms was used. After curve-fitting procedures, peak area ratios of different brain metabolites were calculated. The median total Glx/N acetylaspartate ratio was 1.21 (range 0.64-3.25) in group 1 versus 1.55 (range 1.10-2.75) in group 2 (p = 0.035). The median total Glx/choline ratio was 1.33 (range 0.71-2.52) in group 1 versus 2.14 (range 1.21-3.55) in group 2 (p = 0.019). We concluded that during the first days of life, Glx was elevated in the basal ganglia of neonates with severe HIE. PMID- 11275662 TI - Resuscitation of the asphyxic newborn infant: new insight leads to new therapeutic possibilities. AB - The basic mechanisms leading to cell death in birth asphyxia are becoming better known. Some of these are excitotoxicity, inflammation and oxidative stress. In the so-called therapeutic window - between the primary and secondary energy failure - modulation of these processes may be beneficial, reducing apoptosis and perhaps necrosis. In order to reduce oxidative stress, reoxygenation with low oxygen concentrations, even as low as room air, might be beneficial. Increased oxidative stress might have long-term effects on brain growth and development and there is evidence indicating that exposure to 100% oxygen after birth for only a few minutes might have long-term effects. New guidelines for newborn resuscitation have recently been published but more research is needed in this field, especially regarding resuscitation of preterm infants, where few data exist. PMID- 11275663 TI - Six years of experience with the use of room air for the resuscitation of asphyxiated newly born term infants. AB - In the last 6 years, 830 asphyxiated newly born term infants have been resuscitated with room air (RAR; n = 304) or 100% oxygen (OxR; n = 526) in our hospital. We have studied the time to onset of a regular respiratory pattern, morbidity and mortality, blood gases, reduced glutathione (GSH) and oxidised glutathione (GSSG) and antioxidant enzymes in these infants. No significant differences in the effectiveness of either gas sources or in the final outcome have been found. The RAR group required a shorter time of positive pressure ventilation to attain a spontaneous pattern of respiration. The OxR group showed hyperoxaemia during resuscitation, which was positively correlated with increased GSSG concentrations. Significant oxidative stress was found in the OxR group at 28 days of postnatal life when compared with normal control infants and the RAR group. Oxygen concentrations used during the resuscitation of newly born infants should be strictly monitored. PMID- 11275664 TI - Psychological prevention of early pre-term birth: a reliable benefit. AB - OBJECTIVES: After a previous study had shown the existence of psychological risk factors of pre-term delivery, we designed a study aimed at assessing the effect of psychotherapeutic support of pregnant women hospitalised with pre-term labour, followed by a second multicentric study aimed at demonstrating the reliability of such an intervention. METHODS: Both studies were conducted in two successive cohorts of patients hospitalised with pre-term labour at 18-35 weeks of gestation. The initial study comprised 157 patients in each group, whereas the reliability study comprised 191 patients in the experimental group versus 202 in the control group. In each experimental group, the patients were offered psychotherapeutic support in addition to the usual clinical management. The psychological support included interviews with a psychologist and a collaborative work plan implemented with the nursing staff. RESULTS: The analysis, conducted in the 'intention to treat' manner, shows a significant decrease in the early pre term birth rate (< 35 weeks) from 25.7 to 5.9% (p < 0.0001). After controlling for confounding factors, the adjusted relative risk was 0.16 [95% confidence interval (CI) = 0.07-0.37]. These results were confirmed, at a lesser level, in the reliability study, where the early pre-term birth rate changed from 15.7 to 7.2% (p < 0.02) and the adjusted relative risk was 0.35 (95% CI = 0.16-0.78). CONCLUSION: This study offers new and major results related to the prevention of delivery before 35 weeks of gestation, both in the initial study as well as in the reliability study. Thus, providing this type of psychological support to women hospitalised for pre-term labour, in the context of antenatal care, can help to avoid early pre-term births and their complications in terms of brain damage and neuropsychological development. PMID- 11275665 TI - Pharmacotherapeutical reduction of post-hypoxic-ischemic brain injury in the newborn. AB - Perinatal hypoxia-ischemia (PHI) is a major cause of morbidity and mortality. A substantial part of PHI-related brain damage occurs upon reperfusion and reoxygenation by the excess production of excitatory amino acids, free (pro)radicals and the release of cytokines, triggering programmed cell death. In this respect, several neuroprotective agents have been investigated in neonatal animal models, providing evidence for their usefulness in PHI. Several agents have been shown to be neuroprotective in neonatal animal hypoxia-ischemia models, but only a few agents have been used in clinical studies on term newborns. Although some general information will be provided with respect to focal hypoxia ischemia and neuroprotective agents, this paper focuses on the investigated neuroprotective agents for global PHI and reperfusion brain injury in the newborn, categorized by their mode of action. Future experimental and clinical trials with promising neuroprotective agents need to be performed, including long term follow-up to monitor long-term consequences. Moreover, well-designed combinations of neuroprotective agents with or without other neuroprotective strategies such as brain hypothermia should be given consideration for producing the most promising results in reducing post-hypoxic-ischemic reperfusion injury of the newborn brain. PMID- 11275666 TI - Caries detection methods: can they aid decision making for invasive sealant treatment? AB - The decision to place sealants is a difficult one, and it has been suggested that in a low risk population it may be efficient to wait until caries is detected in the fissure. An invasive sealant technique with fissure preparation may then be indicated. The diagnostic method used in the indication of such a procedure should accurately detect both dentine caries and sound fissures: high sensitivity for dentine caries (at D3 threshold) with high specificity for enamel caries (at D1 threshold). The aims of this study were to assess the diagnostic performance of selected diagnostic methods at normal cut-offs for traditional dentine caries detection and at reduced cut-offs in relation to the desired performance mentioned above, and to assess whether fissure opening allows for accurate visual detection of dentinal caries. Data were obtained from 230 occlusal sites of 101 extracted human molar teeth. Diagnostic methods used on the entire sample were: visual inspection, electrical conductance measurements and laser fluorescence measurements. The sample was then divided into two groups. Group 1 was subjected to visual inspection after application of a dye. Group 2 was subjected to visual inspection after fissure opening only, and after subsequent dye application. Validation was performed by histological investigation. The results with cut-offs normally used in dentine caries detection were roughly in accordance with the literature, except for laser fluorescence. The sensitivity of visual inspection for dentinal caries (D3) was 17% before and 70% after fissure opening. Using reduced cut-offs, a 100% sensitivity (D3) was achieved with 2 methods, but this also resulted in 63 or 87% false positive diagnoses of sound surfaces. Visual inspection and electrical methods both showed a moderate to high sensitivity (D3) with a higher than 50% specificity (D1). It was concluded that visual inspection and electrical methods at reduced cut-offs may aid the indication of invasive sealant treatment. The visual detection of dentinal caries is substantially increased, but not perfect after fissure opening. PMID- 11275667 TI - Effectiveness of three minimal intervention approaches for managing dental caries: survival of restorations after 2 years. AB - The present study was aimed at comparing the effectiveness of three minimally invasive restorative treatment approaches for dentinal lesions in occlusal surfaces. The approaches tested comprised a conventional and a modified conventional treatment and a modified Atraumatic Restorative Treatment (ART). The conventional approach was performed in a university dental clinic whilst the modified-conventional, employing portable equipment, and the modified ART, using hand instruments and a caries removal solution (Caridex, were carried out in the field. Using the split-mouth design, 430 matched contralateral pairs of occlusal cavities were restored with amalgam or glass-ionomers in permanent molars of 152 school children by one dental therapist. The restorations were assessed using a modified USPHS-Ryge criteria. After 2 years, the cumulative survival percentages for occlusal amalgam and glass-ionomer restorations were 92 and 96%, respectively. The survival of all restorations in the conventional, modified conventional and the modified ART group was 96, 96 and 91%, respectively. The survival of occlusal amalgam and glass-ionomer restorations per treatment group was as follows: 94 and 99%, respectively (conventional group); 95 and 97%, respectively (modified-conventional group), and 89 and 93%, respectively (modified ART group). The differences in survival percentage between restorations with amalgam and glass-ionomer, and between the three restorative treatment approaches were not statistically significant. In countries facing scarcity in resources for dental care, ART seems a promising restorative approach to occlusal caries in posterior teeth. However, a longer clinical observation period is required to substantiate this initial conclusion. PMID- 11275668 TI - Tooth-surface progression and reversal changes in fluoridated and no-longer fluoridated communities over a 3-year period. AB - OBJECTIVE: To compare permanent tooth surface-specific progression/reversal changes between fluoridation-ended (F-E) and still-fluoridated (S-F) communities in British Columbia, Canada, over a 3-year period. METHODS: D1D2MFS examinations were contrasted for 2,964 schoolchildren in 1993/94 (grades 2, 3, 8 and 9) and 1996/97 (grades 5, 6, 11 and 12). Generalized Estimating Equation (GEE) models explored the relation between progression/reversal changes and fluoridation status, age, gender, socioeconomic status, and dietary/fluoride histories. RESULTS: Within a scenario of low levels of caries overall, few children had multiple surfaces progressing. At least one smooth surface progressed in 31.4% of subjects; at least one pit-and-fissure (PF) surface progressed in 43.1% of subjects. At least one smooth surface reverted in 89% of subjects who had reversible stages; at least one PF surface reverted in 23.8% of subjects who had reversible stages. GEE (smooth) indicated that odds ratios of progression were twice as large in the F-E site compared to the S-F site, and slightly increased in older participants and in participants exposed to more fluoride technologies. GEE (PF) also indicated that progression was slightly more common in the F-E site; more frequent snacking and lower parental educational attainment had modest associations with increased progression in PF surfaces. For the two types of surfaces, GEE models demonstrated that unerupted surfaces were less likely to progress than sound surfaces. No associations were found between reversals and independent variables. CONCLUSION: Progressions were found to be weakly linked to socio-demographic factors; baseline surface statuses were better predictors of progression. Using the current definitions for disease transitions, F-E communities had more frequent progressions than a S-F community. PMID- 11275669 TI - Effect of dentifrice containing fluoride and/or baking soda on enamel demineralization/remineralization: an in situ study. AB - The additive effect of baking soda on the anticariogenic effect of fluoride dentifrice is not well established. To evaluate it, a crossover in situ study was done in three phases of 28 days. Volunteers, using acrylic palatal appliances containing four human enamel blocks, two sound (to evaluate demineralization) and two with artificial caries lesions (to evaluate remineralization), took part in this study. During each phase, 10% sucrose solution was dripped (3 times a day) only onto the sound blocks. After 10 min, a slurry of placebo, fluoride (F) or fluoride and baking soda (F+NaHCO(3)) dentifrice was dripped onto all enamel blocks. The results showed a higher F concentration in dental plaque formed during treatment with F+NaHCO(3) than placebo (p<0.05), but the difference related to F dentifrice was not significant. The enamel demineralization was lower, and remineralization was greater, after treatment with F+NaHCO(3) than placebo (p<0.05), but the difference related to F dentifrice was not significant. The data suggest that baking soda neither improves nor impairs the effect of F dentifrice on reduction of demineralization and enhancement of remineralization of enamel. PMID- 11275670 TI - Fluoride uptake in human teeth from fluoride-releasing restorative material in vivo and in vitro: two-dimensional mapping by EPMA-WDX. AB - Class V cavities were prepared in the upper and lower left second premolars from an individual under infiltration anesthesia. The cavities were filled with fluoride- releasing resin (TF). One month later, the teeth were extracted. As a control (in vitro), the upper and lower right second premolars were extracted at the time of the cavity preparation in vivo. Immediately after extraction, class V cavities were prepared and filled with TF, and immersed in normal saline solution for 1 month at 37 degrees C. All four premolars were bisected longitudinally and the fluoride uptake around the cavity wall on the cut surface was measured using an electron probe microanalyzer-wavelength dispersive X-ray method. The fluoride uptake was given in the form of a two-dimensional map. Comparison of the observed values of each corresponding part of the tooth in vivo and in vitro revealed no characteristic differences. The maps were quite analogous as a whole. PMID- 11275671 TI - Effect of a water rinse on 'labile' fluoride and other ions in plaque and saliva before and after conventional and experimental fluoride rinses. AB - Labile reservoirs are important in maintaining ion concentrations in oral fluids, especially after a fluoride dentifrice application, where a persistent increase in fluid fluoride can mitigate or reverse caries progression. In this study, the effect of experimental and conventional fluoride rinses on the in vitro and in vivo water-induced release of fluoride, calcium, phosphate, acetate and hydrogen ions from oral reservoirs was examined. At the start of each experiment, 13 subjects rinsed either with a conventional 228-ppm fluoride NaF rinse, a 228-ppm fluoride controlled-release rinse (CR rinse) or received no rinse. Sixty minutes later upper and lower molar plaque samples and 1-min saliva samples were collected. The subjects then rinsed with deionized water for 1 min, and 7 min later, a second set of samples was collected (in vivo study). Plaque fluid and clarified saliva were then recovered from samples by centrifugation, and the remaining plaque mass was sequentially extracted with water and acid to measure the water-extracted and total whole-plaque fluoride (in vitro study). All the samples were analyzed using microtechniques for pH, free calcium, phosphate, organic acids (plaque fluid) and fluoride (plaque fluid, centrifuged saliva and plaque extracts). Results showed that in vivo water rinsing decreased acetate and phosphate in plaque fluid, and fluoride in plaque fluid and saliva, but had no effect on plaque fluid pH. In vivo water rinsing, however, increased plaque fluid free calcium, apparently due to water-induced loss of calcium-binding ions. Water or fluoride-rinse-induced changes in plaque fluid concentration were greater at the lower molar site, suggesting that rinse pooling may influence ion distribution. Before the water rinse, plaque fluid, saliva and whole-plaque total fluoride values were 1.7, 2 and 4 times higher after the CR rinse compared to the NaF rinse. Furthermore, the CR rinse deposited approximately 11 times more water extracted fluoride compared to the NaF rinse, suggesting a 'more efficient' precipitation of 'labile' or 'loosely bound fluoride'. The results presented here, and in previous studies, suggest the possibility of formulating effective fluoride dentifrices with a lower fluoride content than is currently in use. PMID- 11275672 TI - Fluoride deposition in the aged human pineal gland. AB - The purpose was to discover whether fluoride (F) accumulates in the aged human pineal gland. The aims were to determine (a) F-concentrations of the pineal gland (wet), corresponding muscle (wet) and bone (ash); (b) calcium-concentration of the pineal. Pineal, muscle and bone were dissected from 11 aged cadavers and assayed for F using the HMDS-facilitated diffusion, F-ion-specific electrode method. Pineal calcium was determined using atomic absorption spectroscopy. Pineal and muscle contained 297+/-257 and 0.5+/-0.4 mg F/kg wet weight, respectively; bone contained 2,037+/-1,095 mg F/kg ash weight. The pineal contained 16,000+/-11,070 mg Ca/kg wet weight. There was a positive correlation between pineal F and pineal Ca (r = 0.73, p<0.02) but no correlation between pineal F and bone F. By old age, the pineal gland has readily accumulated F and its F/Ca ratio is higher than bone. PMID- 11275673 TI - Comparison of erythritol and xylitol saliva stimulants in the control of dental plaque and mutans streptococci. AB - The effect of 2-month usage of saliva-stimulating pastils containing either erythritol or xylitol was studied in a cohort of 30 subjects assigned to the respective polyol groups (n = 15). The daily consumption level of both polyols was 5.2 g, used in 5 daily chewing episodes. The mean weight of total plaque mass (collectable during a standard period of 3 min from all available tooth surfaces) was reduced significantly in the xylitol-group, while no such effect was observed in the erythritol-group. This reduction in plaque mass was accompanied by a significant reduction in the turbidity readings (A(660)) of aqueous plaque suspensions; no such effect was observed in the erythritol-group. However, plaque protein levels did not differ between baseline and endpoint in either polyol group. The plaque and salivary levels of Streptococcus mutans and plaque levels of total streptococci were reduced significantly in the xylitol-group, while no such effect was detected in the erythritol-group. However, either polyol regimen had no effect on plaque levels of S. sobrinus. The results suggest that systematic use of xylitol-containing saliva stimulants may be more effective in controlling some oral-hygiene-related and caries-associated parameters than similar use of erythritol-containing products. The results also speak for a special relationship between xylitol and S. mutans. However, owing to the great potential of erythritol as a caries-reducing agent -- based on the tetritol nature of erythritol -- the present laboratory results should be considered preliminary and subject to verifying clinical studies. PMID- 11275674 TI - In vitro studies of the penetration of adhesive resins into artificial caries like lesions. AB - Instead of removing the porous carious tissue at a relatively late stage in the disease process, attempts have been made to 'fill' the microporosities of lesions at a much earlier stage of lesion development. This would not only reduce the porosity and therefore access of acid and egress of dissolved material, but also afford some mechanical support to the tissue and perhaps inhibit further attack. Successful infiltration of materials into lesions has been demonstrated previously using resorcinol-formaldehyde which, however, was clinically unacceptable. The advent of dental adhesives with potentially suitable properties has prompted a re-examination of this concept. Artificial lesions of enamel were generated in extracted human teeth using acidified gels. A range of currently available adhesive materials was then used to infiltrate the porosities. The extent of occlusion of the lesion porosities was determined both qualitatively using light microscopy and quantitatively using a chloronaphthalene imbibition technique. The effect of such treatment upon subsequent exposure to acid gels was also investigated. Results showed that up to 60% of the lesion pore volume had been occluded following infiltration with some of the materials and that this treatment was capable of reducing further acid demineralization. The development of such treatment strategies could offer potential noninvasive means of treating early enamel lesions. PMID- 11275675 TI - Protective properties of salivary pellicles from two different intraoral sites on enamel erosion. AB - The purpose of this study was to investigate the protective effect and ultrastructure of salivary pellicles formed in vivo near the orifices of the ducts of parotid and submandibular/sublingual salivary glands. Pellicles were formed by exposing bovine enamel slabs to the oral environment at the buccal aspect of the upper first molars and at the lingual aspect of the lower incisors in 3 subjects over periods of 24 h. Enamel specimens with and without 24-hour pellicles were immersed in citric acid (0.1 and 1%) for periods ranging from 30 s to 5 min, and processed for measurement of surface microhardness (SMH) and transmission electron microscopy (TEM). In comparison to uncovered enamel specimen significantly less decrease in SMH due to acid exposure was observed in pellicle-coated enamel specimens. Pellicles formed at the buccal aspect of the upper molars were less effective in protecting the enamel against acid-induced softening as compared to pellicles formed at the lingual aspect of the lower incisors only after 5 min exposure in 1% citric acid. TEM analysis showed that pellicle layers were dissolved continuously due to acid exposure. However, even after 5 min exposure to 1% citric acid, a residual pellicle layer could be detected on the enamel surface. In conclusion, site-dependent differences of buccally and lingually in vivo formed 24-hour pellicles have minor importance concerning the pellicle-induced protection of the enamel surface against erosive changes. PMID- 11275676 TI - The extent of oral fungal flora in 353 students and possible relationships with dental caries. AB - An epidemiological study was conducted on 353 students to determine the potential relationships between oral saprophytism with Candida albicans and dental status. For each student included, an interview, a dental examination, a mycological investigation and determination of oral pH were conducted. Various factors liable to affect the presence of oral fungus were investigated using the chi(2) test. 58.6% of samples were positive when cultured, with C. albicans in 93.7% of cases. The mean DMF index was 7.6. C. albicans was more frequently isolated in men, smokers, when pH was lower than 7, when dental plaque was abundant and when the time since the teeth had last been brushed was more than 8 h. DMF and F indexes were greater when C. albicans was present but not when it was abundant, while decay was more often present in subjects with abundant C. albicans. Although the specific role of the various factors is difficult to establish, the results suggest that further research to elucidate the possible role of C. albicans in caries aetiology would be valuable. PMID- 11275677 TI - Differences in the salivary glycan pattern between children with high and low caries susceptibility. AB - Interactions between bacterial adhesins of lectin type and the oligosaccharide part of immobilised glycoconjugates on the tooth surface are involved in the specific colonisation of teeth. The specificity of the adhesion process is determined by the carbohydrate specificity of the bacterial lectins and the availability of the corresponding glycosylation pattern. On the other hand the same carbohydrate structures can specifically prevent the binding of bacteria by competitively blocking their adhesion, if sufficient amounts of this distinct carbohydrate structure are available in the secretion. Since carbohydrate binding receptors are also involved in the colonisation of tooth surfaces by cariogenic bacteria, it has been suggested that the architecture of the oligosaccharide portion of soluble glycoconjugates in saliva may play an important role as a constitutional host defence factor in the aetiology of dental caries. Characterising the availability of distinct carbohydrate patterns in saliva by using a pattern of well-described lectins in a competitive lectin inhibition assay we show that in children of a population-based sample a high caries susceptibility is associated with a reduced binding inhibition against the lectin peanut agglutinin (PNA). PNA is specific for the presence of terminal galactosyl residues and binds to the same O-glycan fractions as a surface lectin from Streptococcus mutans. The data suggest that a reduced availability of glycosylation patterns of galactosyl residues detected by the lectin PNA may act as an additional host-derived factor for an increased caries susceptibility. PMID- 11275678 TI - Na(+)/H(+)exchangers: linking osmotic dysequilibrium to modified cell function. AB - The Na(+)/H(+) exchangers (NHEs) are among the major ion transporters involved in cell volume regulation. NHE activation leads to a cellular influx of Na(+) ions and extrusion of H(+) ions, which are readily replenished from intracellular buffers. This will result in a net import of Na(+). In many systems NHE operates in parallel to Cl(-)/ HCO3(-) exchange, resulting in cellular uptake of NaCl. The influx of osmotically obliged water will consequently lead to cell swelling. This makes NHEs suitable to serve as powerful mechanisms for increasing cell volume (CV). The low volume threshold for NHE activation enables the cells to respond to very minute reductions of the CV. By the coupling to the export of H(+) ions cell volume regulatory NHE activation may lead to changes in intracellular pH. On the other hand NHEs are activated by a broad variety of ligands and by intracellular acidosis, which, in turn, may consequently lead to cell swelling. In addition, NHEs are linked to other intracellular proteins and structures, like e.g. the cytoskeleton, which themelves are involved in the regulation of numerous cellular processes. Therefore NHEs link CV regulation to a diversity of cellular functions, both in physiological and pathophysiological conditions. Six isoforms of the Na(+)/H(+) exchanger, termed NHE1--6, have been cloned so far. NHE 1--5 are located in the plasma membrane, whereas NHE6 is sorted to the mitochondrial membrane. NHE1 and NHE6 are the ubiquitously expressed isoforms. The expression of the isoforms NHE2 to NHE5 is restricted to specific tissues and the pattern of their expression, as well as their subcellular localization indicate that they fulfill specialized functions. Cell shrinkage induced activation has been shown for NHE1,2 and 4. In contrast, NHE3 is inhibited by cell shrinkage. In many cells several isoforms are present and assigned to specific membrane domains where they may serve a functional crosstalk between the different ion transporters. PMID- 11275680 TI - Neurofilament expression in cultured rat adenohypophysial cells. AB - The aim of the present work was to investigate in cultured rat adenohypophysial cells: a) the presence of neurofilaments of 200 kDa (NF-H), b) the effect of thyroid hormone (T(3)) and thyrotropin releasing hormone (TRH) on the expression of NF-H and c) the possible role of NF-H on thyrotropin (TSH) secretion. The presence of NF-H was observed by immunocytochemistry in cultured rat adenohypophysial cells. The exposure to T(3) for 12 h produced a significant increase in NF-H expression; whereas incubation with TRH or T(3)+TRH resulted in no change. The cells treated with T(3) or TRH or T(3)+TRH for 24 h showed no alteration. However, incubation for 48 h with TRH or T(3)+TRH caused significant decrease in NF-H expression. Incubation with NF-H antibodies produced a significant inhibition of calcium-induced TSH release in digitonin-permeabilized adenohypophysial cells. These results provide evidence that NF-H is present in cultured rat adenohypophysial cells, and that T(3) and TRH can modify NF-H expression. It can be suggested that in cultured adenohypophysial cells, NF-H may play a role in the secretory process. PMID- 11275679 TI - Differential transcription of ion transporters, NHE1, ATP1B1, NKCC1 in human peripheral blood lymphocytes activated to proliferation. AB - This work, using RT PCR, studied expression of mRNAs encoding ion transporters, the Na/H antiporter (NHE1), the beta subunit of the Na,K-ATPase pump (ATP1B1), the NaK2Cl symporter (NKCC1), and some proteins unrelated to ion transport: the serum and glucocorticoid dependent kinase (hSGK), beta-actin, a glycolytic enzyme (GAPDH), and regulators of proliferation and apoptosis (p53, Bcl-2) during activation of human lymphocytes with phytohemagglutinin for 4-24 h. Within 24 hours the mRNA levels of NHE1, beta-actin, Bcl-2, and p53 increased by more than 100%, the mRNA levels of ATP1B1, GAPDH, and hSGK, by about 50%, while the mRNA levels of NKCC1 decreased transiently. These results indicate a differential transcriptional control of NHE1, ATP1B1, and NKCC1 following a proliferative stimulus of human lymphocytes. PMID- 11275681 TI - Differential regulation of liver-specific and ubiquitously-expressed genes in primary rat hepatocytes by the extracellular matrix. AB - Primary rat hepatocytes were cultured with an extracellular matrix (ECM) overlay, in order to investigate the effect of an ECM on gene expression in hepatocytes. When hepatocytes, isolated by the collagenase-perfusion method, were cultured on type I collagen-coated dishes, the mRNA levels of liver-specific genes (aldolase B, tyrosine aminotransferase and albumin) decreased continuously, while those of ubiquitously-expressed genes (glyceraldehyde 3-phosphate dehydrogenase and beta actin) increased. When a dilute ECM derived from the Engelbreth-Holm-Swarm mouse sarcoma (an EHS gel) was added to the above hepatocytes 3 days after plating, the mRNA levels of liver-specific genes increased, while those of ubiquitously expressed genes decreased. The effects of a rat liver biomatrix (a physiological ECM for rat hepatocytes) on gene expression in primary hepatocytes were similar to those of the EHS gel. A nuclear run-on assay, and 5,6-dichloro-1-beta-d ribofuranosylbenzimidazole or actinomycin D treatments revealed that the transcriptional rates of liver-specific genes were enhanced by the EHS gel overlay, while the apparent stability of the corresponding mRNAs were unchanged. In contrast, the transcriptional rates of ubiquitously-expressed genes were not greatly affected by an EHS gel overlay, while the apparent stability of their mRNAs were decreased. These data suggest that the ECM plays an important role in the maintenance of the differentiated characteristics of primary hepatocytes by inducing the transcription of liver-specific genes and, also, by destabilizing the mRNAs of ubiquitously-expressed genes. PMID- 11275682 TI - Hypertonicity stimulates Cl(-) transport in the intestine of fresh water acclimated eel, Anguilla anguilla. AB - Eel intestinal epithelium when bathed symmetrically with normal Ringer solution develops a net Cl(-) current (short circuit current, Isc) giving rise to a negative transepithelial potential (Vt) at the basolateral side of the epithelium, lower in fresh-water (FW)-acclimated animals with respect to sea water (SW). The aim of the present work was to study the cell response to hypertonic stress of FW eel intestinal epithelium in relation to Cl(-) absorption. The hypertonicity of the external bathing solutions produced first a transient increase of Vt and Isc, then followed (after 10-15 min) by a gradual and sustained increase which reached the maximum value after 40-60 min. The morphometric analysis of the intestine revealed the shrinkage of the cells after 5 min hypertonicity exposure, and then a regulatory volume increase (RVI) response, which parallels the gradual and sustained increase in the electrophysiological parameters. This last phase is inhibited by drugs known to block Cl(-) absorption in eel intestine, such as luminal bumetanide (10 microM), specific inhibitor of Na(+)-K(+)-2Cl(-) cotransport, or basolateral NPPB (0.5 mM), dichloro-DPC (0.5 mM), inhibitors of basolateral Cl(-) conductance. Serosal dimethyl-amiloride (100 microM), specific inhibitor of the Na(+)/H(+) antiport, was ineffective on the hyperosmotic response. Bicarbonate revealed a crucial role as a modulator of hypertonicity response, since in bicarbonate-free conditions or in the presence of serosal 0.25 mM SITS, blocker of HCO(3)(-) transport systems, the Isc response to hypertonicity was lost. In nominally Ca(2+)-free conditions the Isc response to hypertonicity was abolished. The same results were obtained by bilateral addition of 100 microM verapamil or 50 microM nifedipine or 1 mM lanthanum, known Ca(2+) channel blockers, indicating that extracellular Ca(2+) plays a key role for the activation of Cl(-) current in the response to hypertonic stress. The data show that in the eel intestinal epithelium the hypertonicity of the external medium affects cell volume which in turn might represent the signal to increase the rate of Cl(-) transport. This response is sustained by the activation of the luminal Na(+)-K(+)-2Cl(-) cotransporter and the functionality of basolateral Cl(-) channels. PMID- 11275683 TI - Determination of protein-protein interactions of ICIn by the yeast two-hybrid system. AB - ICln is a ubiquitously expressed eukaryotic protein. Expression of the protein in Xenopus laevis oocytes, the knocking-down of the protein in fibroblasts, or the reconstitution of the protein in lipid bilayer led to the assumption that this protein is an ionic channel or a significant part thereof. However, other possible roles for ICln in potential regulatory mechanisms have been postulated, as diverse as regulator of cell morphology by interacting with the Skb1 protein and/or interaction with core spliceosomal proteins. Here we show that ICln is able to interact with SnRNP core proteins SmD1, SmD2, SmD3, SmX5 and SmB/B'. PMID- 11275684 TI - Mechanosensitive channels in prokaryotes. AB - Compared to voltage-dependent or ligand-gated ion channels that have extensively been studied over the last twenty years, there is little knowledge available on structure and function of mechanosensitive (MS) channels that constitute the third major group of ion channels classified according to their gating mechanism. The main purpose of this review is to summarize an area of the MS channel research where major progress has occurred. Cloning of MscL and MscS, the MS channels of Large and Small conductance found in Bacteria and elucidation of the 3D crystal structure of MscL are discussed in conjunction with the physiological role of the MS channels in bacterial osmoregulation. Furthermore, cloning and molecular characterization of MS channels in Archaea Methanococcus jannashii and Thermoplasma acidophilum are described. They present examples of the recent promising developments, which may ultimately lead to the understanding of the biophysical principles and evolutionary origins of mechanosensory transduction. Although they conduct ions and are usually characterized by their ionic conductance and selectivity, the MS channels in prokaryotes may primarily serve to transport osmoticants other than ions. Therefore, throughout this review prokaryotic MS ion channels are referred to as MS channels rather than MS ion channels. PMID- 11275685 TI - cGMP serves as an extracellular regulator of a Ca(2+)-dependent K(+) channel in immortalized human proximal tubule cells. AB - Recently we showed that a K(+) channel in immortalized human kidney epithelial (IHKE-1) cells derived from the proximal tubule is regulated by natriuretic peptides in cell-attached patches and directly regulated by cGMP in excised inside-out oriented membrane patches [1]. The patch clamp technique was used to investigate the regulatory effect of extracellular, non-membrane permeable cGMP on membrane voltage and the regulation of this K(+) channel in outside-out oriented membrane patches. In 7 out of 7 experiments the membrane voltage of IHKE 1 cells depolarized by 3.9 +/- 0.1 mV when the non-membrane permeable cGMP was added to the bath solution. In outside-out oriented membrane patches cGMP inhibited P(o) already at 1 microM (-12 +/- 4%, n=7), at 0.1 mM inhibition of P(o) reached -39 +/- 6% (n=14). cAMP (0.1 mM) only had a weak inhibitory effect (n=7). GTP and ATP (n=7 each) had no significant effect on P(o) from the outside. When cGMP was added to the pipette solution in experiments with outside-out oriented membranes cGMP still inhibited this K(+) channel from the outside by 36 +/- 6% (n=6). In 4 paired experiments 8-Br-cGMP (0.1 mM) showed a significantly higher inhibitory effect on P(o) compared to cGMP (0.1 mM). cGMP inhibits a K(+) channel in human proximal tubule cells from the outside and may serve as an autocrine and paracrine regulatory factor in the kidney. PMID- 11275686 TI - Inhibition of ryanodine binding to sarcoplasmic reticulum vesicles of cardiac muscle by Zn(2+) ions. AB - Using the assay of [(3)H]ryanodine binding to the sarcoplasmic reticulum, the effect of Zn(2+) on ryanodine receptors (RyRs) of cardiac muscle was investigated. There was no obvious change in the binding at [Zn(2+)](f) of less than 0.2 microM. However, a decrease of the binding became significant with raising [Zn(2+)](f) to 0.5 microM. The inhibitory effect of Zn(2+) was [Zn(2+)](f)-dependent, with IC(50/ZnI) of 2.1+/-0.4 microM (mean+/-S.D.). Scatchard analysis indicates that both an increase of K(d) and a decrease of B(max) were responsible for Zn(2+)-induced decrease of the binding. The Hill coefficient for this inhibitory effect of Zn(2+) was between 0.8 and 1.2. The interactions of the effects of Zn(2+) and various modulators of RyR indicate that the inhibitory effect of Zn(2+) was mostly mediated through inhibiting Ca(2+) activation sites (CaA) on RyR. Since the [Zn(2+)](f) dependence was not clearly changed by [Ca(2+)](f), the inhibitory effect of Zn(2+) may not be due to competition of Zn(2+) with Ca(2+) for CaA and probably is indirect. The inhibitory effect of Zn(2+) could not be antagonized by 2 mM dithiothreitol, a thiol-reducing agent, suggesting that the binding of Zn(2+) ions to RyRs of cardiac muscle is not accompanied by obvious change of redox state of the RyRs. In comparison with that seen in skeletal muscle [3], the effects of Zn(2+) on ryanodine binding to the sarcoplasmic reticulum of cardiac muscle show several distinct differences. It is indicated that the effect of Zn(2+) on RyRs may be isoform-dependent. The physiological significance of the effects of Zn(2+) is discussed. PMID- 11275687 TI - Functional enhancement of CFTR expression by mitomycin C. AB - Cystic fibrosis is caused by mutations in the CFTR gene. The most common of these mutations, DeltaF508, results in a protein that is not trafficked to the apical plasma membrane but instead is retained and degraded in the endoplasmic reticulum (ER) by the 26S proteosome. However, this protein is functional upon plasma membrane expression. It has been theoretically estimated that even a modest ( approximately 10%) increase in CFTR-associated chloride conductance can be beneficial in a clinical setting. Thus, understanding basic CFTR biogenesis is important, and identification of prototypical compounds that can increase CFTR expression and trafficking is potentially useful in the development of novel therapeutic strategies to treat cystic fibrosis. We report that mitomycin C (MMC) elicits such a response by increasing CFTR mRNA and protein expression in T-84 and HT-29 cells at very low, non-cytotoxic, pharmacologically relevant concentrations (0.1 microM) leading to enhanced chloride secretion. Thus, MMC may be a useful compound for understanding CFTR regulation and biogenesis. PMID- 11275688 TI - Volume--activated chloride currents in HeLa cells are blocked by tamoxifen but not by a membrane impermeant quaternary analogue. AB - BACKGROUND/AIMS: Tamoxifen has been shown to inhibit volume activated chloride currents in many cell types. Tamoxifen has also been reported to inhibit a number of cation channels as well as cytosolic proteins such as calmodulin. The mechanism of channel block by tamoxifen is not known but three hypotheses can be proposed: i) a direct effect following binding to the channel protein from the aqueous environment or ii) a direct effect on the channel protein after partitioning into the lipid membrane or iii) an indirect mechanism via binding to intracellular regulatory proteins after diffusion across the lipid membrane. The aim of these experiments was to distinguish between these hypotheses using membrane permeant and impermeant antioestrogens. METHODS: Volume activated chloride currents were recorded from single HeLa cells using whole cell patch clamp technique. The ability of tamoxifen and its membrane impermeant quaternary derivative ethyl bromide tamoxifen (EBT) to inhibit these currents was examined. RESULTS: Extracellular tamoxifen at 3 microM inhibited volume activated chloride currents in HeLa cells whereas EBT had no effect up to 10 microM when applied either to the extracellular bathing solution or the intracellular solution via the patch pipette. CONCLUSION: Eliminating the ability of tamoxifen to cross the plasma membrane abolishes its channel blocking activity against volume activated chloride channels in HeLa cells. PMID- 11275690 TI - Interleukin-6 signaling and embryonic mouse submandibular salivary gland morphogenesis. AB - Interleukin-6 (IL-6) is a multifunctional cytokine that mediates cell growth, differentiation, and survival. It was the objective of the present study to investigate the possible function(s) of IL-6 signaling in embryonic mouse submandibular salivary gland (SMG) morphogenesis. After characterizing in vivo mRNA and protein expression of various constituents of this pathway, we utilized in vitro strategies to investigate the phenotypic outcomes of enhanced IL-6 induced signaling and immunoperturbation of IL-6 binding to cognate receptors. These experiments demonstrate: (1) there is a significant increase of IL-6 mRNA with progressive SMG development, and that this is highly correlated with TNF transcript levels; (2) IL-6 and its cognate receptors are immunolocalized in SMG branching epithelia from the canalicular stage to the late terminal bud stage, as are other constituents of the IL-6 pathway; (3) as compared to controls, IL-6 supplemented explants exhibit a substantial increase in overall size and in the number of ductal branches and terminal buds, as well as a highly significant increase in epithelial cell proliferation; (4) SMG explants cultured in the presence of anti-IL-6 neutralizing antibodies exhibit a marked decrease in epithelial ducts and terminal buds, concomitant with a significant decline in cell proliferation and a highly significant increase in apoptosis. Taken together, our experimental results indicate that IL-6 signaling is important to SMG developmental homeostasis. PMID- 11275689 TI - Quantitative gene expression profiles of human liver-derived cell lines exposed to moderate hypoxia. AB - AIMS: To obtain better understanding of molecular events following critical oxygen shortage in liver tissue, we performed a large-scale comparison of gene expression profiles in four human liver cell lines, Chang, Hep3B, HuH7, and HepG2. METHODS: We used Atlas cDNA expression arrays from Clontech for initial screening, and quantitative real-time RT-PCR to assess the statistical significance of observed changes. RESULTS: RT-PCR analysis confirmed significantly changed levels of 24 transcripts after 24 hours incubation in 1% O(2). Among the genes most robustly up-regulated were plasminogen activator inhibitor-1 (PAI-1), insulin-like growth factor binding protein-3, and glyceraldehyde-3-phosphate dehydrogenase. Only PAI-1 was concordantly up regulated in all four cell lines. Conversely, most down-regulated were the stress response genes, including several heat shock proteins, yet only the expression of flap endonuclease-1 was significantly decreased in all cell lines. When comparing individual cell lines, the HepG2 cells displayed a pattern of general down regulation (83%), followed by Hep3B with 58% of genes down-regulated. In the Chang and HuH7 cells, however, the expression of most genes, 50% and 67%, respectively, remained unchanged. CONCLUSION: These studies provide information that is of importance for improved insight into the responses of not only liver tumor cells but also normal liver tissue in the conditions where physiological oxygenation is severely impaired. PMID- 11275692 TI - Granulated decidual cells in the mouse deciduoma: a putative source of decidual prolactin in mice. AB - Decidual cells are endometrial fibroblasts that redifferentiate during pregnancy in several species of mammals. In this work, we describe a subpopulation of resident decidual cells in the mouse endometrium that are joined by intercellular junctions and have cytoplasmic granules. Decidualization was induced in pseudopregnant mice on the 4th day of pseudopregnancy by injection of 30 microl of arachis oil into the uterine lumen. The uteri were collected on day 8 of pseudopregnancy (at 4 p.m., 8 p.m. and 11 p.m.) and on day 9 (at 8 a.m.). The tissues were fixed for light and electron microscopy. During day 8 of pseudopregnancy, granulated cells were present at the antimesometrial pole of the endometrium; they were concentrated at the periphery of the antimesometrial decidua and disappeared on day 9 of pseudopregnancy. The cytoplasm of the granulated decidual cells had acidophilic granules that stained also with periodic acid-Schiff (PAS). These granules stained with anti-rat prolactin antibody in both light and electron microscope immunocytochemical preparations. Vacuoles of various sizes were always present in the granulated cells. A PAS positive and prolactin-stained material was often deposited at the periphery of the vacuoles. Our results indicate that the granulated decidual cells are the source of decidual prolactin which accumulates in cytoplasmic granules. These granulated cells therefore form a transient gland in the mouse antimesometrial endometrium (granulated decidual gland). PMID- 11275691 TI - Morphology of isolated colonic crypts. AB - The traditional paradigm of colonic fluid and electrolyte transport includes a spatial separation of absorptive and secretory processes to surface and crypt cells, respectively. Recent studies of isolated microperfused colonic crypts revealed constitutive Na-dependent fluid absorption while secretion is regulated by one or more neurohumoral agonists. One obvious reason for the difference found in microdissected crypts is their separation from the lamina propria milieu. While it has been shown that isolated crypts are devoid of obvious lamina propria elements, including pericryptal fibroblasts, detailed morphologic information of the content of isolated crypts has been lacking. To characterize the morphology of the isolated crypt, we performed transmission electron microscopy (TEM) and immunofluorescence on microdissected and Ca2+ chelated crypts. Crypt cell type analysis was carried out separately on intact rat colon using light microscopy. TEM revealed a complete lack of either lamina propria cells or extracellular material in crypts isolated by either technique. TEM also revealed a subtle difference between the two isolation methods, with intact basal membranes in microdissected crypts but focal disruption of basal membranes in Ca2+- chelated crypts. Immunofluorescent stains for two basement membrane components (laminin and collagen type IV) revealed the presence of adherent basement membrane only on microdissected crypts; evidence that the plane of separation differs in these two preparations. Crypt cell type analysis on intact rat colon revealed an equal proportion of goblet cells in the right and left colon (approximately 50%) when measuring the middle 70% of the crypts - the area studied during crypt microperfusion. This morphologic analysis will increase our understanding of the observed physiology of isolated colonic crypts. PMID- 11275693 TI - Light and ultrastructural immunocytochemical study of prolactin cells in ovine adenohypophysis. Influence of lactation and weaning. AB - The influence of lactation and weaning on the number, distribution, and structure of mammotroph cells (PRL cells) in ewes was studied using immunocytochemical procedures for light and electron microscopy, as well as morphometric and stereological techniques. The adenohypophyseal gland of 12 ewes of the Segurena breed at different stages of milk production and weaning were analysed, while the same organ of 3 ewes in anestrus served as control. During lactation, PRL cells increased in number and decreased in size. Ultrastructurally, these cells showed a well-developed rough endoplasmic reticulum (RER) and Golgi complex (GC), and large secretory granules. These findings point to increased synthesis and secretion activities during lactation, and a decrease after weaning. One day after weaning 'storage' cells increased in number, and crinophagy became evident 7 days after weaning, when the first 'inactive' cells were seen (10% of total cells). In anestrus these last cells represented 40% of the total, and showed scarce granules, signs of inactivity, lysosomes and lipid droplets. PMID- 11275694 TI - Effect of undernutrition on cranial components and somatotroph-lactotroph pituitary populations in the squirrel monkey (Saimiri sciureus boliviensis). AB - The aim of the present study was to investigate in monkeys the effects of undernutrition on neurocranial and facial components, correlated with a histometric and ultrastructural analysis of somatotroph (growth hormone, GH) and lactotroph (prolactin, PRL) pituitary populations. Twenty Saimiri sciureus boliviensis (Cebidae) of both sexes were employed. The monkeys were born in captivity and when they reached 1 year of age, they were separated into two groups: control and undernourished animals. They were fed ad libitum a 20% and 10% protein diet, respectively. The monkeys were radiographed when they were 3 years old in order to measure the length, width and height of the anterior, middle and posterior components of the neurocranium, as well as those of the masticatory, respiratory and optic components of the face. The volumetric and morphometric indices were then calculated. After the sacrifice, pituitary glands were processed for light and electron microscopy. The quantitative immunohistochemistry revealed a decrease in the volume density and cell density of both GH and PRL cells from malnourished animals when compared to control ones. The ultrastructural study showed changes suggestive of cellular hyperfunction for both types of cells in the former experimental group. Under nutrition also affected the size of the cranial components, with males being more affected than females; brain weight was, however, nonmodified by stress, with the brain/body ratio difference being the same for both sexes. We conclude that in monkeys, experimental undernutrition produces a decrease in the pituitary GH and PRL cell populations, in some way related to changes in the cranio-facial morphometric patterns. PMID- 11275695 TI - Hair growth pattern in nude mice. AB - Nude mice are not bald but instead show an 'abortive' reduced hair growth on different sites of the integument. An albino (NMRI-nu) and a pigmented (C57BL/6 nu) strain of nude mice were examined as to whether the regional distribution pattern of this anagen hair proliferation is subject to the same ontogenetic development as in hairy mice. Hairy mice of both strains served as a comparison. Hair distribution was documented macroscopically by drawing and photography in a total of 415 mice of both sexes up to 421 days of age. Because of the pigmentation of the growing anagen hair follicles, the growth areas in the pigmented nude mice were distinctly visible whereas in the albino mice they were roughly recognisable from the boundaries of hair covering. The regional distribution of the 'abortive' anagen hair pattern in both nude strains corresponded to the wave-like course of the adult hair generations of hairy mice. As in older hairy mice, the hair cycle duration in nude mice was prolonged from an age of 121-180 days, the hair growth areas appeared reduced and less symmetrically orientated. Differences of up to 33% in body mass between the lighter nude and +/nu mice made ontogenetic comparison impossible so that all information is based on direct pattern or age comparison. The significance of experiments on the skin and hair follicles of nude mice is further increased if litters are examined comparatively and the temporal and spatial dimension of the follicle proliferation is considered more carefully than has been the case until now. PMID- 11275696 TI - Morphology of the laminar junction in relation to the shape of the hoof capsule and distal phalanx in adult horses (Equus caballus). AB - The purpose was to investigate whether differences in equine hoof shape, which are inferred to alter foot function, are accompanied by differences in morphology of the laminar junction. Ten fore feet from adult horses were segregated into normal and low-angle groups, depending on the dorsal angle of the hoof wall. Twenty measurements of external hoof shape and four of the enclosed distal phalanx were tested for differences between groups, and for intragroup correlations. Three measurements of laminar morphology (spacing, orientation and degree of bend) were recorded for samples of up to 50 primary epidermal laminae at each of 20 sample sites. Sites were distributed over the foot in 5 circumferential columns and 4 proximodistal rows. Intergroup differences were investigated, as were correlations among sample sites of the laminar variables with the shape measurements. Results show differences in hoof shape between groups (but not bone shape) and laminar morphology. Six shape measurements are significantly different between groups: dorsal angle, medial and lateral angles, lateral sole width, solar circumference, and dorsal length. In the normal group, shape measurements show patterns of correlation among regions of the hoof, and between hoof and bone measurements. In the low-angle group, shape correlations occur largely within one region of the hoof (the heels) and in the bone measurements. Laminar spacing tends to be nonsignificantly greater in the low angle group, while variances for laminar spacing and orientation are significantly greater in this group. Laminar spacing correlates with bone width and coronary circumference (CC) of the hoof in the normal group, but only with CC in the low-angle group. When taken as a whole, and interpreted in light of a model of foot mechanical function, the results appear to indicate a deterioration in structural coherence of the foot in the low-angle group. PMID- 11275697 TI - Scapular insertion of the rabbit latissimus dorsi muscle: gross anatomy and fibre type composition. AB - This paper defines the characteristics and significance of the scapular insertion of the latissimus dorsi muscle (LDM) of the rabbit. In a study of the New Zealand White species (n = 10) the scapular insertion was found to be a consistent anatomical feature of the LDM that made up 12.3% (+/-2.3) of the total muscle weight. The fibres arise from the medial aspect of the body of the LDM and run in a caudocranial direction to be inserted into a broad, thin tendon beneath the scapula ridge. This is morphologically different from the scapular component of the human LDM which is a well-recognized but inconsistent feature and consists of no more than a small leash of fibres running around the lower pole of the scapula. The scapular insertion was deeper red in colour than the body of the muscle and fibre-typing demonstrated a mean slow-fibre composition of 49% (+/ 2.6) compared to 16% (+/-1.7) for the body of the muscle (p < 0.01). Mapping of the fibre types throughout the remainder of the LDM confirmed that the body of the muscle was of fast phenotype but with significantly more slow fibres in the superomedial segment of the muscle than elsewhere. This region of the muscle contributes mainly to the scapular insertion and it is proposed that this part of the muscle takes on a predominantly postural role in stabilising the scapula during movement of the forelimb. PMID- 11275698 TI - Development-related expression of AKAP79 in the striatal compartments of the human brain. AB - The expression of AKAP79 which tethers regulatory proteins within postsynaptic densities has been studied in the two striatal compartments, i.e. patches and matrix, at different stages of the developing human brain by means of immunohistochemistry. The two striatal compartments exhibit various intensities of diffuse immunolabelling and a different number of immunoreactive nerve cells. From the 14th to 20th gestational week a nearly homogeneous distribution of immunoreactive structures in the two compartments of the striatum is seen. Thereafter, a decrease in immunoreactive structures within the matrix is observed (22nd-25th week, intermediate stage). From the 27th week onwards the patch compartment contains distinctly more immunoreactive puncta and nerve cells. Thus, the patches stand out clearly in the immunopreparations. This distribution pattern does not change during proceeding development. AKAP79-immunoreactive nerve cells closely resemble those constituting the class of medium-sized inhibitory projection neurons that receive the dopaminergic input of the striatum. Literature data suggest that AKAP79 may be functionally attributed to dopaminergic inputs. Accordingly, the patterns of AKAP79 expression can at least in part be correlated with the sequential occurrence of dopaminergic innervation. The mature matrix containing a dopaminergic innervation being as dense as in the patches displays distinctly less AKAP79-immunoreactive neurons and puncta than the patches. This discrepancy might indicate that a subpopulation of matrix neurons may, despite dopaminergic input, not express AKAP79. PMID- 11275699 TI - Somatostatin analogs for cancer treatment and diagnosis: an overview. AB - Due to the limited efficacy and considerable toxicity of conventional chemotherapy, novel cytotoxic agents and innovative noncytotoxic approaches to cancer treatment are being developed. Amongst the various hormonal agents, increasing attention is being directed to somatostatin analogs. This is largely due to the demonstration of antineoplastic activity of these compounds in a variety of experimental models in vitro and in vivo and to the elucidation of some aspects of the molecular mechanisms underlying their antineoplastic activity. On the other hand, clinical experience with somatostatin analogs in the treatment of conditions like acromegaly and GEP tumors has shown that they are well tolerated compared to other antineoplastic therapies currently in use. As a consequence, there is much ongoing clinical research to determine whether or not results from experimental studies will translate into clinically useful antineoplastic activity. Besides being used in cancer treatment and palliation, radiolabelled somatostatin analogs are employed for the localization of primary and metastatic tumors expressing somatostatin receptors. The so-called 'somatostatin receptor scintigraphy' is indeed the most important clinical diagnostic investigation for patients with suspected neuroendocrine tumors. Targeted radiotherapy, which is being evaluated in clinical trials, represents an obvious extension of somatostatin scintigraphy. Since the short half-life of native somatostatin makes continuous intravenous infusion mandatory, several long acting analogs have been synthesized. Amongst the hundreds of peptides synthesized, octreotide (which binds mainly to SSTR-2 and SSTR-5 receptor subtypes) has been the most extensively investigated. A thorough analysis of the pharmacological activities and therapeutic efficacy of the native somatostatin and the synthetic analogs (octreotide, lanreotide and vapreotide) reveals that the biological actions of these peptides are not always identical. These differences appear to be related to the different affinities of the natural hormone and synthetic derivatives for the different receptor subtypes. For all the three peptides long-lasting formulations have been developed to provide patients with the convenience of once or twice a month administration and to ensure stable drug serum concentrations between injections. Radiolabelled derivatives of octreotide, lanreotide and vapreotide have been synthesized and used as radiopharmaceuticals for somatostatin receptor scintigraphy and somatostatin receptor-targeted radiotherapy. The safety profile of synthetic somatostatin analogs is well established. Most adverse reactions to these peptides are merely a consequence of their pharmacological activity and consist mainly of gastrointestinal complaints, cholelithiasis and effects on glucose metabolism. They are often of little clinical relevance, thus making somatostatin analogs safe drugs for long-term use. While immediate release preparations are the drugs of choice in the short term, long-acting formulations are better indicated, on an outpatient basis, for the long-term management of chronic conditions. New 'receptor-selective' and 'universal' somatostatin analogs are being developed and combinations of currently available derivatives with other (cytotoxic and/or hormonal) agents are being explored in the search for an efficacious and well-tolerated treatment of the various malignancies. Somatostatin receptor-targeted chemotherapy (with conjugates of somatostatin peptides with cytotoxic drugs) and gene therapy (e.g. transferring the SSTR-2 gene into neoplastic cells), which have been successfully tested in experimental studies, should be applied to human beings in a not too distant future. PMID- 11275700 TI - Antiproliferative effect of somatostatin and analogs. AB - Over the past decade, antiproliferative effects of somatostatin and analogs have been reported in many somatostatin receptor-positive normal and tumor cell types. Regarding the molecular mechanisms involved, somatostatin or analogs mediate their action through both indirect and direct effects. Somatostatin acts through five somatostatin receptors (SSTR1-5) which are variably expressed in normal and tumor cells. These receptors regulate a variety of signal transduction pathways including inhibition of adenylate cyclase, regulation of ion channels, regulation of serine/threonine and tyrosine kinases and phosphatases. This review focuses on recent advances in biological mechanisms involved in the antineoplastic activity of somatostatin and analogs. PMID- 11275701 TI - Established clinical use of octreotide and lanreotide in oncology. AB - The diagnosis and treatment of neuroendocrine tumors have been significantly improved during the last decades. Localization and staging of the disease by somatostatin receptor scintigraphy (Octreoscan) are now the 'gold standard' for the management of these tumors. Treatment with somatostatin analogs has improved quality of life and possibly also survival for patients with neuroendocrine tumors. New long-acting formulations of the somatostatin analogs are as effective as the old regular formulations but will further improve quality of life for the patients. Tumor-targeted therapy with (111)In and (90)Y coupled to somatostatin analogs show promising results but await further studies. PMID- 11275702 TI - The palliative effects of octreotide in cancer patients. AB - Octreotide is an extremely useful compound for palliative care physicians. It appears to be active in a number of different pain states and may be given by the spinal and intraventricular route. Its actions in reducing gut motility and secretions make it a valuable adjunct in the management of inoperable bowel obstruction. The same actions make it a potent antidiarrheal agent. Octreotide will often succeed where other antidiarrheal agents fail. Its ability to reduce gut secretions has led to its use in the treatment of fistulae. It has also been proposed as a useful drug in the management of cachexia and ascites. Most of the existing evidence is based on small numbers of case reports and further larger trials are necessary. PMID- 11275703 TI - Management of breast cancer: is there a role for somatostatin and its analogs? AB - Somatostatin and related peptides are a family of peptides which are ubiquitous and function as endogenous growth inhibitors. Analogs have been developed through the introduction of a D-amino acid in the position 8 of somatostatin moiety which is more resistant to the action of endogenous peptidases than the parental moiety. Both somatostatin and its analogs interact with specific receptors on the cell surface. The five receptor subtypes, SSTR-1 to SSTR-5, which have been characterized so far, have a different affinity for somatostatin and its analogs. This and the fact that receptors are not homogeneously expressed in tissues account for the different activity of these compounds, all of which have demonstrated tumoristatic properties both in vitro and in vivo. The interaction of somatostatin and of somatostatin analogs with specific SSTR receptors is crucial to the antiproliferative mechanisms exerted by these compounds in vitro and in some animal models and the various pathways have been reviewed in detail. However, inhibition of angiogenesis and suppression of lactogenic hormones might represent alternative mechanisms, in particular in breast cancer. The rationale for the use of somatostatin and its analogs in breast cancer patients and to combine these peptides with antihormones, like antiestrogens or prolactin lowering drugs, or cytotoxics has been reviewed together with the results obtained in phase II and comparative trials. The reasons for the limited efficacy shown by these compounds either when used alone or when used in combination with other drugs have also been critically reviewed in the perspective of new trials. PMID- 11275704 TI - Somatostatin, its receptors and analogs, in lung cancer. AB - Despite developments in diagnosis and treatment, lung cancer is the commonest cause of cancer death in Europe and North America. Due to increasing cigarette consumption, the incidence of the disease and resultant mortality is rising dramatically in women. Novel approaches to the management of lung cancer are urgently required. Somatostatin is a tetradecapeptide first identified in the pituitary and subsequently throughout the body particularly in neuroendocrine cells of the pancreas and gastrointestinal tract and the nervous system. The peptide has numerous functions including inhibition of hormone release, immunomodulation and neurotransmission and is an endogenous inhibitor of cell proliferation and angiogenesis. Somatostatin and its analogs, including octreotide (SMS 201-995), somatuline (BIM 23014) and vapreotide (RC-160), act by binding to specific somatostatin receptors (SSTR) of which there are 5 principal subtypes, SSTR-1-5. Although elevated plasma somatostatin levels may be detected in 14-15% of patients, tumor cell expression appears rare. SSTR may be expressed by lung tumors, particularly small cell lung cancer and bronchial carcinoid disease. [(111)In]pentetreotide scintigraphy may have a role to play in the localization and staging of lung cancers both before and following treatment, and in detecting relapsed disease. The potential role of radiolabelled somatostatin analogs as radiotherapeutic agents in the management of lung cancer is currently being explored. Somatostatin analog therapy results in significant growth inhibition of both SSTR-positive and SSTR-negative lung tumors in vivo. Recent work indicates that these agents may enhance the efficacy of chemotherapeutic agents in the treatment of solid tumors including lung cancer. PMID- 11275705 TI - Octreotide in the management of hormone-refractory prostate cancer. AB - Patients with advanced or metastatic prostate cancer (PC), a partially hormone resistant disease, will require some form of hormonal manipulations or some new therapeutic modalities. Octreotide, as somatostatin (SST) analogs, has been found to inhibit the growth of experimental PCs via several mechanisms, as indirect antihormonal and direct antimitogenic actions, mainly due to inhibition of SST receptor subtypes (SSTR-1-5). Sporadic clinical trials with octreotide (alone or with a complete antiandrogen blockade) treatment of patients with advanced stage D2 PC demonstrated promising results. Unfortunately, at present these clinical trials have some disadvantages and leave some uncertainty with regard to the trial design, the SSTR subtype determination and tumor localization with SSTR scintigraphy before the start of a selective SST analog, and finally the randomization in groups according to hormone resistance, dosage regimen and route of administration. PMID- 11275706 TI - Gastrointestinal cancer refractory to chemotherapy: a role for octreotide? AB - Although octreotide has been shown to inhibit the growth of gastrointestinal (GI) tumors in vitro and in vivo, preliminary clinical trials have reported disappointing results for this somatostatin analog in patients with GI cancers. The results of these trials probably reflect the difficulty in assessing the therapeutic potential of an agent such as octreotide in GI cancers. Thus, it is possible that treatment with octreotide could be useful in the stabilization of disease if it is associated with an improvement in survival. On the basis of these considerations five randomized trials were carried out to evaluate the therapeutic potential in patients with GI cancers. Four trials (one in patients with colorectal carcinoma and three in patients with carcinoma of the pancreas) did not show any advantage of octreotide in untreated patients; in contrast, one trial reported that octreotide prolonged survival in patients with GI cancers refractory to chemotherapy. Some clinical features of the latter study (treatment with chemotherapy, different schedules) may explain these conflicting results. Although data from randomized trials suggest that octreotide is not effective in untreated asymptomatic advanced GI cancer patients, further studies are warranted to assess the efficacy of octreotide in chemotherapy refractory patients in order to clarify the impact of octreotide in terms of not only survival but also on the patients' quality of life. PMID- 11275707 TI - Pancreatic cancer: does octreotide offer any promise? AB - The incidence of adenocarcinoma of the pancreas has risen steadily over the past 4 decades. Since pancreatic cancer is diagnosed at an advanced stage, and because of the lack of effective therapies the prognosis of such patients is extremely poor. Despite advances in our understanding of the molecular biology of pancreatic cancer, the systemic treatment of this disease remains unsatisfactory. Systemic chemotherapy and the administration of biologically active molecules such as tumor necrosis factor or interferons have not resulted in significant improvements in response rates or patient survival. New treatment strategies are obviously needed. This paper will discuss current advances in the use of somatostatin analogs in the management of pancreatic cancer. PMID- 11275708 TI - Octreotide for cancer of the liver and biliary tree. AB - Inoperable liver tumors have an unfavorable natural course despite various therapeutic modalities. Octreotide, a somatostatin analog, has shown considerable antitumor activity on animal models of various hepatic tumors and on isolated cell culture lines. In this paper, a review of the experimental evidence is presented. Moreover clinical papers of case reports of uncontrolled studies of patients are also reviewed. The majority of clinical studies provide evidence of a clinical and biochemical response of liver endocrine tumors while regression of tumor size is a rare event. A randomized controlled trial of octreotide in the treatment of advanced hepatocellular carcinoma has shown a significant survival benefit in the treated patients. Literature reports indicate a stimulatory effect of octreotide on Kupffer cells as a possible antitumor mechanism, but other antiproliferative actions of octreotide have been suggested but not proved. Finally the question of the presence and affinity of somatostatin receptors on liver tumor tissue is discussed. In conclusion, according to our experience, octreotide administration is the best available treatment for advanced inoperable hepatocellular carcinoma and future better patient selection, based on receptor subtypes, might further improve the results. PMID- 11275709 TI - Somatostatin analogs in oncology: a look to the future. AB - In the past 15 years considerable advances have been made in our understanding of the molecular pharmacology of the mechanisms whereby somatostatin and its analogs mediate their direct and indirect antineoplastic effects. However, some important issues remain to be resolved, in particular the functional roles of the individual somatostatin receptors (SSTR-1-5) in tumor tissue and up- or downregulation of the hSSTRs with prolonged administration of somatostatin analogs. Answers to these questions are essential before we can maximize the therapeutic efficacy of somatostatin analogs in cancer. For example, is continuous administration more or less effective than intermittent therapy? The role of somatostatin analogs in the management of acromegaly and to a lesser extent neuroendocrine tumors is firmly established. The development of depot preparations of all 3 somatostatin analogs currently available for clinical use will undoubtedly improve both patient compliance and quality of life in patients with these conditions. There are only likely to be minor differences in the therapeutic efficacy of octreotide, lanreotide and vapreotide since all three analogs exert the majority of their antineoplastic effects via hSSTR-2 and hSSTR 5 and at the end of the day, price may well dictate which of these drugs oncologists use to provide symptomatic palliation of acromegaly and neuroendocrine tumors. Apart from some notable exceptions, somatostatin analog therapy has proven to be very disappointing in the management of advanced malignancy. Improvements in the management of solid tumors are likely to come only from combination therapy of somatostatin analogs with cytotoxic agents or other hormones in both advanced malignancy and in the adjuvant setting. Clinical trials with clear-cut objective outcome measures and health-related quality of life assessment are needed to evaluate the therapeutic efficacy of combination treatment in advanced malignancy and as an adjuvant to surgery. Particular attention needs to be paid to possible adverse effects of somatostatin analog therapy on the immune response to cancer. Further studies are required to establish whether the adverse effects of somatostatin analog therapy alone or in combination with cytotoxics or other hormones can be reversed with appropriate immunomodulatory treatment. Targeted somatostatin analog radiotherapy and chemotherapy are currently being investigated and the results of these studies are awaited with interest. Novel approaches using combinations of somatostatin analogs with antiangiogenic drugs or gene therapy are of particular interest and may provide important advances in the management of cancer in the not too distant future. PMID- 11275712 TI - Renal colic: pathophysiology, diagnosis and treatment. PMID- 11275714 TI - Management and aetiology of stones in intestinal urinary reservoirs in adolescents. AB - OBJECTIVE: To review the aetiology and management of reservoir stones in patients with intestinal urinary reservoirs. SUBJECTS AND METHODS: Since 1983 patients with enterocystoplasty have been followed prospectively by protocol. The data sets and notes of 148 patients reconstructed for congenital anomalies were reviewed to retrieve information on the incidence, management and aetiology of reservoir stones. RESULTS: Data were complete on 146 patients, 2 others having been lost to follow-up. Mean follow-up was 3.4 (range 1-14) years. Twenty-three patients formed stones (15.8%). Mean time to stone formation was 45 months (range 1 month to 10 years). In 13 patients the stones were removed by a percutaneous approach. In 9 patients with large stones (>5 cm) an open removal was performed. One patient had a small stone removed through a Kock nipple. All stones were struvite on analysis. All patients with an augmented bladder drained by a supra pubic Mitrofanoff formed stones at some time. The incidence of stones in other groups was: Kock pouch 50%; reservoirs drained by urethral catheterisation 9%; all other abdominal reservoirs 7.5%. No patient who voided spontaneously formed stones. CONCLUSION: Reservoir stones are infective in composition. The incidence is strongly related to the lack of downward, gravitational emptying. Stones up to 5 cm can be removed percutaneously. PMID- 11275713 TI - Investigation of upper tracts after resolution of symptoms due to ureteric calculi. AB - OBJECTIVE: To determine whether patients with proven ureteric calculi on IVU require repeat IVU after resolution of symptoms and passage of calculus on plain X-ray. METHODOLOGY: IVU reports for a 12-month period were obtained and notes and X-rays of those patients with ureteric calculi were reviewed. Presentation, management and subsequent imaging after resolution of symptoms were determined for each patient. All X-rays were reviewed by a uroradiologist. RESULTS: Fifty eight patients were investigated for the study. All initial IVUs showed upper tract dilation or obstruction. Forty-three eventually passed their calculi spontaneously and of these, 18 had KUB, all of which showed passage of the calculus and 25 had repeat IVU, 22 of which were normal. The 3 abnormal IVUs showed persisting calculi which were visible on the plain film. Fifteen patients required surgical intervention and all had repeat IVU, of which 5 were abnormal. CONCLUSION: This study suggests that following resolution of symptoms due to ureteric colic, patients who pass their calculi spontaneously can be followed up by KUB. Only those with persistent calculi on KUB or those who have had surgical intervention require repeat IVU. PMID- 11275715 TI - Symptomatic physiologic hydronephrosis in pregnancy: incidence, complications and treatment. AB - OBJECTIVE: We present the incidence and results of treatment of symptomatic physiologic hydronephrosis in 3,400 pregnant women. METHODS: We analyzed 103 consecutive women who presented with clinical signs and symptoms related to the upper urinary system. Renal sonography, urinalysis, serum creatinine levels, white blood cell (WBC) count, and urine culture were done in all patients at first visit and repeated at least once a month until 1 month after delivery. In patients who manifested acute pyelonephritis, urinalysis, WBC count, erythrocyte sedimentation rate and C-reactive protein levels were repeated every 3 days until normalization, and urine culture as well as renal sonography were performed once a week until 1 month after delivery. Conservative measures (positioning, analgesia, antibiotics) were performed in all patients with symptomatic physiologic hydronephrosis. If the patient's condition was refractory to medical management, drainage of the ureter with a double pigtail stent was performed. RESULTS: Conservative measures were successful in 97 (94%) of 103 patients but 6 (6%) patients had ongoing signs and symptoms of acute pyelonephritis progressing to urosepsis. In all of them, antibiotics were continued and a double pigtail stent was placed resulting in fast regression of symptoms, curing of renal infection and progress of the pregnancies to the term with vaginal delivery. CONCLUSIONS: Symptomatic hydronephrosis in pregnancy can be treated conservatively. If the patient's condition is refractory to medical management, an internal drainage with double pigtail stent may be necessary. PMID- 11275716 TI - Bulbar urethral stricture repair with buccal mucosa graft urethroplasty. AB - OBJECTIVES: Evaluation of the use of buccal mucosa graft as single-stage urethral reconstruction in an adult population with a stenosis of the bulbar urethra. METHODS: In our Department between April 1996 and February 1999, 20 patients with bulbar urethra stenosis underwent single-stage urethroplasty using a buccal mucosa graft. Mean age of patients was 52 years (range 14-70). The etiology of urethral stricture was inflammation (4 cases), iatrogenic (5 cases) and idiopathic (11 cases). A ventral onlay patch (mean length 3.6 cm, range 2.5-5) was employed in all cases. RESULTS: During the follow-up (median 13 months, range 6-28) the overall success rate was 80%. The success rate was 75% for inflammatory strictures, 80% for iatrogenic strictures and 81% for strictures of unknown etiology. CONCLUSIONS: Although longer follow-up is needed, free graft urethroplasty with buccal mucosa graft represents a simple surgical option which has produced encouraging results. This is probably due to the quality of the tissue employed which at present seems to represent the first-choice solution in selected cases. PMID- 11275717 TI - Johanson's staged urethroplasty revisited in the salvage treatment of 68 complex urethral stricture patients: presentation of total urethroplasty. AB - BACKGROUND: To show our experience with the staged Johanson's urethroplasty as a salvage treatment of difficult and complicated groups of patients, and to present the total urethroplasty technique. MATERIAL AND METHODS: During a 12-year period, 68 men with urethral stricture underwent the staged Johanson's urethroplasty. 51 had war-related injuries (75%) resulting in an unhealthy perineal and genital skin with fistulae and/or scarring. 35 patients (52%) had other urethral or vesical problems. 60 patients (88%) had long (0.5-4 cm), multiple or impassable strictures. 58 patients (85%) had strictures of the pendulous urethra. The second stage was performed 2-3 months after the first. Both stages of Johanson's urethroplasty were protected by a stab suprapubic catheter for 3 weeks. Patients were followed up for 23-82 months (mean 52.5). RESULTS: All patients but 4 had improved urine flow (best Qmax ranged between 13.2 and 31.8 ml/s; mean 17.4). 4 patients (6%) needed a revision because of fistula formation or recurrence and 6 patients (9%) developed urinary tract infection postoperatively. CONCLUSIONS: The staged Johanson's urethroplasty is a good treatment for the difficult and complicated urethral strictures which are not suitable for optical urethrotomy, especially those in the pendulous part. In strictures involving all parts of the urethra total urethroplasty could be performed. PMID- 11275718 TI - Incidence and risk factors of bacteriuria after transurethral resection of the prostate. AB - INTRODUCTION: Postoperative bacteriuria is a frequent event after transurethral resection of the prostate, despite the use of prophylactic antibiotics. Certain risk factors have been clearly established (preoperative urinary catheter or bacteriuria, operating time), while others remain uncertain. MATERIALS AND METHODS: We conducted a prospective study in five urology centers, including non catheterized patients with sterile preoperative urine undergoing transurethral resection of the prostate for benign prostatic hyperplasia. All received antibiotic prophylaxis with cefamandole. The incidence of bacteriuria and its risk factors were investigated. RESULTS: The postoperative bacteriuria rate was 26% (26/101), with 8% on removal of the catheter, 14% between the 7th and 10th postoperative days and 5% 1 month postoperatively. Factors associated with bacteriuria on univariate analysis were: operating time, disconnection of the closed urine drainage system and postoperative catheterization > or =3 days. Two variables were associated on multivariate analysis (logistic regression): operating time >52 min (odds ratio 9.0, 95% confidence interval 2.1-39.0) and disconnection of the closed urine drainage system (odds ratio 26.3, 95% confidence interval 6.1; 6.1-113). CONCLUSIONS: The postoperative bacteriuria rate after transurethral resection of the prostate was high in this study, raising the question of the choice and/or duration of prophylactic antibiotics. Prevention of postoperative bacteriuria must be based on careful hemostasis, prevention of postoperative catheter disconnections, and limitation of the duration of postoperative catheterization. PMID- 11275719 TI - Immunocytological analysis of leukocyte subpopulations in urine specimens before and after prostatic massage. AB - OBJECTIVE: To evaluate the presence of leukocyte subpopulations in urine after prostatic massage (VB 3) in symptomatic patients with > or =10 leukocytes/high power field (magnification x1,000) in expressed prostatic secretions, and who were classified as suffering from chronic bacterial prostatitis or inflammatory chronic pelvic pain syndrome. METHODS: 115 consecutive patients were investigated. Granulocytes in centrifuged midstream urine (VB 2) and VB 3 were counted after Papanicolaou stain. Macrophages, B and T lymphocytes were analyzed after immunocytological staining with monoclonal antibodies according to the alkaline phosphatase anti-alkaline phosphatase method. The counts were quantified as the number of cells per view field at a magnification of x400. In all patients, acute or chronic urethritis had been excluded before enrollment in the study. 16 men without signs or symptoms of urogenital inflammation served as controls. RESULTS: Of the 115 patients, 101 men demonstrated > or =10 leukocytes/view field in VB 3. In comparison to VB 2, the leukocyte subpopulations in VB 3 demonstrated an increase in granulocytes (9.2-fold), macrophages (7.6-fold), T lymphocytes (7.6-fold), and B lymphocytes (4-fold). The increase was statistically significant (p<0.001 each). The proportion of these cells in VB 3 was 81.6, 11.1, 5.5, and 1.8%, respectively. As compared to controls, all leukocyte subsets in VB 3 were significantly elevated (p>0.001 each). CONCLUSION: Elevated numbers of leukocytes in VB 3 are indicative of prostatitis provided that urethral inflammation and leukocyturia in VB 2 are excluded. Granulocytes are the predominant cell type of inflammation. The increase in macrophages, T and B lymphocytes in prostatic secretions indicate the participation of both the cellular and humoral immune system in the inflammatory process. PMID- 11275720 TI - Is it necessary to perform urine cytology in screening patients with haematuria? AB - All patients with gross haematuria and those older than 50 years with microscopic haematuria need investigations to rule out the presence of a urological malignancy. OBJECTIVE: To study the role of urine cytology in the evaluation of patients with haematuria. METHODS: Two hundred and eighty-five patients were evaluated. All patients underwent evaluation including urine cytology, flexible cystoscopy, ultrasonography and/or IVU. RESULTS: The mean age of the patients was 62.4 years. Sixty-five percent had gross and 35% microscopic haematuria. Fifty five tumours were discovered (19.2%); of these 48 were transitional cell carcinomas, 3 renal cell carcinomas and 3 carcinomas of the prostate. Thirty seven urinary cytologies were abnormal. The overall sensitivity of urinary cytology was 42.4% and specificity 94.3%. Of 18 patients with positive cytology, all were found to have transitional cell carcinomas on cystoscopy or imaging. Of 19 patients with suspicious cytologies, only 6 were found to have tumours. The remaining 13 patients had no evidence of tumour on combined upper tract imaging (IVU and ultrasound) or on rigid cystoscopy and bladder biopsy. Whilst all the other investigation modalities contributed to diagnoses (and/or exclusion of tumours), no additional tumours were discovered solely by urinary cytology. A moderate cost saving could be made without compromising diagnostic accuracy. CONCLUSION: Our study suggests that performing routine urine cytology is not relevant in the investigation of patients with haematuria, its role is at best supportive. PMID- 11275721 TI - Assessment of fibrin-fibrinogen degradation products (Accu-Dx) test in bladder cancer patients. AB - OBJECTIVES: Measurement of urine fibrin-fibrinogen degradation products has been reported to be a useful marker for bladder malignancies. This is a simple, noninvasive and rapid method in the diagnosis and follow-up of bladder cancer. We performed a prospective study to evaluate the reliability of the Accu-Dx test which detects urinary fibrin and fibrinogen products associated with bladder cancer. MATERIAL AND METHODS: 97 patients were included in this study. 55 patients with bladder cancer were under surveillance, and 35 were evaluated as primary cases. The Accu-Dx test was performed in 7 additional patients presenting with hematuria due to benign disorders. Urine cytology specimens were collected in all, whereas bladder pathology specimens were obtained in 93. The Accu-Dx test was evaluated with regard to the results of urine cytology and pathological assessment, which constituted the gold standard. RESULTS: According to pathology, 69 patients had carcinoma of the bladder. In 48 (69.6%), the Accu-Dx test was positive whereas urine cytology was positive in only 31 (44.9%; p<0.001). Specificity rates were 67.9 and 96.4%, respectively. With higher stages and higher tumor grades, the Accu-Dx test yielded higher positive rates (50% in Ta versus 100% in T2 or higher and 42.9% in G1 versus 94.1% in G3). On the other hand, the test was positive in 6 patients with cystitis of various etiologies. The overall accuracy rate for Accu-Dx was 69%. CONCLUSIONS: The Accu-Dx test is a simple, rapid, reproducible and more importantly a noninvasive method in the detection of bladder malignancies. But it seems inadequate to replace conventional cystoscopy plus pathologic assessment due to its relatively low accuracy and specificity rates. It may be helpful in selecting patients who should be evaluated with rigid endoscopes together with bladder biopsies. PMID- 11275722 TI - Significance of tissue laminin P(1) elastase and fibronectin levels in transitional cell carcinoma of the bladder. AB - OBJECTIVE: Elastase is a serine protease which hydrolyses connective tissue components. Laminin and fibronectin also play an important role in progression and invasion of cancer. The purpose of this study is to investigate the relation between tissue elastase, laminin P(1) and fibronectin levels and tumor characteristics, and analyze the potential of these as prognostic factors in transitional cell carcinoma (TCC) of the bladder. METHODS: Thirty-four patients with TCC of the bladder and 11 controls were included in this study. Elastase and fibronectin levels in tissue homogenates were determined using an enzyme immunoassay and laminin P(1) by radioimmunoassay. Mean follow-up was 43 months. RESULTS: The mean elastase level in bladder carcinoma tissue was 120+/-11.42 ng/homogenate protein, while normal tissue level was 12.36+/-2.70 (p<0.01). The carcinoma and normal tissue mean laminin P(1) levels were 7.02+/-0.37 U and 0.65+/-0.10 U/mg homogenate protein, respectively (p<0.01). The mean fibronectin level was 19.97+/-1.45 ng/mg homogenate protein in the carcinoma tissue and 2.16+/-0.40 in normal tissue (p<0.01). There was no correlation between tumor stage, grade, size, multiplicity and elastase, laminin P(1) and fibronectin levels. CONCLUSION: These results provide evidence that tissue elastase, laminin P(1) and fibronectin levels increase in TCC of the human bladder. Further studies including serum and urine levels should be performed in order to analyze their value as tumor markers in a larger group of patients. PMID- 11275723 TI - Renal tumor size: comparison between computed tomography and surgical measurements. AB - OBJECTIVE: We studied the agreement between renal tumor size as assessed on computed tomography (CT) before surgery and that measured during histopathological examination on the radical nephrectomy specimen. METHODS: We retrospectively analyzed the records of 100 consecutive patients treated with radical nephrectomy for a renal tumor. The tumor size was determined in all patients by the largest diameter shown within the month before surgery on contrast-enhanced CT and as measured postoperatively by the pathologist. A possible influence of the clinical and pathological parameters was assessed in a multivariate analysis. RESULTS: CT estimate and surgical measurement of tumor size were highly correlated (r = 0.9; p<0.001). Median (range) tumor size was 70.0 mm (13-180) and 60.0 mm (10-180) as measured, respectively, on CT and in the specimen, with a significant difference (p = 0.005). Multiple regression did not reveal any significant influence of tumor side, location, type, nuclear grade as well as patient gender, body mass index and radiological center (p>0.3 in all cases). The extent of difference between CT and surgical measurements was significantly influenced by the surgical size of the tumor (p = 0.03): the smaller the tumor, the more the CT overestimated the tumor size. If nephron sparing surgery had been planned for tumors equal to or less than 40 mm, 24 patients would have been selected following the CT estimate, while 27 patients would have met this criterion on the surgical measurement. CONCLUSION: Renal tumors were statistically smaller than the estimate from CT, although this was not systematically the case. This should be kept in mind when issuing recommendations on the optimal cutoff size value under which nephron-sparing surgery is considered equivalent to radical nephrectomy. PMID- 11275724 TI - Endoscopic classification and endourologic therapy in proximal incomplete ureteral duplication pathology. AB - OBJECTIVE: To assess whether patients with proximal incomplete ureteral duplication (PIUD) had functional or anatomical Y junction zone pathology. In such cases, stasis and infection, with or without associated renal scarring, are often found. The results after minimally invasive treatment (retrograde endoincision of ureteral stenosis, localized on one limb) can be evaluated. PATIENTS AND METHODS: In a retrospective analysis of 807 ureteroscopies, we found 29 cases with PIUD pathology dating from July 1996 to March 1999. The investigative protocol involved: intravenous pyelography (IVP) with delayed films, voiding cystourethrogram, and duplex Doppler renal ultrasonography (DDRU) for the resistive index. We used semirigid ureteroscopes (7.5 and 8.5 french) and the Storz cold knife/scissor. The mean follow-up period was 17 months (range 5-33 months). RESULTS: In 22 cases (76%) we found functional obstruction of the Y junction region and in 7 cases (24%) a ureteral stricture shorter than 1 cm of lower or upper limbs. We described four particular morphologies of the Y junction zone: double-barrelled (wide bifid limbs) with larger Y junction, lateral insertion, punctiform insertion and valvular insertion of one of the limbs. The first type is associated with functional obstruction and the others are characteristic of anatomic obstruction. After retrograde cold endoincision, normal PIUD was found in 85% of cases. CONCLUSIONS: Pathological aspects of PIUD may pose interesting problems in diagnosis and therapy. Endoscopic particularities of the Y junction zone could be correlated with IVP and DDRU. Retrograde endoureterotomy appears to be an effective minimally invasive procedure for treatment of anatomical obstruction where the ureteral stenosis of one limb is shorter than 1 cm. PMID- 11275725 TI - Antegrade percutaneous endoluminal treatment of non-malignant ureterointestinal anastomotic strictures following urinary diversion. AB - OBJECTIVE: We report our experience on antegrade percutaneous incision of ureterointestinal anastomosis strictures after urinary diversion. MATERIALS AND METHODS: Since 1994, we have evaluated retrospectively 18 patients with 22 ureterointestinal anastomosis strictures (UAS), who were treated with cold-knife incision. After placement of an 8-french nephrostomy tube, a 0.035-inch guide wire bypassed the stricture under guidance of a centrally opened (5-french) ureter catheter. A wire-mounted cold-knife was pulled through the strictured area retrogradely under fluoroscopic control. Routinely, following the incision, an 8 french external stent was left in place for 6-8 weeks. RESULTS: After stent removal as a primary procedure, the ureteroenteric area has remained patent in 14 of 19 (74%) UAS. In 3 cases undergoing a secondary or repeated procedure, treatment failed. The average follow-up was 23.5 (range 12-39) months. Failures were associated with radiogenic injury of the ureter in 5 UAS and unexplained in 2. No complication was observed. CONCLUSION: Percutaneous endourological management of UAS with the cold-knife incision, when used as a primary treatment, is a safe and effective alternative to open surgical repair and should be considered as an initial approach. PMID- 11275726 TI - Color doppler ultrasonography and spectral analysis of venous flow in diagnosis of varicocele. AB - OBJECTIVE: The standardization of diagnostic criteria for varicocele has not yet been established. This causes difficulty in evaluating both the incidence and clinical studies. Our aim was to establish diagnostic criteria for varicocele in Doppler procedures. METHODS: The characteristics of blood flow in the internal spermatic vein were investigated with color Doppler ultrasonography (CDU) and venous flow spectral analysis in 100 infertile men without clinical varicocele (group I), 100 infertile men with clinical left varicocele (group II), and 50 fertile men without clinical varicocele served as controls (group III). RESULTS: Three types of flow pattern were found in the spectral analysis of venous flow. If the venous flow was directed to the heart and did not change direction with an intra-abdominal pressure increase, it was classified as type I; venous flow directed to the heart, but changing direction with an intra-abdominal pressure increase, was classified as type II, and blood flow directed to the testicles and augmenting with an intra-abdominal pressure increase, was classified as type III. In group I, flow patterns were 39, 56 and 5% on the left side and 55, 42 and 3% on the right side for types I, II and III, respectively. In group II, flow patterns were 0, 35 and 65% on the left side and 61, 38 and 1% on the right side for type I, II and III patterns, respectively. In group III, the figures were 44, 54 and 2% for the left and 54, 46 and 0% for the right. Type II and III flow patterns were seen more frequently than type I in patients with clinical left varicocele (p<0.001). Whereas type I and II flow patterns were more common than type III in subjects without clinical varicocele (p<0.05). A type II flow pattern during normal breathing was seen at a lower rate in the control group than in the other groups (p<0.05). CONCLUSION: Spectral analysis of Doppler waves should be used in combination with CDU for the diagnosis of varicocele. Varicocele should not only be diagnosed with a type II flow pattern which occurs during valsalva. For the diagnosis of varicocele, the main criterion must be a type III pattern flow, as well as a type II pattern during normal breathing. PMID- 11275727 TI - Effect of subinguinal varicocelectomy on sperm parameters and pregnancy rate: a two-group study. AB - OBJECTIVE: To evaluate the changes in semen parameters and pregnancy rates after varicocelectomy. METHODS: We evaluated the results of surgery in 146 men with primary infertility and palpable left varicoceles, compared with 62 men who refused surgery and were treated with tamoxiphene. Patients were selected with strict criteria in order to exclude any other infertility factor in the couple. They were followed up for at least 1 year after treatment. Statistical analyses were performed with the Wilcoxon signed rank test, Kolmogov-Smirnof two-sample test and paired samples t test. RESULTS: After 1 year, the differences in the median values were significant for all parameters. The partners of 62 of the operated men (46.6%) and 8 of the nonoperated (12.9%) became pregnant within 1 year (p<0.001). Thus, the difference between the true 1-year pregnancy rates was 33.7%. Overall, 83.2% of the operated men improved their semen parameters compared to 32.3% of the nonoperated. CONCLUSIONS: Varicocelectomy improves all semen parameters and pregnancy rates significantly. Palpable varicoceles should be operated upon when found in infertile couples. PMID- 11275728 TI - Feasibility of a temporary urethral stent through the striated sphincter in patients in the early phase (6 months) of spinal cord injury. AB - OBJECTIVE: To assess the feasibility of a temporary urethral stent through the striated sphincter in patients in the early phase (before 6 months) of spinal cord injury (SCI) in a department of neurological rehabilitation. METHODS: Fourteen consecutive men with SCI with urinary retention within 6 months after SCI were prospectively treated. Thirteen patients were tetraplegic (C2 to C7) and 1 was paraplegic. All patients were managed with indwelling catheters (10) or intermittent catheterization (4). The Nissenkorn polyurethane urethral stent was inserted across the external sphincter under local anesthesia for an anticipated 4-month duration. RESULTS: No perioperative complications were encountered. Hospital mean stay at the urological department was 1.9 days (range 1-4 days). All patients had good emptying of the bladder (residual urine less than 100 ml) and were free of all types of catheterization. Five stents had to be repositioned in the first 2 weeks, 1 was removed for obstruction at 2.5 months. There was no lithiasis, no upper urinary tract alteration, no symptomatic infection nor local discomfort during follow-up. At a mean of 3.7 months after implantation, 10/14 (71.5%) patients chose sphincterotomy by permanent urethral stent and 4 had stent removal for learning of self-intermittent catheterization (3) and indwelling catheter (1). CONCLUSIONS: The temporary sphincter stent is a new, feasible and reversible technique to manage neuropathic bladder dysfunction in the early phase after SCI. A randomized study on intermittent catheterization should be conducted. It should consider patients' and nursing caregivers' evaluations. PMID- 11275729 TI - Endoscopic puncture of ureterocele as a minimally invasive and effective long term procedure in children. AB - OBJECTIVES: Over the past years the surgical approach to ureterocele has evolved from complicated major surgery to minimally invasive endoscopic treatment. Because of the high rate of secondary surgery in some recently reported series, an upper pole partial nephrectomy is again recommended as the procedure of choice. We have retrospectively evaluated the long-term results of endoscopic puncture of a ureterocele and its long-term effectiveness and applicability in children. METHODS: Over the past 8 years, 34 patients (20 female, 14 male) were treated in our service with primary endoscopic puncture of a ureterocele. The mean age of the patients was 1.1 +/- 4.3 (mean +/- SD) years. Mean follow-up was 6.1 +/- 2.4 years. Antenatally ultrasound detected the ureterocele in 5 (14%) patients, fetal hydronephrosis leading to the postnatal diagnosis in 13 (38%), and 16 (48%) children presented with symptoms of urinary tract infection (UTI). The ureteroceles presented as part of renal duplication in 31 patients (91%), 3 (9%) in a single system and 1 child had bilateral ureteroceles of a duplex system. Twenty (58%) children had intravesical ureteroceles and the remaining 14 (42%) ectopic ureteroceles. Very poorly functioning upper pole moiety presented in 26 (75%) of the cases and nonfunctioning upper poles in 5 (14%). Twenty of 34 children (58%) had initial vesicoureteral reflux (VUR) to the lower moiety, either to the ipsi (60%) or contralateral kidney (40%). A cold knife incision was carried out in 4 (11.7%), puncture by a 3-french Bugbee electrode in 20 (58%), and the stylet of a 3-french ureteral catheter was utilized to puncture the ureterocele in the remaining 10 patients (30.3%). RESULTS: Complete decompression of the ureterocele was observed in 32 of 34 children (94%). Two patients required secondary puncture 2 years following the primary procedure and are doing well. Upper pole moiety function improved postoperatively in 2 infants and remained stable in all 32 patients, no patient presented with deterioration of the renal function. Six of 20 (30%) patients who had initial VUR to the lower pole, accompanied with recurrent UTI, required surgery. Three underwent ureteric reimplantation and another 3 submucosal polytetrafluoroethylene paste (Teflon) injection. Eight (40%) patients presented with spontaneous resolution of VUR to the lower moiety following puncture of the ureterocele. An additional 6 (17.6%) patients developed VUR to the upper moiety following the puncture of the ureterocele, 3 after cold knife incision and 3 after simple puncture. In 2, submucosal Teflon injection solved the VUR and the remaining 4 patients were maintained on prophylactic antibiotics. In 1 child the reflux resolved spontaneously, and none of them presented with UTI. In 2 cases with nonfunctional upper poles, partial nephrectomy was performed due to symptomatic UTI in spite of complete collapse of the ureterocele 1 and 2 years, respectively, following the initial puncture. No difference was observed in the re-operation rate between the patients with ectopic versus intravesical ureterocele (p<0.05). CONCLUSION: We found that endoscopic puncture of a ureterocele presents an easily performed procedure which allows the release of obstructive ureters and avoids major surgery in the majority of the cases even after a long follow-up. PMID- 11275731 TI - Dag-1 carcinoma cell in studying the mechanisms of progression and therapeutic resistance in bladder cancer. AB - OBJECTIVE: We describe a new human bladder carcinoma cell line (DAG-1) established from a resected bladder cancer fragment and maintained in culture for more than 5 years and over 300 passages. METHODS AND RESULTS: Immunological, biochemical and molecular analysis showed that the DAG-1 cells (62 chromosomes) express the cytokeratines 8, 13, 18 and 20 that confirm their epithelial origin as well as numerous cytokine and cytokine receptor mRNAs. They secrete tissue type plasminogen activator (t-PA), urokinase-type plasminogen activator (u-PA), plasminogen activator inhibitors (PAI-1 and PAI-2), and express u-PA receptors (u PAR/CD87) at their surface. DAG-1 cells are resistant to TNFalpha- and IFNgamma induced apoptosis, two cytokines secreted in the urine of Calmette-Guerin bacillus-treated patients and involved in the tumor regression. CONCLUSION: The DAG-1 cell line is a useful tool, both in vitro and in vivo, to study the progression of bladder tumors and their mechanisms of resistance to immunotherapy in relation with PAI-2 and antioxidant enzymes. PMID- 11275730 TI - Impact of urinary tract infection and detrusor pressure on renal tubular function in patients with vesicoureteral reflux. AB - OBJECTIVE: The aim of this study was to evaluate prospectively the impact of bacteriuria with or without pyuria and/or detrusor pressure on renal tubular function in patients with secondary vesicoureteral reflux. METHODS: From October 1994 to December1998, we evaluated 54 patients with secondary vesicoureteral reflux (26 men and 28 women; age 30+/-24 years), of whom 28 had a neurogenic and 26 a nonneurogenic voiding dysfunction. In a reference population (n = 48; 28 men, 20 women; age 38+/-14 years), 43 had a neurogenic and 5 a nonneurogenic voiding dysfunction. Urinary alpha-1-microglobulin was measured immunonephelometrically. Statistical analysis was performed by multiple regression analysis. RESULTS: Patients with vesicoureteral reflux had a significantly higher urinary alpha-1-microglobulin/creatinine ratio. Urinary alpha-1-microglobulin excretion was related to the grade of vesicoureteral reflux, detrusor pressure and compliance, but not to bacteriuria or pyuria, and was diagnostic for vesicoureteral reflux with a sensitivity of 90%, a specificity of 70% and a negative predictive value of 97%. CONCLUSION: Urinary alpha-1 microglobulin excretion is diagnostically useful in patients with secondary vesicoureteral reflux. The use of urinary alpha-1-microglobulin excretion in the follow-up of patients with vesicoureteral reflux has yet to be established. PMID- 11275732 TI - Nitric oxide synthase gene and protein expression are upregulated by Bacille Calmette-Guerin in the rat bladder. AB - OBJECTIVE: We hypothesize that Bacille Calmette-Guerin (BCG) may act through the regulation of various isoforms of nitric oxide synthase (NOS) [inducible (iNOS), endothelial (eNOS), and neuronal (nNOS)] genes and protein expression in rat bladder. METHODS: Adult female Sprague-Dawley rats (200-250 g) were injected transurethrally with BCG (22 rats) and phosphate-buffered saline (22 control rats), and after 2, 4, 6, and 12 h, and 1, 2, 3, 5, 7, 10, and 14 days, the bladders were harvested. Normal and BCG-treated rat bladders were analyzed for mRNA expression for iNOS, eNOS, and nNOS by reverse-transcriptase polymerase chain reaction. Protein expression was determined by Western blotting and immunohistochemistry. RESULTS: mRNA expression for iNOS was induced after 2 h of BCG injection in the rat bladder. Gene expression for iNOS was highest at 6 h to 1 day followed by decreased expression, reaching its lowest level at 5 days. eNOS mRNA expression was detected in control bladders, but its level was higher in the BCG-treated animals. nNOS mRNA expression was present in all samples, but did not change after BCG treatment. Western blotting confirmed these findings. Immunohistochemistry analysis demonstrated that eNOS was present mainly in endothelium, while iNOS was detected in stroma and nNOS in epithelium and smooth muscle of the rat bladder. CONCLUSION: The present study demonstrates, for the first time, that BCG treatment up-regulates gene and protein expression of iNOS and eNOS in normal rat bladders, suggesting that BCG action may be mediated through NOS pathways. PMID- 11275733 TI - Renal actinomycosis mimicking renal carcinoma. AB - The case of a 52-year-old man is reported who presented with night sweats and slight debilitation. Upon CT scan a left-sided renal mass with centrally liquefied areas was detected. The patient underwent nephrectomy for suspected renal cancer with central necrosis. Histologically, the diagnosis of renal actinomycosis was established based on the detection of sulphur granules. Actinomyces israelii is an anaerobic gram-positive bacterium that may cause localized tumour-like infections mainly in the craniocervical region and exceptionally retroperitoneally. Renal actinomycosis is a rare differential diagnosis of renal masses. As nephrectomy may prove hazardous in these cases, the diagnosis should be attempted pre-operatively by ultrasound-guided aspiration and consecutive antibiotic treatment. In selected cases surgery could be avoided at all. PMID- 11275734 TI - Renal cancer. PMID- 11275735 TI - Introduction and conclusions. Improving the management of lower urinary tract symptoms in real life practices. PMID- 11275736 TI - Lower urinary tract symptoms suggestive of benign prostatic obstruction: what is the available evidence for rational management? AB - Because of the need for a more rational management of LUTS suggestive of BPO, EBM is receiving more attention. Treatment decisions should be based on scientific evidence more than on personal preferences of physicians and patients. EBM is more than following rigid treatment guidelines or published advice. It is a process of turning problems of clinical practice into well-defined questions and then finding and appraising available evidence and implementing it into clinical practice. In cases of LUTS suggestive of BPO, much evidence is available on the efficacy and tolerability of different treatment modalities in the short term, whereas long-term data on (cost)-effectiveness are rather scarce. From a patient's point of view, newer pharmacotherapeutic agents can be considered as an appropriate treatment option. alpha(1)-Adrenoceptor antagonists seem to be more efficacious than finasteride and within this group of drugs, selective alpha(1A)/alpha(1D)-adrenoceptor antagonists particularly show a favourable efficacy/tolerability ratio. PMID- 11275737 TI - Lower urinary tract symptoms suggestive of benign prostatic obstruction: how can clinical expertise contribute to rational management? AB - OBJECTIVE: To perform a systematic analysis of clinical expertise on treatment for benign prostatic hyperplasia (lower urinary tract symptoms (LUTS) suggestive of benign prostatic obstruction (BPO)) and to investigate the usefulness of these data in further guideline development. METHODS: A modified Delphi method was used to analyse the opinions of a panel of 15 European urologists on the appropriateness of 4 common treatments for 1,152 "indications" (hypothetical cases) for LUTS suggestive of BPO. Each indication consisted of a unique combination of 9 diagnostic variables, found to be relevant in treatment choice in previous research. The study population was restricted to patients for whom current guidelines do not provide clear indications on the most appropriate treatment. The panellists individually rated the appropriateness of three active treatments (surgery, alpha(1)-adrenoceptor antagonists, finasteride) using a 9 point scale, all in comparison with "watchful waiting". Aggregate panel judgements were calculated from individual ratings for each indication (appropriate, inappropriate, and uncertain). The relationship between diagnostic characteristics and panel opinions was analysed using logistic regression methods. The results were compared to those of an identical panel study including 12 Dutch urologists. RESULTS: Strong agreement existed for 42.5% of the indications, while strong disagreement was found in only 0.1%. For patients who had not previously been treated for LUTS, surgery was considered appropriate in 44% of the indications. For alpha(1)-adrenoceptor antagonists and finasteride these percentages were 56 and 6 respectively. Strong contra-indications were found only for finasteride (34%). Logistic regression analysis demonstrated consistent panel opinions, indicating a strong cumulative impact of almost all diagnostic variables on the panel judgement "appropriate". The figures on appropriateness were highly comparable to the results of the Dutch study (overall agreement 84%, kappa 0,76). A computer program was constructed to facilitate the implementation and evaluation of the panel recommendations in daily clinical practice. CONCLUSIONS: Given the consistency of the panel opinions, the results may be useful in complementing evidence-based guidelines for LUTS suggestive of BPO in the grey area of treatment choice. PMID- 11275738 TI - Lower urinary tract symptoms suggestive of benign prostatic obstruction--what's the long-term effectiveness of medical therapies? AB - The primary treatment goal in patients with lower urinary tract symptoms (LUTS) suggestive of benign prostatic obstruction (BPO) is symptom relief. However, it is also important to assess the effects of medical treatments on disease progression because this can lead to complications such as acute urinary retention or can ultimately require surgical treatment. In a placebo-controlled study finasteride was shown to reduce the incidence of acute urinary retention and the need for surgery in patients with LUTS suggestive of BPO. While alpha(1) adrenoceptor antagonists are superior to finasteride with regard to LUTS relief, systematic studies on their effects on BPO complications and progression to surgery are not available. However, we reviewed a number of smaller studies which consistently indicate that alpha(1)-adrenoceptor antagonists may be at least as effective as finasteride in the prevention of acute urinary retention or the need for surgery in patients with LUTS suggestive of BPO. While this needs to be confirmed in adequately designed and powered studies, such evidence should be taken into consideration when choosing between alpha(1)-adrenoceptor antagonists and finasteride treatment. PMID- 11275740 TI - Lower urinary tract symptoms suggestive of benign prostatic obstruction--Triumph: design and implementation. AB - Lower urinary tract symptoms (LUTS) suggestive of benign prostatic obstruction (BPO) are likely to become an increasing burden for health care budgets in the future due to the high prevalence of this condition in the elderly and the ageing of the population. Data on the cost-effectiveness of the available treatment options in order to promote rational management will therefore be crucial. As this information is currently largely lacking, the European Association of Urology (EAU) has endorsed the formation of a Pan-European Expert Panel that has developed the Triumph (TransEuropean Research Into the Use of Management Policies for LUTS suggestive of BPH in Primary Healthcare) project. The Triumph project will evaluate how LUTS suggestive of BPO is currently managed in real life practice in initially 6 European countries (France, Germany, Italy, Poland, Spain and the UK) and how this condition progresses over time in relation to an initial treatment choice. In other words, how many patients remain on their initial therapy, how many switch to another therapy (including surgery) and within which time frame and how many develop serious complications related to BPO such as acute urinary retention. The clinical data will be retrieved from continuous analysis of large computer-based patient files such as the General Practice Research Database (GPRD) in the UK and the Integrated Primary Care Information (IPCI) database in The Netherlands. Furthermore, a cross-sectional, observational survey will gather similar data in a prospective fashion for at least 1 year from approximately 13,000 patients managed by more than 1,000 primary care physicians in all 6 countries. Both newly diagnosed (around 7,500) and existing (around 5,500) LUTS patients will be enrolled. For the existing patients in whom the diagnosis was made in the past (from January 1997 onwards), the relevant retrospective data (up to 3.5 years) on disease progression will also be collected. Finally, the costs associated with these treatments will be investigated by the development of a national costing database for each participating country that will detail costs related to visits, investigations, prescriptions, surgical interventions, etc. The last patient is anticipated to come out of the initial prospective survey by the end of 2002/beginning of 2003 and the results will hopefully contribute to improved cost-effective management of LUTS suggestive of BPO in the future. PMID- 11275739 TI - Lower urinary tract symptoms suggestive of benign prostatic obstruction: what are the current practice patterns? AB - OBJECTIVE: The current clinical practice patterns for the management of LUTS suggestive of BPO in the US and in various European countries will be reviewed. METHODS: Information was obtained from published scientific articles and IMS/GERS market analysis data. RESULTS: Community-based surveys demonstrate that the prevalence of moderate to severe LUTS in elderly men is high and increases with age. The role of the GP in the initial management of LUTS is growing. In France, Italy and the UK, most patients with LUTS initially visit a GP. More and more patients in the US also first seek medical advice from a primary care physician or an internal medicine specialist. In Germany, both GPs and office-based urologists are involved in the initial management of LUTS. In Poland and Spain, office-based urologists initiate primary therapy for LUTS, although a trend towards involvement of GPs is also seen, especially in Poland. The shift in the initial management of LUTS from secondary to primary care accompanies a decreased incidence of surgery and a growing demand for medical therapy. Currently, there are considerable differences between the medical management of LUTS suggestive of BPO across Europe in real life practice. For example phytotherapy is particularly popular in countries such as Germany, France and Spain, whereas finasteride is more commonly used in Italy, Poland and the UK. alpha(1)-Adrenoceptor antagonists are used in most of these countries as the primary treatment modality. The data furthermore suggest that the current management of patients is often more opinion than evidence-based, which may at least partly be due to the fact that data on long-term effectiveness of treatment options in real life clinical practice are largely lacking. CONCLUSIONS: [corrected] Due to the ageing and longevity of the population, the costs associated with the management of LUTS suggestive of BPO will rise in the future, whereas healthcare budgets will be relatively restricted. In order to improve cost-effective management of LUTS, more and better studies are needed in real life practice in primary care. These studies should not only be based on classical efficacy and safety data, but also on effectiveness of treatment in the long-term and the associated costs. PMID- 11275741 TI - Lower urinary tract symptoms suggestive of benign prostatic obstruction--Triumph: health-economical analysis. AB - Given the ageing of western populations, the cost burden associated with the treatment of LUTS suggestive of BPO will increase substantially over the next few decades. Therefore, from the economic perspective, the primary objective of the Triumph project is the assessment of the cost-effectiveness of treatment options for lower urinary tract symptoms (LUTS) suggestive of benign prostatic obstruction (BPO), formerly referred to as symptomtic benign prostatic hyperplasia (BPH), in medical practice in initially six European countries. All modalities of LUTS treatment used in these countries will be considered. The effectiveness and costs associated with these treatments will be assessed in each country. Data will be gathered from observed medical practice rather than in the setting of a trial. Country-specific aspects will be studied and the outcome of country-specific policies can be predicted. Patient quality of life will also be measured using the I-PSS score as a basis. A number of treatment scenarios will be assessed in terms of both their costs and long-term effects, using a computer simulation. These economic analyses will provide greatly improved insight into the most cost-effective treatments for LUTS suggestive of BPO. PMID- 11275742 TI - Lower urinary tract symptoms suggestive of benign prostatic obstruction--Triumph: the role of general practice databases. AB - The Triumph project aims to document the current management of lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) in general practice and to assess the effectiveness of the initial treatment options used. The first phase of the project will consider existing data sources in primary care. A patient's medical record will contain most, if not all, clinically relevant information, and databases combining the records from a network of computerised general practices can provide longitudinal data for complete populations, linking prescribing records to clinical information on disease progression and outcomes for individual patients. Database research can provide rapid information and offers the ability to conduct studies on a scale that would previously have been prohibited by both time and expense. Within the Triumph project, the THALES, General Practice Research Database (GPRD) and Integrated Primary Care Information (IPCI) databases are, or will be, used to examine the current management of LUTS/BPH in France, the UK and the Netherlands respectively. Preliminary results from the UK General Practice Research Database (GPRD) showed that LUTS/BPH incidence increased linearly from the ages of 45 to 85 years (r(2) = 0.992) and prevalence increased from 3.5% to 35% for men in their late 40s and 80s respectively. With treatment failure defined as a change to another medical therapy, catheterisation or prostatic surgery, and accounting for age and year variation, patients receiving the older alpha(1)-blockers (indoramin and prazosin) appeared to fail significantly earlier than those receiving finasteride. There was no significant difference between finasteride and the newer alpha(1)-blockers (tamsulosin, alfuzosin, terazosin and doxazosin). Patterns of changes between products from the THALES database in France were broadly similar to those seen in the UK. PMID- 11275743 TI - Multiple breeding females in captive groups of golden-headed lion tamarins (Leontopithecus chrysomelas): causes and consequences. AB - In Callitrichidae, reproduction in subordinate females is generally inhibited but occurs in rare cases, possibly in association with the presence of an unrelated male, important food resources or low dispersal opportunities. This study investigates the occurrence of groups with multiple breeding females in captive golden-headed lion tamarins (Leontopithecus chrysomelas), the factors leading to their formation and the consequences for the group. Information obtained from studbook data on the world captive population during 1984-1998 revealed that polygynous groups in captivity are very rare: only 7 cases were discovered. Family groups in which daughters started breeding with a related male were larger than average, had a high number of sexually mature sons and eldest offspring that were well past the age of sexual maturity. Following a breeding attempt, severe aggression frequently occurred, especially if the infants were liveborn. Polygynous groups composed of two related females and an unrelated male tended to remain stable for a longer period than families with breeding daughters. Competition for infant care is probably an important factor determining whether the polygynous situation can persist and for how long. PMID- 11275744 TI - Field kit to characterize physical, chemical and spatial aspects of potential primate foods. AB - An outline is given for a field kit aiming to substantially increase the in situ knowledge gleaned from feeding studies of primates. Measurements are made of colouration (spectrum of non-specular reflection) and many mechanical, chemical and spatial properties of primate foods. PMID- 11275745 TI - Spectral sensitivity of gibbons: implications for photopigments and color vision. PMID- 11275746 TI - Ancient DNA from Megaladapis edwardsi (Malagasy Subfossil): preliminary results using partial cytochrome b sequence. PMID- 11275747 TI - How a subordinate male bonobo leads dominant females up the garden path. PMID- 11275748 TI - Evidence of tree gouging and exudate eating in pygmy slow lorises (Nycticebus pygmaeus). PMID- 11275749 TI - The seasonal diet of the Sichuan snub-nosed monkey (Pygathrix roxellana) in Shennongjia Nature Reserve, China. PMID- 11275750 TI - Female-biased maternal investment in rhesus macaques. PMID- 11275751 TI - Female dominance in African lorises (Otolemur garnettii). PMID- 11275752 TI - A comparison of the sleeping behavior of three sympatric primates. a preliminary report. PMID- 11275753 TI - Absence of intragroup coalitions in adult male red colobus (Colobus badius tephrosceles) in the Kibale National Park, Uganda. PMID- 11275781 TI - Selection and evaluation of children for epilepsy surgery. AB - Major determinants of the success of epilepsy surgery are the proper selection and evaluation of the patient. Candidates should have medically intractable, disabling epilepsy. The evaluation requires a unified, systematic approach by a comprehensive team of health professionals. Key points, common to most evaluations, are enumerated and provide a useful tool for mental organization. In general, the greater the degree of congruent information derived from these investigations, the better the candidacy for surgery. After this, completeness of resection and etiology are other important predictors of success. Children present unique challenges and opportunities owing to the vulnerability and plasticity of the developing nervous system. Surgery for intractable epilepsy can be a rewarding therapy, and in select children, the only chance for cure and substantial development progress. PMID- 11275782 TI - Hydrocephalic children presenting to a Malaysian community-based university hospital over an 8-year period. AB - There are few local statistics on the incidence of hydrocephalus and the outcome of hydrocephalic shunts in the South East Asian region. We report a retrospective study on 285 hydrocephalic patients who underwent shunting procedures between 1990 and 1998 at the University Hospital Science Malaysia, a regional referral center. Multiple logistic regression analysis was applied to predict determinants of outcome in relation to the timing of diagnosis, other congenital abnormalities associated with the hydrocephalus, timing of surgery and cortical thickness from CT scan. The relationship of shunt infection was correlated to the age of the patient and surgical procedure. The predictors for developmental outcome reported by this study were age at diagnosis, type of brain abnormalities and gender. Time of operation and cortical thickness did not contribute to the outcome. PMID- 11275783 TI - Effect of fetal hydrocephalus on the distribution patterns of calcium-binding proteins in the human occipital cortex. AB - Neuronal pathology in the cerebral cortex (CC) of fetal hydrocephalus brains is quite subtle when applying routine Nissl staining. This study is aimed at investigating alterations of interneurons which can be immunolabelled by antibodies against the calcium-binding proteins calretinin (CR), calbindin (CB) and parvalbumin (PV). The subplate (SP) subjacent to the cortex anlage is included as this transient zone plays a pivotal role in the establishment of cortical connections. Nine occipital lobes from cases of fetal hydrocephalus and 9 controls were categorized according to age: 21-25 weeks of gestation (group 1) and 32-36 weeks (group 2). No differences in the distribution, number and morphology of CR-immunoreactive (ir) neurons are seen when comparing hydrocephalic with control brains of group 1. In severely altered hydrocephalic brains of group 2, the distribution and number of CR-, PV- and CB-ir nerve cells are not altered; however, they appear shrunken and processes are less immunolabelled. In extremely altered tissue PV-, CB-ir neurons cannot be detected, whereas the number of CR-ir somata is not reduced. The data indicate that subpopulations of interneurons of the CC may be differentially damaged. The alterations observed in the SP may implicate a possible impairment of transient neuronal circuitries that are essential for the development of cortical connections. On the whole, these neuronal alterations may account for residual deficits observed after shunting. PMID- 11275785 TI - Surgical treatment of hypothalamic hamartoma and refractory seizures: a case report and review of the literature. AB - Refractory gelastic seizures are often associated with hypothalamic hamartoma (HH). Presurgical evaluation in such children often points to a distinct cortical region as the source of the seizures. A case of a child with HH and refractory seizures is presented. Video-EEG monitoring revealed a well-defined epileptic focus in the left frontal region. In accordance with the current understanding of the nature of hamartoma-related seizures, the hamartoma was resected. Follow-up evaluations revealed a marked improvement in seizure frequency and global functioning. PMID- 11275784 TI - Antley-Bixler syndrome, description of two new cases and review of the literature. AB - OBJECTIVE: Antley-Bixler syndrome (ABS) is a rare disorder characterized by multiple malformations of cartilage and bone including multisynostotic osteodysgenesis, midface hypoplasia, choanal atresia or stenosis, femoral bowing, neonatal fractures and multiple joint contractures and, occasionally, urogenital, gastrointestinal or cardiac defects. Since the first report in 1975, at least 34 cases (including this report) have been described. We present 2 cases of congenital hydrocephalus, suggesting the cause of craniosynostosis and midface hypoplasia is associated with ABS. CLINICAL PRESENTATION: The first case was a 1 day-old female with Arnold-Chiari malformation, multiple cranial synostosis, obstructive hydrocephalus and radioulnahumeral synostosis. Ventriculoperitoneal (V-P) shunting was performed when she was 7 days old. She died 42 days later due to cardiopulmonary failure. The second case was a 2-month-old female with bilateral coronal synostosis, obstructive hydrocephalus and brachycephaly. V-P shunting was done soon after her admission, and bilateral lateral canthal advancement with the floating forehead procedure was performed 1 month later. She is still alive with good development. DISCUSSION: The main anomalies of the ABS can be divided into 4 categories: craniofacial, skeletal, extremity and urogenital anomalies. They may be the result of mutation in the fibroblast growth factor receptor 2 (Ser351Cys) gene, which was confirmed in our case 2. Craniosynostosis combined with hydrocephalus created congenital increased intracranial pressure (IICP). Early V-P shunt implantation and surgical release of the closed suture and lateral canthal advancement should be done as soon as possible, ideally when the patient is younger than 3 months. CONCLUSION: Early correction of craniosynostosis was feasible and safe for one of our patients. We offer our experience in the treatment of hydrocephalus and correction of craniosynostosis to relieve IICP. PMID- 11275786 TI - Intramedullary abscess of the spinal cord. AB - A case of intramedullary abscess of the spinal cord in a child aged 3 years is reported. Successful outcome depends on early diagnosis and aggressive management. PMID- 11275787 TI - Occipital aneurysmal bone cyst. PMID- 11275789 TI - An 8-year-old girl with onset of seizures. PMID- 11275788 TI - A 14-year-old female with decreasing visual acuity, severe headache, nausea and vomiting. PMID- 11275790 TI - Von Hippel-Lindau tumour suppressor gene is not involved in sporadic human breast cancer. AB - OBJECTIVE: Mutations of the von Hippel-Lindau (vhl) gene, as well as allelic loss at the gene region (3p25-26) have been described in sporadic cases of the tumour types participating in VHL disease, but also in cancers not associated with the syndrome. In this study, we attempted mutation analysis of the vhl gene, as well as detection of allelic loss at 3p25-26 in sporadic human breast cancer. METHODS: Eighty-two tumour specimens were screened for loss of heterozygosity (LOH) at the vhl region, and compared to the adjacent, histologically normal tissue. Furthermore, mutations within the three exons of vhl in the same panel of tumours were detected using SSCP and heteroduplex analysis and direct sequencing. RESULTS: To our knowledge this is the first mutational analysis reported for the vhl gene in breast cancer, however we failed to reveal any mutations in the specimens examined. All the cases were informative for at least one of the microsatellite markers tested, 24 (29.2%) exhibited LOH at 3p25-26. Clinical and pathological data were available for all tumours examined, however no significant correlations were encountered. CONCLUSION: These results strongly indicate against a critical involvement of the tumour suppressor vhl in breast carcinogenesis. PMID- 11275791 TI - bcl-2 Expression and apoptosis in primary and metastatic breast carcinomas. AB - OBJECTIVES: To evaluate the role of bcl-2 and apoptotic index in the progression from primary to metastatic breast carcinoma and their influence on prognosis. METHODS: bcl-2 expression was examined by immunohistochemistry and apoptotic index by in situ end-labelling in 116 surgical breast carcinomas and lymph node metastases from 50 patients. RESULTS: bcl-2 was observed in 69 cases (59.4%) of primitive carcinomas and 26 cases (65%) of metastatic breast carcinomas and there was agreement of bcl-2 expression between primary and metastatic sites except in 3 cases. bcl-2 expression was significantly associated with several favourable prognostic features, such as small tumour size (p = 0.03) and oestrogen and progesterone-receptor positivity (p < 0.01 and p < 0.001, respectively). A high apoptotic index was significantly associated with a number of poor prognostic factors, including poorly differentiated carcinomas, large tumour size, high Ki67 expression and high mitotic count (p < 0.001 in all cases). The mean apoptotic index was higher in lymph node metastasis than in primary carcinomas (1.19 vs. 0.69, p < 0.01). A low bcl-2 expression and a high apoptotic index were significantly associated with short-relapse free survival rates (p = 0.02 and p < 0.01, respectively), but only apoptotic extent provided independent prognostic information by multivariate analysis. CONCLUSIONS: The evaluation of bcl-2 expression and extent of apoptosis may provide useful prognostic information on breast cancer patients; however while increased apoptosis is strongly associated with the progression from primary carcinomas to lymph node metastases, bcl-2 does not seem to play a significant role in this process. PMID- 11275792 TI - Behavior of murine renal carcinoma cells grown in ectopic or orthotopic sites in syngeneic mice. AB - We examined whether the organ microenvironment modulates the metastatic behavior and the response to doxorubicin (DXR) in murine renal carcinoma (RENCA) cells. Tumor cells were injected into kidney (orthotopic) and subcutis (ectopic) of syngeneic mice. Lung metastases developed in up to 57% (17/30) of animals having kidney tumors but not in those with skin tumors. Tumors growing in the kidney were more resistant to DXR than tumors growing in the subcutis when mice were given intravenous injections of DXR (8 mg/kg) on days 8 and 15 after implantation. In addition, tumor cells cultured from kidney tumors were initially more resistant to DXR than tumor cells cultured from subcutis tumors. After tumor cells were passaged in vitro, all cells exhibited a similar sensitivity to DXR. Additionally, we examined the expression levels of mdr1, EGFR and type IV collagenase by an in situ mRNA hybridization technique. A higher mRNA expression for type IV collagenase and EGFR was found in kidney tumors than in subcutis tumors. These results demonstrate that the organ environment influences the drug responsiveness and the expression of metastasis-related genes in murine renal carcinoma cells. PMID- 11275793 TI - Intrahepatic metastasis by orthotopic implantation of a fragment of murine hepatoma and its related molecules. AB - Intrahepatic metastasis is a major modality in the recurrence of hepatoma. Establishment of the intrahepatic metastasis model would be useful for evaluating new anticancer therapies and analyzing the molecular mechanisms of tumor metastasis. Orthotopic implantation of a fragment of CBO140C12 hepatoma into the liver resulted in the formation of a solitary tumor nodule and its intrahepatic metastasis. In contrast, implantation of ADras3 cancer cells did not show any metastasis on day 21. CBO140C12 cells showed enhancement of the invasive, adhesive and migratory capabilities, as compared with ADras3 cells. Furthermore, mRNA expression and gelatinolytic activity of MMP-9 were detected in CBO140C12 cells, and the expression of mRNA for MT1-MMP in CBO140C12 cells was greater than that in ADras3 cells. Thus, intrahepatic metastasis of CBO140C12 tumor might be involved in the enhancement of the invasiveness of tumor cells via marked expression of MMP-9 and MT1-MMP. PMID- 11275794 TI - Serum anti-p53 antibodies in uterine and ovarian cancer: association with dna sequence copy number abnormalities. AB - The aim of the present study was to evaluate the clinical significance of the serum anti-p53 antibody in patients with uterine and ovarian cancer. Some of the ovarian patients were also evaluated for overexpression of p53 by immunohistochemistry and for cytogenetic alterations by comparative genomic hybridization (CGH). Serum anti-p53 antibodies were determined by an enzyme immunoassay kit. The antibody was detected in 8/30 (27%) of ovarian cancers, in 12/86 (14%) cancers of the uterine cervix, in 5/41 (12%) cancers of the uterine body, and 0/9 (0%) healthy women. The overall survival rate in patients with ovarian cancer was significantly worse in patients with anti-p53 antibody positivity than that in patients with anti-p53-antibody-negative cancers using the log rank test (p = 0.017). There was a significant correlation between the presence of anti-p53 antibody and tissue overexpression of p53 in ovarian cancers. CGH analysis showed that the aberrations in DNA sequence copy number in ovarian cancers were significantly increased in anti-p53-antibody-positive cases compared to antip53-antibody-negative cases including increased copy number on 20q and reduced copy number on 5q and 13q. Although the exact relationship between the presence of serum anti-p53 antibody (specific humoral response) and cytogenetic alterations is still unknown, these findings suggest that the measurement of serum anti-p53 antibody may be useful for the assessment of genetic instability and tumor biological aggressiveness. PMID- 11275795 TI - Automated immunofluorometric assay for MUC1. AB - The aim of the present study was to establish a robust, reliable and fully automated immunofluorometric assay for the breast cancer serum marker MUC1. This would further serve as a prototype assay for evaluation of other MUC1 assays based on new antibody combinations. Using time-resolved fluorescence as tracer signal we developed an automated immunofluorometric assay for MUC1 (MUC1 IFMA). This assay was compared with two commercial assays. The CA15-3 EIA (CanAg) which use the same antibodies as the MUC1 IFMA, and the ETI-CA-15-3 K (Sorin) which use the original antibodies defining the CA 15-3 assay. The three assays showed comparable results. The coefficient of variation was below 10% from 9 to 2,400 kU/l for the MUC1 IFMA, from 15 to 250 kU/l for the CA15-3 EIA, and from 25 to 200 kU/l for the ETI-CA-15-3 K assay. At a specificity of 0.94 the overall diagnostic sensitivities for the MUC1-IFMA, CA15-3 EIA and ETI-CA-15-3 K assays were 0.40, 0.37, and 0.38, respectively. When applied to metastatic breast cancer, all assays had sensitivities close to 0.80. There was a close correlation (Spearman rank = 0.99) between results from the new assay and the CA15-3EIA. The new automated assay was not strictly immunometric as we could not achieve conditions where solid phase or tracer antibodies were in apparent excess. However, the assay performed well at a wide range of assay conditions. The automation, which minimizes imprecision in pipetting and handling of samples, and the high capacity of the AutoDELFIA instrument enabling measurement of all samples in a single run, were important aspects for establishing a reliable assay. The principle of the new automated immunofluorometric assay will be used as a rapid and reliable evaluation of a wide range of monoclonal antibody combinations in our search for the optimal MUC1 assay. This new automated immunofluorometric assay will be useful in the rapid and reliable evaluation of a wide range of monoclonal antibody combinations in our search for the optimal MUC1 assay. PMID- 11275796 TI - Abnormal expression of epidermal growth factor receptor and c-erbB2 in squamous cell carcinoma of the cervix: correlation with human papillomavirus and prognosis. AB - The aim of this study is to assess the expression of epidermal growth factor receptor (EGFR) and c-erbB2 and their correlation with human papillomavirus (HPV) status and prognosis in squamous cell carcinoma of the cervix. The expression of EGFR and c-erbB2 was studied at the protein level using the immunohistochemical (IHC) staining method, at the RNA level using the ribonuclease protection assay and at the DNA level using Southern blot and hybridization method. One hundred and one patients with squamous cell carcinoma of the cervix were recruited. Fifty one patients were of stage 1B/2A and 50 patients were of stage 2B and above. Positive IHC stainings of EGFR and c-erbB2 proteins were found in 74.2 and 19.8% of cases, respectively. DNA amplifications of EGFR and c-erbB2 genes were detected in 35.4 and 17.2%, respectively. Of the patients showing positive EGFR and c-erbB2 staining, only 39.2 and 25%, respectively, showed DNA amplifications. RNA overexpression of EGFR or c-erbB2 was only detected in 2% of cervical cancers and was associated with positive staining and DNA amplifications. HPV was detected in 79.2% of the cases by HPV consensus primers L1, in 57.4% for HPV 16 and 27.7% for HPV 18. The abnormal expression of EGFR and c-erbB2 had no correlation with HPV detection and had no prognostic significance on survival. PMID- 11275797 TI - Carcinoembryonic antigen continuous epitopes determined by the spot method. AB - Carcinoembryonic antigen (CEA) is a heavily glycosylated tumor-associated protein with an N-A1-B1-A2-B2-A3-B3 domain structure. Circulating CEA immunoassays are used for monitoring digestive cancer patients, and radiolabeled anti-CEA monoclonal antibodies (MAb) are used for the diagnosis and therapy of CEA positive tumors. The five major nonoverlapping epitopes (Gold 1-5) have been broadly correlated with the domain organization, but there is no precise localization of the epitopes at the sequence level. In an attempt to identify the peptide sequences corresponding to the five Gold epitopes on the CEA molecule, we prepared a set of 227 overlapping fifteen-mer peptides corresponding to the complete CEA sequence with the SPOT method. Using five high affinity MAbs directed against the five CEA Gold epitopes, we demonstrated that none of these epitopes could be mimicked by a fifteen-mer peptide sequence. However, using rabbit and goat anti-CEA sera, we identified six major continuous antigenic regions. All are included in the Ig-like domains of the CEA: two in the A1 domain (residues 120-134 and 153-164), one each in the A2 (329-337) and A3 domains (508 513), one at the junction between the A3 and B3 domains (553-561) and one in the B3 domain (565-573). A very homologous sequence (common residues VSPRL) was mapped in each of the three A domains. Thus, in terms of occurrence of continuous epitopes, the Ig-like domains A1, A2, A3 and B3 seem to be the most antigenic parts of CEA. These peptide sequences should be good candidates for the future development of site-specific anti-CEA MAbs. PMID- 11275798 TI - Carcinogenesis: mutations and mutagens. AB - In normal human cells there is a steady accumulation of mutations with time. We argue that the great majority of these mutations arise spontaneously and are due to endogenous factors or processes that damage DNA. A small fraction of these mutations converts a normal cell into a cell that is initiated towards the development of cancer. We propose that, in general, these initiated cells are more susceptible to the mutagenic effects of exogenous carcinogenic agents than to the mutagenic effects of endogenous factors. Indeed, it can be assumed that in most instances the initiation event is due to a mutation which causes inactivation or loss of a mutation avoidance gene, such as the p53 gene, or a gene which is involved in the repair of damaged DNA. Recent studies have shown that most of such mutations lead to a considerable enhancement in the mutagenicity of many exogenous agents, whereas the mutagenicity of endogenous factors is less affected. Furthermore, the progressive accumulation of mutations with increasing age implies that more initiated cells are likely to be found in older individuals. Therefore, sensitivity to the carcinogenic effect of exogenous mutagens can generally be assumed to increase in older people. PMID- 11275799 TI - Coexisting linear and disseminated drug eruption: a clinical clue to the understanding of the genetic basis of drug eruptions. PMID- 11275800 TI - EMLA anaesthetic cream for sharp leg ulcer debridement: a review of the clinical evidence for analgesic efficacy and tolerability. AB - Sharp debridement is a fast method of achieving a clean leg ulcer, which promotes healing and enables skin grafting. EMLA cream is the only topical anaesthetic for which there is clinical evidence of analgesic efficacy for debridement. Thirteen clinical investigations of EMLA are reviewed. Four double-blind studies and one open randomised controlled study show that EMLA applied to the ulcer for 30-45 min under occlusion significantly reduces the pain from sharp debridement, decreases the incidence of post-debridement pain and reduces the time needed to achieve a clean ulcer, giving potential savings in healthcare costs. Doses of up to 10 g EMLA result in plasma levels of lidocaine and prilocaine well below toxic levels. Repeated treatment does not change the bacterial flora of the ulcer and rarely causes sensitisation. The treatment of pain in leg ulcer patients is important for patient satisfaction and for patient-perceived quality of life. PMID- 11275801 TI - Familial occurrence of nevus sebaceus of Jadassohn: another case of paradominant inheritance? AB - We describe two sibs (a boy and a girl) suffering from linear nevus sebaceus of Jadassohn. The parents are not affected. This rare pattern of familial occurrence prompts us to suggest that these sibs represent a case of paradominant inheritance of a mosaic disorder. PMID- 11275802 TI - Reduced levels of glutathione S-transferases in patch test reactions to dithranol and sodium lauryl sulphate as demonstrated by quantitative immunocytochemistry: evidence for oxidative stress in acute irritant contact dermatitis. AB - There is increasing evidence that oxidative stress plays a role in the pathogenesis of acute irritant contact dermatitis. As part of on-going studies into the effect of irritant chemicals on the anti-oxidant enzyme systems in the skin, we have examined the changing levels of two classes of glutathione S transferase in patch test reactions to dithranol and sodium lauryl sulphate, using quantitative immunocytochemistry. Although no changes were evident after 6 hrs, significant reductions in the density of staining for glutathione S transferase alpha were seen with both irritants after 48 hrs and 96 hrs. Glutathione S-transferase pi levels were reduced to a lesser degree, reaching significance for dithranol at the 96 hrs time point only, and for sodium lauryl sulphate at 48 hrs only. The results support the hypothesis that oxidative stress plays a role in chemically-induced inflammation, not only in the case of irritants such as dithranol which are known to directly generate reactive oxygen species, but also with chemicals not generally associated with free radical generation. PMID- 11275803 TI - Immunohistochemical detection of interferon-gamma-producing cells in dermatophytosis. AB - Skin lesions of dermatophytosis are thought to be a result of a T cell-dependent inflammatory response that is mediated by various cytokines. We examined whether IFN-gamma-positive cells (as expression of Th1 response) were present in the skin lesions of dermatophytosis in situ by immunohistochemical techniques. Mixtures of CD4-positive T cells and CD8-positive T cells were found to be present in the dermal infiltrates of the lesions. Considerable numbers of CD1a-positive cells were detected in the upper dermis and epidermis. A marked accumulation of CD68 positive cells was found in the upper dermis. IFN-gamma-positive cells were present in the upper dermis of the lesions. The pattern of IFN-gamma staining appeared to be intracellular in mononuclear lymphoid cells. The staining was considered to be highly specific because it could be completely blocked by preabsorption with recombinant IFN-gamma. Our data support the hypothesis that the skin lesions of dermatophytosis may be associated with a Th1 response. Th1 response, which is characterized by IFN-gamma release, is thought to be involved in the host defense against dermatophytes and to reflect cutaneous reaction in dermatophytosis. PMID- 11275804 TI - Detection of cutaneous varicella zoster virus infections by immunofluorescence versus PCR. AB - Detection of localized, clinically atypical cutaneous infections with varicella zoster virus (VZV) has proven difficult, as serum antibody tests sometimes are not sensitive and specific enough for that purpose. Therefore immunofluorescence and an internally controlled PCR for VZV are compared for sensitivity. Detection of PCR products was done by ELISA, and if positive, additionally by agarose gel electrophoresis. Of 60 samples 44 were PCR-positive by ELISA (44 = 100%), of which 37 (84%) were also positive on the agarose gel. Thirty-four samples (77%) were positive by immunofluorescence. No sample was positive by immunofluorescence and negative by PCR. A combination of immunofluorescence and PCR with agarose gel analysis detected 42 samples out of 44 positive by PCR ELISA (95%). These results demonstrate that immunofluorescence is a suitable, fast and inexpensive method for routine diagnostics. Additional sensitivity can be achieved by screening immunofluorescence-negative samples by PCR, which is extremely sensitive but time consuming and labor-intensive. PMID- 11275805 TI - Induction of matrix metalloproteinase-1 in in vitro experimental wound model using a novel three-dimensional culture system. AB - The aim of this study was to determine whether experimentally punched wounds may influence the connective tissue metabolism of human fibroblast in a three dimensional culture supplemented with L-ascorbic acid. This culture was designed for human dermal fibroblasts to organize a three-dimensional dermis-like structure by accumulating self-produced extracellular matrixes. In order to examine the effects of the wound, mRNA expression and production of type I collagen, matrix metalloproteinase-1 (MMP-1) and tissue inhibitors of metalloproteinase-1 (TIMP-1) were sequentially examined up to 72 hrs after the wound was created. The levels of mRNA expression of MMP-1 were higher at 12 and 24 hrs compared with the initial level and then gradually decreased in studies both with and without wounds. However, the levels of mRNA expression of MMP-1 between 6 to 48 hrs were higher in wounds than in non-wounds. The production of proMMP-1 was also significantly enhanced. There were no significant differences in the levels of mRNA expression or the production of type I collagen and TIMP-1 with the presence or absence of a wound. These results indicate that MMP-1 is synthesized to initiate the tissue restoration in response to the disruption of the three dimensional structure. Thus, this culture system provides a new experimental tool with which to examine the direct effect of mechanical changes on connective tissue metabolism in human dermal fibroblasts. PMID- 11275806 TI - Juvenile generalized circinate pustular psoriasis treated with oral cyclosporin A. AB - We report on a 17-year-old boy presenting with relapsing generalized circinate pustular psoriasis exacerbated by streptococcal angina. Because of the severe course in his case, we started systemic treatment with cyclosporin A and corticosteroids. Corticosteroids could easily be tapered down. Cyclosporin A maintenance therapy was realized with 100 mg/day (1.6 mg per kg body weight and day). Side effects were a temporary increase in blood pressure during initiation with 200 mg cyclosporin A/day. After dose reduction no side effects were seen. The pustular lesions disappeared and the PASI score decreased from 40.7 to 4.8. The treatment was found to be well tolerated and effective. PMID- 11275807 TI - Hyperpigmentation of the distal phalanx in healthy Caucasian neonates. AB - Transient alterations in pigmentation are frequently observed in black neonates, but to our knowledge, have not previously been reported in Caucasian infants. In 54 Caucasian newborns, we found at least mild periungual hyperpigmentation similar to the variation in coloration in the periungual region of many black newborns. This pigmentation should be added to the transient benign dermatoses of Caucasian infants. PMID- 11275808 TI - Linear drug eruption. AB - Linear eruptions are sometimes associated with systemic diseases and they may also be induced by various drugs. Paradoxically, such acquired inflammatory skin diseases tend to follow the system of Blaschko's lines. We describe a case of unilateral linear drug eruption caused by ibuprofen, which later became bilateral and generalized. PMID- 11275809 TI - Inflammatory cutaneous metastasis from laryngeal carcinoma. AB - The authors report the case of a 64-year-old man who presented with erythematous infiltrating plaques sited on his right supraclavicular and infraclavicular regions. Five years before a laryngeal epidermoid carcinoma had been surgically removed. After two years the patient underwent lymphadenectomy and radiotherapy for the presence of metastasis in his right cervical lymph nodes. He was disease free until two months before our examination when small red spots appeared on his right supraclavicular region, slightly enlarging. A skin biopsy was performed and histological examination showed the presence of metastatic cells inside dilated lymphatic vessels. Immunohistochemical markers showed only positive staining of atypical cells with monoclonal anti-cytokeratin antibodies (cytokeratin B ORTHO=34betaE12) confirming the epithelial origin of metastastic cells. A diagnosis of inflammatory cutaneous metastasis from laryngeal epidermoid carcinoma was given. This type of metastasis has been frequently observed in patients with breast carcinoma and it has also been described during the course of other malignant tumours. Malignant tumour of the larynx generally spreads to regional lymph nodes or, through blood, to the lungs. Skin metastasis have rarely been described, and always as multiple or solitary nodules. According to our knowledge this is the first case of inflammatory cutaneous metastasis from laryngeal epidermoid carcinoma reported in the literature. PMID- 11275810 TI - Discoid lupus erythematosus developing in areas where fragments of windshield glass had become embedded in the skin. AB - We observed a 26-year-old female patient with discoid lupus erythematosus on her left cheek skin, where fine fragments of windshield glass had been embedded in an automobile accident 8 years previously. She gradually developed general fatigue, morning stiffness of fingers and anti-nuclear antibody. As an etio-pathogenesis of this patient, we speculate that a long exposure to quartz (silica) could give rise to discoid lupus erythematosus in only local damaged areas and at the same time induce systemic immunological changes in some genetically restricted persons; such as production of anti-nuclear antibody. Silica has multi-potential biological effects, especially on immunological functions. PMID- 11275811 TI - Skin metastasis of breast cancer clinically undistinguished from amyopathic dermatomyositis. AB - We report a 65-year-old woman who consulted us on May 25, 1998, showing pruritic, partially flagellate erythema on the back and upper extremities, livedo lesions with erythema on the loins, and erythematous papules on the dorsal finger joints for 2 months. Histopathological findings of erythema on the back showed mononuclear cell infiltration around capillaries and marked edema in the dermis. Laboratory data were within normal range except for positive anti-nuclear antibody. She had undergone total left mastectomy on June 2, 1997 for breast cancer. Supraclavicular lymph node metastasis was found at the beginning of May, 1998. A diagnosis of amyopathic dermatomyositis associated with breast cancer was made. Erythema with itching gradually subsided from the end of August, 1998. Treatment with radiation and chemotherapy reduced lymph node swelling, but complete remission was not obtained. Erythema similar to the previous lesion but without itching re-appeared on the back from January, 2000. Histological findings of erythema showed many carcinoma cells similar to the primary lesion of left breast cancer in the whole dermis. A diagnosis of skin metastasis of breast cancer was made. These findings suggest that skin metastasis should be taken into account for patients with erythema on the trunk similar to dermatomyositis. PMID- 11275812 TI - Delayed adverse reaction to sodium ioxaglic acid-meglumine. AB - A patient had a maculopapular rash, fever, hepatic cytolysis, rhabdomyolysis, eosinophilia and raised total IgEs after coronarography with ioxaglic acid meglumine (Hexabrix). Intradermal test to ioxaglic acid-meglumine was positive 48 hrs later. Histological examination revealed perivascular lymphocytic infiltration, with predominantly CD45Ro+ and CD8+ T cells and apoptotic basal keratinocytes. This case documents a late hypersensitivity reaction to ioxaglic acid-meglumine. PMID- 11275814 TI - Pemphigus of the eyelids. AB - We report the case of a 56-year-old woman who presented with a 2-month history of widespread oral erosion and a 3-day history of small papules on the lower eyelids. No other skin involvement was found. Histopathological examination revealed suprabasal cleft and acantholysis in the lower epidermis of the papule on the lower eyelid and in the lower mucous membrane of the oral mucosa. Intercellular deposits of IgG and C3 were seen in the whole epidermis of the specimen from the papule on the right lower eyelid by direct immunofluorescence study. These deposits were also observed in the biopsy specimen from erosion on the left buccal membrane. Indirect immunofluorescence study using normal human skin as a substrate showed intercellular antibodies directed to the cell surface of the whole epidermis with a titer of 1:40. The titers of antibodies to desmoglein 3 and 1 were 118 and 25.9, respectively, by enzyme-linked immunosorbent assay. The patient was treated with an oral administration of prednisolone (0.75 mg/kg/day) for 9 days, which improved the skin eruptions and oral erosion. The dose of prednisolone was gradually tapered and it took 10 weeks to cease this treatment. These findings suggest that this patient is an unusual case of pemphigus vulgaris (mucosal dominant type) diagnosed from the clinical and histopathological findings, with positive antibodies to desmoglein 3 and 1. PMID- 11275813 TI - Subcutaneous nodules following treatment with aluminium-containing allergen extracts. AB - We describe two patients who developed multiple itching nodules following immunization with vaccines adsorbed on aluminium hydroxide. Both patients had been treated with vaccines for extrinsic asthma and rhinitis for 4 and 10 years respectively. The lesions were persistent and lasted for several years. Histopathological findings were those of a foreign body reaction. Aluminium was most probably involved in the pathogenesis of these lesions because its presence could be demonstrated in macrophages using energy-dispersive X-ray microanalysis. Although some symptomatic relief was achieved with topical corticosteroids and oral antihistamines, treatment was unsuccessful. PMID- 11275815 TI - Guess what. Dermatofibrosarcoma protuberans presenting as an atrophic red plaque. PMID- 11275816 TI - Guess what. B-cell acute lymphoblastic leukemia with aleukemic leukemia cutis. PMID- 11275817 TI - Guess what. Perifolliculitis capitis abscedens et suffodiens. PMID- 11275818 TI - Guess what. Classic Kaposi's sarcoma. PMID- 11275819 TI - Understanding nail disorders. AB - Diagnosing nail dystrophies and understanding their pathogenesis is best accomplished if the origin of the disorder is first established. Exogenous. In this group, the nail plate is altered by exogenous factors and is the site of primary involvement: periungual tissues may be affected secondarily. Endogenous. In this group, the nail plate is altered secondarily by changes occurring in the matrix, proximal nail fold, nail bed, hyponychium or underlying bony phalanx. PMID- 11275820 TI - Photoaging: a daily care. Introduction. PMID- 11275821 TI - What is photoaged skin? PMID- 11275822 TI - Dyspigmentation of aged skin. PMID- 11275823 TI - The role of UVA rays in skin aging. PMID- 11275824 TI - Topical vitamin C in the treatment of photoaged skin. PMID- 11275825 TI - Photoaging: a daily care. Conclusions and perspectives. PMID- 11275826 TI - Critical pediatric equipment availability in Canadian hospital emergency departments. AB - STUDY OBJECTIVE: Of all child visits to emergency departments, 1% to 5% involve critically ill children who require cardiopulmonary resuscitation. Numerous versions of pediatric equipment lists for EDs have been published. Despite these efforts, many EDs remain unprepared for pediatric emergencies. The objectives of this study were to assess the availability of pediatric resuscitation equipment items in Canadian hospital EDs and to identify risk factors for the unavailability of these items. METHODS: Using the updated database of the Canadian Association of Emergency Physicians (CAEP), a questionnaire survey was sent to 737 Canadian hospital EDs with a maximum of 3 mailings to nonresponders. On-site visits to a selected subset of hospital EDs were completed to validate the results obtained by the mailed questionnaire. RESULTS: The response rate was 88.3% (650/737). Results showed the following overall equipment unavailability: intraosseous needle, 15.9%; pediatric drug dose guidelines, 6.6%; infant blood pressure cuff, 14.8%; pediatric defibrillator paddles, 10.5%; infant warming device, 59.4%; infant bag-valve-mask device, 3.5%; infant laryngoscope blade, 3.5%; 3-mm endotracheal tube, 2.5%; and pediatric pulse oximeter, 18.0%. Low percentage of pediatric visits, lack of an on-call pediatrician for the ED, and lack of a pediatric advanced life support-trained physician on staff were independently associated with equipment unavailability. CONCLUSION: This study demonstrated that essential pediatric resuscitation equipment is unavailable in a disturbingly high number of EDs across Canada and has identified several determinants of this unavailability. PMID- 11275827 TI - Critical evaluation of a CLIA-waived streptococcal antigen detection test in the emergency department. AB - STUDY OBJECTIVE: We compare the performance of a Clinical Laboratory Improvement Amendments (CLIA)-waived antigen detection test (ADT) analyzed in the emergency department and a CLIA moderately complex ADT performed in the hospital microbiology laboratory. METHODS: Samples from throat swabs were obtained using a double-headed Culturette II (Becton Dickinson Medical Systems, Sparks, MD) from a consecutive sample of 322 patients 3 years or older who presented to the ED of a university-affiliated pediatric referral hospital with the complaint of sore throat during 1998. One swab was transported to the microbiology laboratory and analyzed using a CLIA moderately complex ADT; negative results were confirmed using sheep blood agar culture. The second swab remained in the ED where a nurse conducted a CLIA-waived ADT. The accepted standard for documentation of group A beta-hemolytic streptococcal (GABHS) infection was either a positive moderately complex ADT or culture result. The time of specimen collection, as well as the time the ED results and microbiology laboratory results were available to treating physicians, were recorded. Main outcome measures were concordance (kappa statistic), sensitivity, and turnaround time (Mann-Whitney U test). RESULTS: Three hundred twenty-two patients (mean age 7.5 years) had both ADTs performed. One hundred one (31%) patients had documented GABHS in the microbiology laboratory; 83 (82%) had a positive ADT result in the microbiology laboratory, and 18 (18%) had a positive culture result after a negative moderately complex ADT result. In 299 patients or 93% (95% confidence interval [CI] 90.8%, 95.8%) of patients, the waived ADT and the moderately complex ADT results were concordant (kappa 0.82; 95% CI 0.78, 0.86; P <.001). The sensitivity of the waived ADT was 80%; the sensitivity of the moderately complex ADT approximated 82% (difference of 2%; 95%CI -3%, 7%). The median times from swab specimen collection to availability of ADT results were 10 minutes (range 3 to 37 minutes) for the waived ADT and 35 minutes (range 5 to 162 minutes) for the moderately complex ADT (P <.001) with a difference of 25 minutes (95% CI 22.4, 27.6 minutes). CONCLUSION: In this study, an ED CLIA-waived rapid streptococcal throat test performed as well as its equivalent CLIA-regulated laboratory test. Further, the ED test provided results more rapidly than the laboratory test. Our results also validate previous work that negative rapid throat test results in pediatric patients in the ED should be confirmed by standard throat culture. PMID- 11275828 TI - Do parents value drowning prevention information at discharge from the emergency department? AB - STUDY OBJECTIVE: We determined parent recall and perceived usefulness of drowning prevention messages included in routine computer-generated discharge instructions. METHODS: All pediatric emergency department patients' computerized discharge instructions included 3 prevention messages: wear a life vest, swim in safe areas, and do not drink alcohol while swimming or boating. Parents were telephoned 1 to 2 weeks after the visit and asked to recall the prevention messages and rate the usefulness of the instructions. Responses were linked with patient characteristics and ED visit variables (day and time of visit, duration of ED visit, severity of condition, diagnostic category, number of tests, and treatments). RESULTS: Of 914 parents who were contacted, 795 were eligible. Of those, 619 (78%) completed the interview. Fifty percent of parents recalled receiving drowning prevention information; of these, 41% recalled unaided the life vest messages and 35% of 155 parents who did not own a life vest stated they would subsequently consider buying their child a life vest. Most (88%) rated the prevention information useful or very useful. No patient or visit variables were associated with usefulness ratings. CONCLUSION: Written injury prevention messages with discharge instructions were well received by parents of children in a pediatric ED. The ED may be a setting where families could receive injury prevention education. PMID- 11275829 TI - Deciphering the authorship code. PMID- 11275830 TI - Pediatric equipment availability and emergency preparedness. PMID- 11275831 TI - Guidelines for preparedness of emergency departments that care for children: a call to action. PMID- 11275833 TI - Emergency medicine in Eastern Europe: the Hungarian experience. AB - The past decade has brought numerous dramatic changes in health care reform to Hungary. The area of acute care medicine has only recently been recognized as an important component in securing proper health care for Hungarians. Emergency medicine as a recognized and accepted specialty is only now beginning to get peer, governmental, and public support. This article describes the past, present, and future of emergency medicine in Hungary and is offered as a model for other Eastern European countries in similar sociopolitical situations. PMID- 11275832 TI - Benign paroxysmal positional vertigo: diagnosis and treatment in the emergency department--a review of the literature and discussion of canalith-repositioning maneuvers. AB - Dizziness is a frequent presenting complaint in emergency department patients. Although seen in patients of all ages, it is more prevalent in patients older than 50 years of age. Vertigo represents a subset of dizziness and is defined as an illusion of movement, usually rotational, of the patient or the patient's surroundings. The illusion of motion may be of oneself (subjective vertigo) or of external objects (objective vertigo). The emergency physician should consider a large differential in the evaluation of vertigo with special attention to whether the vertigo is central or peripheral in origin. PMID- 11275834 TI - New-onset absence status epilepsy presenting as altered mental status in a pediatric patient. AB - Absence status epilepsy (ASE) is an uncommon seizure disorder in children. The primary presentation of new-onset ASE in a pediatric patient is an unusual cause of altered mental status in the emergency department. We describe a previously healthy 8-year-old child who presented with an acutely altered mental state. The patient was awake but confused, with a fluctuating level of alertness and an inability to perform simple routine tasks. The results of general physical and neurologic examination, with the exception of mental status, were normal. Head computed tomography and laboratory test results were normal. Electroencephalographic testing revealed seizure activity consistent with ASE. Administration of intravenous diazepam caused cessation of seizure activity and a return to the patient's baseline mental function. Although rare, ASE should be considered in the differential diagnosis of altered mental status in children. PMID- 11275835 TI - Children in car seats and in the backseat. PMID- 11275836 TI - Commentary: We need to give children a boost before we buckle them. PMID- 11275838 TI - Sympathetic dystrophy. PMID- 11275839 TI - Thyrotoxic hypokalemic periodic paralysis reversed by propranolol without rebound hyperkalemia. PMID- 11275840 TI - Practice model for emergency medicine in both out-of-hospital and in-hospital settings. PMID- 11275841 TI - Octreotide as antidote for sulfonylurea-induced hypoglycemia. PMID- 11275843 TI - Postreduction radiographs for anterior shoulder dislocation: a reappraisal. PMID- 11275847 TI - Care of children in the emergency department: guidelines for preparedness. PMID- 11275845 TI - Human error in medicine: promise and pitfalls, part 2. PMID- 11275850 TI - President's message. The art of mentoring. PMID- 11275851 TI - Your life in your hands: on finding causes, not just treating, breast cancer. PMID- 11275852 TI - One new graduate's experience. PMID- 11275853 TI - Hospitals should replace "sign on bonuses" with "retention bonuses". PMID- 11275854 TI - The lecture I want to give. PMID- 11275855 TI - More on ED nurse practitioners' role. PMID- 11275858 TI - A 29-year-old soldier with heat stroke. PMID- 11275859 TI - The effect of standard care, ibuprofen, and music on pain relief and patient satisfaction in adults with musculoskeletal trauma. AB - OBJECTIVE: The purposes of this study were to determine the most effective nursing intervention to decrease pain for patients with minor musculoskeletal trauma and moderate pain at triage and to examine patient satisfaction. METHODS: Patients were assigned to 1 of 3 intervention groups: (1) standard care (ice, elevation, and immobilization); (2) standard care and ibuprofen; or (3) standard care and music distraction. Patients were monitored for pain ratings for 60 minutes. Patients who sustained minor musculoskeletal trauma within the past 24 hours and presented with pain ratings of 4 or greater were included. Two patient satisfaction questions were asked upon discharge from the emergency department. RESULTS: Seventy-seven patients met the inclusion criteria. No differences in pain ratings between groups were demonstrated. A statistically significant reduction in pain for all patients occurred at 30 minutes (F = 16.18, P <.01) and was maintained at 60 minutes. However, 70% of patients continued to report pain ratings of 4 or greater (on a scale of 1 to 10) at 60 minutes. The reduction in pain was not found to be clinically significant.Eighty-four percent of patients stated that they were more satisfied with their overall care in the emergency department because of the immediate attention to pain relief they received at triage. No differences in satisfaction existed between treatment groups, although patients who reported higher pain ratings expressed statistically significant lower satisfaction with pain management scores (F = 9.375, P =.003). CONCLUSION: None of the therapies-standard care (ice, elevation, immobilization), standard care with ibuprofen, or standard care with music distraction-provided clinically significant pain relief to patients who had minor musculoskeletal trauma (ie, sprains and fractures) and moderate pain at triage. Interestingly, satisfaction scores were sometimes positive, even when pain was not relieved. PMID- 11275860 TI - Livestock trauma in central Texas: cowboys, ranchers, and dudes. AB - INTRODUCTION: Emergency nurses routinely treat a wide variety of animal-related injuries, yet the trauma literature rarely addresses injuries associated with livestock. METHODS: The purpose of this retrospective, descriptive study was to examine the incidence, demographics, severity, and other factors associated with trauma related to livestock in persons admitted to one central texas trauma center during a 5-year period. Patients were classified by activity at the time of injury. The following 5 categories were identified: cowboy, rancher, dude, standing near, and unidentified. Groups were further analyzed by the type of animal involved in the injury, mechanism of injury, patient sex and age, body regions injured, indicators of severity, and county in which the injury occurred. RESULTS: One hundred thirty-six patients met the inclusion criteria. Of these patients, 24% were categorized as cowboys, 18% as ranchers, and 39% as dudes. Six percent were standing near the animal, and activity at the time of injury could not be adequately determined for 13% of patients. Typical demographics, mechanisms of injury, wounds, indicators of severity, and animals were found to be associated with each activity class. DISCUSSION: Horses were responsible for 75% of the traumas sustained by ranchers and 83% of the traumas sustained by dudes. Bulls were responsible for 94% of the cases involving cowboys. Simply standing near livestock was dangerous for 6% of the sample, including young children. Although helmets and steel-toed boots are currently unpopular, wearing them is a simple and important safety strategy. PMID- 11275862 TI - Disabling back injuries in nursing personnel. PMID- 11275861 TI - Intimate partner violence screening and intervention: data from eleven Pennsylvania and California community hospital emergency departments. AB - OBJECTIVE: To provide clinical practice recommendations for screening and interventions for intimate partner violence (IPV) in ED settings. SETTING: Eleven mid-sized community-level hospital emergency departments (20,000 to 40,000 admissions annually) in Pennsylvania and California. PARTICIPANTS: All women (4641) aged 18 years or older who came to the emergency department during 309 selected shifts. METHODS: An anonymous survey inquiring about physical, sexual, and emotional IPV was conducted from 1995 through 1997. In addition, medical records were abstracted for every woman (18 years and older) seen in the 11 participating emergency departments during the study period. RESULTS: The vast majority of both abused and nonabused women supported routine screening for IPV; however, fewer than 25% of women said they were asked about IPV by ED staff. ED screening rates for IPV were higher among women who came to the emergency department because of acute trauma from abuse (39%) than for women who reported that they had been abused within the past year (13%). The prevalence of past year and lifetime IPV was significantly higher when the questionnaire was self administered than when it was administered by a nurse. More than a third of women who had recently been abused and 76% of women who acknowledged experiencing physical or sexual IPV within the past year reported that they did not come to the emergency department for treatment of an injury. Although the majority of women (76% to 90%) agreed with the concept of health care providers reporting IPV to the police, women abused recently were significantly less likely to support this practice. CONCLUSION: The study provides evidence supporting standard protocols for routine screening for IPV among women who come to emergency departments and chart prompts for both screening and interventions. These actions are acceptable to the majority of both abused and nonabused women seen in the emergency department and should be considered in systematic repeated training of health care professionals in emergency departments. This information is important for health care providers who are seeking to improve their identification of and care for abused women. PMID- 11275863 TI - Knowledge assessment and preparation for the Certified Emergency Nurses Examination. PMID- 11275864 TI - Trauma death of a 28-year-old: Two clinicians help a family to view the body and keep a lock of hair. PMID- 11275865 TI - An informal discussion among emergency nurses about their current clinical practice: what's new and what works. PMID- 11275867 TI - Teachable moments: a paradigm shift. PMID- 11275868 TI - Patient wins EMTALA appeal: case underscores that ED documentation of admission/care refusal is crucial. PMID- 11275869 TI - Cutting edge discussions of management, policy, and program issues in emergency care. PMID- 11275870 TI - ED learning units: an innovative teaching method. PMID- 11275871 TI - A critical neonate in the emergency department. PMID- 11275872 TI - ED staff's reporting of impaired drivers: understanding the issues, continuing the work. PMID- 11275873 TI - Overcoming a consent defense to sexual assault. PMID- 11275874 TI - Falls in the elderly: making the difference. PMID- 11275875 TI - A man with wrist pain after a fall. PMID- 11275876 TI - On being a nurse. PMID- 11275878 TI - Radiofrequency energy delivery to the gastroesophageal junction for the treatment of GERD. AB - BACKGROUND: In this multi-center study, the feasibility, safety, and efficacy of radiofrequency (RF) energy delivery to the gastroesophageal junction (GEJ) for the treatment of gastroesophageal reflux disease (GERD) were investigated. METHODS: Forty-seven patients with classic symptoms of GERD (heartburn and/or regurgitation), a daily anti-secretory medication requirement, and at least partial symptom response to drugs were enrolled. All patients had pathologic esophageal acid exposure by 24-hour pH study, a 2 cm or smaller hiatal hernia, grade 2 or less esophagitis, and no significant dysmotility or dysphagia. RF energy was delivered with a catheter and thermocouple-controlled generator to create submucosal thermal lesions in the muscle of the GEJ. GERD symptoms and quality of life were assessed at 0, 1, 4, and 6 months with the short-form health survey (SF-36). Anti-secretory medications were withdrawn 7 days before each assessment of symptoms and pH/motility study. Medication use, endoscopic findings, esophageal acid exposure, and motility were assessed at 0 and 6 months. RESULTS: Thirty-two men and 15 women underwent treatment. At 6 months there were improvements in the median heartburn score (4 to 1, p < or = 0.0001), GERD score (26 to 7, p < or = 0.0001), satisfaction (1 to 4, p < or = 0.0001), mental SF-36 (46.2 to 55.5, p = 0.01), physical SF-36 (41.1 to 51.9, p < or = 0.0001), and esophageal acid exposure (11.7% to 4.8%, p < or = 0.0001). Esophagitis was present in 25 patients before treatment (15 grade 1 and 10 grade 2) and 8 had esophagitis at 6 months (4 grade 1 and 4 grade 2, p = 0.005). At 6 months, 87% no longer required proton pump inhibitor medication. There was no significant change in median lower esophageal sphincter pressure (14.0 to 12.0 mm Hg, p = 0.19), peristaltic amplitude (64 to 66 mm Hg, p = 0.71), or lower esophageal sphincter length (3.0 to 3.0, p = 0.28). There were 3 self-limited complications (fever for 24 hours, odynophagia lasting for 5 days, and a linear mucosal injury that was healed after 3 weeks). CONCLUSION: RF energy delivery significantly improved GERD symptoms, quality of life, and esophageal acid exposure while eliminating the need for anti-secretory medication in the majority of patients with a heterogeneous spectrum of clinical disease severity but with minimal active esophagitis or hiatal hernia. PMID- 11275879 TI - Transoral, flexible endoscopic suturing for treatment of GERD: a multicenter trial. AB - BACKGROUND: A totally transoral outpatient procedure for the treatment of GERD would be appealing. METHODS: A multicenter trial was initiated that included 64 patients with GERD treated with an endoscopic suturing device. Inclusion criteria were 3 or more heartburn episodes per week while not taking medication, dependency on antisecretory medicine, and documented acid reflux by pH monitoring. Exclusion criteria were dysphagia, grade 3 or 4 esophagitis, obesity, and hiatus hernia greater than 2 cm in length. Patients underwent manometry, endoscopy, 24-hour pH monitoring, and symptom severity scoring before and after the procedure. Patients were randomized to a linear or circumferential plication configuration. Adverse procedural events were recorded. RESULTS: Mean 6-month symptom score changes demonstrated procedural efficacy. Heartburn severity and frequency as well as regurgitation all improved (p > 0.0001 for each). Twenty four-hour pH monitoring showed improvement in number of episodes below pH of 4 at 3 and 6 months (p < 0.0007 and 0.0002) and percentage of total time the pH was less than 4 at 6 months (p < 0.011). Plication configuration did not affect symptoms or pH monitoring results. One patient had a self-contained suture perforation that was successfully treated with antibiotics. CONCLUSION: Endoscopic gastroplasty is safe. It is associated with reduced symptoms and medication use at 6 month follow-up in patients with uncomplicated GERD. PMID- 11275880 TI - Endoscopic implantation of Plexiglas (PMMA) microspheres for the treatment of GERD. AB - BACKGROUND: A gelatinous implant containing polymethylmethacrylate (PMMA) beads is successfully used to augment the diminished thickness of the chorium in patients with skin defects and wrinkles. The aim of the present study was to determine whether submucosal injection of PMMA microspheres into the lower esophageal folds decreases the severity of symptoms and acid reflux in patients with GERD. METHODS: Endoscopic submucosal implantation of PMMA was carried out in 10 patients with GERD who were either refractory to or dependent on proton pump inhibitors. Symptom severity score, 24-hour pH monitoring, upper GI endoscopy, and EUS were performed to evaluate the efficacy of implantation. RESULTS: A significant decrease in the symptom severity score and mean total time with esophageal pH less than 4 was noted after the implantation of PMMA (p < 0.05). Seven of 10 patients were taking no medication after PMMA implantation. There were no serious procedure-related complications. CONCLUSIONS: Endoscopic implantation of PMMA into the submucosa of the lower esophageal folds may be a new method for treating GERD. Further studies are required to determine the long term efficacy of the procedure. PMID- 11275881 TI - Endoscopic hemoclipping for upper GI bleeding due to Mallory-Weiss syndrome. AB - BACKGROUND: Endoscopic hemoclipping is known to be highly effective as hemostatic treatment for upper gastrointestinal bleeding. However, the efficacy and safety of hemoclipping for Mallory-Weiss syndrome (MWS) have not been reported. Thus, the aim of the present study was to assess prospectively the usefulness of endoscopic hemoclipping for MWS bleeding. METHODS: This study was conducted from January 1994 to August 1999. Hemoclipping was performed when active bleeding (spurting, streaming or oozing), visible vessels or fresh adhesive clots were found on endoscopic examination. Patients who did not have any of these findings were conservatively treated. Follow-up endoscopy was performed within 24 hours, after 5 days and between 1 and 2 months after the procedure. RESULTS: MWS was diagnosed in a total of 58 patients during the study. Hemoclipping was performed in 26 patients and was technically successful in all cases. The average number of hemoclips used was 2.8 +/- 1.6 (range 1 to 8). The number of hemoclips required for hemostasis depended on the nature of the bleeding. No complications, recurrent bleeding, or deaths resulted. Follow-up endoscopy showed no evidence of hemoclip-induced tissue injury and no impairment of Mallory-Weiss tears. CONCLUSION: Endoscopic hemoclipping provided an effective and safe modality for obtaining hemostasis when bleeding is due to MWS. PMID- 11275882 TI - Effect of antibiotic-loaded hydrophilic stent in the prevention of bacterial adherence: a study of the charge, discharge, and recharge concept using ciprofloxacin. AB - BACKGROUND: Ciprofloxacin prophylaxis significantly prolonged stent patency in cats, but human studies produced conflicting results, possibly due to varying drug levels in bile. The uptake (charge) and release (discharge) of ciprofloxacin from a hydrophilic stent (HS) in an antibiotic solution and the effect of a ciprofloxacin-loaded stent (CHS) in inhibiting Escherichia coli adherence were tested. The adjuvant effect of ciprofloxacin perfusion (recharge) in the inhibition of E coli adherence was also tested. METHODS: Uptake: segments of HS were immersed in 5 mL of ciprofloxacin solutions for 24 hours. Ciprofloxacin remaining in solution was measured to determine the uptake by the HS. Release: CHS were placed in 5 mL water for 24 hours, and released ciprofloxacin was measured. CHS were placed on culture plates with E coli and incubated; diameters of inhibited zones were measured. CHS 0.5 cm in length were incubated in separate 5 mL E coli suspension (10(7) colony forming units [CFU]/mL) in 2% ox bile for 4 hours. E coli adhered on CHS were measured and compared with control HS. An E coli (10(6) CFU/mL) suspension was perfused through a modified Robbins device (MRD)-containing CHS. Stents were removed at regular intervals and processed to determine the adherence of E coli; non-loaded HS served as controls. The experiment was repeated by using CHS together with perfusion of ciprofloxacin solution (0.3 microg/mL) into the MRD for up to 7 days; normal saline solution was used as a control in a second MRD. Stents were removed daily to determine the adherence of E coli. RESULTS: Uptake and release of ciprofloxacin by HS and CHS, respectively, were related to concentration of ciprofloxacin. Between 50% to 90% of the drug was released in 24 hours. Zonal inhibition of E coli growth was proportional to the concentration of ciprofloxacin on the CHS. There was an initial 10-fold reduction in attached E coli on CHS compared with controls, but this effect diminished after 24 hours. With ciprofloxacin perfusion, there was a 100-fold reduction in adhered E coli on CHS, although there was no change in E coli concentration in bile. CONCLUSIONS: There was a free exchange (uptake and release) of ciprofloxacin along a concentration gradient between the antibiotic solution and HS. CHS reduced the number of adhered E coli, but the effect was short-lived. Perfusion of ciprofloxacin offers an adjuvant benefit by enhancing inhibition of E coli adherence on CHS. PMID- 11275883 TI - Is non-Helicobacter pylori, non-NSAID peptic ulcer a common cause of upper GI bleeding? A prospective study of 977 patients. AB - BACKGROUND: Non-Helicobacter pylori, non-NSAID ulcer is relatively common in Western countries. Whether it is a significant problem in the Orient is unclear. The aim of this study was to investigate the incidence of non-H pylori, non-NSAID ulcers presenting with GI bleeding. METHODS: A prospective study was done of 1675 consecutive patients presenting with upper GI bleeding over a period of 12 months. Upper endoscopy was performed with biopsy specimens taken from the antrum and body of the stomach for a biopsy urease test (BUT) and histology for detection of H pylori. Exposure to nonsteroidal anti-inflammatory drugs (NSAID) or aspirin within 4 weeks of hospitalization was carefully scrutinized. A 6-week course of treatment with an H2-receptor antagonist was prescribed for patients who did not use an NSAID and had a negative BUT result. Follow-up endoscopy was performed to confirm H pylori status with a BUT and histology. Positive histology at either initial or follow-up endoscopy was used as the standard for diagnosing H pylori infection. RESULTS: Among 977 patients who were found to have ulcer bleeding, 434 (44%) had exposure to aspirin or an NSAID. Of the 543 non-NSAID users, 431 (79.4%) had a positive BUT and 112 (20.6%) were BUT negative on initial endoscopy. Eighty-nine of 112 patients who were NSAID negative, BUT negative returned for follow-up endoscopy. Forty-nine of 89 (55.1%) were found to have a positive BUT and positive histology at follow-up endoscopy. Only 40 of 977 (4.1%) patients admitted with ulcer bleeding were confirmed to have non-H pylori, non-NSAID ulcers. CONCLUSIONS: Non-H pylori, non-NSAID bleeding ulcer is uncommon. A negative BUT is unreliable for exclusion of H pylori infection during the acute phase of ulcer bleeding. PMID- 11275884 TI - First endoscopic-histologic follow-up in patients with body-predominant atrophic gastritis: when should it be done? AB - BACKGROUND: Body-predominant atrophic gastritis is considered a risk factor for gastric cancer and carcinoid. Timing of follow-up for patients with this disorder has not been defined. This study was undertaken to determine the optimal time for the first endoscopic/histologic follow-up in patients with body-predominant atrophic gastritis. METHODS: Forty-two patients with body-predominant atrophic gastritis were randomly assigned to 1 of 2 follow-up intervals: group A (n = 22) at 24 months and group B (n = 20) at 48 months. At baseline and follow-up patients underwent gastroscopy at which biopsies were obtained from the antrum and body for histopathology and evaluation for enterochromaffin-like cells. RESULTS: In group A patients, 2 antral hyperplastic polyps (9.1%) were present at baseline and 4 antral hyperplastic polyps (18.2%) were found at follow-up. In group B patients, baseline gastroscopy revealed 2 antral hyperplastic polyps (10%) and follow-up 2 antral hyperplastic polyps (10%) and 1 carcinoid tumor (5%) in the body. Atrophy and intestinal metaplasia scores in gastric body and antral mucosa in both groups did not change significantly between baseline and follow up, except an increase in antral mucosa atrophy in group B patients (p = 0.02) was revealed. CONCLUSIONS: The results of this study indicate that performing the first follow-up in patients with body-predominant atrophic gastritis need not be earlier than at 4 years after diagnosis. This interval is satisfactory for detection of potential neoplastic lesions. PMID- 11275885 TI - Prospective study of laparoscopic findings with regard to the development of hepatocellular carcinoma in patients with hepatitis C virus-associated cirrhosis. AB - BACKGROUND: The laparoscopic findings of hepatocarcinogenesis are not well defined. The purpose of this prospective study was to evaluate the predictive value of laparoscopic findings with regard to the development of hepatocellular carcinoma (HCC) in patients with hepatitis C virus (HCV)-associated cirrhosis. METHODS: One hundred nineteen patients with HCV-associated cirrhosis who underwent laparoscopy were enrolled in this study. Ten laparoscopic variables, including degree of development and size of regenerating nodules, irregularity of regenerating nodules, and size of both hepatic lobes, were measured. The predictive value of each finding for the development of HCC was investigated by using univariate and multivariate analyses. RESULTS: HCC developed in a total of 42 of the 119 patients (35.3%) during a mean follow-up period of 62.9 months. The degree of regenerating nodules, the presence of irregular regenerative nodules, and atrophic right lobe were significant predictive factors for HCC with univariate analysis. In particular, 67.1% of patients with irregular regenerative nodules had HCC develop within 5 years. Multivariate analysis revealed that irregular regenerative nodules (relative risk 6.32, p = 0.012), the degree of regenerative nodules (relative risk 4.78, p = 0.029), and atrophic right lobe (relative risk 3.87, p = 0.048) were independent predictive factors. CONCLUSIONS: Morphologic observation of the liver surface by laparoscopy is important and provides information on factors that are statistically significant early predictors of the development of HCC. PMID- 11275886 TI - Efficacy of high-level disinfectants for reprocessing GI endoscopes in simulated use testing. AB - BACKGROUND: There has been recent public concern regarding the adequacy of current practices for flexible endoscope reprocessing. High-level disinfection is defined by the Food and Drug Administration (FDA) as a minimum of 6-log reduction of mycobacteria under a worst-case scenario. Several agents are currently approved by the FDA, but published data on their relative efficacies against mycobacteria are lacking. The objective of this study was to determine the efficacy of these agents for high-level disinfection. METHODS: In simulated-use testing, video endoscopes (5 colonoscopes and 5 duodenoscopes) were each inoculated with 9.0 x 10(7) colony-forming units of Mycobacterium chelonae. Cleaning was performed by using a standardized protocol. Each endoscope was then subjected to chemical disinfection with Cidex (2.0% glutaraldehyde) at 20 degrees C for 20 minutes, Sporox (7.5% hydrogen peroxide) at 20 degrees for 30 minutes, and Steris 20 (0.2% peracetic acid) at 50 degrees C to 56 degrees C for 12 minutes using the Steris System 1 processor. Although not FDA-approved, tests were also conducted by using 70% isopropyl alcohol at 20 degrees C for 20 minutes. These results were compared with disinfection with ethylene oxide gas. All channels were sampled for M chelonae before and after manual cleaning and after disinfection. RESULTS: Cleaning alone resulted in an average log reduction of 3. Cidex, Sporox, Steris 20, ethylene oxide gas, and isopropyl alcohol, in combination with manual cleaning, each achieved a 6-log or greater reduction of the mycobacterial inoculum. No organisms were recovered from any channel after reprocessing with ethylene oxide and Steris 20. CONCLUSIONS: Commercially available high-level disinfectants are equally efficacious for reprocessing flexible GI endoscopes when used in conjunction with cleaning and in accordance with recommended guidelines. PMID- 11275887 TI - EUS predictors of long-term survival in esophageal carcinoma. AB - BACKGROUND: EUS is the most accurate nonsurgical modality for the staging of esophageal cancer, but the ability of EUS to predict outcomes or prognosis is unclear. Patients were examined who had EUS performed for esophageal cancer staging to determine which endosonographic features predict survival. METHOD: Data on 203 patients undergoing EUS for esophageal cancer staging were studied retrospectively. Median survival was calculated for each T-stage and N-stage and according to the presence or absence of celiac axis (CAx) lymphadenopathy as determined by EUS. Kaplan-Meier survival curves were generated for each stage and the log-rank test was used to test for significant differences in survival. Multivariate analysis was performed to test for the relative importance in predicting survival of the EUS stages, also considering age, gender, histology, and type of treatment. RESULTS: Significant differences were found in the ability of EUS-determined T-stage (p = 0.0005), N-stage (p < 0.0001), and presence of CAx nodes (p = 0.0049) to predict survival. Multivariate analysis showed N-stage to predict survival. CONCLUSIONS: Pretreatment EUS can predict survival in esophageal cancer based on initial T-stage, N-stage, and the presence of CAx nodes. The presence of lymphadenopathy at EUS is an important predictor of survival. EUS should be performed in all patients with esophageal cancer, not only for staging patients before therapy, but also as a valuable method of determining prognosis. PMID- 11275888 TI - Assessment of complications of EUS-guided fine-needle aspiration. AB - BACKGROUND: EUS-guided fine-needle aspiration (EUS-FNA) permits both morphologic and cytologic analysis of lesions within or adjacent to the GI tract. Despite increasing use of this technique, the safety and overall complication rates remain poorly defined. METHODS: During a period of 20 months, 322 consecutive patients underwent EUS-FNA in 2 centers. All procedures were performed with the patients under general anesthesia. All complications (including local complications resulting from endoscopy/aspiration or clinical complications after the procedure) were evaluated. Potential risk factors for the development of complications were also analyzed including site and nature of the lesion, presence of portal hypertension, and number of needle passes. RESULTS: A total of 345 lesions were aspirated in 322 patients. EUS-FNA involved the pancreas in 248 cases. Pancreatic lesions included solid (134) and cystic (114) types, which required a mean of 2.5 and 1.4 needle passes, respectively. Complications were observed in 4 (1.2%) patients after aspiration of pancreatic cystic lesions (acute pancreatitis, n = 3; aspiration pneumonia, n = 1) and all cases of pancreatitis resulted from FNA of lesions in the head/uncinate process. No complications resulted from FNA of solid pancreatic lesions. Complications were not observed after FNA of lymph nodes (n = 62) and one case of aspiration pneumonia was observed after FNA of a stromal tumor. EUS-FNA was performed without complication in 16 patients (5%) with portal hypertension. The number of needle passes was not predictive of complications. CONCLUSIONS: Because the overall risk of complications from EUS-FNA was relatively low (1.6%) with no severe or fatal incidents and although the risk appears slightly higher than that for standard EUS alone, the safety of EUS-FNA appears acceptable based on this analysis from an experienced center. PMID- 11275890 TI - EUS-guided fine-needle aspiration combined with flow cytometry and immunocytochemistry in the diagnosis of lymphoma. AB - BACKGROUND: Limited information is available regarding the use of EUS-guided fine needle aspiration (EUS-FNA) in the diagnosis of lymphoproliferative disorders. The aim of this study was to evaluate the yield of this technique in the primary diagnosis of lymphoma. METHODS: The records were reviewed of 38 consecutive patients with GI lesions and/or enlarged lymph nodes identified on imaging studies that raised a suspicion of lymphoma who underwent EUS-FNA of lymph nodes or the gut wall. Final diagnosis was based on clinical follow-up, imaging studies, or surgical findings. RESULTS: Twenty-three patients with lymphoma and 15 patients with benign disease or reactive lymphadenopathy were identified. The overall sensitivity, specificity, and accuracy of EUS-FNA cytology with flow cytometry/immunocytochemistry (FC/IC) for the diagnosis of lymphoma were, respectively, 74%, 93%, and 81%. When comparing patients who had EUS-FNA with FC/IC versus those who had EUS-FNA without FC/IC, sensitivity was 86% versus 44% (p = 0.04), specificity was 100% versus 90% (not significant), and accuracy was 89% versus 68% (not significant). CONCLUSION: EUS-FNA can provide cytology specimens diagnostic for lymphoma. Selective use of FC/IC in patients with suspected lymphoma improves the yield of EUS-FNA and may guide diagnostic evaluation and treatment decisions. PMID- 11275889 TI - EUS with EUS-guided fine-needle aspiration as the first endoscopic test for the evaluation of obstructive jaundice. AB - BACKGROUND: This study assesses the cost savings associated with using endoscopic ultrasound (EUS) before endoscopic retrograde cholangiopancreatography (ERCP) for evaluating patients with suspected obstructive jaundice. METHODS: One hundred forty-seven patients with obstructive jaundice of unknown or possibly neoplastic origin had EUS as their first endoscopic procedure. With knowledge of the final diagnosis and actual management for each patient, their probable evaluation and outcomes and their additional costs were reassessed assuming that ERCP would have been performed as the first endoscopic procedure. Also calculated were the additional costs incurred if EUS were unavailable for use after ERCP and had to be replaced by computed tomography or other procedures. RESULTS: The final diagnoses in these patients included malignancies (65%), choledocholithiasis or cholecystitis (18%), "medical jaundice" (11%), and miscellaneous benign conditions (6%). Fifty-four percent had EUS-guided fine-needle aspiration but only 53% required ERCP after EUS. An EUS-first approach saved an estimated $1007 to $1313/patient, but the cost was $2200 more if EUS was unavailable for use after ERCP. Significant savings persisted through sensitivity analysis. CONCLUSIONS: Performing EUS with EUS-guided fine-needle aspiration as the first endoscopic procedure in patients suspected to have obstructive jaundice can obviate the need for about 50% of ERCPs, helps direct subsequent therapeutic ERCP, and can substantially reduce costs in these patients. PMID- 11275891 TI - Endoscopic therapy for acute diverticular hemorrhage. PMID- 11275892 TI - Mucus outflow from the appendiceal orifice due to an appendiceal mucocele. PMID- 11275893 TI - Small bowel adenocarcinoma. PMID- 11275894 TI - Early-stage gastric adenocarcinoma: revealed after anti-Helicobacter pylori therapy of MALT lymphoma. PMID- 11275895 TI - Initial experience with a steerable, phased vector array ultrasound catheter in the GI tract. AB - BACKGROUND: EUS requires a significant capital outlay. The ability to perform high-resolution phased array scanning and Doppler interrogation by using a catheter that interfaces with a standard US console could increase the accessibility of EUS. Recently, an electronic phased-array US catheter was developed for intracardiac use. To date, this technology has not been applied to the GI tract. The aim of this study is to determine the feasibility and imaging characteristics of a new phased array scanning US catheter in the GI tract. METHODS: Swine were placed under general anesthesia. This study used a 100 cm, 10F, torquable catheter with 4-way tip deflection to greater than 90 degrees. The catheter tip houses a phased vector array transducer with variable frequency (5.5 10 MHz) and variable focal distance. It has pulsed/color and power Doppler capability. The probe was passed through a therapeutic flexible sigmoidoscope into the upper GI tract. Acoustic coupling was achieved via a condom filled with water or by gastric water infusion. Needle visualization experiments were performed with a second endoscope (also passed per oral) with a standard EUS guided fine needle aspiration needle. RESULTS: Acoustic coupling was easily achieved. Resolution of the GI wall into characteristic layers (esophagus 5, stomach 7) was demonstrated. At 5.5 MHz, tissue resolution and Doppler imaging were excellent to greater than 10 cm from the transducer. A 22-gauge EUS-guided fine needle aspiration needle was easily visualized at depth greater than 4 cm. Flow in gastric, hepatic, and pancreatic parenchymal vessels approximately 1 mm diameter was visualized by using power and color Doppler. CONCLUSIONS: This 10F array US catheter is capable of high-resolution two-dimensional imaging of the gut wall as well as high-quality Doppler imaging. The Doppler capabilities of this equipment may have new GI applications. PMID- 11275896 TI - Endoscopic application of photodynamic therapy for cholangiocarcinoma. AB - BACKGROUND: Previous studies indicate that photodynamic therapy provides effective relief from biliary obstruction in advanced cholangiocarcinoma. This report describes a method of applying photodynamic therapy in the biliary tract by using accessories available in the United States. METHODS: Endoscopic retrograde cholangiography was performed to define the proximal and distal extent of intraductal tumor. Patients were injected with 2 mg/kg of sodium porfimer. Forty-eight hours later a commercially available cylindrical diffusing laser fiber was inserted into an 8F biliary catheter equipped with a 0.038 inch side hole at its distal tip. After positioning of a 0.035 inch guidewire proximal to the biliary stricture, the preloaded catheter was advanced over the guidewire by using the monorail technique. Laser light was applied at a power of 400 mW/cm fiber for a total energy of 180 J/cm.(2) RESULTS: Fourteen treatments were performed on 6 patients with tumors of Bismuth types IV (n = 2), III (n = 3), or II (n = 1). By using the preloaded biliary catheter, adequate positioning of the laser fiber was achieved in all patients. A fracture of the diffuser tip occurred during 1 of the treatments. Two patients developed acute cholangitis and 2 patients experienced skin phototoxicity. CONCLUSIONS: Photodynamic therapy for cholangiocarcinoma is safe and technically feasible with a preloaded biliary catheter and a monorail technique for catheter positioning. PMID- 11275899 TI - Preoperative diagnosis of intraductal papillary mucinous tumors of the pancreas by endoscopic pancreatic biopsy. PMID- 11275900 TI - Retroperitoneoscopy in the management of drained infected pancreatic necrosis. PMID- 11275898 TI - Retortamonas intestinalis in the pancreatic juice of a patient with small nodular lesions of the main pancreatic duct. PMID- 11275897 TI - Combined use of electrosurgical incisions and balloon dilatation for the treatment of refractory postoperative pyloric stenosis. AB - BACKGROUND: Drug therapy plus balloon dilatation without gastroscopic incision does not always relieve postoperative pyloric stenosis. METHODS: Five patients with postoperative pyloric stenosis whose symptoms did not improve with drug therapy and balloon dilatation underwent a combination of gastroscopic incision and balloon dilatation. Two or 3 small radial incisions were made in the stenotic muscle of the pylorus electrosurgically at gastroscopy. Then the stenotic muscle layer was loosened and split bluntly along the incisions with balloon dilatation for 15 to 20 minutes. One week later, the combination procedure or balloon dilatation alone was repeated to prevent restenosis. RESULTS: In the 5 patients, the stenosis was improved with the combination therapy. No complications were observed. CONCLUSIONS: Combined use of gastroscopic incision and balloon dilatation may be considered for patients with refractory pyloric stenosis caused by surgical truncal vagotomy. PMID- 11275901 TI - Endoscopic cystogastrostomy during pregnancy. PMID- 11275902 TI - Sclerosing pancreato-cholangitis responsive to corticosteroid therapy: report of 2 case reports and review. PMID- 11275903 TI - Endoscopic incision of a postoperative colonic stricture. PMID- 11275904 TI - Expandable metal stent placement in the treatment of a malignant anastomotic stricture of the transverse colon. PMID- 11275905 TI - Hyponatremic encephalopathy after preparation for colonoscopy. PMID- 11275906 TI - Enterocutaneous fistula: a rare complication of PEG tube placement. PMID- 11275908 TI - Acquired immunodeficiency syndrome-related cryptosporidial cholangitis: resolution with endobiliary prosthesis insertion. PMID- 11275907 TI - Post-polypectomy hemorrhage in von Willebrand disease. PMID- 11275909 TI - Intrahepatic stones might mimic cholangiocarcinoma on ERCP. PMID- 11275910 TI - Endoscopic pancreatic stent insertion for treatment of pseudocyst after distal pancreatectomy. PMID- 11275911 TI - Foreign body removal using a "homemade" loop basket. PMID- 11275912 TI - Endoscopic treatment modalities for GERD: technologic score or scare? PMID- 11275914 TI - Evolving role of echocardiography in the management of atrial fibrillation. AB - The most common cardiovascular arrhythmia is atrial fibrillation (AF), with an estimated prevalence of 2.5 million in 1994. The extent of this public health problem is enormous, particularly in its stroke sequelae. Routine management of this public health problem includes anticoagulation as the primary stroke preventive measure. Echocardiography has been an important adjunctive tool in the evaluation of AF. More innovative and controversial is the putative role of transesophageal echocardiography in the treatment strategy of AF cardioversion to sinus rhythm. The standard of care for AF of less than 1-year duration is to attempt cardioversion to sinus rhythm. An alternative strategy is to utilize the assets of transesophageal echocardiography to visually screen the left atrium for thrombus, thereby playing an active role in the treatment strategy of AF. This review will discuss the role of echocardiography in AF as it was initially used as a diagnostic tool with weak prognostic features, and, more recently, as it can be used today as an adjunctive tool to guide therapy with excellent stroke risk stratification features. PMID- 11275913 TI - Electrocardiographic diagnosis of acute myocardial infarction: Current concepts for the clinician. AB - BACKGROUND: Over the past 2 decades, the 12-lead electrocardiogram has attained special significance for the diagnosis and triage of patients with chest pain because timely detection of myocardial injury and a rapid assessment of myocardium at risk proved pivotal to implementing effective reperfusion therapies during acute myocardial infarction. However, this wealth of information could still be underutilized by clinicians who may restrict their diagnostic quest in patients with chest pain to the more classic electrocardiographic signs. METHODS: The medical literature on electrocardiographic manifestations of acute myocardial infarction was extensively reviewed. RESULTS: The widespread utilization of both coronary angiography and methods to determine myocardial function and metabolism in patients with acute myocardial infarction over the last 10 years has provided the means for rigorous comparisons with electrocardiographic information. We summarize these electrocardiographic signs and patterns in terms of their relevance to the clinician to help reduce the incidence of "nondiagnostic electrocardiograms" and improve timely decision-making. CONCLUSIONS: The electrocardiogram continues to be an invaluable tool in the initial evaluation of patients with chest pain. The plethora of data currently available on electrocardiographic changes correlating with myocardial injury allows clinicians to make faster and better decisions than ever before. PMID- 11275915 TI - Evaluation Study of Congestive Heart Failure and Pulmonary Artery Catheterization Effectiveness (ESCAPE): design and rationale. AB - BACKGROUND: There is little information about how to adjust pharmacologic agents in the treatment of patients with advanced congestive heart failure (CHF). Some studies have suggested that use of pulmonary artery catheterization to guide reductions in filling pressures may improve outcomes for patients with heart failure who are hospitalized with evidence of elevated filling pressures. However, there is no consensus regarding the true utility of this strategy. A randomized clinical trial is needed to test the safety, efficacy, and treatment benefit of pulmonary artery catheterization in patients with advanced CHF. STUDY DESIGN: The Evaluation Study of Congestive Heart Failure and Pulmonary Artery Catheterization Effectiveness (ESCAPE) trial is a multicenter, randomized trial designed to test the long-term safety and efficacy of treatment guided by hemodynamic monitoring and clinical assessment versus that guided by clinical assessment alone in patients hospitalized with New York Heart Association class IV CHF. Five hundred patients will be randomly assigned to receive either medical therapy guided by a combination of clinical assessment and hemodynamic monitoring (PAC arm) or medical therapy guided by clinical assessment alone (CLIN arm). The primary end point of ESCAPE will be the number of days that patients are hospitalized or die during the 6-month period after randomization. Secondary end points will include changes in mitral regurgitation, peak oxygen consumption, and natriuretic peptide levels. Other secondary end points will be pulmonary artery catheter-associated complications, resource utilization, quality of life measures, and patient preferences regarding survival. IMPLICATIONS: The primary goal of ESCAPE will be to provide information about the utility of the pulmonary artery catheter in patients with advanced heart failure, independent of various treatment approaches used by individual physicians. In addition, this study will define current outcomes for this severely compromised population. PMID- 11275916 TI - Regulatory issues for Data and Safety Monitoring Committees. PMID- 11275917 TI - Terms of reference for Data and Safety Monitoring Committees. AB - The Data and Safety Monitoring Committee (DSMC) constitutes one of the most complex elements of a clinical trial. For this reason, it is critical that all participants in and contributors to a trial understand the functions and responsibilities of the DSMC. Failure to understand the complexity of this group's mission can lead to substantial opportunities for confusion among investigators, sponsors, regulators, and the public, especially if the trial that it monitors must be modified or terminated early. PMID- 11275918 TI - The Data and Safety Monitoring Committee: Some final thoughts. PMID- 11275919 TI - Testing the effectiveness of converting patients to long-acting antianginal medications: The Quality of Life in Angina Research Trial (QUART). AB - OBJECTIVE: Our purpose was to test the hypothesis that converting patients with stable angina to long-acting antianginal medications would improve their functional status, symptom control, treatment satisfaction, and quality of life. METHODS AND RESULTS: A single-blind randomized trial of 100 patients with stable coronary artery disease was performed in the outpatient clinic of a Veterans Affairs Health System. Outpatients with chronic stable angina taking at least 2 antianginal medications were studied. Patients were randomized to one of two treatments: optimal adjustment of their usual antianginal medications or conversion to solely long-acting medications (long-acting diltiazem +/- nitroglycerin patches +/- atenolol) with subsequent optimization. The primary outcome was the 3-month change in Seattle Angina Questionnaire scores. Although no differences in physical limitation scores were noted, patients randomized to receive long-acting medications had improved symptom control (3-month improvement in anginal stability [19.1 vs 5.6, P =.02] and anginal frequency [17.8 vs 5.5, P =.006]), more treatment satisfaction (3-month improvement of 8.2 vs 3.0, P =.057), and better quality of life (3-month improvement of 11.2 vs 5.6, P =.09) compared with patients whose pretrial medications were optimized. The improvement in symptom control was statistically significant. CONCLUSION: Converting patients with chronic, stable angina to long-acting antianginal medications resulted in substantial improvements in symptom control with a trend toward better treatment satisfaction and quality of life. PMID- 11275920 TI - Recurrent ischemia after thrombolysis for acute myocardial infarction. AB - BACKGROUND: Reliable predictors have yet to be found for recurrent ischemia after thrombolysis for acute myocardial infarction (AMI), nor do we know whether early angiography can herald recurrent ischemia. This study sought to investigate the relationship between recurrent ischemia and cardiac procedures after thrombolysis for AMI. METHODS: The Global Utilization of Streptokinase and Tissue Plasminogen Activator for Occluded Coronary Arteries (GUSTO-I) trial prospectively studied recurrent ischemia, which was defined as the presence of angina and changes in hemodynamics or the electrocardiogram. Cox regression analysis was used to identify predictors of recurrent ischemia. Other variables examined included time to coronary angiography and revascularization. RESULTS: Of 21,772 US GUSTO-I patients, 6313 (29%) had recurrent ischemia before discharge. Women (hazard ratio [HR] 1.25, 95% confidence interval [CI] 1.17-1.33) and patients with hypercholesterolemia (HR 1.14, 95% CI 1.07-1.22) or prior angina (HR 1.40, 95% CI 1.32-1.49) had a higher likelihood of recurrent ischemia. Current smoking and hours to thrombolysis were inversely related to recurrent ischemia (HR 0.86, 95% CI 0.81-0.92, HR 0.97, 95% CI 0.95- 0.99, respectively). Patients who underwent angiography before recurrent ischemia had a marginally increased risk of ischemia within 12 hours after angiography (HR 1.2, 95% CI 1.1-1.4); ultimately, they had a considerably lower risk 1 week after angiography than did patients without angiography (HR 0.57, 95% CI 0.45-0.72). CONCLUSIONS: Female sex, hypercholesterolemia, prior angina, and nonsmoking status weakly predict recurrent ischemia. Early coronary angiography reduces recurrent ischemia, probably because high-risk patients are identified and revascularized. PMID- 11275921 TI - Prior aspirin users with acute non-ST-elevation coronary syndromes are at increased risk of cardiac events and benefit from enoxaparin. AB - BACKGROUND: The aim of this article was to investigate whether prior aspirin use in patients with acute coronary syndromes affects clinical outcome. The Efficacy Safety Subcutaneous Enoxaparin in Non-Q-Wave Coronary Events Study (ESSENCE) and Thrombolysis in Myocardial Infarction (TIMI) 11B trials have shown superiority of enoxaparin over unfractionated heparin (UFH) in patients with unstable angina and non-ST-segment elevation myocardial infarction (UA/NSTEMI). However, the treatment effect of enoxaparin in the subset of patients reporting prior aspirin use has not been determined. METHODS: The rate of death, myocardial infarction, and urgent revascularization at days 8 and 43 after randomization was compared among patients who received aspirin within the week before randomization with those who did not receive aspirin in the TIMI 11B trial. A total of 3275 patients (84%) were prior aspirin users. RESULTS: The admission diagnosis was similar for prior and nonprior aspirin users. At both day 8 and day 43 the event rate was higher for prior aspirin users than for nonprior aspirin users (odds ratio 1.6 [1.24-2.08], P =.0004 at day 43), even after correction for baseline characteristics. Compared with those prior aspirin users taking UFH, enoxaparin treated prior aspirin users had a reduced rate of the composite end point of death, myocardial infarction, and urgent revascularization at day 8 (odds ratio 0.82 [0.67-1.00], P =.046) and day 43 (odds ratio 0.83 [0.70-0.98], P =.032). CONCLUSION: Patients with UA/NSTEMI and prior aspirin use had a 60% higher risk of death and cardiac ischemic events compared with nonprior aspirin users. On the basis of this subanalysis, enoxaparin is superior to UFH in all patients. In prior aspirin users the benefit is more clearly demonstrated. PMID- 11275922 TI - The absence of high-frequency QRS changes in the presence of standard electrocardiographic QRS changes of old myocardial infarction. AB - BACKGROUND: This study compares the high-frequency QRS components (HF-QRS) in patients with and without standard electrocardiogram (ECG) changes indicative of old myocardial infarction (MI). Previous studies have indicated that patients with an old MI differ in their HF-QRS compared with healthy subjects. The HF-QRS has been reported to be decreased during acute coronary occlusion and increased after reperfusion. However, there is controversy about the appearance of HF-QRS after the acute phase of MI. METHODS: A total of 154 patients were included, 57 with and 97 without QRS changes of old MI on the standard ECG. The patients with old MI were divided into subgroups on the basis of the MI location indicated by the standard ECG. Signal-averaged ECGs from the 12 standard leads were recorded. The root-mean-square values of the HF-QRS were determined within two frequency bands: 150 to 250 Hz and 80 to 300 Hz. RESULTS: There was a large interindividual variation in HF-QRS in patients without MI as well as in those with different MI locations. There were no significant differences between the groups in the summed HF-QRS of all 12 leads or in the pattern of lead distribution of the HF-QRS. Not even the patients with the greatest QRS changes of old MI could be differentiated from those without any changes of old MI on the standard ECG. The results were the same in both analyzed frequency bands. CONCLUSIONS: This study shows, contrary to previous studies, that analysis of HF-QRS cannot differentiate between patients with and without old MI. PMID- 11275923 TI - Joint effects of C-reactive protein and other risk factors on acute coronary events. AB - BACKGROUND: Elevated serum C-reactive protein (CRP) is a predictor of coronary heart disease in population samples. We studied the contribution of the simultaneous presence (joint effects) of elevated CRP and the classic as well as some new risk factors on acute coronary events. METHODS: With a nested case control design and logistic regression analyses, we measured baseline and pre event CRP levels in patients who had myocardial infarction or coronary death (cases) during an 8.5-year follow-up in the Helsinki Heart Study, a coronary primary prevention trial in dyslipidemic middle-aged men. The control patients were participants remaining free of coronary events. RESULTS: Baseline and pre event CRP levels were higher in cases than in control patients (4.4 vs 2.0 mg/L, P <.001 and 6.0 vs 3.6 mg/L, P <.001). The relative risk attributed to elevated CRP was 40% higher with chronic elevation (odds ratio [OR], 3.34) compared with high baseline (OR, 2.24) or pre-event (OR, 2.26) level only. Hypertension, low high-density lipoprotein cholesterol, and high leukocyte count increased the risk only marginally without simultaneous occurrence of high CRP, whereas the joint effects of CRP and these classic risk factors suggested additive effects on coronary risk. In contrast, high levels of immunoglobulin G-class antibodies to oxidized low-density lipoprotein and antiprothrombin antibodies as well as high total immunoglobulin G level increased the risk irrespective of CRP. CONCLUSIONS: Elevated CRP enhances the risks attributed to classic coronary risk factors. PMID- 11275924 TI - Corrected TIMI frame counts correlate with stenosis severity and infarct zone wall motion after thrombolytic therapy. AB - BACKGROUND: The majority of patients with patent infarct-related arteries after thrombolytic therapy have slower than normal flow, which relates to myocardial perfusion. METHODS: To evaluate the relationships between blood levels of creatine kinase (CK) and the corrected Thrombolysis in Myocardial Infarction (TIMI) frame count (CTFC), infarct artery stenosis, and left ventricular function, we studied 397 patients with a first myocardial infarction who underwent angiography at 3 weeks. TIMI flow grades, the CTFC, infarct artery stenosis, and infarct zone wall motion (by contrast ventriculography using the centerline method) were assessed, and CK levels (in units per liter) were measured hourly for the first 4 hours after streptokinase (1.5 x 10(6) U over 30 60 minutes) and then every 4 hours over the next 20 hours, all blinded to treatment and outcome. RESULTS: Infarct artery stenosis and the CTFC, assessed as continuous variables, correlated in patients with patent infarct arteries (r = 0.33, P <.001). Also, there was a significant correlation between the CTFC and the sum of hypokinetic chords in the infarct zone (r = 0.15, P =.01). Patients with total occlusion or markedly slowed infarct artery flow (CTFC >100) had a higher fraction of chords with wall motion >2 SDs below normal (0.65 [0.41, 0.80] vs 0.37 [0.0, 0.67]) compared with patients with normal flow (CTFC < or =27) (P <.001). The rates of increase of median CK levels with respect to TIMI flow grades were 342 U/L/h for TIMI 3 versus 212 U/L/h for TIMI 2 versus 140 U/L/h for TIMI 0-1 (P <.0001). CONCLUSIONS: Prolonged corrected TIMI frame counts correlate with stenosis severity in the infarct artery after infarction, infarct zone regional wall motion, and CK levels. PMID- 11275926 TI - Effect of smoking status and abciximab use on outcome after percutaneous coronary revascularization: Pooled analysis from EPIC, EPILOG, and EPISTENT. AB - BACKGROUND: Tobacco use has been studied in relation to the development of atherosclerotic heart disease and outcome after acute coronary syndromes, though outcome data after percutaneous coronary intervention (PCI) are limited. Tobacco use has been associated with increased fibrinogen levels, thrombin generation, and increased platelet aggregation, and these factors may cause ischemic events after PCI. METHODS: We assessed whether smokers undergoing PCI have more ischemic events and if glycoprotein IIb/IIIa receptor blockade is particularly beneficial in preventing ischemic events in this cohort. We examined clinical outcomes for smokers and nonsmokers by using pooled analysis from 3 large, placebo-controlled trials of the glycoprotein IIb/IIIa antagonist abciximab during PCI. RESULTS: Among 6519 patients, 34% were smokers. The primary end point of death, myocardial infarction, or urgent revascularization within 30 days was reduced by abciximab from 12.3% to 6.4% (relative risk reduction, 48%; P <.001) in smokers and from 11.3% to 5.9% (relative risk reduction, 48%; P <.001) in nonsmokers treated with abciximab. At 6 months, death, myocardial infarction, or urgent revascularization was reduced from 14.4% to 8.5% (relative risk reduction, 41%; P <.001) in smokers and from 14.6% to 8.8% (relative risk reduction, 40%; P <.001) in nonsmokers. Adjusting for baseline differences, smoking was an independent predictor of poor outcome at 30 days (odds ratio, 1.22; 95% confidence interval, 1.02, 1.47; P =.03). CONCLUSIONS: This pooled analysis demonstrates that after PCI, smokers had worse 30-day outcomes than did nonsmokers. However, both groups had a comparable degree of benefit with platelet inhibition through the use of abciximab. PMID- 11275925 TI - Abciximab and early adjunctive percutaneous coronary intervention are associated with improved ST-segment resolution after thrombolysis: Observations from the TIMI 14 Trial. AB - BACKGROUND: Percutaneous coronary intervention (PCI) improves clinical outcomes in selected patients with failed thrombolysis but has not been proven to benefit patients who achieve a patent infarct-related artery. Even after successful epicardial reperfusion, myocardial perfusion may be inadequate. We sought to evaluate whether a strategy that uses a reperfusion regimen containing abciximab and a reduced-dose thrombolytic agent (combination therapy), followed by early adjunctive PCI, would result in improved myocardial perfusion, as assessed by ST segment resolution. METHODS: ST resolution from 90 to 180 minutes after therapy was calculated for all 410 patients from the TIMI 14 trial who had evaluable electrocardiograms at both time points and who were treated with alteplase or reteplase. Patients were grouped according to whether they were treated with combination therapy or full-dose thrombolytic agent alone and whether they underwent PCI between the 90- and 180-minute electrocardiographic measurements. RESULTS: Among 105 patients who underwent adjunctive PCI between 90 and 180 minutes, mean ST resolution from 90 to 180 minutes was significantly greater in those who had received combination therapy versus those who had received full dose thrombolytic alone (54% vs 8%; P =.002). Among 241 patients with TIMI grade 3 flow in the infarct-related artery at 90 minutes, adjunctive PCI significantly improved mean ST resolution in patients who had been treated with combination therapy (57% [PCI] vs 24% [no PCI]; P =.006), but PCI did not have this effect in patients who had received thrombolytic therapy alone (1% [PCI] vs 10% [no PCI]; P =.70). In a multivariate model controlling for factors that would be expected to independently influence 90- to 180-minute ST resolution, abciximab treatment remained significantly associated with greater ST resolution (P =.008). CONCLUSIONS: A strategy that uses a combination reperfusion regimen that includes abciximab, followed by early adjunctive PCI, is associated with greater ST segment resolution, which may reflect enhanced tissue level and microvascular perfusion. Future studies should evaluate prospectively the clinical efficacy of this strategy. PMID- 11275927 TI - Influence of treatment delay on long-term left ventricular function in patients with acute myocardial infarction successfully treated with primary angioplasty. AB - BACKGROUND: Myocardial salvage has been shown to be dependent on the time elapsed from the onset of acute myocardial infarction (AMI) to reperfusion. The aim of this study was to evaluate the importance of time to reperfusion for left ventricular function recovery after primary angioplasty (percutaneous transluminal coronary angioplasty [PTCA]) for AMI. METHODS: Ninety-five patients undergoing long-term successful PTCA for AMI were studied. Echocardiography was performed before and 3, 7, 30, 90, and 180 days after PTCA. End-diastolic volume index (EDVI) and end-systolic volume index (ESVI), ejection fraction, and left ventricular wall motion score index (WMSI) were evaluated. RESULTS: Patients were divided into group A, 23 patients reperfused within 2 hours; group B, 32 patients reperfused between 2 and 4 hours; group C, 22 patients reperfused between 4 and 6 hours; and group D, 18 patients reperfused between 6 and 12 hours. Both EDVI and ESVI were reduced in groups A and B at 90 days. Groups C and D did not show any changes of EDVI and ESVI at any stage throughout the study. Ejection fraction improved only in groups A and B at 30, 90, and 180 days. At study entry, WMSI was similar in all groups. After 7 days, in group A and in group B, WMSI was improved, no changes were observed in group C, and a mild deterioration was observed in group D at 3 and 7 days. Subsequent evaluations showed progressive improvement of WMSI in all groups. CONCLUSIONS: Myocardial salvage is achieved only in patients revascularized within 4 hours from AMI onset. However, revascularization after 6 hours may be worthwhile by preventing ventricular remodeling. PMID- 11275928 TI - Treatment of focal in-stent restenosis with balloon angioplasty alone versus stenting: Short- and long-term results. AB - BACKGROUND: Although both percutaneous transluminal coronary angioplasty (PTCA) and additional stenting can be used for the treatment for focal in-stent restenosis (ISR), no large-scale comparative data on the clinical outcomes after these interventional procedures have been reported. METHODS: In the current study we compared the in-hospital and long-term clinical results of PTCA alone (n = 266 patients, n = 364 lesions) versus stenting (n = 135 patients, n = 161 lesions) for the treatment of focal ISR, defined as a lesion length less than or equal to 10 mm. RESULTS: There were significantly more diabetic patients in the PTCA group than in the stent group (36% vs 26%, P =.04), but other baseline characteristics were similar. Lesion length and preprocedure minimal lumen diameter (MLD) were also similar in the two groups, but the stent group had a larger reference vessel diameter (3.40 +/- 0.73 mm vs 2.99 +/- 0.68 mm, P <.001). Stenting achieved a larger postprocedure MLD than PTCA did (2.95 +/- 0.95 mm vs 2.23 +/- 0.60 mm, P <.001) and a smaller residual diameter stenosis (11% +/- 15% vs 23% +/- 16%, P =.04). Angiographic success was achieved in all cases. The rate of death/Q-wave infarction of urgent revascularization was higher with PTCA than with stent (5.6% vs 0.7%, P =.02). Postprocedure creatine kinase myocardial band enzyme elevation >5 times normal was more frequent with stent (18.5% vs 9.7%, P =.05). At 1 year the two interventional strategies had similar cumulative mortality (4.6% PTCA vs 5.1% stent, P not significant) and target lesion revascularization rate (24.6% PTCA vs 26.5% stent, P not significant). By multivariate analysis, the sole predictor of target lesion revascularization was diabetes (odds ratio 2.4, 95% confidence intervals 1.2-4.7, P =.01). CONCLUSION: Repeat stenting for the treatment of focal ISR had a higher postprocedure creatine kinase myocardial band elevation rate and similar long-term clinical results compared with PTCA alone. PMID- 11275930 TI - Safety, feasibility, and prognostic implications of pharmacologic stress echocardiography in 1482 patients evaluated in an ambulatory setting. AB - BACKGROUND: The outpatient prognostic assessment of coronary artery disease (CAD) by exercise electrocardiography has limitations, including the feasibility of the test and its low positive predictive value in several clinical conditions. In the current study we investigated the safety, feasibility, and prognostic value of pharmacologic stress echocardiography in a large cohort of ambulatory patients. METHODS: The study group was made of 1482 ambulatory patients (969 men, aged 60 +/- 10 years) who underwent stress echocardiography with either dipyridamole (n = 846) or dobutamine (n = 636) for evaluation of suspected or known stable CAD. The pretest likelihood of CAD was intermediate (<70%) in 709 patients and high (> or =70%) in 773 patients. RESULTS: There was no complication during the dipyridamole test, whereas 2 ischemia-dependent, sustained ventricular tachycardias occurred during the dobutamine test. Limiting side effects were observed in 2% of dipyridamole and in 3% of dobutamine stresses. The echocardiogram was positive in 459 patients. During a mean follow-up of 28 +/- 24 months, 58 patients died, 33 had a nonfatal myocardial infarction, and 158 underwent early (< or =3 months) and 64 late (>3 months) revascularization. Multivariate predictors of hard events (death, infarction) were positive echocardiographic results (hazard ratio [HR] 2.9) and resting wall motion score index (WMSI) (HR 2.3). In considering major events (death, infarction, late revascularization) as end points, positive echocardiographic result (HR 4.3), scar (HR 2.2), and resting WMSI (HR 1.7) were independent prognostic predictors. The 5-year survival rates for the ischemic and nonischemic groups were, respectively, 80% and 91% (HR 3.6, 95% confidence interval [CI] 3.8-8.4; P <.0001) considering hard cardiac events and 65% and 88% (HR 2.6, 95% CI 2.1-5.9; P <.0001) considering major events. Multivariate predictors of major events were positive echocardiographic results (HR 8.2) and male sex (HR 2.5) for the intermediate-risk group and positive echocardiographic results (HR 2.9), resting WMSI (HR 1.8), and prior Q-wave myocardial infarction (HR 1.8) for the high-risk group. CONCLUSIONS: Pharmacologic stress echocardiography is safe, highly feasible, and effective in prognostic assessment of ambulatory patients when both a general population and groups selected on the basis of pretest likelihood of CAD are analyzed. It represents a valid complementary tool to exercise electrocardiography for prognostic purposes in outpatients. PMID- 11275929 TI - Clinical significance of ST-segment elevation in lead V1 in patients with acute inferior wall Q-wave myocardial infarction. AB - BACKGROUND: This study was designed to determine the clinical significance of ST segment elevation in the precordial leads (leads V1 and V2) in acute Q-wave inferior wall myocardial infarction. METHODS AND RESULTS: One hundred fifty-eight consecutive patients with acute Q-wave inferior wall myocardial infarction were classified into 3 groups on the basis of the initial ST-change in V1 (group 1 = 29 patients with ST elevation, group 2 = 97 patients with ST depression, and group 3 = 32 patients with no ST-segment change). The right coronary artery was the infarct-related artery in all the patients in group 1. Although there was no significant difference between groups 1 and 2, the number of left ventricular asynergic segments was larger and the incidence of major in-hospital arrhythmias was higher in groups 1 and 2 compared with group 3. Patients in group 1 had a significantly higher incidence of proximal lesion (86%) and right ventricular infarction (69%) than the other 2 groups did. When ST elevation in leads V1 and V2 was considered, 14 of 15 patients (93%) with ST elevation only in V1 had right ventricular infarction, whereas 6 of 14 patients (43%) with ST elevation in both V1 and V2 had right ventricular infarction (P =.011). CONCLUSIONS: ST-segment elevation in V1 on admission in patients with acute Q-wave inferior wall myocardial infarction indicates a right coronary artery lesion associated with a larger infarct size and a higher incidence of major in-hospital arrhythmias. PMID- 11275931 TI - Evaluation and follow-up of patients with left ventricular apical to aortic conduits with 2D and 3D magnetic resonance imaging and Doppler echocardiography: A new look at an old operation. AB - BACKGROUND: Although the interposition of left ventricular apical to descending aorta conduits has diminished with the advent of the Ross-Konno operation, it remains a useful option. We reviewed our institutional experience imaging these conduits and tested the hypothesis that the gradient across the native left ventricular outflow tract (LVOT) by echocardiography correlated with the conduit gradient by cardiac catheterization. In a patient with an unobstructed conduit, no gradient should exist across the native LVOT. METHODS: This was a retrospective review of the echocardiography, cardiac catheterization, magnetic resonance imaging (MRI) data, and history of 9 patients with these conduits over an 8-year period. In 7 of 9 patients, 8 conduit obstruction events were assessed by Doppler interrogation of the native LVOT and by cardiac catheterization. Five patients underwent 6 MRI scans. RESULTS: In all cases of obstruction diagnosed by catheterization (56.3 +/- 21.9 mm Hg), Doppler echocardiography demonstrated gradients across the native LVOT (69.3 +/- 21.2 mm Hg, r = 0.67). Because 2D echocardiography could not visualize the entire conduit in any patient, 2- and 3 dimensional MRI was used successfully to evaluate anatomy and identify the site of obstruction. All patients manifested conduit obstruction. Four (44%) of 9 patients died, 3 underwent the Ross operation, 1 continues to live with his original conduit, and 1 was lost to follow-up. CONCLUSIONS: A gradient by Doppler interrogation of the native LVOT is an indirect means of assessing conduit obstruction. MRI is a useful tool to complement anatomic diagnosis by echocardiography. Conduit obstruction is common, and late mortality rates appear to be high. PMID- 11275932 TI - Diabetes and the associated incidence of subclinical atherosclerosis and coronary artery disease: Implications for management. AB - BACKGROUND: We sought to examine the prevalence, sensitivity, and specificity of coronary calcium (CC), a marker of atherosclerosis, in a population of symptomatic and asymptomatic diabetic persons. METHODS: We used electron beam tomography (EBT) to quantitate CC in 168 symptomatic (chest pain or anginal equivalent) persons with diabetes who underwent coronary angiography and then compared this with a cohort of 155 asymptomatic persons with diabetes. RESULTS: In the 168 symptomatic diabetic persons, 124 (74%) had obstructive coronary artery disease (CAD) by angiography. Receiver-operator characteristic curve analysis was used to maximize sensitivity and specificity for obstructive CAD (>50% stenosis), which established a CC score of 102 as optimal. With use of this cut point, EBT has a sensitivity of 77% and a specificity of 77% for detecting obstructive CAD. Of the 155 asymptomatic diabetic persons, 72% had CC and 48% had a CC score >102. The presumed prevalence of obstructive disease (on the basis of EBT scores and prevalence of CC) among asymptomatic diabetic persons is quite high (as high as symptomatic persons without diabetes). Analyzing the 323 diabetic patients demonstrated no significant age difference in CC scores between women and men. CONCLUSIONS: This study confirms that higher CC scores should be used in diabetic patients to improve the specificity of CC to determine obstructive disease. EBT can allow a noninvasive diagnosis of CAD before clinical presentation, allowing for more therapy for those in which CC is detected. These results suggest that asymptomatic diabetic persons have the same atherogenic burden of those patients with CAD without diabetes. The high prevalence of CC in asymptomatic persons with diabetes supports the need for aggressive management of diabetes and associated risk factors. PMID- 11275933 TI - Evaluation of beta-blocker therapy in patients with dilated cardiomyopathy- Clinical meaning of iodine 123-metaiodobenzylguanidine myocardial single-photon emission computed tomography. AB - BACKGROUND: Patients with heart failure show signs of cardiac sympathetic dysfunction such as elevation of blood norepinephrine (NE) level, as a result of reduction in the number of sympathetic nerves, decrease in myocardial NE content, accelerated NE turnover or spillover of NE, and NE reuptake disorder at sympathetic nerve endings. In dilated cardiomyopathy (DCM), iodine 123 metaiodobenzylguanidine (MIBG) used clinically as a tracer for imaging of the sympathetic function was found to be useful in evaluation of severity and prognosis. METHODS AND RESULTS: A total of 143 (123)I-MIBG myocardial single photon emission computed tomography (SPECT) images were taken at successive intervals on 58 patients with DCM (mean age 54 +/- 11 years), as well as before and after therapy to determine the severity of DCM and the therapeutic effect of beta-blocker. Patients were divided into group A (n = 20), in which left ventricular ejection fraction (LVEF) improved by 10% or more within 6 months after the administration of beta-blocker, and group B (n = 20), in which there was less than a 10% change in LVEF. After (123)I-MIBG myocardial SPECT was taken, the washout rate for the entire left ventricle was calculated from early and delayed images. The estimations of extent score and severity score were based on the polar map prepared from short axial images taken from 17 healthy volunteers (mean age 35 +/- 5 years). There was a significant correlation between LVEF and (123)I-MIBG findings (extent score, severity score, and washout rate) obtained before and after beta-blocker therapy. After beta-blocker therapy, LVEF and (123)I-MIBG findings significantly improved in group A. On the other hand, no change occurred in (123)I-MIBG findings in group B. There was no significant difference in LVEF between group A (32.1% +/- 8.0%) and group B (29.5% +/- 7.2%). Also, there was no significant difference in the washout rate between group A (54.4% +/- 10.9%) and group B (52.9% +/- 7.2%). Comparison of (123)I-MIBG images before beta-blocker therapy of group A and group B revealed that the extent score (26.5 +/- 16.0 vs 44.5 +/- 21.1, respectively; P <.01) and the severity score (24.9 +/- 21.9 vs 58.2 +/- 35.2, respectively; P <.01) on the basis of the early (123)I-MIBG image was significantly lower for group A. CONCLUSIONS: From the above findings, patients with DCM in which (123)I-MIBG uptake is high on early images were expected to show improvement in cardiac function by beta-blocker therapy. Findings also suggested that (123)I-MIBG was useful for examining the severity of DCM, determining the applicability of beta-blocker therapy, estimating the maintenance dosage of beta-blocker, and evaluating prognosis. PMID- 11275934 TI - A semiautomated objective technique for applying the proximal isovelocity surface area method to quantitate mitral regurgitation: Clinical studies with the digital flow map. AB - BACKGROUND: Clinical application of the color Doppler proximal isovelocity surface area (PISA) method to quantify mitral regurgitation (MR) has been limited by the often inaccurate assumption that isovelocity surfaces are hemispheric. This study applied an objective method for selecting the region where the hemispheric geometry holds best on the basis of mathematic analysis of results at different distances from the orifice. We aimed to demonstrate this approach can be applied accurately in the clinical setting and can be semiautomated to promote routine use by extracting velocities from the digital Doppler output and then performing all the calculations automatically. METHODS: In 75 patients with isolated MR, centerline velocities (V(r)) at each distance (r) from the orifice in the proximal flow field were extracted digitally. The automated analysis calculated peak MR flow rates as 2pir(2)V(r) and plotted these against their respective velocities. The optimal value for peak flow rate was obtained mathematically at the site where the slope of this curve was minimal (least inaccuracy). This value was combined with continuous wave Doppler data to provide regurgitant stroke volume (RSV) and orifice area (ROA), which were compared with quantitative Doppler in 75 patients and angiography in 42. RESULTS: RSV and ROA by this optimized, semiautomated PISA method correlated and agreed well with values from quantitative Doppler (y = 0.9x + 1.9, r = 0.90, standard error of the estimate [SEE] = 8.1 mL, mean difference = -0.7 +/- 8.5 mL for RSV; y = 0.9x + 0.02, r = 0.90, SEE = 0.048 cm(2), mean difference = -0.005 +/- 0.1 cm(2) for ROA) and correlated well with angiography (rho = 0.90 for both RSV and ROA). CONCLUSIONS: This objective PISA method for quantifying MR is accurate in the clinical setting and has been semiautomated by use of analysis of digital velocity data to provide a rapid and practical technique suitable to facilitate more extensive application in routine practice. PMID- 11275936 TI - Lack of association among five genetic polymorphisms of the renin-angiotensin system and cardiac hypertrophy in patients with aortic stenosis. AB - BACKGROUND: Patients with aortic stenosis (AS) have left ventricular hypertrophy (LVH). It is thought that LVH in these patients is a consequence of chronic left ventricular pressure overload. However, there is only a poor correlation between the degree of AS and the degree of LVH. Genetic polymorphisms of the renin angiotensin-aldosterone system (RAAS) have been considered to trigger the response of the left ventricle to chronic pressure overload and determine the degree of LVH in patients with AS. METHODS: One hundred five consecutive patients with symptomatic AS were examined by echocardiography and left heart catheterization to determine the severity of AS and LVH. Five genetic polymorphisms of the RAAS (ACE, AGTR1, AGT, CMA, CYP11B2) were analyzed in all patients and the results of genetic analysis were correlated to severity of AS and LVH to determine the importance of the polymorphisms for LVH. RESULTS: All tested genotypes were in Hardy-Weinberg equilibrium and allele frequencies were similar to other study populations. There was no correlation between the severity of AS and the severity of LVH. There was no association between the five tested genotypes of the RAAS and the severity of AS (mean gradient and area of the aortic valve) or LVH (LV muscle mass). CONCLUSION: We conclude that LVH in patients with AS is not determined by the tested genetic polymorphisms of the RAAS. PMID- 11275935 TI - Dilation of the aortic root in children infected with human immunodeficiency virus type 1: The Prospective P2C2 HIV Multicenter Study. AB - BACKGROUND: Vascular lesions have become more evident in human immunodeficiency virus type 1 (HIV)-infected patients as the result of earlier diagnosis, improved treatment, and longer survival. Aortic root dilation in HIV-infected children has not previously been described. This study was undertaken to determine the prevalence of aortic root dilation in HIV-infected children and to evaluate some of the potential pathogenic mechanisms. METHODS: Aortic root measurements were incorporated into the routine echocardiographic surveillance of 280 children of HIV-infected women: an older cohort of 86 HIV-infected children and a neonatal cohort of 50 HIV-infected and 144 HIV-uninfected children. RESULTS: By repeated measures analyses, mean aortic root measurements were significantly increased in HIV-infected children versus HIV-uninfected children (P values of < or =.04 and < or =.005 at 2 and 5 years of age, respectively, for aortic annulus diameter, sinuses of Valsalva, and sinotubular junction). Heart rate, systolic blood pressure, stroke volume, hemoglobin, and hematocrit were not significantly associated with aortic root size. Left ventricular dilation, increased serum HIV RNA levels, and lower CD4 cell count measurements were associated with aortic root dilation at baseline. CONCLUSIONS: Mild and nonprogressive aortic root dilation was seen in children with vertically transmitted HIV infection from 2 to 9 years of age. Aortic root size was not significantly associated with markers for stress-modulated growth; however, aortic root dilation was associated with left ventricular dilation, increased viral load, and lower CD4 cell count in HIV infected children. As prolonged survival of HIV-infected patients becomes more prevalent, some patients may require long-term follow-up of aortic root size. PMID- 11275937 TI - Early and late reactions after the use of iopamidol 340, ioxaglate 320, and iodixanol 320 in cardiac catheterization. AB - BACKGROUND: Although modern contrast agents have tolerability superior to older agents, significant differences remain between the agents currently in use. METHODS: To investigate the incidence of early (<24 hours) and late (>24 hours to 7 days) reactions to 3 contrast agents commonly used in cardiac catheterization, we performed a randomized, prospective, double-blind trial in which 2001 patients received one of the following agents: iopamidol 340, a nonionic monomer; ioxaglate 320, an ionic dimer; and iodixanol 320, a nonionic dimer. Possible reactions to contrast were recorded during the hospital admission and after discharge by means of a questionnaire, telephone follow-up, or both. RESULTS: Early reactions occurred in 22.2% of those receiving ioxaglate, 7.6% of those receiving iodixanol, and 8.8% of those receiving iopamidol (P <.0001). Late skin reactions occurred in 12.2% of those receiving iodixanol, 4.3% of those receiving ioxaglate, and 4.2% of those receiving iopamidol (P <.0001). CONCLUSIONS: The early side effect profile of certain ionic contrast agents suggests that these agents should no longer be used routinely in cardiac catheterization. The use of nonionic agents, however, is associated with late skin reactions, but there are notable differences between the monomeric and dimeric compounds. Although the skin reactions are generally benign, this is not always the case. Patients should be advised accordingly. PMID- 11275938 TI - Serum complement activation in congestive heart failure. AB - BACKGROUND: Although activation of the complement system in myocardial infarction and cardiopulmonary bypass has been shown to contribute to myocardial injury, its role in congestive heart failure (CHF) is unknown. The purpose of this study was to determine the presence of terminal complement activation and its relation to clinical outcomes in patients with CHF. METHODS: We measured serum levels of the terminal complement complex C5b-9 in 36 patients with symptomatic heart failure and left ventricular ejection fraction <40%. We compared the serum C5b-9 levels of these patients with CHF with a group of 12 age-matched control patients. Combined clinical outcomes (death, urgent heart transplantation, or hospitalization with worsening heart failure) at 6 months were determined. RESULTS: The serum C5b-9 [median (25th to 75th percentiles)] levels in 36 patients with CHF [101.5 ng/mL (40 to 164)] were significantly (P =.003) higher than in the 12 control patients [36.5 ng/mL (22 to 50)]. Significantly more of the patients with CHF with the highest levels of C5b-9 (highest 50th percentile) had New York Heart Association class IV symptoms (67% vs 33%; P =.04) and adverse clinical outcomes by 6 months (56% vs 17%; P =.02) compared with the patients with CHF with lower levels (lowest 50th percentile). CONCLUSIONS: We have described a significant elevation in circulating C5b-9, the terminal complement complex, in patients with symptomatic heart failure and have observed an association between high levels of C5b-9 and near-term adverse events. PMID- 11275939 TI - Immunosuppressive therapy in patients with thyroid eye disease: an overview of current concepts. AB - Thyroid eye disease (TED) is a debilitating disease impairing the quality of life of affected patients. Treatment is often not satisfactory. This review summarizes the existing literature and discusses the most widely used forms of treatment for TED such as glucocorticoids (GCs), and other immunosuppressive agents. GCs are the most commonly used treatment in patients with TED. Other immunosuppressive agents such as cyclosporin A, azathioprin, cyclophosphamide and ciamexone have been used, but the results are modest at best and indicate an unfavorable benefit risk relationship. Limited experience indicates that methotrexate may be effective even in patients with refractory TED. Somatostatin analogs, octreotide and lanreotide, may provide a valuable, although costly, therapeutic alternative to GCs. Orbital radiotherapy has been used in the management of TED for almost 60 years. However, its beneficial effects have been questioned recently by several studies, the details of which have not yet been published. Other studies have argued in favor of orbital radiotherapy; however, the benefits appear to be limited to improvement of extraocular muscle dysfunction. PMID- 11275940 TI - Age-related changes of the hypothalamic-pituitary-adrenal axis: pathophysiological correlates. AB - The aim of this review was to examine the evidence for age-related changes of the hypothalamic-pituitary-adrenal (HPA) axis in both physiological and pathological aging, on the basis of the many data in the literature, as well as of our personal findings. A statistically significant circadian rhythmicity of serum cortisol was maintained in elderly subjects, even if with a reduced amplitude of the 24 h fluctuations and a trend to an increase of the serum levels in the evening and at night-time, in comparison with young controls. Furthermore, an age related impairment of HPA sensitivity to steroid feedback was present in elderly people. The occurrence of senile dementia amplified the changes already present in physiological aging. While the cortisol secretion was generally well maintained in aging, the adrenal production of dehydroepiandrosterone and of its sulfate (DHEAS) exhibited an age-related decline. Therefore, the cortisol/DHEAS molar ratio was significantly higher in elderly subjects and even more in demented ones, than in young controls. Due to the opposite effects of cortisol and DHEAS on the brain and particularly on the hippocampal region, the imbalance between glucocorticoids and androgens occurring in physiological and even more in pathological aging, may have adverse effects on the function of this region, whose key role in learning and memory is well known. PMID- 11275941 TI - A programme of iodine supplementation using only iodised household salt is efficient--the case of Poland. AB - BACKGROUND: Iodine prophylaxis in Poland started in 1935 and has been interrupted twice: by World War II and in 1980 for economic reasons. Epidemiological surveys carried out after the Chernobyl accident in 1989 as well as in 1992/1993 and in 1994 as a 'ThyroMobil' study, revealed increased prevalence of goitre in children and adults. Ninety per cent of Poland was classified as an area of moderate iodine deficiency, and 10%, in the seaside area, as mild iodine deficiency territory. Iodine prophylaxis based on iodisation of household salt was introduced again in 1986 as a voluntary model and in 1997 as a mandatory model with 30+/-10 mg KI/kg salt. OBJECTIVE: The evaluation of the obligatory model of iodine prophylaxis in schoolchildren from the same schools in 1994 and 1999. METHODS: Thyroid volume was determined by ultrasonography. Ioduria in casual morning urine samples was measured using Sandell-Kolthoff's method, within the framework of the ThyroMobil study. RESULTS: Goitre prevalence decreased from 38.4 to 7% and urinary iodine concentration increased from 60.4 to 96.2 microg/l mean values between 1994 and 1999. In four schools the prevalence of goitre diminished below 5%. In 1999, 70% of children excreted over 60 microg I/l, and 36% over 100 microg I/l, whereas in 1994 the values were 44 and 13% respectively. CONCLUSION: The present findings indicate that iodine prophylaxis based only on iodised household salt is highly effective. PMID- 11275942 TI - Endocrine factors related to changes in total peripheral vascular resistance after treatment of thyrotoxic and hypothyroid patients. AB - OBJECTIVE: Total peripheral vascular resistance (TPR) decreases in thyrotoxicosis and increases in hypothyroidism. Several mechanisms may be involved, including adaptation to changes in heat production and direct non-genomic effects of tri iodothyronine (T3) on vascular smooth muscle cells. The aim of this study was to see if changes in TPR are related to changes in plasma concentrations of the endothelial hormones adrenomedullin and endothelin-1 as well as other hormones affecting vasculature. DESIGN: A prospective study. SUBJECTS: Eleven hypothyroid patients (pretreatment: thyroid-stimulating hormone (TSH) 68 (38-201) mU/l, T3 0.7 (0.35-1.5) nmol/l, fT4 3.0 (2.0-5.9) pmol/l, median (range)) and 14 with hyperthyroidism (pretreatment: TSH 0.02 (<0.01-0.06) mU/l, T3 6.4 (2.3-13.0) nmol/l, fT4 56.1 (22.9-70.0) pmol/l) were studied before treatment and 3 months after reaching the euthyroid state. Blood collection was carried out simultaneously with the recording of finger arterial pressure (FINAP). Cardiac output and TPR were derived from stroke volume computations by modelling flow from the FINAP signal. RESULTS: Thyroid-function tests of hypothyroid and thyrotoxic patients did not differ after restoration of the euthyroid state. TPR, expressed in arbitrary units (AU), decreased after correction of hypothyroidism (from 1.32+/-0.65 to 0.96+/-0.36 AU, P=0.04) and increased after correction of hyperthyroidism (from 0.75+/-0.18 to 1.10+/-0.35 AU, P=0.007). Adrenomedullin concentrations did not change during the transition from the hypothyroid state 3.2(0.9-11.0) pmol/l to the euthyroid state 4.9(0.9-8.6) pmol/l, but decreased after treatment of hyperthyroidism, from 5.2(0.9-11.0) pmol/l to 2.2(0.9-5.4) pmol/l. Plasma endothelin-1 was undetectable in all samples. Changes in TPR upon treatment correlated with log DeltafT4 (r=-0.65, P=0.001), log DeltaT3, (r=-0.57, P=0.006), Delta noradrenaline (r=0.54, P=0.02) and Delta ANP (atrial natriuretic peptide) (r=-0.59, P=0.004). Multiple linear regression analysis indicated that only T3 was an independent determinant of TPR. Changes in T3 accounted for 46% of the variability in the changes in TPR. CONCLUSIONS: TPR is reduced in thyrotoxicosis and increased in hypothyroidism. Restoration of the euthyroid state normalizes TPR. Changes in TPR are not related to plasma adrenomedullin concentrations, but 46% could be explained by changes in T3. Altered ANP secretion and adrenergic tone may contribute to the T3-induced changes in TPR. PMID- 11275943 TI - Identification of a novel mutation in the autoimmune regulator (AIRE-1) gene in a French family with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy. AB - Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is clinically characterized by the presence of two of the three major clinical symptoms: Addison's disease and/or hypoparathyroidism and/or chronic mucocutaneous candidiasis. Because of its autosomal recessive inheritance, this rare disorder constitutes an interesting model for understanding the molecular background of autoimmunity. Recently, mutations in the autoimmune regulator (AIRE 1) gene have been identified in APECED patients. Here we report, in a large French APECED family, the identification of a novel AIRE-1 missense mutation (Pro326Leu) in association with the Arg257Stop mutation which is detected in more than 80% of mutant Finnish AIRE-1 alleles. This Pro326Leu substitution occurs in the first plant homeodomain (PHD)-type zinc-finger domain of the protein which has been identified in a number of nuclear proteins involved in chromatin mediated transcriptional regulation, such as ATRX, TIF1, KRIP-1 and Mi-2 autoantigen. This mutation highlights the key role of this amino acid in the structure of the PHD domain and confirms that exon 8 constitutes a mutational hotspot. PMID- 11275945 TI - Decreased pituitary sensitivity to glucocorticoids in endurance-trained men. AB - OBJECTIVE: Muscular exercise induces hypothalamo-pituitary-adrenal (HPA) axis activation and when regularly repeated, as in endurance training, leads to HPA axis adaptation. To assess whether non-professional endurance-trained (ET) men with a substantial training load and no clinical or biological features of HPA axis overactivity can present subtle alterations of HPA axis sensitivity to glucocorticoid negative feedback, nine ET men were subjected to HPA axis testing using the dexamethasone-corticotrophin-releasing hormone (CRH) test. DESIGN: Nine endurance-trained men and eight healthy age-matched sedentary men were studied. Morning plasma cortisol and 24 h urinary free cortisol (UFC) were determined and a low dose dexamethasone suppression test (LDDST) was performed followed by CRH stimulation (dexamethasone-CRH test). RESULTS: After a day without physical exercise, at 0800 h, plasma ACTH and cortisol concentrations, and the 24 h UFC and UFC/urinary creatinine (UC) ratio were similar in ET and sedentary men. By contrast, clear differences between the groups were seen in cortisol and ACTH responses to the dexamethasone-CRH test. In eight ET subjects, after LDDST, basal ACTH and cortisol levels were similar to those of sedentary men, whereas one ET subject displayed a poor suppression of cortisol level (131 nmol/l). After injection of CRH, however, three of nine ET men's cortisol levels were not suppressed by dexamethasone but instead displayed significant CRH-induced increase (peak cortisol: 88, 125 and 362 nmol/l). No sedentary subject exhibited any increase in cortisol levels. CONCLUSION: Three of nine ET men with a mean maximum rate of O2 uptake (VO2, max) of 61 ml/kg per min, running 50-70 km per week, were resistant to glucocorticoid suppression during the combined dexamethasone-CRH test. PMID- 11275944 TI - The importance of the unsuppressed glands in the study of intact parathyroid hormone disappearance after parathyroid adenomectomy. AB - BACKGROUND: In the usual techniques for intraoperative intact parathyroid hormone (iPTH) monitoring for primary hyperparathyroidism, the normal glands are implicitly considered suppressed. On the contrary, we believe, as do other researchers, that they are not totally suppressed. METHODS: For this reason, we considered the introduction of an infusion from the unsuppressed normal glands (UNG), described by an influx constant (IC (pg/ml per min)), into the formulation of a two-compartment model. For the blood compartment, we have: C(t)=A.exp( at)+B.exp(-bt)+EV, where A+B+EV=iPTH concentration at zero time (clamping), EV (equilibrium value)=IC/k, 'a' and 'b' are reciprocals of the time constants of the two exponentials and k=rate constant of elimination from the blood. The experimental data were obtained using an IRMA standard method, collecting samples in 20 patients, during and following adenomectomy. RESULTS: In spite of the variability among the patients, all fits were very good, thus confirming the importance of the UNG contribution to the shaping of the disappearance curve. For this reason, the relationship between the constant infusion from the UNG and the basal iPTH level at the induction of anaesthesia (BV), was studied. CONCLUSIONS: The existence of a negative correlation, together with the determination of a regression curve (IC=6.5BV), not only confirmed our assumptions, but also revealed the theoretical possibility of a priori knowledge of the iPTH contribution from the UNG. Hence, there is a theoretical possibility of discriminating between this contribution and that of the remaining (if any) affected gland(s). PMID- 11275946 TI - A new generation IRMA for ACTH with improved specificity: validation in various physiological and pathological conditions. AB - OBJECTIVE: Measurement of plasma ACTH is a key step for the exploration of hypothalamic-pituitary-adrenal disorders. To further improve ACTH recognition a new generation of ACTH IRMA was developed using antibodies directed towards succinylated ACTH (sACTH IRMA). DESIGN: The usefulness of this assay was compared with that of another commercially available ACTH IRMA assay using intact ACTH (ELSA-ACTH) in various pathophysiological situations: patients with low ACTH plasma levels, high ACTH plasma levels with normal or tumoural pituitaries, or ectopic ACTH syndrome, and pregnant women with high proopiomelanocortin (POMC) plasma levels. METHODS: All plasma samples were assayed simultaneously with the two different IRMAs. Comparisons were assessed by plotting the results along the theoretical line of identical values, and by the graphical method of Bland and Altman. RESULTS: In the ELSA-ACTH IRMA, CLIP (or ACTH18-39) showed true cross reactivity, and alpha-melanocyte-stimulating hormone and purified POMC both interfered and induced falsely lower ACTH results; in the sACTH IRMA no peptide showed any cross-reactivity, and only extremely high values of CLIP (50 000 pg/ml) interfered and induced falsely lower ACTH results. In ACTH hypersecretory syndromes, of tumoural (Cushing's disease, ectopic ACTH secretion) or non tumoural (Addison's disease, congenital adrenal hyperplasia) origins a good agreement between the two assays was observed except for very high ACTH plasma values (above 1000 pg/ml) and in some tumours where the sACTH IRMA yielded lower results; in some cases, the presence of circulating CLIP, demonstrated by HPLC studies, may contribute to this discrepancy. It is also likely that the calibration of the ELSA-ACTH kit itself generates higher ACTH values. In normal pregnant women both IRMAs gave highly correlated values, yet lower results were obtained with the sACTH IRMA. CONCLUSION: These data show that the sACTH IRMA has improved qualities of specificity and usefulness for rapid assessment of ACTH plasma levels. PMID- 11275948 TI - Vitamin D and estrogen receptor gene polymorphisms in type 2 diabetes mellitus and in android type obesity. AB - OBJECTIVE: We studied the significance of BsmI restriction enzyme polymorphism of the vitamin D receptor (VDR) gene and the XbaI and PvuII polymorphisms of the estrogen receptor (ER) gene in patients with type 2 diabetes (n=49), android type obesity with normal carbohydrate metabolism (n=29) and healthy controls (n=138). METHODS: The distribution of genotypes in the study groups, as well as their relationship to fasting and 1 h postprandial serum C-peptide levels were analyzed. RESULTS: Postprandial serum C-peptide levels of BB genotypes were significantly higher in the diabetes and obese groups (6.18+/-5.09 ng/ml) compared with other genotypes (2.71+/-2.45 vs. 1.72+/-1.97 ng/ml, respectively, P=0.05). Among patients with type 2 diabetes and obese subjects, the XX allelic variant of the ER gene was more frequent (P=0.00015). Postprandial C-peptide levels of subjects exhibiting XX genotype were significantly lower compared with those with Xx genotype (1.67+/-2.16 vs. 3.8+/-3.72 ng/ml, P=0.021). The BBXx allelic combination of the VDR/ER receptor genes was less frequent in diabetic patients than in healthy subjects or in obese patients. The BBXx genotype was associated with significantly elevated postprandial C-peptide levels in all subjects compared with other combinations (9.65+/-3.14 vs. 1.35+/-2.82 ng/ml, P=0.003). No difference was found in the distribution of the PvuII polymorphism of the ER gene or in the association with the C-peptide levels among study groups. CONCLUSION: Polymorphisms of the VDR/ER receptor genes might play a role in the pathogenesis of type 2 diabetes by influencing the secretory capacity of beta-cells. PMID- 11275947 TI - Final height in isolated GH deficiency type 1A: effects of 5-year treatment with IGF-I. AB - Data concerning final height (FH) in isolated growth hormone deficiency type 1A (IGHD1A) are scanty and controversial. In this paper we report the FH outcome of two girls with IGHD1A who were treated either with GH only (first patient) or with GH during the first 8 years and successively with IGF-I (second patient). In the first patient, FH was only slightly subnormal and slightly taller with respect to target height (TH). Surprisingly, FH was severely subnormal and very far from TH in the patient who underwent IGF-I therapy for >5 years: an auxological outcome similar to the one recently reported in the only two cases in the literature of patients with IGHD1A who have been treated with IGF-I until near FH achievement. We conclude that IGHD1A could have a very heterogeneous phenotypic expression in terms of FH and that IGF-I therapy, even though initiated some years before puberty onset and prolonged for more than 5 years, may not be able to ensure the normalization of height prognosis and the achievement of an FH close to TH. PMID- 11275950 TI - Absence of D147E mutation of CYP11B2 gene in hypertensive patients with increased corticosterone and aldosterone production. AB - OBJECTIVE: 11beta-Hydroxylase and aldosterone synthase are two highly homologous genes involved in different forms of human hypertension and in different animal models of hypertension. It has been shown that the conservative substitution D147E in the human CYP11B2 gene results in an increased production of corticosterone and aldosterone in vitro. A gene conversion between the CYP11B1 and CYP11B2 genes could be responsible for such a substitution. METHODS: In this study we investigated the presence of the mutation D147E of CYP11B2 in a group of 128 patients with primary aldosteronism, 68 patients with essential hypertension and increased corticosterone production and in 48 normal volunteers. RESULTS AND CONCLUSIONS: We did not identify any patient carrying this mutation, indicating that if it exists it is very rare and so has no relevance in determining the increased steroid excretion seen in some subtypes of human hypertension. PMID- 11275949 TI - Relationship between free and total 1,25-dihydroxyvitamin D in conditions of modified binding. AB - OBJECTIVE: A novel assay was employed to study the free 1,25-dihydroxyvitamin D (1,25(OH)2D) concentrations in various populations with different levels of 1,25(OH)2D and vitamin D binding protein (DBP). DESIGN: In 12 healthy women before and after 3 months of oral estrogen/progestagen treatment, 10 pregnant women, and 16 patients with a nephrotic syndrome and normal renal function, the concentrations of total and free 1,25(OH)2D, DBP and albumin were assessed. METHODS: The total concentration of 1,25(OH)2D in serum was assessed using a radioreceptor assay. The free fraction of 1,25(OH)2D was measured using symmetric dialysis. DBP was assessed using single radial immunodiffusion. Serum albumin concentrations were measured on an automated analyzer. RESULTS: In healthy women, the concentrations of total 1,25(OH)2D, free 1,25(OH)2D and DBP were 132+/-19 pmol/l, 92+/-30 fmol/l and 5.59+/-0.43 micromol/l. After 3 months of estrogen/progestagen treatment, total 1,25(OH)2D and DBP levels rose significantly to 175+/-51 pmol/l and 8.32+/-1.59 micromol/l (P< or =0.05 and P< or =0.001); the free 1,25(OH)2D remained unchanged (105+/-39 fmol/l; not significant). Pregnant women had significantly higher levels of total 1,25(OH)2D and DBP (239+/-68 pmol/l and 11.32+/-1.77 micromol/l; both P2.5) in all groups suggests that all activin A in the circulation is bound to follistatin in males. PMID- 11275955 TI - The role of phosphoinositide 3-kinase in spreading osteoclasts induced by colony stimulating factor-1. AB - BACKGROUND: Colony-stimulating factor-1 (CSF-1), a growth and survival factor for osteoclasts, stimulates these cells to spread and migrate towards a gradient of CSF-1. This may support the translocation of osteoclasts to new sites on the bone surface to be resorbed. Phosphoinositide 3-kinase (PI 3-K) is a lipid kinase participating in various signal transduction pathways. OBJECTIVE: To investigate the role of PI 3-K in the CSF-1-induced spreading of osteoclasts. METHODS: In isolated rat osteoclasts treated with or without CSF-1, the distribution of PI 3 K and proteins phosphorylated on tyrosine were investigated using immunofluorescence. In murine osteoclast-like cells grown from bone marrow cells co-cultured with osteoblasts, the activation of the PI 3-K by CSF-1 was determined both in vivo and in vitro. In vivo, the enzyme product in the cell was determined after extraction and separation with thin layer chromatography; in vitro, PI 3-K activity was measured in the pellet immunoprecipitated from the cell lysate. RESULTS: Inhibition of PI 3-K blocked the CSF-1-induced spreading of osteoclasts. In spreading osteoclasts, a portion of PI 3-K was translocated to the periphery where proteins phosphorylated on tyrosine appeared simultaneously. In osteoclast-like cells, CSF-1 stimulated PI 3-K activity. This activity could be immunoprecipitated with antibody against phophotyrosine residues. CONCLUSION: PI 3-K participates in the CSF-1-induced spreading of osteoclasts. The activated PI 3-K may induce the reorganization of the cytoskeleton resulting in spreading and migration. PMID- 11275956 TI - Morphine-induced stimulation of pituitary-adrenocortical activity is mediated by activation of nitric oxide in the early stages of postnatal life in the rat. AB - OBJECTIVE: The first aim of the present study was to determine if morphine, a prototypic mu-opioid agonist drug, affects pituitary-adrenocortical activity in developing rat pups (first and second weeks of postnatal life). The second aim of this study was to explore, in vivo, if nitric oxide (NO) could be involved in the neurohormonal response to morphine in the early stages of postnatal life. METHODS: Plasma ACTH and corticosterone concentrations were determined by RIA in rat pups (n=5-14 rats/experimental group) after they had been killed by decapitation. In a first experiment, 1-day and 1- and 2-week-old rats were treated s.c. with morphine (20 mg/kg) or with vehicle (0.9% NaCl) and killed 5-90 min later. In a second experiment, 2-week-old pups were pretreated s.c. with naltrexone (NAL; 0.4 mg/kg or 10 mg/kg), and injected 1 h later with either morphine (20 mg/kg) or vehicle, and killed 30 min later. Some pups injected with only NAL were killed 60 or 90 min later. On the other hand, pups injected with NAL (10 mg/kg) or NAL and morphine were killed 30 min later. In a third experiment, 2-week-old pups were pretreated s.c. with N-omega-nitro l arginine methyl-ester (L-NAME; 30 mg/kg or 100 mg/kg), and injected 1 h later with either morphine (20 mg/kg) or vehicle, and killed 30 min later. Moreover, some pups injected with L-NAME (100 mg/kg) or L-NAME with morphine were killed 30 min later. In a final experiment, pups were injected s.c. with either S-nitroso-N acetylpenicillamine (SNAP; 5 mg/kg) or vehicle, and killed 60 or 90 min later. RESULTS: Morphine administered to rat pups elicited marked rises in both ACTH and corticosterone secretion. Moreover, these responses increased with advancing postnatal age. In 2-week-old rat pups, NAL, a competitive antagonist at mu-opioid receptors, administered alone increased both plasma ACTH and corticosterone concentrations 30 min later. L-NAME, a specific NO synthase inhibitor, did not affect plasma ACTH and corticosterone concentrations 30 min later when administered alone. NAL, when concomitantly administered with morphine, was unable to block morphine responses. In contrast, morphine responses were blocked by pretreatment (60 min before) with NAL or with L-NAME. Acute injection of SNAP increased both ACTH and corticosterone release. CONCLUSION: Our results suggest that opioids have controversial effects on pituitary-adrenocortical activity in the early postnatal period in the rat, and that endogenous NO is one of the major factors in the response of the pituitary-adrenocortical axis to morphine. PMID- 11275958 TI - Integration, phylogeny, and the hominid cranial base. AB - Basicranial features were examined in catarrhine primates and early hominids in order to demonstrate how information about morphological integration can be incorporated into phylogenetic analysis. Hypotheses purporting to explain the functional and structural relationships of basicranial characters were tested using factor analysis. Characters found to be functionally or structurally related to each other were then further examined in order to determine whether there was evidence that they were phylogenetically independent. If phylogenetic independence could not be demonstrated, then the characters were presumed to be integrated and were grouped into a complex. That complex was then treated as if it were a single character for the purposes of cladistic analysis. Factor analysis revealed that five basicranial features may be structurally related to relative brain size in hominoids. Depending on how one defines phylogenetic independence, as few as two, or as many as all of those characters might be morphologically integrated. A cladistic analysis of early hominids based on basicranial features revealed that the use of integrated complexes had a substantial effect on the phylogenetic position of Australopithecus africanus, a species whose relationships are poorly resolved. Moreover, the use of complexes also had an effect on reanalyses of certain published cladistic data sets, implying that those studies might have been biased by patterns of basicranial integration. These results demonstrate that patterns of morphological integration need to be considered carefully in all morphology-based cladistic analyses, regardless of taxon or anatomical focus. However, an important caveat is that the functional and structural hypotheses tested here predicted much higher degrees of integration than were observed. This result warns strongly that hypotheses of integration must be tested before they can be adequately employed in phylogenetic analysis. The uncritical acceptance of an untested hypothesis of integration is likely to be as disruptive to a cladistic analysis as when integration is ignored. PMID- 11275957 TI - Ghrelin and motilin: two sides of one coin? PMID- 11275960 TI - Egarapithecus narcisoi, a new genus of Pliopithecidae (primates, catarrhini) from the Late Miocene of Spain. AB - Pliopithecid remains from the Spanish locality of Torrent de Febulines (Late Vallesian, MN 10), consisting of right and left mandibular fragments with partial tooth rows and an isolated P(3) probably belonging to the same individual, are described and assigned to Egarapithecus narcisoi gen. et sp. nov. (Pliopithecidae, Crouzeliinae). This is a highly derived species dated at around 9 Ma (Ma = 10(6) years), representing the latest appearance of the family in the European continent. Morphologically it is the most distant member from the inferred primitive pliopithecid morphotype, displaying many autapomorphies that notably accentuate those of the remaining Crouzeliinae. A cladistic analysis based on lower cheek teeth, performed in order to tentatively assess the phylogenetic relationships of Egarapithecus within the Crouzeliinae, indicates that several equally parsimonious cladograms are possible in the light of current evidence. This is due to uncertainties regarding the position of Plesiopliopithecus and Crouzelia (here considered distinct genera), as a result of missing characters and the significant degree of homoplasy apparently involved in crouzeliine dental evolution. Whether Egarapithecus is more closely related to Crouzelia or to Anapithecus (the latter hypothesis tentatively favored here) cannot be definitively resolved with the currently available material and deserves further investigation. It is clear, however, that Egarapithecus is one of the more derived and specialized members of the Pliopithecidae. PMID- 11275961 TI - Pattern of dental development in Hominid XVIII from the Middle Pleistocene Atapuerca-Sima de los Huesos site (Spain). AB - . We describe the pattern of dental development of Hominid XVIII from the Middle Pleistocene Sima de los Huesos (SH) site of the Sierra de Atapuerca (Burgos, Spain). As expected, this pattern is similar to that of modern humans. A delay of development of the lower and upper canines was observed. In contrast, the relative advanced development of the lower second molars and, especially, the upper and lower third molars is noteworthy. This latter feature seems to be common in Pleistocene hominids, and suggests that the pattern of dental development evolved in the genus Homo during the Pleistocene. In European Middle Pleistocene hominids, this pattern probably was facilitated by the extra space available in the mandible and maxilla for developing teeth. PMID- 11275959 TI - Paleopathological and biomolecular study of tuberculosis in a medieval skeletal collection from England. AB - Nine human skeletons of medieval date from a rural English burial site show signs of skeletal tuberculosis. They were subject to polymerase chain reaction (PCR) assays aimed at detecting traces of DNA from infecting mycobacteria, with the purpose both of confirming the paleopathological diagnosis of tuberculosis and determining in individual cases whether disease was due to M. tuberculosis or M. bovis. In all nine cases, evidence for M. tuberculosis complex DNA was found, and in all instances it appeared that disease was due to M. tuberculosis rather than M. bovis. The significance of the findings for understanding tuberculous infection in rural agrarian communities in medieval England is discussed. PMID- 11275962 TI - Geographic variation in tool use on Neesia fruits in orangutans. AB - Geographic variation in the presence of skilled behavior may reflect geographic variation in genetic predispositions or ecological conditions (accompanied by reliable expression during development), or it may reflect the vagaries of invention and the appropriate social conditions for persistence. In this study, we compare the feeding techniques and tool-using skills used by orangutans to extract the nutritious seeds from Neesia fruits between Suaq Balimbing on Sumatra and Gunung Palung on Borneo, and map the distribution of Neesia tool use in Sumatran swamps. We show that neither genetics nor ecology is sufficient to explain the distribution of this tool use, confirming earlier findings on chimpanzees. We conclude that the ability to learn to use tools determines the geographic distribution. It is impossible to distinguish between the history of invention and the conditions for social transmission as the causal factors, but the high density and the social tolerance at Suaq Balimbing create propitious conditions for the maintenance of the skill as a tradition once it has been invented. High orangutan densities in the other Sumatran coastal swamps with Neesia tool use support the conclusion that suitable transmission conditions are the critical factor to explain the geographic distribution of skills such as feeding tool use. PMID- 11275963 TI - Evidence of probable scurvy in subadults from archeological sites in North America. AB - The authors surveyed subadult human skeletons from Native American archeological sites in the United States for evidence of skeletal lesions associated with scurvy. Geographic regions surveyed include the Midatlantic area, the Southeast (Florida), the Southwest, and the Plains. The prevalence of probable subadult scurvy ranged from zero in the Plains samples to 38% in a small sample from Florida. These data indicate the likelihood that scurvy was a significant childhood disease in many Native American groups. Reasons for variation in prevalence remain speculative but include regional and seasonal variation in food types and abundance, cultural patterns of storage and utilization, periodic food shortages, and the relative importance of corn in the diet. These factors are part of a nutritional complex that is related to disease prevalence which can be studied through evidence seen in archeological human remains. PMID- 11275964 TI - Brief communication: neurotraumatological aspects of head injuries resulting from sharp and blunt force in the early medieval period of southwestern Germany. AB - Approximately 10% (33 of 304) of the predominantly male skulls from the 6th through 8th centuries in southwestern Germany exhibit cranial fractures derived from blunt or sharp force trauma. No evidence of fracture healing characterizes 24% (n = 8) of these individuals. All nonhealed fractures were caused by sharp force, and four of these wounds cross the sagittal sinus. The lengths of these straight-edged wounds, produced exclusively by sword blows, measure around 8.0 cm for fatal, and about 5.0 cm for nonfatal wounds. Seventy-six percent (n = 25) of these skulls exhibit some healing, which indicates that these injuries did not lead to immediate death. In this group are all depressed fractures resulting from blunt force blows. Two thirds of the 45 cranial injuries noted on these 33 skulls are located on the left side of these individuals, with a concentration in the frontoparietal region. Bony indications of wound infection occur in four cases (12%). Three crania exhibit circular trepanations in association with fractures. These phenomena are discussed in the context of modern neurotraumatological knowledge. PMID- 11275965 TI - Montagnard ethnicity and genetic relations in Northern Cameroon: comment on "the peopling of sub-Saharan Africa: the case study of Cameroon," by G. Spedini et al. PMID- 11275967 TI - Homo erectus newyorkensis: An Indonesian fossil rediscovered in Manhattan sheds light on the middle phase of human evolution. PMID- 11275968 TI - New fossil hominid calvaria from Indonesia--Sambungmacan 3. AB - A morphologically distinct partial calvaria of Homo cf. erectus from Java, Indonesia is described. The fossil hominid Sambungmacan 3 (Sm 3) was first discovered in 1977 from the banks of the Solo River near the village of Poloyo, Sambungmacan district, in central Java. It was later recovered in a New York City natural history establishment in 1999 and quickly returned to the Indonesian authorities. Examination of Sm 3 shows that the calvaria is well preserved with only portions of the cranial base missing. The most striking characteristics of Sm 3 include: the presence of a vertically rising forehead, more open occipital/nuchal and frontal angles, a more globular vault, and a cranial capacity within the Homo erectus range. Most notably absent in Sm 3 are a number of the classic characters attributed to Homo erectus, such as a strongly expressed angular torus and a continuous supratoral sulcus. The absence of such characters would normally place the calvaria outside the range of Homo erectus (sensu stricto), however, overall quantitative and qualitative morphological assessments of Sm 3 place it within the Homo erectus spectrum. The combination of the morphological characters in Sm 3 may be interpreted in several ways: 1.) the known cranial variation of H. erectus from Indonesia and China is extended; 2.) this calvaria shows evidence of evolutionary change within H. erectus; or 3.) more than one species of Homo existed in the (presumed) Middle Pleistocene of Java.) PMID- 11275969 TI - Endocast of Sambungmacan 3 (Sm 3): a new Homo erectus from Indonesia. AB - A new fossil calvaria, Sambungmacan 3 (Sm 3), described in New Fossil Hominid Calvaria From Indonesia--Sambungmacan 3 by Marquez et al., this volume, yields one of the most advanced and complete endocasts yet recovered from Java. This communication provides a thorough interpretation of the external anatomical landmarks observable on Sm 3. Using computer tomography (CT) and traditional morphological measurements, our comparative paleoneurological analyses show that while Sm 3 has a mosaic of features that are similar to both Indonesian and Chinese H. erectus, it also possesses significant characters reminiscent of later hominins. These include a greater degree of asymmetry characterized by a possible left-occipital, right-frontal petalial pattern, left-right volumetric cerebral asymmetry, and marked asymmetry in Broca's cap. Moreover, the frontal lobe offers a more rounded, shortened appearance in contrast to the flat, elongated appearance of other Indonesian fossils (e.g., Sangiran 17). Another unique trait is exhibited in the transverse plane where the widest breadth of Sm 3 occurs more superiorly than in other Indonesian H. erectus. Thus, the endocast of Sm 3 presents a unique morphology not seen previously in the hominin fossil record. While the strong modern human characteristics in this endocast may not represent a particular ancestry, they do allow us to recognize a new dimension of the remarkable variation in Indonesian Homo erectus. PMID- 11275970 TI - The Sambungmacan 3 Homo erectus calvaria: a comparative morphometric and morphological analysis. AB - The Sambungmacan (Sm) 3 calvaria, discovered on Java in 1977, was illegally removed from Indonesia in 1998 and appeared in New York City in early 1999 at the Maxilla & Mandible, Ltd. natural history shop. Here we undertake an analysis of its phylogenetic and systematic position using geometric morphometrics and comparative morphology. The coordinates of points in the sagittal plane from glabella to opisthion were resampled to yield "lines" of 50 semi-landmarks. Coordinates of glabella, bregma, lambda, inion, and opisthion were also collected and analyzed separately. Casts of Homo erectus fossils from Indonesia, China, and Kenya and of "archaic H. sapiens" from Kabwe and Petralona, as well as 10 modern human crania, were used as the primary comparative sample. The modern humans were well separated from the fossils in a graphical superimposition of Procrustes aligned semi-landmarks as well as in principal component and canonical discriminant analyses. In all of these, Sm 3 falls intermediate between the fossil and modern groups. Morphological comparisons of Sm 3 with a selection of Homo erectus fossils revealed its greatest similarity to specimens from Ngandong and the Sm 1 calvaria. Compared to all other H. erectus, Sm 3 was distinctive in its more vertical supratoral plane, less anteriorly projecting glabella and less sharply angled occiput. In these features it was somewhat similar to modern humans. It is not yet possible to determine if this similarity implies an evolutionary relationship or (more likely) individual or local populational variation. Several features of Sm 3 (small size, gracile supraorbital torus and lack of angular torus, and position in principal component analysis) suggest that it was a female. The use of geometric morphometrics provides a means to statistically test the shapes of such fossils in a manner not easily duplicated by other methods. The intermediate position of Sm 3 between fossil and modern samples in several different subanalyses exemplifies the value of this approach. PMID- 11275971 TI - From Wolff's law to the Utah paradigm: insights about bone physiology and its clinical applications. AB - Efforts to understand our anatomy and physiology can involve four often overlapping phases. We study what occurs, then how, then ask why, and then seek clinical applications. In that regard, in 1960 views, bone's effector cells (osteoblasts and osteoclasts) worked chiefly to maintain homeostasis under the control of nonmechanical agents, and that physiology had little to do with anatomy, biomechanics, tissue-level things, muscle, and other clinical applications. But it seems later-discovered tissue-level mechanisms and functions (including biomechanical ones, plus muscle) are the true key players in bone physiology, and homeostasis ranks below the mechanical functions. Adding that information to earlier views led to the Utah paradigm of skeletal physiology that combines varied anatomical, clinical, pathological, and basic science evidence and ideas. While it explains in a general way how strong muscles make strong bones and chronically weak muscles make weak ones, and while many anatomists know about the physiology that fact depends on, poor interdisciplinary communication left people in many other specialties unaware of it and its applications. Those applications concern 1.) healing of fractures, osteotomies, and arthrodeses; 2.) criteria that distinguish mechanically competent from incompetent bones; 3.) design criteria that should let load-bearing implants endure; 4.) how to increase bone strength during growth, and how to maintain it afterwards on earth and in microgravity situations in space; 5.) how and why healthy women only lose bone next to marrow during menopause; 6.) why normal bone functions can cause osteopenias; 7.) why whole-bone strength and bone health are different matters; 8.) why falls can cause metaphyseal and diaphyseal fractures of the radius in children, but mainly metaphyseal fractures of that bone in aged adults; 9.) which methods could best evaluate whole-bone strength, "osteopenias" and "osteoporoses"; 10.) and why most "osteoporoses" should not have bone-genetic causes and some could have extraosseous genetic causes. Clinical specialties that currently require this information include orthopaedics, endocrinology, radiology, rheumatology, pediatrics, neurology, nutrition, dentistry, and physical, space and sports medicine. Basic science specialties include absorptiometry, anatomy, anthropology, biochemistry, biomechanics, biophysics, genetics, histology, pathology, pharmacology, and cell and molecular biology. This article reviews our present general understanding of this new bone physiology and some of its clinical applications and implications. It must leave to other times, places, and people the resolution of questions about that new physiology, and to understand the many devils that should lie in its details. (Thompson D'Arcy, 1917). PMID- 11275972 TI - Quantitation of the changes in vascularity during arthritis in the knee joint of a mouse with a digital image analysis system. AB - Many joint and bone diseases are caused by, or associated with vascular changes. Particularly in rheumatoid arthritis, vascular sprouting of synovial vessels plays a major role in the generation of joint pathology. To assess the effects of pharmaceuticals that are designed to inhibit neovascularization, we developed a quantitative procedure to measure vascular changes in cross-sections of the mouse knee joint during arthritic inflammation. Arthritis was induced in the knee joint of C57Black6 mice by a single subpatellar injection of methylated BSA after previous immunization. Total vascularity was visualized with a specific monoclonal rat anti-mouse antibody (9F1). Functional vessels were detected with the fluorescent perfusion marker Hoechst 33342. The localization of Hoechst and the vascular marker 9F1 were analyzed in separate images with an automated digital image processing system. By combining the two images, total vascularity and the perfusion status of the vessels during arthritis could be established. The digital image system measures synovial area (SA), number of all blood vessels (NBV) and the number of perfused blood vessels (NpBV). From these parameters the percentage of perfused vessels (perfusion fraction; PF), the vessel density (VD = NBV/SA) and the density of perfused vessels (VDp = NpBV/SA) can be calculated. The measurements showed that the area of synovial tissue had increased during arthritis. Moreover, both the number of blood vessels (NBV) and the number of perfused vessels (NpBV) in the synovial area had increased significantly on Days 4 and 7 after arthritis induction. This procedure enabled quantitation of total vascularity and of functional blood vessels in cross-sections of synovial tissue. It is expected to be a powerful tool, not only to analyze the effects of anti angiogenic therapies in animal models of arthritis, but could also be applicable to study vascular and perfusion changes in vascular related diseases of the skeleton. PMID- 11275973 TI - Anatomy and three-dimensional reconstructions of the brain of the white whale (Delphinapterus leucas) from magnetic resonance images. AB - Magnetic resonance imaging offers a means of observing the internal structure of the brain where traditional procedures of embedding, sectioning, staining, mounting, and microscopic examination of thousands of sections are not practical. Furthermore, internal structures can be analyzed in their precise quantitative spatial interrelationships, which is difficult to accomplish after the spatial distortions often accompanying histological processing. For these reasons, magnetic resonance imaging makes specimens that were traditionally difficult to analyze, more accessible. In the present study, images of the brain of a white whale (Beluga) Delphinapterus leucas were scanned in the coronal plane at 119 antero-posterior levels. From these scans, a computer-generated three-dimensional model was constructed using the programs VoxelView and VoxelMath (Vital Images, Inc.). This model, wherein details of internal and external morphology are represented in three-dimensional space, was then resectioned in orthogonal planes to produce corresponding series of "virtual" sections in the horizontal and sagittal planes. Sections in all three planes display the sizes and positions of such structures as the corpus callosum, internal capsule, cerebral peduncles, cerebral ventricles, certain thalamic nuclear groups, caudate nucleus, ventral striatum, pontine nuclei, cerebellar cortex and white matter, and all cerebral cortical sulci and gyri. PMID- 11275974 TI - Histochemical and biochemical analysis of phospholipase C isoforms in normal human gastric mucosa cells. AB - The expression and activity of PIP2-specific phospholipase C (PLC) in healthy human gastric mucosa cells were investigated by means of Western blotting, immunohistochemistry and in vitro activity assays. The results provide direct evidence for an almost exclusive expression of the PLC beta family and at the same time supply a cellular cartography of each represented isoform of this family. In this context, the putative roles of each isoform in the signaling events regulating the gastric mucosa metabolic machinery are discussed. These data provide a unique map of the specific expression and cellular distribution of the most represented PLC isoforms in healthy human gastric mucosa cells, which may constitute a reference point in future studies aimed at highlighting possible cytochemical and biochemical hallmarks of metaplastic or malignant transformation. PMID- 11275975 TI - Induction and clonal expansion of tumor-specific cytotoxic T lymphocytes from renal cell carcinoma patients after stimulation with autologous dendritic cells loaded with tumor cells. AB - Melanoma and renal cell carcinoma (RCC) are considered to be the most immunogenic tumors in humans. To generate conditions to induce primary T-cell responses against RCC and to allow further expansion of tumor-specific cytotoxic T lymphocytes (CTL) for adoptive transfer, peripheral blood mononuclear cells from RCC patients were stimulated with primary autologous tumor cells or monocyte derived dendritic cells (DC) loaded with either tumor lysate (TU-LY) or apoptotic tumor cells (TU-AP). Whereas repetitive stimulation (4x) with tumor cells alone induced a predominant population of CD3(-) natural killer cells, 4 weeks of stimulation with tumor-loaded DC favored induction and expansion of CD4+ T cells (>80%). However, 2 weekly stimulation cycles with tumor-loaded DC followed by restimulation with autologous irradiated tumor cells alone were optimal for induction of tumor-specific CTL responses in vitro. Using these culture conditions a marked increase of CD4+ T cells was observed during the first 2 weeks of stimulation with tumor-loaded DC. Subsequent restimulation with autologous tumor cells alone gave rise to 500-fold expansion of CD8+ T cells. These CD8+ T cells were shown to exhibit strong major histocompatibility complex class I-restricted cytotoxic activity against the autologous tumor. Comparison of TU-LY and TU-AP as a source of tumor antigen for loading DC did not show any difference in stimulating tumor-specific CTL. Length pattern analysis of the complementary determining region 3 (CDR3) of the T-cell receptor Vbeta chain revealed expansion of oligoclonal CTL populations with outgrowth of 1 or 2 clones after prolonged stimulation with autologous tumor cells. Our study demonstrated an efficient method for generating tumor-specific CTL in vitro that may be used to identify tumor cell antigens or that can be expanded for adoptive T-cell transfer in tumor immunotherapy. PMID- 11275976 TI - Mechanisms of platelet-derived growth factor-induced chemotaxis. PMID- 11275978 TI - Role of Fas and granule exocytosis pathways in tumor-infiltrating T lymphocyte induced apoptosis of autologous human lung-carcinoma cells. AB - We have isolated a cytotoxic T lymphocyte (CTL) clone, Heu161, that reacts specifically with the human autologous lung carcinoma cell line IGR-Heu. We first demonstrated that IGR-Heu lacked Fas-receptor expression and was resistant to CD95-induced apoptosis. To further elucidate the role of Fas in tumor immune surveillance, we have stably transfected IGR-Heu with a Fas-expression vector and isolated CD95-sensitive and -resistant clones. Our data indicated that the resistance of 2 selected Fas-transfected clones to CD95-mediated lysis correlated with down-regulation of caspase-8 or its lack of cleavage and subsequent activation. All Fas transfectants, either sensitive or resistant to anti-Fas agonistic antibody, were as efficiently lysed by the CTL clone as the parental cell line. In addition, neither anti-Fas-blocking antibody nor Fas-Fc molecule inhibited T-cell lysis of Fas-sensitive tumor clone. This cytotoxicity was extracellular Ca(2+)-dependent and abolished in the presence of EGTA, indicating that it was mainly granzyme-mediated. Interestingly, although the caspase inhibitor z-VAD-fmk had no effect on tumor-cell lysis, it efficiently blocked target DNA damage triggered by autologous CTLs via the granule exocytosis pathway, indicating that the latter event was caspase-dependent. The present results suggest that lung carcinoma-specific CTLs use mainly a granule exocytosis dependent pathway to lyse autologous target cells and that these effectors are able to circumvent alteration of the Fas-triggered intracellular signalling pathway via activation of a caspase-independent cytoplasmic death mechanism. PMID- 11275977 TI - The Rho/Rho-kinase and the phosphatidylinositol 3-kinase pathways are essential for spontaneous locomotion of Walker 256 carcinosarcoma cells. AB - Signal transduction pathways controlling spontaneous locomotion of Walker carcinosarcoma cells are not well understood. We have therefore investigated the role of signalling proteins in development of polarity and locomotion of these cells. Treatment of the cells with 100 ng/ml pertussis toxin had no significant effect on the percentage of polarized cells. In contrast, 2 different phosphatidylinositol 3-kinase inhibitors (wortmannin and LY-294002) markedly reduced the proportion of polarized cells. Spontaneous locomotion of the cells was also significantly inhibited by these two inhibitors. In agreement with these data, we observed localization of the p85alpha subunit of phosphatidylinositol 3 kinase predominantly in the membrane fraction of Walker carcinosarcoma cells, indicating constitutive activation of this enzyme. We also investigated a role of Rho family proteins. Spontaneous development of polarity was almost completely suppressed by electroporation of the cells in the presence of 4 microg/ml C(3) exoenzyme, which specifically ADP-ribosylates and inactivates Rho. Two downstream targets of Rho, the Rho-activated kinases I and II, were also detected predominantly in the particulate membrane fraction, suggesting constitutive activation. A specific Rho-kinase inhibitor (Y-27632) blocked spontaneous polarization and migration in a concentration-dependent manner. Our results indicate that constitutive activation of the Rho/Rho-kinase pathway and of phosphatidylinositol 3-kinase plays an essential part in spontaneous development of polarity and cell locomotion of these cells. PMID- 11275979 TI - Correlation between thymidine phosphorylase expression and invasion phenotype in cervical carcinoma cells. AB - The correlation between thymidine phosphorylase (dThdPase) expression and invasion phenotype in human uterine cervical carcinoma cells was investigated using 10 cervical carcinoma cell lines. Semi-quantitative reverse transcription polymerase chain reaction analysis was performed to investigate the mRNA levels of dThdPase and matrix metalloproteinase (MMP)-2 with beta-actin coamplified as an internal standard. dThdPase protein expression levels were detected by highly sensitive enzyme-linked immunosorbent assay. Tumor cell migration along a gradient of substratum-bound fibronectin and invasion into reconstituted basement membrane were evaluated by haptotactic migration and invasion assay. Although dThdPase mRNA and protein expression levels differed remarkably among the cell lines, there was a statistical correlation between them (r = 0.743, p = 0.0139). dThdPase gene and protein expression levels were well correlated with the number of cells that migrated and invaded (p < 0.05). Moreover, there was a close correlation between MMP-2 gene and dThdPase gene and protein expression levels (p < 0.05). Tumor cells that produce dThdPase may have a higher invasive and metastatic potential because of their capacity to pass through tissue barriers. PMID- 11275980 TI - Localization of IQGAP1 is inversely correlated with intercellular adhesion mediated by e-cadherin in gastric cancers. AB - Down-regulation of E-cadherin function is characteristic of cancer cells and might involve the small G-protein Rho family, including Rac1 and Cdc42. IQGAP1 has been reported to be one of the target proteins of Rac1 and Cdc42. To elucidate the role of IQGAP1 in cancer-cell adhesion, its expression was investigated in 47 cases of human gastric cancer by immunohistochemistry and Western blot upon protein fractionation, especially in comparison with E-cadherin and catenin expression. In the non-cancerous columnar epithelium of the stomach, IQGAP1, as well as E-cadherin/catenin, was expressed at the cell-cell boundary. IQGAP1 was frequently observed diffusely in the cytoplasm in intestinal-type tumors (20/22 cases) but was expressed at the cell membrane in diffuse-type tumors (19/25 cases), thus showing significant association with tumor differentiation (p < 0.01). Interestingly, membranous expression of IQGAP1 was inversely correlated with that of E-cadherin (p < 0.05) or alpha-catenin (p < 0.001). These observations were consistent with the Western blot results following protein fractionation. IQGAP1 was dominantly expressed in the soluble fraction in differentiated tumors; however, in undifferentiated tumors, it was mostly in the insoluble fraction. In contrast, both E-cadherin and alpha-catenin were detected only in the insoluble fraction. Thus, subcellular localization of IQGAP1 from the cytoplasm to the cell membrane was correlated with E-cadherin dysfunction and tumor dedifferentiation in gastric carcinogenesis. PMID- 11275982 TI - Establishment of rat hepatocellular carcinoma cell lines with differing metastatic potential in nude mice. AB - For better understanding of cancer metastasis, we have established an in vivo model for induction of highly metastatic hepatocellular carcinomas (HCC) in male F344 rats. From 1 tumor, 4 cell lines with differing metastatic potential (C1, C2, C6, C5F) were established by subcloning using the limited-dilution cloning technique. Two other lines, N1 and L2, arose from another primary HCC and a lung metastatic lesion, respectively. Although cell adhesion of each cell line in culture medium was different, tumors developing in the subcutis of nude mice after transplantation were all moderately differentiated HCC with a trabecular pattern. On subcutaneous injection into nude mice, all 6 cell lines proved to be tumorigenic in the injection site and C5F was highly metastatic to the lung. With injection into the tail vein, N1 and L2 formed frequent metastases in the lung as well as in lymph nodes. Using intraperitoneal injection, C1, C6, N1 and L2 showed marked disseminated growth in the abdominal cavity with bloody ascitis. Northern blot analysis revealed expression of known metastasis-related genes, KAI1 and heparanase, to be decreased in C5F, but no differences in expression of nm23-H1 were evident. A point mutation in the GSK-3beta phosphorylation site of the beta catenin gene was found in L2. These transplantable HCC cell lines that have different metastatic ability should be useful for elucidation of mechanisms of metastasis. PMID- 11275981 TI - Characterization of (123)I-vascular endothelial growth factor-binding sites expressed on human tumour cells: possible implication for tumour scintigraphy. AB - To explore the possibility of vascular endothelial growth factor (VEGF) receptor scintigraphy of primary tumours and their metastases, we analysed the binding properties of (123)I-labelled VEGF(165) ((123)I-VEGF(165)) and (123)I-VEGF(121) to human umbilical vein endothelial cells (HUVECs), several human tumour cell lines (HMC-1, A431, KU812, U937, HEP-1, HEP-G2, HEP-3B and Raji), a variety of primary human tumours (n = 40) and some adjacent non-neoplastic tissues as well as normal human peripheral blood cells in vitro. Two classes of high-affinity (123)I-VEGF(165)-binding site were found on the cell surface of HUVECs. In contrast, one class of high-affinity binding sites for (123)I-VEGF(165) was found on HMC-1, A431, HEP-1, HEP-G2, HEP-3B and U937 cells as well as many primary tumours. For (123)I-VEGF(121), a single class of high-affinity binding site was found on certain cell lines (HUVEC, HEP-1 and HMC-1) and distinct primary tumours (primary melanomas, ductal breast cancers and ovarian carcinomas as well as meningiomas). Tumour cells expressed significantly higher numbers of VEGF receptors compared with normal peripheral blood cells and adjacent non-neoplastic tissues. Immunohistochemical staining revealed that the VEGF receptor Flk-1 is expressed to a much higher extent within malignant tissues compared with neighbouring non-neoplastic cells. We observed significantly greater specific binding of (123)I-VEGF(165) and (123)I-VEGF(121) to a variety of human tumour cells/tissues compared with the corresponding normal tissues or normal peripheral blood cells. In comparison with (123)I-VEGF(121), (123)I-VEGF(165) bound to a higher number of different tumour cell types with a higher capacity. Thus, (123)I VEGF(165) may be a potentially useful tracer for in vivo imaging of solid tumours. PMID- 11275983 TI - Molecular genetic alterations on chromosomes 11 and 22 in ependymomas. AB - Ependymomas arise from the ependymal cells at different locations throughout the brain and spinal cord. These tumors have a broad age distribution with a range from less than 1 year to more than 80 years. In some intramedullary spinal ependymomas, mutations in the neurofibromatosis 2 (NF2) gene and loss of heterozygosity (LOH) on chromosome arm 22q have been described. Cytogenetic studies have also identified alterations involving chromosome arm 11q, including rearrangements at 11q13, in ependymomas. We analyzed 21 intramedullary spinal, 14 ventricular, 11 filum terminale and 6 intracerebral ependymomas for mutations in the MEN1 gene, which is located at 11q13, and mutations in the NF2 gene, which is located at 22q12, as well as for LOH on 11q and 22q. NF2 mutations were found in 6 tumors, all of which were intramedullary spinal and all of which displayed LOH 22q. Allelic loss on 22q was found in 20 cases and was significantly more frequent in intramedullary spinal ependymomas than in tumors in other locations. LOH 11q was found in 7 patients and exhibited a highly significant inverse association with LOH 22q (p<0.001). A hemizygous MEN1 mutation was identified in 3 tumors, all of which were recurrences from the same patient. Interestingly, the initial tumor corresponded to WHO grade II and displayed LOH 11q but not yet a MEN1 mutation. In 2 subsequent recurrences, the tumor had progressed to anaplastic ependymoma (WHO grade III) and exhibited a nonsense mutation in exon 10 of MEN1 (W471X) in conjunction with LOH 11q. This suggests that loss of wild type MEN1 may be involved in the malignant progression of a subset of ependymomas. To conclude, our findings provide evidence for different genetic pathways involved in ependymoma formation and progression, which may allow to define genetically and clinically distinct tumor entities. PMID- 11275984 TI - Chromosomal aberrations in sporadic pituitary tumors. AB - Pituitary adenomas are common intracranial neoplasms that may be hormone secreting or nonfunctional. Genetic defects associated with some pituitary tumors have been identified, although our understanding of the underlying molecular mechanisms remains incomplete. We have studied 75 sporadic pituitary tumors, representing the major clinical subtypes, by comparative genomic hybridization (CGH) with the aim of assessing for DNA copy number changes. CGH revealed chromosomal imbalances in 34 adenomas (45.3%), whereby gains were 4.9 times more frequently observed than losses. Most of the genetic alterations detected by CGH affected entire chromosomes (108/131, 82.4%). Gain of genetic material was observed predominantly on chromosomes X (24/75, 32%), 19 (12/75, 16%), 12 (6/75, 6.7%), 7 and 9 (5/75, 6.7%), whereas loss of DNA sequences most frequently affected chromosomes 11 (4/75, 5.3%), 13 and 10 (3/75, 4%). There were no significant differences in the CGH results for the individual clinical subtypes of pituitary tumors. These results reveal a nonrandom pattern of chromosomal alterations in pituitary tumors, in particular gains of entire chromosomes, and this may contribute to the development of such neoplasms. PMID- 11275985 TI - The LMP1 gene isolated from Russian nasopharyngeal carcinoma has no 30-bp deletion. AB - The Epstein-Barr virus (EBV) is tightly linked to the induction of undifferentiated nasopharyngeal carcinoma (NPC), a tumour endemic in certain areas of southeast Asia. The LMP1 gene encoded by EBV is a classical oncogene due to its ability to transform rodent fibroblasts. LMP1 is absolutely essential for transformation of B cells by the virus and is one of the few EBV genes found to be expressed in NPC. It was originally shown that the LMP1 gene from NPC harbours a deletion of 30 bp in the 3' part of the gene. However, this deletion is also present in the virus spread in healthy people of the areas endemic for NPC and also in other EBV-positive tumours as well as in healthy carriers. We isolated and sequenced the LMP1 gene obtained from tissue of 7 Russian patients with NPC and 1 German patient with an NPC-like tumour of the parotid gland (PG) and compared them with the LMP1 gene isolated from peripheral blood lymphocytes (PBLs) of 6 Russian and 4 German healthy EBV-positive carriers. Neither the Russian NPC cases nor the German NPC-like tumour harboured an LMP1 gene with the 30-bp deletion, while 1 Russian and 2 German carriers contained the LMP1 gene with the 30-bp deletion. In addition, the LMP1 gene isolated from PBLs of the German patient was virtually identical to the gene isolated from the primary tumour. Functional analysis showed no correlation between the presence or absence of the 30-bp deletion and the level of induction of the transcription factors NFkappaB and jun/AP-1 caused by LMP1. These data indicate that the 30-bp deletion is not a factor predisposing for NPC. Comparison of the DNA sequences revealed that the LMP1 genes present in the NPCs most likely represent the "strain" persisting in the general population. PMID- 11275986 TI - Functional inactivation of p73, a homolog of p53 tumor suppressor protein, by human papillomavirus E6 proteins. AB - Human papillomavirus (HPV) is strongly implicated as a causative agent in the etiology of cervical cancer. Of its gene products, E6 binds to and inactivates p53 tumor suppressor protein by ubiquitin/proteasome-dependent degradation. Recently, p73, a novel family of p53, has been identified and demonstrated, like p53, to activate p21(WAF1). Here we show that p73 is also inactivated by HPV-E6, but ubiquitin-mediated proteolysis is not responsive. Yeast two-hybrid and GST pull-down assays indicate a physical interaction between p73 and either HPV-16 or HPV-11 E6 proteins in vivo and in vitro, respectively. The transactivation domain (amino acid residues 1 to 49) is found to be absolutely required for the interaction. Transient co-expression of E6 significantly inhibits the p73-mdiated activation of p21(WAF1) promoter in a p53-defective C33A cell line. Using Gal4 p73 fusion protein, we demonstrate that E6 inhibition of p73 transactivation function is independent of sequence-specific DNA binding, which is confirmed by a direct electrophoretic mobility shift assay. Moreover, E6 inhibits p73 function by interfering with the activity of the amino-terminal activation domain. Co transfection of E6 mutants reveals that the same portion of E6 appears to be responsible for the inactivation of p53 and p73 function. However, the inactivation mechanism of p73 is clearly different from that of p53, because p73, unlike p53, is inactivated by both high- and low-risk E6s and is not susceptible to E6-dependent proteolysis. These overall results, consequently, suggest that in addition to the inactivation of p53, the functional interference of p73 by HPV-E6 may, at least in part, contribute to E6-mediated transformation and hyperproliferation of cervical cells. PMID- 11275987 TI - Opposite regulation of the HPV 20-URR and HPV 27-URR promoters by ultraviolet irradiation and cytokines. AB - Epidemiological evidence implicates ultraviolet radiation and genetic changes (e.g., p53 mutations) as important factors in the etiology of nonmelanoma skin cancer. Little is known about a possible role of cutaneous papillomaviruses in these tumors. We previously reported both positive and negative regulation of the promoter activity of a number of HPV types by UV irradiation. To determine the underlying mechanism, we examined the influence of pro-inflammatory cytokines and MAP-kinases induced by UV irradiation by transfecting the HPV 20-URR and the HPV 27-URR into the RKO, HaCaT and H1299 cell lines expressing wild-type or mutated p53 or lacking p53, respectively. IL-1alpha, IL-1beta, IL-6, IL-17, TNF-alpha, as well as interferon-alpha, -beta and -gamma activated the promoter in the HPV 20 URR but inhibited the HPV 27-URR promoter. The effect of IL-1alpha and UV light was abolished by the addition of IL-1 receptor antagonist. UV irradiation induced a prolonged activation of JNK in HaCaT and H1299 but not in RKO cells, and its dephosphorylation was enhanced in the presence of p53 and the HPV-URRs. PMID- 11275988 TI - Intra-tumoral administration of naked plasmid DNA encoding mouse endostatin inhibits renal carcinoma growth. AB - Endostatin is a C-terminal fragment of collagen XVIII and has potent anti angiogenic and anti-tumor activity. Mouse endostatin-coding sequences were obtained using PCR and linked to the signal sequence of influenzavirus hemagglutinin. The signal-sequence endostatin fragment was subcloned into plasmid vectors under the transcriptional control of cytomegalovirus promoter. Murine renal carcinoma (Renca) cells transfected with endostatin-coding plasmid are shown to secrete full-length endostatin. Endostatin-secreting Renca cells demonstrate slower growth in vivo compared to empty vector-transfected cells, but their in vitro growth is unaffected. Anti-angiogenic activity of secreted endostatin was confirmed in a Matrigel angiogenesis assay in vivo. We report growth inhibition of Renca tumors resulting from intra-tumoral delivery of plasmid vector encoding secretable endostatin. Elevated local concentrations of endostatin resulted from multiple intra-tumoral injections of endotoxin-purified plasmid DNA. Local endostatin levels were high enough to obtain growth arrest of Renca tumors. PMID- 11275989 TI - Effects of docetaxel in combination with radiation on human head and neck cancer cells (ZMK-1) and cervical squamous cell carcinoma cells (CaSki ). AB - The purpose of this study was to determine, as we did for paclitaxel, the cytotoxic and radiosensitizing potential of docetaxel in human head and neck cancer cells (ZMK-1), and in cervical squamous cell carcinoma cells (CaSki). ZMK 1 cells were incubated with docetaxel for 3, 9 or 24 hr before irradiation and 24 hr after irradiation. CaSki cells were incubated with docetaxel 24 hr before and after irradiation. For ZMK-1 cells, the docetaxel concentrations (0.7, 0.7 and 0.35 nM) were determined to obtain approximately equivalent cell survival at the different incubation times (3, 9 and 24 hr, respectively). For CaSki cells, the necessary concentration of docetaxel was 0.07 nM. Radiation doses were given from 0 to 7 Gy. Cell survival was measured by a standard clonogenic assay after a 9 day incubation. Flow cytometry was used to measure the capacity of docetaxel to accumulate cells in the G2/M phase of the cell cycle. We observed a weak accumulation of cells in the G2/M phase for the ZMK-1 cells and a pronounced accumulation for CaSki cells. For docetaxel incubation before irradiation, the isoeffect enhancement ratios for ZMK-1 cells determined at the 37% survival level were 1.18, 2.01, and 2.40 for pre-incubation at 3, 9 and 24 hr, respectively; for CaSki cells the ratio was 1.44. For a docetaxel incubation of 24 hr after irradiation, the isoeffect enhancement ratios determined at the 37% survival level were 1.54 and 1.17 for the ZMK-1, and CaSki cells, respectively. A radiosensitizing effect of docetaxel could be demonstrated unambiguously in the two cell lines used. In contrast to our previously published results with paclitaxel, docetaxel seems to be a better radiosensitizer than paclitaxel. PMID- 11275990 TI - Synergistic chemosensitization and inhibition of progression to androgen independence by antisense Bcl-2 oligodeoxynucleotide and paclitaxel in the LNCaP prostate tumor model. AB - Bcl-2 expression is up-regulated in prostate cancer cells after androgen ablation and associated with development of androgen independence and chemoresistance. We recently reported that antisense Bcl-2 oligodeoxynucleotides (ODNs) delay progression to androgen independence in the androgen-dependent (AD) human LNCaP prostate tumor model. The objectives in this study were to determine whether antisense human Bcl-2 ODN enhances chemosensitivity of paclitaxel and whether combined antisense Bcl-2 ODN and paclitaxel further delays time to androgen independent (AI) progression in the LNCaP tumor model. Semi-quantitative reverse transcriptast-polymerase chain reaction revealed that treatment of LNCaP cells with antisense Bcl-2 ODN decreased Bcl-2 expression in a dose-dependent and sequence-specific manner, whereas Bcl-2 expression was not affected by paclitaxel treatment. Antisense Bcl-2 ODN treatment significantly enhanced paclitaxel chemosensitivity in vitro, reducing cell viability after treatment with 1 nM paclitaxel from 76% to 42%. Characteristic apoptotic DNA laddering was demonstrated after combined treatment with 500 nM antisense Bcl-2 ODN and 1 nM paclitaxel but not with either agent alone. Adjuvant in vivo administration of combined antisense Bcl-2 and polymeric micellar paclitaxel after castration resulted in a significant delay of emergence of AI recurrent LNCaP tumors compared with either agent alone. By 15 weeks post castration, tumor volume in mice treated with antisense Bcl-2 ODN alone or mismatch control ODN plus paclitaxel was >3-fold higher than in mice treated with combined antisense Bcl-2 ODN and paclitaxel. Mean serum prostate-specific antigen levels returned to or were above precastration levels by 11 weeks post castration in mice treated with antisense Bcl-2 ODN alone or mismatch control ODN plus paclitaxel but remained 90% below the pre-castration level in mice treated with combined antisense Bcl-2 ODN and paclitaxel. These findings identify combined antisense Bcl-2 and paclitaxel as a potentially new therapeutic strategy for advanced prostate cancer by enhancing paclitaxel chemosensitivity and delaying progression of hormone refractory prostate cancer. PMID- 11275992 TI - Quantification of matrix metalloproteinase activity in plasma of patients enrolled in a BAY 12-9566 phase I study. AB - The expression of matrix metalloproteinases (MMPs) is often associated with invasiveness or grade of tumours. Increased blood levels of MMP proteins, including MMP-1, MMP-2, MMP-3 and MMP-9 have been detected in various types of cancers. With the exception of one study, MMPs in serum and plasma have been determined using ELISA. In the present study we measured the activity of the MMPs found in human plasma samples using gelatin enzymography and fluorimetric degradation assays. We used plasma samples from healthy control subjects and cancer patients enrolled in a dose-finding study for the MMP inhibitor, BAY 12 9566, to assess the activity of MMPs found in plasma and screen for efficacy of the MMP inhibitor. BAY 12-9566 has inhibitory activity toward MMP-2, MMP-3 and MMP-9. Patients with advanced solid tumours were enrolled in our study and plasma was collected on day 1 before dosing and at steady-state of the drug on day 15. Our results show that BAY 12-9566 was effective in lowering the plasma gelatinolytic activity in the group of 29 patients when considering the data obtained from a fluorimetric gelatinase assay. The data obtained from gelatin enzymography, however, did not reach significance. The fluorimetric degradation assay could be a useful tool to screen plasma from cancer patients in other clinical trials assessing MMP inhibitors. PMID- 11275991 TI - Adenovirus-mediated gene therapy specific for small cell lung cancer cells using a Myc-Max binding motif. AB - Recent clinical trials of gene therapy for patients with thoracic cancers have shown that these treatments were well tolerated with minimal side effects and that we need to further enhance specificity as well as efficiency of gene transfer to target cancer cells. We previously reported that myc-overexpressing SCLC cell lines became selectively sensitive to ganciclovir (GCV) by transducing the herpes simplex virus thymidine kinase (HSV-TK) gene under the control of the Myc-Max response elements (a core nucleotide sequence, CACGTG) and that this construct (MycTK) could be utilized to develop a novel treatment against chemo radio-resistant SCLC. We report here in vivo antitumor effects and safety of a replication-deficient adenoviral vector containing the Myc-Max binding motif (AdMycTK) on SCLC cells. In vitro infection with AdMycTK selectively rendered myc overexpressing SCLC cell lines 63- to 307-fold more sensitive to GCV. In vivo injections with AdMycTK followed by GCV administration markedly suppressed the growth of myc-overexpressing tumors established in the subcutis or in the peritoneal cavity of athymic mice. On the other hand, infection with AdMycTK did not significantly affect either in vitro GCV sensitivity of the cells expressing very low levels of the myc genes or the growth of their subcutaneous tumors. Moreover, we observed no apparent side effects of this treatment including body weight loss or biochemical abnormalities in contrast to the treatment with AdCATK that conferred strong but nonspecific expression of the HSV-TK gene. These results suggested that AdMycTK/GCV therapy is effective on SCLC patients whose tumors overexpress myc family oncogenes. PMID- 11275993 TI - Mutations of p53 gene in human colorectal cancer: distinct frameshifts among populations. AB - We analyzed 57 p53 gene mutations in 181 colorectal cancer patients in Taiwan and compiled data on 475 independent p53 mutations in 1,156 primary colorectal cancer patients worldwide between 1992 to 1998. Transitions at the CpG sites were observed in 31 (54%) and 232 cases (49%), respectively. Frameshift mutations occurring within exons were observed in 11 (20%) and 50 cases (10%), respectively. Among the various populations studied, colorectal cancer in Taiwan had the lowest p53 mutation rate (31%), highest frequency (20%) of frameshift mutations and the second lowest rate (13%) of transversion mutation. Based on their relation to the base runs, the 61 frameshift mutations could be grouped into 4 subclasses. After corrections were made for differences in the base number in a run, the relative mutational frequency at a base run was found to be 9- to 47-fold over that in the no-run residues. The p53 frameshift mutational spectrum found in the cases in Taiwan, with respect to hotspot sequence, was significantly different from those in the selected database (p = 0.008). These data support that the patterns of high frequency of transitions at CpG sites and low frequency of transversions in base substitutions in the p53 gene are similar regardless of patient origin. However, these data also illustrate that frameshift mutations in the p53 gene in colorectal cancer patients are sequence dependent and are distinct among populations. PMID- 11275994 TI - Lymphoproliferative disorders in renal transplant patients in Japan. AB - Post-transplantation lymphoproliferative disorders (PT-LPD) are characterized by a clinically and morphologically heterogeneous group of lymphoid proliferation occurring after organ or bone marrow transplantation. The immunodeficient state provides a basis for lymphomagenesis probably through activation of oncogenic viruses. Twenty-four patients in whom PT-LPD developed after renal transplantation in Japan were analyzed. They received hemodialysis for 4 to 226 (median 13) months before transplantation. In situ hybridization was performed to detect Epstein-Barr virus (EBV). Polymerase chain reaction and Southern hybridization with primers in the tax and pol regions of human T-cell leukemia virus type I (HTLV-1) were performed on DNA extracted from paraffin-embedded specimens. Immunohistochemical analysis revealed that 12 cases were B-cell type, 10 cases (42%) T-cell type and 2 NK-cell type. Five of the T-cell cases were classified as adult T-cell lymphoma with proven HTLV-1 genome in the tumor and seropositivity for the virus. These cases were classified as adult T-cell lymphoma (ALT). More than 80% of B-cell, 30% of T-cell and both NK/T-cell lymphomas were EBV-positive. Co-infection of EBV and HTLV-1 was found in 2 cases with ATL. These findings showed that ATL is common among Japanese renal transplant patients, which might be due to transmission of HTLV-1 via blood transfusion during hemodialysis. PMID- 11275996 TI - Stress of daily activities and risk of breast cancer: a prospective cohort study in Finland. AB - The belief that life stress enhances breast cancer is common, but there are few prospective epidemiological studies on the relationship of life stress and breast cancer. We have investigated the association between stress of daily activities (SDA) and breast-cancer risk in a prospective cohort study of 10,519 Finnish women aged 18 years or more. SDA measures a subject's own appraisal of daily stress. It was assessed in 1975 and 1981 by a self-administered questionnaire, which also provided information on subject characteristics and other known breast cancer risk factors. Follow-up data for breast cancer from 1976 to 1996 were attained through record linkage to the Finnish Cancer Registry. Study subjects were divided into 3 groups based on their SDA scores in 1975: no stress (23% of subjects), some stress (68%) and severe stress (9%). Hazard ratios (HRs) and respective 95% confidence intervals (CIs) for incidence of breast cancer by level of SDA were obtained from the Cox proportional hazards model. We identified 205 incident breast cancers in the cohort. Multivariable-adjusted HRs for breast cancer risk were 1.00 (reference), 1.11 (95% CI 0.78-1.57) and 0.96 (95% CI 0.53 1.73) by increasing level of stress. Neither shifting of the SDA cut-off points nor restricting the analysis to women who reported the same level of SDA in 1975 and 1981 materially altered the results. We found no evidence of an association between self-perceived daily stress and breast-cancer risk. PMID- 11275995 TI - Lung cancer and cigarette smoking in Europe: an update of risk estimates and an assessment of inter-country heterogeneity. AB - Ten case-control studies have been carried out in 6 European countries to investigate the major risk factors for lung cancer. Carcinogenic effect from cigarette smoke was the most relevant interest in our study, which has included 7,609 cases of lung cancer and 10,431 controls, mainly population based. The results indicate elevated odds ratios (ORs; 23.9 among men and 8.7 among women) with attributable risks exceeding 90% for men and close to 60% for women. A large, and statistically significant, variability of the results across countries was detected after adjusting for the most common confounding variables, and after controlling, at least in part, for the instability of the ORs due to the small number of non-smokers in some of the study subsets. This pattern of lung cancer risk associated with cigarettes smoke, across different European regions, reflects inherent characteristics of the studies as well as differences in smoking habits, particularly calendar periods of starting, and it is likely to have been influenced by effect modifiers like indoor radon exposure, occupation, air pollution and dietary habits. PMID- 11275999 TI - Tumour versus patient: vascular and tumour survival versus prognosis. AB - The concept that malignant solid tumour growth depends on angiogenesis is widely recognized. For some tumour types, there is a measurable range of vascularity and the link between prognosis and increased vascular density, best observed at the hotspots at the edge, is now established. What is less discussed are the corollaries: that tumour invasion requires tissue destruction; that the neovasculature must be not only protected but also sustained, especially at the tumour edge; that for tumour survival the edge is the future and the centre is history; and that angiogenesis is essential not only for tumour growth but also for tumour invasion. Different patterns of vascular density in tumour edge and centre have been observed, and these are linked to lymphatic spread and prognosis. The variation is attributable to differing interactions between endothelium and the tumour cell that dictate vascular and tumour survival; this may become relevant to anti-angiogenesis therapies. PMID- 11275997 TI - Tumor invasiveness and liver metastasis of colon cancer cells correlated with cyclooxygenase-2 (COX-2) expression and inhibited by a COX-2-selective inhibitor, etodolac. AB - Non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to reduce the risk and mortality of colorectal cancer (CRC). Although the exact mechanisms remain unclear, the inhibition of cyclooxygenase (COX) by NSAIDs appears to abort, if not prevent, CRC carcinogenesis or metastatic tumor progression. The aim of our study was to investigate the association between COX-2 expression and CRC tumor cell invasiveness. The differences in immunoblot-detectable COX-2 protein contents in primary CRCs, metastatic hepatic lesions and corresponding normal mucosa from the same individual were evaluated in 17 patients. Three different colon cancer cell lines, SW620, Lovo, HT-29 and a metastatic variant of HT-29, HT-29/Inv3, were employed to evaluate COX-2 expression and prostaglandin E(2) (PGE2) production in relation to their invasive abilities in vitro. The effects of a COX-2-selective inhibitor, etodolac, on cell proliferation and invasive activity were also determined. The results showed that 15 of 17 (88%) metastatic CRC cells from the liver and 14 of 17 (82%) primary CRC tissue exhibited much higher levels of COX-2 than corresponding adjacent normal mucosa from the same patient. Among those patients with relatively high COX-2 expression in the primary tumors, almost all exhibited even higher levels of COX-2 in their hepatic metastases. Among the 4 colon cancer cell lines, HT-29/Inv3 manifested the highest COX-2 expression, PGE2 production and in vitro invasive activity. The selective COX-2 inhibitor, etodolac, could especially exert cytotoxicity and markedly suppress the invasive property and PGE(2) production, although not the COX-2 protein level, in HT-29/Inv3 cells. Our results imply that COX-2 expression may be associated with the invasive and metastatic properties of CRC tumor cells. PMID- 11276000 TI - The Hedgehog signalling pathway and cancer. AB - The Hedgehog signalling pathway is important in embryological development and is highly conserved through evolution. Recently Patched, a member of the pathway, was found to be important in Gorlin's syndrome. Inherited Patched gene mutations underlie the syndrome, in which a key feature is multiple basal cell carcinomas (BCCs). The gene is also mutated in sporadic BCCs as well as in sporadic occurrences of other tumours seen in Gorlin's syndrome. The precise mechanism whereby Patched gene mutation leads to tumour development is not known, but BCC is characterized by relentless local invasion and only rarely metastasizes. This suggests that abnormalities of the Hedgehog pathway account for these features. This proposal is discussed in the context of what is already known about the normal function of the Hedgehog pathway and its deregulation in cancer. PMID- 11276001 TI - Retention of the expression of E-cadherin and catenins is associated with shorter survival in grade III ductal carcinoma of the breast. AB - Many studies have investigated the relationship between the E-cadherin/catenin axis and breast cancer biology and yet, unlike the studies in other tumour systems, which have shown a relationship between down-regulation and poor survival, no clear association has emerged in breast. Since accumulating evidence suggests that ductal carcinoma of no special type (NST) represents a diverse group of biologies, this study has focused on grade III ductal carcinoma, in order to reduce the heterogeneity of the study population. A total of 470 breast tumours were studied. Consecutive sections were labelled with antibodies which recognize E-cadherin and the arm proteins with which it interacts: alpha-, beta-, and gamma-catenin. Membrane-bound and cytoplasmic E-cadherin and membrane-bound alpha-catenin expression were associated with a positive oestrogen receptor (ER) status, gamma-catenin with a negative ER status, and, surprisingly, all three with poor survival. Taken together, these findings suggest that a conserved E cadherin/catenin axis may play a part in determining adverse outcome in grade III breast carcinoma. PMID- 11276002 TI - Early distant relapse in "node-negative" breast cancer patients is not predicted by occult axillary lymph node metastases, but by the features of the primary tumour. AB - Early distant relapse occurs in a minority of node-negative breast cancer patients. Whether this poor prognosis can be predicted by the features of the primary tumour, or by the presence of occult metastases in the "negative" lymph nodes (LNs), remains a matter of debate. One hundred and four T(1-2)N(0)M(0) breast carcinoma patients were divided into two groups: group 1 (44%) showing early distant relapse with a median disease-free survival of 25 months, and group 2 (56%) showing no evidence of disease after a median follow-up of 91.5 months. All patients had received locoregional treatment only. All tumours were evaluated for medial/lateral location, histological type, size, grade, mitotic activity, fibrotic focus, necrosis, angiogenesis, growth pattern, and lymphatic vessel permeation. The haematoxylin and eosin-stained slides of all axillary LNs were revised and two additional levels were cut from each paraffin block for cytokeratin immunohistochemistry. In 24 patients (23%), occult metastases were found. These consisted of single cells or small clusters (SCs) in the marginal sinus in 17 patients (16%) and of larger colonies of cells in seven patients (7%). All detected metastases were smaller than 2 mm in diameter (micrometastases). There was no significant correlation between the presence of occult LN metastases (SCs or colonies) and the prognostically important features of the primary tumour. Early metastatic disease was significantly correlated with larger tumour size (p=0.02), higher histological grade (p=0.0008), mitotic activity (p<0.0001), presence of necrosis (p=0.0004), presence of fibrotic foci (p=0.0005), angiogenesis (p=0.0009), and lymphatic vessel permeation (p=0.018). Multiple logistic regression analysis showed that histological grade and the presence of a fibrotic focus were the only independent prognostic factors and that the presence of occult LN metastases was inversely correlated with early distant relapse. Prospective prognostic studies of occult LN metastases should consider the features of the primary tumour in a multivariate analysis. PMID- 11276003 TI - Expression of vascular endothelial growth factor receptor 3 in blood and lymphatic vessels of lung adenocarcinoma. AB - Vascular endothelial growth factor receptor 3 (VEGFR-3) has been proposed as a marker for lymphatic endothelial cells. This study investigated the expression of VEGFR-3 in the tumour vessels of lung adenocarcinoma and evaluated whether VEGFR 3 staining was useful for identifying lymphatic vessels within the tumour stroma. It also explored whether active growth of lymphatic vessels occurred in lung adenocarcinoma. Formalin-fixed, paraffin-embedded specimens obtained from 60 cases of lung adenocarcinoma, including five cases of pure bronchiolo-alveolar carcinoma (BAC) without stromal, vascular, and pleural invasion, were examined. No VEGFR-3-positive vessels were observed in pure BAC, but varying numbers of VEGFR-3-positive vessels were found in 39 of 55 (70.9%) invasive adenocarcinomas. A comparison of serial sections stained for VEGFR-3, CD31, and laminin-1 showed that most of the VEGFR-3-positive vessels appeared to be blood vessels (CD31 positive, laminin-1-positive), but some had the characteristics of lymphatic vessels (variable staining for CD31, little or no staining for laminin-1). VEGFR 3 staining highlighted lymphatic invasion by cancer cells; this invasion could not be detected by CD31 or haematoxylin and eosin (H&E) staining. Active growth of lymphatic vessels (as indicated by nuclear Ki-67 labelling of the endothelium) was observed in five tumours, four of which showed a high level of lymphatic invasion by cancer cells. It was concluded that VEGFR-3 immunostaining did not discriminate clearly between vascular and lymphatic endothelial cells, since expression of VEGFR-3 can be up-regulated in tumour blood vessels. However, VEGFR 3 staining combined with laminin-1 and CD31 staining would be useful for identifying lymphatic vessels and their invasion by tumour cells in a more objective way. Finally, proliferation of lymphatic endothelial cells may occur in association with lymphatic invasion by cancer cells. PMID- 11276004 TI - Decreased expression of TGF-beta cell surface receptors during progression of human oral squamous cell carcinoma. AB - This study examined the immunocytochemical expression of the transforming growth factor-beta (TGF-beta) isoforms TGF-beta1, TGF-beta2, and TGF-beta3, together with the TGF-beta cell surface receptors TbetaR-I and TbetaR-II, in patient matched tissue pairs of normal human oral epithelium, primary squamous cell carcinomas, and metastatic lymph node tumour deposits. There were no significant differences in the intensity of TGF-beta isoform specific staining between the normal oral epithelium, the primary tumours, and the lymph node metastases. By contrast, there was significantly less TbetaR-II in the metastases than in the primary tumour and between the primary tumour and the normal oral epithelium. Similar trends were evident with TbetaR-I, but not at a statistically significant level. This study also examined the structure of TbetaR-I and TbetaR-II in normal human oral keratinocytes in vitro and in 14 human oral carcinoma cell lines with known responses to TGF-beta1. No structural abnormalities of TbetaR-II were present in the normal keratinocytes or in 13 of 14 malignant cell lines; in one line, there were both normal and mutant forms of TbetaR-II, the latter being in the form of a frameshift mutation with the insertion of a single adenine base (bases 709-718, codons 125-128), predicting a truncated receptor having no kinase domain. No defects were present in TbetaR-I. The structures of TbetaR-I and TbetaR-II did not correlate with growth inhibition by TGF-beta1. The data suggest that decreased expression of TGF-beta receptors, rather than structural defects of these genes, may be important in oral epithelial tumour progression. In order to examine the functional significance of a specific decrease in TbetaR-II expression, a dominant-negative TbetaR-II construct (dnTbetaR-II) was transfected into a human oral carcinoma cell line with a normal TGF-beta receptor profile and known to be markedly inhibited by TGF-beta1. In those clones that overexpressed the dnTbetaR-II, growth inhibition and Smad binding activity were decreased, whilst the regulation of Fra-1 and collagenase-1 remained unchanged following treatment with TGF-beta1. The results demonstrate that a decrease in TbetaR-II relative to TbetaR-I leads to selective gene regulation with loss of growth inhibition but continued transcription of AP-1-dependent genes that are involved in the regulation of the extracellular matrix. PMID- 11276005 TI - Vascular endothelial growth factor is an autocrine growth factor in human malignant mesothelioma. AB - Vascular endothelial growth factor (VEGF), a potent mitogen for vascular endothelium, is expressed in malignant pleural mesothelioma (MM). The present report examines the effect of VEGF on MM growth. Four MM cell lines produced significantly higher VEGF levels than normal mesothelial cells (1946+/-14 pg/ml vs. 180+/-17 pg/ml; p<0.001). In addition, MM cells expressed the tyrosine kinase related VEGF receptors Flt-1 and KDR. Recombinant human VEGF phosphorylated both Flt-1 and KDR and increased proliferation of all four MM cell lines in a dose dependent fashion. Neutralizing antibodies against either VEGF, Flt-1 or KDR significantly reduced MM cellular proliferation. In addition, expression of VEGF, Flt-1, and KDR was observed in MM biopsies. Moreover, higher VEGF levels were found in the pleural effusions of MM patients than in the effusions of patients with non-malignant pleural disease (1885.7+/-894.9 pg/ml vs. 266.9+/-180.5 pg/ml; p<0.001). Linear regression analysis showed a significant inverse correlation between serum VEGF levels and MM patient survival (r=0.72; p<0.01). No correlation was found between tumour vessel density and either serum (r=0.26; p=0.42) or pleural effusion (r=0.35; p=0.26) VEGF levels. These results indicate that VEGF, via activation of its tyrosine kinase receptors, may be a key regulator of MM growth. In addition, VEGF production could have an impact on patient survival, not only by promoting tumour angiogenesis but also by directly stimulating tumour growth. PMID- 11276006 TI - Differentiation of liver cell adenomas from well-differentiated hepatocellular carcinomas by comparative genomic hybridization. AB - Liver cell adenomas (LCAs) are rare tumours which may be difficult to differentiate from low-grade hepatocellular carcinomas (HCCs). This study used comparative genomic hybridization (CGH) to look for cytogenetic aberrations which would serve to distinguish between these tumours. For this purpose, ten LCAs and six well-differentiated HCCs were analysed and the results were compared with those reported previously for 15 well-differentiated HCCs. Aberrations were seen in 2/10 LCAs: a gain of chromosome 7p was observed in one and gains of 17q and 20 in a second case. In 6/6 well-differentiated HCCs, up to 13 aberrations were detectable, with a mean of 7.2 aberrations per case in chromosome sites 1q, 4p, 4q, 5p, 5q, 6p, 6q, 7p, 7q, 8p, 8q, 10q, 11p, 13q, 14q, 16p, 16q, 17p, 17q, 20p, 20q, and 21q. Aberrations focused on gains or losses of six chromosome sites, 1q, 4q, 8p, 8q, 16p, and 17p; in all HCC samples, at least two of these sites were affected. None of these aberrations occurred in any of the LCAs analysed. CGH is therefore helpful in distinguishing between LCA and well-differentiated HCC. Detection of one or more of the six most frequent aberrations in HCC supports the diagnosis of carcinoma and makes LCA unlikely. PMID- 11276007 TI - Nuclear localization of beta-catenin is an important prognostic factor in hepatoblastoma. AB - In this study, mutational and immunohistochemical analyses of beta-catenin were performed in 30 hepatoblastomas, to assess the prevalence of alterations of the Wnt pathway with respect to clinicopathological parameters and survival. Four missense mutations of beta-catenin (13.3%) were detected and there was strong immunoreactivity for beta-catenin in the cytoplasm and/or the nucleus in 97% of hepatoblastomas. Nuclear and cytoplasmic staining was demonstrated in 19 of 30 tumours (63%), while ten revealed only cytoplasmic staining. Statistically, this nuclear beta-catenin staining was significantly higher in the embryonal (Fisher exact test; p=0.00393) or undifferentiated type (p=0.00156) of hepatoblastoma than in the fetal type, but there was no difference between clinical stages I and II and clinical stages III and IV (p=0.175). Cumulative survival curves showed that nuclear beta-catenin staining (generalized Wilcoxon test; p=0.0088), undifferentiated histological type (p=0.0305), and clinical stages III and IV (p=0.0107) were significantly correlated with shorter survival time in these patients. Moreover, Cox multivariate analysis provides evidence that nuclear beta catenin staining is the most important prognostic factor for survival (p=0.0090). It is therefore concluded that immunohistochemical analysis of beta-catenin might be a useful clinical tool for estimating the prognosis for patients with hepatoblastoma. PMID- 11276008 TI - CDKN2A/p16 inactivation is related to pituitary adenoma type and size. AB - p16 (CDKN2A, MTS1, INK4A) status at genomic and protein levels was analysed and correlated with clinico-pathological features in 72 pituitary adenomas. Methylation of CpG islands of promoter/exon 1 sequences was found in most gonadotroph, lactotroph, plurihormonal, and null cell adenomas (36 of 44, 82%), but it was rare in somatotroph (1 of 13 cases, 8%) and corticotroph adenomas (1 of 15 cases, 7%). Homozygous CDKN2A deletion was restricted to rare somatotroph (15%) and corticotroph adenomas (13%). Immunohistochemical p16 protein expression was observed in the normal adenohypophysis, whereas it was absent in 60 of 72 (83%) tumours and reduced in another ten (14%) tumours. Staining for p16 was only seen in 5 of 15 (33%) corticotroph, 3 of 13 (23%) somatotroph, 3 of 5 (60%) plurihormonal, and 1 of 19 (5%) null cell adenomas. p16 immunonegativity without CDKN2A methylation or deletion occurred in 22 tumours, including most somatotroph and corticotroph adenomas (15 of 28, 54%). Both CDKN2A alterations and p16 negativity were related to larger tumour size. Patients with p16-negative tumours were older than patients with p16-positive tumours. These data suggest that p16 down-regulation is common in all adenoma types. The mechanisms of p16 down regulation probably involve CDKN2A methylation in most types, but remain to be determined in somatotroph and corticotroph adenomas. These findings also suggest that p16 down-regulation is usually not an initial event, but is acquired during adenoma progression. PMID- 11276009 TI - Relative distribution of tumour cells and reactive cells in follicular lymphoma. AB - Follicular lymphoma is the most common low-grade B-cell lymphoma. It is characterized by at least a partial follicular growth pattern in the majority of cases, by the morphological resemblance of the tumour cells to follicle centre centroblasts and centrocytes, and by the distinctive expression of Bcl-2 protein as a consequence of a translocation between chromosomes 14 and 18, resulting in the juxtaposition of Bcl-2 and the immunoglobulin heavy chain locus. It is not known whether the follicular growth pattern of follicular lymphoma is a consequence of properties of the tumour cells, or whether the tumour cells invade and gradually occupy a niche generated by a normal T-cell-dependent B-cell response. This study has identified cases of follicular lymphoma in which the tumour cells are apparent within a normal reactive germinal centre background. The reactive background has been investigated in these cases and also in cases showing a more characteristic appearance, in which entire malignant follicles appear to be Bcl-2-positive, as assessed by microdissection and analysis of clonality by the polymerase chain reaction (PCR). A reactive oligoclonal background was observed in all cases studied, characteristic of a normal follicle centre response. These data suggest that the progression of follicular lymphoma is dependent on the normal germinal centre microenvironment. Disruption of this dependence might be considered as a novel therapeutic strategy. PMID- 11276010 TI - Gain-of-function mutation at the extracellular domain of KIT in gastrointestinal stromal tumours. AB - Gastrointestinal stromal tumours (GISTs) are the most common mesenchymal tumours of the human gastrointestinal tract. Previous studies of GISTs found gain-of function mutations of the c-kit gene, which encodes a receptor tyrosine kinase (KIT). All the mutations were confined to exon 11, which encodes the juxtamembrane domain. By further examination of the whole coding region of c-kit complementary DNA in 35 GISTs, two were found to show the identical mutation at exon 9, which encodes the extracellular domain. The aims of the present study were to examine the frequency of the extracellular domain mutation and to determine whether the mutation is a gain-of-function type or not. Genomic DNA was extracted from paraffin-embedded tissues of 133 GISTs and exon 9 of the c-kit gene was amplified by polymerase chain reaction. Screening of the mutation was carried out by single-strand conformation polymorphism analysis and direct sequencing was done. Mutant c-kit cDNA was transfected into 293T human embryonic kidney cells and the magnitude of autophosphorylation of the mutant KIT was examined with or without the ligand of KIT, stem cell factor (SCF). In total, seven GIST cases (approximately 5%) were found with the identical mutation at exon 9. The mutant KIT exhibited constitutive autophosphorylation without SCF stimulation. The prognosis of the patients with the extracellular domain mutation was comparable to that of the patients with the juxtamembrane domain mutation. PMID- 11276011 TI - Localized and systemic scleroderma show different histological responses to methotrexate therapy. AB - Although morphoea (localized scleroderma) and systemic sclerosis are distinct disease entities, the skin lesions show identical histological characteristics and both diseases respond favourably to low-dose treatment with methotrexate (MTX). The aim of this study was to find out whether MTX treatment induces different histological response patterns in these two diseases. In seven patients with morphoea and eight with systemic sclerosis, skin biopsies were taken before and after 24 weeks of treatment with low-dose MTX. In the centre and active margin of morphoea lesions, a significant reduction in tenascin staining was seen after 24 weeks of treatment, in contrast to systemic sclerosis. The numbers of mast cells decreased in the active margin of morphoea lesions, whereas in systemic sclerosis no significant change was seen after MTX therapy. Epidermal proliferation and staining of heparan sulphate proteoglycans showed no changes. Although skin lesions from both diseases respond clinically to treatment with MTX, systemic sclerosis shows no change in the immunohistochemical parameters investigated, whereas morphoea does. This difference in dynamic pattern suggests that the apparently similar lesions in localized and systemic sclerosis are not identical. PMID- 11276012 TI - Differences in susceptibility to colonic stem cell somatic mutation in three strains of mice. AB - Different species and different strains of animals commonly show very different sensitivities to carcinogenic regimes, which are often unexplained. A major possible contributory factor is variation in susceptibility to mutation, but this has not been directly demonstrated. This study therefore quantified the colonic stem cell mutation frequency in three strains of mice using two carcinogens. Stem cell mutations were identified using loss of function of glucose 6-phosphate dehydrogenase (G6PD) in individual crypts, a technique validated by several previous studies. The carcinogens dimethylhydrazine (DMH) and ethyl nitrosurea (ENU) were given to Balb/C, C57BL/6J, and C3H mice. In response to DMH, Balb/C mice were most susceptible, with approximately double the stem cell mutation frequency found in C3H and more than ten-fold that found in C57BL/6J (3.3+/-0.71 vs. 1.5+/-0.52 vs. 0.28+/-0.8x10(-4)). In response to ENU, Balb/C mice and C3H mice were equally susceptible, showing a stem cell mutation frequency approximately twice that of C57BL/6J (3.1+/-0.4 vs. 3.1+/-0.65 vs. 1.63+/ 0.28x10(-4)). The observed differences among the strains with respect to somatic mutation following DMH treatment are likely to be due to the previously documented differences in metabolic conversion to the active metabolite. However, as ENU is a directly acting, rapidly inactivated mutagen, strain differences in response to ENU are unlikely to be due to strain-dependent metabolism of the mutagen and are likely to reflect differences in DNA repair efficiency, or possibly in stem cell kinetics among the strains studied. Susceptibility to the induction of colonic stem cell mutation is an important factor in susceptibility to carcinogens, whether due to differences in DNA repair or to other factors. Direct quantification of stem cell mutation frequency allows the separate identification of this component of the carcinogenic cascade and shows that it can make a major contribution to the differing susceptibility of different mouse strains. PMID- 11276013 TI - Differential expression of cell death regulators in response to thapsigargin and adriamycin in Bcl-2 transfected DU145 prostatic cancer cells. AB - Functional overexpression of Bcl-2 has been reported to confer an anti-apoptotic potential in a variety of cell types. The role of Bcl-2 in epithelial cell-cycle control and in interactions with other cell-cycle regulators is not clearly understood. Its expression has been correlated with the hormono- and chemo resistant phenotype in advanced prostate cancer. The aim of this study was to investigate the mechanisms through which Bcl-2 mediates increased cytotoxic chemoresistance by assessing alterations in the expression of cell death regulatory molecules. The DU145 human prostatic adenocarcinoma cell line was stably transfected with a Bcl-2 encoding expression plasmid. Two Bcl-2 transfectants, DKC9 and DKC11, were expanded for further study. The effects of Bcl-2 expression on cellular proliferation, cell death (+/- adriamycin or thapsigargin), and expression of cell-cycle/death regulators (p53, PCNA, Bax, Bak, Bcl-X(L)) were evaluated. Compared with controls, Bcl-2 transfectants showed no difference in the rate of proliferation, a decrease in p53 (approximately two fold), an increase in Bax (approximately two-fold) and PCNA (approximately three fold), and no change in the levels of Bcl-X(L) and Bak proteins. DKC9 and DKC11 also exhibited a significantly increased chemoresistance to adriamycin (0.0025-5 microM) and thapsigargin (0.0025-5 microM) compared with controls. In the presence of thapsigargin or adriamycin, levels of Bcl-2 and its heterodimeric partner Bax were elevated approximately two-fold with no change in Bak in Bcl-2 transfectants in contrast to controls, where Bak was increased (two-fold). This is the first study to demonstrate that Bcl-2 transfection modulates the expression of mutant p53, Bax, and PCNA in prostate cancer cells. Moreover, Bcl-2 overexpression conferred a significant cytotoxic chemoresistance and altered the balance of expression of death promoters (from Bak, a dominant death promoter in controls, to Bax) in response to thapsigargin and adriamycin. PMID- 11276014 TI - Effect of inflammatory cytokines and growth factors on tumour cell adhesion to the peritoneum. AB - In this experimental study, the effect of inflammatory cytokines and growth factors on tumour cell adhesion to the peritoneum was investigated. A reproducible in vitro assay was developed to study the adhesion of CC531 colon carcinoma cells to an autologous monolayer of rat mesothelial cells. Tumour cell adhesion to mesothelium pre-incubated with interleukin-1beta (IL-1beta) and epidermal growth factor (EGF) resulted in at least 60% more tumour cell adhesion at maximal stimulation (p dihydroazadirachtin (3.16 x 10(-8) M) > dansyl dihydroazadirachtin (7.40 x 10(-8) M) > DNP-azadirachtin (7.50 x 10( 8) M) > biotin dihydroazadirachtin (1.27 x 10(-7) M) >> 11-methoxy 22,23 dihydroazadirachtin (6.67 x 10(-7) M). [Originally published in Volume 34, Archives of Insect Biochemistry and Physiology, 34:461-473 (1997).] PMID- 11276069 TI - Third instar nymphs of Rhodnius prolixus exposed to alpha-cyanopyrethroids: from hyperactivity to death. AB - The hyperactivity, incoordination, recovery, and mortality produced by four alpha cyanopyrethroids usually used for Chagas disease vector control (beta cypermethrin, beta-cyfluthrin, lambda-cyhalothrin, and deltamethrin) were evaluated on third instar nymphs of Rhodnius prolixus. All pyrethroids modified the locomotor activity of the nymphs, which increased linearly as a function of the log of insecticide concentration. lambda-Cyhalothrin showed the lowest values of Effective Concentration 50%, Lethal Concentration 50%, Effective Time 50%, and Lethal Time 50% when insecticides were applied by contact with treated filter papers. Recovery from incoordination was observed after topical application of the insecticides. The recovery was inhibited by the simultaneous application of piperonyl butoxide, suggesting that biotransformation by mixed-function microsomal oxidases is involved in the process of recovery. PMID- 11276070 TI - Significance of absorption, oxidation, and binding to toxicity of four ecdysone agonists in multi-resistant cotton leafworm. AB - Treatment of last-instar larvae of multi-resistant cotton leafworm Spodoptera littoralis with four dibenzoylhydrazines, methoxyfenozide (RH-2485), tebufenozide (RH-5992), halofenozide (RH-0345), and RH-5849, resulted in premature molting leading to death. Methoxyfenozide was the most toxic followed by tebufenozide, halofenozide, and RH-5849. To explain differences in toxicity, especially between multi-resistant and laboratory strains, absorption in the body tissues and oxidative metabolism were tested with 14C-labeled ecdysone agonist and a Lineweaver-Burk assay, respectively. Then to address different compound potencies in multi-resistant strains, the potency of the four ecdysone agonists was measured based on their ability to mimic the natural insect molting hormone, 20 hydroxyecdysone (20E) by inducing evagination in isolated imaginal wing discs. Using monoclonal antibody 9B9, the presence of ecdysteroid receptors in imaginal discs in vitro was confirmed. In parallel, Scatchard plot analysis with whole imaginal wing discs cultured with different concentrations of 3H-labeled ponasterone A indicated no significant difference in affinity and in number of target sites for binding between multi-resistant and susceptible laboratory strains. The four compounds tested caused the effect as agonists of 20E in vitro, and typically the order of their toxicities (LC50s) corresponded with that for evagination-induction with whole imaginal discs. PMID- 11276071 TI - Proteins synthesized and secreted by larvae of the ectoparasitic wasp, Eulophus pennicornis. AB - A microscopic examination of Eulophus pennicornis larvae on their host Lacanobia oleracea, revealed that peristaltic waves travelled from the anterior to posterior end of the feeding wasp larvae, and vice versa. In addition, when wasp larvae were immersed in PBS in vitro, they released a variety of proteins, with molecular weights ranging from (at least) 14 to 200 kDa. Amongst these was a protein with an estimated molecular weight similar to that of the 27 kDa parasitism-specific protein (PSP) detected in plasma from parasitized L. oleracea [Richards and Edwards, Insect Biochem Mol Biol 29:557-569 (1999)]. Similar results were obtained when the wasp larvae were incubated on balls of cotton wool soaked in tissue culture medium or sucrose, i.e., conditions that resemble their natural feeding behaviour. These results (and others) indicate that the wasp larvae release proteins, putatively through their mouth. Protein synthesis studies using (35)S-methionine indicated that the wasp larvae synthesize and secrete a variety of proteins in vitro, including one with a molecular weight corresponding to that of the L. oleracea 27 kDa PSP. As expected, only a portion of the total proteins synthesized by the parasitoid larvae were subsequently secreted. In addition, the autoradiogram of secreted proteins contained significantly fewer bands than silver-stained SDS gels of proteins released into PBS or onto cotton wool. Thus, some of the additional bands detected on the latter gels are thought to represent proteins that were not of wasp origin. Instead, these proteins released by the wasp larvae are speculated to be derived from their gut and, as such, probably represent proteins derived from host haemolymph and ingested during feeding. This possibility was supported by an electrophoretic analysis of homogenate supernatants prepared from wasp larvae with or without their gut contents. These studies indicated that the gut contents of the larval parasitoid contributes several distinct bands to the total protein profile. The ability of E. pennicornis larvae to synthesize, secrete, and release proteins is discussed with reference to those produced by endoparasitoid larvae. Published 2001 Wiley-Liss, Inc. PMID- 11276072 TI - Age- and diapause-related acid and alkaline phosphatase activities in the intestine and malpighian tubules of the Colorado potato beetle, Leptinotarsa decemlineata (Say). AB - Specific activities for soluble (s) and membrane (m)-bound acid (ACP) and alkaline phosphatases (ALP) were determined in the midgut, hindgut, and Malpighian tubules for developing, prediapausing, and diapausing adult Colorado potato beetles, Leptinotarsa decemlineata (Say). High ACP activities were found in the hindgut and Malpighian tubules while high ALP activities were found in the Malpighian tubules. Variation in both ACP and ALP activities in each tissue reflects fluctuation in protein synthesis and secretion involved with digestion, excretion, and other unknown functions. Phosphatase activities in the tissues examined show the dynamic nature of diapause in this insect. Diapausing beetles showed increases in phosphatase activity after hormone treatments. JHA treatments increased s-ACP and m-ACP activities in all tissues but 20-HE did not increase activity in any tissue. Allatotropin tended to mimic the effects of JHA treatment. The s-ALP activity was also increased in all tissues whereas m-ALP was increased in the midgut and hindgut by JHA treatment. Malpighian tubule m-ALP activity was only increased by 20-HE treatments. Allatotropin was not as effective in increasing ALP activities as it was with ACP activities. PMID- 11276073 TI - Induction of cortical oscillations in spreading cells by depolymerization of microtubules. AB - Actomyosin-based cortical contractility is a common feature of eukaryotic cells but the capability to produce rhythmic contractions is found in only a few types such as cardiomyocytes. Mechanisms responsible for the acquisition of this capability remain largely unknown. Rhythmic contractility can be induced in non muscle cells by microtubule depolymerization. Spreading epithelial cells and fibroblasts in which microtubules were depolymerized with nocodazole or colcemid underwent rhythmic oscillations of the body that lasted for several hours before the cells acquired a stable, flattened shape. By contrast, control cells spread and flattened into discoid shapes in a smooth and regular manner. Quantitative analysis of the oscillations showed that they have a period of about 50 seconds. The kinase inhibitors, HA 1077 and H7, and the more specific rho-kinase inhibitor, Y 27632, caused the oscillations to immediately cease and the cells to become flat. Transient increases in cytoplasmic calcium preceded the contractile phase of the oscillations. Wrinkle formation by cells plated on elastic substrata indicated that the contractility of colcemid-treated cells increased in comparison to controls but was drastically decreased after HA 1077 addition. These data suggest that an intact microtubular system normally prevents pulsations by moderating excessive rho-mediated actin myosin contractility. Possible mechanistic interactions between rho-mediated and calcium activated contractile pathways that could produce morphological oscillations are discussed. PMID- 11276074 TI - Calyculin-A, an inhibitor for protein phosphatases, induces cortical contraction in unfertilized sea urchin eggs. AB - When an unfertilized sea urchin egg was exposed to calyculin-A (CL-A), an inhibitor of protein phosphatases, for a short period and then lysed, the cortex contracted to exclude cytoplasm and became a cup-shaped mass. We call the contracted cortex "actin cup" since actin filaments were major structural components. Electron microscopic observation revealed that the cup consisted of inner electron-dense layer, middle microfilamentous layer, and outermost granular region. Microfilaments were heavily accumulated in the inner electron-dense layer. The middle layer also contained numerous microfilaments, which were determined to be actin filaments by myosin S1 decoration, and they were aligned so that their barbed ends directed toward the outermost region. Myosin II, Arp2, Arp3, and spectrin were concentrated in the actin cup. Immuno-electron microscopy revealed that myosin II was localized to the electron-dense layer. We further found that the cortical tension of the egg increased just after application of CL A and reached maximum within 10 min. Cytochalasin B or butanedione monoxime blocked the contraction, which suggested that both actin filaments and myosin ATPase activity were required for the contraction. Myosin regulatory light chain (MRLC) in the actin cup was shown to be phosphorylated at the activation sites Ser-19 and Thr-18, by immunoblotting with anti-phosphoepitope antibodies. The phosphorylation of MRLC was also confirmed by a (32)P in vivo labeling experiment. The CL-A-induced cortical contraction may be a good model system for studying the mechanism of cytokinesis. PMID- 11276075 TI - Structural and functional effects of hydrostatic pressure on centrosomes from vertebrate cells. AB - In an attempt to better understand the role of centrioles in vertebrate centrosomes, hydrostatic pressure was applied to isolated centrosomes as a means to disassemble centriole microtubules. Treatments of the centrosomes were monitored by analyzing their protein composition, ultrastructure, their ability to nucleate microtubules from pure tubulin, and their capability to induce parthenogenetic development of Xenopus eggs. Moderate hydrostatic pressure (95 MPa) already affected the organization of centriole microtubules in isolated centrosomes, and also impaired microtubule nucleation. At higher pressure, the protein composition of the peri-centriolar matrix (PCM) was also altered and the capacity to nucleate microtubules severely impaired. Incubation of the treated centrosomes in Xenopus egg extract could restore their capacity to nucleate microtubules after treatment at 95 MPa, but not after higher pressure treatment. However, the centriole structure was in no case restored. It is noteworthy that centrosomes treated with mild pressure did not allow parthenogenetic development after injection into Xenopus eggs, even if they had recovered their capacity to nucleate microtubules. This suggested that, in agreement with previous results, centrosomes in which centriole architecture is impaired, could not direct the biogenesis of new centrioles in Xenopus eggs. Centriole structure could also be affected by applying mild hydrostatic pressure directly to living cells. Comparison of the effect of hydrostatic pressure on cells at the G1/S border or on the corresponding cytoplasts suggests that pro-centrioles are very sensitive to pressure. However, cells can regrow a centriole after pressure-induced disassembly. In that case, centrosomes eventually recover an apparently normal duplication cycle although with some delay. PMID- 11276076 TI - Transport and arrangement of the outer-dynein-arm docking complex in the flagella of Chlamydomonas mutants that lack outer dynein arms. AB - The outer dynein arms of Chlamydomonas flagella are attached to a precise site on the outer doublet microtubules and repeat at a regular interval of 24 nm. This binding is mediated by the outer dynein arm docking complex (ODA-DC), which is composed of three protein subunits. In this study, antibodies against the 83- and 62-kD subunits (DC83 and DC62) of the ODA-DC were used to analyze its state of association with outer arm components within the cytoplasm, and its localization in the axonemes of oda mutants. Immunoprecipitation indicates that DC83 and DC62 are preassembled within the cytoplasm, but that they are not associated with outer arm dynein. Both proteins are lost or greatly diminished in oda1 and oda3, mutants in the structural genes of DC62 and DC83, respectively, demonstrating that their association is necessary for their stable presence in the cytoplasm. Immunoelectron microscopy indicates that DC83 repeats at 24-nm intervals along the length of the doublet microtubules of oda6, which lacks outer arms; thus, outer arm periodicity may be determined by the ODA-DC. Flagellar regeneration and temporary dikaryon experiments indicate that the ODA-DC can be rapidly transported into the flagellum and assembled on the doublet microtubules independently of the outer arms and independently of flagellar growth. Unexpectedly, the intensity of ODA-DC labeling decreased toward the distal ends of axonemes of oda6 but not wild-type cells, suggesting that the outer arms reciprocally contribute to the assembly/stability of the ODA-DC. PMID- 11276077 TI - Protein-flavonol interaction: fluorescence spectroscopic study. AB - Recent studies have shown that various synthetic as well as therapeutically active naturally occurring flavonols possess novel luminescence properties that can potentially serve as highly sensitive monitors of their microenvironments in biologically relevant systems. We report a study on the interactions of bovine serum albumin (BSA) with the model flavonol 3-hydroxyflavone (3HF), using the excited-state proton-transfer (ESPT) luminescence of 3HF as a probe. Upon addition of BSA to the flavonoid solutions, we observe remarkable changes in the absorption, ESPT fluorescence emission and excitation profiles as well as anisotropy (r) values. Complexation of 3HF with protein results in a pronounced shift (20 nm) of the ESPT emission maximum of the probe (from lambda(max)(em) = 513 nm to lambda(max)(em) = 533 nm) accompanied by a significant increase in fluorescence intensity. The spectral data also suggest that, in addition to ESPT, the protein environment induces proton abstraction from 3HF leading to formation of anionic species in the ground state. Fairly high values of anisotropy are observed in the presence of BSA for the tautomer (r = 0.25) as well as anion (r = 0.35) species of 3HF, implying that both the species are located in motion restricted environments of BSA molecules. Analysis of relevant spectroscopic data leads to the conclusions that two binding sites are involved in BSA-3HF interaction, and the interaction is slightly positively cooperative in nature with a similar binding constant of 1.1 - 1.3 x 10(5) M(-1) for both these sites. Proteins 2001;43:75-81. PMID- 11276079 TI - Residue frequencies and pairing preferences at protein-protein interfaces. AB - We used a nonredundant set of 621 protein-protein interfaces of known high resolution structure to derive residue composition and residue-residue contact preferences. The residue composition at the interfaces, in entire proteins and in whole genomes correlates well, indicating the statistical strength of the data set. Differences between amino acid distributions were observed for interfaces with buried surface area of less than 1,000 A(2) versus interfaces with area of more than 5,000 A(2). Hydrophobic residues were abundant in large interfaces while polar residues were more abundant in small interfaces. The largest residue residue preferences at the interface were recorded for interactions between pairs of large hydrophobic residues, such as Trp and Leu, and the smallest preferences for pairs of small residues, such as Gly and Ala. On average, contacts between pairs of hydrophobic and polar residues were unfavorable, and the charged residues tended to pair subject to charge complementarity, in agreement with previous reports. A bootstrap procedure, lacking from previous studies, was used for error estimation. It showed that the statistical errors in the set of pairing preferences are generally small; the average standard error is approximately 0.2, i.e., about 8% of the average value of the pairwise index (2.9). However, for a few pairs (e.g., Ser-Ser and Glu-Asp) the standard error is larger in magnitude than the pairing index, which makes it impossible to tell whether contact formation is favorable or unfavorable. The results are interpreted using physicochemical factors and their implications for the energetics of complex formation and for protein docking are discussed. Proteins 2001;43:89-102. PMID- 11276078 TI - Effect of naturally occurring active site mutations on hepatitis C virus NS3 protease specificity. AB - A comparison of the DNA sequences from all available genotypes of HCV indicate that the active site residues of the NS3 protease are strictly conserved with the exception of positions 123 and 168, which border the S(4) subsite. In genotype 3, the canonic arginine and aspartic acid have been replaced with threonine and glutamine, respectively. To determine if these differences contribute to an altered specificity, we characterized single-chain NS3 proteases from strains 1a, 1b, and 3a with peptide substrates and product inhibitors on the basis of the natural cleavage junction sequences, in addition to polyprotein substrates derived from the 1a strain. No statistically significant differences in specificity were observed. To demonstrate that the active sites were actually different, we generated and evaluated peptide substrates with unnatural extended side-chains. These studies confirmed that there are measurable differences between the NS3 proteases of genotypes 1 and 3. Specifically, a 5-fold difference in K(i) was observed between the proteases from genotypes 1 and 3 when a D-Glu occupied P(5), and a 30-fold difference was seen when this position contained a D homoglutamate. The contribution of residues 123 and 168 toward the altered specificity was then evaluated individually by site-directed mutagenesis. These mutants showed that potency differences within this series could be attributed to the residue that occupied position 123 of the protease. Modeling these unnatural substrate/mutant protease interactions, on the basis of cocrystal structures of enzyme-substrate complexes, provides a structural basis for these observations. Proteins 2001;43:82-88. PMID- 11276080 TI - Clusters in alpha/beta barrel proteins: implications for protein structure, function, and folding: a graph theoretical approach. AB - The alpha/beta barrel fold is adopted by most enzymes performing a variety of catalytic reactions, but with very low sequence similarity. In order to understand the stabilizing interactions important in maintaining the alpha/beta barrel fold, we have identified residue clusters in a dataset of 36 alpha/beta barrel proteins that have less than 10% sequence identity within themselves. A graph theoretical algorithm is used to identify backbone clusters. This approach uses the global information of the nonbonded interaction in the alpha/beta barrel fold for the clustering procedure. The nonbonded interactions are represented mathematically in the form of an adjacency matrix. On diagonalizing the adjacency matrix, clusters and cluster centers are obtained from the highest eigenvalue and its corresponding vector components. Residue clusters are identified in the strand regions forming the beta barrel and are topologically conserved in all 36 proteins studied. The residues forming the cluster in each of the alpha/beta protein are also conserved among the sequences belonging to the same family. The cluster centers are found to occur in the middle of the strands or in the C terminal of the strands. In most cases, the residues forming the clusters are part of the active site or are located close to the active site. The folding nucleus of the alpha/beta fold is predicted based on hydrophobicity index evaluation of residues and identification of cluster centers. The predicted nucleation sites are found to occur mostly in the middle of the strands. Proteins 2001;43:103-112. PMID- 11276081 TI - High throughput docking for library design and library prioritization. AB - The prioritization of the screening of combinatorial libraries is an extremely important task for the rapid identification of tight binding ligands and ultimately pharmaceutical compounds. When structural information for the target is available, molecular docking is an approach that can be used for prioritization. Here, we present the initial validation of a new rapid approach to molecular docking developed for prioritizing combinatorial libraries. The algorithm is tested on 103 individual cases from the protein data bank and in nearly 90% of these cases docks the ligand to within 2.0 A of the observed binding mode. Because the mean CPU time is <5 s/mol, this approach can process hundreds of thousands of compounds per week. Furthermore, if a somewhat less thorough search is performed, the search time drops to 1 s/mol, thus allowing millions of compounds to be docked per week and tested for potential activity. Proteins 2001;43:113-124. PMID- 11276082 TI - Role of the C-terminal chain in human interferongamma stability: an electrostatic study. AB - Electrostatic interactions in two structures of human interferon gamma (hIFNgamma), corresponding to interferon molecule alone and bound to its receptor, were analyzed on the basis of a continuum dielectric model. It was found that a number of titratable groups, mainly basic, show large pK shifts and remain in their neutral forms at physiologically relevant pH. The fact that these groups are largely common to both structures and that most of them belong to the set of most conserved sites suggests that this is a property inherent to the hIFNgamma molecule rather than an artifact of the crystal packing. His111 was also found deprotonated at neutral pH. It was concluded that receptor recognition involving His111 is driven by aromatic coupling of His111 and Tyr52 from the receptor rather than by electrostatic interactions. The structure corresponding to hIFNgamma in complex with its receptor shows a reduction in number and in degree of desolvation of the buried titratable sites. This finding suggested that on receptor binding, hIFNgamma adopts energetically more favorable, relaxed, conformation. It was experimentally shown that in contrast to the full-size hIFNgamma, the construct having 21 amino acid residues deleted from the C terminus is soluble. The hydrophobicity profile analysis suggested that factors other than the exposure of hydrophobic parts of the molecule are responsible for the low stability and propensity for aggregation. On the basis of these results, it was assumed that the electrostatic influence of the C-terminal part contributes particularly to the low solvent exposure of the titratable groups, and hence to the low structural stability and propensity for aggregation of the recombinant hIFNgamma. Proteins 2001;43:125-133. PMID- 11276083 TI - Adaptations of the helix-grip fold for ligand binding and catalysis in the START domain superfamily. AB - With a protein structure comparison, an iterative database search with sequence profiles, and a multiple-alignment analysis, we show that two domains with the helix-grip fold, the star-related lipid-transfer (START) domain of the MLN64 protein and the birch allergen, are homologous. They define a large, previously underappreciated superfamily that we call the START superfamily. In addition to the classical START domains that are primarily involved in eukaryotic signaling mediated by lipid binding and the birch antigen family that consists of plant proteins implicated in stress/pathogen response, the START superfamily includes bacterial polyketide cyclases/aromatases (e.g., TcmN and WhiE VI) and two families of previously uncharacterized proteins. The identification of this domain provides a structural prediction of an important class of enzymes involved in polyketide antibiotic synthesis and allows the prediction of their active site. It is predicted that all START domains contain a similar ligand-binding pocket. Modifications of this pocket determine the ligand-binding specificity and may also be the basis for at least two distinct enzymatic activities, those of a cyclase/aromatase and an RNase. Thus, the START domain superfamily is a rare case of the adaptation of a protein fold with a conserved ligand-binding mode for both a broad variety of catalytic activities and noncatalytic regulatory functions. Proteins 2001;43:134-144. PMID- 11276084 TI - Identification of new repeating motifs in titin. AB - Repeating motifs of 26-28 amino acids have been identified in the PEVK region of the giant elastic protein titin. These motifs, termed PPAK for the four amino acids that often constitute the beginning of the motif, occur 60 times in human soleus titin. PPAK motifs occur in groups of 2-12 that are separated by regions rich in glutamic acid (approximately 45%) and termed polyE segments. The fluctuation of the net charge between the PPAK and polyE regions suggests ionic interactions between these segments and their involvement in the elastic function of titin. Proteins 2001;43:145-149. PMID- 11276085 TI - Elucidating the structural mechanisms for biological activity of the chemokine family. AB - Chemokines are a family of proteins involved in inflammatory and immune response. They share a common fold, made up of a three-stranded beta-sheet, and an overlaying alpha-helix. Chemokines are mainly categorized into two subfamilies distinguished by the presence or absence of a residue between two conserved cysteines in the N-terminus. Although dimers and higher-order quaternary structures are common in chemokines, they are known to function as monomers. Yet, there is quite a bit of controversy on how the actual function takes place. The mechanisms of binding and activation in the chemokine family are investigated using the gaussian network model of proteins, a low-resolution model that monitors the collective motions in proteins. It is particularly suitable for elucidating the global dynamic characteristics of large proteins or the common properties of a group of related proteins such as the chemokine family presently investigated. A sample of 16 proteins that belong to the CC, CXC, or CX(3)C subfamilies are inspected. Local packing density and packing order of residues are used to determine the type and range of motions on a global scale, such as those occurring between various loop regions. The 30s-loop, although not directly involved in the binding interface like the N-terminus and the N-loop, is identified as having a prominent role in both binding/activation and dimerization. Two mechanisms are distinguished based on the communication among the three flexible regions. In these two-step mechanisms, the 30s-loop assists either the N-loop or the N-terminus during binding and activation. The findings are verified by molecular mechanics and molecular dynamics simulations carried out on the detailed structure of representative proteins from each mechanism type. A basis for the construction of hybrids of chemokines to bind and/or activate various chemokine receptors is presented. Proteins 2001;43:150-160. PMID- 11276086 TI - Lattice protein folding with two and four-body statistical potentials. AB - The cooperative folding of proteins implies a description by multibody potentials. Such multibody potentials can be generalized from common two-body statistical potentials through a relation to probability distributions of residue clusters via the Boltzmann condition. In this exploratory study, we compare a four-body statistical potential, defined by the Delaunay tessellation of protein structures, to the Miyazawa-Jernigan (MJ) potential for protein structure prediction, using a lattice chain growth algorithm. We use the four-body potential as a discriminatory function for conformational ensembles generated with the MJ potential and examine performance on a set of 22 proteins of 30-76 residues in length. We find that the four-body potential yields comparable results to the two-body MJ potential, namely, an average coordinate root-mean square deviation (cRMSD) value of 8 A for the lowest energy configurations of all alpha proteins, and somewhat poorer cRMSD values for other protein classes. For both two and four-body potentials, superpositions of some predicted and native structures show a rough overall agreement. Formulating the four-body potential using larger data sets and direct, but costly, generation of conformational ensembles with multibody potentials may offer further improvements. Proteins 2001;43:161-174. PMID- 11276088 TI - Ab initio protein structure prediction using physicochemical potentials and a simplified off-lattice model. AB - This study describes a computational method for ab inito protein structure prediction. Protein conformation has been modeled by using six optimized backbone torsion angles and fixed side chains approximating rotationally averaged real side chains. The approximations aim to keep complexity of the structure description to a minimum without seriously compromising the accuracy of the structural representation. An evolutionary Monte Carlo algorithm has been developed to search through this restricted conformational space to locate low energy protein structures. A simple physicochemical force field has been developed to assess the energies of different conformations within this structural description. The corresponding residue interaction energies are based on hydrophobic, hydrophilic, steric, and hydrogen-bonding potentials. The search procedure has been used to locate native energy minima from primary sequence alone. The 3-D structures of polypeptides up to 38 residues with both beta and alpha secondary structural elements have been accurately predicted. The search procedure has been found to be highly efficient and follows an energetically and structurally plausible pathway to locate native populations. The simple force field described in the study has been compared with a more complex all-atom model and been found to be similarly effective in predicting the structures of proposed independent folding units. Proteins 2001;43:186-202. PMID- 11276087 TI - Structural basis for altered activity of M- and H-isozyme forms of human lactate dehydrogenase. AB - Lactate dehydrogenase (LDH) interconverts pyruvate and lactate with concomitant interconversion of NADH and NAD(+). Although crystal structures of a variety of LDH have previously been described, a notable absence has been any of the three known human forms of this glycolytic enzyme. We have now determined the crystal structures of two isoforms of human LDH-the M form, predominantly found in muscle; and the H form, found mainly in cardiac muscle. Both structures have been crystallized as ternary complexes in the presence of the NADH cofactor and oxamate, a substrate-like inhibitor. Although each of these isoforms has different kinetic properties, the domain structure, subunit association, and active-site regions are indistinguishable between the two structures. The pK(a) that governs the K(M) for pyruvate for the two isozymes is found to differ by about 0.94 pH units, consistent with variation in pK(a) of the active-site histidine. The close similarity of these crystal structures suggests the distinctive activity of these enzyme isoforms is likely to result directly from variation of charged surface residues peripheral to the active site, a hypothesis supported by electrostatic calculations based on each structure. Proteins 2001;43:175-185. PMID- 11276089 TI - Three-dimensional model of the SHBG-like region of anticoagulant protein S: new structure-function insights. AB - Protein S (PS) is a vitamin K-dependent glycoprotein that consists of several modules including a C-terminal sex hormone-binding globulin (SHBG)-like domain that has been subdivided into two laminin LG-type domains. The SHBG-like region of PS is known to bind to a complement regulator molecule, C4b-binding protein (C4BP), coagulation factor Va (FVa) and receptor tyrosine kinases. Inherited PS deficiency has been associated with thromboembolic disease. Yet, study of the mechanisms by which the SHBG-like region of PS serves its essential functions has so far been hampered because of the lack of structural information. Recently, the three-dimensional (3D) structure of LG domains from plasma SHBG, laminin and neurexin have been reported and were found related to the pentraxin family. We used these X-ray structures to build homology models of the SHBG-like region of human PS. We then analyzed previously reported experimental/clinical data in the light of the predicted structures. A potential calcium-binding site is found in the first LG domain of PS and D292 could play a role in this process. This region is close to the interface between the two LG domains and is also surrounded by segments that have been suggested by synthetic peptide studies to be important for C4BP or FVa binding. The 39 point mutations linked to PS deficiencies or reported as neutral variants were rationalized in the 3D structure. Proteins 2001;43:203-216. PMID- 11276091 TI - Transcriptional activity of the TFIIA four-helix bundle in vivo. AB - TFIIA contributes to transcription initiation by stabilizing the TBP-TATA interaction and by mediating the response to transcriptional activators and inhibitors. TFIIA contains a six-stranded beta-sheet domain and a four-helix bundle. The beta-domain makes functional contacts with DNA and TBP. The role of the four-helix bundle was investigated using a structure-based model of this domain (called 4HB). 4HB adopts a highly stable, helical fold, consistent with its structure in the context of TFIIA. Like TBP and other intact transcription factors, 4HB is able to activate transcription in vivo when artificially recruited to a promoter via a heterologous DNA-binding domain. Thus, in addition to making important contacts with TBP and DNA via the beta-domain, TFIIA makes other specific, functional contacts with the transcriptional machinery via the four-helix bundle. Proteins 2001;43:227-232. PMID- 11276090 TI - Ligand-protein inverse docking and its potential use in the computer search of protein targets of a small molecule. AB - Ligand-protein docking has been developed and used in facilitating new drug discoveries. In this approach, docking single or multiple small molecules to a receptor site is attempted to find putative ligands. A number of studies have shown that docking algorithms are capable of finding ligands and binding conformations at a receptor site close to experimentally determined structures. These algorithms are expected to be equally applicable to the identification of multiple proteins to which a small molecule can bind or weakly bind. We introduce a ligand-protein inverse-docking approach for finding potential protein targets of a small molecule by the computer-automated docking search of a protein cavity database. This database is developed from protein structures in the Protein Data Bank (PDB). Docking is conducted with a procedure involving multiple-conformer shape-matching alignment of a molecule to a cavity followed by molecular mechanics torsion optimization and energy minimization on both the molecule and the protein residues at the binding region. Scoring is conducted by the evaluation of molecular-mechanics energy and, when applicable, by the further analysis of binding competitiveness against other ligands that bind to the same receptor site in at least one PDB entry. Testing results on two therapeutic agents, 4H-tamoxifen and vitamin E, showed that 50% of the computer-identified potential protein targets were implicated or confirmed by experiments. The application of this approach may facilitate the prediction of unknown and secondary therapeutic target proteins and those related to the side effects and toxicity of a drug or drug candidate. Proteins 2001;43:217-226. PMID- 11276093 TI - Implications of bulk motion for diffusion-weighted imaging experiments: effects, mechanisms, and solutions. AB - This review article describes the effect of bulk motion on diffusion-weighted imaging experiments, and examines methods for correcting the resulting artifacts. The emphasis throughout the article is on two-dimensional imaging of the brain. The effects of translational and rotational motion on the MR signal are described, and the literature concerning pulsatile brain motion is examined. Methods for ameliorating motion effects are divided into three generic categories. The first is methods that should be intrinsically insensitive to macroscopic motion. These include motion-compensated diffusion-weighting schemes, single-shot EPI, projection reconstruction, and line scanning. Of these, only single-shot EPI and projection reconstruction methods can obtain high-quality images without compromising on sensitivity. The second category of methods is those that can be made insensitive to bulk motion. The methods examined here are FLASH and RARE. It is shown that for both sequences motion insensitivity is in general attained only at the cost of a 50% reduction in sensitivity. The final set of methods examined are those that correct for motion, primarily navigator echoes. The properties and limitations of the navigator echo approach are presented, as are those of methods which attempt to correct the acquired data by minimizing image artifacts. The review concludes with a short summary in which the current status of diffusion imaging in the presence of bulk motion is examined. PMID- 11276094 TI - Methodology of brain perfusion imaging. AB - Numerous techniques have been proposed in the last 15 years to measure various perfusion-related parameters in the brain. In particular, two approaches have proven extremely successful: injection of paramagnetic contrast agents for measuring cerebral blood volumes (CBV) and arterial spin labeling (ASL) for measuring cerebral blood flows (CBF). This review presents the methodology of the different magnetic resonance imaging (MRI) techniques in use for CBV and CBF measurements and briefly discusses their limitations and potentials. PMID- 11276095 TI - Advances in human cardiac 31P-MR spectroscopy: SLOOP and clinical applications. AB - Phosphorus magnetic resonance spectroscopy (31P-MRS) has revealed a lot about the biochemistry of physiological and pathological processes in the heart. Nevertheless, until today, cardiac 31P-MRS has not had any clinical impact, albeit some pioneering studies demonstrated that 31P-MRS can indeed provide diagnostic information. In this paper, the development of techniques for human cardiac 31P-MRS over the past decade is reviewed, and the requirements for a reliable clinical measurement protocol are discussed. Spatial localization with optimal pointspread function (SLOOP) is a new method to achieve spatial localization and absolute quantitation. Its properties are detailed, and preliminary findings in patients with dilated cardiomyopathy or myocardial infarction are presented. PMID- 11276096 TI - MRI in France: the French paradox. AB - Although France is a modern, developed country, which spends nearly 10% of the gross national product on healthcare and has a highly praised level of medicine, the number of modern imaging scanners, such as CT (595), MRI (182), and PET (5), is quite low when compared to other European countries. Politics and a long standing tradition of centralization are prominent among reasons for such an underdevelopment. This situation has resulted in another French paradox not linked to wine consumption. The French life expectancy is very high, but the number of imaging equipment is very low. PMID- 11276097 TI - Diffusion tensor imaging: concepts and applications. AB - The success of diffusion magnetic resonance imaging (MRI) is deeply rooted in the powerful concept that during their random, diffusion-driven displacements molecules probe tissue structure at a microscopic scale well beyond the usual image resolution. As diffusion is truly a three-dimensional process, molecular mobility in tissues may be anisotropic, as in brain white matter. With diffusion tensor imaging (DTI), diffusion anisotropy effects can be fully extracted, characterized, and exploited, providing even more exquisite details on tissue microstructure. The most advanced application is certainly that of fiber tracking in the brain, which, in combination with functional MRI, might open a window on the important issue of connectivity. DTI has also been used to demonstrate subtle abnormalities in a variety of diseases (including stroke, multiple sclerosis, dyslexia, and schizophrenia) and is currently becoming part of many routine clinical protocols. The aim of this article is to review the concepts behind DTI and to present potential applications. PMID- 11276098 TI - A fast FLAIR dual-echo technique for hippocampal T2 relaxometry: first experiences in patients with temporal lobe epilepsy. AB - To evaluate the use of cerebrospinal fluid (CSF) signal nulling in MR T2 measurements of the hippocampus in normal control subjects and patients with temporal lobe epilepsy (TLE), dual-echo acquisitions covering the whole brain were used. T2 relaxation times were estimated in 12 standard Eurospin II MR test objects and in the hippocampi of 10 control subjects, using T2 maps constructed from conventional spin-echo (CSE), fast spin-echo (FSE), and fast FLAIR (FF) dual echo sequences on a 1.5-T MR scanner. Hippocampal T2 values (HCT2) were measured on contiguous coronal 5-mm slices throughout the antero-posterior extent of each hippocampus in the 10 controls and 12 TLE patients, using both CSE and FF. Scan rescan reproducibility in Eurospin II standard MR test objects was high for all sequences. There was a good correlation between T2 values from CSE, FF, and FSE sequences in test objects and in control hippocampi. In controls, the coefficient of variation of mean HCT2 values differed between slice positions, but was lowest for FF, followed by CSE data. The intrarater coefficient of reliability between repeated measurements in control subjects was lowest for FF HCT2, at 2.3%. The interrater coefficient of reliability for CSE HCT2 measurements in controls (4.8%) was slightly lower than the interrater coefficient for FF HCT2 (5.4%). HCT2 measurement with both CSE and FF identified abnormal values in the same 10 hippocampi of 12 patients. Hippocampal dual-echo T2 relaxometry using CSF nulling is reliable in control subjects, and identifies the abnormal hippocampi in patients with TLE. The increases in hippocampal T2 signal demonstrated using FF HCT2 measurements are unlikely to be partial volume effects from CSF. PMID- 11276099 TI - Proton magnetic resonance spectroscopic imaging reveals differences in spinocerebellar ataxia types 2 and 6. AB - The objective of this study was to investigate cerebellar metabolism in patients with autosomal dominant cerebellar ataxia type 1 (ADCA-I) carrying two distinct mutations of spinocerebellar ataxia (SCA). Non-invasive image-guided proton magnetic resonance spectroscopy imaging (1H-MRSI) was performed in 4 patients with SCA2, and 3 patients carrying the SCA6 mutation. For MRSI, we employed a spin-echo sequence (TR = 1500 msec, TE = 135 msec, slice thickness = 15 mm, FOV = 240 mm) and a stimulated-echo sequence (TR = 1500 msec, TE = 20 msec, slice thickness = 15 mm, FOV = 240 mm). Measures included the peak integral ratios of neuronal and glial markers [N-acetylaspartate (NA) to creatine (Cr), choline containing compounds (CHO) to Cr, and lactate (LAC) to Cr]. We found NA:Cr ratios were significantly lower in patients with SCA2 (40.4% lower) compared to patients carrying the SCA6 mutation. CHO:Cr ratios differed between the two mutations using short echo time (30.8% lower in SCA2), but not when applying long echo time 1H-MRSI. Measurements using long echo time revealed LAC peaks in all SCA2 patients. 1H-MRSI revealed metabolic differences between SCA2 and SCA6 patients. NA:Cr ratios were significantly lower in patients with the SCA2 mutation compared to the SCA6 mutation, and LAC signals were obtained in the cerebella of SCA2 patients. In addition, CHO:Cr ratios showed different behavior using short and long TE, indicating differences in relaxation times of choline compounds in SCA2. PMID- 11276100 TI - Proton MR spectroscopy in clinical routine. AB - In vivo magnetic resonance spectroscopy (MRS) addresses metabolic pathways and their steady states in different tissue types. The brain has by tradition, and due to technical limitations in other organs, been one of the tissues most studied by MRS, and both 1H- and 31P-MRS have been used. Although 31P-MRS is outstanding for the evaluation of sources of metabolic energy in the brain, 1H MRS has become the major clinically applied method in neurospectroscopy, as it provides information on markers of neuronal function, myelin, cell membranes, and metabolic active compounds. Furthermore, MR sensitivity is much greater for protons than it is for phosphorus and 1H-MRS, therefore allowing better spatial resolution. This review focuses on neurospectroscopy and diagnostic insights into diverse neurological problems provided by 1H-MRS applied as a clinical tool. PMID- 11276101 TI - Arterial first pass gadolinium-CM dynamics as a function of several intravenous saline flush and Gd volumes. AB - This study was performed to evaluate the dynamics of an arterial first pass gadolinium (Gd) contrast medium (CM) bolus at the descending aorta (DAo), depending on various saline flush and Gd volumes. Using an ultra-fast two- dimensional GE-sequence (Siemens Vision, 1.5-T), 200 sequential cross-sectional images of the addressed vessel (1 slice/s) were obtained. Several saline flush volumes (15 mL, 30 mL, and 60 mL) were applied following the administration of 10 mL Gd (single dose) to a group of 4 normal volunteers (body weight 50-55 kg) using a mechanical MR injector (injection rate = 3.0 mL/s). Additionally, when performing a second test series, the saline volume remained constant, while the Gd volumes were varied from half doses to triple doses (5, 10, 20, and 30 mL Gd were given to every volunteer of the group). The signal intensity versus time (SI/T) curve at a measured region of interest (ROI) within the DAo was calculated. The bolus arrival time (BAT), the maximal signal-to-noise ratio (SNR(max)), the bolus time length (BL; 75% and 80% maximum intensity duration), the slope of the SI/T curve, and the areas below the SI/T curve for both the 80% and 75% maximum intensity duration level (INT(80%) and INT(75%)) were calculated. The increase of saline flush volume from 30 to 60 mL caused significant bolus lengthening of approximately 50% (mean BL = 9.5 s, 10.3 s, and 15.4 s for 15 mL, 30 mL, and 60 mL saline flush volumes, respectively, measured as SI/T duration at the 75% SNR(max) level). Using saline flush volumes equal to or higher than 30 mL increased the slope of the SI/T curve. A continuous increase of INT(75%/80%) by using higher saline flush volumes was found. Different saline and Gd volumes did not affect the SNR(max) and the BAT. Only the low dose (0.05 mmol/kg Gd) showed a 17%-21.6% significantly lower SNR(max). The BL and the INT increased mainly by enlarging of applied Gd volume from single to double dose (BL(75%) and INT(75%) were 9.6 s and 1305, 12.3 s and 2121, 38.5 s and 6181, 37.8 s and 6613 for 5, 10, 20, and 30 mL applied Gd volume, respectively). The arterial bolus length benefits from increasing Gd and saline flush volumes due to increased venous bolus length and wash out effects of Gd within the injection site of the vein. Doses larger than a single dose are not needed to increase the SNR in contrast enhanced magnetic resonance angiography images of the thoracic aorta. PMID- 11276102 TI - Analysis of contrast-enhanced dynamic MR images of the lung. AB - Recent studies have demonstrated the potential of dynamic contrast-enhanced magnetic resonance imaging (MRI) describing pulmonary perfusion. However, breathing motion, susceptibility artifacts, and a low signal-to-noise ratio (SNR) make automatic pixel-by-pixel analysis difficult. In the present work, we propose a novel method to compensate for breathing motion. In order to test the feasibility of this method, we enrolled 53 patients with pulmonary embolism (N = 24), chronic obstructive pulmonary disease (COPD) (N = 14), and acute pneumonia (N = 15). A crucial part of the method, an automatic diaphragm detection algorithm, was evaluated in all 53 patients by two independent observers. The accuracy of the method to detect the diaphragm showed a success rate of 92%. Furthermore, a Bayesian noise reduction technique was implemented and tested. This technique significantly reduced the noise level without removing important clinical information. In conclusion, the combination of a motion correction method and a Bayesian noise reduction method offered a rapid, semiautomatic pixel by-pixel analysis of the lungs with great potential for research and clinical use. PMID- 11276103 TI - Slow clearance gadolinium-based extracellular and intravascular contrast media for three-dimensional MR angiography. AB - The objective of this study was to assess two new slow-clearance contrast media with extracellular and intravascular distribution for magnetic resonance angiography (MRA). Extracellular Gd-DTPA-BC(2)glucA and intravascular Gd(DO3A)(3) lys(16) were developed within the European Biomed2 MACE Program and compared with two reference compounds, intravascular CMD-A2-Gd-DOTA and extracellular GdDOTA, in 12 rats. Pre- and post-contrast three-dimensional MR (TR/TE = 5 msec/2.2 msec; isotropic voxel size 0.86 mm(3)) was acquired for 2 hours. Signal-to-noise enhancement (DeltaSNR) was calculated. Two minutes after injection, all contrast media provided strong vascular signal enhancement. The DeltaSNR for Gd-DTPA BC(2)glucA, Gd(DO3A)(3)-lys(16), CMD-A2-Gd-DOTA, and GdDOTA were 13.0 +/- 1.8, 25.0 +/- 3.2, 25.0 +/- 4.0, and 18.0 +/- 3.4, respectively. Gd-DTPA-BC(2)glucA, Gd(DO3A)(3)-lys(16), and CMD-A2-Gd-DOTA cleared slowly from the circulation, whereas GdDOTA cleared rapidly. Vascular DeltaSNR at 2 hours were 2.9 +/- 0.6, 25.0 +/- 3.2, 25.0 +/- 4.0, and 0.4 +/- 1.0. Gd(DO3A)(3)-lys(16) provided strong vascular and minor background enhancement, and thus may be useful for MRA or perfusion imaging. Gd-DTPA-BC(2)glucA produces persistent enhancement of extracellular water, and thus may allow quantification of extracellular distribution volume and assessment of myocardial viability. PMID- 11276105 TI - Post-radiotherapy contrast enhancement changes in fast dynamic MRI of cervical carcinoma. AB - This pilot study determines fast dynamic gadolinium enhanced MRI contrast enhancement parameters (onset of enhancement and time to peak enhancement) before and after radiotherapy in 10 cervical carcinoma patients. Before radiotherapy, onset of enhancement and time to peak enhancement were early, with a median of 4.5 and 5.2 seconds, respectively. High-grade tumors showed early enhancement, compared with low-grade. After radiotherapy, contrast enhancement patterns differed. In survivors, onset of enhancement after radiotherapy was later than before radiotherapy. In non-survivors, onset of enhancement after radiotherapy was still early. The median difference in onset of enhancement before and after radiotherapy in survivors and non-survivors was an increase of 3.2 and a decrease of 1.1 seconds, respectively. Early onset of enhancement after radiotherapy was a better predictor for survival than a high-signal intensity zone on post radiotherapy unenhanced T1/T2-weighted MRI. It is concluded that enhancement parameters from fast dynamic Gd-enhanced MR images can provide additional functional information with regard to tumor vascularization, and may have prognostic significance. It complements clinical examination and unenhanced MRI in determining the effectiveness of radiotherapy treatment in cervical carcinoma. Future studies will focus on the clinical utility and improvements of the estimation of contrast-enhanced parameters with this new technique. PMID- 11276104 TI - Biodistribution of ultrasmall iron oxide particles in the rat liver. AB - Ferumoxtran, an ultrasmall superparamagnetic iron oxide particle, can be located in several tissue compartments in the liver, namely the extracellular space (blood and interstitium), reticuloendothelial cells, and possibly hepatocytes. To better understand the compartmental distribution of ferumoxtran in the liver, we performed a longitudinal study in the rat using microscopy and magnetic resonance imaging. At light microscopy, no substantial cellular uptake of ferumoxtran was observed before one hour after injection. With a dose of 15 micromol Fe/kg, the number of ferumoxtran particles in the reticuloendothelial cells peaked between one and four hours and with a 150 micromol Fe/kg dose, it peaked between eight and 24 hours. Within hepatocytes, only sparse particles were observed with electron microscopy, at a dose of 150 micromol Fe/kg. Imaging performed up until one hour after ferumoxtran injection showed a significant increase in liver signal intensity on T1-weighted images. These results suggest that ferumoxtran mainly acts as an extracellular agent for at least one hour in the rat and that reticuloendothelial accumulation peaks at later time points. Substantial uptake within hepatocytes did not occur. PMID- 11276106 TI - Accurate estimation of pharmacokinetic contrast-enhanced dynamic MRI parameters of the prostate. AB - Quantitative analysis of contrast-enhanced dynamic MR images has potential for diagnosing prostate cancer. Contemporary fast acquisition techniques can give sufficiently high temporal resolution to sample the fast dynamics observed in the prostate. Data reduction for parametric visualization requires automatic curve fitting to a pharmacokinetic model, which to date has been performed using least squares error minimization methods. We observed that these methods often produce unexpectedly noisy estimates, especially for the typically fast, intermediate parameters time-to-peak and start-of-enhancement, resulting in inaccurate pharmacokinetic parameter estimates. We developed a new curve fit method that focuses on the most probable slope. A set of 10 patients annotated using histopathology was used to compare the conventional and new methods. The results show that our new method is significantly more accurate, especially in the relatively less-enhancing peripheral zone. We conclude that estimation accuracy depends on the curve fit method, which is especially important when evaluating the peripheral zone of the prostate. PMID- 11276107 TI - A targeted contrast agent for magnetic resonance imaging of thrombus: implications of spatial resolution. AB - A preparation of ultra-small superparamagnetic iron oxide (USPIO) particles coupled to an RGD peptide (RGD-USPIO) was investigated as an MR contrast agent, targeted to activated platelets, in both ex vivo and in vivo thrombus models. Thrombus visualization ex vivo was compared using RGD-USPIO and a non-targeted UPSIO. The influence of thrombus visualization on thrombus exposure time to RGD USPIO (ex vivo) and on the spatial resolution of the MR image (ex vivo and in vivo) was assessed. RGD-USPIO resulted in better thrombus visualization than non targeted USPIO ex vivo, and maximum enhancement was achieved after approximately one hour exposure time of the thrombus to RGD-USPIO. The ability to visualize the clots was highly dependent on the spatial resolution of the image. In vivo, an in plane resolution of less than 0.2 x 0.2 mm(2) was required for good clot visualization after contrast enhancement. It is concluded that the achievable resolution and sensitivity is a potential limitation to the usefulness of active vascular targeting in MRI. PMID- 11276109 TI - Investigation of the factors responsible for burns during MRI. AB - Numerous reported burn injuries have been sustained during clinical MRI procedures. The aim of this study was to investigate the possible factors that may be responsible for such burns. Experiments were performed to investigate three possible mechanisms for causing heating in copper wire during MRI: direct electromagnetic induction in a conductive loop, induction in a resonant conducting loop, and electric field resonant coupling with a wire (the antenna effect). Maximum recorded temperature rises were 0.6 degrees C for the loop, 61.1 degrees C for the resonant loop, and 63.5 degrees C for the resonant antenna. These experimental findings suggest that, contrary to common belief, it is unlikely that direct induction in a conductive loop will result in thermal injury. Burn incidents are more likely to occur due to the formation of resonant conducting loops and from extended wires forming resonant antenna. The characteristics of resonance should be considered when formulating safety guidelines. PMID- 11276108 TI - Ultrasmall particulate iron oxides as contrast agents for magnetic resonance spectroscopy: a dose-effect study. AB - Long-distance effects of a superparamagnetic contrast agent (AMI227) were investigated by phosphorus-31 NMR spectroscopy at 7.05 Tesla. In an initial methodological approach, the effects observed on phantoms were compared to the results of theoretical calculations. In a second step, the particles were administered to excised and perfused rat livers (N = 5) and hearts (N = 5) through the perfusion medium for 12 minutes at various concentrations (0.9, 1.8, and 3.6 mM Fe). Organs were subsequently rinsed with the perfusion medium for 42 minutes. During particle perfusion, the spectral lines were shifted and exhibited a strong broadening, although the peak area remained constant, testifying to the inocuity of the material. For hearts only, these disturbances disappeared upon organ rinsing. These through-space susceptibility effects of the particles located in the vessels on phosphorus nuclei, which are strictly confined to the intracellular space, show that high-susceptibility intravascular agents could be useful to evaluate tissue perfusion by contrast-enhanced spectroscopy. PMID- 11276110 TI - Safety aspects and quality assessment in MRI and MRS: a challenge for health care systems in Europe. AB - Despite advances in international cooperation, the application of the safety regulations of different countries remains an important challenge for manufacturers and health care workers in the European Union. Rapid technological development during the last 20 years, and the still controversial nature of certain potential effects of magnetic or electromagnetic fields, make the task particularly difficult for MRI and MRS. As the relevant literature is rather extensive, the present work will limit its goals to four questions: 1) How is MRI and MRS safety regulation managed within the European Union? 2) Concerning direct potential physiological effects, what is presently well known and controlled, and, conversely, what are the remaining open (and often controversial) questions? 3) As metallic implants are probably the main risk in routine MRI, what regulatory strategy is in progress in Europe? 4) As indirect risks related to artifacts must not be underestimated, what European programs have been developed for quality assessment in MRI and MRS? In all of these fields, evidence is provided demonstrating the need for a mutual recognition of common standards for the European Union and the United States. PMID- 11276111 TI - The introduction of MR in the Nordic countries with special reference to Norway: central control versus local initiatives. AB - The introduction of MR technology in Norway differed from that in Denmark, Finland and Sweden. In the latter countries, decisions were made at county or local level while in Norway the process was steered by the government and the national health authorities. For several reasons the steering was not very successful, and the intention of buying one MR unit ended with the purchase of five units. As a counter-reaction, for seven years only university hospitals were allowed to purchase MR equipment. In 1993, the strict regulations were abolished, and during the succeeding years all four Nordic countries experienced similar exponential growth in the number of MR units. PMID- 11276112 TI - Development of clinical MR imaging in Russia and other CIS countries. AB - This paper summarizes the development of magnetic resonance imaging (MRI) in the health systems of the USSR and later in Russia and the neighboring countries. Nuclear magnetic resonance (NMR) has been used as a scientific tool in the USSR since the 1940s. In 1982 the first whole-body MR imager was installed. By the end of 1999, 131 MR systems were operating in Russia. There are substantial regional differences in the number of units installed. Russia has a domestic production of low-field MR units. There are also plans for domestic production of MR contrast agents. Other former USSR republics have much less MR equipment. Most of the MRI centers experience difficulties in maintaining equipment in working condition. Some positive changes in the national health service systems give reason to believe in an increase in the quantity of MR units and the quality of their clinical use. PMID- 11276113 TI - Automated registration of dynamic MR images for the quantification of myocardial perfusion. AB - Cardiac dynamic magnetic resonance imaging (MRI) after contrast media injection suffers from motion induced by free breathing during acquisition. This work presents an automated approach for motion correction of the heart. The registration is based on the multipass/multiresolution iterative minimizing of intrinsic differences between each image and a reference image coupled to a two dimensional/3 parameters rigid body correction. The efficiency of this correction method was evaluated with anatomical landmarks, various cost functions, and for a compartment model fit of the data with 2 parameters: K1, the blood to myocardium transfer coefficient; and Vd, the distribution volume of the contrast media. The variability of K1 and Vd, derived from the fit of the registered images (using the manual correction as a gold standard), was significantly reduced by comparison with the variability obtained from the uncorrected images (P < 0.04). This motion correction method also clearly improves the analysis of dynamic cardiac MRI after contrast media injection in comparison to manual correction. PMID- 11276115 TI - Organization and functional roles of the cytoskeleton in oligodendrocytes. AB - Mature oligodendrocytes are characterized by their numerous cytoplasmic extensions and flat membranous sheets. These sheets contain an extensive cytoskeletal network of microtubules (MTs) that maintain the cellular morphology, are specifically important for cellular sorting, and provide the rails for organelle trafficking. Mitochondria are localized in the primary and secondary processes and follow the tracks of the MTs in the cytoplasmic extensions. Oligodendrocytes express microtubule associated proteins (MAPs), specifically MAP2 and tau, which might be involved in the regulation and stabilization of the dynamic MT network in the myelin-containing cellular processes. Tau and MAP2 heterogeneity increases during oligodendroglia maturation, and in mature oligodendrocytes tau mRNA with four MT binding domains are more prominent than in progenitor cells. Filamentous cell inclusions are a unifying mechanism underlying a variety of late-onset neurodegenerative disorders and have mainly been viewed as neuron-specific. Recent evidence indicated that glial changes occur in CNS degenerative diseases and seem to be a more common feature than previously thought. Glial fibrillary tangles (GFTs) in oligodendrocytes were observed in familial multiple system tauopathy, and glial cytoplasmic inclusions (GCIs) and oligodendroglia degeneration are the histological hallmark of multiple system atrophy (MSA). GCIs are associated with MTs and contain stress proteins and MAPs. Thus, neurons and glial cells share common cytoskeletal pathologies. During health and disease, MAPs might be important regulators of the structural stability and plasticity of the oligodendroglia cytoskeleton. PMID- 11276116 TI - Structure, heterologous expression, and adhesive properties of the P(0)-like myelin glycoprotein IP1 of trout CNS. AB - The IP1 protein of trout CNS myelin as well as an IP1/P(0) chimeric protein were stably expressed in CHO cells. Successful targeting of the recombinant proteins to the membrane surface was verified by immunofluorescence staining. Full-length expression of IP1 could be confirmed by Western blot analysis of proteins extracted from stably transfected CHO-cells. The adhesive properties of IP1 were studied by an in vitro aggregation assay in which microscopic examination was combined with electronic particle counting. While IP1 conveyed only a weak increase in cell aggregation of transfected CHO cells, the IP1/P0 chimera was much more effective. In the presence of specific antibodies, cell aggregation was strongly reduced. The adhesive properties of P(0)-like proteins are discussed considering recent crystallographic data on the atomic structure of the extracellular domain of mammalian P(0). PMID- 11276117 TI - Characterization of GFP-MAL expression and incorporation in rafts. AB - During myelin formation, membrane-associated proteins have to be sorted and transported in specified membrane regions such as compact and non-compact myelin membranes. One protein that may be involved in such a process is the Myelin and Lymphocyte protein MAL (VIP17/ MVP17). MAL was identified as a novel myelin membrane component expressed by oligodendrocytes and Schwann cells. Since MAL has been shown to be important in the apical sorting machinery of polarized cells, we have started to investigate the possible functional role of MAL in sorting myelin membrane-associated molecules. In this study, we have generated cDNA constructs with green fluorescent protein (GFP) either at the N- or C-terminus of MAL. Transfection experiments showed that GFP-MAL expression resembles that of normal MAL, whereas the MAL-GFP fusion construct was not properly transported within the cell. Furthermore, we could demonstrate that GFP-MAL is enriched in detergent insoluble glycolipid-enriched microdomains as already seen for untagged MAL. As a prerequisite for the generation of transgenic mice expressing GFP-MAL under the control of its own regulatory elements, we have generated a cDNA construct with an 8-kb MAL promotor fragment fused to GFP-MAL. Transfection experiments of the Oli-neu oligodendrocyte cell line showed that GFP-MAL was expressed, but only in cells, which were stimulated for differentiation with cAMP. In summary, the results confirm that the fusion protein GFP-MAL is incorporated into detergent insoluble complexes and the 8-kb MAL promotor fragment is sufficient to be activated in oligodendrocytes. PMID- 11276118 TI - Membrane traffic in myelinating oligodendrocytes. AB - In the central nervous system (CNS), the myelin sheath is synthesised by oligodendrocytes as a specialised subdomain of an extended plasma membrane, reminiscent of the segregated membrane domains of polarised cells. Myelination takes place within a relatively short period of time and oligodendrocytes must have adapted membrane sorting and transport mechanisms to achieve such a high rate of myelin synthesis and to maintain the unique organisation of the myelin membrane. In adult life, maintenance of the functional myelin sheath requires a carefully orchestrated balance of myelin synthesis and turnover. Imbalance in these processes may cause dys- or demyelination and disease. This review summarises what is currently known about myelin protein trafficking and mistrafficking in oligodendrocytes. We also present data demonstrating distinct transport pathways for myelin structural proteins and the expression of SNARE proteins in differentiating oligodendrocytes. Myelinating glial cells may well serve as a model system for studying general aspects of membrane trafficking and organisation of membrane domains. PMID- 11276119 TI - Calcium signalling in cells of oligodendroglial lineage. AB - Intracellular Ca2+ is the key signal that regulates the efficacy of neurotransmitter release and synaptic plasticity in neurons but is also an important second messenger involved in the signal transduction and modulation of gene expression in both excitable and non-excitable cells. Glial cells, including cells of oligodendroglial (OLG) lineage, are capable of responding to extracellular stimuli via changes in the intracellular Ca2+. This review article focuses on the mechanisms of Ca2+ signalling in cells of OLG lineage with the goal of providing the basis for understanding the relevance of receptor- and non receptor-mediated signalling to oligodendroglial development, myelination, and demyelination. Conclusions to date indicate that cells of OLG lineage exhibit remarkable plasticity with regard to the expression of ion channels and receptors linked to Ca2+ signalling and that perturbation of [Ca2](i) homeostasis contributes to the pathogenesis of demyelinating diseases. PMID- 11276120 TI - Protein kinase C and mitogen-activated protein kinase signalling in oligodendrocytes. AB - Oligodendrocytes (OL) play a significant physiological role in the central nervous system by creating the myelin sheath that allows for the efficient conduction of nerve impulses. Therefore, it is important to understand which signalling cascades define the proliferation, differentiation, survival, and myelin formation potential of these cells. Currently, much of the knowledge in this field has focused on two sets of protein kinase signalling molecules: Protein kinase C (PKC) and the mitogen-activated protein kinases (MAPKs). The roles of these kinases in OL are complex, and appear to be highly dependent on the developmental stage of the OL. Even so, some broad conclusions can be drawn from the multitude of experiments conducted on the roles of PKC and MAPKs in OL. For instance, PKC appears to have a proliferative effect on immature OL, while at the same time having an inhibitory effect on OL differentiation. In mature OL, the effects of PKC include increased process extension and myelin formation. The extracellular signal-regulated (ERK) members of the MAPK family also appear to increase process extensions in mature OL. On the other hand, the c-Jun N-terminal kinase (JNK) and p38 kinase members of the MAPK family appear to regulate apoptotic events in OL. PMID- 11276121 TI - Differential response of mature TrkA/p75(NTR) expressing human and pig oligodendrocytes: aging, does it matter? AB - A differential morphological response of mature oligodendrocytes (OL) isolated from human and pig brains to the phorbol ester 12-O-tetradecanoylphorbol-13 acetate (TPA) and to the nerve growth factor (NGF) was observed. In both cases, OL regenerate their processes; however, the rate and the extension of the process formation of human OL were behind that of pig OL. Presumably, the advanced age of the human tissue in these experiments might have contributed to this decrease in process formation, an effect that was already observed for rat OL [Yong et al. (1991) J Neurosci Res 29:87-99]. The less effectivity of NGF via TrkA, which was immunocytochemically shown in human OL, and of TPA via the protein kinase C (PKC) pathway, may have its common focus on the mitogen-activated protein kinase (MAPK) cascade. In this context, it was noted that only a few studies on aging of mature OL are available. It is conceivable that age-related changes in the properties of OL could be an important factor for their cellular responsiveness during longer lasting demyelinating diseases such as multiple sclerosis. Hence, this review would like to provide a basis for future investigations on the aging of mature OL. The data presently available suggest a preliminary classification of mature OL into three categories. PMID- 11276122 TI - Molecular pathways of oligodendrocyte apoptosis revealed by mutations in the proteolipid protein gene. AB - A decade after the genetic link was established between mutations in the proteolipid protein gene and two leukodystrophies, Pelizaeus-Merzbacher disease and spastic paraplegia, the molecular mechanisms underlying pathogenesis are beginning to come to light. Data from animal models of these diseases suggest that the absence of proteolipid protein gene products in the central nervous system confers a relatively mild phenotype while missense mutations in and duplications of this gene give rise to mild or severe forms of disease. Previously, we have interpreted the disease process in terms of the accumulation of the mutant proteins in the secretory pathway and, herein, we review the evidence in favor of such a cellular mechanism. Furthermore, on the basis of recent data we suggest that the unfolded protein response may be involved in the pathogenesis of Pelizaeus-Merzbacher disease and spastic paraplegia through a kinase signaling cascade that links the accumulation of mutant proteins in the endoplasmic reticulum of oligodendrocytes with changes in gene regulation, protein synthesis, and possibly apoptosis. PMID- 11276123 TI - 4-Hydroxynonenal, a lipid peroxidation byproduct of spinal cord injury, is cytotoxic for oligodendrocyte progenitors and inhibits their responsiveness to PDGF. AB - Oligodendroglial reactions to compression injury of spinal cord include apoptosis, secondary demyelination, and remyelination failure. Within hours after contusion, the membrane lipid peroxidation (MLP) byproduct, 4-hydroxynonenal (HNE), increases rapidly in gray matter and thereafter in white matter tracts beyond the initial lesion level. Considering that HNE is a mediator and marker of neuronal MLP toxicity in various neurodegenerative conditions, the present study examined its effect on the regeneration potential of oligodendrocyte progenitors, as defined by their capacity to survive, proliferate and migrate in primary culture. Treatment of oligodendroblasts with HNE evoked a time- and dose dependent cytotoxicity resembling apoptosis at aldehyde concentrations known to be produced by neurons and achieved in tissue undergoing peroxidative injury. In addition, sublethal concentrations of HNE inhibited the mitogenic and chemotactic responses of more immature progenitors to platelet-derived growth factor. These effects appear to be mediated in part by the formation of HNE adducts with progenitor proteins located within the plasma membrane and cytoplasmic compartments. Our data are the first to show that HNE can have direct, deleterious effects on oligodendrocyte precursors. The present study also suggests a mechanism by which the striking accumulation of HNE in white matter tracts surrounding the site of spinal cord compression injury and in other ischemic-hypoxic insults associated with MLP could suppress the potential regenerative response of endogenous oligodendrocyte progenitor cells. PMID- 11276124 TI - In vitro injury model for oligodendrocytes: development, injury, and recovery. AB - In this study we investigated the effects of severe hypothermia (cryoinjury) on oligodendrocyte (OL) cell marker expression and morphological features. We used a chemically defined cell culture medium, glial development medium (GDM), which favored the optimal expression of the OL phenotype in CG4 cells. Experiments using CG4 cells cultured in 2% serum or in GDM were conducted in parallel. After severe hypothermia, cells were reanimated at 37 degrees C and 4.5% CO(2) and cultured in either GDM or in medium supplemented with 2% serum. In either medium, around 70% of the total number of cells detached within 2 to 4 hours following reanimation. Oligodendroglial markers such as A2B5, O4, Tf, ferritin, tubulin, and MBP were examined by double and triple immunofluorescence. All of these markers except MBP re-appeared at different times during the recovery period for up to 48 hours. Glial fibrillary acidic protein (GFAP) and heat shock protein 60 (HSP-60) were used as injury markers. The presence of serum induced HSP-60 expression, while GDM did not. All CG4 cells expressed HSP-60 in response to hypothermia independently of the cell culture medium used. Cryoinjury induced a spectrum of morphological changes in CG4 cells. The expression of OL specific markers was also influenced by hypothermia. Moreover both, serum and cryoinjury induced the expression of HSP-60 that colocalized with OL and myelin markers. The expression of GFAP by injured cells but not by normal cells corroborated the state of injury of CG4 cells. PMID- 11276125 TI - Axonal degeneration is an early pathological feature in autoimmune-mediated demyelination in mice. AB - Multiple sclerosis (MS) is an inflammatory disorder of the central nervous system (CNS), characterised by focal destruction of myelin. Although it is evident that the immune system contributes to tissue destruction in MS, it is still unclear as to whether this immune response is a cause or a consequence of the disease process. In addition, there is debate over the contribution of axonal damage to clinical progression. We have described a murine model of relapsing-remitting MS (RR-MS), the most common form of the disease, following immunisation with the myelin component, myelin oligodendrocyte glycoprotein (MOG). We showed that a single injection of a MOG peptide (MOG(35-55)) in NOD/Lt mice induces a paralytic relapsing disease with extensive plaque-like demyelination. This model also mimics many of the immunological features associated with RR-MS. To investigate the relationship between clinical episodes, inflammation, and demyelination/remyelination, we analysed lesions during each attack and remission over the course of the disease, using histological, immunocytochemical, and electron microscopy (EM) techniques. We show that morphological features of lesions in our model resemble those observed in MS. Indeed, severe inflammation and demyelination coincide with the peak of clinical episodes while remissions are characterised by quiescent plaques. Furthermore, axonal damage is evident from the earliest stage of the disease and increases in severity with subsequent relapses. These data establish that in the model of MS-like disease, the peak of clinical episodes coincides with severe inflammation and demyelination and that axonal pathology correlates with clinical progression. PMID- 11276126 TI - Regulation of oligodendrocyte development. AB - Oligodendrocytes are myelinating cells in the central nervous system. Recent studies demonstrated that oligodendrocyte progenitor cells are generated from a restricted region in the ventricular zone. In the rodent spinal cord, progenitor cells appear from narrow and bilateral longitudinal columns in the ventral ventricular zone, and then migrate dorsally. This ventral-specific appearance of oligodendrocyte progenitors may be controlled along the dorso-ventral axis in the spinal cord by extrinsic signals secreted from both the dorsal and ventral cords. The combined action of the Notch signaling pathway and a basic helix-loop-helix class of transcription factors may modulate this early specification of spinal oligodendrocytes and also be involved in multiple steps of oligodendrocyte differentiation. PMID- 11276127 TI - Expression of transcription factors during oligodendroglial development. AB - Transcription factors coordinate the orderly changes of gene expression that underlie all developmental processes including those of oligodendrocytes. In comparison to other systems, relatively little is known about the role of transcription factors during oligodendrocyte development. However, recent years have seen the identification of oligodendroglial transcription factors that, not surprisingly, belong to the same gene families that are also important in other tissues or cell lineages. Some transcription factors such as the bHLH (basic helix-loop-helix) proteins Olig-1 and Olig-2 or the high-mobility-group protein Sox10 are expressed already early during development of the oligodendrocyte lineage, whereas expression of other transcription factors such as the homeodomain protein Gtx/Nkx6.2 only start at the time of terminal differentiation. Once turned on, expression of these proteins can either be permanent as in the above-mentioned cases or transient as exemplified by the POU domain proteins Tst-1/Oct6/SCIP, Brn-1 and Brn-2. Analyses of these transcription factors has already led to the identification of important principles such as functional redundancy between co-expressed proteins, unexpected divergence in the transcription factor repertoire of oligodendrocytes and Schwann cells, and equally unsuspected similarities in transcription factor usage between oligodendrocytes and neurons. Although so far only a small percentage of oligodendroglial transcription factors has been identified, these are excellent candidates for regulators of cell type specification, lineage progression, and terminal differentiation. PMID- 11276128 TI - Re-evaluation of nestin as a marker of oligodendrocyte lineage cells. AB - Maturation of oligodendrocyte progenitors (O2A) is characterized by morphological changes and the sequential expression of specific antigens leading to the formation of myelin membrane. Monoclonal antibodies A2B5, A007, anti-vimentin, and anti-galactocerebroside, recognize oligodendroglia at different stages of development. The neuroepithelial precursor marker nestin is also expressed by the oligodendroglial lineage; we have used enriched populations of progenitors isolated from neonatal rat brain cultures to further examine the cellular distribution of this intermediate filament protein. The phenotypic distribution of nestin positive cells among the oligodendrocyte lineage showed that 65% reacted with A2B5, whereas only 5% were A007(+), and 4% galactocerebroside(+). The remaining 25% of the cells were not labeled and had small cellular bodies devoid of processes, characteristic of the pre-O2A progenitor. Further analysis of the nestin(+) population showed that the majority of the cells were also vimentin(+). Antibody-dependent complement mediated cytolysis of A2B5(+) (O2A cells) and galactocerebroside(+) (mature oligodendrocytes) cells left a population of nestin(+) cells that were induced to proliferate in the presence of growth factors and to differentiate into A2B5(+) and galactocerebroside(+) cells. Proliferating cells maintained in the presence of platelet-derived growth factor or basic fibroblast growth factor retained nestin expression along with A2B5. By contrast, in serum-free medium nestin expression decreased while postmitotic cells acquired A007 and galactocerebroside. Our results suggest that nestin expression is a marker of pre-O2A cells that is maintained in proliferating glial progenitors, but is quickly down-regulated in postmitotic oligodendrocytes (A007(+)/galacto-cerebroside(+)) along with A2B5 and vimentin. However, other glial cells including type 2 astrocytes and some amoeboid microglia also share nestin expression. PMID- 11276129 TI - Tracking oligodendrocytes during development and regeneration. AB - Over the past decade, advances in strategies to tag cells have opened new avenues for examining the development of myelin-forming glial cells and for monitoring transplanted cells in animal models of myelin insufficiency. The strategies for labelling glial cells have encompassed a range of genetic modifications as well as methods for directly attaching labels to cells. Genetically modified oligodendrocytes have been engineered to express enzymatic (e.g., beta galactosidase, alkaline phosphatase), naturally fluorescent (e.g., green fluorescent protein), and antibiotic resistance (e.g., neomycin, zeomycin) reporters. Genes have been introduced in vivo and in vitro with viral or plasmid vectors to somatically label glial cells. To generate germ-line transmission of tagged oligodendrocytes, transgenic mice have been created both by direct injection into mouse fertilized eggs and by "knock-in" of reporters targetted to myelin gene loci in embryonic stem cells. Each experimental approach has advantages and limitations that need to be considered for individual applications. The availability of tagged glial cells has expanded our basic understanding of how oligodendrocytes are specified from stem cells and should continue to fill in the gaps in our understanding of how oligodendrocytes differentiate, myelinate, and maintain their myelin sheaths. Moreover, the ability to select oligodendrocytes by virtue of their acquired antibiotic resistance has provided an important new tool for isolating and purifying oligodendrocytes. Tagged glial cells have also been invaluable in evaluating cell transplant therapies in the nervous system. The tracking technologies that have driven these advances in glial cell biology are continuing to evolve and present new opportunities for examining oligodendrocytes in living systems. Microsc. Res. Tech. 52:766-777, 2001. Published 2001 Wiley-Liss, Inc. PMID- 11276130 TI - Impact of a multidisciplinary day program on disease and healthcare costs in children and adolescents with severe asthma: a two-year follow-up study. AB - For patients whose asthma remains in poor control necessitating high utilization of medical services, a referral to a specialized "center of excellence" is often considered. A decade ago, we evaluated our pediatric asthma program of long-term hospitalization (median stay of 75 days) and found significant decreases in subjects' medical utilization following this intervention. In an effort to contain treatment costs, the former program was markedly altered to one of abbreviated stay with emphasis on family management of asthma. The purpose of the present study was to determine the outcome of children treated in the revised program with regard to disease severity, quality of life, and subsequent utilization of medical resources. Children with severe asthma who were admitted to the program and fulfilled study criteria were consecutively enrolled. Data was obtained concerning disease characteristics, treatment, and quality of life at admission, and at 1 and 2 years following discharge. Medical records for the year prior to program admission and for the 2 years following discharge were coded for medical care encounters. Ninety-eight children, aged 9 months to 18 years (mean age, 10.9 years), were enrolled. They participated in the program for a mean of 15.6 ( +/- 8 SD), median of 15.0, and range of 2-51 treatment days. The group showed significant improvement (P < 0.0001) from admission to 1- and 2-year follow-up in median corticosteroid use, asthma functional severity, perceived competence in asthma management, and quality of life for both caregiver and child. Medical record data showed significant improvement (P < 0.0001) at both 1- and 2-year follow-up in median number of corticosteroid bursts, emergency department visits, hospital days, and overall utilization of medical care encounters. A median total medical encounter cost/patient of $16,250 ($6,972 $25,714 interquartile range (IQR)) for the year prior to program participation was reduced to $1,902 ($505-$6,524 IQR) at 1-year and $690 ($185-$3,550 IQR) at 2 year follow-up (P < 0.0001). We conclude that multidisciplinary care in a short term, outpatient, day treatment program can significantly contribute to improvement in asthma severity, quality of life, and reduction in healthcare costs. PMID- 11276131 TI - Are peak flow and symptom measures good predictors of asthma hospitalizations and unscheduled visits? AB - Epidemiologic studies of pediatric respiratory health often include objective measures such as peak expiratory flow (PEF), and subjective measures such as symptom reports. These measures, however, are poorly correlated with each other, and there is little evidence that PEF is useful in predicting important health outcomes. Within a cohort of 791 inner-city children with asthma, we examined correlations between a series of five peak flow measures and five symptom scores obtained from 2-week diaries. The strongest correlations were found between "total peak flow lability" defined as: [(diary maximum - diary minimum)/diary mean] and "% of days with chest tightness" (r = 0.31). Logistic models evaluated peak flow and symptoms as predictors of an important health outcome: hospitalization or emergency department or unscheduled clinic visit for asthma within 30 days of starting the diary. Each of the peak flow and symptom measures was significantly related to utilization. However, the predictive power of each measure was low (range of area under ROC curve, 0.54-0.67). Models including only peak flow or symptoms had greater prediction than models with risk factors such as atopy, asthma persistence, and age. The prediction from a model with the risk factors and symptoms was not improved by adding a peak flow measure to the model (increase in area under ROC, 0.67-0.68). Stratified analyses suggest that prediction was similar in the fall vs. winter, spring, and summer months. Greater prediction of health outcomes was found among more persistent asthmatics and children who were nonatopic. These findings suggest that in a research setting, peak flow monitoring in children did not add prediction beyond that obtained from symptom reports. Pediatr Pulmonol. 2001; 31:190-197. Published 2001 Wiley-Liss, Inc. PMID- 11276132 TI - Anaerobic fitness in children with asthma: adaptation to maximal intermittent short exercise. AB - Nineteen asthmatic boys (aged 13.4 years, 25-75 percentile: 11.5-15.1 years) performed short bouts of maximal exercise (force-velocity test) to test their anaerobic fitness and tolerance of maximal anaerobic exercise. Fourteen healthy boys (aged 13.9 years, 25-75 percentile: 11.6-15.7 years) matched for anthropometric characteristics including lean body mass (LBM), pubertal stage, and weekly physical activity formed a control group. The maximal anaerobic power (W(ana)) was measured during the force-velocity test. The maximal oxygen uptake (V'(O2max)) was assessed during a standard graded exercise test. Pre- and post exercise pulmonary function was measured by body plethysmography. The asthmatic children exhibited lower W(ana) than the control group (8.2 watt.kg(-1) LBM, 25 75 percentile: 7.55-10.6 vs. 11.3 watt.kg(-1) LBM, 25-75 percentile: 9.46-14.1; P = 0.01). V'(O2max) was also diminished in the asthmatic group (P = 0.01). Multiple stepwise regression models revealed that Tanner's score (P < 0.001) and the diagnosis of asthma (P < 0.01) were the best predictors of W(ana). In conclusion, a diminished anaerobic fitness could contribute to the overall exercise limitation in asthmatic children. PMID- 11276133 TI - Cellular and noncellular components of bronchoalveolar lavage fluid in HIV-1 infected children with radiological evidence of interstitial lung damage. AB - Children with acquired immune deficiency syndrome (AIDS) commonly have recurrent infectious and noninfectious lung complications that ultimately end in death. To study the intensity of alveolar inflammation and to evaluate the clinical utility of bronchoalveolar lavage (BAL) in children with HIV-1 infections, we retrospectively analyzed differential cell counts, lymphocyte subsets, and fibronectin and hyaluronic acid concentrations in BAL fluid of 18 HIV-1-positive children (9 boys, mean age 3.5 years, range 5 months-8 years) with radiological evidence of interstitial lung disease, and 19 control children who had undergone BAL for clinical indications not involving the lung parenchyma (13 boys, mean age 3 years, range 2 months-14 years). BAL fluid from 89% of the HIV-1 infected children showed CD8+ve lymphocytic alveolitis expressing HLA-DR, CD54, and CD 69 antigens. BAL fluid from HIV-infected patients typically contained markedly increased percentages and numbers of lymphocytes (P < 0.0001) and eosinophils (P < 0.04) and significantly higher concentrations of albumin (P < 0.05) and fibronectin (P < 0.0006) than fluids from control children. Whereas BAL cellular components did not differ in P. carinii-positive and P. carinii-negative HIV-1 infected children, fibronectin concentrations were significantly higher in P. carinii-positive than negative children. BAL cell differentials and noncellular components were related neither to severity of disease nor to patients' disease progression. These findings indicate that BAL is useful in studying the intensity of lung inflammation in children with HIV-1 infections and radiologically documented interstitial lung disease, but provides no information on the subsequent clinical course. PMID- 11276135 TI - Serial changes in levels of IL-6 and IL-1beta in premature infants at risk for bronchopulmonary dysplasia. AB - The aim of this study was to define the inflammatory changes occurring in the lungs of infants at risk for bronchopulmonary dysplasia (BPD) over the first 28 days of life, and to define an optimal strategy for steroids therapy in the prevention of BPD. We measured levels of interleukin-6 (IL-6) and interleukin-1 beta (IL-1beta) in tracheal aspirate (TA) samples and blood of premature infants with severe respiratory distress syndrome RDS (n = 45) on the first day of life prior to initiation of surfactant therapy and on days 5-7, 12-14, 19-21, and 26 28. Levels of IL-6 and IL-1beta were determined with a commercially available enzyme-linked immunoassay. Logistic regression analyses were performed in order to examine differences in trends in levels of IL-6 and IL-1beta between groups of infants. Infants were divided into group I (n = 30, FiO(2) < or = 0.35 at 28 days) and group II (n = 15, FiO(2) > 0.35 based on their likelihood of developing BPD at 36 weeks postconceptional age (PCA). The infants were comparable with respect to mean ( +/- SEM) birth weight (895 +/- 33 g vs. 900 +/- 40 g), gestational age (27 +/- 0.38 weeks vs. 27 +/- 0.54 weeks), and severity of respiratory illness at entry into the study (mean airway pressure: 12 +/- 1 cmH(2)O vs. 12 +/- 1 cmH(2)O, and oxygen index: 15 +/- 2 vs. 19 +/- 4) (group I vs. group II, respectively). Logistic regression analyses failed to reveal any significant differences in linear trends of levels of IL-6 and IL-1beta in TA samples between both groups of infants. No particular pattern of change in levels of IL-6 or IL-1beta could be identified among groups of infants. Levels of IL-6 and IL-1beta in TA samples on the first day of life failed to predict the need for FiO(2) > 0.35 at 28 days of age. We could not identify an increasing trend or a specific pattern of changes in postnatal levels of IL-6 or IL-1beta in TA samples of infants who were at greater risk of developing BPD at 36 weeks PCA compared to infants who were not. PMID- 11276134 TI - Evidence of early systemic activation and transendothelial migration of neutrophils in neonates with severe respiratory distress syndrome. AB - Several observations imply that the early inflammatory response involving activated neutrophils, tissue macrophages, and cytokines plays an important role in the pathogenesis of neonatal respiratory distress syndrome (RDS) and progression to bronchopulmonary dysplasia (BPD). The aim of this study was to test the hypothesis that changes in circulating neutrophil number and function and plasma levels of cytokines, consistent with neutrophil activation and migration to the tissues, occur during the early stages of neonatal RDS. For this purpose we measured peripheral blood levels of certain immunological parameters that promote neutrophil activation and transendothelial migration. Twenty preterm neonates with severe RDS and 20 healthy infants matched for gestational age were the subjects. The absolute neutrophil count (ANC), and plasma levels of interleukin-6 (IL-6), granulocyte colony-stimulating factor (G-CSF), and sL selectin using an enzyme-linked immunosorbent assay (ELISA), neutrophil CD11b expression, and respiratory burst activity (RBA) using flow cytometry, were measured within 24 h after birth. The two groups were comparable regarding perinatal characteristics. None of the neonates studied had any clinical or laboratory evidence of infection by the time of blood sampling. The immunological investigation showed that the RDS neonates had significantly lower ANC (P = 0.032), higher expression of the CD11b on neutrophils (P = 0.0065), and higher G CSF and IL-6 plasma levels (P = 0.0047 and P < 0.0001, respectively) in comparison to healthy preterm neonates. The neutrophil RBA and plasma sL-selectin levels did not differ significantly between the two groups. We conclude that in neonates with severe RDS, there is evidence of a systemic neutrophil activation early in the course of the disease, supporting the view of a contributing role of activated neutrophils in the pathogenesis of RDS. PMID- 11276136 TI - Effect of choice of reference equation on analysis of pulmonary function in cystic fibrosis patients. AB - Pulmonary function is an important measure of disease severity and prognosis in cystic fibrosis (CF). It is generally expressed as a percentage of a predicted value, calculated using regression equations derived from a reference population. A number of reference equations are in widespread use. The purposes of this study were to determine: 1) the extent to which, for a given absolute FEV(1) value, percent of predicted (PPFEV(1)) values vary when derived by different reference equations; and 2) whether these differences affect conclusions of longitudinal and cross-sectional analyses. Subjects were all Caucasians 6-18 years old in the 1990 Cystic Fibrosis Foundation Registry. We found clinically important discrepancies in PPFEV(1) when calculated by the methods of Dockery et al. [Am Rev Respir Dis 1983;128:405-412] and Wang et al. [Pediatr Pulmonol 1993;15:75-78] as compared to Knudson et al. [Am Rev Respir Dis 1983;127:725-734] or Polgar and Promadhat [Pulmonary Function Testing in Children 1971; Philadelphia: W.B. Saunders]. In longitudinal analyses, the choice of reference equation resulted in varying apparent rates of decline in FEV(1). For example, among subjects ages 12 14 years in 1990, the decline in PPFEV(1) from 1990-1995 varied between 2-11%, depending on the choice of reference equation. In cross-sectional analyses, the choice of reference equation affected the distribution of subjects classified as having mild, moderate, or severe lung disease. CF physicians should be aware of the impact of choice of reference equation in both clinical care and research. PMID- 11276137 TI - Measurements of resistance by the interrupter technique and of transcutaneous partial pressure of oxygen in young children during methacholine challenge. AB - Measurement of bronchial airway responsiveness requires noninvasive techniques in young children. The study was designed to examine the changes in resistance as measured using the interrupter technique (Rint) at the dose of methacholine (M) that induced a fall in transcutaneous partial pressure in O2 (P(tc)O2) > or = 20% (PD(20)P(tc)O2) in young children. Rint was calculated using the linear back extrapolation method (Rint(L)) and the end-interrupter method (Rint(EI)). Twenty two children (mean age, 5.2 +/- 1.1 years; range, 3.4 - 7.1 years) with nonspecific respiratory symptoms (mainly chronic cough, n = 17) were tested. P(tc)O2, Rint(L), and Rint(EI) were measured before the test, after saline challenge (baseline (B)), after each dose of M delivered by a dosimeter, and after bronchodilator (BD) inhalation. P(tc)O2 decreased significantly during M challenge, from 85 +/- 6 mmHg (B) to 62 +/- 9 mmHg (P < 0.05), and increased after BD inhalation, to 82 +/- 8 mmHg. Rint(L) and Rint(EI) increased significantly during M challenge, from 0.94 +/- 0.2 KPa/L/s and 1.11 +/- 0.19 KPa/L/s (B) to 1.27 +/- 0.35 KPa/L/s and 1.47 +/- 0.37 KPa/L/s, respectively (P < 0.05), and decreased after BD inhalation to 0.80 +/- 0.17 KPa/L/s and 0.95 +/- 0.18 KPa/L/s, respectively. Nineteen of 22 children reached the PD(20)P(tc)O2 at a dose of M ranging from 50-400 microg. At the PD(20)P(tc)O2, significant changes in Rint(L) and Rint(EI) (sensitivity index (SI) > or = 2) were found in 79% and 63% of children, respectively. We conclude that: 1) M challenge using P(tc)O2 is safe in young children; and 2) our findings are not in favor of the use of Rint as the only indicator of bronchial reaction in young children during M challenge. PMID- 11276138 TI - Albuterol delivery in a neonatal ventilated lung model: Nebulization versus chlorofluorocarbon- and hydrofluoroalkane-pressurized metered dose inhalers. AB - The aim of this study was to compare albuterol delivery in a neonatal ventilated lung model, using three delivery methods: 1) jet nebulizer; 2) chlorofluorocarbon pressurized metered dose inhaler (CFC-MDI) actuated into an ACE(R) spacer; and 3) hydrofluoroalkane-pressurized MDI (HFA-MDI) actuated into an ACE(R) spacer. The bench model consisted of a mechanically ventilated infant test lung with ventilator settings to simulate a very low birth weight neonate with moderate lung disease. Albuterol solution (0.5%) was nebulized at the humidifier and temperature port, 125 cm and 30 cm from the Y-piece, respectively. Albuterol metered dose inhalers (MDIs) were actuated into an ACE(R) spacer that was tested in two positions: 1) inline between the endotracheal (ET) tube and the Y-piece; and 2) attached to the ET tube and administered by manual ventilation. Albuterol was collected on a filter at the distal end of the ET tube and was quantitatively analyzed by high performance liquid chromatography. Albuterol delivery by CFC-MDI (position 1, 4.8 +/- 1.0%, vs. position 2, 3.8 +/- 1.6%, P > 0.05) and HFA-MDI (position 1, 5.7 +/- 1.6%, vs. position 2, 5.5 +/- 2.4%, P > 0.05) were significantly greater than delivery by nebulization at 30 cm (0.16 +/- 0.07%) and 125 cm (0.15 +/- 0.03%) from the Y-piece (P < 0.001). A single actuation of albuterol MDI delivered the equivalent of nebulizing 2.5-3.7 mg of albuterol solution. We conclude that albuterol administered by MDI and ACE(R) spacer resulted in more efficient delivery than by nebulization in this mechanically ventilated neonatal lung model. There was no significant difference in drug delivery between CFC-MDI and HFA-MDI; nor did the placement of the spacer significantly affect drug delivery. PMID- 11276139 TI - Physical effects of heliox versus oxygen on measurements of functional residual capacity by the nitrogen washout technique in small lung volumes: a model study. AB - Measurements of functional residual capacity (FRC) by the nitrogen (N(2)) washout technique yield low N(2) signals in neonates and preterm infants, especially when they are on high fractions of inspired oxygen (FiO(2)). Thus, recorded values often lie in the low range of detectability. We hypothesized that using heliox instead of oxygen as a washout gas would affect the electric discharge conditions of N(2) molecules in a standard UV analyzer and thus yield higher N(2) signals. We performed three laboratory experiments using conditions similar to those in neonates with pulmonary disease, reproducing different initial FiO(2) values and very small lung volumes. Standard calibration procedures with physical models between 13.5-87 mL using a calibration syringe and purpose-built small calibration cylinders were carried out, and washout gas was either pure oxygen (as is general practice) or heliox at different ratios. We observed that the calibration line with heliox was shifted upwards and the slope was increased, depending on helium concentration and initial FiO(2). Since this effect was dose dependent with respect to the proportion of helium in the washout gas, this strongly suggests a physical process elicited by the noble gas helium. We conclude that the heliox wash-out modification may help to increase the accuracy of FRC measurements and thus may enable studies of smaller patients or patients on high inspired FiO(2). PMID- 11276141 TI - Oral health and heart disease. PMID- 11276140 TI - Cheyne-Stokes respiration as an additional risk factor for pulmonary hypertension in a boy with trisomy 21 and atrioventricular septal defect. AB - Central ventilation disorders(1) and airway obstruction(2) with chronic hypoxemia are causally related to cor pulmonale. Pulmonary vascular resistance is often reversible, and hypoxic pulmonary hypertension often responds to treatment with supplemental oxygen. Oxygen therapy during sleep may be useful as a temporary palliative treatment in children with obstructive sleep apnea syndrome (3) and Cheyne-Stokes respiration (CSR) in congestive heart failure(4). This type of sleep-related breathing disorder is characterized by periodic crescendo decrescendo alterations in tidal volume. Proposed mechanism include an increased central nervous system sensitivity to changes in arterial PCO2 and PO2, a decrease in total body stores of CO2 and O2 with resulting instability in arterial blood gas tensions in response to changes in ventilation, and an increased circulatory time. Clinical features of obstructive and central sleep related breathing disorders include daytime somnolence, unusual breathing patterns, failure to thrive, and cyanosis masquerading as cyanotic congenital heart disease(2). Down syndrome is often associated with cardiac malformations, left to right shunt, and the development of pulmonary hypertension(5). However, this may be exacerbated by sleep-related breathing disorders, as illustrated in the following case report. PMID- 11276142 TI - Defibrillation, coming to a neighborhood near you. PMID- 11276143 TI - Extra heart beats: usually benign...but not always. PMID- 11276144 TI - Low-dose thiazide diuretics protect bones. PMID- 11276145 TI - Ask the Doctor. I'm 45 years old. At the doctor's office, my blood pressure readings are usually pretty high (the top number may reach 150 or 160), but my home monitor shows numbers more like 130/90. My doctor calls my problem "white coat hypertension" and reassures me that it isn't dangerous. But it bothers me that my pressure shoots up like that. I have plenty of stressful moments every day, and my blood pressure must be going up then, too. Should I be on medication? PMID- 11276146 TI - Weight gain and loss. PMID- 11276148 TI - Respiratory disease. Cough remedies. PMID- 11276147 TI - Colon cancer. The pros and cons of the screening tests. PMID- 11276149 TI - By the way, doctor...I have hypertension, and I take a beta blocker called atenolol and an ACE inhibitor every morning. But I find that my blood pressure is high when I first get up. As the day goes on, my pressure falls. It seems like it's always pretty good by the time I see my doctor, so she thinks everything is fine. But the high numbers worry me. Should I be on another drug? PMID- 11276150 TI - Panic disorder. PMID- 11276151 TI - Stalking. PMID- 11276152 TI - Exercise against depression. PMID- 11276153 TI - GHB: its use and misuse. PMID- 11276154 TI - Drinking trends and thinking trends. PMID- 11276155 TI - Diet and prostate cancer: new grains of hope. PMID- 11276156 TI - Don't fall. PMID- 11276157 TI - Medical memo. Low-dose aspirin gets the nod. PMID- 11276158 TI - On call. I've read that a special treatment can wash excess cholesterol out of my blood. Is it the same as chelation therapy? Does it work? PMID- 11276159 TI - A good reason to cast modesty aside. PMID- 11276160 TI - Reducing osteoporosis risk. PMID- 11276161 TI - New ways to stay dry. PMID- 11276162 TI - By the way, doctor. I recently received an e-mail warning about a risk for breast cancer associated with using antiperspirants. Are you familiar with this theory? Is it valid? PMID- 11276163 TI - By the way, doctor. For the past few months, I've had a pain in my leg, which my doctor calls piriformis syndrome. What can you tell me about this? PMID- 11276164 TI - Minor symptoms: the illnesses of the 21st Century. PMID- 11276165 TI - Osteoporotic fractures of proximal femur: clinical and epidemiological features in a population of the city of Sao Paulo. AB - CONTEXT: It is believed that about 25% of menopausal women in the USA will exhibit some kind of fracture as a consequence of osteoporosis. Fractures of the proximal femur are associated with a greater number of deaths and disabilities and higher medical expenses than all the other osteoporotic fractures together. OBJECTIVE: To study the clinical and epidemiological features of patients with proximal femur fracture in hospitals in Sao Paulo. DESIGN: Transversal and retrospective study. LOCAL: Hospital Sao Paulo and Hospital Servidor Publico Estadual "Francisco Morato Oliveira". PARTICIPANTS: Patients aged sixty-five years or more hospitalized because of proximal femur fracture, from March to November 1996 (N = 73). This group was compared to patients of the same age without fracture of the proximal femur. INTERVENTION: Evaluation of weight, height, body mass index; lifestyle habits (physical activity at home, ingestion of dairy calcium, drinking of coffee, smoking habit), gynecological history (ages at menarche and menopause, number of pregnancies and lactations), previous morbidity, use of medications, history of previous fractures, family history of osteoporosis. MEASUREMENT: The comparison of the different data regarding lifestyle habits between the two groups was made using the chi-squared test. Other data were analyzed using the Mann--Whitney test. P pound 0.05 was considered significant. RESULTS: We noted a predominance of proximal femur fracture among females in relation to males (a female/male ratio of 3.3:1) with a progressive increase in the frequency of proximal femur fracture with age in both sexes. The group with proximal femur fracture, in comparison with the control group, showed a lower body mass index, less physical activity, and a greater number of pregnancies and lactations. Other data were not different. CONCLUSION: In accordance with the literature, we found a predomination of proximal femur fracture in women in relation to men, and a favorable effect of higher body mass index and physical activity for decreasing the frequency of proximal femur fracture. We also discuss the role of pregnancies and lactation on the frequency of proximal femur fracture. PMID- 11276166 TI - Effects of hypertension on maternal adaptations to pregnancy: experimental study on spontaneously hypertensive rats. AB - CONTEXT: Animal models for essential hypertension have been used for understanding the human pathological conditions observed in pregnant hypertensive women. OBJECTIVE: To study the possible effects of pregnancy on hypertension and of hypertension on pregnancy in spontaneously hypertensive rats (SHR), and in their normotensive Wistar-Kyoto (WKY) counterparts. TYPE OF STUDY: Comparative study using laboratory animals. SETTING: Animal Research Laboratory of Clinical Medicine at the Medical School of Botucatu, Sao Paulo State University, Brazil. SAMPLE: Ten to twelve-week-old virgin female normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). The animals were separated into four groups: 15 pregnant spontaneously hypertensive rats (SHR-P), 10 non-pregnant spontaneously hypertensive rats (SHR-NP), 15 pregnant normotensive rats (WKY-P), and 10 non-pregnant normotensive rats (WKY-NP). MAIN MEASUREMENTS: The blood pressure was evaluated by the tail cuff method, in rats either with or without prior training for the handling necessary for tail cuff measurements. The maternal volume expansion was indirectly evaluated by weight gain, and by systemic parameters as hematocrit, hemoglobin, total protein, albumin and sodium retention. The perinatal outcome of pregnancy was evaluated by analysis of resorptions, litter size, rate of low weight and number of stillbirths. RESULTS: The late fall in blood pressure in the pregnant SHR strain and in the normotensive WKY strain can only be detected in rats previously trained to accept the handling necessary for the tail cuff measurement. During pregnancy the body weight gain was significantly higher in WKY than in SHR rats. Systemic parameters were significantly lower in pregnant WKY rats than in non-pregnant WKY rats, while no differences were observed between pregnant and non-pregnant SHR groups. In pregnant WKY rats the sodium retention was higher from the 13th day onwards, while in SHR rats this occurred only on the 21st day. The characteristics of reproductive function such as number and weight of fetus, perinatal mortality and the resorption rate were significantly affected in the SHR strain. CONCLUSION: The SHR strain may be considered as a model for chronic hypovolemic maternal hypertension, with the fetal growth retardation being determined by this hypovolemic state. PMID- 11276167 TI - Use of arteriography for the initial evaluation of patients with intermittent lower limb claudication. AB - CONTEXT: Many patients with intermittent claudication continue to be forwarded to the vascular surgeon for initial evaluation after arteriography has already been accomplished. OBJECTIVE: The main objective of this work was to analyze the usefulness and the need for this procedure. TYPE OF STUDY: Retrospective study. SETTING: The patients were divided into two groups: Group 1, with the arteriography already performed and Group 2 without the initial arteriography. PARTICIPANTS: One hundred patients with intermittent claudication were retrospectively studied. Other specialists had forwarded them for the first evaluation of intermittent claudication, without any previous treatment. MAIN MEASUREMENTS: All patients were treated clinically for at least a 6-month period. The total number of arteriographies performed in the two groups was compared and the need and usefulness of the initial arteriography (of Group 1) was also analyzed. RESULTS: The evolution was similar for both groups. The total number of arteriographies was significantly higher in Group 1 (Group 1 with 53 arteriographies vs. Group 2 with 7 arteriographies). For this group, it was found that arteriography was only useful in five cases (10%), because the surgeries were based on their findings. However, even in those cases, no need for arteriography was observed, as the procedure could have been performed at the time of surgical indication. CONCLUSION: There are no indications for arteriography in the early evaluation of patients with intermittent claudication, because it does not modify the initial therapy, independent of its result. In cases where surgical treatment is indicated, this procedure should only be performed prior to surgery. PMID- 11276168 TI - Height, weight, weight change and risk of breast cancer in Rio de Janeiro, Brazil. AB - CONTEXT: The relationship between body size and breast cancer still remains controversial in considering menopausal status. OBJECTIVE: To evaluate the association of height, weight and weight changes with breast cancer in the city of Rio de Janeiro, Brazil. DESIGN: Case-control study. SETTING: National Cancer Institute (INCA), Rio de Janeiro, Brazil, and State University of Rio de Janeiro (UERJ). SAMPLE: 177 incident cases of invasive breast cancer admitted to the main hospital of INCA between May 1995 and February 1996, and 377 controls recruited from among female visitors to the same hospital. MAIN MEASUREMENTS: Height and weight were measured and information on maximum weight, weight at ages 18 and 30 years, and potential risk factors were ascertained by interview at the hospital. RESULTS: Height was not related to risk of breast cancer among both pre and postmenopausal women. Nevertheless, women in this study were shorter than in studies that have found a positive association. Premenopausal women in the upper quartile of recent body mass index (BMI) and maximum BMI showed a reduced risk of breast cancer (P for trend < or = 0.03). Weight loss between ages 18 and 30 years and from 18 years to present was also associated with breast cancer among premenopausal women. CONCLUSIONS: These findings may merely indicate the known association between leanness and breast cancer. Further studies should explore the role of weight loss on breast cancer risk. PMID- 11276169 TI - Evaluation of invasive and non-invasive methods for the diagnosis of Helicobacter pylori infection in symptomatic children and adolescents. AB - CONTEXT: Multiple diagnostic methods are available for the detection of Helicobacter pylori infection, but at present no single one can be used as the gold standard. OBJECTIVE: The aim of this study was to evaluate the diagnostic accuracy of 3 invasive and 2 non-invasive methods for detection of Helicobacter pylori infection in symptomatic children and adolescents. DESIGN: Prospective cohort study SETTING: Peptic Disease outpatients service, Discipline of Pediatric Gastroenterology, Universidade Federal de Sao Paulo / Escola Paulista de Medicina. PATIENTS: Forty-seven patients who underwent endoscopy because of dyspeptic symptoms. DIAGNOSTIC METHODS: Endoscopy with gastric biopsies for 3 invasive (rapid urease test, histology and culture) and 2 non-invasive methods (a commercial ELISA serology and 13carbon urea breath test - isotope ratio mass spectrometry) for detection of Helicobacter pylori infection. MAIN MEASUREMENTS: Sensitivity, specificity, positive and negative predictive values of each method and agreement and disagreement rates between the methods. RESULTS: Forty-seven patients [mean age, 11y9mo (SD 2y10mo), 27 female and 20 male]; 62% of them were Helicobacter pylori-positive. All methods agreed in 61%, and were negative in 21% and positive in 40%. The greatest concordance between 2 methods occurred between the invasive methods: histology and rapid urease test (89.6%) and histology and culture (87.5%). The greatest sensitivity, considering Helicobacter pylori positive cases, for any combination of 3 or more tests, was achieved by the rapid urease test (S=100%), followed by histology, serology and 13carbon-urea breath test (S=93.1%) and lastly by culture (S=79.3%). The highest specificity was obtained by histology (100%) and culture (100%), followed by the rapid urease test (84.2%), serology (78.9%) and 13carbon-urea breath test (78.9%). CONCLUSIONS: Our results suggest that among invasive methods, an association between the rapid urease test and histology constituted the best choice for the detection of Helicobacter pylori infection. If results of histology and the rapid urease test are different, serology may be recommended. PMID- 11276171 TI - The perimenopause, depressive disorders, and hormonal variability. AB - CONTEXT: Several investigations have postulated that the perimenopause may represent a period of increased psychiatric vulnerability, particularly for mood disorders. This review characterizes the perimenopause, including biological changes, the influence of psychosocial factors and the most common clinical manifestations. Clinic-based studies and community-based surveys addressing the prevalence of depressive symptoms in perimenopausal women are critically reviewed. We also discuss the potential greater vulnerability to mood disturbance during the perimenopause in response to hormonal variability. A therapeutic algorithm for management of depressive symptoms in middle-aged perimenopausal women is also presented. The role of estrogen in the treatment of perimenopausal depressive symptoms is particularly discussed. In addition, we review the existing data regarding the potential efficacy of estrogen as an antidepressant agent (monotherapy, augmentation strategy or prophylaxis). DESIGN: Narrative review. PMID- 11276170 TI - Involuntary weight loss in elderly individuals: assessment and treatment. AB - CONTEXT: The loss of body weight and fat late in life is associated with premature death and increased risk of disability, even after excluding elderly subjects who have a preexisting disease. Although it is important to recognize that periods of substantially positive or negative energy balance and body weight fluctuation occur as a normal part of life, weight losses greater than 5% over 6 months should be investigated. We can divide the major causes of weight loss in the elderly into 4 categories: social, psychiatric, due to medical conditions, and age-related. The clinical evaluation should include a careful history and physical examination. If these fail to provide clues to the weight loss, simple diagnostic tests are indicated. A period of watchful waiting is preferable to blind pursuit of additional diagnostic testing that may yield few useful data, if the results of these initial tests are normal. The first step in managing patients with weight loss is to identify and treat any specific causative or contributing conditions and to provide nutritional support when indicated. Non orexigenic drugs have found an established place in the management of protein energy malnutrition. Early attention to nutrition and prevention of weight loss during periods of acute stress, particularly during hospitalization, may be extremely important, as efforts directed at re-feeding are often unsuccessful. DESIGN: Narrative review. PMID- 11276172 TI - Latent autoimmune diabetes of the adult (LADA) in a Brazilian Indian. AB - CONTEXT: Latent autoimmune diabetes of the adult (LADA) as originally described represents perhaps as many as 10 - 20% of adult-onset patients with diabetes. DESIGN: case report. CASE REPORT: A 38-year-old Brazilian Xavante-Je Indian with Latent Autoimmune Diabetes of the Adult (LADA) is described, coming from the Sangradouro community in Poxoreu, Mato Grosso. The onset of diabetes after reaching 25 years of age, the evolution to insulin deficiency after a period of insulin-independence and the presence of auto-antibodies to glutamic acid decarboxylase (GAD) characteristic of LADA were present. This patient may represent the first case of LADA in a Brazilian with full Indian heritage. Further studies are necessary to verify the prevalence of this new type of diabetes in this population that does not have Caucasoid admixture and has a particular environmental background. PMID- 11276173 TI - Multicentric pheochromocytoma and involvement of the inferior vena cava. AB - CONTEXT: Extension of pheochromocytomas to the inferior vena cava is rare. Multicentric tumors are rare as well, being present in up to 10% of cases. Surgery is the treatment of choice because of the long-term survival free of disease. DESIGN: Case report. CASE REPORT: We report on a case of right adrenal pheochromocytoma with extension to the supra-diaphragmatic vena cava, which underwent surgical excision through thoracophrenic laparotomy without the need for cardiopulmonary bypass. In a 6-year follow-up, another pheochromocytoma was found in the infra-renal Zuckerkandl's organ. Complete surgical excision of the tumor was performed by a median laparotomy and complete retroperitoneal dissection. In both cases, the total removal of the pheochromocytoma has been guaranteed by having margins free of tumor and a normal post-operative level of catecholamines. The pathological study revealed a malignant pheochromocytoma with margins free of neoplasia in both specimens. PMID- 11276174 TI - Cricoarytenoid joint: histological changes during aging. PMID- 11276175 TI - Hair cells in the inner ear of the pirouette and shaker 2 mutant mice. AB - The shaker 2 (sh2) and pirouette (pi) mouse mutants display severe inner ear dysfunction that involves both auditory and vestibular manifestation. Pathology of the stereocilia of hair cells has been found in both mutants. This study was designed to further our knowledge of the pathological characteristics of the inner ear sensory epithelia in both the sh2 and pi strains. Measurements of auditory brainstem responses indicated that both mutants were profoundly deaf. The morphological assays were specifically designed to characterize a pathological actin bundle that is found in both the inner hair cells and the vestibular hair cells in all five vestibular organs in these two mutants. Using light microscope analysis of phalloidin-stained specimens, these actin bundles could first be detected on postnatal day 3. As the cochleae matured, each inner hair cell and type I vestibular hair cell contained a bundle that spans from the region of the cuticular plate to the basal end of the cell, then extends along with cytoplasm and membrane, towards the basement membrane. Abnormal contact with the basement membrane was found in vestibular hair cells. Based on the shape of the cellular extension and the actin bundle that supports it, we propose to name these extensions "cytocauds." The data suggest that the cytocauds in type I vestibular hair cells and inner hair cells are associated with a failure to differentiate and detach from the basement membrane. PMID- 11276176 TI - Postnatal maturation of neuroepithelial bodies and carotid body innervation: a quantitative investigation in the rabbit. AB - The intrapulmonary airways contain oxygen-sensitive chemoreceptors which may be analogous to the arterial chemoreceptors: the neuroepithelial bodies (NEB). While the NEB are prominent in the neonatal lung, physiological studies indicate that the carotid bodies are still relatively inactive at birth. This points to an unequal degree of development of both during the early neonatal period. As a reflexogenic chemoreceptor function depends on a well-developed innervation, we undertook a comparative investigation of the development of the NEB and the carotid body glomus cell innervation. Two morphological aspects of the innervation of NEB and carotid body glomus cells were quantified in rabbits of different age groups. The total sectional area of intracorpuscular and intraglomerular nerve endings per NEB or glomus cell group, respectively, was measured and the area percentage of mitochondria and synaptic vesicles was determined. In the NEB, no significant difference in total sectional area of the nerve endings between the age groups was observed, while in the carotid body there was a significant increase in the adult age group. In addition, the area percentage of mitochondria and synaptic vesicles of the nerve endings did not change significantly with age in the NEB, while in the carotid body these increased and decreased, respectively, with age. These observations point to a shift from morphologically efferent nerve endings, rich in synaptic vesicles, to morphologically afferent nerve endings, rich in mitochondria. Our interpretation of these findings is that, at birth, the NEB innervation is more mature than the carotid body glomus cell innervation and that the latter matures at a later time than the former. These findings support the theory that the NEB may act as complementary chemoreceptors to the carotid body during the early postnatal period. PMID- 11276177 TI - Calcitonin gene-related peptide, substance P and protein gene product 9.5 immunoreactive axonal fibers in the rat footpad skin following partial sciatic nerve injuries. AB - Chronic constriction injury (CCI) and partial ligation (PSNL) of the sciatic nerve induce a similar neuropathic pain syndrome in rats. We examined calcitonin gene-related peptide (CGRP), substance P (SP) and protein gene product (PGP) 9.5 immunoreactive (IR) axons in the footpad skin after the two types of injury. Four and 14 days after CCI, CGRP- and SP-IR axons in the ipsilateral footpad skin disappeared in most rats, but in one third, sparse CGRP- and SP-IR fibers remained. PGP-IR axons dramatically decreased, but some thick fiber fascicles appeared. At the ultrastructural level, these PGP-IR thick fiber fascicles were characterized as unmyelinated axons surrounded by non-IR Schwann cells. Some of these axons were swollen and irregular in shape. In contrast, 4 days after PSNL, CGRP-, SP-, and PGP-IR axons in the ipsilateral footpad skin were present, though significantly reduced in density, in all rats, and by 14 days all IR fiber densities in the footpad skin partially recovered. The loss of CGRP and SP axons in the footpad skin of the CCI model suggests that sensory nerves containing neuropeptides are not essential in transducing stimuli applied to the footpad skin into neuropathic pain, but the abnormal PGP-IR unmyelinated axons in thick fiber fascicles might play a role. The partial loss and rapid recovery of IR axons in the footpad skin after PSNL shows that the two injury models, causing similar behaviors, are associated with very different patterns of cutaneous innervation at the time when the pain syndrome is well developed. PMID- 11276178 TI - Membrane-associated carbonic anhydrase activity in the brain of CA II-deficient mice. AB - Membrane-associated carbonic anhydrase (CA) activity is of importance for transepithelial transport of ions and fluid. Histochemical studies have indicated its presence in the brain, but the data are difficult to evaluate because of interference from cytoplasmic CA isozymes, of which CA II is the predominant one. CA II-deficient mice offer a possibility to study the location of membrane associated CA-activity, without interference from CA II. The location of CA activity in the brain of CA II-deficient and normal mice was studied by the cobalt-phosphate histochemical method, and that of CA I, CA II and CA III by an immunocytochemical method. The brains of both types of mice lacked cytoplasmic isozymes CA I and CA III, and the CA II-deficient mice also lacked CA II. In the normal mice, oligodendrocytes and choroid epithelium stained for CA II in the cytoplasm. In normal and CA (II)D-mice there was an intense membrane associated histochemical CA activity in neuronal processes. Neuronal perikarya were not stained. Endothelial membranes of brain capillaries showed strong histochemical CA-activity. Choroid epithelial cells had histochemical CA activity in the cytoplasm and along apical and baso-lateral cell membranes. The results suggest that membrane-associated CA-activity found along neuronal processes probably modulates pH of the extracellular fluid and thus neuronal activity. CA II and the membrane-associated CA of choroidal epithelium are probably involved in the secretion of cerebrospinal fluid. PMID- 11276179 TI - Topographic representation of the sciatic nerve motor neurons in the spinal cord of the adult rat correlates to region-specific activation patterns of microglia. AB - The frequent use of the adult rat sciatic nerve as a model to study the neuronal responses to injury, nerve regeneration and in transplantation studies, requires a detailed knowledge of the projection pattern of motor neurons into this nerve. Thus, as a first goal we determined this topographical projection of motor neurons and labelled small contingents by applying the fluorescent dye DiI in localised incisions made in the dorsal, rostral, ventral or caudal quadrants of the nerve. As a second goal we analysed with immunohistochemical methods the response of microglial cells within the topographical area corresponding to the incision and within areas outside this location. Uptake of the dye occurred only within the area confined to the incision, thus allowing the identification of the corresponding motor neuron perikarya within the ventral horn, eight to ten days later. In serial transverse sections of the lumbosacral spinal cord the number of labelled cells, their position within the ventral horn, and their longitudinal extent have been determined. The data suggest that the gross projection of the lumbosacral motor neuron column at the mid-thigh level of the sciatic nerve is topographic. In accordance, microglial cells showed fast activation within the injured topographic area, and a less pronounced and delayed response within the non-injured areas of the ventral horn. The graded response of microglial cells suggests that these cells possess a potential of local activation by sensing whether neurons are axotomised or just irritated by axotomy of their neighbours. The topographic organisation proves to be useful in studies on local injuries to the sciatic nerve and when analysing retrograde responses within the lumbosacral spinal cord. PMID- 11276180 TI - Schwann cells in the regenerating fish optic nerve: evidence that CNS axons, not the glia, determine when myelin formation begins. AB - Fish optic nerve fibres quickly regenerate after injury, but the onset of remyelination is delayed until they reach the brain. This recapitulates the timetable of CNS myelinogenesis during development in vertebrate animals generally, and we have used the regenerating fish optic nerve to obtain evidence that it is the axons, not the myelinating glial cells, that determine when myelin formation begins. In fish, the site of an optic nerve injury becomes remyelinated by ectopic Schwann cells of unknown origin. We allowed these cells to become established and then used them as reporters to indicate the time course of pro myelin signalling during a further round of axonal outgrowth following a second upstream lesion. Unlike in the mammalian PNS, the ectopic Schwann cells failed to respond to axotomy and to the initial outgrowth of new optic axons. They only began to divide after the axons had reached the brain. Shortly afterwards, small numbers of Schwann cells began to leave the dividing pool and form myelin sheaths. More followed gradually, so that by 3 months remyelination was almost completed and few dividing cells were left. Moreover, remyelination occurred synchronously throughout the optic nerve, with the same time course in the pre existing Schwann cells, the new ones that colonised the second injury, and the CNS oligodendrocytes elsewhere. The optic axons are the only common structures that could synchronise myelin formation in these disparate glial populations. The responses of the ectopic Schwann cells suggest that they are controlled by the regenerating optic axons in two consecutive steps. First, they begin to proliferate when the growing axons reach the brain. Second, they leave the cell cycle to differentiate individually at widely different times during the ensuing 2 months, during the critical period when the initial rough pattern of axon terminals in the optic tectum becomes refined into an accurate map. We suggest that each axon signals individually for myelin ensheathment once it completes this process. PMID- 11276181 TI - A randomized controlled trial of a passive accessory joint mobilization on acute ankle inversion sprains. AB - BACKGROUND AND PURPOSE: Passive joint mobilization is commonly used by physical therapists as an intervention for acute ankle inversion sprains. A randomized controlled trial with blinded assessors was conducted to investigate the effect of a specific joint mobilization, the anteroposterior glide on the talus, on increasing pain-free dorsiflexion and 3 gait variables: stride speed (gait speed), step length, and single support time. SUBJECTS: Forty-one subjects with acute ankle inversion sprains (<72 hours) and no other injury to the lower limb entered the trial. METHODS: Subjects were randomly assigned to 1 of 2 treatment groups. The control group received a protocol of rest, ice, compression, and elevation (RICE). The experimental group received the anteroposterior mobilization, using a force that avoided incurring any increase in pain, in addition to the RICE protocol. Subjects in both groups were treated every second day for a maximum of 2 weeks or until the discharge criteria were met, and all subjects were given a home program of continued RICE application. Outcomes were measured before and after each treatment. RESULTS: The results showed that the experimental group required fewer treatment sessions than the control group to achieve full pain-free dorsiflexion. The experimental group had greater improvement in range of movement before and after each of the first 3 treatment sessions. The experimental group also had greater increases in stride speed during the first and third treatment sessions. DISCUSSION AND CONCLUSION Addition of a talocrural mobilization to the RICE protocol in the management of ankle inversion injuries necessitated fewer treatments to achieve pain-free dorsiflexion and to improve stride speed more than RICE alone. Improvement in step length symmetry and single support time was similar in both groups. PMID- 11276182 TI - Balance and mobility following stroke: effects of physical therapy interventions with and without biofeedback/forceplate training. AB - BACKGROUND AND PURPOSE: Visual biofeedback/forceplate systems are often used for treatment of balance disorders. In this study, the researchers investigated whether the addition of visual biofeedback/forceplate training could enhance the effects of other physical therapy interventions on balance and mobility following stroke. SUBJECTS: The study included a sample of convenience of 13 outpatients with hemiplegia who ranged in age from 30 to 77 years (mean=60.4, SD=15.4) and were 15 to 538 days poststroke. METHODS: Subjects were assigned randomly to either an experimental group or a control group when the study began, and their cognitive and visual-perceptual skills were tested by a psychologist. Subjects were also assessed using the Berg Balance Scale and the Timed "Up & Go" Test before and after 4 weeks of physical therapy. Both groups received physical therapy interventions designed to improve balance and mobility 2 to 3 times per week. The experimental group trained on the NeuroCom Balance Master for 15 minutes of each 50-minute treatment session. The control group received other physical therapy for 50 minutes. RESULTS: Following intervention, both groups scored higher on the Berg Balance Scale and required less time to perform the Timed "Up & Go" Test. These improvements corresponded to increased independence of balance and mobility in the study population. However, a comparison of mean changes revealed no differences between groups. DISCUSSION AND CONCLUSION: Although both groups demonstrated improvement following 4 weeks of physical therapy interventions, no additional effects were found in the group that received visual biofeedback/forceplate training combined with other physical therapy. PMID- 11276183 TI - Effects of aerobic exercise on energy metabolism in the hypertensive rat heart. AB - BACKGROUND AND PURPOSE: In order to explore the possible effects of physical therapy interventions on people with hypertension, we evaluated the effects of aerobic exercise training on myocardial energy metabolism in an animal model of hypertension. SUBJECTS: We used 36 female spontaneously hypertensive rats (rats with genetically induced hypertension) and 12 normotensive Wistar-Kyoto rats. METHODS: The normotensive rats were sedentary and formed the CONsed group. The spontaneously hypertensive rats were randomly divided into 3 experimental groups (12 rats per group). Hypertensive rats that were sedentary formed the HTNsed group, those that received 8 weeks of exercise training formed the HTNx8 group, and those that received 16 weeks of exercise training formed the HTNx16 group. We measured systolic blood pressure, heart wet weight, maximal activities of cardiac energy metabolism enzymes, glucose transporter content, and total concentrations of protein, glycogen, and triglyceride. RESULTS: Systolic blood pressure was greater than 200 mm Hg in the CONsed group at the time of testing. Exercise training modestly (approximately 11-18 mm Hg) lowered blood pressure in the HTNx8 and HTNx16 groups. Fatty acid enzyme activity was greater in the CONsed group than in HTNsed and HTNx8 groups, but activity was roughly equivalent between the CONsed group and the HTNx16 group. Glucose enzyme activity was greater in the HTNx16 group than in the CONsed group and HTNsed group. Intracellular glycogen concentration was greater in the HTNx8 group than in HTNsed group. CONCLUSION AND DISCUSSION: Results of this study suggest that aerobic exercises may help to normalize cardiac energy metabolism in mammals with hypertension. PMID- 11276184 TI - Physical therapy management of low back pain: an exploratory survey of therapist approaches. AB - BACKGROUND AND PURPOSE: Since the release of acute low back pain management guidelines in 1994, little was known about the effect of these guidelines on clinical practice. The purpose of this study was to examine physical therapists' reported management of acute and subacute lumbar impairment. SUBJECTS: One in 10 registered physical therapists who were randomly selected from southern Ontario, Canada, (n=454) and all registered physical therapists from northern Ontario (n=331) were surveyed. METHODS: In the questionnaire, case scenarios covered 3 areas related to the management of lumbar impairment: (1) physical examination, (2) treatment and recommendations, and (3) therapists' beliefs regarding its management. RESULTS: Five hundred sixty-nine questionnaires were returned (response rate=72.5%). Only data obtained for therapists (n=274) whose weekly workload included more than 10% of people with lumbar impairment were used in the analysis. Overall, patient education, exercise, and electrotherapeutic and thermal modalities were the preferred interventions for acute lumbar impairment (symptom onset of less than 5 weeks) with or without sciatica, whereas exercise and work modification were preferred for subacute lumbar impairment (symptom onset of 5 weeks or longer). There was a trend of using electrotherapeutic and thermal modalities with uncertain effectiveness. Only 46.3% of the therapists agreed or strongly agreed that practice guidelines were useful for managing lumbar impairment. DISCUSSION AND CONCLUSION: Although the physical therapists surveyed, in general, followed the guidelines in managing acute lumbar impairment, they felt uncertain regarding the value of practice guidelines. Future research should focus on identifying effective treatment approaches and exploring the effectiveness of practice guidelines. PMID- 11276185 TI - Effect of setup configurations of split computer keyboards on wrist angle. AB - BACKGROUND AND PURPOSE: Alternative computer keyboards whose halves can be slanted toward each other can reduce a risk factor (ulnar deviation) for work related musculoskeletal disorders (WMSDs) affecting the upper limbs. Two questions that computer keyboard operators face when using keyboards that can be separated into halves (split keyboards) are: (1) At what angle should the keyboard halves be opened? and (2) At what distance apart should the keyboard halves be placed? The objective of this study was to investigate the effects of the opening angle and separation distance between halves of a split keyboard on wrist ulnar deviation and typing efficiency. METHODS: Eleven experienced computer keyboard operators participated in this study and used a split keyboard that was set up in a conventional (nonsplit) format and also in 3 alternative configurations: (1) centers of keyboard halves were separated at 20-cm distance, (2) keyboard halves were separated half of the distance of shoulder width, and (3) keyboard halves were separated at shoulder width distance. RESULTS: The 3 alternative configurations resulted in ulnar deviation of both wrists that were less than ulnar deviation from typing on a conventional setup. There were no differences in ulnar deviations among the 3 alternative configurations. DISCUSSION AND CONCLUSION: The results of this research provide physical therapists and ergonomists with a set of configurations of a split keyboard that they can recommend to their patients or clients. All of the alternative configurations of the split keyboard are beneficial in promoting a neutral wrist position, which theoretically would decrease exposure to WMSDs such as tenosynovitis in the wrist and carpal tunnel syndrome. PMID- 11276186 TI - Exercise in 94 degrees F water for a patient with multiple sclerosis. AB - BACKGROUND AND PURPOSE: The purpose of this case report is to describe the examination, intervention, and outcome of a patient with multiple sclerosis (MS) who participated in a comprehensive rehabilitation program that included aquatic therapy with a pool temperature of 94 degrees F. There are few descriptions of aquatic exercise programs on muscle force, exercise tolerance, and functional outcomes in individuals with MS, and most authors recommend a water temperature of less than 85 degrees F to prevent an exacerbation of symptoms. DESCRIPTION: The patient was a 33-year-old woman. Before, during, and after the aquatic program, she was monitored for body temperature, heart rate, blood pressure, and perceived exertion. She was also assessed for muscle force and functional abilities. OUTCOMES: The patient did not experience heat sensitivity or fatigue throughout the program, and her manual muscle test grades and mobility improved. DISCUSSION: This patient's participation in aquatic therapy, in conjunction with land-based interventions, may have been associated with the improvement in functional abilities. PMID- 11276187 TI - Myofascial trigger points. PMID- 11276188 TI - Predicting falls. PMID- 11276189 TI - Patient/client satisfaction. PMID- 11276191 TI - Trouble with peer review. PMID- 11276192 TI - CTLA-4 up-regulation plays a role in tolerance mediated by CD45. PMID- 11276193 TI - Autoimmunity versus horror autotoxicus: the struggle for recognition. PMID- 11276194 TI - Policing the regulators. PMID- 11276195 TI - Chemokines: not just leukocyte chemoattractants in the promotion of cancer. PMID- 11276196 TI - Extolling the diversity of bacterial endotoxins. PMID- 11276197 TI - Periscope, up! Monitoring microbes in the intestine. PMID- 11276199 TI - A fresh look at tumor immunosurveillance and immunotherapy. AB - Despite major advances in our understanding of adaptive immunity and dendritic cells, consistent and durable responses to cancer vaccines remain elusive and active immunotherapy is still not an established treatment modality. The key to developing an effective anti-tumor response is understanding why, initially, the immune system is unable to detect transformed cells and is subsequently tolerant of tumor growth and metastasis. Ineffective antigen presentation limits the adaptive immune response; however, we are now learning that the host's innate immune system may first fail to recognize the tumor as posing a danger. Recent descriptions of stress-induced ligands on tumor cells recognized by innate effector cells, new subsets of T cells that regulate tumor tolerance and the development of spontaneous tumors in mice that lack immune effector molecules, beckon a reflection on our current perspectives on the interaction of transformed cells with the immune system and offer new hope of stimulating therapeutic immunity to cancer. PMID- 11276200 TI - Thymic selection of CD4+CD25+ regulatory T cells induced by an agonist self peptide. AB - Despite accumulating evidence that regulatory T cells play a crucial role in preventing autoimmunity, the processes underlying their generation during immune repertoire formation are unknown. We show here that interactions with a single self-peptide can induce thymocytes that bear an autoreactive T cell receptor (TCR) to undergo selection to become CD4+CD25+ regulatory T cells. Selection of CD4+CD25+ thymocytes appears to require a TCR with high affinity for a self peptide because thymocytes that bear TCRs with low affinity do not undergo selection into this pathway. Our findings indicate that specificity for self peptides directs the selection of CD4+CD25+ regulatory thymocytes by a process that is distinct from positive selection and deletion. PMID- 11276201 TI - Defining the specific physiological requirements for c-Myc in T cell development. AB - c-Myc is associated with cell growth and cycling in many tissues and its deregulated expression is causally implicated in cancer, particularly lymphomagenesis. However, the contribution of c-Myc to lymphocyte development is unresolved. We show here that the formation of normal lymphocytes by c-Myc-/- cells is selectively defective. c-Myc-/- cells are inefficient, in an age dependent manner, at populating the thymus, and subsequent thymocyte maturation is ineffective: they fail to grow and proliferate normally at the late double negative (DN) CD4-CD8- stage. Because N-Myc expression in thymocytes usually declines at the late DN stage, these results confirm that the nonredundant contributions of Myc family members to development are related to their distinct patterns of developmental gene expression. PMID- 11276202 TI - Gene regulation mediated by calcium signals in T lymphocytes. AB - Modulation of many signaling pathways in antigen-stimulated T and B cells results in global changes in gene expression. Here we investigate the contribution of calcium signaling to gene expression in T cells using cell lines from two severe combined immunodeficiency patients with several cytokine deficiencies and diminished activation of the transcription factor NFAT nuclear factor of activated T cells. These T cells show a strong defect in transmembrane calcium influx that is also apparent in their B cells and fibroblasts. DNA microarray analysis of calcium entry-deficient and control T cells shows that Ca2+ signals both activate and repress gene expression and are largely transduced through the phosphatase calcineurin. We demonstrate an elaborate network of signaling pathways downstream of the T cell receptor, explaining the complexity of changes in gene expression during T cell activation. PMID- 11276203 TI - A point mutation in CD28 distinguishes proliferative signals from survival signals. AB - Upon interaction with its ligand, B7, CD28 becomes phosphorylated on tyrosines. One tyrosine in particular (Y170 in mouse CD28, Y173 in human CD28) has received much attention. This is because it permits CD28 to recruit SH2-containing signaling molecules, including phosphoinositide 3 kinase, Grb2 and Gads. Using mice we employed a transgenic approach to express a tyrosine-->phenylalanine mutant form of CD28 that uncouples these SH2-mediated interactions from CD28. The CD28 mutant is unable to up-regulate expression of the prosurvival protein Bcl xL, rendering the T cells more susceptible to radiation-induced death. Nonetheless, this mutated form of CD28 still prevents the induction of anergy and promotes T cell proliferation, interleukin 2 secretion and B cell help. Thus, we describe a single point mutation within the CD28 cytoplasmic domain that uncouples signals required for proliferation and survival. PMID- 11276204 TI - Extracellular matrix interacts with soluble CD95L: retention and enhancement of cytotoxicity. AB - Fas ligand (CD95L) is synthesized both on the cell surface membrane and in a soluble form. Although CD95L contributes to immune privilege in the cornea and testis, the functions of these alternatively processed proteins are not well understood. Some reports suggest that the cytotoxicity of soluble CD95L is insignificant, whereas others show potent responses in vivo, including hepatocyte apoptosis that causes liver failure. We show here that extracellular matrix proteins interact with soluble CD95L and potentiate its pro-apoptotic activity. The cytotoxicity of supernatants from CD95L-expressing cells was increased by incubation on tissue culture plates coated with these matrix proteins; this effect was mediated by trimeric soluble CD95L. With the use of immunoprecipitation, it was found that CD95L binds directly to fibronectin. In addition, immunohistochemical analysis of the cornea revealed that soluble CD95L binds primarily to extracellular matrix. The retention of soluble CD95L on extracellular matrices is likely to play an important role in the development of peripheral tolerance in immune-privileged sites. PMID- 11276205 TI - A CD14-independent LPS receptor cluster. AB - Bacterial lipopolysaccharide (LPS), the major structural component of the outer wall of Gram-negative bacteria, is a potent initiator of an inflammatory response and serves as an indicator of bacterial infection. Although CD14 has been identified as the main LPS receptor, accumulating evidence has suggested the possible existence of other functional receptor(s). In this study, using affinity chromatography, peptide mass fingerprinting and fluorescence resonance energy transfer, we have identified four new proteins that form an activation cluster after LPS ligation and are involved in LPS signal transduction. Here we present evidence that implicates heat shock proteins 70 and 90, chemokine receptor 4 and growth differentiation factor 5 as the main mediators of activation by bacterial lipopolysaccharide. PMID- 11276206 TI - Leptospiral lipopolysaccharide activates cells through a TLR2-dependent mechanism. AB - Leptospira interrogans are zoonotic pathogens that have been linked to a recent increased incidence of morbidity and mortality in highly populated tropical urban centers. They are unique among invasive spirochetes in that they contain outer membrane lipopolysaccharide (LPS) as well as lipoproteins. Here we show that both these leptospiral outer membrane constituents activate macrophages through CD14 and the Toll-like receptor 2 (TLR2). Conversely, it seems that TLR4, a central component for recognition of Gram-negative LPS, is not involved in cellular responses to L. interrogans. We also show that for intact L. interrogans, it is LPS, not lipoprotein, that constitutes the predominant signaling component for macrophages through a TLR2 pathway. These data provide a basis for understanding the innate immune response caused by leptospirosis and demonstrate a new ligand specificity for TLR2. PMID- 11276208 TI - Dendritic cells express tight junction proteins and penetrate gut epithelial monolayers to sample bacteria. AB - Penetration of the gut mucosa by pathogens expressing invasion genes is believed to occur mainly through specialized epithelial cells, called M cells, that are located in Peyer's patches. However, Salmonella typhimurium that are deficient in invasion genes encoded by Salmonella pathogenicity island 1 (SPI1) are still able to reach the spleen after oral administration. This suggests the existence of an alternative route for bacterial invasion, one that is independent of M cells. We report here a new mechanism for bacterial uptake in the mucosa tissues that is mediated by dendritic cells (DCs). DCs open the tight junctions between epithelial cells, send dendrites outside the epithelium and directly sample bacteria. In addition, because DCs express tight-junction proteins such as occludin, claudin 1 and zonula occludens 1, the integrity of the epithelial barrier is preserved. PMID- 11276207 TI - A pathological function for eotaxin and eosinophils in eosinophilic gastrointestinal inflammation. AB - Although eosinophils have been implicated in the pathogenesis of gastrointestinal disorders, their function has not been established. Using a murine model of oral antigen-induced eosinophil-associated gastrointestinal disease, we report the pathological consequences of eosinophilic inflammation and the involvement of eotaxin and eosinophils. Exposure of mice to enteric-coated antigen promotes an extensive T helper 2-associated eosinophilic inflammatory response involving the esophagus, stomach, small intestine and Peyer's patches as well as the development of gastric dysmotility, gastromegaly and cachexia. Electron microscopy shows eosinophils in proximity to damaged axons, which indicated that eosinophils were mediating a pathologic response. In addition, mice deficient in eotaxin have impaired eosinophil recruitment and are protected from gastromegaly and cachexia. These results establish a critical pathological function for eotaxin and eosinophils in gastrointestinal allergic hypersensitivity. PMID- 11276210 TI - Analyzing functional imaging studies. PMID- 11276211 TI - Are Japanese macaques threatened by neuroscience research? PMID- 11276212 TI - Are Japanese macaques threatened by neuroscience research? PMID- 11276213 TI - Putting noise into neurophysiological models of simple decision making. PMID- 11276215 TI - Color in the cortex revisited. PMID- 11276216 TI - Cdk5 at the junction. PMID- 11276217 TI - Rett syndrome model suggests MECP2 gives neurons the quiet they need to think. PMID- 11276218 TI - A wake-up call from the thalamus. PMID- 11276219 TI - MT signals: better with time. PMID- 11276221 TI - Changing specificity of neurotransmitter regulation of rapid dendritic remodeling during synaptogenesis. AB - Wiring the developing nervous system requires appropriate contact between presynaptic axons and postsynaptic dendrites. Rapid movements of filopodia-like structures on immature dendrites are thought to facilitate initial synaptogenic contact with axons. Here we show that not only can different forms of neurotransmission regulate dendritic filopodial motility, but they do so in a developmentally regulated manner, suggestive of a specific relationship between the action of a neurotransmitter and the corresponding type of synapse being formed. PMID- 11276222 TI - Early development of a somatosensory fovea: a head start in the cortical space race? AB - Star-nosed moles have 11 mechanosensory appendages surrounding each nostril, and primary afferents from a single appendage-the tactile fovea-are greatly over represented in somatosensory cortex. It was found that the foveal appendage led development in the periphery, had the greatest innervated surface area in embryos, and developed mature nerve terminals and epidermal sensory organs first; also, in developing cortex, markers for metabolic activity (cytochrome oxidase) appeared first in the fovea representation. This developmental sequence may provide the fovea with an advantage in a competition for cortical space, and account for the much larger areas of cortex devoted to foveal afferents. PMID- 11276223 TI - Spatial coding of enantiomers in the rat olfactory bulb. AB - Because of their unique properties, enantiomers (pairs of mirror-symmetric, nonsuperimposable molecules that differ only in optical activity and their interaction with other chiral molecules) have been instrumental in demonstrating that olfactory perception relies on molecular shape. To investigate how molecular structure is encoded by the olfactory system, we combined behavioral discrimination tasks with optical imaging of intrinsic signals. We found that rats can behaviorally discriminate members of a wide range of enantiomer pairs, and imaging revealed enantiomer-selective glomeruli in the olfactory bulb, indicating that the spatial pattern of glomerular activity provides sufficient information to discriminate molecular shape. PMID- 11276225 TI - How do dendrites take their shape? AB - Recent technical advances have made possible the visualization and genetic manipulation of individual dendritic trees. These studies have led to the identification and characterization of molecules that are important for different aspects of dendritic development. Although much remains to be learned, the existing knowledge has allowed us to take initial steps toward a comprehensive understanding of how complex dendritic trees are built. In this review, we describe recent advances in our understanding of the molecular mechanisms underlying dendritic morphogenesis, and discuss their cell-biological implications. PMID- 11276224 TI - Altered neuroadaptation in opiate dependence and neurogenic inflammatory nociception in alpha CGRP-deficient mice. AB - The neuropeptide alpha CGRP (calcitonin gene-related peptide) is involved in the complex process of pain signaling, but the precise contribution of alpha CGRP remains unclear. Here we show that mice lacking alpha CGRP display an attenuated response to chemical pain and inflammation. Furthermore, alpha CGRP(-/-) mice do not show changes in heroin self-administration or morphine tolerance, but display a marked decrease in morphine withdrawal signs, suggesting an important contribution of alpha CGRP to opiate withdrawal. PMID- 11276226 TI - Phosphorylation of cofilin by LIM-kinase is necessary for semaphorin 3A-induced growth cone collapse. AB - Semaphorin 3A is a chemorepulsive axonal guidance molecule that depolymerizes the actin cytoskeleton and collapses growth cones of dorsal root ganglia neurons. Here we investigate the role of LIM-kinase 1, which phosphorylates an actin depolymerizing protein, cofilin, in semaphorin 3A-induced growth cone collapse. Semaphorin 3A induced phosphorylation and dephosphorylation of cofilin at growth cones sequentially. A synthetic cell-permeable peptide containing a cofilin phosphorylation site inhibited LIM-kinase in vitro and in vivo, and essentially suppressed semaphorin 3A-induced growth cone collapse. A dominant-negative LIM kinase, which could not be activated by PAK or ROCK, suppressed the collapsing activity of semaphorin 3A. Phosphorylation of cofilin by LIM-kinase may be a critical signaling event in growth cone collapse by semaphorin 3A. PMID- 11276227 TI - Cdk5 is involved in neuregulin-induced AChR expression at the neuromuscular junction. AB - Here we describe an important involvement of Cdk5/p35 in regulating the gene expression of acetylcholine receptor (AChR) at the neuromuscular synapse. Cdk5 and p35 were prominently expressed in embryonic muscle, and concentrated at the neuromuscular junction in adulthood. Neuregulin increased the p35-associated Cdk5 kinase activity in the membrane fraction of cultured C2C12 myotubes. Co immunoprecipitation studies revealed the association between Cdk5, p35 and ErbB receptors in muscle and cultured myotubes. Inhibition of Cdk5 activity not only blocked the NRG-induced AChR transcription, but also attenuated ErbB activation in cultured myotubes. In light of our finding that overexpression of p35 alone led to an increase in AChR promoter activity in muscle, Cdk5 activation is sufficient to mediate the up-regulation of AChR gene expression. Taken together, these results reveal the unexpected involvement of Cdk5/p35 in neuregulin signaling at the neuromuscular synapse. PMID- 11276228 TI - Protein kinase C modulates NMDA receptor trafficking and gating. AB - Regulation of neuronal N-methyl-D-aspartate receptors (NMDARs) by protein kinases is critical in synaptic transmission. However, the molecular mechanisms underlying protein kinase C (PKC) potentiation of NMDARs are uncertain. Here we demonstrate that PKC increases NMDA channel opening rate and delivers new NMDA channels to the plasma membrane through regulated exocytosis. PKC induced a rapid delivery of functional NMDARs to the cell surface and increased surface NR1 immunofluorescence in Xenopus oocytes expressing NMDARs. PKC potentiation was inhibited by botulinum neurotoxin A and a dominant negative mutant of soluble NSF associated protein (SNAP-25), suggesting that receptor trafficking occurs via SNARE-dependent exocytosis. In neurons, PKC induced a rapid delivery of functional NMDARs, assessed by electrophysiology, and an increase in NMDAR clusters on the surface of dendrites and dendritic spines, as indicated by immunofluorescence. Thus, PKC regulates NMDAR channel gating and trafficking in recombinant systems and in neurons, mechanisms that may be relevant to synaptic plasticity. PMID- 11276229 TI - Morphological correlates of functionally defined synaptic vesicle populations. AB - By combining photoconversion of FM1-43-stained vesicles and electron microscopy of hippocampal synapses, we find evidence that the population of morphologically docked synaptic vesicles corresponds to the release-ready neurotransmitter quanta. Furthermore, those synaptic vesicles that are participating in cycles of exo- and endocytosis tend to be closer to the active zone than vesicles that are being held in reserve. PMID- 11276230 TI - A neural code for low-frequency sound localization in mammals. AB - We report a systematic relationship between sound-frequency tuning and sensitivity to interaural time delays for neurons in the midbrain nucleus of the inferior colliculus; neurons with relatively low best frequencies (BFs) showed response peaks at long delays, whereas neurons with relatively high BFs showed response peaks at short delays. The consequence of this relationship is that the steepest region of the function relating discharge rate to interaural time delay (ITD) fell close to midline for all neurons irrespective of BF. These data provide support for a processing of the output of coincidence detectors subserving low-frequency sound localization in which the location of a sound source is determined by the activity in two broad, hemispheric spatial channels, rather than numerous channels tuned to discrete spatial positions. PMID- 11276231 TI - The impact of 'bursting' thalamic impulses at a neocortical synapse. AB - Considerable effort has gone into understanding the mechanisms underlying high frequency 'bursting' of thalamocortical impulses, their sensory information content and their involvement in perception. However, little is known about the influence of such impulses on their cortical targets. Here we follow bursting thalamic impulses to their terminus at the thalamocortical synapse of the awake rabbit, and examine their influence on a class of somatosensory cortical neurons. We show that thalamic bursts potently activate cortical circuits. Initial impulses of each burst have a greatly enhanced ability to elicit cortical action potentials, and later impulses in the burst further raise the probability of eliciting spikes. In some cases, multiple cortical spikes result from a single burst. Moreover, we show that the interval preceding each burst is crucial for generating the enhanced cortical response. The powerful activation of neocortex by thalamocortical bursts is fully consistent with an involvement of these impulses in perceptual/attentional processes. PMID- 11276232 TI - The spatial transformation of color in the primary visual cortex of the macaque monkey. AB - Perceptually, color is used to discriminate objects by hue and to identify color boundaries. The primate retina and the lateral geniculate nucleus (LGN) have cell populations sensitive to color modulation, but the role of the primary visual cortex (V1) in color signal processing is uncertain. We re-evaluated color processing in V1 by studying single-neuron responses to luminance and to equiluminant color patterns equated for cone contrast. Many neurons respond robustly to both equiluminant color and luminance modulation (color-luminance cells). Also, there are neurons that prefer luminance (luminance cells), and a few neurons that prefer color (color cells). Surprisingly, most color-luminance cells are spatial-frequency tuned, with approximately equal selectivity for chromatic and achromatic patterns. Therefore, V1 retains the color sensitivity provided by the LGN, and adds spatial selectivity for color boundaries. PMID- 11276233 TI - Peripheral group I metabotropic glutamate receptors modulate nociception in mice. AB - The metabotropic glutamate receptors (mGluRs) are found throughout the central nervous system, where they modulate neuronal excitability and synaptic transmission. Here we report the presence of phospholipase C-coupled group I mGluRs (mGluR1 and mGluR5) outside the central nervous system on peripheral unmyelinated sensory afferents. Given their localization on predominantly nociceptive afferents, we investigated whether these receptors modulate nociceptive signaling, and found that agonist-induced activation of peripheral group I mGluRs leads to increased sensitivity to noxious heat, a phenomenon termed thermal hyperalgesia. Furthermore, group I mGluR antagonists not only prevent, but also attenuate established formalin-induced pain. Taken together, these results suggest that peripheral mGluRs mediate a component of hyperalgesia and may be therapeutically targeted to prevent and treat inflammatory pain. PMID- 11276234 TI - Thalamocortical NMDA conductances and intracortical inhibition can explain cortical temporal tuning. AB - Cells in cerebral cortex fail to respond to fast-moving stimuli that evoke strong responses in the thalamic nuclei innervating the cortex. The reason for this behavior has remained a mystery. We study an experimentally motivated model of the thalamic input-recipient layer of cat primary visual cortex that accounts for many aspects of cortical orientation tuning. In this circuit, inhibition dominates over excitation, but temporal modulations of excitation and inhibition occur out of phase with one another, allowing excitation to transiently drive cells. We show that this circuit provides a natural explanation of cortical low pass temporal frequency tuning, provided N-methyl-D-aspartate (NMDA) receptors are present in thalamocortical synapses in proportions measured experimentally. This suggests a new and unanticipated role for NMDA conductances in shaping the temporal response properties of cortical cells, and suggests that common cortical circuit mechanisms underlie both spatial and temporal response tuning. PMID- 11276235 TI - Intracortical origin of visual maps. AB - Previous experiments indicate that the shape of maps of preferred orientation in the primary visual cortex does not depend on visual experience. We propose a network model that demonstrates that the orientation and direction selectivity of individual units and the structure of the corresponding angle maps could emerge from local recurrent connections. Our model reproduces the structure of preferred orientation and direction maps, and explains the origin of their interrelationship. The model also provides an explanation for the correlation between position shifts of receptive fields and changes of preferred orientations of single neurons across the surface of the cortex. PMID- 11276236 TI - Activation of the left amygdala to a cognitive representation of fear. AB - We examined the neural substrates involved when subjects encountered an event linked verbally, but not experientially, to an aversive outcome. This instructed fear task models a primary way humans learn about the emotional nature of events. Subjects were told that one stimulus (threat) represents an aversive event (a shock may be given), whereas another (safe) represents safety (no shock will be given). Using functional magnetic resonance imaging (fMRI), activation of the left amygdala was observed in response to threat versus safe conditions, which correlated with the expression of the fear response as measured by skin conductance. Additional activation observed in the insular cortex is proposed to be involved in conveying a cortical representation of fear to the amygdala. These results suggest that the neural substrates that support conditioned fear across species have a similar but somewhat different role in more abstract representations of fear in humans. PMID- 11276237 TI - Circuit mechanisms underlying memory encoding and retrieval in the long axis of the hippocampal formation. AB - Circuits within the hippocampal formation are active during memory processing. Here we used functional magnetic resonance imaging (fMRI) to examine multiple sites across the long axis of the hippocampal formation while subjects performed different phases of an associative memory task, learning to associate faces with names. Viewing faces and hearing names in isolation resulted in separate hippocampal activation patterns. Pairing faces with names resulted a spatially redistributed activation pattern, rather than a simple summation of the activation patterns resulting from viewing faces and hearing names in isolation. Recalling names when cued with faces reactivated a pattern similar to that found during paired training. Finally, the activation patterns representing faces and names were found to be experience dependent, emerging with repeated exposure. Interpreted in the context of hippocampal anatomy and physiology, these findings reveal hippocampal circuit mechanisms that underlie memory encoding and retrieval. PMID- 11276238 TI - More than just mad cow disease. PMID- 11276239 TI - Trading places. PMID- 11276240 TI - Barreling through the outer membrane. PMID- 11276241 TI - Solving the FYVE domain--PtdIns(3)P puzzle. PMID- 11276242 TI - Making the most of metal ions. PMID- 11276243 TI - Unraveling the means to the end in ATP-dependent proteases. PMID- 11276244 TI - Cdc13 subcomplexes regulate multiple telomere functions. PMID- 11276245 TI - How jumping genes were discovered. PMID- 11276246 TI - Picture story. The separator. PMID- 11276247 TI - Comparative architecture of transposase and integrase complexes. AB - Transposases and retroviral integrases promote the movement of DNA segments to new locations within and between genomes. These recombinases function as multimeric protein-DNA complexes. Recent success in solving the crystal structure of a Tn5 transposase--DNA complex provides the first detailed structural information about a member of the transposase/integrase superfamily in its active, DNA-bound state. Here, we summarize the reactions catalyzed by transposases and integrases and review the Tn5 transposase-DNA co-crystal structure. The insights gained from the Tn5 structure and other available structures are considered together with biochemical and genetic data to discuss features that are likely to prove common to the catalytic complexes used by members of this important protein family. PMID- 11276248 TI - The neuronal t-SNARE complex is a parallel four-helix bundle. AB - Assembly of the soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) complex is an essential step for neurotransmitter release in synapses. The presynaptic plasma membrane associated proteins (t-SNAREs), SNAP-25 (synaptosome-associated protein of 25,000 Da) and syntaxin 1A may form an intermediate complex that later binds to vesicle-associated membrane protein 2 (VAMP2). Using spin labeling electron paramagnetic resonance (EPR), we found that the two t-SNARE proteins assemble into a parallel four-helix bundle that consists of two identical syntaxin 1A components and the N-terminal and C-terminal domains of SNAP-25. Although the structure is generally similar to that of the final SNARE complex, the middle region of the helical bundle appears more flexible in the t-SNARE complex. Such flexibility might facilitate interactions between VAMP2 and the t-SNARE complex. PMID- 11276249 TI - The homing endonuclease I-CreI uses three metals, one of which is shared between the two active sites. AB - Homing endonucleases, like restriction enzymes, cleave double-stranded DNA at specific target sites. The cleavage mechanism(s) utilized by LAGLIDADG endonucleases have been difficult to elucidate; their active sites are divergent, and only one low resolution cocrystal structure has been determined. Here we report two high resolution structures of the dimeric I-CreI homing endonuclease bound to DNA: a substrate complex with calcium and a product complex with magnesium. The bound metals in both complexes are verified by manganese anomalous difference maps. The active sites are positioned close together to facilitate cleavage across the DNA minor groove; each contains one metal ion bound between a conserved aspartate (Asp 20) and a single scissile phosphate. A third metal ion bridges the two active sites. This divalent cation is bound between aspartate residues from the active site of each subunit and is in simultaneous contact with the scissile phosphates of both DNA strands. A metal-bound water molecule acts as the nucleophile and is part of an extensive network of ordered water molecules that are positioned by enzyme side chains. These structures illustrate a unique variant of a two-metal endonuclease mechanism is employed by the highly divergent LAGLIDADG enzyme family. PMID- 11276250 TI - Human cystatin C, an amyloidogenic protein, dimerizes through three-dimensional domain swapping. AB - The crystal structure of human cystatin C, a protein with amyloidogenic properties and a potent inhibitor of cysteine proteases, reveals how the protein refolds to produce very tight two-fold symmetric dimers while retaining the secondary structure of the monomeric form. The dimerization occurs through three dimensional domain swapping, a mechanism for forming oligomeric proteins. The reconstituted monomer-like domains are similar to chicken cystatin except for one inhibitory loop that unfolds to form the 'open interface' of the dimer. The structure explains the tendency of human cystatin C to dimerize and suggests a mechanism for its aggregation in the brain arteries of elderly people with amyloid angiopathy. A more severe 'conformational disease' is associated with the L68Q mutant of human cystatin C, which causes massive amyloidosis, cerebral hemorrhage and death in young adults. The structure of the three-dimensional domain-swapped dimers shows how the L68Q mutation destabilizes the monomers and makes the partially unfolded intermediate less unstable. Higher aggregates may arise through the three-dimensional domain-swapping mechanism occurring in an open-ended fashion in which partially unfolded molecules are linked into infinite chains. PMID- 11276251 TI - Substrate-induced activation of a trapped IMC-mediated protein folding intermediate. AB - While several unfolded proteins acquire native structures through distinct folding intermediates, the physiological relevance and importance of such states in the folding kinetics remain controversial. The intramolecular chaperone (IMC) of subtilisin was used to trap a partially folded, stable crosslinked intermediate conformer (CLIC) through a disulfide bond between mutated IMC and subtilisin. The trapped CLIC contains non-native interactions. Here we show that CLIC can be induced into a catalytically active form by incubating it with small peptide substrates. The structure and catalytic properties of the activated crosslinked intermediate conformer (A-CLIC) differ from those of the fully folded enzyme in that A-CLIC lacks any endopeptidase activity toward a large protein substrate. Our results show that a disulfide-linked partially folded protein can be induced to acquire catalytic activity with a substrate specificity that is different from completely folded subtilisin. These results also suggest that protein folding intermediates may also participate in catalytic reactions. PMID- 11276252 TI - Phi-values for BPTI folding intermediates and implications for transition state analysis. AB - Amino acid replacements were used to probe the roles of 14 sites in two well characterized intermediates in the folding pathway of bovine pancreatic trypsin inhibitor (BPTI). One of these intermediates contains one of the three disulfides found in the native protein (30--51). NMR studies have shown that approximately two-thirds of this polypeptide has a native-like conformation. The other intermediate contains two native disulfides (30--51 and 5--55) and has a fully folded conformation. The phi-values for a majority of residues were <1, indicating that the native protein was significantly more destabilized than either intermediate even when the altered residue was located in a well-ordered region of the intermediate. These observations suggest that folding intermediates and transition states may generally be more structured than indicated by phi values alone. PMID- 11276253 TI - Kinetics of unfolding and folding from amide hydrogen exchange in native ubiquitin. AB - Amide hydrogen (NH) exchange is one of the few experimental techniques with the potential for determining the thermodynamics and kinetics of conformational motions at nearly every residue in native proteins. Quantitative interpretation of NH exchange in terms of molecular motions relies on a simple two-state kinetic model: at any given slowly exchanging NH, a closed or exchange-incompetent conformation is in equilibrium with an open or exchange-competent conformation. Previous studies have demonstrated the accuracy of this model in measuring conformational equilibria by comparing exchange data with the thermodynamics of protein unfolding. We report here a test of the accuracy of the model in determining the kinetics of conformational changes in native proteins. The kinetics of folding and unfolding for ubiquitin have been measured by conventional methods and compared with those derived from a comprehensive analysis of the pH dependence of exchange in native ubiquitin. Rate constants for folding and unfolding from these two very different types of experiments show good agreement. The simple model for NH exchange thus appears to be a robust framework for obtaining quantitative information about molecular motions in native proteins. PMID- 11276254 TI - Structure of outer membrane protein A transmembrane domain by NMR spectroscopy. AB - We have determined the three-dimensional fold of the 19 kDa (177 residues) transmembrane domain of the outer membrane protein A of Escherichia coli in dodecylphosphocholine (DPC) micelles in solution using heteronuclear NMR. The structure consists of an eight-stranded beta-barrel connected by tight turns on the periplasmic side and larger mobile loops on the extracellular side. The solution structure of the barrel in DPC micelles is similar to that in n octyltetraoxyethylene (C(8)E(4)) micelles determined by X-ray diffraction. Moreover, data from NMR dynamic experiments reveal a gradient of conformational flexibility in the structure that may contribute to the membrane channel function of this protein. PMID- 11276255 TI - A universal mode of helix packing in RNA. AB - RNA molecules fold into specific three-dimensional shapes to perform structural and catalytic functions. Large RNAs can form compact globular structures, but the chemical basis for close helical packing within these molecules has been unclear. Analysis of transfer, catalysis, in vitro-selected and ribosomal RNAs reveal that helical packing predominantly involves the interaction of single-stranded adenosines with a helix minor groove. Using the Tetrahymena thermophila group I ribozyme, we show here that the near-perfect shape complementarity between the adenine base and the minor groove allows for optimal van der Waals contacts, extensive hydrogen bonding and hydrophobic surface burial, creating a highly energetically favorable interaction. Adenosine is recognized in a chemically similar fashion by a combination of protein and RNA components in the ribonucleoprotein core of the signal recognition particle. These results provide a thermodynamic explanation for the noted abundance of conserved adenosines within the unpaired regions of RNA secondary structures. PMID- 11276256 TI - Simultaneous binding of two proteins to opposite sides of a single transfer RNA. AB - Transfer RNA (tRNA) is a small nucleic acid (typically 76 nucleotides) that forms binary complexes with proteins, such as aminoacyl tRNA synthetases (RS) and Trbp111. The latter is a widely distributed structure-specific tRNA-binding protein that is incorporated into cell signaling molecules. The structure of Trbp111 was modeled onto to the outer, convex side of the L-shaped tRNA. Here we present RNA footprints that are consistent with this model. This binding mode is in contrast to that of tRNA synthetases, which bind to the inside, or concave side, of tRNA. These opposite locations of binding for these two proteins suggest the possibility of a ternary complex. The formation of a tRNA synthetase--tRNA- Trbp111 ternary complex was detected by two independent methods. The results indicate that the tRNA is sandwiched between the two protein molecules. A thermodynamic and functional analysis is consistent with the tRNA retaining its native structure in the ternary complex. These results may have implications for how the translation apparatus is linked to other cellular machinery. PMID- 11276257 TI - Structural analysis of BAG1 cochaperone and its interactions with Hsc70 heat shock protein. AB - BAG-family proteins share a conserved protein interaction region, called the 'BAG domain', which binds and regulates Hsp70/Hsc70 molecular chaperones. This family of cochaperones functionally regulates signal transducing proteins and transcription factors important for cell stress responses, apoptosis, proliferation, cell migration and hormone action. Aberrant overexpression of the founding member of this family, BAG1, occurs in human cancers. In this study, a structure-based approach was used to identify interacting residues in a BAG1- Hsc70 complex. An Hsc70-binding fragment of BAG1 was shown by multidimensional NMR methods to consist of an antiparallel three-helix bundle. NMR chemical shift experiments marked surface residues on the second (alpha 2) and third (alpha 3) helices in the BAG domain that are involved in chaperone binding. Structural predictions were confirmed by site-directed mutagenesis of these residues, resulting in loss of binding of BAG1 to Hsc70 in vitro and in cells. Molecular docking of BAG1 to Hsc70 and mutagenesis of Hsc70 marked the molecular surface of the ATPase domain necessary for interaction with BAG1. The results provide a structural basis for understanding the mechanism by which BAG proteins link molecular chaperones and cell signaling pathways. PMID- 11276258 TI - Nucleotide binding by the histidine kinase CheA. AB - To probe the structural basis for protein histidine kinase (PHK) catalytic activity and the prospects for PHK-specific inhibitor design, we report the crystal structures for the nucleotide binding domain of Thermotoga maritima CheA with ADP and three ATP analogs (ADPNP, ADPCP and TNP-ATP) bound with either Mg(2+) or Mn(2+). The conformation of ADPNP bound to CheA and related ATPases differs from that reported in the ADPNP complex of PHK EnvZ. Interactions of the active site with the nucleotide gamma-phosphate and its associated Mg(2+) ion are linked to conformational changes in an ATP-lid that could mediate recognition of the substrate domain. The inhibitor TNP-ATP binds CheA with its phosphates in a nonproductive conformation and its adenine and trinitrophenyl groups in two adjacent binding pockets. The trinitrophenyl interaction may be exploited for designing CheA-targeted drugs that would not interfere with host ATPases. PMID- 11276259 TI - Multistep assembly of the protein import channel of the mitochondrial outer membrane. AB - Proteins targeted to mitochondria are transported into the organelle through a high molecular weight complex called the translocase of the outer mitochondrial membrane (TOM). At the core of this machinery is a multisubunit general import pore (GIP) of 400 kDa. Here we report the assembly of the yeast GIP that involves two successive intermediates of 250 kDa and 100 kDa. The precursor of the channel lining Tom40 is first targeted to the membrane via the receptor proteins Tom20 and Tom22; it then assembles with Tom5 to form the 250 kDa intermediate exposed to the intermembrane space. The 250 kDa intermediate is followed by the formation of the 100 kDa intermediate that associates with Tom6. Maturation to the 400 kDa complex occurs by association of Tom7 and Tom22. Tom7 functions by promoting both the dissociation of the 400 kDa complex and the transition from the 100 kDa intermediate to the mature complex. These results indicate that the dynamic conversion between the 400 kDa complex and the 100 kDa late intermediate allows integration of new precursor subunits into pre-existing complexes. PMID- 11276260 TI - The leukemia-associated AML1 (Runx1)--CBF beta complex functions as a DNA-induced molecular clamp. AB - We have determined the structure, at 2.6 A resolution, of the AML1 (Runx1) Runt domain--CBF beta--DNA ternary complex, the most common target for mutations in human leukemia. The structure reveals that the Runt domain DNA binding mechanism is unique within the p53 family of transcription factors. The extended C-terminal 'tail' and 'wing' elements adopt a specific DNA-bound conformation that clamps the phosphate backbone between the major and minor grooves of the distorted B form DNA recognition site. Furthermore, the extended 'tail' mediates most of the NF-kappa B/Rel-like base-specific contacts in the major groove. The structure clearly explains the molecular basis for the loss of DNA binding function of the Runt domain--CBF beta complex as a consequence of the human disease-associated mutations in leukemogenesis and cleidocranial dysplasia. PMID- 11276261 TI - Occupational safety and health in small and medium-sized enterprises during social and economic transformation. AB - Small and medium-sized enterprises (SMEs) in Central and Eastern Europe at the turn of the 1980s and 1990s were characterised by many people exposed to hazardous working conditions. Statistics recorded a considerable increase in the number of occupational diseases. They also showed that it was more dangerous to work for a small company. The transition from planned to market economy has brought about an increase in the number of SMEs in Poland. Data on them are provided. The role of the Central Institute for Labour Protection in studying SMEs is presented. A pilot study of employers and employees is discussed. PMID- 11276262 TI - Anthropometry for design for the elderly. AB - This paper presents anthropometric data on elderly people in Australia. Data were collected in the metropolitan city of Sydney, NSW, Australia. In all 171 elderly people (males and females, aged 65 years and above) took part in the study. Mean values, standard deviations, medians, range, and coefficients of variation for the various body dimensions were estimated. Correlation coefficients were also calculated to determine the relationship between different body dimensions for the elderly population. The mean stature of elderly Australian males and females were compared with populations from other countries. The paper discusses design implications for elderly people and provides several examples of application of the anthropometric data. PMID- 11276263 TI - Interactions of some organic solvents: hydrocarbons and chloroalkene. AB - Metabolic and toxicodynamic interactions of some organic solvents in rats repeatedly treated with medium dose levels were examined. It was shown that both n-hexane and ethylbenzene significantly inhibited tetrachloroethylene metabolism during a 2-week period. n-Hexane and tetrachloroethylene enhanced metabolism of ethylbenzene whereas ethylbenzene suppressed n-hexane metabolism only at the end of the experiment. Biochemical changes, especially the drop in the level of non protein sulfhydryl groups in tissues of rats treated with organic solvent mixtures, were significantly less pronounced than those observed after these chemicals were administered separately. These results demonstrate that metabolic interactions between hydrocarbons and chloroalkene may lead to a modification of the biological response to these compounds. PMID- 11276264 TI - Users' demands regarding dental safety glasses. Combining a quantitative approach and grounded theory for the data analysis. AB - Eye infections are common among dentists and many are concerned, but few are using proper eye protection. To understand users' demands behind the low use of safety glasses, all dental teams in Sweden were asked which factors they found most important when choosing dental safety glasses, and rate the importance of 31 statements regarding ergonomic aspects of dental safety glasses in a questionnaire. Data were analysed using the Grounded Theory and a quantitative approach. Results showed that dentists ranked the visual aspects as most important and chair assistants the protective aspects. The highly visual demanding work performed by dentists requires safety glasses that are not yet available on the market, which might explain the low use. PMID- 11276265 TI - Effects of neutral posture on muscle tension during computer use. AB - This study focused on developing a new approach to seated work positions was conducted on 67 office workers who use a Visual Display Terminal (VDT) as a major function of their working day. Muscle tension was measured by surface electromyography (sEMG) while participants were asked to adopt 4 selected working postures. Pain was measured before and after ergonomic intervention on the Nordic scale, which was modified for this study. Adjustable workstations were used to place participants in desired positions during the clinical testing sessions and the extended intervention period. Results indicate the effects of this ergonomic intervention may have positive effects on muscle tension and pain, significant enough to encourage employers to implement training and workstation modifications following these guidelines. PMID- 11276266 TI - Identification of driver model parameters. AB - The paper presents a driver model, which can be used in a computer simulation of a curved ride of a car. The identification of the driver parameters consisted in a comparison of the results of computer calculations obtained for the driver vehicle-environment model with different driver data sets with test results of the double lane-change manoeuvre (Standard No. ISO/TR 3888:1975, International Organization for Standardization [ISO], 1975) and the wind gust manoeuvre. The optimisation method allows to choose for each real driver a set of driver model parameters for which the differences between test and calculation results are smallest. The presented driver model can be used in investigating the driver vehicle control system, which allows to adapt the car construction to the psychophysical characteristics of a driver. PMID- 11276267 TI - Slip resistance of industrial floor surfaces: development of an elastomer suited to in-situ measurement. AB - Slips contribute to 12% of occupational accidents. A slip resistant floor is a mean to prevent slipping accidents occurring in workshops. Floor slip resistance is often evaluated by measuring a friction index, proportional to the force opposing slipping of a reference elastomer on the floor surface under test. When implementing a portable appliance, slip resistance measurements carried out on lubricated floors were not stabilized. The authors advanced the hypothesis of oil impregnating the elastomer. A new elastomer suited to in-situ measurement has been developed to achieve stable measuring conditions. This study highlights the fact that the nature and characteristics of a reference elastomer must be specified when slip resistance measurements are carried out. PMID- 11276268 TI - Effects of Ergorest arm supports on muscle strain and wrist positions during the use of the mouse and keyboard in work with visual display units: a work site intervention. AB - The effects of Ergorest arm supports on wrist angles and musculoskeletal strain in the neck-shoulder-arm region and electrical activity in the shoulder and arm muscles were studied during typing or the use of the mouse in work with a visual display unit (VDU). Twenty-one women were randomized into 3 groups (1 arm support, 2 arm supports, and control). Measurements were carried out before and after the 6-week intervention. The wrist extension of the mouse hand, the muscle activity of the trapezius muscle, and the subjective discomfort ratings indicated that 2 arm supports were better than 1 in work with a mouse. The Ergorest arm support alleviates muscle and joint strain in VDU work when used for both arms. PMID- 11276270 TI - Use of verapamil in the treatment of diarrhea due to microscopic colitis. PMID- 11276271 TI - Therapeutic options in nonulcer dyspepsia. AB - Dyspepsia, defined as pain or discomfort centered in the upper abdomen, affects an estimated 25% of the U.S. population each year; accounts for up to 5% of all visits to primary care physicians, and generates over $1.3 billion in prescription drug costs annually. In the majority of patients evaluated, no clear cause of symptoms can be identified, and the condition is termed functional or nonulcer dyspepsia (NUD). The pathophysiology of NUD remains unclear, but disturbances in gastrointestinal motility or sensation are often found. Clinically, NUD can be subdivided into dysmotility-like (in which discomfort, fullness, bloating, early satiety, or nausea [but not pain] predominate) or ulcer like (in which epigastric pain is predominant). In ulcer-like NUD, antisecretory therapy is useful, but in dysmotility-like NUD, acid suppression is not superior to placebo. Cisapride accelerates gastric emptying and enhances gastric accommodation but probably does not blunt perception. Although cisapride relieves symptoms of dyspepsia without the adverse central nervous system effects often associated with metoclopramide, its cardiac toxicity has led to disuse. Antidepressants are of uncertain efficacy but are widely used. New prokinetics and other enteric neuromodulating agents are being tested in NUD and are likely to find an important place in clinical practice in the future. PMID- 11276272 TI - Increased numbers of women, older individuals, and Blacks receive health care for dyspepsia in the United States. AB - GOALS: The objectives of this research were to use a national probability sample of the U.S. population to determine the demographic characteristics of individuals who obtained care for dyspepsia, to compare these demographic characteristics with those of the U.S. population, and to describe the amount of health care that these individuals received. STUDY: We analyzed data from the 1987 National Medical Expenditure Survey, which is based on a national probability sample of the U.S. adult population. RESULTS: Approximately 3.6 million individuals, or 2% of U.S. adults, obtained care for dyspepsia. Compared with the U.S. population, a predominance of women, individuals 65 years or older, and African Americans obtained care for dyspepsia. Expenditures for health care totaled $2.5 billion. CONCLUSIONS: Given the major impact of dyspepsia on U.S. health care resources, a critical issue facing investigators is to identify the most cost-effective approach to managing these patients. PMID- 11276273 TI - Treatment of elderly patients with nabumetone or diclofenac: gastrointestinal safety profile. AB - GOALS: To evaluate the efficacy and gastrointestinal safety of nabumetone and diclofenac in the treatment of elderly patients with osteoarthritis, participating in a 3-month efficacy trial. BACKGROUND: Elderly patients have an elevated risk for developing gastrointestinal complications with chronic use of nonsteroidal anti-inflammatory drugs. STUDY: This was a randomized, double-blind, parallel-group, multicenter study with a 3-to 14-day placebo washout period and a 12-week active treatment phase. Patients 65 years or older with moderate-to severe osteoarthritis of the knee or hip were randomized to receive 1,000 to 2,000 mg/d of nabumetone or 100 to 150 mg/d of diclofenac. The primary efficacy parameters were the percent of patients improved using a Patients' and Physicians' Global Assessment at endpoint. Gastrointestinal safety was assessed by the incidence of gastrointestinal symptoms and adverse events. RESULTS: Three hundred thirty-five patients (mean age = 72 years) with active osteoarthritis were enrolled. There were no statistically significant differences between the two treatment groups in any of the primary efficacy variables. However, differences between the groups were apparent in the frequency of adverse gastrointestinal events. Two patients in the diclofenac group had evidence of gastrointestinal bleeding and/or ulcer compared with none in the nabumetone group. In addition, significantly more (p < 0.05) patients (4%) receiving diclofenac had alanine transaminase values more than twice the upper limit of normal at endpoint compared with patients receiving nabumetone (0%). CONCLUSIONS: Nabumetone was as effective as diclofenac in the treatment of elderly patients with moderate-to-severe osteoarthritis. However, the gastrointestinal safety profile of nabumetone was superior to that of diclofenac with respect to elevation of liver enzymes. PMID- 11276274 TI - Prognostic value of fibrinolytic tests for hospital outcome in patients with acute upper gastrointestinal hemorrhage. AB - GOALS: We assessed the predictive value of fibrinolytic tests for hospital outcome in a prospective study of 84 nonconsecutive patients with acute upper gastrointestinal hemorrhage. STUDY: Six readily available parameters of activated fibrinolysis (fibrinogen, D-dimer, tissue plasminogen activator [TPA], plasminogen activator inhibitor type 1 [PAI-1], TPA--PAI-1 complexes, and plasmin alpha 2-antiplasmin complexes) were tested for association with hospital outcome. Patients were divided into the following three groups: patients who survived and did not require transfusion or surgery, those who survived without surgery but required transfusion, and those who required surgery or died. RESULTS: Patients with adverse outcome (surgery and/or death) showed significantly higher plasma levels of D-dimer than patients with favorable outcome (p = 0.01). Plasma concentrations of D-dimer >300 ng/mL showed a 20.5% positive predictive value of adverse outcome, with a relative risk of 7.5 (95% CI: 1--57%). Patients who required transfusion showed significantly higher plasma levels of TPA (p = 0.01). A positive correlation between endoscopic bleeding stigmata and D-dimer in the subgroup of patients without liver cirrhosis was found (p = 0.02); however, in the multivariate logistic regression analysis the concentration of D-dimer did not appear as an independent predictor of adverse outcome. CONCLUSIONS: These findings are consistent with the role of increased local fibrinolysis in the digestive tract, particularly of D-dimer, in patients with upper gastrointestinal hemorrhage and adverse outcome. Accordingly, plasma fibrinolytic tests may constitute an appropriate prognostic marker in upper gastrointestinal bleeding. PMID- 11276275 TI - The exacerbation of pancreatic endocrine dysfunction by potent pancreatic exocrine supplements in patients with chronic pancreatitis. AB - BACKGROUND: Chronic pancreatitis often culminates in maldigestion and diabetes. Clinical management is complex as the correction of maldigestion often disturbs diabetic control. STUDY: In the following study, we examined the effects of a potent new commercial pancreatic enzyme on food absorption and blood glucose control. Enzymes were manufactured in enteric-coated mini-microsphere form (0.7- 1.6 mm), designed to prevent gastric acid degradation and facilitate co-migration with food, and given in quantities calculated to cover normal digestion requirements (four capsules with meals, two with snacks; content/capsule: lipase 10,000 USP units, protease 37,500 units, amylase 33,200 units). Forty patients with chronic pancreatitis were screened during a run-in nonenzyme-supplemented phase; only those with stool fat excretion rates over 10 g/d (n = 29) were advanced to a 14-day parallel randomized placebo versus enzyme supplement group comparison. RESULTS: Of these, 62% were diabetic (50% insulin-dependent) and 52% were malnourished (body mass index less than 20 kg/m(2) ). After enzyme supplementation, stool fat and nitrogen excretion decreased, whereas fat absorption increased from 54.0 +/- 9.7% to 80.8 +/- 3.8% per day (p = 0.002) and protein from 80.5 +/- 3.4% to 86.8 +/- 2.2% per day (p = 0.004). Changing treatment from active enzyme supplementation to placebo (and vice versa) resulted in major problems with glucose control; blood glucose levels became abnormal in 28 of 29 patients, one patient required hospitalization for symptomatic hypoglycemia (0.9 mmol/L) during placebo treatment, and one developed diabetic ketoacidosis after recommencing active enzyme supplementation. CONCLUSIONS: In conclusion, high-dose pancreatin mini-microspheres improved, but did not normalize, fat absorption, possibly because of the residual influence of diabetes and malnutrition on absorptive function. In view of the brittle nature of blood glucose control in malnourished insulin-dependent patients, enzyme adjustment should be carefully supervised in-hospital. PMID- 11276276 TI - Long-term prospective study of the effect of ursodeoxycholic acid on cystic fibrosis-related liver disease. AB - GOALS: To evaluate the efficacy of UDCA in arresting the progression of the early multifocal hepatic lesion to overt CF-related NBC. BACKGROUND: Focal biliary cirrhosis is an early hepatic pathologic change related to the ion transport defect in cystic fibrosis. The factors involved in the progression of focal to nodular biliary cirrhosis are not clear. Ursodeoxycholic--a hydrophilic, nontoxic, choleretic, and hepatoprotective exogenous bile acid--has been reported to be effective in the management of cholestatic liver disease. STUDY: For 10 years at 6-month intervals, 70 individuals with cystic fibrosis (38 men and 32 women; age range, 2--29 years) were examined using hepatosplenomegaly, liver function tests, and ultrasound liver scan. Patients demonstrating evidence of liver involvement at the onset or during the study received ursodeoxycholic acid 20 mg/kg body weight. RESULTS: After the administration of ursodeoxycholic acid, the progression of nodular biliary cirrhosis ultrasound changes was arrested, hepatic function was preserved, and no variceal bleeding was observed. No case of focal biliary cirrhosis progressed to nodular biliary cirrhosis. Furthermore, the multifocal, multilobular changes suggestive of focal biliary cirrhosis on ultrasound scan were reversed to normal. CONCLUSION: Ursodeoxycholic acid is effective in improving cholestasis and hepatic dysfunction in nodular biliary cirrhosis and, also, in reversing the early sonography findings suggestive of focal biliary cirrhosis. It is speculated that ursodeoxycholic acid may prove to affect the natural history of cystic fibrosis-related liver disease. PMID- 11276277 TI - Clinical significance of enlarged perihepatic lymph nodes in chronic hepatitis B. AB - GOALS: To evaluate the clinical significance of enlarged perihepatic lymph nodes in patients with chronic hepatitis B. BACKGROUND: Enlargement of perihepatic lymph node is a common finding during ultrasonography in patients with chronic hepatitis. Its relation with liver histology and viremia was reported in chronic hepatitis C, but little has been known about its clinical significance in chronic hepatitis B. STUDY: We evaluated the clinical significance of perihepatic lymphadenopathy in chronic hepatitis B. In 50 patients with biopsy-proven chronic hepatitis B and 15 healthy controls, the perihepatic lymph node volume was evaluated by ultrasonography and its possible correlation with biochemical tests, hepatitis activity index, and hepatitis B viremia was investigated. RESULTS: Perihepatic lymph node was detected in 48 of 50 patients with chronic hepatitis B (volume = 3.4 +/- 2.4 mL) and in two of 15 controls (0.4 mL and 0.6 mL). In chronic hepatitis B, lymph node volume showed a significant correlation with serum aspartate transaminase (r = 0.66), alanine transaminase (r = 0.63), gamma glutamyl-transpeptidase (r = 0.53), histologic activity index (r = 0.59), and necroinflammatory score (r = 0.59; p < 0.05 for all), but not with fibrosis score and serum hepatitis B viremia. CONCLUSIONS: Enlarged perihepatic lymph nodes in chronic hepatitis B can be a good indicator for histologic and biochemical inflammatory activity of the liver, but not for viremia. PMID- 11276279 TI - Endoscopic treatment of acute gastric volvulus causing cardiac tamponade. AB - Acute gastric volvulus occurs when the stomach, or part of the stomach, rotates more than 180 degrees, creating a closed-loop obstruction, which eventually leads to ischemia and strangulation. Acute gastric volvulus may occur in association with a diaphragmatic defect, diaphragmatic elevation of any cause, tumors of the pancreas and stomach, trauma, and congenital abnormalities of mesenteric fixation. We describe an unusual case of an acute gastric volvulus causing cardiac tamponade, which was successfully treated by endoscopic reduction of the gastric volvulus. PMID- 11276278 TI - Intraabdominal vancomycin-resistant enterococcus infections: the new threat. AB - GOALS: To determine the clinical course and outcome in patients with intraabdominal vancomycin-resistant enterococcus infections (VRE-A) and to identify probable risk factors for VRE-A. BACKGROUND: Vancomycin-resistant enterococcus is one of the most notable nosocomial emerging pathogens. The incidence is increasing, especially in the abdominal surgery setting. STUDY: A comparative study of patients with VRE-A and VRE infection in other sites (VRE-O) who were hospitalized for over 1 year. Fisher exact test and Student t test were used; a two-tailed p value of less than 0.05 was considered to be significant. RESULTS: Of 89 nine patients with VRE, six had VRE-A, 24 had VRE-O, and 59 had VRE colonization. The VRE-A group was comprised of one patient with an inoperable Klatskin tumor and biliary sepsis, one with acquired immune deficiency syndrome and an infected pancreatic pseudocyst, two with fecal peritonitis, and two with biliary sepsis after surgery for common bile duct stones. All six patients with VRE-A had recent surgery before VRE isolation, as compared with three in the VRE O group (p = 0.0001). Despite adequate treatment with intravenous chloramphenicol, resulting in eradication of VRE in all six VRE-A cases, the mortality rate remained high at 50%. CONCLUSIONS: Vancomycin-resistant enterococcus should be recognized as an emerging nosocomial pathogen that causes potentially fatal intraabdominal infections in the postsurgical setting. However, the impact of treatment on ultimate outcome needs further evaluation. PMID- 11276280 TI - Ornithine transcarbamylase deficiency unmasked because of gastrointestinal bleeding. AB - Ornithine transcarbamylase (OTC) is a mitochondrial-matrix enzyme that catalyzes conversion of ornithine and carbamyl phosphate to citrulline, the second step in the urea cycle. The urea cycle is the most important pathway to detoxification of ammonia in human beings. Ornithine transcarbamylase deficiency (OTCD) is the most common urea cycle disorder, inherited as an X-linked disorder that can cause fatal hyperammonemia in male newborns. Women with OTCD have a variable expression of their disease, the variability being determined by lyonization (random inactivation) of the X chromosome. We report a case of a 28-year-old woman who presented with hyperammonemic encephalopathy that was precipitated by a gastrointestinal bleed unmasking OTCD. PMID- 11276281 TI - Diaphragm disease of the small bowel: a case without apparent nonsteroidal antiinflammatory drug use. AB - BACKGROUND: Diaphragm-like strictures of the small bowel are an infrequent complication of the treatment of patients with nonsteroidal antiinflammatory drugs (NSAIDs). STUDY: We report a patient with this condition in whom the use of NSAIDs was ruled out by both clinical history and objective blood testing of current aspirin use. RESULTS: He reported a history of recurrent episodes of colic abdominal pain during the past 25 years; he underwent three surgical operations for this condition. Before these symptoms, he had an undefined abdominal process with diarrhea, weight loss, and diffuse edema, which resolved spontaneously without reaching a diagnosis. CONCLUSIONS: We suggest that diaphragm-like strictures might be developed as a nonspecific response to different damaging insults to the intestine and are not necessarily associated with NSAID use. PMID- 11276282 TI - Lupus abdominal crisis owing to rupture of an ileocolic aneurysm with successful angiographic treatment. AB - There are many causes of acute abdominal pain, or abdominal "crises," in patients with systemic lupus erythematosus (SLE), most frequently the causes are serositis or vasculitis. Vasculitis generally causes small vessel abnormalities and may present with symptoms owing to mucosal damage, such as pain, diarrhea, or bleeding. We present a patient with SLE who had the acute onset of severe abdominal pain while hospitalized for a lupus flare and who was found to have a ruptured ileocolic aneurysm with intraperitoneal bleeding. She was successfully managed with angiographic embolization, without further complications. Although angiography is well established as a therapeutic intervention for mesenteric aneurysms of various etiologies, this is the first case of an SLE-related ileocolic aneurysm so managed. This entity should be considered in the differential diagnosis of abdominal pain in patients with lupus, and angiographic embolization should be considered in its management. PMID- 11276283 TI - Use of verapamil for the symptomatic treatment of microscopic colitis. AB - BACKGROUND: The initial use of verapamil for the symptomatic treatment of microscopic colitis is reported. STUDY: Four patients diagnosed with microscopic colitis who failed to respond to conventional therapy are presented. RESULTS: Verapamil was instituted, resulting in prompt and sustained resolution of symptoms. CONCLUSIONS: This therapy was convenient and well tolerated. PMID- 11276284 TI - Endoscopic management of a duodenal duplication cyst associated with biliary obstruction in an adult. AB - Duodenal duplication cysts are distinctly uncommon and most often present in infancy or early childhood. The clinical presentation is generally duodenal obstruction, hemorrhage, or pancreatitis. Duodenal duplication cysts rarely cause biliary obstruction in adults. So far, duodenal duplication cysts have been almost exclusively treated by surgical intervention. This report describes both endoscopic diagnosis and treatment of a large periampullary duodenal duplication cyst associated with biliary obstruction in an adult patient. PMID- 11276285 TI - Lipiodol accumulation in focal peliosis hepatis with sinusoidal dilatation. AB - Peliosis hepatis is a rare benign condition that is histologically characterized by multiple cystic blood-filled spaces in the liver. Although the cause is unknown, the condition occurs in association with several diseases or medications. We report a patient who was found to have a lesion with lipiodol accumulation in the liver 2 months after its intraarterial injection. The lesion was diagnosed and treated as a small hepatocellular carcinoma. However, subsequent right hepatic lobectomy and histologic examination confirmed the diagnosis of focal peliosis hepatis. PMID- 11276286 TI - Antiphospholipid syndrome presenting as portopulmonary hypertension. AB - The association of pulmonary hypertension with portal hypertension, also called portopulmonary hypertension, is a well-described condition. The pathogenesis of this association remains unclear. We describe a 34-year-old female patient with "primary antiphospholipid syndrome" and portopulmonary hypertension. Our finding supports that in situ microthrombosis associated with the presence of anticardiolipin antibodies could be the pathophysiologic explanation for both portal and pulmonary hypertension. PMID- 11276287 TI - Spontaneous fungal peritonitis (Candida glabrata) in a patient with cirrhosis. AB - We report a case of spontaneous fungal peritonitis in a patient with cirrhosis. A 70-year-old woman with cirrhosis secondary to autoimmune hepatitis was admitted with fever and abdominal distention. Paracentesis revealed neutrocytosis, and despite appropriate antibacterial coverage, no clinical improvement was noted and the ascitic fluid white cell count increased on repeat paracentesis. Two consecutive ascitic fluid cultures grew Candida glabrata, and antifungal therapy with amphotericin was initiated, pending sensitivity of the isolate. Because of worsening renal function, amphotericin was discontinued and itraconazole was started, as sensitivity of the isolate was then available. Antifungal therapy resulted in resolution of ascitic fluid neutrocytosis and culture negativity. However, the patient's renal function continued to deteriorate, necessitating hemodialysis. Despite multiple courses of antibiotics, she died of fulminant sepsis and multiorgan failure. PMID- 11276288 TI - Recurrent pneumonia from an ileobronchial fistula complicating Crohn's disease. AB - We report the case of a patient with Crohn's disease and recurrent pneumonia for over 3 years before the discovery of an occult ileopulmonary fistula and review five other cases in the literature. Patients often present with chronic cough productive of feculent sputum, pleuritic chest pain, and signs of pulmonary consolidation that fail to respond completely to antibiotic therapy. Mixed enteric flora is cultured from sputum and bronchial washings in most cases. Bronchoscopy findings range from chronic bronchial inflammation to feculent material in the airways. Barium enema is often diagnostic. Surgery and Crohn's specific therapy are key components of curative therapy. PMID- 11276289 TI - Pseudotumor of the bladder as a manifestation of uncomplicated appendicitis. AB - We report the case of a 67-year-old woman who presented with an apparent bladder tumor and complaints of increased urinary frequency and postvoiding pain. Her diagnostic work-up revealed conflicting findings regarding the source of her bladder disease, and laparoscopy was required to make the final diagnosis of appendicitis. The presentation of appendicitis can be disguised and is often a difficult diagnosis. PMID- 11276290 TI - Co-occurrence of hepatocellular carcinoma and lymphoma in patients with hepatitis C virus cirrhosis. AB - The association of hepatitis C virus (HCV) with neoplasia is not completely understood. Hepatitis C virus is hepatolymphotrophic. It is considered an inducing factor of hepatocellular carcinoma (HCC) and is associated with various types of lymphomas. We describe a patient with HCV cirrhosis who developed gastric lymphoma and HCC, and we review the current data and theories about the oncogenesis of HCV. PMID- 11276291 TI - The Efficacy and Safety of Medicinal Herbs International Conference. 2-3 March 2000, Chapel Hill, North Carolina, USA. PMID- 11276292 TI - Herbal medicine: from the past to the future. AB - A brief discussion of the history of the use of herbal medicines from prehistoric times to the mid-twentieth century precedes an explanation of why usage of such remedies in the United States declined in the 1940s but returned to popularity in the 1980s. The provisions of the Dietary Supplement Health and Education Act of 1994 are presented together with its perceived influence, both positive and negative, on the health of the American people. Possible futures of herbal medicines are considered. The negative viewpoint that they will ultimately be rejected is refuted, and the more optimistic prediction that herbs are ultimately destined to become a part of mainstream medicine is defended. Stumbling blocks to such acceptance are evaluated and methods of overcoming them suggested. The urgent need for the development of a sensible regulatory environment encouraging the approval of botanicals as drugs is emphasized. After predicting a bright future for rational phytomedicines, the author opines that many of them will eventually play significant roles in medicinal practice. PMID- 11276293 TI - Epidemiologic challenges in the study of the efficacy and safety of medicinal herbs. AB - Although clinical trials are needed to prove the efficacy of medicinal herbs and pharmacological studies are essential to the long-term goal of identifying the active ingredients in plants, these will not be forthcoming rapidly enough to meet the acute public health needs for knowledge on efficacy and safety since these substances are currently being widely consumed at various dosages. Resulting from the ongoing 'natural experiment' well-conducted observational epidemiology can bridge the gap and determine whether, as consumed its use is of benefit or detrimental, for whom and when in the course of disease prevention or minimization of disease severity. The classic study designs (cohort and case control) and the more recent development of case-only studies can be put to service for these purposes. The challenges are in dose assessment, understanding mechanisms of effect, determining the relevant time period of exposure for a given disease or symptom, controlling for confounding factors such as disease status, and the special challenges presented by irregular use of medicinal herbs and concurrent use of multiple products and multiple sources. PMID- 11276294 TI - Phytotherapy for benign prostatic hyperplasia. AB - OBJECTIVE: To systematically review the existing evidence regarding the efficacy and safety of phytotherapeutic compounds used to treat men with symptomatic benign prostatic hyperplasia (BPH). DESIGN: Randomized trials were identified searching MEDLINE (1966--1997), EMBASE, Phytodok, the Cochrane Library, bibliographies of identified trials and review articles, and contact with relevant authors and drug companies. The studies were included if men had symptomatic benign prostatic hyperplasia, the intervention was a phytotherapeutic preparation alone or combined, a control group received placebo or other pharmacologic therapies for BPH, and the treatment duration was at least 30 days. Key data were extracted independently by two investigators. RESULTS: A total of 44 studies of six phytotherapeutic agents (Serenoa repens, Hypoxis rooperi, Secale cereale, Pygeum africanum, Urtica dioica, Curcubita pepo) met inclusion criteria and were reviewed. Many studies did not report results in a method allowing meta-analysis. Serenoa repens, extracted from the saw palmetto, is the most widely used phytotherapeutic agent for BPH. A total of 18 trials involving 2939 men were reviewed. Compared with men receiving placebo, men taking Serenoa repens reported greater improvement of urinary tract symptoms and flow measures. Serenoa repens decreased nocturia (weighted mean difference (WMD) = -0.76 times per evening; 95% CI = -1.22 to -0.32; n = 10 studies) and improved peak urine flow (WMD = 1.93 ml s(-1); 95% CI = 0.72 to 3.14, n = 8 studies). Men treated with Serenoa repens rated greater improvement of their urinary tract symptoms versus men taking placebo (risk ratio of improvement = 1.72; 95% CI = 1.21 to 2.44, n = 8 studies). Improvement in symptoms of BPH was comparable to men receiving the finasteride. Hypoxis rooperi (n = 4 studies, 519 men) was also demonstrated to be effective in improving symptom scores and flow measures compared with placebo. For the two studies reporting the International Prostate Symptom Score, the WMD was -4.9 IPSS points (95% CI = -6.3 to -3.5, n = 2 studies) and the WMD for peak urine flow was 3.91 ml s(-1) (95% CI = 0.91 to 6.90, n = 4 studies). Secale cereale (n = 4 studies, 444 men) was found to modestly improve overall urological symptoms. Pygeum africanum (n = 17 studies, 900 men) may be a useful treatment option for BPH. However, review of the literature has found inadequate reporting of outcomes which currently limit the ability to estimate its safety and efficacy. The studies involving Urtica dioica and Curcubita pepo are limited although these agents may be effective combined with other plant extracts such as Serenoa and Pygeum. Adverse events due to phytotherapies were reported to be generally mild and infrequent. CONCLUSIONS: Randomized studies of Serenoa repens, alone or in combination with other plant extracts, have provided the strongest evidence for efficacy and tolerability in treatment of BPH in comparison with other phytotherapies. Serenoa repens appears to be a useful option for improving lower urinary tract symptoms and flow measures. Hypoxis rooperi and Secale cereale also appear to improve BPH symptoms although the evidence is less strong for these products. Pygeum africanum has been studied extensively but inadequate reporting of outcomes limits the ability to conclusively recommend it. There is no convincing evidence supporting the use of Urtica dioica or Curcubita pepo alone for treatment of BPH. Overall, phytotherapies are less costly, well tolerated and adverse events are generally mild and infrequent. Future randomized controlled trials using standardized preparations of phytotherapeutic agents with longer study durations are needed to determine their long-term effectiveness in the treatment of BPH. PMID- 11276295 TI - Efficacy and safety of ginseng. AB - Ginseng (Panax ginseng, C.A. Meyer) has been a popular herbal remedy used in eastern Asian cultures for thousands of years. In North America, the ginseng species indigenous to both Canada and the United States (Panax quinquefolium) represents an important industry for both domestic and export markets. There are numerous theories and claims describing the efficacy of ginseng, which can combat stress, enhance both the central and immune systems and contribute towards maintaining optimal oxidative status against certain chronic disease states and aging. Risk issues concerning the safety of ginseng at recommended dosages are less prominent and scientifically based. While some epidemiological or clinical studies have reported indications of efficacy for specific health benefits or potential toxicity, there are an equal number of studies that provide contradictory evidence. This situation has led to questionable conclusions concerning specific health benefits or risks associated with ginseng. Recent advances in the development of standardized extracts for both Panax ginseng (G 115) and Panax quinquefolius (CNT-2000) have and will continue to assist in the assessment of efficacy and safety standards for ginseng products. This paper reviews the scientific literature and evidence for ginseng efficacy and safety derived mostly from in vitro and animal studies and places emphasis on the need for more randomized, double-blinded, placebo clinical studies that can provide unequivocal conclusions. An example of the efficacy and safety of ginseng is provided with the description of biological activity of a North American ginseng extract (NAGE), which includes illustrating mechanisms for antioxidant activity without prooxidant properties. PMID- 11276296 TI - Efficacy and safety of St. John's wort for the treatment of major depression. AB - OBJECTIVE: Extracts of St. John's wort have been widely used in the treatment of depression. Our aim was to review information related to the efficacy and safety of St. John's wort as an antidepressant. DATA SOURCES: Primary and review articles were identified by a search of Medline (1960 to February 2000) and through secondary sources. STUDY SELECTION: All the articles identified from the data sources were evaluated and all relevant information was included in this review. The pharmacokinetics, mechanism of action, efficacy, side effects and drug interactions of St. John's wort have been examined in various studies. CONCLUSION: St. John's wort is a promising investigational antidepressant, but the data are not yet sufficient to accept hypericum as a first line antidepressant preparation for treatment of depression. Besides the need for dose standardization and adequate trial lengths, there is a need for studies in severely depressed patients and long-term studies to assess the risk of relapse and recurrence. PMID- 11276297 TI - Efficacy and safety of a Ginkgo biloba extract. AB - This review of the literature documents the efficacy of a standard extract of Ginkgo biloba (EGb) in managing signs and symptoms associated with memory disorders and dementia. Analysis of the discrepant findings reveals that study outcomes may vary with the type of population studied, the outcome measurements selected, and the dosing tested. Overall, the efficacy of EGb was more frequently reported in trials enrolling dementia patients than healthy volunteers. In contrast to narrow memory tests, broad cognitive assessments were more likely to detect the treatment effect. Although a dose--response relationship is not yet established, 240 mg day(-1) EGb seems to show a higher rate of treatment response than does 120 mg day(-1). Regarding safety, in all trials reviewed the adverse event profile of EGb was not different from that of the placebo. PMID- 11276298 TI - The efficacy and safety of comfrey. AB - Herbal medication has gathered increasing recognition in recent years with regard to both treatment options and health hazards. Pyrrolizidine alkaloids have been associated with substantial toxicity after their ingestion as tea and in the setting of contaminated cereals have led to endemic outbreaks in Jamaica, India and Afghanistan. In Western Europe, comfrey has been applied for inflammatory disorders such as arthritis, thrombophlebitis and gout and as a treatment for diarrhoea. Only recently was the use of comfrey leaves recognized as a substantial health hazard with hepatic toxicity in humans and carcinogenic potential in rodents. These effects are most likely due to various hepatotoxic pyrrolizidine alkaloids such as lasiocarpine and symphytine, and their related N oxides. The mechanisms by which toxicity and mutagenicity are conveyed are still not fully understood, but seem to be mediated through a toxic mechanism related to the biotransformation of alkaloids by hepatic microsomal enzymes. This produces highly reactive pyrroles which act as powerful alkylating agents. The main liver injury caused by comfrey (Symphytum officinale) is veno-occlusive disease, a non-thrombotic obliteration of small hepatic veins leading to cirrhosis and eventually liver failure. Patients may present with either acute or chronic clinical signs with portal hypertension, hepatomegaly and abdominal pain as the main features. Therapeutic approaches include avoiding intake and, if hepatic failure is imminent, liver transplantation. In view of the known serious hazards and the ban on distributing comfrey in Germany and Canada, it is difficult to understand why comfrey is still freely available in the United States. PMID- 11276299 TI - The efficacy and safety of feverfew (Tanacetum parthenium L.): an update of a systematic review. AB - OBJECTIVE: Feverfew (Tanacetum parthenium L.) is a popular herbal remedy often advocated for the prevention of migraine. The aims of this systematic review are to update the evidence from rigorous clinical trials for or against the efficacy of feverfew for migraine prevention and to provide a safety profile of this herbal remedy. DESIGN: Literature searches were performed using the following databases: Medline, Embase, Biosis, CISCOM and the Cochrane Library (all from their inception to December 1999). Only randomized, placebo-controlled, double blind trials of feverfew mono-preparations for the prevention of migraine in human subjects were included. All articles were read by two independent reviewers. Data were extracted in a pre-defined, standardized fashion. The methodological quality of the trials was evaluated by the Jadad score. For the assessment of safety issues, major reference texts were also consulted. RESULTS: Six trials met the inclusion/exclusion criteria. The majority favour feverfew over placebo. Yet important caveats exist. The data also suggest that feverfew is associated with only mild and transient adverse effects and few other safety concerns. CONCLUSIONS: Feverfew is likely to be effective in the prevention of migraine. There are no major safety problems. PMID- 11276300 TI - Research and future trends in the pharmaceutical development of medicinal herbs from Chinese medicine. AB - Issues concerning the past and future development of medicinal herbs from Chinese medicine (CM) are addressed in this paper. In the Western world, medicinal herbs are becoming increasingly popular and important in the public and scientific communities. In contrast to their regulated status in China and other countries, herbal medicines are regarded as dietary supplements in the US. Accordingly, research must continue worldwide to identify and improve the efficacy of the active principals of herbs both singly and in combination -- from active ingredients, active fractions, and active herbal formulations. While Western medicine currently employs pure, single compounds, either natural or synthetic, CM has long used multiple combinations of compounds in the form of processed natural products, primarily medicinal herbs, to treat and relieve the symptoms of many different human diseases. CM may have fewer and less severe side effects than single pure drugs, making CM especially attractive to the consumer. In effect, CM's focus on combination therapy does serve both ancient and modern theories. However, research using modern analytical and chemical techniques is needed to ensure efficacy and safety, to provide qualitative and quantitative analyses for dietary supplements, and to develop new, effective and safe world class drugs. Drug design is an iterative process. Bioactivity-directed fractionation and isolation identify active natural compounds from single herbs or formulations. These lead structures can be chemically modified and improved through knowledge of structure--activity relationship, mechanism of action, drug metabolism, molecular modelling and combinatorial chemistry studies. Finally, efficacy and toxicity determination as well as clinical trials can contribute to the generation of new drugs from CM. To continue the legacy of CM, as well as the worldwide uses of other medicinal herbs, continued investigation of active formulations, bioactive fractions, and isolated compounds is critical to drug development in the 21st century. PMID- 11276301 TI - alpha-Fetoprotein-induced apoptosis of cancer cells. PMID- 11276302 TI - New aspects of heparin effects. AB - The effects of a 5-day heparin treatment (10 kD, 64 IU/kg, intraperitoneally) on food-procuring behavior and spatial memory in a 12-arm radial maze were studies on Wistar rats. The maximum reinforcement scores in heparinized rats were attained by day 7 and in control rats only by day 16. In total, 75% heparinized and 45% control rats successfully learned the task for 24 days. On day 25 the contents of major transmitters and their metabolites in various brain structures and in the small intestine of control and experimental rats were determined. The rats treated with heparin showed increased concentrations of norepinephrine in the hypothalamus, homovanillic acid in the striatum, and serotonin in the small intestine. Our findings indicate that heparin exhibits a wide range of activities in addition to its anticoagulant effect. PMID- 11276303 TI - Role of thyroid hormones in stress-induced synthesis of heat-shock proteins in the myocardium. AB - Thyroxine in near-physiological doses increased the content of heat-shock proteins in the myocardium and stimulated their accumulation during immobilization stress. Blockade of thyroid functions with methimazole decreased the content of heat-shock proteins in rat myocardium during stress and heat shock and prevented their accumulation during adaptation to short-term immobilizations. PMID- 11276304 TI - Effect of physical training on sulfamethazine acetylation rate. AB - We studied the rate of sulfamethazine acetylation in athletes and untrained controls aging 18-22 years. The rate of sulfamethazine acetylation in controls was characterized by a bimodal distribution: rapid and slow acetylators constituted 42 and 58%, respectively. The rate of sulfamethazine biotransformation in athletes was characterized by a trimodal distribution: ultrarapid, rapid, and slow xenobiotic acetylators constituted 48.4, 22.6, and 29%, respectively. The ultrarapid acetylation phenotype was probably associated with N-acetyltransferase induction and reflected adaptation to physical exercises PMID- 11276305 TI - Effect of endothelin-1 on DNA synthesis in the myocardium of albino rats during early postnatal ontogeny. AB - Effect of intraperitoneal injection of endothelin-1 on DNA synthesis in the myocardium of newborn albino rats was studied by(3)H-thymidine autoradiography. Endothelin-1 injected in a single dose of 10 microg/kg stimulated proliferative processes: the index of labeled nuclei and labeling intensity increased. Repeated (5 times) administration of endothelin-1 in doses of 1 and 10 microg/kg increased labeling intensity, but did not change the index of labeled nuclei. The data suggest that endothelins are involved in morphogenesis of the myocardium during the early postnatal ontogeny. PMID- 11276306 TI - Thiol-selective mechanism of HIV antigen conjugation with serum IgM, IgG, and IgA. AB - The effects of HIV antigen glycoproteins gp160, gp41, and gp36 on thiol-dependent specific (affinity) binding of serum IgM, IgG, and IgA with the corresponding antigens were determined by the formation of signal thiol-containing analytes (free nonprotein SH groups). Free nonprotein SH groups were not found in the reaction mixture, which indicated that HIV antigen glycoproteins blocked this process. The results suggest that the thiol-selective mechanism underlies in vitro conjugation of HIV antigen glycoproteins gp160, gp41, and gp36 with serum immunoglobulins. Previous studies showed that this mechanism of conjugation with immunoglobulins is not characteristic of other lymphotropic viruses (e.g., hepatitis B virus). PMID- 11276307 TI - Effect of bioactive additive mammoleptin on development of transplanted tumors in mice. AB - Bioactive additive mammoleptin used for the treatment of fibrocystic breast disease did not stimulate the growth of primary tumors and metastases in mice with transplanted tumors. Mammoleptin in high doses inhibited the growth of Ehrlich adenocarcinoma and metastatic spreading of solid tumors. PMID- 11276308 TI - Biokinetics of transdermal therapeutic medicinal form of phenazepam. AB - We studied the rate of phenazepam absorption into the blood and its transport to the brain from a transdermal therapeutic system and bioavailability of the drug in this system. Hydrogel matrix consisting of polyvinyl alcohol and 1,2-propylene glycol was used for application. Transdermal application of 0.1-0.4 mg phenazepam in a dose of 14 mg/kg provided a stable level of this drug during application interval (1-48 h), while its bioavailability for blood plasma and brain was 0.63 and 0.2, respectively (determined for 0.4 mg phenazepam). The rate of drug penetration into the blood and brain was 46 and 60 ng/ml/h, respectively. PMID- 11276309 TI - Effect of knotweed extract on alkeran-induced changes in rat ejaculate. AB - We studied the effects of aqueous knotweed extracts in alkeran-induced experimental pathozoospermia. Therapeutic effect of knotweed extract in experimental cytostatic hypogonadism was demonstrated (the preparation improved spermatozoon motility). PMID- 11276310 TI - Regulation of functional activity of bone marrow hemopoietic stem cells by erythroid cells in mice. AB - Transplantation of erythroid and bone marrow cells to irradiated mice stimulated exogenous colony formation. Pretreatment of erythroid cells with specific rabbit antiserum to erythroblasts abolished this effect. The reverse transcriptase polymerase chain reaction revealed the presence of mRNA for interleukin-1alpha, interleukin-1beta, interleukin-3, interleukin-6, and granulocyte-macrophage colony-stimulating factor in erythroid cells. Granulocyte-macrophage colony stimulating factor was found in the conditioned medium from erythroid cells. Thus, erythroid cells stimulated colony-forming activity of bone marrow cells, which was probably mediated via cytokine synthesis (e.g., granulocyte-macrophage colony-stimulating factor). PMID- 11276312 TI - Expression of phosphatidylinositol-3 kinase in lung cancer. AB - The expression of phosphatidylinositol-3 kinase in tumors and homologous tissues from 29 patients with lung cancer, 5 patients with lung metastases of various tumors, and some non-tumorous pulmonary diseases was studied by Western blot analysis. The expression of phosphatidylinositol-3 kinase was increased in these tumors in comparison with histologically intact lung tissue in 5 patients with non-small-cell cancer. In 20 patients expression of phosphatidylinositol-3 kinase was the same as in homologous tissue and in 4 patients it was decreased. No relationship between phosphatidylinositol-3 kinase expression and clinical and morphological characteristics of lung cancer was revealed. PMID- 11276311 TI - Effect of toluene on bioamine-containing structures in the spleen. AB - The effect of toluene administered into the stomach on amino-containing structures in the spleen of random-bred albino mice was studied. It was shown by Falck-Hillarp method that 6 h after treatment the toxicant stimulated splenic mast cell population and inhibited other amino-containing structures. It is therefore suggested that in control mice the major role in bioamine metabolism in the spleen is played by granular fluorescent cells, while after poisoning, mast cells functioning as adapters acquire the primary role. The levels of catecholamines and serotonin in nervous and nonnervous structures peaked 1 week after poisoning and returned to normal after 4 weeks. Presumably, toluene suppresses the immunity starting from the second week after treatment. PMID- 11276313 TI - Regulating effect of epithalone on gastric endocrine cells in pinealectomized rats. AB - Endocrine cells in the stomach of pinealectomized rats after injection of epithalone (pineal gland peptide) were studied by immunohistochemical tests, morphometry, and image analysis microscopic images. A functional relationship was found between the pineal gland and stomach, which is regulated by peptides produced by the pineal gland. PMID- 11276314 TI - Effect of tetrapeptide cortagen on regeneration of sciatic nerve. AB - Intramuscular injection of 10 microg/kg cortagen to rats during 10 days after transsection and suturing of the sciatic nerve increased the growth rate and conduction velocity in the regenerating nerve fibers by 27% and 40%, respectively. PMID- 11276315 TI - Peptide bioregulators inhibit apoptosis. AB - The effects of peptide bioregulators epithalon and vilon on the dynamics of irradiation-induced apoptotic death of spleen lymphocytes in rats indicate that these agents inhibit physiologically programmed cell death. The antiapoptotic effect of vilon was more pronounced, which corroborates the concept on tissue specific effect of peptide bioregulators. PMID- 11276316 TI - Effect of cold stress in early postnatal ontogeny on blood pressure and heart activity in normo- and hypertensive rats. AB - Cold stress in the early postnatal ontogeny caused permanent functional changes in the cardiovascular system, which were different in hypertensive NISAG and normotensive WAG rats. Stress led to elevation of blood pressure and overload on the left heart chambers in adult WAG rats postnatally exposed to cold. At the same time, postnatal exposure to cold stress attenuated functional disturbances typical of hypertensive NISAG rats. PMID- 11276317 TI - Complex analysis of efficiency of transplantation of embryonic nerve tissue to rats with hemiparkinsonism. AB - Effect of transplantation of embryonic ventral mesencephalon preparation containing dopaminergic neurons on repair of the dopaminergic nigrostriatal system was studied in rats with hemiparkinsonism induced by 6-hydroxydopamine. Transplantation of embryonic ventral mesencephalon into denervated striatum led to a more than 50% decrease in apomorphine-induced rotation, recovery of dopamine and DOPAC levels in the brain, and to an increase in DOPAC excretion and the DOPAC-dopamine ratio in daily urine of rats with hemiparkinsonism. Dopaminergic neurons of the transplant survived, forming a network of tyrosine hydroxylase positive processes growing beyond the transplant and reinnervating the adjacent compartments of the striatum. A positive correlation between urinary excretion of DOPAC and brain concentration of dopamine was revealed in denervated rats after transplantation of ventral mesencephalon. Intrastriatal transplantation of cell preparations of embryonic striatum containing no dopaminergic neurons and isolated local injury to the striatum did not affect regeneration of the denervated nigrostratal system. PMID- 11276318 TI - Analysis of protein pool of neuronal populations of cerebellar cortex in rodents of different species. AB - The protein pool of neuronal population of the cerebellar cortex was studied by interference cytometry in rodents occupying different ecological niches and differing by life style, nutrition habits, and motor activity. In all cell populations protein concentrations in the cytoplasm were higher than in the nucleus in all studied rodents and did not depend on the functional characteristics of neurons. The extreme values of protein content were determined for populations of granular and ganglion cells. High protein concentrations per volume unit of cell structure were detected in functionally different cerebellar neurons of gray rats, characterized by high motor activity and a certain degree of synanthropy, while low values were detected in mole rats, slow-moving underground rodents. Therefore, the specific protein pool of neuronal populations of the cerebellar cortex of rodents can be regarded as adaptation to habitation conditions. PMID- 11276319 TI - Focal degradation of cytoplasmic organelles in cardiomyocytes during regenerative and plastic myocardial insufficiency. AB - Focal degradation of cardiomyocyte ultrastructures and cytoplasm, their sequestration, and autophagy were found in Wistar rats with daunomycin-induced regenerative and plastic myocardial insufficiency starting from day 1 after cytostatic treatment. These changes were morphologically manifested in the formation of myelin-like structures, autophagosomes, and secondary lysosomes. Sequestration and partial autophagy of the cytoplasm in cardiomyocytes with diminished or blocked protein synthesis reflect cell regression or involution directed to the adjustment of the cytoplasm volume to functional state of the nucleus. PMID- 11276320 TI - Ultrastructure of nuclear compartment in cardiomyocytes during regenerative and plastic insufficiency of the myocardium. AB - We studied ultrastructure of nuclear compartment in cardiomyocytes during regenerative and plastic insufficiency of the myocardium induced by anthracycline antibiotic daunomycin. A peculiarity of ultrastructural organization of cardiomyocyte nuclei under these conditions is almost complete disappearance of the heterochromatin lumps. The earliest changes in nucleoli under conditions of disturbed DNA-dependent RNA synthesis are segregation of the granular and fibrillar nucleolonema components. Deep alterations in the nucleoli manifested by fragmentation and annulation correlate with pronounced changes in cardiomyocytes ultrastructure, intensive lysis of the myofilaments, reduction of the organelles, and enhanced autophagocytosis. PMID- 11276321 TI - NADPH-positive neurons in heterotopic transplants of embryonic CNS. AB - Neurons of rat neocortex and spinal cord express NADPH-diaphorase after heterotopic allotransplantation into the sciatic nerve. These peculiarity of diaphorase expression in transplanted cells in comparison with brain cells developing in situ suggest an important role of afferent and efferent relationships in the development of NO mechanisms in neurons. PMID- 11276323 TI - Effect of 3,4-benzopyrene on ultrastructure of sinusoidal cells of the liver in adult male rats. AB - Reorganization of sinusoidal cells (endotheliocytes, Kupffer cells, and Ito cells) in the liver microregion of male rat liver exposed to 3,4-benzopyrene was studied. Synchronous activation of the lysosome-vacuole systems in Kupffer cells and endotheliocyte indicates cooperation of these cells in detoxification of benzopyrene and its metabolites. Ito cells lose lipid inclusions and actively proliferate; the appearance of intermediate forms between lipocytes and fibroblasts attests to activation of fibrogenesis in the liver. PMID- 11276322 TI - Effect of mu-opiate receptor agonist tetrapeptide A10 on DNA synthesis and protein content in the myocardium of albino rats. AB - Effect of intraperitoneal injection of tetrapeptide A10 (H-Tyr-D-Orn-Phe-Gly-OH), selective mu-opiate receptor agonist, synthetic analog of dermorphine, in a dose of 100 microg/kg on DNA synthesis and protein content in the myocardium was studied in albino rats. Five injections of tetrapeptide on days 2-6 after birth caused no changes in DNA synthesis 17 days after the last injection, i. e. in 24 day rats. The number of nucleoli and their area increased. In adult males long term (3-week) treatment with tetrapeptide A10 increased the number of nucleoli and the mean and integral optical density of isolated cardiomyocytes stained with amido black B, which probably attested to activation of protein synthesis in the myocardium. Simultaneously, the content of catecholamines in the heart increased. These data are comparable with delayed effects of kappa-opiate receptor agonist dinorphine A1-13 and indicate that morphogenetic properties of opioid peptides in rat myocardium are realized via the same routes. PMID- 11276324 TI - Comparative study of the laryngeal innervation in humans and animals employed in laryngeal transplantation research. AB - Laryngeal transplantation is receiving increased attention. Re-innervation of the transplanted larynx is critical for a successful functional outcome. Different anatomical models (dog, cat, rat, pig) have been employed for experimental purposes. Interspecies similarities and differences are important for extrapolating the experimental results to humans. We present a review of the anatomical course and regional branching patterns of the laryngeal nerves in both humans and animals currently being employed in laryngeal transplantation. The clinical and surgical implications are also discussed. PMID- 11276325 TI - Saint Blase, patron saint of otorhinolaryngology. AB - Otorhinolaryngology is one of the few medical specialities which has a patron saint, Saint Blase (born 317-AD). He was a Doctor and Bishop in Sebaste, Armenia, and he suffered martyrdom under the rule of the Roman Emperor Licinio (Iliria 250 AD - Tsalonica 325 AD). He was acknowledged as having the ability to protect people against throat infections, after curing a child who had choked on a fishbone. The feast of Saint Blase is on February 3rd, and it is celebrated all over the Western world. There are many other Saints related to our speciality, who protect people against ear, nose and throat disorders. We have reviewed the world literature on this subject. PMID- 11276326 TI - Canalplasty: review of 100 cases. AB - Canalplasty is the surgical procedure whereby the external auditory meatus is widened. The indications include exostoses, stenosing external otitis and widening for surgical access. One hundred consecutive ears operated on by one surgeon are reported. The surgical technique is described in detail, paying particular attention to bone removal from the anterior canal wall. In this paper the majority of cases were occasioned by soft tissue rather than bony stenosis. The re-stenosis rate was four per cent and in each case this was associated with the use of a middle temporal artery flap. Partial, transient, delayed facial palsy occurred in two per cent, probably relating to thermal injury transmitted from the burr. A full, spontaneous recovery of facial function occurred in each case. This is a safe and effective technique for canal widening. PMID- 11276327 TI - Hearing aids versus ventilation tubes in persistent otitis media with effusion: a survey of clinical practice. AB - A postal survey was carried out to determine the current clinical practice amongst consultant otolaryngologists in the UK, regarding re-insertion of ventilation tubes or recommendation of hearing aids in cases of recurrence of otitis media with effusion (OME) after ventilation tube extrusion. Amongst the 319 respondents, 15 (4.70 per cent) routinely, 146 (45.77 per cent) sometimes, and 158 (49.53 per cent) either never, or very rarely, recommend hearing aids. Hearing aids and ventilation tubes were both suggested to be equally good options by some consultants but they preferred surgery for a number of reasons. There were inconsistencies in practice and some of the reasons for re-inserting ventilation tubes are not evidence-based. A hearing aid is a non-invasive option and this survey shows a need for a randomized control trial of hearing aids and ventilation tubes in the management of persistent and recurrent OME. PMID- 11276328 TI - How common is hearing impairment in osteogenesis imperfecta? AB - Hearing impairment has long been recognized as a common feature in osteogenesis imperfecta. The figures in some publications could be taken to imply that, with increasing age, the proportion of osteogenesis imperfecta patients with hearing impairment approaches 100 per cent. The incidence of hearing loss in a large survey of 1394 patients with osteogenesis imperfecta was examined. It was found that the most common age of onset was in the second, third and fourth decades of life. At the age of 50 approximately 50 per cent of the patients had symptoms of hearing impairment; over the next 20 years there was little further increase. Differences were shown between patients with different clinical types of osteogenesis imperfecta as delineated in the Sillence classification; hearing loss was significantly less common in the type IV disease than in the type I disorder. Among the 29 families with osteogenesis imperfecta type IA there were distinct differences in the likelihood of hearing loss. These findings provide insights which will be valuable in giving patients advice on the likelihood of developing hearing loss in the future. PMID- 11276329 TI - Suprameatal approach: new surgical approach for cochlear implantation. AB - The conventional technique for cochlear implantation is via a mastoidectomy and posterior tympanotomy. An alternative approach for cochlear implantation is hereto described. The middle ear is entered through a suprameatal approach (SMA) bypassing the mastoid cavity. This surgical approach shortens the duration of the procedure to approximately one hour. The introduction of the cochlear implant electrode array involves drilling in the suprameatal region and the osseous portion of the external auditory canal at a safe distance from the anatomical position of the facial nerve. This prevents possible injury by direct trauma or drill overheating of the chorda tympani or facial nerves. We report 15 consecutive patients who were operated on using the SMA technique. No complications were encountered as a result of this surgical technique but further experience may be necessary. PMID- 11276331 TI - Diathermy epiglottectomy: endoscopic technique. AB - Endoscopic epiglottectomy is usually performed using a surgical laser. Epiglottectomy may be indicated for the treatment of benign or malignant lesions and for the relief of airway obstruction caused by a floppy epiglottis. We present four patients who had endoscopic partial epiglottectomy using monopolar diathermy to treat a floppy epiglottis. All the four patients were male in their sixth and seventh decades with snoring and/or obstructive sleep apnoea syndrome (OSAS) due partly or wholly to a floppy epiglottis. Diathermy epiglottectomy is easily carried out using the laryngoscope, laryngeal instruments and curved rotating microdissection monopolar scissors (used in laparoscopic surgery). This was found to be safe and effective with minimal morbidity. It requires no elaborate preparation and could be performed in hospitals that have no laser facilities. It may be performed in conjunction with other procedures e.g. uvulopalatopharyngoplasty (UPPP) if necessary. PMID- 11276330 TI - Endoscopic repair of unilateral choanal atresia with the KTP laser: a one stage procedure. AB - This paper, describes the endoscopic repair of unilateral choanal atresia with the KTP laser, a one-stage procedure, with no requirement for stenting. Three patients are presented with unilateral choanal atresia, aged six, nine and 38 years-old. The procedure combines the excellent endoscopic visualization, with the good haemostatic and penetrating properties of the KTP laser. Follow up was between 12 months and four years with all choanae remaining patent, no dilatation was required. No surgical complications were noted. PMID- 11276332 TI - Lateral saccular cysts of the larynx. Aetiology, diagnosis and management. AB - Lateral saccular cysts have been diagnosed in 17 patients. Definitive diagnosis was made by computed tomography (CT) that showed a fluid-filled cystic swelling of the saccule with no air fluid level in all patients. Fifty-three per cent of cysts were bulging through the thyrohyoid membrane. They were hugely enlarged in 18 per cent, moderately enlarged in 24 per cent, and slightly enlarged in 12 per cent of patients. One patient (six per cent) showed bilateral cysts. Lateral saccular cysts were primary in origin in 82 per cent of patients and secondary to prolonged intubation, hemilaryngectomy, and laryngoscleroma in 18 per cent of patients. Surgical excision via a lateral cervical approach was performed in eight patients, whereas endoscopic CO2 laser vestibulectomy was performed in nine patients. Endoscopic vestibulectomy with CO2 laser proved to be an efficient and safe procedure for excision of small and medium-sized cysts with a diameter equal to or less than 3 cm in the greatest dimension. The external approach appears more efficient than laser vestibulectomy in excision of huge cysts, as 22 per cent of patients undergoing laser surgery showed a recurrence. PMID- 11276333 TI - Radiotherapy for T1 glottic carcinoma: impact of anterior commissure involvement. AB - The clinical notes of all new patients with T(1) squamous cell carcinoma of the glottis seen in one head and neck cancer unit between 1989 and 1996 were reviewed. Fifty-three patients were treated with radical radiotherapy and of these 42 (79.2 per cent) had no loco-regional recurrence, after a median follow up of seven years. Eleven (20.8 per cent) developed local recurrence and were treated with salvage surgery. Fourteen of the 53 (26.4 per cent) tumours involved the anterior commissure and eight of these 14 (57.1 per cent) developed recurrence, whereas only three of the 19 (15.8 per cent) tumours arising from the anterior half of the fold but not involving the anterior commissure had recurrence. None of the remaining tumours recurred. This difference is statistically significant (p<0.001). Anterior commissure involvement is a predictor of poor response to radiotherapy. This may be the result of understaging as none of the cases had computed tomography (CT) scans performed. Technical radiotherapy factors may also be important, although in all cases of anterior commissure involvement steps were taken to ensure adequate radiation dose to this region. PMID- 11276334 TI - Deliberate removal of incisor teeth to allow access for laryngoscopy. AB - This paper describes a clinical situation where it was impossible to obtain a biopsy of a lesion at the anterior commissure in a patient with progressive hoarseness of voice using standard microlaryngoscopy techniques. Due to anatomical difficulties and a histological suggestion of laryngeal papillomatosis the incisor teeth were deliberately removed to allow an adequate view of the larynx and to facilitate further access. PMID- 11276335 TI - Facial paralysis in Wegener's granulomatosis of the middle ear. AB - A case of Wegener's granulomatosis, which presented as chronic otitis media with facial nerve palsy, is described. Early diagnosis is vital if unnecessary surgical exploration is to be avoided. A false negative cANCA may delay the diagnosis, especially in cases of locoregional disease, and a policy of repeated titres should be adopted, if clinical suspicion is high. PMID- 11276336 TI - Combined risk factors contributing to cerebral venous thrombosis in a young woman. AB - Cerebral venous thrombosis is a rare condition affecting predominantly adolescents or young adults. The presentation is often non-specific, and delay in diagnosis is common. The otolaryngologist may be consulted about the radiological findings of lateral sinus thrombosis and mastoid changes. The association of congenital thrombophilia with unusual presentations of venous thrombosis, especially in young individuals is now well documented. We present a case of lateral and sagittal sinus thrombosis complicated by cerebral venous infarction in a girl with protein C deficiency and masked mastoiditis. Unusual forms of venous thrombosis, including cerebral venous thrombosis may develop in association with a single risk factor for thrombosis, but additional risk factors should be sought especially when thrombosis presents in very young individuals. This case draws attention to the multi-causal nature of cerebral venous thrombosis in young adults, and highlights the issue of masked mastoiditis. A coordinated approach by otolaryngological and haematological teams is recommended in such cases. PMID- 11276337 TI - Sensorineural hearing loss as the initial manifestation of polyarteritis nodosa. AB - A 74-year-old male was referred for the sudden onset of bilateral sudden deafness. The patient had no history of any disease or trauma to the head. Pure tone audiometry revealed bilateral moderate, to severe, sensorineural hearing loss. Auditory brain stem responses (ABRs) showed normal peak and interpeak latencies. These audiological findings suggested that his hearing loss could be attributed to inner ear lesions. However, we felt an alternative explanation for this sudden deafness was likely to exist because the patient also had a month long fever of unknown origin (FUO) and weight loss of 5 kg/month. Using the criteria of The American College of Rheumatology, we made the diagnosis of polyarteritis nodosa (PAN). Serum MPO-ANCA was positive (x 661). For treatment, the patient was begun on prednisolone and cyclophosphamide. Nine months later, fever, hypertension, nephritis, pneumonitis, and arthritis had completely resolved, the MPO-ANCA became negative (MPO-ANCA < x 10). Furthermore, his hearing improved. PMID- 11276338 TI - Tinnitus caused by traumatic posterior auricular artery--internal jugular vein fistula. AB - A patient with an arteriovenous fistula that developed after a traffic accident was recently treated. The patient noticed pulsatile tinnitus in the right orbital region two months after the accident. On the first visit, the preliminary clinical impression of this case was a carotid-cavernous fistula, but angiography showed a fistula between the posterior auricular artery and the internal jugular vein. Although rare, this arteriovenous fistula should be included in the differential diagnosis of pusatile tinnitus in the orbit region. The fistula was controlled by embolization with a platinum coil. This is the first report of an arteriovenous fistula between the posterior auricular artery and internal jugular vein. PMID- 11276339 TI - Recurrent temporal petrositis. AB - The objective of this paper is to present and discuss the common features of temporal petrositis and the different approaches to its management. Petrositis used to be common before the antibiotic era. It can be associated with life threatening complications. The management of this problem used to be by an aggressive surgical approach. However, recent reports are describing good results with more conservative medical treatment and minimal surgical intervention, with the reservation of more aggressive surgical interventions for chronic or refractory cases. PMID- 11276340 TI - Bilateral subcutaneous emphysema of the orbits following nose blowing. AB - Orbital emphysema without evidence of any significant trauma is a rare occurrence. A case is reported here of bilateral subcutaneous emphysema of the orbital, in the absence of facial skeleton trauma, in a healthy adult male following nose blowing. It assumes importance because of potential complications such as loss of vision due to pressure effects and infection. Lamina papyracea is the most common site of bony defect and point of air entry into the orbit. Spontaneous resolution in around two weeks is usual. PMID- 11276342 TI - Isolated primary non-Hodgkin's malignant lymphoma of the larynx. AB - We report a case of glottic primary laryngeal lymphoma. Although the head and neck region is a frequent site of origin of extranodal non-Hodgkin's lymphomas, laryngeal involvement is exceptional. Including this case, about 90 primary laryngeal lymphomas have been reported in the literature. Microscopic study showed a diffuse malignant lymphoma of high-grade malignancy (WF sub-division H). A diffuse, large, B-cell-type NHL was diagnosed histopathologically. The patient was treated with combination chemotherapy, including cyclophosphamide, doxorubicin, vincristine and prednisolone (CHOP), which resulted in complete clinical remission after two courses. Four courses of combination chemotherapy were subsequently performed, making a total of six courses of combination chemotherapy. No recurrence has been observed during the 16-month follow-up period. PMID- 11276341 TI - Supraglottitis complicated by mediastinitis. AB - A rare case of supraglottitis complicated by mediastinitis is presented. Despite aggressive treatment with broadspectrum intravenous antibiotics, the patient persisted to have generalized supraglottitis. Subsequent computed tomography (CT) scanning revealed that she had developed a frank fluid collection starting at the suprasternal notch, extending retrosternally into the superior mediastinum. She recovered with conservative management and did not require aggressive mediastinal drainage as advocated by the literature. PMID- 11276343 TI - Acute tonsillitis complicated by retropharyngeal and thyroid abscess infected with de-repressed beta lactamase Citrobacter mutans. AB - An unusual presentation of acute tonsillitis complicated by retropharyngeal and thyroid abscess is reported. Spontaneous rupture of retropharyngeal abscess resulted in necrotic fistulae between the pharyngeal wall and the retropharyngeal space. PMID- 11276344 TI - Isolated oropharyngeal Kaposi's sarcoma in non AIDS patient: differences and similarities with spindle-cell haemangioendothelioma. AB - Vascular tumours rarely affect the oropharynx and overall they can represent a diagnostic challenge since their clinico-histopathological patterns are not always clear. This case report, of an isolated pharyngeal vascular proliferation, allowed the authors to analyse the similarities and differences between Kaposi's sarcoma and spindle-cell haemangioendothelioma. Moreover, it emphasizes the importance of diagnostic tools, such as the human herpesvirus 8 (HHV8) marker, that sometimes may represent the only reliable test for clearly establishing the diagnosis. PMID- 11276345 TI - True lateral dermoid cyst of the floor of the mouth. AB - Congenital dermoid cysts of the floor of the mouth are relatively rare but when they occur, they do so inevitably in the midline. We present a case of a true lateral dermoid cyst of this region without any intra-oral extension. We discuss the anatomical and histological classification of dermoid cysts within the floor of the mouth. PMID- 11276348 TI - All nasal polyps need histological examination: an audit-based appraisal of clinical practice. PMID- 11276352 TI - Successful marrow transplantation for correction of immunological deficit in lymphopenic agammaglobulinemia and treatment of immunologically induced pancytopenia. Reprinted from Experimental Hematology 1969; 11:-10. PMID- 11276353 TI - Investigator profile: David M. Bodine, PhD. interviewed by Vicki P.Glaser. PMID- 11276354 TI - Correlation coefficients between several parameters and CD34+ cell yield in peripheral blood stem cell harvesting by apheresis. PMID- 11276355 TI - In vivo purging of peripheral blood stem cells obtained by apheresis, using high dose chemotherapy and granulocyte colony-stimulating factor in chronic myelogenous leukemia patients. PMID- 11276356 TI - Cellular therapy of cancer with natural killer cells: will it ever work? PMID- 11276357 TI - Macrophage death and the role of apoptosis in human atherosclerosis. AB - The arterial disease atherosclerosis is responsible for severe morbidity and is the most common cause of death in the Western population. The complete pathogenesis of the disease is unknown, but multiple risk factors have been identified that correlate with the development of its complications such as heart attack and stroke. Evidence suggests that atherosclerosis is an inflammatory disease and the major cell types involved are smooth muscle cells, macrophages, and T lymphocytes. In this paper, we review the function of macrophages in the context of atherosclerosis and we also discuss the role and significance of macrophage death, including apoptosis. There is much evidence, certainly in vitro, suggesting that low-density lipoprotein becomes atherogenic when it undergoes cell-mediated oxidation within the artery wall. Besides inducing apoptosis in vitro, oxidized low-density lipoprotein may also cause extensive DNA damage in intimal cells, which might presage apoptosis. We review the results of experimental and clinical studies, which may indicate how the complications of atherosclerosis could be prevented by using different therapeutical strategies including bone marrow transplantation and gene therapy. PMID- 11276358 TI - Effects of cytokines on the culture and differentiation of dendritic cells in vitro. AB - The ability to culture dendritic cells (DC) in vitro has been integral to the dramatic increase in research in the area of immunotherapy. Over time, a number of methods for generating these cells have been developed. This article will provide an overview of the isolation and generation of DC and will give a detailed description of the role specific cytokines play in this process from the mobilization of precursors to the final maturation of DC. PMID- 11276359 TI - Ex vivo-expanded hematopoietic cell graft recipients exhibit T cell repertoire diversity similar to that seen after conventional stem cell transplants. AB - The feasibility of using ex vivo-expanded hematopoietic progenitor cells to reconstitute hematopoiesis after high-dose chemotherapy is presently being examined. Early studies have shown that myeloid and erythroid hematopoiesis can be successfully reconstituted after high-dose chemotherapy and ex vivo-expanded hematopoietic cell transplantation. The lymphoid reconstitution, however, has not been addressed previously. In this study, we examined the diversity of the T cell receptor V beta chain (TCRBV) repertoires in 5 breast cancer patients who were transplanted with ex vivo-expanded bone marrow mononuclear cells as the only source of hematopoietic graft. Using the TCRBV third complementarity determining region (CDR3) fingerprinting methodology, it is shown that CD4(+) and CD8(+) T cell subsets after ex vivo-expanded hematopoietic cell graft transplants exhibit TCRBV diversities that are similar in complexity when compared to those seen after conventional autologous peripheral blood stem cell transplants (PBSCT). No apparent difference in the extent of CDR3 diversity was found between ex vivo expanded and conventional autologous PBSCT recipients when the CD4(+) and CD8(+) subsets were further separated into CD45RA(+) "naive" and CD45RO(+) "memory" subsets. The diversity of the CD45RA(+) naive subsets was as complex as that of the CD45RO(+) memory subsets. These results indicate that T cell repertoire diversification is not further compromised when ex vivo-expanded hematopoietic cells are used instead of autologous peripheral blood stem cells as the only source of graft. PMID- 11276360 TI - Transforming growth factor: signal transduction pathways, cell cycle mediation, and effects on hematopoiesis. AB - Transforming growth factor-beta (TGF-beta) is a potent growth inhibitor of various cell types including hematopoietic cells. Two receptors, TGFbetaRI and TGFbetaRII, govern the interaction between the cell and the TGF-beta ligand. Primary binding of the ligand occurs with the RII receptor, promoting formation of a heterodimer with RI and activation of signaling. This induces transient association of Smad proteins with the receptors. Smad 3 and 4 may be involved in the TGF-beta-induced G(1) arrest. TGF-beta(1) down-regulates G(1) and G(2) cyclin dependent kinases (cdks) and cyclins in terms of both kinase activity and protein amount. TGF- beta (1) also inhibits phosphorylation of the product of the retinoblastoma tumor suppressor gene (pRb) at multiple serine and threonine residues in human myeloid leukemia cells. The underphosphorylated pRb associates with transcription factor E2F-4 in G(1) phase, whereas the phosphorylated pRb mainly binds to E2F-1 and E2F-3. Because TGF-beta(1) up-regulates p130(pRb family member)/E2F-4 complex formation and down-regulates p107(pRb family member)/E2F-4 complex formation, with E2F-4 levels remaining constant, these results suggest that E2F-4 is switched from p107 to pRb and p130 when cells exit from the cell cycle and arrest in G(1) by the action of TGF-beta(1). The "cdk inhibitor" p27 is both a positive and a negative regulator of TGF-beta(1)-mediated cell cycle control. Although TGF-beta(1) has been reported to be a selected inhibitor of normal primitive hematopoietic stem cells, TGF-beta inhibits both primitive and more differentiated myeloid leukemia cell lines. PMID- 11276361 TI - Inosine monophosphate and aspirin-triggered 15-epi-lipoxin A4 act in concert to regulate neutrophil trafficking: additive actions of two new endogenous anti inflammatory mediators. AB - Regulation of neutrophil (PMN) trafficking by soluble mediators is a critical component in the outcome of host defense, inflammation resolution, and neutrophil mediated tissue injury. Elucidation of the endogenous mediators that protect tissues from excess leukocyte traffic and aberrant PMN activation that can lead to tissue damage and chronic inflammation is of considerable interest, especially the endogenous mechanisms of anti-inflammation. To this end, we recently uncovered two new classes of mediators: inosine monophosphate (IMP) and aspirin triggered 15(R)-epimers of native lipoxin A(4). Here, we examined the combined actions of both classes of compounds in regulating key events in neutrophil trafficking. Neutrophil rolling in mouse microvessels was inhibited by both IMP or 5S,6R,15R-trihydroxy-7,9,13-trans-11-cis-eicosatetraenoic acid (15-epi-LXA(4)) in a concentration-dependent fashion. When combined, IMP (300 nM) and 15-epi-LX (10 nM) demonstrated additive inhibition of neutrophil rolling in microvessels. IMP and 15-epi-LX also significantly inhibited tumor necrosis factor-alpha (TNF alpha)-induced neutrophil accumulation into the mouse air pouch in a dose dependent manner. Again, the combination of low dose IMP (10 microg) and LX analog (5 microg) gave additive inhibition of neutrophil accumulation in this model. These results demonstrate the inhibition of neutrophil trafficking in two separate models by two different classes of small endogenous molecules. The additive inhibition by IMP and aspirin-triggered LX may represent key pathways that protect and resolve inflammatory responses that could be harnessed for treatment. PMID- 11276362 TI - Effects of vascular endothelial growth factor on acute myelogenous leukemia blasts. AB - Vascular endothelial growth factor (VEGF) and its specific receptors are expressed by various malignant cells, including acute myelogenous leukemia (AML) blasts. In this study we performed a detailed characterization of VEGF effects on native human AML blasts derived from a large group of consecutive AML patients with high blast counts in peripheral blood. Exogenous VEGF had divergent effects on spontaneous proliferation and cytokine-dependent (GM-CSF, G-CSF, IL-3) proliferation. Increased, decreased, or unaltered proliferation was observed in the presence of VEGF for various patients, and the VEGF effect differed even in the same patient depending on which exogenous cytokine being present together with VEGF. Similarly, increased, decreased or unaltered interleukin-1beta (IL 1beta) and IL-6 secretion was detected when VEGF was added, and for certain patients the effect of VEGF differed between IL-1beta and IL-6. Exogenous VEGF could also modulate proliferation and differentiation of clonogenic AML progenitors. Constitutive AML blast secretion of VEGF was detected for 40% of patients. Leptin, Flt3-L, IL-4, IL-10, and IL-13 had divergent effects on VEGF release by AML blasts. These results suggest that VEGF can modulate AML blast functions in vivo for a subset of patients. Furthermore, the detection of VEGF in peripheral blood stem cell (PBSC) autografts suggests that VEGF may influence the proliferation and possibly also the survival of contaminating AML cells in PBSC autografts. We conclude that VEGF may influence the functional characteristics of AML cells. Our results suggest that VEGF is important in leukemic hematopoiesis, and the detection of VEGF in PBSC autografts indicates that VEGF may influence the functional phenotype of contaminating AML cells in these grafts. PMID- 11276363 TI - Cytotoxicity of anti-CD64-ricin a chain immunotoxin against human acute myeloid leukemia cells in vitro and in SCID mice. AB - Blast cells from patients with acute myeloid leukemia (AML) commonly express CD64, the high-affinity receptor for immunoglobulin G (FcgammaRI). An immunotoxin (MDX-44) was constructed by coupling humanized anti-CD64 monoclonal antibody (mAb) H22 via a bivalent linker to deglycosylated ricin A-chain (RA). Human leukemia cell lines were incubated with MDX-44 or H22/free RA. The effect of MDX 44 on the proliferation of leukemia cells was assessed by [(3)H]thymidine incorporation. In the presence of interferon-gamma (IFN-gamma), MDX-44 significantly inhibited the proliferation of CD64(+) HL-60, NB4, and U937 cells in 72-h cultures in a dose-dependent manner. The mechanism of action appeared to be the induction of apoptosis, as measured by propidium iodide staining and flow cytometry analysis. However, CD64(-) KG-1a and Daudi cells were not affected by MDX-44/IFN-gamma. Incubating HL-60 cells with MDX-44/IFN-gamma resulted in a 99% decrease in colony-forming units, whereas colony-forming cells in normal bone marrow were not significantly suppressed by such treatment. Cells from 60% of AML patients (6/10) were inhibited by MDX-44/IFN-gamma, and the inhibition was correlated with CD64 expression on these cells (r = 0.65). In a human AML model in NOD/SCID mice, MDX-44/IFN-gamma inhibited 95-98% of peritoneal exudate AML cell proliferation and 85-90% of solid leukemia masses. The effect of MDX-44 on AML cells was dependent on activation of cells by IFN-gamma. MDX-44/IFN-gamma may have value in the therapy of AML cells expressing cell-surface CD64. PMID- 11276364 TI - Long-term bone marrow cultures provide access to early lymphoid progenitors. AB - Long-term bone marrow cultures provide defined systems for studying and manipulating hematopoietic progenitors. Myeloid bone marrow cultures harbor early lymphoid progenitors; however, the nature and phenotype of these progenitors has not been investigated. Phenotypic and molecular markers associated with lymphopoiesis were used to characterize the lymphoid population maintained in these cultures. Cells within myeloid cultures expressed genes associated with lymphopoiesis but did not express the B cell-specific lambda5 gene. Flow cytometry demonstrated that cultures lacked cells expressing markers associated with B cell development. Furthermore, rearrangements of immunoglobulin heavy chain diversity (D) and joining (J(H)) gene segments were not detected in the myeloid cultures suggesting that these conditions support early B cell progenitors. Transferring myeloid cultures to conditions optimal for lymphopoiesis resulted in B cell development that temporally recapitulated events in the bone marrow. We also demonstrate that these lymphoid progenitors are targets for retroviral transduction. This study suggests that long-term cultures provide a useful system to access early lymphoid progenitors and study the events that regulate their differentiation. PMID- 11276365 TI - MGMT expression in murine bone marrow is a major determinant of animal survival after alkylating agent exposure. AB - Myelosuppression is commonly observed after alkylating agent chemotherapy due to low levels of O(6)-alkylguanine DNA alkyltransferase protein (AGT) in hematopoietic progenitors. Mice that lack AGT in all organs, O(6)-methylguanine DNA methyltransferase gene knockout (MGMT(-/-)) mice are extremely hypersensitive to the methylating agent N-methyl-N-nitrosourea (MNU) and exhibit a 10-fold reduction in the LD(90). To determine whether bone marrow damage was the cause of the increased lethality, we transplanted 1 x 10(6) wild-type marrow into MGMT(-/ ) mice and MGMT(-/-) marrow into wild-type mice and observed survival after MNU. Lethally irradiated MGMT(-/-) mice given > or = 25 mg/kg MNU 3 weeks after transplant of wild-type cells survived > 30 days (n = 11), whereas this dose was lethal to control MGMT(-/-) mice 9-12 days post treatment (n = 5). Conversely, lethally irradiated wild-type mice transplanted with MGMT(-/-) cells died after only 20-60 mg/kg MNU within 8-12 days (n = 6). No significant toxicities were found in other organs. Additionally, in an in vivo post transplant competition model, wild-type long-term repopulating cells had a > 200-fold competitive survival advantage over MGMT(-/-) cells, and after MNU treatment completely repopulated the mouse when transplanted at only one-tenth the cell number. We also observed a strong selection for transplanted marrow-derived wild-type stromal elements in the MGMT(-/-) background after drug treatment. These data indicate that alkylating agent hypersensitivity of MGMT(-/-) mice results from hematopoietic damage at the stem level. Thus, DNA repair involving AGT in hematopoietic cells is required for normal host survival following exposure to methylating and chloroethylating agents. PMID- 11276366 TI - Immortalized multipotential mesenchymal cells and the hematopoietic microenvironment. AB - In an attempt to analyze the cellular and molecular basis of the capacity of bone marrow stromal cells to support hematopoiesis in culture, we developed a series of murine stromal cell lines from a single long-term bone marrow culture (BMC). The cytokines produced by these cells were analyzed using immunohistochemical techniques, ribonuclease protection assays (RPA) and RT-PCR. We examined the capacity of these cloned cell lines to replace primary bone marrow-derived stromal cells in long-term bone marrow cultures (LT-BMC) and sought correlations between the capacity to support hematopoiesis in culture with the production of known cytokines. These immortalized lines replicate many of the functions of the hematopoietic microenvironment. They express cytokines known to play a role in hematopoiesis. All of the lines constitutively express mRNA for PBSF (SDF-1), macrophage colony-stimulating factor (M-CSF), stem cell factor (SCF), FLT-3, thrombopoietin (TPO), interleukin 7 (IL-7), leukemia inhibitory factor (LIF), tumor necrosis factor-beta (TNF-beta), and interferon-gamma (IFN-gamma). Most lines also express granulocyte-macrophage colony-stimulating factor (GM-CSF) and G-CSF. They vary in their expression of IL-6, tumor growth factor-beta1 (TGF beta1), TGF-beta2, and TNF-alpha. Growing these lines in the presence of cytokines that influence hematopoiesis alters the levels of cytokine message. The most striking effects were produced by TNF-alpha. In addition to the cytokine mRNAs, the cell lines express factors associated with bone formation such as osteoblast-specific factor-2 (OSF-2) and bone morphogenetic protein-1 (BMP-1). They also express the neural cell-adhesion molecule neuropilin and neurotrophic factors including nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF). Several of the lines can maintain hematopoiesis in culture, as measured by the continuous production of myeloid colony-forming cells (CFU-c), for months. This capacity to support hematopoiesis does not correlate with any pattern of cytokine expression. Several of these lines also support the growth of human hematopoietic cells, and human CFU-c can be detected in the cultures in which CD34(+) bone marrow cells (BMC) are cultured on murine stromal cells. No correlation between the production of any of the known cytokines and the ability to support murine hematopoiesis was detected. In addition, there was no correlation between the capacity to support murine hematopoiesis and the capacity to maintain human HSC. Despite repeated cloning, the lines remain heterogeneous and are capable of producing cells with the properties of fibroblasts, osteoblasts, adipocytes, and myoblasts. In addition to the cytokine mRNAs, the cell lines express factors associated with bone formation such as OSF-2 and BMP 1. They also express the neural cell-adhesion molecule neuropilin and neurotrophic factors including NGF and BDNF. PMID- 11276367 TI - Urokinase-like plasminogen activator receptor expression on disseminated breast cancer cells. AB - Disseminated tumor cells are detected frequently in bone marrow, peripheral blood, and cytokine-mobilized peripheral blood cell products of women undergoing high-dose therapy for breast cancer. Several attempts were made to purge autografts from contaminating cancer cells; however, the biological and clinical impact of these contaminations has not been clarified so far. Expression of distinct phenotypes is a surrogate marker for metastatic behavior of cancer cells. The expression of the urokinase-like plasminogen activator receptor seems to be a factor of high importance. It is not expressed by normal mammary tissue. Disseminated cancer cells from marrow, blood, and stem cell products have been investigated by double-stain technique for urokinase-like plasminogen activator receptor (uPA-R) expressing cytokeratin-positive cells. uPA-R(+)/CK(+) cells could be found in all qualities of samples; however, significantly less in G-CSF mobilized peripheral blood stem cells compared to samples of other provenance (p = 0.02). It can be concluded that epithelial cells of malignant phenotype occur in blood, marrow, and autografts of breast cancer patients. Populations of disseminated tumor cells are phenotypically heterogeneous. Reduced uPA-R expression on cancer cells from leukapheresis samples might suggest a less aggressive nature of these cells compared to disseminated cells found in bone marrow. Furthermore, the data suggest that the phenotype of tumor cell contamination in leukapheresis products differs significantly from those of disseminated cancer cells in bone marrow or blood. PMID- 11276368 TI - Human recombinant interferon-inducible protein-10: intact disulfide bridges are not required for inhibition of hematopoietic progenitors and chemotaxis of T lymphocytes and monocytes. AB - Human recombinant interferon-inducible protein-10 (rIP-10), a C-X-C chemokine, inhibits proliferation of human hematopoietic progenitors responsive to co stimulation by recombinant steel factor (rSLF), is chemotactic for human monocytes and T-lymphocytes, and promotes T-lymphocyte adhesion to endothelial cells. Because chemokines have four conserved cysteines forming two intramolecular disulfide bridges, we decided to investigate their contribution in the biological activity of rIP-10. Since amino acid residues 22-98 of the sequence predicted by the cDNA constitute the naturally occurring IP-10, they were cloned after an initiating methionine into expression vector pET-3d. Subsequently rIP-10 was purified by enzymatic cell lysis, solubilization of refractile bodies with guanidine hydrochloride, renaturation by dialysis against dilute acetic acid, and sequential ion-exchange and reverse-phase high performance liquid chromatography. Purified rIP-10 was reduced with 20 mM dithiothreitol, and chemically modified with 100 mM iodoacetamide (IAA), or S methyl-methanethiosulfonate (MMTS), or N-methylmaleimide (NMM). Radiolabeling experiments demonstrated that 95% of the rIP-10 thiols were modified, and this was confirmed with SDS-PAGE. The biological activity of modified rIP-10 was determined in vitro by inhibition of rSLF-responsive human bone marrow hematopoietic progenitor proliferation and by chemotaxis assays using human T lymphocytes and monocytes. In both assay systems, the biological activity was evident at rIP-10 concentrations of 20-100 ng/ml. The activity was preserved after modification of rIP-10 by IAA or MMTS, but was abolished after modification by NMM. We conclude that disulfide bridges are not essential for the biological activity of rIP-10. PMID- 11276369 TI - Prior cryopreservation of ex vivo-expanded cord blood cells is not detrimental to engraftment as measured in the NOD-SCID mouse model. AB - Cytokine-mediated expansion has been proposed and successfully used to facilitate engraftment post transplantation. This study examined whether cryopreservation following expansion has a detrimental effect on the ability of cells to engraft, using the NOD-SCID mouse model. Cord blood (CB) CD34(+) cells were incubated for 7 days with stem cell factor (SCF), flt-3 ligand (FL), and megakaryocyte growth and development factor (MGDF). Expanded CD34(+) cells were transplanted into NOD SCID mice either fresh or following cryopreservation and thawing. After thawing, recovery of nucleated cells was 94%, of CD34 cells was 63%, and of day-14 progenitors was 17%. The loss of day-14 progenitor cells among the thawed expanded cells did not influence the kinetics of human engraftment in the mouse. Bone marrow (BM) of mice transplanted with thawed expanded CD34(+) cells (14 +/- 3.9%) showed significantly higher levels of human engraftment than mice transplanted with fresh expanded CD34(+) cells (1.5 +/- 0.5%, p = 0.0064). Thawed expanded CD34(+) cells had significantly higher SCID Engrafting Potential (SEP) than freshly expanded CD34(+) cells (p < 0.001). Results suggest that prior cryopreservation does not prevent expanded cells engrafting in NOD-SCID mice. PMID- 11276370 TI - Differential mobilization of CD34+ cells and lymphoma cells in non-Hodgkin's lymphoma patients mobilized with different growth factors. AB - Co-mobilization of CD34(+) cells and tumor has been documented in patients with different types of cancer undergoing peripheral blood stem cell transplantation (PBSCT). Conflicting reports were published regarding the role of various growth factors in tumor cells mobilization, hence we studied the extent of CD34(+) cells and lymphoma cell mobilization in 35 non-Hodgkin's (NHL) patients primed by cyclophosphamide (Cy) in combination with granulocyte colony-stimulating factor (GCSF) (A, 13 patients), granulocyte-macrophage (GM)-CSF (B, 10 patients), or GM CSF followed by G-CSF (C, 12 patients). CD34(+) cells were quantitated by flow cytometry and lymphoma cells by the TaqMan Real Time PCR for bcl-2 gene rearrangement. Successful collection in 4 days of > or = 2 x 10(6) CD34(+) cells/kg needed for prompt engraftment was obtained in 76%, 60%, and 58% of patients in arms A, B, and C, respectively. Lymphoma cell mobilization was detected in 35% patients tested, 78% of which had follicular lymphoma. Lymphoma cell mobilization was similar in the three arms of the study, however, presence of lymphoma cells was prevalent in patients who failed to mobilize the amount of 0.4 x 10(6) CD34(+) cells/kg in 2 days ("poor mobilizers") and reached 42%, compared to 17% in the "successful mobilizers" group of patients. Lymphoma cell contamination in PBSCs was detected proportionately in the peripheral blood and in the bone marrow. We conclude that bcl-2 gene rearrangement is prevalent in patients with follicular histology, and, in these patients, an inverse relationship was observed between mobilization of CD34(+) cells and lymphoma cells. Our results explain the high relative risk (1.98) for mobilization in patients with follicular histology. PMID- 11276371 TI - Mobilization of dendritic cells and NK cells in non-Hodgkin's lymphoma patients mobilized with different growth factors. AB - Conflicting results have been reported regarding the effect of various growth factors on the mobilization of natural killer (NK) cells and dendritic cells in patients undergoing stem cell mobilization for autotransplantation. We compared the extent of mobilization of NK cells and dendritic cells in non-Hodgkin's (NHL) patients undergoing mobilization with granulocyte colony-stimulating factor (G CSF), granulocyte-macrophage (GM)-CSF, or GM-CSF followed by G-CSF. Overall, 35 patients were studied. NK cells and dendritic were quantitated by flow cytometry. NK cells were defined as the sum of CD56(+) cells and CD56/CD16(+) cells. Dendritic cells were defined as the sum of CD80(+) and CD80(+)/CD14(+) cells. NK activity was determined by by microcytotoxicity assay. NK activity correlated well with the total amount of CD56(+) cells mobilized to the peripheral blood. Patients in the three arms of the study mobilized similar amounts of NK cells and NK activity, and patients who lacked NK activity in the peripheral blood, before mobilization, lacked NK activity in their apheresis collections. In contrast to NK cell mobilization, mobilization of dendritic cells/kg was three- to five-fold higher in patients mobilized with GM-CSF-containing regimens compared to patients mobilized with G-CSF alone. We conclude that GM-CSF-containing mobilization regimens are superior for dendritic cell mobilization but similar in the mobilization of NK cells. Therefore, we recommend using GM-CSF-containing regimens for patients undergoing ex vivo or in vivo manipulation of dendritic cells. PMID- 11276372 TI - Costs of diagnosis, treatment, and follow up of patients with acute myeloid leukemia in the netherlands. AB - The aim of the study was to calculate the costs in various places of acute myeloid leukemia (AML). Patients less than 65 years old, who were treated for newly diagnosed AML were included. The cost analysis distinguished between diagnosis, treatment, follow-up (maximum of 2 years), and treatment of relapse. The treatment period was divided into remission induction and consolidation treatment, harvest of bone marrow (BM) or peripheral blood stem cells, and transplantation. The costs of diagnosis amounted to $3,167 (1995 US$). Remission induction treatments cost on average $46,387 and harvest of bone marrow or peripheral blood stem cells costs $6,491. The costs of the transplantation varied between $25,531 and $44,087. Costs of follow-up amounted to $4,167. Relapse treatment, mainly consisting of reinduction therapy, costs on average $24,338. The total average weighted costs of AML patients amounted to $104,386. Treating AML patients is very expensive, and major reductions in costs are not expected in the next future. Considering efficacy and effectiveness, it seemed that choices based on costs could be made between several consolidation techniques and between a specific consolidation technique and/or palliative treatment. PMID- 11276373 TI - Influence of medium components on ex vivo megakaryocyte expansion. AB - Reinfusion of ex vivo-expanded autologous megakaryocytes together with a stem cell transplantation may be useful to prevent or reduce the period of chemotherapy-induced thrombocytopenia. In this study, we analyzed several serum containing and serum-free media to identify the most suitable medium for megakaryocyte expansion. Moreover, two thrombopoietin (Tpo)-mimetic peptides were tested to evaluate whether they could replace Tpo in an expansion protocol. To analyze the effects of different media on megakaryocyte expansion, we used an in vitro liquid culture system. For this purpose, CD34(+) cells were isolated from peripheral blood and cultured for 8 days in the presence of Tpo and interleukin-3 (IL-3). The presence of megakaryocytes was analyzed by flow cytometric analysis after staining for CD41 expression. For our standard culture procedure, megakaryocyte medium (MK medium) supplemented with 10% AB plasma was used. Addition of 5% or 2.5% AB plasma yielded higher numbers of megakaryocytes, implying the presence of inhibitory factors in plasma. However, some plasma components are required for optimal megakaryocyte expansion because addition of less than 1% AB plasma or addition of human serum albumin instead of AB plasma resulted in the formation of lower numbers of megakaryocytes. Two commercially available serum-free media were also tested: Cellgro and Stemspan. If CD34(+) cells were cultured in Cellgro medium similar numbers of megakaryocytes were obtained as when CD34(+) cells were cultured in MK medium supplemented with 10% AB plasma. In MK medium with 2.5% AB plasma, higher numbers of megakaryocytes were cultured than in MK medium supplemented with 10% AB plasma. Therefore, Cellgro medium is not the best alternative medium. In cultures with Stemspan medium, higher numbers of megakaryocytes were obtained compared to MK medium with 10% AB plasma. Stemspan is thus a good alternative for MK medium. Two Tpo-mimetic peptides, AF13948 and PK1M, were tested for their ability to replace Tpo. In cultures with AF13948, comparable numbers of megakaryocytes were obtained as in the presence of Tpo, but in cultures with PK1M the number of megakaryocytes was lower. This study shows that high concentrations of plasma in medium inhibits megakaryocyte formation, but some plasma components are required for optimal megakaryocyte expansion. For an ex vivo expansion protocol, it is worthwhile to test several media, because the number of megakaryocytes differs widely with the medium used. PMID- 11276374 TI - Lexipafant in severe acute pancreatitis: the final word? PMID- 11276375 TI - Pancreatic function tests: when to choose, what to use. AB - Although techniques for high-resolution imaging of the pancreas are constantly being improved, the evaluation of pancreatic function remains crucial for the workup of pancreatic diseases. More than 20 direct and indirect tests are available for the assessment of pancreatic function. Measurement of fecal elastase-1 is recommended as the most suitable test for the initial assessment of pancreatic function. Among other techniques, the pancreolauryl test, and alternatively the BT-PABA (N-benzoyl-L-tyrosyl-p-aminobenzoic acid) or the (13)C mixed-triglyceride test, yield the best sensitivity and specificity. Nevertheless, all indirect tests are of limited value in patients with mild to moderate impairment of pancreatic function. In these patients, the secretin caerulein test remains the gold standard. PMID- 11276378 TI - Genetic testing in acute and chronic pancreatitis. AB - Hereditary pancreatitis (HP) is clinically indistinguishable from pancreatitis with other causes. Patients with HP have an increased chance of developing pancreatitis. Mutations in the cationic trypsinogen gene appear to cause most HP, although there is evidence for mild genetic heterogeneity with defects in other genes. Trypsin stabilization and protection from autolysis appear to play a central role in the pathogenesis of pancreatitis. The role of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) as well as the pancreatic secretory trypsin inhibitor (PSTI) in patients with pancreatitis is intriguing but as yet incompletely understood. Genetic testing may help to identify and manage patients with HP. Healthcare professionals should understand the elements necessary for obtaining informed consent for patients undergoing these tests, the limits in interpreting test results, and the psychosocial issues that may arise from genetic testing. PMID- 11276377 TI - Prophylactic antibiotics and pancreatic necrosis. AB - Recent controlled clinical studies suggest a positive effect of early antibiotic treatment on late morbidity and mortality in severe acute pancreatitis. In contrast, widespread use of antibiotics may lead to an increased number of fungal infections and multiresistant bacteria. Optimal choice, duration, and route of administration of the antibiotic agent(s) are far from being established. The additional administration of an antifungal agent with prophylactic intention cannot be supported by the currently available data. PMID- 11276376 TI - Pancreatic enzyme replacement therapy. AB - Malabsorption due to severe pancreatic exocrine insufficiency is one of the most important late features of chronic pancreatitis. Generally, steatorrhea is more severe and occurs several years prior to malabsorption of other nutrients because synthesis and secretion of lipase are impaired more rapidly, its intraluminal survival is shorter, and the lack of pancreatic lipase activity is not compensated for by nonpancreatic mechanisms. Patients suffer not only from nutritional deficiencies but also from increased nutrient delivery to distal intestinal sites, causing symptoms by profound alteration of upper gastrointestinal secretory and motor functions. Adequate nutrient absorption requires delivery of sufficient enzymatic activity into the duodenal lumen simultaneously with meal nutrients. The following recommendations are based on modern therapeutic concepts: 25,000 to 40,000 units of lipase per meal using pH sensitive pancreatin microspheres, with dosage increases, compliance checks, and differential diagnosis in case of treatment failure. Still, in most patients, lipid digestion cannot be completely normalized by current standard therapy, and future developments are needed to optimize treatment. PMID- 11276379 TI - Update on familial pancreatic cancer. AB - Approximately 5% to 10% of patients with pancreatic cancer have one or more first degree relatives with this disease. A subset of these individuals have a hereditary form of pancreatic cancer designated by association with such hereditary disorders as familial adenomatous polyposis, hereditary nonpolyposis colorectal cancer, hereditary pancreatitis, or familial atypical multiple mole melanoma (FAMMM) syndrome. A subset of those FAMMM kindred with the CDKN2A (p16) germline mutation that expresses both pancreatic cancer and malignant melanoma may constitute a new hereditary pancreatic cancer-prone syndrome. PMID- 11276380 TI - Neoadjuvant, adjuvant, and palliative treatment of pancreatic cancer. AB - Pancreatic cancer remains a highly malignant disease. Curative treatment is only possible for patients diagnosed at a very early stage. Therefore, the vast majority of pancreatic cancer patients receive palliative treatment. Surgical palliation is offered to patients who are found not to have a resectable tumor. The treatment of obstructive jaundice is managed by stenting of the common bile duct or by a surgical bypass. The best possible surgical procedure should be based on the factors that influence hospital mortality, length of survival, and quality of life. In patients with a life expectancy of longer than 3 months, surgical bypass is recommended, with hepaticojejunostomy the treatment of choice. In the same surgical procedure, the relief of duodenal obstruction with a gastroenteric bypass should be achieved. Chemotherapy, radiotherapy, or a combination of both is employed as a neoadjuvant measure, as an adjuvant treatment, or, in most patients, as palliation. As palliative chemotherapy alone, 5-fluorouracil (5-FU) plus folinic acid is still the treatment of choice; however, newer drugs, such as gemcitabine, seem to have similar or marginally better results. Palliative radiochemotherapy with external-beam radiation plus 5 FU and folinic acid seems to lead to better local control of tumor progression but not to better survival, for which distant metastases are the limiting factor. PMID- 11276381 TI - Cholangiography and sphincter of Oddi manometry. PMID- 11276382 TI - Intraductal ultrasound in the biliary tract. AB - Although endoscopic ultrasonography (EUS) represents a major advance in endoscopic imaging, endosonography using dedicated echoendoscopes has some serious drawbacks, including the diameter of the echoendoscope (12 to 13 mm), the lack of intraluminal examination of the pancreatobiliary duct system due to the size of the instrument, unsatisfactory image quality and resolution for small lesions, and the need for a second examination separate from the previous routine endoscopy. Recently developed ultrasonographic miniprobes (diameter, about 2 mm; frequency, 12 to 20 MHz) can be passed through the working channel of standard endoscopes to provide high-frequency ultrasound images. These miniprobes may overcome some of the noted drawbacks and add to the safety and convenience of patients. Moreover, in various diseases of the gastrointestinal tract and the pancreatobiliary ductal system, the diagnostic accuracy of miniprobe ultrasonography has been proven to be superior to that of EUS. Miniprobe ultrasonography is a promising tool that adds new capabilities to the armamentarium of gastroenterologic diagnostic assessment. PMID- 11276383 TI - Precut sphincterotomy: indications, pitfalls, and complications. AB - Precut sphincterotomy is a technique employed to gain access to the common bile duct (CBD) when standard methods using catheters, cannulatomes, and guidewires have failed. It is particularly useful in cases of distal biliary strictures or distal impacted stones and in patients with Billroth II gastrectomies who require papillotomy. It significantly improves the overall success rate of CBD access. This technique should only be used, however, when a therapeutic maneuver is anticipated, and it has no place in diagnostic imaging. In the hands of experienced, skillful endoscopists, the complication rate is comparable with that of standard sphincterotomy. PMID- 11276384 TI - Photodynamic therapy in the biliary tract. AB - Biliary tract cancers are rare malignancies usually characterized by slow growth and a low propensity for metastasis. Despite the relatively localized nature of these cancers, the only therapeutic measure with curative potential to date is surgical intervention. Because symptoms occur late, the diagnosis is rarely made at an early stage, and therefore only about half of the patients can have curative surgery. Patients with advanced disease face a dismal prognosis because palliative treatment options are limited. This review outlines recent advances in treatment of biliary cancer. Encouraging results from prospective, single-arm phase II trials of photodynamic therapy in nonresectable cholangiocarcinoma suggest considerable promise for this new palliative treatment modality. However, the apparent benefit of photodynamic therapy on survival, jaundice, and quality of life must be confirmed in a randomized multicenter trial. PMID- 11276385 TI - Efficacy of biliary scintigraphy in suspected sphincter of Oddi dysfunction. AB - Sphincter of Oddi dysfunction (SOD) can pose diagnostic challenges for the physician. SOD is classified into types I, II, and III, but clinical outcome after sphincterotomy for suspected types II and III SOD has been unpredictable. Therefore, accurate diagnosis of types II and III SOD is important because of the increased risk of sphincterotomy in patients with SOD. Endoscopic sphincter of Oddi manometry (ESOM) is the gold standard for diagnosis of SOD; however, it is associated with significant morbidity and is not an appropriate screening test. Quantitative hepatobiliary scintigraphy (QHBS) has demonstrated good sensitivity as a screening test for SOD in patients following cholecystectomy; however, studies using this methodology are criticized for poor design and patient selection. Recent publications address these criticisms and provide evidence that QHBS and ESOM are comparable diagnostic tools after exclusion of organic biliary obstruction. QHBS can effectively replace invasive ESOM in the diagnostic algorithm of SOD. PMID- 11276387 TI - Diagnosis of high blood pressure in children by means of ambulatory blood pressure monitoring. PMID- 11276386 TI - Gallbladder sludge: what is its clinical significance? AB - Biliary sludge is a mixture of particulate solids that have precipitated from bile. Such sediment consists of cholesterol crystals, calcium bilirubinate pigment, and other calcium salts. Sludge is usually detected on transabdominal ultrasonography. Microscopy of aspirated bile and endoscopic ultrasonography are far more sensitive. Biliary sludge is associated with pregnancy; with rapid weight loss, particularly in the obese; with critical illness involving low or absent oral intake and the use of total parenteral nutrition (TPN); and following gastric surgery. It is also associated with biliary stones with common bile duct obstruction; with certain drugs, such as ceftriaxone and octreotide; and with bone marrow or solid organ transplantation. The clinical course of biliary sludge varies. It often vanishes, particularly if the causative event disappears; other cases wax and wane, and some go on to gallstones. Complications caused by biliary sludge include biliary colic, acute cholangitis, and acute pancreatitis. Asymptomatic patients with sludge or microlithiasis require no therapy. When patients are symptomatic or if complications arise, cholecystectomy is indicated. For the elderly or those at risk from the surgery, endoscopic sphincterotomy can prevent recurrent episodes of pancreatitis. Medical therapy is limited, although some approaches may show promise in the future. PMID- 11276389 TI - The metabolic syndrome and related cardiovascular risk. AB - The metabolic syndrome is a complex association of several risk factors including insulin resistance, dyslipidemia, and essential hypertension. Insulin resistance has been associated with sympathetic activation and endothelial dysfunction, which are the main mechanisms involved in the pathophysiology of hypertension and its related cardiovascular risk. According to the Sixth Report of the Joint National Committee, and guidelines of the World Health Organization/International Society of Hypertension, the presence of multiple risk markers suggests that both hypertension and risk factors should be aggressively managed in order to obtain a better outcome. Primary prevention of obesity at different levels--individual, familial, and social-- starting early in childhood has proven to be cost effective, and will be mandatory to reduce the world epidemic of obesity and its severe consequences. PMID- 11276391 TI - The adrenal and the metabolic syndrome. AB - This review discusses the possible interrelationships between adrenal steroid hormones and the metabolic syndrome. Abnormal regulation of the hypothalamic pituitary-adrenal axis has been proposed. Studies in the United Kingdom associated the metabolic syndrome with low birth weight and hyperactivity of the entire axis. In Italy, increased pituitary responsiveness to stimulation with vasopressin and corticotrophin-releasing hormone was demonstrated in women with central obesity. Swedish researchers have reported that increased stress responses of the axis correlated with a less variable but decreased cortisol level. An allele of the glucocorticoid receptor was also associated with various components of the metabolic syndrome. Evidence also suggests that central obesity is associated with an increased peripheral conversion of cortisol to cortisone and subsequent feedback stimulation of the axis. On the other hand, central fat may have an increased local metabolism in the direction of cortisol. Roles for dehydroepiandrosterone and aldosterone in the syndrome have also been proposed. PMID- 11276392 TI - Syndrome X following renal transplantation. AB - The cardiovascular risk factors that accompany postrenal transplantation include an atherogenic lipid profile, hypertension, new-onset diabetes mellitus, and a chronic prothrombotic state. This picture describes the dysmetabolic syndrome or syndrome X, which can significantly aggravate not only the risk of cardiovascular disease and death in this population, but also the progression of allograft dysfunction. The recognition and aggressive management of the dysmetabolic syndrome in postrenal allograft recipients may have a favorable impact on the incidence of cardiovascular morbidity and mortality in these patients and prolong allograft function. PMID- 11276390 TI - Nonesterified fatty acids in blood pressure control and cardiovascular complications. AB - The fact that cardiovascular risk factors cluster among individuals with the insulin resistance syndrome strongly suggests a common pathogenetic denominator. For many years, abnormalities of nonesterified fatty acid metabolism have been implicated in the disturbances of carbohydrate and lipid metabolism that characterize the cluster. However, until more recently, evidence implicating fatty acids in the hemodynamic and vascular abnormalities that affect patients with this syndrome was lacking. Observations from epidemiological, clinical, and basic science suggest that fatty acids can raise blood pressure and contribute to the development of hypertension. The effects of fatty acids on blood pressure may be mediated in part by inhibition of endothelial nitric oxide synthase activity and endothelium-dependent vasodilation. Fatty acids can also increase alpha1 adrenoceptor-mediated vascular reactivity and induce vascular smooth muscle migration and proliferation. The adverse effects of fatty acids appear to be mediated in part through induction of oxidative stress. Fatty acids interact with other components of the risk factor cluster, including increased angiotensin II, to synergistically augment oxidative stress in cultured vascular smooth muscle cells. Oxidative stress is implicated in the pathogenesis of insulin resistance, hypertension, vascular remodeling, and vascular complications. A clearer definition of the specific reactive oxygen signaling pathways involved and interventions aimed at altering these pathways could lead to more rationale antioxidant therapy and improved outcomes. PMID- 11276393 TI - Sex-related differences in the insulin resistance syndrome. AB - In the insulin resistance (IR) syndrome, sex-specific differences have been reported. First, hypertension more often correlates with hyperinsulinemia in women than in men with the IR syndrome. In addition, salt sensitivity of blood pressure appears to be independent of the activity of the renin-angiotensin system in women, whereas in men there is a strong correlation between the two variables. Secondly, the dyslipidemia found in women with the IR syndrome is characterized by less postprandial plasma insulin, triglycerides, and fatty acid response to a standardized meal. However, this sex difference in lipids disappears after correction for visceral fat mass. Fat physiology and biochemistry differ between the two sexes. In women, adipose cells express less glucocorticoid receptors and less 11beta-hydroxysteroid dehydrogenase. In women visceral fat accumulation appears to be a constant feature of the IR syndrome but in men the syndrome can be present without central obesity. Lastly, during the reproductive years of women, the IR syndrome, such as in pre-eclampsia, may cause fetal growth retardation that has been proposed together with maternal malnutrition to be at the origin of the increased risk for impaired glucose tolerance, hyperinsulinemia, and hypertension in adult life. This gives yet another dimension to this disease in women since in essence they may ultimately transmit this syndrome to both sexes. PMID- 11276395 TI - Recent clinical trial highlights in hypertension. AB - Clinical trials continue to guide patient management. However, the hypotheses generally come from observational studies. Studies on women continue to be too little and too few, particularly in light of the fact that most elderly hypertensive patients are likely to be women. Attention has been drawn to the possibility that hypertension per se may engender symptoms such as headache for example, in addition to harder endpoints such as death. The MICROHOPE study, which was not a blood pressure-lowering study, showed that angiotensin converting enzyme inhibition with ramipril in diabetic patients provided the same beneficial effects previously published for the HOPE study. The doxazosin arm of the ALLHAT study was terminated by the data monitoring and safety board because the doxazosin-treated patients developed congestive heart failure at a greater rate than diuretic-treated patients. Two extensive studies testing two different classes of calcium antagonists against primarily diuretic-based treatment showed that the calcium antagonists were no less effective in terms of preventing hard endpoints. A small, but impressive cross-over study testing the 3-hydroxy-3 methylglutaryl coenzyme A reductase inhibitor, pravastatin, against placebo showed that statin treatment lowers blood pressure in hypercholesterolemic patients with hypertension. Meta- analyses emphasized the value of blood pressure reduction in the elderly and added to the controversy and confusion about the role of calcium antagonists in the first-line treatment of hypertension. The point may be moot since with the current recommendations few hypertensive patients will be adequately treated with a single agent. PMID- 11276396 TI - RhoA/Rho-kinase, vascular changes, and hypertension. AB - Hypertension, the result of a sustained increase in vascular peripheral resistance, is partly due to vascular remodeling and increased vasoconstrictor sensitivity. Stimulation of heterotrimeric G-protein-coupled receptors by various contractile agonists activates intracellular signaling molecules to result in an increase in cytosolic Ca++ and the subsequent phosphorylation of myosin light chain by Ca++/calmodulin-dependent myosin light chain kinase. Additionally, a portion of alpha-adrenergic, serotonergic, and endothelin-1-induced contraction is partially mediated by the calcium-independent activation of the small G protein RhoA and of a downstream target, Rho-kinase. Isolated arteries from hypertensive animals have been shown to have an increased contractile sensitivity to various agonists and to exhibit evidence of remodeling. Recent data suggest that some of these vascular changes may be mediated by increased activity of RhoA/Rho-kinase, potentially introducing a novel therapeutic approach for the treatment of hypertension. PMID- 11276397 TI - How long should telomeres be? AB - What began as a study of the "end-replication problem" took on a new dimension as it became clear that telomeres are a "molecular clock" of replication in human somatic cells. Here we review the biology of telomeres in vitro and in vivo, in mice and humans. We suggest that, in humans, telomeres are involved in the biology of aging and pathobiology of disorders of aging, including cancer and cardiovascular disease. We also propose that the underlying dynamics of telomere biology is in line with broad principles of evolutionary theories. PMID- 11276398 TI - New developments in mechanisms of obesity-induced hypertension: role of adipose tissue. AB - Hypertension develops in almost 60% of obese individuals. Apart from the recent observation of obesity-associated structural changes in kidney structure that may lead to enhanced tubular sodium reabsorbtion, reports of paracrine and hormonal factors derived from adipose tissue have prompted speculations about the role of adipose tissue in the pathophysiology of obesity-induced hypertension. We summarize recent data on leptin's sympathoexcitatory actions, the possible influence of adipose tissue on atrial natriuretic peptide levels, and the formation of vasoactive substances, such as angiotensin II and nonesterified fatty acids, by adipocytes. The mechanisms discussed herein may contribute to the typical findings in obesity-induced hypertension, including volume expansion, sodium retention, enhanced sympathetic nervous system activity, increased activity of the systemic renin-angiotensin system, low atrial natriuretic peptide levels, and disturbed glucose and insulin metabolism. Together, these data strengthen the hypothesis that adipose tissue is potentially a major regulator of cardiovascular-renal function. PMID- 11276399 TI - Genetic rat models of hypertension: relationship to human hypertension. AB - Experimental models of human disease are frequently used to investigate the pathophysiology of disease as well as the mechanisms of action of therapeutics. However, as long as models have been used there have been debates about the utility of experimental models and their applicability for human disease on the phenotypic and genomic level. The recent advances in molecular genetics and genomics have provided powerful tools to study the genetics of multifactorial diseases, such as hypertension. However, studies of such diseases in humans remain challenging in part due to lack of statistical power and genetic heterogeneity within patient populations. For hypertension, various rat models have been developed and used for the identification of susceptibility loci for genetic hypertension. With the advent of "comparative genomics," the application of genetic studies to both human and animal model systems allows for a new paradigm, where comparative genomics can be used to bridge between model utility and clinical relevance. This review discusses recent approaches in genetics to facilitate gene discovery for polygenic disorders with specific focus on how comparative mapping can be used to select target regions in the human genome for large-scale association studies and linkage disequilibrium testing in clinical populations. PMID- 11276401 TI - World and mental health in 2001. PMID- 11276400 TI - Regulation of sodium/potassium ATPase activity: impact on salt balance and vascular contractility. AB - Na+,K+-ATPase distributes ions between the intracellular and extracellular space and is responsible for total-body sodium homeostasis. The activity of this ion pump is regulated by catecholamines and peptide hormones; by the ligand of Na+,K+ ATPase, ouabain; and by direct interaction with cytoskeleton proteins. This review summarizes recent advances in the field of short-term regulation of Na+,K+ ATPase and the implications of these advances for the regulation of blood pressure. Renal Na+,K+-ATPase activity is bidirectionally regulated by natriuretic and antinatriuretic hormones, and a shift in the balance between these forces may lead to salt retention and hypertension. Dopamine plays a key role in this interactive regulation. By inhibiting vascular Na+,K+-ATPase activity, an excess of circulating ouabain may increase calcium concentration in vascular cells and lead to increased vascular contractility. Finally, mutations in cytoskeleton proteins may stimulate renal Na+,K+-ATPase activity by way of protein/protein interaction and lead to salt retention and hypertension. Abnormalities in the systems regulating Na+,K+-ATPase should be explored further in the search for the multiple causes of essential hypertension. PMID- 11276402 TI - New treatments and approaches for attention deficit hyperactivity disorder. PMID- 11276403 TI - Genes and attention deficit hyperactivity disorder. AB - The initial molecular genetic studies of attention deficit hyperactivity disorder (ADHD) evaluated two candidate genes (DAT and DRD4) suggested by dopamine theories of this common disorder and its treatment with stimulant medication. The initial reports of weak associations with ADHD have been replicated by many (but not all) investigators, as is expected for genes with small effects. This literature is reviewed, along with emerging literature generated by active research groups investigating additional genes that might contribute to the genetic basis of this complex disorder. PMID- 11276404 TI - The pathophysiology and treatment of autism. AB - This article critically reviews research done in the past 2 years concerning the pathophysiology and treatment of autism. Recent research in genetics, neuroimaging, neurochemistry, and pharmacologic treatment has advanced the body of knowledge about the pathophysiology of autism. Relatively new imaging technologies (eg, positron emission tomography) are increasingly being applied to the study of subjects with autism and have produced promising results that await replication. Neurochemical and challenge studies continue to suggest a role for 5 HT dysregulation in autism. Additional research is needed to determine the role of neuroendocrine and autoimmune factors in autism. Significant gains have been made in determining which pharmacologic treatments are efficacious in autism. Additional research is needed on agents that might ameliorate the core and associated symptoms of autism. PMID- 11276405 TI - Turn-of-the-century ethical issues in child psychiatric research. AB - National concern in 2000 about increased psychoactive drug prescription for preschoolers accentuated the 1990s thrust for more pharmacologic research in children. Preschoolers are prescribed potent drugs without adequate evidence for efficacy or safety at this plastic age of the rapidly developing brain. Implementation of needed preschool research poses special ethical complications. Children with mental disorder qualify for special protection under both rubrics. Parental informed consent is crucial for preschoolers, who appear incapable of assent because of their preoperational, magical, animistic, egocentric thinking, with inability to comprehend relative risks and benefits. Whether they can dissent is an open question. Possibly for research with direct benefit outweighing the risk, parental permission/consent could override attempted preschooler dissent. Subject recompense should be adjusted for age differences in perception of amount, although parent reimbursement needs to be realistic. Insurance for research risk is desirable. Placebo controls appear justified for preschoolers because there is little evidence base to say that a proven effective treatment already exists. Disruptive behavior disorders, including attention deficit/hyperactivity, have enough evidence of preschool diagnostic validity to justify therapeutic trials. In preschool pharmacologic research, a brief trial of a nonpharmacologic treatment should precede the drug trial to ensure that placebo responders and responders to the alternative treatment are not exposed to drug risk. PMID- 11276407 TI - Adult attention deficit hyperactivity disorder. PMID- 11276406 TI - Antidepressant treatment in children and adolescents: bridging the gap between efficacy and effectiveness. AB - This review of antidepressant treatments in children and adolescents emphasizes the gap between efficacy data derived from randomized clinical trials (RCTs) and the limited effectiveness data from community-based practices. Part one is a brief review of data from randomized, double-blind clinical trials to assess the evidence base for the major approved indications for antidepressants in youths. Part two reviews information gaps in the evidence from RCTs. Part three discusses nonexperimental evidence of the use of antidepressants, including surveys of prescription sales, physician surveys, and population-based data. Part four presents a comprehensive model for assessing the use of antidepressants in youths in the community. The model aims to answer a range of public health-oriented questions and is intended to improve treating physicians' and clinical care providers' ability to manage medications for optimal patient benefit. Suggestions are made for engaging health service providers, health insurers, academicians, advocates, and the government in building the necessary infrastructure to make effectiveness as vital as efficacy to the model of drug therapy evaluation. PMID- 11276408 TI - The genetics of panic disorder. AB - Of the anxiety disorders, panic disorder (PD) has been the most extensively studied from a genetic standpoint. Results of family studies have consistently demonstrated that PD runs in families, and twin studies indicate that genes contribute to this familiality. However, phenotypic and genetic complexity has made finding the specific genes involved in PD a challenge. There is still uncertainty about how best to define the phenotype for genetic studies and whether it is the clinical phenotype of PD or more latent psychologic and biologic traits that are inherited. To date, molecular genetic studies have suggested some chromosomal regions and genes that may contribute to risk, but none of these have been established. We review the genetic epidemiology of PD as well as recent molecular genetic studies of the disorder, and conclude with a discussion of promising strategies that attempt to uncover specific genetic loci involved in the etiology of PD. PMID- 11276409 TI - Genetic counseling for psychiatric disorders. AB - Like other medical conditions, some psychiatric disorders are inherited, whereas others are not. Human genetics research is moving at a rapid pace. Genes for over 450 genetic disorders have been cloned and many disease-causing mutations have also been identified. The explosion of this new knowledge has created many new exciting opportunities in the diagnosis of these heritable disorders. The rapid pace of gene discovery will aid the identification of susceptibility genes for psychiatric disorders. Indeed, we can look forward to answers to many clinical and research questions. These are some of the gifts that the expanding field of human genetics research will continue to bring to medical science. However, as genetic tests for the detection of psychiatric disorders become available, many ethical, legal, and social implications will need to be considered. In this article, we review the principles of genetic counseling for psychiatric disorders, as well as the social and ethical dilemmas that genetic testing may bring. Although medical and scientific advances may bring many gifts, we should approach this new knowledge with caution, as one of the gifts may be a Pandora's box. PMID- 11276411 TI - The genetics of Tourette syndrome. AB - Twin and family studies demonstrate that Tourette syndrome (TS) is a genetic disorder. Early segregation analyses of family data were consistent with the hypothesis of autosomal dominant transmission; however, more recent studies suggest that the mode of inheritance is more complex. Current findings suggest that there are genes of major effect with other genes acting as modifiers. Several genome scans have been completed and several regions of interest have been identified that may harbor susceptibility genes for TS. Work is currently underway to replicate and extend these initial results. PMID- 11276410 TI - The genetics of alcoholism and alcohol abuse. AB - Twin studies have established that there are substantial genetic influences on alcoholism (0.5-0.6) in both men and women. Our knowledge of behaviors predisposing to alcoholism, including anxiety and impulsivity, is advancing rapidly through animal and human studies. Although alcoholism is often comorbid with other substance abuse and psychiatric disorders, recent studies have shown that, with the exception of nicotine, the heritability of alcoholism is largely substance-specific. Increasing understanding of the neurobiology of addiction has identified neural pathways in which genetic variation at candidate genes could influence vulnerability. Some functional variants of these genes have been identified. Recent linkage analyses in humans and rodents have pointed to genomic regions harboring genes that influence alcoholism. Refinement of clinical phenotypes and use of intermediate phenotypes will improve chances of gene identification. All these advances in the understanding of the genetics of alcoholism should facilitate the development of more accurately targeted therapies using molecular diagnostic approaches. PMID- 11276412 TI - The genetics of antisocial behavior. AB - Overall, the evidence from over 100 twin and adoption studies of antisocial behavior suggests that genetic factors account for about half of the variation in risk. However, behavioral genetic studies of antisocial behavior still tend to produce far-ranging estimates of heritability, suggesting that there may be important moderators of these genetic risk factors. In this review, the results of some recent behavioral genetic studies of antisocial behavior that focus on the following issues are examined: 1) developmental changes in the heritability of antisocial behaviors, 2) developmental subtypes of antisocial behavior disorders, 3) sex differences in the heritability of antisocial behavior, 4) cohort differences in the heritability of antisocial behavior, and 5) the genetics of antisocial behavior comorbidity. PMID- 11276414 TI - Induction of flowering in tropical trees by a 30-min reduction in photoperiod: evidence from field observations and herbarium specimens. AB - During the late rainy season in October 1997 we observed. over a range of >100 km, the highly synchronous emergence of flower buds in several deciduous tree species of the semi-deciduous tropical forest in Guanacaste, Costa Rica. Synchronous flowering soon after the rapid decline in day length around the September equinox and in the absence of any notable climatic cues suggested flower induction by declining photoperiod. By combining field observations and the analysis of flowering herbarium collections, we established highly synchronous flowering periods with low interannual and latitudinal variation predicted for photoperiodic flower induction for more than 25 tree species and a few herbs. We describe morphogenetic changes at the shoot apex of three species during flower induction and the suppression and induction of flowering in several herbaceous species by experimental daylight extension. The combined observations provide strong, mainly indirect evidence for photoperiodic induction of flowering in many tropical tree species. At low latitudes with annual variation in day length of 1 hour, flower induction must be caused by a decline in photoperiod of 30 min or less. This is the first report of photoperiodic control of flowering in trees. PMID- 11276413 TI - The genetics of obsessive-compulsive disorder. AB - A significant amount of evidence has been gathered to suggest that obsessive compulsive disorder (OCD) is a heritable disease. Despite this information, few details are understood about the genetics of the disorder. Molecular genetic techniques, especially candidate-gene association studies, have been used to try to appreciate the genetic components of this complex disorder. In addition to the complex inheritance pattern, the heterogeneous nature of the disorder complicates the analyses. This review outlines the evidence that has been gathered to date, implicating OCD as a genetic disorder. In addition, the most interesting and recent findings in the molecular genetic analyses are highlighted. PMID- 11276415 TI - Photoperiodic control of seasonal development and dormancy in tropical stem succulent trees. AB - Tropical stem-succulent trees store large quantities of water in their trunks yet remain leafless during the early and mid dry season. In contrast to most other tropical trees, bud break of vegetative buds is not induced in fully hydrated stem succulents between the winter solstice and the spring equinox by leaf abscission, abnormal rain showers or irrigation. Vegetative buds of leafless trees are therefore in a state of endo-dormancy similar to that of temperate perennial plants during early winter. Highly synchronous bud break regularly occurs soon after the spring equinox, often weeks before the first rainfalls of the wet season. These observations suggested that endo-dormancy and bud break might be induced by declining and increasing photoperiods after the autumn and spring equinoxes, respectively. In phenological field observations, we confirmed highly synchronous bud break after the spring equinox in many trees of five stem succulent species in the northern and southern hemispheres. Shoot growth of potted saplings of Plumeria rubra L. was arrested by a decline in day length below 12 h after the autumn equinox, but continued in saplings maintained in a 13 h photoperiod. Conversely, exposure to a 13-h photoperiod induced bud break of dormant apical buds in saplings and cuttings in January, whereas plants maintained in the natural day length of < 11.7 h remained dormant. Photoperiodic control of endo-dormancy of vegetative buds in stem succulents is thus supported by field observations and experimental variation of the photoperiod. At low latitudes, where annual variation of day length is less than 1 h, bud dormancy is induced and broken by variations in photoperiod of less than 30 min. PMID- 11276416 TI - Temperature response of leaf photosynthetic capacity in seedlings from seven temperate tree species. AB - Seedlings of seven temperate tree species (Acer pseudoplatanus L., Betula pendula Roth, Fagus sylvatica L., Fraxinus excelsior L., Juglans regia L., Quercus petraea Matt. Liebl. and Quercus robur L.) were grown in a nursery under neutral filters transmitting 45% of incident global irradiance. During the second or third year of growth, leaf photosynthetic capacity (i.e., maximal carboxylation rate, Vcmax, maximal photosynthetic electron transport rate, Jmax, and dark respiration, Rd) was estimated for five leaves from each species at five or six leaf temperatures (10, 18, 25, 32, 36 and 40 degrees C). Values of Vcmax and Jmax were obtained by fitting the equations of the Farquhar model on response curves of net CO2 assimilation (A) to sub-stomatal CO2 mole fraction (ci), at high irradiance. Primary parameters describing the kinetic properties of Rubisco (specificity factor, affinity for CO2 and for O2, and their temperature responses) were taken from published data obtained with spinach and tobacco, and were used for all species. The temperature responses of Vcmax and Jmax, which were fitted to a thermodynamic model, differed. Mean values of Vcmax and Jmax at a reference temperature of 25 degrees C were 77.3 and 139 micromol m(-2) s(-1), respectively. The activation energy was higher for Vcmax than for Jmax (mean values of 73.1 versus 57.9 kJ mol(-1)) resulting in a decrease in Jmax/Vcmax ratio with increasing temperature. The mean optimal temperature was higher for Vcmax than for Jmax (38.9 versus 35.9 degrees C). In addition, differences in these temperature responses were observed among species. Temperature optima ranged between 35.9 and above 45 degrees C for Vcmax and between 31.7 and 43.3 degrees C for Jmax, but because of data scatter and the limited range of temperatures tested (10 to 40 degrees C), there were few statistically significant differences among species. The optimal temperature for Jmax was highest in Q. robur, Q. petraea and J. regia, and lowest in A. pseudoplatanus and F. excelsior. Measurements of chlorophyll a fluorescence revealed that the critical temperature at which basal fluorescence begins to increase was close to 47 degrees C, with no difference among species. These results should improve the parameterization of photosynthesis models, and be of particular interest when adapted to heterogeneous forests comprising mixtures of species with diverse ecological requirements. PMID- 11276417 TI - Leaf gas exchange characteristics differ among Sonoran Desert riparian tree species. AB - We investigated leaf gas exchange responses to leaf temperature, leaf-to-air vapor pressure deficit (VPD), and predawn and midday shoot water potential (psipd and psimd, respectively) of two native Sonoran Desert riparian tree species, Fremont cottonwood (Populus fremontii S. Wats.) and Goodding willow (Salix gooddingii Ball), and one exotic riparian tree species, saltcedar (Tamarix chinensis Lour. and related species). Measurements were made at two sites over 2 years that differed climatically. Because multiple linear regression models explained less than 29% of the variation in stomatal conductance (gs) and less than 48% of the variation in net photosynthetic rate (Pn) of all species, we used boundary-line analysis to compare gas exchange responses among species. Gas exchange rates were high in all species. The hyperbolic relationship between Pn and gs suggested that initial reductions in gs at high gs did not inhibit Pn. Reductions in gs of cottonwood and willow occurred at psimd values at or below previously reported xylem cavitation thresholds (-1.6 and -1.4 MPa, respectively), indicating tight stomatal regulation of water loss and a narrow cavitation safety margin. In contrast, reductions in gs of saltcedar occurred at psimd values well above the cavitation threshold (-7.0 MPa), but at much lower psimd values than in cottonwood and willow, suggesting a wider cavitation safety margin and less tight regulation of water loss in saltcedar. High VPD had a smaller effect on leaf gas exchange in willow than in cottonwood. In contrast, willow had a less negative psipd threshold for stomatal closure than cottonwood. Compared with cottonwood and willow, leaf gas exchange of saltcedar was more tolerant of high VPD and low psipd. These physiological characteristics of saltcedar explain its widespread success as an invader of riparian ecosystems containing native Fremont cottonwood and Goodding willow in the Sonoran Desert. PMID- 11276418 TI - Photosynthesis and carbon allocation of six boreal tree species grown in understory and open conditions. AB - One-year-old seedlings of Abies balsamea (L.) Mill, Picea glauca (Moench) Voss, Pinus contorta Loudon, Betula papyrifera Marsh., Populus tremuloides Michx. and Populus balsamifera L. were transplanted in the spring, in pots, to the understory of a mixed P. tremuloides-P. balsamifera stand or to an adjacent open site. Growth and leaf characteristics were measured and photosynthetic light response curves determined in mid-August. Overall, the coniferous seedlings showed less photosynthetic plasticity in response to growth conditions than the deciduous species. Abies balsamea, P. glauca and B. papyrifera responded to the understory environment with higher leaf area ratios, and lower photosynthetic light saturation points and area-based leaf respiration relative to values for open-grown seedlings, while they matched or exceeded the height growth of open grown seedlings. In contrast, seedlings of Pinus contorta, P. tremuloides and P. balsamifera displayed characteristics that were not conducive to survival in the understory. These characteristics included a high light saturation point and leaf dark respiration rate in P. contorta, and lower leaf area variables combined with higher carbon allocation to roots in P. tremuloides and P. balsamifera. By the second growing season, all seedlings of P. tremuloides and P. balsamifera growing in the understory had died. PMID- 11276419 TI - Internal leaf anatomy and photosynthetic resource-use efficiency: interspecific and intraspecific comparisons. AB - Leaf mass per unit area (LMA) and internal leaf anatomy often affect net gas exchange because of their effects on internal CO2 conductance to the site of carboxylation, internal shading, competition for CO2 among carboxylation sites, nitrogen concentration and its partitioning. To evaluate effects of LMA and leaf anatomy on CO2 assimilation, water-use efficiency (WUE) and nitrogen-use efficiency (NUE), we measured LMA, leaf thickness, the thickness of mesophyll components, and gas exchange rates at ambient CO2 concentration in leaves of six woody deciduous and evergreen species with different leaf life spans. In two species, CO2 assimilation was also estimated at saturating CO2 concentrations. There were interspecific differences in all morphological variables studied. Long lived leaves had higher LMA and were thicker than short-lived leaves. Species with high LMA had low assimilation rates and NUE, both in ambient and saturating CO2 concentrations. Thus, in species with high LMA, assimilation was reduced by non-stomatal limitations, possibly because of a lower allocation of N to the photosynthetic machinery than in species with low LMA. Within a species, thicker leaves tended to have a lower tissue density. In intraspecific comparisons under field conditions, increasing internal air volume had positive effects on WUE, probably because of enhanced internal CO2 conductance to the site of carboxylation. We conclude that, in interspecific comparisons, different patterns of N partitioning strongly influence NUE, whereas in intraspecific comparisons, internal leaf anatomy is a key factor regulating resource-use efficiency. PMID- 11276420 TI - Growth stress distribution in leaning trunks of Cryptomeria japonica. AB - The distribution of growth stresses in leaning trunks of Cryptomeria japonica (L.f.) D. Don was determined by measuring the stresses released by the kerf method with strain gauges glued at specified positions along the trunks. Effects of both tree height and peripheral positions on the surface of leaning trunks on surface growth stress were determined. The inner residual growth strains in leaning trunks were also measured. We found high compression stresses in the lower side of leaning trunks that differed greatly from the tensile stresses in normal erect trunks. However, transverse compression stress was found around the tree trunk in both normal and compression wood. In leaning trees, the distribution of internal stresses in the bent trunk portion differed from that in the erect trunk portion, being compressive on the outside and tensile on the inside. The resistant moment introduced by compression stress generated in compression wood is released by the bending of the leaning trunk. The bending stresses are then superimposed on the original internal growth stress. We demonstrated that Poisson's effect of longitudinal stresses should be considered when evaluating transverse surface growth stresses. The existence and intensity of compression wood development can be assessed by growth stress measurements. We conclude that the compressing force of compression wood functions physiologically to give an upward righting response in a leaning trunk. PMID- 11276421 TI - Maturation, topophysis and other factors in relation to rooting in Larix. AB - We examined effects of mist quantity, topophysis (origin of cutting in the crown of 6-year-old trees), maturation state of the donor stock, and time of sticking on rooting and root system quality of cuttings representing five full-sib hybrid larch families obtained with Larix decidua Mill., L. laricina (du Roi) K. Koch, and L. kaempferi (Lamb.) Sarg. (Sieb. and Zucc.) Gord., as parents. Mist frequency, supplemental watering and family all had highly significant effects on percentage of cuttings rooting and root system quality. The high-frequency misting regime yielded both higher rooting percentages and higher quality root systems than the low-frequency regime without supplemental watering. Supplemental watering of the rooting medium in the low-frequency misting regime increased both percent rooting and root system quality to values comparable with those obtained by cuttings in the high-frequency misting regime. Rooting of cuttings from the top, middle and bottom whorls of 5-year-old plantation grown trees tended to decline with increasing height for three of the five families. Overall, height of cutting origin did not significantly affect rooting, but when analyzed separately, two families exhibited a significant decline in rooting toward the top of the trees. Age of donor ortet (ranging from 1 to 7 years) significantly affected both percent rooting and root system quality. Rooting percentages declined linearly with age, and root system quality declined more sharply than percent rooting. The relatively poor root system quality of cuttings from ortets older than 1 year was closely associated with plagiotropic growth. Softwood cuttings (stuck in mid-July) rooted better than hardwood cuttings (stuck in early September) across all families. PMID- 11276422 TI - Size doesn't matter. PMID- 11276423 TI - Twin peaks: the draft human genome sequence. PMID- 11276424 TI - The DNA-repair protein AlkB, EGL-9, and leprecan define new families of 2 oxoglutarate- and iron-dependent dioxygenases. AB - BACKGROUND: Protein fold recognition using sequence profile searches frequently allows prediction of the structure and biochemical mechanisms of proteins with an important biological function but unknown biochemical activity. Here we describe such predictions resulting from an analysis of the 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenases, a class of enzymes that are widespread in eukaryotes and bacteria and catalyze a variety of reactions typically involving the oxidation of an organic substrate using a dioxygen molecule. RESULTS: We employ sequence profile analysis to show that the DNA repair protein AlkB, the extracellular matrix protein leprecan, the disease-resistance-related protein EGL 9 and several uncharacterized proteins define novel families of enzymes of the 2OG-Fe(II) oxygenase superfamily. The identification of AlkB as a member of the 2OG-Fe(II) oxygenase superfamily suggests that this protein catalyzes oxidative detoxification of alkylated bases. More distant homologs of AlkB were detected in eukaryotes and in plant RNA viruses, leading to the hypothesis that these proteins might be involved in RNA demethylation. The EGL-9 protein from Caenorhabditis elegans is necessary for normal muscle function and its inactivation results in resistance against paralysis induced by the Pseudomonas aeruginosa toxin. EGL-9 and leprecan are predicted to be novel protein hydroxylases that might be involved in the generation of substrates for protein glycosylation. CONCLUSIONS: Here, using sequence profile searches, we show that several previously undetected protein families contain 2OG-Fe(II) oxygenase fold. This allows us to predict the catalytic activity for a wide range of biologically important, but biochemically uncharacterized proteins from eukaryotes and bacteria. PMID- 11276425 TI - Survey of transcripts in the adult Drosophila brain. AB - BACKGROUND: Classic methods of identifying genes involved in neural function include the laborious process of behavioral screening of mutagenized flies and then rescreening candidate lines for pleiotropic effects due to developmental defects. To accelerate the molecular analysis of brain function in Drosophila we constructed a cDNA library exclusively from adult brains. Our goal was to begin to develop a catalog of transcripts expressed in the brain. These transcripts are expected to contain a higher proportion of clones that are involved in neuronal function. RESULTS: The library contains approximately 6.75 million independent clones. From our initial characterization of 271 randomly chosen clones, we expect that approximately 11% of the clones in this library will identify transcribed sequences not found in expressed sequence tag databases. Furthermore, 15% of these 271 clones are not among the 13,601 predicted Drosophila genes. CONCLUSIONS: Our analysis of this unique Drosophila brain library suggests that the number of genes may be underestimated in this organism. This work complements the Drosophila genome project by providing information that facilitates more complete annotation of the genomic sequence. This library should be a useful resource that will help in determining how basic brain functions operate at the molecular level. PMID- 11276428 TI - Watching the hands of the Arabidopsis biological clock. AB - Oligonucleotide and cDNA microarrays have been used to analyse the mRNA levels of 8,000 genes in Arabidopsis thaliana throughout the day/night cycle. Genes involved in signal transduction and in various metabolic pathways were found to be coordinately regulated by circadian rhythms and/or by light. PMID- 11276427 TI - Nuclear dynamics: where genes are and how they got there. AB - DNA is highly organized spatially, both within domains of chromatin along each chromosome and within the nucleus as a whole. Recent studies suggest that chromatin localization can affect transcriptional and replicational activity. The similarity between the movements of chromatin nuclear bodies suggests a common mechanism that regulates nuclear dynamics. PMID- 11276426 TI - The cohesin complex: sequence homologies, interaction networks and shared motifs. AB - BACKGROUND: Cohesin is a macromolecular complex that links sister chromatids together at the metaphase plate during mitosis. The links are formed during DNA replication and destroyed during the metaphase-to-anaphase transition. In budding yeast, the 14S cohesin complex comprises at least two classes of SMC (structural maintenance of chromosomes) proteins - Smc1 and Smc3 - and two SCC (sister chromatid cohesion) proteins - Scc1 and Scc3. The exact function of these proteins is unknown. RESULTS: Searches of protein sequence databases have revealed new homologs of cohesin proteins. In mouse, Mmip1 (Mad member interacting protein 1) and Smc3 share 99% sequence identity and are products of the same gene. A phylogenetic tree of SMC homologs reveals five families: Smc1, Smc2, Smc3, Smc4 and an ancestral family that includes the sequences from the Archaea and Eubacteria. This ancestral family also includes sequences from eukaryotes. A cohesion interaction network, comprising 17 proteins, has been constructed using two proteomic databases. Genes encoding six proteins in the cohesion network share a common upstream region that includes the MluI cell-cycle box (MCB) element. Pairs of the proteins in this network share common sequence motifs that could represent common structural features such as binding sites. Scc2 shares a motif with Chk1 (kinase checkpoint protein), that comprises part of the serine/threonine protein kinase motif, including the active-site residue. CONCLUSIONS: We have combined genomic and proteomic data into a comprehensive network of information to reach a better understanding of the function of the cohesin complex. We have identified new SMC homologs, created a new SMC phylogeny and identified shared DNA and protein motifs. The potential for Scc2 to function as a kinase - a hypothesis that needs to be verified experimentally - could provide further evidence for the regulation of sister-chromatid cohesion by phosphorylation mechanisms, which are currently poorly understood. PMID- 11276429 TI - Fungal virulence studies come of age. AB - Sophisticated molecular biological research has revealed many virulence attributes in at least four pathogenic fungi, but the future study of fungal virulence requires investigators to distinguish between molecules that directly interact with the host, molecules that regulate these, and molecules that are always required for fungal growth and survival, independent of the host. PMID- 11276430 TI - Rhesus factors and ammonium: a function in efflux? AB - Completion of fungal, plant and human genomes paved the way to the identification of erythrocytic rhesus proteins and their kidney homologs as ammonium transporters. PMID- 11276431 TI - Brassica genomics: a complement to, and early beneficiary of, the Arabidopsis sequence. AB - Those studying the genus Brassica will be among the early beneficiaries of the now-completed Arabidopsis sequence. The remarkable morphological diversity of Brassica species and their relatives offers valuable opportunities to advance our knowledge of plant growth and development, and our understanding of rapid phenotypic evolution. PMID- 11276433 TI - State of the art review: videoscopic minimally invasive mitral valve surgery. Trekking to a totally endoscopic operation. PMID- 11276434 TI - Introducing multimedia scientific publishing and the new HSF. PMID- 11276435 TI - Recent advances in multivessel coronary grafting without cardiopulmonary bypass. AB - BACKGROUND: Coronary artery bypass grafting (CABG) without the heart lung machine has been possible for easily accessible targets such as the anterior descending or proximal right coronary. Until now technical difficulty in reaching lateral and inferior wall targets imposed significant barriers to multivessel off-pump grafting. To expand the potential for off-pump CABG the authors have devised new exposure and stabilization techniques suitable for all target vessels. In this report we relate our experience with these new techniques and demonstrate that multivessel coronary bypass can be safely performed without cardiopulmonary bypass (CPB). METHODS: From February 8, 1993 to December 16, 1997 a total of 280 patients underwent myocardial revascularization on the beating heart via median sternotomy. Until May 20, 1997 only patients with high preoperative risk factors for CPB were considered for this approach (Group A; N = 122). After this date any patients with favorable anatomy were included (Group B; N = 158) and were subsequently compared with patients operated on using CPB during the same time interval (Group C; N = 114). In Group B patients lateral and/or inferior wall targets were exposed by means of 4 cloth slings (2 through the transverse sinus and 2 behind the inferior vena cava) and by positioning the patients in Trendelenburg with rightward rotation of the table. Regional stabilization of the target artery was obtained with a commercial stabilizing foot plate. RESULTS: Thirty day hospital mortality was only 2 patients (1.6%) in Group A, 3 patients (1.9%) in Group B, and 3 patients (2.6%) in Group C (NS). Postoperative complications were low in both Group A and B. When Group B was compared with a similar cohort in whom CPB was used (Group C), there were statistically significant improvements in ICU and hospital stay demonstrated when CPB was not used (16.8+/-10.7 vs 26.3+/-38.6 hours respectively; p = 0.007, and 4.1+/-1.5 vs 5.5+/-2.4 days respectively, p<0.001). Angiographic followup was available for 78 patients in Groups A and B with a global patency rate (all grafts) of 98.6%, including a patency rate of 96.7% for 60 grafts to obtuse marginal branches of the circumflex). CONCLUSIONS: Multivessel CABG without CPB is possible with results similar to those obtained with pump-oxygenator support using simple exposure and stabilization techniques. PMID- 11276436 TI - Endoscopic repeat sternotomy. AB - BACKGROUND: Repeat cardiac surgery represents eight to twenty five percent of cardiac surgical procedures. Catastrophic hemorrhage is a known complication of repeat sternotomy. A number of techniques have been described to reduce the incidence of injury to the heart and mediastinal structures during reoperation. This paper reports a new endoscopic technique to visualize and lyse the adhesions between the sternum and heart prior to repeat median sternotomy. METHODS: A unique substernal retractor and endoscopic visualization system was developed specifically to facilitate safe and rapid sternotomy in reoperative cardiac cases. Twenty-four patients underwent elective reoperation using the Endoscopic Redo Sternotomy Retractor and instrumentation. There were 5 patients with prior valve surgery and 19 patients with coronary bypass grafts in place. Retrosternal adhesions were divided with special endoscopic cautery or scissors after which a standard reciprocating saw was used to open the sternum without damage to underlying structures. RESULTS: The time required for endoscopic dissection of retrosternal adhesions ranged from 6 to 22 minutes. No injury to any cardiac structure or conduit occurred. CONCLUSIONS: The Endoscopic Redo Sternotomy Retractor provides excellent visualization of all retrosternal structures and adhesions allowing safe and meticulous dissection prior to sternal opening. PMID- 11276432 TI - Fibroblast growth factors. AB - SUMMARY: Fibroblast growth factors (FGFs) make up a large family of polypeptide growth factors that are found in organisms ranging from nematodes to humans. In vertebrates, the 22 members of the FGF family range in molecular mass from 17 to 34 kDa and share 13-71% amino acid identity. Between vertebrate species, FGFs are highly conserved in both gene structure and amino-acid sequence. FGFs have a high affinity for heparan sulfate proteoglycans and require heparan sulfate to activate one of four cell-surface FGF receptors. During embryonic development, FGFs have diverse roles in regulating cell proliferation, migration and differentiation. In the adult organism, FGFs are homeostatic factors and function in tissue repair and response to injury. When inappropriately expressed, some FGFs can contribute to the pathogenesis of cancer. A subset of the FGF family, expressed in adult tissue, is important for neuronal signal transduction in the central and peripheral nervous systems. PMID- 11276437 TI - An intraluminal shunt for off-pump coronary artery bypass grafting. Report of 501 consecutive cases and review of the technique. AB - BACKGROUND: Clinical or subclinical manifestations of coronary ischemia may occur when the target vessel is temporarily occluded during revascularization of the beating unsupported heart. Laboratory evidence has shown that even when the surface electrocardiogram is normal, action potential duration, conduction velocity, and regional electrocardiogram patterns change during clamping of the target coronary artery. In addition, local trauma to the endothelium and/or adjacent atherosclerotic plaque by encircling snares or mechanical clamps can lead to plaque disruption and late stenosis in the native vessel. METHODS: The authors report their experience with a continuous series of patients undergoing coronary grafting on the beating heart using an intraluminal shunt. This simple device maintains distal perfusion and prevents ischemia while at the same time protects the anastomosis from potential suturing errors. The shunt also keeps blood from obscuring the operators vision and thus makes snares and clamps unneccesary. Smooth unhindered removal of the shunt immediately confirms patency of the finished anastomosis. RESULTS: Off-pump coronary grafting was performed in 501 consecutive patients utilizing an intraluminal shunt. Three hundred and seventy three men (74.5%) and 128 women ranging from 34 to 92 years old (mean 60.4 years) underwent a total of 196 internal mammary artery and 596 saphenous vein grafts (1.58 grafts per patient) from November 1983 to December 1996 at the Santa Casa de Sao Paulo Hospital and Hospital Samaritano. Mean shunting time was 14 minutes per anastomosis. Thirty day hospital mortality was 1.39% (7 patients) and all deaths were from non-cardiac causes. Perioperative myocardial infarction occurred in 7 other patients (1.39%) all of whom survived. CONCLUSIONS: In selected cases coronary grafts can be safely constructed on the beating heart without ischemia using a simple and inexpensive intraluminal shunt. The device is easily inserted and removed without damage to the native coronary. In a large series of patients, operative mortality and morbidity were lower than with conventional heart-lung support and cardioplegic arrest. PMID- 11276438 TI - Minimally invasive long saphenous vein harvesting using a laryngoscope. AB - BACKGROUND: Traditional open incisions for long saphenous vein (LSV) harvesting are common sources of post operative complications after coronary artery bypass grafting (CABG). To reduce pain and wound healing complications, minimally invasive harvesting techniques are being developed. We have investigated the use of a conventional laryngoscope for cost effective saphenous removal using short incisions and long subcutaneous tunnels. METHODS: The LSV was exposed through small incisions connected by long subcutaneous tunnels. Soft tissue retraction, visualization and illumination were provided by a sterilized laryngoscope with a #3 or #4 Macintosh blade. Dissection was performed with standard instruments while branch ligation was performed with vascular clips. Thirty two patients undergoing CABG between October 1997 and January 1998 underwent minimally invasive vein harvesting assisted by a laryngoscope. Clinical outcomes were evaluated. RESULTS: There were 27 males and 5 females with a mean age of 62.6 +/- 9.3 years in this study. Adequate saphenous vein was removed in 29 of 32 cases. (In three patients, the vein was so superficial that an open incision proved easier). The length of harvested conduit averaged 38.2 +/- 11.01 centimeters (21 55 centimeters). Harvesting time average 37.1 minutes (+/-10.8 minutes; range from 20 to 62 minutes). Postoperatively, There were no wound dehiscences, infections, cellulitis, or major hematomas. Pain and leg edema were considerably less than with traditional open harvest. CONCLUSIONS: Minimally invasive vein harvesting is less traumatic to the extremity with fewer complications and superior patient satisfaction. Although commercial disposable systems are now available to permit minimally invasive harvesting of the saphenous vein, a conventional laryngoscope can be used with much reduced costs. PMID- 11276439 TI - End-stage heart failure: is there a role for the Batista procedure? AB - BACKGROUND: Medically refractory heart failure is traditionally managed with cardiac transplantation although some limited success has also been obtained in selected patients using dynamic cardiomyoplasty or mechanical assist devices. Recently, a new surgical alternative called partial left ventriculectomy (PLV) was introduced by Batista in 1995. The procedure attempts to relieve symptoms of congestive failure by reducing myocardial mass and restoring the normal mass-to volume ratio of the left ventricle. Despite initial enthusiasm, the results of PLV are not yet known. The aim of this study was to determine survival and clinical outcomes in a group of patients submitted to PLV as a means of surgical treatment for end stage heart disease (ESHD) METHODS: From November 1994 to December 1995, 15 patients with ESHD and dilated cardiomyopathy (DCM) were operated on by the technique described by Randas Batista. We compared preoperative and postoperative assessments of NYHA Functional Class (FC), Quality of Life index (QOL), echocardiographic, ergometric, radioisotopic ventriculography and hemodynamic data at intervals of zero, one, three, six and nine, and twelve months postoperatively. Kaplan-Meier, student t-test and chi square analysis were applied to the numerical and categoric variables. RESULTS: Survival was 80% at one month, 66% at three months, 53% at six months, 47% at nine months and 40% at one year. We also found that 6 of 7 patients (85%) with tricuspid regurgitation (TR) died compared to 4 of 8 patients (50%) without TR. This was the only risk factor indentified which influenced mortality. Post operative echocardiographic evaluations demonstrated reduced left ventricular end diastolic and end-systolic diameters at six months (LVESD 65.5 +/- 8.3 mm preoperatively versus 56.83 +/- 5.74 mm at six months, p=0.007 and LVEDD 73.84 +/ 8.25 mm preoperatively versus 65.33 +/- 5.72 mm at six months, p=0.009). Survivors enjoyed an improved clinical status according to both the NYHA functional class (preoperative Class IV=100% versus postoperative at six months : Class IV = 50%, Class III = 17% and Class II = 33%) and the Quality of Life index (100% were in grade 6 and 7 preoperatively versus 0% at six months). However, statistical significance was not reached in most of these data due to the small number of patients. CONCLUSIONS: Actuarial survival in this series of patients was 53% at six months and 40% at twelve months with survivors showing fewer symptoms and clinical events than preoperatively (100% hospitalized preoperatively versus no patient hospitalized at six months). Therefore, the Batista Operation improves the quality of life patients with dilated cardiomyopathy and can possibly be a new means for bridging to cardiac transplantation in severely ill patients who are not likely to survive long enough to recieve a donor heart. Additional improvements in late results will likely be seen after further experience, evolution of the surgical techniques and better patient selection. PMID- 11276440 TI - Minimally invasive atrial septal defect closure using the subxyphoid approach. AB - BACKGROUND: Atrial septal defects in adults are associated with paradoxical emboli, atrial tachyarrythmias, and congestive heart failure. Surgical closure is highly efficacious with low operative mortality and morbidity. However, in young women sternotomy scars are unsightly reminders of an otherwise curative procedure. Alternatives such as lateral thoracotomy or extended transverse incisions are more cosmetic but associated with breast maldevelopment, numbness and other side effects. The authors are proposing a new surgical approach based on their observation that the right atrium and septum actually lie only 1 inch superior to the xyphisternal junction. METHODS: A 4 inch transverse inframammary incision is used and the linea alba divided. The lower sternum is lifted forward with a commericial cable-pully retractor system (Rultract). Using femoral bypass augmented by a balloon tipped cannula in the superior vena cava, the septal defect is easily visualized and closed with conventional techniques and equipment. RESULTS: Two young women have undergone closure of a patent foremen ovale (N=1) and a large ostium secundum (N=1) defect through this approach. One patient had minor fat necrosis in the incision which subsequently healed without incident. CONCLUSIONS: Close anatomic proximity between the atrial septum and the lower sternum make it feasible to approach ostium secundum defects using a purely subxyphoid exposure. Visualization of the defect is excellent without the need for thoractomy and sternotomy. The use of a small transverse incision in the inframammary crease makes the result cosmetically invisible. PMID- 11276441 TI - The "T-MIDCAB" procedure. Use of extension grafts from the undisturbed internal mammary artery in high-risk patients. AB - BACKGROUND: Minimally invasive direct coronary artery bypass grafting (MIDCAB) is an attractive new alternative for revascularizing patients with high perioperative risk for standard coronary surgery. However, limited surgical exposure through a small thoracotomy makes harvesting the full length of the internal mammary artery (IMA) very difficult and time consuming. We are now employing a new alternative with a "T" shaped bridge graft constructed from the undisturbed IMA using a 4 centimeter interposition segment of donor vessel. We prefer this approach in high risk cases in order to reduce the trauma of the thoracotomy, minimize pain and narcotic use, promote early extubation, and achieve immediate post-operative mobilization and recovery in patients who would otherwise be at risk for a poor outcome with conventional grafting techniques. METHODS: From September 10, 1997 to December 19, 1997 eight high-risk patients underwent at least one "T-MIDCAB" graft from the undisturbed IMA to the coronary artery using a short segment of either radial artery or saphenous vein. All cases were performed using a limited access anterior thoracotomy through the bed of the resected costal cartilage and without intercostal retraction. Five males and three females ranging from 58 to 83 years (average 73 years) were operated using this new concept. Pre-operative ejection fractions ranged from 25% to 80% (mean 43%). Parsonnet scores ranged from 21 to 43 (average 34) with predicted mortalities ranging from 30 to 40%. RESULTS: Eleven "T" grafts were placed (1.38 distals/patient). All 8 patients survived. Postoperative complications were minimal. The average length of stay was only 8 days (range 3 to 9 days). Intensive care unit stay averaged 3 days (range 1 to 4 days). One patient underwent postoperative angiography which demonstrated full patency of the conduit and all anastomoses. CONCLUSIONS: "T-MIDCAB" using a bridge graft of free radial artery or saphenous vein appears to be successful in high risk patients. The authors noted shorter operative times, reduced chest wall trauma and better pain control than with standard MIDCAB and full IMA harvesting. Cautious use of this procedure as an alternative to more morbid types of surgical revascularization is advised. PMID- 11276442 TI - Clinical evaluation of the on-line Sensicath blood gas monitoring system. AB - BACKGROUND: Blood gas analysis is an integral part of the management of open heart surgery and post-surgical intensive care patients. This study was conducted to compare the results of on-line blood gas analysis using the new Sensicath (SC) optical sensor technology against standard blood gas assay. METHODS: Comparative blood gas analysis was performed on 45 postoperative cardiovascular surgical patients and 5 patients during cardiopulmonary bypass. Simultaneous samples were obtained. One sample was sent to the central hospital laboratory for measurement using a Radiometer ABL 500 blood gas analyzer and the second sample was drawn past the optical sensor. Comparisons between the two methods for pH, pO2 and pCO2 were analyzed using linear regression and the method of Bland and Altman for the intensive care unit samples (N = 451) while the T test was used to compare data points obtained during cardiopulmonary bypass (N = 57). Bias, accuracy and precision were also calculated. RESULTS: The regression lines (r2) for pH, pO2, and pCO2 and were 0.69, 0.94, and 0.68 with slopes of 1.000, 0.0957, and 1.052 respectively for postoperative surgical patients. During cardiopulmonary bypass, the T test did not show any significant difference between the Sensicath and the reference values for the arterial and venous pH, pO2 and pCO2. CONCLUSIONS: In summary, this study found the Sensicath to be an accurate, useful tool which permits immediate blood gas analysis without blood exposure and constitutes an advance in the field of intensive care monitoring. PMID- 11276443 TI - Role of cardiopulmonary bypass in single vessel coronary revascularization: implications for MID-CABG. AB - BACKGROUND: Minimally invasive direct coronary artery bypass grafting (MID-CABG) is being utilized for the treatment of coronary artery disease in selected patients. This innovative procedure has generated numerous technical issues relating to coronary revascularization, including whether to perform the revascularization with or without cardiopulmonary bypass (CPB). METHODS: We addressed this issue indirectly by analyzing the 1995 New York State CABG registry, comparing patients who had single vessel bypass without CPB (Non-CPB Group) to a similar cohort of patients who had CABG performed on CPB (CPB Group). The database showed stratification of patients selected for bypass grafting without CPB to a significantly higher risk group, as shown by increased age, higher incidence of reoperation, transmural MI, congestive heart failure, carotid/cerebrovascular disease, and peripheral vascular disease. RESULTS: Patients in the Non-CPB Group had a higher incidence of postoperative malignant ventricular arrhythmias and heart block requiring pacemaker insertion. Otherwise, the incidence of postoperative complications was similar between the two groups. CONCLUSIONS: There were no statistical differences in the hospital mortality or the length of hospitalization between the two groups. In conclusion, the data showed a definite trend toward doing higher risk cases off CPB. These cases had an acceptable early morbidity and mortality outcome. The results were comparable to a group of lower risk patients with single vessel CABG done on cardiopulmonary bypass. However, further follow-up are required to evaluate long-term outcomes and confirm the utility of this surgical option. PMID- 11276445 TI - What's in a name? PMID- 11276444 TI - Autotransplantation procedure for giant left atrium repair. AB - BACKGROUND: Giant left atrium has been associated with bronchopulmonary and left ventricular compression [Kawazoe 1983]. CASE REPORT: We present a patient with severe congestive heart failure (CHF), respiratory insufficiency and a giant left atrium (GLA) following two previous mitral valve procedures and tricuspid valve annuloplasty in the distant past. Mitral prosthetic function and ventricular systolic function were felt to be normal leading to a tentative diagnosis of diastolic restriction from left ventricular compression and pericardial constriction. A pericardial decortication procedure through left thoracotomy was initially done but proved ineffective. Subsequently, full evidence of hemodynamic failure due to the giant left atrium and its respiratory complication was recognized and the patient underwent cardiac autotransplantation procedure [Kosak 1987], with the aim to reduce the left atrial dimensions to normal. CONCLUSIONS: Calcification of posterior left atrial wall prevented a completely satisfactory reduction of atrial size and the severity of ventricular adhesions from the previous pericardial procedure resulted in very long cardiopulmonary bypass time with severe bleeding complications. This case provides ample evidence that GLA can cause respiratory failure and needs to be surgically corrected. PMID- 11276446 TI - The frontier and beyond: the International Society for Minimally Invasive Cardiac Surgery revitalizes our profession. PMID- 11276447 TI - 3D-video- and robot-assisted minimally invasive ASD closure using the Port-Access techniques. AB - BACKGROUND: Video-assisted minimally invasive surgical methods with endovascular based femoral cardiopulmonary bypass (CPB) and balloon occlusion of the aorta (Port-Access technique) were used to close an ostium-secundum atrial septal defect (ASD) in 7 patients. METHODS: Minor modifications were made to the system to provide drainage of the superior vena cava. The surgery was performed through a small (3.5-5cm) right anterolateral thoracotomy with 3D video and robotic arm assistance. RESULTS: The operative procedures were completely uneventful and the patients were discharged four days postoperatively in good condition and with excellent cosmesis. CONCLUSION: Using the modifications described, the Port Access surgical method can be recommended for minimally invasive closure of an ASD. PMID- 11276448 TI - Coronary artery bypass grafting (CABG) without cardiopulmonary bypass (CPB): a strategy for improving results in surgical revascularization. AB - BACKGROUND: Coronary artery bypass grafting (CABG) was performed on patients with cardiopulmonary bypass (CPB group) or without CPB (non-CPB group). A series of CABG patients, performed between January 1, 1995 and September 30, 1997 is included. METHODS: Data were collected and analyzed as determined by the New York State Department of Health. Preoperative comorbidity, postoperative morbidity and mortality were compared. There were 2869 patients in the CPB group and 505 patients in the non-CPB group. RESULTS: Demographics of the two groups were similar but preoperative risk factors were more common in patients undergoing CABG without CPB. Of the non-CPB patients, 31.9% had reoperations as compared to 8.5% in CPB patients (p = 0.00005). The presence of an extensively calcified aorta was more common in the non-CPB patients (5.9% vs. 2.8%, p = 0.0002). Immune deficiency was also more common in the non-CPB group (p = 0.001). Risk-adjusted mortality was similar in the two groups while major complications were much less common when CPB was not utilized. In CPB patients only 84.3% avoided major complications, while among non-CPB patients 90.1% were complication-free (p = 0.0008). CONCLUSIONS: CABG without CPB is an attractive method of surgical revascularization. Increasing age and preoperative comorbidity in patients referred for CABG dictate changes in surgical strategy, of which avoidance of CPB appears most beneficial. PMID- 11276449 TI - Preoperative 3D-reconstructions of ultrafast-CT images for the planning of minimally invasive direct coronary artery bypass operation (MIDCAB). AB - BACKGROUND: The direct left internal mammary artery (LIMA) bypass to the left anterior descending (LAD) without the use of extracorporal circulation through a small anterolateral thoracotomy has become established among the minimally invasive techniques in cardiac surgery. Technical difficulties may occur in patients with an enlarged left ventricle and subsequent lateral positioning of the LAD, a small LAD, or a small LIMA. We used electron beam tomography (EBT) for preoperative visualization of the topographical structures to seek out patients with potential technical difficulties. METHODS: Eighteen patients, mean age 62 +/ 13 years, were entered in this study; in all cases the indication for revascularization was a significant stenosis of the LAD. Preoperatively an ECG triggered EBT was performed. Following the image acquisition, a three-dimensional reconstruction of the data was performed. The LIMA, LAD, first diagonal branch, and chest wall were stained different colors for better visualization. Surgery was performed using a left anterolateral mini-thoracotomy and through this access, the LIMA was dissected and anastomosed using a stabilizer without the use of extracorporal circulation. RESULTS: In all but one of the 18 patients who had a preoperative EBT, the minimally invasive direct coronary artery bypass (MIDCAB) procedure was successfully performed using an anterolateral mini-thoracotomy. Based on the results of the EBT, the 5 centimeter incision was done parasternally in six patients, and more laterally (2-4 cm parasternally) in the other eleven cases. In 13 patients the access penetrated the fourth intercostal space; in four cases the fifth intercostal space was used. In one patient EBT revealed a very laterally positioned and diffusely arteriosclerotic LAD so the patient was operated using a median sternotomy, but without the use of extracorporal circulation. In all 18 patients the preoperatively acquired information of the anatomical topography was confirmed intraoperatively. One case without a preoperative EBT had to be converted to a conventional procedure due to a small, intramyocardial LAD and a very small LIMA. Postoperative angiography revealed patent LIMA grafts and uneventful anastomoses. CONCLUSIONS: For minimally invasive direct coronary artery bypass (MIDCAB) the topography of the LIMA, LAD and intercostal spaces is of major importance. Using the ECG-triggered EBT with subsequent three-dimensional reconstruction these relationships can be visualized. This enables an individual planning of the operation and a minimalization of the skin incision. PMID- 11276450 TI - Double-outlet right ventricle with nonrelated ventricular septal defect: surgical results using the multiple patches technique. AB - OBJECTIVE: Introduce a new surgical technique for biventricular correction of double-outlet right ventricle with noncommitted ventricular septal defect. METHODS: From April 1987 to February 1996, 15 patients with double-outlet right ventricle with noncommitted ventricular septal defect were operated on using a new technique for biventricular repair with multiple bovine pericardial patches to create a tunnel between the left ventricle and the aorta. Ages ranged from two months to 13 years (mean age 4.8 years). Thirteen patients had situs solitus and levocardia, one patient had situs inversus and dextrocardia, and one patient had situs solitus and dextrocardia. Construction of the tunnel began at the right atrium. The ventricular septal defect (VSD) was enlarged anteriorly, if restrictive or small, and the first patch was sutured in the infero-posterior edge of the VSD. The second, third and sometimes the fourth patches were sutured in sequence, through the right ventriculotomy, directing the tunnel to the aortic annulus. RESULTS: Overall mortality was 20%, with two early and one late death. The surviving patients were followed-up for a period ranging from ten months to nine years (mean 33 months), and all were in functional class I (NYHA). Minimal residual ventricular septal defect was observed in one patient, stenosis in two patients and moderate pulmonary insufficiency in one. There was no obstruction of the intraventricular tunnel between the LV and the aorta. CONCLUSION: Based on these data, we conclude that this technical modification for the biventricular repair of the double-outlet right ventricle with noncommitted VSD allows for the construction of a tunnel with adequate internal diameter, respecting the spatial changes between the VSD and aorta. In addition, the intraventricular bovine pericardial tunnel takes up less space, thus reducing the incidence of right ventricle outlet obstruction. PMID- 11276451 TI - Fisics-Incor bovine pericardial bioprostheses: 15 year results. AB - BACKGROUND: From March 1982 to December 1995, 2,607 Fisics-Incor bovine pericardial bioprostheses were implanted in 2,259 patients. Mean age was 47.2 +/- 17.5 years, and 55% were male. Rheumatic fever was present in 1,301 (45.7%) patients. METHODS: One thousand and seventy-three aortic valve replacements, 1,085 mitral replacements, 27 tricuspid replacements, 195 mitral-aortic replacements, and 16 other combined valve replacements were carried out. Combined procedures were performed in 788 (32.9%) patients, the most frequent being tricuspid valve repair (9.2%) and coronary artery bypass grafting (7.7%). RESULTS: Hospital mortality was 8.6% (194 patients), 8.6% for the mitral group, 4.7% for the aortic group, and 12.8% for double-valve replacements. The linear rates for calcification, thromboembolism, rupture, leak and endocarditis were, respectively, 1.1%, 0.2%, 0.9%, 0.1% and 0.5% patient-year. The actuarial survival curve was 56.7 +/- 5.4% in 15 years. Survival free from endocarditis was 91.92%, survival free from thromboembolism was 95 +/- 1.7%, survival free from rupture was 43.7 +/- 9.8%, survival free from leak was 98.9 +/- 4.5%, and survival free from calcification was 48.8 +/- 7.9% in 15 years. In the late postoperative period, 1,614 (80.6%) patients were in New York Heart Association functional Class I. CONCLUSIONS: We conclude that the results with the Fisics Incor bovine pericardial prostheses were satisfactory in our group of patients. PMID- 11276452 TI - Late development of aortic pseudoaneurysm after coarctation repair with fistulization to the bronchial tree. A case report. AB - BACKGROUND: Fistulous communication between the aorta and the tracheobronchial tree is an uncommon and serious cause of hemoptysis secondary to complications of a previous operation performed on the aorta. In cases in which an appropriate surgical intervention is carried out, the survival rate approaches 76%. This surgery is considered one of the most risky operations on the aorta, challenging the surgeon's ability to resolve the problem. METHODS: We present the case report of a 43-year-old female with massive hemoptysis. Her medical history disclosed repair of coarctation of the aorta (15 years before). She underwent emergency left thoracotomy; surgical exploration revealed a false aneurysm from the previous aortic patch repair which communicated to a subsegmental bronchus of the left upper lobe. RESULTS: The thoracic aorta was isolated and clamped, and the previous patch was removed. The bronchial side of the fistula was managed with left superior lobectomy and the aorta was repaired with the placement of a coated woven dacron graft onto healthy aortic tissue. CONCLUSIONS: The patient had an uneventful recovery and remains asymptomatic six months after discharge. PMID- 11276453 TI - Continuous warm blood cardioplegia pitfalls and solutions. AB - BACKGROUND: Continuous warm blood cardioplegia offers superior preservation in both routine and complicated cardiac cases. Management of continuous perfusion is an important task during each case. METHODS: The authors have developed several specific techniques to ensure stable catheter insertion and placement for continuous coronary sinus or antegrade ostial perfusion. RESULTS: Over 3,800 patients have been operated on with continuous warm blood cardioplegia using catheter techniques as described in this article. The overall 30-day mortality was 3.9%. CONCLUSIONS: Safe application of continuous warm blood cardioplegia has many advantages over prior cold techniques, but surgeons must know certain technical modifications to be able to universally apply continuous techniques safely. PMID- 11276454 TI - Current therapy of catamenial pneumothorax. AB - Catamenial pneumothorax, or monthly recurring pneumothorax associated with menstruation, has been reported with increasing frequency in recent years. A representative case illustrates the clinical syndrome, particularly the intraoperative findings. Characteristic of this disorder are a peak incidence in the late twenties or early thirties, recurrent right-sided pneumothoraces occurring at the onset of menstruation, and an association with pelvic endometriosis. Pathologically, there is a consistent pattern of intrathoracic, especially diaphragmatic, foci of ectopic endometrial tissue. There is also a strong association with diaphragmatic fenestrations, though their significance is controversial. Traditional therapy has involved treatment with estrogens, danazol, or thoracotomy with mechanical pleurodesis. These methods have proven, through a large meta-analysis, to be associated with a relatively high rate of recurrence. Subsequent advances in hormonal therapy, along with the development of minimal access surgery, have led to an evolution in management. Despite uncertainty as to the etiology of catamenial pneumothorax, diagnosis of the condition is straightforward and modern treatment is successful in preventing recurrence. PMID- 11276455 TI - A perspective on MIDCAB. PMID- 11276456 TI - Off-pump multivessel coronary artery bypass utilizing the Octopus tissue stabilization system: initial experience in 374 patients from three separate centers. AB - BACKGROUND: Renewed interest in coronary artery bypass without heart-lung support has lead to the development of specialized instrumentation to permit surgical exposure and grafting in all territories of the beating heart. This report summarizes the results of a multicenter, prospective trial using a single system of mechanical stabilization for multivessel revascularization without cardiopulmonary bypass. METHODS: Three principal investigators at different institutions performed off-pump coronary grafting utilizing the Medtronic Octopus suction stabilization system which provides a motionless region encompassing the target coronary artery. Positioning and stabilization strategies evolved during the trial but eventually lead to a consistent approach for accessing all regions of the heart. Clinical data sets were collected prospectively and pooled for evaluation of early and short-term endpoints of success. RESULTS: A total of 374 patients underwent beating heart coronary bypass procedures with only a single death for an in-hospital 30-day mortality rate of 0.26%. There were 140 single vessel revascularizations (37.4% of patients), 119 double coronary grafts (31.8%), 90 triple-vessel grafts (24.1%), and 25 four-vessel grafts (6.7%), for a mean of 1.96 grafts per patient. If the single vessel cases are removed from analysis, the mean number of grafts performed in the multivessel cohort was 2.6 grafts per patient. All anatomic regions of the heart were successfully grafted including traditionally difficult locations such as the obtuse marginal branches of the circumflex and posterior descending branches of the right coronary artery. Only one patient suffered a new neurologic deficit which occurred 15 days postoperatively, for an overall incidence of only 0.26%. No patient required a new intra-aortic balloon pump or dialysis for renal insufficiency. The incidence of atrial fibrillation (12.8%) was age related and essentially unchanged from the overall incidence observed in patients operated at the same institutions using conventional techniques. CONCLUSIONS: Multivessel grafting on the beating heart using the Octopus stabilization system results in remarkably low perioperative mortality and morbidity, with very low incidences of cerebrovascular, renal, and respiratory complications. However, the incidence of postoperative atrial fibrillation is not reduced. Expanded clinical use of beating heart surgery with suction-based stabilization appears to be a promising technique for achieving global revascularization without the need for cardiopulmonary bypass. PMID- 11276457 TI - Robotically-assisted coronary artery bypass surgery: moving toward a completely endoscopic procedure. AB - BACKGROUND: Endoscopic coronary artery bypass grafting (ECABG) has not been possible with traditional techniques. This report details our animal experience determining the feasibility of using a robotically-assisted microsurgical system to perform ECABG. METHODS: Following preliminary work using a cadaveric pig heart model, acute and chronic animal studies were performed. Calves were placed on cardiopulmonary bypass after the left internal mammary artery (LIMA) was harvested. Subxiphoid endoscopic ports (2 instrument, 1 camera) were placed and a robotic system was used to perform ECABG between the LIMA and left anterior descending coronary artery. LIMA graft flow (LIMAQ) was measured, and excised hearts underwent angiographic and histologic analyses. RESULTS: All anastomoses were successfully completed in both the acute and chronic studies (mean time of 33.9 +/- 1.9 and 33.2 +/- 3.4 minutes, respectively). Angiographic patency was 100% in both the acute (8/8) and chronic (6/6) studies, which was confirmed by histology. In the chronic study, there was no difference in LIMAQ between intraoperative and autopsy measurements. CONCLUSIONS: This study shows that ECABG is feasible in an animal model with excellent results. The FDA has recently given approval for clinical trials of this new technology. PMID- 11276458 TI - Low flow veno-venous ECMO via subclavian dialysis catheter for severe respiratory failure. AB - BACKGROUND: We present the case of a 12-year-old female with severe postoperative bacterial pneumonia unresponsive to conventional treatment following a failed renal transplant. CASE REPORT: The patient was placed on low flow veno-venous extracorporeal membrane oxygenation (ECMO) as an adjuvant treatment to antibiotic therapy and maximal ventilatory support. Venous ECMO resulted in rapid improvement and the patient was successfully weaned after 48 hours of circulatory assistance. Two days later, the patient was extubated and safely discharged from the intensive care unit. Eighteen months later, she remains stable on peritoneal dialysis and is awaiting a new donor kidney. CONCLUSIONS: Low flow veno-venous ECMO represents a new therapeutic alternative for critically ill patients whose condition does not meet the conventional ECMO criteria. Further clinical experience is still needed. PMID- 11276459 TI - MIDCAB: impact of epicardial stabilization upon outcomes. AB - BACKGROUND: Minimally invasive direct coronary artery bypass (MIDCAB) has been criticized as compromising anastomotic patency. Epicardial mechanical stabilization devices purportedly facilitate left internal mammary artery (LIMA) anastomosis, thereby enhancing patency and outcome. METHODS: From June 1996 through January 1999, 39 patients underwent MIDCAB via a small left anterior thoracotomy for revascularization of the left anterior descending coronary artery (LAD) without cardiopulmonary bypass (CPB). Immediate postoperative coronary angiography was performed on 38 of the patients. RESULTS: Group 1 consisted of 11 patients who were operated upon without epicardial stabilization. Mean age was 64 years. Two had undergone previous coronary artery bypass (CAB). Predicted mortality was 4.3%. Angiographic anastomotic patency was 60%. Revisions on CPB in three cases increased LIMA patency to 90%. There was one intra-operative death. Average length of stay (LOS) was 5.4 days. Group 2 consisted of 28 patients operated on with mechanical epicardial stabilization. Predicted risk of mortality was 4.4%. Mean age was 66 years. Twelve had undergone previous CAB. Anastomotic patency at angiography was 97.4%. There were no intra-operative deaths and mean LOS was 3.0 days. CONCLUSIONS: We conclude that mechanical epicardial stabilization has transformed the MIDCAB operation into one that offers excellent early patency and clinical outcomes. This operation is of particular value for revascularization of the anterior coronary circulation in patients with previous CAB; clinical results are significantly better than predicted for standard redo CAB. PMID- 11276460 TI - Coronary artery bypass graft surgery in patients with ischemic cardiomyopathy and severe left ventricular dysfunction: short and long-term results. AB - BACKGROUND: We evaluated the prognostic value of preoperative parameters, surgical risk, functional benefits and long-term survival after myocardial revascularization in patients with established ischemic cardiomyopathy. METHODS: Seventy-one patients with ischemic cardiomyopathy, severe left ventricular dysfunction (left ventricular ejection fraction < 30%), and myocardial perfusion evaluated by Thallium-201 scintigraphy, were studied before and after myocardial revascularization, during hospitalization and throughout 48 months (average) of late follow-up. RESULTS: The early postoperative mortality was 2.8% and the five year survival rate was 62.8%. When the survival rate was studied, there was no correlation with 1) the presence of Q-waves on preoperative cardiogram, 2) the presence of ischemia on Tl-201 scintigraphy, 3) the degree of left ventricular ejection fraction, or 4) the presence of angina. There was a statistically significant difference for survivors and non-survivors in the following parameters: 1) functional class IV of CHF, and 2) the presence of left bundle branch block (LBBB). CONCLUSIONS: Our surgical results confirm that myocardial revascularization is a safe procedure, and that it increases late survival and improves the quality of life in patients with ischemic cardiomyopathy and severe left ventricular dysfunction. We also observed that due to heterogeneous coronary and myocardial patterns of ischemic cardiomyopathy, preoperative prognostic parameters are difficult to establish. Preoperative functional class IV congestive heart failure, and LBBB were the main predictors of poor outcome following surgical revascularization for ischemic cardiomyopathy. PMID- 11276461 TI - Continuous transesophageal echocardiographic (TEE) monitoring during port-access cardiac surgery. AB - BACKGROUND: Since the introduction of the closed-chest minimally invasive heart surgery using the Port-Access system a variety of monitoring techniques including fluoroscopy, transesophageal echocardiography (TEE) and invasive pressure measurements have been described. We investigated whether or not single TEE is feasible for perioperative monitoring of the placement, localization and proper function of the endovascular cardiopulmonary bypass (CPB) devices. METHODS: Fifty one patients (35 mitral valve repair or replacement [MVR], 8 coronary artery bypass grafting [CABG], 5 atrial septal defects [ASD] and 3 left atrial myxoma) were subjected to Port-Access surgery (PAS). Intraoperative Omniplane-TEE (2D- and color-flow Doppler techniques) was used as the leading monitoring device for correct positioning of the endopulmonary vent catheter and the venous cannula, and for the visualization of the guide wire and the endoaortic occlusion catheter (Endoclamp). After balloon inflation, its proper positioning and function during endo-aortic occlusion, sufficient delivery of cardioplegia into the coronary ostia, absence of leakage flow and adequate venting were controlled. Left and right radial artery catheters as well as aortic root pressure measurements served as controls. Additional fluoroscopy was used as standby device. RESULTS: In 46 patients (90.1%) sufficient perioperative monitoring was provided by single TEE. In five cases additional intermittent fluoroscopy was necessary for correct positioning of the guide wire (CABG) and the Endoclamp (three MVR and one ASD). Dislocation of the Endoclamp into the left ventricle was observed once but was successfully corrected by TEE guidance. Weaning from CPB and de-airing were easily guided with TEE. We did not observe balloon-mediated aortic injury or aortic valve dysfunction, and myocardial recovery from CPB was uneventful. All cases of MVRs showed sufficient results (68% without evidence of regurgitation, 32% showed residual mitral valve incompetence of less than grade II). Neither perivalvular leakage (MV-replacement) nor shunt- (residual ASD) flow were detectable. CONCLUSIONS: We recommend single TEE as a safe and effective on-line imaging device for monitoring the endovascular CPB system during PAS. Fluoroscopy with its potential risk for the patients and the staff due to x-ray exposure should only be used in the presence of peripheral vascular disease or when echocardiographic imaging is insufficient. PMID- 11276462 TI - Bovine pericardium used as a cardiovascular patch. AB - BACKGROUND: The use of glutaraldehyde preserved bovine pericardium in valvular prostheses is well known. Although widely used clinically as patch material, bovine pericardium has not been extensively studied in this setting. METHODS: With this objective, 21 dogs received a standard bovine pericardial patch to partially replace the aortic, left atrial, or pericardial walls. The dogs were randomly divided into three groups according to implant duration. Group 1 consisted of 6 dogs evaluated surgically after 33 to 43 postoperative days, Group 2 with 7 dogs reoperated after 120 to 165 days, and Group 3 with 8 dogs reoperated after 225 to 305 days. RESULTS: Microscopic and macroscopic evaluation demonstrated: 1) the wrinkled surface of the bovine pericardium adhered to neighboring structures whereas the smooth surface did not adhere to the epicardium; 2) the bovine pericardial patch did not show structural changes in any of the implant sites; 3) the final left atrial patch was significantly smaller than the aortic and pericardial patches for Group 2 and Group 3 dogs; 4) the atrial patch area decreased significantly, whereas the aortic and pericardial areas did not change over time; 5) the pericardial implant was significantly thinner than the aortic and left atrial patches for Group 3 dogs; 6) a layer of fibrous connective tissue was formed on the smooth surface of the left atrial and aortic patches. The internal apposition fibrasis was significantly thicker in the left atrium than in the aorta in Groups 1 and 2; 7) the internal fibrasis layer of the atrial and aortic patches was calcified in Groups 2 and 3; and 8) the internal apposition tissue of the atrial and aortic patches showed neoformation of elastic fibers which clearly increased with implant duration. CONCLUSIONS: The fate of implanted glutaraldehyde-preserved bovine pericardial patches in cardiovascular applications depends on three factors: 1) the contact surface, 2) the tension it is subjected to, and 3) contact with blood flow. PMID- 11276463 TI - Intermittent anterograde normothermic blood cardioplegia: experimental study in rabbits. AB - BACKGROUND: We investigated the degree of myocardial protection provided by intermittent anterograde normothermic blood cardioplegia infusion for 60 minutes at 37 degrees C in normal rabbit hearts. METHODS: Thirty-two New Zealand rabbits were studied and divided into two groups: experimental group and control group. In the experimental group, normothermic blood cardioplegia was infused into the aortic root every 20 minutes over a one-hour period using a two-minute infusion dose. This amounted to an ischemic (unperfused) time of 52 minutes (or 86.6% of the total time). The biochemical investigation was carried out in two phases; Phase I: metabolic study after ischemia with no reperfusion and Phase II: metabolic and functional study after reperfusion. Reperfusion was carried out using a parabiotic perfusion system. Myocardial glycogen and mitochondrial respiration in the ventricular myocardium were established immediately after the end of intermittent cardioplegic solution infusion (Phase I) and after blood reperfusion (Phase II), when left ventricular function (dP/dt max) was also evaluated. RESULTS: At the end of Phase I, there was a significant decrease in myocardial glycogen levels to 58% compared with the control group. In Phase II, the differences in myocardial glycogen between the experimental and the control group were not significant. Mitochondrial respiration analysis did not show significant differences between the experimental and control groups, either in Phase I or II. In Phase I, dP/dtmax values were 903.39 +/- 113.46 mmHg/sec and 1,043 +/- 256.94 mmHg/sec for the experimental and control group, respectively. These differences were not statistically significant. CONCLUSIONS: Intermittent anterograde blood cardioplegia infusion every 20 minutes for 60 minutes at 37 degrees C was an effective myocardial protection method in normal rabbit hearts. PMID- 11276464 TI - Xyphoid MIDCAB: report of the technique and experience with a less invasive MIDCAB procedure. AB - BACKGROUND: Coronary bypass surgery on the beating heart has been in existence since the inception of coronary revascularization. The advent and evolution of the heart-lung machine and cardioplegia have greatly advanced and expanded the realm of bypass surgery, allowing surgeons to perform precise coronary anastomoses in a still field of the arrested heart. The minimally invasive direct coronary artery bypass (MIDCAB) has been used primarily for grafting the left internal mammary artery (LIMA) to left anterior descending artery (LAD) and is gaining acceptance as a less invasive option. Dr. Frederico Benetti in Argentina championed the resurgence of beating heart surgery in 1985, and pioneered the left anterior thoracotomy MIDCAB procedure, which he has further refined to a xiphoid approach. The xiphoid incision is a simpler, less painful approach than that through a left anterior thoracotomy. METHODS: An incision is made through the xiphoid and, if necessary, into the tip of the sternum allowing the left side of the chest to be elevated. The distal LIMA is identified and dissected proximally to about the third interspace to give enough distance for a tensionless anastomosis. The pericardium is opened and the heart positioned to expose the LAD. Local stabilization for the LAD is obtained and the LIMA-to-LAD anastomosis is performed. RESULTS: The xiphoid MIDCAB approach was used for LIMA to-LAD anastomosis in ten patients ranging in age from 52 to 86 years (mean age 73 years). Three patients underwent angioplasty of additionally obstructed vessels (so-called hybrid procedures) following initial MIDCAB. Despite high preoperative-risk profiles and Parsonnet scores, there were no deaths. However, two of the three hybrid patients had major complications resulting in a prolonged hospital stay. CONCLUSIONS: Initial clinical experience with xiphoid MIDCAB proves it is a feasible alternative to intercostal MIDCAB with the possible advantages of reduced pain and chest wall complications. Further investigation into this surgical approach is warranted. PMID- 11276465 TI - The birth of a new society.... my two years as president of ISMICS. PMID- 11276466 TI - Intraoperative monitoring during cardiac surgery: some observations. PMID- 11276467 TI - Development of an off bypass mitral valve repair. AB - BACKGROUND: The Bow Tie Repair (BTR), a single edge-to-edge suture opposing the anterior and posterior leaflets of the mitral valve (MV), has led to satisfactory reduction of mitral regurgitation (MR) with few re-operations and excellent hemodynamic results. The simplicity of the repair lends itself to minimally invasive approaches. A MV grasper has been developed that will coapt both leaflets and fasten the structures with a graduated spiral screw. METHODS: Eleven explanted adult human MVs were mounted in a mock circulatory loop created for simulating a variety of hemodynamic conditions. The MV grasper was used to place a screw in each valve, which was then continuously run for 300,000 to 1,000,000 cycles with a fixed transvalvular pressure gradient. At the completion of these studies, the valves were stressed to a maximal transvalvular gradient for ten minutes. In seven cases, MR was induced and subsequently repaired using the MV screw. In vivo, the MV screw was tested in nine male canines. Through a subcostal incision, the MV grasper entered the left ventricle, approximated the mitral leaflets and deployed the MV screw under direct visualization via an atriotomy. Follow-up transthoracic echocardiograms were done at postoperative week 1, 6, and 12 to identify screw migration, MV regurgitation/stenosis or clot formation. Dogs were sacrificed up to postoperative week 12 to allow gross and histologic assessment. RESULTS: In vitro, no MV screw detached from the valve leaflets or migrated during the durability testing period of 6.8 million cycles, including periods of stress load testing up to 350 mm Hg. The percent regurgitant flow used to assess MR statistically decreased with the placement of the screw from 72 +/- 7% to 34 +/- 17%; p = 0.0025. In vivo, seven dogs whose valves were examined within the first 48 hours revealed leaflet coaptation with an intact MV screw and no evidence of MR. Two dogs, followed for a prolonged period, had serial postoperative echocardiograms demonstrating consistent coaptation, no screw migration, no clot, and no regurgitation or stenosis. In the animal sacrificed at 12 weeks, the MV screw was integrated into the tissue of both leaflets. CONCLUSIONS: The MV screw has provided durable leaflet coaptation and has reduced regurgitation in human MVs. Initial data on the MV screw's biocompatibility and interactions with living valve tissue is promising. Our early success supports further efforts towards the maturation of this prototype into off bypass mitral valve repair technology. PMID- 11276468 TI - Flow measurement in coronary surgery. AB - BACKGROUND: Many of the modern less invasive approaches to coronary artery bypass grafting (CABG) are performed without the use of the heart lung machine and cardiac asystole. Even after the introduction of mechanical stabilizers, the ability to achieve a technically perfect anastomosis is less certain in beating heart bypass surgery. Our group has begun to assess the surgical results of beating heart CABG using Transit Time Flow Measurement (TTFM). Our experience indicates that a meticulous and controlled method of assessing the results of intraoperative flow measurements can improve the quality of information and increases the accuracy of diagnosing technical problems with newly constructed bypass grafts. For this reason, we developed a standard algorithm for using and interpreting intraoperative TTFM. METHODS: From January to August of 1998, 161 patients underwent off-pump CABG with a total of 323 distal anastomoses (2.0 grafts per patient). All completed grafts were tested intraoperatively with TTFM and the decision to accept or revise any individual graft was based on a decision nomogram using key values readily available from the TTFM output. RESULTS: Thirty two grafts (9.9%) were surgically revised based on unsatisfactory flow curves, the Pulsatile Index, or both. All revised grafts were found to have a significant technical error, such as an intimal flap, thrombus, conduit kinking, or dissection. There were no major complications, myocardial infarctions, or deaths in the entire series of patients. CONCLUSIONS: Based on our favorable use of TTFM, we strongly recommend that patency of every graft be assessed whether the operation is performed off pump or on cardiopulmonary bypass. Guidelines for performing and interpreting TTFM ensure a high degree of confidence in the completed graft. The decision to revise a graft can be made based on simple parameters easily acquired from the TTFM device. Any concern about quality or quantity of flow should prompt immediate revision. PMID- 11276469 TI - Ascending aortic atherosclerosis--a complex and challenging problem for the cardiac surgeon. AB - Ascending aortic atherosclerosis is an increasingly recognized problem in cardiac surgery. It is the most important risk factor for perioperative stroke and seems to be in part responsible for postoperative neurobehavioral changes. Patients exhibiting ascending aortic atherosclerosis have a significantly reduced survival rate and are at considerable risk for spontaneous embolic stroke during the long term postoperative course. Preoperative noninvasive diagnosis and intraoperative assessment by inspection or palpation of the aorta are insensitive. Intraoperative epiaortic ultrasound scanning has emerged as a most helpful tool for the diagnosis of ascending aortic atherosclerosis and has revealed major insights into the nature and distribution of this disease. Management strategies range from minimally invasive aortic "no touch" techniques to maximally invasive procedures, including application of deep hypothermic circulatory arrest. Operative modifications in coronary artery bypass grafting include avoidance of aortic crossclamping, alternative methods of aortic crossclamping and placement of all arterial in situ bypass conduits, Y-grafts or extra-anatomical bypass grafts. Other operative strategies include modifications of the arterial cannulation site, replacement of the ascending aorta or ascending aortic endarterectomy. One of the most recently developed methods is embolic capture by intraaortic filters. Increased awareness of ascending aortic atherosclerosis is critical in order to prevent the devastating complications it can cause during cardiac surgery. PMID- 11276470 TI - A method of endoscopic investigation of vascular structures directly through flowing blood. AB - BACKGROUND: Noninvasive cardiovascular diagnosis has improved immensely due to key technological refinements such as digital subtraction angiography, ultrasonography, Doppler flow analysis, and magnetic resonance imaging. Each of these methodologies provides a unique image of the cardiovascular system but will not permit surgical maneuvers or repairs during real time imaging. Our group has developed a new method of endoscopic visualization of the luminal surface of blood vessels directly through flowing blood without interference of the blood or vessel wall. This opens new possibilities in both diagnosis and surgical interventions. METHODS: Transluminal imaging through flowing blood was performed in normal animals using laser frequency light delivered and retrieved via conventional fiberoptic angioscopic instruments. The reflected light energy was reconstructed into a viewable image using a specialized method of optical data processing and filtering systems. Unlike conventional angioscopy, displacement of flowing blood was not needed as the images were obtained with higher frequency laser light. RESULTS: A total of 20 canine experiments were performed between 1996 and 1997 using our endoluminal imaging system. The images obtained revealed details of luminal surfaces, although primitive and low resolution with this first generation of technology. Images of the topography of the femoral, axillary, and subclavian arteries and veins, as well as several intracardiac structures (aorta and aortic valve) were successfully obtained without trauma or physiologic consequence to the animal. CONCLUSIONS: Using conventional fiberoptic angioscopes coupled with laser light of differing wavelengths, it was possible to image the interior of vascular structures through flowing blood. This method visualizes the intraluminal surface in real time and is dependent only on the delivery and capacity of the endoscope. The implications for future cardiovascular diagnosis and corrective surgical procedures are widespread. PMID- 11276471 TI - Minimally invasive replacement of ascending aortic aneurysms: intermediate term results. AB - BACKGROUND: Minimally invasive techniques have gained recent interest in the realm of cardiac surgery. This report describes our initial experience with graft replacement of ascending aortic aneurysms using a superior mini-sternotomy approach. METHODS: Between March 1997 and October 1997, four patients underwent operation for ascending aortic aneurysm via superior mini-sternotomy approach. There were two female and two male patients, ranging in age from 52 to 62 years (mean 53.7 +/- 7.6). All patients had the stigmata of Marfan's syndrome. Mean diameter of the ascending aortas was 6.1 +/- 0.9 cm. Composite graft replacement with coronary reimplantation was performed in all cases. In one patient hemiarch replacement was performed under total circulatory arrest. There was no hospital (30-day) mortality. Mean aortic cross clamp and cardiopulmonary bypass times were 63 +/- 14.1 minutes (range 44 to 78) and 116.7 +/- 43.3 minutes (range 81 to 177), respectively. One patient was re-explored for bleeding. RESULTS: Lengths of hospital stay ranged from 5 to 7 days (mean 5.5 +/- 1). Patients were followed-up for at least 18 months. One patient suffered a fatal stroke in her third postoperative month. All surviving patients were in NYHA Class I at the sixth postoperative month and thereafter. CONCLUSIONS: Minimally invasive graft replacement of ascending aortic aneurysms can be performed safely and effectively. Long term results are likely to be similar to those of conventional cases performed through a full median sternotomy. PMID- 11276472 TI - Aortic homograft root replacement for failed freehand homograft aortic valve: effectiveness of a collagen/thrombin/plasma composite hemostat in the setting of technically complicated homograft to root anastomosis. PMID- 11276474 TI - Non steroidal anti-inflammatory drug-based pain control for minimally invasive direct coronary artery bypass surgery. AB - Minimally invasive direct coronary artery bypass (MIDCAB) surgery has become an attractive alternative technique to treat coronary artery insufficiency. Changes in surgical and anesthesia techniques have led to reduced pulmonary morbidity associated with the operation. Early extubation is typically expected. However, postoperative pain management becomes even more important with early extubation. We describe our technique of a NSAID-based protocol with indomethicin and Torodal that has been safe and effective in over 175 patients following MIDCAB. PMID- 11276473 TI - Endarterectomy for preventing stroke in symptomatic and asymptomatic carotid stenosis. Review of clinical trials and recommendations for surgical therapy. AB - BACKGROUND: Multicenter, randomized trials have demonstrated advantages for surgery over medical therapy in both symptomatic and asymptomatic carotid stenosis of greater than 70%. Controversial interpretations of these trials are debated between medical and surgical camps. The goal of this review is to summarize the current state of knowledge in carotid stenosis and the role of surgery and several advances in operative management. METHODS: Summaries of seven major controlled trials of carotid endarterectomy versus medical therapy are presented along with supportive data from over 90 related publications. Criticisms, deficiencies as well as strengths are offered. RESULTS: All studies in which trial design, clinical variables, case selection, complication definition, and patient follow-up were well conceived and performed showed statistically significant advantages for surgical therapy within a remarkable short interval of follow-up (less than 3 years). Carotid endarterectomy demonstrated a two to four fold reduction in the late incidence of stroke when compared to optimum medical management (risk factor reduction and initiation of antiplatelet therapy). Reduction in stroke risk over time remains stable in surgically treated patients whereas medically treated patients clearly show progression of stenosis and evolution of new neurologic events with time. Several studies indicate that diabetes is a risk factor for stroke with medical therapy that is eliminated by surgical therapy. Advantages were more clearly demonstrated when symptomatic patients (TIAs, stroke, or amaurosis) were studied, but asymptomatic patients received significant benefit as well. The degree of benefit measured was in direct balance to the perioperative risk. Perioperative stroke and death rates must be low (less than 3% combined for asymptomatic patients) in order for statistically significant differences to be detected. However, most centers now can perform carotid endarterectomy within these outcome parameters. CONCLUSIONS: Randomized trials support the safety and efficacy of carotid endarterectomy for stenosis greater than 70% (with or without symptoms). Advantages of surgery over medical therapy were found in less than three years and there is ample evidence to suggest that the differences between these groups would have been even more pronounced had longer follow-up been obtained. Thus for patients who face many years of risk after diagnosis of a carotid lesion, early surgery is the most important and effective intervention for preventing stroke. The results of these trials raised initial concern over increasing health care expenditures from rising surgical case volumes. However, studies of cost effectiveness confirmed that surgery saves health care dollars when compared to the long term care of stroke victims. PMID- 11276475 TI - Endoscopic coronary artery bypass grafting on the beating heart using a computer enhanced telemanipulation system. AB - OBJECTIVE: To develop a technique for computer enhanced endoscopic arterial bypass grafting on the beating heart in an experimental canine model. METHODS: Mongrel dogs (30-35 kg) were used. After single lung ventilation of the right lung was initiated the dogs were placed to the right. The videoscope and the end effectors of the da Vinci telemanipulation system (Intuitive Surgical, Mountain View, CA) were introduced through three ports. Surgery was performed remotely from the console (motion scaling 3:1). After harvesting of the internal thoracic artery (ITA) and preparation of its distal end, the Endostab, a newly developed endoscopic stabilizer, was introduced through an additional port. The anterior wall of the heart was stabilized and the collateral branch (RC) of the left anterior descending artery (LAD) was ligated proximally and distally. The arteriotomy was made and the ITA-graft anastomosed to the RC. The animals were sacrificed and the grafts were checked for patency using bench angiography. RESULTS: In two of five dogs total endoscopic beating heart bypass grafting, including ITA harvest, stabilization, arteriotomy and performance of the anastomosis, was successfully performed using computer enhanced technology and a new endoscopic stabilizer. In two dogs the procedure was completed with femoro femoral cardiopulmonary bypass (CPB) support on the beating or fibrillating heart, respectively. One dog expired due to VT. Hemodynamically, endoscopic stabilization was well tolerated. All four grafts were patent despite a target vessel diameter of less than 1 mm. CONCLUSION: The endoscopic stabilizer can sufficiently immobilize the heart to enable endoscopic beating heart coronary artery bypass grafting by means of a computer controlled instrumentation system. PMID- 11276476 TI - Total inversion of the left lung circulation: morphologic and functional analyses. AB - BACKGROUND: An experimental model for total inversion of left lung circulation was developed. With this model, the authors demonstrate that it is possible to reverse the pulmonary circulation and preserve the normal function and morphology of the lung. METHODS: Eight dogs had their left pulmonary circulation reversed. The blood from the pulmonary artery trunk was diverted to the pulmonary veins, and returned from the pulmonary artery into the left atrium. In order to monitor the flow through the reversed system, color Doppler echocardiography was performed on the ninth postoperative day. The dogs were reoperated after 15 days for re-evaluation. Blood gas analyses from the aorta and the pulmonary artery were used to study the functional status of the lung in both operations. The morphology was studied by comparing biopsies of the lung performed before and after reversal of flow. RESULTS: Blood gas analysis showed no significant difference between the samples of from the aorta and pulmonary artery. Color Doppler echocardiography was a reliable method for the study of the inverted circulation. The histological study showed no differences in the morphology of the lung after the reversed circulation. CONCLUSIONS: Left pulmonary circulation was fully reversed. Pulmonary function and morphology remained normal. PMID- 11276477 TI - Minimally invasive port access surgery reduces operative morbidity for valve replacement in the elderly. AB - BACKGROUND: Although minimally invasive techniques for valvular surgery have rapidly come into widespread use, whether such an approach can be safely applied to elderly patients remains an open question. To help resolve this issue, we reviewed our experience with minimally invasive port access (MIPA) valve surgery in elderly patients and compared it to the results obtained with the standard sternotomy (STD) approach in the same age group. METHODS: From January 1994 through December 1998, 370 consecutive patients at least 70 years of age underwent isolated aortic or mitral valve surgery at our institution. The standard sternotomy operative approach was used in 259 patients (mean age 77.5 years) and the minimally invasive port access approach was used in 111 patients (mean age 76.0; p=.006). A mitral valve procedure was performed more often in the MIPA patients than in the STD patients (49.5% vs. 35.9%; p < .001). RESULTS: Hospital mortality was comparable in the two groups, 9.7% (25/259) in the STD group and 7.2% (8/111) in the MIPA group (p = .50), as was the incidence of many perioperative complications. The MIPA group, however, had a significantly lower incidence of sepsis or wound complications (1.8% vs 7.7%; p = .027), required less fresh frozen plasma transfusion (median 1.0 unit vs 2.0 units; p =.04), and had a shorter length of hospital stay (11.6 days vs 17.6 days; p = .001). CONCLUSIONS: These results indicate that with appropriate surgical techniques the MIPA approach for isolated valve surgery can be safely applied to the elderly patient population with excellent results. In our initial experience the MIPA approach is associated with significantly less plasma transfusion, fewer postoperative complications, and shorter length of hospital stay. PMID- 11276478 TI - Clinical outcomes and angiographic patency in 125 consecutive off-pump coronary bypass patients. AB - BACKGROUND: This study compared clinical outcomes, length of stay, and hospital costs in patients having off-pump coronary bypass (OPCAB) versus conventional bypass surgery (CABG). METHODS: From November 1996 through April 9, 1999, OPCAB was performed for 125 consecutive patients and compared with a contemporaneous, matched control group of 625 CABG patients. Patients were matched according to age, gender, incidence of renal failure, diabetes, pulmonary disease, stroke (CVA), hypertension, peripheral vascular disease, and previous myocardial infarction. Follow-up in the OPCAB patients was 100% and averaged 15 months. RESULTS: An average of 2.0 grafts per patient were performed in the OPCAB group (range 1-5). Ninety-four OPCAB patients (75.2%) had a total of 179 grafts assessed angiographically prior to hospital discharge. All but 4/179 grafts (2.2%) were patent, including 94 of 94 IMA grafts (100%). There were no in hospital deaths in the OPCAB group compared to a mortality rate of 1.4% in the CABG group. OPCAB reduced postoperative hospital stay from 5.5 days in the traditional CABG group to 3.3 days (p=.002), with a decrease in hospital cost of 24% (p = .01). In addition, there was a significant reduction in the rate of transfusion in the OPCAB group (29.6%) compared to the CABG group (56.5%, p = .0001). Two OPCAB patients required postoperative intervention to improve graft patency during the follow-up period. No internal mammary grafts required revision. There was one perioperative CVA and one myocardial infarction in the OPCAB group. CONCLUSIONS: OPCAB surgery reduces hospital cost, postoperative length of stay, and transfusion rate compared to CABG. OPCAB is safe, cost effective, and associated with excellent graft patency and clinical outcomes. PMID- 11276480 TI - A review of transit-time flow measurement for assessing graft patency. AB - Anastomotic quality is a critical issue in minimally invasive coronary artery bypass surgery, particularly "off-pump". It is important to detect a "poor" anastomosis during the procedure so future re-operation can be avoided. Methods such as intraoperative angiography, thermal angiography, probing of the anastomosis, and graft flow measurement have been used intraoperatively to help identify anastomotic errors. With the evolution of stabilizers, graft patency rates for off-pump cases have improved, but many believe they are still not as high as those of the conventional procedure. For off-pump surgery to be accepted and practiced universally, patency rates must be equivalent to those of "on-pump" cases. Transit-time flow measurement has become an increasingly popular non invasive method for assessing anastomotic quality. However, it is difficult to establish whether an anastomosis is patent based on mean graft flow alone. Spectral analysis of graft flow waveforms reveal characteristic patterns that identify intermediate ranges of stenosis between fully patent and totally occluded. Together, these two components of graft flow have been used in the construction of a neural network to help identify "faulty" anastomoses. Transit time flow measurement is a non-invasive tool that can be beneficial in identifying fully patent or nearly occluded grafts, and may also help in distinguishing intermediate stenoses. PMID- 11276479 TI - Contrast-enhanced magnetic resonance angiography for control of minimally invasive coronary artery bypass conduits (MIDCAB/OPCAB). AB - OBJECTIVE: The purpose of this study was to delineate the course and determine the patency of venous and arterial conduits in the early postoperative period following minimally invasive bypass grafting. A less invasive magnetic resonance angiogram was evaluated as alternative to standard contrast angiography and cardiac catheterization. METHODS: Twelve patients (8 males and 4 females) with a mean age of 65.3 (+/- 7.4 ) years were evaluated four to seven days following minimally invasive direct coronary artery bypass surgery (MIDCAB) or off-pump multivessel revascularization with the Octopus stabilizer on the beating heart. Altogether 17 coronary bypass grafts were investigated: 12 left-sided mammary artery grafts to the LAD and five aortocoronary venous bypass grafts. The examination was performed with a 1.5 Tesla Magnetom Vision (Siemens AG, Erlangen) with phased array coil technology. Data acquisition was done with an ultrafast 3D gradient-echosequence in single breathhold and sagittal and coronal views. Contrast enhancement of the vessels was performed with automatic intravenous bolus injection of Gadolinium-DTPA after determination of the individual contrast transit time. Traditional contrast angiography was obtained in all patients during the same time period as a comparison to assess the sensitivity and specificity of the magnetic resonance imaging. RESULTS: All five venous grafts and 11 of the 12 IMA grafts were detected and shown to be patent with the MRA technique. Contrast angiography demonstrated complete patency for all 17 bypass grafts with adequate anastomoses and no evidence of stenosis. The calculated sensitivity for the visualization with MRA was therefore 92% for IMA grafts and 100% for venous grafts. CONCLUSION: The contrast-enhanced ultrafast MRA in single breathhold technique is a reliable, noninvasive method for visualization and determination of the patency of arterial and venous coronary grafts. PMID- 11276481 TI - Noninvasive graft flow and patency assessment following minimally invasive direct coronary artery bypass (MIDCAB) grafting. AB - OBJECTIVE: Assessment of graft patency following minimally invasive direct coronary artery bypass (MIDCAB) surgery is essential in order to determine the efficacy of this technique. This study was conducted to evaluate the role of intraoperative and postoperative noninvasive flow and velocity measurements to follow and predict graft performance. METHODS: Between April 1996 and July 1997, 130 patients had 133 grafts placed using MIDCAB techniques. Intraoperative transit-time ultrasound was used to assess graft patency and flow prior to wound closure. Also, serial transcutaneous doppler examinations were performed to evaluate graft patency on the first postoperative day, at two weeks, and at three months. Peak values for systolic and diastolic waveforms were measured for both flow and velocity, and the diastolic-to-systolic ratio was calculated at each time interval. Recatheterization was performed selectively for inadequate ultrasound flow or doppler velocity, or for patient symptoms. RESULTS: Seven (5.3 %) grafts developed stenosis or occlusion. When compared to normal grafts, mean intraoperative flows, flow ratios, and velocity ratios were lower. Mean postoperative diastolic peak velocity (DPV) to systolic peak velocity (SPV) ratio remained stable over time for normal grafts; however, grafts with stenosis or occlusion demonstrated a diminished DPV/SPV ratio. CONCLUSIONS: Intraoperative transit-time ultrasound and outpatient transcutaneous doppler examinations did not reach a predictive value for graft stenosis or occlusion following MIDCAB surgery in this series of patients. However, these data demonstrate trends that may help identify patients at an increased risk for unfavorable events, guiding the use of postoperative recatheterization in such patients. PMID- 11276482 TI - Intraoperative flow rate measurements of T-grafts: calculating a flow reserve. AB - OBJECTIVE: To evaluate the inflow of the left internal thoracic artery (LITA) and the effect of adding a radial artery T-graft to distal LITA flow, and to calculate the LITA flow reserve. METHODS: Twenty-two patients underwent myocardial revascularization using the radial artery-LITA T-graft in which intraoperative flow measurements were recorded. An ultrasonic flowmeter was used to directly measure flow rates in the T-graft: 1) before completion of the distal anastomoses to measure maximum flow rates (free flow), and 2) after completion of distal anastomoses. RESULTS: The mean free flow rates of the LITA alone, radial artery graft alone, and T-graft (total flow) were 104 +/- 70, 151 +/- 89, and 230 +/- 102 ml/min, respectively. The mean flow rates on bypass of the distal LITA, radial artery graft, and T-graft after the distal anastomoses were completed were 24 +/- 16, 32 +/- 27, and 63 +/- 29 ml/min, respectively. The mean T-graft flow off bypass was 66 +/- 29 ml/min. The mean flow reserve was 70%. CONCLUSION: The LITA has a flow reserve by which proximal flow rates will increase to accommodate the addition of a radial artery T-graft without compromising LITA flow distal to the T anastomosis. PMID- 11276483 TI - Steal syndrome documented by transit time flow measurement technique in an "H" Graft. PMID- 11276484 TI - Incidence and predictors of tias and strokes following coronary artery bypass grafting: report and collective review. AB - BACKGROUND: Neurologic complications account for some of the most devastating problems following coronary artery bypass surgery. In this study we determined the incidence and predictors of perioperative transient ischemic attacks (TIAs) and strokes in patients undergoing coronary artery bypass grafting at our institution. METHODS: Data was prospectively collected from 4,518 consecutive patients undergoing isolated coronary artery bypass grafting at Brigham & Women's Hospital between 1993 and 1997. RESULTS: One hundred and twenty of the 4,518 patients sustained either a TIA (30 patients, 0.7%) or a stroke (90 patients, 2.0%), for an overall incidence of 2.7%. Significant univariate predictors of TIA/stroke included a history of: 1) cerebral vascular disease, 2) peripheral vascular disease, 3) diabetes, 4) renal failure, 5) preoperative myocardial infarction, 6) hypertension, and 7) age > 70 years. Multivariate logistic regression analysis revealed the following significant associations (incidence of TIA/stroke, odds ratio): 1) cerebral vascular disease (6.4%, OR 2.5); 2) peripheral vascular disease (5.3%, OR 1.6); 3) renal failure (5.6%, OR 1.6); 4) myocardial infarction (3.2%, OR 1.5); 5) diabetes (3.7%, OR 1.5); 6) age > 70 (3.5%, OR 1.5). Perioperative TIA/stroke was significantly associated with postoperative low cardiac output and atrial fibrillation. Patients with TIA/stroke had a significantly longer ICU stay (4 vs. 2 median days), length of hospitalization (14 vs. 7 median days), and higher mortality rate (22% vs. 2.6%). CONCLUSIONS: Perioperative TIA/stroke occurred in less than 3% of patients following coronary artery bypass grafting but was associated with significant mortality. The strongest predictors were cerebral and peripheral vascular disease. PMID- 11276485 TI - Changing referral pattern in off-pump coronary artery bypass surgery: a strategy for improving surgical results. AB - BACKGROUND: We have previously shown that a less invasive surgical approach (LISA) can reduce mortality and morbidity in coronary artery bypass grafting (CABG). This appears to have led to the referral of increasingly high risk patients for this procedure as compared to patients undergoing traditional CABG. The purpose of this paper is to compare preoperative risk factors and postoperative complications in both LISA and conventional CABG cases using the New York State database. METHODS: From January 1997 to September 1998, 1,993 patients underwent CABG in our institution: 1,384 with CPB (group A) and 609 without CPB (group B). In group B (LISA), a well defined strategy was followed in an effort to prevent hemodynamic instability during coronary exposure, avoid myocardial ischemia, verify graft patency, and use alternative surgical incisions in reoperations. RESULTS: Analysis of preoperative risk factors using the NYS database showed a significant increase in comorbidities in group B (p < 0.005), while at the same time postoperative complications and risk-adjusted mortality were lower (p = NS). CONCLUSION: Our data demonstrates that by using the LISA, high risk patients can undergo CABG with equal or lower mortality and morbidity than traditional CABG. PMID- 11276486 TI - Minimally invasive coronary artery bypass grafting for myocardial muscle bridging. AB - Myocardial bridging is a congenital anomaly of the left anterior descending coronary artery (LAD), which is associated with myocardial ischemia and infarction, cardiac arrhythmias, and sudden cardiac death. Two cases are reported of symptomatic myocardial bridging refractory to medical management treated by minimally invasive coronary artery bypass grafting without cardiopulmonary bypass. We conclude that minimally invasive coronary artery bypass techniques are appropriate alternatives to endovascular stent placement, muscle bridge division, or aortocoronary grafting with cardiopulmonary bypass for the management of symptomatic myocardial bridging. PMID- 11276487 TI - Minimally invasive axillary-LAD saphenous vein bypass. AB - The left internal mammary artery (LIMA) is the arterial conduit of choice for minimally invasive coronary bypass to the left anterior descending (LAD). However, in redo cases when the LIMA is not available, the use of a saphenous vein graft as an extra-anatomic bypass from the axillary artery to the LAD offers a lower risk alternative than conventional reoperative trans-sternal surgery [Knight 1997]. We report on 3 patients who underwent axillary-LAD saphenous vein bypass. At six months, follow-up by Duplex ultrasound showed patent grafts in all three patients. PMID- 11276488 TI - Twenty-two years and it just keeps going, going, going: The Veterans Affairs Cooperative Study of Coronary Artery Bypass Surgery for Stable Angina. PMID- 11276489 TI - Comparison of MIDCAP versus conventional CABG surgery regarding pain and quality of life. AB - BACKGROUND: This prospective clinical trial focuses on pain and quality of life (QOL) after minimally invasive direct coronary artery bypass (MIDCAB) grafting versus conventional coronary artery bypass grafting (CABG). METHODS: Group A consisted of 65 consecutive MIDCAB patients using an anterolateral mini thoracotomy and the "off-pump" technique. Group B consisted of 95 computer matched patients who underwent conventional CABG with cardiopulmonary bypass (CPB). Pain was graduated using the visual analog scale (VAS), and the verbal rating scale (VRS) [Troidl 1990]. QOL was evaluated at the time of discharge and three months after surgery using modified Nottingham Health Questionnaires that separate physical, social, activity, emotional, pain, and sleeping conditions. RESULTS: Postoperative pain was higher after MIDCAB on postoperative day (POD) 1 (p< 0.002). From POD 4 onwards MIDCAB patients had less pain compared with the conventional group (p<0.04). MIDCAB patients required less pain medication from POD 4 onwards (p<0.05). QOL was significantly better in the MIDCAB group on POD 7 for physical (p< 0.038), activity (p< 0.016), pain (p< 0.041), and sleep (p<0.038) conditions. The three-month questionnaire showed significantly better levels for MIDCAB patients regarding physical (p< 0.03) and pain (p< 0.001) conditions, and a trend for activity (p< 0.08) and emotional (p<0.08) conditions. CONCLUSION: Compared to patients undergoing conventional surgery, MIDCAB patients suffer more pain in the first three postoperative days, probably as a result of the lateral thoracotomy. From POD 4 onwards, MIDCAB patients are significantly better, experiencing less pain and showing better physical, activity, and sleeping conditions even three months after surgery. This can be attributed to the absence of median sternotomy and/or the avoidance of cardiopulmonary bypass. PMID- 11276490 TI - Coronary artery bypass grafting using the Rama technique, a method of coronary stabilization: short-term results. PMID- 11276491 TI - Use of a pulsatile beating heart model for training surgeons in beating heart surgery. AB - BACKGROUND: Coronary artery bypass on the beating heart has undergone resurgence with the introduction of minimally invasive techniques and new stabilizing devices. It is important to develop a method for training surgeons to perform accurate anastomoses despite cardiac motion and to develop the skills needed for consistent results in this demanding field. METHODS: A prosthetic model of the beating heart was created by Limbs and Things, Ltd. (Bristol, UK) and used in our center to simulate clinical situations of beating heart surgery. Anastomotic quality was evaluated using a pre-established set of criteria on patency and suturing with each anastomosis graded on a 12-point scale. RESULTS: The average scores for trainees using the Pulsatile Beating Heart Model were 8.5 while that of the expert surgeon with MIDCAB experience was 11. Defects seen included cross wall suturing and significant narrowing of the toe of the anastomosis. Scores improved with increasing practice during each session. Operators with more clinical experience scored higher. All surgeons felt the model duplicated the exposure and feel of the tissue characteristic of clinical cases. CONCLUSIONS: The beating heart simulator provides excellent training for new as well as experienced surgeons, provides visual feedback of anastomotic errors, and instills increasing confidence in the participants in their ability to construct accurate anastomoses on the beating heart. PMID- 11276493 TI - Endoscopic coronary artery bypass graft (ECABG) procedure with robotic assistance. AB - BACKGROUND: Technical details of the Robotically assisted endoscopic coronary artery bypass graft (ECABG) procedure on the cadaver model are reported. Moreover, this study will provide essential techniques, steps, and procedural development concepts necessary to introduce the ZEUS Robotic Surgical System (Computer Motion, Inc., Goleta, CA) into the human operating room. METHODS: Between August 1998 and March 1999, an ECABG procedure was performed on 10 cadaver torsos. The cadaver torso was placed in the left anterior oblique (LAO) position. The left and right internal mammary arteries (IMA) were taken down endoscopically. The ends of the IMA's were intracorporeally prepared. An upper partial sternotomy was demonstrated for perfusion cannulation and proximal anastomoses for multiple vessel revascularization. An arteriotomy was created with an endoscopic scalpel. The IMA was anastomosed to a chosen coronary using robotic assistance. The patency was verified by probing and injecting of methylene blue. RESULTS: Templates were developed to determine the placement of the robotic arms. Port templates were developed to both harvest the IMA's and perform the desired anastomoses. The following vessels were accessed through the developed port templates and retraction of the heart: left anterior descending (LAD), right coronary artery (RCA), diagonal (D(1)), obtuse marginal (OM(1) and OM(2)), and posterior descending artery (PDA). CONCLUSIONS: The use of robotic assistance during an ECABG procedure on a cadaver model is feasible. This study is a necessary and useful progression from the use of robotics in the animal lab to the use of robotics in the human operating room. PMID- 11276492 TI - Multivessel minimally invasive coronary surgery with endoscopic support. AB - BACKGROUND: Interest in minimally invasive coronary artery bypass (MICAB) grafting and the MICAB experience have been increasing. The purpose of this study was to develop the multivessel minimally invasive coronary revascularization technique and to estimate the effectiveness of the endoscopic support in this operation. METHODS: From January 1998 through April 1999, 190 patients (ages 38 to 72 years) underwent coronary revascularization without cardiopulmonary bypass. Among them, 69 patients (55 males, 14 females) underwent minimally invasive coronary revascularization, from 1 to 3 vessels, through minithoracotomy and ministernotomy with endoscopically dissected internal mammary artery, gastroepiploic artery, and composite grafts. Preoperative risk factors included unstable angina (n = 15), reoperations (n = 8), low ejection fraction (n = 14), renal insufficiency (n = 4), chronic obstructive pulmonary disease (n = 6), cerebrovascular accident (n = 2), diffuse atherosclerosis (n = 4) and diabetes mellitus (n = 7). RESULTS: The operative mortality was 1.5% (1/69). Morbidity included wound infections (n = 1), reoperation for management of bleeding (n = 1), acute graft occlusion (n = 1), perioperative myocardial infarction (n = 1). The number of grafts placed in 69 patients was as follows: single, 54; double, 10; triple, 5. Postoperative angiography and Doppler flow assessment of the coronary anastomoses performed in 22 patients (30%) showed that 97% were patent. CONCLUSIONS: The minimally invasive direct coronary artery bypass grafting operation is safe and effective. Endoscopic support makes the use of minimally invasive technology possible in patients with multivessel coronary disease and makes this operation less traumatic. PMID- 11276494 TI - 3D-visualization improves the dry-lab coronary anastomoses using the Zeus robotic system. AB - BACKGROUND: Robotic surgical instruments enable quick and precise movements and may allow complete endoscopic coronary artery bypass grafting. However, cardiac surgeons will have to become familiar with this technology and endoscopic viewing. We present our training program with special focus on 2D- and 3D visualization. METHODS: A thoracic skeleton, covered with a neoprene suit, served as model for the chest wall. Either a glove, fixed on a metal plate, or a pig heart were placed inside for training. On the glove, a suture line consisting of two lines of 16 points each, with a distance of 2 mm between each point, was stamped. On the pig heart, the LAD was prepared and incised; subsequently an anastomosis was done using the dissected right coronary artery as a graft. The time required was measured for both models. For suturing, the Zeus System (Computer Motion, Goleta, CA) was used and the third robotic arm positioned the endoscopic camera. The scopes were connected to a 3D-camera and the picture was displayed on a headset with two integrated monitors. Visualization was set to either 2D or 3D. Three surgeons were involved in the study. Each one did at least 12 anastomoses on 2D and 3D. RESULTS: The three surgeons involved showed a clear and rapid learning curve. The times required for the suture line decreased from 12.5 +/- 1.6 to 8.5 +/- 0.5 minutes with 2D and from 11.9 +/- 5.4 to 7.8 +/- 0.5 minutes for 3D respectively. This decrease did reach statistical significance (p = 0.03). In the pig heart model, the anastomosis times decreased from 33.2 +/- 8.4 to 15.7 +/- 0.3 minutes with 3D-visualization, and from 36.2 +/- 2.2 to 29.5 +/- 3.3 minutes with 2D. The decrease in anastomosis time did again reach significance (p = 0.025). At the end of the study, the times achieved with 2D visualization were significantly longer than those with 3D (p = 0.01). CONCLUSIONS: A surgical training program is mandatory to become familiar with these new technologies. Both models showed learning curves over an acceptable time course. 3D-visualization facilitated quick and precise movements, thus resulting in shorter anastomosis times. PMID- 11276495 TI - Minimally-invasive aortic root replacement. AB - PURPOSE: We retrospectively analyzed our early results with minimally invasive aortic root replacement. METHODS: Between August 1996 and April 1999, our center performed 137 aortic root replacements. Thirty-seven (27%) were accomplished through a 5 to 8 cm minimally invasive upper hemi-sternotomy incision. All minimally invasive operations were elective. The mean age for this cohort was 46 +/- 12 yrs. Thirty one (84%) of the patients were male and 3 (8%) were reoperations. The average preoperative NYHA classification was 2.4 +/- 0.6 and ejection fraction (EF) was 58% +/- 12%. Valve pathology was congenitally bicuspid in 19 (51%), endocarditis (SBE ) in 5 (14%), calcific degeneration in 4 (11%), annuloaortic ectasia in 3 (8%), rheumatic in 2 (5%) and other etiologies in 4 (11%). Nine patients (24%) had associated ascending aortic or arch aneurysms. RESULTS: The surgical techniques performed through mini-hemisternotomy consisted of 1) full root replacement in 31 (84%), 2) subcoronary replacement in 4 (11%), and 3) hemiroot in 2 (5%). Valve implants consisted of a homograft in 30 (81%), "Freestyle" bioprosthesis in 4 (11%) and a St Jude valved conduit in 3 (8%). Mean cardiopulmonary bypass duration was 193 +/- 47 min. and aortic cross-clamp duration was 157 +/- 40 min. Myocardial protection included systemic hypothermia in all (24 +/- 4 degrees C), antegrade cardioplegia (CP) in 35 (95%) with supplemental retrograde CP in 23 (62%). Three patients (8%) experienced postoperative low cardiac output syndrome (LCO). There was one operative death (3%). There was one (3%) reoperation for bleeding and 13 patients (35%) required blood transfusions. New onset atrial fibrillation occurred in 7 patients (19%) and there were 3 (8%) minor complications. Hospital length of stay (LOS) was 6.7 +/- 4.3 days and LOS was less than 7 days in 29 patients (78%). CONCLUSIONS: Minimally invasive aortic root replacement is feasible for a broad range of aortic valve pathology, can incorporate full root, hemiroot and subcoronary techniques, can be used for homografts and "Freestyle" valves as well as valved conduits, and can be accomplished with acceptable morbidity and mortality. However, the operation takes longer through the smaller incision and therefore requires more careful attention to myocardial protection. PMID- 11276496 TI - Surgical treatment of patients with ischemic cardiomyopathy: the significance of right ventricular function. AB - BACKGROUND: Patients with ischemic cardiomyopathy (ICMP) awaiting heart transplantation (HT) have a high mortality rate, in part because of the lack of donor organs. Given this limitation, we propose to broaden the indications for coronary artery bypass grafting (CABG) in this group and to more accurately select patients with ICMP requiring myocardial revascularization or HT. In this study, we assessed the short and long-term results of CABG in patients with ICMP. We also assessed the role of the right ventricle and the diastolic function of both ventricles in patients with ICMP. Using this information, we propose indications for CABG and/or HT in patients with ICMP. METHODS: We analyzed 49 patients with ICMP undergoing workup as potential heart transplant candidates. The patients were divided into two groups. Group A included 19 patients submitted to isolated CABG based on the preoperative assessment of myocardial viability. Group B consisted of 30 patients determined to be best suited for HT (with five patients actually receiving a donor heart). All patients were assessed by radionuclide ventriculography (RVG) and functional testing in order to assess their myocardial viability. RVGs were obtained prior to coronary bypass as well as at two and twelve months postoperatively. RESULTS: Preoperative data in group A were: left ventricular end-diastolic dimension (LVEDD) 7.0+/-0.32 cm, left ventricular ejection fraction (LVEF) 24.2+/-2.6%, and right ventricular ejection fraction (RVEF) 32.4+/-2.6%. For Group B, LVEDD was 7.7+/-0.29 cm, LVEF was 22+/ 2.7%, and RVEF 26+/-2.6%. The operative mortality in group A was 16.6%. Three patients died in the early postoperative period, two of them due to acute cardiac failure, and one due to cerebral complications. The number of grafted arteries was 3.6+/-0.2. One year after coronary artery bypass, the resting LVEF increased to 33.3% (a 36% improvement compared with preoperative, p < 0.001). Three-year survival was 77% in group A and 26.8% in group B. Examination of the myocardial functional state after CABG showed that the LV diastolic and RV systolic parameters statistically improved. The decrease of RVEF was revealed in the orthostatic test in patients (group A) who died after CABG in the early postoperative period, and in group B that correlated with their parameters of the myocardial viability. CONCLUSIONS: CABG in the patients with ICMP significantly increases the functional reserves of the myocardium of both ventricles, mostly because of the improvement in the diastolic function of the LV. In the RV, the systolic function could restore even in the early postoperative period. Preoperative analysis of the parametric images made after orthostatic test and nitroglycerin intake allow prediction, with 85 % sensitivity and 95% specificity, of the areas of the myocardium which will restore their function immediately after CABG ("hibernated myocardium"). The recovery of regional wall motion amplitude and of the response of the myocardium to coronary revascularization could continue during the first year after revascularization ("stunned myocardium "), most often beginning in the LV. The decrease of the EF of the RV as a response to the "unloading" tests could be a result of the disturbance of the ventricles' interaction as well as the spreading of the scarring of the RV myocardium. This decrease could be a predictor of the possible development of the low cardiac output syndrome after CABG. PMID- 11276497 TI - Symptomatic aneurysm of a saphenous vein graft with compression of the right atrium. AB - A symptomatic aneurysm of a saphenous vein bypass to the right coronary artery in a 77-year-old female patient is presented. Surgical therapy included resection of the aneurysmal saphenous vein graft, reconstruction of the right atrium, and coronary artery bypass grafting (CABG) to the right coronary artery. PMID- 11276498 TI - Two new drainage tubes for minimally invasive cardiac surgery. AB - BACKGROUND: The limited exposure typical of new minimally invasive surgical approaches to cardiac surgery makes it difficult to utilize traditional equipment such as temporary pacing electrodes or the placement of drainage tubes into the lateral pleural space. To overcome these difficulties, we have developed specialized drainage tubes just for limited access cardiac surgery. METHODS: The first device is a drainage tube with temporary pacing function. It has three fixed electrodes and one free pacing wire incorporated into an elliptical, angled 28 Fr silicone drainage tube. This tube is placed in the space between the heart and the diaphragm. The two fixed electrodes provide epicardial contact for ventricular pacing. The third fixed electrode and one free pacing wire are for the purpose of atrial pacing (Japanese patent #2,701,135). The second device is a Y-shaped drainage tube. Frequently, the pleural space is opened during harvesting of an internal mammary graft and then insertion of a chest tube in the thoracic cavity becomes necessary. We developed a new Y-shaped tube where one segment is placed in the retrosternal space and one segment is placed in the pleural cavity. (Japanese Association of Intellectual Copyright #130,591) RESULTS: The drainage pacing device was used in 48 coronary artery bypass grafting (CABG) patients. Drainage function and pacing function were excellent in all patients. The bifurcated drainage device was used in 34 patients achieving effective drainage of both cavities without complication during or after removal. CONCLUSIONS: Due to the limited surgical exposure provided by the newer minimally invasive procedures in cardiac surgery, specialized equipment that can be inserted through small incisions needs to be developed. We report the development of a new drainage-pacing device as well as a bifurcated drain for simultaneous drainage of the mediastinal and pleural cavities. These devices have facilitated minimally invasive cases and were free of complications. PMID- 11276499 TI - [In search of genetic determinants of obesity]. AB - Common obesity is a multifactorial trait that lies at the interface between the biology of body energy regulation and the environment. Obesity involves genetic predisposition but also metabolic, hormonal, behavioural, social and cultural aspects. According to some estimates 40 to 70 percent of the variation in obesity related phenotypes is heritable. Mutations in a single gene cause only about 5% of cases. The number of genes and markers associated or linked with human obesity is more than 200. Interactions between multiple genes, environment and human personality, make the search for obesity-related genes especially challenging. PMID- 11276500 TI - [The role of leptin in human obesity]. AB - The mechanism involved in body mass regulation in humans includes genetic, environmental, and behavioural factors. Human obesity is usually associated with a positive energy balance. Genetic studies in obese mice have revealed the Ob. gene, its products leptin and the leptin receptor to be important factors in the regulation of both appetite and energy expenditure. Leptin is a 16-kilodaltons adipocyte-derived hormone -which circulates in the serum as the free and bound forms. The leptin serum level reflects the amount of energy stored in adipose tissue. Leptin acts through the leptin receptor, -which belongs to the cytokine - receptor family. In rodents as well as in humans, homozygous mutations in genes encoding leptin or the leptin receptor cause early-onset morbid obesity, hyperphagia, and reduced energy expenditure. Recent studies have demonstrated that Ob. gene expression is increased in human obesity. However, mutations of Ob. gene present in the mouse are rare in the human population. PMID- 11276501 TI - [Screening programme for hyperlipidemia in children and adolescents. Prophylactic aspects of atherosclerosis]. AB - Atherosclerotic cardiovascular diseases (CVD), mainly coronary heart disease (CHD) remain the leading cause of death in adult populations of many countries. The following risk factors for atherosclerosis were identified: hypercholesterolemia, hypertension, cigarette smoking and obesity. Scientific reports and epidemiological studies have shown that atherosclerosis begins in childhood. Therefore consensus was obtained that the earlier the prevention begins the better results are achieved. But there are many controversies around early identification of hypercholesterolemia in children. Three options were considered: cholesterol mass screening, selective cholesterol screening and no screening at all. The most acceptable is selective screening performed in children of high risk families (CVD or hypercholesterolemia in the family). It is recommended by the US Expert Panel for the National Cholesterol Education Program for Children and Adolescents (NCEP-Peds). According to the NCEP-Peds, screening should include the following groups: I) children whose parents or grandparents have a history of CVD (under the age of 55 years), 2) children whose parents have a raised blood cholesterol concentration (above 240 mg/dl), 3) children with negative or unknown family history, but having other risk factors (hypertension, obesity, cigarette smoking, high-fat diet). The experts recommend that the examination should be performed in children after the age of 2 years. The NCEP Peds guidelines set total cholesterol levels in serum for children and adolescents from families at risk, below 170 mg/dl, as acceptable. Total cholesterol level between 170 and 199 mg/dl is classified as borderline and 200mg/dl and above--as high. PMID- 11276502 TI - [Levels of lipid peroxides and of some antioxidants in placenta and cord blood of newborns whose mothers smoked during pregnancy]. AB - Cigarette smoking during pregnancy increases the risk of oxidative damage and induces not only intrauterine foetal growth retardation, but also causes disturbances in postnatal growth and development. In the presented studies oxidative damage was estimated through the measurement of lipid peroxides concentration and the level of some antioxidants in placenta and in cord blood of newborns whose mothers smoked during pregnancy. We observed that the concentration of lipid peroxides was higher in cord blood and in placenta tissue (8%) than in the newborns of non-smoking mothers, but the activity of superoxide dismutase and glutathione peroxidase were lower by 20% and 16% respectively. Plasma level of vitamin A (p<0.005), vitamin E (p<0.05), fS-carotene (p< 0.0001) and total plasma antioxidant capacity (p<0.05) were significantly lower in the newborn of smoking than the non-smoking mother group. It is suggested that placental tissue protected the foetus against oxidative stress, but not sufficiently. Antioxidant activities of cord blood may also be insufficient in dismutation of free radicals and their detoxication in order to protect newborns against smoking dependent metabolic disturbances. PMID- 11276503 TI - [Investigations of thyroid function in children and adolescents after treatment of Hodgkin's disease]. AB - Radiotherapy focused on the neck, upper mediastinum or cranium can raise the risk of thyroid dysfunction. The influence of radio- and chemotherapy on thyroid morphology and function after treatment of Hodgkin's disease in 31 children and acute lymphoblastic leucaemia in 36 children was evaluated. Clinical examination, estimation of blood TSH, FT4 levels as well as ultrasonographic scan were performed. In the group of children with Hodgkin s disease the blood level of ATG Antithyreoglobulins andMAB, microsmal antibodies were estimated. The results of clinical examination were normal in all patients. Thyroid function tests were also normal, except for one case with elevated TSH values (22.76 microIU/ml) and decreased FT4 values (0.65 ng/ml). Three patients (9.7%) with Hodgkin s disease and three (8.5%) with ALL (only one had cranial radiotherapy) had an abnormal image of thyroid in ultrasonography (hypoechogenicity, heterogenous ultrasound scan, solid nodule). The results of our investigations indicate that abnormalities found in the ultrasound scan in children and adolescents with Hodgkin's disease in whom treatment has been terminated, can be a first prodrome of thyroid pathology. PMID- 11276505 TI - [Prenatal diagnosis of a brain tumour--an example of diagnostic and therapeutical algorithm]. AB - The authors describe their cooperation in the diagnosis and treatment of a newborn with malignant brain tumour (rare case of carcinoma of the choroid plexus) recognised by means of prenatal sonography and magnetic resonance. The case history is an example of modern algorithm of diagnostic and therapeutic procedures in perinatal medicine and the necessary multicentre collaboration. PMID- 11276504 TI - [Effectiveness of the screening programme for galactosemia. New strategy in Poland]. AB - Galactosemia is an autosomal recessive disease related to deficiency of one of three different enzymes involved in the metabolism of galactose: galactokinase (GALK), galactoso-J-phosphate uridyltransferase (GALT) or UDP-galactose-4 epimerase (GALE). Classic galactosemia is due to GALT deficiency and is the most common. Longitudinal studies have shown that in spite of early diagnosis and early treatment of children with galactosemia detected in the mass screening programme, the results are poor and mental retardation as well as other complications are of similar severity as in children diagnosed clinically without screening. In many investigations it was also proved that some impairments developed already in the prenatal period. Therefore, many countries among them also Poland, stopped mass screening for galactosemia. At present, in Poland the procedure strategy in galactosemic children and their families include: diagnosis of new cases on the basis of clinical symptoms, selective screening in high-risk families, prophylactic lactose-free diet for mothers during pregnancy. Such management can help to prevent clinical manifestations in newborns and prevent death in the early period of life. PMID- 11276506 TI - [Diabetic patient and reproduction]. AB - Current views and opinions on pregnancy in diabetic mothers are presented. Special attention is paid to carbohydrate metabolism, pregestational diabetes mellitus (PGDM), gestational diabetes mellitus (GDM) and unclear heterogenous etiology of gestational diabetes mellitus (GDM). Suggestions for identification, diagnosis and treatment of GDM with an emphasis on early screening strategy are given. Obstetrical management and possible perinatal complications are discussed. PMID- 11276507 TI - [Standards for diagnosis and treatment of phenylketonuria]. AB - The aim of this study is to unify the diagnostic and therapeutic standards in phenylketonuria. The course of the disease, diagnostic methods as well as treatment procedures is presented. Standards of treatment are based on the experiences and research carried out by the Screening Tests Unit of the Department of Public Health and the Clinical Department of Paediatrics of the National Research Institute of Mother and Child in Warsaw. PMID- 11276508 TI - [Early diagnosis of inborn errors of metabolism. New technologies]. AB - Inborn errors of metabolism, in spite of their not very high incidence play more and more evident role in the causes of infant's mortality and morbidity. Therefore early diagnosis and early treatment becomes an important issue in contemporary medicine. The criteria for newborn mass screening, selective screening and supplementary screening as well as the new technologies, among others mass spectrometry, Tandem MS, were discussed. PMID- 11276509 TI - Focus on intrabony defects: guided tissue regeneration. PMID- 11276510 TI - The conservative approach in the treatment of furcation lesions. PMID- 11276511 TI - The role of resective periodontal surgery in the treatment of furcation defects. AB - Resective therapy for the treatment of furcation defects is an essential part of the periodontal therapist's armamentarium. Root resection can successfully treat specific furcation defects that cannot be solved by other surgical and nonsurgical approaches. Complications with these resective procedures are not rare but are usually avoidable when specific endodontic, surgical and restorative guidelines are followed. PMID- 11276512 TI - Focus on furcation defects: guided tissue regeneration. PMID- 11276513 TI - Focus on furcation defects--guided tissue regeneration in combination with bone grafting. PMID- 11276514 TI - Fundamental principles affecting the outcomes of therapy for osseous lesions. PMID- 11276515 TI - Biology of wound healing. PMID- 11276516 TI - Focus on intrabony defects--conservative therapy. PMID- 11276517 TI - Osseous resective surgery. AB - Osseous resective surgery necessitates following certain guidelines for proper recontouring of the alveolar bone and proper management and positioning of the gingival tissues. The results from osseous resective surgery are technique sensitive. It has limited use in treating cases with very deep intrabony or hemiseptal defects, which should be treated with a different surgical approach. If osseous resective surgery is used in advanced lesions, a compromise in the amount of probing depth reduction should be expected. Yet, osseous resective surgery provides the surest method of reducing pockets with an intrabony or hemiseptal osseous component of 3 mm or less, albeit at the expense of some attachment in the neighboring less involved sites. Osseous resective surgery has been and remains one of the principal periodontal treatment modalities because of its proven success (Fig. 17). PMID- 11276518 TI - Diagnosis and epidemiology of periodontal osseous lesions. PMID- 11276519 TI - The treatment of intrabony defects with bone grafts. AB - There is substantial clinical and histological evidence that support the concept that extraoral and intraoral autogenous bone grafts and demineralized freeze dried bone allografts are effective regenerative materials in the treatment of intrabony defects. Moreover, long-term evaluations currently available suggest that the regenerative gains achieved remain clinically stable. Synthetic grafts may result in improved probing depths and clinical attachment levels but have yet to demonstrate the ability to initiate or enhance the formation of a new attachment apparatus. PMID- 11276520 TI - NCDHM. CDS members spread the word at JFK Health world. PMID- 11276521 TI - Negative news by any other name smells just as nasty. PMID- 11276522 TI - The dynamics of change. PMID- 11276523 TI - Seven ways to achieve quality full-mouth surveys. PMID- 11276524 TI - Periodic and quasi-periodic behavior in resource-dependent age structured population models. AB - To describe the dynamics of a resource-dependent age structured population, a general non-linear Leslie type model is derived. The dependence on the resources is introduced through the death rates of the reproductive age classes. The conditions assumed in the derivation of the model are regularity and plausible limiting behaviors of the functions in the model. It is shown that the model dynamics restricted to its omega-limit sets is a diffeomorphism of a compact set, and the period-1 fixed points of the model are structurally stable. The loss of stability of the non-zero steady state occurs by a discrete Hopf bifurcation. Under general conditions, and after the loss of stability of the structurally stable steady states, the time evolution of population numbers is periodic or quasi-periodic. Numerical analysis with prototype functions has been performed, and the conditions leading to chaotic behavior in time are discussed. PMID- 11276525 TI - The migration of cells in multicell tumor spheroids. AB - A mathematical model is proposed to explain the observed internalization of microspheres and 3H-thymidine labelled cells in steady-state multicellular spheroids. The model uses the conventional ideas of nutrient diffusion and consumption by the cells. In addition, a very simple model of the progress of the cells through the cell cycle is considered. Cells are divided into two classes, those proliferating (being in G1, S, G2 or M phases) and those that are quiescent (being in G0). Furthermore, the two categories are presumed to have different chemotactic responses to the nutrient gradient. The model accounts for the spatial and temporal variations in the cell categories together with mitosis, conversion between categories and cell death. Numerical solutions demonstrate that the model predicts the behavior similar to existing models but has some novel effects. It allows for spheroids to approach a steady-state size in a non monotonic manner, it predicts self-sorting of the cell classes to produce a thin layer of rapidly proliferating cells near the outer surface and significant numbers of cells within the spheroid stalled in a proliferating state. The model predicts that overall tumor growth is not only determined by proliferation rates but also by the ability of cells to convert readily between the classes. Moreover, the steady-state structure of the spheroid indicates that if the outer layers are removed then the tumor grows quickly by recruiting cells stalled in a proliferating state. Questions are raised about the chemotactic response of cells in differing phases and to the dependency of cell cycle rates to nutrient levels. PMID- 11276526 TI - Mathematical conservation ecology: a one-predator-two-prey system as case study. AB - A method is presented to analyse the long-term stochastic dynamics of a biological population that is at risk of extinction. From the full ecosystem the method extracts the minimal information to describe the long-term dynamics of that population by a stochastic logistic system. The method is applied to a one predator-two-prey model. The choice of this example is motivated by a study on the near-extinction of a porcupine population by mountain lions whose presence is facilitated by mule deer taking advantage of a change in land use. The risk of extinction is quantified by the expected time of extinction of the population. PMID- 11276527 TI - Bi-trophic food chain dynamics with multiple component populations. AB - Food web models describe the patterns of material and energy flow in communities. In classical food web models the state of each population is described by a single variable which represents, for instance, the biomass or the number of individuals that make up the population. However, in a number of models proposed recently in the literature the individual organisms consist of two components. In addition to the structural component there is an internal pool of nutrients, lipids or reserves. Consequently the population model for each trophic level is described by two state variables instead of one. As a result the classical predator-prey interaction formalisms have to be revised. In our model time budgets with actions as searching and handling provide the formulation of the functional response for both components. In the model, assimilation of the ingested two prey components is done in parallel and the extracted energy is added to a predators reserve pool. The reserves are used for vital processes; growth, reproduction and maintenance. We will explore the top-down modelling approach where the perspective is from the community. We will demonstrate that this approach facilitates a check on the balance equations for mass and energy at this level of organization. Here it will be shown that, if the individual is allowed to shrink when the energy reserves are in short to pay the maintenance costs, the growth process has to be 100% effective. This is unrealistic and some alternative model formulations are discussed. The long-term dynamics of a microbial food chain in the chemostat are studied using bifurcation analysis. The dilution rate and the concentration of nutrients in the reservoir are the bifurcation parameters. The studied microbial bi-trophic food chain with two component populations shows chaotic behaviour. PMID- 11276528 TI - The effect of a receptor layer on the measurement of rate constants. AB - Many cellular reactions involve a reactant in solution binding to or dissociating from a reactant attached to a surface. Most studies assume that the reactions occur on this surface, when in actuality the receptors usually lie in a thin layer on top of it. The effect of this layer is considered, particularly as it relates to the BIAcore measurement device, though the results are applicable to biological systems. A dimensionless parameter measuring the strength of the effect of the receptor layer is found. Asymptotic and singular perturbation techniques are used to analyse association and dissociation kinetics, though the effect of the receptor layer need not be small. Linear and nonlinear integral equations result from the analysis; explicit and asymptotic solutions are constructed for physically realizable cases. In addition, effective rate constants are derived that illustrate the combined effects of transport and the receptor layer on the measured rate constants. All these expressions provide a direct way to estimate rate constants from BIAcore binding data. PMID- 11276529 TI - A mathematical model for prokaryotic protein synthesis. AB - A kinetic model for the synthesis of proteins in prokaryotes is presented and analysed. This model is based on a Markov model for the state of the DNA strand encoding the protein. The states that the DNA strand can occupy are: ready, repressed, or having a mRNA chain of length i in the process of being completed. The case i = 0 corresponds to the RNA polymerase attached, but no nucleotides attached to the chain. The Markov model consists of differential equations for the rates of change of the probabilities. The rate of production of the mRNA molecules is equal to the probability that the chain is assembled to the penultimate nucleotide, times the rate at which that nucleotide is attached. Similarly, the mRNA molecules can also be in different states, including: ready and having an amino acid chain of length j attached. The rate of protein synthesis is the rate at which the chain is completed. A Michaelis-Menten type of analysis is done, assuming that the rate of protein degradation determines the 'slow' time, and that all the other kinetic rates are 'fast'. In the self regulated case, this results in a single ordinary differential equation for the protein concentration. PMID- 11276530 TI - On the ontogeny of the motor circadian rhythm in crayfish. AB - In this paper we attack the problem of understanding the localization of the main structures involved in the motor circadian rhythm of crayfish by analysing its ontogeny. We present experimental results giving the properties of this rhythm in young and adult crayfish. Then we construct a mathematical model (based on a previous one for the electroretinogram circadian rhythm in the same species) simulating those properties. In the process of constructing the model we clarified and made precise various hypotheses about the biological structures involved in them and about the characteristics of the oscillators present in those structures. We also formulate some hypotheses about the general properties of circadian rhythms. Finally, we propose some experiments suggested by the mathematical model. PMID- 11276531 TI - From spikers to bursters via coupling: help from heterogeneity. AB - It has been shown previously that identical spiking cells, incapable of bursting by themselves, may burst under weak diffusive coupling conditions. With stronger coupling, the coupled system reverts to bursting can be recovered by introducing heterogeneity in the model parameters. For a two-cell system, we explain the phenomenon with bifurcation analysis. For larger clusters of coupled cells, we demonstrate by numerical simulation that the phenomenon carries over. In addition, we use a perturbation analysis to deduce the dependence of the heterogeneity parameter used in the bifurcation analysis on the original heterogeneity parameters and the coupling strength. Implications of the phenomenon of emergent bursting are discussed in the context of electrical activity in pancreatic beta cells. PMID- 11276532 TI - Body weight setpoint, metabolic adaption and human starvation. AB - A biological setpoint for fatness has been proposed in the medical literature. This body weight setpoint functions as a point of stable equilibrium. In an underfed state, with resulting weight loss, the body will reduce the relative energy expenditure by metabolic adaption which reduces the rate of weight loss. Previous mathematical models of energy expenditure and weight loss dynamics have not addressed this setpoint mechanism. The setpoint model has been proposed to quantify this biological process and is unique in predicting energy expenditure during weight loss as a function of the setpoint fat-free mass ratio and setpoint energy expenditure, eliminating the various controlling characteristics such as age, gender and heredity. The model is applied to the seminal Minnesota human semistarvation experiment and is used to predict weight vs time on an individual basis and the caloric requirements for weight maintenance at the reduced weight. Comparison is made with the Harris-Benedict equations and the Brody-Kleiber (W3/4) law. PMID- 11276533 TI - Hospitalizing the homeless. PMID- 11276534 TI - The stethoscope at ease. PMID- 11276535 TI - Preventing deaths from long QT syndrome. PMID- 11276536 TI - The stethoscope at ease. PMID- 11276537 TI - Lest we forget. PMID- 11276538 TI - Cervical manipulation: how risky is it? PMID- 11276540 TI - Cervical manipulation: how risky is it? PMID- 11276539 TI - Cervical manipulation: how risky is it? PMID- 11276541 TI - Evidence to action: a tailored multifaceted approach to changing family physician practice patterns and improving preventive care. AB - BACKGROUND: Although there is much room for improvement in the performance of recommended preventive manoeuvres, many inappropriate preventive interventions are being done. We evaluated a multifaceted intervention, delivered by nurses trained in prevention facilitation, to improve prevention in primary care. METHODS: Forty-six health service organizations (HSOs) were recruited from 100 sites in Ontario. After baseline data were collected, we randomly assigned the practices to either an 18-month (July 1997 to December 1998) multifaceted intervention delivered by 1 of 3 nurse facilitators (23 practices) or no intervention (23 practices). The unit of intervention and analysis was the medical practice. The outcome measure was an overall index of preventive performance, which was calculated as the proportion of eligible patients who received 8 recommended preventive manoeuvres less the proportion of eligible patients who received 5 inappropriate preventive manoeuvres. RESULTS: One HSO, in the intervention group, was lost to follow-up. Before the intervention, the index of preventive performance was similar for the intervention and control groups (31.9% [95% confidence interval (CI) 27.3%-36.5%] and 32.1% [95% CI 27.2%-37.0%] respectively). At follow-up the corresponding values were 43.2% (95% CI 38.4% 48.0%) and 31.9% (95% CI 26.8%-37.0%), for an absolute improvement in the intervention group of 11.5% (p < 0.001). The mean proportion of eligible patients who received the recommended manoeuvres was 62.3% (95% CI 58.2%-66.4%) in the intervention group, as compared with 57.4% (95% CI 54.1%-60.7%) in the control group, for an absolute improvement of 7.2% (p = 0.008). The corresponding values for the inappropriate manoeuvres were 19.1% (95% CI 15.6%-22.6%) and 25.5% (95% CI 20.0%-31.0%), for an absolute improvement of 4.4% (p = 0.019). INTERPRETATION: The tailored multifaceted intervention delivered by nurse facilitators was effective in modifying physician practice patterns and significantly improved preventive care performance. PMID- 11276543 TI - Influence of bone densitometry results on the treatment of osteoporosis. AB - BACKGROUND: Measurement of bone mineral density is widely used to diagnose osteoporosis. The objectives of this study are to determine how bone densitometry affects subsequent treatment of osteopenia and osteoporosis with either hormone replacement therapy or bisphosphonates and to examine clinical factors associated with starting either therapy after bone densitometry. METHODS: We conducted a prospective study involving women over 50 years of age who were referred to a tertiary care hospital for the first time to undergo bone density measurement using dual-energy x-ray absorptiometry (DXA). Baseline clinical data were collected through face-to-face interviews before the test. Subsequently, the scans were reviewed and categorized as showing no bone loss, osteopenia or osteoporosis, based on World Health Organization criteria. Three months after DXA, subjects were contacted by telephone to determine their understanding of the test results and any new treatments started or recommended since the scan. RESULTS: Of 383 women recruited at the time of their DXA, 335 (87.5%) completed the 3-month follow-up. Among those reached at follow-up, DXA results showed no bone loss in 119 (35.5%), osteopenia in 137 (40.9%) and osteoporosis in 79 (23.6%). The proportion of subjects with osteoporosis receiving either hormone replacement therapy or bisphosphonate therapy was 15.2% before the test, increasing to 63.3% after the scan. The following factors were independently associated with the initiation of either therapy: actual DXA result showing osteoporosis (odds ratio [OR] 7.2; 95% confidence interval [CI] 1.7-30.3), compared with a normal scan; subjects' perception that their scan showed bone loss (osteopenia, or osteoporosis) (OR 13.5; 95% CI 4.0-45.5) or that they were unclear about the results (OR 5.4; 95% CI 1.6-18.8), compared with the perception that the results were normal; and discussion of the DXA results with a physician (OR 5.5; 95% CI 1.9-16.0). INTERPRETATION: The proportion of women with osteoporosis receiving hormone replacement therapy or bisphosphonate therapy increases after diagnosis with densitometry. However, communication by physicians so that patients understand their test results is a critical component in the initiation of therapy after bone densitometry. PMID- 11276542 TI - Sex-specific determinants of HIV infection among injection drug users in Montreal. AB - BACKGROUND: Sex-specific issues have not been extensively addressed in studies of HIV prevalence, despite the strong implications of differences between men and women in the risk of HIV transmission. The objective of this study was to examine sex-specific behaviours associated with HIV infection among injection drug users in Montreal. METHODS: A total of 2741 active drug users (2209 [80.6%] men) were recruited between 1988 and 1998. Information was sought on sociodemographic characteristics, drug-related behaviour and sexual behaviour, and participants were tested for HIV antibodies. Sex-specific independent predictors of HIV prevalence were assessed by stepwise logistic regression. RESULTS: The overall prevalence of HIV among study subjects was 11.1%; the prevalence was 12.0% among men and 7.5% among women. In multivariate models, a history of sharing syringes with a known seropositive partner (odds ratio [OR] for men 2.44, 95% confidence interval [CI] 1.72-3.46; OR for women 3.03, 95% CI 1.29-7.13) and of sharing syringes in the past 6 months (OR for men 0.61, 95% CI 0.44-0.85; OR for women 0.32, 95% CI 0.14-0.73) were independently associated with HIV infection. Other variables associated with HIV infection were homosexual or bisexual orientation, cocaine rather than heroin as drug of choice, frequency of injection drug use, and obtaining needles at a pharmacy or through needle exchange programs (for men only) and obtaining needles at shooting galleries and being out of treatment (for women only). INTERPRETATION: These results support the hypothesis that risk factors for HIV seropositivity differ between men and women. These sex-related differences should be taken into account in the development of preventive and clinical interventions. PMID- 11276544 TI - Pill-splitting in a long-term care facility. PMID- 11276545 TI - Facilitating the integration of prevention in primary care: a work in progress. PMID- 11276546 TI - The Senate report on end-of-life care: the ball is in our court. PMID- 11276547 TI - Alberta's Bill 11: will trade tribunals set domestic health policy? PMID- 11276548 TI - Injection drug use and despair through the lens of gender. PMID- 11276549 TI - Statins and bones. PMID- 11276550 TI - Problems for clinical judgement: 2. Obtaining a reliable past medical history. AB - Ordinary human reasoning may lead patients to provide an unreliable history of past experiences because of errors in comprehension, recall, evaluation and expression. Comprehension of a question may change depending on the definition of periods of time and prior questions. Recall fails through the loss of relevant information, the fabrication of misinformation and distracting cues. Evaluations may be mistaken because of the "halo effect" and a reluctance to change personal beliefs. Expression is influenced by social culture and the environment. These errors can also occur when patients report a history of present illness, but they tend to be more prominent with experiences that are more remote. An awareness of these specific human fallibilities might help clinicians avoid some errors when eliciting a patient's past medical history. PMID- 11276551 TI - Addiction and the brain: the role of neurotransmitters in the cause and treatment of drug dependence. AB - Recent scientific advances have led to a greater understanding of the neurobiological processes that underlie drug abuse and addiction. These suggest that multiple neurotransmitter systems may play a key role in the development and expression of drug dependence. These advances in our knowledge promise not only to help us identify the underlying cause of drug abuse and dependence, but also to aid the development of effective treatment strategies. PMID- 11276552 TI - The illegality of private health care in Canada. AB - We addressed the question of whether private health care is illegal in Canada by surveying the health insurance legislation of all 10 provinces. Our survey revealed multiple layers of regulation that seem to have as their primary objective preventing the public sector from subsidizing the private sector, as opposed to rendering privately funded practice illegal. Private insurance for medically necessary hospital and physician services is illegal in only 6 of the 10 provinces. Nonetheless, a significant private sector has not developed in any of the 4 provinces that do permit private insurance coverage. The absence of a significant private sector is probably best explained by the prohibitions on the subsidy of private practice by public plans, measures that prevent physicians from topping up their public sector incomes with private fees. PMID- 11276554 TI - Abortion services in Canada: a patchwork quilt with many holes. PMID- 11276553 TI - Bioethics for clinicians: 24. Brain death. AB - Brain death is defined as the complete and irreversible absence of all brain function. It is diagnosed by means of rigorous testing at the bedside. The advent of neurologic or brain death criteria to establish the death of a person was a significant departure from the traditional way of defining death and remains ethically challenging to some. We review the ethical, cultural, religious and legal issues surrounding brain death and outline an approach to establishing a diagnosis of brain death in clinical practice. PMID- 11276555 TI - Feds' approach to genetically modified products criticized. PMID- 11276556 TI - One in 4 Canadians smokes, Ottawa reports. PMID- 11276557 TI - Supplementary fees a necessity, physicians argue. PMID- 11276558 TI - A shortage of doctors? What shortage? PMID- 11276559 TI - Build a simple Web site for your practice in less than an hour. PMID- 11276560 TI - Combination treatment effective option for hypertensive, diabetic patients with microalbuminuria. PMID- 11276561 TI - Navel gazing: a clinical glimpse at body piercing. PMID- 11276562 TI - Alcohol policy in Europe. What can the European Union do? PMID- 11276563 TI - A follow-up study of psychosocial factors and musculoskeletal problems among unskilled female workers with monotonous work. AB - BACKGROUND: Musculoskeletal problems are among the most prevalent occupational health problems in industrialised countries and seem to be common among unskilled, female industrial workers. However, cross-sectional studies only reflect the current situation with regard to both exposure and effect, and selection bias may mask work-related musculoskeletal problems so the general assumption is that prospective cohort studies are more valid and informative. The aim with the present study was to follow-up a group of unskilled female workers and determine whether the number of musculoskeletal ailments reported had changed after 3 years on a group and/or on an individual level. The women had earlier participated in a cross-sectional study correlating exposure with different physical and psychosocial factors at work, at home and during leisure time with their reports of musculoskeletal ailments in the neck, shoulders and thoracic spine. METHODS: One hundred and fifty-three women from the original study group of 173 received a mailed questionnaire, including a visual analogue scale (VAS) and a pain drawing. RESULTS: Ninety-three women were included in the final analyses. Some deterioration in general health and, in particular, in psychological health was observed compared with the earlier study but there was less change in the reporting of musculoskeletal ailments. CONCLUSION: Economic decline and its consequences may have had both a direct and an indirect impact on the deterioration in general health but not in musculoskeletal problems of the women still employed. PMID- 11276564 TI - Psychological distress and quality of life in drug-using and non-drug-using HIV infected women. AB - BACKGROUND: Quantitative research on women infected with human immunodeficiency virus (HIV) has predominantly involved drug users. The present study assessed the psychosocial burden of HIV infection and identified some possible determinants in both drug-using and non-drug-using women. METHOD: Twenty-three hard-drug-using and 55 non-using HIV-positive women aged 18-64 years participated. Psychological distress (SCL-90, with eight scales and total score) and health-related quality of life (Rand SF-36, with eight scales and physical and psychosocial dimension) were measured. A cross-sectional comparison with reference groups (female general population, female psychiatric patients and HIV-positive homosexual men) was made. For analysis the t-test and multiple regression analysis were used. RESULTS: Compared to the general population, both HIV-positive groups had higher (i.e. unfavourable) SCL total scores (t = 8.33 and p < 0.001 and t = 4.97 and p < 0.001) and lower (i.e. unfavourable) Rand SF-36 scores (p < 0.001 on seven or more scales). Compared to psychiatric patients, drug users had similar (n.s.) and non-drug users had lower (t = -9.09 and p < 0.001) SCL scores. Both groups had lower SF-36 scores (p < 0.001 on seven or more scales). Compared to HIV-positive homosexuals, drug users had higher (t = 2.88 and p < 0.01) and non-drug users had similar SCL scores (n.s.). Psychosocial illness burden (SCL and Rand psychosocial dimension) was associated with low self-esteem, poverty, ethnic minority membership and illness stage (Rand only). Child care, drug use/prostitution and illness stage predicted high physical illness burden. CONCLUSION: Women with HIV/AIDS (acquired immune deficiency syndrome) experience considerable distress and poor quality of life, but drug users do more so than non-users. Drug- and gender-related lifestyles affect illness burden. PMID- 11276565 TI - Development of a child health indicator system in Italy. AB - BACKGROUND: The need for a uniform, comprehensive and action-oriented child health indicator system is widely recognised. As part of a Ministry of Health project, a working group was established in Italy in order to develop a proposal for a minimum set of health indicators to be adopted at the regional and local health authority levels, where the planning process takes place. METHODS: The indicators proposed cover 17 areas of perinatal, child and adolescent health. The informing principles for the choice were relevance to the main health problems, availability of a reliable data collection system, feasibility of the collection and analysis process at the two health system levels proposed and extent to which the information provides clues for policy options. RESULTS: The main difficulties arise from a lack of uniform systems of classification and data collection for disabilities, as well as adequate tools for assessing quality of care and quality of life. A basic framework for analysis is suggested, including further breakdown of the indicators proposed, such as analysis by birthweight and by cause of neonatal death and by mother's education and father's employment. The information provided by the health indicators put forward needs to be evaluated within the broader scenario of the child's situation so that associated factors may be identified and clues found for intersectoral policies. Two research projects were started to evaluate the feasibility and reliability of data collection and the impact on the planning process at both the regional and local health authority levels. CONCLUSION: A European-wide initiative is proposed to tackle existing methodological problems effectively and develop a common child health indicator system. PMID- 11276566 TI - Traumatic experience and sleep disturbance in refugee children from the Middle East. AB - BACKGROUND: Sleep disturbance is frequently reported in children after traumatic experiences associated with organised violence. The aim of this study was to identify specific traumatic risk indicators and modifying factors for sleep disturbance among recently arrived refugee children from the Middle East. METHODS: The study group comprises 311, 3-15 year old refugee children from the Middle East. On arrival in Denmark, their parents participated in a structured interview about their childrens' health and history of exile and eventual exposure to war, organised violence and human rights violation. RESULTS: A family history of violence (grandparent's violent death before the birth of the child or parental exposure to torture) as well as a stressful present family situation (father scolds the child more than previously) were the strongest predictors of prevalent sleep disturbance in the children. Arriving in Denmark with both parents rather than one was a modifying factor, so the effect of traumatic experience on sleep patterns later in childhood was mediated through parental presence and behaviour. CONCLUSION: This study indicates that the family environment is of primary importance for childhood sleep disturbance following traumatic experiences connected with war and other organised violence. PMID- 11276567 TI - Employment status of pregnant women in central Poland and the risk of preterm delivery and small-for-gestational-age infants. AB - BACKGROUND: Unemployment is one of the consequences of the ongoing transformation of the Polish economy. The main objective of the present study was to investigate the relationship between women's employment status and preterm delivery (PD) and/or small-for-gestational-age (SGA) infants, in Central Poland during the socio-economic transition. METHODS: The study population comprised 8% random sample of 2,080 women from the Lodz macro region who gave birth to a child during a one-year period (1996-1997). Based on the employment status, three groups were distinguished: the employed (n = 1238), the unemployed (n = 128)--women who lost their job before pregnancy (at least 12 months prior to delivery) and were actively seeking employment, and the housewives (n = 714). Women with chronic medical problems diagnosed before pregnancy were excluded from the study. RESULTS: The rates of preterm delivery were found to be 6.3%, 11.7% and 4.9%, respectively for the employed, unemployed, and housewives. After adjustment for age, marital status, education, maternal height, smoking, own apartment, presence of cervical insufficiency and uterine irritability, an excess risk of PD was observed among the unemployed women (OR = 1.92, 95% CI: 1.01-3.64). Unemployment was associated, though not significantly, with higher rates of SGA births. CONCLUSION: In Central Poland, the unemployed pregnant women are characterised by an excess risk of preterm delivery. The impact of unemployment on the community's health, which seems to be underestimated, should be given more consideration in the national and local policies for public health. PMID- 11276568 TI - Social differentials in the decline of child mortality in nineteenth century Stockholm. AB - BACKGROUND: Mortality had declined dramatically by the end of the nineteenth century and the early twentieth century. Little is known about the development of social differentials in infant and child mortality in Stockholm at the turn of the century. This study investigates social differentials in child mortality during the years 1885, 1891 and 1910 in one parish in Stockholm. METHODS: Individual entries from computerised records originally collected for civil registration purposes in Stockholm for 1878-1925 (the Roteman Archives) were analysed with respect to social class of the head of household and marital status of the mother for 36,718 children aged 0-14 years. Age- and cause-specific mortality rates were calculated for each year of study. Cox' regression analysis was used to analyse the mortality risk (relative rates (RRs) of mortality) by socioeconomic group and by marital status of the mother. RESULTS: Child mortality rates were nearly halved between 1885 and 1910. Socioeconomic differentials in mortality between the four social classes emerged from 1891 as the overall mortality declined. The decline was sharpest in the upper and middle social classes. Children born out of wedlock had higher mortality rates than children of married mothers in all 3 years studied. CONCLUSION: The social differentials in child mortality risk were substantial and the gradient emerged sharper from 1891 to 1910. The results are in line with studies from England and Wales, Germany and the USA for the same time period. The differentials mostly increased because of a greater decline in mortality among higher socioeconomic groups. PMID- 11276569 TI - Long-term illness and psychosomatic complaints in children aged 2-17 years in the five Nordic countries. Comparison between 1984 and 1996. AB - BACKGROUND: The aim of the study was to investigate changes in psychosomatic complaints (PSC) and long-term illness (LTI) and the association between parents' socioeconomic conditions and children's complaints in the five Nordic countries (Denmark, Finland, Iceland, Norway and Sweden). METHODS: The cross-sectional study covered a representative sample of children, aged 2-17 years in each country, altogether 10,219 in 1984 and 10,317 in 1996. The data were collected by mailed questionnaires. The relationship between LTI, PSC and parents' education, income, satisfaction with economy and family situation was analysed using descriptive statistics and logistics regression. RESULTS: LTI and PSC in children have increased in all age groups between 1984 and 1996. The prevalence of LTI was higher among boys than girls, whereas the prevalence of PSC was higher among girls. In both years, the prevalence was higher in low-income, low-educated, one parent families. The general tendency is the same in all countries, although there are some minor differences. The differences in 1984 remained unchanged in 1996 except for LTI that increased in low-educated families for Nordic children as a whole. CONCLUSION: Socially and economically vulnerable families seem to run the greatest risk of having children with LTI and PSC, although the data do not permit a strict causal relationship to be drawn. Also, families with high formal education, solid economy and general satisfaction with their situation seem to be able to offer their children sheltered and stimulating conditions for growth and development, even in times of economic recession. PMID- 11276570 TI - Subjective health complaints in adolescence. A cross-national comparison of prevalence and dimensionality. AB - BACKGROUND: The purpose of this work was to study the prevalence and dimensionality of subjective health complaints in a cross-national population of adolescents. METHODS: The analyses were based on data from a WHO cross-national survey, Health Behaviour in School-aged Children (HBSC). The study included a representative sample of 11, 13 and 15-year-old adolescents from Finland, Norway, Poland and Scotland. Data were collected in 1993-1994 and the total sample included 20,324 adolescents. Subjective health complaints were measured by the HBSC Symptom Checklist (HBSC-SCL), including headaches, abdominal pain, backache, feeling low, irritability, nervousness, sleeping difficulties and dizziness. Descriptive analyses, MANOVA and structural equation modelling (EQS) were conducted. RESULTS: Patterns of reporting were consistent for all four countries. A large number of students reported a high level of symptoms. The reporting of most symptoms increased with age. Girls reported significantly more symptoms than boys and the gender differences also increased with age. Structural equation modelling suggests a model of two correlated factors, which can be labelled psychological and somatic. CONCLUSION: The findings of this study indicate that students report a high level of subjective health complaints already at the age of 11 years. The reporting of most symptoms increases with age and more so for girls than for boys. The finding of two dimensions that differ qualitatively, suggests that these dimensions may have different etiologies. PMID- 11276571 TI - Socio-economic position and coronary heart disease risk factors in youth. Findings from the Young Hearts Project in Northern Ireland. AB - BACKGROUND: This study investigates the existence of socioeconomic differentials in behavioural and biological risk factors for coronary heart disease in young people from Northern Ireland, taking into account differences in biological maturation. METHODS: A school-based prospective study, with measurements in 1989/1990 and 1992/1993. Socio-economic position was based on occupational level of the main family breadwinner. Behavioural risk factors included were physical inactivity, the intake of total energy, dietary fat and a number of micronutrients. Biological risk factors included were blood pressure, body fatness, lipoproteins and cardio-pulmonary fitness. Biological maturation was based on Tanner's stages. PARTICIPANTS: 251 boys and 258 girls who were measured at the age of 12 years and re-examined at the age of 15 years. RESULTS: Cross sectional analyses showed that socio-economic differences in cholesterol intake (in boys) and physical inactivity and total energy intake (in girls) were present at 12 and 15 years of age, while differences in fat and fruit intake and smoking behaviour (in boys and girls) became established at the age of 15 years, with unfavourable levels in subjects in the manual group. Longitudinal analyses confirmed that differences in behavioural risk factors exist or develop during adolescence. No clear pattern of differences in biological risk factors was found by socio-economic position. Adjustment for biological maturation did not materially alter the results. CONCLUSION: Differences in lifestyle by socio economic position seem to become established in adolescence. These differences however, are not (yet) reflected in differences in biological risk factors by socio-economic position. PMID- 11276572 TI - Socioeconomic differences in the consumption of vegetables, fruit and fruit juices. The influence of psychosocial factors. AB - BACKGROUND: The aim was to investigate whether social network and social support factors can explain socioeconomic differences in the risk of consuming low amounts of vegetables, fruit and fruit juices. METHODS: The Malmo Diet and Cancer Study was a prospective cohort study. The present cross-sectional study examined data from a subpopulation of 11,837 individuals that completed baseline examinations in 1992-1994. Dietary habits were assessed using a modified diet history method, and socioeconomic and social network factors were measured with a structured questionnaire. Low consumption was defined as the lowest consumption quartile for vegetables and fruit, while fruit juice consumption was dichotomized to separate users from non-users. RESULTS: Socioeconomic differences were most pronounced regarding the consumption of vegetables and fruit juices. For both sexes, unskilled manual workers had a twice as high risk of low vegetable and fruit juice consumption as higher non-manual employees. No socioeconomic differences in fruit consumption were observed for men, and only moderate differences for women with a higher consumption in higher socioeconomic groups. When the psychosocial variables were introduced in the multivariate model, social participation moderately reduced the socioeconomic differences in vegetable consumption, and the female socioeconomic differences in fruit consumption, but had no effect on the socioeconomic differences in fruit juice consumption. The other psychosocial variables had no effect on the socioeconomic differences. CONCLUSION: Considerable socioeconomic differences in vegetable, fruit and fruit juice consumption were observed. Social participation seemed to be a strong determinant for these food choices. However, this effect was largely independent of the socioeconomic differences. PMID- 11276573 TI - Diabetes and social conditions in Estonia. A population-based study. AB - BACKGROUND: The aim of this study was to investigate aspects of metabolic control, treatment and complications as well as quality of life in patients with diabetes mellitus from a defined population in Estonia. METHODS: We invited 220 randomly selected diabetes patients recruited to a clinical investigation from a local diabetes register of 1,100 patients in Viljandi, Estonia. The main outcome measures were derived from medical history, physical examination (height, weight and blood pressure), laboratory variables (blood glucose and glycated haemoglobin A1 (HbA1 normal reference range 3.2-5.6%), serum total and HDL cholesterol and creatinine), a questionnaire on disease-related knowledge and quality of life variables. RESULTS: In all, 181 diabetes patients were investigated, of whom 90% were diagnosed with type 2 diabetes. The mean diabetes duration was 8.9 years from clinical diagnosis and mean HbA1 level was 7.3%. The overall proportion of patients treated with insulin was 29.8% and with anti-hypertensive drugs 26.5%. Smoking was present in 14.3%. The proportion of patients with various diabetes complications was high (73.5%), mostly consisting of different manifestations of cardiovascular disease. Foot ulcers or gangrene were observed in 11.6%. A low level of quality of life was registered in many patients, mostly due to difficult living conditions. CONCLUSIONS: Diabetes patients in Viljandi showed an acceptable degree of glucose metabolic control, but reported a high degree of diabetes complications, as well as impaired quality of life. The diabetes complications may therefore be due to detrimental factors other than hyperglycaemia, e.g. the standard of care during previous years as well as current social and living conditions. PMID- 11276574 TI - Do health behaviour and psychosocial risk factors explain the European east-west gap in health status? AB - BACKGROUND: Mortality rates are much more favourable in Western European countries than in those of Eastern Europe. Health behaviour and psychosocial factors have been suggested to be important contributors to East-West differences in mortality and health status. METHODS: To compare reported health status as well as health behaviours and psychosocial factors which may be related to unequal health status in different parts of Europe, standardised postal surveys of representative populations samples were conducted in six Eastern and Western European areas. RESULTS: Higher mortality in the eastern populations was associated with more reported morbidity and generally more negative health ratings. Health behaviours and psychosocial factors were also more negative in the East. Multivariate analyses suggested that the East-West difference in health status may be partly explained by differences in health behaviours and psychosocial factors. CONCLUSION: Efforts to promote health in Eastern Europe should concentrate both on the promotion of healthier lifestyles and on improvement of social and economic conditions. PMID- 11276575 TI - Self-reported health in the Republic of Karelia, Russia and in north Karelia, Finland in 1992. AB - BACKGROUND: Major differences in mortality, cardiovascular disease risk factors and health behaviour are known to exist between the populations of eastern Finland and the Republic of Karelia, Russia. Little is known, however, whether similar differences exist in subjective health. METHODS: In spring 1992 a population survey was conducted in North Karelia, Finland and in the area of Pitkaranta, Republic of Karelia, Russia. Random population samples (n = 2,000 in North Karelia and n = 1,000 in Pitkaranta) stratified for age and sex were taken from the population registers. The subjects completed questionnaires and were examined at local health centres. RESULTS: In North Karelia 50% of men reported their health as being quite good or very good, compared to 34% in Pitkaranta (p < 0.0001 for area difference). Among women the corresponding percentages were 58% in North Karelia and 22% in Pitkaranta (p < 0.0001). High household income and education were associated with good self-rated health among both sexes in North Karelia and among women but not men in Pitkaranta. Self-reported physical condition was better in North Karelia than in Pitkaranta (p < 0.0001). Psychosomatic symptoms (p = 0.0002 among men and p < 0.0001 among women) and many somatic symptoms were more prevalent in Pitkaranta than in North Karelia. CONCLUSION: In general, people in North Karelia, Finland feel healthier than people in the neighbouring Republic of Karelia, Russia. Socioeconomic differences in subjective health are less prominent in the Republic of Karelia. PMID- 11276576 TI - Physical and psychological effects of injury. Data from the 1958 British birth cohort study. AB - BACKGROUND: There is only scant evidence for the long-term health effects of road traffic injuries. We therefore assessed the extent to which motor vehicle driver injuries influence limiting long-standing illness and psychological distress using data from a nationwide study (the 1958 British birth cohort) in early adulthood. METHODS: Information was obtained on driver injuries occurring between ages 23 and 33 years and limiting illnesses and psychological distress at age 33 years. The risks of injury-related adverse consequences were derived using logistic regression and expressed as odds ratios (ORs) and 95% confidence intervals. RESULTS: A single injury was associated with limiting illness (OR = 2.01 and 95% CI: 1.38-2.94). The association between a single injury and psychological distress was strong for a recent injury occurring between ages 30 and 33 years (OR = 1.86 and 95% CI: 1.24-2.81), but not for injuries occurring earlier on. The population attributable fraction for limiting illness with one injury was 3.8% (range 1.7-5.3%) and with two or more injuries was 1.0% (range 0.5-1.3%). After controlling for potential confounding factors the corresponding figures were 4.2% (range 2.2-5.6%) and 1.1% (range 0.5-1.3%) respectively. CONCLUSIONS: Driver injuries are associated with a substantial increase in disability and, also in the short term, with increases in psychological distress. These results highlight the need for identifying effective strategies for the prevention of road traffic injuries as well as more effective approaches for rehabilitation of the injured. PMID- 11276577 TI - Registration of external causes of death in the Baltic States 1970-1997. AB - BACKGROUND: Trends in external causes of deaths in the Baltic States--Estonia, Latvia and Lithuania--were analysed against the background of turbulent political, social and economic changes. The reliability of mortality statistics concerning external causes of death in these countries is considered to be good. METHOD: This study is based on data published by the statistical offices of the three Baltic States and on data obtained through interviews with personnel employed at the national statistical offices. The study period was divided, by socio-political and economic factors, into a period of stagnation (1970-1984) and a period of reforms (1985-1997). RESULTS: During 1970-1984 a stable slightly upward trend of external causes of death rates was observed. The curve became S shaped in the reform period: between 1984 and 1988 a marked decrease occurred followed by a rapid increase of rates until 1994, and then by 1997 a fall to the approximate level of 1984. The male to female ratio of external causes of death was between 3.4:1 and 4.2:1. External deaths accounted for 10% to 14% of all deaths before 1984. During the period 1984-1988 the proportion of external deaths was under 10% and peaked in 1994 at 16%. Fluctuations in the trends of external death were more pronounced among males than females in all Baltic countries. CONCLUSION: Trends in external causes of death were similar in Baltic States. High proportions of violent death decreased life-expectancy for both sexes, but markedly for males. Social stresses and alcohol consumption could be considered as factors influencing the mortality rates and specific fluctuations in trends of external death, especially among males. PMID- 11276578 TI - Costs of asthma treatment in Estonia. AB - BACKGROUND: The aim of this study was to describe the distribution of the direct costs of asthma in Estonia by the type of treatment received and to differentiate the costs by age of patients. METHODS: Data were obtained from the databases of national health insurance offices. Persons who had been in a hospital or visited a doctor because of asthma and who also purchased anti-asthmatic drugs during 1997 were identified. The bills of all direct costs of health insurance for each asthma patient could thus be summarised. The data on all purchases of drugs were used to follow the patterns of drug treatment of asthma. RESULTS: The mean annual treatment costs of one asthma patient were 118 EUR. The costs of hospital care accounted for 27%, costs of ambulatory care for 20% and costs of pharmacotherapy for 53% of all treatment costs. Beta-agonists and corticosteroids for inhalation accounted for more than 80% of prescriptions and 90% of pharmacotherapy costs in patients under 45 years. The users of oral corticosteroids had twice as high per capita annual asthma treatment costs as compared to non-users. CONCLUSIONS: The frequency of out-patient visits, hospital admissions, number of prescriptions, total costs and costs of pharmaceuticals increased in parallel with age. The total estimated direct costs of asthma diagnosis and treatment during 1997, as extrapolated from the study population, were 2.1 million EUR, and accounted for 1.4% of direct health care costs in Estonia. PMID- 11276579 TI - A methodological note on combining health and social care expenditures into a single statistic for policy-making purposes. AB - Current national expenditure series in the health sector focus predominantly on spending for medical services. However, as the percentage of elderly individuals grows, national policy makers will increasingly require an expenditure series which includes combined expenditure for social care as well as medical expenditures. In one country, Sweden, national policy makers have begun to relate policy decisions to a 12.0% (1996) figure for combined health and social care expenditures. Calculating such a combined figure presents a number of methodological issues, such as which social care services to include and how to reflect donated care from relatives and friends. An international comparison of this new health and social care figure would enable national decision makers to judge better the efficiency and effectiveness of current policy. PMID- 11276580 TI - Critical reading of epidemiological papers. A guide. AB - Many clinicians, medical practitioners and decision makers have no formal training in epidemiology but need to understand and sometimes evaluate results in epidemiologic studies. This paper attempts to give guidance to non epidemiologists on how to read and evaluate the quality of epidemiologic studies and their results critically. Different methodological issues for evaluating whether the results of a study are causal or by bias, chance or confounding are given. This includes criteria for the choice of an appropriate study design followed by problems of the definition of the study population and the sample selection. We will also point to potential sources of bias in the data collection procedure and list some principles for statistical analysis. Finally, we include comments on how the results should be presented and issues which are related to public health and ethical questions. Although it is not usually possible to perform a perfect study and the correct approach to study design and analysis is often highly dependent on specific features of the population and the surroundings, our paper should help to distinguish between weak and strong investigations and papers. PMID- 11276581 TI - Relationship among growth, steroid production and immunolocalization of transforming growth factor-beta 1 in the normally developing mouse follicles cultured in vitro. AB - This study examined the relationship among growth, steroid production and transforming growth factor-beta 1 (TGF-beta 1) immunolocalization in the mouse follicles cultured in vitro to evaluate the hypothesis that normally developing follicles should express TGF-beta 1 in the granulosa cells around the time of antrum formation. Preantral follicles with 151-175 microns (large category) and 125-150 microns (small category) of initial diameters were used as models for normal and retarded follicles, respectively. Growth rate and timing of antrum formation in both categories were comparable to those of in-vivo grown follicles. At the time of antrum formation, follicular diameters were similar between the two follicle categories; however, antral follicles from the large category showed larger number of granulosa cells, higher estradiol production and proportion of follicles with TGF-beta 1 positive granulosa cells. Two days after antrum formation, there were no differences in the number of granulosa cells and the proportions of follicles with TGF-beta 1 positive granulosa or theca cells between the two categories. Temporal association in large follicles between the increase in estradiol production and proportion of follicles with TGF-beta 1 positive granulosa cells at the time of antrum formation supports our hypothesis. Furthermore, this study demonstrated the usefulness of the follicle culture system in the investigations of follicular physiology. PMID- 11276582 TI - Antigenic variation among equine H 3 N 8 influenza virus hemagglutinins. AB - To provide information on the antigenic variation of the hemagglutinins (HA) among equine H 3 influenza viruses, 26 strains isolated from horses in different areas in the world during the 1963-1996 period were analyzed using a panel of monoclonal antibodies recognizing at least 7 distinct epitopes on the H 3 HA molecule of the prototype strain A/equine/Miami/1/63 (H 3 N 8). The reactivity patterns of the virus strains with the panel indicate that antigenic drift of the HA has occurred with the year of isolation, but less extensively than that of human H 3 N 2 influenza virus isolates, and different antigenic variants co circulate. To assess immunogenicity of the viruses, antisera from mice vaccinated with each of the 7 representative inactivated viruses were examined by neutralization and hemagglutination-inhibition tests. These results emphasize the importance of monitoring the antigenic drift in equine influenza virus strains and to introduce current isolates into vaccine. On the basis of the present results, equine influenza vaccine strain A/equine/Tokyo/2/71 (H 3 N 8) was replaced with A/equine/La Plata/1/93 (H 3 N 8) in 1996 in Japan. The present results of the antigenic analysis of the 26 strains supported the results of a phylogenetic analysis, that viruses belonging to each of the Eurasian and American equine influenza lineages have independently evolved. However, the current vaccine in Japan consists of two American H 3 N 8 strains; A/equine/Kentucky/1/81 and A/equine/La Plata/1/93. It is also therefore recommended that a representative Eurasian strain should be included as a replacement of A/equine/Kentucky/1/81. PMID- 11276583 TI - A rapid and highly sensitive method for diagnosis of equine influenza by antigen detection using immuno-PCR. AB - A rapid and highly sensitive method for diagnosis of influenza by detecting viral antigens using immuno-PCR has been developed. The sensitivity of the immuno-PCR for the detection of the nonstructural protein 1 (NS 1) antigenically common among influenza A viruses was 10(1.0-2.0) times higher than that of RT-PCR for the detection of the viral genome. For the detection of the hemagglutinin (HA) subtype-specific antigen, this assay using anti-HA monoclonal antibodies attained a sensitivity of up to 10(7.0-8.0) times higher than those by virus isolation or by RT-PCR. PMID- 11276584 TI - Mucosal vaccination against influenza: protection of pigs immunized with inactivated virus and ether-split vaccine. AB - Effective vaccinations against swine influenza reduce the economic loss of pig industries, and also may minimize the possibility of emergence of new pandemic viruses, since pigs are intermediate hosts to generate reassortant viruses among avian and mammalian influenza viruses. In this study, we showed that intranasal immunization of pigs with formalin-inactivated or ether-split influenza vaccine (A/Aichi/2/68) induced virus-specific IgG, IgM, and IgA antibodies in their nasal secretions and sera, resulting in complete protection from virus challenge. Antibody response to the challenge virus was not observed in the immunized pigs, suggesting that the replication of the virus in the primary targets, respiratory epithelial cells, was inhibited. The present results indicate that intranasal immunization of pigs with inactivated vaccines is effective to control swine influenza, and also provide a good model, as well as a mouse model, to evaluate an intranasal application of influenza vaccine for humans. PMID- 11276585 TI - Testifying in court. PMID- 11276586 TI - Today's technology at your fingertips. PMID- 11276587 TI - Municipal bond basics. PMID- 11276588 TI - Working with people you don't like. PMID- 11276589 TI - Local dentist finds rewards overseas. PMID- 11276590 TI - Dental care and health insurance. 1938. PMID- 11276591 TI - Dental diversity: what's it all about? PMID- 11276592 TI - Design it right the first time. PMID- 11276593 TI - CDS responds to hygienist shortage. PMID- 11276594 TI - Facts about sensitive teeth. PMID- 11276595 TI - Dental records. Part 1: What and for how long? PMID- 11276596 TI - When to fire an employee. PMID- 11276597 TI - Prolonging the life of your dental handpiece. Interview by Karen A. Gomolka. PMID- 11276598 TI - The quality of sausage. PMID- 11276600 TI - Reducing central venous catheter infections. AB - This article reviews the current literature in relation to the management of central venous catheters and the prevention of catheter related infection. Sources and factors influencing catheter related infection are reviewed. In some areas of catheter management, there are clear recommendations such as the choice of skin preparation and catheter site. Other areas don't have clear guidelines; this results in varying practices and the need for further research. The latest research has been in the areas of impregnated catheters with studies showing some benefits. Recent research has also analysed the effect of the method of fluid and line changes as well as their frequency in relation to catheter related infections. A summary is given outlining interventions which have evidence supporting their practice in the reduction of catheter related infections along with interventions which may be effective in reducing catheter related infection and the need for further research. PMID- 11276601 TI - Credentialling Australian critical care nurses: the pilot study. AB - In October 1998, the Australian College of Critical Care Nurses (ACCCN) launched a model to credential specialist level critical care nurses. This model was 'road tested' during a pilot study, when experienced critical care nurses were invited to apply to become the first Australian credentialled critical care nurses. The pilot study was designed to ensure all individuals taking part in the process were surveyed regarding adequacy of the credentialling package and the credentialling process. Applicants were required to provide evidence of practise at the specialist level, as described in the Competency Standards for Specialist Level Critical Care Nurses. Evidence was presented via curriculum vitae, professional journal and supported by three peer reviewers and two referees. Each application was sent to a three-person assessor panel, which assessed the evidence in the application against the Competency Standards for Specialist Level Critical Care Nurses. A total of six applications from five states and one territory were received by April 1999. Five of the applicants were assessed to have met the Competency Standards and were awarded the credential, Australia Credentialled Critical Care Nurse (ACCN), which they will hold for a period of 3 years. Feedback from assessors, applicants, peer reviewers and referees involved in the pilot study has resulted in the further refinement of the credentialling package and processes. Australian critical care nurses will now have the opportunity to seek to be credentialled four times per year. PMID- 11276602 TI - Instillation of normal saline during endotracheal suctioning: effects on mixed venous saturation. PMID- 11276603 TI - The patients' perception of recovery after coronary angioplasty. AB - Coronary angioplasty and stent placement is associated with short hospital stays. Patients are expected to recover at home, alone, following limited care time with nurses. The purpose of the study was to describe participants' perceptions of recovery after angioplasty. Eight men and three women were interviewed 1 month after discharge from hospital. Verbatim transcripts were analysed for major themes using the qualitative techniques of grounded theory. Data analysis revealed three major categories: awareness of the problem, coping response and appraisal of the situation. These were linked via a problem solving process. In step one, the problem was identified. In step two, coping responses were taken to try and solve the problem. In step three, the results of the coping responses were appraised or evaluated. These categories were further defined by four phases identified as: pre-admission, admission, during the angioplasty and recovery. This paper describes the recovery phase. Awareness of the problem in the recovery phase was associated with 'relief from chest pain' for most participants. In contrast, anxiety continued and was associated with 'uncertainty over future health'. Participants described coping responses of "taking control of their life again" by undertaking both physical and psychological strategies. Finally, the situation was appraised to be either a 'good' or a 'bad' recovery. This appraisal was based on such considerations as the absence of chest pain, improvement in well-being and energy levels. The results of this study highlight patients' concerns and support the need for greater emphasis on their psychosocial needs. This care must be provided within the time constraints of short hospital stays. Nurses must also consider providing support to patients in the pre-admission and recovery phases. PMID- 11276604 TI - Maximising handwashing rates in the critical care unit through yearly performance feedback. AB - Handwashing following patient contact reduces the incidence of nosocomial infections. Despite this, handwashing rates by health care workers (HCWs) are often poor. Feedback on handwashing has been shown to significantly improve its rates. This study determined the optimum time to repeat performance feedback on handwashing rates of hospital staff in order to maximise its incidence. The baseline incidence of handwashing by staff following patient contact was determined by covert observation. This was followed by a period of feedback on handwashing performance by means of histograms displayed in the unit. Handwashing incidence was reassessed 6 and 12 months after the feedback ended. Performance feedback induced significant increases in handwashing incidence amongst nurses (p = 0.0433), resident medical officers (p = 0.0134), specialists (p = 0.0021) and radiographers (p = 0.0001). Non-significant increases were noted in handwashing rates amongst wardsmen/women and physiotherapists. Overall, handwashing incidence declined significantly (p = 0.0001) 12 months post feedback. This study demonstrated that feedback should be repeated within 12 months in order to maximise handwashing rates with the minimum intervention. PMID- 11276605 TI - Chest x-ray quiz. An atrial septal defect. PMID- 11276606 TI - Advanced life support: retention of registered nurses' knowledge 18 months after initial training. AB - In 1996, The Wesley Hospital introduced a 2 day Advanced Life Support (ALS) course, targeted at all critical care registered nurses and medical officers. The purpose of this study was to explore the retention of theoretical knowledge and clinical skills of registered nurses who had successfully completed the 2 day ALS course 18 months previously and to establish effective retesting timeframes. The study utilised a repeated post-test measure design. Forty registered nurses participated in the study. Data were collected during ALS retesting using scores from a theoretical examination and from the results of four practical skill assessments (basic life support, airway management, defibrillation and code management). Using Wilcoxon test, data were analysed with and compared to the participant's original scores from the training program 18 months previously. The findings demonstrate that the participant's theoretical knowledge remained at an equivalent level over the 18 month timeframe. However, 18 months after successfully completing an ALS course, only 75 per cent (n = 30) of participants passed the practical skill assessment components, with the 25 per cent (n = 10) requiring a second attempt to pass. The implications from this study focus on the model of assessment utilised and the dichotomy between theoretical and practical skill assessment results. Additional study is required to determine the optimal timeframe for ALS retesting and educational strategies to help retain skills over time. PMID- 11276607 TI - What is your life like now? Loneliness and elderly individuals residing in nursing homes. AB - Loneliness has been associated with cognitive deterioration, social isolation, hopelessness, and the inability to perform independent activities of daily living. Three factors have been found to increase levels of loneliness among elderly individuals residing in a nursing home: lack of intimate relationships, increased dependency, and loss. Nurses in long-term care facilities are in a position to directly intervene with elderly residents. Individualized interventions will allow the resident to have greater well being and greater quality of life. PMID- 11276608 TI - Alzheimer's dementia. Coping with communication decline. AB - Communication difficulties in patients with Alzheimer's dementia are a major source of caregiver strain because of the psychological and interpersonal burden they present. Nurses are in a critical position for understanding the communication decline and providing education and counseling to decrease this burden. In this article, the communication burden of dementia is examined and practical approaches to facilitating communication are discussed so nurses can increase their role in education and counseling regarding Alzheimer's dementia. PMID- 11276609 TI - Strength training for older adults: prescription guidelines for nurses in advanced practice. PMID- 11276610 TI - Preparing the health work force to serve an aging nation. PMID- 11276611 TI - Theft in nursing homes. PMID- 11276612 TI - Dysphagia among nursing home residents: an assessment and management protocol. AB - The literature suggests that 40% to 60% of nursing home residents have some degree of dysphagia, i.e., difficulty in swallowing. Poorly managed, this can lead to aspiration pneumonia; choking; chronic malnutrition; decreased quality of life; and frustration for residents, family, and staff. This article reviews the nursing literature on dysphagia management and then describes a comprehensive protocol to assess and manage patients with swallowing problems in long term care. PMID- 11276613 TI - The National Center on Women & Aging. PMID- 11276614 TI - High touch. PMID- 11276615 TI - Nutrition and older adults. PMID- 11276616 TI - Influencing health policy through research. PMID- 11276617 TI - Reproductive health consequences of intimate partner violence. A nursing research review. AB - Intimate partner violence is widespread and results in significant negative mental and physical health outcomes for women. This article is a review of nursing research on intimate partner violence and women's reproductive health and focuses on studies published since 1995, building on prior reviews. We begin with research on forced sex and the resulting physical and emotional trauma as well as implications for contraception, STD/HIV prevention, and condom use negotiation. We then discuss several approaches to the study of abuse during pregnancy, including several studies of nursing interventions. We conclude with the clinical implications of these studies. PMID- 11276618 TI - Depression, psychosocial resources, and functional ability in older adults. AB - The purpose of this study was twofold: to examine the differences between two groups of older adults (depressed, nondepressed) for physical health impairment, psychosocial resources, and functional abilities, and to describe the impact of depression, physical health impairment, and psychosocial resources on functional ability. Seventy-eight community-dwelling older adults, age 60 to 75 years, were divided into two groups based on their depression scores. Depressed and nondepressed participants were not significantly different for demographic and illness characteristics, except for income. Depressed participants reported greater physical health impairment, decreased social resources, diminished feelings of economic well-being, and decreased functional abilities when compared to nondepressed participants. Mastery, physical health impairment, and social support were significant predictor variables of functional ability. Mastery was a particularly salient psychological resource for total functional abilities and interventions should be targeted to increasing mastery in older adults. PMID- 11276619 TI - Patient life stories and current situation as told by carers in nursing home wards. AB - Knowing a patient's life story is important for good nursing care of frail and vulnerable elderly people with cognitive impairments. The aim of the study was to compare patients' life stories and current situations as told by carers before and after 1 year of supervision, in which the Resident Assessment Instrument was used as a basis for individualized nursing care. Qualitative content analysis was used to disclose changes and to enable descriptions of patterns. After the intervention, two overall perspectives emerged from the analysis: the patient as a unique person with resources and abilities, despite limitations, and the carers' awareness of their own professional approach. It seemed as if the supervision and the use of a comprehensive and detailed assessment tool contributed to increased knowledge about the patients and to efforts to see them as real persons behind the dementia surface. PMID- 11276620 TI - Caregiving in nursing homes. Views of licensed nurses and nursing assistants. AB - This investigation is a qualitative study of the views held by 36 licensed nurses (25 registered nurses and 11 licensed practical nurses) and 40 nursing assistants regarding caregiving in nursing homes. Because these care providers are most directly involved in the delivery of care, their views are important as determinants of quality of care. Study findings focus on the extent to which nurses and nursing assistants agree on what contributes to good care and how they perceive the work that each does. Also reported are their perceptions regarding factors that make care delivery easy or difficult. Results suggest that nurses and nursing assistants share selected perceptions about the division of labor in the nursing home. Also evident are areas of less agreement among these members of different status sets. A discussion of how these caregivers can work together as effective team members is presented. PMID- 11276621 TI - Fluctuations in established infant pain behaviors. AB - This study compares behaviors that differed across levels of established (e.g., nonprocedural) infant pain with those that differed between periods of greater and lesser distress within any level of infant pain. Sixty-four videotaped infants of two ages (0 to 3 months and 7 to 12 months) and four levels of established infant pain (none, mild, moderate, and severe) were used. Pain was from medical or surgical causes. Behaviors were compared between the most distressed (HI) and the least distressed (LO) video segments per infant and across the four levels of infant pain using a two-level (distress and level of pain) MANOVA. Many behaviors were indicative of high levels of established pain and greater distress. Others increased with greater distress but lower levels of pain. Findings suggest that many behaviors indicative of high distress that constitute the immediate infant pain response are not good indicators of levels of established infant pain. PMID- 11276622 TI - The conceptualization, measurement, and validation of transient mechanical birth trauma. AB - An index of transient mechanical birth trauma (TMBT), consisting of the presence or absence of molding, cephalohematoma, subconjunctival hemorrhage, body bruising, facial bruising, petechiae, forceps marks, diminished arm movements, and sensitivity to sudden position changes, was measured on a convenience sample of 196 healthy newborns. Six dimensions of the Neonatal Behavioral Assessment Scale (NBAS) and other newborn measures also were assessed. Vaginally delivered newborns had more TMBT than those delivered by cesarean section and of newborns delivered vaginally, macrosomics had more TMBT than nonmacrosomics. TMBT positively correlated with range of state, individual reflex items of resistance to left and right arm movement, predominant state during the NBAS exam, and time to complete the NBAS exam. TMBT negatively correlated with newborn state instability and 1- and 5-minute Apgars. The results supported the measure's validity and are discussed in terms of implications for practice and further research to explore TMBT's usefulness. PMID- 11276623 TI - Family role and work adaptation in MI women. AB - The purpose of this longitudinal, randomized clinical study was to assess if family role functioning and work role adaptation in women following a myocardial infarction was improved by a nursing instructional intervention. The 122 participants entered the study at their discharge phase of hospitalization. The treatment group, who received the nursing instructional program, demonstrated increased adaptation related to family role over time, had less symptoms and concerns with sexual activity, and had higher adaptation scores in resuming the paid work role. For both groups, the role most impacted across time was the homemaker role. Nursing implications for assessing role disruption in women with myocardial infarction are described. PMID- 11276624 TI - Cognitive function and quality of life in interferon therapy for melanoma. AB - The purpose of this pilot study was to describe short- and long-term changes in cognitive function and quality of life in patients with melanoma receiving interferon (IFN) alpha-2b. This study used a three-group, repeated measures design in which cognitive function and quality of life were evaluated prior to initiation of treatment at 3-month intervals during treatment and 3 months following the completion of treatment. The sample consisted of 16 adults with Stage II or III melanoma, randomized to one of three treatment groups. Participants in Arm A received high-dose IFN alpha-2b, those in Arm B received low-dose IFN alpha-2b, and those in Arm C received no therapy (control). No significant changes in cognitive function were detected. In participants in Arm A, there was a significant deterioration in the physical well-being dimension of quality of life from baseline to 3 months after beginning therapy. PMID- 11276625 TI - Certified Dialysis Nurse (CDN) examination: exploring the issues. PMID- 11276626 TI - Enriching our practice through culturally competent care. PMID- 11276627 TI - Patient-reported quality of life early in dialysis treatment: effects associated with usual exercise activity. AB - The purpose of this study was to investigate factors associated with quality of life (QoL) early in treatment in a cohort of incident (i.e. newly diagnosed) dialysis patients. This multicenter study investigated QoL reported by patients on chronic hemodialysis (HD) and peritoneal dialysis (PD) at approximately 60 days following the start of treatment. QoL was assessed by the Medical Outcomes Study Short-Form 36 (MOS-SF 36) and by disease-targeted scales from the Kidney Disease Quality of Life (KDQOL). Patient's QoL as measured by the SF-36 was substantially impaired compared to norms for the general population. In univariate analyses, patients' QoL scores were related to demographic variables (age, race, sex, educational level), clinical variables (predialysis BUN and serum creatinine, primary diagnosis of diabetes, cardiovascular comorbidity, average hematocrit and serum albumin in first months of treatment), dialysis variables (HD/PD modality, PD dialysis adequacy, facility patient-staff ratio) and patient's level of usual exercise activity. In multivariate analyses, the most important independent QoL predictor was patient's usual level of exercise activity. Exercise activity independently predicted two performance measures of physical functioning, maximal gait speed and repeated chair rises, as well as patient-perceived physical functioning. Continued study of patient outcomes in relation to adequacy of delivered dialysis, early versus late diagnosis of chronic renal failure (CRF), and patient's usual exercise activity is important because these variables can be the focus for intervention strategies to prevent early deterioration in dialysis patients' functional health status. PMID- 11276628 TI - Symptom distress and day-to-day changes in functional status in chronic hemodialysis patients. AB - The purposes of this study were to examine changes in center-based hemodialysis patients' levels of daily functional status (FS) during the week and to examine the extent to which symptom distress explained variations in daily functional status. A correlational panel study design was used. The sample consisted of 104 chronic incenter hemodialysis patients recruited from two outpatient hemodialysis centers in a mid-Atlantic state. The Inventory of Functional Status was administered daily for 7 consecutive days to assess daily functional status. The Kidney Disease Quality of Life Symptom Scale was administered daily for 7 consecutive days to examine daily symptom distress. Study findings indicated suboptimal levels of FS at baseline, a significant decline in FS the day before the first dialysis session of the week compared to baseline, and a significant decline in FS on the second dialysis day of the week compared to the previous and subsequent nondialysis days. Symptom distress explained 6% of the variation in FS on these 2 days. In conclusion, incenter hemodialysis patients' baseline levels of FS are suboptimal, and FS declines further on the day before the first dialysis session of the week and the second dialysis day of the week. Symptom distress partially accounts for the decline in FS on those days. These findings indicate a need for ongoing FS assessment, implementation of strategies to improve FS, and symptom management in center-based chronic hemodialysis patients. PMID- 11276629 TI - Subjective burden in young and older African-American caregivers of patients with end stage renal disease awaiting transplant. AB - The purposes of this study were to identify the level of subjective burden reported by African American caregivers of patients diagnosed with end stage renal disease (ESRD) awaiting transplantation and identify whether subjective burden, personal strain, and role strain varied by caregiver age. An exploratory descriptive survey design was used. The convenience sample consisted of 78 African American family caregivers. Subjects were obtained from a University transplant clinic in the Mid-South. Caregivers completed the 22-item self administered Burden Interview (BI) and a demographic data form. Data were analyzed using descriptive statistics and the appropriate parametric and nonparametric tests of group differences. Most caregivers, reported little to none or mild to moderate burden. Results also indicated that there was no significant difference in the level of overall burden, personal strain, and role strain reported by young and older caregivers. Additional findings revealed that burden was least in the caregivers of patients who independently performed activities of daily living (ADL) and greatest among the caregivers of patients assisted by someone other than the caregiver with ADL. Findings from this study may help nurses to assist caregivers to identify their level of burden early in the caregiving process. Implementation of early interventions may prevent negative psychological and physical outcomes in these caregivers. PMID- 11276630 TI - Self-care strategies to reduce fluid intake and control thirst in hemodialysis patients. AB - The purpose of this pilot study was to describe how often hemodialysis (HD) patients used particular self-care strategies to reduce fluid intake or relieve thirst, how effective each strategy was perceived to be, and whether they planned to use that strategy in the future. A cross-sectional descriptive design was used. The sample of 20 participants from one Midwest dialysis unit verbally responded to the self-care items that were presented. Most of these patients reported they regularly avoided the sun, took medications with mealtime fluid, limited salt on their foods, avoided salty foods, stayed busy, stayed away from fast food restaurants, drank ice-cold drinks, and drank only when thirsty to reduce fluid intake and relieve thirst. They also planned to use these strategies in the future. Interestingly, the strategies perceived as most effective- measuring daily allotted fluids in a pitcher to drink from all day, spacing liquids over the entire day, and measuring the amount of fluid they drank--were not the same as those they used most. Replication of these findings with larger samples is strongly recommended. PMID- 11276631 TI - Documentation of anemia management interventions. Case study of the anemic patient. AB - Documentation is an integral part of good anemia management. In addition to relaying vital clinical information, documentation demonstrates how nurses and other medical professionals are striving to achieve the standards of care mandated by law, their profession, third party payers, and individual health care providers. Proper documentation of anemia management practices should include a written chronological history of the rationales, plans, interventions, and evaluations initiated to achieve patient-specific Hb/Hct target levels across the spectrum of care. PMID- 11276632 TI - Catheter brushing has made a sweeping difference to our clinical practice. AB - Our experience thus far has been very positive with a zero complication rate. This fact coupled with the benefits of minimizing delays in dialysis schedules, the 'speed' at which the procedure can be performed, and the cost-effectiveness of brushing results in our unit now choosing this technique as first line management for blocked catheters. From the perspective of the patient, there is little or no discomfort associated with the procedure. The simple principles of the brushing technique mean the procedure can be performed at the bedside by an accredited medical officer registrar or registered nurse. Explanation of the procedure and discussion about the benefits of endoluminal brushing versus catheter replacement, coupled with a zero complication rate, has gained the approval of patients. The endoluminal brushing procedure takes approximately 30 minutes to perform, compared with several hours the patient must wait to have a catheter replaced. This results in minimal delays in dialysis schedules and improved client satisfaction. Catheter occlusion and the inconvenience associated with catheter replacement was a very real problem for our HD unit. Our experience to date supports the use of endoluminal brushing as a cost-effective and timely means of restoring catheter patency. PMID- 11276633 TI - Capturing Epogen overfill. PMID- 11276634 TI - Ascorbic acid use in hyporesponders to Epoetin alfa. AB - I.v. ascorbic acid has been used in an effort to mobilize ferritin stores in hyporesponsive HD patients receiving Epoetin alfa. However, not all patients who respond to i.v. ascorbic acid therapy will have subsequent decline in feritin stores (Gastaldello et al., 1995; Tarng & Huang, 1998). Additionally, predicting those patients who will overcome their Epoetin alfa hyporesponsiveness remains unclear. Ascorbic acid's effect on hemosiderin deposits may be another possible mechanism to the increased Epoetin alfa response observed in some HD patients (Hemosiderin is a pathologic deposition of iron in tissues including the spleen, small intestine, and bone marrow). Although there are no well-controlled studies evaluating hemosiderin and i.v. ascorbic acid, it should be noted that subjects with scurvy often present with excessive iron deposits in the tissues, indicating the possible effects of ascorbic acid on hemosiderin metabolism (Bothwell et al., 1964). Ascorbic acid deficiency is often present in many HD patients due to its removal during dialysis and lack of dietary intake (Ponka & Kuhlback, 1983). It remains controversial whether oral ascorbic acid supplementation is indicated in patients receiving HD. Therefore, the Recommended Daily Allowance (RDA) of 60 mg/day should be advised (Makoff, 1999). I.v. ascorbic acid should be considered as a possible adjuvant to therapy in patients who are "iron-overloaded" and hyporesponsive to Epoetin alfa. Although the long-term effects of i.v. ascorbic acid on HD patients is unknown, the potential risk of secondary oxalosis should be considered (Costello, 1991; Pru, Eaton, & Kjellstrand, 1985). It may be necessary to monitor plasma oxalate levels if long-term therapy with i.v. ascorbic acid is used. Clinical studies have examined i.v. ascorbic acid doses from 300 mg-500 mg given up to TIW for a maximum duration of 12 weeks without any significant deleterious effects (Gastaldello et al., 1995; Tarng & Huang, 1998; Tarng et al., 1999). However, large-scale, prospective, and controlled trails are needed to determine the long-term safety and efficacy of i.v. ascorbic acid therapy in iron overloaded HD patients receiving Epoetin alfa. PMID- 11276635 TI - Certified Dialysis Nurse (CDN) examination: pros and cons. PMID- 11276636 TI - The science of quality can work for you. PMID- 11276637 TI - Electrical stimulation as a treatment for stress incontinence. AB - Much research has been conducted into the use of electrical stimulation to restore function in weak/atrophied muscle and it is used widely in the field of muscle rehabilitation. As stress incontinence is a condition which is the result of pelvic floor muscle weakness, it is thought that the symptoms of this condition may be alleviated once the strength and endurance characteristics of this muscle group have been improved. Many studies have been conducted to evaluate the efficacy of various types of electrical stimulation, although definitive conclusions have yet to be drawn. Current forms of electrical stimulation for stress incontinence involve the use of uniform frequencies. In animal studies this type of stimulation has been shown to have drawbacks that are unacceptable when trying to rehabilitate muscle. Consequently, there is a need to develop more physiological patterns of stimulation that will enhance both strength and endurance characteristics without causing premature fatigue. PMID- 11276638 TI - Enabling disabled people: responsibilities of nurse education. AB - This article examines some evidence concerning the approach nurse education takes in relation to disability issues and considers whether it is 'disabling' or 'enabling'. The role and purpose of education and training, along with an analysis of simulation as a teaching method and skills development, are explored. It is argued that nursing curricula has a key responsibility in ensuring that nursing practice is enabling rather than disabling in relation to patients with impairments who may be classed as disabled people by themselves or within society more generally. PMID- 11276639 TI - CliniSorb activated charcoal dressing for odour control. AB - The potential for charcoal cloth in wound management relates to its well demonstrated ability to adsorb small gas molecules. Charcoal has become increasingly used to contain odour and is especially useful in the management of fungating lesions. This article looks at CliniSorb activated charcoal odour control dressing (distributed by CliniMed) and describes its construction, action and place in wound management. PMID- 11276640 TI - Reinforcing the medical stereotype of nursing. PMID- 11276641 TI - Nursing has a chance to stop the erosion of care. PMID- 11276642 TI - Report requests more flexible community care. PMID- 11276643 TI - Students whose application to the nursing register is not supported. PMID- 11276644 TI - Selecting the correct intravenous device: nursing assessment. AB - This article provides information for nursing and medical teams involved in the selection of intravenous (i.v.) devices for patients requiring venous access. For more than 25 years central venous access has been the mainstay for delivery of a variety of i.v. therapies. However, although the use of central venous access has facilitated innovative patient care, it has also precipitated many complications arising from inappropriate catheter selection. The information contained in this article is not intended to replace local policies but is offered as a reference for those involved in developing protocols for assessment of patients requiring therapies via the i.v. route. It aims to assist nurses in relation to matching an appropriate access device to individual patient requirements. PMID- 11276646 TI - Role of community mental health nurses for people with dementia. AB - This is an exploratory study into how eight family carers of people living with dementia perceived the role of their community mental health nurse (CMHN). Using a descriptive qualitative approach with semistructured interviews, areas of the CMHNs' role that the eight carers found supportive were identified and discussed under four main themes: (1) emotional support and advice (2) liaison and networking (3) training and information giving and (4) isolation and loneliness. The study also highlighted certain qualities of the CMHN which were found to be valuable and helpful to the carers. This study suggests that the carers of people living with dementia have many varied needs and they perceive the role of the CMHN as very important and valuable in terms of supporting them in caring for their loved ones. Although not intended as a focus of the study, the carers also highlighted certain desirable qualities in the CMHN that may have implications for all those involved in ensuring a high standard of community mental health nursing service to this client group. PMID- 11276645 TI - Communication is the essence of nursing care. 2: Ethical foundations. AB - This is the second of two articles analysing the communication themes that emerged from a role play situation in the classroom where a group of MSc nursing students carried out a nursing role play. Feedback from the entire group of 19 students was obtained and discussed openly following the presentation of the scenario. The first article examined the following communication issues: breaking bad news; giving information through exploring, questioning, and reassurance; touch; and empathy and humanism (Vol 9(14): 931-8). Attempts were made to give examples of how these key issues could be applied to nursing practice. This second article involves the examination of the ethical issues of truth-telling and autonomy that emerged in the role play and again attempts to integrate the theoretical underpinnings with nursing practice in the knowledge that the arguments advanced are neither right nor wrong. This deduction is based on the fact that each human being is unique; therefore, each interaction that occurs between two people is also unique. Nursing can strive to provide better, more effective and more sensitive communication because ethical communication is, it is argued, the realm in which nurses toll. PMID- 11276647 TI - Interventions in forensic psychiatry: the caregiver's perspective. AB - Although a number of official reports have identified the needs of the caregivers of mentally disordered offenders (forensic clients), there has been limited research in this area. However, some consistent themes have started to emerge from the literature. The burdens being faced by healthcare professionals working with forensic clients are distinct from that of those working with non-forensic clients, with levels of violence directed towards the former being much higher. Levels of professional and social contact are also much reduced for healthcare professionals working in forensic psychiatry. New research seems to show that illness representations (beliefs) have an effect on the coping responses employed by the caregivers of forensic clients. This may be an area that caregivers can focus on in the future to both develop interventions and improve levels of contact and satisfaction with healthcare professionals. PMID- 11276648 TI - The knowledge and attitudes of mental health nurses to electroconvulsive therapy. PMID- 11276649 TI - Outcomes management: from concepts to application. AB - This article provides an introduction to the definition of and rationale for outcomes management and includes a brief review of the outcomes management literature. A model for outcomes management, which links processes that can be changed in care delivery to outcomes that can be measured in a patient population, is reviewed. Guidelines for application of the outcomes management model and practical examples of application to two surgical patient populations are presented. Finally, issues important to outcomes management as a tool for performance improvement are discussed. PMID- 11276650 TI - Clinical decision support systems for outcome measurement and management. AB - Clinical decision support systems are the foundation for outcome management programs through the measurement of specific outcomes, data storage, data analysis, predictive modeling, and risk-adjusted comparison of actual outcomes with predicted outcomes. Many clinical decision support tools or databases are available to clinicians. This article reviews two widely available tools that provide clinical decision support for critical care clinicians, the Project IMPACT and APACHE III Critical Care Series clinical decision support systems. These tools are discussed with regard to risk adjustment methodology, validity, reliability, database size and representation, retrospective and prospective data and analysis, and quality control. Clinical application of clinical decision support systems for benchmarking and use in process improvement and outcome management is reviewed. PMID- 11276652 TI - Right patient? Right bed? A question of appropriateness. AB - Key issues addressing appropriateness of triage decisions and the tools needed to support those decisions are identified in this article. The limitations of current approaches to appropriate utilization such as admission and continued stay criteria, medical management models, bed control and bed management strategies, and rounding practices are discussed, and a systemic model to examine the placement of patients is proposed. Determining which patients can benefit from critical care and which can benefit from an alternative level of care is analyzed through the use of clinical decision support tools that provide both retrospective analysis of current patterns and predictive models to assist the clinician in making continued-stay decisions. Patient populations who are considered for alternative placement are defined. Those populations identified as having the potential to gain limited benefit from the level of intensity of an intensive care unit are managed in alternative sites. The role played by the advanced practice clinician in using clinical decision support tools is discussed. PMID- 11276651 TI - Enhancing outcomes: guidelines, standards, and protocols. AB - Guidelines, protocols, and standards have gained attention as clinical tools designed to enhance clinical decision-making and practice. Yet, evidence has emerged that clinicians are having difficulty integrating new knowledge presented by these tools into practice. This article explores the benefits and barriers to using guidelines and protocols, probes the issues of producing these tools, and examines processes that are critical to constructing valid tools. The key functions important in successful development and implementation of guidelines and protocols are presented, as well as the direction these clinical adjuncts will take in the future. PMID- 11276653 TI - Health information systems: challenges for the 21st century. AB - No longer focused on mere automation of manual processes, healthcare technology is poised to transform practice. Healthcare information systems can extend and enhance the memory; streamline administrative processes; provide access to information where, when, and how it is needed; and manage the cost of care while protecting and improving clinical quality and customer satisfaction. To reap the benefits of information systems, healthcare professionals must take stock of the industry's current position on the road to transformation and determine how to manage the journey ahead. This article explores healthcare information technology trends, discusses emerging technologies such as the Internet and the computerized patient record, and offers future recommendations for achieving technology integration and acceptance. Key to this discussion is the belief that people based skills such as cooperation, leadership, and creative thinking are just as important as--if not more important than--the actual technology. PMID- 11276654 TI - Implementation strategies to manage heart failure outcomes. AB - In the United States, chronic systolic heart failure causes a great economic burden. Pharmacologic and nonpharmacologic therapies must be tailored to the pathophysiologic cause with the ultimate goal of promoting regression or preventing progression of left ventricular remodeling. When this goal is met, symptoms are reduced, quality of life is improved, and morbidity and mortality are decreased. Specific objectives in a nurse-managed heart failure clinic are to improve exercise tolerance, decrease symptoms, and prevent or reduce emergency department visits and acute hospital admissions. Before a nurse-managed outpatient program for heart failure care is implemented, the team must address specific management issues and controversies in heart failure. Actions must focus on chronic disease management rather than just episodic care. Written protocols or algorithms provide guidance in pharmacologic and nonpharmacologic care and ensure that consensus guidelines that offer the best hope of reaching goals are followed. PMID- 11276655 TI - Implementation strategy: one-stop recovery for cardiac surgical patients. AB - "Fast-track" or "rapid recovery" for cardiac surgical patients is enjoying widespread use due to its benefits of increased patient comfort, enhanced quality of care, and cost-savings. Successful implementation of a fast-track program, however, may be challenged by physicians, the institution, or patients and their families. One-Stop Recovery is a fast-track program that emphasizes the benefits of traditional rapid recovery programs while addressing potential challenges. PMID- 11276656 TI - Long-term ventilator management strategies: experiences of two hospitals. AB - Historically, negative clinical and economic outcomes have been associated with patients who need long-term mechanical ventilation. Institutions and clinicians charged with the care of these patients are understandably interested in exploring clinical strategies that assure positive outcomes. This article describes the evidence base for clinical pathways, weaning teams, protocols, and care-managed approaches. In addition, the article describes how different elements of system initiatives designed for a university teaching hospital and a community hospital were implemented and evaluated. The systematic approaches encourage multidisciplinary input and decrease variation, thus improving both quality and cost. PMID- 11276657 TI - Strategies for assessing outcomes in the elderly in acute care. AB - The monitoring of outcomes is an essential component in evaluating healthcare, yet in critical illness, outcome assessments can be challenged by several factors. These factors include changing acuity levels, recurrent illness, readmission to hospitals, and patient attrition. Additional issues, particularly for elderly patients, include selecting appropriate outcome measures, overcoming barriers to patient recruitment for research, and using retention measures. This article explores issues in conducting outcomes research in acute care and uses a longitudinal prospective panel research study of elderly patients after critical illness as an example. Strategies are discussed that can be used in elderly subjects to facilitate accurate data collection in acute care outcomes research. PMID- 11276658 TI - Incorporating outcomes research into clinical practice: the four-step approach. AB - Outcomes research is designed to develop new and generalizable knowledge about care delivery, interventions, and patient outcomes. Advanced practice nurses are asked not only to participate in outcomes research but also to develop and maintain outcomes research projects and databases and to demonstrate the effectiveness of their own care practices through outcomes research. A four-step approach is described as an easy way in which to help advanced practice nurses engage in outcomes research at any site and with any patient population. The four steps are: Step I: identify and describe current practice in nursing-medical care and its outcome(s); Step II: data collection; Step III: data analysis; and Step IV: praxis: putting the research into practice. Participating in outcomes research will allow advanced practice nurses to improve healthcare services and to secure the role of advanced practice nurses in the changing healthcare arena. PMID- 11276659 TI - Leadership: a skill, not a role. AB - The ever-changing healthcare landscape demands leaders who can help navigate through the turbulence. At a time when the demand for leaders in healthcare continues to grow, many wonder whether there is a pending leadership shortage. This perception is fostered by the confusion about how leaders are defined. For many the definition of a leader is synonymous with a job title or role, such as a Manager or Director. Leadership, however, is not a job description; rather, it is influencing others to contribute to a positive outcome. All clinicians, particularly advanced practice nurses, must take the responsibility of finding solutions to the current challenges in healthcare and work through others to achieve a better outcome. In this article leadership is defined by skill sets, not job title, and offers specific strategies to enhance advanced practice nurses' leadership skills. PMID- 11276660 TI - Has the RCN outlived its usefulness? PMID- 11276661 TI - Busman's holiday. PMID- 11276662 TI - Resolve to resist bigotry. PMID- 11276663 TI - In the bag. PMID- 11276664 TI - Blood minded. PMID- 11276665 TI - The government's home loans offer is an admission that nurses aren't paid enough. PMID- 11276666 TI - A lot on its plate. PMID- 11276667 TI - Anaphylaxis: emergency management. PMID- 11276668 TI - Nursing language. PMID- 11276670 TI - Targeting young men. PMID- 11276671 TI - Middle-aged men's health. PMID- 11276669 TI - A training partnership in cervical cytology. PMID- 11276672 TI - NT open learning: clinical supervision--Part 11. Negotiating management and staff needs. PMID- 11276673 TI - Management of a central venous catheter. PMID- 11276674 TI - Practical pointers on Parkinson's. PMID- 11276675 TI - Prescription for change. PMID- 11276676 TI - Willing and able. PMID- 11276677 TI - At the heart of the service. PMID- 11276678 TI - Stress busters. PMID- 11276679 TI - Breaking through the pain barrier. PMID- 11276680 TI - Armed with awareness. PMID- 11276681 TI - Big boys do cry. PMID- 11276682 TI - Breaking away from violence. PMID- 11276683 TI - Who pays the price? PMID- 11276684 TI - Put racism on the record. PMID- 11276685 TI - International trade agreements. PMID- 11276686 TI - Patients with mental illness: general nurses' attitudes and expectations. AB - AIM: To measure the emotional reactions and expectations of 64 nurses in a general hospital to vignettes describing patients with unstable diabetes and a co morbid psychiatric diagnosis. METHOD: A small scale questionnaire survey was used in a within-groups design. RESULTS: Findings suggest that the nurses in the sample were fearful of people with a mental health problem. They were wary of possible unpredictable behaviour. Qualified staff generally felt better equipped to cope with such patients, depending on their psychiatric experience. CONCLUSION: General/adult nurses who have had more exposure to patients with mental health problems during their initial training are more likely to feel adequately prepared for managing people with mental health problems. PMID- 11276687 TI - The implications of drug treatment and testing orders. AB - The Crime and Disorder Act (1998) set out ways to tackle the link between crime and drug use. This article aims to review the Act and its implications on healthcare professionals. It also examines the contents of the Act in relation to the proposed Drug Treatment and Testing Orders (DTTO). PMID- 11276688 TI - Insider/outsider partnerships in an ethnographic study of shared governance. AB - The benefits of research being undertaken by more than one researcher cannot be underestimated. Having one researcher with intimate knowledge of the organisation and another who could provide a dispassionate view, paid dividends in this study into shared governance from an ethnographic perspective. PMID- 11276690 TI - Nursing the patient with coeliac disease. PMID- 11276691 TI - Master plan. PMID- 11276689 TI - Catheter care in the community. AB - The practice of healthcare professionals performing routine catheter changes and the education of patients and carers regarding these procedures is discussed. Patients' sexual needs when catheterisation is used as a method of urinary management are considered, and options for maintaining a normal life style are described. PMID- 11276692 TI - A healthy state of mind. PMID- 11276693 TI - Seller beware. PMID- 11276694 TI - A class above. PMID- 11276695 TI - Under pressure. PMID- 11276696 TI - In all honesty. PMID- 11276697 TI - Happysurfing. PMID- 11276698 TI - A force for the future. PMID- 11276699 TI - They give with one hand.... PMID- 11276700 TI - Give me a break. PMID- 11276701 TI - Prescription for change. PMID- 11276702 TI - Art&science. Children in adult outpatients. PMID- 11276703 TI - The efficacy of bilingual health advocacy in ethnic minority patients with cancer. AB - AIM: This research aims to establish the efficacy of introducing trained bilingual health advocates to non-English speaking cancer patients. METHOD: Male and female Bengali advocates received appropriate training. They were then given a group of patients to manage, while a control group received no such intervention. Outcomes were determined at the baseline and after three months. The study finally concludes in April 2000. RESULTS: The progress so far shows that the advocates had only recruited half of the expected number of Bengali cancer patients. Focus groups showed, however, that healthcare professionals felt that their training was inadequate to overcome the language and cultural barriers, and many were distressed that they were not meeting the needs of minority ethnic patients. CONCLUSION: The authors anticipate that this study will concur with research in other health sectors where bilingual health advocacy has been beneficial, and that future care will be better informed as a result. PMID- 11276705 TI - Dysphagia in stroke patients. AB - Practice profiles are reflective pieces written by nurses in practice and based on continuing professional development articles. This week Maria Rees discusses stroke and dysphagia. Article no. 489. Davies S (1999) Dysphagia in acute strokes. PMID- 11276704 TI - Infusion devices: risks, functions and management. AB - The use of infusion devices to administer intravenous solutions and drugs is common practice throughout a variety of healthcare settings. The risks associated with such practice are wide and varied. Training, maintenance, clinical need and availability of infusion devices are key areas that need to be addressed when managing associated risks. Nurses have a key role to play in addressing these risks. PMID- 11276706 TI - Consumer views of pressure sores: a preliminary survey. AB - AIM: To obtain consumer views about pressure sore development, particularly in A&E departments. METHOD: The local Patient and Carers Association received a freepost survey in the form of a simple fixed choice questionnaire. RESULTS: Fewer than half the questionnaires were returned. Relatively few respondents were aware that pressure sores could be avoided, and few were aware that prolonged bed rest or lying on a hospital trolley could increase the risk. CONCLUSION: The survey raised more questions than it managed to answer. More research is needed into the views of patients and carers before they can be involved in healthcare planning. PMID- 11276707 TI - The provision of leg ulcer services by practice nurses. AB - As part of their implementation programme of leg ulcer guidelines, the group conducted a study of the caseload and attitudes and training of practice nurses with relation to leg ulcer management. They begin by outlining the prevalence and treatment methods of leg ulcers, then outline their study and put forward their resulting recommendations. PMID- 11276708 TI - Foot ulceration in diabetic patients. AB - Foot care is very important if patients with diabetes are to avoid ulceration complications. Kate Springett explains the importance of educating the patient to be aware of signs and symptoms of foot ulceration, and outlines the best management techniques within the scope of a multiprofessional care team. PMID- 11276709 TI - A fertile field. PMID- 11276710 TI - Something's happened to the forums.... PMID- 11276711 TI - Is having strong clinical skills the measure of a good nurse? PMID- 11276712 TI - Working with dinosaurs. PMID- 11276713 TI - Stop closing your eyes to sex and sexuality. PMID- 11276714 TI - Change must come from the top. PMID- 11276715 TI - Dummy run. PMID- 11276716 TI - Don't knock the opportunities PCGs offer community nurses. PMID- 11276717 TI - Excluded. PMID- 11276718 TI - NT learning curve. Resources for better practice. A framework of care for children with cerebral palsy. PMID- 11276719 TI - A nurse-led clinic for peritoneal dialysis. PMID- 11276720 TI - Lymphoedema: a clinical update. PMID- 11276721 TI - Don't ignore it and it will go away. PMID- 11276723 TI - The nurse consultant: an advanced nurse practitioner? PMID- 11276722 TI - Dramatic effects. PMID- 11276724 TI - Some additional benefits of clinical supervision. PMID- 11276725 TI - Central venous catheterisation--2. PMID- 11276726 TI - Care to the end. PMID- 11276727 TI - Embracing technology. The last frontier--technology in the operatory. PMID- 11276729 TI - Indebted to the profession. Young dentists take student loan debt in stride. PMID- 11276728 TI - Liability. PMID- 11276730 TI - Media training. PMID- 11276731 TI - Picture perfect: selecting an intraoral camera system. PMID- 11276732 TI - The name game. PMID- 11276733 TI - Sweet charity? PMID- 11276734 TI - Osseoperception: sensory function and proprioception. AB - Tooth loss and its replacement have significant functional and psychosocial consequences. The removal of intra-dental and periodontal mechanoreception accompanying tooth loss changes the fine proprioceptive control of jaw function and influences the precision of magnitude, direction, and rate of occlusal load application. With the loss of all teeth, complete denture restoration is a compromise replacement which only partially restores function. Implant-supported prostheses restore jaw function more appropriately, with improved psychophysiological discriminatory ability and oral stereognosis. Osseoperception is defined as depending on central influences from corollary discharge from cortico-motor commands to jaw muscles, and contributions from peripheral mechanoreceptors in orofacial and temporomandibular tissues. The processing of central influences is considered with the recognition of the plasticity of neuromotor mechanisms that occurs to accommodate the loss of dental and periodontal inputs. PMID- 11276735 TI - Influence of inflammatory reactions vs. occlusal loading on peri-implant marginal bone level. AB - Plaque accumulation on abutments or implant surfaces induces an inflammatory reaction in the gingiva/alveolar mucosa just as around teeth. The longevity of oral implants can be jeopardized by either peri-implantitis and/or an occlusal overload. In the partially edentulous patient in whom pockets around teeth act as a reservoir for the colonization of the pockets around implants, the risk for inflammatory reactions of the peri-implant soft tissues seems especially more plausible than in the fully edentulous patient. This is especially true for implants with a very rough surface (e.g., plasma-sprayed), because of the positive relationship between surface roughness and supra- as well as subgingival plaque formation. Several medium-term (from 5 to 10 years) clinical studies support this hypothesis, through the observation of ongoing bone loss and subsequent decreasing success/survival percentages. Occlusal overload increases the risk for microfractures at the implant-bone interface in two-stage implants, which can result in significant marginal bone loss and even failure. There is ample evidence that occlusal factors are related to marginal angular defects around two-stage implants. PMID- 11276736 TI - Radiographic determinants of implant performance. AB - This paper reviews and compares the strengths and weaknesses of radiographic techniques including periapical, occlusal, panoramic, direct digital, motion tomography, and computed tomography. Practical considerations for each method, including availability and accessibility, are discussed. To date, digital subtraction radiography is the most versatile and sensitive method for measuring boss loss. It can detect both bone height and bone mass changes on root-form or blade-form dental implants. Criteria for implant success have changed substantially over the past two decades. In clinical trials of dental implants, the outcomes require certain radiographic analyses to address the hypothesis or clinical question adequately. Radiographic methods best suited to the objective assessment of implant performance and hypothesis were reviewed. PMID- 11276737 TI - Clinical experiences with dental implants. AB - The clinical utilization of dental implants has accelerated in recent years, and new applications continue to emerge. Concomitantly, alternative implant systems have introduced conceptually different approaches to treatment using altered protocols. The purpose of this paper is to address some of the background issues pertinent to the long-term success, survival, safety, and effectiveness of these devices. The requirements for clinical acceptance of implants are controlled initially by regulatory bodies; however, the dentist eventually must make a decision on which type of implant should be used in clinical practice. This clinical decision-making process should involve the strategy of using an evidence based approach to ensure quality of care and reduction of liability for negligent care. This is particularly the case when treatment is undertaken in identified high-risk categories. While short- to medium-term data have been accumulated on the success rates of several implant systems, it is apparent that long-term data comparing and contrasting the various advantages and disadvantages of different systems do not exist, and adequate criteria applicable to the collective clinical experience need to be defined. Expanding areas of application are dependent on continuous improvements in implant hardware, surgical protocol development, and rationalized osteopromotive and site installation augmentation technology. Many treatment endeavors are still largely at the pilot study level of development, and long-term prospective clinical trials on large numbers of patients are required to document results adequately and to elucidate the most likely productive areas for future investigation. PMID- 11276738 TI - Clinical experience with one-stage, non-submerged dental implants. AB - This review article describes the scientific documentation of one-stage, non submerged dental implants. In the past 25 years, numerous in vivo studies have demonstrated that non-submerged titanium implants achieve osseointegration as predictable as that of submerged titanium implants. This observation was confirmed in prospective clinical studies, mostly done with the ITI Dental Implant System. ITI implants have been widely documented for up to 10 years of prospective follow-up at various centers. All studies showed success rates well above 90%. In summary, the non-submerged approach is a true alternative to the original healing modality with submerged titanium implants. The non-submerged approach offers several clinical advantages: (i) the avoidance of a second surgical procedure and less chair time per patient, resulting in overall reduced treatment cost; (ii) the lack of microgap at the bone crest level, leading to less crestal bone during healing and resulting in a more favorable crown-to implant length ratio; and (iii) a simplified prosthetic procedure, presenting an ideal basis for cemented implant restorations. Due to these significant clinical advantages, the non-submerged approach will become more important in implant dentistry in the near future, particularly in implant sites without esthetic priority. PMID- 11276739 TI - Two-stage implant systems. AB - Since the advent of osseointegration approximately 20 years ago, there has been a great deal of scientific data developed on two-stage integrated implant systems. Although these implants were originally designed primarily for fixed prostheses in the mandibular arch, they have been used in partially dentate patients, in patients needing overdentures, and in single-tooth restorations. In addition, this implant system has been placed in extraction sites, in bone-grafted areas, and in maxillary sinus elevations. Often, the documentation of these procedures has lagged. In addition, most of the reports use survival criteria to describe results, often providing overly optimistic data. It can be said that the literature describes a true adhesion of the epithelium to the implant similar to adhesion to teeth, that two-stage implants appear to have direct contact somewhere between 50% and 70% of the implant surface, that the microbial flora of the two-stage implant system closely resembles that of the natural tooth, and that the microbiology of periodontitis appears to be closely related to peri implantitis. In evaluations of the data from implant placement in all of the above-noted situations by means of meta-analysis, it appears that there is a strong case that two-stage dental implants are successful, usually showing a confidence interval of over 90%. It also appears that the mandibular implants are more successful than maxillary implants. Studies also show that overdenture therapy is valid, and that single-tooth implants and implants placed in partially dentate mouths have a success rate that is quite good, although not quite as high as in the fully edentulous dentition. It would also appear that the potential causes of failure in the two-stage dental implant systems are peri-implantitis, placement of implants in poor-quality bone, and improper loading of implants. There are now data addressing modifications of the implant surface to alter the percentage of osseointegration. New types of reinforcements for dental implants and the use of growth factors to augment bone regeneration so that implants can be placed more easily are now being actively investigated. PMID- 11276740 TI - Ceramic-coated implant systems. AB - Practitioners have used hydroxyapatite-coated (HA-coated) endosseous and subperiosteal implants in various forms for many years. These have included root forms in both screw and cylindrical shapes, blades, and subperiosteals. The clinical predictability remains controversial and subject to claims and counterclaims. The early days of dental implantology involving root-form implants recommended their placement in fully edentulous cases only, and anterior to the maxillary sinus and mental foramen. Today's philosophy and rationale of dental implantology include the placement of a single implant replacing a missing natural tooth (especially where the teeth adjacent to the edentulous site have no caries or restorative experience). Implants are used to replace the natural dentition in one quadrant/segment, often preceded or accompanied by ridge augmentation and/or sinus grafting if sufficient bone is not present. So we have to address the clinical predictability of survival in terms of indications, quantity, and quality of bone. Clinical data and experience suggest that hydroxyapatite-coated (HA) dental implants may (and possibly should) be used in (1) Type IV bone, (2) fresh extraction sites, (3) grafted maxillary and/or nasal sinuses, or (4) with short implants (< or = 10 mm in length). PMID- 11276741 TI - Determination of the success and failure of root-form osseointegrated dental implants. AB - Permucosal osseointegrated dental implants are a highly effective and predictable treatment modality for edentulism. This review discusses some controversial aspects of the definitions for success and failure of root-form dental implants. The discussion will focus on the underlying pathologies that, if untreated, may lead to loss of the implanted device. Few clinical syndromes are described based on human pathological material and clinical presentation. The theoretical chronological relationship between implant loss and the incidence of pathology of the soft- and hard-tissue seal around implants is also discussed. The review also examines the finding that implant failures are not randomly distributed in the treated populations and that implant loss clusters in specific high-risk groups and individuals. Known risk indicators, and possible risk factors, are discussed, taking into account the patient, the reconstruction, the implant, and implant site-specific factors. Particular emphasis is placed on the need for better determination of whether periodontal patients are at higher risk for implant failures as a consequence of their increased susceptibility to infectious, inflammatory-response-driven tissue breakdown. PMID- 11276742 TI - Materials characteristics of uncoated/ceramic-coated implant materials. AB - In this paper, the biocompatibility of dental implant materials is discussed in the context of both the mechanical characteristics of the materials and the type of surface presented to the surrounding tissues. The proper functioning of the implant depends on whether it possesses the strength necessary to withstand loading within the expected range, with other properties such as elongation being of importance in some instances. A suitable modulus of elasticity may be of major importance in situations when optimum load transmission from the implant into the surrounding bone is key to the successful functioning of the device. Dental implants present a wide range of surfaces to the surrounding tissues based on surface composition, texture, charge energy, and cleanliness (sterility). Metallic implants are characterized by protective oxide layers, but ion release is still common with these materials, and is a function of passivation state, composition, and corrosion potential. An effective surface treatment for titanium appears to be passivation or anodization in a suitable solution prior to implantation. Inert ceramic surfaces exhibit minimal ion release, but are similar to metals in that they do not form a high energy bond to the surrounding bone. Some of the newly developed dental implant alloys such as titanium alloys, which contain zirconium and niobium, and high-strength ceramics such as zirconia may offer some advantages (such as lower modulus of elasticity) over the conventional materials. Calcium phosphate ceramic coatings are commonly used to convert metallic surfaces into a more bioactive state and typically cause faster bone apposition. There is a wide range of ceramic coatings containing calcium and phosphorus, with the primary difference in many of these materials being in the rate of ion release. Although their long-term success rate is unknown, the calcium phosphate surfaces seem to have a higher potential for attachment of osteoinductive agents than do uncoated titanium and other more inert implant materials. PMID- 11276744 TI - In vitro models of biological responses to implants. PMID- 11276743 TI - The biologic tissue responses to uncoated and coated implanted biomaterials. AB - Ultrastructural examination of the morphology and morphometry of the bone supporting uncoated titanium and ceramic implants was assessed in an experimental animal model involving 120 implants placed into the mandibles of 30 adult mongrel dogs. Further, preliminary morphologic and morphometric observations of the bone supporting uncoated and hydroxylapatite-coated endosteal titanium implants was evaluated in a second investigation involving 72 implants placed into the mandibles and maxillae of 6 additional dogs. A densely mineralized collagen fiber matrix was observed directly interfacing with uncoated implants. The only material interposed between the implant and bone matrix was a 20- to 50-nm electron-dense material suggestive of a proteoglycan. Also seen in these same osseointegrated implants were narrow unmineralized zones interposed between the implant and bone matrix. In these zones of remodeling bone, numerous osteoblasts were observed interacting with the collagen fiber matrix. It was shown that a normal homeostasis of anabolic osteoblastic activity and catabolic osteoclastic activity resulted in bone remodeling and the resultant osseointegration of the implants. Hydroxylapatite-coated implants intimately interfaced with healthy bone. The mineralized matrix extended into the microporosity of the HA coating. This matrix contained viable osteocytes. PMID- 11276746 TI - Role of the osteoclast at the bone-implant interface. AB - A thorough understanding of the processes of healing, repair, and remodeling of bone is critical for the establishment and maintenance of osseointegration of dental implants. In this regard, much attention has been paid to the anabolic aspects of bone remodeling, including the cell biology of the osteoblast and the various cytokines and growth factors which regulate these processes. In contrast, there is little information on the bone-resorptive activity that occurs around implants during osseointegration, and of the role of osteoclasts, macrophages, and stromal cells in those catabolic processes associated with bone remodeling. This paper reviews osteoclast cell biology, the interaction of osteoclasts and biomaterials, and the information available on osteoclasts and dental implants, and poses some questions for future research. PMID- 11276745 TI - Implant surface characteristics modulate differentiation behavior of cells in the osteoblastic lineage. AB - This paper reviews the role of surface roughness in the osteogenic response to implant materials. Cells in the osteoblast lineage respond to roughness in cell maturation-specific ways, exhibiting surface-dependent morphologies and growth characteristics. MG63 cells, a human osteoblast-like osteosarcoma cell line, respond to increasing surface roughness with decreased proliferation and increased osteoblastic differentiation. Alkaline phosphatase activity and osteocalcin production are increased. Local factor production is also affected; production of both TGF-beta 1 and PGE2 is increased. On rougher surfaces, MG63 cells exhibit enhanced responsiveness to 1,25-(OH)2D3. Prostaglandins mediate the effects of surface roughness, since indomethacin prevents the increased expression of differentiation markers in these cells. PMID- 11276747 TI - Physical and biological characteristics of implant materials. PMID- 11276748 TI - Soft tissue and epithelial models. AB - The applicability of a biomaterial for the manufacturing of oral implants is determined by its physicochemical and geometric surface properties. Research, therefore, is concerned with the cellular reactions that occur when an implant material comes into contact with body tissues. For permucosal oral implants, this involves both the reaction of bone and gingival cells. In vitro cell culturing- including the use of various analytical techniques like light microscopy, scanning and transmission electron microscopy, confocal laser scanning microscopy, and digital image analysis--is a good tool whereby investigators can obtain more insight into the relevant components of implant-tissue adhesion. In the current overview, the role of cell models in oral implant research is discussed, specifically with reference to responses of epithelial cells and fibroblasts. PMID- 11276749 TI - In vitro models of biological responses to implant microbiological models. AB - To study the etiology and explore possibilities for the therapy of implant associated infections, investigators have developed and utilized various in vitro models. Major contributions have come from the non-oral medical field, where device-related infections can create life-threatening situations. Microbiological models may include (i) models to study the reaction of micro-organisms to the presence of implants, (ii) models to study the reaction of implant-associated micro-organisms to antimicrobial agents, and (iii) models to study the reaction of the host tissues to the presence of implants contaminated with micro organisms. In evaluating the potential usefulness of these models for research in oral implantology, one must consider common features as well as important differences between implanted medical devices and oral implants. Although infections associated with implantable medical devices and oral peri-implant infections share a remarkable number of common features, there are also important differences that need attention when findings from in vitro experiments are extrapolated to clinical relevance. PMID- 11276751 TI - Implant surfaces and interface processes. AB - The past decades and current R&D of biomaterials and medical implants show some general trends. One major trend is an increased degree of functionalization of the material surface, better to meet the demands of the biological host system. While the biomaterials of the past and those in current use are essentially bulk materials (metals, ceramics, polymers) or special compounds (bioglasses), possibly with some additional coating (e.g., hydroxyapatite), the current R&D on surface modifications points toward much more complex and multifunctional surfaces for the future. Such surface modifications can be divided into three classes, one aiming toward an optimized three-dimensional physical microarchitecture of the surface (pore size distributions, "roughness", etc.), the second one focusing on the (bio) chemical properties of surface coatings and impregnations (ion release, multi-layer coatings, coatings with biomolecules, controlled drug release, etc.), and the third one dealing with the viscoelastic properties (or more generally the micromechanical properties) of material surfaces. These properties are expected to affect the interfacial processes cooperatively, i.e., there are likely synergistic effects between and among them: The surface is "recognized" by the biological system through the combined chemical and topographic pattern of the surface, and the viscoelastic properties. In this presentation, the development indicated above is discussed briefly, and current R&D in this area is illustrated with a number of examples from our own research. The latter include micro- and nanofabrication of surface patterns and topographies by the use of laser machining, photolithographic techniques, and electron beam and colloidal lithographies to produce controlled structures on implant surfaces in the size range 10 nm to 100 microns. Examples of biochemical modifications include mono- or lipid membranes and protein coatings on different surfaces. A new method to evaluate, e.g., biomaterial-protein and biomaterial cell interactions--the Quartz Crystal Microbalance--is described briefly. PMID- 11276750 TI - Bone induction by implants coated with cultured osteogenic bone marrow cells. AB - The availability of osteoinductive coatings on dental and orthopedic implants will result in an improved fixation of these devices. Those cases where implants are placed in poor-quality bone or where high failure rates are obtained are especially expected to gain from such coatings. This paper presents a novel, biological approach to obtain bioactive and osteoinductive coatings on bone replacement implant materials. This so-called tissue engineering approach utilizes osteogenic bone marrow cells that are cultured on an implant material to form a bone-like tissue. The implant materials used herein included porous calcium phosphate scaffolds and metallic plates, the latter of which were coated with a biomimetic calcium phosphate coating to facilitate cellular attachment. Bone marrow cells were obtained from a variety of species, including humans, and were grown to facilitate cellular proliferation. The cells were subsequently seeded onto the implants and cultured for an additional week to facilitate osteogenic differentiation and extracellular matrix production. The resulting hybrid implants, encompassing the biomaterial carrier and cultured bone-like tissue, were subsequently implanted subcutaneously in nude mice for 4 weeks, followed by histological examination for de novo bone formation. The results revealed that newly formed bone was seen both in porous implants and on flat metallic surfaces. This bone tissue engineering approach, therefore, offers great potential to enhance bony healing around implants in a compromised bone bed. PMID- 11276752 TI - Peri-implant tissue response to pathological insults. AB - With the increased use of osseointegrated implants and with many implants functioning for long periods of time, the soft tissue barrier around implants has become more important. This paper reviews the soft tissue response around implants under healthy and diseased conditions and presents the etiology of peri implant tissue breakdown. Diagnostic techniques such as probing pocket depth, radiographic evidence, and microbial sampling have been analyzed and modified from the periodontal field and used during the maintenance phase of the dental implant. The long-term goal of implant maintenance is to prevent or to arrest the progression of disease, and to achieve a maintainable implant site. Recent reports indicate that peri-implant tissues can be treated with either non surgical or surgical techniques. PMID- 11276753 TI - Biomechanical and functional behavior of implants. AB - The ability to achieve a long-term stable implant interface is not a significant clinical issue when sufficient uni- or bi-cortical stabilization is available. Clinical outcomes studies suggest that the higher-risk implants are those placed in compromised cortical bone (thin, porous, etc.) in anatomic sites with minimal existing trabecular bone (characterized as type IV bone). In establishing and maintaining an implant interface in such an environment, one needs to consider the impact of masticatory forces. These forces, in turn, have the potential to create localized changes in interfacial stiffness through the viscoelastic properties of bone. Changes in these properties will alter the communication between osteocytes and osteoblasts, leading to an increase in new bone growth, a maintenance of established bone, or a loss (potentially catastrophic) of either cortical or trabecular bone. Therefore, a key to understanding the biomechanical and functional behavior at an implant interface is to control the extent of anticipated modeling and remodeling behavior through an optimal implant design combined with a thorough understanding of how tissues respond to the mechanically active environment. PMID- 11276754 TI - Responses of bone cells to biomechanical forces in vitro. AB - In this paper, we review recent studies of the mechanism by which mechanical loading of bone is transduced into cellular signals of bone adaptation. Current biomechanical theory and in vivo as well as in vitro experiments agree that the three-dimensional network of osteocytes and bone-lining cells provides the cellular basis for mechanosensing in bone, leading to adaptive bone (re)modeling. They also agree that flow of interstitial fluid through the lacunar-canalicular porosity of bone, as a result of mechanical loading, most likely provides the stimulus for mechanosensing, and informs the bone cellular network about the adequacy of the existing bone structure. Important signaling molecules involved in in vivo adaptive bone formation, as well as in in vitro cellular response to fluid flow, are nitric oxide and prostaglandins. The expression of key enzymes for nitric oxide and prostaglandin production in bone cells is altered by fluid shear stress in vitro. Together, these studies have increased our understanding of the cell biology underlying Wolff's Law. This may lead to new strategies for combating disuse-related osteoporosis, and may also be of use in understanding and predicting the long-term integration of bone-replacing implants. PMID- 11276755 TI - In vivo bone response to biomechanical loading at the bone/dental-implant interface. AB - Since dental implants must withstand relatively large forces and moments in function, a better understanding of in vivo bone response to loading would aid implant design. The following topics are essential in this problem. (1) Theoretical models and experimental data are available for understanding implant loading as an aid to case planning. (2) At least for several months after surgery, bone healing in gaps between implant and bone as well as in pre-existing damaged bone will determine interface structure and properties. The ongoing healing creates a complicated environment. (3) Recent studies reveal that an interfacial cement line exists between the implant surface and bone for titanium and hydroxyapatite (HA). Since cement lines in normal bone have been identified as weak interfaces, a cement line at a bone-biomaterial interface may also be a weak point. Indeed, data on interfacial shear and tensile "bond" strengths are consistent with this idea. (4) Excessive interfacial micromotion early after implantation interferes with local bone healing and predisposes to a fibrous tissue interface instead of osseointegration. (5) Large strains can damage bone. For implants that have healed in situ for several months before being loaded, data support the hypothesis that interfacial overload occurs if the strains are excessive in interfacial bone. While bone "adaptation" to loading is a long standing concept in bone physiology, researchers may sometimes be too willing to accept this paradigm as an exclusive explanation of in vivo tissue responses during experiments, while overlooking confounding variables, alternative (non mechanical) explanations, and the possibility that different types of bone (e.g., woven bone, Haversian bone, plexiform bone) may have different sensitivities to loading under healing vs. quiescent conditions. PMID- 11276756 TI - Systemic effectors of alveolar bone mass and implications in dental therapy. PMID- 11276757 TI - Oral disease, cardiovascular disease and systemic inflammation. PMID- 11276758 TI - How periodontal disease may contribute to cardiovascular disease. PMID- 11276759 TI - The risk for endocarditis in dental practice. PMID- 11276760 TI - A history of oral sepsis as a cause of disease. PMID- 11276761 TI - Medical management of the patient with cardiovascular disease. AB - Cigarette smoking, hypertension, hypercholesterolemia, and periodontal disease have been established as major risk factors for cardiovascular disease. Dentists and physicians should work aggressively to educate periodontitis patients about this relationship in an effort to improve the quality of health and contribute to their long-term survival. Blood pressure should be checked at the initial dental visit and at each subsequent visit in patients whose blood pressure is found to be high and/or has a history of hypertension. Dental and medical assistants should receive in-service training to assure competency in measuring blood pressures. All staff should be certified in basic cardiopulmonary resuscitation. Emergency protocol procedures should be in writing and rehearsed regularly. Patients should take their blood pressure medication as usual on the day of the dental procedure. It is helpful for the patients to bring all medications to the office for review at the time of the dental procedure. Good communication should be established between the dentist and physician to maximize good dental and physical health. Because the patient with periodontal disease is at an increased risk for cardiovascular disease, a standardized form should be developed for the convenient exchange of vital information, including but not limited to: blood pressure, medications, allergies, medical conditions and pertinent highlights of dental procedures. Minimize stress in patients with coronary artery disease. This includes providing solid local anesthesia, avoidance of intravascular medication injections, and encouraging relaxation techniques. Antibiotic prophylaxis is indicated in patients with valvular heart disease but does not guarantee the prevention of endocarditis. These patients should be alerted to monitor any symptoms such as fever, chills or shortness of breath. It has also been documented that toothbrushing, flossing and home plaque removers can cause transient bacteremia in periodontal patients. Epinephrine use should be avoided or utilized cautiously in patients with pacemakers or automatic defibrillator devices because of the possibility of refractory arrhythmia. Consultation with patient's cardiologist is advised. Anticoagulation with coumadin is not a contraindication to dental procedures. The prothrombin time or international normalized ratio laboratory values should be checked on the day of the procedure to assure that it is in an acceptable range. Aspirin therapy is not a problem unless the patient is on very high doses for severe arthritis. Continuing medical and dental education credits should emphasize cross-training in both areas to insure comprehensive treatment of the patient with periodontal disease. Smoking cessation, regular exercise, a low-fat diet and good dental hygiene contribute to a healthy cardiovascular system. Patients should understand as best we know the relationship between periodontal and cardiovascular disease to afford them an opportunity to improve their overall dental and physical health. PMID- 11276762 TI - Mechanisms of risk in preterm low-birthweight infants. AB - The periodontal diseases share many common risk factors with preterm low birth weight. Examples are, age, socioeconomic status and smoking (Fig. 5). Studies to date have only shown an association between the two conditions, and this does not indicate a causal relationship. However, since the inflammatory mediators that occur in the periodontal diseases, also play an important part in the initiation of labor, there are plausible biological mechanisms that could link the two conditions. The challenge for the future is to characterize the nature of the factors that predispose a mother to give birth prematurely to infants less than 2500 g and to assign relative probabilities to each. Studies are taking place in many parts of the world to determine the probability of a preterm low-birth weight outcome, the interdependence of the factors that contribute to a birth event and possible casual relationships between these factors. Further information about the details of the effects of maternal infection will come from intervention studies, animal studies and more detailed examination of the mechanisms. PMID- 11276763 TI - The medical necessity of periodontal care. PMID- 11276764 TI - Acute-phase reactants in infections and inflammatory diseases. PMID- 11276766 TI - Periodontal management of the patient with diabetes mellitus. AB - Diabetes mellitus is a common medical disorder that will be encountered by every practicing dentist. Knowledge by the dentist of the general and oral signs and symptoms of undiagnosed or poorly controlled diabetes mellitus are essential, and patients displaying these signs or symptoms should receive medical referral. Patients suspected, or known to suffer from undiagnosed or uncontrolled diabetes mellitus should receive only emergency care until their health status has been properly evaluated. In the event the degree of control of a known diabetic is unknown or the patient is poorly controlled, antibiotic therapy should be administered in conjunction with any necessary surgical procedure or in the presence of oral infection. The practitioner must be prepared to manage diabetic emergencies should they occur in the dental office, and hypoglycemic incidents are most likely. The developing therapies now available for medical management of diabetes mellitus suggest that this condition will be more effectively controlled in the future, but medical consultation is important to proper periodontal patient management. New evidence suggests that advanced periodontal disease may interfere with diabetes mellitus control and the physician should be made aware of the patient's periodontal status. Under most circumstances, the well controlled diabetes mellitus patient can receive safe and effective periodontal therapy with some modification of office protocol, and there is little reason to anticipate that the controlled diabetes mellitus patient cannot look forward to a lifetime of periodontal health if proper and timely periodontal therapy is rendered and the patient maintains effective oral hygiene measures accompanied by appropriate supportive periodontal recall therapy. PMID- 11276765 TI - Hyperglycemia, glycoxidation and receptor for advanced glycation endproducts: potential mechanisms underlying diabetic complications, including diabetes associated periodontitis. PMID- 11276767 TI - Dental implant considerations in the diabetic patient. PMID- 11276768 TI - Medical management of HIV-infected patients. PMID- 11276769 TI - Periodontal management of HIV-infected patients. AB - Recognition and diagnosis of the general oral manifestations and specific periodontal manifestations of HIV infection will continue to be a major responsibility of the dental practitioner. The dental team needs to understand the underlying principles of care for the necrotizing and inflammatory periodontal changes associated with HIV. In addition, the practitioner must take into consideration how the presence of opportunistic infections in the periodontium such as Candida as well as the patient's level of immunosuppression may affect conventional approaches to the common forms of periodontal diseases. By combining local and systemic therapy aimed at both preventing and treating oral lesions and periodontal diseases, combined with new systemic antiviral and vaccine therapies, dental and medical practitioners may together help reduce both the dental morbidity and the overall patient morbidity in the HIV infected patient. PMID- 11276770 TI - Periodontal medicine: the emergence of a new branch of periodontology. PMID- 11276771 TI - Post-menopausal bone loss and its relationship to oral bone loss. PMID- 11276772 TI - Treating patients with Alzheimer's disease. The challenges and rewards. PMID- 11276773 TI - Caring for older patients. PMID- 11276775 TI - Negative publicity and public perception. PMID- 11276774 TI - Spread thin: the state of public dental care in Illinois. PMID- 11276776 TI - Changing relationships. PMID- 11276777 TI - Denial of service: should I worry? PMID- 11276778 TI - Access the markets. PMID- 11276779 TI - How to handle an angry patient. PMID- 11276780 TI - Anil Agarwal gives it away. PMID- 11276781 TI - The children's dental clinic at Cook County Hospital. 1924. PMID- 11276782 TI - Identifying and correcting errors for quality panoramic X-rays. PMID- 11276783 TI - Left hand clapping. PMID- 11276784 TI - Up in smoke. PMID- 11276785 TI - Dentistry: 2000 and beyond. PMID- 11276787 TI - 1,000 years of dentistry. Celebrating a millennium of achievement. PMID- 11276786 TI - Divorce-proofing your practice. PMID- 11276788 TI - Preventing dead-end associateships. PMID- 11276789 TI - How to designate a non-spouse beneficiary for your IRA. PMID- 11276791 TI - Hire smart. Hire right--Part 2. PMID- 11276790 TI - Is your browser Y2K ready? PMID- 11276793 TI - Pathways to a peaceful and fruitful practice. PMID- 11276792 TI - Infection control. PMID- 11276794 TI - The decision to fluoridate the Newcastle water supply from 7 October 1968. PMID- 11276795 TI - The earliest licentiates in dental surgery. PMID- 11276796 TI - Origins of orthognathic surgery. PMID- 11276797 TI - Epidemiology and pharmacoeconomic implications of non-steroidal anti-inflammatory drug-associated gastrointestinal toxicity. AB - Non-steroidal anti-inflammatory drugs (NSAIDs) are widely prescribed and used, especially to treat patients with osteoarthritis and rheumatoid arthritis. Since their introduction as a therapeutic class, a large body of literature has accumulated on the side-effects of these drugs. NSAIDs, through their inhibition of prostaglandin synthesis, can affect the renal and cardiovascular systems. However, the majority of reported side-effects are related to the gastrointestinal (GI) system, and the occurrence of these GI events adds significantly to the disease burden. Several factors have been identified that contribute to the risk of an NSAID-associated GI event. However, when considering risk, especially in clinical trials or observational studies, it is necessary to distinguish between baseline risk and NSAID-attributable risk, since this distinction can affect the results and conclusions of the study; NSAID attributable risk is present in subjects who have few or no risk factors for upper GI toxicity. Safer NSAIDs, such as the new specific cyclooxygenase-2 inhibitors, when targeted to the appropriate patient (i.e. those with NSAID attributable risk), should lead to improved outcomes and reduced costs. PMID- 11276799 TI - Health economic models: a question of balance--summary of an open discussion on the pharmacoeconomic evaluation of non-steroidal anti-inflammatory drugs. AB - Pharmacoeconomics and pharmacoeconomic models are increasingly being used to guide health care decisions. In designing and using these models, an appropriate balance must be struck between scientific rigour and model transparency. It is therefore important to consider carefully how the various model components, such as model perspective, internal and external validity, and choice of comparators and outcomes, should be integrated into the model. These factors are discussed in relation to the pharmacoeconomic evaluation of non-steroidal anti-inflammatory drugs. PMID- 11276798 TI - Celecoxib clinical profile. AB - Celecoxib is the first COX-2-specific inhibitor approved for relief of the signs and symptoms of osteoarthritis (OA) and rheumatoid arthritis (RA), as well as for treatment of familial adenomatous polyposis. For both OA and RA, celecoxib has been shown to be significantly superior in efficacy to placebo and similar in efficacy to traditional non-steroidal anti-inflammatory drugs. Its advantage, however, is its gastrointestinal (GI) safety. Randomized clinical trials as well as long-term outcomes studies have demonstrated that the GI safety profile of celecoxib is superior to that of traditional NSAIDs and similar to that of placebo. Additionally, the renal and cardiovascular safety of celecoxib has also become apparent, as well as its efficacy, tolerability and safety in the elderly population. PMID- 11276800 TI - Epidemiology of rheumatic diseases. AB - Rheumatic diseases are among the oldest diseases recognized. The classification of rheumatic diseases is sometimes difficult due to unknown aetiology and heterogeneity in their clinical presentation. Osteoarthritis (OA) and rheumatoid arthritis (RA) are the two most common rheumatic diseases, accounting for a large percentage of disability worldwide. The economic and social burden of these diseases is great. Their impact on both individuals and society results from a decreased quality of life, lost productivity and increased costs of health care. Without appropriate approaches to patient management and control of these diseases, this impact can be expected to increase as the population ages. One of the challenges in studying OA and RA, and rheumatic diseases in general, is deriving epidemiological data that can be used to understand better the factors that contribute to the initiation and progression of these diseases. Only with such an understanding can significant progress be made in the diagnosis, treatment and management of patients. PMID- 11276801 TI - Overview of the arthritis Cost Consequence Evaluation System (ACCES): a pharmacoeconomic model for celecoxib. AB - Pharmacoeconomic analyses have become useful and essential tools for health care decision makers who increasingly require such analyses prior to placing a drug on a national, regional or hospital formulary. Previous health economic models of non-steroidal anti-inflammatory drugs (NSAIDs) have been restricted to evaluating a narrow range of agents within specific health care delivery systems using medical information derived from homogeneous clinical trial data. This paper summarizes the Arthritis Cost Consequence Evaluation System (ACCES)--a pharmacoeconomic model that has been developed to predict and evaluate the costs and consequences associated with the use of celecoxib in patients with arthritis, compared with other NSAIDs and NSAIDs plus gastroprotective agents. The advantage of this model is that it can be customized to reflect local practice patterns, resource utilization and costs, as well as provide context-specific health economic information to a variety of providers and/or decision makers. PMID- 11276802 TI - Use of the ACCES model to predict the health economic impact of celecoxib in patients with osteoarthritis or rheumatoid arthritis in Norway. AB - A Norwegian customization of the Arthritis Cost Consequence Evaluation System (ACCES) pharmacoeconomic model was used to predict the economic and health impact of the introduction of celecoxib in Norway. The model predicts that use of celecoxib can be expected to result in a reduction in gastrointestinal events with concomitant annual net savings of at least Norwegian krone (NOK) 580 per osteoarthritis (OA) patient and NOK 514 per rheumatoid arthritis (RA) patient. In a cost-effectiveness analysis, celecoxib demonstrated economic dominance (i.e. improved health at reduced cost) compared with the currently available alternatives. In sensitivity analyses, the results of this model have been shown to be relatively robust, with celecoxib demonstrating economic dominance or favourable cost-effectiveness ratios in all analyses. Based on these data, it can be concluded that the introduction of celecoxib into the Norwegian non-steroidal anti-inflammatory drug market, and its use as a first-line agent, will provide societal benefits by improving health care at reduced cost in patients with OA and RA. PMID- 11276803 TI - The Swedish ACCES model: predicting the health economic impact of celecoxib in patients with osteoarthritis or rheumatoid arthritis. AB - The Arthritis Cost Consequence Evaluation System (ACCES) pharmacoeconomic model was used to evaluate the economic and health impact of the recent introduction of celecoxib for treatment of osteoarthritis (OA) and rheumatoid arthritis (RA) in Sweden. The model demonstrates that use of celecoxib can be expected to reduce the incidence of gastrointestinal adverse events, resource utilization and treatment costs. In a cost-effectiveness analysis, celecoxib demonstrated economic dominance (i.e. improved health at reduced cost) compared with the currently available alternatives for OA, and demonstrated economic dominance against a clinically relevant base-case scenario for RA. In sensitivity analyses, the results were shown to be relatively robust; celecoxib demonstrated economic dominance or favourable cost-effectiveness ratios in all analyses. Based on these data, it can be concluded that the use of celecoxib in Sweden will provide societal benefits by improving health care at reduced cost for patients with OA and RA. PMID- 11276804 TI - [New guideline for HIV classification and nomenclature: issues related to the origins of HIVs and biological correlates of HIV-1 subtypes]. PMID- 11276806 TI - [Rotaviruses]. PMID- 11276805 TI - [The physiological role of CK-II in the growth mechanism of HIV-1 as a host mediator]. PMID- 11276807 TI - [Caliciviruses]. PMID- 11276808 TI - [Virus vaccines]. PMID- 11276809 TI - [Astroviruses]. PMID- 11276810 TI - [Enteric adenoviruses: the biological background to viral infection]. PMID- 11276811 TI - [RNA-dependent RNA polymerases of plant plus-sense single-stranded RNA viruses]. PMID- 11276812 TI - [RNA recombination in plant RNA viruses]. PMID- 11276813 TI - [Gene silencing and viruses in plants]. PMID- 11276814 TI - [Plant expression vectors based on plus-sense single-stranded RNA viruses]. PMID- 11276815 TI - [Molecular virology of TT virus (TTV)]. PMID- 11276816 TI - [TTV-like viruses]. PMID- 11276817 TI - [Genome structure of TTV]. PMID- 11276818 TI - [The cellular receptor for measles virus: SLAM (CDw 150)]. PMID- 11276819 TI - [Wither rotavirus vaccines? A perspective from the WHO meeting]. PMID- 11276820 TI - [Early fragments of human tumor virus studies]. PMID- 11276821 TI - The Parkinson's experiment. PMID- 11276822 TI - Cut-rate AIDS. PMID- 11276823 TI - Who's feeling no pain? PMID- 11276824 TI - To tattle vs. to tell. PMID- 11276825 TI - Lessons of a bad heart. PMID- 11276826 TI - Rice-bran products: phytonutrients with potential applications in preventive and clinical medicine. AB - This paper reviews phytonutrients from rice bran that have shown promising disease-preventing and health-related benefits in experimental research studies. Candidate products studied and under investigation include: inositol and related compounds, inositol hexaphosphate (IP6 or phytate), rice oil, ferulic acid, gamma oryzanol, plant sterols, tocotrienols and RICEO, a new rice-bran-derived product. Diseases in which preventive and/or nutraceutical effects have been detected include: cancer, hyperlipidemia, fatty liver, hypercalciuria, kidney stones, and heart disease. In addition, rice-bran products may have potential applications as nutritional ingredients in the context of their utility in functional foods. PMID- 11276828 TI - Protection against drug- and chemical-induced multiorgan toxicity by a novel IH636 grape seed proanthocyanidin extract. AB - Grape seed proanthocyanidins have been demonstrated to exhibit a broad spectrum of pharmacological, therapeutic and chemoprotective properties. In our previous studies, IH636 grape seed proanthocyanidin extract (GSPE, commercially known as ActiVin) demonstrated excellent concentration- and dose-dependent free radical scavenging abilities in both in vitro and in vivo models and provided significantly better protection than vitamins C, E and beta-carotene. GSPE demonstrated significant cytotoxicity towards human breast, lung and gastric adenocarcinoma cells, while enhancing the growth and viability of normal human gastric mucosal cells and macrophage J774A.1 cells. In this study, the bioavailability and protective ability of GSPE were examined against acetaminophen-induced hepatoxicity, amiodarone-induced pulmonary toxicity, doxorubicin-induced cardiotoxicity, cadmium chloride-induced nephrotoxicity, dimethylnitrosamine-induced spleenotoxicity and O-ethyl-S,S-dipropyl phosphorodithioate (MOCAP)-induced neurotoxicity in mice. In each experiment, half of the test animals were orally fed GSPE for 7-10 days prior to drug/chemical exposure, while the other half received no GSPE. Parameters of analysis included changes in serum chemistry [alanine amino-transferase (ALT), blood urea nitrogen and creatine kinase], histopathology and integrity of genomic DNA. The results indicated that GSPE preexposure prior to the drugs/chemicals such as acetaminophen, amiodarone, doxorubicin, cadmium chloride or dimethylnitrosamine treatment, provided near complete protection in terms of serum chemistry changes (ALT, blood urea nitrogen and creatine kinase) and inhibition of both forms of cell death, e.g., apoptosis and necrosis. DNA damage in various tissues triggered by these agents was significantly reduced. Histopathological examination of the organs evaluated reflected similar patterns to those of the serum chemistry and DNA results. MOCAP exposure showed symptoms of severe neurotoxicity coupled with serum chemistry changes in the absence of any significant genomic change or brain pathology. GSPE afforded only partial protection in the brain tissue. These results suggest that GSPE exposure is bioavailable and provides significant multiorgan protection against drug- and chemical-induced toxic assaults. PMID- 11276827 TI - Carnitine protects mitochondria and removes toxic acyls from xenobiotics. AB - The carnitine system is altered by several xenobiotics (drugs and chemicals). These alterations are responsible for most toxic effects, which can be reverted or minimized by L-carnitine administration. Formation of nonmetabolizable acyl coenzyme A (CoA) is a typical step in the biotransformation of pivaloyl antibiotics, valproate and ifosfamide. The elevated levels of acylcarnitine occurring in human urine due to impaired metabolism of specific acyl CoA support the role of L-carnitine as an acceptor of specific, nonmetabolizable acyl CoA. The consequence of this process is a secondary carnitine deficiency. The formation of stable complexes with an essential component of mitochondrial membrane, cardiolipin, and the inhibition of myocardial specific isoform of carnitine-palmitoyl transferase are presumably the basis of adriamycin cardiotoxicity. L-carnitine interacts with cardiolipin, modifying membrane permeability and protecting the functions of the mitochondria. This mechanism can be proposed to explain the protective effects of L-carnitine against adriamycin cardiotoxicity, ammonium acetate and zidovudine-induced mitochondrial ultrastructural and functional alterations. Cisplatin, cephalosporin and carbapenem antibiotics inhibit carnitine reabsorption in renal tubules and cause proximal tubular damage. The study of peroxisomal producing agents belonging to largely different chemical classes showed that these agents caused carnitine system perturbations which may have the potential to be highly relevant biomarkers of exposure to the nongenotoxic peroxisomal proliferating agent class of hepatic tumorigens. PMID- 11276829 TI - Lumbar pedicle: surgical anatomic evaluation and relationships. AB - Although several clinical applications of transpedicular screw fixation in the lumbar spine have been documented for many years, few anatomic studies concerning the lumbar pedicle and adjacent neural structures have been published. The lumbar pedicle and its relationships to adjacent neural structures were investigated through an anatomic study. Our objective is to highlight important considerations in performing transpedicular screw fixation in the lumbar spine. Twenty cadavers were used for observation of the lumbar pedicle and its relations. After removal of whole posterior bony elements including spinous processes, laminae, lateral masses, and inferior and superior facets, the isthmus of the pedicle was exposed. Pedicle width and height (PW and PH), interpedicular distance (IPD), pedicle inferior nerve root distance (PIRD), pedicle-superior nerve root distance (PSRD), pedicle-dural sac distance (PDSD), root exit angle (REA), and nerve root diameter (NRD) were measured. The results indicated that the average distance from the lumbar pedicle to the adjacent nerve roots superiorly, inferiorly and to the dural sac medially at all levels ranged from 2.9 to 6.2 mm, 0.8 to 2.8 mm, and 0.9 to 2.1 mm, respectively. The mean PH and PW at L1-L5 ranged from 10.4 to 18.2 mm and 5.9 to 23.8 mm, respectively. The IPD gradually increased from L1 to L5. The mean REA increased consistently from 35 degrees to 39 degrees. The NRD was between 3.3 and 3.9 mm. Levels of significance were shown for the P < 0.05 and P < 0.01 levels. On the basis of this study, we can say that improper placement of the pedicle screw medially and inferiorly should be avoided. PMID- 11276830 TI - Three-dimensional kinematics of the lumbar spine during treadmill walking at different speeds. AB - The lumbar spine is of primary importance in gait and its development is influenced by the upright posture adopted in human locomotion. However, little is known about the kinematic behavior of the lumbar spine during walking. The aim of this study was to examine (1) lumbar spine kinematics during walking, (2) the effect of walking velocity on lumbar motion patterns and (3) the coupling characteristics of rotation and bending. In 22 volunteers aged 15-57 years, the three-dimensional displacements of T12 to the sacrum were sampled during elementary movements of the trunk and during walking on a treadmill at four walking velocities. A three-dimensional electrogoniometer (CA 6000 Spine Motion Analyzer) sampling at 100 Hz was used. We analyzed maximal primary and coupled motion ranges (ROM) and velocities in each plane. Lumbar ROM during walking did not exceed 40% of maximal active ROM. Transverse plane ROM and frontal and transverse velocities increased with walking velocity. Coupling of rotation and bending during walking was individually variable and dependent on walking velocity. Moreover, the smoothness of the bending-rotation path varied with walking velocity. A simplified envelope of lumbar coupling characteristics during walking is presented, and the existence of an individually variable walking speed that is characterized by a more harmonic lumbar contribution is hypothesized. PMID- 11276831 TI - Can a short spinal cord produce scoliosis? AB - Some patients with scoliosis have a relatively short vertebral canal. This poses the question of whether a short spinal cord may sometimes cause scoliosis. The present paper presents two observations that may support this concept. It presents a scoliosis model demonstrating what effect a short, unforgiving spinal cord might have on the spinal column. The model uses two flexible parallel tubes with the facility to tighten one. It demonstrates that a short, unforgiving spinal cord could produce the abnormal rotatory anatomy observed at the apex in scoliosis, with first lordosis, then lateral deviation and finally a rotation of the vertebral column, with the rotation occurring between the canal and the vertebral body, around the axis of the cord. The anatomy of the apical vertebra is described from two museum specimens, a computed tomography (CT) myelogram and seven magnetic resonance imaging (MRI) studies. The study confirms that the vertebral canal and the intervertebral foraminae retain their original orientation. The spinal cord is eccentric in the canal towards the concavity of the curve; the major component of rotation occurs anterior to the vertebral canal and the axis of this rotation seems to be at the site of the spinal cord. These observations do not establish that a short spinal cord will result in scoliosis, but the results are compatible with this hypothesis, and that impairment of spinal cord growth factors may sometimes be responsible for scoliosis. PMID- 11276832 TI - The influence of cancellous bone density on load sharing in human lumbar spine: a comparison between an intact and a surgically altered motion segment. AB - The aim of the current study is twofold: first, to compare load sharing in compression between an intact and a surgically repaired lumbar spine motion segment L3/4 using a biomechanically validated finite element approach; second, to analyse the influence of bone mineral density on load sharing. Six cadaveric human lumbar spine segments (three segments L2/3 and three segments L4/5) were taken from fresh human cadavers. The intact segments were tested under axial compression of 600 N, first without preload and then following instrumented stabilisation. These results were compared to a finite element model simulating the effect of identical force on the intact segments and the segments with constructs. The predictions of both the intact and the surgically altered finite element model were always within one standard deviation of the mean stiffness as analysed by the biomechanical study. Thus, the finite element model was used to analyse load sharing under compression in an intact and a surgically repaired human lumbar spine segment model, using a variety of E moduli for cancellous bone of the vertebral bodies. In both the intact and the surgically altered model, 89% of the applied load passed through the vertebral bodies and the disc if an E modulus of 25 MPa was used for cancellous bone density. Using 10 MPa- representing soft, osteoporotic bone--this percentage decreased, but it increased using 100 MPa in both the intact and the altered segment. Thus, it is concluded that reconstruction of both the disc and the posterior elements with the implants used in the study recreates the ability of the spine to act as a load-sharing construction in compression. The similarity in load sharing between normal and instrumented spines appears to depend on assumed bone density, and it may also depend on applied load and loading history. PMID- 11276833 TI - Biomechanical compression tests with a new implant for thoracolumbar vertebral body replacement. AB - The authors present an investigation into the biomechanical functioning of a new titanium implant for vertebral body replacement (Synex). Possible indications are fractures and/or dislocations with damage of the anterior column, posttraumatic kyphosis and tumors of the thoracolumbar spine. The construction must be supplemented by a stabilizing posterior or anterior implant. For best fit and contact with adjacent end-plates, Synex is distractable in situ. We performed comparative compression tests with Synex and MOSS ("Harms mesh cage") on human cadaveric specimens of intact vertebrae (L1). The aim of the study was to measure the compressive strength of the vertebral body end-plate in uniaxial loading via both implants to exclude collapse of Synex in vivo. Twelve human cadaveric specimens of intact vertebrae (L1) were divided into two identical groups (matched pairs) according to bone mineral density (BMD), determined using dual energy quantitative computed tomography (DE-QCT). The specimens were loaded with an axial compression force at a constant speed of 5 mm/min to failure, and the displacement was recorded with a continuous load-displacement curve. The mean ultimate compression force (Fmax) showed a tendency towards a higher reading for Synex: 3396 N versus 2719 N (non-significant). The displacement until Fmax was 2.9 mm in the Synex group, which was half as far as in the MOSS group (5.8 mm). The difference was significant (P < 0.001). The compression force was twice as high, and significantly (P < 0.05) higher with Synex at displacements of 1 mm, 1.5 mm and 2 mm. A significant (P < 0.001) correlation (R = 0.89) between Fmax and BMD was found. Synex was found to be at least comparable to MOSS concerning the compressive performance at the vertebral end-plate. A possible consequence of the significantly higher mean compression forces between 1 and 2 mm displacement might be decreased collapse of the implant into the vertebral body in vivo. PMID- 11276834 TI - CT study of craniovertebral rotation in whiplash injury. AB - The present study investigates the diagnostic value of rotatory computed tomography (CT) examinations in normal subjects and patients with whiplash associated disorders (WAD), with the aim of reproducing earlier findings of rotatory CT studies. Forty-seven WAD patients with persistent complaints after a rear-end collision (non-cranial contact acceleration/deceleration trauma) were enrolled in this study. To guarantee a maximally homogeneous study population, only WAD patients with a marked passive cervical retroflexion restriction were included. Transversal CT slices in left and right rotation were made for all cervical levels (the skull included). CT slices in neutral position were used to reconstruct partially depicted vertebrae. Absolute rotatory values were estimated according the method of Penning and Dvorak. For all levels the relative rotatory (RR) value was calculated by dividing absolute rotation values of a particular cervical level by the corresponding total cervical rotation. The measuring error was estimated by comparing the findings of two separately performed measuring procedures. Two age groups of WAD patients were formed. A younger group was matched for age with 26 normal healthy volunteers (the original data of an earlier study). The use of neutral CT slices for reconstruction of a partially depicted cervical vertebra resulted in a measurement error of 1.9 degrees at the level of C0/C1 (occiput/atlas) and 3.5 degrees at C1/C2. Suspected hypermobility as defined by Dvorak was rare in our WAD patients (6.4% C0/C1 and 10.6% C1/C2). RR values at C0/C1 were significantly larger in 79% of the WAD patients. Discriminant analysis of the RR values showed 80% correctly classified WAD patients. Only 11.5% of the normal subjects were classified as false-positive. Since no hypermobility was found at C1/C2, a traumatic lesion of the alar ligaments is less likely. It was concluded that the use of absolute rotation values in rotatory CT scan procedures has a low diagnostic value in WAD patients. Excessive RR values were only found at C0/C1. A traumatic lesion of the ligaments at C0/C1, which prevent vertical translation of the skull with regard to the atlas, is hypothesised. The results of the discriminant analysis of the RR values make this method applicable for the individual WAD patient in daily practice. PMID- 11276835 TI - The relation between initial symptoms and signs and the prognosis of whiplash. AB - Whiplash, a common injury following motor vehicle crashes, is associated with high costs and a prognosis that is variable and difficult to predict. We studied the profile of recovery from whiplash and assessed whether presenting signs and symptoms directly after the crash were predictive of whiplash prognosis. We formed a population-based incident cohort of all 2627 individuals who sustained a whiplash injury resulting from a motor vehicle crash in the province of Quebec, Canada, in 1987, and followed these patients for up to 7 years. The data on signs and symptoms were obtained from the medical charts kept by the universal automobile insurance plan (Societe de l'assurance automobile du Quebec), which covers all 7 million residents of the province, while data on the outcome--the recovery time from whiplash--was obtained from their databases. The median recovery time was 32 days, and 12% of subjects had still not recovered after 6 months. The signs and symptoms that were found to be independently associated with a slower recovery from whiplash, besides female gender and older age, are neck pain on palpation, muscle pain, pain or numbness radiating from the neck to arms, hands or shoulders, and headache. Together, these factors in older females (age 60) predicted a median recovery time of 262 days, compared with 17 days for younger males (age 20) who do not have this profile. In contrast, using a classification of injury severity previously proposed by the Quebec Whiplash Associated Disorders Task Force, the median recovery time varied from 17 to only 123 days. We conclude that whiplash patients presenting with several specific musculoskeletal and neurological signs and symptoms will have a longer recovery period. These patients can easily be identified and closely monitored and targeted for the evaluation of early intervention programmes aimed at managing whiplash patients with a poor prognosis. PMID- 11276836 TI - Posterior stabilization of the cervical spine with hooks and screws. A clinical evaluation of 26 patients with traumatic, degenerative or metastatic lesions, using a new implant system. AB - We operated on 26 patients with cervical spine disorders (13 with traumatic lesions, 3 with spinal stenosis and myelopathy, 1 with osteomyelitis and 9 with metastasis) with posterior stabilization. A new implant system (Cervi-Fix) based on rods, enabling a choice of either screw or laminar hook fixation in a free combination, was used. The system was evaluated for ease of use, for safety, regarding complications related to the system, and for efficacy, regarding loss of correction and signs of instability. The patients were followed for a mean period of 11 months, with ordinary and flexion/extension radiographs and clinical examination. No complications related to the implant system were observed. Loss of correction was observed in one patient. We found constructs with few vertebral fixation points, especially with screws, easy to handle, whereas multiple-claw constructs were time consuming. This implant system seems to be versatile, safe and efficient, but could be improved by the development of instruments for the insertion of the hooks. PMID- 11276837 TI - The effect of surgery and remodelling on spinal canal measurements after thoracolumbar burst fractures. AB - Bone fragments in the spinal canal after thoracolumbar spine injuries causing spinal canal narrowing is a frequent phenomenon. Efforts to remove such fragments are often considered. The purpose of the present study was to evaluate the effects of surgery on spinal canal dimensions, as well as the subsequent effect of natural remodelling, previously described by other authors. A base material of 157 patients operated consecutively for unstable thoracolumbar spine fractures at Sahlgrenska University Hospital in Gothenburg during the years 1980-1988 were evaluated, with a minimum of 5-years follow-up. Of these, 115 had suffered burst fractures. Usually the Harrington distraction rod system was employed. Patients underwent computed tomography (CT) preoperatively, postoperatively and at follow up. From digitized CT scans, cross-sectional area (CSA) and mid-sagittal diameter (MSD) of the spinal canal at the level of injury were determined. The results showed that the preoperative CSA of the spinal canal was reduced to 1.4 cm2 or 49% of normal, after injury. Postoperatively it was widened to 2.0 cm2 or 72% of normal. At the time of follow-up, the CSA had improved further, to 2.6 cm2 or 87%. The extent of widening by surgery depended on the extent of initial narrowing, but not on fragment removal. Remodelling was dependent on the amount of bone left after surgery. The study shows that canal enlargement during surgery is caused by indirect effects when the spine is distracted and put into lordosis. Remodelling will occur if there is residual narrowing. Acute intervention into the spinal canal, as well as subsequent surgery because of residual bone, should be avoided. PMID- 11276838 TI - The use of 'hybrid' allografts in the treatment of fractures of the thoracolumbar spine: first experience. AB - Harvesting autogenous bone grafts of the iliac crest carries complications and lengthens operative times. Allografts are preferred to avoid these problems. Fusion after using allogenic bone grafts has been well studied, by examining trabeculations and remodelling on anteroposterior and lateral radiographs. However, the question remains whether one can rely on radiographs alone to determine fusion. 'Hybrid' fresh-frozen allografts from the femur or tibia were used in 11 adult patients with a mean age of 56.4 years (range: 30-78 years) to stabilize the thoracolumbar spine after anterior decompression for trauma. In one case two adjacent levels were fractured, in another case two fractures occurred at different levels. Fresh-frozen allografts of the femur (in ten cases) and tibia (one case), filled with autogenous cancellous bone graft or pieces of rib, were used to reconstruct the anterior column of the spine. Stabilization was performed by means of a Kaneda device. Anteroposterior and lateral radiographs and, additionally, computed tomography (CT) examinations with reconstructions were used to study fusion. One patient died 1 month after surgery. At follow-up in ten patients, after a mean time of 30.2 months (range: 18-42 months), ten allografts showed a grade I fusion and one a grade III fusion. Additional data from the CT examination with reconstructions, however, showed cross trabeculations in all cases, and a partially united allograft in the patient with a grade III fusion. Cross-trabeculations between the allograft and vertebral body was observed at 6 months, with remodelling occurring at approximately 2 years. Mean loss of correction was minimal, at 3.6 degrees (range 0 degree-16 degrees). Fresh-frozen femoral or tibial allografts worked effectively to maintain correction after trauma when combined with anterior instrumentation. CT examinations with sagittal and coronal reconstructions were more effective for evaluation of fusion compared with anteroposterior and lateral radiographs. The high fusion rate and the low morbidity achieved using allografts in this way supports the exclusive use of allografts in the anterior thoracic and lumbar spine in the future. PMID- 11276839 TI - Surgical treatment of giant cell tumours of the thoracic and lumbar spine: report of nine patients. AB - Giant cell tumours involving vertebral bodies are still difficult to treat, though results are gradually improving. The object of this study was to assess the results of "complete excision", both of previously untreated giant cell tumours and of recurrences, and to consider the possible effects of any tumour contamination during operation. Nine consecutive patients with giant cell tumours of the thoracic and lumbar spine were treated surgically between 1986 and 1995. Four of these patients were referred with recurrent tumours. All operations aimed at complete resection of the tumour, where possible an en-bloc approach was used. The spines were reconstructed with autografts and instrumentation. All patients were regularly reviewed as part of an on-going study. Following the five operations for previously untreated tumours ("primary" operations), there were no local recurrences, but one patient died of pulmonary metastases. One of the four patients operated upon for a recurrence developed a further recurrence, which was excised 2 1/2 years ago. It would seem that giant cell tumours of the thoracic and lumbar spine, including recurrences, should be treated by complete excision. The en-bloc approach is the safest technique. Where an intralesional component is unavoidable, total removal of the (pseudo)capsule should be ensured by preliminary extralesional dissection. Any tumour spill should be meticulously removed. The use of frozen sections to check resection margins is advisable. PMID- 11276840 TI - Direct repair of defect in lumbar spondylolysis and mild isthmic spondylolisthesis by bone grafting, with or without facet joint fusion. AB - Forty-six patients with lumbar spondylolysis and mild isthmic spondylolisthesis were managed with direct repair of the defect with or without facet joint fusion in the affected segment. There were 24 males and 22 females, ranging in age from 15 to 56 years (average, 38.2 years). These patients had experienced clinical symptoms due to spondylolysis for between 4 months and 20 years (average, 5.3 years). Of 46 patients, 28 had no spondylolisthesis, 11 had Meyerding grade I vertebral slippage and 7 had grade II. Direct repair of 98 defects was performed on these patients. Twenty-six patients, in whom the disc adjacent to the defect was determined as degenerative by magnetic resonance imaging (MRI), simultaneously underwent facet joint fusion; 17 in one segment and 9 in two segments. The average period of follow-up was 50 months (24-92 months). Ninety four defects achieved bony healing. As a result, 28 patients were graded as having an excellent outcome, 15 good, and 3 fair. Bone grafting in the defects achieves union between the loose lamina and the anterior element of lumbar vertebrae, and reconstructs the anatomic structure and physiologic functions of the lumbar vertebrae. There was no significant difference in outcome between the spondylolytic/spondylolisthetic patients with non-degenerative disc, who were treated with direct repair of defect only, and those with degenerative disc, who additionally underwent a fusion procedure (P > 0.05). The present series demonstrates a satisfactory result and a high rate of bony healing of the pars defect by this operative procedure in patients with lumbar spondylolysis and mild isthmic spondylolisthesis. Preoperative assessment of the disc degeneration with MRI is of great assistance in making the protocol choice of whether to opt for fusion. PMID- 11276841 TI - Occlusive arterial diseases of the upper and lower extremities found in workers occupationally exposed to vibrating tools. AB - Hand-arm vibration syndrome (HAVS) is primarily a disorder of the fingers and hands. However, in some cases, vibration-exposed workers are observed to have also episodic blanching of the hands and feet. In latter cases, arteriographies of both the upper and lower extremities are necessary to diagnose the background arterial disorders. In this study, eight HAVS subjects with such disorders were examined by arteriography for differential diagnosis in cases of workers' accident compensation. In three HAVS cases with thromboangiitis obliterans, the arteriographic examination revealed obstructive changes in the palm and forearm as well as three below-knee lesions in the lower extremities. In five HAVS cases with arteriosclerosis obliterans, obstruction kinking or coiling, stenosis and/or tapering-off of the proper digital arteries were observed together with two below knee lesions and three high lesions in the lower extremities. From the viewpoint of occupational health, palpation of superficial arteries of both the upper and lower extremities should be routinely performed during both pre-placement and periodic medical examinations for workers exposed to vibrating tools for early detection and/or prevention of any worsening of the background disorders. PMID- 11276842 TI - Experimental studies of vibratory trauma of Corti's organ. I: Electrophysiological measurements. AB - In the present-day environment, vibration concomitant with noise is most frequently observed, but even though it is regarded as only a weak, additional traumatic factor affecting the organ of hearing. This opinion is contrary to a number of reports on the damage of the hearing organ caused by vibration in workers of various branches of industry. As experiments on humans are rather difficult, the harmful effect of vibration is usually examined on laboratory animals. In the majority of studies dealing with this problem the presence of noise (purposely or casually) was found in most cases. The aim of this study was to determine the effect of isolated long-term whole-body vibration and vertical sinusoidal shaking (10 Hz frequency, 5 mm amplitude, and 2 g acceleration) on Corti's organ. The study was carried out on young guinea pigs of both sexes. Eighty four animals (30 control and 54 experimental) with Preyer's reflex and without otoscopically detectable changes were used. A group of 18 animals was subjected to vibration in noiseless shaking apparatus for 30, 90 and 180 days. After a one-month rest, cochlear microphonics were performed under urethane anaesthesia. Our modification of the phase-sensitive detection method was used. Cochlear microphonics at frequencies of 260 Hz, 500 Hz, 1 kHz and 2 kHz was recorded from the apex of the cochlea and for 4 kHz and 8 kHz from the region of the round window. As the cochlear microphonics values showed significant individual differences, all experimental samples were examined by means of non parametric tests. The outcome of the study demonstrated a gradual but considerable cochlear microphonics voltage decrease in the range to 2 kHz. This result pointed to the generation of vibration-induced damage in outer hair cells of the fourth and third turnings of the cochlea in the guinea pigs under study. PMID- 11276843 TI - Relationship between sense of coherence and post-traumatic stress disorder symptoms among firefighters. AB - According to the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV), published by the American Psychiatric Association, the post-traumatic stress disorder (PTSD) is diagnosed when a person: (a) is exposed to a traumatic event that is well outside the range of usual human experience accompanied by intense fear or horror; (b) reexperiences the event in his/her thoughts, dreams and daily life; (c) avoids the stimuli associated with the trauma and numbs his/her emotions; (d) demonstrates symptoms of increased arousal; and (e) manifests these disturbances for a longer period than one month. Since the 1980s, it has been pointed out that PTSD may occur not only among survivors of severe traumatic events but also among those who have rescued the victims of those events. Members of fire brigades constitute a large occupational group exposed to traumatic experiences. The aim of our study was to find an answer to the question of what are the relationships between the level of PTSD symptoms and the sense of coherence (and its three dimensions). In all, 464 firemen were interviewed. PTSD Interview developed by Watson et al. was used to assess the level of PTSD symptoms and the presence]absence of PTSD. The higher level of PTSD symptoms was associated with the lower level of the sense of coherence. A small group (3.9%) of subjects who experienced traumatic events met DSM-IV diagnostic criteria for PTSD. The sense of coherence of these people was significantly lower than that of others. PMID- 11276844 TI - Studies on dermal, ocular and respiratory effects of 4-ethyltoluene in experimental animals. AB - The toxicity of 4-ethyltoluene to experimental animals was studied after single and repeated exposures. It was found that 4-ethyltoluene can be classified as a very mild skin and eye irritant. Sensory respiratory irritation of 4-ethyltoluene was studied in Balb/C male mice using the plethysmographic method. The concentration at which the respiratory rate decreased to 50% (RD50 value) was determined to be 4216 mg/m3 (2795-5850 mg/m3 for 95% confidence interval). To study repeated-dose inhalation toxicity, male and female outbred Wistar rats were exposed in a dynamic inhalation chamber to 4-ethyltoluene vapours at concentrations of 477 or 2337 mg/m3, 6 h/day, 5 days/week for 4 weeks (20 exposure days). No significant changes were observed in food consumption and body weight gain. Statistically significant, concentration-dependent changes in the number of total cells, as well as of macrophages, polymorphonuclear leucocytes and lymphocytes were found in bronchoalveolar lavage. In the fluid of bronchoalveolar lavage, a significant, concentration-related increase was noted in total protein and mucoproteins and the activity of beta-glucuronidase, gamma glutamyl transferase, lactate dehydrogenase and acid phosphatase. Histopathology revealed an increased rate of bronchitis and pneumonia and perivascular lymphoid infiltrations in rats exposed to 2337 mg/m3 of 4-ethyltoluene. PMID- 11276845 TI - Effects of repeated exposure to toluene or amphetamine on locomotor activity in rats. AB - This study was performed to find out whether repeated exposure to toluene might lead to behavioural sensitization estimated on the basis of spontaneous locomotor activity of rats in an open field. As a reference compound amphetamine was used. Toluene at a dose of 740 mg/kg was administered orally, amphetamine at a dose of 1 mg/kg was administered intraperitoneally for 7 consecutive days. Fourteen days after the last administration the amphetamine or toluene challenge was made. Spontaneous locomotor activity was assessed on the first and seventh day of the exposure and on the day of toluene or amphetamine challenge. Statistics employed a two-way analysis of variance (ANOVA). The results showed that oral administration of toluene at a dose of 740 mg/kg may induce behavioural sensitization. The response to toluene was weaker than that to reference chemical -amphetamine. The results did not indicate the cross-sensitization between toluene and amphetamine. PMID- 11276846 TI - Disposition and metabolism of 1,4-dimethylnaphthalene in rat. AB - The distribution, excretion and metabolism of 1,4-dimethylnaphthalene following i.p. administration of a single dose of 28 mg/kg to rats, was investigated using radiotracer [3H] and gas chromatography-mass spectrometry technique (GC-MS). After 72 h, about 97% of the given dose was excreted in urine and faeces. Maximum level of tritium in plasma was observed during the 4th h. after the compound administration. In organs and tissues, the highest concentration during the first hours after administration was detected in the fat, liver, spleen and kidneys. Then gradual decline in tritium was noticed in all examined tissues. In urine the following substances were identified and quantified by GC peak areas: unchanged 1,4-dimethylnaphthalene, 1-hydroxymethyl-4-methylnaphthalene, 4-methyl-1 naphthoic acid, 1,4-dimethylnaphthol, 4-methyl-1-naphthoic aldehyde and 1,4 dimethyl-methylthionaphthalene. PMID- 11276847 TI - Apoptotic effect of cyanobacterial blooms collected from Polish water reservoirs. AB - Recently in many countries, including Poland, the problem of toxicity of cyanobacterial blooms has been of great importance. In many cases it is connected with the increase of microcystins (MCYSTs) concentration in fresh water. This problem is caused by excessive eutrophication of drinking and recreational water bodies. In humans, the most frequent symptoms of the MCYST effect are: cutaneous rash, fever, vomiting, diarrhoea, gastroenteritis and acute damage of the liver. The aim of this work was to estimate apoptotic effects of five different cyanobacterial hepatotoxic extracts containing MC-LR and other variants of MCYSTs (MC-RR, MC-YR, and MC-WR). These effects were analysed in rat hepatocytes- primary target of cyanobacterial hepatotoxins. Morphological changes in hepatocytes were examined by means of fluorescence and differential interference contrast microscopy with the DNA-specific dye, Hoechst 33342. The hepatocytes were treated with each cyanobacterial extracts containing MC-LR in the range between 100 nM-2000 nM for 30 min, 60 min and 120 min. The first characteristic apoptotic changes: shrinking and budding of cells were seen after 30 min, MC-LR = 100 nM. During the next 30 min the percentage of apoptotic cells increased by over 50%, MC-LR at concentrations ranging from 100 to 250 nM (the value dependent on a bloom sample). Highly condensed chromatin and apoptotic bodies were observed in 85-90% of hepatocytes after 120 min of treatment with MC-LR in concentration of 1000 nM. The apoptotic changes in rat hepatocytes confirm the high cytotoxic potential of cyanobacterial bloom samples collected during different months and years from reservoirs of drinking and recreational water in central Poland. PMID- 11276848 TI - Biological monitoring in waste landfills studies. AB - The assessment of exposure to chemicals from waste landfills and their health impact is a crucial, yet extremely difficult task in waste landfill studies. Use of biological markers, both of exposure and of health effects, may be very helpful if adequately chosen and precisely applied. They must be validated before application in risk assessment studies by establishing the relationship between the biomarker, exposure and the health outcome. Biomarkers suitable for the application in the waste landfill studies are reviewed in this paper. PMID- 11276849 TI - Alpha spectrometry for measuring activity of glass-embedded 210PO isotope. AB - Measurements of exposure to radon are performed using numerous research methods which register either temporary or periodic radon concentrations. At the end of the 1990s reports on the possibility of defining past exposure to radon on the basis of measurement of the contents of 210Pb embedded in surface layer of glass were published. The registration of alpha particles emitted with glass was made using the spectrometric system. Use of the system for alpha spectroscopy leads finally to a significant lowering of the bottom threshold of detection and limits the influence of the negative background on the examined glass. PMID- 11276850 TI - Cellular sources of oxidants in the lung. AB - Reactive oxygen species (ROS) are important causative factors of many lung diseases, for instance pulmonary emphysema, adult respiratory distress syndrome (ARDS), lung fibrosis or rejection of transplanted lung. Moreover, recent data show that these molecules also play some physiological roles including signal transduction which regulates such important functions as cell growth and apoptosis. This review describes various cellular sources of ROS in the lung. Special attention was paid to neutrophils, eosinophils, alveolar macrophages and cells of alveolar epithelium. A possible contribution of ROS released by these cells to various lung diseases is discussed. PMID- 11276851 TI - Norma Rae, R.N. AB - Unions are becoming more attractive to nurses who feel increasingly unappreciated and disenfranchised from the decisionmaking process. But unions can cause big headaches for management and the board. The best way to keep unions off your doorstep is to listen to what your nurses want. PMID- 11276852 TI - Best financial practices for trustees. AB - State attorneys general as well as creditors and the IRS are putting more pressure on not-for-profit boards to fulfill their fiduciary responsibilities. Here's a guide to help you avoid liability and ensure a better financial outcome for your organization. PMID- 11276853 TI - CEOs: no shame in self-doubt. PMID- 11276854 TI - Patient safety and the Joint Commission: the latest news. PMID- 11276855 TI - Determining physician need--a priority for trustees. PMID- 11276856 TI - No margin, big mission. Partnerships are key to Crozer-Keystone's community health commitment. AB - Crozer-Keystone Health System in Pennsylvania is committed to providing community health services, even though it has a 1 to 1.5 percent operating margin. How does the system do it? By targeting priorities and engaging in partnerships. PMID- 11276857 TI - Preserving your community's access to essential health care. AB - Reductions in the rates of Medicare and Medicaid payment over the last few years have put tremendous stress on the financial well-being of many hospitals. Trustees are faced with the dual challenge of trying to ensure financial viability of their organizations while at the same time working to maintain access to essential health services for their communities. This is a daunting challenge. PMID- 11276858 TI - Informing juvenile justice policy: directions for behavioral science research. AB - Recent policy initiatives threaten to reduce the rehabilitative mission of the juvenile court or eliminate the court entirely. This article lays out a framework for an empirical assessment of these developments. It first evaluates the available and potential empirical support for three hypotheses about juveniles that might justify maintaining a separate, rehabilitation-oriented juvenile justice system: the hypotheses that, compared to adults, juveniles are more treatable, less culpable, and less deterrable. On the assumption that the continued existence of a rehabilitation-oriented juvenile court can be justified, it then provides suggestions as to how existing intervention strategies for juveniles could benefit from research attention to several substantive and methodological issues. These include refining outcome criteria and sampling strategies, matching offender and program characteristics, reexamining intervention efficacy, and focusing on decision makers and resource allocations. PMID- 11276859 TI - The effects of joint legal custody on mothers, fathers, and children controlling for factors that predispose a sole maternal versus joint legal award. AB - Findings from comparisons of joint and sole custody families that do not control for predivorce differences in demographic and family process variables (factors that may predispose families to choose or be awarded joint custody) are of limited generalizability, since obtained group differences may be attributable to predisposing (self-selection) factors, custody, or both. This study compared a random sample of 254 recently separated, not-yet-divorced families on 71 predivorce variables that might plausibly differentiate between families awarded joint legal versus sole maternal custody. Twenty such factors were identified and controlled for in subsequent comparisons of 52 sole maternal and 26 joint legal custody families 2 years postdivorce. Families with joint custody had more frequent father-child visitation, lower maternal satisfaction with custody arrangements, more rapid maternal repartnering, and fewer child adjustment problems (net of predivorce selection factors). Moreover, these effects did not appear to be moderated by level of predecree parental conflict. No association between custody and fathers' compliance with child support orders was obtained. PMID- 11276860 TI - Children's law: toward a new realism. AB - The realist approach that has dominated American jurisprudence has long had especially great acceptance in regard to children's issues. Ironically, however, decision making on such topics has seemed to be particularly unlikely to be informed by careful assessments of social reality. Symbolism has prevailed over pragmatism. Psycholegal research on children's issues has also often been misdirected. Application of the Convention on the Rights of the Child may point the way to more psychologically minded children's law. PMID- 11276861 TI - Criminal decision making: the development of adolescent judgment, criminal responsibility, and culpability. AB - Theories of judgment in decision making hypothesize that throughout adolescence, judgment is impaired because the development of several psychosocial factors that are presumed to influence decision making lags behind the development of the cognitive capacities that are required to make mature decisions. This study uses an innovative video technique to examine the role of several psychosocial factors -temporal perspective, peer influence, and risk perception--in adolescent criminal decision making. Results based on data collected from 56 adolescents between the ages of 13 and 18 years revealed that detained youth were more likely to think of future-oriented consequences of engaging in the depicted delinquent act and less likely to anticipate pressure from their friends than nondetained youth. Examination of the developmental functions of the psychosocial factors indicates age-based differences on standardized measures of temporal perspective and resistance to peer influence and on measures of the role of risk perception in criminal decision making. Assessments of criminal responsibility and culpability were predicted by age and ethnicity. Implications for punishment in the juvenile justice system are discussed. PMID- 11276862 TI - Interviewing children about real and fictitious events: revisiting the narrative elaboration procedure. AB - Elementary school children participated in a staged event. Two weeks later they were randomly assigned to three interview conditions: (a) a streamlined version of the Narrative Elaboration (NE) procedure involving training in the use of reminder cue cards, (b) exposure to reminder cue cards without training in their use (cue card control group), and (c) a standard interview including no NE training or exposure to reminder cue cards (standard-interview control group). Children in each interview condition were questioned about the staged event and a fictitious event to determine whether children trained in the streamlined NE procedure would provide more information about a staged event than would children in the two control groups and whether the NE interview would result in increased reporting of false information when questioned about a fictitious event. Results indicated that children questioned with the NE interview reported a greater amount of accurate, but not a greater amount of inaccurate, information during cue-card presentation for the staged event than did the cue-card control group. Analyses further indicated that the NE-interview group did not report significantly more false information about the fictitious event than did children in the two control groups. Large standard deviations for the NE-interview children's cue-card recall indicate that the streamlined NE procedure was useful for many children in reporting the staged event, but may have contributed to a small number of children providing false information for the fictitious event. Further research is being conducted to determine which children may be more likely to be helped and which children may be more likely to provide false information regarding a fictitious event. PMID- 11276863 TI - Reducing maltreated children's reluctance to answer hypothetical oath-taking competency questions. AB - Before allowing child witnesses to testify, courts routinely require children to describe what would happen to them if they lied. However, young children often refuse to reason hypothetically if they view the premises as implausible or undesirable, and might be more willing to discuss the consequences of lying if they are asked about another child rather than themselves. On the other hand, children might view themselves as invulnerable to punishment, and therefore believe that whereas other children will be punished for lying, they will not be. In this study, 64 maltreated 5- and 6-year-old children were asked to describe the consequences of lying to three professionals (a judge, a social worker, and a doctor). Participants in the "self" condition were asked what would happen to them if they lied, whereas participants in the "other" condition were asked to describe what would happen to a story child if he or she lied. Asking children about "other" children increased responsiveness, and did not reveal perceptions of invulnerability. The results suggest that young children's understanding that they will be punished for lying may make them reluctant to discuss the consequences of lying, leading to underestimation of their oath-taking competency. PMID- 11276864 TI - Clinical assessment of parents in child protection cases: an empirical analysis. AB - We investigated the content and legal relevance of clinical evaluations of parents conducted in child abuse and neglect cases. The sample consisted of 190 mental health evaluation reports, randomly selected from major providers, that had been completed on parents involved in a large, urban juvenile court system. We coded evaluations on 170 objective and qualitative characteristics in order to assess for criteria recommended in the forensic literature. We compared evaluations across groups categorized by type (e.g., psychological, psychiatric, bonding/parenting, substance abuse) and where the assessments were performed (outside or inside the court). We found numerous substantive failures to meet those criteria for forensic relevance. Evaluations of parents typically were completed in a single session, rarely included a home visit, used few if any sources of information other than the parent, often cited no previous written reports, rarely used behavioral methods, stated purposes in general rather than specific terms, emphasized weaknesses over strengths in reporting results, and often neglected to describe the parent's caregiving qualities or the child's relationship with the parent. Some relevant differences were evident across assessment groups, pointing to examples of more thorough, parenting-specific evaluation practices. We recommend ways to improve current practices in forensic parenting assessment. PMID- 11276865 TI - Cell biology of gingival wound healing. PMID- 11276866 TI - Role of bacterial proteinases in matrix destruction and modulation of host responses. AB - Recently accumulated large bodies of evidence have strongly implicated proteolytic enzymes released by subgingival plaque bacteria in the pathogenicity of periodontal disease. With regard to proteolytic power, however, the contribution from different microbial species considered as periodontal pathogens is not equal. Two of these bacteria, P. gingivalis and T. denticola, have developed an elaborate proteolytic systems composed of several surface-located or secreted enzymes, which apparently serve a role to provide bacteria with nutrients in the form of small peptides and amino acids. Of these two species, proteinases of P. gingivalis are the most intensively studied, and during the last decade an impressive array of information has been accumulated with respect to the biochemical characterization of purified proteinases and structure of the genes encoding them, the regulation of expression and the effects of these enzymes on host systems. In addition, studies on proteinase-deficient isogenic mutants has shed light on both their housekeeping functions and potential role(s) in the pathogenicity of periodontitis. Among several proteinases produced by P. gingivalis, the cysteine proteinases, referred to as gingipains, are clearly in the spotlight. They are the subject of several recent reviews and generally considered as the major virulence factors of this periodontal pathogen (59, 105, 139, 182, 183, 186, 281, 284, 286, 289). Gingipains seem to be key players in subverting host defense systems with, significantly, the complement and neutrophils being the main target. In addition, through uncontrolled activation of kallikrein/kinin pathway and coagulation cascade they contribute to local generation of bradykinin and thrombin, two synergistically working pro inflammatory reagents with a strongly, although indirectly, stimulatory effect on bone resorption. Furthermore, the ability to interact with the cytokine networking systems has the potential to dysregulate the local inflammatory reaction. Finally, gingipains have a strong effect on mechanisms controlling host matrix metalloproteinase activity at the level of gene expression and zymogen activation (Fig. 10). Collectively, at the periodontal lesion site, the non restrained action of gingipains, supported by other proteinases locally produced by subgingival plaque bacteria, would dysregulate most mechanisms controlling inflammatory reaction. Although successful in limiting infection to the periodontium, the ultimate effect of uncontrolled inflammatory processes would be the destruction of periodontal connective tissue, certainly the hallmark of periodontitis. PMID- 11276867 TI - Connective tissue elements as diagnostic aids in periodontology. PMID- 11276868 TI - Regulators of tissue destruction and homeostasis as diagnostic aids in periodontology. PMID- 11276869 TI - Modulation of matrix metalloproteinase activities in periodontitis as a treatment strategy. PMID- 11276870 TI - Modulation of host inflammatory mediators as a treatment strategy for periodontal diseases. AB - Studies ranging from preclinical animal models to human clinical trials support the basic hypothesis that the inhibition of local arachidonic acid metabolites with nonsteroidal anti-inflammatory drugs slows periodontal disease progression. Data on modulation of other host mediators such as cytokines and NO remain restricted to laboratory or preclinical investigations, but appear promising. It is unlikely that such agents, following regulatory approval, will be used ubiquitously in patient populations, but rather may be targeted for at-risk patients. In the emerging field of periodontal medicine, patient cohorts are currently being identified with genetically based inflammatory mediator hyper responses (48, 49, 68, 115). Such cohorts who respond to the endotoxin challenge posed by periodontitis with a robust release of cytokines or prostaglandins may benefit most in terms of slowing periodontitis progression and potentially improving systemic susceptibility (3, 67). PMID- 11276872 TI - Molecular and cell biology of the gingiva. PMID- 11276871 TI - Tissue engineering: a new paradigm for periodontal regeneration based on molecular and cell biology. PMID- 11276873 TI - Role of physical forces in regulating the form and function of the periodontal ligament. PMID- 11276874 TI - Connective tissues of the periodontium--preface. PMID- 11276875 TI - Molecular and cell biology of cementum. PMID- 11276876 TI - Development and general structure of the periodontium. PMID- 11276877 TI - Molecular and cellular biology of alveolar bone. PMID- 11276878 TI - Saving the life of a practice after the death of the dentist. PMID- 11276879 TI - ADA extends warm welcome for dentists coming "home". PMID- 11276880 TI - House suspends PR campaign. Dues drop for 2001. PMID- 11276881 TI - When word of mouth comes back to haunt you. PMID- 11276882 TI - Teens on the Net. PMID- 11276883 TI - Your business. Breaking down bonds. PMID- 11276884 TI - In other words. What you see is what you get. PMID- 11276886 TI - The AMSA block: local anesthesia without collateral numbness. PMID- 11276887 TI - The mouse that roars. PMID- 11276889 TI - Starting out. Are you ethical? PMID- 11276888 TI - Disciplining dentists. State regulatory agency works to protect the public. PMID- 11276890 TI - Media matters. Who do you trust? What do you say? PMID- 11276892 TI - In other words. What stage are you in? PMID- 11276891 TI - Your business. Year-end tax planning strategies to consider now. PMID- 11276893 TI - Managing parents when treating their kids. PMID- 11276894 TI - 1980: a decade of change dawns. PMID- 11276895 TI - Retinal translocation: rationale and results. PMID- 11276896 TI - Mucosal immune system: close encounter in the uncharted world of immunology. PMID- 11276897 TI - Advancements in extended-wear contact lenses. PMID- 11276898 TI - The nature of intelligence. Introduction. PMID- 11276899 TI - g and the one-many problem: is one enough? AB - No one believes that g is the only construct needed to describe individual differences in intelligence. Many believe that g is a dispensable construct. Others object to what they construe as its hegemonic position in the domain of intelligence. In this paper I defend the hegemonic status of g by a brief consideration of diverse criticisms of the g construct. I argue that g is a heritable component of intelligence that accounts for approximately 50% of the covariance among diverse measures of intelligence. It derives from a core information processing ability and it influences a diverse set of social outcomes. The covariances between g and information processing ability and social outcomes are, in large measure, attributable to common genetic influences. PMID- 11276900 TI - General intelligence and the definition of phenotypes. AB - From Spearman's famous 1904 paper to Carroll's recent book on factor analytic results from a multitude of studies, there has been one consistent conclusion: 'g', or general intelligence, is the factor that defines the phenotype for intellectual functioning. It is no overstatement to say that g is undoubtedly the most important psychological construct discovered in this century. It predicts more and is implicated in a wider range of behaviour than any other psychological construct. The empirical support for g is extensive and overwhelming. It would seem that g is the perfect phenotypic definition of intelligence. I argue that it is not the perfect phenotype. If we are to understand intelligence, we need to define a new, more elaborate definition of intelligence taking g as the starting place. It must be remembered that g is a statistical abstraction. Current formulations of g are largely silent about the composition of g. I argue that g is actually made of further separable basic cognitive processes and does not represent a single underlying entity. These basic cognitive processes are integrated into a complex system in the brain that makes them difficult to identify. None the less, until these basic processes are identified and related to brain function there are a number of findings that cannot be explained and this will inhibit scientific progress. PMID- 11276901 TI - Is there a g factor for fitness? AB - Biological fitness is a directly observable quantity with well-known causal components, in contrast to the latent 'g' factor of psychometrics. The study of the causes of variation in fitness should therefore be simpler than the study of variation in mental abilities, but a paucity of data has kept the nature of genetic variation in fitness obscure. We can define an 'f' factor as variation creating positive correlations among components of fitness. There is little doubt that such f factor exist. Perturbations of populations such as mutation or environmental change create such patterns of positive correlation. However, natural selection will tend to minimize variation in any f factor, so it is much less clear whether f causes quantitatively substantial genetic variation within populations. Experimental data are consistent with variation in an f factor within some natural populations. As predicted, f is less important in populations where natural selection has had more opportunity to reshape the correlation matrix. Although one can incorporate variation in g into a study of variation in human fitness, the pace of change in our environment suggests that the results would neither reflect the conditions under which g evolved nor predict future evolutionary changes in g. PMID- 11276902 TI - How can psychological adaptations be heritable? AB - By Fisher's fundamental theorem, selection depletes additive genetic variation. However, moderate heritabilities are invariably obtained for psychological traits, even those that have been under intense selection. Examples include sociosexuality (interest in emotionally uncommitted sex), schizophrenia and sexual orientation, which have all been subject to strong sexual selection. A number of factors can help maintain (or at least slow depletion of) genetic variation. These include antagonistic pleiotropy; geographic or temporal variability in optimal phenotypes (and hence genotypes); mutational pressure (especially in the context of parasite resistance dynamics); and existence of heritable strategic variation or morphs. I discuss the likelihood that these factors maintain heritable variation for intelligence. I then review some evolutionary hypotheses regarding variation in some specific psychological traits. PMID- 11276903 TI - Social complexity and social intelligence. AB - When we talk of the 'nature of intelligence', or any other attribute, we may be referring to its essential structure, or to its place in nature, particularly the function it has evolved to serve. Here I examine both, from the perspective of the evolution of intelligence in primates. Over the last 20 years, the Social (or 'Machiavellian') Intelligence Hypothesis has gained empirical support. Its core claim is that the intelligence of primates is primarily an adaptation to the special complexities of primate social life. In addition to this hypothesis about the function of intellect, a secondary claim is that the very structure of intelligence has been moulded to be 'social' in character, an idea that presents a challenge to orthodox views of intelligence as a general-purpose capacity. I shall outline the principal components of social intelligence and the environment of social complexity it engages with. This raises the question of whether domain specificity is an appropriate characterization of social intelligence and its subcomponents, like theory of mind. As a counter-argument to such specificity I consider the hypothesis that great apes exhibit a cluster of advanced cognitive abilities that rest on a shared capacity for second-order mental representation. PMID- 11276904 TI - IQ gains, WISC subtests and fluid g: g theory and the relevance of Spearman's hypothesis to race. AB - IQ gains over time were calculated for each WISC (Wechsler Intelligence Scale for Children) subtest and the subtests ranked by size of gain. Verbal similarities led at 20 points per generation--larger than gains on Raven's Progressive Matrices. Similarities measures on-the-spot problem-solving (something akin to fluid g); verbal subtests that do not measure this show low rates of gain. WISC subtests were also ranked by their correlations with Raven's, the latter being used as a marker for fluid g. The r between the two hierarchies was calculated to approximate a correlation between IQ gains and fluid g. The result of 0.50 contrasts with the negative correlation between IQ gains and the g generated by factor analysing the WISC battery itself, which is generally viewed as predominately a crystallized g. In sum, it appears that human groups can make massive fluid g gains in a period too short to accommodate radical change in the speed and efficiency of neural processes. Moreover, once gains in intelligent behaviour over historical time are seen to be independent of brain physiology, does g really provide a criterion for assessing their significance? Finally, not only a measure of fluid g (which is highly heritable) but also inbreeding depression are shown to be correlated with IQ gains--gains overwhelmingly environmental in origin. Therefore, correlations between such genetically influenced factors and the size of the black/white IQ gap do not show that the gap has a genetic component. PMID- 11276905 TI - Mutation, selection and the heritability of complex traits. AB - It has been suggested that complex traits, like intelligence, have low heritabilities. This hypothesis stems from the idea that strong selection for higher intelligence has led to the fixation of genetic variation for this trait. The same hypothesis has been framed for complex sexual ornaments used in courtship display. These traits are also subject to directional selection, in this case caused by sexual selection. However, contrary to the hypothesis, comparative data shows that sexual ornaments have higher additive genetic variation than similar non-sexual traits. It appears that the number of variable genes and the effect per genetic locus have increased for sexual ornaments. Theory suggests this is due to selection for extreme phenotypes resulting in condition-dependent expression of sexual traits. Experimental work on the stalk eyed fly, Cyrtodiopsis dalmanni, confirms that the male sexual trait (exaggerated eyespan) is more sensitive to environmental conditions than other non-sexual traits (wing dimensions and female eyespan). This environmental sensitivity has a genetic basis and environmental stress enhances genetic differences. It is likely that genetic variation in intelligence is maintained in a similar fashion. PMID- 11276906 TI - The quantitative and molecular genetics of human intelligence. AB - General cognitive ability or g as measured by IQ tests has been the most intensively studied trait in human behaviour genetics. Although there has been much debate about how best to describe and measure intelligence, this is addressed elsewhere in this volume and here I will assume that there is utility in the concept of general intelligence and review the data showing that, whatever it is, g is familial and influenced by genes. Next, I will briefly describe some of the main quantitative genetics models that have been used in trying to tease out the genetic and environmental sources of variation. Finally, I will outline current molecular genetic research aiming to locate and identify genes influencing human intelligence and try to predict the directions in which such research will lead. PMID- 11276907 TI - Sexual selection for indicators of intelligence. AB - Many traits in many species have evolved through sexual selection specifically to function as 'fitness indicators' that reveal good genes and good health. Sexually selected fitness indicators typically show (1) higher coefficients of phenotypic and genetic variation than survival traits, (2) at least moderate genetic heritabilities and (3) positive correlations with many aspects of an animal's general condition, including body size, body symmetry, parasite resistance, longevity and freedom from deleterious mutations. These diagnostic criteria also appear to describe human intelligence (the g factor). This paper argues that during human evolution, mate choice by both sexes focused increasingly on intelligence as a major heritable component of biological fitness. Many human specific behaviours (such as conversation, music production, artistic ability and humour) may have evolved principally to advertise intelligence during courtship. Though these mental adaptations may be modular at the level of psychological functioning, their efficiencies may be tightly intercorrelated because they still tap into common genetic and neurophysiological variables associated with fitness itself. Although the g factor (like the superordinate factor of fitness itself) probably exists in all animal species, humans evolved an unusually high degree of interest in assessing each other's intelligence during courtship and other social interactions--and, consequently, a unique suite of highly g-loaded mental adaptations for advertising their intelligence to one another through linguistic and cultural interaction. This paper includes nine novel, testable predictions about human intelligence derived from sexual selection theory. PMID- 11276908 TI - The g factor: psychometrics and biology. AB - General ability, defined as psychometric g, arises from the empirical fact that scores on various cognitive tests are positively correlated in the population. The g factor is highly stable across different factor analytic algorithms, across different test batteries and across different populations. Because all cognitive tests, from the simplest to the most complex, regardless of their informational content, are g-loaded to varying degrees, g cannot be described in terms of the tests' content, or even in psychological terms. It is actually a property of the brain. The loadings of various tests on g, from tests of sensory discrimination and reaction time to those of highly complex problem solving, predict those tests' degree of correlation with a number of non-psychometric variables: the test's heritability, inbreeding depression, coefficient of assortative mating, brain size, reaction time, brain nerve conduction velocity, brain glucose metabolic rate and features of brain evoked potentials. Although some of the brain's cognitive functions are modular, the g factor reflects the all-positive correlations among virtually all cognitive functions that show individual differences. I hypothesize that the brain contains no module for general problem solving. Correlations between individuals' performances in various cognitive tasks result from quantitative individual differences in physiological conditions that do not constitute the brain's modular and other neural design features but do influence their speed and efficiency of information processing. PMID- 11276909 TI - Psychometric intelligence differences and brain function. AB - Psychometric intelligence attracts a converging consensus about its phenotypic structure. Mental ability test scores have proven predictive validity. However, although individual differences in mental abilities can be measured, they are not understood. A long-standing aim of the 'London School' of British psychologists, since Galton and Spearman, is to understand the origins of psychometric intelligence differences in terms of individual differences in brain processes. The history of this research is described, as is the rise in interest since the 1970s. The first problem, met since antiquity, is to discover the relevant levels of brain function. Thus, aspects of brain function that 'explain' psychometric intelligence differences are sought at psychometric, cognitive, psychophysical, physiological, neurochemical and genetic levels. The growing points and dead-ends within each of these levels are identified. Special attention is given to research that crosses levels of description of brain function. Two types of multi level brain function research are discussed, 'correlational' and 'circumstantial/experimental,' and examples of each are described. Illustrating both approaches, there is a detailed account of research on inspection time that discusses how psychometric intelligence-brain process correlations at one level (psychophysical) may be expanded using event-related potentials, psychopharmacology and functional magnetic resonance imaging. PMID- 11276910 TI - Intelligence: success and fitness. AB - This chapter presents the consensus among psychometricians regarding the construct of general intelligence ('g') and its measurement. More than any other construct, g illustrates the scientific power of construct validation research. To date, g is carried by more assessment vehicles and saturates more aspects of life than any other dimension of human variation uncovered by psychological science. Phenomena most vital to the core of g's nomological network are reviewed (e.g. abstract learning, information processing, and dealing with novelty). This is followed by coverage of relevant but more peripheral phenomena (e.g. crime, health risk behaviour, and income). Because g constitutes such a ubiquitous aspect of the human condition, its omission in social science research often results in underdetermined causal modelling. Frequently, this constitutes a longstanding error in inductive logic, namely, the Fallacy of the Neglected Aspect. Attending to Carnap's Total Evidence Rule can help to forestall neglected aspects in scientific reasoning. PMID- 11276911 TI - The g factor in non-human animals. AB - Animals possess the attributes we label as 'intelligent' in humans. 'Insight' and 'reasoning' have been demonstrated in chimpanzees, monkeys, racoons, rats, mice, ravens and pigeons. In the rat, the animal species best characterized psychologically and neuroanatomically, reasoning ability correlates with other cognitive capacities and brain size. Other learning task paradigms tested on mice and rats have confirmed consistent individual differences, indicated a neuroanatomical network for learning, and shown the presence of genetic influences for cognitive ability. Animals offer an opportunity to test ideas about intelligence that cannot be performed on humans. Methylazoxymethanol (MAM) administered prenatally can arrest cortical cell division and produce a 'mentally retarded' microcephalic rat. This intellectual deficiency can be ameliorated by postnatal induction of dendritic arborization and synapse formation with naltrexone, suggesting the relevance of neuronal and synapse number for behavioural variation in rat g. Inbred mice lines differ in brain size and behaviour, permitting, through the use of recombinant inbred strains, the determination of genetic loci with quantitative effects on structure and function. Lastly, genetic contributions to g can be directly tested by modifying gene expression and determining the anatomical, physiological, and behavioural benefits. PMID- 11276913 TI - Culture club. PMID- 11276912 TI - Natural selection, mental modules and intelligence. AB - The question of whether intelligence is one trait or many has exercised several generations of researchers, but no consensus is in sight. Evolutionary psychology, with its emphasis on domain-specific mental modules, seems to offer hope for advancing understanding of this question. We know that the mind has been shaped by natural selection to maximize reproductive success. This tells us what the mind must do--it must solve the adaptive problems that the organism confronts. However, whether this functional capacity is manifest in congruent anatomic, physiological, genetic, cognitive or psychometric structures is another matter. Examination of how natural selection shaped other mechanisms suggests that knowing functional demands provides only modest guidance as to the structure of mechanisms. None the less, it remains simultaneously clear that these mechanisms are not entirely general, but have been shaped to cope with specific challenges. Our metaphors for the mind, whether as a digital computer or a Swiss army knife, are misleading because computers and tools are products of intelligent design. In contrast, minds are products of natural selection whose intertwined components are products of incorporated genetic mutations whose effects are widespread and constrained by historical precedents. Our tendencies to describe the structure of the mind in terms of discrete components make it difficult for us to comprehend the mind as a mind. One antidote may be to minimize metaphorical descriptions of postulated structures of mind and focus instead on its function. PMID- 11276914 TI - Friendly infections. PMID- 11276915 TI - The dangerous desk. Mouse-clicking isn't heavy lifting, but it can cause injury. How to keep your job from immobilizing you. PMID- 11276916 TI - Sensitivity, specificity, and predictive accuracies of various noninvasive techniques for detecting hibernating myocardium. PMID- 11276917 TI - Vitamin E and the prevention of atherosclerosis. AB - Successful strategy for the prevention of coronary heart disease (CHD) in particular of atherosclerosis, require a detailed understanding of the underlying mechanism. It is now being recognised that dietary antioxidants, in particular vitamin E, will play an important role in designing future strategies. Although more and more beneficial effects of vitamin E on atherosclerosis are being described, the biochemical and cell biological mechanism underlying these benefits are not yet fully understood, preventing the use of vitamin E as therapeutic agent. Recent new findings have shed new light on the physiological role of vitamin E and suggest that it has a much broader array of biological activities than originally expected. In addition to its well described role as an antioxidant, it is becoming evident that vitamin E also can modulate the immune system, suppress local and chronic inflammation, reduce blood coagulation and thrombus formation, and enhance cell function and survival. This review summarises new findings from in vitro studies and discusses their potential relevance in human atherosclerosis. PMID- 11276918 TI - Are the serum 25-hydroxyvitamin D concentrations in winter associated with forearm bone mineral density in healthy elderly Japanese women? AB - The objective of this study was to investigate whether the serum 25 hydroxyvitamin D (25[OH]D) concentrations in winter are associated with the BMD in elderly Japanese women. The subjects were 117 healthy elderly Japanese women. Serum 25(OH)D concentrations were determined by high-performance liquid chromatography. Forearm BMD in the non-dominant arm was measured by dual-energy X ray absorptiometry (DXA) using a DTX-200 Osteometer. The mean age of the subjects was 66.1 (SD 6.5) years (range: 46-80). The average 25(OH)D concentration was 59.1 nmol/L (SD 16.1), and five of the subjects had low 25(OH)D concentrations (< 30 nmol/L). Forearm BMD decreased linearly with age (r2 = 0.275). There was no linear association between the serum 25(OH)D concentrations and the forearm BMD (p = 0.9483). Multiple regression analysis did not reveal any association between the two (p = 0.5318) when age (p < 0.0001, r2 = 0.271) and weight (p < 0.0001, r2 = 0.153) were taken into account. Our cross-sectional study failed to reveal any association between the serum 25(OH)D concentrations and the forearm BMD in elderly Japanese women, suggesting that 25(OH)D does not play an important role in the determination of BMD. A follow-up study should be conducted to confirm the results of our cross-sectional study. PMID- 11276919 TI - Supplementation of rats with a lutein mixture preserved with vitamin E reduces tissue phylloquinone and menaquinone-4. AB - The modulation of tissue concentrations of vitamin K by a lutein supplement preserved with natural vitamin E was studied in Fischer 344 rats. Vitamin K is necessary for blood coagulation and may be essential for tissue and bone health. Weanling male rats were fed the AIN-93G diet (control) or modified AIN-93G diets containing 0.3, 0.6, 1.2, 2.4 and 4.8 g supplement/100 g diet for 8 weeks. The supplement contained 5% lutein, 0.22% zeaxanthin and 2.2% natural vitamin E as a preservative. Concentrations of trans-phylloquinone in the plasma (nmol/mmol triglycerides) and heart were significantly reduced (P < or = 0.05) in rats fed the supplement. The reductions in trans-phylloquinone in the heart ranged from approximately 20 to 60% of the control. Concentrations of phylloquinone in the liver were significantly lower in the rats fed the supplement at levels > or = 1.2 g/100 g diet than in the control rats. Ratios of cis/trans phylloquinone in liver and heart increased and concentrations of menaquinone-4 in heart decreased as the dietary level of the lutein supplement increased. The results suggest that the lutein supplement affected the absorption, tissue uptake and/or turnover rate of vitamin K. The presence of other components in the supplement confounded the interpretation of the biological effects of lutein alone on vitamin K metabolism. PMID- 11276920 TI - Interactive effect of hesperidin and vitamin E supplements on cholesterol metabolism in high cholesterol-fed rats. AB - Certain bioflavonoids are potent antioxidants and have pharmacologic effects similar to those of vitamin E. Accordingly, the interactive effect of hesperidin and vitamin E was studied with respect to cholesterol metabolism and the antioxidant status. Hesperidin supplement (0.1%, wt/wt) with comparable levels of vitamin E was provided with a high-cholesterol (1%, wt/wt) diet to rats for 5 weeks. The amount of vitamin E included in the hesperidin-free and hesperidin diets was either a low (low-E) or a normal (normal-E) level. The hesperidin supplement and different levels of dietary vitamin E did not significantly alter the concentrations of plasma triglycerides. However, the inclusion of hesperidin significantly lowered the concentration of plasma cholesterol in both the low vitamin E group and the normal-vitamin E group compared to the hesperidin-free groups (p < 0.05). The hepatic triglyceride content was significantly lowered by the hesperidin supplement, as opposed to the plasma triglyceride content, regardless of the vitamin E level in the diet. The hepatic HMG-CoA reductase activity was significantly lowered by the hesperidin supplement with both the low vitamin E and the normal-vitamin E compared to the hesperidin-free groups (p < 0.05). The hepatic HMG-CoA reductase activity was also significantly lowered with an increase in the dietary vitamin E within the hesperidin and hesperidin-free groups. The excretion of fecal neutral sterol and acidic sterols tended to be lower with the hesperidin supplement. Neither dietary hesperidin nor vitamin E significantly changed the hepatic antioxidant enzyme activity. This data indicates that hesperidin lowers the concentration of plasma cholesterol and the hepatic triglyceride content regardless of the dietary vitamin E level. However, the concentration of plasma cholesterol in the hesperidin-free groups was dependent on the dietary vitamin E level. This information may contribute to understanding the interactive effect of hesperidin and vitamin E on cholesterol biosynthesis in high cholesterol-fed rats. PMID- 11276921 TI - Supplementation with vitamin E and/or zinc does not attenuate atherosclerosis in apolipoprotein E-deficient mice fed a high-fat, high-cholesterol diet. AB - Ever since oxidation has been known to be involved in atherogenesis, antioxidants have received considerable attention as potential antiatherogenic agents. The lipid-soluble vitamin E is the main antioxidant carried by lipoproteins. Zinc is a water-soluble trace element that acts as a cofactor of superoxide dismutase (SOD) and has an antioxidant role in several oxidative processes. To test the hypothesis that zinc could adjuvate the antioxidant activity of vitamin E and diminish atherogenesis, we explored how supplementing diet with vitamin E and/or zinc would affect an atherosclerosis-prone animal like Apo E-deficient mice. The increased plasma concentrations of both vitamin E and zinc showed that absorption was high. They had a significant hypolipidemic effect and the supplemented animals had 25% less plasma cholesterol and triglyceride than controls. The SOD activity was significantly higher in washed erythrocytes from mice supplemented with zinc. The plasma of supplemented animals was also significantly more resistant to oxidation. The size of lesions in the proximal aortic region did not differ among groups. Therefore, dietary supplementation resulted in the expected antioxidant effects but there was no substantial attenuation of atherosclerosis in this particular model. PMID- 11276922 TI - The role of folate, antioxidant vitamins and other constituents in fruit and vegetables in the prevention of cardiovascular disease: the epidemiological evidence. AB - Evidence that fruit and vegetables may protect against coronary heart disease is accumulating. It is unclear which constituents of fruit and vegetables are responsible for this protective effect. Folate as a co-substrate in homocysteine metabolism may be important. An intake of about 400 micrograms folate equivalents/day seems to be required to achieve stable low homocysteine blood levels. Five of eight epidemiologic studies show significant inverse associations between folate and cardiovascular disease. These associations could be confounded by antioxidant vitamins and/or other substances. In trials examining an association between folate and cardiovascular disease such confounding must be excluded, before specific recommendations can be given. Observational studies suggest that vitamin C plays a role in the aetiology of cardiovascular disease, but there are no completed intervention trials of this vitamin alone. With regard to vitamin E two cohort studies point to cardiovascular benefits with the long term use of supplements of at least 100 IU/day, but the results of controlled trials are inconclusive. There is some evidence from observational studies of an inverse association between beta-carotene and cardiovascular disease, particularly in smokers. Intervention trials do not support this hypothesis, rather, they suggest a possible harmful effect of beta-carotene supplements in smokers. Nevertheless, protective effects of beta-carotene and vitamin E in different dosages, durations of administration, or different combinations are still possible. The last paragraph of this review discusses limitations of the present and priorities of future research. PMID- 11276923 TI - Associations of age, smoking habits and diabetes with plasma vitamin C of elderly of the POLA study. AB - The objective of this study was to determine the associations of age and sex with plasma vitamin C (vit C) concentration taking into account smoking habits and the presence of age-related pathologies, such as diabetes. The POLA study is a population-based study on age-related eye diseases and their risk factors, and plasma Vitamin C evaluation is part of the biological parameters measured in the 1987 volunteer subjects living in Sete (South of France) and aged more than 60 years. Men had lower average plasma vit C levels than women (31.6 microM.L-1 versus 40.3 microM.L-1, p = 0.001). Plasma vit C was stable as a function of age in women but decreased in men (p = 0.02), enhancing the difference in vit C concentration between men and women with advancing age. Smoking more than 10 cigarettes a day was associated to a lower plasma vit C concentration in men (p = 0.001) but not in women, and diabetic subjects tended to have lower vit C concentrations, the difference being significant only in women (p = 0.003). We conclude that there is a clear influence of sex on plasma vit C. This difference may be due to dietary habits, or metabolism, but may also be due to different sensitivity of age, smoking and to some pathologies. PMID- 11276924 TI - Vitamin B-12-deficiency affects immunoglobulin production and cytokine levels in mice. AB - To clarify the role of B-12 in the immunological function, serum C3, IgM, IgG, IgE contents, splenocytes expression of CD4, CD8, and CD4 positive intracellular IFN-gamma and IL-4 were examined in B-12-deficient mice, and the effect of the administration of CH3-B-12 was also studied. Serum C3, IgM and IgG contents were lower in B-12-deficient mice than in the control mice. On the other hand, serum IgE content was significantly higher in B-12-deficient mice, and the value in CH3 B-12 administered mice, administered CH3-B-12 to B-12-deficient mice for 48 h before the end of feeding period, showed a tendency to recovery. CD4+CD8- cells and CD4+CD8-/CD4-CD8+ ratio in splenocytes were significantly higher in B-12 deficient mice than in control mice. CD4+IFN-gamma+ cells was significantly lower in B-12-deficient mice than in control mice, and CD4+IL-4+ was significantly higher in B-12-deficient mice than in control mice. These results suggest that B 12-deficiency causes CD4+CD8-T cells shift from the T helper type 1 to the T helper type 2, which participate in the IgE production and elevates CD4+CD8-/CD4 CD8+ ratio. Thus, B-12 plays a role in maintaining the immune function in mice. PMID- 11276926 TI - Effects of transient neonatal hyperthyrotropinemia on intellectual quotient and psychomotor performance. AB - Transient neonatal hyperthyrotropinemia (TNH) occurs frequently in areas of iodine deficiency. To evaluate the effect of TNH in intellectual function and psychomotor performance, a historical cohohrt study was performed in 9 years old children with documented TNH at birth. 18 children with TNH who had been born in Mahdieh Hospital were studied at age 9 and compared to 19 matcheal children born at the same time, but having normal thyroid function at birth. Global intelligence (IQ) and psychomotor performance were evaluated with Raven and Bender-Gestalt tests, respectively. Total serum T4 and T3 by commercial RIA and TSH by IRMA. Urine was tested for iodine content by digestion method. Height and weight were similar in two groups at birth and at 9 years of age. Thyroid function tests were similar in the two groups except for TSH at birth which was higher in TNH than in control group (23.4 +/- 8.3 vs 3.6 +/- 1.0 mU/L, P < 0.001). Results of T4, T3, resine uptake, and urinary iodine at 9 years of age were not different between two groups. Mean IQ was 98 +/- 11 and 106 +/- 8 in TNH and normal children, respectively (P < 0.01). There was no significant difference between psychomotor performance in the two groups. There was no correlation between TSH at birth and IQ at 9 years of age. The present finding suggests that TNH can adversely affect longterm intellectual development. PMID- 11276925 TI - Serum free carnitine and total triglycerid levels in children with iron deficiency anemia. AB - Iron deficiency anemia and hyperlipidemia are common public health problems in Turkey. The connection between iron and lipid metabolisms has not been clarified yet. The aim of the study was to determine the effect of iron deficiency on carnitine and lipid metabolism. Study group was consisted of 70 children (mean age 14.7 +/- 1.3 months) suffering from iron deficiency anemia and 20 healthy children (mean age 13.7 +/- 1.2 months) attended to outpatient clinics of Cumhuriyet University, Sivas were enrolled the study as the control group. Assessments of serum free carnitine concentrations, total triglyceride, total cholesterol and VLDL levels were made in both groups. The mean serum free carnitine concentration was significantly lower than the control group (18.9 +/- 0.43 nmol/ml and 45.9 +/- 1.47 nmol/ml respectively, t = 17.5 p < 0.01). Results of our study also indicated higher serum total triglyceride, total cholesterol and VLDL levels in iron deficient patients than the healthy controls. Regression analyses indicated a negative correlation between serum free carnitine and total triglyceride levels in iron deficient patients. This study confirms that iron deficiency anemia may be linked to the endogenous carnitine synthesis in pediatric age group, and thus hyperlipidemia appears to be a risk factor for premature cardiovascular diseases. PMID- 11276927 TI - Serum leptin and lipid profiles in Thai obese and overweight subjects. AB - The weight, height, body mass index (BMI), waist/hip ratio, serum leptin and lipid profiles of 48 overweight (BMI > or = 25.00). Thai males and 166 overweight Thai females, compared with 26 males and 81 females in a control group (BMI = 18.5-24.9 kg/m2), were investigated. Subjects for the study were those persons who turned up regularly for physical check-ups at the out-patient department, general practice section of the Rajvithi Hospital, Bangkok. The study was conducted between March-October, 1998. Statistically significantly higher levels of serum leptin, cholesterol, LDL-C, LDL-C/HDL-C ratio and triglyceride were found in the overweight compared with the control subjects. The median serum leptin concentration in overweight subjects was 19.6 (2.0-60.0 ng/ml) compared with 9.0 (range 1.0-30.0 ng/ml) in the control subjects (p < 0.001). The median values of leptin serum concentrations in the overweight and obese males were significantly higher than those of the overweight and obese females. A total of 66.7% (32 out of 48) of the overweight and obese males had elevated leptin levels, while elevated leptin levels were found in 87.3% (145 out of 166) of the overweight and obese females. A total of 18.8% and 21.1% of the overweight and obese males and females respectively had cholesterol concentrations of > or = 6.48 mmol/l. However, the prevalence of low HDL-C (HDL-C < or = 0.91 mmol/l) was found to be 41.7% in the overweight and obese males and 4.2% in the overweight and obese females. Statistically significant associations were found between weight, height, BMI, waist, hip, waist/hip ratio, HDL-C, and serum leptin in both overweight male and female subjects. A negative correlation was found between serum leptin and LDL-C/HDL-C ratio in both the overweight and obese subjects. PMID- 11276928 TI - Effect of supplemental methionine on plasma homocysteine concentrations in healthy men: a preliminary study. AB - Hyperhomocysteinaemia is an established risk factor for vascular disease. The only source of homocysteine in humans is the amino acid methionine found in dietary protein. In an 8-week study, fasting plasma homocysteine concentrations were examined in a group of healthy male subjects (n = 6) under usual dietary conditions (weeks 1-4) and in response to weekly graded (25, 50 and 75 mg/kg/d) supplementary methionine (weeks 5, 6, 7). Nutrient intakes, including methionine, were calculated from 4 x 3 day food records. Under usual dietary conditions (mean methionine intake; 0.95 +/- 0.51 mg/d) weekly mean plasma homocysteine concentrations for the group were not significantly different (ANOVA) from each other ranging from 6.82 +/- 1.77 to 9.42 +/- 2.73 mumol/l. Doubling (supplementing with 25 mg/kg/d; + 2.04 g/d) or quadrupling (50 mg/kg/d; + 4.08 g/d) methionine intakes did not result in a significant increase in plasma homocysteine (8.56 +/- 3.68 mumol/l and 13.37 +/- 5.09 mumol/l respectively). A significant increase, however, was achieved when diets were supplemented with methionine at the highest level of 75 mg/kg/d (+6.14 g/d) resulting in a mean plasma homocysteine concentration of 18.05 +/- 11.8 mumol/l. Mean plasma homocysteine concentration returned to baseline (8.76 +/- 3.42 mumol/l), 10 days post-supplementation. The results of this study indicate that an increased dietary methionine will only cause elevated fasting homocysteine concentrations if ingested at intakes equivalent to five times usual intake. Because it is very unlikely that such levels could be achieved through dietary means alone we conclude that plasma homocysteine is unlikely to be affected by longer-term changes in food methionine intake. PMID- 11276929 TI - Rapid determination of glutathione peroxidase and thioredoxin reductase activities using a 96-well microplate format: comparison to standard cuvette based assays. AB - Gluthatione peroxidase and thioredoxin reductase are selenocysteine-containing enzymes that are constituents of the cellular antioxidant defense system. Conventional cuvette-based assays for glutathione peroxidase and thioredoxin reductase enzymes are laborious and time consuming. The ability to assay their activities rapidly in multiple samples would aid efforts focused on understanding the impact of these enzymes on the cellular antioxidant defense system. High throughput can be achieved with assays adapted to work in a clinical analyzer but require expensive equipment. Assays designed to work in a 96-well microplate reader provide an alternative methodology for high throughput with reduced instrumentation cost. However, due to differences in the light pathlength when using a 96-well format, the values obtained cannot be compared directly with those obtained using a 1-cm cuvette. Described here are assays for glutathione peroxidase and thioredoxin reductase modified to work in a 96-well format that incorporates light pathlength determinations into the assays. The values obtained using a high throughput 96-well format in conjunction with pathlength determinations are in agreement with those obtained using a standard 1-cm cuvette. While spectrophotometrically derived pathlengths are the most accurate, calculated pathlengths based on assay volume and well size can be used with only a small amount of error introduced. This method can also be applied to many other enzyme assays, thus allowing the rapid analysis of large numbers of samples without the need for expensive equipment. PMID- 11276930 TI - Politics. One gender, two agendas. PMID- 11276931 TI - Primary care. Lock, stock-take and barrel. AB - An analysis of primary care groups' investment plans revealed their main concerns were premises, IT and workforce issues. About a quarter of plans did not address out-of-hours care or community nursing. The plans lacked detail of the process guiding resource allocation. The plans showed little indication of services being organised around the priorities of the local health improvement programme. PMID- 11276932 TI - e-novation. Central reservation. PMID- 11276933 TI - e-novation. To the letter. PMID- 11276934 TI - e-novation. Wobbly at the top. PMID- 11276935 TI - e-novation. Come together. PMID- 11276936 TI - The growing crisis in mental health care. PMID- 11276937 TI - The hospital sector in Germany: an overview. AB - The German health care system has recently experienced far reaching reforms within its hospital sector, including a new financing and remuneration system, the life span of which is to the year 2003. The reforms have been based on the DRG-system already in operation in another country. Other regulations of the new reform involve the system of quality assurance and quality management in German hospitals and a new form of supply of services, the so called "integrated care" system, the introduction of which was to overcome the fragmented system of ambulatory and stationary medical care in Germany. This article, in short, presents an overview of the hospital sector in Germany and reports on the latest developments resulting from implementation of the health care reform. PMID- 11276938 TI - [Man, health, development and economics]. AB - In the next few years the key topics of the health debate in Latin America and the Caribbean will centre on the relationship between health, development and economics. The author suggests that there is a more important issue at the heart of this question, that is whether man should be the central element of any health and social system and the order of priority this consideration should be given with regards to development strategy and economic planning. PMID- 11276939 TI - From district general hospital to local general hospital. AB - Much debate has centred around the future of acute services in the NHS in the UK with particular reference to the role and function of the District General Hospital. This paper explores a new model of hospital development--the 'Local General Hospital' which lies somewhere between the DGH and the cottage or community hospital. The paper uses the case study of Neath General Hospital in South Wales which moved to a LGH model. The paper explores the functional content of the LGH balancing accessibility with cost and clinical effectiveness. The paper makes particular reference to the development of clinical networks which cross organisational boundaries and the support of GPs and consultant medical staff as key success factors in the transition from DGH to LGH. PMID- 11276941 TI - Environmental design effects on Alzheimer symptoms in long-term care residences. AB - As Alzheimer's disease becomes the most extensive and expensive disease in the world, the need to broaden the definition of treatment becomes more and more necessary. Medication is presently the first, and often the last, step in "treatment." But medication alone offers meager solace for the millions of people who live daily with this degenerative disease. The world they live in becomes increasingly strange and their behavior increasingly erratic. In order for most people with this disease to live lives that tend toward normalcy, a combination of pharmacological and non-pharmacological remedies is necessary. Therapeutic settings, activities that make up for cognitive deficits, family involvement, and caregiver approaches must augment medication. One non-pharmacological treatment modality is a physical environment that helps people find their way, jogs their memory, and keeps them safe. This paper recounts an investigative journey to discover what are the components of a multi-approach treatment for Alzheimer's disease, what role the physical environment plays in treatment, and what effects the physical environment has on the behavior of people with Alzheimer's disease. This paper describes and links the work of a small group of investigators and practitioners over the last decade of the 20th century (Zeisel, Hyde, Levkoff 1994; Hyde 1987; Zeisel, 1999; Zeisel and Raia, 2000; Zeisel, Silverstein, Hyde, Powell, and Holmes, 2001). PMID- 11276940 TI - Health insurance and hospitalisation in urban China: bending to the wind of change. AB - The transformation from a planned economy to a market-oriented one in China has had profound impact on access to health care for a vast majority of the Chinese population. This article used the data from the national household health surveys conducted in 1993 and 1998 to show that the coverage of health insurance declined significantly from 54% to 39% among the urban population over the period, while rate of hospital admission came down from 45 inpatients per 1,000 people in 1992 to 30 inpatients per 1,000 people in 1997. It concludes that the declined coverage of health insurance and the rapid rise of medical care costs were the major factors affecting the access of the Chinese population to hospital care services. PMID- 11276943 TI - Measuring and benchmarking quality in hospitals. AB - As the cost of health care increases, but the resources available to deliver patient care decrease, the issue of quality becomes a major concern for everyone either receiving or delivering health care services. Adding to the complexity of the quality issues facing health care institutions have been a number of recent health care articles reporting patient safety issues. These issues have ranged from medical mistakes to medication errors. Factors cited as contributing to these errors have been the shortage of resources, shortage of professionals to provide direct patient care, i.e. nurses, and the increasing cost of providing health care services. PMID- 11276942 TI - Impact of insurance coverage type on laboratory test ordering behaviour of general practitioners. AB - BACKGROUND: There exists much variation between GP's in the use of laboratory tests. Although the requesting pattern of GPs has been extensively described in the literature, little is still known of the factors which influence the GP's test ordering behaviour. AIM: This study aimed to determine whether the payment scheme according to which general practitioners are reimbursed influences the laboratory test ordering behaviour. METHOD: The laboratory test ordering behaviour of the general practitioners of Tilburg, a town with 180,000 citizens in the south of The Netherlands, was studied during a four month period, in relation to the type of insurance coverage of the patients. Two types of insurance were considered: voluntary and compulsory. The data were collected from the laboratory administration and coupled with information obtained from two, interview rounds. RESULTS: Two findings support the hypothesis that the type of insurance coverage of the patient, has an impact on the test ordering behaviour of the physician: The ratio between laboratory requests for sickness fund patients and patients with a private health insurance was found to depend on the fraction of persons with a private health insurance within the family practice. This was tested with multiple linear regression analysis. General practices were divided into two subgroups, those with many > 29%) and few (< 29%), voluntarily insured patients. Where a patient was privately insured it was found that relatively more tests were ordered. In case of general practices with many voluntarily insured patients this distinction disappears. The relative proportion of voluntarily insured patients was found to be an important variable in explaining the test ordering behaviour of general practice physicians in Tilburg. PMID- 11276944 TI - Gaining insight into leadership skills. PMID- 11276945 TI - Status of quality initiatives: one year after the Institute of Medicine's report. AB - Over the past several years there has been an increasing public interest in the quality of health care provided by professionals, institutions and health plans. In the spring of 1998, the President's Advisory Commission on Consumer Protection and Quality in the Health Care Industry issued its final report, Quality First: Better Health Care for All Americans. It identified four areas of needed improvement in health care quality--errors in health services, the underuse of services, the overuse of services, and variation in the use of services. PMID- 11276946 TI - Working toward quality improvement. AB - The National Forum for Health Care Quality Measurement and Reporting was created to develop and implement a national strategy for measuring and reporting health care quality. It grew from a report issued in 1998 by the President's Advisory Commission on Consumer Protection and Quality in the Health Care Industry, which proposed the creation of the Quality Forum as part of an integrated national quality improvement agenda. PMID- 11276947 TI - If we're not merging for for the people, why bother? AB - When Batts Inc., a family business that invented wooden clothes hangers in 1903 was sold to Tyco International, it's ardent competitor, in 1999, nobody bothered to tell its 500 loyal employees in Holland that the plants would be closed, their jobs--perhaps entire careers--ended. And when Lescoa, a family-owned, auto accessory manufacturing company in west Michigan since 1945, was combined with American Bumper, it's competitor from Ionia, the pledge to "not take the company apart" was communicated. However, that pledge was quickly forgotten as American's executives took over, ignoring the family-styled people-culture of decades past. PMID- 11276948 TI - Hospital helps insure more children. AB - Marquette General Hospital's involvement in an initiative to insure children has brought local credibility to a nationwide program and added strength to the hospital's mission. PMID- 11276949 TI - Just cause or just because. AB - Determining the discipline or discharge of an employee is one of the most important and imperfect tasks a manager will probably ever have to do. It is tricky because there are so many questions to answer. Who did what and when? Who's telling the truth and who's bending it? Who knew (or should have known) about rule violations and what was done with this knowledge? What was the intent of the employee and what were the circumstances? PMID- 11276950 TI - Leaping into patient safety. AB - Last November, the Leapfrog Group unveiled an ambitious effort to improve patient safety across the nation. Sponsored by the Business Roundtable, the Leapfrog Group is a consortium of Fortune 500 companies and other large private and public health care purchasers. The group is working to mobilize employer purchasing power to affect big "leaps" in patient safety by educating consumers and rewarding health care providers who meet defined safety standards. PMID- 11276951 TI - Career management: a competitive advantage in today's health care marketplace. AB - A valuable new tool to attract and retain new employees is being used by some of the most progressive companies in Michigan. It is called career management, and it is being used with great success by businesses of all types to give themselves a competitive advantage. PMID- 11276952 TI - Making health care accessible to all. AB - The Muskegon Community Health Project was established in 1994 through a partnership between the W.K. Kellogg Foundation and the Community Foundation for Muskegon County as part of Kellogg's Comprehensive Community Health Models of Michigan (CCHMs) initiative. Muskegon was one of three counties (along with Calhoun and St. Clair) where CCHMs served as a catalyst to bring community members together to identify and implement strategies to improve health care and delivery. PMID- 11276953 TI - American renaissance. AB - A twenty-first century American renaissance is in the making. Powerful social, political, technological, economic and spiritual forces are converging to create new possibilities for our nation and our health care system. We are becoming a designer nation. An increasing percentage of our population are cultural creative with a mandate to create a healthier society that works for everyone. PMID- 11276954 TI - Reluctant conclusion: government fails health care. AB - In the 1940s, 1950s and 1960s, government created modern health care's infrastructure, expanded and guaranteed health services for most Americans, and was the catalyst for new cures and treatments. PMID- 11276955 TI - Kaleidoscope of quality--putting the pieces in place. AB - Quality health care--patients and families expect to receive it. Clinicians work to provide it. Employers and third party payers insist on purchasing it. Regulatory and accrediting bodies set standards and monitor to assure it exists. Finally, a health care issue we all agree on, right? Well, not exactly.... PMID- 11276957 TI - [Expert assessment of medical malpractice]. AB - Conciliation boards are advisory commissions of the medical societies. They deal with clarification of extrajudicial patient claims based on factual or putative treatment mistakes. In perhaps 90% of the cases, an extrajudicial solution is possible, whereby the federal average for recognition is 30%. Within the framework of this study, first the legal basis for the liability of doctors is explained, especially the importance of patient information and consent as well as the responsibility for documentation. The material of the conciliation board of the medical society of Saxony consists of 1375 medical expert opinions rendered between 1 January 1992 and 23 September 2000, of which 165 cases were due to orthopedic treatment. The most common health injuries resulting in compensation claims are described. PMID- 11276956 TI - [Expert assessment of occupational diseases]. AB - Since the enactment of the first Occupational Illness Ordinance (Berufs Krankheiten-Verordnung, BKV) in 1925, not only the prevention of and compensation for workplace accidents but also the competence for recognizing specific occupational diseases has been imposed on the occupational association. Analogous to technological advancement, the BKV has taken into account new hazards with corresponding amplifications. Despite this system of listing specific diseases, it is necessary to perform an investigation of causality for each individual case, which can only be successful with reliable information about the recognizable illness and its differential diagnosis as well as about the legal guidelines for an examination of causality. The mitigation produced by the administration of justice with the theory of properly significant conditions and the comparatively minor requirements for evidence with respect to origin nevertheless do not resolve all of the problems in the practical application of occupational illness law. This uncertainty reoccurs in almost all of the so called "surgical" occupational diseases (BK2101-2110). This is not difficult to recognize in the following descriptions and elucidations of the specific diseases. Recommendations for the practical rendering of an expert opinion are provided. PMID- 11276958 TI - [Intervertebral disk displacement and trauma]. AB - It is difficult to find medical evidence of a correlation between a lumbar disk disease and trauma. One should consider whether the individual degeneration of lumbar disks or the trauma lead to the typical complaints. Disk disease in the population are very common. Therefore the relevance of the individual affection before trauma has to be considered. Spinal trauma with its sudden, incidental onset needs to be differentiated from purposeful and conscious movements. An intervertebral disk disease can be classified as accident related only in cases involving adequate trauma, with no previous complains, and a sudden onset of pain. PMID- 11276959 TI - [Recurrence of idiopathic arteriovenous malformation of the finger. A case report]. AB - We report on a 30-year-old female with a recurrence of an AV malformation after a 20-year interval. Based on the literature, an overview of the diagnosis and therapy of this rare anomaly is given. PMID- 11276960 TI - [Basic principles of medical assessment of the locomotor system]. AB - Rendering a medical expert opinion on the musculoskeletal system is an extraordinary responsibility. Nearly all physicians are confronted with this medical task in their daily clinical practice. Difficulties encountered and misjudgments made in formulating an expert opinion are certainly primarily a result of insufficient knowledge of the sociomedical basis. Proper organization and compliance with a standardized structure are especially important for an expert opinion. Intensified teaching of the principles and importance of the medical expert opinion during medical training will not only result in optimization and acceleration of the sociomedical procedures, it will also awaken pleasure in this many-sided activity. PMID- 11276961 TI - [Expert assessment within the scope of the severe disability regulation and social compensation law]. AB - In the wake of the First and Second World Wars, an extensive system of social compensation and assistance for the war disabled evolved in Germany and later on also for civilians. Depending on the origin of the disability, the benefits are composed either of revenues and pensions and/or tax reductions and nonfinancial assistance. In any case, the extent of these benefits depends on the grade of disability (GdB/MdE) determined by medical examination and on expert opinions in accordance with detailed legal regulations. PMID- 11276962 TI - [Expert assessment of accident sequelae in statutory and private accident insurance]. AB - Public and private accident insurance companies have in common that the insurance risk is the damage to health caused by trauma. Otherwise, they are fundamentally different. Public accident insurance as part of social law has the function of providing compensation for damage. Functional impairment due to trauma is compensated abstractly according to the general labor market. Private accident insurance as part of civil law is an insurance of fixed sums anchored in the agreement. It is completely unrelated to the consequences of trauma-induced limitation of function. PMID- 11276963 TI - Chromosome aberrations and micronucleus frequency in anaesthesiology personnel. AB - Occupational exposure to anaesthetic gases is associated with various adverse health effects. Genetic material is a sensitive target of numerous harmful agents. The aim of this study was to examine whether chromosomal damage could serve to indicate exposure to anaesthetics. Twenty-eight anaesthesiologists, 16 technicians, and 32 control subjects were examined for chromosome aberrations and micronucleus frequency. An increase in chromosome damage was found in both exposed groups. Micronucleus frequency increased significantly, showing higher rates in women. The observed differences between the sexes in respect to the exposure risk call for further, targeted investigation. PMID- 11276964 TI - Renal function in miners intermittently exposed to elemental mercury vapour. AB - The authors investigated renal damage in 45 mercury miners under conditions of relatively short and low-level exposure to elemental (metallic) mercury vapour (Hg0). The analysis included urinary mercury, immunoelectrophoresis of urinary proteins, immunofixation and high-resolution electrophoresis, quantitative analysis of urinary albumin, and urinary alpha 1-microglobulin before and after exposure. The activity of urinary N-acetyl-beta-D-glucosaminidase (NAG) enzyme was determined after exposure. The average duration of exposure of miners was 37 (6-82) days. Urinary mercury significantly increased during exposure. Immunoelectrophoretic changes in the composition of urinary proteins occurred after exposure in 22 of 45 miners, of whom 15 showed high molecular weight (HMW) pattern of urinary proteins and seven showed low molecular weight (LMW) pattern. Only a slight increase in the urinary alpha 1-microglobulin concentration and NAG activity was found in miners with the LMW pattern of urinary proteins. The results point to a slight glomerular and tubular damage in a significant proportion of exposed miners with increased absorption of mercury vapour. PMID- 11276965 TI - The changing pattern of poisoning with psychoactive drugs in Croatia. AB - The aim of this study was to analyse the frequency of poisoning with psychoactive drugs (benzodiazepines, antidepressants and neuroleptics) over the last 15 years in Croatia. The analysis was based on poisoning incidents reported over the phone (hot line) to the Zagreb Poison Control Center and included two periods: 1985 1991 (period I) and 1992-1999 (period II). The data were analysed separately for children and adults. Each phone call was counted as one poisoning incident. Child poisoning with neuroleptics was significantly higher in period II than in period I and so was the adult poisoning with antidepressants, amytriptyline, and combined psychoactive drugs. The frequency of total psychoactive drug poisoning was significantly higher in adults than in children in both periods. From 1992, the frequency of adult poisoning with antidepressants considerably increased as one of the many consequences of war-related stress. The results indicate a need for careful psychiatric evaluation and more critical use of antidepressants in affected individuals. PMID- 11276966 TI - Analysis of drugs of abuse in urine by gas chromatography/mass spectrometry: experience and application. AB - This paper describes quantitative methods for determination of urinary drugs/metabolites. The analysis included indicators of opiate (morphine, codeine, 6-monoacetylmorphine) and methadone (methadone) consumption, indicator of marihuana/hashish consumption (11-nor-9-tetrahydrocannabinol-9-carboxylic acid), indicators of cocaine consumption (cocaine, benzoylecgonine, and ecgonine methyl ester) and of amphetamines consumption (amphetamine, methamphetamine, 3,4 methylenedioxyamphetamine, 3,4-methylenedioxymethamphetamine, and 3,4 methylenedioxyethylamphetamine). The methods included solid-phase extraction of urine, concentration of eluent, derivatisation, and quantitative analysis by gas chromatography/mass spectrometry (GC/MS) on a capillary column in the electron impact and selected ion monitoring (SIM) mode. Sensitivity, reproducibility, and accuracy were determined for all analytes (limit of detection between 3 and 12 ng/ml, precision < 10%, accuracy > 92%). The accuracy was checked through analysis of standard reference materials and participation in an international quality assessment programme. The methods were used in the analysis of spot urine samples of 60 subjects suspected of drug abuse. Negative findings indicated several disadvantages of urine as a biological sample. PMID- 11276967 TI - Household chemicals--common cause of unintentional poisoning. AB - Of 4736 poisoning incidents registered in the Poison Control Centre in Zagreb from 1985 to 1999, household chemicals caused 23%. In the group of cleaning products, 11% of poisoning incidents were caused by corrosives, 9% by liquid detergents and 4% by hypochlorite. Organic solvents caused 18% of household chemical poisonings; among them gasoline and thinners were the most frequent. Cosmetics were responsible for 7% of poisoning incidents; the most frequent were hair shampoo, hydrogen peroxide, and acetone. In the group of other chemicals, the most common were ingestion of thermometer mercury and of silica gel, while poisonings with highly toxic antifreeze, mothballs, or liquid fertilisers were rare. Ingestion or other exposure to household chemicals often caused excessive concern and therapeutic measures. It is therefore advisable to consult a Poison Control Centre in order to get proper information about the composition of a chemical and toxicity of a product. PMID- 11276968 TI - [Hair--a biological source in drug analysis]. AB - Hair testing for drugs of abuse has the advantage of prolonged detection over blood, saliva, or urine analysis and is a useful diagnostic complement to them. This paper describes the main characteristics of hair as a biological sample and the mechanisms of incorporation, retention, and loss of drugs of abuse. The overview is confined to the most common procedures for determination of drugs of abuse, that is, those which involve gas chromatography-mass spectrometry (GC-MS). Each procedure basically includes the same steps: hair washing (decontamination), cutting, extraction/digestion, clean-up, and derivatisation. Although the analysis of hair has been worked out in detail, interpretation of findings of drugs of abuse in hair is complex. The content of hair is not uniform all along; a drug does not necessarily enter through the root; there are differences in hair growth; cosmetic treatment, purity of the taken drug, and the hair colour may affect the concentration of drugs of abuse in hair. The interpretation of findings must take all that into account. Positive findings of drugs of abuse in hair reflect chronic/recurring consumption but do not positively establish the quantity or time of consumption. PMID- 11276969 TI - Fear of flying: an overview. AB - Flying gives the aviator a sense of power and control. Aviators challenge their own skills and test the physical limitations of the plane. They must maintain the balance between fear and joy, sacrifice and love, and risks and rewards of flying in order to continue to fly without reservation. Flying is dangerous. The danger is both real and symbolic, generating fears and anxiety. Interpretation of fear of flying spans from the psychoanalytic-endogenous on one side to the behaviouristic-exogenous on the other side. Unless strictly understood, both models should be taken to consider the interaction between the endogenous and exogenous factors. The therapy of the fear of flying is based on the correlation between the symptoms and underlying dynamics. The prognosis depends on the ability to work through those psychodynamic conflicts. Aviators may continue to fly if the symptoms are minor and the motivation to resolve conflicts is high. PMID- 11276970 TI - [Use of telepathology in histopathologic diagnosis]. PMID- 11276971 TI - [Use of etoposide, ifosfamide and cisplatin (EIP) for treatment of inoperable non small cell lung cancer]. AB - Recent studies have pointed out that chemotherapy can prolong life in advanced inoperable cancer patients. A clinical study to evaluate response and toxicity of the combination of etoposide, ifosfamide and cisplatin (EIP) in the treatment of inoperable non-small cell lung cancer was performed. 25 patients entered the study. Treatment consisted of etoposide 120 mg/m2 given i.v. on days 1-3, ifosfamide 1.5 g/m2 given i.v. on days 1-5 with mesna protection and cisplatin 20 mg/m2 given i.v. on days 1-5. Cycles were repeated every 4 weeks for a maximum of 6 in responders. 16 (64%) patients responded to treatment, 13 (52%) reached partial and 3 (12%) complete remission. In two recurrent cases second remission was achieved after reinstitution of the EIP regimen. Median survival time was 13 months (range 7-48 months) for responders and 5 months (range 2-11 months) for non-responders. Overall treatment was well tolerated with granulocytopenia being the most frequent toxicity. The results are encouraging for further investigations. Application of higher doses of ifosfamide with colony stimulating factors protection is planned. PMID- 11276972 TI - [Lung cancer--differences of incidence between the sexes]. AB - During the last decade increasing incidence of lung cancer among women have been observed in Poland. The aim of the study was to demonstrate differences among men and women with lung cancer. Lung cancer was diagnosed in 785 female and 4619 male in 1995 in Pulmonary Outpatients Departments. Women were younger than man when all histologic types of lung cancer were analysed (59.7 vs 61.9 years p. < 0.001). Particularly younger subjects were those with adenocarcinoma and small cell lung cancer (56.9 and 57.4 years for women and for men respectively 60.2 and 59.6 years, p < 0.001). Although squamous lung cancer was the most prevalent histological type among men (43.7%) and women (24.7%), about two times higher percentage of men had this neoplasm (p. < 0.001). Adenocarcinoma (18% vs 6.6%, p. < 0.001) and small cell lung cancer (18.5% vs 15.5% p. < 0.001) were prevalent in significantly higher percentage among female than male. Nonsmokers were more frequently noticed among women then men (20.4% vs. 1.9%, p. < 0.001), particularly those with adenocarcinoma. Also women smoked less intensively (33.6 pack/years vs. 42.3 pack/years, p < 0.001) except those with squamous cancer. The higher incidence of cancer was observed among mothers (7% vs 3.8% p. < 0.001) and fathers (7.1% vs 5.6%, p. < 0.001) of women than men with lung cancer. PMID- 11276973 TI - [The role of sex as a prognostic factor in lung cancer]. AB - The aim of this study was to demonstrate the prognostic role of the gender. Lung cancer was diagnosed in 785 female and 4619 male registered in Pulmonary Outpatients Departments in 1995. Women were younger than man when all histologic types of lung cancer were analysed (59.7 vs 61.9 years of age p. < 0.001), particularly those with adenocarcinoma(56.9 vs 60.2 years of age, p. < 0.012) and small cell lung cancer (57.4 vs 59.6 years of age, p. < 0.001). Although squamous lung cancer was the most prevalent among men (43.7%) and women (24.7%), about two times higher percentage of men had this neoplasm. Adenocarcinoma (18% vs 6.6%, p. < 0.001) and small cell lung cancer (28.5% vs 15.5% p. < 0.001) were prevalent in significantly higher percentage among female than male. Women were treated more aggressively by surgery (17.1% vs 14.1%, p. = 0.04) but similar percentage of men and women received radiotherapy, chemotherapy and multimodality treatment. Women more frequently survived one year (43% vs 35.7%, p. < 0.04). Significant and independent negative prognostic factors were: gender (RR-1.17 for men), age older than 50 age (RR-1.2), bed performance status (RR-3.28), disseminated disease (RR 2.78) small cell histological type of cancer (RR-1.21) and nonsurgical therapy (RR-3.29). PMID- 11276974 TI - [Clinical significance of pattern types in high resolution computed tomography images of patients with idiopathic pulmonary fibrosis]. AB - The aim of the study was to assess significance of two types of HRCT pattern in patients with idiopathic pulmonary fibrosis (IPF), corresponding to usual interstitial pneumonia. The study population consisted of 34 patients, 11 women and 23 men, mean age 64.5 +/- 10.5 years. The patients were divided into two groups according to HRCT appearance: Group A--reticular pattern with some ground glass attenuation (30 patients); group B--reticular pattern only (4 patients). Age, level of dyspnea, pulmonary function tests were similar in both groups. Longer history of dyspnea and more frequent finger clubbing (all patients) were found in Group B. 30 patients were followed-up for at least 12 months. In this period 4 out of 27 patients died in Group A, and 2 out of 3 patients died in Group B (p < 0.05). CONCLUSION: HRCT showing reticular pattern only corresponds to late phase of IPF and is connected with very short survival. PMID- 11276975 TI - [Pulmonary alveolar proteinosis]. AB - Pulmonary alveolar proteinosis (PAP) is a rare disease characterised by the accumulation of proteinaceous material within alveoli. In order to evaluate the clinical features and the course of PAP we reviewed 7 cases (2F/5M) diagnosed during a 11-year period (1989-1999). The mean age of patients was 40.7 +/- 11.2 years. Diagnosis was obtained by open lung biopsy in all cases. Clinical findings included dyspnea (43%), cough (28%) and crackles (28%). Lung function tests were normal in 5 cases and showed a moderate restrictive pattern in 1 and mild airflow obstruction in 1. Three patients had reduced Dlco (mean was 63% of predicted). Four patients had hypoxemia at rest. Chest X-ray revealed bilateral alveolar opacities (71%), involving perihilar areas and lower lobes. HRCT scans demonstrated diffuse ground glass opacities (83%) with interlobular septa thickening (50%). Three patients were treated with repeated segmental BAL (2 improved). The spontaneous partial remission occurred in 4. PMID- 11276976 TI - [A case of traumatic rupture of the median bronchus]. AB - The case of rupture of median bronchus after road accident was described. Early diagnose and treatment let us to obtain complete reconstruction of bronchial tree and preserve two lung lobes. PMID- 11276977 TI - [Cytomegalic disease as a cause of disseminated lung lesions in an immunosuppressed patient]. AB - A case of disseminated lesions in the lungs was diagnosed just during autopsy as cytomegalic infection. 54-year old patient treated since 10 years because of lymphoplasmacytoid lymphoma of low malignancy was admitted to Institute of Tuberculosis with suspicion of miliary tuberculosis. The high temperature, pemphigus--like skin lesions and disseminated lesions in the chest X-ray appeared immediately after succeeding chemotherapy. Tuberculous bacilli. Aspergillus fumigatus and Pneumocystis carinii were not found in examined materials (BALF, blood, urine, skin and mucous lesions). Patient died after 3 days and typical lesions of cytomegalovirus infection were found only after autopsy in macroscopic picture of the lungs. PMID- 11276979 TI - [The effect of probiotics on the immune system]. PMID- 11276978 TI - [Theophylline--mechanism of action in the respiratory tract]. PMID- 11276980 TI - [Sensitivity to birch pollen--under-appreciated etiology of atopic asthma in towns]. AB - Author presents some epidemiological data of birch pollen allergy. Pathogenesis and also crossreactivity of such allergy is discussed, especially concerning allergy to fruits. Some data about various phenotypes of birch tree are presented. Finally author gives some clinical data of 30 observed cases of young adults whose birch pollen allergy in 63% manifested itself as bronchial asthma. PMID- 11276982 TI - The relevance of a nonword repetition task to assess phonological short-term memory in individuals with Down syndrome. AB - Phonological short-term memory capacity is generally measured with a word span task or a digit span task. Another way to measure it is to use a nonword repetition task. Gathercole and Adams (1993) claimed that this procedure can be used with children as young as two-years old. It seems that in normally developing children the quality of nonword repetition is influenced both by the length of nonwords and by the degree of wordlikeness. Can the phonological short term memory of individuals with Down syndrome be assessed with a nonword repetition task? In order to answer this question, we decided to replicate Gathercole and collaborators' experiments (1991,1993) but with individuals with Down syndrome. The quality of nonword repetition in individuals with Down syndrome is, as in normally developing children, influenced both by the length of nonwords and by their degree of wordlikeness. Furthermore, our results seem to confirm the hypothesis which states that nonwords are temporarily stored in the phonological short-term memory system. As this system has a limited capacity, both normally developing children and people with Down syndrome recall more short nonwords than long nonwords. In conclusion, nonword repetition is a reliable task with which to assess phonological short-term memory in individuals with Down syndrome as well as in normally developing children. PMID- 11276981 TI - Down syndrome and the phonological loop: the evidence for, and importance of, a specific verbal short-term memory deficit. AB - Individuals with Down syndrome are thought to perform poorly on tests of verbal short-term memory, such as measures of word span or digit span. This review critically examines the evidence for a specific deficit in verbal short-term memory in Down syndrome, and outlines a range of possible explanations for such a deficit. The potential implications of a verbal short-term memory impairment for broader aspects of development are outlined, in particular with respect to vocabulary development. Possible intervention strategies, which might improve verbal short-term memory performance in Down syndrome are also considered. However, we argue that further research is needed to fully clarify the nature of a verbal short-term memory deficit in Down syndrome, before the merits of these various intervention approaches can be properly evaluated. PMID- 11276983 TI - Learning to count: a difficult task? AB - This article is concerned with the acquisition of counting skills in pupils with Down syndrome. Data from a larger survey of pupils with severe learning difficulties is explored to investigate the types of errors children make at the earliest stages of learning to count. The pattern of responding was consistent with the view that children with Down syndrome have particular difficulties in tasks utilising auditory sequential memory, in this case learning the number string. The practical implications of these findings are discussed. PMID- 11276985 TI - Verbal short-term memory function. PMID- 11276984 TI - Getting in and staying there: children with Down syndrome in mainstream schools. AB - The proportion of children with Down syndrome in mainstream schools, compared to special schools, has been increasing over the last decade; this is due both to more children going into mainstream schools at five or six and to more children staying in mainstream schools for increasing lengths of time, not uncommonly throughout their school careers. There are, however, wide variations between Local Education Authorities, which is attributed mainly to differing implementation of inclusion policies. Data is drawn together from a number of sources (both previously published and unpublished) which describe some of the processes which take place in making initial placements in mainstream schools, maintaining those placements and transferring out of mainstream schools. Commitment of staff to meeting children's special needs rather than matters relating to the curriculum seem to be of paramount importance both at home and abroad in successful mainstream placements. PMID- 11276986 TI - Lowdown on laminar flow. PMID- 11276987 TI - Aspects of smoke control. PMID- 11276988 TI - Special focus on decontamination. PMID- 11276989 TI - Signs for the times. PMID- 11276990 TI - A handle on door history. PMID- 11276991 TI - Access issues need strategic approach. PMID- 11276992 TI - Tuning in to telemedicine. PMID- 11276993 TI - The future is already here. PMID- 11276994 TI - Human lessons from the M&A (merger & acquisition) wars. PMID- 11276995 TI - Discover the power of wellness. PMID- 11276996 TI - Protecting women's health. PMID- 11276997 TI - Top marks for employee training. PMID- 11276998 TI - Science and access at Merrill Lynch. PMID- 11276999 TI - DataWatch. Voters still put health over tax cuts. PMID- 11277000 TI - Michael and Rose Assarian Cancer Center, Novi, Michigan. PMID- 11277001 TI - Cardiac comprehensive critical care, Methodist Hospital, Indianapolis. PMID- 11277002 TI - Luke Waites Child Development Center, Texas Scottish Rite Hospital for Children, Dallas. PMID- 11277003 TI - Safety in numbers. The five most frequently cited hospital OSHA violations and how to avoid them. PMID- 11277004 TI - Wireless world. Facility managers are behind the curve on handheld wireless devices, but a few leaders are showing the way. PMID- 11277005 TI - Something in the air. Groups turn their attention to Legionella control. PMID- 11277006 TI - Be chemically correct! Advice on managing your facility's cleaning chemicals. PMID- 11277008 TI - A $25 million IOU. PMID- 11277007 TI - My patients got an HMO to take me back. PMID- 11277009 TI - I could not let this patient die. PMID- 11277010 TI - PPMs: going ... going.... PMID- 11277011 TI - Your obligation to treat patients referred by the ER. PMID- 11277012 TI - A new day dawns ... when patients buy their own health care. PMID- 11277013 TI - Value-based partnering in healthcare: a framework for analysis. AB - Value-based partnering is designed to move the healthcare system beyond cost based competition. It recognizes that the healthcare "product" is not a commodity and that much of the value in the system comes from relationships between and among four stakeholders: consumers, providers, health plans, and employers. Given the difficulty of measuring such benefits as quality of care, improved health status, and increased employee productivity, stakeholders within the system traditionally have focused on easily measurable financial considerations such as premium rates. This focus has led to a system that defines relationships in purely financial terms. In contrast, the value-based partnering model presented in this article recognizes the range of factors that stakeholders consider in their relationships with each other. This approach has the potential to change the nature of competition and presents opportunities for those organizations that can effectively partner with other stakeholders and demonstrate value, rather than just lower cost. Moreover, by recognizing the interdependencies among stakeholder groups, the approach creates a strategic reason for employers, health plans, providers, and consumers to exchange information and create long-term alliances. PMID- 11277014 TI - Minimizing the risk of discriminatory suits when terminating poor performers. PMID- 11277015 TI - Interview with Zelda Geyer-Sylvia, Executive Director of M-Care. Interview by Kyle L. Grazier. PMID- 11277016 TI - Customer satisfaction: six strategies for continuous improvement. PMID- 11277017 TI - A millennium mindset: the long boom. PMID- 11277018 TI - Exploring dual commitment among physician executives in managed care. AB - The growth of a medical management specialty is a significant event associated with managed care. Physician executives are lauded for their potential in bridging the clinical and managerial realms. They also serve as a countervailing force to help the medical profession and patients maintain a strong voice in healthcare decision making at the strategic level. However, little is known about their work loyalties. These attitudes are important to explore because they speak to whose interests physician executives consider and represent in their everyday management roles. If physician executives are to maximize their effectiveness in the healthcare workplace, both physicians and organizations must view them as credible sources of authority. This study examines organizational and professional commitment among a national sample of physician executives employed in managed care settings. Data used for the analysis come from a national survey conducted through the American College of Physician Executives in 1996. The findings support the notion that physician executives can and do express simultaneous loyalty to organizational and professional interests. This dual commitment is related to other work attitudes that contribute to success in the management role. In addition, it appears that situational factors increase the chances for dual commitment. These factors derive from a favorable work environment that includes both organizational and professional socialization in the management role. The results of the study are useful in specifying the training and socialization needs of physicians who wish to do management work. They also provide a rationale for collaboration between healthcare organizations and rank-and-file physicians aimed at cultivating physician executives who are credible leaders within the healthcare system. PMID- 11277019 TI - [Respiratory insufficiency caused by extra-pulmonary disease]. PMID- 11277020 TI - [Artificial respiration for acute respiratory insufficiency--state of the art]. PMID- 11277021 TI - [Drug therapy in acute and chronic respiratory insufficiency]. PMID- 11277022 TI - [Therapeutic options in chronic respiratory insufficiency]. PMID- 11277023 TI - [Respiratory insufficiency. Epidemiology, economic burden and health care facilities needed as exemplified by COPD]. PMID- 11277024 TI - [Acute kidney failure]. PMID- 11277025 TI - [Uterine tumor, HCG excess and hyperthyroidism in a 51-year old woman]. PMID- 11277026 TI - [80-year old patient with antibiotic-resistant fever and cholestasis]. PMID- 11277027 TI - [Facial skin manifestations]. PMID- 11277028 TI - [Therapy with preferential and specific COX-2 inhibitors]. PMID- 11277029 TI - [Anorectal continence organ. Comment from a surgical view point of C.Scheurlen, M.Neubrand, M. Kaminski and T.Sauerbruch, Internist (2000)41:1213-1234]. PMID- 11277030 TI - Physicians band together to share I.T. services. PMID- 11277031 TI - Internet eases negotiations for FDA, drug firms. PMID- 11277032 TI - Clinic sets sights on the Internet. PMID- 11277033 TI - For specialists, it's not just the fax. PMID- 11277034 TI - Online managed care transactions gain some ground. PMID- 11277035 TI - Caregivers talk about speech technology. PMID- 11277036 TI - Annual HIMSS survey: HIPAA awareness is growing. PMID- 11277037 TI - E-mail under lock and key. PMID- 11277038 TI - CIOs fall for PDAs PDQ. PMID- 11277039 TI - Medicare. Thompson says Part A funds must pay for Part A. PMID- 11277040 TI - Perspectives. Medicare, insurers, big employers begin to eye practice variations. PMID- 11277041 TI - Just the medicine. Cheaper AIDS drugs. PMID- 11277042 TI - Psychosis and punishment. Should the mentally ill be drugged so they can face execution? PMID- 11277043 TI - 'I can't stress enough how devastating...'. are our borders safe against foot-and mouth? PMID- 11277044 TI - The art of forgetting. With effort, people can purge their memories. PMID- 11277045 TI - For trials of new therapies, cast your net on the Web. PMID- 11277046 TI - Are sealants doing more than preventing caries? AB - Dental sealants are among some of dentistry's most effective tools for preventing tooth decay when applied to pediatric patients' teeth. Yet, a quiet controversy is mounting about some of the chemicals that may be leaching from these powerhouses of prevention. Some scientists hint that sealants may require further study to rule out possible links between some of their components and breast cancer development, prostate enlargement and some as-yet-unknown biological effects. While sealants' preventive power ranks with fluoride, even a remote possibility that they may affect patients' systemic health creates concern. PMID- 11277047 TI - Embracing technology. How (and why) to computerize your office. PMID- 11277048 TI - Northwestern's last class. Interview by Julia A. Jacob. PMID- 11277049 TI - Advertising or public relations: what's right for your practice? PMID- 11277050 TI - Five tips for choosing a dental laboratory. Interview by Karen A. Gomolka. PMID- 11277051 TI - Simple steps to prevent bad breath. PMID- 11277052 TI - The stir over a toothless child and an orange. PMID- 11277053 TI - Urologic laparoscopy: the future is now. AB - Recent interest in urologic laparoscopy is a welcome renaissance of minimally invasive procedures that provide superior benefit to patients. It behooves the urologist to avoid complacency and embrace the resurgence of laparoscopy. Dedicated pursuit and perseverance will facilitate the evolution and future of urologic laparoscopy. PMID- 11277054 TI - Laparoscopic retroperitoneal lymph node dissection. AB - Laparoscopic RPLND for clinical stage I testicular tumors is superior to open surgery in terms of efficiency, morbidity, and costs and has equal diagnostic accuracy. A quality of life study has shown that patients prefer surgery to chemotherapy and laparoscopy to open surgery. Laparoscopic RPLND is also feasible after chemotherapy, and the results are as good as those achieved in clinical stage I disease. Patients with stage IIb lesions can be spared the third and fourth cycle of chemotherapy if diagnostic laparoscopic RPLND is performed to exclude active tumor or remove mature teratoma. Laparoscopic removal of residual tumors following conventional chemotherapy for stage IIb lesions can be performed with minimal morbidity. The long and steep learning curve remains the biggest obstacle in laparoscopic RPLND. Most centers specializing in the treatment of testicular cancer are not trained in laparoscopic surgery and vice versa; however, the growing importance of laparoscopy for oncologic indications should help to overcome this problem in the near future. PMID- 11277056 TI - Laparoscopic live donor nephrectomy. AB - Laparoscopic donor nephrectomy offers numerous advantages when compared with the traditional open approach. For the donor, it has resulted in a shorter hospital stay, fewer postoperative analgesic requirements, earlier return to activities of daily living and employment, and decreased financial loss owing to absence from the workforce. For the recipient, the procedure does not adversely impact on allograft function, graft survival, or patient survival. PMID- 11277055 TI - Laparoscopic management of renal cystic disease. AB - Laparoscopic management of renal cystic disease is a highly effective, safe, and minimally invasive alternative to open surgery and antegrade or retrograde endoscopic procedures. Simple renal cysts can be accessed either transperitoneally or retroperitoneally. Almost all studies of the laparoscopic approach have demonstrated great satisfaction in terms of efficacy, minimal complications, operative time, minimal blood loss, hospital stay, recuperation, and cosmesis over other methods of treating renal cysts. Laparoscopic unroofing of peripelvic cysts is more challenging owing to their proximity to hilar vessels and the collecting system. Such surgery should be considered an advanced laparoscopic procedure. Access may be achieved either transperitoneally or retroperitoneoscopically. The basic principle of adequate exposure is essential for effective treatment. If the cyst is not completely excised, the surgeon must fulgurate the edge and tack perirenal fat in the residual cyst cavity to prevent recurrence and facilitate drainage. Laparoscopic evaluation of complex cysts seems to be sound. The results are promising, and follow-up does not show any increase in peritoneal seeding, tract recurrence, or distant metastases in the small number of neoplasms diagnosed at laparoscopy. Nevertheless, more studies are required with long-term follow-up. Bosniak type IV renal cysts or malignancy in renal cysts can be managed by laparoscopic radical nephrectomy with either access. Laparoscopic cyst marsupialization in patients with ADPKD is the latest emerging indication for laparoscopy in renal cystic disease. This procedure not only effectively reduces pain in some patients but also improves hypertension and stabilizes renal function, delaying renal replacement therapy. Long-term follow up and further evaluation are needed. PMID- 11277057 TI - Retroperitoneal laparoscopic nephropexy. AB - Laparoscopic nephropexy is a suitable and clinically established procedure for the treatment of symptomatic nephroptosis. The availability of a minimally invasive therapy can facilitate decisions regarding the indication after careful selection of patients. PMID- 11277058 TI - Laparoscopic management of female urinary incontinence. AB - Recent technologic developments in laparoscopic reconstructive surgery have generated an interest in the laparoscopic approach to bladder neck suspension. There have been numerous descriptions of a variety of techniques for the laparoscopic approach to bladder neck suspension. Initial reports seemed to suggest satisfactory rates of improvement in the stress urinary incontinence of these patients. Long-term follow-up has shown that although this minimally invasive approach to the management of stress urinary incontinence is associated with a short duration of urinary diversion, minimal postoperative discomfort, and a quick return to a productive life, the durability of the cure has failed the test of time. The laparoscopic bladder neck suspension in 3 and 4 years follow-up has achieved a success rate of only 30%, with a mean time to failure of 18 months. Any new surgical technique applied to the management of stress urinary incontinence must have a minimum of 2 years mean follow-up to determine its true clinical efficacy and durability. PMID- 11277059 TI - Newer techniques in intracorporeal tissue approximation: suturing, tissue adhesives, and microclips. AB - Great strides have been made in the discovery of alternative tissue approximation techniques for use in laparoscopy. Although none of the techniques have eliminated the need for suturing laparoscopically, their potential in achieving this end is promising. When an ideal approximation technique is discovered that is easy to use, safe, and reliably able to hold tissue together laparoscopically, laparoscopic reconstructive surgery should become less formidable and more appealing to urologists. PMID- 11277060 TI - Promontofixation for the treatment of prolapse. AB - Genital prolapse is a common problem in women. The wide variety of surgical techniques used to treat this problem demonstrate how difficult it is to manage. Laparoscopic surgery offers a new approach. It allows a good view of the anterior and posterior compartments so that a global approach for the prolapse is possible by the same surgical route. Traditional promontofixation can be combined with a new approach to the posterior compartment. Laparoscopic promontofixation through installation of an intervesicouterine prosthesis for the treatment of hysterocele and cystocele is associated with paravaginal repair of lateral defects and a Burch anterior colposuspension for urinary stress incontinence. When combined with laparoscopic treatment of rectocele by myorrhaphy and reinforcement of the fascia by means of a prosthesis, it provides a complete range of treatment for all types of feminine prolapse. After 20 years of experience through laparotomy, promontofixation using a triangle has been carried out by laparoscopy at the authors' center since 1991 in an attempt to eliminate the cystocele by solidly anchoring the uterus and bladder floor to the promontory. This laparoscopic technique follows the usual steps for pelvic prolapse repair: 1. Total or subtotal hysterectomy or suspension of the uterus is performed in such a way that it returns to normal physiologic position, and a solid subvesical floor is created. 2. The physiologic axis of the vagina is restored by creating a strong, low posterior point of support and by performing culdoplasty. 3. Evident or latent stress incontinence is treated. It would be pointless to treat the hysterocele on its own because, once the prolapse has been cured, the subvesical mass will disappear and allow urinary incontinence to appear. 4. Reconstruction of the posterior rectovaginal support structures seems to be mandatory and is carried out in almost all cases. The first phase of the laparoscopic approach to pelvic prolapse allowed the authors to explore the technical aspects. Several approaches are possible by laparoscopy. Herein, the authors report 8 years of technical research and assessment. This experience confirms the tremendous potential of laparoscopic surgery for the treatment of all aspects of this pathology by the same route. Stress incontinence, cystocele, hysterocele, rectocele, or enterocele can be treated. The operative time is longer than with the open route, and the surgeon must be highly experienced. Based on their experience, the authors are discovering new concepts. More data are required before a conclusion can be drawn concerning this promising new approach. PMID- 11277061 TI - Use of bowel in laparoscopic urology. AB - Minimally invasive urology is a rapidly expanding field. What once was thought technically impossible is now becoming a reality, especially with the advent of intracorporeal stapling and automated suturing devices. Laparoscopic assistance and pure laparoscopy improve convalescence and cosmesis in comparison with open surgical procedures. Minimally invasive continent urinary stomas, ACE procedures, bladder augmentation, urinary diversion, and urinary undiversion have all been described in clinical practice. Continent urinary diversions and ileal bladder augmentations are being developed. Eventually, even the most challenging urologic procedures will be performed in a minimally invasive manner. PMID- 11277062 TI - Hand-assisted laparoscopic renal surgery. AB - The introduction of hand-assisted laparoscopic surgery (HALS) adds another dimension to standard laparoscopy, and particularly benefits the urologic laparoscopist who performs laparoscopic renal procedures. Hand assistance shortens the initial learning curve for laparoscopic nephrectomy and creates alternatives for more experienced laparoscopists. HALS can be performed on any kidney suitable for intact removal, and is well suited to laparoscopic donor nephrectomy, radical nephroureterectomy, and radical nephrectomy with intact removal. The authors' technique, early experience, and indications for hand assistance are described in detail. PMID- 11277063 TI - Telesurgery. Remote monitoring and assistance during laparoscopy. AB - In comparison to open surgery, laparoscopy results in less postoperative pain, shorter hospitalization, more rapid return to the work force, a better cosmetic result, and a lower incidence of postoperative intra-abdominal adhesions. These advantages are indisputable when comparing large series for cholecystectomy and smaller series for pelvic lymph node dissection, nephrectomy, and bladder neck suspension in experienced hands. Urologists have an obligation to explore the application of these methods to urologic disease and to adjust the standard of care accordingly. Several barriers to the expansion of urologic laparoscopic surgery exist. The experience in extirpative and reconstructive urologic procedures is limited when compared with the data on cholecystectomy. These procedures are technically complex and demand advanced laparoscopic skills and familiarity with laparoscopic anatomy. The steep learning curve translates into long operative times and an unacceptably high rate of complications for inexperienced laparoscopic surgeons. Most practicing urologists have no formal training in advanced laparoscopy, and no formal credentialing guidelines exist. Telesurgical technology may provide one solution to this problem. Through telesurgical mentoring, less experienced surgeons with basic laparoscopic skills could receive training in advanced techniques from a world expert without the need for travel. These systems could also be used to proctor laparoscopic cases for credentialing purposes and to provide a more uniform standard of care. This review has outlined some of the exciting progress made in the field of telesurgery over the past 10 years and described some of the technical and legal obstacles that remain to be surmounted. During the 1990s, urologists were at the forefront of innovation in remote telepresence surgery. As the scope of minimally invasive urologic surgery expands during the first few decades of the twenty first century, telesurgical mentoring should have an increasingly important role. PMID- 11277064 TI - Laparoscopic radical prostatectomy: assessment after 240 procedures. AB - Radical prostatectomy can be successfully performed by transperitoneal laparoscopy by a urologic team experienced in laparoscopy and radical prostatectomy. Operative and postoperative morbidity rates are low. Postoperative pain is minimal, allowing reduction of the length of hospital stay. The oncologic results seem satisfactory based on short-term follow-up. The improvement of the quality of intraoperative vision related to magnification of the image allows a more precise procedure. This subjective improvement of the quality of dissection should reduce the usual functional sequelae of conventional radical prostatectomy, such as incontinence and impotence. This finding needs to be confirmed by a larger series of patients with longer follow-up. Laparoscopic radical prostatectomy is now performed routinely and is proposed as a first-line surgical treatment for localized prostatic cancer at the authors' center. PMID- 11277065 TI - Laparoscopic management of the undescended testicle. AB - The role of laparoscopy with regards to the undescended testicle has been hotly debated since the late 1970s and early 1980s when it was realized that the abdominal testicle could be visualized with the laparoscope. Many enthusiastically embraced the laparoscope for diagnosis and localization of the impalpable undescended testicle, whereas others staunchly maintained that laparoscopy was overly invasive and really facilitated little with regards to orchidopexy. Diagnostic laparoscopy, when compared with the other modalities, holds its own well with regards to accuracy and efficacy. In the early 1990s, the role of laparoscopy expanded to include performance of orchidopexy. It would be naive to believe that the debate cooled with the evolution of laparoscopic orchidopexy. Over the years, since introduction of the operation, there have been many institutions that have examined the role of laparoscopic orchidopexy quite critically. In many people's minds, laparoscopic orchidopexy is a clear competitor to open orchidopexy for the impalpable undescended abdominal testicle; in a few people's minds, it has become the gold standard. PMID- 11277066 TI - Laparoscopic management of intersexual states. AB - Laparoscopy has gained acceptance as the ideal method of surgical treatment of the internal [figure: see text] genital organs in patients with intersex disorders. The intersexual states for which laparoscopy is needed are female and male pseudohermaphroditism, true hermaphroditism, and Turner's syndrome. In these patients, the indications for laparoscopy are the removal of normal gonads and ductal structures that are contrary to the assigned gender and the removal of dysgenetic gonads that are nonfunctional and that present potential for malignancy. In addition to being a minimally invasive surgery, one of the main advantages of this method is the lack of scars, a fact much appreciated by patients and their parents. Generally, gonadectomy is a straightforward operation because the gonads present with an accessible pedicle. Laparoscopic orchidopexy has been standardized and can be performed in patients in whom the testis must be relocated to the scrotum. The removal of ductal structures is also easily performed in most cases, whereas hysterectomy with resection of the vagina may present some difficulties owing to the location of these structures. In patients with a long vaginal component of the urogenital sinus, the distal segment must be removed by a retrograde perineal access, usually performed simultaneously with genitoplasty. Endocrinologists must be aware of the application of this method of treatment in intersex patients, and urologists proficient in laparoscopic techniques must extend their field of work in this area. PMID- 11277067 TI - Laparoscopic Anderson-Hynes dismembered pyeloplasty in children using needlescopic instrumentation. AB - The technique of needlescopic Anderson-Hynes dismembered pyeloplasty is discussed. The recent development of robust 3-mm needlescopic instruments designed to perform dismembered pyeloplasties has added a new dimension to laparoscopic suturing and will significantly enhance the ability to perform microanastomosis--the limiting factor preventing many surgeons from performing this technically demanding operation. The use of such fine instrumentation also reduces the size of instrument cannulae necessary, further reducing the postoperative pain and results in spectacular postoperative cosmetic appearance. PMID- 11277068 TI - Advances in camera, video, and imaging technologies in laparoscopy. AB - The introduction of technological advances, such as HDTV, three-dimensional laparoscopy, and further miniaturization of high-resolution digital video cameras, will allow significantly enhanced opportunities for laparoscopic surgical proficiency and further broadening of laparoscopic applications in urology. These enhancements, coupled with the recent advances in telemedicine and surgical simulation, will improve laparoscopic training and skill acquisition, decrease operative times and costs, minimize morbidity, and improve overall patient care. PMID- 11277069 TI - Laparoscopic nephrectomy and nephroureterectomy in the pediatric patient. AB - The pediatric laparoscopic nephrectomy/nephroureterectomy literature is reviewed. The authors' method for pediatric nephrectomy/nephroureterectomy is presented, as well as the clinical experience from Washington University. PMID- 11277070 TI - Transperitoneal laparoscopic adrenalectomy. AB - Laparoscopic adrenalectomy has been demonstrated by many institutions to be a safe and effective approach to benign adrenal lesions. As surgical skills improve, it will most likely be comparable with open adrenalectomy in overall cost. Although it is technically demanding and takes slightly longer than open surgery, the benefits to the patient in terms of hospital stay, convalescence, and cosmetic results have been conclusively reported. The decision to use a transperitoneal or retroperitoneal access is mostly a function of surgeon preference, with similar outcomes by either approach. It is more important that the surgeon be comfortable with the selected approach. Laparoscopic adrenalectomy has become the gold standard for benign adrenal lesions. PMID- 11277071 TI - Needlescopic urology: current status. AB - Technological miniaturization is an emerging trend that encompasses virtually all surgical subspecialties. Minimization of surgical trauma while maximizing surgical cure through gradual progression from maximally invasive to minimally invasive to ultimately noninvasive technologies must be the goal. Needlescopic techniques represent a natural evolution and sophistication of conventional laparoscopy. At the author's center, needlescopic techniques have been used to particular advantage in four specific clinical applications: adrenalectomy, pediatric orchiopexy, detaching the bladder cuff during laparoscopic nephroureterectomy, and as an adjunctive needlescopic port for retraction purposes during conventional laparoscopic surgery. Given the urologist's natural facility with small-diameter endoscopes, the specialty is uniquely positioned to take a leadership position in this emerging field. Increased experience and careful comparisons with conventional laparoscopy will determine the true role of needlescopic technology in the armamentarium of the urologic surgeon. PMID- 11277072 TI - Laparoscopic adrenalectomy: retroperitoneal approach. AB - The most important procedure in retroperitoneoscopic adrenalectomy is en bloc dissection of the perinephric fat, including the upper pole of the kidney and the adrenal gland, from the surrounding muscles (transversus abdominalis, psoas, and diaphragm) just inside Gerota's fascia. It is not recommended that the surgeon directly identify the adrenal gland at the start of the operation. After dividing the perinephric fat between the adrenal gland and kidney, the upper pole of the kidney is exposed, and dissection proceeds along the renal surface. The lateral retroperitoneoscopic approach promises to be a safer and less invasive treatment for patients with small unilateral adrenal tumors. In all cases, careful patient selection and correct choice of the surgical approach based on tumor size, the patient's condition, and the surgeon's skill are vital to avoid complications during laparoscopic adrenalectomy. PMID- 11277073 TI - Laparoscopic surgery for pheochromocytoma. AB - Because of the excessive production of catecholamines, surgery for pheochromocytoma carries a certain risk that can be reduced by accurate preoperative evaluation and by pretreatment with alpha-blockers. The authors' experience and that of other groups suggests that this inherent risk is not enhanced by the laparoscopic approach, and that, following successful surgery, patients benefit from the minimal invasiveness of this technique. Most surgeons recommend the transperitoneal approach that allows direct access to the adrenal vein, facilitating early ligation. This recommendation does not apply to other pathologies of the adrenal gland. Bilateral adrenal tumors are only seen in patients with familial pheochromocytomas. In this setting, adrenal-sparing surgery should be considered, which can also be performed laparoscopically. Previous adrenal surgery is not a contraindication but will render the procedure more difficult. Laparoscopic excision of paragangliomas is also technically feasible. Laparoscopic adrenal surgery for pheochromocytoma is a difficult and demanding task that must be performed by an experienced surgeon in cooperation with a team of specialists including an internist, endocrinologist, and anesthesiologist. PMID- 11277074 TI - The molecular basis of skeletogenesis. Introduction. PMID- 11277075 TI - Genetic control of the cell proliferation-differentiation balance in the developing skull vault: roles of fibroblast growth factor receptor signalling pathways. AB - Activating mutations of genes encoding the transmembrane tyrosine kinase receptors fibroblast growth factor receptors (FGFRs)1-3, and haploinsufficiency of the transcription factor TWIST, cause human craniosynostosis syndromes that typically involve the coronal suture. We have investigated the functional roles of these genes in development of the coronal suture in mouse fetuses, and tested the effects of increasing FGFR signalling by applying exogenous FGF2 to the suture. The results indicate that the proliferation-differentiation balance in normal sutural development involves a gradient of extracellular FGF from the region of differentiation, in which Fgfr1 is expressed, to the sutural mesenchyme, in which low levels of FGF are associated with Fgfr2 expression in osteogenic stem cells. Experimental increase of sutural FGF levels leads to down regulation of Fgfr2, up-regulation of Fgfr1, up-regulation of the osteogenic differentiation gene Osteopontin, and cessation of proliferation. Twist is expressed in the midsutural mesenchyme and is partially co-expressed with Fgfr2, consistent with the possibility that it is involved in maintaining proliferation through regulating transcription of Fgfr2. PMID- 11277076 TI - Craniosynostosis and related limb anomalies. AB - Many genetically determined craniosynostosis syndromes feature limb anomalies, implying that pathways of cranial suture and limb morphogenesis share some identical components. Identification of heterozygous mutations in FGFR1, FGFR2, FGFR3, TWIST and MSX2 in craniosynostosis has focused particular attention on these genes. Here we explore two themes: use of clinical/molecular analysis to provide new clues to pathophysiology and the contrasting effects of loss- and gain-of-function mutations. Apert syndrome is a severe craniosynostosis/syndactyly disorder usually caused by specific substitutions (Ser252Trp or Pro253Arg) in FGFR2. The relative severity of cranial and limb malformations varies in opposite directions for the two mutations, suggesting that these phenotypes arise by different mechanisms. Clinical and biochemical evidence supports a model in which alternative splice forms of FGFR2 mediate these distinct effects. Pro-->Arg substitutions equivalent the Pro253Arg/FGFR2 mutation occur in both FGFR1 and FGFR3, and are also associated with craniosynostosis. This suggests a common pathological mechanism, whereby enhanced affinity for a limited repertoire of tissue-specific ligand(s) excessively prolongs signalling in the cranial suture. The first MSX2 mutation in craniosynostosis was described in 1993 but this remains the only example. We have recently identified three MSX2 mutations associated with a different cranial phenotype, parietal foramina. DNA binding studies show that the craniosynostosis and parietal foramina arise from gain and loss of function, respectively. PMID- 11277077 TI - The parathyroid hormone-related protein and Indian hedgehog feedback loop in the growth plate. AB - Normal development of the growth plate requires coordinated proliferation and differentiation of chondrocytes and osteoblasts. In previous work, we have shown that Indian hedgehog (IHH), produced by prehypertrophic and hypertrophic chondrocytes, stimulates production of parathyroid hormone-related protein (PTHrP) by perichondrial and early chondrocytic cells. PTHrP then maintains chondrocytes in a proliferative, less differentiated state. Because this less differentiated state delays the production of IHH, IHH and PTHrP may participate in a negative feedback loop that synchronizes and determines the pace of differentiation of chondrocytes in the growth plate. To establish the roles of physiological levels of PTHrP and IHH, we have now injected PTH/PTHrP receptor ( /-) embryonic stem (ES) cells into normal blastocysts to generate mice with chimeric growth plates. The PTH/PTHrP receptor cells leave the proliferative cycle and differentiate prematurely in the middle of the normal proliferative columns. The columns of wild-type cells are longer than normal and the adjacent bone collar is also longer than normal. Patterns of gene expression and the use of chimeras using PTH/PTHrP receptor (-/-); IHH (-/-) ES cells suggest that modified patterns of IHH and PTHrP synthesis explain these abnormalities. Thus, IHH is a master regulator of both chondrocyte and osteoblast differentiation. PMID- 11277078 TI - Cartilage matrix resorption in skeletogenesis. AB - Chondrocytes assemble an extracellular matrix in which the relative composition of type IX versus type II collagen and aggrecan changes during assembly. On maturation and differentiation into hypertrophic cells type IX collagen first loses the NC4 globular domain of the alpha 1(IX) chain that protrudes from the collagen fibril. Subsequently, collagenase 3 (matrix metalloproteinase 13; MMP13) is up-regulated as type X collagen is expressed leading to extensive cleavage and removal of type II collagen and of the remaining COL2 domain of type IX collagen alpha 1(IX) chain. The proteoglycan aggrecan is selectively retained in the extracellular matrix. Inhibition of collagenase leads to arrest of hypertrophy as well as gene expression of MMP13. Thus proteolysis and in particular MMP13 are required for chondrocyte differentiation and for matrix resorption in skeletal development. PMID- 11277079 TI - Retinoid signalling and skeletal development. AB - Metabolites of vitamin A, including retinoic acid (RA), comprise a class of molecules known to be important in development and homeostasis. RA functions through a class of nuclear hormone receptors, the RA receptors (RARs), to regulate gene transcription. In the developing mammalian limb, RA affects the differentiation of many cell lineages, including those of the chondrogenic lineage. In excess, RA is a potent teratogen, causing characteristic skeletal defects in a stage- and dose-dependent manner. Genetic analysis has shown that the absence of RARs leads to severe deficiencies in cartilage formation at certain anatomical locations while promoting ectopic cartilage formation at other sites. Expression of either a dominant-negative or a weak constitutively active RAR in the developing limbs of transgenic mice adversely affects chondrogenesis leading to skeletal malformations. Together, these results show that RAR-mediated signalling plays a fundamental role in skeletogenesis. This chapter will focus on the function of RARs in regulating chondroblast differentiation and the contribution of RA signalling to appositional and longitudinal growth of the skeletal primordia. PMID- 11277080 TI - Cartilaginous condensations and the events taking place within them. PMID- 11277081 TI - Defects in extracellular matrix structural proteins in the osteochondrodysplasias. AB - Mutations in the genes that encode structural proteins of the extracellular matrix affect one or more steps in the diverse set of coordinated events necessary for ordered skeletal development. Depending on the role of the gene product and the severity of the defect, disruption of endochondral ossification and linear growth, the structural integrity and stability of articular cartilage, and/or mineralization can occur. Several themes have emerged from the molecular dissection of these disorders; most of the osteochondrodysplasias that result from defects in structural proteins are inherited in an autosomal dominant fashion; a spectrum of related clinical phenotypes can be produced by distinct mutations in the same gene; haploinsufficiency for the gene product usually produces a milder clinical phenotype than do mutations resulting in synthesis of structurally abnormal proteins. For structural defects, a dominant-negative effect resulting from presence of the abnormal protein in the matrix appears to be the primary determinant of phenotype. Secondary effects on extracellular matrix protein structure can result from defects in post-translational maturation, including hydroxylation, sulfation and proteolytic cleavage, and produce distinct osteochondrodysplasias. Overall, the inherited disorders of skeletogenesis have revealed the exquisite sensitivity of the architecture of the extracellular matrix to the quantity and quality of matrix molecules. PMID- 11277082 TI - Genetic control of bone and joint formation. AB - The form and pattern of the vertebrate skeleton is thought to be strongly influenced by several fundamental morphogenetic behaviours of mesenchymal cells during embryonic development. Recent genetic and developmental studies have identified some of the genes that play an important role in controlling both the aggregation of mesenchymal cells into rough outlines of future skeletal elements (condensations), and in controlling where skeletal precursors cleave or segment to produce separate skeletal elements connected by joints. Members of the bone morphogenetic protein (BMP) family appear to play an important role in both processes. Mouse and human mutations in these genes lead to defects in formation of specific bones and joints, with striking specificity for particular anatomical locations. Results from a range of experiments suggest that these molecules may have multiple functions during normal skeletal development and patterning. A major challenge for the future is to identify genes and pathways that can maintain, repair, or stimulate the regeneration of bone and joint structures at later developmental stages. PMID- 11277083 TI - Early steps in limb patterning and chondrogenesis. AB - The interplay of a number of signalling molecules coordinates growth and patterning of the early embryonic vertebrate limb along the three axes. An unresolved question is how this information translates into proper positioning and patterning of the skeletal elements. More is known about how these elements develop. Cells first form precartilaginous condensations and then subsequently differentiate into chondrocytes. This provides a cartilage template that will ultimately be ossified to give rise to the bony skeleton. Several studies support a role for the bone morphogenetic proteins (BMPs) as extracellular signals that regulate early steps of limb chondrogenesis (cartilage formation). However, they have not clarified the step(s) at which BMPs act, and no genetic evidence is available to date. Here, we have used a retroviral vector to misexpress the BMP antagonist Noggin in the embryonic chick limb. We find that BMP signalling is necessary for the formation of precartilaginous condensations, their differentiation into chondrocytes, and for maintenance of chondrogenesis. These results also indicate that Noggin could be clinically useful to treat diseases involving ectopic cartilage formation. PMID- 11277084 TI - The molecular basis of osteoclast differentiation and activation. AB - Osteoclasts develop from haemopoietic cells of the monocyte-macrophage lineage. Osteoblasts or stromal cells are essentially involved in osteoclastogenesis through cell-cell interaction with osteoclast progenitor cells. Recent findings indicate that osteoblasts/stromal cells express osteoclast differentiation factor (ODF, also called RANKL, TRANCE and OPGL) as a membrane-associated factor in response to several osteotropic factors to support osteoclast differentiation. ODF is a new member of the tumour necrosis factor (TNF) ligand family. Osteoclast precursors, which express RANK, a TNF receptor family member, recognize ODF through cell-cell interactions with osteoblasts/stromal cells, and differentiate into osteoclasts in the presence of macrophage colony-stimulating factor (M-CSF). Osteoclastogenesis inhibitory factor (OCIF, also called OPG), is a secreted TNF receptor, which acts as a decoy receptor for ODF. ODF is responsible for inducing not only differentiation, but also activation of osteoclasts. Interleukin 1 alpha (IL-1 alpha) can be substituted for ODF in inducing the activation of osteoclasts. Recently, it was shown that mouse TNF alpha stimulated the differentiation of M-CSF-dependent mouse bone marrow macrophages into osteoclasts in the presence of M-CSF without any help of osteoblasts/stromal cells. Osteoclast formation induced by TNF alpha was inhibited by antibodies against TNF type 1 receptor (TNFR1) or TNFR2, but not by OCIF. Osteoclasts induced by TNF alpha formed resorption pits on dentine slices only in the presence of IL-1 alpha. These results demonstrate that TNF alpha stimulates osteoclast differentiation through a mechanism independent of the ODF-RANK interaction. TNF alpha and IL-1 alpha may play an important role in pathological bone resorption due to inflammation. PMID- 11277086 TI - Developmental mechanisms of vertebrate limb evolution. AB - Over the past few years, our understanding of the evolution of limbs has been improved by important new discoveries in the fossil record. Additionally, rapid progress has been made in identifying the molecular basis of vertebrate limb development. It is now possible to integrate these two areas of research in order to identify the molecular developmental mechanisms underlying the evolution of paired appendages in vertebrates. After the origin of paired appendages, several vertebrate lineages reduced or eliminated fins and limbs and returned to the limbless condition. Examples include eels, caecilians, snakes, slow worms and several marine mammals. Analyses of fossil and extant vertebrates show that evolution of limblessness frequently occurred together with elongation of the trunk and loss of clear morphological boundaries in the vertebral column. This may be suggestive of a common developmental mechanism linking these two processes. We have addressed this question by analysing python embryonic development at tissue, cellular and molecular levels, and we have identified a developmental mechanism which may account for evolution of limb loss in these animals. PMID- 11277085 TI - Clinical disorders of bone resorption. AB - Clinical disorders in which bone resorption is increased are very common and include Paget's disease of bone, osteoporosis, and the bone changes secondary to cancer, such as occur in myeloma and metastases from breast cancer. Clinical disorders of reduced bone resorption are less common and often have a genetic basis, e.g. in osteopetrosis, and in pycnodysostosis due to cathepsin K deficiency. Bone is metabolically active throughout life. After skeletal growth is complete, remodelling of both cortical and trabecular bone continues and results in an annual turnover of about 10% of the adult skeleton. The commonest disorder of bone resorption is osteoporosis, which affects one in three women over 50 years. Its pathophysiological basis includes genetic predisposition and subtle alterations in systemic and local hormones, coupled with environmental influences. Treatment depends mainly on drugs that inhibit bone resorption, either directly or indirectly. This includes bisphosphonates, oestrogens, synthetic oestrogen-related compounds (SERMs--selective oestrogen receptor modulators) and calcitonin. The most widely used drugs for all disorders of increased bone resorption, including osteoporosis, are the bisphosphonates. Recent elucidation of their mode of action, together with the rapidly increasing knowledge of regulatory mechanisms in bone biology, offers many opportunities for the development of new therapeutic agents. PMID- 11277087 TI - Genetic control of skeletal development. AB - There are three major topics of skeleton biology. The first is skeleton patterning, which addresses how the shape and the location of each specific skeletal element is achieved. The second topic is cell differentiation in the skeleton. There are three specific cell types in the skeleton: the chondrocyte in cartilage, and the osteoblast and osteoclast in bone. The first two cell types are from mesenchymal origin while the third is from monocytic origin. The genes controlling skeleton patterning and cell differentiation are for the most part different. The third aspect of skeleton biology addresses the molecular control of the major function of the skeleton such as skeleton growth, bone mineralization and bone remodelling. Our current knowledge in each of these areas of skeleton biology will be presented in broad terms to set the course for other presentations during the symposium. PMID- 11277088 TI - Regulation of chondrocyte growth and differentiation by fibroblast growth factor receptor 3. AB - Both gain-of-function and loss-of-function mutations in fibroblast growth factor receptor 3 (Fgfr3) have revealed unique roles for this receptor during skeletal development. Loss-of-function alleles of Fgfr3 lead to an increase in the size of the hypertrophic zone, delayed closure of the growth plate and the subsequent overgrowth of long bones. Gain-of-function mutations in Fgfr3 have been genetically linked to autosomal dominant dwarfing chondrodysplasia syndromes where both the size and architecture of the epiphyseal growth plate are altered. Analysis of these phenotypes and the biochemical consequences of the mutations in FGFR3 demonstrate that FGFR3-mediated signalling is an essential negative regulator of endochondral ossification. PMID- 11277089 TI - Defects of human skeletogenesis--models and mechanisms. AB - Heritable diseases of the skeleton are a highly complex group of genetic disorders. Skeletal morphogenesis involves, in principle, four distinct developmental processes: patterning, organogenesis, growth and homeostasis. Defects in patterning affect the number and shape of bones and will result in dysostosis. Organogenesis involves the formation of bone and cartilage as an organ. Defects in growth plate function lead to abnormal proliferation and/or differentiation of chondrocytes resulting in dwarfism and dysplasia. Bone mass, shape and strength are maintained in equilibrium throughout development and adulthood (homeostasis). Animal studies are providing good correlations between specific embryological events and gene function, and consequently a framework for understanding the fundamental pathways that build and pattern bone. Based on the remarkable conservation of basic developmental mechanisms between animal species, connections to human disorders are frequently possible. As examples for recent advances in our understanding of the processes that underlie skeletal pathology, the molecular basis of a patterning defect, synpolydactyly, and a defect of organogenesis, cleidocranial dysplasia, will be presented and discussed. PMID- 11277090 TI - [A comparison of methotrexate with placebo for the maintenance of remission in Crohn's disease. North American Crohn's Study Group Investigators]. PMID- 11277091 TI - Why permutation is even more important in IVRS drug codes schedule generation than in patient randomization schedule generation. PMID- 11277092 TI - Urgent colonoscopy for the diagnosis and treatment of severe diverticular hemorrhage. PMID- 11277093 TI - Differences between generalists and specialists in characteristics of patients receiving gastrointestinal procedures. PMID- 11277094 TI - Management of duodenal adenomas in 98 patients with familial adenomatous polyposis. PMID- 11277095 TI - Impressive amelioration of clinical (NYHA class) and echocardiographic parameters in heart failure patients treated with amiodarone and carvedilol. PMID- 11277096 TI - Study of mitral annulus motion by Doppler tissue imaging and M-mode echocardiography. PMID- 11277097 TI - Toxicogenomics: "the call of the wild chip". PMID- 11277098 TI - Monoallelic expression of the odourant receptor gene and axonal projection of olfactory sensory neurones. AB - BACKGROUND: We have previously generated transgenic mice carrying the murine odourant receptor gene, MOR28, tagged with lacZ. In this animal, the endogenous MOR28 is differently tagged with GFP. It was found that the transgenic and endogenous MOR28 genes are expressed in a mutually exclusive manner and that the two sets of olfactory sensory neurones (OSNs), each expressing either the transgenic or the endogenous MOR28, project their axons to separate glomeruli. RESULTS: Our fluorescent in situ hybridization (FISH) revealed that the two endogenous alleles of MOR28 are also mutually excluded for their transcriptional activation. Therefore, we studied whether there would be any segregation in the projection of the two subsets of OSNs: one set expressing the paternal and the other expressing the maternal allele. It was found that the OSNs for both alleles shared the same glomerulus for their projection, but the projection targets were segregated within the glomerular structure. CONCLUSION: Two subsets of neurones expressing either the transgenic or the endogenous MOR28 target their axons to two separate glomeruli based on the differences in the genetic backgrounds, nature of tagging, and chromosomal locations. In contrast, neurones expressing a maternal or paternal allele share the same glomeruli, but tend to target to segregated areas within the glomerular structure. The segregation was more prominent with increased differences in the genetic background between the two alleles. PMID- 11277100 TI - How many eyes do you need? PMID- 11277099 TI - Molecular characterization of the host-adapted pathogen Verticillium longisporum on the basis of a group-I intron found in the nuclear SSU-rRNA gene. AB - Verticillium wilt of oilseed rape is caused by the host-adapted pathogen Verticillium longisporum comb. nov. With one set of nuclear SSU-rRNA gene primers, a PCR amplification product of ca. 2.5 kb was generated from all isolates of V. longisporum tested (36 from Europe, Japan, and USA), with the exception of two recombinant isolates. On the contrary, all the other phytopathogenic and nonphytopathogenic species of Verticillium tested (18 species, 46 isolates), with the exception of one isolate of V. lecanii and two of Cordyceps sp., generated a product of ca. 1.65 kb. Sequence analysis of the SSU rRNA gene of two typical isolates of V. longisporum (wild radish, Japan, and oilseed rape, Germany) revealed that this dimorphism was due to the presence of an identical 839-bp intron located in a highly conserved insertion position (nt 1165 of Saccharomyces cerevisiae). The intron sequence was classified as group-I intron on the basis of conserved sequence and secondary structural elements. Primers designed from the 839-bp intron sequence amplified only the V. longisporum. Phylogenetic analysis based on SSU-rDNA sequences showed that V. longisporum was closely related to the genera of other filamentous Ascomycetes with fruiting bodies. PMID- 11277101 TI - Ocular involvement in congenital erytropoietic porphyria (Gunther's disease): cytopathological evaluation of conjunctival and corneal changes. PMID- 11277102 TI - Bitot's spots and vitamin A deficiency in a child from the UK. PMID- 11277103 TI - Localised chronic eyedlid disease resulting from long term hydroxyurea therapy. PMID- 11277104 TI - Limbal stem cell deficiency arising from systemic chemotherapy. PMID- 11277105 TI - Orbital entomophthoramycosis in an infant: recovery following surgical debridement, combination antifungal therapy and use of hyperbaric oxygen. PMID- 11277106 TI - On absence seizures and oculomotor phenomena. PMID- 11277107 TI - [An 84-year-old woman with progressive mental deterioration and abnormal behavior]. AB - We report an 84-year-old woman with progressive mental deterioration. She was well until January 1994, when she was 80 years of the age. At that time she developed a delusional ideation, in that she stated that she would be killed by her fellow members of the society for elderly, in which she was belonging. At times, she closed the shutter of her house saying that a stranger was wandering outside of her house. In 1995, she could not identify the face of her son's wife. When she went out for shopping, she lost her way to the home. She prowled about in and out of her home. In 1996, she had to be admitted to a nursing home, where quarrelled with other patients and behaved violently. She was admitted to the neurology service of Hatsuishi Hospital on November 20th, 1997. Family history revealed that her mother was said to be demented. On admission, she was alert and behaved in a good manner. She was disoriented to the time and unable to do serial 7. Her memory was very poor. She did not show aphasia or apraxia. Cranial nerves appeared to be intact. She showed no weakness or muscle atrophy. Gait was normal for her age. Plastic rigidity was noted in four limbs more on the right side. No ataxia was noted. Deep tendon reflexes were exaggerated, however, no Babinski sign was noted. Sensory examination was intact. Her hospital course was characterized by the development of progressive gait disturbance, violent behaviour, and prowling around. On November 30th, 1998, she fell down and suffered from a fracture in the neck of her femur. Although replacement of the femur head was performed, she became unable to walk after this episode. Her mental functions deteriorated further. She developed pneumonia and expired on February 2, 1999. She was discussed in a neurological CPC and the chief discussant arrived at a conclusion that the patient probably had diffuse Lewy body disease, because of the combination of dementia and parkinsonism. Other possibilities discussed in the CPC included Pick's disease, frontotemporal dementia and parkinsonism, and Alzheimer's disease. Post-mortem examination revealed moderate atrophy in the frontal and temporal cortices. Microscopic examination showed atrophy and gliosis in the hippocampus. Many diffuse plaque and neuritic plaques were seen in the frontal cortex by methenamine silver staining. Neurofibrillary tangles were also found. The Meynert nucleus was preserved. The putamen and the substantia nigra were also intact. Pathologic diagnosis was consistent with Alzheimer's disease. PMID- 11277108 TI - Improving care and quality of life for all asthma patients. AB - Asthma is an expensive chronic disease that today affects at least 5 percent of the American population. Oftentimes, many patients are failing to properly manage their conditions. It is no surprise, therefore, that many health care providers are looking for ways to help asthma patients better manage their care, use the most effective medications for their conditions, avoid costly emergency room or hospital visits, and improve their day-to-day quality of life. Sometimes, though, providers still face many challenges such as working with populations that experience poverty and poor social support, lack regular sources of health care, or face environmental situations that aggravate their conditions. However, many are finding out that innovative steps can be taken to help all asthma patients receive effective quality care. PMID- 11277110 TI - Early puberty: a cautionary tale. PMID- 11277109 TI - Costs and respiratory syncytial virus. PMID- 11277111 TI - Phototherapy and mesenteric blood flow--too much, not enough, or the wrong kind? PMID- 11277112 TI - The science and fiction of pertussis vaccines. PMID- 11277114 TI - [MMW title pictures are too provocative. "Which devil is influencing editors?"]. PMID- 11277113 TI - Varicella vaccine question. PMID- 11277115 TI - [Patient demands compensation for damage. He who doesn't react, loses!]. PMID- 11277116 TI - [Heartburn. Only a harmless symptom?]. PMID- 11277117 TI - [Large general practice study with repaglinide. Fewer "anxiety eating" in diabetic patients]. PMID- 11277118 TI - [Patient education program for children and adolescents. Brief asthma management by mouse click]. PMID- 11277119 TI - The three-dimensional Z-plasty for the treatment of depressed adhered scars. PMID- 11277120 TI - Removal of soft tissue injected with liquid silicone with an ultrasound-suction system. PMID- 11277121 TI - Results of Furlow Z-plasty in patients with velocardiofacial syndrome. PMID- 11277122 TI - Cognitive dysfunction in the postoperative patient. PMID- 11277123 TI - Intranasal splint obtained from x-ray film. PMID- 11277124 TI - Importance of the depressor septi nasal muscle in rhinoplasty: anatomic study and clinical application. PMID- 11277125 TI - Digitally based free flap using retrograde arterial flow. PMID- 11277126 TI - Blood clot within a silicone gel prosthesis. PMID- 11277127 TI - Prepuce pollicization: a warning. PMID- 11277128 TI - The "stinger". PMID- 11277129 TI - Spastic paretic stiff-legged gait: joint kinetics. AB - OBJECTIVE: The authors previously suggested that spastic paretic stiff-legged gait, defined as reduced knee flexion in swing associated with upper-motor neuron injury, can be attributed to multiple impairments besides spastic quadriceps activity. This study hypothesizes that subjects with spastic paretic stiff-legged gait have altered kinetics not only about the knee but also about the hip and ankle. DESIGN: Joint kinetic data of 20 subjects with spastic paretic stiff legged gait caused by stroke were compared with data obtained from 20 able-bodied subjects. RESULTS: Significant reductions in the subject group were found in both peak knee-joint power absorption (0.42+/-0.34 vs. 0.99+/-0.27 W/(kg x m x m/sec)) and peak ankle-joint power generation (0.74+/-0.42 vs. 1.51+/-0.17 W/(kg x m x m/sec); both P < 0.0001). The authors observed increases in peak external-hip flexion torque in stance, hip-power generation in loading response, knee extension torque in midstance, ankle-dorsiflexion torque, and ankle-power absorption in stance. There was substantial variability in most torque and power values among subjects, which was significantly greater than that observed in the control subjects. CONCLUSIONS: These findings, in conjunction with previous studies, support the likelihood of multiple mechanisms for reduced knee flexion in swing. Alternatively, some of the joint kinetic differences could be compensations for or associated with reduced knee flexion in swing. The substantial variability among subjects implies that despite a similar visual appearance of reduced knee flexion among subjects with a spastic paretic stiff legged gait pattern, each individual has unique mechanisms associated with this observed gait pattern. PMID- 11277130 TI - Rehabilitation of stroke patients: clinical profile and functional outcome. AB - OBJECTIVE: To describe the nature of functional recovery of 67 Turkish survivors of first-episode stroke who were referred for inpatient rehabilitation and to identify the variables that best predict discharge functional status of these patients. DESIGN: A retrospective, descriptive study of the demographic and clinical profile and the functional status of patients with first-episode stroke. RESULTS: The mean age was 60 (11.8, SD) yr, and 35.8% were men. The mean onset admission interval and length of stay were 62.9 and 97.1 days, respectively. The mean functional status score, as measured by the FIM instrument, at the time of admission was 75 compared with 86.7 at the time of discharge, showing a mean improvement of 11.7. Although rehabilitation gains were similar for the right- and left-side involved groups, patients with right-side paresis had lower FIM scores at the time of admission than did the left-side involved group. Significant predictors of functional status at the time of discharge were admission functional status score and onset-admission interval. CONCLUSION: Knowledge of these predictors can contribute to more appropriate treatment and discharge planning. PMID- 11277131 TI - Static wrist position associated with least median nerve compression: sonographic evaluation. AB - OBJECTIVE: To determine the wrist angle that produces the least compression to the median nerve and to evaluate the usefulness of sonography in determining the optimal position. DESIGN: Seventeen wrists of 17 healthy volunteers received dynamic, high-frequency (8 MHz), high-resolution sonography with the wrist splinted at various positions: 15 degrees of flexion, neutral position, and 15 degrees and 30 degrees of extension. The morphologic changes of the median nerve were evaluated with the wrist positioned at various angles. RESULTS: The neutral position caused significantly lower compression of the median nerve than it did in the other positions. However, in some cases, the lowest pressure was found when the wrist was fixed in 15 degrees of flexion or 15 degrees of extension. Because median nerve compression may decrease the anteroposterior diameter, increase the transverse diameter, and decrease the cross-sectional area, greater anteroposterior diameter, lower flattening ratio (transverse diameter/anteroposterior diameter), and greater cross-sectional area were considered to indicate lower median nerve compression. CONCLUSIONS: Neutral position of the wrist is the best position with the least median nerve compression in most individuals. However, the optimal position may vary from person to person. Sonographic examination can help to determine the splint position that results in the lowest median nerve compression. PMID- 11277132 TI - Early rehabilitation of head-neck edema after curative surgery for orofacial tumors. AB - OBJECTIVE: To evaluate a rehabilitative program for postoperative head-neck edema. DESIGN: Eleven patients completed the study. A series of ten manual lymphatic drainage were initiated and completed early after surgery. On discharge from the hospital, the patients wore "made-to-measure" or customized compression garments for the next several weeks. Tape measurements and sonographic evaluation of the soft-tissue width were used to quantify the extent of the swelling. RESULTS: After 6 wk of therapy, the patients exhibited a statistically significant (P < 0.05; Wilcoxon's test) remission; the remission continued in eight patients who were measured at 12+/-3 wk. CONCLUSIONS: This initial trial demonstrates that sequential therapy of manual lymphatic drainage and compression garments can significantly reduce early postoperative edema after curative surgery for orofacial tumors. The outcome can be quantified by comparing the course of distances between the defined anatomic marks and by sonographic evaluation of soft-tissue width. This pilot study encourages that more controlled, randomized studies, with larger numbers of patients, be conducted to verify these results. PMID- 11277133 TI - occupational physical activity and the development of impaired mobility: the 12 year follow-up of the Baltimore Epidemiologic Catchment Area sample. AB - OBJECTIVE: To examine the association between occupational physical activity and self-reported disability. DESIGN: Population-based case control analysis of a longitudinal population-based study in east Baltimore. Eligible participants were aged 18 to 29 yr in 1981, had complete information on occupation in 1981, no disability with tasks related to the domain of mobility in 1981, and complete information on mobility function in 1993 (n = 174). Occupations were divided into low, moderate, and high metabolic equivalents based on job category in 1981. The main outcome measure was disability defined by self-report of difficulty in one or more of five exercise mobility tasks in 1993. RESULTS: Of 174 eligible participants, 45 (26%) reported the onset of disability at follow-up in 1993. A crude odds ratio of 0.25 (95% confidence interval, 0.06, 0.82) was found for the association of moderate compared with low occupational physical activity and the risk of incident disability in mobility tasks. After adjustments to control for possible confounders, moderate job metabolic activity (1.8-2.9 Mets) was independently protective against disability in this cohort (odds ratio = 0.25; 95% confidence interval = 0.083, 0.783). CONCLUSION: In this cohort of people aged 18 to 29 yr, a moderate amount of occupational physical activity was protective against disability in mobility tasks. PMID- 11277134 TI - Cost-effectiveness of screening x-rays at admission to acute rehabilitation after joint replacement surgery: a retrospective chart review. AB - OBJECTIVE: To investigate the cost-effectiveness of performing routine screening x-rays for patients on admission to an acute rehabilitation facility, after hip or knee replacement surgery, by reviewing the overall incidence of abnormal radiographic findings and determining their impact on patient care and outcome. DESIGN: A retrospective chart review study, in which 592 patients were admitted, after hip or knee replacement surgery, to three acute inpatient rehabilitation facilities under one system. RESULTS: Eight of 592 admissions revealed abnormal screening x-rays, for an overall incidence of 1.35%. All of the eight abnormal radiologic cases remained medically stable throughout their acute rehabilitation stay. The abnormalities did not alter the patients' medical management or length of stay. One case, which had demonstrated normal admission films, revealed a dislocated hip prosthesis on a follow-up x-ray, which was obtained as a result of new onset hip pain. The patient was subsequently transferred back to the acute care hospital for surgical correction. CONCLUSIONS: The authors found a relatively low incidence of abnormal admission x-ray findings; furthermore, the detection of abnormal admission films did not alter patient care or outcome. The results suggested that performing routine admission radiologic studies on all patients after joint replacement surgery or hemiarthroplasty may not be a cost effective screening tool in rehabilitation. PMID- 11277135 TI - Lumbosacral plexopathy in gunshot wounds and motor vehicle accidents: comparison of electrophysiologic findings. AB - OBJECTIVE: To characterize the differences between injuries to the lumbosacral (LS) plexus caused by gunshot wounds (GSW) and motor vehicle crashes (MVC) with regard to the location and extent of involvement. DESIGN: A retrospective review of electrophysiologic data from an electromyography laboratory of a county hospital. Nineteen patients with GSW and ten patients with MVC diagnosed by electromyography with an LS plexopathy were included in the study. Injuries were categorized by the number of anatomic quadrants of the LS plexus: upper anterior, upper posterior, lower anterior, and lower posterior. Comparison of upper vs. lower portions and bilaterality of LS plexus involvement was also made. Statistical analyses were performed with two-tailed Fisher's exact and general association tests. RESULTS: Lower portions of the plexus were involved more frequently in patients with MVC compared those observed in patients with GSW. Upper portions of the LS plexus were more involved compared with the lower portions in patients with GSW injuries. More sections of the plexus were involved in patients with MVC compared with those in patients with GSW. CONCLUSIONS: Compared with patients with MVC, patients with GSW had a greater chance of involvement of the upper portion of the plexus. The reverse was true for the lower portion. Hopefully this information will aid the electromyographer and rehabilitation team in the diagnosis and treatment of traumatic plexopathies caused by different etiologies. PMID- 11277136 TI - Role of weight-bearing flexion and extension myelography in evaluating the intervertebral disc. AB - Magnetic resonance imaging has many advantages compared with myelography and/or computed tomography in evaluating the lumbar spine for herniated nucleus pulposus. The authors have included a series of three patients whose histories and physical examinations were clinically suggestive of herniated nucleus pulposus but whose magnetic resonance imaging scans were interpreted by a radiologist as a disc bulge without nerve root compression. Because all patients had not responded to a conservative care treatment program and surgical intervention was to be considered, subsequent testing with lumbar myelography with weight-bearing flexion and extension views demonstrated more clearly the presence of herniated nucleus pulposus along with compression of the nerve root; it also revealed that a positional change in the disc occurred with flexion and extension. PMID- 11277137 TI - Percutaneous, intramuscular neuromuscular electrical stimulation for the treatment of shoulder subluxation and pain in chronic hemiplegia: a case report. AB - This case report describes the first survivor with chronic stroke who was treated with percutaneous, intramuscular neuromuscular electrical stimulation (NMES) for shoulder subluxation and pain. The patient developed shoulder subluxation and pain within 2 mo of his stroke. After discharge from acute inpatient rehabilitation, he developed shoulder and hand pain, which was treated with subacromial bursa steroid injection and ibuprofen with eventual resolution. The patient remained clinically stable until approximately 15 mo after his stroke when he developed severe shoulder pain associated with shoulder abduction, external rotation, and downward traction. The patient could not tolerate transcutaneous NMES because of the pain of stimulation. At approximately 17 mo post-stroke, the patient's posterior deltoid, middle deltoid, and supraspinatus muscles were percutaneously implanted with intramuscular electrodes. After 6 wk of percutaneous, intramuscular NMES treatment, marked improvements in shoulder subluxation and pain, and modest improvements in activities of daily living and motor function were noted. One year after the onset of treatment, the patient remained pain free, but subluxation had recurred. However, the patient was able to volitionally reduce the subluxation by abducting his shoulder. The patient remained pain free for up to 40 mo after the initiation of percutaneous, intramuscular NMES treatment. This case report demonstrates the feasibility of using percutaneous, intramuscular NMES for treating shoulder subluxation and pain in hemiplegia. PMID- 11277138 TI - Fall during a wheelchair transfer: a case of mismatched brakes. AB - We report the case of a patient who had a transtibial amputation and whose wheelchair had been inadvertently fitted with a push-to-lock brake on one side and a pull-to-lock brake on the other. During a standing-pivot transfer from bed to wheelchair, during which the patient thought that she had applied both brakes, the wheelchair turned away from the patient toward the side of the unlocked brake and the patient fell to the floor. This case report has implications for wheelchair design, wheelchair system management, and for user training. PMID- 11277139 TI - Role of survey research in assessing rehabilitation outcomes. AB - Survey research, although often misperceived as superficial and less scientific than other research methodologies, is nonetheless a valid and reliable technique that physiatrists and researchers use to study a range of important rehabilitation outcomes. Surveys are particularly valuable for describing characteristics in large groups--such as the nature and extent of disability within a population--or for learning about outcomes in a given sample. They are the preferred approach for tracking the perceptions of rehabilitation consumers and for obtaining the data that drive service provision, programmatic development, and policy change. On the national, state, and local levels, the authors describe examples of successful survey research efforts, illustrating the types of data obtained, and the usefulness of those data. PMID- 11277140 TI - Implementing guidelines about colorectal cancer: a national survey of target groups. PMID- 11277141 TI - Preoperative depression and mortality in coronary artery bypass surgery: preliminary findings. AB - BACKGROUND: There is convincing evidence to suggest that depression significantly increases the risk of mortality following myocardial infarction. There are few data concerning depression as a risk factor for mortality following cardiac surgery. The aim of the present observational study was to determine if preoperative depressive symptoms resulted in an increased risk of late mortality following cardiac surgery. METHODS: Preoperative assessments of depressive symptoms were performed on 158 patients undergoing coronary artery bypass surgery. Elevated preoperative depression symptoms were defined as a depression anxiety stress scale score of > or = 10. RESULTS: Twenty-four of the 158 patients ( 15.2%) were classified as having elevated preoperative depressive symptoms. Patients were followed for a median of 25 months (range: 4-38 months). Three of the 24 patients (12.5%) with preoperative depressive symptoms died within the follow-up period, compared with three of the 134 (2.2%) non-depressed patients (odds ratio: 6.24; 95% CI: 1.18-32.98; P = 0.046). There were no other group differences on variables including population demographics, medical risk factors, surgical parameters, and indices of postoperative morbidity. CONCLUSIONS: Elevated depressive symptoms before coronary bypass surgery may be a significant predictor of late death. Prospective studies evaluating the prevalence of depressive symptoms in cardiac surgical patients and their effect on long-term outcome must be undertaken. PMID- 11277142 TI - Surgical resection for pulmonary metastases from colorectal cancer. AB - BACKGROUND: Isolated pulmonary metastases from colorectal cancer are rare. The present study reports on the 15-year experience of the Royal Prince Alfred Unit and discusses means of improving survival outcomes. METHODS: This was a retrospective review, over a 15-year period, of 41 patients who had resectable pulmonary metastases of colorectal origin. RESULTS: Most were asymptomatic at the time of diagnosis. Seventy-two per cent had solitary metastases. The most common procedure performed was a lobectomy. Median follow up was 21 months. Five-year survival was 24%. There were no significant prognostic indicators except for the ability to achieve clear surgical margins. CONCLUSION: Morbidity and mortality have not altered significantly over time. But an improved selection process such as the use of preoperative positron emission tomography will potentially improve survival outcomes. PMID- 11277143 TI - Implementing guidelines about colorectal cancer: a national survey of target groups. AB - BACKGROUND: An Australia-wide postal survey was undertaken to determine surgeons' attitudes towards guidelines and their preferred strategies for dissemination and implementation of guidelines for the management of colorectal cancer, developed by the Australian Cancer Network (ACN) and the Clinical Oncological Society of Australia (COSA). This survey was conducted as a baseline before the release of the definitive guidelines. METHODS: All members of the Royal Australasian College of Surgeons (RACS) with a self-nominated special interest in colorectal surgery and members of the Colorectal Surgical Society of Australia (CSSA) were surveyed. RESULTS: A total of 195 of the 219 surgeons eligible for the study returned questionnaires (89% response rate). Most (86%) were aware that these guidelines were being developed. More than one-half had read at least one draft version. Almost half (44.6%; 95%CI: 37.6-51.9%) agreed that guidelines represented 'cookbook medicine' and one-third (33.3%; 95%CI: 26.9-40.5%) agreed that guidelines might increase the number of malpractice suits. Local adaptation of guidelines and 'academic detailing' were most favourably ranked to assure implementation. Further, 54.9% (95%CI: 47.6-61.9%) of respondents believed that a successful legal defence of a surgeon whose practice had been within the guidelines would encourage uptake. Surgeons operating outside teaching hospitals were more likely to endorse this view than others. CONCLUSIONS: These results demonstrate that an important target group for colorectal cancer guidelines, namely surgeons, appears receptive to clinical practice guidelines. These results could also permit interventions that target attitudinal barriers to implementing guidelines and subgroups of surgeons who have particular concerns. Expensive strategies for implementation ought to be subject to rigorous evaluation for their impact in modifying clinical practice. PMID- 11277144 TI - Colorectal surgery in rural Australia: scars; a surgeon-based audit of workload and standards. AB - BACKGROUND: The collection and measurement of colorectal surgical workload, case management and clinical indicators have been mainly based on metropolitan specialist institutions. The aim of the present study was to examine the workload and standards of colorectal surgery in rural Australia. METHODS: Sixty-nine rural general surgeons in Victoria, Albury and South Australia were invited to complete a questionnaire for each transabdominal colorectal operation performed over a 12 month period from 1 May 1996. Data were collected on comorbidity, operation detail, pathology, complications and intention to use adjuvant cancer therapy. RESULTS: Sixty-two surgeons contributed 877 data forms. The patient average age was 65 years with 60% having pre-existing disease. One-third of operations were emergency presentations of which bowel obstruction was the most common. An anastomosis was performed in 675 patients of whom 22 (3.3%) had a clinical anastomotic leak. For low rectal anastomosis the leak rate was 8.9%. Two-thirds of patients had colorectal cancer and 42% of these cancer patients had advanced (Australian clinicopathological stage C or D) disease. The perioperative mortality rate was 4.6% but in the presence of more than two comorbidities it was 16.4%. Mortality was higher with emergency presentations (8.3%), particularly in patients older than 80 years (15.2%). CONCLUSIONS: The study sampled a very high percentage of rural colorectal surgery performed during the audit period. Colorectal surgery clinical indicators were comparable to other Australian studies. Anti-thrombotic and adjuvant therapy were identified as two areas requiring further education. Major surgery is being performed regularly in south eastern rural Australia at a consistently high standard by surgeons who live and work in their rural community. PMID- 11277145 TI - Choledochal cyst: a changing pattern of presentation. AB - BACKGROUND: The aim of the present paper was to study the spectrum of choledochal cysts and analyse the results of various surgical procedures. METHODS: A prospective study was undertaken, at University Hospital, Varanasi, India, of 10 patients with choledochal cyst who presented between January 1996 and June 2000. The patients had a median age at presentation of 16.0 years (interquartile range (IQR): 13.75 years). All patients underwent ultrasonography and endoscopic retrograde cholangiopancreaticography for confirmation of diagnosis. Cyst excision was performed in eight patients followed by reconstruction by hepaticoduodenostomy and Roux-en-Y hepaticojejunostomy in four patients each. Two patients underwent cystoduodenostomy. RESULTS: One patient who underwent hepaticoduodenostomy had a minor leak that responded to conservative management. All patients were asymptomatic at a median follow up of 36.5 months (IQR: 31 months). CONCLUSIONS: There is a changing trend in the commonest mode and age of presentation; fewer patients are presenting with the classical triad and the mean age of presentation is higher. Reconstruction by hepaticoduodenostomy is equally effective, more physiological and less time consuming as compared to hepaticojejunostomy, if the anastomosis can be achieved without tension. PMID- 11277147 TI - Publication bias in presentations to the Annual Scientific Congress. AB - BACKGROUND: Free papers presented to the Annual Scientific Congress (ASC) of the Royal Australasian College of Surgeons (RACS) were reviewed for the years 1994, 1995 and 1996. Reports were examined for evidence of publication bias. METHODS: Suitable free papers were identified from the proceedings of the meetings and authors were contacted to obtain information about the research reported and any publications resulting from it. RESULTS: Responses were obtained from 302 of 576 presentations considered suitable. A total of 55% of responding authors reported publication of their paper. Basic science papers were most likely to be published. There was a significant bias in favour of publication of positive results (98 of 139 positive vs 76 of 159 inconclusive or negative reports; P < 0.01). Retrospective data were as likely to be published as prospective (51% and 57%, respectively). Reports describing studies of high-level evidence were more likely to be published in journals with a high impact factor. CONCLUSION: The ASC is a comprehensive meeting that attracts a wide range of free papers from most sections of the RACS. There appears to be no evidence of bias in selection of papers for inclusion in the meeting but there is bias in the subsequent publication, which favours positive reports. PMID- 11277146 TI - Australian major incident nomenclature: it may be a 'disaster' but in an 'emergency' it is just a mess. AB - BACKGROUND: A standardized major incident nomenclature has practical applications for medical communication and audit of the medical response to incidents. METHODS: A telephone and fax survey of major incident nomenclature in State and Territory health service emergency management plans and 'disaster' legislation was carried out on 13 August 1999. RESULTS: Within Australia there were a total of 13 different terms to describe incidents that could produce casualties: there were four definitions of the word 'disaster', eight definitions of the word 'emergency' and one definition of the word 'incident'. CONCLUSION: Australia lacks a uniform system of classifying and recording mass casualty incidents. This prevents both the independent clinical audit of the medical response to an incident and the cross-border comparison of the effectiveness of trauma systems to deal with multiple casualties. Australia's geography highlights the need to develop a nomenclature that allows medical practitioners, in isolated environments, to accurately describe an incident and the medical support that is required. The Potential Injury-Creating Event (PICE) nomenclature is a simple system to describe the functional impact of an event upon a community and the level of medical support required. It can be used to provide the basis for the uniform reporting of the medical management of major incidents within Australia. PMID- 11277148 TI - Arthroscopic subacromial decompression with a holmium:YAG laser: review of the literature. AB - BACKGROUND: The Australian Safety and Efficacy Register of New Interventional Procedures-Surgical (ASERNIP-S) undertook to systematically review the literature regarding arthroscopic subacromial decompression (ASD) using a holmium:YAG laser for patients with impingement syndrome, with respect to the safety and efficacy of the procedure. METHODS: Studies on ASD with a holmium:YAG laser were identified using MEDLINE (1984 to July 2000), EMBASE (1974 to August 2000) and Current Contents (1993 to week 33 2000). A number of search terms were used: (laser and shoulder) and (surgery or arthroscop* or acromioplasty or orthopaed* or orthoped* or subacromial decompression or impingement syndrome). The Cochrane Library was searched from 1966 to issue 3 2000, using the search terms 'shoulder and surgery'. Human studies were included for patients with impingement syndrome but without full-thickness rotator cuff tears or rheumatological disorders, and where shoulder pain had been experienced for more than 3 months. A surgeon and reviewer independently assessed the retrieved articles for their inclusion in the review. RESULTS: Seven papers were identified that related to ASD with a holmium:YAG laser. None of the papers for review offered high-quality evidence. There were no properly designed randomized controlled studies. The highest level of evidence came from time series studies. No quantitative analysis could be undertaken for this review. CONCLUSIONS: Given the extremely low level of evidence available for this procedure it was recommended that further research be conducted to establish the safety and efficacy of the technique. This reinforces the conclusion reached in the Cochrane review of interventions for shoulder pain where insufficient evidence was found to either support or refute the efficacy of other interventions for shoulder pain. PMID- 11277149 TI - The development of undergraduate curricula in surgery: III. Assessment. AB - The present review is aimed at providing an overview of the assessment process. The mode of assessment has a powerful influence on the learning behaviour of students. It is therefore important to ensure that there is congruity between the objective, the task and the test. In other words: define it, teach it, examine it. It is difficult to evaluate many of the attributes that we desire in a doctor; and examples of this include empathy, ethical behaviour, problem-solving skills, ability to self-educate and teamwork. Nevertheless, it is generally agreed that it is better to measure uncertainly the significant than to measure reliably and validly the trivial. Furthermore different methods of assessment suit different educational objectives (fitness for purpose) and this supports the use of multiple assessment techniques. PMID- 11277151 TI - Bronchial sleeve resection for a patient with an inflammatory pseudotumour. PMID- 11277150 TI - Severe cutaneous mucormycosis (Zygomycosis) due to Apophysomyces elegans. PMID- 11277152 TI - Ureterocystoplasty: new options. PMID- 11277153 TI - Recurrent idiopathic small bowel volvulus during pregnancy. PMID- 11277154 TI - Double ileocaecal and colonic intussusception due to malignant lymphoma of the caecum in an HIV-positive patient. PMID- 11277156 TI - Celeste Phillips, the mother of family-centered maternity care. PMID- 11277155 TI - Use of Cytotec (misoprostol) for labor induction. PMID- 11277157 TI - Birth center outcomes reported through automated technology. AB - BirthCare HealthCare is a multidisciplinary health care clinic and freestanding birth center in Salt Lake City, Utah. Highlights of a woman's labor assessment and birth outcomes are summarized in a paper birth log. The authors developed a relational database that facilitates efficient outcomes reporting. Examples of outcomes from this database are reported. OBJECTIVES: This study describes clinical outcomes from a freestanding birth center and the automated birth log database used to manage outcomes reporting. DESIGN: Retrospective data obtained primarily from a paper birth log and secondarily from the paper chart were entered into and extracted from an electronic database. PARTICIPANTS: 231 women who were evaluated in 269 encounters for delivery at the birth center between July 1, 1997, and June 30, 1999. MAIN CLINICAL OUTCOME MEASURES: The number of deliveries at the birth center; the percentage of intrapartum, postpartum, and newborn transfers to the inpatient hospital setting; cause-specific transfer rates; method of transfer; medication in labor; low and high birth weights; percentage of low Apgar scores; and average times from admission to birth and birth to discharge. CONCLUSION: Birth center clinical outcomes can be efficiently reported using an automated birth log. Clinical outcomes reported in this study are consistent with finding of previous research that demonstrates that birth centers are a safe alternative site for delivery. PMID- 11277158 TI - Care of the Native American woman: strategies for practice, education, and research. AB - Native Americans, the smallest racial minority in the United States, comprise the fastest growing ethnic group and have a myriad of social and health problems. Women play an important role in health care practices and decision making in this community because many tribes are matrilineal. Practice, education, and research strategies should include identification of beliefs and practices specific to the clan or tribe because there is wide variance in values, lifestyles, and taboos from tribe to tribe. Traditional healers, Native American storytelling, and talking circles can be incorporated into the health care of urban Native American women and their families. PMID- 11277159 TI - Reducing the preterm birth rate: a population health strategy. AB - The rate of preterm birth has been increasing in Canada and the United States. Efforts to prevent preterm birth have been largely ineffective. A population health strategy that integrates disease prevention and health promotion is needed. In this article, the five categories of health determinants proposed by the Federal, Provincial and Territorial Advisory Committee on Population Health are used as a framework to discuss risk factors and propose policies and interventions to reduce the preterm birth rate. PMID- 11277160 TI - Neonatal skin care: evaluation of the AWHONN/NANN research-based practice project on knowledge and skin care practices. Association of Women's Health, Obstetric and Neonatal Nurses/National Association of Neonatal Nurses. AB - OBJECTIVE: To develop and evaluate an evidence-based clinical practice guideline for assessment and routine care of neonatal skin, educate nurses about the scientific basis for practices recommended in the guideline, and design procedures that facilitate implementation of the project guideline into clinical practice. DESIGN: Descriptive report of the collaborative neonatal skin care research-based practice project of the Association of Women's Health, Obstetric and Neonatal Nurses and the National Association of Neonatal Nurses. SETTING: Neonatal intensive-care unit (NICU) and special-care nurseries and well-baby nurseries in 51 hospitals located throughout the United States. PARTICIPANTS: Member site coordinators (N = 51), nurses who work at the selected sites, and the neonates observed during both the pre- and postimplementation phases of the project (N = 2,820). METHOD: An evidence-based clinical practice guideline was developed, sites were selected from all respondents of the call for sites, site coordinator training was provided, data collection was facilitated by project specific data collection tools, and the project was evaluated by the science team. MAIN OUTCOME MEASURES: Diversity and numbers of sites represented, patient representation, site coordinator knowledge of neonatal skin care pre- and postimplementation, use of project-designed implementation tools, satisfaction with project guideline and the data collection process, changes in practices and product use, and site coordinators' experiences during guideline implementation. RESULTS: Fifty-one sites completed the project, representing NICU, special-care, and well-baby nurseries in both academic and community hospital settings in 27 states. Registered nurses working in these sites totaled 4,754 full-time equivalent positions (FTEs) (in NICU/special-care and well-baby nurseries). Site coordinators demonstrated increased knowledge of research-based neonatal skin care and satisfaction with the implementation tools and data collection process. Product use changed, reflecting acquisition of new knowledge. Barriers to implementation of the guideline were identified. CONCLUSIONS: The AWHONN/NANN Neonatal Skin Care Research-Based Practice Project demonstrated increased knowledge among site coordinators who received training, facilitated changes in neonatal skin care as defined by the practice guideline, and thus advanced evidence-based clinical practice. PMID- 11277161 TI - Neonatal skin care: clinical outcomes of the AWHONN/NANN evidence-based clinical practice guideline. Association of Women's Health, Obstetric and Neonatal Nurses and the National Association of Neonatal Nurses. AB - OBJECTIVE: To test the effectiveness of an evidence-based clinical practice guideline for neonatal skin care on selected clinical outcomes for newborns in neonatal intensive-care units (NICU), special-care units (SCU), and well-baby nurseries. DESIGN: Prospective evaluation of the collaborative neonatal skin care research-based practice project of the Association of Women's Health, Obstetric and Neonatal Nurses and the National Association of Neonatal Nurses. SETTING: NICU and well-baby units in 51 hospitals located throughout the United States. PARTICIPANTS: Member site coordinators (N = 51) and the neonates (N= 2,820) observed during both the pre- and postimplementation phases of the project. METHOD: Site coordinators received specialized education in neonatal skin care and implemented an evidence-based clinical practice guideline addressing 10 aspects of neonatal skin care. Baseline observations of skin condition, care practices, and environment of newly admitted neonates were collected by site coordinators. Postimplementation observations were then completed. MAIN OUTCOME MEASURES: Skin condition was assessed with the Neonatal Skin Condition Score (NSCS), which ranges from a score of three (best condition) to a score of nine (worst condition), based on dryness, erythema, and skin breakdown. Changes in frequency of selected skin care practices were used to assess the effectiveness and feasibility of using the practice guideline in everyday clinical practice. Aspects of the care environment with potential effect on skin integrity were monitored to determine risk factors. RESULTS: Fifty-one site coordinators made 11,468 systematic assessments of 2,464 NICU and SCU newborns and 356 well newborns. Baseline skin scores were better in well newborns compared with premature newborns. After implementation of the guideline, skin condition was improved, as reflected by less visible dryness, redness, and skin breakdown in both the NICU/SCU and well newborns. The guideline was integrated into care, as evidenced by increased use of emollients, particularly with premature infants, and decreased frequency of bathing. A relationship was shown between selected aspects of the environment and alterations in skin integrity. CONCLUSIONS: Use of the AWHONN/NANN Neonatal Skin Care Research-Based Clinical Practice Guideline was successfully implemented at 51 sites, and effectiveness was demonstrated by changed care practices and improved skin condition in premature and full-term newborns. The results of this project support a wider dissemination of the project's practice guideline for neonatal skin care. PMID- 11277162 TI - Specialized care for twin gestations: improving newborn outcomes and reducing costs. AB - OBJECTIVE: To compare newborn outcomes and costs of hospital stays for twins born to mothers receiving care in a specialized twin clinic with a research-based care protocol and one consistent caregiver versus twins whose mothers received standard prenatal care. DESIGN AND SETTING: A retrospective, historical cohort study conducted in a high-risk obstetric clinic in central Texas. PATIENTS: Thirty women pregnant with twins received specialized care. The comparison group consisted of 41 women pregnant with twins who received standard care. INTERVENTIONS: An advanced practice nurse provided prenatal care, which included weekly clinic visits, home visits, and 24-hour availability for phone support. OUTCOME MEASURES: Gestational age at birth, birth weight, length of stay in the neonatal intensive-care unit (NICU), and hospital charges for the newborns. RESULTS: No newborns of less than 30 weeks gestation were born to women in the specialized care group, the mean birth weight was 249 g (SD +/- 77) higher, days in the NICU were reduced from a mean of 17 to 7, and hospital charges were $30,000 less per infant. CONCLUSIONS: Newborn outcomes were improved and length of stay and hospital charges were significantly reduced for newborns whose mothers had received care in the specialized twin clinic. PMID- 11277163 TI - Effect of acupressure by Sea-Bands on nausea and vomiting of pregnancy. AB - OBJECTIVE: To determine the effect of continuous acupressure at P6 applied by Sea Bands with acupressure buttons on the frequency and severity of nausea and vomiting of pregnancy during the 1 st trimester. DESIGN: A two-group, quasi experimental, posttest-only and posttest-repeated measure. SETTING: Seventeen medical clinics or offices in southern Michigan. PARTICIPANTS: Convenience sample of English-speaking, healthy pregnant women in their 1 st trimester, who had at least one episode of nausea, vomiting, or both before their prenatal clinic/office visit where they were recruited. After being accepted for the study, the women were randomly assigned to treatment or placebo groups. INTERVENTION: Treatment group 1 applied SeaBands with acupressure buttons to both wrists for 4 days and removed the Sea-Bands for 3 subsequent days. Placebo group 2 applied the Sea-Bands without acupressure buttons to both wrists on the same time schedule as group 1. MAIN OUTCOME MEASURE: Self-report daily diaries of the number of times per day that participants experienced nausea, the severity of nausea, the number of vomiting episodes per day, and the severity of vomiting. RESULTS: Mann-Whitney U procedures revealed that the treatment group had significantly less frequency and severity of nausea and vomiting of pregnancy while wearing the Sea-Bands than did the placebo group. The treatment group also had significantly less frequency and severity of nausea and vomiting of pregnancy while wearing the SeaBands than when not wearing the Sea-Bands. CONCLUSIONS: Sea Bands with acupressure buttons are a noninvasive, inexpensive, safe, and effective treatment for the nausea and vomiting of pregnancy. PMID- 11277164 TI - Mothers recovering from cocaine addiction: factors affecting parenting skills. AB - OBJECTIVE: To identify factors that may influence parenting by mothers who are recovering from cocaine addiction. DESIGN: Exploratory descriptive, with in-depth unstructured interviews. SETTING: Interviews were conducted in the woman's home or in a treatment center. PARTICIPANTS: A convenience sample of 11 women recovering from cocaine addiction who were mothers of children 3 years of age and younger. RESULTS: A content analysis was used to analyze the interview data. Two themes, personal/psychologic factors and environmental/contextual factors, and four subthemes emerged. They identify issues that may affect parenting by mothers being treated for cocaine addiction. Subthemes included low self-esteem, difficulty developing a maternal identity, isolation from friends and family, and chronic life stress. CONCLUSION: This study provides a better understanding of the sources contributing to vulnerability in the parenting role for mothers recovering from cocaine addiction and will assist nurses in providing care for these mothers and their children. PMID- 11277165 TI - Improving adherence to abnormal Pap smear follow-up. AB - OBJECTIVE: To gain a better understanding of factors that affect follow-up and the strategies that have been found to improve follow-up after an abnormal Papanicolaou (Pap) smear test. DATA SOURCES: A computer-based search of the literature was conducted using MEDLINE with the keywords adherence, nonadherence, compliance, follow-up, and abnormal Pap smears. STUDY SELECTION: Research studies published between 1985 and 1999 in the English language were included. If relevant studies were cited in the articles reviewed, these studies also were reviewed. A total of 25 studies were reviewed. DATA EXTRACTION: A critical review of these studies was conducted, with special attention to implications for clinical practice as well as future research. The studies fell into two groups: factors associated with nonadherence and strategies developed to improve adherence. DATA SYNTHESIS: A number of factors were identified that affect adherence to follow-up among women with abnormal Pap smears. Some of these factors involve characteristics of the woman, such as demographics, social support, lack of understanding, and fear. Factors that pertain to the health care system, such as inconvenient clinic hours, male providers, and insensitive staff, also were identified. A number of strategies have been successful in improving follow-up, including telephone counseling, educational programs, and economic incentives. CONCLUSIONS: Nurses play a crucial role in facilitating adherence to recommendations for follow-up. They can identify women at risk for poor follow up, increase awareness of the factors that affect follow-up, and implement strategies shown to be successful in improving adherence. Future research should take into account the multifactoral nature of adherence as well as the patient's perspective. In addition, studies should be designed with special attention to generalizability and should include women from populations most at risk for cervical cancer. PMID- 11277166 TI - Reflections on a decade of outcomes management in women's services. AB - Outcomes management was theorized as a strategy for redesigning and continually enhancing interdisciplinary health care processes through the use of outcomes measurements. Regardless of the rapidly changing and oftentimes turbulent health care environment, this strategy successfully facilitated positive outcomes in one women's service for 10 years. Outcomes affected during these years included reduction in cost per case, length of stay, infant prematurity, and cesarean delivery rates. PMID- 11277167 TI - Surviving the big one. PMID- 11277168 TI - Severity and risk adjusting relating to obstetric outcomes, DRG assignment, and reimbursement. AB - Obstetric risk has important implications for reporting and benchmarking quality in today's managed health care environment. Administrative data, including diagnosis related group (DRG) information collected by hospitals, is used by payers and governmental groups for reimbursement, monitoring quality, and setting financial rates. Obstetric conditions that affect the patient experience are coded but do not often contribute to the overall DRG assignment. This strategy, therefore, may provide comparisons that are misleading to consumers and payers. Additionally, financial rates often do not provide adequate reimbursement for the cost incurred in caring for high-risk patients. Finally, risk prediction strategies have historically been used to both identify vulnerable patients for early management and make more equitable comparisons of groups of patients. PMID- 11277169 TI - Notice of duplicate publication. PMID- 11277170 TI - Mesenchymal stem cells: heading into the clinic. AB - In recent years, there has been an increasing interest in non-hematopoietic pluripotent progenitor cells that are found in the bone marrow. Mesenchymal stem cells (MSCs) are the first non-hematopoietic progenitors to be isolated from the bone marrow and extensively characterized. In addition to their ability to support hematopoiesis, MSCs can differentiate into osteocytes, chondrocytes, tenocytes, adipocytes and smooth muscle cells. This article will review our current understanding of bone marrow stroma and MSCs and their potential therapeutic role in the setting of hematopoietic stem cell transplantation. PMID- 11277171 TI - Peripheral blood progenitor cell mobilisation alters myeloid, but not erythroid, progenitor cell self-renewal kinetics. AB - Transplantation of progenitor cells which have been mobilised into the bloodstream (PBPC) following the administration of G-CSF results in more rapid neutrophil recovery than transplantation of bone marrow (BM). The reasons for the accelerated neutrophil engraftment are not clear, but would be explained by increased self-replication of myeloid progenitor cells (CFU-GM). We have used a CFU-GM replating assay to investigate myeloid progenitor self-replication, and quantification of subcolony formation during erythroid burst formation to quantify erythroid progenitor self-renewal. Secondary colony formation by CFU-GM, grown from PBPC and then replated was increased compared with secondary colony formation by BM CFU-GM (P = 0.0001); erythroid subcolony formation was not altered. There was no difference between the replating abilities of PBPC CFU-GM derived from allogeneic donors (normal individuals) and autologous donors (patients with malignant disease) although differences were found between subgroups of autologous donors. The increased replication of PBPC could not be accounted for by a reduction in progenitor cell apoptosis; PBPC CFU-GM contained slightly fewer apoptotic CD34+ cells than BM CFU-GM. The increased replication by PBPC CFU-GM was reversible because it declined when CFU-GM colonies were passaged through three sequential CFU-GM replating cycles. This decline in self replication was more rapid than the decline seen in replated BM CFU-GM. The self replication of PBPC CFU-GM, and subcolony formation by BFU-E could be further enhanced by exposure to cytokines in vitro. We conclude that mobilisation alters the replication kinetics of myeloid, but not of erythroid, progenitor cells, that mobilisation-induced events are of limited duration and that in vitro exposure to cytokines may modify PBPC progenitor cell kinetics. PMID- 11277172 TI - Hematopoietic recovery of ex vivo perfusion culture expanded bone marrow and unexpanded peripheral blood progenitors after myeloablative chemotherapy. AB - Ex vivo culture of hematopoietic progenitor cells for autologous transplantation has generated world-wide interest, since it offers the prospect of using a limited cell number, and may allow more efficient gene transfer and passive elimination of contaminating tumor cells. In this study, we expanded bone marrow (BM) cells from 10 breast cancer patients to determine whether small BM aliquots can durably restore hematopoiesis, and whether thrombopoietin (TPO) improves hematopoietic reconstitution after myeloablative chemotherapy. We used the AastromReplicell System (ARS), performing a computer-controlled, stromal-based cell expansion process with frequent medium, cytokine and gas exchange. For the inoculation of 9 x 10(8) MNC, a median BM volume of 97.8 ml (range, 72.4-272) was harvested. We found a median 4.5-fold nucleated cell expansion, an 18-fold CFU-GM expansion, and 69% of input LTC-IC numbers. Nucleated and Lin-/CD34+ cells were infused with a median of 43.5 x 10(6)/kg (range, 34.1-71.7) and 2.8 x 10(5)/kg (range, 0.95-5.9), respectively. Despite tumor cell detection by immunocytochemical staining in 3/10 patients before expansion, tumor cells were not detectable in 9/10, and in one patient 1 log reduced post ARS culture. Following high-dose STAMP V chemotherapy, all patients received 12-day expanded BM cells. The median time to engraftment was 17 days (range, 11-20) for WBC >1000/microl, and 28 days (range, 21-55) for platelets >20,000/microl. A correlation between post-expansion Lin-/CD34+ cells and engraftment for ANC >500/microl, WBC >1000/microl and platelets >20,000/microl was observed. Hematopoiesis has been maintained for a median of 15 (range, 6-24) months. Our results demonstrate that transplantation of ex vivo expanded small BM aliquots allows hematopoietic reconstitution after myeloablative chemotherapy. Ex vivo generated ARS cells can reduce the risk of tumor cell reinoculation with autotransplants and may be valuable in settings in which only small stem cell doses are available, eg when using cord blood transplants or in non-mobilizing patients. PMID- 11277173 TI - Recombinant human thrombopoietin (rhTPO) after autologous bone marrow transplantation: a phase I pharmacokinetic and pharmacodynamic study. AB - Thrombocytopenia following myelotoxic therapy is a common problem and when severe (<20,000/microl) can lead to severe morbidity and mortality. Thrombopoietin (TPO) is a naturally occurring glycosylated peptide which stimulates the differentiation of bone marrow stem cells into megakaryocyte progenitor cells, induces the expression of megakaryocyte differentiation markers, promotes megakaryocyte proliferation, polyploidization and, ultimately, the formation of increased numbers of platelets in the circulation. TPO has now been produced by recombinant technology and has entered clinical trials. This open label phase I study was designed to determine the safety, tolerance and pharmacokinetics of recombinant thrombopoietin (rhTPO) when administered to patients after undergoing high-dose chemotherapy followed by autologous bone marrow transplantation. rhTPO was administered intravenously by bolus injection at doses ranging from 0.3 to 4.8 microg/kg/day every 3 days to 30 patients and 0.6 microg/kg daily to three patients. rhTPO was begun the day after marrow infusion and continued until platelet recovery to >20,000/microl. G-CSF was concomitantly administered to promote myeloid recovery. Serious adverse events or neutralizing antibodies to rhTPO were not observed during the study. Median platelet recovery after ABMT was 19 days (range, 11-41). Neither the dose nor the schedule of rhTPO appeared to have any impact upon the time course of platelet recovery. In this phase I study, rhTPO was found to be well tolerated without the development of neutralizing antibodies and without compromising neutrophil recovery. Platelet recovery was similar for all doses studied warranting further evaluation in phase II and III trials designed to test for platelet recovery efficacy. PMID- 11277175 TI - Transfer of idiotypic protein primed allogeneic marrow grafts elicits potent graft-versus-myeloma effects in mice. AB - The active immunization of bone marrow (BM) donors with myeloma immunoglobulin (Ig) results in an idiotypic T cell response that can be transferred to the recipient. Using a murine model we evaluated the effectiveness, side-effects and underlying mechanisms of this approach. Balb/c (H-2d) mice were given a dose of HOPC-1F myeloma cells secreting the monoclonal IgG2a followed by lethal total body irradiation (7.5 Gy) 2 days later and a subsequent transplantation of 2 x 10(7) allogeneic MHC-matched DBA/2-derived marrow cells. Donors were pre immunized with three i.p. injections of HOPC(IgG2a) or control Ig given with incomplete Freund's adjuvants (IFA) spaced 1 week apart. In some experiments, donor-spleen cells were additionally transferred 2 h post transplant. Injection of HOPC-myeloma led to death of all animals after a median survival time (MST) of 42 days. A lethal dose of TBI followed by transfer of unmanipulated marrow grafts plus splenocytes resulted in moderate antimyeloma effects with 8% of mice achieving long-term survival. Nearly the same results were obtained after transplantation of BM immunized with the control Ig. In contrast, transplantation of marrow grafts from HOPC(IgG2a) immunized donors exerted a significant GVM effect with 63% long-term survival for more than 180 days. The additional transfer of 2 x 10(7) immune splenocytes derived from the same donor resulted in even stronger anti-myeloma effects (FFR 87%). No increase in the incidence of severe acute GVHD was observed. In vitro data suggest that allogeneic CD8+ idiotype-specific T cells may be the major effector cells. Our results demonstrate that active immunization of the donor with the myeloma-specific Ig can induce powerful graft-versus-myeloma effects after allogeneic BMT. PMID- 11277174 TI - A short course of induction chemotherapy followed by two cycles of high-dose chemotherapy with stem cell rescue for chemotherapy naive metastatic breast cancer. AB - A single cycle of high-dose chemotherapy with stem cell support (HDC) in women with responsive metastatic breast cancer (BC) consistently achieves over 50% complete and near complete response (CR/nCR). This significant cytoreduction results in a median event-free survival (EFS) of 8 months, and approximately 20% 3-year and 16% 5-year EFS in selected patients. To improve long-term outcomes, new strategies to treat minimal residual tumor burden are needed. Increased total dose delivered can be achieved with two cycles of HDC. Critical design issues include shortening induction chemotherapy to avoid development of drug resistance and the use of different agents for each HDC cycle. We have determined the maximum tolerated dose (MTD) for paclitaxel combined with high-dose melphalan in the context of a double transplant and explored the impact of a short induction phase. Between June 1994 and August 1996, we enrolled 32 women with metastatic BC on to this phase I double transplant trial. Induction consisted of doxorubicin 30 mg/m2/day days 1-3 given for 2 cycles every 14 days with G-CSF 5 microg/kg s.c. days 4-12. Stem cell collection was performed by leukapheresis in each cycle when the WBC recovered to above 1000/microl. Patients with stable disease or better response to induction were eligible to proceed with HDC. HDC regimen I (TxM) included paclitaxel with dose escalation from 0 to 300 mg/m2 given on day 1 and melphalan 180 mg/m2 in two divided doses given on day 3. HDC regimen II was CTCb (cyclophosphamide 6 g/m2, thiotepa 500 mg/m2, and carboplatin 800 mg/m2 total doses) delivered by 96-h continuous infusion. At the first dose level of 150 mg/m2 paclitaxel by 3 h infusion, four of five patients developed dose-limiting toxicity consisting of diffuse skin erythema and capillary leak syndrome. Only two of these five completed the second transplant. Subsequently, paclitaxel was delivered by 24-h continuous infusion together with 96 h of dexamethasone and histamine receptor blockade. This particular toxicity was not observed again. No toxic deaths occurred and dose-limiting toxicity was not encountered. Three patients were removed from study prior to transplant: one for insurance refusal and two for disease progression. All others completed both cycles of transplant. Complete and near complete response (CR/nCR) after completion of therapy was achieved in 23 (72%) of 32 patients. The median EFS is 26 months. The median overall survival has not yet been reached. At a median follow-up of 58 months, EFS and overall survival are 41% and 53%, respectively. This double transplant approach is feasible, safe, and tolerable. Treatment duration is only 14 weeks and eliminates lengthy induction chemotherapy. The observed event-free and overall survivals are promising and are better than expected following a single transplant. Whilst selection biases may in part contribute to this effect, a much larger phase II double transplant trial is warranted in preparation for a potential randomized comparison of standard therapy vs single vs double transplant. PMID- 11277176 TI - Prognostic significance of the immunocytochemical detection of contaminating tumor cells (CTC) in apheresis products of patients with high-risk breast cancer treated with high-dose chemotherapy and stem cell transplantation. AB - The aim of this study was to determine whether the detection of CTC in the apheresis product contribute significantly to treatment failure of patients with high-risk breast carcinoma treated with high-dose chemotherapy (HDC) and stem cell transplantation (SCT). Patients were with stage II and III adenocarcinoma of the breast with > or = 10 axillary lymph nodes affected after primary surgery (> or = 10 N+) who had received HDC with SCT. We analyzed retrospectively the presence of CTC as assessed by immunocytochemistry (ICC) in the apheresis products obtained after standard adjuvant chemotherapy. We compared the clinical outcome of patients who received HDC and SCT with or without CTC-positive apheresis. One hundred and twenty-seven apheresis products samples were obtained from 51 patients. Fourteen (27.4%) of these samples were CTC positive. After a median follow-up of 4.6 years, 20 patients have relapsed, 14 died from progression of their disease and 30 patients remain alive and free of progression. For the whole group of patients the 5 year probabilities of DFS and OS were 60% (IC 95%, 47-75%) and 71% (IC 95%, 55-83%), respectively. However, the 5 year probabilities of DFS were 23% (IC 95%, 0-46) and 75% (IC 95%, 60-89) for patients with CTC positive and negative, respectively. The 5 year probabilities of OS were 42% (IC 95%, 15-68) and 83% (IC 95%, 70-95) for patients with CTC positive and negative, respectively. Both univariate and multivariate analysis showed that the presence of CTC in the apheresis product was the only prognostic factor associated with a higher incidence of clinically overt disease relapse (P = 0.002) and shorter survival (P = 0.003). The presence of cytokeratin-positive metastatic cells in the apheresis product increases the risk of relapse after HDC and SCT in patients with stage II and III adenocarcinoma of the breast with > or = 10 N+. PMID- 11277177 TI - Parainfluenza virus type 3 infections in a hematology unit. AB - Parainfluenza virus type 3 (PIV3) is associated with a high mortality rate in BMT recipients with lower respiratory tract infections. We describe nine patients with hematological malignancies (five having undergone either allogeneic or autologous stem cell transplantation) identified as having PIV3 infection during a 2-month period in a Hematology Unit. Four patients with infiltrates on chest radiograph received intravenous ribavirin therapy; all survived. The infection was community-acquired in two patients, while nosocomial origin of the disease was evident, or presumed, in the remaining seven. The policy implemented to control the spread of PIV3 was as follows: (1) nasopharyngeal samples for antigen detection were obtained from all patients presenting with respiratory symptoms; (2) all diagnosed (or suspected) PIV3-positive hematological patients were nursed following contact isolation precautions, preferably in the Infectious Diseases Unit; and (3) staff were given further education on hospital hygiene. Our experience shows that it may be possible to avoid mortality for PIV3 lower respiratory tract infection in immunocompromised patients by early commencement of intravenous ribavirin. It is also possible, even without closing the ward, to contain nosocomial spread of PIV3 by implementing systematic nasopharyngeal sampling for rapid diagnostics, and by strict adherence to cohorting and contact isolation precautions. PMID- 11277178 TI - Screening for cytomegalovirus (CMV) infection in allogeneic bone marrow transplantation using a quantitative whole blood polymerase chain reaction (PCR) method: analysis of potential risk factors for CMV infection. AB - Potential risk factors for CMV infection and the use of quantitative CMV PCR screening to guide pre-emptive anti-CMV therapy were reviewed retrospectively in 32 allogeneic bone marrow transplant patients accrued over a 2-year period. Significant CMV PCR positivity (an indicator of CMV infection) developed in 34% of patients. When analysed by recipient CMV IgG serostatus, 69% of seropositive recipients developed significant CMV PCR positivity while none of the seronegative recipients did so (P = 0.00007). Considering only the seropositive recipients, 100% of those who received the low intensity campath 1H/fludarabine/melphalan 'mini-allograft' conditioning regimen developed significant CMV PCR positivity, while only 44% of those who had received cyclophosphamide/TBI did so (P = 0.0337). The mean time to first episode of significant CMV PCR positivity for those who had received campath/fludarabine/melphalan was 25 days while for those who had received cyclophosphamide/TBI, this was 66 days (P = 0.0372). For the first episode of significant CMV PCR positivity, the mean index and peak CMV PCR counts for those who had received campath/fludarabine/melphalan were 4.54 and 5.22 log copies/ml respectively, while for cyclophosphamide/TBI, the corresponding figures were 3.85 and 4.12 log copies/ml respectively (P = 0.2986 and P = 0.0472 for index and peak values). 85% of those who had significant CMV PCR positivity with the campath/fludarabine/melphalan regimen developed more than one such episode, while 50% of those receiving cyclophosphamide/TBI regimen did so (P = 0.491). Significant CMV PCR positivity was associated with symptoms in a proportion of patients (pyrexia 45%, cough 18%, rise in AST 72%). No patient developed overt CMV disease. CMV PCR is useful for guiding pre-emptive anti-CMV therapy and for monitoring response. PMID- 11277179 TI - Severe cardiac toxicity in hematological stem cell transplantation: predictive value of reduced left ventricular ejection fraction. AB - Eighty patients receiving hematological stem cell transplantation (HCT) with a preparative regimen consisting of total body irradiation (12.5 Gy), cyclophosphamide (4000 or 4500 mg/m2), and thiotepa (400 mg/m2) were evaluated for the development of cardiac toxicity. Patients in whom the pretransplant cumulative dose of anthracycline was more than or equal to 300 mg/m2 showed a lower left ventricular ejection fraction (EF) before HCT compared to patients with less than 300 mg/m2 (0.61 +/- 0.09 vs 0.67 +/- 0.06, P = 0.0010). Patients who had undergone more than or equal to six courses of chemotherapy showed a decreased EF before HCT compared to those after less than six courses (0.67 +/- 0.05 vs 0.63 +/- 0.09, P = 0.03). Three of seven patients (43%) whose pretransplant EF had been less than or equal to 0.55 developed severe cardiac toxicity, characterized by congestive heart failure (CHF) compared with none of 83 patients (0%) whose pretransplant EF had been more than 0.55 (P = 0.00026). Of the three patients who developed severe cardiac toxicity, two were given more than 300 mg/m2 of cumulative anthracycline and underwent 23 courses and six courses of chemotherapy, while the other patient received only two courses of chemotherapy with a total dose of 139 mg/m2 of anthracycline. These results indicate that an increased cumulative dose of anthracycline and number of chemotherapy treatments are correlated with a decrease of the EF and that the EF before HCT is useful for predicting the risk of cardiac complications for recipients who have received chemotherapy. PMID- 11277180 TI - Engraftment-associated hypophosphatemia--the role of cytokine release and steep leukocyte rise post stem cell transplantation. AB - Hypophosphatemia associated with bone marrow transplantation has been infrequently reported. The suggested mechanism is phosphate uptake by the replicating cells. Various cytokines are associated with the development of hypophosphatemia. The present study evaluated the interrelationship between cytokine release, the rise in WBC and the development of hypophosphatemia during the engraftment period. Blood samples were obtained from 60 patients undergoing peripheral blood stem cell transplant, on the day of admission and then daily from the day of transplant until discharge. Hypophosphatemia developed in 62% of the patients. The median day of minimal phosphorus level was +8 and it antedated engraftment by 2 days. There was a significant correlation between the day of minimal phosphorus level and the day of maximal WBC and a significant correlation between the fall in phosphorus level and WBC rise. IL-6 and IL-8 showed similar kinetics. Higher IL-6 and IL-8 levels were directly associated with lower phosphorus levels. In conclusion, hypophosphatemia commonly occurs in the post transplant period. We assume that both a direct effect of cytokine release and an increased consumption by the dividing WBCs contribute to its appearance. As its occurrence usually antedates engraftment it can be used as a forerunner for WBC recovery. PMID- 11277181 TI - Renal function in long-term survivors of stem cell transplantation in childhood. A prospective trial. AB - The aim of this prospective study was to assess glomerular and tubular renal function before, and 1 and 2 years after hematological stem cell therapy (HSCT) in children and adolescents. 137 consecutive patients undergoing HSCT, for malignant diseases, were included in a prospective trial. Forty-four patients were followed for up to 1 year after HSCT and 36 for up to 2 years, without relapse. Ninety healthy school children were used as a control group. The following parameters were investigated: inulin clearance (GFR), urinary excretion of albumin, alpha1-microglobulin (alpha1-MG), calcium, beta-N acetylglucosaminidase (beta-NAG) and Tamm-Horsfall protein (THP), tubular phosphate reabsorption (TP/Cl(cr)) and percent reabsorption of amino acids (TAA). Significantly lower GFR was found 1 and 2 years after HSCT but within the normal range in the period before HSCT. There was no correlation between GFR within the first month after HSCT and long-term outcome of GFR. Tubular dysfunction was found in 14-45% of patients 1 and 2 years after HSCT depending on the parameter investigated. Pathological values 1 and 2 years after HSCT were found for alpha1 MG excretion in 40% and 39%, respectively, for TP/Cl(cr) in 44% and 45%, for beta NAG in 26% and 19%. Median TP/Cl(cr) was significantly lower 2 years after HSCT than before. TAA was mildly impaired in 7/14 patients before, in 5/29 one and in 9/29 2 years after HSCT, but median TAA was within normal range at all times. The median excretion of albumin, THP and calcium was within the normal range at all investigations. No influence of ifosfamide pre-treatment on the severity of tubulopathy was found. The investigation of tubular renal function should be part of a long-term follow-up in children after HSCT. PMID- 11277182 TI - Treatment of post-transplant lymphoproliferative disease with induction chemotherapy followed by haploidentical peripheral blood stem cell transplantation and Rituximab. AB - Management of monoclonal lymphoproliferative disease following stem cell transplantation is difficult and previous attempts to eradicate tumor using chemotherapy or radiation therapy alone have not been successful. We report successful early eradication of an EBV negative, B cell non-Hodgkin's lymphoma in a child who received a T cell-depleted, maternal haploidentical bone marrow transplant for severe combined immunodeficiency disease. Our treatment strategy involved combining conventional induction chemotherapy with re-transplantation using the paternal donor as a source of peripheral blood stem cells, followed by treatment with anti-CD 20 monoclonal antibody (Rituximab). This strategy exploits the potential graft-versus-tumor activity of the mature T cells in the graft, while providing a source of stem cells to confer long-term immune function. The administration of Rituximab in the early post-transplant course may provide additional anti-tumor activity without affecting the new stem cell compartment. PMID- 11277183 TI - Intense immunosuppression followed by purified blood CD34+ cell autografting in a patient with refractory juvenile rheumatoid arthritis. AB - A 15-year-old boy with refractory juvenile rheumatoid arthritis (JRA) underwent intense immunosuppressive therapy followed by purified blood CD34+ cell autografting. He had been taking prednisolone (PDN) daily or every other day combined with methotrexate once a week to control the disease for 7 years. He suffered from psychological complications and a very short stature due to the adverse effects of these drugs. CD34+ cells were purified in bulk from G-CSF mobilized PBSC using an Isolex 300. After the administration of cyclophosphamide (200 mg/kg) and anti-lymphocyte globulin (45 mg/kg), 3.6 x 10(6)/kg purified CD34+ cells were infused. His post-transplant course was uneventful except for herpes-zoster infection. He is now more than 1 year post transplant and has not taken any immunosuppressive medication. His rate of growth has increased (>10 cm/year) due to the effects of the cessation of PDN and the administration of recombinant human growth hormone (rGH), in contrast to the gain of 2 cm in the preceding 3 years with rGH treatment. Although the durability of this remission is unknown, intense immunosuppressive therapy followed by purified blood CD34+ cell autografting might be acceptable for adolescent patients with refractory JRA to achieve a drug-free period for physical and psychological maturation. PMID- 11277184 TI - Successful treatment of thrombocytopenia and hemolytic anemia with IvIG in a patient with lupus-like syndrome after mismatched related PBSCT. AB - Hematopoietic stem cell transplantation (HSCT) is a treatment option for autoimmune diseases but can also cause clinical features similar to those of autoimmune diseases. In some of these cases the autoimmune-like condition is associated with autoimmune cytopenia, a complication that can be unresponsive to established treatment strategies and which may be fatal. The majority of cases reported on immune hemolytic anemia have been of alloimmune origin due to ABO red blood cell antigen incompatibilities between donor and recipient. We now report a patient with a lupus-like syndrome, presenting with severe thrombocytopenia and hemolytic anemia 9 months after HLA-mismatch, ABO compatible-related PBSCT who experienced no response to high-dose steroids, but who had a sustained response to repeated IvIG therapy. PMID- 11277185 TI - Four patients with AL amyloidosis treated with high-dose chemotherapy and autologous stem cell transplantation. AB - Four patients with AL amyloidosis underwent high-dose chemotherapy and autologous stem cell transplantation at our institutions. Here, we report the clinical courses and outcomes in these patients. Two patients with multi-organ amyloid deposits including cardiac involvement died within 12 days after high-dose chemotherapy. However, in the other two patients, one of whom was suffering from amyloid-related cardiac failure, a significant improvement of organ function was achieved. PMID- 11277186 TI - Preoperative high-dose chemotherapy with peripheral blood stem cell support as a primary management of locally advanced breast cancer. AB - Locally advanced breast cancer (LABC) has a poor prognosis with a high risk of local recurrence and distant metastasis. Preoperative combined chemotherapy with or without granulocyte colony-stimulating factor (G-CSF) support does not improve the long-term survival rate. In our report, two patients with LABC received preoperative high-dose chemotherapy with peripheral blood stem cell support (HDC/PBSCs). Prior to high-dose chemotherapy with peripheral blood stem cell support, they both received induction chemotherapy of cyclophosphamide, epirubicin and 5-fluorouracil (FEC) for two cycles which resulted in a partial response. PBSC mobilization and collection were carried out following the second cycle of induction chemotherapy followed by G-CSF. High-dose cyclophosphamide (2500 mg/m2), carboplatin (600 mg/m2) and etoposide (600 mg/m2) were administered with PBSC support. A radical mastectomy was performed followed by adjuvant chemotherapy with FEC regimen for four cycles, followed by local irradiation, and endocrine therapy with tamoxifen. Both patients achieved a remarkable response in the primary lesion after HDC/PBSCs and tolerated the whole treatment well. Preoperative HDC/PBSCs as a new strategy in the treatment of LABC seems practical but it needs to be studied more deeply. PMID- 11277187 TI - High-dose chemotherapy with autologous PBSCT as post-remission therapy for AML. PMID- 11277189 TI - Comparison of four methods for determination of total protein concentrations in pleural and peritoneal fluid from dogs. AB - OBJECTIVE: To compare 4 techniques for determination of total protein concentrations in peritoneal and pleural effusions from dogs. SAMPLE POPULATION: 23 peritoneal and 12 pleural fluid samples from 35 dogs with various abnormalities. PROCEDURE: Samples were collected into tubes containing EDTA, centrifuged, and stored at -20 C until total protein concentrations were assessed. Protein concentration in each sample was determined by use of urine test strips, refractometry, and Bradford and biuret techniques. Accuracy of each method was determined, using dilutions of human control sera. RESULTS: There was good correlation among results of all quantitative procedures. Results of the biuret technique were more accurate than results of the Bradford assay. Refractometry underestimated protein concentration in samples with < 20 g of protein/L. Results of urine test strips correctly classified effusion samples into 2 groups on the basis of total protein concentrations less than or greater than 20 g/L. CONCLUSIONS AND CLINICAL RELEVANCE: Results of any of these 4 techniques can be used to rapidly and efficiently differentiate peritoneal and pleural fluid from dogs into transudates and exudates on the basis of total protein concentration less than or greater than 20 g/L, respectively. PMID- 11277188 TI - Cytotoxic effects of peroxynitrite, polymorphonuclear neutrophils, free-radical scavengers, inhibitors of myeloperoxidase, and inhibitors of nitric oxide synthase on bovine mammary secretory epithelial cells. AB - OBJECTIVE: To determine cytotoxic effects of activated polymorphonuclear neutrophils (PMN) and peroxynitrite on bovine mammary secretory epithelial cells before and after addition of nitric oxide synthase inhibitors, myeloperoxidase (MPO) inhibitors, and free-radical scavengers. SAMPLE POPULATION: Polymorphonuclear neutrophils from 3 lactating cows. PROCEDURE: Cells from the bovine mammary epithelial cell line MAC-T were cultured. Monolayers were treated with activated bovine PMN, lipopolysaccharide (LPS), phorbol 12-myristate 13 acetate (PMA), 3-morpholino-sydnonimine (SIN-1), 4-amino-benzoic acid hydrazide (ABAH), NG-monomethyl-L-arginine, histidine, and superoxide dismutase (SOD). At 24 hours, activity of lactate dehydrogenase in culture medium was used as a relative index of cell death. Tyrosine nitration of proteins in MAC-T cell lysates was determined by visual examination of immunoblots. RESULTS: Lipopolysaccharide, PMA, and < or = 0.1 mM SIN-1 were not toxic to MAC-T cells. Activated PMN, > or = 6 mg of histidine/ml, and 0.5 mM SIN-1 were toxic. Together, histidine and 500,000 activated PMN/ml also were toxic. NG-monomethyl-L arginine did not have an effect, but ABAH decreased PMN-mediated cytotoxicity. Ten and 50 U of SOD/ml protected MAC-T cells from cytotoxic effects of 0.5 mM SIN 1. Compared with control samples, nitration of MAC-T tyrosine residues decreased after addition of 500,000 PMN/ml or > or = 6 mg of histidine/ml. Superoxide dismutase increased and SIN-1 decreased tyrosine nitration of MAC-T cell proteins in a dose-responsive manner. CONCLUSIONS AND CLINICAL RELEVANCE: Peroxynitrite, MPO, and histidine are toxic to mammary secretory epithelial cells. Superoxide dismutase and inhibition of MPO activity mitigate these effects. Nitration of MAC T cell tyrosine residues may be positively associated with viability. PMID- 11277190 TI - Reduction of diameter of the left ventricle of dogs by plication of the left ventricular free wall. AB - OBJECTIVE: To determine effects of reducing the diameter of the left ventricle of dogs by plication of the left ventricular free wall. ANIMALS: 8 healthy adult mixed-breed dogs. PROCEDURE: Left lateral thoracotomy and a T-shaped pericardiotomy were performed. The free wall of the left ventricle was imbricated with 3 interrupted transfixing sutures applied in a horizontal mattress pattern, using 3-0 polypropylene suture assembled on a straight cutting needle. Surgeons were careful to avoid the coronary vessels. Echocardiography was performed 24 hours before and 48 hours after surgery. Electrocardiography was performed before and 1, 2, 7, 15, 21, 30, and 60 days after surgery. RESULTS: Echocardiographic measurements revealed that the diameter of the left ventricle was reduced by a mean of 23.5%. Electrocardiography revealed ventricular premature complexes 24 hours after surgery that regressed without treatment during the first week after surgery. CONCLUSIONS AND CLINICAL RELEVANCE: Plication of the left ventricular free wall of dogs can reduce end-diastolic and end-systolic dimensions of the left ventricle. The technique is simple and does not require cardiopulmonary bypass. According to Laplace's law, the reduction of cardiac diameter leads to reduction on free-wall tension and may improve left ventricular function in dilatated hearts. Thus, additional studies involving dogs with dilated cardiomyopathy should be conducted. PMID- 11277191 TI - Methodology and validity of assessing kinematics of the thoracolumbar vertebral column in horses on the basis of skin-fixated markers. AB - OBJECTIVE: To determine the validity of using skin-fixated markers to assess kinematics of the thoracolumbar vertebral column in horses. ANIMALS: 5 Dutch Warmblood horses without abnormalities of the vertebral column. PROCEDURE: Kinematics of T6, T10, T13, T17, L1, L3, L5, S3, and both tuber coxae were determined by use of bone-fixated and skin-fixated markers. Three-dimensional coordinate data were collected while horses were walking and trotting on a treadmill. Angular motion patterns were calculated and compared on the basis of 2 dimensional analysis of data from skin-fixated markers and 3-dimensional analysis of data from bone-fixated markers. RESULTS: Flexion-extension of thoracolumbar vertebrae and axial rotation of the sacrum were satisfactorily determined at both the walk and trot, using skin-fixated markers. Data from skin-fixated markers were accurate for determining lateral bending at the walk in the midthoracic and lower lumbar portion of the vertebral column only. However, at the trot, data from skin-fixated markers were valid for determining lateral bending for all thoracolumbar vertebrae. CONCLUSIONS AND CLINICAL RELEVANCE: Caution should be taken when interpreting data obtained by use of skin-fixated markers on lateral bending motions during the walk in horses. For determination of other rotations at the walk and all rotations at the trot, use of skin-fixated markers allows valid calculations of kinematics of the vertebral column. Understanding to what extent movements of skin-fixated markers reflect true vertebral motion is a compulsory step in developing noninvasive methods for diagnosing abnormalities of the vertebral column and related musculature in horses. PMID- 11277192 TI - Evaluation of the safety of an abbreviated course of injections of allergen extracts (rush immunotherapy) for the treatment of dogs with atopic dermatitis. AB - OBJECTIVE: To evaluate the safety of an abbreviated course of injections of allergen extracts (rush immunotherapy) for the treatment of dogs with atopic dermatitis. ANIMALS: 30 dogs with atopic dermatitis examined at a veterinary dermatology referral practice for treatment with allergen-specific immunotherapy. PROCEDURE: A catheter was placed in a vein in each dog. Dogs were constantly observed throughout the procedure. Allergen extracts were administered in increasing concentrations every 30 minutes for 6 hours to a maintenance concentration of 20,000 protein nitrogen units/ml. Epinephrine, oxygen, and emergency treatment were available as needed. RESULTS: In 22 (73%) dogs, rush immunotherapy safely replaced the prolonged induction period (15 weeks) of weekly injections that consists of increasing concentrations of allergen extract. In 7 (23%) dogs, the induction period was abbreviated to 4 weeks. Of the 8 dogs that developed problems during rush immunotherapy, increased pruritus necessitated premature cessation of rush immunotherapy in 7, and 1 developed generalized wheals. Oral administration of prednisolone (1 mg/kg of body weight) resulted in resolution of adverse effects in all 8 dogs. CONCLUSION AND CLINICAL RELEVANCE: Rush immunotherapy performed by personnel at a veterinary hospital is a safe method for treatment of dogs with atopic dermatitis. PMID- 11277194 TI - Estimation of the probability for exceeding a threshold concentration of furosemide at various intervals after intravenous administration in horses. AB - OBJECTIVE: To estimate the probability for exceeding a threshold concentration of furosemide commonly used for regulatory purposes after IV administration of furosemide in horses. ANIMALS: 12 mature healthy horses (6 Thoroughbreds and 6 Quarter Horses). PROCEDURE: Venous blood was collected from each horse prior to and 0.25, 0.5, 0.75, 1, 2, 3, 4, 4.5, 5, and 6 hours after administration of 250 or 500 mg of furosemide. Concentrations of furosemide were determined, using an ELISA. Concentration of furosemide was modeled as a function of time, accounting for inter- and intrahorse variabilities. On the basis of pharmacokinetic data, the probability for exceeding a concentration of 100 ng/ml as a function of time was determined, using a semiparametric smooth functional averaging method. A bootstrap approach was used to assess inherent variation in this estimated probability. RESULTS: The estimated probability of exceeding the threshold of 100 ng of furosemide/ml ranged from 11.6% at 4 hours to 2.2% at 5.5 hours after IV administration of 250 mg of furosemide/horse and 34.2% at 4 hours to 12.3% at 5.5 hours after IV administration of 500 mg of furosemide/horse. The probability of a horse being falsely identified in violation of regulatory concentrations was inversely associated with time and positively associated with dose. CONCLUSIONS AND CLINICAL RELEVANCE: Interhorse variability with respect to pharmacokinetics of furosemide will result in misclassification of some horses as being in violation of regulatory concentrations. PMID- 11277195 TI - Screening method for identification of beta-lactams in bovine urine by use of liquid chromatography and a microbial inhibition test. AB - OBJECTIVE: To develop a multiple-residue screening method for the detection of beta-lactams in bovine urine. ANIMALS: 6 clinically normal Holstein cows and 6 calves. PROCEDURE: Pooled urine obtained from cows was used as a negative-control sample or spiked with varying concentrations of 6 beta-lactam antibiotics. Urine samples were prepared for liquid chromatography by diluting 1 ml of urine with 9 ml of 0.01M KH2PO4, 0.01 M Na2PO4, and filtering. Filtrate (2,000 ml) was eluted with a mobile phase in a gradient program. A fraction corresponding to each beta lactam of interest was collected and evaporated to < 1 ml, and water then was added to achieve a 1 ml volume. The collected fraction was tested, using a microbial inhibition test. Then, calves were fed milk spiked with a mixture of 5 beta-lactam antibiotics at a concentration 40X the FDA tolerance in milk. Three hours following the feeding, urine samples were obtained from the calves and tested, as described for the urine samples for the cows. RESULTS: The lowest concentrations of amoxicillin, ampicillin, cephapirin, cloxacillin, desfuroylceftiofurcysteine, and penicillin G that were consistently detected in urine were 100, 10, 100, 250, 1,000, and 10 ng/ml, respectively. Amoxicillin, ampicillin, cephapirin, cloxacillin, desacetylcephapirin, and penicillin G were detected in urine samples of 6/6, 5/6, 0/6, 6/6, 2/6, and 3/6 calves respectively, fed antibiotic-spiked milk. CONCLUSIONS AND CLINICAL RELEVANCE: The integrated method described can be used to detect or identify beta-lactam antibiotics in bovine urine. This method can be used to test cattle for beta lactam residues. PMID- 11277193 TI - Respiratory reflexes in spontaneously breathing anesthetized dogs in response to nasal administration of sevoflurane, isoflurane, or halothane. AB - OBJECTIVE: To characterize respiratory reflexes elicited by nasal administration of sevoflurane (Sevo), isoflurane (Iso), or halothane (Hal) in anesthetized dogs. ANIMALS: 8 healthy Beagles. PROCEDURE: A permanent tracheostomy was created in each dog. Two to 3 weeks later, dogs were anesthetized by IV administration of thiopental and alpha-chloralose. Nasal passages were isolated such that inhalant anesthetics could be administered to the nasal passages while the dogs were breathing 100% O2 via the tracheostomy. Respiratory reflexes in response to administration of each anesthetic at 1.2 and 2.4 times the minimum alveolar concentration (MAC) and the full vaporizer setting (5%) were recorded. Reflexes in response to administration of 5% of each anesthetic also were recorded following administration of lidocaine to the nasal passages. RESULTS: Nasal administration of Sevo, Iso, and Hal induced an immediate ventilatory response characterized by a dose-dependent increase in expiratory time and a resulting decrease in expired volume per unit of time. All anesthetics had a significant effect, but for Sevo, the changes were smaller in magnitude. Responses to administration of each anesthetic were attenuated by administration of lidocaine to the nasal passages. CONCLUSIONS AND CLINICAL RELEVANCE: Nasal administration of Sevo at concentrations generally used for mask induction of anesthesia induced milder reflex inhibition of breathing, presumably via afferent neurons in the nasal passages, than that of Iso or Hal. Respiratory reflexes attributable to stimulation of the nasal passages may contribute to speed of onset and could promote a smoother induction with Sevo, compared with Iso or Hal. PMID- 11277196 TI - Influence of age and sex on plasma lipid and lipoprotein concentrations and associated enzyme activities in cats. AB - OBJECTIVE: To determine effects of age and sex on plasma lipid and lipoprotein metabolism in cats. ANIMALS: 33 kittens and 16 adolescent, 23 adult, and 10 senior cats. PROCEDURE: Plasma concentrations of cholesterol, triglyceride, and lipoprotein-cholesterol and activities of lipoprotein lipase, hepatic lipase, and lecithin:cholesterol acyl transferase (LCAT) were measured and compared within and among groups. RESULTS: Plasma cholesterol and triglyceride concentrations were significantly higher in 5- and 7-week-old kittens, compared with the same kittens after weaning and cats in the other age groups. Cholesterol concentration was significantly less in 20-week-old kittens, compared with adolescent and adult cats. Lipid and lipoprotein-cholesterol concentrations were not significantly different among the adolescent, adult, and senior groups, nor did sex influence lipid and lipoprotein-cholesterol concentrations in these groups. Activities of lipoprotein and hepatic lipases were significantly less in senior cats, compared with the other groups. Activity of LCAT was highest in 20-week-old kittens and was greater in sexually intact adult and adolescent females, compared with their male counterparts. After castration, activities of hepatic lipase and LCAT significantly decreased in adolescent male cats. CONCLUSIONS AND CLINICAL RELEVANCE: The upper limits of reference ranges for plasma cholesterol and triglyceride concentrations should be increased for kittens < 8 weeks of age. Low cholesterol concentrations in adolescent cats likely reflect high tissue demands for growth and steroidogenesis. Decrease in lipoprotein and hepatic lipase activity in senior cats could predispose this age group to hypertriglyceridemia, particularly in insulin-resistant cats or those fed a high fat diet. PMID- 11277197 TI - Comparative efficacy of tricaine methanesulfonate and clove oil for use as anesthetics in red pacu (Piaractus brachypomus). AB - OBJECTIVE: To compare the anesthetic efficacy and physiologic changes associated with exposure to tricaine methanesulfonate and clove oil (100% eugenol). ANIMALS: 15 adult cultured red pacu (Piaractus brachypomus). PROCEDURE: Fish were exposed to each of 6 anesthetic concentrations in a within-subjects complete crossover design. Stages of anesthesia and recovery were measured, and physiologic data were collected before and during anesthesia. RESULTS: Interval to induction was more rapid and recovery more prolonged in fish exposed to eugenol, compared with those exposed to tricaine methanesulfonate. The margin of safety for eugenol was narrow, because at the highest concentration, most fish required resuscitation. Mixed venous-arterial PO2 consistently decreased with anesthesia, while PCO2 consistently increased with anesthesia in all fish regardless of anesthetic agent. The increase in PCO2 was accompanied by a decrease in pH, presumably secondary to respiratory acidosis. Anesthesia was associated with increased blood glucose, potassium, and sodium concentrations as well as Hct and hemoglobin. Fish anesthetized with eugenol were more likely to react to a hypodermic needle puncture than fish anesthetized with tricaine methanesulfonate. CONCLUSIONS AND CLINICAL RELEVANCE: Anesthesia induced with tricaine methanesulfonate or eugenol contributes to hypoxemia, hypercapnia, respiratory acidosis, and hyperglycemia in red pacu. Similar to tricaine methanesulfonate, eugenol appears to be an effective immobilization compound, but eugenol is characterized by more rapid induction, prolonged recovery, and a narrow margin of safety. Care must be taken when using high concentrations of eugenol for induction, because ventilatory failure may occur rapidly. In addition, analgesic properties of eugenol are unknown. PMID- 11277198 TI - Role of excessive maternal iron in the pathogenesis of congenital leukoencephalomalacia in captive black rhinoceroses (Diceros bicornis). AB - OBJECTIVE: To investigate the possibility that excessive maternal iron (overload) may contribute to development of congenital leukoencephalomalacia in captive black rhinoceroses. SAMPLE POPULATION: Tissue specimens and serum samples from 18 rhinoceroses in 2 kindreds harboring 4 (possibly 5) affected female calves. PROCEDURE: Fresh and archival sera and necropsy tissue specimens were evaluated to determine the nature and extent of iron overload in captive and wild black rhinoceroses as well as other rhinoceros species. RESULTS: Quantitative serum and tissue assays of iron and iron analytes, corroborated by histopathologic findings, indicated that these kindreds carried the greatest body burdens of iron yet found among captive black rhinoceroses. Fourteen of 18 rhinoceroses had the highest serum ferritin concentrations measured among 64 black rhinoceroses in captivity in the United States. Dams of affected calves had serum ferritin concentrations 2 orders of magnitude higher than clinically normal humans, equids, or free-ranging rhinoceroses. A neonatal serum sample from 1 affected female calf had a high ferritin concentration (approx 100-fold increase), but a male sibling of another affected female did not, suggesting a possible sex disparity in fetal response to maternal iron overload. Morphologic hallmarks of hemochromatosis were prominent in dams and grandams of affected calves. CONCLUSIONS AND CLINICAL RELEVANCE: Excessive maternal iron may affect female fetuses more than males, possibly inducing leukoencephalomalacia by catalyzing production of highly toxic hydroxyl free radicals during crucial periods of in utero development. Reduction of maternal iron overload may decrease the probability of developing leukoencephalomalacia and some other disorders commonly affecting rhinoceroses in captivity. PMID- 11277199 TI - Petrographic and geochemic evaluation of equine enteroliths. AB - OBJECTIVE: To characterize the texture, mineralogic features, and chemical features of enteroliths obtained from horses. SAMPLE POPULATION: Enteroliths from 13 horses with colic. PROCEDURE: Enteroliths were harvested from 13 horses that underwent ventral midline celiotomy for treatment of colic or necropsy because of colonic obstruction and rupture caused by enteroliths. Dietary and environmental history were determined via questionnaires or evaluation of medical records. In 7 horses that underwent surgical treatment for enterolithiasis, samples of colonic contents were obtained via an enterotomy in the pelvic flexure. Colonic concentrations of magnesium (Mg), phosphorus (P), sulfur (S), sodium (Na), calcium (Ca), and potassium (K) were determined. Enteroliths were analyzed via electron microprobe analysis and X-ray diffraction. RESULTS: Enteroliths varied widely regarding degree of porosity, presence and distribution of radiating texture, and composition and size of the central nidus. A distinct concentric banding was identifiable in all enteroliths. Struvite was the predominant component of all enteroliths, although Mg vivianite was identified in 5 enteroliths, and there were variable quantities of Na, S, K, and Ca in the struvite within enteroliths. Despite an abundance of Ca in colonic fluids, Mg phosphate minerals were preferentially formed, compared with Ca-phosphates (apatite), in equine enteroliths. CONCLUSIONS AND CLINICAL RELEVANCE: Enteroliths comprise 2 major Mg phosphates: struvite and Mg vivianite. There is wide variability in macrotexture and ionic concentrations between and within enteroliths. PMID- 11277200 TI - Comparison of detomidine and romifidine as premedicants before ketamine and halothane anesthesia in horses undergoing elective surgery. AB - OBJECTIVE: To compare detomidine hydrochloride and romifidine as premedicants in horses undergoing elective surgery. ANIMALS: 100 client-owned horses. PROCEDURE: After administration of acepromazine (0.03 mg/kg, IV), 50 horses received detomidine hydrochloride (0.02 mg/kg of body weight, IV) and 50 received romifidine (0.1 mg/kg, IV) before induction and maintenance of anesthesia with ketamine hydrochloride (2 mg/kg) and halothane, respectively. Arterial blood pressure and blood gases, ECG, and heart and respiratory rates were recorded. Induction and recovery were timed and graded. RESULTS: Mean (+/- SD) duration of anesthesia for all horses was 104 +/- 28 minutes. Significant differences in induction and recovery times or grades were not detected between groups. Mean arterial blood pressure (MABP) decreased in both groups 30 minutes after induction, compared with values at 10 minutes. From 40 to 70 minutes after induction, MABP was significantly higher in detomidine-treated horses, compared with romifidine-treated horses, although more romifidine-treated horses received dobutamine infusions. In all horses, mean respiratory rate ranged from 9 to 11 breaths/min, PaO2 from 200 to 300 mm Hg, PaCO2 from 59 to 67 mm Hg, arterial pH from 7.33 to 7.29, and heart rate from 30 to 33 beats/min, with no significant differences between groups. CONCLUSIONS AND CLINICAL RELEVANCE: Detomidine and romifidine were both satisfactory premedicants. Romifidine led to more severe hypotension than detomidine, despite administration of dobutamine to more romifidine-treated horses. Both detomidine and romifidine are acceptable alpha2 adrenoceptor agonists for use as premedicants before general anesthesia in horses; however, detomidine may be preferable when maintenance of blood pressure is particularly important. PMID- 11277201 TI - von Willebrand disease phenotype and von Willebrand factor marker genotype in Doberman Pinschers. AB - OBJECTIVE: To define the relationship between clinical expression of a type-1 von Willebrand disease phenotype and genotype at 2 von Willebrand factor marker loci in Doberman Pinschers. ANIMALS: 102 client-owned Doberman Pinschers. PROCEDURES: Dogs were recruited on the basis of plasma von Willebrand factor concentration, clinical history, and pedigree. Blood samples and response to a history questionnaire were obtained for each dog. Plasma von Willebrand factor concentration was measured by use of an ELISA, and genotyping was performed via polymerase chain reaction for 1 intragenic and 1 extragenic von Willebrand factor marker. Amplification product size was determined by use of polyacrylamide gel electrophoresis (intragenic marker) or automated sequence analysis (extragenic marker). Western blots were prepared from a subset of dogs with low plasma von Willebrand factor concentration to evaluate multimer distribution. RESULTS: Strong associations were detected between plasma von Willebrand factor concentration and von Willebrand factor marker genotype. Twenty-five dogs had substantial reduction in plasma von Willebrand factor concentration and multiple hemorrhagic events. All were homozygous for a 157-base-pair intragenic marker allele and homozygous or compound heterozygous for 1 of 4 extragenic marker alleles. These marker genotypes were exclusively detected in dogs with low plasma von Willebrand factor concentration, although some dogs with these genotypes did not have abnormal bleeding. CONCLUSIONS AND CLINICAL RELEVANCE: Type-1 von Willebrand disease in Doberman Pinschers is associated with the von Willebrand factor gene locus; however, the expression pattern in this breed appears more complex than that of a simple recessive trait. PMID- 11277202 TI - Antioxidant prevention of Heinz body formation and oxidative injury in cats. AB - OBJECTIVE: To determine the effectiveness of 3 antioxidants in preventing Heinz body anemia in cats. DESIGN: Prospective study. ANIMALS: 44 specific-pathogen free healthy cats. PROCEDURE: Cats were housed individually, divided randomly into 4 groups, and given the following orally every 12 hours: empty gelcaps (control cats), N-acetylcysteine (NAC, 100 mg/kg of body weight), vitamin E (d,l alpha-tocopherol; 400 IU), or ascorbate (250 mg). After 2 weeks, Heinz bodies were induced by dietary onion powder (OP; 1% or 3% of dry matter) or propylene glycol (PG, 8% wt/vol in drinking water) for an additional 3 weeks. Intake of treated water or food was recorded daily. Body weight, PCV, Heinz body and reticulocyte percentages, reduced glutathione concentration, and total antioxidant status were measured twice weekly in all cats. RESULTS: Heinz body percentage and degree of anemia did not differ significantly among cats receiving antioxidants and control cats except in cats that ingested water containing PG, in which antioxidant supplementation was associated with a decrease in water intake. Of cats that were fed a diet that contained OP, cats that received NAC had significantly higher reduced glutathione concentrations, compared with other cats in the experiment. Total antioxidant status did not consistently correlate with antioxidant supplementation or type of oxidant administered (ie, OP or PG). CONCLUSIONS AND CLINICAL RELEVANCE: Although the effect of antioxidant supplementation on Heinz body anemia in cats was minimal, antioxidants may have subclinical biochemical effects such as GSH sparing that may be important against milder forms of oxidative stress. PMID- 11277203 TI - Effects of the angiotensin converting enzyme inhibitor benazepril in cats with induced renal insufficiency. AB - OBJECTIVE: To determine effects of the angiotensin converting enzyme inhibitor benazepril in cats with induced renal insufficiency. ANIMALS: 32 cats. PROCEDURE: Renal mass was surgically reduced, and cats were assigned to 1 of 4 eight-cat groups. Group 1 received placebo, whereas groups 2, 3, and 4 received benazepril hydrochloride orally once daily for approximately 6.5 months at the following doses: group 2, 0.25 to 0.50 mg/kg of body weight; group 3, 0.50 to 1.00 mg/kg; and group 4, 1.00 to 2.00 mg/kg. Arterial blood pressures, glomerular filtration rate (GFR), and renal plasma flow were determined before treatment and during the treatment period. Other determinants of renal hemodynamics were measured by use of micropuncture techniques. Renal biopsy specimens were examined microscopically. RESULTS: Compared with cats that received placebo, mean systolic arterial blood pressure was significantly less and GFR significantly greater in cats that received benazepril. Glomerular capillary pressure and the ratio of efferent to afferent arteriolar vascular resistance were also significantly less in treated cats. However, histologic differences in renal specimens were not detected. CONCLUSIONS AND CLINICAL RELEVANCE: Treatment with benazepril sustained single nephron GFR in remnant nephrons of cats with induced renal insufficiency. Administration of benazepril was also associated with a small but significant reduction in degree of systemic hypertension and an increase in whole kidney GFR. Benazepril may be an effective treatment to slow the rate of progression of renal failure in cats with renal disease. PMID- 11277204 TI - Detection of free radicals in ischemic and reperfused canine gracilis muscle flaps by use of spin-trapping electron paramagnetic resonance spectroscopy. AB - OBJECTIVE: To determine whether free radicals are produced in ischemic and reperfused canine skeletal muscle, whether free radicals can be detected from effluent blood by use of spin-trapping electron paramagnetic resonance (EPR) spectroscopy, and whether free radical-induced skeletal muscle damage is detectable by use of light microscopy. ANIMALS: 6 healthy mixed-breed dogs. PROCEDURES: Dogs were anesthetized and both gracilis muscles were isolated, leaving only the major vascular pedicle intact. Ischemia was induced in 1 flap for 4 hours; the contralateral flap served as the control. Ischemic flaps were then reperfused for 15 minutes. alpha-Phenyl-N-tert-butylnitrone, a spin-trapping agent, was administered intravenously to each dog 1 hour prior to reperfusion. Following reperfusion, effluent blood samples from muscle flaps were obtained and processed for EPR spectroscopy. Muscle biopsy specimens were obtained for histologic evaluation, and dogs were euthanatized. RESULTS: Spin adducts were not detected in blood from control flaps. However, spin adducts were detected in all ischemic-reperfused muscle flaps. Principal signals identified were characteristic of oxygen- and carbon-centered radicals. Significantly more muscle damage was detected in ischemic-reperfused flaps, compared with control flaps. CONCLUSIONS AND CLINICAL RELEVANCE: Free radicals may be an important component of injury induced by ischemia and reperfusion of canine skeletal muscle. Spin trap adducts of free radicals can be detected in effluent blood of canine muscle flaps by use of spin-trapping EPR spectroscopy. Spin-trapping EPR spectroscopy may be useful for the study of antioxidants and free radical scavengers in attenuating ischemia and reperfusion-mediated skeletal muscle damage. PMID- 11277205 TI - Results of intradermal tests in horses without atopy and horses with chronic obstructive pulmonary disease. AB - OBJECTIVE: To compare results of intradermal tests (IDT), conducted using environmental allergens, in horses without atopy and horses with chronic obstructive pulmonary disease (COPD). ANIMALS: 38 horses (22 horses without atopy and 16 horses with COPD). PROCEDURE: All horses were examined (physical examination, hematologic examination, serum biochemical analyses, examination of bronchoalveolar lavage fluid). An IDT was conducted, using a full panel of 73 allergens consisting of grasses, weeds, trees, molds, and insects. Results of the IDT were evaluated 30 minutes and 4, 6, and 24 hours after injection of allergens. Horses without atopy were euthanatized, and gross and histologic changes of lung parenchyma were assessed. RESULTS: Horses without atopy had a greater number of positive immediate and late-phase reactions than did horses with COPD. Horses with COPD did not have a significantly greater number of positive reactions than horses without atopy at any time period for any allergen group (grasses, weeds, trees, molds, and insects). CONCLUSIONS AND CLINICAL RELEVANCE: Positive results of IDT document allergen-specific hypersensitivity but do not necessarily distinguish clinically relevant reactions from subclinical reactivity in horses with COPD. Interpreting the clinical relevance of results of IDT requires a thorough knowledge of the medical history, physical examination findings, and environment of each animal. PMID- 11277206 TI - Bacteriologic and histologic features in mice after intranasal inoculation of Brucella melitensis. AB - OBJECTIVE: To characterize effects of intranasal inoculation of virulent Brucella melitensis strain 16M in mice. ANIMALS: Female Balb/c mice, 6 to 8 weeks old. PROCEDURE: Studies were designed to elucidate gross morphologic lesions, bacterial burden in target organs, and histologic changes in tissues following experimental intranasal inoculation of mice with B melitensis 16M, which could be used to characterize a model for testing vaccine efficacy. RESULTS: Measurable splenomegaly was evident at 3 and 7 weeks after inoculation. A demonstrable increase in splenic colony-forming units (CFU) from infected mice increased over time with increasing dose when comparing inocula of 10(3), 10(4), and 10(5) CFU. Recovery of brucellae from the lungs was possible early in infection with 10(1), 10(3), and 10(5) CFU, but only the group inoculated with 10(5) CFU consistently yielded quantifiable bacteria. At a dose of 10 CFU, few organisms were located in the spleen. Bacteria were recovered up to 140 days after inoculation in mice given 10(3) CFU. At an inoculum of 10(5) CFU, bacterial counts were highest early in infection. Histologic examination of tissues revealed an increase in white pulp and marginal zone in the spleen and lymphohistiocytic hepatitis. CONCLUSION AND CLINICAL RELEVANCE: Changes in the spleen and liver increased with increases in dose and with increased time following intranasal inoculation with B melitensis 16M. Surprisingly, histologic changes were not observed in the lungs of inoculated mice. PMID- 11277207 TI - Clinical evaluation of established optimal immobilizing doses of medetomidine ketamine in captive reindeer (Rangifer tarandus tarandus). AB - OBJECTIVE: To evaluate clinical effects and repeatability of clinical effects for an optimal immobilizing dose of a combination of medetomidine hydrochloride (MED) and ketamine hydrochloride (KET) in reindeer (Rangifer tarandus tarandus). ANIMALS: 12 healthy 6- to 8-month old reindeer. PROCEDURE: Each reindeer was immobilized once with an initial dose (combination of 0.06 mg of MED/kg of body weight and 0.3 mg KET/kg) and twice with an optimal dose of MED-KET. Reversal was achieved with 5 mg of atipamezole/mg of MED injected 45 minutes after MED-KET administration. Observational variables were recorded. Oxygen saturation of arterial hemoglobin measured by pulse oximetry (Spo2), respiratory rate (RR), heart rate (HR), and rectal temperature (RT) were recorded 10, 25, and 40 minutes after immobilization. RESULTS: Mean time to first sign of sedation and time until a recumbent animal lifted its head were significantly reduced for reindeer given the optimal dose, compared with the initial dose. Mean Spo2 remained > 90% during initial immobilization; this value was significantly lower for the optimal dose, but increased during immobilization from 85 to 89%. At all doses, RR increased significantly throughout the recorded period; however, RT and HR were constant. Except for time until reindeer stood, all time variables, Spo2, RR, RT, and HR were repeatable. CONCLUSION AND CLINICAL RELEVANCE: mmobilization of captive reindeer achieved by use of the optimal dose established here is clinically acceptable, although Spo2 should be carefully monitored. Administration of the optimal dose produced the same clinical effect during repeated immobilization of the same reindeer. PMID- 11277209 TI - Cytokeratins of the matrices of the chestnut (torus carpeus) and periople in horses with acute laminitis. AB - OBJECTIVES: To determine whether there is a change in the expression of cytokeratins in the epidermal cells of the non-weight-bearing parts of the limb in horses with acute laminitis and thus determine whether the morphologic changes that develop in the periople and chestnut (torus carpeus) of horses early in acute laminitis are caused by inhibition of keratinocyte differentiation. ANIMALS: 8 horses with acute laminitis. PROCEDURE: Tissue specimens were obtained from the chestnuts of all 8 horses and from the stratum externum of the hoof wall of 3 horses. Tissue specimens were obtained within 48 hours of the first clinical signs of laminitis. The cytokeratins were characterized by 1- and 2-dimensional gel electrophoresis, and the tissue distribution of the cytokeratins was studied by immunohistochemical staining. RESULTS: The biochemical findings indicated that the epidermal cells of tissues from horses affected by laminitis contained the same set of cytokeratins as corresponding tissues from clinically normal horses. Immunohistochemistry on sections from specimens of horses with laminitis versus clinically normal horses indicated a difference in the expression of cytokeratin in the basal cells in the matrix of the stratum externum of the hoof wall and in the matrix of the chestnut of horses with laminitis in which the most severe morphologic changes were observed. CONCLUSIONS: Inhibition of keratinocyte differentiation, as observed by immunohistochemical changes, in cells in parts of the chestnut and periople may indirectly indicate that the observed epidermal changes in horses with laminitis are primary and are unaffected by weight bearing. PMID- 11277208 TI - Effect of contact stress in bones of the distal interphalangeal joint on microscopic changes in articular cartilage and ligaments. AB - OBJECTIVE: To examine articular cartilage of the distal interphalangeal (DIP) joint and distal sesamoidean impar ligament (DSIL) as well as the deep digital flexor tendon (DDFT) for adaptive responses to contact stress. SAMPLE POPULATION: Specimens from 21 horses. PROCEDURE: Pressure-sensitive film was inserted between articular surfaces of the DIP joint. The digit was subjected to a load. Finite element models (FEM) were developed from the data. The navicular bone, distal phalanx, and distal attachments of the DSIL and DDFT were examined histologically. RESULTS: Analysis of pressure-sensitive film revealed significant increases in contact area and contact load at dorsiflexion in the joints between the distal phalanx and navicular bone and between the middle phalanx and navicular bone. The FEM results revealed compressive and shear stresses. Histologic evaluation revealed loss of proteoglycans in articular cartilage from older horses (7 to 27 years old). Tidemark advancement (up to 14 tidemarks) was observed in articular cartilage between the distal phalanx and navicular bone in older clinically normal horses. In 2 horses with navicular syndrome, more tidemarks were evident. Clinically normal horses had a progressive increase in proteoglycans in the DSIL and DDFT. CONCLUSIONS AND CLINICAL RELEVANCE: Load on the navicular bone and associated joints was highest during dorsiflexion. This increased load may be responsible for microscopic changes of tidemark advancement and proteoglycan depletion in the articular cartilage and of proteoglycan production in the DSIL and DDFT Such microscopic changes may represent adaptive responses to stresses that may progress and contribute to lameness. PMID- 11277210 TI - Aberrations of the p53 tumor suppressor gene in various tumors in dogs. AB - OBJECTIVE: To evaluate aberrations of the p53 tumor suppressor gene in naturally developing tumors in dogs. SAMPLE POPULATION: Tumor specimens from 15 dogs with various tumors, including malignant lymphoma (7 dogs), monocytic leukemia (1), mammary gland adenoma (1), mammary gland benign mixed tumor (1), rhabdomyosarcoma (1), colon cancer (1), and osteosarcoma (3). PROCEDURE: Aberrations of the p53 gene in these tumor tissues were examined by reverse transcriptase-polymerase chain reaction and single-strand conformation polymorphism analysis, using 3 fragments that covered the entire open reading frame of the canine p53 gene, followed by nucleotide sequencing of the abnormal bands. RESULTS: Point mutations, deletions, and insertions resulting in a number of amino acid substitutions of wild-type p53 were detected in 7 of the 15 tumor specimens from dogs with malignant lymphoma, monocytic leukemia, rhabdomyosarcoma, colon cancer, and osteosarcoma. Of these 7 dogs, 2 had aberrations of the p53 gene on both alleles, whereas 5 had aberrations of the p53 gene on 1 allele and concurrently lacked the wild-type p53 transcript. Many of the aberrations of the p53 gene detected in these tumors were located in the transactivation, DNA binding, and oligomerization domains. CONCLUSIONS AND CLINICAL RELEVANCE: Various naturally developing tumors in dogs often have inactivation of the p53 tumor suppressor gene, which may be 1 of the multiple step-wise genetic changes during tumorigenesis. This study indicates that p53 gene can be a target for gene therapy for tumors in dogs. PMID- 11277212 TI - Usefulness of dobutamine stress tests for detection of cardiac abnormalities in dogs with experimentally induced early left ventricular dysfunction. AB - OBJECTIVE: To determine whether dobutamine stress tests (DST) can be used to detect cardiac dysfunction in dogs with early left ventricular dysfunction (ELVD) induced by rapid right ventricular pacing (RRVP). ANIMALS: 7 adult male Beagles. PROCEDURE: A pacemaker was surgically implanted in each dog at the level of the right ventricular apex. Electrocardiography, Doppler sphygmomanometry, and Doppler echocardiography were performed before and during a DST prior to activation of the pacemaker and every 3 to 4 days during the period of RRVP. Dobutamine stress tests were performed by infusing dobutamine at incremental dosages ranging from 12.5 to 42.5 microg/kg of body weight/min. RESULTS: Clinical signs of congestive heart failure were not observed during the pacing period. However, all dogs developed ELVD associated with significant changes in values for most Doppler echocardiographic variables obtained prior to DST Adverse cardiac effects were not detected during DST. Most Doppler echocardiographic indices of cardiac function were significantly altered in response to dobutamine infusion during the pacing period, compared with prepacing values. However, a dobutamine-induced 2-fold increase in cardiac output was maintained. CONCLUSIONS AND CLINICAL RELEVANCE: Dobutamine stress tests can be safely performed in dogs with experimentally induced ELVD. Dobutamine stress tests may be a sensitive, noninvasive diagnostic method, complementary to standard clinical examinations, for detection of early cardiac dysfunction in dogs asymptomatic for dilated cardiomyopathy. PMID- 11277211 TI - Effect of increased dietary protein and decreased dietary carbohydrate on performance and body composition in racing Greyhounds. AB - OBJECTIVE: To determine effects of increased dietary protein and decreased dietary carbohydrate on hematologic variables, body composition, and racing performance in Greyhounds. ANIMALS: 8 adult Greyhounds. PROCEDURE: Dogs were fed a high-protein (HP; 37% metabolizable-energy [ME] protein, 33% ME fat, 30% ME carbohydrate) or moderate-protein (MP; 24% ME protein, 33% ME fat, 43% ME carbohydrate) extruded diet for 11 weeks. Dogs subsequently were fed the other diet for 11 weeks (crossover design). Dogs raced a distance of 500 m twice weekly. Rectal temperature, hematologic variables before and after racing, plasma volume, total body water, body weight, average weekly food intake, and race times were measured at the end of each diet period. RESULTS: When dogs were fed the MP diet, compared with the HP diet, values (mean +/- SD) differed significantly for race time (32.43 +/- 0.48 vs 32.61 +/- 0.50 seconds), body weight (32.8 +/- 2.5 vs 32.2 +/- 2.9 kg), Hct before (56 +/- 4 vs 54 +/- 6%) and after (67 +/- 3 vs 64 +/- 8%) racing, and glucose (131 +/- 16 vs 151 +/- 27 mg/dl) and triglyceride (128 +/- 17 vs 104 +/- 28 mg/dl) concentrations after racing. CONCLUSIONS AND CLINICAL RELEVANCE: Greyhounds were 0.18 seconds slower (equivalent to 0.08 m/s or 2.6 m) over a distance of 500 m when fed a diet with increased protein and decreased carbohydrate. Improved performance attributed to feeding meat to racing Greyhounds apparently is not attributable to increased dietary protein and decreased dietary carbohydrate. PMID- 11277213 TI - Rapid SPECT simulation of downscatter in non-uniform media. AB - A rotation-based Monte Carlo (MC) simulation method (RMC) has been developed, designed for rapid calculation of downscatter through non-uniform media in SPECT. A possible application is downscatter correction in dual isotope SPECT. With RMC, only a fraction of all projections of a SPECT study have to be MC simulated in a standard manner. The other projections can be estimated rapidly using the results of these standard MC calculations. For efficiency, approximations have to be made in RMC with regard to the final scatter angle of the detected photons. Further speed-up is obtained by combining RMC with convolution-based forced detection (CFD) instead of forced detection (FD), which is a more common variance reduction technique for MC. The RMC method was compared with standard MC for 99mTc downscatter in a 201Tl window (72 keV+/-10%) using a digital thorax phantom. The resulting scatter projections are in good agreement (maximum bias a few per cent of the largest value in the projection), but RMC with CFD is about three orders in magnitude faster than standard MC with FD and up to 25 times faster than standard MC with CFD. Using RMC combined with CFD, the generation of 64 almost noise-free downscatter projections (64 x 64) takes only a couple of minutes on a 500 MHz Pentium processor. Therefore, rotation-based Monte Carlo could serve as a practical tool for downscatter correction schemes in dual isotope SPECT. PMID- 11277214 TI - Concepts for shuttling multileaf collimators for intensity-modulated radiation therapy. AB - Shuttling multileaf collimators (SMLCs) can increase the MU efficiency of intensity-modulated radiation therapy compared with the multiple-static-field (MSF-MLC) technique or dynamic MLC (DMLC) technique with conventional MLCs. In a previous paper (Phys. Med. Biol. 45 3343-58) a particular SMLC was shown, for highly modulated intensity distributions, to increase the MU efficiency compared with the MSF-MLC technique. In this companion paper, two new arrangements similar to that described in the earlier paper, but with less mechanical complexity, are shown to be constructionally simpler but less MU efficient. Additionally another new concept of SMLC is shown which also increases the MU efficiency compared with the MSF-MLC technique and often improves the MU efficiency compared with the previously reported SMLC for highly modulated intensity distributions. It also leads to zero tongue-and-groove underdose in the direction orthogonal to that of the shuttling elements (so-called across-the-rows). PMID- 11277215 TI - Collimator scatter and 2D dosimetry in small proton beams. AB - Monte Carlo simulations have been performed to determine the influence of collimator-scattered protons from a 150 MeV proton beam on the dose distribution behind a collimator. Slit-shaped collimators with apertures between 2 and 20 mm have been simulated. The Monte Carlo code GEANT 3.21 has been validated against one-dimensional dose measurements with a scintillating screen, observed by a CCD camera. In order to account for the effects of the spatial response of the CCD/scintillator system, the line-spread function was determined by comparison with measurements made with a diamond detector. The line-spread function of the CCD/scintillator system is described by a Gaussian distribution with a standard deviation of 0.22 mm. The Monte Carlo simulations show that protons that hit the collimator on the entrance face and leave it through the wall of the aperture make the largest scatter contribution. Scatter on air is the major contribution to the extent of the penumbra. From the energy spectra it is derived that protons with a relative biological effectiveness greater than 1 cause at most 1% more damage in tissue than what would be expected from the physical dose. PMID- 11277216 TI - Real-time dose calculation and visualization for the proton therapy of ocular tumours. AB - A new real-time dose calculation and visualization was developed as part of the new 3D treatment planning tool OCTOPUS for proton therapy of ocular tumours within a national research project together with the Hahn-Meitner Institut Berlin. The implementation resolves the common separation between parameter definition, dose calculation and evaluation and allows a direct examination of the expected dose distribution while adjusting the treatment parameters. The new tool allows the therapist to move the desired dose distribution under visual control in 3D to the appropriate place. The visualization of the resulting dose distribution as a 3D surface model, on any 2D slice or on the surface of specified ocular structures is done automatically when adapting parameters during the planning process. In addition, approximate dose volume histograms may be calculated with little extra time. The dose distribution is calculated and visualized in 200 ms with an accuracy of 6% for the 3D isodose surfaces and 8% for other objects. This paper discusses the advantages and limitations of this new approach. PMID- 11277217 TI - On-line reconstruction of low boron concentrations by in vivo gamma-ray spectroscopy for BNCT. AB - Boron neutron capture therapy (BNCT) is a radiation therapy in which the neutron capture reaction of 10B is used for the selective destruction of tumours. At the High Flux Reactor (HFR) in Petten, a therapy facility with an epithermal neutron beam has been built. In the first instance, patients with brain tumours will be treated. The doses delivered to the tumour and to the healthy tissue depend on the thermal neutron fluence and on the boron concentrations in these regions. An accurate determination of the patient dose during therapy requires knowledge of these time-dependent concentrations. For this reason, a gamma-ray telescope system, together with a reconstruction formalism, have been developed. By using a gamma-ray detector in a telescope configuration, boron neutron capture gamma-rays of 478 keV emitted by a small specific region can be detected. The reconstruction formalism can calculate absolute boron concentrations using the measured boron gamma-ray detection rates. Besides the boron gamma-rays, a large component of 2.2 MeV gamma-rays emitted at thermal neutron capture in hydrogen is measured. Since the hydrogen distribution is almost homogeneous within the head, this component can serve as a measure of the total number of thermal neutrons in the observed volume. By using the hydrogen gamma-ray detection rate for normalization of the boron concentration, the reconstruction tool eliminates the greater part of the influence of the inhomogeneity of the thermal neutron distribution. MCNP calculations are used as a tool for the optimization of the detector configuration. Experiments on a head phantom with 5 ppm 10B in healthy tissue showed that boron detection with a standard deviation of 3% requires a minimum measuring time of 2 min live time. From two position-dependent measurements, boron concentrations in two compartments (healthy tissue and tumour) can be determined. The reconstruction of the boron concentration in healthy tissue can be done with a standard deviation of 6%. The gamma-ray telescope can also be used for in vivo dosimetry. PMID- 11277218 TI - A film technique for the determination of output factors and end effect times for the Leksell Gamma Knife. AB - The relative output factors of the four helmets for a model B Leksell Gamma Knife and the end effect times for each helmet have been measured. For the three helmets with the smallest-diameter collimators a technique employing Kodak XV-2 film was used. The measured output factors are in good agreement with the values recommended by the manufacturer. The end effect times vary with the collimator size, with the shorter time occurring with the smaller collimator. PMID- 11277219 TI - The measurement of thermal neutron flux depression for determining the concentration of boron in blood. AB - Boron neutron capture therapy (BNCT) is a form of targeted radiotherapy that relies on the uptake of the capture element boron by the volume to be treated. The treatment procedure requires the measurement of boron in the patient's blood. The investigation of a simple and inexpensive method for determining the concentration of the capture element 10B in blood is described here. This method, neutron flux depression measurement, involves the determination of the flux depression of thermal neutrons as they pass through a boron-containing sample. It is shown via Monte Carlo calculations and experimental verification that, for a maximum count rate of 1 x 10(4) counts/s measured by the detector, a 10 ppm 10B sample of volume 20 ml can be measured with a statistical precision of 10% in 32 +/- 2 min. For a source activity of less than 1.11 x 10(11) Bq and a maximum count rate of less than 1 x 10(4) counts/s, a 10 ppm 10B sample of volume 20 ml can be measured with a statistical precision of 10% in 58 +/- 3 min. It has also been shown that this technique can be applied to the measurement of the concentration of any element with a high thermal neutron cross section such as 157Gd. PMID- 11277220 TI - Dose measurements in inhomogeneous bone/tissue and lung/tissue phantoms for angiography using synchrotron radiation. AB - For angiography using synchrotron radiation we measured the absorbed dose distribution in inhomogeneous phantoms with thin LiF:Mg, Cu, P, LiF:Mg, Ti thermoluminescent dosimeters (TLDs) in tissue and lung substitutes, and with Mg2SiO4:Tb TLDs in bone substitute for 33.32 keV monoenergetic photons from synchrotron radiation. The energy responses of the TLDs were measured in air for 10-40 keV monoenergetic photons. The values at 30 keV became smaller by 30% for LiF:Mg, Cu, P and larger by 22% for Mg2SiO4:Tb than the ratio of the mass energy absorption coefficients of the TLDs to that of air. These values were used to modify the calculated response of the TLDs in each phantom material. The absorbed dose distribution obtained was compared with that calculated using the Monte Carlo transport code EGS4 expanded to a low-energy region, and their agreement was confirmed taking linear polarization into account. In the bone substitute the dose increased by a factor of 3.9, while behind the bone the dose decreased drastically because of photon attenuation. In the lung substitute a slight dose difference from that in soft tissue was observed because of its different density. The LiF:Mg, Cu, P TLDs exhibited a better energy response, higher sensitivity and wider linear regions than did the other tissue-equivalent TLDs in the low-energy region. PMID- 11277221 TI - Long-term stability of liquid ionization chambers with regard to their qualification as local reference dosimeters for low dose-rate absorbed dose measurements in water. AB - The long-term sensitivity and calibration stability of liquid ionization chambers (LICs) has been studied at a local and a secondary standards dosimetry laboratory over a period of 3 years. The chambers were transported several times by mail between the two laboratories for measurements. The LICs used in this work are designed for absorbed dose measurements in the dose rate region of 0.1-100 mGy min(-1) and have a liquid layer thickness of 1 mm and a sensitive volume of 16.2 mm3. The liquids used as sensitive media in the chambers are mixtures of isooctane (C8H18) and tetramethylsilane (Si(CH3)4) in different proportions (about 2 to 1). Operating at a polarizing voltage of 300 V the leakage current of the chambers was stable and never exceeded 3% of the observable current at a dose rate of about 1 mGy min(-1). The volume sensitivity of the chambers was measured to be of the order of 10(-9) C Gy(-1) mm3. No systematic changes in the absorbed dose to water calibration was observed for any of the chambers during the test period (sigma < 0.2%). Variations in chamber dose response with small changes in the polarizing voltage as well as sensitivity changes with accumulated absorbed dose were also investigated. Measurements showed that the LIC response varies by 0.15% per 1% change in applied voltage around 300 V. No significant change could be observed in the LIC sensitivity after a single absorbed dose of 15 kGy. The results indicate that the LIC can be made to serve as a calibration transfer instrument and a reference detector for absorbed dose to water determinations providing good precision and long-term reproducibility. PMID- 11277222 TI - Determination of absorbed dose calibration factors for therapy level electron beam ionization chambers. AB - Over several years the National Physical Laboratory (NPL) has been developing an absorbed dose calibration service for electron beam radiotherapy. To test this service, a number of trial calibrations of therapy level electron beam ionization chambers have been carried out during the last 3 years. These trials involved 17 UK radiotherapy centres supplying a total of 46 chambers of the NACP, Markus, Roos and Farmer types. Calibration factors were derived from the primary standard calorimeter at seven energies in the range 4 to 19 MeV with an estimated uncertainty of +/-1.5% at the 95% confidence level. Investigations were also carried out into chamber perturbation, polarity effects, ion recombination and repeatability of the calibration process. The instruments were returned to the radiotherapy centres for measurements to be carried out comparing the NPL direct calibration with the 1996 IPEMB air kerma based Code of Practice. It was found that, in general, all chambers of a particular type showed the same energy response. However, it was found that polarity and recombination corrections were quite variable for Markus chambers-differences in the polarity correction of up to 1% were seen. Perturbation corrections were obtained and were found to agree well with the standard data used in the IPEMB Code. The results of the comparison between the NPL calibration and IPEMB Code show agreement between the two methods at the +/-1% level for the NACP and Farmer chambers, but there is a significant difference for the Markus chambers of around 2%. This difference between chamber types is most likely to be due to the design of the Markus chamber. PMID- 11277223 TI - Quantifying effects of lead shielding in electron beams: a Monte Carlo study. AB - Lead shielding in contact with the patient's skin is often encountered in radiotherapy with electron beams. The influence of the lead shielding on dose distributions in the patient cannot fully be assessed using modern treatment planning systems. In this work the problem of quantifying the effect of lead shielding on dose distributions is addressed. Monte Carlo dose calculations were performed in a half-blocked water phantom shielded by lead, using a realistic model for the fluence of an electron linear accelerator. Electron beam energies of 6-20 MeV and lead thicknesses of 1-7 mm are used for 10 x 10 cm2 and 5 x 5 cm2 fields. The perturbation of the particle fluence and dose distributions in water introduced by the lead shielding is quantified. The effect of oblique electron beams on the dose perturbation is shown. A fictitious clinical example, the shielding of an eye in electron beam treatment, is used to demonstrate the usefulness of Monte Carlo based treatment planning algorithms that can incorporate the effects of lead shielding. PMID- 11277224 TI - Backscatter factors for mammography calculated with Monte Carlo methods. AB - The objective of this study is to establish a comprehensive set of backscatter factors for mammography based on the exposure model proposed by the European Protocol on Dosimetry in Mammography. The Monte Carlo calculated backscatter factors (BSFs) presented in this study are for various exposure conditions encountered in mammographic practice as well as in calibration procedures. The data demonstrate the variation of the BSF as a function of the exposure parameters, hence enabling a better match with calibration conditions and, at the same time, reviewing the BSF data already recommended by the European Protocol. Furthermore, earlier data for BSF for general diagnostic radiology are validated. PMID- 11277225 TI - Narrow stereotactic beam profile measurements using N-vinylpyrrolidone based polymer gels and magnetic resonance imaging. AB - In this work, polymer gel-MRI dosimetry (using VIPAR gels), radiographic film and a PinPoint ion chamber were used for profile measurements of 6 MV x-ray stereotactic beams of 5 and 10 mm diameter. The VIPAR gel-MRI method exhibited a linear dose response up to 32 Gy. VIPAR gels were found to resolve the penumbra region quite accurately, provided that the in-plane image resolution of the related T2-map is adequate (< or = 0.53 mm). T2-map slice thickness had no significant effect on beam profile data. VIPAR measurements performed with a spatial resolution of 0.13 mm provided penumbra widths (80%-20% distance) of 1.34 and 1.70 mm for the 5 and 10 mm cones respectively. These widths were found to be significantly smaller than those obtained with the film (2.23 mm for the 5 mm cone, 2.45 mm for the 10 mm cone) and PinPoint (2.25 mm for the 5 mm cone, 2.52 mm for the 10 mm cone) methods. Regarding relative depth dose measurements, good correlation between VIPAR gel and PinPoint data was observed. In conclusion, polymer gel-MRI dosimetry can provide relatively accurate profile data for very small beams used in stereotactic radiosurgery since it can overcome, to some extent, the problems related to the finite size of conventional detectors. PMID- 11277226 TI - Studies of magnetization transfer and relaxation in irradiated polymer gels- interpretation of MRI-based dosimetry. AB - Magnetization transfer and NMR relaxation rates were measured for water protons in two types of polymer gels developed for radiation dosimetry with MRI in order to quantify the contributions of different relaxation processes to the radiation response in such gels. Measurements included the rate of magnetization transfer between proton pools and the ratio of the sizes of exchanging pools, R1 and R2. A model of relaxation in irradiated gels is presented to explain their properties. The model incorporates three proton pools: free water, macromolecular and interfacial. Two pools are insufficient to model the data. In these systems, radiation-induced polymerization appears to increase the size of a solid-like macromolecular proton pool but does not affect the rate constant of magnetization transfer per proton from macromolecular protons to the free water protons. The relation between R1 and the pool size ratio is consistent with free water exchanging with a macromolecular pool with an R1 of approximately 8 Hz. In addition, the rate of magnetization transfer is not limited by the rate of chemical exchange between the free water and the interfacial protons, and magnetization transfer most probably occurs via labile proton exchange rather than via bound water molecules. PMID- 11277227 TI - Study on examinee's dose delivered in computed tomography. AB - Dose profiles are presented resulting from computed tomography (CT). The profiles are positioned at the central axis, 1 cm away from the outer surface of the phantom, for single and multiple scans. A Hitachi W-1000 scanner is used with a thermoluminescent dosimeter (TLD), and standard dosimetry head and trunk phantoms. Regression equations are found linking the dose resulting from scattered radiation associated with a single scan to the distance from the scanning centre. The impact on the CT dose index value (CTDI) for varying integrating lengths is analysed. Some problems associated with CT dose measurement are noted, which may assist in the practical application of IBSS (International Basic Standard of Radiation Protection and Safety of Radiation Sources) guide levels. PMID- 11277228 TI - Correction of scatter in megavoltage cone-beam CT. AB - The role of scatter in a cone-beam computed tomography system using the therapeutic beam of a medical linear accelerator and a commercial electronic portal imaging device (EPID) is investigated. A scatter correction method is presented which is based on a superposition of Monte Carlo generated scatter kernels. The kernels are adapted to both the spectral response of the EPID and the dimensions of the phantom being scanned. The method is part of a calibration procedure which converts the measured transmission data acquired for each projection angle into water-equivalent thicknesses. Tomographic reconstruction of the projections then yields an estimate of the electron density distribution of the phantom. It is found that scatter produces cupping artefacts in the reconstructed tomograms. Furthermore, reconstructed electron densities deviate greatly (by about 30%) from their expected values. The scatter correction method removes the cupping artefacts and decreases the deviations from 30% down to about 8%. PMID- 11277229 TI - X-ray emission from a compact hot plasma: applications to radiology and mammography. AB - Hot electrons confined in a compact magnetized plasma (in a 1 litre chamber) and heated by electron cyclotron resonance were used to provide an intense reproducible x-ray emission. Since electrons were generated in situ, the use of filaments and high voltages at the base of the x-ray tubes was avoided. The source can be pulsed or operated in a highly stable continuous mode. The spatial and energetic characteristics of the hot electrons were determined via their bremsstrahlung by comparison with theoretical calculations. In this test x-ray source the average energy of the hot electron flux was in the range 10-40 keV, depending on the operating parameters: intermagnet distance (6-9 cm), power of the 2.45 GHz microwave (200-1000 W), chamber pressure (4 x 10(-5)-9 x 10(-5) Torr). The insertion of solid targets into the electron flux leads to various focal spot sizes, enabling application in radiology and mammography. The images of a mammographic phantom were obtained in a few seconds with good contrast (microcalcifications being successfully detected). PMID- 11277230 TI - Total variation norm for three-dimensional iterative reconstruction in limited view angle tomography. AB - An iterative Bayesian reconstruction algorithm for limited view angle tomography, or ectomography, based on the three-dimensional total variation (TV) norm has been developed. The TV norm has been described in the literature as a method for reducing noise in two-dimensional images while preserving edges, without introducing ringing or edge artefacts. It has also been proposed as a 2D regularization function in Bayesian reconstruction, implemented in an expectation maximization algorithm (TV-EM). The TV-EM was developed for 2D single photon emission computed tomography imaging, and the algorithm is capable of smoothing noise while maintaining edges without introducing artefacts. The TV norm was extended from 2D to 3D and incorporated into an ordered subsets expectation maximization algorithm for limited view angle geometry. The algorithm, called TV3D-EM, was evaluated using a modelled point spread function and digital phantoms. Reconstructed images were compared with those reconstructed with the 2D filtered backprojection algorithm currently used in ectomography. Results show a substantial reduction in artefacts related to the limited view angle geometry, and noise levels were also improved. Perhaps most important, depth resolution was improved by at least 45%. In conclusion, the proposed algorithm has been shown to improve the perceived image quality. PMID- 11277231 TI - A mathematical comparison of two models of the electrical properties of biological tissues. AB - The purpose of this paper is to compare two models of the electrical properties of tissue, which may be used to relate the effective conductivity to the volume fraction f of cells in the tissue. Both models assume that tissue comprises spherical cells, which behave electrically as dipoles. The first model, developed by Hanai, describes the tissue as a concentrated suspension of weakly conducting spheres in a conducting medium, with each sphere experiencing a uniform mean field. The second approach, developed by Chiew and Glandt, explicitly describes the effect of a random but statistically homogeneous cell structure on the average field and magnitude of the dipole interaction. The two analyses are identical to first order in f, but differ in the way in which the interactions between the dipoles are accounted for. The model developed by Chiew and Glandt appears to offer a more robust theoretical framework for describing the electrical properties of tissue. The comparison aims to contribute to an improved understanding of the relationship between the electrical properties and spatial structure of tissue. PMID- 11277232 TI - Determining changes in NIR absorption using a layered model of the human head. AB - A theoretical approach is presented to determine absorption changes in different compartments of a layered structure from distributions of times of flight of photons. In addition resulting changes in spatial profiles of time-integrated intensity and mean time of flight are calculated. The capability of a single distance, time-domain method to determine absorption changes with depth resolution is tested on a layered phantom. We apply this method to in vivo measurements on the human head (motor stimulation, Valsalva manoeuvre) and introduce a small-sized time-domain experimental set-up suitable for bedside monitoring. PMID- 11277233 TI - Polyhydramnios and arterio-arterial placental anastomoses may beneficially affect monochorionic twin pregnancies. AB - Our objective was to appraise whether an increased amniotic fluid pressure by polyhydramnios can beneficially affect monochorionic twins that are haemodynamically connected by arterio-venous plus arterio-arterial placental anastomoses. We assessed the effects of polyhydramnios in monochorionic twin placentas, combining (a) data from previous in vitro placental perfusion experiments in singleton term placentas under simulated normal and increased amniotic fluid pressures with (b) logical deduction from observations made in monochorionic twins. Our hypothesis is that in monochorionic placentas, an increased amniotic fluid pressure increases the placental microvascular resistance but not the resistance of placental chorionic plate arteries. Hence, an increased amniotic fluid pressure increases the microvascular resistance of the joint cotyledon, the arterio-venous resistance, but not the arterioarterial resistance. This proposed mechanism reduces arterio-venous but not oppositely directed arterio-arterial transfusion. Therefore, reversal of the normal direction of net foeto-foetal transfusion may develop, which will reduce the circulatory imbalance that evolved between the monochorionic foetal twins. In contrast, in monochorionic twins connected by unidirectional or bidirectional arterio-venous anastomoses reversal of the normal direction of net foeto-foetal transfusion will not occur. In conclusion, reversal of the normal direction of net foeto-foetal transfusion, induced by polyhydramnios, is protective against the onset and severity of twin-twin transfusion syndrome in monochorionic twins connected by arterio-venous plus arterio-arterial anastomoses, but not by unidirectional or bidirectional arterio-venous anastomoses. PMID- 11277234 TI - A scattering phase function for blood with physiological haematocrit. AB - Though the optics of red blood cells as well as whole blood has been studied extensively, an effective scattering phase function for whole blood is still needed. The interference of waves scattered by neighbouring cells cannot be neglected in highly concentrated suspensions such as whole blood. As a result, the phase function valid for single erythrocytes may fail to describe a single scattering process in whole blood with physiological haematocrit (Hct approximately 0.4). In this study we compared the results obtained in goniophotometric measurements of blood samples with the results of angle-resolved Monte Carlo simulations. The results show that a Henyey-Greenstein phase function with an anisotropy factor of 0.972 is an adequate approximation for the effective scattering phase function of whole blood with high haematocrit at a wavelength of 514 nm. PMID- 11277235 TI - EXAFS study on the local atomic structures around iron in glycosylated haemoglobin. AB - Samples with 5.1%, 9.8% and 15.3% HbA1c were extracted from normal subjects and patients with slight and serious diabetes respectively. Extended x-ray absorption fine structure spectra of Fe K absorption were collected at the EXAFS experimental station of the Beijing Synchrotron Radiation Facility. The step-by step fluorescent mode was employed with a count time of 10 s per point. Several independent scans were averaged to eliminate the statistical noise. Reference backscattering amplitudes and phaseshifts were calculated using the curve wave theory (FEFF code) of EXAFS. Apart from the nitrogen neighbours around the central iron atom, oxygen neighbours are also found. The Fe-N bond length increases by about 0.02 A for the sample with 15.3% HbA1c compared with the others, but the Fe-O bond length is almost unchanged. With increasing of HbA1c concentration, the content of Hb increases and the content of HbO2 decreases. This demonstrates that the glycosylation of haemoglobin will decrease its ability to carry oxygen. PMID- 11277236 TI - Monte Carlo dosimetry of the Buchler high dose rate 192Ir source. AB - In this study a complete set of dosimetric data is presented for the high dose rate (HDR) source from Amersham used in the Buchler remote afterloading HDR unit. These data have been calculated by means of the Monte Carlo simulation code GEANT taking into account the detailed geometry of the source. Absolute dose rate distributions in water were calculated around this source and are presented as conventional 2D Cartesian look-up tables. All dosimetric quantities recommended by the AAPM Task Group 43 report have been calculated. Quantities determined are: dose rate constant, radial dose function, anisotropy function, anisotropy factor and anisotropy constant. The dose rate distributions of the Buchler HDR source are compared with those of other HDR sources used in brachytherapy, showing that the differences are large in zones near the long source axis due to oblique filtration. These Monte Carlo simulated data in water can be used for clinical applications. PMID- 11277237 TI - Medical image registration. AB - Radiological images are increasingly being used in healthcare and medical research. There is, consequently, widespread interest in accurately relating information in the different images for diagnosis, treatment and basic science. This article reviews registration techniques used to solve this problem, and describes the wide variety of applications to which these techniques are applied. Applications of image registration include combining images of the same subject from different modalities, aligning temporal sequences of images to compensate for motion of the subject between scans, image guidance during interventions and aligning images from multiple subjects in cohort studies. Current registration algorithms can, in many cases, automatically register images that are related by a rigid body transformation (i.e. where tissue deformation can be ignored). There has also been substantial progress in non-rigid registration algorithms that can compensate for tissue deformation, or align images from different subjects. Nevertheless many registration problems remain unsolved, and this is likely to continue to be an active field of research in the future. PMID- 11277238 TI - Continuing the fight to retain control of the patient-physician relationship. PMID- 11277239 TI - Tibial osteotomy coincident with long stem total knee arthroplasty: a surgical technique. AB - Tibial deformity secondary to previous fracture or osteotomy requires corrective osteotomy in some patients undergoing total knee arthroplasty (TKA). This can be performed in either two stages or coincident with the arthroplasty. Literature on coincident tibial osteotomy and TKA has been published previously, but details of the complicated surgical technique are lacking for surgeons performing this procedure for the first time. This article details the preoperative planning involved and the intraoperative technique used in tibial osteotomy coincident with TKA. PMID- 11277240 TI - Effect of quadriceps contraction on tangential patellar radiography. AB - This study examined whether the addition of quadriceps contraction to standard Merchant views provides additional useful information in the evaluation of patients with extensor mechanism malalignment. Fifteen patients (23 knees) with anterior knee pain due to lateral patellar compression syndrome and 22 control patients (44 knees) underwent standard Merchant views with the quadriceps relaxed and with an isometric isotonic contraction. Congruence and lateral patellar angles were measured for all groups. Although the congruence angle differed significantly between the control and symptomatic groups with the quadriceps contracted (P< or = .001), this difference also was seen without quadriceps contraction. There was also no significant difference within each group on addition of quadriceps contraction. No significant difference existed between the two groups for lateral patellar angle with quadriceps contraction. The addition of a controlled isometric quadriceps contraction did not add to the diagnostic yield of the standard Merchant view in terms of a predictable change in measured radiographic parameters. PMID- 11277241 TI - Effect of a patella-stabilizing brace on lateral subluxation of the patella: assessment using kinematic MRI. AB - This study investigated the effect of a special patella brace on patients with lateral subluxation of the patella using kinematic magnetic resonance imaging (MRI). Fifteen patients were assessed with and without application of the brace (Shields Patella Stabilizing Brace, Hely & Weber, Santa Paula, Calif) using active-movement, against-resistance kinematic MRI of the patello-femoral joint. Kinematic MRI examinations were evaluated using previously described qualitative criteria to determine patello-femoral relationships. Eleven (73%) patellofemoral joints had improvement (55%) or correction (45%) of lateral subluxation of the patella. The brace failed to alter the position of the patella in four (27%) patients who had patella alta (two patients) or were extremely overweight (two patients). This study provided objective findings that application of the brace improved or corrected lateral subluxation of the patella in the majority of patients. This information has important implications for the conservative treatment of patients with this form of patellar malalignment. PMID- 11277242 TI - Tendon-to-bone healing of a semitendinosus tendon autograft used for ACL reconstruction in a sheep model. AB - Anterior cruciate ligament (ACL) reconstruction was performed in a single hind limb of 30 sheep using a doubled semitendinosus tendon graft. Three additional animals were used as controls. Histologic and biomechanical analysis was performed from 4-52 weeks postoperatively. Perpendicular collagen fibers were found connecting the tendon graft to the bone tunnels at 8 weeks. These fibers were seen circumferentially at 12 weeks. By 24 weeks, the bone tunnel was well defined, and no further changes were observed at 52 weeks. Tendon incorporation within the femoral and tibial tunnels was similar at each interval. Although the small sample size did not permit statistical testing, the reconstruction strength was similar up to 12 weeks (15%-19% of controls). This increased at 24 (28%) and 52 (40%) weeks. The stiffness primarily increased from 4-8 weeks (18%-39%) and 24 52 weeks (52%-82%). Up to 12 weeks, failures occurred by graft pull-out from the bone tunnel. All 24- and 52-week specimens ruptured through the intra-articular portion of the graft, further indicating sufficient graft incorporation within the bone tunnels. PMID- 11277243 TI - Early postoperative assessment technique: a time-to-remission technique after condylar replacement. AB - Inconsistencies in reports of clinical outcome occur on many levels. Using life table techniques for remission can increase reproducibility, facilitate comparisons with other studies, and answer patient questions about rehabilitation time. The complementary rate of Kaplan-Meier's cumulative survival rate was calculated for pain and knee mobility after unicondylar and bicondylar knee replacement in osteoarthritis and rheumatoid arthritis. Pain relief was quick, often within 2 months. Eventually, >50% had attained both pain alleviation and normal mobility. The probability of regaining at least 90 degrees flexion was greater than regaining 0 degrees extension. The probability of increasing the preoperative extension capacity was higher than that of increasing the flexion capacity. PMID- 11277244 TI - Simultaneous avulsion fracture of the tibial tubercle with avulsion of the patellar ligament. PMID- 11277245 TI - Rotational positioning of the femoral component in total knee arthroplasty. PMID- 11277246 TI - Orienting the femoral component at total knee arthroplasty. PMID- 11277248 TI - Flexion space balancing using a prosthesis with asymmetrical posterior femoral condyles without external rotation. PMID- 11277247 TI - Determining proper femoral component rotational alignment during total knee arthroplasty. PMID- 11277249 TI - Positioning the femoral component: the effect of proper ligament balance. PMID- 11277250 TI - Preoperative calculation of the femoral transepicondylar axis: a combined radiographic and mathematical method. PMID- 11277251 TI - Analytical method development for 18 pesticides in house dust and settled residues using SEC, SPE, TMS methylation, and GC-MS. AB - An analytical method is developed to analyze eighteen pesticides in carpet dust and also dust that has settled on surfaces in order to determine the potential exposure of children to pesticide residues. For nonacid pesticides, the extract after centrifugation and filtration is cleaned up using size-exclusion chromatography (SEC) and then analyzed by gas chromatography (GC) coupled with a mass spectrometer (MS). The best solvent for extraction is ethyl acetate cyclohexane (3:1). The recoveries of spiked nonacid pesticides from 2 g of dust are between 72% and 110% with a variation between 4.2% and 25.6%, and the detection limit is 10 to 50 ng/g dust, depending on the pesticide. For acid pesticides, the dust is extracted with a saturated Ca(OH)2 solution, centrifuged, cleaned up by polyvinylbenzene/polystyrene-type solid-phase extraction cartridges, and methylated with trimethylsilyldiazomethane (TMS). Acid pesticides on filter paper samples are extracted with acidified acetone (3 mM H3PO4) and methylated with TMS. Methylation with TMS is fast and easy to perform. Methyl esters of the pesticides are completely separated and detected at low levels by GC-MS in the selective ion monitoring mode. The average recoveries of pesticides from 2 g of dust are between 81% and 104%. The average recoveries of pesticides spiked on filter paper are between 88% and 113%. A capillary column with a stationary phase of trifluoropropylmethyl polysiloxane gives the best separation and sensitivity for most pesticides on the GC-MS. PMID- 11277252 TI - Estimation of molar heat capacities in solution from gas chromatographic data. AB - The temperature dependence of retention data (retention or capacity factors) is measured for 35 aliphatic ketones and aldehydes as model compounds on a dimethylpolysiloxane stationary phase. A novel model is derived to determine the heat of solution and the solution molar heat capacities from the fits of the log natural of the difference of the retention factor and the column temperature (T) versus 1/T and the temperature arrangement. The convex curvature present in the residual plots of a former defined equation of ours disappears when applying a newly defined model. A detailed statistical analysis clearly shows the superiority of the refined model to the earlier one in a broader temperature range. The validation of this model is made through a comparison of heat capacity values taken from literature determined by different methods. The molar heat capacity of a pure liquid oxo compound is similar to that of when the same compound is solvated in a stationary phase. PMID- 11277253 TI - Determination of carvedilol in human cardiac tissue by high-performance liquid chromatography. AB - A new high-performance liquid chromatographic method has been developed for the determination of the beta-receptor blocker carvedilol in human cardiac tissue. After homogenizing tissue samples in a microdismembrator, carvedilol and the internal standard naftopidil are extracted with acetone. The extract is evaporated to dryness and reconstituted in a potassium acetate buffer of pH 3.5. Samples are cleaned up with solid-phase extraction columns. Carvedilol and the internal standard show recoveries of 69.8 +/- 12.2% and 63.9 +/- 9.34%, respectively. The linearity range for carvedilol is 0.01-0.35 ng/mg (parts per billion) tissue (wet weight), and the limit of quantitation is 0.01 ng/mg. The percentage coefficient of variation of the intra-assay varies between 1.45 and 5.38% and the interassay between 4.25 and 6.96%. To use as an application of the assay, the cardiac carvedilol tissue level in a patient on oral carvedilol therapy for congestive heart failure is reported. PMID- 11277254 TI - My separation is initially fine using a methanol-water mobile phase at pH 7; but over a week or so, the performance rapidly decreases and is not usable. PMID- 11277255 TI - One of the capillary columns in my dual column assembly degraded faster than the other. PMID- 11277256 TI - High-performance liquid chromatographic method development and validation for the simultaneous quantitation of naproxen sodium and pseudoephedrine hydrochloride impurities. AB - A reversed-phase high-performance liquid chromatographic procedure for the simultaneous determination of impurities associated with pseudoephedrine hydrochloride (PSEH) and naproxen sodium (NapNa) is developed and validated. The method is developed using a Waters Spherisorb cyano column (5 microm, 250 x 4.6 mm). An isocratic elution in a water-acetonitrile-methanol-triethylamine mixture (850:75:75:5) is adjusted to a pH of 3.7 +/- 0.02 with formic acid as the mobile phase. The UV detection was set at 260 nm, and the wavelength was switched to 235 nm before the elution of the last component, 2-ethyl-6-methoxy-naphthalene (EMN). The method is shown to be linear at a concentration range of 0.24 to 1.92 microg/mL for benzaldehyde, benzoic acid, and 2-(methylamino)-propiophenone hydrochloride, which are known impurities of PSEH. The NapNa impurities, 2-(6' hydroxy-2'-naphthyl) propionic acid, 2-hydroxy-6-methoxy-naphthalene, 1-(6' methoxy-2'-naphthyl) ethanol, 2-acetyl-6-methoxy-naphthalene, and EMN are also demonstrated to be linear at a concentration range of 0.44 to 3.52 microg/mL. Under the chromatographic conditions of the method, all impurities are resolved from the active components. PMID- 11277257 TI - Oligoacetic acid characterization by isocratic and linear salt gradient anion exchange chromatography. AB - A homologous series of four oligomers of acetic acid (namely acetic acid, succinic acid, tricarballylic acid, and tetracarboxylic acid) is characterized using isocratic and linear salt gradient anion-exchange chromatography. The double logarithmic plot of the isocratic retention factors versus the salt concentration gives straight lines for all samples. These straight lines (with the exception of the line for the strongly retained succinate peak) have a common intersection point--something which is proved to be a direct consequence of the stoichiometric mass-action ion-exchange model. The characteristic charge and the equilibrium ion-exchange constant (and the corresponding Gibbs free energy of ion exchange, or deltaG(exchange)) are determined from the isocratic experiments. The characteristic charge agrees satisfactorily with the number of carboxylic acid groups in the samples, and the deltaG(exchange) value decreases linearly with the characteristic charge. Succinic acid always gives two chromatographic peaks despite the proven chemical purity of the sample. The characteristic charge that is calculated for both of the succinic acid peaks is approximately two. The deltaG(exchange) value calculated for the weakly retained succinic acid peak falls in the free energy versus characteristic charge straight line defined by the other homologues. The deltaG(exchange) value of the strongly retained peak is lower than that of the weakly retained peak by 1.85 kJ/mol. The two succinic acid peaks are explained in terms of an equilibrium between two conformers in solution -one binding the solution counterions tightly and the other loosely. An analysis of all samples under a linear salt gradient provides retention times that increase linearly with the number of functional groups. Using an appropriate model (along with the isocratically determined characteristic charge and ion exchange constant), we predict theoretically the linear gradient retention times, which agree reasonably well with the experimental ones. PMID- 11277258 TI - High-performance liquid chromatographic analysis for the characterization of triterpenoids from Ganoderma. AB - A high-performance liquid chromatographic (HPLC) analysis of triterpenoids from Ganoderma is developed and validated in an attempt to explore a way to differentiate a number of species of the genus Ganoderma. Results show that 64 samples examined in this study could be divided into 18 groups based on characteristics of the HPLC pattern of triterpenoids. This result also conforms with those of the morphological examination and the interfertility test by di monokaryotic mating. The HPLC analysis of triterpenoids further reveals that differentiation among samples from three different regions each of the two species G. lucidum and G. tsugae is workable. Even then, an incorrect designation is found for two of the groups of samples that were originally classified as G. resinaceum but showed different morphological characteristics and mating incompatibility. In conclusion, an HPLC analysis of triterpenoids is a simple and easy way to differentiate among different species of the genus Ganoderma. PMID- 11277259 TI - Long-term effect of incadronate disodium (YM-175) on fracture healing of femoral shaft in growing rats. AB - The aim of this study was to investigate the long-term effect of incadronate on fracture healing of the femoral shaft in rats. Female Sprague-Dawley 8-week-old rats were injected subcutaneously (sc) with either vehicle (V group) or two doses of incadronate (10 microg/kg and 100 microg/kg) three times a week for 2 weeks. Right femoral diaphysis was then fractured and fixed with intramedullary stainless wire. Just after fracture, incadronate treatment was stopped in pretreatment groups (P groups: P-10 and P-100) or continued in continuous treatment groups (C groups: C-10 and C-100). All rats were killed at 25 weeks or 49 weeks after surgery. Fractured femur was evaluated radiologically and mechanically and then stained in Villanueva bone stain and embedded in methyl methacrylate. Undecalcified cross-sections from the fracture area were evaluated microradiologically and histomorphometrically. Radiographic observation showed that the fracture line disappeared in all groups. Cross-sectional area in the C 100 group was the biggest among all groups and in the C-10 group was larger than that in the V group at 25 weeks. Histological and histomorphometric observations showed that the process of fracture healing was delayed under continuous treatment with incadronate as evidenced by the delay of both lamellar cortical shell formation and resolution of original cortex in C groups. Percent linear labeling perimeter, mineral apposition rate (MAR), and bone formation rate (BFR) in C groups significantly decreased compared with the other groups, indicating that the callus remodeling was suppressed under continuous treatment, especially with a high dose. Mechanical study showed that the stiffness and ultimate load of the fractured femur in the C 100 group were the highest among all groups at both 25 weeks and 49 weeks. In conclusion, this study showed that long-term continuous treatment with incadronate delayed the process of fracture healing of femur in rats, especially under high dose but it did not impair the recovery of mechanical integrity of the fracture. PMID- 11277260 TI - Can bisphosphonates be given to patients with fractures? PMID- 11277261 TI - Stimulation of protein kinase C activity in cells expressing human parathyroid hormone receptors by C- and N-terminally truncated fragments of parathyroid hormone 1-34. AB - The parathyroid hormone (PTH) fragment PTH(1-34) stimulates adenylyl cyclase, phospholipase C (PLC), and protein kinase C's (PKCs) in cells that express human, opossum, or rodent type 1 PTH/PTH-related protein (PTHrP) receptors (PTHR1s). Certain carboxyl (C)-terminally truncated fragments of PTH(1-34), such as human PTH(1-31) [hPTH-(1-31)NH2], stimulate adenylyl cyclase but not PKCs in rat osteoblasts or PLC and PKCs in mouse kidney cells. The hPTH(1-31)NH2 peptide does fully stimulate PLC in HKRK B7 porcine renal epithelial cells that express 950,000 transfected hPTHR1s per cell. Amino (N)-terminally truncated fragments, such as bovine PTH(3-34) [bPTH(3-34)], hPTH(3-34)NH2, and hPTH(13-34), stimulate PKCs in Chinese hamster ovary (CHO) cells expressing transfected rat receptors, opossum kidney cells, and rat osteoblasts, but an intact N terminus is needed to stimulate PLC via human PTHR1s in HKRK B7 cells. We now report that the N terminally truncated analogs bPTH(3-34)NH2 and hPTH(13-34)OH do activate PKC via human PTHR1s in HKRK B7 cells, although less effectively than hPTH(1-34)NH2 and hPTH(1-31)NH2. Moreover, in a homologous human cell system (normal foreskin fibroblasts), these N-terminally truncated fragments stimulate PKC activity as strongly as hPTH(1-34)NH2 and hPTH(1-31)NH2. Thus, it appears that unlike their opossum and rodent equivalents, hPTHR1s can stimulate both PLC and PKCs when activated by C-terminally truncated fragments of PTH(1-34). Furthermore, hPTHR1s, like the PTHR1s in rat osteoblasts, opossum kidney cells, and rat PTHR1 transfected CHO cells also can stimulate PKC activity by a mechanism that is independent of PLC. The efficiency with which the N-terminally truncated PTH peptides stimulate PKC activity depends on the cellular context in which the PTHR1s are expressed. PMID- 11277262 TI - Parathyroid hormone/parathyroid hormone-related peptide type 1 receptor in human bone. AB - The parathyroid hormone/parathyroid hormone-related peptide (PTH/PTHrP) receptor (denoted as PTH-1R) is a key signaling factor through which calcium-regulating hormones PTH and PTHrP exert their effects on bone. There are contradictory reports regarding the capability of osteoclasts to express PTH-1R. To address this issue in humans, bone biopsy specimen samples from 9 normal controls and 16 patients with moderate to severe secondary renal hyperparathyroid bone disease (2 degrees HPT) with elevated PTH levels were studied to determine whether osteoclasts in the bone microenvironment express PTH-1R messenger RNA (mRNA) and protein. We report that osteoclasts express the PTH-1R mRNA but the protein is detected only in patients with 2 degrees HPT. The PTH-1R mRNA and protein also were found in osteoblasts, osteocytes, and bone marrow cells. Receptor expression was higher in osteoclasts and osteoblasts of patients with 2 degrees HPT than normal controls (98.0 +/- 1.1% vs. 65.7 +/- 14.3% and 65.8 +/- 3.4% vs. 39.1 +/- 6.2%; p < 0.01, respectively). Approximately half of osteoclasts found in bone of patients with 2 degrees HPT have the PTH-1R protein. In patients with 2 degrees HPT, a positive relationship exists between erosion depth, a parameter of osteoclastic activity, and the percentage of osteoclasts with PTH-1R protein (r = 0.58; p < 0.05). In normal controls, an inverse relationship exists between the percentage of osteoblasts with receptor mRNA, mRNA signals/cell, and serum PTH levels (r = -0.82 and p < 0.05 and r = -0.78 and p < 0.01, respectively). The results provide the novel evidence of PTH-1R in human osteoclasts and suggest a functional role for the receptors in 2 degrees HPT. PMID- 11277263 TI - Mechanical consequences of bone loss in cancellous bone. AB - The skeleton is continuously being renewed in the bone remodeling process. This prevents accumulation of damage and adapts the architecture to external loads. A side effect is a gradual decrease of bone mass, strength, and stiffness with age. We investigated the effects of bone loss on the load distribution and mechanical properties of cancellous bone using three-dimensional (3D) computer models. Several bone loss scenarios were simulated. Bone matrix was removed at locations of high strain, of low strain, and random throughout the architecture. Furthermore, resorption cavities and thinning of trabeculae were simulated. Removal of 7% of the bone mass at highly strained locations had deleterious effects on the mechanical properties, while up to 50% of the bone volume could be removed at locations of low strain. Thus, if remodeling would be initiated only at highly strained locations, where repair is likely needed, cancellous bone would be continuously at risk of fracture. Thinning of trabeculae resulted in relatively small decreases in stiffness; the same bone loss caused by resorption cavities caused large decreases in stiffness and high strain peaks at the bottom of the cavities. This explains that a reduction in the number and size of resorption cavities in antiresorptive drug treatment can result in large reductions in fracture risk, with small increases in bone mass. Strains in trabeculae surrounding a cavity increased by up to 1,000 microstrains, which could lead to bone apposition. These results give insight in the mechanical effects of bone remodeling and resorption at trabecular level. PMID- 11277264 TI - Caspase-dependent cleavage of cadherins and catenins during osteoblast apoptosis. AB - As transmembrane, Ca2+-dependent cell-cell adhesion molecules, cadherins play a central role in tissue morphogenesis and homeostasis. Stable adhesion is dependent on interactions of the cytoplasmic domain of the cadherins with a group of intracellular proteins, the catenins. In the present study, we have detected the expression of alpha-, beta-, and gamma-catenins in human osteoblasts, which assemble with cadherins to form two distinct complexes containing cadherin and alpha-catenin, with either beta- or gamma-catenin. In osteoblasts undergoing apoptosis, proteolytic cleavage of N-cadherin and beta- and gamma- catenins but not alpha-catenin was associated with the activation of caspase-3 and prevented by the caspase inhibitor Z-VAD-fmk. The pattern of cadherin/catenin cleavage detected in apoptotic osteoblasts was reproduced in vitro by recombinant caspase 3. The presence of a 90-kDa extracellular domain fragment of N-cadherin in conditioned medium from apoptotic cells indicates that additional extracellular or membrane-associated proteases also are activated. Disruption of N-cadherin mediated cell-cell adhesion with function-blocking antibodies induced osteoblast apoptosis, activation of caspases, and cleavage of beta-catenin. These findings provide compelling evidence that N-cadherin-mediated cell-cell adhesion promotes osteoblast survival and suggest that the underlying mechanism may involve activation of beta-catenin signaling. PMID- 11277265 TI - Human osteoclast cathepsin K is processed intracellularly prior to attachment and bone resorption. AB - Cathepsin K is a member of the papain superfamily of cysteine proteases and has been proposed to play a pivotal role in osteoclast-mediated bone resorption. We have developed a sensitive cytochemical assay to localize and quantify osteoclast cathepsin K activity in sections of osteoclastoma and human bone. In tissue sections, osteoclasts that are distant from bone express high levels of cathepsin K messenger RNA (mRNA) and protein. However, the majority of the cathepsin K in these cells is in an inactive zymogen form, as assessed using both the cytochemical assay and specific immunostaining. In contrast, osteoclasts that are closer to bone contain high levels of immunoreactive mature cathepsin K that codistributes with enzyme activity in a polarized fashion toward the bone surface. Polarization of active enzyme was clearly evident in osteoclasts in the vicinity of bone. The osteoclasts apposed to the bone surface were almost exclusively expressing the mature form of cathepsin K. These cells showed intense enzyme activity, which was polarized at the ruffled border. These results suggest that the in vivo activation of cathepsin K occurs intracellularly, before secretion into the resorption lacunae and the onset of bone resorption. The processing of procathepsin K to mature cathepsin K occurs as the osteoclast approaches bone, suggesting that local factors may regulate this process. PMID- 11277266 TI - Epigenetic regulation of human bone morphogenetic protein 6 gene expression in prostate cancer. AB - Bone morphogenetic proteins (BMPs), belonging to the transforming growth factor beta (TGF-beta) superfamily, are multifunctional molecules that regulate bone induction and organ development. Among BMPs, BMP-6 has been shown to be overexpressed in prostate cancer and is speculated to be associated with bone forming skeletal metastasis. We investigated the regulatory mechanism of the BMP 6 gene expression in prostate cancer cell lines DU-145, LNCaP, PC-3, and PC-3M with regard to the methylation status of the CpG island in the 5' flanking region of the human BMP-6 gene. By sequence-specific analysis of methylated cytosines, we show here that the methylation status of the CpG loci around the Sp1 site of the BMP-6 promoter is related to its steady-state expression and an alternative splicing of messenger RNA (mRNA) in prostate cancer cell lines. Furthermore, a study of clinical cases of benign and malignant prostate lesion by in situ hybridization showed that BMP-6 expression was high at both primary and secondary sites in cases of advanced cancer with metastasis. Demethylation of the CpG loci around the Spl binding site was shown in cases with high BMP-6 expression by sequencing analysis of the methylated cytosine from paraffin-embedded materials. Our results suggested that during cancer progression, besides inactivation of tumor suppressor genes by hypermethylation, activation of certain genes like BMP 6 by selective demethylation was a common epigenetic event giving a variable character to the invading and metastasizing cancer cells. PMID- 11277267 TI - The bone morphogenetic proteins antagonist Noggin inhibits membranous ossification. AB - Bone morphogenetic proteins (BMPs) are expressed and secreted during fracture repair. Although they are likely to be required for this process, little is known about their physiological role in bone regeneration. Noggin is a protein that specifically binds and inactivates several BMPs. It plays fundamental roles during early embryonal development and limb morphogenesis by this BMP inactivating activity. This study shows that Noggin can modify bone formation in vivo in the adult animal and, thus, indirectly, that BMP signaling is indispensable in this process. A noggin mutein (hNgdeltaB2-Fc) engineered so as to display increased bioavailability was used. Bilateral titanium bone chambers were inserted in 70 rats, and side comparisons for bone formation in the chambers were done. The hNgdeltaB2-Fc had no effect on total amount of tissue formed in the chamber but decreased the amount of bone compared with both buffer controls and a control made up of an Fc-tagged IL-6Ralpha protein, which had no effects of its own. Also, wild-type noggin inhibited bone formation. Thus, endogenous BMP signaling is necessary for normal bone regeneration. PMID- 11277268 TI - Down-regulation of procollagen alpha1[I]] messenger RNA by titanium particles correlates with nuclear factor kappaB (NF-kappaB) activation and increased rel A and NF-kappaB1 binding to the collagen promoter. AB - Previously, we showed that exposure of human osteoblasts to titanium particles stimulates protein tyrosine phosphorylation (PTP), activates the transcription factor nuclear factor kappaB (NF-kappaB), and causes an approximately 50% decrease in the steady-state messenger RNA (mRNA) level of procollagen alpha1[I]. In this study, we identify three NF-kappaB binding sites within the human procollagen alpha1[I] gene promoter, show that titanium particles stimulate their binding of the NF-kappaB subunits Rel A (p65) and NF-kappaB1 (p50), and find NF kappaB activation correlates with collagen gene suppression by titanium particles in osteoblasts. Protein tyrosine kinase (PTK) inhibitors, which significantly reduce the suppressive effect of titanium particles on collagen gene expression, inhibited NF-kappaB binding activity showing that titanium particle stimulation of PTK signals in osteoblasts are critical for both NF-kappaB activation and collagen gene expression. The antioxidant pyrrolidine dithiocarbamate (PDTC), which also inhibits the titanium particle suppression of collagen, abrogated the titanium particle activation of NF-kappaB, suggesting the involvement of redox signals in NF-kappaB-mediated collagen gene expression. The RNA polymerase II inhibitor actinomycin D (Act D) decreased procollagen alpha1[I] mRNA expression and effectively blocked the titanium-induced suppressive effect, suggesting that titanium particles activate a cascade of signals in osteoblasts, which result in a suppression of procollagen alpha1[I] mRNA. Collectively, these results show that titanium particles can activate NF-kappaB signaling in osteoblasts and suggest that NF-kappaB binding to the collagen gene promoter has a functional role in the down-regulation of procollagen alpha1[I] gene transcription. PMID- 11277269 TI - Collagen metabolism is markedly altered in the hypertrophic cartilage of growth plates from rats with growth impairment secondary to chronic renal failure. AB - Skeletal growth depends on growth plate cartilage activity, in which matrix synthesis by chondrocytes is one of the major processes contributing to the final length of a bone. On this basis, the present work was undertaken to ascertain if growth impairment secondary to chronic renal insufficiency is associated with disturbances of the extracellular matrix (ECM) of the growth plate. By combining stereological and in situ hybridization techniques, we examined the expression patterns of types II and X collagens and collagenase-3 in tibial growth plates of rats made uremic by subtotal nephrectomy (NX) in comparison with those of sham operated rats fed ad libitum (SAL) and sham-operated rats pair-fed with NX (SPF). NX rats were severely uremic, as shown by markedly elevated serum concentrations of urea nitrogen, and growth retarded, as shown by significantly decreased longitudinal bone growth rates. NX rats showed disturbances in the normal pattern of chondrocyte differentiation and in the rates and degree of substitution of hypertrophic cartilage with bone, which resulted in accumulation of cartilage at the hypertrophic zone. These changes were associated with an overall decrease in the expression of types II and X collagens, which was especially marked in the abnormally extended zone of the hypertrophic cartilage. Unlike collagen, the expression of collagenase-3 was not disturbed severely. Electron microscopic analysis proved that changes in gene expression were coupled to alterations in the mineralization as well as in the collagen fibril architecture at the hypertrophic cartilage. Because the composition and structure of the ECM have a critical role in regulating the behavior of the growth plate chondrocytes, results obtained are consistent with the hypothesis that alteration of collagen metabolism in these cells could be a key process underlying growth retardation in uremia. PMID- 11277270 TI - Regulation of the 1b isoform of the plasma membrane calcium pump by 1,25 dihydroxyvitamin D3 in rat osteoblast-like cells. AB - The first isogene of the plasma membrane calcium pump (PMCA1) is expressed on the apical plasma membrane of osteoblasts, but its regulation by 1,25 dihydroxyvitamin D3 [1,25(OH)2D3] has not been studied in this cell type. We studied 1,25(OH)2D3 effects on PMCA1 function, protein, messenger RNA (mRNA), and isoform expression in osteoblasts. Of seven rat and human immortalized osteoblast like cell lines studied, PMCA1 mRNA expression was confirmed in all. Only ROS 17/2.8 cells expressed measurable PMCA1 protein by Western analysis. Immunocytochemistry indicated that PMCA1 was expressed predominantly on the plasma membrane of ROS 17/2.8 cells. The 1,25(OH)2D3 but not 24,25 dihydroxyvitamin D3 [24,25(OH)2D3] treatment of confluent ROS 17/2.8 cells resulted in an approximate 3- to 5-fold dose-dependent increase in PMCA1 expression of message and protein as assessed by Western and Northern analysis and vesicular 45Ca uptake of membrane vesicles. 1,25(OH)2D3 had no effect on PMCA1 posttranscriptional splicing. The 1b isoform of PMCA was expressed under all experimental conditions. 1,25(OH)2D3 favored increased expression of the 5.5 kilobases (kb) over the 7.5-kb PMCA1b transcript, with a 2-fold proportional increase in the smaller transcript relative to the larger transcript evident at the highest dose of 1,25(OH)2D3 studied. The resultant proportional increase in the smaller 5.5-kb transcript may increase mRNA stability and account for the increase in PMCA1b protein and function with 1,25(OH)2D3. These data provide evidence for the role of 1,25(OH)2D3 and PMCA1b in the regulation of calcium transport in bone cells. PMID- 11277271 TI - Nitric oxide synthase isoforms during fracture healing. AB - We have shown previously that nitric oxide (NO) has regulatory effects on fracture healing. Our aim here was to investigate the temporal expression patterns of the three NO synthase (NOS) isoforms that are responsible for the generation of NO by semiquantitative competitive polymerase chain reaction (PCR) and immunoblot analysis after femoral fractures in rats. We found that 4 days after fracture, there were increases in the levels of messenger RNA (mRNA) for all three NOS isoforms, with peaks for the inducible NOS (iNOS; 35-fold increase, p < 0.05) at day 4, the endothelial NOS (eNOS; 5-fold increase, p < 0.05) at day 7, and the neuronal NOS (bNOS; 16-fold increase, p < 0.05) at day 21. At a protein level, the time course expression of NOS isoforms was consistent with the results of those at the mRNA level. In addition, we have previously reported a 2.5-fold increase in NOS activity detected by [3H]arginine to [3H]citrulline conversion at day 15 compared with that at day 4 after fracture. The findings that the expression of NOS isoforms during fracture healing is type specific and time dependent are important and may have clinical applications in the regulation of bone repair by NOS inhibitors or stimulators at different stages after injury. PMID- 11277272 TI - Long-term effects of withdrawal of bisphosphonate incadronate disodium (YM175) on bone mineral density, mass, structure, and turnover in the lumbar vertebrae of ovariectomized rats. AB - This study was designed to evaluate the long-term effects of incadronate disodium (YM175) after its withdrawal on cancellous bone mass in ovariectomized (OVX) rats. Thirteen-week-old female SD rats were randomized into four groups: sham operated, OVX, low-YM, and high-YM (0.01 mg/kg or 0.1 mg/kg subcutaneously [sc], three times a week after OVX) groups. After 4 weeks of treatment with vehicle or YM175, rats from each group were killed at time points of 0 (baseline), 3, 6, 9, and 12 months after withdrawal of the agent. Bone mineral density (BMD) of the lumbar vertebrae was measured by dual-energy X-ray absorptiometry (DXA). Bone volume (BV/TV), trabecular number and trabecular separation (Tb.N and Tb.Sp), eroded surface (ES/BS), osteoclast number and osteoclast surface (N.Oc/BS and Oc.S/BS), osteoid surface (OS/BS), and bone formation rate (BFR/BS) were measured as histomorphometric parameters of the fifth lumbar vertebra. BMD, BV/TV, Tb.N, and Tb.Sp in YM175-treated groups were maintained at the same level as in the sham group until 12 months after withdrawal in the high-YM group and until 3 months after withdrawal in the low-YM group. YM175 decreased both bone formative and resorptive parameters in histomorphometry. Serum bone-specific alkaline phosphatase (ALP) and urinary deoxypyridinoline at both doses of YM175 also showed a suppressive effect of this agent on bone turnover. These results indicate that YM175, after withdrawal, still maintains bone volume dose dependently by depressing bone resorption and formation in OVX rats. Intermittent YM175 treatment with a long interval may be sufficient to maintain the bone volume and structure in OVX rats. PMID- 11277273 TI - The prospects of estimating trabecular bone tissue properties from the combination of ultrasound, dual-energy X-ray absorptiometry, microcomputed tomography, and microfinite element analysis. AB - Osteoporosis commonly is assessed by bone quantity, using bone mineral density (BMD) measurements from dual-energy X-ray absorptiometry (DXA). However, such a measure gives neither information about the integrity of the trabecular architecture nor about the mechanical properties of the constituting trabeculae. We investigated the feasibility of deriving the elastic modulus of the trabeculae (the tissue modulus) from computer simulation of mechanical testing by microfinite element analysis (muFEA) in combination with measurements of ultrasound speed of sound (SOS) and BMD measurements. This approach was tested on 15 postmortem bovine bone cubes. The apparent elastic modulus of the specimens was estimated from SOS measurements in combination with BMD. Then the trabecular morphology was reconstructed using microcomputed tomography (muCT). From the reconstruction a mesh for muFEA was derived, used to simulate mechanical testing. The tissue modulus was found by correlating the apparent moduli of the specimens as assessed by ultrasound with the ones as determined with muFEA. A mean tissue modulus of 4.5 GPa (SD, 0.69) was found. When adjusting the muFEA-determined elastic moduli of the entire specimens with their calculated tissue modulus, an overall correlation of R2 = 96% with ultrasound-predicted values was obtained. We conclude that the apparent elastic stiffness characteristics as determined from ultrasound correlate linearly with those from muFEA. From both methods in combination, the elastic stiffness of the mineralized tissue can be determined as an estimator for mechanical tissue quality. This method can already be used for biopsy specimens, and potentially could be applicable in vivo as well, when clinical CT or magnetic resonance imaging (MRI) tools with adequate resolution reach the market. In this way, mechanical bone quality could be estimated more accurately in clinical practice. PMID- 11277274 TI - Effect of pamidronate in preventing local bone loss after total hip arthroplasty: a randomized, double-blind, controlled trial. AB - Acute periprosthetic bone loss occurs after total hip arthroplasty. Bone loss undermines the support of the implant and may contribute to prosthetic failure. At present, there is no established prophylaxis for this process. We studied the effect of a single-dose infusion of 90 mg of pamidronate on early periprosthetic bone mineral density (BMD), biochemical markers of bone turnover, radiological, and clinical outcome in a 26-week, prospective, randomized, double-blinded study of 47 men and women undergoing total hip arthroplasty. Pamidronate therapy led to a significant reduction in bone loss compared with placebo for both the proximal femur and the pelvis (repeated measures analysis of variance [ANOVA]); p = 0.001 and p = 0.01, respectively). Pamidronate therapy was associated with suppression of all biochemical markers of bone turnover compared with placebo (repeated measures ANOVA; p < 0.05 for all comparisons), with the exception of urinary free deoxypyridinoline. Pamidronate did not interfere with the clinical improvement in symptoms after total hip arthroplasty, or radiological outcome, and was not associated with an increase in adverse events. This study provides clinical data on the efficacy and safety of bisphosphonates for the prevention of bone loss after total hip arthroplasty and supports the establishment of larger-scale clinical trials to determine the long-term clinical efficacy of this intervention using implant failure as the primary endpoint. PMID- 11277275 TI - Protective effect of short-tem calcitriol or cyclical etidronate on bone loss after cardiac or lung transplantation. AB - Bone loss is most rapid in the immediate period after cardiac or lung transplantation. This randomized study compared the efficacy of 6 months of treatment with either calcitriol (1,25-dihydroxyvitamin D3; 0.5 microg/day) or two cycles of etidronate plus calcium in preventing bone loss in 41 patients undergoing cardiac or lung transplantation. Patients were followed for 18 months after cessation of treatment. Bone mineral density (BMD) was measured by dual energy X-ray absorptiometry (DXA). There were no significant differences between groups with respect to age or cumulative dose of prednis(ol)one or cyclosporin over the 2 years. Bone loss did not differ between groups after 6 months and, despite 6 months prophylaxis with either agent, bone loss was significant in both groups at 6 months and 12 months. However, compared with an untreated reference group, both therapies offered significant protection at 6 months and etidronate provided significant protective carryover after therapy had been discontinued. These data suggest short-term prophylaxis with calcitriol or cyclical etidronate is partially effective in reducing bone loss after cardiac or lung transplantation but treatment needs to be continued for a longer term. PMID- 11277276 TI - Risk factors for hip fracture in Asian men and women: the Asian osteoporosis study. AB - The objectives of the Asian Osteoporosis Study (AOS) were to determine risk factors for hip fracture in men and women in four Asian countries, that is, Singapore, Malaysia, Thailand, and the Philippines. A total of 451 men and 725 women (aged 50 years and over) with hip fractures were compared with an equal number of community controls. A standardized questionnaire was administered by interview. The following relative risks (RRs) were found in women and men by multiple logistic regression: dietary calcium intake < 498 mg/day, 2.0 for women (95% CI, 1.5-2.8) and 1.5 for men (95% CI, 1.0-2.2); no load bearing activity in the immediate past, 2.0 for women (95% CI, 1.4-2.7) and 3.4 for men (95% CI, 2.3 5.1); no vigorous sport activities in young adulthood, 7.2 for women (95% CI, 4.0 13.0) and 2.4 for men (95% CI, 1.6-3.6); cigarette smoking, 1.5 for men (95% CI, 1.0-2.1); alcohol consumption 7 days a week, 2.9 for women (95% CI, 1.0-8.6) and 1.9 for men (95% CI, 1.1-3.2); fell twice or more in the last 12 months, 3.0 for women (95% CI, 1.8-4.8) and 3.4 for men (95% CI, 1.8-6.6); a history of fractures after 50 years of age, 1.8 for women (95% CI, 1.1-2.9) and 3.0 for men (95% CI, 1.6-5.6); a history of stroke, 3.8 for women (95% CI, 2.0-7.1) and 3.6 for men (95% CI, 1.8-7.1); use of sedatives, 2.5 for women (95% CI, 1.0-6.3) and 3.0 for men (95% CI, 1.0-9.7); and use of thyroid drugs, 7.1 for women (95% CI, 2.0-25.9) and 11.8 for men (95% CI, 1.3-106.0). Women who were 1.56 m or taller had an RR of 2.0 (95% CI, 1.3-3.0) for hip fracture and men who were 1.69 m or taller had an RR of 1.9 (95% CI, 1.2-3.1) for hip fracture. Based on these findings, primary preventive programs for hip fracture could be planned in Asia. PMID- 11277277 TI - Use of inhaled corticosteroids and risk of fractures. AB - Treatment with systemic corticosteroids is known to increase the risk of fractures but little is known of the fracture risks associated with inhaled corticosteroids. A retrospective cohort study was conducted using a large UK primary care database (the General Practice Research Database [GPRD]). Inhaled corticosteroid users aged 18 years or older were compared with matched control patients and to a group of noncorticosteroid bronchodilator users. Patients with concomitant use of systemic corticosteroids were excluded. The study comprised 170,818 inhaled corticosteroid users, 108,786 bronchodilator users, and 170,818 control patients. The average age was 45.1 years in the inhaled corticosteroid, 49.3 years in the bronchodilator, and 45.2 years in the control groups. In the inhaled corticosteroid cohort, 54.5% were female. The relative rates (RRs) of nonvertebral, hip, and vertebral fractures during inhaled corticosteroid treatment compared with control were 1.15 (95% CI, 1.10-1.20), 1.22 (95% CI, 1.04 1.43), and 1.51 (95% CI, 1.22-1.85), respectively. No differences were found between the inhaled corticosteroid and bronchodilator groups (nonvertebral fracture RR = 1.00; 95% CI, 0.94-1.06). The rates of nonvertebral fractures among users of budesonide (RR = 0.95; 95% CI, 0.85-1.07) and fluticasone propionate (RR = 1.03; 95% CI, 0.71-1.49) were similar to the rate determined for users of beclomethasone dipropionate. We conclude that users of inhaled corticosteroids have an increased risk of fracture, particularly at the hip and spine. However, this excess risk may be related more to the underlying respiratory disease than to inhaled corticosteroid. PMID- 11277279 TI - Low back pain in Norwegian helicopter aircrew. AB - The size and consequences of low back pain (LBP) in Norwegian helicopter aircrew has been investigated in a retrospective and prospective survey. With 50.5% reporting such pain in a 2-yr period, and Sea King aircrew reporting LBP on on almost half (49.3%) of the missions flown, the magnitude of the problem equals that reported from other air forces. Pilots reported LBP six times more often than other crewmembers and almost half (48.6%) felt the pain influenced the quality of work. This could have flight safety implications. Crewmembers with total flying time over 2,000 h have a significantly higher incidence of sick leave than those with less than 2,000 h. Only 1 pilot out of 10 with total flying time under 500 h had had flight-related LBP. PMID- 11277278 TI - Moderate alcohol consumption suppresses bone turnover in adult female rats. AB - Chronic alcohol abuse is a major risk factor for osteoporosis but the effects of moderate drinking on bone metabolism are largely uninvestigated. Here, we studied the long-term dose-response (0, 3, 6, 13, and 35% caloric intake) effects of alcohol on cancellous bone in the proximal tibia of 8-month-old female rats. After 4 months of treatment, all alcohol-consuming groups of rats had decreased bone turnover. The inhibitory effects of alcohol on bone formation were dose dependent. A reduction in osteoclast number occurred at the lowest level of consumption but there were no further reductions with higher levels of consumption. An imbalance between bone formation and bone resorption at higher levels of consumption of alcohol resulted in trabecular thinning. Our observations in rats raise the concern that moderate consumption of alcoholic beverages in humans may reduce bone turnover and potentially have detrimental effects on the skeleton. PMID- 11277280 TI - Aircraft control forces and EMG activity in a C-130 Hercules during strength critical maneuvers. AB - BACKGROUND: The force levels required to operate aircraft controls should be readily generated by pilots, without undue fatigue or exertion. However, maximum pilot applied forces, as specified in aircraft design standards, were empirically derived from the subjective comments of test pilots, and may not be applicable for the majority of pilots. Further, experienced RNZAF Hercules flying instructors have indicated that endurance and fatigue are problems for Hercules pilots. The aim of this study was to quantify aircraft control forces during emergency maneuvers in a Hercules aircraft and compare these forces with design standards. In addition, EMG data were recorded as an indicator of muscle fatigue during flight. METHODS: Six subjects were tested in a C-130 Hercules aircraft. The maneuvers performed were low-level dynamic flight, one engine-off straight and-level flight, and a two-engines-off simulated approach. The variables recorded were pilot-applied forces and EMG activity. RESULTS: Left rudder pedal force and vastus lateralis activity were both significantly greater during engine off maneuvers than during low-level dynamic flight (p < 0.05). Maximum aircraft control forces for all controls were within 10% of the design standards. The mean EMG activity across all muscles and maneuvers was 26% MVC, with a peak of 61% MVC in vastus lateralis during the two-engine-off approach. The median frequency of the vastus lateralis EMG signal decreased 13.0% and 16.0% for the one engine-off and two-engine-off maneuvers, respectively. CONCLUSION: The forces required to fly a Hercules aircraft during emergency maneuvers are similar to the aircraft design standards. However, the levels of vastus lateralis muscle activation observed during the engine-off maneuvers can be sustained for approximately 1 min only. Thus, if two engines fail more than 1 min before landing, pilots may have to alternate control of the aircraft to share the workload and enable the aircraft to land safely. PMID- 11277281 TI - Effect of "inside-out" and "outside-in" attitude displays on off-axis tracking in pilots and nonpilots. AB - BACKGROUND: Pilots employing helmet-mounted displays spend sustained periods of time looking off-axis, necessitating the inclusion of attitude symbology on the helmet to maintain spatial awareness. We examined how fundamentally different attitude references, a moving-horizon ("inside-out") or a moving-aircraft ("outside-in"), affected pilot and nonpilot attitude control when looking on- or off-axis. Both a rear-view and a side-view outside-in perspective were depicted to investigate the effect of control-display compatibility. METHODS: Subjects performed a compensatory pitch-roll tracking task either looking on-axis or 90 degrees off-axis using three symbologies: 1) a compressed pitch ladder with horizon line; 2) a 3-D aircraft representation viewed from the rear; and 3) a 3-D aircraft representation viewed from the side. Tracking error in roll and pitch, control bias, and subjective ratings were collected and analyzed. RESULTS: There was no significant difference in the tracking performance of U.S. Air Force pilots in pitch and roll using the inside-out or outside-in rear-view formats on- and off-axis, although they preferred the inside-out format. Nonpilots tracked significantly better using the outside-in rear-view format, which they also preferred. Both groups tracked poorly using the outside-in side-view format and control-display compatibility had no important effect. CONCLUSIONS: Pilots are equally adept using outside-in and inside-out displays. Given that an outside-in display may better reflect a person's inherent frame of reference for orientation (as evidenced by the nonpilots' superior performance with it), the results seem to indicate that pilots, through experience, have adapted to an inside-out frame of reference. PMID- 11277282 TI - The effects of +Gz force on the bone mineral density of fighter pilots. AB - HYPOTHESIS: Bone is a metabolically active tissue which responds to high strain loading. The purpose of this study was to examine the bone response to high +Gz force loading generated during high performance flying. METHODS: The bone response to +Gz force loading was monitored in 10 high performance RAAF pilots and 10 gender-, age-, height-, weight-matched control subjects. The pilots were stationed at the RAAF base at Pearce, Western Australia, all completing the 1-yr flight training course. The pilots flew the Pilatus PC-9 aircraft, routinely sustaining between 2.0 and 6.0 +Gz. Bone mineral density (BMD) and bone mineral content (BMC) were measured at baseline and 12 mo, using the Hologic QDR 2000+ bone densitometer. RESULTS: After controlling for change in total body weight and fat mass, the pilots experienced a significant increase in BMD and BMC for thoracic spine, pelvis, and total body, in the magnitude of 11.0%, 4.9%, and 3.7%, respectively. However, no significant changes in bone mineral were observed in the pilots lumbar spine, arms or legs. The control group experienced a significant decrease in pelvic BMC, with no other bone mineral changes observed at any site. CONCLUSIONS: These findings suggest that site specific BMD is increased in response to high +Gz forces generated during high performance flying in a PC-9. PMID- 11277283 TI - Safety of long distance aeromedical transport of the cardiac patient: a retrospective study. AB - BACKGROUND: The purpose of this study is to provide data regarding the safety of long distance air transport of cardiac patients, establish a time frame for safe transport, and assess current guidelines for postmyocardial infarct (post-MI) transport. METHODS: Retrospective analysis of all long distance aeromedical transports performed by Montreal-based Skyservice Lifeguard from January 1 to October 1, 1998. RESULTS: 109 cardiac patients were transported; 83 by air ambulance (AA), and 26 commercially (C). Diagnoses included MI (63%), unstable angina (31%), congestive heart failure (21%), and arrhythmia (17%). Patients were transported a mean of 7 d (AA) vs. 13.7 d (C) after presentation. Inflight complications, occurring in 10% of AA and 4% of C flights, were minor (chest pain, desaturation, and hypotension), and resolved quickly. In 51 post-MI AA patients, complication rate for transport > 7 d after admission was 0% (vs. 14% <7 d), and > 72 h after last chest pain was 6% (vs. 18% <72 h). Comparing uncomplicated (n = 25) vs. complicated (n = 26) MI reveals fewer complications for transport 0-3 d (13% vs. 50%) and 4-7 d (9% vs. 14%) after admission, and 48 72 h after last chest pain (0% vs. 100%). CONCLUSIONS: AA transport of cardiac patients can safely be performed earlier than guidelines for C flights. AA transport appears safe after complicated MI by day 7 or > 72 h chest pain free, and after uncomplicated MI by day 3 or > 48 h chest pain free. Future guidelines for aeromedical transport post-MI should distinguish between C and AA. PMID- 11277284 TI - A motion sickness maximum around the 0.2 Hz frequency range of horizontal translational oscillation. AB - BACKGROUND: Low frequency translational oscillation can provoke motion sickness in land vehicles, ships and aircraft. Although controlled motion experiments indicate a progressive increase in nauseogenicity as frequency decreases toward 0.2 Hz, few data are available on the existence of a definite maximum nauseogenic potential of motion around 0.2 Hz, or decreased nauseogenicity below this frequency. HYPOTHESIS: Nauseogenicity should be maximal around 0.2 Hz. METHODS: We selected 12 subjects for high motion sickness susceptibility, and they were exposed to horizontal sinusoidal motion (1.0 m.s(-2) peak acceleration) at 3 different frequencies (0.1, 0.2 and 0.4 Hz), at 1-wk intervals at the same time of day, according to a factorial design. Subjects were seated comfortably in the upright position with head erect. Fore-aft motion was through the body and head X axis. Motion was stopped (motion endpoint) at moderate nausea or after 30 min. RESULTS: The proportion of subjects experiencing moderate nausea was maximal at the intermediate frequency: 8/12 at 0.1 Hz, 12/12 at 0.2 Hz, 7/12 at 0.4 Hz. The mean time to motion endpoint was significantly (p < 0.01) shorter at the intermediate frequency: 18.0 min at 0.1 Hz; 11.2 min at 0.2 Hz; 20.2 min at 0.4 Hz. Similar frequency patterns emerged for other sickness ratings. The equivalent times to achieve moderate nausea using estimated values to correct for subjects who reached the 30 min time cut-off were: 22.7 min at 0.1 Hz; 11.2 min at 0.2 Hz; 28.1 min at 0.4 Hz. CONCLUSIONS: A maximum nauseogenic potential around 0.2 Hz was substantiated. PMID- 11277285 TI - Cardiovascular and humoral readjustment after different levels of head-up tilt in humans. AB - PURPOSE: To get a more complete picture of cardiovascular regulation after postural changes, this investigation directly monitored volume-related, hemodynamic, and endocrine variables during and after 30 min of passive head-up tilt (HUT) of various degrees. It was hypothesized that the return of variables to pre-tilt control level is of system-specific duration and different from what is found after lower body negative pressure (LBNP). DESIGN: We tested 7 persons on 5 different days using, in random order, no (HUT0) or different intensity (12 degrees , 30 degrees , 53 degrees , and 70 degrees ) of passive orthostasis (HUT12, HUT30, HUT53, HUT70). Data were collected before (supine), during, and after (supine) HUT and compared with synchronous data from HUT0. RESULTS: There was graded alteration with the sine of tilt angle for all hormones and directly volume-related variables. The effects of HUT70 were of the same magnitude as previously documented by others. After HUT, hemodynamic variables and catecholamines returned to control levels most rapidly. Heart rate depression, as observed in a companion LBNP study in the same subjects, did not occur. Vasopressin, PRA, plasma volume and Z0 returned to nominal values more slowly. Plasma aldosterone was still elevated 50 min after reassuming supine posture. CONCLUSION: Besides specific dose-responses within hemodynamic, volume-dependent, and hormonal variables after orthostatic loading of different degree, the return to control levels after HUT occurs with distinctly different time-courses, which are not identical with those seen after LBNP-simulated orthostasis. PMID- 11277286 TI - Lower body adynamia as a factor to reduce the risk of hypobaric decompression sickness. AB - BACKGROUND: We define lower body adynamia (LBA) as restricted lower body movement, particularly walking, during both the denitrogenation phase at site pressure and during the exercise phase while at altitude. HYPOTHESIS: Our null hypothesis is that subjects who are adynamic in the lower body but do upper body exercise will be at similar risk of decompression sickness (DCS) and venous gas emboli (VGE) as subjects who randomly walk but do no planned exercise while at altitude. METHODS: We selected a data set that contained 1401 altitude exposures with the following conditions: a) walking was part of the exercise at altitude; or b) there was no planned exercise done at altitude but walking was not restricted; or c) LBA was inforced, but upper body exercise was done at altitude. We used logistic regression (LR) on all 1401 exposures, a log logistic survival analysis (SA) on a subset of data from "a" and "c" (n = 234), and estimated a model for how the incidence of VGE changes through time. RESULTS: The estimated probabilities of DCS and VGE with 95% confidence intervals (Cls) from the LR with a simulation of a 3-h oxygen prebreathe, a 4-h exposure to 4.3 psia in a male, and exercise and LBA conditions as described above are: (see text). CONCLUSION: LBA that includes upper body exercise appears to be as protective against DCS and VGE as random walking by subjects who did no prescribed exercise while at altitude, and is more protective than exercise that included walking. Our conclusions are based on an assumption that we have adequately controlled, through our data selection process and the use of multivariable models, important variables in tests that were not done at the Johnson Space Center. PMID- 11277287 TI - Pulmonary function in men after repeated sessions of oxygen breathing at 0.25 MPa for 90 min. AB - HYPOTHESIS: We wanted to evaluate the pulmonary effects of discontinuous oxygen breathing (15 min O2, 2 min air breaks, 15:2), at 0.25 MPa once a day for 90 min O2 (6 sequences) over 10 d. This sequence, which has never been evaluated, is currently used in our hyperbaric therapy center. METHODS: Clinical and functional pulmonary status (questionnaire, spirometry, flow/volume loop, pulmonary diffusing capacity for carbon monoxide) was assessed in 10 non-smoking healthy volunteers after one exposure at 0.25 MPa consisting of 90 min of discontinuous oxygen breathing (15:2) and in 10 non-smoking patients who received a hyperbaric treatment consisting of 90 min of the same discontinuous O2 breathing (15:2) once a day over 10 d. The patients received daily intravenous methylprednisolone (1 mg x kg(-1)) and nicergoline (60 mg). RESULTS: There were no respiratory symptoms in either group. As expected, for a single exposure of that duration, lung function did not change in volunteers; however, a significant decrease in maximal expiratory flows (MEF) at 50 (-15%) and 25% (-33%) of forced vital capacity (p < 0.05) without change in forced vital capacity (FVC) appeared in patients treated over 10 d. CONCLUSION: Repetition of the 15:2 oxygen breathing sequence for 90 min once a day over 10 d led to greater flow limitation in peripheral airways than reported after continuous oxygen breathing of 210 min at 0.3 MPa which showed a 7% decrement in MEF50 and a 12% decrement in MEF25. No studies reporting these indexes were found in the 0.2-0.25 MPa range. Similar decrements in MEF50 and MEF25 with steady FVC have been reported after 14 d of daily hyperbaric therapy (0.24 MPa) with 30:5 sequence (-9% and -13%, respectively), 80% of the patients were symptom free. Similarily, our patients were all symptom free and remained so 1 yr after the study, hence, this toxicity is of weak clinical significance in subjects free of inflammatory lung diseases. HBO therapy, though safe, is not totally without effect on the lung. PMID- 11277288 TI - Type 2 diabetes in an aviator, protein diet vs. traditional diet: case report. AB - An experienced helicopter pilot with hypertension, hyper-triglyceridemia, elevated cholesterol, obesity, and diabetes is treated with a high-protein, low carbohydrate diet. In 3 months, he loses 35 lbs, is normotensive without medication, cholesterol and triglycerides show significant reduction, fasting blood glucose and 2-h post glucose load are normal. At follow-up 1 yr later he has maintained hemoglobin A1C in the low 5 range. The protein diet is discussed and compared with the traditional dietary approach for type 2 diabetes. PMID- 11277289 TI - Resynchronization of blood pressure circadian rhythm after westward trans-7 meridian flight with and without melatonin treatment. AB - Blood pressure (BP) of a healthy 37-yr-old male traveling from Milan to Houston was monitored for 36 h before the flight and continued for 5 d after the arrival. The rhythmometric analysis of BP data was made to investigate the rate of adaptation to a rapid rest-activity cycle shift. Since two trips were evaluated, during the second one the subject took melatonin (3 mg) before the nocturnal rest. In the first trip the BP circadian rhythm synchronization occurred on the 5th day. In the second trip melatonin promoted an immediate but unstable adaptation to the new rest-activity cycle. PMID- 11277290 TI - USAF pilot selection and training. PMID- 11277291 TI - Where are we going? PMID- 11277292 TI - The SD/LOC syndrome: spatial disorientation and loss of consciousness. PMID- 11277293 TI - Nonlinear behavior of trabecular bone at small strains. AB - Study of the behavior of trabecular bone at strains below 0.40 percent is of clinical and biomechanical importance. The goal of this work was to characterize, with respect to anatomic site, loading mode, and apparent density, the subtle concave downward stress-strain nonlinearity, that has been observed recently for trabecular bone at these strains. Using protocols designed to minimize end artifacts, 155 cylindrical cores from human vertebrae, proximal tibiae, proximal femora, and bovine proximal tibiae were mechanically tested to yield at 0.50 percent strain per second in tension or compression. The nonlinearity was quantified by the reduction in tangent modulus at 0.20 percent and 0.40 percent strain as compared to the initial modulus. For the pooled data, the mean +/- SD percentage reduction in tangent modulus at 0.20 percent strain was 9.07+/- 3.24 percent in compression and 13.8 +/- 4.79 percent in tension. At 0.40 percent strain, these values were 23.5 +/- 5.71 and 35.7+/- 7.10 percent, respectively. The magnitude of the nonlineari't depended on both anatomic site (p < 0.001) and loading mode (p < 0.001), and in tension was positively correlated with density. Calculated values of elastic modulus and yield properties depended on the strain range chosen to define modulus via a linear curve fit (p < 0.005). Mean percent differences in 0.20 percent offset yield strains were as large as 10.65 percent for some human sites. These results establish that trabecular bone exhibits nonlinearity at low strains, and that this behavior can confound intersite comparisons of mechanical properties. A nonlinear characterization of the small strain behavior of trabecular bone was introduced to characterize the initial stress-strain behavior more thoroughly. PMID- 11277294 TI - Estimation of bone matrix apparent stiffness variation caused by osteocyte lacunar size and density. AB - The role of osteocyte lacunar size and density on the apparent stiffness of bone matrix was predicted using a mechanical model from the literature. Lacunar size and lacunar density for different bones from different gender and age groups were used to predict the range of matrix apparent stiffness values for human cortical and cancellous tissue. The results suggest that bone matrix apparent stiffness depends on tissue type (cortical versus cancellous), age, and gender, the magnitudes of the effects being significant but small in all cases. Males had a higher predicted matrix apparent stiffness than females for vertebral cancellous bone (p< I0(-7)) and the difference increased with age (p =0.0007). In contrast, matrix apparent stiffness was not different between males and females forfemoral cortical bone and increased with age in both males (p < 0.0001) and females (p < 0.0364). Osteocyte lacunar density and size may cause significant gender and age related variations in bone matrix apparent stiffness. The magnitude of variations in matrix apparent stiffness was small within the physiological range of lacunar size and density for healthy bone, whereas the variations can be profound in certain pathological cases. It was proposed that the mechanical effects of osteocyte density be uncoupled from their biological effects by controlling lacunar size in normal bone. PMID- 11277295 TI - Dynamic behavior of lung surfactant. AB - We have previously developed an adsorption-limited model to describe the exchange of lung surfactant and its fractions to and from an air-liquid interface in oscillatory surfactometers. Here we extend this model to allow for diffusion in the liquid phase. Use of the model in conjunction with experimental data in the literature shows that diffusion-limited transport i.s important for characterizing the transient period from the start of oscillations to the achievement of steady-state conditions. Matching previous data shows that upon high levels of film compression, large changes occur in adsorption rate, desorption rate, and diffusion constant, consistent with what one might expect if the subsurface region was greatly enriched in DPPC. Collapse of the surfactant film that occurs during compression leads to a .significant elevation of surfactant concentration immediately heneath the interface, consistent with the subsurface depot of surfactant that has heen postulated by other investigators. Modeling studies also uncovered a phenomenon of surfactant behavior in which the interfacial tension remains constant at its minimum equilibrium value while the film is compressed, hut without collapse of the film. The phenomenon was due to desorption of surfactant from the interface and termed "pseudo-film collapse.'' The new model also gave improved agreement with steady-state oscillatory cycling in a pulsating bubble surfactometer. PMID- 11277296 TI - Model of human/liquid cooling garment interaction for space suit automatic thermal control. AB - The Wissler human thermoregulation model was augmented to incorporate simulation of a space suit thermal control system that includes interaction with a liquid cooled garment (LCG) and ventilation gas flow through the suit. The model was utilized in the design process of an automatic controller intended to maintain thermal neutrality of an exercising subject wearing a liquid cooling garment. An experimental apparatus was designed and built to test the efficacy of specific physiological state measurements to provide feedback data for input to the automatic control algorithm. Control of the coolant inlet temperature to the LCG was based on evaluation of transient physiological parameters that describe the thermal state of the subject, including metabolic rate, skin temperatures, and core temperature. Experimental evaluation of the control algorithm function was accomplished in an environmental chamber under conditions that simulated the thermal environment of a space suit and transient metabolic work loads typical of astronaut extravehicular activity (EVA). The model was also applied to analyze experiments to evaluate performance of the automatic control system in maintaining thermal comfort during extensive transient metabolic profiles for a range of environmental temperatures. Finally, the model was used to predict the efficacy of the LCG thermal controller for providing thermal comfort for a variety of regiments that may be encountered in future space missions. Simulations with the Wissler model accurately predicted the thermal interaction between the subject and LCG for a wide range of metabolic profiles and environmental conditions and matched the function of the automatic temperature controller for inlet cooling water to the LCG. PMID- 11277297 TI - Sagittal profile of the femoral condyles and its application to femorotibial contact analysis. AB - Measurements of the sagittal profiles of the articular surfaces of 24 femoral condyles were performed using a laser range finder. An algebraic algorithm was developed to reconstruct the measured sagittal profiles with simple geometry. In particular, it has been shown that a two-circular-arc model provides a very accurate reconstruction of the actual profiles in the femorotibial contact region. The average sagittal profile was used for a femorotibial contact analysis of TKA implants. The contact analysis was performed by using a rigid-body-spring model extended to the case of nonlinear force-deformation behavior of the tibial polyethylene component. PMID- 11277298 TI - Local dynamic stability versus kinematic variability of continuous overground and treadmill walking. AB - This study quantified the relationships between local dynamic stabiliht and variabilitr during continuous overground and treadmill walking. Stride-to-stride standard deviations were computed from temporal and kinematic data. Marimum finite-time Lyapunov exponents were estimated to quantify local dynamic stability. Local stability of gait kinematics was shown to be achieved over multiple consecutive strides. Traditional measures of variability poorly predicted local stability. Treadmill walking was associated with significant changes in both variability and local stability. Thus, motorized treadmills may produce misleading or erroneous results in situations where changes in neuromuscular control are likely to affect the variability and/or stability of locomotion. PMID- 11277299 TI - The generalized torque approach for analyzing the results of pedaling tests. AB - This paper deals with the analysis of experimental data obtained using an ergometer apparatus. A straightforward analysis method based on the power equation and the concept of generalized torques is presented. This method makes it possible to study the influence of the net muscle joint torques and gravity and inertia forces on the crank torque. The assumptions and limitations of the proposed method are discussed and this method is compared with the methods of analysis proposed by other researchers. In order to assess the validity of the method, some experimental data are elaborated. Results show that the method can highlight the effect of training and the pedaling technique of an athlete. PMID- 11277300 TI - Repetitive gait of passive bipedal mechanisms in a three-dimensional environment. AB - The inherent dynamics of bipedal, kneed mechanisms are studied with emphasis on the existence and stability, of repetitive gait in a three-dimensional environment, in the absence of external, active control. The investigation is motivated by observations that sustained anthropomorphic locomotion is largely a consequence of geometric and inertial properties of the mechanism. While the modeling excludes active control, the energy dissipated in ground and knee collisions is continuously re-injected by considering gait down slight inclines. The paper describes the dependence of the resulting passive gait in vertically constrained and unconstrained mechanisms on model parameters, such as ground compliance and ground slope. We also show the possibility of achieving statically unstable gait with appropriate parameter choices. PMID- 11277301 TI - Determination of the compressive material properties of the supraspinatus tendon. AB - A methodology was developed for determining the compressive properties of the supraspinatus tendon, based on finite element principles. Simplified three dimensional models ure re reated based on anatomical thickness measurements of unloaded supraspinatus tendons over 15 points. The tendon material was characterized as a composite structure of' longitudinally arranged collagen fibers within an extrafibrillar matrix. The matrix was formulated as a hyperelastic material described by the Ogden form of the strain energy potential. The hyperelastic material parameters were parametrically manipulated until the analytical load-displacement results were similar to the results obtaizned from indentation testinrg. In the geometrically averaged tendon, the average ratio of experimental to theoretical maximum indentation displacement was 1.00 (SD: 0.01). The average normalization of residuals was 2.1 g (SD: 0.9 g). Therefjore, the compressive material properties of the supraspinatus tendo'n extrafibrillar matrix were adequately derived with a first-order hyperelastic formulation. The initial comnpressive elastic modulus ranged from 0.024 to 0.090 MPa over the tendon surface and increased nonlinearly with additional compression. Using these material properties, the stresses induced during acromional impingement can be analyzed. PMID- 11277302 TI - A biphasic, anisotropic model of the aortic wall. AB - A biphasic, anisotropic elastic model of the aortict wall is developed and compared to literature values of experimental measurements of vessel wall radii, thickness, and hvdraulic conductivity as a function of intraluminal pressure. The model gives good predictions using a constant wall modulus for pressures less than 60 mmHg, but requires a strain-dependent modulus for pressures greater than this. In both bovine and rabbit aorta, the tangential modulus is found to be approximately 20 times greater than the radial modulus. These moduli lead to predictions that, when perfused in a cylindrical geometry, the aortic volume and its specific hydraulic coonductivity are relatively independent of perfusion pressure, in agreement with experimental measurements. M, the parameter that relates specific hydraulic conductivy, to tissue dilation, is found to be a positive quantity correcting a previous error in the literature. PMID- 11277303 TI - Modeling of the deformation of flexible tubes using a single law: application to veins of the lower limb in man. AB - The topic of this study mainly concerns a representative model of the behavior of flexible ducts such as elastic tubes or veins. This model is based on a phenomenological approach of the inflation and collapse of the tube. It leads to a single "universal" analytical expression of the tube law, valid fir a wide range of' positive and negative transmural pressures, which presents a significant improvement compared to previous theoretical studies defined with different expressions on restricted ranges of pressure. Moreover, the theoretical approaches most often require simplif'ing hypotheses--no longitudinal tension, no surrounding tissues--which are quite unrealistic both in the physiological case and in the experimental setup. These theoretical models can therefore be expected only roughly to describe the actual behavior of such vessels. The representative model, on the contrary, allows one to account for the deformation--inflating as well as collapse--of elastic tubes or veins with better accuracy. The tube law is a function of six parameters chosen in order to fit the experimental data. A comparison between results obtained in our laboratory using silicone tubes and representative models is presented. The model is then applied to physiological data obtained in vivo on human leg veins. PMID- 11277304 TI - Numerical simulation of streaming potentials due to deformation-induced hierarchical flows in cortical bone. AB - Bone is a very dynamic tissue capable of modiA,fing its composition, microstructure, and overall geometry in response to the changing biomechanical needs. Streaming potential has been hypothesized as a mechanotransduction mechanism that may allow osteocytes to sense their biomechanical environment. A correct understanding of the mechanism for streaming potential will illuminate our understanding of bone remodeling, such as the remodeling associated with exercise hypertrophy, disuse atrophy, and the bone remodeling arounid implants. In the current research, a numerical model based on the finite element discretization is proposed to simulate the fluid flows through the complicated hierarchical flow system and to calculate the concomitant stress generated potential (SGP) as a result of applied mechanical loading. The lacunae-canaliculi and the matrix microporosity are modeled together as discrete one-dimensional flow channels superposed in a biphasic poroelastic matrix. The cusplike electric potential distribution surrounding the Haversian canal that was experimentallv observed and reported in the literature earlier was successfully reproduced by the current numerical calculation. PMID- 11277305 TI - Hydrodynamic modeling of cerebrospinal fluid motion within the spinal cavity. AB - The fluid that resides within cranial and spinal cavities, cerebrospinal fluid (CSF), moves in a pulsatile fashion to and from the cranial cavity. This motion can be measured hy magnetic resonance imaging (MRI) and may he of clinical importance in the diagnosis of several brain and spinal cord disorders such as hydrocephalus, Chiari malformation, and syringomyelia. In the present work, a geometric and hydrodynamic characterization of an anatomically relevant spinal canal model is presented. We found that inertial effects dominate the flow field under normal physiological flow rates. Along the length of the spinal canal, hydraulic diameter was found to vary significantly from 5 to 15 mm. The instantaneous Reynolds number at peak flow rate ranged from 150 to 450, and the Womersle number ranged from 5 to 17. Pulsatile flow calculations are presented for an idealized geometric representation of the spinal cavity. A linearized Navier-Stokes model of the pulsatile CSF flow was constructed based on MRI flow rate measurements taken on a healthy volunteer. The numerical model was employed to investigate effects of cross-sectional geometry and spinal cord motion on unsteady velocity, shear stress, and pressure gradientfields. The velocity field was shown to be blunt, due to the inertial character of the flow, with velocity peaks located near the boundaries of the spinal canal rather than at the midpoint between boundaries. The pressure gradient waveform was found to be almost exclusively dependent on the flow waveform and cross-sectional area. Characterization of the CSF dynamics in normal and diseased states may be important in understanding the pathophysiology of CSF related disorders. Flow models coupled with MRI flow measurements mnay become a noninvasive tool to explain the abnormal dynamics of CSF in related brain disorders as well as to determine concentration and local distribution of drugs delivered into the CSF space. PMID- 11277306 TI - Pulsatile flow in an end-to-side vascular graft model: comparison of computations with experimental data. AB - Various hemodynamic factors have been implicated in vascular graft intimal hyperplasia, the major mechanism contributing to chronic failure of small diameter grafts. However, a thorough knowledge of the graft flow field is needed in order to determine the role of hemodynamics and how these factors affect the underlying biological processes. Computational fluid dynamics offers much more versatility and resolution than in vitro or in vivo methods, yet computations must be validated by careful comparison with experimental data. Whereas numerous numerical and in vitro simulations of arterial geometries have been reported, direct point-by-point comparisons of the two techniques are rare in the literature. We have conducted finite element computational analyses for a model of an end-to-side vascular graft and compared the results with experimental data obtained using laser-Doppler velocimetry. Agreement for velocity profiles is found to be good, with some clear differences near the recirculation zones during the deceleration and reverse-flow segments of the flow waveform. Wall shear stresses are determined from velocity gradients, whether by computational or experimental methods, and hence the agreement for this quantity, while still good, is less consistent than for velocity itself from the wall shear stress numerical results, we computed four variables that have been cited in the development of intiimal hyperplasia-the time-averaged wall shear stress, an oscillating shear index, and spatial and temporal wall shear stress gradients in order to illustrate the versatility of numerical methods. We conclude that the computational approach is a valid alternative to the experimental approach for quantitative hemodynamic studies. Where differences in velocity were found by the two methods, it was generally attributed to the inability of the numerical method to model the fluid dynamics when flow conditions are destabilizing. Differences in wall shear, in the absence of destabilizing phenomena, were more likely to be caused by difficulties in calculating wall shear from relatively low resolution in vitro data. PMID- 11277307 TI - Waveform dependence of pulsatile flow in a stenosed channel. AB - Bloodflow in arteries often shows a rich variety of vortical flows, which are dominated by the complex geometry of blood vessels, the dynamic pulsation of blood flow, and the complicated boundary conditions. With a two-dimensional model of unsteady flow in a stenosed channel, the pulsatile influence on such vortical fluid dynamics has been numerically studied in terms of waveform dependence on physiological pulsation. Results are presented for unsteady flows downstream of the stenosed portion with variation in the wavefiorms of systole and diastole. Overall, a train of propagating vortex waves is observed for all the cases, but it shows great sensitivity to the waveforms. The generation and development of the vortex waves may be linked to the presence of an adverse pressure gradient within a specific interval between two points of inflection of the systolic waveform. The adverse pressure gradient consists of a global pressure gradient that is found to be closely related to the dynamnics of' the pulsation, and a local pressure gradient, which is obsented to be dominated by the nonlinear vortex dynamics. PMID- 11277308 TI - Fluid dynamics of a textured blood-contacting surface. AB - This study examined the fluid dynamics of a textured blood-contacting surface using a computational fluid-dynamic modeling technique. The texture consisted of a regular array of microfibers of length 50 or 100 microm, spaced 100 microm apart, projecting perpendicularly to the surface. The results showed that the surface texture served as a flow-retarding solid boundary for a laminar viscous flow, resulting in a lowered wall shear stress on the hase-plane surface. However, the maximum wall shear stress on the fibers was much higher than the shear stress on the nontextured phase plane. At all fractions of fiber height down past 10 microm, the permeability of the textured region greatly exceeded the analytically predictable permeability of an equivalent array of infinite-height fihers. The lowered suiface shear stress appears to explain in part the enhanced deposition of formed blood elements on the textured surface. PMID- 11277309 TI - Pancreatic exocrine function in patients with type 1 and type 2 diabetes mellitus. AB - Reduced exocrine pancreatic function has been observed in a high percentage of patients with type 1 diabetes in the past. There are only few data for type 2 diabetes available and they are contradictory. In this study we investigated exocrine pancreatic function in 105 controls and 114 patients with type 1 or type 2 diabetes mellitus by means of an indirect test (faecal elastase-1 concentration). This test has good sensitivity and specificity for moderate and severe pancreatic insufficiency as compared to the gold standard. Reduced faecal elastase-1 concentrations were found in 56.7% of type 1 patients, 35% of type 2 patients and 18.1% of the controls. Elastase-1 concentrations did not correlate with alcohol consumption, diabetes duration or diabetes therapy. The data found for type 1 patients correspond to those reported in earlier studies. The results for type 2 diabetics show that exocrine pancreatic function is also impaired in a high percentage in this group of patients. Pathogenetic concepts to explain these findings as consequences of diabetes complications or insulin deficiency are still under debate. Observations from autopsies and the data of the controls in this study suggest that chronic pancreatitis might be a common problem. In consequence, diabetes secondary to exocrine disease could be much more frequent than believed so far. PMID- 11277311 TI - Chronic effects of streptozotocin-induced diabetes on the ultrastructure of rat ventricular and papillary muscle. AB - Contractile dysfunctions have been demonstrated in different experimental models of diabetes which have similar characteristics to many of the abnormalities found in the clinical setting. Administration of streptozotocin (STZ) to young adult rats induces beta-cell necrosis of the pancreas which gives rise to hypoinsulinaemia and hyperglycaemia, features which are also seen in untreated type 1 clinical diabetes. We have investigated the chronic effects of STZ-induced diabetes on contraction in rat ventricular myocytes and ultrastructure of cardiac muscle. Diabetes was induced in male Wistar rats (230-270 g) with a single injection of STZ (60 mg kg(-1)). At 2 and 10 months after STZ treatment, the amplitude of contraction was larger in diabetic compared to control myocytes. Time to peak contraction was significantly longer at 2 months but appeared to normalise at 10 months after STZ treatment. In contrast, time to half relaxation of contraction was not significantly different after 2 months but was significantly reduced at 10 months after STZ treatment compared to control. Transmission electron microscope examination of cardiac muscle showed that the ultrastructure of cardiac muscle, especially structures associated with contraction, were not greatly altered after STZ treatment. Sarcomere lengths were not significantly different in papillary or ventricular muscle at 4 or 8 months after STZ treatment compared to control. Our data provide evidence that morphological defects in contractile myofilaments and associated structures cannot explain contractile dysfunctions seen in ventricular myocytes from STZ treated animals. PMID- 11277310 TI - Effect of dietary supplementation with the pyridoindole antioxidant stobadine on antioxidant state and ultrastructure of diabetic rat myocardium. AB - Consistent with the postulated role of oxidative stress in the etiology of late diabetic complications, pharmacological interventions based on biological antioxidants have been suggested. The aim of the present study was to investigate the effect of dietary supplementation with the pyridoindole antioxidant stobadine on the myocardial antioxidant status and ultrastructure of streptozotocin diabetic rats. Diabetic male Wistar rats were fed for 32 weeks a standard diet or a diet supplemented with stobadine (0.05% w/w). Control rats received a standard diet or stobadine-supplemented diet (0.16% w/w). Plasma levels of glucose, cholesterol and triglycerides were increased significantly by diabetes. Activities of superoxide dismutase and catalase were markedly elevated in the diabetic myocardium. Myocardial levels of conjugated dienes increased after eight months of diabetes, in spite of significantly increased myocardial alpha tocopherol and coenzyme Q9 content. The long-term treatment of diabetic animals with stobadine (i) reduced plasma cholesterol and triglyceride levels yet left the severe hyperglycemia unaffected, (ii) reduced oxidative damage of myocardial tissue as measured by conjugated dienes, (iii) reversed myocardial levels of alpha-tocopherol and coenzyme Q9 to near control values, (iv) reduced elevated activity of superoxide dismutase in the diabetic myocardium, and (v) attenuated angiopathic and atherogenic processes in the myocardium as assessed by electron microscopy examination. These results are in accordance with the postulated prooxidant role of chronic hyperglycemia and provide further evidence that development of pathological changes in diabetic myocardium is amenable to pharmacological intervention by biological antioxidants. PMID- 11277312 TI - Relationship of residual beta-cell function, metabolic control and chronic complications in type 2 diabetes mellitus. AB - As the relationships between C-peptide levels and metabolic control and chronic complications are poorly known in type 2 diabetes, due to the slow decline of beta-cell function, we evaluated these associations in a cohort of type 2 diabetic patients. After excluding insulin-treated subjects, 1,533 patients were divided according to their C-peptide fasting levels in quartiles. Patients within the lowest C-peptide quartile showed significantly higher duration of diabetes, prevalence of retinopathy and values of HDL-cholesterol, albumin excretion rate and HbA1c, while BMI, diastolic blood pressure, percentages of hypertension and metabolic syndrome, and values of triglycerides and uric acid were significantly higher in the highest C-peptide quartile. The associations between C-peptide and duration of diabetes, AER, HbA1c, retinopathy and the components of the metabolic syndrome remained significant, after multiple adjustments. In conclusion, these data support the hypothesis that a reduced insulin secretion is associated with a longer duration of diabetes and a greater prevalence of microvascular complications, while higher insulin levels are associated with the components of the metabolic syndrome. PMID- 11277313 TI - Effect of ciprofibrate on lipoprotien metabolism and oxidative stress parameters in patients with type 2 diabetes mellitus and atherogenic lipoprotein phenotype. AB - The effect of ciprofibrate therapy on plasma lipids and lipoproteins, HDL and LDL subfraction profile, fractional esterification rate of HDL cholesterol (FER(HDL)) and the resistance of LDL and serum lipids to oxidation was studied in 24 males with type 2 diabetes and atherogenic lipoprotein phenotype (ALP). We also examined the effect of ciprofibrate therapy on oxidative DNA damage in peripheral lymphocytes. No differences in glucose, HbA1C and BMI levels were found after three months of ciprofibrate therapy. Ciprofibrate significantly decreased total cholesterol and triglyceride levels by 5.5% and 50% (p = 0.05; 0.001, respectively) and increased HDL-cholesterol levels by 8.5% (p = 0.05). FER(HDL) and LDL subfraction profile were also favorably affected. However, no effect on HDL subclasses was found. There were no statistically significant differences in lipid resistance to oxidation measured in serum and in LDL (lag time and Vmax) before and after therapy. No significant effect of ciprofibrate was found on oxidative DNA damage. The evaluation of the relationship between oxidative damage of purines with lag time in LDL and maximal rate of serum lipid oxidation showed significant correlations after therapy (r = -0.58; 0.47, p = 0.01; 0.05, respectively), but only trends before starting ciprofibrate treatment. Type 2 diabetes mellitus represents a complex metabolic disorder expressed in glucose and lipoprotein disturbances and increased oxidative stress. Ciprofibrate therapy favorably affected major features of lipid abnormalities of diabetic patients, but the level of oxidative stress assessed by in vitro and in vivo methods was not changed. The evaluation of expected logical correlations between the parameters of lipoprotein metabolism, lipid resistance in serum and LDL, and oxidative DNA damage showed that those correlations were more relevant and significant after ciprofibrate treatment and were not related with glucose homeostasis. PMID- 11277314 TI - In vitro effect of zinc on superoxide anion production by polymorphonuclear leukocytes of diabetic patients. AB - We evaluated the ability of zinc to modulate the superoxide anion released by isolated, phorbol myristate acetate (PMA)-stimulated polymorphonuclear leukocytes (PMNs). In the presence of zinc at physiological level (20 micromol/l), PMNs released 20% less superoxide anions than untreated cells, whereas at higher zinc concentration (200 micromol/l), a 40%-increase in superoxide anion production was detected. The direction (suppression or stimulation) of superoxide anion generation was dependent upon extracellular zinc level and did not discriminate type 1 and 2 diabetic subjects from normal controls. The mechanism(s) of dose dependent pro- and antioxidative effects of zinc on isolated PMNs in vitro as well as the implication of this finding for the modulatory effect of Zn on PMN superoxide anion production in vivo need further investigation. PMID- 11277315 TI - Insulin release form isolated, human islets after acute or prolonged exposure to glimepiride. PMID- 11277316 TI - Radiation induced chemotherapy sensitization in trimodality therapy of stage III non small cell lung cancer. A preliminary report. AB - The overall cure rate of locally advanced non-small cell lung carcinoma (NSCLC) remains poor. Although there have been encouraging reports of preoperative use of chemotherapy, more recent trend is the trimodal approach of radiation, chemo, and surgical-therapies. With the trimodal therapy, increased tumor response and resectability are reported, however, there are increased treatment related side effects. We observed that a relatively small dose of radiation given prior to induction chemotherapy greatly enhanced the tumor response to the chemotherapy without increased toxicity. A total of 18 patients (8 IIIA and 10 IIIB) were initially given 20 Gy of radiation therapy in 10 fractions and then received 2 courses of Taxol combination chemotherapy. The overall response rate was 83% (15/18) and 13 out of 18 patients underwent surgery. There was one postoperative death (not therapy related). It is speculated that the small dose of radiation therapy may have sensitized the tumor to subsequent chemotherapy, and we suggest a new hypothesis of "Radiation therapy induced chemotherapy sensitization". PMID- 11277317 TI - Clinicopathological features of gastric stromal tumors. AB - Stromal tumors in the gastrointestinal (GI) tract consist of myogenic tumors, neurogenic tumors and gastrointestinal stromal tumors (GISTs). Mutations in the c kit gene have been found in GISTs, and GISTs with c-kit mutations showed aggressive clinical behavior and histological features. In the present study, we classified stromal tumors into four groups according to histological differentiation and c-kit mutation: myogenic tumors, neurogenic tumors, c-kit mutation (-) GISTs and c-kit mutation (+) GISTs, and examined their clinicopathological importance and validity using data obtained from 125 patients with gastric stromal tumors. There was no difference in preoperative symptoms and signs among the four groups. GISTs with c-kit mutations were large and showed invasion into neighboring structures compared with the other tumors, indicating the clinically aggressive features of mutation (+) GISTs. In histological examinations, c-kit mutation (+) GISTs were higher in cellularity (P < 0.0001) and mitotic cell count (P = 0.0086), and showed frequent histological necrosis (P = 0.0058) and hemorrhage (P = 0.0170), and consequently, were higher in histological grade (P = 0.0001). In prognostic analyses, overall, cause-specific and disease-free survival of patients in the mutation (+) GIST group was the poorest among the four groups. No significant differences were found among the other three groups of myogenic tumors, neurogenic tumors and c-kit mutation (-) GISTs, indicating a similar aggressiveness in clinical presentation and histological features. Thus, this classification is considered to be clinically and pathologically important in the diagnosis of gastric stromal tumors. PMID- 11277318 TI - Role of increased arterial inflow in arm edema after modified radical mastectomy. AB - Chronic arm edema is a common finding after modified radical mastectomy and its pathophysiology is unclear. In a prospective study the value of increased arterial inflow and venous abnormalities after mastectomy was evaluated. Arterial and venous blood flow in axillary vessels of 39 patients with arm swelling and 16 patients without swelling were investigated by Doppler ultrasound. In patients with arm edema the arterial flow on the surgical treated side was 689.73+/-44.6 (mean+/-sem) ml/min and 427.73+/-30.8 ml/min on the contralateral side (p<0.05). In those without swelling the flow was 447.75+/-37.8 ml/min on the treated side and 354.95+/-28.7 ml/min on the contralateral side (p>0.05). The difference between arterial flow measurements on the treated sides of the patients with and without arm swelling was statistically significant. There was no significant difference between the measurements on the contralateral sides of both groups. Venous abnormalities were not detected in both groups of patients. We, therefore, conclude that modified radical mastectomy causes increased inflow in ipsilateral arm and it may play an important role in the etiology of arm swelling in breast cancer patients. PMID- 11277319 TI - Supportive treatment in weight-losing cancer patients due to the additive adverse effects of radiation treatment and/or chemotherapy. AB - The reversal of anorexia and weight loss especially in patients with advanced cancer suffering from radiation treatment (RT) -related complications and debilitated furtherly during RT would be a welcome relief. The purpose of this study is to evaluate the feasibility of supportive treatment with megestrol acetate (MA) in our weight-losing cancer patients increasingly experiencing anorexia, smell, taste, and weight loss due to the additive adverse effects of RT +/- chemotherapy and how MA changes the additive role of the severity of RT reactions on such patients. From June 1997 to October 1998, 100 eligible patients were enrolled on a randomized, placebo-controlled clinical trial. Of the 100 patients, 46 received MA during RT and 4 after the end of the RT, and 50 received placebo for 3 months. Subjective parameters were assessed by a brief questionnaire form based on scoring from 1 to 5, according to the degree of the loss or change for each parameter of malnutrition, appetite, taste and smell developed by us. At the end of the study a statistically significant weight gain was achieved in the patient group receiving MA compared to the placebo group (+3 to +5 kg versus -3.7 to -5.9 kg, p=0.000). Significant improvements were seen in performance status (p=0.000), appetite (p=0.000), malnutrition (p=0.000), loss of taste (p=0.000) and smell qualities (p=0.02) in the MA group compared to the placebo group. In the MA group there was no statistically significant difference related to the weight changes according to the grade of either the acute or late RT effects (p=0.65 and 0.07, respectively). Whereas, in the placebo group a statistically significant additive effect of the acute and late RT effects was detected on weight loss (p=0.008 and 0.007, respectively). We observed no side effects of MA in a 3-month time follow-up. The use of MA 480 mg/day during RT was effective in reversing anorexia and weight loss in spite of the acute RT effects, and helped most patients to well tolerate specific tumor therapy. Further evaluation of its mechanisms of action on RT-related adverse effects, tumor response relationships, and effect on patient survival are researched. PMID- 11277320 TI - Treatment of iatrogenic ureteral injuries during various operations for malignant conditions. AB - In the present study we report twenty-nine patients with iatrogenic injuries and management during various operations for malignant conditions. The patients were reviewed in order to identify and study the incidence, type of treatment administered and outcome. The study group was composed of 29 patients with 31 iatrogenic injuries between 1992 and 1999. General surgical, gynecological and urological procedures accounted for 24 (83%), 4 (14%), and 1 (3%) injuries respectively. Twenty-eight injuries were diagnosed at operation and three after an interval of 5, 17 and 45 days. Of the injuries, 51% occurred in the lower third of the ureter, 30% in the upper third and 19% in the middle third. Complete transsection, excision, ligation and partial transection accounted for 19 (61%), 9 (29%), 2 (7%), and 1 (3%) respectively. Treatment consisted of end-to-end ureteroureteral anastomosis in 18 cases; ileal interposition in 4 cases; transureteroureterostomy and ureteroneocystostomy in 2 cases; surgical repair, nephrectomy, ureterocutaneostomy and ileal loop in each of the remaining cases. Primary healing was obtained in all patients. General surgical procedures are the most common cause of iatrogenic injuries during malignant conditions. The proper identification and, when necessary, identification of the ureter at the pelvic brim, should decrease the incidence of iatrogenic ureteral injury. When identified at injury and treated immediately such injuries seldom lead to loss of renal function. We do not advocate to perform nephrectomy for any type of ureteric injury since the preservation of the kidney should be the aim of a surgeon. PMID- 11277321 TI - Oxidant and antioxidant status in larynx squamous cell carcinomas. AB - In this study we reviewed 20 patients with laryngeal squamous cell carcinoma to evaluate the oxidant and antioxidant status on tissue level. Superoxide Dismutase (SOD) and Catalase levels, two important antioxidant enzymes, and Malondialdehyde (MDA) levels, a valuable index of lipid peroxidation, were compared in cancerous and normal tissues of the patients. In cancerous tissue SOD activities were significantly lower than in the normal tissue, while there was no significant difference in MDA levels and Catalase activities. It was also observed that SOD activities significantly decreased as the histopathologic malignancy grades increased in cancerous tissue. These changes in oxidant and antioxidant status in carcinomatous tissue of the larynx are considered to be of great interest. PMID- 11277322 TI - Linkage of persistent cholangitis after bilioenterostomy with biliary carcinogenesis in hamsters. AB - Biliary carcinoma occurring after bilioenterostomy has been reported as a late complication of this surgical procedure. The present study was designed to determine if bilioenterostomy promotes biliary carcinogenesis, and also to clarify the relationship between biliary inflammation and biliary carcinogenesis in hamsters. Syrian hamsters underwent a simple laparotomy (SL), choledochoduodenostomy (CD) or choledochojejunostomy (CJ). All hamsters received subcutaneous injections of the chemical carcinogen, N-nitrosobis (2-oxopropyl) amine (BOP), and were sacrificed 20 weeks after surgery. Neoplastic lesions in the biliary tree were histologically examined, and the presence and degree of cholangitis was also evaluated with special reference to biliary carcinogenesis. The incidence of bile duct carcinoma was not significantly different among the three groups. Numerous bile duct carcinomas, however, were recognized in the bilioenterostomized animals, especially in the CJ group. Moreover, significant correlations between biliary carcinogenesis and the presence of cholangitis were noted in both the CD and CJ groups, but not in the SL control group. Severe cholangitis was evident in the CJ group, and the number of biliary carcinomas was well correlated with the degree of cholangitis. In conclusion, the risk of carcinoma in the biliary tract may increase when persistent cholangitis is present after biliary reconstruction. PMID- 11277323 TI - Inhibition of N-nitrosodiethylamine-induced hepatocarcinogenesis by Picroliv. AB - Picroliv, an iridoid glycoside mixture prepared from the roots and rhizomes of Picrorhiza kurroa was found to be an effective inhibitor of hepatocarcinogenesis induced by N-Nitrosodiethylamine (NDEA) in rats. Animals administered with NDEA had large hepatic nodules and the liver weight was increased to 6.17 +/- 1.0 g/100 g.b.wt. as compared to the normal liver weight 2.84 +/- 0.08 g/100 g.b.wt. Picroliv administration (200 mg/Kg.b.wt) reduced the liver weight to 3.30 +/- 0.23 g/100 g.b.wt. Oral administration of Picroliv reduced NDEA-induced elevation of gamma-glutamyltranspeptidase (gamma-GT) in serum and liver to that of normal rats. Moreover, elevated levels of bilirubin, alkaline phosphatase (ALP), glutamatepyruvate transaminase (GPT) and serum peroxides were also found to be significantly reduced by Picroliv administration. Similar observations were noticed in glutathione (GSH) and glutathione S-transferase (GST) levels. Histopathological analysis of the Picroliv treated rat liver resembles that of a normal liver except for a few alterations such as hepatocytomegalia and karyomegalia in some focii. The results are indicative of the chemopreventive potential of Picroliv against chemically-induced liver tumours. PMID- 11277324 TI - Drug-induced modulation of carcinoembryonic antigen (CEA) expression in neoplastic cells from a patient with rectal cancer. AB - Treatment with 5-fluorouracil (5-FU) or recombinant interferon-gamma (IFN), alone or in combination, was found to increase carcinoembryonic antigen (CEA) expression in several carcinoma cell lines. In this study we examined the in vitro effect of these agents on CEA expression of tumor cells, obtained from a patient operated for rectal cancer. The results showed that exposure of cancer cells to 5-FU or to IFN resulted in increased CEA levels in terms of percentage of CEA-positive cells and mean fluorescence values, as indicated by FACS analysis. However, drug combination did not induce CEA expression higher than that provided by single agents alone. Treatment with 5-FU or with IFN produced a reduction of the total number of viable cells. Moreover, Western blot analysis revealed that exposure of cancer cells to each drug was followed by a substantial increase of the total cellular CEA content. On the contrary, 5-FU in combination with IFN did not increase the expression of the antigen more than that obtained by single agents. Noteworthy, exposure of CEA-negative cells from adjacent normal rectal tissue to both agents alone or in combination, did not result in CEA induction. In conclusion, the present results suggest new approaches aimed at (a) increasing the sensitivity of diagnostic procedures based on detection of CEA positive tumor cells; (b) facilitating the recognition of CEA-positive cancer cells by immune responses induced by anti-CEA peptide vaccines. PMID- 11277325 TI - Immunohistochemical assessment of Ki-67 as prognostic cellular proliferation marker in anal canal carcinoma. AB - In order to define new prognostic factors useful for therapeutic decision-making, the Authors conducted a study on anal canal carcinomas in which Ki-67 proliferation index is correlated with pathological variables and clinical outcome. The Ki-67-detectable antigen is expressed in all stages of the cells cycle except G0. Thus, Ki-67 index can measure cell proliferation and it could be considered an indicator of prognosis. Thirty-one patients with anal canal carcinoma were evaluated. The specimens were formalin-fixed, paraffin-embedded and used for immunostaining of Ki-67 antigen. We found a significant correlation between Ki-67 score and depth of invasion and lymph node involvement. No correlation was found between high Ki-67 value and neoplastic relapse. These results suggest that Ki-67 positivity carries different significance in different cancers. Additional studies are required to ascertain whether more aggressive therapeutic procedures should be applied in the subset of patients with a high growth fraction. PMID- 11277326 TI - Utility of the serum tumor markers: CYFRA 21.1, carcinoembryonic antigen (CEA), and squamous cell carcinoma antigen (SCC) in squamous cell lung cancer. AB - The aim of this study is to assess the clinical usefulness of serum assays of carcinoembryonic antigen (CEA), squamous cell carcinoma antigen (SCC), and CYFRA 21.1 in the diagnosis of squamous cell lung cancer. Sixty patients with squamous cell, and twenty-four patients with nonsquamous cell histology of nonsmall cell lung cancer were enrolled in this study. Serum CEA, SCC, and CYFRA 21.1 levels were obtained by commercially available kits. Upper cutoff levels were 10 ng/ml, 3.5 ng/ml, and 3.5 ng/ml, respectively. In squamous cell lung cancer, percentages and 95% confidence interval (CI) of the patients with elevated levels were as follows: for CEA 23.3% (13-36), for SCC 20.0% (10-32), and for CYFRA 21.1 85.0% (73-93). The positivity rate of CYFRA 21.1 was more significant than CEA and SCC in both squamous and nonsquamous cell lung cancer. None of the markers were significant in differentiating squamous/nonsquamous histology. Only tumor marker CEA was significantly elevated in metastatic squamous cell lung cancer (p=0.004). A novel tumor marker CYFRA 21.1 can be used as a reliable tumor marker in diagnosing squamous cell lung cancer. In addition, CEA has an important role in determining metastatic disease. PMID- 11277327 TI - Adriamycin induced G2/M cell cycle arrest in transitional cell cancer cells with wt p53 and p21(WAF1/CIP1) genes. AB - Adriamycin (ADM), widely used for systemic and local treatment of bladder tumors, triggers apoptosis in bladder cancer cells. Here we investigated the effect of ADM on cell cycle progression and expression of cell cycle regulating proteins in bladder cancer cell lines with various p53 and p21(WAF1/CIP1) status. Flowcytometric analysis was used to estimate the cell cycle distribution of T24, HT-1376, RT4, and SCaBER bladder cancer cell lines. Cell cycle regulating proteins were analyzed by Immunoblot. Treatment of RT4 cells, bearing wild type p53 and p21(WAF1/CIP1), with ADM induced expression of both proteins and cell cycle arrest, not in G1, as was anticipated, but in the G2 phase. Simultaneously, Retinoblastoma (Rb) protein expression was decreased. Expression of PCNA, which is a target gene of E2F, was not changed. The results suggest that even if the tumor cells bear wild type (wt) p53 and wt p21(WAF1/CIP1) and both proteins accumulate due to genotoxic stimuli, the cell cycle arrest might happen not in the G1 but in the G2 phase. PMID- 11277329 TI - Early tumor growth in metastatic organs influenced by the microenvironment is an important factor which provides organ specificity of colon cancer metastasis. AB - We have previously demonstrated that liver metastases in nude mice and lung metastases in nude rats occurred specifically, when KM12SM human colon carcinoma cells were inoculated orthotopically into the cecal wall of nude mice and rats. To clarify the relationship between the tumor growth potential in the metastatic organs and the metastatic organ preference in these two metastatic models, we have evaluated the in vitro cell growth activities affected by the organ conditioned medium (CM) from the liver and lung, and the in vivo growth activities of the ectopic implanted tumors in the liver and lung. The tumorigenicity of the ectopic implanted tumors was 100% in mouse liver, 33% in rat liver, 50% in mouse lung, and 75% in rat lung. The crude liver CM of the animals showed inhibitory activities for KM12SM cell growth in a dosage-dependent manner, and the crude lung CM stimulated KM12SM cell growth. The liver CM of nude mice inhibited the KM12SM cell growth more strongly compared with the CM of nude rats, and the lung CM of nude rats was more strongly stimulated compared with the CM of nude mice. The liver CM of nude mice had non-heparin binding factors, which stimulated or inhibited KM12SM cell growth, in a molecular weight range of 50 to 100 kDa. By contrast, the liver CM of nude rats showed no growth stimulating activity for KM12SM cells. These results suggest that the metastatic organ specificity of KM12SM cells may depend on the early tumor growth influenced by the microenvironment in metastatic organs. PMID- 11277328 TI - Mutated p53 in tumors, mutant p53 and p53-specific antibodies in the circulation in patients with gastric cancer. AB - The accumulation of mutated p53 in tumor cells results in the presence in the circulation of mutant p53 (p53m) and the production of p53-specific antibodies (p53Ab). We examined the relationships among these phenomena and analyzed their clinical implications in 62 patients with gastric cancer at various stages. Expression of p53 in tumors was studied by an immunohistochemical method and circulating p53m and p53Ab were quantitated with commercially available enzyme linked immunosorbent assay kits. The detectable expression of p53 in tumors and circulating p53Ab was recognized in 28 (45.2%) and 7 (11.3%) of the 62 patients, respectively. The number of patients with higher levels of circulating p53m increased with the progression of the depth of cancer invasion. Patients with any positive findings for the three p53-related parameters had a poorer prognosis, and the difference was statistically significant in patients with p53Ab. When survival was analyzed in terms of the combination of the three p53-related parameters (detectable expression of p53 in tumor cells, high levels of p53m and p53Ab in the circulation), a significantly poorer prognosis was associated with an increase in the number of positive parameters. Analysis of p53 in tumor cells, together with analysis of circulating p53m and p53Ab, could improve the accuracy of prognosis in patients with gastric cancer. PMID- 11277330 TI - Stimulation of megakaryocytopoiesis and platelet production during growth of an experimental lymphoma. AB - The effect of malignant tumor growth on host's megakaryocytopoiesis and platelet production was studied in mice bearing transplantable Dalton's lymphoma. Tumor growth was paralleled by thrombocytosis, neutrophilia, and anemia. Platelet 51Cr half-life was normal but incorporation of 75Selenomethionine into circulating platelets was significantly enhanced in the tumor bearers suggesting stimulated thrombopoiesis while platelet life span remained unchanged. Megakaryocytes and their precursors, the small acetyl cholinesterase positive cells, were found in increased numbers in the bone marrow (BM) and particularly in the spleen where five to eight-fold rise was observed at the log phase of tumor growth. In addition, a remarkable increase in the number of megakaryocyte progenitors (CFU MK and MK CFU-S) was observed both in the BM and spleen. Stimulation of these progenitors was more pronounced in the spleen than in the marrow, and the change was noticeable even from the third day of tumor bearing. Therefore, the results suggest that thrombocytosis associated with the growth of this experimental lymphoma was due to accelerated platelet production following stimulated megakaryocytopoiesis especially in the spleen. PMID- 11277331 TI - Evidence of chromosome instability in chronic pancreatitis. AB - In the current study we analyzed chromosome instability on peripheral blood lymphocytes cultured from 7 untreated patients with chronic pancreatitis (CP) by assessing telomeric associations (TAS), chromosome aberrations (CA) and sister chromatid exchanges (SCE). Seven healthy individuals were also analyzed. Mean frequencies of TAS were significantly higher in CP patients (X +/- SE: 11.00 +/- 2.37) compared to controls (1.00 +/- 0.30) (p<0.001). Chromosomes preferentially involved in TAS were: 9, 20, 16 and 21, being the most affected arms: 9p, 20q, 16p, 9q and 21q. All these terminal bands were coincident with cancer breakpoints (p<0.03), two of them (40%) were specifically associated to pancreatic carcinoma rearrangements. Three bands (60%) were coincident with oncogene location. The mean frequency of CA was significantly higher in patients (3.88 +/- 0.80) compared to controls (0.63 +/- 0.49) (p<0.001). Chromosomes 1, 2 and 13 were the most damaged. No specifically affected breakpoints were found. SCE analysis showed higher levels in patients (8.33 +/- 0.70) than in controls (6.62 +/- 0.34) (p<0.025), but no differences were observed in cell cycle kinetics. Our results clearly indicate that CP patients exhibit chromosome instability, showing the presence of an unstable genome that could be related to the cancer development observed in this disease. PMID- 11277332 TI - Altered expression of jack fruit lectin specific glycoconjugates in benign and malignant human colorectum. AB - Lectins bind to terminal non reducing sugars of glycoconjugates in cell membranes and secretions. These glycoproteins can be very characteristic, both for the differentiation state of a tissue and for its grade of malignancy. The carbohydrate structures of cellular glycoconjugates in normal, adenoma, and carcinoma of human colorectum were analysed using HRP conjugated Jack Fruit Lectin (JFL). Fifty two rectal carcinomas, 18 adenomas and 25 normal rectal tissues were used for the study. Diaminobenzidine (DAB) was used as the visualant. Normal rectal tissues showed positive reaction with intense staining in more than 60% of the cells. Adenomas were seen to have moderate to intense staining with a range of 30 to 60% positive cells. The lowest levels of JFL staining were observed in invasive tumours. JFL binding was found to decrease with carcinomatous cells compared to adenomas and normal cells. A significant negative association was found between JFL binding and histologic abnormality (r = -0.85, P <0.0000). These results suggest that there are alterations in the carbohydrate structures of cellular glycoconjugates, which can be related to goblet cell differentiation, in normal, benign and malignant human colorectal tissue. PMID- 11277333 TI - P53 overexpression in normal oral mucosa of heavy smokers. AB - The observation that p53 alterations are early events in the tumorigenesis of head and neck cancer and the association with cigarette smoking have prompted us to search for p53 overexpression in the oral mucosa of heavy smokers who have no overt precancerous or cancerous lesions. Formalin-fixed paraffin embedded tissue sections, obtained from oral mucosa of 30 otherwise healthy heavy smokers, were evaluated for tobacco related changes, and immunostained with a mouse monoclonal antibody p53-DO7 for p53 immunoreactivity. Histopathological evaluation revealed hyperplastic changes in twenty eight samples (93%), eight of which also demonstrated dysplastic changes. Positive immunoreaction for p53 was detected in six (20%) of the tissue samples. The study provided significant information about the frequency of hyperplasia, dysplasia, and p53 overexpression in individuals who were heavy smokers. It is suggested, also, that chemoprevention might have some impact in this particular group of individuals. PMID- 11277334 TI - Functional membrane changes due to tumor induction in rat pituitary cell cultures. AB - Membrane functions in tumorous cells are different from those in healthy cells. The aim of the present study was to investigate the changes in pituitary cell membrane functions and hormone secretion after tumor induction in vivo and in vitro. Prolactinomas were induced in vivo in female Wistar rats with estrone acetate. Normal anterior pituitaries and prolactinomas of female Wistar rats were dissociated enzymatically and mechanically, then cultured on collagen-treated plastic dishes. Some normal anterior pituitary cultures were treated with benz(c)acridines as tumorigenic agents in vitro. Intracellular 3',5'-cyclic adenosine monophosphate (cAMP) levels were determined by a competitive binding technique, membrane fluidity was assayed by fluorescence anisotropy, and ATP-ase activities were estimated via ATP loss. The results indicated decreased membrane fluidity in tumorous cell cultures. However, in vitro benz(c)acridine treatment exerted more pronounced effects than those observed after in vivo estrone treatment. The ATP-ase activities were highly increased in benz(c)acridine treated cells and in estrogen-induced prolactinoma cells, more strongly so in the former ones. The intracellular cAMP levels were higher than normal in both of them. The results concerning the ACTH, alpha-MSH, PRL and GH levels of normal and tumorous cell cultures were published in our previous study. Our findings show that the tumorous transformation of pituitary cells can cause significant changes in functional membrane parameters and hormone secretion. Decreased membrane fluidity was accompanied by an increased exocytosis (hormone release) and adenylate cyclase activity in tumorous cells. PMID- 11277335 TI - Pseudomixoma peritonei: a case report. AB - Pseudomixoma Peritonei (PMP) is an uncommon neoplasm characterised by mucinous ascites and multifocal amorphous mucous substances involving the peritoneal surface, omentum and bowel loops. Although the origin of the Pseudomixoma Peritonei is still unclear, it could be due to the perforation of an ovarian mucinous cystoadenoma or an appendiceal mucocele. The further pelvic dissemination of the endotumor material, which adhere itself into the peritoneal surface, may induce an intra-abdominal transformation of the peritoneal mesothelium into mucin-producing tissue. A case of Pseudomyxoma Peritonei (PMP) which occurred in a young woman is reported. PMID- 11277336 TI - Two siblings with chronic myelogenous leukemia. AB - In the present study we report the occurrence of Chronic Myelogenous Leukemia in two siblings (brother and sister) 47 and 62 years old respectively. These two cases raised the question of a possible genetic prodiathesis or a possible trasmission (environmental factor) cause. PMID- 11277337 TI - Home monitoring with the ClearPlan Easy Fertility Monitor for fertility awareness. AB - Many couples planning a pregnancy have had to gauge the best time for sexual intercourse from subjective indications, e.g. cervical mucus changes, or retrospective information, e.g. temperature and calendar methods. Between women, and within a woman, there are considerable variations in menstrual cycle length and underlying hormonal patterns. Although fertility test kits are available, most still rely on visual interpretation by the user. A new combination of immunochemical testing and computing technology is now available in the ClearPlan Easy Fertility Monitor. This paper describes the features of this accurate, simple-to-use, home-based fertility monitor, which gathers data on hormonal patterns and takes account of the individuality of menstrual cycles in women. These data and information about other events, such as sexual intercourse, can be stored in the monitor and accessed by the couple and their medical professional for retrospective review. The monitor is useful for couples who have just started trying to conceive, or those who have been planning a pregnancy for some time. PMID- 11277339 TI - Trial protocol and sample result of a study comparing the Clearplan Easy Fertility Monitor with serum hormone and vaginal ultrasound measurements in the determination of ovulation. AB - This paper presents a preliminary result from a study designed to evaluate the ability of the ClearPlan Easy Fertility Monitor to predict ovulation, compared with conventional methods of ovulation determination, i.e. serum hormone measurements (estradiol and luteinizing hormone) and vaginal ultrasound scans (follicle size and endometrium thickness). Fifty-five female volunteers will be monitored over 3 consecutive monthly cycles. Preliminary data gathered from one volunteer are included in this presentation. These data indicate that the ClearPlan Easy Fertility Monitor predicted ovulation as accurately as serum hormone measurements and vaginal ultrasound scans. The results show that this monitor has potential as a home-based ovulation predictor for couples attempting to conceive. PMID- 11277340 TI - Acceptability of home monitoring as an aid to conception. AB - This paper considers the concept of consumer acceptance of medical products and its importance to successful healthcare provision. The critical dimensions ('domains') of acceptance for a product designed to assist family planning, and the importance of using high-quality psychometric scales to assess these domains, are discussed. Qualitative and quantitative data from acceptance studies with a personal contraceptive monitor, PERSONA, are presented. These data indicate that the monitor is user-friendly and acceptable to couples, and underline the importance of user acceptability for home monitors designed to assist family planning--including those used for conception purposes. PMID- 11277341 TI - Potential fertility--defining the window of opportunity. AB - The ClearPlan Easy Fertility Monitor predicts the times of high potential fertility, i.e. the times when sexual intercourse is most likely to result in conception. The test procedure identifies the times of specified changes in the concentration of urinary hormone metabolites during each ovarian cycle. The results are displayed in terms of prior knowledge about the time-specific probabilities of conception. This paper contains a summary of the scientific background that led to the development of the ClearPlan Easy Fertility Monitor. PMID- 11277342 TI - Aldosterone escape during angiotensin-converting enzyme inhibitor therapy in essential hypertensive patients with left ventricular hypertrophy. AB - Continuous angiotensin-converting enzyme (ACE) inhibitor therapy does not necessarily produce significant decreases in plasma aldosterone levels (aldosterone escape). We examined the role of aldosterone escape in 75 essential hypertensive patients treated with an ACE inhibitor (enalapril maleate [34 patients], imidapril hydrochloride [24 patients] or trandolapril [17 patients]) for 40 weeks. With treatment, blood pressure decreased and plasma renin activity increased, while plasma aldosterone concentrations did not change. Aldosterone escape was observed in 38 of the 75 patients and in 17 of 37 patients with left ventricular hypertrophy before treatment. Left ventricular mass index did not change in patients with aldosterone escape but decreased significantly in patients without aldosterone escape. The present study demonstrated a high incidence of aldosterone escape in patients with essential hypertension despite the use of ACE inhibitors. The results also suggest that aldosterone escape may reverse the beneficial effects of an ACE inhibitor on left ventricular hypertrophy. PMID- 11277343 TI - Efficacy and tolerability of Hairgain in individuals with hair loss: a placebo controlled, double-blind study. AB - This randomized, placebo-controlled, double-blind study was designed to investigate the efficacy and tolerability of a new agent for the treatment of hair loss, based on a marine protein, minerals and vitamins. Sixty subjects with hair loss of different aetiologies participated in the 6-month blinded phase of the study. Objective assessments indicated that the treatment was effective and subjective assessments showed a statistically significant positive effect of treatment. Exposure to the active preparation for a further 6 months in an open phase indicated a further improvement in hair growth. Exposure of the patients previously treated with placebo to the active preparation for 12 months gave similar results. Tolerability was good and no side-effects were reported. The product investigated may provide an alternative to pharmacotherapy for the treatment of hair-loss problems in individuals with androgenic alopecia. PMID- 11277344 TI - Anti-pituitary antibody-induced multiple endocrine disorders in mice. AB - Circulating anti-pituitary antibodies (APAs) have been detected in patients with autoimmune diseases, although the role of autoantibodies in the pathogenesis of autoimmune diseases is still unclear. With the aim of elucidating the autoimmune mechanisms involved in patients with multiple endocrine disorders, we evaluated the pathological changes in the pituitary gland, thyroid, pancreas and adrenal gland of mice, both wild-type and using a murine model of autoimmune thyroid disease [MRL-lpr/lpr] that had been immunized with murine, rat, porcine or human pituitary glands. In four of seven mice, a 22 kD band corresponding to APA was detected by Western blotting in the serum from mice that had been immunized with human pituitary tissues but not in the serum from mice immunized with rat or pig tissue. Inflammatory changes were detected in all groups of mice, occurring in the hypophysis, pancreas and adrenal glands but not in the thyroid. In conclusion, APA-induced autoimmune endocrine disorders are likely to be important for studying the mechanisms involved in autoimmune syndromes. PMID- 11277345 TI - Efficacy of acetylcholinesterase inhibitors versus nootropics in Alzheimer's disease: a retrospective, longitudinal study. AB - The aim of this study was to investigate the efficacy of nootropics (piracetam, aniracetam, nimodopine and dihydroergicristine) versus acetylcholinesterase inhibitors (AChE-Is) (tacrine and donepezil) in the treatment of Alzheimer's disease. This is a retrospective study of 510 patients with Alzheimer's disease. To determine clinical efficacy of treatment, we used the mean change over time in scores for the following tests: the Mini-Mental State Examination (MMSE); the Cambridge Cognitive Examination for the Elderly; and the Functional Rating Scale for Symptoms of Dementia. In all patients and in patients with severe Alzheimer's disease (baseline MMSE < 11), no significant differences were seen in the neuropsychological test scores between the two treatment groups. In patients with moderate dementia (baseline MMSE between 11 and 20), however, there was a significantly greater deterioration, as shown on the CAMCOG scale, after 12 months' treatment for patients receiving AChE-Is compared with those receiving nootropics (-4.38 for AChE-Is group versus 1.48 for nootropics group). For patients with mild dementia (baseline MMSE score between 21 and 26), there was a significantly greater deterioration on the MMSE scale for each time-point in the nootropics group compared with the AChE-Is group. In conclusion, we did not find any strong evidence that a difference in efficacy exists between AChE-Is and nootropics in the treatment of Alzheimer's disease. PMID- 11277346 TI - Cardiac beriberi among illegal mainland Chinese immigrants. AB - The most common symptoms of chronic beriberi due to thiamine deficiency include dyspnoea, fatigue, leg oedema, lower extremity weakness and numbness. When collapsed peripheral circulation, metabolic acidosis, or shock are present, the disease has advanced from chronic beriberi to pernicious or fulminating beriberi heart failure (Shoshin beriberi). We report two patients with fulminating beriberi; both of whom had been incarcerated at a detention centre for 5 months before hospitalization. A prolonged monotonous diet, low in thiamine, was a major risk factor in both patients. Thiamine deficiency should be considered for any patient with symptoms and signs compatible with beriberi. PMID- 11277348 TI - C-reactive protein values and erythrocyte sedimentation rates after total hip and total knee arthroplasty. AB - We studied the changes in serum C-reactive protein levels (CRP) and erythrocyte sedimentation rates (ESR) in patients with primary osteoarthritis, who underwent uncomplicated arthroplasty. Of the 28 patients studied, 12 had cementless total hip replacement (THR), and 16 underwent cemented total knee replacement (TKR) with a tourniquet. In both groups serum CRP levels increased rapidly after surgery, peaking on day 2 (THR 23.17 mg/dl, TKR 26.02 mg/dl), and dropping gradually to pre-operative values on day 21 in THR patients and at the end of the second month in TKR patients. ESR peaked on day 5 after operation (THR 100.5 mm/h, TKR 101.3 mm/h), dropping close to pre-operative values at the end of the third month in THR patients and at the end of the ninth month in TKR patients, although, even after a year, ESRs were slightly above their pre-operative values. Serum CRP levels changed more rapidly than ESRs and returned to normal more rapidly. CRP and ESR values tended to be higher in TKR than in THR patients. PMID- 11277347 TI - The CAG regimen (low-dose cytarabine, aclarubicin hydrochloride and granulocyte colony-stimulating factor) for the treatment of elderly acute myelomonocytic leukaemia: a case study. AB - Elderly patients with acute myelomonocytic leukaemia (AMMoL) frequently have a poor quality of life after induction of remission using high-intensity treatment; we seek a more appropriate regimen for such patients. An 86-year-old man was hospitalized with a diagnosis of AMMoL (FAB classification M4), of abnormal karyotype, and complications of diabetes mellitus and complete right bundle branch block. He was treated with CAG therapy (cytarabine 10 mg/m2 subcutaneously every 12 h for 14 consecutive days; aclarubicin hydrochloride 10 mg/m2 per day, bolus intravenously for 4 consecutive days; granulocyte colony-stimulating factor 100 microg/day, subcutaneous injection for 14 consecutive days) every 3 months. White blood cell counts were at their lowest (around 600 - 800/microl) 12 days after the end of therapy, but returned to about 2000 - 2300/microl 30 days after stopping therapy. No symptoms of drug-related toxicity, except slight nausea, were found. Complete remission with a good quality of life was induced and lasted over 2 years suggesting that CAG therapy might prove effective in elderly patients with AMMoL. PMID- 11277349 TI - Correlates of insight in serious mental illness. AB - This study extends research into insight by examining its relationship to a variety of demographic, clinical, neurocognitive, and psychosocial variables among a broad diagnostic sample of 211 adults with serious mental illness. Participants completed a full battery of instruments measuring these variables. Results support a relationship between ratings of poor insight and a psychotic (vs. mood) diagnosis, increased psychiatric symptoms, poorer social skills, and negative medication attitudes. Minorities and those with a substance abuse diagnosis were also more likely to be rated as having poor insight. No relationship was found between level of insight and age, gender, education level, neurocognitive deficits, hospitalization history, size of one's social network, or quality of life measures. Results are discussed in the context of improving the measurement and assessment of insight, conceptualizing interventions aimed at addressing level of insight, and improving outcomes for patients with severe and persistent mental illness. Findings also support a need for continued investigation of how mental illness is understood, experienced, and expressed across diverse groups of people living with mental illness. PMID- 11277350 TI - Characteristics of patients with schizophrenia who do not believe they are mentally ill. AB - The purpose of this study was to use a very simple self-report measure to identify patients who did not believe they were mentally ill and describe their characteristics. The study included 177 inpatients and outpatients with schizophrenia. Multivariate regression methods analyzed the relationship between illness belief and sociodemographic, clinical, and attitudinal factors. Thirty seven percent of subjects did not believe they were mentally ill. Younger age, fewer depressive symptoms, lower perceived medication efficacy, greater satisfaction with current mental health, and less concern about mental illness stigma were associated with not believing one was mentally ill. Outpatients with fewer hospitalizations were less likely to believe they were ill. Inpatients with more hospitalizations were less likely to believe they were ill and had poor medication adherence. Readily identifying patients who do not believe they are mentally ill may be useful to clinicians and policymakers when matching at-risk patients with adherence interventions. PMID- 11277351 TI - Phobic, panic, and major depressive disorders and the five-factor model of personality. AB - This study investigated five-factor model personality traits in anxiety (simple phobia, social phobia, agoraphobia, and panic disorder) and major depressive disorders in a population-based sample. In the Baltimore Epidemiologic Catchment Area Follow-up Study, psychiatrists administered the Schedules for Clinical Assessment in Neuropsychiatry to 333 adult subjects who also completed the Revised NEO Personality Inventory. All of the disorders except simple phobia were associated with high neuroticism. Social phobia and agoraphobia were associated with low extraversion. In addition, lower-order facets of extraversion, agreeableness, and conscientiousness were associated with certain disorders (i.e., low positive emotions in panic disorder; low trust and compliance in certain phobias; and low competence, achievement striving, and self-discipline in several disorders). This study emphasizes the utility of lower-order personality assessments and underscores the need for further research on personality/psychopathology etiologic relationships. PMID- 11277352 TI - Anger, impulsivity, social support, and suicide risk in patients with posttraumatic stress disorder. AB - An emerging literature suggests that posttraumatic stress disorder (PTSD) patients are at an increased risk for suicide. The objective of this study was: a) to reexamine the relationship between PTSD and suicide by comparing suicide risks of persons with PTSD, to persons with anxiety disorder and to matched controls; and b) to examine the relationship between anger, impulsivity, social support and suicidality in PTSD and other anxiety disorders. Forty-six patients suffering from PTSD were compared with 42 non-PTSD anxiety disorder patients and with 50 healthy controls on measures of anger, impulsivity, social support, and suicide risk. Persons with PTSD had the highest scores on the measures of suicide risk, anger, and impulsivity and the lowest scores on social support. Multivariate analysis revealed that in the PTSD group, impulsivity was positively correlated with suicide risk and anger was not. PTSD symptoms of intrusion and avoidance were only mildly correlated with suicide risk at the bivariate level but not at the multivariate level. For the PTSD and anxiety disorder groups, the greater the social support, the lower the risk of suicide. For the controls, social support and impulsivity were not related to suicide risk, whereas anger was. These findings suggest that persons with PTSD are at higher risk for suicide and that in assessing suicide risk among persons with PTSD, careful attention should be paid to levels of impulsivity, which may increase suicide risk, and to social support, which may reduce the risk. PMID- 11277353 TI - Life stress and the symptoms of major depression. AB - Life stress has been found to be associated with onset of depression and with greater severity of depressive symptoms. It is unclear, though, if life stress is related to particular classes or specific symptoms in depression. The association between severe life events and depressive symptoms was tested in 59 individuals diagnosed by Research Diagnostic Criteria with endogenous primary nonpsychotic major depression. As predicted, life stress was associated principally with cognitive-affective symptoms, not somatic symptoms. There also was a consistent association across different assessment methods between severe events and suicidal ideation. Finally, associations held specifically for severe events occurring before onset, not for severe events occurring after onset. Symptom variation in major depression is related specifically to severe stressors before onset and includes primarily cognitive-affective types of symptoms. There is an especially pronounced association of prior severe stress with suicidal ideation. The implications of stress-symptom associations are addressed for enlarging understanding of symptom heterogeneity and subtype distinctions in major depression. PMID- 11277354 TI - Neuropsychological evaluation of higher functioning homeless persons: a comparison of an abbreviated test battery to the mini-mental state exam. AB - This study examined neuropsychological functioning in a heterogeneous population of persons who were homeless (N = 60) and compared the value of the Abbreviated Halstead-Reitan Test Battery with the Mini-Mental State Exam (MMSE). A high incidence of neuropsychological dysfunction was evident with 80% of patients showing impaired test battery performance and 35% showing an impaired MMSE. Performance on the Trail Making Test, Part B was especially impaired. Patients impaired on Trails B more often showed impaired test battery performance, suggesting it may be a better screening tool than the MMSE. Neuropsychological performance was not significantly affected by the patients' gender, age, diagnosis, or past psychiatric and medical history. Regression analysis suggested that 29% of the variance in test battery performance was accounted for by the patients' education. Results support previous findings that large numbers of people who are homeless are neuropsychologically impaired; this should be considered when planning treatment and rehabilitation. PMID- 11277355 TI - The neglect of response bias in mental health research. AB - A number of researchers have observed that response biases, defined as when subjects respond to items in research instruments in ways that do not coincide with the intent or content of the instrument, suffuse measurements and assessments of mental disorders. They cautioned that the response bias problem has been neglected in mental health research at the price of substantial error. Have the cautions been heeded? Or does the neglect of response bias continue? Articles published in 1998 in three major psychiatric journals were examined: Archives of General Psychiatry, American Journal of Psychiatry, and the Journal of Nervous and Mental Disease. The articles were examined to determine whether response biases were mentioned and whether systematic efforts were made to attend to their influence on the findings of the study. Each article was assessed twice by independent raters. The examination indicates that a very small minority of the articles reviewed mentioned response bias and that among those mentioning it, a minority attempted to control for bias effects. Cautions offered about response bias have not been heeded. Accordingly, the issue is one of how to incorporate concerns about response bias into the institutional structures that influence the culture of mental health research. PMID- 11277356 TI - Frontal lobe dysfunctions, dissociation, and trauma self-reports in forensic psychiatric patients. PMID- 11277357 TI - Transliminality, brain function, and synesthesia. PMID- 11277358 TI - The relationship of pathologic skin picking to obsessive-compulsive disorder. PMID- 11277359 TI - S100B protein, glia and Gilles de la Tourette syndrome. AB - Activated glial cells play an important role in a variety of neurological disorders. This study examines S100B protein levels in the serum of patients with Gilles de la Tourette syndrome, as potential marker for glial cell function. Two groups of children were examined: 61 reference patients and 33 patients with Gilles de la Tourette syndrome. It was found that S100B serum concentrations in the reference group decrease with increasing age. Furthermore it was found that the mean S100B concentration in serum of children with Gilles de la Tourette syndrome is significantly higher than in the reference group. These preliminary results suggest that glial tissue might be involved in the pathophysiology of the syndrome. PMID- 11277360 TI - Neurodevelopmental risks in twin-to-twin transfusion syndrome: preliminary findings. AB - The purpose of this study was to determine the neurodevelopmental risks in patients with twin-to-twin transfusion syndrome, a rare but serious complication of monochorionic twin gestations. From a total sample of 94 twins with twin-to twin transfusion syndrome, admitted during 1989 and 1993, 49 patients survived and 40 patients were followed to a mean age of 24 months. Neurological status and psychomotor development (Denver and Griffiths Developmental Tests) were determined. Parameters of the neonatal period were evaluated for their potential prediction. Of the 40 tested patients 18 showed a normal psychomotor development. Thirteen patients exibited a specific delay in language development and/or showed minor neurological dysfunctions. Nine twins had severe psychomotor retardation in combination with cerebral palsy. Major neurological sequelae were found, more common in recipients than in donors (6/19 vs 3/21). Correspondingly, neonatal ultrasound showed more pathological results (especially periventricular leucomalacia) in recipients. Neither anaemia nor polycythaemia at birth can predict developmental outcome. Apart from a high prenatal mortality rate, both twins, donators as well as recipients, are highly at risk for brain damage of different aetiology, associated with abnormal neonatal cerebral ultrasound. PMID- 11277361 TI - Hypoxic-ischaemic brain damage in immature rats: effects of adrenoceptor modulation. AB - The purpose of the present study was to evaluate the role of adrenergic receptors in the cascade leading to hypoxic-ischaemic brain injury in neonatal rats. The effect of adrenergic agents (prazosin, yohimbine, idazoxan and clonidine) administered before or after hypoxia-ischaemia was evaluated with respect to mortality and brain injury. Rat pups of either 7 or 8 days of age were subjected to unilateral carotid artery ligation combined with hypoxia (6% or 8% O2 in N2). The mortality was higher in hypoxic-ischaemic groups pre-treated with the alpha adrenergic receptor antagonists prazosin (48%) or yohimbine (53%) than in saline controls (7%). After 2 weeks the severity of the brain injury was evaluated in the surviving rats. Unilateral brain injury, evaluated by brain weight deficit of the injured ipsilateral hemisphere compared with the contralateral hemisphere, was 17.8 +/- 4.9% and 27.1 +/- 4.0% in pre- and post-treated saline groups, respectively. Post-treatment with clonidine, an alpha2-adrenergic agonist, reduced brain injury by 45% (p < 0.05) compared with saline controls. Pre treatment with the same drug was not effective. Idazoxan had no effect on brain injury in this animal model. The results indicate that activation of central alpha2-adrenergic or imidazole receptors provides neuroprotection during reperfusion after hypoxic-ischaemic brain injury in neonatal rats. PMID- 11277362 TI - What's new in neuromuscular disorders? Nuclear envelope and Emery-Dreifuss muscular dystrophy. PMID- 11277363 TI - The long-term use of vigabatrin and lamotrigine in patients with severe childhood onset epilepsy. AB - Our purpose was to compare the initial efficacy and retention at 5 years of lamotrigine (LTG) and vigabatrin (VGB) in patients with severe childhood-onset epilepsies, especially with regard to loss of efficacy. Add-on therapy in 95 young patients with severe epilepsies in a neuropaediatric department was prospectively followed in open label studies for up to 5 years. VGB group: 56 patients (mean age: 11.1 years). LTG group: 39 patients (mean age: 13.6 years). Definition of initial responders: more than 50% reduction in seizure-frequency after 6 weeks of VGB treatment or after 4 months of LTG treatment. Definition of retention at 5 years: percentage of patients still taking LTG or VGB after 5 years. Definition of loss of efficacy: return to the baseline seizure frequency. The results were: VGB group: after 6 weeks 18 of 56 patients (32%) were initial responders. The retention at 5 years was five of 56 patients (8.9%). One patient (1.8%) was still seizure free. A loss of efficacy occurred in 10 of the 18 initial responders, usually within the first 9 months after the initial response. In the LTG group, after 4 months, 11 of 39 patients (28%) were initial responders. The retention at 5 years was 10 of 39 patients (25.6%). Five patients (12.8%) were still seizure free. The rate of adverse events was equal in both groups (41%). All but one occurred within the first 6 months of treatment. Our study in patients with difficult to treat childhood epilepsies suggests that in clinical practice patients were less likely to be discontinued from LTG than from VGB within 5 years, after similiar initial efficacy. This was mainly due to a loss of efficacy in VGB treatment early after initial response. PMID- 11277364 TI - Primary neonatal thalamic haemorrhage and epilepsy with continuous spike-wave during sleep: a longitudinal follow-up of a possible significant relation. AB - Epilepsy with continuous spike-waves during sleep was diagnosed in a child who suffered primary neonatal thalamic haemorrhage, and who was followed from birth to 17 years of age. Early cognitive development was normal. Acquired behavioural problems and cognitive stagnation could be directly related to the epilepsy and not to the initial lesion and posthaemorrhagic hydrocephalus. This case and long term follow-up data on a few children who suffered primary neonatal thalamic haemorrhage suggest that epilepsy with continuous spike-waves during sleep can be a sequel. Disturbances of thalamocortical interactions could play a role in the still poorly understood syndrome of epilepsy with continuous spike-waves during sleep. PMID- 11277365 TI - Why and how to assess the aetiological diagnosis of children with intellectual disability/mental retardation and other neurodevelopmental disorders: description of the Finnish approach. AB - One of the most important tasks of a physician who has patients with delayed development is to assess the cause of the problem. To help with this work, an aetiological classification based on timing and type of the injury to the central nervous system (CNS), has been created. The main divisions are: genetic causes; CNS malformations and multiple malformation syndromes of unknown origin; external prenatal factors; paranatally acquired disorders (-1 to +4 weeks from delivery); postnatally acquired disorders; and untraceable or unclassified causes. In the classification, the earliest factor injuring the CNS is the primary diagnosis. The first stage, which consists of a quite simple workup, reveals the timing of the injury and allows the possibility for family counselling. The overview of the second stage assessment is also given. The 'aetiological tree' illustrates the classification method and serves as a reference and teaching aid. It can also be used for genetic counselling to demonstrate the situation. This method has been used for almost 20 years and has been proven to enhance diagnostic activity and family counselling. PMID- 11277366 TI - What's new in neuro-oncology. PMID- 11277367 TI - Increased urinary excretion of tubular enzymes and proteins in children with epilepsy. AB - Cross-sectional analysis of children undergoing treatment with anti-epileptic drugs has shown an increased urinary excretion of tubular enzymes and proteins. This has usually been interpreted as a consequence of subclinical renal-tubular damage or enzyme induction. We measured excretion of tubular enzymes and proteins in 29 children who suffered from epileptic seizures and in 27 control children. Investigations were undertaken at diagnosis before the start of treatment and 3-4 months later. At diagnosis we found a slightly, but statistically significant increased excretion of N-acetyl-beta-glucosaminidase and alpha1-microglobulin. There was no significant difference between patients with an idiopathic and symptomatic aetiology of seizures or between patients with different seizure types. At the second investigation, in children treated with carbamazepine or valproate, no further increase occurred. We conclude that the increased excretion of tubular enzymes and proteins in children with epilepsy is most probably not due to a side-effect of the anti-epileptic drugs, but to a physiological alteration associated with the epilepsy itself. While the cause is unknown, the influence of serotonin metabolism is discussed. PMID- 11277368 TI - Anti-epileptic drug treatment in children: hyperhomocysteinaemia, B-vitamins and the 677C-->T mutation of the methylenetetrahydrofolate reductase gene. AB - The aim of the study was to observe the influence of carbamazepine and valproic acid on plasma total homocysteine and B-vitamin status and the gene-drug interaction with the 677C-->T mutation of the methylenetetrahydrofolate reductase (MTHFR) gene. Plasma total homocysteine concentrations were determined in 136 epileptic children taking anti-epileptic drugs as monotherapy. Nutritional (folate, B12 and B6 vitamins) and genetic (MTHFR 677 C-->T) determinants of plasma homocysteine were studied in a random sample of 59 of the 136 epileptic children. Total homocysteine concentrations were significantly increased (p < 0.05) and folate and vitamin B6 levels were significantly decreased (p < 0.01) in the children taking anti-epileptic drugs compared with our reference ranges. In the carbamazepine-treated group, significantly positive correlation was found between duration of treatment and homocysteine concentration (p < 0.01). Homocysteine concentrations showed a significantly negative correlation with vitamin levels (folate: p = 0.002, and vitamin B12: p = 0.017) only in the carbamazepine treated group. In children treated with carbamazepine up to 3 years, total homocysteine concentration correlated negatively only with folate (p = 0.003), while in patients treated for more than 3 years, total homocysteine correlated negatively only with vitamin B12 values (p = 0.007). The lowering action of carbamazepine treatment on folate levels seems to be associated with hyperhomocysteinaemia, which seems to be related to the homozygous condition for the MTHFR 677C-->T mutation. Valproic acid treatment, although also associated with hyperhomocysteinaemia, only shows a lowering effect on vitamin B6 levels, which seems to be independent of the MTHFR genotype. PMID- 11277369 TI - Hashimoto's thyroiditis--a rare but treatable cause of encephalopathy in children. AB - Hashimoto's encephalopathy is a very rare complication of Hashimoto's thyroiditis. It is a progressive or relapsing encephalopathy associated with elevation of thyroid specific autoantibodies. Patients usually present when euthyroid and this diagnosis should be considered in any unexplained encephalopathy or progressive cognitive decline in the euthyroid patient. PMID- 11277370 TI - Focal upper limb neuropathy in a child. AB - A 9-year-old girl presented with a 4-week history of right upper limb weakness. Clinical assessment and neurophysiological studies suggested an atypical brachial plexitis. She re-presented 3 1/2 years later with progressive muscle weakness involving both legs and left arm and hand. There had been no interval improvement in her right upper limb. Clinical, neurophysiological and pathological findings were consistent with chronic inflammatory demyelinating polyneuropathy. She responded to a single course of intravenous immunoglobulin--and review more than 6 years after treatment confirms that she remains functionally normal. Focal upper limb neuropathy preceding a diffuse demyelinating process by several years has not been previously described in a child. Long-term follow-up of this patient allows us to comment on the natural history of her condition and the apparent long-term efficacy of intravenous immunoglobulin in this case. PMID- 11277371 TI - Central nervous system malformations: locations of known human mutations. PMID- 11277372 TI - Further evidence of neurological sequelae associated with interferon therapy in the paediatric population. PMID- 11277373 TI - Routine daily chest radiographs in ventilated, very low birth weight infants. AB - The aim of this study was to assess the frequency of new abnormalities on routine chest radiographs of ventilated, very low birth weight (VLBW) infants during the acute stage of their illness. Infants were identified who had had at least three daily routine chest radiographs. The appearance of their subsequent radiographs was compared to that obtained on the 1st day of ventilatory support and the timing of new abnormalities (malposition of the endotracheal or nasogastric tube, pulmonary interstitial emphysema, pleural effusion, pulmonary oedema, lobar collapse or consolidation) noted. A total of 100 radiographs were examined from 30 VLBW infants, median gestational age 27 weeks (range 23-32 weeks). New abnormalities were present on the radiographs of 24 infants and on 50% of the radiographs examined. The commonest abnormalities noted were pulmonary interstitial emphysema, collapse and consolidation. Conclusion. Routine daily chest radiographs in mechanically ventilated, very low birth weight infants during the acute stage of their respiratory illness can yield new information important in patient care, new abnormalities being demonstrated in 50% of radiographs examined. PMID- 11277374 TI - Steroids in full term infants with respiratory failure and pulmonary hypertension due to meconium aspiration syndrome. AB - Persistent pulmonary hypertension of the newborn (PPHN) due to meconium aspiration syndrome (MAS), has a high morbidity and mortality especially in centres with limited access to extra-corporeal membrane oxygenation or nitric oxide therapy. In such a setting, we conducted a pilot study to evaluate the effect of dexamethasone on severe respiratory failure with PPHN due to MAS with a view to exploring possible justification for randomised controlled trials in similar patients. We prospectively managed a consecutive case series of 14 infants over a 3-year period with the above mentioned diagnosis, who were ventilated and with an oxygenation index >25. Dexamethasone was commenced in a dose of 0.5 mg/kg per day and given for up to a maximum of 9 days in a reducing schedule. Differences between time points were analysed using analysis of variance for repeated measures. The mean age of commencing dexamethasone was 79.9 h. There was a rapid, significant improvement (P < 0.05) in the respiratory status in 13 of these infants after commencing dexamethasone, allowing weaning from the ventilator and eventual extubation at a mean age of 8.7 days. One infant died. Two infants had infective episodes. Conclusion. Dexamethasone, if started early in infants with respiratory failure and persistent pulmonary hypertension of the newborn due to meconium aspiration syndrome may be effective in improving gas exchange, and possibly avoiding extra-corporeal membrane oxygenation. A randomised controlled trial of using dexamethasone early in similar patients and setting is warranted. PMID- 11277375 TI - Short stature as the only presenting feature in a patient with an isodicentric (Y)(q11.23) and gonadoblastoma. A clinical and molecular cytogenetic study. AB - A 13-year-old phenotypically female patient presented with short stature (height SDS -2.6), but without any Turner stigmata or other dysmorphic features. Chromosome analysis showed mosaicism for an isodicentric (idic) (Y)(q11.23) containing cell line and a 45,X cell line. Subsequent gonadectomy revealed a left streak ovary and a right ovary of abnormal appearance, which on histological examination appeared to contain a gonadoblastoma. DNA analysis showed that the proposed critical region of the gonadoblastoma locus on the Y chromosome was contained within the patient's idic (Y). Conclusion. The case described here shows that patients with 45,X/46,X, isodicentric (Yp) mosaicism and a female phenotype (1) can lack external virilisation but still have a gonadoblastoma and (2) do not necessarily have Turner stigmata but can present with only short stature. This case also underlines the importance of karyotyping patients with unexplained short stature to enable gonadectomy if Y-derived material is detected. PMID- 11277376 TI - Cerebral oedema in enuretic children during low-dose desmopressin treatment: a preventable complication. AB - Seven cases of cerebral oedema have been observed in enuretic children during low dose desmopressin (DDAVP) treatment given in a dose of 7-21 microg daily in the Czech Republic between 1995 and 1999, after the drug started to be marketed for this indication and delivered in simple bottles with a dropper. All seven children (age 5-11 years, four boys) experienced a period of unconsciousness but all recovered without sequelae. In most cases, safety measures were underestimated and natraemia was not regularly controlled. Two children developed cerebral oedema after excessive water intake in preparation for uroflowmetry, another one drank much during a hot summer day, in one diabetes insipidus was not recognised and two children were clearly non-compliant with reduced fluid intake on a long-term basis. Only in one child, no risk factor was found. Conclusion. Proper selection and instruction of patients is needed to avert cerebral oedema during treatment with desmopressin for nocturnal enuresis. PMID- 11277377 TI - Canalisation in human growth: a widely accepted concept reconsidered. AB - According to the concept of canalisation, infants and children stay within one or two growth channels, and therefore, any crossing of height centiles always warrants further evaluation. In view of evidence against this concept we re investigated the variability of individual growth in the First Zurich Longitudinal Growth Study. The investigation is based on height measurements of 232 children (112 females, 120 males) measured at annual intervals during childhood and half-yearly during adolescence. Height data were transformed into height standard deviation scores (SDS) and canalisation defined by the width of an individual's growth channel, i.e., by the differences between maximum and minimum height SDS, in the individual series of measurements. Many subjects of the First Zurich Longitudinal Growth Study crossed numerous centiles with patterns that often seemed to show characteristic features. For approximately two thirds of the subjects, the SDS channel during the whole growth process covers more than one SDS. In childhood, between the age of two and age of minimal height velocity, only about one fourth of the subjects have an SDS channel below 0.5, indicating acceptable canalisation. During childhood, growth in boys appeared slightly more canalised than in girls (P = 0.02). Conclusion. The present investigation does not support the concept of strict canalisation of individual growth. We suggest to consider crossing of centiles a normal event in child development, though in a clinical setting crossing centiles should still be taken seriously, at least at first until a medical cause for this has been excluded. PMID- 11277378 TI - Adenocarcinoma of the colon developing on the basis of Crohn's disease in childhood. AB - Colorectal carcinoma rarely affects children and has a dismal prognosis with 5 year survival rates as low as 2.5%-7% despite apparently radical surgery. Here we report the case of an adenocarcinoma of the sigmoid colon in a 15-year-old girl preceded by uncertain abdominal complaints of 5 years' duration. Pathological work-up revealed a tumour with lymph node metastases (pT3NI). Immunohistochemical evidence of p53 overexpression by the tumour cells raised the suspicion of an underlying Li-Fraumeni syndrome. In addition, there were aphthoid ulceration, fissuration of the non-tumorous mucosa, along with a mixed transmural infiltrate composed of macrophages, eosinophils, and non-typical giant cells, which were compatible with simultaneous Crohn's disease. Anamnestic data concerning the occurrence of idiopathic inflammatory bowel disease or colorectal carcinoma in the patient's relatives were non-contributory. The present results suggest a possible relationship between Crohn's disease and colon cancer due to the defective p53 gene product. PMID- 11277379 TI - Simultaneous occurrence of the haemolytic uraemic syndrome and acute post infectious glomerulonephritis. AB - We report on two children, a 12-year-old boy and a 6-year-old girl, with simultaneous occurrence of clinical and laboratory features consistent with both diarrhoea-negative haemolytic uraemic syndrome (D-HUS) and acute post-infectious glomerulonephritis (APGN). Both presented with acute renal insufficiency, hypertension and oedema. Laboratory evaluation revealed micro-angiopathic anaemia with burr cells, thrombocytopenia, elevated lactic dehydrogenase and low complement C3. Urinalysis showed marked proteinuria and haematuria. Renal biopsy was characteristic of APGN, but not of HUS. The outcome was good in both children. Conclusion. The simultaneous occurrence of diarrhoea-negative haemolytic uraemic syndrome and acute post-infectious glomerulonephritis is rare. The outcome is generally good as is expected in the latter condition in contrast to the former. PMID- 11277380 TI - Ascorbate acid concentration in airways lining fluid from infants who develop chronic lung disease of prematurity. AB - Chronic lung disease of prematurity (CLD) remains a common cause of morbidity and mortality in preterm infants. Oxygen toxicity remains a major risk factor for the development of CLD and as a consequence the antioxidant status of CLD babies is a major focus of interest. In the present study, we determined whether ascorbate, urate, and total glutathione concentrations were decreased in infants who developed CLD when compared to those who did not. From 34 preterm infants, 141 serial bronchoalveolar lavage fluid (BALF) and plasma samples were collected: 12 developed CLD (median gestation 26 weeks, range 23-28 weeks, median birth weight 780 g, range 630-1070 g), 16 developed and recovered from respiratory distress syndrome (RDS) (median gestation 31 weeks, range 26-39 weeks, median birth weight 1820 g, range 840-4160 g), and six were ventilated for non-respiratory reasons, (median gestation 35 weeks, range 32-38 weeks, median birth weight 2180 g, range 1100-2860 g). Following birth, the concentration of BALF ascorbate, urate and glutathione decreased over the 1st week in all three groups. Thereafter, BALF ascorbate increased in RDS and control infants during the 2nd week but this increase was delayed by 2 weeks in the CLD infants. No differences were noted between the RDS and CLD groups for urate and total glutathione in BALF or urate in plasma. BALF protein concentration was similar in all three groups except for a rise at day 7 in the CLD group but this did not reach statistical significance. Conclusion. A delayed increase in bronchoalveolar lavage fluid ascorbate concentration might be associated with an increased risk of developing chronic lung disease of prematurity. PMID- 11277381 TI - Subnormal response of plasma glucose concentration to glucagon despite adequate glycogenolysis: the importance of kinetic measurements. AB - The plasma glucose concentration response to a glucagon bolus is considered an important diagnostic tool in hypoglycemia of unknown origin. The response of plasma glucose concentration to glucagon can however also be misleading in the differential diagnosis. In a 3-week-old male infant suffering recurrent severe preprandial hypoglycemia and dependent on continuous i.v. glucose infusion, extensive diagnostic screening including a liver biopsy did not lead to a diagnosis. Based on an insufficient glycemic response (twice) to a glucagon bolus, a disorder of glycogenolysis was suspected. Glucose production and gluconeogenesis were measured (glycogenolysis calculated) during diminishing i.v. glucose infusion and after a glucagon bolus. Reducing glucose infusion resulted in a steep increase in glycogenolysis and gluconeogenesis, maintaining total glucose turnover (production plus infusion) constant at +/-9 mg x kg(-1) x min( 1) (+/-60% gluconeogenesis, +/-40% glycogenolysis). Plasma glucose concentration however decreased from 4.9 mmol/l to 3.4 mmol/l. Glucagon increased glucose production by 50% but resulted in only a minor increase in glucose concentration. Conclusion. As glucose concentration depends on the balance between glucose production and utilization (uptake), facilitated glucose uptake rather than impaired glycogenolysis explains the hypoglycemic episodes in this patient. A subnormal response of plasma glucose to glucagon therefore does not necessarily imply a disturbance in glycogenolysis. In cases of hypoglycemia of unknown origin, measurement of glucose kinetics with stable isotopes is indicated. PMID- 11277382 TI - Idiopathic palmoplantar eccrine hidradenitis in children. AB - Idiopathic palmoplantar eccrine hidradenitis (IPPH) is a recently described disorder characterized by painful erythematous plantar nodules and in three cases, showed a typical neutrophilic infiltrate around and within the eccrine sweat apparatus. Five cases of IPPH on the soles of the feet in healthy children are reported. The disorder presented after intense physical activity in four cases. The course was benign and self-limiting. Complete bed rest for several days without any medical therapy led to alleviation of the pain and disappearance of all the lesions. Conclusion. Idiopathic palmoplantar eccrine hidradenitis may be more common than reported. Paediatricians should be aware of it in order to avoid unnecessary diagnostic tests and treatments. PMID- 11277383 TI - Heparinised whole blood or citrated blood for exchange transfusion. PMID- 11277384 TI - Hyperbilirubinaemia, glucose-6-phosphate dehydrogenase deficiency and Gilbert syndrome. PMID- 11277385 TI - Intra-abdominal hemorrhage due to a ruptured corpus luteum cyst in a girl with congenital afibrinogenemia. PMID- 11277388 TI - The in-vitro bleeding time for the screening of platelet function in the newborn: assessment of a normal reference range. PMID- 11277387 TI - Compound heterozygosity for factor V Leiden and prothrombin G20210A mutations in a child with Budd-Chiari syndrome. PMID- 11277386 TI - Desmopressin in a long-term treatment of children with primary nocturnal enuresis -a symptomatic therapy? PMID- 11277389 TI - Neuralgic amyotrophy associated with parvovirus B19 infection in a child. PMID- 11277390 TI - In situ hybridization for the identification of yeastlike organisms in tissue section. AB - The identification of yeast and yeastlike organisms in tissue sections can be very difficult. Biopsy tissues may be limited, with only occasional organisms present. In addition, several common species have overlapping histologic features. Deoxyribonucleic acid probes were designed to detect both the 18S and 28S ribosomal ribonucleic acid sequences of five fungal organisms with a high degree of specificity for each fungus. Each of these organisms--Blastomyces dermatitidis, Coccidioides immitis, Cryptococcus neoformans, Histoplasma capsulatum, and Sporothrix schenckii--can be manifested histologically as round, yeastlike structures, often within a similar size range. Probes were tested against 98 archived, formalin-fixed, paraffin-embedded tissue specimens, each of which had culture-proved involvement by one of these organisms. Assessment of accuracy was based on the presence of yeastlike organisms in consecutive Grocott's methanemine silver (GMS)-stained tissue sections, and agreement with culture results. The results indicated that GMS had a greater overall sensitivity in detecting fungal organisms (95.9%) compared with in situ hybridization (ISH; 82.7%). ISH with oligonucleotide deoxyribonucleic acid probes, however, was more specific, with all species-specific probes yielding 100% specificity (compared with 96.2-100% specificity based on morphology alone). ISH also had a higher positive predictive value (100% in all cases) compared with GMS (83.3-100%). In addition, four cases with rare organisms present (4.1% of cases tested) were detected by ISH but not by GMS staining. These results show that ISH, directed against ribosomal ribonucleic acid, provides a rapid, accurate technique for the identification of yeastlike organisms in histologic tissue sections. Its primary strength lies in the ability to speciate organisms accurately that are too few or atypical to identify based solely on morphologic features. PMID- 11277391 TI - Translocation (10;11;22)(p14;q24;q12) characterized by fluorescence in situ hybridization in a case of Ewing's tumor. AB - It is well recognized that the identification by classic cytogenetics of t(11;22)(q24;q12) is a useful aid in the accurate diagnosis of Ewing's sarcoma and related tumors. This translocation induces the EWS/FLI-1 fusion transcript, which can be detected by reverse transcription-polymerase chain reaction. Recent studies have also used fluorescence in situ hybridization (FISH) to demonstrate the translocation. The authors coupled classic cytogenetics and FISH on tumor cells from the original specimen, the local recurrence, and the pulmonary metastasis as well as from the xenografted tumors in a case of extraosseous Ewing's sarcoma. FISH analysis not only confirmed the cytogenetic results but also allowed the identification of a tumor-specific chromosome change, consistent with a complex translocation, t(10;11;22), as well as revealed other chromosomal rearrangements on both metaphases and interphase nuclei of each material. In addition this technique served to identify, in the interphase nuclei of the original tumor, the clone that became dominant, from the cytogenetic point of view, in the lung metastasis and in the nude mice xenografted tumors. Current results indicate that the use of FISH on metaphases and interphase nuclei is an easy and reliable approach to complement or even to substitute classic cytogenetic studies for the detection of specific chromosomal rearrangements, especially for determining complex translocations and for describing tumoral clones with different cytogenetic markers. PMID- 11277392 TI - PCR techniques for clonality assays. AB - Clonal overgrowths represent the hallmark of neoplastic proliferations, and their demonstration has been proved useful clinically for the diagnosis of malignant lymphomas based on the detection of specific and dominant immunoglobulin and/or T cell receptor gene rearrangements. Nonrandom genetic alterations can also be used to test clonal expansions and the clonal evolution of neoplasms, especially analyzing hypervariable deoxyribonucleic acid (DNA) regions from patients heterozygous for a given marker. These tests rely basically on the demonstration of loss of heterozygosity (LOH) resulting from either hemizygosity (nonrandom interstitial DNA deletions) or homozygosity of mutant alleles observed in neoplasms. LOH analyses identify clonal expansions of a tumor cell population, and point to monoclonal proliferation when multiple and consistent LOH are demonstrated. Based on the methylation-related inactivation of one X chromosome in female subjects, X-linked markers (e.g., androgen receptor gene) will provide clonality information using LOH analyses after DNA digestion with methylation sensitive restriction endonucleases. Therefore, both non-X-linked and X-linked analyses give complementary information, related and not related to the malignant transformation pathway respectively. Applied appropriately, these tools can establish the clonal evolution of tumor cell populations (tumor heterogeneity), identify early relapses, distinguish recurrent tumors from other metachronic neoplasms, and differentiate field transformation from metastatic tumor growths in synchronic and histologically identical neoplasms. PMID- 11277393 TI - An effective strategy of using molecular testing to screen mentally retarded individuals for fragile X syndrome. AB - Fragile X syndrome (FXS) is the most common form of familial mental retardation (MR). It is caused by the expansion of the CGG repeat in the FMR1 gene on the X chromosome. To date, FXS is not treatable, but can be prevented by prenatal genetic examination. Identifying women who carry a full mutation or premutation FMR1 gene is thus very important, and can be done by tracing family members of FXS subjects. However, most of the FXS subjects in Taiwan as well as those in many other countries have not been identified. In this study the authors attempt to develop reliable and inexpensive tests suitable for a large-scale screen of subjects with MR for FXS. Together with their previous study, a total of 311 male and 160 female subjects with MR were screened with nonradioactive Southern blot assay using mixed deoxyribonucleic acid from three subjects of the same sex. From these subjects, nine male subjects and one female FXS subject were diagnosed. All male subjects were also screened with nonradioactive polymerase chain reaction (PCR). These nine male FXS subjects were also detected on the basis of PCR amplification failure. No false-negative results were discerned. The PCR procedure was simplified further by combining it with an analysis of a blood spot on filter paper, which is a much simpler and cheaper method for sample collection and DNA preparation. This method was then used to screen 104 boys with MR. Two of them were suspected, and later confirmed with Southern blot assay, as subjects with FXS. This study suggests that simple PCR combined with blood spot analysis could be a reliable, inexpensive test that is feasible for a large-scale screening of male subjects with MR for FXS. However, Southern blot assay with mixed deoxyribonucleic acid is appropriate for screening female subjects. Based on this strategy, most FXS subjects could be identified easily for further management. PMID- 11277394 TI - A novel polymorphism in the PTC gene allows easy identification of allelic loss in basal cell nevus syndrome lesions. AB - Basal cell nevus syndrome (BCNS; also nevoid basal cell carcinoma syndrome [NBCCS]; Gorlin's syndrome) is an autosomal dominant syndrome characterized by multiple basal cell carcinomas, keratocysts, and developmental skeletal defects. Mutation of the human homologue of Drosophila patched (PTC) gene is considered to be the molecular defect in BCNS. PTC mutations have been observed in sporadic tumors including basal cell and ovarian carcinomas and medulloblastoma. The authors report a novel C/T polymorphism in the PTC gene. Forty-eight normal blood samples were screened for the presence of the polymorphism using direct radioactive and automated sequencing of polymerase chain reaction (PCR) products and restriction enzyme digestion. Results demonstrated 20 homozygous T (43%), 11 homozygous C (23%), and 17 heterozygous C/T (35%). The presence of this polymorphism has permitted us to directly detect allelic loss in BCNS, sporadic keratocysts, and basal cell carcinoma (BCC). Further, four BCNS keratocysts and two BCNS-BCC and three non-BCNS keratocysts showed allelic loss of complementary DNA from lesions when compared with their corresponding blood genomic DNA. PMID- 11277395 TI - Physical state of HPV16 and chromosomal mapping of the integrated form in cervical carcinomas. AB - Using a procedure based on restriction enzyme cleavage, self-ligation, and inverse polymerase chain reaction (rliPCR), the authors investigated 18 cervical intraepithelial neoplasia III (CIN III) cases and 37 invasive squamous carcinomas for integration of human papillomavirus type 16 (HPV16). All eighteen CIN III cases (severe dysplasia or high-grade squamous intraepithelial lesion) were found to harbor episomal HPV, but one of the samples contained mixed episomal and integrated forms. Seventeen of 37 invasive cervical carcinoma samples were identified previously as containing the completely integrated HPV16 genome by using PCR covering the entire E1/E2 gene, and this was confirmed by rliPCR in 16 cases. One case, however, showed a low level of episomal deoxyribonucleic acid in addition to the predominant integrated form. Of the remaining 20 carcinoma samples showing episomal forms in the previous analysis, 14 were found to contain integrated forms using rliPCR, and four contained multimeric episomal forms. Thus, in total, 31 of 37 of the carcinomas (84%) showed the integrated HPV16 genome. The rliPCR product from five carcinoma cases was cloned into a plasmid vector and used as a template for "primer walking" deoxyribonucleic acid sequencing to deduce human sequences flanking the integrated HPV genome. Based on this information, bacterial artificial chromosome (BAC) and P1-derived artificial chromosome (PAC) clones were obtained and used as probes in fluorescent in situ hybridization experiments on human metaphase chromosomes. The results of the fluorescent in situ hybridization experiments showed evidence for HPV16 integration in chromosome regions 1q25, 3q28, 6p25, 11p13, and 18q22. Sixteen carcinoma samples, containing episomal HPV16, were sequenced in the long control region. Evidence for changes in E2 binding or silencer YY1 sequences was found in only two samples. PMID- 11277396 TI - Poor storage and handling of tissue mimics mitochondrial DNA depletion. AB - Analysis of the mitochondrial DNA (mtDNA) is an important part in the diagnosis of mitochondrial disorders. Besides point mutations and deletions in the mitochondrial genome a reduction in the amount of mtDNA molecules (mtDNA depletion) can also be the reason for mitochondrial defects. The DNA stability in clinical samples is essential for proper performance and interpretation of DNA based diagnosis. The stability of mtDNA was compared with that of nuclear DNA under poor handling and storage conditions. Fresh and thawed muscle tissue specimens were kept at different temperatures for a certain period of time before DNA isolation. Quantitative Southern blot analysis revealed a time-dependent decrease in the amount of mtDNA compared with nuclear DNA in thawed tissue specimens. Therefore, the current study demonstrates that proper specimen storage is a critical issue in quantitative mtDNA analysis and that poor handling and storage of tissue may mimic a severe mtDNA depletion. PMID- 11277397 TI - Gene expression analysis of the catalytic subunit of human telomerase (hEST2) in the differential diagnosis of serous effusions. AB - Diagnostic accuracy in effusion cytology based on morphologic examination is not always satisfactory. Therefore, various diagnostic adjuncts such as immunocytochemistry or deoxyribonucleic acid cytometry are employed in this diagnostic field. Recently, demonstration of telomerase activity has been proposed as a possible marker for malignancy. In this study a seminested reverse transcription-polymerase chain reaction (RT-PCR) strategy for expression analysis of the catalytic subunit of human telomerase (hEST2) was used in 58 serous effusions. RT-PCR results correlated with cytologic diagnoses in 14 of 17 malignant effusions. In eight effusions cytologically suspicious for malignancy, PCR results were in accordance with the clinical follow-up. However, hEST2 RT-PCR was also positive in six of 15 cytologically benign effusions that consisted predominantly of inflammatory and mesothelial cells. Using the telomeric repeat amplification protocol, it could be demonstrated that cultured, proliferating benign mesothelial cells may present a weak telomerase activity, as is known in other benign cells including activated lymphocytes. In conclusion, the simple and rapid method of hEST2 RT-PCR serves to support the cytologic diagnosis of malignancy, but false-positive PCR results resulting from activated lymphocytes and proliferating mesothelial cells must be considered. PMID- 11277398 TI - Re: Molecular detection of M. tuberculosis DNA in tuberculosis and sarcoidosis. PMID- 11277399 TI - A microsatellite fluorescent method for linkage analysis in familial retinoblastoma and deletion detection at the RB1 locus in retinoblastoma and osteosarcoma. AB - Linkage analysis at the retinoblastoma locus (RB1) is essential for identifying individuals at risk and to offer adequate genetic counseling in familial retinoblastoma. It can also be used to detect large deletions involving RB1, which accounts for 15% of the genetic alterations in hereditary retinoblastoma. These studies are usually carried out with lengthy Southern blot analyses of relatively uninformative restriction fragment length polymorphisms. The authors report an alternative, reliable protocol for genotyping the RB1 locus using two pairs of highly informative intragenic and flanking microsatellites linked closely to the RB1 gene, and analysis of the fluorescent-labeled polymerase chain reaction products with automatic sizing technology. This methodology has successfully identified high risk carriers in five of the five pedigrees of familial retinoblastoma studied. In addition, gross deletions affecting the RB1 gene were identified in two of 12 sporadic bilateral retinoblastomas, and loss of heterozygosity at the RB1 locus has been detected in one of three osteosarcomas using the same experimental protocol. The described protocol is simpler and faster than conventional Southern blot methodologies and can identify a larger number of informative cases. PMID- 11277400 TI - Gene structure and larval expression of cnox-2Am from the coral Acropora millepora. AB - We have cloned a Hox-like gene, cnox-2Am, from a staghorn coral, Acropora millepora, an anthozoan cnidarian, and characterised its embryonic and larval expression. cnox-2Am and its orthologs in other cnidarians and Trichoplax most closely resemble the Gsx and, to a lesser extent, Hox 3/4 proteins. Developmental northern blots and in situ hybridisation are consistent in showing that cnox-2Am message appears in the planula larva shortly after the oral/aboral axis is formed following gastrulation. Expression is localised in scattered ectodermal cells with a restricted distribution along the oral/aboral body axis. They are most abundant along the sides of the cylindrical larva, rare in the oral region and absent from the aboral region. These cells, which on morphological grounds we believe to be neurons, are of two types; one tri-or multipolar near the basement membrane and a second extending projections in both directions from a mid ectodermal nucleus. Anti-RFamide staining reveals neurons with a similar morphology to the cnox-2Am-expressing cells. However, RFamide-expressing neurons are more abundant, especially at the aboral end of the planula, where there is no cnox-2Am expression. The pattern of expression of cnox-2Am resembles that of Gsx orthologs in Drosophila and vertebrates in being expressed in a spatially restricted portion of the nervous system. PMID- 11277401 TI - Epimorphic regeneration of the distal part of the planarian pharynx. AB - The totipotent stem cells called neoblasts seem to be concerned with the remarkable regeneration ability of planarians. However, the pharynx is able to regenerate after the amputation of its distal part, in spite of a lack of neoblasts in the pharynx. The process of regeneration has been referred to as morphallaxis, based on conventional histochemical observations. We examined it again immuno-histochemically using anti-Dugesia japonica proliferating cell nuclear antigen (DjPCNA) antibody for neoblasts and anti-D. japonica myosin heavy chain-A (DjMHC-A) antibody for pharynx muscle fibers. This immuno-histochemical study, together with observations of the regeneration process of planarians irradiated with X-rays in particular regions, revealed that after the amputation, neoblasts from outside the pharynx entered that organ, moved through the mesenchyme of the pharynx to the wounded area, and differentiated into the cells that had been lost there. We show here that the regeneration after amputation of the distal part of the pharynx is an 'epimorphic' process. PMID- 11277403 TI - Pathfinding analysis in a glia-less gcm mutant in Drosophila. AB - Many lines of evidence suggest that glial cells function as guide post cells for axonal pathfinding. However, due to the difficulty in completely eliminating glial cells during development, their functions in axonal pathfinding have not been critically evaluated. In Drosophila gcm mutant embryos, glial cells were genetically eliminated providing us with a unique opportunity to investigate glial functions in nervous system formation. We showed that even in the absence of glial cells the initial axonal extension of pioneer neurons was essentially normal. However, at later stages, axon bundle formation and pathfinding were disturbed in the absence of glial cells, and abnormal migration of glial cells led to misrouting of axons. This indicates that glial cells are required for correct pathfinding at later stages. We propose that glial cells function in a stage-specific manner; they are not required for the initial extension of pioneers but essential for the subsequent extension of pioneers and followers as well as axon bundle formation. PMID- 11277402 TI - Multiple function of poxn gene in larval PNS development and in adult appendage formation of Drosophila. AB - The gene pox-neuro (poxn), which encodes a transcriptional regulator including a paired domain, specifies the differences between mono-innervated external sensory (m-es) organs and poly-innervated external sensory (p-es) organs in Drosophila. Here, we analyse the function of poxn in the development of the larval peripheral nervous system (PNS) and in other developmental aspects using a loss-of-function mutant of poxn. We observed that, in addition to the transformation of p-es into m-es organs in the mutant embryo, the external structure of the trichome-like sensilla (hairs) misdifferentiates into that of the campaniform-like sensilla (papillae) in the second and third larval instars. We also observed that POXN is expressed in a cell associated with the external structure of the trichome-like sensilla in the first and second instar larvae. These results imply that poxn is required in two distinct steps in the development of the larval PNS: (1) development of the larval p-es organs during embryogenesis and (2) re-formation of larval sensory hairs after each larval moult. In addition to its expression in the developing PNS, POXN is also expressed in concentric domains of the leg and antennal imaginal discs of early third instar larvae, and in the region of the wing disc that will form the wing hinge. The loss of poxn function results in defects of segmentation of the tarsus and antenna and in a distortion in the wing hinge. These results indicate that the poxn gene plays crucial roles in the morphogenesis of the appendages, in addition to its role in the early specification of neuronal identity. PMID- 11277404 TI - Neurogenesis in the developing visual system of the branchiopod crustacean Triops longicaudatus (LeConte, 1846): corresponding patterns of compound-eye formation in Crustacea and Insecta? AB - In the discussion on arthropod phylogeny, the structural evolution of compound eyes and optic ganglia in Crustacea and Insecta is an important topic. On the one hand, many morphological features as well as developmental aspects of the visual system in Insecta and Crustacea correspond in so much detail that eye design in these two groups is likely to have a common euarthropodan ancestor. On the other hand, however, some authors advocate a convergent evolution of the crustacean and insect visual system founding their arguments on differences in the arrangement of the visual neuropils and the fibre connections between Malacostraca and Entomostraca (the "entomostracan enigma"). Therefore, information about cellular aspects of visual system formation in entomostracan Crustacea is likely to enliven this debate, but is not yet available. To fill this gap, we examined the proliferation of neuronal stem cells in the developing visual system of the tadpole shrimp Triops longicaudatus (LeConte, 1846) (Entomostraca, Branchiopoda, Phyllopoda, Calmanostraca, Notostraca) by in vivo incorporation of the proliferation marker bromodeoxyuridine and subsequent immunohistochemical detection. Our results indicate that in the developing visual system of T. longicaudatus, three band-shaped zones containing neuronal stem cells are present corresponding to the proliferation zones found in Malacostraca. We therefore conclude that the ontogenetic mechanisms of visual-system formation are evolutionarily conserved (homologous) in Branchiopoda, Malacostraca, and Insecta. PMID- 11277405 TI - Maelstrom is required to position the MTOC in stage 2-6 Drosophila oocytes. AB - The factors that determine intracellular polarity are largely unknown. In Drosophila oocytes one of the earliest polar events is the positioning of the microtubule-organizing center (MTOC). Here we present data that are consistent with the hypothesis that maelstrom is required for posterior positioning of the MTOC. PMID- 11277406 TI - Isolation of an early neural maker gene abundantly expressed in the nervous system of the ascidian, Halocynthia roretzi. AB - Ascidian tadpole larvae possess a primitive nervous system, which is a prospective prototype of the chordate nervous system. It is composed of relatively few cells but sufficient for complex larval behavior. Here we report on HrETR-1, a gene zygotically expressed in a large proportion of the developing neural cells of the ascidian, Halocynthia roretzi. HrETR-1 is an early neural marker which can be used for analyzing neural differentiation. HrETR-1 expression intensified in most neural cells of genes isolated to date, in both central and peripheral nervous systems including palps as early as the 110-cell stage. Using this gene as a probe, we characterized neural cells in the nervous system as well as confirming their origins. Also, we recognized three types of peripheral epidermal neurons which presumably correlate to the larval neurons previously reported for another ascidian. Among these, five bilateral neurons located in the anterior region of the trunk appeared to be derived from a8.26 blastomeres. PMID- 11277407 TI - Immunohistochemistry for the age of molecular morphology. PMID- 11277408 TI - Comparison of cyclin A and MIB-1 expression in astrocytic tumors using image based cell analysis system. AB - Traditional prognostic indicators for astrocytic tumors include tumor size, type, and histologic grade. Data suggest that tumor growth fraction assessed by MIB-1 is an important predicator of survival. Cyclin A, like MIB-1, is a recently described specific marker of proliferation, detectable primarily in S phase of the cell cycle as it undergoes progression to G2 phase. Thirty-seven cases of astrocytic tumors--14 cases of World Health Organization grade 1 and 2 (low grade tumors), 8 cases of grade 3 (anaplastic astrocytoma), and 15 cases of grade 4 (glioblastoma multiforme)--were simultaneously evaluated using routine paraffin immunohistochemical methods with commercial antibodies against MIB-1 and cyclin A. The results were quantitated using a Cell Analysis System (CAS) 200 image analyzer. The mean percentage positive nuclear area for MIB-1 was 3.32% in grade 1 and 2, 19.27% in grade 3, and 24.00% in grade 4 astrocytic tumors. Cyclin A showed a similar pattern of positivity in the same cases: 2.84% in grade 1 and 2, 16.27% in grade 3, and 24.88% in grade 4 astrocytic tumors. The data suggest that both proliferation markers correlated significantly with histologic grade. Cyclin A appears to be as good an indicator of brain tumor proliferation as MIB-1. Because cyclin A is detectable primarily in the S phase of the cell cycle, the fraction of cells positive for cyclin A should allow for a more accurate indicator of tumor progression. PMID- 11277410 TI - Chromosomal abnormalities and p53 gene mutation in a cardiac angiosarcoma. AB - Angiosarcoma is the most common malignant neoplasm of the heart. However, to the authors' knowledge, no cytogenetic study of cardiac angiosarcoma has been reported. In the current study, an angiosarcoma from the right atrium of a 29 year-old man was investigated. Examination of tissue sections indicated that the tumor was a high grade epithelioid angiosarcoma of the heart. Cytogenetic analysis of tumor cells revealed a hyperdiploid clonal population with chromosomal numerical changes and one structural rearrangement, which was defined as: 55, XY, +der(1;17) (q10:q10), +2, +7, +8, +8, +19, +20, +21, +22. Multicolor fluorescent in situ hybridization on paraffin-embedded tissue sections illustrated polysomy of chromosome 8 in tumor cells. In addition, immunohistochemical analysis showed high expression of mutated p53 gene products in tumor cell nuclei. These findings demonstrate the involvement of chromosomal anomalies and gene mutation in cardiac angiosarcoma and suggest they play a role in neoplasia of the heart. PMID- 11277409 TI - Apoptosis, bcl-2 expression, and p53 expression in gastrointestinal stromal/smooth muscle tumors. AB - The authors investigated the relations between outcome and apoptosis, immunohistochemical demonstration of bcl-2 protein, and immunohistochemical staining for p53 protein in patients with gastrointestinal stromal/smooth muscle tumors (GIST). Patients whose tumors demonstrated cellular apoptosis using the terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate biotin nick end labeling (TUNEL) assay had an improved survival over those whose tumors did not improve. In contrast, patients whose tumors demonstrated staining for bcl-2 protein had a decreased survival compared with those whose tumors did not demonstrate bcl-2. There was no relation between p53 immunoreactivity and survival. These results suggest that inhibition of apoptosis may be associated with malignant behavior in patients with gastrointestinal stromal/smooth muscle tumors. PMID- 11277411 TI - Immunohistochemical and reverse transcription-polymerase chain reaction expression analysis of tyrosinase and microphthalmia-associated transcription factor in angiomyolipomas. AB - Angiomyolipomas (AMLs) show a characteristic immunoreactivity with melanocyte differentiation markers such as monoclonal antibody (mAb) HMB45, which detects melanocyte differentiation antigen gp100 and mAb A103 reacting with Melan-A/MART 1. Monoclonal antibody T311 to tyrosinase (a key enzyme of melanogenesis) and mAb D5 to the microphthalmia (Mitf) antigen are two newly available markers of melanocytic differentiation. The authors tested 15 AMLs with T311 and D5 by immunohistochemistry and a subset of 3 cases by reverse transcription-polymerase chain reaction for their expression of tyrosinase and Mitf mRNA. T311 showed poor sensitivity in AMLs because only focal staining was seen in 1 out of 15 cases, although tyrosinase mRNA was found in all tested cases. Mitf mRNA was present in 3 of 3 tested cases, and D5 was positive in 15 of 15 AMLs. However, D5 immunostaining often was focal and not as homogeneous as A103, which was analyzed in a previous study. D5 staining also could be seen in other cell types such as normal renal tubular cells, macrophages, and renal cell carcinoma. The current results show that in contrast with HMB45 and A103, T311 has little or no value in the diagnosis of AMLs. D5 may be useful in a panel of antibodies in the diagnosis of AMLs. PMID- 11277412 TI - Standardization in immunohistochemistry. AB - Over the last few years, the role of immunohistochemical analysis has been changing from that of an ancillary diagnostic technique to that of a stand-alone diagnostic method, the results of which determine treatment approach. With this change comes a need for increasing standardization of both preanalytical and analytical methods, so that results emanating from different laboratories are directly comparable. These efforts have been aided to some degree by the development of automated staining systems and by the increased use of test kits, but interlaboratory reproducibility for immunohistochemical methods is still far lower than that of most clinical analytical measurements. In this article, the author presents some approaches to further reduce intra- and interlaboratory variation in the performance of immunohistochemical assays, focusing attention on guidelines recently published by NCCLS. PMID- 11277413 TI - Oligodendroglioma: HMB-45 positivity using catalyzed signal amplification method: an immunohistochemical (HMB-45, CD31, p53, Mib-1) and ultrastructural study. AB - Although melanin synthesis and the presence of melanosomes are exceptionally reported in nervous system tumors, there is no record of melanotic oligodendrogliomas in the literature. The purpose of the current study was to evaluate whether melanosomes are immunohistochemically and ultrastructurally detectable in nonmelanotic oligodendrogliomas and to verify whether these data are related to prognosis. Thirty surgical specimens (19 primary lesions and 11 recurrences) from 19 patients were examined. Median survival was 80 months. Immunohistochemical studies were performed using the monoclonal antibodies HMB 45, CD31. Mib-1, and p53. Using catalyzed signal amplification (CSA), HMB-45 positivity was noticed in 3 (10%) of the oligodendrogliomas being studied. No correlation with survival was found. Ultrastructural examination displayed the presence of melanosomelike structures. Tumor vascularization, estimated by means of CD31 antibody, was increased in 6 of 19 primary lesions but there was no significant correlation with survival. Nine of the19 primary lesions were p53 negative. In these cases, survival was longer than in p53-positive tumors (P = 0.0213). Proliferation rate, evaluated with Mib-1, was unrelated to survival, but proved greater in recurrences (10 of 11 cases) than in primary tumors (7 of 19 lesions; P = 0.007). PMID- 11277414 TI - Apomucin expression and association with Lewis antigens during gastric development. AB - In normal stomach, MUC5AC and MUC6 apomucins are associated with Lewis types 1 and 2, respectively, and this association is lost during gastric carcinogenesis. The expression of gastric (MUC5AC, MUC6) and intestinal (MUC2, MUC4) apomucins and Lewis antigens during gastric development, using single and double labeling immunohistochemistry on fetal tissues (15-41 weeks), was analyzed and related to the tumor expression patterns. Apomucin expression in other fetal tissues was also analyzed. In gastric samples, MUC2 is detected in 14 of 19 showing no correlation with fetal age, and MUC4 is not detected. MUC5AC and MUC6 are always highly detected and are coexpressed and associated with both types of Lewis antigens. These patterns change progressively with the development of the adult gastric morphology. MUC2 is detected in the small intestine, colon, and pancreas; MUC4 is expressed in the colon; MUC5AC is detected in the small intestine; and MUC6 is found in the duodenum and pancreas. The patterns of apomucin expression and association with Lewis antigens during development are complex, but there is a trend toward the establishment of the adult pattern, with the exception of MUC4, which is not detected. These patterns found in fetal stomach indicate that alterations reported in gastric tumors do not fully recapitulate a developmental phenotype. PMID- 11277415 TI - Progesterone receptor immunohistochemical quantitation compared with cytosolic assay: correlation with prognosis in breast cancer. AB - Quantitation of estrogen and progesterone receptors (PR) represents the standard of care in the treatment of patients with breast cancer. Historically this was performed by cytosolic assay; current methods utilize immunohistochemical staining, which may be quantitated visually or by image cytometry. Formalin-fixed paraffin embedded sections from 95 breast carcinomas were immunostained with an avidin-biotin complex technique. steam antigen retrieval, and a monoclonal PR antibody (1/40 Biogenex). Nuclear immunostain was quantitated visually as the percentage of immunopositive nuclei, scored as 0 to 4. By image cytometry, the percentage of positively staining nuclear area (PPNA) was determined in 15 hpf using the CAS 200 Image Analyzer. Dextran-coated charcoal (DCC) ligand binding assay data were divided into negative (<10 fmol), low positive (10-50), or positive (>50). A statistically significant correlation was found between stage (P = 0.0001), the presence of nodal metastases (P = 0.0001), cytosolic assay (P = 0.036), image cytometry (P = 0.01), and disease-free survival. Only stage (P = 0.0001) and PR quantitation per cytosolic assay (P = 0.0001) correlated with overall survival. The method of choice for the assessment of PR hormone status in breast carcinomas is the DCC ligand binding assay. This method correlates with both survival and disease-free survival. Image cytometric quantitation of PR immunohistochemical staining correlates only with disease-free survival. The commonly used method of visual quantitation of PR immunostaining fails to relate either to survival or disease-free survival. PMID- 11277416 TI - Automated immunohistochemical staining of formalin-fixed and paraffin-embedded tissues using a catalyzed signal amplification method. AB - An immunohistochemical assay using catalyzed signal amplification (CSA), which is based on the peroxidase catalyzed deposition of biotinylated tyramide, is a highly sensitive method to visualize weak immunohistochemical signals originating from rare antigens or masked antigens in formalin-fixed, paraffin-embedded (FFPE) tissues. However, CSA methods are hampered by poor reproducibility and the complexity of their staining procedures. In this study, we aimed to apply the CSA procedure to a capillary gap-based, automated immunostainer, TechMate Horizon, to perform immunohistochemical signal amplification effectively and reproducibly. A variety of cellular antigens previously considered to be undetectable in FFPE human specimens were selected and examined with the automated immunostainer. Compared with the manual CSA staining method that takes more than 2 hours, the automated CSA method took less than 2 hours to complete. The staining results from the automated CSA method presented higher reproducibility, as well as lower background owing to well-regulated, punctual staining and washing at every step of the procedure. Conclusively, the automation of the CSA method enabled us to perform the time-consuming and complicated CSA amplification technique with minimal effort in an accurate, consistent, and reproducible manner. PMID- 11277417 TI - In situ hybridization detection of calcitonin mRNA in routinely fixed, paraffin embedded tissue sections: a comparison of different types of probes combined with tyramide signal amplification. AB - In situ hybridization (ISH) is a powerful molecular tool used to visualize nucleic acids in tissues and cells. However, its use is limited by the relative lack of sensitivity in detecting low copy numbers of nucleic acids. Several strategies have been developed to improve the threshold levels of in situ detection of nucleic acid by amplification of either target nucleic acid sequences before ISH (such as in situ polymerase chain reaction) or after the hybridization procedure (such as tyramide signal amplification [TSA]). The authors compared the use of different types of probes to detect calcitonin mRNA in 10 cases of formalin-fixed, paraffin-embedded medullary thyroid carcinoma with and without TSA. In addition, dot blot hybridization was used to compare the signal amplification by TSA with oligonucleotide. cDNA, and cRNA probes. These results show that cRNA probes are the most sensitive types of probes for detecting mRNA molecules in formalin-fixed, paraffin-embedded tissue sections and that tyramide amplification can increase the sensitivity for detection of calcitonin mRNA in formalin-fixed, paraffin-embedded tissue sections at least 2- to 4-fold with cRNA probes. PMID- 11277418 TI - Cytokeratin immunoreactivity in mouse tissues: study of different antibodies with a new detection system. AB - The cross-reactivity of a group of monoclonal antibodies (MABs) generated against human cytokeratins (CKs) was investigated in mouse tissues. Formalin-fixed and paraffin-embedded sections of lung, stomach, small and large intestine, liver, and kidney were immunostained with MABs after epitope retrieval with enzyme digestion. AE1/AE3, a "cocktail" of two MABs that recognizes basic and acidic CKs, 5D3 MAB to low molecular weight CKs (8, 18, and 19), and monospecific MABs to CK 7 and 20 were tested. Additionally, CK 17 and 34betaE12 MABs to high molecular weight CKs were evaluated in the same organs and in sections from skin and preputial glands. We employed the new universal animal system (ARK) as the detection system. The results showed intense reactivity for the first group of antibodies used, with topographic distribution similar to that in human tissues, with the exception of CK 7 in lung parenchyma, which displayed reactivity only in type II pneumocytes, with negativity of adjacent bronchial epithelium. Also of note was the lack of reaction of liver hepatocytes and renal tubular cells to AE1/AE3 and 5D3 MABs. Regarding the second group of antibodies, no reaction was obtained for CK 17 in the tissues tested. On the contrary, 34betaE12 MAB yielded intense reactivity in cells of epidermis and hair follicles. Compared to other detection systems used previously in this animal, ARK produced a well-defined reactivity at the cellular level without any background. We conclude that a useful panel of anti-CK antibodies commonly used in human pathology can be applied successfully to mouse tissues after enzyme digestion, leading to a more accurate definition of cellular populations in this laboratory animal. PMID- 11277419 TI - Routine formaldehyde fixation irreversibly reduces immunoreactivity of Bcl-2 in the nuclear compartment of breast cancer cells, but not in the cytoplasm. AB - Bcl-2 and Bax belong to a family of proteins involved in apoptosis regulation and are believed to reside in the cellular cytoplasm. The authors recently reported interphase nuclear localization of both proteins after immunofluorescence staining of formaldehyde- and methanol-fixed human and rodent cell monolayers. In addition, the authors' data confirmed earlier reports on Bcl-2 immunoreactivity of mitotic chromosomes in human cells. In their experience, nuclear or mitotic staining of Bcl-2, in contrast with cytoplasmic Bcl-2 immunoreactivity, is rarely observed in formaldehyde-fixed, paraffin-embedded breast cancer specimens. Therefore, the authors wondered if nuclear and mitotic Bcl-2 immunoreactivity could be irreversibly reduced by certain fixation procedures, including formaldehyde fixation. Here the authors investigated the effects of various routinely used fixation protocols and antigen retrieval techniques on Bcl-2 and Bax immunoreactivity in monolayers of MCF-7 human breast cancer cells. Whereas nuclear and mitotic immunoreactivity for Bcl-2 was clearly present after formaldehyde and methanol fixation, it was completely absent in cells fixed in acetone, methanol, or formaldehyde alone. In addition, it was found that in particular nuclear and mitotic Bcl-2, and to a lesser extent cytoplasmic Bcl-2 immunoreactivity, decreased after prolonged formaldehyde fixation, whereas Bax immunoreactivity diminished only slightly. Heat-mediated antigen retrieval after prolonged formaldehyde fixation elevated cytoplasmic, but not nuclear and mitotic, Bcl-2 immunoreactivity. PMID- 11277420 TI - An immunocytochemical approach to the demonstration of intracellular processing of mast cell carboxypeptidase. AB - Newly synthesized secretory proteins are transported from the rough endoplasmic reticulum to the Golgi complex where they can undergo posttranslational modification and are then packaged for secretion by concentration within membrane bound very small progranules that fuse to form large immature granules. The contents of these vesicles are thought to be then processed, forming mature secretory granules. After acquiring their mature appearance, the secretory granules reside in the cytoplasm until they are secreted. In this study, we raised antibodies against the first 15 N-terminal amino acids of mast cell pro carboxypeptidase and the last 14 C-terminal amino acids of mast cell carboxypeptidase. Immunohistochemical localization of the two peptides was carried out in human breast tissue and rat tissue (ear, skin, peritoneum, and tongue). In all cases, both epitopes were demonstrated only in mast cell secretory granules. However, mast cells from 3-week-old rats were more positive for the pro-enzyme compared to 3-month-old rats. Human mast cells in breast tissues were mostly negative for the pro-enzyme and positive for the carboxypeptidase. On the basis of these observations, it seems that posttransitional modification of the pro-enzyme to form the active enzyme occurs in the mast cell secretory granules. PMID- 11277421 TI - Topoisomerase alpha II, retinoblastoma gene product, and p53: potential relationships with aggressive behavior and malignant transformation in recurrent respiratory papillomatosis. AB - Recurrent respiratory papillomatosis (RRP) has a juvenile aggressive form and an adult more indolent form. Most cases of RRP are cytologically benign; however, some undergo malignant transformation. At present, there are no known markers that help identify patients at risk for aggressive disease. We investigated by immunohistochemistry expressions of topoisomerase alpha II, MIB-1, p53, p21, E cadherin, retinoblastoma (RB) gene protein product, HER-2/neu, and steroid hormone receptors in a case of juvenile respiratory papillomatosis with malignant transformation to determine whether these markers are associated with malignant transformation. Histologic examination of the pulmonary lobectomy specimen revealed well-differentiated squamous carcinoma and invasive papillomatosis. Increased staining was found in areas of invasive papillomatosis for topoisomerase alpha II, p53, and MIB-1, with highest labeling indices in areas of squamous carcinoma. Staining intensity for RB gene protein product showed gradual decline from benign papilloma (3+) and invasive papillomatosis (2+) to squamous carcinoma (0-1+). Expression of p21 was similar in benign papilloma and invasive papillomatosis but showed reduction in squamous carcinoma. Expressions of E cadherin, HER-2/neu, and steroid hormone receptors did not appear to correlate with biologic behavior. Increased topoisomerase alpha II and p53 expression along with reduced RB gene protein product and p21 expression may serve as markers of transformation to invasive papillomatosis and squamous carcinoma. PMID- 11277422 TI - Neuroendocrine differentiation in bronchial carcinomas of classic squamous-cell type: an immunohistochemical study of 29 cases applying the tyramide signal amplification technique. AB - With regard to the cellular origin of bronchial squamous-cell carcinomas, there are some clinicopathologic and experimental data indicating a link between neuroendocrine (NE) bronchial tumors and the traditionally non-NE squamous-cell carcinomas. Against this background, 29 consecutively resected bronchial squamous cell carcinomas were examined immunohistochemically (IHC) by means of the specific NE cell marker chromogranin A (CgA), using not only conventional IHC methods, but also the technique with increased sensitivity, offered by the tyramide signal amplification (TSA) procedure. Whereas none of the 29 tumors displayed CgA immunoreactive (IR) cells using the conventional IHC procedure, 10 were found to display a fine granular CgA IR in the neoplastic parenchymal cells using the TSA technique. This incidence is higher than previously reported. However, the CgA IR cells never formed any majority cell population of the neoplastic parenchyma; when present, most of them occurred as micronodules or larger confluent areas in the peripheral most undifferentiated parts of the carcinomatous sheets. Single CgA IR cells were detected only rarely in the spinocellular or keratinized areas. It can be speculated that the observations conform with the recently proposed hypothesis that there is a reservoir of NE progenitor cells in the bronchial mucosa capable of proliferation. PMID- 11277423 TI - Immunohistochemical assessment of an asymptomatic glucagonoma in a patient with hypergastrinemia and marked antral angiodysplasia. AB - A 58-year-old patient had been treated for recurrent gastritis. Numerous gastroscopies indicated hemorrhagic gastritis combined with increasingly severe anemia. The patient was admitted with a hemoglobin of 4.4 g/dL. Gastroscopy showed marked antral angiodysplasia. Serum samples for gastrin were taken and found to be elevated (170-250 U/mL). The search for a gastrin-producing tumor with abdominal ultrasound, computed tomography, octreotide scan, and secretin test was negative, but angiography detected a pancreas tumor with a 2-cm diameter. Partial pancreatectomy and partial gastrectomy were performed. Immunohistochemical examination of the tumor did not show a gastrinoma but did show glucagon-reactive tissue. Further tumors or elevated plasma hormone levels were not detected, and a multiple endocrine neoplasia type I syndrome could be excluded. We thus found antral angiodysplasia with hypergastrinemia leading to detection of a glucagonoma diagnosed by immunohistochemistry. After more than 4 years of follow-up, the patient is without any symptoms or signs of relapse or secondary hormone syndrome. PMID- 11277424 TI - Expression studies on AUX1-like genes in Medicago truncatula suggest that auxin is required at two steps in early nodule development. AB - Medicago truncatula contains a family of at least five genes related to AUX1 of Arabidopsis thaliana (termed MtLAX genes for Medicago truncatula-like AUX1 genes). The high sequence similarity between the encoded proteins and AUX1 implies that the MtLAX genes encode auxin import carriers. The MtLAX genes are expressed in roots and other organs, suggesting that they play pleiotropic roles related to auxin uptake. In primary roots, the MtLAX genes are expressed preferentially in the root tips, particularly in the provascular bundles and root caps. During lateral root and nodule development, the genes are expressed in the primordia, particularly in cells that were probably derived from the pericycle. At slightly later stages, the genes are expressed in the regions of the developing organs where the vasculature arises (central position for lateral roots and peripheral region for nodules). These results are consistent with MtLAX being involved in local auxin transport and suggest that auxin is required at two common stages of lateral root and nodule development: development of the primordia and differentiation of the vasculature. PMID- 11277425 TI - Pathogenicity determinants in the complex virus population of a Plum pox virus isolate. AB - Several subisolates were separated from a single Plum pox virus (PPV) isolate, PPV-PS. In spite of an extremely high sequence conservation (more than 99.9% similarity), different subisolates differed largely in pathogenicity in herbaceous hosts and infectivity in woody plants. The severity of symptomatology did not seem to correlate with virus accumulation. Sequence analysis and site directed mutagenesis demonstrated that single amino acid changes in the helper component (HC) protein caused a drastic effect on virus symptoms in herbaceous hosts and notably modified virus infectivity in peach seedlings. These results indicate that HC variation might play an important role in virulence evolution of natural plant virus infections. Moreover, the analysis of Potato virus X (PVX)-HC chimeras showed that the identified HC amino acid changes had parallel effects on the severity of symptoms caused by PPV and on HC-induced enhancement of PVX pathogenicity, indicating that HC functions in potyvirus symptomatology and in synergism with other viruses have overlapping determinants. PMID- 11277426 TI - Arabidopsis thaliana genes expressed in the early compatible interaction with root-knot nematodes. AB - In the compatible interaction between Arabidopsis thaliana and the endoparasitic nematode Meloidogyne incognita, galls are formed on the roots of the host plant. Differential display was used to identify alterations of gene expression in young A. thaliana root galls caused by M. incognita. Six genes were confirmed as plant genes by DNA gel blot hybridizations. Significant homology was found with a trypsin inhibitor, peroxidase, mitochondrial uncoupling protein, endomembrane protein, 20S proteasome alpha-subunit, and diaminopimelate decarboxylase. The cellular and temporal expression of each of the six genes was analyzed by mRNA in situ hybridizations. PMID- 11277427 TI - Definition of tissue-specific and general requirements for plant infection in a phytopathogenic fungus. AB - Although plant diseases are usually characterized by the part of the plant that is affected (e.g., leaf spots, root rots, wilts), surprisingly little is known about the factors that condition the ability of pathogens to colonize different plant tissues. Here we demonstrate that the leaf blast pathogen Magnaporthe grisea also can infect plant roots, and we exploit this finding to distinguish tissue-specific and general requirements for plant infection. Tests of a M. grisea mutant collection identified some mutants that were defective specifically in infection of either leaves or roots, and others such as the map kinase mutant pmk1 that were generally defective in pathogenicity. Conservation of a functional PMK1-related MAP kinase in the root pathogen Gaeumannomyces graminis was also demonstrated. Exploitation of the ability of M. grisea to infect distinct plant tissues thus represents a powerful tool for the comprehensive dissection of genetic determinants of tissue specificity and global requirements for plant infection. PMID- 11277428 TI - Transposon impala, a novel tool for gene tagging in the rice blast fungus Magnaporthe grisea. AB - impala, a Tc1-mariner transposable element from Fusarium oxysporum, was introduced into the rice blast fungus Magnaporthe grisea to develop transposon based insertional mutagenesis. A construct (pNIL160) containing an autonomous impala copy inserted in the promoter of niaD encoding Aspergillus nidulans nitrate reductase was introduced by transformation into a M. grisea nitrate reductase-deficient mutant. impala excision was monitored by restoration of prototrophy for nitrate. Southern analysis of niaD+ revertants revealed that impala was able to excise and reinsert at new loci in M. grisea. As observed for its host Fusarium oxysporum, impala inserted at a TA site left a typical excision footprint of 5 bp. We have shown that a defective impala copy was inactive in M. grisea, yet it can be activated by a functional impala transposase. A transformant carrying a single copy of pNIL160 was used to generate a collection of 350 revertants. Mutants either altered for their mycelial growth (Rev2) or nonpathogenic (Rev77) were obtained. Complementation of Rev77 with a 3-kb genomic fragment from a wild-type locus was successful, demonstrating the tagging of a pathogenicity gene by impala. This gene, called ORP1, is essential for penetration of host leaves by M. grisea and has no sequence homology to known genes. PMID- 11277429 TI - Expression of the Avirulence gene Avr9 of the fungal tomato pathogen Cladosporium fulvum is regulated by the global nitrogen response factor NRF1. AB - Here we describe the role of the Cladosporium fulvum nitrogen response factor 1 (Nrf1) gene in regulation of the expression of avirulence gene Avr9 and virulence on tomato. The Nrf1 gene, which was isolated by a polymerase chain reaction-based strategy, is predicted to encode a protein of 918 amino acid residues. The protein contains a putative zinc finger DNA-binding domain that shares 98% amino acid identity with the zinc finger of the major nitrogen regulatory proteins AREA and NIT2 of Aspergillus nidulans and Neurospora crassa, respectively. Functional equivalence of Nrf1 to areA was demonstrated by complementation of an A. nidulans areA loss-of-function mutant with Nrf1. Nrf1-deficient transformants of C. fulvum obtained by homologous recombination were unable to utilize nitrate and nitrite as a nitrogen source. In contrast to what was observed in the C. fulvum wild type, the Avr9 gene was no longer induced under nitrogen-starvation conditions in Nrf1-deficient strains. On susceptible tomato plants, the Nrf1-deficient strains were as virulent as wild-type strains of C. fulvum, although the expression of the Avr9 gene was strongly reduced. In addition, Nrf1-deficient strains were still avirulent on tomato plants containing the functional Cf-9 resistance gene, indicating that in planta, apparently sufficient quantities of stable AVR9 elicitor are produced. Our results suggest that the NRF1 protein is a major regulator of the Avr9 gene. PMID- 11277430 TI - Elicitin genes expressed in vitro by certain tobacco isolates of Phytophthora parasitica are down regulated during compatible interactions. AB - Phytophthora spp. secrete proteins called elicitins in vitro that can specifically induce hypersensitive response and systemic acquired resistance in tobacco. In Phytophthora parasitica, the causal agent of black shank, most isolates virulent on tobacco are unable to produce elicitins in vitro. Recently, however, a few elicitin-producing P. parasitica strains virulent on tobacco have been isolated. We investigated the potential diversity of elicitin genes in P. parasitica isolates belonging to different genotypes and with various virulence levels toward tobacco as well as elicitin expression pattern in vitro and in planta. Although elicitins are encoded by a multigene family, parAl is the main elicitin gene expressed. This gene is highly conserved among isolates, regardless of the elicitin production and virulence levels toward tobacco. Moreover, we show that elicitin-producing P. parasitica isolates virulent on tobacco down regulate parAl expression during compatible interactions, whichever host plant is tested. Conversely, one elicitin-producing P. parasitica isolate that is pathogenic on tomato and avirulent on tobacco still expresses parAl in the compatible interaction. Therefore, some P. parasitica isolates may evade tobacco recognition by down regulating parA1 in planta. The in planta down regulation of parA1 may constitute a suitable mechanism for P. parasitica to infect tobacco without deleterious consequences for the pathogen. PMID- 11277431 TI - The contribution of syringopeptin and syringomycin to virulence of Pseudomonas syringae pv. syringae strain B301D on the basis of sypA and syrB1 biosynthesis mutant analysis. AB - Sequencing of an approximately 3.9-kb fragment downstream of the syrD gene of Pseudomonas syringae pv. syringae strain B301D revealed that this region, designated sypA, codes for a peptide synthetase, a multifunctional enzyme involved in the thiotemplate mechanism of peptide biosynthesis. The translated protein sequence encompasses a complete amino acid activation module containing the conserved domains characteristic of peptide synthetases. Analysis of the substrate specificity region of this module indicates that it incorporates 2,3 dehydroaminobutyric acid into the syringopeptin peptide structure. Bioassay and high performance liquid chromatography data confirmed that disruption of the sypA gene in strain B301D resulted in the loss of syringopeptin production. The contribution of syringopeptin and syringomycin to the virulence of P. syringae pv. syringae strain B301D was examined in immature sweet cherry with sypA and syrB1 synthetase mutants defective in the production of the two toxins, respectively. Syringopeptin (sypA) and syringomycin (syrB1) mutants were reduced in virulence 59 and 26%, respectively, compared with the parental strain in cherry, whereas the syringopeptin-syringomycin double mutant was reduced 76% in virulence. These data demonstrate that syringopeptin and syringomycin are major virulence determinants of P. syringae pv. syringae. PMID- 11277432 TI - The nodulation protein NodG shows the enzymatic activity of an 3-oxoacyl-acyl carrier protein reductase. AB - The acyl carrier protein NodF is required for the synthesis of unusual polyunsaturated fatty acids that confer specificity to lipochitin oligosaccharide nodulation (Nod) factors of Rhizobium leguminosarum. In this study, homogeneous NodF protein was used as a ligand to identify proteins of R. leguminosarum that specifically interact with NodF and presumably are involved in the biosynthesis or transfer of the unusual fatty acids. The N-terminal amino acid sequence of a 29-kDa protein that interacts strongly with NodF revealed high similarity to NodG of Rhizobium sp. N33 and to NodG of Sinorhizobium meliloti We cloned and sequenced the gene coding for the NodG-like protein of R. leguminosarum and found it to be the product of the constitutively expressed gene fabG. FabG is the 3 oxoacyl-acyl carrier protein reductase that catalyzes the first reduction step in each cycle of fatty acid elongation. FabG of R. leguminosarum and NodG of Rhizobium sp. N33 were expressed in Escherichia coli. In both cases, the purified protein showed 3-oxoacyl-acyl carrier protein reductase activity in vitro. Therefore, NodG has the same biochemical function as FabG, and the high degree of similarity at the protein and DNA level suggest that nodG is a duplication of the housekeeping genefabG. PMID- 11277433 TI - Comparison of benefit to sugarcane plant growth and 15N2 incorporation following inoculation of sterile plants with Acetobacter diazotrophicus wild-type and Nif- mutants strains. AB - The ability of the nitrogen-fixing bacterial endophyte Acetobacter diazotrophicus strain PAl5 to enhance the growth of sugarcane SP70-1143 was evaluated in the growth chamber, greenhouse, and field by comparing plants inoculated with wild type and Nif mutant MAd3A in two independent experiments. The wild-type and Nif mutant strains colonized sugarcane plants equally and persisted in mature plants. In N-deficient conditions, sugarcane plants inoculated with A. diazotrophicus PAl5 generally grew better and had a higher total N content 60 days after planting than did plants inoculated with mutant MAd3A or uninoculated plants. These results indicate that the transfer of fixed N from A. diazotrophicus to sugarcane might be a significant mechanism for plant growth promotion in this association. When N was not limiting, growth enhancement was observed in plants inoculated with either wild-type or Nif- mutants, suggesting the additional effect of a plant growth promoting factor provided by A. diazotrophicus. A 15N2 incorporation experiment demonstrated that A. diazotrophicus wild-type strains actively fixed N2 inside sugarcane plants, whereas the Nif- mutants did not. PMID- 11277434 TI - Alcohol oxidase is a novel pathogenicity factor for Cladosporium fulvum, but aldehyde dehydrogenase is dispensable. AB - Cladosporiumfulvum is a mitosporic ascomycete pathogen of tomato. A study of fungal genes expressed during carbon starvation in vitro identified several genes that were up regulated during growth in planta. These included genes predicted to encode acetaldehyde dehydrogenase (Aldh1) and alcohol oxidase (Aox1). An Aldh1 deletion mutant was constructed. This mutant lacked all detectable ALDH activity, had lost the ability to grow with ethanol as a carbon source, but was unaffected in pathogenicity. Aox1 expression was induced by carbon starvation and during the later stages of infection. The alcohol oxidase enzyme activity has broadly similar properties (Km values, substrate specificity, pH, and heat stability) to yeast enzymes. Antibodies raised to Hansenula polymorpha alcohol oxidase (AOX) detected antigens in Western blots of starved C. fulvum mycelium and infected plant material. Antigen reacting with the antibodies was localized to organelles resembling peroxisomes in starved mycelium and infected plants. Disruption mutants of Aox1 lacked detectable AOX activity and had markedly reduced pathogenicity as assayed by two different measures of fungal growth. These results identify alcohol oxidase as a novel pathogenicity factor and are discussed in relation to peroxisomal metabolism of fungal pathogens during growth in planta. PMID- 11277435 TI - Cucumber mosaic virus-plant interactions: identification of 3a protein sequences affecting infectivity, cell-to-cell movement, and long-distance movement. AB - Mutants of the Cucumber mosaic virus (CMV) movement protein (MP) were generated and analyzed for their effects on virus movement and pathogenicity in vivo. Similar to the wild-type MP, mutants M1, M2, and M3, promoted virus movement in eight plant species. Mutant M3 showed some differences in pathogenicity in one host species. Mutant M8 showed some host-specific alterations in movement in two hypersensitive hosts of CMV. Mutant M9 showed altered pathogenicity on three hosts and was temperature sensitive for long-distance movement, demonstrating that cell-to-cell and long-distance movement are distinct movement functions for CMV. Four mutants (M4, M5, M6, and M7) were debilitated from movement in all hosts tested. Mutants M4, M5, and M6 could be complemented in trans by the wild type MP expressed transgenically, although not by each other or by mutant M9 (at the restrictive temperature). Mutant M7 showed an inability to be complemented in trans. From these mutants, different aspects of the CMV movement process could be defined and specific roles for particular sequence domains assigned. The broader implications of these functions are discussed. PMID- 11277436 TI - Relative effects on virulence of mutations in the sap, pel, and hrp loci of Erwinia chrysanthemi. AB - We constructed strains of Erwinia chrysanthemi EC16 with multiple mutations involving three virulence systems in this bacterium, namely pel (coding for the major pectate lyases pelABCE), hrp (hypersensitive response and pathogenicity), and sap (sensitivity to antimicrobial peptides). The relative effects on virulence of those mutations have been analyzed on potato tubers and chicory leaves. In potato tubers, the sap mutation (BT105) had a greater effect in the reduction of the virulence than the pel (CUCPB5006) and hrp (CUCPB5039) mutations. This reduction was similar to that observed in the pel-hrp double mutant (CUCPB5037). The analysis of the strains affected in Pel-Sap (BT106), Hrp Sap (BT107), and Pel-Hrp-Sap (BT108) suggested that the effects of these mutations are additive. In chicory leaves, the mutation in the sap locus appeared to have a greater effect than in potato tubers. The competitive indices of strains BT105, UM1005 (Pel-), CUCPB5039, and CUCPB5037 have been estimated in vivo and in vitro. These results indicate that the mutation in the hrp locus can be complemented in vivo by coinfection, whereas the mutations in pel and sap cannot. PMID- 11277437 TI - Immunocytochemical localization of HrpA and HrpZ supports a role for the Hrp pilus in the transfer of effector proteins from Pseudomonas syringae pv. tomato across the host plant cell wall. AB - The Hrp pilus, composed of HrpA subunits, is an essential component of the type III secretion system in Pseudomonas syringae. We used electron microscopy (EM) and immunocytochemistry to examine production of the pilus in vitro from P. syringae pv. tomato strain DC3000 grown under hrp-inducing conditions on EM grids. Pili, when labeled with antibodies to HrpA, developed rapidly in a nonpolar manner shortly after the detection of the hrpA transcript and extended up to 5 microm into surrounding media. Structures at the base of the pilus were clearly differentiated from the basal bodies of flagella. The HrpZ protein, also secreted via the type III system, was found by immunogold labeling to be associated with the pilus in vitro. Accumulation and secretion of HrpA and HrpZ were also examined quantitatively after the inoculation of wild-type DC3000 and hrpA and hrpZ mutants into leaves of Arabidopsis thaliana. The functional pilus crossed the plant cell wall to generate tracks of immunogold labeling for HrpA and HrpZ. Mutants that produced HrpA but did not assemble pili were nonpathogenic, did not secrete HrpA protein, and were compromised for the accumulation of HrpZ. A model is proposed in which the rapidly elongating Hrp pilus acts as a moving conveyor, facilitating transfer of effector proteins from bacteria to the plant cytoplasm across the formidable barrier of the plant cell wall. PMID- 11277438 TI - Biological activity of the rolB-like 5' end of the A4-orf8 gene from the Agrobacterium rhizogenes TL-DNA. AB - The iaaM gene from different plant-associated bacteria encodes a tryptophan monooxygenase (IaaM) that catalyzes the synthesis of indole-3-acetamide (IAM), a precursor of indole-3-acetic acid (IAA). Unlike the IaaM proteins from other bacteria, Agrobacterium spp. T-DNA-encoded IaaM proteins carry a 200 amino acid N terminal extension with low homology to various members of the RolB protein family. This family is composed of 18 highly divergent T-DNA-encoded proteins, the basic functions of which are still largely undetermined. Deletion of the 5' rolB-like extension of the iaaM gene from Agrobacterium tumefaciens strain Ach5 did not lead to a reduction in IAM synthesis in plants. When expressed in tobacco, the rolB-like fragment did not affect growth or morphology. An iaaM homolog (A4-orf8) from the TL-DNA of Agrobacterium rhizogenes strain A4 also was investigated. Neither the full-size A4-orf8 gene nor the 5'-truncated form induced detectable IAM synthesis. Plants expressing the rolB-like part of the A4 orf8 gene, however, were dwarfed and mottled to various extents and synthesized abnormally high amounts of glucose, fructose, sucrose, and starch. PMID- 11277439 TI - The C-terminal dilysine motif for targeting to the endoplasmic reticulum is not required for Cf-9 function. AB - The tomato resistance gene Cf-9 encodes a membrane-anchored, receptor-like protein that mediates specific recognition of the extracellular elicitor protein AVR9 of Cladosporium fulvum. The C-terminal dilysine motif (KKRY) of Cf-9 suggests that the protein resides in the endoplasmic reticulum. Previously, two conflicting reports on the subcellular location of Cf-9 were published. Here we show that the AARY mutant version of Cf-9 is still functional in mediating AVR9 recognition, suggesting that functional Cf-9 resides in the plasma membrane. The data presented here and in reports by others can be explained by masking the dilysine signal of Cf-9 with other proteins. PMID- 11277440 TI - The Arabidopsis downy mildew resistance gene, RPP13-Nd, functions independently of NDR1 and EDS1 and does not require the accumulation of salicylic acid. AB - RPP13-Nd-mediated resistance prevents parasitism by five isolates of Peronospora parasitica (At) in a transgenic Arabidopsis. Columbia background. We tested the effect of a number of known disease resistance mutations on the RPP13-Nd function and found that resistance remained unaltered in plants carrying mutations in either EDS1 or NDR1 and in double ndr1-1/eds1-2 mutant lines. Furthermore, we found that pbs2, pad4-1, npr1-1, and rps5-1, which compromise resistance to a number of P. parasitica (At) isolates, had no affect on RPP13-Nd function. In addition, RPP13-Nd-mediated resistance remained unchanged in a background of salicylic acid depletion (nahG). We conclude that RPP13-Nd is the first Arabidopsis R gene product reported to act via a novel signaling pathway that is independent of salicylic acid-mediated responses and is completely independent of NDR1 and EDS1. PMID- 11277441 TI - A bacterial artificial chromosome library of Lotus japonicus constructed in an Agrobacterium tumefaciens-transformable vector. AB - We constructed a BAC library of the model legume Lotus japonicus with a 6-to 7 fold genome coverage. We used vector PCLD04541, which allows direct plant transformation by BACs. The average insert size is 94 kb. Clones were stable in Escherichia coli and Agrobacterium tumefaciens. PMID- 11277442 TI - Stable RK2-derived cloning vectors for the analysis of gene expression and gene function in gram-negative bacteria. AB - The construction of several stable RK2-derived cloning vectors for the analysis of gene expression and function in gram-negative bacteria is reported. Plasmid stability is conferred by the RK2 par locus or by insertion of the spsAB or spsCD symbiotic plasmid stability loci from pNGR234a of Rhizobium sp. NGR234. The vectors carry multiple cloning sites with protection against read-through transcriptional activity of vector sequences. Vector derivatives with the constitutive nptII promoter or a promoterless gusA gene are suitable for the study of gene function or regulation in bacteria. PMID- 11277444 TI - Visual-field bias in the judgment of facial expression of emotion. AB - The left and right hemispheres of the brain are differentially related to the processing of emotions. Although there is little doubt that the right hemisphere is relatively superior for processing negative emotions, controversy exists over the hemispheric role in the processing of positive emotions. Eighty right-handed normal male participants were examined for visual-field (left-right) differences in the perception of facial expressions of emotion. Facial composite (RR, LL) and hemifacial (R, L) sets depicting emotion expressions of happiness and sadness were prepared. Pairs of such photographs were presented bilaterally for 150 ms, and participants were asked to select the photographs that looked more expressive. A left visual-field superiority (a right-hemisphere function) was found for sad facial emotion. A hemispheric advantage in the perception of happy expression was not found. PMID- 11277443 TI - Genetic organization of the hrp gene cluster and dspAE/BF operon in Erwinia herbicola pv. gypsophilae. AB - Erwinia herbicola pv. gypsophilae induces gall formation in gypsophila that is dependent on the existence of a pathogenicity plasmid (pPATHEhg). We previously demonstrated the presence of several hrp genes on this plasmid. By employing transposon mutagenesis and sequencing, a functional hrp gene cluster on the pPATHEhg has now been characterized completely. The hrp genes of E. herbicola pv. gypsophilae are remarkably similar to and colinear with those of Erwinia amylovora and Pantoea stewartii and generally showed 60 to 90% nucleotide or deduced amino acid identity. E. herbicola pv. gypsophilae, however, lacks hrpW, which is present in E. amylovora. Additionally, E. herbicola pv. gypsophilae mutants deficient in harpin production retained pathogenicity and were slightly reduced in their ability to elicit a hypersensitive response (HR) in tobacco. The "disease specific" region, dspA/EB/F, exhibited 60 to 74% identity with the dspA/EB/F loci of E. amylovora and P. stewartii, respectively. Mutations in dspA/E abolished pathogenicity of E. herbicola pv. gypsophilae but not HR elicitation on tobacco. Inactivation of HrpL reduced plant-induced transcription of dspA/E by three orders, indicating Hrp-dependent regulation. PMID- 11277446 TI - The spectrally dependent monotic component in the decreasing-loudness aftereffect: implications for dynamic auditory localization. AB - Listeners exposed to a tone increasing in intensity report an aftereffect of decreasing loudness in a steady tone heard afterward. In the present study, the spectral dependence of the monotic decreasing-loudness aftereffect (adapting and testing 1 ear) was compared with (a) the spectral dependence of the interotic decreasing-loudness aftereffect (adapting 1 ear and testing the other ear) and (b) a non-adaptation control condition. The purpose was to test the hypothesis that the decreasing-loudness aftereffect may concern the sensory processing associated with dynamic localization. The hypothesis is based on two premises: (a) dynamic localization requires monaural sensory processing, and (b) sensory processing is reflected in spectral selectivity. Hence, the hypothesis would be supported if the monotic aftereffect were more spectrally dependent and stronger than the interotic aftereffect; A. H. Reinhardt-Rutland (1998) showed that the hypothesis is supported with regard to the related increasing-loudness aftereffect. Two listeners were exposed to a 1-kHz adapting stimulus. From responses of "growing softer" or "growing louder" to test stimuli changing in intensity, nulls were calculated; test carrier frequencies ranged from 0.5 kHz to 2 kHz. Confirming the hypothesis, the monotic aftereffect peaked at around the 1 kHz test carrier frequency. In contrast, the interotic aftereffect showed little evidence of spectrally dependent peaking. Except when test and adaptation carrier frequencies differed markedly, the interotic aftereffect was smaller than the monotic aftereffect. PMID- 11277445 TI - The contributions of memory and attention processes to cognitive abilities. AB - In two experiments, the contributions of memory and attention processes to the cognitive abilities of reasoning and perceptual speed were investigated. Two measures of speed of information retrieval from long-term and short-term memory (Posner paradigm, Sternberg paradigm) and two attention measures (continuous attention test, attention switching test) were included in the first experiment (N = 220). The memory tests led to correlations with the measures of cognitive abilities, whereas the attention test did not. The same tests as well as one additional memory test and one attention test (working memory test, test of covert orientation) were administered in the second experiment (N = 116). Again, the memory tests led to the larger correlations with the measures of cognitive abilities. Two components were obtained in components analysis, of which the first was characterized by high loadings of the memory tests and the second by high loadings of the attention tests. Only the memory component contributed to the prediction of cognitive abilities. PMID- 11277447 TI - Sampling methods for identifying differences in source reliability. AB - This study explored observer performance in using across-trial and within-trial variability information to weight sources based on reliability. Three trained observers performed a multi-element, visual signal-detection task under 3 block type conditions: a fixed block condition and 2 random conditions. In the fixed block condition, the observers had both within- and across-trial variability information to identify differences in source reliability. The random conditions eliminated the across-trial information, leaving only within-trial variability information in 1 case and neither within- nor across-trial variability information in another case. There was a significant block-type effect. Observers could use differences in across-trial variability of individual sources to assign weights. Although there was a difference in the weights assigned to reliable and unreliable sources in the partial-random condition (in which within-trial variability information was available), only 1 participant showed a significant difference in the weight assignment relative to the full-random condition. Thus, altogether, observers were not very efficient at using within-trial variability to weight sources accordingly. PMID- 11277448 TI - The effect of informative and uninformative cueing of attention on feature integration. AB - In the present study, the authors observed the effect of informative and uninformative attentional cueing on visual search for targets that were defined by a simple feature or by conjunctions of features. Three different types of attentional cueing were tested in three experiments: peripheral informative cueing, peripheral uninformative cueing, and central informative cueing. Participants showed a greater effect of cueing in detecting a conjunction of features than in detecting unique features only when attention was oriented by either peripheral or central informative cueing. This differential cueing effect was not observed when attention was oriented by peripheral uninformative cueing. The results suggest that voluntarily oriented attention plays a more important role in feature integration than automatically oriented attention does. The results also pose limits on the generalizability of K. A. Briand's (1998) proposal regarding the role of automatically oriented attention in feature integration. PMID- 11277449 TI - A test of the overconfidence phenomenon using audio signals. AB - The existence of the overconfidence phenomenon was examined using a signal detection paradigm. Fifty-five participants were asked to decide whether they heard a signal or noise only, and to rate how certain they were of their decisions. The results confirmed the existence of overconfidence as well as the "hard-easy" effect. PMID- 11277450 TI - Item-cued directed forgetting of related words and pictures in children and adults: selective rehearsal versus cognitive inhibition. AB - The main purpose of this study was to compare the relative importance of selective rehearsal and cognitive inhibition in accounting for developmental changes in the directed-forgetting paradigm developed by R. A. Bjork (1972). In two experiments, children in Grades 2 and 5 and college students were asked to remember some words or pictures and to forget others when items were categorically related. Their memory for both items and the associated remember or forget cues was then tested with recall and recognition. Fifth graders recognized more of the forget-cued words than college students did. The pattern of results suggested that age differences in rehearsal and source monitoring (i.e., remembering whether a word had been cued remember or forget) were better explanatory mechanisms for children's forgetting inefficiencies than retrieval inhibition was. The results are discussed in terms of a multiple process view of inhibition. PMID- 11277451 TI - Coloring single stroop elements: reducing automaticity or slowing color processing? AB - Automaticity theory and the effect of coloring a single element were tested with all or only 1 element colored in Stroop tasks. The 312 participants in 5 experiments indicated stimulus presentation color by key press. Experiments 1 and 2 replicated those of D. Besner, J. A. Stoltz, and C. Boutilier (1997) with some changes, and revealed similar results: less Stroop interference with only 1 letter colored. Besner et al. (1997) interpreted the results as indicating that coloring a single letter eliminates automatic reading processes. The cause of that reduction in Stroop interference was investigated in Experiments 3, 4, and 5 using color words, bars, and rectangles. The effect of coloring 1 element was to increase color-naming time by the same amount for congruent and neutral, nonverbal stimuli, but not for incongruent stimuli. The results are interpreted in terms of automaticity theory, and a continuous flow approach to the Stroop effect is presented. PMID- 11277452 TI - Distinguishing short-term memory from working memory. AB - The aim of the present research was to determine whether short-term memory and working memory could be distinguished. In two studies, 7- to 13-year-olds (N = 155, N = 132) were administered tasks thought to assess short-term memory as well as tasks thought to assess working memory. Both exploratory and confirmatory factor analyses distinguished short-term memory tasks from working memory tasks. In addition, performance on working memory tasks was related to word decoding skill but performance on short-term memory tasks was not. Finally, performance on both short-term memory and working memory tasks were associated with age-related increases in processing speed. Results are discussed in relation to models of short-term and working memory. PMID- 11277453 TI - Working memory capacity and strategy use. AB - In this study, we examine the role of strategy use in working memory (WM) tasks by providing short-term memory (STM) task strategy training to participants. In Experiment 1, the participants received four sessions of training to use a story formation (i.e., chaining) strategy. There were substantial improvements from pretest to posttest (after training) in terms of both STM and WM task performance. Experiment 2 demonstrated that WM task improvement did not occur for control participants, who were given the same amount of practice but were not provided with strategy instructions. An assessment of participants' strategy use on the STM task before training indicated that more strategic participants displayed better WM task performance and better verbal skills. These results support our hypothesis that strategy use influences performance on WM tasks. PMID- 11277454 TI - Perceptual and semantic sources of category-specific effects: event-related potentials during picture and word categorization. AB - In two experiments the effect of object category on event-related potentials (ERPs) was assessed while subjects performed superordinate categorizations with pictures and words referring to objects from natural (e.g., animal) and artifactual (e.g., tool) categories. First, a category probe was shown that was presented as name in Experiment 1 and as picture in Experiment 2. Thereafter, the target stimulus was displayed. In both experiments, analyses of the ERPs to the targets revealed effects of category at about 160 msec after target onset in the pictorial modality, which can be attributed to category-specific differences in perceptual processing. Later, between about 300-500 msec, natural and artifactual categories elicited similar ERP effects across target and category modalities. These findings suggest that perceptual as well as semantic sources contribute to category-specific effects. They support the view that semantic knowledge associated with different categories is represented in multiple subsystems that are similarly accessed by pictures and words. PMID- 11277455 TI - The relation of tip-of-the-tongue states and retrieval time. AB - The tip-of-the-tongue state (TOT) is the phenomenological experience that a word is on the verge of being recalled. Participants rated TOTs as either emotional or nonemotional. In Experiment 1, given general-information questions, participants spent more time attempting retrieval during emotional TOTs than during nonemotional TOTs or n-TOTs (retrieval failures not accompanied by TOTs). Experiment 2 replicated the effect that TOTs show longer retrieval times than n TOTs. In Experiment 3, with word definitions as stimuli, retrieval times were longer for emotional TOTs. Experiment 4 showed the same relation between retrieval times and TOTs even when participants made retrospective decisions about whether they had experienced a TOT before they retrieved the correct target. Valence of emotion was correlated with correct resolution of the TOT. These results are discussed in the context of a metacognitive model, in which TOTs serve to monitor and control cognition. PMID- 11277456 TI - What happens if you retest autobiographical memory 10 years on? AB - Burt (1992a, 1992b) reported data on the autobiographical memory of diarists for events that had occurred on average 3.3 years earlier. This paper reports data on 11 of the diarists, who were recontacted after a further 10 years and who agreed to a retest of their memory. Estimates of event date and event duration from the two recall attempts were compared. As predicted, duration estimation was extremely stable and showed no detrimental effects of the additional 10 years of retention interval. Estimation of event date was predicted to show an increase in forward telescoping due to the increased remoteness of the event sample, but, contrary to this prediction, backward telescoping dominated dating errors. A combination of the establishment of a recent boundary and Kemp's (1999) associative model of dating is proposed as an explanation for these results. It is argued that the nature of dating errors may depend on the time of the event's occurrence in the life span and the age of the individual dating the events. PMID- 11277457 TI - Naming the color of a word: is it responses or task sets that compete? AB - Subjects named the colors in which high- and low-frequency words and pronounceable nonwords, otherwise matched, were displayed. Color naming was slower for all three item types than for visually equivalent strings of nonalphanumeric symbols but was no slower for words than for nonwords, nor for high-frequency words than for low-frequency words. Unpronounceable letter strings had intermediate color-naming latencies. However, frequency and lexical status had large effects on latency for reading the same words and pseudowords aloud. Interference is thus predicted not by the strength of association between a letter string and its pronunciation but by the presence of word-like constituents. We argue that the interference from an unprimed noncolor word is due to, and isolates, one of two components of the classic Stroop effect: competition from the whole task set of reading. The other component, response competition, occurs only when lexical access is sufficiently primed. PMID- 11277459 TI - Propositional reasoning: the differential contribution of "rules" to the difficulty of complex reasoning problems. AB - In Experiment 1, complex propositional reasoning problems were constructed as a combination of several types of logical inferences: modus ponens, modus tollens, disjunctive modus ponens, disjunctive syllogism, and conjunction. Rule theories of propositional reasoning can account for how one combines these inferences, but the difficulty of the problems can be accounted for only if a differential psychological cost is allowed for different basic rules. Experiment 2 ruled out some alternative explanations for these differences that did not refer to the intrinsic difficulty of the basic rules. It was also found that part of the results could be accounted for by the notion of representational cost, as it is used in the mental model theory of propositional reasoning. However, the number of models as a measure of representational cost seems to be too coarsely defined to capture all of the observed effects. PMID- 11277458 TI - Primacy in causal strength judgments: the effect of initial evidence for generative versus inhibitory relationships. AB - The order in which people receive information has a substantial effect on subsequent judgment and inference. Our focus is on the order of covariation evidence in causal learning. The first experiment shows that the initial presentation of evidence suggesting a generative causal relationship (the joint presence or joint absence of cause and effect) leads to higher judged causal strength than does the initial presentation of evidence suggesting an inhibitory relationship (the presence of cause or effect in the absence of the other). Additional studies show that this primacy effect is unlikely to be due to fatigue or to an insufficient number of learning trials. These results are not readily explained by current contingency-based or associative theories of causal induction. PMID- 11277460 TI - The influence of retrieval processes in verbal overshadowing. AB - Recent studies of eyewitness memory have observed deleterious effects of producing a verbal description on later identification accuracy of a previously viewed face, an effect termed verbal overshadowing (Schooler & Engstler-Schooler, 1990). The present research investigated whether the phenomenon of verbal overshadowing may be constrained by variation in participants' initial retrieval processes, such that verbalization of a previously viewed stimulus could produce either positive or negative influences on subsequent attempts at recollection. To assess the validity of this hypothesis, we manipulated participants' response criterion during the verbal description task. As predicted, variation in response criterion significantly influenced not only the quality of the description generated but also accuracy on a subsequent identification task. This retrieval based effect was found to persist despite either a postdescription delay (Experiment 1) or source-monitoring instructions at the time of the identification task (Experiment 2). We conclude that retrieval-based processes exert a powerful influence over the accuracy of verbalization and subsequent identification of a target face. PMID- 11277461 TI - Dissociating retention and access in working memory: an age-comparative study of mental arithmetic. AB - In two experiments, young and older adults solved arithmetic chain tasks with single-digit operands, with or without a concurrent memory load of three or six digits. Variables in the arithmetic tasks had to be replaced by digits from the screen or from the memory set. A task-irrelevant concurrent load impaired neither speed nor accuracy of arithmetic in younger adults. In Experiment 2, this was also true for older adults. A large decrease in arithmetic performance was observed, however, when variables in the arithmetic task had to be substituted by digits from the memory list. Older adults had specific problems with this condition in Experiment 1, where the substitution involved two successive steps, but not in Experiment 2, where the substitution from memory could be done in a single step. The results are difficult to reconcile with models assuming a common resource for storage and processing. Rather, they are compatible with the hypothesis that a concurrent memory load interferes with a processing task only during the points of access to working memory. Further, even though access to working memory was found to be the critical source of concurrent-load interference, it was found to be insensitive to the effects of adult aging. PMID- 11277462 TI - Involvement of short-term memory in complex mental calculation. AB - The aim of this study was to examine the involvement of the short-term memory system in complex mental addition by manipulating the phonological and visual similarity of two numbers to be added. The phonological similarity of the problems appeared to have a major effect on both speed and accuracy. However, the manipulation of visual similarity failed to have any measurable impact. This suggests that the phonological loop, rather than the visual-spatial sketch pad, would be used preferably for temporary storage of addends. An interpretation of these results in terms of the nature of the internal code underlying this task is discussed. PMID- 11277463 TI - Long-term working memory in text production. AB - In reading and other high-level cognitive tasks, Ericsson and Kintsch (1995) proposed that the limited capacity of short-term working memory (STWM) is supplemented by long- term working memory (LTWM) for individuals with a high degree of domain-specific knowledge. In Experiment 1, college students (N = 80) wrote persuasive and narrative texts concerning baseball; domain-specific knowledge about baseball and verbal ability was assessed. The results showed that verbal ability and domain-specific knowledge independently affected writing skill, supporting the view that literacy depends on both knowledge sources and refuting one argument raised in support of the LTWM hypothesis. Experiment 2 (N = 42) replicated this outcome and tested the prediction that a high degree of domain-specific knowledge would lessen interference on a secondary task. The data supported the interference prediction, offering evidence that LTWM plays a role in the production of text. PMID- 11277464 TI - Cross-modal repetition priming of heterographic homophones. AB - In two cross-modal priming experiments in French, we investigated the effects of auditorily presented heterographic homophones (an English example is /meId/) on the subsequent visual recognition of the dominant (MADE) and subordinate (MAID) printed forms. When only pronounceable, regular nonwords were used as distractor items in the lexical decision task, both dominant and subordinate forms were facilitated by the homophone prime relative to an unrelated word prime. When pseudohomophones were added among the nonword distractors, dominant targets continued to show facilitation while subordinate targets showed an inhibitory trend. These results provide evidence for inhibition-based selection in the processing of ambiguous words in the absence of any biasing context. PMID- 11277465 TI - The postdiction superiority effect in metacomprehension of text. AB - Metacomprehension accuracy for texts was greater after, rather than before, answering test questions about the texts-a postdiction superiority effect. Although postdiction superiority was found across successive sets of test questions and across successive texts, there was no improvement in metacomprehension accuracy after participants had taken more tests. Neither prediction nor postdiction gamma correlations with test performance improved with successive tests. Although the results are consistent with retrieval hypotheses, they contradict predictions made by test knowledge hypotheses, which state that increasing knowledge of the nature of the tests should increase metacomprehension accuracy. PMID- 11277466 TI - Categorical perception of relative orientation in visual object recognition. AB - The purpose of the present investigation was to determine whether the orientation between an object's parts is coded categorically for object recognition and physical discrimination. In three experiments, line drawings of novel objects in which the relative orientation of object parts varied by steps of 30 degrees were used. Participants performed either an object recognition task, in which they had to determine whether two objects were composed of the same set of parts, or a physical discrimination task, in which they had to determine whether two objects were physically identical. For object recognition, participants found it more difficult to compare the 0 degrees and 30 degrees versions and the 90 degrees and 60 degrees versions of an object than to compare the 30 degrees and 60 degrees versions, but only at an extended interstimulus interval (ISI). Categorical coding was also found in the physical discrimination task. These results suggest that relative orientation is coded categorically for both object recognition and physical discrimination, although metric information appears to be coded as well, especially at brief ISIs. PMID- 11277467 TI - Comprehending illocutionary force. AB - According to speech act theory (Searle, 1969), utterances have both a propositional content and an illocutionary force (the speech act performed with the utterance). Four experiments were conducted to examine whether utterance comprehension involves speech act recognition. Participants in all experiments first read remarks that could be characterized by a particular speech act (e.g., beg). A recognition probe reaction time procedure was used in Experiments 1 and 2; participants indicated whether a probe word had literally appeared in the last remark that they had read. Participants were significantly slower at making this judgment (and made significantly more errors) when the probe represented the speech act performed with the prior remark than when it did not. A lexical decision task was used in Experiments 3 and 4, and participants were significantly faster at verifying target words representing the speech act performed with a remark, relative to control words. Overall, the results suggest that speech act recognition may be an important component of the comprehension of conversational remarks. PMID- 11277469 TI - Heart failure management: 25 years of progress. PMID- 11277468 TI - The pointedness effect on representational momentum. AB - An observer's memory for the final position of a moving object is shifted forward in the direction of that object's motion. It is called representational momentum (RM). This study addressed stimulus-specific effects on RM. In Experiment 1, participants showed larger memory shift for an object moving in its typical direction of motion than when it moved in a nontypical direction of motion. In Experiment 2, participants indicated larger memory shift for a pointed pattern moving in the direction of its point than when it moved in the opposite direction. In Experiment 3, we again examined the influences of knowledge about objects' typical motions and the pointedness of objects, because we did not control the shape (pointedness) of objects in Experiment 1. The results showed that only pointedness affected the magnitude of memory shift and that the effect was smaller than the momentum effect. PMID- 11277470 TI - Growth and change in the prescribing of anti-depressants in New Zealand: 1993 1997. AB - AIMS: To examine changes in the prescribing of anti-depressants in New Zealand from 1993-1997, in terms of expenditure, the number of dispensings and days of therapy supplied. METHOD: Data on subsidised dispensings of anti-depressant drugs during 1993 to 1997 were obtained from PHARMAC and analysed using SAS. RESULTS: The overall size of the anti-depressant market increased considerably over the study period. Government expenditure rose 2.25 times, and 1.65 times as many days of anti-depressant medication were supplied in 1997 as in 1993. Most of this was due to the growth in prescribing of newer anti-depressants, but the use of older drugs remained constant. CONCLUSIONS: In common with other countries, the use of newer agents is contributing to increased overall use of anti-depressant medication and government expenditure in New Zealand. Use of older drugs has not diminished substantially. PMID- 11277471 TI - Regional variation in anti-depressant dispensings in New Zealand: 1993-1997. AB - AIMS: To examine regional differences in the prescribing of anti-depressants in New Zealand from 1993 to 1997, and to examine the composition and dynamics of these differences. METHODS: Data on every subsidised dispensing of anti depressant drugs 1993 to 1997 were obtained from PHARMAC and analysed using SAS. Each dispensing was allocated to a regional council area on the basis of the location of the dispensing pharmacy. RESULTS: Prescribing of anti-depressants increased with time in all regions. However, there was substantial regional variation in prescribing rates per-capita, the highest being 2.28 to 2.49 times the lowest in every year. Regions also varied substantially in the mix of newer and older drugs used, although newer drugs became increasingly important in every region. CONCLUSIONS: Regional differences in anti-depressant prescribing are large. Further research with different data sources is required to explore the reasons for this variation. PMID- 11277472 TI - Non-invasive methods for measuring data quality in general practice. AB - AIM: To develop non-invasive methods of measuring the quality of data recorded in general practice. METHODS: Laboratory and pharmaceutical claims data from fourteen practices (44 doctors) from the FirstHealth network of general practices were examined to determine the extent to which valid minimum bounds on expected rates of diagnosis coding could be established. These were compared with recorded rates in patient notes to measure completeness of diagnosis recording. Data completeness was measured for demographic data and a marker for the accuracy of gender coding was developed from diagnosis data. RESULTS: Minimum rates of diagnosis could be established for asthma, diabetes (NIDDM and IDDM), ischaemic heart disease, hypothyroidism, bipolar affective disorder and Parkinson's disease. Minimum bounds for the number of patients requiring monitoring of warfarin and digoxin levels were also established. These expected minimum rates were combined with measures of completeness of age, gender, ethnicity and smoking data, and a gender coding accuracy measure, to produce a set of fourteen data quality indicators. Pass/fail thresholds on each indicator were set and each of the fourteen practices was scored on the number of passes they achieved. The scores ranged from three to nine out of fourteen passses. CONCLUSIONS: Non invasive data quality measures may be useful in providing feedback to general practitioners as part of a data quality improvement cycle. The sensitivity of this method will decline as data quality improves. PMID- 11277473 TI - Delays in the investigation of allegations of child sexual abuse in the Wellington city district 1995-1996: a retrospective study. AB - AIMS: To determine the duration of the statutory investigation process after referral of alleged chid sexual abuse and to assess which components of this process are most prone to delay. METHODS: Retrospective review of police, Child Youth and Family (CYF) and medical records for 123 young persons <17 years old for whom a referral regarding alleged sexual abuse was made to the Wellington Serious Abuse Team from January 1995 to December 1996. RESULTS: There were 82 (66.7%) females and 41 (33.3%) males referred. Maori and Pacific Island children were over-represented in the sample. The median time from referral to evidential interview or diagnostic interview was 47 days. This period was longer for children <5 years of age (66 days) compared with children > or =5 years of age (45.5 days), although this difference was not statistically significant. Although 53.3% of children alleged genital contact, only 26% were referred for a medical assessment. The time from initiation of investigation to completion was a median of 141 days. Reasons for delay were difficult to delineate but appeared to relate to inadequate staffing. CONCLUSIONS: There is an unreasonable delay in the investigation of alleged child sexual abuse. This is particularly concerning in younger children. PMID- 11277474 TI - Clinical use of beta-blocker therapy in patients with heart failure: a practical guide. PMID- 11277475 TI - Spontaneous intramural aortic haematoma. PMID- 11277476 TI - Treatment delays for patients with acute myocardial infarction within the Coromandel region of New Zealand. AB - AIM: To assess treatment delays incurred by Coromandel patients requiring thrombolytic therapy for acute myocardial infarction at Thames Hospital. METHODS: A chart search was undertaken at Thames Hospital to identify all patients admitted from July 1993 to July 1998 with a diagnosis of acute myocardial infarction who were treated with thrombolytic therapy. Times of pain onset, general practitioner (GP) assessment, transportation, hospital arrival and thrombolytic administration were noted. Additional information, when required, was obtained from the patient's GP or the St Johns Ambulance service. RESULTS: 153 patients were thrombolysed at Thames Hospital over this period, mean age 65.1 years, 36.6% in heart failure. The mean time from pain onset to GP contact was 157.2 minutes and varied from 63.2 minutes in Coromandel Township to 272.5 minutes in Pauanui. Delays from GP contact to thrombolysis were longer for patients living in outlying areas versus Thames and its environs, 169.4 +/- 45.9 versus 125.2 +/- 50.4 minutes (mean +/-SD) respectively, p<0.001. This contributed to a total delay from pain onset to thrombolysis of 316.7 +/- 145.8 minutes for patients in outlying areas versus 269.1 +/- 185.8 minutes for local patients (p=0.014). CONCLUSIONS: Delays in providing thrombolytic therapy for acute infarct patients reflect not only transport times but also delays in seeking initial medical assessment and hospital triage times. Transport times become particularly significant for those outside of Thames and its environs. Only with improved patient education and local delivery of thrombolytic therapy will these delays be adequately addressed. PMID- 11277477 TI - Outcome following heart transplantation in Maori and Polynesian patients: a comparison with European New Zealanders. AB - AIM: To compare demographic, clinical and outcome data of Maori and Polynesian with New Zealand European heart transplant patients. METHODS: A retrospective analysis was made of data from the 104 patients who underwent heart transplantation at Green Lane Hospital over a period of twelve years, of whom 79 were European, 23 Maori/Polynesian, and two Asian. Clinical characteristics, blood group, HLA matching and outcomes of recipients were compared. RESULTS: There was no significant difference in age and gender between the two groups. Maori and Polynesian patients were heavier, had a greater body mass index and were more likely to have rheumatic heart disease than their European counterparts. Maori/Polynesian patients were predominantly blood group A, whilst European patients were mainly group O. The waiting time for a donor heart was similar in both groups. There was no significant difference in number of rejection episodes and survival. CONCLUSIONS: Green Lane Hospital has the largest international experience of heart transplantation in Maori and Polynesian patients. Although there are some differences in clinical profile, outcome in terms of rejection episodes and survival is similar in the two groups. PMID- 11277478 TI - Turning off growth signals in tumours: a new approach to cancer chemotherapy. PMID- 11277479 TI - Getting health back on track. PMID- 11277480 TI - Anxiety and pulmonary embolism. PMID- 11277481 TI - Health and the World Trade Organization. PMID- 11277482 TI - Is chlamydial infection underdiagnosed--particularly in teenage males? PMID- 11277483 TI - Pacific peoples killed by New Zealand tobacco industry exports. PMID- 11277484 TI - Trauma system coordination in New Zealand--are we going forwards or backwards? PMID- 11277485 TI - Prevalence of viral hepatitis. PMID- 11277486 TI - Re-run: what isn't medical practice? PMID- 11277488 TI - Professional misconduct. PMID- 11277487 TI - Medicolegal diary: standards of conduct. PMID- 11277489 TI - Effects of the actin-stabilizing drug, jasplakinolide, on pigment granule motility in isolated retinal pigment epithelial (RPE) cells of green sunfish, Lepomis cyanellus. AB - The retinal pigment epithelium (RPE) of teleosts contains pigment granules that migrate in response to changes in light condition. Dissociated, cultured RPE cells in vitro can be triggered to aggregate or disperse pigment granules by the application of cAMP or dopamine, respectively. Previous research using the actin disrupting drug, cytochalasin D, suggested that pigment granule motility is actin dependent. To further examine the role of actin in pigment granule motility, we tested the effects of the actin-stabilizing drug, jasplakinolide, on pigment granule motility. Pigment granules in previously dispersed RPE cells remained dispersed after jasplakinolide exposure (0.1-1 microM), but the drug halted movement of most pigment granules and stimulated rapid bi-directional movements in a small subset of granules. Jasplakinolide also blocked net pigment granule aggregation and interfered with the maintenance of full aggregation. Although jasplakinolide did not block pigment granule dispersion, it did alter the motility of dispersing granules compared to control cells; rather than the normal saltatory, primarily centrifugal movements, granules of jasplakinolide-treated cells demonstrated slow, creeping centrifugal movements and more rapid bi directional movements. Jasplakinolide also altered cell morphology; the length and thickness of apical projections increased, and enlarged, paddle-like structures, which contained F-actin appeared at the tips of projections. Actin antibody labeling of jasplakinolide-treated cells revealed a more reticulated network of actin compared to antibody-labeled control cells. These results indicate that jasplakinolide-induced disruption of the actin network compromises normal pigment granule dispersion and aggregation in isolated RPE cells, thus providing further evidence that these movements are actin dependent. PMID- 11277490 TI - Applications of spectral imaging: detection and analysis of human melanoma and its precursors. AB - Light-based imaging has extensive applications for medicine and biology, and recent advances in optical imaging modalities, such as confocal and multi-photon scanning fluorescence microscopy, bioluminescence, optical coherence tomography, and spectral imaging, have opened new avenues for visualizing and recording over time dynamic changes in genetic, developmental, and disease mechanisms that cannot be captured by conventional light microscopy. In the present article, we focus on spectral imaging, and using human melanoma and its precursor lesions as an example, we describe the ability of spectral imaging to detect early-stage disease, capture gene expression profiles in tissue specimens, and visualize gene functions in tumors growing in living animals. PMID- 11277491 TI - Interaction of major coat color gene functions in mice as studied by chemical analysis of eumelanin and pheomelanin. AB - Melanocytes produce two chemically distinct types of melanin pigments, eumelanin and pheomelanin. These pigments can be quantitatively analyzed by acidic permanganate oxidation or reductive hydrolysis with hydriodic acid to form pyrrole-2,3,5-tricarboxylic acid or aminohydroxyphenylalanine, respectively. About 30 coat color genes in mice have been cloned, and functions of many of those genes have been elucidated. However, little is known about the interacting functions of these loci. In this study, we used congenic mice to eliminate genetic variability, and analyzed eumelanin and pheomelanin contents of hairs from mice mutant at one or more of the major pigment loci, i.e., the albino (C) locus that encodes tyrosinase, the slaty (Slt) locus that encodes tyrosinase related protein 2 (TRP2 also known as dopachrome tautomerase, DCT), the brown (B) locus that encodes TRP1, the silver (Si) locus that encodes a melanosomal silver protein, the agouti (A) locus that encodes agouti signaling protein (ASP), the extension (E) locus that encodes melanocortin-1 receptor, and the mahogany (Mg) locus that encodes attractin. We also measured total melanin contents after solubilization of hairs in hot Soluene-350 plus water. Hairs were shaved from 2-3 month-old congenic C57BL/6J mice. The chinchilla (c(ch)) allele is known to encode tyrosinase, whose activity is about one third that of wild type (C). Phenotypes of chinchilla (c(ch)/c(ch)) mice that are wild type or mutant at the brown and/or slaty, loci indicate that functioning TRP2 and TRP1 are necessary, in addition to high levels of tyrosinase, for a full production of eumelanin. The chinchilla allele was found to reduce the amount of pheomelanin in lethal yellow and recessive yellow mice to less than one fifth of that in congenic yellow mice that were wild type at the albino locus. This indicates that reduction in tyrosinase activity affects pheomelanogenesis more profoundly compared with eumelanogenesis. Hairs homozygous for mutation at the slaty locus contain 5,6 dihydroxyindole-2-carboxylic acid (DHICA)-poor melanin, and this chemical phenotype was retained in hairs that were mutant at both the brown locus and the slaty locus. Hair from mice mutant at the brown locus, but not at the slaty locus, do not contain DHICA-poor melanin. This indicates that the proportion of DHICA in eumelanin is determined by TRP2, but not by TRP1. Mutation at the slaty locus (Slt(lt)) was found to have no effect on pheomelanogenesis, supporting a role of TRP2 only in eumelanogenesis. The mutation at silver (si) locus showed an effect similar to brown, a partial suppression of eumelanogenesis. The mutation at mahogany (mg) locus partially suppressed the effect of lethal yellow (Ay) on pheomelanogenesis, supporting a role of mahogany in interfering with agouti signaling. These results show that combination of double mutation study of congenic mice with chemical analysis of melanins is useful in evaluating the interaction of pigment gene functions. PMID- 11277493 TI - Herpesvirus connection in the expression of autoimmune vitiligo in Smyth line chickens. AB - The Smyth line (SL) chicken is an animal model for human vitiligo, a common acquired depigmentary disorder affecting about 1-2% of people worldwide. The vitiligo-like depigmentation in SL chickens typically develops when the birds are between 6 and 14 weeks of age and may affect 70-95% of hatch mates. The development of SL vitiligo is considered to depend on two interacting components, namely an inherent melanocyte defect and an autoimmune reaction to melanocytes. Recently, a role for an environmental factor in the expression of vitiligo was suggested by the observation that only 10% of SL chicks imported from the University of Massachusetts (UM) and reared in isolation at biosecurity level 2 (BSL 2) at the University of Arkansas (UA) exhibited vitiligo. Following further assessment of environmental differences between UA and UM SL chickens, three environmental factors that may have influenced the expression of SL vitiligo were identified. Included were housing condition, status of Mycoplasma synoviae infection, and turkey herpesvirus (HVT) vaccination status. Studies were subsequently conducted at UA and UM to assess the role of these environmental factors in the expression of SL vitiligo. M. synoviae infection was not found necessary for vitiligo expression in SL chickens. However, HVT emerged as a strong candidate for an important environmental factor in SL vitiligo. The connection between HVT and SL vitiligo was confirmed for both BSL 2 and conventional housing. Therefore, the observations reported here suggest a strong causative link between HVT infection and SL vitiligo. PMID- 11277492 TI - Tyrosine levels regulate the melanogenic response to alpha-melanocyte-stimulating hormone in human melanocytes: implications for pigmentation and proliferation. AB - Melanocyte-stimulating hormone (alpha-MSH) increases cytosolic levels of cAMP as well as tyrosinase activity in murine melanocytes. These activities depend upon the presence of melanin precursors and may differ in human melanocytes. In this study, we demonstrate that high levels of tyrosine (3.7 mM), the chief melanin precursor, reduced the proliferative effect of alpha-MSH and altered human melanocyte morphology as compared to treatment with low (25-30 microM, half physiological) levels of tyrosine. The anti-proliferative effect of high levels of tyrosine was not restricted to alpha-MSH; tyrosine also reduced proliferation induced by forskolin, a direct activator of the cAMP pathway. Exposure to low tyrosine levels and alpha-MSH induced a dendritic morphology; in the presence of high tyrosine and alpha-MSH, melanocytes displayed large, pigmented cell bodies and less dendricity. Exposure to alpha-MSH in the presence of low tyrosine for up to 9 days did not appreciably increase melanin levels, but culturing the human melanocytes in high levels of tyrosine with alpha-MSH increased melanin levels 10 50-fold, depending on the pigmentation background of the donor. A greater induction of melanin accumulation was observed in melanocytes derived from light skinned donors than was observed in cells obtained from dark-skinned donors. The poor ability of alpha-MSH to stimulate melanin synthesis was not caused by a lack of induction of melanogenic proteins, as alpha-MSH increased the expression of microphthalmia (MITF), tyrosinase, dopachrome tautomerase (DCT), and Pmel-17, compared to untreated cells or cells stimulated by phorbol ester alone, regardless of tyrosine levels. DCT levels were greatly induced by low tyrosine with alpha-MSH, but were dramatically decreased by high tyrosine with alpha-MSH. Interestingly, in this same medium (high tyrosine), MITF levels also decreased after 2 weeks and were barely detectable by the third week. Despite the absence of MITF at 3 weeks of treatment in high tyrosine medium, tyrosinase levels remained high, thereby suggesting that additional factors must be responsible for tyrosinase transcription in human melanocytes. Our results indicate that tyrosine levels can regulate the proliferative activity induced by alpha-MSH, as well as the extent of melanogenesis in normal human melanocytes. The significance of this work is that tyrosine levels may be part of the mechanism that switches melanocytes out of a proliferative status and into a melanin-synthesizing, terminally differentiated phenotype. PMID- 11277494 TI - New era of cell re-discovery led to the control of melanogenesis/melanoma: a scientific journey into terra incognita. PMID- 11277495 TI - Some thoughts on a current trend in pigment cell research. PMID- 11277496 TI - Increased blood levels of human S100 in melanoma chick embryo xenografts' circulation. AB - The chorioallantoic membrane (CAM) of the fertilized egg allows grafting of human melanomas for short-term investigations and offers the opportunity to investigate the behavior of metastasizing cells and the release of S100beta into peripheral blood. Tissue from one primary melanoma as well as cutaneous and subcutaneous metastases of 10 melanoma patients with elevated levels of S100 in the peripheral blood before surgery were transplanted onto the CAM of chick embryos at day 5/6 of development. Grafts were nourished by the host blood supply 2 days after transplantation. Histologically, 3 days after grafting, metastasizing melanoma cells could be found near the vessels of the host membrane, penetrating the endothelial layer and entering the blood system. Growth conditions remained stable for 6 days after transplantation. Blood samples were taken from a larger CAM vessel before collecting the xenografts 5 days after grafting. Measurement of human S100 in peripheral blood was performed in a blinded manner. No negative control showed elevated levels of human S100 protein. Samples deriving from melanoma xenografts contained highly elevated levels of S100 protein in 80% of cases. The data strongly support the concept of graft-host interaction concerning adherence of tumors and extravasation of human melanoma cells. PMID- 11277497 TI - A diepitopic sequential oligopeptide carrier (SOCn) as mimic of the sm autoantigen: synthesis, conformation and biological assays. AB - Anti-Sm (Sm: U1-U6 RNA-protein complex) antibodies are usually considered highly specific for systemic lupus erythematosus (SLE), while anti-U1RNP (U1RNP: U1RNA protein complex) are thought of as diagnostic criteria for the mixed connective tissue disease (MCTD). However, both antibody specificities coexist in SLE and MCTD, in varying percentages. Although the anti-Sm/anti-U1RNP immunological cross reactivity has been initially attributed to a common motif, PPXY(Z)PP (where X, Y, Z are various amino acids), found in the Sm, U1-A and U1-C autoantigens, it appears that the conformational features of the Sm epitopes also play an important role in the immunoreactivity. The PPGMRPP and PPGIRGP main epitopes of the Sm antigen were coupled in duplicate to the tetrameric Ac-(Lys-Aib-Gly)4-OH, SOC4, carrier to form the [(PPGMRPP)2, (PPGIRGP)2]-SOC4 construct as a mimic of the native Sm. It was found that: (i) the 3(10) helical structure of SOC4 allows the epitopes to adopt an exposed orientation, similar to their free forms, that facilitates their recognition from the anti-Sm antibodies, and (ii) the U1-RNP cross-reactivity is minimized. PMID- 11277498 TI - Insights into peptide and protein function: a convergent approach. AB - From viruses to multicellular organisms, life is inseparable from the genetic instructions aimed at regulating its maintenance, division, multiplication, differentiation and death (apoptosis). Over the past 15 years, structural studies have begun to resolve the complex reactions involved in these fundamental processes in biology. The three-dimensional representations of the complexes formed with peptides and/or proteins have allowed interpretation of the biochemical data and formulation of novel hypotheses about the control and execution of these processes. Moreover, they have opened the way to rational approaches for designing compounds able to interfere with these crucial events in normal or pathological conditions. Various results obtained in our laboratory in these fields are briefly summarized in this review. PMID- 11277499 TI - Antibiotic activity of pentadecapeptides modelled from amino acid descriptors. AB - Pentadecapeptides based on modified murine lactoferricin (LFM) sequences show varying degrees of antibacterial activity against Escherichia coli and Staphylococcus aureus. By means of projections to latent structures (PLS), a good correlation is obtained if the biological activity is modelled as a function of variables describing peptide properties, e.g. alpha-helicity, hydrophobicity/hydrophilicity and charge. Using variables derived from a principal component analysis (PCA) of all naturally occurring amino acids, it is possible to describe the amino acid content of the peptides using three variables per amino acid position. The resulting descriptor matrix is then used to develop quantitative structure-activity relationships (QSAR). It is shown that the theoretically derived descriptors model the activity of the peptides better than the earlier model, and that properties of the peptides other than antibacterial activity can be predicted. PMID- 11277501 TI - Influence of N-terminal residue stereochemistry on the prolyl amide geometry and the conformation of 5-tert-butylproline type VI beta-turn mimics. AB - The effects of N-terminal amino acid stereochemistry on prolyl amide geometry and peptide turn conformation were investigated by coupling both L- and D-amino acids to (2S, 5R)-5-tert-butylproline and L-proline to generate, respectively, N (acetyl)dipeptide N'-methylamides 1 and 2. Prolyl amide cis- and trans-isomers were, respectively, favored for peptides 1 and 2 as observed by proton NMR spectroscopy in water, DMSO and chloroform. The influence of solvent composition on amide proton chemical shift indicated an intramolecular hydrogen bond between the N'-methylamide proton and the acetamide carbonyl for the major conformer of dipeptides (S)-1, that became less favorable in (R)-1 and 2. The coupling constant (3J(NH,alpha)) values for the cis-isomer of (R)-1 indicated a phi2 dihedral angle value characteristic of a type VIb beta-turn conformation in solution. X-ray crystallographic analysis of N-acetyl-D-leucyl-5-tert butylproline N'-methylamide (R)-lb showed the prolyl residue in a type VIb beta turn geometry possessing an amide cis-isomer and psi3-dihedral angle having a value of 157 degrees, which precluded an intramolecular hydrogen bond. Intermolecular hydrogen bonding between the leucyl residues of two turn structures within the unit cell positioned the N-terminal residue in a geometry where their phi2 and psi2 dihedral angle values were not characteristic of an ideal type VIb turn. The circular dichroism spectra of tert-butylprolyl peptides (S)- and (R)-1b were found not to be influenced by changes in solvent composition from water to acetonitrile. The type B spectrum exhibited by (S)-1b has been previously assigned to a type VIa beta-turn conformation [Halab L, Lubell WD. J. Org. Chem. 1999; 64: 3312-3321]. The type C spectrum exhibited by the (R)-lb has previously been associated with type II' beta-turn and alpha-helical conformations in solution and appears now to be also characteristic for a type VIb geometry. PMID- 11277500 TI - Synthesis, conformational analysis and biological activities of lanthionine analogs of a cell adhesion modulator. AB - Cell adhesion is critical for many biological processes, such as hemostasis, wound healing, tumor metastasis and inflammation. Integrins are important mediators of cell adhesion. The integrin alpha4beta1, also known as VLA-4, is a cell surface receptor involved in inflammation. A cyclic peptide, 1-FCA-Arg-c[Cys Asp-Thz-Cys]-OH, is a potent antagonist to VLA-4 with an IC50 of 2.4 nM. In the current study, we synthesized the lanthionine analogs of 1-FCA-Arg-c[Cys-Asp-Thz Cys]-OH and determined the conformations of both the parent compound and its lanthionine analog in solution by NMR and computer simulations. The lanthionine analog retains its selectivity to VLA-4 with high nanomolar potency. Both molecules adopt similar topological arrangements in their conformations, while some important differences remain in the sulfur bridge region, which may cause the difference in potency. PMID- 11277502 TI - STFA '98: an international conference on slipping, tripping and falling accidents. PMID- 11277503 TI - The perceptions of managers and accident subjects in the service industries towards slip and trip accidents. AB - Slips and trips are a major cause of injury and lost time, and remain so despite specific initiatives. The work presented here focuses on how accident subjects and their managers each perceive the circumstances of an accident with respect to causal responsibility. To investigate this issue, a research questionnaire was designed, piloted and then applied to 33 occupational slip and trip accidents reported to a Local Authority Inspectorate. The results showed important differences in the attribution of causal responsibility between those who experience and those who investigate slips and trip accidents. The levels of agreement between individual accident subjects and their managers was at best fair (using kappa). The consequences of this and the limited scope of investigation carried out by managers is highlighted and the need for improved training and the development of a practical model of risk assessment and investigation for managers is advanced. PMID- 11277504 TI - Identification of risk factors and countermeasures for slip, trip and fall accidents during the delivery of mail. AB - Risk factors for slip, trip and fall accidents (STFA) during the delivery of mail were identified using a range of accident-centred and accident-independent methods. Key factors included slippery underfoot conditions, non-weather related environmental hazards (e.g., uneven paving, steps, inadequate lighting), poor slip resistance from footwear, unsafe working practices, management safety practices, and underlying organisational influences. Intervention measures were recommended that target STFA risks at three levels: slip resistance, exposure to hazardous conditions, and employee behaviour in the face of hazardous conditions. The use of a participative approach to intervention selection and design enabled allowance for the organisational context to be made. PMID- 11277505 TI - Serious stair injuries can be prevented by improved stair design. AB - An estimated 2.5 million injuries, and a further 4000 deaths in the UK in 1995 were due to home accidents. About 230,000 of these injuries and 497 deaths resulted from falls on stairs. When exposure is taken into account, stairs are one of the most hazardous locations in buildings. Of these falls, those where the person falls forward are most likely to cause severe injuries. One aspect of stair design, the "going" (the horizontal distance between two consecutive nosings) can be changed to decrease the potential number of serious trip accidents. Builders should therefore be encouraged to build stairs with larger goings. PMID- 11277506 TI - An investigation of underfoot accidents in a MAIM (Merseyside Accident Information Model) database. AB - Underfoot accidents taken from a study of patient interviews have been analysed to investigate factors that may be implicated in the causes of the accidents. Within the sample of patients in this study women holding items are more at risk than men from underfoot accidents. Carrying items such as shopping and handbags may obstruct the line of sight to underfoot hazards, affect balance and adversely affect reflex corporal movements that may help prevent injury. PMID- 11277508 TI - Slip, trip and fall accidents: relationship to building features and use of coroners' reports in ascribing cause. AB - Coroners' reports of 1035 deaths possibly related to building features were examined to assess their usefulness in identifying the cause of slip, trip and fall (STF) fatalities. Of the total, falls accounted for over 80%. Of the STF deaths, 61.4% related to falls on stairs, 6.7% to falls from steps or ladders, and 5.5% to falls from windows or roofs. About 60% of total fall fatalities involved infirm persons; alcohol was involved in 60% of the falls in the under-50 age group. Information in coroners' reports is generally insufficient to link building features to injuries; better approaches to data collection are required. PMID- 11277507 TI - Slips, trips and falls in different work groups--with reference to age and from a preventive perspective. AB - Inter-record linkage between two Swedish databases on population and injury was effected to provide information on occupational slip, trip and fall (STF) accidents. The text descriptions in more than 1600 accident reports from occupational groups with high incidence rates of STF accidents were categorised by gender and age and the factors contributing to the accidents studied. Both older male and female workers had higher rates of reported STF accidents than younger workers, but it was established that within any one occupation the workplace hazards were common to all. Both for men and for women, the initial approach to the prevention of STF accidents should be to improve orderliness in the workplace. PMID- 11277509 TI - Development of a portable test device for assessing on-site floor slipperiness: an interim report. AB - The main objective was to design and construct a prototype portable slipmeter with the capability of measuring static, transitional kinetic and steady-state kinetic coefficient of friction properties of on-site floors. The second objective was to evaluate its operation in the laboratory, using a commercial force platform as reference. The prototype was found to be capable of measuring the described frictional characteristics of floor surfaces, using three different test wheels and two modes of operation, impact and non-impact testing. The results anticipate that the slipmeter may prove to be more valid than any traditional measurement technique. The study continues with biomechanical trials and will be completed during the year 2000. PMID- 11277510 TI - The effect of surface roughness and contaminant on the dynamic friction of porcelain tile. AB - Surface roughness affects friction, but it is not clear what surface roughness characteristics are better correlated with friction. The average of the maximum height above the mean line in each cut-off length (Rpm) and the arithmetical average of surface slope (deltaa) had the highest correlation with dynamic friction coefficient in a previous study. The previous study was expanded to two different footwear materials and four different contaminants on a porcelain tile in the current investigation. The results showed that dynamic friction decreased as the interface speed and glycerol content in the contaminant were increased due to the hydrodynamic lubrication effect. Deltaa had the highest correlation with friction for most of the test conditions with neolite. For Four S rubber, friction coefficient appeared to have the highest correlation with the parameters related to the surface void volume at 30% glycerol content, related to the surface slope at 70 and 85% glycerol contents, and related to the peak to valley distance at 99% glycerol content. A good indicator of surface slip resistance probably should consist of the surface parameters representing the surface slope, the surface void volume and the surface peak-to-valley distance with the coefficients determined by the system parameters. PMID- 11277511 TI - The effect of roughness, floor polish, water, oil and ice on underfoot friction: current safety footwear solings are less slip resistant than microcellular polyurethane. AB - Research over a period of about 18 years has shown that a microcellular polyurethane known as AP66033 is the most slip-resistant safety footwear soling material on oily and wet surfaces. In recent years it has been replaced in commercially available footwear by a dual density polyurethane (DDP) which has a dense outer layer and a soft microcellular backing. This research programme has compared the slip resistance of AP66033 with DDP and some rubber solings. In addition, data were obtained on the effects of soling and floor roughness, and floor polish on slip resistance. Some data were also obtained for walking on ice. The coefficient of friction (CoF) of the solings was measured on 19 water wet surfaces in three conditions: (I) when the solings were new, (II) following abrasion to create maximum roughness and (III) after polishing. The CoF was measured on four oily surfaces after each of 11 abrasion or polishing treatments. The profound effects of the roughening of all soles and of floor roughness on the CoF were demonstrated for both wet and oily surfaces. The superior slip resistance of AP66033 was confirmed for oily and wet conditions; however, some rubbers not suitable for safety footwear achieved higher CoF values on wet floors. All of the floor polishes reduced the CoF of all floors when contaminated with water. The mean CoF of DDP solings was lower than the mean for AP66033 on wet and oily surfaces. No safety footwear soling provided adequate grip on dry ice and the CoF was reduced by water on the ice. A rubber used for rock climbing footwear was one of the most slip-resistant solings on wet surfaces in the laboratory but recorded the lowest CoF on ice. It is concluded that the incidence of occupational injuries caused by slipping could be reduced by the following: (A) returning to safety footwear soled with the microcellular polyurethane AP66033; (B) abrading all new and smooth footwear solings with a belt sanding machine coated with P100 grit; (C) avoiding the use of floor polish; (D) informing the general public about the poor slip resistance of ordinary footwear on ice and the lowering of slip resistance in cold weather. PMID- 11277512 TI - An improved synthesis of 1,2,4-oxadiazoles on solid support. AB - The use of tetra-N-butylammonium fluoride (TBAF) as a mild and efficient reagent for the cyclodehydration of O-acyl amidoximes has been extended to the synthesis of 1,2,4-oxadiazoles on solid support. Argopore MB-CHO resin (Argonaut Technologies) was reductively aminated and subsequently acylated with 4 cyanobenzoyl chloride. Conversion of the nitrile to the amidoxime and acylation with a range of acid chlorides in parallel followed by treatment with TBAF under ambient conditions afforded a library of 3,5-disubstituted 1,2,4-oxadiazoles. PMID- 11277513 TI - 2,4-Thiazolidinediones as potent and selective human beta3 agonists. AB - Methylsulfonamide substituted 2,4-thiazolidinedione 22c is a potent (EC50=0.01 microM, IA=1.19) and selective (more than 110-fold over beta1 and beta2 agonist activity) beta3 agonist. This compound has also been proven to be active and selective in an in vivo mode. PMID- 11277514 TI - Peptidomimetic inhibitors of protein farnesyltransferase show potent antimalarial activity. AB - Malaria continues to represent a very serious health problem in the tropics. The current methods of clinical treatment are showing deficiencies due to the increased incidence of resistance in the parasite. In the present paper we report the design, synthesis, and evaluation of potential antimalarial agents against a novel target, protein farnesyltransferase. We show that the most potent compounds are active against Plasmodium falciparum in vitro at submicromolar concentrations. PMID- 11277515 TI - Phenylcyclohexene and phenylcyclohexadiene substituted compounds having retinoid antagonist activity. AB - Retinoids are natural and synthetic analogues of the hormone retinoic acid. Systemic retinoid agonist therapy is usually associated with toxic side effects, such as mucocutaneous toxicity, which may be alleviated by the use of topical retinoid antagonists. We report the synthesis and biological activity of a new series of potent, RAR-specific antagonists substituted with phenylcyclohexene and phenylcyclohexadiene groups. PMID- 11277516 TI - Design, synthesis, and intracellular localization of a fluorescently labeled DNA binding polyamide related to the antibiotic distamycin. AB - The design and synthesis of the lipophilic (9) and fluorescent (10) conjugates of a structural analogue of distamycin and their in vitro cellular localization studies are reported. Confocal laser scanning microscopy (CLSM) indicates that 10 rapidly enters human ovarian adenocarcinoma (SKOV-3) cells with principal uptake in mitochondria and uniform cytoplasmic distribution. PMID- 11277517 TI - The synthesis of cyclic tetrapeptoid analogues of the antiprotozoal natural product apicidin. AB - A novel synthetic strategy is described which may be used to prepare analogues of the antimalarial, fungal metabolite apicidin. Compared to the natural product, one analogue shows potent and selective activity in vitro against the parasite Trypanosoma brucei and low mammalian cell toxicity. PMID- 11277518 TI - Synthesis of phenylglycine derivatives as potent and selective antagonists of group III metabotropic glutamate receptors. AB - The syntheses of a range of ring and alpha-substituted 4-phosphonophenylglycines are described. A brief discussion of the antagonist activities of compounds 4-10 on group I, II and III metabotropic glutamate (mGlu) receptors expressed in the neonatal rat spinal cord is included. PMID- 11277520 TI - Synthesis and characterization of bis(mu-hydroxo)diiron(III) complex of N-(4 nitro-2-hydroxy)phenylmethyl-N-(2-pyridylethyl)-N-(2-pyridylmethyl)amine and hydroxylation reaction of alkane. AB - Bis(mu-hydroxo)diiron(III) complex, [Fe2III(mu-OH)2(NE)2](NO3)2 x 3H2O (3) (N-(4 nitro-2-hydroxy)phenylmethyl-N-(2-pyridylethyl)-N-(2-pyridylmethyl)amine = HNE, where H denotes a dissociable proton of the p-nitrophenol group), has been prepared and characterized by X-ray crystallography, electronic and magnetic spectroscopies. PMID- 11277519 TI - NO-independent stimulators of soluble guanylate cyclase. AB - SARs around a novel type of guanylate cyclase stimulator which act by a mechanism different from classical NO-donors are described. Several pyrazolopyridinylpyrimidines are shown to relax aortic rings and revealed a long lasting blood pressure lowering effect in rats after oral application. PMID- 11277521 TI - Bisphosphonates derived from fatty acids are potent growth inhibitors of Trypanosoma cruzi. AB - We have investigated the effect of a series of bisphosphonates derived from fatty acids against Trypanosoma cruzi proliferation in in vitro assays. Some of these drugs proved to be potent inhibitors against the intracellular form of the parasite exhibiting IC50 values at the low micromolar level. As bisphosphonates are FDA clinically approved for treatment of bone resorption, their potential innocuousness makes them good candidates to control tropical diseases. PMID- 11277522 TI - A novel fluorinated tryptamine with highly potent serotonin 5-HT1A receptor agonist properties. AB - Synthesis and biological evaluation of a novel fluorinated tryptamine analogue are described. This new compound 1-(4-fluoro-5-methoxyindol-3-yl)pyrrolidine (2) was found to be a potent serotonin 5-HT1A agonist. PMID- 11277523 TI - Anti-MRSA cephems. Part 1: C-3 substituted thiopyridinium derivatives. AB - Sixteen novel cephalosporin derivatives with activity against methicillin resistant Staphylococcus aureus (MRSA) are described. The compounds were synthesized using substituted thiopyridones, generated either by cyclization of functionalized precursors, or by direct alkylation of the enolate of 2-methyl substituted pyrones. The most active compound in vitro against a strain of MRSA (A27223) displayed an MIC of 0.5 microg/mL. The most efficacious compound in vivo had a PD50 of 2.1 mg/kg. PMID- 11277524 TI - Discovery of zoniporide: a potent and selective sodium-hydrogen exchanger type 1 (NHE-1) inhibitor with high aqueous solubility. AB - Zoniporide (CP-597,396) is a potent and selective inhibitor of NHE-1, which exhibits high aqueous solubility and acceptable pharmacokinetics for intravenous administration. The discovery, synthesis, activities, and rat and dog pharmacokinetics of this compound are presented. The potency and selectivity of zoniporide may be due to the conformation that the molecule adopts due to the presence of a cyclopropyl and a 5-quinolinyl substituent on the central pyrazole ring of the molecule. PMID- 11277525 TI - Synthesis of metabolically blocked paclitaxel analogues. AB - The stereospecific syntheses of the metabolically blocked 6-alpha-F, Cl, Br paclitaxel, and 6-alpha-F-10-acetyldocetaxel are described and in vitro and in vivo activity is presented. PMID- 11277526 TI - Synthesis and antitumor activity of novel paclitaxel-chlorambucil hybrids. AB - The syntheses and antitumor activity of three paclitaxel-chlorambucil hybrids are presented. Hybrid 3 showed significant in vivo efficacy. PMID- 11277527 TI - Synthesis and use of FSCPX, an irreversible adenosine A1 antagonist, as a 'receptor knock-down' tool. AB - A new preparative synthetic route for the irreversible adenosine A1 antagonist 8 cyclopentyl-3-N-[3-((3-(4-fluorosulphonyl)benzoyl)-oxy)-propyl]-1-N-propyl xanthine (FSCPX, 1) is described. The availability of ample amounts of the irreversible antagonist FSCPX allowed us to use FSCPX as a research tool for adenosine A1 receptors in in vivo experiments. After verification of the irreversible antagonistic function of FSCPX in in vitro experiments, FSCPX was used successfully as a 'receptor knock-down' tool in in vivo experiments on conscious rats. PMID- 11277528 TI - Spatial requirements of the antagonist binding site of the NK2 receptor. AB - Computer-aided modelling has been used to identify a putative antagonist binding site in the tachykinin NK2 receptor. In order to validate the implied spatial requirements for this region, a series of compounds, based on the potent antagonist GR 149861 have been synthesised and their binding affinities established. Our findings suggest the presence of a large hydrophobic cavity in the putative binding crevice of GR 149861. PMID- 11277529 TI - Effects of two-carbon bridge region methoxylation of benztropine: discovery of novel chiral ligands for the dopamine transporter. AB - 6-Methoxylated and 8-oxygenated benztropines were prepared and evaluated for their DAT and SERT activity (binding and uptake inhibition). Methoxylation at the two-carbon bridge of benztropine produced a novel class of potent and selective DAT ligands. An interesting enantioselectivity was also observed for this new class of chiral benztropines. The inactivity of the 8-oxygenated analogues seems to point out that, unlike cocaine and its analogues, interactions of benztropine ligands with DAT may be strongly governed by the nitrogen atom. PMID- 11277530 TI - An efficient conversion of 5-nitroisatin into 5-nitroindole derivative. AB - Our process research on OPC-35564 revealed that a mixed borohydride reducing agent (ZrCl4/NaBH4) in DME (Itsuno system) afforded a rapid and direct conversion of N-alkyl-nitroisatin into nitroindole nucleus. Comparison with other reducing agents indicated the superiority of the present system and the key function of ZrCl4. For the manipulation of base-labile isatin, a useful procedure for its N alkylation using Cu2CO3 is also presented. PMID- 11277531 TI - Diaryl ester prodrugs of FR900098 with improved in vivo antimalarial activity. AB - The fosmidomycin derivative FR900098 represents an inhibitor of the 1-deoxy-D xylulose 5-phosphate (DOXP) reductoisomerase with potent antimalarial activity. Prodrugs of FR900098 with increased activity after oral administration were obtained by chemical modification of the phosphonate moiety to yield phosphodiaryl esters. One diaryl ester prodrug demonstrated efficacy in mice infected with the rodent malaria parasite Plasmodium vinckei comparable to i.p. drug administration. PMID- 11277532 TI - Structure elucidation and conformational properties of a novel bioactive clerodane diterpene using a combination of high field NMR spectroscopy, computational analysis and X-ray diffraction. AB - The structure of a novel CC clerodane type diterpenoid, namely (+)-19-acetoxy-cis clerodan-3-ene-15-oic acid 1 was elucidated and its conformational properties were explored using a combination of high field NMR spectroscopy and computational analysis. The structural analysis provided results consistent with those obtained by a single X-ray diffraction study of its dicyclohexylammonium salt. The new clerodane type diterpene isolated in large quantities from Cistus monspeliensis L. leaves was found to exhibit significant antibacterial activity against Staphyloccoci (MIC50 = 0.085 mM) and therefore represents a promising lead compound. Interestingly the deacetylated derivative 2 of compound 1 was inactive. PMID- 11277533 TI - Novel inhibitors of the enzyme estrone sulfatase (ES). AB - We report the initial results of our study into a series of simple 4-sulfamated phenyl alkyl ketones as potential inhibitors of the enzyme estrone sulfatase. The results of the study show that these compounds are potent inhibitors, possessing greater inhibitory activity than COUMATE, but weaker activity than EMATE. Furthermore, the compounds are observed to be irreversible inhibitors. PMID- 11277534 TI - N-Aminoindoline derivatives as inhibitors of 5-lipoxygenase. AB - N-Aminoindoline derivatives were prepared and their 5-lipoxygenase inhibitory activities were evaluated in vitro and compared with those of phenidone and NDGA. Compound 4 presents the most effective 5-LO inhibition. PMID- 11277535 TI - 7-Alkyl- and 7-cycloalkyl-5-aryl-pyrrolo[2,3-d]pyrimidines--potent inhibitors of the tyrosine kinase c-Src. AB - 7-Substituted-5-aryl-pyrrolo[2,3-d]pyrimidines have been prepared starting from alpha-bromoacetophenones. These compounds represent a novel class of potent inhibitors of the tyrosine kinase pp60(c-Src) with good specificity towards other tyrosine kinases (EGF-R, v-Abl). PMID- 11277536 TI - 7-Pyrrolidinyl- and 7-piperidinyl-5-aryl-pyrrolo[2,3-d]pyrimidines--potent inhibitors of the tyrosine kinase c-Src. AB - 7-Heterocyclyl-5-aryl-pyrrolo[2,3-d]pyrimidines represent a new class of highly potent and selective inhibitors of the tyrosine kinase pp60(c-Src). PMID- 11277537 TI - Access to the inaccessible sequence of cpn 60.1 (195-217) by temporary oxazolidine protection of selected amide bonds. AB - The solid-phase peptide synthesis of a reportedly inaccessible peptide sequence of chaperonin 60.1 (195-219) is described using oxazolidine containing dipeptide building blocks ('pseudo-proline' dipeptide units). Two attempts at the synthesis of the chaperonin 60.1 sequence are outlined using one and two pseudo-proline units, respectively, and these results are compared with the outcome of an ordinary stepwise (double) coupling procedure. The only successful synthesis is that combining two pseudo-proline building blocks. PMID- 11277538 TI - Synthesis and in vitro cytotoxicity of novel long chain busulphan analogues. AB - Two novel long chain alkanediol dimethanesulphonates, analogues of busulphan, were synthesized. Their in vitro cytotoxicity was evaluated against six solid tumor cell lines (A2780, H322, LL, WiDr, C26-10 and UMSCC-22B). 2 Tetradecylbutane-1,4-diol dimethanesulphonate was proved to be the most active compound exhibiting IC50 values between 20.82 and 26.36 microM. PMID- 11277540 TI - Cancer screening in perspective. PMID- 11277539 TI - Controlling antibiotic resistance: will we someday see limited prescribing autonomy? PMID- 11277541 TI - The patient safety grid: toxic cascades in health care settings. PMID- 11277542 TI - Restless legs syndrome. PMID- 11277543 TI - Role of Papanicolaou tests. PMID- 11277544 TI - Counseling to reduce noise-induced hearing loss. PMID- 11277545 TI - Noise-induced hearing loss in shooters of shoulder firearms. PMID- 11277547 TI - Cancer screening guidelines. AB - Numerous medical organizations have developed cancer screening guidelines. Faced with the broad, and sometimes conflicting, range of recommendations for cancer screening, family physicians must determine the most reasonable and up-to-date method of screening. Major medical organizations have generally achieved consensus on screening guidelines for breast, cervical and colorectal cancer. For breast cancer screening in women ages 50 to 70, clinical breast examination and mammography are generally recommended every one or two years, depending on the medical organization. For cervical cancer screening, most organizations recommend a Papanicolaou test and pelvic examination at least every three years in patients between 20 and 65 years of age. Annual fecal occult blood testing along with flexible sigmoidoscopy at five-year to 10-year intervals is the standard recommendation for colorectal cancer screening in patients older than 50 years. Screening for prostate cancer remains a matter of debate. Some organizations recommend digital rectal examination and a serum prostate-specific antigen test for men older than 50 years, while others do not. In the absence of compelling evidence to indicate a high risk of endometrial cancer, lung cancer, oral cancer and ovarian cancer, almost no medical organizations have developed cancer screening guidelines for these types of cancer. PMID- 11277548 TI - Acute and chronic paronychia. AB - Paronychia is one of the most common infections of the hand. Clinically, paronychia presents as an acute or a chronic condition. It is a localized, superficial infection or abscess of the paronychial tissues of the hands or, less commonly, the feet. Any disruption of the seal between the proximal nail fold and the nail plate can cause acute infections of the eponychial space by providing a portal of entry for bacteria. Treatment options for acute paronychias include warm-water soaks, oral antibiotic therapy and surgical drainage. In cases of chronic paronychia, it is important that the patient avoid possible irritants. Treatment options include the use of topical antifungal agents and steroids, and surgical intervention. Patients with chronic paronychias that are unresponsive to therapy should be checked for unusual causes, such as malignancy. PMID- 11277549 TI - Osteoporosis: part II. Nonpharmacologic and pharmacologic treatment. AB - Family physicians will frequently encounter patients with osteoporosis, a condition that is often asymptomatic until a fracture occurs. Treatment of the fracture can be initiated without further diagnostic testing. Thereafter, treatment of osteoporosis includes (1) prevention of further bone loss through weight-bearing exercise, tobacco and alcohol avoidance, hormone replacement therapy in women, and raloxifene and calcium supplementation; (2) treatment of fracture-related pain with analgesics and calcitonin; (3) building bone mass when feasible with alendronate; and (4) modifying behaviors that increase the risk of falls. Patients without fracture who are at risk for osteoporosis can also benefit from these preventive measures. Furthermore, women of all ages should be encouraged to maintain a daily calcium intake of 1,000 to 1,500 mg and to participate in weight-bearing exercise for 30 minutes three times weekly to reduce their risk of falls and fractures. Persons at risk should avoid medications known to compromise bone density, such as glucocorticoids, thyroid hormones and chronic heparin therapy. PMID- 11277546 TI - Antimicrobial resistance: a plan of action for community practice. AB - Antibiotic resistance was once confined primarily to hospitals but is becoming increasingly prevalent in family practice settings, making daily therapeutic decisions more challenging. Recent reports of pediatric deaths and illnesses in communities in the United States have raised concerns about the implications and future of antibiotic resistance. Because 20 percent to 50 percent of antibiotic prescriptions in community settings are believed to be unnecessary, primary care physicians must adjust their prescribing behaviors to ensure that the crisis does not worsen. Clinicians should not accommodate patient demands for unnecessary antibiotics and should take steps to educate patients about the prudent use of these drugs. Prescriptions for targeted-spectrum antibiotics, when appropriate, can help preserve the normal susceptible flora. Antimicrobials intended for the treatment of bacterial infections should not be used to manage viral illnesses. Local resistance trends may be used to guide prescribing decisions. PMID- 11277550 TI - Endometrial biopsy. AB - Endometrial biopsy is an office procedure that serves as a helpful tool in diagnosing various uterine abnormalities. The technique is fairly easy to learn and may be performed without assistance. The biopsy is obtained through the use of an endometrial suction catheter that is inserted through the cervix into the uterine cavity. Twirling the catheter while moving it in and out of the uterine cavity enhances uptake of uterine tissue, which is aspirated into the catheter and removed. Endometrial biopsy is useful in the work-up of abnormal uterine bleeding, cancer screening, endometrial dating and infertility evaluation. Contraindications to the procedure include pregnancy, acute pelvic inflammatory disease, and acute cervical or vaginal infections. Postoperative infection is rare but may be further prevented through the use of prophylactic antibiotic therapy. Intraoperative and postoperative cramping are frequent side effects. PMID- 11277551 TI - Asymptomatic microscopic hematuria in adults: summary of the AUA best practice policy recommendations. AB - The American Urological Association (AUA) convened the Best Practice Policy Panel on Asymptomatic Microscopic Hematuria to formulate policy statements and recommendations for the evaluation of asymptomatic microhematuria in adults. The recommended definition of microscopic hematuria is three or more red blood cells per high-power microscopic field in urinary sediment from two of three properly collected urinalysis specimens. This definition accounts for some degree of hematuria in normal patients, as well as the intermittent nature of hematuria in patients with urologic malignancies. Asymptomatic microscopic hematuria has causes ranging from minor findings that do not require treatment to highly significant, life-threatening lesions. Therefore, the AUA recommends that an appropriate renal or urologic evaluation be performed in all patients with asymptomatic microscopic hematuria who are at risk for urologic disease or primary renal disease. At this time, there is no consensus on when to test for microscopic hematuria in the primary care setting, and screening is not addressed in this report. However, the AUA report suggests that the patient's history and physical examination should help the physician decide whether testing is appropriate. PMID- 11277552 TI - Insulin resistance syndrome. AB - Insulin resistance can be linked to diabetes, hypertension, dyslipidemia, cardiovascular disease and other abnormalities. These abnormalities constitute the insulin resistance syndrome. Because resistance usually develops long before these diseases appear, identifying and treating insulin-resistant patients has potentially great preventive value. Insulin resistance should be suspected in patients with a history of diabetes in first-degree relatives; patients with a personal history of gestational diabetes, polycystic ovary syndrome or impaired glucose tolerance; and obese patients, particularly those with abdominal obesity. Present treatment consists of sensible lifestyle changes, including weight loss to attain healthy body weight, 30 minutes of accumulated moderate-intensity physical activity per day and increased dietary fiber intake. Pharmacotherapy is not currently recommended for patients with isolated insulin resistance. PMID- 11277553 TI - Asthma medications. PMID- 11277554 TI - Asthma: taking medicines safely. PMID- 11277555 TI - Outdoor air pollution: possible health effects. PMID- 11277556 TI - AAP technical report on the prevention of pneumococcal infections. American Academy of Pediatrics. PMID- 11277557 TI - The influence of previous experience on predictive motor control. AB - Anticipating the consequences of our motor commands is a fundamental component of sensorimotor control. For example, when one hand pulls on an object held in the other, the restraining hand generates an anticipatory increase in grip force thereby preventing the object from slipping. To investigate how such anticipation is learned subjects held an object, whose properties were under computer control, between their hands. This allowed instant changes in the behaviour of the manipulated object on a trial by trial basis. The extent of grip force modulation seen in one hand, when the other pulled on the object was found to depend in a systematic way on the object's properties experienced over at least the previous three trials. PMID- 11277558 TI - Contrast response characteristics of long-range lateral interactions in cat striate cortex. AB - Single-cell responses in visual cortex to a target falling within their receptive field can be modified by collinear flanking stimuli concurrently presented outside the receptive field. Here, we report the presence of four types of contrast-dependent lateral effects: (1) facilitation at low target contrasts and suppression at high contrasts, (2) facilitation that increases with contrast, (3) suppression that increases with contrast, and (4) suppression at low contrasts with facilitation at high contrasts. We propose a sensitivity modulation model that accounts for all the four types of lateral effects by changes in two parameters. In this model, activation of neighboring neurons changes the sensitivities of the target neuron to both the direct feedforward input and inhibitory, divisive feedback from neighboring neurons. PMID- 11277559 TI - NMDA-mediated social learning of fear-induced conditioned analgesia to biting flies. AB - Although fear conditioning has received extensive neurobiological attention little is known about social learning whereby one individual may learn and acquire the fear responses of another. A 30 min exposure to intact biting flies (stable fly, Stomoxys colcitrans L.) elicits in individual fly-naive mice analgesia and active self burying responses to avoid the flies. Fly-naive observer mice that witnessed other demonstrator mice being attacked by biting flies exhibited analgesia and self-burying to avoid flies when exposed 24 h later to altered flies whose biting mouth parts were removed. The opiate antagonist naloxone, while reducing the analgesic responses elicited by exposure to a fly stressed demonstrator, did not affect either the subsequent conditioned analgesia or self-burying. However, the specific NMDA receptor antagonist NPC 12626, given to observers prior to, but not after, presentation of fly attacked demonstrators blocked the socially determined conditioned analgesia and self burying avoidance. This supports NMDA involvement in the mediation of the social transmission and long-term (24h) retention of conditioned analgesia and fear. PMID- 11277560 TI - Serotonergic nuclei of the raphe are not affected in human ageing. AB - Sleep disorders increase with ageing. The serotonergic system has been linked with sleep regulation. In fatal familial insomnia, a prion disease with insomnia as one major clinical feature, we recently observed a disturbance in the serotonergic system as likely substrate of typical symptoms. Using immunohistochemistry for the serotonin synthesizing enzyme, tryptophan hydroxylase, we investigated the serotonergic median raphe nuclei (dorsal raphe nucleus, superior central nucleus, and raphe obscurus nucleus) in brains of an older (n = 12; age range 62-84 years) and a younger group (n = 10; age range 5-29 years). We found no significant difference between age groups in the percentage of neurons able to synthesize serotonin. Other changes might relate to sleep disturbances in the elderly. PMID- 11277561 TI - NGF enhances depolarization effects on SNAP-25 expression: induction of SNAP-25b isoform. AB - The 25 kDa synaptosomal associated protein (SNAP-25), which is implicated in neuronal plasticity and neurosecretion, exists as two isoforms generated by alternative splicing of exons 5a and 5b. The aim of the present study was to characterize factors influencing isoform expression. We report that chronic depolarization of PC12 cells alone or in the presence of NGF induces the expression of isoform-b, in addition to a 1.8- to 3-fold increase in SNAP-25 mRNA and protein as determined by immunoblotting and combined RT-PCR and Southern blot analysis. When cerebellar granule neurons were cultured in elevated K+, the predominant isoform switched from SNAP-25a to SNAP-25b. Taken together these results suggested that chronic depolarization regulates the transcription and processing of SNAP-25 mRNA. PMID- 11277562 TI - Diadenosine pentaphosphate is more potent than ATP at P2X receptors in isolated rat vagus nerve. AB - The effects of diadenosine pentaphosphate (Ap5A), diadenosine tetraphosphate (Ap4A), alpha,beta-methyleneATP (alpha,beta-meATP), and ATP were studied on the excitability of unmyelinated axons in isolated rat vagus nerve by means of a computerized threshold tracking technique. All purinergic compounds produced an increase in excitability, however, only the effects of alpha,beta-meATP and of Ap5A were strongly reduced by 2'- (or 3') -O-(2,4,6-trinitrophenyl)-ATP (TNP ATP), a selective blocker for P2X1, P2X3, and heteromeric P2X2/3 receptors. The rank order of potency for TNP-ATP-sensitive excitation was determined as follows (30 microM each): alpha,beta-meATP >Ap5A >> Ap4A = ATP. These data suggest that Ap5A might be an important naturally occurring agonist for P2X receptors at the axonal membrane of unmyelinated, including nociceptive, nerve fibres. PMID- 11277563 TI - Entorhinal cortex disruption causes memory deficit in early Alzheimer's disease as shown by PET. AB - Voxel-based mapping of the correlations between cognitive scores and resting state brain glucose utilization measured by PET has recently emerged as a novel way to reveal in living patients with Alzheimer's disease (AD) the neural systems whose disruption underlies particular neuropsychological, especially mnemonic, deficits. We have now applied this approach using a novel cognitive paradigm designed to selectively assess verbal episodic memory, and show that in early AD disruption of the left entorhinal cortex underlies this memory deficit, consistent with post mortem data showing that this brain area is affected earliest and most severely by tau pathology in AD. PMID- 11277564 TI - Agouti-related peptide prevents steroid-induced luteinizing hormone and prolactin surges in female rats. AB - We studied the effect of Agrp (agouti-related peptide) on LH (luteinizing hormone) and PRL (prolactin) surges in ovariectomized rats primed with estradiol and progesterone. The rats displayed characteristic LH and PRL surges that were completely abolished by starving. Injection of either 1 nmol or 3 nmol Agrp (83 132), a potent antagonist of the orexigenic MC3 and MC4 receptors, completely prevented both the LH and PRL surges. We also investigated the effects of either a single or double injection of anti-Agrp serum to fasted animals, which were without LH and PRL surges. A single injection of the antiserum was without effect, but the rats that received double injection of anti-Agrp serum partially reinstated both the LH and PRL surges. Although the onset of LH and PRL surges was significantly delayed in the double treated group, the highest levels of the surges for both hormones were statistically indistinguishable compared with the control group. These data give a clear indication that endogenous Agrp may be involved in LH and PRL surges during starvation, providing further evidence that the melanocortin system is important for these hormonal surges in female rats. PMID- 11277565 TI - Increased serum S100beta protein concentrations following severe head injury in humans: a biochemical marker of brain death? AB - This study investigated S100beta protein as a biochemical serum marker of brain damage in severe head injury and brain death victims. Blood samples obtained from 15 patients with severe head injury admitted to the trauma intensive care unit (ICU), five patients with a diagnosis of brain death due to hemorrhage following cerebral aneurysm rupture, and five healthy individuals were investigated. The S100beta protein serum concentrations were analyzed with a immunoradiometric assay kit. The 15 patients with severe head injury were followed up for 6 months. Outcome was considered either death or recovery with ICU discharge. S100beta concentrations were closely related to brain damage. Among the severe head injury victims, higher S100beta concentrations were detected in those patients that progressed to death. The individuals with brain death had similar mean S100beta concentrations, irrespective of its cause (either trauma or vascular rupture). S100beta protein is a promising serum outcome predictor for severe head injury victims and may contribute to the early diagnosis of brain death. PMID- 11277566 TI - Involvement of apolipoprotein E in herpes simplex encephalitis. AB - APOE polymorphism may influence risk for cold sores and, by interacting with latent HSV-1, risk for Alzheimer's disease (AD). APOE genotype also influences outcome after brain injury. We sought to determine whether APOE genotype influences risk for herpes simplex encephalitis (HSE), whether apoE is involved in the response to HSE and if APOE genotype influences outcome from HSE. There was increased immunoreactivity of neurons, neuropil and glia for apoE areas of brain damaged by HSE. APOE genotypes for cases of HSE (n = 57) were similar to those of controls (n = 41). APOE genotypes for survivors of HSE were similar to those of patients who died. We conclude that apoE is involved in the response to damage associated with HSE, as in other forms of brain injury. However, APOE genotype does not appear to influence either the risk of developing HSE or subsequent mortality. PMID- 11277567 TI - Orphanin FQ: an endogenous antagonist of rat brain dopamine transporter. AB - Orphanin FQ, also known as nociceptin (NC),is a well-known ligand for opioid receptor-like ORLI receptor. This heptadecapeptide was identified as potently inhibiting the uptake of rat dopamine transporter (rDAT) which is stably expressed in CHO cells (designated D8 cells). Further kinetic analysis proved that this occurs through competitive inhibition with an IC50 of about 1.9 microM. Orphanin FQ also inhibits [3H]dopamine uptake by rat striatal synaptosomes, which confirmed the effect of orphanin FQ on D8 cells. Orphanin FQ was also found to inhibit GABA transporter type I (GATI) but not the serotonin transporter. These results suggest that orphanin FQ is an endogenous antagonist of dopamine transport and that it affects locomotion and other activities at least partly by inhibiting dopamine transporter and directly affecting dopamine transmission or by inhibiting GABA transporter to indirectly change dopaimne transmission. PMID- 11277569 TI - Emotional expression boosts early visual processing of the face: ERP recording and its decomposition by independent component analysis. AB - To investigate the hypothesis that early visual processing of stimuli might be boosted by signals of emotionality, we analyzed event related potentials (ERPs) of twelve right-handed normal subjects. Gray-scale still images of faces with emotional (fearful and happy) or neutral expressions were presented randomly while the subjects performed gender discrimination of the faces. The results demonstrated that the faces with emotion (both fear and happiness) elicited a larger negative peak at about 270 ms (N270) over the posterior temporal areas, covering a broad range of posterior visual areas. The result of independent component analysis (ICA) on the ERP data suggested that this posterior N270 had a synchronized positive activity at the frontal-midline electrode. These findings confirm that the emotional signal boosts early visual processing of the stimuli. This enhanced activity might be implemented by the amygdalar re-entrant projections. PMID- 11277568 TI - Task-related EEG-EEG coherence depends on dopaminergic activity in Parkinson's disease. AB - We investigated whether functional coupling between different cortical areas is impaired in Parkinson's disease, using corticocortical coherence as a surrogate measure of coupling. We recorded scalp EEG from different sites in seven parkinsonian patients while they tracked a visual target using their wrist, or copied the same movement from memory. Differences in EEG-EEG coherence between the tracking and copying tasks and their respective controls, visual tracking alone and fixation of a stationary target, were determined on and off levodopa. After levodopa we found extensive task-specific and broad band cortico-cortical coherence. Off levodopa cortico-cortical coherence was much reduced. Ascending dopaminergic projections from the ventral mesencephalon may therefore be important in determining the pattern and extent of corticocortical coupling during executive tasks. PMID- 11277570 TI - Peripheral PAR-2 triggers thermal hyperalgesia and nociceptive responses in rats. AB - Protease-activated receptor-2 (PAR-2), a member of the G protein-coupled, seven trans-membrane domain receptor family, is activated by trypsin/tryptase and present in various tissues including the primary sensory neurons, playing a role in development of neurogenic inflammation. The present study examined if activation of peripheral PAR-2 could modulate nociception in the rat. Expression of mRNA for PAR-2 was confirmed in the L4-6 dorsal root ganglia, but not spinal cord. The PAR-2-activating peptide SLIGRL-NH2 administered by the intraplantar (i.pl.) route, produced thermal, but not mechanical, hyperalgesia in the rat, although the PAR-2-inactive control peptide LSIGRL-NH2 had no effect. Not only the PAR-2-activating but also inactive peptides elicited nociceptive behavior (licking/biting) in the intact rats, whereas only the former peptide produced such behavior in the rats that had received repeated administration of compound 48/80 for mast cell depletion. These data provide novel evidence that activation of peripheral PAR-2 is pro-nociceptive, producing thermal hyperalgesia and also triggering pain sensation, by itself, independently of mast cell degranulation. PMID- 11277571 TI - Mitochondrial cytochrome c oxidase subunit III is selectively down-regulated by aluminum exposure in PC12S cells. AB - Aluminum (Al) has been implicated in several neurological diseases including dialysis dementia and Alzheimer's disease (AD). One possible mechanism of Al neurotoxicity could involve alteration of mitochondrial gene expression. We exposed PC12 cells to 0.1-100 microM AlCl3 for 6h at pH 7.4. Internalized Al, measured by atomic absorption spectrometry, was linearly proportional to the extracellular Al concentration. Northern blot analyses showed that cytochrome c oxidase subunit III (COX III) mRNA was significantly reduced by 70% after addition of 1 microM AlCl3. Higher concentrations of AlCl3 did not show a significant further effect. These results suggest that Al neurotoxicity involves a specific impairment of cytochrome c oxidase. PMID- 11277572 TI - The distribution of neuronal calcium sensor-1 protein in the developing and adult rat retina. AB - We have examined the distribution of the neuronal calcium-binding protein, neuronal calcium sensor 1 (NCS-1) in the developing and adult rat retina using subcellular fractionation of the rat retina and immunohistochemistry. NCS-1 immunoreactivity was situated primarily in the ganglion cells, a class of amacrine cells, and in the inner plexiform layer (IPL). During development, NCS-1 protein expression closely followed that of the synaptic vesicle protein, synaptophysin, increasing dramatically in the IPL at postnatal day 3, the time when conventional synapses are formed in the retina. These findings suggest that NCS-1 plays a role in synaptogenesis in the retina and in synaptic transmission at conventional synapses but not ribbon synapses in the adult rat retina. PMID- 11277573 TI - Effect on human attention of exposure to the electromagnetic field emitted by mobile phones. AB - This study examined the effect of exposure to the electromagnetic field emitted by mobile phones on human attention. Three measures of attention were administered to 72 teenagers, 37 of whom were mobile phone users. The results showed that the mobile phone users performed better on one of the three measures of attention than did the non-mobile phone users. The results suggest that exposure to the electromagnetic field emitted by mobile phones may have a mild facilitating effect on attention functions, which is consistent with previous observations that exposure to the electromagnetic field has a facilitating effect on cognitive processing. The possibility that mobile phone users may be naturally better at multiple tasking tasks was discussed. PMID- 11277574 TI - Retrograde degeneration of neurite membrane structural integrity of nerve growth cones following in vitro exposure to mercury. AB - Inhalation of mercury vapor (Hg0) inhibits binding of GTP to rat brain tubulin, thereby inhibiting tubulin polymerization into microtubules. A similar molecular lesion has also been observed in 80% of brains from patients with Alzheimer disease (AD) compared to age-matched controls. However the precise site and mode of action of Hg ions remain illusive. Therefore, the present study examined whether Hg ions could affect membrane dynamics of neurite growth cone morphology and behavior. Since tubulin is a highly conserved cytoskeletal protein in both vertebrates and invertebrates, we hypothesized that growth cones from animal species could be highly susceptible to Hg ions. To test this possibility, the identified, large Pedal A (PeA) neurons from the central ring ganglia of the snail Lymnoea stagnalis were cultured for 48 h in 2 ml brain conditioned medium (CM). Following neurite outgrowth, metal chloride solution (2 microl) of Hg, Al, Pb, Cd, or Mn (10(-7) M) was pressure applied directly onto individual growth cones. Time-lapse images with inverted microscopy were acquired prior to, during, and after the metal ion exposure. We demonstrate that Hg ions markedly disrupted membrane structure and linear growth rates of imaged neurites in 77% of all nerve growth cones. When growth cones were stained with antibodies specific for both tubulin and actin, it was the tubulin/microtubule structure that disintegrated following Hg exposure. Moreover, some denuded neurites were also observed to form neurofibrillary aggregates. In contrast, growth cone exposure to other metal ions did not effect growth cone morphology, nor was their motility rate compromised. To determine the growth suppressive effects of Hg ions on neuronal sprouting, cells were cultured either in the presence or absence of Hg ions. We found that in the presence of Hg ions, neuronal somata failed to sprout, whereas other metalic ions did not effect growth patterns of cultured PeA cells. We conclude that this visual evidence and previous biochemical data strongly implicate Hg as a potential etiological factor in neurodegeneration. PMID- 11277575 TI - The expression of calcitonin gene-related peptide in dorsal horn neurons of the mouse lumbar spinal cord. AB - Using immunohistochemistry and in situ hybridization the expression of calcitonin gene-related peptide (CGRP) was studied in the mouse spinal cord under normal conditions and after unilateral rhizotomy and after local colchicine treatment. Under normal conditions a dense plexus of CGRP-immunoreactive (IR) fibres was observed in the superfical layers of dorsal horns with lower numbers of fibers in deeper laminae. Seven days after unilateral rhizotomy, there was a marked reduction of CGRP-IR fibres in the ipsilateral superfical layers and distinctly CGRP-IR neurons could be detected in the ipsilateral lamina III. CGRP mRNA positive neurons were observed in lamina III in both the ipsilateral and contralateral dorsal horn. Colchicine treatment did not markedly increase the number of CGRP-IR neurons. The results suggest that CGRP is synthesized in local dorsal horn neurons of the mouse, and these neurons presumably participate in sensory processing. PMID- 11277576 TI - Extension of glial processes by activation of Ca2+-permeable AMPA receptor channels. AB - AMPA type-glutamate receptor channels (AMPARs) assembled without the GluR2 (GluR B) subunit are characterized by high Ca2+ permeability, and are expressed abundantly in cerebellar Bergmann glial cells. Here we show that the morphology of cultured Bergmann glia-like fusiform cells derived from the rat cerebellum was changed by manipulating expression of Ca2+-permeable AMPARs using adenoviral vector-mediated gene transfer. Converting endogenous Ca2+-permeable AMPARs into Ca2+-impermeable channels by viral-mediated transfer of GluR2 gene induced retraction of glial processes. In contrast, overexpression of Ca2+-permeable AMPARs markedly elongated glial processes. The process extension was blocked by 2,3-Dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX), a specific antagonist of AMPAR. These results indicate that glutamate regulates the morphology of glial processes by activating Ca2+-permeable AMPARs. PMID- 11277577 TI - Tetrahydrocannabinol (THC) interferes with conditioned retching in Suncus murinus: an animal model of anticipatory nausea and vomiting (ANV). AB - Little is understood about effective countermeasures to the expression of anticipatory nausea and vomiting (ANV) in chemotherapy patients. We present a model of ANV based on the emetic reactions of the Suncus murinus (musk shrew). Following two pairings of a novel distinctive contextual cue with the emetic effects of an injection of lithium chloride, the context acquired the potential to elicit retching in the absence of the toxin. The expression of this conditioned retching reaction was completely suppressed by pretreatment with THC at a dose that did not suppress general activity. This provides the first experimental evidence in support of anecdotal reports that THC suppresses ANV. PMID- 11277578 TI - Reversible deafferentation of abducens motoneurons and internuclear neurons with tetanus neurotoxin. AB - Tetanus neurotoxin (TeNT) is a blocker of synaptic vesicle exocytosis in central synapses with preferential affinity for inhibitory neurotransmission. Following its intramuscular injection, TeNT is retrogradely and trans-synaptically transported towards the premotor terminals. Therefore, we have used TeNT as a tool to study the consequences of functional deafferentation on motoneurons following its peripheral administration. For this, we injected the toxin into the lateral rectus muscle at doses of 5 or 0.5 ng/kg and recorded the discharge activity of abducens motoneurons and internuclear neurons in the alert cat. Our results showed that: (i) TeNT blocked selectively the afferent inhibitory signals on abducens neurons only when used at a low dose, whereas both excitatory and inhibitory synaptic drive was lost after the high dose treatment; (ii) all effects were reversible within one month; and (iii) strikingly, the internuclear neurons of the abducens nucleus showed similar discharge alterations to the motoneurons, suggesting a TeNT action on shared common afferences. PMID- 11277579 TI - Edg2 receptor distribution in adult rat brain. AB - The Edg (endothelial differentiation gene) receptors are recently discovered G protein coupled receptors which are activated by endogenous lysophospholipids. The cellular activities mediated by Edg receptors are reminiscent of those normally associated with Trk receptor activation and include modulation of cell growth, differentiation, proliferation and migration as well as apoptotic and cytoskeletal effects. In this study we have investigated immunohistochemically the distribution of one family member, the Edg2 receptor, within the adult rat brain and shown the protein expression to be most prominent in white matter tract regions. This suggests a possible role for the Edg2 receptor in nerve cell myelination. PMID- 11277580 TI - Expression of monocarboxylate transporter MCT1 in normal and neoplastic human CNS tissues. AB - Expression of monocarboxylate transporter MCT1 was studied in archival tissues from human CNS using antibodies to the carboxyl-terminal end of MCT1. Sections of neocortex, hippocampus and cerebellum of brains from 10 adult autopsy patients who died from other than CNS disease, and from archival surgical biopsy specimens of 83 primary CNS and eight non-CNS tumors were studied. MCT1 immunoreactivity was present in microvessels and, ependymocytes of normal CNS tissues similar to that reported for MCT1 expression in rat brains. MCT1 immunoreactivity was strongest in ependymomas, hemangioblastomas and high grade glial neoplasms, and weakest in low grade gliomas. Increased MCT1 expression in high grade glial neoplasms may provide a potential therapeutic target for treatment of some CNS neoplasms. PMID- 11277581 TI - N100 evoked potential latency variation and startle in schizophrenia. AB - The contribution of evoked potential (EP) latency jitter, a measure of CNS temporal variability, on startle and EP gating defects in schizophrenic subjects has not been characterized. The amplitude of the N100/P200 EP complex (peak to trough) derived using a time-locked averaging procedure, N100 EP latency jitter derived from single trial analysis, acoustic startle response and clinical symptoms were measured in 51 schizophrenic subjects. N100 latency jitter was inversely correlated with N100/P200 EP amplitude in both cross-sectional and longitudinal analysis. Subjects with elevated EP gating ratios (>0.5) had similar latency jitter values for initial (S1) and test (S2) stimuli, while subjects with a low gating ratio (0-0.5) had a lower level of S1 latency jitter. Temporal variability thus plays a significant and complex role in previously reported sensory gating deficits in schizophrenic subjects. PMID- 11277582 TI - Cell migration from the corticostriatal angle to the basal telencephalon in rat embryos. AB - Lateral cortical stream (LCS) has been described as a flow of cell migration found in the lateral cortex of the embryonic telencephalon of mammals. The destinations of the cell migration were reported as the ventrolateral neocortex, the pyriform cortex, endopyriform nucleus and the claustrum. At the same time, however, other destinations of LCS have been suggested in the ventral telencephalon. Therefore, we investigated the additional destinations of the LCS using a combination of several molecular biological techniques. Using an expression vector of modified green fluorescent protein (GFP) introduced into the ventricular zone (VZ) around the corticostriatal angle, both tangential and radial cell migration was revealed in the LCS. The radial cell migration in the LCS supplied cells to the ventro-lateral neocortex, pyriform cortex and to the level of the lateral olfactory tract. In the second experiment, we injected COS-1 cells transfected with a Sema3A expression vector into one side of the neocortex. The cell supply to the destination of the LCS ceased due to the formation of a large necrosis in the LCS, which was triggered by the Sema3A-COS cell injection, and the dense cell layer in the olfactory tubercle shrunk on the side where COS cells were injected. These data indicated that the majority of neurons in the dense cell layer of the olfactory tubercle reached this point through the LCS. One of the origins of the cells in the LCS would be in the corticostriatal angle. PMID- 11277583 TI - Circadian rhythm in NO synthase I transcript expression and its photoperiodic regulation in the rat pineal gland. AB - The photoneural regulation of nitric oxide synthase type I (NOS I) expression in the rat pineal was investigated using semiquantitative RT-PCR. NOS I transcript expression exhibited a daily rhythm with peak values during the night hours. The daily rhythm in NOS I transcript expression persisted under constant dark conditions and was abolished under constant light conditions. The extent of nocturnal NOS I expression was found to be dependent on the photoperiod. It was attenuated under 20 h light and 4 h dark (L:D 20:4) compared with 12 h light and 12 h dark (L:D 12:12). The present findings indicate that, in the rat pineal, NOS I transcript expression exhibits a true circadian rhythm. Further, photoperiod induced changes in circadian transcript expression appear to be responsible for reported changes in NOS I protein expression. PMID- 11277584 TI - Degeneration of nociceptive nerve terminals in human peripheral neuropathy. AB - Patients with peripheral neuropathy have symptoms involving small-diameter nociceptive nerves and elevated thermal thresholds. Nociceptive nerves terminate in the epidermis of the skin and are readily demonstrated with the neuronal marker, protein gene product 9.5 (PGP 9.5). To investigate the pathological characteristics of elevated thermal thresholds, we performed PGP 9.5 immunocytochemistry on 3 mm punch skin biopsies (the forearm and the leg) from 55 normal subjects and 35 neuropathic patients. Skin innervation was evaluated by quantifying epidermal nerve densities. Epidermal nerve densities were reduced in neuropathic patients compared to normal subjects. Epidermal nerve densities were variably correlated with thermal thresholds. The proportion of neuropathic patients with reduced epidermal nerve densities was larger than the proportion of neuropathic patients with elevated thermal thresholds. These results indicated that degeneration of epidermal nerve terminals preceded the elevation of thermal thresholds. Skin biopsy together with immunocytochemical demonstration of epidermal innervation offers a new approach to evaluate small-fiber sensory neuropathy. PMID- 11277585 TI - DYT1 mutation in Japanese patients with primary torsion dystonia. AB - A GAG deletion at position 946 in the DYT1 gene has been identified as one of the gene mutations responsible for autosomal dominant primary torsion dystonia. We examined 178 Japanese patients with various forms of dystonia, and found the mutation in six patients (3.4%) from three families. Five of them had early clinical onset (before age 12) with initial involvement of a limb. To our knowledge, this is the first report of the frequency and the clinical features of DYT1 mutation in oriental patients, and the clinical presentation of the mutation in these patients was similar to that of Jewish or non-Jewish Caucasian patients. PMID- 11277586 TI - Modulation of visceral nociceptive responses of rat spinal dorsal horn neurons by sympathectomy. AB - We determined whether sympathectomy modulates visceral nociception under physiological or inflammatory conditions. Recordings of sacral spinal dorsal horn neurons with sustained responses were performed in pentobarbitone-anesthetized rats. Graded colorectal distension (CRD, 20-100 mmHg) was used as a visceral nociceptive stimulus. Inflammation was induced by intracolonic instillation of turpentine (25%). Sympathectomy was produced by administering 6-hydroxydopamine. Inflammation produced an increase in the CRD-evoked responses. The CRD-evoked responses were attenuated following sympathectomy both under control and inflammatory conditions. These changes in the CRD-evoked responses were associated with corresponding changes in spontaneous discharge rate. The convergent input evoked by noxious pinch of the skin was not changed by any of the experimental conditions. The results indicate that sympathectomy attenuates visceral nociceptive responses and spontaneous activity of sacral spinal cord neurons, without effect on convergent cutaneous inputs, both under physiological and inflammatory conditions. PMID- 11277587 TI - Dopamine modulates sodium currents in cochlear spiral ganglion neurons. AB - Lateral olivocochlear (LOC) efferent neurons, putatively dopaminergic, synapse on afferent dendrites of type I spiral ganglion neurons (SGNs) in the cochlea and depress their activity. To investigate the underlying mechanisms, whole-cell patch clamp recordings were obtained from mouse SGNs. Dopamine (DA), and D1-like (D1, D5) and D2-like (D2, D3 and D4) receptor agonists, reduced AP amplitude and induced a slow transient depolarization. Under voltage clamp, D1-like and D2-like agonists induced a dose-dependent inward current that was reversibly blocked by their receptor antagonists. The inward current was blocked by tetrodotoxin (TTX), implicating Na+ channels. The reduction of AP amplitude and voltage-gated Na+ current by DA and DA agonists provides a mechanism for suppressing spike activity in type I afferent neurons. PMID- 11277588 TI - Heat hyperalgesia following partial sciatic ligation in rats: interacting nature and nurture. AB - As in humans, levels of neuropathic pain produced by nerve injury are highly variable among animals. This variability was attributed to genetic and environmental factors. For example, we reported that chronic neuropathic sensory disorders developing following total (autotomy) or partial nerve injury (allodynia and hyperalgesia) depended on the diet rats consumed. Here we investigated the interaction between genetic and dietary factors in the development of heat hyperalgesia in rats following partial sciatic ligation (the PSL model). We show that heat sensitivity of intact rats and levels of heat hyperalgesia of PSL-injured rats were highly variable across eight different rat strains and seven different diets. Thus, genetic and environmental variables interact in determination of levels of chronic neuropathic sensory disorders in rats. PMID- 11277589 TI - Changes in response modulation of cat perigeniculate neurons related to EEG state and application of neuromodulators. AB - Spike activity of single perigeniculate (PGN) neurons was recorded in the anaesthetized (N2O/halothane) and paralysed cat during presentation of moving gratings of optimal spatial frequency. Typically, the ongoing (tonic, spontaneous) activity of PGN cells increased during a rise in EEG delta power accompanied by a reduction and often a total loss of spike rate modulation by the moving grating. The opposite behaviour was found when the EEG delta power vanished. Micro-iontophoretically applied acetylcholine (ACh) had an effect similar to a decrease in EEG delta power, decreasing ongoing activity but increasing the response modulation depth. The opposite effect could be achieved with the excitatory action of serotonin (5-HT), mimicking a strengthened EEG delta power. These data support previous data indicating that PGN neurons contribute to spatio-temporal tuning of subcortical visual activity in a state dependent way. PMID- 11277590 TI - Motor imagery in normal subjects and Parkinson's disease patients: an H215O PET study. AB - Motor imagery paradigms can be used to investigate motor preparation. We used positron emission tomography to compare regional cerebral blood flow (rCBF) in patients with Parkinson's disease and normal controls under three conditions: rest, motor imagery and motor execution. In controls, imagery activated bilateral dorsolateral and mesial frontal cortex, inferior parietal cortex and precuneus. Motor execution additionally activated primary motor cortex (p < 0.001). Between group, for imagery there was relative reduction in dorsolateral and mesial frontal activation in the patient group (p<0.01). For execution, there was impaired activation of right dorsolateral frontal cortex and basal ganglia (p<0.01). Our results support the notion that underfunctioning of mesial frontal and dorsolateral prefrontal cortex may underlie motor preparation in Parkinson's disease but also suggest that akinesia may occur in the absence of impaired mesial frontal cortex activation. PMID- 11277592 TI - Cloning and tissue distribution of a novel isoform of the rat GABA(B)R1 receptor subunit. AB - We have identified a novel splice variant of the metabotropic GABA(B) receptor (R) subunit I, designated GABA(B)R1f, from a rat hippocampus cDNA library screen. GABA(B)R1f shares sequence homology with rat GABA(B)R1a, with the exception of an in-frame deletion of exon 4, resulting in a 21 bp deletion in the coding sequence of the N-terminal extracellular domain. In addition, GABA(B)R1f also contains a 93 bp in-frame insertion in a region of the sequence corresponding to the second extracellular loop and the fifth transmembrane domain, similar to that found in rat GABA(B)R1c. While being ubiquitously (but variably) expressed, reverse transcription polymerase chain reaction analysis revealed the GABA(B)R1f isoform to be most prevalent in peripheral vs central tissues, suggesting a potential role for this novel isoform in either the mediation of inhibitory transmission in these various tissues, or in as yet defined actions unrelated to central synaptic regulatory mechanisms attributable to GABA(B)R. PMID- 11277591 TI - Expression and regulation of Na pump isoforms in cultured cerebellar granule cells. AB - We investigated expression of Na pump isoforms in cultured cerebellar granule cells and measured in situ ion pump activities of the isoforms, to elucidate functions of Na pump isoforms in neurons. The cells expressed three Na pump isoforms (alpha1, alpha2 and alpha3 isoforms), however the alpha1 isoform acted as a main ion pump under basal conditions. The ion pump activity of the alpha3/ alpha2 isoforms increased remarkably after stimulation of the neurons with glutamate, therefore the alpha3/alpha2 isoforms as well as the alpha1 isoform acted as ion pumps after the stimulation. The glutamate effects were mainly mediated by non-NMDA receptors. These results suggest that alpha1 isoform and alpha3/alpha2 isoforms are functionally important under basal conditions and after neuronal excitation, respectively. PMID- 11277593 TI - 15-deoxy-delta12,14-prostaglandin J2, a specific ligand for peroxisome proliferator-activated receptor-gamma, induces neuronal apoptosis. AB - Although considerable research has shown a role for peroxisome proliferator activated receptors (PPAR) in adipose differentiation and in the regulation of inflammation, little is known about its possible functions in neurons. We investigated the role of PPARgamma in primary cultures of cortical neurons and human neuroblastoma SH-SYSY cells. Incubation of cortical neurons with the specific PPARgamma ligand 15-Deoxy-delta12,14-prostaglandin J2 (15d-PGJ2) induced morphological changes including neurite degeneration and nuclear condensation that were consistent with neurons dying by apoptosis. The morphological changes associated with incubation of cortical neurons with 15d-PGJ2 were prevented following pretreatment of neurons with the general caspase inhibitor, Z-VAD. These results highlight a novel role for PPARgamma in neurons and suggest that unwarranted activation of PPARgamma may contribute to the neuronal apoptosis associated with certain neurodegenerative disorders including Alzheimer's disease (AD). PMID- 11277594 TI - The metabolic evidence of synergistic interaction between DAMGO and DPDPE on undifferentiated SH-SY5Y cells. AB - Recent studies have demonstrated the analgesic synergy between mu- and delta opioid receptor, but evidence obtained at the cellular level is scanty. This work was designed to find the evidence of synergy between the actions of D-Ala2-Mephe4 glyol5 enkephalin (DAMGO) and D-Phe2, D-Phe5 enkephalin (DPDPE) on undifferentiated SH-SY5Y cells. Microphysiometer was used to detect the functional activity of cells by measuring the real-time extracellular acidification rate (ECAR). The results demonstrate the unequivocal synergy between DAMGO and DPDPE at least within certain ratios. In addition, combined administration of the two drugs in the synergistic ratios attenuates receptor desensitization. These data indicate that DAMGO and DPDPE have a synergistic effect at cellular level. PMID- 11277595 TI - Impaired cerebral glucose metabolism and cognitive functioning predict deterioration in mild cognitive impairment. AB - The objective of this study was to assess whether reduced glucose metabolism (rCMRGlu) and cognitive functioning could predict development of Alzheimer's disease (AD) in subjects with mild cognitive impairment (MCI). Twenty MCI patients underwent baseline and follow-up investigations of rCMRGlu, as measured by PET, and cognitive function measured by neuropsychological test assessments. Subjects were clinically followed up with an average interval of 36.5 months. Two groups were obtained after the second clinical assessment. Nine patients were diagnosed as AD and classified as progressive MCI (P-MCI), whereas 11 patients remained clinically stable and were classified as stable MCI (S-MCI). There were no differences in demographic variables or baseline MMSE between the two subgroups. Logistic regression indicated the two variables that most effectively predicted future development of AD were rCMRGlu from the left temporoparietal area and performance on the block design. These combined measures gave an optimal 90% correct classification rate, whereas only rCMRGlu or neuropsychology alone gave 75% and 65% correct classification, respectively. Measures of temporoparietal cerebral metabolism and visuospatial function may aid in predicting the evolution to AD for patients with MCI. PMID- 11277596 TI - Analgesic domains of interferon-alpha. AB - Interferon-alpha (IFNalpha) is not only an immunoregulatory factor, but is also an analgesic molecule. We ever reported that there exist distinct domains in IFNalpha molecule that mediate immune and analgesic effects respectively and inferred that the analgesic domain locates around the 122nd Tyr residue of IFNalpha molecule in the tertiary structure. After the 36th Phe residue, which was located closely to the 122nd Tyr residue in the tertiary structure, was mutated to Ser using site-directed mutagenesis, the analgesic activity of this mutant lost completely, but the antiviral activity of IFNalpha still maintained 40.5% of wild type IFNalpha. The results suggest that the 36th Phe residue is one of the constituent for the analgesic domain of IFNalpha and inferred that the analgesic domain of IFNalpha consists of the 122nd Tyr and the residues around the 122nd in the tertiary structure, which include the 36th Phe. PMID- 11277597 TI - Dynamics of cortical activation in a hemianopic patient. AB - Although residual vision in patients with cortical blindness is common, its brain mechanisms are poorly known. To study these mechanisms we measured neuromagnetic responses to visual stimuli in a patient with right posterior cerebral lesion and left visual field hemianopia. His vision had partially recovered with intensive training before our measurements. Compared with the processing in the healthy side, early occipital responses were attenuated for both passive viewing of checkerboard reversal patterns and a letter identification task. In both conditions there were prominent longer-latency responses at the right superior temporal cortex. We suggest that the activation in the superior temporal cortex can partially compensate for the failure to produce synchronized population responses at the early stages of visual cortical processing. PMID- 11277598 TI - N- and L- but not P/Q-type calcium channels contribute to neuropeptide release from rat skin in vitro. AB - Here we directly demonstrate the liberation of CGRP from rat skin in vitro induced by high extracellular concentrations of KCl. The EC50 was 52 mM KCl and saturation was reached from 80 mM KCl. The release was entirely dependent on the presence of extracellular calcium ions. It was reduced by nonsubtype selective inhibition of voltage-operated calcium channels (VOCCs). Application of selective antagonists suggest expression of L-type and N-type but not P/Q-type VOCCs in cutaneous nociceptive terminals. These may be activated by any suprathreshold depolarizing stimuli to induce neurogenic inflammation. The expression pattern greatly differs from central nociceptive terminals where, in addition, P/Q-type VOCC have been found. PMID- 11277599 TI - Quinolinic acid induces oxidative stress in rat brain synaptosomes. AB - The oxidative action of quinolinic acid (QUIN), and the protective effects of glutathione (GSH), and 2-amino-5-phosphonovaleric acid (APV), were tested in rat brain synaptosomes, Reactive oxygen species (ROS) formation was quantified after the exposure of synaptosomes to increasing concentrations of QUIN (25-500 microM). The potency of QUIN to induce lipid peroxidation (LP) was tested as a regional index of thiobarbituric acid-reactive substances (TBARS) production, and the antioxidant actions of both GSH (50 microM) and APV (250 microM) on QUIN induced LP were evaluated in synaptosomes prepared from different brain regions. QUIN induced concentration-dependent increases in ROS formation and TBARS in all regions analyzed, but increased production of fluorescent peroxidized lipids only in the striatum and the hippocampus, whereas both GSH and APV decreased this index. These results suggest that the excitotoxic action of QUIN involves regional selectivity in the oxidative status of brain synaptosomes, and may be prevented by substances exhibiting antagonism at the NMDA receptor. PMID- 11277600 TI - Modulation of P-CREB and expression of c-fos in cochlear nucleus and superior olive following electrical intracochlear stimulation. AB - Investigating activity-dependent plasticity in the auditory brain stem of the adult rat, we observed that electrical intracochlear stimulation led to a tonotopically localized modulation of the phosphorylation of the cAMP response element binding protein (CREB) and an equally localized expression of the immediate early gene product c-Fos in cochlear nucleus and superior olive. As P CREB is thought to act as transcription factor on one promoter site of the c-fos gene, we compared immunolabeling for P-CREB and c-fos in adjacent brain sections. Following 2h sustained stimulation in previously deafened animals, labelling for P-CREB declined in regions where c-Fos labelling increased. This suggests that the level or state of P-CREB (e.g. whether it is phosphorylated or not) are affected by intracochlear stimulation in a process that appears to be linked to the stimulation-dependent expression of c-Fos in auditory brain stem nuclei. PMID- 11277601 TI - Activation of glutamate uptake by guanosine in primary astrocyte cultures. AB - Guanine-based purines have been shown to modulate the effects of glutamate, which is essential for brain function and mediates excitotoxicity. In the search for a mechanism involving the interaction between purine nucleoside guanosine and glutamate, we found that guanosine dose-dependently, significantly (63%) and potently (EC50 =2.47 microM) enhanced glutamate uptake in cultured astrocytes. This effect was not inhibited by the blocker of nucleoside transporter dipyridamole nor by the adenosine antagonist theophylline, suggesting an extracellular site of action without the involvement of adenosine receptors. These results indicate a regulatory role of guanosine on extracellular levels of glutamate, possibly contributing for protecting neural cells against glutamate induced excitotoxicity. PMID- 11277602 TI - A dose-finding and discontinuation study of clomipramine in panic disorder. AB - Eighty-one panic disorder patients with or without agoraphobia were treated with flexible doses of clomipramine under single-blind conditions. Fifty-seven (70.3%) reached operational criteria for full remission in 16.2 +/- 6.5 weeks, with a mean dose of 89.1 +/- 8.2 mg/day. Fifty-four (81%) of them received a continuous post-remission maintenance treatment at full doses of clomipramine for 4-6 months. No patient relapsed during the clomipramine maintenance phase. Their medication was then tappered and discontinued with placebo substitution under double-blind conditions. Fifty-one (63%) patients were followed-up until relapse or recurrence for up to 3 years, with periodic assessments. Three different outcome groups were identified: the first (n = 19, 19; 37.2%) experienced an early/immediate relapse (5.2 +/- 4.9 weeks after drug discontinuation); the second group (n = 22, 22; 43.1%) experienced recurrence after 42.9 +/- 35 weeks following discontinuation; and the third group (n = 10, 10; 19.6%) remained assymptomatic and functionally well throughout the follow-up. Predictors of early relapse were: (1) higher baseline score in the Beck Depression Inventory; (2) higher global score on the phobic avoidance scale after the full remission criteria; and (3) the need for higher clomipramine doses to reach full remission. The need for long-term or intermittent maintenance for most patients is emphasized. PMID- 11277603 TI - Selective imidazoline I2 ligands do not show antidepressant-like activity in the forced swim test in mice. AB - Clonidine is an adrenergic agonist with high affinity for alpha2 adrenoceptors that also has affinity for imidazoline receptors. Clonidine has previously been shown to reduce immobility in the forced swim test (FST) in mice. In the present study, this effect was blocked by idazoxan (0.06 mg/kg s.c.) and by yohimbine (1.0 mg/kg s.c.) suggesting that clonidine's effects in this test are mediated via its action at alpha2 sites. Imidazoline I2 site ligands have been shown to inhibit monoamine oxidase and thus may also have antidepressant activity. Three compounds with selective affinity for I2 receptors (BU224, BU239, BDF 8082) were also tested in the FST. These compounds showed no activity either alone or in combination with a subthreshold dose of imipramine in the FST. These results suggest that I2 receptor ligands do not show antidepressant-like activity in the FST in mice. Furthermore the activity of the mixed alpha2/I1 agonist clonidine is most likely to be due to its action at alpha2 sites. PMID- 11277604 TI - Effects of d-amphetamine on the performance of rats in an animal analogue of the A-X continuous performance test. AB - Schizophrenia patients subjected to the A-X Continuous Performance Test (A-X CPT) show cognitive deficits that are thought to reflect impaired representation and maintenance of context information. An issue deserving attention is to what extent the acute amphetamine model of schizophrenia also models these cognitive deficits. The present experiment examined the effect of acute d-amphetamine (AMP) on the performance of rats in an animal analogue of the A-X CPT. Subjects first learned to solve an A --> X+, B --> X-, A --> Y- discrimination task, with A and B representing visual features; X and Y designating auditory target stimuli; --> signifying a serial presentation; and + and - referring to food reinforcement and non-reinforcement, respectively. Frequency of food-magazine visits was the dependent measure. After mastering the discrimination, rats received test trials under either saline or 0.5 mg/kg AMP (s.c.). At test, the interval between feature and target presentation was varied; reinforcement contingencies were maintained. AMP significantly impaired performance on the A --> X+/B --> X- discrimination by increasing the response level on B --> X- trials. AMP did not significantly affect performance on the A --> X+/A --> Y- discrimination. However, AMP also increased magazine responding in the absence of the presentation of features and targets. A parsimonious conclusion based on these preliminary results is that acute AMP does not affect processing of context information provided by the visual features in this procedure. It rather has a more non-specific response-enhancing effect, especially with respect to stimuli associated with the delivery of food. PMID- 11277605 TI - Discriminative stimulus properties of indorenate, a 5-HT1A, 5-HT1B and 5-HT2C agonist: a study in rats. AB - Indorenate (INDO), initially described as an antihypertensive agent, also has some effects on behaviour, with anxiolytic and anorectic actions being reported. The aim of the present experiment was to examine the activity of INDO at the behavioural level at various serotonin (5-hydroxytryptamine, 5-HT) receptor sites by comparing its stimulus properties with those of other 5-HT receptor agonists and by examining its interactions with some 5-HT antagonists. Rats were trained to discriminate between 10.0 mg/kg INDO (administered intraperitoneally (90 min before the start of the session) from saline. A Fixed Ratio 10 (FR10) schedule of reinforcement was in effect in each drug condition. During generalization test sessions, the discrimination index (DI, responses to drug lever/responses to drug + saline lever) was calculated from the responses emitted before the first reinforcer of the session. DI was a function of the dose of INDO employed. Generalization to the discriminative stimulus properties of INDO was observed with the 5-HT1A receptor agonist 8-OH-DPAT (1.0 mg/kg produced 90% generalization) and the 5-HT(1B/2C) receptor agonist 1-(3-trifluoromethylphenyl) piperazine (TFMPP) (3.0 mg/kg produced up to 75% generalization). Yohimbine (5.6 mg/kg), buspirone (1.0 mg/kg), 6-chloro-2-(1-piperaziny)pyrazine (1.0 mg/kg) and m-chlorophenylpiperazine (mCPP) (1.0 mg/kg) induced a DI of 70%, 50% and 48% and 55%, respectively. In generalization tests, ritanserin (0.01-1.0 mg/kg) induced saline-like responding. NAN-190 (3.0 mg/kg), a 5-HT1A receptor antagonist, was able to reduce the DI of INDO to 50%. Although the 5-HT(2C/2A) receptor antagonists cinanserin (10.0 mg/kg) and metergoline (0.3 mg/kg) were able to reduce the stimulus properties of INDO to 60% and 30%, respectively, only ritanserin (1.0 mg/kg) reduced the stimulus properties of INDO to 25% with a clear dose-response relationship. The results suggest that INDO acts as an agonist at 5-HT1A receptor sites, but its activity at 5-HT(1B/2C) receptor sites also contributes to its discriminative function. PMID- 11277606 TI - Investigation of preference for nightly triazolam versus placebo in moderate social alcohol drinkers. AB - This study was designed to examine whether the widely prescribed benzodiazepine hypnotic triazolam has reinforcing effects in moderate social alcohol drinkers, without histories of drug abuse or insomnia, in the context of its use as a hypnotic. Eleven healthy adult volunteers who met criteria for 'good sleepers' participated in a 60-session double-blind choice study which was conducted on an outpatient basis with participants sleeping at home. Twenty three-session sampling/choice tests were conducted sequentially to provide 20 evaluations of the reinforcing effects of 0.25 mg/70 kg triazolam versus placebo, ingested orally 30 min before bedtime. Each three-session test consisted of two sampling sessions, in which participants received exposure to each of the two drug conditions in different colored capsules, followed by one choice session, in which participants were asked to choose one of the two colour-coded capsules for self-administration. Four participants exhibited a significant choice of triazolam, three, a significant choice of placebo (i.e. triazolam avoidance), and four, a random (i.e. non-significant) choice between triazolam and placebo. The reasons provided by participants were consistent with their choices and with the expected effects of triazolam versus placebo. Analyses of post-sleep questionnaires indicated that triazolam did not produce a clinically meaningful improvement in sleep. The finding that triazolam functioned as a reinforcer in participants without insomnia suggests that triazolam has reinforcing effects in some individuals for which hypnotic treatment is not clinically indicated, and that health care professionals must continue to assess the risk/benefit ratio of benzodiazepine hypnotic prescription. PMID- 11277608 TI - Hypericum perforatum (St John's Wort): a non-selective reuptake inhibitor? A review of the recent advances in its pharmacology. AB - Hypericum possesses a unique pharmacology in that it displays the pharmacology of many classes of antidepressants and new mechanisms not typical of standard antidepressants. The most potent of all its action is the moderate to high potency for inhibition of the reuptake of monoamines, serotonin, dopamine and noradrenaline and the amino-acid neurotransmitters GABA and glutamate. Unlike standard reuptake inhibitors, hypericum exerts this reuptake inhibition non competitively by enhancing intracellular Na+ ion concentrations. At a receptor level, chronic treatment with hypericum downregulates beta1-adrenoceptor, upregulates post-synaptic 5-HT1A receptors and 5-HT2 receptors. Although the major constituent responsible for the antidepressant effect is thought to be hyperforin, other constituents such as hypericin, pseudohypericin, flavonoids and oligomeric procyanidines may also play a direct or indirect role. While reuptake inhibition may more than likely be responsible for most of the antidepressant effect, other mechanisms may also contribute alone or in combination to exert the overall antidepressant action. PMID- 11277607 TI - The 5-HT1A receptor in schizophrenia: a promising target for novel atypical neuroleptics? AB - Increasing attention is being directed towards the role of the serotonergic system in the neurochemistry of schizophrenia and antipsychotic drug treatment. This review considers the 5-HT1A receptor in this context. In patients with schizophrenia, the majority of post-mortem studies have reported increases in 5 HT1A receptor density in the prefrontal cortex in the approximate range 15-80%. Although the pathophysiological significance of this finding is unclear, given the location of a major proportion of these receptors on pyramidal cells, it may reflect an abnormal glutamatergic network. In terms of drug treatment, 5-HT1A agonists clearly display anticataleptic activity in rats. In addition, 5-HT1A agonists consistently increase dopamine release in the prefrontal cortex in rodents, which is an effect that might be predicted to improve negative symptoms. 5-HT1A agonists augment classical neuroleptics in some rat models of antipsychotic action and may be capable of modulating the glutamatergic network therapeutically. Despite the encouraging preclinical data, there is a paucity of clinical studies of 5-HT1A agonist augmentation of neuroleptics in the treatment of schizophrenia. However, the clinical relevance may be clarified by the atypical antipsychotic drugs clozapine, quetiapine and ziprasidone which combine D2 receptor antagonism and 5-HT1A agonism. In conclusion, given the increased prefrontal 5-HT1A receptor density in the illness, and the anticataleptic activity of 5-HT1A agonists combined with their ability to evoke prefrontal dopamine release, there is now a sufficient rationale to examine thoroughly the role of the 5-HT1A receptor in schizophrenia and antipsychotic drug treatment. PMID- 11277609 TI - Treatment of bipolar affective disorder in clinical practice. AB - A case note survey of 100 outpatients with a clinical diagnosis of bipolar affective disorder in a UK inner city teaching hospital revealed monotherapy with a mood stabilizer in only 23% of patients, mostly lithium (15%). Overall, 51% of patients were prescribed lithium, 19% carbamazepine and 5% valproate with only 8% receiving a combination of two mood stabilizers. Treatment appeared to be inadequate in 13/51 of patients on lithium, 9/19 of those on carbamazepine and 1/5 of those on valproate. Antipsychotics were used as monotherapy in 20% of patients and combined with a mood stabilizer in 43% of patients. Only 6% of patients were on atypical antipsychotics. These findings suggest that the treatment for many patients does not match recommendations. Clearer evidence on the place of combination mood stabilizers and adjunctive antipsychotics, particularly atypicals is needed in the treatment of bipolar affective disorder. PMID- 11277610 TI - Intermittent, luteal phase nefazodone treatment of premenstrual dysphoric disorder. AB - Three outpatients who fulfilled full DSM-IV diagnostic criteria for premenstrual dysphoric disorder (PDD) were successfully treated with intermittent (luteal phase) nefazodone. They received the medication at low doses of up to 100 mg/day (50 mg b.i.d.), for 2 weeks through the luteal phase of the menstrual cycle only. All the patients reported a marked symptomatic improvement, including full remission of their emotional symptoms, and two achieved in addition full remission of their somatic symptoms. Side-effects reported during the treatment were mild. The use of luteal phase nefazodone seems to be a promising treatment strategy for the management of PDD. It offers advantages over daily dosing throughout the menstrual cycle, such as reduced incidence and severity of side effects, and avoids the stigma that may accompany the continuous use of psychopharmacological treatment, with the advantage that compliance may be improved. PMID- 11277611 TI - Neuroleptic malignant-like syndrome after abrupt withdrawal of baclofen. AB - We present the case of a 36-year-old man who presented with a clinically neuroleptic malignant-like syndrome involving disorientation, signs of autonomic dysfunction, rigidity and raised total creatine kinase level, but in the absence of any neuroleptic medication. He had, however, abruptly stopped taking his long term baclofen in the days prior to presentation. He improved markedly after the reintroduction of baclofen, and we postulate that his clinical syndrome resulted from the sudden withdrawal of this drug. We concur with the concept that neuroleptic malignant syndrome represents a spectrum of disorders, and add it to the list of possible sequelae after abrupt withdrawal of baclofen. PMID- 11277612 TI - Venlafaxine occupation at the noradrenaline reuptake site: in-vivo determination in healthy volunteers. AB - Venlafaxine, a serotonin and noradrenaline reuptake inhibitor, is an effective antidepressant at doses of 75 mg p.o. daily and above. Preclinical and healthy volunteer studies have demonstrated that venlafaxine is more potent at the serotonin than at the noradrenaline reuptake site, with noradrenergic blocking effects being observed at doses >75 mg p.o. in man. We used the Multiple Organs Coincidences Counter and [11C] meta hydroxy ephedrine (MHED) to test whether significant occupation of cardiac sympathetic neurones was achieved in man in vivo after the acute administration of venlafaxine 75 mg p.o. in nine healthy volunteers. MHED is a tracer which binds at the noradrenaline reuptake site. This study demonstrates that the [11C]MHED signal is significantly reduced after the administration of venlafaxine 75 mg p.o. thus showing that noradrenaline reuptake blockade is observable at this dose. This effect is predominantly seen in volunteers who received > 1 mg/kg venlafaxine. PMID- 11277613 TI - Antithrombin III enhances inducible nitric oxide synthase gene expression in vascular smooth muscle cells. AB - Evidence suggests that antithrombin III (ATIII) exerts anti-inflammatory properties in addition to its anti-coagulative mechanisms. In animal models of sepsis, ATIII affected cytokine plasma concentrations with a decrease of pro inflammatory cytokines. In addition to cytokines, excessive production of nitric oxide (NO) derived from inducible nitric oxide synthase (iNOS) might represent another important mediator of the cytotoxic events during sepsis. Regarding ATIII as a potential anti-inflammatory modulator, one may speculate that ATIII inhibits the synthesis of iNOS-derived NO. However, our data demonstrate that ATIII further stimulates iNOS gene expression when applied together with either interleukin-1 beta or the combination of lipopolysaccharide plus interferon gamma. The most prominent synergistic effects on NO synthesis were found when ATIII was given at higher concentrations (1, 5, and 10 U/ml). Although the mechanisms of ATIII signal transduction remain to be established, intensification of interleukin-1 beta or interferon-gamma/lipopolysaccharide-induced NO synthesis by ATIII does not attribute to the anti-inflammatory properties of ATIII. PMID- 11277614 TI - The immune response modifier resiquimod mimics CD40-induced B cell activation. AB - Members of the imidazoquinoline molecule family, including imiquimod and resiquimod (R-848), have potent antiviral and antitumor activities. Imiquimod cream (5%) (Aldara) is currently indicated for treatment of external genital and perianal warts. Previous characterization of these compounds has focused upon their ability to activate monocytes and dendritic cells, but recent studies have shown that resiquimod also stimulates B lymphocytes to proliferate and express an activated phenotype. This suggests that resiquimod could potentially serve as an effective vaccine adjuvant in stimulating a humoral immune response. This study shows that resiquimod mimics effects of the T-dependent CD40 signal in both mouse and human B cell lines. Resiquimod, like CD40, stimulates antibody secretion, cytokine production, protection from apoptosis, and CD80 upregulation. In addition, it shows synergy with signals delivered by the B cell antigen receptor and heightens CD40-mediated B cell activation, demonstrating that resiquimod can enhance antigen-specific responses in B lymphocytes. PMID- 11277615 TI - Epidermal growth factor regulates astrocyte expression of the interleukin-4 receptor via a MAPK-independent pathway. AB - Human astrocytes express the interleukin (IL)-4 receptor alpha chain (IL-4R alpha) in vitro and in vivo but mechanisms governing astrocyte IL-4R alpha expression have not been established. We hypothesized that epidermal growth factor (EGF) and IL-4, agents that profoundly affect astrocyte proliferation, might also alter IL-4R alpha expression. Exposure to EGF for 24 h enhanced IL-4R alpha mRNA levels; in contrast, IL-4 yielded no increase. Immunoblotting demonstrated that EGF but not IL-4 increased astrocyte IL-4R alpha protein after 2--4 days of exposure. Similarly, EGF but not IL-4 strongly activated phosphorylation of p42/p44 extracellular regulated kinase isoforms, a reaction blocked by the mitogen-activated protein kinase (MAPK) inhibitor, PD98059. PD98059 also blocked EGF-stimulated DNA synthesis but not IL-4R alpha mRNA levels, while antibody to the EGF receptor (erbB1) blocked both EGF effects. Data suggest that astrocyte IL-4R alpha expression is upregulated by EGF but not by IL 4 in an EGF-receptor-dependent manner and that mechanisms are independent of MAPK activation. PMID- 11277616 TI - Functional CD4(+) and CD8(+) T-cell responses induced by autologous mitomycin C treated Epstein-Barr virus transformed lymphoblastoid cell lines. AB - Epstein-Barr virus (EBV) gene expression in tumor cells of posttransplant lymphoproliferative disorder (PTLD) patients resembles that of EBV transformed B cell lines (LCL). EBV-specific cytotoxic T-lymphocytes can be generated by stimulating peripheral blood lymphocytes with autologous LCL. We describe a standardized method for the growth inactivation and cryopreservation of LCL for optimal T-cell stimulation and analyzed the function and phenotype of responding T-cells. LCL growth was completely blocked by mitomycin C treatment (McLCL) and McLCL could be cryopreserved while retaining excellent APC function. McLCL stimulated both CD4(+) and CD8(+) T-cells as measured by HLA-DR and CD25 expression using FACS analysis. EBV-specific CTL activity and T-cell proliferation were induced and immunocytochemical staining showed CD4(+) and (granzyme B positive) CD8(+) T-cells rosetting with McLCL. Granzymes A and B, IFN gamma, and IL-6 were detected at significant levels in the supernatant. Thus, ex vivo T-cell activation with cryopreserved McLCL results in activation of both CD4(+) and CD8(+) T-cells producing a Th1-like cytokine profile, making this a suitable protocol for adoptive therapy of PTLD. PMID- 11277617 TI - Analysis of IL-12 receptor beta 2 chain expression of circulating T lymphocytes in patients with atopic asthma. AB - Th2 cell predominance relative to Th1 cells contributes to pathological immune responses in patients with atopic asthma. IL-12 is a key cytokine in the induction of Th1 cells, and downregulation of IL-12 production is reported in these patients. However, IL-12 receptor expression of their T lymphocytes has not been clarified. In this study, expression of IL-12 receptor beta 2 on T cells and secretion of cytokines which affect IL-12 receptor beta 2 expression by their PBMC were examined. We found that IL-12 receptor beta 2 expression of the T cells is reduced. This is partly due to the diminished production of IL-12 and enhanced secretion of IL-4 by their PBMC. IL-18 production is not significantly modulated in these patients. Furthermore, intrinsic defects of the CD4(+) T cells, which reduce their IL-12 receptor beta 2 expression in response to IL-12 and/or IL-18 stimulation, are evident and are importantly involved in the Th1/Th2 imbalance of patients with atopic asthma. PMID- 11277618 TI - Differential stimulation of helper and cytotoxic T cells by dendritic cells after infection by Yersinia enterocolitica in vitro. AB - Dendritic cells (DC) are antigen-presenting cells crucial for initiating immune responses like sensitization of T cells to foreign antigens. We have previously shown that infection of DC by enteropathogenic Yersinia enterocolitica in vitro leads to a transient suppression in the immunostimulatory capacity for autologous enriched total T cells. In this study, we found that killed Yersinia could replace live bacteria in this aspect, and that yersinial lipopolysaccharide (LPS) antigen could be detected intracellularly over a time course of 8 days. A suppressive effect on T cell proliferation after stimulation with Yersinia infected compared to uninfected DC was seen for CD4+ T cells isolated by immunomagnetic separation techniques over the whole time course of 8 days, whereas CD8+ T cells followed to exhibit a suppressed proliferation rate starting on day 5 post infection till the end of the time course. In contrast, enriched total T cells stimulated by Yersinia-infected DC showed weaker proliferation till day 6 post infection compared to stimulation by uninfected DC, but not thereafter. Mixing of purified CD4+ and CD8+ T cells at day 8 post infection could reconstitute the effect seen for enriched total T cells. Thus, helper in concert with cytotoxic T cells might contribute to the immune responses, that are necessary for control of Yersinia-infections. PMID- 11277619 TI - IL-12 prevents tolerance induction with mouse thyroglobulin by priming pathogenic T cells in experimental autoimmune thyroiditis: role of IFN-gamma and the costimulatory molecules CD40l and CD28. AB - Deaggregated mouse thyroglobulin (dMTg) induces tolerance to experimental autoimmune thyroiditis (EAT), a Th1-cell-mediated disease. To test whether IL-12, a potent activator of Th1 cells, can overcome tolerance induction, different doses of IL-12 were given to CBA/J mice during the critical interval of 2--3 days after dMTg administration. After challenge with MTg/LPS, dMTg/IL-12-pretreated mice showed more extensive thyroiditis than immunized controls, but comparable levels of anti-MTg and T cell proliferation. Without challenge, few MTg antibodies were produced. In contrast, pretreatment with dMTg/poly A:U or dMTg/IL 1, two other T cell activators which also interfere with tolerance induction, induced antibodies before challenge, but not more severe thyroiditis. Mice pretreated with IL-12 without dMTg developed thyroiditis comparable to immunized controls, but less severe thyroiditis than dMTg/IL-12-pretreated mice. Clearly, IL-12 not only blocked tolerance induction, but also primed antigen-specific T cells during the tolerogenic period of dMTg pretreatment, resulting in stronger thyroiditis than immunization only. Neither treatment with anti-IFN-gamma nor the use of IFN-gamma knockout mice altered the capacity of IL-12 to prevent tolerance induction. However, both anti-CD28 and anti-CD40L antibodies diminished the priming effect by dMTg/IL-12. The mechanisms of IL-12 action include priming of MTg-specific T cells and the involvement of T cell costimulatory molecules. PMID- 11277621 TI - Seeking a vaccine against Coccidioides immitis and serologic studies: expectations and realities. AB - The studies reported in this review indicate that, whereas the expectations from the molecular approach help to excite and enlighten us, the realities suggest that even some less modern approaches may provide the necessary practical solutions to problems of serology and vaccination against coccidioidomycosis. Concurrent conduct of the two approaches should maximize the chances for success. PMID- 11277620 TI - Dexamethasone modulates TCR zeta chain expression and antigen receptor-mediated early signaling events in human T lymphocytes. AB - Dexamethasone is a potent anti-inflammatory and immunosupressive agent that has complex, yet incompletely defined, effects on the immune response. Here, we explored the effect of dexamethasone on the expression of TCR zeta chain and TCR/CD3-induced early signaling events in human T lymphocytes. Immunoblotting studies using TCR zeta chain specific mAb showed a dose-dependent biphasic effect of dexamethasone on TCR zeta chain expression, that is, it was increased when cells were incubated with 10 nM, whereas the expression was decreased when incubated with 100 nM dexamethasone. The dose-dependent biphasic effect of dexamethsone on the TCR zeta chain expression was also revealed by FACS analysis of permeabilized cells. Time course studies showed that upregulation of the TCR zeta chain at 10 nM dexamethasone reached maximum levels at 24 h and remained elevated up to 48 h. Other subunits of the TCR/CD3 complex were minimally affected under these conditions. The increased expression of the TCR zeta chain following treatment with 10 nM dexamethasone correlated with increased anti-CD3 antibody-induced tyrosine phosphorylation of the TCR zeta chain and downstream signaling intermediate ZAP-70 and PLC gamma with faster kinetics. Similarly, the induction of TCR zeta chain expression at 10 nM dexamethasone correlated with increased and more sustained TCR/CD3-mediated [Ca(2+)](i) response. Reporter gene assays using TCR zeta chain promoter-driven luciferase gene constructs in Jurkat cells showed that treatment with 10 nM dexamethasone increased TCR zeta chain promoter activity and that the region between -160 and +58 was responsible for the observed effect. These results suggest that dexamethasone primarily acts at the transcriptional level and differentially modulates TCR zeta chain expression and antigen receptor-mediated early signaling events in human peripheral T lymphocytes. PMID- 11277622 TI - Organization and distribution pattern of MGLR-3, a novel retrotransposon in the rice blast fungus Magnaporthe grisea. AB - A specific telomere was deleted in spontaneous, gain-of-virulence mutants derived from a rice pathogen of Magnaporthe grisea. Three different types of transposons, including Pot2, Mg-SINE, and a novel, 6-kb-long LTR (long terminal repeat)-type retrotransposon designated MGLR-3, were identified on this chromosomal end. The 114-bp-long telomeric repeat is immediately followed by the 3' LTR of MGLR-3. A truncated copy of Pot2 immediately flanks the 5' LTR, suggesting that this telomere was generated by a transposition event of MGLR-3 into this Pot2 element, causing the breakage of a chromosome. The subsequent addition of a telomeric repeat to the 3' LTR of MGLR-3 most probably repaired the broken end of the chromosome. Mg-SINE is located 25 bp away from the truncated Pot2 element. MGLR-3 exhibited strong homology to various gypsy-class retrotransposons, including grh and MAGGY in M. grisea. MGLR-3 is ubiquitous regardless of the host of origin. PMID- 11277623 TI - Regulation of hisHF transcription of Aspergillus nidulans by adenine and amino acid limitation. AB - The hisHF gene of Aspergillus nidulans encodes imidazole-glycerole-phosphate (IGP) synthase, consisting of a glutamine amidotransferase and a cyclase domain. The enzyme catalyzes the fifth and sixth steps of histidine biosynthesis, which results in an intermediate of the amino acid and an additional intermediate of purine biosynthesis. An A. nidulans hisHF cDNA complemented a Saccharomyces cerevisiae his7Delta strain and Escherichia coli hisH and hisF mutant strains. The genomic DNA encoding the hisHF gene was cloned and its sequence revealed two introns within the 1659-bp-long open reading frame. The transcription of the hisHF gene of A. nidulans is activated upon amino acid starvation, suggesting that hisHF is a target gene of cross pathway control. Adenine but not histidine, both end products of the biosynthetic pathways connected by the IGP synthase, represses hisHF transcription. In contrast to other organisms HISHF overproduction did not result in any developmental phenotype of the fungus in hyphal growth or the asexual life cycle. hisHF overexpression caused a significantly reduced osmotic tolerance and the inability to undergo the sexual life cycle leading to acleistothecial colonies. PMID- 11277624 TI - ADHII in Aspergillus nidulans is induced by carbon starvation stress. AB - In Aspergillus nidulans there are three NAD(+)-dependent alcohol dehydrogenases (ADHs) that are capable of utilizing ethanol as a substrate. ADHI is the physiological enzyme of ethanol catabolism and ADHIII is induced under conditions of anaerobiosis. The physiological role of ADHII (structural gene alcB) is unknown. We have measured beta-galactosidase in a transformant with an alcB::lacZ fusion and have shown that alcB is maximally expressed under conditions of carbon starvation. The behavior of the alcB::lacZ transformant suggests a hierarchy of repressing carbon sources characteristic of repression by the general carbon catabolite repressor protein, CreA, but in a creA(d)30 background the transformant shows only partial derepression of beta-galactosidase on 1% glucose compared to the creA+ strain. Our results suggest that, in addition to carbon catabolite repression acting via CreA, a CreA-independent mechanism is involved in induction of alcB on carbon starvation. PMID- 11277625 TI - Genetic and physical mapping of two centromere-proximal regions of chromosome IV in Aspergillus nidulans. AB - Chromosome IV is the smallest chromosome of Aspergillus nidulans. The centromere proximal portion of the chromosome was mapped physically using overlapping clones of a cosmid genomic library. Two contiguous segments of a physical map, based on restriction mapping of cosmid clones, were generated, together covering more than 0.4 Mb DNA. A reverse genetic mapping approach was used to establish a correlation between physical and genetic maps; i.e., marker genes were integrated into physically mapped segments and subsequently mapped by mitotic and meiotic recombination. The resulting data, together with additional classical genetic mapping, lead to a substantial revision of the genetic map of the chromosome, including the position of the centromere. Comparison of physical and genetic maps indicates that meiotic recombination is low in subcentromeric DNA, its frequency being reduced from 1 crossover per 0.8 Mb to approximately 1 crossover per 5 Mb per meiosis. The portion of the chromosome containing the functional centromere was not mapped because repeat-rich regions hindered further chromosome walking. The size of the missing segment was estimated to be between 70 and 400 kb. PMID- 11277627 TI - The Coachella valley fringe-toed lizard (Uma inornata): genetic diversity and phylogenetic relationships of an endangered species. AB - A phylogeny was reconstructed for 23 populations of fringe-toed lizards (genus Uma) from the three most northern species of the genus, including the Mojave fringe-toed lizard U. scoparia, the Colorado Desert fringe-toed lizard U. notata, and the endangered Coachella Valley fringe-toed lizard U. inornata. The outgroup taxa were the zebra-tailed lizard, Callisaurus draconoides; the lesser earless lizard, Holbrookia maculata; and the greater earless lizard, Cophosaurus texanus. Evaluation of 1630 combined nucleotide sequence from the mitochondrial genes ATPase 6 and cytochrome b yielded 10 most parsimonious trees. Reweighting the characters using the rescaled consistency index eliminated eight of these trees. The remaining two trees differ only in the placement of two individuals from the Superstition Mountains which either formed a monophlyetic unit or grouped with one individual from the Anza-Borrego population. The preferred phylogeny, one more consistent with geography, had two primary clades: one consisting of U. scoparia and the other placing U. inornata inside the clade containing U. notata. Uma inornata was most closely related to nearby U. notata notata, as opposed to more distant U. notata rufopunctata. PMID- 11277626 TI - A study of the protein secretory pathway of Aspergillus niger using a glucoamylase-GFP fusion protein. AB - The effect of various treatments that block protein secretion was visualized in Aspergillus niger using a strain expressing a glucoamylase-GFP fusion protein. Cold shock caused the retention of the fusion protein in a reticulate network (ER) with brighter nodes that may represent Golgi bodies. Treatment of germlings with brefeldin A (BFA) also initially caused accumulation within the ER but prolonged exposure led to the formation and targeting of the fusion protein to vacuoles from the ER. Disruption of actin with cytochalasin A initially led to a faint diffuse accumulation and ultimately to the formation of aggregated bodies which were not vacuoles, suggesting that the actin cytoskeleton is important in secretory vesicle transport. Disruption of microtubules with nocodazole led to hyperbranching but did not cause intracellular accumulation, suggesting that microtubules play a role in directing vesicle transport rather than vesicle movement per se. Treatment of regenerating protoplasts confirmed that BFA and cytochalasin but not nocodazole inhibited protein secretion. When germlings were subjected to carbon starvation, vacuolation was rapidly initiated throughout the hyphae and GFP fluorescence was visible in some of the vacuoles, indicating retargeting of the fusion protein from the secretory pathway to the vacuoles. PMID- 11277628 TI - Phylogenetic relationships of Australian members of the family percichthyidae inferred from mitochondrial 12S rRNA sequence data. AB - Phylogenetic relationships among 20 Australian species of the family Percichthyidae were investigated from sequence data of two portions of the mitochondrial 12S rRNA gene. The molecular data indicate that Australian genera within this family cluster into three distinct clades. The first clade is composed of some species currently ascribed to the genus Macquaria, along with Nannatherina, Nannoperca, and Bostockia, the second of Maccullochella and two catadromous Macquaria species, and the third of Gadopsis. However, the positioning of Gadopsis within this family was unresolved. Monophyly within each genus was well supported, except for Macquaria, which is clearly polyphyletic. The molecular data were used to examine two hypotheses of Australian percichthyid evolution and favor a freshwater origin for the family. PMID- 11277629 TI - Molecular phylogenetic analysis of the dragonfly genera Libellula, Ladona, and Plathemis (Odonata: Libellulidae) based on mitochondrial cytochrome oxidase I and 16S rRNA sequence data. AB - Molecular phylogenetic relationships among members of the odonate genus Libellula (Odonata: Anisoptera: Libellulidae) were examined using 735 bp of mitochondrial COI and 416 bp of 16S ribosomal RNA gene sequences. Considerable debate exists over several relationships within Libellula, as well over the status of two putative genera often placed as subgenera within Libellula: Ladona and Plathemis. Parsimony and maximum-likelihood analyses of the separate and combined data sets indicate that Plathemis is basal and monophyletic and that Ladona is the sister clade to the remainder of Libellula sensu stricto (s.s.) (all species within the genus Libellula, excluding Plathemis and Ladona). Moreover, two European taxa, Libellula fulva and L. depressa, were found to occupy a sister group relationship within the Ladona clade. Relationships within Libellula s.s. are less well resolved. However, monophyletic lineages within the genus are largely consistent with morphologically based subgeneric classifications. Although tree topologies from each analysis differed in some details, the differences were in no case statistically significant. The analysis of the combined COI and 16S data yielded trees with overall stronger support than analyses of either gene alone. Several analyses failed to support the monophyly of Libellula sensu lato due to the inclusion of one or more outgroup species. However, statistical comparisons of topologies produced by unconstrained analyses and analyses in which the monophyly of Libellula was constrained indicate that any differences are nonsignificant. Based on morphological data, we therefore reject the paraphyly of Libellula and accept the outgroup status of Orthemis ferruginea and Pachydiplax longipennis. PMID- 11277630 TI - Recombination among multiple mitochondrial pseudogenes from a Passerine genus. AB - PCR products of a fragment of the mitchondrial protein coding subunit 5 of NADH dehydrogenase (ND5) from eight individuals representing five species of the South American bird genus Conirostrum were cloned. The 130 clones, which were subsequently sequenced, constituted 55 different sequences. Due to the observed differences in substitution patterns 58% of the cloned sequences were identified as pseudogenes. Recombination could be traced in 19% of the inferred nuclear pseudogenes, but this figure probably represents a significant underestimation of the factual recombination events. The nonrecombined pseudogenes consisted of multiple haplotypes found to diverge from 1 to 16% from the mitochondrial gene. The number of mitochondrial nuclear copies and their apparent frequent recombination suggest that pseudogenes constitute a serious potential risk in confounding phylogenetic studies and population genetic analysis. PMID- 11277631 TI - Perception and history: molecular phylogeny of a diverse group of neotropical frogs, the 30-chromosome hyla (anura: hylidae). AB - We used 16S rRNA, 12S rRNA, and cytochrome-b sequence to investigate the history of the "30-chromosome" Hyla, a diverse assemblage of neotropical treefrogs. Three aspects of these frogs were examined: (1) phylogenetic relationships among constituent species groups, among the species of one of these groups (Hyla leucophyllata group), and among populations of Hyla leucophyllata; (2) the apparent age of cladogenetic events; and (3) the phylogeography of H. leucophyllata. Mixed success in resolving the phylogeny is not because of a lack of character variation; levels of genetic divergence are high and suggest pre Pleistocene diversification, even among populations. Close temporal proximity of ancient cladogenetic events might make resolution of the topology difficult using any character set. At the population level, current geographic proximity is a poor predictor of phylogenetic affinity. A long history of dispersal and colonization may complicate, or even preclude, the accurate recovery of the history of this species in the Amazon Basin. It remains to be seen whether the patterns found here will prove common among neotropical frogs. PMID- 11277632 TI - A combined molecular approach to phylogeny of the jumping spider subfamily dendryphantinae (araneae: salticidae). AB - Four gene regions were sequenced for 30 species of jumping spiders, most from the subfamily Dendryphantinae, to investigate their molecular phylogeny and evolution. These are three regions from the mitochondria (ca. 560 bp of 16S plus adjacent tRNA, 1047 bp of cytochrome oxidase 1 (CO1), and 414 bp of NADH1 (ND1) and one region from the nuclear genome (ca. 750 bp of 28S). Parsimony and likelihood analyses of these gene regions separately and together support the monophyly of the dendryphantines as delimited previously by morphological characters. A group of elongate-bodied genera are placed as basal among the dendryphantines, and previously proposed relationships of Poultonella, Paraphidippus, and Sassacus vitis are confirmed. Comparison of overall rates of molecular evolution indicates striking differences across the gene regions, with highest divergence in ND1, CO1, 16S, and 28S in decreasing order. All four regions are characterized by both within- and among-site rate variation. Phylogenetic results from CO1 conflict conspicuously with phylogenetic results from the other genes and morphological data. Attempts to account for potential sources of this conflict (e.g., accommodating biased base composition, high homoplasy, within- and among-site rate variation, etc.) are largely unsuccessful. PMID- 11277633 TI - Phylogenetic utility of mitochondrial COI and nuclear Gpdh genes in Drosophila. AB - Phylogenetic utility of the mitochondrial COI (cytochrome oxidase subunit I) and nuclear Gpdh (glycerol-3-phosphate dehydrogenase) genes was studied in the Drosophila melanogaster species group. The rate of substitution was higher in the COI gene than in the Gpdh gene. In addition, multiple substitutions, not only for transitional but also for transversional substitutions, occurred faster in the COI gene. None of the trees obtained using the COI gene supported the well established monophyly of the ananassae subgroup. In addition, the incongruence length difference test, Templeton test, and partitioned Bremer support revealed that the trees based on the COI data are considerably different from those based on the Gpdh and the combined data set. Thus, the COI gene did not show good phylogenetic performance in the melanogaster group. The present analyses based on the Gpdh gene and the combined data set revealed that the ananassae subgroup branched off first in the melanogaster group followed by the montium subgroup and further by the melanogaster subgroup in contrast to the most recent phylogenetic hypothesis based on Amy multigenes. PMID- 11277634 TI - Genetically distinct populations in an Asian soldier-producing aphid, Pseudoregma bambucicola (Homoptera: Aphididae), identified by DNA fingerprinting and molecular phylogenetic analysis. AB - To estimate genetic structure of a soldier-producing aphid, Pseudoregma bambucicola, samples from natural populations throughout southeastern Asia were analyzed by a DNA fingerprinting technique. We unexpectedly found that P. bambucicola comprises two geographic groups, the northern group and the southern group, which are genetically distinct from each other but morphologically almost indistinguishable. Molecular phylogenetic and statistical analyses based on mitochondrial ribosomal DNA sequences demonstrated that the northern and southern groups of P. bambucicola are not closely related but constitute distinct lineages in the genus Pseudoregma. Detailed morphological reexamination revealed that the two groups could be distinguished by the number of setae on the 8th abdominal tergite of 1st instar nymphs and soldiers. From these results, it was suggested that P. bambucicola should be divided into two species. The northern group from Japan, Taiwan, Hong Kong, and northern Vietnam retains the name P. bambucicola, whereas we suggest that the name P. carolinensis (R. Takahashi, 1941, Tenthredo 3, 208-220) should be used for the southern group from Thailand, Malay Peninsula, Java, Irian Jaya, and Micronesia. The morphological resemblance between P. bambucicola and P. carolinensis might be due to shared ancestral characters of the genus Pseudoregma. PMID- 11277635 TI - Molecular phylogenetics and historical biogeography among salamandrids of the "true" salamander clade: rapid branching of numerous highly divergent lineages in Mertensiella luschani associated with the rise of Anatolia. AB - Phylogenetic relationships among salamandrids of the "true" salamander clade are investigated using 2019 aligned base positions (713 parsimony informative) of 20 mitochondrial DNA sequences from the genes encoding ND1 (subunit one of NADH dehydrogenase), tRNA(Ile), tRNA(Gln), tRNA(Met), ND2, tRNA(Trp), tRNA(Ala), tRNA(Asn), tRNA(Cys), tRNA(Tyr), and COI (subunit I of cytochrome c oxidase), plus the origin for light-strand replication (O(L)) between the tRNA(Asn) and the tRNA(Cys) genes. Parsimony analysis produces a robust phylogenetic estimate for the relationships of the major groups of "true" salamanders. Strong support is provided for the sister taxon relationship of Chioglossa and Mertensiella caucasica and for the placement of Salamandra and Mertensiella luschani as sister taxa. These relationships suggest two vicariant events between Europe and Anatolia caused by the formation of seaways in the Mediterranean Basin. Molecular divergence indicates an Early Miocene separation of Chioglossa and M. caucasica and a Late Miocene separation of Salamandra and M. luschani. The traditional phylogenetic hypothesis of a monophyletic Mertensiella is statistically rejected, indicating that southwestern and northeastern Anatolian populations have separate historical biogeographic origins. Therefore, we recommend placement of M. luschani in the genus Salamandra. Within M. luschani, six highly divergent lineages showing 7.6 to 10.1% pairwise sequence divergence are identified. Tests using four-taxon subsamples suggest that these lineages diverged nearly simultaneously in the Late Miocene, approximately 6 to 8 million years ago, when extensive uplifting of Anatolia occurred in response to the Arabian collision. PMID- 11277636 TI - Intraspecific phylogeography of Lacerta vivipara and the evolution of viviparity. AB - The lacertid lizard Lacerta vivipara is one of the few squamate species with two reproductive modes. We present the intraspecific phylogeny obtained from neighbor joining and maximum-parsimony analyses of the mtDNA cytochrome b sequences for 15 individuals from Slovenian oviparous populations, 34 individuals from western oviparous populations of southern France and northern Spain, 92 specimens from European and Russian viviparous populations, and 3 specimens of the viviparous subspecies L. v. pannonica. The phylogeny indicates that the evolutionary transition from oviparity to viviparity probably occurred once in L. vivipara. The western oviparous group from Spain and southern France is phylogenetically most closely related to the viviparous clade. However, the biarmed W chromosome characterizing the western viviparous populations is an apomorphic character, whereas the uniarmed W chromosome, existing both in the western oviparous populations and in the geographically distant eastern viviparous populations, is a plesiomorphic character. This suggests an eastern origin of viviparity. Various estimates suggest that the oviparous and viviparous clades of L. vivipara split during the Pleistocene. Our results are discussed in the framework of general evolutionary models: the concept of an oviparity-viviparity continuum in squamates, the cold climate model of selection for viviparity in squamates, and the contraction-expansion of ranges in the Pleistocene resulting in allopatric differentiation. PMID- 11277637 TI - A molecular biogeographic analysis of the relationship between North American melanoploid grasshoppers and their Eurasian and South American relatives. AB - The Melanoplinae constitute one of the two largest subfamilies of Acrididae. Distributed mainly throughout the New World and parts of Eurasia, this group of grasshoppers includes over 100 genera and 800 species. Over the past five decades there has been considerable speculation on the origins of North and South American taxa. The most favored hypothesis proposes an ancient division of Laurasian taxa accompanying the separation of North America and Eurasia, with subsequent radiations within those continents, followed by a recent incursion of Nearctic melanoplines into the southern hemisphere with the establishment of the Isthmus of Panama. This research tests that scenario by phylogenetic analysis using as characters portions of five mitochondrial gene sequences, totaling 2285 bp. Three tree-building methods, maximum-parsimony, neighbor-joining, and maximum likelihood, strongly support the different view that melanopline grasshoppers originated somewhere in the Americas and spread to the Old World. The feasibility of these findings is discussed within a geological context. PMID- 11277638 TI - Evolutionary assembly of the milkweed fauna: cytochrome oxidase I and the age of Tetraopes beetles. AB - The insects that feed on the related plant families Apocynaceae and Asclepiadaceae (here collectively termed "milkweeds") comprise a "component community" of highly specialized, distinctive lineages of species that frequently sequester toxic cardiac glycosides from their host plants for defense against predators and are thus often aposematic, advertising their consequent unpalatability. Such sets of specialized lineages provide opportunities for comparative studies of the rate of adaptation, diversification, and habitat related effects on molecular evolution. The cerambycid genus Tetraopes is the most diverse of the new world milkweed herbivores and the species are generally host specific, being restricted to single, different species of Asclepias, more often so than most other milkweed insects. Previous work revealed correspondence between the phylogeny of these beetles and that of their hosts. The present study provides analyses of near-complete DNA sequences for Tetraopes and relatives that are used to establish a molecular clock and temporal framework for Tetraopes evolution with their milkweed hosts. PMID- 11277639 TI - Evolution of mitochondrial and ribosomal gene sequences in anophelinae (Diptera: Culicidae): implications for phylogeny reconstruction. AB - In this study, two mitochondrial genes, cyt b and ND5, and the D2 expansion segment of the 28S nuclear ribosomal gene were used to reconstruct a phylogeny of the mosquito subfamily Anophelinae. The ingroup consisted of all three genera of Anophelinae and five of six subgenera of Anopheles. Six genera of Culicinae were used as the outgroup. Extreme conservation at the protein level coupled with rapid saturation of synonymous positions probably accounted for the lack of meaningful phylogenetic signal in the cyt b gene. In contrast, abundant variation at all codon positions of the ND5 gene allowed recovery of the basal and most of the recent relationships. Phylogenetic analysis of D2 produced results consistent with those of ND5. Combined analysis indicated well-supported monophyletic Anophelinae (with Chagasia basal), Anopheles + Bironella, and subgeneric clades within the genus Anopheles. Moreover, subgenera Nyssorhynchus and Kerteszia were supported as a monophyletic lineage. The Kishino-Hasegawa test could not reject the monophyly of Anopheles, whereas the recently proposed hypothesis of close affinity of Bironella to the subgenus Anopheles was rejected by the analyses of ND5 and combined data sets. The lack of resolution of Bironella and Anopheles clades, or basal relationships among subgeneric clades within Anopheles, suggests their rapid diversification. Recovery of relationships consistent with morphology and previous molecular studies provides evidence of substantial phylogenetic signal in D2 and ND5 genes at levels of divergence from closely related species to subfamily in mosquitoes. PMID- 11277640 TI - Molecular phylogeography of the cosmopolitan bryozoan Celleporella hyalina: cryptic speciation? PMID- 11277641 TI - Second-line therapy for potentially platinum-sensitive recurrent ovarian cancer: what is optimal treatment? PMID- 11277642 TI - Carboplatin alone vs carboplatin plus epidoxorubicin as second-line therapy for cisplatin- or carboplatin-sensitive ovarian cancer. AB - OBJECTIVE: The aim of the study was to analyze the benefit/toxicity profile of a second-line treatment with carboplatin alone or carboplatin plus another non cross-resistant drug (epidoxorubicin) in ovarian cancer patients sensitive to cisplatin-based chemotherapy at first-line treatment. METHODS: We conducted a randomized clinical trial. Women with epithelial ovarian cancer FIGO Stage II--IV who had a complete or partial response to first-line treatment with cisplatin or carboplatin-based regiments and subsequently progressed or relapsed more than 6 months after discontinuation of first-line treatment were eligible for the study. A total of 190 subjects entered the study. They were randomly allocated to either 300 mg/m(2) of carboplatin every 28 days for five cycles (95 patients) or 120 mg/m(2) of epidoxorubicin and 300 mg/m(2) of carboplatin every 28 days for five cycles (95 patients). RESULTS: A complete response was reported, respectively, in 32 (36%) women allocated to carboplatin alone and in 28 (31.8%) of those allocated to carboplatin plus epidoxorubicin. The corresponding figures for partial response were 18 (20.2%) and 26 (29.9%). Comparing the frequency of complete response, partial response, no change, and progression, the differences between the two groups were not significant (chi(2)(3) 5.10, P = 0.16). The median duration of response was 16 months in the carboplatin alone and 20 months in the carboplatin plus epidoxorubicin group (P = not significant). The 3-year percentage of survival was 29% in the carboplatin alone and 42% in the carboplatin plus epidoxorubicin group; this difference was not statistically significant. The frequency of leukopenia, anemia, and thrombocytopenia grade 3-4 was higher in the epidoxorubicin plus carboplatin than in the carboplatin alone group. Alopecia G3 was present in 88% of women treated with epidoxorubicin plus carboplatin. CONCLUSIONS: The general results of this study do not show any marked differences in response to second-line treatment among women treated with single-agent (carboplatin) or multiagent (carboplatin plus epidoxorubicin) schedules. Toxicity, particularly hematological, was more relevant in women treated with the multiagent schedule. PMID- 11277643 TI - Characterization and tumorigenicity of human ovarian surface epithelial cells immortalized by SV40 large T antigen. AB - OBJECTIVES: Epithelial ovarian cancers are considered to arise from neoplastic transformation of the ovarian surface epithelium (OSE). However, the earliest events in ovarian carcinogenesis have not been clearly defined because patients are often diagnosed in the advanced stages and useful in vivo and in vitro experimental systems using human OSE cells are lacking. We aimed to improve the availability of experimental models for the study of human ovarian carcinogenesis. METHODS: Subcultured human OSE cells were transfected with SV40 large T antigen. Resulting OSE cell lines were characterized using immunocytochemistry and tested tumorigenicity. RESULTS: Six immortalized OSE cell lines were obtained. All cell lines essentially retained the original morphological features of normal OSE cells and showed higher proliferation rates and saturation density. Although they were all nontumorigenic in athymic mice, OSE2b-2 sv cells, which were selected in soft agar from colonies of an SV40 large T antigen-expressing transfectant, OSE2b sv, produced tumors on the peritoneal surface, mesothelium, and diaphragm and induced ascites after being injected intraperitoneally. Solid tumors also grew when mice were inoculated subcutaneously. The tumor cells were formed in a solid sheet arrangement and no evidence of glandular or squamous differentiation was present. They were weakly immunostained with an antibody against cytokeratin, and intercellular junctions resembling attachment devices were ultrastructurally present between cells. The tumors were histologically diagnosed as undifferentiated carcinomas. CONCLUSIONS: The established cell lines may provide a model system to investigate the mechanisms of cytogenic and molecular changes from normal OSE cells through the various steps of transformation. PMID- 11277644 TI - Evaluation of chemoresistance markers in women with epithelial ovarian carcinoma. AB - OBJECTIVE: Preliminary studies have suggested that lung resistance protein (LRP), multidrug resistance protein (MRP), and p27 may be useful markers of chemoresistance. Our goal was to evaluate the expression of LRP, MRP, and p27 in normal ovaries and chemosensitive and chemoresistant ovarian carcinomas. METHODS: Fourteen women with normal ovaries and fifty women with epithelial ovarian carcinoma who underwent cytoreductive surgery from 1996 through 1998 had specimens stained with immunocytochemistry for LRP, MRP, and p27. All women received paclitaxel/platinum-based chemotherapy. Twenty-nine women had a disease free survival (DFS) of at least 12 months after completion of chemotherapy (sensitive) and 21 women had persistent disease during treatment (resistant). RESULTS: Evaluation of LRP expression revealed significant differences between the normal ovaries and cancers in both the epithelial (57% vs 90%, P = 0.03) and stromal (86% vs 32%, P = 0.001) components. Evaluation of MRP expression revealed significant differences between normal ovaries and cancers in the epithelial component (7% vs 66%, P = 0.001) but not in the stromal component (14% vs 4%, P = 0.1). Evaluation of p27 revealed significant reductions in expression in cancers compared with normal ovaries for both the epithelial (90% vs 55%, P = 0.02) and stromal (88% vs 5%, P = 0.001) components. Comparison between the chemosensitive and chemoresistant groups revealed no significant differences in expression of LRP and MRP, in either the epithelial or stromal component, but significantly lower levels of p27 were expressed in the epithelial component of the chemoresistant group (P = 0.01). CONCLUSIONS: The expression of LRP, MRP, and p27 is significantly different in ovarian cancers compared with normal ovaries. Low levels of p27 expression are associated with chemoresistance; however, LRP and MRP expression are not prognostic for chemosensitivity. PMID- 11277645 TI - Epithelial ovarian carcinoma and European birthplace of grandparents. AB - OBJECTIVE: The aim of this study was to determine whether the risk of ovarian carcinoma was related to latitude or to genetically based patterns of European geographic origin. PATIENTS AND METHODS: We studied the countries of origin of European-born grandparents of 168 newly diagnosed patients in two hospitals in New York City, compared with 159 controls from similar neighborhoods. We measured the risk of this cancer associated with having one or more white, non-Jewish grandparents born in North Europe versus none or in South Europe versus none. We also classified geographic origins in other ways to reflect the two main trends in genetic variations between Europeans mapped by Cavalli-Sforza et al. (The History and Geography of Human Genes, Princeton University Press, Princeton, 1994). Unconditional logistic regression was used to control for age, parity, years of use of oral contraception, age at menarche, education, Catholic religion, and area of residence and for numbers of Jewish grandparents, siblings, and first-degree relatives with breast or ovarian cancer. RESULTS: Approximately half of the subjects had least one white, non-Jewish grandparent born in Europe. There was no significant effect of ancestral latitude: among women born in the United States the odds ratio (OR) and 95% confidence limits associated with North European ancestry were 0.87 (0.47--1.63) compared with a reference group of women with no such ancestry. The corresponding OR for South Europe was 0.73 (0.39- 1.74). Using the genetically based classifications of countries of origin, however, we found significant differences between cases and controls; ancestries from North West Europe and those from countries concentrically near Spain showed lower risks of ovarian carcinoma. CONCLUSIONS: The results support the hypothesis that the previously observed effects of latitude must act through environmental effects or through gene-environment interactions. Other variations in risk related to geographic origins are consistent with known patterns of genetic differences, but require confirmation in larger, population-based studies. PMID- 11277646 TI - Significance of "normal" endometrial cells in cervical cytology from asymptomatic postmenopausal women receiving hormone replacement therapy. AB - OBJECTIVE: The aim of this study is to assess the significance of normal endometrial cells identified in screening Pap smears from asymptomatic postmenopausal women taking hormone replacement therapy (HRT). METHODS: Endometrial histology reported from 93 asymptomatic postmenopausal women receiving HRT noted to have normal endometrial cells on a screening Pap smear was reviewed. Information regarding HRT, endometrial sampling, and interval between the sentinel Pap and sampling was extracted from the record. RESULTS: Endometrial biopsies were obtained an average of 1.7 months after the Pap smears. Eighteen of the ninety-three histology specimens (19%, 95% CI: 12--27%) identified abnormalities, in four cases precancerous or cancerous lesions. These 18 abnormalities included 7 endometrial polyps; 7 cases of simple hyperplasia, 1 with atypia; 3 cases of complex hyperplasia, 1 with atypia; and 1 endometrial adenocarcinoma. CONCLUSION: Normal endometrial cells identified on a screening Pap smear in this population may be an indication of endometrial disease. PMID- 11277647 TI - Characteristics and survival of cervical cancer patients managed at adjacent urban public and university medical centers. AB - OBJECTIVE: The goal of this work was to compare characteristics and survival of cervical cancer patients at adjacent public and university hospitals to define the effects of poverty and ethnicity on disease. METHODS: A retrospective chart review was conducted of cervical cancer patients managed by gynecologic oncologists at two adjacent urban hospitals between 1992 and 1998. Continuous variables were compared by t test, categorical variables by chi(2), and survival by the Kaplan-Meier and log-rank methods. RESULTS: In all, 372 patients were identified, with 209 (56%) at the public hospital and 163 (44%) at the adjacent university hospital. Ethnic distribution differed between the two hospitals: 100 (52%) versus 46 (28%) African-American, 56 (29%) versus 13 (8%) Hispanic, 31 (16%) versus 96 (60%) Caucasian, and 5 (3%) versus 6 (4%) other (P < 0.001). In addition, public hospital patients presented with more advanced cancers (stages II--IV) than those managed at the university hospital, 96 (48%) versus 53 (34%) (P = 0.008), and squamous cancers were more common at the public hospital, 154 (89%) versus 120 (76%) (P = 0.03). However, with a median follow-up of 17 months, stage-adjusted survival did not differ between the two institutions. CONCLUSIONS: The higher proportions of advanced and squamous cervical cancers encountered at the public hospital likely reflect suboptimal screening. Equal access to gynecologic oncologists eliminated disparities in stage-adjusted survival. Efforts at earlier diagnosis should be directed at indigent, especially minority women. PMID- 11277648 TI - Detection of human papillomaviruses in cervical neoplasias using multiple sets of generic polymerase chain reaction primers. AB - OBJECTIVE: The aim of this study was to evaluate precisely the differences in the spectra of human papillomavirus (HPV) types detected by different generic primer pairs commonly used for detection of this extraordinarily heterogeneous virus. METHODS: Three sets of polymerase chain reaction (PCR) primers for the L1 open reading frame (ORF) and two sets for E6/E7 ORFs were used to detect HPVs in DNAs from 107 cervical tissues, including 77 cervical neoplasias. HPV types were determined by analysis of restriction fragment length polymorphisms (RFLPs) and nucleotide sequencing. RESULTS: A high overall detection rate of HPV in cervical neoplasias (76/77, 98.7%) was achieved by polymerase chain reaction (PCR) amplification with multiple sets of generic primers, while the detection rate for each individual primer pair varied from 48/77 (62%) to 70/77 (91%). Only in 34 of 77 cases (44%) were HPV DNAs positive for all sets of primer pairs. Further determination of HPV types by RFLPs and nucleotide sequencing showed inconsistencies between the PCR primer pairs used. CONCLUSION: Our study revealed that the HPV detection rate is critically affected by the choice of PCR primers, and that appropriate use of combinations of generic PCR primer sets followed by RFLP analyses is both necessary and sufficient for typing most HPVs in cervical lesions. More precise methods such as sequencing would be necessary in only a few cases. PMID- 11277649 TI - Concomitant radiotherapy and paclitaxel for high-risk endometrial cancer: first feasibility study. AB - OBJECTIVE: Postoperative radiotherapy (RT) is the most used adjuvant treatment in high risk endometrial cancer (HREC), and it appears to reduce the incidence of pelvic relapses but doesn't seem to improve survival. Paclitaxel (P) has shown in vitro and clinical activity against endometrial cancer, and it is also a potent radiosensitizer by blocking dividing cells in G2/M phase. This is the first study that verifies the feasibility of a treatment with concomitant weekly chemotherapy and RT to potentially reduce the incidence of local and distant relapses in order to improve survival in HREC. PATIENTS AND METHODS: Thirteen patients with HREC have entered the feasibility study at San Raphael Hospital University of Milan. All patients underwent total abdominal hysterectomy, bilateral salpingo oophorectomy, and surgical staging. Four patients presented stage IC disease, 2 women had IIB stage tumors, 5 patients revealed IIIA stage disease, and 2 had stage IIIC. The patients received P (60 mg./m(2)) via a continuous 1-h infusion once weekly during the 5 weeks of RT (mean radiation dose of 50.4 Gy). At the end of RT three additional consolidation courses of P (80 mg/m(2)) were subministered. Eleven patients received only pelvic irradiation; in 2 cases radiotherapy was performed on an extended field. RESULTS: Eleven of the 13 enrolled patients have completed the radiochemotherapy regimen. A total of 100 courses of P were performed. All patients completed the RT. Adverse effects were evaluated. Hematological toxicity was mild: four cycles (4%) were delayed 1 week because of grade 1 neutropenia. No severe thrombocytopenia was identified. No hemotrasfusions were performed. One cycle was delayed for fever. Gastrointestinal adverse effects were observed in 2 patients, in which the cycles were delayed 1 week because of diarrhea. One cycle was delayed 1 week because of dermatitis. One patient developed a subocclusion 8 weeks after the end of the treatment, with medical resolution. No patients developed hypersensitivity reactions. CONCLUSIONS: Concomitant P and RT is safe and acceptable treatment in patients with HREC. Prospective clinical studies are necessary to evaluate the benefits of this regimen for the long-term survival rate. PMID- 11277650 TI - Phase II study of vinorelbine in the treatment of platinum-resistant ovarian carcinoma. AB - OBJECTIVE: The purpose of this phase II study was to evaluate on an intent-to treat basis the activity and toxicity of single-agent vinorelbine (VRL) as second line chemotherapy of patients with platinum-resistant ovarian cancer. Platinum resistant disease was defined as disease refractory to or relapsing within 12 months after finishing platinum-containing chemotherapy. METHODS: VRL (30 mg/m(2)) was administered intravenously as a bolus injection days 1 and 8 every 21 days. Initially, four courses of VRL were given. Patients with responding or stable disease received four more courses of VRL to a maximum of eight courses. RESULTS: Twenty-eight of 33 eligible patients were considered evaluable for response. The overall response rate was 21% (7/33) (95% CI: 7--35). Median time to progression was 3.1 months and median survival was 10.1 months. Toxicity was generally mild. Leukopenia was the dose-limiting toxicity. CALGB grade III/IV infection was observed in 15/0% of patients. The most important nonhematologic toxicities were nausea and constipation. Grade III/IV nausea was observed in 6/0% and grade III/IV constipation in 3/3% of patients. Peripheral neurotoxicity was only a minor problem with no grade III/IV toxicity. No patients stopped treatment because of toxicity and no toxic death was reported. CONCLUSION: VRL was generally well tolerated, but the activity in platinum-resistant ovarian cancer was only modest, although fully comparable to other second-line treatments. Further studies are required to define the role of VRL in combination chemotherapy for ovarian cancer. PMID- 11277651 TI - Ultrasound, physical examination, and CA 125 measurement for the detection of recurrence after conservative surgery for early borderline ovarian tumors. AB - BACKGROUND: Borderline ovarian tumors often affect women of childbearing age and the prognosis is outstanding. Given the young age of several patients and the good prognosis, fertility-sparing surgery is considered adequate for stage I tumors. However, women treated conservatively have a relatively small but well defined risk of recurrence and no study has specifically addressed the optimal follow-up technique. METHODS: From 1981 to 1997, 164 women underwent fertility sparing surgery for stage I borderline ovarian tumor and were followed prospectively. After surgery all women underwent physical examination and ultrasound examination every 3 months for 2 years after first diagnosis and every 6 months thereafter. Measurement of serum CA 125 levels was planned every 6 months in patients with a serous tumor. RESULTS: With a median follow-up of 71, months 28 women treated with fertility-sparing surgery (28/164 = 17%) had either recurrence of borderline tumor (23) or recurrence with carcinoma. Complete details of follow-up procedures are available for 24 women and they represent the study population. An abnormal adnexal mass was detected in 18 of 19 women with recurrent borderline tumor. One patient had diagnosis due to persistent free fluid. All five women with invasive carcinoma had diagnosis of a complex adnexal mass. Gynecologic examination was suspicious (palpable mass) in 7 cases and obviously abnormal (large mass or nodules) in another 7. CA 125 serum levels were elevated in 8 cases. CONCLUSION: Transvaginal ultrasound is currently the most effective diagnostic technique for the follow-up of young patients treated conservatively for early borderline ovarian tumor. PMID- 11277652 TI - Flow cytometric and clinicopathologic study of complete hydatidiform moles with special reference to the significance of cytometric aneuploidy. AB - OBJECTIVE: As complete hydatidiform moles (CMs) have been studied less with respect to aneuploidy and its clinical implications, the significance of cytometric aneuploidy in CMs was evaluated. METHODS: Two hundred thirty-nine CMs were studied clinicopathologically and analyzed by flow cytometry using formalin fixed paraffin-embedded tissues. RESULTS: Of 239 CMs, 182 were diploid, 30 were tetraploid, and 27 were aneuploid (nontriploid/tetraploid aneuploid). There were no significant histologic differences among the diploid, tetraploid, and aneuploid CMs. Persistent disease developed in 20 of 114 CMs (17.6%) (16 of 77 diploid, 4 of 18 tetraploid, and none of 19 aneuploid CMs). Eight diploid and three tetraploid CMs were invasive, and one patient each with diploid CM and tetraploid CM developed choriocarcinoma and none of 19 patients with aneuploid CMs had sequelae. CONCLUSION: These results suggest that aneuploid CMs are associated with less risk for persistent disease than diploid or tetraploid CMs. DNA ploidy status may be an independent predictor of persistent disease. PMID- 11277653 TI - Presence of aberrant tumor-reactive immunoglobulins in the circulation of patients with ovarian cancer. AB - OBJECTIVES: Cancer patients generally exhibit circulating tumor-reactive immunoglobulins; however, these antibodies fail to eradicate tumors or prevent their progression. This study identifies and characterizes an aberrant tumor reactive IgG population present in women with ovarian cancer. METHODS: In this pilot study, IgG was isolated from the sera of women with advanced-stage ovarian cancer (stages III and IV, n = 62) and age-matched female volunteers (n = 50) by affinity chromatography. These IgGs were characterized on the basis on their aberrant binding to concanavalin A affinity columns. Subsequently, the concanavalin A-binding moiety was localized following IgG fragmentation, analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and characterized by oligosaccharide profiling. RESULTS: The level of concanavalin A-binding IgG in our control population was 8.9 +/- 2.9%, whereas in ovarian cancer patients, the level of concanavalin A-binding IgG was 38.8 +/- 7.4%. In the patients with ovarian cancer, 87.5 +/- 5.7% of the tumor-reactive IgG was demonstrated to be concanavalin A-binding. Based on oligosaccharide profiling of the fragmented concanavalin A-binding IgG, the aberrant lectin binding appeared to be the consequence of altered glycosylation of one of the two Fc chains. CONCLUSIONS: While our previous studies have identified the presence of circulating IgG reactive with specific tumor-associated antigens and its association with poor prognosis, this report demonstrated the presence of an aberrantly glycosylated IgG population in cancer patients. This altered IgG appeared to be the primary class of tumor-reactive antibodies in these women. PMID- 11277655 TI - Multimodality therapy in early-stage neuroendocrine carcinoma of the uterine cervix. AB - OBJECTIVE: Patients with early-stage neuroendocrine cervical carcinoma (NECC) have a high mortality rate despite aggressive therapy. The rarity of this tumor precludes initiation of a randomized, prospective trial. We reviewed our experience in early stage disease and performed a meta-analysis of the literature to identify prognostic factors and determine optimal multimodality therapy. METHODS: Eleven women with International Federation of Gynecology and Obstetrics (FIGO) early stage (IB--IIA) NECC were treated with surgery and chemotherapy at our institutions between 1978 and 1998. Administration of radiation therapy was recorded, but not required for inclusion in this study. A gynecologic pathologist reviewed all histopathologic sections. Medical records were retrospectively reviewed and clinical data obtained. Twenty-three early-stage NECC patients who were similarly treated during the study interval were identified by a Medline search of the English literature and included in the analysis. The Kaplan--Meier method and log-rank test were used for survival analysis. RESULTS: The overall 2 year survival rate for the 34 patients was 38%. The median age was 37 years (range, 20--75 years). Median cervical tumor diameter was 3.2 cm (range 0.5--11.0 cm). Lymphovascular space invasion was present in 21 (78%) of 27 patients (7 unknown). Fifteen (52%) of twenty-nine had lymph node metastases (5 unknown). Fifteen patients received postoperative platinum/etoposide (PE), seven received vincristine/adriamycin/cyclophosphamide (VAC), two received alternating cycles of VAC and PE, and ten received other chemotherapy regimens. Twenty women were treated with radiation therapy. The presence of lymph node metastases was a poor prognostic factor (P < 0.001). PE and VAC chemotherapy was associated with increased survival (P < 0.01). CONCLUSION: NECC is a highly lethal variant of cervical cancer. The presence of lymph node metastases is the most important prognostic variable. Postoperative VAC or PE appears most likely to improve chances for survival. PMID- 11277654 TI - Complications associated with intraperitoneal chemotherapy catheters. AB - PURPOSE: The goal of this work was to determine the complication rate and any predisposing risk factors associated with subcutaneous intraperitoneal (ip) catheters used in the treatment of patients with advanced ovarian cancer. METHODS: We retrospectively reviewed the charts of 301 patients who had a subcutaneous Bardport catheter placed for administration of ip chemotherapy at Memorial Sloan--Kettering Cancer Center (MSKCC) from December 1989 to May 1997. RESULTS: Thirty (10%) patients were identified as having catheter-related complications, with 19 (6.3%) experiencing inflow obstruction and 11 (3.6%) experiencing infection. Only 21 of 301 (7%) required cessation of chemotherapy prior to its expected completion, with 14 (4.6%) occurring in the malfunction group and 7 (2.3%) in the infection group. Three hundred thirteen patients received an ip catheter; however, 12 patients who received their ip chemotherapy elsewhere were excluded when determining the complication rate. Overall, 218 of 313 (69.6%) catheters were placed at the time of laparotomy, 61 of 313 (19.5%) catheters were placed at the time of laparoscopy, and 34 of 313 (10.9%) were placed as a separate procedure. In the malfunction group, 18 of 19 (94.7%) patients had their catheters placed at the time of laparotomy, none were placed at the time of laparoscopy, and 1 of 19 (5.3%) was placed as a separate procedure. In the infection group, 8 of 11 (72.7%) catheters were placed at laparotomy, 2 of 11 (18.3%) were placed at the time of laparoscopy, and 1 of 11 (9.0%) was placed as a separate procedure. Complications occurred in 3 of 54 (5.5%) patients who received platinum alone, 11 of 134 (8.2%) who received platinum in combination, 2 of 43 (4.7%) who received paclitaxel alone, 13 of 61 (21.3%) who received mitoxantrone alone or in combination, and 1 of 9 (11.1%) who received other regimens. CONCLUSION: Subcutaneous ip catheters are associated with a lower rate of catheter-related complications than previously reported, perhaps due in part to both avoiding insertion of ip catheters at the time of bowel surgery and placing ip catheters at the time of laparoscopy. PMID- 11277656 TI - Compliance with and acute hematologic toxic effects of chemoradiation in indigent women with cervical cancer. AB - OBJECTIVES: The goals of this work were to describe the compliance with and acute hematologic toxic effects of chemoradiation for cervical cancer in indigent women and to explore the likelihood that chemoradiation is effective outside research settings. We hypothesized that if compliance and toxicity are not limiting in this high-risk group of patients, the effectiveness of chemoradiation for cervical cancer in community settings is likely to mirror the efficacy demonstrated in clinical trials. METHODS: This study is a retrospective review of prospectively maintained data on women with newly diagnosed cervical cancer treated with chemoradiation between August 1998 and August 2000. Cisplatin was given weekly at 40 mg/m(2) to a maximum of six courses. A WBC count <3000/mm(3) resulted in cancellation of cisplatin but not radiation, and patients were transfused for hemoglobin <9 g/dl. Statistical analysis was performed using the t test, chi(2) test, and Fisher's exact test. RESULTS: In all, 19 of 65 patients treated (29.2%) missed at least one chemotherapy cycle, with 10 (15.4%) due to missed appointments, 8 (12.3%) due to a low WBC count, and 1 due to increased creatinine. Nineteen patients (29.2%) received RBC transfusion during chemoradiation, and two (3%) had platelets <75,000/mm(3). Noncompliant patients had a lower mean total point A dose (7986 cGy vs 8413 cGy, P = 0.04) and longer overall treatment duration (79 days vs 51 days, P < 0.001). No patient had a fatal hematologic complication. CONCLUSION: Nearly a third of the indigent women treated with chemoradiation for cervical cancer do not complete the prescribed treatment, and a similar number require blood transfusions. In indigent and minority women, the effectiveness of chemoradiation protocols may not mirror the efficacy obtained in clinical trials. PMID- 11277657 TI - The role of cytoreductive surgery in the management of stage IV uterine papillary serous carcinoma. AB - OBJECTIVE: The aim of this study was to evaluate the survival impact of cytoreductive surgery and other prognostic determinants in patients with Stage IV uterine papillary serous carcinoma (UPSC). METHODS: All patients with FIGO Stage IV UPSC diagnosed between January 1, 1989 and December 31, 1998 were identified from tumor registry databases. Individual patient data were collected retrospectively. Survival analysis and comparisons were performed using the method of Kaplan and Meier, the log-rank test, and the Cox proportional hazards regression model. Predictors of surgical outcome were evaluated using the log rank test. RESULTS: Thirty-one patients underwent primary cytoreductive surgery for Stage IV UPSC (median age, 65 years). The median survival for all patients was 14.4 months. Optimal cytoreduction was defined as residual disease < or =1 cm in maximal diameter. The only significant predictor of a suboptimal surgical outcome was the presence of disease in three or more anatomic regions. Overall, 16 of 31 patients (51.6%) completed primary surgery with optimal disease status. Optimal cytoreduction was associated with a median survival of 26.2 months, compared with 9.6 months for patients left with suboptimal residual disease (P < 0.001). At 24 months, 57.1% of optimally cytoreduced patients were still alive, compared with just 6.7% of patients left with suboptimal disease. Furthermore, patients with only microscopic residual tumor had a significantly longer median survival (30.4 months) than both patients with 0.1- to 1.0-cm residual disease (20.5 months) and those left with suboptimal disease (P = 0.004). Postoperative platinum-based chemotherapy was associated with a median survival of 17.1 months, compared with 9.5 months without such therapy (P = 0.018). Patients receiving the combination of platinum + paclitaxel had a median survival rate of 29.1 months versus 14.4 months for patients receiving platinum + cyclophosphamide +/- doxorubicin (P = 0.054). On multivariate analysis, the only statistically significant predictor of survival was the cytoreductive surgical outcome. CONCLUSIONS: The strongest predictor of overall survival for patients with Stage IV UPSC was the amount of residual disease following surgery. Recommended management for this group of patients should consist of maximal surgical cytoreduction followed by platinum-based chemotherapy, preferably in combination with paclitaxel. PMID- 11277658 TI - Routes of lymphatic spread: a study of 112 consecutive patients with endometrial cancer. AB - OBJECTIVE: The goal of this work was to assess different patterns of lymphatic spread to pelvic and para-aortic lymph nodes (LNs) in endometrial cancer as a function of the location of tumor within the uterus. METHODS: Between 1984 and 1999, 625 patients with endometrial cancer were managed with hysterectomy and node dissection at our institution. The present study includes the 112 (18%) patients who had positive pelvic and/or para-aortic LNs; 41 (37%) of them had cervical involvement. RESULTS: The external iliac was the most commonly involved pelvic LN site both in patients with tumor limited to the corpus and in those with cervical invasion. Isolated pelvic LN metastases to a single site were more frequently observed in external iliac LNs and obturator LNs in patients with tumor confined to the uterine corpus, whereas they occurred more commonly in external iliac and common iliac LNs in patients with cervical involvement. Metastasis to the common iliac LNs was more frequent in patients with disease extension to the cervix. In fact, common iliac LNs were positive in 67% of patients with cervical invasion, compared with only 30% of those with tumor confined to the uterine corpus (P < 0.01). Para-aortic LN invasion was significantly associated with obturator LN status. In fact, para-aortic LNs were positive in 64% of patients with positive obturator LNs compared with 23% of patients with negative obturator LNs (P = 0.01). All patients with positive para aortic LNs and tumor invading the cervix had positive common iliac LNs. By contrast, when tumor was limited to the corpus, common iliac LNs were involved in only 27% of patients with positive para-aortic LNs. CONCLUSION: External iliac LNs are the most commonly involved LNs in endometrial cancer. Compared with carcinomas limited to the uterine corpus, endometrial cancers invading the cervix spread more readily to the common iliac LNs. Furthermore, these data suggest that para-aortic LN metastases spread via a route shared by the common iliac LNs when tumor involves the cervix but spread predominantly via a route common to the obturator LNs (and/or external iliac LNs) when the primary tumor site is the corpus only. PMID- 11277659 TI - Scalp and cranial bone metastasis of endometrial carcinoma: a case report and literature review. AB - OBJECTIVE: The aim of this study is to report a rare case of scalp and cranial bone metastasis of endometrial carcinoma and review the literature. METHOD: We report a 45-year-old multiparous woman with FIGO Stage 1A Grade II endometrial adenocarcinoma who presented 3 years following total abdominal hysterectomy and bilateral salpingo-oophorectomy with scalp and cranial bone metastasis. Similar cases in the literature are reviewed. RESULTS: The patient metastatic workup revealed local, distant, scalp, and cranial bone metastasis. She died within 6 months. The poor prognosis is similar to that of six other cases reviewed. CONCLUSION: Scalp and cranial bone metastasis following endometrial carcinoma is extremely rare. It is a reflection of a widely disseminated disease and poor prognosis. However, single bone metastasis has a better postmetastatic survival with the help of local radiotherapy than scalp and multiple bone metastasis. PMID- 11277660 TI - Recurrent metastatic fallopian tube carcinoma in pregnancy. AB - BACKGROUND: Fallopian tube cancer is the rarest of all gynecologic cancers. An extensive literature search on Medline reveals no previous case reports of fallopian tube carcinoma in association with a term pregnancy. CASE: A woman with surgical stage IIB fallopian tube carcinoma was treated with limited staging laparotomy, as per the patient's fertility wishes, followed by adjuvant cis platinum and paclitaxel (Taxol). One year following chemotherapy, she conceived. She was noted to have an asymptomatic intraabdominal recurrence at 16 weeks. The patient completed 37 weeks of pregnancy without further therapy according to her wishes. She subsequently underwent a cesarean section with optimal tumor reduction surgery. Carboplatin and paclitaxel were reinstituted, achieving partial response. She is presently alive with stable disease status 6 months after completing her salvage chemotherapy. CONCLUSION: This is the first case report of recurrent fallopian tube cancer in pregnancy. PMID- 11277661 TI - Management of recurrent juvenile granulosa cell tumor of the ovary. AB - BACKGROUND: Juvenile granulosa cell tumors of the ovary are a rare form of neoplasm that makes up less than 5% of ovarian tumors in childhood and adolescence. About 90% are diagnosed in stage I with a favorable prognosis. More advanced stages (FIGO stages II--IV) have a poor prognosis. CASE: A patient was initially diagnosed at age 17 with FIGO stage IIIC disease and treated with a right salpingo-oophorectomy, debulking, and staging followed by six cycles of carboplatin and etoposide chemotherapy. Tumor recurrence in the liver and adjacent to the spleen occurred 13 months after completion of primary therapy. Aggressive surgical removal of tumor followed by six cycles of bleomycin and taxol as salvage chemotherapy resulted in 44 months of disease-free survival. On November 27, 2000, she had a cesarean delivery of a 2335-g normal male due to a breech presentation. Exploration revealed no evidence of tumor. CONCLUSION: This is the second case report of a patient with advanced juvenile granulosa cell tumor to become pregnant after apparently successful chemotherapy. These results are encouraging, but the best treatment for extensive and recurrent disease has yet to be determined. PMID- 11277662 TI - Neuroendocrine small cell carcinoma of the uterine cervix showing polypoid growth and complicated by pregnancy. AB - BACKGROUND: Neuroendocrine small cell carcinoma of the uterine cervix is an aggressive disease, and it rarely is complicated by pregnancy. CASE: A polypoid tumor was found in the uterine cervix in a 27-year-old Japanese woman at 27 weeks of gestation. No polyp had been detected at 14 weeks of gestation. The polyp was excised and diagnosed as neuroendocrine small cell carcinoma by histological examination, including Grimelius, neuron-specific enolase, and chromogranin staining. A healthy infant was born by cesarean section at 29 weeks of gestation: this was followed by radical hysterectomy with pelvic lymphadenectomy. After surgery, four cycles of combination chemotherapy with cisplatin and etoposide were administered, and the patient is disease-free as of 13 months after surgery. CONCLUSION: When a polypoid lesion is found, especially when it demonstrates rapid growth, it may be necessary to excise and histologically examine the polyp even during pregnancy. PMID- 11277663 TI - Primary medical management of recurrent aggressive angiomyxoma of the vulva with a gonadotropin-releasing hormone agonist. AB - BACKGROUND: An aggressive angiomyxoma of the pelvis is a locally infiltrative lesion treated with wide local excision. Recurrence is common. A potential medical treatment alternative is reported. CASE: A 34-year-old woman presented with her second recurrence of a vulvar angiomyxoma following two prior surgical excisions. Analysis of the recurrent tumor for estrogen and progesterone receptors was strongly positive. The patient was treated with 3 months of a gonadotropin-releasing hormone (GnRH) agonist. Comparison of pre- and posttreatment magnetic resonance imaging scans showed complete radiographic resolution of the tumor. Physical examination confirmed these findings. CONCLUSION: Medical management with a GnRH agonist may obviate the need for radical exenterative surgery for a recurrent aggressive angiomyxoma of the vulva. PMID- 11277664 TI - Response to "Inguinal node status by ultrasound in vulva cancer" (Abang Mohammed et al. Gynecol Oncol 2000;77:93-6). PMID- 11277666 TI - LLETZ. PMID- 11277668 TI - LLETZ--evidence of its efficacy against HPV infection. PMID- 11277669 TI - Response Mode Effects and Moral Values. AB - In five studies, we measured the extent to which subjects weight moral product attributes in different response modes. We found that nonprice judgments such as likelihood of purchase ratings were more reflective of expressed moral attitudes than were pricing responses, and that holistic price evaluations were especially unlikely to reflect moral considerations. Post-task ratings confirmed the preference results, as did an experiment controlling for the influence of task goals. Our results have implications for compatibility theories of preference elicitation, the predictability of respondent ratings of attribute unacceptability, and the measurement of utilities for morally charged attributes. Copyright 2001 Academic Press. PMID- 11277670 TI - Individualism-Collectivism as a Boundary Condition for Effectiveness of Minority Influence in Decision Making. AB - Results of this experiment demonstrate that individualists and collectivists react differently to minority influence. Based on the distinction between objectivity and preference norms in the minority influence literature, we hypothesize that individualism and collectivism influence (A) responses to minority influence (focusing on the target of influence) and (B) effectiveness of minority influence (focusing on the influence agent). Our results replicate past research and demonstrate improved decision quality for individuals exposed to a minority perspective. Moreover, minority influence targets with high horizontal individualism and low horizontal collectivism made higher quality decisions. Influence targets with high vertical collectivism demonstrated higher quality decisions when the influence agent held a high status position in the group. Results also demonstrate that influence agents with high vertical individualism experienced less role stress than those with low vertical individualism. Finally, influence agents with low role stress were more effective in influencing the decision making of others. We discuss our findings in terms of boundary conditions to the minority influence process. Copyright 2000 Academic Press. PMID- 11277671 TI - Beyond the Obvious: Chronic Vividness of Imagery and the Use of Information in Decision Making. AB - The authors investigate two competing hypotheses about how chronic vividness of imagery interacts with the vividness and salience of information in decision making. Results from four studies, covering a variety of decision domains, indicate that chronic imagery vividness rarely amplifies the effects of vivid and salient information. Imagery vividness may, in fact, attenuate the effects of vivid and salient information. This is because, relative to nonvivid imagers, vivid imagers rely less on information that appears obvious and rely more on information that seems less obvious. This tendency is so robust that vividness of imagery may amplify the effects of vivid information only when this information is the only information available in the decision field. The findings seem to reflect vivid imagers' tendency to totally immerse themselves in a decision problem and scrutinize the available information creatively. Copyright 2000 Academic Press. PMID- 11277672 TI - Who's Being Served? "Self-Serving" Attributions in Social Hierarchies. AB - Making external attributions for negative events, though often considered "self serving," also implies that the attributor is not in control of critical resources. We hypothesized that making external attributions for negative events will lead to impressions of powerlessness. Because individuals in high-status roles are expected to have power and control, external attributions may violate these role expectations; thus, we further hypothesized that status would moderate the relationship between attributions and interpersonal outcomes. Specifically, more negative impressions and affect will be directed toward high-status individuals who make external attributions than toward their lower status counterparts. Three studies were conducted, one using a role-play methodology, one using an experimentally created hierarchy, and one using vignettes. The results supported our hypotheses: external attributions can be highly disserving for people in high-status positions. Copyright 2000 Academic Press. PMID- 11277673 TI - Trust, Confidence, and Expertise in a Judge-Advisor System. AB - The relationship between trust, confidence, and expertise in Judge-Advisor Systems is examined in two experiments with Judge-Advisor pairs, one with strangers and another with participants in ongoing relationships. There was expertise asymmetry so that Judges had less expertise than their Advisors. The dyads could receive money for accurate Judge decisions. Either the Judge or Advisor had the power to allocate this money between dyad members, before task interaction in study one and after task completion in study two. Because Judges were more dependent on Advisors than vice versa, it was predicted that trust would be more important to Judges. Results were supportive. Judges had higher and more variable ratings of trust in their partner than did Advisors, suggesting that Judges were more motivated to evaluate trust. High confidence by Advisors had a positive impact on Judges' ratings of trust and tendency to follow their advice. Judges' trust in their Advisors was significantly related their taking the advice and being confident in their final decisions. Although participants in study two had higher levels of trust in their partners, they allocated less money to them. The implications for establishing trust are discussed. Copyright 2001 Academic Press. PMID- 11277674 TI - Normative and Descriptive Analyses of Simpson's Paradox in Decision Making. AB - Suppose that you are evaluating two delivery companies. Your investigation shows that Company B has a better on-time rate for small packages and also for large packages. Despite Company B's performance, however, Company A has a better overall on-time rate. This situation exemplifies Simpson's Paradox, in which the judged relationship between two variables (e.g., company and performance) differs depending on whether that relationship is viewed within subcategories of a third variable (e.g., package size) or in the aggregate. A normative analysis is presented arguing that the reasonableness of using the third variable depends upon the sample size as well as the separation between and variability within categories. To test subjects' abilities to behave appropriately in Simpson's Paradox situations, we examined responses to variations in these factors in five studies. Results showed that subjects had little or no sensitivity to differing stimulus set sizes. Also, subjects were sensitive to relationship strength and the variability within and between groups, and in nonnormative ways. Subjects' judgment behavior is related to a broader perspective concerning selection among multiple available levels of analysis based on a consideration of argument strength. Copyright 2000 Academic Press. PMID- 11277675 TI - Children and guns in a well child cohort. AB - BACKGROUND: The purpose of this study was to estimate the extent of and to identify predictors of preadolescent gun use in a well child cohort with matched parent and child data. METHODS: We analyzed self-report questionnaires from children and their parents using conditional logistic regression models. Questionnaires were given to 3,145 ten- to twelve-year-old children and 3,145 parents enrolled by their pediatricians in a prevention cohort study. RESULTS: Thirty-two percent of the children lived in households with guns. Children and parents generally agreed about the presence of guns in their homes; 17% had access to unlocked guns in their homes; 22% had fired guns. In this preadolescent cohort, firing guns was associated with being male, having guns in the home, having friends who use guns, and initiation of alcohol use. CONCLUSIONS: In this well child cohort, significant numbers of preadolescent, healthy boys in white, middle-class U.S. homes have access to guns, are using guns, and have friends who use guns. These children are also early alcohol adopters. Safety interventions with parents of preadolescents about the risks for accidental injury, death, and suicide due to child gun use may prove beneficial. PMID- 11277676 TI - Page for patients. Menopause. PMID- 11277677 TI - Factors that influence passive smoking in infancy: a study among mothers of newborn babies in The Netherlands. AB - BACKGROUND: The aim of this study was to assess the factors that influence smoking in the presence of the infant by mothers, partners, other family members, and friends. METHODS: An observational study using questionnaires was performed with smoking and nonsmoking parents of babies between 1 and 14 months old attending Dutch well-baby clinics between February and May 1996. The main measures were prevention of passive smoking in children by mothers and the relation with self-reported attitudes, social influence, and self-efficacy. RESULTS: A total of 1702 parents completed the questionnaire (63%). A total of 1551 questionnaires were completed by the mother. Sixty-five percent of the mothers prevented passive smoking by their child. This figure was 55% for smokers and 69% for nonsmokers. Attitude was the factor that most explained preventive behavior among both smokers and nonsmokers. Among the respondents, a lack of prevention of passive smoking was significantly related to (1) a negative attitude and 2) a negative social influence exerted by their partner, (3) lower self-efficacy in reducing passive smoking, and (4) increasing age of the child. (5) Finally, a lack of prevention is associated with the mother's self-efficacy in asking others not to smoke. This association strongly differs between smoking and nonsmoking mothers. CONCLUSION: The results suggest that health education efforts should focus on attitude and self-efficacy, assuming that these precede actual behavior, and in particular on the health consequences of the exposure of young children to tobacco smoke. The information should not be restricted to parents of newborn babies; it should also focus on parents with older children. Particular attention should be paid to smokers with a low educational level. The results also indicate that education should strengthen the ability of nonsmoking parents to deal with smokers and the ability of smoking parents to deal with their own smoking behavior. PMID- 11277678 TI - Smoking initiation and cessation by gender and educational level in Catalonia, Spain. AB - BACKGROUND: The role of gender and socioeconomic status in smoking has been characterized in the United States and Northern European countries. However, there is scarce information of the dynamic of the tobacco epidemic in Southern European countries. The aim of this study was to analyze smoking initiation and cessation according to level of education and gender in Catalonia, Spain. METHODS: Data from the Catalan Health Survey (1994), a cross-sectional study based in a representative sample of the noninstitutionalized population of Catalonia, was used. The relative risks and 95% confidence interval of smoking initiation were computed by means of Cox's regression. The odds ratios and 95% confidence intervals of quitting smoking were derived from logistic regression models. Direct responses from 4,370 men and 5,213 women ages 25 years or over were included for analysis. RESULTS: Ever smoking was inversely related to level of education in men. Males with the highest educational level tended to have a lower probability of being a smoker at a given age than those with less than primary school (relative risk = 0.6; 95% confidence interval: 0.5-0.7). This pattern appeared with small variation across age groups. In women, a reverse trend was present: the higher the level of education the higher the relative risk of starting smoking (relative risk = 4.6; 95% confidence interval: 3.1-6.7). Quitting smoking was more likely among men and women with higher education as compared to men and women with less than primary school (men: odds ratio = 1.5; 95% confidence interval 1.1-2.1; women: odds ratio = 4.6; 95% confidence interval: 2.1-10.4). CONCLUSIONS: The differential effect of education according to gender may reflect different phases of the smoking epidemic. In Catalonia, the transition of smoking from upper and lower socioeconomic groups occurred recently among men, and women have currently begun to experience this transition. PMID- 11277679 TI - Direct measurement of low-density-lipoprotein cholesterol is more effective than total cholesterol for the purpose of lipoprotein screening. AB - BACKGROUND: We have developed a new method for chemically measuring blood low density-lipoprotein (LDL) cholesterol. In the present study, we simulated guidelines of the National Cholesterol Education Program (NCEP) using our LDL cholesterol measurements. METHODS: Blood samples were collected from 1,069 individuals (519 males, 550 females) who were referred to our laboratory at Niigata University Hospital for lipoprotein analysis. LDL cholesterol levels were determined according to our assay protocol, which has been published previously. Subjects were categorized by NCEP guidelines and identified "false positives" and "false negatives" on the basis of LDL cholesterol levels measured by our method. RESULTS: The sensitivity of the NCEP guidelines is 87.5% and the specificity is 87.1%, provided we assume that every individual has fewer than two risk factors for coronary heart disease. If we assume that every individual has two or more risk factors, the sensitivity and specificity of the guidelines are 99 and 56.8%, respectively. CONCLUSION: This study presents an opportunity to reevaluate guidelines for routine lipoprotein screening. The chance that individuals with higher LDL cholesterol and lower high-density-lipoprotein cholesterol levels in serum would be missed at initial classification should be zero. The chance that individuals with desirable lipid levels would undergo further lipoprotein analysis should be decreased. Since the new method can be implemented cost effectively in routine lipoprotein screening, direct measurement of LDL cholesterol could replace total cholesterol. PMID- 11277680 TI - Seattle 5 a Day worksite program to increase fruit and vegetable consumption. AB - BACKGROUND: 5 a Day for Better Health is a simple message encouraging people to eat more fruits and vegetables. The Seattle 5 a Day worksite investigators designed and evaluated an intervention, organized on stages of behavioral change, to increase worksitewide fruit and vegetable consumption. METHODS: We recruited 28 worksites with cafeterias and randomized 14 to intervention and 14 to control. The intervention addressed both changes in the work environment and individual level behavior change. In each worksite, an employee advisory board, with study interventionist assistance, implemented the program. By surveying cross-sectional samples of 125 employees per worksite, we compared worksite mean fruit and vegetable consumption at 2-year follow-up with that at baseline. Unobtrusive site level indicators including plate observation and cafeteria checklist were also used. RESULTS: The difference at 2 years was 0.5 for the intervention worksites and 0.2 for the control worksites, with an intervention effect of 0.3 daily serving (P < 0.05). Other measures of fruit and vegetable consumption, including unobtrusive indicators, supported the effectiveness of the intervention. CONCLUSIONS: This simple 5 a Day intervention is feasible and acceptable for use in worksites with cafeterias. There was a significant differential increase in fruit and vegetable consumption in the intervention worksites. This kind of worksite intervention can achieve important health benefits on a population basis, because of its potential to reach large numbers of people. PMID- 11277681 TI - Effectiveness of altered incentives in a food safety inspection program. AB - BACKGROUND: The Los Angeles County Department of Health Services revamped its retail food establishment inspection program in 1998 to include, among other novel provisions, the prominent posting of grades. The purpose of the present study is to examine results from the second year of the revamped program to determine the longer-term impact of the program. METHODS: An analysis of program results for Year 1 led to several program refinements: stratification of risk categories, certification of food handlers, and an external fraud hotline. Outcome measures used included routine inspection scores; closure rates; scores from owner-initiated inspections and departmentally initiated inspections; inspection frequencies; compliance with food handler certification requirements; and frequency of use of the external fraud hotline. RESULTS: Inspection scores continued to increase in 1999, while the rate of closures decreased. The owner initiated inspections also resulted in improved scores, generally maintained on subsequent inspections. Higher risk establishments were inspected more frequently than were lower risk establishments. Over 30,000 food handlers were certified. The external fraud hotline received 16 fraud complaints. CONCLUSIONS: Results indicate that retail food establishments are taking stronger action to assure compliance with food safety practices. PMID- 11277682 TI - Where's the fat? Trends in U.S. diets 1965-1996. AB - BACKGROUND: Controlling fat intake has been an ongoing health concern since the late 1950s. This study examines 30-year trends in food sources of fat intake. It focuses on both total fat and specific fatty acid classes to ascertain if there are trend differences by age, sex, or race/ethnicity. METHODS: Nationally representative cross-sectional U.S. Department of Agriculture surveys from 1965, 1977-1978, 1989-1991, and 1994-1996 form the basis of this analysis, which compares 45,357 adults aged 18 years and older. Food files linked over time are used to create comparable food groups and nutrient values. RESULTS: The proportion of fat in the diet from grain-based mixed dishes, higher-fat snack foods, and higher fat potatoes has increased to partially offset reductions in fat from dairy, red meat, and added fat categories. Food sources of fat differ by race/ethnicity and age. The percentage of fat from fast foods and ethnic foods increased over time from 1 to 11% of total fat. The ratio of visible to invisible fat declined considerably. CONCLUSION: While animal product-based sources of fat continue to require emphasis, the shift toward fast foods, fried foods, and grain based mixed dish and edible oil sources requires more focus. PMID- 11277683 TI - Physician perceptions about administration of immunizations outside of physician offices. AB - BACKGROUND: Expanding nonphysician participation in the administration of immunizations has been suggested as a means of increasing immunization rates. However, there is little information about physician interest in collaborating with nonphysicians to provide out-of-office immunizations. METHODS: All active members of the Iowa Academy of Family Physicians were surveyed by mail. Physicians reported on their collaboration histories, their willingness to collaborate in the future, their concerns with collaboration, and whether they approved of their patients' using nonphysicians for immunizations. RESULTS: Of 898 eligible physicians, 476 (53%) returned questionnaires that were analyzed. Seventy-five percent (n = 357) of the physicians reported that they had voluntarily collaborated with a person outside their office to provide immunizations. Ninety-five percent (n = 452) of physicians indicated a willingness to collaborate in some form in the future. However, physicians had concerns about (a) being able to be kept informed about immunizations their patients receive outside of their offices, (b) adequate training of the nonphysician to administer immunizations and respond to complications of immunization, and (c) loss of preventive health opportunities if patients ceased coming to physicians for routine immunizations. CONCLUSION: The majority of family physicians have collaborated to deliver immunizations and indicate support for nonphysician participation. Almost all physicians would consider future collaborative arrangements although they have concerns about record keeping and the safety of out-of-office immunization programs. PMID- 11277684 TI - Evidence that smokeless tobacco use is a gateway for smoking initiation in young adult males. AB - BACKGROUND: This study was designed to test the hypothesis that smokeless tobacco (SLT) serves as a gateway drug for smoking among young adult males. Methods. A cohort (n = 7,865) of U.S. Air Force recruits who claimed to have never smoked cigarettes was followed prospectively for 1 year. The participants were male, 32.9% were ethnic minorities, and their average age was 19.84 years (SD = 2.29). Among recruits entering basic military training, 403 (5.1%) reported current SLT use and 198 (2.5%) reported a past history of SLT use. RESULTS: At the 1-year follow-up current SLT users were 233% more likely to have initiated smoking than nonusers (odds ratio = 2.33, 95% CI = 1.84-2.94). Similarly, recruits who reported past SLT use were 227% more likely to begin smoking than participants who had never used SLT (odds ratio = 2.27, 95% CI = 1.64-3.15). SLT use remained a potent predictor of smoking initiation in a multivariate logistic model that included demographic factors and other risk factors for initiation. CONCLUSIONS: SLT use appears to be an important predictor of smoking initiation among young adult males. This study suggests that smoking prevention and cessation programs should also include strategies related to SLT use. PMID- 11277685 TI - Population-based prevention of eating disorders: an application of the Rose prevention model. AB - BACKGROUND: Several decades of concerted research on eating disorders have generated a broad range of proposed causal influences, but much of this etiologic research does not elucidate practical avenues for preventive interventions. Translating etiologic theory into community health interventions depends on the identification of key leverage points, factors that are amenable to public health intervention and provide an opportunity to maximize impact on the outcome of interest. Population-based preventive strategies, elaborated by epidemiologist Geoffrey Rose, can maximize the impact of public health interventions. In the case of eating disorders, Rose's model is instructive: Dieting stands out as risk behavior that may both fit Rose's model well and be a key leverage point for preventive intervention. METHODS: Grounded in Rose's work, this article lodges a theoretical argument for the population-based prevention of eating disorders. In the introductory section, existing research on the epidemiology of dieting is reviewed, showing that it is extremely common among adolescent girls and women and that the behavior has been implicated as a causal factor for disordered eating. Next, new evidence is offered to build a case for how a population-wide reduction in dieting may be an effective strategy for prevention of eating pathology. Finally Rose's prevention framework is used to introduce a unique and provocative perspective on the prevention of eating disorders. RESULTS: Dieting is a normative behavior in our culture with psychological and physiological effects in the causal chain leading to eating pathology. This behavior may represent an ideal target for population-based prevention. CONCLUSIONS: Theoretical and empirical evidence suggests that a population-wide reduction in dieting may be a justifiable and effective strategy for prevention of eating pathology. PMID- 11277686 TI - A computer simulation model of mass media interventions directed at tobacco use. AB - OBJECTIVES: The goal of this study was to develop a simulation model to examine the effects of tobacco control mass media interventions on smoking rates and smoking-attributable deaths. METHODS: The model projects the number of smokers and smoking-related deaths. Based on empirical and theoretical research, the effects of media interventions, varying in magnitude and duration, directed at all smokers and directed specifically at youth under age 18 are modeled. RESULTS: The model predicts that sustained media interventions of sufficient magnitude and duration directed at all smokers have the potential to substantially reduce the number of smokers and premature deaths, with the effects growing over time. For the same expenditures, youth interventions would appear to have smaller and more delayed effects. CONCLUSIONS: Media interventions, particularly those targeted at the general population and of sufficient scale and duration, have the ability to substantially reduce smoking rates and save lives, but their effects are likely to depend on how they are implemented. PMID- 11277687 TI - Relationships of physical activity with dietary behaviors among adults. AB - BACKGROUND: Physical activity and diet are important influences on health, but few data are available about the relationship between these two factors. The purpose of this study was to examine relationships between physical activity and dietary quality and to identify determinants of the combination of sedentary behavior and suboptimal diet. METHODS: The design of this study was cross sectional. The setting was a large managed-care organization and the participants were 1,322 racially diverse men and women ages 25-91 years. We categorized subjects' physical activity into vigorous, moderate, and sedentary based on answers to two validated interviewer-administered questions about intensity and duration of specified activities. Dietary assessment was by means of a validated short food frequency questionnaire. We defined suboptimal diet as consuming unhealthful quantities of at least two of the following five food groups: fruits, vegetables, whole grain foods, whole-fat dairy foods, and red and processed meats. RESULTS: Seven hundred fifty-four (57%) subjects were sedentary and 617 (47%) consumed a suboptimal diet. Using multiple linear regression, we found that sedentary individuals consumed smaller amounts of foods and nutrients considered to be healthful, such as fruits and vegetables, fiber, calcium, folate, and vitamins A, C, and E, than more active participants. For nutrients considered to be harmful, such as saturated fat, trans fat, and dietary cholesterol, the association with physical activity was inverse. In multiple logistic regression analyses, the strongest sociodemographic correlates of the joint presence of inactivity and poor diet were less education [odds ratio for 1-year decrease 1.14 (95% confidence interval 1.06, 1.22)], nonwhite race [1.48 (1.05, 2.07)], and nonmarried status [1.49 (1.06, 2.10)]. CONCLUSIONS: Physical activity and diet quality are correlated behaviors. Suboptimal diet and sedentary behavior tend to cluster in individuals who are less educated, not married, and of nonwhite race. Programs that target diet and activity together, informed by their joint determinants, may attain enhanced outcomes. PMID- 11277688 TI - Challenges for rotavirus vaccines. PMID- 11277689 TI - H9N2 influenza A viruses from poultry in Asia have human virus-like receptor specificity. AB - H9N2 influenza A viruses are currently widespread in chickens, quail, and other poultry in Asia and have caused a few cases of influenza in humans. In this study, we found that H9N2 viruses from Hong Kong live bird markets have receptor specificity similar to that of human H3N2 viruses. In addition, the neuraminidase of poultry H9N2 viruses has mutations in its hemadsorbing site, a characteristic resembling that of human H2N2 and H3N2 viruses but differing from that of other avian viruses. Peculiar features of surface glycoproteins of H9N2 viruses from Hong Kong suggest an enhanced propensity for introduction into humans and emphasize the importance of poultry in the zoonotic transmission of influenza viruses. PMID- 11277690 TI - Membrane topology of coronavirus E protein. AB - Coronavirus small envelope protein E has two known biological functions: it plays a pivotal role in virus envelope formation, and the murine coronavirus E protein induces apoptosis in E protein-expressing cultured cells. The E protein is an integral membrane protein. Its C-terminal region extends cytoplasmically in the infected cell and in the virion toward the interior. The N-terminal two-thirds of the E protein is hydrophobic and lies buried within the membrane, but its orientation in the lipid membrane is not known. Immunofluorescent analyses of cells expressing biologically active murine coronavirus E protein with a hydrophilic short epitope tag at the N-terminus showed that the epitope tag was exposed cytoplasmically. Immunoprecipitation analyses of the purified microsomal membrane vesicles that contain the same tagged E protein revealed the N-terminal epitope tag outside the microsomal membrane vesicles. These analyses demonstrated that the epitope tag at the N-terminus of the E protein was exposed cytoplasmically. Our data were consistent with an E protein topology model, in which the N-terminal two-thirds of the transmembrane domain spans the lipid bilayer twice, exposing the C-terminal region to the cytoplasm or virion interior. PMID- 11277691 TI - The genome sequence of Yaba-like disease virus, a yatapoxvirus. AB - The genome sequence of Yaba-like disease virus (YLDV), an unclassified member of the yatapoxvirus genus, has been determined. Excluding the terminal hairpin loops, the YLDV genome is 144,575 bp in length and contains inverted terminal repeats (ITRs) of 1883 bp. Within 20 nucleotides of the termini, there is a sequence that is conserved in other poxviruses and is required for the resolution of concatemeric replicative DNA intermediates. The nucleotide composition of the genome is 73% A+T, but the ITRs are only 63% A+T. The genome contains 151 tightly packed open reading frames (ORFs) that either are > or =180 nucleotides in length or are conserved in other poxviruses. ORFs within 23 kb of each end are transcribed toward the termini, whereas ORFs within the central region of the genome are encoded on either DNA strand. In the central region ORFs have a conserved position, orientation, and sequence compared with vaccinia virus ORFs and encode many enzymes, transcription factors, or structural proteins. In contrast, ORFs near the termini are more divergent and in seven cases are without counterparts in other poxviruses. The YLDV genome encodes several predicted immunomodulators; examples include two proteins with similarity to CC chemokine receptors and predicted secreted proteins with similarity to MHC class I antigen, OX-2, interleukin-10/mda-7, poxvirus growth factor, serpins, and a type I interferon-binding protein. Phylogenic analyses indicated that YLDV is very closely related to yaba monkey tumor virus, but outside the yatapoxvirus genus YLDV is more closely related to swinepox virus and leporipoxviruses than to other chordopoxvirus genera. PMID- 11277692 TI - Rinderpest virus C and V proteins interact with the major (L) component of the viral polymerase. AB - Rinderpest virus, like other Morbilliviruses, expresses three proteins from the single P gene. In addition to the P protein, which interacts both with the viral polymerase (L) and the nucleocapsid (N) protein, the virus expresses a C and a V protein from the same gene. The functions of these two proteins in the viral life cycle are not clear. Although both C and V proteins are dispensable, in that viable viruses can be made that express neither, each seems to play a role in optimum viral replication. We have used the yeast-two hybrid system, binding to coexpressed fusions of C and V to glutathione-S-transferase, and studies of the native size of these proteins to investigate interactions of the rinderpest virus C and V proteins with other virus-encoded proteins. The V protein was found to interact with both the N and L proteins, while the C protein was found to bind to the L protein, and to self-associate in high-molecular-weight aggregates. PMID- 11277693 TI - A critical role for inducible nitric oxide synthase in host survival following coxsackievirus B4 infection. AB - Coxsackieviral infections have been linked etiologically to multiple diseases. The serotype CB4 is associated with acute pancreatitis and autoimmune type 1 diabetes. To delineate the mechanisms of host survival after an acute infection with CB4 (strain E2), we have investigated the role of nitric oxide (NO), generated by the inducible form of nitric oxide synthase (NOS2), in viral clearance and pancreatic beta-cell maintenance. Mice deficient in NOS2 (NOS2-/- mice) and their wild-type (wt) counterparts were injected with CB4, after which both groups developed severe pancreatitis, hepatitis, and hypoglycemia within 3 days. Within 4 to 7 days postinfection (p.i.), most of the NOS2-/- mice died and at a strikingly higher mortality rate than wt mice. Histological examination of pancreata from both infected NOS2-/- and infected wt mice revealed early and complete destruction of the pancreatic acinar tissue, but intact, insulin-stained islets. When examined up to 8 weeks p.i., neither surviving NOS2-/-mice nor surviving wt mice developed hyperglycemia. However, the clearance of infectious CB4 was different between the mice. The spleens of NOS2-/- survivors were cleared of infectious virus with kinetics similar to that of wt mice, but the livers, pancreata, kidneys, and hearts of the NOS2-/- groups cleared virus more slowly than those of the wt group. This delayed clearance was particularly prominent in the livers of infected NOS2-/- mice, which also showed prolonged histopathological features of viral hepatitis. Taken together, this outcome suggests that NOS2 (and NO) is not required for the prevention of pancreatic beta cell depletion after CB4 infection. Instead the critical actions of NOS2 apparently occur early in the host immune response, allowing mice to survive and clear virus. Moreover, the data support the existence of an organ-specific dependency on NO for a rapid clearance of CB4. PMID- 11277694 TI - Membrane-associated respiratory syncytial virus F protein expressed from a human rhinovirus type 14 vector is immunogenic. AB - Human rhinovirus (HRV) replicons have the potential to serve as respiratory vaccine vectors for mucosal immunization in humans. However, since many vaccine immunogens of interest are glycosylated, an important concern is whether HRV replicons are capable of expressing glycosylated proteins. The human respiratory syncytial virus (RSV) fusion (F) protein was chosen as a model glycoprotein and the HRV replicon DeltaP1FVP3 was generated by inserting the F protein-coding sequence in frame and in lieu of the 5' proximal 1489 nucleotides of the capsid coding segment in the HRV-14 genome. When transfected into H1-HeLa cells, DeltaP1FVP3 replicated and led to the expression of the F protein. Inhibition with guanidine demonstrated that F-protein expression was dependent on DeltaP1FVP3 replication and did not result from translation of input RNA. Although most of the F protein remained as an immature, glycosylated precursor (F0), a readily detectable fraction of the protein was processed into the mature glycosylated subunit F1, an event known to occur within the Golgi apparatus. Packaged DeltaP1FVP3 replicons were generated in transfected HeLa cells by coexpression of homologous HRV capsid proteins using the vaccinia virus/T7 RNA polymerase hybrid system. Packaged replicon RNAs were capable of infecting fresh cells, leading to accumulation of the F protein as in RNA-transfected cells. Mice immunized with HeLa cell lysates containing F protein expressed from DeltaP1FVP3 produced neutralizing antibodies against RSV. These results indicate that an HRV 14 replicon can express a foreign glycosylated protein, providing further support for the potential of HRV replicons as a vaccine delivery system. PMID- 11277695 TI - Modified HPV16 E7 Genes as DNA Vaccine against E7-Containing Oncogenic Cells. AB - Therapeutic vaccines against tumors associated with human papillomaviruses (HPV) should elicit cellular immune responses against early HPV antigens, primarily the oncoproteins E7 and E6. Because of safety concerns, the direct use of an unmodified oncogene is impossible in human DNA vaccination. Therefore, we introduced three point mutations into the pRb-binding site of HPV16 E7 oncogene to eliminate its transformation potential. The resultant gene was denoted E7GGG. The rates of expression and the cellular localization of E7 and E7GGG proteins were comparable. In immunization-challenge experiments, the efficacy of plasmids containing the E7, E7GGG, or fusion genes of HPV16 E7, viz. L1DeltaCE7(1-60) (M. Muller et al., 1997, Virology 234, 93-111), and Sig/E7/LAMP-1 (T. C. Wu et al., 1995, Proc. Natl. Acad. Sci. USA 92, 11671-11675), was compared. While tumors developed in all animals immunized with the wild-type E7 gene, a significant proportion of mice remained tumor-free after vaccination with the E7GGG gene. The fusion gene L1DeltaCE7(1-60) induced negligible protection, but Sig/E7/LAMP-1 conferred the highest protection. Intradermal immunization by gene gun proved superior to i.m. inoculation. In "therapeutic" experiments, a 1-day delay between inoculation of oncogenic cells and the start of DNA immunization resulted in partial therapeutic effect, but a 3-day delay produced a substantially lower immunization effect. A combination of Sig/E7/LAMP-1 and E7GGG genes did not enhance the immune response. These results demonstrate a significant enhancement of HPV16 E7 immunogenicity after mutagenesis of the pRb-binding site, but the mutated E7 gene did not excel the Sig/E7/LAMP-1 fusion gene. PMID- 11277696 TI - Coreceptor choice and T cell depletion by R5, X4, and R5X4 HIV-1 variants in CCR5 deficient (CCR5delta32) and normal human lymphoid tissue. AB - Coreceptor utilization by HIV-1 is an important determinant of pathogenesis. However, coreceptor selectivity is defined in vitro, while in vivo critical pathogenic events occur in lymphoid tissues. Using pharmacological inhibitors, we recently provided evidence that coreceptor selectivity by the R5X4 dual-tropic isolate 89.6 was more restricted in ex vivo infected lymphoid tissue than in vitro [S. Glushakova, Y. Yi, J. C. Grivel, A. Singh, D. Schols, E. De Clercq, R. G. Collman, and L. Margolis (1999). J. Clin. Invest. 104, R7-R11]. Here we extend those observations using CCR5-deficient (CCR5Delta32) lymphoid tissue as well as additional primary isolates. We definitively show that neither CCR5 nor secondary coreceptors used in vitro mediate 89.6 infection in lymphoid tissue. We also demonstrate that restricted coreceptor use in lymphoid tissue ex vivo compared with in vitro utilization occurs with other dual-tropic primary isolates and is not unique to 89.6. For all strains tested that are dual tropic in vitro, severe CD4 T cell depletion in lymphoid tissue correlated with preferential CXCR4 use in this ex vivo system. PMID- 11277697 TI - Inhibition of HIV infection by the cytokine midkine. AB - The growth factor midkine (MK) has been reported to bind heparan sulfate and nucleolin, two components of the cell surface implicated in the attachment of HIV 1 particles. Here we show that synthetic and recombinant preparations of MK inhibit in a dose-dependent manner infection of cells by T-lymphocyte- and macrophage-tropic HIV-1 isolates. The binding of labeled MK to cells is prevented by excess unlabeled MK or by the anti-HIV pseudopeptide HB-19 that blocks HIV entry by forming a stable complex with the cell-surface-expressed nucleolin. MK mRNA is systematically expressed in adult peripheral blood lymphocytes from healthy donors, while its expression becomes markedly but transiently increased upon in vitro treatment of lymphocytes with IL-2 or IFN-gamma and activation of T lymphocytes by PHA or antibodies specific to CD3/CD28. In MK-producing lymphocytes, MK is detectable at the cell surface where it colocalizes with the surface-expressed nucleolin. Finally, by using MK-producing CD4(+) and CD4(-) cell clones we show that HIV infection in cell cultures could be inhibited in both an autocrine and a paracrine manner. The potent and distinct anti-HIV action of MK along with its enhanced expression in lymphocytes by various physiological stimuli suggests that MK is a cytokine that could be implicated in HIV-induced pathogenesis. PMID- 11277698 TI - Antigenic sites of poliovirus type 3 eliciting IgA monoclonal antibodies in orally immunized mice. AB - A panel of neutralizing IgA monoclonal antibodies was produced from mice orally inoculated with poliovirus type 3 Sabin and cholera toxin as adjuvant. Low levels of neutralizing antibodies were elicited in mice after several boosts, but only in the presence of cholera toxin. Characterization of IgA MAbs by neutralization escape virus mutants showed that all but one neutralizing MAbs against type 3 poliovirus were directed to antigenic site N-AgIII, which was previously found by us to be the major target of mucosal immune response to Sabin 1 in the mouse. Our data indicate that residue 236 of VP3, not previously reported, is also involved in forming site N-AgIII in addition to formerly described VP3 (aa 58-59) and VP1 (aa 286-290) residues. Unlike poliovirus type 1 IgA MAbs, all IgA MAbs herein described neutralized the wild-type parental poliovirus. PMID- 11277699 TI - A new strategy in design of +RNA virus infectious clones enabling their stable propagation in E. coli. AB - Infectious clone methodology is a valuable tool of modern experimental virology. However, its use is often constrained by the instability of infectious clone constructs during propagation in E. coli. To circumvent this problem, we have devised a strategy that could be suitable for design of +RNA virus molecular clones in general. An infectious clone is assembled as "infectious DNA," and expression of problem regions present in the viral cDNA is prevented during propagation in E. coli by insertion of short introns. To demonstrate the feasibility of this approach, a highly unstable Japanese encephalitis flavivirus infectious clone has been successfully converted into a remarkably stable infectious DNA construct with the specific infectivity of 10(6) pfu/microg in cell culture. The proposed strategy may be useful in the design of self amplifying gene therapy vectors and development of new immunization methodologies, and could facilitate creation of molecular repositories of existing viral vaccines. PMID- 11277700 TI - Chemical modification patterns of active and inactive as well as procapsid-bound and unbound DNA-packaging RNAof bacterial virus Phi29. AB - During replication, the lengthy genome of dsDNA viruses is translocated with remarkable velocity into the limited space within the preformed procapsid. We previously found that a viral-encoded RNA (pRNA) played a key role in bacterial virus phi29 DNA translocation. Design of mutant pRNA sets containing two and three inactive mutant pRNAs, respectively, led to the conclusion that the stoichiometry of pRNA in DNA packaging is the common multiple of 2 and 3. Together with studies using binomial distribution of mutant and wild-type pRNA, it has been confirmed that six pRNAs of phi29 form a hexagonal complex to drive the DNA translocating machine. These findings have brought about commonality between viral DNA packaging and other universal DNA/RNA-riding processes including DNA replication and RNA transcription. Chemical modification was used to compare the structures of active and inactive as well as free and procapsid bound pRNA. Our results explain why certain pRNA mutants are inactive in DNA packaging while remaining competent in procapsid binding, since the mutations were located in a domain involved in DNA translocation that is dispensable for procapsid binding. A mutant pRNA that had reduced procapsid binding was revealed to have a structural alteration within the procapsid-binding region that may account for the binding deficiency. Chemical probing of procapsid-bound pRNA revealed a large area of protection, while a 3-base bulge, C(18)C(19)A(20), was accessible to chemicals. A pRNA with a deletion of this 3-base bulge was fully competent to form dimers, bind procapsids, and inhibit phi29 virion assembly in vitro; however, its activity in DNA packaging and virion assembly was completely lost. The results suggest that this bulge is not involved in procapsid binding but may interact with other DNA-packaging components. A computer model showing the location of the CCA bulge was presented. PMID- 11277701 TI - An infectious clone of the West Nile flavivirus. AB - West Nile (WN) virus is the most widespread among flaviviruses, but until recently it was not known on the American continent. We describe here design of a subgenomic replicon, as well as a full-length infectious clone of the lineage II WN strain, which appeared surprisingly stable compared to other flavivirus infectious clones. This infectious clone was used to investigate effects of 5'- and 3'-nonrelated sequences on virus replication and infectivity of synthetic RNA. While a long nonrelated sequence at the 3'-end delayed but did not prevent establishment of the productive infectious cycle, a much shorter extra sequence at the 5'-end completely abrogated virus replication. Replacement of the conserved 5'-adenosine residue substantially delayed, but did not prevent, establishment of virus infection. In all cases, the recovered virus had restored its authentic 5'- and 3'-end genome sequences. However, the presence of extensive nonrelated sequences at both 5'- and 3'-ends could not be repaired. PMID- 11277702 TI - Potent broad cross-neutralizing sera inhibit attachment of primary HIV-1 isolates (groups M and O) to peripheral blood mononuclear cells. AB - Gp160-directed antibody-mediated neutralization is thought to function by at least two different mechanisms that impair virus entry into the host cell: inhibition of virus attachment and inhibition of virus-cell membrane fusion. Previously, the neutralization spectra of sera derived from human immunodeficiency virus type 1 (HIV-1) infected patients were determined using 17 primary isolates belonging to HIV-1 group M (env clades A-H) and group O. The sera could be categorized as potent broad cross-neutralizing, limited cross neutralizing, and nonneutralizing sera. The aim of this study was to examine whether the neutralizing capacity of polyclonal human sera correlates with their capacity to inhibit the attachment of infectious virions to the surface of peripheral blood mononuclear cells. A 100% correlation was found between the broad cross-neutralizing capacity and the ability to inhibit binding of primary isolates belonging to different genetic clades and groups to peripheral blood mononuclear cells. These results may indicate that broad cross-neutralizing antibodies are directed against those conserved regions on gp120 that interact with the cell receptor(s) and that those antibodies can therefore interfere with the binding of virus to the host cell. PMID- 11277703 TI - Porcine HveC, a member of the highly conserved HveC/nectin 1 family, is a functional alphaherpesvirus receptor. AB - Human herpesvirus entry mediator C (HveC) is an alphaherpesvirus receptor which binds to virion glycoprotein D (gD). We identified porcine HveC and studied its interaction with pseudorabies virus (PrV) and herpes simplex virus type 1 (HSV-1) gD. Porcine and human HveC have 96% amino acid identity and HveC from African green monkey, mouse, hamster, and cow are similarly conserved. Porcine HveC mediates entry of HSV-1, HSV-2, PrV, and bovine herpesvirus type 1. Truncated soluble forms of HSV-1 and PrV gD bind competitively to porcine HveC. Biosensor analysis shows that PrV gD binds with a 10-fold higher affinity than HSV-1 gD. Monoclonal antibodies against human HveC recognize the porcine homologue and can block gD binding and entry of HSV-1 and PrV. Porcine HveC is functionally indistinguishable from human HveC. Our results are consistent with the suggestion that HveC is a pan-alphaherpesvirus receptor that interacts with a conserved structural domain of gD. PMID- 11277704 TI - Selected concepts and investigations in compound classification, molecular descriptor analysis, and virtual screening. PMID- 11277705 TI - Internet software for the calculation of the lipophilicity and aqueous solubility of chemical compounds. AB - In this paper we describe an Internet Java-based technology that allows scientists to make their analytical software available worldwide. The implementation of this technology is exemplified by programs for the calculation of the lipophilicity and water solubility of chemical compounds available at http://www.lnh.unil.ch/~itetko/logp. Both these molecular properties are key parameters in quantitative structure-activity relationship studies and are used to provide invaluable information for the overall understanding of the uptake distribution, biotransformation, and elimination of a wide variety of chemicals. The compounds can be analyzed in batch or single-compound mode. The single compound analysis offers the possibility to compare our results with several popular lipophilicity calculation methods, including CLOGP, KOWWIN, and XLOGP. The chemical compounds are analyzed according to SMILES line notation that can be prepared with the JME molecular editor of Peter Ertl. Conversion to SMILES from 56 formats is also available using the molecular structure information interchange hub developed by Pat Walters and Matt Stahl. PMID- 11277706 TI - JChemTidy: a tool for converting chemical Web document collections to an XHTML representation. AB - A robot-based procedure is described for traversing a collection of hyperlinked documents written in HTML and converting these to the XML-compliant and well formed XHTML representation. Transcluded chemical content invoked using or HTML calls are converted to the XHTML recommended form. Additional attributes such as title or derived chemical attributes such as a SMILES descriptor are added to improve the indexing of the resulting document collection. Conformance tests for the popular Web browsers are reported. PMID- 11277707 TI - Chemical registries--in the fourth decade of service. AB - Methods for precise yet usable descriptions of virtually any topic are essential, especially for identification of chemical compounds and materials. Systematic nomenclature and precise chemical structures, if known, are the ultimate in description of chemical compounds. However, there has always been a pervasive need for brief, yet precise methods to register chemical compounds for use in both indexing and retrieval of additional information. The most predominant chemical registration system is the Chemical Abstracts Service (CAS) Registry System, begun in 1965. The history and use of this system will be described, and its importance to a number of disciplines, not just chemistry, will be described. PMID- 11277708 TI - The journals crisis: redirecting the blame. AB - Publishers and librarians are the usual culprits blamed for the "journals crisis", typically defined as an ever-tightening spiral of increasing serial prices and decreasing number of subscribers. An alternative viewpoint is expressed in this article-the author community bears the brunt of the blame for this situation. The glut of published articles, the surplus of journals, and the growing publication rate per academic chemist drive the crisis. Therefore, any global publication paradigm change must originate from within the author community. PMID- 11277709 TI - A new and simple approach to chemical complexity. Application to the synthesis of natural products. AB - A new approach to describe the complexity of chemical structures is proposed. This index is simple and can be easily programmed. It derives from Whitlock's index and, despite its empirical character, provides results paralleling those obtained with the mathematical Bertz index. It has been applied to follow the strategic progress of some natural product syntheses. PMID- 11277710 TI - Traditional Chinese medicine database and application on the Web. AB - To study traditional Chinese medicines and exchange related information through the worldwide Web, we developed a traditional Chinese medicine database and a program for searching and displaying data in the database on the Web. In this paper, the traditional Chinese medicine database is briefly introduced; the methods used in developing the program, including ISAPI (Microsoft Internet Server Application Programming Interface), VRML (Virtual Reality Model Language), and JavaScript are described; and three application examples are also given. PMID- 11277711 TI - A simple method for aligning many protein sequences. AB - A simple extension of the Needleman and Wunsch algorithm for aligning pairs of protein sequences allows it to be used for the efficient generation of very large multiple-sequence alignments whose members are similar. This technique could have applications in a broad range of high-volume genomics projects. PMID- 11277712 TI - Database mining using soft computing techniques. An integrated neural network fuzzy logic-genetic algorithm approach. AB - Two different soft computing (SC) techniques (a competitive learning neural network and an integrated neural network-fuzzy logic-genetic algorithm approach) are employed in the analysis of a database subset obtained from the Cambridge Structural Database. The chemical problem chosen for study is relevant to the relationship between various metric parameters in transition metal imido (LnMdNZ, Z = carbon-based substituent) complexes and the chemical consequences of such relationships. The SC techniques confirmed and quantified the suspected relationship between the metal-nitrogen bond length and the metal-nitrogen substituent bond angle for transition metal imidos: increased metal-nitrogen carbon angles correlate with shortened metal-nitrogen distances. The mining effort also yielded an unexpected correlation between the NC distance and the MNC angle-shorter NC correlate with larger MNC. A fuzzy inference system is used to construct an MNred-NC-MNC hypersurface. This hypersurface suggests a complicated interdependence among NC, MNred, and the angle subtended by these two bonds. Also, major portions of the hypersurface are very flat, in regions where MNC is approaching linearity. The relationships are also seen to be influenced by whether the imido substituent is an alkyl or aryl group. Computationally, the present results are of particular interest in two respects. First, SC classification was able to isolate an "outlier" cluster. Identification of outliers is important as they may correspond to unreported experimental errors in the database or novel chemical entities, both of which warrant further investigation. Second, the SC database mining not only confirmed and quantified a suspected relationship (MNred versus MNC) within the data but also yielded a trend that was not suspected (NC versus MNC). PMID- 11277713 TI - A probabilistic derivation of the partial least-squares algorithm. AB - Traditionally the partial least-squares (PLS) algorithm, commonly used in chemistry for ill-conditioned multivariate linear regression, has been derived (motivated) and presented in terms of data matrices. In this work the PLS algorithm is derived probabilistically in terms of stochastic variables where sample estimates calculated using data matrices are employed at the end. The derivation, which offers a probabilistic motivation to each step of the PLS algorithm, is performed for the general multiresponse case and without reference to any latent variable model of the response variable and also without any so called "inner relation". On the basis of the derivation, some theoretical issues of the PLS algorithm are briefly considered: the complexity of the original motivation of PLS regression which involves an "inner relation"; the original motivation behind the prediction stage of the PLS algorithm; the relationship between uncorrelated and orthogonal latent variables; the limited possibilities to make natural interpretations of the latent variables extracted. PMID- 11277714 TI - QSRR correlation of free-radical polymerization chain-transfer constants for styrene. AB - Quantitative structure-reactivity relationships (QSRR) are deduced for kinetic chain-transfer constants for 90 agents on styrene polymerization at 60 degrees C. Three- and five-parameter correlations were obtained with R2 of 0.725 and 0.818, respectively. The descriptors involved in the correlations are consistent with the proposed mechanism of the chain-transfer reactions. PMID- 11277715 TI - Allowed boundary sequences for fused polycyclic patches and related algorithmic problems. AB - We consider sequences that encode boundary circuits of fused polycycles made up of polygonal faces with p sides, p < or = 6. We give a constructive algorithm for recognizing such sequences when p = 5 or 6. A simpler algorithm is given for planar hexagonal sequences. Hexagonal and pentagonal sequences of length at most 8 are tabulated, the former corresponding to planar benzenoid hydrocarbons CxHy with y up to 14. PMID- 11277716 TI - Inversion of simulated positron annihilation lifetime spectrum using a neural network. AB - Inversion of positron annihilation lifetime spectroscopy, based on a neural network Hopfield model, is presented in this paper. From a previous reported density function for lysozyme in water a simulated spectrum, without the superposition of statistical fluctuation and spectrometer resolution effects, was generated. These results were taken as the exact results from which the neural network was trained. The precision of the inverted density function was analyzed taking into account the number of neurons and the learning time of the neural network. A fair agreement was obtained when comparing the neural network results with the exact results. For example, the maximum of the density function, with a precision of 0.4% for the percentual relative error, was obtained for 64 neurons. PMID- 11277717 TI - On finding nonisomorphic connected subgraphs and distinct molecular substructures. AB - The problem of finding all nonisomorphic subgraphs of a given graph (all distinct substructures of a given molecular structure) is discussed. A computer program is introduced that first generates all connected subgraphs and then uses a combination of well-discriminating graph invariants to eliminate duplicates. The program is broadly applicable, in particular for molecular graphs which may or may not contain unsaturation or heteroatoms. The number of distinct substructures (Ns), proposed earlier as a measure of a compound's complexity which takes into account its symmetry, is thus automatically obtained. As was to be expected, due to the nature of the problem the computational effort increases exponentially with problem size, whence in most cases complexity measures other than Ns are to be preferred. PMID- 11277718 TI - QSAR study of steroid benchmark and dipeptides based on MEDV-13. AB - A molecular electronegativity distance vector based on 13 atomic types, called MEDV-13, is a descriptor for predicting the biological activities of molecules based on the quantitative structure-activity relations (QSAR). The MEDV-13 uses a modified electrotopological state (E-state) index to substitute for the relative eletronegativity (q) of non-hydrogen atoms in the molecule of interest in the MEDV and a topological distance for the relative distance (d) in the MEDV. For an organic molecule containing several chemical elements such as C, H, O, N, S, F, Cl, Br, I, and P, the MEDV-13 includes at best 91 descriptors. Then it is essential to employ a principal component regression (PCR) technique to derive a QSAR model relating the biological activities to the MEDV-13. The MEDV-13 is used to study the QSAR of the corticosteroid-binding globulin (CBG) binding affinity of the steroids and the activity inhibiting angiotensin-converting enzyme (ACE) of dipeptides, and resulting models have a comparable quality to the current three-dimensional (3D) methods such as CoMFA though the MEDV-13 is a descriptor based on two-dimensional topological information. PMID- 11277719 TI - Similarity calculations using two-dimensional molecular representations. AB - Molecular similarity calculations are important for rational drug design. Time constraints prevent these techniques being used on large data sets or on large molecules. By reducing the molecular representation to a two-dimensional form, the alignment of the molecules can be greatly speeded up. The accuracy of the resulting similarity values can be improved by using a neural network. PMID- 11277720 TI - Structural diversity of small molecule libraries. AB - A novel method for assessing structural diversity is presented. Maximum common subgraph identity is used as the measure of similarity between two chemical structures. A conditional probability treatment of similarity distributions for libraries of chemical structures is used to define diversity. This evaluation method together with the evaluation of traditional physicochemical properties is used to assess a large number of chemical libraries and to understand structural differences between these. PMID- 11277722 TI - Prediction of drug solubility by the general solubility equation (GSE). AB - The revised general solubility equation (GSE) proposed by Jain and Yalkowsky is used to estimate the aqueous solubility of a set of organic nonelectrolytes studied by Jorgensen and Duffy. The only inputs used in the GSE are the Celsius melting point (MP) and the octanol water partition coefficient (K(ow)). These are generally known, easily measured, or easily calculated. The GSE does not utilize any fitted parameters. The average absolute error for the 150 compounds is 0.43 compared to 0.56 with Jorgensen and Duffy's computational method, which utilitizes five fitted parameters. Thus, the revised GSE is simpler and provides a more accurate estimation of aqueous solubility of the same set of organic compounds. It is also more accurate than the original version of the GSE. PMID- 11277721 TI - Metric and multidimensional scaling: efficient tools for clustering molecular conformations. AB - The application of metric and multidimensional scaling to conformer ensembles was demonstrated in this work. An automated process was devised to cluster and assign group memberships and cluster representatives. The method allows rapid clustering, leading to intuitive results that can be visually inspected. Multidimensional scaling was found to be superior to metric scaling for clustering conformers. The performance of different hierarchical clustering algorithms was compared using multidimensional plots, and the group average method was found to perform best. PMID- 11277723 TI - Correlation of the solubilities of gases and vapors in methanol and ethanol with their molecular structures. AB - Two four-parameter quantitative structure-property relations, with R2 = 0.95 and R2 = 0.97, respectively, gave good correlations for the solubilities of 87 gases and vapors in methanol and 61 in ethanol. All the descriptors used are derived solely from the structures of the molecules, making it possible to predict solubilities for unavailable or unknown compounds. PMID- 11277724 TI - Topology of membrane proteins. AB - Integral membrane proteins play important roles in living cells. Due to difficulties of experimental techniques, theoretical approaches, i.e., topology prediction methods, are important for structure determination of this class of proteins. Here we show a detailed comparison of transmembrane topology prediction methods. According to this comparison, we conclude that the topology of integral membrane proteins is determined by the maximum divergence of the amino acid composition of sequence segments. These segments are located in different areas of the cell, which can be characterized by different physicochemical properties. The results of these prediction methods compared to the X-ray diffraction data of several transmembrane proteins will also be discussed. PMID- 11277726 TI - Codes in platonic, Archimedean, Catalan, and related polyhedra: a model for maximum addition patterns in chemical cages. AB - The notion of d-code is extended to general polyhedra by defining maximum sets of vertices with pairwise separation > or =d. Codes are enumerated and classified by symmetry for all regular and semiregular polyhedra and their duals. Partial results are also given for the series of medials of Archimedean polyhedra. In chemistry, d-codes give a model for maximal addition to or substitution in polyhedral frameworks by bulky groups. Some illustrative applications from the chemistry of fullerenes and boranes are described. PMID- 11277725 TI - New description of molecular chirality and its application to the prediction of the preferred enantiomer in stereoselective reactions. AB - A new representation of molecular chirality as a fixed-length code is introduced. This code describes chiral carbon atoms using atomic properties and geometrical features independent of conformation and is able to distinguish between enantiomers. It was used as input to counterpropagation (CPG) neural networks in two different applications. In the case of a catalytic enantioselective reaction the CPG network established a correlation between the chirality codes of the catalysts and the major enantiomer obtained by the reaction. In the second application-enantioselective reduction of ketones by DIP-chloride-the series of major and minor enantiomers produced from different substrates were clustered by the CPG neural network into separate regions, one characteristic of the minor products and the other characteristic of the major products. PMID- 11277727 TI - Getting discriminant functions of antibacterial activity from physicochemical and topological parameters. AB - Linear discriminant analysis has been demonstrated to be a very useful tool in the selection and design of new drugs. Up to now we have used it through the search of a topological pattern of activity. In this work our goal is to calculate a complete set of physicochemical parameters using semiempirical (quantum chemical) calculations as well as topological indices (TIs) and try to find out any discriminant function for antibacterial activity through the combined use of both types of descriptors. The physicochemical parameters, such as heat of formation, HOMO, LUMO, dipole moment, polarizability, hyperpolarizability, PM3 generated IR vibrational frequencies, etc., were calculated using PM3 Hamiltonian implemented within the MOPAC97 package. Among the TIs, connectivity as well as topological charge indices stands as the most representatives. The obtained results suggest that one of the maxima and minima vibrational frequencies play an important role in the antibacterial activity. These frequencies are associated with the torsional molecular vibration (N3) and the stretching vibration (N5) of X-H groups (X = C, N, O). Furthermore, the differences between the maxima and minima values showed an even better discriminant ability than the values themselves. The additional use of the topological indices provided a clear improvement in the discriminant function and also provided a straightforward way to predict the values of such frequencies, so that the results can be applied to a large set of compounds searching for new candidates as antibacterials. PMID- 11277729 TI - Application of BCUT metrics and genetic algorithm in binary QSAR analysis. AB - The application of three-dimensional H-suppressed BCUT metrics (BCUTs) in binary QSAR analysis was investigated using carbonic anhydrase II inhibitors and estrogen receptor ligands as test cases. Variable selection was accomplished with a genetic algorithm (GA). Highly predictive binary QSAR models were obtained for both sets of compounds within 200 GA generations. The derived binary QSAR models were validated with two sets of compounds not included in the training sets. The results indicate that BCUTs are very useful molecular descriptors, and the genetic algorithm is a very efficient variable selection tool in binary QSAR analysis. PMID- 11277728 TI - Mini-fingerprints detect similar activity of receptor ligands previously recognized only by three-dimensional pharmacophore-based methods. AB - Mini-fingerprints (MFPs) are short binary bit string representations of molecular structure and properties, composed of few selected two-dimensional (2D) descriptors and a number of structural keys. MFPs were specifically designed to recognize compounds with similar activity. Here we report that MFPs are capable of detecting similar activities of some druglike molecules, including endothelin A antagonists and alpha(1)-adrenergic receptor ligands, the recognition of which was previously thought to depend on the use of multiple point three-dimensional (3D) pharmacophore methods. Thus, in these cases, MFPs and pharmacophore fingerprints produce similar results, although they define, in terms of their complexity, opposite ends of the spectrum of methods currently used to study molecular similarity or diversity. For each of the studied compound classes, comparison of MFP bit settings identified a consensus or signature pattern. Scaling factors can be applied to these bits in order to increase the probability of finding compounds with similar activity by virtual screening. PMID- 11277730 TI - Prediction of surface tension, viscosity, and thermal conductivity for common organic solvents using quantitative structure-property relationships. AB - Predictive models for the surface tension, viscosity, and thermal conductivity of 213 common organic solvents are reported. The models are derived from numerical descriptors which encode information about the topology, geometry, and electronics of each compound in the data set. Multiple linear regression and computational neural networks are used to train and evaluate models based on statistical indices and overall root-mean-square error. Eight-descriptor models were developed for both surface tension and viscosity, while a nine-descriptor model was developed for thermal conductivity. In addition, a single nine descriptor model was developed for prediction of all three properties. The results of this study compare favorably to previously reported prediction methods for these three properties. PMID- 11277731 TI - Prediction of C60 solubilities from solvent molecular structures. AB - Models predicting fullerene solubility in 96 solvents at 298 K were developed using multiple linear regression and feed-forward computational neural networks (CNN). The data set consisted of a diverse set of solvents with solubilities ranging from -3.00 to 2.12 log (solubility) where solubility = (1 x 10(4))(mole fraction of C60 in saturated solution). Each solvent was represented by calculated molecular structure descriptors. A pool of the best linear models, as determined by rms error, was developed, and a CNN model was developed for each of the linear models. The best CNN model was chosen based on the lowest value of a specified cost function and had an architecture of 9-3-1. The 76-compound training set for this model had a root-mean-square error of 0.255 log solubility units, while the 10-compound cross-validation set had an rms error of 0.253. The 10-compound external prediction set had an rms error of 0.346 log solubility units. PMID- 11277732 TI - QSAR modeling with the electrotopological state: TIBO derivatives. AB - Quantitative structure-activity relationships (QSAR), based on the atom level E state indices and calculated molecular properties (log P, MR), have been developed for the affinity of a large set of TIBO derivatives against HIV-1 reverse transcriptase (HIV-1 RT) utilizing multiple linear regression techniques. A model with five descriptors, including four atom level E-state indices (carbon atoms 2, 4, 8, and 9) and calculated log P, showed good statistics both in the regression (r2 = 0.85 and s = 0.52) and leave-one-out cross-validation (q2 = 0.80 and s(PRESS) = 0.56) for the training set of 41 compounds. The statistics for the prediction of anti-HIV activity in the test set of 24 TIBO derivatives were r2 = 0.80 and s = 0.64, respectively. The model descriptors indicate the importance of lipophilic and electronic contributions toward HIV-1 RT inhibition of TIBO derivatives used in this study. PMID- 11277733 TI - Exploring chemical rings in a simple topological-descriptor space. AB - A new method for organizing chemical rings based on their topology is presented. It uses three simple descriptors that characterize separate aspects of ring topology. These descriptors are integers and can thus be interpreted as the coordinates of discrete cells in a three-dimensional space. The descriptor values of any ring topology correspond to the coordinates of some cell. A database of rings can be distributed in this descriptor space by assigning each of them to the corresponding cell. This approach is applied to a database of 40 182 different ring topologies, derived from a comprehensive collection of chemical rings extracted from the CAS Registry File. This database is distributed among 7387 cells, and the population statistics and spatial distribution of these cells are discussed. An examination of selected cells shows that ring topologies which are similar tend to be close together in descriptor space. Some results of using this space to study ring diversity are presented. It is found that the distribution of the ring-topology database is not highly compact but has many significant voids. It is also found that the distribution of medicinally relevant rings in this space shows the influence of certain structural constraints on drug molecules. PMID- 11277734 TI - Estimation of the aqueous solubility of organic molecules by the group contribution approach. AB - Several group contribution methods to estimate the aqueous solubility of organic molecules are proposed and evaluated for their ability to predict the water solubility of new molecules. The learning set consisted of 1168 organic compounds with experimental data taken from the literature after critical evaluation. The best method, based on a new fragment atom scheme, leads to a squared correlation coefficient of 0.95 and an average absolute calculation error of 0.50 log unit, which is superior to other group contribution methods currently available. One of the advantages of this model is that it has upper and lower limits so that the predicted solubilities cannot be unrealistily high or low. PMID- 11277735 TI - Evaluation of ligand overlap by atomic parameters. AB - The overlap of ligand atoms has been analyzed for 32 common enzyme systems. The ligand alignment was determined by superposition of the experimentally determined protein structures. Comparison of the overlapping atoms in terms of atomic contribution to partition coefficient and molar refractivity shows that in most cases ligand atoms overlap with atoms of a similar nature, as should be expected. A new statistic, the mean maximal atomic deviation (MeMAD), is determined and validated for each system studied by comparison to randomized data. In almost all cases, the MeMAD is well separated from the randomized outcome. This result indicates the validity of the atom-based physicochemical parameters in 3-D QSAR methods. PMID- 11277736 TI - Modeling of nontraditional structures of carbon. AB - The present contribution deals with one of the possible structure types of nongraphitic carbon adsorbents. The proposed structure consists of infinite strips of condensed aromatic rings arranged into a hexagonal honeycomb-like structure with the edge formed only by sp2 carbon atoms. Quantum chemical calculations were performed at the UHF/STO-3G level using the CRYSTAL95 program package for periodical approach. The extremely high adsorption ability of some low-density paramagnetic carbon forms can be elucidated by the presence of the sp2 honeycomb carbon structure and the specific properties of its electronic structure. PMID- 11277737 TI - A quantum mechanical/neural net model for boiling points with error estimation. AB - We present QSPR models for normal boiling points employing a neural network approach and descriptors calculated using semiempirical MO theory (AM1 and PM3). These models are based on a data set of 6000 compounds with widely varying functionality and should therefore be applicable to a diverse range of systems. We include cross-validation by simultaneously training 10 different networks, each with different training and test sets. The predicted boiling point is given by the mean of the 10 results, and the individual error of each compound is related to the standard deviation of these predictions. For our best model we find that the standard deviation of the training error is 16.5 K for 6000 compounds and the correlation coefficient (R2) between our prediction and experiment is 0.96. We also examine the effect of different conformations and tautomerism on our calculated results. Large deviations between our predictions and experiment can generally be explained by experimental errors or problems with the semiempirical methods. PMID- 11277738 TI - Neural network based temperature-dependent quantitative structure property relations (QSPRs) for predicting vapor pressure of hydrocarbons. AB - A neural network based quantitative structure-property relationship (QSPR) was developed for the vapor pressure-temperature behavior of hydrocarbons based on a data set for 274 compounds. The optimal QSPR model was developed based on a 7-29 1 back-propagation neural network architecture using valance molecular connectivity indices (1chi(v), 3chi(v), 4chi(v)), molecular weight, and temperature as input parameters. The average absolute errors in vapor pressure predictions for the test, validation, and overall data sets were 8.2% (0.036 log P units or 23.2 kPA), 9.2% (0.039 log P units or 26.8 kPA), and 10.7% (0.046 log P units or 31.1 kPA), respectively. The performance of the QSPR for temperature dependent vapor pressure, which was developed from a simple set of molecular descriptors, displayed accuracy of better than or well within the range of other available estimation methods. PMID- 11277739 TI - Glanvillic acids A and B and methyl capucinoate A, new metabolites isolated from the Caribbean sponges Plakortis halichondrioides and Plakinastrella onkodes. AB - Glanvillic acids A (2) and B (3) and the cytotoxic cyclic peroxides methyl capucinoate A (4) and 5 were isolated from the Dominican marine sponges Plakortis halichondrioides and Plakinastrella onkodes, respectively. The structures have been elucidated by spectroscopic analysis of 4 and 5 and of methyl glanvillates A (6) and B (7). PMID- 11277740 TI - New eudesmane sesquiterpenes from the root of Lindera strychnifolia. AB - Strychnistenolide (1) and its acetate 2 were isolated from the root of Lindera strychnifolia, along with a novel rearranged type of secoeudesmane, strychnilactone (3). Their structures were elucidated by extensive analysis of their NMR spectra, including 2D NMR techniques, together with an X-ray analysis for 3. Strychnistenolide exists as a single stereoisomer in CHCl(3), but in pyridine is epimerized. PMID- 11277741 TI - Six new diarylheptanoids from the seeds of Alpinia blepharocalyx. AB - Chromatographic separation of part of an EtOH extract of the seeds of Alpinia blepharocalyx resulted in the isolation of six new (1-6) and two known (7, 8) diarylheptanoids together with 12 known compounds. The structures of the new compounds, including their absolute stereochemistry, were elucidated by spectroscopic and chemical methods as (3S,5S)- (1) and (3S,5R)-3-hydroxy-5 methoxy-1-(4-hydroxyphenyl)-7-phenyl-6E-heptene (2), (3S,5S)- (3) and (3S,5R)-3 hydroxy-5-ethoxy-1-(4-hydroxyphenyl)-7-phenyl-6E-heptene (4), (3S)-methoxy-1,7 bis(4-hydroxyphenyl)-6E-hepten-5-one (5), and 1,7-bis(4-hydroxyphenyl)hepta-4E,6E dien-3-one (6). Among the isolated compounds, 5, (3S,5S)-3,5-dihydroxy-1,7-bis(4 hydroxyphenyl)heptane (8), 4'-hydroxy-5,6-dehydrokawain (14), and/or phloroglucinol (20) showed significant antiproliferative activity against murine colon 26-L5 carcinoma (ED(50): 5, 5.2 microM; 8, 12.8 microM; 14, 20.7 microM; 20, 26.4 microM) and human HT-1080 fibrosarcoma (ED(50): 5, 10.1 microM; 14, 20.1 microM; 20, 20.9 microM) cells. PMID- 11277743 TI - Cheilanthane sesterterpenes, protein kinase inhibitors, from a marine sponge of the genus Ircinia. AB - Four cheilanthane sesterterpenoids, 25-hydroxy-13(24),15,17-cheilanthatrien-19,25 olide (1), 13,16-epoxy-25-hydroxy-17-cheilanthen-19,25-olide (2), 25-hydroxy 13(24),17-cheilanthadien-16,19-olide (3), and 16,25-dihydroxy-13(24),17 cheilanthadien-19,25-olide (4), were isolated from the marine sponge Irciniasp. Compounds 1, 3, and 4 are new natural products. The four compounds inhibit MSK1 (mitogen and stress activated kinase) and MAPKAPK-2 (mitogen activated protein kinase activated protein kinase), two protein kinases involved in mitogen and stress signal transduction. PMID- 11277742 TI - Isoterchebulin and 4,6-O-isoterchebuloyl-D-glucose, novel hydrolyzable tannins from Terminalia macroptera. AB - Two new hydrolyzable tannins, isoterchebulin (1) and 4,6-O-isoterchebuloyl-D glucose (2), together with six known tannins, 3-8, were isolated from the bark of Terminalia macroptera. Their structures were elucidated by extensive 1D and 2D NMR studies, MS, and chemical transformations. Biological activities of all compounds were evaluated against the snail Biomphalaria glabrata, the bacteria Bacillus subtilis and Pseudomonas fluorescens, the nematode Caenorhabditis elegans, and four cancer cell lines (Hep G2, MCF-7/S, MDA-MB-231, and 5637 cells). All compounds except 3 showed antimicrobial activities against B. subtilis (MIC 8-64 microg/mL), whereas only 1 was active against C. elegans (100 microg/mL) and B. glabrata(LC(100) = 60 microg/mL). 3 and 8 were toxic against 5637 cells with LC(50) = 84.66 and 41.40 microM, respectively. PMID- 11277744 TI - Pitipeptolides A and B, new cyclodepsipeptides from the marine cyanobacterium Lyngbya majuscula. AB - Two new cyclodepsipeptides have been isolated from a population of the marine cyanobacterium Lyngbya majuscula collected at Piti Bomb Holes, Guam. They appear to be unique to this particular Guamanian collection and have been named pitipeptolides A (1) and B (2). Their structures have been elucidated by spectroscopic techniques and by characterization of degradation products. Distinctive features include the presence of a 2,2-dimethyl-3-hydroxy-7-octynoic acid residue in 1 and a 2,2-dimethyl-3-hydroxy-7-octenoic acid residue in 2, previously shown to be biosynthetic signatures of cyanobacterial metabolites. Pitipeptolides A (1) and B (2) exhibit weak cytotoxicity against LoVo cancer cells, but possess moderate antimycobacterial activity and stimulate elastase activity. PMID- 11277745 TI - Characterization of spirolides a, c, and 13-desmethyl c, new marine toxins isolated from toxic plankton and contaminated shellfish. AB - Three additional marine toxins, spirolides A (1), C (3), and 13-desmethyl-C (7), were isolated from contaminated scallops and phytoplankton collections obtained from a Nova Scotian aquaculture site, as well as from batch cultures of the dinoflagellate Alexandrium ostenfeldii obtained as a single-cell isolate from these phytoplankton assemblages. The structures of these new spirolide derivatives, characterized by mass spectrometry and NMR, indicate a close relationship with spirolides B (2) and D (4) isolated previously from contaminated shellfish in the same area. All of these compounds display "fast acting" toxicity in the traditional bioassay used for monitoring shellfish, and this is related to the presence of a cyclic imine function in all these compounds. Those spirolides containing a vicinal dimethyl group in the seven membered ring are resistant to oxalic acid hydrolysis, whereas those that do not are readily hydrolyzed. These observations suggest that the extra methyl group on the seven-membered imine ring of 3, 4, and 7 appears to block the process of imine hydrolysis perhaps by stereochemical interference. PMID- 11277746 TI - Three new phenolic compounds from a manipulated plant cell culture, Mirabilis jalapa. AB - Bioassay-guided fractionation of an organic extract of the cell mass from a manipulated plant cell culture of Mirabilis jalapa led to the isolation and subsequent identification of three new phenolic compounds, 1, 2, and 3. The isoflavone 1 and dehydrorotenoid 2 were identified as the principal antifungal principles from this plant cell culture with IC(50)'s of 25 and 48 microg/mL, respectively, against the test organism, Candida albicans DSY1024. The rotenoid 3 was inactive at 200 microg/mL in this assay. PMID- 11277747 TI - Briaexcavatolides K-N, new briarane diterpenes from the gorgonian Briareum excavatum. AB - Four new briarane diterpenes, briaexcavatolides K-N (1-4), along with a known diterpene, 5, have been isolated from the Taiwanese gorgonian Briareum excavatum. The structures of the new metabolites were established by extensive spectral analyses. Furthermore, the structure, including the relative configuration of briaexcavatolide K (1), was confirmed by a single-crystal X-ray analysis. Briaexcavatolides K and L (1 and 2) are the only briarane diterpenes known to possess hydroxyl groups at the C-8beta and C-17alpha positions, respectively. Cytotoxicity of these metabolites toward various cancer cell lines also is described. PMID- 11277748 TI - Three new furan derivatives and a new fatty acid from a Taiwanese marine sponge Plakortis simplex. AB - Three new furan derivatives, plakorsins A-C (6-8), together with a new fatty acid, plakortic acid (9), have been isolated from the Taiwanese marine sponge Plakortis simplex in addition to the known metabolites chondrillin (1), 6-epi chondrillin (2), 2-oxo-2,5-dihydrofuran-5-acetic acid methyl ester (methyl 1,4 epoxy-1-oxo-2-hexenoate) (3), dimethyl beta-ketoadipate (4), and the monomethyl ester of cis,cis-muconic acid (5). The structures of these compounds were established mainly on the basis of spectral data and chemical methods. Biological studies revealed that compound 7 exhibited cytotoxic activity against COLO-250 and KB-16 cells, and compound 1 is active against KB-16 cells. PMID- 11277749 TI - Five novel neuroprotective triterpene esters of Ulmus davidiana var. japonica. AB - Investigation of the constituents of the stem and root barks of Ulmus davidiana var. japonica resulted in the isolation of five new triterpene esters named ulmicin A-E (1-5). Using several spectroscopic techniques, their structures were determined to be 3beta,11alpha,15alpha-trihydroxylup-20(29)-ene-11-(3'-methoxy-4' hydroxybenzoyl ester) (1), 3beta,11alpha,15alpha-trihydroxylup-20(29)-ene-11-(4' hydroxybenzoyl ester) (2), 3beta,11alpha,15alpha-trihydroxylup-20(29)-ene-11-(3' methoxy-4'-hydroxybenzoyl)-15-(4'-hydroxybenzoyl ester) (3), 3beta,11alpha,15alpha-trihydroxylup-20(29)-ene-11,15-di(3'-methoxy-4' hydroxybenzoyl ester) (4), and 3beta,11alpha,15alpha-trihydroxylup-20(29)-ene-11 (3'-methoxy-4'-hydroxybenzoyl)-15-(benzoyl ester) (5). All five compounds showed significant neuroprotective activities against glutamate-induced neurotoxicity in primary cultures of rat cortical cells. PMID- 11277750 TI - Regio- and stereoselectivity in the coupling reaction of secologanin with dopamine derivatives. AB - The coupling reaction of tetraacetylsecologanin with dopamine and its N-benzyl derivative was investigated. In both series, stereoisomers at C-1, as well as regioisomer normal and neo compounds, were formed. Moreover, the N-unsubstituted products were partially lactamized, and the N-benzyl derivatives epimerized at C 1. In the products, the R configuration of C-1 over the S and the formation of the normal structure over the neo one predominated. The epimerization of both epimers gave an equilibrium of R and S in a ratio of 7:3 and was interpreted by cleavage of the C-1-N-2 bond. The fact that lactamization was much faster in the R than in the S series was explained on the basis of the supposed transition states. The structure, the configuration of C-1, and in several cases the conformations were established by detailed NMR studies and supported by chemical correlations. PMID- 11277751 TI - Cortamidine oxide, a novel disulfide metabolite from the New Zealand basidiomycete (mushroom) Cortinarius species. AB - Three disulfide metabolites were isolated from the fruiting bodies of the basidiomycete (mushroom) Cortinarius sp., collected in the Catlins, New Zealand. The structures of these compounds were determined as the unsymmetrical disulfide cortamidine oxide (1), 2,2'-dithiobis(pyridine N-oxide) (2), and the symmetrical disulfide 3. Both 1 and 2 showed significant antimicrobial activity and cytotoxicity. 2,2'-Dithiobis(pyridine N-oxide) (2) and the symmetrical disulfide 3 are assumed to be artifacts of the isolation procedure. PMID- 11277752 TI - Resolution and absolute configuration of naturally occurring auronols. AB - Resolution of racemic 2-benzyl-2-hydroxy-1-benzofuran-3(2H)-ones (auronols) and CD data of the ensuing enantiomers permit assessment of the absolute configuration at the single stereocenter. PMID- 11277753 TI - Synthesis of (+)-cyclozonarone and the absolute configuration of naturally occurring (-)-cyclozonarone. AB - Synthesis of (+)-cyclozonarone (1) has been achieved using (-)-polygodial (3) as chiral starting material. The absolute configuration of naturally occurring (-) cyclozonarone was established as 5R,10R by comparison of spectral data and optical rotation with those of (+)-cyclozonarone. PMID- 11277754 TI - Hipposulfates A and B, new sesterterpene sulfates from an Okinawan sponge, Hippospongia cf. metachromia. AB - Two new sesterterpene sulfates, hipposulfates A (1) and B (2), have been isolated from an Okinawan sponge, Hippospongia cf. metachromia and their structures elucidated by interpretation of spectroscopic data. Both compounds contain an enolsulfate functionality. Hipposulfate A (1) showed moderate cytotoxicity. PMID- 11277755 TI - A simple isolation method for the major catechins in green tea using high-speed countercurrent chromatography. AB - An efficient protocol for the preparative purification of the major tea catechins (-)-epicatechin gallate (3) and epigallocatechin gallate (5) has been developed, employing liquid-liquid partitioning, high-speed countercurrent chromatography, and gel chromatography for final purification. The method is suitable for scale up to significantly larger quantities. PMID- 11277756 TI - Isolation and structures of haterumadioxins A and B, cytotoxic endoperoxides from the Okinawan sponge Plakortis lita. AB - Cytotoxic endoperoxides, haterumadioxins A (1) and B (2), were isolated from the Okinawan sponge Plakortis lita. Their structures were determined by spectroscopic analysis. The absolute stereostructure of 1 was determined by degradation reactions and the modified Mosher's method. Haterumadioxins showed significant cytotoxicity against 38 human cancer cell lines. PMID- 11277757 TI - Three new triterpenes from the seeds of Combretum quadrangulare and their hepatoprotective activity. AB - Three new triterpenes of the lupane type, 2alpha,6beta-dihydroxybetulinic acid (1) and 6beta-hydroxyhovenic acid (2), and an oleanane type, 6beta-hydroxyarjunic acid (3), together with several known compounds, have been isolated from the MeOH extract of the seeds of Combretum quadrangulare. The structures of these compounds were elucidated on the basis of spectroscopic analysis, and their hepatoprotective activities were tested for D-GalN/TNF-alpha-induced cell death in primary cultured mouse hepatocytes. PMID- 11277758 TI - Five new insecticidal sesquiterpenoids from Celastrus angulatus. AB - Five new sesquiterpene polyol esters were isolated from the root bark of Celastrus angulatus by bioassay-guided fractionation. Their structures were determined by spectral data interpretation as 1alpha,2alpha,6beta,8beta,13 pentaacetoxy-9beta-benzoyloxy-4beta-hydroxy-beta-dihydroagarofuran (1), 1alpha,2alpha,6beta-triacetoxy-8alpha-(beta-furancarbonyloxy)-9beta-benzoyloxy-13 isobutanoyloxy-4beta-hydroxy-beta-dihydroagarofuran (2), 1alpha,2alpha,6beta triacetoxy-8beta-isobutanoyloxy-9beta-(beta-furancarbonyloxy)-13-(alpha methyl)butanoyloxy-4beta-hydroxy-beta-dihydroagarofuran (3), 1alpha,2alpha,6beta triacetoxy-8alpha,13-diisobutanoyloxy-9beta-benzoyloxy-4beta-hydroxy-beta dihydroagarofuran (4), and 1alpha,2alpha,6beta-triacetoxy-8alpha-isobutanoyloxy 9beta-benzoyloxy-13-(alpha-methyl)butanoyloxy-4beta-hydroxy-beta dihydroagarofuran (5). Compounds 1-5 exhibited insecticidal activity against the larval of Mythimna separata. PMID- 11277759 TI - Total synthesis of heliannone A and (R,S)-heliannone B, two bioactive flavonoids from Helianthus annuus cultivars. AB - The total synthesis of heliannone A (1) and (R,S)-heliannone B (2), two bioactive flavonoids originally isolated from Helianthus annuus cultivars, is described. PMID- 11277760 TI - 3-Acetoxyspathulenol, a new aromadendrane-type natural product from the soft Coral Parerythropodium fulvum. AB - From the lipophilic extract of the soft coral Parerythropodium fulvum a new sesquiterpene with an aromadendrane-type carbon skeleton, 3-acetoxyspathulenol (1), was isolated, and the known compounds spathulenol (2) and tridensenone (3) were identified. The structure of the new compound was determined by interpretation of its spectroscopic data, including 1D and 2D (1)H and (13)C NMR (COSY, HMQC, HMBC) and MS. PMID- 11277761 TI - Microbial transformation of hypoestenone. AB - Preparative-scale fermentation of hypoestenone (1) with Mucor ramannianus (ATCC 9628) has resulted in the isolation of 8(9)alpha-epoxyhypoestenone (2) and the new metabolites 8(9)alpha-epoxy-12,13-anhydrohypoestenone (3), 17 hydroxyhypoestenone (4), 13alpha,18-dihydroxyhypoestenone (5), and 13beta,18 dihydroxyhypoestenone (6). Structure elucidation of these metabolites was based primarily on 1D and 2D NMR analyses. PMID- 11277762 TI - Two new lactones with mosquito larvicidal activity from three Hortonia species. AB - The structures of the butenolides 1 and 2, isolated from the endemic plants Hortonia floribunda, H. angustifolia, and H. ovalifolia, collected in Sri Lanka, have been elucidated by spectroscopic analysis. These compounds exhibited potent mosquito larvicidal activity against the second instar larvae of Aedes aegypti. PMID- 11277763 TI - New diterpene alkaloids from the roots of Spiraea japonica. AB - Chemical investigation on an ethanol extract from the roots of Spiraea japonica var. acuta resulted in the isolation of three new diterpene alkaloids named spiramide (1) and spiratine A (2) and spiratine B (3). Structures of 1-3 were elucidated primarily on the basis of 1D and 2D NMR experiments. PMID- 11277764 TI - Inhibition of glucose and dehydroascorbic acid uptakes by resveratrol in human transformed myelocytic cells. AB - Resveratrol (trans-3,5,4'-trihydroxystilbene) is a natural phytochemical found in grapes and wine. Numerous biological effects of resveratrol have been reported in the last 10 years. In this paper, the competitive inhibition of intracellular uptake of glucose and dehydroascorbic acid in U937 and HL-60 cells by resveratrol is reported. PMID- 11277765 TI - Potent antiviral potamogetonyde and potamogetonol, new furanoid labdane diterpenes from Potamogeton malaianus. AB - New furanoid labdane diterpenes, potamogetonyde (3) and potamogetonol (4), together with two known compounds, potamogetonin (1) and 15,16-epoxy-12-oxo 8(17),13(16),14-labdatrien-20,19-olide (2), were isolated from the CH(2)Cl(2) extract of Potamogeton malaianus. The chemical structures of 1-4 were elucidated by the analyses of their spectral data, mainly by 1D and 2D NMR techniques. Potamogetonyde (3) and potamogetonol (4) exhibited potent antiviral (HSV-1) activity with respective IC(50) values of 8 and 3 microg/mL. Compounds 1-4 possessed cytotoxicity toward insect cells (fall armyworm and mosquito larvae, IC(50) of 11-72 microg/mL). Furanoid diterpenes 3 and 4 also exhibited cytotoxicity against the Vero cell line with respective IC(50)'s of 31 and 28 microg/mL, while 1 and 2 were inactive at 50 microg/mL. Compounds 1-4 were inactive (at 20 microg/mL) against KB and BC cell lines and showed only weak antimycobacterial activity against Mycobacterium tuberculosis H37Ra with minimum inhibitory concentrations of 50-100 microg/mL. PMID- 11277766 TI - New cytotoxic isomalabaricane triterpenes from the sponge Jaspis species. AB - Three new isomalabaricane triterpenes, 29-hydroxystelliferin E (1), 29 hydroxystelliferin A (2), and stelliferin G (3), together with the known triterpene 3-epi-29-hydroxystelliferin E (4), were isolated from the organic extract of the sponge Jaspis sp. collected in the South Pacific. All the compounds isolated showed antiproliferative activity against melanoma cells (MALME-3M). PMID- 11277767 TI - Oxygenated sesquiterpenoids from a nonpoisonous sardinian chemotype of giant fennel (Ferula communis). AB - The roots of the nonpoisonous chemotype of giant fennel (Ferula communis) from Sardinia afforded a novel cadinanetriol (1), whose structure was established by spectroscopic data and confirmed by synthesis from the E,E-Delta (1(10),5) germacradiene allohedycariol. A number of known compounds were also identified. Despite the lack of morphological differences, a broad chemical diversity exists within giant fennel, underlying the contrasting data on its poisonous properties. PMID- 11277768 TI - A new macrocyclic trichothecene, 12,13-deoxyroridin E, produced by the marine derived fungus Myrothecium roridum collected in Palau. AB - A new macrocyclic trichothecene, 12,13-deoxyroridin E (1), and three known compounds, roridin E (2), verrucarin A (3), and verrucarin J (4), were obtained as cytotoxic components from the marine-derived fungus Myrothecium roridum, isolated in Palau. 12,13-Deoxyroridin E is the second example of a macrocyclic trichothecene possessing a double bond at C-12-C-13 and was about 80-fold less cytotoxic than roridin E, the epoxide variant. PMID- 11277769 TI - Daucane sesquiterpenes from Ferula hermonis. AB - The roots of Ferula hermonis Boiss yielded two new daucane esters, 14-(4' hydroxybenzoyloxy)dauc-4,8-diene (1) and 14-(4'-hydroxy-3'-methoxybenzoyloxy)dauc 4,8-diene (2), together with the four known sesquiterpenes jaeschkeanadiol p hydroxybenzoate (3), jaeschkeanadiol benzoate (4), jaeschkeanadiol (5), and epoxyjaeschkeanadiol (6). The identities of the isolated compounds were ascertained primarily using NMR and MS data. Compounds 1 and 3 exhibited antimicrobial activity against Staphylococcus aureus with IC(50) 1.5 and 3.5 microg/mL, respectively, and against Methicillin-resistant S. aureus with IC(50) 2.0 and 4.0 microg/mL, respectively. PMID- 11277770 TI - Spiroiminodihydantoin is the major product of the 8-oxo-7,8-dihydroguanosine reaction with peroxynitrite in the presence of thiols and guanosine photooxidation by methylene blue. AB - [reaction: see text]. The potent oxidant, peroxynitrite, will oxidize 8-oxo-7,8 dihydroguanosine to give several products. In the presence of a thiol agent, the major final product has been determined to be a spiroiminodihydantoin compound. Additionally, we have found that the spiroiminodihydantoin, and not the previously reported 4-hydroxy-8-oxo-4,8-dihydroguanosine, is the major final product formed during the methylene blue-mediated photooxidation of guanosine. PMID- 11277771 TI - Synthesis of oligopyridines and their metal complexes as precursors to topological stereoisomers. AB - [structure: see text]. Palladium-based carbon-carbon coupling reactions in sequence with halogen-exchange chemistry on a series of heterocycles lead to an efficient synthetic strategy for oligopyridines that bind metal ions such as ruthenium to form coordination racks. The particular structures are designed to form terpyridine subunits for octahedral binding. Reaction of 4,6 diiodopyrimidine produces a "double-bay" terpyridine from which binuclear coordination complexes have been formed. PMID- 11277772 TI - Single-step formation of structurally defined bicyclic peptides via S(N)Ar cyclization. AB - [structure: see text]. A solid-phase methodology for macrocyclization via an S(N)Ar reaction has been developed for the unambiguous formation of bicyclic peptidic compounds in a single cyclization step. The cyclization strategy involves the reaction of a 3,5-dihydroxybenzoyl group with two nitrofluorobenzoyl moieties. The symmetry of the dihydroxy aromatic ring results in a single product, and the remaining nitro groups are subsequently reduced to anilines and acylated. PMID- 11277773 TI - Synthetic studies toward the C5-C20 domain of the azaspiracids. AB - [structure: see text]. An approach toward the C5-C20 THF-fused trioxadispiroketal portion of the azaspiracids is reported. The highly substituted azaspiracid D ring (C16-C19) was prepared by the one-pot conversion of a tetraol into a tetrahydrofuran. Efforts to establish the C10 and C13 spiroketal centers via an oxonium-initiated bis-spiroketalization under both kinetic and thermodynamic conditions have yielded the (10R,13S)-trioxadispiroketal 19 as the major product, which is diastereomeric with the (10R,13R) relative configuration assigned to the azaspiracids. PMID- 11277774 TI - Synthesis of a 2,9-dioxabicyclo[3.3.1]nonane via double intramolecular hetero Michael addition: entry to the F-G ring system of the azaspiracids. AB - [structure: see text]. An effective approach to form a 1,3-disubstituted 2,9 dioxabicyclo[3.3.1]nonane system representing the core of the F and G rings (C28 C34) of the azaspiracid natural products has been developed. The double intramolecular hetero-Michael addition (DIHMA) of a diol upon an ynone generated the bicyclic ketal in a highly diastereoselective fashion. PMID- 11277775 TI - Highly diastereoselective Michael addition of alpha-hydroxy acid derivatives and enantioselective synthesis of (+)-crobarbatic acid. AB - [reaction: see text]. Michael addition of enolates of 2a and 2b to alpha,beta unsaturated esters took place selectively on different faces (Si and Re, respectively) of the double bond to give the corresponding products 4 and 5, respectively, with >98% de. Subsequent hydrolysis of these Micheal adducts gives 3,4-disubstituted gamma-lactones with high enantiomeric excesses. PMID- 11277776 TI - A 2'-acylamido cap that increases the stability of oligonucleotide duplexes. AB - [structure: see text]. Reported here is the synthesis of oligonucleotides containing a 2'-acylamido-2'-deoxyuridine residue at their 3'-terminus. Compared to control sequences bearing a thymidine residue, the quinolone-capped duplexes give higher UV melting points. In the case of (5'-ACGCGU-NA-2')2, where NA denotes a nalidixic acid residue, the melting point increase is up to 22 degrees C over that of (ACGCGT)2. PMID- 11277777 TI - Benzyltriethylammonium dichloroiodate/sodium bicarbonate combination as an inexpensive, environmentally friendly, and mild iodinating reagent for anilines. AB - [reaction: see text]. Monoiodinated anilines were prepared in good to excellent yields by the action of benzyltriethylammonium dichloroiodate on anilines in the presence of sodium bicarbonate and methanol. The iodinating reagent was prepared in an environmentally friendly fashion without the use of organic solvents. PMID- 11277778 TI - Phenylene ethynylene diazonium salts as potential self-assembling molecular devices. AB - [reaction: see text]. Functionalized diazonium salts for molecular electronic devices are prepared by the reaction of the corresponding anilines with NOBF4 in sulfolane-acetonitrile solvent. PMID- 11277779 TI - Diastereoselective synthesis of 1-benzyltetrahydroisoquinoline derivatives from amino acids via 1,4 chirality transfer. Part 1. AB - [structure: see text]. L-(-)-phenylalanine, L-(+)-valine, and L-(-)-proline were used in the diastereoselective synthesis of benzyltetrahydroisoquinoline derivatives. PMID- 11277780 TI - Potent integrin antagonists from a small library of RGD-including cyclic pseudopeptides. AB - [structure: see text]. A small library of cyclic RGD pseudopentapeptides incorporating stereoisomeric 6,5- and 7,5-fused bicyclic lactams was synthesized with the aim of developing active and selective integrin antagonists. The solid phase synthesis and activity of these RGD derivatives is described. The approach led to two of the most active known inhibitors of alpha(V)beta3 receptor. PMID- 11277782 TI - Nonconventional carbon additions to azomethines. aryl amination/indoline synthesis by direct aryl radical addition to azomethine nitrogen. AB - [reaction: see text]. The generality of a new method for aryl amination has been defined. Ketimines derived from o-bromophenethylamine cyclize to the N substituted indoline when treated with nBu3SnH and a radical initiator. The pH neutral conditions tolerate base- and acid-sensitive functionality. The observed regioselectivity is nonconventional for addition reactions involving carbon radicals and carbon-heteroatom pi-bonds. PMID- 11277781 TI - Acylation of indole under Friedel-Crafts conditions-an improved method to obtain 3-acylindoles regioselectively. AB - [reaction: see text]. The reaction of unsubstituted indole with different acylating agents such as acid chlorides, anhydrides, nitriles, and amino acid derivatives in the presence of Lewis acid is reported. PMID- 11277784 TI - Isomerization of linear to angular [3]phenylene and PAHs under flash vacuum pyrolysis conditions. AB - [structure: see text]. Flash vacuum pyrolysis (FVP) of linear [3]phenylene affords its angular counterpart and the same mixture of polycyclic aromatic hydrocarbon isomers as that observed on FVP of angular [3]phenylene. A mechanism, supported by a 13C labeling study, is proposed to explain these results. PMID- 11277783 TI - Effect of a 17alpha-(3-hydroxypropyl)-17beta-acetyl substituent pattern on the glucocorticoid and progestin receptor binding of 11beta-arylestra-4,9-dien-3 ones. AB - [structure: see text]. Replacing the 17alpha-acetoxy substituent in an antiprogestational 17beta-acetyl-11beta-arylestra-4,9-dien-3-one by 3 hydroxypropyl significantly diminished glucocorticoid receptor binding with little effect on progestin receptor binding. PMID- 11277785 TI - Benzobicyclo[8.4.0(1,6)0(8,13]tetradecenones: ring CD-taxoid models by planar tether controlled cycloaddition. AB - [structure: see text]. A general approach to functionalized hexahydroanthracene dione skeletons is described. The planar dicarbonyl triene 15 cyclized spontaneously upon oxidation of the precursor diol 14 to the tricyclicdiketone 16. The adducts 16 and 2 were further transformed to the corresponding epoxides and oxetanes as a model for the D ring of taxoids. PMID- 11277786 TI - 2'-C-branched ribonucleosides. 2. Synthesis of 2'-C-beta-trifluoromethyl pyrimidine ribonucleosides. AB - [structure: see text]. The first synthesis of 2'-C-beta-trifluoromethyl pyrimidine ribonucleosides is described. 1,2,3,5-Tetra-O-benzoyl-2-C-beta trifluoromethyl-alpha-D-ribofuranose (3) is prepared from 1,3,5-tri-O-benzoyl alpha-D-ribofuranose (1) in three steps and converted to 3,5-di-O-benzoyl-2-C beta-trifluoromethyl-alpha-D-1-ribofuranosyl bromide (5). The 1-bromo derivative (5) is found to be a powerful reaction intermediate for the synthesis of ribonucleosides. The reaction of silylated pyrimidines with (5) in the presence of HgO/HgBr2 affords exclusively the beta-anomers (6-8). Deprotection of (6-8) with ammonia in methanol yields the 2'-C-beta-trifluoromethyl nucleosides (9-11). PMID- 11277788 TI - The H-phosphonate approach to the synthesis of phosphopeptides on solid phase. AB - [reaction: see text]. Ammonium tert-butyl H-phosphonate was used for the phosphorylation of Tyr- and Ser-containing peptides synthesized by an Fmoc strategy. This reaction, leading to a monoprotected peptide phosphate, was found to be highly efficient and generally applicable. Moreover, the method employed avoids undesired side reactions during chain elongation (pyrophosphate formation and beta-elimination catalyzed by piperidine). PMID- 11277787 TI - Intramolecular O-arylation of phenols with phenylboronic acids: application to the synthesis of macrocyclic metalloproteinase inhibitors. AB - [reaction: see text]. The copper acetate mediated intramolecular O-arylation of phenols with phenylboronic acid pseudopeptides is the key step in the preparation of macrocyclic biphenyl ether hydroxamic acid inhibitors of collagenase 1 and gelatinases A and B. The intramolecular macrocyclization was found to be mild and tolerant of common chemical functionality. This methodology should provide a general route to macrocyclic biphenyl ethers. PMID- 11277789 TI - Preparation of silyl enol ethers using (bistrimethylsilyl)acetamide in ionic liquids. AB - [reaction: see text]. Ionic liquids have been used for the preparation of silyl enol ethers from aldehydes and ketones with (bistrimethylsilyl)acetamide (BSA) in good yields. PMID- 11277790 TI - An hermaphrodite [2]rotaxane: preparation and analysis of structure. AB - [structure: see text]. The pictured rotaxane is assembled in water by capping a substituted cyclodextrin composed of the wheel and axle components. The unusual dimeric structure of the rotaxane reflects the thermodynamics of the assembly process. In N,N-dimethylformamide, the corresponding monomer is the predominant product. PMID- 11277791 TI - Rigid, cross-conjugated macrocycles: a cyclic alternative to 4,4'-bipyridines in supramolecular chemistry. AB - [structure: see text]. The synthesis of two fully conjugated, rigid macrocyclic analogues to 4,4'-bipyridine is described. The use of these macrocycles in self assembly processes is demonstrated by axial coordination to metalloporphyrins, and one system (R = Me) has been characterized by single-crystal X-ray crystallography. PMID- 11277792 TI - An enantioselective synthesis of benzylidene-protected syn-3,5-dihydroxy carboxylate esters via osmium, palladium, and base catalysis. AB - [reaction: see text]. The enantioselective syntheses of several protected syn-3,5 dihydroxy carboxylic esters have been achieved from the corresponding achiral 1,3 dieneoates. The route relies upon an enantio- and regioselective Sharpless dihydroxylation and a palladium-catalyzed reduction to form delta-hydroxy-1 enoates. The resulting delta-hydroxy-1-enoates are subsequently converted into benzylidene-protected 3,5-dihydroxy carboxylic esters in one step. The benzylidene-protected 3,5-dihydroxy carboxylic esters are produced in good overall yields (25% to 51%) and high enantiomeric excesses (80% to >95%). PMID- 11277793 TI - N-methoxy-N-acylnitrenium ions: application to the formal synthesis of (-) TAN1251A. AB - [structure: see text]. A formal synthesis of the muscarinic M(1) receptor antagonist (-)-TAN1251A (7) from L-tyrosine is described. Central to this venture has been the construction of the 1-azaspiro[4.5]decane skeleton present in the natural product by an N-methoxy-N-acylnitrenium ion-induced spirocyclization. The dienone generated in this transformation, 10, was converted to (-)-TAN1251A via tricycle 9, an intermediate in Kawahara's recent synthesis of racemic 7. PMID- 11277794 TI - Group-selective hydroalumination: a novel route to stereogenic tert-alcohol centers. AB - [reaction: see text]. A new group-selective reaction is described, that is, the hydroalumination of bis-alkynyl alcohols armed with an adjacent stereogenic center, giving stereo-defined tert-alcohols of potential utility in natural product synthesis. PMID- 11277795 TI - Stereoselective halogenations of alkenes and alkynes in ionic liquids. AB - [reaction: see text]. Room-temperature ionic liquids, 1-butyl-3-methylimidazolium hexafluorophosphate, 1-butyl-3-methylimidazolium tetrafluoroborate, 1-butyl-3 methylimidazolium bromide, and 1-butyl-3-methylimidazolium chloride, are used as "green" recyclable alternative to chlorinated solvents for the stereoselective halogenation of alkenes and alkynes. PMID- 11277796 TI - Dicobalt octacarbonyl catalyzed carbonylated cycloaddition of triynes to functionalized tetracycles. AB - [reaction: see text]. We have demonstrated that a dicobalt octacarbonyl catalyzed double [2 + 2 + 1] carbonylative cycloaddition reaction of triyne can be carried out to yield a novel 5.5.5.6 tetracyclic di-enone system. PMID- 11277797 TI - A xanthate transfer radical process for the introduction of the trifluoromethyl group. AB - [reaction: see text]. S-Trifluoromethyl xanthates efficiently add to unactivated alkenes by a radical mechanism to give adducts with a trifluoromethyl group at the least hindered terminus of the olefin. PMID- 11277798 TI - Highly stereoselective hydrocarbation of terminal alkynes via Pt-catalyzed hydrosilylation/Pd-catalyzed cross-coupling reactions. AB - [reaction: see text]. The formal addition of an aryl-H or alkenyl-H bond across a terminal alkyne has been accomplished by the combination of platinum-catalyzed hydrosilylation followed by palladium-catalyzed cross-coupling. The use of the t Bu3P-Pt(DVDS) catalyst in combination with tetramethyldisiloxane gave excellent regio- and stereoselectivity with a number of alkyne substrates. Subsequent, fluoride-promoted cross-coupling proceeded in high yield and stereospecificity for a variety of aryl halides. PMID- 11277799 TI - Convenient and efficient Suzuki-Miyaura cross-coupling catalyzed by a palladium/diazabutadiene system. AB - [structure: see text]. A Pd(OAc)2/diazabutadiene system has been developed for the catalytic cross-coupling of aryl halides with arylboronic acids. A combination of the diazabutadiene DAB-Cy (1, N,N'-dicyclohexyl-1,4-dizabutadiene) and Pd(OAc)2 was found to form an excellent catalyst for the Suzuki-Miyaura cross coupling of various aryl bromides and activated aryl chlorides with arylboronic acids. PMID- 11277801 TI - Creation of novel chiral cryptophanes by a self-assembling method utilizing a pyridyl-Pd(II) interaction. AB - [structure: see text]. This Letter demonstrates the molecular design of novel self-assembled chiral cryptophanes. Mediated by square-planar Pd(II) complexes, racemic pyridyl cyclotriveratrylene derivative rac-2 self-assembles into mixtures of racemic chiral cryptophanes and meso cryptophanes (1), which interconvert with each other, and the rates are remarkably enhanced by the addition of a slight excess of rac-2. On the other hand, optically resolved P-2 or M-2 self-assembles into the chiral cryptophane as the only product. PMID- 11277800 TI - A ring expansion-annulation strategy for the synthesis of substituted azulenes. Preparation and Suzuki coupling reactions of 1-azulenyl triflates. AB - [structure: see text]. A new strategy for the synthesis of substituted azulenes is reported, based on the reaction of beta'-bromo-alpha-diazo ketones with rhodium carboxylates. The key transformation involves intramolecular addition of a rhodium carbenoid to an arene pi-bond, electrocyclic ring opening, beta elimination, tautomerization, and trapping to produce 1-hydroxyazulene derivatives. The synthetic utility of the method is enhanced by the ability of the triflate derivatives to participate in Suzuki coupling reactions, as illustrated in a synthesis of the antiulcer drug egualen sodium (KT1-32). PMID- 11277802 TI - Design and synthesis of novel scaffolds for drug discovery: hybrids of beta-D glucose with 1,2,3,4-tetrahydrobenzo[e][1,4]diazepin-5-one, the corresponding 1 oxazepine, and 2- and 4-pyridyldiazepines. AB - [structure: see text]. We describe the syntheses of novel tricyclic scaffolds that incorporate a fusion of a substituted pyranose ring with the seven-membered rings of 1,2,3,4-tetrahydrobenzo[e][1,4]diazepin-5-one and the corresponding oxazepine and pyridyldiazepine to generate the benzodiazepines, and the related heterocycles. In each instance, the pyranose rings contain three protected hydroxyls, suitable for selective derivatization. PMID- 11277803 TI - Polymer-aided stereodivergent synthesis (PASS): a new concept for the discrete preparation of optical antipodes. AB - [structure: see text]. A new concept for the discrete preparation of optical antipodes is reported. The approach makes use of a cyclization/cleavage procedure that has been applied to polymer-supported quasi-meso compound 1, containing a polymeric leaving group and a regular leaving group. Two-directional cyclization leads to the formation and separation of quasi-enantiomers 2 and 3 simultaneously, with one being immobilized and one free in solution. Treatment of 2 and 3 with appropriate nucleophiles gives discrete enantiomers 4 and 5. PMID- 11277806 TI - Stem cells step closer to the clinic: paralysis partially reversed in rats with ALS-like disease. PMID- 11277807 TI - Blood groups differ on donor deferral. PMID- 11277810 TI - From the Food and Drug Administration. PMID- 11277813 TI - From the Surgeon General. Global mental health: its time has come. PMID- 11277814 TI - Outcomes of angioplasty vs thrombolysis by hospital angioplasty volume. PMID- 11277815 TI - Outcomes of angioplasty vs thrombolysis by hospital angioplasty volume. PMID- 11277817 TI - Timing of antiretroviral treatment initiation. PMID- 11277819 TI - Chlamydia trachomatis and cervical squamous cell carcinoma. PMID- 11277820 TI - Chlamydia trachomatis and cervical squamous cell carcinoma. PMID- 11277821 TI - Chlamydia trachomatis and cervical squamous cell carcinoma. PMID- 11277822 TI - Chlamydia trachomatis and cervical squamous cell carcinoma. PMID- 11277824 TI - Y2k revisited: a human component? PMID- 11277825 TI - Effects of atorvastatin on early recurrent ischemic events in acute coronary syndromes: the MIRACL study: a randomized controlled trial. AB - CONTEXT: Patients experience the highest rate of death and recurrent ischemic events during the early period after an acute coronary syndrome, but it is not known whether early initiation of treatment with a statin can reduce the occurrence of these early events. OBJECTIVE: To determine whether treatment with atorvastatin, 80 mg/d, initiated 24 to 96 hours after an acute coronary syndrome, reduces death and nonfatal ischemic events. DESIGN AND SETTING: A randomized, double-blind trial conducted from May 1997 to September 1999, with follow-up through 16 weeks at 122 clinical centers in Europe, North America, South Africa, and Australasia. PATIENTS: A total of 3086 adults aged 18 years or older with unstable angina or non-Q-wave acute myocardial infarction. INTERVENTIONS: Patients were stratified by center and randomly assigned to receive treatment with atorvastatin (80 mg/d) or matching placebo between 24 and 96 hours after hospital admission. MAIN OUTCOME MEASURES: Primary end point event defined as death, nonfatal acute myocardial infarction, cardiac arrest with resuscitation, or recurrent symptomatic myocardial ischemia with objective evidence and requiring emergency rehospitalization. RESULTS: A primary end point event occurred in 228 patients (14.8%) in the atorvastatin group and 269 patients (17.4%) in the placebo group (relative risk [RR], 0.84; 95% confidence interval [CI], 0.70-1.00; P =.048). There were no significant differences in risk of death, nonfatal myocardial infarction, or cardiac arrest between the atorvastatin group and the placebo group, although the atorvastatin group had a lower risk of symptomatic ischemia with objective evidence and requiring emergency rehospitalization (6.2% vs 8.4%; RR, 0.74; 95% CI, 0.57-0.95; P =.02). Likewise, there were no significant differences between the atorvastatin group and the placebo group in the incidence of secondary outcomes of coronary revascularization procedures, worsening heart failure, or worsening angina, although there were fewer strokes in the atorvastatin group than in the placebo group (12 vs 24 events; P =.045). In the atorvastatin group, mean low-density lipoprotein cholesterol level declined from 124 mg/dL (3.2 mmol/L) to 72 mg/dL (1.9 mmol/L). Abnormal liver transaminases (>3 times upper limit of normal) were more common in the atorvastatin group than in the placebo group (2.5% vs 0.6%; P<.001). CONCLUSION: For patients with acute coronary syndrome, lipid-lowering therapy with atorvastatin, 80 mg/d, reduces recurrent ischemic events in the first 16 weeks, mostly recurrent symptomatic ischemia requiring rehospitalization. PMID- 11277826 TI - Glycine antagonist in neuroprotection for patients with acute stroke: GAIN Americas: a randomized controlled trial. AB - CONTEXT: Elucidation of the ischemic cascade has helped stimulate development of neuroprotective drugs aimed at limiting brain injury in the hours following an ischemic stroke. To date, none of these drugs has shown clinical efficacy. OBJECTIVE: To examine the efficacy of gavestinel (GV150526), an antagonist of the glycine site of the N-methyl-D-aspartate receptor, as a neuroprotective therapy for acute ischemic stroke when administered within 6 hours of symptom onset. DESIGN: The Glycine Antagonist in Neuroprotection (GAIN) Americas trial, a randomized, double-blind placebo-controlled trial with enrollment from April 1998 to October 1999. SETTING: One hundred thirty-two hospital centers across the United States and Canada. PATIENTS: The primary efficacy population consisted of 1367 ischemic stroke patients with a predefined level of limb weakness and functional independence prior to stroke, stratified at randomization by age (75 years) and initial stroke severity (National Institutes of Health [NIH] Stroke Scale scores of 2-5, 6-13, or >/=14). INTERVENTION: Patients were randomly assigned to receive an intravenous loading dose (800 mg) plus 5 maintenance doses (200 mg every 12 hours) of gavestinel (n = 701) or placebo (n = 666) for 3 days. MAIN OUTCOME MEASURE: Functional capability at 3 months, measured by the Barthel Index (BI), with scores trichotomized as dead/0-55, 60 90, and 95-100, compared between the gavestinel and placebo groups. RESULTS: Treatment groups were well matched for baseline characteristics. For each group, median NIH Stroke Scale was 12, median age was 72 years, and median time to treatment was 5.2 hours. No statistically significant improvement on the 3-month BI trichotomy was demonstrated for gavestinel (P =.79). The proportion who were functionally independent (BI score = 95-100) was 39% in the gavestinel group and 37% in the placebo group. No statistically significant difference in 3-month survival was observed using Kaplan-Meier curves (P =.11). No other secondary end point suggested an advantage for gavestinel. Among the 333 patients (24%) who received recombinant tissue-type plasminogen activator, there was also no benefit for gavestinel (P =.53). There were no serious safety issues. CONCLUSION: In this study, gavestinel administered up to 6 hours after an acute ischemic stroke did not improve functional outcome at 3 months. PMID- 11277827 TI - Epidemiology of invasive Streptococcus pneumoniae infections in the United States, 1995-1998: Opportunities for prevention in the conjugate vaccine era. AB - CONTEXT: Pneumococcal polysaccharide vaccine is recommended for elderly persons and adults with certain chronic illnesses. Additionally, a recently licensed pneumococcal 7-valent conjugate vaccine has been recommended for use in young children and could dramatically change the epidemiology of pneumococcal disease. OBJECTIVES: To assess pneumococcal disease burden in the United States, estimate the potential impact of new vaccines, and identify gaps in vaccine recommendations. DESIGN AND SETTING: Analysis of data from the Active Bacterial Core Surveillance (ABCs)/Emerging Infections Program Network, an active, population-based system in 9 states. PATIENTS: A total of 15 860 cases of invasive pneumococcal disease occurring between January 1, 1995, and December 31, 1998. MAIN OUTCOME MEASURES: Age- and race-specific pneumoccocal disease incidence rates per 100 000 persons, case-fatality rates, and vaccine preventability. RESULTS: In 1998, overall incidence was 23.2 cases per 100 000, corresponding to an estimated 62 840 cases in the United States. Incidence was highest among children younger than 2 years (166.9) and adults aged 65 years or older (59.7). Incidence among blacks was 2.6 times higher than among whites (95% confidence interval [CI], 2.4-2.8). Overall, 28.6% of case-patients were at least 65 years old and 85.9% of cases in this age group were due to serotypes included in the 23-valent polysaccharide vaccine; 19.3% of case-patients were younger than 2 years and 82.2% of cases in this age group were due to serotypes included in the 7-valent conjugate vaccine. Among patients aged 2 to 64 years, 50.6% had a vaccine indication as defined by the Advisory Committee on Immunization Practices (ACIP). The case-fatality rate among patients aged 18 to 64 years with an ACIP indication was 12.1% compared with 5.4% for those without an indication (relative risk, 2.2; 95% CI, 1.7-2.9). CONCLUSIONS: Young children, elderly persons, and black persons of all ages are disproportionately affected by invasive pneumococcal disease. Current ACIP recommendations do not address a subset of persons aged 18 to 64 years but do include those at highest risk for death from invasive pneumococcal disease. PMID- 11277828 TI - Association of cancer with AIDS-related immunosuppression in adults. AB - CONTEXT: Large-scale studies are needed to determine if cancers other than Kaposi sarcoma, non-Hodgkin lymphoma, and cervical cancer occur in excess in persons with human immunodeficiency virus (HIV) infection or acquired immunodeficiency syndrome (AIDS). OBJECTIVES: To examine the general cancer pattern among adults with HIV/AIDS and to distinguish immunosuppression-associated cancers from other cancers that may occur in excess among persons with HIV/AIDS. DESIGN, SETTING, AND SUBJECTS: Analysis of linked population-based AIDS and cancer registry data from 11 geographically diverse areas in the United States, including 302 834 adults aged 15 to 69 years with HIV/AIDS. The period of study varied by registry between 1978 and 1996. MAIN OUTCOME MEASURE: Relative risks (RRs) of cancers, calculated by dividing the number of observed cancer cases by the number expected based on contemporaneous population-based incidence rates. We defined cancers potentially influenced by immunosuppression by 3 criteria: (1) elevated overall RR in the period from 60 months before to 27 months after AIDS; (2) elevated RR in the 4- to 27-month post-AIDS period; and (3) increasing trend in RR from before to after AIDS onset. RESULTS: Expected excesses were observed for the AIDS defining cancers, but non-AIDS-defining cancers also occurred in statistically significant excess (n = 4422; overall RR, 2.7; 95% confidence interval [CI], 2.7 2.8). Of individual cancers, only Hodgkin disease (n = 612; RR, 11.5; 95% CI, 10.6-12.5), particularly of the mixed cellularity (n = 217; RR, 18.3; 95% CI, 15.9-20.9) and lymphocytic depletion (n = 36; RR, 35.3; 95% CI, 24.7-48.8) subtypes; lung cancer (n = 808; RR, 4.5; 95% CI, 4.2-4.8); penile cancer (n = 14; RR, 3.9; 95% CI, 2.1-6.5); soft tissue malignancies (n = 78; RR, 3.3; 95% CI, 2.6 4.1); lip cancer (n = 20; RR, 3.1; 95% CI, 1.9-4.8); and testicular seminoma (n = 115; RR, 2.0; 95% CI, 1.7-2.4) met all 3 criteria for potential association with immunosuppression. CONCLUSION: Although occurring in overall excess, most non AIDS-defining cancers do not appear to be influenced by the advancing immunosuppression associated with HIV disease progression. Some cancers that met our criteria for potential association with immunosuppression may have occurred in excess in persons with HIV/AIDS because of heavy smoking (lung cancer), frequent exposure to human papillomavirus (penile cancer), or inaccurately recorded cases of Kaposi sarcoma (soft tissue malignancies) in these persons. However, Hodgkin disease, notably of the mixed cellularity and lymphocytic depletion subtypes, and possibly lip cancer and testicular seminoma may be genuinely influenced by immunosuppression. PMID- 11277829 TI - Detection of peanut allergens in breast milk of lactating women. AB - CONTEXT: Most individuals who react to peanuts do so on their first known exposure. A potential but unproven route of occult exposure resulting in sensitization to peanut is via breast milk during lactation. OBJECTIVE: To investigate the ability of maternal dietary peanut protein to pass into breast milk during lactation. DESIGN AND SETTING: Clinical investigation conducted at 2 North American hospitals from March 1999 to October 2000. PATIENTS: Twenty-three healthy, lactating women aged 21 to 35 years. INTERVENTION: Each woman consumed 50 g of dry roasted peanuts, after which breast milk samples were collected at hourly intervals. MAIN OUTCOME MEASURES: Presence in breast milk of total peanut protein, analyzed by a sandwich enzyme-linked immunosorbent assay, and 2 major peanut allergens, Ara h 1 and Ara h 2, detected by immunoblot analysis. RESULTS: Peanut protein was detected in 11 of 23 subjects. It was detected in 10 subjects within 2 hours of ingestion and in 1 subject within 6 hours. The median peak peanut protein concentration in breast milk was 200 ng/mL (mean, 222 ng/mL; range, 120-430 ng/mL). Both major peanut allergens Ara h 1 and Ara h 2 were detected. CONCLUSIONS: Peanut protein is secreted into breast milk of lactating women following maternal dietary ingestion. Exposure to peanut protein during breastfeeding is a route of occult exposure that may result in sensitization of at-risk infants. PMID- 11277830 TI - An 8-year-old boy with autism. PMID- 11277831 TI - A 52-year-old suicidal man, 1 year later. PMID- 11277832 TI - Lipid-lowering therapy in acute coronary syndromes. PMID- 11277833 TI - Neuroprotection in acute ischemic stroke. PMID- 11277834 TI - msJAMA. Electronic medical records: saving trees, saving lives. PMID- 11277835 TI - msJAMA. Patient participation in electronic medical records. PMID- 11277836 TI - msJAMA. Electronic medical records: a decade of experience. PMID- 11277837 TI - msJAMA. Privacy protections for cybercharts: an update on the law. PMID- 11277838 TI - msJAMA. Grassroots computing: palmtops in health care. PMID- 11277840 TI - msJAMA. The patient-owned, population-based electronic medical record: a revolutionary resource for clinical medicine. PMID- 11277847 TI - Conducting statewide needs assessments for persons with traumatic brain injury. AB - The Traumatic Brain Injury Act of 1996 (Public Law 104-166) gave new authority to the Health Resources and Services Administration (HRSA) in the United States Department of Health and Human Services (DHHS) to establish a grant program for states to assist in addressing the needs of persons with traumatic brain injury (TBI). The resulting State Demonstration Grant Program has made available two categories of grants: planning and implementation. Planning grants are awarded to assist states in building infrastructure through the development of four core components. One of the core components is a statewide needs and resource assessment encompassing the full spectrum of services, from initial acute treatment through rehabilitation and long-term community supports. In 1999, assessments completed in 11 states were subjected to a comparative analysis to identify trends across states and to extract salient issues for the State Demonstration Grant Program. This article summarizes the context of the HRSA funded needs assessments and contrasts the methods of needs assessment utilized. Over-arching issues are discussed, including exemplary qualitative and quantitative methods, and the diversity of theoretical models employed in designing assessments and interpreting findings. Several limitations in approaches were also identified, including shortcomings of convenience samples for conducting mail surveys and the unlikely validity of using a needs and resource discrepancy approach to identify gaps in services. PMID- 11277848 TI - A survey of state programs to finance rehabilitation and community services for individuals with brain injury. AB - This article will review and compare existing state-funded programs and their approaches to financing, administering, and delivering an array of services, including rehabilitation and home and community-based services and supports to individuals with traumatic brain injury (TBI) and their families. A state-by state chart provides funding and operational characteristics of the programs. The state menu of services provided and how they are delivered are also described. The article addresses how these programs fit within their overall state service delivery system, trends and changes that programs may be undergoing, and the influences that shape these programs. PMID- 11277849 TI - Coordinated and adequately funded state streams for rehabilitation of newly injured persons with tbi. AB - Traumatic brain injury (TBI) rates are highest among families with the lowest income levels. A paucity of appropriate funding streams for low-income, recently injured TBI patients is reported to cause delays in provision of early postacute rehabilitation or to cause patients to be discharged without receiving rehabilitation. There are also reports of patients remaining in hospitals with minimal care or being returned home, both because of a lack of a discharge site. The purposes of this exploratory study were to: (1) identify model aspects of existing publicly supported and administered programs for postacute individuals with TBI; (2) present results of a survey measuring to what extent state Medicaid programs fund postacute rehabilitation services for recently injured patients with TBI; (3) present results of interviews with trauma center social workers affiliated with the National Institute on Disability and Rehabilitation Research (NIDRR) TBI Model Systems projects to determine whether and how delays in receiving Medicaid coverage occur; and (4) make recommendations for improved systems of care for postacute individuals with TBI. PMID- 11277850 TI - The use of Medicaid waivers and their impact on services. AB - This article examines Medicaid waivers to provide services to adults with traumatic brain injury (TBI) who are Medicaid eligible in the community and who would otherwise be placed in nursing homes or who meet nursing home levels of care. Medicaid waivers for individuals with TBI were addressed through a review of six TBI Waivers (Colorado, Idaho, Kansas, New York, Rhode Island, and Vermont). A summary of the services covered by the waivers is provided. Review of the data suggests two processes, obtaining a waiver and implementing a waiver. Four characteristics were linked to obtaining waivers (a) social-medical political climate, (b) similarity to other waivers, (c) ability to strengthen access and reduce barriers, and (d) expenditure of resources. Similarly, four theoretical characteristics were posited to be associated with managing waivers: (a) cost effectiveness, developmental process of waiver implementation, (c) ability to improve access and reduce barriers, and (d) expenditure of resources to manage the waiver. PMID- 11277851 TI - Community supports for individuals with challenging behavior after brain injury: an analysis of the New York state behavioral resource project. AB - OBJECTIVE: To articulate a framework for supporting individuals with behavioral challenges following brain injury, to describe a state-funded clinical intervention program established to provide community support to individuals with challenging behaviors following brain injury using such a framework, to present data from which it can be concluded that such a program is cost-effective, and to offer policy recommendations. DESIGN: Retrospective analysis of cost data and living arrangements for individuals with behavioral challenges served for a period of 3-4 years through the New York State Home and Community-Based Services Traumatic Brain Injury (HCBS-TBI) Waiver program and the State Behavioral Resource Project. SUBJECTS: 80 individuals with TBI and severe behavioral challenges that included criminal behavior, physical threats, assault and physical aggression, sexually inappropriate behavior, substance abuse, refusal to participate in intervention programs, and psychiatric disturbances, all of whom were at least 2 years postinjury (X = 7.33 years postinjury). RESULTS: The majority of individuals (82% of the 1996 cohort; 89% of the 1997 cohort) with challenging behaviors were living successfully in their home communities 3-4 years after the initiation of behavioral supports. In addition, the costs of serving these individuals decreased in the first year of community integration and for that last year that a cost analysis was completed. CONCLUSIONS: Many individuals with behavioral impairments can be supported successfully in community settings, so long as those supports are flexibly implemented to meet the changing needs of those individuals. Moreover, even with intensive supports, it is cost-effective to serve this population in the community. PMID- 11277852 TI - Educating students with TBI: themes and recommendations. AB - Ten educational consultants and researchers, each with extensive experience working with children with traumatic brain injury (TBI) in school settings, identified seven themes related to serving this population in public schools. These themes are discussed under the headings (1) incidence of TBI and prevalence of persistent educational disability, (2) diversity and central tendencies within the population, (3) assessment, (4) intervention and support in school settings, (5) training and support for educators, (6) intervention and support for families, and (7) systems change and flexibility. For each theme, a set of recommendations is provided, forming an educational research and policy agenda for pediatric TBI. PMID- 11277853 TI - The changing face of publicly funded employment services. AB - The federal-state vocational rehabilitation (VR) program has been a significant stream of funding for individuals with traumatic brain injury (TBI). However, the success of this traditional rehabilitation approach with people with brain injuries remains limited. Recently, the use of specialized employment services, such as Supported Employment, has proven to be far more effective, but funding for this population remains limited. In addition, even individuals who receive specialized employment services often require more intensive, ongoing supports to remain employed. In response, many states have developed innovative services and approaches that are improving the placement and retention rates of individuals with TBI. In this article, the authors provide an overview of selected federal funding initiatives related to the provision of employment services to people with brain injuries, including recent amendments to the Rehabilitation Act. The article also highlights progressive state employment policies and makes suggestions for creative use of government employment monies. PMID- 11277855 TI - Possible applications for dopaminergic agents following traumatic brain injury: part 2. PMID- 11277856 TI - Effect of membrane composition and of co-encapsulation of immunostimulants in a liposome-entrapped crotoxin. AB - Crotoxin isolated from the venom of Crotalus durissus terrificus (South American rattlesnake) was incorporated into liposomes by the dehydration-rehydration vesicle method using different membrane compositions and the co-encapsulation of immunostimulants. Crotoxin was also encapsulated into liposomes formed from a non phospholipid amphiphile, a mixture of polyoxyethylene 2-cetyl ether, dicetyl phosphate and cholesterol. The preparations were characterized in relation to stability, toxicity and the protection of mice against whole venom after immunization. All liposome preparations were quite stable, retaining more than 75% of the originally encapsulated crotoxin after 1 week of incubation at physiological temperature. Co-encapsulation with lipopolysaccharide increased the leakage of crotoxin. In contrast, co-encapsulation of the lipid moiety of lipopolysaccharide did not influence the stability of liposomes. Toxicity of liposomes was dependent on membrane composition. Liposomes made with phospholipids that were resistant to phospholipase A(2) activity were less toxic. Mice immunized with three doses of the 1 x LD50 of crotoxin encapsulated into liposomes, and with associated immunostimulants, were protected against challenge with 8 x subcutaneous LD50 of C. durissus terrificus venom. Using the same immunization schedule, liposomes made from a non-phospholipid mixture and without immunostimulants achieved 100% protection. PMID- 11277857 TI - Synthesis and analysis of ethylated tetracycline, an antibiotic derivative that inhibits the growth of tetracycline-resistant XL1-Blue bacteria. AB - Bacterial resistance to antibiotics is a significant problem in medical care facilities, causing increased fatalities due to infection. The present study demonstrates that antibiotic structures can be selectively altered in a manner that revives their ability to inhibit bacterial growth. The antibiotic tetracycline was ethylated at the position of the phenolic hydroxy group with the use of diazoethane, forming an ethyl ether functional group. This derivative was dissolved in Luria-Bertani (LB) agar medium, then placed in tissue culture for screening against a tetracycline-resistant bacterial strain. The growth of this bacterial strain, designated XL1-Blue, was inhibited by the ethylated form of tetracycline. The procedure for synthesizing ethylated tetracycline utilizes diazoethane and is presented with the molecular structures and IR spectra. The ethylated form of tetracycline was stable at -20 degrees C for many weeks, and was soluble in LB agar plate medium. Ethylated tetracycline induced growth inhibition of XL1-Blue bacteria within the first 24 h of incubation. The level of bacterial growth inhibition was greater than 30%. Calculation of the partition coefficient, log P, was accomplished and indicates that ethylated tetracycline has an increased lipophilic tendency relative to unmodified tetracycline, and therefore has greater solubility in lipid bilayers. PMID- 11277859 TI - Using an experimental design for the optimization of LVV-haemorphin-7 and VV haemorphin-7 extraction by an organic solvent mixture in the course of bovine haemoglobin peptic hydrolysis. AB - Mixture design was used to improve the extraction of two opioid peptides (LVV haemorphin-7 and VV-haemorphin-7) by water-immiscible solvents in the course of bovine haemoglobin peptic hydrolysis. Because of the loss of the peptic activity, these two haemorphins did not appear in the aqueous phase when the peptic reaction was achieved in the presence of butan-2-ol alone. We have shown that it is possible to use octan-1-ol, as a co-solvent, to recover the peptic activity. However, when the hydrolysis was achieved with octan-1-ol alone, the two haemorphins appeared in the aqueous phase, but were not extracted by the organic phase. We therefore investigated haemorphin extraction in the course of the hydrolysis reaction by a mixture of butan-2-ol and octan-1-ol. We have determined the optimal conditions with respect to the extraction of opioid peptides and the stability of the pepsin. To design a future continuous-stirred-tank reactor, we propose a biphasic system composed of 45% water, 45% butan-2-ol and 10% octan-1 ol for the extraction of the two haemorphins in the course of bovine haemoglobin peptic hydrolysis. PMID- 11277858 TI - Anthrax-toxin-mediated delivery of a 19 kDa antigen of Mycobacterium tuberculosis into the cytosol of mammalian cells. AB - PA63, the proteolytically activated 63 kDa fragment of protective antigen (PA, 83 kDa), mediates translocation of lethal factor (LF) and oedema factor into the cytosol. The N-terminal 254 amino acids of LF (LFn) are required for binding to PA63 and mediating translocation of active ligands fused to either the N- or C terminus. Here we report translocation of a 19 kDa antigen of Mycobacterium tuberculosis into the cytosol of mammalian cells when fused to the C-terminus of LFn (LFn-19kDa). The fusion protein was non-toxic to J774A.1 macrophage cells in combination with PA and retained the ability to bind to PA63 when incubated with Chinese hamster ovary K1 cells. The data show the efficacy of anthrax toxin to mediate translocation of M. tuberculosis antigens into the cytosol of mammalian cells and may prove useful in delivering proteins and peptides carrying immunodominant mycobacterial antigens into the cytosol. PMID- 11277860 TI - Erythrocyte-mediated delivery of dexamethasone in patients with chronic obstructive pulmonary disease. AB - Human erythrocytes from ten patients with chronic obstructive pulmonary disease (COPD) were loaded with increasing amounts of dexamethasone 21-phosphate and were re-infused into the original donors. Drug-loaded erythrocytes acted as circulating bioreactors, converting the non-diffusible dexamethasone 21-phosphate into the diffusible dexamethasone. Pharmacokinetic analyses on these patients showed that a single administration of drug-loaded erythrocytes was able to maintain detectable dexamethasone concentrations in blood for up to seven days. This continuous release of dexamethasone was paralleled by the suspension of beta2-agonist and oral corticosteroid treatments by all of the patients. Thus dexamethasone 21-phosphate-loaded erythrocytes are safe carriers for corticosteroid analogues and are a useful alternative to frequent oral or inhaled drugs in elderly patients with COPD. PMID- 11277861 TI - Solution stability studies of the subunit components of meningococcal C oligosaccharide-CRM197 conjugate vaccines. AB - Spectroscopic methods were used to detect modifications in the structures of CRM197, the mutant diphtheria toxin, and meningococcal C capsular oligosaccharide following their conjugation and incubation at various temperatures. Meningococcal C oligosaccharide-CRM197 conjugate vaccines obtained from two different manufacturers were incubated at -20, 4, 23, 37 or 55 degrees C for 5 weeks or subjected to ten cycles of freeze-thawing. The CRM197 carrier protein and the saccharide components of the treated vaccines were monitored by CD and NMR spectroscopic techniques. CD data indicated incubation temperature-dependent conformational changes in the carrier protein from vaccine A. Modifications appeared in both secondary and tertiary structures of the conjugated CRM(197) when incubated at 23 degrees C or above. This was characteristic of the 'open' conformation previously observed for this protein component. The NMR spectra also indicated modification of the structure of the conjugated CRM197 component of vaccine A when incubated at 23 degrees C or above, but failed to show any modification in the conjugated oligosaccharide. On the other hand, the structure of the oligosaccharide chains in vaccine B appeared to be degraded following incubation at 55 degrees C, even though the thermal effect on the conjugated CRM197 was less apparent. Repeated freeze-thawing did not affect the CD or NMR spectra. In conclusion, the two meningococcal C oligosaccharide-CRM197 conjugate vaccines were stable when stored at their recommended temperatures, but were differently affected by elevated temperatures. The conjugates differ in their conjugation chemistry, attachment positions, oligosaccharide chain length and loading, as well as recommended pH and storage buffer, and their different stability properties can probably be attributed to a combination of these factors. PMID- 11277863 TI - Biochemical and immunological properties of human cardiac troponin I fragments. AB - Cardiac troponin I (cTnI) is the inhibitory subunit of the troponin complex and is a biochemical marker for myocardial infarction (MI). It is found in human serum within 4-6 h following MI. One of us has shown [Morjana (1998) Biotechnol. Appl. Biochem. 28, 105-111] that MI patient serum TnI is cleaved at the N- and C terminals and that the TnI fragments exist as a complex with tropinin C (TnC) and troponin T (TnT). In the present study, we have generated C-terminal truncated TnI fragments and studied their immunological and biochemical properties. Human recombinant TnI (rTnI) expressed in Escherichia coli is cleaved into a major fragment with a molecular mass of 17500 Da using CNBr. The major CNBr fragment contains the first 153 amino acids of human cTnI (TnI153). Cleavage of the rTnI with the endoproteinase Asp-N generates a smaller TnI fragment (TnI88, residues 6 96). TnI153 has higher immunological activity than that of rTnI and lower activity than that of TnI88, as judged by the Stratus II TnI Immunoassay. TnI153 exhibits biochemical and immunological properties similar to those of intact TnI. It binds TnC at a molar ratio of 1:1 and forms a ternary complex with TnC and TnT. TnC enhances the immunological activity of TnI153, but has little effect on the activity of TnI88. The TnI153-TnC complex exhibits higher immunological activity than rTnI-TnC and TnI88-TnC, and much higher activity than free rTnI, TnI153 and TnI88. The presence of TnT has no effect on the immunological activity of the TnI153-TnC complex, suggesting that the addition of TnT does not interfere with TnI153 recognition by TnI monoclonal antibodies. Free TnI153 and TnI88 degrade rapidly in human serum. TnC protects TnI153 from proteolytic degradation, but offers less protection for TnI88. The TnI88-TnC complex lost 80% of its immunological activity after incubation for 2 days in human serum at 37 degrees C. However, there was no loss in the immunological activity of the TnI153-TnC complex under the same conditions. A cTnI fragment (TnI80, residues 1-80), expressed in E. coli as a fusion protein, exhibits immunological activity and stability similar to that of TnI88. PMID- 11277862 TI - Chromosome localization and gene-copy-number quantification of three random integrations in Chinese-hamster ovary cells and their amplified cell lines using fluorescence in situ hybridization. AB - We have used fluorescence in situ hybridization (FISH) to localize three random Chinese-hamster ovary (CHO) cell chromosomal integration sites and determine gene copy number in their amplified cell lines. Metaphase FISH showed all three to have integrated into different chromosome positions on different chromosomes. All three Geneticin parent cell lines were found to have single integration sites by Southern-blot analysis, and these data were confirmed using both metaphase and interphase FISH. Following amplification, metaphase FISH showed that amplification in two of the cell lines occurred by duplication in the chromosome arm where the integration occurred. However, for one cell line, amplification resulted in the translocation of amplified genes from one marker chromosome to another. Interphase FISH showed an expected increase in gene copy number upon amplification with methotrexate. PMID- 11277864 TI - Improved preparation and crystallization of 25 kDa human fibroblast growth factor 9. AB - We prepared 25 kDa human fibroblast growth factor-9 (hFGF-9 N33) on a large scale after overproduction in Escherichia coli MM294 (DE3)/pTG931. The purification was performed by a combination of hydrophobic chromatography and HPLC with an ion exchange column, a heparin affinity column and a gel filtration column. This improved procedure was rapid and simple, and the purified hFGF-9 N33 was found to be homogeneous as judged by various criteria, such as amino acid analysis, N terminal amino acid sequence, C-terminal amino acid analysis and biological activity. Furthermore, as determined by low endotoxin and DNA content, the protein was of high purity. In addition, the hFGF-9 N33 prepared in the present study was easily crystallized by the vapour diffusion method. PMID- 11277865 TI - Activation of diacetyldapsone and a preliminary evaluation of a cyclodextrin diacetyldapsone complex in cultured lung cells. AB - A diacetyldapsone-2-hydroxypropyl-beta-cyclodextrin complex (DADDS-CD) was evaluated with regard to the ability of cultured lung cells to activate DADDS to the active metabolite dapsone. The same system was used to assess the effect of the complex on cell growth. The complex was incubated with cells for periods of 24 to 72 h, followed by extraction of metabolites from the incubation medium and analysis by HPLC. In addition, the Trypan Blue exclusion technique was used to assess cell viability during this time period. Results indicated that lung cells could activate DADDS to dapsone and that, while the complex appeared to delay cell growth in the first 24 h period, no significant difference was seen between cells incubated in the presence and absence of the complex at 72 h. These results indicate that DADDS-CD has significant potential as a drug-delivery system for DADDS in the lung based upon the ability of the cells to activate DADDS. The mixed effects of the complex on cell growth may have important implications when considering the frequency of administration of the complex to the lung. PMID- 11277866 TI - A rapid method for detecting conformational changes during differentiation and apoptosis of HL60 cells by Fourier-transform infrared spectroscopy. AB - The conformational changes of HL60 cells during differentiation and apoptosis are still not clearly understood. This study uses a new method, Fourier-transform infrared (FT-IR) spectroscopy, to analyse the changes of HL60 cells. We detected several differences among normal HL60 cells, differentiated cells induced by all trans retinoic acid (ATRA) and PMA, and apoptotic cells induced by vincristine (VCR). The results suggest that the alpha-helix content of the membrane protein of differentiated HL60 cells induced by ATRA and PMA increased,and that the beta sheet content of membrane proteins of apoptotic cells induced by VCR also increased. Also, C-O(H)-stretching modes of serine, threonine and tyrosine residues of cell proteins were shown to be distinctly different, but no significant differences were detected between cells differentiated along different lineages by ATRA and PMA. PMID- 11277867 TI - Regulated secretion of proinsulin/insulin from human hepatoma cells transduced by recombinant adeno-associated virus. AB - To employ hepatocytes as surrogate beta-cells for gene therapy of diabetes, a regulatory system was devised in this study by placing the human insulin cDNA under the control of the phosphoenolpyruvate carboxykinase (PEPCK) promoter, followed by the cytomegalovirus immediate early promoter-driven enhanced-green fluorescent-protein open reading frame. The expression cassette was inserted into the adeno-associated virus vector between two inverted terminal repeats, and used to produce recombinant adeno-associated virus (rAAV). HepG2 human hepatoma cells were transduced by rAAV at the desired multiplicity of infection, followed by treatment with various concentrations of retinoic acid, dexamethasone, dibutyryl cAMP (dbcAMP) and 3-isobutyl-1-methylxanthine (IBMX). The cell-culture media were collected at 8, 16 and 24 h later. Proinsulin/insulin levels were determined with human proinsulin/insulin radioimmunoassay kits. Transduction of HepG2 cells by rAAV showed that green fluorescence was produced as early as 12 h after rAAV infection. Flow-cytometrical analysis demonstrated that transduction efficiency increased with the numbers of transducing rAAV particles used. The transduced hepatocytes were shown to secrete immunoreactive proinsulin/insulin, which were stimulated by the concentrations of retinoic acid, dexamethasone and dbcAMP in the culture medium. High conversion from proinsulin into insulin occurred when these cells were treated with dexamethasone and dbcAMP. The presence of IBMX enhanced the secretion of proinsulin/insulin from the dbcAMP-treated cells. We conclude that rAAV is a promising vector for gene therapy of diabetes. Regulated secretion of proinsulin/insulin can be obtained in the rAAV-transduced HepG2 cells conferred with the PEPCK promoter via rAAV-mediated gene transfer. PMID- 11277868 TI - Corticotrophin-releasing hormone and human parturition. AB - Corticotrophin-releasing hormone (CRH), the hypothalamic peptide that controls function of the pituitary-adrenal axis in response to stress, is expressed in abundance in the human placenta and is present in high concentrations in maternal and fetal plasma during late pregnancy. A number of lines of evidence now imply a role for this hormone in the control of parturition and fetal maturation in humans. It has been proposed that CRH, through interactions with oestrogen, adrenal steroids, prostaglandins and oxytocin, establishes positive-feedback loops that initiate parturition and labour. Excessive production of placental CRH has also been linked to preterm labour. However, there are a number of significant gaps in our knowledge of the function of this peptide in pregnancy. This review examines current evidence regarding the putative role of CRH in human parturition. PMID- 11277869 TI - Potential local regulatory functions of inhibins, activins and follistatin in the ovary. AB - The changing pattern of granulosa cell expression of inhibin/activin subunits and follistatin during follicle development and their differential regulation by extrinsic and intraovarian factors supports evidence from functional studies, mostly in vitro, that these proteins have important roles in folliculogenesis, oocyte maturation and corpus luteum function. Gonadal inhibins function as negative feedback hormones to regulate the synthesis and secretion of pituitary FSH, a key determinant of follicle development, but there is little supportive evidence for a peripheral endocrine role for ovary-derived activins or follistatin in this regard. However, activins and follistatin are expressed in numerous other tissues, including anterior pituitary, and they are firmly implicated as local intrapituitary regulators of FSH secretion. Intraovarian actions of granulosa cell-derived activins include the promotion of granulosa cell proliferation and upregulation of FSH receptors, P450arom, oestrogen synthesis, granulosa cell LH receptors and enhancement of oocyte maturation. Through its activin-binding role, follistatin can reverse each of these activin induced responses. In addition to their endocrine feedback role, granulosa derived inhibins can sensitize theca cells to LH, thereby enhancing the production of androgens, an essential requirement for follicular oestrogen synthesis. Activins can oppose this effect and suppress thecal androgen production. Granulosa cells overproduce inhibin a subunit precursor relative to betaA/betaB subunit precursors and evidence indicates that different parts of the inhibin a subunit precursor have intrinsic biological activities distinct from inhibin alphabetaA/B dimer, and serve as additional local modulators of follicle and corpus luteum function. PMID- 11277870 TI - Fetomaternal interactions and influences during equine pregnancy. AB - The equine embryo takes 6 days to traverse the oviduct and, when it finally enters the uterus, it remains spherical in shape and moves continually throughout the uterine lumen until day 17 after ovulation to deliver its maternal recognition of pregnancy signal to the entire endometrium. Between day 25 and day 35 after ovulation, the trophoblast cells of a discrete annulate portion of the chorion multiply rapidly and acquire an invasive phenotype and, between day 36 and day 38, migrate deeply into the maternal endometrium to form the equine unique endometrial protuberances known as endometrial cups. These cups secrete large quantities of a gonadotrophic hormone (eCG) into the maternal circulation which, in conjunction with pituitary FSH, stimulates the development of accessory luteal structures in the maternal ovaries to supplement the supply of progesterone to maintain the pregnancy until the placenta can assume this role at about day 100. The non-invasive allantochorion extends slowly to fill the uterus by days 80-85 and its microcotyledonary architecture, which provides both haemotrophic and histotrophic nutrition for the growing fetus, is not fully established until days 120-140. The fetoplacental unit synthesizes large quantities of steroid hormones during the second half of pregnancy, using fetal C 19 precursors secreted by the enlarged fetal gonads for the production of oestrogens and maternal C-21 precursors for the synthesis of progesterone and large quantities of 5alpha-reduced progestagens. Near term, additional pregnenelone is secreted by the fetal adrenal glands so that the mare exhibits the unusual phenomenon of foaling while maternal serum progestagen concentrations are increasing and oestrogen concentrations are decreasing. PMID- 11277871 TI - Chemosensation and genetic individuality. AB - Numerous studies have shown that there are measurable behavioural consequences that can result from the olfactory recognition of alleles borne at the major histocompatibility complex (MHC). These consequences include simple individual recognition, disassortative mate preference, discrimination of kin from non-kin and whether a pregnancy is carried to term. Such a system, which can influence the reproductive behaviour of a species, will have profound effects on its genetic constitution and survival. The likely mechanism responsible for the production of MHC-related odours involves soluble MHC molecules that carry allele specific odoriferous molecules from the blood via the kidneys into the urine, from where they are released into the environment. The ability of soluble MHC molecules to signal genetic individuality in this way may have evolved before the appearance of an acquired immune system in our immediate ancestors, the protochordates. PMID- 11277872 TI - An X-Y paint set and sperm FISH protocol that can be used for validation of cattle sperm separation procedures. AB - X and Y chromosome paints were developed from sorted yak chromosomes for sexing cattle spermatozoa. Clear hybridization signals were obtained for every spermatozoon using a modified sperm decondensation protocol and fluorescence in situ hybridization (FISH). The procedure was evaluated using the established Beltsville sperm sexing technology, which separates spermatozoa by flow cytometry into X- and Y-bearing fractions. Close agreement was found between the assessment of sperm separation by flow cytometry and by FISH with the X-Y paint set. The FISH method is a simple, reliable and robust procedure for assessing the effectiveness of separation of X and Y spermatozoa. PMID- 11277873 TI - Localization of primordial germ cells or their precursors in stage X blastoderm of chickens and their ability to differentiate into functional gametes in opposite-sex recipient gonads. AB - This study was performed to determine the distribution of primordial germ cells and their precursors in stage X blastoderm of chickens. The blastoderm (Barred Plymouth Rock chickens) isolated from the yolk was separated into three portions: the central disc, the marginal zone and the area opaca. The dissociated blastodermal cells derived from the central disc, marginal zone and area opaca were transferred into a recipient blastoderm (White Leghorn chicken) from which a cell cluster was removed from the centre of the central disc. The manipulated embryos were cultured in host eggshells until hatching. The chicks were raised until sexual maturity and test mated with Barred Plymouth Rock chickens to assess the donor cell contribution to the recipient germline. Germline chimaeric chickens were produced efficiently (46.7%, 7/15) when the blastodermal cells derived from the central disc were transferred into recipient embryos of the same sex, whereas no germline chimaeric chickens were produced when the blastodermal cells derived from the marginal zone or area opaca were transferred into recipient embryos of the same sex (0/12). Germline chimaeric chickens were also produced by transfer of blastodermal cells derived from the central disc (6.7%, 1/15), marginal zone (10.0%, 1/10) or area opaca (11.1%, 1/9) into recipient embryos of the opposite sex. It is concluded that primordial germ cells are induced during or shortly after stage X and that the cells derived from the central disc have the highest potential to give rise to germ cells. Cells derived from the marginal zone and area opaca can also give rise to germ cells, although the frequency is low. PMID- 11277874 TI - Secretion of matrix metalloproteinases 2 and 9 and tissue inhibitor of metalloproteinases into follicular fluid during follicle development in equine ovaries. AB - Extensive tissue remodelling is required in equine ovaries for follicle growth and development and also migration of the follicle to the ovulatory fossa, where ovulation occurs. The mechanisms for these processes are largely unexplored. Matrix metalloproteinases (MMPs) and their endogenous tissue inhibitors (TIMPs) are important for control of breakdown of extracellular matrix during tissue remodelling. The aims of this study were to determine the pattern and sites of secretion of the gelatinases MMP-2 and -9 and TIMPs into follicular fluid during follicle development in mare ovaries. The predominant gelatinase detected in follicular fluid was MMP-2, which was present in similar amounts throughout follicular development, as demonstrated by zymography. MMP-9 was also present in follicular fluid and secretion increased significantly (P < 0.05) with development of follicles from < 10 mm to 11-20 mm in diameter. Follicular fluid also contained TIMP-1, TIMP-2, unglycosylated and glycosylated TIMP-3, and TIMP 4, as shown by reverse zymography. The abundance of TIMPs remained largely unchanged during follicle development. MMP-2 and -9 were localized by immunohistochemistry to stromal cells and granulosa and theca cells. TIMP-1, -2, 3 and -4 were present in granulosa and theca cells of the follicle and in stromal cells and also associated with extracellular matrix of the ovarian stromal tissue. The MMPs and TIMPs are likely to be involved in the regulation of the breakdown of extracellular matrix during tissue remodelling for follicle development and migration to the ovulation fossa in mares. PMID- 11277876 TI - Inhibition of neutrophil chemotaxis by pig seminal plasma in vitro: a potential method for modulating post-breeding inflammation in sows. AB - The aim of this study was to determine the regulatory role of pig seminal plasma in post-breeding uterine inflammation. Polymorphonuclear neutrophil (PMN) chemotaxis of lipopolysaccharide (LPS)-activated blood plasma or heat-inactivated blood plasma plus LPS containing increasing concentrations of seminal plasma was assessed in chemotactic chambers. Seminal plasma was diluted serially with McCoy's medium to concentrations of 50.0, 25.0, 12.5, 6.2 or 3.1% (v/v) and added to normal or heat-inactivated pig blood plasma that was activated with LPS before or after incubation in a 37 degrees C waterbath for 30 min. Chemotaxis was determined using blood-derived PMNs and was expressed as a percentage of the positive control of LPS-activated blood plasma. A linear dose-dependent suppression of chemotaxis by seminal plasma was observed for blood plasma activated before or after addition of seminal plasma. Compared with the positive control, concentrations of seminal plasma < 6.2% failed to suppress PMN chemotaxis (P < 0.05). A dose-dependent suppressive effect of seminal plasma on heat stable chemotactic components of pig blood plasma was also observed (P < 0.05). A marked suppression was observed at concentrations of seminal plasma > 12.5% of the sample volume (P < 0.05). These results indicate that seminal plasma suppresses chemotactic blood plasma components regardless of formation sequence (pre- or post-activation) or source (normal or heat-inactivated blood plasma). These results indicate that seminal plasma may be necessary in diluted boar semen used for artificial insemination to regulate post-breeding inflammation in sows. PMID- 11277875 TI - Relationship of Fas ligand expression and atresia during bovine follicle development. AB - The Fas antigen (Fas) is a cell surface receptor that may be involved in the initiation and progression of follicle cell apoptosis during atresia. Fas initiates apoptosis in sensitive cells after binding Fas ligand (FasL). Other experiments have shown that expression of Fas mRNA and responsiveness to Fas mediated apoptosis vary in bovine granulosa and theca cells during follicle development. In the present study, FasL mRNA content was measured and Fas and FasL protein expression was examined in bovine granulosa and theca cells of healthy dominant follicles and the two largest atretic subordinate follicles on day 5 of the oestrous cycle (day 0 = oestrus), and of dominant follicles from the first wave of follicle development after they had become atretic and showed no growth for 4 days. FasL mRNA content was higher in granulosa cells from atretic compared with healthy follicles. FasL mRNA content was also higher in theca cells from atretic subordinate compared with healthy dominant follicles on day 5, but did not differ between theca cells from healthy and atretic dominant follicles. Immunohistochemical staining for FasL was more intense in theca compared with granulosa cells and in atretic compared with healthy follicles. Immunohistochemical staining for Fas was more intense in granulosa compared with theca cells and in atretic subordinate compared with healthy dominant follicles on day 5. Immune cells, known to express Fas and FasL, were localized in the theca, but not the granulosa, cell layer of all follicles. Higher concentrations of Fas and FasL in cells from atretic follicles, together with the previous demonstration of increased responsiveness of granulosa cells from subordinate follicles to FasL-induced apoptosis, support a potential role for FasL-mediated apoptosis during ovarian follicle atresia. PMID- 11277877 TI - Expression of heparanase mRNA in bovine placenta during gestation. AB - This study reports the identification and sequence of a partial cDNA for bovine heparanase and the expression of its mRNA in the placenta during gestation. The 364 amino acid residues deduced from the 1092 bp cDNA fragment share 81.9% and 80.5% identity with amino acid sequences of human and rat heparanase, respectively. Northern blot hybridization showed that two mRNAs (2.0 and 3.5 kb) are strongly expressed in placenta, and weakly expressed in the kidney, lung, spleen and non-pregnant uterus. In the placenta, these transcripts were detected in the cotyledon at all stages of gestation examined, and in the intercotyledonary fetal membrane and caruncle on day 60, day 120 and day 260. Quantitative real-time RT-PCR analysis showed very low expression of heparanase mRNA in the conceptus before implantation (day 17), but high expression in the cotyledon-containing fetal membrane (days 27-34) after implantation. Furthermore, heparanase mRNA was detected in the cotyledon, intercotyledonary fetal membrane and caruncle after days 60-64 of gestation. However, no significant expression of heparanase mRNA was observed in intercaruncular endometrium at all stages of gestation examined. These results demonstrate that heparanase mRNA is expressed in the placentome, indicating that heparanase may play a role in implantation, and in placental development and function. PMID- 11277878 TI - Association of endometriosis in horses with differentiation of periglandular myofibroblasts and changes of extracellular matrix proteins. AB - Periglandular fibrosis and cystic dilation of uterine glands are associated with equine endometriosis. The presence of extracellular matrix proteins (collagen type I, III and IV, laminin and fibronectin) in healthy and endometriotic specimens was demonstrated by immunohistochemistry. The distribution of collagen I, but not collagen III, was dependent on the stage of the oestrous cycle. The arrangement of collagen I and collagen III in endometriotic specimens was similar to that in normal endometrium. In periglandular fibrosis, collagen IV, laminin and fibronectin deposition outside the basement membrane was observed. In these regions, stromal cells were characterized immunohistochemically as myofibroblasts because of their expression of a-smooth muscle actin, and occasionally tropomyosin and desmin. Periglandular differentiation of contractile cells could be interpreted as a reaction to support the extrusion of secretions in cystic dilated glands. Moreover, the changes of extracellular matrix proteins are characteristic for neoplastic lesions, although further development of endometriosis to benign or malignant tumours is not known in horses. Knowledge of the factors responsible for these fibroblastic modulations may be the key to explaining the pathogenesis of endometriosis. PMID- 11277879 TI - Immunolocalization of steroidogenic enzymes in the corpus luteum and placenta of the Japanese black bear, Ursus thibetanus japonicus, during pregnancy. AB - The Japanese black bear, Ursus thibetanus japonicus, is a seasonal breeder and shows delayed implantation for several months during pregnancy. The objective of this study was to clarify the steroidogenic capability of the corpus luteum and placenta during pregnancy, including both delayed implantation and fetal development, by immunolocalization of steroidogenic enzymes in these organs of the Japanese black bear. Ovaries and placentae from 15 wild Japanese black bears, which had been killed legally by hunters and were thought to be pregnant, were used in an immunocytochemical study to localize the cholesterol side chain cleavage cytochrome P450 (P450scc), 3beta-hydroxysteroid dehydrogenase (3betaHSD), 17alpha-hydroxylase cytochrome P450 (P450c17) and aromatase cytochrome P450 (P450arom) by the avidin-biotin-peroxidase complex method using polyclonal antisera raised in mammals against P450scc, 3betaHSD, P450c17 and P450arom. P450scc and 3betaHSD were localized in all luteal cells throughout pregnancy. P450c17 was present in a few luteal cells, especially in the outer area of the corpus luteum throughout pregnancy, but the number of positively immunostained cells decreased during the post-implantation period. Cells positively immunostained for P450c17 were significantly smaller than negatively immunostained cells (P < 0.01). P450arom was present sporadically in a few luteal cells throughout pregnancy, but the number of positively immunostained cells decreased during the post-implantation period. The size of cells positively immunostained for P450arom was not significantly different from that of negatively immunostained cells. The whole placenta was negatively immunostained for P450scc, 3betaHSD and P450c17, but P450arom was present in the syncytiotrophoblasts and endothelial cells of maternal blood vessels. These results indicate that, in the Japanese black bear, corpora lutea are a source of progesterone which may play an important role in the maintenance of delayed implantation and fetal development during pregnancy. Corpora lutea have a minimum capability to synthesize androgen in small luteal cells and oestrogen in normal sized luteal cells during pregnancy, and placentae have the ability to synthesize oestrogen during late pregnancy. PMID- 11277881 TI - Effects of arginine- and lysine-vasopressin on phospholipase C activity, intracellular calcium concentration and prostaglandin F2alpha secretion in pig endometrial cells. AB - Oxytocin and vasopressin are related peptides that have receptors in the uterus. Species from families other than Suidae produce only arginine-vasopressin; in contrast, pigs apparently express both arginine- and lysine-vasopressin. The aim of this study was to determine whether arginine- or lysine-vasopressin would activate phospholipase C, increase intracellular calcium concentration [Ca(2+)](i) and stimulate PGF(2alpha) production in enriched cultures of stromal, glandular epithelial and luminal epithelial cells from pig endometrium. Cells were obtained from gilts on day 16 after oestrus by differential enzymatic digestion and sieve separation. After 96 h in culture, the cells were treated with 0 or 100 nmol arginine- or lysine-vasopressin l(-1). The responses to 100 nmol oxytocin l(-1) and 100 nmol GnRH l(-1) were used as positive and negative controls, respectively. Consistent with previous results, oxytocin stimulated phospholipase C activity (P < 0.05), increased [Ca(2+)](i) (P < 0.05) and promoted PGF(2alpha) secretion (P < 0.05) from stromal and glandular epithelial cells. Activity of phospholipase C, [Ca(2+)](i) and PGF(2alpha) release were also increased (P < 0.05) by arginine-vasopressin in stromal cells, but the responses were less (P < 0.01) than those induced by oxytocin. An oxytocin antagonist attenuated the [Ca(2+)](i) response of stromal cells to both oxytocin and arginine-vasopressin. Sequential treatment of cells with oxytocin and arginine vasopressin indicated that oxytocin desensitized the response to oxytocin, but arginine-vasopressin did not similarly desensitize the response to oxytocin. In glandular and luminal epithelial cells, arginine-vasopressin did not stimulate phospholipase C activity, [Ca(2+)](i) or PGF(2alpha) secretion. Neither GnRH nor lysine-vasopressin induced phospholipase C activity, increased [Ca(2+)](i) or stimulated PGF(2alpha) production in any endometrial cell type. These results indicate that oxytocin receptors can bind arginine-vasopressin more readily than they bind lysine-vasopressin. Type 1 vasopressin receptors may also exist in endometrium predominantly on cells other than stromal, glandular epithelial and luminal epithelial cells, as in previous studies both arginine-vasopressin and lysine-vasopressin stimulated phospholipase C activity in endometrial explants to a similar extent as oxytocin. PMID- 11277880 TI - Improvement of semen quality by nocturnal scrotal cooling and moderate behavioural change to reduce genital heat stress in men with oligoasthenoteratozoospermia. AB - A questionnaire assessing factors that might cause an increase in scrotal temperature was completed by patients with reproducible oligoasthenoteratozoospermia of idiopathic nature or caused by varicocele. Evaluation by means of a grading scale revealed increased scrotal heat stress in oligoasthenoteratozoospermic patients compared with normozoospermic men (P < 0.01). In addition, long-term determination of 24 h scrotal temperature profiles showed that compared with semen donors, oligoasthenoteratozoospermic patients frequently had scrotal temperatures above 35.5 degrees C despite the same environmental temperatures (P < 0.05). In 88% of cases, maximum scrotal temperatures were measured during rest or sleep phases, whereas minimum values were recorded during physical activity or frequent change of position. Nocturnal scrotal cooling by means of an air stream resulted in a decrease in scrotal temperature of approximately 1 degrees C. Furthermore, a highly significant increase in sperm concentration (P < 0.0001) and total sperm output (P < 0.0001) was achieved after nocturnal scrotal cooling for 12 weeks together with a moderate decrease in factors leading to genital heat stress. A significant improvement in sperm motility (P < 0.05) and sperm morphology (P < 0.05) was also observed, but this improvement was markedly less pronounced than the changes in sperm concentration. This study shows the importance of genital heat stress as a cofactor in fertility impairment in men and indicates nocturnal scrotal cooling as a therapeutic option. PMID- 11277882 TI - Reproductive seasonality in domestic sows kept outdoors without boars. AB - Plasma progesterone, LH and prolactin concentrations were measured twice a week in mature sows kept outdoors without boars in two experiments to examine whether perception of daylength change underlies seasonal infertility in domestic pigs. In Expt 1, melatonin implants inserted on 12 April or 22 May to block perception of the increasing daylength did not affect the oestrous cycle significantly, since only two untreated control sows became seasonally anoestrous. In Expt 2, all control sows became anoestrous for 131 +/- 42.5 days (n = 11). Melatonin implants inserted at the spring equinox (n = 9) prevented seasonal anoestrus (P < 0.001), but timed daily oral melatonin administration was less effective (P < 0.05): 5 of 11 sows became anoestrous for 132 +/- 45.6 days. In both experiments, there were significant low-amplitude seasonal rhythms in mean plasma prolactin and LH concentrations. Prolactin reached maximum concentrations 2-4 weeks before the summer solstice, whereas LH reached a nadir 4-6 weeks after the summer solstice. Neither anoestrus nor melatonin administration altered these patterns consistently. Endogenous plasma melatonin was higher (P < 0.001) during darkness (12.5 ng l(-1)) than during daytime (8.9 ng l(-1)) in untreated sows, but only clearly so during spring and summer. Melatonin implants increased mean daily plasma melatonin to 146 ng l(-1), whereas melatonin fed at 15:00 h increased values to 40-60 ng l(-1) 2-4 h after administration, but daytime concentrations were unchanged. Melatonin administration, despite decreasing seasonal anoestrus, did not prevent the seasonal increase in plasma prolactin and had no significant effect on plasma LH; therefore, its role in regulating seasonal changes in the reproduction of domestic sows remains uncertain. PMID- 11277883 TI - Regulation of lipid peroxidation by nitric oxide and PGF2alpha during luteal regression in rats. AB - Corpus luteum regression is related to an increased generation of reactive oxygen species. Although several studies indicate that PGF(2alpha) is involved in regression of the corpus luteum in mammalian species through an increase in reactive oxygen species, the exact mechanism remains unknown. In the present study, the relationship between nitric oxide and PGF(2alpha) in regulation of lipid peroxidation was studied. Ovarian tissue from pseudopregnant rats at mid- (day 5) or late phase or at the time of regression (day 9 of pseudopregnancy) of corpus luteum development was used. Thiobarbituric acid reactants, used as a lipid peroxidation index, were higher on day 9 of pseudopregnancy than on day 5. In contrast, glutathione content (an antioxidant metabolite) was lower on day 9 than on day 5 of pseudopregnancy. These results indicate that there was an enhanced oxidative status in ovarian tissue during luteolysis. Administration of N(omega)-nitro-L-arginine methyl ester (L-NAME: 600 micromol l(-1)), a competitive nitric oxide synthase (NOS) inhibitor, led to a decrease in basal thiobarbituric acid reactant content in ovarian tissue from rats on day 9 of pseudopregnancy only, indicating that during regression of the corpus luteum, NO could act as intermediary in ovarian lipid peroxidation. Administration of a luteolytic dose (3 microg kg(-1) body weight i.p.) of a synthetic PGF(2alpha) increased thiobarbituric acid reactant content in ovaries from rats on day 9 of pseudopregnancy. As this effect was reversed partially by L-NAME, it is proposed that during regression of corpora lutea, PGF(2alpha) and NO are involved in regulation of lipid peroxidation. As this effect was only reversed partially, it is possible that there is another mechanism involving PGF(2alpha) (but not the NO NOS pathway) in regulation of ovarian lipid peroxidation. Furthermore, the administration of PGF(2alpha) enhanced ovarian NOS activity, whereas cyclooxygenase inhibition (by indomethacin treatment in vivo) reduced it. As western blotting of ovarian homogenates obtained from PGF(2alpha)-injected rats increased inducible NOS (iNOS) content, it is concluded that PGF(2alpha) enhances both activity and synthesis of NO in rat ovarian tissues during luteolysis. Taken together, these results indicate that in ovaries with regressing corpora lutea, both NO and PGF(2alpha) are involved in part in regulation of lipid peroxidation. PMID- 11277884 TI - Effect of transient early hyperthyroidism on onset of puberty in Suffolk ram lambs. AB - The thyroid function and sexual development of eight 6-week-old Suffolk ram lambs were studied. The lambs were divided into either control or treatment groups and housed indoors. From 6 to 12 weeks of age, four lambs in the treatment group received 15 mg kg(-1) body weight per day of 6-propyl 2-thiouracil orally to suppress normal thyroid function. During the same period, thyroxine and tri iodothyronine were injected s.c. at the rate of 8 and 16 microg kg(-1) body weight per day, respectively, to induce a hyperthyroid state. Four control lambs received sham injection and oral excipient. Concentrations of thyroxine, tri iodothyronine, FSH, testosterone and insulin-like growth factor I were determined in blood collected by indwelling jugular catheters once a week, every 20 min from 09:00 to 15:20 h. Scrotal circumference was recorded each week. Semen collection was attempted by electro-ejaculation between weeks 17 and 36. Lambs were castrated at week 36 and testicular histology was examined. During the treatment period only, the concentration of thyroid hormones was higher in treated lambs than in controls (P < 0.05). From week 6 to week 9 only, concentrations of FSH in treated lambs were lower than in controls (P < 0.05). Insulin-like growth factor I concentrations were lower in treated lambs than in controls from week 10 to week 13 (P < 0.05). Frequency of testosterone pulses was higher (P < 0.01) in the treated lambs but concentrations were similar in the control and treated lambs throughout the experiment. Scrotal circumference was greater in treated lambs from week 26 to week 36 (P < 0.05). Treated lambs produced viable spermatozoa earlier than did control lambs. At week 36, sperm concentration in treated lambs was higher than in controls (P < 0.01) but semen volumes were similar (P > 0.1). Diameter of the seminiferous tubules in treated lambs was larger than in controls (P < 0.05). In conclusion, transient neonatal hyperthyroidism decreased FSH and insulin-like growth factor I concentrations temporarily, increased testosterone pulses and sperm production and advanced puberty in Suffolk ram lambs. PMID- 11277885 TI - A skin surgery fable. AB - A fable is presented to inform the reader of the existence of a valuable, but under-utilized educational resource that might be utilized in the teaching of dermatologic surgery. PMID- 11277886 TI - Complications of nail surgery: a review of the literature. AB - BACKGROUND: The realm of nail unit surgery encompasses the dermatologist as well as the hand surgeon. Nail surgery complications may include allergy to anesthetic, infection, hematoma, nail deformity, and persistent pain and swelling. OBJECTIVE: To review the pertinent literature regarding nail unit surgery complications. METHODS: A Medline literature search was performed for relevant publications. RESULTS: Nail unit surgery complications appear to be relatively infrequent. The majority of postoperative nail deformity complications result from nail matrix damage. CONCLUSION: Complications may be reduced to a minimum by preventive measures, such as careful patient selection, sterile technique, and gentle treatment of the nail matrix. PMID- 11277887 TI - Nail biopsy: indications and methods. AB - Nail biopsy is a safe and useful technique for diagnosis and management of many nail conditions. A basic understanding of nail anatomy and biology is a prerequisite for a successful nail biopsy. The patient must be adequately prepared and there needs to be excellent anesthesia and hemostasis. The type of nail biopsy depends largely on the location of the pathology in the nail unit. The techniques of nail biopsy by location in the nail unit and by lesion type are discussed. PMID- 11277888 TI - Warts of the nail unit: surgical and nonsurgical approaches. AB - BACKGROUND: Warts are the most common nail tumor and mostly affect children and young adults. Periungual warts are usually due to HPV-1, 2, and 4. Development of periungual warts is favored by maceration and trauma, especially nail biting. OBJECTIVE: To discuss the biology, clinical features, and medical and surgical treatment of periungual warts. METHODS: Review of the literature and personal experience. RESULTS: The natural course of warts makes aggressive approaches restricted to selected cases. Medical treatments, usually topical, include keratolytic agents, virucidal agents, and immunomodulators. All choices have been utilized successfully, but keratolytic agents are the best first-line approach. Surgical treatments include cryotherapy, surgical excision, electrosurgery, infrared coagulation, localized heating with a radio-frequency heat generator and laser therapy, especially the Er:YAG laser, which has an excellent safety profile. CONCLUSIONS: Definitive cure is not guaranteed by any therapy and periungual warts can recur and become larger after correct treatment. PMID- 11277889 TI - Glomus tumors of the nail unit: a plastic surgeon's approach. PMID- 11277890 TI - Nail unit matrix transplantation: a plastic surgeon's approach. AB - The growth of the nail is primarily from the nail bed and mostly from the germinal matrix component, and to some extent from the sterile matrix as well. The purpose of this article is to describe the approach and technique of a composite nail, nail bed, hyponychium, and perionychium transplant when the total nail and nail bed need reconstruction. A case is presented when a nail bed junctional nevus is excised and reconstruction with nail unit matrix transplantation and the result is shown. PMID- 11277891 TI - Mohs micrographic surgery of the nail unit and squamous cell carcinoma. AB - The nail unit can be a challenging anatomic location for surgical removal of neoplasms. Although uncommon, malignancies do affect this specialized epithelial structure. In particular, Bowen's disease and more invasive squamous cell carcinoma (SCC) are the most common neoplasms to affect the nail unit and surrounding structures. Other neoplasms such as basal cell carcinoma and malignant melanoma can also affect the nail unit, but less frequently, and will not be discussed in the scope of this review. Mohs micrographic surgery continues to be the treatment of choice because of the procedure's tissue-sparing qualities. A clear understanding of the anatomy and the histology of the nail unit, a review of the technique of Mohs surgery of the nail anatomy, as well as a review of the literature are presented. PMID- 11277892 TI - Split nail deformities: a surgical approach. PMID- 11277893 TI - Surgical anatomy of the nail unit. AB - Surgical procedures in the nail unit require intimate knowledge of the gross and microscopic anatomy of the distal digit in order to obtain accurate diagnostic information and achieve the desired results with minimal impact on appearance and function. This review of the surgical anatomy of the nail unit discusses normal anatomy with an emphasis on those aspects necessary to perform surgery with minimal residual impact. PMID- 11277894 TI - Pincer nails: definition and surgical treatment. AB - BACKGROUND: There are four main types of ingrown nail. These are distal nail embedding, juvenile (subcutaneous) ingrown nail, hypertrophy of the lateral nail fold (lip), and pincer nail. OBJECTIVE: The etiology of pincer nail may be hereditary or acquired. The mechanism of the most common form, an enlarged base of the distal bony phalanx, is discussed. METHODS: Use of roentgenogram and magnetic resonance imaging highlights exophytes of the base and dorsal hyperostosis of the distal phalanx. RESULTS: Global assessment may lead in mild cases to medical therapy. Usually, however, the lateral matrix horn must be surgically removed or cauterized by phenol. Dermal grafting under the nail matrix provides excellent long-term results. PMID- 11277895 TI - Surgical correction of mucous cysts of the nail unit. PMID- 11277896 TI - The treatment of nail apparatus melanoma with Mohs micrographic surgery. PMID- 11277897 TI - Nail unit enchondromas and osteochondromas: a surgical approach. AB - The anatomic singularities of the nail unit explain why bony tumors and osseous outgrowths of the distal phalanx quickly interfere with the nail apparatus. Osteochondromas (or exostosis) are benign osteocartilaginous outgrowths most commonly located on the hallux. They are by far the most common bony lesions affecting the nail unit. While frequently considered benign tumors, subungual exostoses are better regarded as reactional osseous outgrowths rather than true tumors. Enchondromas (better named chondromas) are intraosseous tumors whose location at the distal phalanx is rare. Large tumors may be responsible for phalanx deformity. PMID- 11277898 TI - Metastatic tumors to the nail unit: subungual metastases. AB - BACKGROUND: Cutaneous metastases are variable in location and morphology. Metastatic tumor can present as a subungual lesion in either an oncology patient or a previously cancer-free individual. However, the diagnosis of a subungual metastasis is often not initially considered since the symptoms and appearance of the subungual tumor frequently mimic those of other conditions. OBJECTIVE: To describe the clinical characteristics, radiographic changes, and pathologic findings of the subungual metastases in two women with metastatic carcinoma and to discuss the features of metastatic tumor lesions to the subungual area and distal digits previously reported in oncology patients. METHODS: The clinical presentation, radiologic studies, and pathologic examination of metastatic subungual tumor lesions were described in two oncology patients: a woman with breast cancer and a woman with renal cell carcinoma. The published reports of cancer patients with subungual metastases were reviewed and the following variables were evaluated: the primary origin of the cancer, the histology of the primary tumor, the temporal relationship between the onset of symptoms or the appearance of subungual metastasis and the diagnosis of the visceral malignancy, the symptoms and the morphology of the subungual metastases, the clinical differential diagnosis of subungual metastases, the relationship between the site of origin of the primary tumor and the incidence of metastases either to the fingers and the thumbs or to the toes, the distribution of subungual metastases, the incidence of radiologically confirmable bony involvement of the distal phalanx by metastatic tumor in the digit containing the subungual metastasis, and the prognosis of patients in whom the diagnosis of a subungual metastasis has been confirmed. RESULTS: Subungual metastases most frequently occur in patients with primary tumors of the lung (41%), genitourinary tract organs (17%, of which the kidney represents 11%), and breast (9%). The histology of the primary tumors that was most common included renal cell carcinoma and squamous cell carcinoma. The appearance of the subungual tumor was the first sign of a previously unsuspected primary malignancy in 44% of the patients with subungual metastases. Subungual metastases were frequently painful and most often presented as either an erythematous enlargement or swelling of the distal digit or a red to violacious nodule that distorted either the nail plate or the soft tissue of the distal digit, or both. The lesion was often initially mistaken as an acute infection. The lesion involved one or more digits of the hands in 92% of patients with subungual metastases; symmetrical subungual metastases and metastatic tumor restricted only to the great toes were less commonly observed. In patients with subungual metastases that involved the digits of their hands, the most frequent sites of primary tumor origin were the lung (35%) and the genitourinary tract organs (25%). Radiologic evidence of bony involvement of the respective distal digit was either initially present or subsequently developed in 92% of patients with subungual metastases. Patients with subungual metastases have a poor prognosis; their survival following the diagnosis of the subungual tumor is usually only a few months. CONCLUSION: The clinical differential diagnosis of a new periungual or subungual lesion (with or without an associated nail plate dystrophy) should include tumor metastasis to the nail unit not only in oncology patients, but also in previously cancer-free individuals. PMID- 11277899 TI - Longitudinal melanonychia and melanoma: an unusual case presentation. PMID- 11277900 TI - Treatment of myxoid cysts. PMID- 11277901 TI - Use of dermal grafts in reconstructing deep nasal defects and shaping the ala nasi. AB - BACKGROUND: Deep postsurgical defects on the nose and alar rim pose a challenge to repair. Several techniques are available to reestablish normal contour. If a depressed area is anticipated, dermal grafts can be used to fill the defect and soften contour irregularities. The technique is simple and can prevent the need for a more complicated repair. OBJECTIVE: To reestablish normal contour over nasal defects by using dermal grafts as a tissue filler in conjunction with graft and flap reconstruction. METHODS: Fifteen patients had contour deformities that could be improved with dermal graft insertion under their full-thickness skin grafts or flaps. Fourteen of these patients had nasal defects and one had a vermilion border defect that resulted from tumor removal by Mohs surgery. Patient selection, dermal graft harvesting, and surgical technique are described. RESULTS: All patients showed cosmetic benefit from dermal grafting. Contour was improved in each case. Fourteen patients had no significant complications. One flap showed tip necrosis in a patient who was a smoker. No resorption of the dermal grafts occurred over the 1- to 9-month follow-up period. No cyst formation occurred. CONCLUSION: Dermal grafts can be used during the initial repair of postsurgical nasal defects. These dermis grafts effectively fill the defect and restore contour. The technique is simple, easily mastered, and can obviate the need for more complicated repair. PMID- 11277902 TI - Sutureless skin closure in varicose vein surgery: preliminary results. AB - OBJECTIVE: To describe the preliminary results of a special method of wound closure in varicose vein surgery using the tissue adhesive butyl-2-cyanoacrylate. METHODS: Eighteen consecutive young women (mean age 23 years) underwent partial stripping of the greater saphenous vein for varicose veins of the lower limbs by an external phleboextractor. Their wounds were closed without sutures by means of the adhesive butyl-2-cyanoacrylate. The cutaneous edges were drawn together by linear traction between forceps and the adhesive was applied and allowed to set. Less than 0.5 ml of adhesive was required to complete the entire procedure. Wounds were evaluated at 7 days for infection, dehiscence, and tissue reactions. At 6 months all wounds were rated for cosmesis using a validated visual analog scale, that is, a 100 mm line with "worst scar" at the right end of the line and "best scar" at the left end. All patients were interviewed about their acceptance of tissue adhesive skin closure. RESULTS: The mean time required to close the epidermis with the adhesive was 117 seconds. All patients were followed up for 6 months. At 7 days no adverse outcomes had occurred. Results of wound evaluation at 6 months by the visual analog scale showed scores of 22.2 +/- 13.8 mm (optimal). The percentage of optimal scores was 94.4%, and only one patient (5.6%) had a suboptimal score. Inquiry into the patient's opinions suggested that this procedure was very acceptable. CONCLUSION: Preliminary results with sutureless skin closure in varicose vein surgery have been very encouraging. This fast and cosmetic method of wound repair can replace the need for skin sutures in varicose vein surgery. PMID- 11277903 TI - Sugaring: an ancient method of hair removal. AB - BACKGROUND: Excess body hair can be removed by several methods including shaving, waxing, electrolysis, and lasers. We report herein an ancient technique of hair removal. OBJECTIVE: To describe the technique of sugaring for removal of unwanted hair. METHODS: Skin and paste preparation are described and paste application explained. RESULTS: Hairless and exfoliated skin is achieved. CONCLUSION: Sugaring is a cost effective and practical method of hair removal heretofore unreported in the medical literature. PMID- 11277904 TI - Dog-ear graft technique. PMID- 11277905 TI - Dermatoscopy/ELM for the evaluation of nail-apparatus pigmentation. PMID- 11277906 TI - Regarding tissue expansion and limited or wide undermining. PMID- 11277907 TI - Subtotal keloid excision--a preferable preventative regarding recurrence. PMID- 11277908 TI - A simple maneuver to facilitate dual chondrocutaneous flap segment anastomosis. PMID- 11277909 TI - Commentary on malignant eccrine spiradenoma. PMID- 11277910 TI - Commentary on immediate postoperative laser resurfacing improves second intention healing. PMID- 11277911 TI - Commentary on endovenous laser. PMID- 11277912 TI - Structure-function relationships of hormone-sensitive lipase. AB - Research into the structure-function relationships of lipases and esterases has increased significantly during the past decade. Of particular importance has been the deduction of several crystal structures, providing a new basis for understanding these enzymes. The generated insights have, together with cloning and expression, aided studies on structure-function relationships of hormone sensitive lipase (HSL). Novel phosphorylation sites have been identified in HSL, which are probably important for activation of HSL and lipolysis. Functional and structural analyses have revealed features in HSL common to lipases and esterases. In particular, the catalytic core with a catalytic triad has been unveiled. Furthermore, the investigations have given clear suggestions with regard to the identity of functional and structural domains of HSL. In the present paper, these studies on HSL structure-function relationships and short term regulation are reviewed, and the results presented in relation to other discoveries in regulated lipolysis. PMID- 11277913 TI - An element within the 5' untranslated region of human Hsp70 mRNA which acts as a general enhancer of mRNA translation. AB - The untranslated regions of mRNAs encoding heat-shock proteins have been reported to contain elements important to the post-transcriptional regulation of these key components of the stress response. In this report we describe an element from the 5'UTR of human Hsp70 mRNA that increases the efficiency of mRNA translation. Cloning of this region upstream of the coding sequence of two different reporter genes (firefly luciferase and chloramphenicol acetyltransferase) increases expression of the reporter under normal cell culture conditions by up to an order of magnitude. This effect was observed in three different promoter contexts (HSP, SV40 and CMV) and in six cell lines. The increase in protein production is not accompanied by any alteration in mRNA levels, suggesting that the element facilitates translation. 5' or 3' truncated sequences are ineffective in enhancing reporter expression, suggesting that the activity arises from the secondary structure of the element, rather than from some smaller defined motif. Computer analysis of this region revealed that it is able to form stable secondary structures (DeltaG approximately -292.6 kJ x mol(-1)). The Hsp70 element does not seem to act as an internal ribosome entry site. Incorporation of the sequence into plasmids used for DNA vaccination produces increased antibody responses, confirming that the sequence is functional in primary cells. These data suggest that the 5'UTR of human Hsp70 mRNA plays an important role in determining Hsp70 expression levels, and that it contains an element of general utility in enhancing recombinant protein expression systems. PMID- 11277914 TI - The human neutrophil lipocalin supports the allosteric activation of matrix metalloproteinases. AB - The human neutrophil lipocalin (HNL), a member of the large family of lipocalins that exhibit various physiological functions, is coexpressed in granulocytes with progelatinase B (MMP-9). Part of it is covalently bound to the proenzyme and therefore may play a possible role in the activation process of promatrix metalloproteinases. We now report that HNL is able to accelerate the direct activation of promatrix metalloproteinases slightly. A significant enhancement of the activity could be demonstrated for the HgCl2- and the plasma kallikrein induced activation of all three secretory forms of proMMP-9 and of proMMP-8. The same activating effects were exerted by HNL isolated from granulocytes as well as by the recombinant forms expressed by the yeast Pichia pastoris or by Escherichia coli. This demonstrates that the carbohydrate moiety is not essential for the biological activity of HNL. Activation and activity enhancement are obviously mediated by entrapping the remaining N-terminal sequence residues of the partially truncated proenzyme into the hydrophobic binding pocket of the HNL. In conclusion these results document that HNL can exert an enzyme-activating effect in the regulation of inflammatory and pathophysiological responses of granulocytes in the physiological activation of MMPs that have been subject to limited proteolytic processing. PMID- 11277915 TI - Peptidomics of the pars intercerebralis-corpus cardiacum complex of the migratory locust, Locusta migratoria. AB - The pars intercerebralis-corpora cardiaca system (PI-CC) of insects is the endocrinological equivalent of the hypothalamus-pituitary system of vertebrates. Peptide profiles of the pars intercerebralis and the corpora cardiaca were characterized using simple sampling protocols in combination with MALDI-TOF and electrospray ionization double quadrupole time of flight (ESI-Qq-TOF) mass spectrometric technologies. The results were compared with earlier results of conventional sequencing methods and immunocytochemical methods. In addition to many known peptides, several m/z signals corresponding to putative novel peptides were observed in the corpora cardiaca and/or pars intercerebralis. Furthermore, for a number of peptides evidence was provided about their localization and MALDI TOF analysis of the released material from the corpora cardiaca yielded information on the hormonal status of particular brain peptides. PMID- 11277916 TI - FeMo cofactor biosynthesis in a nifE- mutant of Rhodobacter capsulatus. AB - In all diazotrophic micro-organisms investigated so far, mutations in nifE, one of the genes involved in the biosynthesis of the FeMo cofactor (FeMoco), resulted in the accumulation of cofactorless inactive dinitrogenase. In this study, we have found that strains of the phototrophic non-sulfur purple bacterium Rhodobacter capsulatus with mutations in nifE, as well as in the operon harbouring the nifE gene, were capable of reducing acetylene and growing diazotrophically, although at distinctly lower rates than the wild-type strain. The diminished rates of substrate reduction were found to correlate with the decreased amounts of the dinitrogenase component (MoFe protein) expressed in R. capsulatus. The in vivo activity, as measured by the routine acetylene-reduction assay, was strictly Mo-dependent. Maximal activity was achieved under diazotrophic growth conditions and by supplementing the growth medium with molybdate (final concentration 20-50 microM). Moreover, in these strains a high proportion of ethane was produced from acetylene ( approximately 10% of ethylene) in vivo. However, in in vitro measurements with cell-free extracts as well as purified dinitrogenase, ethane production was always found to be less than 1%. The isolation and partial purification of the MoFe protein from the nifE mutant strain by Q-Sepharose chromatography and subsequent analysis by EPR spectroscopy and inductively coupled plasma MS revealed that FeMoco is actually incorporated into the protein (1.7 molecules of FeMoco per tetramer). On the basis of the results presented here, the role of NifNE in the biosynthetic pathway of the FeMoco demands reconsideration. It is shown for the first time that NifNE is not essential for biosynthesis of the cofactor, although its presence guarantees formation of a higher content of intact FeMoco-containing MoFe protein molecules. The implications of our findings for the biosynthesis of the FeMoco will be discussed. PMID- 11277917 TI - Amine oxidase-like activity of polyphenols. Mechanism and properties. AB - Polyphenols in several oxidation systems gained amine oxidase-like activity, probably due to the formation of the corresponding quinones. In the presence of Cu(II), o- and p-phenolic compounds exhibited amine oxidase-like activity, whereas only the o-phenolic compounds showed the activity in the presence of 1,1 diphenyl-2-picrylhydrazyl radical. The activity was determined by measuring the conversion of benzylamine to benzaldehyde by HPLC. Moreover, gallic acid, chlorogenic acid, and caffeic acid, which are plant polyphenols, converted the lysine residue of bovine serum albumin to alpha-amino-adipic semialdehyde residue, indicating lysyl oxidase-like activity. We also characterized the activity of pyrocatechol, hydroquinone, and pyrogallol in the presence of Cu(II). The oxidative deamination was accelerated at a higher pH, and required O2 and transition metal ions. Furthermore, EDTA markedly inhibited the reaction but not beta-aminopropionitrile, which is a specific inhibitor of lysyl oxidase. Catalase significantly inhibited the oxidation, implying the participation of hydroxyl radical in the reaction, but superoxide dismutase stimulated the oxidation, probably due to its radical formation activity. We discussed the mechanism of the oxidative deamination by polyphenols and the possible significance of the activity for biological systems. PMID- 11277918 TI - Mechanism of phosphoryl transfer by nucleoside diphosphate kinase pH dependence and role of the active site Lys16 and Tyr56 residues. AB - Nucleoside diphosphate (NDP) kinase phosphorylates nucleoside diphosphates with little specificity for the base and the sugar. Although nucleotide analogues used in antiviral therapies are also metabolized to their triphosphate form by NDP kinase, their lack of the 3'-hydroxyl of the ribose, which allows them to be DNA chain terminators, severely impairs the catalytic efficiency of NDP kinase. We have analyzed the kinetics parameters of several mutant NDP kinases modified on residues (Lys16, Tyr56, Asn119) interacting with the gamma-phosphate and/or the 3'-OH of the Mg2+-ATP substrate. We compared the relative contributions of the active-site residues and the substrate 3'-OH for point mutations on Lys16, Tyr56 and Asn119. Analysis of additional data from pH profiles identify the ionization state of these residues in the enzyme active form. X-ray structure of K16A mutant NDP kinase shows no detectable rearrangement of the residues of the active site. PMID- 11277919 TI - Structural and catalytic properties and homology modelling of the human nucleoside diphosphate kinase C, product of the DRnm23 gene. AB - The human DRnm23 gene was identified by differential screening of a cDNA library obtained from chronic myeloid leukaemia-blast crisis primary cells. The over expression of this gene inhibits differentiation and induces the apoptosis of myeloid precursor cell lines. We overproduced in bacteria a truncated form of the encoded protein lacking the first 17 N-terminal amino acids. This truncated protein was called nucleoside diphosphate (NDP) kinase CDelta. NDP kinase CDelta had similar kinetic properties to the major human NDP kinases A and B, but was significantly more stable to denaturation by urea and heat. Analysis of denaturation by urea, using size exclusion chromatography, indicated unfolding without the dissociation of subunits, whereas renaturation occurred via a folded monomer. The stability of the protein depended primarily on subunit interactions. Homology modelling of the structure of NDP kinase CDelta, based on the crystal structure of NDP kinase B, indicated that NDP kinase CDelta had several additional stabilizing interactions. The overall structure of the two enzymes appears to be identical because NDP kinase CDelta readily formed mixed hexamers with NDP kinase A. It is possible that mixed hexamers can be observed in vivo. PMID- 11277920 TI - The uptake by cells of 2-arachidonoylglycerol, an endogenous agonist of cannabinoid receptors. AB - It is not yet clear if the endocannabinoid 2-arachidonoylglycerol (2-AG) is transported into cells through the same membrane transporter mediating the uptake of the other endogenous cannabinoid, anandamide (N-arachidonoylethanolamine, AEA), and whether this process (a) is regulated by cells and (b) limits 2-AG pharmacological actions. We have studied simultaneously the facilitated transport of [14C]AEA and [3H]2-AG into rat C6 glioma cells and found uptake mechanisms with different efficacies but similar affinities for the two compounds (Km 11.0 +/- 2.0 and 15.3 +/- 3.1 microM, Bmax 1.70 +/- 0.30 and 0.24 +/- 0.04 nmol.min 1.mg protein-1, respectively). Despite these similar Km values, 2-AG inhibits [14C]AEA uptake by cells at concentrations (Ki = 30.1 +/- 3.9 microM) significantly higher than those required to either 2-AG or AEA to inhibit [3H]2 AG uptake (Ki = 18.9 +/- 1.8 and 20.5 +/- 3.2 microM, respectively). Furthermore: (a) if C6 cells are incubated simultaneously with identical concentrations of [14C]AEA and [3H]2-AG, only the uptake of the latter compound is significantly decreased as compared to that observed with [3H]2-AG alone; (b) the uptake of [14C]AEA and [3H]2-AG by cells is inhibited with the same potency by AM404 (Ki = 7.5 +/- 0.7 and 10.2 +/- 1.7 microM, respectively) and linvanil (Ki = 9.5 +/- 0.7 and 6.4 +/- 1.2 microM, respectively), two inhibitors of the AEA membrane transporter; (c) nitric oxide (NO) donors enhance the uptake of both [14C]AEA and [3H]2-AG, thus suggesting that 2-AG action can be regulated through NO release; (d) AEA and 2-AG induce a weak release of NO that can be blocked by a CB1 cannabinoid receptor antagonist, and significantly enhanced in the presence of AM404 and linvanil, thus suggesting that transport into C6 cells limits the action of both endocannabinoids. PMID- 11277921 TI - Diversity of O-linked glycosylation patterns between species. Characterization of 25 carbohydrate chains from oviducal mucins of Rana ridibunda. AB - Amphibia egg jelly coats are formed by components secreted along the oviduct. These secretion products overlay the oocytes as they pass along the different oviducal portions. Mucin type glycoproteins are the major constituents of the egg jelly coats. In this study, the O-linked carbohydrate chains of the jelly coats surrounding the eggs of Rana ridibunda were released by alkaline borohydride treatment. Fractionation of the mixture of O-linked oligosaccharide-alditols was achieved by a combination of chromatographic techniques including gel-permeation chromatography, ion-exchange chromatography and high-performance liquid chromatography using an amino-bonded silica column. The primary structures of these O-glycans were determined by one-dimensional and two-dimensional 1H-NMR spectroscopy and matrix-assisted laser-desorption-ionization-time-of flight mass spectrometry. 25 oligosaccharide structures, possessing a core consisting of Gal(beta1-3)GalNAc-ol with or without branching through a GlcNAc residue linked (beta1-6) to the GalNAc residue (core type 2 or core type 1, respectively) are described. The most representative antennae are: HSO3(6)[Fuc(alpha1-3)]GlcNAc; Gal(beta1-2)Gal; Gal(beta1-2)Gal(alpha1-3)[Fuc(alpha1-2)]Gal; GlcA(beta1-3) Gal(beta1-3)[Fuc(alpha1-2)]Gal; GalNAc(alpha1-4)Gal(beta1-4)Gal; Gal(beta1 3)GalNAc(alpha1-4)Gal(beta1-4)Gal and GlcA(beta1-3)Gal(beta1-3)GalNAc. These results confirm the species-specific O-glycosylation of Amphibia oviducal mucins. The significance of this observation should be linked to a symbiotic role of carbohydrates involved in host-parasite interactions. PMID- 11277922 TI - Receptor mediated uptake of peptides that bind the human transferrin receptor. AB - A biopanning process designed to find peptide epitopes specific for cell surface receptors has been used in this study to select seven- and 12-amino-acid peptides capable of binding to and internalizing with the human transferrin receptor (hTfR). Through sequential rounds of negative and positive selection, two peptide sequences were identified that specifically bind to the hTfR. Phage containing the sequences HAIYPRH or THRPPMWSPVWP were inhibited from binding the hTfR in a dose-dependent fashion when peptides of the same sequence were present in a competition assay. Interestingly, transferrin did not compete with either of these sequences for receptor binding, suggesting that these peptides bind a site on the hTfR distinct from the transferrin binding site. When either of these sequences was expressed as a fusion to green fluorescent protein (GFP), the recombinant GFP molecule was internalized in cells expressing the hTfR. These studies suggest that the two peptides can be used to target other proteins into the endosomal pathway. Further, they provide a strategy for identifying peptides that bind to other cell surface receptors that can be used for both diagnostic and therapeutic purposes. PMID- 11277923 TI - Glucose-6-phosphate dehydrogenase-6-phosphogluconolactonase. A novel bifunctional enzyme in malaria parasites. AB - Plasmodium falciparum glucose 6-phosphate dehydrogenase (Pf Glc6PD), compared to other Glc6PDs has an additional 300 amino acids at the N-terminus. They are not related to Glc6PD but are similar to a family of proteins (devb) of unknown function, some of which are encoded next to Glc6PD in certain bacteria. The human devb homologue has recently been shown to have 6-phosphogluconolactonase (6PGL) activity. This suggests Pf Glc6PD may be a bifunctional enzyme, the evolution of which has involved the fusion of adjacent genes. Further functional analysis of Pf Glc6PD has been hampered because parts of the gene could not be cloned. We have isolated and sequenced the corresponding Plasmodium berghei gene and shown it encodes an enzyme (Pb Glc6PD) with the same structure as the P. falciparum enzyme. Pb Glc6PD is 950 amino acids long with significant sequence similarity in both the devb and Glc6PD domains with the P. falciparum enzyme. The P. berghei enzyme does not have an asparagine-rich segment between the N and C halves and it contains an insertion at the same point in the Glc6PD region as the P. falciparum enzyme but the insertion in the P. berghei is longer (110 versus 62 amino acids) and unrelated in sequence to the P. falciparum insertion. Though expression of this enzyme in bacteria produced largely insoluble protein, conditions were found where the full-length enzyme was produced in a soluble form which was purified via a histidine tag. We show that this enzyme has both Glc6PD and 6PGL activities. Thus the first two steps of the pentose phosphate pathway are catalysed by a single novel bifunctional enzyme in these parasites. PMID- 11277924 TI - Acidic pH generated by H+-ATPase pumps triggers the activity of a fusogenic protein associated with rat liver endoplasmic reticulum. AB - Fusogenic protein (FP) is a glycoprotein ( approximately 50 kDa), previously purified by us from rat liver endoplasmic reticulum, which explicates fusogenic activity at acidic pH in vitro. To suggest a possible role of FP in membrane fusion, the topology of the protein in the membrane and the conditions in which FP is operating in microsomes have been investigated. Anti-FP polyclonal antibodies inhibited pure FP activity, but not the protein activity in microsomes, suggesting interaction of antibodies with a part of FP concealed in intact membranes. FP activity in microsomes was lost after treatment with Pronase. Western blot analysis of Pronase-treated microsomes showed that the proteolysis removed a fragment ( approximately 5 kDa). This fragment is exposed on the outer surface of microsomes and involved in fusogenic activity, whereas the largest part of FP is embedded in microsomal vesicles. Therefore, FP can be affected by modifications on the cytosolic and luminal sides of microsomal membranes. Indeed, when microsomal lumen was acidified by H+-ATPase activity, binding and fusion of fluorescent labelled liposomes to microsomes occurred. Direct involvement of FP in the fusogenic event was observed by reconstituting pure FP in liposomes with a preformed H+ gradient. FP triggered a fusion process in response to the acidic interior of liposomes, despite an exterior 7.4 pH unable to promote fusogenic protein activity. As intracellular membrane fusion occurs at neutral pH involving the cytosolic sides of membranes, FP may participate in this event by exploiting the acidic pH formed in the lumen of endoplasmic reticulum through H+-translocating ATPase activity. PMID- 11277925 TI - The structure of the PII-ATP complex. AB - PII is a signal transduction protein that is part of the cellular machinery used by many bacteria to regulate the activity of glutamine synthetase and the transcription of its gene. The structure of PII was solved using a hexagonal crystal form (form I). The more physiologically relevant form of PII is a complex with small molecule effectors. We describe the structure of PII with ATP obtained by analysis of two different crystal forms (forms II and III) that were obtained by co-crystallization of PII with ATP. Both structures have a disordered recognition (T) loop and show differences at their C termini. Comparison of these structures with the form I protein reveals changes that occur on binding ATP. Surprisingly, the structure of the PII/ATP complex differs with that of GlnK, a functional homologue. The two proteins bind the base and sugar of ATP in a similar manner but show differences in the way that they interact with the phosphates. The differences in structure could account for the differences in their activities, and these have been attributed to a difference in sequence at position 82. It has been demonstrated recently that PII and GlnK form functional heterotrimers in vivo. We construct models of the heterotrimers and examine the junction between the subunits. PMID- 11277926 TI - Monoclonal antibodies to mycobacterial DNA gyrase A inhibit DNA supercoiling activity. AB - DNA gyrase is an essential type II topoisomerase found in bacteria. We have previously characterized DNA gyrase from Mycobacterium tuberculosis and Mycobacterium smegmatis. In this study, several monoclonal antibodies were generated against the gyrase A subunit (GyrA) of M. smegmatis. Three, MsGyrA:C3, MsGyrA:H11 and MsGyrA:E9, were further analyzed for their interaction with the enzyme. The monoclonal antibodies showed high degree of cross-reactivity with both fast-growing and slow-growing mycobacteria. In contrast, none recognized Escherichia coli GyrA. All the three monoclonal antibodies were of IgG1 isotype falling into two distinct types with respect to epitope recognition and interaction with the enzyme. MsGyrA:C3 and MsGyrA:H11 IgG, and their respective Fab fragments, inhibited the DNA supercoiling activity catalyzed by mycobacterial DNA gyrase. The epitope for the neutralizing monoclonal antibodies appeared to involve the region towards the N-terminus (residues 351-415) of the enzyme in a conformation-dependent manner. These monoclonal antibodies would serve as valuable tools for structure-function analysis and immunocytological studies of mycobacterial DNA gyrase. In addition, they would be useful for designing peptide inhibitors against DNA gyrase. PMID- 11277927 TI - Exonucleolytic proofreading by p53 protein. AB - The tumour suppressor p53 protein plays an important role in maintaining genetic integrity. Recently, p53 was shown to have an intrinsic 3'-->5' exonuclease activity. The current study has extended the characterization of purified wild type recombinant p53-associated 3'-->5' exonuclease function to demonstrate proofreading activity. p53-associated 3'-->5' exonuclease shows clear preference for degradation of ssDNA over dsDNA substrate. On partial duplex structures, this exonucleolytic activity displays a marked preference for excision of a mismatched vs. a correctly paired 3' terminus which enables the p53 protein to act as a proofreader. However, p53 displays variation in excision of mismatched base pairs. The results demonstrate that p53 exhibits mispair excision with a specificity of A:A > A:G > A:C opposite the template adenine residue and with a specificity of G:A > G:G > G:T opposite the template guanine residue. Hence, the observed specificity of mismatch excision shows that p53 exonucleolytic proofreading preferentially repairs transversion mutations. As part of an investigation of the functional interaction between p53 and DNA polymerase, the influence of p53 on the accuracy of DNA synthesis was determined with exonuclease deficient murine leukemia virus (MLV) reverse transcriptase (RT), representing a relatively low fidelity enzyme. Using an in vitro biochemical assay with 3' terminal mismatch-containing DNA template primers, it was shown that wild-type recombinant p53 protein enhanced the DNA replication fidelity of MLV RT. A functional interaction between the exonuclease (p53) and polymerase (MLV RT) activities was observed; excision of mispairs by p53 was followed by further elongation onto correctly base-paired 3'-termini by MLV RT. Furthermore, the formation of 3'-mispair and subsequent mispair extension by the enzyme were decreased substantially in the presence of p53. The fact that the exonuclease deficient MLV RT is more accurate in the presence of p53, suggests that p53 protein may function as an external proofreading exonuclease for viral enzyme. The observed decrease in initial nucleotide misincorporation and 3'-terminal mispair extension by MLV RT in the presence of p53, indicates the mechanism by which p53 affects the DNA replication fidelity of exonuclease-deficient DNA polymerase. PMID- 11277928 TI - Characterization of the Cph1 holo-phytochrome from Synechocystis sp. PCC 6803. AB - The cph1 gene from the unicellular cyanobacterium Synechoycstis sp. PCC 6803 encodes a protein with the characteristics of plant phytochromes and histidine kinases of two-component phospho-relay systems. Spectral and biochemical properties of Cph1 have been intensely studied in vitro using protein from recombinant systems, but virtually nothing is known about the situation in the natural host. In the present study, His6-tagged Cph1 was isolated from Synechocystis cells. The cph1-his gene was expressed either under the control of the natural cph1 promoter or over-expressed using the strong promoter of the psbA2 gene. Upon purification with nickel affinity chromatography, the presence of Cph1 in extracts was confirmed by immunoblotting and Zn2+-induced fluorescence. The Cph1 extracts exhibited a red/far-red photoactivity characteristic of phytochromes. Difference spectra were identical with those of the phycocyanobilin adduct of recombinant Cph1, implying that phycocyanobilin is the chromophore of Cph1 in Synechocystis. PMID- 11277929 TI - Insect silk contains both a Kunitz-type and a unique Kazal-type proteinase inhibitor. AB - Insect silk is made up of structural fibrous (fibroins) and sticky (sericins) proteins, and contains a few small peptides of hitherto unknown functions. We demonstrate that two of these peptides inhibit bacterial and fungal proteinases (subtilisin, proteinase K and pronase). These 'silk proteinase inhibitors' 1 and 2 (SPI 1 and 2) are produced in the middle section of the silk-secreting glands prior to cocoon spinning and their production is controlled at transcription level. The full length cDNA of pre-SPI 1 contains 443 nucleotides and encodes a peptide of 76 amino-acid residues, of which 20 make up a signal sequence. The mature SPI 1 (6056.7 Da, 56 residues) is a typical thermostable Kunitz-type proteinase inhibitor with Arg in P1 position. The cDNA of pre-SPI 2 consists of 260 nucleotides and yields a putative secretory peptide of 58 amino-acid residues. The functional SPI 2 (3993 Da, 36 residues) is a single-domain Kazal type proteinase inhibitor with unique structural features: free segment of the N terminus is reduced to a single amino-acid residue, lack of CysI and CysV precludes formation of the A-ring and provides increased flexibility to the C ring, and absence of several residues around the normal position of CysV shortens and changes the alpha helix segment of the protein. The structure reveals that the length and arrangement of the B-ring, including exposure of the P1 residue, and the position of the C-terminus relative to the B-loop, are essential for the activity of the Kazal-type inhibitors. PMID- 11277930 TI - Processing of tumor necrosis factor by the membrane-bound TNF-alpha-converting enzyme, but not its truncated soluble form. AB - Tumour necrosis factor (TNF)-alpha-converting enzyme (TACE) is a membrane protein belonging to the ADAM (a disintegrin and metalloproteinase) family that cleaves various membrane proteins, including the proform of TNF-alpha. In this study, we constructed expression vectors for the membrane-bound full-length TACE (mTACE) and its truncated soluble form (sTACE). When a human TNF-alpha expression vector was introduced into human 293 cells, processing of TNF-alpha to its mature form was enhanced by coexpressing mTACE, and this processing was inhibited by a metalloproteinase inhibitor. On the other hand, coexpression of sTACE had no effect on the processing of TNF-alpha, although the culture medium of sTACE transfected cells could cleave a peptide containing the TNF-alpha cleavage site. Fas ligand (FasL)-transfected 293 cells released a considerable amount of soluble FasL, and coexpression of neither mTACE nor sTACE enhanced this shedding. Immunoprecipitation and Western blotting analysis with cells that were cotransfected with TACE and TNF-alpha indicated that both mTACE and sTACE could interact with the proform of TNF-alpha. In the same assay, neither mTACE nor sTACE interacted with FasL. The catalytic domain-lacking TACE mutant, which could also interact TNF-alpha, showed a dominant negative effect on not only TNF-alpha secretion but also FasL secretion. These results suggest that binding of the membrane-anchored but not the soluble form of TACE to TNF-alpha results in efficient ectodomain shedding, and that FasL secretase is a metalloproteinase similar, but not identical, to TACE. PMID- 11277931 TI - Defined sequence segments of the small heat shock proteins HSP25 and alphaB crystallin inhibit actin polymerization. AB - The interaction of small heat shock proteins (sHSPs) with the actin cytoskeleton has been described and some members of this family, e.g. chicken and murine HSP25 (HSP27), inhibit the polymerization of actin in vitro. To analyse the molecular basis of this interaction, we synthesized a set of overlapping peptides covering the complete sequence of murine HSP25 and tested the effect of these peptides on actin polymerization in vitro by fluorescence spectroscopy and electron microscopy. Two peptides comprising the sequences W43 to R57 (peptide 6) and I92 to N106 (peptide 11) of HSP25 were found to be potent inhibitors of actin polymerization. Phosphorylation of N-terminally extended peptide 11 at serine residues known to be phosphorylated in vivo resulted in decline of their inhibitory activity. Interestingly, peptides derived from the homologous peptide 11 sequence of murine alphaB-crystallin showed the same behaviour. The results suggest that both HSP25 and alphaB-crystallin have the potential to inhibit actin polymerization and that this activity is regulated by phosphorylation. PMID- 11277932 TI - Transport of L-carnitine in isolated cerebral cortex neurons. AB - The accumulation of carnitine was measured in cerebral cortex neurons isolated from adult rat brain. This process was found to be lowered by 40% after preincubation with ouabain and with SH-group reagents (N-ethylmaleimide and mersalyl). The initial velocity of carnitine transport was found to be inhibited by 4-aminobutyrate (GABA) in a competitive way (Ki = 20.9 +/- 2.4 mM). However, of various inhibitors of GABA transporters, only nipecotic acid and very high concentrations of 1-[2-([(diphenylmethylene)amino]oxy)ethyl]-1,2,5,6-tetrahydro-3 pyridinecarboxylic acid hydrochloride (NO-711) acid decreased carnitine accumulation while betaine, taurine and beta-alanine had no effect. The GABA transporters expressed in Xenopus laevis oocytes did not transport carnitine. Moreover, carnitine was not observed to diminish the accumulation of GABA in cerebral cortex neurons, which further excluded a possible involvement of the GABA transporter GAT1 in the process of carnitine accumulation, despite the expression of this protein in the cells under study. The absence of carnitine transporter OCTN2 in rat cerebral cortex neurons (K. A. Nalecz, D. Dymna, J. E. Mroczkowska, A. Broer, S. Broer, M. J. Nalecz and R. Cecchelli, unpublished results), together with the insensitivity of carnitine accumulation towards betaines, implies that a novel transporting protein is present in these cells. PMID- 11277933 TI - Human phosphatidylinositol 4-kinase isoform PI4K92. Expression of the recombinant enzyme and determination of multiple phosphorylation sites. AB - Human phosphatidylinositol 4-kinase, isoform PI4K92, was expressed as His6 tagged protein in Sf9 cells reaching a level of approximately 5% of cellular protein. The enzyme can be purified nearly to homogeneity in a single step by absorption/desorption on Ni/nitriloacetic acid agarose magnetic beads. High Km values in the millimolar range for ATP and PtdIns as well as only a moderate inhibition by adenosine and a sensitivity to Wortmannin (IC50 approximately 300 nM) characterize the enzyme as a type 3 PI4K. The enzyme produces PtdIns4P as product. The isolated enzyme is a phosphoprotein, additionally phosphate is incorporated by incubation with ATP/Mg or ATP/Mn. Phosphorylation sites were mapped by MALDI-MS and LC-MS/MS at the following positions: S258, T263, S266, S277, S294, T423, S496, T504. Accordingly, a stretch of 81 amino acids between the common and the C-terminal catalytic domain was designated phosphorylation domain. PMID- 11277934 TI - Trichosanthin interacts with acidic ribosomal proteins P0 and P1 and mitotic checkpoint protein MAD2B. AB - Trichosanthin is a ribosome-inactivating protein with multiple pharmacological properties. By a yeast two-hybrid system, ribosomal phosphoproteins P0 and P1 and a putative mitotic checkpoint protein, MAD2B, were found to interact with an active-site mutated trichosanthin (TCS). The interactions were verified by an in vitro binding assay of recombinant wild-type TCS and target proteins. The interaction domain of P0 was mapped to amino acids 220-273, which had been previously reported to be involved in the interaction with P1 and P2 in yeast. Consistent with our previous finding that the last seven residues of TCS are not essential for an active conformation, the same deletion did not affect the interaction with P0. Our present study suggests that TCS may disrupt the binding of elongation factors to the P-complex, in addition to the well-known N glycosidase activity for ribosome inactivation. PMID- 11277935 TI - Cytotoxic mechanism of the ribotoxin alpha-sarcin. Induction of cell death via apoptosis. AB - Alpha-sarcin is a ribosome-inactivating protein that has been well characterized in vitro, but little is known about its toxicity in living cells. We have analyzed the mechanism of internalization of alpha-sarcin into human rhabdomyosarcoma cells and the cellular events that result in the induction of cell death. No specific cell surface receptor for alpha-sarcin has been found. The toxin is internalized via endocytosis involving acidic endosomes and the Golgi, as deduced from the ATP requirement and the effects of NH4Cl, monensin and nigericin on its cytotoxicity. Specific cleavage of 28S rRNA in cultured rhabdomyosarcoma cells, associated with protein biosynthesis inhibition, has been detected. alpha-Sarcin kills rhabdomyosarcoma cells via apoptosis: incubation of cells with alpha-sarcin at a concentration below its IC50 induces internucleosomal genomic DNA fragmentation, reversion of membrane asymmetry, activation of caspase-3-like activity and cleavage of poly(ADP-ribose)polymerase. Apoptosis is not a general direct consequence of protein biosynthesis inhibition, as deduced from the comparative analysis of the effects of alpha-sarcin and cycloheximide; the latter does not induce apoptosis even at concentrations far beyond its IC50, where protein biosynthesis is null. Experiments with a catalytically inactive alpha-sarcin mutant, neither toxic nor apoptotic, reveal that induced apoptosis is directly related to the effects of catalytic activity of the toxin on the ribosomes. The caspase inhibitor z-VAD-fmk does not suppress protein synthesis inhibition by alpha-sarcin. Together, these data suggest that alpha-sarcin-induced caspase activation is a pathway downstream of the 28S rRNA catalytic cleavage and consequent protein biosynthesis inhibition. PMID- 11277936 TI - Solution structure of microcin J25, the single macrocyclic antimicrobial peptide from Escherichia coli. AB - The three-dimensional solution structure of microcin J25, the single cyclic representative of the microcin antimicrobial peptide class produced by enteric bacteria, was determined using two-dimensional 1H NMR spectroscopy and molecular modeling. This hydrophobic 21-residue peptide exhibits potent activity directed to Gram-negative bacteria. Its primary structure, cyclo( V1GIGTPISFY10GGGAGHVPEY20F-), has been determined previously [Blond, A., Peduzzi, J., Goulard, C., Chiuchiolo, M. J., Barthelemy, M., Prigent, Y., Salomon, R.A., Farias, R.N., Moreno, F. & Rebuffat, S. (1999) Eur. J. Biochem., 259, 747-755]. Conformational parameters (3JNHCalphaH coupling constants, quantitative nuclear Overhauser enhancement data, chemical shift deviations, temperature coefficients of amide protons, NH-ND exchange rates) were obtained in methanol solution. Structural restraints consisting of 190 interproton distances inferred from NOE data, 11 phi backbone dihedral angle and 9 chi1 angle restraints derived from the coupling constants and three hydrogen bonds in agreement with the amide exchange rates were used as input for simulated annealing calculations and energy minimization in the program XPLOR. Microcin J25 adopts a well-defined compact structure consisting of a distorted antiparallel beta sheet, which is twisted and folded back on itself, thus resulting in three loops. Residues 7-10 and 17-20 form the more regular part of the beta sheet. The region encompassing residues Gly11-His16 consists of a distorted beta hairpin, which divides into two small loops and is stabilized by an inverse gamma turn and a type I' beta turn. The reversal of the chain leading to the Phe21-Pro6 loop results from a mixed beta/gamma turn. A cavity, in which the hydrophilic Ser8 side-chain is confined, is delimited by two crab pincer-like regions that comprise residues 6-8 and 18-1. PMID- 11277937 TI - Extensive deproteinization of Dictyostelium discoideum RNase P reveals a new catalytic activity. AB - Nuclear Dictyostelium discoideum RNase P was subjected to vigorous deproteinization procedures. After treatment with proteinase K followed by phenol extraction of samples containing D. discoideum RNase P activity, a new enzymatic activity was recovered. The proteinase K/phenol/SDS treated enzyme cleaves Schizossacharomyces pombe tRNAser (supS1), D. discoideum tRNASer and tRNALeu precursors several nucleotides upstream of the cleavage site of RNase P, liberating products with 5'-hydroxyl ends. This activity seems to be associated with one or two RNA molecules copurifying with D. discoideum RNase P activity as judged by its inhibition in the presence of micrococcal nuclease, which is in contrast to its resistance to proteinase K/phenol/SDS treatment. PMID- 11277938 TI - Identification of the rat adrenal zona fasciculata/reticularis specific protein, inner zone antigen (IZAg), as the putative membrane progesterone receptor. AB - Using immunological methods, a protein specific to the inner zones of the rat adrenal cortex, and called inner zone antigen (IZAg), was previously shown to have two interrelated forms of 26 kDa (IZAg1) and 55-60 kDa (IZAg2), and to have an action on steroid hydroxylation. After two-dimensional gel electrophoresis, and immunoaffinity column purification, N-terminal amino-acid analysis showed that the first 12 amino acids were identical to those of a recently described putative membrane located progesterone receptor (PPMR). RT-PCR was then used to generate the cDNA of this protein, using RNA extracted from rat adrenals. A glutathione S-transferase (GST)-fusion construct was expressed in Escherichia coli, and shown to generate an immunoreactive product of molecular mass consistent with its identification as IZAg1. More detailed examination of the distribution of this protein, not only in the zona fasciculata/reticularis of the adrenal cortex, but also in the Leydig cell, kidney and liver, suggest it may have a role in steroid hormone synthesis and/or metabolism. PMID- 11277939 TI - A new structural type for Haemophilus influenzae lipopolysaccharide. Structural analysis of the lipopolysaccharide from nontypeable Haemophilus influenzae strain 486. AB - Structural elucidation of the sialylated lipopolysaccharide (LPS) of non-typeable Haemophilus influenzae (NTHi) strain 486 has been achieved by the application of high-field NMR techniques and ESI-MS along with composition and linkage analyses on O-deacylated LPS and oligosaccharide samples. It was found that the LPS contains the common element of H. influenzae, L-alpha-D-Hepp-(1-->2)-[PEtn-->6]-L alpha-D-Hepp-(1-->3)-[beta-D-Glcp-(1-->4)]-L-alpha-D-Hepp-(1-->5)-[PPEtn-->4] alpha-Kdop-(2-->6)-Lipid A, but instead of glycosyl substitution of the terminal heptose residue (HepIII) at the O2 position observed in other H. influenzae strains, HepIII is chain elongated at the O3 position by either lactose or sialyllactose (i.e. alpha-Neu5Ac-(2-->3)-beta-D-Galp-(1-->4)-beta-D-Glcp). The LPS is substituted by an O-acetyl group linked to the O2 position of HepIII and phosphocholine (PCho) which was located at the O6 position of a terminal alpha-D Glcp residue attached to the central heptose, a molecular environment different from what has been reported earlier for PCho. In addition, minor substitution by O-linked glycine to the LPS was observed. By investigation of LPS from a lpsA mutant of NTHi strain 486, it was demonstrated that the lpsA gene product also is responsible for chain extension from HepIII in this strain. The involvement of lic1 in expression of PCho was established by investigation of a lic1 mutant of NTHi strain 486. PMID- 11277940 TI - Protein kinase C regulates transcription of the human guanylate cyclase C gene. AB - Guanylate cyclase C is the receptor for the bacterial heat-stable enterotoxins and guanylin family of peptides, and mediates its action by elevating intracellular cGMP levels. Potentiation of ligand-stimulated activity of guanylate cyclase C in human colonic T84 cells is observed following activation of protein kinase C as a result of direct phosphorylation of guanylate cyclase C. Here, we show that prolonged exposure of cells to phorbol esters results in a decrease in guanylate cyclase C content in 4beta-phorbol 12-myristate 13-acetate treated cells, as a consequence of a decrease in guanylate cyclase C mRNA levels. The reduction in guanylate cyclase C mRNA was inhibited when cells were treated with 4beta-phorbol 12-myristate 13-acetate (PMA) in the presence of staurosporine, indicating that a primary phosphorylation event by protein kinase C triggered the reduction in RNA levels. The reduction in guanylate cyclase C mRNA levels was not due to alterations in the half-life of guanylate cyclase C mRNA, but regulation occurred at the level of transcription of guanylate cyclase C mRNA. Expression in T84 cells of a guanylate cyclase C promoter-luciferase reporter plasmid, containing 1973 bp of promoter sequence of the guanylate cyclase C gene, indicated that luciferase activity was reduced markedly on PMA treatment of cells, and the protein kinase C-responsive element was present in a 129-bp region of the promoter, containing a HNF4 binding element. Electrophoretic mobility shift assays using an oligonucleotide corresponding to the HNF4 binding site, indicated a decrease in binding of the factor to its cognate sequence in nuclear extracts prepared from PMA-treated cells. We therefore show for the first time that regulation of guanylate cyclase C activity can be controlled at the transcriptional level by cross-talk with signaling pathways that modulate protein kinase C activity. We also suggest a novel regulation of the HNF4 transcription factor by protein kinase C. PMID- 11277941 TI - Defects of type I procollagen metabolism correlated with decrease of prolidase activity in a case of lethal osteogenesis imperfecta. AB - We have studied the structure and metabolism of type I procollagen in a case of perinatal lethal osteogenesis imperfecta (OI) type II. Cultured skin fibroblasts from the proband synthesized both normal and abnormal forms of type I procollagen. Some abnormal, overmodified molecules were secreted by OI cells, although less efficiently than normal molecules from control cells. The OI fibroblasts accumulated large amounts of abnormal proalpha1(I) and proalpha2(I) chains intracellularly. The extracellular collagenolytic activity was decreased compared to control cells. Furthermore, OI cells produced less type I procollagen and demonstrated lower capacity to synthesize DNA than control cells. We have found that in contrast to prolinase activity, the activity of prolidase (an enzyme essential for collagen synthesis and cell growth) is also significantly reduced in OI cells. No differences were found in the amount of the enzyme protein recovered from both the OI and control cells. However, we found that expressions of beta1 integrin and insulin-like growth factor-I receptor (receptors known to play an important role in up regulation of prolidase activity) were decreased in OI cells compared to control cells. The decrease in prolidase activity may provide an important mechanism of altered cell growth and collagen metabolism involved in producing the perinatal lethal form of the OI phenotype. PMID- 11277942 TI - Formation and properties of hybrid photosynthetic F1-ATPases. Demonstration of different structural requirements for stimulation and inhibition by tentoxin. AB - A hybrid ATPase composed of cloned chloroplast ATP synthase beta and gamma subunits (betaC and gammaC) and the cloned alpha subunit from the Rhodospirillum rubrum ATP synthase (alphaR) was assembled using solubilized inclusion bodies and a simple single-step folding procedure. The catalytic properties of the assembled alpha3Rbeta3CgammaC were compared to those of the core alpha3Cbeta3CgammaC complex of the native chloroplast coupling factor 1 (CF1) and to another recently described hybrid enzyme containing R. rubrum alpha and beta subunits and the CF1 gamma subunit (alpha3Rbeta3RgammaC). All three enzymes were similarly stimulated by dithiothreitol and inhibited by copper chloride in response to reduction and oxidation, respectively, of the disulfide bond in the chloroplast gamma subunit. In addition, all three enzymes exhibited the same concentration dependence for inhibition by the CF1 epsilon subunit. Thus the CF1 gamma subunit conferred full redox regulation and normal epsilon binding to the two hybrid enzymes. Only the native CF1 alpha3Cbeta3CgammaC complex was inhibited by tentoxin, confirming the requirement for both CF1 alpha and beta subunits for tentoxin inhibition. However, the alpha3Rbeta3CgammaC complex, like the alpha3Cbeta3CgammaC complex, was stimulated by tentoxin at concentrations in excess of 10 microm. In addition, replacement of the aspartate at position 83 in betaC with leucine resulted in the loss of stimulation in the alpha3Rbeta3CgammaC hybrid. The results indicate that both inhibition and stimulation by tentoxin require a similar structural contribution from the beta subunit, but differ in their requirements for alpha subunit structure. PMID- 11277943 TI - High affinity binding of paclitaxel to human serum albumin. AB - Paclitaxel, a very potent antitumor agent is a hydrophobic molecule with low aqueous solubility. Its currently used formula (Taxol) contains the drug in a 1 : 1 (v/v) mixture of ethanol and Cremophor EL. To minimize vehicle-related toxicity, we developed a novel, water-soluble formulation in which paclitaxel is bound noncovalently to human serum albumin. For this purpose, studies of the paclitaxel-albumin binding equilibrium were performed. Paclitaxel dissolved in ethanol was added to the aqueous solution of human serum albumin. Precipitated paclitaxel was removed and unbound drug was separated by ultrafiltration. Paclitaxel concentration was measured by RP-HPLC. Binding data were evaluated based both on the Scatchard plot and the general binding equation describing binding equilibria with the stepwise stoichiometric binding constants. The Scatchard plot was found to be curvilinear with a slight positive slope of the final part. Parameters of high affinity specific binding were determined from the initial part of the curve (nsp = 1.3 and Ksp = 1.7 x 10(6) M(-1)). Stoichiometric binding constants were estimated by fitting the general binding equation to the experimental data (K1 = 2.4 x 10(6) M(-1) and K2 = 1.0 x 10(5) M(-1)). Saturation of the protein with paclitaxel, similarly to other ligands of albumin, could not be reached. The greatest observed value of r (number of paclitaxel molecules bound to one albumin molecule) was 6.6. PMID- 11277944 TI - Itraconazole: an effective oral antifungal for onychomycosis. PMID- 11277945 TI - Post herpetic neuralgia and the dermatologist. PMID- 11277946 TI - So moisturizers may cause trouble! PMID- 11277947 TI - Human cowpox infection in Sharkia Governorate, Egypt. AB - BACKGROUND AND OBJECTIVE: In the last few years, outbreaks of an apparently specific dermatosis occurred during the months of late summer and early autumn in our locality. Our aim in this study was to reveal the underlying etiology of this dermatosis. SUBJECTS AND METHODS: Sixty patients with the disease were studied clinically, epidemiologically, histopathologically, and ultrastructurally. RESULTS: The results of all methods suggest that the dermatosis is most probably a human cowpox infection. Electron microscopy showed unenveloped cowpox virons. CONCLUSIONS: This apparently specific dermatosis is due to human cowpox infection. Future investigations will be needed to better define this important zoonosis frequently passed by cats to humans and carried by rodents. PMID- 11277948 TI - Frequency of toenail onychomycosis in patients with cutaneous manifestations of chronic venous insufficiency. AB - BACKGROUND: Chronic venous insufficiency (CVI) can originate onychopathy per se. We have anecdotally observed nail changes in patients with CVI, but there are few studies which determine the frequency of both onychopathy and onychomycosis in these patients OBJECTIVE: The aim of the study was to determine the frequency of nail pathology and onychomycosis in patients with CVI PATIENTS AND METHODS: We included 36 adult patients, both men and women, aged from 18 to 59 years, with clinically documented venous leg ulcers. All patients were examined by a dermatologist and the venous leg ulcers were classified according to severity in three grades. The nail changes were described and a mycological examination was performed. We obtained a small fragment of the nail for histological examination. In 27 patients, we also performed functional studies to determine the type of venous insufficiency. RESULTS: The ratio of women to men was 5 : 1. The mean age of patients was 46.39 +/- 8.51 years, men being slightly younger than women. Ten patients had ulcers of grade I severity, 12 had grade II, and 14 had grade III. The overall time of evolution of the cutaneous lesions was 11.02 +/- 10.11. Fourteen patients had superficial venous insufficiency, whereas 13 had deep venous insufficiency. Twenty-two (61.11%) of our patients had nail alterations. These nail changes were related more to the type of vascular affection than with the severity of cutaneous involvement. In more than half of the cases (59.09%), onychomycosis was the cause of the nail changes. The overall frequency of onychomycosis was 36.11%. The etiologic agent of onychomycosis was isolated in 38.46% of the cases, and Trichophyton rubrum was the most frequent agent. The histologic examination of the nail plate showed a low sensitivity (62%) but a high specificity (100%) in the detection of nail plate parasitization. No clinical differences could be established between the nail changes observed in patients with true onychomycosis and those with nonfungal onychopathy. CONCLUSIONS: Nail changes are common in patients with venous leg ulcer, and onychomycosis accounts for slightly more than half of the cases. We therefore recommend a routine mycological examination in patients wit nail changes and cutaneous manifestations of CVI, to diagnose or rule out onychomycosis, and therefore avoid overtreating patients without onychomycosis with antimycotics. PMID- 11277949 TI - Childhood cutaneous tuberculosis: a study over 25 years from northern India. AB - AIMS: We undertook this study to analyse the pattern of childhood cutaneous tuberculosis prevailing in northern India over the past 25 years and to highlight differences from and similarities to adult tuberculosis. MATERIALS AND METHODS: Clinical records of children with cutaneous tuberculosis who attended the Nehru Hospital attached to the Postgraduate Institute of Medical Education and Research, Chandigarh, India, between January 1975 to December 1999 were analysed. RESULTS: Four hundred and two patients with cutaneous tuberculosis were examined during the 25-year period of observation. These patients formed 0.1% of the total dermatology outpatients. Seventy-five (18.7%) of these 402 cases were children ( 10 mm). Histopathologic reports were available for evaluation in all 75 children. Out of 30 cases of LV, 24 (80%) showed classical tuberculous histology. In contrast, out of 40 cases with SFD, only 19 (47.5%) showed classical histology. Classical tuberculous histology was noted in all 3 (100%) cases of TVC and 1 (100%) case each with tuberculids and gumma. Tubercle bacilli could be demonstrated in 4/30 (13.3%) cases with LV and 9/40 (22.5%) cases with SFD. Systemic involvement was seen in 16 (21.3%) children, of whom 3 (18.8%) had LV and 13 (81.3%) SFD. The lungs were the most common organs involved in 8 (50.0%) children followed by bone(s) in 4 (25.0%), abdomen in 2 (12.5%), and both lung and bone in 1 (6.3%) child. In contrast to adults, girls outnumbered boys in the childhood series; SFD was a common form of presentation in contrast to LV and TVC, tuberculous gumma and tuberculids were noted less often. In both children and adults, Mantoux reactivity did not correlate with the extent of the disease; patients with disseminated disease were found to be less often vaccinated with BCG and regional lymphadenopathy was noted more often in patients with disseminated disease. CONCLUSIONS: In the whole spectrum of cutaneous tuberculosis, there is a proportion of patients with dissemination (systemic involvement) who are of great epidemiological significance as they require a change in the standard therapeutic regimens recommended for cutaneous tuberculosis. PMID- 11277950 TI - Elevated plasma homocysteine levels in patients on isotretinoin therapy for cystic acne. AB - BACKGROUND: The use of Isotretinoin (Iso) for cystic acne (CA) therapy includes marked side-effects such as dyslipidemia, increased liver enzymes, and reduction of biotinidase activity. Moreover, Homocysteine (Hcy), an amino acid, is metabolized in the liver requiring folate, vitamin B6, vitamin B12, and the activity of enzymes, i.e. cystathionine-beta-synthase. Increased blood levels of Hcy are associated with premature occlusive vascular disease. OBJECTIVE: The aim of this study was the evaluation of Hcy levels and the responsible vitamins for its metabolism in patients with CA on Iso treatment. METHODS AND RESULTS: Twenty eight patients with CA were submitted to laboratory examinations before (Value 1) and after (Value 2) 45 days on Iso (0.5 mg/kg/24 h) therapy. Blood levels of Hcy and vitamin B6 were evaluated by HPLC methods, and folate and vitamin B12 using a commercial Kit. Hcy levels (Value 1 = 7.86 +/- 1.6 micromol/L; Value 2 = 13.65 +/ 3.3 micromol/L; P < 0.001) were statistically significantly increased in patients on treatment. Vitamins were unaltered, and lipids and liver enzymes increased. Significant correlation between Hcy levels, vitamins, and liver enzymes was found. Methionine loading tests performed in nine patient-volunteers showed an abnormal response post-treatment. CONCLUSIONS: It is suggested that the elevated Hcy levels in patients after 45 days on Iso therapy could be due either to the 'inhibition' of cystathionine-beta-synthase by the drug and/or their liver dysfunction. Daily vitamin supplementation along with frequent evaluations of Hcy blood levels are recommended for the prevention of a premature occlusive vascular disease. PMID- 11277951 TI - Cutaneous lymphomas in Tokyo: analysis of 62 cases in a dermatology clinic. AB - BACKGROUND: The epidemiology of cutaneous lymphomas revealed that the incidence of lymphomas differed depending on various factors including area, race, and sex, among others. OBJECTIVE: This study was undertaken to analyse the incidence of cutaneous lymphomas in Tokyo. METHODS: The clinical records and histologic material from 50 patients with lymphomas of the skin and 12 patients with lymphomatoid papulosis seen during the last 10 years at the Department of Dermatology, The Jikei University School of Medicine, Tokyo, have been reviewed. RESULTS: T-cell lymphomas including mycosis fungoides (MF)-Sezary's syndrome (SS) complex and adult T-cell leukemia/lymphoma (ATL) were more frequent than B-cell lymphomas. The incidence of ATL is associated with the number of human T-cell lymphotropic virus type 1 (HTLV-1) carriers in the general population. Cutaneous B-cell lymphoma (CBCL) is not as rare as previously thought in Japan. CONCLUSIONS: Although the frequency of these cases depends on many unrelated factors, these values can provide a rough indication of the incidence of cutaneous lymphomas in Tokyo. The incidence of cutaneous lymphomas may be influenced by changes in environmental factors including viral infections. PMID- 11277952 TI - Which intercurrent infections are associated with maculopapular cutaneous drug reactions? A case-control study. AB - BACKGROUND: Patients with lymphotrophic viral infections are at increased risk for cutaneous drug reactions (CDRs). However, the association between other intercurrent infections and maculopapular CDRs has not been evaluated by epidemiologic methods. OBJECTIVE: We conducted a case-control study in order to evaluate the exposure to intercurrent infections in patients with maculopapular CDRs. METHODS: Data were obtained through assessment of files of 53 patients hospitalized for maculopapular CDRs in the Department of Dermatology and 159 control patients. Exposure to intercurrent infections was recorded in patients and controls. RESULTS: An intercurrent infectious disease was documented in 31/53 (58.5%) of patients with CDRs, as compared to 12/159 (7.5%) patients in the control group (OR 17.26, 95% CI: 7.24-42.00). Maculopapular CDRs were associated with respiratory tract infections (OR 20.53, 95% CI: 5.20-94.45), and urinary tract infections (OR 20.61, 95% CI: 2.36-465.99), but not with skin infections (OR 3.83, 95% CI: 0.85-17.87) or other infections. CONCLUSIONS: Our study implies that maculopapular CDRs are associated with respiratory tract infections as well as urinary tract infections. Further study is needed to evaluate the role of intercurrent infections in the pathogenesis of CDRs. PMID- 11277954 TI - Chronic lymphocytic leukemia presenting as cutaneous and bone involvement. PMID- 11277953 TI - Application of computerized image analysis in pigmentary skin diseases. AB - BACKGROUND: Melanocyte number and the amount of melanin pigment are related to diagnosis and treatment of pigmentary skin diseases. Various histologic methods are used, such as Fontana-Masson stain for melanin pigment or immunohistochemical stain for melanocytes. Recently, computerized image analysis has been applied to many fields to avoid interobserver bias. In this study, we applied a computerized image analysis to assess the melanin content and melanocyte density of human epidermis. METHODS: We evaluated the skin biopsy specimens (paraffin blocks) from normal human skin (33 +/- 6.6, n = 11) and diseased skins; vitiligo (32 +/- 10.0, n = 8), melasma (35 +/- 8.6, n = 11), and lentigo senilis (40 +/- 7.2, n = 11) (mean age +/- SD). Each specimen was stained with Fontana-Masson for melanin pigments and immunohistochemical method for melanocytes. Quantitative analysis of melanin pigment and melanocyte number (density) were investigated through two methods: (1) two dermatologists measured the visual scales; and (2) computerized image analysis was used to measure melanin content indices (MCI). The data were evaluated using one-way ANOVA. RESULTS: The visual scale of the Fontana-Masson stain was the highest for lentigo senilis (3.8 +/- 0.40), followed by melasma (2.6 +/- 0.67), normal skin (1.8 +/- 0.60) and vitiligo (0) (P < 0.05). These findings were consistent with objective measurements made by computerized image analysis. MCI values were 120.3 +/- 20.74 for lentigo senilis, 81.1 +/- 19.27 for melasma, 45.5 +/- 16.92 for normal skin, and 0.3 +/- 0.30 for vitiligo in decreasing order (P < 0.05). MC/1E (melanocyte number per 1 mm epidermis) was about two fold larger in lentigo senilis (18.1 +/- 8.92) than melasma (9.7 +/- 2.40) or normal skin (9.3 +/- 2.67) (P < 0.05). MC/1B (melanocyte number per 1 mm basal layer) was about 1.5 fold higher in lentigo senilis (13.5 +/- 4.17), compared to normal skin (9.0 +/- 3.55) (P < 0.05). Melasma showed increased melanocyte numbers compared to normal skin, but it was not statistically significant (P > 0.05). CONCLUSION: We believe this computerized image analysis could be useful tool for diagnosis and comparison of interval changes in pigmentary diseases like melasma or lentigo senilis by quantifying melanin pigments or melanocytes in skin biopsy specimens. PMID- 11277955 TI - Prurigo in a patient with intestinal strongyloidiasis. PMID- 11277956 TI - Nevus lipomatosus cutaneous superficialis (Hoffmann-Zurhelle) with localized scleroderma like appearance. PMID- 11277957 TI - Intravenous pyogenic granuloma. PMID- 11277958 TI - Subacute cutaneous lupus erythematosus with pityriasis-like cutaneous manifestations. PMID- 11277959 TI - Poroid hidradenoma. PMID- 11277960 TI - Duration of improvement in psoriasis after treatment with tazarotene 0.1% gel plus clobetasol propionate 0.05% ointment: comparison of maintenance treatments. PMID- 11277962 TI - Once-daily desloratadine improves the signs and symptoms of chronic idiopathic urticaria: a randomized, double-blind, placebo-controlled study. AB - BACKGROUND: Chronic idiopathic urticaria (CIU) is the most common type of chronic urticaria, and pruritus is the most prominent symptom. Antihistamines are the first-line treatment for CIU. Sedation and anticholinergic adverse effects are often experienced with the first-generation antihistamines and there is a risk of cardiovascular adverse effects and drug interactions with some second-generation agents. Hence, new treatment options are needed. Desloratadine is a new, potent, nonsedating antihistamine that has an excellent cardiovascular safety profile. METHODS: This was a multicenter, randomized, double-blind, placebo-controlled study designed to determine the efficacy and safety of desloratadine in the treatment of moderate-to-severe CIU. A total of 190 patients, aged 12-79 years, with at least a 6-week history of CIU and who were currently experiencing a flare of at least moderate severity, were randomly assigned to therapy with desloratadine 5 mg or placebo once daily for 6 weeks. Twice daily, patients rated the severity of CIU symptoms (pruritus, number of hives, and size of largest hive), as well as the impact of CIU symptoms on sleep and daily activity. Patients and investigators jointly evaluated therapeutic response and overall condition. Safety evaluations included the incidence of treatment-emergent adverse events, discontinuations due to adverse events, and changes from baseline in vital signs, laboratory parameters, and ECG intervals. RESULTS: Desloratadine was superior to placebo in controlling pruritus and total symptoms after the first dose and maintained this superiority to the end of the study. Measures of sleep, daily activity, therapeutic response, and global CIU status were also significantly better with desloratadine after the first dose; these clinical benefits were also maintained throughout the 6-week study. No significant adverse events occured. CONCLUSIONS: Desloratadine 5 mg daily is a safe and effective treatment for CIU with significant benefits within 24 h and maintained through the treatment period. PMID- 11277961 TI - Treatment of larva migrans cutanea (creeping eruption): a comparison between albendazole and traditional therapy. AB - BACKGROUND: Creeping eruption (CE), which is characteristic of tropical and subtropical regions, is being increasingly frequently observed in Italy. The presence on the beaches of stray animals infected by nematodes of the Ancylostoma species favors contact between human skin and the larva-infested soil. MATERIALS AND METHODS: Our experience with 56 patients (13 cryotherapy, one thiabendazole together with cryotherapy, six thiabendazole, two albendazole with cryotherapy, and 34 albendazole) is described. RESULTS: A prompt and definitive cure was achieved in all 56 patients. The therapeutic effectiveness of the various methods used is therefore equivalent. CONCLUSIONS: We believe that albendazole should be considered the first choice for treatment. It is extremely well tolerated and patient compliance is good. PMID- 11277964 TI - An interesting outbreak of leishmaniasis in a family in the state of Rio de Janeiro. PMID- 11277963 TI - Subcutaneous dermatofibroma showing a depressed surface. PMID- 11277965 TI - The dark side. PMID- 11277966 TI - The fork'ed path to mitosis. AB - A concurrence of genomic, reverse genetic and biochemical approaches has cracked the decade-long enigma concerning the identity of the transcription factors that control gene expression at the G2/M transition in the budding yeast cell cycle. PMID- 11277968 TI - Interferon-gamma-induced changes in synaptic activity and AMPA receptor clustering in hippocampal cultures. AB - Extended release of interferon-gamma (IFN-gamma) in the nervous system during immunological and infectious conditions may trigger demyelinating disorders and cause disturbances in brain function. The aim of this study was to examine the effects of IFN-gamma on neuronal function in rat hippocampal cell cultures by using whole cell patch clamp analysis together with quantitative immunocytochemistry. Acute application of IFN-gamma to differentiated neurons in culture caused no immediate neurophysiological responses, but recordings after 48 h of incubation displayed an increase in frequency of AMPA receptor (AMPAR) mediated spontaneous excitatory postsynaptic currents (EPSCs). Quantitative immunocytochemistry for the AMPAR subunit GluR1 showed no alteration in receptor clustering at this time point. However, prolonged treatment with IFN-gamma for 2 weeks resulted in a significant reduction in AMPAR clustering on dendrites but no marked differences in EPSC frequency between treated neurons and controls could be observed. On the other hand, treatment of hippocampal neurons for 4 weeks, instituted at an immature stage (1 day in culture), caused a significant reduction in spontaneous EPSC frequency. These neurons developed with no overt alterations in dendritic arborization or in the appearance of dendritic spines as visualized by alpha-actinin immunocytochemistry. Nonetheless, there was a marked reduction in AMPAR clustering on dendrites. These observations show that a key immunomodulatory molecule, IFN-gamma, can cause long-term modifications of synaptic activity and perturb glutamate receptor clustering. PMID- 11277967 TI - Biocompatibility of silicon-based arrays of electrodes coupled to organotypic hippocampal brain slice cultures. AB - In this study we examined the passive biocompatibility of a three-dimensional microelectrode array (MEA), designed to be coupled to organotypic brain slice cultures for multisite recording of electrophysiological signals. Hippocampal (and corticostriatal) brain slices from 1-week-old (and newborn) rats were grown for 4-8 weeks on the perforated silicon chips with silicon nitride surfaces and 40 microm sized holes and compared with corresponding tissue slices grown on conventional semiporous membranes. In terms of preservation of the basic cellular and connective organization, as visualized by Nissl staining, Timm sulphide silver-staining, microtubule-associated protein 2 (MAP2) and glial fibrillary acidic protein (GFAP) immunostaining, the slice cultures grown on chips did not differ from conventionally grown slice cultures. Neither were there any signs of astrogliosis or neurodegeneration around the upper recording part of the 47 microm-high platinum-tip electrodes. Slice cultures grown on a separate set of chips with platinum instead of silicon nitride surfaces also displayed normal MAP2 and GFAP immunostaining. The width of the GFAP-rich zone (glia limitans) at the bottom surface of the slice cultures was the same ( approximately 20 microm) in cultures grown on chips with silicon nitride and platinum surfaces and on conventional insert membranes. The slice cultures grown on chips maintained a normal, subfield differentiated susceptibility to the glutamate receptor agonist N-methyl-D-aspartate (NMDA) and the neurotoxin trimethyltin (TMT), as demonstrated by the cellular uptake of propidium iodide (PI), which was used as a reproducible and quantifiable marker for neuronal degeneration. We conclude that organotypic brain slice cultures can grow on silicon-based three-dimensional microelectrode arrays and develop normally with display of normal subfield differentiated susceptibilities to known excito- and neurotoxins. From this it is anticipated that the set-up, designed for recording of electrophysiological parameters, can be used for long-term studies of defined neuronal networks and provide valuable information on both normal, neurotoxicological and neuropathological conditions. PMID- 11277969 TI - Modulation of oscillatory neural activities by cholinergic activation of interneurons in the olfactory center of a terrestrial slug. AB - The neurons in the procerebrum (PC) of the terrestrial slug Limax marginatus show regular oscillation of their membrane potential, and the oscillation has been implicated in olfactory processing. The neural mechanisms for the generation and modulation of the oscillation have been poorly understood. In the present work, we examined the ionic conductances evoked by acetylcholine (ACh) in the PC neurons and the effects of ACh application on the population activities of intrinsic and extrinsic neurons. The PC neurons are categorized into bursting neurons, which are putative local inhibitory neurons, and nonbursting neurons, which likely mediate the input and output of information in the PC. Bath application of ACh augmented the local field potential oscillation in the PC. Perforated patch recording from single PC neurons revealed that ACh has direct excitatory effects on bursting neurons, while it suppresses the activity of nonbursting neurons, possibly via augmented inhibitory synaptic input from bursting neurons. The correlation between the membrane potential of bursting neurons and the frequency of oscillation suggests that bursting neurons are the main determinant of the oscillation frequency. Application of ACh also resulted in a reduction of the oscillation amplitude in the olfactory nerve, suggesting that the frequency modulation in the oscillatory network could change the activities in the follower neurons. PMID- 11277970 TI - Effect of LPS on the permeability of the blood-brain barrier to insulin. AB - Insulin has emerged as an important neuropeptide. Central actions of insulin appear to oppose those in the periphery. Insulin is transported across the blood brain barrier (BBB) by a saturable transport system. The permeability of the BBB to insulin is altered by various events, but no studies exist that have examined the permeability of the BBB to insulin during infection or inflammation, states which can induce peripheral insulin resistance. We looked at the effects of lipopolysaccharide (LPS), a bacterial endotoxin and a powerful cytokine releaser, on the permeability of the BBB to human insulin in CD-1 mice. Intraperitoneal injections of LPS significantly increased the uptake by the brain of 131I-insulin and disrupted the BBB to 125I-albumin. After subtraction of the brain/serum ratio for 125I-albumin, brain/serum ratios for insulin were increased: 10.38 +/- 0.70 microl/g (LPS) vs. 3.62 +/- 0.27 microl/g (no LPS), P<0.0001, showing that LPS increased the uptake of insulin independent of BBB disruption. This increase in insulin uptake was due to enhanced saturable transport. Pretreatment with indomethacin 10 min before LPS injections enhanced BBB disruption, but not insulin transport. Pretreatment with the nitric oxide (NO) synthase inhibitor aminoguanidine had no effect on insulin or albumin uptake, but pretreatment with NG-nitro-L-arginine methyl ester (L-NAME) enhanced insulin transport, but not BBB disruption. We conclude that LPS increases the saturable transport of insulin across the BBB independent of disruption and prostaglandins with potentiation by NO inhibition. Such increased transport could potentiate the central effects of insulin and so contribute to the peripheral insulin resistance seen with infection and inflammation. PMID- 11277971 TI - Limbic dopaminergic adaptation to a stressful stimulus in a rat model of depression. AB - The dopaminergic mesolimbic system has a key role in motivation and reward, and stressful stimuli appear to alter its functionality. Since stress is considered to be one of the primary factors that mediate the expression of depressive behavior, dopamine and its metabolites in the nucleus accumbens of control and Flinders Sensitive Line rats, an animal model of depression, were examined prior to and after a forced swim test. In both types of rats, the levels of dopamine metabolites markedly decreased after the forced swimming, albeit to different extents. In contrast, 60 min after the swim test, dopamine levels were elevated only in the control rats. The accumbal dopaminergic activity is discussed in relation to the behavior of 'depressed' and normal rat lines subjected to a stressful event. PMID- 11277972 TI - Enriched environment during development is protective against lead-induced neurotoxicity. AB - It is known that children of lower socioeconomic status have a disproportionately higher risk of being exposed to lead and have a more negative outcome from that exposure than children who are raised under more fortunate circumstances. Yet, little is known about how environmental factors may influence the injurious effects on the brain of a neurotoxin such as lead. The present study used an animal model of lead poisoning to examine the extent to which different environmental milieus may modify the effects of lead on the developing brain. Young rats were raised in either enriched or impoverished environments and drank either distilled water or water with lead. Lead-exposed rats raised in the impoverished environment had spatial learning deficits and significantly decreased neurotrophic factor gene expression in the hippocampus. In contrast, the animals raised in the enriched environment were significantly protected against the behavioral and neurochemical toxicity of lead. These results demonstrate that impoverished environment may accentuate while enriched environment may ameliorate neurobehavioral and neurochemical toxicity from developmental lead exposure. PMID- 11277973 TI - Neuroprotective effects of GDNF against 6-OHDA in young and aged rats. AB - In young adult rats, glial cell line-derived neurotrophic factor (GDNF) can completely protect against 6-hydroxydopamine-induced loss of nigral dopamine neurons when administered 6 h prior to the 6-hydroxydopamine. The present study was undertaken to determine if GDNF would provide similar protective effects in aged rats. Male, Fischer 344 x Brown Norway hybrid rats of 3, 18 and 24 months of age were given an intranigral injection of GDNF or vehicle followed 6 h later with an intranigral injection of 6-hydroxydopamine. Nigral dopamine neuron cell survival, and striatal and nigral dopamine and DOPAC levels, were evaluated 2 weeks after the lesions. In vehicle treated animals cell survival on the lesioned side ranged from 15 to 27%. GDNF promoted significant cell survival in the nigra of all three age groups; however, the percent survival was lowest in the 24-month old animals (85% at 3 months, 75% at 18 months, 56% at 24 months). Similarly, dopamine levels in the striatum and substantia nigra on the lesioned side remained significantly greater in the GDNF treated animals compared to the vehicle treated animals. As with the cell survival experiment, the protective effects of GDNF on dopamine levels were less in the 24-month-old animals. GDNF pretreatment also protected against 6-hydroxydopamine-induced reductions in striatal DOPAC levels in all age groups. Overall, these results indicate that GDNF can protect nigrostriatal dopamine neurons against the effects of 6 hydroxydopamine in aged as well as young adult rats. However, the extent of protection is less in the aged (24-month-old) animals. PMID- 11277974 TI - Sciatic and vagal sensory inputs converge onto non-baroreceptive neurones of the rostral ventrolateral medulla. AB - Previous data suggested that somatic and vagal sensory afferent inputs may converge in the rostral ventrolateral medulla oblongata (RVLM). The aim of the present study was to establish the existence of convergence between inputs mediated via the cervical vagus and contralateral sciatic nerves using in vivo intracellular recordings. The majority of RVLM neurones that received input from the vagus or the sciatic nerves also responded to stimulation of the other nerve. In 72% of the neurones the response was excitation or inhibition to stimulation of both nerves, respectively. The most frequent response type was a short excitation in response to stimulation of both nerves. Only 8% of the neurones exhibited a visible response to stimulation of the aortic depressor nerve. The results provided experimental evidence that non-baroreceptive neurones of the RVLM are involved in coordination of somatic and visceral sensory inputs. PMID- 11277975 TI - Early release of cytochrome C and activation of caspase-3 in hyperglycemic rats subjected to transient forebrain ischemia. AB - The mechanisms underlying the aggravating effect of hyperglycemia on brain damage are still elusive. The present study was designed to test our hypothesis that hyperglycemia-mediated damage is caused by mitochondrial dysfunction with mitochondrial release of cytochrome c (cyt c) to the cytoplasm, which leads to activation of caspase-3, the executioner of cell death. We induced 15 min of forebrain ischemia, followed by 0.5, 1, and 3 h of recirculation in sham, normoglycemic and hyperglycemic rats. Release of cyt c was observed in the neocortex and CA3 in hyperglycemic rats after only 0.5 h of reperfusion, when no obvious neuronal damage was observed. The release of cyt c persisted after 1 and 3 h of reperfusion. Activation of caspase-3 was observed after 1 and 3 h of recovery in hyperglycemic animals. No cyt c release or caspase-3 activation was observed in sham-operated controls while a mild increase of cyt c was observed in normoglycemic ischemic animals after 1 and 3 h of reperfusion. The findings that there is caspase activation and cyt c relocation support a notion that the biochemical changes that constitute programmed cell death occur after ischemia and contribute, at least in part, to hyperglycemia-aggravated ischemic neuronal death. PMID- 11277976 TI - Identification of 5-HT(3A) and 5-HT(3B) receptor subunits in mammalian retinae: potential pre-synaptic modulators of photoreceptors. AB - Although serotonin (5-HT) is found in the mammalian retina only at low levels, considerable evidence suggests that it plays a role in visual processing. Pharmacological experiments indicate that numerous receptors for 5-HT are present in the mammalian retina. One of these is the ionotropic 5-HT(3) receptor. So far, two subunits for this receptor have been identified in the nervous system, 5 HT(3A) and 5-HT(3B). Co-expression of these subunits in Xenopus oocytes is sufficient to reconstitute native 5-HT(3) receptor properties. Thus, it is believed that a native neuronal 5-HT(3) receptor is multimeric similar to the related acetylcholine receptor family. To determine whether this receptor is expressed in the mammalian retina, we first performed reverse transcription polymerase chain reaction and first demonstrated the presence of transcripts for both the 5-HT(3A) and 5-HT(3B) receptor subunits. Then using a well-characterized polyclonal antiserum against the 5-HT(3A) receptor subunit, we demonstrated 5 HT(3A) receptor immunoreactivity (IR) in the rabbit, rat, and human retina. This IR was localized specifically to the rod photoreceptor terminals in all three species, suggesting that this receptor may modulate the rod signaling pathway by controlling the output at the rod terminals. PMID- 11277977 TI - Interleukin-6 promotes post-traumatic healing in the central nervous system. AB - The central nervous system (CNS) is an immune-privileged site where the role of immune cells and mediators in traumatic brain injury is poorly understood. Previously we have demonstrated that interleukin (IL)-6, a cytokine that acts on a wide range of tissues influencing cell growth and differentiation, is an agonist for vascular endothelial growth factor (VEGF), in in vitro vascularization assays for brain microvessel endothelial cells. In this present work we focus on the role of IL-6 in promoting tissue repair in the CNS in vivo. An aseptic cerebral injury (ACI) was created in the right parietal cortex, using both wild type (C57Bl/6J) and IL-6-deficient (C57Bl/6J-IL-6-/-) mice to study the consequences of the absence of IL-6 on the pathology of brain injuries. We monitored the immediate, early, and late responses to this traumatic injury by characterizing several histologic features in the CNS at days 1, 4, 7 and 14 following injury. Acellular necrosis, cellular infiltration, and re vascularization were characterized in the injured tissues, and each of these histologic features was individually graded and totaled to assign a healing index. IL-6-deficient mice were found to have a comparatively slower rate of recovery and healing. Furthermore, fluorescein isothiocyanate (FITC)-dextran intravenous injection demonstrated leaky vessels in IL-6-deficient but not in wild type animals following ACI. Additionally, chronic expression of IL-6 in the CNS using transgenic GFAP-IL-6 mice resulted in more rapid healing following ACI. The accelerated tissue repair in GFAP-IL-6 transgenic animals is primarily due to extensive re-vascularization as detected by endothelial cell markers. Combined, this data suggests an important role of IL-6 in tissue repair processes following traumatic injury in the CNS. PMID- 11277978 TI - Haloperidol downregulates phospholipase A(2) signaling in rat basal ganglia circuits. AB - Our laboratory has developed an in vivo method to quantitatively evaluate phospholipase A(2) (PLA(2))-mediated signal transduction in brain regions of rodents. In this method, quantitative autoradiography is used to identify brain uptake of intravenously injected, radiolabeled arachidonic acid ([3H]AA). Dopamine D(2) receptors are coupled to G-proteins that activate PLA(2), releasing AA from the stereospecifically numbered (sn) 2 position of phospholipids, and regional [3H]AA uptake is proportional to the rate of release. In the present experiment, the D(2) antagonist haloperidol (1.0 mg/kg i.p.) or the drug vehicle was administered to male adult rats for 21 days. Rats were infused 3 days later with 1.75 mCi/kg [3H]AA (i.v.), anesthetized and decapitated 20 min after infusion onset, and brains were processed for quantitative autoradiography. Chronic haloperidol significantly decreased [3H]AA incorporation in two primary dopaminergic basal ganglia-frontal cortex circuits, the mesocorticolimbic and nigrostriatal systems, while insignificant changes in AA incorporation were noted in other brain regions. These results suggest that one mechanism by which haloperidol exerts its effect is by downregulating D(2)-mediated PLA(2) signaling involving AA release in basal ganglia-frontal cortex circuitry. PMID- 11277979 TI - Energy fuel utilization by fetal versus young rabbit brain: a 13C MRS isotopomer analysis of [U-(13)C]glucose metabolites. AB - The principle substrate for brain metabolism is glucose, which provides both energy and the carbon skeletons of glutamate and glutamine, via the TCA cycle. The existence of two distinct cerebral metabolic compartments, neurons and glia, involved in glutamate and glutamine synthesis, respectively, is a widely accepted concept. In previous work, the relative glucose flux via pyruvate dehydrogenase (PDH) and pyruvate carboxylase (PC) in adult rabbit brain, using 13C NMR isotopomer analysis of glutamate and glutamine, was quantified. In this work, manifestation of cerebral compartmentation in the near-term fetal rabbit was investigated, using the above approach. Following infusion of [U-13C]glucose into maternal circulation (1 mg/kg per min) for 60-70 min, fetal brains were excised and brain extracts were studied by 13C NMR. The labelling patterns of fetal cerebral metabolites differed from those observed in the young adult brain. The most significant differences were found for glutamine labelling patterns. We suggested that these differences are a result of increased utilization of non labeled fuels, mainly beta-hydroxybutyrate (beta-HBA) in the glia, the site of glutamine synthesis. In addition, we have shown that acute exposure to elevated beta-HBA levels leads to increased uptake, but not utilization, into the fetal rabbit brain; no increase in uptake is observed in the adult brain. We have also demonstrated that during short-term starvation, although no changes are detected in plasma and cerebral glucose levels in the fetal and young adult brain, amino acid levels and energy metabolism are altered in the young adult brain. PMID- 11277980 TI - Differential expression of cellular prion protein in mouse brain as detected with multiple anti-PrP monoclonal antibodies. AB - The normal cellular prion protein (PrP(C)) plays an essential role in the development of prion diseases. Indirect evidence has suggested that different PrP(C) glycoforms may be expressed in different brain regions and perform distinct functions. However, due to a lack of monoclonal antibodies (Mabs) that are specific for mouse PrP(C), the expression of PrP(C) in the mouse brain has not been studied in great detail. We used Mabs specific for either the N-terminus or the C-terminus of the mouse PrP(C) to study its expression in the mouse brain by immunoblotting and immunohistochemistry. Immunoblotting studies demonstrated that the expression of PrP(C) differed quantitatively as well as qualitatively in different regions of the brain. The anti-C-terminus Mabs reacted with all three molecular weight bands of PrP(C); the anti-N-terminus Mabs only reacted with the 39-42 kDa PrP(C). The results from immunohistochemical staining revealed the spatial distribution of PrP(C) in the mouse brain, which were consistent with that from immunoblotting. Although expression of PrP(C) has been reported to be required for long-term survival of Purkinje cells, we were unable to detect PrP(C) in the Purkinje cell layer in the cerebellum with multiple anti-PrP Mabs. Our findings suggest that PrP(C) variants, i.e. various glycoforms and truncated forms, might be specifically expressed in different regions of mouse brain and might have different functions. PMID- 11277981 TI - Glucocorticoids and serotonin alter glucocorticoid receptor (GR) but not mineralocorticoid receptor (MR) mRNA levels in fetal mouse hippocampal neurons, in vitro. AB - Studies utilizing rats and guinea pigs have demonstrated that the hypothalamo pituitary-adrenal (HPA) axis can be programmed by glucocorticoids during fetal life. Such programming is believed to occur, at least partially, at the level of hippocampal glucocorticoid receptors (GR) and mineralocorticoid receptors (MR). Studies have also demonstrated that serotonin up regulates GR levels within the developing hippocampus. However, the cell type in which these changes take place has not been determined. We hypothesized that dexamethasone, corticosterone and serotonin exposure modify GR and MR mRNA levels in fetal mouse hippocampal cultures, and that these effects are confined to neurons. Cultures were derived from CD1 mouse fetuses on day 18 of gestation (n=8 dams). Fetal hippocampi were dissected, then mechanically and chemically dispersed. Cultures were exposed to dexamethasone, corticosterone or serotonin (1-100 nM) for 4 days. Levels of GR and MR mRNA were examined by in situ hybridization and high-resolution silver emulsion autoradiography. Four days exposure to dexamethasone or corticosterone (10 or 100 nM) decreased levels of GR mRNA within neurons. There was no significant change in MR mRNA in either experiment. Exposure to serotonin (100 nM) significantly increased expression of GR mRNA in hippocampal neurons. MR mRNA levels were unaffected by serotonin treatment. Dexamethasone, corticosterone or serotonin exposure did not alter expression of GR mRNA within glial cells. We conclude that synthetic and endogenous glucocorticoids, as well as serotonin, can influence neuronal levels of GR mRNA during hippocampal development. However, whether these effects are permanent remains to be determined. PMID- 11277982 TI - Interleukin-1beta induces expression of neuropeptide Y in primary astrocyte cultures in a cytokine-specific manner: induction in human but not rat astrocytes. AB - Previous studies have demonstrated that astrocyte cultures express neuropeptide Y (NPY) in a regulated manner, namely, phorbol ester leads to an increase in proNPY mRNA and NPY production. In this respect, the behavior of astrocytes derived from the human fetal or rat neonatal brain is similar (Regul. Pept. 75 (1998) 293). Since astrocytes can be exposed to high levels of IL-1beta, we addressed the question: Does IL-1beta regulate NPY expression by the astrocytes? Primary astrocytes derived from the human fetal or rat neonatal cortex were cultured in serum-free medium. IL-1beta, but not IL-6 or TNF-alpha, led to an increase in NPY production dose-dependently. IL-1beta action manifested in the human but not in the rat astrocytes and it was completely abolished by IL-1 receptor antagonist. The responsiveness to IL-1beta did not diminish upon sub-culture of the astrocytes (five passages). In addition, IL-1beta led to an increase in the abundance of proNPY-mRNA, which was preceded by a rapid and transient increase in cFos-mRNA and a rapid and sustained increase in JunB-mRNA. In contrast to cFos/JunB, IL-1beta did not alter the abundance of cJun-mRNA. In summary, we demonstrate that IL-1beta induction of NPY expression in astrocytes is species- and cytokine-specific and that IL-1 receptor is involved. Moreover, induction of NPY expression is preceded by a rapid increase in the expression of two transcription factors (cFos, JunB) that have been previously (Oncogene 9 (1994) 2369; J. Neurochem. 70 (1998) 1887) implicated in transcriptional regulation of the human NPY gene. PMID- 11277983 TI - Expression of mutant amyloid precursor proteins decreases adhesion and delays differentiation of Hep-1 cells. AB - The amyloid precursor protein (APP) is a type I integral membrane protein and is processed to generate several intra-cellular and secreted fragments. The physiological role of APP and its processed fragments is unclear. Several mutations have been discovered in APP, which are causative of early-onset, familial, neurological disease, including Alzheimer's disease (FAD). These mutations alter the processing of APP and lead to excess production and extra cellular deposition of A-beta peptide (Abeta). We have examined the role of APP in a cell culture model of endothelial cell function. The endothelial cell line, Hep-1, was stably transfected with wild-type (wt) and FAD mutant forms of APP (mAPP). Secretion of sAPPalpha was reduced in cell lines over-expressing mAPP when these cells were grown on several different substrates. Levels of secreted Abeta were increased as measured by ELISA in the mutant cell lines. Cell adhesion to laminin-, fibronectin-, collagen I-, and collagen IV-coated culture flasks was reduced in all mAPP-expressing cell lines, while in lines over-expressing wt-APP, adhesiveness was slightly increased. Cell lines over-expressing mAPP differentiated more slowly into capillary network-like structures on Matrigel than those expressing wt-APP. No differences were detected among all cell lines in a migration/invasion assay. The results suggest that APP may have a role in cell adhesiveness and maturation of endothelial cells into capillary-like networks. The reduction in adhesion and differentiation in mutant cell lines may be due to reduced amounts of sAPPalpha released into the culture media or toxic effects of increased extracellular Abeta. PMID- 11277984 TI - Fourth ventricular injection of CART peptide inhibits short-term sucrose intake in rats. AB - Expression of CART (cocaine-amphetamine-regulated transcript) in the rat hypothalamus is modulated by nutritional status, and injection of synthetic CART peptide into the forebrain ventricular system suppresses food intake, indicating a possible role in hypothalamic control of energy homeostasis. Its recent identification in cell bodies and central terminals of vagal afferent neurons additionally suggests a role in brainstem mechanisms of meal termination and satiety. We demonstrate here that CART[55-102] (0.2 nmol) suppresses short-term sucrose intake and overnight chow intake in non-food-deprived rats even more when delivered into the fourth ventricle as compared to the lateral ventricle. At the threshold dose (0.02-0.08 nmol) no readily noticeable motor impairments were observed. The results are consistent, but do not prove a site of action within the brainstem, possibly in mediating vagal satiety signals at the level of the NTS. PMID- 11277985 TI - Oligodendrocyte killing by quinolinic acid in vitro. AB - Quinolinic acid, which is produced by macrophages and microglia, can kill neurons in vivo and in vitro. To test whether quinolinic acid is toxic to oligodendrocytes, glial cells cultured from the brains of 2-day-old rats were incubated with quinolinic acid at concentrations known to kill neurons. The cells were then fixed and immunostained with MAbO4 to mark immature and mature oligodendrocytes and anti-myelin basic protein (MBP) to mark mature oligodendrocytes. The data indicated up to 54% reductions in the numbers of O4 positive cells in cultures after incubation with quinolinic acid. Apoptosis of O4 positive cells began during the first 6 h, and some of the apoptotic cells became fragmented. Further apoptosis, and clumping of dead MBP-positive oligodendrocytes, occurred during longer incubation with quinolinic acid. Thus, quinolinic acid arising from macrophages and microglia during autoimmune disease may take part in a mechanism of oligodendrocyte injury and killing. PMID- 11277986 TI - Platelet secretion of beta-amyloid is increased in hypercholesterolaemia. AB - Tissue accumulation of the cytotoxic beta-amyloid peptide (Abeta) occurs in Alzheimer's disease (AD), one possible source being the platelet. AD and cardiovascular disease may share some risk factors, including hypercholesterolaemia which is associated with increased platelet activity. We examined platelet Abeta release under resting and collagen-stimulated conditions in normocholesterolaemic and hypercholesterolaemic individuals. Resting platelet Abeta efflux was greater in hypercholesterolaemics than in normocholesterolaemics. Collagen-stimulated Abeta release was concentration dependent and increased in hypercholesterolaemics. Resting Abeta release correlated positively with plasma total cholesterol and low-density lipoprotein (LDL) cholesterol, and inversely with platelet count. These data indicate that abnormal platelet Abeta release occurs in hypercholesterolaemia. PMID- 11277987 TI - Potent, hydroxyl radical-scavenging effect of apomorphine with iron and dopamine perfusion in rat striatum. AB - In dopaminergic neurons, free radicals are likely produced via dopamine metabolism by monoamine oxidase or via its auto-oxidation, a process facilitated by transition metals. In this study we examined the effect and possible mechanisms of apomorphine to reduce iron- and dopamine-induced 2,3 dihydroxybenzoic acid (2,3-DHBA) formation by microdialysis. We have shown that (1) FeSO(4).7H(2)O reduced both the release of dopamine and the output of dihydroxyphenylacetic acid (DOPAC); (2) apomorphine may reduce FeSO(4).7H(2)O induced increases of 2,3-DHBA formation; (3) apomorphine has substantially reduced DOPAC output in early phase and blocked dopamine-induced increase of 2,3 DHBA levels. It is concluded that apomorphine is a potent hydroxyl radical scavenger in vivo, especially for the dopamine formation. PMID- 11277988 TI - Involvement of prostanoid release in the mediation of UTP-induced cerebrovascular contraction in the rat. AB - The interaction between uridine-5'-triphosphate (UTP) and prostanoids was studied in isolated rat middle cerebral arteries (MCAs). The strong contractions in MCA segments induced by UTP were weakened significantly by indomethacin and more markedly by the thromboxane receptor antagonist ICI 192605. Thromboxane A(2) (TXA(2)) release by MCAs was below the detection limit of the chemiluminescence enzyme immunoassay, but increased TXA(2) formation was detected in basilar arteries in the presence of UTP. Prostacyclin (PGI(2)) formation by MCAs also increased in the presence of UTP. These results suggest that UTP stimulates the release of both TXA(2) and PGI(2) from the rat MCA but the vascular effect of TXA(2) is dominant. PMID- 11277989 TI - Antinociception induced by PAG-microinjected dipyrone (metamizol) in rats: involvement of spinal endogenous opioids. AB - Dipyrone microinjection into the periaqueductal gray matter (PAG) elicits antinociception in rats by activating endogenous opioidergic circuits in PAG and the rostral ventromedial medulla. We have now found that endogenous opioids in the spinal cord are also involved. Responses of dorsal spinal neurons to noxious stimulation of a hindpaw were diminished (to 38-44%) by dipyrone microinjection (100 microg/0.5 microl) into the PAG. This was abolished by application of naloxone (50 microg/50 microl) to the spinal cord. The fact that dipyrone, a non opioid analgesic, activates opioidergic circuits may be clinically important. PMID- 11277990 TI - Decreased CRH mRNA expression in the fetal guinea pig hypothalamus following maternal nutrient restriction. AB - The regulation of corticotropin-releasing hormone (CRH) mRNA expression following maternal nutrient restriction was examined in the fetal hypothalamus. Pregnant guinea pigs were food restricted for 48 h or fed normally during late gestation. After nutrient restriction, CRH mRNA levels in the hypothalamic paraventricular nucleus of the fetus were determined using in situ hybridization and were found to be significantly decreased (P<0.0001) compared to controls. In conclusion, we have successfully sequenced the coding sequence of the guinea pig CRH gene, and have shown that a short period (48 h) of maternal nutrient restriction inhibits CRH mRNA expression in the fetal hypothalamus. PMID- 11277991 TI - Low intensity spinal cord stimulation may induce cutaneous vasodilation via CGRP release. AB - This study examined whether spinal cord stimulation (SCS) at intensities below motor threshold (MT) produces cutaneous vasodilation through sympathetic inhibition and/or antidromic activation of sensory fibers. SCS was applied to anesthetized rats with stimulus parameters used clinically, i.e. 50 Hz, 0.2 ms and stimulus intensities at 30, 60 or 90% of MT. SCS-induced vasodilation was not attenuated by hexamethonium, an autonomic ganglion blocking agent, but was abolished by CGRP-(8-37), an antagonist of the calcitonin gene-related peptide (CGRP) receptor. We concluded that SCS-induced vasodilation under the conditions of this study was mediated by peripheral release of CGRP via antidromic activation of sensory fibers. PMID- 11277992 TI - Basic fibroblast growth factor and fibroblast growth factor receptors in adult olfactory epithelium. AB - Basic fibroblast growth factor (FGF2) stimulates proliferation of the globose basal cells, the neuronal precursor in the olfactory epithelium. The present study investigates the expression of basic fibroblast growth factor and fibroblast growth factor receptors in the adult olfactory epithelium. FGF2 immunoreactivity was expressed widely in the olfactory epithelium, with the highest density of immunoreactivity in the supporting cells. In contrast, most cells in the epithelium expressed FGF2 mRNA. Fibroblast growth factor receptor-1 (FGFr1) immunoreactivity was densest in the basal cell and neuronal layers of the olfactory epithelium and on the apical surface of supporting cells. In the lamina propria FGF2 immunoreactivity and mRNA were densest in cells close to the olfactory nerve bundles. FGFr1 immunoreactivity was heaviest on the olfactory ensheathing cells. Using reverse transcriptase-polymerase chain reaction analysis, the olfactory epithelium was shown to express only three receptor splice variants, including one (FGFr1c) with which basic fibroblast growth factor has high affinity. Other receptor splice variants were present in the lamina propria. Taken together, these observations indicate endogenous sources of FGF2 within the olfactory epithelium and lamina propria and suggest autocrine and paracrine pathways via which FGF2 might regulate olfactory neurogenesis. The observation of only three receptor splice variants in the olfactory epithelium limits the members of the fibroblast growth factor family which could act in the olfactory epithelium. The widespread distribution of receptors suggests that fibroblast growth factors may have roles other than proliferation of globose basal cells. PMID- 11277993 TI - Retrograde labelling of mitral/tufted cells in the mouse accessory olfactory bulb following local injections of the lipophilic tracer DiI into the vomeronasal amygdala. AB - It has recently become apparent that there are two classes of vomeronasal receptor neurons that project to functionally separate anterior and posterior sub regions of the mammalian accessory olfactory bulb. However, anterograde tracing of the projections from these sub-regions, in the mouse, has revealed that the processing pathways are not segregated at the level of the vomeronasal amygdala. Both sub-regions have overlapping projections to the superficial lamina of the medial and posterior medial cortical nuclei of the amygdala. However, differential projections have been found in the opossum, in which only the posterior sub-region projects to the deeper laminae of the medial amygdala. Therefore, there may be species differences in these projections that are important for the control of reproductive behaviour. This study used an alternative approach of retrogradely tracing mitral/tufted cell projections from different nuclei of the vomeronasal amygdala back to the accessory olfactory bulb of mice. Local injections of the lipophilic tracer DiI were made into the antero dorsal and postero-ventral divisions of the medial amygdala, and into the postero medial cortical amygdala. In each case, provided the DiI affected the superficial lamina Ia, labelled mitral/tufted cells were found distributed throughout the anterior-posterior extent of the accessory olfactory bulb. These results confirm that mitral/tufted cells of the anterior and posterior sub-regions of the accessory olfactory bulb project to both the medial and postero-medial cortical nuclei of the amygdala. There was no evidence for differential projections from the anterior and posterior sub-regions accessory olfactory bulb in mice, as has been reported to occur in other species. PMID- 11277994 TI - Mutation Arg336 to Lys in Saccharomyces cerevisiae phosphoenolpyruvate carboxykinase originates an enzyme with increased oxaloacetate decarboxylase activity. AB - Saccharomyces cerevisiae phosphoenolpyruvate (PEP) carboxykinase catalyzes one of the first reactions in the biosynthesis of carbohydrates. Apart from the physiologically important reaction, the enzyme also presents low oxaloacetate decarboxylase and pyruvate kinase-like activities. Data from the crystalline structure of homologous Escherichia coli PEP carboxykinase suggest that Arg(333) may be involved in stabilization of enolpyruvate, a postulated reaction intermediate. In this work, the equivalent Arg(336) from the S. cerevisiae enzyme was changed to Lys or Gln. Kinetic analyses of the varied enzymes showed that a positive charge at position 336 is critical for catalysis of the main reaction, and further suggested different rate limiting steps for the main reaction and the secondary activities. The Arg336Lys altered enzyme showed increased oxaloacetate decarboxylase activity and developed the ability to catalyze pyruvate enolization. These last results support the proposal that enolpyruvate is an intermediate in the PEP carboxykinase reaction and suggest that in the Arg336Lys PEP carboxykinase a proton donor group has appeared. PMID- 11277995 TI - TNF-alpha inhibits UCP-1 expression in brown adipocytes via ERKs. Opposite effect of p38MAPK. AB - Tumor necrosis factor-alpha (TNF-alpha) activates extracellular-regulated kinases (ERKs) and p38 mitogen-activated protein kinase (p38MAPK), and inhibits the expression of uncoupling protein-1 (UCP-1) and adipocyte-specific genes in rat fetal brown adipocytes. MEK inhibition with PD98059 abolished the inhibitory effect of TNF-alpha on UCP-1, but not on adipogenic genes. In contrast, inhibition of p38MAPK with SB203580 potentiated the negative effect of TNF-alpha on UCP-1 and adipogenic genes. The inhibitory action of TNF-alpha was partially correlated with changes in C/EBPalpha and beta protein levels and in their DNA binding activity, suggesting a role for these transcription factors. However, other transcription factors might explain the different regulation of UCP-1 and adipogenic genes by ERKs. PMID- 11277996 TI - Effect of divalent cations on the ATPase activity of Escherichia coli SecA. AB - It was found that Ca(2+) stimulates the intrinsic SecA ATPase activity in the absence as well as in the presence of liposome. On the other hand, Mg(2+), the general cofactor for ATPase, did not affect the intrinsic SecA ATPase but reduced the portion of ATPase activity enhanced by Ca(2+). The enhancement of SecA ATPase activity correlated well with the increase in 8-anilino-1-naphthalene-sulfonic acid binding of SecA, suggesting that increased exposure of hydrophobic residues stimulates the enzyme activity. PMID- 11277997 TI - An internal region of the RpoH heat shock transcription factor is critical for rapid degradation by the FtsH protease. AB - The proteolysis of regulatory proteins plays an important role in the control of gene expression. The Escherichia coli heat shock sigma factor RpoH (sigma(32)) is highly unstable. Its instability is determined by interactions with the DnaK chaperone machine, RNA polymerase and the ATP-dependent protease FtsH. Bradyrhizobium japonicum expresses three RpoH proteins of which RpoH(1) is highly stable. To determine which regions of E. coli RpoH determine protein lability, we generated a number of truncated versions and hybrid proteins. Truncation of N terminal amino acids had no, and deletion of C-terminal amino acids only a minor effect on stability of RpoH. A major determinant of RpoH lability was mapped to a region of about 85 amino acids (residues 36-122) roughly comprising the sigma factor region 2. This is the first demonstration of an internal RpoH region being responsible for FtsH-mediated degradation. PMID- 11277998 TI - Pro-atherogenic factors induce telomerase inactivation in endothelial cells through an Akt-dependent mechanism. AB - Advanced aging may contribute to impairment of angiogenesis and development of vascular diseases. Telomerase was shown to delay endothelial cell (EC) senescence. Therefore, we determined the regulation of telomerase activity in EC. Inhibition of phosphoinositol 3-kinase (PI3K) suppressed telomerase activity, whereas inhibitors directed against ERK1/2 or protein kinase C had no effect. Dominant negative Akt significantly reduced telomerase activity. Moreover, pro atherogenic stimuli such as oxidized low density lipoprotein led to an inactivation of Akt and diminished telomerase activity. Thus, the PI3K/Akt pathway plays an important role in the regulation of telomerase activity. Pro atherosclerotic factors impair telomerase activity and thereby may promote EC aging. PMID- 11277999 TI - Hypericin photo-induced apoptosis involves the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and activation of caspase-8. AB - Hypericin (HYP) is a photosensitizing pigment from Hypericum perforatum that displays cytotoxic effects in neoplastic cell lines. Therefore, HYP is presently under consideration as a new anticancer drug in photodynamic therapy. Here, we investigated the mechanism of action of HYP photo-induced apoptosis of Jurkat cells compared to the cytostatic drug paclitaxel (PXL). Both photoactivated HYP and PXL similarly increased the activity of caspase-8 and caspase-3, and drug induced apoptosis of Jurkat cells was completely blocked by inhibitors of caspase 8 (Z-IETD-FMK) and caspase-3 (Z-DEVD-FMK). The involvement of death receptors was analyzed using neutralizing monoclonal antibodies against Fas (SM1/23), FasL (NOK 2) and TNF-R1 (MAB225), and a polyclonal rabbit anti-human TNF-related apoptosis inducing ligand (TRAIL) antiserum. TRAIL antibody blocked TRAIL-induced and HYP photo-induced, but not PXL-induced apoptosis of Jurkat cells. In contrast, PXL induced, but not HYP-induced apoptosis was blocked by the SM1/23 and NOK-2 antibodies. Anti-TNF-R1 antibody had no effect. These findings suggest that HYP photo-induced apoptosis of Jurkat cells is mediated in part by the TRAIL/TRAIL receptor system and subsequent activation of upstream caspases. PMID- 11278000 TI - Nucleotide-induced conformational changes in the human multidrug resistance protein MRP1 are related to the capacity of chemotherapeutic drugs to accumulate or not in resistant cells. AB - Intracellular accumulation of anthracycline derivatives was measured in a human embryonic kidney cell line (HEK) and a resistant subline (HEK/multidrug resistance protein (MRP1)) overexpressing MRP1 at the plasma membrane surface. Two compounds (daunorubicin and doxorubicin) were rejected outside the multidrug resistant cells. On the contrary, three compounds (4'-deoxy-4'-iodo-doxorubicin, 4-demethoxy-daunorubicin and 3'-(3-methoxymorpholino)doxorubicin) accumulated equally within sensitive HEK cells and resistant HEK/MRP1 cells. Our main objective here was to characterize the MRP1 conformational changes mediated by the binding of these anthracycline derivatives and to determine whether these conformational changes are related to MRP1-mediated drug transport. MRP1 was reconstituted in lipid vesicles as previously described [Manciu, L., Chang, X.B., Riordan, J.R. and Ruysschaert, J.-M. (2000) Biochemistry 39, 13026-13033]. The reconstituted protein was shown to conserve its ATPase and drug transport activity. Acrylamide quenching of Trp fluorescence was used to monitor drug dependent conformational changes. Binding of drugs (4-demethoxy-daunorubicin and 3'-(3-methoxymorpholino)doxorubicin) which accumulate in resistant cells immobilizes MRP1 in a conformational state that is insensitive to ATP binding whereas drugs rejected outside the resistant cells (daunorubicin, doxorubicin) favor a conformational change which may be a required step in the transport process. PMID- 11278001 TI - Development and application of bioluminescent Caenorhabditis elegans as multicellular eukaryotic biosensors. AB - We describe a novel approach to assess toxicity to the free-living nematode Caenorhabditis elegans that relies on the ability of firefly luciferase to report on endogenous ATP levels. We have constructed bioluminescent C. elegans with the luc gene under control of a constitutive promoter. Light reduction was observed in response to increasing temperature, concentrations of copper, lead and 3,5 dichlorophenol. This was due to increased mortality coupled with decreased metabolic activity in the surviving animals. The light emitted by the transgenic nematodes gave a rapid, real-time indication of metabolic status. This forms the basis of rapid and biologically relevant toxicity tests. PMID- 11278002 TI - Effects of FTDP-17 mutations on the in vitro phosphorylation of tau by glycogen synthase kinase 3beta identified by mass spectrometry demonstrate certain mutations exert long-range conformational changes. AB - In vitro phosphorylation of recombinant wild-type 2N4R tau and FTDP-17 exonic mutant forms P301L, V337M and R406W by glycogen synthase kinase 3beta (GSK3beta) was examined by two dimensional phosphopeptide mapping analysis on thin layer cellulose plates. Comparison of these peptide maps with those generated from wild type 1N4R tau isoform from which the phosphopeptide constituents and sites of phosphorylation had been determined previously, enabled us to monitor directly changes in phosphorylation of the individual tau proteins. No differences were found in the phosphorylation of wild-type, P301L or V337M tau by GSK3beta but the R406W mutant showed at least two clear differences from the other three tau proteins. The peptides, identified by mass spectrometry corresponding to phosphorylation at both threonine 231 and serine 235 (spot 3), serines 396, 400 and 404 (spot 6a) and serines 195 and 199 (spot 6b) were absent from the R406W peptide map. The findings imply that the R406W mutation in tau exerts long-range conformational effects on the structure of tau. PMID- 11278003 TI - Tyrosine-1290 of tetanus neurotoxin plays a key role in its binding to gangliosides and functional binding to neurones. AB - Tetanus toxin acts by blocking the release of glycine from inhibitory neurones within the spinal cord. An initial stage in the toxin's action is binding to acceptors on the nerve surface and polysialogangliosides are a component of these acceptor moieties. Using site-directed mutagenesis, we identify tyrosine-1290 of tetanus toxin as a key residue that is involved in ganglioside binding. This residue, which is located at the centre of a shallow pocket on the beta-trefoil domain of the tetanus H(c) fragment, is also shown to play a key role in the functional binding of tetanus toxin to spinal cord neurones leading to the inhibition of neurotransmitter release. PMID- 11278004 TI - Sphingolipids of the mycopathogen Sporothrix schenckii: identification of a glycosylinositol phosphorylceramide with novel core GlcNH(2)alpha1-->2Ins motif. AB - Acidic glycosphingolipid components were extracted from the yeast form of the dimorphic mycopathogen Sporothrix schenckii. Two minor and the major fraction from the yeast form (Ss-Y1, -Y2, and -Y6, respectively) have been isolated. By a combination of 1- and 2-D 1H-nuclear magnetic resonance (NMR) spectroscopy, electrospray ionization mass spectrometry (ESI-MS), and gas chromatography/mass spectrometry (GC/MS), Ss-Y6 was determined to be triglycosylinositol phosphorylceramide with a novel glycan structure, Manalpha1-->3Manalpha1- >6GlcNH(2)alpha1-->2Ins1-P-1Cer (where Ins=myo-inositol, P=phosphodiester). While the GlcNH(2)alpha1-->6Ins1-P- motif is found widely distributed in eukaryotic GPI anchors, the linkage GlcNH(2)alpha1-->2Ins1-P- has not been previously observed in any glycolipid. Ss-Y1 and Ss-Y2 were both found to have the known glycan structure Manalpha1-->3Manalpha1-->2Ins1-P-1Cer. Together with the results of a prior study [Toledo et al. (2001) Biochem. Biophys. Res. Commun. 280, 19-24] which showed that the mycelium form expresses GIPCs with the structures Manalpha1 ->6Ins1-P-1Cer and Manalpha1-->3Manalpha1-->6Ins1-P-1Cer, these results demonstrate that S. schenckii can synthesize glycosylinositol phosphorylceramides with at least three different core linkages. PMID- 11278005 TI - Cyclosporin A increases basal intracellular calcium and calcium responses to endothelin and vasopressin in human coronary myocytes. AB - Cyclosporin A (CsA) is a widely used immunosuppressive agent with severe side effects including hypertension. Here, we investigated the effects of CsA on intracellular free calcium ([Ca(2+)](i)) and the mechanisms involved in vasoconstriction in cultured human coronary myocytes. We used the Fura-2 technique for Ca(2+) imaging. Acute application of CsA at therapeutic concentrations (0.1-10 micromol/l) had no effect. Chronic exposure to CsA (1 micromol/l) for 24 h induced a small (20 nmol/l) but highly significant increase of basal [Ca(2+)](i) and enhanced the occurrence of spontaneous Ca(2+) oscillations. Endothelin- and vasopressin-induced rises of [Ca(2+)](i) were also enhanced. The demonstration that CsA increases basal [Ca(2+)](i) in addition to its impact on agonist receptor stimulation is of major importance for new therapeutic approaches. PMID- 11278007 TI - Patterns of shell penetration by Chorus giganteus juveniles (Gastropoda: Muricidae) on the mussel Semimytilus algosus. AB - Patterns are described for shell penetration by the sublittoral muricid snail Chorus giganteus during predatory attacks on the mussel Semimytilus algosus. Location, form and size of shell penetrations were observed in relation to the size of the predator. The results suggested that positions of the perforations on the mussel shells were related to size of the attacking snail. Smaller snails perforated areas near the shell ligament and in the central zone of the shell, while larger snails more frequently attacked shell borders, principally on the ventral side. These observations may be related to: (a) changes in the process of manipulation of the prey during development of the foot and the shell tooth of the predator, (b) changes in internal structure of the snails related to the shell perforation mechanism, or (c) learned behavior acquired experientially by the snails during early growth. Although in other studies of muricid penetration patterns larger boreholes made in shells of the prey were positively correlated with increasing predator size, this relation did not appear to hold with C. giganteus, as larger specimens often made relatively small shell perforations. Areas of boreholes made in the mussel shells by this snail varied from 0.01 to 1.1 mm(2), and were unusually variable in size and shape, especially when compared with literature results on bores characteristic of other muricid species. PMID- 11278008 TI - Quantification of radular marks as a method for estimating grazing of intertidal gastropods on rocky shores. AB - Wax discs have been used previously on intertidal rocky shores to record the grazing activity of gastropods. This study has evaluated this methodology for recording grazing of four common intertidal microalgal grazers on intertidal shores in New South Wales, Australia. In the laboratory, the four species examined-the patellid limpet, Cellana tramoserica (Sowerby), the trochid, Austrocochlea porcata (A. Adams), the neritid, Nerita atramentosa Reeve and the littorinid, Bembicium nanum (Lamarck)-made distinctive marks in the wax. These allowed identification of each species or combinations of species grazing over the different discs. Field experiments showed that the intensity of grazing, as indicated by the mean number of scratches per disc, was positively related to the number of gastropods in the surrounding area during low tide for C. tramoserica. The number of scratches per disc in any area was correlated with the percentage of discs scratched. The relationship for C. tramoserica was found at two scales in sites (approximately 3x3 m) and also in plots (50x50 cm) within sites. Therefore, densities that were measured when these limpets were inactive during low tide provided good estimates of grazing activity during high tide. This is largely because these limpets do not move far between where they rest and where they feed. The amount of microalgal food in the vicinity was not correlated with density, nor with grazing intensity. No relationship between density and grazing intensity was found for N. atramentosa, although experiments were only done in the field at one spatial scale (in sites, 3x3 m). Results obtained in the laboratory and in the field show that wax discs are useful to distinguish grazing by different species of gastropods on Australian rocky shores and allow tests of hypotheses about grazing activity at different spatial scales. PMID- 11278009 TI - Absence of population genetic differentiation in the New Zealand greenshell mussel Perna canaliculus (Gmelin 1791) as assessed by allozyme variation. AB - Genetic variation in the endemic New Zealand greenshell mussel, Perna canaliculus (Gmelin 1791), was examined using starch-gel electrophoresis at seven protein coding loci (Idh; Acon-1; Acon-2; Gpd; Pgi; Pgm; Pgd) in 35 populations (N=1038 mussels). For all loci and all populations, Fisher's exact tests indicated highly significant departures from Hardy-Weinberg equilibrium (HWE), but this overall result was caused by significant heterozygote deficiencies at only two loci (Pgm and Pgd), and in only three northern populations (Kuaotunu, Te Haumi and Days Bay). Allelic and genotypic differentiation between population pairs at individual loci and across all loci were nonsignificant, and genotypic disequilibrium at each locus pair was also nonsignificant for all populations. Genetic variation in all populations was high (mean heterozygosity, 0.210+/ 0.113), while Nei's D among populations was very low (0.002+/-0.002). Low population subdivision (θ=-0.001+/-0.002) and high levels of gene flow (Nm(p)=10.18; Nm(θ)=infinity) also indicated that the single panmictic unit model best explains population genetic homogeneity in P. canaliculus over a north south distance >2000 km. Lack of genetic subdivision in this species is discussed in light of two previous allozyme studies, with differing results: one suggested that a north-south division exists between greenshell mussel stocks, and the other suggested that population structure in this species can be explained through isolation by distance model modified by local hydrology. PMID- 11278010 TI - Physiological performance of juvenile southern flounder, Paralichthys lethostigma (Jordan and Gilbert, 1884), in chronic and episodic hypoxia. AB - Dissolved oxygen (DO) is proving to be one of the most important abiotic factors determining growth and survival of juvenile estuarine fish. In shallow, throughout estuarine systems, low DO can occur in two broad categories: a diel oscillating pattern resulting in repeated nocturnal hypoxia due to the photosynthesis-respiration cycle of algal populations, or as prolonged bottom water hypoxia or anoxia caused by stratification. A series of laboratory experiments was conducted to characterize the physiological performance of juvenile southern flounder, Paralichthys lethostigma, (55-65 mm TL) exposed to four treatments of DO: (1) constant normoxia (6.50+/-0.50 mg O(2) l(-1)), (2) constant hypoxia (2.79+/-0.19 mg O(2) l(-1)), (3) constant intermediate hypoxia (4.74+/-0.18 mg O(2) l(-1)), and (4) an oscillating oxygen environment cycling dielly between the normoxic and hypoxic levels (2.8-6.2 mg O(2) l(-1), daily mean=4.40 mg O(2) l(-1)). Routine respiration was positively correlated with DO level and increased significantly during the day in the oscillating treatment in response to increasing DO. Ventilation rates were negatively correlated with the DO level in the constant treatments and increased significantly at night in the oscillating treatment in response to nocturnal hypoxia. Similarly, hematocrit levels were negatively related to DO levels in the constant treatments after 5 and 26 days of exposure to the treatments. Hematocrit levels also increased significantly the oscillating treatment, apparently in response to the episodic nocturnal hypoxia. Growth was significantly reduced in the 2.8 mg O(2) l(-1) treatment and the oscillating treatment but not in the 4.7 mg O(2) l(-1) treatment. Acclimation was evident by an increase in growth rates from week 2 to week 3 and a decrease in hematocrit levels between 5 and 26 days of exposure in the 2.7 and 4.5 mg O(2) l(-1) treatments but was not evident in the normoxic or oscillating treatments. These results suggest that a juvenile fish must remain in even moderately low DO in order for acclimation to occur. The research presented demonstrates that correctly assessing habitat quality in terms of DO requires knowledge of a fish's physiological and environmental history. PMID- 11278011 TI - The autofluorescent age pigment lipofuscin: key to age, growth and productivity of the Antarctic amphipod Waldeckia obesa (Chevreux, 1905). AB - Peracarid crustaceans are among the most important taxa in terms of biodiversity and carbon-flow within the Weddell Sea benthos; however, very few data on their age, growth and productivity are available. This study uses the pigment lipofuscin as an age marker in the scavenging amphipod Waldeckia obesa (Chevreux, 1905) from the eastern Weddell Sea. Resin brain sections of 159 trap-caught specimens (1.2 to 7.7 mm coxal plate length L(cox) equal to 5 to 31 mm total length) were recorded digitally by confocal microscopy, and images were analysed. A modal progression analysis of the lipofuscin concentration-frequency distribution revealed five regularly spaced modes presumed to reflect consecutive annual age classes. Single females outside the range of mode V occurred, indicating maximum age of up to 8 years in females. No regular modes were obvious from the comparable length-frequency distribution of 386 individuals. Average yearly pigment accumulation was linear, and accumulation rates did not differ between sexes. The estimates of the growth parameters L(infinity) and k of the von Bertalanffy growth function were 7.47 mm L(cox) and 0.50 per year in females, respectively, and 6.92 mm L(cox) and 0.60 per year in males, respectively. Mortality, estimated from catch curves, amounted to 0.27 per year in females and 0.43 per year in males. P/B ratio, calculated from the mass specific growth rate method, was 0.38 per year for the pooled population (0.25 per year in females, 0.31 per year in males, 2.26 per year in juveniles). The results are discussed with regard to advantages and drawbacks of the methodology, and are compared with results from warmer water habitats. PMID- 11278012 TI - Change in the concentrations of iron in different size fractions during a phytoplankton bloom in controlled ecosystem enclosures. AB - To observe micronutrient dynamics in the plankton ecosystem, controlled ecosystem enclosure (CEE) experiments were conducted in Saanich Inlet, B.C., Canada. Two CEEs (2.5 m in diameter, 16 m in length, one for Fe studies and the other for biological studies) were launched for the period 22 July to 5 August 1996 and enriched with 10 uM nitrate and 5.2 nM Fe (13% of total Fe) on day 1. Sampling from three integrated depths, intervals 0-4, 4-8 and 8-12 m, was performed on days 0, 1, 2, 3, 4, 5, 7, 9 and 11. Iron concentrations were measured for five size fractions: >25 um particles, 2-25 um particles, 0.2-2 um particles, 0.2 um 200 kDa small colloidal particles and <200 kDa soluble species. The sediment in the Fe enclosure was also collected on every sampling day after day 2 and its Fe was determined. Size-fractionated particulate organic carbon and total chlorophyll-a were also analyzed.The Fe in small colloidal particles (200 kDa-0.2 um) comprised 78% of the traditionally defined dissolved phase (<0.2 um) on day 1. Of all the size fractions of Fe, the small colloidal particulate fraction decreased most significantly during the phytoplankton bloom. In the dissolved fraction (<0.2 um), the small colloidal particle fraction comprised 79% of the decrease. The decrease in concentration of Fe in small colloidal particles was larger than that of total Fe from day 1 to day 4. In contrast, the >25 um Fe particles increased over the same period. These results suggest that Fe in small colloidal particles changed to >25 um Fe particles during phytoplankton growth. A large amount of Fe was kept in the surface layer with the phytoplankton, and transported to the deep layer by phytoplankton sedimentation, at the end of the bloom. From these results, the small colloidal particulate Fe seems to be the most dynamic size fraction and a high percentage of Fe in small colloidal particles changed to large particles due to chemical/physical aggregation and/or physical adsorption to suspended particles such as phytoplankton cells. PMID- 11278013 TI - Effects of periodic hypoxia on mortality, feeding and predation in an estuarine epifaunal community. AB - The York River Estuary, a tributary of the Chesapeake Bay, USA, experiences periodic low oxygen stress (hypoxia), yet epifaunal species form dense communities there. We studied hypoxia tolerance of common epifaunal species in the York River by exposing sessile and mobile epifauna to high and low oxygen concentrations in laboratory aquaria. Mortality in hypoxia varied among species, ranging from 0% to 100%, with trends of decreased tolerance by mobile species relative to sessile species. While most species tested experienced some mortality after being exposed to hypoxia (at 1 mg O(2)/l or 0.5 mg O(2)/l) for 5 days, many species had a median lethal time (LT(50)) in hypoxia greater than 1 week (3 of 6 species at 1 mg O(2)/l and 6 of 14 species at 0.5 mg O(2)/l), the maximum duration of typical hypoxic episodes in the York River, suggesting that hypoxia may cause little mortality for some species in this system. However, hypoxia had sub-lethal effects on behavior in all species tested. Epifaunal animals responded to hypoxia with behaviors that moved them higher in the water column or by entering resting states until hypoxia passed. Feeding and predation by a variety of taxa (the hydroid Obelia bicuspidata, the mud crab Neopanope sayi, juvenile blue crabs Callinectes sapidus, the flatworm Stylochus ellipticus, and the nudibranch Doridella leucolena) decreased during hypoxia, despite varying mortality responses to low oxygen stress, suggesting that short hypoxic episodes may create predation refuges for prey species. At least one highly tolerant species (O. bicuspidata) showed substantially decreased growth in hypoxia. Although relatively high tolerance of hypoxia by many estuarine epifaunal species limits serious disturbance during brief hypoxic episodes, hypoxia's greatest impact on York River epifaunal communities might be through its indirect effects on behavior and predation. PMID- 11278018 TI - New method for coating tympanostomy tubes to prevent tube occlusions. AB - OBJECTIVE: tympanostomy tube insertion is currently the most common surgical procedure requiring general anesthesia performed on children. Occlusion of the tube and prolonged otorrhea through the tube are typical problems associated with the use of middle-ear ventilation tubes. In this study, a new method for coating ventilation tubes is introduced that prevents occlusion of the tube lumen by granulation tissue, blood clot or pus. METHODS: human serum albumin (HSA) was used to coat standard tympanostomy tubes of different materials. Fibronectin, a typical protein in serum and exudates and one of the most adhesive glycoproteins, was used as a model representative of exudates of the ear. RESULTS: when compared with the binding on uncoated tubes, the binding of fibronectin on HSA-coated tubes was inhibited from 59 to 85%, depending on the tube material used. CONCLUSIONS: HSA-coating markedly reduced the binding of fibronectin on tube surfaces in vitro. The study shows the potential role of HSA-coating in preventing the adherence of foreign material to tympanostomy tubes and reducing tube occlusions. PMID- 11278019 TI - Post-transplant lymphoproliferative disease in tonsils of children with liver transplantation. AB - OBJECTIVE: To assess the incidence and characteristics of post-transplant lymphoproliferative disease (PTLD) in tonsils of the liver transplanted children. METHODS: All patients under 14 years of age recipients of a liver transplant at the institution and operated on for tonsillectomy under suspicion of malignancy were included in this study. RESULTS: Seven patients underwent surgery on their tonsils under suspicion of PTLD. One case of B-cell lymphoma, and three cases of polymorphic diffuse B-cell hyperplasia were found. This represents an incidence of 1.4% of PTLD in the tonsils of the 283 pediatric liver transplants performed at the hospital. CONCLUSION: The incidence of PTLD in tonsils after liver transplantation is very low at the institution. However, it is very important to follow-up allograft recipients for early diagnosis of this entity. PMID- 11278020 TI - Prospective study of morbidity after tonsillectomy in children. AB - Post-operative morbidity was prospectively studied in 384 children after tonsillectomy or adenotonsillectomy, using visual analogue scores to record symptom levels, and questionnaires to monitor satisfaction scores from the children and their families. Assessments were performed between the 7th and 14th day post-operatively. Two hundred children were assessed before the introduction of a pre-admission programme which consisted of an instructional videotape session in the ward and an advice booklet. Department practice was also changed to provide a bottle of paracetamol on discharge routinely for each child. Following these changes in practice a further 184 children were assessed. The provision of relatively simple measures in the programme increased parental satisfaction rates (P<0.05) and reduced GP contact rates (35--17%, P<0.05) post operatively. The actual levels of morbidity were unchanged despite the provision of analgesia. PMID- 11278021 TI - Disordered breathing during sleep in patients with mucopolysaccharidoses. AB - OBJECTIVE: Obstructive sleep apnoea (OSA) has been reported as a feature of children with mucopolysaccharidoses (MPS). However, the incidence and severity of OSA with respect to disease type is poorly defined. The aim of the present study was to measure objectively the degree of OSA in a group of children with a range of MPS syndromes. METHODS: In a cross-sectional study, cardiopulmonary sleep studies were performed during unsedated sleep in 26 children with MPS over a period of 2 years. Scores of OSA severity based upon clinical history and upon objective sleep study data were made in each case and compared. RESULTS: OSA was present in 24/26 patients, and ranged in severity from mild to severe. OSA was most marked in MPS type IH (Hurler syndrome) followed by types IHS (Hurler- Scheie syndrome) and II (Hunter syndrome). Frequent arousals and poor sleep quality, not suspected clinically, were noted in several patients. There was agreement between the clinical and objective scoring systems in only 17/26 patients (65%) with clinical history scores tending to underestimate the most severe cases (5/26 cases) and overestimate the severity in the mild cases (4/26 cases). CONCLUSIONS: Obstructive respiratory problems are frequent in MPS patients and there are differences in severity of OSA between the different MPS types. Assessments of the severity of OSA based upon clinical history alone are inadequate. Our results suggest that objective sleep studies are necessary to evaluate these cases, to monitor clinical outcome and to assess the effects of therapeutic intervention. Prospective studies in larger numbers of patients are needed to validate these observations. PMID- 11278022 TI - Otoacoustic emissions and auditory assessment in infants at risk for early brain damage. AB - The importance of early hearing screening has long been recognized, as the prognosis for the hearing impaired child is improved when the diagnosis is made as early as possible, and the intervention is begun immediately. For clinical screening of hearing impairment, the recording of otoacoustic emissions was recommended. As some risk factors for early brain damage are at the same time also risk factors for dysfunction of auditory system, we presumed that infants at risk for brain damage have hearing impairment more frequently than the rest of the population of the same age. We were interested in the role of otoacoustic emission testing during the assessment of auditory function in these infants. There were 110 infants at risk for brain damage included in the study. After thorough otorhinolaryngological examination, auditory function was estimated by recording of otoacoustic emissions, tympanometry, pure tone audiometry and, when necessary, auditory brainstem responses. Otoacoustic emissions were recorded by Madsen-Electronics Celesta 503 in an acoustically treated sound room. We registered spontaneous as well as transient and distortion product otoacoustic emissions. The neurologist formed two groups with different degrees of neurological risk. The collected results of auditory function were compared with the degree of neurological risk. For the statistical analysis, the procedure chi(2) and Fischer test were used. Spontaneous otoacoustic emission was detected in 38.2% of examinees. Evoked otoacoustic emissions were registered in 87.3% of infants. The testing had to be repeated in 32.7% of infants. We observed evoked otoacoustic emissions to be present also in a child with sensorineural hearing impairment and no auditory brainstem responses. Up to 32.7% of infants at risk for brain damage were hard of hearing. Conductive hearing loss was discovered with 25.4% of infants, and eight (7.3%) had sensorineural hearing impairment. In the group of examinees with only risk factors 3.6% had sensorineural impairment and in a group with abnormal motor development, there were 18.5% with that kind of hearing loss. Fischer test confirmed a statistically significant difference between the groups. Infants at risk for brain damage have more frequently impaired auditory function than their peers. For this reason, it is especially important to focus attention on the hearing condition when dealing with this population. Recording of evoked otoacoustic emissions is very helpful in pediatric audiometry, but any interpretation of the results should consider the possibility of auditory neuropathy. PMID- 11278023 TI - Changes in external ear resonance after ventilation tube (Grommet) insertion in children with otitis media with effusion. AB - OBJECTIVE: As otitis media with effusion is common in children, the effects of a ventilation tube should be taken into account in the prescription of hearing aids for children. In ears with a ventilation tube, the external auditory canal communicates directly with the middle ear space, and so the impedance of the middle ear may change. Consequently, this will affect external-ear resonance. The aim of this study is to observe the effects of the tympanic membrane perforations caused by the ventilation tube on external-ear resonance. We selected 30 ears with otitis media with effusion to measure external-ear resonance before and after ventilation tube insertion. We compared the external-ear resonance of a control group with that of the otitis media with the effusion group and two types of ventilation-tube groups, respectively. In the subjects who have otitis media with effusion, the average gain of the peak resonance was larger than that in the control group. After ventilation-tube insertion, the amplitude of the gain decreased to the same level as the control group, but a characteristic negative gain appeared around 1000 Hz in about half of all cases. This negative gain was observed more frequently in the ventilation tube with a larger diameter. The raised peak resonance gain in the otitis media with effusion group decreased to a level roughly the same as that of the control group after ventilation-tube insertion. Provision of an additional gain in the low frequencies around 1000 Hz should be considered for patients with a ventilation tube when prescribing hearing aids. PMID- 11278024 TI - Relationship between NRT measurements and behavioral levels in children with the Nucleus 24 cochlear implant may change over time: preliminary report. AB - OBJECTIVE: Response from spiral ganglion cells to electrical stimulation via the Nucleus 24 cochlear implant can be measured using the neural response telemetry system. The purpose of this study was to assess, in children, the correlation between the neural response threshold and the behavioral levels used for cochlear implant programming process. METHODS: The neural response telemetry test was administered to 23 children (mean age at implantation: 4 years) with the Nucleus 24 cochlear implant. Four intra-cochlear electrodes (electrodes 5, 10, 15 and 20) were tested. The neural response threshold at 3, 6, 9 and 12 months post implantation was compared with the behavioral threshold and the maximum comfort level estimated during the same periods: a Pearson's correlation test was performed for each tested electrode. RESULTS: On apical electrodes, the correlation with the behavioral threshold remained significant from 3 to 12 months post-implantation (r ranging from 0.696 to 0.909, P<0.05), and the correlation with the maximum comfort level was also significant throughout the study period, except on electrode 15 at 9 months (tendency to significance). On basal and intermediate electrodes, statistical correlations were found only at some points of time; nonetheless, at 12 months post-implantation, a significant correlation with behavioral levels could be clearly demonstrated both on electrode 15 (r=0.914--0.778, P<0.05) and on electrode 10 (r=0.845--0.720, P<0.05). CONCLUSIONS: This preliminary study suggests that the correlation between the neural response threshold and behavioral levels may improve from the base towards the apex of the cochlea. However, a significant correlation can be demonstrated for all tested electrodes at 12 months post-implantation. During the first months post-implantation care must be exercised when interpreting neural response telemetry measurements: a positive test does not necessarily mean that the stimulus delivered to the acoustic nerve will be centrally processed with the result of an auditory perception. PMID- 11278025 TI - Extranasopharyngeal angiofibroma arising from the nasal septum. AB - Extra nasopharyngeal origin of angiofibroma is very rare. The nasal septum is a very rare site of extra nasopharyngeal angiofibroma with only two cases reported in the medical literature. We report here a case of a vascular mass arising from the nasal septum of an 8 year old boy. Histopathology confirmed it to be a case of angiofibroma. A review is also made of the other reported cases of angiofibroma arising from the nasal cavity. The likely theory of origin of the tumor and the management is also discussed. PMID- 11278026 TI - Multiple bilateral orbital abscesses secondary to nasal furunculosis. AB - Orbital inflammation secondary to sinus inflammation is a well known entity and has been widely reported. However, nasal furunculosis resulting in orbital inflammation is a rare occurrence. We present a case of a 2-year-old boy who developed multiple bilateral orbital abscesses secondary to nasal furunculosis. To our knowledge such a case has not been reported before. PMID- 11278027 TI - Otic capsule fracture with preservation of hearing and delayed-onset facial paralysis. AB - The unusual occurrence of an otic capsule fracture with preservation of hearing is presented. In addition, the patient suffered facial paralysis beginning 6 days after the injury that rapidly recovered. Fifteen-month follow-up reveals stable hearing thresholds. The course of a fracture through the inner ear could be an important factor in determining the potential for hearing preservation. PMID- 11278028 TI - Duplication of the external auditory canal: a report of three cases. AB - OBJECTIVE: Malformations of the first branchial cleft are uncommon and only sporadically reported in the literature. They may present as inflammatory openings on the neck, bland cysts or fistula associated with the external auditory canal. In this retrospective study, clinical features and anatomical relationships are described in three pediatric cases. Therapeutical guidelines for surgical management of first branchial cleft anomalies are discussed. PATIENTS: Between 1997 and 1999 three patients aged 9 months, 2 and 7 years with first branchial cleft anomalies were included in this study. All patients were treated surgically, wide exposure and superficial parotidectomy was necessary for complete removal in two of three cases. RESULTS: Exploring patients histories revealed previous infections with repeated incision and drainage procedures as well as inadequate operative resections. Clinically, purulent drainage from the ear, swelling in the parotid area and abscess formation with persistent drainage after incision in the neck or parotid area were noted. CONCLUSIONS: From our case series two of three patients underwent inadequate incision and drainage procedures to combat infection followed by scar tissue formation. Because of the variable relation to the facial nerve this led to difficulties in identifying and protecting the nerve during definite surgery. Management of first branchial cleft anomalies must include the facilities to achieve ear surgery and superficial parotidectomy including facial nerve exposure. PMID- 11278029 TI - The presence of the dam affects the efficiency of gentling and feeding on the early establishment of the stockperson-lamb relationship. AB - This experiment investigates how the maternal presence influences the effect of additional human contact in early age on the reaction of lambs to their stockperson. Forty twin-born lambs were involved in this experiment during their first 4 days of life. Ten pairs of twins were reared artificially from 12h of age. One of each litter (AF, n=10) received 6.5+/-0.7 sessions of 30min of separation from the twin (with a wire fence) with 5min of gentling and feeding (suckling from a bottle and from a bucket fitted with a rubber teat). The other twin was not treated. Ten pairs of twins were reared with their dam and received 6.6+/-0.7 sessions of treatment. One twin (MAF, n=10) received the same treatment as AF. The other twin (M, n=10) was separated for 30min from the dam and had no human contact. From the age of 70+/-7h, lambs were tested in a social isolation test (alone for 1min, with the familiar stockman for 2min, alone again for 1min), in a Preference test (2min) between an unfamiliar maternal ewe and the familiar stockman, and, for the AF lambs only, in a Preference test (2min) between their familiar and an unfamiliar stockman. Eight AF lambs learned to suck on their own from the bucket of milk by the end of the experiment compared to only one MAF (P<0.001). AF lambs approached the human more (P<0.01), vocalised less (P<0.01) and walked less (P<0.01) during the social isolation test than animals reared with their mother (M and MAF). AF did not show any preference between the stockman and the unfamiliar maternal ewe while M and MAF lambs chose the ewe (P<0.05). AF lambs discriminated the familiar from an unfamiliar stockman only if they had learned to suck from the bucket during the treatment. Nevertheless MAF lambs vocalised less than M (P<0.05) in the presence of the stockman during the social isolation test, indicating a possible reduction of isolation distress. These results show that artificially reared lambs are preferentially motivated to interact with a familiar human after only a few days of contact. Moreover, they highlight the difficulty in using a feeding reward to improve the human-lamb relationship when lambs are reared permanently with their dams. However, the results suggest that early gentling improves the human-animal relationship whatever the maternal environment. PMID- 11278030 TI - Social facilitation of predatory, sheep-chasing behaviour in Norwegian Elkhounds, grey. AB - Sheep grazing on unfenced mountain pasture may be attacked by loose dogs, which may chase and kill sheep and separate lambs from their mothers. We have earlier shown that testing dogs individually towards sheep with an electronic dog collar may effectively reduce the chasing propensity 1 year later. The aim of this study was to investigate whether and how a non-chasing and a chasing companion dog influence sheep chasing in test dogs (20 Norwegian Elkhounds, grey), and whether this varies with sex or age. The test dogs' predatory behaviour towards sheep was examined by observation of the dogs in a fenced enclosure with sheep in 5min tests together with, first, a non-chasing (Hamilton stoever, a hound breed) and 2 3 days later a chasing (Border collie) companion dog. Physical contact with sheep was prevented. Initially, the test dogs exhibited a higher chasing motivation towards sheep in tests with a chasing, as compared with a non-chasing, companion (P<0.001). During the entire test, 60% of the dogs attacked sheep when accompanied by the non-chaser. All dogs attacked sheep when the chasing companion was present (P=0.008), although in only 8 of 20 cases the companion dog chased simultaneously with the test dog. In the chasing-companion tests, the attack latency was shorter (P<0.001) than in tests with a non-chasing companion. The attack severity was higher when attacking alone than when attacking together with the chasing companion (P=0.033). In these tests, the higher the attack severity, the shorter was the attack latency (P=0.006). A sequence analysis on chasing companion tests showed that test dogs generally started with observing or showing interest in sheep, followed by attacks, which increased in severity. No sex differences were observed. In non-chasing companion tests, dogs aged up to 2.5 years exhibited a weaker initial hunting motivation than older dogs (P=0.025), but during the entire test were more inclined to attack (P=0.019). Taken together, our findings indicate that a companion dog showing intentions of predatory behaviour stimulates predatory chase in another dog, while a non chasing companion has a limited influence on this. In tests certifying dogs for their refrainment of chasing sheep, well trained Border collies approaching sheep on command might be used to reveal the full predatory potential of the dog being tested. PMID- 11278031 TI - Behavioural differences between three breed groups of hunting dogs confronted with domestic sheep. AB - When running free in open fields, domestic dogs occasionally display predatory behaviour towards domestic sheep. This has not yet been studied scientifically. The aim of the present study was to investigate the inclination to chase sheep in three breed groups of hunting dogs that are most frequently used in areas with grazing sheep. We studied 41 elkhounds, 29 hare hunting dogs and 68 English setters. Behaviours indicative of motivation for chasing or attacking sheep were examined in three different ways. A path test examined functional traits such as hunting ability, contact willingness, reactivity to sudden noise, and response towards a lone sheep. In a sheep confrontation test, loose-leashed dogs were observed in a fenced enclosure with sheep and given electric shocks through an electronic dog collar if within 1-2m from the sheep. A questionnaire to the dog owners supplied information on their dog's previous experience with sheep and behavioural responses to various types of novel stimuli. No significant sex differences were found. The elkhounds showed the highest interest in a lone sheep in the path test, and displayed the highest initial hunting motivation, the highest percentage of dogs starting a sheep attack, the highest attack severity, and were most frequently given el. shocks. The hare hunting dogs were intermediate, while setters showed the lowest values for these variables. Dogs reported as showing low fearfulness more frequently acted as potential sheep chasers in the tests. Dogs up to 3 years of age showed a more pronounced initial hunting motivation and more frequent attacks than older dogs, although there were no age differences in the number of el. shocks given in the test. The latter may be related to the more frequent abruption of attacks among younger dogs. The main factors predicting a high hunting motivation and attack severity were lack of previous opportunity to chase sheep, low fearfulness towards gunshots and unfamiliar people, and general interest in sheep shown when encountering them. Probability of sheep chasing differed between dog breeds and age groups. Previous experience and certain character traits were indicative of a high predatory motivation towards sheep. PMID- 11278032 TI - Behavioural changes and aversive conditioning in hunting dogs by the second-year confrontation with domestic sheep. AB - Domesticated dogs occasionally exhibit predatory behaviour towards domestic sheep when running loose in pasture. Both young and old dogs of either sex may chase sheep. Electronic dog collars applying electric shocks are utilised as one method of training dogs to refrain from attacking sheep. This device is used for a number of other training purposes which have raised concern for the welfare of the dogs being trained. This study aims at testing long-term learning effects of previous sheep tests on sheep chasing in hunting dog breeds (Norwegian elkhounds (grey), English setters, and hare hunting dogs), in particular with use of electronic dog collars, in addition to uncovering potential secondary negative effects on dogs' behaviour and mental stability. The dogs (N=114) were subjected to three tests for two subsequent years, the second year being reported here. Dogs were tested for reactions to different stimuli, including a sheep, in a path test. In a sheep confrontation test, dogs were fenced in with a sheep group and given el. shocks when approaching 1-2m from sheep. A questionnaire to the dog owners reported differences in dogs' behaviour between the years.Dogs showed weaker or delayed behavioural responses in both tests in the second year. No dogs showed interest in or attacked a lone sheep in the path test in the second year, while almost two thirds of them did so the first year. In the sheep confrontation test, the dogs exhibited comparatively hesitant initial hunting motivation the second year, being more evident in dogs which received el. shocks the first year. No dogs chased or attacked sheep as their first response in this test, while half of them did so the first year. The proportion of dogs attacking sheep during the entire test was reduced to almost one fourth. The number of el. shocks administered reduced by the second year, and only one of the dogs that received el. shocks the first year received el. shocks the second year. The owners reported no negative effect on the dogs' behaviour during the year ensuing el. shock treatment. Eighteen of the 24 dogs reported by owners to exhibit behavioural changes lost their previous interest in sheep.The second-year tests indicate that aversive conditioning with the use of electronic dog collar may be an efficient method for reducing the probability of a dog chasing or attacking grazing sheep. No adverse effects were observed with our test procedure. PMID- 11278033 TI - A concept of welfare based on reward evaluating mechanisms in the brain: anticipatory behaviour as an indicator for the state of reward systems. AB - In this review we attempt to link the efficiency by which animals behave (economy of animal behaviour) to a neuronal substrate and subjective states to arrive at a definition of animal welfare which broadens the scope of its study. Welfare is defined as the balance between positive (reward, satisfaction) and negative (stress) experiences or affective states. The state of this balance may range from positive (good welfare) to negative (poor welfare). These affective states are momentary or transient states which occur against the background of and are integrated with the state of this balancing system. As will be argued the efficiency in behaviour requires that, for instance, satisfaction is like a moving target: reward provides the necessary feedback to guide behaviour; it is a not steady-state which can be maintained for long. This balancing system is reflected in the brain by the concerted action of opioid and mesolimbic dopaminergic systems. The state of this system reflects the coping capacity of the animal and is determined by previous events. In other words, this integrative approach of behavioural biology and neurobiology aims at understanding how the coping capacity of animals may be affected and measured. We argue that this balancing system underlies the economy of behaviour. Furthermore we argue that among other techniques anticipation in Pavlovian conditioning is an easy and useful tool to assess the state of this balancing system: for estimating the state of an animal in terms of welfare we focus on the conditions when an animal is facing a challenge. PMID- 11278034 TI - Thermoprecipitation of lysozyme from egg white using copolymers of N isopropylacrylamide and acidic monomers. AB - Thermoprecipitation of lysozyme from egg white was demonstrated using copolymers of N-isopropylacrylamide with acrylic acid, methacrylic acid, 2-acryloylamido-2 methylpropane-sulfonic acid and itaconic acid, respectively. Polymers synthesized using molar feed ratio of N-isopropylacrylamide:acidic monomers of 98:2 exhibited lower critical solution temperatures in the range of 33--35 degrees C. These polymers exhibited electrostatic interactions with lysozyme and inhibited its bacteriolytic activity. The concentration of acidic groups required to attain 50% relative inhibition of lysozyme by the polymers, was 10(4)--10(5) times lower than that required for the corresponding monomers. This was attributed to the multimeric nature of polymer-lysozyme binding. More than 90% lysozyme activity was recovered from egg white. Polymers exhibited reusability up to at least 16 cycles with retention of >85% recovery of specific activity from aqueous solution. In contrast, copolymer comprising natural inhibitor of lysozyme i.e. poly (N-isopropylacrylamide-co-O-acryloyl N-acetylglucosamine) lost 50% recovery of specific activity. Thermoprecipitation using these copolymers, which enables very high recovery of lysozyme from egg white, would be advantageous over pH sensitive polymers, which generally exhibit lower recovery. PMID- 11278035 TI - Lipase catalyzed synthesis of organic acid esters of lactic acid in non-aqueous media. AB - Lipases from Rhizomucor miehei (Lipozyme IM20) and porcine pancreas (PPL) were employed as catalysts for the esterification reaction between the hydroxyl group of lactic acid and the carboxyl group of organic acids. Reactions were carried out at both shake-flask and bench-scale levels. Various parameters, such as solvent, temperature, substrate and enzyme concentrations, effect of buffer volume, buffer pH and water volume, were investigated for optimization of yields. While ethylmethyl ketone (EMK) was found to be the best solvent for shake-flask reactions, chloroform gave higher yields at bench-scale level. Detailed studies were carried out with respect to the synthesis of palmitoyl and stearoyl lactic acids. At shake-flask level, maximum yields of 37.5 and 40% were observed in case of palmitoyl and stearoyl lactic acids, respectively, with Lipozyme IM20; at bench-scale level, the maximum yields were 85.1 and 99% respectively, when PPL was employed. Of all the organic acids employed (C(2)--C(18)), only lauric, palmitic and stearic acids gave yields above 50%. At bench-scale level, PPL could be reused for up to three cycles with yields above 40%. Esters prepared were found to conform to Food Chemical Codex (FCC) specifications in terms of acid value, ester value, sodium and lactic acid contents. PMID- 11278036 TI - Removal of ethylacetate vapor from waste gases by a trickle-bed air biofilter. AB - Biofilter system is a relatively new process that has been proven to be more cost effective than traditional technologies such as carbon adsorption, liquid scrubbing, condensation, thermal incineration, and catalytic incineration for removing low-strength volatile organic compounds from waste gases. The trickle bed air biofilter (TBAB) performance for ethylacetate (EA) removal was evaluated under different influent loadings. In the pseudo-steady states, the elimination capacity increased, but the removal efficiency decreased with increased influent loading. More than 95 and 90% removal efficiencies could be achieved for EA loadings below 490 and 810 g m(-3) h(-1), respectively. The TBAB appears to be very effective for controlling EA emission under low to high loading conditions, and the effectiveness could be maintained over 190 days of laboratory operation. PMID- 11278037 TI - Immobilization of sugar-non-specific nucleases by utilizing the streptavidin- biotin interaction. AB - Due to their high enzymatic activity, the sugar-non-specific endonucleases from Serratia marcescens and Anabaena can be used for a number of applications, such as the removal of contaminating genetic material from biological preparations, footprinting studies, and the determination of nucleic acids in biochemical samples. These methods would benefit from immobilized nucleases. For this purpose, a single cysteine residue was added at the N-terminus of the Serratia and Anabaena nucleases and subsequently modified with a maleimide-biotin conjugate. Alternatively, a biotin acceptor domain was fused to the Anabaena nuclease, allowing biotinylation during expression in E. coli without a further chemical step. The attachment of biotin-modified nucleases to streptavidin-coated paramagnetic beads and to streptavidin-coated surface plasmon resonance sensor chips (to study interactions with substrate and inhibitor) worked well when aggregates present in the protein preparations were removed by ultrafiltration. These methods should be of general use for similar enzyme systems. PMID- 11278038 TI - Hydrophobic interaction chromatography of proteins. AB - In this article, an overview of hydrophobic interaction chromatography (HIC) of proteins is given. After a brief description of protein hydrophobicity and hydrophobic interactions, we present the different proposed theories for the retention mechanism of proteins in HIC. Additionally, the main parameters to consider for the optimization of fractionation processes by HIC and the stationary phases available were described. Selected examples of protein fractionation by HIC are also presented. PMID- 11278039 TI - An aerobic fixed-phase biofilm reactor system for the degradation of the low molecular weight aromatic compounds occurring in the effluents of anaerobic digestors treating olive mill wastewaters. AB - An aerobic co-culture, prepared by combining Ralstonia sp. LD35 and Pseudomonas putida DSM1868, was recently found to be capable of extensively degrading many of the hydroxylated and/or methoxylated benzoic, phenylacetic and 3-phenyl-2 propenoic acids occurring in the olive mill wastewaters (OMWs). In the perspective of developing a biotechnological process for the degradation of low molecular weight (MW) aromatic compounds occurring in the effluents of anaerobic digestors treating OMWs, the capability of this bacterial co-culture of biodegrading a synthetic mix of the above mentioned compounds and the aromatic compounds of an anaerobic OMW-treatment plant effluent in the physiological state of immobilised cells was investigated. Two aerobic fixed-bed biofilm reactors were developed by immobilising the co-culture cells on Manville silica beads and on polyurethane foam cubes. Both supports were found to give rise to a microbiologically stable and biologically active biofilm. The two biofilm reactors were found to be similarly capable of rapidly and completely biodegrading the components of a synthetic mix of nine monocyclic aromatic acids typically present in OMWs and the low-MW aromatic compounds occurring in the anaerobic effluent in batch conditions. However, in the same conditions, the silica bead-packed reactor was found to be more effective in the removal of high MW phenolic compounds from the anaerobic effluent with respect to the polyurethane cube-packed reactor. These results are encouraging in the perspective of using the co-culture as immobilized cells for developing a continuous biotechnological process for the post-treatment of effluents with low MW aromatic compounds produced by anaerobic digestors treating OMWs. PMID- 11278040 TI - Genetic engineering of the Trichoderma reesei endoglucanase I (Cel7B) for enhanced partitioning in aqueous two-phase systems containing thermoseparating ethylene oxide--propylene oxide copolymers. AB - Endoglucanases (endo-1,4-beta-D-glucan-4-glucanohydrolase, EC 3.2.1.4) are industrially important enzymes. In this study endoglucanase I (EGI or Cel7B) of the filamentous fungi Trichoderma reesei has been genetically engineered to investigate the influence of tryptophan rich peptide extensions (tags) on partitioning in an aqueous two-phase model system. EGI is a two-domain enzyme and is composed of a N-terminal catalytic domain and a C-terminal cellulose binding domain, separated by a linker. The aim was to find an optimal tag and fusion position, which further could be utilised for large scale extractions. Peptide tags of different length and composition were attached at various localisations of EGI. The fusion proteins were expressed from T. reesei with the use of the gpdA promoter from Aspergillus nidulans. Variations in secreted levels between the engineered proteins were obtained. The partitioning of EGI in an aqueous two phase system composed of a thermoseparating ethylene oxide-propylene oxide random copolymer (EO(50)PO(50)) and dextran, could be significantly improved by relatively minor genetic engineering. The (Trp-Pro)(4) tag added after a short stretch of the linker, containing five proline residues, gave in the highest partition coefficient of 12.8. The yield in the top phase was 94%. The specific activity was 83% of the specific activity of unmodified EGI on soluble substrate. The efficiency of a tag fused to a protein is shown by the tag efficiency factor (TEF). A hypothetical TEF of 1.0 would indicate full tag exposure and optimal contribution to the protein partitioning by the fused tag. The location of the fusion point after the sequence of five proline residues in the linker of EGI is the most beneficial in two-phase separation. The highest TEF (0.97) was obtained with the (Trp-Pro)(2) tag at this position, indicating full exposure and intactness of the tag. However, the peptide tag composed of (Trp-Pro)(4) improved the partition properties the most but had lower TEF in comparison to (Trp Pro)(2). PMID- 11278042 TI - Synthesis and influenza virus sialidase inhibitory activity of analogues of 4 Guanidino-Neu5Ac2en (Zanamivir) modified in the glycerol side-chain. AB - Analogues of 4-Guanidino-Neu5Ac2en (Zanamivir) have been prepared containing carbamate substituents at the 7-hydroxy position. (4S,5R,6R)-5-Acetylamino-6-[1R [(6-aminohexyl)carbamoyloxy]-2R,3-dihydroxypropyl]-4-guanidino-5,6-dihydro-4H pyran-2carboxylic acid and (4S,5R,6R)-5-Acetylamino-6-[1R-[heptylcarbamoyloxy] 2R,3-dihydroxypropyl]-4-guanidino-5,6-dihydro4H-pyran2-carboxylic acid were the two analogues possessing activity comparable to Zanamivir, showing potent inhibition of influenza virus sialidases and good antiviral activity in vitro. PMID- 11278041 TI - Cyclic amide derivatives as potential prodrugs II: N-hydroxymethylsuccinimide- / isatin esters of some NSAIDs as prodrugs with an improved therapeutic index. AB - Ester prodrugs of aspirin 1a, ibuprofen 1b, naproxen 1c and indomethacin 1d were synthesized using N-Hydroxymethylsuccinimide (HMSI) 3 and N-hydroxymethylisatin (HMIS) 4 as promoieties to reduce their gastrointestinal toxicity and improve bioavailability. Additionally, the kinetics of hydrolysis of the synthesized prodrugs 5a-d and 6a-d were studied at 37 degrees C in non-enzymatic simulated gastric fluid (SGF; hydrochloric acid buffer pH = 1.2); 0.02 M phosphate buffer (pH = 7.4); 80% human plasma and 10% rat liver homogenate. The results indicate higher chemical stability of the ester prodrugs in non-enzymatic SGF (t(1/2) congruent with 6.5-18.6 h) and rapid conversion to the parent drugs in 80% human plasma (t(1/2) congruent with 11.4-235 min) as well as in 10% rat liver homogenates (t(1/2) congruent with 12.0-90.0 min). As a general pattern, the HMSI esters 5a-d revealed higher chemical stability than the corresponding HMIS analogues 6a-d. The pH-rate profile of 5c and 6a indicated maximum stability of the former at pH = 1.2-8.0 and of the latter at pH = 1.2-4.0. The distribution coefficient (D(7.4)) values of the prodrugs 5a-d, 6a-d and the parent drugs 1a-d in an n-octanol/phosphate buffer (pH = 7.4) system indicated enhanced lipophilic properties of the prodrugs. Furthermore, the HMIS ester prodrugs 6a-d are more lipophilic than the corresponding HMSI derivatives 5a-d. In vivo ulcerogenicity studies using scanning electron microscopy on stomach specimens of rats treated with an oral dose for 4 d revealed that the synthesized ester prodrugs are significantly less irritating to gastric mucosa than the parent drugs. These results suggested HMSI and/or HMIS esters possess good potential as prodrugs with an improved therapeutic index for oral delivery of NSAIDs. PMID- 11278043 TI - Synthesis and anti-Pneumocystis carinii pneumonia activity of novel dicationic dibenzothiophenes and orally active prodrugs. AB - Dicationic carbazoles have been found to be highly active against a rat model of Pneumocystis carinii pneumonia (PCP). Unfortunately, amidoxime derivatives, designed as prodrugs, were inactive against PCP even though the corresponding amidines were highly active. In the present work, a series of 2,8- and 3,7-bis cationic dibenzothiophenes was synthesized and assayed for anti-PCP activity. Three of the compounds proved to be more potent and less toxic than a standard anti-PCP drug (pentamidine) when given intravenously. Unlike the carbazoles, a dibenzothiophene amidoxime prodrug given orally reduced the parasite load by more than 99%. While no quantitative correlation was seen between anti-PCP activity and DNA binding, a strong level of DNA binding was found to be necessary for antimicrobial activity. PMID- 11278044 TI - Carbonic anhydrase inhibitors. Part 60(#). The topical intraocular pressure lowering properties of metal complexes of a heterocyclic sulfonamide: influence of the metal ion upon biological activity. AB - Metal complexes of a sulfonamide possessing strong carbonic anhydrase (CA) inhibitory properties have been obtained from the sodium salt of the sulfonamide or from the free sulfonamide in the presence of ammonia, and the following metal ions: Mg(II); Zn(II); Mn(II); Cu(II); Co(II); Ni(II); Be(II); Cd(II); Pb(II); Al(III); Fe(III) and La(III). The original sulfonamide, 5-(3,4 dichlorophenylureido)-1,3,4-thiadiazole-2-sulfonamide and its complexes were assayed for in vitro inhibition of three CA isozymes, CA I, II and IV, some of which play a critical role in ocular fluid secretion. All these compounds (the sulfonamide and its metal complexes) behave as very powerful inhibitors against the three investigated CA isozymes. The parent sulfonamide possesses strong topical pressure lowering effects in rabbits when applied as a 1% solution directly into the eye, but some of its metal complexes, such as the Mg(II); Zn(II); Mn(II) and Cu(II) derivatives, lower intraocular pressure (IOP) in experimental animals much better. Ex vivo data showed a 98.5-99.9% inhibition of CA II and IV in ocular fluids and tissues of the rabbits treated with these agents, proving that the IOP lowering properties are due to CA inhibition. The influence of the different metal ions upon the efficiency of the obtained complexes as pressure lowering drugs are discussed, considering the possibility to design in this way more selective pharmacological agents from this class. PMID- 11278045 TI - Novel anthraquinone derivatives with redox-active functional groups capable of producing free radicals by metabolism: are free radicals essential for cytotoxicity? AB - The mode of action of antitumour anthraquinone derivatives (i.e. mitoxantrone) is not clearly established yet. It includes, among others, intercalation and binding to DNA, bioreduction and aerobic redox cycling. A series of anthraquinone derivatives, with potentially bioreducible groups sited in the side chain, have been synthesized and biologically evaluated. Their redox and cytotoxic activities were screened. Derivatives which bear a 2-(dimethylamino)ethylamino substituent, known to confer high DNA affinity, demonstrated cytotoxicity but not redox activity (beside the anthraquinone reduction). Conversely, derivatives which showed redox activity were not cytotoxic toward the P388 cell line. The results suggest that bioreduction is not the main mode of action in the cytotoxicity of anthraquinones. PMID- 11278046 TI - Synthesis and evaluation of the in vivo trypanocidal activity of water soluble organotin compounds. AB - A series of (3-(2-methoxy)ethoxypropyl)tin derivatives were synthesized as potential trypanocidal drugs. The series included an alkyltin trichloride, a dialkyltin dichloride and the corresponding dialkyltin oxide, and six dialkyltin dithio derivatives. Compounds were evaluated for trypanocidal activity using in vitro cultures of Trypanosoma equiperdum and mice infected with the same strain of parasite for in vivo tests. Two of the title derivatives, the bis (3-(2 methoxy)ethoxypropyl)tin dichloride 2 and the corresponding bis (3-(2 methoxy)ethoxypropyl)tin oxide 3, appeared to be water soluble reagents. Furthermore, they are the first examples of organotin compounds presenting interesting in vivo trypanocidal activity. PMID- 11278047 TI - Synthesis and anti-HIV activity of alpha-thiophenoxy-hydroxyethylamide derivatives. AB - A series of new anti-HIV derivatives containing a novel alpha thiophenoxyhydroxyethylamide core have been synthesized, using S phenylbenzenethiosulfonate as the thiosulfenylating reagent. Some of the new synthesized compounds (1a, 1c, 1g, 1i, 1j and 1l) inhibited HIV replication in cell culture assays (syncytia formation) with effective concentrations (EC(50)) ranging from 0.1-1 microM. Incorporation of thiophenoxy substitution within various pseudomimetic peptide backbones provided a series of highly potent HIV inhibitors. PMID- 11278048 TI - In vitro antimicrobial activity of aromatic diamidines and diimidazolines related to pentamidine. AB - Geometric isomers of pentamidine analogues were screened for antimicrobial activity in vitro. cis isomers demonstrated good antibacterial activity compared to their trans counterparts. Both isomers were moderately active against opportunistic pathogens that afflict AIDS patients with minimum inhibitory concentrations in the range of 3.12-12.5 microg/mL. PMID- 11278049 TI - On the formation of steroidal amidoesters of 4-[N,N-bis(2 chloroethyl)amino]benzoic acid and their cytotoxic activity. AB - The 4-[N,N-bis(2-chloroethyl)amino]benzoate of 17beta-acetamido-5alpha-androstan 3beta-ol, 17beta-acetamido-5-androsten-3beta-ol, 3beta-acetamido-5alpha-androstan 17beta-ol and 3alpha-acetamido-5beta-androstan-17beta-ol have been prepared and their antineoplastic effect evaluated against MIA Pa-Ca-2 pancreatic carcinoma, T47D breast carcinoma and A431 squamus cell carcinoma. Among the compounds tested, the compound 17beta-acetamido-3beta-hydroxy-5-androsten-4-[N, N-bis(2 chloroethyl)amino]benzoate appeared to possess a significant cytotoxic effect against A431 cells. PMID- 11278050 TI - Symmetry-based inhibitors of HIV-1 protease. Design, synthesis and preliminary structure-activity studies of acylated 2,3-diamino-1-hydroxypropanes and 2,4 diamino-1-hydroxybutanes. AB - Two series of peptidomimetics containing a novel C(2) pseudosymmetrical hydroxyalkyldiamino core structure were prepared from amino acid starting materials and tested for inhibitory activity against the HIV-1 protease (HIV-1 Pr) and the virus in cell culture. In the 2,3-diamino-1-hydroxypropane series, compound 6a, containing P1/P1(I) benzyl and P2/P2(I) Fmoc substituents, displayed modest HIV-1 Pr inhibition (IC(50) = 430 nM). The corresponding 2,4-diamino-1 hydroxybutane derivative (6b) was the best inhibitor of the series (IC(50) = 160 nM). Interestingly, 6a and 6b showed satisfactory inhibition of HIV replication in cell culture (ED(50) = 340 and 110 nM, respectively), a result which suggests good cell membrane penetration by this class of compounds. PMID- 11278051 TI - Synthesis and study of some 4-aza and 17a-azasteroidal isoxazoles. AB - 4-Aza-5 alpha-androstano[2,3-d]isoxazole 4 and 17a-aza-D-homo-5-androsteno[17,16 c]isoxazole 7 have been prepared by treating alpha,beta-unsaturated-beta chloroaldehydes 3 and 6 [1, 2] with hydroxylamine hydrochloride in pyridine. [16,17-d]Isoxazole derivatives 4 and 8 were found to be unstable in most of the organic solvents. beta-Cyanolactam derivatives 5 and 9 have also been prepared in both the series. PMID- 11278052 TI - Benzimidazole condensed ring systems. XI. Synthesis of some substituted cycloalkyl pyrido[1,2-a]benzimidazoles with anticipated antineoplastic activity. AB - As part of a research project on the synthesis of a number of pyrido[1,2 a]benzimidazole derivatives with possible antineoplastic activity, and as a result of the interesting antineoplastic activity recorded for one such compound (NSC 649900), some new cycloalkylpyrido[1,2-a]benzimidazoles were prepared and evaluated for such activity. Compound (7c, NSC 682011) exhibited a good in vitro antineoplastic activity especially against most of the leukaemia cell lines. This compound has been selected by the NCI for further testing in a new in vivo anticancer hollow fibre assay. PMID- 11278053 TI - Anatomical and physiological parameters affecting gastrointestinal absorption in humans and rats. AB - Anatomical and physiological parameters of the gastrointestinal (GI) tract dramatically affect the rate and extent of absorption of ingested compounds. These parameters must be considered by nutritionists, pharmacologists and toxicologists when describing or modeling absorption. Likewise, interspecies extrapolation (e.g. from rat to human) requires species-to-species comparison of these parameters. The present paper (1) describes the alimentary canal and the barrier to absorption; (2) relates the major sites of absorption; (3) compares the dimensions and surface areas of human and rat intestinal tracts; (4) discusses motility of the gut and transit times through regions of the alimentary canal; (5) explains how luminal contents are altered by physical, chemical and metabolic processes; and (6) describes the flow of blood and lymph from the GI tract to the systemic circulation, including the enterohepatic circulation. Despite strong morphological similarities between humans and rats at the microscopic level, gross anatomical differences in the relative absorptive surface areas provide a basis for concluding that the human GI tract is capable of absorbing materials faster and to a greater extent than that of the rat. Differences in the environment of the GI lumen of the two species make it possible to infer which substances are more likely to be present in a dissolved/non-ionized state for each species. Taken together, these differences may be of sufficient magnitude to alter the assessment of risks/benefits for a given compound when those risks/benefits are based on interspecies extrapolations. PMID- 11278054 TI - Antioxidant activity of a Rhus verniciflua Stokes ethanol extract. AB - A fractionated ethanol extract derived from Rhus Verniciflua Stokes (RVS) was assessed in both organic and aqueous media for the purpose of characterizing the mechanisms of antioxidant activity. RVS, an indigenous plant to Korea, was initially extracted with ethanol and characterized to contain a 90 KDa-ABTS reactive protein possessing 0.662 ng/mg copper. This characterization suggested that a primary component of RVS was Laccase, an oxidase enzyme complex. RVS exhibited a significant (P < 0.01) concentration-dependent inhibition of linoleic acid oxidation in an emulsion system up to 48 hours of incubation. Free radical scavenging activity of both a stable radical (e.g DPPH) and hydroxyl (e.g. *OH) radical followed a concentration-dependent pattern in different model systems. Using a liposome model with peroxyl radicals generated by AAPH, a significant extension of both the lag phase and a reduction of peak propagation of peroxyl radicals by RVS over a concentration range of 1 to 10 microg/ml was observed. RVS ethanol extract was also found to protect human low-density lipoprotein (LDL) from oxidative modification, mediated by cupric ion at 37 degrees C. Finally, RVS was found to be effective at protecting against plasmid DNA strand breakage induced by peroxyl free radicals in an aqueous medium. Our findings show that the ethanol fraction derived from RVS contained significant antioxidant activity in both polar and non-polar mediums. PMID- 11278055 TI - DNA strand breakage and oxygen tension: effects of beta-carotene, alpha tocopherol and ascorbic acid. AB - DNA damage is involved in carcinogenesis, aging and other degenerative diseases. The relationship between DNA strand breakage and beta-carotene (0.1-1.6 microM) was examined under different O(2) tensions and with other antioxidants: alpha tocopherol (5-80 microM), ascorbic acid (10-160 microM) and mixtures of these antioxidants. Supercoiled plasmid DNA pBR322 was incubated with 2,2'-azobis (2 amidinopropane) dihydrochloride (AAPH) to induce DNA strand breaks in the presence of antioxidants under 15, 150, and 760 torr of O(2) tension. Under 15 torr of O(2) tension, beta-carotene, alpha-tocopherol, ascorbic acid and mixtures of these antioxidants provided a dose-dependent protection against AAPH-induced DNA strand breaks. The best protection was achieved in the mixture of antioxidants. Under 150 torr of oxygen tension, the antioxidant effect of beta carotene was diminished at > or = 0.8 microM. A prooxidant effect was found at 0.8 > or = microM beta-carotene, producing more single- and double-strand breaks. alpha-Tocopherol and ascorbic acid exhibited dose-dependent antioxidant effects at 150 torr of oxygen tension. Under 760 torr of O(2) tension, the prooxidant effect of 0.8 microM beta-carotene was significant, causing supercoiled DNA to completely breakdown to circular and linear forms. In addition, 760 torr of O(2) tension attenuated the antioxidant effects of alpha-tocopherol and ascorbic acid. Thus, beta-carotene causes concentration-dependent DNA breakdown at high O(2) tension. The protection of DNA from the prooxidant effects of beta-carotene afforded by alpha-tocopherol and/or ascorbic acid was limited at high O(2) tension. PMID- 11278057 TI - Effects of chlorpropham (CIPC) on the hemopoietic system of rats. AB - Male F344 rats were given 0 or 3% chlorpropham in the diet and at 2, 4, 6, 8 or 13 weeks of administration, five rats in each group were examined for hematology, plasma clinical chemistry and pathology. Marked splenomegaly and hepatomegaly were observed in treated rats at 2-13 weeks of administration. Red blood cell counts, hemoglobin concentration, packed cell volume and platelet counts were significantly decreased and methemoglobin level, mean corpuscular volume and white blood cell counts were significantly increased in treated rats at 2-13 weeks of administration. The covalent binding of m-chloraniline m-CA, (the hydrolytic metabolite of chlorpropham) was observed in hemoglobin or splenic protein of treated rats, but only small amounts of free m-CA were present in blood or spleen. Congestion, hemosiderin deposits, extramedullary hemopoiesis and lymphoid atrophy in spleen and hyperplasia of hemopoietic cells in bone marrow were observed in treated rats at 2-13 weeks and fibrosis in splenic capsule were observed in treated rats at 4-13 weeks. The pathological changes in spleen rather than hematological changes progressed during administration, suggesting splenotoxicity of CIPC in rats. PMID- 11278056 TI - Teratological studies in defatted jojoba meal-supplemented rats. AB - To look for possible developmental effects in the offspring of jojoba meal treated Wistar rats, and to distinguish between the effects of reduced food intake and the specific developmental effects of jojoba meal itself, mated female rats were divided into three groups of 20 rats. They received during gestation: (a) normal rodent food (control group); (b) normal rodent food supplemented with 3% defatted jojoba meal (jojoba group); or (c) normal rodent food pair-fed with the jojoba group (pair-fed group). The jojoba meal group showed approximately 30% inhibition of food intake. Ten rats from each group were killed on gestation day 21. Compared to the control group, foetal body weight was reduced in both the jojoba and pair-fed groups, with a greater reduction in the jojoba group. Skeletal ossification was retarded to the same extent in both the jojoba and pair fed groups. The other 10 rats from each group were left to produce litters. Compared with controls, the body weight of the pups was lower in both the jojoba and pair-fed groups; the reduction was slightly greater in the jojoba group, but this difference disappeared after 1 week. The offspring showed no other abnormalities and reproduced normally. We conclude that, at the dose used, the retardation in foetal skeletal ossification, induced by jojoba meal supplementation during gestation, is due to food intake inhibition. Moreover, the lower birth weight of the young of jojoba-treated dams compared with the pair-fed group is merely due to a lower body weight gain during gestation. PMID- 11278058 TI - Disposition, accumulation and toxicity of iron fed as iron (II) sulfate or as sodium iron EDTA in rats. AB - A study was performed to provide data on the disposition, accumulation and toxicity of sodium iron EDTA in comparison with iron (II) sulfate in rats on administration via the diet for 31 and 61 days. Clinical signs, body weights, food consumption, food conversion efficiency, hematology, clinical chemistry and pathology of selected organs were used as criteria for disclosing possible harmful effects. Determination of iron and total iron binding capacity in blood plasma and non-heme iron analysis in liver, spleen and kidneys were used to assess the disposition and accumulation of iron originating from sodium iron EDTA or iron (II) sulfate. It was concluded that, under the conditions of the present study, iron is accumulated from the diet in liver, spleen and kidneys in a dose dependent manner, and iron derived from FeEDTA is taken up and/or accumulated less efficiently in liver and spleen than iron from FeSO(4). Moreover, feeding iron up to 11.5 and 11.2 mg/kg body weight/day, derived from FeSO(4) and FeEDTA, respectively, did not result in tissue iron excess nor in any other toxicologically significant effects. PMID- 11278059 TI - Changes in susceptibility to acetaminophen-induced liver injury by the organic anion indocyanine green. AB - The non-metabolizable organic anion indocyanine green (ICG) has been shown previously to reduce markedly the biliary secretion of acetaminophen, particularly the glutathione conjugate of APAP (APAP-GSH), suggesting that this APAP metabolite may compete with other xenobiotics for excretion into the bile via a canalicular organic anion transport process. This study was conducted to determine whether changes in the biliary disposition of APAP induced by ICG could lead to alterations in susceptibility to APAP hepatotoxicity. To investigate this, groups of overnight-fasted male CD-1 mice received 30 micromol ICG/kg, intravenously, immediately prior to APAP dosing (500 mg/kg, ip). Controls were given propylene glycol vehicle. Mice were killed at 4 h after APAP challenge for immunochemical analysis of cytosolic protein arylation and determination of non protein sulfhydryl (NPSH) depletion, or at 12 and 24 h for biochemical and histological assessment of liver injury. Elevated plasma sorbitol dehydrogenase activity and centrilobular hepatocellular necrosis was present in control mice receiving APAP at 12 and 24 h. Treatment with ICG did not alter susceptibility to APAP toxicity when measured at 12 h after challenge. However, the severity of histologic lesions in the ICG-APAP group was significantly lower at 24 h after challenge. Furthermore, treatment with ICG did not alter APAP-induced glutathione depletion or cytosolic protein arylation. These data suggest that the organic anion ICG has a protective effect on APAP toxicity that promotes a faster recovery from liver injury. PMID- 11278061 TI - Insecticidal and vertebrate toxicity associated with ethnobotanicals used as post harvest protectants in Ghana. AB - Six plant species (Cassia sophera, Chamaecrista nigricans, Mitragyna inermis, Ocimum americanum, Securidaca longepedunculata and Synedrella nodiflora) traditionally used in Ghana to control insect pests of stored grain and legumes were screened in the laboratory at three concentrations (0.5, 1 and 5%, w/w) against four common storage pests (Rhyzopertha dominica, Callosobruchus maculatus, Sitophilus zeamais and Prostephanus truncatus). All the plants showed some ability to control all or some of the test insect species. Levels of efficacy varied according to test concentration with the highest concentration tested providing the best control. The S. longepedunculata plant induced the highest percent mortality and was the best at reducing emergence of the F(1) generation. The six plants were also incorporated into standard rat diet at two concentrations (1 and 5%, w/w) and fed to rats over a 6-week period to assess potential deleterious effects against vertebrates. None of the plants demonstrated any neurotoxicological or neurobehavioural effects to the rats over the course of the trial. However, S. longepedunculata and C. nigricans caused a significant reduction in rat growth rate when incorporated at 5% in the diet, induced cell hyperplasia in the liver, and reduced the mean weight of the liver and kidneys, compared to the control group of rats. Kidney pathology was affected only by the 5% concentration of S. longepedunculata which caused a reduced accumulation of alpha2mu-globulin. The implications of these results are discussed in the context of farmer usage of insecticidal plants for stored product protection. PMID- 11278060 TI - Subclinical, non-erythematous irritation with an open assay model (washing): sodium lauryl sulfate (SLS) versus sodium laureth sulfate (SLES). AB - Compared to exaggerated hand washing procedures, an open non-exaggerated assay better approximates consumer surfactant use. Our goal was to observe skin surface modifications induced by an open test with regard to discriminating between surfactant solutions. This human in vivo assay provided information about the effect of only three washes at the laboratory and a week of at-home use. Dorsal hand and volar forearm were compared. The results demonstrated that this clinical model permits exploration of subclinical surfactant-induced irritation. Both the volar forearm and the dorsal hand are capable of discriminating between the effects of sodium lauryl sulfate (SLS) and sodium laureth sulfate (SLES). Squamometry proved to be a sensitive assessment technique for detecting surfactant-induced subclinical skin surface alterations and for differentiating surfactant effects in this open application assay, in as few as three washes. PMID- 11278062 TI - Bacterial nucleoid, DNA replication, segregation and cell division. PMID- 11278063 TI - Isolation of the Escherichia coli nucleoid. AB - Numerous protocols for the isolation of bacterial nucleoids have been described based on treatment of cells with sucrose-lysozyme-EDTA and subsequent lysis with detergents in the presence of counterions (e.g., NaCl, spermidine). Depending on the lysis conditions both envelope-free and envelope-bound nucleoids could be obtained, often in the same lysate. To investigate the mechanism(s) involved in compacting bacterial DNA in the living cell, we wished to isolate intact nucleoids in the absence of detergents and high concentrations of counterions. Here, we compare the general lysis method using detergents with a procedure involving osmotic shock of Escherichia coli spheroplasts that resulted in nucleoids free of envelope fragments. After staining the DNA with DAPI (4',6 diamidino-2-phenylindole) and cell lysis by either isolation procedure, free floating nucleoids could be readily visualized in fluorescence microscope preparations. The detergent-salt and the osmotic-shock nucleoids appeared as relatively compact structures under the applied ionic conditions of 1 M and 10 mM, respectively. RNase treatment caused no dramatic changes in the size of either nucleoid. PMID- 11278064 TI - Transcription induces a supercoil domain barrier in bacteriophage Mu. AB - In enteric bacteria, chromosomes are partitioned into domains that exhibit restricted supercoil movement. The most common domain barrier detected by gammadelta resolution assays is random with respect to sequence and occurs more frequently in cells growing rapidly in rich medium compared to cells in stationary phase. Transcription generates both positive and negative supercoiling movement. To address the question of whether transcription causes the appearance of new domain boundaries, a transcriptionally active MudI element was substituted for a MudJr-1 element that resides within the cobT gene of Salmonella typhimurium. Mu-specific transcription from the phage early promoter was placed under control of either the wild type (c(+)) or the temperature-sensitive (cts62) repressor. Using a resolution assay with res sites at six chromosomal locations, domain structure was normal in cells carrying the MudAr-1 prophage with a wild type Mu repressor. However, in cells with a MudAr-1 prophage harboring the cts62 repressor, a new domain barrier appeared in > 90% of the cells. Supercoil movement was restricted ahead of but not behind the transcription machinery. We conclude that the strong Mu early promoter induces the appearance of a domain barrier within the limits of a MudAr-1 prophage. PMID- 11278065 TI - Polarization of the Escherichia coli chromosome. A view from the terminus. AB - The E. coli chromosome replication arms are polarized by motifs such as RRNAGGGS oligomers, found preferentially on leading strands. Their skew increases regularly from the origin to dif (the site in the center of the terminus where chromosome dimer resolution occurs), to reach a value of 90% near dif. Convergent information indicates that polarization in opposite directions from the dif region controls tightly the activity of dif, probably by orienting mobilization of the terminus at cell division. Another example of polarization is the presence, in the region peripheral to the terminus, of small non-divisible zones whose inversion interferes with spatial separation of sister nucleoids. The two phenomena may contribute to the organization of the Ter macrodomain. PMID- 11278066 TI - DNA degradation in the terminus region of resolvase mutants of Escherichia coli, and suppression of this degradation and the Dif phenotype by recD. AB - We recently proposed that guillotining of dimer chromosomes occurs at cell division in resolvase mutants of Escherichia coli. This was based on the abnormal pattern of cell division observed in 10-14% of the cells in microcolonies of xerC, xerD and dif mutants. A prediction of this guillotining is that DNA degradation should occur in the terminus region, in the vicinity of the dif locus. We have tested this by DNA-DNA hybridization and have observed that dif was absent in about 22% of the chromosomes in exponentially growing xerC mutants. A locus 206 kb from dif was not affected by this degradation. We have also observed that degradation did not occur in xerC recD mutants, and that the low efficiency of plating associated with the Dif phenotype was suppressed in this strain. A model is proposed in which rapid degradation of the terminus region does not occur in recD mutants following guillotining, and that this permits the initiation of repair of broken dimer chromosomes prior to completion of cell division. PMID- 11278067 TI - The form of chromosomal DNA molecules in bacterial cells. AB - The circular concept of the bacterial chromosome was based initially on experiments involving conjugation mapping and autoradiographic imaging of DNA. This view was then supported by DNA fragment mapping, genome sequencing, and the analysis of linear DNA produced by a single cleavage of chromosomal DNA. A circular chromosome is also indicated by the existence of a mechanism for segregating dimeric chromosomes produced by recombination and the replication of DNA on both sides of the replication terminus. The evidence for circularity is reviewed here and found to be compatible with either a circular or a linear chromosomal DNA molecule. Moving pictures of ethidium-stained DNA revealed most chromosomal DNA as a rosette form with loops emanating from a dense node or as a network of strands lacking a node. This description applies to Escherichia coli, Agrobacterium tumefaciens, Pyrococcus endeavorii, Vibrio cholerae, and both the linear-mapping chromosome of Streptomyces lividans and its circular-mapping derivative. Networks without nodes were found for two linear-mapping Borrelia species. For the E. coli chromosome, open-form circles of various sizes were found only at extremely low frequency. The node of the rosette was reduced in size or eliminated in recA mutants, as well as by treatment with either ribonuclease, topoisomerase IV, 1 M NaCl, or lysozyme. A model is presented for the bacterial chromosome in which the DNA is compacted by many points of strand association (including recombination junctions, tangles and knots) created during the repair of DNA damage that occurs many times in each chromosome replication cycle. PMID- 11278068 TI - Chromosome separation and segregation in dinoflagellates and bacteria may depend on liquid crystalline states. AB - The patterns characteristic of certain liquid crystals called 'twisted nematics' or 'cholesterics' have been observed in thin sections of both dinoflagellates and bacterial chromosomes. These liquid crystals have also been obtained in vitro in concentrated DNA solutions. A large part of DNA in prokaryotic chromosomes forms such a twisted liquid crystal, whilst the remainder consists of lateral loops and is less concentrated. These semi-ordered phases could help chromosome separation to occur during and after DNA replication. We suggest that, owing to chemical differences, one of the two replicated filaments is immiscible with the rest of DNA in this chromosome. This immiscibility occurs in the context of an ordered liquid, with the DNA closely layered by a regular twist, a situation proposed to strongly minimize entangling after replication and hence to facilitate segregation. PMID- 11278069 TI - HU-GFP and DAPI co-localize on the Escherichia coli nucleoid. AB - The heterodimeric HU protein, one of the most abundant DNA binding proteins, plays a pleiotropic role in bacteria. Among others, HU was shown to contribute to the maintenance of DNA superhelical density in Escherichia coli. By its properties HU shares some traits with histones and HMG proteins. More recently, its specific binding to DNA recombination and repair intermediates suggests that HU should be considered as a DNA damage sensor. For all these reasons, it will be of interest to follow the localization of HU within the living bacterial cells. To this end, we constructed HU-GFP fusion proteins and compared by microscopy the GFP green fluorescence with images of the nucleoid after DAPI staining. We show that DAPI and HU-GFP colocalize on the E. coli nucleoid. HU, therefore, can be considered as a natural tracer of DNA in the living bacterial cell. PMID- 11278070 TI - Genome organisation and chromatin structure in Escherichia coli. AB - We have analysed the complete sequence of the Escherichia coli K12 isolate MG1655 genome for chromatin-associated protein binding sites, and compared the predicted location of predicted sites with experimental expression data from 'DNA chip' experiments. Of the dozen proteins associated with chromatin in E. coli, only three have been shown to have significant binding preferences: integration host factor (IHF) has the strongest binding site preference, and FIS sites show a weak consensus, and there is no clear consensus site for binding of the H-NS protein. Using hidden Markov models (HMMs), we predict the location of 608 IHF sites, scattered throughout the genome. A subset of the IHF sites associated with repeats tends to be clustered around the origin of replication. We estimate there could be roughly 6000 FIS sites in E. coli, and the sites tend to be localised in two regions flanking the replication termini. We also show that the regions upstream of genes regulated by H-NS are more curved and have a higher AT content than regions upstream of other genes. These regions in general would also be localised near the replication terminus. PMID- 11278071 TI - DNA supercoiling and transcription in Escherichia coli: The FIS connection. AB - The nucleoid-associated protein FIS modulates the topology of DNA in a growth phase dependent manner functioning homeostatically to counteract excessive levels of negative superhelicity. We propose that this is achieved by at least two mechanisms: the physical constraint of low levels of negative superhelicity by FIS binding to DNA and by a reduction in the expression and effectiveness of DNA gyrase. In addition, high levels of expression of the fis gene do themselves require a high negative superhelical density. On DNA substrates containing phased high affinity binding sites, as exemplified by the upstream activating sequence of the tyrT promoter, FIS forms tightly bent DNA structures, or microloops, that are necessary for the optimal expression of the promoter. We suggest that these microloops compensate in part for the FIS-induced lowering of the superhelical density. PMID- 11278072 TI - Roles of Escherichia coli histone-like protein HU in DNA replication: HU-beta suppresses the thermosensitivity of dnaA46ts. AB - The HU protein is a small, basic, heat-stable DNA-binding protein that is well conserved in prokaryotes and is associated with the bacterial nucleoid. In enterobacteria, including Escherichia coli, HU is a heterotypic dimer, HUalphabeta, composed of two closely related sub-units encoded by the hupA and hupB genes, respectively. HU was shown to participate in vitro in the initiation of DNA replication as an accessory factor to assist the action of DnaA protein in the unwinding of oriC DNA. To further elucidate the role of HU in the regulation of the DNA replication initiation process, we tested the synchrony phenotype in the absence of either one or both HU sub-units. The hupAB mutant exhibits an asynchronous initiation, the hupA mutant shows a similar reduced synchrony, whereas the hupB mutant shows a normal phenotype. Using a thermosensitive dnaA46 strain (dnaA46ts), an initiation mutant, we reveal a special role of HUbeta. The presence of a plasmid overproducing HUbeta in a dnaA46ts lacking HU (hupAB background) compensates for the thermosensitivity of this initiation mutant. Moreover, the overproduction of HUbeta confers to dnaA46ts a pattern of asynchrony similar to that of a dnaAcos, the intragenic suppressor of dnaA46ts. We show that the relative ratio of HUalpha versus HUbeta is greatly perturbed in dnaA46ts which accumulates little, if any, HUbeta. Therefore, the suppression of thermosensitivity in dnaA46hupAB by HUbeta may be caused by an unexpected absence of HUbeta in the dnaA46ts mutant. Visibly the HU composition is sensitive to the different states of DnaA, and may play a role during the regulation of the initiation process of the DNA replication by affecting subsequent events along the cell cycle. PMID- 11278073 TI - Structural basis for preferential binding of H-NS to curved DNA. AB - The Escherichia coli H-NS protein is a nucleoid-associated protein involved in transcription regulation and DNA compaction. H-NS exerts its role in DNA condensation by non-specific interactions with DNA. With respect to transcription regulation preferential binding sites in the promoter regions of different genes have been reported. In this paper we describe the analysis of H-NS-DNA complexes on a preferred H-NS binding site by atomic force microscopy. On the basis of these data we present a model for the specific recognition of DNA by H-NS as a function of DNA curvature. PMID- 11278074 TI - H-NS and H-NS-like proteins in Gram-negative bacteria and their multiple role in the regulation of bacterial metabolism. AB - In Escherichia coli, the H-NS protein plays an important role in the structure and the functioning of bacterial chromosome. A homologous protein has also been identified in several enteric bacteria and in closely related organisms such as Haemophilus influenzae. To get information on their structure and their function, we identified H-NS-like proteins in various microorganisms by different procedures. In silico analysis of their amino acid sequence and/or in vivo experiments provide evidence that more than 20 proteins belong to the same class of regulatory proteins. Moreover, large scale technologies demonstrate that, at least in E. coli, the loss of motility in hns mutants results from a lack of flagellin biosynthesis, due to the in vivo repression of flagellar gene expression. In contrast, several genes involved in adaptation to low pH are strongly induced in a H-NS deficient strain, resulting in an increased resistance to acidic stress. Finally, expression profiling and phenotypic analysis suggest that, unlike H-NS, its paralogous protein StpA does not play any role in these processes. PMID- 11278076 TI - Mutagenesis of the downstream region of the Escherichia coli hns promoter. AB - The promoter of hns, the structural gene for the abundant nucleoid-associated protein H-NS of Escherichia coli, contains, downstream of the initiation site, two four bp-long 'CG clamps', one of which overlaps the potential target sequence (CCAAT) of CspA, the cold-shock transcriptional enhancer of this gene. To establish the role of these potential regulatory signals during the cold-shock activation of hns, the CCCCAAT sequence has been subjected to mutagenesis, weakening the strength of the CG clamp and scrambling or inverting the CCAAT sequence. The resulting mutated hns promoters were placed in front of a reporter gene (cat) and their activity was studied in cells subjected to cold-shock under conditions where the increase in the concentration of CspA is either large or small. Our results allow us to conclude that although not essential, the CCCCAAT sequence, mainly due to the presence of the CG clamp, may play an important role in the CspA-mediated regulation of hns expression at both transcriptional and translational levels. PMID- 11278075 TI - The nucleoid-associated protein StpA binds curved DNA, has a greater DNA-binding affinity than H-NS and is present in significant levels in hns mutants. AB - The StpA protein is closely related to H-NS, the well-characterised global regulator of gene expression which is a major component of eubacterial chromatin. Despite sharing a very high degree of sequence identify and having biochemical properties in common with H-NS, the physiological function of StpA remains unknown. We show that StpA exhibits similar DNA-binding activities to H-NS. Although both display a strong preference for binding to curved DNA, StpA binds DNA with a four-fold higher affinity than H-NS, with K(d)s of 0.7 microM and 2.8 microM, respectively. It has previously been reported that expression of stpA is derepressed in an hns mutant. We have quantified the amount of StpA protein produced under this condition and find it to be only one-tenth the level of H-NS protein in wild-type cells. Our findings explain why the presence of StpA does not compensate for the lack of H-NS in an hns mutant, and why the characteristic pleiotropic hns mutant phenotype is observed. PMID- 11278077 TI - The GemA protein of phage Mu and the GyrB gyrase subunit of Escherichia coli: the search for targets and interactions leading to the reversion of Mu-induced mutations. AB - The mutant bacteriophage Mugem2(Ts), known to synchronize the division of infected cells, to relax DNA supercoiling and, as prophage, to give rise to precisely excised revertants, has been thought to overexpress the gemA-mor operon, and genetic evidence suggests that the B subunit of DNA gyrase (GyrB) is the target of action of GemA. In two different double hybrid tests presented here, we find no evidence of GemA-GyrB protein-protein interaction. We do observe a GemA-GemA interaction, however, indicating that GemA can dimerize. In lacZ::Mu lysogens, overexpression of the gemA-mor operon from a plasmid, under control of the L-arabinose inducible p(araBAD) promoter, does not permit the recovery of Lac(+) revertants. These observations suggest that GyrB is not the direct target of GemA action and that the various phenotypes of Mugem2(Ts) are not caused by overexpression of the gemA-mor operon. PMID- 11278078 TI - A possible role for L24 of Bacillus subtilis in nucleoid organization and segregation. AB - The condensation of DNA in bacterial nucleoids during cell cycle is a complex and dynamic process. Proteins displaying the physico-chemical properties of histones are known to contribute to this process. During a search for B. subtilis nucleoid associated proteins, HBsu and L24 were identified as the most abundant proteins in nucleoid containing fractions. Purified L24 binds and condenses DNA in vitro. In this paper we describe immunofluorescence studies that demonstrated that L24 is located at the poles of the nucleoids in exponentially growing cells. In contrast, the protein is dispersed in the cytoplasm during stationary phase. Moreover, overexpression of the rplX gene encoding L24 disrupts nucleoid segregation and positioning. PMID- 11278079 TI - Molecular components of the archaeal nucleosome. AB - Here we describe the organization of the archaeal nucleosome, in which four archaeal histones are circumscribed by approximately 80 bp of DNA. Through a combination of sequence comparisons, 3D structural studies, site-directed mutagenesis and assays for DNA binding, we have assigned functions to most of the individual residues in the histone fold of the representative archaeal histone rHMfB. By SELEX selection, the sequences of DNA molecules that are most readily bound and wrapped by rHMfB into archaeal nucleosomes in vitro have been identified, and these define DNA structures that position archaeal nucleosome assembly. PMID- 11278082 TI - Franz Kuhn, his contribution to anaesthesia and emergency medicine. AB - Franz Kuhn (1866-1929), a German surgeon, made a significant practical and scientific contribution towards the development of modern anaesthesia and emergency medicine. He developed modern, scientifically based concepts in close correlation to practical inventions for every day use. All of his studies and developments were patient orientated and led to remarkable improvements in patient safety. Kuhn was a major protagonist of endotracheal intubation, perfected his flexo-metallic endotracheal tubes, worked on different techniques of intubating the trachea, applied positive pressure to the lungs during thoracic surgery and developed anaesthesia machines. In the early 20th century, he wrote several papers on this topic including a remarkable monograph, dealing with the techniques, indications in anaesthesia and emergency medicine and his experiences of endotracheal intubation. Due to a dispute with Sauerbruch on the methods of avoiding a pneumothorax during thoracic surgery and the development of local and regional anaesthesia techniques, the value of his work and his revolutionary ideas were not appreciated until 40 years later. PMID- 11278084 TI - European Resuscitation Council Guidelines 2000 for Automated External Defibrillation. A statement from the Basic Life Support and Automated External Defibrillation Working Group(1) and approved by the Executive Committee of the European Resuscitation Council. AB - The European Resuscitation Council (ERC) last issued guidelines for Automated External Defibrillators (AEDs) in 1998 [1]. The American Heart Association, together with representatives from the International Liaison Committee on Resuscitation (ILCOR), then undertook a series of evidence-based evaluations of the science of resuscitation [2] which culminated in the publication of "Guidelines 2000 for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care" in August 2000 [3,4]. The Basic Life Support and Automated External Defibrillation Working Group (BLS&AED Working Group of the ERC) has considered this document and has recommended changes in the ERC AED guidelines. These are presented in this paper. PMID- 11278083 TI - European Resuscitation Council Guidelines 2000 for Adult Basic Life Support. A statement from the Basic Life Support and Automated External Defibrillation Working Group(1) and approved by the Executive Committee of the European Resuscitation Council. AB - The European Resuscitation Council (ERC) last issued guidelines for Basic Life Support (BLS) in 1998 [1]. These were based on the "Advisory Statements" of the International Liaison Committee on Resuscitation (ILCOR) published in 1997 [2]. Following this, the American Heart Association, together with representatives from ILCOR, undertook a series of evidence-based evaluations of the science of resuscitation [3] which culminated in the publication of "Guidelines 2000 for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care" in August 2000 [4,5]. The Basic Life Support and Automated External Defibrillation Working Group (BLS&AED Group) has considered this document and has recommended changes in the ERC BLS guidelines. These are presented in this paper. PMID- 11278085 TI - European Resuscitation Council Guidelines 2000 for Adult Advanced Life Support. A statement from the Advanced Life Support Working Group(1) and approved by the Executive Committee of the European Resuscitation Council. AB - The European Resuscitation Council (ERC) last issued guidelines for Basic Life Support (BLS) in 1998 [1]. These were based on the 1997 International Liaison Committee on Resuscitation (ILCOR) Advisory Statements [2]. In 1999 and 2000 representatives of ILCOR, at the invitation of the American Heart Association, met on a number of occasions in Dallas to agree a Consensus on Science upon which future guidelines would be based. Representatives from the ERC played a prominent role in the deliberations, which culminated in the publication of "Guidelines 2000 for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care--A Consensus on Science" in August 2000 [3]. The consensus was evidence-based wherever possible. The ERC ALS Working Group has considered this document and has recommended some changes in the guidelines that will be suitable for European practice. These changes, together with a summary of the Sequence of Actions in ALS, are presented in this paper. PMID- 11278086 TI - European Resuscitation Council Guidelines 2000 for Basic Paediatric Life Support. A statement from the Paediatric Life Support Working Group and approved by the Executive Committee of the European Resuscitation Council. AB - The European Resuscitation Council (ERC) last issued guidelines for Paediatric Life Support (PLS) in 1998 [1]. These were based on the "Advisory Statements" of the International Liaison Committee on Resuscitation (ILCOR) published in 1997 [2]. Following this, the American Heart Association, together with representatives from ILCOR, undertook a series of evidence-based evaluations of the science of resuscitation which culminated in the publication of "Guidelines 2000 for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care" in August 2000 [3,4]. The Paediatric Life Support Working Party of the European Resuscitation Council has considered this document and the supporting scientific literature and has recommended changes to the ERC Basic PLS guidelines. These are presented in this paper. There have been few major changes to the ERC recommended guidelines as some of the changes agreed in "Guidelines 2000" had already been introduced into Europe subsequent to the 1998 ILCOR "Advisory Statements" (Fig. 1). PMID- 11278087 TI - European Resuscitation Council Guidelines 2000 for Advanced Paediatric Life Support. A statement from Paediatric Life Support Working Group and approved by the Executive Committee of the European Resuscitation Council. AB - The European Resuscitation Council (ERC) last issued guidelines for Paediatric Life Support (PLS) in 1998 [1]. These were based on the "Advisory Statements" of the International Liaison Committee on Resuscitation (ILCOR) published in 1997 [2]. Following this, the American Heart Association, together with representatives from ILCOR, undertook a series of evidence-based evaluations of the science of resuscitation which culminated in the publication of "Guidelines 2000 for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care" in August 2000 [3,4]. The Paediatric Life Support Working Party of the European Resuscitation Council has considered this document and the supporting scientific literature and has recommended changes to the ERC Advanced PLS guidelines. These are presented in this paper. There have been few major changes to the ERC recommended guidelines as some of the changes agreed in "Guidelines 2000" had already been introduced into Europe subsequent to the 1998 ILCOR "Advisory Statements" (Fig. 1). PMID- 11278088 TI - European Resuscitation Council Guidelines 2000 for Newly Born Life Support. A statement from the Paediatric Life Support Working Group and approved by the Executive Committee of the European Resuscitation Council. AB - The European Resuscitation Council (ERC) last issued guidelines for the resuscitation of the newly born infant in 1999 [1]. This was an "Advisory Statement" of the International Liaison Committee on Resuscitation (ILCOR). Following this, the American Heart Association and the Neonatal Resuscitation Programme Steering Committee of the American Academy of Paediatrics and representatives of the World Health Organisation, together with representatives from ILCOR, undertook a series of evidence-based evaluations of the science of resuscitation which culminated in the publication of "Guidelines 2000 for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care" in August 2000 [2,3]. The Paediatric Life Support Working Party of the European Resuscitation Council has considered this document and the supporting scientific literature and presents the ERC Newly Born Guidelines in this paper. Readers will find few changes to the ILCOR Advisory Statement recommendations as the new evidence that has emerged since its publication in 1999 has been confirmatory of the ILCOR recommendations. PMID- 11278089 TI - Update on the emergency medical treatment of anaphylactic reactions for first medical responders and for community nurses. PMID- 11278090 TI - "Probability of successful defibrillation" as a monitor during CPR in out-of hospital cardiac arrested patients. AB - The frequency spectrum of the ECG in ventricular fibrillation (VF) correlates with myocardial perfusion and might predict defibrillation success defined as return of spontaneous circulation (ROSC). The predictive power increases when more spectral variables are combined, but the complex information can be difficult to handle during the intensity of CPR. We therefore developed a method for expressing this multidimensional information in a single reproducible variable reflecting the probability of defibrillation success. This is based on the highest performing predictor for ROSC after 883 shocks given to 156 patients with VF. This was a combination of two decorrelated spectral features based on a principal component analysis of an original feature set with information on centroid frequency, peak power frequency, spectral flatness and energy. The function "Probability of defibrillation success" (P(ROSC)(v)) was developed by a 2-dimensional histogram technique. P(ROSC)(v) discriminated between shocks followed by ROSC and No-ROSC (P<0.0001). The present methodology indicates a possible way to develop a CPR monitor. PMID- 11278091 TI - Resuscitation decision index: a new approach to decision-making in prehospital CPR. AB - Retrospective and prospective studies have been undertaken to assess physicians' practice-patterns by studying cardiopulmonary resuscitation (CPR) case summaries. Most summaries reveal similar influences by the physician, patient and situation related variables on the patterns of resuscitation. The initiation of resuscitation efforts is addressed frequently, but, very few studies discuss the topic of termination of resuscitation. Prehospital emergencies are addressed very rarely. The objective of this study was to introduce a new methodological approach towards initiation and termination of resuscitation efforts in prehospital situations. The subject studied were the physicians' decisions concerning initiation/withholding, termination/withdrawal and the resulting early survival rates. The result is termed the "Resuscitation decision index" (RDI). The "RDI" could be a tool allowing comparisons on a quantitative level, between different EMS systems or disciplines and giving an insight into the decision process. The "RDI" can enhance audit of resuscitation. The process of decision making can be used to help future theoretical decision-making strategies. PMID- 11278092 TI - Eliciting patient disutilities for the adverse outcomes of cardiopulmonary resuscitation. AB - BACKGROUND: in helping patients decide about treatments, such as whether to authorize cardiopulmonary resuscitation (CPR), physicians typically present information about the possible outcomes and their likelihoods. The aim of this study was to elicit patient disutilities for the adverse outcomes of cardiopulmonary resuscitation (CPR) using the methodology of NH Anderson's functional theory of cognition and to determine how patients integrate the disutility and the likelihood of an outcome. METHODS: 77 French adults rated scenarios of possible outcomes of CPR on a linear scale with anchors "what would be the best (or worst) for me." In 25 of the 27 scenarios, the result would be either total recovery or one of five adverse outcomes (chest injury, mild reversible brain damage, severe irreversible brain damage, death after intensive care, immediate death) with one of five likelihoods (one to five chances out of ten). In the other two, the only possible result was either total recovery or immediate death. RESULTS: the mean disutilities relative to 0 for chest injury and 100 for severe brain damage were 13 for mild brain injury, 68 for death after intensive care, and 69 for immediate death. The graphs of the ratings of each adverse outcome in relation to its frequency were fan-shaped, showing that participants integrated this information multiplicatively. CONCLUSIONS: the functional theory of cognition provides an alternate method of eliciting patient utilities for the outcomes of CPR and supports clinicians' assumption that people combine utility and likelihood multiplicatively. PMID- 11278093 TI - Causes of airway obstruction during cuffed oropharyngeal airway use. AB - This study investigated the cause for needing airway maneuvers to maintain a patent airway during the use of cuffed oropharyngeal airway (COPA). Twenty adult patients (29.4+/-6.8 years-old, ASA 1-2) scheduled for minor gynecological surgery who required brief manipulations of the airway despite COPA use following the manufacture's guidelines, were enrolled in this study. To obtain airway patency, 15 patients required only the head-tilt maneuver. In eight of the 15 patients, the laryngeal inlet was opened partially (n=4) or completely (n=4). Despite lifting the epiglottis, the laryngeal inlet was incomplete at the level of pharyngeal view. The patency of the laryngeal inlet was decided by the extent of the distance between the posterior pharyngeal wall and the lateral glossoepiglottic fold, which was made by hyoid bone. In the other seven patients, the head-tilt maneuver elevated the epiglottis and completely opened the laryngeal inlet. Five patients required both the jaw-thrust and head-tilt maneuver. Of these patients lifting the epiglottis was incomplete in three and the laryngeal inlet was partially collapsed in one even after the airway manipulations. The airways in these three patients, however, became patent after manipulations despite the persisting partial obstruction. PMID- 11278094 TI - Reducing CPR artefacts in ventricular fibrillation in vitro. AB - CPR creates artefacts on the ECG, and a pause in CPR is therefore mandatory during rhythm analysis. This hands-off interval is harmful to the already marginally circulated tissues during CPR, and if the artefacts could be removed by filtering, the rhythm could be analyzed during ongoing CPR. Fixed coefficient filters used in animals cannot solve this problem in humans, due to overlapping frequency spectra for artefacts and VF signals. In the present study, we established a method for mixing CPR-artefacts (noise) from a pig with human VF (signal) at various signal-to-noise ratios (SNR) from -10 dB to +10 dB. We then developed a new methodology for removing CPR artefacts by applying a digital adaptive filter, and compared the results with this filter to that of a fixed coefficient filter. The results with the adaptive filter clearly outperformed the fixed coefficient filter for all SNR levels. At an original SNR of 0 dB, the restored SNRs were 9.0+/-0.7 dB versus 0.9+/-0.7 dB respectively (P<0.0001). PMID- 11278095 TI - Simultaneous sternothoracic cardiopulmonary resuscitation: a new method of cardiopulmonary resuscitation. AB - No existing device for cardiopulmonary resuscitation (CPR) is designed to exploit both the "cardiac pump" and the "thoracic pump" effect simultaneously. The purpose of this study was to measure the haemodynamic effect of a new simultaneous sternothoracic cardiopulmonary resuscitation (SST-CPR) device that could compress the sternum and constrict the thoracic cavity simultaneously in a canine cardiac arrest model. After 4 min of ventricular fibrillation, 24 mongrel dogs were randomized to receive standard CPR (n=12) or SST-CPR (n=12). SST-CPR generated a new pattern of the aortic pressure curve presumed to be the result of both sternal compression and thoracic constriction. SST-CPR resulted in significantly higher mean arterial pressure than standard CPR (68.9+/-16.1 vs. 30.5+/-10.0 mmHg, P<0.01). SST-CPR generated higher coronary perfusion pressure than standard CPR (47.0+/-11.4 vs. 17.3+/-8.9 mmHg, P<0.01). End tidal CO(2) tension was also higher during SST-CPR than standard CPR (11.6+/-6.1 vs. 2.17+/ 3.3 mmHg, P<0.01). In this preliminary animal model study, simultaneous sternothoracic cardiopulmonary resuscitation generated better haemodynamic effects than standard, closed chest cardiopulmonary resuscitation. PMID- 11278096 TI - Does paddle force applied during defibrillation meet advanced life support guidelines of the European Resuscitation Council? AB - OBJECTIVES: to determine whether paddle force applied during defibrillation meets the 12 kg (approximately 120 N) force recommended by the advanced life support (ALS) guidelines of the European Resuscitation Council (ERC). MATERIALS AND METHODS: an adult mannequin was "defibrillated" using standard defibrillation paddles instrumented to measure paddle force. Paddle force was recorded at the time the discharge buttons on the paddle handles were depressed. RESULTS: 54 doctors and nurses performed simulated defibrillation on a mannequin. Median sternal paddle force was 60.6 N (range 26.1-132.8 N) and median apical paddle force was 59.5 N (range 18.6-118.5 N). Only 3/54 operators (5.6%) applied sternal paddle force equal to or in excess of ERC recommendations. No operator applied apical paddle force equal to or in excess of ERC recommendations. CONCLUSIONS: force applied to defibrillation paddles does not meet guidelines of the European Resuscitation Council. Greater emphasis during advanced life support training should be placed on the importance of firm paddle force during defibrillation. PMID- 11278097 TI - Lesion of the bulbospinal noradrenergic pathways blocks desipramine-induced inhibition of the C-fiber evoked nociceptive reflex in rats. AB - Desipramine-induced inhibition of spinal cord nociceptive transmission was studied in rats with or without lesion of the bulbospinal noradrenergic system by recording the C-fiber evoked nociceptive reflex from a hind limb. Bulbospinal noradrenergic projections were lesioned by injecting intrathecally 20 microg of 6 hydroxydopamine 2 weeks before the electrophysiological experiments. Results show that desipramine (5, 10 and 20 mg/kg intraperitoneally) produced dose-dependent inhibition of the C reflex response duration in rats having intact noradrenergic bulbospinal systems. The inhibitory effect of desipramine was reduced or even abolished in rats pre-treated with 6-hydroxydopamine. In addition, [3H] noradrenaline uptake was significantly lower in spinal cord slices arising from 6 hydroxydopamine lesioned animals, as compared to that from intact rats. These observations support the notion that the antinociceptive activity of antidepressants with noradrenergic selectivity depends on a normal rate of endogenous noradrenaline released by bulbospinal neurons. PMID- 11278098 TI - D2 dopamine receptor-G protein coupling. Cross-regulation of agonist and guanosine nucleotide binding sites. AB - The cross regulation of agonist binding to D2 dopamine receptors and guanosine nucleotide binding to G proteins was studied using membranes of Chinese hamster ovary cells expressing rat D2short dopamine receptors. All guanosine nucleotides studied caused a concentration-dependent loss of high-affinity agonist binding sites of D2 receptors with potencies corresponding to their affinity to bind to G proteins in these membranes. On the other hand, the dopaminergic agonists, but not antagonists, decreased the affinities of guanosine diphosphate and guanosine monophosphate, but not of guanosine 5'-(gamma-thiotriphosphate). The cross regulation of ligand binding to D2 dopamine receptors and G proteins suggests the existence of several conformational states of these proteins during the signal transduction and that the high-affinity state of agonist binding is a transient state of the agonist-receptor-G protein complex, where no nucleotides are bound. PMID- 11278099 TI - 5-HT7 receptor mRNA expression in human trigeminal ganglia. AB - The present study investigated, by using the method of reverse transcriptase polymerase chain reaction, whether the 5-HT7 receptor mRNA is expressed in human trigeminal ganglia. Trigeminal ganglia were excised post mortem from five human subjects. Oligonucleotide primers were selected based upon unique regions of complementary DNA sequence for the cloned human 5-HT7 receptor. Sequence analysis revealed the presence of a single message that matched the sequence of the cloned human 5-HT7 receptor. The present results may direct future efforts to determine the potential excitatory role of the 5-HT7 receptor in the neuropathological events that develop in migraine headache. PMID- 11278100 TI - Perceptual phenomena after unilateral arm amputation: a pre-post-surgical comparison. AB - Painful and non-painful phantom phenomena occur frequently after amputations but are rarely investigated in the perioperative stage. The goal of the present study was the assessment of phantom phenomena, pain and changes in primary somatosensory cortex prior to and after upper limb amputation. Two patients who suffered from metastatic carcinoma were examined 2 days prior to and 7 days after the amputation of an arm using comprehensive psychometric assessments and neuroelectric source imaging. Both patients reported phantom limb pain that was similar to their pre-amputation pain. In one patient, reorganization of the mouth area into the deafferented hand area took place immediately after the amputation. In the other patient reorganization had occurred prior to the amputation possibly related to non-use of the arm several years prior to the amputation. PMID- 11278101 TI - Induction of phosphatidylinositol 3-kinase and serine-threonine kinase-like immunoreactivity in rabbit spinal cord after transient ischemia. AB - The mechanism of spinal cord injury has been thought to be related with tissue ischemia, and spinal motor neuron cells are suggested to be vulnerable to ischemia. To evaluate the mechanism of such vulnerability of motor neurons, we attempted to make a reproducible model of rabbit spinal cord ischemia. Using this model, the inductions of phosphatidylinositol 3-kinase (PI3-k) and serine threonine kinase (Akt) were investigated with immunohistochemical analyses for up to 7 days of the reperfusion following 15 min of ischemia in rabbit spinal cord. It has been demonstrated that both PI3-k and its downstream effector, Akt mediate growth factor-induced neuronal survival. Spinal cord sections from animals sacrificed at 8 h, 1, 2, and 7 days following the 15 min of ischemia were immunohistochemically evaluated using monoclonal antibodies for PI3-k and Akt. Following the 15 min of ischemia, the majority of the motor neurons showed selective cell death at 7 days of reperfusion. Immunoreactivity of PI3-k and Akt were induced at 8 h of reperfusion selectively in motor neuron cells. No glial cells and inter neurons were stained in the spinal cord sections. The activation of PI3-k and Akt protein at the early stage of reperfusion may be one of the factors responsible for the delay in neuronal death after spinal cord ischemia. PMID- 11278102 TI - Comparative study of glutamate mediated gamma-aminobutyric acid release from nitric oxide synthase and tyrosine hydroxylase immunoreactive cells of the Cebus apella retina. AB - The effects of excitatory amino acids (EAAs) upon transporter-mediated gamma aminobutyric acid (GABA) release were investigated in cells containing tyrosine hydroxylase (TH) or nitric oxide synthase (NOS) in retina of the primate Cebus apella. Retinas were treated in vitro with 50 microM Kainate (KA) or 5 mM L Glutamate (L-Glu), for 30 min at 37 degrees C, in an Mg2+-free Locke's solution with or without Ca2+. The effects of EAAs were measured immunocytochemically by determining the GABA content in TH or NOS-immunoreactive cells in the inner retina, after stimulation. L-Glu and KA induced a Ca2+-independent GABA release from most GABA-immunoreactive cells of the inner retina. Double label experiments indicated that this release occurs in NOS+/GABA+ cells, but not in TH+/GABA+ cells suggesting that these cell subpopulations may be differentiated in some functional aspects. PMID- 11278103 TI - Endogenous acetylcholine facilitates epileptogenesis in immature rat neocortex. AB - In the presence of the gamma-amino butyric acid-A (GABAA) antagonist bicuculline methiodide (50 microM), synchronous spontaneous and evoked potentials were recorded extracellularly from the deep layers of immature neocortex (postnatal days 10-31, P10-P31) in vitro. Addition of the anticholinesterase eserine (10 microM) depressed the amplitude (by 29.5+/-6.6%, n=13) and duration (by 26.3+/ 4.7%, n=11) of the evoked field potentials in 13/19 slices (68%), and increased significantly the rates of occurrence of spontaneous epileptiform discharges or induced them in 9/19 slices (47%). All these effects were blocked by the muscarinic antagonist atropine (2.5 microM, n=3), suggesting that they were mediated by the activation of muscarinic receptors by endogenous acetylcholine. The cholinergic inhibitory effect is unlikely to terminate seizures, while the excitatory effect, could conceivably promote or aggravate their manifestation. In conclusion, these findings demonstrate that endogenous acetylcholine may contribute to epileptogenesis in immature neocortex. PMID- 11278104 TI - Effect of hypergravity on the mouse basal expression of NGF and BDNF in the retina, visual cortex and geniculate nucleus: correlative aspects with NPY immunoreactivity. AB - We investigated the effect of hypergravitation on Nerve growth factor (NGF) and Brain-derived-neurotrophic factor (BDNF) expression in the visual cortex, geniculate nucleus (GN), and retina of adult male mice. The results showed that altered gravity causes an increase in NGF and BDNF in the visual cortex and GN which resulted to be associated with an up-regulation of cells immunoreactive to neuropeptide Y (NPY) in the visual cortex and GN. We also found a decrease in NGF, BDNF, and NPY in the mouse retina exposed to hypergravity. These findings suggest that alteration in gravitational environment differentially affects local neurotrophic factors and NPY expression. The possible functional significance of these observations is discussed. PMID- 11278105 TI - Effect of AF64A, a cholinergic neurotoxin, on footshock stimulation-induced locus coeruleus excitation in rats. AB - We studied the effect of ethylcholine mustard aziridinium ion (AF64A), a cholinergic neurotoxin, on the footshock stimulation (FS)-induced excitation of the locus coeruleus (LC) neurons in rats. The FS-evoked LC excitation was significantly reduced in AF64A-treated rats, in comparison with normal rats. In particular, the early component of LC excitation was less pronounced. The number of choline acetyltransferase immunoreactive neurons in the septal complex was significantly lower than those in normal rats, except for in the ventral subgroup. These findings suggest that the cholinergic neuron system is involved in the early component of LC excitation in rats. PMID- 11278106 TI - Complement inhibition does not reduce post-hypoxic-ischemic cerebral injury in 21 day-old rats. AB - Hypoxic-ischemic (HI) cerebral injury frequently follows resuscitation and is a recognized cause of permanent long-term neurologic disability in children. Complement activation has been shown to participate in post-ischemic injury to a variety of tissues and organs. To test the hypothesis that complement activation participates in post-HI cerebral injury in immature rats, 21-day-old rats were subjected to right common carotid artery ligation and 8% O(2). This combination of ischemia and hypoxia resulted in the development of significant neuronal loss, edema, and atrophy in the right cerebral hemisphere. However, intraperitoneal administration of the complement inhibitors soluble complement receptor type 1 or cobra venom factor did not reduce the neuronal loss, edema, or atrophy. Therefore, complement activation did not contribute significantly to the cerebral injury observed in this immature rat model. PMID- 11278107 TI - Anti-apoptotic effect of trans-resveratrol on paclitaxel-induced apoptosis in the human neuroblastoma SH-SY5Y cell line. AB - Paclitaxel, an anticancer drug, induces apoptosis in human neuroblastoma cell line SH-SY5Y. The addition of trans-resveratrol, a natural antioxidant present in grapes and red wine, to SH-SY5Y cultures exposed to paclitaxel significantly reduces cellular death. The neuroprotective action of trans-resveratrol is due neither to its antioxidant capacity nor to interference with the polymerization of tubulin induced by paclitaxel. However, trans-resveratrol is able to inhibit the activation of caspase 7 and degradation of poly-(ADP-ribose)-polymerase which occur in SH-SY5Y exposed to paclitaxel. Resveratrol, therefore, exerts its anti apoptotic effect by modulating the signal pathways that commit these neuronal like cells to apoptosis. PMID- 11278108 TI - Human balance control during cutaneous stimulation of the plantar soles. AB - Previous work on human postural control of upright stance, performed in the absence of visual and vestibular orientation cues, suggests that somatosensory cues in the feet enable subjects to maintain equilibrium during low-frequency platform tilts. Here we confirm earlier studies which indicated that stimulation of plantar cutaneous mechanoreceptors can lead to postural responses. Yet, this stimulation did not modify considerably the postural reactions of normal subjects and vestibular loss patients during platform tilts. We therefore suggest that it is necessary to differentiate between (i) cues from plantar cutaneous receptors involved in exteroceptive functions, like the evaluation of the support structure or of relative foot-to-surface motion, and (ii) cues from deep receptors which subserve proprioceptive functions like the control of center of pressure shifts within the limits of the foot support base. PMID- 11278109 TI - Episodic retrieval is reflected by a process specific increase in human electroencephalographic theta activity. AB - Is an increase in theta during retrieval due (primarily) to the access of a stored code or to more general processes? The electroencephalogram was recorded while subjects performed a recognition task with pictures. According to the event related desynchronization/synchronization method, the percentage of band power changes was calculated during encoding and retrieval for a theta and three alpha bands. Significant results were obtained (with minor exceptions) only in the theta band. The increase in theta was significantly larger during retrieval than during encoding but did not differ significantly between new and successfully retrieved old pictures. Because a memory trace is lacking for new pictures, the increase in theta during retrieval reflects primarily general processing demands of a complex episodic memory system. PMID- 11278110 TI - A functional role for adenosine A3 receptors: modulation of synaptic plasticity in the rat hippocampus. AB - Adenosine modulates hippocampal synaptic plasticity, namely long-term potentiation (LTP) and long-term depression (LTD), through activation of A1 and A2A receptors. We now report a novel role for the recently described adenosine A3 receptor in the regulation of synaptic plasticity in the CA1 area of hippocampal slices. Activation of adenosine A3 receptors by (1-[2-chloro-6-[[(3 iodophenyl)methyl]amino]-9H-purin-p-yl]-1-deoxy-N-methyl-beta-D ribofuranuronamide (Cl-IBMECA) (100 nM) increased the magnitude of theta-burst induced LTP (from 1.2+/-0.6% in the control solution to 25.5+/-0.8% in the presence of Cl-IBMECA) and attenuated LTD (from 30.0+/-5.5% decrease in the control solution to 13.6+/-6.6% decrease in the presence of Cl-IBMECA). The selective adenosine A3 receptor antagonist, MRS 1191 (5-10 microM), prevented the effects of Cl-IBMECA. These findings indicate a functional role for adenosine A3 receptors in the modulation of synaptic plasticity. PMID- 11278111 TI - Neuroprotective effects of estrogen against beta-amyloid toxicity are mediated by estrogen receptors in cultured neuronal cells. AB - Although estrogen is known to exert beneficial effects on Alzheimer's disease, its underlying cellular mechanisms have not been clear. In this study we investigated whether or not neuroprotective effects of estrogen are mediated by estrogen receptors (ERs). Treatment of estrogen (1.8 nM) reduced beta-amyloid (Abeta)-induced death of ER-expressing W4 cells. This effect of estrogen was blocked by a specific ER blocker ICI 182,780. When estrogen was treated to HT22 cells, which lack functional ERs, Abeta-induced cell death was not affected. Transfection of HT22 cells with human ERalpha, but not ERbeta, restored protective action of estrogen against Abeta. Hoechst staining revealed that estrogen protected ERalpha-expressing cells by blocking Abeta-induced apoptosis. These results indicate that estrogen blocks Abeta-induced cell death via ERalpha dependent pathways. PMID- 11278118 TI - Humoral immune responses to phocine herpesvirus-1 in Pacific harbor seals (Phoca vitulina richardsii) during an outbreak of clinical disease. AB - Infection with phocine herpesvirus type-1 (PHV-1) has been associated with morbidity and high mortality in neonatal harbor seals (Phoca vitulina). A PHV-1 specific indirect enzyme linked immunosorbent assay (ELISA) was developed to sequentially measure the serological status of 106 harbor seal neonates admitted to a Pacific coast rehabilitation center (total number of sera tested was 371). Early in the season (February-April), the majority of pups had low serum levels of PHV-1 specific antibody. A dramatic increase in PHV-1 specific antibody, involving the majority of hospitalized pups, was observed during a 4-week period in May. This coincided with a high incidence of PHV-1 associated adrenal lesions and mortality. Although there was overall agreement between the timing of seroconversion to PHV-1 and histological evidence of PHV-1 infection, 82.4% of individual pups with adrenalitis had no evidence of a humoral response to PHV-1 at the time of their death. This suggests either a rapid disease course, or an inability to develop a humoral response in some neonatal seals. PMID- 11278119 TI - Development of disabled, replication-defective gene transfer vectors from the Jembrana disease virus, a new infectious agent of cattle. AB - Jembrana disease virus (JDV) is a newly isolated and characterised bovine lentivirus. It causes an acute disease in Bali cattle (Bos javanicus), which can be readily transmitted to susceptible cattle with 17% mortality. There is as yet no treatment or preventive vaccine. We have developed a gene transfer vector system based on JDV that has three components. The first of the components is a bicistronic transfer vector plasmid that was constructed to contain cis-sequences from the JDV genome, including 5'- and 3'-long terminal repeats (LTRs), 0.4kb of truncated gag and 1.1kb of 3'-env, a multiple cloning site to accommodate the gene(s) of interest for transfer, and an internal ribosome entry site plus the neomycin phosphotransferase (Neo) gene cassette for antibiotic selection. The second element is a packaging plasmid that contains trans-sequences, including gag, pol, vif, tat and rev, but without the env and packaging signals. The third is a plasmid encoding the G glycoprotein of vesicular stomatitis virus (VSV-G) to supply the vector an envelope for pseudotyping. Cotransfection of 293T cells with these three plasmid components produced VSV-G pseudotyped, disabled, replication defective, bicistronic JDV vectors encoding the green fluorescent protein (EGFP) and the Neo resistance selection maker simultaneously with a titre range of (0.4 1.2)x10(6)CFU/ml. Transduction of several replicating primary and transformed cells from cattle, primate and human sources and importantly growth-arrested cells with the JDV vectors showed high efficiency of EGFP gene transfer at 35 75%, which was stable and the expression of EGFP was long term. Furthermore, these JDV vectors were designed to suit the inclusion and expression of genes corresponding to JDV specific proteins, such as gag or env, for the development of vaccines for Jembrana disease. This strategy should also be applicable to other bovine diseases as well. The design and construction of the JDV vector system should facilitate the study of the lentivirology and pathogenesis of the diseases associated with JDV or other bovine virus infections. To our knowledge, this is the first such vector system developed from a cattle virus. PMID- 11278120 TI - Efficacy of spectinomycin against Mycoplasma bovis induced pneumonia in conventionally reared calves. AB - Sixteen 3-week-old calves were intratracheally inoculated with Mycoplasma bovis. Follow-up consisted of regular bronchoalveolar lavages (BALs) and clinical examinations. Animals were slaughtered from 4 to 21 days after inoculation. Counts were made of the mycoplasmas and other bacteria systematically isolated from the BAL liquids and lung lobes after slaughter. On the 6th day, spectinomycin 20mg/kg was given intramuscularly in three repeated doses at 24h intervals to six randomly chosen calves. All animals had developed a persistent M. bovis infection with a maximum BAL count on the 6th day (start of treatment). Co-occuring Pasteurella multocida infection was found in most animals with a maximum rate on the 14th day. The extent of lung surface lesions varied widely (0 64%) but was greater in the later slaughtered calves. Average counts of M. bovis and P. multocida in the BAL liquids were lower in treated calves than in untreated ones but the difference was not statistically significant. However, M. bovis and P. multocida counts in the lungs of the treated group were significantly lower than in the untreated group (p=0.003 and 0.009, respectively). PMID- 11278121 TI - Use of ESAT-6 in the interferon-gamma test for diagnosis of bovine tuberculosis following skin testing. AB - The whole blood interferon-gamma (IFN-gamma) test has proven to be a practical ancillary test for re-testing cattle for bovine tuberculosis 8-28 days following tuberculin skin testing. An improvement in the specificity of the IFN-gamma test could further reduce culling of false positive animals. The primary aim of this study was to evaluate a single mycobacterial antigen, ESAT-6 in the IFN-gamma test for use in skin test-positive cattle. These skin test-positive cattle comprised 51 Mycobacterium bovis-infected animals from tuberculosis-infected herds and 85 non-infected animals from tuberculosis-free herds. The test based on ESAT-6 had a higher specificity than the test based on purified protein derivative (PPD) tuberculin, but this was offset by a small decrease in sensitivity. Use of a lower cut-off in the ESAT-6-based test improved the sensitivity, while still maintaining a very high specificity. A secondary aim in the study was to assess the ESAT-6 and PPD-based tests for detecting bovine tuberculosis in skin test-negative animals from a persistently infected herd. The PPD-based test detected the majority of the lesioned or M. bovis-culture positive animals, while the ESAT-6-based test detected a smaller proportion. The false negatives in the IFN-gamma test from both the skin test-negative and positive groups were predominantly M. bovis-culture positive animals with no visible lesions. The current study has shown that a defined specific antigen such as ESAT 6 can markedly improve the specificity of the IFN-gamma test for re-testing skin test-positive animals. An ESAT-6-based IFN-gamma test could be particularly useful to reduce the false positive rate, yet still maintain an acceptable level of sensitivity. PMID- 11278122 TI - A novel method for isolation of Brachyspira (Serpulina) hyodysenteriae from pigs with swine dysentery in Italy. AB - Brachyspira (Serpulina) hyodysenteriae was isolated from 10 of 11 pigs with clinically suspected swine dysentery in six herds in northern Italy. All strains were successfully isolated in the selective blood agar modified medium with spectinomycin and rifampin (BAM-SR) currently used in our laboratory to isolate B. (S.) pilosicoli of human origin, after pre-treatment of intestinal material with spectinomycin and rifampin in foetal calf serum. Isolates had phenotypic characteristics typical of B. (S.) hyodysenteriae. PMID- 11278123 TI - Virulence of Pasteurella multocida recA mutants. AB - In order to determine the role of the RecA protein in the virulence of Pasteurella multocida, a recA mutant was constructed and used in studies of virulence and competition in relation to wild-type strain. To achieve this, firstly, the recA gene was isolated and sequenced, showing an Escherichia coli like SOS box and encoding a protein of 354 amino acids which has the closest identity with the Haemophilus influenzae RecA protein. Further, the recA mutant was constructed, by inactivating this gene by single recombination of a suicide plasmid containing an internal region of the P. multocida recA gene, and shown to be more sensitive to UV radiation than the parental strain. The P. multocida mutant was slightly attenuated in virulence, as indicated by the LD(50), the time of death of infected animals, and a failure to compete with the wild-type strain in mixed infections. Compared to the parent strain, the mutant had a similar growth rate but a longer lag phase. These data suggest that the diminished virulence of the recA mutant as well as its failure in competition were more a consequence of the long lag phase rather than a direct effect of the inactivation of the recA gene on genes involved in virulence. PMID- 11278124 TI - Expression and partial characterization of an elastase from Chromobacterium violaceum. AB - Chromobacterium violaceum was recovered at necropsy from the lungs, liver, spleen, and an interscapular abscess of a Chinese red panda (strain 98-9187) [J. Vet. Diagn. Invest. 12 (2000) 177]. As the lungs exhibited extensive, necrotizing lesions harboring bacterial aggregates, we sought to determine whether C. violaceum produced an elastase that might in part account for these lesions. The C. violaceum type strain (ATCC 12472(T)) and strain 98-9187 were shown to exhibit elastolytic activity by elastin Congo red and elastin nutrient agar assays. The activity was isolated from the periplasmic fraction and was present throughout the growth cycle. Activity increased markedly in late logarithmic phase growth. In elastin-limiting medium, activity rapidly decreased in early stationary phase indicating a tight regulation of yield. The activity was optimal at neutral pH and was sensitive to the metalloproteinase inhibitors EDTA and 1,10 phenanthroline. Activity was restored upon addition of zinc indicating the enzyme is a zinc metalloproteinase. A band corresponding to purified elastase activity was present at approximately 30kDa in a denaturing polyacrylamide gel. PMID- 11278125 TI - Differentiation of avian pathogenic Escherichia coli (APEC) strains by random amplified polymorphic DNA (RAPD) analysis. AB - Here we describe the application of a random amplified polymorphic DNA (RAPD) analysis for molecular genetic typing avian pathogenic Escherichia coli (APEC) strains. The RAPD technique was shown to be highly reproducible. Stable banding patterns with a high discriminatory capacity were obtained using two different primers. Overall, 55 E. coli strains were analyzed with a RAPD technique. The RAPD analysis showed that the E. coli strains isolated from poultry in Thailand and Sweden could be grouped into 50 of RAPD types by using these two different primer sets. Most of these different E. coli RAPD types were not geographically restricted. There was, as expected, a tendency of higher genetic relationship among E. coli strains isolated from the same farm. It is suggested that the RAPD technique may provide a rapid, low cost, simple and powerful tool to study the clonal epidemiology of avian E. coli infections. PMID- 11278126 TI - Molecular epidemiology of Salmonella Heidelberg in an equine hospital. AB - From 1992 to 1997, multi-drug resistant (MDR) Salmonella Heidelberg isolates were cultured from a number of horses hospitalised in a veterinary hospital in Victoria, Australia. To examine the relationships between the cases, 28 isolates from the hospital were compared by pulsed field gel electrophoresis (PFGE), IS200 element profiles, antimicrobial resistance patterns, plasmid profiles and phage typing. The PFGE patterns following digestion with XbaI and BlnI restriction endonucleases showed that the isolates from the veterinary hospital originated from a common source. These isolates also had indistinguishable IS200 profiles. However, PFGE was more discriminatory than IS200 profiles. All the veterinary hospital isolates and one independent isolate had the same antimicrobial resistance pattern and had at least one plasmid in common. Localisation of antimicrobial resistance genes indicated that the veterinary hospital isolates had more than one plasmid carrying resistance genes and that the genes encoding sulphathiazole and trimethoprim resistance were not on these plasmids. Phage typing was ineffective as 22 of the 28 isolates were untypeable. In conclusion, the combination of different methods used for epidemiological studies suggested that a single strain of MDR S. Heidelberg was isolated from horses admitted to the hospital for 6 years and caused salmonellosis in susceptible horses within that period with no apparent correlation between the antimicrobials used and retention of its MDR phenotype. PMID- 11278127 TI - Motor features in psychotic disorders. I. Factor structure and clinical correlates. AB - The dimensional structure of motor disorders remains largely unknown. This study aimed to ascertain the factor structure of motor signs and their clinical correlates in psychotic disorders. A sample of consecutive admissions of psychotic patients (n=187) was utilized to examine the factor structure of motor disorders as assessed by the Modified Rogers Scale (MRS). The relationship between motor dimensions and external variables was analyzed. A comparative examination of alternative factor solutions revealed that a six-factor structure, explaining 59% of the total variance, best fitted the 36 MRS items. This solution comprised the components of motor poverty, agitation, stereotypy/mannerisms, proskinetic, negativistic and dyskinetic. All the motor dimensions significantly improved over the psychotic episode. Motor dimensions differentially correlated with the syndromes of psychoses, with the association between motor poverty and the negative syndrome being particularly strong. Residual motor pathology, but not the acute one, was related to various clinical variables. Residual symptoms of motor poverty and stereotypy/mannerisms were associated with poor premorbid adjustment, more illness severity and a diagnosis of schizophrenia. It is concluded that the factor structure of motor disorders and its clinical correlates are rather more complex than generally acknowledged. PMID- 11278128 TI - Motor features in psychotic disorders. II. Development of diagnostic criteria for catatonia. AB - Existing diagnostic criteria for catatonia have been based exclusively on theoretical assumptions. The present study aimed to develop empirical criteria for diagnosing catatonia. The same patient population (n=187) described in Part I (Peralta, V., Cuesta, M.J., Motor features in psychotic disorders. I. Factor structure and clinical correlates. Schizophr. Res., 00, 000-000) was used in this study. Fourteen catatonic signs with potential diagnostic value were cluster analyzed to derive groups with and without a catatonic syndrome. Taking the cluster solution as the reference standard, the diagnostic value for individual signs was examined by means of multiple discriminant analysis, ROC analysis, and the parameters of sensibility, specificity, positive predictive power and negative predictive power. Cluster analysis divided the sample into a catatonic group (n=32) and a noncatatonic group (n=155). Discriminant analysis showed that 11 of the 14 potential diagnostic signs discriminated among groups: immobility/stupor, mutism, negativism, oppositionism, posturing, catalepsy, automatic obedience, echophenomena, rigidity, verbigeration and withdrawal. ROC analysis showed that any combination of three or more of these symptoms best fitted the cluster-derived catatonic syndrome. In conclusion, patients displaying three or more classical motor signs may be diagnosed with confidence as suffering from a catatonic syndrome. PMID- 11278129 TI - Risk factors for suicide in young people suffering from schizophrenia: a long term follow-up study. AB - Ten per cent of patients with schizophrenia commit suicide, but assessment of risk is difficult. Large case-control studies with a long follow-up period are needed. These should focus on patients from one age group to give clinicians the details required to identify those at highest risk.We present a case-control study of 63 patients who committed suicide and 63 controls from a consecutive admission series of patients with a diagnosis of schizophrenia. All patients were under the age of 30 at admission.Risk factors for suicide were male gender, chronic illness with frequent relapses (OR 6.0), frequent short hospitalisation, a negative attitude towards treatment (OR non-compliance 7.0), impulsive behaviour (OR acting out 6.4, OR involuntary commitment 17), parasuicide (OR suicide attempt 4.8, OR highly lethal suicide attempt 11), high pre-morbid IQ (OR 4.3), psychosis (OR 7.0) and depression (OR 36). However, early onset of a defect state (OR 6.3) and a daily activity (OR 4.2) were protective factors. Identified risk factors could help clinicians to target high-risk patients and form the basis for interventions aimed at reducing suicide. PMID- 11278130 TI - Relationships between depression and psychotic symptoms of schizophrenia during an acute episode and stable period. AB - The aim of the present study is to explore the relationship between depression and psychotic symptoms of schizophrenia over the course of illness. Sixty-eight patients meeting DSM-IV criteria for schizophrenia were enrolled, 27 in an acute episode, 41 when stable. Assessments were performed using the Calgary Depression Scale for Schizophrenia (CDSS) for depression and the Positive and Negative Syndrome Scale (PANSS) for psychotic symptoms. When considering patients in an acute episode (52% depressed), the CDSS score was correlated only with the PANSS positive sub-scale score. For patients in the stable period (38% depressed), the CDSS score was correlated with positive as well as negative and general psychopathology sub-scale scores. Hence, the relationship between depression and other symptoms of schizophrenia appear to differ during different stages of illness. PMID- 11278131 TI - Male gender is associated with deficit schizophrenia: a meta-analysis. AB - An association between deficit schizophrenia and male gender could be expected, since male schizophrenic subjects have been repeatedly found more severe than females on several dimensions of severity. Surprisingly, very few studies have confirmed such an association. We performed a more definitive test of this association using a meta-analysis. A pooled odds ratio was computed based on the 23 studies that reported the gender ratio in deficit vs. non-deficit schizophrenia. We tested for the heterogeneity of the association and examined the potential impact of the sampling method, the method used to assess the deficit syndrome, the breadth of diagnoses included and the mean duration of illness. A highly significant association between male gender and deficit schizophrenia was observed (pooled odds ratio=1.75). There was no definitive evidence that differences across studies in sampling methods, breadth of diagnoses included, mean duration of illness and methods to assess the deficit syndrome affected the strength of the association. However, the studies using the "Proxy Deficit Syndrome" method to assess the deficit syndrome yielded qualitatively weaker evidence. This significant association between male gender and deficit schizophrenia may reflect the influence of a gender related factor (e.g. sexual hormones) or gender differences in the liability to different etiologies of schizophrenia. The role of gender as a potential confounder must be closely examined in studies comparing deficit and non-deficit SZ. PMID- 11278132 TI - The factor structure for positive and negative symptoms in South African Xhosa patients with schizophrenia. AB - Most studies investigating the symptom dimensions of schizophrenia utilising the Scale for the Assessment of Negative Symptoms (SANS) and the Scale for the Assessment of Positive Symptoms (SAPS) favour a three factor model. This study sought to investigate the factor structure of both the global and individual items of the SANS and SAPS in a large sample of South African Xhosa patients with schizophrenia. A total of 422 subjects participated. Both principal components and factor analytical procedures were applied. For the global items, a two-factor solution representing positive and negative symptoms accounted for 59.9% of the variance. Alternatively, the three-dimensional model of negative, psychotic and disorganisation factors was supported by a five-factor solution if the more heterogeneous items of attention and alogia were ignored. Analysis of the individual items yielded a five-factor solution with the negative symptoms splitting into diminished expression and disordered relating, and the positive symptoms separating into factors for psychosis, thought disorder and bizarre behaviour. Our findings are very similar to those from other parts of the world, providing evidence that the factor structure for the symptoms of schizophrenia is relatively resistant to cultural influences. This is particularly true for negative symptoms. PMID- 11278133 TI - Suspiciousness as a specific risk factor for major depressive episodes in schizophrenia. AB - OBJECTIVE: Serious depression is a common and important complication of schizophrenia. In a prospective, population-based study, we tested the hypothesis that suspiciousness increases the risk for the later development of depression in schizophrenia. METHOD: Data came from the Epidemiological Catchment Area (ECA) study. Baseline clinical and demographic features were used to predict the onset of new episodes of depression at 1 year follow-up. As ECA diagnoses were based on lay interviews, which may have low sensitivity compared with clinical diagnoses, two overlapping groups of putative schizophrenia patients were defined. RESULTS: Suspiciousness was associated with an increased risk of new episodes of depression in both patient groups, after accounting for demographic variables. There was no association between an increased risk of depression and either disorganization or hallucinations and delusions. CONCLUSIONS: Suspiciousness appears to be a specific risk factor for depression in psychotic groups. Interventions that decrease suspiciousness, or mitigate its isolating effects, might decrease the risk of serious depression and suicide. PMID- 11278134 TI - Obstetric complications in women with schizophrenia. AB - It is not known whether schizophrenic women have increased incidence of complications during pregnancy and delivery. Data from the Danish Medical Birth Register were used to compare 2212 births to 1537 schizophrenic women in Denmark with a random sample of all deliveries in Denmark during 1973-1993 (122931 births to 72742 women). The schizophrenic women had fewer antenatal care visits. They were at lower risk of pre-eclampsia, but tended to have lower Apgar scores. There were no other differences in the incidence of specific complications such as placenta previa, placental abruption, and abnormal fetal presentation. Schizophrenic women were at increased risk of interventions such as Cesarean section, vaginal assisted delivery, amniotomy, and pharmacological stimulation of labor. There were no important differences between the deliveries to schizophrenic women who gave birth before and after their first admission to a psychiatric department. These results show no evidence that schizophrenic women have a greater frequency of specific obstetric complications than non schizophrenic women. Nevertheless, they are at increased risk for interventions during delivery. PMID- 11278136 TI - Depression in schizophrenia: recognition and management in the USA. AB - The recognition of depression as a distinct syndrome within schizophrenia is a relatively recent development. The International Survey of Depression in Schizophrenia was designed to evaluate current clinical practice and prescribing trends in the management of the depressive component of schizophrenia. A 48-item questionnaire, comprising fixed-response questions and questions stimulated by case scenarios, was distributed to 37513 psychiatrists in the USA. A total of 43484 psychiatrists in Canada, Australia and 21 European countries also received the questionnaire. A total of 1128 US psychiatrists responded. Analysis of the data revealed that US psychiatrists identify symptoms of depression in approximately one-third of patients with schizophrenia, and largely appreciate the magnitude of the resultant burden on patients and their families. Responses to questions regarding treatment approaches and case scenarios demonstrated that the level of adjunctive prescribing of antidepressants in the USA is often higher than in other regions. Levels of awareness of depression in patients with schizophrenia and recognition of the need for effective management appear to be high among US psychiatrists. However, more than a quarter of these specialists rarely or never prescribe adjunctive antidepressant medications. Disparities in treatment approaches varying from the existing scientific evidence base underscore the need for further investigation into ways of optimizing the management of this serious coexisting condition. PMID- 11278135 TI - Low birthweight in schizophrenia: prematurity or poor fetal growth? AB - In the general population, low birthweight (LBW) is associated with neurological and psychological problems during childhood and adolescence. LBW may result from premature birth or poor fetal growth, and the independent effects of these two events on childhood development are not fully understood. The rate of low weight births is increased in schizophrenia and is associated with social withdrawal during childhood and an early onset of illness. However, it is unclear whether this LBW reflects poor fetal growth or premature birth, or whether these two risk factors have distinct implications for childhood functioning and age at onset of schizophrenia. Subjects included 270 patients with schizophrenia for whom a detailed history of obstetric events could be obtained. The rate of low weight births was high and was associated with poorer premorbid functioning and an earlier age at illness onset. The rate of both premature births and poor fetal growth was high relative to the normal population. Prematurity, but not poor fetal growth, was associated with premorbid social withdrawal and an early age at illness onset. Poor fetal growth, but not prematurity, was associated with low educational achievement. These results suggest that poor fetal growth and prematurity are associated with distinct patterns of childhood maladjustment in individuals who develop schizophrenia. PMID- 11278137 TI - The Schizophrenia Suicide Risk Scale (SSRS): development and initial validation. AB - BACKGROUND: Estimations about the lifetime risk of suicide in schizophrenia vary between 4 and 10%. At present, there does not exist a suicide risk scale developed particularly for schizophrenic patients. The aims of the present study were to: (1) develop a clinically useful semi-structured scale for the estimation of short-term suicide risk among schizophrenic patients, and (2) to carry out an initial validation of the scale. METHODS: A 25-item Schizophrenia Suicide Risk Scale (SSRS) was constructed on the base of the literature. The SSRS scores of 69 living schizophrenic patients (LS group) were compared with the scores of 69 schizophrenic suicides (SS group) whose data had been collected previously from The Finnish nationwide and representative psychological autopsy study. Internal consistency of the SSRS was evaluated with Cronbach alpha. The most important SSRS items predicting suicide were identified with a logistic regression analysis. Sensitivity, specificity, positive predictive value, and negative predictive value of the SSRS in predicting suicide with various cut-off scores were calculated. RESULTS: In the final logistic regression model, the following SSRS items significantly predicted suicide: suicide plans communicated to someone during the past 3 months; one or more previous suicide attempts; loss of professional skills demanding job; depression observed during an interview; and suicide plans communicated during an interview. With high cut-off scores the specificity of the SSRS became satisfactory, but the sensitivity dropped below 32%. Internal consistency of the anamnestic history of the SSRS was low, which suggests that anamnestic risk factors for suicide in schizophrenia are multifactorial. Internal consistency of the interview-based items was high, and present state risk factors seemed to consist of two separate factors, depression anxiety and irritability. CONCLUSIONS: The SSRS may be clinically useful in identifying schizophrenic patients with a particularly high risk for suicide. However, the SSRS seems not to be a practical screening instrument for suicide risk in schizophrenia, and it is probably impossible to construct a suicide risk scale with both high sensitivity and high specificity in this disorder. PMID- 11278138 TI - Duration of untreated psychosis predicts treatment outcome in an early psychosis program. AB - For patients first presenting with a non-affective psychotic disorder, the duration of untreated psychosis (DUP; the time between the onset of positive psychotic symptoms and the initiation of appropriate treatment) varies widely, from a few weeks to several years. A number of studies report that a longer DUP is associated with poorer clinical outcomes. We studied DUP and its association with clinical outcomes in a group of patients with schizophrenia and related psychotic disorders treated in the naturalistic clinical setting of an early psychosis program. DUP was determined for 19 patients with a non-affective psychotic disorder (schizophrenia, schizoaffective disorder or schizophreniform disorder) and no previous treatment for psychosis, by use of the IRAOS, a retrospective structured interview carried out with patients and their families. Positive and Negative Syndrome Scale (PANSS) and Global Assessment of Function (GAF) ratings were available at baseline and 6month follow-up. For analysis, patients were categorized into a short DUP (n=9) or long DUP (n=10) group. The median DUP (57weeks) was used as the dividing point. At baseline, the two groups did not differ significantly on positive symptoms or total PANSS ratings. However, negative symptoms were more severe in the long DUP group at baseline (P=0.029), and the long DUP group had a significantly higher mean rating for the passive/apathetic social withdrawal item of the PANSS (P=0.024). At 6month follow up, the long DUP group had significantly higher ratings for positive symptoms (P=0.028) and had lower GAF scores (P=0.044). Significantly more (P=0.033) long DUP patients had enduring positive psychotic symptoms. The results confirm both the wide range of DUP among patients first presenting with schizophrenia and related psychotic disorders and the association of long DUP, defined as greater than approximately 1year, with a poorer clinical outcome. This study highlights the importance of collecting data regarding DUP and supports the view that patients with a long DUP are likely to be less responsive to treatment in general and will require greater resources and more intensive interventions. PMID- 11278139 TI - Familiality of symptom dimensions in schizophrenia. AB - The division of schizophrenic symptoms into three core dimensions - psychomotor poverty, reality distortion, and disorganisation - is well established. When factor analytic studies have included affective symptoms they have identified two additional dimensions - manic and depressive. Whether these five dimensions represent underlying psychopathology of a genetic or environmental aetiology remains unclear. The aims of this study were to perform factor analysis of symptoms in a group of familial schizophrenic patients and to investigate the familiality of the symptom dimensions identified, and their relationship to clinical characteristics. Symptoms were recorded, using the Operational Criteria Checklist for Psychotic Illness, for 155 Caucasian subjects with an RDC diagnosis of schizophrenia, schizoaffective disorder, or psychosis of unknown origin, from 61 families multiply affected with schizophrenia. Factor analysis indicated five symptom dimensions: depressive, manic, reality distortion, disorganisation, and psychomotor poverty. The psychomotor poverty, disorganisation, and manic dimensions were shown to be familial. Psychomotor poverty, disorganisation, and reality distortion were all associated with deterioration from premorbid functioning and chronic course of the disorder. In addition, psychomotor poverty was significantly related to poor premorbid functioning, as well as to single marital status and unemployment at onset. Disorganisation was significantly related to single marital status and unemployment at onset. The familiality of the psychomotor poverty, disorganisation, and manic dimensions supports their use in the delineation of homogeneous subsets for genetic studies. PMID- 11278140 TI - Genetic epidemiology and schizophrenia: a study of reproductive fitness. AB - Genetic epidemiological studies have demonstrated markedly reduced rates in reproduction among schizophrenic patients. According to evolutionary theory, behavioral and psychological phenotypes are selected based on ecological "fit". Where differential survival or reproductive success exists, genotype frequencies are altered in subsequent generations. In the case of schizophrenia, lower rates of reproduction constitute a negative selection factor that should reduce genes in the population associated with the expression of the disease--ultimately leading to decreases in prevalence. However, studies reveal a stable prevalence of about 1% over time. Attempts to explain the apparent contradiction between negative selection and stable prevalence have taken several forms. One explanation suggests that reproductive rates in relatives of schizophrenic patients are increased--compensating for reproductive loss in affected family members. Family data from schizophrenic patients at the Maryland Psychiatric Research Center were compared with those of healthy volunteers and volunteers with schizophrenia spectrum personality (SSP) disorders. Controlling for important socio-cultural and demographic variables, a multiple regression model revealed a significant increase in the number of siblings associated with schizophrenia. No differences in reproductive fitness were found among normal and SSP volunteers. This observed pattern in reproductive fitness provides one mechanism by which prevalence rates can remain stable despite lower reproductive rates among individuals with schizophrenia. Evidence of increased reproductive fitness in relatives suggests the need to consider the complex interactions of proximate and ultimate (evolutionary) mechanisms in the expression of schizophrenia. PMID- 11278141 TI - Household crowding in early adulthood and schizophrenia are unrelated in Denmark: a nested case-control study. AB - A number of studies have found that being born in an urban area is a risk factor for developing schizophrenia. It has been hypothesized that increased exposure to infectious agents through household crowding might account for this association. Using Danish longitudinal registers, we have established a population-based sample of 191 cases of schizophrenia where the first admission occurred between 1981 and 1993. These cases were compared with 17413 individually matched controls of the same gender and age. Information regarding parents' and siblings' psychiatric history, urbanization, season and place of birth, and square meter per dweller were included in a conditional logistic regression model. We found square meter per dweller to be insignificant and without any trend when included as a risk factor for schizophrenia, whereas previous findings of schizophrenia associated with being born in an urban area and with schizophrenia in parents and siblings were replicated. PMID- 11278142 TI - Time trends in first admissions for schizophrenia and paranoid psychosis in Stockholm County, Sweden. AB - Several studies have reported decreasing time trends in first diagnosed schizophrenia patients. The aim of this study was to analyze time trends for first admissions with a diagnosis of schizophrenia or a diagnosis of either schizophrenia or paranoid psychosis during 1978-1994 in Stockholm County, Sweden, with a population of around 1.8million. Information about first psychiatric admission with the diagnosis schizophrenia or paranoid psychosis for residents of Stockholm County was obtained from the Swedish population-based psychiatric inpatient register. Age-adjusted average yearly changes in first hospitalization rates were estimated in a Poisson regression model. Time trends in first admission rates were calculated from 1978 to 1994, while admissions during 1971 to 1977 were observed only to eliminate later re-admissions. First admissions for schizophrenia declined by 1.9% annually for females and by 1.3% for males, while first admissions for schizophrenia and paranoid psychosis together were unchanged over the study period for both genders. Our results indicate that the incidence of schizophrenia and paranoid psychosis taken together was essentially the same over the studied time period in Stockholm County, and that the apparent decline in first admission rates for schizophrenia may be an effect of changes in clinical diagnosis over time. PMID- 11278143 TI - Unilateral olfactory perception and magical ideation. AB - We assessed olfactory detection thresholds and discrimination abilities in 40 healthy right-handers (20 women and 20 men). All subjects were also required to complete the Magical Ideation (MI) scale, a well-validated 30-item schizotypy inventory. Over both nostrils, we found elevated thresholds for subjects with high MI scores (at or above the median score of 9.0) compared with those with low scores. In men but not women, specifically left-nostril acuity was inversely correlated to MI raw scores. MI was unrelated to olfactory discrimination performance. These results suggest an association, at least in healthy men, between even moderate signs of schizotypy and deficits in odor detection. The selective impairment of left-nostril performance adds to the growing evidence for left temporal lobe functional abnormalities in people high on MI. This laterality effect is known from previous studies in patients with schizophrenia. However, as a rule, in psychiatric patients olfactory identification rather than simple detection performance was found to be impaired, indicating that the integration of odor information is affected at different levels of processing in schizotypy compared with schizophrenia. Work with completely normal subjects may reasonably complement clinical studies of olfactory perception. Among its advantages are the good subject compliance and the absence of medication effects. PMID- 11278144 TI - Pain insensitivity in the relatives of schizophrenia patients. AB - The psychiatric literature contains anecdotal reports of diminished pain sensitivity in schizophrenia that date back to Kraepelin. Yet, the phenomenon of pain insensitivity in schizophrenia remains largely unstudied. For example, it is not clear if pain insensitivity is a consequence of the illness or if it is also present in the well relatives of schizophrenia patients. To explore this issue, we examined pain thresholds and pain tolerances in healthy young adults. Compared with controls with no family history of psychopathology (n=21), participants with a family history of schizophrenia (n=32) showed elevated pain thresholds and pain tolerances to finger pressure. Pain insensitivity was also significantly correlated with elevated scores on measures of self-referential thinking, magical ideation, and perceptual disturbances. Finally, a sizeable minority (19%) of well relatives of schizophrenia patients showed extreme pain insensitivity compared to other participants. The pattern of findings suggests that pain insensitivity may warrant further exploration as a potential marker of underlying liability to psychosis. PMID- 11278145 TI - Number of older siblings of individuals diagnosed with schizophrenia. AB - One of the most consistent epidemiological findings in schizophrenia research is the small excess of late winter/early spring births. There is also evidence that schizophrenia is associated with urban birth and with later birth order. One interpretation of these three findings is that respiratory viral infections brought into the household by children in crowded areas could disrupt foetal brain development and predispose to schizophrenia in later life. To further explore this hypothesis, case register data were used to assess if schizophrenics with a greater number of older siblings are more likely to be born in urban areas and during late winter/early spring months. Data from the Dublin and Three County Case Register were compiled relating to 2969 patients with schizophrenia and 5904 patients with neurosis. We used logistic regression analysis to determine if the number of older siblings differentiated schizophrenia from neurosis after controlling for the effects of gender, urban/rural birth, season of birth and sibship size, and to examine whether any interactions existed. The number of older siblings did not predict a diagnosis of schizophrenia over neurosis. There was no interaction between number of older siblings and urban birth, between number of older siblings and spring birth, or between number of older siblings, season of birth and urban birth. These data do not support the hypothesis that schizophrenia, by comparison with neurosis, is associated with an increased number of older siblings or that there is an interaction between number of older siblings, urban birth or season of birth. PMID- 11278146 TI - Antipsychotic prescription use and costs for persons with schizophrenia in the 1990s: current trends and five year time series forecasts. AB - Real advances in schizophrenia pharmacotherapy have been made over this decade with the development of more efficacious treatment options with fewer side effects. These advances have high per-unit direct costs that may have a profound effect on drug budgets of systems caring for persons with schizophrenia. The objective of this study was to describe the changes in utilization and cost for antipsychotic prescriptions by atypical, clozapine, decanoate products, and traditional neuroleptics in a large naturalistic setting, i.e. the Georgia Medicaid population. Secondly, this study forecasted the categorized antipsychotic prescription utilization through the year 2002. Administrative claims data spanning 1990-1997 for Medicaid eligible persons suffering from schizophrenia in the state of Georgia were supplemented with psychiatric institutional data obtained from the Georgia Department of Human Resources. A total of 16227 Medicaid-eligible recipients had a code indicative of schizophrenia (ICD-9-CM=295.(**)) and were at least 16 years of age at the time of their first diagnosis. The mean recipient prescription use and expenditures were tallied for each month of the study and stratified by prescription category (atypical, clozapine, decanoate, and traditional antipsychotic). ARIMA time series models were identified and estimated using these monthly PMPM utilization and expenditures estimates to forecast 5 years beyond the last month of the study. The total use of antipsychotics increased modestly throughout the study period, and the use of atypicals, clozapine, and decanoate products increased substantially, while a decrease was observed for traditional antipsychotics. In 1995 dollars, antipsychotic expenditures increased from a mean of approximately $10 PMPM in 1990 to $95 projected for the year 2002. This transition from traditional oral antipsychotics to atypicals and decanoate products has a profound effect on drug expenditures for systems paying for the care of persons with schizophrenia. Further studies to determine the value of the transitions of therapy described in this study need to be evaluated using a system-wide- or Medicaid perspective. PMID- 11278147 TI - Cytokine profiles in drug-naive schizophrenic patients. AB - A large body of evidence concerning immunological abnormalities in schizophrenic patients seems to suggest a role of the immune system in the multifactorial pathogenesis of schizophrenia. We investigated the production of various cytokines [interleukin (IL)-2, IL-4, IL-10, interferon (INF)-gamma] in drug-free (n=26) and drug-naive (n=7) schizophrenic patients and in healthy controls (n=33). Production of IL-2 and INF-gamma was significantly higher (respectively P=0.021 and P=0.001) in patients than in controls. These findings provide further evidence that immunological abnormalities are present in some schizophrenic patients. PMID- 11278148 TI - Selective impairments of theory of mind in people with schizophrenia. AB - "Theory of mind" (ToM) means the ability to represent others' intentions, knowledge and beliefs and interpret them. Children with autism typically fail tasks aimed at assessing their understanding of false beliefs. These features of autism are strikingly similar to some negative features of schizophrenia. Mental abilities were studied in 35 schizophrenics (DSM-IV) and 17 normal controls. Subjects heard four ToM stories and simultaneously were shown cartoons depicting the action occurring in the stories. All stories involved false beliefs or deception. As for the current symptomatology, schizophrenics were divided according to Liddle's three-dimensional model (reality distortion, psychomotor poverty, disorganisation). Our results show significant differences between schizophrenics and normal controls in all ToM stories, with schizophrenic people performing worse than controls. In first-order stories (a false belief about the state of the world) significant differences were found among symptom dimensions, with the psychomotor poverty group performing worse than disorganisation subjects and reality distortion ones. As for second-order stories (a false belief about the belief of another character), the psychomotor poverty group performed worse than the other groups only in one of the four ToM stories. More research in separating ToM deficits from attention disturbances is needed. PMID- 11278149 TI - Physostigmine and cognition in schizotypal personality disorder. AB - There is evidence that reduced cholinergic activity may play a role in the pathophysiology of cognitive impairment in the schizophrenia spectrum. We tested the effects of physostigmine, an anticholinesterase inhibitor, on visuospatial working memory as evaluated by the Dot test, and on verbal learning and recall as measured by a serial learning task in patients with schizotypal personality disorder. Physostigmine tended to improve the Dot test, but not serial verbal learning performance in these patients. PMID- 11278150 TI - Differential neuropsychological patterns of frontal- and temporal-lobe dysfunction in patients with schizophrenia. AB - The frontal and temporal lobes have been implicated as pathogenic sites for schizophrenia, although there is a marked heterogeneity of brain function and structure between individual patients. It is currently unclear whether some patients with schizophrenia exhibit primarily frontal lobe dysfunction, while others exhibit primarily temporal-lobe dysfunction. The current investigation examined this issue in a preliminary way by using neurocognitive tests to discriminate test performances of patients with schizophrenia from patients without schizophrenia who had definitive neurological evidence of either frontal- or temporal-lobe dysfunction. Of the patients with schizophrenia, 20.7% were classified as having a frontal lobe dysfunction profile, while 19.3% had a temporal lobe dysfunction profile. Results further clarify neurobiological heterogeneity in schizophrenia by demonstrating that a substantial number of patients with schizophrenia exhibit either primarily frontal- or temporal-lobe dysfunction. Results may partially explain the inadequacy of neurobiological models for schizophrenia that do not consider these differential patterns of dysfunction. PMID- 11278151 TI - Effects of olanzapine and other antipsychotics on cognitive function in chronic schizophrenia: a longitudinal study. AB - This study aimed to determine the effect of olanzapine and other antipsychotic drugs on cognitive functions after 6months of treatment. Baseline, 3month and 6month psychopathological and cognitive evaluations were made. Thirty-eight partially responsive outpatients with DSM-IV chronic schizophrenia diagnosis were included in the study. On the indication of their attending psychiatrists, 21 patients initiated treatment with olanzapine, and 17 remained on their previous treatment with other antipsychotic drugs. Cognitive assessments were blind to medication and psychopathological status. The olanzapine group presented a significantly greater improvement in negative symptomatology and verbal memory than the comparison group in repeated-measures of MANOVAs between baseline, 3month and 6month assessments. These differences remained statistically significant after covarying out gender, treatment with other atypical antipsychotics, biperidene doses and changes in positive and negative symptoms. In order to match previous differences between groups, cognitive baseline scores for each test were introduced as covariates, resulting in a significant improvement for the olanzapine group in negative symptomatology and the interference task of the Stroop test.We then re-analyzed the data, dividing the comparison group into two groups: risperidone-treated patients (n=9) and patients receiving conventional antipsychotic drugs (n=8). Post-hoc analyses between groups were carried out with baseline cognitive assessment as covariate. The olanzapine group improved significantly more than the risperidone group in negative symptomatology and in the interference task of Stroop test. The improvement in the number of categories of the Wisconsin Card Sorting Test was higher in risperidone patients than in those receiving olanzapine or conventional antipsychotic treatment. Conventional antipsychotic drugs did not present a significant improvement over atypical antipsychotic drugs in any cognitive function. In summary, in patients suffering from chronic schizophrenia, atypical antipsychotic agents were associated with slight differential improvements over time in attentional, verbal memory and executive functions compared with conventional neuroleptic drugs. No differential improvements were found in social functioning, verbal fluency, non-verbal domains of memory or visuo-motor abilities. PMID- 11278152 TI - Stroop interference and facilitation effects in first-episode schizophrenic patients. AB - In the Stroop test, interference occurs in naming the print color of a word when the word is itself the name of another color. Facilitation occurs when the word is the same as the print color. Previous studies on selective attention in schizophrenia using the Stroop interference effects have yielded contradicting results. Constraints included limited sample size and the recruitment of medicated chronic patients. We studied the Stroop interference and facilitation effects in a relatively large sample of first-episode schizophrenic patients (n=56), a substantial proportion of whom were medication-naive (n=30) at the time of initial testing. We have also carried out longitudinal follow-up assessments when patients reached a clinically stable state, as well as 4months after recovery from the episode. We found that the Stroop interference effect was not increased in first-episode schizophrenic patients, whether medication-naive or not. This effect did not change over the follow-up period. In addition, we detected an increase in Stroop facilitation effect in medicated schizophrenic patients, but only in the initial assessment soon after they had received medication. After sustained treatment, the increase in facilitation was normalized. These observations supported previous findings of a normal Stroop interference effect amongst schizophrenic patients. The increased facilitation effect for patients in their early phase of treatment (but not later) may represent an acute effect of anti-psychotic medication. Its nature and significance require further investigation. PMID- 11278153 TI - Memory and attention deficits in drug naive patients with schizophrenia. AB - The present study was designed to evaluate the integrity of cognitive fronto temporal processes in drug naive patients with schizophrenia. The evaluation of drug naive patients discards the potential influence of medication, and may allow the specification of cognitive impairments that are truly illness-related. Subcomponents of long-term memory as well as several measures of attention were examined. A group of 16 patients who had never taken antipsychotics and a group of 20 normal controls underwent tests of alertness, information maintaining, and sustained and selective attention, as well as tests of explicit and implicit recall. The psychopathological manifestations of patients were also assessed with the BPRS, PANSS, ESRS clinical scales. Attention test performances revealed that drug naive patients presented a decrease in their ability to respond promptly to a stimulus, sustain their attention on a task, display normal selective attention strategies, and maintain information for on-line processing. The results also suggest that the drug naive patients are impaired when both strategic and associative processes must be triggered to explicitly recover information in long term memory. In contrast, the results revealed that implicit access to perceptual mental representations is spared in schizophrenic patients. Finally, features of the patients' clinical symptomatology and some cognitive deficits were also shown to be correlated. Overall, results showed that, in relation to normals, drug naive patients were mildly impaired, with little intersubject variability, and that not all cognitive processes were equally disturbed in relation to the normal subjects' performances. Results support the idea that an important part of the impairments seen in schizophrenia is present before the introduction of neuroleptic medication and chronic illness. PMID- 11278154 TI - Neuropsychological change in schizophrenia after 6 weeks of clozapine. AB - Neuropsychological change after 6weeks of clozapine treatment was examined in 18 treatment-refractory patients to test anticipated domain-specific cognitive improvements. The first aim of this study was to test the assumption that increased homogeneity of sample and treatment would yield an experimental design with sufficient sensitivity to detect general intellectual changes with clozapine that were not apparent in one previous investigation. The second aim was to test predictions derived from a domain-specific review of all other investigations with clozapine suggesting salient gains on tests sensitive to motor and mental speed, visual spatial manipulation, and new learning of verbal material. The results showed that the comprehensive neuropsychological test battery was sensitive to general cognitive changes with clozapine, and supported the hypothesized domain-specific gains on tests of motor and mental speed, visual spatial manipulation and new verbal learning. Novel gains were also apparent on tests of new learning with nonverbal material. The results are discussed in relation to aspects of experimental design necessary for the evaluation of prospective medication-induced changes in cognitive skill, particularly in future investigations designed to differentiate between second-generation antipsychotic medications. PMID- 11278155 TI - Substance abuse in schizophrenia: a review of the literature and a study of correlates in Sweden. AB - The purpose of the current study is twofold: (a) to provide an overall synthesis of recent research on substance abuse in schizophrenia and (b) to present findings in a Swedish patient sample. Studies conducted since 1990 have found a wide range of abuse prevalence rates, with male gender and younger age as primary correlates. Less certainty exists regarding substance abuse as an independent risk factor for schizophrenia and its further impact on illness course. In a sample of 87 patients attending a psychiatric clinic in Malmo, lifetime prevalence of substance abuse was 48.3%, with abuse debut primarily preceding first contact for psychotic symptoms. Significant correlates of abuse were male gender, family history of substance abuse, and increased rates of hospitalization and criminality, with poorer outcome found in previous as well as current abusers. Alcohol abuse, either solely or in combination with other substances, was the main type of substance abuse. Although the specific factors (social, behavioural, genetic) that predispose schizophrenic patients to substance abuse remain unclear, the predominantly male profile of abusers might suggest a link between substance abuse and the poorer clinical outcome frequently observed, especially in male schizophrenic patients. PMID- 11278156 TI - Cannabis use is associated with schizotypy and attentional disinhibition. AB - While most neurochemical research into the pathogenesis of schizophrenia (SZ) has focused on the dopaminergic, glutamatergic, and serotonergic systems, the exact nature and cause of this disorder have proven intractable. Given the recent discovery and elucidation of the endogenous cannabinioid system, a re-examination of the cannabis-induced exacerbation hypothesis of SZ is warranted. The purpose of the present study was to assess whether current cannabis users exhibit personality correlates and neurocognitive deficits similar to those observed in SZ patients. 15 current cannabis users, 15 drug-free controls, and 10 past cannabis users were assessed on tasks which assess attentional inhibition, spatial working memory, olfactory identification, and schizotypal personality. Current cannabis users demonstrated deficits in attentional inhibition, decreased reaction time, and significantly higher scores on the schizotypal personality questionnaire (SPQ) compared with the non-using and past cannabis using groups. No group differences were found on the working memory or olfactory identification tasks. These results suggest that cannabis use can mimic attentional deficits seen in acute schizophrenia and is associated with schizotypal personality, thus setting the stage for a possible cannabinoid model of SZ. PMID- 11278157 TI - Rural and urban differences in patients with a dual diagnosis. AB - OBJECTIVES: To evaluate the differences between two cohorts of patients with severe mental illness (schizophrenia-spectrum or bipolar disorder) and co occurring substance-use disorders, living in either predominantly rural areas or urban areas. METHODS: Two study groups of patients with a dual diagnosis, recruited using the same criteria, were evaluated, including 225 patients from New Hampshire and 166 patients from two cities in Connecticut. The two study groups were compared on demographic characteristics, housing, legal problems, psychiatric and substance use diagnoses, substance use and abuse, psychiatric symptoms, and quality of life. RESULTS: Patients in the Connecticut study group had higher rates of cocaine-use disorder, more involvement in the criminal justice system, more homelessness, and were more likely to be from minority backgrounds. The Connecticut group also had a higher proportion of patients with schizophrenia and more severe symptoms, as well as lower rates of marriage, educational attainment, and work than the New Hampshire study group. Alcohol-use disorder was higher in the New Hampshire group. Subsequent analyses within the Connecticut group indicated that although African American patients had higher rates of cocaine-use disorder than white patients, cocaine disorder and not minority status was most strongly related to criminal involvement and homelessness. CONCLUSIONS: Because of the substances abused and the greater degree of psychiatric illness severity, patients with a dual diagnosis who are living in urban areas may require greater ancillary services, such as residential programs, Assertive Community Treatment, and jail diversion programs in order to treat their disorders successfully. PMID- 11278158 TI - Social functioning, psychopathology, and medication side effects in relation to substance use and abuse in schizophrenia. AB - OBJECTIVE: To identify correlates of self-reported substance use and problems associated with that use in people with schizophrenia. METHODS: A sample of 404 patients with schizophrenia, schizoaffective disorder, or schizophreniform disorder and longitudinal assessments of substance use were examined. Three groups were formed according to consistency of substance use over time: No/Low Alcohol, Alcohol Only, and Drug Use (with or without alcohol use). Similar groups were formed regarding problems associated with alcohol use. Groups were compared on demographics, psychiatric history, psychopathology, medication side effects, and social functioning. RESULTS: Substance users were more likely to be young, male, and to have lower levels of education. Substance users generally had fewer negative symptoms, more social contacts, and better social-leisure functioning. However, substance users, especially drug users, also were rated as having more interpersonal and family problems, had an earlier age at first psychiatric hospitalization, and were more likely to have been recently hospitalized. Patients reporting problems with alcohol use reported more frequent alcohol and drug use, greater severity of akathisia, and problems in interpersonal, family, and self-efficacy domains. CONCLUSIONS: Many of the same variables that correlate with substance use disorder also correlated with moderate substance use in this sample of people with schizophrenia. Although moderate users of alcohol and drugs may have better social functioning in some areas, they also are likely to have substantial problems in interpersonal relationships, especially those involving family members. PMID- 11278159 TI - Evidence for a mitochondrial oxidative phosphorylation defect in brains from patients with schizophrenia. AB - In-vivo imaging studies and post-mortem studies have demonstrated an impairment of energy metabolism in brains of patients with schizophrenia. Decreased oxidative metabolism has been consistently documented in the frontal lobes. However, the biochemical basis of these changes is unclear. The changes could be caused by reduced requirement of the cells for metabolic energy or an abnormality in energy generation. Neurons generate energy through the respiratory chain in the mitochondria. The respiratory chain consists of five enzyme complexes (I-V). The purpose of the present study was to assess mitochondrial function and test the hypothesis of an underlying oxidative phosphorylation defect in schizophrenia. We analysed spectrophotometrically post-mortem brain specimens of frontal cortex, temporal cortex, basal ganglia, and cerebellum of 12 patients who met the DSM-IV criteria for schizophrenia and of 13 healthy controls for the specific activities of respiratory chain enzymes in the mitochondria. The major finding was that the activity of complex IV was significantly reduced in the frontal cortex (40.9+/-6.7 vs. 87.3+/-12, P=0.003) and in the temporal cortex (39.5+/-6.8 vs. 78+/-10.8, P=0.006) of schizophrenics. In addition, the activity of complexes I+III was significantly reduced in the temporal cortex (2.2+/-0.6 vs. 4.4+/-0.5, P=0.01) and basal ganglia (1.6+/-0.5 vs. 3.4+/-0.3, P=0.015) in schizophrenia. All other enzyme activities showed no differences to healthy controls. The results confirm a defect of oxidative phosphorylation in brains from patients with schizophrenia, which may contribute to impaired energy generation. PMID- 11278160 TI - Estrogen - a potential treatment for schizophrenia. AB - Estrogen has been shown in animal studies to modulate both the dopamine and serotonin neurotransmitter systems - the main neurotransmitters implicated in the pathogenesis of schizophrenia. A double blind, 28 day, placebo-controlled study was conducted with three groups of women of child-bearing age (N=12 in each group) who received standardized antipsychotic medication plus 50mcg transdermal estradiol or 100mcg transdermal estradiol or transdermal placebo. Analyses show that women receiving 100mcg of estradiol made greater improvements in the symptoms of schizophrenia than both the 50mcg estradiol and placebo groups. Women receiving 50mcg estradiol had more improvement in their symptoms compared with the placebo group. The 100mcg estradiol group had significantly lower mean lutenizing hormone (LH) and higher mean prolactin levels across the study period compared with both the 50mcg and placebo groups. The addition of 100mcg adjunctive transdermal estrogen significantly enhanced the treatment of acute, severe psychotic symptoms in women with schizophrenia. The differential response of adding 50mcg versus 100mcg estradiol on the types of symptom affected may be related to the estrogen effect on LH and prolactin. The positive impact of estrogen treatment on psychotic symptoms by a direct effect on dopamine and serotonin systems or via an indirect prolactin-mediated effect may be very useful in the overall treatment of women with schizophrenia. PMID- 11278162 TI - Apolipoprotein E4 stimulates sulfation of glycosaminoglycans in neural cells. AB - Apolipoprotein E externally added to neuroblastoma cells in culture stimulates [35S]sulfate incorporation on cell and extracellular matrix glycosaminoglycans (sGAG). This stimulation is mainly observed for ApoE4 compared to ApoE3. The increase in sulfation is not due to increased synthesis as there is no corresponding increase in the [3H]glucosamine incorporation. Since the presence of ApoE is a risk factor for Alzheimer's disease (AD) and the presence of sGAG could facilitate the assembly of the main components, beta-amyloid and tau proteins, of the aberrant structures found in AD, the present study indicates a possible relation between those factors. PMID- 11278161 TI - Saccadic and smooth-pursuit eye movements in deficit and non-deficit schizophrenia. AB - We have analyzed eye movement performances in schizophrenics showing primary negative or deficit symptoms (n=16) and non-deficit schizophrenics (n=55), and compared them with those of controls (n=34) in order to study the relationships between negative symptoms and eye movement abnormalities. Patients were subtyped into deficit and non-deficit subgroups using the Schedule for the Deficit Syndrome. Three oculomotor paradigms were used: smooth pursuit, a reflexive saccade paradigm and an antisaccadic task. The smooth pursuit gain was significantly decreased (and the rate of catch-up saccades increased) in schizophrenics as compared with controls, but no difference was observed between patient groups. In the reflexive saccade paradigm, no difference was found between controls and patients, except for latency in deficit patients. In the antisaccade paradigm, the number of errors and the latency of successful antisaccades were significantly increased in schizophrenics as compared with controls. The latency of successful antisaccades in both directions was significantly increased in deficit patients as compared with non-deficit patients. The latency of rightward successful antisaccades was significantly increased as compared with the latency of leftward antisaccades in deficit patients only. However, when patients were classified into negative and non negative groups using the PANSS, no difference was found in the antisaccade paradigm. Smooth pursuit impairment does not seem to depend on the primary enduring negative symptoms.In deficit schizophrenics, the abnormalities observed in the antisaccadic task are consistent with prefrontal dysfunction, and may suggest parietal lobe dysfunction as well. PMID- 11278163 TI - Conformational properties of serine proteinase inhibitors (serpins) confer multiple pathophysiological roles. AB - Serine proteinase inhibitors (Serpins) are irreversible suicide inhibitors of proteases that regulate diverse physiological processes such as coagulation, fibrinolysis, complement activation, angiogenesis, apoptosis, inflammation, neoplasia and viral pathogenesis. The molecular structure and physical properties of serpins permit these proteins to adopt a number of variant conformations under physiological conditions including the native inhibitory form and several inactive, non-inhibitory forms, such as complexes with protease or other ligands, cleaved, polymerised and oxidised. Alterations of a serpin which affect its structure and/or secretion and thus reduce its functional levels may result in pathology. Serpin dysfunction has been implicated in thrombosis, emphysema, liver cirrhosis, immune hypersensitivity and mental disorders. The loss of inhibitory activity of serpins necessarily results in an imbalance between proteases and their inhibitors, but it may also have other physiological effects through the generation of abnormal concentrations of modified, non-inhibitory forms of serpins. Although these forms of inhibitory serpins are detected in tissues and fluids recovered from inflammatory sites, the important questions of which conditions result in generation of different molecular forms of serpins, what biological function these forms have, and which of them are directly linked to pathologies and/or may be useful markers for characterisation of disease states, remain to be answered. Elucidation of the biological activities of non-inhibitory forms of serpins may provide useful insights into the pathogenesis of diseases and suggest new therapeutic strategies. PMID- 11278164 TI - Molecular requirements for the internalisation step of endocytosis: insights from yeast. AB - Molecular genetic studies of endocytosis using the unicellular eukaryote Saccharomyces cerevisiae (budding yeast) have led to the identification of many cellular components, both proteins and lipids, required for this process. While initially, many of these requirements (e.g. for actin, various actin-associated proteins, the ubiquitin conjugation system, and for ergosterol and sphingolipids) appeared to differ from known requirements for endocytosis in higher eukaryotes (e.g. clathrin, AP-2, dynamin), it now seems that endocytosis in higher and lower eukaryotes share many requirements. Often, what were initially identified as actin cytoskeleton-associated proteins in S. cerevisiae, are now revealing themselves as clathrin-coated pit- and vesicle-associated proteins in higher eukaryotes. So rather than delineating two endocytic pathways, one actin-based and one clathrin-based, the combined studies on higher and lower eukaryotes are revealing interesting interplay in both systems between the actin cytoskeleton, clathrin coats, and lipids in the formation of endocytic vesicles at the plasma membrane. Recent results from the yeast system show that the Arp2/3p complex, Wiskott-Aldrich syndrome protein (WASP), and WASP-interacting protein (WIP), proteins involved in the nucleation step of actin filament assembly, play a major role in the formation of endocytic vesicles. This discovery suggests models whereby endocytic vesicles may be actively pushed from the plasma membrane and into the cell by newly forming and rapidly extending actin filaments. PMID- 11278165 TI - Identification of the protein product of the Coch gene (hereditary deafness gene) as the major component of bovine inner ear protein. AB - In order to better understand the cause of hereditary hearing impairment, we have performed a proteomic analysis of the inner ear proteins using two-dimensional gel electrophoresis. In the process of analysis, we have found very unique properties of the bovine homologue of the human COCH gene product. The COCH gene is responsible for one of the hereditary hearing impairments, DFNA9, and was recently suggested to be a possible genetic factor contributing to Meniere's disease. The Coch protein constitutes 70% of bovine inner ear proteins and is composed of 16 different protein spots, with charge and size heterogeneity. Heterogeneity of this protein suggests that the Coch gene is processed in several ways, at the transcriptional and/or posttranslational level. Much knowledge has accumulated about the hereditary hearing impairment genes; however, little research has been done regarding the protein products of those genes. This is the first report to characterize the Coch protein. Study of the Coch protein might provide more information on the mechanism of hearing and vestibular disorders. PMID- 11278166 TI - Effects of alveolar surfactant aggregates on T-lymphocyte proliferation. AB - The effects of alveolar large aggregate (LA) and small aggregate (SA) surfactant subfractions isolated from healthy adult rats on mitogen-stimulated proliferative responses of human peripheral blood mononuclear cells (PBMC) was examined. Various concentrations of total surfactant suppressed proliferation of stimulated lymphocytes by up to 95% of mitogen-stimulated cells alone. LA subfractions of total surfactant had no effect on proliferation, whereas SA significantly enhanced the lymphocyte proliferation at lower concentrations (7.8 microg/ml) compared to mitogen-stimulated cells alone. Higher concentrations of SA (62.5 microg/ml) inhibited lymphocyte proliferation. This concentration-dependent effect of SA on proliferation of PBMC was also present when cells were stimulated with various lectins including anti-CD3, concanavalin A and phytohemagglutinin. Analysis of the supernatant of mitogen-stimulated cell cultures treated with inhibitory concentrations of SA showed decreased amounts of interleukin (IL)-2, compared to cells alone, which could be reversed by adding exogenous IL-2 to the cell cultures with the SA. These results suggest that alveolar surfactant subfractions have distinct functions within the alveoli, both biophysically and with respect to their effects on the host's immunomodulatory responses. PMID- 11278167 TI - Stearoyl coenzyme A desaturase 1 expression and activity are increased in the liver during iron overload. AB - In humans, hepatic iron overload can lead to hepatocellular carcinoma development. Iron related dysregulation of hepatic genes could play a role in this phenomenon. We previously found that the carbonyl-iron overloaded mouse was a useful model to study the mechanisms involved in the development of hepatic lesions related to iron excess. The aim of the present study was to identify hepatic genes overexpressed in conditions of iron overload by using this model. A suppressive subtractive hybridization was performed between hepatic mRNAs extracted from control and 3% carbonyl-iron overloaded mice during 8 months. This methodology allowed us to identify stearoyl coenzyme A desaturase 1 (SCD1) mRNA overexpression in the liver of iron loaded mice. The corresponding enzymatic activity was also found to be significantly increased. In addition, we demonstrated that both SCD1 mRNA expression and activity were increased in another iron overload model in mice obtained by a single iron-dextran subcutaneous injection. Moreover, we found, in both models, that SCD1 mRNA was not only influenced by the quantity of iron in the liver but also by the duration of iron overload since SCD1 mRNA upregulation was not detected in earlier stages of iron overload. In addition, we found that cellular repartition likely influenced SCD1 mRNA expression. In conclusion, we demonstrated that iron excess in the liver induced both the expression of SCD1 mRNA and its corresponding enzymatic activity. The level and duration of iron overload, as well as cellular repartition of iron excess in the liver likely play a role in this induction. The fact that the expression and activity of SCD1, an enzyme adding a double bound into saturated fatty acids, are induced in two models of iron overload in mice leads to the conclusion that iron excess in the liver may enhance the biosynthesis of unsaturated fatty acids. PMID- 11278169 TI - The use of Pseudomonas acyl-CoA synthetase to form acyl-CoAs from dicarboxylic fatty acids. AB - Pseudomonas acyl-CoA synthetase is shown to act on saturated dicarboxylic acids with a chain length of C10 or greater to produce conjugates containing a single CoA unit. The synthetase can, therefore, be used to generate novel acyl-CoA analogues for studies on proteins that utilise, bind to, or are modulated by acyl CoAs. PMID- 11278168 TI - Enhanced expression of a-series gangliosides in fibroblasts of patients with peroxisome biogenesis disorders. AB - Peroxisome biogenesis disorders (PBD) are classified into Zellweger syndrome (ZS), infantile Refsum disease (IRD) and neonatal adrenoleukodystrophy. Disturbances in the differentiation of neural cells such as migration arrest are characteristic of PBD. So far the pathogenesis of these disturbances is not clearly understood. We describe an altered metabolism of glycosphingolipids in PBD which has not yet been investigated. We observed an increased amount of a series gangliosides, GM2, GM1 and GD1a, in the fibroblasts of patients with ZS and IRD. Gangliosides GM1 and GD1a were not present in detectable amounts in normal subjects. A key step in the synthesis of a-series gangliosides is a transfer of GalNAc to ganglioside GM3, so we determined the level of ganglioside GM3 by immunohistochemical methods. We found a granular structure, which was positive toward anti-ganglioside GM3 antibody in the cytoplasm of the patients' fibroblasts. In control cells, the cell membrane was slightly positive toward anti-GM3 antibody. These results may help to clarify the pathogenesis of PBD with respect to the functional roles of glycosphingolipids in cell differentiation, proliferation and apoptosis. PMID- 11278170 TI - Structural models of human apolipoprotein A-I: a critical analysis and review. AB - Human apolipoprotein (apo) A-I has been the subject of intense investigation because of its well-documented anti-atherogenic properties. About 70% of the protein found in high density lipoprotein complexes is apo A-I, a molecule that contains a series of highly homologous amphipathic alpha-helices. A number of significant experimental observations have allowed increasing sophisticated structural models for both the lipid-bound and the lipid-free forms of the apo A I molecule to be tested critically. It seems clear, for example, that interactions between amphipathic domains in apo A-I may be crucial to understanding the dynamic nature of the molecule and the pathways by which the lipid-free molecule binds to lipid, both in a discoidal and a spherical particle. The state of the art of these structural studies is discussed and placed in context with current models and concepts of the physiological role of apo A-I and high-density lipoprotein in atherosclerosis and lipid metabolism. PMID- 11278171 TI - Oil-bodies as substrates for lipolytic enzymes. AB - Plant seeds store triacylglycerols (TAGs) in intracellular organelles called oil bodies or oleosomes, which consist of oil droplets covered by a coat of phospholipids and proteins. During seed germination, the TAGs of oil-bodies hydrolysed by lipases sustain the growth of the seedlings. The mechanism whereby lipases gain access to their substrate in these organelles is largely unknown. One of the questions that arises is whether the protein/phospholipid coat of oil bodies prevents the access of lipase to the oil core. We have investigated the susceptibility of almond oil-bodies to in vitro lipolysis by various purified lipases with a broad range of biochemical properties. We have found that all the enzymes assayed were capable of releasing on their own free fatty acids from the TAG of oil-bodies. Depending on the lipase, the specific activity measured on oil bodies using the pH-stat technique was found to range from 18 to 38% of the specific activity measured on almond oil emulsified by gum arabic. Some of these lipases are known to have a dual lipase/phospholipase activity. However, no correlation was found to exist between the ability of a lipase to readily and efficiently hydrolyse the TAG content of oil-bodies and the presence of a phospholipase activity. Kinetic studies indicate that oil-bodies behave as a substrate as other proteolipid organelles such as milk fat globules. Finally we have shown that a purified water-soluble plant lipase on its own can easily hydrolyse oil-bodies in vitro. Our results suggest that the lipolysis of oil bodies in seedlings might occur without any pre-hydrolysis of the protein coat. PMID- 11278172 TI - Dual effects of lysophosphatidic acid on human airway smooth muscle cell proliferation and survival. AB - Lysophosphatidic acid (LPA) is a phospholipid growth mediator found in serum at 2 20 microM. In many cell types, including human airway smooth muscle (HASM) cells, LPA-induced proliferation occurs at 10-100 microM LPA. At these concentrations LPA forms Ca2+ precipitates. The potential involvement of Ca2+ and Ca2+ LPA precipitates in LPA-induced HASM cell mitogenesis was investigated. In the absence of extracellular Ca2+, 10 and 30 microM LPA stimulated HASM cell mitogenesis. However, with 100 microM LPA in the absence of extracellular Ca2+, HASM cells exhibited a profound shape change and loss of viability, determined to be apoptosis by both DNA staining and assessment of cytosolic nucleosomal reactivity. A bioassay based on the adenosine 3':5'-cyclic monophosphate response of C62B rat glioma cells was used to measure the bioactivity of LPA solutions prepared in Ca2+ free and Ca2+ containing medium. After 24 h, a 100 microM LPA solution in Ca2+ free medium contained markedly greater bioactivity than a 100 microM LPA solution made in Ca2+ containing medium. In summary, formation of Ca2+ LPA precipitates decreases the amount of biologically active LPA in solution, and high concentrations of bioactive LPA achieved in Ca2+ free but not in Ca2+ containing medium induce apoptosis of HASM cells. PMID- 11278174 TI - Lysophosphatidylcholine derived from deer antler extract suppresses hyphal transition in Candida albicans through MAP kinase pathway. AB - A family of 2-lysophosphatidylcholines (lyso-PCs) was isolated from deer antler extract, guided exclusively by hyphal transition inhibitory activity in Candida albicans. Structural determination of the isolated lyso-PCs by spectroscopic methods, including infrared spectroscopy, 1H nuclear magnetic resonance (NMR), 13C NMR, 2D correlation spectroscopy NMR, fast atom bombardment mass spectrometry and tandem mass spectrometry, confirmed that the natural products were composed of at least four different lyso-PCs varying in fatty acid moiety at the sn-1 position of the glycerol backbone. The major lyso-PCs were confirmed as 1 stearoyl-, 1-oleoyl-, 1-linoleoyl- and 1-palmitoyl-2-lyso-sn-glycero-3 phosphatidylcholines. Lyso-PC specifically suppressed the morphogenic transition from yeast to hyphae in C. albicans, without affecting the growth of either yeast or hyphae. Lyso-PC exerted hyphal transition that suppressed activity in the broad spectrum of the Candida species, such as C. albicans, Candida krusei, Candida guilliermondii and Candida parapsilosis. Northern analysis indicated that the suppression was mediated through the mitogen-activated protein kinase pathway. PMID- 11278173 TI - The expression of PPAR-associated genes is modulated through postnatal development of PPAR subtypes in the small intestine. AB - In this study, we found that the mRNA level of peroxisome proliferator-activated receptor (PPAR) alpha, but not of PPARdelta, was elevated in the jejunum during the postnatal development of the rat. Moreover, we found that the expressions of PPAR-dependent genes, such as acyl-CoA oxidase, L-FABP, and I-FABP, were also increased during the postnatal development of the small intestine. Electrophoretic mobility shift assay revealed that both the PPARalpha-9-cis retinoic acid receptor alpha (RXRalpha) heterodimer and the PPARdelta-RXRalpha heterodimer bound to the peroxisome proliferator response element (PPRE) of acyl CoA oxidase and L-FABP genes. The binding of the PPARalpha-RXRalpha heterodimer to the PPREs of the various genes was enhanced by the addition of PPARalpha, with a concomitant reduction of the binding of PPARdelta-RXRalpha to the PPREs. Furthermore, the binding activity of PPARalpha-RXRalpha, but not PPARdelta RXRalpha, to the PPREs was enhanced by the addition of a PPAR ligand, WY14,643. The GAL4-PPAR-chimera reporter assay showed that WY14,643 transactivated the reporter gene through action of PPARalpha, but not through PPARdelta, in Caco-2 cells. Furthermore, oral administration of a PPAR ligand, clofibrate, during 3 consecutive days of the weanling period caused a parallel increase in the mRNA levels of these PPAR-dependent genes. These results suggest that acyl-CoA oxidase, L-FABP and the other PPAR-dependent genes in the small intestine may be coordinately modulated during postnatal development by the disproportional expression of PPARalpha over PPARdelta. PMID- 11278175 TI - Impaired spreading of surfactant phospholipids in the lungs of newborn rats with pulmonary hypoplasia as a model of congenital diaphragmatic hernia induced by nitrofen. AB - In order to clarify the pathological outcome of congenital diaphragmatic hernia (CDH), we devised an animal model of CDH by administration of 2,4-dichlorophenyl p-nitrophenyl ether (nitrofen) to pregnant rats, and determined the level and distribution of lung surfactant using the monoclonal antibody toward sphingomyelin and disaturated phosphatidylcholine (disat-PC). In control rats, the concentration of disat-PC was found to increase greatly from 16 to 18 days of gestation. Intragastric administration of nitrofen to pregnant rats at day 9 of gestation resulted in CDH in 42.7% of fetuses delivered after 20 days of gestation. In nitrofen-treated fetuses, the concentration of disat-PC in the lungs was lower than those in control fetuses, and surfactant apoprotein SP-A was similarly reduced in nitrofen-treated fetuses. However, the concentration of disat-PC in nitrofen-treated fetuses was higher than that in control fetuses at 18 days of gestation, indicating a synthetic potential of surfactant in nitrofen treated fetuses comparable to that at the late stage of normal gestation. Immunohistochemical study with the antibody revealed that surfactant phospholipid was mainly in the form of intracellular granules in nitrofen-treated fetuses, probably causing the hypoplastic lungs and then CDH, in contrast to the uniform distribution on the pulmonary alveolar surface in control fetuses. PMID- 11278176 TI - Immunoadjuvant activity of interferon-gamma-liposomes co-administered with influenza vaccines. AB - In an attempt to potentiate the relatively low immunogenicity of the currently used influenza vaccines, especially in high-risk groups, monovalent and divalent subunit vaccine preparations were co-administered with free or liposome associated murine interferon gamma (mIFNgamma) as an adjuvant. Recombinant murine IFNgamma was entrapped (50-70% efficiency) in two types of large multilamellar vesicles: mIFNgamma-LIP A-'conventional' liposomes, and mIFNgamma-LIP B- 'surface depleted' liposomes, in which 60 and 8% of the associated cytokine was located at the external liposome membrane, respectively. Subunit preparations containing the viral surface proteins hemagglutinin and neuraminidase (HN) were injected once, i.p. (0.5 microg each), into BALB/c mice, alone and combined with free or liposomal mIFNgamma (mIFNgamma-LIP, 0.5 or 3.0 microg). Sera were tested 3-16 weeks post-vaccination by hemagglutination inhibition (HI), and by ELISA for IgG1 and IgG2a antibodies (Abs). In addition, protective immunity against intranasal viral infection was assayed at 11 and 17 weeks post-vaccination. The results showed that: (a) Vaccination with HN alone produces very low HI and IgG titers and does not afford any protection. (b) Although co-administration with free mIFNgamma (particularly using 3.0 microg) markedly enhances HI titer as well as the IgG1 and IgG2a levels, protection is negligible (0-33%). (c) In most cases, mIFNgamma-LIP is significantly more potent than free mIFNgamma (2-40-fold increase in Ab titer), and the low dose (0.5 microg) is generally more efficient than the high dose. Up to 83% of the mice co-vaccinated with mIFNgamma-LIP were protected against viral challenge. (d) Both the IgG2a level and the HI titer appear to be crucial for protection. (e) Although the two liposomal preparations differ in their cytokine release profile in vivo and in their bioactivity in vitro, their adjuvant activity is comparable. PMID- 11278177 TI - The relationship between 1H-NMR mobile lipid intensity and cholesterol in two human tumor multidrug resistant cell lines (MCF-7 and LoVo). AB - The high resolution proton nuclear magnetic resonance (1H-NMR) spectra of two different cell lines exhibiting multidrug resistance (MDR) as demonstrated by the expression of the well-known energy-driven, membrane-bound 170 kDa P-glycoprotein pump known as Pgp were investigated. In particular, the mobile lipid (ML) profile, and the growth and biochemical characteristics of MCF-7 (human mammary carcinoma) and LoVo (human colon adenocarcinoma) sensitive and resistant tumor cells were compared. The results indicate that both MCF-7 and LoVo resistant cells have a higher ML intensity than their respective sensitive counterparts. However, since sensitive and resistant cells of each pair grow in the same manner, variations in growth characteristics do not appear to be the cause of the ML changes as has been suggested by other authors in non-resistant tumor cells. In order to investigate further the origin of the ML changes, lipid analyses were conducted in sensitive and resistant cell types. The results of these experiments show that resistant cells of both cell types have a greater amount of esterified cholesterol and saturated cholesteryl ester and triglyceride fatty acid than their sensitive counterparts. From a thorough analysis of the data obtained in this paper utilizing numerous techniques including biological, biophysical and biochemical ones, it is hypothesized that cholesterol and triglyceride play a pivotal role in inducing changes in NMR ML signals. The importance of these lipid variations in MDR is discussed in view of the controversy regarding the origin of ML signals and the paramount role played by the Pgp pump in resistance. PMID- 11278178 TI - The novel sequences of major plasma apolipoproteins in the eel Anguilla japonica. AB - cDNAs encoding major plasma apolipoproteins (apo) were cloned from the eel Anguilla japonica liver and their nucleotide sequences determined. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that eel lipoproteins contain apolipoproteins of 28 kDa and 14 kDa as major components. Each of the two apolipoproteins showed two isoforms having different isoelectric points as demonstrated by two-dimensional electrophoresis. The two 28 kDa components had different N-terminal amino acid sequences, whereas the two 14 kDa components had an identical one. Then cDNA clones encoding these apolipoproteins were isolated from a cDNA library constructed from the eel liver. An acidic 28 kDa component (28 kDa-1) consisted of 259 amino acids including a putative signal peptide of 27 residues, whereas a basic 28 kDa component (28 kDa-2) was composed of 260 amino acids containing a putative signal peptide of 23 residues. The tandem repeating units, which are characteristic of apolipoproteins, for 28 kDa-1 showed 27.8% identity to that of porcine apoA-IV, although mammalian apoA-IV is about 40 kDa and much larger than 28 kDa-1. However, the repeating units of 28 kDa-2 showed 52.5% identity to that of Atlantic salmon apoA-I. The 14 kDa apolipoprotein consisted of 142 amino acids containing a putative signal peptide of 20 residues. It has a novel sequence differing from apolipoproteins of other vertebrates. The transcriptional expressions of 28 kDa-1, 28 kDa-2, and 14 kDa components were all restricted to the liver, except for the transcripts of 28 kDa-2 which were also slightly expressed in the intestine. PMID- 11278179 TI - t-Butyl hydroperoxide and oxidized low density lipoprotein enhance phospholipid hydrolysis in lipopolysaccharide-stimulated retinal pericytes. AB - Free radicals induced by organic peroxides or oxidized low density lipoprotein (oxLDL) play a critical role in the development of atherosclerosis. In investigating this process, and the concomitant inflammatory response, the role of pericytes, cells supporting the endothelial ones in blood vessels, has received little attention. In this study we tested the hypothesis that tert-butyl hydroperoxide (t-BuOOH) and oxLDL, administered in sublethal doses to the culture medium of retinal pericytes, function as prooxidant signals to increase the stimulation of the peroxidation process induced by lipopolysaccharide (LPS). Confluent cell monolayers were exposed to t-BuOOH (25-400 microM), native LDL or oxLDL (3.4-340 nmol hydroperoxides/mg protein, 1-100 micro). LPS (1 microg/ml), t BuOOH (200 microM), and oxLDL (100 microM), but not native LDL, incubated for 24 h with cells, markedly increased lipid peroxidation, cytosolic phospholipase A2 (cPLA2) activity and arachidonic acid (AA) release in a time- and dose-dependent manner. AACOCF(3), a potent cPLA2 inhibitor, and the antioxidant alpha-tocopherol strongly inhibited the prooxidant-stimulated AA release. Long-term exposure to maximal concentrations of t-BuOOH (400 microM) or oxLDL (100 microM) had a sharp cytotoxic effect on the cells, described by morphological and biochemical indices. The presence of t-BuOOH or oxLDL at the same time, synergistically increased phospholipid hydrolysis induced by LPS alone. 400 microM t-BuOOH or 100 microM oxLDL had no significant effect on the stimulation of an apoptosis process estimated by DNA laddering and light and electron microscopy. The results indicate that (i) pericytes may be the target of extensive oxidative damage; (ii) activation of cPLA2 mediates AA liberation; (iii) as long-term regulatory signals, organic peroxide and specific constituents of oxLDL increase the pericyte ability to degrade membrane phospholipids mediated by LPS which was used, in the present study, to simulate in vitro an inflammatory burst in the retinal capillaries. PMID- 11278180 TI - Ethyl docosahexaenoate-associated decrease in fetal brain lipid peroxide production is mediated by activation of prostanoid and nitric oxide pathways. AB - Previously we have shown that intraamniotic administration of ethyl docosahexaenoate (Et-DHA) to pregnant rats resulted in decreased lipid peroxidation in the fetal brain, under a variety of conditions (S. Glozman, P. Green, E. Yavin, J. Neurochem. 70 (1998) 2482-2491). In the present study we examine the potential mechanisms to explain this effect. This was done by a pharmacological approach, utilizing brain slice preparations from Et-DHA treated or control rats in the presence of various agents and examining the formation of products in the tissue slices or incubation medium. Et-DHA treated brains produced 2-3-fold more prostanoids (PN) than control brains, indicating cyclooxygenase (COX) activation. Indomethacin at 50 microM inhibited PN formation and also abolished Et-DHA induced decrease in lipid peroxides, as evident by the levels of thiobarbituric acid reactive substances (TBARS) released in the medium. The phospholipase A2 inhibitors quinacrine and p-bromophenacyl bromide added at 0.1 mM concentration each to either slices from controls or Et-DHA treated fetal brains, decreased TBARS production. Et-DHA treated brains released 2.2-fold more nitric oxide (NO) than control brains and NO synthase (NOS) inhibitors abolished this effect. Increasing the concentration of NO by the addition of an NO donor greatly decreased the concentration of the TBARS in the medium. These results suggest that at least some of the effect of Et-DHA on decreased lipid peroxidation may be explained by a shift of oxygen species utilization via enzymatically regulated, therefore metabolically controlled, COX and NOS activities. PMID- 11278181 TI - MR imaging and spectroscopy of brain development. AB - MR imaging provides an invaluable tool for the study of brain development in vivo. Current MR imaging techniques allow noninvasive methodologies, without ionizing radiation, that provide a diversity of information on structure, metabolism, and function of the developing brain. This article focuses on the application of conventional and advanced MR imaging techniques, including quantitative morphometric MR imaging, diffusion-weighted, functional MR, and MR spectroscopic imaging to the study of early human brain development. PMID- 11278182 TI - Fetal brain MR imaging. AB - In the past 10 years, improvements in MR imaging and faster imaging techniques have dramatically increased the use of in-utero fetal brain MR imaging. Challenging abnormalities now can be diagnosed prenatally through a careful analysis of morphology and signal changes. Using illustrations of normal brain development as a starting point, this article illustrates and discusses major brain malformations and specific morphologic changes, destructive lesions, and isolated ventriculomegalies, and the advantages and disadvantages of T1 and T2 sequences are provided. PMID- 11278183 TI - Neonatal MR imaging. AB - Recently, MR imaging has become the technique of choice in evaluating neonatal central nervous system diseases. It is the only imaging technique that can discriminate myelinated from neonatal unmyelinated white matter; it offers the highest sensitivity in detecting acute anoxic injury of the neonatal brain; and with proper coils and sequences, it can exquisitely depict neonatal brain anatomy and locate pathology, offering a robust and reliable tool in the prognostic assessment of neonatal central nervous system disease. PMID- 11278184 TI - Diffusion-weighted imaging of the brain in neonates and infants. AB - Diffusion-weighted imaging provides novel and interesting insights into normal development and pathologic processes occurring in neonates and infants. Both myelinated and unmyelinated white matter show restricted diffusion. Focal infarction in perinatal stroke and more global injury associated with hypoxic ischemia encephalopathy produce high-signal lesions in the brain, in the acute phase. Diffusion-weighted imaging also demonstrates abnormalities of a variety of other diseases, when conventional imaging is relatively uninformative. PMID- 11278185 TI - Imaging of the developmentally delayed child. AB - Neuroimaging can aid in determining the causes of developmental delay in a child. Information from neuroimaging examinations may help guide further testing and treatment in these children. This article emphasizes the neuroimaging approaches, modalities, and features in the child with developmental delay. The common causative categories of developmental delay are reviewed. PMID- 11278186 TI - Neuroimaging of epilepsy in children. AB - Epilepsy is, above all, a pediatric disease. The electroclinical expression of epilepsy depends on several factors, including the presence of a subjacent brain anomaly and the age of the child. Genetic, congenital, and perinatal disorders account for most of the cases. MR imaging should identify any abnormality of the brain, in order to understand the condition and, in selected cases, to be able to treat the patient surgically. PMID- 11278187 TI - Holoprosencephaly: new concepts. AB - Holoprosencephaly represents a broad spectrum of malformations resulting from a lack of separation of the structures of the forebrain. Recent discoveries in the fields of genetics and developmental neurobiology have advanced our knowledge of this complex disorder. By combining this basic-science knowledge with observations of brain morphology, we can better understand the embryology and genetic factors that influence brain development and, ultimately, form more accurate classification systems and stratification measures for predicting patient outcome. PMID- 11278188 TI - MR spectroscopy in pediatric neuroradiology. AB - MR spectroscopy of the pediatric brain now has entered the clinical arena as a result of enhanced technology, complementary new sequences, and proof of clinical utility. Armed with a knowledge of the variation in metabolite concentrations over time, with myelination and brain growth, proton MR spectroscopy can be helpful in differential diagnosis, management, and prognostication of pediatric disease processes. Although other molecules can be interrogated, hydrogen proton spectroscopy is the mainstay. The development of multiplanar techniques, performed in a time-efficient fashion, has enabled more robust spectra to be obtained from larger volumes of brain, permitting spatial localization of different metabolites, such as lactate. With the introduction of gene therapy and other new interventions, a noninvasive tool such as MR spectroscopy may prove to be invaluable. PMID- 11278189 TI - Ophthalmologic disease in children. AB - MR imaging has become essential in the evaluation of a spectrum of ophthalmologic diseases in children. Orbit, globe, optic nerve, and optic tract disease processes that are frequently evaluated by MR imaging include congenital malformations, and inflammatory, neoplastic, and traumatic lesions. MR imaging evaluation aids in differentiating these lesions for diagnosis, the extent of disease, treatment planning, follow-up, and prognosis. PMID- 11278190 TI - Perfusion imaging in the pediatric patient. AB - The prevalence of cerebrovascular disease in children is much higher than most clinicians and neuroradiologists suspect, when all primary and secondary causes are considered. Most signal alterations found on MR imaging in childhood central nervous system pathologic conditions result from causes other than a decrease in tissue perfusion. In addition to conventional MR imaging, the ability to assess changes in tissue water by diffusion imaging and tissue perfusion by perfusion weighted imaging can prove useful to asses cerebral hemodynamics in various pathologic disorders. Exogenous contrast bolus dynamic perfusion-weighted imaging is especially useful in children to differentiate between ischemic injury and other conditions that may alter T2 relaxation, such as demyelination and edema. Perfusion imaging has proved to be a robust and valuable tool to assess the hemodynamic component in childhood CNS disease related to neoplasms and complications from their therapy, cerebrovascular occlusive disease, childhood CNS arteriopathies and trauma. PMID- 11278191 TI - Functional MR imaging in pediatrics. AB - Functional magnetic resonance (fMR) imaging can show neuronal structures underlying specific perceptual and cognitive processes. With the aid of fMR imaging, the development of brain functions can be followed, and deviation from the normal pattern can be established quickly. This article discusses the unique issues of fMR imaging in the pediatric population (e.g., the occurrence of a negative blood oxygenation-level dependent [BOLD] signal during visual stimulation in the age group in whom the synaptic density is the highest; in older children, when synaptic pruning has proceeded, the BOLD signal takes on the positive characteristics seen in adults). fMR imaging also suggests prospectively important applications in the diagnostic workup of children: an early diagnosis of functional deficit can reduce residual deficits to a minimum because remediation, such as specialized training, can be started at an early stage. PMID- 11278192 TI - A hierarchical axis of object processing stages in the human visual cortex. AB - How are objects represented in the human visual cortex? Two conflicting theories suggest either a holistic representation, in which objects are represented by a collection of object templates, or a part-based representation, in which objects are represented as collections of features or object parts. We studied this question using a gradual object-scrambling paradigm in which pictures of objects (faces and cars) were broken in a stepwise manner into an increasing number of blocks. Our results reveal a hierarchical axis oriented anterior--posteriorly in the organization of ventral object-areas. Along this axis, representations are arranged in bands of increasing sensitivity to image scrambling. The axis starts in early visual areas through retinotopic areas V4/V8 and continues into the lateral-occipital sulcus dorsally and the posterior fusiform girus ventrally, corresponding together to the previously described object-related lateral occipital complex (LOC). Regions showing the highest sensitivity to scrambling tended to be located at the most anterior-lateral regions of the complex. In these more anterior regions, breaking the images into 16 parts produced a significant reduction in activation. Interestingly, activation was not affected when images were cut in two halves, either horizontally or vertically. Car images generally produced a weaker activation compared to faces in the lateral occipital complex but showed the same tendency of increased scrambling sensitivity along the anterior--posterior axis. These results suggest the existence of a hierarchical axis along ventral occipito-temporal object-areas, in which the neuronal properties shift from sensitivity to local object features to a more global and holistic representation. PMID- 11278193 TI - Where is 'dorsal V4' in human visual cortex? Retinotopic, topographic and functional evidence. AB - In flattened human visual cortex, we defined the topographic homologue of macaque dorsal V4 (the 'V4d topologue'), based on neighborhood relations among visual areas (i.e. anterior to V3A, posterior to MT+, and superior to ventral V4). Retinotopic functional magnetic resonance imaging (fMRI) data suggest that two visual areas ('LOC' and 'LOP') are included within this V4d topologue. Except for an overall bias for either central or peripheral stimuli (respectively), the retinotopy within LOC and LOP was crude or nonexistent. Thus the retinotopy in the human V4d topologue differed from previous reports in macaque V4d. Unlike some previous reports in macaque V4d, the human V4d topologue was not significantly color-selective. However, the V4d topologue did respond selectively to kinetic motion boundaries, consistent with previous human fMRI reports. Because striking differences were found between the retinotopy and functional properties of the human topologues of 'V4v' and 'V4d', it is unlikely that these two cortical regions are subdivisions of a singular human area 'V4'. PMID- 11278194 TI - Somatotopy in human primary motor and somatosensory hand representations revisited. AB - High-resolution functional magnetic resonance imaging of healthy volunteers was used to study the functional anatomy of the human primary motor (M1) and somatosensory (S1) cortical hand representations during simple movements of thumb, little finger and wrist and a sequential movement of the middle three fingers. Rest served as a control state. The results demonstrated an orderly somatotopy in both M1 and S1, even though the cortical areas active with individual movements significantly overlapped. Moreover, the activation patterns in M1 and S1 differed in three aspects: (i) S1 activation was distributed into significantly more clusters than M1 and the primary cluster was smaller; (ii) the overlaps of areas active with different movements were significantly larger in M1 than in S1; (iii) the difference between the three-finger sequential movement and the single-finger movements was more pronounced in S1 than in M1. The sequence activated S1 cortex was distributed into significantly more clusters. There was also a trend for a bigger volume difference between sequence and the single finger movements in S1 than M1. These data suggest that while the distributed character dominates in M1 and S1, a somatotopic arrangement exists for both M1 and S1 hand representations, with the S1 somatotopy being more discrete and segregated, in contrast to the more integrated and overlapping somatotopy in M1. PMID- 11278195 TI - Remembering the color of objects: an ERP investigation of source memory. AB - Subjects studied pictures of common objects outlined in either red or green and were asked to memorize the objects and their associated colors. Event-related potentials (ERPs) were recorded during subsequent inclusion (i.e. item) and exclusion (i.e. source) memory tasks. The main goal of the experiment was to determine if brain signatures for familiarity and recollection, two behavioral processes thought to account for episodic memory performance, would be observed in the pattern of ERP results. For correctly recognized items, early, posterior old/new effects were recorded (approximately 300--600 ms) that did not differ in magnitude or scalp distribution between item and source memory tasks. A subsequent long-duration occipitally focused negativity (approximately 800 ms peak) was evident in the source but not the item memory task. The ERPs associated with 'source errors' in the source memory task also showed robust early old/new effects. However, 'source error' ERPs lacked frontal scalp activity compared to those associated with correct source attribution. The data suggest that a recollective response may require frontal involvement whereas a decision based on familiarity may not. PMID- 11278196 TI - Age-related volumetric changes of brain gray and white matter in healthy infants and children. AB - To date there is little information about brain development during infancy and childhood, although several quantitative studies have shown volume changes in adult brains. We performed three-dimensional magnetic resonance imaging (3D-MRI) in 28 healthy children aged 1 month to 10 years. We examined the volumes of whole brain and frontal and temporal lobes with an advanced method for segmenting images into gray matter (GM), white matter (WM) and cerebrospinal fluid (CSF) compartments. Growth spurts of whole brain and frontal and temporal lobes could be seen during the first 2 years after birth. During this period the frontal lobes grew more rapidly than the temporal lobes, the right--left asymmetry was more noticeable in the temporal lobes than in the frontal lobes and the increase in GM was larger than that in WM in the temporal lobes. Subsequently, WM volume increased at a higher rate than GM volume throughout childhood. Quantitative information on normal brain development may play a pivotal role in clarifying brain neurodevelopmental abnormalities. PMID- 11278197 TI - Caffeine-induced telencephalic vesicle evagination in early post-implantation mouse embryos involves cAMP-dependent protein kinase (PKA) inhibition. AB - Other studies have shown that caffeine accelerates telencephalic vesicle evagination in early post-implantation mouse embryos. The present study examines the effect of caffeine on gene modulation in post-implantation mouse embryos. Using mRNA differential display, we observed that caffeine increased gene expression of the regulatory subunit (RI alpha) of cAMP-dependent protein kinase (PKA). RT--PCR analysis confirmed an increase in expression of this gene in caffeine-exposed embryos when compared with saline-treated controls. Using a fluorescent substrate of PKA, we found that PKA activity in the presence of cAMP was lower in caffeine-treated embryos than in controls. Treatment with H89 and PKI(12-24)amide, two inhibitors of PKA activity, mimicked the effects of caffeine on telencephalic vesicle formation. Together these data suggest that in early post-implantation mouse embryos caffeine modulates gene expression of the RI alpha subunit of PKA and that caffeine-induced inhibition of PKA activity plays a role in early telencephalic evagination. PMID- 11278198 TI - Dissociating neural correlates of cognitive components in mental calculation. AB - Mental calculation is a complex cognitive operation that is composed of a set of distinct functional processes. Using functional magnetic resonance imaging (fMRI), we mapped brain activity in healthy subjects performing arithmetical tasks and control tasks evoking a comparable load on visuo-constructive, linguistic, attentional and mnemonic functions. During calculation, as well as non-mathematical tasks, similar cortical networks consisting of bilateral prefrontal, premotor and parietal regions were activated, suggesting that most of these cortical areas do not exclusively represent modules for calculation but support more general cognitive operations that are instrumental but not specific to mental arithmetic. Significant differences between calculation and the non mathematical tasks were found in parietal sub-regions, where non-arithmetic number or letter substitution tasks preferentially activated the superior parietal lobules whereas calculation predominantly elicited activation of the left dorsal angular gyrus and the medial parietal cortices. We interpret the latter activations to reflect sub-processes of mental calculation that are related to the processing of numerical representations during exact calculation and to arithmetical fact retrieval. Finally, we found that more complex calculation tasks involving the application of calculation rules increased activity in left inferior frontal areas that are known to subserve linguistic and working memory functions. Taken together, these findings help to embed the specific cognitive operation of calculation into a neural framework that provides the required set of instrumental components. This result may further inform the cognitive modeling of calculation and adds to the understanding of neuropsychological deficit patterns in patients. PMID- 11278199 TI - Relationship between EEG potentials and intracellular activity of striatal and cortico-striatal neurons: an in vivo study under different anesthetics. AB - The functions of the basal ganglia are achieved through excitation of striatal output neurons (SONs) by converging cortical glutamergic afferents. We assessed the relationship between different patterns of activity in cortico-striatal (C-S) cells and the electrical behavior of SONs in vivo. Intracellular activities of rat C-S neurons in the orofacial motor cortex and of SONs, located in the projection field of this cortical region, were recorded under different anesthetics, which generate various temporal patterns of cortical activity. A surface electroencephalogram (EEG) of the orofacial motor cortex was simultaneously performed with intracellular recordings and EEG waves were used as correlates of a coherent synaptic activity in cortical neurons. Under barbiturate anesthesia C-S neurons showed rhythmic (5--7 Hz) supra-threshold depolarizations in phase with large amplitude EEG waves. The correlative activity of SONs was characterized by large amplitude oscillation-like synaptic depolarizations that could trigger action potentials. Under ketamine-xylazine anesthesia C-S neurons exhibited a step-like behavior consisting of depolarizing plateaus (up states), leading to multiple spike discharges, interrupted by hyperpolarizing periods (down states). The related activity of SONs was step-like membrane potential fluctuations with firing confined to the early part of the striatal up state. In C-S neurons and SONs up states coincided with slow recurrent EEG waves (approximately 1 Hz). Finally, under neurolept-analgesia an apparently disorganized EEG activity was associated with a lack of rhythmic discharge in C-S neurons. This uncorrelated activity in C-S neurons resulted in an absence of spontaneous firing as well as of large amplitude synaptic depolarizations in SONs. In the present study we demonstrate that SONs shape their input-output relationship by filtering out uncorrelated synaptic activity and that a minimal synchronization in the cortico-striatal afferents is required to produce significant synaptic depolarization in SONs. PMID- 11278200 TI - Prefrontal gray matter volume reduction in first episode schizophrenia. AB - Functional measures have consistently shown prefrontal abnormalities in schizophrenia. However, structural magnetic resonance imaging (MRI) findings of prefrontal volume reduction have been less consistent. In this study, we evaluated prefrontal gray matter volume in first episode (first hospitalized) patients diagnosed with schizophrenia, compared with first episode patients diagnosed with affective psychosis and normal comparison subjects, to determine the presence in and specificity of prefrontal abnormalities to schizophrenia. Prefrontal gray and white matter volumes were measured from first episode patients with schizophrenia (n = 17), and from gender and parental socio-economic status-matched subjects with affective (mainly manic) psychosis (n = 17) and normal comparison subjects (n = 17), age-matched within a narrow age range (18- 29 years). Total (left and right) prefrontal gray matter volume was significantly reduced in first episode schizophrenia compared with first episode affective psychosis and comparison subjects. Follow-up analyses indicated significant left prefrontal gray matter volume reduction and trend level reduction on the right. Schizophrenia patients showed 9.2% reduction on the left and 7.7% reduction on the right compared with comparison subjects. White matter volumes did not differ among groups. These data suggest that prefrontal cortical gray matter volume reduction is selectively present at first hospitalization in schizophrenia but not affective psychosis. PMID- 11278202 TI - The Mesothelium, Teflon or Velcro? Mesothelium in endometriosis pathogenesis. AB - Sampson's transplantation theory for the pathogenesis of peritoneal endometriosis is widely accepted. The events that take place, however, on the cellular and subcellular level during the transition of endometrial tissue in the abdominal cavity into peritoneal endometriosis remain controversial. The mesothelium plays a central role in the debate on this subject. The interaction between endometrium and peritoneum has been studied in an in-vitro model using amnion, peritoneum and mesothelial cells in culture on the one hand and cyclic and menstrual endometrium on the other hand. The results of these studies indicate that (i) an intact mesothelial lining prevents adhesion of shed endometrial tissue, (ii) shed endometrial tissue adheres to the peritoneal extracellular matrix and (iii) menstrual effluent creates its own adhesion sites by damaging the mesothelial lining thus exposing the extracellular matrix. Therefore we conclude that the mesothelium has the properties of Teflon, while the extracellular matrix resembles Velcro. PMID- 11278201 TI - Is there any physiological role for gonadotrophin oligosaccharide heterogeneity in humans? I. Gondatrophins are synthesized and released in multiple molecular forms. A matter of fact. AB - Carbohydrates attached to the protein core of all glycoprotein hormones play an essential role in the function of the molecule, influencing a number of intracellular and extracellular processes. As with other members of the glycoprotein hormone family, pituitary gonadotrophins are not produced as single or unique molecules but rather as arrays of isoforms that differ from each other mainly in the structure of their oligosaccharide attachments. In both experimental animals and in humans, the abundance of the different isoforms varies depending on the endocrine status of the donor present at the time of collection of the tissue or sample. Conditions characterized by an oestrogen enriched hormonal milieu (eg. the preovulatory phase of the menstrual cycle) promote the formation and secretion of variants with relatively low sialic acid and/or sulphate content, whereas physiological deficiency of this sex steroid (as in the postmenopause) favours the production of highly sialylated, long-lived gonadotrophin variants. When tested individually, less sialylated isoforms exhibit higher receptor-binding and in-vitro biological activity but shorter plasma half-life than their more sialylated counterparts. Both the hormonal regulation and the functional differences among the naturally occurring isoforms strongly suggest that gonadotrophin heterogeneity represents a distinctly different mechanism through which the pituitary gland may regulate the intensity and duration of the gonadotrophic stimulus. Nevertheless, whereas the existence of the alternatively glycosylated variants of gonadotrophins in both the pituitary and in serum is currently without doubt, the physiological role of this phenomenon is still a controversial issue and a matter of debate. PMID- 11278204 TI - Gender reassignment and assisted reproduction: present and future reproductive options for transsexual people. AB - Transsexual people who want transition to their desired gender have to undergo hormonal and surgical treatments, which lead to irreversible loss of their reproductive potential. This paper argues that transsexual people should be offered the same options as any person that risks losing their germ cells because of treatment for a malignant disease. Indeed, transsexual women (male-to-female transsexual patients) may be given the option to store spermatozoa before they start hormonal therapy, so that their gametes may be used in future relationships. This may be especially important for the many transsexual women who identify as lesbians after their transition. Conversely, transsexual men (female-to-male transsexual patients) may be offered storage of oocytes or ovarian tissue, possibly obtained at the time of their oophorectomy. Current technology offers transsexual people the possibility to obtain children who are genetically their own in their future relationships and the option of gamete banking should therefore be discussed before starting hormonal and surgical reassignment treatment. This is particularly important for transsexual people who are diagnosed and treated at a young age. PMID- 11278203 TI - Embryo implantation and GnRH antagonists: ovarian actions of GnRH antagonists. AB - The gonadotrophin-releasing hormone (GnRH) antagonists, cetrorelix and ganirelix, have both been approved for ovarian stimulation to prevent a premature LH surge. Since GnRH receptors and their gene expression have been detected in human ovary, concern has risen over whether GnRH antagonists might affect ovarian function. Three large trials which compared GnRH agonists (used in the standard protocol worldwide), with the new antagonist treatment found no significant differences concerning the most important goals, e.g. pregnancy rate, fertilization and quality of oocytes. However, the concentration of oestradiol, and the pregnancy and implantation rates were lower in GnRH antagonist-treated patients. These findings again fuelled the debate about the possible extrapituitary effects of GnRH antagonists. Here, we review the conflicting data in the literature on the ovarian effects of GnRH antagonists and discuss our own results. In our view, it is unlikely that GnRH antagonists have a relevant impact on ovarian steroidogenesis and function; however, GnRH antagonists may exert other effects on the ovary. PMID- 11278205 TI - Avoiding multiple pregnancies: sailing uncharted seas. AB - Single embryo transfer is being proposed as the solution to avoid multiple pregnancies in IVF. Nevertheless, in my opinion, although this is the right solution, it is still not the correct one at the present time. Mainly, it is unfair to the majority of infertile couples and it will also severely limit the physician's capacity to resolve unfavourable IVF cases. Furthermore, current IVF technology is far from perfect and the impact of single embryo transfer needs to be evaluated in patients over 38 years of age, poor responders, and also in regard to blastocyst transfer and the development of pre-implantation genetic diagnosis. PMID- 11278206 TI - To blastocyst or not to blastocyst? That is the question. AB - Recent advances in culture media preparations have allowed for cleavage embryos to be developed to the blastocyst stage. Blastocysts are regarded as having increased implantation potential, and two blastocysts are typically transferred, which reduces the occurrence of high order multiple gestations. However, with current techniques, most cleavage embryos do not become blastocysts and it is not clear how many of these embryos would have implanted had they been replaced at the cleavage stage. Furthermore, experience with blastocyst cryopreservation is lacking and the overall benefit of blastocyst culture is unknown, unless we consider the combined pregnancy rates of both fresh and frozen blastocysts. PMID- 11278207 TI - Poor responder-high responder: the importance of soluble vascular endothelial growth factor receptor 1 in ovarian stimulation protocols. AB - This study was designed to detect vascular endothelial growth factor (VEGF) and its soluble receptor (sVEGFR-1) in follicular fluid specimens and to evaluate the importance of sVEGFR-1 with respect to ovarian response to gonadotrophin stimulation. A total of 69 patients was treated for IVF with recombinant human follicle stimulating hormone (FSH). Concentrations of VEGF and sVEGFR-1 were quantified in follicular fluids from oocyte retrievals. Patients were designated to three groups with respect to the number of harvested oocytes: group A, 1-5 oocytes; group B, 6-10 oocytes; group C, >10 oocytes. In group A, 1133 +/- 870 pg VEGF/ml follicular fluid per oocyte were quantified, in group B 426 +/- 262 pg VEGF/ml per oocyte, and in group C 274 +/- 179 pg VEGF/ml per oocyte. Soluble VEGFR-1 concentrations resulted in 1200 +/- 523 pg/ml follicular fluid per oocyte in group A, 255 +/- 193 pg/ml per oocyte in group B, and 79 +/- 69 pg/ml per oocyte in group C. No free sVEGFR-1 could be detected in any follicular fluid. An index to estimate the biological activity of VEGF by dividing VEGF/sVEGFR-1 revealed an increasing availability of VEGF with higher ovarian response to gonadotrophin therapy. In group A this index was 1.03, in group B 1.71, and in group C 3.21. A delicate balance between VEGF and sVEGFR-1 is necessary to allow an adequate ovarian reaction to gonadotrophin therapy. Excess of bio-active VEGF increases the risk for ovarian hyperstimulation syndrome. Excess of sVEGFR-1 results in poor response and goes in parallel with reduced chances for conception. PMID- 11278208 TI - Development of endometriosis-like lesions after transplantation of human endometrial fragments onto the chick embryo chorioallantoic membrane. AB - The chick embryo chorioallantoic membrane (CAM) bioassay was used to investigate the early pathogenesis of endometriosis. Endometrial fragments were explanted onto the CAM. The grafts including the surrounding CAM were excised at 24, 48 or 72 h after explantation, fixed and embedded in paraffin. Immunohistochemical analysis was used to distinguish endometrial cells. To identify cells of human origin, in-situ hybridization was performed using a probe specific for human chromosome 1. After 24 h, direct contact between endometrial stromal as well as epithelial cells and the mesenchymal layer of the CAM was observed. Invasion of both stromal cells and intact endometrial glands into the mesenchymal layer was observed after 48 h. At 72 h, endometriosis-like lesions were observed in the mesenchymal layer. Positive staining with antibodies to vimentin and pan cytokeratin was observed in the invading cells as well as in the lesions. In the lesions these positively stained cells showed in-situ hybridization signals for human chromosome 1, confirming their human origin. In conclusion, after 3 days of incubation, endometriosis-like lesions consisting of human endometrial glands and stromal cells were found in the mesenchymal layer of the CAM. These lesions apparently resulted from the invasion of intact human epithelial structures and stromal cells. PMID- 11278209 TI - Administration of cyclophosphamide at different stages of follicular maturation in mice: effects on reproductive performance and fetal malformations. AB - This study assessed reproductive performance, fetal viability and teratogenicity in female mice exposed to cyclophosphamide across a timeline corresponding to different stages of follicle maturation. Pregnancies were established in female Balb/c mice 1-4 weeks after administration of a non-sterilizing dose of cyclophosphamide (75 mg/kg). Each mating group represented a different stage of follicular growth at the time of cyclophosphamide exposure. The number of corpora lutea, pregnancies and fetal resorptions were determined. Surviving fetuses were evaluated for gross malformations. Results indicated that conceptions attributable to follicles exposed to cyclophosphamide at a mature stage had a significantly lower number of implantation sites, 4.82 +/- 1.01 versus 8.27 +/- 0.81 in controls (P = 0.001) and a high resorption rate, 56% +/- 0.11 versus 34% +/- 0.07 in controls (P = 0.05). The proportion of corpora lutea in this group which resulted in viable fetuses was extremely low, 0.2 +/- 0.06 versus 0.51 +/- 0.07 in controls (P = 0.001). Malformation rate was more than 10 times higher in all treated groups (P < 0.05) and a particularly high incidence of 33% (P = 0.0014) was observed in conceptions attributable to oocytes exposed to cyclophosphamide at the earliest stages of follicle growth. With an extended interval between exposure and mating the malformation rate gradually decreased towards normal values in the 12th week group. This study suggests that the effect of cyclophosphamide on female gametes and subsequently on future reproduction is influenced by the stage of oocyte maturation at the time of exposure. Early fertilization post-chemotherapy can result in a high rate of pregnancy failure and high malformation rate. This should be taken into account when considering the use of oocyte retrieval, IVF and embryo cryopreservation in patients currently undergoing chemotherapy. PMID- 11278210 TI - Chorion releases a factor that inhibits oxytocin-stimulated myometrial contractility in the pregnant guinea pig. AB - It was postulated that chorion releases a substance necessary for the maintenance of uterine quiescence during pregnancy. A decrease in the release of this substance at the end of the pregnancy would be necessary for normal myometrial activation. This hypothesis was tested by demonstrating the ability of chorion to inhibit oxytocin-stimulated myometrial contractility in vitro. Tissues were obtained from timed pregnant Duncan-Hartley guinea pigs either at pre-term or near-term gestation. Myometrial strips were placed in organ baths for isometric tension measurement and contractions stimulated by oxytocin (10(-8) mol/l). Fetal membranes or conditioned medium from chorion were added directly to the organ bath. Near-term chorion and chorion conditioned-medium decreased oxytocin stimulated contractile activity to 39% and 49% respectively. Neither pre-term nor near-term amnion reduced oxytocin-stimulated myometrial contractile activity. Relaxation induced by pre-term chorion was greater than near-term chorion (23% and 41% of the oxytocin-induced basal level respectively; P < 0.05). Further, chorion-induced relaxation was independent of the gestational age of the myometrium. Human chorion from a term, not-in-labour woman also inhibited oxytocin-stimulated guinea pig myometrial contractility. It was concluded that the chorion releases a substance or substances that reduce oxytocin-stimulated myometrial contractility and may be involved in the maintenance of uterine quiescence during pregnancy. PMID- 11278211 TI - Comparable clinical outcome using the GnRH antagonist ganirelix or a long protocol of the GnRH agonist triptorelin for the prevention of premature LH surges in women undergoing ovarian stimulation. AB - This multicentre, randomized study was performed to assess the efficacy and safety of 0.25 mg ganirelix (Orgalutran, Antagon) treatment, using triptorelin (Decapeptyl) in a long protocol as a reference treatment. In total, 236 subjects were randomized to treatment with ganirelix (0.25 mg, s.c.) and 119 to triptorelin (0.1 mg, s.c.) treatment (treatment ratio 2:1). Treatment with ganirelix started on day 6 of stimulation, whereas treatment with triptorelin started on menstrual cycle day 21 to 24 of the previous cycle (i.e. the midluteal phase). The ganirelix regimen was on average 17 days shorter (9 versus 26 days) compared to the triptorelin regimen. The median total dose of recombinant FSH (Puregon) used was 450 IU less (1350 versus 1800 IU) in the ganirelix protocol. The initial follicular growth was faster and, consequently, oestradiol concentrations were higher in the ganirelix group. On the day of human chorionic gonadotrophin (HCG), the mean number of follicles > or = 11 mm was 10.1 and 10.7 and the median serum oestradiol concentration was 1090 and 1370 pg/ml in the ganirelix and triptorelin groups respectively. Per attempt, 7.9 and 9.6 oocytes (mean) were retrieved in the ganirelix and triptorelin groups respectively. The fertilization rates (64.0% ganirelix and 64.9% triptorelin) and the mean number of good quality embryos (2.7 and 2.9) were comparable in both treatment groups. The implantation rate was identical (22.9%). The ongoing pregnancy rate per attempt was 31.0 and 33.9% in the ganirelix and triptorelin groups respectively. The ganirelix regimen showed an improved local tolerance in that the percentage of subjects with at least one local skin reaction was 2-fold lower than in the triptorelin group (11.9 versus 24.1%). Taking all data together, it may be concluded that ganirelix offers a new treatment regimen in ovarian stimulation that is short, safe and well-tolerated, optimizing convenience for the patient. PMID- 11278212 TI - Follicular development and hormone concentrations following recombinant FSH administration for anovulation associated with polycystic ovarian syndrome: prospective, randomized comparison between low-dose step-up and modified step down regimens. AB - The present study compared ovarian performance and hormone concentrations, after ovulation induction, in polycystic ovarian syndrome (PCOS) patients, using recombinant human FSH (rhFSH) in low-dose step-up and modified step-down regimens. Twenty-six women with clomiphene citrate-resistant chronic anovulatory infertility were treated with rhFSH in two consecutive cycles according to two different low-dose regimens: (i) the classic chronic low-dose step-up protocol, the starting dose being 75 IU; (ii) a modified step-down protocol where the starting dose was 300 IU followed by 3 days free of treatment, then rhFSH 75 IU daily was given and stepwise dose increments were performed exactly the same as in the step-up method. Each woman received both treatment approaches, in a randomized order, with an interval of > or = 1 month between treatments. The total number of follicles that were > 10, > 14 and > 17 mm in diameter on the day of human chorionic gonadotrophin (HCG) administration, and thus cycles with HCG cancelled, were significantly increased with the step-up approach. The total number of rhFSH ampoules tended to be higher with the step-down schedule despite the fact that both the mean duration of treatment and the threshold dose were similar with the two low-dose approaches. A physiological step-down approach for ovulation induction in PCOS patients may be more appropriate in order to avoid multifollicular cycles than the step-up approach. PMID- 11278213 TI - Leptin during assisted reproductive cycles: the effect of ovarian stimulation and of very early pregnancy. AB - Leptin may have a role in human reproduction. The impact of IVF and of very early pregnancy on serum leptin concentrations was studied in 66 infertile patients, of whom 19 became pregnant. Ovarian suppression was accompanied by a fall in leptin concentrations (21 +/- 4%, mean +/- SE; P < 0.01) from the mid-luteal phase, and ovarian stimulation by a rise (76 +/- 8%; P < 0.0001) from suppression. The mid luteal concentration of leptin after stimulation was 28 +/- 7% higher than that during the preceding normal cycle (P < 0.001). Concentrations of leptin and oestradiol were related before treatment, at ovarian suppression and at 8 days after oocyte retrieval. In addition, the rises in leptin and oestradiol concentrations during stimulation were correlated, but only in those patients who became pregnant (r = 0.69; P = 0.001). Women with a successful pregnancy had higher concentrations of leptin (18.7 +/- 4.8 microg/l) at 12 days after embryo transfer than those who had miscarriages (10.0 +/- 1.9 microg/l; P < 0.001), or those failing to become pregnant (11.6 +/- 1.2 microg/l; P < 0.0001). We concluded that leptin concentrations are influenced by ovarian function and that the relationship between leptin and oestrogen (but not a single leptin concentration), may be an important factor for the outcome of IVF. PMID- 11278214 TI - Comparison of LH concentrations in the early and mid-luteal phase in IVF cycles after treatment with HMG alone or in association with the GnRH antagonist Cetrorelix. AB - Luteinizing hormone (LH) is mandatory for the maintenance of the corpus luteum. Ovarian stimulation for IVF has been associated with a defective luteal phase. The luteal phases of two groups of patients with normal menstrual cycles and no endocrinological cause of infertility were retrospectively analysed in IVF cycles. Thirty-one infertile patients stimulated with human menopausal gonadotrophins (HMG) for IVF to whom the gonadotrophin-releasing hormone (GnRH) antagonist Cetrorelix 0.25 mg was also administered to prevent the LH surge (group I) were compared with 31 infertile patients stimulated with HMG alone (group II). Despite differences in the stimulation outcome, luteal LH serum concentrations were similar in the two groups. LH values dropped from 2.3 +/- 1 IU/l on the day of human chorionic gonadotrophin (HCG) administration to 1.1 +/- 0.7 IU/l on day HCG +2 in group I (P < 0.0001) and from 5.1 +/- 3 to 1.2 +/- 1.7 IU/l (P < 0.0001) in group II. In the mid-luteal phase, LH concentrations were low in both groups. Our results suggest that suppressed LH concentrations in the early and mid-luteal phase may not be attributed solely to the GnRH-antagonist administration. Pituitary LH secretion may be inhibited by supraphysiological steroid serum concentrations via long-loop feedback and/or by the central action of the exogenously administered HCG via a short-loop mechanism. PMID- 11278215 TI - Circulating follistatin concentrations are higher and activin concentrations are lower in polycystic ovarian syndrome. AB - Familial polycystic ovarian syndrome (PCOS) has been proposed to be linked to a site near the follistatin gene. We studied the concentrations of circulating follistatin, activin A and inhibin B in well-characterized subjects with PCOS (n = 108) and controls without PCOS (n = 20). Mean (+/- SEM) concentrations of follistatin were higher (P < 0.05) in PCOS (0.27 +/- 0.03 ng/ml) than controls (0.15 +/- 0.02 ng/ml) and activin A were lower (P < 0.05) in PCOS (0.20 +/- 0.01ng/ml) than controls (0.24 +/- 0.02 ng/ml). Inhibin B concentrations were not different between the two groups: PCOS (0.06 +/- 0.01ng/ml), and controls (0.06 +/- 0.01ng/ml). It is proposed that higher concentrations of follistatin with lower concentrations of activin A may relate to follicular development not proceeding beyond 8-10 mm and may be partly responsible for the lack of pre ovular follicle development in PCOS. PMID- 11278216 TI - Simultaneous evaluation of basal FSH and oestradiol response to GnRH analogue (F G-test) allows effective drug regimen selection for IVF. AB - To determine whether preliminary assessment of ovarian reserve by simultaneous evaluation of basal follicle-stimulating hormone (FSH) and oestradiol response to gonadotrophin releasing hormone (GnRH) analogue (F-G-test) can be used to tailor individually the drug regimen for ovarian stimulation, the in-vitro fertilization (IVF) results of 238 patients were retrospectively analysed. Sixty-two women with abnormal response to the test (DeltaE2 <180 pmol/l and/or FSH >9.5 mIU/ml) had commenced buserelin nasal spray in the mid-luteal phase and discontinued it on cycle day 1. Ovarian stimulation was started on cycle day 3 with 375 IU/day of gonadotrophin. Fifty-three patients completed the treatment cycle (group A). A total of 176 women with normal response to the test (DeltaE2 >180 pmol/l and FSH <9.5 mIU/ml) had continued the GnRH analogue throughout the stimulation cycle and a starting dose of 225 IU/day of gonadotrophin was used from cycle day 3. A total of 158 patients completed the treatment cycle (group B). Group A had significantly higher age (34.9 +/- 4.2 versus 33.2 +/- 4.2) (P < 0.05) and basal FSH (9.2 +/- 3.8 versus 7.0 +/- 2.2) (P < 0.05) and required a higher total dose of gonadotrophin. The numbers of oocytes retrieved and embryos transferred were significantly lower. However, fertilization, clinical pregnancies, and implantation rates were similar in both groups. It was concluded that simultaneous evaluation of basal FSH and oestradiol response to GnRH analogue can be useful in identifying subcategories of women with reduced ovarian reserve who may benefit from reduced GnRH analogue administration and a higher starting dose of gonadotrophin. PMID- 11278217 TI - Effects of human hydrosalpinx fluid on in-vitro murine fertilization. AB - Patients with hydrosalpinges show a decrease of both fertility and clinical outcome of IVF and embryo transfer treatment. Several reports have demonstrated the negative effects of hydrosalpinx fluid (HSF) on embryo development and implantation. The aim of this study was to determine whether human HSF, collected from infertile patients, might exhibit a deleterious effect on gametes and fertilization using a murine IVF system. Murine gametes were co-incubated during IVF until first cleavage with human HSF diluted to 50% from four patients (HSF1 4). It was demonstrated that HSF affected fertilization, as determined by the count of the 2-cell embryos. Pre-incubation of spermatozoa with HSF during capacitation significantly lowered the percentage of 2-cell embryos (P < 0.05). While HSF1-3 had no significant effect on motility and viability of spermatozoa, HSF4 almost completely affected their survival. In contrast, pre-incubation of ovulated oocytes surrounded by their cumulus cells with HSF before IVF did not impede first cleavage. Taken together, these results suggest that HSF has a cytotoxic effect on spermatozoa and/or impairs the fertilization process, probably by altering capacitation/acrosome reaction and/or ligand(s)-receptor(s) interactions. Hydrosalpinges may be partly associated with sterility through HSF inhibitory effects on fertilization. PMID- 11278218 TI - IVF following impossible or failed surgical reversal of tubal sterilization. AB - Microsurgical re-anastomosis or IVF offer ways of reversing previous tubal sterilization. This retrospective study analysed 56 attempts of IVF in 37 couples after impossible or failed surgical sterilization reversal. Efficacy of IVF in this group (TL) was compared with that of a tubal pathology control group (TP) at all stages of IVF (stimulation, fertilization and implantation). Depending on patient age, significantly fewer oocytes were produced after ovarian stimulation in the TL group than in the control (TP) group (P = 0.023 for all TL patients; P = 0.02 when patients aged >38 years were excluded). The total number of embryos available for transfer was significantly lower in the TL group (P = 0.0042), but this was age-related, since when women aged >38 years were excluded there was no significant difference between the two groups. The ongoing pregnancy rate was similar in both groups, the probability of ongoing pregnancy appearing to depend on patient age rather than on previous fertility. PMID- 11278219 TI - A postal survey of embryo transfer practice in the UK. AB - In this postal survey a questionnaire was sent to all unit directors in the UK to determine their attitudes to the factors influencing embryo transfer practice. They were requested to rate each step on a scale of 1-10, where 1 was irrelevant and 10 very important. A total of 80 practitioners from 40 units replied. Over 50% of the corresponding practitioners were consultants, 33% were middle-grade clinicians, and 12% were infertility nurse specialists. The factor that got the highest rating was the need for a standardized protocol for all unit staff regarding embryo transfer technique. The second critical factor voted by the respondents was the presence of blood on the embryo transfer catheter. Not touching the uterine fundus was deemed to be the third most important factor while the type of embryo transfer catheter used was a very close fourth. Prolonged bed rest following embryo transfer was voted the least important factor to influence the outcome. The wide variations in practice and choice of catheters encountered in this survey are indications of the divided opinion and lack of concrete evidence on which to base any firm decisions. The need for large clinical studies to assess clearly whether higher pregnancy rates will result from modifications in embryo transfer practice is highlighted. PMID- 11278221 TI - A randomized study comparing IVF in the unstimulated cycle with IVF following clomiphene citrate. AB - The efficiency of IVF in unstimulated cycles was compared with that following ovarian stimulation with clomiphene citrate in a simple protocol with ultrasound monitoring only. A total of 132 couples with no previous IVF attempts, selected by female age <35 years, indication for intracytoplasmic sperm injection or infertility caused by tubal factor or unexplained infertility were randomized to the two protocols. Randomization yielded two comparable groups. The clomiphene group (68 couples) performed significantly better than the unstimulated group (64 couples) in terms of number of cycles with oocyte harvest (90/111 or 81% versus 65/114 or 57%; chi(2) = 9.21, P < 0.002), embryo transfers per started cycle (59/111 or 53% versus 29/114 or 25%; chi(2) = 18.14, P < 0.0001), live intrauterine pregnancy rate per started cycle (20/111 or 18% versus 4/114 or 4%; chi(2) = 12.42, P < 0.0001), live intrauterine pregnancy rate per embryo transfer (20/59 or 34% versus 4/29 or 14%; chi(2) = 3.96, P = 0.047), but not in terms of implantation rate (22/85 or 26% versus 4/29 or 14%; chi(2) = 1.65). Only two twin pregnancies occurred. Modest side-effects were recorded following clomiphene. Accordingly, a simple clomiphene citrate protocol, but not IVF in unstimulated cycles, seems compatible with the concept of 'friendly IVF', yielding a fair pregnancy rate both per cycle started and per embryo transfer in selected patients. The results do not substantiate any important negative anti-oestrogenic effects of clomiphene. PMID- 11278220 TI - Comparison of triple serum screening and pregnancy outcome in oocyte donation versus IVF pregnancies. AB - The current study compared triple serum screening results and outcomes in 37 oocyte donation (OD) and 46 self oocyte IVF-conceived singletons of similarly aged women (28.8 +/- 4.4 years and 30.7 +/- 4.5 years respectively). Both groups were followed from their embryo transfer and throughout pregnancy. Although the daily pattern of first-trimester serum beta-human chorionic gonadotrophin (HCG) was similar in both groups, higher mid-gestation HCG serum concentrations were found, i.e. 1.38 and 1.32 multiples of the median (median MoM) for IVF and OD respectively, in comparison with 0.99 median MoM from the same reference laboratory. Only the OD group had significantly increased alpha fetoprotein (AFP) concentrations (1.45 median MoM) (P = 0.002) compared with the reference laboratory. A total of 11% of the IVF and 13% of the OD women were found to be screen positive. In neither group were chromosomal abnormalities detected and no fetal or neonatal deaths were recorded. Seven (15%) of the OD and seven (19%) of the IVF women had an adverse obstetric outcome. Of those cases, six IVF and four OD women had serum HCG > or = 1.2 MoM and five OD women had AFP >1.2 MoM. Therefore, in those pregnancies the high serum HCG concentrations may alert for adverse obstetric outcome rather than indicating a high risk for Down's syndrome fetuses. PMID- 11278222 TI - High frequency of XY disomy in spermatozoa of severe oligozoospermic men. AB - Frequencies of disomy and diploidy in spermatozoa for chromosomes X, Y and 18 were compared among severe oligozoospermic men (<5x10(6) spermatozoa/ml), oligozoospermic men (5-20x10(6) spermatozoa/ml), and normospermic men using three colour fluorescence in-situ hybridization (FISH). Semen samples were collected from 10 severe oligozoospermic men aged 26-49 years, 10 oligozoospermic men aged 27-48 years and seven normospermic men aged 25-31 years. Karyotypes in lymphocytes obtained from peripheral blood were all 46,XY. In severe oligozoospermic men, analysis of 200 interphases per individual using FISH showed XY constitutions for sex chromosomes in all cells. A minimum of 10 000 sperm nuclei per individual for each chromosome was evaluated in severe oligozoospermic men and oligozoospermic men, and a minimum of 6000 sperm nuclei per individual in normospermic men. In total, 245 707 sperm nuclei were evaluated. The hybridization efficiency was 99.8%. The severe oligozoospermic men showed significantly higher frequencies of XY disomy (0.41%) and diploidy (0.49%) compared with oligozoospermic men (0.16%, P < 0.01; 0.22%, P < 0.05) and normospermic men (0.18%, P < 0.05; 0.21%, P < 0.05) (Mann-Whitney U-test). The data suggest that when severe oligozoospermic men undergo intracytoplasmic sperm injection, there can be an increase in the rate of conceptuses with 47,XXY chromosomes. PMID- 11278223 TI - Round spermatids from infertile men exhibit decreased protamine-1 and -2 mRNA. AB - During spermiogenesis, histone-to-protamine exchange causes chromatin condensation. Spermatozoa from infertile men are known to exhibit an increased protamine-1 (PRM1) to protamine-2 (PRM2) protein ratio. Since patients undergoing testicular sperm extraction (TESE) followed by intracytoplasmic sperm injection (ICSI) reveal low fertilization rates, whether the outcome of ICSI could be related to the percentage of round spermatids expressing PRM1-mRNA and PRM2-mRNA was investigated. Applying in-situ hybridization, 55 testicular biopsies from men undergoing TESE/ICSI were investigated. The percentage of PRM1-mRNA and PRM2-mRNA positive spermatids was significantly (P < 0.0001) decreased in men with at least qualitatively normal spermatogenesis (PRM1-mRNA: 58.4 +/- 13.8%; PRM2-mRNA: 56.4 +/- 11.3%) and impaired spermatogenesis (PRM1-mRNA: 32.6 +/- 10.8%; PRM2-mRNA: 31.7 +/- 11.1%) compared with men with obstructive azoospermia and quantitatively normal spermatogenesis (PRM1-mRNA: 79.9 +/- 4.6%; PRM2-mRNA: 78.1 +/- 5.7%). A positive correlation (r(PRM1) = 0.733; r(PRM2) = 0.784; P < 0.001) was demonstrated between the score and the percentage of PRM1-mRNA and PRM2-mRNA positive spermatids. While successful fertilization was neither related to the score, nor to the percentage of PRM1-mRNA and PRM2-mRNA positive spermatids, a significant (P < 0.05) relationship was demonstrated between successful fertilization and the PRM1-mRNA to PRM2-mRNA ratio. Therefore, the PRM1-mRNA to PRM2-mRNA ratio in round spermatids may serve as a possible predictive factor for the outcome of ICSI. PMID- 11278224 TI - XX males without SRY gene and with infertility. AB - The case of a 28 year old male with normal male phenotype, in whom repeated seminal analysis showed complete azoospermia, is presented. Peripheral blood culture for chromosome studies revealed 46 chromosomes with XX constitution. Polymerase chain reaction (PCR) analysis of genomic DNA failed to detect the presence of the sex-determining region of the Y chromosome (SRY). A literature review of all SRY-negative XX males with normal male phenotype showed that this case is the sixth reported case but the first to be diagnosed during the investigations of infertility. The frequency, aetiology and diagnosis of this rare syndrome are also reviewed. PMID- 11278225 TI - A microscopic and biochemical study of fragmentation phenotypes in stage appropriate human embryos. AB - The occurrence of a pleiomorphic population of cytoplasmic fragments is a common characteristic of early human embryos fertilized in vitro. Here, temporal, spatial, fine structural, and biochemical aspects of fragmentation were examined in fragmented monospermic and dispermic pronuclear to early cleavage stages human embryos classified as stage-appropriate during the first 3.5 days of culture. The morphodynamics of certain common patterns of fragmentation and the movement and composition of fragments were analysed by time-lapse video, mitochondrial fluorescent probes, and transmission electron microscopy. Plasma membrane and nuclear DNA integrity were assessed by annexin V staining, and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) and single cell alkaline gel electrophoresis ('comet') assays respectively. Developmental competence for affected embryos was related to outcome after embryo transfer. The results demonstrate that certain common forms of spontaneous fragmentation affecting early human embryos are not lethal, and that clusters of apparent fragments are often transient structures, which disappear by resorption or lysis. The findings suggest that the occurrence and fate of fragments characteristic of these phenotypes may be related to oncosis-like processes associated with transient and focal ATP deficiencies in blastomeres and mitochondrial deficiencies or absence in extracellular fragments. PMID- 11278226 TI - Preliminary findings in germinal vesicle transplantation of immature human oocytes. AB - Transplanting a germinal vesicle (GV) from an aged woman's oocyte into a younger ooplasm has been proposed as a possible way to reduce the incidence of oocyte aneuploidy which is considered to be responsible for age-related infertility. In this study, we have assessed the efficiency of each step involved in nuclear transplantation-specifically cell survival, nuclear-cytoplasmic reconstitution, and the capacity of the reconstituted oocytes for in-vitro maturation. In addition, we have evaluated the fertilizability and karyotypic status of the manipulated oocytes by intracytoplasmic sperm injection (ICSI) and fluorescent in situ hybridization technique respectively. Nuclear transplantation was accomplished with an overall efficiency of 73%. Due to the limited availability of materials, most nuclear transplantation procedures were performed between sibling oocytes. The maturation rate of 62% following reconstitution was comparable with that of control oocytes, as was the incidence of aneuploidy among the reconstituted oocytes. The ICSI results of the reconstituted oocytes yielded a survival rate of 77%, a fertilization rate of 52%, and a satisfactory early embryonic cleavage. Furthermore, in a limited number of observations where the nucleus of an aged oocyte was transferred into a younger ooplasm, there was an appropriate chromosomal segregation. These findings demonstrate that human oocytes reconstituted with GV nuclei are able to undergo maturation, fertilization, and early embryo cleavage, and maintain a normal ploidy. Although in-vitro maturation seems to be a limiting step, this technique would allow us to investigate further the nuclear-ooplasmic relationship during meiotic maturation. PMID- 11278227 TI - Effects of low O2 and ageing on spindles and chromosomes in mouse oocytes from pre-antral follicle culture. AB - To assess their quality, spindles were analysed in mouse oocytes from pre-antral follicle culture. High or low oxygen tension was present during the last 16 or 20 h post human chorionic gonadotrophin (HCG)/epidermal growth factor (EGF) addition. Most oocytes from pre-antral follicle culture possessed typical anastral spindles with flat poles resembling those of ovulated, in-vivo-matured oocytes of sexually mature mice, while denuded oocytes in-vitro matured to metaphase II (MII) formed significantly longer, slender spindles with pointed, narrow poles. Spindles in oocytes from follicle culture were only slightly shorter and less compact at the equator as compared with those of oocytes matured in vivo. Chromosomes were well aligned at the equator in MII oocytes obtained from follicle culture with high oxygen. Maturation rate was significantly reduced by lowering oxygen tension to 5% O2. Prolonged culture and the presence of only 5% O2 dramatically increased the percentage of MII oocytes with unaligned chromosomes. These observations indicate that sufficient oxygen supply and time of retrieval after initiation of resumption of maturation by HCG as well as the microenvironment and cell-cell interactions between oocytes and their somatic compartment are critical in affecting the oocyte's capacity to mature to MII, to form a functional spindle, and to align chromosomes correctly. PMID- 11278228 TI - Amino acids promote human blastocyst development in vitro. AB - Supplementation of culture media with amino acids has been shown to benefit preimplantation embryo development in several species. This randomized study analysed the in-vitro development of human embryos obtained after IVF in the presence or absence of a combination of amino acids from the 2- to 4-cell stage to the blastocyst stage. A total of 129 human embryos was randomly distributed between three serum-free chemically defined sequential media: (i) glucose-free Earle's balanced salt solution (EBSS) with glutamine (Gln) prior to morula stage, supplemented with glucose for blastocyst formation; (ii) glucose-free EBSS with glutamine and non-essential amino acids (AA) for cleavage stage development, and supplemented with all 20 AA for blastocyst formation (Earle's+AA); and (iii) a sequential commercial medium containing amino acids (K-SCIM). Embryos were individually cultured for successive periods of 24 h. On day 6 of development, blastocysts were differentially labelled and the numbers of trophectoderm and inner cell mass cells, mitoses and dead cells were examined. Blastocyst development was similar for the three sequential media. The mixture of AA significantly increased total blastocyst cell numbers from 61.8 +/- 4.2 with Earle's+Gln to 99.3 +/- 8.4 with Earle's+AA and 100.2 +/- 9.4 with K-SCIM (P = 0.005). This increase was present in both the trophectoderm and inner cell mass lineages (P < 0.02). Furthermore, the dead cell index was significantly lower with Earle's+AA (P = 0.047). PMID- 11278229 TI - Differential effects of repeated ovarian stimulation on cytoplasmic and spindle organization in metaphase II mouse oocytes matured in vivo and in vitro. AB - The effects of four rounds of ovarian stimulation spaced 1-6 weeks apart on the normality of metaphase II (MII) spindle formation, chromosomal alignment and cytoplasmic organization were examined in intact ovulated mouse oocytes and at MII for oocytes obtained at the germinal vesicle stage from the same ovaries and matured in vitro. The terminal deoxynucleotidyl transferase-mediated dUDP nick end labelling assay was used to identify DNA strand breaks in chromosomes, and histological studies of ovaries between and at each round of ovarian stimulation were performed. The results demonstrate a progressive and significant increase in the frequency of spindle defects with each round of ovarian stimulation, including those spaced weeks apart. Oocytes with spindle defects were also characterized by the occurrence of detached chromosomes and cytoplasmic asters. In contrast, in-vitro matured oocytes derived from the same ovaries were normal. No evidence of DNA strand breaks with repeated rounds of ovarian stimulation was detected in ovulated or in-vitro matured oocytes. The development and persistence of nodules of hypertrophied granulosa in regions where follicular growth occurs suggest that a progressively increasing proportion of oocytes in the ovulatory pathway may experience an intrafollicular milieu that has negative consequences for competence. The results are discussed with respect to ovarian and oocyte biological ageing and possible adverse implications for human oocyte competence with repeated hyperstimulation. PMID- 11278230 TI - Polycystic ovaries and eating disorders: Are they related? AB - A cross-sectional observational study was used to investigate the reported association between polycystic ovarian syndrome and bulimia nervosa in a group of young, post-menarcheal women in the normal population. Volunteers aged 18-25 years were recruited from two universities and two general practice surgeries in Oxford. A total of 230 women completed an interviewer-based eating disorder examination, which was used to diagnose bulimia nervosa and its variants, and to assess eating behaviour. Transabdominal ultrasound was used to diagnose the presence or absence of polycystic ovaries. Symptoms of polycystic ovarian syndrome were assessed using menstrual history, anthropometric measurements, clinical observation of acne and hirsutism, and biochemical analysis of a fasting blood sample. A total of 30% of all participants described episodes of overeating and 4% had used extreme methods of weight control. Two women were diagnosed with bulimia nervosa, and five women with binge-eating disorder; however, these diagnoses were not associated with polycystic ovaries. Scores for dieting and overall eating disorder symptoms in the polycystic ovary groups were not significantly higher than those for women with normal ovaries, and therefore the suggestion that polycystic ovaries predispose towards the development of eating disorders is not supported by this study. PMID- 11278231 TI - Long-term results of laparoscopic myomectomy: recurrence rate in comparison with abdominal myomectomy. AB - Laparoscopic myomectomy is still a debated procedure and there are conflicting opinions regarding the recurrence rate. Laparoscopic myomectomy may present a higher risk of recurrence compared with abdominal myomectomy. The aim of this investigation was to analyse the recurrence rate of myomas after surgery. From January 1991 to June 1998, 165 myomectomies were performed for symptomatic myomas measuring at least 3 cm in diameter and numbering seven or less per patient. During the first 3 years of this survey, 81 patients were randomized for abdominal or laparoscopic myomectomy. Transvaginal ultrasound examination was performed within 15-30 days of surgery and every 6 months for a post-operative period of 40 months. The two groups had similar pre-operative clinical features and the number and volume of myomas did not differ between the two groups. At the end of the study the group of abdominal myomectomies showed nine recurrences (23%) against 11 (27%) of the laparoscopic group. In order to evaluate the recurrence rate in relation to several risk factors, laparoscopic myomectomies were performed from 1991 in 84 patients who agreed to follow-up (and were not in the randomized group). Of these, 78 patients were evaluated with transvaginal ultrasound for a mean interval of 26 months and 17 (21.78%) recurrences were found. Most recurrences (75%) were seen at ultrasound between 10 and 30 months after surgery. The patient's age, pre- and post-operative gravidity and parity had no influence on recurrence. Neither the number of myomas removed nor the depth of penetration or size were positively associated with the risk of recurrence. However, an associated risk factor was pre-operative gonadotrophin releasing hormone agonist treatment (P < 0.02). None of the women with recurrence required additional surgery. We conclude that laparoscopic myomectomy is a reliable procedure. The recurrence rate is similar to that seen after abdominal myomectomy. PMID- 11278233 TI - Tubal curettage: a new conservative treatment for haemorrhagic interstitial pregnancies. AB - Haemorrhagic interstitial pregnancies are commonly treated by cornual resection. This invasive procedure may increase the risk of uterine rupture in subsequent pregnancies. We report here a case of a haemorrhagic interstitial pregnancy, associated with a viable intrauterine pregnancy in a salpingectomized woman, which was treated successfully by curettage of the uterine cornu. PMID- 11278232 TI - Measurement of serum CA-125 concentrations does not improve the value of Chlamydia trachomatis antibody in predicting tubal pathology at laparoscopy. AB - Chlamydia antibody testing (CAT) has been used to predict tubal pathology associated with Chlamydia infection, the leading cause of pelvic inflammatory disease (PID). Tubal pathology not related to C. trachomatis is unlikely to be identified by CAT alone. A correlation between serum CA-125 concentrations and the severity of adnexal inflammation during acute PID was demonstrated. The objectives of this study were to determine the prevalence of C. trachomatis infection in an Asian infertile population and to assess the role of a combination of serum CA-125 and CAT in the prediction of tubal pathology as shown by laparoscopy. A total of 110 consecutive women attending an infertility clinic for work-up were recruited. Blood was taken for CAT and CA-125 on the day of hospital admission and an endocervical swab was taken for culture of C. trachomatis prior to laparoscopy. Two (1.8%) women had C. trachomatis found in the endocervix and 28 (25.5%) women had CAT of > or = 1:32. Serum CA-125 concentrations were > 35 IU/ml in 11 (10%) women. The discriminative capacity of CAT in the diagnosis of tubal pathology including both proximal and distal obstruction was not improved by measuring serum CA-125, regardless of the threshold values of serum CA-125 concentration. PMID- 11278234 TI - Three blastocyst stage embryo transfer resulting in a quintuplet pregnancy. AB - High-order pregnancies are associated with high morbidity and mortality and the incidence is increased as a drastic complication of assisted reproductive technology. This case presents a high-order pregnancy achieved by transfer of three blastocyst stage embryos resulting in a quintuplet pregnancy including a monochorionic triplet. Following the selective termination of the monochorionic triplet, two healthy children were born. The mechanism of monochorionic development and its association with assisted reproductive technology are discussed. PMID- 11278235 TI - The Human Rights Act (1998) and its impact on reproductive issues. AB - The Human Rights Act (HR Act) 1998 (UK) (Human Rights Act, 1998) came into effect on October 2, 2000. Instead of taking a case to the European Court of Human Rights in Strasbourg, litigants can enforce their rights in the UK. The Act will have an unprecedented effect in virtually all areas of the UK legal systems. In line with those countries who have incorporated the 'Convention' in domestic law, litigation is expected to increase. The extensive body of Convention law, as well as decisions of the domestic courts of other states which have incorporated the Convention, now becomes an integral part of UK jurisprudence. Broadly, the Act applies to public and not private bodies. The relevant bodies which embody reproductive issues and concerns are for example the National Health Service (NHS) and the regulatory bodies such as the Human Fertilisation and Embryology Authority (HFEA) (Human Fertilisation and Embryology Authority Act, 1990) and the Human Genetics Advisory Commission (HGAC). A profound impact on the NHS practice, interpretations of the HFEA Act and its Code of Practice can be envisaged in relation to the Convention rights. Cases involving reproductive issues are already emerging in relation to the HR Act and which include sex selection, the present embryo transfer policy, interpretation of fatherless offspring and the provision of fertility services under the NHS. This review is intended to raise awareness of the HR Act 1998 for persons interested in human reproductive issues and how the HR Act could impact on the current laws and practice. Whilst it is only possible to speculate on what might happen in relation to the HR Act, what is certain is that UK law will radically change to accommodate the requirements of the HR Act 1998. PMID- 11278236 TI - Prevention of twin pregnancies after IVF/ICSI by single embryo transfer. ESHRE Campus Course Report. AB - Twin pregnancies constitute the most serious complication for both mother and children after IVF/ICSI treatment, but transfer of at least two 'best looking' embryos remains the standard policy. This is due to our inability and reluctance to identify both the 'twin prone' patient and the top quality embryo. Some centres now electively transfer a single embryo (eSET) when particular embryo quality and patient criteria are met. Results from several centres were presented during an ESHRE Campus Course, held on May 6(th) 2000. Sound clinical trials are needed to clarify several points of discussion. What is the clinical profile of patients in whom eSET should be considered? Will the overall (ongoing) pregnancy rate of the IVF/ICSI programme decrease if eSET is performed in these patients? What is the twinning rate when eSET is a routine policy? Will the financial gain by avoiding perinatal hospitalization costs of prevented twins be balanced by the likely need to perform a number of extra IVF/ICSI cycles? What will be gained by freezing the extra number of high quality embryos? Should eSET be performed at the 2 pronuclear stage, the early cleaving embryo or the blastocyst stage? Common sense dictates that eSET as a concept should be applied from now onwards. PMID- 11278237 TI - Should ICSI be the treatment for all cases of in-vitro conception? PMID- 11278238 TI - LH and ovulation induction. PMID- 11278239 TI - Natural variation in the human sex ratio. PMID- 11278241 TI - Cell adhesion regulates the interaction between Nck and p21-activated kinase. AB - The p21-activated kinases (PAKs) are important mediators of cytoskeletal reorganization, cell motility and transcriptional events regulated by the Rho family GTPases Rac and Cdc42. PAK activation by serum components is strongly dependent on cell adhesion to the extracellular matrix (ECM). PAK binds directly to the Nck adapter protein, an interaction thought to play an important role in regulation and localization of PAK activity. This report demonstrates that the interaction of PAK with Nck is regulated dynamically by cell adhesion. PAK-Nck binding is rapidly lost after cell detachment and rapidly restored after re adhesion to the ECM protein fibronectin, suggesting a rapidly reversible mode of regulation. Furthermore, the loss of Nck binding correlates with changes in the phosphorylation state of PAK in nonadherent cells, as evidenced by electrophoretic mobility shift and phosphorylation within a sequence known to mediate interaction with Nck. The ability of cell adhesion to regulate PAK phosphorylation and interaction with Nck may contribute to the anchorage dependence of PAK activation as well as to the localization of activated PAK within a cell. PMID- 11278242 TI - Leucine zipper-mediated homodimerization of the p21-activated kinase-interacting factor, beta Pix. Implication for a role in cytoskeletal reorganization. AB - Pix, a p21-activated kinase-interacting exchange factor, is known to be involved in the regulation of Cdc42/Rac GTPases. The 85-kDa betaPix-a protein contains an Src homology 3 domain, the tandem Dbl homology and Pleckstrin homology domains, a proline-rich region, and a GIT1-binding domain. In addition to those domains, betaPix-a also contains a putative leucine zipper domain at the C-terminal end. In this study, we demonstrate that the previously identified putative leucine zipper domain mediates the formation of betaPix-a homodimers. Using in vitro and in vivo methodologies, we show that deletion of the leucine zipper domain is sufficient to abolish betaPix-a homodimerization. In NIH3T3 fibroblast cells, expression of wild type betaPix-a induces the formation of membrane ruffles. However, cells expressing the leucine zipper domain deletion mutant could not form membrane ruffle structures. Moreover, platelet-derived growth factor mediated cytoskeletal changes were completely blocked by the leucine zipper domain deletion mutant. The results suggest that the leucine zipper domain enables betaPix-a to homodimerize, and homodimerization is essential for betaPix a signaling functions leading to the cytoskeletal reorganization. PMID- 11278244 TI - Domain-specific mutations of a transforming growth factor (TGF)-beta 1 latency associated peptide cause Camurati-Engelmann disease because of the formation of a constitutively active form of TGF-beta 1. AB - Transforming growth factor (TGF)-beta1 is secreted as a latent form, which consists of its mature form and a latency-associated peptide (beta1-LAP) in either the presence or the absence of additional latent TGF-beta1-binding protein. We recently reported that three different missense mutations (R218H, R218C, and C225R) of beta1-LAP cause the Camurati-Engelmann disease (CED), an autosomal dominant disorder characterized by hyperosteosis and sclerosis of the diaphysis of the long bones. Pulse-chase experiments using fibroblasts from CED patients and expression experiments of the mutant genes in an insect cell system suggest that these mutations disrupt the association of beta1-LAP and TGF-beta1 and the subsequent release of the mature TGF-beta1. Furthermore, the cell growth of fibroblasts from a CED patient and mutant gene-transfected fibroblasts was suppressed via TGF-beta1. The growth suppression observed was attenuated by neutralizing antibody to TGF-beta1 or by treatment of dexamethasone. On the other hand, the proliferation of human osteoblastic MG-63 cells was accelerated by coculture with CED fibroblasts. These data suggest that the domain-specific mutations of beta1-LAP result in a more facile activation of TGF-beta1, thus causing CED. PMID- 11278243 TI - TIMP-3 is a potent inhibitor of aggrecanase 1 (ADAM-TS4) and aggrecanase 2 (ADAM TS5). AB - The proteoglycan aggrecan is an important major component of cartilage matrix that gives articular cartilage the ability to withstand compression. Increased breakdown of aggrecan is associated with the development of arthritis and is considered to be catalyzed by aggrecanases, members of the ADAM-TS family of metalloproteinases. Four endogenous tissue inhibitors of metalloproteinases (TIMPs) regulate the activities of functional matrix metalloproteinases (MMPs), enzymes that degrade most components of connective tissue, but no endogenous factors responsible for the regulation of aggrecanases have been found. We show here that the N-terminal inhibitory domain of TIMP-3, a member of the TIMP family that has functional properties distinct from other TIMPs, is a strong inhibitor of human aggrecanases 1 and 2, with K(i) values in the subnanomolar range. This truncated inhibitor, which lacks the C-terminal domain that is responsible for interactions with molecules other than active metalloproteinases, is produced at high yield by bacterial expression and folding from inclusion bodies. This provides a starting point for developing a biologically available aggrecanase inhibitor suitable for the treatment of arthritis. PMID- 11278245 TI - Bcl-B, a novel Bcl-2 family member that differentially binds and regulates Bax and Bak. AB - A novel human member of the Bcl-2 family was identified, Bcl-B, which is closest in amino acid sequence homology to the Boo (Diva) protein. The Bcl-B protein contains four Bcl-2 homology (BH) domains (BH1, BH2, BH3, BH4) and a predicted carboxyl-terminal transmembrane (TM) domain. The BCL-B mRNA is widely expressed in adult human tissues. The Bcl-B protein binds Bcl-2, Bcl-X(L), and Bax but not Bak. In transient transfection assays, Bcl-B suppresses apoptosis induced by Bax but not Bak. Deletion of the TM domain of Bcl-B impairs its association with intracellular organelles and diminishes its anti-apoptotic function. Bcl-B thus displays a unique pattern of selectivity for binding and regulating the function of other members of the Bcl-2 family. PMID- 11278247 TI - The RING heterodimer BRCA1-BARD1 is a ubiquitin ligase inactivated by a breast cancer-derived mutation. AB - BRCA1-BARD1 constitutes a heterodimeric RING finger complex associated through its N-terminal regions. Here we demonstrate that the BRCA1-BARD1 heterodimeric RING finger complex contains significant ubiquitin ligase activity that can be disrupted by a breast cancer-derived RING finger mutation in BRCA1. Whereas individually BRCA1 and BARD1 have very low ubiquitin ligase activities in vitro, BRCA1 combined with BARD1 exhibits dramatically higher activity. Bacterially purified RING finger domains comprising residues 1-304 of BRCA1 and residues 25 189 of BARD1 are capable of polymerizing ubiquitin. The steady-state level of transfected BRCA1 in vivo was increased by co-transfection of BARD1, and reciprocally that of transfected BARD1 was increased by BRCA1 in a dose-dependent manner. The breast cancer-derived BARD1-interaction-deficient mutant, BRCA1(C61G), does not exhibit ubiquitin ligase activity in vitro. These results suggest that the BRCA1-BARD1 complex contains a ubiquitin ligase activity that is important in prevention of breast and ovarian cancer development. PMID- 11278246 TI - Akt participation in the Wnt signaling pathway through Dishevelled. AB - Inactivation of glycogen synthase kinase 3beta (GSK3beta) and the resulting stabilization of free beta-catenin are critical steps in the activation of Wnt target genes. While Akt regulates GSK3alpha/beta in the phosphatidylinositide 3 OH kinase signaling pathway, its role in Wnt signaling is unknown. Here we report that expression of Wnt or Dishevelled (Dvl) increased Akt activity. Activated Akt bound to the Axin-GSK3beta complex in the presence of Dvl, phosphorylated GSK3beta and increased free beta-catenin levels. Furthermore, in Wnt overexpressing PC12 cells, dominant-negative Akt decreased free beta-catenin and derepressed nerve growth factor-induced differentiation. Therefore, Akt acts in association with Dvl as an important regulator of the Wnt signaling pathway. PMID- 11278248 TI - Gene correction of the apolipoprotein (Apo) E2 phenotype to wild-type ApoE3 by in situ chimeraplasty. AB - Apolipoprotein (apo) E is a polymorphic plasma protein, synthesized mainly by liver. Here, we evaluate whether synthetic DNA-RNA oligonucleotides (chimeraplasts) can convert a dysfunctional isoform, apoE2 (C --> T, R158C), which causes Type III hyperlipidemia and premature atherosclerosis, into apoE3. First, we treated recombinant Chinese hamster ovary cells stably secreting apoE2 with a 68-mer apoE2 to apoE3 chimeraplast. About one-third of apoE2 was converted to apoE3, and the repair was stable through 12 passages. Subcloning treated cells produced both apoE2 and apoE3 clones. Direct sequencing and reverse transcription polymerase chain reaction confirmed the genotype, whereas phenotypic change was verified by isoelectric focusing and immunoblotting of secreted proteins. Second, we established that the APOE2 gene can be targeted both in vivo, using transgenic mice overexpressing human apoE2, and in chromosomal context, using cultured lymphocytes from a patient homozygous for the epsilon2 allele. We conclude that chimeraplasty has the potential to convert the apoE2 mutation in patients with Type III hyperlipidemia to apoE3. PMID- 11278251 TI - Smurf1 interacts with transforming growth factor-beta type I receptor through Smad7 and induces receptor degradation. AB - Smad7 is an inhibitory Smad that acts as a negative regulator of signaling by the transforming growth factor-beta (TGF-beta) superfamily proteins. Smad7 is induced by TGF-beta, stably interacts with activated TGF-beta type I receptor (TbetaR-I), and interferes with the phosphorylation of receptor-regulated Smads. Here we show that Smurf1, an E3 ubiquitin ligase for bone morphogenetic protein-specific Smads, also interacts with Smad7 and induces Smad7 ubiquitination and translocation into the cytoplasm. In addition, Smurf1 associates with TbetaR-I via Smad7, with subsequent enhancement of turnover of TbetaR-I and Smad7. These results thus reveal a novel function of Smad7, i.e. induction of degradation of TbetaR-I through recruitment of an E3 ligase to the receptor. PMID- 11278252 TI - Gene expression profile of antithrombotic protein c defines new mechanisms modulating inflammation and apoptosis. AB - Human protein C is a natural anticoagulant factor, and a recombinant activated form of the molecule (rhAPC) is completing clinical evaluation for treatment of severe sepsis. Because of the pathophysiologic role of endothelial dysfunction in severe inflammatory disease and sepsis, we explored the possibility that rhAPC might directly modulate endothelial function, independent of its anticoagulant activity. Using broad transcriptional profiling, we show that rhAPC directly modulates patterns of endothelial cell gene expression clustering into anti inflammatory and cell survival pathways. rhAPC directly suppressed expression of p50 and p52 NFkappaB subunits, resulting in a functional decrease in NFkappaB binding at target sites. Further, rhAPC blocked expression of downstream NFkappaB regulated genes following tumor necrosis factor alpha induction, including dose dependent suppression of cell adhesion expression and functional binding of intracellular adhesion molecule 1, vascular cell adhesion molecule 1, and E selectin. Further, rhAPC modulated several genes in the endothelial apoptosis pathway, including the Bcl-2 homologue protein and inhibitor of apoptosis protein. These pathway changes resulted in the ability of rhAPC to inhibit the induction of apoptosis by the potent inducer, staurosporine. This new mechanistic understanding of endothelial regulation and the modulation of tumor necrosis factor-induced endothelial dysfunction creates a novel link between coagulation, inflammation, and cell death and provides insight into the molecular basis for the efficacy of APC in systemic inflammation and sepsis. PMID- 11278253 TI - Direct transcriptional activation of human caspase-1 by tumor suppressor p53. AB - The tumor suppressor protein p53 is a sequence-specific DNA-binding protein, and its biological responses are very often mediated by transcriptional activation of various target genes. Here we show that caspase-1 (interleukin-1beta converting enzyme), which plays a role in the production of proinflammatory cytokines and in apoptosis, is a transcriptional target of p53. Caspase-1 mRNA levels increased upon overexpression of p53 by transfection in MCF-7 cells. Human caspase-1 promoter showed a sequence homologous to the consensus p53-binding site. This sequence bound to p53 in gel shift assays. A caspase-1 promoter-reporter construct was activated 6-8-fold by cotransfection with normal p53 but not by mutant p53 (His(273)) in HeLa, as well as MCF-7, cells. Mutation of the p53 binding site in caspase-1 promoter abolished transactivation by p53. Treatment of p53-positive MCF-7 cells with the DNA-damaging drug, doxorubicin, which increases p53 levels, enhanced caspase-1 promoter activity 4-5-fold, but similar treatment of MCF-7-mp53 (a clone of MCF-7 cells expressing mutant p53) and p53-negative HeLa cells with doxorubicin did not increase caspase-1 promoter activity. Doxorubicin treatment increased caspase-1 mRNA levels in MCF-7 cells but not in MCF-7-mp53 or HeLa cells. These results show that endogenous p53 can regulate caspase-1 gene expression. PMID- 11278254 TI - A cysteine-rich adipose tissue-specific secretory factor inhibits adipocyte differentiation. AB - A 12.5-kDa cysteine-rich adipose tissue-specific secretory factor (ADSF/resistin) is a novel secreted protein rich in serine and cysteine residues with a unique cysteine repeat motif of CX(12)CX(8)CXCX(3)CX(10)CXCXCX(9)CC. A single 0.8 kilobase mRNA coding for this protein was found in various murine white adipose tissues including inguinal and epididymal fats and also in brown adipose tissue but not in any other tissues examined. Two species of mRNAs with sizes of 1.4 and 0.8 kilobases were found in rat adipose tissue. Sequence analysis indicates that this is because of two polyadenylation signals, the proximal one with the sequence AATACA with a single base mismatch from murine AATAAA and the distal consensus sequence AATAAA. The mRNA level was markedly increased during 3T3-L1 and primary preadipocyte differentiation into adipocytes. Its expression in adipose tissue is under tight nutritional and hormonal regulation; the mRNA level was very low during fasting and increased 25-fold when fasted mice were refed a high carbohydrate diet. It was also very low in adipose tissue of streptozotocin diabetes and increased 23-fold upon insulin administration. Upon treatment with the conditioned medium from COS cells transfected with the expression vector, conversion of 3T3-L1 cells to adipocytes was inhibited by 80%. The regulated expression pattern suggesting this factor as an adipose sensor for the nutritional state of the animals and the inhibitory effect on adipocyte differentiation implicate its function as a feedback regulator of adipogenesis. PMID- 11278255 TI - Phosphopantothenoylcysteine synthetase from Escherichia coli. Identification and characterization of the last unidentified coenzyme A biosynthetic enzyme in bacteria. AB - Phosphopantothenoylcysteine synthase catalyzes the formation of (R)-4'-phospho-N pantothenoylcysteine from 4'-phosphopantothenate and l-cysteine: this enzyme, involved in the biosynthesis of coenzyme A (CoA), has not previously been identified. Recently it was shown that the NH(2)-terminal domain of the Dfp protein from bacteria catalyzes the next step in CoA biosynthesis, the decarboxylation of (R)-4'-phospho-N-pantothenoylcysteine to form 4' phosphopantetheine (Kupke, T., Uebele, M., Schmid, D., Jung, G., Blaesse, M., and Steinbacher, S. (2000) J. Biol. Chem. 275, 31838-31846). We have partially purified phosphopantothenoylcysteine decarboxylase from Escherichia coli and demonstrated that the protein encoded by the dfp gene, here renamed coaBC, also has phosphopantothenoylcysteine synthetase activity, using CTP rather than ATP as the activating nucleoside 5'-triphosphate. This discovery completes the identification of all the enzymes involved in the biosynthesis of coenzyme A in bacteria. PMID- 11278257 TI - FP prostanoid receptor activation of a T-cell factor/beta -catenin signaling pathway. AB - FP prostanoid receptors are G-protein-coupled receptors (GPCR) that consist of two known isoforms, FP(A) and FP(B). These isoforms, which are generated by alternative mRNA splicing, are identical except for their carboxyl-terminal domains. Previously we have shown that stimulation of both isoforms with prostaglandin F(2alpha) (PGF(2alpha)) activates the small G-protein Rho, leading to morphological changes consisting of cell rounding and the formation of cell aggregates. Following the removal of PGF(2alpha), however, FP(A)-expressing cells show rapid reversal of cell rounding, whereas FP(B)-expressing cells do not. We now show that acute treatment of FP(B)-expressing cells with PGF(2alpha) leads to a subcellular reorganization of beta-catenin, a decrease in the phosphorylation of cytoplasmic beta-catenin, and persistent stimulation of Tcf/Lef-mediated transcriptional activation. This does not occur in FP(A)-expressing cells and may underlie the differences between these isoforms with respect to the reversal of cell rounding. The Tcf/beta-catenin signaling pathway is known to mediate the actions of Wnt acting through the heptahelical receptor, Frizzled, and has not been associated previously with GPCR activation. Our findings expand the signaling possibilities for GPCRs and suggest novel roles for FP receptors in normal tissue development and malignant transformation. PMID- 11278256 TI - Biochemical analysis of transcriptional repression by Drosophila histone deacetylase 1. AB - To study the mechanisms by which deacetylases regulate transcription by RNA polymerase II, we investigated the biochemical properties of purified recombinant Drosophila histone deacetylase 1 (dHDAC1, also known as dRPD3). We found that purified dHDAC1 and Gal4-dHDAC1 polypeptides possess substantial deacetylase activity. Thus, deacetylation by dHDAC1 does not require any additional cofactors. Gal4-dHDAC1, but not dHDAC1, was observed to repress transcription in vitro by about 2-3-fold from chromatin templates, but not from naked DNA templates, in a Gal4 site-dependent manner. This magnitude of repression is similar to that commonly seen by deacetylases in vivo, as assessed by treatment of cells with deacetylase inhibitors. Transcriptional repression by Gal4-dHDAC1 was blocked by the deacetylase inhibitor, FR901228, and thus, deacetylase activity correlates with repression. Single round transcription analyses showed that Gal4-dHDAC1 reduces the absolute number of productive initiation complexes with chromatin templates. Moreover, with chromatin templates that were assembled with completely purified components, Gal4-dHDAC1 was found to deacetylate nucleosomal histones as well as to repress transcription. These experiments provide biochemical evidence for the requirement of chromatin for transcriptional repression by dHDAC1 and further show that dHDAC1 acts to repress the transcription initiation process. PMID- 11278258 TI - Huntingtin's neuroprotective activity occurs via inhibition of procaspase-9 processing. AB - Huntington's Disease is an inherited neurodegenerative disease that affects the medium spiny neurons in the striatum. The disease is caused by the expansion of a polyglutamine sequence in the N terminus of Huntingtin (Htt), a widely expressed protein. Recently, we have found that Htt is an antiapoptotic protein in striatal cells and acts by preventing caspase-3 activity. Here we report that Htt overexpression in other CNS-derived cells can protect them from more than 20 days exposure to fatal stimuli. In particular, we found that cytochrome c continues to be released from mitochondria into the cytosol of cells that overexpress normal Htt. However, procaspase-9 is not processed, indicating that wild-type Htt (wtHtt) acts downstream of cytochrome c release. These data show that Htt inhibits neuronal cell death by interfering with the activity of the apoptosome complex. PMID- 11278259 TI - Binding of CO at the Pro2 side is crucial for the activation of CO-sensing transcriptional activator CooA. (1)H NMR spectroscopic studies. AB - CooA is a heme-containing transcriptional activator that anaerobically binds to DNA at CO atmosphere. To obtain information on the conformational transition of CooA induced by CO binding to the heme, we assigned ring current-shifted (1)H NMR signals of CooA using two mutants whose axial ligands of the heme were replaced. In the absence of CO, the NMR spectral pattern of H77Y CooA, in which the axial histidine (His(77)) was replaced with tyrosine, was similar to that of wild-type CooA. In contrast, the spectra of CooADeltaN5, in which the NH(2) termini including the other axial ligand (Pro(2)) were deleted, were drastically modulated. We assigned three signals of wild-type CooA at -4.5, -3.6, and -2.8 ppm to delta(1)-, alpha-, and delta(2)-protons of Pro(2), respectively. The Pro(2) signals were undetectable in the upfield region of the spectrum of the CO bound state, which confirms that CO displaces Pro(2). Interestingly, the Pro(2) signals were observed for CO-bound H77Y CooA, implying that CO binds to the trans position of Pro(2) in H77Y CooA. The abolished CO-dependent transcriptional activity of H77Y CooA is therefore the consequence of Pro(2) ligation. These observations are consistent with the view that the movement of the NH(2) terminus triggers the conformational transition to the DNA binding form. PMID- 11278260 TI - A high affinity acceptor for phospholipase A2 with neurotoxic activity is a calmodulin. AB - One of the high affinity binding proteins for ammodytoxin C, a snake venom presynaptically neurotoxic phospholipase A(2), has been purified from porcine cerebral cortex and characterized. After extraction from the membranes, the toxin binding protein was isolated in a homogenous form using wheat germ lectin Sepharose, Q-Sepharose, and ammodytoxin-CH-Sepharose chromatography. The specific binding of (125)I-ammodytoxin C to the isolated acceptor was inhibited to different extents by some neurotoxic phospholipases A(2), ammodytoxins, bee venom phospholipase A(2), agkistrodotoxin, and crotoxin; but not by nontoxic phospholipases A(2), ammodytin I(2), porcine pancreatic phospholipase A(2), and human type IIA phospholipase A(2); suggesting the significance of the acceptor in the mechanism of phospholipase A(2) neurotoxicity. The isolated acceptor was identified as calmodulin by tandem mass spectrometry. Since calmodulin is generally considered as an intracellular protein, the identity of this acceptor supports the view that secretory phospholipase A(2) neurotoxins have to be internalized to exert their toxic effect. Moreover, since ammodytoxin is known to block synaptic transmission, its interaction with calmodulin as an acceptor may constitute a valuable probe for further investigation of the role of the latter in this Ca(2+)-regulated process. PMID- 11278261 TI - The ARG tyrosine kinase interacts with Siva-1 in the apoptotic response to oxidative stress. AB - The Abl family of mammalian nonreceptor tyrosine kinases consists of c-Abl and ARG (Abl-related gene). Certain insights are available regarding the involvement c-Abl in the response of cells to stress. ARG, however, has no known function in cell signaling. The present studies demonstrate that ARG associates with the proapoptotic Siva-1 protein. The functional significance of the ARG-Siva-1 interaction is supported by the finding that ARG is activated by oxidative stress and that this response involves ARG-mediated phosphorylation of Siva-1 on Tyr(48). The proapoptotic effects of Siva-1 are accentuated in cells stably expressing ARG and are inhibited in ARG-deficient cells. Moreover, the proapoptotic effects of Siva-1 are abrogated by mutation of the Tyr(48) site. We also show that the apoptotic response to oxidative stress is attenuated in ARG deficient cells and that this defect is corrected by reconstituting ARG expression. These findings support a model in which the activation of ARG by oxidative stress induces apoptosis by a Siva-1-dependent mechanism. PMID- 11278262 TI - Isolation of a novel interleukin-1-inducible nuclear protein bearing ankyrin repeat motifs. AB - We isolated a novel gene termed interleukin (IL)-1-inducible nuclear ankyrin repeat protein (INAP), of which expression was specifically induced by IL-1 in OP9 stromal cells. The INAP has ankyrin-repeat motifs and shares weak amino acid sequence homology with Bcl-3 and other IkappaB family members. The human genomic INAP gene found in the NCBI data base is located at chromosome 3q3.11. Northern blot analyses revealed that INAP was not expressed in any examined tissues without stimulation, but INAP expression was rapidly and transiently induced by IL-1 although not by tumor necrosis factor alpha nor by phorbol 12-myristate 13 acetate in OP9 cells. Immunoblots with anti-INAP-specific antibody demonstrated that INAP was rapidly and specifically produced by IL-1 stimulation and was predominantly localized in the nucleus. Immunofluorescence stainings showed that the INAP newly synthesized by IL-1 stimulation was promptly translocated into the nucleus, and FLAG-tagged INAP forcibly expressed in NIH/3T3 cells was also specifically localized in the nucleus. The possible interaction of INAP with RelA/p65, NF-kappaB1/p50, NF-kappaB2/p52, C/EBPbeta, and retinoid X receptor was examined, but we could detect none of these interactions in the nuclear extracts of IL-1-stimulated cells. Unlike Bcl-3 and other IkappaB family members, INAP may play a unique role in IL-1-induced specific gene expression and/or signal transduction in the nucleus. PMID- 11278263 TI - Protein kinase G activates the JNK1 pathway via phosphorylation of MEKK1. AB - We recently obtained evidence that treatment of human colon cancer cells with exisulind (sulindac sulfone) and related compounds induces apoptosis by activation of protein kinase G (PKG) and c-Jun kinase (JNK1). The present study further explores this mechanism. We demonstrate that in NIH3T3 cells a constitutively active mutant of PKG causes a dose-dependent activation of JNK1 and thereby transactivates c-Jun and stimulates transcription from the AP-1 enhancer element. The activation of JNK1 and the transactivation of c-Jun by this mutant of PKG were inhibited by a dominant negative MEKK1. In vitro assays showed that a purified PKG directly phosphorylated the N-terminal domain of MEKK1. PKG also directly phosphorylated a full-length MEKK1, and this was associated with enhanced MEKK1 phosphorylation. Thus, it appears that PKG activates JNK1 through a novel PKG-MEKK1-SEK1-JNK1 pathway, by directly phosphorylating and activating MEKK1. PMID- 11278264 TI - Heat shock protein 90 mediates the balance of nitric oxide and superoxide anion from endothelial nitric-oxide synthase. AB - The balance of nitric oxide (.NO) and superoxide anion (O(2)) plays an important role in vascular biology. The association of heat shock protein 90 (Hsp90) with endothelial nitric-oxide synthase (eNOS) is a critical step in the mechanisms by which eNOS generates.NO. As eNOS is capable of generating both.NO and O(2), we hypothesized that Hsp90 might also mediate eNOS-dependent O(2) production. To test this hypothesis, bovine coronary endothelial cells (BCEC) were pretreated with geldanamycin (GA, 10 microg/ml; 17.8 microm) and then stimulated with the calcium ionophore, (5 microm). GA significantly decreased -stimulated eNOS dependent nitrite production (p < 0.001, n = 4) and significantly increased stimulated eNOS-dependent O(2) production (p < 0.001, n = 8). increased phospho eNOS(Ser-1179) levels by >1.6-fold over vehicle (V)-treated levels. Pretreatment with GA by itself or with increased phospho-eNOS levels. In unstimulated V treated BCEC cultures low amounts of Hsp90 were found to associate with eNOS. Pretreatment with GA and/or increased the association of Hsp90 with eNOS. These data show that Hsp90 is essential for eNOS-dependent.NO production and that inhibition of ATP-dependent conformational changes in Hsp90 uncouples eNOS activity and increases eNOS-dependent O(2) production. PMID- 11278265 TI - An Escherichia coli mutant defective in lipid export. AB - Escherichia coli phospholipids and lipopolysaccharide, made on the inner surface of the inner membrane, are rapidly transported to the outer membrane by mechanisms that are not well characterized. We now report a temperature-sensitive mutant (WD2) with an A270T substitution in a trans-membrane region of the ABC transporter MsbA. As shown by (32)P(i) and (14)C-acetate labeling, export of all major lipids to the outer membrane is inhibited by approximately 90% in WD2 after 30 min at 44 degrees C. Transport of newly synthesized proteins is not impaired. Electron microscopy shows reduplicated inner membranes in WD2 at 44 degrees C, consistent with a key role for MsbA in lipid trafficking. PMID- 11278266 TI - Molecular determinants of ion permeation and selectivity in inositol 1,4,5 trisphosphate receptor Ca2+ channels. AB - We tested the hypothesis that key residues in a putative intraluminal loop contribute to determination of ion permeation through the intracellular Ca(2+) release channel (inositol 1,4,5-trisphosphate receptors (IP(3)Rs)) that is gated by the second messenger inositol 1,4,5-trisphosphate (IP(3)). To accomplish this, we mutated residues within the putative pore forming region of the channel and analyzed the functional properties of mutant channels using a (45)Ca(2+) flux assay and single channel electrophysiological analyses. Two IP(3)R mutations, V2548I and D2550E, retained the ability to release (45)Ca(2+) in response to IP(3). When analyzed at the single channel level; both recombinant channels had IP(3)-dependent open probabilities similar to those observed in wild-type channels. The mutation V2548I resulted in channels that exhibited a larger K(+) conductance (489 +/- 13 picosiemens (pS) for V2548I versus 364 +/- 5 pS for wild type), but retained a Ca(2+) selectivity similar to wild-type channels (P(Ca(2+)):P(K(+)) approximately 4:1). Conversely, D2550E channels were nonselective for Ca(2+) over K(+) (P(Ca(2+)):P(K(+)) approximately 0.6:1), while the K(+) conductance was effectively unchanged (391 +/- 4 pS). These results suggest that amino acid residues Val(2548) and Asp(2550) contribute to the ion conduction pathway. We propose that the pore of IP(3)R channels has two distinct sites that control monovalent cation permeation (Val(2548)) and Ca(2+) selectivity (Asp(2550)). PMID- 11278267 TI - The highly conserved protein methyltransferase, Skb1, is a mediator of hyperosmotic stress response in the fission yeast Schizosaccharomyces pombe. AB - The p21-activated kinase, Shk1, is required for cell viability, establishment and maintenance of cell polarity, and proper mating response in the fission yeast, Schizosaccharomyces pombe. Previous genetic studies suggested that a presumptive protein methyltransferase, Skb1, functions as a positive modulator of Shk1. However, unlike Shk1, Skb1 is not required for viability or mating of S. pombe cells and contributes only modestly to the regulation of cell morphology under normal growth conditions. Here we demonstrate that Skb1 plays a more significant role in regulating cell growth and polarity under conditions of hyperosmotic stress. We provide evidence that the inability of skb1Delta cells to properly maintain cell polarity in hyperosmotic conditions results from inefficient subcellular targeting of F-actin. We show that Skb1 localizes to cell ends, sites of septation, and nuclei of S. pombe cells. Hyperosmotic shock results in substantial delocalization of Skb1 from cell ends and nuclei, as well as stimulation of Skb1 protein methyltransferase activity. Taken together, our results demonstrate a new role for Skb1 as a mediator of hyperosmotic stress response in fission yeast. We show that the protein methyltransferase activity of the human Skb1 homolog, Skb1Hs, is also stimulated by hyperosmotic stress in fission yeast, providing evidence for evolutionary conservation of a role for Skb1-related proteins as mediators of hyperosmotic stress response, as well as mechanisms involved in regulating this novel class of protein methyltransferases. PMID- 11278268 TI - Effects of the NIK aly mutation on NF-kappaB activation by the Epstein-Barr virus latent infection membrane protein, lymphotoxin beta receptor, and CD40. AB - Homozygosity for the aly point mutation in NF-kappaB-inducing kinase (NIK) results in alymphoplasia in mice, a phenotype similar to that of homozygosity for deletion of the lymphotoxin beta receptor (LTbetaR). We now find that NF-kappaB activation by Epstein-Barr virus latent membrane protein 1 (LMP1) or by an LMP1 transmembrane domain chimera with the LTbetaR signaling domain in human embryonic kidney 293 cells is selectively inhibited by a wild type dominant negative NIK comprised of amino acids 624-947 (DN-NIK) and not by aly DN-NIK. In contrast, LMP1/CD40 is inhibited by both wild type (wt) and aly DN-NIK. LMP1, an LMP1 transmembrane domain chimera with the LTbetaR signaling domain, and LMP1/CD40 activate NF-kappaB in wt or aly murine embryo fibroblasts. Although wt and aly NIK do not differ in their in vitro binding to tumor necrosis factor receptor associated factor 1, 2, 3, or 6 or in their in vivo association with tumor necrosis factor receptor-associated factor 2 and differ marginally in their very poor binding to IkappaB kinase beta (IKKbeta), only wt NIK is able to bind to IKKalpha. These data are compatible with a model in which activation of NF-kappaB by LMP1 and LTbetaR is mediated by an interaction of NIK or a NIK-like kinase with IKKalpha that is abrogated by the aly mutation. On the other hand, CD40 mediates NF-kappaB activation through a kinase that interacts with a different component of the IKK complex. PMID- 11278270 TI - Peroxisome proliferator-activated receptor-gamma ligands inhibit adipocyte 11beta -hydroxysteroid dehydrogenase type 1 expression and activity. AB - Peroxisome proliferator-activated receptor-gamma (PPARgamma) has been shown to play an important role in the regulation of expression of a subclass of adipocyte genes and to serve as the molecular target of the thiazolidinedione (TZD) and certain non-TZD antidiabetic agents. Hypercorticosteroidism leads to insulin resistance, a variety of metabolic dysfunctions typically seen in diabetes, and hypertrophy of visceral adipose tissue. In adipocytes, the enzyme 11beta hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) converts inactive cortisone into the active glucocorticoid cortisol and thereby plays an important role in regulating the actions of corticosteroids in adipose tissue. Here, we show that both TZD and non-TZD PPARgamma agonists markedly reduced 11beta-HSD-1 gene expression in 3T3-L1 adipocytes. This diminution correlated with a significant decrease in the ability of the adipocytes to convert cortisone to cortisol. The half-maximal inhibition of 11beta-HSD-1 mRNA expression by the TZD, rosiglitazone, occurred at a concentration that was similar to its K(d) for binding PPARgamma and EC(50) for inducing adipocyte differentiation thereby indicating that this action was PPARgamma-dependent. The time required for the inhibitory action of the TZD was markedly greater for 11beta-HSD-1 gene expression than for leptin, suggesting that these genes may be down-regulated by different molecular mechanisms. Furthermore, whereas regulation of PPARgamma inducible genes such as phosphoenolpyruvate carboxykinase was maintained when cellular protein synthesis was abrogated, PPARgamma agonist inhibition of 11beta HSD-1 and leptin gene expression was ablated, thereby supporting the conclusion that PPARgamma affects the down-regulation of 11beta-HSD-1 indirectly. Finally, treatment of diabetic db/db mice with rosiglitazone inhibited expression of 11beta-HSD-1 in adipose tissue. This decrease in enzyme expression correlated with a significant decline in plasma corticosterone levels. In sum, these data indicate that some of the beneficial effects of PPARgamma antidiabetic agents may result, at least in part, from the down-regulation of 11beta-HSD-1 expression in adipose tissue. PMID- 11278269 TI - Cyclic AMP inhibits Akt activity by blocking the membrane localization of PDK1. AB - Akt is a protein serine/threonine kinase that plays an important role in the mitogenic responses of cells to variable stimuli. Akt contains a pleckstrin homology (PH) domain and is activated by phosphorylation at threonine 308 and serine 473. Binding of 3'-OH phosphorylated phosphoinositides to the PH domain results in the translocation of Akt to the plasma membrane where it is activated by upstream kinases such as (phosphoinositide-dependent kinase-1 (PDK1). Over expression of constitutively active forms of Akt promotes cell proliferation and survival, and also stimulates p70 S6 kinase (p70S6K). In many cells, an increase in levels of intracellular cyclic AMP (cAMP) diminishes cell growth and promotes differentiation, and in certain conditions cAMP is even antagonistic to the effect of growth factors. Here, we show that cAMP has inhibitory effects on the phosphatidylinositol 3-kinase/PDK/Akt signaling pathway. cAMP potently inhibits phosphorylation at threonine 308 and serine 473 of Akt, which is required for the protein kinase activities of Akt. cAMP also negatively regulates PDK1 by inhibiting its translocation to the plasma membrane, despite not affecting its protein kinase activities. Furthermore, when we co-expressed myristoylated Akt and PDK1 mutants which constitutively co-localize in the plasma membrane, Akt activity was no longer sensitive to raised intracellular cAMP concentrations. Finally, cAMP was also found to inhibit the lipid kinase activity of PI3K and to decrease the levels of phosphatidylinositol 3,4,5-triphosphate in vivo, which are required for the membrane localization of PDK1. Collectively, these data strongly support the theory that the cAMP-dependent signaling pathway inhibits Akt activity by blocking the coupling between Akt and its upstream regulators, PDK, in the plasma membrane. PMID- 11278272 TI - Hyaluronic acid induces survival and proliferation of human myeloma cells through an interleukin-6-mediated pathway involving the phosphorylation of retinoblastoma protein. AB - Originating from a post-switch memory B cell or plasma cell compartment in peripheral lymphoid tissues, malignant myeloma cells accumulate in the bone marrow of patients with multiple myeloma. In this favorable microenvironment their growth and survival are dependent upon both soluble factors and physical cell-to-cell and cell-to-extracellular matrix contacts. In this report we show that hyaluronan (HA), a major nonprotein glycosaminoglycan component of the extracellular matrix in mammalian bone marrow, is a survival and proliferation factor for human myeloma cells. The effect of HA is mainly mediated through a gp 80-interleukin 6 (IL-6) receptor pathway by a CD44-independent mechanism, suggesting that HA retains and concentrates IL-6 close to its site of secretion, thus favoring its autocrine activity. In addition, we show that HA-mediated survival and proliferation of myeloma cells is associated with a down-regulation in the expression of p27(kip1) cyclin-dependent kinase inhibitor and a hyperphosphorylation of the retinoblastoma protein (pRb). These data suggest that HA could be an important component in the myeloma cell physiopathology in vivo by potentiating autocrine and/or paracrine IL-6 activities. PMID- 11278271 TI - The CK2 phosphorylation of vitronectin. Promotion of cell adhesion via the alpha(v)beta 3-phosphatidylinositol 3-kinase pathway. AB - Phosphorylation of vitronectin (Vn) by casein kinase II was previously shown to occur at Thr50 and Thr57 and to augment a major physiological function of vitronectin-cell adhesion and spreading. Here we show that this phosphorylation increases cell adhesion via the alpha(v)beta3 (not via the alpha(v)beta5 integrin), suggesting that alpha(v)beta3 differs from alpha(v)beta5 in its biorecognition profile. Although both the phospho (CK2-PVn) and non-phospho (Vn) analogs of vitronectin (simulated by mutants Vn(T50E,T57E), and Vn(T50A,T57A), respectively) trigger the alpha(v)beta3 as well as the alpha(v)beta5 integrins, and equally activate the ERK pathway, these two forms are different in their activation of the focal adhesion kinase/phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB) pathway. Specifically, we show (i) that, upon exposure of cells to Vn/CK2-PVn, their PKB activation depends on the availability of the alpha(v)beta3 integrin on their surface; (ii) that upon adhesion of the beta3-transfected cells onto the CK2-PVn, the extent of PKB activation coincides with the enhanced adhesion of these cells, and (iii) that both the PKB activation and the elevation in the adhesion of these cells is PI3K-dependent. The occurrence of a cell surface receptor that specifically distinguishes between a phosphorylated and a non-phosphorylated analog of Vn, together with the fact that it preferentially activates a distinct intra-cellular signaling pathway, suggest that extra-cellular CK2 phosphorylation may play an important role in the regulation of cell adhesion and migration. PMID- 11278273 TI - Lithium and valproate decrease inositol mass and increase expression of the yeast INO1 and INO2 genes for inositol biosynthesis. AB - Bipolar affective disorder (manic-depressive illness) is a chronic, severe, debilitating illness affecting 1-2% of the population. The Food and Drug Administration-approved drugs lithium and valproate are not completely effective in the treatment of this disorder, and the mechanisms underlying their therapeutic effects have not been established. We are employing genetic and molecular approaches to identify common targets of lithium and valproate in the yeast Saccharomyces cerevisiae. We show that both drugs affect molecular targets in the inositol metabolic pathway. Lithium and valproate cause a decrease in intracellular myo-inositol mass and an increase in expression of both a structural (INO1) and a regulatory (INO2) gene required for inositol biosynthesis. The opi1 mutant, which exhibits constitutive expression of INO1, is more resistant to inhibition of growth by lithium but not by valproate, suggesting that valproate may inhibit the Ino1p-catalyzed synthesis of inositol 1 phosphate. Consistent with this possibility, growth in valproate leads to decreased synthesis of inositol monophosphate. Thus, both lithium and valproate perturb regulation of the inositol biosynthetic pathway, albeit via different mechanisms. This is the first demonstration of increased expression of genes in the inositol biosynthetic pathway by both lithium and valproate. Because inositol is a key regulator of many cellular processes, the effects of lithium and valproate on inositol synthesis have far-reaching implications for predicting genetic determinants of responsiveness and resistance to these agents. PMID- 11278275 TI - DNA sequence-dependent differences in TATA-binding protein-induced DNA bending in solution are highly sensitive to osmolytes. AB - The complex formed between the TATA-binding protein (TBP) and the "TATA box" of eukaryotic class II promoters is the foundation for assembly of the complex to which RNA polymerase II is ultimately recruited. TBP binds productively to canonical and diverse variant TATA sequences with >100-fold differences in transcription efficiency. Co-crystals of canonical sequences and >11 variant sequences bound to various TBP molecules all have approximately 80 degrees bends. In contrast, the bend angles for TBP-TATA complexes in solution, derived from distance distributions, are approximately 80 degrees for a canonical sequence but range from 30 degrees to 62 degrees for five variant sequences. We show in this study that the osmolytes used to crystallize TBP-TATA complexes induce profound increases in the DNA bends of two transcriptionally active TBP-bound variant sequences to a common angle of approximately 80 degrees but have little effect on a transcriptionally inactive variant. The effect of osmolyte on the TBP-induced DNA bend of a variant TATA box sequence is also manifest in the kinetics of association, demonstrating a functional consequence of an osmolyte-induced structural change. PMID- 11278274 TI - Ester bond-containing tea polyphenols potently inhibit proteasome activity in vitro and in vivo. AB - It has been discovered that proteasome inhibitors are able to induce tumor growth arrest or cell death and that tea consumption is correlated with cancer prevention. Here, we show that ester bond-containing tea polyphenols, such as (-) epigallocatechin-3-gallate (EGCG), potently and specifically inhibit the chymotrypsin-like activity of the proteasome in vitro (IC(50) = 86-194 nm) and in vivo (1-10 microm) at the concentrations found in the serum of green tea drinkers. Atomic orbital energy analyses and high performance liquid chromatography suggest that the carbon of the polyphenol ester bond is essential for targeting, thereby inhibiting the proteasome in cancer cells. This inhibition of the proteasome by EGCG in several tumor and transformed cell lines results in the accumulation of two natural proteasome substrates, p27(Kip1) and IkappaB alpha, an inhibitor of transcription factor NF-kappaB, followed by growth arrest in the G(1) phase of the cell cycle. Furthermore, compared with their simian virus-transformed counterpart, the parental normal human fibroblasts were much more resistant to EGCG-induced p27(Kip1) protein accumulation and G(1) arrest. Our study suggests that the proteasome is a cancer-related molecular target of tea polyphenols and that inhibition of the proteasome activity by ester bond containing polyphenols may contribute to the cancer-preventative effect of tea. PMID- 11278276 TI - DNA bends in TATA-binding protein-TATA complexes in solution are DNA sequence dependent. AB - The TATA-binding protein (TBP) initiates assembly of transcription preinitiation complexes on eukaryotic class II promoters, binding to and restructuring consensus and variant "TATA box" sequences. The sequence dependence of the DNA structure in TBP-TATA complexes has been investigated in solution using fluorescence resonance energy transfer. The mean 5'dye-3'dye distance varies significantly among oligomers bearing the adenovirus major late promoter sequence (AdMLP) and five single-site variants bound to Saccharomyces cerevisiae TBP, consistent with solution bend angles for AdMLP of 76 degrees and for the variants ranging from 30 degrees to 62 degrees. These solution bends contrast sharply with the corresponding co-crystal structures, which show approximately 80 degrees bends for all sequences. Transcription activities for these TATA sequences are strongly correlated with the solution bend angles but not with TBP-DNA binding affinities. Our results support a model in which transcription efficiency derives primarily from the sequence-dependent structure of the TBP-TATA binary complex. Specifically, the distance distribution for the average solution structure of the TBP-TATA complex may reflect the sequence-dependent probability for the complex to assume a conformation in which the TATA box DNA is severely bent. Upon assumption of this geometry, the binary complex becomes a target for binding and correctly orienting the other components of the preinitiation complex. PMID- 11278277 TI - Role of ATM in oxidative stress-mediated c-Jun phosphorylation in response to ionizing radiation and CdCl2. AB - Ionizing radiation-induced phosphorylation of the transcription factor c-Jun is impaired in cells derived from individuals with ataxia telangiectasia (AT), in which the ATM gene is mutated. We demonstrate here that ATM modulates c-Jun phosphorylation following exposure to ionizing radiation as well as treatment with CdCl(2), a potent pro-oxidant. Exposure of AT and control fibroblasts to CdCl(2) induced a biphasic increase in c-Jun phosphorylation on serine residues 63 and 73, with the extent of the second phase being markedly greater in AT cells than in control cells. Heme oxygenase-1, a marker of oxidative stress, was also significantly induced in AT fibroblasts. Expression of recombinant ATM in AT fibroblasts, however, reduced the extent of the effects of CdCl(2) on both c-Jun phosphorylation and heme oxygenase-1 induction. Our data suggest that ATM contributes to oxidative stress-mediated signaling that leads to c-Jun phosphorylation by acting as a sensor of ionizing radiation-induced oxidative stress and by modulating intracellular redox homeostasis. PMID- 11278278 TI - Comodulation of CXCR4 and CD26 in human lymphocytes. AB - We provide convergent and multiple evidence for a CD26/CXCR4 interaction. Thus, CD26 codistributes with CXCR4, and both coimmunoprecipitate from membranes of T (CD4(+)) and B (CD4(-)) cell lines. Upon induction with stromal cell-derived factor 1alpha (SDF-1alpha), CD26 is cointernalized with CXCR4. CXCR4-mediated down-regulation of CD26 is not induced by antagonists or human immunodeficiency virus (HIV)-1 gp120. SDF-1alpha-mediated down-regulation of CD26 is not blocked by pertussis toxin but does not occur in cells expressing mutant CXCR4 receptors unable to internalize. Codistribution and cointernalization also occurs in peripheral blood lymphocytes. Since CD26 is a cell surface endopeptidase that has the capacity to cleave SDF-1alpha, the CXCR4.CD26 complex is likely a functional unit in which CD26 may directly modulate SDF-1alpha-induced chemotaxis and antiviral capacity. CD26 anchors adenosine deaminase (ADA) to the lymphocyte cell surface, and this interaction is blocked by HIV-1 gp120. Here we demonstrate that gp120 interacts with CD26 and that gp120-mediated disruption of ADA/CD26 interaction is a consequence of a first interaction of gp120 with a domain different from the ADA binding site. SDF-1alpha and gp120 induce the appearance of pseudopodia in which CD26 and CXCR4 colocalize and in which ADA is not present. The physical association of CXCR4 and CD26, direct or part of a supramolecular structure, suggests a role on the function of the immune system and the pathophysiology of HIV infection. PMID- 11278280 TI - Mechanism of factor Va inactivation by plasmin. Loss of A2 and A3 domains from a Ca2+-dependent complex of fragments bound to phospholipid. AB - The coagulation cofactor Va (FVa) is a noncovalent heterodimer consisting of a heavy chain (FVaH) and a light chain (FVaL). Previously, the fibrinolytic effector plasmin (Pn) has been shown to inhibit FVa function. To understand this mechanism, the fragmentation profile of human FVa by Pn and the noncovalent association of the derived fragments were determined in the presence of Ca(2+) using anionic phospholipid (aPL)-coated microtiter wells and large (1 microm) aPL micelles as affinity matrices. Following Pn inactivation of aPL-bound FVa, a total of 16 fragments were observed and their NH(2) termini sequenced. These had apparent molecular weights and starting residues as follows (single letter abbreviation is used): 50(L1766), 48(L1766), 43(Q1828), 40(Q1828), 30(S1546), 12(T1657), and 7(S1546) kDa from FVaL; and 65(A1), 50(A1), 45(A1), 34(S349), 30(L94), 30(M110), and 3 small <5(W457, W457, and K365) kDa from FVaH. Of these, 50(L1766), 48(1766), 43(Q1828), and 40(Q1828) spanning the C1/C2 domains, and 30(L94), but not the similar 30(M110), positioned within the A1 domain remained associated with aPL. These were detected antigenically during Pn- or tissue plasminogen activator-mediated lysis of fibrin clot formed in plasma. Chelation by EDTA dissociated the 30(L94)-kDa fragment, which was observed to associate with intact FVaL upon recalcification, indicating that the Leu-94 to Lys-109 region of the A1 domain plays a critical role in the FVaL and FVaH Ca(2+) dependent association. By using domain-specific monoclonal antibodies and an assay for thrombin generation, loss of FVa prothrombinase function was coincident with proteolysis at sites in the A2 and A3 domains resulting in their dissociation. Inactivation of FV or FVa by Pn was independent of the thrombophilic R506Q mutation. These results identify the molecular composition of Pn-cleaved FVa that remains bound to membrane as largely A1-C1/C2 in the presence of Ca(2+) and suggest that Pn inhibits FVa by a process involving A2 and A3 domain dissociation. PMID- 11278279 TI - UVA induces Ser381 phosphorylation of p90RSK/MAPKAP-K1 via ERK and JNK pathways. AB - UVA exposure plays an important role in the etiology of skin cancer. The family of p90-kDa ribosomal S6 kinases (p90(RSK)/MAPKAP-K1) are activated via phosphorylation. In this study, results show that UVA-induced phosphorylation of p90(RSK) at Ser(381) through ERKs and JNKs, but not p38 kinase pathways. We provide evidence that UVA-induced p90(RSK) phosphorylation and kinase activity were time- and dose-dependent. Both PD98059 and a dominant negative mutant of ERK2 blocked ERKs and p90(RSK) Ser(381) phosphorylation, as well as p90(RSK) activity. A dominant negative mutant of p38 kinase blocked UVA-induced phosphorylation of p38 kinase, but had no effect on UVA-induced Ser(381) phosphorylation of p90(RSK) or kinase activity. UVA-induced p90(RSK) phosphorylation and kinase activity were markedly attenuated in JnK1(-/-) and JnK2(-/-) cells. A dominant negative mutant of JNK1 inhibited UVA-induced JNKs and p90(RSK) phosphorylation and kinase activity, but had no effect on ERKs phosphorylation. PD169316, a novel inhibitor of JNKs and p38 kinase, inhibited phosphorylation of p90(RSK), JNKs, and p38 kinase, but not ERKs. However, SB202190, a selective inhibitor of p38 kinase, had no effect on p90(RSK) or JNKs phosphorylation. Significantly, ERKs and JNKs, but not p38 kinase, immunoprecipitated with p90(RSK) when stimulated by UVA and p90(RSK) was a substrate for ERK2 and JNK2, but not p38 kinase. These data indicate clearly that p90(RSK) Ser(381) may be phosphorylated by activation of JNKs or ERKs, but not p38 kinase. PMID- 11278281 TI - Intercellular adhesion molecule-1 (ICAM-1) gene expression in human T cells is regulated by phosphotyrosyl phosphatase activity. Involvement of NF-kappaB, Ets, and palindromic interferon-gamma-responsive element-binding sites. AB - Intercellular adhesion molecule-1 (ICAM-1) plays an important role in adhesion phenomena involved in the immune response. The strength of adhesion has been shown to be modulated by changes in ICAM-1 gene expression. In T cells, signaling pathways are intimately regulated by an equilibrium between protein-tyrosine kinases and protein tyrosine phosphatases (PTP). The use of bis-peroxovanadium (bpV) compounds, a class of potent PTP inhibitors, enabled us to investigate the involvement of phosphotyrosyl phosphatases in the regulation of ICAM-1 gene expression in human T cells. Here, we demonstrate for the first time that inhibition of PTP results in an increase of ICAM-1 surface expression on both human T lymphoid and primary mononuclear cells. The crucial role played by the NF kappaB-, Ets-, and pIgammaRE-binding sites in bpV[pic]-mediated activation of ICAM-1 was demonstrated using various 5' deletion and site-specific mutants of the ICAM-1 gene promoter driving the luciferase reporter gene. Co-transfection experiments with trans-dominant mutants and electrophoretic mobility shift assays confirmed the importance of constitutive and inducible transcription factors that bind to specific responsive elements in bpV-dependent up-regulation of ICAM-1 surface expression. Altogether, these observations suggest that expression of ICAM-1 in human T cells is regulated by phosphotyrosyl phosphatase activity through NF-kappaB-, Ets-, and STAT-1-dependent signaling pathways. PMID- 11278282 TI - Localization and stability of introns spliced from the Pem homeobox gene. AB - RNA splicing generates two products in equal molar amounts, mature mRNAs and spliced introns. Although the mechanism of RNA splicing and the fate of the spliced mRNA products have been well studied, very little is known about the fate and stability of most spliced introns. Research in this area has been hindered by the widely held view that most vertebrate introns are too unstable to be detectable. Here, we report that we are able to detect all three spliced introns from the coding region of the Pem homeobox gene. By using a tetracycline (tet) regulated promoter, we found that the half-lives of these Pem introns ranged from 9 to 29 min, comparable with those of short lived mRNAs such as those encoding c fos and c-myc. The half-lives of the Pem introns correlated with both their length and 5' to 3' orientation in the Pem gene. Subcellular fractionation analysis revealed that spliced Pem introns and pre-mRNA accumulated in the nuclear matrix, high salt-soluble, and DNase-sensitive fractions within the nucleus. Surprisingly, we found that all three of the spliced Pem introns were also in the cytoplasmic fraction, whereas Pem pre-mRNAs, U6 small nuclear RNA, and a spliced intron from another gene were virtually excluded from this fraction. This indicates either that spliced Pem introns are uniquely exported to the cytoplasm for degradation or they reside in a unique soluble nuclear fraction. Our study has implications for understanding the regulation of RNA metabolism, as the stability of introns and the location of their degradation may dictate the following: (i) the stability of nearby mRNAs that compete with spliced introns for rate-limiting nucleases, (ii) the rate at which free nucleotides are available for further rounds of transcription, and (iii) the rate at which splicing factors are recycled. PMID- 11278283 TI - MST, a physiological caspase substrate, highly sensitizes apoptosis both upstream and downstream of caspase activation. AB - The human serine/threonine kinase, mammalian STE20-like kinase (MST), is considerably homologous to the budding yeast kinases, SPS1 and STE20, throughout their kinase domains. The cellular function and physiological activation mechanism of MST is unknown except for the proteolytic cleavage-induced activation in apoptosis. In this study, we show that MST1 and MST2 are direct substrates of caspase-3 both in vivo and in vitro. cDNA cloning of MST homologues in mouse and nematode shows that caspase-cleaved sequences are evolutionarily conserved. Human MST1 has two caspase-cleavable sites, which generate biochemically distinct catalytic fragments. Staurosporine activates MST either caspase-dependently or independently, whereas Fas ligation activates it only caspase-dependently. Immunohistochemical analysis reveals that MST is localized in the cytoplasm. During Fas-mediated apoptosis, cleaved MST translocates into the nucleus before nuclear fragmentation is initiated, suggesting it functions in the nucleus. Transiently expressed MST1 induces striking morphological changes characteristic of apoptosis in both nucleus and cytoplasm, which is independent of caspase activation. Furthermore, when stably expressed in HeLa cells, MST highly sensitizes the cells to death receptor-mediated apoptosis by accelerating caspase-3 activation. These findings suggest that MST1 and MST2 play a role in apoptosis both upstream and downstream of caspase activation. PMID- 11278284 TI - Elevated AKT activity protects the prostate cancer cell line LNCaP from TRAIL induced apoptosis. AB - We find that the prostate cancer cell lines ALVA-31, PC-3, and DU 145 are highly sensitive to apoptosis induced by TRAIL (tumor-necrosis factor-related apoptosis inducing ligand), while the cell lines TSU-Pr1 and JCA-1 are moderately sensitive, and the LNCaP cell line is resistant. LNCaP cells lack active lipid phosphatase PTEN, a negative regulator of the phosphatidylinositol (PI) 3 kinase/Akt pathway, and demonstrate a high constitutive Akt activity. Inhibition of PI 3-kinase using wortmannin and LY-294002 suppressed constitutive Akt activity and sensitized LNCaP cells to TRAIL. Treatment of LNCaP cells with TRAIL alone induced cleavage of the caspase 8 and XIAP proteins. However, processing of BID, mitochondrial release of cytochrome c, activation of caspases 7 and 9, and apoptosis did not occur unless TRAIL was combined with either wortmannin, LY 294002, or cycloheximide. Blocking cytochrome c release by Bcl-2 overexpression rendered LNCaP cells resistant to TRAIL plus wortmannin treatment but did not affect caspase 8 or BID processing. This indicates that in these cells mitochondria are required for the propagation rather than the initiation of the apoptotic cascade. Infection of LNCaP cells with an adenovirus expressing a constitutively active Akt reversed the ability of wortmannin to potentiate TRAIL induced BID cleavage. Thus, the PI 3-kinase-dependent blockage of TRAIL-induced apoptosis in LNCaP cells appears to be mediated by Akt through the inhibition of BID cleavage. PMID- 11278285 TI - Light-induced photoreceptor apoptosis in vivo requires neuronal nitric-oxide synthase and guanylate cyclase activity and is caspase-3-independent. AB - Apoptosis is the mode of photoreceptor cell death in inherited and induced retinal degeneration. However, the molecular mechanisms of photoreceptor cell death in human cases and animal models of retinal dystrophies remain undefined. Exposure of Balb/c mice to excessive levels of white light results in photoreceptor apoptosis. This study delineates the molecular events occurring during and subsequent to the induction of retinal degeneration by exposure to white light in Balb/c mice. We demonstrate an early increase in intracellular calcium levels during photoreceptor apoptosis, an event that is accompanied by significant superoxide generation and mitochondrial membrane depolarization. Furthermore, we show that inhibition of neuronal nitric-oxide synthase (nNOS) by 7-nitroindazole is sufficient to prevent retinal degeneration implicating a key role for neuronal nitric oxide (NO) in this model. We demonstrate that inhibition of guanylate cyclase, a downstream effector of NO, also prevents photoreceptor apoptosis demonstrating that guanylate cyclase too plays an essential role in this model. Finally, our results demonstrate that caspase-3, frequently considered to be one of the key executioners of apoptosis, is not activated during retinal degeneration. In summary, the data presented here demonstrate that light-induced photoreceptor apoptosis in vivo is mediated by the activation of nNOS and guanylate cyclase and is caspase-3-independent. PMID- 11278286 TI - Heterodimerization of mineralocorticoid and glucocorticoid receptors at a novel negative response element of the 5-HT1A receptor gene. AB - Negative regulation of neuronal serotonin (5-HT1A) receptor levels by glucocorticoids in vivo may contribute to depression. Both types I (mineralocorticoid) and II (glucocorticoid) receptors (MR and GR, respectively) participate in corticosteroid-induced transcriptional repression of the 5-HT1A gene; however, the precise mechanism is unclear. A direct repeat 6-base pair glucocorticoid response element (GRE) half-site 5'-TGTCCT separated by 6 nucleotides was conserved in human, mouse, and rat 5-HT1A receptor promoters. In SN-48 neuronal cells that express MR, GR, and 5-HT1A receptors, deletion or inactivation of the nGRE (negative GRE) eliminated negative regulation of the rat 5-HT1A or heterologous promoters by corticosteroids, whereas its inclusion conferred corticosteroid-induced inhibition to a heterologous promoter. Bacterially expressed recombinant MR and GR preferentially bound to the nGRE as a heterodimer, as identified in nuclear extracts of MR/GR-transfected COS-7 cells, and with higher affinity than MR or GR homodimers. In SN48 and COS-7 cells, concentration-dependent coactivation of MR and GR was required for maximal inhibitory action by corticosteroids and was abrogated in the L501P-GR mutant lacking DNA binding activity. Corticosteroid-mediated transcriptional inhibition was greater for MR/GR in combination than for MR or GR alone. These data represent the first identification of an nMRE/GRE and indicate that heterodimerization of MR and GR mediates direct corticosteroid-induced transrepression of the 5-HT1A receptor promoter. PMID- 11278287 TI - 14-3-3 is involved in p75 neurotrophin receptor-mediated signal transduction. AB - The low affinity neurotrophin receptor (p75NTR) has been shown to mediate the apoptosis signaling to neural cells. However, the specific mechanisms of intracellular signal transduction of this process are largely unknown. To understand p75NTR-mediated signal transduction, we previously identified a protein that interacts with the intracellular domain of p75NTR, and we named it p75NTR-associated cell death executor (NADE). To elucidate further the signaling mechanisms utilized by p75NTR and NADE, we screened for NADE-binding protein(s) with the yeast two-hybrid method, and we identified 14-3-3epsilon as a NADE binding protein in vivo. To examine whether 14-3-3epsilon affects the induction of p75NTR-mediated apoptosis, wild type or various deletion mutant forms of 14-3 3epsilon were co-expressed in HEK293, PC12nnr5, and oligodendrocytes. Interestingly, transient expression of the mutant form of 14-3-3epsilon lacking the 208-255 amino acid region blocked nerve growth factor-dependent p75NTR/NADE mediated apoptosis, although this mutant form of 14-3-3epsilon continued to associate with NADE. These results suggest that 14-3-3epsilon plays an important role in the modulation of nerve growth factor-dependent p75NTR/NADE-mediated apoptosis. PMID- 11278288 TI - Involvement of deacylation in activation of substrate hydrolysis by Drosophila acetylcholinesterase. AB - Insect acetylcholinesterase (AChE), an enzyme whose catalytic site is located at the bottom of a gorge-like structure, hydrolyzes its substrate over a wide range of concentrations (from 2 microm to 300 mm). AChE is activated at low substrate concentrations and inhibited at high substrate concentrations. Several rival kinetic models have been developed to try to describe and explain this behavior. One of these models assumes that activation at low substrate concentrations partly results from an acceleration of deacetylation of the acetylated enzyme. To test this hypothesis, we used a monomethylcarbamoylated enzyme, which is considered equivalent to the acylated form of the enzyme and a non-hydrolyzable substrate analog, 4-oxo-N,N,N-trimethylpentanaminium iodide. It appears that this substrate analog increases the decarbamoylation rate by a factor of 2.2, suggesting that the substrate molecule bound at the activation site (K(d) = 130 +/- 47 microm) accelerates deacetylation. These two kinetic parameters are consistent with our analysis of the hydrolysis of the substrate. The location of the active site was investigated by in vitro mutagenesis. We found that this site is located at the rim of the active site gorge. Thus, substrate positioning at the rim of the gorge slows down the entrance of another substrate molecule into the active site gorge (Marcel, V., Estrada-Mondaca, S., Magne, F., Stojan, J., Klaebe, A., and Fournier, D. (2000) J. Biol. Chem. 275, 11603-11609) and also increases the deacylation step. This results in an acceleration of enzyme turnover. PMID- 11278290 TI - Transcriptional cross-talk between Smad, ERK1/2, and p38 mitogen-activated protein kinase pathways regulates transforming growth factor-beta-induced aggrecan gene expression in chondrogenic ATDC5 cells. AB - In chondrogenesis, members of the transforming growth factor-beta (TGF-beta) superfamily play critical roles by inducing gene expression of cartilage-specific molecules. By using a chondrogenic cell line, ATDC5, we investigated the TGF-beta mediated signaling pathways involved in expression of the aggrecan gene (Agc). At confluency, TGF-beta induced Agc expression within 3 h, and cycloheximide blocked this induction, indicating that de novo protein synthesis is essential for this response. At this stage, TGF-beta induced rapid, transient phosphorylation of Smad2, extracellular signal-activated kinase 1/2 (ERK1/2), and p38 mitogen activated protein kinase (MAPK). Inhibition of the Smad pathways by transfection with a dominant negative Smad4 construct significantly reduced TGF-beta-induced Agc expression, indicating that Smad signaling is essential for this response. Furthermore, an inhibitor of the ERK1/2 pathway, U0126, or inhibitors of the p38 MAPK pathway, SB203580 and SKF86002, repressed TGF-beta-induced Agc expression in a dose-dependent manner, indicating that ERK1/2 or p38 MAPK activation is also required for TGF-beta-induced Agc expression in confluent ATDC5 cells. In differentiated ATDC5 cells, persistently high basal levels of ERK1/2 and p38 MAPK phosphorylation correlated with elevated basal Agc expression, which was inhibited by incubation with inhibitors of these pathways. Whereas Smad2 was rapidly phosphorylated by TGF-beta and involved in the initial activation of Agc expression in confluent cells, Smad2 activation was not required for maintaining the high level of Agc expression. Taken together, these results suggest an important role for transcriptional cross-talk between Smad and MAPK pathways in expression of early chondrocytic phenotypes and identify important changes in the regulation of Agc expression following chondrocyte differentiation. PMID- 11278289 TI - Glutathione S-transferase mu modulates the stress-activated signals by suppressing apoptosis signal-regulating kinase 1. AB - Apoptosis signal-regulating kinase 1 (ASK1) is a mitogen-activated protein kinase kinase kinase that can activate the c-Jun N-terminal kinase and the p38 signaling pathways. It plays a critical role in cytokine- and stress-induced apoptosis. To further characterize the mechanism of the regulation of the ASK1 signal, we searched for ASK1-interacting proteins employing the yeast two-hybrid method. The yeast two-hybrid assay indicated that mouse glutathione S-transferase Mu 1-1 (mGSTM1-1), an enzyme involved in the metabolism of drugs and xenobiotics, interacted with ASK1. We subsequently confirmed that mGSTM1-1 physically associated with ASK1 both in vivo and in vitro. The in vitro binding assay indicated that the C-terminal portion of mGSTM1-1 and the N-terminal region of ASK1 were crucial for binding one another. Furthermore, mGSTM1-1 suppressed stress-stimulated ASK1 activity in cultured cells. mGSTM1-1 also blocked ASK1 oligomerization. The ASK1 inhibition by mGSTM1-1 occurred independently of the glutathione-conjugating activity of mGSTM1-1. Moreover, mGSTM1-1 repressed ASK1 dependent apoptotic cell death. Taken together, our findings suggest that mGSTM1 1 functions as an endogenous inhibitor of ASK1. This highlights a novel function for mGSTM1-1 insofar as mGSTM1-1 may modulate stress-mediated signals by repressing ASK1, and this activity occurs independently of its well-known catalytic activity in intracellular glutathione metabolism. PMID- 11278292 TI - Orphan receptor promiscuity in the induction of cytochromes p450 by xenobiotics. AB - The mechanisms by which different classes of chemicals induce the same cytochrome P450 (CYP) or the same chemical differentially induces more than one CYP are not well understood. We show that in primary hepatocytes and in vivo in liver (transfected by particle-mediated delivery) two orphan nuclear receptors, constitutive androstane receptor and pregnane X receptor (PXR1), transactivate a CYP gene via the same response element in a xenobiotic-specific manner. The constitutive androstane receptor mediates the barbiturate activation of expression of CYP2B1 and CYP3A1. PXR1 activates both genes in response to synthetic steroids. To exert their effect the receptors bind to the same direct repeat site (DR4) within the phenobarbital response element of the CYP2B1 promoter and to the same DR3 site in the pregnane X response element of CYP3A1. The receptors are therefore promiscuous with respect to DNA binding but not ligand binding. Differences in enhancer half-site spacing may influence the efficiency of interactions between the receptor and the transcription machinery and hence form the basis for the differential induction of CYP2B1 and CYP3A1 in response to barbiturates and synthetic steroids. PMID- 11278293 TI - Coiled coil region of streptokinase gamma-domain is essential for plasminogen activation. AB - The specific functions of the amino acid residues in the streptokinase (SK) gamma domain were analyzed by studying the interactions of human plasminogen (HPlg) and SK mutants prepared by charge-to-alanine mutagenesis. SK with mutations of groups of amino acids outside the coiled coil region of SK gamma-domain, SK(K278A,K279A,E281A,K282A), and SK(D360A,R363A) had similar HPlg activator activities as wild-type SK. However, significant changes of the functions of SK with mutations within the coiled coil region were observed. Both SK(D322A,R324A,D325A) and SK(R330A,D331A,K332A,K334A) had decreased amounts of complex formation with microplasminogen and failed to activate HPlg. SK(D328A,R330A) had a 21-fold reduced catalytic efficiency for HPlg activation. The studies of SK with single amino acid mutation to Ala demonstrate that Arg(324), Asp(325), Lys(332), and Lys(334) play important roles in the formation of a HPlg.SK complex. On the other hand, amino acid residues Asp(322), Asp(328), and Arg(330) of SK are involved in the virgin enzyme induction. Potential contact between Lys(332) of SK and Glu(623) of human microplasmin and strong interactions between Asp(328) and Lys(330), Asp(331) and Lys(334), and Asp(322) and Lys(334) of SK are noticed. These interactions are important in maintaining a coiled coil conformation. Therefore, we conclude that the coiled coil region of SK gamma domain, SK(Leu(314)-Ala(342)), plays very important roles in HPlg activation by participating in virgin enzyme induction and stabilizing the activator complex. PMID- 11278295 TI - Dantrolene inhibition of ryanodine receptor Ca2+ release channels. Molecular mechanism and isoform selectivity. AB - As an inhibitor of Ca(2+) release through ryanodine receptor (RYR) channels, the skeletal muscle relaxant dantrolene has proven to be both a valuable experimental probe of intracellular Ca(2+) signaling and a lifesaving treatment for the pharmacogenetic disorder malignant hyperthermia. However, the molecular basis and specificity of the actions of dantrolene on RYR channels have remained in question. Here we utilize [(3)H]ryanodine binding to further investigate the actions of dantrolene on the three mammalian RYR isoforms. The inhibition of the pig skeletal muscle RYR1 by dantrolene (10 microm) was associated with a 3-fold increase in the K(d) of [(3)H]ryanodine binding to sarcoplasmic reticulum (SR) vesicles such that dantrolene effectively reversed the 3-fold decrease in the K(d) for [(3)H]ryanodine binding resulting from the malignant hyperthermia RYR1 Arg(615) --> Cys mutation. Dantrolene inhibition of the RYR1 was dependent on the presence of the adenine nucleotide and calmodulin and reflected a selective decrease in the apparent affinity of RYR1 activation sites for Ca(2+) relative to Mg(2+). In contrast to the RYR1 isoform, the cardiac RYR2 isoform was unaffected by dantrolene, both in native cardiac SR vesicles and when heterologously expressed in HEK-293 cells. By comparison, the RYR3 isoform expressed in HEK-293 cells was significantly inhibited by dantrolene, and the extent of RYR3 inhibition was similar to that displayed by the RYR1 in native SR vesicles. Our results thus indicate that both the RYR1 and the RYR3, but not the RYR2, may be targets for dantrolene inhibition in vivo. PMID- 11278294 TI - Involvement of the acid sphingomyelinase pathway in uva-induced apoptosis. AB - The sphingomyelin-ceramide pathway is an evolutionarily conserved ubiquitous signal transduction system that regulates many cell functions including apoptosis. Sphingomyelin (SM) is hydrolyzed to ceramide by different sphingomyelinases. Ceramide serves as a second messenger in mediating cellular effects of cytokines and stress. In this study, we find that acid sphingomyelinase (SMase) activity was induced by UVA in normal JY lymphoblasts but was not detectable in MS1418 lymphoblasts from Niemann-Pick type D patients who have an inherited deficiency of acid SMase. We also provide evidence that UVA can induce apoptosis by activating acid SMase in normal JY cells. In contrast, UVA-induced apoptosis was inhibited in MS1418 cells. Exogenous SMase and its product, ceramide (10-40 micrometer), induced apoptosis in JY and MS1418 cells, but the substrate of SMase, SM (20-80 micrometer), induced apoptosis only in JY cells. These results suggest that UVA-induced apoptosis by SM is dependent on acid SMase activity. We also provide evidence that induction of apoptosis by UVA may occur through activation of JNKs via the acid SMase pathway. PMID- 11278296 TI - Anti-CD3 and concanavalin A-induced human T cell proliferation is associated with an increased rate of arachidonate-phospholipid remodeling. Lack of involvement of group IV and group VI phospholipase A2 in remodeling and increased susceptibility of proliferating T cells to CoA-independent transacyclase inhibitor-induced apoptosis. AB - In this study arachidonate-phospholipid remodeling was investigated in resting and proliferating human T lymphocytes. Lymphocytes induced to proliferate with either the mitogen concanavalin A or with anti-CD3 (OKT3) in combination with interleukin 2 (OKT3/IL-2) showed a greatly accelerated rate of [3H]arachidonate phospholipid remodeling compared with resting lymphocytes or with lymphocytes stimulated with OKT3 or IL-2 alone. The concanavalin A-stimulated cells showed a 2-fold increase in the specific activity of CoA-independent transacylase compared with unstimulated cells, indicating that this enzyme is inducible. Stimulation with OKT3 resulted in greatly increased quantities of the group VI calcium independent phospholipase A2 but not of the quantity of group IV cytosolic phospholipase A2. However, group IV phospholipase A2 became phosphorylated in OKT3-stimulated cells, as determined by decreased electrophoretic mobility. Incubation of cells with the group VI phospholipase A2 inhibitor, bromoenol lactone, or the dual group IV/group VI phospholipase A2 inhibitor, methyl arachidonyl fluorophosphonate, did not block arachidonate-phospholipid remodeling resting or proliferating T cells, suggesting that these phospholipases A2 were not involved in arachidonate-phospholipid remodeling. The incubation of nonproliferating human lymphocytes with inhibitors of CoA-independent transacylase had little impact on cell survival. In contrast, OKT3/IL-2 stimulated T lymphocytes were very sensitive to apoptosis induced by CoA independent transacylase inhibitors. Altogether these results indicate that increased arachidonate-phospholipid remodeling is associated with T cell proliferation and that CoA-independent transacylase may be a novel therapeutic target for proliferative disorders. PMID- 11278297 TI - Glyceroneogenesis and the source of glycerol for hepatic triacylglycerol synthesis in humans. AB - Glyceroneogenesis, i.e. the synthesis of the glycerol moiety of triacylglycerol from pyruvate, has been suggested to be quantitatively important in both the liver and adipose tissue during fasting. However, the actual contribution of glyceroneogenesis to triacylglycerol synthesis has not been quantified in vivo in human studies. In the present study we have measured the contribution of glycerol and pyruvate to in vivo synthesis of hepatic triacylglycerol in nonpregnant and pregnant women after an overnight fast. Five nonpregnant women were administered [(13)C(3)]glycerol tracer as prime constant rate infusion, and the appearance of tracer in plasma glucose and triacylglycerol was quantified using gas chromatography-mass spectrometry. The contribution of pyruvate to hepatic triacylglycerol was quantified in nonpregnant and pregnant women using the deuterium labeling of body water method. The appearance of [(2)H] in hydrogens on C(1) and C(3) of triacylglycerol was measured following periodate oxidation of the glycerol isolated from hydrolyzed triacylglycerol. After a 16-h fast, approximately 6.1% of the plasma triacylglycerol pool was derived from plasma glycerol, whereas 10 to 60% was derived from pyruvate in nonpregnant women and pregnant women early in gestation. Our data suggest that glyceroneogenesis from pyruvate is quantitatively a major contributor to plasma triacylglycerol synthesis and may be important for the regulation of very low density lipoprotein triacylglycerol production. Our data also suggest that 3-glycerol phosphate is in rapid equilibrium with the triosephosphate pool, resulting in rapid labeling of the triose pool by the administered tracer glycerol. Because the rate of flux of triosephosphate to glucose during fasting far exceeds that to triacylglycerol, more glycerol ends up in glucose than in triacylglycerol. Alternatively, there may be two distinct pools of 3-glycerol phosphate in the liver, one involved in generating triosephosphate from glycerol and the other involved in glyceride glycerol synthesis. PMID- 11278298 TI - Phosphorylation and regulation of G-protein-activated phospholipase C-beta 3 by cGMP-dependent protein kinases. AB - Among the drugs that are known to relax the vascular smooth muscle and regulate other cellular functions, beta-adrenergic agonists and nitric oxide-containing compounds are some of the most effective ones. The mechanisms of these drugs are thought to lower agonist-induced intracellular [Ca(2+)] by increasing intracellular cAMP and cGMP, activating their respective protein kinases. However, the physiological targets of cyclic nucleotide-dependent protein kinases are not clear. The molecular basis for the regulation of intracellular Ca(2+) by signaling pathways coupled to cyclic nucleotides is not well defined. G-protein activated phospholipase C (PLC-beta) catalyzes the hydrolysis of phosphatidylinositol 4,5-bisphosphates to generate diacylglycerol and inositol 1,4,5-triphosphate, leading to the activation of protein kinase C and the mobilization of intracellular Ca(2+). In this study, we shown that G-protein activated PLC enzymes are the potential targets of cGMP-dependent protein kinases (PKG). PKG can directly phosphorylate PLC-beta2 and PLC-beta3 in vitro with purified proteins and in vivo with metabolic labeling. Phosphorylation of PLC beta leads to the inhibition of G-protein-activated PLC-beta3 activity by 50-70% in COS-7 cell transfection assays. By using phosphopeptide mapping and site directed mutagenesis, we further identified two key phosphorylation sites for the regulation of PLC-beta3 by PKG (Ser(26) and Ser(1105)). Mutation at these two sites (S26A and S1105A) of PLC-beta3 completely blocked the phosphorylation of PLC-beta3 protein catalyzed by PKG. Furthermore, mutation of these serine residues removed the inhibitory effect of PKG on the activation of the mutant PLC beta3 proteins by G-protein subunits. Our results suggest a molecular mechanism for the regulation of G-protein-mediated intracellular [Ca(2+)] by the NO-cGMP dependent signaling pathway. PMID- 11278299 TI - Amyloid precursor protein and amyloid beta peptide in human platelets. Role of cyclooxygenase and protein kinase C. AB - The main component of Alzheimer's disease (AD) senile plaques is amyloid-beta peptide (Abeta), a proteolytic fragment of the amyloid precursor protein (APP). Platelets contain both APP and Abeta and may contribute to the perivascular amyloid deposition seen in AD. However, no data are available concerning the biochemical mechanism(s) involved in their formation and release by these cells. We found that human platelets released APP and Abeta following activation with collagen or arachidonic acid. Inhibition of platelet cyclooxygenase (COX) reduced APP but not Abeta release following those stimuli. In contrast, activation of platelets by thrombin and calcium ionophore caused release of both APP and Abeta in a COX-independent fashion. Ex vivo studies showed that, despite suppression of COX activity, administration of aspirin did not modify Abeta or APP levels in serum or plasma, suggesting that this enzyme plays only a minor role in vivo. We examined the regulation of APP cleavage and release from activated platelets and found that cleavage requires protein kinase C (PKC) activity and is regulated by the intracellular second messengers phosphatidylinositol 2-phosphate and Ca(2+). Our data provide the first evidence that in human platelets COX is a minor component of APP secretion whereas PKC plays a major role in the secretory cleavage of APP. By contrast, Abeta release may represent secretion of preformed peptide and is totally independent of both COX and PKC activity. PMID- 11278300 TI - Identification of human macrophage inflammatory proteins 1alpha and 1beta as a native secreted heterodimer. AB - Chemokines are secreted proteins that function as chemoattractants for leukocytes. The chemokines macrophage inflammatory protein 1alpha and 1beta (MIP 1alpha and MIP-1beta) now have been shown to be secreted from activated human monocytes and peripheral blood lymphocytes (PBLs) as a heterodimer. Immunoprecipitation and immunoblot analysis revealed that antibodies to either MIP-1alpha or MIP-1beta precipitated a protein complex containing both MIP-1alpha and MIP-1beta under normal conditions from culture supernatants and lysates of these cells. Mass spectrometry of the complexes, precipitated from the culture supernatants of monocytes and PBLs, revealed the presence of NH(2)-terminal truncated MIP-1alpha (residues 5-70) together with either intact MIP-1beta or NH(2)-terminal truncated MIP-1beta (residues 3-69), respectively. The secreted MIP-1alpha/beta heterodimers were dissociated into their component monomers under acidic conditions. Exposure of monocytes or PBLs to monensin induced the accumulation of heterodimers composed of NH(2)-terminal truncated MIP-1alpha and full-length MIP-1beta in the Golgi complex. The mixing of recombinant chemokines in vitro demonstrated that heterodimerization of MIP-1alpha and MIP-1beta is specific and that it occurs at physiological conditions, pH 7.4, and in the range of nanomolar concentrations. The data presented here provide the first biochemical evidence for the existence of chemokine heterodimers under natural conditions. Formation of heterodimers of MIP-1alpha/beta may have an impact on intracellular signaling events that contribute to CCR5 and possibly to other chemokine receptor functions. PMID- 11278301 TI - Ultraviolet radiation inhibits interleukin-2-induced tyrosine phosphorylation and the activation of STAT5 in T lymphocytes. AB - UV radiation was recently found to hinder interferon-gamma from exerting its biological effects by inhibiting the phosphorylation of signal transducer and activator of transcription (STAT)-1, a crucial signal transducing protein in the interferon-gamma pathway. Because this activity by UV may contribute to its immunosuppressive properties we studied whether this is specific for STAT1 or whether UV also affects other members of the STAT family. STAT5 is crucially involved in signaling of interleukin (IL)-2, enabling up-regulation of the IL-2 receptor alpha chain, an essential component of the high affinity IL-2 receptor. Exposure of the murine T cell line CTLL to IL-2 caused tyrosine phosphorylation of STAT5 that was remarkably reduced when cells were exposed to UV. Accordingly, STAT5 binding activity was significantly impaired in UV-exposed cells. In contrast, IL-2-induced tyrosine phosphorylation of the kinases Jak1 and Jak3 located upstream of STAT5 was not affected by UV. The effect of UV on STAT5 phosphorylation was antagonized by orthovanadate, implying involvement of a phosphatase in this process. Accordingly, up-regulation of the IL-2 receptor alpha chain was reduced in cells that were treated with IL-2 plus UV. Because STAT5-mediated IL-2 effects are vital for normal immune functions, inhibition of STAT5 signaling by UV may contribute to its well known immunosuppressive properties. PMID- 11278303 TI - Matrix regulation of skeletal cell apoptosis. Role of calcium and phosphate ions. AB - Previously, we noted that inorganic phosphate (P(i)), a major component of bone extracellular matrix, induced osteoblast apoptosis (Meleti, Z., Shapiro, I. M., and Adams, C. S. (2000) Bone (NY) 27, 359-366). Since Ca(2+) along with P(i) is released from bone during the resorption process, we advanced the hypothesis that Ca(2+) modulates P(i)-mediated osteoblast apoptosis. To test this hypothesis, osteoblasts were incubated with both ions, and cell death was determined. We noted that a modest increase in the medium Ca(2+) concentrations ([Ca(2+)](e)) of 0.1-1 mm caused a profound and rapid enhancement in P(i)-dependent death of cultured osteoblasts. An elevation in [Ca(2+)](e) alone had no effect on osteoblast viability, whereas Ca(2+) channel blockers failed to inhibit killing of ion pair-treated cells. These results indicated that P(i)-mediated cell death is not dependent on a sustained increase in the cytosolic Ca(2+) concentration. Terminal dUTP nick-end labeling analysis and measurement of caspase-3 activity of the ion pair-treated cells suggested that death was apoptotic. Apoptosis was confirmed using caspase-3 and endonuclease inhibitors. The mitochondrial membrane potential and cytosolic Ca(2+) status of the treated cells were evaluated. After incubation with [Ca(2+) ](e) and P(i), a decrease in mitochondrial fluorescence was noted, suggesting that the ions decreased the mitochondrial transmembrane potential. Subsequent to the fall in mitochondrial membrane potential, there was a transient elevation in the cytosolic Ca(2+) concentration. Results of the study suggest that the ion pair conspire at the level of the plasma membrane to induce intracellular changes that result in loss of mitochondrial function. The subsequent increase in the cytosolic Ca(2+) concentration may trigger downstream events that transduce osteoblast apoptosis. PMID- 11278302 TI - Transforming growth factor-beta receptor-associated protein 1 is a Smad4 chaperone. AB - Members of the transforming growth factor-beta (TGF-beta) superfamily signal through unique cell membrane receptor serine-threonine kinases to activate downstream targets. TRAP1 is a previously described 96-kDa cytoplasmic protein shown to bind to TGF-beta receptors and suggested to play a role in TGF-beta signaling. We now fully characterize the binding properties of TRAP1, and show that it associates strongly with inactive heteromeric TGF-beta and activin receptor complexes and is released upon activation of signaling. Moreover, we demonstrate that TRAP1 plays a role in the Smad-mediated signal transduction pathway, interacting with the common mediator, Smad4, in a ligand-dependent fashion. While TRAP1 has only a small stimulatory effect on TGF-beta signaling in functional assays, deletion constructs of TRAP1 inhibit TGF-beta signaling and diminish the interaction of Smad4 with Smad2. These are the first data to identify a specific molecular chaperone for Smad4, suggesting a model in which TRAP1 brings Smad4 into the vicinity of the receptor complex and facilitates its transfer to the receptor-activated Smad proteins. PMID- 11278304 TI - Presence and activation of nuclear phosphoinositide 3-kinase C2beta during compensatory liver growth. AB - Highly purified liver nuclei incorporated radiolabeled phosphate into phosphatidylinositol 4-phosphate (PtdIns(4)P), PtdIns(4,5)P(2), and PtdIns(3,4,5)P(3). When nuclei were depleted of their membrane, no radiolabeling of PtdIns(3,4,5)P(3) could be detected showing that within the intranuclear region there are no class I phosphoinositide 3-kinases (PI3K)s. In membrane depleted nuclei harvested 20 h after partial hepatectomy, the incorporation of radiolabel into PtdIns(3)P was observed together with an increase in immunoprecipitable PI3K-C2beta activity, which is sensitive to wortmannin (10 nm) and shows strong preference for PtdIns over PtdIns(4)P as a substrate. On Western blots PI3K-C2beta revealed a single immunoreactive band of 180 kDa, whereas 20 h after partial hepatectomy gel shift of 18 kDa was noticed, suggesting that observed activation of enzyme is achieved by proteolysis. When intact membrane depleted nuclei were subjected to short term (20 min) exposure to micro-calpain, similar gel shift together with an increase in PI3K-C2beta activity was observed, when compared with the nuclei harvested 20 h after partial hepatectomy. Moreover, the above-mentioned gel shift and increase in PI3K-C2beta activity could be prevented by the calpain inhibitor calpeptin. The data presented in this report show that, in the membrane-depleted nuclei during the compensatory liver growth, there is an increase in PtdIns(3)P formation as a result of PI3K-C2beta activation, which may be a calpain-mediated event. PMID- 11278305 TI - A G1127S change in calcium-binding epidermal growth factor-like domain 13 of human fibrillin-1 causes short range conformational effects. AB - Human fibrillin-1, an extracellular matrix glycoprotein, has a modular organization that includes 43 calcium-binding epidermal growth factor-like (cbEGF) domains arranged as multiple tandem repeats. A missense mutation that changes a highly conserved glycine to serine (G1127S) has been identified in cbEGF13, which results in a variant of Marfan syndrome, a connective tissue disease. Previous experiments on isolated cbEGF13 and a cbEGF13-14 pair indicated that the G1127S mutation caused defective folding of cbEGF13 but not cbEGF14. We have used limited proteolysis methods and two-dimensional NMR spectroscopy to identify the structural consequences of this mutation in a covalently linked cbEGF12-13 pair and a cbEGF12-14 triple domain construct. Protease digestion studies of the cbEGF12-13 G1127S mutant pair indicated that both cbEGF12 and 13 retained similar calcium binding properties and thus tertiary structure to the normal domain pair, because all identified cleavage sites showed calcium dependent protection from proteolysis. However, small changes in the conformation of cbEGF13 G1127S, revealed by the presence of a new protease-sensitive site and comparative two-dimensional NOESY data, suggested that the fold of the mutant domain was not identical to the wild-type, but was native-like. Additional cleavage sites identified in cbEGF12-14 G1127S indicated further subtle changes within the mutant domain but not the flanking domains. We have concluded the following in this study. (i) Covalent linkage of cbEGF12 preserves the native like fold of cbEGF13 G1127S and (ii) conformational effects introduced by G1127S are localized to cbEGF13. This study demonstrates that missense mutations in fibrillin-1 cbEGF domains can cause short range structural effects in addition to long range effects previously observed with a E1073K mutation in cbEGF12. PMID- 11278306 TI - Prion protein binds copper within the physiological concentration range. AB - The prion protein is known to be a copper-binding protein, but affinity and stoichiometry data for the full-length protein at a physiological pH of 7 were lacking. Furthermore, it was unknown whether only the highly flexible N-terminal segment with its octarepeat region is involved in copper binding or whether the structured C-terminal domain is also involved. Therefore we systematically investigated the stoichiometry and affinity of copper binding to full-length prion protein PrP(23-231) and to different N- and C-terminal fragments using electrospray ionization mass spectrometry and fluorescence spectroscopy. Our data indicate that the unstructured N-terminal segment is the cooperative copper binding domain of the prion protein. The prion protein binds up to five copper(II) ions with half-maximal binding at approximately 2 microm. This argues strongly for a direct role of the prion protein in copper metabolism, since it is almost saturated at about 5 microm, and the exchangeable copper pool concentration in blood is about 8 microm. PMID- 11278307 TI - G-protein-coupled receptor stimulation of the p42/p44 mitogen-activated protein kinase pathway is attenuated by lipid phosphate phosphatases 1, 1a, and 2 in human embryonic kidney 293 cells. AB - Sphingosine 1-phosphate, lysophosphatidic acid, and phosphatidic acid bind to G protein-coupled receptors to stimulate intracellular signaling in mammalian cells. Lipid phosphate phosphatases (1, 1a, 2, and 3) are a group of enzymes that catalyze de-phosphorylation of these lipid agonists. It has been proposed that the lipid phosphate phosphatases exhibit ecto activity that may function to limit bioavailability of these lipid agonists at their receptors. In this study, we show that the stimulation of the p42/p44 mitogen-activated protein kinase pathway by sphingosine 1-phosphate, lysophosphatidic acid, and phosphatidic acid, all of which bind to G(i/o)-coupled receptors, is substantially reduced in human embyronic kidney 293 cells transfected with lipid phosphate phosphatases 1, 1a, and 2 but not 3. This was correlated with reduced basal intracellular phosphatidic acid and not ecto lipid phosphate phosphatase activity. These findings were supported by results showing that lipid phosphate phosphatases 1, 1a, and 2 also abrogate the stimulation of p42/p44 mitogen-activated protein kinase by thrombin, a peptide G(i/o)-coupled receptor agonist whose bioavailability at its receptor is not subject to regulation by the phosphatases. Furthermore, the lipid phosphate phosphatases have no effect on the stimulation of p42/p44 mitogen-activated protein kinase by other agents that do not use G proteins to signal, such as serum factors and phorbol ester. Therefore, these findings show that the lipid phosphate phosphatases 1, 1a, and 2 may function to perturb G-protein-coupled receptor signaling per se rather than limiting bioavailability of lipid agonists at their respective receptors. PMID- 11278308 TI - Anesthetic-like interactions of nitric oxide with albumin and hemeproteins. A mechanism for control of protein function. AB - Noncovalent bonding interactions of nitric oxide (NO) with human serum albumin (HSA), human hemoglobin A, bovine myoglobin, and bovine cytochrome c oxidase (CcO) have been explored. The anesthetic nitrous oxide (NNO) occupies multiple sites within each protein, but does not bind to heme iron. Infrared (IR) spectra of NNO molecules sequestered within albumin, with NO present, support the binding of NO and NNO to the same sites with comparable affinities. Perturbations of IR spectra of the Cys(34) thiol of HSA indicate NO, NNO, halothane, and chloroform can induce similar changes in protein structure. Experiments evaluating the relative affinities of binding of NO and carbon monoxide (CO) to iron(II) sites of the hemeproteins led to evidence of NO binding to noniron, nonsulfur sites as well. With HbA, IR spectra of cysteine thiols and/or the iron(II) N-O stretching region denote changes in protein structure due to NO, NNO, or CO occupying noniron sites with an order of decreasing affinities of NO > NNO > CO. Loss of NO from some, not all, noniron sites in hemeproteins is very slow (t(1/2) approximately hours). These findings provide examples in which NO and anesthetics alter the structure and properties of protein similarly, and support the hypothesis that some physiological effects of NO (and possibly CO) result from anesthetic-like noncovalent bonding to sites within protein or other tissue components. Such bonding may be involved in mechanisms for control of oxygen transport, mitochondrial respiration, and activation of soluble guanylate cyclase by NO. PMID- 11278309 TI - Caveolin-1 interacts with androgen receptor. A positive modulator of androgen receptor mediated transactivation. AB - Androgen receptor (AR) belongs to the steroid hormone nuclear receptor superfamily. It functions as an androgen-dependent transcriptional factor that regulates genes for cell proliferation and differentiation. Caveolin is a principal component of caveolae membranes serving as a scaffold protein of many signal transduction pathways. Recent results correlate caveolin-1 expression with androgen sensitivity in murine prostate cancer. Furthermore, immunohistochemical staining of patient specimens suggests that caveolin expression may be an independent predictor of progression of prostate cancer. In this study, we investigate the potential interactions between AR signaling and caveolin-1 and demonstrate that overexpression of caveolin-1 potentiates ligand-dependent AR activation. Conversely, down-regulation of caveolin-1 expression by a caveolin-1 antisense expression construct can down-regulate ligand-dependent AR activation. Association between these two molecules is also demonstrated by co-localization of AR with caveolin-rich, low-density membrane fractions isolated by an equilibrium sucrose gradient centrifugation method. Co-immunoprecipitation and glutathione S-transferase fusion protein pull-down experiments demonstrate that interaction between AR and caveolin-1 is an androgen-dependent process, offering further evidence for a physiological role of this interaction. Using a mammalian two-hybrid assay system, we determine that the NH(2) terminus region of caveolin 1 is responsible for the interaction with both the NH(2)-terminal domain and the ligand-binding domain of AR. PMID- 11278310 TI - Adenosine nucleotides acting at the human P2Y1 receptor stimulate mitogen activated protein kinases and induce apoptosis. AB - For the widely distributed P2Y receptors for nucleotides, the transductional and functional responses downstream of their coupling to G proteins are poorly characterized. Here we describe apoptotic induction and the associated differential stimulation of mitogen-activated protein (MAP) kinase family members by the human P2Y(1) receptor. The potent P2Y(1) receptor agonist, 2-methylthio ADP (2-MeSADP), stimulated the extracellular-signal regulated kinases (ERK1/2) (EC(50) approximately 5 nm) as well as several, but not all isoforms detected, of the stress-activated protein kinase (SAPK) family. Phospho-isoforms of p38 were unaffected. The induced kinase activity was blocked by the P2Y(1) receptor selective antagonist, adenosine-2'-phosphate-5'-phosphate, but unaffected by pertussis toxin. In addition, the endogenous ligand ADP, and significantly also 2 MeSATP, induced concentration-dependent phosphorylation changes in the same MAP kinase family members. The sustained activation of ERK1/2 was associated with Elk 1 phosphorylation that was abolished by the MEK1 inhibitor, PD 98059. However, the concomitant transient activation of the SAPKs was not sufficient to induce c Jun or ATF-2 phosphorylation. The transient phase of the ERK activity was partially inhibited either by the phosphatidylinositol 3-kinase inhibitor, LY 294002, or the PKC inhibitor, Go 6976. In addition, the Src inhibitor, PP1, or expression of dominant negative Ras also attenuated the transient phase of ERK phosphorylation. In contrast, inhibition of Ras or Src had no effect on the sustained ERK activity, which was critically dependent on phosphatidylinositol 3 kinase. The transient SAPK activity was suppressed by expression of a dominant negative form of MKK4. Furthermore, this kinase-deficient mutant inhibited 2 MeSADP-induced caspase-3 stimulation and the associated decrease in cell number. In conclusion, adenosine di- and triphosphate stimulation of the human P2Y(1) receptor can transiently activate the Ras-ERK cascade via the cooperative effects of phosphatidylinositol 3-kinase, Src and PKC. The sustained ERK stimulation, via a Ras-insensitive pathway, culminates in Elk-1 activation without inducing a proliferation effect. The transient SAPK activity did not evoke transcription factor phosphorylation but was required for the P2Y(1) receptor-mediated apoptotic function. PMID- 11278311 TI - Importance of the P4' residue in human granzyme B inhibitors and substrates revealed by scanning mutagenesis of the proteinase inhibitor 9 reactive center loop. AB - The cytotoxic lymphocyte serine proteinase granzyme B induces apoptosis of abnormal cells by cleaving intracellular proteins at sites similar to those cleaved by caspases. Understanding the substrate specificity of granzyme B will help to identify natural targets and develop better inhibitors or substrates. Here we have used the interaction of human granzyme B with a cognate serpin, proteinase inhibitor 9 (PI-9), to examine its substrate sequence requirements. Cleavage and sequencing experiments demonstrated that Glu(340) is the P1 residue in the PI-9 RCL, consistent with the preference of granzyme B for acidic P1 residues. Ala-scanning mutagenesis demonstrated that the P4-P4' region of the PI 9 RCL is important for interaction with granzyme B, and that the P4' residue (Glu(344)) is required for efficient serpin-proteinase binding. Peptide substrates based on the P4-P4' PI-9 RCL sequence and containing either P1 Glu or P1 Asp were cleaved by granzyme B (k(cat)/K(m) 9.5 x 10(3) and 1.2 x 10(5) s(-1) M(-1), respectively) but were not recognized by caspases. A substrate containing P1 Asp but lacking P4' Glu was cleaved less efficiently (k(cat)/K(m) 5.3 x 10(4) s(-1) M(-1)). An idealized substrate comprising the previously described optimal P4-P1 sequence (Ile-Glu-Pro-Asp) fused to the PI-9 P1'-P4' sequence was efficiently cleaved by granzyme B (k(cat)/K(m) 7.5 x 10(5) s(-1) M(-1)) and was also recognized by caspases. This contrasts with the literature value for a tetrapeptide comprising the same P4-P1 sequence (k(cat)/K(m) 6.7 x 10(4) s(-1) M( 1)) and confirms that P' residues promote efficient interaction of granzyme B with substrates. Finally, molecular modeling predicted that PI-9 Glu(344) forms a salt bridge with Lys(27) of granzyme B, and we showed that a K27A mutant of granzyme B binds less efficiently to PI-9 and to substrates containing a P4' Glu. We conclude that granzyme B requires an extended substrate sequence for specific and efficient binding and propose that an acidic P4' substrate residue allows discrimination between early (high affinity) and late (lower affinity) targets during the induction of apoptosis. PMID- 11278312 TI - Hepatitis C virus core protein activates nuclear factor kappa B-dependent signaling through tumor necrosis factor receptor-associated factor. AB - Hepatitis C virus (HCV) core protein, a viral nucleocapsid, has been shown to affect various intracellular events including the nuclear factor kappaB (NF kappaB) signaling supposedly associated with inflammatory response, cell proliferation, and apoptosis. In order to elucidate the effect of HCV core protein on the NF-kappaB signaling in HeLa and HepG2 cells, a reporter assay was utilized. HCV core protein significantly activated NF-kappaB signaling in a dose dependent manner not only in HeLa and HepG2 cells transiently transfected with core protein expression plasmid, but also in HeLa cells induced to express core protein under the control of doxycycline. HCV core protein increased the DNA binding affinity of NF-kappaB in the electrophoretic mobility shift assay. Acetyl salicylic acid, an IKKbeta-specific inhibitor, and dominant negative form of IKKbeta significantly blocked NF-kappaB activation by HCV core protein, suggesting HCV core protein activates the NF-kappaB pathway mainly through IKKbeta. Moreover, the dominant negative forms of TRAF2/6 significantly blocked activation of the pathway by HCV core protein, suggesting HCV core protein mimics proinflammatory cytokine activation of the NF-kappaB pathway through TRAF2/6. In fact, HCV core protein activated interleukin-1beta promoter mainly through NF kappaB pathway. Therefore, this function of HCV core protein may play an important role in the inflammatory reaction induced by this hepatotropic virus. PMID- 11278313 TI - Palmitoylation of caveolin-1 in endothelial cells is post-translational but irreversible. AB - Caveolin-1 is a palmitoylated protein involved in assembly of signaling molecules in plasma membrane subdomains termed caveolae and in intracellular cholesterol transport. Three cysteine residues in the C terminus of caveolin-1 are subject to palmitoylation, which is not necessary for caveolar targeting of caveolin-1. Protein palmitoylation is a post-translational and reversible modification that may be regulated and that in turn may regulate conformation, membrane association, protein-protein interactions, and intracellular localization of the target protein. We have undertaken a detailed analysis of [(3)H]palmitate incorporation into caveolin-1 in aortic endothelial cells. The linkage of palmitate to caveolin-1 was hydroxylamine-sensitive and thus presumably a thioester bond. However, contrary to expectations, palmitate incorporation was blocked completely by the protein synthesis inhibitors cycloheximide and puromycin. In parallel experiments to show specificity, palmitoylation of aortic endothelial cell-specific nitric-oxide synthase was unaffected by these reagents. Inhibitors of protein trafficking, brefeldin A and monensin, blocked caveolin-1 palmitoylation, indicating that the modification was not cotranslational but rather required caveolin-1 transport from the endoplasmic reticulum and Golgi to the plasma membrane. In addition, immunophilin chaperones that form complexes with caveolin-1, i.e. FK506-binding protein 52, cyclophilin A, and cyclophilin 40, were not necessary for caveolin-1 palmitoylation because agents that bind immunophilins did not inhibit palmitoylation. Pulse-chase experiments showed that caveolin-1 palmitoylation is essentially irreversible because the release of [(3)H]palmitate was not significant even after 24 h. These results show that [(3)H]palmitate incorporation is limited to newly synthesized caveolin-1, not because incorporation only occurs during synthesis but because the continuous presence of palmitate on caveolin-1 prevents subsequent repalmitoylation. PMID- 11278314 TI - Collectrin, a collecting duct-specific transmembrane glycoprotein, is a novel homolog of ACE2 and is developmentally regulated in embryonic kidneys. AB - Collectrin, a novel homolog of angiotensin-converting enzyme-related carboxypeptidase (ACE2), was identified during polymerase chain reaction-based cDNA subtraction and up-regulated in 5/6 ablated kidneys at hypertrophic phase. Collectrin, with 222 amino acids, has an apparent signal peptide and a transmembrane domain; the sequence is conserved in mouse, rat, and human and shares 81.9% identity. Human collectrin has 47.8% identity with non-catalytic extracellular, transmembrane, and cytosolic domains of ACE2; however, unlike ACE and ACE2, collectrin lacks active dipeptidyl carboxypeptidase catalytic domains. The collectrin mRNA transcripts are expressed exclusively in the kidney. In situ hybridization reveals its mRNA expression in renal collecting ducts, and immunohistochemistry shows that it is localized to the luminal surface and cytoplasm of collecting ducts. Immunoprecipitation studies, using [35S]methionine labeled renal cortical and inner medullar collecting duct cells, i.e. M-1 and mIMCD-3, indicate that the protein size is approximately 32 kDa. During the development of mouse kidney, mRNA signal is detectable at day 13 of gestation, and the protein product is observed in the ureteric bud branches. Its expression is progressively increased during later stages of the gestation extending into the neonatal periods and then is decreased in adult life. Up-regulated expression of collectrin in the hypertrophic kidneys after renal ablation and restricted spatio-temporal expression during development indicates a possible role(s)in the process of progressive renal failure and renal organogenesis. PMID- 11278315 TI - Activation of human prothrombin by arginine-specific cysteine proteinases (Gingipains R) from porphyromonas gingivalis. AB - The effect of 95- (HRgpA) and 50-kDa gingipain R (RgpB), arginine-specific cysteine proteinases from periodontopathogenic bacterium Porphyromonas gingivalis on human prothrombin activation was investigated. Each enzyme released thrombin from prothrombin in a dose- and time-dependent manner with the former enzyme, containing adhesion domains, being 17-fold more efficient than the single chain RgpB. A close correlation between the generation of fibrinogen clotting activity and amidolytic activity indicated that alpha-thrombin was produced by gingipains R, and this was confirmed by SDS-polyacrylamide gel electrophoresis, thrombin active site labeling, and amino-terminal sequence analysis of prothrombin digestion fragments. Significantly, the catalytic efficiency of HRgpA to generate thrombin (k(cat)/K(m) = 1.2 x 10(6) m(-)1 s(-)1) was 100-fold higher than that of RgpB (k(cat)/K(m) = 1.2 x 10(4) m(-)1 s(-)1). The superior prothrombinase activity of HRgpA over RgpB correlates with the fact that only the former enzyme was able to clot plasma, and kinetic data indicate that prothrombin activation can occur in vivo. At P. gingivalis-infected periodontitis sites HRgpA may be involved in the direct production of thrombin and, therefore, in the generation of prostaglandins and interleukin-1, both have been found to be associated with the development and progression of the disease. Furthermore, by taking into account that the P. gingivalis bacterium has been immunolocalized in carotid atherosclerotic plaques at thrombus formation sites (Chiu, B. (1999) Am. Heart J. 138, S534-S536), our results indicate that bacterial proteinases may potentially participate in the pathogenesis of cardiovascular disease associated with periodontitis. PMID- 11278316 TI - Unique biochemical nature of carp retinol-binding protein. N-linked glycosylation and uncleavable NH2-terminal signal peptide. AB - Retinol transport and metabolism have been well characterized in mammals; however, very little is known in fish. To study the mechanism by which fish retinol-binding protein (RBP) is able to remain in plasma besides its small molecular size, we isolated RBP cDNA from a carp liver cDNA library. Comparison of the deduced amino acid sequence with that of known vertebrate RBPs showed that carp RBP has high homology to the other cloned vertebrate RBPs, but it lacks the COOH-terminal tetrapeptide, RNL(S)L, which is most likely involved in the interaction with transthyretin in mammalian RBPs. In addition, the primary structure of carp RBP contains two consensus N-linked glycosylation sites that represent a unique feature. We have obtained experimental evidence, by in vitro and in vivo expression experiments, that both sites are indeed glycosylated. We have also characterized the protein as a complex type N-linked glycoprotein by lectin binding assay, neuraminidase and endoglycosidase H and F digestion. Inhibition of glycosylation by tunicamycin treatment of transfected cells caused a great reduction of RBP secretion. Since kidney filtration of anionic proteins is less than half that of neutral protein of the same size, this finding strongly suggests that the amount of carp RBP filtration through kidney glomeruli may be reduced by a glycosylation-dependent increase in the molecular size and negative charge of the protein. A second unique feature of carp RBP as secretory protein is the presence of a nonconserved NH(2)-terminal hydrophobic domain, which functions as an insertion signal but is not cleaved cotranslationally and remains in the secreted RBP. PMID- 11278317 TI - The human tumor suppressor arf interacts with spinophilin/neurabin II, a type 1 protein-phosphatase-binding protein. AB - The INK4a gene, one of the most often disrupted loci in human cancer, encodes two unrelated proteins, p16(INK4a) and p14(ARF) (ARF) both capable of inducing cell cycle arrest. Although it has been clearly demonstrated that ARF inhibits cell cycle via p53 stabilization, very little is known about the involvement of ARF in other cell cycle regulatory pathways, as well as on the mechanisms responsible for activating ARF following oncoproliferative stimuli. In search of factors that might associate with ARF to control its activity or its specificity, we performed a yeast two-hybrid screen. We report here that the human homologue of spinophilin/neurabin II, a regulatory subunit of protein phosphatase 1 catalytic subunit specifically interacts with ARF, both in yeast and in mammalian cells. We also show that ectopic expression of spinophilin/neurabin II inhibits the formation of G418-resistant colonies when transfected into human and mouse cell lines, regardless of p53 and ARF status. Moreover, spinophilin/ARF coexpression in Saos-2 cells, where ARF ectopic expression is ineffective, somehow results in a synergic effect. These data demonstrate a role for spinophilin in cell growth and suggest that ARF and spinophilin could act in partially overlapping pathways. PMID- 11278318 TI - The ERK signaling cascade inhibits gonadotropin-stimulated steroidogenesis. AB - The response of granulosa cells to luteinizing hormone (LH) and follicle stimulating hormone (FSH) is mediated mainly by cAMP/protein kinase A (PKA) signaling. Notably, the activity of the extracellular signal-regulated kinase (ERK) signaling cascade is elevated in response to these stimuli as well. We studied the involvement of the ERK cascade in LH- and FSH-induced steroidogenesis in two granulosa-derived cell lines, rLHR-4 and rFSHR-17, respectively. We found that stimulation of these cells with the appropriate gonadotropin induced ERK activation as well as progesterone production downstream of PKA. Inhibition of ERK activity enhanced gonadotropin-stimulated progesterone production, which was correlated with increased expression of the steroidogenic acute regulatory protein (StAR), a key regulator of progesterone synthesis. Therefore, it is likely that gonadotropin-stimulated progesterone formation is regulated by a pathway that includes PKA and StAR, and this process is down-regulated by ERK, due to attenuation of StAR expression. Our results suggest that activation of PKA signaling by gonadotropins not only induces steroidogenesis but also activates down-regulation machinery involving the ERK cascade. The activation of ERK by gonadotropins as well as by other agents may be a key mechanism for the modulation of gonadotropin-induced steroidogenesis. PMID- 11278319 TI - Vascular endothelial growth factor receptor-1 and neuropilin-2 form complexes. AB - The products of the neuropilin-1 (Np-1) and neuropilin-2 (Np-2) genes are receptors for factors belonging to the class 3 semaphorin family and participate in the guidance of growing axons to their targets. In the presence of heparin like molecules, both receptors also function as receptors for the heparin-binding 165-amino acid isoform of vascular endothelial growth factor (VEGF(165)). Both receptors are unable to bind to the 121-amino acid isoform of vascular endothelial growth factor (VEGF(121)), which lacks a heparin-binding domain. Interestingly, complexes corresponding in size to (125)I-VEGF(121).neuropilin complexes are formed when (125)I-VEGF(121) is bound and cross-linked to porcine aortic endothelial cells co-expressing VEGFR-1 and either Np-1 or Np-2. These complexes do not seem to represent complexes of (125)I-VEGF(121) with a truncated form of VEGFR-1, presumably formed as a result of the presence of Np-1 or Np-2 in the cells, because such truncated forms could not be detected with anti-VEGFR-1 antibodies. Antibodies directed against VEGFR-1 co-immunoprecipitated the (125)I VEGF(121).Np-2 sized cross-linked complex along with (125)I-VEGF(121).VEGFR-1 complexes from cells expressing both VEGFR-1 and Np-2 but not from control cells, indicating that VEGFR-1 and Np-2 associate with each other. To perform the reciprocal experiment we have expressed in porcine aortic endothelial cells a Np 2 receptor containing an in-frame myc epitope at the C terminus. Surprisingly, the myc-tagged Np-2 receptor lost most of its VEGF(165) binding capacity but not its semaphorin-3F binding ability. Nevertheless, when Np-2myc was co-expressed in cells with VEGFR-1, it partially regained its VEGF(165) binding ability. Antibodies directed against the myc epitope co-immunoprecipitated (125)I VEGF(165).Np-2myc and (125)I- VEGF(165).VEGFR-1 complexes from cells co expressing VEGFR-1 and Np-2myc, indicating again that VEGFR-1 associates with Np 2. Our experiments therefore indicate that Np-2, and possibly also Np-1, associate with VEGFR-1 and that such complexes may be part of a cell membrane associated signaling complex. PMID- 11278320 TI - Involvement of Bcl-2 and Bax in photodynamic therapy-mediated apoptosis. Antisense Bcl-2 oligonucleotide sensitizes RIF 1 cells to photodynamic therapy apoptosis. AB - Photodynamic therapy (PDT), a promising treatment modality, is an oxidative stress that induces apoptosis in many cancer cells in vitro and tumors in vivo. Understanding the mechanism(s) involved in PDT-mediated apoptosis may improve its therapeutic efficacy. Although studies suggest the involvement of multiple pathways, the triggering event(s) responsible for PDT-mediated apoptotic response is(are) not clear. To investigate the role of Bcl-2 in PDT-mediated apoptosis, we employed Bcl-2-antisense and -overexpression approaches in two cell types differing in their responses toward PDT apoptosis. In the first approach, we treated radiation-induced fibrosarcoma (RIF 1) cells, which are resistant to silicon phthalocyanine (Pc 4)-PDT apoptosis, with Bcl-2-antisense oligonucleotide. This treatment resulted in sensitization of RIF 1 cells to PDT mediated apoptosis as demonstrated by i) cleavage of poly(ADP-ribose) polymerase, ii) DNA ladder formation, iii) terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive cells, and iv) DEVDase activity. This treatment also resulted in oligonucleotide concentration-dependent decrease in cell viability and down-regulation of Bcl-2 protein with a concomitant increase in apoptosis. However, the level of Bax, a pro-apoptotic member of Bcl-2 family, remained unaltered. In the second approach, an overexpression of Bcl-2 in PDT apoptosis-sensitive human epidermoid carcinoma (A431) cells resulted in enhanced apoptosis and up-regulation of Bax following PDT. In both the approaches, the increased Bax/Bcl-2 ratio was associated with an increased apoptotic response of PDT. Our data also demonstrated that PDT results in modulation of other Bcl-2 family members in a way that the overall ratio of pro-apoptotic and anti apoptotic member proteins favors apoptosis. PMID- 11278321 TI - Molecular cloning and characterization of a zinc finger protein involved in Id-1 stimulated mammary epithelial cell growth. AB - Id proteins are dominant negative regulators of basic helix-loop-helix transcription factors. Previous work in our laboratory has shown that constitutive expression of Id-1 in SCp2 mouse mammary epithelial cells inhibits their differentiation and induces proliferation, invasion, and migration. Id-1 expression also correlates with the invasive and aggressive potential of human breast cancer cells. However, little is known about Id-1 target genes that are important for regulating normal and transformed breast epithelial cell phenotypes. Now we report the cloning of a novel zinc finger protein, Zfp289, using degenerate primers to specifically amplify cDNAs from Id-1-transfected SCp2 cells. Zfp289 has homology with a yeast zinc finger protein, the GTPase activating protein Gcs-1, which was initially identified as a gene required for the re-entry of cells into the cell cycle after stationary phase growth. Zfp289 mRNA expression pattern correlates with Id-1 expression in SCp2 mammary epithelial cells under various experimental conditions as well as in the mouse mammary gland at different stages of development. It is predominantly present in the cytoplasm of the cells as evident from green fluorescent protein fusion protein localization. SCp2 mammary epithelial cells with constitutive expression of Zfp289 have a higher S-phase index, compared with control cells, when cultured in a serum-free medium. We conclude that the novel zinc finger protein Zfp289, which may represent the mammalian homologue of Gcs-1, is potentially an important mediator of the Id-1-induced proliferation pathway in mammary epithelial cells. PMID- 11278322 TI - Lefty proteins exhibit unique processing and activate the MAPK pathway. AB - Lefty polypeptides, novel members of the transforming growth factor-beta (TGF beta) superfamily, are involved in the formation of embryonic lateral patterning. Members of the TGF-beta superfamily require processing for their activation, suggesting cleavage to be an essential step for lefty activation. Transfection of different cell lines with lefty resulted in expression of a 42-kDa protein, which was proteolytically processed to release two polypeptides of 34 and 28 kDa. Since members of the proprotein convertase (PC) family cleave different TGF-beta factors and are involved in the establishment of embryonic laterality, we studied their role in lefty processing. Cotransfection analysis showed that PC5A processed the lefty precursor to the 34-kDa form in vivo, whereas furin, PACE4, PC5B, and PC7 had a limited activity. None of these PCs showed activity in the processing of the lefty polypeptide to the 28-kDa lefty form. The mutation of the consensus sequences for PC cleavage in the lefty protein allowed the lefty cleavage sites to be identified. Mutations of the sequence RGKR to GGKG (amino acids 74-77) and of RHGR to GHGR (amino acids 132-135) prevented the proteolytic processing of the lefty precursor to the 34- and 28-kDa forms, respectively. To identify the biologically active form of lefty, we studied the effect of lefty treatment on pluripotent P19 cells. Lefty did not induce Smad2 or Smad5 phosphorylation, Smad2/Smad4 heterodimerization, or nuclear translocation of Smad2 or Smad4, but activated the MAPK pathway in a time- and dose-dependent fashion. Further analysis showed the 28-kDa (but not the 34-kDa) polypeptide to induce MAPK activity. Surprisingly, the 42-kDa lefty protein was also capable of inducing MAPK activity, indicating that the lefty precursor is biologically active. The data support a molecular model of processing as a mechanism for regulation of lefty signaling. PMID- 11278323 TI - Cloning and biological activity of epigen, a novel member of the epidermal growth factor superfamily. AB - High throughput sequencing of a mouse keratinocyte library was used to identify an expressed sequence tag with homology to the epidermal growth factor (EGF) family of growth factors. We have named the protein encoded by this expressed sequence tag Epigen, for epithelial mitogen. Epigen encodes a protein of 152 amino acids that contains features characteristic of the EGF superfamily. Two hydrophobic regions, corresponding to a putative signal sequence and transmembrane domain, flank a core of amino acids encompassing six cysteine residues and two putative N-linked glycosylation sites. Epigen shows 24-37% identity to members of the EGF superfamily including EGF, transforming growth factor alpha, and Epiregulin. Northern blotting of several adult mouse tissues indicated that Epigen was present in testis, heart, and liver. Recombinant Epigen was synthesized in Escherichia coli and refolded, and its biological activity was compared with that of EGF and transforming growth factor alpha in several assays. In epithelial cells, Epigen stimulated the phosphorylation of c-erbB-1 and mitogen-activated protein kinases and also activated a reporter gene containing enhancer sequences present in the c-fos promoter. Epigen also stimulated the proliferation of HaCaT cells, and this proliferation was blocked by an antibody to the extracellular domain of the receptor tyrosine kinase c-erbB-1. Thus, Epigen is the newest member of the EGF superfamily and, with its ability to promote the growth of epithelial cells, may constitute a novel molecular target for wound-healing therapy. PMID- 11278324 TI - Dopamine D2 receptor dimer formation: evidence from ligand binding. AB - We have examined the binding of two radioligands ([(3)H]spiperone and [(3)H]raclopride) to D(2) dopamine receptors expressed in Chinese hamster ovary cells. In saturation binding experiments in the presence of sodium ions, both radioligands labeled a similar number of sites, whereas in the absence of sodium ions [(3)H]raclopride labeled about half the number of sites labeled by [(3)H]spiperone. In competition experiments in the absence of sodium ions, however, raclopride was able to inhibit [(3)H]spiperone binding fully. In saturation analyses with [(3)H]spiperone in the absence of sodium ions raclopride exerted noncompetitive effects, decreasing the number of sites labeled by the radioligand. These data are interpreted in terms of a model where the receptor exists as a dimer, and in the absence of sodium ions, raclopride exerts negative cooperativity across the dimer both for its own binding and the binding of spiperone. A model of the receptor has been produced that provides a good description of the experimental phenomena described here. PMID- 11278325 TI - Metabotropic glutamate 1alpha and adenosine A1 receptors assemble into functionally interacting complexes. AB - Recently, evidence has emerged that seven transmembrane G protein-coupled receptors may be present as homo- and heteromers in the plasma membrane. Here we describe a new molecular and functional interaction between two functionally unrelated types of G protein-coupled receptors, namely the metabotropic glutamate type 1alpha (mGlu(1alpha) receptor) and the adenosine A1 receptors in cerebellum, primary cortical neurons, and heterologous transfected cells. Co immunoprecipitation experiments showed a close and subtype-specific interaction between mGlu(1alpha) and A1 receptors in both rat cerebellar synaptosomes and co transfected HEK-293 cells. By using transiently transfected HEK-293 cells a synergy between mGlu(1alpha) and A1 receptors in receptor-evoked [Ca(2+)](i) signaling has been shown. In primary cultures of cortical neurons we observed a high degree of co-localization of the two receptors, and excitotoxicity experiments in these cultures also indicate that mGlu(1alpha) and A1 receptors are functionally related. Our results provide a molecular basis for adenosine/glutamate receptors cross-talk and open new perspectives for the development of novel agents to treat neuropsychiatric disorders in which abnormal glutamatergic neurotransmission is involved. PMID- 11278326 TI - N-terminal domains of the class ia phosphoinositide 3-kinase regulatory subunit play a role in cytoskeletal but not mitogenic signaling. AB - Phosphoinositide (PI) 3-kinases are required for the acute regulation of the cytoskeleton by growth factors. We have shown previously that in the MTLn3 rat adenocarcinoma cells line, the p85/p110alpha PI 3-kinase is required for epidermal growth factor (EGF)-stimulated lamellipod extension and formation of new actin barbed ends at the leading edge of the cell. We have now examined the role of the p85alpha regulatory subunit in greater detail. Microinjection of recombinant p85alpha into MTLn3 cells blocked both EGF-stimulated mitogenic signaling and lamellipod extension. In contrast, a truncated p85(1-333), which lacks the SH2 and iSH2 domains and does not bind p110, had no effect on EGF stimulated mitogenesis but still blocked EGF-stimulated lamellipod extension. Additional deletional analysis showed that the SH3 domain was not required for inhibition of lamellipod extension, as a construct containing only the proline rich and breakpoint cluster region (BCR) homology domains was sufficient for inhibition. Although the BCR domain of p85 binds Rac, the effects of the p85 constructs were not because of a general inhibition of Rac signaling, because sorbitol-induced JNK activation in MTLn3 cells was not inhibited. These data show that the proline-rich and BCR homology domains of p85 are involved in the coupling of p85/p110 PI 3-kinases to regulation of the actin cytoskeleton. These data provide evidence of a distinct cellular function for the N-terminal domains of p85. PMID- 11278327 TI - Expression studies on clustered trypanosomatid box C/D small nucleolar RNAs. AB - We analyzed three chromosomal loci of the trypanosomatid Leptomonas collosoma encoding box C/D small nucleolar RNAs (snoRNAs). All the snoRNAs that were analyzed here carry two sequences complementary to rRNA target sites and obey the +5 rule for guide methylation. Studies on transgenic parasites carrying the snoRNA-2 gene in the episomal expression vector (pX-neo) indicated that no promoter activity was found immediately adjacent to this gene. Deleting the flanking sequences of snoRNA-2 affected the expression; in the absence of the 3' flanking (but not 5'-flanking) sequence, the expression was almost completely abolished. The snoRNA genes are transcribed as polycistronic RNA. All snoRNAs can be folded into a common stem-loop structure, which may play a role in processing the polycistronic transcript. snoRNA B2, a member of a snoRNA cluster, was expressed when cloned into the episomal vector, suggesting that each gene within a cluster is individually processed. Studies with permeable cells indicated that snoRNA gene transcription was relatively sensitive to alpha-amanitin, thus supporting transcription by RNA polymerase II. We propose that snoRNA gene expression, similar to protein-coding genes in this family, is regulated at the processing level. PMID- 11278328 TI - Agrin-induced activation of acetylcholine receptor-bound Src family kinases requires Rapsyn and correlates with acetylcholine receptor clustering. AB - During neuromuscular synaptogenesis, neurally released agrin induces aggregation and tyrosine phosphorylation of acetylcholine receptors (AChRs) by acting through both the receptor tyrosine kinase MuSK (muscle-specific kinase) and the AChR associated protein, rapsyn. To elucidate this signaling mechanism, we examined tyrosine phosphorylation of AChR-associated proteins, particularly addressing whether agrin activates Src family kinases bound to the AChR. In C2 myotubes, agrin induced tyrosine phosphorylation of these kinases, of AChR-bound MuSK, and of the AChR beta and delta subunits, as observed in phosphotyrosine immunoblotting experiments. Kinase assays revealed that the activity of AChR associated Src kinases was increased by agrin, whereas phosphorylation of the total cellular kinase pool was unaffected. In both rapsyn-deficient myotubes and staurosporine-treated C2 myotubes, where AChRs are not clustered, agrin activated MuSK but did not cause either Src family or AChR phosphorylation. In S27 mutant myotubes, which fail to aggregate AChRs, no agrin-induced phosphorylation of AChR bound Src kinases, MuSK, or AChRs was observed. These results demonstrate first that agrin leads to phosphorylation and activation of AChR-associated Src-related kinases, which requires rapsyn, occurs downstream of MuSK, and causes AChR phosphorylation. Second, this activation intimately correlates with AChR clustering, suggesting that these kinases may play a role in agrin-induced AChR aggregation by forming an AChR-bound signaling cascade. PMID- 11278329 TI - Tumor cell invasion is promoted by activation of protease activated receptor-1 in cooperation with the alpha vbeta 5 integrin. AB - The first prototype of the protease activated receptor (PAR) family, the thrombin receptor PAR1, plays a central role both in the malignant invasion process of breast carcinoma metastasis and in the physiological process of placental implantation. The molecular mechanism underlying PAR1 involvement in tumor invasion and metastasis, however, is poorly defined. Here we show that PAR1 increases the invasive properties of tumor cells primarily by increased adhesion to extracellular matrix components. This preferential adhesion is accompanied by the cytoskeletal reorganization of F-actin toward migration-favoring morphology as detected by phalloidin staining. Activation of PAR1 increased the phosphorylation of focal adhesion kinase and paxillin, and the induced formation of focal contact complexes. PAR1 activation affected integrin cell-surface distribution without altering their level of expression. The specific recruitment of alpha(v)beta(5) to focal contact sites, but not of alpha(v)beta(3) or alpha(5)beta(1), was observed by immunofluorescent microscopy. PAR1 overexpressing cells showed selective reciprocal co-precipitation with alpha(v)beta(5) and paxillin but not with alpha(v)beta(3) that remained evenly distributed under these conditions. This co-immunoprecipitation failed to occur in cells containing the truncated form of PAR1 that lacked the entire cytoplasmic portion of the receptor. Thus, the PAR1 cytoplasmic tail is essential for conveying the cross-talk and recruiting the alpha(v)beta(5) integrin. While PAR1 overexpressing cells were invasive in vitro, as reflected by their migration through a Matrigel barrier, invasion was further enhanced by ligand activation of PAR1. Moreover, the application of anti-alpha(v)beta(5) antibodies specifically attenuated this PAR1 induced invasion. We propose that the activation of PAR1 may lead to a novel cooperation with the alpha(v)beta(5) integrin that supports tumor cell invasion. PMID- 11278330 TI - Modulation of the basolateral and apical step of transepithelial organic anion secretion in proximal tubular opossum kidney cells. Acute effects of epidermal growth factor and mitogen-activated protein kinase. AB - The organic anion transport system in the proximal tubule of the kidney is of major importance for the excretion of a variety of endogenous and potentially toxic exogenous substances. Furthermore, the clearance of model substrates (e.g. para-aminohippurate) of this system is used for the determination of renal blood flow. We investigated regulation of organic anion secretion in a way that allowed us to examine simultaneously regulation of overall transepithelial secretion and to estimate the separate contributions of regulation of the basolateral and apical transport steps to this overall regulation. The data were verified by measurement of initial basolateral uptake rate and initial apical efflux rate. Opossum kidney cells were used as a suitable model system for proximal tubule cells, and [14C]para-aminohippurate was utilized as an organic anion. Stimulation of protein kinase C inhibited transepithelial secretion because of inhibition of both apical efflux and basolateral uptake. Inhibition of the mitogen-activated protein kinase (MAPK) kinase MEK reduced transepithelial secretion via inhibition of basolateral uptake and apical efflux. Epidermal growth factor (EGF) enhanced transepithelial secretion via stimulation of basolateral uptake but did not affect apical efflux. EGF induced stimulation of basolateral uptake was abolished by inhibition of MEK. EGF led to phosphorylation of ERK1/2, which was also abolished by inhibition of MEK. Thus, EGF stimulated basolateral uptake of organic anions via MAPKs. Transepithelial organic anion secretion can be regulated at two sites, at least: basolateral uptake and apical efflux. Both steps are under control of protein kinase C and MAPK. The pathophysiologically relevant growth factor EGF enhances transepithelial secretion via stimulation of basolateral uptake. EGF stimulates basolateral uptake via MEK and ERK1/2. Thus, renal organic anion extraction may be modulated, especially under pathophysiological conditions. PMID- 11278331 TI - The role of the membrane in the inactivation of factor va by plasmin. Amino acid region 307-348 of factor V plays a critical role in factor Va cofactor function. AB - The mechanism of inactivation of bovine factor Va by plasmin was studied in the presence and absence of phospholipid vesicles (PCPS vesicles). Following 60-min incubation with plasmin (4 nm) membrane-bound factor Va (400 nm) is completely inactive, whereas in the absence of phospholipid vesicles following a 1-h incubation period, the cofactor retains 90% of its initial cofactor activity. Amino acid sequencing of the fragments deriving from cleavage of factor Va by plasmin demonstrated that while both chains of factor Va are cleaved by plasmin, only cleavage of the heavy chain correlates with inactivation of the cofactor. In the presence of a membrane surface the heavy chain of the bovine cofactor is first cleaved at Arg(348) to generate a fragment of M(r) 47,000 containing the NH(2)-terminal part of the cofactor (amino acid residues 1-348) and a M(r) 42,000 fragment (amino acid residues 349-713). This cleavage is associated with minimal loss in cofactor activity. Complete loss of activity of the membrane-bound cofactor coincides with three cleavages at the COOH-terminal portion of the M(r) 47,000 fragment: Lys(309), Lys(310), and Arg(313). These cleavages result in the release of the COOH terminus of the molecule and the production of a M(r) 40,000 fragment containing the NH(2)-terminal portion of the factor Va molecule. Factor Va was treated with plasmin in the absence of phospholipid vesicles followed by the addition of PCPS vesicles and activated protein C (APC). A rapid inactivation of the cofactor was observed as a result of cleavage of the M(r) 47,000 fragment at Arg(306) by APC and appearance of a M(r) 39,000 fragment. These data suggest a critical role of the amino acid sequence 307-348 of factor Va. A 42-amino acid peptide encompassing the region 307-348 of human factor Va (N42R) was found to be a good inhibitor of factor Va clotting activity with an IC(50) of approximately 1.3 microm. These data suggest that plasmin is a potent inactivator of factor Va and that region 307-348 of the cofactor plays a critical role in cofactor function and may be responsible for the interaction of the cofactor with factor Xa and/or prothrombin. PMID- 11278332 TI - Binding of heterochromatin protein 1 to the nuclear envelope is regulated by a soluble form of tubulin. AB - We have previously shown that the mouse heterochromatin protein 1 homologue M31 interacts dynamically with the nuclear envelope. Using quantitative in vitro assays, we now demonstrate that this interaction is potently inhibited by soluble factors present in mitotic and interphase cytosol. As indicated by depletion and order-of-addition experiments, the inhibitory activity co-isolates with a 55-kDa protein, which binds avidly to the nuclear envelope and presumably blocks M31 binding sites. Purification of this protein and microsequencing of tryptic peptides identify it as alpha2/6:beta2-tubulin. Consistent with this observation, bona fide tubulin, isolated from rat brain and maintained in a nonpolymerized state, abolishes binding of M31 to the nuclear envelope and aborts M31-mediated nuclear envelope reassembly in an in vitro system. These observations provide a new example of "moonlighting," a process whereby multimeric proteins switch function when their aggregation state or localization is altered. PMID- 11278333 TI - Activation of the phosphatidylinositol 3-kinase/Akt pathway protects against interleukin-3 starvation but not DNA damage-induced apoptosis. AB - Baf-3 cells are dependent on interleukin-3 (IL-3) for their survival and proliferation in culture. To identify anti-apoptotic pathways, we performed a retroviral-insertion mutagenesis on Baf-3 cells and selected mutants that have acquired a long term survival capacity. The phenotype of one mutant, which does not overexpress bcl-x and proliferates in the absence of IL-3, is described. We show that, in this mutant, Akt is constitutively activated leading to FKHRL1 phosphorylation and constitutive glycolytic activity. This pathway is necessary for the mutant to survive following IL-3 starvation but is not sufficient or necessary to protect cells from DNA damage-induced cell death. Indeed, inhibition of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in Baf-3 cells does not prevent the ability of IL-3 to protect cells against gamma-irradiation-induced DNA damage. This protective effect of IL-3 rather correlates with the expression of the anti-apoptotic Bcl-x protein. Taken together, these data demonstrate that the PI3K/Akt pathway is sufficient to protect cells from growth factor starvation induced apoptosis but is not required for IL-3 inhibition of DNA damage-induced cell death. PMID- 11278334 TI - Phosphorylation of protein phosphatase inhibitor-1 by Cdk5. AB - Protein phosphatase inhibitor-1 is a prototypical mediator of cross-talk between protein kinases and protein phosphatases. Activation of cAMP-dependent protein kinase results in phosphorylation of inhibitor-1 at Thr-35, converting it into a potent inhibitor of protein phosphatase-1. Here we report that inhibitor-1 is phosphorylated in vitro at Ser-67 by the proline-directed kinases, Cdk1, Cdk5, and mitogen-activated protein kinase. By using phosphorylation state-specific antibodies and selective protein kinase inhibitors, Cdk5 was found to be the only kinase that phosphorylates inhibitor-1 at Ser-67 in intact striatal brain tissue. In vitro and in vivo studies indicated that phospho-Ser-67 inhibitor-1 was dephosphorylated by protein phosphatases-2A and -2B. The state of phosphorylation of inhibitor-1 at Ser-67 was dynamically regulated in striatal tissue by glutamate-dependent regulation of N-methyl-d-aspartic acid-type channels. Phosphorylation of Ser-67 did not convert inhibitor-1 into an inhibitor of protein phosphatase-1. However, inhibitor-1 phosphorylated at Ser-67 was a less efficient substrate for cAMP-dependent protein kinase. These results demonstrate regulation of a Cdk5-dependent phosphorylation site in inhibitor-1 and suggest a role for this site in modulating the amplitude of signal transduction events that involve cAMP-dependent protein kinase activation. PMID- 11278335 TI - Inhibition of cell growth and spreading by stomach cancer-associated protein tyrosine phosphatase-1 (SAP-1) through dephosphorylation of p130cas. AB - SAP-1 (stomach cancer-associated protein-tyrosine phosphatase-1) is a transmembrane-type protein-tyrosine phosphatase that is abundant in the brain and certain cancer cell lines. With the use of a "substrate-trapping" approach, p130(cas), a major focal adhesion-associated phosphotyrosyl protein, has now been identified as a likely physiological substrate of SAP-1. Expression of recombinant SAP-1 induced the dephosphorylation of p130(cas) as well as that of two other components of the integrin-signaling pathway (focal adhesion kinase and p62(dok)) in intact cells. In contrast, expression of a substrate-trapping mutant of SAP-1 induced the hyperphosphorylation of these proteins, indicating a dominant negative effect of this mutant. Overexpression of SAP-1 induced disruption of the actin-based cytoskeleton as well as inhibited various cellular responses promoted by integrin-mediated cell adhesion, including cell spreading on fibronectin, growth factor-induced activation of extracellular signal regulated kinase 2, and colony formation. Finally, the enzymatic activity of SAP 1, measured with an immunocomplex phosphatase assay, was substantially increased by cell-cell adhesion. These results suggest that SAP-1, by mediating the dephosphorylation of focal adhesion-associated substrates, negatively regulates integrin-promoted signaling processes and, thus, may contribute to contact inhibition of cell growth and motility. PMID- 11278336 TI - Peroxisome proliferator-activated receptor gamma ligands suppress the transcriptional activation of cyclooxygenase-2. Evidence for involvement of activator protein-1 and CREB-binding protein/p300. AB - We investigated whether peroxisome proliferator-activated receptor gamma (PPARgamma) ligands (ciglitazone, troglitazone, and 15-deoxy-Delta(12,14) prostaglandin J(2)) inhibited cyclooxygenase-2 (COX-2) induction in human epithelial cells. Ligands of PPARgamma inhibited phorbol ester (phorbol 12 myristate 13-acetate, PMA)-mediated induction of COX-2 and prostaglandin E(2) synthesis. Nuclear run-offs revealed increased rates of COX-2 transcription after treatment with PMA, an effect that was inhibited by PPARgamma ligands. PMA mediated induction of COX-2 promoter activity was inhibited by PPARgamma ligands; this suppressive effect was prevented by overexpressing a dominant negative form of PPARgamma or a PPAR response element decoy oligonucleotide. The stimulatory effects of PMA were mediated by a cyclic AMP response element in the COX-2 promoter. Treatment with PMA increased activator protein-1 (AP-1) activity and the binding of c-Jun, c-Fos, and ATF-2 to the cyclic AMP response element, effects that were blocked by PPARgamma ligands. These findings raised questions about the mechanism underlying the anti-AP-1 effect of PPARgamma ligands. The induction of c-Jun by PMA was blocked by PPARgamma ligands. Overexpression of either c-Jun or CREB-binding protein/p300 partially relieved the suppressive effect of PPARgamma ligands. When CREB-binding protein and c-Jun were overexpressed together, the ability of PPARgamma ligands to suppress PMA-mediated induction of COX-2 promoter activity was essentially abrogated. Bisphenol A diglycidyl ether, a compound that binds to PPARgamma but lacks the ability to activate transcription, also inhibited PMA-mediated induction of AP-1 activity and COX-2. Taken together, these findings are likely to be important for understanding the anti-inflammatory and anti-cancer properties of PPARgamma ligands. PMID- 11278337 TI - Retention of the Alzheimer's amyloid precursor fragment C99 in the endoplasmic reticulum prevents formation of amyloid beta-peptide. AB - gamma-Secretase is a membrane-associated endoprotease that catalyzes the final step in the processing of Alzheimer's beta-amyloid precursor protein (APP), resulting in the release of amyloid beta-peptide (Abeta). The molecular identity of gamma-secretase remains in question, although recent studies have implicated the presenilins, which are membrane-spanning proteins localized predominantly in the endoplasmic reticulum (ER). Based on these observations, we have tested the hypothesis that gamma-secretase cleavage of the membrane-anchored C-terminal stump of APP (i.e. C99) occurs in the ER compartment. When recombinant C99 was expressed in 293 cells, it was localized mainly in the Golgi apparatus and gave rise to abundant amounts of Abeta. Co-expression of C99 with mutant forms of presenilin-1 (PS1) found in familial Alzheimer's disease resulted in a characteristic elevation of the Abeta(42)/Abeta(40) ratio, indicating that the N terminal exodomain of APP is not required for mutant PS1 to influence the site of gamma-secretase cleavage. Biogenesis of both Abeta(40) and Abeta(42) was almost completely eliminated when C99 was prevented from leaving the ER by addition of a di-lysine retention motif (KKQN) or by co-expression with a dominant-negative mutant of the Rab1B GTPase. These findings indicate that the ER is not a major intracellular site for gamma-secretase cleavage of C99. Thus, by inference, PS1 localized in this compartment does not appear to be active as gamma-secretase. The results suggest that presenilins may acquire the characteristics of gamma secretase after leaving the ER, possibly by assembling with other proteins in peripheral membranes. PMID- 11278338 TI - CD15 expression in mature granulocytes is determined by alpha 1,3 fucosyltransferase IX, but in promyelocytes and monocytes by alpha 1,3 fucosyltransferase IV. AB - The CD15 carbohydrate epitope is expressed in mature human neutrophils, monocytes, and promyelocytes. We aimed to determine the alpha1,3 fucosyltransferase responsible for the expression of CD15 in each subpopulation of leukocytes. Three alpha1,3-fucosyltransferases, FUT4, FUT7, and FUT9, are expressed in human leukocytes. We demonstrated that FUT9 exhibits 20-fold stronger activity for CD15 synthesis than FUT4, whereas FUT4 exhibits 4.5-fold stronger activity for CDw65 synthesis than FUT9. By competitive reverse transcriptase-polymerase chain reaction, FUT9 was found to be strongly expressed in mature granulocytes and peripheral blood mononuclear cell, but not in monocytes. CD34(+) and CD15(+) cells in cord blood and myeloid cell lines (HL-60 and U937) did not express FUT9 at all. FUT4 transcripts were ubiquitously expressed in all blood cells and all cultured cell lines, with HL-60 and U937 cells in particular expressing a number of FUT4 transcripts. Transfection of the FUT9 gene into Jurkat and U937 cells demonstrated that FUT9 has the potential to express CD15 in myeloid and lymphoid cells. These findings suggest that the expression of CD15 in mature granulocytes is directed by FUT9, whereas it is determined in promyelocytes and monocytes by FUT4. Measurement of CD15 synthesizing activity in cell homogenates of each cell population using the polylactosamine acceptor further supported these conclusions. PMID- 11278339 TI - Insulin stimulates PKCzeta -mediated phosphorylation of insulin receptor substrate-1 (IRS-1). A self-attenuated mechanism to negatively regulate the function of IRS proteins. AB - Incubation of rat hepatoma Fao cells with insulin leads to a transient rise in Tyr phosphorylation of insulin receptor substrate (IRS) proteins. This is followed by elevation in their P-Ser/Thr content, and their dissociation from the insulin receptor (IR). Wortmannin, a phosphatidylinositol 3-kinase (PI3K) inhibitor, abolished the increase in the P-Ser/Thr content of IRS-1, its dissociation from the IR, and the decrease in its P-Tyr content following 60 min of insulin treatment, indicating that the Ser kinases that negatively regulate IRS-1 function are downstream effectors of PI3K. PKCzeta fulfills this criterion, being an insulin-activated downstream effector of PI3K. Overexpression of PKCzeta in Fao cells, by infection of the cells with adenovirus-based PKCzeta construct, had no effect on its own, but it accelerated the rate of insulin-stimulated dissociation of IR.IRS-1 complexes and the rate of Tyr dephosphorylation of IRS 1. The insulin-stimulated negative regulatory role of PKCzeta was specific and could not be mimic by infecting Fao cells with adenoviral constructs encoding for PKC alpha, delta, or eta. Because the reduction in P-Tyr content of IRS-1 was accompanied by a reduced association of IRS-1 with p85, the regulatory subunit of PI3K, it suggests that this negative regulatory process induced by PKCzeta, has a built-in attenuation signal. Hence, insulin triggers a sequential cascade in which PI3K-mediated activation of PKCzeta inhibits IRS-1 functions, reduces complex formation between IRS-1 and PI3K, and inhibits further activation of PKCzeta itself. These findings implicate PKCzeta as a key element in a multistep negative feedback control mechanism of IRS-1 functions. PMID- 11278340 TI - Interaction of hematopoietic progenitor kinase 1 and c-Abl tyrosine kinase in response to genotoxic stress. AB - The c-Abl protein tyrosine kinase is activated by certain DNA-damaging agents and regulates induction of the stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK). The hematopoietic progenitor kinase 1 (HPK1) has also been shown to act upstream to the SAPK/JNK signaling pathway. We report here that exposure of hematopoietic Jurkat T cells to genotoxic agents is associated with activation of HPK1. The results demonstrate that exposure of Jurkat cells to DNA damaging agents is associated with translocation of active c-Abl from nuclei to cytoplasm and binding of c-Abl to HPK1. Our findings also demonstrate that c-Abl phosphorylates HPK1 in cytoplasm and stimulates HPK1 activity. The functional significance of the c-Abl-HPK1 interaction is supported by the demonstration that this complex regulates SAPK/JNK activation. Overexpression of c-Abl(K-R) inhibits HPK1-induced activation of SAPK/JNK. Conversely, the dominant negative mutant of HPK1 blocks c-Abl-mediated induction of SAPK/JNK. These findings indicate that activation of HPK1 and formation of HPK1/c-Abl complexes are functionally important in the stress response of hematopoietic cells to genotoxic agents. PMID- 11278341 TI - The Ca2+-sensing receptor activates cytosolic phospholipase A2 via a Gqalpha dependent ERK-independent pathway. AB - The Ca(2+)-sensing receptor (CaR) stimulates a number of phospholipase activities, but the specific phospholipases and the mechanisms by which the CaR activates them are not defined. We investigated regulation of phospholipase A(2) (PLA(2)) by the Ca(2+)-sensing receptor (CaR) in human embryonic kidney 293 cells that express either the wild-type receptor or a nonfunctional mutant (R796W) CaR. The PLA(2) activity was attributable to cytosolic PLA(2) (cPLA(2)) based on its inhibition by arachidonyl trifluoromethyl ketone, lack of inhibition by bromoenol lactone, and enhancement of the CaR-stimulated phospholipase activity by coexpression of a cDNA encoding the 85-kDa human cPLA(2). No CaR-stimulated cPLA(2) activity was found in the cells that expressed the mutant CaR. Pertussis toxin treatment had a minimal effect on CaR-stimulated arachidonic acid release and the CaR-stimulated rise in intracellular Ca(2+) (Ca(2+)(i)), whereas inhibition of phospholipase C (PLC) with completely inhibited CaR-stimulated PLC and cPLA(2) activities. CaR-stimulated PLC activity was inhibited by expression of RGS4, an RGS (Regulator of G protein Signaling) protein that inhibits Galpha(q) activity. CaR-stimulated cPLA(2) activity was inhibited 80% by chelation of extracellular Ca(2+) and depletion of intracellular Ca(2+) with EGTA and inhibited 90% by treatment with W7, a calmodulin inhibitor, or with KN-93, an inhibitor of Ca(2+), calmodulin-dependent protein kinases. Chemical inhibitors of the ERK activator, MEK, and a dominant negative MEK, MEK(K97R), had no effect on CaR-stimulated cPLA(2) activity but inhibited CaR-stimulated ERK activity. These results demonstrate that the CaR activates cPLA(2) via a Galpha(q), PLC, Ca(2+) CaM, and calmodulin-dependent protein kinase-dependent pathway that is independent the ERK pathway. PMID- 11278342 TI - L-plastin peptide activation of alpha(v)beta(3)-mediated adhesion requires integrin conformational change and actin filament disassembly. AB - L-plastin (LPL) is a leukocyte actin binding protein previously implicated in the activation of the integrin alpha(M)beta(2) on polymorphonuclear neutrophils. To determine the role for LPL in integrin activation, K562 cell adhesion to vitronectin via alpha(v)beta(3), a well-studied model for activable integrins, was examined. Cell permeant versions of peptides based on the N-terminal sequence of LPL and the LPL headpiece domain both activated alpha(v)beta(3)-mediated adhesion. In contrast to adhesion induced by treatment with phorbol 12-myristate 13-acetate (PMA), LPL peptide-activated adhesion was independent of integrin beta(3) cytoplasmic domain tyrosines and was not inhibited by cytochalasin D. Also in contrast to PMA, LPL peptides synergized with RGD ligand or Mn(2+) for generation of a conformational change in alpha(v)beta(3) associated with the high affinity state of the integrin, as determined by binding of a ligand-induced binding site antibody. Although LPL and ligand showed synergy for ligand-induced binding site expression when actin depolymerization was inhibited by jasplakinolide, LPL peptide-induced adhesion was inhibited. Thus, both actin depolymerization and ligand-induced integrin conformational change are required for LPL peptide-induced adhesion. We hypothesize that the critical steps of increased integrin diffusion and affinity enhancement may be linked via modulation of the function of the actin binding protein L-plastin. PMID- 11278343 TI - Prion protein contains a second endoplasmic reticulum targeting signal sequence located at its C terminus. AB - Prion protein (PrP) is synthesized at the membrane of the endoplasmic reticulum (ER) in three different topological forms as follows: a fully translocated one ((sec)PrP) and two with opposite orientations in the membrane ((Ntm)PrP and (Ctm)PrP). We asked whether other signal sequences exist in the PrP, other than the N-terminal signal sequence, that contribute to its topological diversity. In vitro translocation assays showed that PrP lacking its N-terminal signal sequence could still translocate into ER microsomes, although at reduced efficiency. Deletion of each of the two hydrophobic regions in PrP revealed that the C terminally located hydrophobic region (TM2) can function as second signal sequence in PrP. Translocation mediated by the TM2 alone can occur post translationally and yields mainly (Ctm)PrP, which is implicated in some forms of neurodegeneration in prion diseases. We conclude that, in vitro, PrP can insert into ER membranes co- and post-translationally and can use two different signal sequences. We propose that the unusually complex topology of PrP results from the differential utilization of two signal sequences in PrP. PMID- 11278344 TI - Identification and characterization of an Entamoeba histolytica upstream regulatory element 3 sequence-specific DNA-binding protein containing EF-hand motifs. AB - The hgl5 gene of Entamoeba histolytica is negatively regulated through the upstream regulatory element 3 (URE3) DNA motif TATTCTATT. This motif is also present and significant in the function of the E. histolytica fdx gene promoter. A yeast one-hybrid screen was used to identify an E. histolytica cDNA encoding a protein (URE3-BP) that recognized this DNA motif. Analysis of the predicted amino acid sequence demonstrated the presence of two EF-hand motifs but identified no canonical DNA binding motifs. URE3-BP, expressed in bacteria, demonstrated Ca(2+) dependent and sequence-specific recognition of the URE3 DNA sequence as assessed by electrophoretic mobility shift assays. Antibodies raised against URE3-BP blocked the formation of the URE3 DNA-protein complex by native nuclear extracts. The URE3-BP protein was present in the E. histolytica nucleus and cytoplasm with an apparent molecular mass of 22.6 kDa. Our results represent the first use of a yeast genetic screen to identify, on the basis of function, a DNA-binding protein of an early branching eukaryote. Since the URE3 DNA can modulate gene expression in both a positive and negative manner, this protein may have more than one mechanism of interaction with transcriptional machinery. Characterization of URE3 BP should provide insight into transcription regulation and virulence control in this parasite. PMID- 11278345 TI - Phosphorylation of the N-ethylmaleimide-sensitive factor is associated with depolarization-dependent neurotransmitter release from synaptosomes. AB - Critical to SNARE protein function in neurotransmission are the accessory proteins, soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein (SNAP), and NSF, that play a role in activation of the SNAREs for membrane fusion. In this report, we demonstrate the depolarization-induced, calcium dependent phosphorylation of NSF in rat synaptosomes. Phosphorylation of NSF is coincident with neurotransmitter release and requires an influx of external calcium. Phosphoamino acid analysis of the radiolabeled NSF indicates a role for a serine/threonine-specific kinase. Synaptosomal phosphorylation of NSF is stimulated by phorbol esters and is inhibited by staurosporine, chelerythrine, bisindolylmaleimide I, calphostin C, and Ro31-8220 but not the calmodulin kinase II inhibitor, Kn-93, suggesting a role for protein kinase C (PKC). Indeed, NSF is phosphorylated by PKC in vitro at Ser-237 of the catalytic D1 domain. Mutation of this residue to glutamic acid or to alanine eliminates in vitro phosphorylation. Molecular modeling studies suggest that Ser-237 is adjacent to an inter-subunit interface at a position where its phosphorylation could affect NSF activity. Consistently, mutation of Ser-237 to Glu, to mimic phosphorylation, results in a hexameric form of NSF that does not bind to SNAP-SNARE complexes, whereas the S237A mutant does form complex. These data suggest a negative regulatory role for PKC phosphorylation of NSF. PMID- 11278346 TI - Nickel resistance and chromatin condensation in Saccharomyces cerevisiae expressing a maize high mobility group I/Y protein. AB - Expression of a maize cDNA encoding a high mobility group (HMG) I/Y protein enables growth of transformed yeast on a medium containing toxic nickel concentrations. No difference in the nickel content was measured between yeast cells expressing either the empty vector or the ZmHMG I/Y2 cDNA. The ZmHMG I/Y2 protein contains four AT hook motifs known to be involved in binding to the minor groove of AT-rich DNA regions. HMG I/Y proteins may act as architectural elements modifying chromatin structure. Indeed, a ZmHMG I/Y2-green fluorescent protein fusion protein was observed in yeast nuclei. Nickel toxicity has been suggested to occur through an epigenetic mechanism related to chromatin condensation and DNA methylation, leading to the silencing of neighboring genes. Therefore, the ZmHMG I/Y2 protein could prevent nickel toxicity by interfering with chromatin structure. Yeast cell growth in the presence of nickel and yeast cells expressing the ZmHMG I/Y2 cDNA increased telomeric URA3 gene silencing. Furthermore, ZmHMG I/Y2 restored a wild-type level of nickel sensitivity to the yeast (Delta)rpd3 mutant. Therefore, nickel resistance of yeast cells expressing the ZmHMG I/Y2 cDNA is likely achieved by chromatin structure modification, restricting nickel accessibility to DNA. PMID- 11278347 TI - The 1.8-A crystal structure of a matrix metalloproteinase 8-barbiturate inhibitor complex reveals a previously unobserved mechanism for collagenase substrate recognition. AB - The individual zinc endoproteinases of the tissue degrading matrix metalloproteinase (MMP) family share a common catalytic architecture but are differentiated with respect to substrate specificity, localization, and activation. Variation in domain structure and more subtle structural differences control their characteristic specificity profiles for substrates from among four distinct classes (Nagase, H., and Woessner, J. F. J. (1999) J. Biol. Chem. 274, 21491-21494). Exploitation of these differences may be decisive for the design of anticancer or other drugs, which should be highly selective for their particular MMP targets. Based on the 1.8-A crystal structure of human neutrophil collagenase (MMP-8) in complex with an active site-directed inhibitor (RO200-1770), we identify and describe new structural determinants for substrate and inhibitor recognition in addition to the primary substrate recognition sites. RO200-1770 induces a major rearrangement at a position relevant to substrate recognition near the MMP-8 active site (Ala206-Asn218). In stromelysin (MMP-3), competing stabilizing interactions at the analogous segment hinder a similar rearrangement, consistent with kinetic profiling of several MMPs. Despite the apparent dissimilarity of the inhibitors, the central 2-hydroxypyrimidine-4,6-dione (barbiturate) ring of the inhibitor RO200-1770 mimics the interactions of the hydroxamate-derived inhibitor batimastat (Grams, F., Reinemer, P., Powers, J. C., Kleine, T., Pieper, M., Tschesche, H., Huber, R., and Bode, W. (1995) Eur. J. Biochem. 228, 830-841) for binding to MMP-8. The two additional phenyl and piperidyl ring substituents of the inhibitor bind into the S1' and S2' pockets of MMP-8, respectively. The crystal lattice contains a hydrogen bond between the O(gamma) group of Ser209 and N(delta)1 of His207 of a symmetry related molecule; this interaction suggests a model for recognition of hydroxyprolines present in physiological substrates. We also identify a collagenase-characteristic cis peptide bond, Asn188-Tyr189, on a loop essential for collagenolytic activity. The sequence conservation pattern at this position marks this cis-peptide bond as a determinant for triple-helical collagen recognition and processing. PMID- 11278348 TI - Modified phosphatidylethanolamine as the active component of oxidized low density lipoprotein promoting platelet prothrombinase activity. AB - We analyzed the influence of the atherogenic oxidized low density lipoproteins (LDL) on the activity of the platelet prothrombinase complex, a major contributor to overall thrombin formation in vivo. Platelet dependent thrombin generation was found to be strongly stimulated by in vitro oxidized LDL. The enhancement was additive to that observed with the platelet agonist thrombin. Oxidized LDL increased the platelet binding of annexin-V, suggesting that the augmented surface exposure of aminophospholipids promoted the prothrombinase activity. All of the stimulatory activity of the oxidized LDL could be recovered in the microemulsions prepared from the lipid portion of the modified particles. Phospholipid vesicles were prepared containing the total lipids of the oxidized LDL but lacking specifically in one lipid component. Following the selective removal of the ethanolamine phospholipids (PE) from the LDL lipids, the platelet dependent thrombin formation was markedly reduced. Vesicles enriched with the isolated PE fraction alone enhanced the thrombin generation. Analyses with autoxidized phospholipids indicated that oxidation products of unsaturated diacyl PE were mainly responsible for the increased prothrombinase activity. Oxidized LDL and its PE fraction lost their stimulatory activity after treatment with NaCNBH(3), a chemical reductant of Schiff base adducts. Phospholipid vesicles supplemented with synthetic aldehyde-PE adducts largely reproduced the stimulation of the thrombin generation. We conclude that the oxidized LDL particles elicit a pronounced prothrombotic response by increasing the activity of the platelet prothrombinase complex. Specific oxidative modifications of the LDL-associated ethanolamine phospholipids are mainly responsible for this stimulation. PMID- 11278349 TI - Structure-function analysis of the zinc-binding region of the Clpx molecular chaperone. AB - The ClpX heat shock protein of Escherichia coli is a member of the universally conserved Hsp100 family of proteins, and possesses a putative zinc finger motif of the C(4) type. The ClpX is an ATPase which functions both as a substrate specificity component of the ClpXP protease and as a molecular chaperone. Using an improved purification procedure we show that the ClpX protein is a metalloprotein complexed with Zn(II) cations. Contrary to other Hsp100 family members, ClpXZn(II) exists in an oligomeric form even in the absence of ATP. We show that the single ATP-binding site of ClpX is required for a variety of tasks, namely, the stabilization of the ClpXZn(II) oligomeric structure, binding to ClpP, and the ClpXP-dependent proteolysis of the lambdaO replication protein. Release of Zn(II) from ClpX protein affects the ability of ClpX to bind ATP. ClpX, free of Zn(II), cannot oligomerize, bind to ClpP, or participate in ClpXP dependent proteolysis. We also show that ClpXDeltaCys, a mutant protein whose four cysteine residues at the putative zinc finger motif have been replaced by serine, behaves in similar fashion as wild type ClpX protein whose Zn(II) has been released either by denaturation and renaturation, or chemically by p hydroxymercuriphenylsulfonic acid. PMID- 11278350 TI - Further characterization of the helicase activity of eIF4A. Substrate specificity. AB - Eukaryotic initiation factor (eIF) 4A is the archetypal member of the DEAD box family of RNA helicases and is proposed to unwind structures in the 5' untranslated region of mRNA to facilitate binding of the 40 S ribosomal subunit. The helicase activity of eIF4A has been further characterized with respect to substrate specificity and directionality. Results confirm that the initial rate and amplitude of duplex unwinding by eIF4A is dependent on the overall stability, rather than the length or sequence, of the duplex substrate. eIF4A helicase activity is minimally dependent on the length of the single-stranded region adjacent to the double-stranded region of the substrate. Interestingly, eIF4A is able to unwind blunt-ended duplexes. eIF4A helicase activity is also affected by substitution of 2'-OH (RNA) groups with 2'-H (DNA) or 2'-methoxyethyl groups. These observations, taken together with results from competitive inhibition experiments, suggest that eIF4A may interact directly with double-stranded RNA, and recognition of helicase substrates occurs via chemical and/or structural features of the duplex. These results allow for refinement of a previously proposed model for the mechanism of action of eIF4A helicase activity. PMID- 11278351 TI - Assembled F1-(alpha beta ) and Hybrid F1-alpha 3beta 3gamma -ATPases from Rhodospirillum rubrum alpha, wild type or mutant beta, and chloroplast gamma subunits. Demonstration of Mg2+versus Ca2+-induced differences in catalytic site structure and function. AB - Refolding together the expressed alpha and beta subunits of the Rhodospirillum rubrum F(1)(RF(1))-ATPase led to assembly of only alpha(1)beta(1) dimers, showing a stable low MgATPase activity. When incubated in the presence of AlCl(3), NaF and either MgAD(T)P or CaAD(T)P, all dimers associated into closed alpha(3)beta(3) hexamers, which also gained a low CaATPase activity. Both hexamer ATPase activities exhibited identical rates and properties to the open dimer MgATPase. These results indicate that: a) the hexamer, as the dimer, has no catalytic cooperativity; b) aluminium fluoride does not inhibit their MgATPase activity; and c) it does enable the assembly of RrF(1)-alpha(3)beta(3) hexamers by stabilizing their noncatalytic alpha/beta interfaces. Refolding of the RrF(1) alpha and beta subunits together with the spinach chloroplast F(1) (CF(1))-gamma enabled a simple one-step assembly of two different hybrid RrF(1) alpha(3)beta(3)/CF(1)gamma complexes, containing either wild type RrF(1)-beta or the catalytic site mutant RrF(1)beta-T159S. They exhibited over 100-fold higher CaATPase and MgATPase activities than the stabilized hexamers and showed very different catalytic properties. The hybrid wild type MgATPase activity was, as that of RrF(1) and CF(1) and unlike its higher CaATPase activity, regulated by excess free Mg(2+) ions, stimulated by sulfite, and inhibited by azide. The hybrid mutant had on the other hand a low CaATPase but an exceptionally high MgATPase activity, which was much less sensitive to the specific MgATPase effectors. All these very different ATPase activities were regulated by thiol modulation of the hybrid unique CF(1)-gamma disulfide bond. These hybrid complexes can provide information on the as yet unknown factors that couple ATP binding and hydrolysis to both thiol modulation and rotational motion of their CF(1)-gamma subunit. PMID- 11278352 TI - Identification of a truncated form of the G-protein regulator AGS3 in heart that lacks the tetratricopeptide repeat domains. AB - AGS3, a 650-amino acid protein encoded by an approximately 4-kilobase (kb) mRNA enriched in rat brain, is a Galpha(i)/Galpha(t)-binding protein that competes with Gbetagamma for interaction with Galpha(GDP) and acts as a guanine nucleotide dissociation inhibitor for heterotrimeric G-proteins. An approximately 2-kb AGS3 mRNA (AGS3-SHORT) is enriched in rat and human heart. We characterized the heart enriched mRNA, identified the encoded protein, and determined its ability to interact with and regulate the guanine nucleotide-binding properties of G proteins. Screening of a rat heart cDNA library, 5'-rapid amplification of cDNA ends, and RNase protection assays identified two populations of cDNAs (1979 and 2134 nucleotides plus the polyadenylation site) that diverged from the larger 4 kb mRNA (AGS3-LONG) in the middle of the protein coding region. Transfection of COS-7 cells with AGS3-SHORT cDNAs resulted in the expression of a major immunoreactive AGS3 polypeptide (M(r) approximately 23,000) with a translational start site at Met(495) of AGS3-LONG. Immunoblots indicated the expression of the M(r) approximately 23,000 polypeptide in rat heart. Glutathione S-transferase AGS3-SHORT selectively interacted with the GDP-bound versus guanosine 5'-O-(3 thiotriphosphate) (GTPgammaS)-bound conformation of Galpha(i2) and inhibited GTPgammaS binding to Galpha(i2). Protein interaction assays with glutathione S transferase-AGS3-SHORT and heart lysates indicated interaction of AGS3-SHORT with Galpha(i1/2) and Galpha(i3), but not Galpha(s) or Galpha(q). Immunofluorescent imaging and subcellular fractionation following transient expression of AGS3 SHORT and AGS3-LONG in COS-7 and Chinese hamster ovary cells indicated distinct subcellular distributions of the two forms of AGS3. Thus, AGS3 exists as a short and long form, both of which apparently stabilize the GDP-bound conformation of Galpha(i), but which differ in their tissue distribution and trafficking within the cell. PMID- 11278353 TI - Regulation of Stat3 activation by MEK kinase 1. AB - Stat3 is a latent transcription factor activated by various cytokines and growth factors. Phosphorylation on Tyr-705 is a prerequisite for dimer formation, nuclear translocation, binding to its cognate DNA sequences, and regulation of the target gene transcription. Ser-727 phosphorylation of Stat3 plays an additional role in the regulation of transcription. MEK kinase 1 (MEKK1) is a mitogen-activated protein kinase (MAPK) kinase kinase (MAPKKK) that activates the c-Jun NH(2)-terminal kinase signaling pathway. Here we report that MEKK1 is involved in the regulation of Stat3 activation by growth factors. Kinase-inactive MEKK1 inhibits Stat3 phosphorylation on tyrosine and serine, and its transcriptional activity stimulated by epidermal growth factor and platelet derived growth factor in different cell types. In contrast, active MEKK1 induces Stat3 tyrosine and serine phosphorylation leading to a functionally active Stat3 capable of binding DNA and enhancing transcription. Ser-727 is phosphorylated by MEKK1 in vitro, whereas Tyr-705 phosphorylation induced by MEKK1 involves Src and Janus kinases in vivo. These data demonstrate for the first time a novel role of MEKK1 to modulate tyrosine kinases that results in the activation of specific members of STAT family. PMID- 11278354 TI - Reassessment of the Ca2+ sensing property of a type I metabotropic glutamate receptor by simultaneous measurement of inositol 1,4,5-trisphosphate and Ca2+ in single cells. AB - Transient transfection of Chinese hamster ovary or baby hamster kidney cells expressing the Group I metabotropic glutamate receptor mGlu1alpha with green fluorescent protein-tagged pleckstrin homology domain of phospholipase Cdelta1 allows real-time detection of inositol 1,4,5-trisphosphate. Loading with Fura-2 enables simultaneous measurement of intracellular Ca(2+) within the same cell. Using this technique we have studied the extracellular calcium sensing property of the mGlu1alpha receptor. Quisqualate, in extracellular medium containing 1.3 mm Ca(2+), increased inositol 1,4,5-trisphosphate in all cells. This followed a typical peak and plateau pattern and was paralleled by concurrent increases in intracellular Ca(2+) concentration. Under nominally Ca(2+)-free conditions similar initial peaks in inositol 1,4,5-trisphosphate and Ca(2+) concentration occurred with little change in either agonist potency or efficacy. However, sustained inositol 1,4,5-trisphosphate production was substantially reduced and the plateau in Ca(2+) concentration absent. Depletion of intracellular Ca(2+) stores using thapsigargin abolished quisqualate-induced increases in intracellular Ca(2+) and markedly reduced inositol 1,4,5-trisphosphate production. These data suggest that the mGlu1alpha receptor is not a calcium sensing receptor because the initial response to agonist is not sensitive to extracellular Ca(2+) concentration. However, prolonged activation of phospholipase C requires extracellular Ca(2+), while the initial burst of activity is highly dependent on Ca(2+) mobilization from intracellular stores. PMID- 11278355 TI - Insulin-mediated GLUT4 translocation is dependent on the microtubule network. AB - The GLUT4 facilitative glucose transporter is recruited to the plasma membrane by insulin. This process depends primarily on the exocytosis of a specialized pool of vesicles containing GLUT4 in their membranes. The mechanism of GLUT4 vesicle exocytosis in response to insulin is not understood. To determine whether GLUT4 exocytosis is dependent on intact microtubule network, we measured insulin mediated GLUT4 exocytosis in 3T3-L1 adipocytes in which the microtubule network was depolymerized by pretreatment with nocodazole. Insulin-mediated GLUT4 translocation was inhibited by more than 80% in nocodazole-treated cells. Phosphorylation of insulin receptor substrate 1 (IRS-1), activation of IRS-1 associated phosphatidylinositide 3-kinase, and phosphorylation of protein kinase B/Akt-1 were not inhibited by nocodazole treatment indicating that the microtubule network was not required for proximal insulin signaling. An intact microtubule network is specifically required for insulin-mediated GLUT4 translocation since nocodazole treatment did not affect insulin-mediated GLUT1 translocation or adipsin secretion. By using in vitro microtubule binding, we demonstrated that both GLUT4 vesicles and IRS-1 bind specifically to microtubules, implicating microtubules in both insulin signaling and GLUT4 translocation. Vesicle binding to microtubules was not mediated through direct binding of GLUT4 or insulin-responsive aminopeptidase to microtubules. A model microtubule-dependent translocation of GLUT4 is proposed. PMID- 11278356 TI - Endoplasmic reticulum (ER)-associated degradation of T cell receptor subunits. Involvement of ER-associated ubiquitin-conjugating enzymes (E2s). AB - Degradation of proteins from the endoplasmic reticulum is fundamental to quality control within the secretory pathway, serves as a way of regulating levels of crucial proteins, and is utilized by viruses to enhance pathogenesis. In yeast two ubiquitin-conjugating enzymes (E2s), UBC6p and UBC7p are implicated in this process. We now report the characterization of murine homologs of these E2s. MmUBC6 is an integral membrane protein that is anchored via its hydrophobic C terminal tail to the endoplasmic reticulum. MmUBC7, which is not an integral membrane protein, shows significant endoplasmic reticulum colocalization with MmUBC6. Overexpression of catalytically inactive MmUBC7 significantly delayed degradation from the endoplasmic reticulum of two T cell antigen receptor subunits, alpha and CD3-delta, and suggests a role for the ubiquitin conjugating system at the initiation of retrograde movement from the endoplasmic reticulum. These findings also implicate, for the first time, a specific E2 in degradation from the endoplasmic reticulum in mammalian cells. PMID- 11278357 TI - Interferon gamma (IFNgamma ) and tumor necrosis factor alpha synergism in ME-180 cervical cancer cell apoptosis and necrosis. IFNgamma inhibits cytoprotective NF kappa B through STAT1/IRF-1 pathways. AB - We investigated the molecular mechanism of the synergism between interferon gamma (IFNgamma) and tumor necrosis factor alpha (TNFalpha) documented in a variety of biological occasions such as tumor cell death and inflammatory responses. IFNgamma/TNFalpha synergistically induced apoptosis of ME-180 cervical cancer cells. IFNgamma induced STAT1 phosphorylation and interferon regulatory factor 1 (IRF-1) expression. Transfection of phosphorylation-defective STAT1 inhibited IFNgamma/TNFalpha-induced apoptosis, whereas IRF-1 transfection induced susceptibility to TNFalpha. Dominant-negative IkappaBalpha transfection sensitized ME-180 cells to TNFalpha. IFNgamma pretreatment attenuated TNFalpha- or p65-induced NF-kappaB reporter activity, whereas it did not inhibit p65 translocation or DNA binding of NF-kappaB. IRF-1 transfection alone inhibited TNFalpha-induced NF-kappaB activity, which was reversed by coactivator p300 overexpression. Caspases were activated by IFNgamma/TNFalpha combination; however, caspase inhibition did not abrogate IFNgamma/TNFalpha-induced cell death. Instead, caspase inhibitors directed IFNgamma/TNFalpha-treated ME-180 cells to undergo necrosis, as demonstrated by Hoechst 33258/propidium iodide staining and electron microscopy. Taken together, our results indicate that IFNgamma and TNFalpha synergistically act to destroy ME-180 tumor cells by either apoptosis or necrosis, depending on caspase activation, and STAT1/IRF-1 pathways initiated by IFNgamma play a critical role in IFNgamma/TNFalpha synergism by inhibiting cytoprotective NF-kappaB. IFNgamma/TNFalpha synergism appears to activate cell death machinery independently of caspase activation, and caspase activation seems to merely determine the mode of cell death. PMID- 11278358 TI - Hypochlorite-modified low density lipoprotein inhibits nitric oxide synthesis in endothelial cells via an intracellular dislocalization of endothelial nitric oxide synthase. AB - Hypochlorous acid/hypochlorite, generated by the myeloperoxidase/H(2)O(2)/halide system of activated phagocytes, has been shown to oxidize/modify low density lipoprotein (LDL) in vitro and may be involved in the formation of atherogenic lipoproteins in vivo. Accordingly, hypochlorite-modified (lipo)proteins have been detected in human atherosclerotic lesions where they colocalize with macrophages and endothelial cells. The present study investigates the influence of hypochlorite-modified LDL on endothelial synthesis of nitric oxide (NO) measured as formation of citrulline (coproduct of NO) and cGMP (product of the NO activated soluble guanylate cyclase) upon cell stimulation with thrombin or ionomycin. Pretreatment of human umbilical vein endothelial cells with hypochlorite-modified LDL led to a time- and concentration-dependent inhibition of agonist-induced citrulline and cGMP synthesis compared with preincubation of cells with native LDL. This inhibition was neither due to a decreased expression of endothelial NO synthase (eNOS) nor to a deficiency of its cofactor tetrahydrobiopterin. Likewise, the uptake of l-arginine, the substrate of eNOS, into the cells was not affected. Hypochlorite-modified LDL caused remarkable changes of intracellular eNOS distribution including translocation from the plasma membrane and disintegration of the Golgi location without altering myristoylation or palmitoylation of the enzyme. In contrast, cyclodextrin known to deplete plasma membrane of cholesterol and to disrupt caveolae induced only a disappearance of eNOS from the plasma membrane that was not associated with decreased agonist-induced citrulline and cGMP formation. The present findings suggest that mislocalization of NOS accounts for the reduced NO formation in human umbilical vein endothelial cells treated with hypochlorite-modified LDL and point to an important role of Golgi-located NOS in these processes. We conclude that inhibition of NO synthesis by hypochlorite-modified LDL may be an important mechanism in the development of endothelial dysfunction and early pathogenesis of atherosclerosis. PMID- 11278359 TI - Calorimetric investigations of the structural stability and interactions of colicin B domains in aqueous solution and in the presence of phospholipid bilayers. AB - The effects of pH and temperature on the stability of interdomain interactions of colicin B have been studied by differential-scanning calorimetry, circular dichroism, and fluorescence spectroscopy. The calorimetric properties were compared with those of the isolated pore-forming fragment. The unfolding profile of the full-length toxin is consistent with two endothermic transitions. Whereas peak A (T(m) = 55 degrees C) most likely corresponds to the receptor/translocation domain, peak B (T(m) = 59 degrees C) is associated with the pore-forming domain. By lowering the pH from 7 to 3.5, the transition temperature of peaks A and B are reduced by 25 and 18 degrees C, respectively, due to proton exchange upon denaturation. The isolated pore-forming fragment unfolds at much higher temperatures (T(m) = 65 degrees C) and is stable throughout a wide pH range, indicating that intramolecular interactions between the different colicin B domains result in a less stable protein conformation. In aqueous solution circular dichroism spectra have been used to estimate the content of helical secondary structure of colicin B ( approximately 40%) or its pore-forming fragment ( approximately 80%). Upon heating, the ellipticities at 222 nm strongly decrease at the transition temperature. In the presence of lipid vesicles the differential-scanning calorimetry profiles of the pore-forming fragment exhibit a low heat of transition multicomponent structure. The heat of transition of membrane-associated colicin B (T(m) = 54 degrees C at pH 3.5) is reduced and its secondary structure is conserved even at intermediate temperatures indicating incomplete unfolding due to strong protein-lipid interactions. PMID- 11278360 TI - Cystic fibrosis pathogens activate Ca2+-dependent mitogen-activated protein kinase signaling pathways in airway epithelial cells. AB - Much of the pulmonary disease in cystic fibrosis is associated with polymorphonuclear leukocyte-dominated airway inflammation caused by bacterial infection. Respiratory epithelial cells express the polymorphonuclear chemokine interleukin-8 (IL-8) in response to ligation of asialylated glycolipid receptors, which are increased on damaged or regenerating cells and those with cystic fibrosis transmembrane conductance regulator mutations. Because both Pseudomonas aeruginosa and Staphylococcus aureus, the most common pathogens in cystic fibrosis, bind asialylated glycolipid receptors such as asialoGM1, we postulated that diverse bacteria can activate a common epithelial signaling pathway to elicit IL-8 expression. P. aeruginosa PAO1 but not pil mutants and S. aureus RN6390 but not the agr mutant RN6911 stimulated increases in [Ca(2+)](i) in 1HAEo airway epithelial cells. This response stimulated p38 and ERK1/2 mitogen activated protein kinase (MAPK) signaling cascades resulting in NF-kappaB activation and IL-8 expression. Ligation of the asialoGM1 receptor or thapsigargin-elicited Ca(2+) release activated this pathway, whereas P. aeruginosa lipopolysaccharide did not. The rapid kinetics of epithelial activation precluded bacterial invasion of the epithelium. Recognition of asialylated glycolipid receptors on airway epithelial cells provides a common pathway for Gram-positive and Gram-negative organisms to initiate an epithelial inflammatory response. PMID- 11278361 TI - Repressor element silencing transcription factor/neuron-restrictive silencing factor (REST/NRSF) can act as an enhancer as well as a repressor of corticotropin releasing hormone gene transcription. AB - The repressor element-1/neuron-restrictive silencing element (RE-1/NRSE) mediates transcriptional repression by the repressor element silencing transcription factor/neuron-restrictive silencing factor (REST/NRSF) in many neuron-specific genes. REST/NRSF is expressed most highly in non-neural tissues, where it is thought to repress gene transcription, but is also found in developing neurons and at low levels in the brain. Its null mutation in vivo results in embryonic lethality in mice. While the RE-1/NRSE-mediated repressive influence of REST/NRSF is well established, results in transgenic studies have suggested that the action of the system is more complex. Here, we report that transcription of the corticotropin releasing hormone (CRH) gene is regulated by REST/NRSF, in part through the RE-1/NRSE. Expression of transfected Crh-luciferase constructs was down-regulated by REST/NRSF in a RE-1/NRSE-dependent fashion in both muscle derived L6 and REST/NRSF co-transfected neuronal PC12 cells. Treatment of L6 cells with trichostatin A revealed that REST/NRSF repression depends, in part, on histone deacetylase activity in these cells. In another neuronal cell line, NG108, REST/NRSF also repressed expression from constructs containing an intact RE-1/NRSE. However, unexpectedly, REST/NRSF up-regulated expression levels of constructs lacking an intact RE-1/NRSE. These results suggest that REST/NRSF can act as both a repressor of Crh transcription, via the Crh RE-1/NRSE, and an enhancer of Crh transcription, via a mechanism independent of the Crh RE-1/NRSE. PMID- 11278362 TI - p21-activated protein kinase gamma-PAK suppresses programmed cell death of BALB3T3 fibroblasts. AB - In response to stress stimulants, cells activate opposing signaling pathways for cell survival and programmed cell death. p21-activated protein kinase gamma-PAK is involved in both cell survival and cell death pathways. Many stress stimulants activate gamma-PAK as a full-length enzyme and as a proteolytic fragment. Caspase mediated proteolytic activation parallels cell death and appears to be a pro apoptotic factor in stress-induced cell death. Here, we show that activation of full-length gamma-PAK promotes cell survival and suppresses stress-induced cell death. Expression of constitutively active gamma-PAK-T402E, which mimics activated full-length gamma-PAK, stimulates cell survival of BALB3T3 fibroblasts in response to tumor necrosis factor alpha, growth factor withdrawal, and UVC light. This stimulation of cell survival is mainly due to protection of cells from cell death rather than by stimulation of proliferation. Expression of gamma PAK-T402E increases phosphorylation of the pro-apoptotic Bcl-2 family protein Bad and protects from cell death induced by ectopic expression of Bad. In response to tumor necrosis factor alpha, expression of gamma-PAK-T402E increases the early but reduces the late activation of ERK, JNK, and p38. Our results indicate that the ubiquitous gamma-PAK may have a crucial function in cell survival by regulating the pro-apoptotic activity of Bad and the stress-induced activation of ERK, JNK, and p38 pathways. PMID- 11278363 TI - Interactions of the human T-cell leukemia virus type-II integrase with the conserved CA in the retroviral long terminal repeat end. AB - Retroviral integrases (INs) interact with termini of retroviral DNA in the conserved 5'-C(A/G)T. For most integrases, modifications of critical moieties in the major and minor grooves of these sequences decrease 3'-processing. However, for human immunodeficiency virus type-2 (HTLV-2) IN, the replacement of the guanine with 6-methylguanine or hypoxanthine not only reduced 3'-processing, but also promoted cleavage at a second site. This novel cleavage activity required an upstream ACA, unique to the HTLV-2 U5 end. 3'-Processing assays with additional isosteric modifications at Gua and filter binding experiments revealed that the mechanism of the second site cleavage differed among the major groove, minor groove, and mismatch modifications. Importantly, the decrease in 3'-processing activity noted with the minor groove and mismatch modifications were attributed to a decrease in binding. Major groove modifications, however, decreased the level of 3'-processing, but did not affect binding. This suggests that integrase binds the viral end through the minor groove, but relies on major groove contacts for 3'-processing. Several modifications were also examined in strand transfer and disintegration substrates. HTLV-2 IN showed reduced activity with strand transfer and disintegration substrates containing major groove, but not minor groove modifications. This suggests major groove interactions at guanine also provide an important role in these reactions. PMID- 11278364 TI - Rapamycin-insensitive regulation of 4e-BP1 in regenerating rat liver. AB - In cultured cells, growth factor-induced phosphorylation of two translation modulators, p70 S6 kinase and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1), is blocked by nanomolar concentrations of the immunosuppressant rapamycin. Rapamycin also attenuates liver regeneration after partial hepatectomy, but it is not known if this growth-suppressive effect is due to dephosphorylation of p70 S6 kinase and/or 4E-BP1. We found that partial hepatectomy induced a transient increase in liver p70 S6 kinase activity and 4E BP1 phosphorylation as compared with sham-operated rats. The amount of p70 S6 kinase protein in regenerating liver did not increase, but active kinase from partially hepatectomized animals was highly phosphorylated. Phosphorylated 4E-BP1 from regenerating liver was unable to form an inhibitory complex with initiation factor 4E. Rapamycin blocked the activation of p70 S6 kinase in response to partial hepatectomy in a dose-dependent manner, but 4E-BP1 phosphorylation was not inhibited. By contrast, functional phosphorylation of 4E-BP1 induced by injection of cycloheximide or growth factors was partially reversed by the drug. The mammalian target of rapamycin (mTOR) has been proposed to directly phosphorylate 4E-BP1. Western blot analysis using phospho-specific antibodies showed that phosphorylation of Thr-36/45 and Ser-64 increased in response to partial hepatectomy in a rapamycin-resistant manner. Thus, rapamycin inhibits p70 S6 kinase activation and liver regeneration, but not functional phosphorylation of 4E-BP1, in response to partial hepatectomy. These results indicate that the effect of rapamycin on 4E-BP1 function in vivo can be significantly different from its effect in cultured cells. PMID- 11278365 TI - Identification of the anti-angiogenic site within vascular basement membrane derived tumstatin. AB - Components of vascular basement membrane are involved in regulating angiogenesis. Recently, tumstatin (the NC1 domain of alpha3 chain of type IV collagen) was identified as possessing anti-angiogenic activity. In the present study, the anti angiogenic activity of tumstatin was localized to the putative 54-132-amino acid Tum-5 domain, and the activity mediated by alpha(v)beta(3) integrin interaction in an RGD-independent manner. The recombinant Tum-5 produced in Escherichia coli and Pichia Pastoris specifically inhibited proliferation and caused apoptosis of endothelial cells with no significant effect on nonendothelial cells. Tum-5 also inhibited tube formation of endothelial cells on Matrigel and induced G1 endothelial cell cycle arrest. Moreover, anti-angiogenic effect of Tum-5 was also examined in vivo using both a Matrigel plug assay in C57BL/6 mice and human prostate cancer (PC-3) xenografts in nude mice. The in vivo results demonstrate that Tum-5 at 1 mg/kg significantly inhibited growth of PC-3 tumors in association with a decrease in CD31 positive vasculature. These in vivo studies also show that, at molar equivalents, human Tum-5 is at least 10-fold more active than human endostatin. In addition, these studies for the first time suggest that through the action of endogenous inhibitors, alpha(v)beta(3) integrin may also function as a negative regulator of angiogenesis. Taken together, these findings demonstrate that Tum-5, a domain derived from tumstatin, is an effective inhibitor of tumor-associated angiogenesis and a promising candidate for the treatment of cancer. PMID- 11278366 TI - Tumor suppressor MMAC/PTEN inhibits cytokine-induced NFkappaB activation without interfering with the IkappaB degradation pathway. AB - The phosphoinositide 3-kinase (PI 3-kinase) pathway has been implicated in the activation of the proinflammatory transcription factor nuclear factor kappaB (NFkappaB). To investigate the role of this pathway in NFkappaB activation, we employed mutated in multiple advanced cancers/phosphatase and tensin homologue (MMAC/PTEN), a natural antagonist of PI 3-kinase activity. Our results show that cytokine-induced DNA binding and transcriptional activities of NFkappaB were both inhibited in a glioma cell line that was stably transfected with MMAC/PTEN. The ability of interleukin-1 (IL-1) to induce inhibitor (IkappaB) degradation or nuclear translocation of NFkappaB was, however, unaffected by MMAC/PTEN expression, suggesting that PI 3-kinase utilizes another equally important mechanism to control NFkappaB activation. It is conceivable that NFkappaB is directly phosphorylated through such a mechanism because treatment with protein phosphatase 2A significantly reduced its DNA binding activity. Moreover, IL-1 induced phosphorylation of p50 NFkappaB was potently inhibited in MMAC/PTEN expressing cells. Whereas the mediators of NFkappaB phosphorylation remain to be identified, IL-1 was found to induce physical interactions between the PI 3 kinase target Akt kinase and the IkappaB.IkappaB kinase complex. Physical interactions between these proteins were antagonized by MMAC/PTEN consistent with their potential involvement in NFkappaB activation. Taken together, our observations suggest that PI 3-kinase regulates NFkappaB activation through a novel phosphorylation-dependent mechanism. PMID- 11278367 TI - Identification and characterization of a novel nuclear factor of activated T cells-1 isoform expressed in mouse brain. AB - The nuclear factor of activated T-cells (NFAT) family transcription factors play a key role in the control of cytokine gene expression in T-cells. Although initially identified in T-cells, recent data have unveiled unanticipated roles for NFATs in the development, proliferation, and differentiation of other tissues. Here we report the identification, cDNA cloning, and functional characterization of a new isoform of NFAT1 highly expressed in mouse brain. This isoform, which we named NFAT1-D, is identical to NFAT1 throughout the N-terminal regulatory domain and the portion of the Rel domain which includes the minimal region required for specific binding to DNA and interaction with AP-1. The homology stops sharply upstream of the 3'-boundary of the Rel homology domain and is followed by a short unique C-terminal region. NFAT1-D was expressed at high levels in all brain districts and was found as a constitutively active transcription complex. Transfection of a NFAT/luciferase reporter in the neuronal cell line PC12, which also expresses NFAT1-D, showed that these cells expressed a constitutive NFAT activity that was enhanced after nerve growth factor-induced differentiation but was resistant to the immunosuppressant cyclosporin A. NFAT1-D was, however, inducibly activated in a cyclosporin A-sensitive manner when expressed in T-cells, suggesting that the activity of NFAT proteins might be controlled by their specific cellular context. PMID- 11278368 TI - HIV-1 Tat protein interacts with mammalian capping enzyme and stimulates capping of TAR RNA. AB - HIV gene expression is subject to a transcriptional checkpoint, whereby negative transcription elongation factors induce an elongation block that is overcome by HIV Tat protein in conjunction with P-TEFb. P-TEFb is a cyclin-dependent kinase that catalyzes Tat-dependent phosphorylation of Ser-5 of the Pol II C-terminal domain (CTD). Ser-5 phosphorylation confers on the CTD the ability to recruit the mammalian mRNA capping enzyme (Mce1) and stimulate its guanylyltransferase activity. Here we show that Tat spearheads a second and novel pathway of capping enzyme recruitment and activation via a direct physical interaction between the C terminal domain of Tat and Mce1. Tat stimulates the guanylyltransferase and triphosphatase activities of Mce1 and thereby enhances the otherwise low efficiency of cap formation on a TAR stem-loop RNA. Our findings suggest that multiple mechanisms exist for coupling transcription elongation and mRNA processing. PMID- 11278369 TI - Examination of donor substrate conversion in yeast transketolase. AB - The cleavage of the donor substrate d-xylulose 5-phosphate by wild-type and H263A mutant yeast transketolase was studied using enzyme kinetics and circular dichroism spectroscopy. The enzymes are able to catalyze the cleavage of donor substrates, the first half-reaction, even in the absence of any acceptor substrate yielding d-glyceraldehyde 3-phosphate as measured in the coupled optical test according to Kochetov (Kochetov, G. A. (1982) Methods Enzymol. 90, 209-223) and compared with the H263A variant. Overall, the H263A mutant enzyme is less active than the wild-type. However, an increase in the rate constant of the release of the enzyme-bound glycolyl moiety was observed and related to a stabilization of the "active glycolaldehyde" (alpha-carbanion) by histidine 263. Chemically synthesized dl-(alpha,beta-dihydroxyethyl)thiamin diphosphate is bound to wild-type transketolase with an apparent K(D) of 4.3 +/- 0.8 microm (racemate) calculated from titration experiments using circular dichroism spectroscopy. Both enantiomers are cleaved by the enzyme at different rates. In contrast to the enzyme-generated alpha-carbanion of (alpha,beta-dihydroxyethyl)thiamin diphosphate formed by decarboxylation of hydroxylactylthiamin diphosphate after incubation of transketolase with beta-hydroxypyruvate, the synthesized dl (alpha,beta-dihydroxyethyl)thiamin diphosphate did not work as donor substrate when erythrose 4-phosphate is used as acceptor substrate in the coupled enzymatic test according to Sprenger (Sprenger, G. A., Schorken, U., Sprenger, G., and Sahm, H. (1995) Eur. J. Biochem. 230, 525-532). PMID- 11278370 TI - Vanadate facilitates interferon alpha-mediated apoptosis that is dependent on the Jak/Stat pathway. AB - Type I interferon (IFN)-dependent inhibition of cell growth can occur either in the absence or presence of apoptosis. The mechanisms that determine whether or not cells undergo apoptosis after exposure to IFN-alpha are not clear. This study shows that a variety of cell lines that display growth inhibition but not apoptosis in response to IFN-alpha will undergo programmed cell death when low concentrations of the protein-tyrosine phosphatase inhibitor vanadate are added with IFN-alpha. In contrast, the combination of tumor necrosis factor-alpha with vanadate did not trigger apoptosis in these cells. Caspase-3 activity was detected only in cells exposed to IFN-alpha and vanadate but not to IFN-alpha or vanadate alone. The ability of IFN-alpha and vanadate to induce apoptosis did not require expression of p53 and was blocked by N-acetyl-l-cysteine. Activation of the Jak/Stat pathway and expression of IFN-inducible genes was not altered by incubation of cells with IFN-alpha and vanadate compared with IFN-alpha alone. However, mutant cells lacking Stat1, Stat2, Jak1, or Tyk2, or cells expressing kinase inactive Jak1 or Tyk2 did not undergo apoptosis in the presence of IFN alpha and vanadate. These results suggest that IFN-alpha stimulation of Stat dependent genes is necessary, but not sufficient, for this cytokine to induce apoptosis. Another signaling cascade that involves the activity of a protein tyrosine phosphatase and/or the generation of reactive oxygen species may play an important role in promoting IFN-alpha-induced apoptosis. PMID- 11278371 TI - Two domains of the human bZIP transcription factor TCF11 are necessary for transactivation. AB - TCF11 is a bZIP transcription factor of the CNC subfamily. It has been implicated in the regulation of the antioxidant response and is vital during embryonic development, but its precise biological functions have not yet been fully worked out. Structural characterization of the gene and several of its products indicates that complex regulatory mechanisms are employed. To investigate how altering the structure of the gene products might influence their activity we have mapped functional domains within the protein. We show that two separate domains are required for transactivation by full-length TCF11: an N-terminal acidic domain and a serine-rich stretch adjacent to the CNC-bZIP domains. A naturally occurring shorter isoform (identical to LCR-F1) produced by internal initiation of translation is unable to transactivate in our assay. However, the shorter form could interfere with the transactivating ability of the longer form, which indicates a control mechanism for keeping the activity of TCF11 at a desired level. We show that TCF11 and the closely related CNC-bZIP factor p45 NF E2 show different cell type-specific activation patterns with full-length TCF11 being active in COS-1 cells but silent in erythroid cells (K562), whereas p45 NF E2 is active in K562 cells and silent in COS-1 cells. Domain swapping experiments show that cell type-specific activity is not fully determined by dimerization/DNA binding domains or transactivation domains alone. The resulting profile of activity is most likely achieved by interaction of the domains and their cell specific environment. PMID- 11278372 TI - Stabilization and activation of p53 by the coactivator protein TAFII31. AB - Regulation of the stability of p53 is key to its tumor-suppressing activities. mdm2 directly binds to the amino-terminal region of p53 and targets it for degradation through the ubiquitin-proteasome pathway. The coactivator protein TAF(II)31 binds to p53 at the amino-terminal region that is also required for interaction with mdm2. In this report, we demonstrate that expression of TAF(II)31 inhibits mdm2-mediated ubiquitination of p53 and increases p53 levels. TAF(II)31-mediated p53 stabilization results in activation of p53-mediated transcriptional activity and leads to p53-dependent growth arrest in fibroblasts. UV-induced stabilization of p53 coincides with an increase in p53-associated TAF(II)31 and a corresponding decrease in mdm2-p53 interaction. Non-p53 binding mutant of TAF(II)31 fails to stabilize p53. Our results suggest that direct interaction of TAF(II)31 and p53 not only mediates p53 transcriptional activation but also protects p53 from mdm2-mediated degradation, thereby resulting in activation of p53 functions. PMID- 11278373 TI - Forkhead family transcription factor FKHRL1 is expressed in human megakaryocytes. Regulation of cell cycling as a downstream molecule of thrombopoietin signaling. AB - FKHRL1, a member of the Forkhead transcription factor family, is one of the downstream molecules of phosphatidylinositol 3-kinase-Akt. This molecule is a mammalian homolog of DAF-16, which plays an important role in the longevity of Caenorhabditis elegans. In this study we found that Akt and FKHRL1 proteins were detectable in highly purified normal human megakaryocytes and that these molecules were actually phosphorylated by thrombopoietin (TPO). To clarify the functional role of FKHRL1 in TPO signaling, we established a tetracycline inducible system in the human TPO-dependent leukemia cell line UT-7/TPO. Induced expression of active FKHRL1 led to cell cycle arrest at G0/G1 phase in this cell line. These results suggest that FKHRL1 plays an important role in the cell cycle of megakaryocytic cells as one of the downstream target molecules of phosphatidylinositol 3-kinase-Akt, presumably mediated through the activation or inactivation of cell cycle-associated gene(s). PMID- 11278374 TI - Degradation of membrane-bound ganglioside GM2 by beta -hexosaminidase A. Stimulation by GM2 activator protein and lysosomal lipids. AB - According to a recent hypothesis, glycosphingolipids originating from the plasma membrane are degraded in the acidic compartments of the cell as components of intraendosomal and intralysosomal vesicles and structures. Since most previous in vitro investigations used micellar ganglioside GM2 as substrate, we studied the degradation of membrane-bound ganglioside GM2 by water-soluble beta hexosaminidase A in the presence of the GM2 activator protein in a detergent free, liposomal assay system. Our results show that anionic lipids such as the lysosomal components bis(monoacylglycero)phosphate or phosphatidylinositol stimulate the degradation of GM2 by beta-hexosaminidase A up to 180-fold in the presence of GM2 activator protein. In contrast, the degradation rate of GM2 incorporated into liposomes composed of neutral lysosomal lipids such as dolichol, cholesterol, or phosphatidylcholine was significantly lower than in negatively charged liposomes. This demonstrates that both, the GM2 activator protein and anionic lysosomal phospholipids, are needed to achieve a significant degradation of membrane-bound GM2 under physiological conditions. The interaction of GM2 activator protein with immobilized membranes was studied with surface plasmon resonance spectroscopy at an acidic pH value as it occurs in the lysosomes. Increasing the concentration of bis(monoacylglycero)phosphate in immobilized liposomes led to a significant drop of the resonance signal in the presence of GM2 activator protein. This suggests that in the presence of bis(monoacylglycero)phosphate, which has been shown to occur in inner membranes of the acidic compartment, GM2 activator protein is able to solubilize lipids from the surface of immobilized membrane structures. PMID- 11278375 TI - Complete cysteine-scanning mutagenesis and site-directed chemical modification of the Tn10-encoded metal-tetracycline/H+ antiporter. AB - Bacterial Tn10-encoded metal-tetracycline/H(+) antiporter was the first found drug exporter and has been studied as a paradigm of antiporter-type major facilitator superfamily transporters. Here the 400 amino acid residues of this protein were individually replaced by cysteine except for the initial methionine. As a result, we could obtain a complete map of the functionally or structurally important residues. In addition, we could determine the precise boundaries of all the transmembrane segments on the basis of the reactivity with N-ethylmaleimide (NEM). The NEM binding results indicated the presence of a transmembrane water filled channel in the transporter. The twelve transmembrane segments can be divided into three groups; four are totally embedded in the hydrophobic interior, four face a putative water-filled channel along their full length, and the remaining four face the channel for half their length, the other halves being embedded in the hydrophobic interior. These three types of transmembrane segments are mutually arranged with a 4-fold symmetry. The competitive binding of membrane permeable and -impermeable SH reagents in intact cells indicates that the transmembrane water-filled channel has a thin barrier against hydrophilic molecules in the middle of the transmembrane region. Inhibition and stimulation of NEM binding in the presence of tetracycline reflects the substrate-induced protection or conformational change of the Tn10-encoded metal-tetracycline/H(+) antiporter. The mutations protected from NEM binding by tetracycline were mainly located around the permeability barrier in the N-terminal half, suggesting the location of the substrate binding site. PMID- 11278376 TI - A full biological response to autoantibodies in Graves' disease requires a disulfide-bonded loop in the thyrotropin receptor N terminus homologous to a laminin epidermal growth factor-like domain. AB - We observed amino acid homology between the cysteine-rich N terminus of the thyrotropin receptor (TSHR) ectodomain and epidermal growth factor-like repeats in the laminin gamma1 chain. Thyroid-stimulating autoantibodies (TSAb), the cause of Graves' disease, interact with this region of the TSHR in a manner critically dependent on antigen conformation. We studied the role of the cluster of four cysteine (Cys) residues in this region of the TSHR on the functional response to TSAb in Graves' patients' sera. As a benchmark we also studied TSH binding and action. Removal in various permutations of the four cysteines at TSHR positions 24, 29, 31, and 41 (signal peptide residues are 1-21) revealed Cys(41) to be the key residue for receptor expression. Forced pairing of Cys(41) with any one of the three upstream Cys residues was necessary for trafficking to the cell surface of a TSHR with high affinity TSH binding similar to the wild-type receptor. However, for a full biological response to TSAb, forced pairing of Cys(41) with Cys(29) or with Cys(31), but not with Cys(24), retained functional activity comparable with the wild-type TSHR. These data suggest that an N-terminal disulfide-bonded loop between Cys(41) and Cys(29) or its close neighbor Cys(31) comprises, in part, the highly conformational epitope for TSAb at the critical N terminus of the TSHR. Amino acid homology, as well as cysteine pairing similar to the laminin gamma1 chain epidermal growth factor-like repeat 11, suggests conformational similarity between the two molecules and raises the possibility of molecular mimicry in the pathogenesis of Graves' disease. PMID- 11278377 TI - The regulation of uncoupling protein-2 gene expression by omega-6 polyunsaturated fatty acids in human skeletal muscle cells involves multiple pathways, including the nuclear receptor peroxisome proliferator-activated receptor beta. AB - Fatty acids have been postulated to regulate uncoupling protein (UCP) gene expression in skeletal muscle in vivo. We have identified, at least in part, the mechanism by which polyunsaturated fatty acids increase UCP-2 expression in primary culture of human muscle cells. omega-6 fatty acids and arachidonic acid induced a 3-fold rise in UCP-2 mRNA levels possibly through transcriptional activation. This effect was prevented by indomethacin and mimicked by prostaglandin (PG) E(2) and carbaprostacyclin PGI(2), consistent with a cyclooxygenase-mediated process. Incubation of myotubes for 6 h with 100 micrometer arachidonic acid resulted in a 150-fold increase in PGE(2) and a 15 fold increase in PGI(2) in the culture medium. Consistent with a role of cAMP and protein kinase A, both prostaglandins induced a marked accumulation of cAMP in human myotubes, and forskolin reproduced the effect of arachidonic acid on UCP-2 mRNA expression. Inhibition of protein kinase A with H-89 suppressed the effect of PGE(2), whereas cPGI(2) and arachidonic acid were still able to increase ucp-2 gene expression, suggesting additional mechanisms. We found, however, that the MAP kinase pathway was not involved. Prostaglandins, particularly PGI(2), are potent activators of the peroxisome proliferator-activated receptors. A specific agonist of peroxisome proliferator-activated receptor (PPAR) beta (L165041) increased UCP-2 mRNA levels in myotubes, whereas activation of PPARalpha or PPARgamma was ineffective. These results suggest thus that ucp-2 gene expression is regulated by omega-6 fatty acids in human muscle cells through mechanisms involving at least protein kinase A and the nuclear receptor PPARbeta. PMID- 11278378 TI - alpha 7 nicotinic receptor transduces signals to phosphatidylinositol 3-kinase to block A beta-amyloid-induced neurotoxicity. AB - Multiple lines of evidence, from molecular and cellular to epidemiological, have implicated nicotinic transmission in the pathogenesis of Alzheimer's disease (AD). Here we show the signal transduction mechanism involved in nicotinic receptor-mediated protection against beta-amyloid-enhanced glutamate neurotoxicity. Nicotine-induced protection was suppressed by an alpha7 nicotinic receptor antagonist (alpha-bungarotoxin), a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002 and wortmannin), and a Src inhibitor (PP2). Levels of phosphorylated Akt, an effector of PI3K, and Bcl-2 were increased by nicotine. The alpha7 nicotinic receptor was physically associated with the PI3K p85 subunit and Fyn. These findings indicate that the alpha7 nicotinic receptor transduces signals to PI3K in a cascade, which ultimately contributes to a neuroprotective effect. This might form the basis of a new treatment for AD. PMID- 11278379 TI - Cell surface heparan sulfate proteoglycans participate in factor VIII catabolism mediated by low density lipoprotein receptor-related protein. AB - We have demonstrated previously that catabolism of a coagulation factor VIII (fVIII) from its complex with von Willebrand factor (vWf) is mediated by low density lipoprotein receptor-related protein (LRP) (Saenko, E. L., Yakhyaev, A. V., Mikhailenko, I., Strickland, D. K., and Sarafanov, A. G. (1999) J. Biol. Chem. 274, 37685-37692). In the present study, we found that this process is facilitated by cell surface heparan sulfate proteoglycans (HSPGs). This was demonstrated by simultaneous blocking of LRP and HSPGs in model cells, which completely prevented fVIII internalization and degradation from its complex with vWf. In contrast, the selective blocking of either receptor had a lesser effect. In vivo studies of clearance of (125)I-fVIII-vWf complex in mice also demonstrated that the simultaneous blocking of HSPGs and LRP led to a more significant prolongation of fVIII half-life (5.5-fold) than blocking of LRP alone (3.5-fold). The cell culture and in vivo experiments revealed that HSPGs are also involved in another, LRP-independent pathway of fVIII catabolism. In both pathways, HSPGs act as receptors providing the initial binding of fVIII-vWf complex to cells. We demonstrated that this binding occurs via the A2 domain of fVIII, since A2, but not other portions of fVIII or isolated vWf, strongly inhibited cell surface binding of fVIII-vWf complex, and the affinities of A2 and fVIII-vWf complex for the cells were similar. The A2 site involved in binding to heparin was localized to the region 558-565, based on the ability of the corresponding synthetic peptide to inhibit A2 binding to heparin, used as a model for HSPGs. PMID- 11278380 TI - Regulation of von Willebrand factor binding to the platelet glycoprotein Ib-IX by a membrane skeleton-dependent inside-out signal. AB - The platelet receptor for von Willebrand factor (vWF), glycoprotein Ib-IX (GPIb IX), mediates initial platelet adhesion and activation. We show here that the receptor function of GPIb-IX is regulated intracellularly via its link to the filamin-associated membrane skeleton. Deletion of the filamin binding site in GPIb(alpha) markedly enhances ristocetin- (or botrocetin)-induced vWF binding and allows GPIb-IX-expressing cells to adhere to immobilized vWF under both static and flow conditions. Cytochalasin D (CD) that depolymerizes actin also enhances vWF binding to wild type GPIb-IX. Thus, vWF binding to GPIb-IX is negatively regulated by the filamin-associated membrane skeleton. In contrast to native vWF, binding of the isolated recombinant vWF A1 domain to wild type and filamin binding-deficient mutants of GPIb-IX is comparable, suggesting that the membrane skeleton-associated GPIb-IX is in a state that prevents access to the A1 domain in macromolecular vWF. In platelets, there is a balance of membrane skeleton associated and free forms of GPIb-IX. Treatment of platelets with CD increases the free form and enhances vWF binding. CD also reverses the inhibitory effects of prostaglandin E1 on vWF binding to GPIb-IX. Thus, GPIb-IX-dependent platelet adhesion is doubly controlled by vWF conformation and a membrane skeleton dependent inside-out signal. PMID- 11278381 TI - Sumo-1 modification regulates the DNA binding activity of heat shock transcription factor 2, a promyelocytic leukemia nuclear body associated transcription factor. AB - Heat shock transcription factor 2 (HSF2) is a transcription factor that regulates heat shock protein gene expression, but the mechanisms regulating the function of this factor are unclear. Here we report that HSF2 is a substrate for modification by the ubiquitin-related protein SUMO-1 and that HSF2 colocalizes in cells with SUMO-1 in nuclear granules. Staining with anti-promyelocytic leukemia antibodies indicates that these HSF2-containing nuclear granules are PML bodies. Our results identify lysine 82 as the major site of SUMO-1 modification in HSF2, which is located in a "wing" within the DNA-binding domain of this protein. Interestingly, SUMO-1 modification of HSF2 results in conversion of this factor to the active DNA binding form. This is the first demonstration that SUMO-1 modification can directly alter the DNA binding ability of a transcription factor and reveals a new mechanism by which SUMO-1 modification can regulate protein function. PMID- 11278383 TI - Determination of photosystem II subunits by matrix-assisted laser desorption/ionization mass spectrometry. AB - Photosystem II of higher plants and cyanobacteria is composed of more than 20 polypeptide subunits. The pronounced hydrophobicity of these proteins hinders their purification and subsequent analysis by mass spectrometry. This paper reports the results obtained by application of matrix-assisted laser desorption/ionization mass spectrometry directly to isolated complexes and thylakoid membranes prepared from cyanobacteria and spinach. Changes in protein contents following physiopathological stimuli are also described. Good correlations between expected and measured molecular masses allowed the identification of the main, as well as most of the minor, low molecular weight components of photosystem II. These results open up new perspectives for clarifying the functional role of the various polypeptide components of photosystems and other supramolecular integral membrane complexes. PMID- 11278382 TI - Protein kinase C-associated kinase (PKK), a novel membrane-associated, ankyrin repeat-containing protein kinase. AB - A novel murine membrane-associated protein kinase, PKK (protein kinase C associated kinase), was cloned on the basis of its physical association with protein kinase Cbeta (PKCbeta). The regulated expression of PKK in mouse embryos is consistent with a role for this kinase in early embryogenesis. The human homolog of PKK has over 90% identity to its murine counterpart, has been localized to chromosome 21q22.3, and is identical to the PKCdelta-interacting kinase, DIK (Bahr, C., Rohwer, A., Stempka, L., Rincke, G., Marks, F., and Gschwendt, M. (2000) J. Biol. Chem. 275, 36350-36357). PKK comprises an N terminal kinase domain and a C-terminal region containing 11 ankyrin repeats. PKK exhibits protein kinase activity in vitro and associates with cellular membranes. PKK exists in three discernible forms at steady state: an underphosphorylated form of 100 kDa; a soluble, cytosolic, phosphorylated form of 110 kDa; and a phosphorylated, detergent-insoluble form of 112 kDa. PKK is initially synthesized as an underphosphorylated soluble 100-kDa protein that is quantitatively converted to a detergent-soluble 110-kDa form. This conversion requires an active catalytic domain. Although PKK physically associates with PKCbeta, it does not phosphorylate this PKC isoform. However, PKK itself may be phosphorylated by PKCbeta. PKK represents a developmentally regulated protein kinase that can associate with membranes. The functional significance of its association with PKCbeta remains to be ascertained. PMID- 11278384 TI - Deoxycytidyl transferase activity of the human REV1 protein is closely associated with the conserved polymerase domain. AB - The REV1 protein is a member of the growing family of translesion DNA polymerases. A cDNA of the human REV1 gene that we had originally isolated encoded 1250 amino acids residues, which was one amino acid shorter than previously reported ones. The shorter form of REV1 was named REV1S. All individuals examined expressed equivalent amounts of REV1S and REV1 mRNA, suggesting that the REV1S mRNA is a splicing variant. We show that the REV1S protein also possesses deoxycytidyl transferase activity that inserts a dCMP opposite a DNA template apurinic/apyrimidinic site. Deletion and point mutation analysis of the REV1S protein revealed that the domain required for deoxycytidyl transferase and DNA binding activities of the REV1S protein are located in a conserved domain of translesion DNA polymerases. This result indicates that the structure of the catalytic site of the deoxycytidyl transferase closely resembles that of the translesion DNA polymerases. Therefore, the molecular mechanism of the dCMP transfer reaction of the REV1S protein and maybe also the REV1 protein might be the same as that of the dNTP transfer reaction of the translesion DNA polymerases. PMID- 11278385 TI - Extracellular signal-regulated kinase/90-KDA ribosomal S6 kinase/nuclear factor kappa B pathway mediates phorbol 12-myristate 13-acetate-induced megakaryocytic differentiation of K562 cells. AB - Two signaling pathways, the extracellular signal-regulated kinase (ERK)/mitogen activated protein kinase (MAPK)-dependent pathway and the nuclear factor-kappaB (NF-kappaB)-dependent pathway, have been known to mediate megakaryocytic differentiation of K562 cells induced by phorbol 12-myristate 13-acetate (PMA). In this study, we examined whether 90-kDa ribosomal S6 kinase (RSK), known as a substrate of ERK/MAPK and a signal-inducible IkappaBalpha kinase, would link two pathways during the differentiation. RSK1 was activated in a time- and dose dependent manner during the PMA-induced differentiation. Overexpression of wild type or dominant inhibitory mutant (D205N) of RSK1 enhanced or suppressed PMA stimulated NF-kappaB activation and megakaryocytic differentiation as shown by morphology, nonspecific esterase activity, and expression of the CD41 megakaryocytic marker, respectively. In addition, overexpression of the dominant inhibitory mutant (S32A/S36A) of IkappaBalpha inhibited PMA-stimulated and RSK1 enhanced megakaryocytic differentiation, indicating that NF-kappaB mediates a signal for megakaryocytic differentiation downstream of RSK1. PMA-stimulated activation of ERK/MAPK, RSK1, and NF-kappaB and the PMA-induced megakaryocytic differentiation were prevented by pretreatment with PD98059, a specific inhibitor of the mitogen-activated ERK kinase (MEK). Therefore, these results demonstrate that the sequential ERK/RSK1/NF-kappaB pathway mediates PMA-stimulated megakaryocytic differentiation of K562 cells. PMID- 11278386 TI - A novel role of neuregulin in skeletal muscle. Neuregulin stimulates glucose uptake, glucose transporter translocation, and transporter expression in muscle cells. AB - Neuregulins regulate the expression of acetylcholine receptor genes and induce development of the neuromuscular junction in muscle. In studying whether neuregulins regulate glucose uptake in muscle, we analyzed the effect of a recombinant neuregulin, (r)heregulin-beta1-(177-244) (HRG), on L6E9 muscle cells, which express the neuregulin receptors ErbB2 and ErbB3. L6E9 responded acutely to HRG by a time- and concentration-dependent stimulation of 2-deoxyglucose uptake. HRG-induced stimulation of glucose transport was additive to the effect of insulin. The acute stimulation of the glucose transport induced by HRG was a consequence of the translocation of GLUT4, GLUT1, and GLUT3 glucose carriers to the cell surface. The effect of HRG on glucose transport was dependent on phosphatidylinositol 3-kinase activity. HRG also stimulated glucose transport in the incubated soleus muscle and was additive to the effect of insulin. Chronic exposure of L6E9 cells to HRG potentiated myogenic differentiation, and under these conditions, glucose transport was also stimulated. The activation of glucose transport after chronic HRG exposure was due to enhanced cell content of GLUT1 and GLUT3 and to increased abundance of these carriers at the plasma membrane. However, under these conditions, GLUT4 expression was markedly down regulated. Muscle denervation is associated with GLUT1 induction and GLUT4 repression. In this connection, muscle denervation caused a marked increase in the content of ErbB2 and ErbB3 receptors, which occurred in the absence of alterations in neuregulin mRNA levels. This fact suggests that neuregulins regulate glucose transporter expression in denervated muscle. We conclude that neuregulins regulate glucose uptake in L6E9 muscle cells by mechanisms involving the recruitment of glucose transporters to the cell surface and modulation of their expression. Neuregulins may also participate in the adaptations in glucose transport that take place in the muscle fiber after denervation. PMID- 11278387 TI - Angiotensin II type I receptor modulates intracellular free Mg2+ in renally derived cells via Na+-dependent Ca2+-independent mechanisms. AB - Treatment of Madin-Darby canine kidney (MDCK) cells with the peptide hormone angiotensin II (Ang II) results in an increase in the concentrations of cytosolic free calcium ([Ca(2+)](i)) and sodium ([Na(+)](i)) with a concomitant decrease in cytosolic free Mg(2+) concentration ([Mg(2+)](i)). In the present study we demonstrate that this hormone-induced decrease in [Mg(2+)](i) is independent of [Ca(2+)](i) but dependent on extracellular Na(+). [Mg(2+)](i), [Ca(2+)](i), and [Na(+)](i) were measured in Ang II-stimulated MDCK cells by fluorescence digital imaging using the selective fluoroprobes mag-fura-2AM, fura-2AM, and sodium binding benzofuran isophthalate (acetoxymethyl ester), respectively. Ang II decreased [Mg(2+)](i) and increased [Na(+)](i) in a dose-dependent manner. These effects were inhibited by irbesartan (selective AT(1) receptor blocker) but not by PD123319 (selective AT(2) receptor blocker). Imipramine and quinidine (putative inhibitors of the Na(+)/Mg(2+) exchanger) and removal of extracellular Na(+) abrogated Ang II-mediated [Mg(2+)](i) effects. In cells pretreated with thapsigargin (reticular Ca(2+)-ATPase inhibitor), Ang II-stimulated [Ca(2+)](i) transients were attenuated (p < 0.01), whereas agonist-induced [Mg(2+)](i) responses were unchanged. Clamping the [Ca(2+)](i) near 50 nmol/liter with 1,2 bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) inhibited Ang II-induced [Ca(2+)](i) increases but failed to alter Ang II induced [Mg(2+)](i) responses. Benzamil, a selective blocker of the Na(+)/Ca(2+) exchanger, inhibited [Na(+)](i) but not [Mg(2+)](i) responses. Our data demonstrate that in MDCK cells, AT(1) receptors modulate [Mg(2+)](i) via a Na(+) dependent Mg(2+) transporter that is not directly related to [Ca(2+)](i). These data support the notion that rapid modulation of [Mg(2+)](i) is not simply a result of Mg(2+) redistribution from intracellular buffering sites by Ca(2+) and provide evidence for the existence of a Na(+)-dependent, hormonally regulated transporter for Mg(2+) in renally derived cells. PMID- 11278388 TI - Matrix GLA protein modulates differentiation induced by bone morphogenetic protein-2 in C3H10T1/2 cells. AB - Matrix GLA protein (MGP) is ubiquitously expressed with high accumulation in bone and cartilage, where it was found to associate with bone morphogenetic proteins (BMP) during protein purification. To test whether MGP affects BMP-induced differentiation, three sets of experiments were performed. First, pluripotent C3H10T1/2 cells transfected with human MPG (hMGP) or antisense to hMGP (AS-hMGP) were treated with BMP-2. In cells overexpressing hMGP, osteogenic and chondrogenic differentiation was inhibited indicating decreased BMP-2 activity. Conversely, in cells overexpressing AS-hMGP, BMP-2 activity was enhanced. Second, cells were prepared from homozygous and heterozygous MPG-deficient mice aortas. When treated with BMP-2, these cells underwent chondrogenic and osteogenic differentiation, respectively, whereas controls did not. Third, FLAG-tagged hMGP with the same biological effect as native hMGP inhibited BMP-induced differentiation, when exogenously added to culture media. Together, these results suggest that MGP modulates BMP activity. To test whether hMGP fragments would retain the effect of full-length hMGP, three subdomains were overexpressed in C3H10T1/2 cells. In cells expressing the mid-region, alone (amino acids (aa) 35 54) or in combination with the N terminus (aa 1-54) but not the C terminus (aa 35 84), osteogenic differentiation was enhanced and occurred even without added BMP 2. Thus, two subdomains had the opposite effect of full-length hMGP, possibly due to different expression levels or domain characteristics. PMID- 11278389 TI - Stimulation of p300-mediated transcription by the kinase MEKK1. AB - p300 and CREB-binding protein (CBP) are related transcriptional coactivators that possess histone acetyltransferase activity. Inactivation of p300/CBP is part of the mechanism by which adenovirus E1A induces oncogenic transformation of cells. Recently, the importance of p300/CBP has been demonstrated directly in several organisms including mouse, Drosophila, and Caenorhabditis elegans where p300/CBP play an indispensable role in differentiation, in patterning, and in cell fate determination and proliferation during development. CBP/p300s are modified by phosphorylation during F9 cell differentiation as well as adenovirus infection, suggesting that phosphorylation may play a role in the regulation of p300/CBP activity. Here we show that the mitogen-activated/extracellular response kinase kinase 1 (MEKK1) enhances p300-mediated transcription. We identify several domains within p300 that can respond to MEKK1-induced transcriptional activation. Interestingly, activation of p300-mediated transcription by MEKK1 does not appear to require the downstream kinase JNK and may involve either a direct phosphorylation of p300 by MEKK1 or by other non-JNK MEKK1-directed downstream kinases. Finally, we present evidence that p300 is important for MEKK1 to induce apoptosis. Taken together, these results identify MEKK1 as a kinase that is likely to be involved in the regulation of the transactivation potential of p300 and support a role of p300 in MEKK1-induced apoptosis. PMID- 11278390 TI - Dominant negative function by an alternatively spliced form of the interferon inducible protein kinase PKR. AB - The double-stranded RNA (dsRNA)-activated protein kinase PKR (protein kinase dsRNA-dependent) plays an important role in the regulation of protein synthesis by phosphorylating the alpha-subunit of eukaryotic initiation factor 2. Through this activity, PKR is thought to mediate the antiviral and antiproliferative actions of interferon. Here, we show that the human T cell leukemia Jurkat cells express an alternatively spliced form of PKR with a deletion of exon 7 (PKRDeltaE7), resulting in a truncated protein that retains the two dsRNA-binding motifs. PKRDeltaE7 exhibits a dominant negative function by inhibiting both PKR autophosphorylation and eukaryotic initiation factor 2 alpha-subunit phosphorylation in vitro and in vivo. Reverse transcriptase-polymerase chain reaction assays showed that PKRDeltaE7 is expressed in a broad range of human tissues at variable levels. Interestingly, expression of PKRDeltaE7 is higher in Jurkat cells than in normal peripheral blood mononuclear cells, raising the possibility of a role in cell proliferation and/or transformation. Thus, expression of alternatively spliced forms of PKR may represent a novel mechanism of PKR autoregulation with important implications in the control of cell proliferation. PMID- 11278391 TI - Signal transduction by the CEACAM1 tumor suppressor. Phosphorylation of serine 503 is required for growth-inhibitory activity. AB - CEACAM1 is a cell-cell adhesion molecule that mediates homophilic cell adhesion. In addition, CEACAM1 was also shown to suppress the growth of prostate, breast, and colon tumors. Structural and functional analyses showed that the adhesion activity of CEACAM1 is mediated by its extracellular domain while its cytoplasmic domain is necessary and sufficient for growth-inhibitory activity. The signal pathways leading to CEACAM1-mediated growth suppression are not known. We studied the importance of phosphorylation of serine 503 in this growth-inhibitory signaling pathway. Full-length CEACAM1 was found to be phosphorylated in vivo in both tyrosine and serine residues. Mutation of tyrosine 488 to phenylalanine did not abolish the tumor-suppressive activity of CEACAM1, suggesting that phosphorylation at tyrosine 488 is not critical for CEACAM1's tumor-suppressive activity. Although expression of CEACAM1's cytoplasmic domain inhibited the growth of DU145 prostate cancer cells in vivo, mutation of serine 503 to alanine abolished the growth-inhibitory activity. In addition, the change of serine 503 to aspartic acid produced tumor-suppressive activity similar to that of the wild type CEACAM1. These results suggested that phosphorylation at serine 503 is essential for CEACAM1's growth-inhibitory function in vivo. PMID- 11278393 TI - The cell cycle inhibitor p16(INK4A) sensitizes lymphoblastic leukemia cells to apoptosis by physiologic glucocorticoid levels. AB - The cyclin-dependent kinase inhibitor p16(INK4A) is frequently inactivated in childhood T-cell acute lymphoblastic leukemia. To investigate possible consequences of this genetic alteration for tumor development, we conditionally expressed p16(INK4A) in the T-cell acute lymphoblastic leukemia line CCRF-CEM, which carries a homozygous deletion of this gene. In agreement with its reported function, p16(INK4A) expression was associated with hypophosphorylation of the retinoblastoma protein pRB and stable cell cycle arrest in G(0)/G(1), documenting that the pRB/E2F pathway is functional in these cells. Unexpectedly, p16(INK4A) expression increased the sensitivity threshold for glucocorticoid (GC)-induced apoptosis from therapeutic to physiologic levels. As a possible explanation for this phenomenon, we found that p16(INK4A)-arrested cells had elevated GC receptor expression associated with enhanced GC-mediated transcriptional activity and increased responsiveness of the GC-regulated cyclin D3 gene. These data are supported by our previous findings that GC receptor levels critically influence GC sensitivity and imply that p16(INK4A) inactivation, in addition to allowing unrestricted proliferation, represents a mechanism by which lymphoid tumor cells might escape cell death triggered by endogenous GC. PMID- 11278392 TI - Transient activation of Jun N-terminal kinases and protection from apoptosis by the insulin-like growth factor I receptor can be suppressed by dicumarol. AB - The insulin-like growth factor I receptor (IGF-IR) activated by its ligands insulin-like growth factor (IGF)-I or IGF-II mediates suppression of apoptosis and contributes to tumorigenesis and cell growth. Here we investigated the activation of the stress-activated protein kinases including Jun N-terminal Kinases and p38 MAPK by IGF-I in interleukin-3-dependent FL5.12 lymphocytic cells that overexpress the IGF-IR (FL5.12/WT). We have shown previously that IGF-I protects these cells from apoptosis induced by interleukin-3 withdrawal but does not promote proliferation. IGF-I induced a rapid and transient activation of JNK that peaked at 40 min that was paralleled by a transient and robust phosphorylation of c-Jun. p38 was constitutively phosphorylated in FL5.12/WT cells. Activation of the JNK pathway by IGF-I occurred in the presence of phosphatidylinositol 3-kinase inhibitors and could be enhanced by anisomycin. Analysis of a series of FL5.12 cells expressing mutated IGF-IRs and analysis of 32D/IGF-IR cells showed that neither the C terminus of the receptor nor IRS-1 and IRS-2 were required for JNK activation, although tyrosine 950 was essential for full activation. The JNK inhibitor dicumarol suppressed IGF-I-mediated activation of JNK and phosphorylation of c-Jun but did not affect p38 and IkappaB phosphorylation or activation of AKT. IGF-I-mediated protection from apoptosis in FL5.12/WT cells was completely suppressed by dicumarol and partially suppressed by a p38 inhibitor. In the breast carcinoma cell line MCF-7, treatment with dicumarol also induced apoptosis. These data indicate that transient activation of JNK by IGF-I is mediated by signals that are distinct from those leading to phosphatidylinositol 3-kinase and AKT activation. The data further suggest that the SAPK pathways contribute to suppression of apoptosis by the IGF-IR. PMID- 11278394 TI - Transcript expression in Saccharomyces cerevisiae at high salinity. AB - Transcript expression of Saccharomyces cerevisiae at high salinity was determined by microarray analysis of 6144 open reading frames (ORFs). From cells grown in 1 m NaCl for 10, 30, and 90 min, changes in transcript abundance >2-fold were classified. Salinity-induced ORFs increased over time: 107 (10 min), 243 (30 min), and 354 (90 min). Up-regulated, functionally unknown ORFs increased from 17 to 149 over this period. Expression patterns were similar early, with 67% of up regulated transcripts after 10 min identical to those at 30 min. The expression profile after 90 min revealed different up-regulated transcripts (identities of 13% and 22%, respectively). Nucleotide and amino acid metabolism exemplified the earliest responses to salinity, followed by ORFs related to intracellular transport, protein synthesis, and destination. Transcripts related to energy production were up-regulated throughout the time course with respiration associated transcripts strongly induced at 30 min. Highly expressed at 90 min were known salinity stress-induced genes, detoxification-related responses, transporters of the major facilitator superfamily, metabolism of energy reserves, nitrogen and sulfur compounds, and lipid, fatty acid/isoprenoid biosynthesis. We chose severe stress conditions to monitor responses in essential biochemical mechanisms. In the mutant, Deltagpd1/gpd2, lacking glycerol biosynthesis, the stress response was magnified with a partially different set of up-regulated ORFs. PMID- 11278395 TI - Activated JNK phosphorylates the c-terminal domain of MLK2 that is required for MLK2-induced apoptosis. AB - MAP kinase signaling pathways are important mediators of cellular responses to a wide variety of stimuli. Signals pass along these pathways via kinase cascades in which three protein kinases are sequentially phosphorylated and activated, initiating a range of cellular programs including cellular proliferation, immune and inflammatory responses, and apoptosis. One such cascade involves the mixed lineage kinase, MLK2, signaling through MAP kinase kinase 4 and/or MAP kinase kinase 7 to the SAPK/JNK, resulting in phosphorylation of transcription factors including the oncogene, c-jun. Recently we showed that MLK2 causes apoptosis in cultured neuronal cells and that this effect is dependent on activation of the JNK pathway (Liu, Y. F., Dorow, D. S., and Marshall, J. (2000) J. Biol. Chem. 275, 19035-19040). Furthermore, dominant-negative MLK2 blocked apoptosis induced by polyglutamine-expanded huntingtin protein, the product of the mutant Huntington's disease gene. Here we show that as well as activating the stress signaling pathway, MLK2 is a target for phosphorylation by activated JNK. Phosphopeptide mapping of MLK2 proteins revealed that activated JNK2 phosphorylates multiple sites mainly within the noncatalytic C-terminal region of MLK2 including the C-terminal 100 amino acid peptide. In addition, MLK2 is phosphorylated in vivo within several of the same C-terminal peptides phosphorylated by JNK2 in vitro, and this phosphorylation is increased by cotransfection of JNK2 and treatment with the JNK activator, anisomycin. Cotransfection of dominant-negative JNK kinase inhibits phosphorylation of kinase negative MLK2 by anisomycin-activated JNK. Furthermore, we show that the N terminal region of MLK2 is sufficient to activate JNK but that removal of the C terminal domain abrogates the apoptotic response. Taken together, these data indicate that the apoptotic activity of MLK2 is dependent on the C-terminal domain that is the main target for MLK2 phosphorylation by activated JNK. PMID- 11278396 TI - S-nitrosation controls gating and conductance of the alpha 1 subunit of class C L type Ca(2+) channels. AB - Modulation of smooth muscle, L-type Ca(2+) channels (class C, Ca(V)1.2b) by thionitrite S-nitrosoglutathione (GSNO) was investigated in the human embryonic kidney 293 expression system at the level of whole-cell and single-channel currents. Extracellular administration of GSNO (2 mM) rapidly reduced whole-cell Ba(2+) currents through channels derived either by expression of alpha1C-b or by coexpression of alpha1C-b plus beta2a and alpha2-delta. The non-thiol nitric oxide (NO) donors 2,2-diethyl-1-nitroso-oxhydrazin (2 mM) and 3 morpholinosydnonimine-hydrochloride (2 mM), which elevated cellular cGMP levels to a similar extent as GSNO, failed to affect Ba(2+) currents significantly. Intracellular administration of copper ions, which promote decomposition of the thionitrite, antagonized its inhibitory effect, and loading of cells with high concentrations of dithiothreitol (2 mM) prevented the effect of GSNO on alpha1C-b channels. Intracellular loading of cells with oxidized glutathione (2 mM) affected neither alpha1C-b channel function nor their modulation by GSNO. Analysis of single-channel behavior revealed that GSNO inhibited Ca(2+) channels mainly by reducing open probability. The development of GSNO-induced inhibition was associated with the transient occurrence of a reduced conductance state of the channel. Our results demonstrate that GSNO modulates the alpha1 subunit of smooth muscle L-type Ca(2+) channels by an intracellular mechanism that is independent of NO release and stimulation of guanylyl cyclase. We suggest S nitrosation of intracellularly located sulfhydryl groups as an important determinant of Ca(2+) channel gating and conductance. PMID- 11278397 TI - Overexpression of LAT1/CD98 light chain is sufficient to increase system L-amino acid transport activity in mouse hepatocytes but not fibroblasts. AB - l-amino acid transporter-1 (LAT1) is a highly conserved gene identified as a light chain of the CD98 amino acid transporter and cellular activation marker. In our previous studies we found increased expression of LAT1 in primary human cancers. We have demonstrated also that LAT1 response to arginine availability is lost in transformed and tumorigenic cells such that expression is constitutively high. System l-amino acid transport activity correlates with changes in LAT1. To assess the functional relevance of increased LAT1 expression and the requirement for 4F2 heavy chain, we overexpressed these CD98 subunits together and separately in nontransformed hepatocytes and fibroblasts. Antigen tags in the expression constructs confirmed that expressed proteins were localized to both cytoplasmic and plasma membrane locations within the cells. Overexpression of LAT1 alone in mouse hepatocytes, but not fibroblasts, was sufficient to increase system l transport, and these cells displayed a growth advantage in conditions of limited arginine. Our results suggest that loss of regulation leading to constitutive expression of LAT1 can contribute to oncogenesis. We hypothesize that the altered LAT1 expression observed in hepatocarcinogenesis gives cells a growth or survival advantage through increased transport activity in a tumor microenvironment of limited amino acid availability. PMID- 11278398 TI - Binding of pyridine nucleotide coenzymes to the beta-subunit of the voltage sensitive K+ channel. AB - The beta-subunit of the voltage-sensitive K(+) (K(v)) channels belongs to the aldo-keto reductase superfamily, and the crystal structure of K(v)beta2 shows NADP bound in its active site. Here we report that K(v)beta2 displays a high affinity for NADPH (K(d) = 0.1 micrometer) and NADP(+) (K(d) = 0.3 micrometer), as determined by fluorometric titrations of the recombinant protein. The K(v)beta2 also bound NAD(H) but with 10-fold lower affinity. The site-directed mutants R264E and N333W did not bind NADPH, whereas, the K(d)(NADPH) of Q214R was 10-fold greater than the wild-type protein. The K(d)(NADPH) was unaffected by the R189M, W243Y, W243A, or Y255F mutation. The tetrameric structure of the wild-type protein was retained by the R264E mutant, indicating that NADPH binding is not a prerequisite for multimer formation. A C248S mutation caused a 5-fold decrease in K(d)(NADPH), shifted the pK(a) of K(d)(NADPH) from 6.9 to 7.4, and decreased the ionic strength dependence of NADPH binding. These results indicate that Arg-264 and Asn-333 are critical for coenzyme binding, which is regulated in part by Cys 248. The binding of both NADP(H) and NAD(H) to the protein suggests that several types of K(v)beta2-nucleotide complexes may be formed in vivo. PMID- 11278399 TI - Bcl-XL expression correlates with primary macrophage differentiation, activation of functional competence, and survival and results from synergistic transcriptional activation by Ets2 and PU.1. AB - Depriving primary bone marrow-derived macrophages of colony-stimulating factor-1 (CSF-1) induces programmed cell death by apoptosis. We show that cell death is accompanied by decreases in the expression of anti-apoptotic Bcl-x(L) protein and the Ets2 and PU.1 proteins of the Ets transcription factor family. Macrophages require both priming and triggering signals independent of CSF-1 to kill neoplastic cells or microorganisms, and this activation of macrophage competence is accompanied by increased expression of bcl-x(L), ets2, and PU.1. Furthermore, we show that only Ets2 and PU.1, but not Ets1, function in a synergistic manner to transactivate the bcl-x promoter. The synergy observed between PU.1 and Ets2 is dependent on the transactivation domains of both proteins. Although other transcription factors like Fos, c-Jun, Myc, STAT3, and STAT5a are implicated in the activation of macrophage competence or in CSF-1 signaling, no synergy was observed between Ets2 and these transcription factors on the bcl-x promoter. We demonstrate that the exogenous expression of both Ets2 and PU.1 in macrophages increases the number of viable cells upon CSF-1 depletion and that Ets2 and PU.1 can functionally replace Bcl-x(L) in inhibiting Bax-induced apoptosis. Together, these results demonstrate that PU.1 and Ets2 dramatically increase bcl-x activation, which is necessary for the cytocidal function and survival of macrophages. PMID- 11278400 TI - Pou homeodomain protein OCT1 is implicated in the expression of the caudal related homeobox gene Cdx-2. AB - The caudal homeobox gene Cdx-2 is a transcriptional activator for approximately a dozen genes specifically expressed in pancreatic islets and intestinal cells. It is also involved in preventing the development of colorectal tumors. Studies using "knockout" approaches demonstrated that Cdx-2 is haplo-insufficient in certain tissues including the intestines but not the pancreatic islets. The mechanisms, especially transcription factors, which regulate Cdx-2 expression, are virtually unknown. We found previously that Cdx-2 expression could be autoregulated in a cell type-specific manner. In this study, we located an octamer (OCT) binding site within the mouse Cdx-2 gene promoter. This site, designated as Cdx-2(P)OCT, is involved in the expression of the Cdx-2 promoter. Both pancreatic and intestinal cell lines were found to express a number of POU (OCT binding) homeodomain proteins examined by electrophoretic mobility shift assay. However, it appears that Cdx-2(P)OCT interacts only with OCT1 in the nuclear extracts of the intestinal cell lines examined, although it interacts with OCT1 and at least two other POU proteins that are to be identified in the pancreatic InR1-G9 cell nuclear extract. Co-transfecting OCT1 cDNA but not five other POU gene cDNAs activates the Cdx-2 promoter in the pancreatic InR1-G9 and the intestinal Caco-2 cell lines. In contrast, Cdx-2(P)OCT cannot act as an enhancer element if it is fused to a thymidine kinase promoter. Furthermore, Cdx 2(P)OCT-thymidine kinase fusion promoters cannot be activated by OCT1 co transfection. Cell type-specific expression, cell type-specific binding affinity of POU proteins to the cis-element Cdx-2(P)OCT, and the DNA content-dependent activation of Cdx-2 promoter via Cdx-2(P)OCT by OCT1 suggest that POU proteins play important and complicated roles in modulating Cdx-2 expression in cell type specific manners. PMID- 11278401 TI - Nuclear import of the U1A splicesome protein is mediated by importin alpha /beta and Ran in living mammalian cells. AB - U1A is a component of the uracil-rich small nuclear ribonucleoprotein. The molecular mechanism of nuclear import of U1A was investigated in vivo and in vitro. When recombinant deletion mutants of U1A are injected into the BHK21 cell cytoplasm, the nuclear localization signal (NLS) of U1A is found in the N terminal half of the central domain (residues 100-144 in mouse U1A). In an in vitro assay, it was found that the U1A-NLS accumulated in only a portion of the nuclei in the absence of cytosolic extract. In contrast, the addition of importin alpha/beta and Ran induced the uniform nuclear accumulation of U1A-NLS in all cells. Furthermore, U1A was found to bind the C-terminal portion of importin alpha. In addition, the in vitro nuclear migration of full-length U1A was found to be exclusively dependent on importin alpha/beta and Ran. Moreover, in living cells, the full-length U1A accumulated in the nucleus in a Ran-dependent manner, and nuclear accumulation was inhibited by the importin beta binding domain of importin alpha. These results suggest that the nuclear import of U1A is mediated by at least two distinct pathways, an importin alpha/beta and Ran-dependent and an -independent pathway in permeabilized cells, and that the latter pathway may be suppressed in intact cells. PMID- 11278403 TI - Caspase-mediated cleavage of hematopoietic progenitor kinase 1 (HPK1) converts an activator of NFkappaB into an inhibitor of NFkappaB. AB - Hematopoietic progenitor kinase 1 (HPK1), a mammalian Ste20-related protein kinase, is a potent stimulator of the stress-activated protein kinases (SAPKs/JNKs). Here we report activation of NFkappaB transcription factors by HPK1 that was independent of SAPK/JNK activation. Overexpression of a dominant negative SEK1 significantly inhibited SAPK/JNK activation, whereas NFkappaB stimulation by HPK1 remained unaffected. Furthermore, activation of NFkappaB required the presence of full-length, kinase-active HPK1, whereas the isolated kinase domain of HPK1 was sufficient for activation of SAPK/JNK. We also demonstrate that overexpression of a dominant-negative IKKbeta blocks HPK1 mediated NFkappaB activation suggesting that HPK1 acts upstream of the IkappaB kinase complex. In apoptotic myeloid progenitor cells HPK1 was cleaved at a DDVD motif resulting in the release of the kinase domain and a C-terminal part. Although expression of the isolated HPK1 kinase domain led to SAPK/JNK activation, the C-terminal part inhibited NFkappaB activation. This dominant negative effect was not only restricted to HPK1-mediated but also to NIK- and tumor necrosis factor alpha-mediated NFkappaB activation, suggesting an impairment of the IkappaB kinase complex. Thus HPK1 activates both the SAPK/JNK and NFkappaB pathway in hematopoietic cells but is converted into an inhibitor of NFkappaB activation in apoptotic cells. PMID- 11278402 TI - Modulation of cell growth by the hepatitis C virus nonstructural protein NS5A. AB - Hepatitis C virus nonstructural protein, NS5A, is a phosphoprotein produced from the processing of the viral polyprotein precursor. NS5A associates with several cellular proteins in mammalian cells, and the biological consequences of this interaction are currently unknown. To this end, five stable NS5A-expressing murine and human cell lines were established. Tetracycline-regulated NIH3T3 cells and rat liver epithelial cells as well as the constitutive, NS5A-expressing, human Chang liver, HeLa, and NIH3T3 cells all exhibited cell growth retardation compared with the control cells. Cell cycle analysis by flow cytometry indicated that the NS5A-expressing human epitheloid tumor cells had a reduced S phase and an increase in the G(2)/M phase, which could be explained by a p53-dependent induction of p21(Waf1/Cip1) protein and mRNA levels. NS5A interacts with Cdk1 in vivo and in vitro, and a significant portion of the p21(Waf1/Cip1) was found to be in a complex with Cdk2 in the NS5A-expressing human hepatic cell line. Cdk1 and cyclin B1 proteins were also reduced in human Chang liver cells consistent with the increase in G(2)/M phase. Our results suggest that the NS5A protein causes growth inhibition and cell cycle perturbations by targeting the Cdk1/2 cyclin complexes. PMID- 11278404 TI - A mechanistic model for Ncd directionality. AB - Ncd is a kinesin-related protein that drives movement to the minus-end of microtubules. Pre-steady-state kinetic experiments have been employed to investigate the cooperative interactions between the motor domains of the MC1 dimer and to establish the ATPase mechanism. Our results indicate that the active sites of dimeric Ncd free in solution are not equivalent; ADP is held more tightly at one site than at the other. Upon microtubule binding, fast release of ADP from the first motor domain is stimulated at 18 s(-1), yet rate-limiting ADP release from the second motor domain occurs at 1.4 s(-1). We propose that the head with the low affinity for ADP binds the microtubule first to establish the directional bias of the microtubule.Ncd intermediate where one motor domain is bound to the microtubule with the second head detached and directed toward the minus-end of the microtubule. The force generating cycle is initiated as ATP binds to the empty site of the microtubule-bound head. ATP hydrolysis at head 1 is required for head 2 to bind to the microtubule. The kinetics indicate that two ATP molecules are required for a single step and force generation for minus-end directed movement generated by this non-processive dimeric motor. PMID- 11278405 TI - Toxoplasma gondii ADP-ribosylation factor 1 mediates enhanced release of constitutively secreted dense granule proteins. AB - Toxoplasma gondii dense granules are morphologically similar to dense matrix granules in specialized secretory cells, yet are secreted in a constitutive, calcium-independent fashion. We previously demonstrated that secretion of dense granule proteins in permeabilized parasites was augmented by the non-hydrolyzable GTP analogue guanosine 5'-3-O-(thio)triphosphate (GTPgammaS) (Chaturvedi, S., Qi, H., Coleman, D. L., Hanson, P., Rodriguez, A., and Joiner, K. A. (1998) J. Biol. Chem. 274, 2424-2431). As now demonstrated by pharmacological and electron microscopic approaches, GTPgammaS enhanced release of dense granule proteins in the permeabilized cell system. To investigate the role of ADP-ribosylation factor 1 (ARF1) in this process, a cDNA encoding T. gondii ARF1 (TgARF1) was isolated. Endogenous and transgenic TgARF1 localized to the Golgi of T. gondii, but not to dense granules. An epitope-tagged mutant of TgARF1 predicted to be impaired in GTP hydrolysis (Q71L) partially dispersed the Golgi signal, with localization to scattered vesicles, whereas a mutant impaired in nucleotide binding (T31N) was cytosolic in location. Both mutants caused partial dispersion of a Golgi/trans Golgi network marker. TgARF1 mutants inhibited delivery of the secretory reporter, Escherichia coli alkaline phosphatase, to dense granules, precluding an in vivo assessment of the role of TgARF1 in release of intact dense granules. To circumvent this limitation, recombinant TgARF1 was purified using two separate approaches, and used in the permeabilized cell assay. TgARF1 protein purified on a Cibacron G3 column and able to bind GTP stimulated dense granule secretion in the permeabilized cell secretion assay. These results are the first to show that ARF1 can augment release of constitutively secreted vesicles at the target membrane. PMID- 11278406 TI - Identification of specific pore residues mediating KCNQ1 inactivation. A novel mechanism for long QT syndrome. AB - KCNQ1 inactivation bears electrophysiological characteristics different from classical N- and C-type inactivation in Shaker-like potassium channels. However, the molecular site of KCNQ1 inactivation has not yet been determined. KCNQ2 channels do not exert a fast inactivation in contrast to KCNQ1 channels. By expressing functional chimeras between KCNQ1 and KCNQ2 in Xenopus oocytes, we mapped the region of this inactivation to transmembrane domain S5 and the pore loop H5 and finally narrowed down the site to positions Gly(272) and Val(307) in KCNQ1. Exchanging these two amino acids individually with the analogous KCNQ2 residue abolished inactivation. Furthermore, a KCNQ1-like inactivation was introduced into KCNQ2 by mutagenesis in the corresponding region, confirming its relevance for the inactivation process. As KCNQ1 inactivation involves the regions S5 and H5, it exhibits a geography distinct from N- or C-type inactivation. Native cardiac I(Ks) channels comprising KCNQ1 and accessory MinK subunits do not inactivate because of the functional interaction of KCNQ1 with MinK. Mutations in KCNQ1 can lead to long QT1 syndrome, an inherited form of arrhythmia. The long QT1 mutant KCNQ1(L273F) displays a pronounced KCNQ1 inactivation. Here we show that when expressing mutant I(Ks) channels formed from KCNQ1(L273F) and MinK, MinK association no longer eliminates KCNQ1 inactivation. This results in smaller repolarizing currents in the heart and therefore represents a novel mechanism leading to long QT syndrome. PMID- 11278407 TI - Sphingosine 1-phosphate and activation of endothelial nitric-oxide synthase. differential regulation of Akt and MAP kinase pathways by EDG and bradykinin receptors in vascular endothelial cells. AB - Sphingosine 1-phosphate (S1P) is a platelet-derived sphingolipid that elicits numerous biological responses in endothelial cells mediated by a family of G protein-coupled EDG receptors. Stimulation of EDG receptors by S1P has been shown to activate the endothelial isoform of nitric-oxide synthase (eNOS) in heterologous expression systems (Igarashi, J., and Michel, T. (2000) J. Biol. Chem. 275, 32363-32370). However, the signaling pathways that modulate eNOS regulation by S1P/EDG in vascular endothelial cells remain less well understood. We now report that S1P treatment of bovine aortic endothelial cells (BAEC) acutely increases eNOS enzyme activity; the EC(50) for S1P activation of eNOS is approximately 10 nm. The magnitude of eNOS activation by S1P in BAEC is equivalent to that elicited by the agonist bradykinin. S1P treatment activates Akt, a protein kinase implicated in phosphorylation of eNOS. S1P treatment of BAEC leads to eNOS phosphorylation at Ser(1179), a residue phosphorylated by Akt; an eNOS mutant in which this Akt phosphorylation site is inactivated shows attenuated S1P-induced eNOS activation. S1P-induced activation both of Akt and of eNOS is inhibited by pertussis toxin, by the phosphoinositide 3-kinase inhibitor wortmannin, and by the intracellular calcium chelator BAPTA (1,2 bis(aminophenoxy)ethane-N,N,N',N'-tetraacetic acid). By contrast to S1P, activation of G protein-coupled bradykinin B2 receptors neither activates kinase Akt nor promotes Ser(1179) eNOS phosphorylation despite robustly activating eNOS enzyme activity. Understanding the differential regulation of protein kinase pathways by S1P and bradykinin may lead to the identification of new points for eNOS regulation in vascular endothelial cells. PMID- 11278408 TI - Estrogen receptor binding to DNA is not required for its activity through the nonclassical AP1 pathway. AB - In the classical signaling pathway, the estrogen receptor (ER) binds directly to estrogen response elements (EREs) to regulate gene transcription. To test the hypothesis that the nonclassical pathway involves ER interactions with other proteins rather than direct binding to DNA, mutations were introduced into the DNA binding domain (DBD) of the mouse ERalpha. The effects of these DBD mutations were examined in DNA binding assays using reporter constructs containing either EREs (classical) or AP1 (nonclassical) response elements. Using the AP1 reporter, there was a reversal of ER action relative to that seen with the ERE reporter. Estradiol induced suppression, and the antiestrogen ICI 182,780 stimulated transcription of the AP1 reporter. DBD mutations in the proximal (P-box) of the first zinc finger of the ER (E207A/G208A and E207G/G208S) eliminated ERE binding. These mutants were inactive using the ERE reporter but retained partial or full activity with the AP1 reporter. The DBD mutant ERs interacted with Jun when tested in mammalian cell two-hybrid assays. Two mutations (K366D and I362R) in the ER ligand binding domain known to alter coactivator interactions impaired transcriptional responses using either the ERE or AP1 reporters. We concluded that ER action through the AP1 response element involves interactions with other promoter-bound proteins instead of, or in addition to, direct binding to DNA. Interactions with coactivators were required for both pathways. These data supported a model in which ER-mediated transcriptional activation or repression is dependent on the ligand and the nature of the response element in the target gene. PMID- 11278409 TI - Identification and characterization of a transcriptional regulator for the lck proximal promoter. AB - The lck gene encodes a protein-tyrosine kinase that plays a key role in signaling mediated through T cell receptor (TCR) and pre-TCR complexes. Transcription of the lck gene is regulated by two independent promoter elements: the proximal and distal promoters. Previous studies employing transgenic mice demonstrated that the sequence between -584 and -240 from the transcription start site in the mouse lck proximal promoter is required for its tissue-specific expression in the thymus. In this study, we demonstrate that a Kruppel-like zinc finger protein, mtbeta (BFCOL1, BERF-1, ZBP-89, ZNF148), previously cloned as a protein that binds to the CD3delta gene enhancer, binds to the -365 to -328 region of the lck proximal promoter. mtbeta is ubiquitously expressed in various cell lines and mouse tissues. Overexpressed mtbeta is more active in T-lineage cells than B lineage cells for transactivating an artificial promoter consisting of the mtbeta binding site and a TATA box. Activity of the lck proximal promoter was significantly impaired by mutating the mtbeta binding site or by reducing mtbeta protein expression level by using antisense mRNA. Our results indicate that mtbeta activity is regulated in a tissue-specific manner and that mtbeta is a critical transactivator for the lck proximal promoter. PMID- 11278410 TI - Filamin associates with Smads and regulates transforming growth factor-beta signaling. AB - Members of the Smad proteins transmit signals triggered by the ligands of transforming growth factor (TGF)-beta superfamily. Ligand-activated receptors induce phosphorylation of so-called receptor-regulated Smads, which then accumulate in the nucleus to participate in target gene transcription, in collaboration with Smad-interacting proteins. We performed yeast two-hybrid screening and identified filamin, a cytoskeletal actin-binding protein 280, as a Smad5-interacting protein. Filamin was found to be associated not only with Smad5 but also with other Smad proteins, including TGF-beta/activin receptor-regulated Smad2. TGF-beta signaling was defective in filamin-deficient human melanoma cells M2 compared with a filamin-transfected subline A7, as determined by TGF-beta responsive reporter gene activation and Smad2 nuclear accumulation. M2 cells restored TGF-beta responsiveness following transient transfection of full-length filamin encoding vector. The defective TGF-beta signaling in M2 cells seemed to be due to impaired receptor-induced serine phosphorylation of Smad2. These results suggest that filamin plays an important role in Smad-mediated signaling. PMID- 11278411 TI - Cryo-electron microscopic localization of protein L7/L12 within the Escherichia coli 70 S ribosome by difference mapping and Nanogold labeling. AB - The Escherichia coli ribosomal protein L7/L12 is central to the translocation step of translation, and it is known to be flexible under some conditions. The assignment of electron density to L7/L12 was not possible in the recent 2.4 A resolution x-ray crystallographic structure (Ban, N., Nissen, P., Hansen, J., Moore, P. B., and Steitz, T. A. (2000) Science 289, 905-920). We have localized the two dimers of L7/L12 within the structure of the 70 S ribosome using two reconstitution approaches together with cryo-electron microscopy and single particle reconstruction. First, the structures were determined for ribosomal cores from which protein L7/L12 had been removed by treatment with NH(4)Cl and ethanol and for reconstituted ribosomes in which purified L7/L12 had been restored to core particles. Difference mapping revealed that the reconstituted ribosomes had additional density within the L7/L12 shoulder next to protein L11. Second, ribosomes were reconstituted using an L7/L12 variant in which a single cysteine at position 89 in the C-terminal domain was modified with Nanogold (Nanoprobes, Inc.), a 14 A gold derivative. The reconstruction from cryo-electron microscopy images and difference mapping placed the gold at four interfacial positions. The finding of multiple sites for the C-terminal domain of L7/L12 suggests that the conformation of this protein may change during the steps of elongation and translocation. PMID- 11278412 TI - Stromal and epithelial expression of tumor markers hyaluronic acid and HYAL1 hyaluronidase in prostate cancer. AB - Hyaluronic acid (HA), a glycosaminoglycan, regulates cell adhesion and migration. Hyaluronidase (HAase), an endoglycosidase, degrades HA into small angiogenic fragments. Using an enzyme-linked immunosorbent assay-like assay, we found increased HA levels (3-8-fold) in prostate cancer (CaP) tissues when compared with normal (NAP) and benign (BPH) tissues. The majority ( approximately 75-80%) of HA in prostate tissues was found to exist in the free form. Primary CaP fibroblast and epithelial cells secreted 3-8-fold more HA than respective NAP and BPH cultures. Only CaP epithelial cells and established CaP lines secreted HAase and the secretion increased with tumor grade and metastasis. The pH activity profile and optimum (4.2; range 4.0-4.3) of CaP HAase was identical to the HYAL1 type HAase present in human serum and urine. Full-length HYAL1 transcript and splice variants were detected in CaP cells by reverse transcriptase-polymerase chain reaction, cloning, and sequencing. Immunoblotting confirmed secretion of a approximately 60-kDa HYAL1-related protein by CaP cells. Immunohistochemistry showed minimal HA and HYAL1 staining in NAP and BPH tissues. However, a stromal and epithelial pattern of HA and HYAL1 expression was observed in CaP tissues. While high HA staining was observed in tumor-associated stroma, HYAL1 staining in tumor cells increased with tumor grade and metastasis. The gel-filtration column profiles of HA species in NAP, BPH, and CaP tissues were different. While the higher molecular mass and intermediate size HA was found in all tissues, the HA fragments were found only in CaP tissues. In particular, the high-grade CaP tissues, which showed both elevated HA and HYAL1 levels, contained angiogenic HA fragments. The stromal-epithelial HA and HYAL1 expression may promote angiogenesis in CaP and may serve as prognostic markers for CaP. PMID- 11278413 TI - Yop1p, the yeast homolog of the polyposis locus protein 1, interacts with Yip1p and negatively regulates cell growth. AB - Rab proteins are small GTPases that are essential elements of the protein transport machinery of eukaryotic cells. Each round of membrane transport requires a cycle of Rab protein nucleotide binding and hydrolysis. We have recently characterized a protein, Yip1p, which appears to play a role in Rab mediated membrane transport in Saccharomyces cerevisiae. In this study, we report the identification of a Yip1p-associated protein, Yop1p. Yop1p is a membrane protein with a hydrophilic region at its N terminus through which it interacts specifically with the cytosolic domain of Yip1p. Yop1p could also be coprecipitated with Rab proteins from total cellular lysates. The TB2 gene is the human homolog of Yop1p (Kinzler, K. W., Nilbert, M. C., Su, L.-K., Vogelstein, B., Bryan, T. M., Levey, D. B., Smith, K. J., Preisinger, A. C., Hedge, P., McKechnie, D., Finniear, R., Markham, A., Groffen, J., Boguski, M. S., Altschul, S. F., Horii, A., Ando, H. M., Y., Miki, Y., Nishisho, I., and Nakamura, Y. (1991) Science 253, 661-665). Our data demonstrate that Yop1p negatively regulates cell growth. Disruption of YOP1 has no apparent effect on cell viability, while overexpression results in cell death, accumulation of internal cell membranes, and a block in membrane traffic. These results suggest that Yop1p acts in conjunction with Yip1p to mediate a common step in membrane traffic. PMID- 11278414 TI - Analysis of glomerulosclerosis and atherosclerosis in lecithin cholesterol acyltransferase-deficient mice. AB - To evaluate the biochemical and molecular mechanisms leading to glomerulosclerosis and the variable development of atherosclerosis in patients with familial lecithin cholesterol acyl transferase (LCAT) deficiency, we generated LCAT knockout (KO) mice and cross-bred them with apolipoprotein (apo) E KO, low density lipoprotein receptor (LDLr) KO, and cholesteryl ester transfer protein transgenic mice. LCAT-KO mice had normochromic normocytic anemia with increased reticulocyte and target cell counts as well as decreased red blood cell osmotic fragility. A subset of LCAT-KO mice accumulated lipoprotein X and developed proteinuria and glomerulosclerosis characterized by mesangial cell proliferation, sclerosis, lipid accumulation, and deposition of electron dense material throughout the glomeruli. LCAT deficiency reduced the plasma high density lipoprotein (HDL) cholesterol (-70 to -94%) and non-HDL cholesterol (-48 to -85%) levels in control, apoE-KO, LDLr-KO, and cholesteryl ester transfer protein-Tg mice. Transcriptome and Western blot analysis demonstrated up regulation of hepatic LDLr and apoE expression in LCAT-KO mice. Despite decreased HDL, aortic atherosclerosis was significantly reduced (-35% to -99%) in all mouse models with LCAT deficiency. Our studies indicate (i) that the plasma levels of apoB containing lipoproteins rather than HDL may determine the atherogenic risk of patients with hypoalphalipoproteinemia due to LCAT deficiency and (ii) a potential etiological role for lipoproteins X in the development of glomerulosclerosis in LCAT deficiency. The availability of LCAT-KO mice characterized by lipid, hematologic, and renal abnormalities similar to familial LCAT deficiency patients will permit future evaluation of LCAT gene transfer as a possible treatment for glomerulosclerosis in LCAT-deficient states. PMID- 11278415 TI - Activation of annexin 7 by protein kinase C in vitro and in vivo. AB - Annexin 7, a Ca(2+)/GTP-activated membrane fusion protein, is preferentially phosphorylated in intact chromaffin cells, and the levels of annexin 7 phosphorylation increase quantitatively in proportion to the extent of catecholamine secretion. Consistently, various protein kinase C inhibitors proportionately reduce both secretion and phosphorylation of annexin 7 in these cells. In vitro, annexin 7 is quantitatively phosphorylated by protein kinase C to a mole ratio of 2.0, and phosphorylation is extraordinarily sensitive to variables such as pH, calcium, phospholipid, phorbol ester, and annexin 7 concentration. Phosphorylation of annexin 7 by protein kinase C significantly potentiates the ability of the protein to fuse phospholipid vesicles and lowers the half-maximal concentration of calcium needed for this fusion process. Furthermore, other protein kinases, including cAMP-dependent protein kinase, cGMP dependent protein kinase, and protein-tyrosine kinase pp60(c-)(src), also label annexin 7 with high efficiency but do not have this effect on membrane fusion. In the case of pp60(c-)(src), we note that this kinase, if anything, modestly suppresses the membrane fusion activity of annexin 7. These results thus lead us to hypothesize that annexin 7 may be a positive mediator for protein kinase C action in the exocytotic membrane fusion reaction in chromaffin cells. PMID- 11278416 TI - Neprilysin degrades both amyloid beta peptides 1-40 and 1-42 most rapidly and efficiently among thiorphan- and phosphoramidon-sensitive endopeptidases. AB - To identify the amyloid beta peptide (Abeta) 1-42-degrading enzyme whose activity is inhibited by thiorphan and phosphoramidon in vivo, we searched for neprilysin (NEP) homologues and cloned neprilysin-like peptidase (NEPLP) alpha, NEPLP beta, and NEPLP gamma cDNAs. We expressed NEP, phosphate-regulating gene with homologies to endopeptidases on the X chromosome (PEX), NEPLPs, and damage induced neuronal endopeptidase (DINE) in 293 cells as 95- to 125-kDa proteins and found that the enzymatic activities of PEX, NEPLP alpha, and NEPLP beta, as well as those of NEP and DINE, were sensitive to thiorphan and phosphoramidon. Among the peptidases tested, NEP degraded both synthetic and cell-secreted Abeta1-40 and Abeta1-42 most rapidly and efficiently. PEX degraded cold Abeta1-40 and NEPLP alpha degraded both cold Abeta1-40 and Abeta1-42, although the rates and the extents of the digestion were slower and less efficient than those exhibited by NEP. These data suggest that, among the endopeptidases whose activities are sensitive to thiorphan and phosphoramidon, NEP is the most potent Abeta-degrading enzyme in vivo. Therefore, manipulating the activity of NEP would be a useful approach in regulating Abeta levels in the brain. PMID- 11278417 TI - Uridine phosphorylase association with vimentin. Intracellular distribution and localization. AB - Uridine phosphorylase (UPase), a key enzyme in the pyrimidine salvage pathway, is associated with the intermediate filament protein vimentin, in NIH 3T3 fibroblasts and colon 26 cells. Affinity chromatography was utilized to purify UPase from colon 26 and NIH 3T3 cells using the uridine phosphorylase inhibitor 5'-amino benzylacyclouridine linked to an agarose matrix. Vimentin copurification with UPase was confirmed using both Western blot analysis and MALDI-MS methods. Separation of cytosolic proteins using gel filtration chromatography yields a high molecular weight complex containing UPase and vimentin. Purified recombinant UPase and recombinant vimentin were shown to bind in vitro with an affinity of 120 pm and a stoichiometry of 1:2. Immunofluorescence techniques confirm that UPase is associated with vimentin in both NIH 3T3 and colon 26 cells and that depolymerization of the microtubule system using nocodazole results in UPase remaining associated with the collapsed intermediate filament, vimentin. Our data demonstrate that UPase is associated with both the soluble and insoluble pools of vimentin. Approximately 60-70% of the total UPase exists in the cytosol as a soluble protein. Sequential extraction of NIH 3T3 or colon 26 cells liberates an additional 30-40% UPase activity associated with a detergent extractable fraction. All pools of UPase have been shown to possess enzymatic activity. We demonstrate for the first time that UPase is associated with vimentin and the existence of an enzymatically active cytoskeleton-associated UPase. PMID- 11278418 TI - Isolation and characterization of senescence-induced cDNAs encoding deoxyhypusine synthase and eucaryotic translation initiation factor 5A from tomato. AB - Full-length cDNA clones encoding deoxyhypusine synthase (DHS) and eucaryotic initiation factor 5A (eIF-5A) have been isolated from a cDNA expression library prepared from tomato leaves (Lycopersicon esculentum, cv. Match) exposed to environmental stress. DHS mediates the first of two enzymatic reactions that activate eIF-5A by converting a conserved lysine to the unusual amino acid, deoxyhypusine. Recombinant protein obtained by expressing tomato DHS cDNA in Escherichia coli proved capable of carrying out the deoxyhypusine synthase reaction in vitro in the presence of eIF-5A. Of particular interest is the finding that DHS mRNA and eIF-5A mRNA show a parallel increase in abundance in senescing tomato flowers, senescing tomato fruit, and environmentally stressed tomato leaves exhibiting programmed cell death. Western blot analyses indicated that DHS protein also increases at the onset of senescence. It is apparent from previous studies with yeast and mammalian cells that hypusine-modified eIF-5A facilitates the translation of a subset of mRNAs mediating cell division. The present study provides evidence for senescence-induced DHS and eIF-5A in tomato tissues that may facilitate the translation of mRNA species required for programmed cell death. PMID- 11278419 TI - Differential responses to nerve growth factor and epidermal growth factor in neurite outgrowth of PC12 cells are determined by Rac1 activation systems. AB - Neurite outgrowth of PC12 cells is induced by nerve growth factor (NGF) but not by epidermal growth factor (EGF). This differential response has been explained by the duration of mitogen-activated protein kinase (MAPK) activation; NGF induces sustained MAPK activation but EGF leads short-lived activation. However, precise mechanisms have not yet been understood. Here we demonstrate the difference between NGF and EGF in regulation of Rac1, a small GTPase involved in neurite outgrowth, in PC12 cells. NGF phosphoinositide 3-kinase dependently induces transient activation of Rac1 and accumulation of active Rac1 at protrusion sites on the cell surface, inducing filamentous actin-rich protrusions and subsequent neurite formation in a Rac1-dependent manner. On the other hand, EGF phosphoinositide 3-kinase independently induces more transient Rac1 activation but neither accumulates active Rac1 nor forms Rac1- and filamentous actin-rich protrusions. Difference in the Rac1 localization between NGF and EGF was also observed with the localization of exogenously expressed green fluorescent protein-tagged Rac1. The Rac1-mediated protrusion by NGF is independent of MAPK cascade, but the subsequent neurite extension requires the cascade. Thus, the differential activation of Rac1 and localization of active Rac1 contribute to the difference in the ability of NGF and EGF to induce neurite outgrowth, and we propose that the MAPK cascade-independent prompt activation of Rac1 and recruitment of active Rac1 at the protrusion sites trigger the initiation of neurite formation. PMID- 11278420 TI - The activity of anandamide at vanilloid VR1 receptors requires facilitated transport across the cell membrane and is limited by intracellular metabolism. AB - The endogenous ligand of CB(1) cannabinoid receptors, anandamide, is also a full agonist at vanilloid VR1 receptors for capsaicin and resiniferatoxin, thereby causing an increase in cytosolic Ca(2+) concentration in human VR1-overexpressing (hVR1-HEK) cells. Two selective inhibitors of anandamide facilitated transport into cells, VDM11 and VDM13, and two inhibitors of anandamide enzymatic hydrolysis, phenylmethylsulfonyl fluoride and methylarachidonoyl fluorophosphonate, inhibited and enhanced, respectively, the VR1-mediated effect of anandamide, but not of resiniferatoxin or capsaicin. The nitric oxide donor, sodium nitroprusside, known to stimulate anandamide transport, enhanced anandamide effect on the cytosolic Ca(2+) concentration. Accordingly, hVR1-HEK cells contain an anandamide membrane transporter inhibited by VDM11 and VDM13 and activated by sodium nitroprusside, and an anandamide hydrolase activity sensitive to phenylmethylsulfonyl fluoride and methylarachidonoyl fluorophosphonate, and a fatty acid amide hydrolase transcript. These findings suggest the following. (i) Anandamide activates VR1 receptors by acting at an intracellular site. (ii) Degradation by fatty acid amide hydrolase limits anandamide activity on VR1; and (iii) the anandamide membrane transporter inhibitors can be used to distinguish between CB(1) or VR1 receptor-mediated actions of anandamide. By contrast, the CB(1) receptor antagonist SR141716A inhibited also the VR1-mediated effect of anandamide and capsaicin on cytosolic Ca(2+) concentration, although at concentrations higher than those required for CB(1) antagonism. PMID- 11278421 TI - Progesterone stimulates adipocyte determination and differentiation 1/sterol regulatory element-binding protein 1c gene expression. potential mechanism for the lipogenic effect of progesterone in adipose tissue. AB - Fatty acid synthase (FAS), a nutritionally regulated lipogenic enzyme, is transcriptionally controlled by ADD1/SREBP1c (adipocyte determination and differentiation 1/sterol regulatory element-binding protein 1c), through insulin mediated stimulation of ADD1/SREBP1c expression. Progesterone exerts lipogenic effects on adipocytes, and FAS is highly induced in breast tumor cell lines upon progesterone treatment. We show here that progesterone up-regulates ADD1/SREBP1c expression in the MCF7 breast cancer cell line and the primary cultured preadipocyte from rat parametrial adipose tissue. In MCF7, progesterone induced ADD1/SREBP1c and Metallothionein II (a well known progesterone-regulated gene) mRNAs, with comparable potency. In preadipocytes, progesterone increased ADD1/SREBP1c mRNA dose-dependently, but not SREBP1a or SREBP2. Run-on experiments demonstrated that progesterone action on ADD1/SREBP1c was primarily at the transcriptional level. The membrane-bound and mature nuclear forms of ADD1/SREBP1 protein accumulated in preadipocytes cultured with progesterone, and FAS induction could be abolished by adenovirus-mediated overexpression of a dominant negative form of ADD1/SREBP1 in these cells. Finally, in the presence of insulin, progesterone was unable to up-regulate ADD1/SREBP1c mRNA in preadipocytes, whereas its effect was restored after 24 h of insulin deprivation. Together these results demonstrate that ADD1/SREBP1c is controlled by progesterone, which, like insulin, acts by increasing ADD1/SREBP1c gene transcription. This provides a potential mechanism for the lipogenic actions of progesterone on adipose tissue. PMID- 11278422 TI - Yes-associated protein and p53-binding protein-2 interact through their WW and SH3 domains. AB - To understand the role of the Yes-associated protein (YAP), binding partners of its WW1 domain were isolated by a yeast two-hybrid screen. One of the interacting proteins was identified as p53-binding protein-2 (p53BP-2). YAP and p53BP-2 interacted in vitro and in vivo using their WW1 and SH3 domains, respectively. The YAP WW1 domain bound to the YPPPPY motif of p53BP-2, whereas the p53BP-2 SH3 domain interacted with the VPMRLR sequence of YAP, which is different from other known SH3 domain-binding motifs. By mutagenesis, we showed that this unusual SH3 domain interaction was due to the presence of three consecutive tryptophans located within the betaC strand of the SH3 domain. A point mutation within this triplet, W976R, restored the binding selectivity to the general consensus sequence for SH3 domains, the PXXP motif. A constitutively active form of c-Yes was observed to decrease the binding affinity between YAP and p53BP-2 using chloramphenicol acetyltransferase/enzyme-linked immunosorbent assay, whereas the overexpression of c-Yes did not modify this interaction. Since overexpression of an activated form of c-Yes resulted in tyrosine phosphorylation of p53BP-2, we propose that the p53BP-2 phosphorylation, possibly in the WW1 domain-binding motif, might negatively regulate the YAP.p53BP-2 complex. PMID- 11278423 TI - Activation of protein kinase A and atypical protein kinase C by A(2A) adenosine receptors antagonizes apoptosis due to serum deprivation in PC12 cells. AB - We found in the present study that stimulation of A(2A) adenosine receptors (A(2A)-R) prevents apoptosis in PC12 cells. This A(2A)-protective effect was blocked by protein kinase A (PKA) inhibitors and was not observed in a PKA deficient PC12 variant. Stimulation of PKA also prevented apoptosis, suggesting that PKA is required for the protective effect of A(2A)-R. A general PKC inhibitor, but not down-regulation of conventional and novel PKCs, readily blocked the protective effect of A(2A)-R stimulation and PKA activation, suggesting that atypical PKCs (aPKCs) serve a critical role downstream of PKA. Consistent with this hypothesis, stimulation of A(2A)-R or PKA enhanced nuclear aPKC activity. In addition, the A(2A)-protective effect was blocked by a specific inhibitor of one aPKC, PKCzeta, whereas overexpression of a dominant-positive PKCzeta enhanced survival. In contrast, inhibitors of MAP kinase and phosphatidylinositol 3-kinase did not modulate the A(2A)-protective effect. Dominant-negative Akt also did not alter the A(2A)-protective effect, whereas it significantly reduced the protective action of nerve growth factor. Collectively, these data suggest that aPKCs can function downstream of PKA to mediate the A(2A) R-promoted survival of PC12 cells. Furthermore, the results indicate that different extracellular stimuli can employ distinct signaling pathways to protect against apoptosis induced by the same insult. PMID- 11278424 TI - Calsenilin is a substrate for caspase-3 that preferentially interacts with the familial Alzheimer's disease-associated C-terminal fragment of presenilin 2. AB - Calsenilin is a member of the recoverin family of neuronal calcium-binding proteins that we have previously shown to interact with presenilin 1 (PS1) and presenilin 2 (PS2) holoproteins. The expression of calsenilin can regulate the levels of a proteolytic product of PS2 (Buxbaum, J. D., Choi, E. K., Luo, Y., Lilliehook, C., Crowley, A. C., Merriam, D. E., and Wasco, W. (1998) Nat. Med. 4, 1177-1181) and reverse the presenilin-mediated enhancement of calcium signaling (Leissring, M. A., Yamasaki, T. R., Wasco, W., Buxbaum, J. D., Parker, I., and LaFerla, F. M. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 8590-8593). Here, we have used cultured mammalian cells that transiently or stably express calsenilin to extend the characterization of calsenilin and of the calsenilin-PS2 interaction. We have found that calsenilin has the ability to interact with endogenous 25-kDa C-terminal fragment (CTF) that is a product of regulated endoproteolytic cleavage of PS2 and that the presence of the N141I PS2 mutation does not significantly alter the interaction of calsenilin with PS2. Interestingly, when the 25-kDa PS2 CTF and the 20-kDa PS2 CTF are both present, calsenilin preferentially interacts with the 20-kDa CTF. Increases in the 20-kDa fragment are associated with the presence of familial Alzheimer's disease associated mutations (Kim, T., Pettingell, W. H., Jung, Y., Kovacs, D. M., and Tanzi, R. E. (1997) Science 277, 373-376). However, the finding that the production of the 20-kDa fragment is regulated by the phosphorylation of PS2 (Walter, J., Schindzielorz, A., Grunberg, J., and Haass, C. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 1391-1396) suggests that it is a regulated physiological event that also occurs in the absence of the familial Alzheimer's disease associated mutations in PS2. Finally, we have demonstrated that calsenilin is a substrate for caspase-3, and we have used site-directed mutagenesis to map the caspase-3 cleavage site to a region that is proximal to the calcium binding domain of calsenilin. PMID- 11278425 TI - Oxidation of thymine to 5-formyluracil in DNA promotes misincorporation of dGMP and subsequent elongation of a mismatched primer terminus by DNA polymerase. AB - 5-Formyluracil (fU) is a major oxidative thymine lesion generated by ionizing radiation and reactive oxygen species. In the present study, we have assessed the influence of fU on DNA replication to elucidate its genotoxic potential. Oligonucleotide templates containing fU at defined sites were replicated in vitro by Escherichia coli DNA polymerase I Klenow fragment deficient in 3'-5' exonuclease. Gel electrophoretic analysis of the reaction products showed that fU constituted very weak replication blocks to DNA synthesis, suggesting a weak to negligible cytotoxic effect of this lesion. However, primer extension assays with a single dNTP revealed that fU directed incorporation of not only correct dAMP but also incorrect dGMP, although much less efficiently. No incorporation of dCMP and dTMP was observed. When fU was substituted for T in templates, the incorporation efficiency of dAMP (f(A) = V(max)/K(m)) decreased to (1/4) to (1/2), depending on the nearest neighbor base pair, and that of dGMP (f(G)) increased 1.1-5.6-fold. Thus, the increase in the replication error frequency (f(G)/f(A) for fU versus T) was 3.1-14.3-fold. The misincorporation rate of dGMP opposite fU (pK(a) = 8.6) but not T (pK(a) = 10.0) increased with pH (7.2-8.6) of the reaction mixture, indicating the participation of the ionized (or enolate) form of fU in the mispairing with G. The resulting mismatched fU:G primer terminus was more efficiently extended than the T:G terminus (8.2-11.3-fold). These results show that when T is oxidized to fU in DNA, fU promotes both misincorporation of dGMP at this site and subsequent elongation of the mismatched primer, hence potentially mutagenic. PMID- 11278427 TI - Peflin and ALG-2, members of the penta-EF-hand protein family, form a heterodimer that dissociates in a Ca2+-dependent manner. AB - Peflin, a newly identified 30-kDa Ca(2+)-binding protein, belongs to the penta-EF hand (PEF) protein family, which includes the calpain small subunit, sorcin, grancalcin, and ALG-2 (apoptosis-linked gene 2). We prepared a monoclonal antibody against human peflin. The antibody immunoprecipitated a 22-kDa protein as well as the 30-kDa protein from the lysate of Jurkat cells. Western blotting of the immunoprecipitates revealed that the 22-kDa protein corresponds to ALG-2. This was confirmed by Western blotting of the immunoprecipitates of epitope tagged peflin or ALG-2 whose cDNA expression constructs were transfected to human embryonic kidney (HEK) 293 cells. Gel filtration of the cytosolic fraction of Jurkat cells revealed co-elution of peflin and ALG-2 in fractions eluting earlier than recombinant ALG-2, further supporting the notion of heterodimerization of the two PEF proteins. Surprisingly, peflin dissociated from ALG-2 in the presence of Ca(2+). Peflin and ALG-2 co-localized in the cytoplasm, but ALG-2 was also detected in the nuclei as revealed by immunofluorescent staining and subcellular fractionation. Peflin was recovered in the cytosolic fraction in the absence of Ca(2+) but in the membrane/cytoskeletal fraction in the presence of Ca(2+). These results suggest that peflin has features common to those of other PEF proteins (dimerization and translocation to membranes) and may modulate the function of ALG-2 in Ca(2+) signaling. PMID- 11278428 TI - Biochemical analysis of point mutations in the 5'-3' exonuclease of DNA polymerase I of Streptococcus pneumoniae. Functional and structural implications. AB - To define the active site of the 5'-3' exonucleolytic domain of the Streptococcus pneumoniae DNA polymerase I (Spn pol I), we have constructed His-tagged Spn pol I fusion protein and introduced mutations at residues Asp(10), Glu(88), and Glu(114), which are conserved among all prokaryotic and eukaryotic 5' nucleases. The mutations, but not the fusion to the C-terminal end of the wild-type, reduced the exonuclease activity. The residual exonuclease activity of the mutant proteins has been kinetically studied, together with potential alterations in metal binding at the active site. Comparison of the catalytic rate and dissociation constant of the D10G, E114G, and E88K mutants and the control fusion protein support: (i) a critical function of Asp(10) in the catalytic event, (ii) a role of Glu(114) in the exonucleolytic reaction, being secondarily involved in both catalysis and DNA binding, and (iii) a nonessential function of Glu(88) for the exonuclease activity of Spn pol I. Moreover, the pattern of metal activation of the mutant proteins indicates that none of the three residues is a metal ligand at the active site. These findings and those previously obtained with D190A mutant of Spn pol I are discussed in relation to structural and mutational data for related 5' nucleases. PMID- 11278426 TI - Secoisolariciresinol dehydrogenase purification, cloning, and functional expression. Implications for human health protection. AB - Matairesinol is a central precursor in planta in the biosynthesis of numerous lignans, including that of the important antiviral and anticancer agent, podophyllotoxin. In this study, the approximately 32-kDa NAD-dependent secoisolariciresinol dehydrogenase, which catalyzes the enantiospecific conversion of (-)-secoisolariciresinol into (-)-matairesinol in Forsythia intermedia, was purified >6,000-fold to apparent homogeneity. The 831-base pair cDNA clone encoding this 277-amino acid protein was next obtained from a library constructed from F. intermedia stem tissue, whose fully functional recombinant protein, produced by expression of this cDNA in Escherichia coli, catalyzed the same enantiospecific conversion via the corresponding lactol intermediate. A homologous secoisolariciresinol dehydrogenase gene was also isolated from a Podophyllum peltatum rhizome cDNA library, whose 834-base pair cDNA clone encoded a 278-amino acid protein with a calculated molecular mass of approximately 32 kDa. Expression of this protein in E. coli produced a fully functional recombinant protein that also catalyzed the enantiospecific conversion of (-) secoisolariciresinol into (-)-matairesinol via the intermediary lactol. Various kinetic parameters were defined and established conversion of the intermediary lactol as being rate-limiting. With this overall enzymatic conversion now unambiguously defined, the entire biochemical pathway to the lignans, secoisolariciresinol and matairesinol, has been elucidated. Last, both secoisolariciresinol and matairesinol are metabolized in the gut of mammals, following digestion of high fiber dietary grains, seeds, and berries, into the so called "mammalian" lignans, enterodiol and enterolactone, respectively; these in turn confer significant protection against the onset of breast and prostate cancers. PMID- 11278429 TI - Isopentenyl pyrophosphate, a mycobacterial non-peptidic antigen, triggers delayed and highly sustained signaling in human gamma delta T lymphocytes without inducing eown-modulation of T cell antigen receptor. AB - The Vgamma9Vdelta2 T cell subset, which represents up to 90% of the circulating gammadelta T cells in humans, was shown to be activated, via the T cell receptor (TcR), by non-peptidic phosphorylated small organic molecules. These phosphoantigens, which are not presented by professional antigen-presenting cells, induce production of high amounts of interferon-gamma and tumor necrosis factor (TNF-alpha). To date, the specific signals triggered by these antigens have not been characterized. Here we analyze proximal and later intracellular signals triggered by isopentenyl pyrophosphate (IPP), a mycobacterial antigen that specifically stimulates Vgamma9Vdelta2 T cells, and compare these to signals induced by the non-physiological model using an anti-CD3 antibody. During antigenic stimulation we noticed that, except for the proximal p56(lck) signal, which is triggered early, the signals appear to be delayed and highly sustained. This delay, which likely accounts for the delay observed in TNF-alpha production, is discussed in terms of the ability of the antigen to cross-link and recruit transducing molecules mostly anchored to lipid rafts. Moreover, we demonstrate that, in contrast to anti-CD3 antibody, IPP does not induce down-modulation of the TcR.CD3 complex, which likely results in the highly sustained signaling and release of high levels of TNF-alpha. PMID- 11278431 TI - Granulocyte colony-stimulating factor induces ERK5 activation, which is differentially regulated by protein-tyrosine kinases and protein kinase C. Regulation of cell proliferation and survival. AB - Granulocyte colony-stimulating factor (G-CSF) plays a major role in the regulation of granulopoiesis. Treatment of cells with G-CSF has been shown to activate multiple signal transduction pathways. We show here that Erk5, a novel member of the MAPK family, and its specific upstream activator MEK5 were activated in response to incubation of cells with G-CSF. Different from other members of the MAPK family including Erk1/2, JNK, and p38, maximal activation of Erk5 by G-CSF required the C-terminal region of the G-CSF receptor. Genistein, a specific inhibitor of protein-tyrosine kinases, blocked G-CSF-induced Erk5 activation. In contrast, inhibition of protein kinase C activity increased G-CSF mediated activation of Erk5 and MEK5, whereas stimulation of protein kinase C activity inhibited activation of the two kinases by G-CSF. The proliferation of BAF3 cells in response to G-CSF was inhibited by expression of a dominant negative MEK5 but potentiated by expression of a constitutively active MEK5. Expression of the constitutively active MEK5 also increased the survival of BAF3 cells cultured in the absence of or in low concentrations of G-CSF. Together, these data implicate Erk5 as an important signaling component in the biological actions of G-CSF. PMID- 11278432 TI - Divergent N-terminal sequences of a deubiquitinating enzyme modulate substrate specificity. AB - Ubiquitin-specific processing proteases (UBPs) are characterized by a conserved core domain with surrounding divergent sequences, particularly at the N-terminal end. We previously cloned two isoforms of a testis UBP, UBP-t1 and UBP-t2, which contain identical core regions but distinct N termini that target the two isoforms to different subcellular locations (Lin, H., Keriel, A., Morales, C. R., Bedard, N., Zhao, Q., Hingamp, P., Lefrancois, S., Combaret, L., and Wing, S. S. (2000) Mol. Cell. Biol. 20, 6568-6578). To determine whether the N termini also influence the biochemical functions of the UBP, we expressed UBP-t1, UBP-t2, and the common core domain, UBP core, in Escherichia coli. The three isoforms cleaved branched triubiquitin at >20-fold faster rates than linear diubiquitin, suggesting that UBP-testis functions as an isopeptidase. Both N-terminal extensions inhibited the ability of UBP-core to generate free ubiquitin when linked in a peptide bond with itself, another peptide, or to small adducts. The N terminal extension of UBP-t2 increased the ability of UBP-core to cleave branched triubiquitin. UBP-core removed ubiquitin from testis ubiquitinated proteins more rapidly than UBP-t2 and UBP-t1. Thus, UBP enzymes appear to contain a catalytic core domain, the activities and specificities of which can be modulated by N terminal extensions. These divergent N termini can alter localization and confer multiple functions to the various members of the large UBP family. PMID- 11278430 TI - Hypotension, autonomic failure, and cardiac hypertrophy in transgenic mice overexpressing the alpha 1B-adrenergic receptor. AB - alpha(1)-Adrenergic receptors (alpha(1A), alpha(1B), and alpha(1D)) are regulators of systemic arterial blood pressure and blood flow. Whereas vasoconstrictory action of the alpha(1A) and alpha(1D) subtypes is thought to be mainly responsible for this activity, the role of the alpha(1B)-adrenergic receptor (alpha(1B)AR) in this process is controversial. We have generated transgenic mice that overexpress either wild type or constitutively active alpha(1B)ARs. Transgenic expression was under the control of the isogenic promoter, thus assuring appropriate developmental and tissue-specific expression. Cardiovascular phenotypes displayed by transgenic mice included myocardial hypertrophy and hypotension. Indicative of cardiac hypertrophy, transgenic mice displayed an increased heart to body weight ratio, which was confirmed by the echocardiographic finding of an increased thickness of the interventricular septum and posterior wall. Functional deficits included an increased isovolumetric relaxation time, a decreased heart rate, and cardiac output. Transgenic mice were hypotensive and exhibited a decreased pressor response. Vasoconstrictory regulation by alpha(1B)AR was absent as shown by the lack of phenylephrine-induced contractile differences between ex vivo mesenteric artery preparations. Plasma epinephrine, norepinephrine, and cortisol levels were also reduced in transgenic mice, suggesting a loss of sympathetic nerve activity. Reduced catecholamine levels together with basal hypotension, bradycardia, reproductive problems, and weight loss suggest autonomic failure, a phenotype that is consistent with the multiple system atrophy-like neurodegeneration that has been reported previously in these mice. These results also suggest that this receptor subtype is not involved in the classic vasoconstrictory action of alpha(1)ARs that is important in systemic regulation of blood pressure. PMID- 11278433 TI - Inhibition of human endogenous retrovirus-K10 protease in cell-free and cell based assays. AB - A full-length and C-terminally truncated version of human endogenous retrovirus (HERV)-K10 protease were expressed in Escherichia coli and purified to homogeneity. Both versions of the protease efficiently processed HERV-K10 Gag polyprotein substrate. HERV-K10 Gag was also cleaved by human immunodeficiency virus, type 1 (HIV-1) protease, although at different sites. To identify compounds that could inhibit protein processing dependent on the HERV-K10 protease, a series of cyclic ureas that had previously been shown to inhibit HIV 1 protease was tested. Several symmetric bisamides acted as very potent inhibitors of both the truncated and full-length form of HERV-K10 protease, in subnanomolar or nanomolar range, respectively. One of the cyclic ureas, SD146, can inhibit the processing of in vitro translated HERV-K10 Gag polyprotein substrate by HERV-K10 protease. In addition, in virus-like particles isolated from the teratocarcinoma cell line NCCIT, there is significant accumulation of Gag and Gag-Pol precursors upon treatment with SD146, suggesting the compound efficiently blocks HERV-K Gag processing in cells. This is the first report of an inhibitor able to block cell-associated processing of Gag polypeptides of an endogenous retrovirus. PMID- 11278434 TI - Structural and functional features of the transmembrane domain of the Na,K-ATPase beta subunit revealed by tryptophan scanning. AB - In oligomeric P2-ATPases such as Na,K- and H,K-ATPases, beta subunits play a fundamental role in the structural and functional maturation of the catalytic alpha subunit. In the present study we performed a tryptophan scanning analysis on the transmembrane alpha-helix of the Na,K-ATPase beta1 subunit to investigate its role in the stabilization of the alpha subunit, the endoplasmic reticulum exit of alpha-beta complexes, and the acquisition of functional properties of the Na,K-ATPase. Single or multiple tryptophan substitutions in the beta subunits transmembrane domain had no significant effect on the structural maturation of alpha subunits expressed in Xenopus oocytes nor on the level of expression of functional Na,K pumps at the cell surface. Furthermore, tryptophan substitutions in regions of the transmembrane alpha-helix containing two GXXXG transmembrane helix interaction motifs or a cysteine residue, which can be cross-linked to transmembrane helix M8 of the alpha subunit, had no effect on the apparent K(+) affinity of Na,K-ATPase. On the other hand, substitutions by tryptophan, serine, alanine, or cysteine, but not by phenylalanine of two highly conserved tyrosine residues, Tyr(40) and Tyr(44), on another face of the transmembrane helix, perturb the transport kinetics of Na,K pumps in an additive way. These results indicate that at least two faces of the beta subunits transmembrane helix contribute to inter- or intrasubunit interactions and that two tyrosine residues aligned in the beta subunits transmembrane alpha-helix are determinants of intrinsic transport characteristics of Na,K-ATPase. PMID- 11278435 TI - Akt, MAPK (Erk1/2), and p38 act in concert to promote apoptosis in response to ErbB receptor family inhibition. AB - The ErbB receptor family is implicated in the malignant transformation of several tumor types and is overexpressed frequently in breast, ovarian, and other tumors. The mechanism by which CI-1033 and gemcitabine, either singly or in combination, kill tumor cells was examined in two breast lines, MDA-MB-453 and BT474; both overexpress the ErbB-2 receptor. CI-1033, a potent inhibitor of the ErbB family of receptor tyrosine kinases, reduced levels of activated Akt in MDA-MB-453 cells. This effect alone, however, did not induce apoptosis in these cells. Gemcitabine treatment resulted in a moderate increase in the percentage of apoptotic cells that was accompanied by activation of p38 and MAPK (ERK1/2). CI 1033 given 24 h after gemcitabine produced a significant increase in the apoptotic fraction over treatment with either drug alone. During the combined treatment p38 remained activated, whereas Akt and activated MAPK were suppressed. Substitution of CI-1033 with the phosphatidylinositol 3-kinase inhibitor LY294002 and the MAPK/ERK kinase inhibitor PD 098059 in combination with gemcitabine produced the same results as the combination of CI-1033 and gemcitabine. p38 suppression by SB203580 prevented the enhanced cell kill by CI-1033. In contrast to MDA-MB-453, BT474 cells exhibited activated p38 under unstressed conditions as well as activated Akt and MAPK. Treatment of BT474 cells with CI-1033 inhibited both the phosphorylation of Akt and MAPK and resulted in a 47% apoptotic fraction. Gemcitabine did not cause apoptosis in the BT474 cells. These data indicate that suppression of Akt and MAPK in the presence of activated p38 results in cell death and a possible mechanism for the enhanced apoptosis produced by the combination of CI-1033 and gemcitabine in MDA-MB-453 cells. Furthermore, tumors that depend on ErbB receptor signaling for survival and exhibit activated p38 in the basal state may be susceptible to apoptosis by CI 1033 as a single agent. PMID- 11278436 TI - The tyrosine kinase ACK1 associates with clathrin-coated vesicles through a binding motif shared by arrestin and other adaptors. AB - One target for the small GTPase Cdc42 is the nonreceptor tyrosine kinase activated Cdc42-associated kinase (ACK), which binds selectively to Cdc42.GTP. We report that ACK1 can associate directly with the heavy chain of clathrin. A central region in ACK1 containing a conserved motif behaves as a clathrin adaptor and competes with beta-arrestin for a common binding site on the clathrin N terminal head domain. Overexpressed ACK1 perturbs clathrin distribution, an activity dependent on the presence of C-terminal "adaptor" sequences that are also present in the related nonkinase gene 33. ACK1 interacts with the adaptor Nck via SH3 interactions but does not form a trimeric complex with p21-activated serine/threonine kinase, which also binds Nck. Stable low level expression of green fluorescent protein-ACK1 in NIH 3T3 cells has been used to localize ACK1 to clathrin-containing vesicles. The co-localization of ACK1 in vivo with clathrin and AP-2 indicates that it participates in trafficking, underlying an ability to increase receptor-mediated transferrin uptake. PMID- 11278437 TI - Thrombin regulates vascular smooth muscle cell growth and heat shock proteins via the JAK-STAT pathway. AB - The growth-stimulating effects of thrombin are mediated primarily via activation of a G protein-coupled receptor, PAR-1. Because PAR-1 has no intrinsic tyrosine kinase activity, yet requires tyrosine phosphorylation events to induce mitogenesis, we investigated the role of the Janus tyrosine kinases (JAKs) in thrombin-mediated signaling. JAK2 was activated rapidly in rat vascular smooth muscle cells (VSMC) treated with thrombin, and signal transducers and activators of transcription (STAT1 and STAT3) were phosphorylated and translocated to the nucleus in a JAK2-dependent manner. AG-490, a JAK2-specific inhibitor, and a dominant negative JAK2 mutant inhibited thrombin-induced ERK2 activity and VSMC proliferation suggesting that JAK2 is upstream of the Ras/Raf/MEK/ERK pathway. To elucidate the functional significance of JAK-STAT activation, we studied the effect of thrombin on heat shock protein (Hsp) expression, based upon the following: 1) reports that thrombin stimulates reactive oxygen species production in VSMC; 2) the putative role of Hsps in modulating cellular responses to reactive oxygen species; and 3) the presence of functional STAT1/3-binding sites in Hsp70 and Hsp90beta promoters. Indeed, thrombin up-regulated Hsp70 and Hsp90 protein expression via enhanced binding of STATs to cognate binding sites in the Hsp70 and Hsp90 promoters. Together, these results suggest that JAK-STAT pathway activation is necessary for thrombin-induced VSMC growth and Hsp gene expression. PMID- 11278438 TI - A novel group of phospholipase A2s preferentially expressed in type 2 helper T cells. AB - We report a novel phospholipase A(2) (PLA(2)), group XII (GXII) PLA(2), distinct from other cysteine-rich groups with a catalytic histidine motif, by its 20-kDa size and distribution of the 14 cysteine residues within the protein. Alternative spliced forms with distinct subcellular localization, designated GXII-1 and GXII 2, were identified by reverse transcription-polymerase chain reaction. Importantly, GXII PLA(2)s, in particular GXII-2 PLA(2), and group V PLA(2), but not group X PLA(2), were selectively expressed in murine type 2 helper T (Th2) clones and in vitro differentiated mouse CD4 Th2 cells as compared with type 1 helper T clones and cells. Stimulation with anti-CD3 appreciably up-regulated expression of GXII PLA(2)s and group V PLA(2) by steady state analysis of the Th2 cells as compared with type 1 helper T cells. These results suggest that group XII and group V PLA(2)s might participate in helper T cell immune response through release of immediate second signals and generation of downstream eicosanoids. PMID- 11278439 TI - Muc13, a novel human cell surface mucin expressed by epithelial and hemopoietic cells. AB - Transmembrane mucins are glycoproteins involved in barrier function in epithelial tissues. To identify novel transmembrane mucin genes, we performed a tblastn search of the GenBanktrade mark EST data bases with a serine/threonine-rich search string, and a rodent gene expressed in bone marrow was identified. We determined the cDNA sequence of the human orthologue of this gene, MUC13, which localizes to chromosome band 3q13.3 and generates 3.2-kilobase pair transcripts encoding a 512-amino acid protein comprised of an N-terminal mucin repeat domain, three epidermal growth factor-like sequences, a SEA module, a transmembrane domain, and a cytoplasmic tail (GenBanktrade mark accession no. ). MUC13 mRNA is expressed most highly in the large intestine and trachea, and at moderate levels in the kidney, small intestine, appendix, and stomach. In situ hybridization in murine tissues revealed expression in intestinal epithelial and lymphoid cells. Immunohistochemistry demonstrated the human MUC13 protein on the apical membrane of both columnar and goblet cells in the gastrointestinal tract, as well as within goblet cell thecae, indicative of secretion in addition to presence on the cell surface. MUC13 is cleaved, and the beta-subunit containing the cytoplasmic tail undergoes homodimerization. Including MUC13, there are at least five cell surface mucins expressed in the gastrointestinal tract. PMID- 11278440 TI - A novel MaxiK splice variant exhibits dominant-negative properties for surface expression. AB - We identified a novel MaxiK alpha subunit splice variant (SV1) from rat myometrium that is also present in brain. SV1 has a 33-amino acid insert in the S1 transmembrane domain that does not alter S1 overall hydrophobicity, but makes the S0-S1 linker longer. SV1 was transfected in HEK293T cells and studied using immunocytochemistry and electrophysiology. In non-permeabilized cells, N-terminal c-Myc- or C-terminal green fluorescent protein-tagged SV1 displayed no surface labeling or currents. The lack of SV1 functional expression was due to endoplasmic reticulum (ER) retention as determined by colabeling experiments with a specific ER marker. To explore the functional role of SV1, we coexpressed SV1 with the alpha (human SLO) and beta1 (KCNMB1) subunits of the MaxiK channel. Coexpression of SV1 inhibited surface expression of alpha and beta1 subunits approximately 80% by trapping them in the ER. This inhibition seems to be specific for MaxiK channel subunits since SV1 was unable to prevent surface expression of the Kv4.3 channel or to interact with green fluorescent protein. These results indicate a dominant-negative role of SV1 in MaxiK channel expression. Moreover, they reveal down-regulation by splice variants as a new mechanism that may contribute to the diverse levels of MaxiK channel expression in non-excitable and excitable cells. PMID- 11278441 TI - Structural and functional identification of major histocompatibility complex class I-restricted self-peptides as naturally occurring molecular mimics of viral antigens. Possible role in CD8+ T cell-mediated, virus-induced autoimmune disease. AB - Structural similarity (molecular mimicry) between viral epitopes and self peptides can lead to the induction of autoaggressive CD4(+) as well as CD8(+) T cell responses. Based on the flexibility of T cell receptor/antigen/major histocompatibility complex recognition, it has been proposed that a self-peptide could replace a viral epitope for T cell recognition and therefore participate in pathophysiological processes in which T cells are involved. To address this issue, we used, as a molecular model of viral antigen, the H-2D(b)-restricted immunodominant epitope nucleoprotein (NP)-(396-404) (FQPQNGQFI) of lymphocytic choriomeningitis virus (LCMV). We identified peptide sequences from murine self proteins that share structural and functional homology with LCMV NP-(396-404) and that bound to H-2D(b) with high affinity. One of these self-peptides, derived from tumor necrosis factor receptor I (FGPSNWHFM, amino acids 302-310), maintained LCMV-specific CD8(+) T cells in an active state as observed both in vitro in cytotoxic assays and in vivo in a model of virus-induced autoimmune diabetes, the rat insulin promoter-LCMV NP transgenic mouse. The natural occurrence and molecular concentration at the surface of H-2(b) spleen cells of tumor necrosis factor receptor I-(302-310) were determined by on-line micro-high pressure liquid chromatography/mass spectrometry and supported its biological relevance. PMID- 11278442 TI - Ligand binding and functional properties of betaglycan, a co-receptor of the transforming growth factor-beta superfamily. Specialized binding regions for transforming growth factor-beta and inhibin A. AB - Betaglycan, also known as the transforming growth factor-beta (TGF-beta) type III receptor, is a membrane-anchored proteoglycan that binds TGF-beta via its core protein. Deletion mutagenesis analysis has revealed two regions of betaglycan ectodomain capable of binding TGF-beta: one at the amino-terminal half, the endoglin-related region (Lopez-Casillas, F., Payne, H., Andres, J. L., and Massague, J. (1994) J. Cell Biol. 124, 557-568), and the other at the carboxyl terminal half, the uromodulin-related region (Pepin, M.-C., Beauchemin, M., Plamondon, J., and O'Connor-McCourt, M. D. (1994) Proc. Natl. Acad. Sci. U. S. A 91, 6997-7001). In the present work we have functionally characterized these ligand binding regions. Similar to the wild type receptor, both regions bind TGF beta2 with higher affinity than TGF-beta1. However, only the endoglin-related region increases the TGF-beta2 labeling of the TGF-beta type II receptor, the so called "TGF-beta -presentation" function of the wild type receptor. Despite this preference, both regions as well as the wild type receptor mediate the TGF-beta2 dependent Smad2 phosphorylation, indicating that they can function indistinguishably as TGF-beta-enhancing co-receptors. On the other hand, we found that the recently described ability of the wild type betaglycan to bind inhibin A is a property of the core protein that resides in the uromodulin-related region. Binding competition experiments indicate that this region binds inhibin and TGF beta with the following relative affinities: TGF-beta2 > inhibin A > TGF-beta1. All together, the present results suggest that betaglycan ectodomain is endowed with two bona fide independent ligand binding domains that can perform specialized functions as co-receptors of distinct members of the TGF-beta superfamily. PMID- 11278443 TI - Cell cycle and biochemical effects of PD 0183812. A potent inhibitor of the cyclin D-dependent kinases CDK4 and CDK6. AB - Progression through the G1 phase of the cell cycle requires phosphorylation of the retinoblastoma gene product (pRb) by the cyclin D-dependent kinases CDK4 and CDK6, whose activity can specifically be blocked by the CDK inhibitor p16(INK4A). Misregulation of the pRb/cyclin D/p16(INK4A) pathway is one of the most common events in human cancer and has lead to the suggestion that inhibition of cyclin D dependent kinase activity may have therapeutic value as an anticancer treatment. Through screening of a chemical library, we initially identified the [2,3 d]pyridopyrimidines as inhibitors of CDK4. Chemical modification resulted in the identification of PD 0183812 as a potent and highly selective inhibitor of both CDK4 and CDK6 kinase activity, which is competitive with ATP. Flow cytometry experiments showed that of the cell lines tested, only those expressing pRb demonstrated a G1 arrest when treated with PD 0183812. This arrest correlated in terms of incubation time and potency with a loss of pRb phosphorylation and a block in proliferation, which was reversible. These results suggest a potential use of this chemical class of compounds as therapeutic agents in the treatment of tumors with functional pRb, possessing cell cycle aberrations in other members of the pRb/cyclin D/p16(INK4A) pathway. PMID- 11278444 TI - Protein-tyrosine kinase Pyk2 mediates endothelin-induced p38 MAPK activation in glomerular mesangial cells. AB - Endothelin-1 (ET-1), a member of a family of 21 amino acid peptides possessing vasoconstrictor properties, is known to stimulate mesangial cell proliferation. In this study, ET-1 (100 nm) induced a rapid activation of p21(ras) in human glomerular mesangial cells (HMC). Inhibition of Src family tyrosine kinase activation with [4-Amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine] or chelation of intracellular free calcium with 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester significantly decreased ET 1dependent p21(ras) activation and suggested the involvement of the cytoplasmic proline-rich tyrosine kinase Pyk2. We have observed that Pyk2 was expressed in HMC and was tyrosine-phosphorylated within 5 min of ET-1 treatment. ET-1-induced activation of Pyk2 was further confirmed using phospho-specific anti-Pyk2 antibodies. Surprisingly, Src kinase activity was required upstream of ET-1 induced autophosphorylation of Pyk2. To determine whether Pyk2 autophosphorylation mediated ET-1-dependent p21(ras) activation, adenovirus mediated transfer was employed to express a dominant-negative form of Pyk2 (CRNK). CRNK expression inhibited ET-1-induced endogenous Pyk2 autophosphorylation, but did not abolish ET-1-mediated increases in GTP-bound p21(ras) levels. ET-1-induced activation of the p38 MAPK (but not ERK) pathway was inhibited in HMC and in rat glomerular mesangial cells expressing the dominant-negative form of Pyk2. These findings suggest that the engagement of Pyk2 is important for ET-1-mediated p38 MAPK activation and hence the biological effect of this peptide in mesangial cells. PMID- 11278445 TI - Identification of the mechanisms regulating the differential activation of the mapk cascade by epidermal growth factor and nerve growth factor in PC12 cells. AB - In PC12 cells, epidermal growth factor (EGF) transiently stimulates the mitogen activated protein (MAP) kinases, ERK1 and ERK2, and provokes cellular proliferation. In contrast, nerve growth factor (NGF) stimulation leads to the sustained activation of the MAPKs and subsequently to neuronal differentiation. It has been shown that both the magnitude and longevity of MAPK activation governs the nature of the cellular response. The activations of MAPKs are dependent upon two distinct small G-proteins, Ras and Rap1, that link the growth factor receptors to the MAPK cascade by activating c-Raf and B-Raf, respectively. We found that Ras was transiently stimulated upon both EGF and NGF treatment of PC12 cells. However, EGF transiently activated Rap1, whereas NGF stimulated prolonged Rap1 activation. The activation of the ERKs was due almost exclusively (>90%) to the action of B-Raf. The transient activation of the MAPKs by EGF was a consequence of the formation of a short lived complex assembling on the EGF receptor itself, composed of Crk, C3G, Rap1, and B-Raf. In contrast, NGF stimulation of the cells resulted in the phosphorylation of FRS2. FRS2 scaffolded the assembly of a stable complex of Crk, C3G, Rap1, and B-Raf resulting in the prolonged activation of the MAPKs. Together, these data provide a signaling link between growth factor receptors and MAPK activation and a mechanistic explanation of the differential MAPK kinetics exhibited by these growth factors. PMID- 11278447 TI - Monitoring receptor oligomerization using time-resolved fluorescence resonance energy transfer and bioluminescence resonance energy transfer. The human delta opioid receptor displays constitutive oligomerization at the cell surface, which is not regulated by receptor occupancy. AB - Oligomerization of the human delta-opioid receptor and its regulation by ligand occupancy were explored following expression in HEK293 cells using each of co immunoprecipitation of differentially epitope-tagged forms of the receptor, bioluminescence resonance energy transfer and time-resolved fluorescence resonance energy transfer. All of the approaches identified constitutively formed receptor oligomers, and the time-resolved fluorescence studies confirmed the presence of such homo-oligomers at the cell surface. Neither the agonist ligand [d-Ala(2),d-Leu(5)]enkephalin nor the inverse agonist ligand ICI174864 were able to modulate the oligomerization status of this receptor. Interactions between co expressed delta-opioid receptors and beta(2)-adrenoreceptors were observed in co immunoprecipitation studies. Such hetero-oligomers could also be detected using bioluminescence resonance energy transfer although the signal obtained was substantially smaller than for homo-oligomers of either receptor type. Signal corresponding to the delta-opioid receptor-beta(2)-adrenoreceptor hetero-oligomer was increased in the presence of agonist for either receptor. However, substantial levels of this hetero-oligomer were not detected at the cell surface using time-resolved fluorescence resonance energy transfer. These studies demonstrate that, following transient transfection of HEK293 cells, constitutively formed oligomers of the human delta-opioid receptor can be detected by a variety of approaches. However, these are not regulated by ligand occupancy. They also indicate that time-resolved fluorescence resonance energy transfer represents a means to detect such oligomers at the cell surface in populations of intact cells. PMID- 11278446 TI - ATM-dependent phosphorylation of human Rad9 is required for ionizing radiation induced checkpoint activation. AB - ATM (ataxia-telangiectasia-mutated) is a Ser/Thr kinase involved in cell cycle checkpoints and DNA repair. Human Rad9 (hRad9) is the homologue of Schizosaccharomyces pombe Rad9 protein that plays a critical role in cell cycle checkpoint control. To examine the potential signaling pathway linking ATM and hRad9, we investigated the modification of hRad9 in response to DNA damage. Here we show that hRad9 protein is constitutively phosphorylated in undamaged cells and undergoes hyperphosphorylation upon treatment with ionizing radiation (IR), ultraviolet light (UV), and hydroxyurea (HU). Interestingly, hyperphosphorylation of hRad9 induced by IR is dependent on ATM. Ser(272) of hRad9 is phosphorylated directly by ATM in vitro. Furthermore, hRad9 is phosphorylated on Ser(272) in response to IR in vivo, and this modification is delayed in ATM-deficient cells. Expression of hRad9 S272A mutant protein in human lung fibroblast VA13 cells disturbs IR-induced G(1)/S checkpoint activation and increased cellular sensitivity to IR. Together, our results suggest that the ATM-mediated phosphorylation of hRad9 is required for IR-induced checkpoint activation. PMID- 11278448 TI - Structure-function of the putative I-domain within the integrin beta 2 subunit. AB - The central region (residues 125-385) of the integrin beta(2) subunit is postulated to adopt an I-domain-like fold (the beta(2)I-domain) and to play a critical role in ligand binding and heterodimer formation. To understand structure-function relationships of this region of beta(2), a homolog-scanning mutagenesis approach, which entails substitution of nonconserved hydrophilic sequences within the beta(2)I-domain with their homologous counterparts of the beta(1)I-domain, has been deployed. This approach is based on the premise that beta(1) and beta(2) are highly homologous, yet recognize different ligands. Altogether, 16 segments were switched to cover the predicted outer surface of the beta(2)I-domain. When these mutant beta(2) subunits were transfected together with wild-type alpha(M) in human 293 cells, all 16 beta(2) mutants were expressed on the cell surface as heterodimers, suggesting that these 16 sequences within the beta(2)I-domain are not critically involved in heterodimer formation between the alpha(M) and beta(2) subunits. Using these mutant alpha(M)beta(2) receptors, we have mapped the epitopes of nine beta(2)I-domain specific mAbs, and found that they all recognized at least two noncontiguous segments within this domain. The requisite spatial proximity among these non-linear sequences to form the mAb epitopes supports a model of an I-domain-like fold for this region. In addition, none of the mutations that abolish the epitopes of the nine function-blocking mAbs, including segment Pro(192)-Glu(197), destroyed ligand binding of the alpha(M)beta(2) receptor, suggesting that these function-blocking mAbs inhibit alpha(M)beta(2) function allosterically. Given the recent reports implicating the segment equivalent to Pro(192)-Glu(197) in ligand binding by beta(3) integrins, these data suggest that ligand binding by the beta(2) integrins occurs via a different mechanism than beta(3). Finally, both the conformation of the beta(2)I domain and C3bi binding activity of alpha(M)beta(2) were dependent on a high affinity Ca(2+) binding site (K(d) = 105 microm), which is most likely located within this region of beta(2). PMID- 11278449 TI - Muscarinic acetylcholine receptor regulation of TRP6 Ca2+ channel isoforms. Molecular structures and functional characterization. AB - In this study, we report the molecular cloning of cDNAs encoding three distinct isoforms of rat (r) TRP6 Ca(2+) channels. The longest isoform, rTRP6A, contains 930 amino acid residues; rTRP6B lacks 54 amino acids (3-56) at the N terminus, and rTRP6C is missing an additional 68 amino acids near the C terminus. Transient transfection of COS cells with expression vectors encoding rTRP6A or rTRP6B increased Ca(2+) influx and gave rise to a novel Ba(2+) influx after activation of M(5) muscarinic acetylcholine receptors. By contrast, passive depletion of intracellular Ca(2+) stores with thapsigargin did not induce Ba(2+) influx in cells expressing rTRP6 isoforms. Ba(2+) influx was also stimulated in rTRP6A expressing cells after exposure to the diacylglycerol analog, 1-oleoyl-2-acetyl sn-glycerol (OAG), but rTRP6B-expressing cells failed to show OAG-induced Ba(2+) influx. Expression of a rTRP6 N-terminal fragment of rTRP6B or rTRP6A antisense RNA blocked M(5) muscarinic acetylcholine receptor-dependent Ba(2+) influx in COS cells that were transfected with rTRP6 cDNAs. Together these results suggest that rTRP6 participates in the formation of Ca(2+) channels that are regulated by a G protein-coupled receptor, but not by intracellular Ca(2+) stores. In contrast to the results we obtained with rTRP6A and rTRP6B, cells expressing rTRP6C showed no increased Ca(2+) or Ba(2+) influxes after stimulation with carbachol and also did not show OAG-induced Ba(2+) influx. Glycosylation analysis indicated that rTRP6A and rTRP6B are glycosylated in COS cells, but that rTRP6C is mostly not glycosylated. Together these results suggest that the N terminus (3-56 amino acids) is crucial for the activation of rTRP6A by diacylglycerol and that the 735 802 amino acid segment located just downstream from the 6th transmembrane segment may be required for processing of the rTRP6 protein. PMID- 11278450 TI - Riboflavin synthase of Escherichia coli. Effect of single amino acid substitutions on reaction rate and ligand binding properties. AB - Conserved amino acid residues of riboflavin synthase from Escherichia coli were modified by site-directed mutagenesis. Replacement or deletion of phenylalanine 2 afforded catalytically inactive proteins. S41A and H102Q mutants had substantially reduced reaction velocities. Replacements of various other conserved polar residues had little impact on catalytic activity. (19)F NMR protein perturbation experiments using a fluorinated intermediate analog suggest that the N-terminal sequence motif MFTG is part of one of the substrate-binding sites of the protein. PMID- 11278451 TI - A GTP-dependent vertebrate-type phosphoenolpyruvate carboxykinase from Mycobacterium smegmatis. AB - This is the first report on a bacterial verterbrate-type GTP-dependent phosphoenolpyruvate carboxykinase (PCK). The pck gene of Mycobacterium smegmatis was cloned. The recombinant PCK was overexpressed in Escherichia coli in a soluble form and with high activity. The purified enzyme was found to be monomeric (72 kDa), thermophilic (optimum temperature, 70 degrees C), very stable upon storage at 4 degrees C, stimulated by thiol-containing reducing agents, and inhibited by oxalate and by alpha-ketoglutarate. The requirement for a divalent cation for activity was fulfilled best by Mn(2+) and Co(2+) and poorly by Mg(2+). At 37 degrees C, the highest V(m) value (32.5 units/mg) was recorded with Mn(2+) and in the presence of 37 mm dithiothreitol (DTT). The presence of Mg(2+) (2 mm) greatly lowered the apparent K(m) values for Mn(2+) (by 144-fold in the presence of DTT and by 9.4-fold in the absence of DTT) and Co(2+) (by 230-fold). In the absence of DTT but in the presence of Mg(2+) (2 mm) as the co-divalent cation, Co(2+) was 21-fold more efficient than Mn(2+). For producing oxaloacetate, the enzyme utilized both GDP and IDP; ADP served very poorly. The apparent K(m) values for phosphoenolpyruvate, GDP, and bicarbonate were >100, 66, and 8300 micrometer, respectively, whereas those for GTP and oxaloacetate (for the phosphoenolpyruvate formation activity) were 13 and 12 microm, respectively. Thus, this enzyme preferred the gluconeogenesis/glycerogenesis direction. This property fits the suggestion that in M. smegmatis, pyruvate carboxylase is not anaplerotic but rather gluconeogenic (Mukhopadhyay, B., and Purwantini, E. (2000) Biochim. Biophys. Acta. 1475, 191-206). Both in primary structure and kinetic properties, the mycobacterial PCK was very similar to its vertebrate-liver counterparts and thus could serve as a model for these enzymes; examples for several immediate targets are presented. PMID- 11278453 TI - Activation of the Rap1 guanine nucleotide exchange gene, CalDAG-GEF I, in BXH-2 murine myeloid leukemia. AB - Here we report the recurrent proviral activation of the Rap1-specific guanine nucleotide exchange factor CalDAG-GEF I (Kawasaki, H., Springett, G. M., Toki, S., Canales, J. J., Harlan, P., Blumenstiel, J. P., Chen, E. J., Bany, I. A., Mochizuki, N., Ashbacher, A., Matsuda, M., Housman, D. E., and Graybiel, A. M. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 13278-13283; Correction (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 318) gene in BXH-2 acute myeloid leukemia. We also show that CalDAG-GEF I encodes two protein isoforms, a full-length isoform (CalDAG-GEF Ia) and a C-terminally truncated isoform (CalDAG-GEF Ib). Expression of the full-length CalDAG-GEF Ia isoform in Rat2 fibroblasts enhances growth in low serum, whereas expression in Swiss 3T3 cells causes morphological transformation and increased saturation density. In FDCP1 myeloid cells, CalDAG GEF Ia expression increases growth and saturation density in the presence of the diacylglycerol analogs phorbol 12-myristate 13-acetate (PMA), which activates CalDAG-GEF Ia exchange activity. Likewise, in 32Dcl3 myeloblast cells, CalDAG-GEF Ia expression increases cell adherence to fibronectin in response to PMA and calcium ionophore and allows higher saturation densities and prolonged growth on fibronectin-coated plates. These effects were correlated with increased Rap1, but not Ras, protein activation following PMA and calcium ionophore treatment. Our results suggest that Rap1-GTP delivers signals that favor progression through the cell cycle and morphological transformation. The identification of CalDAG-GEF I as a proto-oncogene in BXH-2 acute myeloid leukemia is the first evidence implicating Rap1 signaling in myeloid leukemia. PMID- 11278452 TI - Physical and functional interactions of Galphaq with Rho and its exchange factors. AB - Recent reports have shown that several heterotrimeric protein-coupled receptors that signal through Galpha(q) can induce Rho-dependent responses, but the pathways that mediate the interaction between Galpha(q) and Rho have not yet been identified. In this report we present evidence that Galpha(q) expressed in COS-7 cells coprecipitates with the Rho guanine nucleotide exchange factor (GEF) Lbc. Furthermore, Galpha(q) expression enhances Rho-dependent responses. Coexpressed Galpha(q) and Lbc have a synergistic effect on the Rho-dependent rounding of 1321N1 astrocytoma cells. In addition, serum response factor-dependent gene expression, as assessed by the SRE.L reporter gene, is synergistically activated by Galpha(q) and Rho GEFs. The synergistic effect of Galpha(q) on this response is inhibited by C3 exoenzyme and requires phospholipase C activation. Surprisingly, expression of Galpha(q), in contrast to that of Galpha(12) and Galpha(13), does not increase the amount of activated Rho. We also observe that Galpha(q) enhances SRE.L stimulation by activated Rho, indicating that the effect of Galpha(q) occurs downstream of Rho activation. Thus, Galpha(q) interacts physically and/or functionally with Rho GEFs; however this does not appear to lead to or result from increased activation of Rho. We suggest that Galpha(q) generated signals enhance responses downstream of Rho activation. PMID- 11278454 TI - Binding sites for Rev and ASF/SF2 map to a 55-nucleotide purine-rich exonic element in equine infectious anemia virus RNA. AB - The equine infectious anemia virus (EIAV) Rev protein (ERev) negatively regulates its own synthesis by inducing alternative splicing of its mRNA. This bicistronic mRNA contains four exons; exons 1 and 2 encode Tat, and exons 3 and 4 encode Rev. When Rev is expressed, exon 3 is skipped to produce an mRNA that contains only exons 1, 2, and 4. The interaction of ERev with its cis-acting RNA response element, the RRE, is also essential for nuclear export of intron-containing viral mRNAs that encode structural and enzymatic gene products. The primary ERev binding site and the manner in which ERev interacts with RNA or cellular proteins to exert its regulatory function have not been defined. We have performed in vitro RNA binding experiments to show that recombinant ERev binds to a 55 nucleotide, purine-rich tract proximal to the 5' splice site of exon 3. Because of its proximity to the 5' splice site and since it contains elements related to consensus exonic splicing enhancer sequences, we asked whether cellular proteins recognize the EIAV RRE. The cellular protein, ASF/SF2, a member of the serine- and arginine-rich family of splicing factors (SR proteins) bound to repeated sequences within the 55-nucleotide RRE region. Electrophoretic mobility shift and UV cross-linking experiments indicated that ERev and SR proteins bind simultaneously to the RRE. Furthermore, in vitro protein-protein interaction studies revealed an association between ERev and SR proteins. These data suggest that EIAV Rev-induced exon skipping observed in vivo may be initiated by simultaneous binding of Rev and SR proteins to the RRE that alter the subsequent assembly or catalytic activity of the spliceosomal complex. PMID- 11278455 TI - Genomic structure of the promoters of the human estrogen receptor-alpha gene demonstrate changes in chromatin structure induced by AP2gamma. AB - Expression of human estrogen receptor-alpha (ERalpha) involves the activity from several promoters that give rise to alternate untranslated 5' exons. However, the genomic locations of the alternate 5' exons have not been reported previously. We have developed a contig map of the human ERalpha gene that includes all of the known alternate 5' exons. By using S1 nuclease and 5'- rapid amplification of cDNA ends, the cap sites for the alternate ERalpha transcripts E and H were identified. DNase I-hypersensitive sites specific to ERalpha-positive cells were associated with each of the cap sites. A DNase I-hypersensitive site, HS1, was localized to binding sites for AP2 in the untranslated region of exon 1 and was invariably present in the chromatin structure of ERalpha-positive cells. Overexpression of AP2gamma in human mammary epithelial cells generated the HS1 hypersensitive site. The ERalpha promoter was induced by AP2gamma in mammary epithelial cells, and trans-activation was dependent upon the region of the promoter containing the HS1 site. These results demonstrate that AP2gamma trans activates the ERalpha gene in hormone-responsive tumors by inducing changes in the chromatin structure of the ERalpha promoter. These data are further evidence for a critical role for AP2 in the oncogenesis of hormone-responsive breast cancers. PMID- 11278456 TI - Biochemical basis for the dominant inheritance of hypermethioninemia associated with the R264H mutation of the MAT1A gene. A monomeric methionine adenosyltransferase with tripolyphosphatase activity. AB - Methionine adenosyltransferase (MAT) catalyzes the synthesis of S adenosylmethionine (AdoMet), the main alkylating agent in living cells. Additionally, in the liver, MAT is also responsible for up to 50% of methionine catabolism. Humans with mutations in the gene MAT1A, the gene that encodes the catalytic subunit of MAT I and III, have decreased MAT activity in liver, which results in a persistent hypermethioninemia without homocystinuria. The hypermethioninemic phenotype associated with these mutations is inherited as an autosomal recessive trait. The only exception is the dominant mild hypermethioninemia associated with a G-A transition at nucleotide 791 of exon VII. This change yields a MAT1A-encoded subunit in which arginine 264 is replaced by histidine. Our results indicate that in the homologous rat enzyme, replacement of the equivalent arginine 265 by histidine (R265H) results in a monomeric MAT with only 0.37% of the AdoMet synthetic activity. However the tripolyphosphatase activity is similar to that found in the wild type (WT) MAT and is inhibited by PP(i). Our in vivo studies demonstrate that the R265H MAT I/III mutant associates with the WT subunit resulting in a dimeric R265H-WT MAT unable to synthesize AdoMet. Tripolyphosphatase activity is maintained in the hybrid MAT, but is not stimulated by methionine and ATP, indicating a deficient binding of the substrates. Our data indicate that the active site for tripolyphosphatase activity is functionally active in the monomeric R265H MAT I/III mutant. Moreover, our results provide a molecular mechanism that might explain the dominant inheritance of the hypermethioninemia associated with the R264H mutation of human MAT I/III. PMID- 11278457 TI - A regulatory hydrophobic area in the flexible joint region of plasminogen activator inhibitor-1, defined with fluorescent activity-neutralizing ligands. Ligand-induced serpin polymerization. AB - We have characterized the neutralization of the inhibitory activity of the serpin plasminogen activator inhibitor-1 (PAI-1) by a number of structurally distinct organochemicals, including compounds with environment-sensitive spectroscopic properties. In contrast to latent and reactive center-cleaved PAI-1 and PAI-1 in complex with urokinase-type plasminogen activator (uPA), active PAI-1 strongly increased the fluorescence of the PAI-1-neutralizing compounds 1 anilinonaphthalene-8-sulfonic acid and 4,4'-dianilino-1,1'-bisnaphthyl-5,5' disulfonic acid. The fluorescence increase could be competed by all tested nonfluorescent neutralizers, indicating that all neutralizers bind to a common hydrophobic area preferentially accessible in active PAI-1. Activity neutralization proceeded through two consecutive steps as follows: first step is conversion to forms displaying substrate behavior toward uPA, and second step is to forms inert to uPA. With some neutralizers, the second step was associated with PAI-1 polymerization. Vitronectin reduced the susceptibility to the neutralizers. Changes in sensitivity to activity neutralization by point mutations were compatible with the various neutralizers having overlapping, but not identical, binding sites in the region around alpha-helices D and E and beta strand 1A, known to act as a flexible joint when beta-sheet A opens and the reactive center loop inserts as beta-strand 4A during reaction with target proteinases. The defined binding area may be a target for development of compounds for neutralizing PAI-1 in cancer and cardiovascular diseases. PMID- 11278458 TI - Characterization of the nuclear import pathway for HIV-1 integrase. AB - The karyophilic properties of the human immunodeficiency virus, type I (HIV-1) pre-integration complex (PIC) allow the virus to infect non-dividing cells. To better understand the mechanisms responsible for nuclear translocation of the PIC, we investigated nuclear import of HIV-1 integrase (IN), a PIC-associated viral enzyme involved in the integration of the viral genome in the host cell DNA. Accumulation of HIV-1 IN into nuclei of digitonin-permeabilized cells does not result from passive diffusion but rather from an active transport that occurs through the nuclear pore complexes. HIV-1 IN is imported by a saturable mechanism, implying that a limiting cellular factor is responsible for this process. Although IN has been previously proposed to contain classical basic nuclear localization signals, we found that nuclear accumulation of IN does not involve karyopherins alpha, beta1, and beta2-mediated pathways. Neither the non hydrolyzable GTP analog, guanosine 5'-O-(thiotriphosphate), nor the GTP hydrolysis-deficient Ran mutant, RanQ69L, significantly affects nuclear import of IN, which depends instead on ATP hydrolysis. Therefore these results support the idea that IN import is not mediated by members of the karyopherin beta family. More generally, in vitro nuclear import of IN does not require addition of cytosolic factors, suggesting that cellular factor(s) involved in this active but atypical pathway process probably remain associated with the nuclear compartment or the nuclear pore complexes from permeabilized cells. PMID- 11278459 TI - Granzyme B-mediated apoptosis proceeds predominantly through a Bcl-2-inhibitable mitochondrial pathway. AB - Cytotoxic T lymphocytes kill virus-infected and tumor cell targets through the concerted action of proteins contained in cytolytic granules, primarily granzyme B and perforin. Granzyme B, a serine proteinase with substrate specificity similar to the caspase family of apoptotic cysteine proteinases, is capable of cleaving and activating a number of death proteins in target cells. Despite the ability to engage the death pathway at multiple entry points, the preferred mechanism for rapid induction of apoptosis by granzyme B has yet to be clearly established. Here we use time lapse confocal microscopy to demonstrate that mitochondrial cytochrome c release is the primary mode of granzyme B-induced apoptosis and that Bcl-2 is a potent inhibitor of this pivotal event. Caspase activation is not required for cytochrome c release, an activity that correlates with cleavage and activation of Bid, which we have found to be cleaved more readily by granzyme B than either caspase-3 or caspase-8. Bcl-2 blocks the rapid destruction of targets by granzyme B by blocking mitochondrial involvement in the process. PMID- 11278460 TI - The human sex-determining gene SRY is a direct target of WT1. AB - The product of the Wilms' tumor gene, WT1, is essential for male sex determination and differentiation in mammals. In addition to causing Wilms' tumor, mutations in WT1 often cause two distinct but overlapping urogenital defects in men, Denys-Drash syndrome and Frasier syndrome. In this study we investigated the regulation of the sex determination gene SRY by WT1. Our results showed that WT1 up-regulates the SRY gene through the proximal early growth response gene-1-like DNA-binding sequences in the core promoter. Mutant WT1 proteins in Denys-Drash syndrome patients were unable to activate this promoter. These mutants did not act in a dominant negative manner, as expected over the wild-type WT1 in this promoter. We also found that WT1 could transactivate the endogenous SRY gene. These observations, together with the overlapping expression patterns of WT1 and SRY in human gonads, led us to propose that WT1 regulates SRY in the initial sex determination process in humans and activates a cascade of genes ultimately leading to the complete organogenesis of the testis. PMID- 11278461 TI - Characterization of a hypoxia-inducible factor (HIF-1alpha ) from rainbow trout. Accumulation of protein occurs at normal venous oxygen tension. AB - The mammalian hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription factor that controls the induction of several genes involved in glycolysis, erythropoiesis, and angiogenesis when cells are exposed to hypoxic conditions. Until now, the expression and function of HIF-1alpha have not been studied in fish, which experience wide fluctuations of oxygen tensions in their natural environment. Using electrophoretic mobility shift assay, we have ascertained that a hypoxia-inducible factor is present in rainbow trout cells. We have also cloned the full-length cDNA (3605 base pairs) of the HIF-1alpha from rainbow trout with a predicted protein sequence of 766 amino acids that showed a 61% similarity to human and mouse HIF-1alpha. Polyclonal antibodies against the N-terminal part (amino acids 12-363) and the C-terminal part (amino acids 330-730) of rainbow trout HIF-1alpha protein recognized rainbow trout and chinook salmon HIF-1alpha protein in Western blot analysis. Also, the human and mouse HIF-1alpha proteins were recognized by the N-terminal rainbow trout anti-HIF-1alpha antibody but not by the C-terminal HIF-1alpha antibody. The accumulation of HIF-1alpha was studied by incubating rainbow trout and chinook salmon cells at different oxygen concentrations from 20 to 0.2% O(2) for 1 h. The greatest accumulation of HIF 1alpha protein occurred at 5% O(2) (38 torr), a typical oxygen tension of venous blood in normoxic animals. The protein stability experiments in the absence or presence of a proteasome inhibitor, MG-132, demonstrated that the inhibitor is able to stabilize the protein, which normally is degraded via the proteasome pathway both in normoxia and hypoxia. Notably, the hypoxia response element of oxygen-dependent degradation domain is identical in mammalian, Xenopus, and rainbow trout HIF-1alpha proteins, suggesting a high degree of evolutionary conservation in degradation of HIF-1alpha protein. PMID- 11278462 TI - Focal adhesion kinase activates Stat1 in integrin-mediated cell migration and adhesion. AB - Recent studies suggest that focal adhesion kinase (FAK) is important for cell migration. We now suggest a mechanism by which FAK activates the signal transducer and activator of transcription (STAT) pathway, regulating cell adhesion and migration. In particular, we observe that FAK is capable of activating Stat1, but not Stat3. Co-immunoprecipitation and in vitro binding assays demonstrate that Stat1 is transiently and directly associated with FAK during cell adhesion, and Stat1 is activated in this process. FAK with a C terminal deletion (FAKDeltaC14) completely abolishes this interaction, indicating this association is dependent on the C-terminal domain of FAK, which is required for FAK localization at focal contacts. Moreover, Stat1 activation during cell adhesion is diminished in FAK-deficient cells, correlating with decreased migration in these cells. Finally, we show that depletion of Stat1 results in an enhancement of cell adhesion and a decrease in cell migration. Thus, our results have demonstrated, for the first time, a critical signaling pathway from integrin/FAK to Stat1 that reduces cell adhesion and promotes cell migration. PMID- 11278463 TI - The human gC1qR/p32 gene, C1qBP. Genomic organization and promoter analysis. AB - gC1qR is an ubiquitously expressed cell protein that interacts with the globular heads of C1q (gC1q) and many other ligands. In this study, the 7.8-kilobase pair (kb) human gC1qR/p32 (C1qBP) gene was cloned and found to consist of 6 exons and 5 introns. Analysis of a 1.3-kb DNA fragment at the 5'-flanking region of this gene revealed the presence of multiple TATA, CCAAT, and Sp1 binding sites. Luciferase reporter assays performed in different human cell lines demonstrated that the reporter gene was ubiquitously driven by this 1.3-kb fragment. Subsequent 5' and 3' deletion of this fragment confined promoter elements to within 400 base pairs (bp) upstream of the translational start site. Because the removal of the 8-bp consensus TATATATA at -399 to -406 and CCAAT at -410 to -414 did not significantly affect the transcription efficiency of the promoter, GC rich sequences between this TATA box and the translation start site may be very important for the promoter activity of the C1qBP gene. One of seven GC-rich sequences in this region binds specifically to PANC-1 nuclear extracts, and the transcription factor Sp1 was shown to bind to this GC-rich sequence by the supershift assay. Primer extension analysis mapped three major transcription start regions. The farthest transcription start site is 49 bp upstream of the ATG translation initiation codon and is in close proximity of the specific SP1 binding site. PMID- 11278464 TI - Distinct signaling pathways for MCP-1-dependent integrin activation and chemotaxis. AB - Transmigration of monocytes to the subendothelial space is the initial step of atherosclerotic plaque formation and inflammation. Integrin activation and chemotaxis are two important functions involved in monocyte transmigration. To delineate the signaling cascades leading to integrin activation and chemotaxis by monocyte chemoattractant protein-1 (MCP-1), we have investigated the roles of MAPK and Rho GTPases in THP-1 cells, a monocytic cell line. MCP-1 stimulated beta1 integrin-dependent, but not beta2 integrin-dependent cell adhesion in a time-dependent manner. MCP-1-mediated cell adhesion was inhibited by a MEK inhibitor but not by a p38-MAPK inhibitor. In contrast, MCP-1-mediated chemotaxis was inhibited by the p38-MAPK inhibitor but not by the MEK inhibitor. The inhibitor of Rho GTPase, C3 exoenzyme, and a Rho kinase inhibitor abrogated MCP-1 dependent chemotaxis but not integrin-dependent cell adhesion. Further, C3 exoenzyme and the Rho kinase inhibitor blocked MCP-1-dependent p38-MAPK activation. These data indicate that ERK is responsible for integrin activation, that p38-MAPK and Rho are responsible for chemotaxis mediated by MCP-1, and that Rho and the Rho kinase are upstream of p38-MAPK in MCP-1-mediated signaling. This study demonstrates that two distinct MAPKs regulate two dependent signaling cascades leading to integrin activation and chemotaxis induced by MCP-1 in THP-1 cells. PMID- 11278466 TI - A pancreatic beta -cell-specific enhancer in the human PDX-1 gene is regulated by hepatocyte nuclear factor 3beta (HNF-3beta ), HNF-1alpha, and SPs transcription factors. AB - The PDX-1 transcription factor plays a key role in pancreas development. Although expressed in all cells at the early stages, in the adult it is mainly restricted to the beta-cell. To characterize the regulatory elements and potential transcription factors necessary for human PDX-1 gene expression in beta-cells, we constructed a series of 5' and 3' deletion fragments of the 5'-flanking region of the gene, fused to the luciferase reporter gene. In this report, we identify by transient transfections in beta- and non-beta-cells a novel beta-cell-specific distal enhancer element located between -3.7 and -3.45 kilobases. DNase I footprinting analysis revealed two protected regions, one binding the transcription factors SP1 and SP3 and the other hepatocyte nuclear factor 3beta (HNF-3beta) and HNF-1alpha. Cotransfection experiments suggest that HNF-3beta, HNF-1alpha, and SP1 are positive regulators of the herein-described human PDX-1 enhancer element. Furthermore, mutations within each motif abolished the binding of the corresponding factor(s) and dramatically impaired the enhancer activity, therefore suggesting cooperativity between these factors. PMID- 11278465 TI - The tyrosine kinase Hck is an inhibitor of HIV-1 replication counteracted by the viral vif protein. AB - The virus infectivity factor (Vif) protein facilitates the replication of human immunodeficiency virus type 1 (HIV-1) in primary lymphocytes and macrophages. Its action is strongly dependent on the cellular environment, and it has been proposed that the Vif protein counteracts cellular activities that would otherwise limit HIV-1 replication. Using a glutathione S-transferase pull-down assay, we identified that Vif binds specifically to the Src homology 3 domain of Hck, a tyrosine kinase from the Src family. The interaction between Vif and the full-length Hck was further assessed by co-precipitation assays in vitro and in human cells. The Vif protein repressed the kinase activity of Hck and was not itself a substrate for Hck phosphorylation. Within one single replication cycle of HIV-1, Hck was able to inhibit the production and the infectivity of vif deleted virus but not that of wild-type virus. Accordingly, HIV-1 vif- replication was delayed in Jurkat T cell clones stably expressing Hck. Our data demonstrate that Hck controls negatively HIV-1 replication and that this inhibition is suppressed by the expression of Vif. Hck, which is present in monocyte-macrophage cells, represents the first identified cellular inhibitor of HIV-1 replication overcome by Vif. PMID- 11278467 TI - Expression and function of the collagen receptor GPVI during megakaryocyte maturation. AB - In this report, the expression and function of the platelet collagen receptor glycoprotein VI (GPVI) were studied in human megakaryocytes during differentiation and maturation of mobilized blood and cord blood derived CD34(+) cells. By flow cytometry, using an anti-GPVI monoclonal antibody or convulxin, a GPVI-specific ligand, GPVI was detected only on CD41(+) cells including some CD41(+)/CD34(+) cells, suggesting expression at a stage of differentiation similar to CD41. These results were confirmed at the mRNA level using reverse transcription-polymerase chain reaction. GPVI expression was low during megakaryocytic differentiation but increased in the more mature megakaryocytes (CD41(high)). As in platelets, megakaryocyte GPVI associates with the Fc receptor gamma chain (FcRgamma). The FcR gamma chain was detected at the RNA and protein level at all stages of megakaryocyte maturation preceding the expression of GPVI. The other collagen receptor, alpha(2)beta(1) integrin (CD49b/CD29), had a pattern of expression similar to GPVI. Megakaryocytic GPVI was recognized as a 55-kDa protein by immunoblotting and ligand blotting, and thus it presented a slightly lower apparent molecular mass than platelet GPVI (58 kDa). Megakaryocytes began to adhere to immobilized convulxin via GPVI after only 8-10 days of culture, at a time when megakaryocytes were maturing. At this stage of maturation, they also adhered to immobilized collagen by alpha(2)beta(1) integrin-dependent and independent mechanisms. Convulxin induced a very similar pattern of protein tyrosine phosphorylation in megakaryocytes and platelets including Syk, FcRgamma, and PLC(gamma)2. Our results showed that GPVI is expressed early during megakaryocytic differentiation but functionally allows megakaryocyte adherence to collagen only at late stages of differentiation when its expression increases. PMID- 11278468 TI - Identification of tyrosine residues in vascular endothelial growth factor receptor-2/FLK-1 involved in activation of phosphatidylinositol 3-kinase and cell proliferation. AB - Activation of vascular endothelial growth factor receptor-2 (VEGFR-2) plays a critical role in vasculogenesis and angiogenesis. However, the mechanism by which VEGFR-2 activation elicits these cellular events is not fully understood. We recently constructed a chimeric receptor containing the extracellular domain of human CSF-1R/c-fms, fused with the entire transmembrane and cytoplasmic domains of murine VEGFR-2 (Rahimi, N., Dayanir, V., and Lashkari, K. (2000) J. Biol. Chem. 275, 16986-16992). In this study we used VEGFR-2 chimera (herein named CKR) to elucidate the signal transduction relay of VEGFR-2 in porcine aortic endothelial (PAE) cells. Mutation of tyrosines 799 and 1173 individually on CKR resulted in partial loss of CKR's ability to stimulate cell growth. Double mutation of these sites caused total loss of CKR's ability to stimulate cell growth. Interestingly, mutation of these sites had no effect on the ability of CKR to stimulate cell migration. Further analysis revealed that tyrosines 799 and 1173 are docking sites for p85 of phosphatidylinositol 3-kinase (PI3K). Pretreatment of cells with wortmannin, an inhibitor of PI3K, and rapamycin, a potent inhibitor of S6 kinase, abrogated CKR-mediated cell growth. However, expression of a dominant negative form of ras (N(17)ras) and inhibition of the mitogen-activated protein kinase (MAPK) pathway by PD98059 did not attenuate CKR stimulated cell growth. Altogether, these results demonstrate that activation of VEGFR-2 results in activation of PI3K and that activation of PI3K/S6kinase pathway, but not Ras/MAPK, is responsible for VEGFR-2-mediated cell growth. PMID- 11278469 TI - Regulation of membrane targeting of the G protein-coupled receptor kinase 2 by protein kinase A and its anchoring protein AKAP79. AB - The beta2 adrenergic receptor (beta2AR) undergoes desensitization by a process involving its phosphorylation by both protein kinase A (PKA) and G protein coupled receptor kinases (GRKs). The protein kinase A-anchoring protein AKAP79 influences beta2AR phosphorylation by complexing PKA with the receptor at the membrane. Here we show that AKAP79 also regulates the ability of GRK2 to phosphorylate agonist-occupied receptors. In human embryonic kidney 293 cells, overexpression of AKAP79 enhances agonist-induced phosphorylation of both the beta2AR and a mutant of the receptor that cannot be phosphorylated by PKA (beta2AR/PKA-). Mutants of AKAP79 that do not bind PKA or target to the beta2AR markedly inhibit phosphorylation of beta2AR/PKA-. We show that PKA directly phosphorylates GRK2 on serine 685. This modification increases Gbetagamma subunit binding to GRK2 and thus enhances the ability of the kinase to translocate to the membrane and phosphorylate the receptor. Abrogation of the phosphorylation of serine 685 on GRK2 by mutagenesis (S685A) or by expression of a dominant negative AKAP79 mutant reduces GRK2-mediated translocation to beta2AR and phosphorylation of agonist-occupied beta2AR, thus reducing subsequent receptor internalization. Agonist-stimulated PKA-mediated phosphorylation of GRK2 may represent a mechanism for enhancing receptor phosphorylation and desensitization. PMID- 11278470 TI - Protein kinase C alpha -mediated negative feedback regulation is responsible for the termination of insulin-like growth factor I-induced activation of nuclear phospholipase C beta1 in Swiss 3T3 cells. AB - Previous studies from several independent laboratories have demonstrated the existence of an autonomous phosphoinositide (PI) cycle within the nucleus, where it is involved in both cell proliferation and differentiation. Stimulation of Swiss 3T3 cells with insulin-like growth factor-I (IGF-I) has been shown to induce a transient and rapid increase in the activity of nuclear-localized phospholipase C (PLC) beta1, which in turn leads to the production of inositol trisphosphate and diacylglycerol in the nucleus. Nuclear diacylglycerol provides the driving force for the nuclear translocation of protein kinase C (PKC) alpha. Here, we report that treatment of Swiss 3T3 cells with Go6976, a selective inhibitor of PKC alpha, caused a sustained elevation of IGF-I-stimulated nuclear PLC activity. A time course study revealed an inverse relationship between nuclear PKC activity and the activity of nuclear PLC in IGF-I-treated cells. A time-dependent association between PKC alpha and PLC beta1 in the nucleus was also observed following IGF-I treatment. Two-dimensional phosphopeptide mapping and site-directed mutagenesis demonstrated that PKC promoted phosphorylation of PLC beta1 at serine 887 in the nucleus of IGF-I-treated cells. Overexpression of either a PLC beta1 mutant in which the PKC phosphorylation site Ser(887) was replaced by alanine, or a dominant-negative PKC alpha, resulted in a sustained activation of nuclear PLC following IGF-I stimulation. These results indicate that a negative feedback regulation of PLC beta1 by PKC alpha plays a critical role in the termination of the IGF-I-dependent signal that activates the nuclear PI cycle. PMID- 11278471 TI - Tumor necrosis factor-alpha induction of endothelial ephrin A1 expression is mediated by a p38 MAPK- and SAPK/JNK-dependent but nuclear factor-kappa B independent mechanism. AB - Tumor necrosis factor-alpha (TNF-alpha) is a multifunctional cytokine that induces a broad spectrum of responses including angiogenesis. Angiogenesis promoted by TNF-alpha is mediated, at least in part, by ephrin A1, a member of the ligand family for Eph receptor tyrosine kinases. Although TNF-alpha induces ephrin A1 expression in endothelial cells, the signaling pathways mediating ephrin A1 induction remain unknown. In this study, we investigated the signaling mechanisms of TNF-alpha-dependent induction of ephrin A1 in endothelial cells. Both TNFR1 and TNFR2 appear to be involved in regulating ephrin A1 expression in endothelial cells, because neutralizing antibodies to either TNFR1 or TNFR2 inhibited TNF-alpha-induced ephrin A1 expression. Inhibition of nuclear factor kappaB (NF-kappaB) activation by a trans-dominant inhibitory isoform of mutant IkappaBalpha did not affect ephrin A1 induction, suggesting that NF-kappaB proteins are not major regulators of ephrin A1 expression. In contrast, ephrin A1 induction was blocked by inhibition of p38 mitogen-activated protein kinase (MAPK) or SAPK/JNK, but not p42/44 MAPK, using either selective chemical inhibitors or dominant-negative forms of p38 MAPK or TNF receptor-associated factor 2. These findings indicate that TNF-alpha-induced ephrin A1 expression is mediated through JNK and p38 MAPK signaling pathways. Taken together, the results of our study demonstrated that induction of ephrin A1 in endothelial cells by TNF alpha is mediated through both p38 MAPK and SAPK/JNK, but not p42/44 MAPK or NF kappaB, pathways. PMID- 11278472 TI - 3-Hydroxy-3-methylglutaryl-CoA reductase inhibitors block calcium-dependent tyrosine kinase Pyk2 activation by angiotensin II in vascular endothelial cells. involvement of geranylgeranylation of small G protein Rap1. AB - We recently reported the calcium-dependent activation of tyrosine kinase Pyk2 by angiotensin II (Ang II) in pulmonary vein endothelial cells (PVEC). Since Pyk2 has no calcium binding domain, and neither Ca(2+) nor Ca(2+)/calmodulin directly activates Pyk2, it is not clear how Ca(2+) transduces the signal to activate Pyk2, a key tyrosine kinase, in the early events of Ang II signaling. In the present study, we investigated the mechanism of the calcium-dependent activation of Pyk2 in response to Ang II by using 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors and isoprenoid intermediates in PVEC. We have obtained substantial evidence indicating that Ang II activates Pyk2 through calcium-mediated activation of the geranylgeranylated small G protein Rap1 and the Rap1 association with Pyk2. Thus, the small G protein Rap1 is an intermediary signaling molecule linking Ang II-induced calcium signal to Pyk2 activation in PVEC. In addition, our results indicate that 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors, a class of cholesterol-lowering drugs, could interrupt Ang II signaling independent of cholesterol lowering in endothelial cells. PMID- 11278473 TI - Functional analysis of type 1alpha cGMP-dependent protein kinase using green fluorescent fusion proteins. AB - The cGMP-dependent protein kinases (PKGs) are ubiquitous effector enzymes that regulate a variety of physiological processes in response to nitric oxide and natriuretic agonists. We have constructed green fluorescent fusion proteins (GFP) using full-length (PKG-GFP) and truncations encoding either the regulatory domain of PKG1alpha (G1alphaR-GFP) or the catalytic domains of PKG1alpha (GFP-G1C) to examine the enzymatic properties and intracellular location. When transiently transfected into mammalian cells, these constructs were detected on Western blots at the expected sizes using anti-GFP antibodies. The GFP-G1C and the full-length PKG1alpha-GFP fusion proteins were found to have constitutive activity both in vivo and in vitro. The G1alphaR-GFP protein was found to dimerize with endogenous type 1 PKG and behaved in a dominant negative manner both in vivo and in vitro. When expressed transiently in either HEK-293 or A549 epithelial cells, the fusion proteins encoding the amino-terminal regulatory domains (PKG-GFP, G1alphaR-GFP) were present in the cytosol and were rarely observed in the nucleus. In contrast, the GFP-G1C (lacking regulatory domains) concentrated in the nucleus. Of the fusion proteins containing the regulatory region, the constitutive PKG-GFP protein was present in a more centralized location, whereas the G1alphaR-GFP protein colocalized with F-actin on stress fibers and in dynamic regions of the plasma membrane. Microscopic and immunoprecipitation studies indicated that both the G1alphaR-GFP and the PKG-GFP fusion proteins colocalized with vasodilator stimulated phosphoprotein (VASP). These constructs thus represent novel tools with which to visualize inactive, and activated, PKG1alpha in vivo, and we have used them to demonstrate two functionally independent domains. In addition, we show for the first time in living cells that PKG is found in dynamic membrane regions in association with VASP. PMID- 11278474 TI - The glial and the neuronal glycine transporters differ in their reactivity to sulfhydryl reagents. AB - The neuronal (GlyT2) and glial (GlyT1) glycine transporters, two members of the Na(+)/Cl(-)-dependent neurotransmitter transporter superfamily, differ by many aspects, such as substrate specificity and Na(+) coupling. We have characterized under voltage clamp their reactivity toward the membrane impermeant sulfhydryl reagent [2-(trimethylammonium)-ethyl]-methanethiosulfonate (MTSET). In Xenopus oocytes expressing GlyT1b, application of MTSET reduced to the same extent the Na(+)-dependent charge movement, the glycine-evoked current, and the glycine uptake, indicating a complete inactivation of the transporters following cysteine modification. In contrast, this compound had no detectable effect on the glycine uptake and the glycine-evoked current of GlyT2a. The sensitivities to MTSET of the two transporters can be permutated by suppressing a cysteine (C62A) in the first extracellular loop (EL1) of GlyT1b and introducing one at the equivalent position in GlyT2a, either by point mutation (A223C) or by swapping the EL1 sequence (GlyT1b-EL1 and GlyT2a-EL1) resulting in AFQ <--> CYR modification. Inactivation by MTSET was five times faster in GlyT2a-A223C than in GlyT2a-EL1 or GlyT1b, suggesting that the arginine in position +2 reduced the cysteine reactivity. Protection assays indicate that EL1 cysteines are less accessible in the presence of all co-transported substrates: Na(+), Cl(-), and glycine. Application of dithioerythritol reverses the inactivation by MTSET of the sensitive transporters. Together, these results indicate that EL1 conformation differs between GlyT1b and GlyT2a and is modified by substrate binding and translocation. PMID- 11278475 TI - Factor VIIa modified in the 170 loop shows enhanced catalytic activity but does not change the zymogen-like property. AB - Factor VIIa (VIIa) is an unusual trypsin-type serine proteinase that appears to exist in an equilibrium between minor active and dominant zymogen-like inactive conformational states. The binding of tissue factor to VIIa is assumed to shift the equilibrium into the active state. The proteinase domain of VIIa contains a unique structure: a loop formed by a disulfide bond between Cys310 and Cys329, which is five residues longer than those of other trypsin types. To examine the functional role of the loop region, we prepared two mutants of VIIa. One of the mutants, named VII-11, had five extra corresponding residues 316-320 of VII deleted. The other mutant, VII-31, had all of the residues in its loop replaced with those of trypsin. Functional analysis of the two mutants showed that VIIa-11 (Kd = 41 nm) and VIIa-31 (Kd = 160 nm) had lower affinities for soluble tissue factor as compared with the wild-type VIIa (Kd = 11 nm). The magnitude of tissue factor-mediated acceleration of amidolytic activities of VIIa-11 (7-fold) and that of VIIa-31 (2-fold) were also smaller than that of wild-type VIIa (30-fold). In the absence of tissue factor, VIIa-31 but not VIIa-11 showed enhanced activity; the catalytic efficiencies of VIIa-31 toward various chromogenic substrates were 2-18-fold greater than those of the wild-type VIIa. Susceptibility of the alpha-amino group of Ile-153 of VIIa-31 to carbamylation was almost the same as that of wild-type VIIa, suggesting that VIIa-31 as well as wild-type VIIa exist predominantly in the zymogen-like state. Therefore, the tested modifications in the loop region had adverse effects on affinity for tissue factor, disturbed the tissue factor-induced conformational transition, and changed the catalytic efficiency of VIIa, but they did not affect the equilibrium between active and zymogen-like conformational states. PMID- 11278476 TI - Heterologous inhibition of G protein-coupled receptor endocytosis mediated by receptor-specific trafficking of beta-arrestins. AB - We have observed an unexpected type of nonreciprocal "cross-regulation" of the agonist-induced endocytosis of G protein-coupled receptors by clathrin-coated pits. Isoproterenol-dependent internalization of beta2-adrenergic receptors in stably transfected HEK293 cells was specifically blocked (>65% inhibition) by vasopressin-induced activation of V2 vasopressin receptors co-expressed at similar levels. In contrast, activation of beta2 receptors caused no detectable effect on V2 receptor internalization in the same cells. Several pieces of evidence suggest that this nonreciprocal inhibition of endocytosis is mediated by receptor-specific intracellular trafficking of beta-arrestins. First, previous studies showed that the activation of V2 but not beta2 receptors caused pronounced recruitment of beta-arrestins to endocytic membranes (Oakley, R. H., Laporte, S. A., Holt, J. A., Barak, L. S., and Caron, M. G. (1999) J. Biol. Chem. 274, 32248-32257). Second, overexpression of arrestin 2 or 3 (beta-arrestin 1 or 2) abolished the V2 receptor-mediated inhibition of beta2 receptor internalization. Third, mutations of the V2 receptor that block endomembrane recruitment of beta-arrestins eliminated the V2 receptor-dependent blockade of beta2 receptor internalization. These results identify a novel type of heterologous regulation of G protein-coupled receptors, define a new functional role of receptor-specific intracellular trafficking of beta-arrestins, and suggest an experimental method to rapidly modulate the functional activity of beta-arrestins in intact cells. PMID- 11278477 TI - Cytosine methylation enhances mitoxantrone-DNA adduct formation at CpG dinucleotides. AB - Recently, we have shown that mitoxantrone can be activated by formaldehyde in vitro to form DNA adducts that are specific for CpG and CpA sites in DNA. The CpG specificity of adduct formation prompted investigations into the effect of cytosine methylation (CpG) on adduct formation, since the majority of CpG dinucleotides in the mammalian genome are methylated and hypermethylation in subsets of genes is associated with various neoplasms. Upon methylation of a 512 base pair DNA fragment (containing the lac UV5 promoter) using HpaII methylase, three CCGG sites downstream of the promoter were methylated at C5 of the internal cytosine residue. In vitro transcription studies of mitoxantrone-reacted DNA revealed a 3-fold enhancement in transcriptional blockage (and hence adduct formation) exclusively at these methylated sites. In vitro cross-linking assays also revealed that methylation enhanced mitoxantrone adduct formation by 2-3 fold, and methylation of cytosine at a single potential drug binding site on a duplex oligonucleotide also enhanced adduct levels by 3-fold. Collectively, these results indicate preferential adduct formation at methylated CpG sites. However, adducts at these methylated sites exhibited the same stability as nonmethylated sites, suggesting that cytosine methylation increases drug accessibility to DNA rather than being involved in kinetic stabilization of the adduct. PMID- 11278478 TI - Cyclic nucleotides modulate store-mediated calcium entry through the activation of protein-tyrosine phosphatases and altered actin polymerization in human platelets. AB - Agonists elevate the cytosolic calcium concentration in human platelets via a receptor-operated mechanism, involving both Ca(2+) release from intracellular stores and subsequent Ca(2+) entry, which can be inhibited by platelet inhibitors, such as prostaglandin E(1) and nitroprusside which elevate cAMP and cGMP, respectively. In the present study we investigated the mechanisms by which cAMP and cGMP modulate store-mediated Ca(2+) entry. Both prostaglandin E(1) and sodium nitroprusside inhibited thapsigargin-evoked store-mediated Ca(2+) entry and actin polymerization. However, addition of these agents after induction of store-mediated Ca(2+) entry did not affect either Ca(2+) entry or actin polymerization. Furthermore, prostaglandin E(1) and sodium nitroprusside dramatically inhibited the tyrosine phosphorylation induced by depletion of the internal Ca(2+) stores or agonist stimulation without affecting the activation of Ras or the Ras-activated phosphatidylinositol 3-kinase or extracellular signal related kinase (ERK) pathways. Inhibition of cyclic nucleotide-dependent protein kinases prevented inhibition of agonist-evoked Ca(2+) release but it did not have any effect on the inhibition of Ca(2+) entry or actin polymerization. Phenylarsine oxide and vanadate, inhibitors of protein-tyrosine phosphatases prevented the inhibitory effects of the cGMP and cAMP elevating agents on Ca(2+) entry and actin polymerization. These results suggest that Ca(2+) entry in human platelets is directly down-regulated by cGMP and cAMP by a mechanism involving the inhibition of cytoskeletal reorganization via the activation of protein tyrosine phosphatases. PMID- 11278479 TI - Role of the ERK pathway in the activation of store-mediated calcium entry in human platelets. AB - Extracellular signal-regulated kinases (ERKs), are common participants in a broad variety of signal transduction pathways. Several studies have demonstrated the presence of ERKs in human platelets and their activation by the physiological agonist thrombin. Here we report the involvement of the ERK cascade in store mediated Ca(2+) entry in human platelets. Treatment of dimethyl-bis-(o aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid-loaded platelets with thapsigargin to deplete the intracellular Ca(2+) stores resulted in a time- and concentration-dependent activation of ERK1 and ERK2. Incubation with either U0126 or PD 184352, specific inhibitors of mitogen-activated protein kinase kinase (MEK), prevented thapsigargin-induced ERK activation. Furthermore, U0126 and PD 184352 reduced Ca(2+) entry stimulated by thapsigargin or thrombin, in a concentration-dependent manner. The role of ERK in store-mediated Ca(2+) entry was found to be independent of phosphatidylinositol 3- and 4-kinases, the tyrosine kinase pathway, and actin polymerization but sensitive to treatment with inhibitors of Ras, suggesting that the ERK pathway might be a downstream effector of Ras in mediating store-mediated Ca(2+) entry in human platelets. In addition, we have found that store depletion stimulated ERK activation does not require PKC activity. This study demonstrates for the first time a novel mechanism for regulation of store-mediated Ca(2+) entry in human platelets involving the ERK cascade. PMID- 11278480 TI - Heat shock factor-4 (HSF-4a) represses basal transcription through interaction with TFIIF. AB - The heat shock transcription factors (HSFs) regulate the expression of heat shock proteins (hsps), which are critical for normal cellular proliferation and differentiation. One of the HSFs, HSF-4, contains two alternative splice variants, one of which possesses transcriptional repressor properties in vivo. This repressor isoform inhibits basal transcription of hsps 27 and 90 in tissue culture cells. The molecular mechanisms of HSF-4a isoform-mediated transcriptional repression is unknown. Here, we present evidence that HSF-4a inhibits basal transcription in vivo when it is artificially targeted to basal promoters via the DNA-binding domain of the yeast transcription factor, GAL4. By using a highly purified, reconstituted in vitro transcription system, we show that HSF-4a represses basal transcription at an early step during preinitiation complex assembly, as pre-assembled preinitiation complexes are refractory to the inhibitory effect on transcription. This repression occurs by the HSF-4a isoform, but not by the HSF-4b isoform, which we show is capable of activating transcription from a heat shock element-driven promoter in vitro. The repression of basal transcription by HSF-4a occurs through interaction with the basal transcription factor TFIIF. TFIIF interacts with a segment of HSF-4a that is required for the trimerization of HSF-4a, and deletion of this segment no longer inhibits basal transcription. These studies suggest that HSF-4a inhibits basal transcription both in vivo and in vitro. Furthermore, this is the first report identifying an interaction between a transcriptional repressor with the basal transcription factor TFIIF. PMID- 11278481 TI - Pdx-1 is required for activation in vivo from a duodenum-specific enhancer. AB - The purine metabolic gene adenosine deaminase (ADA) is expressed along a defined spatiotemporal pattern in the developing mammalian small intestine, where high level expression is limited to the villous epithelium of the duodenum. This activation is observed in rodents as the intestine completes the final maturation resulting in adult crypt-villus structures at 2-3 weeks postpartum. A regulatory module responsible for this pattern of expression has been identified in the second intron of the human ADA gene. Of the multiple duodenal proteins that can interact with this small duodenal enhancer region, the studies contained in this work describe the identification of five of these proteins as the dispersed homeobox protein PDX-1. This transcription factor exhibits a profile of expression in the small intestine similar to that observed for ADA, making it an ideal candidate factor for the duodenum-specific ADA enhancer. Loss of PDX-1 binding, via a PDX-1 mutated enhancer transgenic construction, resulted in complete loss of high-level activation in the duodenum, demonstrating the absolute requirement for this factor in vivo. However, co-transfection experiments suggest that other proteins that bind the enhancer are also required for enhancer function because PDX-1 alone was incapable of significant transactivation. PMID- 11278482 TI - Modification of alternative splicing of Bcl-x pre-mRNA in prostate and breast cancer cells. analysis of apoptosis and cell death. AB - There is ample evidence that deregulation of apoptosis results in the development, progression, and/or maintenance of cancer. Since many apoptotic regulatory genes (e.g. bcl-x) code for alternatively spliced protein variants with opposing functions, the manipulation of alternative splicing presents a unique way of regulating the apoptotic response. Here we have targeted oligonucleotides antisense to the 5'-splice site of bcl-x(L), an anti-apoptotic gene that is overexpressed in various cancers, and shifted the splicing pattern of Bcl-x pre-mRNA from Bcl-x(L) to Bcl-x(S), a pro-apoptotic splice variant. This approach induced significant apoptosis in PC-3 prostate cancer cells. In contrast, the same oligonucleotide treatment elicited a much weaker apoptotic response in MCF-7 breast cancer cells. Moreover, although the shift in Bcl-x pre mRNA splicing inhibited colony formation in both cell lines, this effect was much less pronounced in MCF-7 cells. These differences in responses to oligonucleotide treatment were analyzed in the context of expression of Bcl-x(L), Bcl-x(S), and Bcl-2 proteins. The results indicate that despite the presence of Bcl-x pre-mRNA in a number of cell types, the effects of modification of its splicing by antisense oligonucleotides vary depending on the expression profile of the treated cells. PMID- 11278483 TI - The mechanism of a bacterial plasminogen activator intermediate between streptokinase and staphylokinase. AB - The therapeutic properties of plasminogen activators are dictated by their mechanism of action. Unlike staphylokinase, a single domain protein, streptokinase, a 3-domain (alpha, beta, and gamma) molecule, nonproteolytically activates human (h)-plasminogen and protects plasmin from inactivation by alpha(2)-antiplasmin. Because a streptokinase-like mechanism was hypothesized to require the streptokinase gamma-domain, we examined the mechanism of action of a novel two-domain (alpha,beta) Streptococcus uberis plasminogen activator (SUPA). Under conditions that quench trace plasmin, SUPA nonproteolytically generated an active site in bovine (b)-plasminogen. SUPA also competitively inhibited the inactivation of plasmin by alpha(2)-antiplasmin. Still, the lag phase in active site generation and plasminogen activation by SUPA was at least 5-fold longer than that of streptokinase. Recombinant streptokinase gamma-domain bound to the b plasminogen.SUPA complex and significantly reduced these lag phases. The SUPA b.plasmin complex activated b-plasminogen with kinetic parameters comparable to those of streptokinase for h-plasminogen. The SUPA-b.plasmin complex also activated h-plasminogen but with a lower k(cat) (25-fold) and k(cat)/K(m) (7.9 fold) than SK. We conclude that a gamma-domain is not required for a streptokinase-like activation of b-plasminogen. However, the streptokinase gamma domain enhances the rates of active site formation in b-plasminogen and this enhancing effect may be required for efficient activation of plasminogen from other species. PMID- 11278484 TI - Casein kinase II sites in the intracellular C-terminal domain of the thyrotropin releasing hormone receptor and chimeric gonadotropin-releasing hormone receptors contribute to beta-arrestin-dependent internalization. AB - We have previously shown that the mammalian gonadotropin-releasing hormone receptor (GnRHR), a unique G-protein-coupled receptor (GPCR) lacking an intracellular carboxyl tail (C-tail), does not follow a beta-arrestin-dependent internalization pathway. However, internalization of a chimeric GnRHR with the thyrotropin-releasing hormone receptor (TRHR) C-tail does utilize beta-arrestin. Here, we have investigated the sites within the intracellular C-tail domain that are important for conferring beta-arrestin-dependent internalization. In contrast to the chimeric GnRHR with a TRHR C-tail, a chimeric GnRHR with the catfish GnRHR C-tail is not beta-arrestin-dependent. Sequence comparisons between these chimeric receptors show three consensus phosphorylation sites for casein kinase II (CKII) in the TRHR C-tail but none in the catfish GnRHR C-tail. We thus investigated a role for CKII sites in determining GPCR internalization via beta arrestin. Sequential introduction of three CKII sites into the chimera with the catfish C-tail (H354D,A366E,G371D) resulted in a change in the pattern of receptor phosphorylation and beta-arrestin-dependence, which only occurred when all three sites were introduced. Conversely, mutation of the putative CKII sites (T365A,T371A,S383A) in the C-tail of a beta-arrestin-sensitive GPCR, the TRHR, resulted in decreased receptor phosphorylation and a loss of beta-arrestin dependence. Mutation of all three CKII sites was necessary before a loss of beta arrestin-dependence was observed. Visualization of beta-arrestin/GFP redistribution confirmed a loss or gain of beta-arrestin sensitivity for receptor mutants. Internalization of receptors without C-tail CKII sites was promoted by a phosphorylation-independent beta-arrestin mutant (R169E), suggesting that these receptors do not contain the necessary phosphorylation sites required for beta arrestin-dependent internalization. Apigenin, a specific CKII inhibitor, blocked the increase in receptor internalization by beta-arrestin, thus providing further support for the involvement of CKII. This study presents evidence of a novel role for C-tail CKII consensus sites in targeting these GPCRs to the beta-arrestin dependent pathway. PMID- 11278486 TI - The mechanism of PAK activation. Autophosphorylation events in both regulatory and kinase domains control activity. AB - The p21-activated kinases (PAKs), in common with many kinases, undergo multiple autophosphorylation events upon interaction with appropriate activators. The Cdc42-induced phosphorylation of PAK serves in part to dissociate the kinase from its partners PIX and Nck. Here we investigate in detail how autophosphorylation events affect the catalytic activity of PAK by altering the autophosphorylation sites in both alpha- and betaPAK. Both in vivo and in vitro analyses demonstrate that, although most phosphorylation events in the PAK N-terminal regulatory domain play no direct role in activation, a phosphorylation of alphaPAK serine 144 or betaPAK serine 139, which lie in the kinase inhibitory domain, significantly contribute to activation. By contrast, sphingosine-mediated activation is independent of this residue, indicating a different mode of activation. Thus two autophosphorylation sites direct activation while three others control association with focal complexes via PIX and Nck. PMID- 11278485 TI - Phosphorylation-independent association of CXCR2 with the protein phosphatase 2A core enzyme. AB - Protein phosphatase 2A (PP2A) is postulated to be involved in the dephosphorylation of G protein-coupled receptors. In the present study, we demonstrate that the carboxyl terminus of CXCR2 physically interacts with the PP2A core enzyme, a dimer formed by PP2Ac and PR65, but not with the PP2Ac monomer, suggesting direct interaction of the receptor with PR65. The integrity of a sequence motif in the C terminus of CXCR2, KFRHGL, which is conserved in all CC and CXC chemokine receptors, is required for the receptor binding to the PP2A core enzyme. CXCR2 co-immunoprecipitates with the PP2A core enzyme in HEK293 cells and in human neutrophils. Overexpression of dominant negative dynamin 1 (dynamin 1 K44A) in CXCR2-expressing cells blocks the receptor association with the PP2A core enzyme, and an internalization-deficient mutant form of CXCR2 (I323A,L324A) also exhibits impaired association with the PP2A core enzyme, suggesting that the receptor internalization is required for the receptor binding to PP2A. A phosphorylation-deficient mutant of CXCR2 (331T), which has previously been shown to undergo internalization in HEK293 cells, binds to an almost equal amount of the PP2A core enzyme in comparison with the wild-type CXCR2, suggesting that the interaction of the receptor with PP2A is phosphorylation-independent. The dephosphorylation of CXCR2 is reversed by treatment of the cells with okadaic acid. Moreover, pretreatment of the cells with okadaic acid increases basal phosphorylation of CXCR2 and attenuates CXCR2-mediated calcium mobilization and chemotaxis. Taken together, these data indicate that PP2A is involved in the dephosphorylation of CXCR2. We postulate that this interaction results from direct binding of the regulatory subunit A (PR65) of PP2A to the carboxyl terminus of CXCR2 after receptor sequestration and internalization. PMID- 11278487 TI - An essential role of Glu-243 and His-239 in the phosphotransfer reaction catalyzed by pyruvate dehydrogenase kinase. AB - This study was undertaken to examine the mechanistic significance of two highly conserved residues positioned in the active site of pyruvate dehydrogenase kinase, Glu-243 and His-239. We used site-directed mutagenesis to convert Glu-243 to Ala, Asp, or Gln and His-239 to Ala. The resulting mutant kinases demonstrated a greatly reduced capacity for phosphorylation of pyruvate dehydrogenase. The Glu 243 to Asp mutant had approximately 2% residual activity, whereas the Glu-243 to Ala or Gln mutants exhibited less than 0.5 and 0.1% residual activity, respectively. Activity of the His-239 to Ala mutant was decreased by approximately 90%. Active-site titration with [alpha-(32)P]ATP revealed that neither Glu-243 nor His-239 mutations affected nucleotide binding. All mutant kinases showed similar or even somewhat greater affinity than the wild-type kinase toward the protein substrate, pyruvate dehydrogenase complex. Furthermore, neither of the mutations affected the inter-subunit interactions. Finally, pyruvate dehydrogenase kinase was found to possess a weak ATP hydrolytic activity, which required Glu-243 and His-239 similar to the kinase activity. Based on these observations, we propose a mechanism according to which the invariant glutamate residue (Glu-243) acts as a general base catalyst, which activates the hydroxyl group on a serine residue of the protein substrate for direct attack on the gamma phosphate. The glutamate residue in turn might be further polarized through interaction with the neighboring histidine residue (His 239). PMID- 11278488 TI - The v-Src SH3 domain facilitates a cell adhesion-independent association with focal adhesion kinase. AB - Integrins facilitate cell attachment to the extracellular matrix, and these interactions generate cell survival, proliferation, and motility signals. Integrin signals are relayed in part by focal adhesion kinase (FAK) activation and the formation of a transient signaling complex initiated by Src homology 2 (SH2)-dependent binding of Src family protein-tyrosine kinases to the FAK Tyr-397 autophosphorylation site. Here we show that in viral Src (v-Src)-transformed NIH3T3 fibroblasts, an adhesion-independent FAK-Src signaling complex occurs. Co expression studies in human 293T cells showed that v-Src could associate with and phosphorylate a Phe-397 FAK mutant at Tyr-925 promoting Grb2 binding to FAK in suspended cells. In vitro, glutathione S-transferase fusion proteins of the v-Src SH3 but not c-Src SH3 domain bound to FAK in lysates of NIH3T3 fibroblasts. The v Src SH3-binding sites were mapped to known proline-X-X-proline (PXXP) SH3-binding motifs in the FAK N- (residues 371-377) and C-terminal domains (residues 712-718 and 871-882) by in vitro pull-down assays, and these sites are composed of a PXXPXXPhi (where Phi is a hydrophobic residue) v-Src SH3 binding consensus. Sequence comparisons show that residues in the RT loop region of the c-Src and v Src SH3 domains differ. Substitution of c-Src RT loop residues (Arg-97 and Thr 98) for those found in the v-Src SH3 domain (Trp-97 and Ile-98) enhanced the binding of distinct NIH3T3 cellular proteins to a glutathione S-transferase fusion protein of the c-Src (Trp-97 + Ile-98) SH3 domain. FAK was identified as a c-Src (Trp-97 + Ile-98) SH3 domain target in fibroblasts, and co-expression studies in 293T cells showed that full-length c-Src (Trp-97 + Ile-98) could associate in vivo with Phe-397 FAK in an SH2-independent manner. These studies establish a functional role for the v-Src SH3 domain in stabilizing an adhesion independent signaling complex with FAK. PMID- 11278489 TI - Biochemical characterization of the reverse activity of rat brain ceramidase. A CoA-independent and fumonisin B1-insensitive ceramide synthase. AB - We have previously purified a membrane-bound ceramidase from rat brain and recently cloned the human homologue. We also observed that the same enzyme is able to catalyze the reverse reaction of ceramide synthesis. To obtain insight into the biochemistry of this enzyme, we characterized in this study this reverse activity. Using sphingosine and palmitic acid as substrates, the enzyme exhibited Michaelis-Menten kinetics; however, the enzyme did not utilize palmitoyl-CoA as substrate. Also, the activity was not inhibited in vitro and in cells by fumonisin B1, an inhibitor of the CoA-dependent ceramide synthase. The enzyme showed a narrow pH optimum in the neutral range, and there was very low activity in the alkaline range. Substrate specificity studies were performed, and the enzyme showed the highest activity with d-erythro-sphingosine (Km of 0.16 mol %, and Vmax of 0.3 micromol/min/mg), but d-erythro-dihydrosphingosine and the three unnatural stereoisomers of sphingosine were poor substrates. The specificity for the fatty acid was also studied, and the highest activity was observed for myristic acid with a Km of 1.7 mol % and a Vmax of 0.63 micromol/min/mg. Kinetic studies were performed to investigate the mechanism of the reaction, and Lineweaver-Burk plots indicated a sequential mechanism. Two competitive inhibitors of the two substrates were identified, l-erythro-sphingosine and myristaldehyde, and inhibition studies indicated that the reaction followed a random sequential mechanism. The effect of lipids were also tested. Most of these lipids showed moderate inhibition, whereas the effects of phosphatidic acid and cardiolipin were more potent with total inhibition at around 2.5-5 mol %. Paradoxically, cardiolipin stimulated ceramidase activity. These results define the biochemical characteristics of this reverse activity. The results are discussed in view of a possible regulation of this enzyme by the intracellular pH or by an interaction with cardiolipin and/or phosphatidic acid. PMID- 11278490 TI - The radioresistance to killing of A1-5 cells derives from activation of the Chk1 pathway. AB - Checkpoints respond to DNA damage by arresting the cell cycle to provide time for facilitating repair. In mammalian cells, the G(2) checkpoint prevents the Cdc25C phosphatase from removing inhibitory phosphate groups from the mitosis-promoting kinase Cdc2. Both Chk1 and Chk2, the checkpoint kinases, can phosphorylate Cdc25C and inactivate its in vitro phosphatase activity. Therefore, both Chk1 and Chk2 are thought to regulate the activation of the G(2) checkpoint. Here we report that A1-5, a transformed rat embryo fibroblast cell line, shows much more radioresistance associated with a much stronger G(2) arrest response when compared with its counterpart, B4, although A1-5 and B4 cells have a similar capacity for nonhomologous end-joining DNA repair. These phenotypes of A1-5 cells are accompanied by a higher Chk1 expression and a higher phosphorylation of Cdc2. On the other hand, Chk2 expression increases slightly following radiation; however, it has no difference between A1-5 and B4 cells. Caffeine or UCN-01 abolishes the extreme radioresistance with the strong G(2) arrest and at the same time reduces the phosphorylation of Cdc2 in A1-5 cells. In addition, Chk1 but not Chk2 antisense oligonucleotide sensitizes A1-5 cells to radiation-induced killing and reduces the G(2) arrest of the cells. Taken together these results suggest that the Chk1/Cdc25C/Cdc2 pathway is the major player for the radioresistance with G(2) arrest in A1-5 cells. PMID- 11278491 TI - Structural and functional analysis of missense mutations in fumarylacetoacetate hydrolase, the gene deficient in hereditary tyrosinemia type 1. AB - Hereditary tyrosinemia type 1 (HT1) is an autosomal recessive disease caused by a deficiency of the enzyme involved in the last step of tyrosine degradation, fumarylacetoacetate hydrolase (FAH). Thus far, 34 mutations in the FAH gene have been reported in various HT1 patients. Site-directed mutagenesis of the FAH cDNA was used to investigate the effects of eight missense mutations found in HTI patients on the structure and activity of FAH. Mutated FAH proteins were expressed in Escherichia coli and in mammalian CV-1 cells. Mutations N16I, F62C, A134D, C193R, D233V, and W234G lead to enzymatically inactive FAH proteins. Two mutations (R341W, associated with the pseudo-deficiency phenotype, and Q279R) produced proteins with a level of activity comparable to the wild-type enzyme. The N16I, F62C, C193R, and W234G variants were enriched in an insoluble cellular fraction, suggesting that these amino acid substitutions interfere with the proper folding of the enzyme. Based on the tertiary structure of FAH, on circular dichroism data, and on solubility measurements, we propose that the studied missense mutations cause three types of structural effects on the enzyme: 1) gross structural perturbations, 2) limited conformational changes in the active site, and 3) conformational modifications with no significant effect on enzymatic activity. PMID- 11278492 TI - Characterization of the glycosylation profiles of Alzheimer's beta -secretase protein Asp-2 expressed in a variety of cell lines. AB - Amyloid 39-42 beta -peptides are the main components of amyloid plaques found in the brain of Alzheimer's disease patients. Amyloid 39-42 beta-peptide is formed from amyloid precursor protein by the sequential action of beta- and gamma secretases. Asp-2 is a transmembrane aspartic protease expressed in the brain, shown to have beta-secretase activity. Mature Asp-2 has four N-glycosylation sites. In this report we have characterized the carbohydrate structures in this glycoprotein expressed in three different cell lines, namely Chinese hamster ovary, CV-1 origin of SV40, and baculovirus-infected SF9 cells. Biantennary and triantennary oligosaccharides of the "complex" type were released from glycoprotein expressed in the mammalian cells, whereas mannose-rich glycans were identified from glycoprotein synthesized in the baculovirus-infected cells. Site directed mutagenesis of the asparagine residues at amino acid positions 153, 172, 223, and 354 demonstrate that the protease activity of Asp-2 is dependent on its glycosylation. PMID- 11278493 TI - Specific phosphorylation of threonine by the Dictyostelium myosin II heavy chain kinase family. AB - Dictyostelium myosin II heavy chain kinase A (MHCK A), MHCK B, and MHCK C contain a novel type of protein kinase catalytic domain that displays no sequence identity to the catalytic domain present in conventional serine, threonine, and/or tyrosine protein kinases. Several proteins, including myelin basic protein, myosin regulatory light chain, caldesmon, and casein were phosphorylated by the bacterially expressed MHCK A, MHCK B, and MHCK C catalytic domains. Phosphoamino acid analyses of the proteins showed that 91 to 99% of the phosphate was incorporated into threonine with the remainder into serine. Acceptor amino acid specificity was further examined using a synthetic peptide library (MAXXXX(S/T)XXXXAKKK; where X is any amino acid except cysteine, tryptophan, serine, and threonine and position 7 contains serine and threonine in a 1.7:1 ratio). Phosphorylation of the peptide library with the three MHCK catalytic domains resulted in 97 to 99% of the phosphate being incorporated into threonine, while phosphorylation with a conventional serine/threonine protein kinase, the p21-activated kinase, resulted in 80% of the phosphate being incorporated into serine. The acceptor amino acid specificity of MHCK A was tested directly by substituting serine for threonine in a synthetic peptide and a glutathione S transferase fusion peptide substrate. The serine-containing substrates were phosphorylated at a 25-fold lower rate than the threonine-containing substrates. The results indicate that the MHCKs are specific for the phosphorylation of threonine. PMID- 11278494 TI - Lack of abundance of cytoplasmic cyclosporin A-binding protein renders free living Leishmania donovani resistant to cyclosporin A. AB - The majority of the effects of cyclosporin A (CsA) on cells is caused by the inhibition of phosphatase activity of calcineurin (CN) by the cyclophilin A (CyPA)-CsA complex formed in the cytoplasm. Although CsA inhibits the proliferation of a large number of parasites, not all are susceptible. The presence of structurally altered CyPA with lower affinity for CsA had been suggested to be the cause of resistance. We report here the identification and cloning of a high affinity CsA-binding protein (LdCyP) from Leishmania donovani, a trypanosomatid parasite that is naturally resistant to CsA. The translated LdCyP consists of 187 amino acids with a cleavable 21-amino acid hydrophobic NH(2)-terminal extension. Modeling studies confirmed that all the residues of human CyPs responsible for interaction with CsA are sequentially and conformationally conserved in LdCyP. The purified recombinant protein displayed biochemical parameters comparable to human CyPs. Reverse transcription-polymerase chain reaction analysis confirmed that LdCyP was abundantly expressed. Immunoblot experiments and direct CsA binding studies revealed that LdCyP located in the subcellular organelles constituted the bulk of the CsA binding activity present in L. donovani, whereas the level of binding activity in the cytosol was conspicuously low. CsA selectively facilitated the secretion of LdCyP in the culture medium. Based on these results, it is concluded that the insensitivity of L. donovani to CsA is probably due to the paucity of CsA binding activity in the cytoplasm of the parasite. We suggest that LdCyP, located in the secretory pathway, may function as a chaperone by binding to membrane proteins rather than as the mediator of CN inhibition. PMID- 11278495 TI - Foreign DNA integration. Genome-wide perturbations of methylation and transcription in the recipient genomes. AB - In hamster cells transgenic for the DNA of adenovirus type 12 (Ad12) or for the DNA of bacteriophage lambda, the patterns of DNA methylation in specific cellular genes or DNA segments remote from the site of transgene insertion were altered. In the present report, a wide scope of cellular DNA segments and genes was analyzed. The technique of methylation-sensitive representational difference analysis (MS-RDA) was based on a subtractive hybridization protocol after selecting against DNA segments that were heavily methylated and hence rarely cleaved by the methylation-sensitive endonuclease HpaII. The MS-RDA protocol led to the isolation of several cellular DNA segments that were indeed more heavily methylated in lambda DNA-transgenic hamster cell lines. By applying the suppressive subtractive hybridization technique to cDNA preparations from nontransgenic and Ad12-transformed or lambda DNA-transgenic hamster cells, several cellular genes with altered transcription patterns were cloned from Ad12 transformed or lambda DNA-transgenic hamster cells. Many of the DNA segments with altered methylation, which were isolated by a newly developed methylation sensitive amplicon subtraction protocol, and cDNA fragments derived from genes with altered transcription patterns were identified by their nucleotide sequences. In control experiments, no differences in gene expression or DNA methylation patterns were detectable among individual nontransgenic BHK21 cell clones. In one mouse line transgenic for the DNA of bacteriophage lambda, hypermethylation was observed in the imprinted Igf2r gene in DNA from heart muscle. Two mouse lines transgenic for an adenovirus promoter-indicator gene construct showed hypomethylation in the interleukin 10 and Igf2r loci. We conclude that the insertion of foreign DNA into an established mammalian genome can lead to alterations in cellular DNA methylation and transcription patterns. It is conceivable that the genes and DNA segments affected by these alterations depend on the site(s) of foreign DNA insertion. PMID- 11278496 TI - Taf(II) 250 phosphorylates human transcription factor IIA on serine residues important for TBP binding and transcription activity. AB - Transcription factor IIA (TFIIA) is a positive acting general factor that contacts the TATA-binding protein (TBP) and mediates an activator-induced conformational change in the transcription factor IID (TFIID) complex. Previously, we have found that phosphorylation of yeast TFIIA stimulates TFIIA.TBP.TATA complex formation and transcription activation in vivo. We now show that human TFIIA is phosphorylated in vivo on serine residues that are partially conserved between yeast and human TFIIA large subunits. Alanine substitution mutation of serine residues 316 and 321 in TFIIA alphabeta reduced TFIIA phosphorylation significantly in vivo. Additional alanine substitutions at serines 280 and 281 reduced phosphorylation to undetectable levels. Mutation of all four serine residues reduced the ability of TFIIA to stimulate transcription in transient transfection assays with various activators and promoters, indicating that TFIIA phosphorylation is required globally for optimal function. In vitro, holo-TFIID and TBP-associated factor 250 (TAF(II)250) phosphorylated TFIIA on the beta subunit. Mutation of the four serines required for in vivo phosphorylation eliminated TFIID and TAF(II)250 phosphorylation in vitro. The NH(2)-terminal kinase domain of TAF(II)250 was sufficient for TFIIA phosphorylation, and this activity was inhibited by full-length retinoblastoma protein but not by a retinoblastoma protein mutant defective for TAF(II)250 interaction or tumor suppressor activity. TFIIA phosphorylation had little effect on the TFIIA.TBP.TATA complex in electrophoretic mobility shift assay. However, phosphorylation of TFIIA containing a gamma subunit Y65A mutation strongly stimulated TFIIA.TBP.TATA complex formation. TFIIA-gammaY65A is defective for binding to the beta-sheet domain of TBP identified in the crystal structure. These results suggest that TFIIA phosphorylation is important for strengthening the TFIIA.TBP contact or creating a second contact between TFIIA and TBP that was not visible in the crystal structure. PMID- 11278497 TI - A novel family of chitin-binding proteins from insect type 2 peritrophic matrix. cDNA sequences, chitin binding activity, and cellular localization. AB - The peritrophic matrix is a prominent feature of the digestive tract of most insects, but its function, formation, and even its composition remain contentious. This matrix is a molecular sieve whose toughness and elasticity are generated by glycoproteins, proteoglycans, and chitin fibrils. We now describe a small, highly conserved protein, peritrophin-15, which is an abundant component of the larval peritrophic matrices of the Old World screwworm fly, Chrysomya bezziana, and sheep blowfly, Lucilia cuprina. Their deduced amino acid sequences code for a 8-kDa secreted protein characterized by a highly conserved and novel register of six cysteines. Two Drosophila homologues have also been identified from unannotated genomic sequences. Recombinant peritrophin-15 binds strongly and specifically to chitin; however, the stoichiometry of binding is low (1:10,000 N acetyl glucosamine). We propose that peritrophin-15 caps the ends of the chitin polymer. Immunogold studies localized peritrophin-15 to the peritrophic matrix and specific vesicles in cells of the cardia, the small organ of the foregut responsible for peritrophic matrix synthesis. The vesicular contents are disgorged at the base of microvilli underlying the newly formed peritrophic matrix. This is the first time that the process of synthesis and integration of a peritrophic matrix protein into the nascent peritrophic matrix has been observed. PMID- 11278498 TI - Dimeric fragment of the insulin receptor alpha-subunit binds insulin with full holoreceptor affinity. AB - The insulin receptor (IR) is a dimeric receptor, and its activation is thought to involve cross-linking between monomers initiated by binding of a single insulin molecule to separate epitopes on each monomer. We have previously shown that a minimized insulin receptor consisting of the first three domains of the human IR fused to 16 amino acids from the C-terminal of the alpha-subunit was monomeric and bound insulin with nanomolar affinity (Kristensen, C., Wiberg, F. C., Schaffer, L., and Andersen, A. S. (1998) J. Biol. Chem. 273, 17780-17786). To investigate the insulin binding properties of dimerized alpha-subunits, we have reintroduced the domains containing alpha-alpha disulfide bonds into this minireceptor. When inserting either the first fibronectin type III domain or the full-length sequence of exon 10, the receptor fragments were predominantly secreted as disulfide-linked dimers that both had nanomolar affinity for insulin, similar to the affinity found for the minireceptor. However, when both these domains were included we obtained a soluble dimeric receptor that bound insulin with 1000-fold higher affinity (4-8 pm) similar to what was obtained for the solubilized holoreceptor (14-24 pm). Moreover, dissociation of labeled insulin from this receptor was accelerated in the presence of unlabeled insulin, demonstrating another characteristic feature of the holoreceptor. This is the first direct demonstration showing that the alpha-subunit of IR contains all the epitopes required for binding insulin with full holoreceptor affinity. PMID- 11278499 TI - The water channel aquaporin-8 is mainly intracellular in rat hepatocytes, and its plasma membrane insertion is stimulated by cyclic AMP. AB - We previously found that water transport across hepatocyte plasma membranes occurs mainly via a non-channel mediated pathway. Recently, it has been reported that mRNA for the water channel, aquaporin-8 (AQP8), is present in hepatocytes. To further explore this issue, we studied protein expression, subcellular localization, and regulation of AQP8 in rat hepatocytes. By subcellular fractionation and immunoblot analysis, we detected an N-glycosylated band of approximately 34 kDa corresponding to AQP8 in hepatocyte plasma and intracellular microsomal membranes. Confocal immunofluorescence microscopy for AQP8 in cultured hepatocytes showed a predominant intracellular vesicular localization. Dibutyryl cAMP (Bt(2)cAMP) stimulated the redistribution of AQP8 to plasma membranes. Bt(2)cAMP also significantly increased hepatocyte membrane water permeability, an effect that was prevented by the water channel blocker dimethyl sulfoxide. The microtubule blocker colchicine but not its inactive analog lumicolchicine inhibited the Bt(2)cAMP effect on both AQP8 redistribution to cell surface and hepatocyte membrane water permeability. Our data suggest that in rat hepatocytes AQP8 is localized largely in intracellular vesicles and can be redistributed to plasma membranes via a microtubule-depending, cAMP-stimulated mechanism. These studies also suggest that aquaporins contribute to water transport in cAMP stimulated hepatocytes, a process that could be relevant to regulated hepatocyte bile secretion. PMID- 11278500 TI - SPIN90 (SH3 protein interacting with Nck, 90 kDa), an adaptor protein that is developmentally regulated during cardiac myocyte differentiation. AB - In the yeast two-hybrid screening, we have isolated a cDNA clone from a human heart library using Nck Src homology 3 (SH3) domains as bait. The full-length cDNA, which encoded 722 amino acids, was identified as a VIP54-related gene containing an SH3 domain, proline-rich motifs, a serine/threonine-rich region, and a long C-terminal hydrophobic region. We refer to this protein as SPIN90 (SH3 Protein Interacting with Nck, 90 kDa). The amino acid sequence of the SH3 domain has the highest homology with those of Fyn, Yes, and c-Src. SPIN90 was broadly expressed in human tissues; in particular, it was highly expressed in heart, brain, and skeletal muscle, and its expression was developmentally regulated during cardiac myocyte differentiation. SPIN90 is able to bind to the first and third SH3 domains of Nck, in vitro, and is colocalized with Nck at sarcomere Z discs within cardiac myocytes. Moreover, treatment with antisera raised against SPIN90 disrupted sarcomere structure, suggesting that this protein may play an important role in the maintenance of sarcomere structure and/or in the assembly of myofibrils into sarcomeres. PMID- 11278501 TI - Gaf-1, a gamma -SNAP-binding protein associated with the mitochondria. AB - The role of alpha/beta-SNAP (Soluble NSF Attachment Protein) in vesicular trafficking is well established; however, the function of the ubiquitously expressed gamma-SNAP remains unclear. To further characterize the cellular role of this enigmatic protein, a two-hybrid screen was used to identify new, gamma SNAP-binding proteins and to uncover potentially novel functions for gamma-SNAP. One such SNAP-binding protein, termed Gaf-1 (gamma-SNAP associate factor-1) specifically binds gamma- but not alpha-SNAP. The full-length Gaf-1 (75 kDa) is ubiquitously expressed and is found stoichiometrically associated with gamma-SNAP in cellular extracts. This binding is distinct from other SNAP interactions since no alpha-SNAP or NSF coprecipitated with Gaf-1. Subcellular fractionation and immunofluorescence analysis show that Gaf-1 is peripherally associated with the outer mitochondrial membrane. Only a fraction of gamma-SNAP was mitochondrial with the balance being either cytosolic or associated with other membrane fractions. GFP-gamma-SNAP and the C-terminal domain of Gaf-1 both show a reticular distribution in HEK-293 cells. This reticular structure colocalizes with Gaf-1 and mitochondria as well as with microtubules but not with other cytoskeletal elements. These data identify a class of gamma-SNAP interactions that is distinct from other members of the SNAP family and point to a potential role for gamma-SNAP in mitochondrial dynamics. PMID- 11278502 TI - Scp160p, an RNA-binding, polysome-associated protein, localizes to the endoplasmic reticulum of Saccharomyces cerevisiae in a microtubule-dependent manner. AB - Scp160p is an RNA-binding protein containing 14 tandemly repeated heterogenous nuclear ribonucleoprotein K-homology domains, which are implicated in RNA binding. Scp160p interacts with free and membrane-bound polysomes that are dependent upon the presence of mRNA. Despite its presence on cytosolic polysomes, Scp160p is predominantly localized to the endoplasmic reticulum (ER). Accumulation of Scp160p-ribosome complexes at the ER requires the function of microtubules but is independent of the actin cytoskeleton. We propose that the multi-K-homology-domain protein Scp160p functions as an RNA binding platform, interacting with polysomes that are transported to the ER. PMID- 11278503 TI - CYP2A6*6, a novel polymorphism in cytochrome p450 2A6, has a single amino acid substitution (R128Q) that inactivates enzymatic activity. AB - By using the polymerase chain reaction technique combined with restriction enzyme fragment length polymorphism (PCR-RFLP), a novel polymorphism of CYP2A6, CYP2A6*6, was detected in 0.4% of the Japanese population. To study the enzymatic properties of the CYP2A6.6 protein with a single amino acid substitution of arginine 128 to glutamine, both this isozyme and the CYP2A6.1 protein (wild-type) were produced in insect cells using a baculovirus system. Coumarin 7 hydroxylation, which reflects CYP2A6 activity, was significantly reduced (one eighth of normal) in cell lysate from CYP2A6*6-transfected Sf9 cells compared with that lysate from CYP2A6*1-transfected cells. To clarify the mechanism of inactivation of the CYP2A6.6 enzyme, the heme content and reduced CO difference spectrum were examined. Although CYP2A6.6 retained about one-half the heme content of CYP2A6.1, the reduced CO-bound Soret peak was completely lost. These results suggest that the inactivation of CYP2A6.6 is mainly due to disordering of the holoprotein structure rather than a failure of heme incorporation. PMID- 11278504 TI - Identification of essential residues in 2',3'-cyclic nucleotide 3' phosphodiesterase. Chemical modification and site-directed mutagenesis to investigate the role of cysteine and histidine residues in enzymatic activity. AB - 2',3'-Cyclic nucleotide 3'-phosphodiesterase (CNP; EC ) catalyzes in vitro hydrolysis of 3'-phosphodiester bonds in 2',3'-cyclic nucleotides to produce 2' nucleotides exclusively. N-terminal deletion mapping of the C-terminal two-thirds of recombinant rat CNP1 identified a region that possesses the catalytic domain, with further truncations abolishing activity. Proteolysis and kinetic analysis indicated that this domain forms a compact globular structure and contains all of the catalytically essential features. Subsequently, this catalytic fragment of CNP1 (CNP-CF) was used for chemical modification studies to identify amino acid residues essential for activity. 5,5'-Dithiobis-(2-nitrobenzoic acid) modification studies and kinetic analysis of cysteine CNP-CF mutants revealed the nonessential role of cysteines for enzymatic activity. On the other hand, modification studies with diethyl pyrocarbonate indicated that two histidines are essential for CNPase activity. Consequently, the only two conserved histidines, His-230 and His-309, were mutated to phenylalanine and leucine. All four histidine mutants had k(cat) values 1000-fold lower than wild-type CNP-CF, but K(m) values were similar. Circular dichroism studies demonstrated that the low catalytic activities of the histidine mutants were not due to gross changes in secondary structure. Taken together, these results demonstrate that both histidines assume critical roles for catalysis. PMID- 11278505 TI - Insulin signals to prenyltransferases via the Shc branch of intracellular signaling. AB - We assessed the roles of insulin receptor substrate-1 (IRS-1) and Shc in insulin action on farnesyltransferase (FTase) and geranylgeranyltransferase I (GGTase I) using Chinese hamster ovary (CHO) cells that overexpress wild-type human insulin receptors (CHO-hIR-WT) or mutant insulin receptors lacking the NPEY domain (CHO DeltaNPEY) or 3T3-L1 fibroblasts transfected with adenoviruses that express the PTB or SAIN domain of IRS-1 and Shc, the pleckstrin homology (PH) domain of IRS 1, or the Src homology 2 (SH2) domain of Shc. Insulin promoted phosphorylation of the alpha-subunit of FTase and GGTase I in CHO-hIR-WT cells, but was without effect in CHO-DeltaNPEY cells. Insulin increased FTase and GGTase I activities and the amounts of prenylated Ras and RhoA proteins in CHO-hIR-WT (but not CHO DeltaNPEY) cells. Overexpression of the PTB or SAIN domain of IRS-1 (which blocked both IRS-1 and Shc signaling) prevented insulin-stimulated phosphorylation of the FTase and GGTase I alpha-subunit activation of FTase and GGTase I and subsequent increases in prenylated Ras and RhoA proteins. In contrast, overexpression of the IRS-1 PH domain, which impairs IRS-1 (but not Shc) signaling, did not alter insulin action on the prenyltransferases, but completely inhibited the insulin effect on the phosphorylation of IRS-1 and on the activation of phosphatidylinositol 3-kinase and Akt. Finally, overexpression of the Shc SH2 domain completely blocked the insulin effect on FTase and GGTase I activities without interfering with insulin signaling to MAPK. These data suggest that insulin signaling from its receptor to the prenyltransferases FTase and GGTase I is mediated by the Shc pathway, but not the IRS-1/phosphatidylinositol 3 kinase pathway. Shc-mediated insulin signaling to MAPK may be necessary (but not sufficient) for activation of prenyltransferase activity. An additional pathway involving the Shc SH2 domain may be necessary to mediate the insulin effect on FTase and GGTase I. PMID- 11278506 TI - A stable organic free radical in anaerobic benzylsuccinate synthase of Azoarcus sp. strain T. AB - The novel enzyme benzylsuccinate synthase initiates anaerobic toluene metabolism by catalyzing the addition of toluene to fumarate, forming benzylsuccinate. Based primarily on its sequence similarity to the glycyl radical enzymes, pyruvate formate-lyase and anaerobic ribonucleotide reductase, benzylsuccinate synthase was speculated to be a glycyl radical enzyme. In this report we use EPR spectroscopy to demonstrate for the first time that active benzylsuccinate synthase from the denitrifying bacterium Azoarcus sp. strain T harbors an oxygen sensitive stable organic free radical. The EPR signal of the radical was centered at g = 2.0021 and was characterized by a major 2-fold splitting of about 1.5 millitesla. The strong similarities between the EPR signal of the benzylsuccinate synthase radical and that of the glycyl radicals of pyruvate formate-lyase and anaerobic ribonucleotide reductase provide evidence that the benzylsuccinate synthase radical is located on a glycine residue, presumably glycine 828 in Azoarcus sp. strain T benzylsuccinate synthase. PMID- 11278507 TI - Cloning and characterization of adenylate kinase from Chlamydia pneumoniae. AB - Chlamydiae proliferate only within the infected host cells and are thought to be "energy parasites," because they take up ATP from the host cell as an energy source. In the present study, we isolated from Chlamydia pneumoniae the gene encoding adenylate kinase (AK). Using the enzyme produced in Escherichia coli, its properties were characterized. K(m) values for AMP and for ADP of the purified C. pneumoniae AK (AKcpn) were each 330 microm, which is significantly higher than the reported values of other AKs, whereas K(m) for ATP was 24 microm, which was rather lower than others. AKcpn contains 1 g atom of zinc/mol of 24,000 dalton protein. Mass spectrometric analysis of AKcpn and analysis of properties of mutated AKcpn strongly suggested that zinc is associated with four cysteine residues in the LID domain of the enzyme. The apo-AKcpn that lost zinc retained AK activity, although K(m) for AMP of apo-AKcpn increased about 2-fold and V(max) decreased about one-half from that of holo-AKcpn. The apo-AKcpn was more thermolabile and sensitive to trypsin digestion than the holo-AKcpn. Moreover, the recovery in vitro of the AK activity during the renaturation process of the denatured apo-AKcpn was dependent on zinc. A mutated protein in which cysteine residues in the LID domain were substituted by other amino acids lost both zinc and enzyme activity. The mutated protein was more sensitive to protease than the apo-AKcpn. These results indicate that zinc in AKcpn, although not essential for the catalysis, stabilizes the enzyme and probably plays a crucial role in proper folding of the protein. Furthermore, the catalytic properties of AKcpn suggest a distinctive regulatory mechanism in the metabolism compared with AKs in other organisms. PMID- 11278508 TI - Expression cloning of a Na+-independent aromatic amino acid transporter with structural similarity to H+/monocarboxylate transporters. AB - A cDNA was isolated from rat small intestine by expression cloning which encodes a novel Na+-independent transporter for aromatic amino acids. When expressed in Xenopus oocytes, the encoded protein designated as TAT1 (T-type amino acid transporter 1) exhibited Na+-independent and low-affinity transport of aromatic amino acids such as tryptophan, tyrosine, and phenylalanine (Km values: approximately 5 mm), consistent with the properties of classical amino acid transport system T. TAT1 accepted some variations of aromatic side chains because it interacted with amino acid-related compounds such as l-DOPA and 3-O-methyl DOPA. Because TAT1 accepted N-methyl- and N-acetyl-derivatives of aromatic amino acids but did not accept their methylesters, it is proposed that TAT1 recognizes amino acid substrates as anions. Consistent with this, TAT1 exhibited sequence similarity (approximately 30% identity at the amino acid level) to H+/monocarboxylate transporters. Distinct from H+/monocarboxylate transporters, however, TAT1 was not coupled with the H+ transport but it mediated an electroneutral facilitated diffusion. TAT1 mRNA was strongly expressed in intestine, placenta, and liver. In rat small intestine TAT1 immunoreactivity was detected in the basolateral membrane of the epithelial cells suggesting its role in the transepithelial transport of aromatic amino acids. The identification of the amino acid transporter with distinct structural and functional characteristics will not only facilitate the expansion of amino acid transporter families but also provide new insights into the mechanisms of substrate recognition of organic solute transporters. PMID- 11278509 TI - Potential role for the BLM helicase in recombinational repair via a conserved interaction with RAD51. AB - Bloom's syndrome (BS) is an autosomal recessive disorder that predisposes individuals to a wide range of cancers. The gene mutated in BS, BLM, encodes a member of the RecQ family of DNA helicases. The precise role played by these enzymes in the cell remains to be determined. However, genome-wide hyper recombination is a feature of many RecQ helicase-deficient cells. In eukaryotes, a central step in homologous recombination is catalyzed by the RAD51 protein. In response to agents that induce DNA double-strand breaks, RAD51 accumulates in nuclear foci that are thought to correspond to sites of recombinational repair. Here, we report that purified BLM and human RAD51 interact in vitro and in vivo, and that residues in the N- and C-terminal domains of BLM can independently mediate this interaction. Consistent with these observations, BLM localizes to a subset of RAD51 nuclear foci in normal human cells. Moreover, the number of BLM foci and the extent to which BLM and RAD51 foci co-localize increase in response to ionizing radiation. Nevertheless, the formation of RAD51 foci does not require functional BLM. Indeed, in untreated BS cells, an abnormally high proportion of the cells contain RAD51 nuclear foci. Exogenous expression of BLM markedly reduces the fraction of cells containing RAD51 foci. The interaction between BLM and RAD51 appears to have been evolutionarily conserved since the C-terminal domain of Sgs1, the Saccharomyces cerevisiae homologue of BLM, interacts with yeast Rad51. Furthermore, genetic analysis reveals that the SGS1 and RAD51 genes are epistatic indicating that they operate in a common pathway. Potential roles for BLM in the RAD51 recombinational repair pathway are discussed. PMID- 11278510 TI - The dimerization interface of the metastasis-associated protein S100A4 (Mts1): in vivo and in vitro studies. AB - The S100 calcium-binding proteins are implicated in signal transduction, motility, and cytoskeletal dynamics. The three-dimensional structure of several S100 proteins revealed that the proteins form non-covalent dimers. However, the mechanism of the S100 dimerization is still obscure. In this study we characterized the dimerization of S100A4 (also named Mts1) in vitro and in vivo. Analytical ultracentrifugation revealed that apoS100A4 was present in solution as a mixture of monomers and dimers in a rapidly reversible equilibrium (K(d) = 4 +/ 2 microm). The binding of calcium promoted dimerization. Replacement of Tyr-75 by Phe resulted in the stabilization of the dimer. Helix IV is known to form the major part of the dimerization interface in homologous S100 proteins. By using the yeast two-hybrid system we showed that only a few residues of helix IV, namely Phe-72, Tyr-75, Phe-78, and Leu-79, are essential for dimerization in vivo. A homology model demonstrated that these residues form a hydrophobic cluster on helix IV. Their role is to stabilize the structure of individual subunits rather than provide specific interactions across the dimerization surface. Our mutation data showed that the specificity at the dimerization surface is not particularly stringent, which is consistent with recent data indicating that S100 proteins can form heterodimers. PMID- 11278511 TI - Structure of human thymidylate synthase suggests advantages of chemotherapy with noncompetitive inhibitors. AB - Thymidylate synthase (TS) is a major target in the chemotherapy of colorectal cancer and some other neoplasms. The emergence of resistance to the treatment is often related to the increased levels of TS in cancer cells, which have been linked to the elimination of TS binding to its own mRNA upon drug binding, a feedback regulatory mechanism, and/or to the increased stability to intracellular degradation of TS.drug complexes (versus unliganded TS). The active site loop of human TS (hTS) has a unique conformation resulted from a rotation by 180 degrees relative to its orientation in bacterial TSs. In this conformation, the enzyme must be inactive, because the catalytic cysteine is no longer positioned in the ligand-binding pocket. The ordered solvent structure obtained from high resolution crystallographic data (2.0 A) suggests that the inactive loop conformation promotes mRNA binding and intracellular degradation of the enzyme. This hypothesis is supported by fluorescence studies, which indicate that in solution both active and inactive forms of hTS are present. The binding of phosphate ion shifts the equilibrium toward the inactive conformation; subsequent dUMP binding reverses the equilibrium toward the active form. Thus, TS inhibition via stabilization of the inactive conformation should lead to less resistance than is observed with presently used drugs, which are analogs of its substrates, dUMP and CH(2)H(4)folate, and bind in the active site, promoting the active conformation. The presence of an extension at the N terminus of native hTS has no significant effect on kinetic properties or crystal structure. PMID- 11278512 TI - A regulatory role of the PetM subunit in a cyanobacterial cytochrome b6f complex. AB - To investigate the function of the PetM subunit of the cytochrome b6f complex, the petM gene encoding this subunit was inactivated by insertional mutagenesis in the cyanobacterium Synechocystis PCC 6803. Complete segregation of the mutant reveals a nonessential function of PetM for the structure and function of the cytochrome b6f complex in this organism. Photosystem I, photosystem II, and the cytochrome b6f complex still function normally in the petM- mutant as judged by cytochrome f re-reduction and oxygen evolution rates. In contrast to the wild type, however, the content of phycobilisomes and photosystem I as determined from 77 K fluorescence spectra is reduced in the petM- strain. Furthermore, whereas under anaerobic conditions the kinetics of cytochrome f re-reduction are identical, under aerobic conditions these kinetics are slower in the petM- strain. Fluorescence induction measurements indicate that this is due to an increased plastoquinol oxidase activity in the mutant, causing the plastoquinone pool to be in a more oxidized state under aerobic dark conditions. The finding that the activity of the cytochrome b6f complex itself is unchanged, whereas the stoichiometry of other protein complexes has altered, suggests an involvement of the PetM subunit in regulatory processes mediated by the cytochrome b6f complex. PMID- 11278513 TI - Structural characterization of pyrrolic cross-links in collagen using a biotinylated Ehrlich's reagent. AB - The structures of pyrrolic forms of cross-links in collagen have been confirmed by reacting collagen peptides with a biotinylated Ehrlich's reagent. This reagent was synthesized by converting the cyano group of N-methyl-N-cyanoethyl-4 aminobenzaldehyde to a carboxylic acid, followed by conjugation with biotin pentyl-amine. Derivatization of peptides from bone collagen both stabilized the pyrroles and facilitated selective isolation of the pyrrole-containing peptides using a monomeric avidin column. Reactivity of the biotinylated reagent with collagen peptides was similar to that of the standard Ehrlich reagent, but heat denaturation of the tissue before enzyme digestion resulted in the loss of about 50% of the pyrrole cross-links. Identification of a series of peptides by mass spectrometry confirmed the presence of derivatized pyrrole structures combined with between 1 and 16 amino acid residues. Almost all of the pyrrole-containing peptides appeared to be derived from N-terminal telopeptide sequences, and the nonhydroxylated (lysine-derived) form predominated over pyrrole cross-links derived from helical hydroxylysine. PMID- 11278514 TI - Detection and binding properties of GABA(A) receptor assembly intermediates. AB - Density gradient centrifugation of native and recombinant gamma-aminobutyric acid, type A (GABA(A)) receptors was used to detect assembly intermediates. No such intermediates could be identified in extracts from adult rat brain or from human embryonic kidney (HEK) 293 cells transfected with alpha(1), beta(3), and gamma(2) subunits and cultured at 37 degrees C. However, subunit dimers, trimers, tetramers, and pentamers were found in extracts from the brain of 8-10-day-old rats and from alpha(1)beta(3)gamma(2) transfected HEK cells cultured at 25 degrees C. In both systems, alpha(1), beta(3), and gamma(2) subunits could be identified in subunit dimers, indicating that different subunit dimers are formed during GABA(A) receptor assembly. Co-transfection of HEK cells with various combinations of full-length and C-terminally truncated alpha(1) and beta(3) or alpha(1) and gamma(2) subunits and co-immunoprecipitation with subunit-specific antibodies indicated that even subunits containing no transmembrane domain can assemble with each other. Whereas alpha(1)gamma(2), alpha(1)Ngamma(2), alpha(1)gamma(2)N, and alpha(1)Ngamma(2)N, combinations exhibited specific [(3)H]Ro 15-1788 binding, specific [(3)H]muscimol binding could only be found in alpha(1)beta(3) and alpha(1)beta(3)N, but not in alpha(1)Nbeta(3) or alpha(1)Nbeta(3)N combinations. This seems to indicate that a full-length alpha(1) subunit is necessary for the formation of the muscimol-binding site and for the transduction of agonist binding into channel gating. PMID- 11278515 TI - A gamma-2 herpesvirus nucleocytoplasmic shuttle protein interacts with importin alpha 1 and alpha 5. AB - Herpesvirus saimiri (HVS) is the prototype gamma-2 herpesvirus. This is an increasing important subfamily of herpesviruses due to the identification of the first human gamma-2 herpesvirus, Kaposi's sarcoma-associated herpesvirus. The HVS open reading frame (ORF) 57 protein is a multifunctional trans-regulatory protein homologous to genes identified in all classes of herpesviruses. Recent analysis has demonstrated that ORF 57 has the ability to bind viral RNA and to shuttle between the nucleus and cytoplasm, and is required for efficient nuclear export of viral transcripts. Here we have investigated the nucleocytoplasmic shuttling mechanism utilized by the ORF 57 protein. The yeast two-hybrid system was employed to identify interacting cellular proteins using ORF 57 as bait. We demonstrate that ORF 57 interacts with importin alpha isoforms 1 and 5. In addition, the binding of ORF 57 to importin alpha was mediated by the importin alpha hydrophobic internal armadillo repeats. An ORF 57 amino-terminal arginine rich sequence, which functions as a nuclear localization sequence, was also required for this interaction. Furthermore, the ORF 57 protein is responsible for the redistribution of importin alpha into the nucleoli. These results identify novel cellular interactions essential for the functioning of this important herpesvirus regulatory protein. PMID- 11278516 TI - In vivo proteolytic degradation of the Escherichia coli acyltransferase HlyC. AB - Escherichia coli hemolysin (HlyA) is the prototype toxin of a major family of exoproteins produced by Gram-negative bacteria known as "repeats in toxins." Only fatty acid-acylated HlyA molecules at residues Lys564 and Lys690 are able to damage the target cell membrane. Fatty acylation of pro-HlyA is dependent on the co-synthesized acyltransferase HlyC and the acylated form of acyl-carrier protein. By using a collection of hlyA and hlyC mutant strains, the processing of HlyC was investigated. HlyC was not detected by Western blot in an E. coli strain encoding hlyC and hlyA, but it was present in a strain encoding only hlyC. The hlyC mRNA pattern, however, was similar in both strains indicating that the turnover of HlyC does not occur at the transcriptional level. HlyC was detected in Western blots of cell lysates from an E. coli strain encoding HlyC and a HlyA derivative where both acylation sites were substituted. Similar results were obtained when HlyC was expressed in a hlyA mutant strain lacking part of a putative HlyC binding domain, indicating that this particular HlyA region affects HlyC stability. We did not detect HlyC in cell lysates from hlyC mutants with different abilities to acylate pro-HlyA, suggesting that the degradation of HlyC is not related to the HlyA acylation process. The protease systems ClpAP, ClpXP, and FtsH were found to be responsible for the HlyA-dependent processing of HlyC. PMID- 11278517 TI - Induction of apoptosis through B-cell receptor cross-linking occurs via de novo generated C16-ceramide and involves mitochondria. AB - B-cells, triggered via their surface B-cell receptor (BcR), start an apoptotic program known as activation-induced cell death (AICD), and it is widely believed that this phenomenon plays a role in the restriction and focusing of the immune response. Although both ceramide and caspases have been proposed to be involved in AICD, the contribution of either and the exact molecular events through which AICD commences are still unknown. Here we show that in Ramos B-cells, BcR triggered cell death is associated with an early rise of C16 ceramide that derives from activation of the de novo pathway, as demonstrated using a specific inhibitor of ceramide synthase, fumonisin B1 (FB1), and using pulse labeling with the metabolic sphingolipid precursor, palmitate. There was no evidence for activation of sphingomyelinases or hydrolysis of sphingomyelin. Importantly, FB1 inhibited several specific apoptotic hallmarks such as poly(A)DP-ribose polymerase cleavage and DNA fragmentation. Electron microscopy revealed morphological evidence of mitochondrial damage, suggesting the involvement of mitochondria in BcR-triggered apoptosis, and this was inhibited by FB1. Moreover, a loss of mitochondrial membrane potential was observed in Ramos cells after BcR cross-linking, which was inhibited by the addition of FB1. Interestingly, benzyloxycarbonyl-Val-Ala-dl-Asp, a broad spectrum caspase inhibitor did not inhibit BcR-induced mitochondrial membrane permeability transition but did block DNA fragmentation. These results suggest a crucial role for de novo generated C16 ceramide in the execution of AICD, and they further suggest an ordered and more specific sequence of biochemical events in which de novo generated C16 ceramide is involved in mitochondrial damage resulting in a downstream activation of caspases and apoptosis. PMID- 11278518 TI - Characterization of a novel isoform of caspase-9 that inhibits apoptosis. AB - We have identified a novel isoform of rat caspase-9 in which the C terminus of full-length caspase-9 is replaced with an alternative peptide sequence. Casp-9 CTD (where CTD is carboxyl-terminal divergent) is expressed in multiple tissues, with the relative highest expression observed in ovary and heart. Casp-9-CTD was found primarily in the cytoplasm and was not detected in the nucleus. Structural predictions suggest that in contrast to full-length caspase-9, casp-9-CTD will not be processed. Our model is supported by reduced protease activity of casp-9 CTD preparations in vitro and by the lack of detectable processing of casp-9-CTD proenzyme or the induction of cell death following transfection into cells. Both neuronal and non-neuronal cell types transfected with casp-9-CTD were resistant to death evoked by trophic factor deprivation or DNA damage. In addition, cytosolic lysates prepared from cells permanently expressing exogenous casp-9-CTD were resistant to caspase induction by cytochrome c in reconstitution assays. Taken together, our observations indicate that casp-9-CTD acts as a dominant negative variant. Its expression in various tissues indicates a physiological role in regulating cell death. PMID- 11278519 TI - The activity of guanine exchange factor NET1 is essential for transforming growth factor-beta-mediated stress fiber formation. AB - To examine signaling pathways underlying transforming growth factor-beta (TGF beta)-mediated changes in cell morphology, we used a microarray system to identify downstream target genes that may play a role in this process. Through this approach, we found that the NET1 gene was induced upon TGF-beta treatment in several cell types. NET1 is a guanine nucleotide exchange factor for RhoA whose activity has been implicated in stress fiber formation. In the Swiss 3T3 cell line, TGF-beta induces NET1 expression, and this correlated with an increase in stress fiber formation. Overexpression of the wild type NET1 gene increases stress fiber formation, and overexpression of a dominant negative NET1 mutant (L392E) prevented TGF-beta dependent increase in stress fiber formation. Furthermore, treatment of the cells with a RhoA kinase inhibitor Y-27632 blocks TGF-beta-induced stress fiber formation. By using a stable cell line expressing dominant negative Smad3, we found that the Smad signaling pathway is essential for the induction of NET1, which in turn leads to the increase of Rho activity. Taken together, those data suggest that induction of NET1 is important for the increase of Rho activity upon TGF-beta treatment, which may represent the critical trigger for a variety of downstream events in different cells. Our results support the presence of a novel signaling pathway by which TGF-beta may regulate the formation of stress fibers and reorganization of cytoskeletal structures. PMID- 11278520 TI - Cleavage of factor VIII heavy chain is required for the functional interaction of a2 subunit with factor IXA. AB - Factor VIII circulates as a noncovalent heterodimer consisting of a heavy chain (HC, contiguous A1-A2-B domains) and light chain (LC). Cleavage of HC at the A1 A2 and A2-B junctions generates the A1 and A2 subunits of factor VIIIa. Although the isolated A2 subunit stimulates factor IXa-catalyzed generation of factor Xa by approximately 100-fold, the isolated HC, free from the LC, showed no effect in this assay. However, extended reaction of HC with factors IXa and X resulted in an increase in factor IXa activity because of conversion of the HC to A1 and A2 subunits by factor Xa. HC cleavage by thrombin or factor Xa yielded similar products, although factor Xa cleaved at a rate of approximately 1% observed for thrombin. HC showed little inhibition of the A2 subunit-dependent stimulation of factor IXa activity, suggesting that factor IXa-interactive sites are masked in the A2 domain of HC. Furthermore, HC showed no effect on the fluorescence anisotropy of fluorescein-Phe-Phe-Arg-factor IXa in the presence of factor X, whereas thrombin-cleaved HC yielded a marked increase in this parameter. These results indicate that HC cleavage by either thrombin or factor Xa is essential to expose the factor IXa-interactive site(s) in the A2 subunit required to modulate protease activity. PMID- 11278521 TI - Identification and characterization of a low oxygen response element involved in the hypoxic induction of a family of Saccharomyces cerevisiae genes. Implications for the conservation of oxygen sensing in eukaryotes. AB - An organism's ability to respond to changes in oxygen tension depends in large part on alterations in gene expression. The oxygen sensing and signaling mechanisms in eukaryotic cells are not fully understood. To further define these processes, we have studied the Delta9 fatty acid desaturase gene OLE1 in Saccharomyces cerevisiae. We have confirmed previous data showing that the expression of OLE1 mRNA is increased in hypoxia and in the presence of certain transition metals. OLE1 expression was also increased in the presence of the iron chelator 1,10-phenanthroline. A 142-base pair (bp) region 3' to the previously identified fatty acid response element was identified as critical for the induction of OLE1 in response to these stimuli using OLE1 promoter-lacZ reporter constructs. Electromobility shift assays confirmed the presence of an inducible band shift in response to hypoxia and cobalt. Mutational analysis defined the nonameric sequence ACTCAACAA as necessary for transactivation. A 20-base pair oligonucleotide containing this nonamer confers up-regulation by hypoxia and inhibition by unsaturated fatty acids when placed upstream of a heterologous promoter in a lacZ reporter construct. Additional yeast genes were identified which respond to hypoxia and cobalt in a manner similar to OLE1. A number of mammalian genes are also up-regulated by hypoxia, cobalt, nickel, and iron chelators. Hence, the identification of a family of yeast genes regulated in a similar manner has implications for understanding oxygen sensing and signaling in eukaryotes. PMID- 11278522 TI - Role of the N-terminal forkhead-associated domain in the cell cycle checkpoint function of the Rad53 kinase. AB - Forkhead-associated (FHA) domains are multifunctional phosphopeptide-binding modules and are the hallmark of the conserved family of Rad53-like checkpoint protein kinases. Rad53-like kinases, including the human tumor suppressor protein Chk2, play crucial roles in cell cycle arrest and activation of repair processes following DNA damage and replication blocks. Here we show that ectopic expression of the N-terminal FHA domain (FHA1) of the yeast Rad53 kinase causes a growth defect by arresting the cell cycle in G(1). This phenotype was highly specific for the Rad53-FHA1 domain and not observed with the similar Rad53-FHA2, Dun1-FHA, and Chk2-FHA domains, and it was abrogated by mutations that abolished binding to a phosphothreonine-containing peptide in vitro. Furthermore, replacement of the RAD53 gene with alleles containing amino acid substitutions in the FHA1 domain resulted in an increased DNA damage sensitivity in vivo. Taken together, these data demonstrate that the FHA1 domain contributes to the checkpoint function of Rad53, possibly by associating with a phosphorylated target protein in response to DNA damage in G(1). PMID- 11278523 TI - Phosphorylation of Ser363, Thr370, and Ser375 residues within the carboxyl tail differentially regulates mu-opioid receptor internalization. AB - Prolonged activation of opioid receptors leads to their phosphorylation, desensitization, internalization, and down-regulation. To elucidate the relationship between mu-opioid receptor (MOR) phosphorylation and the regulation of receptor activity, a series of receptor mutants was constructed in which the 12 Ser/Thr residues of the COOH-terminal portion of the receptor were substituted to Ala, either individually or in combination. All these mutant constructs were stably expressed in human embryonic kidney 293 cells and exhibited similar expression levels and ligand binding properties. Among those 12 Ser/Thr residues, Ser(363), Thr(370), and Ser(375) have been identified as phosphorylation sites. In the absence of the agonist, a basal phosphorylation of Ser(363) and Thr(370) was observed, whereas [d-Ala(2),Me-Phe(4),Gly(5)-ol]enkephalin (DAMGO)-induced receptor phosphorylation occurs at Thr(370) and Ser(375) residues. Furthermore, the role of these phosphorylation sites in regulating the internalization of MOR was investigated. The mutation of Ser(375) to Ala reduced the rate and extent of receptor internalization, whereas mutation of Ser(363) and Thr(370) to Ala accelerated MOR internalization kinetics. The present data show that the basal phosphorylation of MOR could play a role in modulating agonist-induced receptor internalization kinetics. Furthermore, even though mu-receptors and delta-opioid receptors have the same motif encompassing agonist-induced phosphorylation sites, the different agonist-induced internalization properties controlled by these sites suggest differential cellular regulation of these two receptor subtypes. PMID- 11278524 TI - Determination of the functional epitopes of human interleukin-18-binding protein by site-directed mutagenesis. AB - The human interleukin (IL)-18-binding protein (hIL-18BP) is a naturally occurring antagonist of IL-18, a proinflammatory cytokine that is related to IL-1beta and has an important role in defense against microbial invaders. As its name implies, the hIL-18BP binds to IL-18 with high affinity and prevents the interaction of IL 18 with its receptor. We genetically modified the C terminus of hIL-18BP by appending a 15-amino acid biotinylation recognition site and a six-histidine tag and then performed site-directed mutagenesis to determine the functional epitopes that mediate efficient binding to IL-18. The mutated IL-18BPs were secreted from mammalian cells, captured by metal affinity chromatography, biotinylated in situ, eluted, and immobilized on streptavidin-coated chips. Using surface plasmon resonance, we identified seven amino acids of hIL-18BP which, when changed individually to alanine, caused an 8-750-fold decrease in binding affinity, largely because of increased off-rates. These seven amino acids localized to the predicted beta-strand c and d of hIL-18BP immunoglobulin-like domain, and most had hydrophobic side chains. Just two amino acids, tyrosine 97 and phenylalanine 104, contributed approximately 50% of the binding free energy. Information obtained from these studies could contribute to the design of molecular antagonists of IL-18 for treatment of inflammatory diseases. PMID- 11278525 TI - Replication protein A as a "fidelity clamp" for DNA polymerase alpha. AB - The current view of DNA replication in eukaryotes predicts that DNA polymerase alpha (pol alpha)-primase synthesizes the first 10-ribonucleotide-long RNA primer on the leading strand and at the beginning of each Okazaki fragment on the lagging strand. Subsequently, pol alpha elongates such an RNA primer by incorporating about 20 deoxynucleotides. pol alpha displays a low processivity and, because of the lack of an intrinsic or associated 3'--> 5' exonuclease activity, it is more error-prone than other replicative pols. Synthesis of the RNA/DNA primer catalyzed by pol alpha-primase is a critical step in the initiation of DNA synthesis, but little is known about the role of the DNA replication accessory proteins in its regulation. In this paper we provide evidences that the single-stranded DNA-binding protein, replication protein A (RP A), acts as an auxiliary factor for pol alpha playing a dual role: (i) it stabilizes the pol alpha/primer complex, thus acting as a pol clamp; and (ii) it significantly reduces the misincorporation efficiency by pol alpha. Based on these results, we propose a hypothetical model in which RP-A is involved in the regulation of the early events of DNA synthesis by acting as a "fidelity clamp" for pol alpha. PMID- 11278526 TI - Conformation of a purified "spontaneously" inserting thylakoid membrane protein precursor in aqueous solvent and detergent micelles. AB - Subunit W of photosystem II (PsbW) is a single-span thylakoid membrane protein that is synthesized with a cleavable hydrophobic signal peptide and integrated into the thylakoid membrane by an apparently spontaneous mechanism. In this study, we have analyzed the secondary structure of the pre-protein at early stages of the insertion pathway, using purified recombinant pre-PsbW. We show that the protein remains soluble in Tris buffer after removal of detergent. Under these conditions pre-PsbW contains no detectable alpha-helix, whereas substantial alpha-helical structure is present in SDS micelles. In aqueous buffer, the tryptophan fluorescence emission characteristics are intermediate between those of solvent-exposed and hydrophobic environments, suggesting the formation of a partially folded structure. If denaturants are excluded from the purification protocol, pre-PsbW purifies instead as a 180-kDa oligomer with substantial alpha helical structure. Mature-size PsbW was prepared by removal of the presequence, and we show that this protein also contains alpha-helix in detergent but in lower quantities than the pre-protein. We therefore propose that pre-PsbW contains alpha-helical structure in both the mature protein and the signal peptide in nonpolar environments. We propose that pre-PsbW acquires its alpha-helical structure only during the later, membrane-bound stages of the insertion pathway, after which it forms a "helical hairpin"-type loop intermediate in the thylakoid membrane. PMID- 11278527 TI - Glycoprotein quality control in the endoplasmic reticulum. Mannose trimming by endoplasmic reticulum mannosidase I times the proteasomal degradation of unassembled immunoglobulin subunits. AB - Quality control in the endoplasmic reticulum must discriminate nascent proteins in their folding process from terminally unfolded molecules, selectively degrading the latter. Unassembled Ig-mu and J chains, two glycoproteins with five N-linked glycans and one N-linked glycan, respectively, are degraded by cytosolic proteasomes after a lag from synthesis, during which glycan trimming occurs. Inhibitors of mannosidase I (kifunensine), but not of mannosidase II (swainsonine), prevent the degradation of mu chains. Kifunensine also inhibits J chain dislocation and degradation, without inhibiting secretion of IgM polymers. In contrast, glucosidase inhibitors do not significantly affect the kinetics of mu and J degradation. These results suggest that removal of the terminal mannose from the central branch acts as a timer in dictating the degradation of transport incompetent, glycosylated Ig subunits in a calnexin-independent way. Kifunensine does not inhibit the degradation of an unglycosylated substrate (lambda Ig light chains) or of chimeric mu chains extended with the transmembrane region of the alpha T cell receptor chain, implying the existence of additional pathways for extracting proteins from the endoplasmic reticulum lumen for proteasomal degradation. PMID- 11278529 TI - Folding intermediates of a model three-helix bundle protein. Pressure and cold denaturation studies. AB - The stability and equilibrium unfolding of a model three-helix bundle protein, alpha(3)-1, by guanidine hydrochloride (GdnHCl), hydrostatic pressure, and temperature have been investigated. The combined use of these denaturing agents allowed detection of two partially folded states of alpha(3)-1, as monitored by circular dichroism, intrinsic fluorescence emission, and fluorescence of the hydrophobic probe bis-ANS (4,4'-dianilino-1,1'-binaphthyl-5,5'-disulfonic acid). The overall free-energy change for complete unfolding of alpha(3)-1, determined from GdnHCl unfolding data, is +4.6 kcal/mol. The native state is stabilized by 1.4 kcal/mol relative to a partially folded pressure-denatured intermediate (I(1)). Cold denaturation at high pressure gives rise to a second partially (un)folded conformation (I(2)), suggesting a significant contribution of hydrophobic interactions to the stability of alpha(3)-1. The free energy of stabilization of the native-like state relative to I(2) is evaluated to be -2.5 kcal/mol. Bis-ANS binding to the pressure- and cold-denatured states indicates the existence of significant residual hydrophobic structure in the partially (un)folded states of alpha(3)-1. The demonstration of folding intermediates of alpha(3)-1 lends experimental support to a number of recent protein folding simulation studies of other three-helix bundle proteins that predicted the existence of such intermediates. The results are discussed in terms of the significance of de novo designed proteins for protein folding studies. PMID- 11278528 TI - Cotranscription and intergenic splicing of human P2Y11 and SSF1 genes. AB - The P2Y(11) receptor is an ATP receptor positively coupled to the cAMP and phosphoinositide pathways. Ssf1 is a Saccharomyces cerevisiae nuclear protein, which plays an important role in mating. The gene encoding the human orthologue of SSF1 is adjacent to the P2Y(11) gene on chromosome 19. During the screening of placenta cDNA libraries, we isolated a chimeric clone resulting from the intergenic splicing between the P2Y(11) and SSF1 genes. The fusion protein was stably expressed in CHO-K1 cells where it generated a cAMP response to ATP qualitatively indistinguishable from that of the P2Y(11) receptor. According to both Western blotting and cAMP response, the expression of the fusion protein in the transfected cells was clearly lower than that of the P2Y(11) receptor. Both P2Y(11) and SSF1 probes detected a 5.6-kb messenger RNA with a similar pattern of intensity in each of 11 human tissues. The ubiquitous presence of chimeric transcripts and their up-regulation during granulocytic differentiation indicate that the transgenic splicing between the P2Y(11) and the SSF1 genes is a common and regulated phenomenon. There are very few examples of intergenic splicing in mammalian cells, and this is the first case involving a G-protein-coupled receptor. PMID- 11278530 TI - Unusually stable and long-lived ligand-induced conformations of integrins. AB - Integrins are a large family of cell surface receptors that are involved in a wide range of biological processes. The integrin alpha(IIb)beta3 (glycoprotein IIb-IIIa) is a major platelet glycoprotein heterodimeric receptor that mediates platelet aggregation and is currently a target for pharmaceutical intervention. Ligand binding to the receptor has been shown to induce conformational changes by physical methods and the exposure of neoepitopes (the ligand-induced binding sites). Here we show that the antagonist XP280 induces a conformation that is stable to treatment with SDS and that the protein retains this conformation for several days even after dissociation of the inhibitor. These ligand-induced conformational changes take place with purified protein and on intact platelets. They are competable with an RGDS peptide and are stable to reduction but not boiling or treatment with EDTA. The retention of an altered conformation in the absence of the ligand implies the possibility of ligand-induced alteration of biological function even in the absence of ligand. Finally, similar behavior is observed with the integrin alpha(v)beta3, suggesting that access to SDS stable conformations may be conserved throughout the integrin superfamily. The unusual stability, long-lived nature, and potential generality of these conformations could have profound implications for integrin biology. PMID- 11278531 TI - Cyclopentenone prostaglandins as potential inducers of intracellular oxidative stress. AB - In the present study, we find that cyclopentenone prostaglandins (PGs) of the J(2) series, naturally occurring derivatives of PGD(2), are potential inducers of intracellular oxidative stress that mediates cell degeneration. Based on an extensive screening of diverse chemical agents on induction of intracellular production of reactive oxygen species (ROS), we found that the cyclopentenone PGs, such as PGA(2), PGJ(2), Delta(12)-PGJ(2), and 15-deoxy-Delta(12,14)-PGJ(2), showed the most potent pro-oxidant effect on SH-SY5Y human neuroblastoma cells. As the intracellular events that mediate the PG cytotoxicity, we observed (i) the cellular redox alteration represented by depletion of antioxidant defenses, such as glutathione and glutathione peroxidase; (ii) a transient decrease in the mitochondrial membrane potential (Deltapsi); (iii) the production of protein bound lipid peroxidation products, such as acrolein and 4-hydroxy-2-nonenal; and (iv) the accumulation of ubiquitinated proteins. These events correlated well with the reduction in cell viability. In addition, the thiol compound, N acetylcysteine, could significantly inhibit the PG-induced ROS production, thereby preventing cytotoxicity, suggesting that the redox alteration is closely related to the pro-oxidant effect of cyclopentenone PGs. More strikingly, the lipid peroxidation end products, acrolein and 4-hydroxy-2-nonenal, detected in the PG-treated cells potently induced the ROS production, which was accompanied by the accumulation of ubiquitinated proteins and cell death, suggesting that the membrane lipid peroxidation products may represent one of the causative factors that potentiate the cytotoxic effect of cyclopentenone PGs by accelerating intracellular oxidative stress. These data suggest that the intracellular oxidative stress, represented by ROS production/lipid peroxidation and redox alteration, may underlie the well documented biological effects, such as antiproliferative and antitumor activities, of cyclopentenone PGs. PMID- 11278532 TI - Direct activation of cloned K(atp) channels by intracellular acidosis. AB - ATP-sensitive K(+) (K(ATP)) channels may be regulated by protons in addition to ATP, phospholipids, and other nucleotides. Such regulation allows a control of cellular excitability in conditions when pH is low but ATP concentration is normal. However, whether the K(ATP) changes its activity with pH alterations remains uncertain. In this study we showed that the reconstituted K(ATP) was strongly activated during hypercapnia and intracellular acidosis using whole-cell recordings. Further characterizations in excised patches indicated that channel activity increased with a moderate drop in intracellular pH and decreased with strong acidification. The channel activation was produced by a direct action of protons on the Kir6 subunit and relied on a histidine residue that is conserved in all K(ATP). The inhibition appeared to be a result of channel rundown and was not seen in whole-cell recordings. The biphasic response may explain the contradictory pH sensitivity observed in cell-endogenous K(ATP) in excised patches. Site-specific mutations of two residues showed that pH and ATP sensitivities were independent of each other. Thus, these results demonstrate that the proton is a potent activator of the K(ATP). The pH-dependent activation may enable the K(ATP) to control vascular tones, insulin secretion, and neuronal excitability in several pathophysiologic conditions. PMID- 11278533 TI - Direct interaction between the subunit RAP30 of transcription factor IIF (TFIIF) and RNA polymerase subunit 5, which contributes to the association between TFIIF and RNA polymerase II. AB - The general transcription factor IIF (TFIIF) assembled in the initiation complex, and RAP30 of TFIIF, have been shown to associate with RNA polymerase II (pol II), although it remains unclear which pol II subunit is responsible for the interaction. We examined whether TFIIF interacts with RNA polymerase II subunit 5 (RPB5), the exposed domain of which binds transcriptional regulatory factors such as hepatitis B virus X protein and a novel regulatory protein, RPB5-mediating protein. The results demonstrated that RPB5 directly binds RAP30 in vitro using purified recombinant proteins and in vivo in COS1 cells transiently expressing recombinant RAP30 and RPB5. The RAP30-binding region was mapped to the central region (amino acids (aa) 47-120) of RPB5, which partly overlaps the hepatitis B virus X protein-binding region. Although the middle part (aa 101-170) and the N terminus (aa 1-100) of RAP30 independently bound RPB5, the latter was not involved in the RPB5 binding when RAP30 was present in TFIIF complex. Scanning of the middle part of RAP30 by clustered alanine substitutions and then point alanine substitutions pinpointed two residues critical for the RPB5 binding in in vitro and in vivo assays. Wild type but not mutants Y124A and Q131A of RAP30 coexpressed with FLAG-RAP74 efficiently recovered endogenous RPB5 to the FLAG RAP74-bound anti-FLAG M2 resin. The recovered endogenous RPB5 is assembled in pol II as demonstrated immunologically. Interestingly, coexpression of the central region of RPB5 and wild type RAP30 inhibited recovery of endogenous pol II to the FLAG-RAP74-bound M2 resin, strongly suggesting that the RAP30-binding region of RPB5 inhibited the association of TFIIF and pol II. The exposed domain of RPB5 interacts with RAP30 of TFIIF and is important for the association between pol II and TFIIF. PMID- 11278534 TI - Cloning and characterization of a ninth member of the UDP-GalNAc:polypeptide N acetylgalactosaminyltransferase family, ppGaNTase-T9. AB - We have cloned, expressed and characterized the gene encoding a ninth member of the mammalian UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase (ppGaNTase) family, termed ppGaNTase-T9. This type II membrane protein consists of a 9-amino acid N-terminal cytoplasmic region, a 20-amino acid hydrophobic/transmembrane region, a 94-amino acid stem region, and a 480-amino acid conserved region. Northern blot analysis revealed that the gene encoding this enzyme is expressed in a broadly distributed manner across many adult tissues. Significant levels of 5- and 4.2-kilobase transcripts were found in rat sublingual gland, testis, small intestine, colon, and ovary, with lesser amounts in heart, brain, spleen, lung, stomach, cervix, and uterus. In situ hybridization to mouse embryos (embryonic day 14.5) revealed significant hybridization in the developing mandible, maxilla, intestine, and mesencephalic ventricle. Constructs expressing this gene transiently in COS7 cells resulted in no detectable transferase activity in vitro against a panel of unmodified peptides, including MUC5AC (GTTPSPVPTTSTTSAP) and EA2 (PTTDSTTPAPTTK). However, when incubated with MUC5AC and EA2 glycopeptides (obtained by the prior action of ppGaNTase-T1), additional incorporation of GalNAc was achieved, resulting in new hydroxyamino acid modification. The activity of this glycopeptide transferase is distinguished from that of ppGaNTase-T7 in that it forms a tetra-glycopeptide species from the MUC5AC tri-glycopeptide substrate, whereas ppGaNTase-T7 forms a hexa-glycopeptide species. This isoform thus represents the second example of a glycopeptide transferase and is distinct from the previously identified form in enzymatic activity as well as expression in embryonic and adult tissues. These findings lend further support to the existence of a hierarchical network of differential enzymatic activity within the diversely regulated ppGaNTase family, which may play a role in the various processes governing development. PMID- 11278535 TI - Characterization of heteromultimeric G protein-coupled inwardly rectifying potassium channels of the tunicate tadpole with a unique pore property. AB - Two cDNAs that encode the G protein-coupled inwardly rectifying K(+) channel (GIRK, Kir3) of tunicate tadpoles (tunicate G protein-coupled inwardly rectifying K(+) channel-A and -B; TuGIRK-A and -B) have been isolated. The deduced amino acid sequences showed approximately 60% identity with the mammalian Kir3 family. Detected by whole mount in situ hybridization, both TuGIRK-A and -B were expressed similarly in the neural cells of the head and neck region from the tail bud stage to the young tadpole stage. By co-injecting cRNAs of TuGIRK-A and G protein beta(1)/gamma(2) subunits (Gbetagamma) in Xenopus oocytes, an inwardly rectifying K(+) current was expressed. In contrast, coinjection of TuGIRK-B with Gbetagamma did not express any current. When both TuGIRK-A and -B were coexpressed together with Gbetagamma, an inwardly rectifying K(+) current was also detected. The properties of this current clearly differed from those of TuGIRK-A current, since it displayed a characteristic decline of the macroscopic conductance at strongly hyperpolarized potentials. TuGIRK-A/B current also differed from TuGIRK-A current in terms of the lower sensitivity to the Ba(2+) block, the higher sensitivity to the Cs(+) block, and the smaller single channel conductance. Taken together, we concluded that TuGIRK-A and -B form functional heteromultimeric G protein-coupled inwardly rectifying K(+) channels in the neural cells of the tunicate tadpole. By introducing a mutation of Lys(161) to Thr in TuGIRK-B, TuGIRK-A/B channels acquired a higher sensitivity to the Ba(2+) block and a slightly lower sensitivity to the Cs(+) block, and the decrease in the macroscopic conductance at hyperpolarized potentials was no longer observed. Thus, the differences in the electrophysiological properties between TuGIRK-A and TuGIRK-A/B channels were shown to be, at least partly, due to the presence of Lys(161) at the external mouth of the pore of the TuGIRK-B subunit. PMID- 11278536 TI - Assembly of Tom6 and Tom7 into the TOM core complex of Neurospora crassa. AB - Translocation of preproteins across the mitochondrial outer membrane is mediated by the translocase of the outer mitochondrial membrane (TOM) complex. We report the molecular identification of Tom6 and Tom7, two small subunits of the TOM core complex in the fungus Neurospora crassa. Cross-linking experiments showed that both proteins were found to be in direct contact with the major component of the pore, Tom40. In addition, Tom6 was observed to interact with Tom22 in a manner that depends on the presence of preproteins in transit. Precursors of both proteins are able to insert into the outer membrane in vitro and are assembled into authentic TOM complexes. The insertion pathway of these proteins shares a common binding site with the general import pathway as the assembly of both Tom6 and Tom7 was competed by a matrix-destined precursor protein. This assembly was dependent on the integrity of receptor components of the TOM machinery and is highly specific as in vitro-synthesized yeast Tom6 was not assembled into N. crassa TOM complex. The targeting and assembly information within the Tom6 sequence was found to be located in the transmembrane segment and a flanking segment toward the N-terminal, cytosolic side. A hybrid protein composed of the C terminal domain of yeast Tom6 and the cytosolic domain of N. crassa Tom6 was targeted to the mitochondria but was not taken up into TOM complexes. Thus, both segments are required for assembly into the TOM complex. A model for the topogenesis of the small Tom subunits is discussed. PMID- 11278537 TI - A plant gene encoding a Myb-like protein that binds telomeric GGTTTAG repeats in vitro. AB - A gene (AtTRP1) encoding a telomeric repeat-binding protein has been isolated from Arabidopsis thaliana. AtTRP1 is a single copy gene located on chromosome 5 of A. thaliana. The protein AtTRP1 encoded by this gene is not only homologous to the Myb DNA-binding motifs of other telomere-binding proteins but also is similar to several initiator-binding proteins in plants. Gel retardation assay revealed that the 115 residues on the C terminus of this protein, including the Myb motif, are sufficient for binding to the double-stranded plant telomeric sequence. The isolated DNA-binding domain of AtTRP1 recognizes each telomeric repeat centered on the sequence GGTTTAG. The almost full-length protein of AtTRP1 does not form any complex at all with the DNA fragments carrying four or fewer GGTTTAG repeats. However, it forms a complex with the sequence (GGTTTAG)(8) more efficiently than with the sequence (GGTTTAG)(5). These data suggest that the minimum length of a telomeric DNA for AtTRP1 binding consists of five GGTTTAG repeats and that the optimal AtTRP1 binding may require eight or more GGTTTAG repeats. It also implies that this protein AtTRP1 may bind in vivo primarily to the ends of plant chromosomes, which consist of long stretches of telomeric repeats. PMID- 11278538 TI - Identification of critical residues in bovine IFNAR-1 responsible for interferon binding. AB - Interferons have antiviral, antigrowth and immunomodulatory effects. The human type I interferons, IFN-alpha, IFN-beta, and IFN-omega, induce somewhat different cellular effects but act through a common receptor complex, IFNAR, composed of subunits IFNAR-1 and IFNAR-2. Human IFNAR-2 binds all type I IFNs but with lower affinity and different specificity than the IFNAR complex. Human IFNAR-1 has low intrinsic binding of human IFNs but strongly affects the affinity and differential ligand specificity of the IFNAR complex. Understanding IFNAR-1 interactions with the interferons is critical to elucidating the differential ligand specificity and activation by type I IFNs. However, studies of ligand interactions with human IFNAR-1 are compromised by its low affinity. The homologous bovine IFNAR-1 serendipitously binds human IFN-alphas with nanomolar affinity. Exploiting its strong binding of human IFN-alpha2, we have identified residues important for ligand binding. Mutagenesis of any of five aromatic residues of bovine IFNAR-1 caused strong decreases in ligand binding, whereas mutagenesis of proximal neutral or charged residues had smaller effects. These residues were mapped onto a homology model of IFNAR-1 to identify the ligand binding face of IFNAR-1, which is consistent with previous structure/function studies of human IFNAR-1. The topology of IFNAR-1/IFN interactions appears novel when compared with previously studied cytokine receptors. PMID- 11278539 TI - Copper converts the cellular prion protein into a protease-resistant species that is distinct from the scrapie isoform. AB - Several lines of evidence have suggested that copper ions play a role in the biology of both PrP(C) and PrP(Sc), the normal and pathologic forms of the prion protein. To further investigate this intriguing connection, we have analyzed how copper ions affect the biochemical properties of PrP(C) extracted from the brains of transgenic mice and from transfected cells. We report that the metal rapidly and reversibly induces PrP(C) to become protease-resistant and detergent insoluble. Although these two properties are commonly associated with PrP(Sc), we demonstrate using a conformation-dependent immunoassay that copper-treated PrP is structurally distinct from PrP(Sc). The effect of copper requires the presence of at least one of the five octapeptide repeats normally present in the N-terminal half of the protein, consistent with the idea that the metal alters the biochemical properties of PrP by directly binding to this region. These results suggest potential roles for copper in prion diseases, as well as in the physiological function of PrP(C). PMID- 11278540 TI - Mutational analysis and reconstituted expression of the biosynthetic genes involved in the formation of 3-amino-5-hydroxybenzoic acid, the starter unit of rifamycin biosynthesis in amycolatopsis Mediterranei S699. AB - To investigate a novel branch of the shikimate biosynthesis pathway operating in the formation of 3-amino-5-hydroxybenzoic acid (AHBA), the unique biosynthetic precursor of rifamycin and related ansamycins, a series of target-directed mutations and heterologous gene expressions were investigated in Amycolatopsis mediterranei and Streptomyces coelicolor. The genes involved in AHBA formation were inactivated individually, and the resulting mutants were further examined by incubating the cell-free extracts with known intermediates of the pathway and analyzing for AHBA formation. The rifL, -M, and -N genes were shown to be involved in the step(s) from either phosphoenolpyruvate/d-erythrose 4-phosphate or other precursors to 3,4-dideoxy-4-amino-d-arabino-heptulosonate 7-phosphate. The gene products of the rifH, -G, and -J genes resemble enzymes involved in the shikimate biosynthesis pathway (August, P. R., Tang, L., Yoon, Y. J., Ning, S., Muller, R., Yu, T.-W., Taylor, M., Hoffmann, D., Kim, C.-G., Zhang, X., Hutchinson, C. R., and Floss, H. G. (1998) Chem. Biol. 5, 69-79). Mutants of the rifH and -J genes produced rifamycin B at 1% and 10%, respectively, of the yields of the wild type; inactivation of the rifG gene did not affect rifamycin production significantly. Finally, coexpressing the rifG-N and -J genes in S. coelicolor YU105 under the control of the act promoter led to significant production of AHBA in the fermented cultures, confirming that seven of these genes are indeed necessary and sufficient for AHBA formation. The effects of deletion of individual genes from the heterologous expression cassette on AHBA formation duplicated the effects of the genomic rifG-N and -J mutations on rifamycin production, indicating that all these genes encode proteins with catalytic rather than regulatory functions in AHBA formation for rifamycin biosynthesis by A. mediterranei. PMID- 11278541 TI - Mitotic reorganization of the intermediate filament protein nestin involves phosphorylation by cdc2 kinase. AB - The intermediate filament protein nestin is expressed during early stages of development in the central nervous system and in muscle tissues. Nestin expression is associated with morphologically dynamic cells, such as dividing and migrating cells. However, little is known about regulation of nestin during these cellular processes. We have characterized the phosphorylation-based regulation of nestin during different stages of the cell cycle in a neuronal progenitor cell line, ST15A. Confocal microscopy of nestin organization and (32)P in vivo labeling studies show that the mitotic reorganization of nestin is accompanied by elevated phosphorylation of nestin. The phosphorylation-induced alterations in nestin organization during mitosis in ST15A cells are associated with partial disassembly of nestin filaments. Comparative in vitro and in vivo phosphorylation studies identified cdc2 as the primary mitotic kinase and Thr(316) as a cdc2 specific phosphorylation site on nestin. We generated a phosphospecific nestin antibody recognizing the phosphorylated form of this site. By using this antibody we observed that nestin shows constitutive phosphorylation at Thr(316), which is increased during mitosis. This study shows that nestin is reorganized during mitosis and that cdc2-mediated phosphorylation is an important regulator of nestin organization and dynamics during mitosis. PMID- 11278542 TI - Role of the Lewis(x) glycan determinant in corneal epithelial cell adhesion and differentiation. AB - In this study we demonstrate that in corneal epithelium there is cell-cell contact-regulated expression of a 145-kDa glycoprotein (GP) bearing the glycan determinant Lewis(x) (Le(x)) (Galbeta(1,4)[Fucalpha(1,3)]GlcNAc). This glycoprotein (Le(x)-GP) was expressed in confluent/contact-inhibited cultures but not in sparse cultures of corneal epithelium. In contrast, a 135-kDa glycoprotein bearing precursor, unfucosylated, lactosamine-containing glycans (Galbeta1 4GlcNAcbeta1-R) was expressed in sparse cultures. Immunofluorescence staining and confocal microscopy of confluent cultures revealed that in corneal epithelium, Le(x) antigen is located in high density at sites of cell-cell adhesion. In in vitro cell-cell adhesion assays, anti-Le(x), but not anti-sialyl-Le(x) monoclonal antibodies, inhibited the formation of corneal epithelial cell-cell adhesion. Also, when added to confluent cultures, antibodies to Le(x) disrupted the monolayer and caused tightly packed polygonal cells to round up. Analysis of the expression of Fut genes that encode alpha-1,3-fucosyltransferases, the enzymes that generate the Le(x) determinant, revealed that confluent/contact-inhibited cultures of rabbit corneal epithelium contain markedly elevated levels of Fut4 and Fut3/5/6 gene transcripts compared with sparse cultures. These data suggest that the Fut4 and Fut3/5/6 genes are targets of cell-cell contact-regulated signals and that Fut gene products direct cell-cell contact-associated expression of Le(x) on the Le(x)-GP in corneal epithelium. Immunohistochemical analysis revealed that the expression of Le(x) antigen in the epithelium of adult and developing corneas is related to the stage of differentiation of the cells. Although early differentiated cells robustly expressed Le(x), relatively undifferentiated cells did not, and the expression level was relatively low in terminally differentiated cells. Overall, these data provide evidence that a Le(x)-bearing glycoprotein plays a role through the Le(x) determinant in corneal epithelial cell-cell adhesion, and these data suggest that Le(x)-mediated cell cell interactions contribute to mechanisms that mediate corneal epithelial cell differentiation. PMID- 11278543 TI - Zonula occludens toxin structure-function analysis. Identification of the fragment biologically active on tight junctions and of the zonulin receptor binding domain. AB - Zonula occludens toxin (Zot) is an enterotoxin elaborated by Vibrio cholerae that increases intestinal permeability by interacting with a mammalian cell receptor with subsequent activation of intracellular signaling leading to the disassembly of the intercellular tight junctions. Zot localizes in the bacterial outer membrane of V. cholerae with subsequent cleavage and secretion of a carboxyl terminal fragment in the host intestinal milieu. To identify the Zot domain(s) directly involved in the protein permeating effect, several zot gene deletion mutants were constructed and tested for their biological activity in the Ussing chamber assay and their ability to bind to the target receptor on intestinal epithelial cell cultures. The Zot biologically active domain was localized toward the carboxyl terminus of the protein and coincided with the predicted cleavage product generated by V. cholerae. This domain shared a putative receptor-binding motif with zonulin, the Zot mammalian analogue involved in tight junction modulation. Amino acid comparison between the Zot active fragment and zonulin, combined with site-directed mutagenesis experiments, confirmed the presence of an octapeptide receptor-binding domain toward the amino terminus of the processed Zot. PMID- 11278544 TI - Interaction of p130 with, and consequent inhibition of, the catalytic subunit of protein phosphatase 1alpha. AB - The protein p130 was originally isolated from rat brain as an inositol 1,4,5 trisphosphate-binding protein with a domain organization similar to that of phospholipase C-delta1 but which lacks phospholipase C activity. Yeast two-hybrid screening of a human brain cDNA library for clones that encode proteins that interact with p130 has now led to the identification of the catalytic subunit of protein phosphatase 1alpha (PP1calpha) as a p130-binding protein. The association between p130 and PP1calpha was also confirmed in vitro by an overlay assay, a "pull-down" assay, and surface plasmon resonance analysis. The interaction of p130 with PP1calpha resulted in inhibition of the catalytic activity of the latter in a p130 concentration-dependent manner. Immunoprecipitation and immunoblot analysis of COS-1 cells that stably express p130 and of mouse brain extract with antibodies to p130 and to PP1calpha also detected the presence of a complex of p130 and PP1calpha. The activity of glycogen phosphorylase, which is negatively regulated by dephosphorylation by PP1calpha, was higher in COS-1 cells that stably express p130 than in control COS-1 cells. These results suggest that, in addition to its role in inositol 1,4,5-trisphosphate and Ca(2+) signaling, p130 might also contribute to regulation of protein dephosphorylation through its interaction with PP1calpha. PMID- 11278545 TI - Differentiated 3T3L1 adipocytes are composed of heterogenous cell populations with distinct receptor tyrosine kinase signaling properties. AB - Various studies have demonstrated that the platelet-derived growth factor (PDGF) receptor in adipocytes can activate PI 3-kinase activity without affecting insulin-responsive glucose transporter (GLUT4) translocation. To investigate this phenomenon of receptor signaling specificity, we utilized single cell analysis to determine the cellular distribution and signaling properties of PDGF and insulin in differentiated 3T3L1 adipocytes. The insulin receptor was highly expressed in a large percentage of the cell population (>95%) that also expressed caveolin 2 and GLUT4 with very low levels of the PDGF receptor. In contrast, the PDGF receptor was only expressed in approximately 10% of the differentiated 3T3L1 cell population with relatively low levels of the insulin receptor, caveolin 2, and GLUT4. Consistent with this observation, insulin stimulated the phosphorylation of Akt in the caveolin 2- and GLUT4-positive cells, whereas PDGF primarily stimulated Akt phosphorylation in the caveolin 2- and GLUT4-negative cell population. Furthermore, transfection of the PDGF receptor in the insulin receptor-, GLUT4-, and caveolin 2-positive cells resulted in the ability of PDGF to stimulate GLUT4 translocation. These data demonstrate that differentiated 3T3L1 adipocytes are not a homogeneous population of cells, and the lack of PDGF receptor expression in the GLUT4-positive cell population accounts for the inability of the endogenous PDGF receptor to activate GLUT4 translocation. PMID- 11278546 TI - Ca2+ release through ryanodine receptors regulates skeletal muscle L-type Ca2+ channel expression. AB - Skeletal muscle obtained from mice that lack the type 1 ryanodine receptor (RyR 1), termed dyspedic mice, exhibit a 2-fold reduction in the number of dihydropyridine binding sites (DHPRs) compared with skeletal muscle obtained from wild-type mice (Buck, E. D., Nguyen, H. T., Pessah, I. N., and Allen, P. D. (1997) J. Biol. Chem. 272, 7360-7367 and Fleig, A., Takeshima, H., and Penner, R. (1996) J. Physiol. (Lond.) 496, 339-345). To probe the role of RyR-1 in influencing L-type Ca(2+) channel (L-channel) expression, we have monitored functional L-channel expression in the sarcolemma using the whole-cell patch clamp technique in normal, dyspedic, and RyR-1-expressing dyspedic myotubes. Our results indicate that dyspedic myotubes exhibit a 45% reduction in maximum immobilization-resistant charge movement (Q(max)) and a 90% reduction in peak Ca(2+) current density. Calcium current density was significantly increased in dyspedic myotubes 3 days after injection of cDNA encoding either wild-type RyR-1 or E4032A, a mutant RyR-1 that is unable to restore robust voltage-activated release of Ca(2+) from the sarcoplasmic reticulum (SR) following expression in dyspedic myotubes (O'Brien, J. J., Allen, P. D., Beam, K., and Chen, S. R. W. (1999) Biophys. J. 76, A302 (abstr.)). The increase in L-current density 3 days after expression of either RyR-1 or E4032A occurred in the absence of a change in Q(max). However, Q(max) was increased 85% 6 days after injection of dyspedic myotubes with cDNA encoding the wild-type RyR-1 but not E4032A. Because normal and dyspedic myotubes exhibited a similar density of T-type Ca(2+) current (T current), the presence of RyR-1 does not appear to cause a general overall increase in protein synthesis. Thus, long-term expression of L-channels in skeletal myotubes is promoted by Ca(2+) released through RyRs occurring either spontaneously or during excitation-contraction coupling. PMID- 11278547 TI - A functional interaction with CBP contributes to transcriptional activation by the Wilms tumor suppressor WT1. AB - The Wilms tumor gene WT1 encodes a zinc finger transcription factor that is required for normal kidney development. WT1 was identified as a transcriptional repressor, based on its suppression of promoter reporters, but analysis of native transcripts using high density microarrays has uncovered transcriptional activation, rather than repression, of potential target genes. We report here that WT1 binds to the transcriptional coactivator CBP, leading to synergistic activation of a physiologically relevant promoter. The physical interaction between WT1 and CBP is evident in vitro and in vivo, and the two proteins are co immunoprecipitated from embryonic rat kidney cells. The WT1-CBP association requires the first two zinc fingers of WT1 and the adenovirus 5 E1A-binding domain of CBP. Overexpression of this domain of CBP is sufficient to inhibit WT1 mediated transcriptional activation of a promoter reporter, as is co-transfection of E1A. Retrovirally driven expression of either the CBP fragment or of E1A in human hematopoietic cells suppresses the induction by WT1 of its endogenous target gene, p21(Cip1). These observations support a model of WT1 as a transcriptional activator of genes required for cellular differentiation. PMID- 11278548 TI - Prostaglandin E2 increases growth and motility of colorectal carcinoma cells. AB - Chronic use of nonsteroidal anti-inflammatory drugs results in a significant reduction of risk and mortality from colorectal cancer in humans. All of the mechanism(s) by which nonsteroidal anti-inflammatory drugs exert their protective effects are not completely understood, but they are known to inhibit cyclooxygenase activity. The cyclooxygenase enzymes catalyze a key reaction in the conversion of arachidonic acid to prostaglandins, such as prostaglandin E(2) (PGE(2)). Here we demonstrate that PGE(2) treatment of LS-174 human colorectal carcinoma cells leads to increased motility and changes in cell shape. The prostaglandin EP(4) receptor signaling pathway appears to play a role in transducing signals which regulate these effects. PGE(2) treatment results in an activation of phosphatidylinositol 3-kinase/protein kinase B pathway that is required for the PGE(2)-induced changes in carcinoma cell motility and colony morphology. Our results suggest that PGE(2) might enhance the invasive potential of colorectal carcinoma cells via activation of major intracellular signal transduction pathways not previously reported to be regulated by prostaglandins. PMID- 11278549 TI - Trax (translin-associated factor X), a primarily cytoplasmic protein, inhibits the binding of TB-RBP (translin) to RNA. AB - Trax (Translin-associated factor X) has been shown to interact with TB RBP/Translin by its coimmunoprecipitation and in yeast two-hybrid assays. Here we demonstrate that Trax is widely expressed, does not bind to DNA or RNA, but forms heterodimers with TB-RBP under reducing conditions. The heterodimer of TB-RBP and Trax inhibits TB-RBP binding to RNA, but enhances TB-RBP binding to specific single stranded DNA sequences. The in vitro interactions between TB-RBP and Trax are confirmed by similar interactions in the yeast two-hybrid system. Cell fractionation and confocal microscope studies reveal that Trax is predominantly cytoplasmic. In contrast, TB-RBP is present in both the nuclei and cytoplasm of transfected cells and uses a highly conserved nuclear export signal to exit nuclei. In addition to a leucine zipper, two basic domains in TB-RBP are essential for RNA binding, but only one of these domains is needed for DNA binding. Trax restores DNA binding to TB-RBP containing an altered form of this domain. These data suggest that Trax-TB.RBP interactions modulate the DNA- and RNA-binding activity of TB-RBP. PMID- 11278550 TI - A family of AP-2 proteins down-regulate manganese superoxide dismutase expression. AB - Manganese superoxide dismutase (Mn-SOD) is a primary antioxidant enzyme whose expression is essential for life in oxygen. Mn-SOD has tumor suppressor activity in a wide variety of tumors and transformed cell systems. Our initial observations revealed that Mn-SOD expression was inversely correlated with expression of AP-2 transcription factors in normal human fibroblasts and their SV 40 transformed counterparts. Thus we hypothesized that AP-2 may down-regulate Mn SOD expression. To examine the functional role of AP-2 on Mn-SOD promoter transactivation we cotransfected AP-2-deficient HepG2 cells with a human Mn-SOD promoter-reporter construct and expression vectors encoding each of the three known AP-2 family members. Our results indicated that AP-2 could significantly repress Mn-SOD promoter activity, and that this repression was both Mn-SOD promoter and AP-2-specific. The three AP-2 proteins appeared to play distinct roles in Mn-SOD gene regulation. Moreover, although all three AP-2 proteins could repress the Mn-SOD promoter, AP-2alpha and AP-2gamma were more active in this regard than AP-2beta. Transcriptional repression by AP-2 was not a general effect in this system, because another AP-2-responsive gene, c-erbB-3, was transactivated by AP-2. Repression of Mn-SOD by AP-2 was dependent on DNA binding, and expression of AP-2B, a dominant negative incapable of DNA binding, relieved the repression on Mn-SOD promoter and reactivated Mn-SOD expression in the AP-2 abundant SV40-transformed fibroblast cell line MRC-5VA. These results indicate that AP-2-mediated transcriptional repression contributes to the constitutively low expression of Mn-SOD in SV40-transformed fibroblasts and suggest a mechanism for Mn-SOD down-regulation in cancer. PMID- 11278552 TI - Enhanced mu-opioid responses in the spinal cord of mice lacking protein kinase Cgamma isoform. AB - The protein kinase C (PKC)gamma isoform is a major pool of the PKC family in the mammalian spinal cord. PKCgamma is distributed strategically in the superficial layers of the dorsal horn and, thus, may serve as an important biochemical substrate in sensory signal processing including pain. Here we report that mu opioid receptor-mediated analgesia/antinociception and activation of G-proteins in the spinal cord are enhanced in PKCgamma knockout mice. In contrast, delta- and kappa-opioidergic and ORL-1 receptor-mediated activation of G-proteins in PKCgamma knockout mice was not altered significantly relative to the wild-type mice. Deletion of PKCgamma had no significant effect on the mRNA product of spinal mu-opioid receptors but caused an increase of maximal binding of the mu opioid receptor agonist [3H][d-Ala(2),N-Me-Phe(4),Gly(5)-ol]enkephalin in spinal cord membranes obtained from PKCgamma knockout mice. These findings suggest that deletion of PKCgamma genes protects the functional mu-opioid receptors from degradation by phosphorylation. More importantly the present data provide direct evidence that PKCgamma constitutes an essential pathway through which phosphorylation of mu-opioid receptors occurs. PMID- 11278551 TI - Characterization of a rat neuronal nicotinic acetylcholine receptor alpha7 promoter. AB - Neuronal nicotinic acetylcholine receptors (nAChRs) containing the alpha7 subunit are expressed in the central nervous system, autonomic nervous system, retina, adrenal medulla, and PC12 cells. alpha7 nAChRs have been implicated in several important biological activities apart from synaptic transmission such as mediating neurite growth and presynaptic control of neurotransmitter release. A 178-base pair promoter was sufficient to drive high level expression of the alpha7 gene in PC12 cells. The alpha7 promoter was also cell-specific, expressing in PC12 cells but not in L6 rat muscle cells. Within our minimal rat alpha7 nAChR promoter we identified two sequences important for basal level expression. Mutation of a GC-rich sequence at -172 relative to the translational start site led to an increase in activity of the promoter, indicating the presence of a negative regulatory element. Upstream stimulatory factor-1 acted to regulate alpha7 expression positively by binding to an E-box at -116. A site directly adjacent to the upstream stimulatory factor-1 binding site was shown to bind Egr 1. Sp1 and Sp3 binding also occurred downstream from or overlapping the Egr-1 binding site in the rat alpha7 promoter. Several transcription factors interact in close proximity to control expression of the rat alpha7 nicotinic receptor gene. PMID- 11278553 TI - NCK and PAK participate in the signaling pathway by which vascular endothelial growth factor stimulates the assembly of focal adhesions. AB - Vascular endothelial growth factor (VEGF)-induced endothelial cell migration is a key step in the angiogenic response and is mediated, in part, by an accelerated rate of focal adhesion complex assembly and disassembly. We investigated the signaling pathway by which VEGF regulates focal adhesion complex assembly by examining the signaling proteins involved. VEGF stimulated the tyrosine phosphorylation of the SH2 domain-containing signaling proteins NCK and CRK in human umbilical vein endothelial cells. The signaling pathways that couple the kinase insert domain-containing receptor to NCK and CRK is most likely mediated by another cellular protein, as NCK and CRK were tyrosine-phosphorylated in response to VEGF in cells expressing receptors mutated at each of several candidate SH2 domain-interacting cytosolic tyrosines. In the absence of VEGF treatment, NCK (but not CRK) associated with the p21 GTPase-activated kinase PAK. PAK catalytic activity was augmented after VEGF treatment; an association of PAK with 60- and 90-kDa tyrosine-phosphorylated proteins accompanied this. VEGF stimulated the recruitment of PAK to focal adhesions, and FAK immunoprecipitated with both NCK and PAK in VEGF-treated (but not untreated) human umbilical vein endothelial cells. Inhibition of NCK protein expression using antisense oligonucleotides led to the inhibition of both VEGF-induced focal adhesion assembly and VEGF-induced cell migration, demonstrating a necessary role of NCK in these cellular responses. PMID- 11278555 TI - Caspase remodeling of the spectrin membrane skeleton during lens development and aging. AB - Terminal differentiation of lens fiber cells resembles the apoptotic process in that organelles are lost, DNA is fragmented, and changes in membrane morphology occur. However, unlike classically apoptotic cells, which are disintegrated by membrane blebbing and vesiculation, aging lens fiber cells are compressed into the center of the lens, where they undergo cell-cell fusion and the formation of specialized membrane interdigitations. In classically apoptotic cells, caspase cleavage of the cytoskeletal protein alpha-spectrin to approximately 150-kDa fragments is believed to be important for membrane blebbing. We report that caspase(s) cleave alpha-spectrin to approximately 150-kDa fragments and beta spectrin to approximately 120- and approximately 80-kDa fragments during late embryonic chick lens development. These fragments continue to accumulate with age so that in the oldest fiber cells of the adult lens, most, if not all, of the spectrin is cleaved to discrete fragments. Thus, unlike classical apoptosis, where caspase-cleaved spectrin is short lived, lens fiber cells contain spectrin fragments that appear to be stable for the lifetime of the organism. Moreover, fragmentation of spectrin results in reduced membrane association and thus may lead to permanent remodeling of the membrane skeleton. Partial and specific proteolysis of membrane skeleton components by caspases may be important for age related membrane changes in the lens. PMID- 11278554 TI - Water immersion stress prevents caerulein-induced pancreatic acinar cell nf-kappa b activation by attenuating caerulein-induced intracellular Ca2+ changes. AB - Prior stress ameliorates caerulein-induced pancreatitis in rats. NF-kappaB is a proinflammatory transcription factor activated during caerulein pancreatitis. However, the effects of prior stress on pancreatic NF-kappaB activation are unknown. In the current study, the effect of prior water immersion stress on caerulein and tumor necrosis factor-alpha (TNF-alpha)-induced NF-kappaB activation in the pancreas was evaluated. Water immersion of rats for up to 6 h prevents supramaximal caerulein-induced pancreatic IkappaB-alpha degradation and NF-kappaB activation in vivo. NF-kappaB activity is also inhibited in vitro in pancreatic acini prepared from water-immersed animals. TNF-alpha-induced NF kappaB activation in pancreas or in pancreatic acini is unaffected by prior water immersion. Chelation of intracellular Ca(2+) by 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetate/acetoxymethyl ester has similar effects to water immersion in preventing caerulein but not TNF-alpha-induced NF-kappaB activation in pancreas. Both the spike response and the sustained rise in [Ca(2+)](i) in response to supramaximal caerulein stimulation are reduced markedly in acini prepared from water-immersed animals as compared with normal animals. Our findings indicate that, in addition to Ca(2+)-dependent mechanisms, Ca(2+) independent signaling events also may lead to NF-kappaB activation in pancreatic acinar cells. Water immersion stress prevents supramaximal caerulein-induced NF kappaB activation in pancreas in vivo and in vitro by affecting intracellular Ca(2+) homeostasis. PMID- 11278556 TI - Lipid phosphate phosphatases in Arabidopsis. Regulation of the AtLPP1 gene in response to stress. AB - An Arabidopsis thaliana gene (AtLPP1) was isolated on the basis that it was transiently induced by ionizing radiation. The putative AtLPP1 gene product showed homology to the yeast and mammalian lipid phosphate phosphatase enzymes and possessed a phosphatase signature sequence motif. Heterologous expression and biochemical characterization of the AtLPP1 gene in yeast showed that it encoded an enzyme (AtLpp1p) that exhibited both diacylglycerol pyrophosphate phosphatase and phosphatidate phosphatase activities. Kinetic analysis indicated that diacylglycerol pyrophosphate was the preferred substrate for AtLpp1p in vitro. A second Arabidopsis gene (AtLPP2) was identified based on sequence homology to AtLPP1 that was also heterologously expressed in yeast. The AtLpp2p enzyme also utilized diacylglycerol pyrophosphate and phosphatidate but with no preference for either substrate. The AtLpp1p and AtLpp2p enzymes showed differences in their apparent affinities for diacylglycerol pyrophosphate and phosphatidate as well as other enzymological properties. Northern blot analyses showed that the AtLPP1 gene was preferentially expressed in leaves and roots, whereas the AtLPP2 gene was expressed in all tissues examined. AtLPP1, but not AtLPP2, was regulated in response to various stress conditions. The AtLPP1 gene was transiently induced by genotoxic stress (gamma ray or UV-B) and elicitor treatments with mastoparan and harpin. The regulation of the AtLPP1 gene in response to stress was consistent with the hypothesis that its encoded lipid phosphate phosphatase enzyme may attenuate the signaling functions of phosphatidate and/or diacylglycerol pyrophosphate that form in response to stress in plants. PMID- 11278557 TI - The missing link in coronavirus assembly. Retention of the avian coronavirus infectious bronchitis virus envelope protein in the pre-Golgi compartments and physical interaction between the envelope and membrane proteins. AB - One missing link in the coronavirus assembly is the physical interaction between two crucial structural proteins, the membrane (M) and envelope (E) proteins. In this study, we demonstrate that the coronavirus infectious bronchitis virus E can physically interact, via a putative peripheral domain, with M. Deletion of this domain resulted in a drastic reduction in the incorporation of M into virus-like particles. Immunofluorescent staining of cells coexpressing M and E supports that E interacts with M and relocates M to the same subcellular compartments that E resides in. E was retained in the pre-Golgi membranes, prior to being translocated to the Golgi apparatus and the secretory vesicles; M was observed to exhibit similar localization and translocation profiles as E when coexpressed with E. Deletion studies identified the C-terminal 6-residue RDKLYS as the endoplasmic reticulum retention signal of E, and site-directed mutagenesis of the -4 lysine residue to glutamine resulted in the accumulation of E in the Golgi apparatus. The third domain of E that plays a crucial role in virus budding is a putative transmembrane domain present at the N-terminal region, because deletion of the domain resulted in a free distribution of the mutant protein and in dysfunctional viral assembly. PMID- 11278558 TI - Role of p53 in HER2-induced proliferation or apoptosis. AB - HER2 oncogene overexpression has been associated either with proliferation or differentiation and apoptosis. The role of p53 on these different chances was investigated. Wild type (wt) p53-IGROV1 cells showed growth inhibition and apoptosis after HER2 transfection, whereas no anti-proliferative effect was observed in its mutated p53 sub-line unless wt p53 was cotransfected with HER2. Stable HER2 transfectants derived from wt p53 line treated with heregulin-beta1 or epidermal growth factor showed a decrease in proliferation due to a G(2)/M cell cycle block despite normal mitogen-activated protein kinase activation. In these HER2 transfectants, c-Myc and p53 expression were increased, whereas MDM2 was dramatically down-modulated. By contrast, growth factors stimulation of HER2 transfectants with mutated-p53 induced progression through the cell cycle. Together, our data point to a regulatory role for p53 in HER2 signaling. PMID- 11278559 TI - Versican V1 proteolysis in human aorta in vivo occurs at the Glu441-Ala442 bond, a site that is cleaved by recombinant ADAMTS-1 and ADAMTS-4. AB - Mature human aorta contains a 70-kDa versican fragment, which reacts with a neoepitope antiserum to the C-terminal peptide sequence DPEAAE. This protein therefore appears to represent the G1 domain of versican V1 (G1-DPEAAE(441)), which has been generated in vivo by proteolytic cleavage at the Glu(441)-Ala(442) bond, within the sequence DPEAAE(441)-A(442)RRGQ. Because the equivalent aggrecan product (G1-NITEGE(341)) and brevican product (G1-EAVESE(395)) are generated by ADAMTS-mediated cleavage of the respective proteoglycans, we tested the capacity of recombinant ADAMTS-1 and ADAMTS-4 to cleave versican at Glu(441)-Ala(442). Both enzymes cleaved a recombinant versican substrate and native human versican at the Glu(441)-Ala(442) bond and the mature form of ADAMTS-4 was detected by Western analysis of extracts of aortic intima. We conclude that versican V1 proteolysis in vivo can be catalyzed by one or more members of the ADAMTS family of metalloproteinases. PMID- 11278560 TI - Modulation of oncogenic DBL activity by phosphoinositol phosphate binding to pleckstrin homology domain. AB - The Dbl family guanine nucleotide exchange factors (GEFs) contain a region of sequence similarity consisting of a catalytic Dbl homology (DH) domain in tandem with a pleckstrin homology (PH) domain. PH domains are involved in the regulated targeting of signaling molecules to plasma membranes by protein-protein and/or protein-lipid interactions. Here we show that Dbl PH domain binding to phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol 3,4,5-triphosphate results in the inhibition of Dbl GEF activity on Rho family GTPase Cdc42. Phosphatidylinositol 4,5-bisphosphate binding to the PH domain significantly inhibits the Cdc42 interactive activity of the DH domain suggesting that the DH domain is subjected to the PH domain modulation under the influence of phosphoinositides (PIPs). We generated Dbl mutants unable to interact with PIPs. These mutants retained GEF activity on Cdc42 in the presence of PIPs and showed a markedly enhanced activating potential for both Cdc42 and RhoA in vivo while displaying decreased cellular transforming activity. Immunofluorescence analysis of NIH3T3 transfectants revealed that whereas the PH domain localizes to actin stress fibers and plasma membrane, the PH mutants are no longer detectable on the plasma membrane. These results suggest that modulation of PIPs in both the GEF catalytic activity and the targeting to plasma membrane determines the outcome of the biologic activity of Dbl. PMID- 11278561 TI - Two new elastin cross-links having pyridine skeleton. Implication of ammonia in elastin cross-linking in vivo. AB - Isolation and structure analysis of two amino acids from bovine ligamentum nuchae elastin hydrolysates revealed the presence of pyridine cross-links in elastin. The structures of these amino acids were determined to have 3,4,5- and 2,3,5 trisubstituted pyridine skeletons both with three carboxylic acids and a mass of 396 (C(18)H(28)N(4)0(6)) identified as 4-(4-amino-4-carboxybutyl)-3,5-di-(3-amino 3-carboxypropyl)-pyridine and 2-(4-amino-4-carboxybutyl)-3,5-di-(3-amino-3 carboxypropyl)-pyridine. We have named these pyridine cross-links desmopyridine (DESP) and isodesmopyridine (IDP), respectively. Structure analysis of these pyridine cross-links implied that the formation of these cross-links involved the condensation reaction between ammonia and allysine. The elastin incubated with ammonium chloride showed that DESP and IDP levels increased as the allysine content decreased. DESP and IDP were measured by high pressure liquid chromatography (HPLC) with UV detection and were found in a variety of bovine tissues. The DESP/desmosine (DES) and IDP/isodesmosine (IDE) ratios in aorta elastin were higher than in other tissues. DESP and IDP contents in human aorta elastin were found to be gradually increased with age. The concentration of IDP was significantly elevated in aorta elastin of rat with chronic liver cirrhosis induced by carbon tetrachloride (mean +/- S.D.; 11.1 +/- 0.9 nmol/mg elastin) when compared with normal rats (5.9 +/- 1.5 nmol/mg elastin). Although DESP and IDP are present at only trace concentrations in the tissue elastin, these pyridine cross-links may be useful biomarkers for the aortic elastin damaged by ammonia. PMID- 11278563 TI - Identification of a novel human tankyrase through its interaction with the adaptor protein Grb14. AB - Tankyrase is an ankyrin repeat-containing poly(ADP-ribose) polymerase originally isolated as a binding partner for the telomeric protein TRF1, but recently identified as a mitogen-activated protein kinase substrate implicated in regulation of Golgi vesicle trafficking. In this study, a novel human tankyrase, designated tankyrase 2, was isolated in a yeast two-hybrid screen as a binding partner for the Src homology 2 domain-containing adaptor protein Grb14. Tankyrase 2 is a 130-kDa protein, which lacks the N-terminal histidine/proline/serine-rich region of tankyrase, but contains a corresponding ankyrin repeat region, sterile alpha motif module, and poly(ADP-ribose) polymerase homology domain. The TANKYRASE 2 gene localizes to chromosome 10q23.2 and is widely expressed, with mRNA transcripts particularly abundant in skeletal muscle and placenta. Upon subcellular fractionation, both Grb14 and tankyrase 2 associate with the low density microsome fraction, and association of these proteins in vivo can be detected by co-immunoprecipitation analysis. Deletion analyses implicate the N terminal 110 amino acids of Grb14 and ankyrin repeats 10-19 of tankyrase 2 in mediating this interaction. This study supports a role for the tankyrases in cytoplasmic signal transduction pathways and suggests that vesicle trafficking may be involved in the subcellular localization or signaling function of Grb14. PMID- 11278562 TI - The prion protein has RNA binding and chaperoning properties characteristic of nucleocapsid protein NCP7 of HIV-1. AB - Transmissible spongiform encephalopathies are fatal neurodegenerative diseases associated with the accumulation of a protease-resistant form of the prion protein (PrP). Although PrP is conserved in vertebrates, its function remains to be identified. In vitro PrP binds large nucleic acids causing the formation of nucleoprotein complexes resembling human immunodeficiency virus type 1 (HIV-1) nucleocapsid-RNA complexes and in vivo MuLV replication accelerates the scrapie infectious process, suggesting possible interactions between retroviruses and PrP. Retroviruses, including HIV-1 encode a major nucleic acid binding protein (NC protein) found within the virus where 2000 NC protein molecules coat the dimeric genome. NC is required in virus assembly and infection to chaperone RNA dimerization and packaging and in proviral DNA synthesis by reverse transcriptase (RT). In HIV-1, 5'-leader RNA/NC interactions appear to control these viral processes. This prompted us to compare and contrast the interactions of human and ovine PrP and HIV-1 NCp7 with HIV-1 5'-leader RNA. Results show that PrP has properties characteristic of NCp7 with respect to viral RNA dimerization and proviral DNA synthesis by RT. The NC-like properties of huPrP map to the N terminal region of huPrP. Interestingly, PrP localizes in the membrane and cytoplasm of PrP-expressing cells. These findings suggest that PrP is a multifunctional protein possibly participating in nucleic acid metabolism. PMID- 11278564 TI - Conditional liver-specific expression of simian virus 40 T antigen leads to regulatable development of hepatic neoplasm in transgenic mice. AB - Adaptive epigenetic changes and toxicity often accompany constitutive expression of a transgene or knockout of an endogenous gene in mice. These considerations potentially limit the usefulness of transgenic technology in studying the in vivo functions of a gene. Using conditional gene expression technology, it is possible to override such restrictions to achieve temporal and tissue-specific manipulation of gene expression in vivo. Based on the tetracycline regulatory system, we established a binary transgenic model in which the conditional expression of two transgenes, SV40 T antigen (TAg) and lacZ, can be tightly regulated in the liver by administration of tetracycline. The mouse albumin or mouse major urinary protein promoter was used to achieve liver-specific expression of the tetracycline-responsive transcriptional activator (tTA) in one set of transgenic mice. These mice were crossed with transgenic mice carrying either TAg or lacZ under the control of the tTA-regulated promoter. Analyses of mice transgenic for both tTA and TAg (or lacZ) revealed that the liver-specific expression of the transgenes could be suppressed to undetectable levels and regulated in a reversible fashion by tetracycline administration and withdrawal. Mice with tTA and TAg transgenes developed hepatocellular adenomas and hyperplasia that could be prevented by continuous tetracycline administration. Our report demonstrates the value of this binary transgenic model in studying the physiological functions of any potential genes of interest in a liver-specific manner. PMID- 11278565 TI - Structural plasticity of an invariant hydrophobic triad in the switch regions of Rab GTPases is a determinant of effector recognition. AB - Rab GTPases function as regulatory components of an evolutionarily conserved machinery that mediates docking, priming, and fusion of vesicles with intracellular membranes. We have previously shown that the active conformation of Rab3A is stabilized by a substantial hydrophobic interface between the putative conformational switch regions (Dumas, J. J., Zhu, Z., Connolly, J. L., and Lambright, D. G. (1999) Structure 7, 413-423). A triad of invariant hydrophobic residues at this switch interface (Phe-59, Trp-76, and Tyr-91) represents a major interaction determinant between the switch regions of Rab3A and the Rab3A specific effector Rabphilin3A (Ostermeier, C., and Brunger, A. T. (1999) Cell 96, 363-374). Here, we report the crystal structure of the active form of Rab5C, a prototypical endocytic Rab GTPase. As is true for Rab3A, the active conformation of Rab5C is stabilized by a hydrophobic interface between the switch regions. However, the conformation of the invariant hydrophobic triad (residues Phe-58, Trp-75, and Tyr-90 in Rab5C) is dramatically altered such that the resulting surface is noncomplementary to the switch interaction epitope of Rabphilin3A. This structural rearrangement reflects a set of nonconservative substitutions in the hydrophobic core between the central beta sheet and the alpha2 helix. These observations demonstrate that structural plasticity involving an invariant hydrophobic triad at the switch interface contributes to the mechanism by which effectors recognize distinct Rab subfamilies. Thus, the active conformation of the switch regions conveys information about the identity of a particular Rab GTPase as well as the state of the bound nucleotide. PMID- 11278566 TI - A purple acid phosphatase from sweet potato contains an antiferromagnetically coupled binuclear Fe-Mn center. AB - A purple acid phosphatase from sweet potato is the first reported example of a protein containing an enzymatically active binuclear Fe-Mn center. Multifield saturation magnetization data over a temperature range of 2 to 200 K indicates that this center is strongly antiferromagnetically coupled. Metal ion analysis shows an excess of iron over manganese. Low temperature EPR spectra reveal only resonances characteristic of high spin Fe(III) centers (Fe(III)-apo and Fe(III) Zn(II)) and adventitious Cu(II) centers. There were no resonances from either Mn(II) or binuclear Fe-Mn centers. Together with a comparison of spectral properties and sequence homologies between known purple acid phosphatases, the enzymatic and spectroscopic data strongly indicate the presence of catalytic Fe(III)-Mn(II) centers in the active site of the sweet potato enzyme. Because of the strong antiferromagnetism it is likely that the metal ions in the sweet potato enzyme are linked via a mu-oxo bridge, in contrast to other known purple acid phosphatases in which a mu-hydroxo bridge is present. Differences in metal ion composition and bridging may affect substrate specificities leading to the biological function of different purple acid phosphatases. PMID- 11278567 TI - N-linked glycosylation of the HIV type-1 gp120 envelope glycoprotein as a major determinant of CCR5 and CXCR4 coreceptor utilization. AB - The variable V1V2 and V3 regions of the human immunodeficiency virus type-1 (HIV 1) envelope glycoprotein (gp120) can influence viral coreceptor usage. To substantiate this we generated isogenic HIV-1 molecularly cloned viruses that were composed of the HxB2 envelope backbone containing the V1V2 and V3 regions from viruses isolated from a patient progressing to disease. We show that the V3 amino acid charge per se had little influence on altering the virus coreceptor phenotype. The V1V2 region and its N-linked glycosylation degree were shown to confer CXCR4 usage and provide the virus with rapid replication kinetics. Loss of an N-linked glycosylation site within the V3 region had a major influence on the virus switching from the R5 to X4 phenotype in a V3 charge-dependent manner. The loss of this V3 N-linked glycosylation site was also linked with the broadening of the coreceptor repertoire to incorporate CCR3. By comparing the amino acid sequences of primary HIV-1 isolates, we identified a strong association between high V3 charge and the loss of this V3 N-linked glycosylation site. These results demonstrate that the N-linked glycosylation pattern of the HIV-1 envelope can strongly influence viral coreceptor utilization and the R5 to X4 switch. PMID- 11278568 TI - Enterophilins, a new family of leucine zipper proteins bearing a b30.2 domain and associated with enterocyte differentiation. AB - Enterocyte terminal differentiation occurs at the crypt-villus junction through the transcriptional activation of cell-specific genes, many of which code for proteins of the brush border membrane such as intestinal alkaline phosphatase, sucrase-isomaltase, or the microvillar structural protein villin. Several studies have shown that this sharp increase in specific mRNA levels is intimately associated with arrest of cell proliferation. We isolated several clones from a guinea pig intestine cDNA library. They encode new proteins characterized by an original structure associating a carboxyl-terminal B30.2/RFP-like domain and a long leucine zipper at the amino terminus. The first member of this novel gene family codes for a 65-kDa protein termed enterophilin-1, which is specifically expressed in enterocytes before their final differentiation. Enterophilin-1 is the most abundant in the small intestine but is still present in significant amounts in colonic enterocytes. In Caco-2 cells, a similar 65-kDa protein was recognized by a specific anti-enterophilin-1 antibody, and its expression was positively correlated with cell differentiation status. In addition, transfection of HT-29 cells with enterophilin-1 full-length cDNA slightly inhibited cell growth and promoted an increase in alkaline phosphatase activity. Taken together, these data identify enterophilins as a new family of proteins associated with enterocyte differentiation. PMID- 11278569 TI - Initial steps of ferulic acid polymerization by lignin peroxidase. AB - The major products of the initial steps of ferulic acid polymerization by lignin peroxidase included three dehydrodimers resulting from beta-5' and beta beta'coupling and two trimers resulting from the addition of ferulic acid moieties to decarboxylated derivatives of beta-O-4'- and beta-5'-coupled dehydrodimers. This is the first time that trimers have been identified from peroxidase-catalyzed oxidation of ferulic acid, and their formation appears to be favored by decarboxylation of dehydrodimer intermediates. After initial oxidation, the coupling reactions appear to be determined by the chemistry of ferulic acid phenoxy radicals, regardless of the enzyme and of whether the reaction is performed in vitro or in vivo. This claim is supported by our finding that horseradish peroxidase provides a similar product profile. Furthermore, two of the dehydrodimers were the two products obtained from laccase-catalyzed oxidation (Tatsumi, K. S., Freyer, A., Minard, R. D., and Bollag, J.-M. (1994) Environ. Sci. Technol. 28, 210-215), and the most abundant dehydrodimer is the most prominent in grass cell walls (Ralph, J., Quideau, S., Grabber, J. H., and Hatfield, R. D. (1994) J. Chem. Soc. Perkin Trans. 1, 3485-3498). Our results also indicate that the dehydrodimers and trimers are further oxidized by lignin peroxidase, suggesting that they are only intermediates in the polymerization of ferulic acid. The extent of polymerization appears to be dependent on the ionization potential of formed intermediates, H(2)O(2) concentration, and, probably, enzyme stability. PMID- 11278570 TI - The H2 sensor of Ralstonia eutropha. Biochemical characteristics, spectroscopic properties, and its interaction with a histidine protein kinase. AB - Previous genetic studies have revealed a multicomponent signal transduction chain, consisting of an H(2) sensor, a histidine protein kinase, and a response regulator, which controls hydrogenase gene transcription in the proteobacterium Ralstonia eutropha. In this study, we isolated the H(2) sensor and demonstrated that the purified protein forms a complex with the histidine protein kinase. Biochemical and spectroscopic analysis revealed that the H(2) sensor is a cytoplasmic [NiFe]-hydrogenase with unique features. The H(2)-oxidizing activity was 2 orders of magnitude lower than that of standard hydrogenases and insensitive to oxygen, carbon monoxide, and acetylene. Interestingly, only H(2) production but no HD formation was detected in the D(2)/H(+) exchange assay. Fourier transform infrared data showed an active site similar to that of standard [NiFe]-hydrogenases. It is suggested that the protein environment accounts for a restricted gas diffusion and for the typical kinetic parameters of the H(2) sensor. EPR analysis demonstrated that the [4Fe-4S] clusters within the small subunit were not reduced under hydrogen even in the presence of dithionite. Optical spectra revealed the presence of a novel, redox-active, n = 2 chromophore that is reduced by H(2). The possible involvement of this chromophore in signal transduction is discussed. PMID- 11278571 TI - Genetic analysis of the Escherichia coli FtsZ.ZipA interaction in the yeast two hybrid system. Characterization of FtsZ residues essential for the interactions with ZipA and with FtsA. AB - The recruitment of ZipA to the septum by FtsZ is an early, essential step in cell division in Escherichia coli. We have used polymerase chain reaction-mediated random mutagenesis in the yeast two-hybrid system to analyze this interaction and have identified residues within a highly conserved sequence at the C terminus of FtsZ as the ZipA binding site. A search for suppressors of a mutation that causes a loss of interaction (ftsZ(D373G)) identified eight different changes at two residues within this sequence. In vitro, wild type FtsZ interacted with ZipA with a high affinity in an enzyme-linked immunosorbent assay, whereas FtsZ(D373G) failed to interact. Two mutant proteins examined restored this interaction significantly. In vivo, the alleles tested are significantly more toxic than the wild type ftsZ and cannot complement a deletion. We have shown that a fusion, which encodes the last 70 residues of FtsZ in the two-hybrid system, is sufficient for the interaction with FtsA and ZipA. However, when the wild type sequence is compared with one that encodes FtsZ(D373G), no interaction was seen with either protein. Mutations surrounding Asp-373 differentially affected the interactions of FtsZ with ZipA and FtsA, indicating that these proteins bind the C terminus of FtsZ differently. PMID- 11278572 TI - Cdc42 is a substrate for caspases and influences Fas-induced apoptosis. AB - Fas-mediated apoptosis results in the activation of caspases, which subsequently cleave cellular substrates that are essential for normal cell viability. In the present study, we show that the Ras-related GTP-binding protein Cdc42 is susceptible to caspase-catalyzed proteolysis in a number of cell lines, including NIH3T3 fibroblasts, human breast cancer cells (e.g. T47D), and COS-7 cells. Both caspase-3 and caspase-7 were able to catalyze the cleavage of Cdc42, whereas caspase-6 and caspase-8 were without effect. The susceptibility to the caspase stimulated degradation is specific; although Rac can also serve as a caspase substrate, neither Rho nor Ras is degraded. Caspase sensitivity is conferred by a consensus sequence (DXXD) that lies immediately upstream of the Rho insert regions (residues 122-134) of Cdc42 and Rac. The removal of a stretch of residues (120) that includes the insert region or site-directed mutagenesis of either aspartic acid 118 or 121 within a constitutively active background (i.e. Cdc42(F28L)) as well as a wild-type Cdc42 background yields Cdc42 molecules that provide a marked protection against Fas ligand-induced apoptosis. Overall, these results are consistent with a model in which Cdc42 acts downstream of Fas, perhaps to influence the rate of apoptosis, with the ultimate caspase-mediated degradation of Cdc42 then allowing for a maximal apoptotic response. PMID- 11278573 TI - Osteopontin gene regulation by oscillatory fluid flow via intracellular calcium mobilization and activation of mitogen-activated protein kinase in MC3T3-E1 osteoblasts. AB - Recently fluid flow has been shown to be a potent physical stimulus in the regulation of bone cell metabolism. However, most investigators have applied steady or pulsing flow profiles rather than oscillatory fluid flow, which occurs in vivo because of mechanical loading. Here oscillatory fluid flow was demonstrated to be a potentially important physical signal for loading-induced changes in bone cell metabolism. We selected three well known biological response variables including intracellular calcium (Ca(2+)i), mitogen-activated protein kinase (MAPK) activity, and osteopontin (OPN) mRNA levels to examine the response of MC3T3-E1 osteoblastic cells to oscillatory fluid flow with shear stresses ranging from 2 to -2 Newtons/m(2) at 1 Hz, which is in the range expected to occur during routine physical activities. Our results showed that within 1 min, oscillatory flow induced cell Ca(2+)i mobilization, whereas two MAPKs (ERK and p38) were activated over a 2-h time frame. However, there was no activation of JNK. Furthermore 2 h of oscillatory fluid flow increased steady-state OPN mRNA expression levels by approximately 4-fold, 24 h after exposure to fluid flow. The presence of both ERK and p38 inhibitors and thapsigargin completely abolished the effect of oscillatory flow on steady-state OPN mRNA levels. In addition, experiments using a variety of pharmacological agents suggest that oscillatory flow induces Ca(2+)i mobilization via the L-type voltage-operated calcium channel and the inositol 1,4,5-trisphosphate pathway. PMID- 11278574 TI - Plasma membrane Ca(2+)-ATPase associates with the cytoskeleton in activated platelets through a PDZ-binding domain. AB - The plasma membrane Ca(2+)-ATPase (PMCA) plays an essential role in maintaining low cytosolic Ca(2+) in resting platelets. During platelet activation PMCA is phosphorylated transiently on tyrosine residues resulting in inhibition of the pump that enhances elevation of Ca(2+). Tyrosine phosphorylation of many proteins during platelet activation results in their association with the cytoskeleton. Consequently, in the present study we asked if PMCA interacts with the platelet cytoskeleton. We observed that very little PMCA is associated with the cytoskeleton in resting platelets but that approximately 80% of total PMCA (PMCA1b + PMCA4b) is redistributed to the cytoskeleton upon activation with thrombin. Tyrosine phosphorylation of PMCA during activation was not associated with the redistribution because tyrosine-phosphorylated PMCA was not translocated specifically to the cytoskeleton. Because PMCA b-splice isoforms have C-terminal PSD-95/Dlg/ZO-1 homology domain (PDZ)-binding domains, a C-terminal peptide was used to disrupt potential PDZ domain interactions. Activation of saponin permeabilized platelets in the presence of the peptide led to a significant decrease of PMCA in the cytoskeleton. PMCA associated with the cytoskeleton retained Ca(2+)-ATPase activity. These results suggest that during activation active PMCA is recruited to the cytoskeleton by interaction with PDZ domains and that this association provides a microenvironment with a reduced Ca(2+) concentration. PMID- 11278575 TI - Phosphoinositide 3-kinase facilitates antigen-stimulated Ca(2+) influx in RBL-2H3 mast cells via a phosphatidylinositol 3,4,5-trisphosphate-sensitive Ca(2+) entry mechanism. AB - This study presents evidence that phosphoinositide 3-kinase (PI3K) plays a concerted role with phospholipase Cgamma in initiating antigen-mediated Ca(2+) signaling in mast cells via a phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P(3))-sensitive Ca(2+) entry pathway. Exogenous PI(3,4,5)P(3) at concentrations close to its physiological level induces instantaneous Ca(2+) influx into RBL-2H3 cells. This PI(3,4,5)P(3)-induced intracellular Ca(2+) increase is independent of phospholipase C activity or the depletion of internal stores. Moreover, inhibition of PI3K by LY294002 or by overexpression of the dominant negative inhibitor Deltap85 suppresses the Ca(2+) response to the cross linking of the high affinity receptor for IgE (FcepsilonRI). Concomitant treatment of RBL-2H3 cells with LY294002 or Deltap85 and 2-aminoethyl diphenylborate, a cell-permeant antagonist of D-myo-inositol 1,4,5-trisphosphate receptors, abrogates antigen-induced Ca(2+) signals, whereas either treatment alone gives rise to partial inhibition. Conceivably, PI(3,4,5)P(3)-sensitive Ca(2+) entry and capacitative Ca(2+) entry represent major Ca(2+) influx pathways that sustain elevated [Ca(2+)]i to achieve optimal physiological responses. This study also refutes the second messenger role of D-myo-inositol 1,3,4,5 tetrakisphosphate in regulating FcepsilonRI-mediated Ca(2+) response. Considering the underlying mechanism, our data suggest that PI(3,4,5)P(3) directly stimulates a Ca(2+) transport system in plasma membranes. Together, these data provide a molecular basis to account for the role of PI3K in the regulation of FcepsilonRI mediated degranulation in mast cells. PMID- 11278576 TI - Association between the 15-kDa selenoprotein and UDP-glucose:glycoprotein glucosyltransferase in the endoplasmic reticulum of mammalian cells. AB - Mammalian selenocysteine-containing proteins characterized with respect to function are involved in redox processes and exhibit distinct expression patterns and cellular locations. A recently identified 15-kDa selenoprotein (Sep15) has no homology to previously characterized proteins, and its function is not known. Here we report the intracellular localization and identification of a binding partner for this selenoprotein which implicate Sep15 in the regulation of protein folding. The native Sep15 isolated from rat prostate and mouse liver occurred in a complex with a 150-kDa protein. The latter protein was identified as UDP glucose:glycoprotein glucosyltransferase (UGTR), the endoplasmic reticulum (ER) resident protein, which was previously shown to be involved in the quality control of protein folding. UGTR functions by glucosylating misfolded proteins, retaining them in the ER until they are correctly folded or transferring them to degradation pathways. To determine the intracellular localization of Sep15, we expressed a green fluorescent protein-Sep15 fusion protein in CV-1 cells, and this protein was localized to the ER and possibly other perinuclear compartments. We determined that Sep15 contained the N-terminal signal peptide that was essential for translocation and that it was cleaved in the mature protein. However, C-terminal sequences of Sep15 were not involved in trafficking and retention of Sep15. The data suggest that the association between Sep15 and UGTR is responsible for maintaining the selenoprotein in the ER. This report provides the first example of the ER-resident selenoprotein and suggests a possible role of the trace element selenium in the quality control of protein folding. PMID- 11278577 TI - Crystal structure of a mutant hERalpha ligand-binding domain reveals key structural features for the mechanism of partial agonism. AB - The crystal structure of a triple cysteine to serine mutant ERalpha ligand binding domain (LBD), complexed with estradiol, shows that despite the presence of a tightly bound agonist ligand, the protein exhibits an antagonist-like conformation, similar to that observed in raloxifen and 4-hydroxytamoxifen-bound structures. This mutated receptor binds estradiol with wild type affinity and displays transcriptional activity upon estradiol stimulation, but with limited potency (about 50%). This partial activity is efficiently repressed in antagonist competition assays. The comparison with available LBD structures reveals key features governing the positioning of helix H12 and highlights the importance of cysteine residues in promoting an active conformation. Furthermore the present study reveals a hydrogen bond network connecting ligand binding to protein trans conformation. These observations support a dynamic view of H12 positioning, where the control of the equilibrium between two stable locations determines the partial agonist character of a given ligand. PMID- 11278578 TI - A naturally processed rat major histocompatibility complex class I-associated viral peptide as target structure of borna disease virus-specific CD8+ T cells. AB - The first naturally processed peptide synthesized by a virus and recognized by classical CD8(+) T cells in association with the RT1.A(l) major histocompatibility complex class I molecule of the Lewis rat is reported. Borna disease virus-specific CD8(+) T cells recognize syngeneic target cells pulsed with peptides extracted from Borna disease virus-infected cells. The predicted peptide sequence ASYAQMTTY from the viral p40 protein coeluted with the cytotoxic T-lymphocyte-reactive fraction was identified among natural ligands by tandem mass spectrometry. Numerous naturally processed peptides derived from intracellular bacteria, viruses, or tumors and recognized by CD8(+) T cells of man and mice are known, leading to a better understanding of cellular immune mechanisms against pathogens in these two species. In contrast, for the rat little information exists with regard to the function and role of CD8(+) T cells as part of their cellular immune defense system. This first naturally processed viral epitope in the rat contributes to the understanding of the rat cellular immune response and might trigger the identification of more cytotoxic T lymphocyte epitopes in this animal. PMID- 11278579 TI - Expression of CaT-like, a novel calcium-selective channel, correlates with the malignancy of prostate cancer. AB - The regulation of intracellular Ca(2+) plays a key role in the development and growth of cells. Here we report the cloning and functional expression of a highly calcium-selective channel localized on the human chromosome 7. The sequence of the new channel is structurally related to the gene product of the CaT1 protein cloned from rat duodenum and is therefore called CaT-like (CaT-L). CaT-L is expressed in locally advanced prostate cancer, metastatic and androgen insensitive prostatic lesions but is undetectable in healthy prostate tissue and benign prostatic hyperplasia. Additionally, CaT-L is expressed in normal placenta, exocrine pancreas, and salivary glands. New markers with well defined biological function that correlate with aberrant cell growth are needed for the molecular staging of cancer and to predict the clinical outcome. The human CaT-L channel represents a marker for prostate cancer progression and may serve as a target for therapeutic strategies. PMID- 11278581 TI - Characterization of binding properties of urinary trypsin inhibitor to cell associated binding sites on human chondrosarcoma cell line HCS-2/8. AB - Urinary trypsin inhibitor (UTI) forms membrane complexes with UTI-binding proteins (UTI-BPs) and initiates modulation of urokinase-type plasminogen activator (uPA) expression, which results in UTI-mediated suppression of cell invasiveness. It has been established that suppression of uPA expression and invasiveness by UTI is mediated through inhibition of protein kinase C-dependent signaling pathways and that human chondrosarcoma cell line HCS-2/8 expresses two types of UTI-BPs; a 40-kDa UTI-BP (UTI-BP(40)), which is identical to link protein (LP), and a 45-kDa UTI-BP (UTI-BP(45)). Here we characterize binding properties of UTI-BPs.UTI complexes in the cells. In vitro ligand blot, cell binding and competition assays, and Scatchard analyses demonstrate that both UTI BP(40) and UTI-BP(45) bind (125)I-UTI. A deglycosylated form of UTI (NG-UTI), from which the chondroitin-sulfate side chain has been removed, binds only to UTI BP(40). Additional experiments, using various reagents to block binding of (125)I UTI and NG-UTI to the UTI-BP(40) and UTI-BP(45) confirm that the chondroitin sulfate side chain of UTI is required for its binding to UTI-BP(45). Analysis of binding of (125)I-UTI and NG-UTI to the cells suggests that low affinity binding sites are the UTI-BP(40) (which can bind NG-UTI), and the high affinity sites are the UTI-BP(45). In addition, UTI-induced suppression of phorbol ester stimulated up-regulation of uPA is inhibited by reagents that were shown to prevent binding of UTI to the 40- and 45-kDa proteins. We conclude that UTI must bind to both of the UTI-BPs to suppress uPA up-regulation. PMID- 11278580 TI - Inhibition of cytosolic phospholipase A2 by annexin I. Specific interaction model and mapping of the interaction site. AB - Annexins (ANXs) display regulatory functions in diverse cellular processes, including inflammation, immune suppression, and membrane fusion. However, the exact biological functions of ANXs still remain obscure. Inhibition of phospholipase A(2) (PLA(2)) by ANX-I, a 346-amino acid protein, has been observed in studies with various forms of PLA(2). "Substrate depletion" and "specific interaction" have been proposed for the mechanism of PLA(2) inhibition by ANX-I. Previously, we proposed a specific interaction model for inhibition of a 100-kDa porcine spleen cytosolic form of PLA(2) (cPLA(2)) by ANX-I (Kim, K. M., Kim, D. K., Park, Y. M., and Na, D. S. (1994) FEBS Lett. 343, 251-255). Herein, we present an analysis of the inhibition mechanism of cPLA(2) by ANX-I in detail using ANX-I and its deletion mutants. Deletion mutants were produced in Escherichia coli, and inhibition of cPLA(2) activity was determined. The deletion mutant ANX-I-(1-274), containing the N terminus to amino acid 274, exhibited no cPLA(2) inhibitory activity, whereas the deletion mutant ANX-I-(275-346), containing amino acid 275 to the C terminus, retained full activity. The protein protein interaction between cPLA(2) and ANX-I was examined using the deletion mutants by immunoprecipitation and mammalian two-hybrid methods. Full-length ANX I and ANX-I-(275-346) interacted with the calcium-dependent lipid-binding domain of cPLA(2). ANX-I-(1-274) did not interact with cPLA(2). Immunoprecipitation of A549 cell lysate with anti-ANX-I antibody resulted in coprecipitation of cPLA(2). These results are consistent with the specific interaction mechanism rather than the substrate depletion model. ANX-I may function as a negative regulator of cPLA(2) in cellular signal transduction. PMID- 11278582 TI - Involvement of pRB-related p107 protein in the inhibition of S phase progression in response to genotoxic stress. AB - pRB family pocket proteins consisting of pRB, p107, and p130 are thought to act as a set of growth regulators that inhibit the cell cycle transition from G1 to S phases by virtue of their interaction with E2F transcription factors. When cells are committed to progressing through the cell cycle at the late G1 restriction point, they are hyperphosphorylated by G1 cyclin-cyclin-dependent kinase and are functionally inactivated. Consistent with such a G1 regulatory role, pRB and p130 are abundantly expressed in quiescent cells. In contrast, p107 is present at low levels in the hypophosphorylated form in quiescent cells. As cells progress toward late G1 to S phases, the levels of p107 increase, and the majority become hyperphosphorylated, suggesting a possible role of p107 in post-G1 cell cycle regulation. In this study, we have demonstrated that a nonphosphorylatable and thus constitutively active p107 has the potential to inhibit S phase progression. The levels of the phosphorylation-resistant p107 required for the S phase inhibition are significantly less than those of endogenous p107. We further show herein that the exposure of cells to the DNA-damaging agent, cisplatin, provokes S phase arrest, which is concomitantly associated with the accumulation of hypophosphorylated p107. Furthermore, the S phase inhibitory response to cisplatin is augmented by the ectopic expression of wild type p107, although it is diminished by the adenovirus E1A oncoprotein, which counteracts the pocket protein functions. Because p107 is a major pRB family protein expressed in S phase cells, our results indicate that p107 participates in an inhibition of cell cycle progression in response to DNA damage in S phase cells. PMID- 11278583 TI - Multiple effector domains within SNT1 coordinate ERK activation and neuronal differentiation of PC12 cells. AB - Differentiation of neuronal precursor cells in response to neurotrophic differentiation factors is accompanied by the activation of membrane-anchored SNT signaling adaptor proteins. Two classes of differentiation factors, the neurotrophins and fibroblast growth factors, induce rapid tyrosine phosphorylation of SNT1(FRS2alpha), which in turn enables SNT1 to recruit Shp2 tyrosine phosphatase and Grb2 adaptor protein in complex with the Ras GDP/GTP exchange factor Sos. To determine effector functions of SNT that promote neuronal differentiation of PC12 pheochromocytoma cells, we engineered a chimeric protein, SNT1(IRS)CX, bearing the effector region of SNT1 and the insulin receptor recognition domains of IRS2. Insulin promoted tyrosine phosphorylation of SNT1(IRS)CX in transfected PC12 cells accompanied by sustained activation of ERK1/2 mitogen-activated protein kinases and neuronal differentiation. The SNT1(IRS)CX-mediated response was dependent on endogenous Ras, MEK, and Shp2 activities. Mutagenesis of SNT1(IRS)CX identified three classes of effector motifs within SNT critical for both sustained ERK activation and neuronal differentiation: 1) four phosphotyrosine motifs that mediate recruitment of Grb2, 2) two phosphotyrosine motifs that mediate recruitment of Shp2, and 3) a C terminal motif that functions by helping to recruit Sos. We discuss possible mechanisms by which three functionally distinct SNT effector motifs collaborate to promote a downstream biochemical and biological response. PMID- 11278584 TI - Disruption of 3-phosphoinositide-dependent kinase 1 (PDK1) signaling by the anti tumorigenic and anti-proliferative agent n-alpha-tosyl-l-phenylalanyl chloromethyl ketone. AB - The anti-tumorigenic and anti-proliferative effects of N-alpha-tosyl-l phenylalanyl chloromethyl ketone (TPCK) have been known for more than three decades. Yet little is known about the discrete cellular targets of TPCK controlling these effects. Previous work from our laboratory showed TPCK, like the immunosuppressant rapamycin, to be a potent inhibitor of the 70-kilodalton ribosomal S6 kinase 1 (S6K1), which mediates events involved in cell growth and proliferation. We show here that rapamycin and TPCK display distinct inhibitory mechanisms on S6K1 as a rapamycin-resistant form of S6K1 was TPCK-sensitive. Additionally, we show that TPCK inhibited the activation of the related kinase and proto-oncogene Akt. Upstream regulators of S6K1 and Akt include phosphoinositide 3-kinase (PI 3-K) and 3-phosphoinositide-dependent kinase 1 (PDK1). Whereas TPCK had no effect on either mitogen-regulated PI 3-K activity or total cellular PDK1 activity, TPCK prevented phosphorylation of the PDK1 regulatory sites in S6K1 and Akt. Furthermore, whereas both PDK1 and the mitogen activated protein kinase (MAPK) are required for full activation of the 90 kilodalton ribosomal S6 kinase (RSK), TPCK inhibited RSK activation without inhibiting MAPK activation. Consistent with the capacity of RSK and Akt to mediate a cell survival signal, in part through phosphorylation of the pro apoptotic protein BAD, TPCK reduced BAD phosphorylation and led to cell death in interleukin-3-dependent 32D cells. Finally, in agreement with results seen in embryonic stem cells lacking PDK1, protein kinase A activation was not inhibited by TPCK showing TPCK specificity for mitogen-regulated PDK1 signaling. TPCK inhibition of PDK1 signaling thus disables central kinase cascades governing diverse cellular processes including proliferation and survival and provides an explanation for its striking biological effects. PMID- 11278585 TI - Novel alternatively spliced exon in the extracellular ligand-binding domain of the pituitary adenylate cyclase-activating polypeptide (PACAP) type 1 receptor (PAC1R) selectively increases ligand affinity and alters signal transduction coupling during spermatogenesis. AB - The expression of the paracrine signaling hormone pituitary adenylate cyclase activating polypeptide (PACAP) is regulated in a cyclical fashion during the 12 day spermatogenic cycle of the adult rat testis. The precise functions of PACAP in the development of germ cells are uncertain, but cycle- and stage-specific expression may augment cAMP-regulated gene expression in germ cells and associated Sertoli cells. Here we report the existence of a heretofore unrecognized exon in the extracellular domain of the PACAP type 1 receptor (PAC1R) that is alternatively spliced during the spermatogenic cycle in the rat testis. This splice variant encodes a full-length receptor with the insertion of an additional 72 base pairs encoding 24 amino acids (exon 3a) between coding exons 3 and 4. The PAC1R(3a) mRNA is preferentially detected in seminiferous tubules and is expressed at the highest levels in round spermatids and Sertoli cells. Analyses of ligand binding and signaling functions in stably transfected HEK293 cells expressing the two receptor isoforms reveals a 6-fold increase in the affinity of the PAC1R(3a) to bind PACAP-38, and alterations in its coupling to both cAMP and inositol phosphate signaling pathways relative to the wild type PAC1R. These findings suggest that the extracellular region between coding exons 3 and 6 of PAC1R may play an important role in the regulation of the relative ligand affinities and the relative coupling to G(s) (cAMP) and G(q) (inositol phosphates) signal transduction pathways during spermatogenesis. PMID- 11278587 TI - Amino acid residue mutations uncouple cooperative effects in Escherichia coli D-3 phosphoglycerate dehydrogenase. AB - d-3-Phosphoglycerate dehydrogenase from Escherichia coli contains two Gly-Gly sequences that occur at junctions between domains. A previous study (Grant, G. A., Xu, X. L., and Hu, Z. (2000) Biochemistry 39, 7316-7319) determined that the Gly-Gly sequence at the junction between the regulatory and substrate binding domain functions as a hinge between the domains. Mutations in this area significantly decrease the ability of serine to inhibit activity but have little effect on the K(m) and k(cat). Conversely, the present study shows that mutations to the Gly-Gly sequence at the junction of the substrate and nucleotide binding domains, which form the active site cleft, have a significant effect on the k(cat) of the enzyme without substantially altering the enzyme's sensitivity to serine. In addition, mutation of Gly-294, but not Gly-295, has a profound effect on the cooperativity of serine inhibition. Interestingly, even though cooperativity of inhibition can be reduced significantly, there is little apparent effect on the cooperativity of serine binding itself. An additional mutant, G336V,G337V, also reduces the cooperativity of inhibition, but in this case serine binding also is reduced to the point at which it cannot be measured by equilibrium dialysis. The double mutant G294V,G336V demonstrates that strain imposed by mutation at one hinge can be relieved partially by mutation at the other hinge, demonstrating linkage between the two hinge regions. These data show that the two cooperative processes, serine binding and catalytic inhibition, can be uncoupled. Consideration of the allowable torsional angles for the side chains introduced by the mutations yields a range of values for these angles that the glycine residues likely occupy in the native enzyme. A comparison of these values with the torsional angles found for the inhibited enzyme from crystal coordinates provides potential beginning and ending orientations for the transition from active to inhibited enzyme, which will allow modeling of the dynamics of domain movement. PMID- 11278586 TI - Mechanisms governing subcellular localization and function of human RGS2. AB - RGS proteins negatively regulate heterotrimeric G proteins at the plasma membrane. RGS2-GFP localizes to the nucleus, plasma membrane, and cytoplasm of HEK293 cells. Expression of activated G(q) increased RGS2 association with the plasma membrane and decreased accumulation in the nucleus, suggesting that signal induced redistribution may regulate RGS2 function. Thus, we identified and characterized a conserved N-terminal domain in RGS2 that is necessary and sufficient for plasma membrane localization. Mutational and biophysical analyses indicated that this domain is an amphipathic alpha-helix that binds vesicles containing acidic phospholipids. However, the plasma membrane targeting function of the amphipathic helical domain did not appear to be essential for RGS2 to attenuate signaling by activated G(q). Nevertheless, truncation mutants indicated that the N terminus is essential, potentially serving as a scaffold that binds receptors, signaling proteins, or nuclear components. Indeed, the RGS2 N terminus directs nuclear accumulation of GFP. Although RGS2 possesses a nuclear targeting motif, it lacks a nuclear import signal and enters the nucleus by passive diffusion. Nuclear accumulation of RGS2 does not limit its ability to attenuate G(q) signaling, because excluding RGS2 from the nucleus was without effect. RGS2 may nonetheless regulate signaling or other processes in the nucleus. PMID- 11278588 TI - A novel promoter element, photoreceptor conserved element II, directs photoreceptor-specific expression of nocturnin in Xenopus laevis. AB - Nocturnin is a vertebrate circadian clock-regulated gene, and in Xenopus laevis its mRNA is specifically expressed in retinal photoreceptor cells. We have investigated the transcriptional regulatory mechanism that drives this precise spatial expression pattern of the nocturnin gene. A deletion series of the nocturnin 5'-flanking sequence driving the green fluorescence protein (GFP) reporter was used to generate transgenic Xenopus tadpoles. We found that a construct containing 2.6 kilobase pairs of 5'-flanking sequence targeted high level GFP reporter expression specifically to photoreceptor cells, in a pattern identical to endogenous nocturnin. This photoreceptor-specific expression pattern was maintained with several further deletions of 5'-upstream sequence, including a short 59-base pair fragment. Within this region of 59 base pairs, three perfect repeats of a novel protein binding site were identified by electrophoretic mobility shift assay. Competitions using varying oligonucleotide sequences demonstrated that the sequence required for protein binding is CAGACAGGCTATA, designated photoreceptor-conserved element II (PCE II). The protein complex that binds to this element is enriched in retinal extracts, and mutations of PCE II which fail to bind the protein complex also fail to direct GFP reporter expression to photoreceptors. These results indicate that the PCE II in the proximal promoter of the nocturnin gene is sufficient for driving the photoreceptor-specific expression of nocturnin. PMID- 11278589 TI - Regulated nuclear-cytoplasmic localization of CCAAT/enhancer-binding protein delta in osteoblasts. AB - Insulin-like growth factor I (IGF-I) plays a central role in skeletal growth by promoting bone cell replication and differentiation. Prostaglandin E2 (PGE2) and parathyroid hormone enhance cAMP production in cultured rat osteoblasts and stimulate IGF-I expression through a transcriptional mechanism mediated by cAMP dependent protein kinase (PKA). We previously showed that PGE2 activated the transcription factor CCAAT/enhancer-binding protein delta (C/EBPdelta) in osteoblasts and induced its binding to a DNA element within the IGF-I promoter. We report here that a PKA-dependent pathway stimulates nuclear translocation of C/EBPdelta. Under basal conditions, C/EBPdelta was cytoplasmic but rapidly accumulated in the nucleus after PGE2 treatment (t(1/2) < 30 min). Nuclear translocation occurred without concurrent protein synthesis and was maintained in the presence of hormone. Nuclear localization required PKA and was blocked by a dominant-interfering regulatory subunit of the enzyme, even though C/EBPdelta was not a PKA substrate. Upon removal of hormonal stimulus, C/EBPdelta quickly exited the nucleus (t(1/2) < 12 min) through a pathway blocked by leptomycin B. Mutagenesis studies indicated that the basic domain of C/EBPdelta was necessary for nuclear localization and that the leucine zipper region permitted full nuclear accumulation. We thus define a pathway for PKA-mediated activation of C/EBPdelta through its regulated nuclear import. PMID- 11278590 TI - Colipase residues Glu64 and Arg65 are essential for normal lipase-mediated fat digestion in the presence of bile salt micelles. AB - Pancreatic triglyceride lipase (PTL) requires colipase for activity. Various constituents in meals and in bile, particularly bile acids, inhibit PTL. Colipase restores activity to lipase in the presence of inhibitory substances like bile acids. Presumably, colipase functions by anchoring and orienting PTL at the oil water interface. The x-ray structure of the colipase.PTL complex supports this model. In the x-ray structure, colipase has a hydrophobic surface positioned to bind substrate and a hydrophilic surface, lying opposite the hydrophobic surface, with two putative lipase-binding domains, Glu(45)/Asp(89) and Glu(64)/Arg(65). To determine whether the hydrophilic surface interacts with PTL in solution, we introduced mutations into the putative PTL binding domains of human colipase. Each mutant was expressed, purified, and assessed for activity against various substrates. Most of the mutants showed impaired ability to reactivate PTL, with mutations in the Glu(64)/Arg(65) binding site causing the greatest effect. Analysis indicated that the mutations decreased the affinity of the colipase mutants for PTL and prevented the formation of PTL.colipase complexes. The impaired function of the mutants was most apparent when assayed in micellar bile salt solutions. Most mutants stimulated PTL activity normally in monomeric bile salt solutions. We also tested the mutants for their ability to bind substrate and anchor lipase to tributyrin. Even though the ability of the mutants to anchor PTL to an interface decreased in proportion to their activity, each mutant colipase bound to tributyrin to the same extent as wild type colipase. These results demonstrate that the hydrophilic surface of colipase interacts with PTL in solution to form active colipase.PTL complexes, that bile salt micelles influence that binding, and that the proper interaction of colipase with PTL requires the Glu(64)/Arg(65) binding site. PMID- 11278591 TI - The crystal structures of Apo and complexed Saccharomyces cerevisiae GNA1 shed light on the catalytic mechanism of an amino-sugar N-acetyltransferase. AB - The yeast enzymes involved in UDP-GlcNAc biosynthesis are potential targets for antifungal agents. GNA1, a novel member of the Gcn5-related N-acetyltransferase (GNAT) superfamily, participates in UDP-GlcNAc biosynthesis by catalyzing the formation of GlcNAc6P from AcCoA and GlcN6P. We have solved three crystal structures corresponding to the apo Saccharomyces cerevisiae GNA1, the GNA1 AcCoA, and the GNA1-CoA-GlcNAc6P complexes and have refined them to 2.4, 1.3, and 1.8 A resolution, respectively. These structures not only reveal a stable, beta intertwined, dimeric assembly with the GlcNAc6P binding site located at the dimer interface but also shed light on the catalytic machinery of GNA1 at an atomic level. Hence, they broaden our understanding of structural features required for GNAT activity, provide structural details for related aminoglycoside N acetyltransferases, and highlight the adaptability of the GNAT superfamily members to acquire various specificities. PMID- 11278592 TI - Sphingosine 1-phosphate activates Akt, nitric oxide production, and chemotaxis through a Gi protein/phosphoinositide 3-kinase pathway in endothelial cells. AB - Sphingosine 1-phosphate (SPP) binds to members of the endothelial differentiation gene family (EDG) of receptors and leads to diverse signaling events including cell survival, growth, migration and differentiation. However, the mechanisms of how SPP activates these proangiogenic pathways are poorly understood. Here we show that SPP signals through the EDG-1 receptor to the heterotrimeric G protein G(i), leading to activation of the serine/threonine kinase Akt and phosphorylation of the Akt substrate, endothelial nitric-oxide synthase (eNOS). Inhibition of G(i) signaling, and phosphoinositide 3-kinase (PI 3-kinase) activity resulted in a decrease in SPP-induced endothelial cell chemotaxis. SPP also stimulates eNOS phosphorylation and NO release and these effects are also attenuated by inhibition of G(i) signaling, PI 3-kinase, and Akt. However, inhibition of NO production did not influence SPP-induced chemotaxis but effectively blocked the chemotactic actions of vascular endothelial growth factor. Thus, SPP signals through G(i) and PI 3-kinase leading to Akt activation and eNOS phosphorylation. PMID- 11278593 TI - Human corneal GlcNac 6-O-sulfotransferase and mouse intestinal GlcNac 6-O sulfotransferase both produce keratan sulfate. AB - Human corneal N-acetylglucosamine 6-O-sulfotransferase (hCGn6ST) has been identified by the positional candidate approach as the gene responsible for macular corneal dystrophy (MCD). Because of its high homology to carbohydrate sulfotransferases and the presence of mutations of this gene in MCD patients who lack sulfated keratan sulfate in the cornea and serum, hCGn6ST protein is thought to be a sulfotransferase that catalyzes sulfation of GlcNAc in keratan sulfate. In this report, we analyzed the enzymatic activity of hCGn6ST by expressing it in cultured cells. A lysate prepared from HeLa cells transfected with an intact form of hCGn6ST cDNA or culture medium from cells transfected with a secreted form of hCGn6ST cDNA showed an activity of transferring sulfate to C-6 of GlcNAc of synthetic oligosaccharide substrates in vitro. When hCGn6ST was expressed together with human keratan sulfate Gal-6-sulfotransferase (hKSG6ST), HeLa cells produced highly sulfated carbohydrate detected by an anti-keratan sulfate antibody 5D4. These results indicate that hCGn6ST transfers sulfate to C-6 of GlcNAc in keratan sulfate. Amino acid substitutions in hCGn6ST identical to changes resulting from missense mutations found in MCD patients abolished enzymatic activity. Moreover, mouse intestinal GlcNAc 6-O-sulfotransferase had the same activity as hCGn6ST. This observation suggests that mouse intestinal GlcNAc 6-O-sulfotransferase is the orthologue of hCGn6ST and functions as a sulfotransferase to produce keratan sulfate in the cornea. PMID- 11278594 TI - The propeptide of the transforming growth factor-beta superfamily member, macrophage inhibitory cytokine-1 (MIC-1), is a multifunctional domain that can facilitate protein folding and secretion. AB - Macrophage inhibitory cytokine-1 (MIC-1) is a divergent member of the transforming growth factor-beta (TGF-beta) superfamily. While it is synthesized in a pre-pro form, it is unique among superfamily members because it does not require its propeptide for correct folding or secretion of the mature peptide. To investigate factors that enable these propeptide independent events to occur, we constructed MIC-1/TGF-beta1 chimeras, both with and without a propeptide. All chimeras without a propeptide secreted less efficiently compared with the corresponding constructs with propeptide. Folding and secretion were most affected after replacement of the predicted major alpha-helix in the mature protein, residues 56-68. Exchanging the human propeptide in this chimera with either the murine MIC-1 or TGF-beta1 propeptide resulted in secretion of the unprocessed, monomeric chimera, suggesting a specific interaction between the human MIC-1 propeptide and mature peptide. Propeptide deletion mutants enabled identification of a region between residues 56 and 78, which is important for the interaction between the propeptide and the mature peptide. Cotransfection experiments demonstrated that the propeptide must be in cis with the mature peptide for this phenomenon to occur. These results suggest a model for TGF-beta superfamily protein folding. PMID- 11278595 TI - Casein kinase I associates with members of the centaurin-alpha family of phosphatidylinositol 3,4,5-trisphosphate-binding proteins. AB - Mammalian casein kinases I (CKI) belong to a family of serine/threonine protein kinases involved in diverse cellular processes including cell cycle progression, membrane trafficking, circadian rhythms, and Wnt signaling. Here we show that CKIalpha co-purifies with centaurin-alpha(1) in brain and that they interact in vitro and form a complex in cells. In addition, we show that the association is direct and occurs through the kinase domain of CKI within a loop comprising residues 217-233. These residues are well conserved in all members of the CKI family, and we show that centaurin-alpha(1) associates in vitro with all mammalian CKI isoforms. To date, CKIalpha represents the first protein partner identified for centaurin-alpha(1). However, our data suggest that centaurin alpha(1) is not a substrate for CKIalpha and has no effect on CKIalpha activity. Centaurin-alpha(1) has been identified as a phosphatidylinositol 3,4,5 trisphosphate-binding protein. Centaurin-alpha(1) contains a cysteine-rich domain that is shared by members of a newly identified family of ADP-ribosylation factor guanosine trisphosphatase-activating proteins. These proteins are involved in membrane trafficking and actin cytoskeleton rearrangement, thus supporting a role for CKIalpha in these biological events. PMID- 11278596 TI - Identification of an amino acid residue in multidrug resistance protein 1 critical for conferring resistance to anthracyclines. AB - Murine multidrug resistance protein 1 (mrp1), unlike human MRP1, does not confer resistance to anthracyclines. Previously, we have shown that a human/murine hybrid protein containing amino acids 959-1187 of MRP1 can confer resistance to these drugs. We have now examined the functional characteristics of mutant proteins in which we have converted individual amino acids in the comparable region of mrp1 to those present at the respective locations in MRP1. These mutations had no effect on the drug resistance profile conferred by mrp1 with the exception of converting glutamine 1086 to glutamate, as it is in the corresponding position (1089) in MRP1. This mutation created a protein that conferred resistance to doxorubicin without affecting vincristine resistance, or the ability of mrp1 to transport leukotriene C(4) (LTC(4)) and 17beta-estradiol 17-(beta-d-glucuronide) (E(2)17betaG). Furthermore, mutation Q1086D conferred the same phenotype as mutation Q1086E while the mutation Q1086N did not detectably alter the drug resistance profile of mrp1, suggesting that an anionic side chain was required for anthracycline resistance. To confirm the importance of MRP1 E1089 for conferring resistance to anthracyclines, we mutated this residue to Gln, Asp, Ala, Leu, and Lys in the human protein. The mutation E1089D showed the same phenotype as MRP1, while the E1089Q substitution markedly decreased resistance to anthracyclines without affecting LTC(4) and E(2)17betaG transport. Conversion of Glu-1089 to Asn, Ala, or Leu had a similar effect on resistance to anthracyclines, while conversion to a positive amino acid, Lys, completely eliminated resistance to anthracyclines and vincristine without affecting transport of LTC(4), E(2)17betaG, and the GSH-dependent substrate, estrone-3 sulfate. These results demonstrate that an acidic amino acid residue at position 1089 in predicted TM14 of MRP1 is critical for the ability of the protein to confer drug resistance particularly to the anthracyclines, but is not essential for its ability to transport conjugated organic anions such as LTC(4) and E(2)17betaG. PMID- 11278597 TI - The Bohr effect of hemoglobin intermediates and the role of salt bridges in the tertiary/quaternary transitions. AB - Understanding mechanisms in cooperative proteins requires the analysis of the intermediate ligation states. The release of hydrogen ions at the intermediate states of native and chemically modified hemoglobin, known as the Bohr effect, is an indicator of the protein tertiary/quaternary transitions, useful for testing models of cooperativity. The Bohr effects due to ligation of one subunit of a dimer and two subunits across the dimer interface are not additive. The reductions of the Bohr effect due to the chemical modification of a Bohr group of one and two alpha or beta subunits are additive. The Bohr effects of monoliganded chemically modified hemoglobins indicate the additivity of the effects of ligation and chemical modification with the possible exception of ligation and chemical modification of the alpha subunits. These observations suggest that ligation of a subunit brings about a tertiary structure change of hemoglobin in the T quaternary structure, which breaks some salt bridges, releases hydrogen ions, and is signaled across the dimer interface in such a way that ligation of a second subunit in the adjacent dimer promotes the switch from the T to the R quaternary structure. The rupture of the salt bridges per se does not drive the transition. PMID- 11278598 TI - Involvement of a triton-insoluble floating fraction in Dictyostelium cell-cell adhesion. AB - We have isolated and characterized a Triton-insoluble floating fraction (TIFF) from Dictyostelium. Ten major proteins were consistently detected in TIFF, and six species were identified by mass spectrometry as actin, porin, comitin, regulatory myosin light chain, a novel member of the CD36 family, and the phospholipid-anchored cell adhesion molecule gp80. TIFF was enriched with many acylated proteins. Also, the sterol/phospholipid ratio of TIFF was 10-fold higher than that of the bulk plasma membrane. Immunoelectron microscopy showed that TIFF has vesicular morphology and confirmed the association of gp80 and comitin with TIFF membranes. Several TIFF properties were similar to those of Dictyostelium contact regions, which were isolated as a cytoskeleton-associated membrane fraction. Mass spectrometry demonstrated that TIFF and contact regions shared the same major proteins. During development, gp80 colocalized with F-actin, porin, and comitin at cell-cell contacts. These proteins were also recruited to gp80 caps induced by antibody cross-linking. Filipin staining revealed high sterol levels in both gp80-enriched cell-cell contacts and gp80 caps. Moreover, sterol sequestration by filipin and digitonin inhibited gp80-mediated cell-cell adhesion. This study reveals that Dictyostelium TIFF has structural properties previously attributed to vertebrate TIFF and establishes a role for Dictyostelium TIFF in cell-cell adhesion during development. PMID- 11278599 TI - The cytoplasmic tail of alpha 1,2-fucosyltransferase contains a sequence for golgi localization. AB - The Golgi apparatus has a central role in the glycosylation of proteins and lipids. There is a sequential addition of carbohydrates by glycosyltransferases that are distributed within the Golgi in the order in which the glycosylation occurs. The mechanism of glycosyltransferase retention is considered to involve their transmembrane domains and flanking regions, although we have shown that the cytoplasmic tail of alpha1,2-fucosyltransferase is important for its Golgi localization. Here we show that the removal of the alpha1,2-fucosyltransferase cytoplasmic tail altered its function of fucosylation and its localization site. When the tail was removed, the enzyme moved from the Golgi to the trans Golgi network, suggesting that the transmembrane is responsible for retention and that the cytoplasmic tail is responsible for localization. The cytoplasmic tail of alpha1,2-fucosyltransferase contains 8 amino acids (MWVPSRRH), and mutating these to alanine indicated a role for amino acids 3 to 7 in localization with a particular role of Ser(5). Mutagenesis of Ser(5) to amino acids containing an hydroxyl (Tyr and Thr) demonstrated that the hydroxyl at position 5 is important. Thus, the cytoplasmic tail, and especially a single amino acid, has a predominant role in the localization and thus the function of alpha1,2-fucosyltransferase. PMID- 11278600 TI - Erk is essential for growth, differentiation, integrin expression, and cell function in human osteoblastic cells. AB - Extracellular signal-regulated kinases (Erks), members of the mitogen-activated protein kinase superfamily, play an important role in cell proliferation and differentiation. In this study we employed a dominant negative approach to determine the role of Erks in the regulation of human osteoblastic cell function. Human osteoblastic cells were transduced with a pseudotyped retrovirus encoding either a mutated Erk1 protein with a dominant negative action against both Erk1 and Erk2 (Erk1DN cells) or the LacZ protein (LacZ cells) as a control. Both basal and growth factor-stimulated MAPK activity and cell proliferation were inhibited in Erk1DN cells. Expression of Erk1DN protein suppressed both osteoblast differentiation and matrix mineralization by decreasing alkaline phosphatase activity and the deposition of bone matrix proteins. Cell adhesion to collagen, osteopontin, and vitronectin was decreased in Erk1DN cells as compared with LacZ cells. Cell spreading and migration on these matrices were also inhibited. In Erk1DN cells, expression of alphabeta(1), alpha(v)beta(3), and alpha(v)beta(5) integrins on the surface was decreased. Metabolic labeling indicated that the synthesis of these integrins was inhibited in Erk1DN cells. These data suggest that Erks are not only essential for the growth and differentiation of osteoblasts but also are important for osteoblast adhesion, spreading, migration, and integrin expression. PMID- 11278601 TI - Transcriptional repression by thyroid hormone receptors. A role for receptor homodimers in the recruitment of SMRT corepressor. AB - Nuclear hormone receptors, such as the thyroid hormone receptors (T3Rs) and retinoid X receptors (RXRs), are ligand-regulated transcription factors that control key aspects of metazoan gene expression. T3Rs can bind to DNA either as receptor homodimers or as heterodimers with RXRs. Once bound to DNA, nuclear hormone receptors regulate target gene expression by recruiting auxiliary proteins, denoted corepressors and coactivators. We report here that T3R homodimers assembled on DNA exhibit particularly strong interactions with the SMRT corepressor, whereas T3R.RXR heterodimers are inefficient at binding to SMRT. Mutants of T3R that exhibit enhanced repression properties, such as the v Erb A oncoprotein or the T3Rbeta-Delta432 mutant found in human resistance to thyroid hormone syndrome, display enhanced homodimerization properties and exhibit unusually strong interactions with the SMRT corepressor. Significantly, the topology of a DNA binding site can determine whether that site recruits primarily homodimers or heterodimers and therefore whether corepressor is efficiently or inefficiently recruited to the resulting receptor-DNA complex. We suggest that T3R homodimers, and not heterodimers, may be important mediators of transcriptional repression and that the nature of the DNA binding site, by selecting for receptor homodimers or heterodimers, can influence the ability of the receptor to recruit corepressor. PMID- 11278602 TI - Sustained nitric oxide production in macrophages requires the arginine transporter CAT2. AB - The aberrant production of nitric oxide (NO) contributes to the pathogenesis of diseases as diverse as cancer and arthritis. Sustained NO production via the inducible enzyme, nitric-oxide synthase 2 (NOS2), requires extracellular arginine uptake. Three closely related cationic amino acid transporter genes (Cat1-3) encode the transporters that mediate most arginine uptake in mammalian cells. Because CAT2 is induced coordinately with NOS2 in numerous cell types, we investigated a possible role for CAT2-mediated arginine transport in regulating NO production. The complexity of arginine transport systems and their biochemically similar transport properties called for a genetic approach to determine the role of CAT2. CAT2-deficient mice were generated and found to be healthy and fertile in contrast to Cat1(-/-) animals. Analysis of cytokine activated macrophages from Cat2(-/-) mice revealed a 92% reduction in NO production and a 95% reduction in l-Arg uptake. The reduction in NO production was not due to differences in NOS2 protein expression, NOS2 activity, or intracellular l-arginine content. In conclusion, our results show that sustained abundant NO synthesis by macrophages requires arginine transport via the CAT2 transporter. PMID- 11278603 TI - The ERBB2/HER2 receptor differentially interacts with ERBIN and PICK1 PSD 95/DLG/ZO-1 domain proteins. AB - Identification of protein complexes associated with the ERBB2/HER2 receptor may help unravel the mechanisms of its activation and regulation in normal and pathological situations. Interactions between ERBB2/HER2 and Src homology 2 or phosphotyrosine binding domain signaling proteins have been extensively studied. We have identified ERBIN and PICK1 as new binding partners for ERBB2/HER2 that associate with its carboxyl-terminal sequence through a PDZ (PSD-95/DLG/ZO-1) domain. This peptide sequence acts as a dominant retention or targeting basolateral signal for receptors in epithelial cells. ERBIN belongs to the newly described LAP (LRR and PDZ) protein family, whose function is crucial in non vertebrates for epithelial homeostasis. Whereas ERBIN appears to locate ERBB2/HER2 to the basolateral epithelium, PICK1 is thought to be involved in the clustering of receptors. We show here that ERBIN and PICK1 bind to ERBB2/HER2 with different mechanisms, and we propose that these interactions are regulated in cells. Since ERBIN and PICK1 tend to oligomerize, further complexity of protein networks may participate in ERBB2/HER2 functions and specificity. PMID- 11278604 TI - Chinese hamster ovary (CHO) cells may express six beta 4-galactosyltransferases (beta 4GalTs). Consequences of the loss of functional beta 4GalT-1, beta 4GalT-6, or both in CHO glycosylation mutants. AB - Six beta4-galactosyltransferase (beta4GalT) genes have been cloned from mammalian sources. We show that all six genes are expressed in the Gat(-)2 line of Chinese hamster ovary cells (Gat(-)2 CHO). Two independent mutants termed Pro(-)5Lec20 and Gat(-)2Lec20, previously selected for lectin resistance, were found to have a galactosylation defect. Radiolabeled biantennary N-glycans synthesized by Pro( )5Lec20 were proportionately less ricin-bound than similar species from parental CHO cells, and Lec20 cell extracts had a markedly reduced ability to transfer Gal to GlcNAc-terminating acceptors. Northern blot analysis revealed a severe reduction in beta4GalT-1 transcripts in Pro(-)5Lec20 cells. The Gat(-)2Lec20 mutant expressed beta4GalT-1 transcripts of reduced size due to a 311-base pair deletion in the beta4GalT-1 gene coding region. Northern analysis with probes from the remaining five beta4GalT genes revealed that Gat(-)2 CHO and Gat( )2Lec20 cells express all six beta4GalT genes. Unexpectedly, the beta4GalT-6 gene is not expressed in either Pro(-)5 or Pro(-)5Lec20 cells. Thus, in addition to a deficiency in beta4GalT-1, Pro(-)5Lec20 cells lack beta4GalT-6. Nevertheless, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry data of N-glycans released from cellular glycoproteins showed that both the beta4GalT 1(-) (Gat(-)2Lec20) and beta4GalT-1(-)/beta4GalT-6(-) (Pro(-)5Lec20) mutants have a similar Gal deficiency, affecting neutral and sialylated bi-, tri-, and tetraantennary N-glycans. By contrast, glycolipid synthesis was normal in both mutants. Therefore, beta4GalT-1 is a key enzyme in the galactosylation of N glycans, but is not involved in glycolipid synthesis in CHO cells. beta4GalT-6 contributes only slightly to the galactosylation of N-glycans and is also not involved in CHO cell glycolipid synthesis. These CHO glycosylation mutants provide insight into the variety of in vivo substrates of different beta4GalTs. They may be used in glycosylation engineering and in investigating roles for beta4GalT-1 and beta4GalT-6 in generating specific glycan ligands. PMID- 11278605 TI - Structure and expression of the TREX1 and TREX2 3' --> 5' exonuclease genes. AB - The TREX1 and TREX2 genes encode mammalian 3'-->5' exonucleases. Expression of the TREX genes in human cells was investigated using a reverse transcription polymerase chain reaction strategy. Our results show that TREX1 and TREX2 are expressed in all tissues tested, providing direct evidence for the expression of these genes in human cells. Potential transcription start sites are identified for the TREX genes using rapid amplification of cDNA ends to recover the 5' flanking regions of the TREX transcripts. The 5'-flanking sequences indicate transcription initiation from consensus putative promoters identified -140 and 650 base pairs upstream of the TREX1 open reading frame (ORF) and -623 and -753 base pairs upstream of the TREX2 ORF. Novel TREX1 and TREX2 cDNAs are identified that contain protein-coding sequences generated from exons positioned in genomic DNA up to 18 kilobases 5' to the TREX1 ORF and up to 25 kilobases 5' to the TREX2 ORF. These novel cDNAs and sequences in the GenBank data base indicate that transcripts containing the TREX1 and TREX2 ORFs are produced using a variety of mechanisms that include alternate promoter usage, alternative splicing, and varied sites for 3' cleavage and polyadenylation. These initial studies have revealed previously unrecognized complexities in the structure and expression of the TREX1 and TREX2 genes. PMID- 11278606 TI - Melanoma chondroitin sulfate proteoglycan regulates matrix metalloproteinase dependent human melanoma invasion into type I collagen. AB - Tumor cell adhesion and proteolysis of the extracellular matrix proteins surrounding the cells are tightly linked processes in tumor invasion. In this study, we sought to identify components of the cell surface of a vertical growth phase melanoma cell line, WM1341D, that mediate invasive cellular behavior. We determined by antisense inhibition that melanoma chondroitin sulfate proteoglycan (MCSP) and membrane-type 3 matrix metalloproteinase (MT3-MMP) expressed on WM1341D are required for invasion of type I collagen and degradation of type I gelatin. MT3-MMP co-immunoprecipitated with MCSP in WM1341D melanoma cells cultured on type I collagen or laminin. The association between MT3-MMP and MCSP was largely disrupted by removing chondroitin sulfate glycosaminoglycan (CS) from the cell surface, suggesting CS could mediate the association between the two cell surface core proteins. Recombinant MT3-MMP and MT3-MMP from whole cell lysates of WM1341D cells were specifically eluted from CS- conjugated affinity columns. The results indicate that MT3-MMP possesses the potential to promote melanoma invasion and proteolysis and that the formation of a complex between MT3 MMP and MCSP may be a crucial step in activating these processes. PMID- 11278607 TI - The tumor-sensitive calmodulin-like protein is a specific light chain of human unconventional myosin X. AB - Human calmodulin-like protein (CLP) is an epithelial-specific Ca(2+)-binding protein whose expression is strongly down-regulated in cancers. Like calmodulin, CLP is thought to regulate cellular processes via Ca(2+)-dependent interactions with specific target proteins. Using gel overlays, we identified a approximately 210-kDa protein binding specifically and in a Ca(2+)-dependent manner to CLP, but not to calmodulin. Yeast two-hybrid screening yielded a CLP-interacting clone encoding the three light chain binding IQ motifs of human "unconventional" myosin X. Pull-down experiments showed CLP binding to the IQ domain to be direct and Ca(2+)-dependent. CLP interacted strongly with IQ motif 3 (K(d) approximately 0.5 nm) as determined by surface plasmon resonance. Epitope-tagged myosin X was localized preferentially at the cell periphery in MCF-7 cells, and CLP colocalized with myosin X in these cells. Myosin X was able to coprecipitate CLP and, to a lesser extent, calmodulin from transfected COS-1 cells, indicating that CLP is a specific light chain of myosin X in vivo. Because unconventional myosins participate in cellular processes ranging from membrane trafficking to signaling and cell motility, myosin X is an attractive CLP target. Altered myosin X regulation in (tumor) cells lacking CLP may have as yet unknown consequences for cell growth and differentiation. PMID- 11278608 TI - NF-kappaB activation is involved in regulation of cystic fibrosis transmembrane conductance regulator (CFTR) by interleukin-1beta. AB - Interleukin-1 beta (IL-1beta) regulates the levels of cystic fibrosis transmembrane conductance regulator (CFTR) mRNA and protein in the T84 human carcinoma cell line. Here, we studied the role of the transcription factor NF kappaB in this regulation. Initially, T84 cells were pretreated with the NF kappaB inhibitor pyrrolidine dithiocarbamate. Cells were then stimulated with IL 1beta, and CFTR mRNA levels were determined after 4 h by Northern blot analysis. As a result of PDTC treatment, IL-1beta stimulation of CFTR mRNA was blocked. On the other hand, daunorubicin, an NF-kappaB activator, increased the steady-state levels of CFTR mRNA. Furthermore, after treatment with IL-1beta for 1 h, cytoplasmic IkappaBalpha degradation occurred simultaneously with translocation of p65 into the nucleus. The T84 cells were also transduced with an adenoviral vector expressing a dominant negative form of IkappaBalpha, which prevents IkappaBalpha phosphorylation and the subsequent nuclear translocation of NF kappaB. After viral transduction, the cells were stimulated with IL-1beta for 4 h, and CFTR mRNA levels were measured by Northern blot analysis. The stimulation of CFTR, induced by IL-1beta, was also blocked in the presence of the dominant negative mutant. These results indicate that NF-kappaB is involved in the pathway by which IL-1beta regulates CFTR. PMID- 11278609 TI - Hyaluronan synthase elevation in metastatic prostate carcinoma cells correlates with hyaluronan surface retention, a prerequisite for rapid adhesion to bone marrow endothelial cells. AB - Bone marrow is the primary site of metastasis in patients with advanced stage prostate cancer. Prostate carcinoma cells metastasizing to bone must initially adhere to endothelial cells in the bone marrow sinusoids. In this report, we have modeled that interaction in vitro using two bone marrow endothelial cell (BMEC) lines and four prostate adenocarcinoma cell lines to investigate the adhesion mechanism. Highly metastatic PC3 and PC3M-LN4 cells were found to adhere rapidly and specifically (70-90%) to BMEC-1 and trHBMEC bone marrow endothelial cells, but not to human umbilical vein endothelial cells (15-25%). Specific adhesion to BMEC-1 and trHBMEC was dependent upon the presence of a hyaluronan (HA) pericellular matrix assembled on the prostate carcinoma cells. DU145 and LNCaP cells were only weakly adherent and retained no cell surface HA. Maximal BMEC adhesion and HA encapsulation were associated with high levels of HA synthesis by the prostate carcinoma cells. Up-regulation of HA synthase isoforms Has2 and Has3 relative to levels expressed by normal prostate corresponded to elevated HA synthesis and avid BMEC adhesion. These results support a model in which tumor cells with up-regulated HA synthase expression assemble a cell surface hyaluronan matrix that promotes adhesion to bone marrow endothelial cells. This interaction could contribute to preferential bone metastasis by prostate carcinoma cells. PMID- 11278610 TI - The SOCS box of SOCS-1 accelerates ubiquitin-dependent proteolysis of TEL-JAK2. AB - Fusion of the TEL gene on 12p13 to the JAK2 tyrosine kinase gene on 9p24 has been found in human leukemia. TEL-mediated oligomerization of JAK2 results in constitutive activation of the tyrosine kinase (JH1) domain and confers cytokine independent proliferation on interleukin-3-dependent Ba/F3 cells. Forced expression of the JAK inhibitor gene SOCS1/JAB/SSI-1 induced apoptosis of TEL JAK2-transformed Ba/F3 cells. This suppression of TEL-JAK2 activity was dependent on SOCS box-mediated proteasomal degradation of TEL-JAK2 rather than on kinase inhibition. Degradation of JAK2 depended on its phosphorylation and its high affinity binding with SOCS1 through the kinase inhibitory region and the SH2 domain. It has been demonstrated that von Hippel-Lindau disease (VHL) tumor suppressor gene product possesses the SOCS box that forms a complex with Elongin B and C and Cullin-2, and it functions as a ubiquitin ligase. The SOCS box of SOCS1/JAB has also been shown to interact with Elongins; however, ubiquitin ligase activity has not been demonstrated. We found that the SOCS box interacted with Cullin-2 and promoted ubiquitination of TEL-JAK2. Furthermore, overexpression of dominant negative Cullin-2 suppressed SOCS1-dependent TEL-JAK2 degradation. Our study demonstrates the substrate-specific E3 ubiquitin-ligase like activity of SOCS1 for activated JAK2 and may provide a novel strategy for the suppression of oncogenic tyrosine kinases. PMID- 11278611 TI - Evidence for distinct cholesterol domains in fiber cell membranes from cataractous human lenses. AB - Previous studies in our laboratory have provided direct evidence for the existence of distinct cholesterol domains within the plasma membranes of human ocular lens fiber cells. The fiber cell plasma membrane is unique in that it contains unusually high concentrations of cholesterol, with cholesterol to phospholipid (C/P) mole ratios ranging from 1 to 4. Since membrane cholesterol content is disturbed in the development of cataracts, it was hypothesized that perturbation of cholesterol domain structure occurs in cataracts. In this study, fiber cell plasma membranes were isolated from both normal (control) and cataractous lenses and assayed for cholesterol and phospholipid. Control and cataractous whole lens membranes had C/P mole ratios of 3.1 and 1.7, respectively. Small angle x-ray diffraction approaches were used to directly examine the structural organization of the cataractous lens plasma membrane versus control. Both normal and cataractous oriented membranes yielded meridional diffraction peaks corresponding to a unit cell periodicity of 34.0 A, consistent with the presence of immiscible cholesterol domains. However, comparison of diffraction patterns indicated that cataractous lens membranes contained more pronounced and better defined cholesterol domains than controls, over a broad range of temperature (5-40 degrees C) and relative humidity (52-92%) levels. In addition, diffraction analyses of the sterol-poor regions of cataractous membranes indicated increased membrane rigidity as compared with control membranes. Modification of the membrane lipid environment, such as by oxidative insult, is believed to be one potential mechanism for the formation of highly resolved cholesterol domains despite significantly reduced cholesterol content. The results of this x-ray diffraction study provide evidence for fundamental changes in the lens fiber cell plasma membrane structure in cataracts, including the presence of more prominent and highly ordered, immiscible cholesterol domains. PMID- 11278612 TI - Role of hydrophobic residues in the C1b domain of protein kinase C delta on ligand and phospholipid interactions. AB - The C1 domains of conventional and novel protein kinase C (PKC) isoforms bind diacylglycerol and phorbol esters with high affinity. Highly conserved hydrophobic residues at or near the rim of the binding cleft in the second cysteine-rich domain of PKC-delta (PKC-deltaC1b) were mutated to probe their roles in ligand recognition and lipid interaction. [(3)H]Phorbol 12,13-dibutyrate (PDBu) binding was carried out both in the presence and absence of phospholipids to determine the contribution of lipid association to the ligand affinity. Lipid dependence was determined as a function of lipid concentration and composition. The binding properties of a high affinity branched diacylglycerol with lipophilicity similar to PDBu were compared with those of PDBu to identify residues important for ligand selectivity. As expected, Leu-20 and Leu-24 strongly influenced binding. Substitution of either by aspartic acid abolished binding in either the presence or absence of phosphatidylserine. Mutation of Leu 20 to Arg or of Leu-24 to Lys caused a dramatic (340- and 250-fold, respectively) reduction in PDBu binding in the presence of lipid but only a modest reduction in the weaker binding of PDBu observed in the absence of lipid, suggesting that the main effect was on C1 domain -phospholipid interactions. Mutation of Leu-20 to Lys or of Trp-22 to Lys had modest (3-fold) effects and mutation of Phe-13 to Tyr or Lys was without effect. Binding of the branched diacylglycerol was less dependent on phospholipid and was more sensitive to mutation of Trp-22 to Tyr or Lys, especially in the presence of phospholipid, than was PDBu. In terms of specific PKC isoforms, our results suggest that the presence of Arg-20 in PKC zeta may contribute to its lack of phorbol ester binding activity. More generally, the results emphasize the interplay between the C1 domain, ligand, and phospholipid in the ternary binding complex. PMID- 11278614 TI - Cloning and characterization of IL-1HY2, a novel interleukin-1 family member. AB - The interleukin-1 (IL-1) family members play an important role in the process of inflammation and host defense. We describe here the identification and characterization of a novel member of the IL-1 family, IL-1HY2. The human IL-1HY2 protein shares significant amino acid sequence similarity (37%) with the IL-1 receptor antagonist and has a predicted three-dimensional structure similar to that of the IL-1 receptor antagonist. The IL-1HY2 gene is located in close proximity to other IL-1 family genes on human chromosome 2, and the genomic organization of the IL-1HY2 gene is highly conserved with other IL-1 family members. IL-1HY2 protein is secreted from mammalian cells, and the purified recombinant IL-1HY2 protein binds soluble IL-1 receptor type I. IL-1HY2 is expressed in human skin, spleen, and tonsil. Immunohistochemical analysis showed that the IL-1HY2 protein is expressed in the basal epithelia of skin and in proliferating B cells of the tonsil. These data suggest that IL-1HY2 is a novel IL-1 family member and that it may participate in a network of IL-1 family members to regulate adapted and innate immune responses. PMID- 11278613 TI - Akt/PKB activity is required for Ha-Ras-mediated transformation of intestinal epithelial cells. AB - Phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB/Akt) is thought to serve as an oncogenic signaling pathway which can be activated by Ras. The role of PI3K/Akt in Ras-mediated transformation of intestinal epithelial cells is currently not clear. Here we demonstrate that inducible expression of oncogenic Ha-Ras results in activation of PKB/Akt in rat intestinal epithelial cells (RIE iHa-Ras), which was blocked by treatment with inhibitors of PI3K activity. The PI3K inhibitor, LY-294002, partially reversed the morphological transformation induced by Ha-Ras and resulted in a modest stimulation of apoptosis. The most pronounced phenotypic alteration following inhibition of PI3K was induction of G(1) phase cell cycle arrest. LY-294002 blocked the Ha-Ras-induced expression of cyclin D1, cyclin-dependent kinase (CDK) 2, and increased the levels of p27(kip). Both LY-294002 and wortmannin significantly reduced anchorage-independent growth of RIE-iHa-Ras cells. Forced expression of both the constitutively active forms of Raf (DeltaRaf-22W or Raf BXB) and Akt (Akt-myr) resulted in transformation of RIE cells that was not achieved by transfection with either the Raf mutant construct or Akt-myr alone. These findings delineate an important role for PI3K/Akt in Ras-mediated transformation of intestinal epithelial cells. PMID- 11278615 TI - Eicosanoids inhibit the G-protein-gated inwardly rectifying potassium channel (Kir3) at the Na+/PIP2 gating site. AB - We previously showed that activation of the human endothelin A receptor (HETAR) by endothelin-1 (Et-1) selectively inhibits the response to mu opioid receptor (MOR) activation of the G-protein-gated inwardly rectifying potassium channel (Kir3). The Et-1 effect resulted from PLA2 production of an eicosanoid that inhibited Kir3. In this study, we show that Kir3 inhibition by eicosanoids is channel subunit-specific, and we identify the site within the channel required for arachidonic acid sensitivity. Activation of the G-protein-coupled MOR by the selective opioid agonist D-Ala(2)Glyol, enkephalin, released Gbetagamma that activated Kir3. The response to MOR activation was significantly inhibited by Et 1 activation of HETAR in homomeric channels composed of either Kir3.2 or Kir3.4. In contrast, homomeric channels of Kir3.1 were substantially less sensitive. Domain deletion and channel chimera studies suggested that the sites within the channel required for Et-1-induced inhibition were within the region responsible for channel gating. Mutation of a single amino acid in the homomeric Kir3.1 to produce Kir3.1(F137S)(N217D) dramatically increased the channel sensitivity to arachidonic acid and Et-1 treatment. Complementary mutation of the equivalent amino acid in Kir3.4 to produce Kir3.4(S143T)(D223N) significantly reduced the sensitivity of the channel to arachidonic acid- and Et-1-induced inhibition. The critical aspartate residue required for eicosanoid sensitivity is the same residue required for Na(+) regulation of PIP(2) gating. The results suggest a model of Kir3 gating that incorporates a series of regulatory steps, including Gbetagamma, PIP(2), Na(+), and arachidonic acid binding to the channel gating domain. PMID- 11278617 TI - The Escherichia coli RNA polymerase.anti-sigma 70 AsiA complex utilizes alpha carboxyl-terminal domain upstream promoter contacts to transcribe from a -10/-35 promoter. AB - During infection of Escherichia coli, the phage T4 early protein AsiA inhibits open complex formation by the RNA polymerase holoenzyme Efinal sigma(70) at -10/ 35 bacterial promoters through binding to region 4.2 of the final sigma(70) subunit. We used the -10/-35 lacUV5 promoter to study the properties of the Efinal sigma(70). AsiA complex in the presence of the glutamate anion. Under these experimental conditions, inhibition by AsiA was significantly decreased. KMnO(4) probing showed that the observed residual transcriptional activity was due to the slow transformation of the ternary complex Efinal sigma(70). AsiA.lacUV5 into an open complex. In agreement with this observation, affinity of the enzyme for the promoter was 10-fold lower in the ternary complex than in the binary complex Efinal sigma(70).lacUV5. A tau plot analysis of abortive transcription reactions showed that AsiA binding to Efinal sigma(70) resulted in a 120-fold decrease in the second-order on-rate constant of the reaction of Efinal sigma(70) with lacUV5 and a 55-fold decrease in the rate constant of the isomerization step leading to the open complex. This ternary complex still responded to activation by the cAMP.catabolite activator protein complex. We show that compensatory Efinal sigma(70)/promoter upstream contacts involving the C terminal domains of alpha subunits in Efinal sigma(70) become essential for the binding of Efinal sigma(70). AsiA to the lacUV5 promoter. PMID- 11278616 TI - Phosphorylation of xanthine dehydrogenase/oxidase in hypoxia. AB - The enzyme xanthine oxidase (XO) has been implicated in the pathogenesis of several disease processes, such as ischemia-reperfusion injury, because of its ability to generate reactive oxygen species. The expression of XO and its precursor xanthine dehydrogenase (XDH) is regulated at pre- and posttranslational levels by agents such as lipopolysaccharide and hypoxia. Posttranslational modification of the protein, for example through thiol oxidation or proteolysis, has been shown to be important in converting XDH to XO. The possibility of posttranslational modification of XDH/XO through phosphorylation has not been adequately investigated in mammalian cells, and studies have reported conflicting results. The present report demonstrates that XDH/XO is phosphorylated in rat pulmonary microvascular endothelial cells (RPMEC) and that phosphorylation is greatly increased ( approximately 50-fold) in response to acute hypoxia (4 h). XDH/XO phosphorylation appears to be mediated, at least in part, by casein kinase II and p38 kinase as inhibitors of these kinases partially prevent XDH/XO phosphorylation. In addition, the results indicate that p38 kinase, a stress activated kinase, becomes activated in response to hypoxia (an approximately 4 fold increase after 1 h of exposure of RPMEC to hypoxia) further supporting a role for this kinase in hypoxia-stimulated XDH/XO phosphorylation. Finally, hypoxia-induced XDH/XO phosphorylation is accompanied by a 2-fold increase in XDH/XO activity, which is prevented by inhibitors of phosphorylation. In summary, this study shows that XDH/XO is phosphorylated in hypoxic RPMEC through a mechanism involving p38 kinase and casein kinase II and that phosphorylation is necessary for hypoxia-induced enzymatic activation. PMID- 11278618 TI - The UL5 and UL52 subunits of the herpes simplex virus type 1 helicase-primase subcomplex exhibit a complex interdependence for DNA binding. AB - Herpes simplex virus type 1 encodes a heterotrimeric helicase-primase complex composed of the products of the UL5, UL52, and UL8 genes. The UL5 protein contains seven motifs found in all members of helicase Superfamily 1 (SF1), and the UL52 protein contains several conserved motifs found in primases; however, the contributions of each subunit to the biochemical activities of the subcomplex are not clear. In this work, the DNA binding properties of wild type and mutant subcomplexes were examined using single-stranded, duplex, and forked substrates. A gel mobility shift assay indicated that the UL5-UL52 subcomplex binds more efficiently to the forked substrate than to either single strand or duplex DNA. Although nucleotides are not absolutely required for DNA binding, ADP stimulated the binding of UL5-UL52 to single strand DNA whereas ATP, ADP, and adenosine 5'-O (thiotriphosphate) stimulated the binding to a forked substrate. We have previously shown that both subunits contact single-stranded DNA in a photocross linking assay (Biswas, N., and Weller, S. K. (1999) J. Biol. Chem. 274, 8068 8076). In this study, photocross-linking assays with forked substrates indicate that the UL5 and UL52 subunits contact the forked substrates at different positions, UL52 at the single-stranded DNA tail and UL5 near the junction between single-stranded and double-stranded DNA. Neither subunit was able to cross-link a forked substrate when 5-iododeoxyuridine was located within the duplex portion. Photocross-linking experiments with subcomplexes containing mutant versions of UL5 and wild type UL52 indicated that the integrity of the ATP binding region is important for DNA binding of both subunits. These results support our previous proposal that UL5 and UL52 exhibit a complex interdependence for DNA binding (Biswas, N., and Weller, S. K. (1999) J. Biol. Chem. 274, 8068-8076) and indicate that the UL52 subunit may play a more active role in helicase activity than had previously been thought. PMID- 11278619 TI - Control of mitochondrial redox balance and cellular defense against oxidative damage by mitochondrial NADP+-dependent isocitrate dehydrogenase. AB - Mitochondria are the major organelles that produce reactive oxygen species (ROS) and the main target of ROS-induced damage as observed in various pathological states including aging. Production of NADPH required for the regeneration of glutathione in the mitochondria is critical for scavenging mitochondrial ROS through glutathione reductase and peroxidase systems. We investigated the role of mitochondrial NADP(+)-dependent isocitrate dehydrogenase (IDPm) in controlling the mitochondrial redox balance and subsequent cellular defense against oxidative damage. We demonstrate in this report that IDPm is induced by ROS and that decreased expression of IDPm markedly elevates the ROS generation, DNA fragmentation, lipid peroxidation, and concurrent mitochondrial damage with a significant reduction in ATP level. Conversely, overproduction of IDPm protein efficiently protected the cells from ROS-induced damage. The protective role of IDPm against oxidative damage may be attributed to increased levels of a reducing equivalent, NADPH, needed for regeneration of glutathione in the mitochondria. Our results strongly indicate that IDPm is a major NADPH producer in the mitochondria and thus plays a key role in cellular defense against oxidative stress-induced damage. PMID- 11278620 TI - Early events in glycosylphosphatidylinositol anchor addition. substrate proteins associate with the transamidase subunit gpi8p. AB - The addition of glycosylphosphatidylinositol (GPI) anchors to proteins occurs by a transamidase-catalyzed reaction mechanism soon after completion of polypeptide synthesis and translocation. We show that placental alkaline phosphatase becomes efficiently GPI-anchored when translated in the presence of semipermeabilized K562 cells but is not GPI-anchored in cell lines defective in the transamidase subunit hGpi8p. By studying the synthesis of placental alkaline phosphatase, we demonstrate that folding of the protein is not influenced by the addition of a GPI anchor and conversely that GPI anchor addition does not require protein folding. These results demonstrate that folding of the ectodomain and GPI addition are two distinct processes and can be mutually exclusive. When GPI addition is prevented, either by synthesis of the protein in the presence of cell lines defective in GPI addition or by mutation of the GPI carboxyl-terminal signal sequence cleavage site, the substrate forms a prolonged association with the transamidase subunit hGpi8p. The ability of the transamidase to recognize and associate with GPI anchor signal sequences provides an explanation for the retention of GPI-anchored protein within the ER in the absence of GPI anchor addition. PMID- 11278621 TI - Fli-1 inhibits collagen type I production in dermal fibroblasts via an Sp1 dependent pathway. AB - Fibrosis is characterized by the excessive deposition of extracellular matrix (ECM), especially collagen. Because Ets factors are implicated in physiological and pathological ECM remodeling, the aim of this study was to investigate the role of Ets factors in collagen production. We demonstrate that the expression of collagenous proteins and collagen alpha2(I) (COL1A2) mRNA was inhibited following stable transfection of Fli-1 in dermal fibroblasts. Subsequent analysis of the COL1A2 promoter identified a critical Ets binding site that mediates Fli-1 inhibition. In contrast, Ets-1 stimulates COL1A2 promoter activity. In vitro binding assays demonstrate that both Fli-1 and Ets-1 form DNA-protein complexes with sequences present in COL1A2 promoter. Furthermore, Fli-1 binding to the COL1A2 is enhanced via Sp1-dependent interaction. Studies using Fli-1 dominant interference and DNA binding mutants indicate that Fli-1 inhibition is mediated by both direct (DNA binding) and indirect (via protein-protein interaction) mechanisms and that Sp1 is an important mediator of the Fli-1 function. Furthermore, experiments using the Gal4 system in the context of different cell types as well as experiments with the COL1A2 promoter in different cell lines demonstrate that the direction and magnitude of the effect of Fli-1 is promoter- and cell context-specific. We propose that Fli-1 inhibits COL1A2 promoter activity by competition with Ets-1. In addition, we postulate that another factor (co-repressor) may be required for maximal inhibition after recruitment to the Fli-1-Sp1 complex. We conclude that the ratio of Fli-1 to Ets-1 and the presence of co-regulatory proteins ultimately control ECM production in fibroblasts. PMID- 11278622 TI - MEKK2 is required for T-cell receptor signals in JNK activation and interleukin-2 gene expression. AB - The c-Jun N-terminal kinases (JNKs) are members of the mitogen-activated protein kinase (MAPK) gene family and are essential for cell proliferation, differentiation, and apoptosis. Previously we found that activation of JNK in T cells required costimulation of both T-cell receptor and auxiliary receptors such as CD28. In this study, we cloned a full-length human MEK kinase (MEKK) 2 cDNA from Jurkat T-cells and demonstrated that it was a major upstream MAPK kinase kinase for the JNK cascade in T-cells. The human MEKK2 cDNA encoded a polypeptide of 619 amino acids and was the human counterpart of the reported murine MEKK2. It was 94% homologous with human and murine MEKK3 at the catalytic domains and 60% homologous at the N-terminal noncatalytic region. Northern blot analysis showed that MEKK2 was ubiquitously expressed, with the highest level in peripheral blood leukocytes. In T cells, MEKK2 was found to be a strong activator of JNK but not of extracellular signal-regulated kinase MAPKs and to activate JNK-dependent AP-1 reporter gene expression. MEKK2 also synergized with anti-CD3 antibody to activate JNK in T cells, and stimulation of T cells led to induction of MEKK2 tyrosine phosphorylation. Significantly, the JNK activation induced by anti-CD3 and anti-CD28 antibodies, but not by 12-O-tetradecanoylphorbol-13-acetate and Ca(2+) ionophore A23187, was inhibited by dominant negative MEKK2 mutants. AP-1 and interleukin-2 reporter gene induction in T-cells was also inhibited by dominant negative MEKK2 mutants. Taken together, our results showed that human MEKK2 is a key signaling molecule for T-cell receptor/CD3-mediated JNK MAPK activation and interleukin-2 gene expression. PMID- 11278623 TI - Matrix-dependent proteolysis of surface transglutaminase by membrane-type metalloproteinase regulates cancer cell adhesion and locomotion. AB - Cell invasion requires cooperation between adhesion receptors and matrix metalloproteinases (MMPs). Membrane type (MT)-MMPs have been thought to be primarily involved in the breakdown of the extracellular matrix. Our report presents evidence that MT-MMPs in addition to the breakdown of the extracellular matrix may be engaged in proteolysis of adhesion receptors on tumor cell surfaces. Overexpression of MT1-MMP by glioma and fibrosarcoma cells led to proteolytic degradation of cell surface tissue transglutaminase (tTG) at the leading edge of motile cancer cells. In agreement, structurally related MT1-MMP, MT2-MMP, and MT3-MMP but not evolutionary distant MT4-MMP efficiently degraded purified tTG in vitro. Because cell surface tTG represents a ubiquitously expressed, potent integrin-binding adhesion coreceptor involved in the binding of cells to fibronectin (Fn), the proteolytic degradation of tTG by MT1-MMP specifically suppressed cell adhesion and migration on Fn. Reciprocally, Fn in vitro and in cultured cells protected its surface receptor, tTG, from proteolysis by MT1-MMP, thereby supporting cell adhesion and locomotion. In contrast, the proteolytic degradation of tTG stimulated migration of cells on collagen matrices. Together, our observations suggest both an important coreceptor role for cell surface tTG and a novel regulatory function of membrane-anchored MMPs in cancer cell adhesion and locomotion. Proteolysis of adhesion proteins colocalized with MT-MMPs at discrete regions on the surface of migrating tumor cells might be controlled by composition of the surrounding ECM. PMID- 11278624 TI - Inhibition of cytochrome c release in Fas-mediated signaling pathway in transgenic mice induced to express hepatitis C viral proteins. AB - Persistent hepatitis C virus (HCV) infection often progresses to chronic hepatitis, cirrhosis, and hepatocellular carcinoma. Numerous viruses have been reported to escape from apoptotic mechanism to maintain persistent infection. In the present study, we characterized the effect of HCV proteins on the Fas signal using HCV transgenic mice, which expressed core, E1, E2, and NS2 proteins, regulated by the Cre/loxP switching system. The transgene expression of HCV transgenic mice caused resistance to Fas antibody stimulated lethality. Apoptotic cell death in the liver of HCV protein expressing mice was significantly reduced compared with nonexpressing mice. Histopathological analysis and DNA fragmentation analysis revealed that the HCV proteins suppressed Fas-mediated apoptotic cell death. To identify the target pathway of HCV proteins, we characterized caspase activity. The activation of caspase-9 and -3/7 but not caspase-8 was inhibited by HCV proteins. Cytochrome c release from mitochondria was inhibited in HCV protein expressing mice. These results indicated that the expression of HCV proteins may directly or indirectly inhibit Fas-mediated apoptosis and death in mice by repressing the release of cytochrome c from mitochondria, thereby suppressing caspase-9 and -3/7 activation. These results suggest that HCV may cause persistent infection, as a result of suppression of Fas-mediated cell death. PMID- 11278625 TI - Cloning and characterization of ELL-associated proteins EAP45 and EAP20. a role for yeast EAP-like proteins in regulation of gene expression by glucose. AB - RNA polymerase II elongation factor ELL was recently purified from rat liver as a component of a multiprotein complex containing ELL and three ELL-associated proteins (EAPs) of approximately 45 (EAP45), approximately 30 (EAP30), and approximately 20 (EAP20) kDa (Shilatifard, A. (1998) J. Biol. Chem. 273, 11212 11217). Cloning of cDNA encoding the EAP30 protein revealed that it shares significant sequence similarity with the product of the Saccharomyces cerevisiae SNF8 gene (Schmidt, A. E., Miller, T., Schmidt, S. L., Shiekhattar, R., and Shilatifard, A. (1999) J. Biol. Chem. 274, 21981-21985), which is required for efficient derepression of glucose-repressed genes. Here we report the cloning of cDNAs encoding the EAP45 and EAP20 proteins. In addition, we identify the S. cerevisiae VPS36 and YJR102c genes as potential orthologs of EAP45 and EAP20 and show that they are previously uncharacterized SNF genes with properties very similar to SNF8. PMID- 11278626 TI - Two unrelated phosphoenolpyruvate carboxylase polypeptides physically interact in the high molecular mass isoforms of this enzyme in the unicellular green alga Selenastrum minutum. AB - In the chlorophyte Selenastrum minutum, phosphoenolpyruvate carboxylase (PEPC) exists as two kinetically distinct classes of isoforms sharing the same 102-kDa catalytic subunit (p102). Class 1 PEPC is homotetrameric, whereas Class 2 PEPCs consist of three large protein complexes. The different Class 2 PEPCs contain p102 and 130-, 73-, and 65-kDa polypeptides in different stoichiometric combinations. Immunoblot, immunoprecipitation, and chemical cross-linking studies indicated that p102 physically interacts with the 130-kDa polypeptide (p130) in Class 2 PEPCs. Immunological data and mass spectrometric and sequence analyses revealed that p102 and p130 are not closely related even if a p130 tryptic peptide had significant similarity to a conserved PEPC C-terminal domain from several sources. Evidence supporting the hypothesis that p130 has PEPC activity includes the following. (i) Specific activity expressed relative to the amount of p102 was lower in Class 1 than in Class 2 PEPCs; (ii) reductive pyridoxylation of both p102 and p130 was inhibited by magnesium-phosphoenolpyruvate; and (iii) biphasic phosphoenolpyruvate binding kinetics were observed with Class 2 PEPCs. These data support the view that unicellular green algae uniquely express, regulate, and assemble divergent PEPC polypeptides. This probably serves an adaptive purpose by poising these organisms for survival in different environments varying in nutrient content. PMID- 11278627 TI - ABCR, the ATP-binding cassette transporter responsible for Stargardt macular dystrophy, is an efficient target of all-trans-retinal-mediated photooxidative damage in vitro. Implications for retinal disease. AB - A large body of experimental and clinical data have documented the damaging effects of light exposure on photoreceptor cells although the identities of the biologically relevant molecular targets of photodamage are still uncertain. Several lines of evidence point to retinoids or retinoid derivatives as chromophores that can mediate light damage. We report here that ABCR, a photoreceptor-specific transporter involved in the recycling of all-trans retinal, is unusually sensitive to photooxidation damage mediated by all-trans retinal in vitro. Partial loss of ABCR function is responsible for Stargardt macular dystrophy, which is associated with accumulation of A2E, a diretinoid adduct within the retinal pigment epithelium. Photodamage to ABCR causes it to aggregate in SDS gels and results in the loss of retinal-stimulated ATPase activity. Peripherin/RDS and ROM-1, two structural proteins that colocalize with ABCR at the outer segment disc rim, are also significantly more susceptible to all-trans-retinal-mediated photodamage than are the major proteins from the rod outer segment. These observations imply that there may be specific protein targets of photodamage within the outer segment, and they may be especially relevant to assessing the risk of light exposure in those individuals who already have diminished ABCR activity due to mutation in one or both copies of the ABCR gene. PMID- 11278628 TI - Purification and characterization of human laminin-8. Laminin-8 stimulates cell adhesion and migration through alpha3beta1 and alpha6beta1 integrins. AB - Recently identified laminin isoforms containing the alpha4 chain have been shown to be expressed in the basement membrane of restricted organs such as heart, skeletal muscle, and blood vessels, especially those in embryos. We screened 38 human cell lines for the expression of the laminin alpha4 chain by reverse transcriptase-polymerase chain reaction and found that T98G glioblastoma cells express only alpha4, but not other alpha chains. Laminin-8, an isoform containing the alpha4 and beta1 chains, was purified from conditioned medium of T98G cells by gel filtration and immunoaffinity chromatography using a monoclonal antibody against laminin beta1 chain. The purified laminin isoform was composed of disulfide-linked 230-, 220-, and 200-kDa subunits, which immunoblot analysis identified as the beta1, gamma1, and alpha4 chains. Purified laminin-8 had cell adhesive activity comparable to laminin-1 but significantly weaker than laminin-5 and laminin-10/11. T98G cells adhering to laminin-8 became more elongated than those adhering to other laminin isoforms and extended multiple pseudopods. Cell adhesion to laminin-8 was abolished by an antibody against the integrin beta1 subunit or a combination of antibodies against the integrin alpha3 and alpha6 subunits, but not by either anti-alpha3 or anti-alpha6 antibody alone, suggesting that both alpha3beta1 and alpha6beta1 integrins serve as adhesion receptors for laminin-8. Consistent with these observations, K562 erythroleukemic cells transfected with either integrin alpha3 or alpha6 cDNA were capable of adhering to laminin-8 when beta1 integrins were stimulated by the beta1-activating antibody 8A2. Despite its moderate cell adhesive activity, laminin-8 was significantly potent in promoting cell migration when compared with other laminin isoforms and fibronectin. Cell migration on laminin-8 was completely inhibited by a combination of antibodies against alpha3 and alpha6 integrins, and substantially inhibited by anti-alpha3 antibody alone, suggesting that laminin-8 mediated cell migration is predominantly mediated by alpha3beta1 integrin. Given its potency to stimulate cell migration and preferential localization to the basement membrane of capillaries and embryonic tissues, laminin-8 may play a role in processes requiring enhanced cell migration during development, wound healing, and angiogenesis. PMID- 11278629 TI - Characterization of a powerful high affinity antagonist that inhibits biological activities of human interleukin-13. AB - Interleukin-13 (IL-13), a predominantly Th2-derived cytokine, appears to play a central pathological role in asthma, atopic dermatitis, allergic rhinitis, some parasitic infections, and cancer. We hypothesized that an IL-13 antagonist may have profound therapeutic utility in these conditions. We, therefore, mutagenized human IL-13 in which Glu at position 13 was substituted by a Lys residue. This highly purified recombinant IL-13 variant, IL-13E13K, bound with 4-fold higher affinity to the IL-13 receptor than wild-type IL-13 but retained no detectable proliferative activity on the TF-1 hematopoietic cell line. IL-13E13K competitively inhibited IL-13- and IL-4-dependent TF-1 proliferation. It also inhibited IL-13-induced STAT-6 (signal transduction and activator of transducer 6) activation in immune cells and cancer cells and reversed IL-13-induced inhibition of CD14 expression on human primary monocytes. These results demonstrate that high affinity binding and signal generation can be uncoupled efficiently in a ligand receptor interaction. These results also suggest that IL 13E13K may be a useful antagonist for the treatment of allergic, inflammatory, and parasitic diseases or even malignancies in which IL-13 plays a central role. PMID- 11278630 TI - Mechanism of dihydropyridine interaction with critical binding residues of L-type Ca2+ channel alpha 1 subunits. AB - We investigated the mechanism of interaction of individual L-type channel amino acid residues with dihydropyridines within a dihydropyridine-sensitive alpha1A subunit (alpha1A(DHP)). Mutation of individual residues in repeat III and expression in Xenopus oocytes revealed that Thr(1393) is not required for dihydropyridine interaction but that bulky side chains (tyrosine, phenylalanine) in this position sterically inhibit dihydropyridine coordination. In position 1397 a side chain carbonyl group was required for high antagonist sensitivity. Agonist function required the complete amide group of a glutamine residue. Val(1516) and Met(1512) side chains were required for agonist (Val(1516)) and antagonist (Val(1516), Met(1512)) sensitivity. Replacement of Ile(1504) and Ile(1507) by alpha1A phenylalanines was tolerated. Substitution of Thr(1393) by phenylalanine or Val(1516) by alanine introduced voltage dependence of antagonist action into alpha1A(DHP), suggesting that these residues form part of a mechanism mediating voltage dependence of dihydropyridine sensitivity. Our data provide important insight into dihydropyridine binding to alpha1A(DHP) which could facilitate the development of alpha1A-selective modulators. By modulating P/Q type Ca(2+) channels such drugs could serve as new anti-migraine therapeutics. PMID- 11278631 TI - A novel anti-angiogenic form of antithrombin with retained proteinase binding ability and heparin affinity. AB - Latent antithrombin, an inactive antithrombin form with low heparin affinity, has previously been shown to efficiently inhibit angiogenesis and tumor growth. We now show that heat treatment similar to that used for preparation of latent antithrombin also transforms antithrombin to another form, which we denote prelatent, with potent anti-angiogenic and anti-tumor activity but with retained proteinase- and heparin-binding properties. The ability of prelatent antithrombin to inhibit angiogenesis is presumably due to a limited conformational change, which may partially resemble that in latent antithrombin. Such a change is evidenced by a different cleavage pattern of prelatent than of native antithrombin by nontarget proteinases. Prelatent antithrombin exerts its anti angiogenic effect by a similar mechanism as latent antithrombin, i.e. by inhibiting focal adhesion formation and focal adhesion kinase activity, thereby leading to decreased proliferation of endothelial cells. The proteinase inhibitory fractions in commercial antithrombin preparations, which have been heat treated during production, also have anti-angiogenic activity, comparable with that of the prelatent antithrombin form. PMID- 11278632 TI - Class II recombinant phosphoribosyl diphosphate synthase from spinach. Phosphate independence and diphosphoryl donor specificity. AB - A recombinant form of spinach (Spinacia oleracea) phosphoribosyl diphosphate (PRPP) synthase isozyme 3 resembling the presumed mature enzyme has been synthesized in an Escherichia coli strain in which the endogenous PRPP synthase gene was deleted, and has been purified to near homogeneity. Contrary to other PRPP synthases the activity of spinach PRPP synthase isozyme 3 is independent of P(i), and the enzyme is inhibited by ribonucleoside diphosphates in a purely competitive manner, which indicates a lack of allosteric inhibition by these compounds. In addition spinach PRPP synthase isozyme 3 shows an unusual low specificity toward diphosphoryl donors by accepting dATP, GTP, CTP, and UTP in addition to ATP. The kinetic mechanism of the enzyme is an ordered steady state Bi Bi mechanism with K(ATP) and K(Rib-5-P) values of 170 and 110 micrometer, respectively, and a V(max) value of 13.1 micromol (min x mg of protein)(-1). The enzyme has an absolute requirement for magnesium ions, and maximal activity is obtained at 40 degrees C at pH 7.6. PMID- 11278633 TI - Identification of the binding site for fibrinogen recognition peptide gamma 383 395 within the alpha(M)I-domain of integrin alpha(M)beta2. AB - The leukocyte integrin alpha(M)beta(2) (Mac-1, CD11b/CD18) is a cell surface adhesion receptor for fibrinogen. The interaction between fibrinogen and alpha(M)beta(2) mediates a range of adhesive reactions during the immune inflammatory response. The sequence gamma(383)TMKIIPFNRLTIG(395), P2-C, within the gamma-module of the D-domain of fibrinogen, is a recognition site for alpha(M)beta(2) and alpha(X)beta(2). We have now identified the complementary sequences within the alpha(M)I-domain of the receptor responsible for recognition of P2-C. The strategy to localize the binding site for P2-C was based on distinct P2-C binding properties of the three structurally similar I-domains of alpha(M)beta(2), alpha(X)beta(2), and alpha(L)beta(2), i.e. the alpha(M)I- and alpha(X)I-domains bind P2-C, and the alpha(L)I-domain did not bind this ligand. The Lys(245)-Arg(261) sequence, which forms a loop betaD-alpha5 and an adjacent helix alpha5 in the three-dimensional structure of the alpha(M)I-domain, was identified as the binding site for P2-C. This conclusion is supported by the following data: 1) mutant cell lines in which the alpha(M)I-domain segments (245)KFG and Glu(253)-Arg(261) were switched to the homologous alpha(L)I-domain segments failed to support adhesion to P2-C; 2) synthetic peptides duplicating the Lys(245)-Tyr(252) and Glu(253)-Arg(261) sequences directly bound the D fragment and P2-C derivative, gamma384-402, and this interaction was blocked efficiently by the P2-C peptide; 3) mutation of three amino acid residues within the Lys(245)-Arg(261) segment, Phe(246), Asp(254), and Pro(257), resulted in the loss of the binding function of the recombinant alpha(M)I-domains; and 4) grafting the alpha(M)(Lys(245)-Arg(261)) segment into the alpha(L)I-domain converted it to a P2-C-binding protein. These results demonstrate that the alpha(M)(Lys(245)-Arg(261)) segment, a site of the major sequence and structure difference among alpha(M)I-, alpha(X)I-, and alpha(L)I-domains, is responsible for recognition of a small segment of fibrinogen, gammaThr(383)-Gly(395), by serving as ligand binding site. PMID- 11278634 TI - Apoptosis in motion. An apical, P35-insensitive caspase mediates programmed cell death in insect cells. AB - Activation of caspases by proteolytic processing is a critical step during apoptosis in metazoans. Here we use high resolution time lapse microscopy to show a tight link between caspase activation and the morphological events delineating apoptosis in cultured SF21 cells from the moth Spodoptera frugiperda, a model insect system. The principal effector caspase, Sf-caspase-1, is proteolytically activated during SF21 apoptosis. To define the potential role of initiator caspases in vivo, we tested the effect of cell-permeable peptide inhibitors on pro-Sf-caspase-1 processing. Anti-caspase peptide analogues prevented apoptosis induced by diverse signals, including UV radiation and baculovirus infection. IETD-fmk potently inhibited the initial processing of pro-Sf-caspase-1 at the junction (TETD-G) of the large and small subunit, a cleavage that is blocked by inhibitor of apoptosis Op-IAP but not pancaspase inhibitor P35. Because Sf caspase-1 was inhibited poorly by IETD-CHO, our data indicated that the protease responsible for the first step in pro-Sf-caspase-1 activation is a distinct apical caspase. Thus, Sf-caspase-1 activation is mediated by a novel, P35 resistant caspase. These findings support the hypothesis that apoptosis in insects, like that in mammals, involves a cascade of caspase activations. PMID- 11278635 TI - Ligand-dependent formation of retinoid receptors, receptor-interacting protein 140 (RIP140), and histone deacetylase complex is mediated by a novel receptor interacting motif of RIP140. AB - Receptor-interacting protein 140 (RIP140) interacts with retinoic acid receptor and retinoid X receptor in a ligand-dependent manner and suppresses retinoic acid (RA) induction of its target genes. The receptor-interacting motif is mapped to a C-terminal peptide sequence (LTKTNPILYYMLQK) of RIP140. The functional role of this motif in mediating the suppressive effects of RIP140 on RA induction is demonstrated in mutation studies. RA induces coimmunoprecipitation of histone deacetylase 3 with retinoic acid receptor/retinoid X receptor in the presence of wild type RIP140, but not in the presence of the C-terminal motif-deleted RIP140. A decrease in histone acetylation on the promoter region that carries a RA response element is associated with the expression of wild type RIP140, but not with expression of the mutant RIP140, in a dose-dependent manner. These data provide a molecular explanation for RIP140 acting as a novel ligand-dependent, negative modulator of RA-regulated gene expression. PMID- 11278637 TI - Activity of pulmonary surfactant protein-D (SP-D) in vivo is dependent on oligomeric structure. AB - Pulmonary surfactant protein-D (SP-D) is a member of the collectin family of C type lectins that is synthesized in many tissues including respiratory epithelial cells in the lung. SP-D is assembled predominantly as dodecamers consisting of four homotrimeric subunits each. Association of these subunits is stabilized by interchain disulfide bonds involving two conserved amino-terminal cysteine residues (Cys-15 and Cys-20). Mutant recombinant rat SP-D lacking these residues (RrSP-Dser15/20) is secreted in cell culture as trimeric subunits rather than as dodecamers. In this study, transgenic mice that express this mutant were generated to elucidate the functional importance of SP-D oligomerization in vivo. Expression of RrSP-Dser15/20 failed to correct the pulmonary phospholipid accumulation and emphysema characteristic of SP-D null (mSP-D-/-) mice. Expression of high concentrations of the mutant protein in wild-type mice reduced the abundance of disulfide cross-linked oligomers of endogenous SP-D in the bronchoalveolar lavage fluid and demonstrated a phenotype that partially overlapped with that of the SP-D-/- mice; the animals developed emphysema and foamy macrophages without the associated abnormalities in alveolar phospholipids typical of SP-D-/- mice. Development of foamy macrophages in SP-D-deficient mice is not secondary to the increased abundance of surfactant phospholipids. Disulfide cross-linked SP-D oligomers are required for the regulation of surfactant phospholipid homeostasis and the prevention of emphysema and foamy macrophages in vivo. PMID- 11278638 TI - Growth hormone regulates phosphorylation and function of CCAAT/enhancer-binding protein beta by modulating Akt and glycogen synthase kinase-3. AB - Growth hormone (GH) regulates transcription factors associated with c-fos, including C/EBPbeta. Two forms of C/EBPbeta, liver-activating protein (LAP) and liver inhibitory protein (LIP), are dephosphorylated in GH-treated 3T3-F442A fibroblasts. GH-induced dephosphorylation of LAP and LIP is reduced when cells are preincubated with phosphatidylinositol 3'-kinase (PI3K) inhibitors. GH activates Akt and inhibits glycogen synthase kinase-3 (GSK-3). Lithium, a GSK-3 inhibitor, increases GH-dependent dephosphorylation of LAP and LIP. Both are in vitro substrates of GSK-3, suggesting that GSK-3 inactivation contributes to GH promoted dephosphorylation of C/EBPbeta. Alkaline phosphatase increases binding of LAP homodimers and decreases binding of LIP homodimers to c-fos, suggesting that dephosphorylation of C/EBPbeta modifies their ability to bind DNA. Both alkaline phosphatase- and GH-mediated dephosphorylation comparably increase binding of endogenous LAP in 3T3-F442A cells. In cells overexpressing LAP and GSK 3, LAP binding decreases, suggesting that GSK-3-mediated phosphorylation interferes with LAP binding. Expression of constitutively active GSK-3 reduced GH stimulated c-fos promoter activity. These studies indicate that PI3K/Akt/GSK-3 mediates signaling between GH receptor and the nucleus, promoting dephosphorylation of C/EBPbeta. Dephosphorylation increases binding of LAP complexes to the c-fos promoter and may contribute to the participation of C/EBPbeta in GH-stimulated c-fos expression. PMID- 11278639 TI - Potent blockade of hepatocyte growth factor-stimulated cell motility, matrix invasion and branching morphogenesis by antagonists of Grb2 Src homology 2 domain interactions. AB - Hepatocyte growth factor (HGF) stimulates mitogenesis, motogenesis, and morphogenesis in a wide range of cellular targets during development, homeostasis and tissue regeneration. Inappropriate HGF signaling occurs in several human cancers, and the ability of HGF to initiate a program of protease production, cell dissociation, and motility has been shown to promote cellular invasion and is strongly linked to tumor metastasis. Upon HGF binding, several tyrosines within the intracellular domain of its receptor, c-Met, become phosphorylated and mediate the binding of effector proteins, such as Grb2. Grb2 binding through its SH2 domain is thought to link c-Met with downstream mediators of cell proliferation, shape change, and motility. We analyzed the effects of Grb2 SH2 domain antagonists on HGF signaling and observed potent blockade of cell motility, matrix invasion, and branching morphogenesis, with ED(50) values of 30 nm or less, but only modest inhibition of mitogenesis. These compounds are 1000 10,000-fold more potent anti-motility agents than any previously characterized Grb2 SH2 domain antagonists. Our results suggest that SH2 domain-mediated c-Met Grb2 interaction contributes primarily to the motogenic and morphogenic responses to HGF, and that these compounds may have therapeutic application as anti metastatic agents for tumors where the HGF signaling pathway is active. PMID- 11278640 TI - Identification of amino acid residues in the ETS transcription factor Erg that mediate Erg-Jun/Fos-DNA ternary complex formation. AB - Jun, Fos, and Ets proteins belong to distinct families of transcription factors that target specific DNA elements often found jointly in gene promoters. Physical and functional interactions between these families play important roles in modulating gene expression. Previous studies have demonstrated a direct interaction between the DNA-binding domains of the two partners. However, the molecular details of the interactions have not been investigated so far. Here we used the known three-dimensional structures of the ETS DNA-binding domain and Jun/Fos heterodimer to model an ETS-Jun/Fos-DNA ternary complex. Docking procedures suggested that certain ETS domain residues in the DNA recognition helix alpha3 interact with the N-terminal basic domain of Jun. To support the model, different Erg ETS domain mutants were obtained by deletion or by single amino acid substitutions and were tested for their ability to mediate DNA binding, Erg-Jun/Fos complex formation, and transcriptional activation. We identified point mutations that affect both the DNA binding properties of Erg and its physical interaction with Jun (R367K), as well as mutations that essentially prevent transcriptional synergy with the Jun/Fos heterodimer (Y371V). These results provide a framework of the ETS/bZIP interaction linked to the manifestation of functional activity in gene regulation. PMID- 11278641 TI - Identification and characterization of a conformational heparin-binding site involving two fibronectin type III modules of bovine tenascin-X. AB - Tenascin-X is known as a heparin-binding molecule, but the localization of the heparin-binding site has not been investigated until now. We show here that, unlike tenascin-C, the recombinant fibrinogen-like domain of tenascin-X is not involved in heparin binding. On the other hand, the two contiguous fibronectin type III repeats b10 and b11 have a predicted positive charge at physiological pH, hence a set of recombinant proteins comprising these domains was tested for interaction with heparin. Using solid phase assays and affinity chromatography, we found that interaction with heparin was conformational and involved both domains 10 and 11. Construction of a three-dimensional model of domains 10 and 11 led us to predict exposed residues that were then submitted to site-directed mutagenesis. In this way, we identified the basic residues within each domain that are crucial for this interaction. Blocking experiments using antibodies against domain 10 were performed to test the efficiency of this site within intact tenascin-X. Binding was significantly reduced, arguing for the activity of a heparin-binding site involving domains 10 and 11 in the whole molecule. Finally, the biological significance of this site was tested by cell adhesion studies. Heparan sulfate cell surface receptors are able to interact with proteins bearing domains 10 and 11, suggesting that tenascin-X may activate different signals to regulate cell behavior. PMID- 11278642 TI - The role of water molecules in the association of cytochrome P450cam with putidaredoxin. An osmotic pressure study. AB - We have investigated the osmotic pressure dependence of the association between ferric cytochrome P450cam and putidaredoxin (Pdx) to gain an insight into the role of water molecules in the P450cam-reduced Pdx complexation amenable to physiological electron transfer. The association constant was evaluated from the electron transfer rates from reduced Pdx to P450cam. The natural logarithm of the association constant K(a) was linearly reduced by the osmotic pressure, and osmotic stress yields uptake of 25 waters upon association. In contrast, uptake of only 13 waters is observed from the osmotic pressure dependence of the association in the nonphysiological redox partners P450cam and oxidized Pdx. Although general protein-protein associations proceed through dehydration around the complex interface, the interfacial waters could mediate hydrogen-bonding interactions. Therefore, about 10 more interfacial waters imply an additional water-mediated hydrogen-bonding network in the P450cam.reduced Pdx complex, which does not exist in the complex with oxidized Pdx. It is also possible that the water-mediated hydrogen-bonding interactions support a high P450cam affinity for reduced (K(a) = 0.83 microm(-1)) relative to oxidized (K(a) = 0.058 microm(-1)) Pdx. This study points to a novel role of solvents in assisting redox state dependent interaction between P450cam and Pdx. PMID- 11278643 TI - Calcium influx and signaling in yeast stimulated by intracellular sphingosine 1 phosphate accumulation. AB - In mammalian cells, intracellular sphingosine 1-phosphate (S1P) can stimulate calcium release from intracellular organelles, resulting in the activation of downstream signaling pathways. The budding yeast Saccharomyces cerevisiae expresses enzymes that can synthesize and degrade S1P and related molecules, but their possible role in calcium signaling has not yet been tested. Here we examine the effects of S1P accumulation on calcium signaling using a variety of yeast mutants. Treatment of yeast cells with exogenous sphingosine stimulated Ca(2+) accumulation through two distinct pathways. The first pathway required the Cch1p and Mid1p subunits of a Ca(2+) influx channel, depended upon the function of sphingosine kinases (Lcb4p and Lcb5p), and was inhibited by the functions of S1P lyase (Dpl1p) and the S1P phosphatase (Lcb3p). The biologically inactive stereoisomer of sphingosine did not activate this Ca(2+) influx pathway, suggesting that the active S1P isomer specifically stimulates a calcium-signaling mechanism in yeast. The second Ca(2+) influx pathway stimulated by the addition of sphingosine was not stereospecific, was not dependent on the sphingosine kinases, occurred only at higher doses of added sphingosine, and therefore was likely to be nonspecific. Mutants lacking both S1P lyase and phosphatase (dpl1 lcb3 double mutants) exhibited constitutively high Ca(2+) accumulation and signaling in the absence of added sphingosine, and these effects were dependent on the sphingosine kinases. These results show that endogenous S1P-related molecules can also trigger Ca(2+) accumulation and signaling. Several stimuli previously shown to evoke calcium signaling in wild-type cells were examined in lcb4 lcb5 double mutants. All of the stimuli produced calcium signals independent of sphingosine kinase activity, suggesting that phosphorylated sphingoid bases might serve as messengers of calcium signaling in yeast during an unknown cellular response. PMID- 11278644 TI - A dominant negative mutant of Bacillus anthracis protective antigen inhibits anthrax toxin action in vivo. AB - PA63, a proteolytically activated 63-kDa form of anthrax protective antigen (PA), forms heptameric oligomers and has the ability to bind and translocate the catalytic moieties, lethal factor (LF), and edema factor (EF) into the cytosol of mammalian cells. Acidic pH triggers oligomerization and membrane insertion by PA63. A disordered amphipathic loop in domain II of PA (2beta2-2beta3 loop) is involved in membrane insertion by PA63. Because conditions required for membrane insertion coincide with those for oligomerization of PA63 in mammalian cells, residues constituting the 2beta2-2beta3 loop were replaced with the residues of the amphipathic membrane-inserting loop of its homologue iota-b toxin secreted by Clostridium perfringens. It was hypothesized that such a molecule might assemble into hetero-heptameric structures with wild-type PA ultimately leading to the inhibition of cellular intoxication. The mutation blocked the ability of PA to mediate membrane insertion and translocation of LF into the cytosol but had no effect on proteolytic activation, oligomerization, or binding LF. Moreover, an equimolar mixture of purified mutant PA (PA-I) and wild-type PA showed complete inhibition of toxin activity both in vitro on J774A.1 cells and in vivo in Fischer 344 rats thereby exhibiting a dominant negative effect. In addition, PA-I inhibited the channel-forming ability of wild-type PA on the plasma membrane of CHO-K1 cells thereby indicating protein-protein interactions between the two proteins resulting in the formation of mixed oligomers with defective functional activity. Our findings provide a basis for understanding the mechanism of translocation and exploring the possibility of the use of this PA molecule as a therapeutic agent against anthrax toxin action in vivo. PMID- 11278645 TI - Antagonism between PTEN/MMAC1/TEP-1 and androgen receptor in growth and apoptosis of prostatic cancer cells. AB - PTEN/MMAC1/TEP-1 (PTEN) tumor suppressor and androgen receptor play important roles in prostatic tumorigenesis by exerting opposite effects on homeostasis of prostatic epithelium. Here, we describe a mutual repression and selective dominance between PTEN and the androgen receptor (AR) in the growth and the apoptosis of prostatic cancer cells. On the one hand, PTEN and an inhibitor of phosphoinositide 3-kinase repressed the transcriptional activity of the AR as well as androgen-induced cell proliferation and production of prostate-specific antigen. On the other hand, androgens protected prostate cancer cells from PTEN induced apoptosis in an AR-dependent manner. Whereas the repression of the transcriptional activity of the AR by PTEN is likely to involve the down regulation of AKT, androgens protected prostate cancer cells from PTEN-induced apoptosis without an effect on AKT activity, demonstrating a differential involvement of AKT in the interaction between PTEN and the AR. Our data suggest that the loss of PTEN function may induce tumorigenesis through unopposed activity of the AR as well as contribute to the resistance of prostate cancers to androgen ablation therapy. PMID- 11278646 TI - Effect of up-regulating individual steps in the reverse cholesterol transport pathway on reverse cholesterol transport in normolipidemic mice. AB - Cholesterol acquired by extrahepatic tissues (from de novo synthesis or lipoproteins) is returned to the liver for excretion in a process called reverse cholesterol transport (RCT). We undertook studies to determine if RCT could be enhanced by up-regulating individual steps in the RCT pathway. Overexpression of 7alpha-hydroxylase, Scavenger receptor B1, lecithin:cholesterol acyltransferase (LCAT), or apoA-I in the liver did not stimulate cholesterol efflux from any extrahepatic tissue. In contrast, infusion of apoA-I.phospholipid complexes (rHDL) that resemble nascent HDL markedly stimulated cholesterol efflux from tissues into plasma. Cholesterol effluxed to rHDL was initially unesterified but by 24 h this cholesterol was largely esterified and had shifted to normal HDL (in mice lacking cholesteryl ester transfer protein) or to apoB containing lipoproteins (in cholesteryl ester transfer protein transgenic mice). Most of the cholesterol effluxed into plasma in response to rHDL came from the liver. However, an even greater proportion of effluxed cholesterol was cleared by the liver resulting in a transient increase in liver cholesterol concentrations. Fecal sterol excretion was not increased by rHDL. Thus, although rHDL stimulated cholesterol efflux from most tissues and increased net cholesterol movement from extrahepatic tissues to the liver, cholesterol flux through the entire RCT pathway was not increased. PMID- 11278647 TI - Tumor suppressor p53 protein is a new target for the metastasis-associated Mts1/S100A4 protein: functional consequences of their interaction. AB - A physical and functional interaction between the Ca(2+)-binding protein Mts1 (S100A4) and the tumor suppressor p53 protein is shown here for the first time. We demonstrate that Mts1 binds to the extreme end of the C-terminal regulatory domain of p53 by several in vitro and in vivo approaches: co-immunoprecipitation, affinity chromatography, and far Western blot analysis. The Mts1 protein in vitro inhibits phosphorylation of the full-length p53 and its C-terminal peptide by protein kinase C but not by casein kinase II. The Mts1 binding to p53 interferes with the DNA binding activity of p53 in vitro and reporter gene transactivation in vivo, and this has a regulatory function. A differential modulation of the p53 target gene (p21/WAF, bax, thrombospondin-1, and mdm-2) transcription was observed upon Mts1 induction in tet-inducible cell lines expressing wild type p53. Mts1 cooperates with wild type p53 in apoptosis induction. Our data imply that the ability of Mts1 to enhance p53-dependent apoptosis might accelerate the loss of wild type p53 function in tumors. In this way, Mts1 can contribute to the development of a more aggressive phenotype during tumor progression. PMID- 11278648 TI - Calmodulin enhances the stability of the estrogen receptor. AB - The estrogen receptor mediates breast cell proliferation and is the principal target for chemotherapy of breast carcinoma. Previous studies have demonstrated that the estrogen receptor binds to calmodulin-Sepharose in vitro. However, the association of endogenous calmodulin with endogenous estrogen receptors in intact cells has not been reported, and the function of the interaction is obscure. Here we demonstrate by co-immunoprecipitation from MCF-7 human breast epithelial cells that endogenous estrogen receptors bind to endogenous calmodulin. Estradiol treatment of the cells had no significant effect on the interaction. However, incubation of the cells with tamoxifen enhanced by 5-10-fold the association of calmodulin with the estrogen receptor and increased the total cellular content of estrogen receptors by 1.5-2-fold. In contrast, the structurally distinct calmodulin antagonists trifluoperazine and CGS9343B attenuated the interaction between calmodulin and the estrogen receptor and dramatically reduced the number of estrogen receptors in the cell. Neither of these agents altered the amount of estrogen receptor mRNA, suggesting that calmodulin stabilizes the protein. This hypothesis is supported by the observation that, in the presence of Ca2+, calmodulin protected estrogen receptors from in vitro proteolysis by trypsin. Furthermore, overexpression of wild type calmodulin, but not a mutant calmodulin incapable of binding Ca2+, increased the concentration of estrogen receptors in MCF-7 cells, whereas transient expression of a calmodulin inhibitor peptide reduced the estrogen receptor concentration. These data demonstrate that calmodulin binds to the estrogen receptor in intact cells in a Ca2+-dependent, but estradiol-independent, manner, thereby modulating the stability and the steady state level of estrogen receptors. PMID- 11278649 TI - Inhibition of interleukin-4- and CD40-induced IgE germline gene promoter activity by 2'-aminoethoxy-modified triplex-forming oligonucleotides. AB - Elevated levels of IgE are intimately associated with a number of allergic diseases, such as allergic rhinitis or asthma. Therefore, prevention of IgE production in human B-cells represents an attractive therapeutic target. IL-4 induced IgE germline gene transcription represents a crucial early step during IgE isotype switch differentiation. Gene induction is orchestrated by the coordinated action of the transcription factors STAT6 (signal transducer and activator of transcription), NF-kappaB, PU.1, and C/EBP. This study shows that 2' aminoethoxy-modified oligonucleotides, which partially overlap with the STAT6 and the adjacent PU.1/NF-kappaB binding site, inhibit DNA binding of all three proteins with high affinity in a dose- and time-dependent fashion in vitro. Loss of protein binding correlated strongly with increasing DNA triplex formation. Importantly, the oligomers also effectively displaced pre-bound recombinant NF kappaB p50 from double-stranded DNA in vitro. Functionally, the oligonucleotides led to a selective inhibition of IL-4-induced reporter gene activity from a construct driven by the IgE germline gene promoter in human B-cells. These data confirm the critical role of this cytokine-responsive regulatory region in IgE germline gene induction and further support the concept of specific modulation of gene expression by DNA triplex formation induced with chemically modified oligonucleotides. PMID- 11278650 TI - Mutational analysis of the fractalkine chemokine domain. Basic amino acid residues differentially contribute to CX3CR1 binding, signaling, and cell adhesion. AB - Fractalkine (FKN/CX3CL1) is a unique member of the chemokine gene family and contains a chemokine domain (CD), a mucin-like stalk, a single transmembrane region, and a short intracellular C terminus. This structural distinction affords FKN the property of mediating capture and firm adhesion of FKN receptor (CX3CR1) expressing cells under physiological flow conditions. Shed forms of FKN also exist, and these promote chemotaxis of CX3CR1-expressing leukocytes. The goal of the present study was to identify specific residues within the FKN-CD critical for FKN-CX3CR1 interactions. Two residues were identified in the FKN-CD, namely Lys-7 and Arg-47, that are important determinants in mediating an FKN-CX3CR1 interaction. FKN-K7A and FKN-R47A mutants exhibited 30-60-fold decreases in affinity for CX3CR1 and failed to arrest efficiently CX3CR1-expressing cells under physiological flow conditions. However, these mutants had differential effects on chemotaxis of CX3CR1-expressing cells. The FKN-K7A mutant acted as an equipotent partial agonist, whereas the FKN-R47A mutant had marked decreased potency and efficacy in measures of chemotactic activity. These data identify specific structural features of the FKN-CD that are important in interactions with CX3CR1 including steady state binding, signaling, and firm adhesion of CX3CR1-expressing cells. PMID- 11278651 TI - Cloning and characterization of LUN, a novel ring finger protein that is highly expressed in lung and specifically binds to a palindromic sequence. AB - We isolated cDNAs encoding a novel RING finger protein (LUN), the mRNAs of which were expressed at high levels in the lung. In situ hybridization revealed that LUN mRNAs were expressed in the alveolar epithelium of the lung. The LUN gene locus was assigned to chromosome 9p21, which contains candidate tumor suppressor genes associated with loss of heterozygosity in more than 86% of small cell lung cancers. We clarified that LUN is localized to the nucleus and reveals Zn(2+) dependent DNA binding activity. The region from amino acids 51 to 374 of LUN is responsible for DNA binding. Furthermore, we identified a novel palindromic binding consensus (5'-TCCCAGCACTTTGGGA-3') for the LUN binding. Interestingly, this LUN binding palindromic sequence is found in the upstream transcriptional regulatory region of the E-cadherin gene and two intervening regions of the talin gene. Our results suggested that LUN might be an important trans-acting transcriptional regulator for lung cancer-associated genes including E-cadherin and talin genes. PMID- 11278652 TI - An inhibitory role of Rho in the vasopressin-mediated translocation of aquaporin 2 into cell membranes of renal principal cells. AB - Vasopressin regulates water reabsorption in renal collecting duct principal cells by a cAMP-dependent translocation of the water channel aquaporin-2 (AQP2) from intracellular vesicles into the cell membrane. In the present work primary cultured inner medullary collecting duct cells were used to study the role of the proteins of the Rho family in the translocation of AQP2. Clostridium difficile toxin B, which inhibits all members of the Rho family, Clostridium limosum C3 toxin, which inactivates only Rho, and the Rho kinase inhibitor, Y-27632, induced both depolymerization of actin stress fibers and AQP2 translocation in the absence of vasopressin. The data suggest an inhibitory role of Rho in this process, whereby constitutive membrane localization is prevented in resting cells. Expression of constitutively active RhoA induced formation of actin stress fibers and abolished AQP2 translocation in response to elevation of intracellular cAMP, confirming the inhibitory role of Rho. Cytochalasin D induced both depolymerization of the F-actin cytoskeleton and AQP2 translocation, indicating that depolymerization of F-actin is sufficient to induce AQP2 translocation. Thus Rho is likely to control the intracellular localization of AQP2 via regulation of the F-actin cytoskeleton. PMID- 11278653 TI - Characterization of mutant neutrophil elastase in severe congenital neutropenia. AB - Severe congenital neutropenia is a heritable human disorder characterized by neutropenia and acute myelogenous leukemia. We recently determined that the majority of cases result from de novo or autosomal dominantly inherited heterozygous mutations in ELA2, encoding neutrophil elastase. Neutrophil elastase is a chymotryptic serine protease localized in granules of neutrophils and monocytes and is the major target of inhibition of the serpin alpha(1) antitrypsin. The mutations causing severe congenital neutropenia consist of amino acid missense substitutions, in-frame deletion, splice donor mutation producing a deletion, splice acceptor mutation causing insertion of novel residues, and protein truncating mutations of the carboxyl terminus resulting from nonsense substitutions and deletions leading to frameshifts. We have expressed 14 mutant forms of neutrophil elastase in vitro and have characterized their biochemical properties. The mutations have variable effects on proteolytic activity, eliminating the possibility that the disease results from haploinsufficiency. There is no evidence that the mutant enzymes are cytotoxic. The mutant enzymes retain vulnerability to inhibition by alpha(1)-antitrypsin, but demonstrate variable avidity for interaction with this serpin. Somewhat surprisingly, the mutant enzymes inhibit the wild type enzyme when both are coexpressed within the same cell, suggesting the potential to interfere with normal subcellular trafficking or post-translational processing. PMID- 11278654 TI - Palmitate-induced apoptosis can occur through a ceramide-independent pathway. AB - Cytotoxic accumulation of long chain fatty acids has been proposed to play an important role in the pathogenesis of diabetes mellitus and heart disease. To explore the mechanism of cellular lipotoxicity, we cultured Chinese hamster ovary cells in the presence of media supplemented with fatty acid. The saturated fatty acid palmitate, but not the monounsaturated fatty acid oleate, induced programmed cell death as determined by annexin V positivity, caspase 3 activity, and DNA laddering. De novo ceramide synthesis increased 2.4-fold with palmitate supplementation; however, this was not required for palmitate-induced apoptosis. Neither biochemical nor genetic inhibition of de novo ceramide synthesis arrested apoptosis in Chinese hamster ovary cells in response to palmitate supplementation. Rather, our data suggest that palmitate-induced apoptosis occurs through the generation of reactive oxygen species. Fluorescence of an oxidant sensitive probe was increased 3.5-fold with palmitate supplementation indicating that production of reactive intermediates increased. In addition, palmitate induced apoptosis was blocked by pyrrolidine dithiocarbamate and 4,5-dihydroxy 1,3-benzene-disulfonic acid, two compounds that scavenge reactive intermediates. These studies suggest that generation of reactive oxygen species, independent of ceramide synthesis, is important for the lipotoxic response and may contribute to the pathogenesis of diseases involving intracellular lipid accumulation. PMID- 11278655 TI - 10E4 antigen of Scrapie lesions contains an unusual nonsulfated heparan motif. AB - The carbohydrate antigen on heparan sulfate recognized by monoclonal antibody 10E4 is uniquely codistributed with the abnormal prion protein, PrP(Sc), even in the earliest detectable brain lesions of scrapie-infected mice. Determining the chemical structure of 10E4 antigen is, therefore, an important aspect of structure elucidation of scrapie lesions, and a prerequisite for designing experiments to understand its role in scrapie pathogenesis. Toward this aim, we have examined preparations of heparan sulfate, with differing sulfate contents, for binding by 10E4 antibody. The highest antigenicity was observed in a preparation (HS-1) with the lowest sulfate content. HS-1 was partially depolymerized with heparin lyase III, and oligosaccharide fragments examined for 10E4 antigen expression by the neoglycolipid technology. An antigen-positive and two antigen-negative tetrasaccharides were isolated and examined by electrospray mass spectrometry. The antigen-positive tetrasaccharide sequence on heparan sulfate was thus deduced to contain a unique unsulfated motif that includes an N unsubstituted glucosamine in the sequence, UA-GlcN-UA-GlcNAc. Antibody binding experiments with neoglycolipids prepared from a series of heparin/heparan sulfate disaccharides, and the trisaccharide derived from the antigen-positive tetrasaccharide after removal of the terminal hexuronic acid, show that both the penultimate glucosamine and the outer nonsulfated hexuronic acid are important for 10E4 antigenicity. PMID- 11278656 TI - Particle formation by a conserved domain of the herpes simplex virus protein VP22 facilitating protein and nucleic acid delivery. AB - VP22, a structural protein of herpes simplex virus, exhibits unusual trafficking properties which we proposed might be exploited in gene and protein delivery applications. To pursue the use of the protein itself for cargo delivery into cells, we developed an expression system for the C-terminal half of VP22, residues 159-301 (VP22.C1), and purified the protein in high yields. Addition of short oligonucleotides (ODNs) induced the assembly of novel particles, which were regular spheres with a size range of 0.3 to 1.0 microm in diameter, incorporating both protein and ODN. Following the particles in living cells using fluorescently tagged ODNs, we show that they enter efficiently within 2-4 h, and reside stably in the cell cytoplasm for up to several days. Remarkably, however, light activation induced particle disruption and release of the protein and ODN to the nucleus and cytoplasm within seconds, a process that we have captured by time lapse microscopy. In addition to delivering antisense ODNs, ribozymes, and RNA/DNA hybrids, the VP22.C1 protein could also be modified to include peptides or proteins. These particles have the potential for delivery of a wide range of therapeutic agents in gene therapy and vaccine development. PMID- 11278657 TI - The hairpin loop but not the bulged C of the iron responsive element is essential for high affinity binding to iron regulatory protein-1. AB - Vertebrates control intracellular iron concentration principally through the interaction of iron regulatory proteins with mRNAs that contain an iron responsive element, a small hairpin with a bulged C. The hairpin loop and bulged C have previously been assumed to be critical for binding and have been proposed to make direct contact with the iron regulatory proteins. However, we show here that a U or G can be substituted for the bulged C provided that specific nucleotides are also present within internal loops. The K(d), IC(50) and chemical modifications of the iron responsive element variants are similar to the wild type. Results are more consistent with a role in which the C-bulge functions to orient the hairpin for optimal protein binding rather than to directly contact the protein. Characterization of these novel iron responsive element variants may facilitate the identification of additional mRNAs whose expression is controlled by iron regulatory proteins, as well as provide insight into the nature of a critical RNA-protein interaction. PMID- 11278658 TI - Escherichia coli ykfE ORFan gene encodes a potent inhibitor of C-type lysozyme. AB - The complete nucleotide sequences of over 37 microbial and three eukaryote genomes are already publicly available, and more sequencing is in progress. Despite this accumulation of data, newly sequenced microbial genomes continue to reveal up to 50% of functionally uncharacterized "anonymous" genes. A majority of these anonymous proteins have homologues in other organisms, whereas the rest exhibit no clear similarity to any other sequence in the data bases. This set of unique, apparently species-specific, sequences are referred to as ORFans. The biochemical and structural analysis of ORFan gene products is of both evolutionary and functional interest. Here we report the cloning and expression of Escherichia coli ORFan ykfE gene and the functional characterization of the encoded protein. Under physiological conditions, the protein is a homodimer with a strong affinity for C-type lysozyme, as revealed by co-purification and co crystallization. Activity measurements and fluorescence studies demonstrated that the YkfE gene product is a potent C-type lysozyme inhibitor (K(i) approximately 1 nm). To denote this newly assigned function, ykfE has now been registered under the new gene name Ivy (inhibitor of vertebrate lysozyme) at the E. coli genetic stock center. PMID- 11278659 TI - Mechanism of cyclosporin-induced inhibition of intracellular collagen degradation. AB - The immunosuppressant cyclosporin A (CsA) markedly inhibits collagen degradation by an intracellular phagocytic pathway in fibroblasts, an effect that can lead to massive gingival overgrowth. We used a collagen bead model of collagen phagocytosis to determine whether CsA inhibits internalization by blocking efflux of calcium from endoplasmic reticulum (ER) and mitochondrial calcium stores. CsA caused dose-dependent inhibition of phagocytosis of collagen-coated (but not bovine serum albumin-coated) beads. Chelation of intracellular Ca(2+) with BAPTA/AM or inhibition of Ca(2+)-ATPase of ER stores with thapsigargin reduced collagen bead phagocytosis. Measurement of intracellular calcium by ratio fluorometry showed increases in response to collagen-coated beads. Preincubation with CsA or thapsigargin caused a >3-fold decrease in intracellular calcium elevations in response to stimulation with collagen beads. Direct measurements of Ca(2+) in mitochondrial and ER stores showed that CsA only slightly inhibited collagen bead-induced discharge of calcium from mitochondria, but almost completely blocked discharge from ER stores. We reduced the numbers of mitochondria with chronic ethidium bromide treatment to test for the importance of ER/mitochondrial interactions. In these cells, CsA delayed collagen bead induced calcium discharge from mitochondria. Collectively, these data indicate that CsA inhibits collagen phagocytosis by blocking calcium release from ER stores and may perturb functional interactions between the ER and mitochondria that regulate calcium stores. PMID- 11278660 TI - Transcription factor AP-2 functions as a repressor that contributes to the liver specific expression of serum amyloid A1 gene. AB - We previously identified transcription factor AP-2 as the nuclear factor that interacts with the tissue-specific repressor element in the rat serum amyloid A1 (SAA1) promoter. In this report, we provide evidence for a second AP-2-binding site and show that both AP-2 sites participate in mediating the transcription repression of SAA1 promoter. This proximal AP-2 site overlaps with the NFkappaB binding site known to be essential for SAA1 promoter activity. Protein binding competition experiments demonstrated that AP-2 and NFkappaB binding to these overlapping sites were mutually exclusive. Furthermore, the addition of AP-2 easily displaced prebound NFkappaB, whereas NFkappaB could not displace AP-2. These results thus suggest that one mechanism by which AP-2 negatively regulates SAA1 promoter activity may be by antagonizing the function of NFkappaB. Consistent with a repression function, transient expression of AP-2 in HepG2 cells inhibited conditioned medium-induced SAA1 promoter activation. This inhibition was dependent on functional AP-2-binding sites, since mutation of AP-2 binding sites abolished inhibitory effects of AP-2 in HepG2 cells as well as resulted in derepression of the SAA1 promoter in HeLa cells. In addition to SAA1, we found that several other liver gene promoters also contain putative AP-2 binding sites. Some of these sequences could specifically inhibit AP-2.DNA complex formation, and for the human complement C3 promoter, overexpression of AP 2 also could repress its cytokine-mediated activation. Finally, stable expression of AP-2 in hepatoma cells significantly reduced the expression of endogenous SAA, albumin, and alpha-fetoprotein genes. Taken together, our results suggest that AP 2 may function as a transcription repressor to inhibit the expression of not only SAA1 gene but also other liver genes in nonhepatic cells. PMID- 11278661 TI - Androgen receptor specifically interacts with a novel p21-activated kinase, PAK6. AB - The androgen receptor (AR) is a hormone-dependent transcription factor that plays important roles in male sexual differentiation and development. Transcription activation by steroid hormone receptors, such as the androgen receptor, is mediated through interaction with cofactors. We recently identified a novel AR interacting protein, provisionally termed PAK6, that shares a high degree of sequence similarity with p21-activated kinases (PAKs). PAK6 is a 75-kDa protein that contains a putative amino-terminal Cdc42/Rac interactive binding motif and a carboxyl-terminal kinase domain. A domain-specific and ligand-dependent interaction between AR and PAK6 was further confirmed in vivo and in vitro. Northern blot analysis revealed that PAK6 is highly expressed in testis and prostate tissues. Most importantly, immunofluorescence studies showed that PAK6 cotranslocates into the nucleus with AR in response to androgen. Transient transfection experiments showed that PAK6 specifically repressed AR-mediated transcription. This report identifies a novel function for a PAK-homologous protein and suggests a potential unique mechanism by which other signal transduction pathways may cross-talk with AR pathways to regulate AR function in normal and malignant prostate cells. PMID- 11278662 TI - Stopped-flow kinetic analysis of replication protein A-binding DNA: damage recognition and affinity for single-stranded DNA reveal differential contributions of k(on) and k(off) rate constants. AB - Replication protein A (RPA) is a heterotrimeric protein required for many DNA metabolic functions, including replication, recombination, and nucleotide excision repair (NER). We report the pre-steady-state kinetic analysis of RPA binding DNA substrates using a stopped-flow assay to elucidate the kinetics of DNA damage recognition. The bimolecular association rate, k(on), for RPA binding to duplex DNA substrates is greatest for a 1,3d(GXG), intermediate for a 1,2d(GpG) cisplatin-DNA adduct, and least for an undamaged duplex DNA substrate. RPA displays a decreased k(on) and an increased k(off) for a single-stranded DNA substrate containing a single 1,2d(GpG) cisplatin-DNA adduct compared with an undamaged DNA substrate. The k(on) for RPA-binding single-stranded polypyrimidine sequences appears to be diffusion-limited. There is minimal difference in k(on) for varying length DNA substrates; therefore, the difference in equilibrium binding affinity is mainly attributed to the k(off). The k(on) for a purine-rich 30-base DNA is reduced by a factor of 10 compared with a pyrimidine-rich DNA of identical length. These results provide insight into the mechanism of RPA-DNA binding and are consistent with RPA recognition of DNA-damage playing a critical role in NER. PMID- 11278663 TI - Nuclear factor 1 interferes with Sp1 binding through a composite element on the rat poly(ADP-ribose) polymerase promoter to modulate its activity in vitro. AB - Poly(ADP-ribose) polymerase-1 (PARP-1) catalyzes the rapid and extensive poly(ADP ribosyl)ation of nuclear proteins in response to DNA strand breaks, and its expression, although ubiquitous, is modulated from tissue to tissue and during cellular differentiation. PARP-1 gene promoters from human, rat, and mouse have been cloned, and they share a structure common to housekeeping genes, as they lack a functional TATA box and contain multiple GC boxes, which bind the transcriptional activator Sp1. We have previously shown that, although Sp1 is important for rat PARP1 (rPARP) promoter activity, its finely tuned modulation is likely dependent on other transcription factors that bind the rPARP proximal promoter in vitro. In this study, we identified one such factor as NF1-L, a rat liver isoform of the nuclear factor 1 family of transcription factors. The NF1-L site on the rPARP promoter overlaps one of the Sp1 binding sites previously identified, and we demonstrated that binding of both factors to this composite element is mutually exclusive. Furthermore, we provide evidence that NF1-L has no effect by itself on rPARP promoter activity, but rather down-regulates the Sp1 activity by interfering with its ability to bind the rPARP promoter in order to modulate transcription of the rPARP gene. PMID- 11278664 TI - Differential biosynthesis of polysialic or disialic acid Structure by ST8Sia II and ST8Sia IV. AB - ST8Sia II (STX) and ST8Sia IV (PST) are polysialic acid (polySia) synthases that catalyze polySia formation of neural cell adhesion molecule (NCAM) in vivo and in vitro. It still remains unclear how these structurally similar enzymes act differently in vivo. In the present study, we performed the enzymatic characterization of ST8Sia II and IV; both ST8Sia II and IV have pH optima of 5.8 6.1 and have no requirement of metal ions. Because the pH dependence of ST8Sia II and IV enzyme activities and the pK profile of His residues are similar, we hypothesized that a histidine residue would be involved in their catalytic activity. There is a conserved His residue (cf. His(348) in ST8Sia II and His(331) in ST8Sia IV, respectively) within the sialyl motif VS in all sialyltransferase genes cloned to date. Mutant ST8Sia II and IV enzymes in which this His residue was changed to Lys showed no detectable enzyme activity, even though they were folded correctly and could bind to CDP-hexanolamine, suggesting the importance of the His residue for their catalytic activity. Next, the degrees of polymerization of polySia in NCAM catalyzed by ST8Sia II and IV were compared. ST8Sia IV catalyzed larger polySia formation of NCAM than ST8Sia II. We also analyzed the (auto)polysialylated enzymes themselves. Interestingly, when ST8Sia II or IV itself was sialylated under conditions for polysialylation, the disialylated compound was the major product, even though polysialylated compounds were also observed. These results suggested that both ST8Sia II and IV catalyze polySia synthesis toward preferred acceptor substrates such as NCAM, whereas they mainly catalyze disialylation, similarly to ST8Sia III, toward unfavorable substrates such as enzyme themselves. PMID- 11278665 TI - MAPK/ERK overrides the apoptotic signaling from Fas, TNF, and TRAIL receptors. AB - The tumor necrosis factor (TNF), Fas, and TNF-related apoptosis-inducing ligand (TRAIL) receptors (R) are highly specific physiological mediators of apoptotic signaling. We observed earlier that a number of FasR-insensitive cell lines could redirect the proapoptotic signal to an anti-apoptotic ERK1/2 signal resulting in inhibition of caspase activation. Here we determine that similar mechanisms are operational in regulating the apoptotic signaling of other death receptors. Activation of the FasR, TNF-R1, and TRAIL-R, respectively, rapidly induced subsequent ERK1/2 activation, an event independent from caspase activity. Whereas inhibition of the death receptor-mediated ERK1/2 activation was sufficient to sensitize the cells to apoptotic signaling from FasR and TRAIL-R, cells were still protected from apoptotic TNF-R1 signaling. The latter seemed to be due to the strong activation of the anti-apoptotic factor NF-kappaB, which remained inactive in FasR or TRAIL-R signaling. However, when the cells were sensitized with cycloheximide, which is sufficient to sensitize the cells also to apoptosis by TNF-R1 stimulation, we noticed that adenovirus-mediated expression of constitutively active MKK1 could rescue the cells from apoptosis induced by the respective receptors by preventing caspase-8 activation. Taken together, our results show that ERK1/2 has a dominant protecting effect over apoptotic signaling from the death receptors. This protection, which is independent of newly synthesized proteins, acts in all cases by suppressing activation of the caspase effector machinery. PMID- 11278666 TI - The cell cycle control element of histone H4 gene transcription is maximally responsive to interferon regulatory factor pairs IRF-1/IRF-3 and IRF-1/IRF-7. AB - Interferon regulatory factors (IRFs) are transcriptional mediators of interferon responsive signaling pathways that are involved in antiviral defense, immune response, and cell growth regulation. To investigate the role of IRF proteins in the regulation of histone H4 gene transcription, we compared the transcriptional contributions of IRF-1, IRF-2, IRF-3, and IRF-7 using transient transfection assays with H4 promoter/luciferase (Luc) reporter genes. These IRF proteins up regulate reporter gene expression but IRF-1, IRF-3, and IRF-7 are more potent activators of the H4 promoter than IRF-2. Forced expression of different IRF combinations reveals that IRF-2 reduces IRF-1 or IRF-3 dependent activation, but does not affect IRF-7 function. Thus, IRF-2 may have a dual function in histone H4 gene transcription by acting as a weak activator at low dosage and a competitive inhibitor of other strongly activating IRFs at high levels. IRF-1/IRF 3 and IRF-1/IRF-7 pairs each mediate the highest levels of site II-dependent promoter activity and can up-regulate transcription by 120-150-fold. We also find that interferon gamma up-regulates IRF-1 and site II-dependent promoter activity. This up-regulation is not observed when the IRF site is mutated or if cells are preloaded with IRF-1. Our results indicate that IRF-1, IRF-2, IRF-3, and IRF-7 can all regulate histone H4 gene expression. The pairwise utilization of distinct IRF factors provides a flexible transcriptional mechanism for integration of diverse growth-related signaling pathways. PMID- 11278667 TI - Tissue factor pathway inhibitor inhibits endothelial cell proliferation via association with the very low density lipoprotein receptor. AB - Tissue factor pathway inhibitor (TFPI) contains three Kunitz-type proteinase inhibitor domains and is a potent inhibitor of tissue factor-mediated coagulation. Here, we report that TFPI inhibits the proliferation of basic fibroblast growth factor-stimulated endothelial cells. A truncated form of TFPI, containing only the first two Kunitz-type proteinase inhibitor domains, has very little antiproliferative activity, suggesting that the carboxyl-terminal region of TFPI is responsible for this activity. Binding studies revealed that full length TFPI, but not the truncated TFPI molecule, is recognized by the very low density lipoprotein receptor (VLDL receptor) indicating that this receptor is a novel high affinity endothelial cell receptor for TFPI. The antiproliferative activity of TFPI on endothelial cells is inhibited by the receptor-associated protein, a known antagonist of ligand binding by the VLDL receptor, and by anti VLDL receptor antibodies. These results confirm that the antiproliferative activity of TFPI is mediated by the VLDL receptor and suggest that this receptor ligand system may be a useful target for the development of new anti-angiogenic and antitumor agents. PMID- 11278668 TI - Heparin-binding histidine and lysine residues of rat selenoprotein P. AB - Selenoprotein P is a plasma protein that has oxidant defense properties. It binds to heparin at pH 7.0, but most of it becomes unbound as the pH is raised to 8.5. This unusual heparin binding behavior was investigated by chemical modification of the basic amino acids of the protein. Diethylpyrocarbonate (DEPC) treatment of the protein abolished its binding to heparin. DEPC and [(14)C]DEPC modification, coupled with amino acid sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry of peptides, identified several peptides in which histidine and lysine residues had been modified by DEPC. Two peptides from one region (residues 80-95) were identified by both methods. Moreover, the two peptides that constituted this sequence bound to heparin. Finally, when DEPC modification of the protein was carried out in the presence of heparin, these two peptides did not become modified by DEPC. Based on these results, the heparin-binding region of the protein sequence was identified as KHAHLKKQVSDHIAVY. Two other peptides (residues 178-189 and 194-234) that contain histidine-rich sequences met some but not all of the criteria of heparin-binding sites, and it is possible that they and the histidine-rich sequence between them bind to heparin under some conditions. The present results indicate that histidine is a constituent of the heparin-binding site of selenoprotein P. The presence of histidine, the pK(a) of which is 7.0, explains the release of selenoprotein P from heparin binding as pH rises above 7.0. It can be speculated that this property would lead to increased binding of selenoprotein P in tissue regions that have low pH. PMID- 11278669 TI - Clostridium perfringens epsilon toxin induces a rapid change of cell membrane permeability to ions and forms channels in artificial lipid bilayers. AB - Epsilon toxin is a potent toxin produced by Clostridium perfringens types B and D, which are responsible for a rapidly fatal enterotoxemia in animals. One of the main properties of epsilon toxin is the production of edema. We have previously found that epsilon toxin causes a rapid swelling of Madin-Darby canine kidney cells and that the toxin does not enter the cytosol and remains associated with the cell membrane by forming a large complex (Petit, L., Gibert, M., Gillet, D., Laurent-Winter, C., Boquet, P., and Popoff, M. R. (1997) J. Bacteriol. 179, 6480 6487). Here, we report that epsilon toxin induced in Madin-Darby canine kidney cells a rapid decrease of intracellular K(+), and an increase of Cl(-) and Na(+), whereas the increase of Ca(2+) occurred later. The entry of propidium iodide that was correlated with the loss of cell viability monitored by the 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test indicates that epsilon toxin formed large pores. In artificial lipid bilayers, epsilon toxin caused current steps with a single-channel conductance of 60 pS in 100 mm KCl, which represented general diffusion pores. The channels were slightly selective for anions, but cations could also penetrate. Epsilon toxin formed wide and water filled channels permeable to hydrophilic solutes up to a molecular mass of at least 1 kDa, which probably represents the basic mechanism of toxin action on target cells. PMID- 11278670 TI - The crystal structure of a novel mammalian lectin, Ym1, suggests a saccharide binding site. AB - Ym1, a secretory protein synthesized by activated murine peritoneal macrophages, is a novel mammalian lectin with a binding specificity to GlcN. Lectins are responsible for carbohydrate recognition and for mediating cell-cell and cell extracellular matrix interactions in microbes, plants, and animals. Glycosaminoglycan heparin/heparan sulfate binding ability was also detected in Ym1. We report here the three-dimensional structure of Ym1 at 2.5-A resolution by x-ray crystallography. The crystal structure of Ym1 consists of two globular domains, a beta/alpha triose-phosphate isomerase barrel domain and a small alpha + beta folding domain. A notable electron density of sugar is detected in the Ym1 crystal structure. The saccharide is located inside the triose-phosphate isomerase domain at the COOH terminal end of the beta-strands. Both hydrophilic and hydrophobic interactions are noted in the sugar-binding site in Ym1. Despite the fact that Ym1 is not a chitinase, structurally, Ym1 shares significant homology with chitinase A of Serratia marcescens. Ym1 and chitinase A have a similar carbohydrate binding cleft. This study provides new structure information, which will lead to better understanding of the biological significance of Ym1 and its putative gene members. PMID- 11278671 TI - Neuron-specific Bcl-2 homology 3 domain-only splice variant of Bak is anti apoptotic in neurons, but pro-apoptotic in non-neuronal cells. AB - We have identified and characterized N-Bak, a neuron-specific isoform of the pro apoptotic Bcl-2 family member Bak. N-Bak is generated by neuron-specific splicing of a novel 20-base pair exon, which changes the previously described Bak, containing Bcl-2 homology (BH) domains BH1, BH2, and BH3, into a shorter BH3-only protein. As demonstrated by reverse transcription-polymerase chain reaction and RNase protection assay, N-Bak transcripts are expressed only in central and peripheral neurons, but not in other cells, whereas the previously described Bak is expressed ubiquitously, but not in neurons. Neonatal sympathetic neurons microinjected with N-Bak resisted apoptotic death caused by nerve growth factor (NGF) removal, whereas microinjected Bak accelerated NGF deprivation-induced death. Overexpressed Bak killed sympathetic neurons in the presence of NGF, whereas N-Bak did not. N-Bak was, however, still death-promoting when overexpressed in non-neuronal cells. Thus, N-Bak is an anti-apoptotic BH3-only protein, but only in the appropriate cellular environment. This is the first example of a neuron-specific Bcl-2 family member. PMID- 11278672 TI - Delineation of functional regions within the subunits of the Saccharomyces cerevisiae cell adhesion molecule a-agglutinin. AB - a-Agglutinin from Saccharomyces cerevisiae is a cell adhesion glycoprotein expressed on the surface of cells of a mating type and consists of an anchorage subunit Aga1p and a receptor binding subunit Aga2p. Cell wall attachment of Aga2p is mediated through two disulfide bonds to Aga1p (Cappellaro, C., Baldermann, C., Rachel, R., and Tanner, W. (1994) EMBO J. 13, 4737-4744). We report here that purified Aga2p was unstable and had low molar specific activity relative to its receptor alpha-agglutinin. Aga2p co-expressed with a 149-residue fragment of Aga1p formed a disulfide-linked complex with specific activity 43-fold higher than Aga2p expressed alone. Circular dichroism of the complex revealed a mixed alpha/beta structure, whereas Aga2p alone had no periodic secondary structure. A 30-residue Cys-rich Aga1p fragment was partially active in stabilization of Aga2p activity. Mutation of either or both Aga2p cysteine residues eliminated stabilization of Aga2p. Thus the roles of Aga1p include both cell wall anchorage and cysteine-dependent conformational restriction of the binding subunit Aga2p. Mutagenesis of AGA2 identified only C-terminal residues of Aga2p as being essential for binding activity. Aga2p residues 45-72 are similar to sequences in soybean Nod genes, and include residues implicated in interactions with both Aga1p (including Cys(68)) and alpha-agglutinin. PMID- 11278674 TI - Quantifying the impact of membrane microtopology on effective two-dimensional affinity. AB - Just as interactions of soluble proteins are affected by the solvent, membrane protein binding is influenced by the surface environment. This is particularly true for adhesion receptors because their function requires tightly apposed membranes. We sought to demonstrate, and further, to quantify the possible scale of this phenomenon by comparing the effective affinity and kinetic rates of an adhesion receptor (CD16b) placed in three distinct environments: red blood cells (RBCs), detached Chinese hamster ovary (CHO) cells, and K562 cells. Effective affinity reflects both the intrinsic receptor-ligand kinetics and the effectiveness of their presentation by the host membranes. Expression of CD16b, a low affinity Fcgamma receptor, was established by either transfection or spontaneous insertion via its glycosylphosphatidylinositol anchor. Binding to IgG coated RBCs, measured using a micropipette method, indicated a 50-fold increase in effective affinity for receptors on RBCs over CHO and K562 cells, whereas the off rates were similar for all three. Electron microscopy confirmed that specific tight contacts were broad in RBC-RBC conjugates but sparse in CHO-RBC conjugates. We suggest that through modulation of surface roughness the cytoskeleton can greatly impact the effectiveness of adhesion molecules, even those with no cytoplasmic structures. Implications for locomotion and static adhesion are discussed. PMID- 11278673 TI - Studies of insulin secretory responses and of arachidonic acid incorporation into phospholipids of stably transfected insulinoma cells that overexpress group VIA phospholipase A2 (iPLA2beta ) indicate a signaling rather than a housekeeping role for iPLA2beta. AB - A cytosolic 84-kDa group VIA phospholipase A(2) (iPLA(2)beta) that does not require Ca(2+) for catalysis has been cloned from several sources, including rat and human pancreatic islet beta-cells and murine P388D1 cells. Many potential iPLA(2)beta functions have been proposed, including a signaling role in beta-cell insulin secretion and a role in generating lysophosphatidylcholine acceptors for arachidonic acid incorporation into P388D1 cell phosphatidylcholine (PC). Proposals for iPLA(2)beta function rest in part on effects of inhibiting iPLA(2)beta activity with a bromoenol lactone (BEL) suicide substrate, but BEL also inhibits phosphatidate phosphohydrolase-1 and a group VIB phospholipase A(2). Manipulation of iPLA(2)beta expression by molecular biologic means is an alternative approach to study iPLA(2)beta functions, and we have used a retroviral construct containing iPLA(2)beta cDNA to prepare two INS-1 insulinoma cell clonal lines that stably overexpress iPLA(2)beta. Compared with parental INS 1 cells or cells transfected with empty vector, both iPLA(2)beta-overexpressing lines exhibit amplified insulin secretory responses to glucose and cAMP-elevating agents, and BEL substantially attenuates stimulated secretion. Electrospray ionization mass spectrometric analyses of arachidonic acid incorporation into INS 1 cell PC indicate that neither overexpression nor inhibition of iPLA(2)beta affects the rate or extent of this process in INS-1 cells. Immunocytofluorescence studies with antibodies directed against iPLA(2)beta indicate that cAMP-elevating agents increase perinuclear fluorescence in INS-1 cells, suggesting that iPLA(2)beta associates with nuclei. These studies are more consistent with a signaling than with a housekeeping role for iPLA(2)beta in insulin-secreting beta cells. PMID- 11278675 TI - Energetics and specificity of Rat DNA polymerase beta interactions with template primer and gapped DNA substrates. AB - Interactions between rat polymerase beta (pol beta) and the template-primer, as well as gapped DNAs, were studied using the quantitative fluorescence titration technique. Stoichiometries of rat pol beta complexes with DNA substrates are much higher than stoichiometries predicted by the structures of co-crystals. The data can be understood in the context of the two single-stranded (ss)DNA-binding modes of the enzyme, the (pol beta)(16) and (pol beta)(5) binding modes, which differ by the number of nucleotides occluded by the protein. The 8-kDa domain of the enzyme engages the double-stranded (ds)DNA downstream from the primer, while the 31-kDa domain has similar affinity for the ss-ds DNA junction and the dsDNA. The affinity of rat pol beta for the gapped DNA is not affected by the size of the gap. The results indicate a plausible model for recognition of the gapped DNA by rat pol beta. The enzyme binds the ss-ds DNA junction of the gap using the 31-kDa domain. This binding induces an allosteric transition, resulting in the association of the 8-kDa domain with the dsDNA, leading to an amplification of the affinity for the gap. The 5' terminal phosphate, downstream from the primer, has little effect on the affinity, but affects the ssDNA conformation of the gap. PMID- 11278676 TI - The allosteric effector l-lactate induces a conformational change of 2x6-meric lobster hemocyanin in the oxy state as revealed by small angle x-ray scattering. AB - Hemocyanins are multisubunit respiratory proteins found in many invertebrates. They bind oxygen highly cooperatively. However, not much is known about the structural basis of this behavior. We studied the influence of the physiological allosteric effector l-lactate on the oxygenated quaternary structure of the 2x6 meric hemocyanin from the lobster Homarus americanus employing small angle x-ray scattering (SAXS). The presence of 20 mm l-lactate resulted in different scattering curves compared with those obtained in the absence of l-lactate. The distance distribution functions p(r) indicated a more compact molecule in presence of l-lactate, which is also reflected in a reduction of the radius of gyration by about 0.2 nm (3%). Thus, we show for the first time on a structural basis that a hemocyanin in the oxy state can adopt two different conformations. This is as predicted from the analysis of oxygen binding curves according to the "nesting" model. A comparison of the distance distribution functions p(r) obtained from SAXS with those deduced from electron microscopy revealed large differences. The distance between the two hexamers as deduced from electron microscopy has to be shortened by up to 1.1 nm to agree well with the small angle x-ray curves. PMID- 11278677 TI - SDS-induced phenoloxidase activity of hemocyanins from Limulus polyphemus, Eurypelma californicum, and Cancer magister. AB - Phenoloxidase, widely distributed among animals, plants, and fungi, is involved in many biologically essential functions including sclerotization and host defense. In chelicerates, the oxygen carrier hemocyanin seems to function as the phenoloxidase. Here, we show that hemocyanins from two ancient chelicerates, the horseshoe crab Limulus polyphemus and the tarantula Eurypelma californicum, exhibit O-diphenoloxidase activity induced by submicellar concentrations of SDS, a reagent frequently used to identify phenoloxidase activity. The enzymatic activity seems to be restricted to only a few of the heterogeneous subunits. These active subunit types share similar topological positions in the quaternary structures as linkers of the two tightly connected 2 x 6-mers. Because no other phenoloxidase activity was found in the hemolymph of these animals, their hemocyanins may act as a phenoloxidase and thus be involved in the primary immune response and sclerotization of the cuticle. In contrast, hemolymph of a more recent arthropod, the crab Cancer magister, contains both hemocyanin with weak phenoloxidase activity and another hemolymph protein with relatively strong phenoloxidase activity. The chelicerate hemocyanin subunits showing phenoloxidase activity may have evolved into a separate phenoloxidase in crustaceans. PMID- 11278679 TI - Beta -amyloid-(1-42) impairs activity-dependent cAMP-response element-binding protein signaling in neurons at concentrations in which cell survival Is not compromised. AB - Cognitive impairment is a major feature of Alzheimer's disease and is accompanied by beta-amyloid (Abeta) deposition. Transgenic animal models that overexpress Abeta exhibit learning and memory impairments, but neuronal degeneration is not a consistent characteristic. We report that levels of Abeta-(1-42), which do not compromise the survival of cortical neurons, may indeed interfere with functions critical for neuronal plasticity. Pretreatment with Abeta-(1-42), at sublethal concentrations, resulted in a suppression of cAMP-response element-binding protein (CREB) phosphorylation, induced by exposure to either 30 mm KCl or 10 microm N-methyl-d-aspartate. The effects of Abeta-(1-42) seem to involve mechanisms unrelated to degenerative changes, since Abeta-(25-35), a toxic fragment of Abeta, at sublethal concentrations did not interfere with activity dependent CREB phosphorylation. Furthermore, caspase inhibitors failed to counteract the Abeta-(1-42)-evoked suppression of CREB activation. Abeta-(1-42) also interfered with events downstream of activated CREB. The Abeta-(1-42) treatment suppressed the activation of the cAMP response element-containing brain derived neurotrophic factor (BDNF) exon III promoter and the expression of BDNF exon IIII mRNA induced by neuronal depolarization. In view of the critical role of CREB and BDNF in neuronal plasticity, including learning and memory, the observations indicate a novel pathway through which Abeta may interfere with neuronal functions and contribute to cognitive deficit in Alzheimer's disease before the stage of massive neuronal degeneration. PMID- 11278678 TI - Biochemical and biological characterization of a human Rac2 GTPase mutant associated with phagocytic immunodeficiency. AB - The Rho GTPase, Rac2, is expressed only in hematopoietic cell lineages, suggesting a specific cellular function in these cells. Genetic targeting studies in mice showed that Rac2 is an essential regulator of neutrophil chemotaxis, L selectin capture and rolling, and superoxide production. Recently, a dominant negative mutation of Rac2, D57N, has been reported to be associated with a human phagocytic immunodeficiency. To understand further the cellular phenotypes associated with this D57N Rac2 mutant we examined its biochemical characteristics and functional effects when expressed in primary murine bone marrow cells. When compared with wild type (WT) Rac2, D57N Rac2 displayed approximately 10% GTP binding ability resulting from a markedly enhanced rate of GTP dissociation and did not respond to the guanine nucleotide exchange factors. These results suggest that D57N Rac2 may act in a dominant negative fashion in cells by sequestering endogenous guanine nucleotide exchange factors. When expressed in hematopoietic cells, D57N Rac2 reduced endogenous activities of not only Rac2, but also Rac1 and decreased cell expansion in vitro in the presence of growth factors due to increased cell apoptosis. Unexpectedly, D57N expression had no effect on proliferation. In contrast, expansion of cells transduced with WT Rac2 and a dominant active mutant, Q61L, was associated with significantly increased proliferation. Transplantation of transduced bone marrow cells into lethally irradiated recipients showed that the percentage of D57N-containing peripheral blood cells decreased markedly from 40% at 1 month to <5% by 3 months postinjection. Neutrophils derived in vitro from the transduced progenitor cells containing D57N demonstrated markedly impaired migration and O(2)(-) responses to formyl-methionyl-leucyl-phenylalanine, reflecting the same cellular phenotype in these differentiated cells as those described previously in patient cells. These data suggest that the phenotypic abnormalities associated with D57N Rac2 may involve not only neutrophil cellular functions, but also abnormal cell survival in other hematopoietic cells and that overexpression of Rac leads to increased proliferation of normal cells in vitro, whereas deficiency of Rac leads to increased apoptosis. PMID- 11278680 TI - Purification and characterization of an immunomodulatory endometrial protein, glycodelin. AB - Human glycodelin is synthesized by endometrial cells in the late secretory phase and early pregnancy under hormonal regulation. Whereas the precise physiological functions of glycodelin are unknown, its expression during embryonic nidation and its inhibition of T cell proliferation suggest an immunomodulatory role. We purified human glycodelin from first trimester human decidual cytosol by using a rapid two-step high-performance liquid chromatography method and investigated its effects on human monocyte migration. Human U937 cells were used as a model of monocyte chemotaxis in Boyden chamber migration assays. N-Formyl-Met-Leu-Phe and the beta-chemokine RANTES (regulated on activation normal T cell expressed and secreted) were used as monocyte chemoattractants. Purified glycodelin inhibited monocyte migration in a dose-dependent fashion (IC50 = 550 nm). Glycodelin activity was totally reversed by heat inactivation (95 degrees C x 15 min) and neutralized by pretreatment with specific anti-glycodelin antibodies. Deglycosylated glycodelin was equipotent to intact glycodelin in the monocyte migration assay. 125I-Glycodelin binding to whole U937 cells revealed a single, saturable site with a Kd = 48 +/- 21 nm by Scatchard analysis. Cross-linking studies indicated that glycodelin binds to a high molecular mass (approximately 250 kDa) protein complex at the monocyte cell surface. Our findings support the hypothesis that glycodelin reduces the local maternal inflammatory response toward the implantation of a semiallogeneic conceptus. PMID- 11278683 TI - Interactions of the cold shock protein CspB from Bacillus subtilis with single stranded DNA. Importance of the T base content and position within the template. AB - The cold shock protein CspB from Bacillus subtilis binds T-based single-stranded DNA (ssDNA) with high affinity (Lopez, M. M., Yutani, K., and Makhatadze, G. I. (1999) J. Biol. Chem. 274, 33601-33608). In this paper we report the results of CspB interactions with non-homogeneous ssDNA templates containing continuous and non-continuous stretches of T bases. The analysis of CspB-ssDNA interactions was performed using fluorescence spectroscopy, analytical centrifugation and isothermal titration calorimetry. We show that (i) there is a strong correlation between the CspB affinity and stoichiometry and the T content in the oligonucleotide that is independent of which other bases are incorporated into the sequence of ssDNA; (ii) the binding properties of CspB to ssDNA templates with continuous or non-continuous stretches of T bases with similar T content is very similar, and (iii) the mechanism of interaction between CspB and the T-based non-homogeneous ssDNA is mainly through the bases (a stretch of three T bases located in the middle of the ssDNA templates makes the binding independent of the ionic strength). The biological relevance of these results to the role of CspB as an RNA chaperone is discussed. PMID- 11278681 TI - Yeast Mps1p phosphorylates the spindle pole component Spc110p in the N-terminal domain. AB - The yeast spindle pole body (SPB) component Spc110p (Nuf1p) undergoes specific serine/threonine phosphorylation as the mitotic spindle apparatus forms, and this phosphorylation persists until cells enter anaphase. We demonstrate that the dual specificity kinase Mps1p is essential for the mitosis-specific phosphorylation of Spc110p in vivo and that Mps1p phosphorylates Spc110p in vitro. Phosphopeptides generated by proteolytic cleavage were identified and sequenced by mass spectrometry. Ser(60), Thr(64), and Thr(68) are the major sites in Spc110p phosphorylated by Mps1p in vitro, and alanine substitution at these sites abolishes the mitosis-specific isoform in vivo. This is the first time that phosphorylation sites of an SPB component have been determined, and these are the first sites of Mps1p phosphorylation identified. Alanine substitution for any one of these phosphorylated residues, in conjunction with an alanine substitution at residue Ser(36), is lethal in combination with alleles of SPC97, which encodes a component of the Tub4p complex. Consistent with a specific dysfunction for the alanine substitution mutations, simultaneous mutation of all four serine/threonine residues to aspartate does not confer any defect. Sites of Mps1p phosphorylation and Ser(36) are located within the N-terminal globular domain of Spc110p, which resides at the inner plaque of the SPB and binds the Tub4p complex. PMID- 11278684 TI - A vertebrate aldo-keto reductase active with retinoids and ethanol. AB - Enzymes of the short chain and medium chain dehydrogenase/reductase families have been demonstrated to participate in the oxidoreduction of ethanol and retinoids. Mammals and amphibians contain, in the upper digestive tract mucosa, alcohol dehydrogenases of the medium chain dehydrogenase/reductase family, active with ethanol and retinol. In the present work, we searched for a similar enzyme in an avian species (Gallus domesticus). We found that chicken does not contain the homologous enzyme from the medium chain dehydrogenase/reductase family but an oxidoreductase from the aldo-keto reductase family, with retinal reductase and alcohol dehydrogenase activities. The amino acid sequence shows 66-69% residue identity with the aldose reductase and aldose reductase-like enzymes. Chicken aldo-keto reductase is a monomer of M(r) 36,000 expressed in eye, tongue, and esophagus. The enzyme can oxidize aliphatic alcohols, such as ethanol, and it is very efficient in all-trans- and 9-cis-retinal reduction (k(cat)/K(m) = 5,300 and 32,000 mm(-1).min(-1), respectively). This finding represents the inclusion of the aldo-keto reductase family, with the (alpha/beta)(8) barrel structure, into the scenario of retinoid metabolism and, therefore, of the regulation of vertebrate development and tissue differentiation. PMID- 11278685 TI - Physical interaction with Yes-associated protein enhances p73 transcriptional activity. AB - Specific protein-protein interactions are involved in a large number of cellular processes and are mainly mediated by structurally and functionally defined domains. Here we report that the nuclear phosphoprotein p73 can engage in a physical association with the Yes-associated protein (YAP). This association occurs under physiological conditions as shown by reciprocal co immunoprecipitation of complexes from lysates of P19 cells. The WW domain of YAP and the PPPPY motif of p73 are directly involved in the association. Furthermore, as required for ligands to group I WW domains, the terminal tyrosine (Y) of the PPPPY motif of p73 was shown to be essential for the association with YAP. Unlike p73alpha, p73beta, and p63alpha, which bind to YAP, the endogenous as well as exogenously expressed wild-type p53 (wt-p53) and the p73gamma isoform do not interact with YAP. Indeed, we documented that YAP interacts only with those members of the p53 family that have a well conserved PPXY motif, a target sequence for WW domains. Overexpression of YAP causes an increase of p73alpha transcriptional activity. Differential interaction of YAP with members of the p53 family may provide a molecular explanation for their functional divergence in signaling. PMID- 11278686 TI - Interaction between GC box binding factors and Smad proteins modulates cell lineage-specific alpha 2(I) collagen gene transcription. AB - Type I collagen is produced predominantly in mesenchymal cells, but molecular mechanisms responsible for cell type-specific expression are virtually unknown. During fibrogenic process in the liver, activated hepatic stellate cells (HSC) are the main producers of type I collagen, whereas parenchymal hepatocytes produce little, if any, of this protein. We have previously reported that Sp1 and an interacting unknown factor(s) bind to the -313 to -255 sequence of the alpha2(I) collagen gene (COL1A2) and play essential roles for basal and TGF-beta stimulated transcription in skin fibroblasts and HSC. Recently, Smad3 has been shown to bind to this region, and its interaction with Sp1 has been implicated in TGF-beta-elicited COL1A2 stimulation. The present study demonstrates predominant binding of Sp3 rather than Sp1 to this regulatory element in parenchymal hepatocytes. In these cells, this region did not exhibit strong enhancer activity or mediate the effect of TGF-beta. Transfection of HSC with an Sp3 expression plasmid abolished the COL1A2 response to TGF-beta, whereas overexpression of Sp1 in hepatocytes increased basal COL1A2 transcription and conferred TGF-beta responsiveness. Functional and physical interactions between Sp1 and Smad3, but not between Sp3 and Smad3, were demonstrated using the bacterial GAL4 system and immunoprecipitation-Western blot analyses. These results indicate that cell lineage-specific interactions between GC box binding factors and Smad protein(s) may account, at least in part, for differential COL1A2 transcription and TGF-beta responsiveness in HSC and parenchymal hepatocytes. PMID- 11278687 TI - Kinetic and mechanistic studies of prolyl oligopeptidase from the hyperthermophile Pyrococcus furiosus. AB - Prolyl oligopeptidase (POP) is widely distributed in mammals, where it is implicated in neuropeptide processing. It is also present in some bacteria and archaea. Because POP is found in mesophilic and hyperthermophilic organisms, and is distributed among all three phylogenetic domains, studies of its function and structure could lead to new insights about the evolution of enzyme mechanisms and thermostability. Kinetic studies were conducted on the POP of the hyperthermophilic archaeon Pyrococcus furiosus (Pfu) 85 degrees C in both H(2)O and D(2)O. Pfu POP displayed many similarities to mammalian POPs, however the solvent isotope effect (k(0)/k(1)) was 2.2 at both high and low pH, indicating that general base/acid catalysis is the rate-limiting step. The pH-rate profiles indicated a three-deprotonation process with pK(a) values of 4.3, 7.2, and 9.1. The temperature dependence of these values revealed a heat of ionization of 4.7 kJ/mol for pK(es1) and 22 kJ/mol for pK(es2), suggesting the catalytic involvement of a carboxyl group and an imidazole group, respectively. Temperature dependence of the catalytic rate was assessed at pH 6.0 and 7.6. Entropy values of -119 and -143 Jmol(-1)K(-1) were calculated at the respective pH values, with a corresponding difference in enthalpy of 8.5 kJ/mol. These values suggest that two or three hydrogen bonds are broken during the transition state of the acidic enzyme form, whereas only one or two are broken during the transition state of the basic enzyme form. A model has been constructed for Pfu POP based on the crystal structure of porcine POP and the sequence alignment. The similarities demonstrated for POPs from these two organisms reflect the most highly conserved characteristics of this class of serine protease, whereas the differences between these enzymes highlights the large evolutionary distance between them. Such fundamental information is crucial to our understanding of the function of proteins at high temperature. PMID- 11278688 TI - Isolation of a zebrafish rod opsin promoter to generate a transgenic zebrafish line expressing enhanced green fluorescent protein in rod photoreceptors. AB - To exploit zebrafish as a transgenic model, tissue-specific promoters must be identified. We isolated a 20-kilobase (kbp) zebrafish rod opsin genomic clone, which consists of 18 kbp of 5'-flanking region, the entire coding region, and 0.5 kbp of 3'-flanking sequence. Polymerase chain reaction, Southern blotting, and DNA sequencing revealed the rod opsin gene lacks introns. The transcription start site was localized 94 nucleotides upstream of the translation initiation site. Sequence alignment with orthologous promoters revealed conserved cis-elements including glass, NRE, OTX/Bat-1, Ret-1/PCE-1, Ret-4, and TATA box. A 1.2-kbp promoter fragment was cloned upstream of the enhanced green fluorescent protein (EGFP) cDNA and microinjected into 1- to 2-cell stage zebrafish embryos. EGFP expression was detected in the ventral-nasal eye at 3 days postfertilization and spread throughout the eye. Progeny of the positive founder fish, which were identified by polymerase chain reaction amplification of fin genomic DNA, exhibited EGFP expression in the retina, confirming the germline transmission of the transgene. Frozen eye sections demonstrated the EGFP expression was rod specific and exhibited a similar developmental expression profile as the rod opsin protein. This stable transgenic line provides a novel tool for identification of genes regulating development and maintenance of rod photoreceptors. PMID- 11278689 TI - NADH oxidase activity of mitochondrial apoptosis-inducing factor. AB - Apoptosis-inducing factor (AIF) is a mitochondrial flavoprotein, which translocates to the nucleus during apoptosis and causes chromatin condensation and large scale DNA fragmentation. Here we report the biochemical characterization of AIF's redox activity. Natural AIF purified from mitochondria and recombinant AIF purified from bacteria (AIFDelta1-120) exhibit NADH oxidase activity, whereas superoxide anion (O(2)(-)) is formed. AIFDelta1-120 is a monomer of 57 kDa containing 1 mol of noncovalently bound FAD/mol of protein. ApoAIFDelta1-120, which lacks FAD, has no NADH oxidase activity. However, native AIFDelta1-120, apoAIFDelta1-120, and the reconstituted (FAD-containing) holoAIFDelta1-120 protein exhibit a similar apoptosis-inducing potential when microinjected into the cytoplasm of intact cells. Inhibition of the redox function, by external addition of superoxide dismutase or covalent derivatization of FAD with diphenyleneiodonium, failed to affect the apoptogenic function of AIFDelta1-120 assessed on purified nuclei in a cell-free system. Conversely, blockade of the apoptogenic function of AIFDelta1-120 with the thiol reagent para chloromercuriphenylsulfonic acid did not affect its NADH oxidase activity. Altogether, these data indicate that AIF has a marked oxidoreductase activity which can be dissociated from its apoptosis-inducing function. PMID- 11278690 TI - Identification of a 350-kDa ClpP protease complex with 10 different Clp isoforms in chloroplasts of Arabidopsis thaliana. AB - A 350-kDa ClpP protease complex with 10 different subunits was identified in chloroplast of Arabidopsis thaliana, using Blue-Native gel electrophoresis, followed by matrix-assisted laser desorption ionization time-of-flight and nano electrospray tandem mass spectrometry. The complex was copurified with the thylakoid membranes, and all identified Clp subunits show chloroplast targeting signals, supporting that this complex is indeed localized in the chloroplast. The complex contains chloroplast-encoded pClpP and six nuclear-encoded proteins nCpP1 6, as well as two unassigned Clp homologues (nClpP7, nClpP8). An additional Clp protein was identified in this complex; it does not belong to any of the known Clp genes families and is here assigned ClpS1. Expression and accumulation of several of these Clp proteins have never been shown earlier. Sequence and phylogenetic tree analysis suggests that nClpP5, nClpP2, and nClpP8 are not catalytically active and form a new group of Clp higher plant proteins, orthologous to the cyanobacterial ClpR protein, and are renamed ClpR1, -2, and 3, respectively. We speculate that ClpR1, -2, and -3 are part of the heptameric rings, whereas ClpS1 is a regulatory subunit positioned at the axial opening of the ClpP/R core. Several truncations and errors in intron and exon prediction of the annotated Clp genes were corrected using mass spectrometry data and by matching genomic sequences with cDNA sequences. This strategy will be widely applicable for the much needed verification of protein prediction from genomic sequence. The extreme complexity of the chloroplast Clp complex is discussed. PMID- 11278691 TI - Regulation of Id2 gene expression by the insulin-like growth factor I receptor requires signaling by phosphatidylinositol 3-kinase. AB - The Id proteins play an important role in proliferation, differentiation, and tumor development. We report here that Id gene expression can be regulated by the insulin-like growth factor I receptor (IGF-IR), a receptor that also participates in the regulation of cellular proliferation and differentiation. Specifically, we found that the IGF-IR activated by its ligand was a strong inducer of Id2 gene expression in 32D murine hemopoietic cells. This activation was not simply the result of cellular proliferation, as Id2 gene expression was higher in 32D cells stimulated by IGF-I than in cells exponentially growing in interleukin-3. The up regulation of Id2 gene expression was largely dependent on the presence of insulin receptor substrate-1, a major substrate of the IGF-IR and a potent activator of the phosphatidylinositol 3-kinase (PI3K) pathway. The role of PI3K activity in the up-regulation of Id2 gene expression by the IGF-IR was confirmed by different methods and in different cell types. In 32D cells, the up-regulation of Id2 gene expression by the PI3K pathway correlated with interleukin-3 independence and inhibition of differentiation. PMID- 11278692 TI - CARD11 and CARD14 are novel caspase recruitment domain (CARD)/membrane-associated guanylate kinase (MAGUK) family members that interact with BCL10 and activate NF kappa B. AB - The caspase recruitment domain (CARD) is a protein-binding module that mediates the assembly of CARD-containing proteins into apoptosis and NF-kappaB signaling complexes. We report here that CARD protein 11 (CARD11) and CARD protein 14 (CARD14) are novel CARD-containing proteins that belong to the membrane associated guanylate kinase (MAGUK) family, a class of proteins that functions as molecular scaffolds for the assembly of multiprotein complexes at specialized regions of the plasma membrane. CARD11 and CARD14 have homologous structures consisting of an N-terminal CARD domain, a central coiled-coil domain, and a C terminal tripartite domain comprised of a PDZ domain, an Src homology 3 domain, and a GUK domain with homology to guanylate kinase. The CARD domains of both CARD11 and CARD14 associate specifically with the CARD domain of BCL10, a signaling protein that activates NF-kappaB through the IkappaB kinase complex in response to upstream stimuli. When expressed in cells, CARD11 and CARD14 activate NF-kappaB and induce the phosphorylation of BCL10. These findings suggest that CARD11 and CARD14 are novel MAGUK family members that function as upstream activators of BCL10 and NF-kappaB signaling. PMID- 11278693 TI - In vivo interaction between dynamitin and MacMARCKS detected by the fluorescent resonance energy transfer method. AB - Dynamitin is a subunit of the dynactin complex regulating microtubule-dependent motor functions, and MacMARCKS (Macrophage-enriched myristoylated alanine-rich protein kinase C substrate) is a major protein kinase C substrate regulating integrin activation. The interaction between dynamitin and MacMARCKS has been implicated in integrin-dependent cell spreading. However, the in vivo interaction of these two proteins in living cells has not been demonstrated. Spatial and temporal information about the interaction is also lacking. In this study, we used the fluorescent resonance energy transfer method to demonstrate in vivo interaction between MacMARCKS and dynamitin with cyan fluorescent protein (CFP) conjugated dynamitin as the donor fluorophore and yellow fluorescent protein (YFP)-conjugated MacMARCKS as the acceptor fluorophore. The interaction of these two fusion proteins was studied both in vitro and in vivo, and typical fluorescent resonance energy transfer was observed; the CFP emission peak increased while the YFP emission peak decreased when protein interaction was abolished. Spatial and temporal information was obtained in RAW macrophage cells. In resting macrophage cells, dynamitin-MacMARCKS interaction is concentrated at the cell periphery, although the majority of dynamitin is distributed at the perinuclear region of the cells. When cells were treated with phorbol 12 myristate 13-acetate, both proteins concentrated to perinuclear regions of the cells, and yet the interaction disappeared as the cell spread. Similar events were also observed in 293 cells. Thus, we conclude that dynamitin and MacMARCKS indeed interact in living cells. PMID- 11278694 TI - Regulation of STRA13 by the von Hippel-Lindau tumor suppressor protein, hypoxia, and the UBC9/ubiquitin proteasome degradation pathway. AB - In this study, we focus on different modes of regulation of STRA13, a human ortholog of the mouse basic helix-loop-helix transcriptional factor, previously identified by us as a new von Hippel-Lindau tumor suppressor gene (VHL) target. The gene was overexpressed in VHL-deficient cell lines and tumors, specifically clear cell renal carcinomas and hemangioblastomas. Introduction of wild type VHL transgene into clear cell renal carcinoma restored low level expression of STRA13. Overexpression was also detected in many common malignancies with an intact VHL gene, suggesting the existence of another, VHL-independent pathway of STRA13 regulation. Similar to many other von Hippel-Lindau tumor-suppressor protein (pVHL) targets, the expression of STRA13 on the mRNA level was hypoxia sensitive, indicating oxygen-dependent regulation of the gene, presumably through the pVHL/hypoxia-inducible factor 1 (HIF-1) pathway. The yeast two-hybrid screening revealed interaction of the STRA13 protein with the human ubiquitin conjugating enzyme (UBC9) protein, the specificity of which was confirmed in mammalian cells. By adding the proteasome inhibitor acetyl-leucinyl-leucinyl norleucinal, we demonstrated that the 26 S proteasome pathway regulates the stability of pSTRA13. Co-expression of STRA13 and UBC9 led to an increase of the pSTRA13 ubiquitination and subsequent degradation. These data established that UBC9/STRA13 association in cells is of physiological importance, presenting direct proof of UBC9 involvement in the ubiquitin-dependent degradation of pSTRA13. Hypoxia treatment of mammalian cells transiently expressing STRA13 protein showed that stability of pSTRA13 is not affected by hypoxia or VHL. Thus, STRA13, a new pVHL target, is regulated in cells on multiple levels. We propose that STRA13 may play a critical role in carcinogenesis, since it is a potent transcriptional regulator, abundant in a variety of common tumors. PMID- 11278696 TI - Aberrant expression of human mucin gene MUC5B in gastric carcinoma and cancer cells. Identification and regulation of a distal promoter. AB - In gastric cancer, altered expression of MUC1, MUC2, MUC5AC, and MUC6 mucin genes has already been described. We show in this report by the means of in situ hybridization, reverse transcriptase-polymerase chain reaction, and transfection assays that MUC5B is also abnormally expressed in gastric carcinomatous tissues and cell lines. We thus undertook to elucidate the molecular mechanisms that regulate the transcription of MUC5B in gastric cancer cells. To this end, high expressing (KATO-III) and low expressing (AGS) gastric cancer cell lines were chosen to study human mucin gene MUC5B expression and promoter activity. Sequencing of the promoter region revealed a distal TATA box located 1 kilobase upstream of the proximal TATA box. Functional activity of the promoter was addressed by using deletion mutants covering 2044 nucleotides upstream of the MUC5B transcription start site. We identified a distal promoter 10 times more active than the proximal promoter in KATO-III cells. In AGS cells, both promoters, much less active, showed the same range of activity. Binding assays allowed us to show that the transcription factor ATF-1 binds to a cis-element present in the distal promoter. Sp1, which binds to both promoters specifically transactivates the proximal promoter. Treatment of transfected cells with PMA, cholera toxin A subunit, and calcium ionophore showed that only PMA led to a substantial activation of the distal promoter. MUC5B 5'-flanking region having a high GC content, influence of methylation on the MUC5B expression was assessed. Our results indicate that repression of MUC5B expression visualized in AGS cells is due in part to the presence of numerous methylated cytosine residues throughout the 5'-flanking region. Altogether these results demonstrate that MUC5B expression in gastric cancer cells is governed by a highly active distal promoter that is up-regulated by protein kinase C and that repression is under the influence of methylation. PMID- 11278695 TI - The human immunodeficiency virus type 1 Vpu protein inhibits NF-kappa B activation by interfering with beta TrCP-mediated degradation of Ikappa B. AB - The human immunodeficiency virus type 1 (HIV-1) Vpu protein binds to the CD4 receptor and induces its degradation by cytosolic proteasomes. This process involves the recruitment of human betaTrCP (TrCP), a key member of the SkpI-Cdc53 F-box E3 ubiquitin ligase complex that specifically interacts with phosphorylated Vpu molecules. Interestingly, Vpu itself, unlike other TrCP-interacting proteins, is not targeted for degradation by proteasomes. We now report that, by virtue of its affinity for TrCP and resistance to degradation, Vpu, but not a phosphorylation mutant unable to interact with TrCP, has a dominant negative effect on TrCP function. As a consequence, expression of Vpu in HIV-infected T cells or in HeLa cells inhibited TNF-alpha-induced degradation of IkappaB-alpha. Vpu did not inhibit TNF-alpha-mediated activation of the IkappaB kinase but instead interfered with the subsequent TrCP-dependent degradation of phosphorylated IkappaB-alpha. This resulted in a pronounced reduction of NF kappaB activity. We also observed that in cells producing Vpu-defective virus, NF kappaB activity was significantly increased even in the absence of cytokine stimulation. However, in the presence of Vpu, this HIV-mediated NF-kappaB activation was markedly reduced. These results suggest that Vpu modulates both virus- and cytokine-induced activation of NF-kappaB in HIV-1-infected cells. PMID- 11278697 TI - Conserved cysteines in the sialyltransferase sialylmotifs form an essential disulfide bond. AB - The sialyltransferase gene family is comprised of 16 cloned enzymes. All members contain two conserved protein domains, termed the S- and L-sialylmotifs, that participate in substrate binding. Of only six invariant amino acids, two are cysteines, with one found in each sialylmotif. Although the recombinant soluble form of ST6Gal I has six cysteines, quantitative analysis indicated the presence of only one disulfide linkage, and thiol reducing agents dithiothreitol and beta mercaptoethanol inactivated the enzyme. Analysis of site-directed mutants showed that alanine or serine mutants of invariant Cys(181) or Cys(332) exhibit no detectable activity, either by direct assay or by staining of the transfected cells with Sambucus nigra agglutinin, which recognizes the product NeuAcalpha2,6Galbeta1,4GlcNAc on glycoproteins. In contrast, alanine mutations of charged residues adjacent to either cysteine showed little or no effect on enzyme activity. Immunofluorescence microscopy showed that although the wild type sialyltransferase is properly localized in the Golgi apparatus, the inactive cysteine mutants are retained in the endoplasmic reticulum. The results suggest that the invariant cysteine residues in the L- and S-sialylmotifs participate in the formation of an intradisulfide linkage that is essential for proper conformation and activity of ST6Gal I. PMID- 11278698 TI - Akt and Bcl-xL promote growth factor-independent survival through distinct effects on mitochondrial physiology. AB - A comparison of Akt- and Bcl-x(L)-dependent cell survival was undertaken using interleukin-3-dependent FL5.12 cells. Expression of constitutively active Akt allows cells to survive for prolonged periods following growth factor withdrawal. This survival correlates with the expression level of activated Akt and is comparable in magnitude to the protection provided by the anti-apoptotic gene Bcl x(L). Although both genes prevent cell death, Akt-protected cells can be distinguished from Bcl-x(L)-protected cells on the basis of increased glucose transporter expression, glycolytic activity, mitochondrial potential, and cell size. In addition, Akt-expressing cells require high levels of extracellular nutrients to support cell survival. In contrast, Bcl-x(L)-expressing cells deprived of interleukin-3 survive in a more vegetative state, in which the cells are smaller, have lower mitochondrial potential, reduced glycolytic activity, and are less dependent on extracellular nutrients. Thus, Akt and Bcl-x(L) suppress mitochondrion-initiated apoptosis by distinct mechanisms. Akt-mediated survival is dependent on promoting glycolysis and maintaining a physiologic mitochondrial potential. In contrast, Bcl-x(L) maintains mitochondrial integrity in the face of a reduced mitochondrial membrane potential, which develops as a result of the low glycolytic rate in growth factor-deprived cells. PMID- 11278699 TI - Mechanical strain induces specific changes in the synthesis and organization of proteoglycans by vascular smooth muscle cells. AB - In the mechanically active environment of the artery, cells sense mechanical stimuli and regulate extracellular matrix structure. In this study, we explored the changes in synthesis of proteoglycans by vascular smooth muscle cells in response to precisely controlled mechanical strains. Strain increased mRNA for versican (3.2-fold), biglycan (2.0-fold), and perlecan (2.0-fold), whereas decorin mRNA levels decreased to a third of control levels. Strain also increased versican, biglycan, and perlecan core proteins, with a concomitant decrease in decorin core protein. Deformation did not alter the hydrodynamic size of proteoglycans as evidenced by molecular sieve chromatography but increased sulfate incorporation in both chondroitin/dermatan sulfate proteoglycans and heparan sulfate proteoglycans (p < 0.05 for both). Using DNA microarrays, we also identified the gene for the hyaluronan-linking protein TSG6 as mechanically induced in smooth muscle cells. Northern analysis confirmed a 4.0-fold increase in steady state mRNA for TSG6 following deformation. Size exclusion chromatography under associative conditions showed that versican-hyaluronan aggregation was enhanced following deformation. These data demonstrate that mechanical deformation increases specific vascular smooth muscle cell proteoglycan synthesis and aggregation, indicating a highly coordinated extracellular matrix response to biomechanical stimulation. PMID- 11278700 TI - Relationships between the activities in vitro and in vivo of various kinds of ribozyme and their intracellular localization in mammalian cells. AB - Nineteen different functional RNAs were synthesized for an investigation of the actions of ribozymes, in vitro and in vivo, under the control of two different promoters, tRNA or U6, which localize transcripts either in the cytoplasm or in the nucleus. No relationships were found between the activities of these RNAs in cultured cells and the kinetic parameters of their respective chemical cleavage reactions in vitro, indicating that in no case was chemical cleavage the rate limiting step in vivo. For example, a hepatitis delta virus (HDV) ribozyme, whose activity in vitro was almost 3 orders of magnitude lower than that of a hammerhead ribozyme, still exhibited similar activity in cells when an appropriate expression system was used. As expected, external guide sequences, the actions of which depend on nuclear RNase P, were more active in the nucleus. Analysis of data obtained with cultured cells clearly demonstrated that the cytoplasmic ribozymes were significantly more active than the nuclear ribozymes, suggesting that mature mRNAs in the cytoplasm might be more accessible to antisense molecules than are pre-mRNAs in the nucleus. Our findings should be useful for the future design of intracellularly active functional molecules. PMID- 11278701 TI - Expression of the Caldariomyces fumago chloroperoxidase in Aspergillus niger and characterization of the recombinant enzyme. AB - The Caldariomyces fumago chloroperoxidase was successfully expressed in Aspergillus niger. The recombinant enzyme was produced in the culture medium as an active protein and could be purified by a three-step purification procedure. The catalytic behavior of recombinant chloroperoxidase (rCPO) was studied and compared with that of native CPO. The specific chlorination activity (47 units/nmol) of rCPO and its pH optimum (pH 2.75) were very similar to those of native CPO. rCPO catalyzes the oxidation of various substrates in comparable yields and selectivities to native CPO. Indole was oxidized to 2-oxindole with 99% selectivity and thioanisole to the corresponding R-sulfoxide (enantiomeric excess >98%). Incorporation of (18)O from labeled H(2)18O(2) into the oxidized products was 100% in both cases. PMID- 11278702 TI - Differential activation of the Rac pathway by Ha-Ras and K-Ras. AB - Ras proteins are key regulators of cell growth and differentiation. Mammalian cells express three closely related Ras proteins: Ha-Ras, K-Ras, and N-Ras. We have compared the abilities of the Ha-Ras and K-Ras isoforms to activate the Rac effector pathway, using three Rac-dependent readouts: induction of membrane ruffling and pinocytosis, stimulation of cell motility, and Pak binding. The total surface area of membrane ruffles induced by K-RasV12 was 2-fold greater than that induced by Ha-RasV12. Likewise, the number of K-RasV12-induced pinocytic vesicles per cell was approximately 2-fold greater than that induced by Ha-RasV12. In a wound healing assay, K-RasV12-injected cells migrated twice as fast as Ha-RasV12-injected cells. Moreover, the Pak binding activity of Rac, which is indicative of the amount of GTP-bound Rac, was higher in K-RasV12 expressing cells than Ha-RasV12-expressing cells. These results suggest that K Ras activates Rac more efficiently than Ha-Ras. The preferential activation of Rac by K-Ras is dependent on the mode of membrane anchoring and impacts on the ability of K-Ras to regulate cell survival. PMID- 11278703 TI - Subunit-subunit interactions in trimeric arginase. Generation of active monomers by mutation of a single amino acid. AB - The structure of the trimeric, manganese metalloenzyme, rat liver arginase, has been previously determined at 2.1-A resolution (Kanyo, Z. F., Scolnick, L. R., Ash, D. E., and Christianson, D. W., (1996) Nature 383, 554-557). A key feature of this structure is a novel S-shaped oligomerization motif at the carboxyl terminus of the protein that mediates approximately 54% of the intermonomer contacts. Arg-308, located within this oligomerization motif, nucleates a series of intramonomer and intermonomer salt links. In contrast to the trimeric wild type enzyme, the R308A, R308E, and R308K variants of arginase exist as monomeric species, as determined by gel filtration and analytical ultracentrifugation, indicating that mutation of Arg-308 shifts the equilibrium for trimer dissociation by at least a factor of 10(5). These monomeric arginase variants are catalytically active, with k(cat)/K(m) values that are 13-17% of the value for wild-type enzyme. The arginase variants are characterized by decreased temperature stability relative to the wild-type enzyme. Differential scanning calorimetry shows that the midpoint temperature for unfolding of the Arg-308 variants is in the range of 63.6-65.5 degrees C, while the corresponding value for the wild-type enzyme is 70 degrees C. The three-dimensional structure of the R308K variant has been determined at 3-A resolution. At the high protein concentrations utilized in the crystallizations, this variant exists as a trimer, but weakened salt link interactions are observed for Lys-308. PMID- 11278704 TI - The Gab1 docking protein links the b cell antigen receptor to the phosphatidylinositol 3-kinase/Akt signaling pathway and to the SHP2 tyrosine phosphatase. AB - B cell antigen receptor (BCR) signaling causes tyrosine phosphorylation of the Gab1 docking protein. This allows phosphatidylinositol 3-kinase (PI3K) and the SHP2 tyrosine phosphatase to bind to Gab1. In this report, we tested the hypothesis that Gab1 acts as an amplifier of PI3K- and SHP2-dependent signaling in B lymphocytes. By overexpressing Gab1 in the WEHI-231 B cell line, we found that Gab1 can potentiate BCR-induced phosphorylation of Akt, a PI3K-dependent response. Gab1 expression also increased BCR-induced tyrosine phosphorylation of SHP2 as well as the binding of Grb2 to SHP2. We show that the pleckstrin homology (PH) domain of Gab1 is required for BCR-induced phosphorylation of Gab1 and for Gab1 participation in BCR signaling. Moreover, using confocal microscopy, we show that BCR ligation can induce the translocation of Gab1 from the cytosol to the plasma membrane and that this requires the Gab1 PH domain as well as PI3K activity. These findings are consistent with a model in which the binding of the Gab1 PH domain to PI3K-derived lipids brings Gab1 to the plasma membrane, where it can be tyrosine-phosphorylated and then act as an amplifier of BCR signaling. PMID- 11278705 TI - Optimized RNA targets of two closely related triple KH domain proteins, heterogeneous nuclear ribonucleoprotein K and alphaCP-2KL, suggest Distinct modes of RNA recognition. AB - The KH domain mediates RNA binding in a wide range of proteins. Here we investigate the RNA-binding properties of two abundant RNA-binding proteins, alphaCP-2KL and heterogeneous nuclear ribonucleoprotein (hnRNP) K. These proteins constitute the major poly(C) binding activity in mammalian cells, are closely related on the basis of the structures and positioning of their respective triplicated KH domains, and have been implicated in a variety of post transcriptional controls. By using SELEX, we have obtained sets of high affinity RNA targets for both proteins. The primary and secondary structures necessary for optimal protein binding were inferred in each case from SELEX RNA sequence comparisons and confirmed by mutagenesis and structural mapping. The target sites for alphaCP-2KL and hnRNP K were both enriched for cytosine bases and were presented in a single-stranded conformation. In contrast to these shared characteristics, the optimal target sequence for hnRNP K is composed of a single short "C-patch" compatible with recognition by a single KH domain whereas that for alphaCP-2KL encompassed three such C-patches suggesting more extensive interactions. The binding specificities of the respective SELEX RNAs were confirmed by testing their interactions with native proteins in cell extracts, and the importance of the secondary structure in establishing an optimized alphaCP-2KL-binding site was supported by comparison of SELEX target structure with that of the native human alpha-globin 3'-untranslated region. These data indicate that modes of macromolecular interactions of arrayed KH domains can differ even among closely related KH proteins and that binding affinities are substantially dependent on the presentation of the target site within the RNA secondary structure. PMID- 11278706 TI - Role of cysteinyl residues in sensing Pb(II), Cd(II), and Zn(II) by the plasmid pI258 CadC repressor. AB - The cadCA operon of Staphylococcus aureus plasmid pI258 confers resistance to salts of the soft metals lead, cadmium, and zinc. The operon is regulated by CadC, a member of the ArsR family of metal-responsive transcriptional repressors. In this study the role of the five cysteine residues of CadC in soft metal ion sensing was investigated. Cys-7, Cys-11, Cys-52, Cys-58, and Cys-60 were changed individually to glycine or serine residues. The effect of the cadC mutations was examined in Escherichia coli using a green fluorescent protein reporter system. None of the mutations affected the ability of CadC to repress gfp expression. Neither Cys-11 nor Cys-52 was required for in vivo response to Pb(II), Zn(II), or Cd(II). Cys-7, Cys-58, or Cys-60 mutations each reduced or eliminated soft metal sensing. Wild-type and mutant CadC proteins were purified, and the effect of the substitutions on DNA binding was determined using a restriction enzyme protection assay. Binding of wild-type CadC protected cad operator DNA from digestion at the single SspI site, and the addition of Pb(II), Zn(II), or Cd(II) resulted in deprotection. Chemical modification of the cysteine residues in CadC had no effect on protection but eliminated deprotection. C11G and C52G proteins exhibited wild-type properties in vitro. C7G, C58S, and C60G proteins were able to be protected from SspI digestion but had reduced responses to soft metal ions. The results indicate that Cys-7, Cys-58, and Cys-60 are involved in sensing those soft metals and suggest that they are ligands to Pb(II), Zn(II), and Cd(II). PMID- 11278707 TI - Calcium- and syntaxin 1-mediated trafficking of the neuronal glycine transporter GLYT2. AB - Previously we demonstrated the existence of a physical and functional interaction between the glycine transporters and the SNARE protein syntaxin 1. In the present report the physiological role of the syntaxin 1-glycine transporter 2 (GLYT2) interaction has been investigated by using a brain-derived preparation. Previous studies, focused on syntaxin 1-transporter interactions using overexpression systems, led to the postulation that syntaxin is somehow implicated in protein trafficking. Since syntaxin 1 is involved in exocytosis of neurotransmitter and also interacts with GLYT2, we stimulated exocytosis in synaptosomes and examined its effect on surface-expression and transport activity of GLYT2. We found that, under conditions that stimulate vesicular glycine release, GLYT2 is rapidly trafficked first toward the plasma membrane and then internalized. When the same experiments were performed with synaptosomes inactivated for syntaxin 1 by a pretreatment with the neurotoxin Bont/C, GLYT2 was unable to reach the plasma membrane but still was able to leave it. These results indicate the existence of a SNARE-mediated regulatory mechanism that controls the surface-expression of GLYT2. Syntaxin 1 is involved in the arrival to the plasma membrane but not in the retrieval. Furthermore, by using immunogold labeling on purified preparations from synaptosomes, we demonstrate that GLYT2 is present in small synaptic-like vesicles. GLYT2-containing vesicles may represent neurotransmitter transporter that is being trafficked. The results of our work suggest a close correlation between exocytosis of neurotransmitter and its reuptake by transporters. PMID- 11278708 TI - The CGM1a (CEACAM3/CD66d)-mediated phagocytic pathway of Neisseria gonorrhoeae expressing opacity proteins is also the pathway to cell death. AB - Phagocytosis of Opa+ Neisseria gonorrhoeae (gonococcus, GC) by neutrophils is in part dependent on the interaction of Opa proteins with CGM1a (CEACAM3/CD66d) antigens, a neutrophil-specific receptor. However, the signaling pathways leading to phagocytosis have not been characterized. Here we show that interaction of OpaI bacteria with neutrophils or CGM1a-transfected DT40 cells induces calcium flux, which correlates with phagocytosis of bacteria. We identified an immunoreceptor tyrosine-based activation motif (ITAM) in CGM1a, and showed that the ability of CGM1a to transduce signals and mediate phagocytosis was abolished by mutation of the ITAM tyrosines. We also demonstrated that CGM1a-ITAM-mediated bacterial phagocytosis is dependent on Syk and phospholipase C activity in DT40 cells. Unexpectedly, the activation of the CGM1a-ITAM phagocytic pathway by Opa+ GC results in induction of cell death. PMID- 11278709 TI - Basic fibroblast growth factor-induced activation of novel CREB kinase during the differentiation of immortalized hippocampal cells. AB - Growth factors bind to their specific receptors on the responsive cell surface and thereby initiate dramatic changes in the proliferation, differentiation, and survival of their target cells. In the present study we have examined the mechanism by which growth factor-induced signals are propagated to the nucleus, leading to the activation of transcription factor, cis-acting cAMP response element (CRE)-binding protein (CREB), in immortalized hippocampal progenitor cells (H19-7). During the differentiation of H19-7 cells by basic fibroblast growth factor (bFGF) a critical regulatory Ser(133) residue of CREB was phosphorylated followed by an increase of CRE-mediated gene transcription. Expression of S133A CREB mutants blocked the differentiation of H19-7 cells by bFGF. Although the kinetics of CREB phosphorylation by EGF was transient, bFGF induced a prolonged pattern of CREB phosphorylation. Interestingly, bFGF-induced CREB phosphorylation and subsequent CRE-mediated gene transcription is not likely to be mediated by any of previously known signaling pathways that lead to phosphorylation of CREB, such as mitogen-activated protein kinases, protein kinase A, protein kinase C, phosphatidylinositol 3-kinase-p70(S6K), calcium/calmodulin dependent protein kinase, and casein kinase 2. By using in vitro in gel kinase assay the presence of a novel 120-kDa bFGF-inducible CREB kinase was identified. These findings identify a new growth factor-activated signaling pathway that regulates gene expression at the CRE. PMID- 11278710 TI - The binding of oxidized low density lipoprotein (ox-LDL) to ox-LDL receptor-1 reduces the intracellular concentration of nitric oxide in endothelial cells through an increased production of superoxide. AB - Oxidized low density lipoprotein (ox-LDL) has been suggested to affect endothelium-dependent vascular tone through a decreased biological activity of endothelium-derived nitric oxide (NO). Oxidative inactivation of NO is regarded as an important cause of its decreased biological activity, and in this context superoxide (O(2)) is known to inactivate NO in a chemical reaction during which peroxynitrite is formed. In this study we examined the effect of ox-LDL on the intracellular NO concentration in bovine aortic endothelial cells and whether this effect is influenced by ox-LDL binding to the endothelial receptor lectin like ox-LDL receptor-1 (LOX-1) through the formation of reactive oxygen species and in particular of O(2). ox-LDL induced a significant dose-dependent decrease in intracellular NO concentration both in basal and stimulated conditions after less than 1 min of incubation with bovine aortic endothelial cells (p < 0.01). In the same experimental conditions ox-LDL also induced O(2) generation (p < 0.001). In the presence of radical scavengers and anti-LOX-1 monoclonal antibody, O(2) formation induced by ox-LDL was reduced (p < 0.001) with a contemporary rise in intracellular NO concentration (p < 0.001). ox-LDL did not significantly modify the ability of endothelial nitric oxide synthase to metabolize l-arginine to l citrulline. The results of this study show that one of the pathophysiological consequences of ox-LDL binding to LOX-1 may be the inactivation of NO through an increased cellular production of O(2). PMID- 11278711 TI - A novel tobacco mitogen-activated protein (MAP) kinase kinase, NtMEK1, activates the cell cycle-regulated p43Ntf6 MAP kinase. AB - Two-hybrid screening of a tobacco BY-2 cell suspension cDNA library using the p43(Ntf6) mitogen-activated protein (MAP) kinase as bait resulted in the isolation of a cDNA encoding a protein with features characteristic of a MAP kinase kinase (MEK), which has been called NtMEK1. Two-hybrid interaction analysis and pull-down experiments showed a physical interaction between NtMEK1 and the tobacco MAP kinases p43(Ntf6) and p45(Ntf4), but not p43(Ntf3). In kinase assays NtMEK1 preferentially phosphorylated p43(Ntf6). Functional studies in yeast showed that p43(Ntf6) could complement the yeast MAP kinase mutant mpk1 when co-expressed with NtMEK1, and that this complementation depended on the kinase activity of p43(Ntf6). Expression analysis showed that the NtMEK1 and ntf6 genes are co-expressed both in plant tissues and following the induction of cell division in leaf pieces. These data suggest that NtMEK1 is an MEK for the p43(Ntf6) MAP kinase. PMID- 11278712 TI - Enac degradation in A6 cells by the ubiquitin-proteosome proteolytic pathway. AB - Amiloride-sensitive epithelial Na(+) channels (ENaC) are responsible for trans epithelial Na(+) transport in the kidney, lung, and colon. The channel consists of three subunits (alpha, beta, gamma) each containing a proline rich region (PPXY) in their carboxyl-terminal end. Mutations in this PPXY domain cause Liddle's syndrome, an autosomal dominant, salt-sensitive hypertension, by preventing the channel's interactions with the ubiquitin ligase Neural precursor cell-expressed developmentally down-regulated protein (Nedd4). It is postulated that this results in defective endocytosis and lysosomal degradation of ENaC leading to an increase in ENaC activity. To show the pathway that degrades ENaC in epithelial cells that express functioning ENaC channels, we used inhibitors of the proteosome and measured sodium channel activity. We found that the inhibitor, MG-132, increases amiloride-sensitive trans-epithelial current in Xenopus distal nephron A6 cells. There also is an increase of total cellular as well as membrane associated ENaC subunit molecules by Western blotting. MG-132-treated cells also have increased channel density in patch clamp experiments. Inhibitors of lysosomal function did not reproduce these findings. Our results suggest that in native renal cells the proteosomal pathway is an important regulator of ENaC function. PMID- 11278713 TI - Plasminogen activator inhibitor type 2 contains mRNA instability elements within exon 4 of the coding region. Sequence homology to coding region instability determinants in other mRNAs. AB - Plasminogen activator inhibitor type 2 (PAI-2) is a serine protease inhibitor that inhibits urokinase. Constitutive and regulated PAI-2 gene expression involves post-transcriptional events, and an AU-rich mRNA instability motif within the 3'-untranslated region of PAI-2 mRNA is required for this process (Maurer, F., Tierney, M., and Medcalf, R. L. (1999) Nucleic Acids Res. 27, 1664 1673). Here we show that instability determinants are present within various exons of the PAI-2 coding region, most notably within exon 4. Deletion of exon 4 from the full-length PAI-2 cDNA results in a doubling in the half-life of PAI-2 mRNA, whereas a 28-nucleotide region within exon 4 contains binding sites for cytoplasmic proteins. Inducible stabilization of PAI-2 mRNA in HT-1080 cells treated with phorbol ester and tumor necrosis factor does not alter the binding of proteins to the exon 4 instability determinant, but resulted in a transient increase in the binding of factors to the AU-rich RNA instability element. Hence, PAI-2 mRNA stability is influenced by elements located within both the coding region and the 3'-untranslated region and that cytoplasmic mRNA binding factors may influence steady state and inducible PAI-2 mRNA expression. Finally a 10 nucleotide region flanking the exon 4 protein-binding site is homologous to instability elements within five other transcripts, suggesting that a common coding region determinant may exist. PMID- 11278714 TI - Glycosylation signals that separate the trimerization from the mhc class II binding domain control intracellular degradation of invariant chain. AB - Invariant chain (Ii) serves as a chaperone for folding and intracellular transport of major histocompatibility complex class II (MHCII) molecules. Early in biosynthesis, Ii associates with MHCII molecules and directs their intracellular transport to endocytic compartments where vesicular proteinases sequentially release Ii from the MHCII heterodimer. The detachment of Ii makes the MHCII groove susceptible for binding of antigenic peptides. We investigated the role of N-linked glycosylation in the controlled intracellular degradation of Ii. Motifs for asparagine-linked glycosylation were altered, and mutated Ii (IiNmut) was transiently expressed in COS cells. The half-life of IiNmut was strongly reduced compared with wild-type Ii although the sensitivity of the N glycan-free polypeptide to in vitro proteinase digestion was not substantially increased. Inhibition of vesicular proteinases revealed endosomal degradation of IiNmut. Intracellular proteolysis of IiNmut is substantially impaired by serine proteinase inhibitors. Thus, a considerable amount of IiNmut is degraded in nonacidic intracellular compartments. The data suggest that N-linked glycosylation of Ii hinders premature proteolysis in nonacidic vesicles resulting in Ii degradation in acidic MHC class II-processing compartments. PMID- 11278715 TI - Stathmin family proteins display specific molecular and tubulin binding properties. AB - Stathmin family phosphoproteins (stathmin, SCG10, SCLIP, and RB3/RB3'/RB3") are involved in signal transduction and regulation of microtubule dynamics. With the exception of stathmin, they are expressed exclusively in the nervous system, where they display different spatio-temporal and functional regulations and hence play at least partially distinct and possibly complementary roles in relation to the control of development, plasticity, and neuronal activities. At the molecular level, each possesses a specific "stathmin-like domain" and, with the exception of stathmin, various combinations of N-terminal extensions involved in their association with intracellular membrane compartments. We show here that each stathmin-like domain also displays specific biochemical and tubulin interaction properties. They are all able to sequester two alpha/beta tubulin heterodimers as revealed by their inhibitory action on tubulin polymerization and by gel filtration. However, they differ in the stabilities of the complexes formed as well as in their interaction kinetics with tubulin followed by surface plasmon resonance as follows: strong stability and slow kinetics for RB3; medium for SCG10, SCLIP, and stathmin; and weak stability and rapid kinetics for RB3'. These results suggest that the fine-tuning of their stathmin-like domains contributes to the specific functional roles of stathmin family proteins in the regulation of microtubule dynamics within the various cell types and subcellular compartments of the developing or mature nervous system. PMID- 11278716 TI - Base excision and DNA binding activities of human alkyladenine DNA glycosylase are sensitive to the base paired with a lesion. AB - The human alkyladenine DNA glycosylase has a broad substrate specificity, excising a structurally diverse group of damaged purines from DNA. To more clearly define the structural and mechanistic bases for substrate specificity of human alkyladenine DNA glycosylase, kinetics of excision and DNA binding activities were measured for several different damaged and undamaged purines within identical DNA sequence contexts. We found that 1,N(6)-ethenoadenine (epsilonA) and hypoxanthine (Hx) were excised relatively efficiently, whereas 7,8 dihydro-8-oxoguanine, O(6)-methylguanine, adenine, and guanine were not. Single turnover kinetics of excision of Hx and epsilonA paired with T showed that excision of Hx was about four times faster than epsilonA, whereas binding assays showed that the binding affinity was about five times greater for epsilonA than for Hx. The opposing pyrimidine base had a significant effect on the kinetics of excision and DNA binding affinity of Hx but a small effect on those for epsilonA. Surprisingly, replacing a T with a U opposite Hx dramatically reduced the excision rate by a factor of 15 and increased the affinity by a factor of 7-8. The binding affinity of human alkyladenine DNA glycosylase to a DNA product containing an abasic site was similar to that for an Hx lesion. PMID- 11278717 TI - A plant 3'-phosphoesterase involved in the repair of DNA strand breaks generated by oxidative damage. AB - Two novel, structurally and functionally distinct phosphatases have been identified through the functional complementation, by maize cDNAs, of an Escherichia coli diphosphonucleoside phosphatase mutant strain. The first, ZmDP1, is a classical Mg(2+)-dependent and Li(+)-sensitive diphosphonucleoside phosphatase that dephosphorylates both 3'-phosphoadenosine 5'-phosphate (3'-PAP) and 2'-PAP without any discrimination between the 3'- and 2'-positions. The other, ZmDP2, is a distinct phosphatase that also catalyzes diphosphonucleoside dephosphorylation, but with a 12-fold lower Li(+) sensitivity, a strong preference for 3'-PAP, and the unique ability to utilize double-stranded DNA molecules with 3'-phosphate- or 3'-phosphoglycolate-blocking groups as substrates. Importantly, ZmDP2, but not ZmDP1, conferred resistance to a DNA repairdeficient E. coli strain against oxidative DNA-damaging agents generating 3'-phosphate- or 3'-phosphoglycolate-blocked single strand breaks. ZmDP2 shares a partial amino acid sequence similarity with a recently identified human polynucleotide kinase 3'-phosphatase that is thought to be involved in DNA repair, but is devoid of 5'-kinase activity. ZmDP2 is the first DNA 3' phosphoesterase thus far identified in plants capable of converting 3'-blocked termini into priming sites for reparative DNA polymerization. PMID- 11278718 TI - Atpase activity and multimer formation of Pilq protein are required for thin pilus biogenesis in plasmid R64. AB - Plasmid R64 pilQ gene is essential for the formation of thin pilus, a type IV pilus. The pilQ product contains NTP binding motifs and belongs to the PulE VirB11 family of NTPases. The pilQ gene was overexpressed with an N-terminal His tag, and PilQ protein was purified. Purified His tag PilQ protein displayed ATPase activity with a V(max) of 0.71 nmol/min/mg of protein and a K(m) of 0.26 mm at pH 6.5. By gel filtration chromatography, PilQ protein was eluted at the position corresponding to 460 kDa, suggesting that PilQ protein forms a homooctamer. To analyze the relationship between structure and function of PilQ protein, amino acid substitutions were introduced within several conserved motifs. Among 11 missense mutants, 7 mutants exhibited various levels of reduced DNA transfer frequencies in liquid matings. Four mutant genes (T234I, K238Q, D263N, and H328A) were overexpressed with a His tag. The purified mutant PilQ proteins contained various levels of reduced ATPase activity. Three mutant PilQ proteins formed stable multimers similar to wild-type PilQ, whereas the PilQ D263N multimer was unstable. PilQ D263N monomer exhibited low ATPase activity, while PilQ D263N multimer did not. These results indicate that ATPase activity of the PilQ multimer is essential for R64 thin pilus biogenesis. PMID- 11278719 TI - Homo- and heterooligomeric SNARE complexes studied by site-directed spin labeling. AB - SNARE (soluble NSF acceptor protein receptor) proteins are thought to mediate membrane fusion by assembling into heterooligomeric complexes that connect the fusing membranes and initiate the fusion reaction. Here we used site-directed spin labeling to map conformational changes that occur upon homo- and heterooligomeric complex formation of neuronal SNARE proteins. We found that the soluble domains of synaptobrevin, SNAP-25, and syntaxin 1 are unstructured. At higher concentrations, the SNARE motif of syntaxin 1 forms homooligomeric helical bundles with at least some of the alpha-helices aligned in parallel. In the assembled SNARE complex, mapping of thirty side chain positions yielded spectra which are in good agreement with the recently published crystal structure. The loop region of SNAP-25 that connects the two SNARE motifs is largely unstructured. C-terminal truncation of synaptobrevin resulted in complexes that are completely folded N-terminal of the truncation but become unstructured at the C-terminal end. The binary complex of syntaxin and SNAP-25 consists of a parallel four helix-bundle with properties resembling that of the ternary complex. PMID- 11278720 TI - Identification of anaplastic lymphoma kinase as a receptor for the growth factor pleiotrophin. AB - Pleiotrophin (PTN) is a secreted growth factor that induces neurite outgrowth and is mitogenic for fibroblasts, epithelial, and endothelial cells. During tumor growth PTN can serve as an angiogenic factor and drive tumor invasion and metastasis. To identify a receptor for PTN, we panned a phage display human cDNA library against immobilized PTN protein as a bait. From this we isolated a phage insert that was homologous to an amino acid sequence stretch in the extracellular domain (ECD) of the orphan receptor tyrosine kinase anaplastic lymphoma kinase (ALK). In parallel with PTN, ALK is highly expressed during perinatal development of the nervous system and down-modulated in the adult. Here we show in cell-free assays as well as in radioligand receptor binding studies in intact cells that PTN binds to the ALK ECD with an apparent Kd of 32 +/- 9 pm. This receptor binding is inhibited by an excess of PTN, by the ALK ECD, and by anti-PTN and anti-ECD antibodies. PTN added to ALK-expressing cells induces phosphorylation of both ALK and of the downstream effector molecules IRS-1, Shc, phospholipase C gamma, and phosphatidylinositol 3-kinase. Furthermore, the growth stimulatory effect of PTN on different cell lines in culture coincides with the endogenous expression of ALK mRNA, and the effect of PTN is enhanced by ALK overexpression. From this we conclude that ALK is a receptor that transduces PTN-mediated signals and propose that the PTN-ALK axis can play a significant role during development and during disease processes. PMID- 11278721 TI - Targeting of Kruppel-associated box-containing zinc finger proteins to centromeric heterochromatin. Implication for the gene silencing mechanisms. AB - Kruppel-associated box-containing zinc finger proteins (KRAB-ZFPs) repress transcription via functional interaction with the corepressor KRAB-associated protein-1 (KAP-1). KAP-1 directly interacts with heterochromatin protein 1 (HP1), a dose-dependent regulator of heterochromatin-mediated silencing. Here we show that two KRAB-ZFPs that we previously identified, KRAZ1 and KRAZ2, are targeted to foci of centromeric heterochromatin containing HP1alpha through the interaction with KAP-1. Centromeric targeting potential of KRAZ1 and KAP-1 is strictly correlated with their silencing activities; a KRAB mutant of KRAZ1 that is unable to bind KAP-1 and KAP-1 deletions unable to bind HP1 cannot localize to centromeric foci nor repress transcription. We provide evidence that this correlation is likely to be functionally relevant. First, overexpression of the VP16 transactivation domain fused with the KAP-1 deletion that binds to KRAB but not to HP1 leads to dramatic redistribution of KRAZ1 from centromeric foci and simultaneously converts KRAZ1-mediated silencing into strong transcriptional activation. Second, a specific inhibitor of histone deacetylases, trichostatin A, effectively redistributes KRAZ1 and KAP-1 from centromeric foci and partially relieves their silencing activities. These data strongly suggest that KRAB ZFPs/KAP-1 silence transcription by dynamic recruitment of the target locus to the specific gene silencing compartment, centromeric heterochromatin, in a histone deacetylase-dependent manner. PMID- 11278722 TI - High affinity interaction of yeast transcriptional regulator, Mot1, with TATA box binding protein (TBP). AB - Yeast Mot1, an essential ATP-dependent regulator of basal transcription, removes TATA box-binding protein (TBP) from TATA sites in vitro. Complexes of Mot1 and Spt15 (yeast TBP), radiolabeled in vitro, were immunoprecipitated with anti-TBP (or anti-Mot1) antibodies in the absence of DNA, showing Mot1 binds TBP in solution. Mot1 N-terminal deletions (residues 25-801) abolished TBP binding, whereas C-terminal ATPase domain deletions (residues 802-1867) did not. Complex formation was prevented above 200 mm salt, consistent with electrostatic interaction. Correspondingly, TBP variants lacking solvent-exposed positive charge did not bind Mot1, whereas a mutant lacking positive charge within the DNA binding groove bound Mot1. ATPase-defective mutant, Mot1(D1408N), which inhibits growth when overexpressed (but is suppressed by co-overexpression of TBP), bound TBP normally in vitro, suggesting it forms nonrecyclable complexes. N-terminal deletions of Mot1(D1408N) were not growth-inhibitory. C-terminal deletions were toxic when overexpressed, and toxicity was ameliorated by TBP co-overproduction. Residues 1-800 of Mot1 are therefore necessary and sufficient for TBP binding. The N terminus of 89B, a tissue-specific Drosophila Mot1 homolog, bound the TBP like factor, dTRF1. Native Mot1 and derivatives deleterious to growth localized in the nucleus, whereas nontoxic derivatives localized to the cytosol, suggesting TBP binding and nuclear transport of Mot1 are coupled. PMID- 11278723 TI - Activation of caspase-12, an endoplastic reticulum (ER) resident caspase, through tumor necrosis factor receptor-associated factor 2-dependent mechanism in response to the ER stress. AB - When accumulation of a malfolded protein in the endoplastic reticulum (ER) is induced by various adverse conditions, such as hypoxia, glucose starvation, and perturbation of calcium homeostasis, cells respond to the stress by increasing transcription of genes encoding ER molecular chaperones, a process known as unfolded protein response. The signaling is initiated by IRE1s, ER stress sensors. Alternatively, excessive stress to the ER results in apoptosis. Caspase 12 is known to be essential for this ER stress-induced apoptosis. In this study, we analyzed the detailed regulatory mechanisms of IRE1s during ER stress. We identified c-Jun N-terminal inhibitory kinase (JIK) as a binding partner of IRE1alpha, and JIK was seen to modulate IRE1alpha-TRAF2 (tumor necrosis factor receptor-associated factor 2) complex formation and the resultant alteration to c Jun N-terminal kinase signaling from IRE1s in response to ER stress. We also demonstrated that TRAF2 interacts with procaspase-12 and promotes the clustering of procaspase-12 and its activation by cleavage in response to ER stress. These results indicate that TRAF2 plays crucial roles not only in the signaling of the c-Jun N-terminal kinase pathway but also in activation of caspase-12 to transduce signals from IRE1s. Thus, we provide a missing link in the ER stress-induced apoptosis-signaling pathway, one which connects the stress sensor molecule IRE1 and the activation of caspase-12. PMID- 11278724 TI - A two-receptor pathway for catabolism of Clara cell secretory protein in the kidney. AB - Clara cell secretory protein (CCSP) is a transport protein for lipophilic substances in bronchio-alveolar fluid, plasma, and uterine secretion. It acts as a carrier for steroid hormones and polychlorinated biphenyl metabolites. Previously, the existence of receptors for uptake of CCSP.ligand complexes into the renal proximal tubules had been suggested. Using surface plasmon resonance analysis, we demonstrate that CCSP binds to cubilin, a peripheral membrane protein on the surface of proximal tubular cells. Binding to cubilin results in uptake and lysosomal degradation of CCSP in cultured cells. Surprisingly, internalization of CCSP is blocked not only by cubilin antagonists but also by antibodies directed against megalin, an endocytic receptor that does not bind CCSP but associates with cubilin. Consistent with a role of both receptors in renal uptake of CCSP in vivo, patients deficient for cubilin or mice lacking megalin exhibit a defect in tubular uptake of the protein and excrete CCSP into the urine. These findings identify a cellular pathway consisting of a CCSP binding protein (cubilin) and an endocytic coreceptor (megalin) responsible for tissue-specific uptake of CCSP and associated ligands. PMID- 11278726 TI - Identification of the active oligomeric state of an essential adenine DNA methyltransferase from Caulobacter crescentus. AB - Caulobacter crescentus contains one of the two known prokaryotic DNA methyltransferases that lacks a cognate endonuclease. This endogenous cell cycle regulated adenine DNA methyltransferase (CcrM) is essential for C. crescentus cellular viability. DNA methylation catalyzed by CcrM provides an obligatory signal for the proper progression through the cell cycle. To further our understanding of the regulatory role played by CcrM, we sought to investigate its biophysical properties. In this paper we employed equilibrium ultracentrifugation, velocity ultracentrifugation, and chemical cross-linking to show that CcrM is dimeric at physiological concentrations. However, surface plasmon resonance experiments in the presence of S-adenosyl-homocysteine evince that CcrM binds as a monomer to a defined hemi-methylated DNA substrate containing the canonical methylation sequence, GANTC. Initial velocity experiments demonstrate that dimerization of CcrM does not affect DNA methylation. Collectively, these findings suggest that CcrM is active as a monomer and provides a possible in vivo role for dimerization as a means to stabilize CcrM from premature catabolism. PMID- 11278725 TI - Phosphorylation of the tumor necrosis factor receptor CD120a (p55) recruits Bcl-2 and protects against apoptosis. AB - Ligation of the tumor necrosis factor alpha receptor CD120a initiates responses as diverse as apoptosis and the expression of NF-kappaB-dependent pro-survival genes. How these opposing responses are controlled remains poorly understood. Here we demonstrate that phosphorylation by p42(mapk/erk2) inhibits the apoptotic activity of CD120a while preserving its ability to activate NF-kappaB. Phosphorylated CD120a is re-localized from the Golgi complex to tubular structures of the endoplasmic reticulum wherein it recruits Bcl-2. Antisense mediated down-regulation of Bcl-2 antagonized the localization of CD120a to tubular structures and reversed the protection from apoptosis conferred by receptor phosphorylation. We propose that phosphorylation of CD120a represents a novel, Bcl-2-dependent mechanism by which the apoptotic activity of the receptor may be regulated. Thus, oncogenic activation of p42(mapk/erk2) may serve to inhibit the apoptotic activity of this death receptor while preserving NF-kappaB dependent responses and may thus indirectly contribute to a failure to eliminate cells bearing oncogenes of the Ras-Raf-MEK-p42(mapk/erk2) pathway. PMID- 11278727 TI - Hypoxia-induced proliferative response of vascular adventitial fibroblasts is dependent on g protein-mediated activation of mitogen-activated protein kinases. AB - Hypoxia has been shown to act as a proliferative stimulus for adventitial fibroblasts of the pulmonary artery. The signaling pathways involved in this growth response, however, remain unclear. We tested the hypothesis that hypoxia induced proliferation of fibroblasts would be dependent on distinct (compared with serum) activation and utilization patterns of mitogen-activated protein (MAP) kinases initiated by Galpha(i/o) proteins. We found that hypoxia stimulated increases in DNA synthesis and growth of quiescent fibroblasts in the absence of exogenous mitogens and also markedly augmented serum-stimulated growth responses. Hypoxia caused a transient activation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), the time course and pattern of which was somewhat similar to that induced by serum but which was of lesser magnitude. On the other hand, hypoxia-induced activation of p38 MAP kinase was biphasic, whereas serum-stimulated activation of p38 MAP kinase was transient, and the magnitude of activation was greater for hypoxia compared with that of serum stimulation. ERK1/2, JNK1, and p38 MAP kinase but not JNK2 were necessary for hypoxia-induced proliferation because PD98059, SB202190, and JNK1 antisense oligonucleotides nearly ablated the growth response. JNK2 appeared to act as a negative modulator of hypoxia-induced growth because JNK2 antisense oligonucleotides led to an increase in DNA synthesis. In serum-stimulated cells, antisense JNK1 oligonucleotides and PD98059 had inhibitory effects on proliferation, whereas SB202190 led to an increase in DNA synthesis. Pertussis toxin, which blocks Galpha(i/o)-mediated signaling, markedly attenuated hypoxia induced DNA synthesis and activation of ERK and JNK but not p38 MAP kinase. We conclude that hypoxia itself can act as a growth promoting stimulus for subsets of bovine neonatal adventitial fibroblasts largely through Galpha(i/o)-mediated activation of a complex network of MAP kinases whose specific contributions to hypoxia-induced proliferation differ from traditional serum-induced growth signals. PMID- 11278729 TI - Co-operative effect of c-Src tyrosine kinase and Stat3 in activation of hepatocyte growth factor expression in mammary carcinoma cells. AB - We have previously shown coexpression of hepatocyte growth factor (HGF) and its receptor Met in the invasive tumor front of human breast carcinomas. We have also demonstrated secretion of HGF, constitutive activation of Met, and increased invasion in a murine breast carcinoma cell line, SP1. These observations suggest the presence of an HGF autocrine loop in some breast carcinoma cells, which confers increased survival, growth, and invasiveness during tumor progression and metastasis. c-Src tyrosine kinase, which is critical in regulating the expression of many genes, is activated in SP1 carcinoma cells, as well as in most human breast cancers. We therefore examined the role of c-Src kinase in HGF expression in breast carcinoma cells. Expression of activated c-Src in SP1 cells increased transcription from the HGF promoter and expression of HGF mRNA and protein, while dominant negative c-Src had the opposite effect. Using deletion analysis, we showed that the region between -254 and -70 base pairs was required for c-Src responsiveness of the HGF promoter. This region contains two putative consensus sequences (at -110 and -149 base pairs) for the Stat3 transcription factor, which bind protein complexes containing Stat3 (but not Stat1, -5A, or -5B). Coexpression of activated c-Src and Stat3 synergistically induced strong HGF promoter activity in SP1 cells, as well as in a nonmalignant epithelial cell line, HC11 (HGF negative). c-Src kinase activity correspondingly increased the tyrosine 705 phosphorylation and DNA binding affinity of Stat3 (but not Stat1, 5A, or -5B). Collectively, our data indicate a cooperative effect of c-Src kinase and Stat3 in the activation of HGF transcription and protein expression in breast carcinoma cells. This process may be important in overriding the strong repression of HGF expression in nonmalignant epithelium, and thereby promote tumorigenesis. PMID- 11278728 TI - J-domain protein, Jac1p, of yeast mitochondria required for iron homeostasis and activity of Fe-S cluster proteins. AB - J-proteins are molecular chaperones with a characteristic domain predicted to mediate interaction with Hsp70 proteins. We have previously isolated yeast mutants of the mitochondrial Hsp70, Ssq1p, in a genetic screen for mutants with altered iron homeostasis. Here we describe the isolation of mutants of the J domain protein, Jac1p, using the same screen. Mutant jac1 alleles predicted to encode severely truncated proteins (lacking 70 or 152 amino acids) were associated with phenotypes strikingly similar to the phenotypes of ssq1 mutants. These phenotypes include activation of the high affinity cellular iron uptake system and iron accumulation in mitochondria. In contrast to iron accumulation, Fe-S proteins of mitochondria were specifically deficient. In jac1 mutants, like in ssq1 mutants, processing of the Yfh1p precursor protein from intermediate to mature forms was delayed. In the genetic backgrounds used in this study, jac1 null mutants were found to be viable, permitting analysis of genetic interactions. The Deltajac1 Deltassq1 double mutant was more severely compromised for growth than either single mutant, suggesting a synthetic or additive effect of these mutations. Overexpression of Jac1p partially suppressed ssq1 slow growth and vice versa. Similar mitochondrial localization and similar mutant phenotypes suggest that Ssq1p and Jac1p are functional partners in iron homeostasis. PMID- 11278730 TI - A family of 16-kDa pancreatic secretory stress proteins form highly organized fibrillar structures upon tryptic activation. AB - A group of 16-kDa proteins, synthesized and secreted by rat pancreatic acinar cells and composed of pancreatic stone protein (PSP/reg) and isoforms of pancreatitis-associated protein (PAP), show structural homologies, including conserved amino acid sequences, cysteine residues, and highly sensitive N terminal trypsin cleavage sites, as well as conserved functional responses in conditions of pancreatic stress. Trypsin activation of recombinant stress proteins or counterparts contained in rat pancreatic juice (PSP/reg, PAP I and PAP III) resulted in conversion of 16-kDa soluble proteins into 14-kDa soluble isoforms (pancreatic thread protein and pancreatitis-associated thread protein, respectively) that rapidly polymerize into insoluble sedimenting structures. Activated thread proteins show long lived resistance to a wide spectrum of proteases contained in pancreatic juice, including serine proteases and metalloproteinases. In contrast, PAP II, following activation with trypsin or pancreatic juice, does not form insoluble structures and is rapidly digested by pancreatic proteases. Scanning and transmission electron microscopy indicate that activated thread proteins polymerize into highly organized fibrillar structures with helical configurations. Through bundling, branching, and extension processes, these fibrillar structures form dense matrices that span large topological surfaces. These findings suggest that PSP/reg and PAP I and III isoforms consist of a family of highly regulated soluble secretory stress proteins, which, upon trypsin activation, convert into a family of insoluble helical thread proteins. Dense extracellular matrices, composed of helical thread proteins organized into higher ordered matrix structures, may serve physiological functions within luminal compartments in the exocrine pancreas. PMID- 11278731 TI - Static pressure regulates connective tissue growth factor expression in human mesangial cells. AB - Connective tissue growth factor (CTGF) is overexpressed in a variety of fibrotic disorders such as renal fibrosis and atherosclerosis. Fibrosis is a common final pathway of renal diseases of diverse etiology, including inflammation, hemodynamics, and metabolic injury. Mechanical strains such as stretch, shear stress, and static pressure are possible regulatory elements in CTGF expression. In this study, we examined the ability of static pressure to modulate CTGF gene expression in cultured human mesangial cells. Low static pressure (40-80 mm Hg) stimulated cell proliferation via a protein kinase C-dependent pathway. In contrast, high static pressure (100-180 mm Hg) induced apoptosis in human mesangial cells. This effect was reversed by treatment with CTGF antisense oligonucleotide but not with transforming growth factor beta1-neutralizing antibody or protein kinase C inhibitor. High static pressure not only up regulated the expression of CTGF, but also the expression of extracellular matrix proteins (collagen I and IV, laminin). This up-regulation of extracellular matrix proteins was also reversed by treatment with CTGF antisense oligonucleotide. As judged by mRNA expression of a total of 1100 genes, including apoptosis associated genes using DNA microarray techniques, recombinant CTGF protein induced apoptosis by down-regulation of a number of anti-apoptotic genes. Overexpression of CTGF in mesangial cells by transient transfection had similar effects. Taken together, these results suggest that high blood pressure up regulates CTGF expression in mesangial cells. High levels of CTGF in turn enhance extracellular matrix production and induce apoptosis in mesangial cells, and may contribute to remodeling of mesangium and ultimately glomerulosclerosis. PMID- 11278732 TI - Serine proteinase inhibitor 3 and murinoglobulin I are potent inhibitors of neuropsin in adult mouse brain. AB - Extracellular serine protease neuropsin (NP) is expressed in the forebrain limbic area of adult brain and is implicated in synaptic plasticity. We screened for endogenous NP inhibitors with recombinant NP (r-NP) from extracts of the hippocampus and the cerebral cortex in adult mouse brain. Two SDS-stable complexes were detected, and after their purification, peptide sequences were determined by amino acid sequencing and mass spectrometry, revealing that target molecules were serine proteinase inhibitor-3 (SPI3) and murinoglobulin I (MUG I). The addition of the recombinant SPI3 to r-NP resulted in an SDS-stable complex, and the complex formation followed bimolecular kinetics with an association rate constant of 3.4 +/- 0.22 x 10(6) M(-1) s(-1), showing that SPI3 was a slow, tight binding inhibitor of NP. In situ hybridization histochemistry showed that SPI3 mRNA was expressed in pyramidal neurons in the hippocampal CA1-CA3 subfields, as was NP mRNA. Alternatively, the addition of purified plasma MUG I to r-NP resulted in an SDS-stable complex, and MUG I inhibited degradation of fibronectin by r-NP to 24% at a r-NP/MUG I molar ratio of 1:2. Immunofluorescence histochemistry showed that MUG I localized in the hippocampal neurons. These findings indicate that SPI3 and MUG I serve to inactivate NP and control the level of NP in adult brain, respectively. PMID- 11278733 TI - Structure-activity relationship studies of melanin-concentrating hormone (MCH) related peptide ligands at SLC-1, the human MCH receptor. AB - Melanin-concentrating hormone (MCH) is a cyclic nonadecapeptide involved in the regulation of feeding behavior, which acts through a G protein-coupled receptor (SLC-1) inhibiting adenylcyclase activity. In this study, 57 analogues of MCH were investigated on the recently cloned human MCH receptor stably expressed in HEK293 cells, on both the inhibition of forskolin-stimulated cAMP production and guanosine-5'-O-(3-[(35)S]thiotriphosphate ([(35)S]- GTPgammaS) binding. The dodecapeptide MCH-(6-17) (MCH ring between Cys(7) and Cys(16), with a single extra amino acid at the N terminus (Arg(6)) and at the C terminus (Trp(17))) was found to be the minimal sequence required for a full and potent agonistic response on cAMP formation and [(35)S]- GTPgammaS binding. We Ala-scanned this dodecapeptide and found that only 3 of 8 amino acids of the ring, namely Met(8), Arg(11), and Tyr(13), were essential to elicit full and potent responses in both tests. Deletions inside the ring led either to inactivity or to poor antagonists with potencies in the micromolar range. Cys(7) and Cys(16) were substituted by Asp and Lys or one of their analogues, in an attempt to replace the disulfide bridge by an amide bond. However, those modifications were deleterious for agonistic activity. In [(35)S]- GTPgammaS binding, these compounds behaved as weak antagonists (K(B) 1-4 microm). Finally, substitution in MCH-(6-17) of 6 out of 12 amino acids by non-natural residues and concomitant replacement of the disulfide bond by an amide bond led to three compounds with potent antagonistic properties (K(B) = 0.1-0.2 microm). Exploitation of these structure-activity relationships should open the way to the design of short and stable MCH peptide antagonists. PMID- 11278734 TI - Cysteine cross-linking defines part of the dimer and B/C domain interface of the Escherichia coli mannitol permease. AB - Part of the dimer and B/C domain interface of the Escherichia coli mannitol permease (EII(mtl)) has been identified by the generation of disulfide bridges in a single-cysteine EII(mtl), with only the activity linked Cys(384) in the B domain, and in a double-cysteine EII(mtl) with cysteines at positions 384 and 124 in the first cytoplasmic loop of the C domain. The disulfide bridges were formed in the enzyme in inside-out membrane vesicles and in the purified enzyme by oxidation with Cu(II)-(1,10-phenanthroline)(3), and they were visualized by SDS polyacrylamide gel electrophoresis. Discrimination between possible disulfide bridges in the dimeric double-cysteine EII(mtl) was done by partial digestion of the protein and the formation of heterodimers, in which the cysteines were located either on different subunits or on one subunit. The disulfide bridges that were identified are an intersubunit Cys(384)-Cys(384), an intersubunit Cys(124)-Cys(124), an intersubunit Cys(384)-Cys(124), and an intrasubunit Cys(384)-Cys(124). The disulfide bridges between the B and C domain were observed with purified enzyme and confirmed by matrix-assisted laser desorption ionization time of flight mass spectrometry. Mannitol did not influence the formation of the disulfide between Cys(384) and Cys(124). The close proximity of the two cysteines 124 was further confirmed with a separate C domain by oxidation with Cu(II)-(1,10 phenanthroline)(3) or by reactions with dimaleimides of different length. The data in combination with other work show that the first cytoplasmic loop around residue 124 is located at the dimer interface and involved in the interaction between the B and C domain. PMID- 11278735 TI - Biochemical and pharmacological criteria define two shedding activities for TRANCE/OPGL that are distinct from the tumor necrosis factor alpha convertase. AB - A number of structurally and functionally diverse membrane proteins are released from the plasma membrane in a process termed protein ectodomain shedding. Ectodomain shedding may activate or inactivate a substrate or change its properties, such as converting a juxtacrine into a paracrine signaling molecule. Here we have characterized the activities involved in protein ectodomain shedding of the tumor necrosis factor family member TRANCE/OPGL in different cell types. The criteria used to evaluate these activities include (a) cleavage site usage, (b) response to activators and inhibitors of intracellular signaling pathways, and (c) sensitivity to tissue inhibitors of metalloproteases (TIMPs). At least two TRANCE shedding activities emerged, both of which are distinct from the tumor necrosis factor alpha convertase. One of the TRANCE sheddases is induced by the tyrosine phosphatase inhibitor pervanadate but not by phorbol esters, whereas the other is refractory to both of these stimuli. Furthermore, the pervanadate regulated sheddase activity is sensitive to TIMP-2 but not TIMP-1, which is consistent with the properties of a membrane type matrix metalloprotease. This study provides insights into the properties of different activities involved in protein ectodomain shedding and has implications for the functional regulation of TRANCE by potentially more than one protease. PMID- 11278736 TI - A novel class of oxylipins, sn1-O-(12-oxophytodienoyl)-sn2-O-(hexadecatrienoyl) monogalactosyl Diglyceride, from Arabidopsis thaliana. AB - The cyclic derivative of 13(S)-hydroperoxolinolenic acid, 12-oxophytodienoic acid, serves as a signal transducer in higher plants, mediating mechanotransductory processes and plant defenses against a variety of pathogens, and also serves as a precursor for the biosynthesis of jasmonic acid, a mediator of plant herbivore defense. Biosynthesis of 12-oxophytodienoic acid from alpha linolenic acid occurs in plastids, mainly in chloroplasts, and is thought to start with free linolenic acid liberated from membrane lipids by lipase action. In Arabidopsis thaliana, the glycerolipid fraction contains esterified 12 oxophytodienoic acid, which can be released enzymatically by sn1-specific, but not by sn2-specific, lipases. The 12-oxophytodienoyl glycerolipid fraction was isolated, purified, and characterized. Enzymatic, mass spectrometric, and NMR spectroscopic data allowed us to establish the structure of the novel oxylipin as sn1-O-(12-oxophytodienoyl)-sn2-O-(hexadecatrienoyl)-monogalactosyl diglyceride. The novel class of lipids is localized in plastids. Purified monogalactosyl diglyceride was not converted to the sn1-(12-oxophytodienoyl) derivative by the combined action of (soybean) lipoxygenase and (A. thaliana) allene oxide synthase, an enzyme ensemble that converts free alpha-linolenic acid to free 12 oxophytodienoic acid. When leaves were wounded, a significant and transient increase in the level of (12-oxophytodienoyl)-monogalactosyl diglyceride was observed. In A. thaliana, the major fraction of 12-oxophytodienoic acid occurs esterified at the sn1 position of the plastid-specific glycerolipid, monogalactosyl diglyceride. PMID- 11278737 TI - Characterization of a novel synGAP isoform, synGAP-beta. AB - We cloned a cDNA encoding a novel synGAP, synGAP-d (GenBank(TM) accession number ), from a rat brain cDNA library. The clone consisted of 4801 nucleotides with a coding sequence of 3501 nucleotides, encoded a protein consisting of 1166 amino acids with >99% homology with 1092 amino acid overlaps to synGAP, and contained a 13-nucleotide insertion to the previously reported synGAP mRNAs, which suggested that the clone was a splice variant of synGAP. We also found that there are at least seven variants in the 3' portion of the synGAP mRNA and that they encoded five different protein isoforms. The coding sequence of these C-terminal variants were classified into alpha1, alpha2, beta1, beta2, beta3, beta4, and gamma, and synGAP-d was classified as the beta1 form. The previously reported synGAPs (synGAP-a, -b, and -c and p135synGAP) can be classified as the alpha1 isoform. All isoforms were expressed specifically in the brain. Unexpectedly, the beta isoform, which lacks a C-terminal PSD-95-binding motif ((S/T)XV), was more restricted to the postsynaptic density fraction than the motif-containing alpha1 isoform. The beta isoform did not interact with PSD-95 but specifically interacted with a nonphosphorylated alpha subunit of Ca(2+)/calmodulin-dependent protein kinase II through its unique C-terminal tail. PMID- 11278738 TI - Rantes activates Jak2 and Jak3 to regulate engagement of multiple signaling pathways in T cells. AB - The chemokine RANTES (regulated on activation normal T cell expressed and secreted) and its cognate receptor CC chemokine receptor 5 (CCR5) have been implicated in regulating immune cell function. Previously we reported that in T cells, RANTES activation of CCR5 results in Stat1 and Stat3 phosphorylation activation, leading to Stat1:1 and Stat1:3 dimers that exhibit DNA binding activity and the transcriptional induction of a Stat-inducible gene, c-fos. Given that RANTES and CCR5 have been implicated in T cell activation, we have studied RANTES-induced signaling events in a CCR5-expressing T cell line, PM1. RANTES treatment of PM1 T cells results in the rapid phosphorylation-activation of CCR5, Jak2, and Jak3. RANTES-inducible Jak phosphorylation is insensitive to pertussis toxin inhibition, indicating that RANTES-CCR5-mediated tyrosine phosphorylation events are not coupled directly to Galpha(i) protein-mediated events. In addition to Jaks, several other proteins are rapidly phosphorylated on tyrosine residues in a RANTES-dependent manner, including the Src kinase p56(lck), which associates with Jak3. Additionally our data confirm that the amino-terminally modified RANTES proteins, aminooxypentane-RANTES and Met-RANTES, are agonists for CCR5 and induce early tyrosine phosphorylation events that are indistinguishable from those inducible by RANTES with similar kinetics. Our data also demonstrate that RANTES activates the p38 mitogen-activated protein (MAP) kinase pathway. This is evidenced by the rapid RANTES-dependent phosphorylation and activation of p38 MAP kinase as well as the activation of the downstream effector of p38, MAP kinase activated protein (MAPKAP) kinase-2. Pharmacological inhibition of RANTES dependent p38 MAP kinase activation blocks MAPKAP kinase-2 activity. Thus, activation of Jak kinases and p38 MAP kinase by RANTES regulates the engagement of multiple signaling pathways. PMID- 11278739 TI - Distinct roles for Ku protein in transcriptional reinitiation and DNA repair. AB - Transcriptional reinitiation is a distinct phase of the RNA polymerase II transcription cycle. Prior work has shown that reinitiation is deficient in nuclear extracts from Chinese hamster ovary cells lacking the 80-kDa subunit of Ku, a double-strand break repair protein, and that activity is rescued by expression of the corresponding cDNA. We now show that Ku increases the amount or availability of a soluble factor that is limiting for reinitiation, that the factor increases the number of elongation complexes associated with the template at all times during the reaction, and that the factor itself does not form a tight complex with DNA. The factor may consist of a preformed complex of transcription proteins that is stabilized by Ku. A Ku mutant, lacking residues 687-728 in the 80-kDa subunit, preferentially suppresses transcription in Ku containing extracts, suggesting that Ku interacts directly with proteins required for reinitiation. The Ku mutant functions normally in a DNA end-joining system, indicating that the functions of Ku in transcription and repair are genetically separable. Based on our results, we present a model in which Ku is capable of undergoing a switch between a transcription factor-associated and a repair-active state. PMID- 11278740 TI - Human biliverdin reductase is autophosphorylated, and phosphorylation is required for bilirubin formation. AB - Biliverdin reductase (BVR) reduces heme oxygenase (HO) activity product, biliverdin, to bilirubin. BVR is unique in having dual pH/dual cofactor requirements. Using Escherichia coli-expressed human BVR and COS cells, we show that BVR is autophosphorylated and that phosphorylation is required for its activity. An "in blot" autophosphorylation assay showed that BVR is a renaturable phosphoprotein. Controls for the experiments were HO-1 and HO-2; both are phosphoproteins but are not autophosphorylated. Autophosphorylation was pH dependent, with activity at pH 8.7 being most prominent. In addition, 2'(3')-O (2,4,6-trinitrophenyl)adenosine 5'-triphosphate fluorescence titration of BVR gave a lower K(d) at pH 8.7 than at pH 7.4 (15.5 versus 28.0 micrometer). Mn(2+) was required for binding of the ATP analogue and for autophosphorylation; the autokinase activity was lost when treated at 60 degrees C for 10 min. The loss of transferred phosphates by alkaline treatment suggested that BVR is a serine/threonine kinase. Potato acid phosphatase treatment reversibly inactivated the enzyme. The enzyme was also inactivated by treatment with the serine/threonine phosphatase, protein phosphatase 2A; okadaic acid attenuated the inhibition. Titration of protein phosphatase 2A-released phosphates indicated a 1:6 molar ratio of BVR to phosphate. The BVR immunoprecipitated from COS cell lysates was a phosphoprotein, and its activity and phosphorylation levels increased in response to H(2)O(2). The results define a previously unknown mechanism for regulation of BVR activity and are discussed in the context of their relevance to heme metabolism. PMID- 11278741 TI - Mutant Cu/Zn-superoxide dismutase proteins have altered solubility and interact with heat shock/stress proteins in models of amyotrophic lateral sclerosis. AB - Mutations in the Cu/Zn-superoxide dismutase (SOD-1) gene are responsible for a familial form of amyotrophic lateral sclerosis. In humans and experimental models, death of motor neurons is preceded by formation of cytoplasmic aggregates containing mutant SOD-1 protein. In our previous studies, heat shock protein 70 (HSP70) prolonged viability of cultured motor neurons expressing mutant human SOD 1 and reduced formation of aggregates. In this paper, we report that mutant SOD-1 proteins have altered solubility in cells relative to wild-type SOD-1 and can form a direct association with HSP70 and other stress proteins. Whereas wild-type human and endogenous mouse SOD-1 were detergent-soluble, a portion of mutant SOD 1 was detergent-insoluble in protein extracts of NIH3T3 transfected with SOD-1 gene constructs, spinal cord cultures established from G93A SOD-1 transgenic mouse embryos, and lumbar spinal cord from adult G93A transgenic mice. A direct association of HSP70, HSP40, and alphaB-crystallin with mutant SOD-1 (G93A or G41S), but not wild-type or endogenous mouse SOD-1, was demonstrated by coimmunoprecipitation. Mutant SOD-1.HSP70 complexes were predominantly in the detergent-insoluble fraction. However, only a small percentage of total cellular mutant SOD-1 was detergent-insoluble, suggesting that mutation-induced alteration of protein conformation may not in itself be sufficient for direct interaction with heat shock proteins. PMID- 11278742 TI - Nuclear import of Spo12p, a protein essential for meiosis. AB - In Saccharomyces cerevisiae, Spo12p is involved in mitosis and is essential for meiosis. We found that Spo12p is imported into the nucleus by the karyopherin Kap121p. A complex containing Spo12p and Kap121p was isolated from cytosol and was also reconstituted with recombinant proteins, indicating that this interaction is direct. Spo12p was mislocalized to the cytosol in pse1-1, a temperature-sensitive strain harboring a mutation of Kap121p, at the permissive temperature, confirming an essential role for Kap121p in Spo12p import. Spo12p was also mislocalized in a pse1-1/pse1-1 homozygous strain, suggesting it is imported via the same pathway in diploid cells. Furthermore, we found that pse1 1/pse1-1 shows a sporulation defect similar to that of spo12Delta/spo12Delta. In addition, we have characterized the Spo12p nuclear localization signal, mapped it to residues 76-130, and identified residues within this region that are important for nuclear localization signal function. PMID- 11278743 TI - Crystal structure of the Mycobacterium tuberculosis beta-ketoacyl-acyl carrier protein synthase III. AB - Mycolic acids (alpha-alkyl-beta-hydroxy long chain fatty acids) cover the surface of mycobacteria, and inhibition of their biosynthesis is an established mechanism of action for several key front-line anti-tuberculosis drugs. In mycobacteria, long chain acyl-CoA products (C(14)-C(26)) generated by a type I fatty-acid synthase can be used directly for the alpha-branch of mycolic acid or can be extended by a type II fatty-acid synthase to make the meromycolic acid (C(50) C(56)))-derived component. An unusual Mycobacterium tuberculosis beta-ketoacyl acyl carrier protein (ACP) synthase III (mtFabH) has been identified, purified, and shown to catalyze a Claisen-type condensation between long chain acyl-CoA substrates such as myristoyl-CoA (C(14)) and malonyl-ACP. This enzyme, presumed to play a key role in initiating meromycolic acid biosynthesis, was crystallized, and its structure was determined at 2.1-A resolution. The mtFabH homodimer is closely similar in topology and active-site structure to Escherichia coli FabH (ecFabH), with a CoA/malonyl-ACP-binding channel leading from the enzyme surface to the buried active-site cysteine residue. Unlike ecFabH, mtFabH contains a second hydrophobic channel leading from the active site. In the ecFabH structure, this channel is blocked by a phenylalanine residue, which constrains specificity to acetyl-CoA, whereas in mtFabH, this residue is a threonine, which permits binding of longer acyl chains. This same channel in mtFabH is capped by an alpha helix formed adjacent to a 4-amino acid sequence insertion, which limits bound acyl chain length to 16 carbons. These observations offer a molecular basis for understanding the unusual substrate specificity of mtFabH and its probable role in regulating the biosynthesis of the two different length acyl chains required for generation of mycolic acids. This mtFabH presents a new target for structure based design of novel antimycobacterial agents. PMID- 11278744 TI - The p38 MAPK pathway is required for cell growth inhibition of human breast cancer cells in response to activin. AB - Activin, a member of the TGFbeta family inhibits cell growth in various target tissues. Activin interacts with a complex of two receptors that upon activation phosphorylate specific intracellular mediators, the Smad proteins. The activated Smads interact with diverse DNA binding proteins and co-activators of transcription in a cell-specific manner, thus leading to various activin biological effects. In this study, we investigated the role and mechanism of action of activin in the human breast cancer T47D cells. We found that activin treatment of T47D cells leads to a dramatic decrease in cell growth. Thus activin appears as a potent cell growth inhibitor of these breast cancer cells. We show that activin induces the Smad pathway in these cells but also activates the p38 mitogen-activated protein kinase pathway, further leading to phosphorylation of the transcription factor ATF2. Finally, specific inhibitors of the p38 kinase (SB202190, SB203580, and PD169316) but not an inactive analogue (SB202474) or the MEK-1 inhibitor PD98059 completely abolish the activin-mediated cell growth inhibition of T47D cells. Together, these results define a new role for activin in human breast cancer T47D cells and highlight a new pathway utilized by this growth factor in the mediation of its biological effects in cell growth arrest. PMID- 11278745 TI - P-glycoprotein does not protect cells against cytolysis induced by pore-forming proteins. AB - P-glycoprotein (P-gp) is an ATP-dependent drug pump that confers multidrug resistance (MDR). In addition to its ability to efflux toxins, P-gp can also inhibit apoptosis induced by a wide array of cell death stimuli that rely on activation of intracellular caspases for full function. We therefore hypothesized that P-gp may have additional functions in addition to its role in effluxing xenotoxins that could provide protection to tumor cells against a host response. There have been a number of contradictory reports concerning the role of P-gp in regulating complement activation. Given the disparate results obtained by different laboratories and our published results demonstrating that P-gp does not affect cell death induced by another membranolytic protein, perforin, we decided to assess the role of P-gp in regulating cell lysis induced by a number of different pore-forming proteins. Testing a variety of different P-gp-expressing MDR cell lines produced following exposure of cells to chemotherapeutic agents or by retroviral gene transduction in the complete absence of any drug selection, we found no difference in sensitivity of P-gp(+ve) or P-gp(-ve) cells to the pore forming proteins complement, perforin, or pneumolysin. Based on these results, we conclude that P-gp does not affect cell lysis induced by pore-forming proteins. PMID- 11278746 TI - Lefty inhibits receptor-regulated Smad phosphorylation induced by the activated transforming growth factor-beta receptor. AB - Transforming growth factor-beta (TGF-beta) is a pleiotropic cytokine that regulates growth and differentiation of diverse types of cells. TGF-beta actions are directed by ligand-induced activation of TGF-beta receptors with intrinsic serine/threonine kinase activity that trigger phosphorylation of receptor regulated Smad (R-Smad) protein. Phosphorylated R-Smad proteins bind to Smad4, and the complexes formed move into the nucleus, where they act as components of a transcriptional complex. Here, we show that TGF-beta signaling is inhibited by lefty, a novel member of the TGF-beta superfamily. Lefty perturbed TGF-beta signaling by inhibiting the phosphorylation of Smad2 following activation of the TGF-beta receptor. Moreover, lefty inhibited the events that lie downstream from R-Smad phosphorylation, including heterodimerization of R-Smad proteins with Smad4 and nuclear translocation of the R-Smad.Smad4 complex. Lefty repressed TGF beta-induced expression of reporter genes for the p21, cdc25, and connective tissue growth factor promoters and of a reporter gene driven by the Smad-binding element. Similarly, lefty inhibited both BMP-mediated Smad5 phosphorylation and gene transcription. The action of lefty does not appear to depend on protein synthesis, including synthesis of inhibitory Smad proteins. Thus, lefty provides a repressed state of TGF-beta- or BMP-responsive genes and participates in negative modulation of TGF-beta and BMP signaling by inhibition of phosphorylation of R-Smad proteins. PMID- 11278747 TI - Overexpression of cyclooxygenase-2 is sufficient to induce tumorigenesis in transgenic mice. AB - The cyclooxygenase (COX)-2 gene encodes an inducible prostaglandin synthase enzyme that is overexpressed in adenocarcinomas and other tumors. Deletion of the murine Cox-2 gene in Min mice reduced the incidence of intestinal tumors, suggesting that it is required for tumorigenesis. However, it is not known if overexpression of Cox-2 is sufficient to induce tumorigenic transformation. We have derived transgenic mice that overexpress the human COX-2 gene in the mammary glands using the murine mammary tumor virus promoter. The human Cox-2 mRNA and protein are expressed in mammary glands of female transgenic mice and were strongly induced during pregnancy and lactation. Female virgin Cox-2 transgenic mice showed precocious lobuloalveolar differentiation and enhanced expression of the beta-casein gene, which was inhibited by the Cox inhibitor indomethacin. Mammary gland involution was delayed in Cox-2 transgenic mice with a decrease in apoptotic index of mammary epithelial cells. Multiparous but not virgin females exhibited a greatly exaggerated incidence of focal mammary gland hyperplasia, dysplasia, and transformation into metastatic tumors. Cox-2-induced tumor tissue expressed reduced levels of the proapoptotic proteins Bax and Bcl-x(L) and an increase in the anti-apoptotic protein Bcl-2, suggesting that decreased apoptosis of mammary epithelial cells contributes to tumorigenesis. These data indicate that enhanced Cox-2 expression is sufficient to induce mammary gland tumorigenesis. Therefore, inhibition of Cox-2 may represent a mechanism-based chemopreventive approach for carcinogenesis. PMID- 11278748 TI - Yeast V-ATPase complexes containing different isoforms of the 100-kDa a-subunit differ in coupling efficiency and in vivo dissociation. AB - The 100 kDa a-subunit of the yeast vacuolar (H(+))-ATPase (V-ATPase) is encoded by two genes, VPH1 and STV1. These genes encode unique isoforms of the a-subunit that have previously been shown to reside in different intracellular compartments in yeast. Vph1p localizes to the central vacuole, whereas Stv1p is present in some other compartment, possibly the Golgi or endosomes. To compare the properties of V-ATPases containing Vph1p or Stv1p, Stv1p was expressed at higher than normal levels in a strain disrupted in both genes, under which conditions V ATPase complexes containing Stv1p appear in the vacuole. Complexes containing Stv1p showed lower assembly with the peripheral V(1) domain than did complexes containing Vph1p. When corrected for this lower degree of assembly, however, V ATPase complexes containing Vph1p and Stv1p had similar kinetic properties. Both exhibited a K(m) for ATP of about 250 microm, and both showed resistance to sodium azide and vanadate and sensitivity to nanomolar concentrations of concanamycin A. Stv1p-containing complexes, however, showed a 4-5-fold lower ratio of proton transport to ATP hydrolysis than Vph1p-containing complexes. We also compared the ability of V-ATPase complexes containing Vph1p or Stv1p to undergo in vivo dissociation in response to glucose depletion. Vph1p-containing complexes present in the vacuole showed dissociation in response to glucose depletion, whereas Stv1p-containing complexes present in their normal intracellular location (Golgi/endosomes) did not. Upon overexpression of Stv1p, Stv1p-containing complexes present in the vacuole showed glucose-dependent dissociation. Blocking delivery of Vph1p-containing complexes to the vacuole in vps21Delta and vps27Delta strains caused partial inhibition of glucose-dependent dissociation. These results suggest that dissociation of the V-ATPase complex in vivo is controlled both by the cellular environment and by the 100-kDa a-subunit isoform present in the complex. PMID- 11278749 TI - Rab8b and its interacting partner TRIP8b are involved in regulated secretion in AtT20 cells. AB - Rab proteins are a family of small GTPases that regulate intracellular vesicle traffic. Rab8b, because of its homology with Rab8, has been suggested to function in vesicle transport to the plasma membrane. Using the yeast two-hybrid system, we identified a Rab8b interacting clone, termed TRIP8b, from a rat brain cDNA library. The gene encodes a 66-kDa protein with homology to the peroxisomal targeting signal 1 receptor. The interaction between Rab8b and TRIP8b was further verified by in vitro binding assays and co-immunoprecipitation studies. Additional experiments with Rab8b mutants demonstrated that Rab8b requires a guanine nucleotide but not prenylation for its interaction with TRIP8b. Western immunoblot analysis showed that TRIP8b was primarily expressed in brain. Subcellular fractionation of AtT20 cells revealed that TRIP8b was present in both cytosolic and membrane fractions. To investigate the function of Rab8b and TRIP8b in secretion, we examined the release of ACTH from AtT20 cells. Results from stable cell lines expressing Rab8b or TRIP8b indicated that both proteins had a stimulatory effect on cAMP-induced secretion of ACTH. In summary, these data suggest that Rab8b and TRIP8b interact with each other and are involved in the regulated secretory pathway in AtT20 cells. PMID- 11278750 TI - Cloning and expression of a novel nuclear matrix-associated protein that is regulated during the retinoic acid-induced neuronal differentiation. AB - Retinoic acid (RA), a derivative of vitamin A, is essential for the normal patterning and neurogenesis during development. RA treatment induces growth arrest and terminal differentiation of a human embryonal carcinoma cell line (NT2) into postmitotic central nervous system neurons. Using RNA fingerprinting by arbitrarily primed polymerase chain reaction, we identified a novel serine/threonine-rich protein, RA-regulated nuclear matrix-associated protein (Ramp), that was down-regulated during the RA-induced differentiation of NT2 cells. Prominent mRNA expression of ramp could be detected in adult placenta and testis as well as in all human fetal tissues examined. The genomic clone of ramp has been mapped to the telomere of chromosome arm 1q, corresponding to band 1q32.1-32.2. Associated with the nuclear matrix of NT2 cells, Ramp translocates from the interphase nucleus to the metaphase cytoplasm during mitosis. During the late stage of cytokinesis, Ramp concentrates at the midzone of the dividing daughter cells. The transcript expression of ramp is closely correlated with the cell proliferation rate of NT2 cells. Moreover, overexpression of Ramp induces a transient increase in the proliferation rate of NT2 cells. Taken together, our data suggest that Ramp plays a role in the proliferation of the human embryonal carcinoma cells. PMID- 11278751 TI - Chimeras of X+, K+-ATPases. The M1-M6 region of Na+, K+-ATPase is required for Na+-activated ATPase activity, whereas the M7-M10 region of H+, K+-ATPase is involved in K+ de-occlusion. AB - In this study we reveal regions of Na(+),K(+)-ATPase and H(+),K(+)-ATPase that are involved in cation selectivity. A chimeric enzyme in which transmembrane hairpin M5-M6 of H(+),K(+)-ATPase was replaced by that of Na(+),K(+)-ATPase was phosphorylated in the absence of Na(+) and showed no K(+)-dependent reactions. Next, the part originating from Na(+),K(+)-ATPase was gradually increased in the N-terminal direction. We demonstrate that chimera HN16, containing the transmembrane segments one to six and intermediate loops of Na(+),K(+)-ATPase, harbors the amino acids responsible for Na(+) specificity. Compared with Na(+),K(+)-ATPase, this chimera displayed a similar apparent Na(+) affinity, a lower apparent K(+) affinity, a higher apparent ATP affinity, and a lower apparent vanadate affinity in the ATPase reaction. This indicates that the E(2)K form of this chimera is less stable than that of Na(+),K(+)-ATPase, suggesting that it, like H(+),K(+)-ATPase, de-occludes K(+) ions very rapidly. Comparison of the structures of these chimeras with those of the parent enzymes suggests that the C-terminal 187 amino acids and the beta-subunit are involved in K(+) occlusion. Accordingly, chimera HN16 is not only a chimeric enzyme in structure, but also in function. On one hand it possesses the Na(+)-stimulated ATPase reaction of Na(+),K(+)-ATPase, while on the other hand it has the K(+) occlusion properties of H(+),K(+)-ATPase. PMID- 11278752 TI - Murine equivalent of the human histo-blood group ABO gene is a cis-AB gene and encodes a glycosyltransferase with both A and B transferase activity. AB - We have cloned murine genomic and complementary DNA that is equivalent to the human ABO gene. The murine gene consists of at least six coding exons and spans at least 11 kilobase pairs. Exon-intron boundaries are similar to those of the human gene. Unlike human A and B genes that encode two distinct glycosyltransferases with different donor nucleotide-sugar specificities, the murine gene is a cis-AB gene that encodes an enzyme with both A and B transferase activities, and this cis-AB gene prevails in the mouse population. Cloning of the murine AB gene may be helpful in establishing a mouse model system to assess the functionality of the ABO genes in the future. PMID- 11278753 TI - Evidence that the peptidylprolyl isomerase domain of the hsp90-binding immunophilin FKBP52 is involved in both dynein interaction and glucocorticoid receptor movement to the nucleus. AB - We have previously shown that immunoadsorption of the FKBP52 immunophilin component of steroid receptor.hsp90 heterocomplexes is accompanied by coadsorption of cytoplasmic dynein, a motor protein involved in retrograde transport of vesicles toward the nucleus. Coimmunoadsorption of dynein is competed by an expressed fragment of FKBP52 comprising its peptidylprolyl isomerase (PPIase) domain (Silverstein, A. M., Galigniana, M. D., Kanelakis, K. C., Radanyi, C., Renoir, J.-M., and Pratt, W. B. (1999) J. Biol. Chem. 52, 36980 36986). Here we show that cotransfection of 3T3 cells with the FKBP52 PPIase domain and a green fluorescent protein (GFP) glucocorticoid receptor (GR) chimera inhibits dexamethasone-dependent movement of the GFP-GR from the cytoplasm to the nucleus. Cotransfection with FKBP12 does not affect GFP-GR movement. Inhibition of movement by the FKBP52 PPIase domain is abrogated in cells treated with colcemid to eliminate microtubules prior to steroid addition. After withdrawal of colcemid, microtubules reform, and PPIase inhibition of GFP-GR movement is restored. These observations are consistent with the notion that FKBP52 targets retrograde movement of the GFP-GR along microtubules by linking the receptor to the dynein motor. Here, we also show that native GR.hsp90 heterocomplexes immunoadsorbed from L cell cytosol contain dynein and that GR.hsp90 heterocomplexes assembled in reticulocyte lysate contain cytoplasmic dynein in a manner that is competed by the PPIase domain of FKBP52. PMID- 11278754 TI - Direct binding of the signal-transducing adaptor Grb2 facilitates down-regulation of the cyclin-dependent kinase inhibitor p27Kip1. AB - Ectopic expression of Jab1/CSN5 induces specific down-regulation of the cyclin dependent kinase (Cdk) inhibitor p27 (p27(Kip1)) in a manner dependent upon transportation from the nucleus to the cytoplasm. Here we show that Grb2 and Grb3 3, the molecules functioning as an adaptor in the signal transduction pathway, specifically and directly bind to p27 in the cytoplasm and participate in the regulation of p27. The interaction requires the C-terminal SH3-domain of Grb2/3-3 and the proline-rich sequence contained in p27 immediately downstream of the Cdk binding domain. In living cells, enforcement of the cytoplasmic localization of p27, either by artificial manipulation of the nuclear/cytoplasmic transport signal sequence or by coexpression of ectopic Jab1/CSN5, markedly enhances the stable interaction between p27 and Grb2. Overexpression of Grb2 accelerates Jab1/CSN5-mediated degradation of p27, while Grb3-3 expression suppresses it. A p27 mutant unable to bind to Grb2 is transported into the cytoplasm in cells ectopically expressing Jab1/CSN5 but is refractory to the subsequent degradation. These findings indicate that Grb2 participates in a negative regulation of p27 and may directly link the signal transduction pathway with the cell cycle regulatory machinery. PMID- 11278755 TI - The role of the M6-M7 loop (L67) in stabilization of the phosphorylation and Ca(2+) binding domains of the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA). AB - The amino acid sequence (L67) intervening between the M6 and M7 transmembrane segments of the Ca(2+) transport ATPase was subjected to mutational analysis. Mutation of Pro(820) to Ala interferes with protein expression even though transcription occurs at normal levels. Single mutations of Lys(819) or Arg(822) to Ala, Phe, or Glu allow good expression, but produce strong inhibition of ATPase activity. The main defect produced by these mutations is strong interference with enzyme phosphorylation by ATP in the presence of Ca(2+), and also by P(i) in the absence of Ca(2+). The Lys(819) and Arg(822) mutants undergo slight and moderate reduction of Ca(2+) binding affinity, respectively. Reduction of overall steady state ATPase velocity is then due to inhibition of phosphorylated intermediate formation. On the other hand, a cluster of conservative mutations of Asp(813), Asp(815), and Asp(818) to Asn interferes strongly with enzyme activation by Ca(2+) binding and formation of phosphorylated enzyme intermediate by utilization of ATP. Enzyme phosphorylation by Pi in the absence of Ca(2+) undergoes slight or no inhibition by the triple aspartate mutation. Therefore, the triple mutation interferes mainly with the calcium dependent activation of the ATPase. The effect of the triple mutation can be to a large extent reproduced by single mutation of Asp(813) (but not of Asp(815) or Asp(818)) to Asn. Functional and structural analysis of the experimental data demonstrates that the L67 loop plays an important role in protein folding and function. This role is sustained by linking the cytosolic catalytic domain and the transmembrane Ca(2+) binding domain through a network of hydrogen bonds. PMID- 11278757 TI - The PTPmu protein-tyrosine phosphatase binds and recruits the scaffolding protein RACK1 to cell-cell contacts. AB - PTPmu, an Ig superfamily receptor protein-tyrosine phosphatase, promotes cell cell adhesion and interacts with the cadherin-catenin complex. The signaling pathway downstream of PTPmu is unknown; therefore, we used a yeast two-hybrid screen to identify additional PTPmu interacting proteins. The membrane-proximal catalytic domain of PTPmu was used as bait. Sequencing of two positive clones identified the scaffolding protein RACK1 (receptor for activated protein C kinase) as a PTPmu interacting protein. We demonstrate that RACK1 interacts with PTPmu when co-expressed in a recombinant baculovirus expression system. RACK1 is known to bind to the src protein-tyrosine kinase. This study demonstrates that PTPmu association with RACK1 is disrupted by the presence of constituitively active src. RACK1 is thought to be a scaffolding protein that recruits proteins to the plasma membrane via an unknown mechanism. We have shown that the association of endogenous PTPmu and RACK1 in a lung cell line is increased at high cell density. We also demonstrate that the recruitment of RACK1 to both the plasma membrane and cell-cell contact sites is dependent upon the presence of the PTP mu protein in these cells. Therefore, PTPmu may be one of the proteins that recruits RACK1 to points of cell-cell contact, which may be important for PTPmu dependent signaling in response to cell-cell adhesion. PMID- 11278756 TI - Ski-interacting protein interacts with Smad proteins to augment transforming growth factor-beta-dependent transcription. AB - Transforming growth factor-beta (TGF-beta) signaling requires the action of Smad proteins in association with other DNA-binding factors and coactivator and corepressor proteins to modulate target gene transcription. Smad2 and Smad3 both associate with the c-Ski and Sno oncoproteins to repress transcription of Smad target genes via recruitment of a nuclear corepressor complex. Ski-interacting protein (SKIP), a nuclear hormone receptor coactivator, was examined as a possible modulator of transcriptional regulation of the TGF-beta-responsive promoter from the plasminogen activator inhibitor gene-1. SKIP augmented TGF-beta dependent transactivation in contrast to Ski/Sno-dependent repression of this reporter. SKIP interacted with Smad2 and Smad3 proteins in vivo in yeast and in mammalian cells through a region of SKIP between amino acids 201-333. In vitro, deletion of the Mad homology domain 2 (MH2) domain of Smad3 abrogated SKIP binding, like Ski/Sno, but the MH2 domain of Smad3 alone was not sufficient for protein-protein interaction. Overexpression of SKIP partially overcame Ski/Sno dependent repression, whereas Ski/Sno overexpression attenuated SKIP augmentation of TGF-beta-dependent transcription. Our results suggest a potential mechanism for transcriptional control of TGF-beta signaling that involves the opposing and competitive actions of SKIP and Smad MH2-interacting factors, such as Ski and/or Sno. Thus, SKIP appears to modulate both TGF-beta and nuclear hormone receptor signaling pathways. PMID- 11278758 TI - A role for the Ppz Ser/Thr protein phosphatases in the regulation of translation elongation factor 1Balpha. AB - In vivo 32P-labeled yeast proteins from wild type and ppz1 ppz2 phosphatase mutants were resolved by bidimensional electrophoresis. A prominent phosphoprotein, which in ppz mutants showed a marked shift to acidic regions, was identified by mixed peptide sequencing as the translation elongation factor 1Balpha (formerly eEF1beta). An equivalent shift was detected in cells overexpressing HAL3, a inhibitory regulatory subunit of Ppz1. Subsequent analysis identified the conserved Ser-86 as the in vivo phosphorylatable residue and showed that its phosphorylation was increased in ppz cells. Pull-down experiments using a glutathione S-transferase (GST)-EF1Balpha fusion version allowed to identify Ppz1 as an in vivo interacting protein. Cells lacking Ppz display a higher tolerance to known translation inhibitors, such as hygromycin and paromomycin, and enhanced readthrough at all three nonsense codons, suggesting that translational fidelity might be affected. Overexpression of a GST-EF1Balpha fusion counteracted the growth defect associated to high levels of Ppz1 and this effect was essentially lost when the phosphorylatable Ser-86 is replaced by Ala. Therefore, the Ppz phosphatases appear to regulate the phosphorylation state of EF1Balpha in yeast, and this may result in modification of the translational accuracy. PMID- 11278759 TI - The sonic hedgehog receptor patched associates with caveolin-1 in cholesterol rich microdomains of the plasma membrane. AB - The Hedgehog signaling pathway is involved in early embryonic patterning as well as in cancer; however, little is known about the subcellular localization of the Hedgehog receptor complex of Patched and Smoothened. Since Hh has been found in lipid rafts in Drosophila, we hypothesized that Patched and Smoothened might also be found in these cholesterol-rich microdomains. In this study, we demonstrate that both Smoothened and Patched are in caveolin-1-enriched/raft microdomains. Immunoprecipitation studies show that Patched specifically interacts with caveolin-1, whereas Smoothened does not. Fractionation studies show that Patched and caveolin-1 can be co-isolated from buoyant density fractions that represent caveolae/raft microdomains and that Patched and caveolin-1 co-localize by confocal microscopy. Glutathione S-transferase fusion protein experiments show that the interaction between Patched and caveolin-1 involves the caveolin-1 scaffolding domain and a Patched consensus binding site. Immunocytochemistry data and fractionation studies also show that Patched seems to be required for transport of Smoothened to the membrane. Depletion of plasmalemmal cholesterol influences the distribution of the Hh receptor complex in the caveolin enriched/raft microdomains. These data suggest that caveolin-1 may be integral for sequestering the Hh receptor complex in these caveolin-enriched microdomains, which act as a scaffold for the interactions with the Hh protein. PMID- 11278761 TI - Functional role and immunocytochemical localization of the gamma a and gamma b forms of the Na,K-ATPase gamma subunit. AB - The gamma subunit of the Na,K-ATPase is a member of the FXYD family of type 2 transmembrane proteins that probably function as regulators of ion transport. Rat gamma is present primarily in the kidney as two main splice variants, gamma(a) and gamma(b), which differ only at their extracellular N termini (TELSANH and MDRWYL, respectively; Kuster, B., Shainskaya, A., Pu, H. X., Goldshleger, R., Blostein, R., Mann, M., and Karlish, S. J. D. (2000) J. Biol. Chem. 275, 18441 18446). Expression in cultured cells indicates that both variants affect catalytic properties, without a detectable difference between gamma(a) and gamma(b). At least two singular effects are seen, irrespective of whether the variants are expressed in HeLa or rat alpha1-transfected HeLa cells, i.e. (i) an increase in apparent affinity for ATP, probably secondary to a left shift in E(1) <--> E(2) conformational equilibrium and (ii) an increase in K(+) antagonism of cytoplasmic Na(+) activation. Antibodies against the C terminus common to both variants (anti-gamma) abrogate the first effect but not the second. In contrast, gamma(a) and gamma(b) show differences in their localization along the kidney tubule. Using anti-gamma (C-terminal) and antibodies to the rat alpha subunit as well as antibodies to identify cell types, double immunofluorescence showed gamma in the basolateral membrane of several tubular segments. Highest expression is in the medullary portion of the thick ascending limb (TAL), which contains both gamma(a) and gamma(b). In fact, TAL is the only positive tubular segment in the medulla. In the cortex, most tubules express gamma but at lower levels. Antibodies specific for gamma(a) and gamma(b) showed differences in their cortical location; gamma(a) is specific for cells in the macula densa and principal cells of the cortical collecting duct but not cortical TAL. In contrast, gamma(b) but not gamma(a) is present in the cortical TAL only. Thus, the importance of gamma(a) and gamma(b) may be related to their partially overlapping but distinct expression patterns and tissue-specific functions of the pump that these serve. PMID- 11278760 TI - Bradykinin B2 receptors activate Na+/H+ exchange in mIMCD-3 cells via Janus kinase 2 and Ca2+/calmodulin. AB - We used a cultured murine cell model of the inner medullary collecting duct (mIMCD-3 cells) to examine the regulation of the ubiquitous sodium-proton exchanger, Na+/H+ exchanger isoform 1 (NHE-1), by a prototypical G protein coupled receptor, the bradykinin B2 receptor. Bradykinin rapidly activates NHE-1 in a concentration-dependent manner as assessed by proton microphysiometry of quiescent cells and by 2'-7'-bis[2-carboxymethyl]-5(6)-carboxyfluorescein fluorescence measuring the accelerated rate of pH(i) recovery from an imposed acid load. The activation of NHE-1 is blocked by inhibitors of the bradykinin B2 receptor, phospholipase C, Ca2+/calmodulin (CaM), and Janus kinase 2 (Jak2), but not by pertussis toxin or by inhibitors of protein kinase C and phosphatidylinositol 3'-kinase. Immunoprecipitation studies showed that bradykinin stimulates the assembly of a signal transduction complex that includes CaM, Jak2, and NHE-1. CaM appears to be a direct substrate for phosphorylation by Jak2 as measured by an in vitro kinase assay. We propose that Jak2 is a new indirect regulator of NHE-1 activity, which modulates the activity of NHE-1 by increasing the tyrosine phosphorylation of CaM and most likely by increasing the binding of CaM to NHE-1. PMID- 11278762 TI - Syntaxin 7 complexes with mouse Vps10p tail interactor 1b, syntaxin 6, vesicle associated membrane protein (VAMP)8, and VAMP7 in b16 melanoma cells. AB - Syntaxin 7 is a mammalian target soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) involved in membrane transport between late endosomes and lysosomes. The aim of the present study was to use immunoaffinity techniques to identify proteins that interact with Syntaxin 7. We reasoned that this would be facilitated by the use of cells producing high levels of Syntaxin 7. Screening of a large number of tissues and cell lines revealed that Syntaxin 7 is expressed at very high levels in B16 melanoma cells. Moreover, the expression of Syntaxin 7 increased in these cells as they underwent melanogenesis. From a large scale Syntaxin 7 immunoprecipitation, we have identified six polypeptides using a combination of electrospray mass spectrometry and immunoblotting. These polypeptides corresponded to Syntaxin 7, Syntaxin 6, mouse Vps10p tail interactor 1b (mVti1b), alpha-synaptosome-associated protein (SNAP), vesicle-associated membrane protein (VAMP)8, VAMP7, and the protein phosphatase 1M regulatory subunit. We also observed partial colocalization between Syntaxin 6 and Syntaxin 7, between Syntaxin 6 and mVti1b, but not between Syntaxin 6 and the early endosomal t-SNARE Syntaxin 13. Based on these and data reported previously, we propose that Syntaxin 7/mVti1b/Syntaxin 6 may form discrete SNARE complexes with either VAMP7 or VAMP8 to regulate fusion events within the late endosomal pathway and that these events may play a critical role in melanogenesis. PMID- 11278763 TI - Structure-function analysis of Bag1 proteins. Effects on androgen receptor transcriptional activity. AB - Bag1 is a regulator of heat shock protein 70 kDa (Hsp70/Hsc70) family proteins that interacts with steroid hormone receptors. Four isoforms of Bag1 have been recognized: Bag1, Bag1S, Bag1M (RAP46/HAP46), and Bag1L. Although Bag1L, Bag1M, and Bag1 can bind the androgen receptor (AR) in vitro, only Bag1L enhanced AR transcriptional activity. Bag1L was determined to be a nuclear protein by immunofluorescence microscopy, whereas Bag1, Bag1S, and Bag1M were predominantly cytoplasmic. Forced nuclear targeting of Bag1M, but not Bag1 or Bag1S, resulted in potent AR coactivation, indicating that Bag1M possesses the necessary structural features provided it is expressed within the nucleus. The ability of Bag1L to enhance AR activity was reduced with the removal of an NH(2)-terminal domain of Bag1L, which was found to be required for efficient nuclear localization and/or retention. In contrast, deletion of a conserved ubiquitin like domain from Bag1L did not interfere with its nuclear targeting or AR regulatory activity. Thus, both the unique NH(2)-terminal domain and the COOH terminal Hsc70-binding domain of Bag1L are simultaneously required for its function as an AR regulator, whereas the conserved ubiquitin-like domain is expendable. PMID- 11278764 TI - Glucocorticoid receptor-mediated protection from apoptosis is associated with induction of the serine/threonine survival kinase gene, sgk-1. AB - We previously demonstrated that activation of the glucocorticoid receptor (GR) initiates an antiapoptotic signal in the immortalized human mammary epithelial cell line MCF10A that is dependent on the GR's transcriptional activity. In this study, we show that the survival role of GR activation extends to protecting human breast cancer cells undergoing apoptosis after growth factor deprivation. Serum and glucocorticoid-regulated kinase-1 (sgk), a gene previously identified as a direct transcriptional target of the activated GR in a rat mammary tumor cell line, was rapidly induced after GR activation in human mammary epithelial cells. Furthermore, in the absence of all growth factors, ectopic sgk expression inhibited apoptosis, suggesting that SGK is a survival kinase. Finally, kinase dead SGK expression inhibited the protection from apoptosis usually seen after GR activation. These findings suggest that SGK is an important downstream target of GR-mediated survival signaling and that it is distinct from other survival kinases because it can be primarily regulated at the level of transcription. PMID- 11278765 TI - Transcriptional regulation of the TFIIH transcription repair components XPB and XPD by the hepatitis B virus x protein in liver cells and transgenic liver tissue. AB - Human hepatitis B virus is a risk factor for the development of hepatocellular carcinoma. The hepatitis B virus x protein (HBx) has been shown to inactivate the p53 tumor suppressor protein and impair DNA repair, cell cycle, and apoptosis mechanisms. Herein we report that HBx represses two components of the transcription-repair factor TFIIH, XPB (p89), and XPD (p80), both in p53 proficient and p53-deficient liver cells. This inhibition is observed while HBx maintains its transactivation function. Expression of HBx in liver cells results in down-regulation of endogenous XPB and XPD mRNAs and proteins; this inhibition is not observed with other TFIIH subunits, XPA or PCNA. In liver tissue from HBx transgenics, XPB and XPD proteins are down-regulated in comparison to matched normal liver tissue. HBx has been shown to interact with Sp1 transcription factor and affects its DNA binding activity. Sp1 is essential for the basal promoter activity of XPB in liver cells and Drosophila SL2 cells. In the Sp1-deficient SL2 cells, HBx-induced XPB and XPD inhibition is Sp1-dependent. In summary, our results provide evidence that HBx represses the expression of key TFIIH proteins at least in part through Sp1 elements; this repression may impair TFIIH function in DNA repair mechanisms. PMID- 11278766 TI - A novel quaternary structure of the dimeric alpha-crystallin domain with chaperone-like activity. AB - alphaB-crystallin, a member of the small heat-shock protein family and a major eye lens protein, is a high molecular mass assembly and can act as a molecular chaperone. We report a synchrotron radiation x-ray solution scattering study of a truncation mutant from the human alphaB-crystallin (alphaB57-157), a dimeric protein that comprises the alpha-crystallin domain of the alphaB-crystallin and retains a significant chaperone-like activity. According to the sequence analysis (more than 23% identity), the monomeric fold of the alpha-crystallin domain should be close to that of the small heat-shock protein from Methanococcus jannaschii (MjHSP16.5). The theoretical scattering pattern computed from the crystallographic model of the dimeric MjHSP16.5 deviates significantly from the experimental scattering by the alpha-crystallin domain, pointing to different quaternary structures of the two proteins. A rigid body modeling against the solution scattering data yields a model of the alpha-crystallin domain revealing a new dimerization interface. The latter consists of a strand-turn-strand motif contributed by each of the monomers, which form a four-stranded, antiparallel, intersubunit composite beta-sheet. This model agrees with the recent spin labeling results and suggests that the alphaB-crystallin is composed by flexible building units with an extended surface area. This flexibility may be important for biological activity and for the formation of alphaB-crystallin complexes of variable sizes and compositions. PMID- 11278767 TI - Protein compactness measured by fluorescence resonance energy transfer. Human carbonic anhydrase ii is considerably expanded by the interaction of GroEL. AB - Nine single-cysteine mutants were labeled with 5-(2 iodoacetylaminoethylamino)naphthalene-1-sulfonic acid, an efficient acceptor of Trp fluorescence in fluorescence resonance energy transfer. The ratio between the fluorescence intensity of the 5-(2-acetylaminoethylamino)naphthalene-1-sulfonic acid (AEDANS) moiety excited at 295 nm (Trp absorption) and 350 nm (direct AEDANS absorption) was used to estimate the average distances between the seven Trp residues in human carbonic anhydrase II (HCA II) and the AEDANS label. Guanidine HCl denaturation of the HCA II variants was also performed to obtain a curve that reflected the compactness of the protein at various stages of the unfolding, which could serve as a scale of the expansion of the protein. This approach was developed in this study and was used to estimate the compactness of HCA II during heat denaturation and interaction with GroEL. It was shown that thermally induced unfolding of HCA II proceeded only to the molten globule state. Reaching this state was sufficient to allow HCA II to bind to GroEL, and the volume of the molten globule intermediate increased approximately 2.2-fold compared with that of the native state. GroEL-bound HCA II expands to a volume three to four times that of the native state (to approximately 117,000 A(3)), which correlates well with a stretched and loosened-up HCA II molecule in an enlarged GroEL cavity. Recently, we found that HCA II binding causes such an inflation of the GroEL molecule, and this probably represents the mechanism by which GroEL actively stretches its protein substrates apart (Hammarstrom, P., Persson, M., Owenius, R., Lindgren, M., and Carlsson, U. (2000) J. Biol. Chem. 275, 22832-22838), thereby facilitating rearrangement of misfolded structure. PMID- 11278768 TI - Inhibition of mitogen-activated protein kinase kinase induces apoptosis of human chondrocytes. AB - We previously have reported that the mitogen-activated protein kinase (MAPK) pathway is stimulated by adhesion of human chondrocytes to anti-beta(1)-integrin antibodies or collagen type II in vitro. These mechanisms most likely prevent chondrocyte dedifferentiation to fibroblast-like cells and chondrocyte death. To investigate whether this pathway plays an essential role for the differentiation, phenotype, and survival of chondrocytes, we blocked mitogen-activated protein kinase/extracellular signal-regulated kinase (Erk) (MEK), a kinase upstream of the kinase Erk by using U0126. Exposure of chondrocytes to U0126 caused activation of caspase-3 in a dose-dependent manner. Western blot analysis with an antibody specific for dually phosphorylated Erk shows that collagen type II induced phosphorylation of Erk1/2 was specifically blocked by U0126 in a dose dependent manner. Immunohistochemical analysis showed that treated chondrocytes were caspase-3 positive. In treated chondrocytes, the cleavage of 116-kDa poly(ADP-ribose)polymerase resulted in the 85-kDa apoptosis-related cleavage fragment and was associated with caspase-3 activity. Analysis by electron microscopy showed typical morphological signs of apoptosis, such as crescent shaped clumps of heterochromatin, and a degraded pericellular matrix. Thus, these results indicate that the MEK/Erk signal transduction pathway is involved in the maintenance of chondrocytes differentiation and survival. These data stimulate further investigations on the role of mitogen-activated protein kinase pathways in human chondrocytes. PMID- 11278769 TI - Identification of 12 new yeast mitochondrial ribosomal proteins including 6 that have no prokaryotic homologues. AB - Mitochondrial ribosomal proteins were studied best in yeast, where the small subunit was shown to contain about 35 proteins. Yet, genetic and biochemical studies identified only 14 proteins, half of which were predictable by sequence homology with prokaryotic ribosomal components of the small subunit. Using a recently described affinity purification technique and tagged versions of yeast Ykl155c and Mrp1, we isolated this mitochondrial ribosomal subunit and identified a total of 20 proteins, of which 12 are new. For a subset of the newly described ribosomal proteins, we showed that they are localized in mitochondria and are required for the respiratory competency of the yeast cells. This brings to 26 the total number of proteins described as components of the mitochondrial small ribosomal subunit. Remarkably, almost half of the previously and newly identified mitochondrial ribosomal components showed no similarity to any known ribosomal protein. Homologues could be found, however, in predicted protein sequences from Schizosaccharomyces pombe. In more distant species, putative homologues were detected for Ykl155c, which shares conserved motifs with uncharacterized proteins of higher eukaryotes including humans. Another newly identified ribosomal protein, Ygl129c, was previously shown to be a member of the DAP-3 family of mitochondrial apoptosis mediators. PMID- 11278770 TI - Internalization of transthyretin. Evidence of a novel yet unidentified receptor associated protein (RAP)-sensitive receptor. AB - Transthyretin (TTR) is a plasma carrier of thyroxine and retinol-binding protein (RBP). Though the liver is the major site of TTR degradation, its cellular uptake is poorly understood. We explored TTR uptake using hepatomas and primary hepatocytes and showed internalization by a specific receptor. RBP complexed with TTR led to a 70% decrease of TTR internalization, whereas TTR bound to thyroxine led to a 20% increase. Different TTR mutants showed differences in uptake, suggesting receptor recognition dependent on the structure of TTR. Cross-linking studies using hepatomas and (125)I-TTR revealed a approximately 90-kDa complex corresponding to (125)I-TTR bound to its receptor. Given previous evidence that a fraction of TTR is associated with high-density lipoproteins (HDL) and that in the kidney, megalin, a member of the low-density lipoprotein receptor family (LDLr) internalizes TTR, we hypothesized that TTR and lipoproteins could share related degradation pathways. Using lipid-deficient serum in uptake assays, no significant changes were observed showing that TTR uptake is not lipoprotein dependent or due to TTR-lipoprotein complexes. However, competition studies showed that lipoproteins inhibit TTR internalization. The scavenger receptor SR BI, a HDL receptor, and known LDLr family hepatic receptors did not mediate TTR uptake as assessed using different cellular systems. Interestingly, the receptor associated protein (RAP), a ligand for all members of the LDLr, was able to inhibit TTR internalization. Moreover, the approximately 90-kDa TTR-receptor complex obtained by cross-linking was sensitive to the presence of RAP. To confirm that RAP sensitivity observed in hepatomas did not represent a mechanism absent in normal cells, primary hepatocytes were tested, and similar results were obtained. The RAP-sensitive TTR internalization together with displacement of TTR uptake by lipoproteins, further suggests that a common pathway might exist between TTR and lipoprotein metabolism and that an as yet unidentified RAP sensitive receptor mediates TTR uptake. PMID- 11278772 TI - Discrete targeting signals direct Pmp47 to oleate-induced peroxisomes in Saccharomyces cerevisiae. AB - Pmp47 is a peroxisomal membrane protein consisting of six transmembrane domains (TMDs). We previously showed that the second matrix loop containing a basic cluster of amino acids is important for peroxisomal targeting, and similar basic targeting motifs have been found in other peroxisomal membrane proteins. However, this basic cluster by itself targets to peroxisomes very poorly. We have developed a sensitive quantitative localization assay based on the targeting of Pmp47-GFP fusion proteins to identify the important elements of the basic cluster and to search for other targeting information on Pmp47. Our data suggest that side-chain structure and position as well as charge are important for targeting by the basic cluster. Analysis of other regions of Pmp47 indicates that all TMDs except TMD2 can be eliminated or substituted without significant loss of targeting. TMD2 plus an adjacent cytoplasmic-oriented sequence is crucial for targeting. Cytoplasmic-oriented sequences from two other peroxisomal membrane proteins, ScPex15p and ScPmp22, could partially substitute for the analogous sequence in Pmp47. Targeting with high fidelity to oleate-induced peroxisomes required the following elements: the cytoplasmic-oriented sequence and TMD2, a short matrix loop containing a basic cluster, and a membrane-anchoring TMD. PMID- 11278771 TI - Down-regulation of cholesterol 7alpha-hydroxylase (CYP7A1) gene expression by bile acids in primary rat hepatocytes is mediated by the c-Jun N-terminal kinase pathway. AB - Cholesterol 7alpha-hydroxylase (CYP7A1), the rate-limiting enzyme in the neutral pathway of bile acid biosynthesis, is feedback-inhibited at the transcriptional level by hydrophobic bile acids. Recent studies show that bile acids are physiological ligands for farnesoid X receptor (FXR). Activated FXR indirectly represses CYP7A1 transcription through induction of small heterodimer protein (SHP-1). In this study, we provide evidence that bile acids rapidly down-regulate CYP7A1 transcription via activation of the JNK/c-Jun pathway. Furthermore, we demonstrate that SHP-1 is also a direct target of activated c-Jun. In primary rat hepatocyte cultures, taurocholate (TCA) strongly activated JNK in a time- and concentration-dependent manner. Tumor necrosis factor-alpha, a potent activator of JNK, also rapidly activated JNK and down-regulated CYP7A1 mRNA levels. Overexpression of dominant-negative JNK1 or a transactivating domain mutant of c Jun significantly blocked the ability of TCA to down-regulate CYP7A1 mRNA. In contrast, overexpression of wild-type c-Jun (c-Jun(wt)) enhanced the repression of CYP7A1 by TCA. Moreover, overexpression of c-Jun(wt) resulted in increased SHP 1 promoter activity. Mutation of a putative AP-1 (c-Jun) element suppressed c-Jun mediated activation of the SHP-1 promoter construct. These results indicate that the bile acid-activated JNK pathway plays a pivotal role in regulating CYP7A1 levels in primary rat hepatocytes. PMID- 11278773 TI - Insulin and insulin-like growth factor I receptors utilize different G protein signaling components. AB - We examined the role of heterotrimeric G protein signaling components in insulin and insulin-like growth factor I (IGF-I) action. In HIRcB cells and in 3T3L1 adipocytes, treatment with the Galpha(i) inhibitor (pertussis toxin) or microinjection of the Gbetagamma inhibitor (glutathione S-transferase-betaARK) inhibited IGF-I and lysophosphatidic acid-stimulated mitogenesis but had no effect on epidermal growth factor (EGF) or insulin action. In basal state, Galpha(i) and Gbeta were associated with the IGF-I receptor (IGF-IR), and after ligand stimulation the association of IGF-IR with Galpha(i) increased concomitantly with a decrease in Gbeta association. No association of Galpha(i) was found with either the insulin or EGF receptor. Microinjection of anti-beta arrestin-1 antibody specifically inhibited IGF-I mitogenic action but had no effect on EGF or insulin action. beta-Arrestin-1 was associated with the receptors for IGF-I, insulin, and EGF in a ligand-dependent manner. We demonstrated that Galpha(i), betagamma subunits, and beta-arrestin-1 all play a critical role in IGF-I mitogenic signaling. In contrast, neither metabolic, such as GLUT4 translocation, nor mitogenic signaling by insulin is dependent on these protein components. These results suggest that insulin receptors and IGF-IRs can function as G protein-coupled receptors and engage different G protein partners for downstream signaling. PMID- 11278774 TI - Characterization of a novel KRAB/C2H2 zinc finger transcription factor involved in bone development. AB - Osteogenic differentiation involves a cascade of coordinated gene expression that regulates cell proliferation and matrix protein formation in a defined temporo spatial manner. Here we have used differential display to identify a novel zinc finger transcription factor (AJ18) that is induced during differentiation of bone cells in vitro and in vivo. The 64-kDa protein, encoded by a 7- kilobase mRNA, contains a Kruppel-associated box (KRAB) domain followed by 11 successive C(2)H(2) zinc finger motifs. AJ18 mRNA, which is also expressed in kidney and brain, is developmentally regulated in embryonic tibiae and calvariae, with little expression in neonate and adult animals. During osteogenic differentiation in vitro AJ18 mRNA is expressed as cells approach confluence and declines as bone formation occurs. Using bacterially expressed, His-tagged AJ18 in a target detection assay, we identified a consensus binding sequence of 5'-CCACA-3', which forms part of the consensus element for Runx2, a master gene for osteogenic differentiation. Overexpression of AJ18 suppressed Runx2-mediated transactivation of an osteocalcin promoter construct in transient transfection assays and reduced alkaline phosphatase activity in bone morphogenetic protein-induced C3H10T1/2 cells. These studies, therefore, have identified a novel zinc finger transcription factor in bone that can modulate Runx2 activity and osteogenic differentiation. PMID- 11278775 TI - The dynamics of the relay loop tryptophan residue in the Dictyostelium myosin motor domain and the origin of spectroscopic signals. AB - Steady-state and time-resolved fluorescence measurements were performed on a Dictyostelium discoideum myosin II motor domain construct retaining a single tryptophan residue at position 501, located on the relay loop. Other tryptophan residues were mutated to phenylalanine. The Trp-501 residue showed a large enhancement in fluorescence in the presence of ATP and a small quench in the presence of ADP as a result of perturbing both the ground and excited state processes. Fluorescence lifetime and quantum yield measurements indicated that at least three microstates of Trp-501 were present in all nucleotide states examined, and these could not be assigned to a particular gross conformation of the motor domain. Enhancement in emission intensity was associated with a reduction of the contribution from a statically quenched component and an increase in a component with a 5-ns lifetime, with little change in the contribution from a 1-ns lifetime component. Anisotropy measurements indicated that the Trp-501 side chain was relatively immobile in all nucleotide states, and the fluorescence was effectively depolarized by rotation of the whole motor domain with a correlation time on 50-70 ns. Overall these data suggest that the backbone of the relay loop remains structured throughout the myosin ATPase cycle but that the Trp-501 side chain experiences a different weighting in local environments provided by surrounding residues as the adjacent converter domain rolls around the relay loop. PMID- 11278776 TI - The myosin relay helix to converter interface remains intact throughout the actomyosin ATPase cycle. AB - Crystal structures of the myosin motor domain in the presence of different nucleotides show the lever arm domain in two basic angular states, postulated to represent prestroke and poststroke states, respectively (Rayment, I. (1996) J. Biol. Chem. 271, 15850-15853; Dominguez, R., Freyzon, Y., Trybus, K. M., and Cohen, C. (1998) Cell 94, 559-571). Contact is maintained between two domains, the relay and the converter, in both of these angular states. Therefore it has been proposed by Dominguez et al. (cited above) that this contact is critical for mechanically driving the angular change of the lever arm domain. However, structural information is lacking on whether this contact is maintained throughout the actin-activated myosin ATPase cycle. To test the functional importance of this interdomain contact, we introduced cysteines into the sequence of a "cysteine-light" myosin motor at position 499 on the lower cleft and position 738 on the converter domain (Shih, W. M., Gryczynski, Z., Lakowicz, J. L., and Spudich, J. A. (2000) Cell 102, 683-694). Disulfide cross-linking could be induced. The cross-link had minimal effects on actin binding, ATP-induced actin release, and actin-activated ATPase. These results demonstrate that the relay/converter interface remains intact in the actin strongly bound state of myosin and throughout the entire actin-activated myosin ATPase cycle. PMID- 11278777 TI - The Cdc42 target ACK2 directly interacts with clathrin and influences clathrin assembly. AB - The Ras-related GTP-binding protein Cdc42 has been implicated in a diversity of biological functions including the regulation of intracellular trafficking and endocytosis. While screening for Cdc42 targets that influence these activities, we identified the protein-tyrosine kinase ACK2 (for activated Cdc42-associated kinase 2) as a new binding partner for clathrin. ACK2 binds clathrin via a domain that is conserved among a number of other clathrin-binding proteins including the arrestins and AP-2. Overexpression of ACK2 in NIH3T3 cells results in an inhibition of transferrin receptor endocytosis because of a competition between ACK2 and AP-2 for clathrin. Activated Cdc42 weakens the interaction between ACK2 and clathrin and thus reverses the ACK2-mediated inhibition of endocytosis. Overexpression of ACK2 increases the amount of clathrin present in fractions enriched in clathrin-coated vesicles. Taken together, our data suggest that ACK2 may represent a novel clathrin-assembly protein and participate in the regulation of receptor-mediated endocytosis. PMID- 11278778 TI - The yeast ALG11 gene specifies addition of the terminal alpha 1,2-Man to the Man5GlcNAc2-PP-dolichol N-glycosylation intermediate formed on the cytosolic side of the endoplasmic reticulum. AB - The initial steps in N-linked glycosylation involve the synthesis of a lipid linked core oligosaccharide followed by the transfer of the core glycan to nascent polypeptides in the endoplasmic reticulum (ER). Here, we describe alg11, a new yeast glycosylation mutant that is defective in the last step of the synthesis of the Man(5)GlcNAc(2)-PP-dolichol core oligosaccharide on the cytosolic face of the ER. A deletion of the ALG11 gene leads to poor growth and temperature-sensitive lethality. In an alg11 lesion, both Man(3)GlcNAc(2)-PP dolichol and Man(4)GlcNAc(2)-PP-dolichol are translocated into the ER lumen as substrates for the Man-P-dolichol-dependent sugar transferases in this compartment. This leads to a unique family of oligosaccharide structures lacking one or both of the lower arm alpha1,2-linked Man residues. The former are elongated to mannan, whereas the latter are poor substrates for outerchain initiation by Ochlp (Nakayama, K.-I., Nakanishi-Shindo, Y., Tanaka, A., Haga Toda, Y., and Jigami, Y. (1997) FEBS Lett. 412, 547-550) and accumulate largely as truncated biosynthetic end products. The ALG11 gene is predicted to encode a 63.1-kDa membrane protein that by indirect immunofluorescence resides in the ER. The Alg11 protein is highly conserved, with homologs in fission yeast, worms, flies, and plants. In addition to these Alg11-related proteins, Alg11p is also similar to Alg2p, a protein that regulates the addition of the third mannose to the core oligosaccharide. All of these Alg11-related proteins share a 23-amino acid sequence that is found in over 60 proteins from bacteria to man whose function is in sugar metabolism, implicating this sequence as a potential sugar nucleotide binding motif. PMID- 11278779 TI - Glucose transporter asymmetries in the bovine blood-brain barrier. AB - The transport of glucose across the mammalian blood-brain barrier is mediated by the GLUT1 glucose transporter, which is concentrated in the endothelial cells of the cerebral microvessels. Several studies supported an asymmetric distribution of GLUT1 protein between the luminal and abluminal membranes (1:4) with a significant proportion of intracellular transporters. In this study we investigated the activity and concentration of GLUT1 in isolated luminal and abluminal membrane fractions of bovine brain endothelial cells. Glucose transport activity and glucose transporter concentration, as determined by cytochalasin B binding, were 2-fold greater in the luminal than in the abluminal membranes. In contrast, Western blot analysis using a rabbit polyclonal antibody raised against the C-terminal 20 amino acids of GLUT1 indicated a 1:5 luminal:abluminal distribution. Western blot analysis with antibodies raised against either the intracellular loop of GLUT1 or the purified erythrocyte protein exhibited luminal:abluminal ratios of 1:1. A similar ratio was observed when the luminal and abluminal fractions were exposed to the 2-N-4[(3)H](1-azi-2,2,2, trifluoroethyl)benzoxyl-1,3-bis-(d-mannos-4-yloxyl)-2-propylamine ([(3)H]ATB BMPA) photoaffinity label. These observations suggest that either an additional glucose transporter isoform is present in the luminal membrane of the bovine blood-brain barrier or the C-terminal epitope of GLUT1 is "masked" in the luminal membrane but not in the abluminal membranes. PMID- 11278780 TI - Cell adhesion and migration properties of beta 2-integrin negative polymorphonuclear granulocytes on defined extracellular matrix molecules. Relevance for leukocyte extravasation. AB - Regulated adhesion of leukocytes to the extracellular matrix is essential for transmigration of blood vessels and subsequent migration into the stroma of inflamed tissues. Although beta(2)-integrins play an indisputable role in adhesion of polymorphonuclear granulocytes (PMN) to endothelium, we show here that beta(1)- and beta(3)-integrins but not beta(2)-integrin are essential for the adhesion to and migration on extracellular matrix molecules of the endothelial cell basement membrane and subjacent interstitial matrix. Mouse wild type and beta(2)-integrin null PMN and the progranulocytic cell line 32DC13 were employed in in vitro adhesion and migration assays using extracellular matrix molecules expressed at sites of extravasation in vivo, in particular the endothelial cell laminins 8 and 10. Wild type and beta(2)-integrin null PMN showed the same pattern of ECM binding, indicating that beta(2)-integrins do not mediate specific adhesion of PMN to the extracellular matrix molecules tested; binding was observed to the interstitial matrix molecules, fibronectin and vitronectin, via integrins alpha(5)beta(1) and alpha(v)beta(3), respectively; to laminin 10 via alpha(6)beta(1); but not to laminins 1, 2, and 8, collagen type I and IV, perlecan, or tenascin-C. PMN binding to laminins 1, 2, and 8 could not be induced despite surface expression of functionally active integrin alpha(6)beta(1), a major laminin receptor, demonstrating that expression of alpha(6)beta(1) alone is insufficient for ligand binding and suggesting the involvement of accessory factors. Nevertheless, laminins 1, 8, and 10 supported PMN migration, indicating that differential cellular signaling via laminins is independent of the extent of adhesion. The data demonstrate that adhesive and nonadhesive interactions with components of the endothelial cell basement membrane and subjacent interstitium play decisive roles in controlling PMN movement into sites of inflammation and illustrate that beta(2)-integrins are not essential for such interactions. PMID- 11278781 TI - Analysis of the cyclic nucleotide binding domain of the HERG potassium channel and interactions with KCNE2. AB - Mutations in the cyclic nucleotide binding domain (CNBD) of the human ether-a-go go-related gene (HERG) K+ channel are associated with LQT2, a form of hereditary Long QT syndrome (LQTS). Elevation of cAMP can modulate HERG K+ channels both by direct binding and indirect regulation through protein kinase A. To assess the physiological significance of cAMP binding to HERG, we introduced mutations to disrupt the cyclic nucleotide binding domain. Eight mutants including two naturally occurring LQT2 mutants V822M and R823W were constructed. Relative cAMP binding capacity was reduced or absent in CNBD mutants. Mutant homotetramers carry little or no K+ current despite normal protein abundance and surface expression. Co-expression of mutant and wild-type HERG resulted in currents with altered voltage dependence but without dominant current suppression. The data from co-expression of V822M and wild-type HERG best fit a model where one normal subunit within a tetramer allows nearly normal current expression. The presence of KCNE2, an accessory protein that associates with HERG, however, conferred a partially dominant current suppression by CNBD mutants. Thus KCNE2 plays a pivotal role in determining the phenotypic severity of some forms of LQT2, which suggests that the CNBD of HERG may be involved in its interaction with KCNE2. PMID- 11278782 TI - Both phosphorylation and caspase-mediated cleavage contribute to regulation of the Ste20-like protein kinase Mst1 during CD95/Fas-induced apoptosis. AB - The serine/threonine kinase Mst1, a mammalian homolog of the budding yeast Ste20 kinase, is cleaved by caspase-mediated proteolysis in response to apoptotic stimuli such as ligation of CD95/Fas or treatment with staurosporine. Furthermore, overexpression of Mst1 induces morphological changes characteristic of apoptosis in human B lymphoma cells. Mst1 may therefore represent an important target for caspases during cell death which serves to amplify the apoptotic response. Here we report that Mst1 has two caspase cleavage sites, and we present evidence indicating that cleavage may occur in an ordered fashion and be mediated by distinct caspases. We also show that caspase-mediated cleavage alone is insufficient to activate Mst1, suggesting that full activation of Mst1 during apoptosis requires both phosphorylation and proteolysis. Another role of phosphorylation may be to influence the susceptibility of Mst1 to proteolysis. Autophosphorylation of Mst1 on a serine residue close to one of the caspase sites inhibited caspase-mediated cleavage in vitro. Finally, Mst1 appears to function upstream of the protein kinase MEKK1 in the SAPK pathway. In conclusion, Mst1 activity is regulated by both phosphorylation and proteolysis, suggesting that protein kinase and caspase pathways work in concert to regulate cell death. PMID- 11278783 TI - A possible role of Ku in mediating sequential repair of closely opposed lesions. AB - One of the hallmarks of ionizing radiation exposure is the formation of clustered damage that includes closely opposed lesions. We show that the Ku70/80 complex (Ku) has a role in the repair of closely opposed lesions in DNA. DNA containing a dihydrouracil (DHU) close to an opposing single strand break was used as a model substrate. It was found that Ku has no effect on the enzymatic activity of human endonuclease III when the substrate DNA contains only DHU. However, with DNA containing a DHU that is closely opposed to a single strand break, Ku inhibited the nicking activity of human endonuclease III as well as the amount of free double strand breaks induced by the enzyme. The inhibition on the formation of a free double strand break by Ku was found to be much greater than the inhibition of human endonuclease III-nicking activity by Ku. Furthermore, there was a concomitant increase in the formation of Ku-DNA complexes when endonuclease III was present. Similar results were also observed with Escherichia coli endonuclease III. These results suggest that Ku reduces the formation of endonuclease III-induced free double strand breaks by sequestering the double strand breaks formed as a Ku-DNA complex. In doing so, Ku helps to avoid the formation of the intermediary free double strand breaks, possibly helping to reduce the mutagenic event that might result from the misjoining of frank double strand breaks. PMID- 11278784 TI - MalR-mediated regulation of the Streptococcus pneumoniae malMP operon at promoter PM. Influence of a proximal divergent promoter region and competition between MalR and RNA polymerase proteins. AB - The Streptococcus pneumoniae mal regulon contains two operons, malXCD and malMP involved in the uptake and utilization of maltosaccharides. Both operons are transcribed from two divergent promoters, P(X) and P(M), and are negatively regulated by the MalR transcriptional repressor. Purified MalR protein binds to two DNA regions that encompasses both promoters, thus occupying its two operators, O(M) and O(X). However, the levels of occupation and repression were different, being higher when MalR was bound to O(M) than when it was anchored to O(X). Competition experiments between MalR and the Escherichia coli RNA polymerase on promoters P(M) and P(X) showed that the affinity of either protein for the promoter/operator DNA sequences was important to determine the frequency of transcription initiation. In addition to the control exerted by MalR, expression from promoter P(M) was affected by upstream sequences located within or close to P(X) promoter. PMID- 11278785 TI - Regulation of sterol regulatory element-binding proteins in hamster intestine by changes in cholesterol flux. AB - A control chow diet or diets containing 1% cholesterol (cholesterol-enriched) or 4% cholestyramine and 0.15% lovastatin (cholesterol-depletion) were fed to hamsters for 2 weeks. Sterol regulatory element-binding protein (SREBP)-1a, SREBP 1c, SREBP-2, 3-hydroxy-3-methylglutaryl-coenzyme A reductase, 3-hydroxy-3 methylglutaryl-coenzyme A synthase, and LDL receptor mRNA levels and SREBP-1 and 2 protein expression were estimated in villus cell populations from duodenum, jejunum, and ileum. SREBP-1a was a minor transcript in hamster intestine, and its gene expression was not altered by changes in dietary cholesterol flux. In contrast, SREBP-1c gene expression was increased by dietary cholesterol and decreased by cholesterol depletion. mRNA levels for SREBP-2 and the other sterol responsive genes were increased in intestines of animals on the cholesterol depletion diet but minimally suppressed if at all, by the diet enriched in cholesterol. In general, the amount of the precursor form of SREBP-1 was higher in cells of the upper villus and lower in cells of the lower villus. SREBP-2 precursor was higher in cells of the lower villus and lower in cells of the upper villus. Protein expression of precursor correlated with the location of gene expression for SREBPs. The amount of precursor mass of SREBP-2 was not altered by cholesterol feeding but was increased by cholesterol depletion. The mature form of SREBP-2 was in very low abundance and difficult to detect in intestines of animals fed control chow or cholesterol. It was readily detectable and increased in intestines of animals on the cholesterol-depletion diet. The diets did not significantly alter the amount of precursor or mature forms of SREBP-1. Cholesterol feeding had no effect on cholesterol or fatty acid synthesis, whereas synthesis of these lipids was increased in intestines of hamsters on the cholesterol-depleted diet. These results suggest that SREBP-1a has little or no role in regulating intestinal cholesterol synthesis. It is postulated that under basal conditions, SREBP-1c regulates intestinal fatty acid synthesis and SREBP-2 regulates cholesterol synthesis. Following marked changes in cholesterol flux across the intestine, SREBP-2 assumes the role of SREBP-1 and regulates both cholesterol and fatty acid synthesis in intestine. PMID- 11278786 TI - Activation of cell division protein FtsZ. Control of switch loop T3 conformation by the nucleotide gamma-phosphate. AB - The effect of bound nucleotide on the conformation of cell division protein FtsZ from Methanococcus jannaschii has been investigated using molecular dynamics and site-directed mutagenesis. The molecular dynamics indicate that the gamma phosphate of GTP induces a conformational perturbation in loop T3 (Gly88-Gly99 segment), in a position structurally equivalent to switch II of Ha-ras-p21. In the simulated GTP-bound state, loop T3 is pulled by the gamma-phosphate into a more compact conformation than with GDP, related to that observed in the homologous proteins alpha- and beta-tubulin. The existence of a nucleotide induced structural change in loop T3 has been confirmed by mutating Thr92 into Trp (T92W-W319Y FtsZ). This tryptophan (12 A away from gamma-phosphate) shows large differences in fluorescence emission, depending on which nucleotide is bound to FtsZ monomers. Loop T3 is located at a side of the contact interface between two FtsZ monomers in the current model of FtsZ filament. Such a structural change may bend the GDP filament upon hydrolysis by pushing against helix H8 of next monomer, thus, generating force on the membrane during cell division. A related curvature mechanism may operate in tubulin activation. PMID- 11278787 TI - In vitro RNA synthesis from exogenous dengue viral RNA templates requires long range interactions between 5'- and 3'-terminal regions that influence RNA structure. AB - Viral replicases of many positive-strand RNA viruses are membrane-bound complexes of cellular and viral proteins that include viral RNA-dependent RNA polymerase (RdRP). The in vitro RdRP assay system that utilizes cytoplasmic extracts from dengue viral-infected cells and exogenous RNA templates was developed to understand the mechanism of viral replication in vivo. Our results indicated that in vitro RNA synthesis at the 3'-untranslated region (UTR) required the presence of the 5'-terminal region (TR) and the two cyclization (CYC) motifs suggesting a functional interaction between the TRs. In this study, using a psoralen-UV cross linking method and an in vitro RdRP assay, we analyzed structural determinants for physical and functional interactions. Exogenous RNA templates that were used in the assays contained deletion mutations in the 5'-TR and substitution mutations in the 3'-stem-loop structure including those that would disrupt the predicted pseudoknot structure. Our results indicate that there is physical interaction between the 5'-TR and 3'-UTR that requires only the CYC motifs. RNA synthesis at the 3'-UTR, however, requires long range interactions involving the 5'-UTR, CYC motifs, and the 3'-stem-loop region that includes the tertiary pseudoknot structure. PMID- 11278788 TI - Evidence for a functional monomeric form of the bacteriophage T4 DdA helicase. Dda does not form stable oligomeric structures. AB - The active form of many helicases is oligomeric, possibly because oligomerization provides multiple DNA binding sites needed for unwinding of DNA. In order to understand the mechanism of the bacteriophage T4 Dda helicase, the potential requirement for oligomerization was investigated. Chemical cross-linking and high pressure gel filtration chromatography provided little evidence for the formation of an oligomeric species. The specific activity for ssDNA stimulated ATPase activity was independent of Dda concentration. Dda was mutated to produce an ATPase-deficient protein (K38A Dda) by altering a residue within a conserved, nucleotide binding loop. The helicase activity of K38A Dda was inactivated, although DNA binding properties were similar to Dda. In the presence of limiting DNA substrate, the rate of unwinding by Dda was not changed; however, the amplitude of product formation was reduced in the presence of increasing concentrations of K38A Dda. The reduction was between that expected for a monomeric or dimeric helicase based on simple competition for substrate binding. When unwinding of DNA was measured in the presence of excess DNA substrate, addition of K38A Dda caused no reduction in the observed rate for strand separation. Taken together, these results indicate that oligomerization of Dda is not required for DNA unwinding. PMID- 11278789 TI - MAP kinases mediate UVB-induced phosphorylation of histone H3 at serine 28. AB - Histone H3 phosphorylation is related closely to chromatin remodeling and chromosome condensation. H3 phosphorylation at serine 28 is coupled with mitotic chromosome condensation in diverse mammalian cell lines. However, the pathway that mediates phosphorylation of H3 at serine 28 is unknown. In the present study, ERK1, ERK2, or p38 kinase strongly phosphorylated H3 at serine 28 in vitro. JNK1 or JNK2 was able also to phosphorylate H3 at serine 28 in vitro but to a lesser degree. UVB irradiation markedly induced phosphorylation of H3 at serine 28 in JB6 Cl 41 cells. PD 98059, a MEK1 inhibitor, and SB 202190, a p38 kinase inhibitor, efficiently repressed UVB-induced H3 phosphorylation at serine 28. Expression of dominant negative mutant (DNM) ERK2 in JB6 Cl 41 cells totally blocked UVB-induced phosphorylation of H3 at serine 28. Additionally, DNM p38 kinase or DNM JNK1 partially blocked UVB-induced H3 phosphorylation at serine 28. Furthermore, UVB-induced H3 phosphorylation at serine 28 was inhibited in Jnk1(-/ ) cells but not in Jnk2(-/-) cells. These results suggest that UVB-induced H3 phosphorylation at serine 28 may be mediated by mitogen-activated protein kinases. PMID- 11278790 TI - Rat seminal vesicle FAD-dependent sulfhydryl oxidase. Biochemical characterization and molecular cloning of a member of the new sulfhydryl oxidase/quiescin Q6 gene family. AB - Rat FAD-dependent sulfhydryl oxidase was purified; partial sequencing indicated that it was homologous to human quiescin Q6. A cDNA (GenBank accession no. AF285078) was cloned from rat seminal vesicles, and active recombinant sulfhydryl oxidase was expressed in Chinese hamster ovary epithelial cells. This 2472 nucleotide cDNA has an open reading frame of 1710 base pairs, encoding a protein of 570 amino acids including a 32-amino acid leader sequence and two potential sites for N-glycosylation. One of them is used and the 64,000 M(r) purified protein was transformed to 61,000 by the action of endoglycosidase F. Northern blotting and reverse transcription-polymerase chain reaction analyses showed that there were small amounts of sulfhydryl oxidase in the rat testis, prostate, lung, heart, kidney, spleen, and liver, and that the gene was highly expressed in seminal vesicles and epididymis. Rat sulfhydryl oxidase cDNA corresponds to the human cell growth inhibiting factor cDNA, which could be a differently spliced form of quiescin Q6. Comparing sulfhydryl oxidase sequences with those of human quiescin Q6 and mammalian and Caenorhabditis elegans quiescin Q6-related genes established the existence of a new family of FAD-dependent sulfhydryl oxidase/quiescin Q6-related genes containing protein-disulfide isomerase-type thioredoxin and yeast ERV1 domains. PMID- 11278791 TI - Parathyroid hormone-induced bone resorption does not occur in the absence of osteopontin. AB - Osteopontin is an RGDS-containing protein that acts as a ligand for the alpha(v)beta(3) integrin, which is abundantly expressed in osteoclasts, cells responsible for bone resorption in osteopenic diseases such as osteoporosis and hyperparathyroidism. However, the role of osteopontin in the process of bone resorption has not yet been fully understood. Therefore, we investigated the direct function of osteopontin in bone resorption using an organ culture system. The amount of (45)Ca released from the osteopontin-deficient bones was not significantly different from the basal release from wild type bones. However, in contrast to the parathyroid hormone (PTH) enhancement of the (45)Ca release from wild type bones, PTH had no effect on (45)Ca release from organ cultures of osteopontin-deficient bones. Because PTH is located upstream of receptor activator of NF-kappaB ligand (RANKL), that directly promotes bone resorption, we also examined the effect of RANKL. Soluble RANKL with macrophage-colony stimulating factor enhanced (45)Ca release from the bones of wild type fetal mice but not from the bones of osteopontin-deficient mice. To obtain insight into the cellular mechanism underlying the phenomena observed in osteopontin-deficient bone, we investigated the number of tartrate-resistant acid phosphatase (TRAP) positive cells in the bones subjected to PTH treatment in cultures. The number of TRAP-positive cells was increased significantly by PTH in wild type bone; however, no such PTH-induced increase in TRAP-positive cells was observed in osteopontin-deficient bones. These results indicate that the absence of osteopontin suppressed PTH-induced increase in bone resorption via preventing the increase in the number of osteoclasts in the local milieu of bone. PMID- 11278792 TI - Structural and functional analysis of a bipolar replication terminus. Implications for the origin of polarity of fork arrest. AB - We have delineated the amino acid to nucleotide contacts made by two interacting dimers of the replication terminator protein (RTP) of Bacillus subtilis with a novel naturally occurring bipolar replication terminus by converting RTP to a site-directed chemical nuclease and mapping its cleavage sites on the terminus. The data show a relatively symmetrical arrangement of the amino acid to base contacts, and a comparison of the bipolar contacts with that of a normal unipolar terminus suggests that the DNA-protein contacts play an important determinative role in generating polarity from structurally symmetrical RTP dimers. The amino acid to nucleotide contacts provided distance constraints that enabled us to build a three-dimensional model of the protein-DNA complex. The model is consistent with features of the bipolar Ter.RTP complex derived from mutational and cross-linking data. The bipolar terminus arrested Escherichia coli DNA replication and DnaB helicase and T7 RNA polymerase in vitro in both orientations. RTP arrested the unwinding of duplex DNA on the bipolar Ter DNA substrate regardless of the length of the duplex DNA. The latter result suggested further that the terminus arrested authentic DNA unwinding by the helicase rather than just translocation of helicase on DNA. PMID- 11278793 TI - H+-coupled pantothenate transport in the intracellular malaria parasite. AB - Pantothenate, the precursor of coenzyme A, is an essential nutrient for the intraerythrocytic stage of the malaria parasite Plasmodium falciparum. Pantothenate enters the malaria-infected erythrocyte via new permeation pathways induced by the parasite in the host cell membrane (Saliba, K. J., Horner, H. A., and Kirk, K. (1998) J. Biol. Chem. 273, 10190-10195). We show here that pantothenate is taken up by the intracellular parasite via a novel H(+)-coupled transporter, quite different from the Na(+)-coupled transporters that mediate pantothenate uptake into mammalian cells. The plasmodial H(+):pantothenate transporter has a low affinity for pantothenate (K(m) approximately 23 mm) and a stoichiometry of 1 H(+):1 pantothenate. It is inhibited by low concentrations of the bioflavonoid phloretin and the thiol-modifying agent p-chloromercuribenzene sulfonate. On entering the parasite, pantothenate is phosphorylated (and thereby trapped) by an unusually high affinity pantothenate kinase (K(m) approximately 300 nm). The combination of H(+)-coupled transporter and kinase provides the parasite with an efficient, high affinity pantothenate uptake system, which is distinct from that of the host and is therefore an attractive target for antimalarial chemotherapy. PMID- 11278794 TI - Specific association of a set of molecular chaperones including HSP90 and Cdc37 with MOK, a member of the mitogen-activated protein kinase superfamily. AB - We have recently identified and cloned a novel member of mitogen-activated protein kinase superfamily protein, MOK (Miyata, Y., Akashi, M., and Nishida, E. (1999) Genes Cells 4, 299-309). To address its regulatory mechanisms, we searched for cellular proteins that specifically associate with MOK by co immunoprecipitation experiments. Several cellular proteins including a major 90 kDa molecular chaperone HSP90 were found associated with MOK. Treatment of cells with geldanamycin, an HSP90-specific inhibitor, rapidly decreased the protein level of MOK, and the decrease was attributed to enhanced degradation of MOK through proteasome-dependent pathways. Our data suggest that the association with HSP90 may regulate intracellular protein stability and solubility of MOK. Experiments with a series of deletion mutants of MOK indicated that the region encompassing the protein kinase catalytic subdomains I-IV is required for HSP90 binding. Closely related protein kinases (male germ cell-associated kinase and male germ cell-associated kinase-related kinase) were also found to associate with HSP90, whereas conventional mitogen-activated protein kinases (extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase/stress-activated protein kinase) were not associated with HSP90. In addition, we found that other molecular chaperones including Cdc37, HSC70, HSP70, and HSP60 but not GRP94, FKBP52, or Hop were detected specifically in the MOK-HSP90 immunocomplexes. These results taken together suggest a role of a specific set of molecular chaperones in the stability of signal-transducing protein kinases. PMID- 11278797 TI - Proteolytic activation of ETK/Bmx tyrosine kinase by caspases. AB - Etk/Bmx is a member of the Btk/Tec family of kinases, which are characterized by having a pleckstrin homology domain at the N terminus, in addition to the Src homology 3 (SH3), SH2, and the catalytic domains, shared with the Src family kinases. Etk, or Btk kinases in general, has been implicated in the regulation of apoptosis. To test whether Etk is the substrate for caspases during apoptosis, in vitro translated [(35)S]methionine-labeled Etk was incubated with different apoptotic extracts and recombinant caspases, respectively. Results showed that Etk was proteolyzed in all conditions tested with identical cleavage patterns. Caspase-mediated cleavage of Etk generated a C-terminal fragment, containing the complete SH2 and tyrosine kinase domains, but without intact pleckstrin homology and SH3 domains. This fragment has 4-fold higher kinase activity than that of the full-length Etk. Ectopic expression of the C-terminal fragment of Etk sensitized the PC3 prostate cancer cells to apoptosis in response to apoptosis-inducing stimuli. The finding, together with an earlier report that Etk is potentially antiapoptotic, suggests that Etk may serve as an apoptotic switch, depending on the forms of Etk existing inside the cells. To our knowledge, this is the first case where the activity of a tyrosine kinase is induced by caspase cleavage. PMID- 11278796 TI - Functionally different AU- and G-rich cis-elements confer developmentally regulated mRNA stability in Trypanosoma cruzi by interaction with specific RNA binding proteins. AB - Post-transcriptional regulatory mechanisms have been suggested to be the main point of control of gene expression in kinetoplastid parasites. We have previously shown that Trypanosoma cruzi SMUG mucin mRNA steady-state level is developmentally regulated by post-transcriptional mechanisms, being stable in the epimastigote insect vector stage, but unstable in the trypomastigote infective stage of the parasite. Its turnover is controlled by an AU-rich element (ARE) localized in the 3'-untranslated region, since a reporter gene lacking this sequence was stable in the trypomastigote stage (Di Noia, J. M., D'Orso, I., Sanchez, D. O., and Frasch, A. C. (2000) J. Biol. Chem. 275, 10218-10227). Here, we show by gel mobility shift assay that the 44-nt ARE sequence interacts with a set of stage-specific AU-rich element RNA-binding proteins (ARE-BPs). The epimastigote stage AU-rich element RNA-binding protein, named E-ARE-BP, and the trypomastigote stage ARE-BPs, named T-ARE-BPs, are efficiently competed by poly(U). UV cross-linking analysis showed that E-ARE-BP has an apparent molecular mass of 100 kDa and is different from the 45-50-kDa ARE-BPs present in other stages of the parasite. Transfection experiments allowed the identification of a novel cis-element that might be responsible for a positive effect on mRNA stability. It is a G-rich element, named GRE, composed by two contiguous CGGGG pentamers. The factors that recognize GRE were different from the ones that bind to ARE, in both molecular masses and subcellular localization. Thus, ARE and GRE are functionally different cis-elements, which might regulate mucin expression throughout the parasite life cycle. PMID- 11278795 TI - Cholesterol, a cell size-dependent signal that regulates glucose metabolism and gene expression in adipocytes. AB - Enlarged fat cells exhibit modified metabolic capacities, which could be involved in the metabolic complications of obesity at the whole body level. We show here that sterol regulatory element-binding protein 2 (SREBP-2) and its target genes are induced in the adipose tissue of several models of rodent obesity, suggesting cholesterol imbalance in enlarged adipocytes. Within a particular fat pad, larger adipocytes have reduced membrane cholesterol concentrations compared with smaller fat cells, demonstrating that altered cholesterol distribution is characteristic of adipocyte hypertrophy per se. We show that treatment with methyl-beta cyclodextrin, which mimics the membrane cholesterol reduction of hypertrophied adipocytes, induces insulin resistance. We also produced cholesterol depletion by mevastatin treatment, which activates SREBP-2 and its target genes. The analysis of 40 adipocyte genes showed that the response to cholesterol depletion implicated genes involved in cholesterol traffic (caveolin 2, scavenger receptor BI, and ATP binding cassette 1 genes) but also adipocyte-derived secretion products (tumor necrosis factor alpha, angiotensinogen, and interleukin-6) and proteins involved in energy metabolism (fatty acid synthase, GLUT 4, and UCP3). These data demonstrate that altering cholesterol balance profoundly modifies adipocyte metabolism in a way resembling that seen in hypertrophied fat cells from obese rodents or humans. This is the first evidence that intracellular cholesterol might serve as a link between fat cell size and adipocyte metabolic activity. PMID- 11278798 TI - An active site tyrosine influences the ability of the dimethyl sulfoxide reductase family of molybdopterin enzymes to reduce S-oxides. AB - Dimethyl sulfoxide reductase (DMSOR), trimethylamine-N-oxide reductase (TMAOR), and biotin sulfoxide reductase (BSOR) are members of a class of bacterial oxotransferases that contain the bis(molybdopterin guanine dinucleotide)molybdenum cofactor. The presence of a Tyr residue in the active site of DMSOR and BSOR that is missing in TMAOR has been implicated in the inability of TMAOR, unlike DMSOR and BSOR, to utilize S-oxides. To test this hypothesis, Escherichia coli TMAOR was cloned and expressed at high levels, and site-directed mutagenesis was utilized to generate the Tyr-114 --> Ala and Phe variants of Rhodobacter sphaeroides DMSOR and insert a Tyr residue into the equivalent position in TMAOR. Although all of the mutants turn over in a manner similar to their respective wild-type enzymes, mutation of Tyr-114 in DMSOR results in a decreased specificity for S-oxides and an increased specificity for trimethylamine-N-oxide (Me(3)NO), with a greater change observed for DMSOR-Y114A. Insertion of a Tyr into TMAOR results in a decreased preference for Me(3)NO relative to dimethyl sulfoxide. Kinetic analysis and UV-visible absorption spectra indicate that the ability of DMSOR to be reduced by dimethyl sulfide is lost upon mutation of Tyr-114 and that TMAOR does not exhibit this activity even in the Tyr insertion mutant. PMID- 11278799 TI - Distinct binding determinants for ERK2/p38alpha and JNK map kinases mediate catalytic activation and substrate selectivity of map kinase phosphatase-1. AB - Mitogen-activated protein (MAP) kinase phosphatase 1 (MKP-1/CL100) is an inducible nuclear dual specificity protein phosphatase that can dephosphorylate and inactivate both mitogen- and stress-activated protein kinases in vitro and in vivo. However, the molecular mechanism responsible for the substrate selectivity of MKP-1 is unknown. In addition, it has been suggested that the signal transducers and activators of transcription 1 (STAT1) transcription factor is a physiological non-MAP kinase substrate for MKP-1. We have used the yeast two hybrid assay to demonstrate that MKP-1 is able to interact selectively with the extracellular signal-regulated kinase 1/2 (ERK1/2), p38alpha, and c-Jun NH(2) terminal kinase (JNK) MAP kinase isoforms. Furthermore, this binding is accompanied by catalytic activation of recombinant MKP-1 protein in vitro, and these end points show an absolute correlation with MKP-1 substrate selectivity in vivo. In contrast, MKP-1 does not interact with STAT1. Recombinant STAT1 does not cause catalytic activation of MKP-1; nor does MKP-1 block tyrosine phosphorylation of STAT1 in vivo. Both binding and catalytic activation of MKP-1 are abrogated by mutation of a conserved docking site in ERK2, p38alpha, and JNK1 MAP kinases. Within MKP-1, MAP kinase binding is mediated by the amino-terminal noncatalytic domain of the protein. However, mutation of a conserved cluster of positively charged residues within this domain abolishes the binding and activation of MKP-1 by ERK2 and p38alpha but not JNK1, indicating that there are distinct binding determinants for these MAP kinase isoforms. We conclude that the substrate selectivity of MKP-1 is determined by specific protein-protein interactions coupled with catalytic activation of the phosphatase and that these interactions are restricted to members of the MAP kinase family of enzymes. PMID- 11278800 TI - Programming the transcriptional state of replicating methylated dna. AB - CpG methylation is maintained in daughter chromatids by the action of DNA methyltransferase at the replication fork. An opportunity exists for transcription factors at replication forks to bind their cognate sequences and thereby prevent remethylation by DNA methyltransferase. To test this hypothesis, we injected a linearized, methylated, and partially single-stranded reporter plasmid into the nuclei of Xenopus oocytes and followed changes in the transcriptional activity after DNA replication. We find that dependent on Gal4 VP16, the action of DNA methyltransferase, and replication-coupled chromatin assembly DNA replication provides a window of time in which regulatory factors can activate or repress gene activity. Demethylation in the promoter region near the GAL4 binding sites of the newly synthesized DNA did not occur even though the Gal4 binding sites were occupied and transcription was activated. We conclude that "passive" demethylation at the replication fork is not simply dependent on the presence of DNA binding transcriptional activators. PMID- 11278801 TI - Cyclobutane pyrimidine dimers and bulky chemical DNA adducts are efficiently repaired in both strands of either a transcriptionally active or promoter-deleted APRT gene. AB - Both prokaryotic and eukaryotic cells have the capacity to repair DNA damage preferentially in the transcribed strand of actively expressed genes. However, we have found that several types of DNA damage, including cyclobutane pyrimidine dimers (CPDs) are repaired with equal efficiency in both the transcribed and nontranscribed strands of the adenine phosphoribosyltransferase (APRT) gene in Chinese hamster ovary cells. We further found that, in two mutant cell lines in which the entire APRT promoter region has been deleted, CPDs are still efficiently repaired in both strands of the promoterless APRT gene, even though neither strand appears to be transcribed. These results suggest that efficient repair of both strands at this locus does not require transcription of the APRT gene. We have also mapped CPD repair in exon 3 of the APRT gene in each cell line at single nucleotide resolution. Again, we found similar rates of CPD repair in both strands of the APRT gene domain in both APRT promoter-deletion mutants and their parental cell line. Our findings suggest that current models of transcription-coupled repair and global genomic repair may underestimate the importance of factors other than transcription in governing the efficiency of nucleotide excision repair. PMID- 11278802 TI - Three RNA polymerase II carboxyl-terminal domain kinases display distinct substrate preferences. AB - CDK7, CDK8, and CDK9 are cyclin-dependent kinases (CDKs) that phosphorylate the C terminal domain (CTD) of RNA polymerase II. They have distinct functions in transcription. Because the three CDKs target only serine 5 in the heptad repeat of model CTD substrates containing various numbers of repeats, we tested the hypothesis that the kinases differ in their ability to phosphorylate CTD heptad arrays. Our data show that the kinases display different preferences for phosphorylating individual heptads in a synthetic CTD substrate containing three heptamer repeats and specific regions of the CTD in glutathione S-transferase fusion proteins. They also exhibit differences in their ability to phosphorylate a synthetic CTD peptide that contains Ser-2-PO(4). This phosphorylated peptide is a poor substrate for CDK9 complexes. CDK8 and CDK9 complexes, bound to viral activators E1A and Tat, respectively, target only serine 5 for phosphorylation in the CTD peptides, and binding to the viral activators does not change the substrate preference of these kinases. These results imply that the display of different CTD heptads during transcription, as well as their phosphorylation state, can affect their phosphorylation by the different transcription-associated CDKs. PMID- 11278803 TI - Mutant presenilins disturb neuronal calcium homeostasis in the brain of transgenic mice, decreasing the threshold for excitotoxicity and facilitating long-term potentiation. AB - Mutant human presenilin-1 (PS1) causes an Alzheimer's-related phenotype in the brain of transgenic mice in combination with mutant human amyloid precursor protein by means of increased production of amyloid peptides (Dewachter, I., Van Dorpe, J., Smeijers, L., Gilis, M., Kuiperi, C., Laenen, I., Caluwaerts, N., Moechars, D., Checler, F., Vanderstichele, H. & Van Leuven, F. (2000) J. Neurosci. 20, 6452-6458) that aggravate plaques and cerebrovascular amyloid (Van Dorpe, J., Smeijers, L., Dewachter, I., Nuyens, D., Spittaels, K., van den Haute, C., Mercken, M., Moechars, D., Laenen, I., Kuiperi, C., Bruynseels, K., Tesseur, I., Loos, R., Vanderstichele, H., Checler, F., Sciot, R. & Van Leuven, F. (2000) J. Am. Pathol. 157, 1283-1298). This gain of function of mutant PS1 is approached here in three paradigms that relate to glutamate neurotransmission. Mutant but not wild-type human PS1 (i) lowered the excitotoxic threshold for kainic acid in vivo, (ii) facilitated hippocampal long-term potentiation in brain slices, and (iii) increased glutamate-induced intracellular calcium levels in isolated neurons. Prominent higher calcium responses were triggered by thapsigargin and bradykinin, indicating that mutant PS modulates the dynamic release and storage of calcium ions in the endoplasmatic reticulum. In reaction to glutamate, overfilled Ca(2+) stores resulted in higher than normal cytosolic Ca(2+) levels, explaining the facilitated long-term potentiation and enhanced excitotoxicity. The lowered excitotoxic threshold for kainic acid was also observed in mice transgenic for mutant human PS2[N141I] and was prevented by dantrolene, an inhibitor of Ca(2+) release from the endoplasmic reticulum. PMID- 11278804 TI - A single carboxyl mutant of the multidrug transporter EmrE is fully functional. AB - EmrE, a multidrug transporter from Escherichia coli removes toxic compounds from the cell in exchange with protons. Glu-14 is the only charged residue in the putative membrane domains and is fully conserved in more than 50 homologues of the protein. This residue was shown to be an essential part of the binding site, common to protons and substrate. EmrE bearing a single carboxylic residue, Glu 14, shows uptake and binding properties similar to those of the wild type. This suggests that a small protein bearing only 110 amino acids with a single carboxyl in position 14 is the most basic structure that shows ion-coupled transport activity. The role of Glu-14 in substrate binding was examined by using dicyclohexylcarbodiimide, a hydrophobic carbodiimide that is known to react with carboxyls. Tetraphenylphosphonium binding to both wild type and the single carboxyl mutant is inhibited by dicyclohexylcarbodiimide in a dose-dependent manner. Ethidium and other substrates of EmrE prevent this inhibition with an order of potency in accord with their apparent affinities. This suggests that dicyclohexylcarbodiimide binding is sterically prevented by the substrate, supporting the contention that Glu-14, the reactive residue, is part of the substrate-binding site. PMID- 11278805 TI - Transcriptional activation of mouse sst2 somatostatin receptor promoter by transforming growth factor-beta. Involvement of Smad4. AB - The sst2 somatostatin receptor is an inhibitory G protein-coupled receptor, which exhibits anti-tumor properties. Expression of sst2 is lost in most human pancreatic cancers. We have cloned 2090 base pairs corresponding to the genomic DNA region upstream of the mouse sst2 (msst2) translation initiation codon (ATG). Deletion reporter analyses in mouse pituitary AtT-20 and human pancreatic cancer PANC-1, BxPC-3, and Capan-1 cells identify a region from nucleotide -260 to the ATG codon (325 base pairs) showing maximal activity, and a region between nucleotides -2025 and -260 likely to comprise silencer or transcriptional suppressor elements. In PANC-1 and AtT-20 cells, transforming growth factor (TGF) beta up-regulates msst2 transcription. Transactivation is mediated by Smad4 and Smad3. The cis-acting region responsible for such regulation is comprised between nucleotides -1115 and -972 and includes Sp1 and CAGA-box sequences. Expression of Smad4 in Smad4-deficient Capan-1 and BxPC-3 cells restores TGF-beta-dependent and -independent msst2 transactivation. Expression of Smad4 in BxPC-3 cells reestablishes both endogenous sst2 expression and somatostatin-mediated inhibition of cell growth. These findings demonstrate that msst2 is a new target gene for TGF-beta transcription regulation and underlie the possibility that loss of Smad4 contributes to the lack of sst2 expression in human pancreatic cancer, which in turn may contribute to a stimulation of tumor growth. PMID- 11278806 TI - Muscle specificity encoded by specific serum response factor-binding sites. AB - Serum response factor (SRF) is a MADS box transcription factor that regulates muscle-specific and growth factor-inducible genes by binding the consensus sequence CC(A/T)6GG, known as a CArG box. Because SRF expression is not restricted solely to muscle, its expression alone cannot account for the muscle specificity of some of its target genes. To understand further the role of SRF in muscle-specific transcription, we created transgenic mice harboring lacZ transgenes linked to tandem copies of different CArG boxes with flanking sequences. CArG boxes from the SM22 and skeletal alpha-actin promoters directed highly restricted expression in developing smooth, cardiac, and skeletal muscle cells during early embryogenesis. In contrast, the CArG box and flanking sequences from the c-fos promoter directed expression throughout the embryo, with no preference for muscle cells. Systematic swapping of the core and flanking sequences of the SM22 and c-fos CArG boxes revealed that cell type specificity was dictated in large part by sequences immediately flanking the CArG box core. Sequences that directed widespread embryonic expression bound SRF more strongly than those that directed muscle-restricted expression. We conclude that sequence variations among CArG boxes influence cell type specificity of expression and account, at least in part, for the ability of SRF to distinguish between growth factor-inducible and muscle-specific genes in vivo. PMID- 11278807 TI - The role of the synaptic protein snap-25 in the potency of botulinum neurotoxin type A. AB - Botulinum neurotoxin serotype A (BoNT/A) is distinguished from BoNT/E by longer duration of paralysis and greater potency. The proteolytic activity of BoNT/A in cultures of dissociated spinal cord neurons persists beyond 80 days, whereas BoNT/E activity persists for less than 1 day (Keller, J. E., Neale, E. A. Oyler, G., and Adler, M. (1999) FEBS Lett. 456, 137-142). This single quality of toxin activity can account for the differences observed in the duration of muscle block. In the present work we sought to understand the basis for the apparent greater potency of BoNT/A. BoNT/E cleaves a 26-amino acid fragment from the C terminus of the synaptic protein SNAP-25 whereas BoNT/A removes only nine residues creating a 197-amino acid fragment (P197) that is 95% the length of SNAP 25. We show that inhibition of neurotransmitter release by BoNT/E is equivalent to the damage caused to SNAP-25. However, synaptic blockade by BoNT/A is greater than the extent of SNAP-25 proteolysis. These findings can be explained if P197 produces an inhibitory effect on neurotransmitter release. A mathematical model of the experimentally determined relationship between SNAP-25 damage and blockade of neurotransmission supports this interpretation. Furthermore, neurotransmitter release following complete cleavage of SNAP-25 can be achieved by P197, but with about 5-fold less sensitivity to external Ca(2+). In this case, vesicular release is restored by increasing intracellular Ca(2+). These data demonstrate that P197 competes with intact SNAP-25, but is unable to initiate normal synaptic vesicle fusion in physiological concentrations of Ca(2+). PMID- 11278808 TI - Prostate apoptosis response-4 enhances secretion of amyloid beta peptide 1-42 in human neuroblastoma IMR-32 cells by a caspase-dependent pathway. AB - Prostate apoptosis response-4 (Par-4) is a leucine zipper protein that promotes neuronal cell death in Alzheimer's disease (AD). Neuronal degeneration in AD may result from extracellular accumulation of amyloid beta peptide (Abeta) 1-42. To examine the effect of Par-4 on Abeta secretion and to reconcile amyloid/apoptosis hypotheses of AD, we generated IMR-32 cell lines that overexpress Par-4 and/or its leucine zipper domain. Overexpression of Par-4 did not significantly affect levels of the endogenously expressed beta amyloid precursor protein but drastically increased the Abeta(1-42)/Abeta(total) ratio in the conditioned media about 6-8 h after trophic factor withdrawal. Time course analysis of caspase activation reveals that Par-4 overexpression exacerbated caspase activation, which is detectable within 2 h after trophic factor withdrawal. Furthermore, inhibition of caspase activity by the broad spectrum caspase inhibitor BD-fmk significantly attenuated the Par-4-induced increase in Abeta 1-42 production. In addition, the effects of Par-4 on secretion of Abeta 1-42 were consistently blocked by co-expression of the leucine zipper domain, indicating that the effect of Par-4 on Abeta secretion may require its interaction with other protein(s). These results suggest that Par-4 increases secretion of Abeta 1-42 largely through a caspase-dependent pathway after apoptotic cascades are initiated. PMID- 11278809 TI - Selective roles of retinoic acid receptor and retinoid x receptor in the suppression of apoptosis by all-trans-retinoic acid. AB - Retinoic acids exert profound effects on many biological processes including cell proliferation, differentiation, and morphogenesis. We previously reported that all-trans-retinoic acid (t-RA) protected mesangial cells from H(2)O(2)-triggered apoptosis by suppressing the activator protein 1 (AP-1) pathway. It was via inhibition of c-fos and c-jun expression and suppression of c-Jun N-terminal kinase (JNK) activation. In this report, we investigated the involvement of retinoic acid receptor (RAR) and retinoid X receptor (RXR) in the antiapoptotic effect of t-RA in H(2)O(2)-exposed cells. We found that pretreatment with RAR pan antagonist (AGN193109) or RXR pan-antagonist (HX531) attenuated the antiapoptotic effect of t-RA. Similarly, transient transfection with a dominant-negative mutant of RAR or a dominant-negative RXR diminished the antiapoptotic effect of t-RA. Both RAR and RXR antagonists reversed the suppressive effect of t-RA on AP-1 activity. However, the roles of RAR and RXR in the suppression of AP-1 components by t-RA were found to be different. RAR antagonist reversed the suppressive effect of t-RA on both c-fos and c-jun, whereas RXR antagonist reversed the effect of t-RA on c-fos but not c-jun. Furthermore, suppression of JNK activation by t-RA was observed even in the presence of RAR and RXR antagonists. Consistently, suppression of JNK by t-RA was not affected by overexpression of either the dominant-negative RAR or the dominant-negative RXR. These data elucidated that the antiapoptotic effect of t-RA is mediated by both nuclear receptor-dependent and -independent mechanisms. PMID- 11278810 TI - Regulated heterogeneity in 12-kDa P-protein phosphorylation and composition of ribosomes in maize (Zea mays L.). AB - Maize (Zea mays L.) possesses four distinct approximately 12-kDa P-proteins (P1, P2a, P2b, P3) that form the tip of a lateral stalk on the 60 S ribosomal subunit. RNA blot analyses suggested that the expression of these proteins was developmentally regulated. Western blot analysis of ribosomal proteins isolated from various organs, kernel tissues during seed development, and root tips deprived of oxygen (anoxia) revealed significant heterogeneity in the levels of these proteins. P1 and P3 were detected in ribosomes of all samples at similar levels relative to ribosomal protein S6, whereas P2a and P2b levels showed considerable developmental regulation. Both forms of P2 were present in ribosomes of some organs, whereas only one form was detected in other organs. Considerable tissue-specific variation was observed in levels of monomeric and multimeric forms of P2a. P2b was not detected in root tips, accumulated late in seed embryo and endosperm development, and was detected in soluble ribosomes but not in membrane-associated ribosomes that copurified with zein protein bodies of the kernel endosperm. The phosphorylation of the 12-kDa P-proteins was also developmentally and environmentally regulated. The potential role of P2 heterogeneity in P-protein composition in the regulation of translation is discussed. PMID- 11278811 TI - Mouse LYVE-1 is an endocytic receptor for hyaluronan in lymphatic endothelium. AB - The glycosaminoglycan hyaluronan is a key substrate for cell migration in tissues during inflammation, wound healing, and neoplasia. Unlike other matrix components, hyaluronan (HA) is turned over rapidly, yet most degradation occurs not locally but within distant lymph nodes, through mechanisms that are not yet understood. While it is not clear which receptors are involved in binding and uptake of hyaluronan within the lymphatics, one likely candidate is the lymphatic endothelial hyaluronan receptor LYVE-1 recently described in our laboratory (Banerji, S., Ni, J., Wang, S., Clasper, S., Su, J., Tammi, R., Jones, M., and Jackson, D.G. (1999) J. Cell Biol. 144, 789-801). Here we present evidence that LYVE-1 is involved in the uptake of hyaluronan by lymphatic endothelial cells using a new murine LYVE-1 orthologue identified from the EST data base. We show that mouse LYVE-1 both binds and internalizes hyaluronan in transfected 293T fibroblasts in vitro and demonstrate using immunoelectron microscopy that it is distributed equally among the luminal and abluminal surfaces of lymphatic vessels in vivo. In addition, we show by means of specific antisera that expression of mouse LYVE-1 remains restricted to the lymphatics in homozygous knockout mice lacking a functional gene for CD44, the closest homologue of LYVE-1 and the only other Link superfamily HA receptor known to date. Together these results suggest a role for LYVE-1 in the transport of HA from tissue to lymph and imply that further novel hyaluronan receptors must exist that can compensate for the loss of CD44 function. PMID- 11278812 TI - Interferon alpha /beta promotes cell survival by activating nuclear factor kappa B through phosphatidylinositol 3-kinase and Akt. AB - Interferons (IFNs) play critical roles in host defense by modulating gene expression via activation of signal transducer and activator of transcription (STAT) factors. IFN-alpha/beta also activates another transcription factor, nuclear factor kappaB (NF-kappaB), which protects cells against apoptotic stimuli. NF-kappaB activation requires the IFN-dependent association of STAT3 with the IFNAR1 chain of the IFN receptor. IFN-dependent NF-kappaB activation involves the sequential activation of a serine kinase cascade involving phosphatidylinositol 3-kinase (PI-3K) and Akt. Whereas constitutively active PI 3K and Akt induce NF-kappaB activation, Ly294002 (a PI-3K inhibitor), dominant negative PI-3K, and kinase-dead Akt block IFN-dependent NF-kappaB activation. Moreover, dominant-negative PI-3K blocks IFN-promoted degradation of kappaBox alpha. Ly294002, a dominant-negative PI-3K construct, and kinase-dead Akt block IFN-promoted cell survival, enhancing apoptotic cell death. Therefore, STAT3, PI 3K, and Akt are components of an IFN signaling pathway that promotes cell survival through NF-kappaB activation. PMID- 11278813 TI - Disease-associated mutations in the extracytoplasmic loops of cystic fibrosis transmembrane conductance regulator do not impede biosynthetic processing but impair chloride channel stability. AB - Consistent with its function as a chloride channel regulated entirely from the cytoplasmic side of the plasma membrane, the cystic fibrosis transmembrane conductance regulator (CFTR) glycoprotein exposes little of its mass on the exterior surface of cells. The first and fourth extracytoplasmic loops (ELs) contain approximately 15 and 30 residues, respectively; the other four ELs are extremely short. To examine the influence of missense mutants in ELs detected in patients with cystic fibrosis, we have expressed them in mammalian (baby hamster kidney (BHK21)) cells and assessed their biosynthetic processing and chloride channel activity. In contrast to previous findings that 18 of 30 disease associated missense mutations in cytoplasmic loops caused retention of the nascent polypeptides in the endoplasmic reticulum, all the EL mutants studied matured and were transported to the cell surface. This pronounced asymmetry is consistent with the notion that endoplasmic reticulum quality control of nascent CFTR is exerted primarily on the cytoplasmic side of the membrane. Although this set of EL mutations has little effect on CFTR maturation, most of them seriously compromise its chloride channel activity. Substitutions at six different positions in EL1 and single positions in EL2 and EL4 all destabilized the open state, some of them severely, indicating that the ELs contribute to the stability of the CFTR ion pore. PMID- 11278814 TI - Phosphorylation of Smad7 at Ser-249 does not interfere with its inhibitory role in transforming growth factor-beta-dependent signaling but affects Smad7 dependent transcriptional activation. AB - Smad proteins are major components in the intracellular signaling pathway of transforming growth factor-beta (TGF-beta), and phosphorylation is an important mechanism in regulation of their functions. Smad7 was identified as a potent inhibitor of TGF-beta-dependent signaling. We have identified serine 249 in Smad7 as a major phosphorylation site, the phosphorylation of which was not affected by TGF-beta1. Abrogation of the phosphorylation by substitution of Ser-249 with alanine or aspartic acid residues did not affect the ability of Smad7 to inhibit TGF-beta1 and BMP7 signaling. No differences were found in the stability or in the intracellular distribution of Smad7 mutants compared with the wild-type molecule. However, Smad7 fused to the DNA-binding domain of GAL4 induced transcription from a reporter with mutated TATA minimal promoter in a Ser-249 dependent manner. Moreover, a reporter with the SV40 minimal promoter was inhibited by GAL4-Smad7, and this effect was also dependent on Ser-249 phosphorylation. The amplitude of effects on transcriptional regulation was dependent on cell type. Our results suggest that phosphorylation of Smad7, unlike phosphorylation of the receptor-regulated Smads, does not regulate TGF-beta signaling but rather affects TGF-beta-independent effects of Smad7 on transcriptional regulation. PMID- 11278815 TI - Energetics of target peptide binding by calmodulin reveals different modes of binding. AB - Thermodynamic parameters of interactions of calcium-saturated calmodulin (Ca(2+) CaM) with melittin, C-terminal fragment of melittin, or peptides derived from the CaM binding regions of constitutive (cerebellar) nitric-oxide synthase, cyclic nucleotide phosphodiesterase, calmodulin-dependent protein kinase I, and caldesmon (CaD-A, CaD-A*) have been measured using isothermal titration calorimetry. The peptides could be separated into two groups according to the change in heat capacity upon complex formation, DeltaC(p). The calmodulin dependent protein kinase I, constitutive (cerebellar) nitric-oxide synthase, and melittin peptides have DeltaC(p) values clustered around -3.2 kJ.mol(-1).K(-1), consistent with the formation of a globular CaM-peptide complex in the canonical fashion. In contrast, phosphodiesterase, the C-terminal fragment of melittin, CaD A, and CaD-A* have DeltaC(p) values clustered around -1.6 kJ.mol(-1).K(-1), indicative of interactions between the peptide and mostly one lobe of CaM, probably the C-terminal lobe. It is also shown that the interactions for different peptides with Ca(2+)-CaM can be either enthalpically or entropically driven. The difference in the energetics of peptide/Ca(2+)-CaM complex formation appears to be due to the coupling of peptide/Ca(2+)-CaM complex formation to the coil-helix transition of the peptide. The binding of a helical peptide to Ca(2+) CaM is dominated by favorable entropic effects, which are probably mostly due to hydrophobic interactions between nonpolar groups of the peptide and Ca(2+)-CaM. Applications of these findings to the design of potential CaM inhibitors are discussed. PMID- 11278816 TI - Features of the parkin/ariadne-like ubiquitin ligase, HHARI, that regulate its interaction with the ubiquitin-conjugating enzyme, Ubch7. AB - We recently reported the identification of a RING finger-containing protein, HHARI (human homologue of Drosophila ariadne), which binds to the human ubiquitin conjugating enzyme UbcH7 in vitro. We now demonstrate that HHARI interacts and co localizes with UbcH7 in mammalian cells, particularly in the perinuclear region. We have further defined a minimal interaction region of HHARI comprising residues 186-254, identified individual amino acid residues essential for the interaction, and determined that the distance between the RING1 finger and IBR (in between RING fingers) domains is critical to maintaining binding. We have also established that the RING1 finger of HHARI cannot be substituted for by the highly homologous RING finger domains of either of the ubiquitin-protein ligase components c-CBL or Parkin, despite their similarity in structure and their independent capabilities to bind UbcH7. Furthermore, mutation of the RING1 finger domain of HHARI from a RING-HC to a RING-H2 type abolishes interaction with UbcH7. These studies demonstrate that very subtle changes to the domains that regulate recognition between highly conserved components of the ubiquitin pathway can dramatically affect their ability to interact. PMID- 11278817 TI - Epidermal growth factor induction of apolipoprotein A-I is mediated by the Ras MAP kinase cascade and Sp1. AB - Insulin induces apolipoprotein A-I, apoA-I gene transcription via a membrane receptor with intrinsic tyrosine kinase activity. This finding prompted us to ask whether the gene is stimulated by epidermal growth factor (EGF), EGF a peptide hormone that binds to another member of the receptor superfamily with tyrosine kinase activity. Our data showed that like insulin, EGF increased abundance of apoA-I protein and transcription of the gene in human hepatoma, Hep G2 cells. The effects of both hormones appeared direct because their induction of apoA-I gene transcription was not affected by the protein synthesis inhibitor, cycloheximide. Although both insulin and EGF stimulate apoA-I expression, each hormone binds to a distinct membrane receptor thus suggesting differential intracellular signaling. Therefore, we used a panel of inhibitors to define the pathway(s) that mediate the actions of these hormones. Whereas, the actions of EGF required only the Ras-mitogen-activated protein, MAP kinase, those of insulin were mediated by equal participation of both the Ras-MAP kinase and protein kinase C, PKC cascades. Despite differences in signaling pathways triggered by each hormone receptor, the activation of apoA-I transcription required the participation of a single transcription factor, Sp1. Furthermore, EGF induction of transcription was attenuated by mutating the MAP kinase site at amino acid, Thr(266) rendering Sp1 phosphorylation deficient. In summary, EGF stimulation of apoA-I expression is mediated solely by the Ras-MAP kinase cascade and enhanced activity of this pathway requires Sp1 with an intact phosphorylation site at Thr(266). However, insulin induction of this gene is different and requires both Ras-MAP kinase and PKC pathways but their actions are also mediated by Sp1. PMID- 11278818 TI - Cross-talk between 1,25-dihydroxyvitamin D3 and transforming growth factor-beta signaling requires binding of VDR and Smad3 proteins to their cognate DNA recognition elements. AB - 1,25-Dihydroxyvitamin D(3) (vitamin D) and transforming growth factor-beta (TGF beta) regulate diverse biological processes including cell proliferation and differentiation through modulation of the expression of target genes. Members of the Smad family of proteins function as effectors of TGF-beta signaling pathways whereas the vitamin D receptor (VDR) confers vitamin D signaling. We investigated the molecular mechanisms by which TGF-beta and vitamin D signaling pathways interact in the regulation of the human osteocalcin promoter. Synergistic activation of the osteocalcin gene promoter by TGF-beta and vitamin D was observed in transient transfection experiments. However, in contrast to a previous report by Yanagisawa, J., Yanagi, Y., Masuhiro, Y., Suzawa, M., Watanabe, M., Kashiwagi, K., Toriyabe, T., Kawabata, M., Miyazono, K., and Kato, S. (1999) Science, 283, 1317-1321, synergistic activation was not detectable when the osteocalcin vitamin D response element (VDRE) alone was linked to a heterologous promoter. Inclusion of the Smad binding elements (SBEs) with the VDRE in the heterologous promoter restored synergistic activation. Furthermore, this synergy was dependent on the spacing between VDRE and SBEs. The Smad3-Smad4 heterodimer was found to bind in gel shift assay to two distinct DNA segments of the osteocalcin promoter: -1030 to -989 (SBE3) and -418 to -349 (SBE1). Deletion of SBE1, which is proximal to the VDRE, but not the distal SBE3 in this promoter reporter abolished TGF-beta responsiveness and eliminated synergistic co activation with vitamin D. Thus the molecular mechanism, whereby Smad3 and VDR mediate cross-talk between the TGF-beta and vitamin D signaling pathways, requires both a VDRE and a SBE located in close proximity to the target promoter. PMID- 11278819 TI - The zinc finger transcription factor, MOK2, negatively modulates expression of the interphotoreceptor retinoid-binding protein gene, IRBP. AB - The human and murine MOK2 orthologue genes encode Kruppel/TFIIIA-related zinc finger proteins, which are factors able to recognize both DNA and RNA through their zinc finger motifs. MOK2 proteins have been shown to bind to the same 18 base pair (bp)-specific sequence in duplex DNA. This MOK2-binding site was found within introns 7 and 2 of human PAX3 and interphotoreceptor retinoid-binding protein (IRBP) genes, respectively. As these two genes are expressed in the brain as MOK2, we have suggested that PAX3 and IRBP genes are two potentially important target genes for the MOK2 protein. In this study, we focused our attention on IRBP as a potential MOK2 target gene. Sequence comparison and binding studies of the 18-bp MOK2-binding sites present in intron 2 of human, bovine, and mouse IRBP genes show that the 3'-half sequence is the essential core element for MOK2 binding. Very interestingly, 8-bp of this core sequence are found in a reverse orientation, in the IRBP promoter. We demonstrate that MOK2 can bind to the 8-bp sequence present in the IRBP promoter and repress its transcription when transiently overexpressed in retinoblastoma Weri-RB1 cells. In the IRBP promoter, it appears that the TAAAGGCT MOK2-binding site overlaps with the photoreceptor specific CRX-binding element. We suggest that MOK2 represses transcription by competing with the cone-rod homeobox protein (CRX) for DNA binding, thereby decreasing transcriptional activation by CRX. Furthermore, we show that Mok2 expression in the developing mouse and in the adult retina seems to be concordant with IRBP expression. PMID- 11278820 TI - The beta ' subunit of Escherichia coli RNA polymerase is not required for interaction with initiating nucleotide but is necessary for interaction with rifampicin. AB - Using a modification of a highly selective affinity labeling protocol, we demonstrated that the alpha(2)beta subassembly of Escherichia coli RNA polymerase efficiently and specifically interacts with the initiating purine nucleotide. Isolated beta is also active in this reaction. In contrast, neither beta nor alpha(2)beta is able to interact with a chimeric molecule composed of rifampicin attached to an initiation substrate. Based on these results, we conclude that the RNA polymerase initiation site, specific for purine nucleotides, which ultimately become the 5'-end of the transcript, is essentially complete in the absence of the largest subunit, beta'. However, the rifampicin binding center is formed only in the alpha(2)betabeta' core enzyme. We interpret our results in light of the high resolution structure of core RNA polymerase from Thermus aquaticus. PMID- 11278821 TI - The kinetic pathway of RNA binding to the Escherichia coli transcription termination factor Rho. AB - The Escherichia coli transcription termination factor Rho is structurally and functionally homologous to hexameric helicases that assemble into ring structures. Using stopped-flow fluorescence and presteady-state ATPase kinetics, we have determined the kinetic pathway of poly(C) RNA binding to Rho hexamer, both in the presence and in absence of ATP. These studies indicate a four-step sequential mechanism of RNA binding and reveal the respective roles of the primary and secondary RNA binding sites in initiation and ATPase activation of Rho. The primary RNA binding sites of Rho hexamer interact with poly(C) RNA at a diffusion-limited rate constant close to 8 x 10(8) m(-1) s(-1), resulting in the Rho-RNA species PR1, which subsequently isomerizes to PR2 with a rate constant 21 s(-1). The PR2 isomerizes to PR3 with a rate constant of 32 s(-1) in the presence of ATP, and the formation of PR4 from PR3 results in a species that is fully competent in hydrolyzing ATP at the RNA-stimulated rate. The PR3 to PR4 isomerization occurs at a relatively slow rate of 4.1 s(-1); thus, the presteady state ATPase kinetics show a distinct lag due to the slow initiation step. The interactions of the RNA with the primary sites trigger ring opening, and we propose that during the last two steps, the RNA migrates into the central channel and interacts with the secondary sites, resulting in the activation of the ATPase activity. The primary RNA binding sites, in addition to promoting sequence specific initiation, kinetically facilitate loading of the RNA into the secondary sites, which are relatively inaccessible, since they are present in the central channel. These studies reveal common features used by hexameric helicases to bind nucleic acids in an efficient and specific manner. PMID- 11278822 TI - The serine/threonine kinase PAK4 prevents caspase activation and protects cells from apoptosis. AB - The serine/threonine kinase PAK4 was identified first as an effector molecule for the Rho GTPase Cdc42. PAK4 differs from other members of the PAK family both in sequence and function. Previously we have shown that an important function of this kinase is to mediate the induction of filopodia in response to activated Cdc42. Studies with a constitutively active PAK4 mutant have shown that it also has a role in promoting anchorage-independent growth, an important hallmark of oncogenic transformation. Here we show that another function of PAK4 is to protect cells against apoptotic cell death. Expression of wild-type or constitutively active PAK4 delays the onset of apoptosis in response to tumor necrosis factor alpha stimulation, UV irradiation, and serum starvation. Consistent with an antiapoptotic function, expression of PAK4 leads to an increase in phosphorylation of the proapoptotic protein Bad and an inhibition of caspase activation. PMID- 11278823 TI - A proteolytic NH2-terminal truncation of cardiac troponin I that is up-regulated in simulated microgravity. AB - In a tail suspension rat model, we investigated changes in myofilament protein during cardiac adaptation in simulated microgravity. Contractile force and velocity of cardiac muscle were decreased in the tail suspension rats as compared with the control. Ca(2+)-dependent actomyosin ATPase activity was also decreased; however, sensitivity of cardiac muscle to Ca(2+) activation was unchanged. There was no change in expression of myosin heavy chain, tropomyosin, troponin T, or troponin I isoforms in hearts of tail suspension rats. A novel finding is a fragment of cardiac troponin I (cTnI) that had increased amounts in the heart of tail suspension rats. Binding of this cTnI fragment by a monoclonal antibody that specifically recognizes the COOH terminus indicates an intact COOH terminus. NH(2)-terminal sequence analysis of the cTnI fragment revealed truncations primarily of amino acids 1-26 and 1-27 and smaller amounts of 1-30, including Ser(23) and Ser(24), which are substrates of protein kinase A phosphorylation. This cTnI fragment is present in normal cardiac muscle and incorporated into myofibrils, indicating a role in regulating contractility. This proteolytic modification of cTnI up-regulated during simulated microgravity suggests a potential role of the NH(2)-terminal segment of cTnI in functional adaptations of cardiac muscle. PMID- 11278824 TI - Inhibition of gap-junctional communication induces the trans-differentiation of osteoblasts to an adipocytic phenotype in vitro. AB - Osteoblasts and adipocytes are thought to differentiate from a common stromal progenitor cell. These two phenotypically mature cell types show a high degree of plasticity, which can be observed when cells are grown under specific culture conditions. Gap junctions are abundant among osteoblastic cells in vivo and in vitro, whereas they are down-regulated during adipogenesis. Gap junctional communication (GJC) modulates the expression of genes associated with the mature osteoblastic phenotype. Inhibition of GJC utilizing 18-alpha-glycyrrhetinic acid (AGRA) blocks the maturation of pre-osteoblastic cells in vitro. Moreover, cytoplasmic lipid droplets are detectable at the end of the culture period, suggesting that GJC inhibition may favor an adipocytic phenotype. We used several human osteoblastic cell lines, as well as bone-derived primary osteoblastic cells, to show that confluent cultures of human osteoblastic cells grown under osteogenic conditions developed an adipocytic phenotype after 3 days of complete inhibition of GJC using AGRA or oleamide, two dissimilar nontoxic reversible inhibitors. Development of an adipogenic phenotype was confirmed by the accumulation of triglyceride droplets and the increase in mRNA expression of the adipocytic markers peroxisome proliferator-activated receptor gamma2 and lipoprotein lipase. Glycyrrhizic acid, a noninhibitory AGRA analog, or alpha bromopalmitate, a nondegradable fatty acid, had no effect. Modulation of skeletal GJC may represent a new pharmacological target by which inhibition of marrow adipogenesis can take place with the parallel enhancement of osteoblastogenesis, thus providing a novel therapeutic approach to the treatment of human age-related osteopenic diseases and postmenopausal osteoporosis. PMID- 11278826 TI - Expression of Semliki Forest virus E1 protein in Escherichia coli. Low pH-induced pore formation. AB - Exposure of Semliki Forest virus 1 to mildly acidic conditions results in conformational changes of the viral spike proteins, which in turn leads to a pore formation across its membrane. The ability to form a pore has been ascribed to the ectodomain of the Semliki Forest virus (SFV) E1 spike protein. To elucidate whether the E1 protein per se is sufficient for low pH-dependent pore formation, we expressed E1 in Escherichia coli in an inducible manner using the pET11c expression system. The data obtained clearly showed that the E1 protein was expressed in the bacterial cell membrane and that exposure of E. coli expressing the SFV E1 protein to low pH (<6.2) resulted in a permeability change of the membrane. Thus, we conclude that the E1 protein of SFV per se is sufficient to promote pore formation under mildly acidic conditions. PMID- 11278825 TI - Polyamine enhancement of the synthesis of adenylate cyclase at the translational level and the consequential stimulation of the synthesis of the RNA polymerase sigma 28 subunit. AB - The effects of polyamines on the synthesis of various final sigma subunits of RNA polymerase were studied using Western blot analysis. Synthesis of final sigma(28) was stimulated 4.0-fold and that of final sigma(38) was stimulated 2.3-fold by polyamines, whereas synthesis of other final sigma subunits was not influenced by polyamines. Stimulation of final sigma(28) synthesis was due to an increase in the level of cAMP, which occurred through polyamine stimulation of the synthesis of adenylate cyclase at the level of translation. Polyamines were found to increase the translation of adenylate cyclase mRNA by facilitating the UUG codon dependent initiation. Analysis of RNA secondary structure suggests that exposure of the Shine-Dalgarno sequence of mRNA is a prerequisite for polyamine stimulation of the UUG codon-dependent initiation. PMID- 11278827 TI - Asialoglycoprotein receptor deficiency in mice lacking the major receptor subunit. Its obligate requirement for the stable expression of oligomeric receptor. AB - The asialoglycoprotein receptor is an abundant hetero-oligomeric endocytic receptor that is predominantly expressed on the sinusoidal surface of the hepatocytes. A number of physiological and pathophysiological functions have been ascribed to this hepatic lectin (HL), the removal of desialylated serum glycoproteins and apoptotic cells, clearance of lipoproteins, and the sites of entry for hepatotropic viruses. The assembly of two homologous subunits, HL-1 and HL-2, is required to form functional, high affinity receptors on the cell surface. However, the importance of the individual subunits for receptor transport to the cell surface is controversial. We have previously generated HL-2 deficient mice and showed that the expression of HL-1 was significantly reduced, and the functional activity as the asialoglycoprotein receptor was virtually eliminated. However, we failed to detect phenotypic abnormalities. To explore the significance of the major HL-1 subunit for receptor expression and function in vivo, we have disrupted the HL-1 gene in mice. Homozygous HL-1-deficient animals are superficially normal. HL-2 expression in the liver is virtually abrogated, indicating that HL-1 is strictly required for the stable expression of HL-2. Although these mice are almost unable to clear asialo-orosomucoid, a high affinity ligand for asialoglycoprotein receptor, they do not accumulate desialylated glycoproteins or lipoproteins in the plasma. PMID- 11278828 TI - Mitochondrial expression and function of GAS-1 in Caenorhabditis elegans. AB - A mutation in the gene gas-1 alters sensitivity to volatile anesthetics, fecundity, and life span in the nematode Caenorhabditis elegans. gas-1 encodes a close homologue of the 49-kDa iron protein subunit of Complex I of the mitochondrial electron transport chain from bovine heart. gas-1 is widely expressed in the nematode neuromuscular system and in a subcellular pattern consistent with that of a mitochondrial protein. Pharmacological studies indicate that gas-1 functions partially via presynaptic effects. In addition, a mutation in the gas-1 gene profoundly decreases Complex I-dependent metabolism in mitochondria as measured by rates of both oxidative phosphorylation and electron transport. An increase in Complex II-dependent metabolism also is seen in mitochondria from gas-1 animals. There is no apparent alteration in physical structure in mitochondria from gas-1 nematodes compared with those from wild type. These data indicate that gas-1 is the major 49-kDa protein of complex I and that the GAS-1 protein is critical to mitochondrial function in C. elegans. They also reveal the importance of mitochondrial function in determining not only aging and life span, but also anesthetic sensitivity, in this model organism. PMID- 11278829 TI - A quantitative molecular model for modulation of mammalian translation by the eIF4E-binding protein 1. AB - Translation initiation is a key point of regulation in eukaryotic gene expression. 4E-binding proteins (4E-BPs) inhibit initiation by blocking the association of eIF4E with eIF4G, two integral components of the mRNA cap-binding complex. Phosphorylation of 4E-BP1 reduces its ability to bind to eIF4E and thereby to compete with eIF4G. A novel combination of biophysical and biochemical tools was used to measure the impact of phosphorylation and acidic side chain substitution at each potentially modulatory site in 4E-BP1. For each individual site, we have analyzed the effects of modification on eIF4E binding using affinity chromatography and surface plasmon resonance analysis, and on the regulatory function of the 4E-BP1 protein using a yeast in vivo model system and a mammalian in vitro translation assay. We find that modifications at the two sites immediately flanking the eIF4E-binding domain, Thr(46) and Ser(65), consistently have the most significant effects, and that phosphorylation of Ser(65) causes the greatest reduction in binding affinity. These results establish a quantitative framework that should contribute to understanding of the molecular interactions underlying 4E-BP1-mediated translational regulation. PMID- 11278830 TI - Tropomyosin-troponin regulation of actin does not involve subdomain 2 motions. AB - Dynamic properties of F-actin structure prompted suggestions (Squire, J. M., and Morris, E. P. (1998) FASEB J. 12, 761-771) that actin subdomain 2 movements play a role in thin-filament regulation. Using fluorescently labeled yeast actin mutants Q41C, Q41C/C374S, and D51C/C374S and azidonitrophenyl putrescine (ANP) Gln(41)-labeled alpha-actin, we monitored regulation-linked changes in subdomain 2. These actins had fully regulated acto-S1 ATPase activities, and emission spectra of regulated Q41C(AEDANS)/C374S and D51C(AEDANS)/C374S filaments did not reveal any calcium-dependent changes. Fluorescence energy transfer in these F actins mostly occurred from Trp(340) and Trp(356) to 5 (2((acetyl)amino)ethyl)amino-naphthalene-1-sulfonate (AEDANS)-labeled Cys(41) or Cys(51) of adjacent same strand protomers. Our results show that fluorescence energy transfer between these residues is similar in the mostly blocked (-Ca(2+)) and closed (+Ca(2+)) states. Ca(2+) also had no effect on the excimer band in the pyrene-labeled Q41C-regulated actin, indicating virtually no change in the overlap of pyrenes on Cys(41) and Cys(374). ANP quenching of rhodamine phalloidin fluorescence showed that neither Ca(2+) nor S1 binding to regulated alpha-actin affects the phalloidin-probe distance. Taken together, our results indicate that transitions between the blocked, closed, and open regulatory states involve no significant subdomain 2 movements, and, since the cross-linked alpha-actin remains fully regulated, that subdomain 2 motions are not essential for actin regulation. PMID- 11278831 TI - Uncoupling of 3'-phosphatase and 5'-kinase functions in budding yeast. Characterization of Saccharomyces cerevisiae DNA 3'-phosphatase (TPP1). AB - Polynucleotide kinase is a bifunctional enzyme containing both DNA 3'-phosphatase and 5'-kinase activities seemingly suited to the coupled repair of single-strand nicks in which the phosphate has remained with the 3'-base. We show that the yeast Saccharomyces cerevisiae is able to repair transformed dephosphorylated linear plasmids by non-homologous end joining with considerable efficiency independently of the end-processing polymerase Pol4p. Homology searches and biochemical assays did not reveal a 5'-kinase that would account for this repair, however. Instead, open reading frame YMR156C (here named TPP1) is shown to encode only a polynucleotide kinase-type 3'-phosphatase. Tpp1p bears extensive similarity to the ancient L-2-halo-acid dehalogenase and DDDD phosphohydrolase superfamilies, but is specific for double-stranded DNA. It is present at high levels in cell extracts in a functional form and so does not represent a pseudogene. Moreover, the phosphatase-only nature of this gene is shared by Saccharomyces mikatae YMR156C and Arabidopsis thaliana K15M2.3. Repair of 3' phosphate and 5'-hydroxyl lesions is thus uncoupled in budding yeast as compared with metazoans. Repair of transformed dephosphorylated plasmids, and 5'-hydroxyl blocking lesions more generally, likely proceeds by a cycle of base removal and resynthesis. PMID- 11278832 TI - Cloning, expression, and characterization of a human inosine triphosphate pyrophosphatase encoded by the itpa gene. AB - ITP and dITP exist in all cells. dITP is potentially mutagenic, and the levels of these nucleotides are controlled by inosine triphosphate pyrophosphatase (EC ). Here we report the cloning, expression, and characterization of a 21.5-kDa human inosine triphosphate pyrophosphatase (hITPase), an enzyme whose activity has been reported in many animal tissues and studied in populations but whose protein sequence has not been determined before. At the optimal pH of 10.0, recombinant hITPase hydrolyzed ITP, dITP, and xanthosine 5'-triphosphate to their respective monophosphates whereas activity with other nucleoside triphosphates was low. K(m) values for ITP, dITP, and xanthosine 5'-triphosphate were 0.51, 0.31, and 0.57 mm, respectively, and k(cat) values were 580, 360, and 640 s(-1), respectively. A divalent cation was absolutely required for activity. The gene encoding the hITPase cDNA sequence was localized by radiation hybrid mapping to chromosome 20p in the interval D20S113-D20S97, the same interval in which the ITPA inosine triphosphatase gene was previously localized. A BLAST search revealed the existence of many similar sequences in organisms ranging from bacteria to mammals. The function of this ubiquitous protein family is proposed to be the elimination of minor potentially mutagenic or clastogenic purine nucleoside triphosphates from the cell. PMID- 11278833 TI - Targeting of tumor cells by cell surface urokinase plasminogen activator dependent anthrax toxin. AB - Urokinase plasminogen activator receptor (uPAR) binds pro-urokinase plasminogen activator (pro-uPA) and thereby localizes it near plasminogen, causing the generation of active uPA and plasmin on the cell surface. uPAR and uPA are overexpressed in a variety of human tumors and tumor cell lines, and expression of uPAR and uPA is highly correlated to tumor invasion and metastasis. To exploit these characteristics in the design of tumor cell-selective cytotoxins, we constructed mutated anthrax toxin-protective antigen (PrAg) proteins in which the furin cleavage site is replaced by sequences cleaved specifically by uPA. These uPA-targeted PrAg proteins were activated selectively on the surface of uPAR expressing tumor cells in the presence of pro-uPA and plasminogen. The activated PrAg proteins caused internalization of a recombinant cytotoxin, FP59, consisting of anthrax toxin lethal factor residues 1-254 fused to the ADP-ribosylation domain of Pseudomonas exotoxin A, thereby killing the uPAR-expressing tumor cells. The activation and cytotoxicity of these uPA-targeted PrAg proteins were strictly dependent on the integrity of the tumor cell surface-associated plasminogen activation system. We also constructed a mutated PrAg protein that selectively killed tissue plasminogen activator-expressing cells. These mutated PrAg proteins may be useful as new therapeutic agents for cancer treatment. PMID- 11278834 TI - Stimulation of nuclear export and inhibition of nuclear import by a Ran mutant deficient in binding to Ran-binding protein 1. AB - Receptor-mediated nucleocytoplasmic transport is dependent on the GTPase Ran and Ran-binding protein 1 (RanBP1). The acidic C terminus of Ran is required for high affinity interaction between Ran and RanBP1. We found that a novel Ran mutant with four of its five acidic C-terminal amino acids modified to alanine (RanC4A) has an approximately 20-fold reduced affinity for RanBP1. We investigated the effects of RanC4A on nuclear import and export in permeabilized HeLa cells. Although RanC4A promotes accumulation of the nuclear export receptor CRM1 at the cytoplasmic nucleoporin Nup214, it strongly stimulates nuclear export of GFP NFAT. Since RanC4A exhibits an elevated affinity for CRM1 and other nuclear transport receptors, this suggests that formation of the export complex containing CRM1, Ran-GTP, and substrate is a rate-limiting step in export, not release from Nup214. Conversely, importin alpha/beta-dependent nuclear import of bovine serum albumin, coupled to a classical nuclear localization sequence is strongly inhibited by RanC4A. Inhibition can be reversed by additional importin alpha, which promotes the formation of an importin alpha/beta complex. These results provide physiological evidence that release of Ran-GTP from importin beta by RanBP1 and importin alpha is critical for the recycling of importin beta to a transport-competent state. PMID- 11278835 TI - Inhibition of insulin-induced activation of Akt by a kinase-deficient mutant of the epsilon isozyme of protein kinase C. AB - Akt, also known as protein kinase B, is a protein-serine/threonine kinase that is activated by growth factors in a phosphoinositide (PI) 3-kinase-dependent manner. Although Akt mediates a variety of biological activities, the mechanisms by which its activity is regulated remain unclear. The potential role of the epsilon isozyme of protein kinase C (PKC) in the activation of Akt induced by insulin has now been examined. Expression of a kinase-deficient mutant of PKCepsilon (epsilonKD), but not that of wild-type PKCepsilon or of kinase-deficient mutants of PKCalpha or PKClambda, with the use of adenovirus-mediated gene transfer inhibited the phosphorylation and activation of Akt induced by insulin in Chinese hamster ovary cells or L6 myotubes. Whereas the epsilonKD mutant did not affect insulin stimulation of PI 3-kinase activity, the phosphorylation and activation of Akt induced by a constitutively active mutant of PI 3-kinase were inhibited by epsilonKD, suggesting that epsilonKD affects insulin signaling downstream of PI 3 kinase. PDK1 (3'-phosphoinositide-dependent kinase 1) is thought to participate in Akt activation. Overexpression of PDK1 with the use of an adenovirus vector induced the phosphorylation and activation of Akt; epsilonKD inhibited, whereas wild-type PKCepsilon had no effect on, these actions of PDK1. These results suggest that epsilonKD inhibits the insulin-induced phosphorylation and activation of Akt by interfering with the ability of PDK1 to phosphorylate Akt. PMID- 11278836 TI - Regulation of the level of Vesl-1S/Homer-1a proteins by ubiquitin-proteasome proteolytic systems. AB - The vesl-1S/homer-1a gene is up-regulated during seizure and long term potentiation. Other members of the Vesl family, Vesl-1L, -2, and -3, are constitutively expressed in the brain. We examined the regulatory mechanisms governing the expression level of Vesl-1S protein, either an exogenously introduced one in COS7 or human embryonic kidney 293T cells or an endogenous one in rat brain neurons in cultures. In both cases, application of proteasome inhibitors increased the amount of Vesl-1S protein but not that of Vesl-1L, -2, or -3 protein. Deletion analyses revealed that the C-terminal 11-amino acid region was responsible for the proteolysis of Vesl-1S by proteasomes. Application of proteasome inhibitors promoted ubiquitination of Vesl-1S protein but not that of the Vesl-1S deletion mutant, which evaded proteasome-mediated degradation. These results indicate that ubiquitin-proteasome systems are involved in the regulation of the expression level of Vesl-1S protein. PMID- 11278837 TI - The N-terminal helix of Xenopus cyclins A and B contributes to binding specificity of the cyclin-CDK complex. AB - Mitotic cyclins A and B contain a conserved N-terminal helix upstream of the cyclin box fold that contributes to a significant interface between cyclin and cyclin-dependent kinase (CDK). To address its contribution on cyclin-CDK interaction, we have constructed mutants in conserved residues of the N-terminal helix of Xenopus cyclins B2 and A1. The mutants showed altered binding affinities to Cdc2 and/or Cdk2. We also screened for mutations in the C-terminal lobe of CDK that exhibited different binding affinities for the cyclin-CDK complex. These mutations were at residues that interact with the cyclin N-terminal helix motif. The cyclin N-terminal helix mutations have a significant effect on the interaction between the cyclin-CDK complex and specific substrates, Xenopus Cdc6 and Cdc25C. These results suggest that the N-terminal helix of mitotic cyclins is required for specific interactions with CDKs and that to interact with CDK, specific substrates Cdc6 and Cdc25C require the CDK to be associated with a cyclin. The interaction between the cyclin N-terminal helix and the CDK C terminal lobe may contribute to binding specificity of the cyclin-CDK complex. PMID- 11278838 TI - Vitamin C inhibits the enzymatic activity of Streptococcus pneumoniae hyaluronate lyase. AB - Enzyme activity measurement showed that L-ascorbic acid (vitamin C (Vc)) competitively inhibits the hyaluronan degradation by Streptococcus pneumoniae hyaluronate lyase. The complex crystal structure of this enzyme with Vc was determined at 2.0 A resolution. One Vc molecule was found to bind to the active site of the enzyme. The Vc carboxyl group provides the negative charges that lead the molecule into the highly positively charged cleft of the enzyme. The Vc ring system forms hydrophobic interactions with the side chain of Trp-292, which is one of the aromatic patch residues of this enzyme responsible for the selection of the cleavage sites on the substrate chain. The binding of Vc inhibits the substrate binding at hyaluronan 1, 2, and 3 (HA1, HA2, and HA3) catalytic positions. The high concentration of Vc in human tissues probably provides a low level of natural resistance to the pneumococcal invasion. This is the first time that Vc the direct inhibition on the bacterial "spreading factor" was reported, and Vc is also the first chemical that has been shown experimentally to have an inhibitory effect on bacterial hyaluronate lyase. These studies also highlight the possible structural requirement for the design of a stronger inhibitor of bacterial hyaluronate lyase. PMID- 11278839 TI - Rab3a binding and secretion-enhancing domains in Rim1 are separate and unique. Studies in adrenal chromaffin cells. AB - Rim1 was identified in brain by its ability to bind Rab3a-GTP and has been postulated to be a Rab3a effector protein. Like Rabphilin3, it modulates secretion and contains a zinc finger and two C2 domains. We have investigated the structural basis for the ability of Rim1 to bind Rab3a-GTP and to stimulate exocytosis in chromaffin cells. Both full-length and N-terminal Rim1 enhance secretion 40-50% in both intact and permeabilized cells. The abilities of Rim1 to enhance secretion and to bind Rab3a-GTP reside on distinct and relatively small domains that act independently. A approximately 30-amino acid sequence immediately N-terminal of the zinc finger constitutes the minimal Rab3a-GTP binding domain. This short sequence is not found in Rabphilin3 and is entirely different from the zinc finger and flanking regions of Rabphilin3 that bind Rab3a GTP. The zinc finger domain in Rim1 is unnecessary for Rab3a-GTP binding but, alone, enhances secretion. An analysis of the characteristics of the enhancement of secretion in permeabilized chromaffin cells indicates that N-terminal Rim1 does not alter the sensitivity of secretion to Ca(2+) but, instead, increases the rate of ATP-dependent priming of secretion. PMID- 11278840 TI - Helical stalk segments S4 and S5 of the plasma membrane H+-ATPase from Saccharomyces cerevisiae are optimized to impact catalytic site environment. AB - The stalk segments of P-type ion-translocating enzymes are presumed to play important roles in energy coupling. In this work, stalk segments S4 and S5 of the yeast H(+)-ATPase were examined for helical character, optimal length, and segment orientation by a combination of proline substitution, insertion/deletion mutagenesis, and second-site suppressor analyses. The substitution of various residues for helix-disrupting proline in both S4 (L353P,L353G; A354P; and G371P) and S5 (D676P and I684P) resulted in highly defective or inactive enzymes supporting the importance of helical character and/or the maintenance of essential interactions. The contiguous helical nature of transmembrane segment M5 and stalk element S5 was explored and found to be favorable, although not essential. The deletion or addition of one or more amino acids at positions Ala(354) in S4 and Asp(676) in S5, which were intended to either rotate helical faces or extend/reduce the length of helical segments, resulted in enzyme destabilization that abolished most enzyme assembly. Second-site suppressor mutations were obtained to primary site mutations G371A (S4) and D676G (S5) and were analyzed with a molecular structure model of the H(+)-ATPase. Primary site mutations were predicted to alter the site of phosphorylation either directly or indirectly. The suppressor mutations either directly changed packing around the primary site or altered the environment of the site of phosphorylation. Overall, these data support the view that stalk segments S4 and S5 of the H(+)-ATPase are helical elements that are optimized for length and interactions with other stalk elements and can influence the phosphorylation domain. PMID- 11278841 TI - Phosphorylation regulates intracellular trafficking of beta-secretase. AB - beta-Secretase (BACE) is a transmembrane aspartyl protease, which generates the N terminus of Alzheimer's disease amyloid beta-peptide. Here, we report that BACE can be phosphorylated within its cytoplasmic domain at serine residue 498 by casein kinase 1. Phosphorylation exclusively occurs after full maturation of BACE by propeptide cleavage and complex N-glycosylation. Phosphorylation/dephosphorylation affects the subcellular localization of BACE. BACE wild type and an S498D mutant that mimics phosphorylated BACE are predominantly located within juxtanuclear Golgi compartments and endosomes, whereas nonphosphorylatable BACE S498A accumulates in peripheral EEA1-positive endosomes. Antibody uptake assays revealed that reinternalization of BACE from the cell surface is independent of its phosphorylation state. After reinternalization, BACE wild type as well as BACE S498D are efficiently retrieved from early endosomal compartments and further targeted to later endosomal compartments and/or the trans-Golgi network. In contrast, nonphosphorylatable BACE S498A is retained within early endosomes. Our results therefore demonstrate regulated trafficking of BACE within the secretory and endocytic pathway. PMID- 11278842 TI - Two different combinations of RNA-binding domains determine the RNA binding specificity of nucleolin. AB - Nucleolin is an abundant nucleolar protein involved in several steps of ribosome biogenesis. The protein is highly conserved through evolution and possesses four RNA-binding domains (RBD), which are likely to determine its RNA binding specificity. Previous studies have shown that nucleolin interacts with two different RNA targets. The first is a small stem-loop structure, the nucleolin recognition element (NRE), found all along the pre-ribosomal RNA. The second is a short single-stranded RNA sequence, the evolutionary conserved motif (ECM), located five nucleotides downstream of the first processing site in the pre ribosomal RNA 5' external transcribed spacer. Biochemical, genetic, and structural studies have shown that the first two RBD of nucleolin are necessary and sufficient for the specific interaction of nucleolin with the NRE motif. In this work, we have studied the interaction of nucleolin with the ECM sequence. Deletion and mutational analyses showed that all four RBDs of hamster nucleolin were required for the interaction with the ECM sequence. This RNA binding specificity is conserved between hamster and Xenopus laevis, whereas the Xenopus protein does not interact with the NRE. Nucleolin is the first example of a protein that requires four RBDs for its interaction with an RNA target, demonstrating that a single protein can use different combinations of RBD to interact specifically with several RNA sequences. PMID- 11278843 TI - Phosphorylation of a synaptic vesicle-associated protein by an inositol hexakisphosphate-regulated protein kinase. AB - Despite the fact that inositol hexakisphosphate (InsP(6)) is the most abundant inositol metabolite in cells, its cellular function has remained an enigma. In the present study, we present the first evidence of a protein kinase identified in rat cerebral cortex/hippocampus that is activated by InsP(6). The substrate for the InsP(6)-regulated protein kinase was found to be the synaptic vesicle associated protein, pacsin/syndapin I. This brain-specific protein, which is highly enriched at nerve terminals, is proposed to act as a molecular link coupling components of the synaptic vesicle endocytic machinery to the cytoskeleton. We show here that the association between pacsin/syndapin I and dynamin I can be increased by InsP(6)-dependent phosphorylation of pacsin/syndapin I. These data provide a model by which InsP(6)-dependent phosphorylation regulates synaptic vesicle recycling by increasing the interaction between endocytic proteins at the synapse. PMID- 11278844 TI - The cost of exposing a hydrophobic loop and implications for the functional role of 4.5 S RNA in the Escherichia coli signal recognition particle. AB - The signal recognition particle (SRP) is an RNA-protein complex that directs ribosomes to the rough endoplasmic reticulum membrane by binding to targeting signals found on the nascent chain of proteins destined for export to the endoplasmic reticulum. We found evidence from studies with fragments of the protein component of the Escherichia coli SRP that a long hydrophobic loop (the so-called "finger loop") is detrimental to the stability of its signal peptide binding domain, the M domain. This hydrophobic loop is highly conserved and thus may have a critical role in the function of the SRP. Given our previously reported evidence that 4.5 S RNA stabilizes the tertiary fold of the M domain (Zheng, N., and Gierasch, L. M. (1997) Mol. Cell 1, 79-87), we now propose that the functional requirement for 4.5 S RNA resides in its ability to counteract the destabilizing influence of the finger loop. PMID- 11278845 TI - Atp-bound topoisomerase ii as a target for antitumor drugs. AB - Topoisomerase II (TOP2) poisons interfere with the breakage/reunion reaction of TOP2 resulting in DNA cleavage. In the current studies, we show that two different classes (ATP-sensitive and -insensitive) of TOP2 poisons can be identified based on their differential sensitivity to the ATP-bound conformation of TOP2. First, in the presence of 1 mm ATP or the nonhydrolyzable analog adenosine 5'-(beta,gamma-imino)triphosphate, TOP2-mediated DNA cleavage induced by ATP-sensitive TOP2 poisons (e.g. doxorubicin, etoposide, mitoxantrone, and 4' (9-acridinylamino)methanesulfon-m-anisidide) was 30-100-fold stimulated, whereas DNA cleavage induced by ATP-insensitive TOP2 poisons (e.g. amonafide, batracylin, and menadione) was only slightly (less than 3-fold) affected. In addition, ADP was shown to strongly antagonize TOP2-mediated DNA cleavage induced by ATP sensitive but not ATP-insensitive TOP2 poisons. Second, C427A mutant human TOP2alpha, which exhibits reduced ATPase activity, was shown to exhibit cross resistance to all ATP-sensitive but not ATP-insensitive TOP2 poisons. Third, using ciprofloxacin competition assay, TOP2-mediated DNA cleavage induced by ATP sensitive but not ATP-insensitive poisons was shown to be antagonized by ciprofloxacin. These results suggest that ATP-bound TOP2 may be the specific target of ATP-sensitive TOP2 poisons. Using Lac repressor-operator complexes as roadblocks, we show that ATP-bound TOP2 acts as a circular clamp capable of entering DNA ends and sliding on unobstructed duplex DNA. PMID- 11278846 TI - Selective suppression of CCAAT/enhancer-binding protein beta binding and cyclooxygenase-2 promoter activity by sodium salicylate in quiescent human fibroblasts. AB - The anti-inflammatory actions of salicylates cannot be explained by inhibition of cyclooxygenase (COX) activity. This study demonstrates that sodium salicylate at a therapeutic concentration suppressed COX-2 gene transcription induced by phorbol 12-myristate 13-acetate and interleukin 1beta by inhibiting the binding of CCAAT/enhancer-binding protein beta to its promoter region of COX-2. By contrast, salicylate did not inhibit nuclear factor kappaB-dependent COX-2 induction by tumor necrosis factor alpha. The inhibitory effect of sodium salicylate was restricted to serum-deprived quiescent cells. These findings indicate that contrary to the current view that salicylate acts via inhibition of nuclear factor kappaB the pharmacological actions of aspirin and salicylates are mediated by inhibiting CCAAT/enhancer-binding protein beta binding and transactivation. These findings have a major impact on the conceptual understanding of the mechanism of action of salicylates and on new drug discovery and design. PMID- 11278847 TI - Magnesium-dependent association and folding of oligonucleosomes reconstituted with ubiquitinated H2A. AB - The MgCl2-induced folding of defined 12-mer nucleosomal arrays, in which ubiquitinated histone H2A (uH2A) replaced H2A, was analyzed by quantitative agarose gel electrophoresis and analytical centrifugation. Both types of analysis showed that uH2A arrays attained a degree of compaction similar to that of control arrays in 2 mM MgCl2. These results indicate that attachment of ubiquitin to H2A has little effect on the ability of nucleosomal arrays to form higher order folded structures in the ionic conditions tested. In contrast, uH2A arrays were found to oligomerize at lower MgCl2 concentrations than control nucleosomal arrays, suggesting that histone ubiquitination may play a role in nucleosomal fiber association. PMID- 11278848 TI - The p38 mitogen-activated kinase pathway regulates the human interleukin-10 promoter via the activation of Sp1 transcription factor in lipopolysaccharide stimulated human macrophages. AB - Interleukin-10 (IL-10), a pleiotropic cytokine that inhibits inflammatory and cell-mediated immune responses, is produced by a wide variety of cell types including T and B cells and monocytes/macrophages. Regulation of pro- and anti inflammatory cytokines has been suggested to involve distinct signaling pathways. In this study, we investigated the regulation of the human IL-10 (hIL-10) promoter in the human monocytic cell line THP-1 following activation with lipopolysaccharide (LPS). Analysis of hIL-10 promoter sequences revealed that DNA sequences located between base pairs -652 and -571 are necessary for IL-10 transcription. A computer analysis of the promoter sequence between base pairs 652 and -571 revealed the existence of consensus sequences for Sp1, PEA1, YY1, and Epstein-Barr virus-specific nuclear antigen-2 (EBNA-2)-like transcription factors. THP-1 cells transfected with a plasmid containing mutant Sp1 abrogated the promoter activity, whereas plasmids containing the sequences for PEA1, YY1, and EBNA-2-like transcription factors did not influence hIL-10 promoter activity. To understand the events upstream of Sp1 activation, we investigated the role of p38 and extracellular signal-regulated kinase mitogen-activated protein kinases by using their specific inhibitors. SB202190 and SB203580, the p38-specific inhibitors, inhibited LPS-induced IL-10 production. In contrast, PD98059, a specific inhibitor of extracellular signal-regulated kinase kinases, failed to modulate IL-10 production. Furthermore, SB203580 inhibited LPS-induced activation of Sp1, as well as the promoter activity in cells transfected with a plasmid containing the Sp1 consensus sequence. These results suggest that p38 mitogen activated protein kinase regulates LPS-induced activation of Sp1, which in turn regulates transcription of the hIL-10 gene. PMID- 11278849 TI - beta -Amyloid peptide-induced apoptosis regulated by a novel protein containing a g protein activation module. AB - Degeneration of neurons in Alzheimer's disease is mediated by beta-amyloid peptide by diverse mechanisms, which include a putative apoptotic component stimulated by unidentified signaling events. This report describes a novel beta amyloid peptide-binding protein (denoted BBP) containing a G protein-coupling module. BBP is one member of a family of three proteins containing this conserved structure. The BBP subtype bound human beta-amyloid peptide in vitro with high affinity and specificity. Expression of BBP in cell culture induced caspase dependent vulnerability to beta-amyloid peptide toxicity. Expression of a signaling-deficient dominant negative BBP mutant suppressed sensitivity of human Ntera-2 neurons to beta-amyloid peptide mediated toxicity. These findings suggest that BBP is a target of neurotoxic beta-amyloid peptide and provide new insight into the molecular pathophysiology of Alzheimer's disease. PMID- 11278850 TI - Cytochrome c oxidase-deficient patients have distinct subunit assembly profiles. AB - Cytochrome c oxidase (COX) deficiency is the most common respiratory chain defect in childhood and is clinically heterogeneous. We report a study of six patients with COX deficiencies. Two of the patients had as yet undefined defects, three patients had Surf-1 mutations, and one patient had a 15-base pair deletion in the COX III subunit. We show that quantitative measurements of steady-state levels of subunits by monoclonal antibody reactivity, when used in combination with a discontinuous sucrose gradient methods, provide an improved diagnosis of COX deficiencies by distinguishing between kinetic, stability, and assembly defects. The two mutants of undefined etiology had a full complement of subunits with one stable and the other partially unstable to detergent solubilization. Both are likely to carry mutations in nuclear-encoded subunits of the complex. The three Surf-1 mutants and the COX III mutant each had reduced steady-state levels of subunits but variable associations of the residual subunits. This information, as well as aiding in diagnosis, helps in understanding the genotype-phenotype relationships of COX deficiencies and provides insight into the mechanism of assembly of the enzyme complex. PMID- 11278851 TI - Paclitaxel induces prolonged activation of the Ras/MEK/ERK pathway independently of activating the programmed cell death machinery. AB - Paclitaxel is a widely used chemotherapeutic agent and is known to induce programmed cell death (apoptosis) in a variety of cell types, but the precise underlying mechanisms are poorly understood. To elucidate these mechanisms, we challenged human esophageal squamous cancer cell lines with paclitaxel and investigated its effects upon signal transduction pathways. Physiologically relevant concentrations of paclitaxel (1-1,000 nm) induced apoptosis. All three mitogen-activated protein kinase (MAPK) family members, c-Jun N-terminal kinase (JNK), p38 MAPK, and extracellular signal-regulated kinase (ERK) were activated upon paclitaxel treatment. Interestingly, JNK activation and p38 MAPK activation were delayed and peaked at 48 h, whereas ERK activity was sustained over 72 h. In addition, Ras activation and MAPK/ERK kinase (MEK) phosphorylation were observed in concordance with ERK activation. While ERK activation was completely ablated by MEK inhibitors, immunoprecipitation and Western blot analysis revealed that neither MEK-1 nor MEK-2 was involved, but instead another member of the MEK family may potentially participate. Although pretreatment with a general caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone rescued the cell death, it did not prevent Ras or ERK activation. Furthermore, inhibition of JNK, p38 MAPK, or MEK did not alter PARP cleavage and the cell death induced by paclitaxel. These results in aggregate suggest that the delayed activation of JNK, p38 MAPK, and ERK was not linked to activation of the cell death machinery. PMID- 11278852 TI - The role of leucine 191 of Escherichia coli uracil DNA glycosylase in the formation of a highly stable complex with the substrate mimic, ugi, and in uracil excision from the synthetic substrates. AB - Uracil DNA glycosylase (UDG), a highly conserved DNA repair enzyme, initiates the uracil excision repair pathway. Ugi, a bacteriophage-encoded peptide, potently inhibits UDGs by serving as a remarkable substrate mimic. Structure determination of UDGs has identified regions important for the exquisite specificity in the detection and removal of uracils from DNA and in their interaction with Ugi. In this study, we carried out mutational analysis of the Escherichia coli UDG at Leu191 within the 187HPSPLS192 motif (DNA intercalation loop). We show that with the decrease in side chain length at position 191, the stability of the UDG-Ugi complexes regresses. Further, while the L191V and L191F mutants were as efficient as the wild type protein, the L191A and L191G mutants retained only 10 and 1% of the enzymatic activity, respectively. Importantly, however, substitution of Leu191 with smaller side chains had no effect on the relative efficiencies of uracil excision from the single-stranded and a corresponding double-stranded substrate. Our results suggest that leucine within the HPSPLS motif is crucial for the uracil excision activity of UDG, and it contributes to the formation of a physiologically irreversible complex with Ugi. We also envisage a role for Leu191 in stabilizing the productive enzyme-substrate complex. PMID- 11278853 TI - JFC1, a novel tandem C2 domain-containing protein associated with the leukocyte NADPH oxidase. AB - We have employed a yeast two-hybrid system to screen a B lymphoblast-derived cDNA library, searching for regulatory components of the NADPH oxidase. Using as bait the C-terminal half of p67(phox), which contains both Src homology 3 domains, we have cloned JFC1, a novel human 62-kDa protein. JFC1 possesses two C2 domains in tandem. The C2A domain shows homology with the C2B domain of synaptotagmins. JFC1 mRNA was abundantly expressed in bone marrow and leukocytes. The expression of JFC1 in neutrophils was restricted to the plasma membrane/secretory vesicle fraction. We confirmed JFC1-p67(phox) association by affinity chromatography. JFC1-containing beads pulled down both p67(phox) and p47(phox) subunits from neutrophil cytosol, but when the recombinant proteins were used, only p67(phox) bound to JFC1, indicating that JFC1 binds to the cytosolic complex via p67(phox) without affecting the interaction between p67(phox) and p47(phox). In contrast to synaptotagmins, JFC1 was unable to bind to inositol 1,3,4,5-tetrakisphosphate but did bind to phosphatidylinositol 3,4,5-trisphosphate and to a lesser extent to phosphatidylinositol 3,4-diphosphate. From the data presented here, it is proposed that JFC1 is acting as an adaptor protein between phosphatidylinositol 3 kinase products and the oxidase cytosolic complex. PMID- 11278854 TI - The recruitment of SOX/OCT complexes and the differential activity of HOXA1 and HOXB1 modulate the Hoxb1 auto-regulatory enhancer function. AB - Regionally restricted expression patterns of Hox genes in developing embryos rely on auto-, cross-, and para-regulatory transcriptional elements. One example is the Hoxb1 auto-regulatory element (b1-ARE), which drives expression of Hoxb1 in the fourth rhombomere of the hindbrain. We previously showed that HOXB1 and PBX1 activate transcription from the b1-ARE by binding to sequences required for the expression of a reporter gene in rhombomere 4 in vivo. We now report that in embryonal carcinoma cells, which retain characteristics of primitive neuroectodermal cells, the b1-ARE displays higher basal and HOX/PBX-induced activities than in other cell backgrounds. We have identified a bipartite-binding site for SOX/OCT heterodimers within the b1-ARE that accounts for its cell context-specific activity and is required for maximal transcriptional activity of HOX/PBX complexes in embryonal carcinoma cells. Furthermore, we found that in an embryonal carcinoma cell background, HOXB1 has a significantly higher transcriptional activity than its paralog HOXA1. We map the determinants for this differential activity within the HOXB1 N-terminal transcriptional activation domain. By using analysis in transgenic and HOXA1 mutant mice, we extended these findings on the differential activities of HOXA1 and HOXB1 in vivo, and we demonstrated that they are important for regulating aspects of HOXB1 expression in the hindbrain. We found that mutation of the SOX/OCT site and targeted inactivation of Hoxa1 both impair the response of the b1-ARE to retinoic acid in transgenic mice. Our results show that Hoxa1 is the primary mediator of the response of b1-ARE to retinoic acid in vivo and that this function is dependent on the binding of SOX/OCT heterodimers to the b1-ARE. These results uncover novel functional differences between Hox paralogs and their modulators. PMID- 11278856 TI - Damaged DNA-binding protein DDB stimulates the excision of cyclobutane pyrimidine dimers in vitro in concert with XPA and replication protein A. AB - Human cells contain a protein that binds to UV-irradiated DNA with high affinity. This protein, damaged DNA-binding protein (DDB), is a heterodimer of two polypeptides, p127 and p48. Recent in vivo studies suggested that DDB is involved in global genome repair of cyclobutane pyrimidine dimers (CPDs), but the mechanism remains unclear. Here, we show that in vitro DDB directly stimulates the excision of CPDs but not (6-4)photoproducts. The excision activity of cell free extracts from Chinese hamster AA8 cell line that lacks DDB activity was increased 3-4-fold by recombinant DDB heterodimer but not p127 subunit alone. Moreover, the addition of XPA or XPA + replication protein A (RPA), which themselves enhanced excision, also enhanced the excision in the presence of DDB. DDB was found to elevate the binding of XPA to damaged DNA and to make a complex with damaged DNA and XPA or XPA + RPA as judged by both electrophoretic mobility shift assays and DNase I protection assays. These results suggest that DDB assists in the recognition of CPDs by core NER factors, possibly through the efficient recruitment of XPA or XPA.RPA, and thus stimulates the excision reaction of CPDs. PMID- 11278855 TI - Involvement of the pro-oncoprotein TLS (translocated in liposarcoma) in nuclear factor-kappa B p65-mediated transcription as a coactivator. AB - In this study, we have demonstrated that translocated in liposarcoma (TLS), also termed FUS, is an interacting molecule of the p65 (RelA) subunit of the transcription factor nuclear factor kappaB (NF-kappaB) using a yeast two-hybrid screen. We confirmed the interaction between TLS and p65 by the pull-down assay in vitro and by a coimmunoprecipitation experiment followed by Western blot of the cultured cell in vivo. TLS was originally identified as part of a fusion protein with CHOP arising from chromosomal translocation in human myxoid liposarcomas. TLS has been shown to be involved in TFIID complex formation and associated with RNA polymerase II. However, the role of TLS in transcriptional regulation has not yet been clearly elucidated. We found that TLS enhanced the NF kappaB-mediated transactivation induced by physiological stimuli such as tumor necrosis factor alpha, interleukin-1beta, and overexpression of NF-kappaB inducing kinase. TLS augmented NF-kappaB-dependent promoter activity of the intercellular adhesion molecule-1 gene and interferon-beta gene. These results suggest that TLS acts as a coactivator of NF-kappaB and plays a pivotal role in the NF-kappaB-mediated transactivation. PMID- 11278857 TI - The role of the Src homology 3-Src homology 2 interface in the regulation of Src kinases. AB - The regulatory fragment of Src kinases, comprising Src homology (SH) 3 and SH2 domains, is responsible for controlled repression of kinase activity. We have used a multidisciplinary approach involving crystallography, NMR, and isothermal titration calorimetry to study the regulatory fragment of Fyn (FynSH32) and its interaction with a physiological activator: a fragment of focal adhesion kinase that contains both phosphotyrosine and polyproline motifs. Although flexible, the preferred disposition of SH3 and SH2 domains in FynSH32 resembles the inactive forms of Hck and Src, differing significantly from LckSH32. This difference, which results from variation in the SH3-SH2 linker sequences, will affect the potential of the regulatory fragments to repress kinase activity. This surprising result implies that the mechanism of repression of Src family members may vary, explaining functional distinctions between Fyn and Lck. The interaction between FynSH32 and focal adhesion kinase is restricted to the canonical SH3 and SH2 binding sites and does not affect the dynamic independence of the two domains. Consequently, the interaction shows no enhancement by an avidity effect. Such an interaction may have evolved to gain specificity through an extended recognition site while maintaining rapid dissociation after signaling. PMID- 11278858 TI - The early interaction of the outer membrane protein phoe with the periplasmic chaperone Skp occurs at the cytoplasmic membrane. AB - Spheroplasts were used to study the early interactions of newly synthesized outer membrane protein PhoE with periplasmic proteins employing a protein cross-linking approach. Newly translocated PhoE protein could be cross-linked to the periplasmic chaperone Skp at the periplasmic side of the inner membrane. To study the timing of this interaction, a PhoE-dihydrofolate reductase hybrid protein was constructed that formed translocation intermediates, which had the PhoE moiety present in the periplasm and the dihydrofolate reductase moiety tightly folded in the cytoplasm. The hybrid protein was found to cross-link to Skp, indicating that PhoE closely interacts with the chaperone when the protein is still in a transmembrane orientation in the translocase. Removal of N-terminal parts of PhoE protein affected Skp binding in a cumulative manner, consistent with the presence of two Skp-binding sites in that region. In contrast, deletion of C-terminal parts resulted in variable interactions with Skp, suggesting that interaction of Skp with the N-terminal region is influenced by parts of the C terminus of PhoE protein. Both the soluble as well as the membrane-associated Skp protein were found to interact with PhoE. The latter form is proposed to be involved in the initial interaction with the N-terminal regions of the outer membrane protein. PMID- 11278859 TI - Decreased expression of specific genes in yeast cells lacking histone H1. AB - Chromatin plays an important role in regulating eukaryotic gene expression. Chromatin is composed of DNA wrapped around a nucleosome core (consisting of two copies of the well conserved histones H2A, H2B, H3, and H4) and a more variable linker histone H1. Various in vitro and in vivo studies have implicated histone H1 as a repressor of gene expression or as an activator, but its exact role is still unclear. Sequencing of the yeast genome has led to the identification of a putative histone H1 gene. Biochemical studies demonstrated that yeast does indeed possess a bona fide histone H1. However, deletion of the unique yeast H1 gene is not associated with any phenotypes, and it was questioned whether it plays any role. To address this issue, we performed whole-genome microarray analysis to identify genes that are affected by H1 removal. Surprisingly, deletion of the gene encoding histone H1 does not result in increased gene expression but rather in a modest reduction. Northern blot analysis of selected genes confirmed the results obtained with the microarray analysis. A similar effect was observed with an integrated lacZ reporter. Thus, our data demonstrate that removal of yeast histone H1 only results in decreased gene expression. PMID- 11278860 TI - Binding of heparin/heparan sulfate to fibroblast growth factor receptor 4. AB - Fibroblast growth factors (FGFs) are heparin-binding polypeptides that affect the growth, differentiation, and migration of many cell types. FGFs signal by binding and activating cell surface FGF receptors (FGFRs) with intracellular tyrosine kinase domains. The signaling involves ligand-induced receptor dimerization and autophosphorylation, followed by downstream transfer of the signal. The sulfated glycosaminoglycans heparin and heparan sulfate bind both FGFs and FGFRs and enhance FGF signaling by mediating complex formation between the growth factor and receptor components. Whereas the heparin/heparan sulfate structures involved in FGF binding have been studied in some detail, little information has been available on saccharide structures mediating binding to FGFRs. We have performed structural characterization of heparin/heparan sulfate oligosaccharides with affinity toward FGFR4. The binding of heparin oligosaccharides to FGFR4 increased with increasing fragment length, the minimal binding domains being contained within eight monosaccharide units. The FGFR4-binding saccharide domains contained both 2-O-sulfated iduronic acid and 6-O-sulfated N-sulfoglucosamine residues, as shown by experiments with selectively desulfated heparin, compositional disaccharide analysis, and a novel exoenzyme-based sequence analysis of heparan sulfate oligosaccharides. Structurally distinct heparan sulfate octasaccharides differed in binding to FGFR4. Sequence analysis suggested that the affinity of the interaction depended on the number of 6-O-sulfate groups but not on their precise location. PMID- 11278861 TI - Interaction sites of the G protein beta subunit with brain G protein-coupled inward rectifier K+ channel. AB - G protein-coupled inward rectifier K(+) channels (GIRK channels) are activated directly by the G protein betagamma subunit. The crystal structure of the G protein betagamma subunits reveals that the beta subunit consists of an N terminal alpha helix followed by a symmetrical seven-bladed propeller structure. Each blade is made up of four antiparallel beta strands. The top surface of the propeller structure interacts with the Galpha subunit. The outer surface of the betagamma torus is largely made from outer beta strands of the propeller. We analyzed the interaction between the beta subunit and brain GIRK channels by mutating the outer surface of the betagamma torus. Mutants of the outer surface of the beta(1) subunit were generated by replacing the sequences at the outer beta strands of each blade with corresponding sequences of the yeast beta subunit, STE4. The mutant beta(1)gamma(2) subunits were expressed in and purified from Sf9 cells. They were applied to inside-out patches of cultured locus coeruleus neurons. The wild type beta(1)gamma(2) induced robust GIRK channel activity with an EC(50) of about 4 nm. Among the eight outer surface mutants tested, blade 1 and blade 2 mutants (D1 and CD2) were far less active than the wild type in stimulating GIRK channels. However, the ability of D1 and CD2 to regulate type I and type II adenylyl cyclases was not very different from that of the wild type beta(1)gamma(2). As to the activities to stimulate phospholipase Cbeta(2), D1 was more potent and CD2 was less potent than the wild type beta(1)gamma(2). Additionally we tested four beta(1) mutants in which mutated residues are located in the top Galpha/beta interacting surface. Among them, mutant W332A showed far less ability than the wild type to activate GIRK channels. These results suggest that the outer surface of blade 1 and blade 2 of the beta subunit might specifically interact with GIRK and that the beta subunit interacts with GIRK both over the outer surface and over the top Galpha interacting surface. PMID- 11278862 TI - From consensus sequence peptide to high affinity ligand, a "library scan" strategy. AB - A wide variety of proteins have been shown to recognize and bind to specific amino acid sequences on other proteins. These sequences can be readily identified using combinatorial peptide libraries. However, peptides containing these preferred sequences ("consensus sequence peptides") typically display only modest affinities for the consensus sequence-binding site on the intact protein. In this report, we describe a parallel synthesis strategy that transforms consensus sequence peptides into high affinity ligands. The work described herein has focused on the Lck SH2 domain, which binds the consensus peptide acetyl-Tyr(P) Glu-Glu-Ile-amide with a K(D) of 1.3 micrometer. We employed a strategy that creates a series of spatially focused libraries that challenge specific subsites on the target protein with a diverse array of functionality. The final lead compound identified in this study displayed a 3300-fold higher affinity for the Lck SH2 domain than the starting consensus sequence peptide. PMID- 11278863 TI - The G protein beta subunit is a determinant in the coupling of Gs to the beta 1 adrenergic and A2a adenosine receptors. AB - The signaling specificity of five purified G protein betagamma dimers, beta(1)gamma(2), beta(2)gamma(2), beta(3)gamma(2), beta(4)gamma(2), and beta(5)gamma(2), was explored by reconstituting them with G(s) alpha and receptors or effectors in the adenylyl cyclase cascade. The ability of the five betagamma dimers to support receptor-alpha-betagamma interactions was examined using membranes expressing the beta(1)-adrenergic or A2a adenosine receptors. These receptors discriminated among the defined heterotrimers based solely on the beta isoform. The beta(4)gamma(2) dimer demonstrated the highest coupling efficiency to either receptor. The beta(5)gamma(2) dimer coupled poorly to each receptor, with EC(50) values 40-200-fold higher than those observed with beta(4)gamma(2). Strikingly, whereas the EC(50) of the beta(1)gamma(2) dimer at the beta(1)-adrenergic receptor was similar to beta(4)gamma(2), its EC(50) was 20 fold higher at the A2a adenosine receptor. Inhibition of adenylyl cyclase type I (AC1) and stimulation of type II (AC2) by the betagamma dimers were measured. betagamma dimers containing Gbeta(1-4) were able to stimulate AC2 similarly, and beta(5)gamma(2) was much less potent. beta(1)gamma(2), beta(2)gamma(2), and beta(4)gamma(2) inhibited AC1 equally; beta(3)gamma(2) was 10-fold less effective, and beta(5)gamma(2) had no effect. These data argue that the beta isoform in the betagamma dimer can determine the specificity of signaling at both receptors and effectors. PMID- 11278864 TI - Role of phosphoinositide 3-kinase in monocyte recruitment under flow conditions. AB - Chemokines such as the monocyte chemol attractant protein-1 (MCP-1) convert monocyte rolling to firm arrest under physiological flow conditions via integrin activation and simultaneously activate phosphoinositide 3-kinase (PI3K). Here we used adenoviral gene transfer and biochemical inhibitors to manipulate PI3K dependent pathways in human monocytes. In in vitro lipid kinase assays from purified human monocytes, we showed that MCP-1 activates the "classical" PI3Kalpha pathway and not PI3Kgamma, a PI3K isoform thought to be activated only by the betagamma complex of heterotrimeric G proteins. The activity of PI3Kalpha in purified human monocytes was evident within 30 s. MCP-1-induced monocyte arrest was significantly inhibited both by wortmannin (n = 4; p < 0.01) and LY294002 (n = 4; p < 0.01) with restoration of the rolling phenotype (p < 0.05 for both inhibitors, compared with rolling of control monocytes after MCP-1 treatment). To test the hypothesis that activation of PI3K is sufficient to induce monocyte adhesion, we transduced the monocytic THP-1 cell line with a recombinant adenovirus (Ad) carrying a constitutively active mutant of PI3K (Ad.BD110). We examined the ability of these cells to adhere to human vascular endothelium (HUVEC) transduced with adenoviruses carrying E-selectin, intercellular adhesion molecule-1 (ICAM-1), and VCAM-1. Under flow conditions, ICAM-1- and VCAM-1-dependent firm adhesion of Ad.BD110-transduced THP-1 cells was enhanced compared with THP-1 cells infected with control Ad (n = 4; p < 0.01 for both). Adhesion augmented by constitutive PI3K activation was entirely abrogated by pretreatment with wortmannin (n = 3; p < 0.01). In contrast, a constitutively active Akt construct had no effect on THP-1 adhesion (n = 3; p = NS). We conclude that PI3K activation is necessary and sufficient to enhance monocytic adhesion under physiological flow conditions. BD110-expressing THP-1 cells should provide a useful tool for identifying the signaling pathways downstream of PI3K that are necessary for monocyte recruitment relevant to a variety of human vascular pathologies. PMID- 11278866 TI - Modulation of the degree and pattern of methyl-esterification of pectic homogalacturonan in plant cell walls. Implications for pectin methyl esterase action, matrix properties, and cell adhesion. AB - Homogalacturonan (HG) is a multifunctional pectic polysaccharide of the primary cell wall matrix of all land plants. HG is thought to be deposited in cell walls in a highly methyl-esterified form but can be subsequently de-esterified by wall based pectin methyl esterases (PMEs) that have the capacity to remove methyl ester groups from HG. Plant PMEs typically occur in multigene families/isoforms, but the precise details of the functions of PMEs are far from clear. Most are thought to act in a processive or blockwise fashion resulting in domains of contiguous de-esterified galacturonic acid residues. Such de-esterified blocks of HG can be cross-linked by calcium resulting in gel formation and can contribute to intercellular adhesion. We demonstrate that, in addition to blockwise de esterification, HG with a non-blockwise distribution of methyl esters is also an abundant feature of HG in primary plant cell walls. A partially methyl-esterified epitope of HG that is generated in greatest abundance by non-blockwise de esterification is spatially regulated within the cell wall matrix and occurs at points of cell separation at intercellular spaces in parenchymatous tissues of pea and other angiosperms. Analysis of the properties of calcium-mediated gels formed from pectins containing HG domains with differing degrees and patterns of methyl-esterification indicated that HG with a non-blockwise pattern of methyl ester group distribution is likely to contribute distinct mechanical and porosity properties to the cell wall matrix. These findings have important implications for our understanding of both the action of pectin methyl esterases on matrix properties and mechanisms of intercellular adhesion and its loss in plants. PMID- 11278865 TI - Enhanced antiviral and antiproliferative properties of a STAT1 mutant unable to interact with the protein kinase PKR. AB - We have previously reported a physical association between STAT1 and the protein kinase double-stranded RNA-activated protein kinase (PKR). PKR inhibited STAT1 function in a manner independent of PKR kinase activity. In this report, we have further characterized the properties of both molecules by mapping the sites of their interaction. A STAT1 mutant unable to interact with PKR displays enhanced interferon gamma (IFN-gamma)-induced transactivation capacity compared with STAT1. This effect appears to be mediated by the higher capacity of STAT1 mutant to heterodimerize with STAT3. Furthermore, expression of STAT1 mutant in STAT1(-/ ) cells enhances both the antiviral and antiproliferative effects of IFNs as opposed to STAT1. We also provide evidence that STAT1 functions as an inhibitor of PKR in vitro and in vivo. That is, phosphorylation of eIF-2alpha is enhanced in STAT1(-/-) than STAT1(+/+) cells in vivo, and this correlates with higher activation capacity of PKR in STAT1(-/-) cells. Genetic experiments in yeast demonstrate the inhibition of PKR activation and eIF-2alpha phosphorylation by STAT1 but not by STAT1 mutant. These data substantiate our previous findings on the inhibitory effects of PKR on STAT1 and implicate STAT1 in translational control through the modulation of PKR activation and eIF-2alpha phosphorylation. PMID- 11278867 TI - Mutation of a single conserved tryptophan in multidrug resistance protein 1 (MRP1/ABCC1) results in loss of drug resistance and selective loss of organic anion transport. AB - Multidrug resistance protein 1 (MRP1/ABCC1) belongs to the ATP-binding cassette transporter superfamily and is capable of conferring resistance to a broad range of chemotherapeutic agents and transporting structurally diverse conjugated organic anions. In this study, we found that substitution of a highly conserved tryptophan at position 1246 with cysteine (W1246C-MRP1) in the putative last transmembrane segment (TM17) of MRP1 eliminated 17beta-estradiol 17-(beta-d glucuronide) (E(2)17betaG) transport by membrane vesicles prepared from transiently transfected human embryonic kidney cells while leaving the capacity for leukotriene C(4)- and verapamil-stimulated glutathione transport intact. In addition, in contrast to wild-type MRP1, leukotriene C(4) transport by the W1246C MRP1 protein was no longer inhibitable by E(2)17betaG, indicating that the mutant protein had lost the ability to bind the glucuronide. A similar phenotype was observed when Trp(1246) was replaced with Ala, Phe, and Tyr. Confocal microscopy of cells expressing Trp(1246) mutant MRP1 molecules fused at the C terminus with green fluorescent protein showed that they were correctly routed to the plasma membrane. In addition to the loss of E(2)17betaG transport, HeLa cells stably transfected with W1246C-MRP1 cDNA were not resistant to the Vinca alkaloid vincristine and accumulated levels of [(3)H]vincristine comparable to those in vector control-transfected cells. Cells expressing W1246C-MRP1 were also not resistant to cationic anthracyclines (doxorubicin, daunorubicin) or the electroneutral epipodophyllotoxin VP-16. In contrast, resistance to sodium arsenite was only partially diminished, and resistance to potassium antimony tartrate remained comparable to that of cells expressing wild-type MRP1. This suggests that the structural determinants required for transport of heavy metal oxyanions differ from those for chemotherapeutic agents. Our results provide the first example of a tryptophan residue being so critically important for substrate specificity in a eukaryotic ATP-binding cassette transporter. PMID- 11278868 TI - Transgenic MUC1 interacts with epidermal growth factor receptor and correlates with mitogen-activated protein kinase activation in the mouse mammary gland. AB - MUC1 is a large (>400 kDa), heavily glycosylated transmembrane protein that is aberrantly expressed on greater than 90% of human breast carcinomas and subsequent metastases. The precise function of MUC1 overexpression in tumorigenesis is unknown, although various domains of MUC1 have been implicated in cell adhesion, cell signaling, and immunoregulation. Stimulation of the MDA-MB 468 breast cancer line as well as mouse mammary glands with epidermal growth factor results in the co-immunoprecipitation of MUC1 with a tyrosine phosphorylated protein of approximately 180 kDa. We have generated transgenic lines overexpressing full-length (MMF), cytoplasmic tail deleted (DeltaCT), or tandem repeat deleted (DeltaTR)-human MUC1 under the control of the mouse mammary tumor virus promoter to further examine the role of MUC1 in signaling and tumorigenesis. Immunoprecipitation experiments revealed that full-length transgenic MUC1 physically associates with all four erbB receptors, and co localizes with erbB1 in the lactating gland. Furthermore, we detected a sharp increase in ERK1/2 activation in MUC1 transgenic mammary glands compared with Muc1 null and wild-type animals. These results point to a novel function of increased MUC1 expression, potentiation of erbB signaling through the activation of mitogenic MAP kinase pathways. PMID- 11278870 TI - The fission yeast copper-sensing transcription factor Cuf1 regulates the copper transporter gene expression through an Ace1/Amt1-like recognition sequence. AB - Transcriptional regulation of genes encoding critical components of copper transport is essential for copper homeostasis and growth in yeast. Analysis of regulatory regions in the promoter of the ctr4(+) copper transporter gene in fission yeast Schizosaccharomyces pombe reveals the identity of a conserved copper-signaling element (CuSE), which is recognized by the transcription factor Cuf1. We demonstrate that CuSE is necessary for transcriptional activation in response to copper deprivation conditions. Interestingly, the CuSE element bears a strong sequence similarity to the recognition site, denoted MRE (metal regulatory element), which is recognized by a distinct class of copper sensors required for copper detoxification, including Ace1 from Saccharomyces cerevisiae and Amt1 from Candida glabrata. When a consensus MRE from S. cerevisiae is introduced into S. pombe, transcription is induced by copper deprivation in a Cuf1-dependent manner, similar to regulation by Mac1, the nuclear sensor for regulating the expression of genes encoding components involved in copper transport in S. cerevisiae. UV-cross-linking experiments show that the Cuf1 protein directly binds the CuSE. These results demonstrate that the Cuf1 nutritional copper-sensing factor possesses a module that functions similarly to domains found in the Ace1/Amt1 class of metalloregulatory factors, which allows the protein to act through a closely related MRE-like sequence to regulate copper transport gene expression in S. pombe. PMID- 11278869 TI - Identification of sperm-specific proteins that interact with A-kinase anchoring proteins in a manner similar to the type II regulatory subunit of PKA. AB - The cAMP-dependent protein kinase (PKA) is targeted to specific subcellular compartments through its interaction with A-kinase anchoring proteins (AKAPs). AKAPs contain an amphipathic helix domain that binds to the type II regulatory subunit of PKA (RII). Synthetic peptides containing this amphipathic helix domain bind to RII with high affinity and competitively inhibit the binding of PKA with AKAPs. Addition of these anchoring inhibitor peptides to spermatozoa inhibits motility (Vijayaraghavan, S., Goueli, S. A., Davey, M. P., and Carr, D. W. (1997) J. Biol. Chem. 272, 4747-4752). However, inhibition of the PKA catalytic activity does not mimic these peptides, suggesting that the peptides are disrupting the interaction of AKAP(s) with proteins other than PKA. Using the yeast two-hybrid system, we have now identified two sperm-specific human proteins that interact with the amphipathic helix region of AKAP110. These proteins, ropporin (a protein previously shown to interact with the Rho signaling pathway) and AKAP-associated sperm protein, are 39% identical to each other and share a strong sequence similarity with the conserved domain on the N terminus of RII that is involved in dimerization and AKAP binding. Mutation of conserved residues in ropporin or RII prevents binding to AKAP110. These data suggest that sperm contains several proteins that bind to AKAPs in a manner similar to RII and imply that AKAPs may have additional and perhaps unique functions in spermatozoa. PMID- 11278871 TI - Molecular and biochemical characterization of a novel oxysterol-binding protein (OSBP2) highly expressed in retina. AB - We are interested in understanding the possible function(s) of the oxysterol binding proteins in mediating oxysterol cytotoxicity in the retina. In this study we describe the cloning, localization, and biological activity of a novel oxysterol-binding protein (OSBP2), and complete the molecular characterization of the previously known OSBP1. Both OSBP genes contain 14 exons and have similar exon sizes and splice sites suggesting they may have arisen from a gene duplication event. OSBP1 is located in chromosome 11q12.1, and OSBP2 is located in 22q12. At the protein level they share 63% overall similarity and although they have unique N termini, both have similar pleckstrin homology domains within the N terminus region. Northern blot analyses indicate that OSBP1 is broadly expressed in human and monkey tissues. OSBP2 is detected mainly in retina, testis, and fetal liver. Western blot analysis using peptide antibodies specific to OSBP1 and OSBP2 detected the proteins in different subcellular fractions in the retinal monkey tissue. OSBP1 is detected mainly in the soluble or cytosolic fraction and nuclei whereas OSBP2 is detected exclusively in the detergent soluble fraction suggesting association with membranes. Immunohistochemical localization of OSBP1 and OSBP2 in the monkey retina placed these two proteins in similar but distinct areas of the inner retina. OSBP2 was found to bind 7 ketocholesterol but to have very little affinity for cholesterol or 25 hydroxycholesterol. PMID- 11278873 TI - Ivermectin, an unconventional agonist of the glycine receptor chloride channel. AB - The effects of the antihelmintic, ivermectin, were investigated in recombinantly expressed human alpha(1) homomeric and alpha(1)beta heteromeric glycine receptors (GlyRs). At low (0.03 microm) concentrations ivermectin potentiated the response to sub-saturating glycine concentrations, and at higher (> or =0.03 microm) concentrations it irreversibly activated both alpha(1) homomeric and alpha(1)beta heteromeric GlyRs. Relative to glycine-gated currents, ivermectin-gated currents exhibited a dramatically reduced sensitivity to inhibition by strychnine, picrotoxin, and zinc. The insensitivity to strychnine could not be explained by ivermectin preventing the access of strychnine to its binding site. Furthermore, the elimination of a known glycine- and strychnine-binding site by site-directed mutagenesis had little effect on ivermectin sensitivity, demonstrating that the ivermectin- and glycine-binding sites were not identical. Ivermectin strongly and irreversibly activated a fast-desensitizing mutant GlyR after it had been completely desensitized by a saturating concentration of glycine. Finally, a mutation known to impair dramatically the glycine signal transduction mechanism had little effect on the apparent affinity or efficacy of ivermectin. Together, these findings indicate that ivermectin activates the GlyR by a novel mechanism. PMID- 11278872 TI - The hepatitis B virus-X protein activates a phosphatidylinositol 3-kinase dependent survival signaling cascade. AB - The hepatitis B virus-X (HBx) protein is known as a multifunctional protein that not only coactivates transcription of viral and cellular genes but coordinates the balance between proliferation and programmed cell death, by inducing or blocking apoptosis. In this study the role of the HBx protein in activation of phosphatidylinositol 3-kinase (PI3K) was investigated as a possible cause of anti apoptosis in liver cells. HBx relieved serum deprivation-induced and pro-apoptic stimuli-induced apoptosis in Chang liver (CHL) cells. Treatment with 1-d-3-deoxy 3-fluoro-myo-inositol, an antagonist to PI3K, which blocks the formation of 3' phosphorylated phosphatidyl inositol in CHL cells transformed by HBx (CHL-X) but not normal Chang liver (CHL) cells, showed a marked loss of viability with evidence of apoptosis. Similarly, treatment with wortmannin, an inhibitor of PI3K, stimulated apoptosis in HBx-transformed CHL cells but not in normal cells, confirming that HBx blocks apoptosis through the PI3K pathway. The serine 47 threonine kinase, Akt, one of the downstream effectors of PI3K-dependent survival signaling was 2-fold higher in HBx-transformed CHL (CHL-X) cells than CHL cells. Phosphorylation of Akt at serine 473 and Bad at serine 136 were induced by HBx, which were specifically blocked by wortmannin and dominant negative mutants of Akt and Bad, respectively. We also demonstrated that HBx inhibits caspase 3 activity and HBx down-regulation of caspase 3 activity was blocked by the PI3K inhibitor. Regions required for PI3K phosphorylation on the HBx protein overlap with the known transactivation domains. HBx blocks apoptosis induced by serum withdrawal in CHL cells in a p53-independent manner. The results indicate that, unlike other DNA tumor viruses that block apoptosis by inactivating p53, the hepatitis B virus achieves protection from apoptotic death through a HBx-PI3K-Akt Bad pathway and by inactivating caspase 3 activity that is at least partially p53 independent in liver cells. Moreover, these data suggest that modulation of the PI3K activity may represent a potential therapeutic strategy to counteract the occurrence of apoptosis in human hepatocellular carcinoma. PMID- 11278874 TI - Roles of the C termini of alpha -, beta -, and gamma -subunits of epithelial Na+ channels (ENaC) in regulating ENaC and mediating its inhibition by cytosolic Na+. AB - The amiloride-sensitive epithelial Na(+) channels (ENaC) in the intralobular duct cells of mouse mandibular glands are inhibited by the ubiquitin-protein ligase, Nedd4, which is activated by increased intracellular Na(+). In this study we have used whole-cell patch clamp methods in mouse mandibular duct cells to investigate the role of the C termini of the alpha-, beta-, and gamma-subunits of ENaC in mediating this inhibition. We found that peptides corresponding to the C termini of the beta- and gamma-subunits, but not the alpha-subunit, inhibited the activity of the Na(+) channels. This mechanism did not involve Nedd4 and probably resulted from the exogenous C termini interfering competitively with the protein protein interactions that keep the channels active. In the case of the C terminus of mouse beta-ENaC, the interacting motif included betaSer(631), betaAsp(632), and betaSer(633). In the C terminus of mouse gamma-ENaC, it included gammaSer(640). Once these motifs were deleted, we were able to use the C termini of beta- and gamma-ENaC to prevent Nedd4-mediated down-regulation of Na(+) channel activity. The C terminus of the alpha-subunit, on the contrary, did not prevent Nedd4-mediated inhibition of the Na(+) channels. We conclude that mouse Nedd4 interacts with the beta- and gamma-subunits of ENaC. PMID- 11278875 TI - 12/15-lipoxygenase translocation enhances site-specific actin polymerization in macrophages phagocytosing apoptotic cells. AB - The enzyme 12/15-lipoxygenase (12/15-LO) introduces peroxyl groups in a position specific manner into unsaturated fatty acids in certain cells, but the role of such enzymatic lipid peroxidation remains poorly defined. Here we report a novel function for 12/15-LO in mouse peritoneal macrophages. When macrophages were coincubated with apoptotic cells, the enzyme translocated from cytosol to the plasma membrane and was more extensively concentrated at sites where macrophages bound apoptotic cells, colocalizing with polymerized actin of emerging filopodia. Disruption of F-actin did not prevent the 12/15-LO translocation. In contrast, inhibition of the 12/15-LO activity, or utilization of genetically engineered macrophages in which the 12/15-LO gene has been disrupted, greatly reduced actin polymerization in phagocytosing macrophages. Lysates of 12/15-LO-deficient macrophages had significantly lower ability to promote in vitro actin polymerization than the lysates of wild type macrophages. These studies suggest that the 12/15-LO enzyme plays a major role in local control of actin polymerization in macrophages in response to interaction with apoptotic cells. PMID- 11278876 TI - Exchangeability of N termini in the ligand-gated porins of Escherichia coli. AB - The ferric siderophore transporters of the Gram-negative bacterial outer membrane manifest a unique architecture: Their N termini fold into a globular domain that lodges within, and physically obstructs, a transmembrane porin beta-barrel formed by their C termini. We exchanged and deleted the N termini of two such siderophore receptors, FepA and FhuA, which recognize and transport ferric enterobactin and ferrichrome, respectively. The resultant chimeric proteins and empty beta-barrels avidly bound appropriate ligands, including iron complexes, protein toxins, and viruses. Thus, the ability to recognize and discriminate these molecules fully originates in the transmembrane beta-barrel domain. Both the hybrid and the deletion proteins also transported the ferric siderophore that they bound. The FepA constructs showed less transport activity than wild type receptor protein, but the FhuA constructs functioned with turnover numbers that were equivalent to wild type. The mutant proteins displayed the full range of transport functionalities, despite their aberrant or missing N termini, confirming (Braun, M., Killmann, H., and Braun, V. (1999) Mol. Microbiol. 33, 1037-1049) that the globular domain within the pore is dispensable to the siderophore internalization reaction, and when present, acts without specificity during solute uptake. These and other data suggest a transport process in which siderophore receptors undergo multiple conformational states that ultimately expel the N terminus from the channel concomitant with solute internalization. PMID- 11278877 TI - Peculiar 2-aminopurine fluorescence monitors the dynamics of open complex formation by bacteriophage T7 RNA polymerase. AB - The kinetics of promoter binding and open complex formation in bacteriophage T7 RNA polymerase was investigated using 2-aminopurine (2-AP) modified promoters. 2 AP serves as an ideal probe to measure the kinetics of open complex formation because its fluorescence is sensitive to both base-unpairing and base-unstacking and to the nature of the neighboring bases. All four 2-AP bases in the TATA box showed an increase in fluorescence with similar kinetics upon binding to the T7 RNA polymerase, indicating that the TATA sequence becomes unpaired in a concerted manner. The 2-AP at -4 showed a peculiarly large increase in fluorescence upon binding to the T7 RNA polymerase. Based on the recent crystal structure of the T7 RNA polymerase-DNA complex, we propose that the large fluorescence increase is due to unstacking of the 2-AP base at -4 from the guanine at -5, during open complex formation. The unstacking may be a critical event in directing and placing the template strand correctly in the T7 RNA polymerase active site upon promoter melting for template directed RNA synthesis. Based on equilibrium fluorescence and stopped-flow kinetic studies, we propose that a fast form of T7 RNA polymerase binds promoter double-stranded DNA by a three-step mechanism. The initial collision complex or a closed complex, ED(c) is formed with a K(d) of 1.8 microm. This complex isomerizes to an open complex, ED(o1), in an energetically unfavorable reaction with an equilibrium constant of 0.12. The ED(o1) further isomerizes to a more stable open complex, ED(o2), with a rate constant around 300 s(-1). Thus, in the absence of the initiating nucleotide, GTP, the overall equilibrium constant for closed to open complex conversion is 0.5 and the net rate of open complex formation is nearly 150 s(-1). PMID- 11278878 TI - Dependence of the bi-functional nature of a sialyltransferase from Neisseria meningitidis on a single amino acid substitution. AB - The L1 immunotype strain 126E of Neisseria meningitidis has been shown to have an N-acetyl-neuraminic acid-containing lipooligosaccharide in which an alpha-linked galactose from a P(k) epitope is substituted at the O6 position (Wakarchuk, W. W., Gilbert, M., Martin, A., Wu, Y., Brisson, J. R., Thibault, P., and Richards, J. C. (1998) Eur. J. Biochem. 254, 626-633). Using a synthetic P(k)-epitope containing acceptor in glycosyltransferase reactions, we were able to show by NMR analysis of the reaction product that the 126E(L1)-derived sialyltransferase can make both alpha-2,3 and alpha-2,6 linkages to the terminal galactose. Gene disruption experiments showed that the lst gene in 126E(L1) was responsible for the in vivo addition of the alpha-2,6-linked N-acetyl-neuraminic acid residue. By site-directed mutagenesis it was possible to change the MC58(L3)-derived enzyme into a bifunctional enzyme with a single amino acid change at position 168, where a glycine was changed to an isoleucine. We performed a gene replacement experiment where the 126E(L1) alpha-2,3/6-sialyltransferase was replaced by allelic exchange with the monofunctional MC58(L3) alpha-2,3-sialyltransferase and with the mutant MC58(L3) allele G168I. We observed that the level of LOS sialylation with the G168I allele was very similar to that of the wild type 126E(L1), indicating that residue 168 is the critical residue for the alpha-2,6 sialyltransferase activity in vitro as well as in vivo. PMID- 11278879 TI - The essential HupB and HupN proteins of Pseudomonas putida provide redundant and nonspecific DNA-bending functions. AB - A protein mixture containing two major components able to catalyze a beta recombination reaction requiring nonspecific DNA bending was obtained by fractionation of a Pseudomonas putida extract. N-terminal sequence analysis and genomic data base searches identified the major component as an analogue of HupB of Pseudomonas aeruginosa and Escherichia coli, encoding one HU protein variant. The minor component of the fraction, termed HupN, was divergent enough from HupB to predict a separate DNA-bending competence. The determinants of the two proteins were cloned and hyperexpressed, and the gene products were purified. Their activities were examined in vitro in beta-recombination assays and in vivo by complementation of the Hbsu function of Bacillus subtilis. HupB and HupN were equally efficient in all tests, suggesting that they are independent and functionally redundant DNA bending proteins. This was reflected in the maintenance of in vivo activity of the final sigma54 Ps promoter of the toluene degradation plasmid, TOL, which requires facilitated DNA bending, in DeltahupB or DeltahupN strains. However, hupB/hupN double mutants were not viable. It is suggested that the requirement for protein-facilitated DNA bending is met in P. putida by two independent proteins that ensure an adequate supply of an essential cellular activity. PMID- 11278880 TI - The major CD9 and CD81 molecular partner. Identification and characterization of the complexes. AB - By associating with specific partner molecules and with each other, the tetraspanins are thought to assemble multimolecular complexes that may be especially relevant with respect to metastasis. We have previously identified a 135-kDa molecule (CD9P-1) as a major molecular partner of CD9 in cancer cell lines. This molecule was identified, after immunoaffinity purification and mass spectrometry analysis, as the protein encoded by the KIAA1436 gene and the human ortholog of a rat protein known as FPRP. Cross-linking experiments detected a complex of the size of CD9 plus CD9P-1, showing that these glycoproteins directly associate with each other, probably in the absence of any other molecule. The use of chimeric CD9/CD82 molecules revealed the role of the second half of CD9, comprising the large extracellular loop and the fourth transmembrane domain. CD9P 1 was also shown to form separate complexes with CD81 and with an unidentified 175-kDa molecule. It also associated with other tetraspanins under conditions maintaining tetraspanin/tetraspanin interactions. The identification of a protein strongly linked to the tetraspanin web and the production of a specific monoclonal antibody will help to further characterize the role of this "web" under physiological and pathological conditions. PMID- 11278881 TI - A single residue at the active site of CD38 determines its NAD cyclizing and hydrolyzing activities. AB - CD38 is a multifunctional enzyme involved in metabolizing two Ca(2+) messengers, cyclic ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP). When incubated with NAD, CD38 predominantly hydrolyzes it to ADP-ribose (NAD glycohydrolase), but a trace amount of cADPR is also produced through cyclization of the substrate. Site-directed mutagenesis was used to investigate the amino acid important for controlling the hydrolysis and cyclization reactions. CD38 and its mutants were produced in yeast, purified, and characterized by immunoblot. Glu-146 is a conserved residue present in the active site of CD38. Its replacement with Phe greatly enhanced the cyclization activity to a level similar to that of the NAD hydrolysis activity. A series of additional replacements was made at the Glu-146 position including Ala, Asn, Gly, Asp, and Leu. All the mutants exhibited enhanced cyclase activity to various degrees, whereas the hydrolysis activity was inhibited greatly. E146A showed the highest cyclase activity, which was more than 3-fold higher than its hydrolysis activity. All mutants also cyclized nicotinamide guanine dinucleotide to produce cyclic GDP. This activity was enhanced likewise, with E146A showing more than 9-fold higher activity than the wild type. In addition to NAD, CD38 also hydrolyzed cADPR effectively, and this activity was correspondingly depressed in the mutants. When all the mutants were considered, the two cyclase activities and the two hydrolase activities were correlated linearly. The Glu-146 replacements, however, only minimally affected the base-exchange activity that is responsible for synthesizing NAADP. Homology modeling was used to assess possible structural changes at the active site of E146A. These results are consistent with Glu-146 being crucial in controlling specifically and selectively the cyclase and hydrolase activities of CD38. PMID- 11278883 TI - Constitutive and agonist-dependent homo-oligomerization of the thyrotropin releasing hormone receptor. Detection in living cells using bioluminescence resonance energy transfer. AB - The ability of G-protein-coupled receptors (GPCRs) to interact to form new functional structures, either forming oligomers with themselves or forming associations with other intracellular proteins, has important implications for the regulation of cellular events; however, little is known about how this occurs. Here, we have employed a newly emerging technology, bioluminescence resonance energy transfer (BRET), used to study protein-protein interactions in living cells, to demonstrate that the thyrotropin-releasing hormone receptor (TRHR) forms constitutive homo-oligomers. This formation of TRHR homo-oligomers in the absence of ligand was shown by demonstration of an energy transfer between TRHR molecules fused to either donor, Renilla luciferase (Rluc) or acceptor, enhanced yellow fluorescent protein (EYFP) molecules. This interaction was shown to be specific, since energy transfer was not detected between co-expressed tagged TRHRs and either complementary tagged gonadotropin-releasing hormone (GnRH) or beta(2)-adrenergic receptors. Furthermore, generation of a BRET signal between the TRHRs could only be inhibited by co-expression of the wild-type TRHR and not by other GPCRs. Agonist stimulation led to a time- and dose-dependent increase in the amount of energy transfer. Inhibition of receptor internalization by co-expression of dynamin mutant K44A did not affect the interaction between TRHRs, suggesting that clustering of receptors within clathrin-coated pits is not sufficient for energy transfer to occur. BRET also provided evidence for the agonist-induced oligomerization of another GPCR, the GnRH receptor (GnRHR), and the presence of an agonist-induced interaction of the adaptor protein, beta arrestin, with TRHR and the absence of an interaction of beta-arrestin with GnRHR. This study supports the usefulness of BRET as a powerful tool for studying GPCR aggregations and receptor/protein interactions in general and presents evidence that the functioning unit of TRHRs exists as homomeric complexes. PMID- 11278882 TI - Oxidized low density lipoprotein decreases macrophage expression of scavenger receptor B-I. AB - Scavenger receptor class B type I (SR-BI) has recently been identified as a high density lipoprotein (HDL) receptor that mediates bidirectional flux of cholesterol across the plasma membrane. We have previously demonstrated that oxidized low density lipoprotein (OxLDL) will increase expression of another class B scavenger receptor, CD36 (Han, J., Hajjar, D. P., Febbraio, M., and Nicholson, A. C. (1997) J. Biol. Chem. 272, 21654-21659). In studies reported herein, we evaluated the effects of OxLDL on expression of SR-BI in macrophages to determine how exposure to this modified lipoprotein could alter SR-BI expression and cellular lipid flux. OxLDL decreased SR-BI expression in a dose- and time-dependent manner. Incubation with OxLDL had no effect on the membrane distribution of SB-BI, and it decreased expression of both cytosolic and membrane protein. Consistent with its effect on SR-BI protein expression, OxLDL decreased SR-BI mRNA in a dose-dependent manner. The ability of OxLDL to decrease SR-BI expression was dependent on the degree of LDL oxidation. OxLDL decreased both [(14)C]cholesteryl oleate/HDL uptake and efflux of [(14)C]cholesterol to HDL in a time-dependent manner. Incubation of macrophages with 7-ketocholesterol, but not free cholesterol, also inhibited expression of SR-BI. Finally, we demonstrate that the effect of OxLDL on SR-BI is dependent on the differentiation state of the monocyte/macrophage. These results imply that in addition to its effect in inducing foam cell formation in macrophages through increased uptake of oxidized lipids, OxLDL may also enhance foam cell formation by altering SR-BI-mediated lipid flux across the cell membrane. PMID- 11278884 TI - Structure of the bound dioxygen species in the cytochrome oxidase reaction of cytochrome cd1 nitrite reductase. AB - Reduction of dioxygen to water is a key process in aerobic life, but atomic details of this reaction have been elusive because of difficulties in observing active oxygen intermediates by crystallography. Cytochrome cd(1) is a bifunctional enzyme, capable of catalyzing the one-electron reduction of nitrite to nitric oxide, and the four-electron reduction of dioxygen to water. The latter is a cytochrome oxidase reaction. Here we describe the structure of an active dioxygen species in the enzyme captured by cryo-trapping. The productive binding mode of dioxygen in the active site is very similar to that of nitrite and suggests that the catalytic mechanisms of oxygen reduction and nitrite reduction are closely related. This finding has implications to the understanding of the evolution of oxygen-reducing enzymes. Comparison of the dioxygen complex to complexes of cytochrome cd(1) with stable diatomic ligands shows that nitric oxide and cyanide bind in a similar bent conformation to the iron as dioxygen whereas carbon monoxide forms a linear complex. The significance of these differences is discussed. PMID- 11278885 TI - Regulation of nuclear factor kappa B transactivation. Implication of phosphatidylinositol 3-kinase and protein kinase C zeta in c-Rel activation by tumor necrosis factor alpha. AB - Transactivation by c-Rel (nuclear factor kappaB) was dependent on phosphorylation of several serines in the transactivation domain, indicating that it is a phosphorylation-dependent Ser-rich domain. By Ser --> Ala mutational and deletion analysis, we have identified two regions in this domain: 1) a C-terminal region (amino acids 540-588), which is required for basal activity; and 2) the 422-540 region, which responds to external stimuli as tumor necrosis factor (TNF) alpha or phorbol myristate acetate plus ionomycin. Ser from 454 to 473 were shown to be required for TNFalpha-induced activation, whereas Ser between 492 and 519 were required for phorbol myristate acetate plus ionomycin activation. Phosphatidylinositol 3-kinase (PI3K) and protein kinase C (PKC) zeta were identified as downstream signaling molecules of TNFalpha-activation of c-Rel transactivating activity. Interestingly, dominant negative forms of PI3K inhibited PKCzeta activation and dominant negative PKCzeta inhibited PI3K mediated activation of c-Rel transactivating activity, indicating a cross-talk between both enzymes. We have identified the critical role of different Ser for PKCzeta- and PI3K-mediated responses. Interestingly, those c-Rel mutants not only did not respond to TNFalpha but also acted as dominant negative forms of nuclear factor kappaB activation. PMID- 11278886 TI - Delta 3,5,delta 2,4-dienoyl-CoA isomerase is a multifunctional isomerase. A structural and mechanistic study. AB - Delta(3,5),Delta(2,4)-Dienoyl-CoA isomerase (DI), an auxiliary enzyme of unsaturated fatty acid beta-oxidation, was purified from rat mitochondria and peroxisomes and subjected to N-terminal sequencing to facilitate a mechanistic study of this enzyme. The mature mitochondrial DI from rat heart was lacking its 34 N-terminal amino acid residues that have the properties of a mitochondrial targeting sequence. The peroxisomal isomerase was identified as a product of the same gene with a truncated and ragged N terminus. Expression of the cDNA coding for the mature mitochondrial DI in Escherichia coli yielded an enzyme preparation that was as active as the native DI. Because the recombinant DI also exhibited Delta(3,5,7),Delta(2,4,6)-trienoyl-CoA isomerase (TI) activity, both isomerases reside on the same protein. Mutations of any of the 3 acidic amino acid residues located at the active site (Modis, Y., Filppula, S. A., Novikov, D. K., Norledge, B., Hiltunen, J. K., and Wierenga, R. K. (1998) Structure 6, 957-970) caused activity losses. In contrast to only a 10-fold decrease in activity upon replacement of Asp(176) by Ala, substitutions of Asp(204) by Asn and of Glu(196) by Gln resulted in 10(5)-fold lower activities. Such activity losses are consistent with the direct involvement of these latter two residues in the proposed proton transfers at carbons 2 and 6 or 8 of the substrates. Probing of the wild-type and mutants forms of the enzyme with 2,5-octadienoyl-CoA as substrate revealed low Delta(2),Delta(3)-enoyl-CoA isomerase and Delta(5),Delta(4)-enoyl-CoA isomerase activities catalyzed by Glu(196) and Asp(204), respectively. Altogether, these data reveal that positional isomerizations of the diene and triene are facilitated by simultaneous proton transfers involving Glu(196) and Asp(204), whereas each residue alone can catalyze, albeit less efficiently, a monoene isomerization. PMID- 11278887 TI - Analysis of gene induction and arrest site transcription in yeast with mutations in the transcription elongation machinery. AB - In vitro, transcript elongation by RNA polymerase II is impeded by DNA sequences, DNA-bound proteins, and small ligands. Transcription elongation factor SII (TFIIS) assists RNA polymerase II to transcribe through these obstacles. There is however, little direct evidence that SII-responsive arrest sites function in living cells nor that SII facilitates readthrough in vivo. Saccharomyces cerevisiae strains lacking elongation factor SII and/or containing a point mutation in the second largest subunit of RNA polymerase II, which slows the enzyme's RNA elongation rate, grow slowly and have defects in mRNA metabolism, particularly in the presence of nucleotide-depleting drugs. Here we have examined transcriptional induction in strains lacking SII or containing the slow polymerase mutation. Both mutants and a combined double mutant were defective in induction of GAL1 and ENA1. This was not due to an increase in mRNA degradation and was independent of any drug treatment, although treatment with the nucleotide depleting drug 6-azauracil exacerbated the effect preferentially in the mutants. These data are consistent with mutants in the Elongator complex, which show slow inductive responses. When a potent in vitro arrest site was transcribed in these strains, there was no perceptible effect upon mRNA accumulation. These data suggest that an alternative elongation surveillance mechanism exists in vivo to overcome arrest. PMID- 11278888 TI - Amino acid residues involved in gating identified in the first membrane-spanning domain of the rat P2X(2) receptor. AB - The first hydrophobic segment of the rat P2X(2) receptor extends from residue Leu(29) to Val(51). In the rat P2X(2) receptor, we mutated amino acids in this segment and adjoining flanking regions (Asp(15) through Thr(60)) individually to cysteine and expressed the constructs in human embryonic kidney cells. Whole-cell recordings were used to measure membrane currents evoked by brief (2-s) applications of ATP (0.3-100 microM). Currents were normal except for Y16C, R34C, Y43C, Y55C, and Q56C (no currents but normal membrane expression by immunohistochemistry), Q37C (small currents), and F44C (normal current but increased sensitivity to ATP, as well as alphabeta-methylene-ATP). We used methanethiosulfonates of positive, negative, or no charge to test the accessibility of the substituted cysteines. D15C, P19C, V23C, V24C, G30C, Q37C, F44C, and V48C were strongly inhibited by neutral, membrane-permeant methanethiosulfonates. Only V48C was also inhibited by positively and negatively charged methanethiosulfonates, consistent with an extracellular position; however, accessibility of V48C was increased by channel opening. V48C could disulfide with I328C, as shown by the large increase in ATP-evoked current caused by reducing agents. The results suggest that Val(48) at the outer end of the first hydrophobic segment takes part in the gating movement of channel opening. PMID- 11278889 TI - Activation loop sequences confer substrate specificity to phosphoinositide 3 kinase alpha (PI3Kalpha ). Functions of lipid kinase-deficient PI3Kalpha in signaling. AB - Phosphoinositide 3-kinases (PI3Ks) are dual specificity lipid and protein kinases. While the lipid-dependent PI3K downstream signaling is well characterized, little is known about PI3K protein kinase signaling and structural determinants of lipid substrate specificity across the various PI3K classes. Here we show that sequences C-terminal to the PI3K ATP-binding site determine the lipid substrate specificity of the class IA PI3Kalpha (p85/p110alpha). Transfer of such activation loop sequences from class II PI3Ks, class III PI3Ks, and a related mammalian target of rapamycin (FRAP) into p110alpha turns the lipid substrate specificity of the resulting hybrid protein into that of the donor protein, while leaving the protein kinase activity unaffected. All resulting hybrids lacked the ability to produce phosphatidylinositol 3,4,5-trisphosphate in intact cells. Amino acid substitutions and structure modeling showed that two conserved positively charged (Lys and Arg) residues in the activation loop are crucial for the functionality of class I PI3Ks as phosphatidylinositol 4,5 bisphosphate kinases. By transient transfecion of 293 cells, we show that p110alpha hybrids, although unable to support lipid-dependent PI3K signaling, such as activation of protein kinase B/Akt and p70(S6k), retain the capability to associate with and phosphorylate insulin receptor substrate-1, with the same specificity and higher efficacy than wild type PI3Kalpha. Our data lay the basis for the understanding of the class I PI3K substrate selectivity and for the use of PI3Kalpha hybrids to dissect PI3Kalpha function as lipid and protein kinase. PMID- 11278890 TI - Calcium-activated potassium channels sustain calcium signaling in T lymphocytes. Selective blockers and manipulated channel expression levels. AB - To maintain Ca(2+) entry during T lymphocyte activation, a balancing efflux of cations is necessary. Using three approaches, we demonstrate that this cation efflux is mediated by Ca(2+)-activated K(+) (K(Ca)) channels, hSKCa2 in the human leukemic T cell line Jurkat and hIKCa1 in mitogen-activated human T cells. First, several recently developed, selective and potent pharmacological inhibitors of K(Ca) channels but not K(V) channels reduce Ca(2+) entry in Jurkat and in mitogen activated human T cells. Second, dominant-negative suppression of the native K(Ca) channel in Jurkat T cells by overexpression of a truncated fragment of the cloned hSKCa2 channel decreases Ca(2+) influx. Finally, introduction of the hIKCa1 channel into Jurkat T cells maintains rapid Ca(2+) entry despite pharmacological inhibition of the native small conductance K(Ca) channel. Thus, K(Ca) channels play a vital role in T cell Ca(2+) signaling. PMID- 11278891 TI - Molecular regulation of the endothelin-1 gene by hypoxia. Contributions of hypoxia-inducible factor-1, activator protein-1, GATA-2, AND p300/CBP. AB - Endothelin-1 (ET-1) is a peptide hormone with potent vasoconstrictor properties which is synthesized and secreted predominantly by vascular endothelial cells. Its production is regulated by numerous stimuli including ischemia and hypoxia, and the enhanced levels that occur during myocardial ischemia may contribute to the progression of heart failure. We reported previously a preliminary characterization of a hypoxia-inducible factor-1 (HIF-1) binding site in the human ET-1 promoter which contributed to the activation of ET-1 expression in endothelial cells. We report here that the HIF-1 binding site alone is not sufficient for the response to hypoxia but requires an additional 50 base pairs of flanking sequence that includes binding sites for the factors activator protein-1 (AP-1), GATA-2, and CAAT-binding factor (NF-1). Mutation of any one of these sites or the HIF-1 site eliminated induction by hypoxia. Mutations of the AP-1 and GATA-2 sites, but not the HIF-1 site, were complemented by overexpressing AP-1, GATA-2, HIF-1alpha, or the activator protein p300/CBP, restoring the response to hypoxia. Binding studies in vitro confirmed physical associations among GATA-2, AP-1, and HIF-1 factors. Overexpression or depletion of p300/CBP modulated the level of ET-1 promoter expression as well as the endogenous ET-1 transcript but did not change the fold induction by hypoxia in either case. Regulation of the ET-1 promoter by hypoxia in non-endothelial cells required overexpression of GATA-2 and HIF-1alpha. The results support essential roles for AP-1, GATA-2, and NF-1 in stabilizing the binding of HIF-1 and promoting recruitment of p300/CBP to the ET-1 hypoxia response complex. PMID- 11278892 TI - Drosophila heparan sulfate 6-O-sulfotransferase (dHS6ST) gene. Structure, expression, and function in the formation of the tracheal system. AB - Heparan sulfate, one of the most abundant components of the cell surface and the extracellular matrix, is involved in a variety of biological processes such as growth factor signaling, cell adhesion, and enzymatic catalysis. The heparan sulfate chains have markedly heterogeneous structures in which distinct sequences of sulfate groups determine specific binding properties. Sulfation at each different position of heparan sulfate is catalyzed by distinct enzymes, sulfotransferases. In this study, we identified and characterized Drosophila heparan sulfate 6-O-sulfotransferase (dHS6ST). The deduced primary structure of dHS6ST exhibited several common features found in those of mammalian HS6STs. We confirmed that, when the protein encoded by the cDNA was expressed in COS-7 cells, it showed HS6ST activity. Whole mount in situ hybridization revealed highly specific expression of dHS6ST mRNA in embryonic tracheal cells. The spatial and temporal pattern of dHS6ST expression in these cells clearly resembles that of the Drosophila fibroblast growth factor (FGF) receptor, breathless (btl). RNA interference experiments demonstrated that reduced dHS6ST activity caused embryonic lethality and disruption of the primary branching of the tracheal system. These phenotypes were reminiscent of the defects observed in mutants of FGF signaling components. We also show that FGF-dependent mitogen activated protein kinase activation is significantly reduced in dHS6ST double stranded RNA-injected embryos. These findings indicate that dHS6ST is required for tracheal development in Drosophila and suggest the evolutionally conserved roles of 6-O-sulfated heparan sulfate in FGF signaling. PMID- 11278893 TI - Hydroxyeicosanoids bind to and activate the low affinity leukotriene B4 receptor, BLT2. AB - Leukotriene B(4), an arachidonate metabolite, is a potent chemoattractant of leukocytes involved in various inflammatory diseases. Two G-protein-coupled receptors for leukotriene B(4) have been cloned and characterized. BLT1 (Yokomizo, T., Izumi, T., Chang, K., Takuwa, Y., and Shimizu, T. (1997) Nature 387, 620-624) is a high affinity receptor exclusively expressed in leukocytes, and BLT2 (Yokomizo, T., Kato, K., Terawaki, K., Izumi, T., and Shimizu, T. (2000) J. Exp. Med. 192, 421-432) is a low affinity receptor expressed more ubiquitously. Here we report the binding profiles of various BLT antagonists and eicosanoids to either BLT1 or BLT2 using the membrane fractions of Chinese hamster ovary cells stably expressing the receptor. BLT antagonists are grouped into three classes: BLT1-specific U-75302, BLT2-specific LY255283, and BLT1/BLT2 dual-specific ZK 158252 and CP 195543. We also show that 12(S) hydroxyeicosatetraenoic acid, 12(S)-hydroperxyeicosatetraenoic acid, and 15(S) hydroxyeicosatetraenoic acid competed with [(3)H]LTB(4) binding to BLT2, but not BLT1, dose dependently. These eicosanoids also cause calcium mobilization and chemotaxis through BLT2, again in contrast to BLT1. These findings suggest that BLT2 functions as a low affinity receptor, with broader ligand specificity for various eicosanoids, and mediates distinct biological and pathophysiological roles from BLT1. PMID- 11278894 TI - Phorbol esters and related analogs regulate the subcellular localization of beta 2-chimaerin, a non-protein kinase C phorbol ester receptor. AB - The novel phorbol ester receptor beta2-chimaerin is a Rac-GAP protein possessing a single copy of the C1 domain, a 50-amino acid motif initially identified in protein kinase C (PKC) isozymes that is involved in phorbol ester and diacylglycerol binding. We have previously shown that, like PKCs, beta2-chimaerin binds phorbol esters with high affinity in a phospholipid-dependent manner (Caloca, M. J., Fernandez, M. N., Lewin, N. E., Ching, D., Modali, R., Blumberg, P. M., and Kazanietz, M. G. (1997) J. Biol. Chem. 272, 26488-26496). In this paper we report that like PKC isozymes, beta2-chimaerin is translocated by phorbol esters from the cytosolic to particulate fraction. Phorbol esters also induce translocation of alpha1 (n)- and beta1-chimaerins, suggesting common regulatory mechanisms for all chimaerin isoforms. The subcellular redistribution of beta2-chimaerin by phorbol esters is entirely dependent on the C1 domain, as revealed by deletional analysis and site-directed mutagenesis. Interestingly, beta2-chimaerin translocates to the Golgi apparatus after phorbol ester treatment, as revealed by co-staining with the Golgi marker BODIPY-TR-ceramide. Structure relationship analysis of translocation using a series of PKC ligands revealed substantial differences between translocation of beta2-chimaerin and PKCalpha. Strikingly, the mezerein analog thymeleatoxin is not able to translocate beta2-chimaerin, although it very efficiently translocates PKCalpha. Phorbol esters also promote the association of beta2-chimaerin with Rac in cells. These data suggest that chimaerins can be positionally regulated by phorbol esters and that each phorbol ester receptor class has distinct pharmacological properties and targeting mechanisms. The identification of selective ligands for each phorbol ester receptor class represents an important step in dissecting their specific cellular functions. PMID- 11278895 TI - Differential binding regulation of microtubule-associated proteins MAP1A, MAP1B, and MAP2 by tubulin polyglutamylation. AB - The major neuronal post-translational modification of tubulin, polyglutamylation, can act as a molecular potentiometer to modulate microtubule-associated proteins (MAPs) binding as a function of the polyglutamyl chain length. The relative affinity of Tau, MAP2, and kinesin has been shown to be optimal for tubulin modified by approximately 3 glutamyl units. Using blot overlay assays, we have tested the ability of polyglutamylation to modulate the interaction of two other structural MAPs, MAP1A and MAP1B, with tubulin. MAP1A and MAP2 display distinct behavior in terms of tubulin binding; they do not compete with each other, even when the polyglutamyl chains of tubulin are removed, indicating that they have distinct binding sites on tubulin. Binding of MAP1A and MAP1B to tubulin is also controlled by polyglutamylation and, although the modulation of MAP1B binding resembles that of MAP2, we found that polyglutamylation can exert a different mode of regulation toward MAP1A. Interestingly, although the affinity of the other MAPs tested so far decreases sharply for tubulins carrying long polyglutamyl chains, the affinity of MAP1A for these tubulins is maintained at a significant level. This differential regulation exerted by polyglutamylation toward different MAPs might facilitate their selective recruitment into distinct microtubule populations, hence modulating their functional properties. PMID- 11278896 TI - Epiplakin, a novel member of the Plakin family originally identified as a 450-kDa human epidermal autoantigen. Structure and tissue localization. AB - A 450-kDa human epidermal autoantigen was originally identified as a protein that reacted with the serum from an individual with a subepidermal blistering disease. Molecular cloning of this protein has now shown that it contains 5065 amino acids and has a molecular mass of 552 kDa. As reported previously this protein, which we call epiplakin, belongs to the plakin family, but it has some very unusual features. Epiplakin has 13 domains that are homologous to the B domain in the COOH-terminal region of desmoplakin. The last five of these B domains, together with their associated linker regions, are particularly strongly conserved. However, epiplakin lacks a coiled-coil rod domain and an amino-terminal domain, both of which are found in all other known members of the plakin family. Furthermore, no dimerization motif was found in the sequence. Thus, it is likely that epiplakin exists in vivo as a single-chain structure. Epitope mapping experiments showed that the original patient's serum recognized a sequence unique to epiplakin, which was not found in plectin. Immunofluorescence staining revealed the presence of epiplakin in whole sheets of epidermis and esophagus, in glandular cells of eccrine sweat and parotid glands and in mucous epithelial cells in the stomach and colon. PMID- 11278897 TI - Identification of dual alpha 4beta1 integrin binding sites within a 38 amino acid domain in the N-terminal thrombin fragment of human osteopontin. AB - Previous work from our laboratory demonstrates that the alpha(4)beta(1) integrin is an adhesion receptor for OPN and that alpha(4)beta(1) binding site(s) are present in the N-terminal thrombin fragment of osteopontin (OPN) (Bayless, K. J., Meininger, G. A., Scholtz, J. M., and Davis, G. E. (1998) J. Cell Sci. 111, 1165 1174). The work presented here identifies two alpha(4)beta(1) binding sites within a recombinantly produced N-terminal thrombin fragment of human OPN. Initial experiments, using wild-type OPN containing an RGD sequence or an OPN-RGE mutant, showed identical alpha(4)beta(1)-dependent cell adhesive activity. A strategy to localize alpha(4)beta(1) binding sites within the thrombin fragment of osteopontin involved performing a series of truncation analyses. Removal of the last 39 amino acids (130) completely eliminated adhesion, indicating all binding activity was present within that portion of the molecule. Combined mutation and deletion analyses of this region revealed the involvement of dual alpha(4)beta(1) binding sites. Synthetic peptides for both regions in OPN, ELVTDFPTDLPAT (131) and SVVYGLR (162), were found to block alpha(4)beta(1) dependent adhesion. The first peptide when coupled to Sepharose bound the alpha(4)beta(1) integrin directly whereas a mutated ELVTEFPTELPAT peptide showed a dramatically reduced ability to bind. These data collectively demonstrate that dual alpha(4)beta(1) integrin binding sites are present in a 38 amino acid domain within the N-terminal thrombin fragment of OPN. PMID- 11278898 TI - A novel plant ferritin subunit from soybean that is related to a mechanism in iron release. AB - Ferritin is a multimeric iron storage protein composed of 24 subunits. Ferritin purified from dried soybean seed resolves into two peptides of 26.5 and 28 kDa. To date, the 26.5-kDa subunit has been supposed to be generated from the 28-kDa subunit by cleavage of the N-terminal region. We performed amino acid sequence analysis of the 28-kDa subunit and found that it had a different sequence from the 26.5-kDa subunit, thus rendering it novel among known soybean ferritins. We cloned a cDNA encoding this novel subunit from 10-day-old seedlings, each of which contained developed bifoliates, an epicotyl and a terminal bud. The 26.5 kDa subunit was found to be identical to that identified previously lacking the C terminal 16 residues that correspond to the E helix of mammalian ferritin. However, the corresponding region in the 28-kDa soybean ferritin subunit identified in this study was not susceptible to cleavage. We present evidence that the two different ferritin subunits in soybean dry seeds show differential sensitivity to protease digestions and that the novel, uncleaved 28-kDa ferritin subunit appears to stabilize the ferritin shell by co-existing with the cleaved 26.5-kDa subunit. These data demonstrate that soybean ferritin is composed of at least two different subunits, which have cooperative functional roles in soybean seeds. PMID- 11278899 TI - Role of a JAK3-dependent biochemical signaling pathway in platelet activation and aggregation. AB - Here we provide experimental evidence that identifies JAK3 as one of the regulators of platelet function. Treatment of platelets with thrombin induced tyrosine phosphorylation of the JAK3 target substrates STAT1 and STAT3. Platelets from JAK3-deficient mice displayed a decrease in tyrosine phosphorylation of STAT1 and STAT3. In accordance with these data, pretreatment of human platelets with the JAK3 inhibitor WHI-P131 markedly decreased the base-line enzymatic activity of constitutively active JAK3 and abolished the thrombin-induced tyrosine phosphorylation of STAT1 and STAT3. Following thrombin stimulation, WHI P131-treated platelets did not undergo shape changes indicative of activation such as pseudopod formation. WHI-P131 inhibited thrombin-induced degranulation/serotonin release as well as platelet aggregation. Highly effective platelet inhibitory plasma concentrations of WHI-P131 were achieved in mice without toxicity. WHI-P131 prolonged the bleeding time of mice in a dose dependent manner and improved event-free survival in a mouse model of thromboplastin-induced generalized and invariably fatal thromboembolism. To our knowledge, WHI-P131 is the first anti-thrombotic agent that prevents platelet aggregation by inhibiting JAK3. PMID- 11278900 TI - Prostaglandin receptor subtypes, EP3C and EP4, mediate the prostaglandin E2 induced cAMP production and sensitization of sensory neurons. AB - Although a number of prostaglandin E(2) (PGE(2)) receptor subtypes have been cloned, limited studies have been performed to elucidate subtypes that subserve specific actions of this eicosanoid, in part because of a paucity of selective receptor antagonists. Using reverse transcription-polymerase chain reaction (PCR) and antisense oligonucleotides, we examined which prostaglandin E(2) receptor (EP receptor) subtypes are expressed in sensory neurons and which mediate the PGE(2) induced increase in cAMP production and augmentation of peptide release. Reverse transcription-PCR of cDNA isolated from rat sensory neurons grown in culture revealed PCR products for the EP1, EP2, EP3C, and EP4 receptor subtypes but not the EP3A or EP3B. Preexposing neuronal cultures for 48 h to antisense oligonucleotides of EP3C and EP4 mRNA diminished expression of the respective receptors by approximately 80%, abolished the PGE(2)-stimulated production of cAMP, and blocked the ability of PGE(2) to augment release of immunoreactive substance P and calcitonin gene-related peptide. Pretreating with individual antisense against the EP2, EP3C, or EP4 receptors or combinations of missense oligonucleotides had no effect on PGE(2)-induced activity. Treatment with antisense to EP3C and EP4 receptor subtypes did not alter the ability of forskolin to increase cAMP or enhance peptide release. These results demonstrate that sensory neurons are capable of expressing multiple EP receptor subtypes but that only the EP3C and EP4 receptors mediate PGE(2)-induced sensitization of sensory neurons. PMID- 11278901 TI - Roles of water in heme peroxidase and catalase mechanisms. AB - A water molecule is coproduced with the Compound I intermediate in the reactions of native heme peroxidases and catalases with hydrogen peroxide. As a result of water release/rebinding from/to the coproduct formation site the Compound I intermediate may exist in two forms: a "wet" form, Compound I(H(2)O), in which a water molecule is present at or near the site of coproduct water formation, and Compound I, in which the coproduct water formation site is "dry." It is postulated that the absence or presence of a water molecule at this site provides the structural basis for a redox pathway switching mechanism, such that the transition states for 2-electron equivalent reduction of Compound I intermediates are accessible in the dry form, but that in the wet form only 1-electron equivalent processes are possible, unless release of water can be stimulated. This concept provides the basis of a general mechanism in which the classical functional distinction between catalases and peroxidases, as well as the more complex behavior observed in halide oxidation and halogenation reactions, appear as particular cases in which variations in the degree of retention of water at the coproduct formation site influence Compound I reactivity. PMID- 11278903 TI - Alteration of a nonconserved active site residue in the chemotaxis response regulator CheY affects phosphorylation and interaction with CheZ. AB - CheY is a response regulator in the well studied two-component system that mediates bacterial chemotaxis. Phosphorylation of CheY at Asp(57) enhances its interaction with the flagellar motor. Asn(59) is located near the phosphorylation site, and possible roles this residue may play in CheY function were explored by mutagenesis. Cells containing CheY59NR or CheY59NH exhibited hyperactive phenotypes (clockwise flagellar rotation), and CheY59NR was characterized biochemically. A continuous enzyme-linked spectroscopic assay that monitors P(i) concentration was the primary method for kinetic analysis of phosphorylation and dephosphorylation. CheY59NR autodephosphorylated at the same rate as wild-type CheY and phosphorylated similarly to wild type with acetyl phosphate and faster (4-14x) with phosphoramidate and monophosphoimidazole. CheY59NR was extremely resistant to CheZ, requiring at least 250 times more CheZ than wild-type CheY to achieve the same dephosphorylation rate enhancement, whereas CheY59NA was CheZ sensitive. However, several independent approaches demonstrated that CheY59NR bound tightly to CheZ. A submicromolar K(d) for CheZ binding to CheY59NR-P or CheY.BeF(3)(-) was inferred from fluorescence anisotropy measurements of fluoresceinated-CheZ. A complex between CheY59NR-P and CheZ was isolated by analytical gel filtration, and the elution position from the column was indistinguishable from that of the CheZ dimer. Therefore, we were not able to detect large CheY-P.CheZ complexes that have been inferred using other methods. Possible structural explanations for the specific inhibition of CheZ activity as a result of the arginyl substitution at CheY position 59 are discussed. PMID- 11278902 TI - Marked differences between metalloproteases meprin A and B in substrate and peptide bond specificity. AB - Meprin A and B are highly regulated, secreted, and cell-surface metalloendopeptidases that are abundantly expressed in the kidney and intestine. Meprin oligomers consist of evolutionarily related alpha and/or beta subunits. The work herein was carried out to identify bioactive peptides and proteins that are susceptible to hydrolysis by mouse meprins and kinetically characterize the hydrolysis. Gastrin-releasing peptide fragment 14-27 and gastrin 17, regulatory molecules of the gastrointestinal tract, were found to be the best peptide substrates for meprin A and B, respectively. Peptide libraries and a variety of naturally occurring peptides revealed that the meprin beta subunit has a clear preference for acidic amino acids in the P1 and P1' sites of substrates. The meprin alpha subunit selected for small (e.g. serine, alanine) or hydrophobic (e.g. phenylalanine) residues in the P1 and P1' sites, and proline was the most preferred amino acid at the P2' position. Thus, although the meprin alpha and beta subunits share 55% amino acid identity within the protease domain and are normally localized at the same tissue cell surfaces, they have very different substrate and peptide bond specificities indicating different functions. Homology models of the mouse meprin alpha and beta protease domains, based on the astacin crystal structure, revealed active site differences that can account for the marked differences in substrate specificity of the two subunits. PMID- 11278904 TI - Dissecting a charged network at the active site of orotidine-5'-phosphate decarboxylase. AB - The crystal structure of yeast orotidine-5'-phosphate decarboxylase in complex with the postulated transition state analog, 6-hydroxyuridine-5'-phosphate, reveals contacts between this inhibitor and a novel quartet of charged residues (Lys-59, Asp-91, Lys-93, and Asp-96) within the active site. The structure also suggests a possible interaction between O2 of the 6-hydroxyuridine-5'-phosphate pyrimidine ring and Gln-215. Here we report the results of mutagenesis of each of the charged active site residues and Gln-215. The activities of the Q215A and wild-type enzymes were equal indicating that any interactions between this residue and the pyrimidine ring are dispensable for efficient decarboxylation. For the D91A and K93A mutant enzymes, activity was reduced by more than 5 orders of magnitude and substrate binding could not be detected by isothermal calorimetry. For the D96A mutant enzyme, k(cat) was reduced by more than 5 orders of magnitude, and isothermal calorimetry indicated an 11-fold decrease in the affinity of this enzyme for the substrate in the ground state. For the K59A enzyme, k(cat) was reduced by a factor of 130, and K(m) had increased by a factor of 900. These results indicate that the integrity of the network of charged residues is essential for transition state stabilization. PMID- 11278905 TI - Discrimination between native and non-native disulfides by protein-disulfide isomerase. AB - The folding assistant and chaperone protein-disulfide isomerase (PDI) catalyzes disulfide formation, reduction, and isomerization of misfolded proteins. PDI substrates are not restricted to misfolded proteins; PDI catalyzes the dithiothreitol (DTT)-dependent reduction of native ribonuclease A, microbial ribonuclease, and pancreatic trypsin inhibitor, suggesting that an ongoing surveillance by PDI can test even native disulfides for their ability to rearrange. The mechanism of reduction is consistent with an equilibrium unfolding of the substrate, attack by the nucleophilic cysteine of PDI followed by direct attack of DTT on a covalent intermediate between PDI and the substrate. For native proteins, the rate constants for PDI-catalyzed reduction correlate very well with the rate constants for uncatalyzed reduction by DTT. However, the rate is weakly correlated with disulfide stability, surface exposure, or local disorder in the crystal. Compared with native proteins, scrambled ribonuclease is a much better substrate for PDI than predicted from its reactivity with DTT; however, partially reduced bovine pancreatic trypsin inhibitor (des(14-38)) is not. An extensively unfolded polypeptide may be required by PDI to distinguish native from non-native disulfides. PMID- 11278906 TI - Exposure on cell surface and extensive arginine methylation of ewing sarcoma (EWS) protein. AB - In contrast to the knowledge regarding the function of chimeric Ewing sarcoma (EWS) fusion proteins that arise from chromosomal translocation, the cellular function of the RNA binding EWS protein is poorly characterized. EWS protein had been found mainly in the nucleus. In this report we show that EWS protein is not only found in the nucleus and cytosol but also on cell surfaces. After cell surface biotinylation, isoelectric focusing of membrane fraction, avidin-agarose extraction of biotinylated proteins, and SDS-polyacrylamide gel electrophoresis, EWS protein was identified by matrix-assisted laser desorption ionization and nanoelectrospray tandem mass spectrometry of in-gel-digested peptides. These analyses revealed that the protein, having repeated RGG motifs, is extensively asymmetrically dimethylated on arginine residues, the sites of which have been mapped by mass spectrometric methods. Out of a total of 30 Arg-Gly sequences, 29 arginines were found to be at least partially methylated. The Arg-Gly-Gly sequence was present in 21 of the 29 methylation sites, and in contrast to other methylated proteins, only 11 (38%) methylated arginine residues were found in the Gly-Arg-Gly sequence. The presence of Gly on the C-terminal side of the arginine residue seems to be a prerequisite for recognition by a protein-arginine N methyltransferase (PRMT) catalyzing this asymmetric dimethylation reaction. One monomethylarginine and no symmetrically methylated arginine residue was found. The present findings imply that RNA-binding EWS protein shuttles from the nucleus to the cell surface in a methylated form, the role of which is discussed. PMID- 11278907 TI - Biochemical characterization of Gyp6p, a Ypt/Rab-specific GTPase-activating protein from yeast. AB - Gyp6p from yeast belongs to the GYP family of Ypt/Rab-specific GTPase-activating proteins, and Ypt6p is its preferred substrate (Strom, M., Vollmer, P., Tan, T. J., and Gallwitz, D. (1993) Nature 361, 736-739). We have investigated the kinetic parameters of Gyp6p/Ypt6p interactions and find that Gyp6p accelerates the intrinsic GTPase activity of Ypt6p (0.0002 min(-1)) by a factor of 5 x 10(6) and that they have a very low affinity for its preferred substrate (K(m) = 592 micrometer). Substitution with alanine of several arginines, which Gyp6p shares with other GYP family members, resulted in significant inhibition of GAP activity. Replacement of arginine-155 with either alanine or lysine abolished its GAP activity, indicating a direct involvement of this strictly conserved arginine in catalysis. Physical interaction of the catalytically inactive Gyp6(R155A) mutant GAP with Ypt6 wild-type and Ypt6 mutant proteins could be demonstrated with the two-hybrid system. Short N-terminal and C-terminal truncations of Gyp6p resulted in a complete loss of GAP activity and Ypt6p binding, showing that in contrast to two other Gyp proteins studied previously, most of the 458 amino acid long Gyp6p sequence is required to form a three-dimensional structure that allows substrate binding and catalysis. PMID- 11278908 TI - The Tctex1/Tctex2 class of dynein light chains. Dimerization, differential expression, and interaction with the LC8 protein family. AB - The Tctex1/Tctex2 family of dynein light chains associates with the intermediate chains at the base of the soluble dynein particle. These components are essential for dynein assembly and participate in specific motor-cargo interactions. To further address the role of these light chains in dynein activity, the structural and biochemical properties of several members of this polypeptide class were examined. Gel filtration chromatography and native gel electrophoresis indicate that recombinant Chlamydomonas flagellar Tctex1 exists as a dimer in solution. Furthermore, yeast two-hybrid analysis suggests that this association also occurs in vivo. In contrast, both murine and Chlamydomonas Tctex2 are monomeric. To investigate protein-protein interactions involving these light chains, outer arm dynein from Chlamydomonas flagella was cross-linked using dimethylpimelimidate. Immunoblot analysis of the resulting products revealed the interaction of LC2 (Tctex2) with LC6, which is closely related to the highly conserved LC8 protein found in many enzyme systems, including dynein. Northern dot blot analysis demonstrated that Tctex1/Tctex2 family light chains are differentially expressed both in a tissue-specific and developmentally regulated manner in humans. These data provide further support for the existence of functionally distinct populations of cytoplasmic dynein with differing light chain content. PMID- 11278909 TI - The crystal structure of the inhibitor-complexed carboxypeptidase D domain II and the modeling of regulatory carboxypeptidases. AB - The three-dimensional crystal structure of duck carboxypeptidase D domain II has been solved in a complex with the peptidomimetic inhibitor, guanidinoethylmercaptosuccinic acid, occupying the specificity pocket. This structure allows a clear definition of the substrate binding sites and the substrate funnel-like access. The structure of domain II is the only one available from the regulatory carboxypeptidase family and can be used as a general template for its members. Here, it has been used to model the structures of domains I and III from the former protein and of human carboxypeptidase E. The models obtained show that the overall topology is similar in all cases, the main differences being local and because of insertions in non-regular loops. In both carboxypeptidase D domain I and carboxypeptidase E slightly different shapes of the access to the active site are predicted, implying some kind of structural selection of protein or peptide substrates. Furthermore, emplacement of the inhibitor structure in the active site of the constructed models showed that the inhibitor fits very well in all of them and that the relevant interactions observed with domain II are conserved in domain I and carboxypeptidase E but not in the non-active domain III because of the absence of catalytically indispensable residues in the latter protein. However, in domain III some of the residues potentially involved in substrate binding are well preserved, together with others of unknown roles, which also are highly conserved among all carboxypeptidases. These observations, taken together with others, suggest that domain III might play a role in the binding and presentation of proteins or peptide substrates, such as the pre-S domain of the large envelope protein of duck hepatitis B virus. PMID- 11278910 TI - The Mycobacterium tuberculosis pks2 gene encodes the synthase for the hepta- and octamethyl-branched fatty acids required for sulfolipid synthesis. AB - Multidrug-resistant tuberculosis is a major global health emergency. Cell wall lipids of Mycobacterium tuberculosis can play crucial roles in the pathogenesis. The enzymes involved in their synthesis can be ideal new drug targets against tuberculosis, because many such lipids are unique to this pathogen. A variety of multiple methyl-branched fatty acids are among such unique lipids. We have identified seven genes highly homologous to the mas gene, which is known to be involved in the production of one class of such multiple methyl-branched fatty acids. One of these mas-like genes, pks2, was disrupted using a phage-mediated delivery of the disruption construct. Gene disruption by homologous recombination was confirmed by polymerase chain reaction analysis of the flanking regions of the introduced disrupted gene and by Southern analysis. Thin-layer and radio gas chromatographic analyses of lipids derived from [1-14C]propionic acid and gas chromatography/mass spectrometry analysis of the fatty acids and hydroxy fatty acids showed that the pks2 mutant was incapable of producing hepta- and octamethyl phthioceranic acids and hydroxyphthioceranic acids that are the major acyl constituents of sulfolipids. Consequently, pks2 mutant does not produce sulfolipids. Sulfolipid deficiency in pks2 mutant was confirmed by two dimensional thin-layer chromatographic analysis of lipids derived from [1 14C]propionic acid and 35SO4(-2). With this sulfolipid-deficient mutant, it should be possible to test for the postulated important roles for sulfolipids in the pathogenesis of M. tuberculosis. PMID- 11278911 TI - The conserved active site motif A of Escherichia coli DNA polymerase I is highly mutable. AB - Escherichia coli DNA polymerase I participates in DNA replication, DNA repair, and genetic recombination; it is the most extensively studied of all DNA polymerases. Motif A in the polymerase active site has a required role in catalysis and is highly conserved. To assess the tolerance of motif A for amino acid substitutions, we determined the mutability of the 13 constituent amino acids Val(700)-Arg(712) by using random mutagenesis and genetic selection. We observed that every residue except the catalytically essential Asp(705) can be mutated while allowing bacterial growth and preserving wild-type DNA polymerase activity. Hence, the primary structure of motif A is plastic. We present evidence that mutability of motif A has been conserved during evolution, supporting the premise that the tolerance for mutation is adaptive. In addition, our work allows identification of refinements in catalytic function that may contribute to preservation of the wild-type motif A sequence. As an example, we established that the naturally occurring Ile(709) has a previously undocumented role in supporting sugar discrimination. PMID- 11278912 TI - Phospholipase D activation by norepinephrine is mediated by 12(s)-, 15(s)-, and 20-hydroxyeicosatetraenoic acids generated by stimulation of cytosolic phospholipase a2. tyrosine phosphorylation of phospholipase d2 in response to norepinephrine. AB - Norepinephrine (NE) stimulates phospholipase D (PLD) through a Ras/MAPK pathway in rabbit vascular smooth muscle cells (VSMC). NE also activates calcium influx and calmodulin (CaM)-dependent protein kinase II-dependent cytosolic phospholipase A(2) (cPLA(2)). Arachidonic acid (AA) released by cPLA(2)-catalyzed phospholipid hydrolysis is then metabolized into hydroxyeicosatetraenoic acids (HETEs) through lipoxygenase and cytochrome P450 4A (CYP4A) pathways. HETEs, in turn, have been shown to stimulate Ras translocation and to increase MAPK activity in VSMC. This study was conducted to determine the contribution of cPLA(2)-derived AA and its metabolites (HETEs) to the activation of PLD. NE induced PLD activation was reduced by two structurally distinct CaM antagonists, W-7 and calmidazolium, and by CaM-dependent protein kinase II inhibition. Blockade of cPLA(2) activity or protein depletion with selective cPLA(2) antisense oligonucleotides abolished NE-induced PLD activation. The increase in PLD activity elicited by NE was also blocked by inhibitors of lipoxygenases (baicalein) and CYP4A (17-octadecynoic acid), but not of cyclooxygenase (indomethacin). AA and its metabolites (12(S)-, 15(S)-, and 20-HETEs) increased PLD activity. PLD activation by AA and HETEs was reduced by inhibitors of Ras farnesyltransferase (farnesyl protein transferase III and BMS-191563) and MEK (U0126 and PD98059). These data suggest that HETEs are the mediators of cPLA(2) dependent PLD activation by NE in VSMC. In addition to cPLA(2), PLD was also found to contribute to AA release for prostacyclin production via the phosphatidate phosphohydrolase/diacylglycerol lipase pathway. Finally, a catalytically inactive PLD(2) (but not PLD(1)) mutant inhibited NE-induced PLD activity, and PLD(2) was tyrosine-phosphorylated in response to NE by a MAPK dependent pathway. We conclude that NE stimulates cPLA(2)-dependent PLD(2) through lipoxygenase- and CYP4A-derived HETEs via the Ras/ERK pathway by a mechanism involving tyrosine phosphorylation of PLD(2) in rabbit VSMC. PMID- 11278913 TI - Infection by Trypanosoma cruzi. Identification of a parasite ligand and its host cell receptor. AB - The infective trypomastigote stage of Trypanosoma cruzi expresses a set of surface glycoproteins that are known collectively as Tc85 and belong to the gp85/trans-sialidase supergene family. A member of this family, Tc85-11, with adhesive properties to laminin and cell surfaces was recently cloned. In this report, the Tc85-11 domain for cell binding and its corresponding receptor on epithelial cell LLC-MK(2) are described. Using synthetic peptides corresponding to the Tc85-11 carboxyl-terminal segment, we show that the mammalian cell-binding domain colocalizes to the most conserved motif of the trypanosome gp85/trans sialidase supergene family (VTVXNVFLYNR). Even though Tc85-11 binds to laminin, the 19-residue cell-binding peptide (peptide J) does not contain the laminin binding site, because it does not bind to laminin or inhibit cell binding to this glycoprotein. The host cell receptor for the peptide was characterized as cytokeratin 18. Addition of anti-cytokeratin antibodies to the culture medium significantly inhibited the infection of epithelial cells by T. cruzi. Tc85-11 is a multiadhesive glycoprotein, encoding at least two different binding sites, one for laminin and one for cytokeratin 18, that allow the parasite to overcome the barriers imposed by cell membranes, extracellular matrices, and basal laminae to reach the definitive host cell. This is the first description of a direct interaction between cytokeratin and a protozoan parasite. PMID- 11278914 TI - The heme-regulated eukaryotic initiation factor 2alpha kinase. A potential regulatory target for control of protein synthesis by diffusible gases. AB - Nitric oxide (NO) has been reported to inhibit protein synthesis in eukaryotic cells by increasing the phosphorylation of the alpha-subunit of eukaryotic initiation factor (eIF) 2. However, the mechanism through which this increase occurs has not been characterized. In this report, we examined the effect of the diffusible gases nitric oxide (NO) and carbon monoxide (CO) on the activation of the heme-regulated eIF2alpha kinase (HRI) in rabbit reticulocyte lysate. Spectral analysis indicated that both NO and CO bind to the N-terminal heme-binding domain of HRI. Although NO was a very potent activator of HRI, CO markedly suppressed NO induced HRI activation. The NO-induced activation of HRI was transduced through the interaction of NO with the N-terminal heme-binding domain of HRI and not through S-nitrosylation of HRI. We postulate that the regulation of HRI activity by diffusible gases may be of wider physiological significance, as we further demonstrate that NO generators increase eIF2alpha phosphorylation levels in NT2 neuroepithelial and C2C12 myoblast cells and activate HRI immunoadsorbed from extracts of these non-erythroid cell lines. PMID- 11278915 TI - Investigation of invariant serine/threonine residues in mevalonate kinase. Tests of the functional significance of a proposed substrate binding motif and a site implicated in human inherited disease. AB - Mevalonate kinase serine/threonine residues have been implicated in substrate binding and inherited metabolic disease. Alignment of >20 mevalonate kinase sequences indicates that Ser-145, Ser-146, Ser-201, and Thr-243 are the only invariant residues with alcohol side chains. These residues have been individually mutated to alanine. Structural integrity of the mutants has been demonstrated by binding studies using fluorescent and spin-labeled ATP analogs. Kinetic characterization of the mutants indicates only modest changes in K(m)((ATP)). K(m) for mevalonate increases by approximately 20-fold for S146A, approximately 40-fold for T243A, and 100-fold for S201A. V(max) changes for S145A, S201A, and T243A are < or =3-fold. Thus, the 65-fold activity decrease associated with the inherited human T243I mutation seems attributable to the nonconservative substitution rather than any critical catalytic function. V(max) for S146A is diminished by 4000-fold. In terms of V/K(MVA), this substitution produces a 10(5)-fold effect, suggesting an active site location and catalytic role for Ser-146. The large k(cat) effect suggests that Ser-146 productively orients ATP during catalysis. K(D(Mg-ATP)) increases by almost 40-fold for S146A, indicating a specific role for Ser-146 in liganding Mg(2+)-ATP. Instead of mapping within a proposed C-terminal ATP binding motif, Ser-146 is situated in a centrally located motif, which characterizes the galactokinase/homoserine kinase/ mevalonate kinase/phosphomevalonate kinase protein family. These observations represent the first functional demonstration that this region is part of the active site in these related phosphotransferases. PMID- 11278916 TI - The integrin alpha 7 cytoplasmic domain regulates cell migration, lamellipodia formation, and p130CAS/Crk coupling. AB - The integrin alpha(7)beta(1) is the major laminin-binding integrin in skeletal, heart, and smooth muscle and is a receptor for laminin-1 and -2. It mediates myoblast migration on laminin-1 and -2 and thus might be involved in muscle development and repair. Previously we have shown that alpha(7)B as well as the alpha(7)A and -C splice variants induce cell motility on laminin when transfected into nonmotile HEK293 cells. In this study we have investigated the role of the cytoplasmic domain of alpha(7) in the laminin-induced signal transduction of alpha(7)beta(1) integrin regulating cell adhesion and migration. Deletion of the cytoplasmic domain did not affect assembly of the mutated alpha(7)Deltacyt/beta(1) heterodimer on the cell surface or adhesion of alpha(7)Deltacyt-transfected cells to laminin. The motility of these cells on the laminin-1/E8 fragment, however, was significantly reduced to the level of mock transfected cells; lamellipodia formation and polarization of the cells were also impaired. Adhesion to the laminin-1/E8 fragment induced tyrosine phosphorylation of the focal adhesion kinase, paxillin, and p130(CAS) as well as the formation of a p130(CAS)-Crk complex in wild-type alpha(7)B-transfected cells. In alpha(7)BDeltacyt cells, however, the extent of p130(CAS) tyrosine formation was reduced and formation of the p130(CAS)-Crk complex was impaired, with unaltered levels of p130(CAS) and Crk protein levels. These findings indicate adhesion dependent regulation of p130(CAS)/Crk complex formation by the cytoplasmic domain of alpha(7)B integrin after cell adhesion to laminin-1/E8 and imply alpha(7)B controlled lamellipodia formation and cell migration through the p130(CAS)/Crk protein complex. PMID- 11278917 TI - Insulin-like growth factor-I (IGF-I) receptor activation rescues UV-damaged cells through a p38 signaling pathway. Potential role of the IGF-I receptor in DNA repair. AB - The activated insulin-like growth factor-I receptor (IGF-IR) is implicated in mitogenesis, transformation, and anti-apoptosis. To investigate the role of the IGF-IR in protection from UV-mimetic-induced DNA damage, 4-nitroquinoline N-oxide (4-NQO) was used. In this study we show that the activation of the IGF-IR is capable of rescuing NWTb3 cells overexpressing normal IGF-IRs from 4-NQO-induced DNA damage as demonstrated by cellular proliferation assays. This action was specific for the IGF-IR since cells expressing dominant negative IGF-IRs were not rescued from 4-NQO UV-mimetic treatment. DNA damage induced by 4-NQO in NWTb3 cells was significantly decreased after IGF-IR activation as measured by comet assay. IGF-I was also able to overcome the cell cycle arrest, observed after 4 NQO treatment, thereby enhancing the ability of NWTb3 cells to enter S phase. Interestingly, the p38 mitogen-activated protein kinase pathway was shown to represent the main signaling pathway involved in the IGF-IR-mediated rescue of UV like damaged cells. The ability of the IGF-IR to induce DNA repair was also demonstrated by infecting NWTb3 cells with UV-irradiated adenovirus. Activation of the IGF-IR resulted in enhanced beta-galactosidase reporter gene activity demonstrating repair of the damaged DNA. This study indicates a direct role of the IGF system in the rescue of damaged cells via DNA repair. PMID- 11278918 TI - Identification and characterization of a mammalian enzyme catalyzing the asymmetric oxidative cleavage of provitamin A. AB - In vertebrates, symmetric versus asymmetric cleavage of beta-carotene in the biosynthesis of vitamin A and its derivatives has been controversially discussed. Recently we have been able to identify a cDNA encoding a metazoan beta,beta carotene-15,15'-dioxygenase from the fruit fly Drosophila melanogaster. This enzyme catalyzes the key step in vitamin A biosynthesis, symmetrically cleaving beta-carotene to give two molecules of retinal. Mutations in the corresponding gene are known to lead to a blind, vitamin A-deficient phenotype. Orthologs of this enzyme have very recently been found also in vertebrates and molecularly characterized. Here we report the identification of a cDNA from mouse encoding a second type of carotene dioxygenase catalyzing exclusively the asymmetric oxidative cleavage of beta-carotene at the 9',10' double bond of beta-carotene and resulting in the formation of beta-apo-10'-carotenal and beta-ionone, a substance known as a floral scent from roses, for example. Besides beta-carotene, lycopene is also oxidatively cleaved by the enzyme. The deduced amino acid sequence shares significant sequence identity with the beta,beta-carotene-15,15' dioxygenases, and the two enzyme types have several conserved motifs. To establish its occurrence in different vertebrates, we then attempted and succeeded in cloning cDNAs encoding this new type of carotene dioxygenase from human and zebrafish as well. As regards their possible role, the apocarotenals formed by this enzyme may be the precursors for the biosynthesis of retinoic acid or exert unknown physiological effects. Thus, in contrast to Drosophila, in vertebrates both symmetric and asymmetric cleavage pathways exist for carotenes, revealing a greater complexity of carotene metabolism. PMID- 11278919 TI - RGD-containing peptides inhibit fibrinogen binding to platelet alpha(IIb)beta3 by inducing an allosteric change in the amino-terminal portion of alpha(IIb). AB - To determine the molecular basis for the insensitivity of rat alpha(IIb)beta(3) to inhibition by RGD-containing peptides, hybrids of human and rat alpha(IIb)beta(3) and chimeras of alpha(IIb)beta(3) in which alpha(IIb) was composed of portions of human and rat alpha(IIb) were expressed in Chinese hamster ovary cells and B lymphocytes, and the ability of the tetrapeptide RGDS to inhibit fibrinogen binding to the various forms of alpha(IIb)beta(3) was measured. These measurements indicated that sequences regulating the sensitivity of alpha(IIb)beta(3) to RGDS are located in the seven amino-terminal repeats of alpha(IIb). Moreover, replacing the first three or four (but not the first two) repeats of rat alpha(IIb) with the corresponding human sequences enhanced sensitivity to RGDS, whereas replacing the first two or three repeats of human alpha(IIb) with the corresponding rat sequences had little or no effect. Nevertheless, RGDS bound to Chinese hamster ovary cells expressing alpha(IIb)beta(3) regardless whether the alpha(IIb) in the heterodimers was human, rat, or a rat-human chimera. These results indicate that the sequences determining the sensitivity of alpha(IIb)beta(3) to RGD-containing peptides are located in the third and fourth amino-terminal repeats of alpha(IIb). Because RGDS binds to both human and rat alpha(IIb)beta(3), the results suggest that differences in RGDS sensitivity result from differences in the allosteric changes induced in these repeats following RGDS binding. PMID- 11278920 TI - Dual growth arrest pathways in astrocytes and astrocytic tumors in response to Raf-1 activation. AB - Normal human fibroblasts have been shown to undergo a p16(Ink4a)-associated senescence-like growth arrest in response to sustained activation of the Ras/Raf/MEK/ERK pathway. We noted a similar p16(Ink4a)-associated, senescence like arrest in normal human astrocytes in response to expression of a conditional form of Raf-1. While HPV16 E7-mediated functional inactivation of the p16(Ink4a)/pRb pathway in astrocytes blocked the p16(Ink4a)-associated growth arrest in response to activation of Raf-1, it also revealed a second p21(Cip1) associated, senescence-associated, beta-galactosidase-independent growth arrest pathway. Importantly, the p21(Cip1)-associated pathway was present not only in normal astrocytes but also in p53-, p14(ARF)-, and p16(Ink4a)/pRb-deficient high grade glioma cells that lacked the p16(Ink4a)-dependent arrest mechanism. These results suggest that normal human cells have redundant arrest pathways, which can be activated by Raf-1, and that even tumors that have dismantled p16(Ink4a) dependent growth arrest pathways are potentially regulated by a second p21(Cip1) dependent growth arrest pathway. PMID- 11278922 TI - Rvb1p and Rvb2p are essential components of a chromatin remodeling complex that regulates transcription of over 5% of yeast genes. AB - Eukaryotic Rvb1p and Rvb2p are two highly conserved proteins related to the helicase subset of the AAA+ family of ATPases. Conditional mutants in both genes show rapid changes in the transcription of over 5% of yeast genes, with a similar number of genes being repressed and activated. Both Rvb1p and Rvb2p are required for maintaining the induced state of many inducible promoters. ATP binding and hydrolysis by Rvb1p and Rvb2p is individually essential in vivo, and the two proteins are associated with each other in a high molecular weight complex that shows ATP-dependent chromatin remodeling activity in vitro. Our findings show that Rvb1p and Rvb2p are essential components of a chromatin remodeling complex and determine genes regulated by the complex. PMID- 11278923 TI - Overlapping destinations for two dual targeted glycyl-tRNA synthetases in Arabidopsis thaliana and Phaseolus vulgaris. AB - In plant mitochondria, some of the tRNAs are encoded by the mitochondrial genome and resemble their prokaryotic counterparts, whereas the remaining tRNAs are encoded by the nuclear genome and imported from the cytosol. Generally, mitochondrial isoacceptor tRNAs all have the same genetic origin. One known exception to this rule is the group of tRNA(Gly) isoacceptors in dicotyledonous plants. A mitochondrion-encoded tRNA(Gly) and at least one nucleus-encoded tRNA(Gly) coexist in the mitochondria of these plants, and both are required to allow translation of all four GGN glycine codons. We have taken advantage of this atypical situation to address the problem of tRNA/aminoacyl-tRNA synthetase coevolution in plants. In this work, we show that two different nucleus-encoded glycyl-tRNA synthetases (GlyRSs) are imported into Arabidopsis thaliana and Phaseolus vulgaris mitochondria. The first one, GlyRS-1, is similar to human or yeast glycyl-tRNA synthetase, whereas the second, GlyRS-2, is similar to Escherichia coli glycyl-tRNA synthetase. Both enzymes are dual targeted, GlyRS-1 to mitochondria and to the cytosol and GlyRS-2 to mitochondria and chloroplasts. Unexpectedly, GlyRS-1 seems to be active in the cytosol but inactive in mitochondrial fractions, whereas GlyRS-2 is likely to glycylate both the organelle-encoded tRNA(Gly) and the imported tRNA(Gly) present in mitochondria. PMID- 11278924 TI - Cleavage of a C-terminal peptide is essential for heptamerization of Clostridium perfringens epsilon-toxin in the synaptosomal membrane. AB - Activation of Clostridium perfringens epsilon-protoxin by tryptic digestion is accompanied by removal of the 13 N-terminal and 22 C-terminal amino acid residues. In this study, we examined the toxicity of four constructs: an epsilon protoxin derivative (PD), in which a factor Xa cleavage site was generated at the C-terminal trypsin-sensitive site; PD without the 13 N-terminal residues (DeltaN PD); PD without the 23 C-terminal residues (DeltaC-PD); and PD without either the N- or C-terminal residues (DeltaNC-PD). A mouse lethality test showed that DeltaN PD was inactive, as is PD, whereas DeltaC-PD and DeltaNC-PD were equally active. DeltaC-PD and DeltaNC-PD, but not the other constructs formed a large SDS resistant complex in rat synaptosomal membranes as demonstrated by SDS polyacrylamide gel electrophoresis. When DeltaNC-PD and DeltaC-PD, both labeled with (32)P and mixed in various ratios, were incubated with membranes, eight distinct high molecular weight bands corresponding to six heteropolymers and two homopolymers were detected on a SDS-polyacrylamide gel, indicating the active toxin forms a heptameric complex. These results indicate that C-terminal processing is responsible for activation of the toxin and that it is essential for its heptamerization within the membrane. PMID- 11278925 TI - Evidence for a central apolipoprotein A-I domain loosely bound to lipids in discoidal lipoproteins that is capable of penetrating the bilayer of phospholipid vesicles. AB - Previous evidence indicated that discoidal reconstituted high density lipoproteins (rHDL) of apolipoprotein A-I (apoA-I) can interact with lipid membranes (Tricerri, M. A., Corsico, B., Toledo, J. D., Garda, H. A., and Brenner, R. R. (1998) Biochim. Biophys. Acta 1391, 67-78). With the aim of studying this interaction, photoactivable reagents and protein cleavage with CNBr and hydroxylamine were used. The generic hydrophobic reagent 3-(trifluoromethyl) 3-(m-[125I]iodophenyl)diazirine gave information on the apoA-I regions in contact with the lipid phase in the rHDL discs. Two protein regions loosely bound to lipids were detected: a C-terminal domain and a central one located between residues 87 and 112. They consist of class Y amphipathic alpha-helices that have a different distribution of the charged residues in their polar faces by comparison with class A helices, which predominate in the rest of the apoA-I molecule. The phospholipid analog 1-O-hexadecanoyl-2-O-[9-[[[2-[125I]iodo-4 (trifluoro-methyl-3-H-diazirin-3-yl)benzyl]oxy]carbonyl]nonanoyl]-sn-glycero-3 phosphocholine, which does not undergo significant exchange between membranes and lipoproteins, was used to identify the apoA-I domain directly involved in the interaction of rHDL discs with membranes. By incubating either rHDL or lipid-free apoA-I with lipid vesicles containing 125I-TID-PC, only the 87-112 apoA-I segment becomes labeled after photoactivation. These results indicate that the central domain formed by two type Y helices swings away from lipid contact in the discoidal lipoproteins and is able to insert into membrane bilayers, a process that may be of great importance for the mechanism of cholesterol exchange between high density lipoproteins and cell membranes. PMID- 11278926 TI - The desymmetrization of bicyclic beta -diketones by an enzymatic retro-Claisen reaction. A new reaction of the crotonase superfamily. AB - The enzyme 6-oxocamphor hydrolase, which catalyzes the desymmetrization of 6 oxocamphor to yield (2R,4S)-alpha-campholinic acid, has been purified with a factor of 35.7 from a wild type strain of Rhodococcus sp. NCIMB 9784 grown on (1R)-(+)-camphor as the sole carbon source. The enzyme has a subunit molecular mass of 28,488 Da by electrospray mass spectrometry and a native molecular mass of approximately 83,000 Da indicating that the active protein is trimeric. The specific activity was determined to be 357.5 units mg(-)1, and the K(m) was determined to be 0.05 mm for the natural substrate. The N-terminal amino acid sequence was obtained from the purified protein, and using this information, the gene encoding the enzyme was cloned. The translation of the gene was found to bear significant homology to the crotonase superfamily of enzymes. The gene is closely associated with an open reading frame encoding a ferredoxin reductase that may be involved in the initial step in the biodegradation of camphor. A mechanism for 6-oxocamphor hydrolase based on sequence homology and the known mechanism of the crotonase enzymes is proposed. PMID- 11278927 TI - Structural characterization of protein kinase A as a function of nucleotide binding. Hydrogen-deuterium exchange studies using matrix-assisted laser desorption ionization-time of flight mass spectrometry detection. AB - Transient state kinetic studies indicate that substrate phosphorylation in protein kinase A is partially rate-limited by conformational changes, some of which may be associated with nucleotide binding (Shaffer, J., and Adams, J. A. (1999) Biochemistry 38, 12072-12079). To assess whether specific structural changes are associated with the binding of nucleotides, hydrogen-deuterium exchange experiments were performed on the enzyme in the absence and presence of ADP. Four regions of the protein are protected from exchange in the presence of ADP. Two regions encompass the catalytic and glycine-rich loops and are integral parts of the active site. Conversely, protection of probes in the C terminus is consistent with nucleotide-induced domain closure. One protected probe encompasses a portion of helix C, a secondary structural element that does not make any direct contacts with the nucleotide but has been reported to undergo segmental motion upon the activation of some protein kinases. The combined data suggest that binding of the nucleotide has distal structural effects that may include stabilizing the closed state of the enzyme and altering the position of a critical helix outside the active site. The latter represents the first evidence that the nucleotide alone can induce changes in helix C in solution. PMID- 11278928 TI - Structural and functional studies of CCAAT/enhancer-binding protein epsilon. AB - CCAAT/enhancer-binding protein (C/EBP) epsilon is a critical transcription factor for differentiation of myeloid cells. Structural and functional relationships of C/EBPepsilon were explored by recombinant protein studies, gene mutation, and transactivation assays. Evidence strongly suggested that C/EBPepsilon does not have disulfide bonds. Transactivation analysis of C/EBPepsilon having mutations of each of three conserved cysteines (C345, C148S, and C280S) indicated that the three mutant proteins had almost the same activity as the wild type. Dimer formation of C/EBPepsilon was not detected using both reducing and non-reducing SDS-polyacrylamide gel electrophoresis with Western blot analysis from either bacterial or mammalian expressed C/EBPepsilon. Furthermore, C/EBPepsilon mutant C280S gave a gel band similar to that for wild type, suggesting that this C terminal, conserved cysteine is not involved in disulfide bond formation in vivo, even though previous data for C/EBPbeta suggested that dimers may form in vitro utilizing this conserved cysteine residue. Mutational studies of conserved residues in the activating domain 1 (ADM1) and ADM2 of the amino region of the gene indicated that negative charge is critical for transactivational activity of C/EBPepsilon. Mutational analyses of hydrophobic amino acids in ADM1 suggested that these residues do not play a key role in transactivational activity. Further mutational studies indicated that, although the N-terminal 32-amino acid peptide of C/EBPepsilon isoform p32 did not greatly influence the transactivation activity compared with p30 isoform, this peptide does modulate transactivation activity. Domain swapping experiments substituting the ADM1 domain of various C/EBPs for C/EBPepsilon showed that the C/EBPalpha and -delta but not -beta ADM1 markedly enhanced the chimeric C/EBPepsilon transcriptional activity. Based on mutational data and possible mRNA structure, we hypothesized about the effect of mRNA structure on translation of the two major C/EBPepsilon isoforms: p32 and p30. The data suggested a very stable 8-base pair double helical structure with one strand sequence including the initial codon for p32 and complementary strand with the initial codon for p30. PMID- 11278929 TI - Heat shock protein-chaperoned peptides but not free peptides introduced into the cytosol are presented efficiently by major histocompatibility complex I molecules. AB - The studies reported here bear on the events in the cytosol that lead to trafficking of peptides during antigen processing and presentation by major histocompatibility complex (MHC) I molecules. We have introduced free antigenic peptides or antigenic peptides bound to serum albumin or to cytosolic heat shock proteins hsp90 (and its endoplasmic reticular homologue gp96) or hsp70 into the cytosol of living cells and have monitored the presentation of the peptides by appropriate MHC I molecules. The experiments show that (i) free peptides or serum albumin-bound peptides, introduced into the cytosol, become ligands of MHC I molecules at a far lower efficiency than peptides chaperoned by any of the heat shock proteins tested and (ii) treatment of cells with deoxyspergualin, a drug that binds hsp70 and hsp90 with apparent specificity, abrogates the ability of cells to present antigenic peptides through MHC I molecules, and introduction of additional hsp70 into the cytosol overcomes this abrogation. These results suggest for the first time a functional role for cytosolic chaperones in antigen processing. PMID- 11278930 TI - Heparin enhances the specificity of antithrombin for thrombin and factor Xa independent of the reactive center loop sequence. Evidence for an exosite determinant of factor Xa specificity in heparin-activated antithrombin. AB - Heparin activates the primary serpin inhibitor of blood clotting proteinases, antithrombin, both by an allosteric conformational change mechanism that specifically enhances factor Xa inactivation and by a ternary complex bridging mechanism that promotes the inactivation of thrombin and other target proteinases. To determine whether the factor Xa specificity of allosterically activated antithrombin is encoded in the reactive center loop sequence, we attempted to switch this specificity by mutating the P6-P3' proteinase binding sequence excluding P1-P1' to a more optimal thrombin recognition sequence. Evaluation of 12 such antithrombin variants showed that the thrombin specificity of the serpin allosterically activated by a heparin pentasaccharide could be enhanced as much as 55-fold by changing P3, P2, and P2' residues to a consensus thrombin recognition sequence. However, at most 9-fold of the enhanced thrombin specificity was due to allosteric activation, the remainder being realized without activation. Moreover, thrombin specificity enhancements were attenuated to at most 5-fold with a bridging heparin activator. Surprisingly, none of the reactive center loop mutations greatly affected the factor Xa specificity of the unactivated serpin or the several hundred-fold enhancement in factor Xa specificity due to activation by pentasaccharide or bridging heparins. Together, these results suggest that the specificity of both native and heparin-activated antithrombin for thrombin and factor Xa is only weakly dependent on the P6-P3' residues flanking the primary P1-P1' recognition site in the serpin-reactive center loop and that heparin enhances serpin specificity for both enzymes through secondary interaction sites outside the P6-P3' region, which involve a bridging site on heparin in the case of thrombin and a previously unrecognized exosite on antithrombin in the case of factor Xa. PMID- 11278931 TI - A novel osteoblast-derived C-type lectin that inhibits osteoclast formation. AB - We have cloned and expressed murine osteoclast inhibitory lectin (mOCIL), a 207 amino acid type II transmembrane C-type lectin. In osteoclast formation assays of primary murine calvarial osteoblasts with bone marrow cells, antisense oligonucleotides for mOCIL increased tartrate-resistant acid phosphatase-positive mononucleate cell formation by 3-5-fold, whereas control oligonucleotides had no effect. The extracellular domain of mOCIL, expressed as a recombinant protein in Escherichia coli, dose-dependently inhibited multinucleate osteoclast formation in murine osteoblast and spleen cell co-cultures as well as in spleen cell cultures treated with RANKL and macrophage colony-stimulating factor. Furthermore, mOCIL acted directly on macrophage/monocyte cells as evidenced by its inhibitory action on adherent spleen cell cultures, which were depleted of stromal and lymphocytic cells. mOCIL completely inhibited osteoclast formation during the proliferative phase of osteoclast formation and resulted in 70% inhibition during the differentiation phase. Osteoblast OCIL mRNA expression was enhanced by parathyroid hormone, calcitriol, interleukin-1alpha and -11, and retinoic acid. In rodent tissues, Northern blotting, in situ hybridization, and immunohistochemistry demonstrated OCIL expression in osteoblasts and chondrocytes as well as in a variety of extraskeletal tissues. The overlapping tissue distribution of OCIL mRNA and protein with that of RANKL strongly suggests an interaction between these molecules in the skeleton and in extraskeletal tissues. PMID- 11278932 TI - Replication factors MCM2 and ORC1 interact with the histone acetyltransferase HBO1. AB - The minichromosome maintenance (MCM) proteins, together with the origin recognition complex (ORC) proteins and Cdc6, play an essential role in eukaryotic DNA replication through the formation of a pre-replication complex at origins of replication. We used a yeast two-hybrid screen to identify MCM2-interacting proteins. One of the proteins we identified is identical to the ORC1-interacting protein termed HBO1. HBO1 belongs to the MYST family, characterized by a highly conserved C2HC zinc finger and a putative histone acetyltransferase domain. Biochemical studies confirmed the interaction between MCM2 and HBO1 in vitro and in vivo. An N-terminal domain of MCM2 is necessary for binding to HBO1, and a C2HC zinc finger of HBO1 is essential for binding to MCM2. A reverse yeast two hybrid selection was performed to isolate an allele of MCM2 that is defective for interaction with HBO1; this allele was then used to isolate a suppressor mutant of HBO1 that restores the interaction with the mutant MCM2. This suppressor mutation was located in the HBO1 zinc finger. Taken together, these findings strongly suggest that the interaction between MCM2 and HBO1 is direct and mediated by the C2HC zinc finger of HBO1. The biochemical and genetic interactions of MYST family protein HBO1 with two components of the replication apparatus, MCM2 and ORC1, suggest that HBO1-associated HAT activity may play a direct role in the process of DNA replication. PMID- 11278933 TI - ATF-7, a novel bZIP protein, interacts with the PRL-1 protein-tyrosine phosphatase. AB - We have identified a novel basic leucine zipper (bZIP) protein, designated ATF-7, that physically interacts with the PRL-1 protein-tyrosine phosphatase (PTPase). PRL-1 is a predominantly nuclear, farnesylated PTPase that has been linked to the control of cellular growth and differentiation. This interaction was initially found using the yeast two-hybrid system. ATF-7 is most closely related to members of the ATF/CREB family of bZIP proteins, with highest homology to ATF-4. ATF-7 homodimers can bind specifically to CRE elements. ATF-7 is expressed in a number of different tissues and is expressed in association with differentiation in the Caco-2 cell model of intestinal differentiation. We have confirmed the PRL-1.ATF 7 interaction and mapped the regions of ATF-7 and PRL-1 important for interaction to ATF-7's bZIP region and PRL-1's phosphatase domain. Finally, we have determined that PRL-1 is able to dephosphorylate ATF-7 in vitro. Further insight into ATF-7's precise cellular roles, transcriptional function, and downstream targets are likely be of importance in understanding the mechanisms underlying the complex processes of maintenance, differentiation, and turnover of epithelial tissues. PMID- 11278934 TI - The M1 receptor is required for muscarinic activation of mitogen-activated protein (MAP) kinase in murine cerebral cortical neurons. AB - Muscarinic acetylcholine receptors (mAChR) in the central nervous system are involved in learning and memory, epileptic seizures, and processing the amyloid precursor protein. The M(1) receptor is the predominant mAChR subtype in the cortex and hippocampus. Although the five mAChR fall into two broad functional groups, all five subtypes, when expressed in recombinant systems, can activate the mitogen-activated protein kinase (MAPK) pathway. The MAPK pathway has been implicated in learning and memory, amyloid protein processing, and neuronal plasticity. We used M(1) knock-out mice to determine the role of this receptor subtype in signal transduction in the mouse forebrain. In primary cortical cultures from mice lacking the M(1) mAChR, agonist-stimulated phosphoinositide hydrolysis was reduced by more than 60% compared with cultures from wild type mice. Although muscarinic agonists induced robust activation of MAPK in cortical cultures from wild type mice, mAChR-mediated activation of MAPK was virtually absent in cultures from M(1)-deficient mice. These results indicate that the M(1) mAChR is the major subtype that mediates activation of phospholipase C and MAPK in mouse forebrain. PMID- 11278935 TI - Physiological levels of mammalian uncoupling protein 2 do not uncouple yeast mitochondria. AB - We assessed the ability of human uncoupling protein 2 (UCP2) to uncouple mitochondrial oxidative phosphorylation when expressed in yeast at physiological and supraphysiological levels. We used three different inducible UCP2 expression constructs to achieve mitochondrial UCP2 expression levels in yeast of 33, 283, and 4100 ng of UCP2/mg of mitochondrial protein. Yeast mitochondria expressing UCP2 at 33 or 283 ng/mg showed no increase in proton conductance, even in the presence of various putative effectors, including palmitate and all-trans retinoic acid. Only when UCP2 expression in yeast mitochondria was increased to 4 microg/mg, more than an order of magnitude greater than the highest known physiological concentration, was proton conductance increased. This increased proton conductance was not abolished by GDP. At this high level of UCP2 expression, an inhibition of substrate oxidation was observed, which cannot be readily explained by an uncoupling activity of UCP2. Quantitatively, even the uncoupling seen at 4 microgram/mg was insufficient to account for the basal proton conductance of mammalian mitochondria. These observations suggest that uncoupling of yeast mitochondria by UCP2 is an overexpression artifact leading to compromised mitochondrial integrity. PMID- 11278936 TI - Molecular cloning and characterization of a novel mammalian endo-apyrase (LALP1). AB - Here we describe the cloning, localization, and characterization of a novel mammalian endo-apyrase (LALP1) in human and mouse. The predicted human LALP1 gene encodes a 604-amino acid protein, whereas the mouse Lalp1 gene encodes a 606 amino acid protein. The human and mouse genes have 88% amino acid sequence identity. These genes share considerable homologies with hLALP70, a recently discovered mammalian lysosomal endo-apyrase. The human LALP1 gene resides on chromosome 10q23-q24 and contains 12 exons and 11 introns covering a genomic region of approximately 46 kilobase pairs. The subcellular localization and enzymatic activity of LALP1 indicated that LALP1 is indeed an endo-apyrase with substrate preference for nucleoside triphosphates UTP, GTP, and CTP. PMID- 11278937 TI - Basic fibroblast growth factor-induced proliferation of primary astrocytes. evidence for the involvement of sphingomyelin biosynthesis. AB - We recently reported that the marked decrease in cellular ceramide in primary astrocytes is an early event associated with the mitogenic activity of basic fibroblast growth factor (bFGF) (Riboni, L., Viani, P., Bassi, R., Stabieini, A., and Tettamanti, G. (2000) GLIA 32, 137-145). Here we show that a rapid activation of sphingomyelin biosynthesis appears to be the major mechanism responsible for the fall in ceramide levels induced by bFGF. When quiescent astrocytes were treated with bFGF, an increased amount of newly synthesized ceramide (from either l-[(3)H]serine or [(3)H]sphingosine) was directed toward the biosynthesis of sphingomyelin. Conversely, bFGF did not appear to affect ceramide levels by other metabolic pathways involved in ceramide turnover such as sphingomyelin degradation and ceramide biosynthesis, degradation, and glucosylation. Enzymatic studies demonstrating a relevant and rapid increase in sphingomyelin synthase activity after bFGF treatment have provided a convincing explanation for the activation of sphingomyelin biosynthesis. The bFGF-induced increase in sphingomyelin synthase appears to depend on a post-translational activation mechanism. Moreover, in the presence of brefeldin A, the activation of sphingomyelin biosynthesis was abolished, suggesting that the enzyme is located in a compartment other than the Golgi apparatus. Also the phosphatidylcholine specific phospholipase C inhibitor D609 exerted a potent inhibitory effect on sphingomyelin biosynthesis. Finally, we demonstrate that inhibition of sphingomyelin biosynthesis by brefeldin A or D609 led to a significant inhibition of bFGF-stimulated mitogenesis. All this supports that, in primary astrocytes, the early activation of sphingomyelin synthase is involved in the bFGF signaling pathway leading to proliferation. PMID- 11278938 TI - Role of the phospholipase C-inositol 1,4,5-trisphosphate pathway in calcium release-activated calcium current and capacitative calcium entry. AB - We investigated the putative roles of phospholipase C, polyphosphoinositides, and inositol 1,4,5-trisphosphate (IP(3)) in capacitative calcium entry and calcium release-activated calcium current (I(crac)) in lacrimal acinar cells, rat basophilic leukemia cells, and DT40 B-lymphocytes. Inhibition of phospholipase C with blocked calcium entry and I(crac) activation whether in response to a phospholipase C-coupled agonist or to calcium store depletion with thapsigargin. Run-down of cellular polyphosphoinositides by concentrations of wortmannin that block phosphatidylinositol 4-kinase completely blocked calcium entry and I(crac). The membrane-permeant IP(3) receptor inhibitor, 2-aminoethoxydiphenyl borane, blocked both capacitative calcium entry and I(crac). However, it is likely that 2 aminoethoxydiphenyl borane does not inhibit through an action on the IP(3) receptor because the drug was equally effective in wild-type DT40 B-cells and in DT40 B-cells whose genes for all three IP(3) receptors had been disrupted. Intracellular application of another potent IP(3) receptor antagonist, heparin, failed to inhibit activation of I(crac). Finally, the inhibition of I(crac) activation by or wortmannin was not reversed or prevented by direct intracellular application of IP(3). These findings indicate a requirement for phospholipase C and for polyphosphoinositides for activation of capacitative calcium entry. However, the results call into question the previously suggested roles of IP(3) and IP(3) receptor in this mechanism, at least in these particular cell types. PMID- 11278939 TI - Intra-endosomal pH-sensitive recruitment of the Arf-nucleotide exchange factor ARNO and Arf6 from cytoplasm to proximal tubule endosomes. AB - Kidney proximal tubule epithelial cells have an extensive apical endocytotic apparatus that is critical for the reabsorption and degradation of proteins that traverse the glomerular filtration barrier and that is also involved in the extensive recycling of functionally important apical plasma membrane transporters. We show here that an Arf-nucleotide exchange factor, ARNO (ADP ribosylation factor nucleotide site opener) as well as Arf6 and Arf1 small GTPases are located in the kidney proximal tubule receptor-mediated endocytosis pathway, and that ARNO and Arf6 recruitment from cytosol to endosomes is pH dependent. In proximal tubules in situ, ARNO and Arf6 partially co-localized with the V-ATPase in apical endosomes in proximal tubules. Arf1 was localized both at the apical pole of proximal tubule epithelial cells, but also in the Golgi. By Western blot analysis ARNO, Arf6, and Arf1 were detected both in purified endosomes and in proximal tubule cytosol. A translocation assay showed that ATP driven endosomal acidification triggered the recruitment of ARNO and Arf6 from proximal tubule cytosol to endosomal membranes. The translocation of both ARNO and Arf6 was reversed by V-type ATPase inhibitors and by uncouplers of endosomal intralumenal pH, and was correlated with the magnitude of intra-endosomal acidification. Our data suggest that V-type ATPase-dependent acidification stimulates the selective recruitment of ARNO and Arf6 to proximal tubule early endosomes. This mechanism may play an important role in the pH-dependent regulation of receptor-mediated endocytosis in proximal tubules in situ. PMID- 11278940 TI - c-Src tyrosine kinase binds the beta 2-adrenergic receptor via phospho-Tyr-350, phosphorylates G-protein-linked receptor kinase 2, and mediates agonist-induced receptor desensitization. AB - The nonreceptor tyrosine kinase Src has been implicated in the switching of signaling of beta2-adrenergic receptors from adenylylcyclase coupling to the mitogen-activated protein kinase pathway. In the current work, we demonstrate that Src plays an active role in the agonist-induced desensitization of beta2 adrenergic receptors. Both the expression of dominant-negative Src and treatment with the 4-amine-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) inhibitor of Src kinase activity blocks agonist-induced desensitization. Agonist triggers tyrosine phosphorylation of the beta2-adrenergic receptor and recruitment and activation of Src. Because phosphorylation of the Tyr-350 residue of the beta2-adrenergic receptor creates a conditional, canonical SH2-binding site on the receptor, we examined the effect of the Y350F mutation on Src phosphorylation, Src recruitment, and desensitization. Mutant beta2-adrenergic receptors with a Tyr-to-Phe substitution at Tyr-350 do not display agonist induced desensitization, Src recruitment, or Src activation. Downstream of binding to the receptor, Src phosphorylates and activates G-protein-linked receptor kinase 2 (GRK2), a response obligate for agonist-induced desensitization. Constitutively active Src increases GRK phosphorylation, whereas either expression of dominant-negative Src or treatment with the PP2 inhibitor abolishes tyrosine phosphorylation of GRK and desensitization. Thus, in addition to its role in signal switching to the mitogen-activated protein kinase pathway, Src recruitment to the beta2-adrenergic receptor and activation are obligate for normal agonist-induced desensitization. PMID- 11278941 TI - DNA binding by the ETS-domain transcription factor PEA3 is regulated by intramolecular and intermolecular protein.protein interactions. AB - The control of DNA binding by eukaryotic transcription factors represents an important regulatory mechanism. Many transcription factors are controlled by cis acting autoinhibitory modules that are thought to act by blocking promiscuous DNA binding in the absence of appropriate regulatory cues. Here, we have investigated the determinants and regulation of the autoinhibitory mechanism employed by the ETS-domain transcription factor, PEA3. DNA binding is inhibited by a module composed of a combination of two short motifs located on either side of the ETS DNA-binding domain. A second type of protein, Ids, can act in trans to mimic the effect of these cis-acting inhibitory motifs and reduce DNA binding by PEA3. By using a one-hybrid screen, we identified the basic helix-loop-helix-leucine zipper transcription factor USF-1 as an interaction partner for PEA3. PEA3 and USF-1 form DNA complexes in a cooperative manner. Moreover, the formation of ternary PEA3.USF-1.DNA complexes requires parts of the same motifs in PEA3 that form the autoinhibitory module. Thus the binding of USF-1 to PEA3 acts as a switch that modifies the autoinhibitory motifs in PEA3 to first relieve their inhibitory action, and second, promote ternary nucleoprotein complex assembly. PMID- 11278943 TI - The use of prothrombin(S525C) labeled with fluorescein to directly study the inhibition of prothrombinase by antithrombin during prothrombin activation. AB - Serine 525 of human prothrombin was mutated to cysteine and covalently labeled with fluorescein to make II(S525C)-fluorescein. Kinetics of cleavage of this derivative by prothrombinase are identical to those of wild-type prothrombin. Cleavage is coincident with a 50% increase in fluorescence intensity and the product is catalytically inactive. Thus, it allows convenient monitoring of prothrombin activation without generating active thrombin. The kinetics of inhibition of factor Xa (FXa) by antithrombin (AT) and AT-heparin were measured by monitoring activation of II(S525C)-fluorescein and the hydrolysis of the chromogenic substrate S2222 in the presence of AT. With S2222 as the substrate the rate constant for inhibition of FXa, Ca(2+), and unilamellar vesicles of phosphatidylcholine and phosphatidylserine (75:25) (PCPS) vesicles by AT was 3.51 x 10(3) m(-1) s(-1); when factor Va (FVa) was included the rate constant was 1.55 x 10(3) m(-1) s(-1). In the absence of FVa, II(S525C)-fluorescein had no effect on inhibition. When II(S525C)-fluorescein was the substrate, however, FVa at saturating concentrations profoundly protected FXa from inhibition by AT, increasing the half-life from 3 min with FXa, Ca(2+), PCPS, and II(S525C) fluorescein, to greater than 69 min when FVa was included. Thus, both FVa and prothrombin are necessary for this level of protection. In the absence of prothrombin, FVa decreased the second order rate constant for inhibition by the AT-heparin complex from 1.58 x 10(7) m(-1) s(-1), for FXa, Ca(2+), and PCPS, to 7.72 x 10(6) m(-1) s(-1). II(S525C)-fluorescein and factor Va together reduced the rate constant to less than 1% of that for FXa, Ca(2+), and PCPS. At a heparin concentration of 0.2 unit/ml, this corresponds to a half-life increase from 1 s to 136 s. PMID- 11278942 TI - Identification of Barx2b, a serum response factor-associated homeodomain protein. AB - CC(A/T)(6)GG or serum response elements represent a common regulatory motif important for regulating the expression of many smooth muscle-specific genes. They are multifunctional elements that bind serum response factor (SRF) and are important for serum induction of genes, expression of muscle-specific genes, and differentiation of vascular smooth muscle cells. In the current study, a yeast two-hybrid screen was used to identify proteins from mouse intestine that interact with SRF. A novel homeodomain-containing transcription factor, called Barx2b, was identified that specifically interacts with SRF and promotes the DNA binding activity of SRF. Northern blotting, RNase protection analysis, and Western blotting revealed that Barx2b mRNA and protein are expressed in several smooth muscle-containing tissues, as well as in skeletal muscle and brain. In vitro binding studies using bacterial fusion proteins revealed that the DNA binding domain of SRF interacts with a region of Barx2b located amino-terminal of the homeobox domain. The results of these studies support the hypothesis that interaction of SRF with different homeodomain-containing proteins may play a critical role in determining the cell-specific functions of SRF. PMID- 11278944 TI - Two novel Xenopus homologs of mammalian LP(A1)/EDG-2 function as lysophosphatidic acid receptors in Xenopus oocytes and mammalian cells. AB - Lysophosphatidic acid (LPA) induces diverse biological responses in many types of cells and tissues by activating its specific G protein-coupled receptors (GPCRs). Previously, three cognate LPA GPCRs (LP(A1)/VZG-1/EDG-2, LP(A2)/EDG-4, and LP(A3)/EDG-7) were identified in mammals. By contrast, an unrelated GPCR, PSP24, was reported to be a high affinity LPA receptor in Xenopus laevis oocytes, raising the possibility that Xenopus uses a very different form of LPA signaling. Toward addressing this issue, we report two novel Xenopus genes, xlp(A1)-1 and xlp(A1)-2, encoding LP(A1) homologs (approximately 90% amino acid sequence identity with mammalian LP(A1)). Both xlp(A1)-1 and xlp(A1)-2 are expressed in oocytes and the nervous system. Overexpression of either gene in oocytes potentiated LPA-induced oscillatory chloride ion currents through a pertussis toxin-insensitive pathway. Injection of antisense oligonucleotides designed to inhibit xlp(A1)-1 and xlp(A1)-2 expression in oocytes eliminated their endogenous response to LPA. Furthermore, retrovirus-mediated heterologous expression of xlp(A1)-1 or xlp(A1)-2 in B103 rat neuroblastoma cells that are unresponsive to LPA conferred LPA-induced cell rounding and adenylyl cyclase inhibition. These results indicate that XLP(A1)-1 and XLP(A1)-2 are functional Xenopus LPA receptors and demonstrate the evolutionary conservation of LPA signaling over a range of vertebrate phylogeny. PMID- 11278945 TI - Isolation and characterization of EMILIN-2, a new component of the growing EMILINs family and a member of the EMI domain-containing superfamily. AB - EMILIN (elastin microfibril interfase located Protein) is an elastic fiber associated glycoprotein consisting of a self-interacting globular C1q domain at the C terminus, a short collagenous stalk, an extended region of potential coiled coil structure, and an N-terminal cysteine-rich domain (EMI domain). Using the globular C1q domain as a bait in the yeast two-hybrid system, we have isolated a cDNA encoding a novel protein. Determination of the entire primary structure demonstrated that this EMILIN-binding polypeptide is highly homologous to EMILIN. The domain organization is superimposable, one important difference being a proline-rich (41%) segment of 56 residues between the potential coiled-coil region and the collagenous domain absent in EMILIN. The entire gene (localized on chromosome 18p11.3) was isolated from a BAC clone, and it is structurally almost identical to that of EMILIN (8 exons, 7 introns with identical phases at the exon/intron boundaries) but much larger (about 40 versus 8 kilobases) than that of EMILIN. Given these findings we propose to name the novel protein EMILIN-2 and the prototype member of this family EMILIN-1 (formerly EMILIN). The mRNA expression of EMILIN-2 is more restricted compared with that of EMILIN-1; highest levels are present in fetal heart and adult lung, whereas, differently from EMILIN-1, adult aorta, small intestine, and appendix show very low expression, and adult uterus and fetal kidney are negative. Finally, the EMILIN-2 protein is secreted extracellularly by in vitro-grown cells, and in accordance with the partial coexpression in fetal and adult tissues, the two proteins shown extensive but not absolute immunocolocalization in vitro. PMID- 11278946 TI - X-ray absorption studies of human matrix metalloproteinase-2 (MMP-2) bound to a highly selective mechanism-based inhibitor. comparison with the latent and active forms of the enzyme. AB - Malignant tumors express high levels of zinc-dependent endopeptidases called matrix metalloproteinases (MMPs), which are thought to facilitate tumor metastasis and angiogenesis by hydrolyzing components of the extracellular matrix. Of these enzymes, gelatinases A (MMP-2) and B (MMP-9), have especially been implicated in malignant processes, and thus, they have been a target for drugs designed to block their activity. Therefore, understanding their molecular structure is key for a rational approach to inhibitor design. Here, we have conducted x-ray absorption spectroscopy of the full-length human MMP-2 in its latent, active, and inhibited states and report the structural changes at the zinc ion site upon enzyme activation and inhibition. We have also examined the molecular structure of MMP-2 in complex with SB-3CT, a recently reported novel mechanism-based synthetic inhibitor that was designed to be highly selective in gelatinases. It is shown that SB-3CT directly binds the catalytic zinc ion of MMP 2. Interestingly, the novel mode of binding of the inhibitor to the catalytic zinc reconstructs the conformational environment around the active site metal ion back to that of the proenzyme. PMID- 11278947 TI - Scavenger receptor class B type I-mediated reverse cholesterol transport is inhibited by advanced glycation end products. AB - Cellular interactions of advanced glycation end products (AGE) are mediated by AGE receptors. We demonstrated previously that class A scavenger receptor types I and II (SR-A) and CD36, a member of class B scavenger receptor family, serve as the AGE receptors. In this study, we investigated whether scavenger receptor class B type I (SR-BI), another receptor belonging to class B scavenger receptor family, was also an AGE receptor. We used Chinese hamster ovary (CHO) cells overexpressed hamster SR-BI (CHO-SR-BI cells). (125)I-AGE-bovine serum albumin (AGE-BSA) was endocytosed in a dose-dependent fashion and underwent lysosomal degradation by CHO-SR-BI cells. (125)I-AGE-BSA exhibited saturable binding to CHO SR-BI cells (K(d) = 8.3 microg/ml). Endocytic uptake of (125)I-AGE-BSA by CHO-SR BI cells was completely inhibited by oxidized low density lipoprotein (LDL) and acetylated LDL, whereas LDL exerted only a weak inhibitory effect (<20%). Cross competition experiments showed that AGE-BSA had no effect on HDL binding to these cells and vice versa. Interestingly, however, SR-BI-mediated selective uptake of HDL-CE was completely inhibited by AGE-BSA in a dose-dependent manner (IC(50) <10 microg/ml). Furthermore, AGE-BSA partially inhibited (by <30%) the selective uptake of HDL-CE in human hepatocarcinoma HepG2 cells (IC(50) <30 microg/ml). In addition, [(3)H]cholesterol efflux from CHO-SR-BI cells to HDL was significantly inhibited by AGE-BSA in a dose-dependent manner (IC(50) <30 microg/ml). Our results indicate that AGE proteins, as ligands for SR-BI, effectively inhibit both SR-BI-mediated selective uptake of HDL-CE and cholesterol efflux from peripheral cells to HDL, suggesting that AGE proteins might modulate SR-BI mediated cholesterol metabolism in vivo. PMID- 11278948 TI - The basic helix-loop-helix protein, sharp-1, represses transcription by a histone deacetylase-dependent and histone deacetylase-independent mechanism. AB - Many aspects of neurogenesis and neuronal differentiation are controlled by basic helix-loop-helix (bHLH) proteins. One such factor is SHARP-1, initially identified on the basis of its sequence similarity to hairy. Unlike hairy, and atypically for bHLHs, SHARP-1 is expressed late in development, suggestive of a role in terminal aspects of differentiation. Nevertheless, the role of SHARP-1 and the identity of its target genes remain unknown. During the course of a one hybrid screen for transcription factors that bind to regulatory domains of the M1 muscarinic acetylcholine receptor gene, we isolated the bHLH transcription factor SHARP-1. In this study, we investigated the functional role of SHARP-1 in regulating transcription. Fusion proteins of SHARP-1 tethered to the gal4 DNA binding domain repress both basal and activated transcription when recruited to either a TATA-containing or a TATAless promoter. Furthermore, we identified two independent repression domains that operate via distinct mechanisms. Repression by a domain in the C terminus is sensitive to the histone deacetylase inhibitor trichostatin A, whereas repression by the bHLH domain is insensitive to TSA. Furthermore, overexpression of SHARP-1 represses transcription from the M(1) promoter. This study represents the first report to assign a function to, and to identify a target gene for, the bHLH transcription factor SHARP-1. PMID- 11278949 TI - Binding of 13-HODE and 15-HETE to phospholipid bilayers, albumin, and intracellular fatty acid binding proteins. implications for transmembrane and intracellular transport and for protection from lipid peroxidation. AB - Transport and utilization of fatty acids (FA) in cells is a multistep process that includes adsorption to and movement across the plasma membrane and binding to intracellular fatty acid binding proteins (FABP) in the cytosol. We monitored the transbilayer movement of several polyunsaturated FA and oxidation products (13-hydroxy octadecadienoic acid (HODE) and 15-hydroxytetraenoic acid (HETE)) in unilamellar protein-free phospholipid vesicles containing a fluorescent pH probe. All FA diffused rapidly by the flip-flop mechanism across the model membrane, as revealed by pH changes inside the vesicle. This result suggests that FA oxidation products generated in the cell could cross the plasma or nuclear membrane spontaneously without a membrane transporter. To illuminate features of extra- and intracellular transport, the partitioning of unsaturated FA and oxidized FA between phospholipid vesicles and albumin or FABP was studied by the pyranin assay. These experiments showed that all polyunsaturated FA and oxidized FA (13 HODE and 15-HETE) desorbed rapidly from the phospholipid bilayer to bind to bovine serum albumin, which showed a slight preference for the unsaturated FA over the oxidized FA. FABP rapidly bound FA in the presence of phospholipid bilayers, with a preference of 13-HODE over the unsaturated FA and with a specificity depending on the type of FABP. Liver FABP was significantly more effective than intestinal FABP in binding 13-HODE in the presence of vesicles. The more effective binding of the FA metabolite, 13-HODE, than its precursor 18:2 by FABP may help protect cellular membranes from potential damage by monohydroxy fatty acids and may contribute a pathway for entry of 13-HODE into the nucleus. PMID- 11278950 TI - Phosphorylation by cdc2-CyclinB1 kinase releases cytoplasmic dynein from membranes. AB - Movement of various cargoes toward microtubule minus ends is driven by the microtubule motor cytoplasmic dynein (CD). Many cargoes are motile only during certain cell cycle phases, suggesting that CD function may be under cell cycle control. Phosphorylation of the CD light intermediate chain (DLIC) has been suggested to play a crucial role in modulating CD function during the Xenopus embryonic cell cycle, where CD-driven organelle movement is active in interphase but greatly reduced in metaphase. This down-regulation correlates with hyperphosphorylation of DLIC and release of CD from the membrane. Here we investigate the role of the key mitotic kinase, cdc2-cyclinB1, in this process. We show that DLIC within the native Xenopus CD complex is an excellent substrate for purified Xenopus cdc2-glutathione S-transferase (GST) cyclinB1 (cdc2 GSTcyclinB1) kinase. Mass spectrometry of native DLIC revealed that a conserved cdc2 site (Ser-197) previously implicated in the metaphase modulation of CD remains phosphorylated in interphase and so is unlikely to be the key regulatory site. We also demonstrate that incubating interphase membranes with cdc2 GSTcyclinB1 kinase results in substantial release of CD from the membrane. These data suggest that phosphorylation of DLIC by cdc2 kinase leads directly to the loss of membrane-associated CD and an inhibition of organelle movement. PMID- 11278951 TI - Ca2+-independent smooth muscle contraction. a novel function for integrin-linked kinase. AB - Smooth muscle contraction follows an increase in cytosolic Ca(2+) concentration, activation of myosin light chain kinase, and phosphorylation of the 20-kDa light chain of myosin at Ser(19). Several agonists acting via G protein-coupled receptors elicit a contraction without a change in [Ca(2+)](i) via inhibition of myosin light chain phosphatase and increased myosin phosphorylation. We showed that microcystin (phosphatase inhibitor)-induced contraction of skinned smooth muscle occurred in the absence of Ca(2+) and correlated with phosphorylation of myosin light chain at Ser(19) and Thr(18) by a kinase distinct from myosin light chain kinase. In this study, we identify this kinase as integrin-linked kinase. Chicken gizzard integrin-linked kinase cDNA was cloned, sequenced, expressed in E. coli, and shown to phosphorylate myosin light chain in the absence of Ca(2+) at Ser(19) and Thr(18). Subcellular fractionation revealed two distinct populations of integrin-linked kinase, including a Triton X-100-insoluble component that phosphorylates myosin in a Ca(2+)-independent manner. These results suggest a novel function for integrin-linked kinase in the regulation of smooth muscle contraction via Ca(2+)-independent phosphorylation of myosin, raise the possibility that integrin-linked kinase may also play a role in regulation of nonmuscle motility, and confirm that integrin-linked kinase is indeed a functional protein-serine/threonine kinase. PMID- 11278952 TI - Cyclic green fluorescent protein produced in vivo using an artificially split PI PfuI intein from Pyrococcus furiosus. AB - A cyclic protein was produced in vivo using the intein from Pyrococcus furiosus PI-PfuI in a novel approach to create a circular permutation of the precursor protein by introducing new termini in the intein domain. Green fluorescent protein (GFP) was cyclized with this method in vivo on milligram scales. There was no by-product of linear or polymerized species isolated, unlike with other in vitro or in vivo cyclization methods utilizing inteins. Cyclized GFP unfolded at half the rate of the linear form upon chemical denaturation and required >2 days in 7 m guanidine hydrochloride until a residual fast folding phase (consistent with a persistent cis-proline) had disappeared. Cyclic GFP might become a novel tool for studying the role of termini and backbone topology in various biological processes such as protein degradation and translocation in vivo as well as in vitro. PMID- 11278953 TI - The tumor suppressor protein p53 requires a cofactor to activate transcriptionally the human BAX promoter. AB - An important regulator of the proapoptotic BAX is the tumor suppressor protein p53. Unlike the p21 gene, in which p53-dependent transcriptional activation is mediated by a response element containing two consensus p53 half-sites, it previously was reported that activation of the BAX element by p53 requires additional sequences. Here, it is demonstrated that the minimal BAX response element capable of mediating p53-dependent transcriptional activation consists of two p53 half-sites plus an adjacent 6 base pairs (5'-GGGCGT-3'). This GC-rich region constitutes a "GC box" capable both of binding members of the Sp family of transcription factors, including Sp1 in vitro, and of conferring Sp1-dependent transcriptional activation on a minimal promoter in cells. Mutations within this GC box abrogated the ability of p53 to activate transcription without affecting the affinity of p53 for its binding site, demonstrating that these 6 bases are required for p53-dependent activation. In addition, a positive correlation was observed between the ability of p53 to activate transcription in cells and the ability of Sp1 to bind this response element in vitro. Mutations that inhibited Sp1 binding also blocked the ability of p53 to activate transcription through this element. Together, these results suggest a model in which p53 requires the cooperation of Sp1 or a Sp1-like factor to mediate transcriptional activation of the human BAX promoter. PMID- 11278954 TI - Triplex-induced recombination in human cell-free extracts. Dependence on XPA and HsRad51. AB - Triple helix-forming oligonucleotides (TFOs) can bind to polypurine/polypyrimidine regions in DNA in a sequence-specific manner. Triple helix formation has been shown to stimulate recombination in mammalian cells in both episomal and chromosomal targets containing direct repeat sequences. Bifunctional oligonucleotides consisting of a recombination donor domain tethered to a TFO domain were found to mediate site-specific recombination in an intracellular SV40 vector target. To elucidate the mechanism of triplex-induced recombination, we have examined the ability of intermolecular triplexes to provoke recombination within plasmid substrates in human cell-free extracts. An assay for reversion of a point mutation in the supFG1 gene in the plasmid pSupFG1/G144C was established in which recombination in the extracts was detected upon transformation into indicator bacteria. A bifunctional oligonucleotide containing a 30-nucleotide TFO domain linked to a 40-nucleotide donor domain was found to mediate gene correction in vitro at a frequency of 46 x 10(-)5, at least 20-fold above background and over 4-fold greater than the donor segment alone. Physical linkage of the TFO to the donor was unnecessary, as co-mixture of separate TFO and donor segments also yielded elevated gene correction frequencies. When the recombination and repair proteins HsRad51 and XPA were depleted from the extracts using specific antibodies, the triplex-induced recombination was diminished, but was either partially or completely restored upon supplementation with the purified HsRad51 or XPA proteins, respectively. These results establish that triplex-induced, intermolecular recombination between plasmid targets and short fragments of homologous DNA can be detected in human cell extracts and that this process is dependent on both XPA and HsRad51. PMID- 11278955 TI - Molecular cloning of MIS, a myeloid inhibitory siglec, that binds protein tyrosine phosphatases SHP-1 and SHP-2. AB - We describe the molecular cloning and characterization of a novel myeloid inhibitory siglec, MIS, that belongs to the family of sialic acid-binding immunoglobulin-like lectins. A full-length MIS cDNA was obtained from murine bone marrow cells. MIS is predicted to contain an extracellular region comprising three immunoglobulin-like domains (V-set amino-terminal domain followed by two C set domains), a transmembrane domain and a cytoplasmic tail with two immunoreceptor tyrosine-based inhibitory motif (ITIM)-like sequences. The closest relative of MIS in the siglec family is human siglec 8. Extracellular regions of these two siglecs share 47% identity at the amino acid level. Southern blot analysis suggests the presence of one MIS gene. MIS is expressed in the spleen, liver, heart, kidney, lung and testis tissues. Several isoforms of MIS protein exist due to the alternative splicing. In a human promonocyte cell line, MIS was able to bind Src homology 2-containing protein-tyrosine phosphatases, SHP-1 and SHP-2. This binding was mediated by the membrane-proximal ITIM of MIS. Moreover, MIS exerted an inhibitory effect on FcgammaRI receptor-induced calcium mobilization. These data suggest that MIS can play an inhibitory role through its ITIM sequences. PMID- 11278956 TI - Peroxisome proliferator-activated receptor-gamma activation inhibits interleukin 1beta -mediated platelet-derived growth factor-alpha receptor gene expression via CCAAT/enhancer-binding protein-delta in vascular smooth muscle cells. AB - CCAAT/enhancer-binding protein (C/EBP)-binding motifs have been identified in the promoter regions of interleukin (IL)-6, tumor necrosis factor-alpha, and platelet derived growth factor-alpha receptor (PDGFalphaR). Recently, peroxisome proliferator-activated receptors (PPARs) have been suggested to be important immunomodulatory mediators. Although many studies have demonstrated that the interaction between C/EBPs and PPARs plays a central role in lipid metabolism, expression and function of these factors are unknown in vascular smooth muscle cells (VSMCs). In the present study, we clarified a functional relationship between C/EBPs and PPARgamma in the regulation of IL-1beta-induced PDGFalphaR expression in VSMCs. PPARgamma activators, troglitazone and 15-deoxy-Delta(12,14) prostaglandin J(2), inhibited IL-1beta-induced PDGFalphaR expression and suppressed PDGF-induced proliferation activity of VSMCs. Electromobility shift and supershift assays for a C/EBP motif in the PDGFalphaR promoter region revealed that PPARgamma activators suppressed IL-1beta-induced DNA binding activity of C/EBPdelta and beta. PPARgamma activators also suppressed IL-1beta induced C/EBPdelta expression. In contrast, overexpression of C/EBPdelta reversed the suppressive effect of PPARgamma activators on PDGFalphaR expression almost completely. From these results, we conclude that the inhibitory effect of PPARgamma activators on PDGFalphaR expression is mainly mediated by C/EBPdelta suppression. Regulation of C/EBPdelta by PPARgamma activators probably plays critical roles in modulating inflammatory responses in the arterial wall. PMID- 11278957 TI - Antioxidant system within yeast peroxisome. Biochemical and physiological characterization of CbPmp20 in the methylotrophic yeast Candida boidinii. AB - Candida boidinii Pmp20 (CbPmp20), a protein associated with the inner side of peroxisomal membrane, belongs to a recently identified protein family of antioxidant enzymes, the peroxiredoxins, which contain one cysteine residue. Pmp20 homologs containing the putative peroxisome targeting signal type 1 have also been identified in mammals and lower eukaryotes. However, the physiological function of these Pmp20 family proteins has been unclear. In this study, we investigated the biochemical and physiological functions of recombinant CbPmp20 protein in methanol-induced peroxisomes of C. boidinii using the PMP20-deleted strain of C. boidinii (pmp20Delta strain). The His(6)-tagged CbPmp20 fusion protein was found to have glutathione peroxidase activity in vitro toward alkyl hydroperoxides and H(2)O(2). Catalytic activity and dimerization of His(6) CbPmp20 depended on the only cysteine residue corresponding to Cys(53). The pmp20Delta strain was found to have lost growth ability on methanol as a carbon and energy source. The pmp20Delta growth defect was rescued by CbPmp20, but neither CbPmp20 lacking the peroxisome targeting signal type 1 sequence nor CbPmp20 haboring the C53S mutation retrieved the growth defect. Interestingly, the pmp20Delta strain had a more severe growth defect than the cta1Delta strain, which lacks catalase, another antioxidant enzyme within the peroxisome. During incubation of these strains in methanol medium, the cta1Delta strain accumulated H(2)O(2), whereas the pmp20Delta strain did not. Therefore, it is speculated to be the main function of CbPmp20 is to decompose reactive oxygen species generated at peroxisomal membrane surface, e.g. lipid hydroperoxides, rather than to decompose H(2)O(2). In addition, we detected a physiological level of reduced glutathione in peroxisomal fraction of C. boidinii. These results may indicate a physiological role for CbPmp20 as an antioxidant enzyme within peroxisomes rich in reactive oxygen species. PMID- 11278958 TI - Mitogen-activated protein kinase phosphatase 1 activity is necessary for oxidized phospholipids to induce monocyte chemotactic activity in human aortic endothelial cells. AB - Entrapment and oxidation of low density lipoproteins (LDL) in the sub-endothelial space is a key process in the initiation of atherosclerotic lesion development. Functional changes induced by oxidized lipids in endothelial cells are early events in the pathogenesis of atherosclerosis. Oxidized-l-alpha-1-palmitoyl-2 arachidonoyl-sn-glycero-3-phosphocholine (ox-PAPC), a major component of minimally modified/oxidized-LDL (MM-LDL) mimics the biological activities assigned to MM-LDL both in vitro in a co-culture model as well as in vivo in mice. We hypothesized that ox-PAPC initiates gene expression changes in endothelial cells that result in enhanced endothelial/monocyte interactions. To analyze the gene expression changes that oxidized lipids induce in endothelial cells, we used a suppression subtractive hybridization procedure to compare mRNA from PAPC-treated human aortic endothelial cells (HAEC) with that of ox-PAPC treated cells. We report here the identification of a gene, mitogen-activated protein kinase phosphatase 1 (MKP-1), that is rapidly and transiently induced in ox-PAPC-treated HAEC. Inhibition of MKP-1 using either the phosphatase inhibitor sodium orthovanadate or antisense oligonucleotides prevents the accumulation of monocyte chemotactic activity in ox-PAPC-treated HAEC supernatants. Furthermore, we show that decreased monocyte chemotactic activity in HAEC treated with sodium orthovanadate or MKP-1 antisense oligonucleotides is due to decreased MCP-1 protein. Our results implicate a direct role for MKP-1 in ox-PAPC-induced signaling pathways that result in the production of MCP-1 protein by ox-PAPC treated HAEC. PMID- 11278959 TI - Involvement of fibronectin type II repeats in the efficient inhibition of gelatinases A and B by long-chain unsaturated fatty acids. AB - The matrix metalloproteinases gelatinase A (MMP-2) and gelatinase B (MMP-9) are implicated in the physiological and pathological breakdown of several extracellular matrix proteins. In the present study, we show that long-chain fatty acids (e.g. oleic acid, elaidic acid, and cis- and trans-parinaric acids) inhibit gelatinase A as well as gelatinase B with K(i) values in the micromolar range but had only weak inhibitory effect on collagenase-1 (MMP-1), as assessed using synthetic or natural substrates. The inhibition of gelatinases depended on fatty acid chain length (with C18 > C16, C14, and C10), and the presence of unsaturations increased their inhibitory capacity on both types of gelatinase. Ex vivo experiments on human skin tissue sections have shown that micromolar concentrations of a long-chain unsaturated fatty acid (elaidic acid) protect collagen and elastin fibers against degradation by gelatinases A and B, respectively. In order to understand why gelatinases are more susceptible than collagenase-1 to inhibition by long-chain fatty acids, the possible role of the fibronectin-like domain (a domain unique to gelatinases) in binding inhibitory fatty acids was investigated. Affinity and kinetic studies with a recombinant fibronectin-like domain of gelatinase A and with a recombinant mutant of gelatinase A from which this domain had been deleted pointed to an interaction of long-chain fatty acids with the fibronectin-like domain of the protease. Surface plasmon resonance studies on the interaction of long-chain fatty acids with the three individual type II modules of the fibronectin-like domain of gelatinase A revealed that the first type II module is primarily responsible for binding these compounds. PMID- 11278960 TI - Human ClC-3 is not the swelling-activated chloride channel involved in cell volume regulation. AB - Volume regulation is essential for normal cell function. A key component of the cells' response to volume changes is the activation of a channel, which elicits characteristic chloride currents (I(Cl, Swell)). The molecular identity of this channel has been controversial. Most recently, ClC-3, a protein highly homologous to the ClC-4 and ClC-5 channel proteins, has been proposed as being responsible for I(Cl, Swell). Subsequently, however, other reports have suggested that ClC-3 may generate chloride currents with characteristics clearly distinct from I(Cl, Swell). Significantly different tissue distributions for ClC-3 have also been reported, and it has been suggested that two isoforms of ClC-3 may be expressed with differing functions. In this study we generated a series of cell lines expressing variants of ClC-3 to rigorously address the question of whether or not ClC-3 is responsible for I(Cl, Swell). The data demonstrate that ClC-3 is not responsible for I(Cl, Swell) and has no role in regulatory volume decrease, furthermore, ClC-3 is not activated by intracellular calcium and fails to elicit chloride currents under any conditions tested. Expression of ClC-3 was shown to be relatively tissue-specific, with high levels in the central nervous system and kidney, and in contrast to previous reports, is essentially absent from heart. This distribution is also inconsistent with the previous proposed role in cell volume regulation. PMID- 11278961 TI - Tumor targeting of mono-, di-, and tetravalent anti-p185(HER-2) miniantibodies multimerized by self-associating peptides. AB - Multimerization of antibody fragments increases the valency and the molecular weight, both identified as key features in the design of the optimal targeting molecule. Here, we report the construction of mono-, di-, and tetrameric variants of the anti-tumor p185(HER-2) single chain Fv fragment 4D5 by fusion of self associating peptides to the carboxyl terminus. Dimeric miniantibodies with a synthetic helix-turn-helix domain and tetrameric ones with the multimerization domain of the human p53 protein were produced in functional form in the periplasm of Escherichia coli. We have directly compared these molecules and the single chain Fv fragment in the targeting of SK-OV-3 xenografts. Tetramerization of the 4D5 antibody fragment resulted in increased serum persistence, significantly reduced off-rate, due to the avidity effect, both in surface plasmon resonance measurements on purified p185(HER-2) and on SK-OV-3 cells. The (99m)technetium tricarbonyl-labeled tetrameric 4D5-p53 miniantibody localized with the highest dose at the tumor and remained stably bound for at least 72 h. The highest total dose was 4.3% injected dose/g after 24 h, whereas the highest tumor-to-blood ratio was found to be 13.5:1 after 48 h, with a total dose of 3.2% injected dose/g. The tetramer shows no higher avidity than the dimer, presumably since the simultaneous binding to more than two antigen molecules on the surface of cells is not possible, and the improvement in performance over the dimer must at least be due in part to the molecular weight. These results demonstrate that multimerization by self-associating peptides can be used for the development of more effective targeting molecules for medical diagnostics and therapy. PMID- 11278962 TI - The TXP motif in the second transmembrane helix of CCR5. A structural determinant of chemokine-induced activation. AB - CCR5 is a G-protein-coupled receptor activated by the chemokines RANTES (regulated on activation normal T cell expressed and secreted), macrophage inflammatory protein 1alpha and 1beta, and monocyte chemotactic protein 2 and is the main co-receptor for the macrophage-tropic human immunodeficiency virus strains. We have identified a sequence motif (TXP) in the second transmembrane helix of chemokine receptors and investigated its role by theoretical and experimental approaches. Molecular dynamics simulations of model alpha-helices in a nonpolar environment were used to show that a TXP motif strongly bends these helices, due to the coordinated action of the proline, which kinks the helix, and of the threonine, which further accentuates this structural deformation. Site directed mutagenesis of the corresponding Pro and Thr residues in CCR5 allowed us to probe the consequences of these structural findings in the context of the whole receptor. The P84A mutation leads to a decreased binding affinity for chemokines and nearly abolishes the functional response of the receptor. In contrast, mutation of Thr-82(2.56) into Val, Ala, Cys, or Ser does not affect chemokine binding. However, the functional response was found to depend strongly on the nature of the substituted side chain. The rank order of impairment of receptor activation is P84A > T82V > T82A > T82C > T82S. This ranking of impairment parallels the bending of the alpha-helix observed in the molecular simulation study. PMID- 11278963 TI - Factor IXa:factor VIIIa interaction. helix 330-338 of factor ixa interacts with residues 558-565 and spatially adjacent regions of the a2 subunit of factor VIIIa. AB - The physiologic activator of factor X consists of a complex of factor IXa, factor VIIIa, Ca(2+) and a suitable phospholipid surface. In one study, helix 330 (162 in chymotrypsin) of the protease domain of factor IXa was implicated in binding to factor VIIIa. In another study, residues 558-565 of the A2 subunit of factor VIIIa were implicated in binding to factor IXa. We now provide data, which indicate that the helix 330 of factor IXa interacts with the 558-565 region of the A2 subunit. Thus, the ability of the isolated A2 subunit was severely impaired in potentiating factor X activation by IXa(R333Q) and by a helix replacement mutant (IXa(helixVII) in which helix 330-338 is replaced by that of factor VII) but it was normal for an epidermal growth factor 1 replacement mutant (IXa(PCEGF1) in which epidermal growth factor 1 domain is replaced by that of protein C). Further, affinity of each 5-dimethylaminonaphthalene-1-sulfonyl (dansyl)-Glu-Gly-Arg-IXa (dEGR-IXa) with the A2 subunit was determined from its ability to inhibit wild-type IXa in the tenase assay and from the changes in dansyl fluorescence emission signal upon its binding to the A2 subunit. Apparent K(d(A2)) values are: dEGR-IXa(WT) or dEGR-IXa(PCEGF1) approximately 100 nm, dEGR IXa(R333Q) approximately 1.8 micrometer, and dEGR-IXa(helixVII) >10 micrometer. In additional experiments, we measured the affinities of these factor IXa molecules for a peptide comprising residues 558-565 of the A2 subunit. Apparent K(d(peptide)) values are: dEGR-IXa(WT) or dEGR-IXa(PCEGF1) approximately 4 micrometer, and dEGR-IXa(R333Q) approximately 62 micrometer. Thus as compared with the wild-type or PCEGF1 mutant, the affinity of the R333Q mutant for the A2 subunit or the A2 558-565 peptide is similarly reduced. These data support a conclusion that the helix 330 of factor IXa interacts with the A2 558-565 sequence. This information was used to model the interface between the IXa protease domain and the A2 subunit, which is also provided herein. PMID- 11278964 TI - Ataxia telangiectasia mutated (ATM) kinase and ATM and Rad3 related kinase mediate phosphorylation of Brca1 at distinct and overlapping sites. In vivo assessment using phospho-specific antibodies. AB - Recent studies have provided evidence that breast cancer susceptibility gene products (Brca1 and Brca2) suppress cancer, at least in part, by participating in DNA damage signaling and DNA repair. Brca1 is hyperphosphorylated in response to DNA damage and co-localizes with Rad51, a protein involved in homologous recombination, and Nbs1.Mre11.Rad50, a complex required for both homologous recombination and nonhomologous end joining repair of damaged DNA. Here, we report that there is a qualitative difference in the phosphorylation states of Brca1 between ionizing radiation (IR) and UV radiation. Brca1 is phosphorylated at Ser-1423 and Ser-1524 after IR and UV; however, Ser-1387 is specifically phosphorylated after IR, and Ser-1457 is predominantly phosphorylated after UV. These results suggest that different types of DNA-damaging agents might signal to Brca1 in different ways. We also provide evidence that the rapid phosphorylation of Brca1 at Ser-1423 and Ser-1524 after IR (but not after UV) is largely ataxia telangiectasia mutated (ATM) kinase-dependent. The overexpression of catalytically inactive ATM and Rad3 related (ATR) kinase inhibited the UV-induced phosphorylation of Brca1 at these sites, indicating that ATR controls Brca1 phosphorylation in vivo after the exposure of cells to UV light. Moreover, ATR associates with Brca1; ATR and Brca1 foci co-localize both in cells synchronized in S phase and after exposure of cells to DNA-damaging agents. ATR can itself phosphorylate the region of Brca1 phosphorylated by ATM (Ser-Gln cluster in the C terminus of Brca1, amino acids 1241-1530). However, there are additional uncharacterized ATR phosphorylation site(s) between residues 521 and 757 of Brca1. Taken together, our results support a model in which ATM and ATR act in parallel but somewhat overlapping pathways of DNA damage signaling but respond primarily to different types of DNA lesion. PMID- 11278965 TI - NFAT4 movement in native smooth muscle. A role for differential Ca(2+) signaling. AB - The transcription factor NFAT (nuclear factor of activated T-cells) plays a central role in mediating Ca(2+)-dependent gene transcription in a variety of cell types. Sustained increases in intracellular calcium concentration ([Ca(2+)]i) are presumed to be required for NFAT dephosphorylation by the Ca(2+)/calmodulin-dependent protein calcineurin and its subsequent nuclear translocation. Here, we provide the first identification and characterization of NFAT in native smooth muscle, showing that NFAT4 is the predominant isoform detected by reverse transcriptase-polymerase chain reaction and Western blot analysis. PDGF induces NFAT4 translocation in smooth muscle, leading to an increase in NFAT transcriptional activity. NFAT4 activation by PDGF depends on Ca(2+) entry through voltage-dependent Ca(2+) channels, because its nuclear accumulation is prevented by the Ca(2+) channel blocker nisoldipine and the K(+) channel opener pinacidil. Interestingly, elevation of [Ca(2+)]i by membrane depolarization or ionomycin treatment are not effective stimuli for NFAT4 nuclear accumulation, indicating that Ca(2+) influx is necessary but not sufficient for NFAT4 activation. In contrast, membrane depolarization readily activates the Ca(2+)-dependent transcription factor CREB (cAMP-responsive element-binding protein). The calcineurin blockers CsA and FK506 also prevented the PDGF-induced NFAT4 nuclear localization. These results indicate that both the nature of the calcium signal and PDGF-induced modulation of nuclear import-export of NFAT are critical for NFAT4 activation in this tissue. PMID- 11278966 TI - The transmembrane domain of syntaxin 1A is critical for cytoplasmic domain protein-protein interactions. AB - Assembly of the plasma membrane proteins syntaxin 1A and SNAP-25 with the vesicle protein synaptobrevin is a critical step in neuronal exocytosis. Syntaxin is anchored to the inner face of presynaptic plasma membrane via a single C-terminal membrane-spanning domain. Here we report that this transmembrane domain plays a critical role in a wide range of syntaxin protein-protein interactions. Truncations or deletions of the membrane-spanning domain reduce synaptotagmin, alpha/beta-SNAP, and synaptobrevin binding. In contrast, deletion of the transmembrane domain potentiates SNAP-25 and rbSec1A/nsec-1/munc18 binding. Normal partner protein binding activity of the isolated cytoplasmic domain could be "rescued" by fusion to the transmembrane segments of synaptobrevin and to a lesser extent, synaptotagmin. However, efficient rescue was not achieved by replacing deleted transmembrane segments with corresponding lengths of other hydrophobic amino acids. Mutations reported to diminish the dimerization of the transmembrane domain of syntaxin did not impair the interaction of full-length syntaxin with other proteins. Finally, we observed that membrane insertion and wild-type interactions with interacting proteins are not correlated. We conclude that the transmembrane domain, via a length-dependent and sequence-specific mechanism, affects the ability of the cytoplasmic domain to engage other proteins. PMID- 11278968 TI - Revisiting the lysogenization control of bacteriophage lambda. Identification and characterization of a new host component, HflD. AB - Upon infection to the Escherichia coli cell, the genome of bacteriophage lambda either replicates to form new progenies (lytic growth) or integrates into the host chromosome (lysogenization). The lambda CII protein is a key determinant in the lysis-lysogeny decision. It is a short-lived transcription activator for the lambda genes essential for lysogeny establishment. In this study, we isolated a new class of hfl (high frequency lysogenization) mutants of E. coli, using a new selection for enhancement of CII-stimulated transcription. The gene affected was termed hflD, which encodes a protein of 213 amino acids. An hflD-disrupted mutant indeed showed an Hfl phenotype, indicating that HflD acts to down-regulate lysogenization. HflD is associated peripherally with the cytoplasmic membrane. Its interaction with CII was demonstrated in vitro using purified proteins as well as in vivo using the bacterial two-hybrid system. Pulse-chase examinations demonstrated that the HflD function is required for the rapid in vivo degradation of CII, although it interfered with FtsH-mediated CII proteolysis in an in vitro reaction system using detergent-solubilized components. We suggest that HflD is a factor that sequesters CII from the target promoters and recruits it to the membrane where the FtsH protease is localized. PMID- 11278969 TI - Reaction of human myoglobin and H2O2. Electron transfer between tyrosine 103 phenoxyl radical and cysteine 110 yields a protein-thiyl radical. AB - The sequence of human myoglobin (Mb) is similar to that of other species except for a unique cysteine at position 110 (Cys(110)). Adding hydrogen peroxide (H(2)O(2)) to human Mb affords Trp(14)-peroxyl, Tyr(103)-phenoxyl, and Cys(110) thiyl radicals and coupling of Cys(110)-thiyl radicals yields a homodimer through intermolecular disulfide bond formation (Witting, P. K., Douglas, D. J., and Mauk, A. G. (2000) J. Biol. Chem. 275, 20391-20398). Treating a solution of wild type Mb and H(2)O(2) with 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) at DMPO:protein /= 100 mol/mol only DMPO-Tyr(103) radicals were present. The DMPO-dependent decrease in DMPO-Cys(110) was matched by a near 1:1 stoichiometric increase in DMPO-Tyr(103). In contrast, reaction of the Y103F human Mb with H(2)O(2) gave no DMPO-Cys(110) at DMPO:protein /= 100 mol/mol (i.e. conditions that consistently gave DMPO-Tyr(103) in the case of wild type Mb). No detectable homodimer was formed by incubation of the Y103F variant with H(2)O(2). However, the homodimer was detected in a mixture of both the Y103F and C110A variants of human Mb upon treatment with H(2)O(2) (C110A:Y103F:H(2)O(2) 2:1:5 mol/mol/mol); the yield of this homodimer increased with increasing ratios of C110A:Y103F. Together, these data suggest that addition of H(2)O(2) to human Mb can produce Cys(110)-thiyl radicals through an intermolecular electron transfer reaction from Cys(110) to a Tyr(103)-phenoxyl radical. PMID- 11278967 TI - Saturated fatty acids, but not unsaturated fatty acids, induce the expression of cyclooxygenase-2 mediated through Toll-like receptor 4. AB - Results from our previous studies demonstrated that activation of Toll-like receptor 4 (Tlr4), the lipopolysaccharide (LPS) receptor, is sufficient to induce nuclear factor kappaB activation and expression of inducible cyclooxygenase (COX 2) in macrophages. Saturated fatty acids (SFAs) acylated in lipid A moiety of LPS are essential for biological activities of LPS. Thus, we determined whether these fatty acids modulate LPS-induced signaling pathways and COX-2 expression in monocyte/macrophage cells (RAW 264.7). Results show that SFAs, but not unsaturated fatty acids (UFAs), induce nuclear factor kappaB activation and expression of COX-2 and other inflammatory markers. This induction is inhibited by a dominant-negative Tlr4. UFAs inhibit COX-2 expression induced by SFAs, constitutively active Tlr4, or LPS. However, UFAs fail to inhibit COX-2 expression induced by activation of signaling components downstream of Tlr4. Together, these results suggest that both SFA-induced COX-2 expression and its inhibition by UFAs are mediated through a common signaling pathway derived from Tlr4. These results represent a novel mechanism by which fatty acids modulate signaling pathways and target gene expression. Furthermore, these results suggest a possibility that propensity of monocyte/macrophage activation is modulated through Tlr4 by different types of free fatty acids, which in turn can be altered by kinds of dietary fat consumed. PMID- 11278970 TI - Uncoupling protein 3 (UCP3) stimulates glucose uptake in muscle cells through a phosphoinositide 3-kinase-dependent mechanism. AB - UCP3 is a mitochondrial membrane protein expressed in humans selectively in skeletal muscle. To determine the mechanisms by which UCP3 plays a role in regulating glucose metabolism, we expressed human UCP3 in L6 myotubes by adenovirus-mediated gene transfer and in H(9)C(2) cardiomyoblasts by stable transfection with a tetracycline-repressible UCP3 construct. Expression of UCP3 in L6 myotubes increased 2-deoxyglucose uptake 2-fold and cell surface GLUT4 2.3 fold, thereby reaching maximally insulin-stimulated levels in control myotubes. Wortmannin, LY 294002, or the tyrosine kinase inhibitor genistein abolished the effect of UCP3 on glucose uptake, and wortmannin inhibited UCP3-induced GLUT4 cell surface recruitment. UCP3 overexpression increased phosphotyrosine associated phosphoinositide 3-kinase (PI3K) activity 2.2-fold compared with control cells (p < 0.05). UCP3 overexpression increased lactate release 1.5- to 2 fold above control cells, indicating increased glucose metabolism. In H(9)C(2) cardiomyoblasts stably transfected with UCP3 under control of a tetracycline repressible promotor, removal of doxycycline resulted in detectable levels of UCP3 at 12 h and 2.2-fold induction at 7 days compared with 12 h. In parallel, glucose transport increased 1.3- and 2-fold at 12 h and 7 days, respectively, and the stimulation was inhibited by wortmannin or genistein. p85 association with membranes was increased 5.5-fold and phosphotyrosine-associated PI3K activity 3.8 fold. In contrast, overexpression of UCP3 in 3T3-L1 adipocytes did not alter glucose uptake, suggesting tissue-specific effects of human UCP3. Thus, UCP3 stimulates glucose transport and GLUT4 translocation to the cell surface in cardiac and skeletal muscle cells by activating a PI3K dependent pathway. PMID- 11278971 TI - A homologue of N-ethylmaleimide-sensitive factor in the malaria parasite Plasmodium falciparum is exported and localized in vesicular structures in the cytoplasm of infected erythrocytes in the brefeldin A-sensitive pathway. AB - N-Ethylmaleimide-sensitive factor (NSF) and its homologues play a central role in vesicular trafficking in eukaryotic cells. We have identified a NSF homologue in Plasmodium falciparum (PfNSF). The reported PfNSF gene sequence (GenBank accession number CAB10575) indicated that PfNSF comprises 783 amino acids with a calculated molecular weight of 89,133. The overall identities of its gene and amino acid sequences with those of rat NSF are 50.9 and 48.8%, respectively. Reverse transcription-polymerase chain reaction analysis and Northern blotting with total P. falciparum RNA indicated expression of the PfNSF gene. Polyclonal antibodies against a conserved region of NSF specifically recognized an 89-kDa polypeptide in the parasite cells. After homogenization of the parasite cells, approximately 90% of an 89-kDa polypeptide is associated with particulate fraction, suggesting membrane-bound nature of PfNSF. PfNSF was present within both the parasite cells and the vesicular structure outside of the parasite cells. The export of PfNSF outside of the parasite cells appears to occur at the early trophozoite stage and to terminate at the merozoite stage. The export of PfNSF is inhibited by brefeldin A, with 9 microM causing 50% inhibition. Immunoelectromicroscopy indicated that intracellular PfNSF was associated with organelles such as food vacuoles and that extracellular PfNSF was associated with vesicular structures in the erythrocyte cytoplasm. These results indicate that PfNSF expressed in the malaria parasite is exported to the extracellular space and then localized in intraerythrocytic vesicles in a brefeldin A-sensitive manner. It is suggested that a vesicular transport mechanism is involved in protein export targeted to erythrocyte membranes during intraerythrocytic development of the malaria parasite. PMID- 11278972 TI - New insights into host factor requirements for prokaryotic beta-recombinase mediated reactions in mammalian cells. AB - The prokaryotic beta-recombinase catalyzes site-specific recombination between two directly oriented minimal six sites in mammalian cells, both on episomic and chromatin-integrated substrates. Using a specific recombination activated gene expression system, we report the site-specific recombination activity of an enhanced green fluorescent protein (EGFP) fused version of beta-recombinase (beta EGFP). This allows expression of active beta-recombinase detectable in vivo and in fixed cells by fluorescence microscopy. In addition, cellular viability is compatible with a substantial level of expression of the beta-EGFP protein. Using fluorescence-activated cell sorting, we have been able to enrich cell populations expressing this fusion protein. Application of this strategy has allowed us to study in more depth the host factor requirements for this system. Previous work showed that eukaryotic HMG1 protein was necessary and sufficient to help beta recombinase activity in vitro. The influence of ectopic expression of HMG1 protein in the recombination process has been analyzed, indicating that HMG1 overexpression does not lead to a significant increase on the efficiency of beta recombinase-mediated recombination both on episomal substrates and chromatin associated targets. In addition, beta-recombinase-mediated recombination has been demonstrated in HMG1 deficient cells at the same levels as in wild type cells. These data demonstrate the existence of cellular factors different from HMG-1 that can act as helpers for beta-recombinase activity in the eukaryotic environment. PMID- 11278973 TI - Expression analysis of the nrdHIEF operon from Escherichia coli. Conditions that trigger the transcript level in vivo. AB - Escherichia coli has two aerobic ribonucleotide reductases encoded by the nrdAB and nrdHIEF operons. While NrdAB is active during aerobiosis, NrdEF is considered a cryptic enzyme with no obvious function. Here, we present evidence that nrdHIEF expression might be important under certain circumstances. Basal transcript levels were dramatically enhanced (25-75-fold), depending on the growth-phase and the growth-medium composition. Likewise, a large increase of >100-fold in nrdHIEF mRNA was observed in bacteria lacking Trx1 and Grx1, the two main NrdAB reductants. Moreover, nrdHIEF expression was triggered in response to oxidative stress, particularly in mutants missing hydroperoxidase I and alkyl-hydroperoxide reductase activities (69.7-fold) and in cells treated with oxidants (up to 23.4 fold over the enhanced transcript level possessed by cells grown on minimal medium). The mechanism(s) that triggers nrdHIEF expression remains unknown, but our findings exclude putative global regulators like RpoS, Fis, cAMP, OxyR, SoxR/S, or RecA. What we have learned about nrdHIEF expression indicates strong differences between its regulation and that of the nrdAB operon and of genes coding for components of both thioredoxin/glutaredoxin pathways. We propose that E. coli might optimize the responses to different stimuli by co-evolving the expression levels for its multiple reductases and electron donors. PMID- 11278974 TI - DNA synthesis and mitotic clonal expansion is not a required step for 3T3-L1 preadipocyte differentiation into adipocytes. AB - Upon differentiation induction of 3T3-L1 preadipocytes by a hormone mixture containing 1-isobutyl-3-methylxanthine, dexamethasone, and insulin, the preadipocytes undergo approximately 2 rounds of mitotic clonal expansion, which just precedes the adipogenic gene expression program and has been thought to be an essential early step for differentiation initiation. By inducing 3T3-L1 preadipocytes with each individual hormone, it was determined that the mitotic clonal expansion was induced only by insulin and not by 1-isobutyl-3 methylxanthine or dexamethasone. Cell number counting and fluorescence-activated cell-sorting analysis indicated that a significant fraction of 3T3-L1 preadipocytes differentiated into adipocytes without mitotic clonal expansion when induced with the combination of 1-isobutyl-3-methylxanthine and dexamethasone. Furthermore, when normally induced 3T3-L1 preadipocytes were treated with PD98059 (an inhibitor of mitogen-activated protein kinase/extracellular signal-regulated kinase kinase 1) to block the activation of extracellular signal-regulated kinase (Erk) 1 and Erk2, the mitotic clonal expansion was blocked, but adipocyte differentiation was not affected. These observations were confirmed by bromodeoxyuridine labeling. The differentiated adipocytes induced with 1-isobutyl-3-methylxanthine and dexamethasone or standard hormone mixture plus PD98059 were not labeled by bromodeoxyuridine. Thus, it is evident that 3T3-L1 preadipocytes could differentiate into adipocytes without DNA synthesis and mitotic clonal expansion. Our results also suggested that activation of Erk1 and Erk2 is essential to but not sufficient for induction of mitotic clonal expansion. PMID- 11278976 TI - Tat1, a novel sulfate transporter specifically expressed in human male germ cells and potentially linked to rhogtpase signaling. AB - RhoGTPases (Rho, Rac, and Cdc42) are known to regulate multiple functions, including cell motility, adhesion, and proliferation; however, the signaling pathways underlying these pleiotropic effects are far from fully understood. We have recently described a new RhoGAP (GTPase activating protein for RhoGTPases) gene, MgcRacGAP, primarily expressed in male germ cells, at the spermatocyte stage. We report here the isolation, through two-hybrid cloning, of a new partner of MgcRacGAP, very specifically expressed in the male germ line and showing structural features of anion transporters. This large protein (970 amino acids and a predicted size of 109 kDa), we provisionally designated Tat1 (for testis anion transporter 1), is closely related to a sulfate permease family comprising three proteins in human (DRA, Pendrin, and DTD); it is predicted to be an integral membrane protein with 14 transmembrane helices and intracytoplasmic NH(2) and COOH termini. In situ hybridization studies demonstrate that Tat1 and MgcRacGAP genes are coexpressed in male germ cells at the spermatocyte stage. On testis sections, Tat1 protein can be immunodetected in spermatocytes and spermatids associated with plasma membrane. Two-hybrid and in vitro binding assays demonstrate that MgcRacGAP stably interacts through its NH(2)-terminal domain with the Tat1 COOH-terminal region. Expression of Tat1 protein in COS7 cells generates a 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene and chloride sensitive sulfate transport. Therefore we conclude that Tat1 is a novel sulfate transporter specifically expressed in spermatocytes and spermatids and interacts with MgcRacGAP in these cells. These observations raise the possibility of a new regulatory pathway linking sulfate transport to Rho signaling in male germ cells. PMID- 11278975 TI - Inhibition of oxidized low-density lipoprotein-induced apoptosis in endothelial cells by nitric oxide. Peroxyl radical scavenging as an antiapoptotic mechanism. AB - Proatherogenic oxidized low-density lipoprotein (oxLDL) induces endothelial apoptosis. We investigated the anti-apoptotic effects of intracellular and extracellular nitric oxide (*NO) donors, iron chelators, cell-permeable superoxide dismutase (SOD), glutathione peroxidase mimetics, and nitrone spin traps. Peroxynitrite (ONOO-)-modified oxLDL induced endothelial apoptosis was measured by DNA fragmentation, TUNEL assay, and caspase-3 activation. Results indicated the following: (i) the lipid fraction of oxLDL was primarily responsible for endothelial apoptosis. (ii) Endothelial apoptosis was potently inhibited by *NO donors and lipophilic phenolic antioxidants. OxLDL severely depleted Bcl-2 levels in endothelial cells and *NO donors restored Bcl-2 protein in oxLDL-treated cells. (iii) The pretreatment of a lipid fraction derived from oxLDL with sodium borohydride or potassium iodide completely abrogated apoptosis in endothelial cells, suggesting that lipid hydroperoxides induce apoptosis. (iv) Metalloporphyrins dramatically inhibited oxLDL-induced apoptosis in endothelial cells. Neither S-nitrosation of caspase-3 nor induction of Hsp70 appeared to play a significant role in the antiapoptotic mechanism of *NO in oxLDL-induced endothelial apoptosis. We propose that cellular lipid peroxyl radicals or lipid hydroperoxides induce an apoptotic signaling cascade in endothelial cells exposed to oxLDL, and that *NO inhibits apoptosis by scavenging cellular lipid peroxyl radicals. PMID- 11278977 TI - COL5A1 exon 14 splice acceptor mutation causes a functional null allele, haploinsufficiency of alpha 1(V) and abnormal heterotypic interstitial fibrils in Ehlers-Danlos syndrome II. AB - We studied four affected individuals from a family of three generations with Ehlers-Danlos Syndrome II. Type V collagen transcripts of affected individuals were screened by reverse transcriptase-polymerase chain reaction. Amplification of the exon 9-28 region of alpha1(V) yielded normal and larger products from the proband. Sequencing of cDNA revealed a 100-base pair insertion from the 3'-end of intron 13 between exons 13 and 14 in one allele. The genomic defect was identified as an A(-2)--> G substitution at the exon 14 splice acceptor site. A cryptic acceptor site -100 nucleotide within intron 13 is used instead of the mutant splice site. The insertion shifts the reading frame +1 and results in a stop codon within exon 17. The mutant transcript was much less abundant than normal allele product in untreated cultured fibroblasts but was approximately equimolar in cycloheximide-treated cells, suggesting that the mutation causes nonsense-mediated decay of mRNA. By RNase protection experiments, the level of mutant transcript was determined to be 8% that of the normal transcript in untreated proband fibroblasts. Relative to type I collagen, proband fibroblasts secreted only 65% of the amount of type V collagen secreted by normal controls. Selective salt precipitation of proband secreted collagen provided supportive evidence that the alpha chain composition of type V collagen remains alpha1(V)(2)alpha2(V) even in the context of alpha1(V) haploinsufficiency. Type V collagen incorporates into type I collagen fibrils in the extracellular matrix and is thought to regulate fibril diameter. Transmission electron micrographs of type I collagen fibrils in a proband dermal biopsy showed greater heterogeneity in fibril diameter than in a matched control. The proband had a greater proportion of both larger and smaller fibrils and occasional fibrils with a cauliflower configuration. Unlike the genotype/phenotype relationship seen for type I collagen defects and osteogenesis imperfecta, the null allele in this family appears to cause clinical features similar to those seen in cases with structural alterations in type V collagen. PMID- 11278978 TI - Human RAD52 exhibits two modes of self-association. AB - The human RAD52 protein plays an important role in the earliest stages of chromosomal double-strand break repair via the homologous recombination pathway. Individual subunits of RAD52 self-associate into rings that can then form higher order complexes. RAD52 binds to double-strand DNA ends, and recent studies suggest that the higher order self-association of the rings promotes DNA end joining. Earlier studies defined the self-association domain of RAD52 to a unique region in the N-terminal half of the protein. Here we show that there are in fact two experimentally separable self-association domains in RAD52. The N-terminal self-association domain mediates the assembly of monomers into rings, and the previously unidentified domain in the C-terminal half of the protein mediates higher order self-association of the rings. PMID- 11278979 TI - Pathways of epoxyeicosatrienoic acid metabolism in endothelial cells. Implications for the vascular effects of soluble epoxide hydrolase inhibition. AB - Epoxyeicosatrienoic acids (EETs) are products of cytochrome P-450 epoxygenase that possess important vasodilating and anti-inflammatory properties. EETs are converted to the corresponding dihydroxyeicosatrienoic acid (DHET) by soluble epoxide hydrolase (sEH) in mammalian tissues, and inhibition of sEH has been proposed as a novel approach for the treatment of hypertension. We observed that sEH is present in porcine coronary endothelial cells (PCEC), and we found that low concentrations of N,N'-dicyclohexylurea (DCU), a selective sEH inhibitor, have profound effects on EET metabolism in PCEC cultures. Treatment with 3 microM DCU reduced cellular conversion of 14,15-EET to 14,15-DHET by 3-fold after 4 h of incubation, with a concomitant increase in the formation of the novel beta oxidation products 10,11-epoxy-16:2 and 8,9-epoxy-14:1. DCU also markedly enhanced the incorporation of 14,15-EET and its metabolites into PCEC lipids. The most abundant product in DCU-treated cells was 16,17-epoxy-22:3, the elongation product of 14,15-EET. Another novel metabolite, 14,15-epoxy-20:2, was present in DCU-treated cells. DCU also caused a 4-fold increase in release of 14,15-EET when the cells were stimulated with a calcium ionophore. Furthermore, DCU decreased the conversion of [3H]11,12-EET to 11,12-DHET, increased 11,12-EET retention in PCEC lipids, and produced an accumulation of the partial beta-oxidation product 7,8-epoxy-16:2 in the medium. These findings suggest that in addition to being metabolized by sEH, EETs are substrates for beta-oxidation and chain elongation in endothelial cells and that there is considerable interaction among the three pathways. The modulation of EET metabolism by DCU provides novel insight into the mechanisms by which pharmacological or molecular inhibition of sEH effectively treats hypertension. PMID- 11278980 TI - Regulatory interaction between the cystic fibrosis transmembrane conductance regulator and HCO3- salvage mechanisms in model systems and the mouse pancreatic duct. AB - The pancreatic duct expresses cystic fibrosis transmembrane conductance regulator (CFTR) and HCO3- secretory and salvage mechanisms in the luminal membrane. Although CFTR plays a prominent role in HCO3- secretion, the role of CFTR in HCO3 salvage is not known. In the present work, we used molecular, biochemical, and functional approaches to study the regulatory interaction between CFTR and the HCO3- salvage mechanism Na+/H+ exchanger isoform 3 (NHE3) in heterologous expression systems and in the native pancreatic duct. We found that CFTR regulates NHE3 activity by both acute and chronic mechanisms. In the pancreatic duct, CFTR increases expression of NHE3 in the luminal membrane. Thus, luminal expression of NHE3 was reduced by 53% in ducts of homozygote DeltaF508 mice. Accordingly, luminal Na+-dependent and HOE694- sensitive recovery from an acid load was reduced by 60% in ducts of DeltaF508 mice. CFTR and NHE3 were co immunoprecipitated from PS120 cells expressing both proteins and the pancreatic duct of wild type mice but not from PS120 cells lacking CFTR or the pancreas of DeltaF508 mice. The interaction between CFTR and NHE3 required the COOH-terminal PDZ binding motif of CFTR, and mutant CFTR proteins lacking the C terminus were not co-immunoprecipitated with NHE3. Furthermore, when expressed in PS120 cells, wild type CFTR, but not CFTR mutants lacking the C-terminal PDZ binding motif, augmented cAMP-dependent inhibition of NHE3 activity by 31%. These findings reveal that CFTR controls overall HCO3- homeostasis by regulating both pancreatic ductal HCO3- secretory and salvage mechanisms. PMID- 11278981 TI - Purification and characterization of a novel phosphorus-oxidizing enzyme from Pseudomonas stutzeri WM88. AB - The ptxD gene from Pseudomonas stutzeri WM88 encoding the novel phosphorus oxidizing enzyme NAD:phosphite oxidoreductase (trivial name phosphite dehydrogenase, PtxD) was cloned into an expression vector and overproduced in Escherichia coli. The heterologously produced enzyme is indistinguishable from the native enzyme based on mass spectrometry, amino-terminal sequencing, and specific activity analyses. Recombinant PtxD was purified to homogeneity via a two-step affinity protocol and characterized. The enzyme stoichiometrically produces NADH and phosphate from NAD and phosphite. The reverse reaction was not observed. Gel filtration analysis of the purified protein is consistent with PtxD acting as a homodimer. PtxD has a high affinity for its substrates with Km values of 53.1 +/- 6.7 microm and 54.6 +/- 6.7 microm, for phosphite and NAD, respectively. Vmax and kcat were determined to be 12.2 +/- 0.3 micromol x min(-1) x mg(-1) and 440 min(-1). NADP can substitute poorly for NAD; however, none of the numerous compounds examined were able to substitute for phosphite. Initial rate studies in the absence or presence of products and in the presence of the dead end inhibitor sulfite are most consistent with a sequential ordered mechanism for the PtxD reaction, with NAD binding first and NADH being released last. Amino acid sequence comparisons place PtxD as a new member of the d-2 hydroxyacid NAD-dependent dehydrogenases, the only one to have an inorganic substrate. To our knowledge, this is the first detailed biochemical study on an enzyme capable of direct oxidation of a reduced phosphorus compound. PMID- 11278982 TI - c-Jun N-terminal kinase activation by hydrogen peroxide in endothelial cells involves SRC-dependent epidermal growth factor receptor transactivation. AB - The phenotypic properties of the endothelium are subject to modulation by oxidative stress, and the c-Jun N-terminal kinase (JNK) pathway is important in mediating cellular responses to stress, although activation of this pathway in endothelial cells has not been fully characterized. Therefore, we exposed endothelial cells to hydrogen peroxide (H(2)O(2)) and observed rapid activation of JNK within 15 min that involved phosphorylation of JNK and c-Jun and induction of AP-1 DNA binding activity. Inhibition of protein kinase C and phosphoinositide 3-kinase did not effect JNK activation. In contrast, the tyrosine kinase inhibitors, genistein, herbimycin A, and 4-amino-5-(4-chlorophenyl)-7-(t butyl)pyrazolo[3,4-d]pyrimidine (PP2) significantly attenuated H(2)O(2)-induced JNK activation as did endothelial cell adenoviral transfection with a dominant negative form of Src, implicating Src as an upstream activator of JNK. Activation of JNK by H(2)O(2) was also inhibited by AG1478 and antisense oligonucleotides directed against the epidermal growth factor receptor (EGFR), implicating the EGFR in this process. Consistent with this observation, H(2)O(2) stimulated EGFR tyrosine phosphorylation and complex formation with Shc-Grb2 that was abolished by PP2, implicating Src in H(2)O(2)-induced EGFR activation. Tyrosine phosphorylation of the EGFR by H(2)O(2) did not involve receptor autophosphorylation at Tyr(1173) as assessed by an autophosphorylation-specific antibody. These data indicate that H(2)O(2)-induced JNK activation in endothelial cells involves the EGFR through an Src-dependent pathway that is distinct from EGFR ligand activation. These data represent one potential pathway for mediating oxidative stress-induced phenotypic changes in the endothelium. PMID- 11278983 TI - Dimethyl sulfoxide affects the selection of splice sites. AB - Depending on the cell lines and cell types, dimethyl sulfoxide (Me2SO) can induce or block cell differentiation and apoptosis. Although Me2SO treatment alters many levels of gene expression, the molecular processes that are directly affected by Me2SO have not been clearly identified. Here, we report that Me2SO affects splice site selection on model pre-mRNAs incubated in a nuclear extract prepared from HeLa cells. A shift toward the proximal pair of splice sites was observed on pre mRNAs carrying competing 5'-splice sites or competing 3'-splice sites. Because the activity of recombinant hnRNP A1 protein was similar when added to extracts containing or lacking Me2SO, the activity of endogenous A1 proteins is probably not affected by Me2SO. Notably, in a manner reminiscent of SR proteins, Me2SO activated splicing in a HeLa S100 extract. Moreover, the activity of recombinant SR proteins in splice site selection in vitro was improved by Me2SO. Polar solvents like DMF and formamide similarly modulated splice site selection in vitro but formamide did not activate a HeLa S100 extract. We propose that Me2SO improves ionic interactions between splicing factors that contain RS-domains. The direct impact of Me2SO on alternative splicing may explain, at least in part, the different and sometimes opposite effects of Me2SO on cell differentiation and apoptosis. PMID- 11278984 TI - Secretion of surfactant protein C, an integral membrane protein, requires the N terminal propeptide. AB - Proteolytic processing of surfactant protein C (SP-C) proprotein in multivesicular bodies of alveolar type II cells results in a 35-residue mature peptide, consisting of a transmembrane domain and a 10-residue extramembrane domain. SP-C mature peptide is stored in lamellar bodies (a lysosomal-like organelle) and secreted with surfactant phospholipids into the alveolar space. This study was designed to identify the peptide domain of SP-C required for sorting and secretion of this integral membrane peptide. Deletion analyses in transiently transfected PC12 cells and isolated mouse type II cells suggested the extramembrane domain of mature SP-C was cytosolic and sufficient for sorting to the regulated secretory pathway. Intratracheal injection of adenovirus encoding SP-C mature peptide resulted in secretion into the alveolar space of wild type mice but not SP-C (-/-) mice. SP-C secretion in null mice was restored by the addition of the N-terminal propeptide. The cytosolic domain, consisting of the N- terminal propeptide and extramembrane domain of mature SP-C peptide, supported secretion of the transmembrane domain of platelet-derived growth factor receptor. Collectively, these studies indicate that the N-terminal propeptide of SP-C is required for intracellular sorting and secretion of SP-C. PMID- 11278986 TI - Allosteric communication of tryptophan synthase. Functional and regulatory properties of the beta S178P mutant. AB - The alpha(2)beta(2) tryptophan synthase complex is a model enzyme for understanding allosteric regulation. We report the functional and regulatory properties of the betaS178P mutant. Ser-178 is located at the end of helix 6 of the beta subunit, belonging to the domain involved in intersubunit signaling. The carbonyl group of betaSer-178 is hydrogen bonded to Gly-181 of loop 6 of the alpha subunit only when alpha subunit ligands are bound. An analysis by molecular modeling of the structural effects caused by the betaS178P mutation suggests that the hydrogen bond involving alphaGly-181 is disrupted as a result of localized structural perturbations. The ratio of alpha to beta subunit concentrations was calculated to be 0.7, as for the wild type, indicating the maintenance of a tight alpha-beta complex. Both the activity of the alpha subunit and the inhibitory effect of the alpha subunit ligands indole-3-acetylglycine and d,l-alpha-glycerol 3-phosphate were found to be the same for the mutant and wild type enzyme, whereas the beta subunit activity of the mutant exhibited a 2-fold decrease. In striking contrast to that observed for the wild type, the allosteric effectors indole-3-acetylglycine and d,l-alpha-glycerol-3-phosphate do not affect the beta activity. Accordingly, the distribution of l-serine intermediates at the beta site, dominated by the alpha-aminoacrylate, is only slightly influenced by alpha subunit ligands. Binding of sodium ions is weaker in the mutant than in the wild type and leads to a limited increase of the amount of the external aldimine intermediate, even at high pH, whereas binding of cesium ions exhibits the same affinity and effects as in the wild type, leading to an increase of the alpha aminoacrylate tautomer absorbing at 450 nm. Crystals of the betaS178P mutant were grown, and their functional and regulatory properties were investigated by polarized absorption microspectrophotometry. These findings indicate that (i) the reciprocal activation of the alpha and beta activity in the alpha2beta2 complex with respect to the isolated subunits results from interactions that involve residues different from betaSer-178 and (ii) betaSer-178 is a critical residue in ligand-triggered signals between alpha and beta active sites. PMID- 11278987 TI - Intrinsic DNA distortion of the bacteriophage Mu momP1 promoter is a negative regulator of its transcription. A novel mode of regulation of toxic gene expression. AB - The momP1 promoter of the bacteriophage Mu mom operon is an example of a weak promoter. It contains a 19-base pair suboptimal spacer between the -35 (ACCACA) and -10 (TAGAAT) hexamers. Escherichia coli RNA polymerase is unable to bind to momP1 on its own. DNA distortion caused by the presence of a run of six T nucleotides overlapping the 5' end of the -10 element might prevent RNA polymerase from binding to momP1. To investigate the influence of the T(6) run on momP1 expression, defined substitution mutations were introduced by site-directed mutagenesis. In vitro probing experiments with copper phenanthroline ((OP)(2)Cu) and DNase I revealed distinct differences in cleavage patterns among the various mutants; in addition, compared with the wild type, the mutants showed an increase (variable) in momP1 promoter activity in vivo. Promoter strength analyses were in agreement with the ability of these mutants to form open complexes as well as to produce momP1-specific transcripts. No significant role is attributed to the overlapping and divergently organized promoter, momP2, in the expression of momP1 activity, as determined by promoter disruption analysis. These data support the view that an intrinsic DNA distortion in the spacer region of momP1 acts in cis as a negative element in mom operon transcription. This is a novel mechanism of regulation of toxic gene expression. PMID- 11278985 TI - Up-regulation of apoptosis inhibitory protein IAP-2 by hypoxia. Hif-1-independent mechanisms. AB - Hypoxia is a key determinant of tissue pathology during tumor development and organ ischemia. However, little is known regarding hypoxic regulation of genes that are directly involved in cell death or death resistance. Here we report the striking induction by severe hypoxia of the anti-apoptotic protein IAP-2. Hypoxic cells with IAP-2 up-regulation became resistant to apoptosis. IAP-2 was induced by hypoxia per se rather than by the secondary effects of hypoxia, including ATP depletion and cell injury. The inductive response did not relate to alterations of cellular redox status or arrest of mitochondrial respiration. On the other hand, IAP-2 induction was attenuated by actinomycin D, suggesting a role for gene transcription. In vitro nuclear run-on assays demonstrated specific increases in IAP-2 transcriptional activity after hypoxia exposure. HIF-1, the primary transcription factor that is responsible for multiple gene activation under hypoxia, does not have a role in IAP-2 expression. HIF-1 and IAP-2 were induced by different degrees of hypoxia; severe hypoxia or anoxia was required for IAP-2 induction. Moreover, cobalt chloride and desferrioxamine activated HIF-1 but not IAP-2. Finally, IAP-2 was induced by severe hypoxia in mouse embryonic stem cells that were deficient of HIF-1. Thus, this study not only provides the first demonstration of hypoxic regulation of an anti-apoptotic gene but also suggests the participation of novel hypoxia-responsive transcription mechanisms. PMID- 11278988 TI - A novel ganglioside isolated from renal cell carcinoma. AB - In renal cell carcinoma (RCC), the level of higher gangliosides is correlated with degree of metastatic potential, and cell lines derived from metastatic deposits of RCC are characterized by high expression of disialogangliosides (Saito, S., Orikasa, S., Ohyama, C., Satoh, M., and Fukushi, Y. (1991) Int. J. Cancer 49, 329-334 and Saito, S., Orikasa, S., Satoh, M., Ohyama, C., Ito, A., and Takahashi, T. (1997) Jpn. J. Cancer Res. (Gann) 88, 652-659). We now report two disialogangliosides, G1 and G2, found in the RCC cell line TOS-1. G1 from TOS 1 cells was characterized as having a novel hybrid structure between ganglio series (region I as in Structure; same as the terminal structure of ganglioside GM2), and the lacto-series type 1 (region II). The characterization was based on reactivity with various monoclonal antibodies (mAbs) with defined epitope specificity, as well as monosaccharide and fatty acid component analysis, (1)H NMR spectroscopy, and electrospray ionization mass spectrometry of the intact compound. G1 showed strong reactivity with mAb RM2, raised originally against TOS 1 cells, and weak cross-reactivity with anti-GM2 mAb MK-1-8. The antigen is hereby termed GalNAc disialosyl Lc4Cer (IV4GalNAcIV3NeuAcIII6NeuAcLc4; abbreviated GalNAcDSLc4). G2 was identified by 1H NMR and mass spectrometry as having a structure similar to Structure but without the GalNAcbeta1-->4 substitution and showed strong reactivity with mAb FH9 reported previously to be specific for disialosyl lacto-series type 1 (disialosyl Lc(4)) having vicinal alpha2-->3 and alpha2-->6 sialosyl residues, an antigen associated with human colonic cancer. Clinicopathological studies indicate that expression of these disialoganglioside antigens in RCC tissue is correlated with the metastatic potential of RCC. PMID- 11278990 TI - Inhibition of the nuclear factor kappa B (NF-kappa B) pathway by tetracyclic kaurene diterpenes in macrophages. Specific effects on NF-kappa B-inducing kinase activity and on the coordinate activation of ERK and p38 MAPK. AB - The anti-inflammatory action of most terpenes has been explained in terms of the inhibition of nuclear factor kappaB (NF-kappaB) activity. Ent-kaurene diterpenes are intermediates of the synthesis of gibberellins and inhibit the expression of NO synthase-2 and the release of tumor necrosis factor-alpha in J774 macrophages challenged with lipopolysaccharide. These diterpenes inhibit NF-kappaB and IkappaB kinase (IKK) activation in vivo but failed to affect in vitro the function of NF-kappaB, the phosphorylation and targeting of IkappaBalpha, and the activity of IKK-2. Transient expression of NF-kappaB-inducing kinase (NIK) activated the IKK complex and NF-kappaB, a process that was inhibited by kaurenes, indicating that the inhibition of NIK was one of the targets of these diterpenes. These results show that kaurenes impair the inflammatory signaling by inhibiting NIK, a member of the MAPK kinase superfamily that interacts with tumor necrosis factor receptor-associated factors, and mediate the activation of NF kappaB by these receptors. Moreover, kaurenes delayed the phosphorylation of p38, ERK1, and ERK2 MAPKs, but not that of JNK, in response to lipopolysaccharide treatment of J774 cells. The absence of a coordinate activation of MAPK and IKK might contribute to a deficient activation of NF-kappaB that is involved in the anti-inflammatory activity of these molecules. PMID- 11278991 TI - Specific phosphorylation of nucleophosmin on Thr(199) by cyclin-dependent kinase 2-cyclin E and its role in centrosome duplication. AB - The kinase activity of cyclin-dependent kinase 2 (CDK2)-cyclin E is required for centrosomes to initiate duplication. We have recently found that nucleophosmin (NPM/B23), a phosphoprotein primarily found in nucleolus, associates with unduplicated centrosomes and is a direct substrate of CDK2-cyclin E in centrosome duplication. Upon phosphorylation by CDK2-cyclin E, NPM/B23 dissociates from centrosomes, which is a prerequisite step for centrosomes to initiate duplication. Here, we identified that threonine 199 (Thr(199)) of NPM/B23 is the major phosphorylation target site of CDK2-cyclin E in vitro, and the same site is phosphorylated in vivo. NPM/T199A, a nonphosphorylatable NPM/B23 substitution mutant (Thr(199) --> Ala) acts as dominant negative when expressed in cells, resulting in specific inhibition of centrosome duplication. As expected, NPM/T199A remains associated with the centrosomes. These observations provide direct evidence that the CDK2-cyclin E-mediated phosphorylation on Thr(199) determines association and dissociation of NPM/B23 to the centrosomes, which is a critical control for the centrosome to initiate duplication. PMID- 11278992 TI - Conformational change of elongation factor Tu (EF-Tu) induced by antibiotic binding. Crystal structure of the complex between EF-Tu.GDP and aurodox. AB - Aurodox is a member of the family of kirromycin antibiotics, which inhibit protein biosynthesis by binding to elongation factor Tu (EF-Tu). We have determined the crystal structure of the 1:1:1 complex of Thermus thermophilus EF Tu with GDP and aurodox to 2.0-A resolution. During its catalytic cycle, EF-Tu adopts two strikingly different conformations depending on the nucleotide bound: the GDP form and the GTP form. In the present structure, a GTP complex-like conformation of EF-Tu is observed, although GDP is bound to the nucleotide binding site. This is consistent with previous proposals that aurodox fixes EF-Tu on the ribosome by locking it in its GTP form. Binding of EF-Tu.GDP to aminoacyl tRNA and mutually exclusive binding of kirromycin and elongation factor Ts to EF Tu can be explained on the basis of the structure. For many previously observed mutations that provide resistance to kirromycin, it can now be understood how they prevent interaction with the antibiotic. An unexpected feature of the structure is the reorientation of the His-85 side chain toward the nucleotide binding site. We propose that this residue stabilizes the transition state of GTP hydrolysis, explaining the acceleration of the reaction by kirromycin-type antibiotics. PMID- 11278993 TI - Inhibition of endosomal insulin-like growth factor-I processing by cysteine proteinase inhibitors blocks receptor-mediated functions. AB - The receptor for the type 1 insulin-like growth factor (IGF-I) has been implicated in cellular transformation and the acquisition of an invasive/metastatic phenotype in various tumors. Following ligand binding, the IGF-I receptor is internalized, and the receptor.ligand complex dissociates as the ligand is degraded by endosomal proteinases. In the present study we show that the inhibition of endosomal IGF-I-degrading enzymes in human breast and murine lung carcinoma cells by the cysteine proteinase inhibitors, E-64 and CA074 methyl ester, profoundly altered receptor trafficking and signaling. In treated cells, intracellular ligand degradation was blocked, and although the receptor and two substrates, Shc and Insulin receptor substrate, were hyperphosphorylated on tyrosine, IGF-I-induced DNA synthesis, anchorage-independent growth, and matrix metalloproteinase synthesis were inhibited. The results suggest that ligand processing by endosomal proteinases is a key step in receptor signaling and function and a potential target for therapy. PMID- 11278995 TI - A HECT domain E3 enzyme assembles novel polyubiquitin chains. AB - Although polyubiquitin chains linked through Lys(29) of ubiquitin have been implicated in the targeting of certain substrates to proteasomes, the signaling properties of these chains are poorly understood. We previously described a ubiquitin-protein isopeptide ligase (E3) from erythroid cells that assembles polyubiquitin chains through either Lys(29) or Lys(48) of ubiquitin (Mastrandrea, L. D., You, J., Niles, E. G., and Pickart, C. M. (1999) J. Biol. Chem. 274, 27299 27306). Here we describe the purification of this E3 based on its affinity for a linear fusion of ubiquitin to the ubiquitin-conjugating enzyme UbcH5A. Among five major polypeptides in the affinity column eluate, the activity of interest was assigned to the product of a previously cloned human cDNA known as KIAA10 (Nomura, N., Miyajima, N., Sazuka, T., Tanaka, A., Kawarabayasi, Y., Sato, S., Nagase, T., Seki, N., Ishikawa, K., and Tabata, S. (1994) DNA Res. 1, 27-35). The KIAA10 protein is a member of the HECT (homologous to E6-AP carboxyl terminus) domain family of E3s. These E3s share a conserved C-terminal (HECT) domain that functions in the catalysis of ubiquitination, while their divergent N-terminal domains function in cognate substrate binding (Huibregtse, J. M., Scheffner, M., Beaudenon, S., and Howley, P. M. (1995) Proc. Natl. Acad. Sci. U. S. A. 92, 2563 2567). Recombinant KIAA10 catalyzed the assembly of both Lys(29)- and Lys(48) linked polyubiquitin chains. Surprisingly, the C-terminal 428 residues of KIAA10 were both necessary and sufficient for this activity, suggesting that the ability to assemble polyubiquitin chains may be a general property of HECT domains. The N terminal domain of KIAA10 interacted in vitro with purified 26 S proteasomes and with the isolated S2/Rpn1 subunit of the proteasome's 19 S regulatory complex, suggesting that the N-terminal domains of HECT E3s may function in proteasome binding as well as substrate binding. PMID- 11278994 TI - Pyridoxal phosphate binding sites are similar in human heme-dependent and yeast heme-independent cystathionine beta-synthases. Evidence from 31P NMR and pulsed EPR spectroscopy that heme and PLP cofactors are not proximal in the human enzyme. AB - Two classes of cystathionine beta-synthases have been identified in eukaryotes, the heme-independent enzyme found in yeast and the heme-dependent form found in mammals. Both classes of enzymes catalyze a pyridoxal phosphate (PLP)-dependent condensation of serine and homocysteine to produce cystathionine. The role of the heme in the human enzyme and its location relative to the PLP in the active site are unknown. (31)P NMR spectroscopy revealed that spin-lattice relaxation rates of the phosphorus nucleus in PLP are similar in both the paramagnetic ferric (T(1) = 6.34 +/- 0.01 s) and the diamagnetic ferrous (T(1) = 5.04 +/- 0.06 s) enzyme, suggesting that the two cofactors are not proximal to each other. This is also supported by pulsed EPR studies that do not provide any evidence for strong or weak coupling between the phosphorus nucleus and the ferric iron. However, the (31)P signal in the reduced enzyme moved from 5.4 to 2.2 ppm, and the line width decreased from 73 to 16 Hz, providing the first structural evidence for transmission to the active site of an oxidation state change in the heme pocket. These results are consistent with a regulatory role for the heme as suggested by previous biochemical studies from our laboratory. The (31)P chemical shifts of the resting forms of the yeast and human enzymes are similar, suggesting that despite the difference in their heme content, the microenvironment of the PLP is similar in the two enzymes. The addition of the substrate, serine, resulted in an upfield shift of the phosphorus resonance in both enzymes, signaling formation of reaction intermediates. The resting enzyme spectra were recovered following addition of excess homocysteine, indicating that both enzymes retained catalytic activity during the course of the NMR experiment. PMID- 11278996 TI - Heavy chain ferritin enhances serine hydroxymethyltransferase expression and de novo thymidine biosynthesis. AB - We have elucidated a biochemical mechanism whereby changes in iron metabolism cause changes in folate-dependent one-carbon metabolism. Although animal and clinical studies have demonstrated that perturbations in iron status and metabolism alter folate metabolism, the biochemical mechanisms underlying these associations have yet to be identified. The effect of altered ferritin expression on folate metabolism was determined in human MCF-7 cells and SH-SY5Y neuroblastoma. Cells expressing rat heavy chain ferritin (HCF) exhibited markedly increased expression of the folate-dependent enzyme cytoplasmic serine hydroxymethyltransferase (cSHMT). These effects were not seen when rat light chain ferritin was expressed. Additionally, cSHMT expression was not altered when HCF expression was induced in MCF-7 cells cultured with supplemental ferric citrate. This indicates that cSHMT expression is increased by elevated HCF concentrations, independent of increased iron availability, suggesting that cSHMT expression may respond to HCF-induced chelation of the regulatory iron pool. Increased HCF expression did not alter cSHMT mRNA levels, but did increase translation rates of cSHMT mRNA. The increase in translation was mediated, at least in part, through the cSHMT 5'-untranslated region of the transcript. MCF-7 cells with increased expression of cSHMT displayed increased efficiency of de novo thymidylate biosynthesis, indicating that thymidylate synthesis is normally limited by cSHMT activity in MCF-7 cells. Our data suggest that the iron regulatory pool may play an important role in regulating folate metabolism and thereby thymidine biosynthesis. PMID- 11278997 TI - Thiazolidinediones but not metformin directly inhibit the steroidogenic enzymes P450c17 and 3beta -hydroxysteroid dehydrogenase. AB - Androgen biosynthesis requires 3beta-hydroxysteroid dehydrogenase type II (3betaHSDII) and the 17alpha-hydroxylase and 17,20-lyase activities of cytochrome P450c17. Thiazolidinedione and biguanide drugs, which are used to increase insulin sensitivity in type 2 diabetes, lower serum androgen concentrations in women with polycystic ovary syndrome. However, it is unclear whether this is secondary to increased insulin sensitivity or to direct effects on steroidogenesis. To investigate potential actions of these drugs on P450c17 and 3betaHSDII, we used "humanized yeast" that express these steroidogenic enzymes in microsomal environments. The biguanide metformin had no effect on either enzyme, whereas the thiazolidinedione troglitazone inhibited 3betaHSDII (K(I) = 25.4 +/- 5.1 microm) and both activities of P450c17 (K(I) for 17alpha-hydroxylase, 8.4 +/- 0.6 microm; K(I) for 17,20-lyase, 5.3 +/- 0.7 microm). The action of troglitazone on P450c17 was competitive, but it was mainly a noncompetitive inhibitor of 3betaHSDII. The thiazolidinediones rosiglitazone and pioglitazone exerted direct but weaker inhibitory effects on both P450c17 and 3betaHSDII. These differential effects of the thiazolidinediones do not correlate with their effects on insulin sensitivity, suggesting that distinct regions of the thiazolidinedione molecule mediate these two actions. Thus, thiazolidinediones inhibit two key enzymes in human androgen synthesis contributing to their androgen-lowering effects, whereas metformin affects androgen synthesis indirectly, probably by lowering circulating insulin concentrations. PMID- 11278999 TI - Classes of thiols that influence the activity of the skeletal muscle calcium release channel. AB - The skeletal muscle Ca(2+) release channel/ryanodine receptor (RyR1) is a prototypic redox-responsive ion channel. Nearly half of the 101 cysteines per RyR1 subunit are kept in a reduced (free thiol) state under conditions comparable with resting muscle. Here we assessed the effects of physiological determinants of cellular redox state (oxygen tension, reduced (GSH) or oxidized (GSSG) glutathione, and NO/O(2) (released by 3-morpholinosydnonimine)) on RyR1 redox state and activity. Oxidation of approximately 10 RyR1 thiols (from approximately 48 to approximately 38 thiols/RyR1 subunit) had little effect on channel activity. Channel activity increased reversibly as the number of thiols was further reduced to approximately 23/subunit, whereas more extensive oxidation (to approximately 13 thiols/subunit) inactivated the channel irreversibly. Neither S nitrosylation nor tyrosine nitration contributed to these effects. The results identify at least three functional classes of RyR1 thiols and suggest that 1) the channel may be protected from oxidation by a large reservoir of functionally inert thiols, 2) the channel may be designed to respond to moderate oxidative stress by a change in activation setpoint, and 3) the channel is susceptible to oxidative injury under more extensive conditions. PMID- 11278998 TI - Conserved regions of the Drosophila erect wing protein contribute both positively and negatively to transcriptional activity. AB - Genetic studies of the Drosophila erect wing (ewg) gene have revealed that ewg has an essential function in the embryonic nervous system and is required for the specification of certain muscle cells. We have found that EWG is a site-specific transcriptional activator, and we report here that evolutionarily conserved regions of EWG contribute both positively and negatively to transcriptional activity. Using gel mobility shift assays, we have shown that an EWG dimer binds specifically to DNA. In transfection assays, EWG activated expression of a reporter gene bearing specific binding sites. Analysis of deletion mutants and fusions of EWG to the Gal4 DNA binding domain has identified a transcriptional activation domain in the C terminus of EWG. Deletion analysis also revealed a novel inhibitory region in the N terminus of EWG. Strikingly, both the activation domain and the inhibitory region are conserved in EWG homologs including human nuclear respiratory factor 1 (NRF-1) and the sea urchin P3A2 protein. The strong conservation of elements that determine transcriptional activity suggests that the EWG, NRF-1, and P3A2 family of proteins shares common mechanisms of action and has maintained common functions across evolution. PMID- 11279000 TI - Alteration of the co-substrate selectivity of deacetoxycephalosporin C synthase. The role of arginine 258. AB - Deacetoxycephalosporin C synthase is an iron(II) 2-oxoglutaratedependent oxygenase that catalyzes the oxidative ring-expansion of penicillin N to deacetoxycephalosporin C. The wild-type enzyme is only able to efficiently utilize 2-oxoglutarate and 2-oxoadipate as a 2-oxoacid co-substrate. Mutation of arginine 258, the side chain of which forms an electrostatic interaction with the 5-carboxylate of the 2-oxoglutarate co-substrate, to a glutamine residue reduced activity to about 5% of the wild-type enzyme with 2-oxoglutarate. However, other aliphatic 2-oxoacids, which were not co-substrates for the wild-type enzyme, were utilized by the R258Q mutant. These 2-oxoacids "rescued" catalytic activity to the level observed for the wild-type enzyme as judged by penicillin N and G conversion. These co-substrates underwent oxidative decarboxylation as observed for 2-oxoglutarate in the normal reaction with the wild-type enzyme. Crystal structures of the iron(II)- 2-oxo-3-methylbutanoate (1.5 A), and iron(II)-2-oxo-4 methylpentanoate (1.6 A) enzyme complexes were obtained, which reveal the molecular basis for this "chemical co-substrate rescue" and help to rationalize the co-substrate selectivity of 2-oxoglutaratedependent oxygenases. PMID- 11279001 TI - Reconstitution of transcription from the human U6 small nuclear RNA promoter with eight recombinant polypeptides and a partially purified RNA polymerase III complex. AB - The human U6 small nuclear (sn) RNA core promoter consists of a proximal sequence element, which recruits the multisubunit factor SNAP(c), and a TATA box, which recruits the TATA box-binding protein, TBP. In addition to SNAP(c) and TBP, transcription from the human U6 promoter requires two well defined factors. The first is hB", a human homologue of the B" subunit of yeast TFIIIB generally required for transcription of RNA polymerase III genes, and the second is hBRFU, one of two human homologues of the yeast TFIIIB subunit BRF specifically required for transcription of U6-type RNA polymerase III promoters. Here, we have partially purified and characterized a RNA polymerase III complex that can direct transcription from the human U6 promoter when combined with recombinant SNAP(c), recombinant TBP, recombinant hB", and recombinant hBRFU. These results open the way to reconstitution of U6 transcription from entirely defined components. PMID- 11279002 TI - Identification of transcriptional enhancer factor-4 as a transcriptional modulator of CTP:phosphocholine cytidylyltransferase alpha. AB - CTP:phosphocholine cytidylyltransferase (CCT) is the rate-limiting and regulated enzyme of mammalian phosphatidylcholine biosynthesis. There are three isoforms, CCTalpha, CCTbeta1, and CCTbeta2. The mouse CCTalpha gene promoter is regulated by an enhancer element (Eb) located between -103 and -82 base pairs (5' GTTTTCAGGAATGCGGAGGTGG-3') upstream from the transcriptional start site (Bakovic, M., Waite, K., Tang, W., Tabas, I., and Vance, D. E. (1999) Biochim. Biophys. Acta 1436, 147-165). To identify the Eb-binding protein(s), we screened a mouse embryo cDNA library by the yeast one-hybrid system and obtained 19 positive clones. Ten cDNA clones were identified as transcriptional enhancer factor-4 (TEF 4). The TEF-binding consensus sequence, 5'-(A/T)(A/G)(A/G)(A/T)ATG(C/T)(G/A)-3', was identified within the Eb binding region. Gel-shift analysis using radiolabeled Eb fragment as a probe showed that cell extracts from yeast expressing hemagglutinin-tagged TEF-4 caused a marked band retardation that could be prevented with an anti-hemagglutinin antibody. When COS-7 cells were transfected with TEF-4, CCTalpha promoter-luciferase reporter activity and CCTalpha mRNA levels increased. A TEF-4 deletion mutant containing a DNA-binding domain, mTEA(+), stimulated the CCTalpha promoter activity, whereas protein lacking the DNA binding domain, mTEA(-), did not. Unexpectedly, when the ATG core of the TEF-4 binding consensus within the Eb region was mutated, promoter activity was enhanced rather than decreased. Thus, TEF-4 might act as a dual transcriptional modulator as follows: as a suppressor via its direct binding to the Eb element and as an activator via its interactions with the basal transcriptional machinery. These results provide the first evidence that TEF-4 is an important regulator of CCTalpha gene expression. PMID- 11279004 TI - Inhibition of cell migration by Abl family tyrosine kinases through uncoupling of Crk-CAS complexes. AB - c-Abl and the Abl-related gene product (Arg) are nonreceptor tyrosine kinases that regulate the actin cytoskeleton of cells by direct association with F-actin and localization to focal contacts. However, the biological significance of this interaction is not known. We show here that transfection of COS-7 cells with a kinase-inactive form of c-Abl (Abl) promotes c-Crk II/p130(CAS) (Crk-CAS) coupling, enhancing cell migration. Moreover, embryonic fibroblast cells isolated from mice devoid of endogenous Abl and Arg (abl-/- arg-/-) demonstrate increased Crk-CAS coupling and motility. Conversely, expression of a kinase-active form of Abl or reconstitution of abl-/- arg-/- cells with wild-type Abl prevents Crk-CAS coupling and inhibits cell migration. Thus, Abl and Arg kinases play a critical role in preventing cell migration through regulation of Crk and CAS adaptor protein complexes, which are necessary for cell movement. PMID- 11279003 TI - Critical proliferation-independent window for basic fibroblast growth factor repression of myogenesis via the p42/p44 MAPK signaling pathway. AB - In many cell types including myoblasts, growth factors control proliferation and differentiation, in part, via the mitogen-activated protein kinase (MAPK) pathway (also known as the extracellular regulated kinase (Erk) pathway). In C2C12 myoblast cells, insulin-like growth factor-1 and basic fibroblast growth factor (bFGF) activate MAPK/Erk, and both growth factors promote myoblast proliferation. However, these factors have opposing roles with respect to differentiation; insulin-like growth factor-1 enhances muscle cell differentiation, whereas bFGF inhibits the expression of the muscle-specific transcription factors MyoD and myogenin. Cells treated with bFGF and PD98059, a specific inhibitor of the MAPK pathway, show enhanced expression of the muscle-specific transcription factors MyoD and myogenin as compared with cells not exposed to this inhibitor. Inhibiting MAPK activity also enhances myoblast fusion and the expression of the late differentiation marker myosin heavy chain. Basic FGF mediated repression of muscle-specific genes does not result from continued cell proliferation, since bFGF-treated cells progress through only one round of cell division. We have identified a critical boundary 16 to 20 h after plating during which bFGF induced MAPK activity is able to repress myogenic gene expression and differentiation. Thus, the targets of MAPK that regulate myogenesis are functional at this time and their identification is in progress. PMID- 11279005 TI - The specificity of receptor binding by vascular endothelial growth factor-d is different in mouse and man. AB - Human vascular endothelial growth factor-D (VEGF-D) binds and activates VEGFR-2 and VEGFR-3, receptors expressed on vascular and lymphatic endothelial cells. As VEGFR-2 signals for angiogenesis and VEGFR-3 is thought to signal for lymphangiogenesis, it was proposed that VEGF-D stimulates growth of blood vessels and lymphatic vessels into regions of embryos and tumors. Here we report the unexpected finding that mouse VEGF-D fails to bind mouse VEGFR-2 but binds and cross-links VEGFR-3 as demonstrated by biosensor analysis with immobilized receptor domains and bioassays of VEGFR-2 and VEGFR-3 cross-linking. Mutation of amino acids in mouse VEGF-D to those in the human homologue indicated that residues important for the VEGFR-2 interaction are clustered at, or are near, the predicted receptor-binding surface. Coordinated expression of VEGF-D and VEGFR-3 in mouse embryos was detected in the developing skin where the VEGF-D gene was expressed in a layer of cells beneath the developing epidermis and VEGFR-3 was localized on a network of vessels immediately beneath the VEGF-D-positive cells. This suggests that VEGF-D and VEGFR-3 may play a role in establishing vessels of the skin by a paracrine mechanism. Our study of receptor specificity suggests that VEGF-D may have different biological functions in mouse and man. PMID- 11279006 TI - Functional signal peptides bind a soluble N-terminal fragment of SecA and inhibit its ATPase activity. AB - The selective recognition of pre-secretory proteins by SecA is essential to the process of protein export from Escherichia coli, yet very little is known about the requirements for recognition and the mode of binding of precursors to SecA. The major reason for this is the lack of a soluble system suitable for biophysical study of the SecA-precursor complex. Complicating the development of such a system is the likelihood that SecA interacts with the precursor in a high affinity, productive manner only when it is activated by binding to membrane and SecYEG. A critical aspect of the precursor/SecA interaction is that it is regulated by various SecA ligands (nucleotide, lipid, SecYEG) to facilitate the release of the precursor, most likely in a stepwise fashion, for translocation. Several recent reports show that functions of SecA can be studied using separated domains. Using this approach, we have isolated a proteolytically generated N terminal fragment of SecA, which is stably folded, has high ATPase activity, and represents an activated version of SecA. We report here that this fragment, termed SecA64, binds signal peptides with significantly higher affinity than does SecA. Moreover, the ATPase activity of SecA64 is inhibited by signal peptides to an extent that correlates with the ability of these signal peptides to inhibit either SecA translocation ATPase or in vitro protein translocation, arguing that the interaction with SecA64 is functionally significant. Thus, SecA64 offers a soluble, well defined system to study the mode of recognition of signal peptides by SecA and the regulation of signal peptide release. PMID- 11279007 TI - Localization of disulfide bonds in the frizzled module of Ror1 receptor tyrosine kinase. AB - The frizzled (FRZ) module is a novel module type that was first identified in G protein-coupled receptors of the frizzled and smoothened families and has since been shown to be present in several secreted frizzled-related proteins, in some modular proteases, in collagen XVIII, and in various receptor tyrosine kinases of the Ror family. The FRZ modules constitute the extracellular ligand-binding region of frizzled receptors and are known to mediate signals of WNT family members through these receptors. With an eye toward defining the structure of this important module family, we have expressed the FRZ domain of rat Ror1 receptor tyrosine kinase in Pichia pastoris. By proteolytic digestion and amino acid sequencing the disulfide bonds were found to connect the 10 conserved cysteines in a 1-5, 2-4, 3-8, 6-10, and 7-9 pattern. Circular dichroism and differential scanning calorimetry studies on the recombinant protein indicate that the disulfide-bonded FRZ module corresponds to a single, compact, and remarkably stable folding domain possessing both alpha-helices and beta-strands. PMID- 11279008 TI - Hemin-induced activation of the thioredoxin gene by Nrf2. A differential regulation of the antioxidant responsive element by a switch of its binding factors. AB - Thioredoxin plays an important role in various cellular processes through redox regulation. Here, we have demonstrated that thioredoxin expression is transcriptionally induced in K562 cells by hemin (ferriprotoporphyrin IX) through activation of a regulatory region positioned from -452 to -420 bp of the thioredoxin gene. Insertion of a mutation in the antioxidant responsive element (ARE)/AP-1 consensus binding sequence in this region abolished the response to hemin. With electrophoretic mobility shift and DNA affinity assays, we have shown that the NF-E2p45/small Maf complex constitutively binds to the ARE. The binding of the Nrf2/small Maf complex to ARE was induced by hemin, whereas the binding of Jun/Fos proteins to ARE was induced by phorbol 12-myristate 13-acetate, but not hemin. Hemin induced nuclear translocation of Nrf2 but did not affect nuclear expression of Jun/Fos proteins. Overexpression of Nrf2 augmented the response to hemin in a dose-dependent manner. In contrast, overexpression of the dominant negative mutant of Nrf2 suppressed hemin-induced activation through the ARE. We show here hemin-induced activation of the thioredoxin gene by Nrf2 through the ARE and propose a novel mechanism of the regulation of the ARE through a switch of its binding factors. PMID- 11279009 TI - Autologous biological response modification of the gonadotropin receptor. AB - It is generally held with respect to heterotrimeric guanine nucleotide binding protein-coupled receptors that binding of ligand stabilizes a conformation of receptor that activates adenylyl cyclase. It is not formally appreciated if, in the case of G-protein-coupled receptors with large extracellular domains (ECDs), ECDs directly participate in the activation process. The large ECD of the glycoprotein hormone receptors (GPHRs) is 350 amino acids in length, composed of seven leucine-rich repeat domains, and necessary and sufficient for high affinity binding of the glycoprotein hormones. Peptide challenge experiments to identify regions in the follicle-stimulating hormone (FSH) receptor (FSHR) ECD that could bind its cognate ligand identified only a single synthetic peptide corresponding to residues 221-252, which replicated a leucine-rich repeat domain of the FSHR ECD and which had intrinsic activity. This peptide inhibited human FSH binding to the human FSHR (hFSHR) and also inhibited human FSH-induced signal transduction in Y-1 cells expressing recombinant hFSHR. The hFSHR-(221-252) domain was not accessible to anti-peptide antibody probes, suggesting that this domain resides at an interface between the hFSHR ECD and transmembrane domains. CD spectroscopy of the peptide in dodecyl phosphocholine micelles showed an increase in the ordered structure of the peptide. CD and NMR spectroscopies of the peptide in trifluoroethanol confirmed that hFSHR-(221-252) has the propensity to form ordered secondary structure. Importantly and consistent with the foregoing results, dodecyl phosphocholine induced a significant increase in the ordered secondary structure of the purified hFSHR ECD as well. These data provide biophysical evidence of the influence of environment on GPHR ECD subdomain secondary structure and identify a specific activation domain that can autologously modify GPHR activity. PMID- 11279010 TI - Insect cells encode a class II alpha-mannosidase with unique properties. AB - Previously, we cloned and characterized an insect (Sf9) cell cDNA encoding a class II alpha-mannosidase with amino acid sequence and biochemical similarities to mammalian Golgi alpha-mannosidase II. Since then, it has been demonstrated that other mammalian class II alpha-mannosidases can participate in N-glycan processing. Thus, the present study was performed to evaluate the catalytic properties of the Sf9 class II alpha-mannosidase and to more clearly determine its relationship to mammalian Golgi alpha-mannosidase II. The results showed that the Sf9 enzyme is cobalt-dependent and can hydrolyze Man(5)GlcNAc(2) to Man(3)GlcNAc(2), but it cannot hydrolyze GlcNAcMan(5)GlcNAc(2). These data establish that the Sf9 enzyme is distinct from Golgi alpha-mannosidase II. This enzyme is not a lysosomal alpha-mannosidase because it is not active at acidic pH and it is localized in the Golgi apparatus. In fact, its sensitivity to swainsonine distinguishes the Sf9 enzyme from all other known mammalian class II alpha-mannosidases that can hydrolyze Man(5)GlcNAc(2). Based on these properties, we designated this enzyme Sf9 alpha-mannosidase III and concluded that it probably provides an alternate N-glycan processing pathway in Sf9 cells. PMID- 11279011 TI - The low density lipoprotein receptor-related protein modulates levels of matrix metalloproteinase 9 (MMP-9) by mediating its cellular catabolism. AB - Members of the matrix metalloproteinase (MMP) family of enzymes participate in matrix remodeling and share a number of structural and functional features. The activity of this family of proteinases is carefully regulated at the level of zymogen activation and by a family of specific inhibitors termed tissue inhibitors of metalloproteinases (TIMP). It is now becoming clear that levels of certain MMPs are modulated by their association with cellular receptors that mediate their rapid internalization and degradation. In the current investigation we report that the amount of MMP-9 in conditioned cell culture medium is significantly increased when mouse embryonic fibroblasts are grown in the presence of the 39-kDa receptor-associated protein (RAP), an antagonist of ligand binding to low density lipoprotein receptor-related protein (LRP). In vitro assays reveal that the MMP-9.TIMP-1 complex binds to LRP with high affinity and that the binding determinant for LRP appears to reside on MMP-9. Cell lines expressing LRP mediate the internalization of 125I-labeled MMP-9.TIMP-1 complexes, whereas cell lines genetically deficient in LRP show a diminished capacity to mediate the cellular catabolism of MMP-9.TIMP-1 complexes. The results demonstrate that LRP is a functional receptor for MMP-9 and suggest a major role for LRP in modulating remodeling of the extracellular matrix by regulating extracellular proteinase activity. PMID- 11279012 TI - The C terminus of sprouty is important for modulation of cellular migration and proliferation. AB - The Drosophila Sprouty (SPRY) protein has been shown to inhibit the actions of epidermal growth factor and fibroblast growth factor. However, the role of mammalian SPRY proteins has not been clearly elucidated. We postulated that human Sprouty2 (hSPRY2) is an inhibitor of cellular migration and proliferation. Indeed, using stably transfected HeLa cells, which expressed hemagglutinin (HA) tagged hSPRY2 or hSPRY2 tagged at the C terminus with red fluorescent protein, we demonstrated that hSPRY2 inhibits the migration of cells in response to serum, epidermal growth factor, fibroblast growth factor, and platelet-derived growth factor. Additionally, hSPRY2 also inhibited the growth of HeLa cells in response to serum. Previously, two C-terminal domains on hSPRY2, which are necessary for its colocalization with microtubules (residues 123-177) or translocation to membrane ruffles (residues 178-194), have been identified (Lim, J., Wong, E. S., Ong, S. H., Yusoff, P., Low, B. C., and Guy, G. R. (2000) J. Biol. Chem. 275, 32837-32845). Therefore, using TAT-tagged hSPRY2 and its mutants, we determined the role of these two C-terminal domains in the inhibition of cell migration and proliferation. Our data show that the deletion of either of these two regions in hSPRY2 abrogates its ability to modulate cell migration in response to different growth factors and proliferation in response to serum. Therefore, we conclude that hSPRY2 inhibits the actions of a number of growth factors, and its C terminus, which is homologous among various SPRY isoforms, is important in mediating its biological activity. PMID- 11279013 TI - ATP-dependent nucleosome remodeling and histone hyperacetylation synergistically facilitate transcription of chromatin. AB - Drosophila nucleosome remodeling factor (NURF) is an ISWI-containing protein complex that facilitates nucleosome mobility and transcriptional activation in an ATP-dependent manner. Numerous studies have implicated histone acetylation in transcriptional activation. We investigated the relative contributions of these two chromatin modifications to transcription in vitro of a chromatinized adenovirus E4 minimal promoter that contains binding sites for the GAL4-VP16 activator. We found that NURF could remodel chromatin and stimulate transcription irrespective of the acetylation status of histones. In contrast, hyperacetylation of histones in the absence of NURF was unable to stimulate transcription, suggesting that NURF-dependent chromatin remodeling is an obligatory step in E4 promoter activation. When chromatin templates were first hyperacetylated and then incubated with NURF, significantly greater transcription stimulation was observed. The results suggest that changes in chromatin induced by acetylation of histones and the mobilization of nucleosomes by NURF combine synergistically to facilitate transcription. Experiments using single and multiple rounds of transcription indicate that these chromatin modifications stimulate transcription preinitiation as well as reinitiation. PMID- 11279015 TI - Human T-cell leukemia virus type I tax repression of p73beta is mediated through competition for the C/H1 domain of CBP. AB - The Tax protein, encoded by the human T-cell leukemia virus type I (HTLV-I), is required for high level viral transcription and HTLV-I-associated malignant transformation. Although the precise mechanism of malignant transformation by Tax is unclear, it is well established that Tax represses the transcription function of the tumor suppressor p53, possibly accelerating the accumulation of genetic mutations that are critical in HTLV-I-mediated malignant transformation. Tax repression of p53 transcription function appears to occur, at least in part, through competition for the cellular coactivator CBP/p300. In this study, we characterize the effect of Tax on the p53 family member, p73. We demonstrate that Tax also represses the transcription function of p73beta and that the repression is reciprocal in vivo, consistent with the idea that both transcription factors may compete for CBP/p300 in vivo. We provide evidence showing that both Tax and p73 interact strongly with the C/H1 domain of CBP and that their binding to this region is mutually exclusive in vitro. This finding provides evidence supporting the idea that reciprocal transcriptional repression between Tax and p73 is mediated through coactivator competition. PMID- 11279016 TI - Overproduction of bacterial protein disulfide isomerase (DsbC) and its modulator (DsbD) markedly enhances periplasmic production of human nerve growth factor in Escherichia coli. AB - Production of eukaryotic proteins with multiple disulfide bonds in the Escherichia coli periplasm often encounters difficulty in obtaining soluble products with native structure. Human nerve growth factor beta (NGF) contains three disulfide bonds between nonconsecutive cysteine residues and forms insoluble aggregates when expressed in E. coli. We now report that overexpression of Dsb proteins known to catalyze formation and isomerization of disulfide bonds can substantially enhance periplasmic production of NGF. A set of pACYC184-based plasmids that permit dsb expression under the araB promoter were introduced into cells carrying a compatible plasmid that expresses NGF. The efficiency of periplasmic production of NGF fused to the OmpT signal peptide was strikingly improved by coexpression of DsbCD or DsbABCD proteins (up to 80% of total NGF produced). Coexpression of DsbAB was hardly effective, whereas that of DsbAC increased the total yield but not the periplasmic expression. These results suggest synergistic roles of DsbC and DsbD in disulfide isomerization that appear to become limiting upon NGF production. Furthermore, recombinant NGF produced with excess DsbCD (or DsbABCD) was biologically active judged by the neurite outgrowth assay using rat PC12 cells. PMID- 11279017 TI - Retinoblastoma protein is functionally distinct from its homologues in affecting glucocorticoid receptor-mediated transcription and apoptosis. AB - The cell cycle regulator, retinoblastoma protein, is known to potentiate glucocorticoid receptor-activated transcription through the interaction of its pocket domain with the transcription coactivator, hBRM. We now show that glucocorticoid receptor-induced apoptosis is also dependent on both the retinoblastoma protein and hBRM. p107 and p130, which share extensive sequence homology with the pocket domain of the retinoblastoma protein but not its N terminal region, also interact with hBRM but do not support either glucocorticoid receptor-dependent activity. This difference arises from the divergent N-terminal domain of the retinoblastoma protein, which, when fused to the pocket domains, confers upon p107 and p130 the ability to influence glucocorticoid receptor activities. This effect probably results from the promotion of glucocorticoid receptor-targeted chromatin remodeling by the hBRM-containing SWI/SNF complex because the N-terminal domain of the retinoblastoma protein enhances glucocorticoid receptor-hBRM interactions. These results highlight that, besides the interaction between hBRM and the pocket domain of RB, the N-terminal region of the retinoblastoma protein is also essential for glucocorticoid receptor induced apoptosis and the potentiation of glucocorticoid receptor-mediated transcription and provide a basis for functional distinction between the retinoblastoma protein and its homologues. PMID- 11279018 TI - Down-regulation of intrinsic P-glycoprotein expression in multicellular prostate tumor spheroids by reactive oxygen species. AB - Intrinsic expression of the multidrug resistance (MDR) transporter P-glycoprotein (Pgp) may be regulated by reactive oxygen species (ROS). A transient expression of Pgp was observed during the growth of multicellular tumor spheroids. Maximum Pgp expression occurred in tumor spheroids with a high percentage of quiescent, Ki-67-negative cells, elevated glutathione levels, increased expression of the cyclin-dependent kinase inhibitors p27Kip1 and p21WAF-1 as well as reduced ROS levels and minor activity of the mitogen-activated kinase (MAPK) members c-Jun amino-terminal kinase (JNK), extracellular signal-regulated kinase ERK1,2, and p38 MAPK. Raising intracellular ROS by depletion of glutathione with buthionine sulfoximine (BSO) or glutamine starvation resulted in down-regulation of Pgp and p27Kip1, whereas ERK1,2 and JNK were activated. Down-regulation of Pgp was furthermore observed with low concentrations of hydrogen peroxide and epidermal growth factor, indicating that ROS may regulate Pgp expression. The down regulation of Pgp following BSO treatment was abolished by agents interfering with receptor tyrosine kinase signaling pathways, i.e. the protein kinase C inhibitors bisindolylmaleimide I (BIM-1) and Ro-31-8220, the p21ras farnesyl protein transferase inhibitor III, the c-Raf inhibitor ZM 336372 and PD98059, which inhibits ERK1,2 activation. ROS involved as second messengers in receptor tyrosine kinase signaling pathways may act as negative regulators of Pgp expression. PMID- 11279019 TI - Influence of moderate temperatures on myristoyl-CoA metabolism and acyl-CoA thioesterase activity in the psychrophilic antarctic yeast Rhodotorula aurantiaca. AB - The inability of psychrophilic microorganisms to grow at moderate temperatures (>20 degrees C) presently represents an unresolved thermodynamic paradox. Here we report for the psychrophilic yeast Rhodotorula aurantiaca A19, isolated from Antarctic ice, that the inability to grow at temperatures close to 20 degrees C is associated with profound alterations in cell morphology and integrity. High performance liquid chromatography analysis of the intracellular acyl-CoA esters revealed an abnormal accumulation of myristoyl-CoA (C14-CoA) in cells cultivated close to the nonpermissive temperature. Its concentration (500 microm) was found to be 28-fold higher than in cells cultivated at 0 degrees C. If one considers its ability to disrupt membrane bilayers and to inhibit many cellular enzymes and functions, intracellular myristoyl-CoA accumulation in the psychrophile R. aurantiaca represents one of the principal causes of growth arrest at moderate temperatures. Intracellular acyl-CoA concentrations are believed to be regulated by thioesterase activity. Thus in an attempt to explore the mechanism by which temperature disrupts myristoyl-CoA metabolism, we isolated and characterized a long chain acyl-CoA thioesterase. The monomeric 80-kDa thioesterase from the psychrophilic yeast shows a very strong specificity for myristoyl-CoA. The affinity for substrate and the catalytic efficiency of the thioesterase are optimal below 5 degrees C (temperatures habitually experienced by the strain) and dramatically decrease with increasing temperature. The loss of affinity for substrate is related to the intracellular increase of myristoyl-CoA concentration. Our observations reveal one of the probable mechanisms by which temperature fixes the limit of growth for this psychrophilic yeast. PMID- 11279020 TI - Profiling changes in gene expression during differentiation and maturation of monocyte-derived dendritic cells using both oligonucleotide microarrays and proteomics. AB - Dendritic cells (DCs) are antigen-presenting cells that play a major role in initiating primary immune responses. We have utilized two independent approaches, DNA microarrays and proteomics, to analyze the expression profile of human CD14(+) blood monocytes and their derived DCs. Analysis of gene expression changes at the RNA level using oligonucleotide microarrays complementary to 6300 human genes showed that approximately 40% of the genes were expressed in DCs. A total of 255 genes (4%) were found to be regulated during DC differentiation or maturation. Most of these genes were not previously associated with DCs and included genes encoding secreted proteins as well as genes involved in cell adhesion, signaling, and lipid metabolism. Protein analysis of the same cell populations was done using two-dimensional gel electrophoresis. A total of 900 distinct protein spots were included, and 4% of them exhibited quantitative changes during DC differentiation and maturation. Differentially expressed proteins were identified by mass spectrometry and found to represent proteins with Ca(2+) binding, fatty acid binding, or chaperone activities as well as proteins involved in cell motility. In addition, proteomic analysis provided an assessment of post-translational modifications. The chaperone protein, calreticulin, was found to undergo cleavage, yielding a novel form. The combined oligonucleotide microarray and proteomic approaches have uncovered novel genes associated with DC differentiation and maturation and has allowed analysis of post-translational modifications of specific proteins as part of these processes. PMID- 11279022 TI - The structure of the multidrug resistance protein 1 (MRP1/ABCC1). crystallization and single-particle analysis. AB - Multidrug resistance protein 1 (MRP1/ABCC1) is an ATP-binding cassette (ABC) polytopic membrane transporter of considerable clinical importance that confers multidrug resistance on tumor cells by reducing drug accumulation by active efflux. MRP1 is also an efficient transporter of conjugated organic anions. Like other ABC proteins, including the drug resistance conferring 170-kDa P glycoprotein (ABCB1), the 190-kDa MRP1 has a core structure consisting of two membrane-spanning domains (MSDs), each followed by a nucleotide binding domain (NBD). However, unlike P-glycoprotein and most other ABC superfamily members, MRP1 contains a third MSD with five predicted transmembrane segments with an extracytosolic NH(2) terminus. Moreover, the two nucleotide-binding domains of MRP1 are considerably more divergent than those of P-glycoprotein. In the present study, the first structural details of MRP1 purified from drug-resistant lung cancer cells have been obtained by electron microscopy of negatively stained single particles and two-dimensional crystals formed after reconstitution of purified protein with lipids. The crystals display p2 symmetry with a single dimer of MRP1 in the unit cell. The overall dimensions of the MRP1 monomer are approximately 80 x 100 A. The MRP1 monomer shows some pseudo-2-fold symmetry in projection, and in some orientations of the detergent-solubilized particles, displays a stain filled depression (putative pore) appearing toward the center of the molecule, presumably to enable transport of substrates. These data represent the first structural information of this transporter to approximately 22-A resolution and provide direct structural evidence for a dimeric association of the transporter in a reconstituted lipid bilayer. PMID- 11279021 TI - The zinc finger domain of Tzfp binds to the tbs motif located at the upstream flanking region of the Aie1 (aurora-C) kinase gene. AB - Our previous studies showed that Aie1 (aurora-C), is a novel testis kinase belonging to the aurora kinase family (). In this report, we describe a testis zinc finger protein (Tzfp) that binds to the upstream flanking sequence of the Aie1 gene. The mouse Tzfp gene, mapped to chromosome 7 B2-B3, encodes a 465-amino acid transcription factor containing a conserved N-terminal BTB/POZ domain and three C-terminal PLZF-like C(2)H(2) zinc fingers. The zinc finger domain of Tzfp binds to the TGTACAGTGT motif (Tzfp binding site, termed tbs) located at the upstream flanking sequence of the Aie1 gene by gel mobility shift, DNase I footprinting, and competition analyses. When the C-terminal zinc fingers of Tzfp were fused to the transactivation domain of VP16, the chimera activated transcription of a reporter construct containing multiple copies of the tbs. In contrast, the same chimera did not activate the reporter gene when an essential nucleotide fifth C was mutated to A at the tbs. Furthermore, we showed that the N terminal BTB/POZ domain of TZFP has a repressor activity. Taken together, our results indicate that Tzfp recognizes a sequence-specific motif (tbs) and may play a role in the regulation of the genes carrying the tbs. PMID- 11279023 TI - The Quinone-binding sites of the Saccharomyces cerevisiae succinate-ubiquinone oxidoreductase. AB - The Saccharomyces cerevisiae succinate dehydrogenase (SDH) of the mitochondrial electron transport chain oxidizes succinate and reduces ubiquinone. Using a random mutagenesis approach, we identified functionally important amino acid residues in one of the anchor subunits, Sdh4p. We analyzed three point mutations (F69V, S71A, and H99L) and one nonsense mutation (Y89OCH) that truncates the Sdh4p subunit at the third predicted transmembrane segment. The F69V and the S71A mutations result in greatly impaired respiratory growth in vivo and quinone reductase activities in vitro, with negligible effects on enzyme stability. In contrast, the Y89OCH and the H99L mutations elicit large structural perturbations that impair assembly as evidenced by reduced covalent FAD levels, membrane associated succinate-phenazine methosulfate reductase activities, and thermal stability. We propose that the Phe-69 and the Ser-71 residues are involved in the formation of a quinone-binding site, whereas the His-99 residue is at the interface of the peripheral and the membrane domains. In addition, the properties of the Y89OCH mutation are consistent with the interpretation that the third transmembrane segment is not involved in catalysis but rather plays an important structural role. The mutant enzymes are differentially sensitive to a quinone analog inhibitor, providing further evidence for a two-quinone binding model in the yeast SDH. PMID- 11279024 TI - Regulation of beta-catenin structure and activity by tyrosine phosphorylation. AB - beta-Catenin plays a dual role as a key effector in the regulation of adherens junctions and as a transcriptional coactivator. Phosphorylation of Tyr-654, a residue placed in the last armadillo repeat of beta-catenin, decreases its binding to E-cadherin. We show here that phosphorylation of Tyr-654 also stimulates the association of beta-catenin to the basal transcription factor TATA binding protein. The structural bases of these different affinities were investigated. Our results indicate that the beta-catenin C-terminal tail interacts with the armadillo repeat domain, hindering the association of the armadillo region to the TATA-binding protein or to E-cadherin. Phosphorylation of beta-catenin Tyr-654 decreases armadillo-C-terminal tail association, uncovering the last armadillo repeats. In a C-terminal-depleted beta-catenin, the presence of a negative charge at Tyr-654 does not affect the interaction of the TATA binding protein to the armadillo domain. However, in the case of E-cadherin, the establishment of ion pairs dominates its association with beta-catenin, and its binding is greatly dependent on the absence of a negative charge at Tyr-654. Thus, phosphorylation of Tyr-654 blocks the Ecadherin-beta-catenin interaction, even though the steric hindrance of the C-tail is no longer present. These results explain how phosphorylation of beta-catenin in Tyr-654 modifies the tertiary structure of this protein and the interaction with its different partners. PMID- 11279025 TI - The FadR.DNA complex. Transcriptional control of fatty acid metabolism in Escherichia coli. AB - In Escherichia coli, the expression of fatty acid metabolic genes is controlled by the transcription factor, FadR. The affinity of FadR for DNA is controlled by long chain acyl-CoA molecules, which bind to the protein and modulate gene expression. The crystal structure of FadR reveals a two domain dimeric molecule where the N-terminal domains bind DNA, and the C-terminal domains bind acyl-CoA. The DNA binding domain has a winged-helix motif, and the C-terminal domain resembles the sensor domain of the Tet repressor. The FadR.DNA complex reveals how the protein interacts with DNA and specifically recognizes a palindromic sequence. Structural and functional similarities to the Tet repressor and the BmrR transcription factors suggest how the binding of the acyl-CoA effector molecule to the C-terminal domain may affect the DNA binding affinity of the N terminal domain. We suggest that the binding of acyl-CoA disrupts a buried network of charged and polar residues in the C-terminal domain, and the resulting conformational change is transmitted to the N-terminal domain via a domain spanning alpha-helix. PMID- 11279026 TI - Refined characterization of corneodesmosin proteolysis during terminal differentiation of human epidermis and its relationship to desquamation. AB - Corneodesmosin is a putative adhesion glycoprotein located in the extracellular part of the desmosomes in the upper layers of the epidermis. Synthesized by granular keratinocytes as a 52-56-kDa protein, corneodesmosin is progressively proteolysed during corneocyte maturation. This processing is a prerequisite for desquamation. Two glycine- and serine-rich domains of the protein might take on the conformation of adhesive secondary structures similar to glycine loops. Corneodesmosin proteolysis was further characterized. Deglycosylation experiments and reactivity with lectins demonstrated that the corneodesmosin carbohydrate moiety does not prevent the proteolysis. Immunoblotting, immunohistochemistry, and immunoelectron microscopy experiments using affinity-purified anti-peptide antibodies raised to four of the five structural domains of corneodesmosin and a monoclonal antibody against its fifth central domain showed that the first step in corneodesmosin processing is the cleavage of its extremities and probably occurs before its incorporation into desmosomes. Then the glycine loop-related domains are cleaved, first the N-terminal and then part of the C-terminal domain. At the epidermis surface, the multistep proteolytic cleavage leaves intact only the central domain, which was detected on exfoliated corneocytes and probably lacks adhesive properties. Importantly, corneodesmosin was demonstrated to be a preferred substrate of two serine proteases involved in desquamation, the stratum corneum tryptic and chymotryptic enzymes. PMID- 11279027 TI - Receptor-mediated inhibition of G protein-coupled inwardly rectifying potassium channels involves G(alpha)q family subunits, phospholipase C, and a readily diffusible messenger. AB - G protein-coupled inwardly rectifying K+ (GIRK) channels can be activated or inhibited by distinct classes of receptor (G(alpha)i/o- and G(alpha)q-coupled), providing dynamic regulation of cellular excitability. Receptor-mediated activation involves direct effects of G(beta)gamma subunits on GIRK channels, but mechanisms involved in GIRK channel inhibition have not been fully elucidated. An HEK293 cell line that stably expresses GIRK1/4 channels was used to test G protein mechanisms that mediate GIRK channel inhibition. In cells transiently or stably cotransfected with 5-HT1A (G(alpha)i/o-coupled) and TRH-R1 (G(alpha)q coupled) receptors, 5-HT (5-hydroxytryptamine; serotonin) enhanced GIRK channel currents, whereas thyrotropin-releasing hormone (TRH) inhibited both basal and 5 HT-activated GIRK channel currents. Inhibition of GIRK channel currents by TRH primarily involved signaling by G(alpha)q family subunits, rather than G(beta)gamma dimers: GIRK channel current inhibition was diminished by Pasteurella multocida toxin, mimicked by constitutively active members of the G(alpha)q family, and reduced by minigene constructs that disrupt G(alpha)q signaling, but was completely preserved in cells expressing constructs that interfere with signaling by G(beta)gamma subunits. Inhibition of GIRK channel currents by TRH and constitutively active G(alpha)q was reduced by, an inhibitor of phospholipase C (PLC). Moreover, TRH- R1-mediated GIRK channel inhibition was diminished by minigene constructs that reduce membrane levels of the PLC substrate phosphatidylinositol bisphosphate, further implicating PLC. However, we found no evidence for involvement of protein kinase C, inositol trisphosphate, or intracellular calcium. Although these downstream signaling intermediaries did not contribute to receptor-mediated GIRK channel inhibition, bath application of TRH decreased GIRK channel activity in cell-attached patches. Together, these data indicate that receptor-mediated inhibition of GIRK channels involves PLC activation by G(alpha) subunits of the G(alpha)q family and suggest that inhibition may be communicated at a distance to GIRK channels via unbinding and diffusion of phosphatidylinositol bisphosphate away from the channel. PMID- 11279028 TI - Cooperative activation by GATA-4 and YY1 of the cardiac B-type natriuretic peptide promoter. AB - YY1, a multifunctional protein essential for embryonic development, is a known repressor or activator of transcription. In cardiac and skeletal myocytes, YY1 has been described essentially as a negative regulator of muscle-specific genes. In this study, we report that YY1 is a transcriptional activator of the B-type natriuretic peptide (BNP) gene, which encodes one of the heart major secretory products. YY1 binds an element within the proximal cardiac BNP promoter, in close proximity to the high affinity binding sites for the zinc finger GATA proteins. We show that YY1 cooperates with GATA-4 to synergistically activate BNP transcription. Structure-function analysis revealed that the DNA binding domain of YY1 is sufficient for cooperative interaction with GATA-4, likely through corecruitment of the CREB-binding protein coactivator. The results suggest that YY1 and GATA factors are components of transcriptionally active complexes present in cardiac and other GATA-containing cells. PMID- 11279029 TI - Biosynthesis of 9-cis-retinoic acid in vivo. The roles of different retinol dehydrogenases and a structure-activity analysis of microsomal retinol dehydrogenases. AB - Retinoic acid is generated by a two-step mechanism. First, retinol is converted into retinal by a retinol dehydrogenase, and, subsequently, retinoic acid is formed by a retinal dehydrogenase. In vitro, several enzymes are suggested to act in this metabolic pathway. However, little is known regarding their capacity to contribute to retinoic acid biosynthesis in vivo. We have developed a versatile cell reporter system to analyze the role of several of these enzymes in 9-cis retinoic acid biosynthesis in vivo. Using a Gal4-retinoid X receptor fusion protein-based luciferase reporter assay, the formation of 9-cis-retinoic acid from 9-cis-retinol was measured in cells transfected with expression plasmids encoding different combinations of retinol and retinal dehydrogenases. The results suggested that efficient formation of 9-cis-retinoic acid required co expression of retinol and retinal dehydrogenases. Interestingly, the cytosolic alcohol dehydrogenase 4 failed to efficiently catalyze 9-cis-retinol oxidation. A structure-activity analysis showed that mutants of two retinol dehydrogenases, devoid of the carboxyl-terminal cytoplasmic tails, displayed greatly reduced enzymatic activities in vivo, but were active in vitro. The cytoplasmic tails mediate efficient endoplasmic reticulum localization of the enzymes, suggesting that the unique milieu in the endoplasmic reticulum compartment is necessary for in vivo activity of microsomal retinol dehydrogenases. PMID- 11279030 TI - Ponericins, new antibacterial and insecticidal peptides from the venom of the ant Pachycondyla goeldii. AB - The antimicrobial, insecticidal, and hemolytic properties of peptides isolated from the venom of the predatory ant Pachycondyla goeldii, a member of the subfamily Ponerinae, were investigated. Fifteen novel peptides, named ponericins, exhibiting antibacterial and insecticidal properties were purified, and their amino acid sequences were characterized. According to their primary structure similarities, they can be classified into three families: ponericin G, W, and L. Ponericins share high sequence similarities with known peptides: ponericins G with cecropin-like peptides, ponericins W with gaegurins and melittin, and ponericins L with dermaseptins. Ten peptides were synthesized for further analysis. Their antimicrobial activities against Gram-positive and Gram-negative bacteria strains were analyzed together with their insecticidal activities against cricket larvae and their hemolytic activities. Interestingly, within each of the three families, several peptides present differences in their biological activities. The comparison of the structural features of ponericins with those of well-studied peptides suggests that the ponericins may adopt an amphipathic alpha helical structure in polar environments, such as cell membranes. In the venom, the estimated peptide concentrations appear to be compatible with an antibacterial activity in vivo. This suggests that in the ant colony, the peptides exhibit a defensive role against microbial pathogens arising from prey introduction and/or ingestion. PMID- 11279031 TI - The zinc finger protein 202 (ZNF202) is a transcriptional repressor of ATP binding cassette transporter A1 (ABCA1) and ABCG1 gene expression and a modulator of cellular lipid efflux. AB - The zinc finger gene 202 (ZNF202) located within a hypoalphalipoproteinemia susceptibility locus on chromosome 11q23 is a transcriptional repressor of various genes involved in lipid metabolism. To provide further evidence for a functional linkage between ZNF202 and hypoalphalipoproteinemia, we investigated the effect of ZNF202 expression on ATP binding cassette transporter A1 (ABCA1) and ABCG1. ABCA1 is a key regulator of the plasma high density lipoprotein pool size, whereas ABCG1 is another mediator of cellular cholesterol and phospholipid efflux in human macrophage. We demonstrate here that the full-length ZNF202m1 isoform binds to GnT repeats within the promotors of ABCA1 (-229/-210) and ABCG1 (-572/-552). ZNF202m1 expression in HepG2 cells dose-dependently repressed the promotor activities of ABCA1 and ABCG1. This transcriptional effect required the presence of the SCAN domain in ZNF202 and the functional integrity of a TATA box at position -24 of ABCA1, whereas the presence of GnT binding motifs was nonessential. The state of ZNF202 SCAN domain oligomerization affected the ability of the adjacent ZNF202 Kruppel-associated box domain to recruit the transcriptional corepressor KAP1. Overexpression of ZNF202m1 in RAW264.7 macrophages prevented the induction of ABCA1 gene expression by 20(S)OH cholesterol and 9-cis-retinoic acid, further substantiating the interference of ZNF202 in critical elements of transcriptional activation. Finally, HDL and apoAImediated lipid efflux was significantly reduced in RAW264.7 cells stably expressing ZNF202m1. In conclusion, we have identified ABCA1 and ABCG1 as target genes for ZNF202-mediated repression and thus, provide evidence for a functional linkage between ZNF202 and hypoalphalipoproteinemia. PMID- 11279032 TI - Human UDP-galactose 4-epimerase. Accommodation of UDP-N-acetylglucosamine within the active site. AB - UDP-galactose 4-epimerase catalyzes the interconversion of UDP-galactose and UDP glucose during normal galactose metabolism. One of the key structural features in the proposed reaction mechanism for the enzyme is the rotation of a 4' ketopyranose intermediate within the active site pocket. Recently, the three dimensional structure of the human enzyme with bound NADH and UDP-glucose was determined. Unlike that observed for the protein isolated from Escherichia coli, the human enzyme can also turn over UDP-GlcNAc to UDP-GalNAc and vice versa. Here we describe the three-dimensional structure of human epimerase complexed with NADH and UDP-GlcNAc. To accommodate the additional N-acetyl group at the C2 position of the sugar, the side chain of Asn-207 rotates toward the interior of the protein and interacts with Glu-199. Strikingly, in the human enzyme, the structural equivalent of Tyr-299 in the E. coli protein is replaced with a cysteine residue (Cys-307) and the active site volume for the human protein is calculated to be approximately 15% larger than that observed for the bacterial epimerase. This combination of a larger active site cavity and amino acid residue replacement most likely accounts for the inability of the E. coli enzyme to interconvert UDP-GlcNAc and UDP-GalNAc. PMID- 11279033 TI - Dipeptidyl peptidase I is essential for activation of mast cell chymases, but not tryptases, in mice. AB - Dipeptidyl peptidase I (DPPI) is the sole activator in vivo of several granule associated serine proteases of cytotoxic lymphocytes. In vitro, DPPI also activates mast cell chymases and tryptases. To determine whether DPPI is essential for their activation in vivo, we used enzyme histochemical and immunohistochemical approaches and solution-based activity assays to study these enzymes in tissues and bone marrow-derived mast cells (BMMCs) from DPPI +/+ and DPPI -/- mice. We find that DPPI -/- mast cells contain normal amounts of immunoreactive chymases but no chymase activity, indicating that DPPI is essential for chymase activation and suggesting that DPPI -/- mice are functional chymase knockouts. The absence of DPPI and chymase activity does not affect the growth, granularity, or staining characteristics of BMMCs and, despite prior predictions, does not alter IgE-mediated exocytosis of histamine. In contrast, the level of active tryptase (mMCP-6) in DPPI -/- BMMCs is 25% that of DPPI +/- BMMCs. These findings indicate that DPPI is not essential for mMCP-6 activation but does influence the total amount of active mMCP-6 in mast cells and therefore may be an important, but not exclusive mechanism for tryptase activation. PMID- 11279034 TI - Apolipoprotein A-II modulates the binding and selective lipid uptake of reconstituted high density lipoprotein by scavenger receptor BI. AB - High density lipoprotein (HDL) represents a mixture of particles containing either apoA-I and apoA-II (LpA-I/A-II) or apoA-I without apoA-II (LpA-I). Differences in the function and metabolism of LpA-I and LpA-I/A-II have been reported, and studies in transgenic mice have suggested that apoA-II is pro atherogenic in contrast to anti-atherogenic apoA-I. The molecular basis for these observations is unclear. The scavenger receptor BI (SR-BI) is an HDL receptor that plays a key role in HDL metabolism. In this study we investigated the abilities of apoA-I and apoA-II to mediate SR-BI-specific binding and selective uptake of cholesterol ester using reconstituted HDLs (rHDLs) that were homogeneous in size and apolipoprotein content. Particles were labeled in the protein (with (125)I) and in the lipid (with [(3)H]cholesterol ether) components and SR-BI-specific events were analyzed in SR-BI-transfected Chinese hamster ovary cells. At 1 microg/ml apolipoprotein, SR-BI-mediated cell association of palmitoyloleoylphosphatidylcholine-containing AI-rHDL was significantly greater (3-fold) than that of AI/AII-rHDL, with a lower K(d) and a higher B(max) for AI rHDL as compared with AI/AII-rHDL. Unexpectedly, selective cholesterol ester uptake from AI/AII-rHDL was not compromised compared with AI-rHDL, despite decreased binding. The efficiency of selective cholesterol ester uptake in terms of SR-BI-associated rHDL was 4-5-fold greater for AI/AII-rHDL than AI-rHDL. These results are consistent with a two-step mechanism in which SR-BI binds ligand and then mediates selective cholesterol ester uptake with an efficiency dependent on the composition of the ligand. ApoA-II decreases binding but increases selective uptake. These findings show that apoA-II can exert a significant influence on selective cholesterol ester uptake by SR-BI and may consequently influence the metabolism and function of HDL, as well as the pathway of reverse cholesterol transport. PMID- 11279035 TI - Functional analysis of the trypanosomal AAA protein TbVCP with trans-dominant ATP hydrolysis mutants. AB - TbVCP is a member of the AAA (ATPases Associated with a variety of cellular Activities) family of proteins containing two ATPase domains. Southern analysis indicates TbVCP to have a single-locus, two-copy, genomic organization. One copy, but not both, can be disrupted by targeted gene replacement, suggesting that TbVCP is essential for trypanosome viability. Site-directed mutagenesis of the ATP hydrolysis motifs indicates that the second conserved ATPase domain is essential for TbVCP activity. Constitutive overexpression of TbVCP with a single mutation in the second hydrolysis motif or with mutations in both hydrolysis motifs was not possible. Regulated overexpression of these mutants resulted in cell death as a dominant negative phenotype. In each case cell growth arrested at 24-h post-induction and at all stages of the cell cycle as judged by replication of nuclear and kinetoplast genomes. Onset of growth arrest coincided with the development of severe and characteristic morphological alterations for each mutant. Neither constitutive nor regulated overexpression of wild type TbVCP or the single first hydrolysis domain mutant had any overt effect on cell viability or morphology. However, the distinct phenotype of the double mutant indicates that the first hydrolysis domain, although not essential, does modulate overall TbVCP function. Finally, yeast complementation studies demonstrated that TbVCP can functionally replace the yeast homologue Cdc48p, indicating that protein.protein interactions essential to function have been maintained over great phylogenetic distances. PMID- 11279036 TI - Coupling of cholesterol and cone-shaped lipids in bilayers augments membrane permeabilization by the cholesterol-specific toxins streptolysin O and Vibrio cholerae cytolysin. AB - Vibrio cholerae cytolysin (VCC) forms oligomeric pores in lipid bilayers containing cholesterol. Membrane permeabilization is inefficient if the sterol is embedded within bilayers prepared from phosphatidylcholine only but is greatly enhanced if the target membrane also contains ceramide. Although the enhancement of VCC action is stereospecific with respect to cholesterol, we show here that no such specificity applies to the two stereocenters in ceramide; all four stereoisomers of ceramide enhanced VCC activity in cholesterol-containing bilayers. A wide variety of ceramide analogs were as effective as D-erythro ceramide, as was diacylglycerol, suggesting that the effect of ceramide exemplifies a general trend of lipids with a small headgroup to augment the activity of VCC. Incorporation of these cone-shaped lipids into cholesterol containing bilayers also gave similar effects with streptolysin O, another cholesterol-specific but structurally unrelated cytolysin. In contrast, the activity of staphylococcal alpha-hemolysin, which does not share with the other toxins the requirement for cholesterol, was far less affected by the presence of lipids with a conical shape. The collective data indicate that sphingolipids and glycerolipids do not interact with the cytolysins specifically. Instead, lipids that have a conical molecular shape appear to effect a change in the energetic state of membrane cholesterol that in turn augments the interaction of the sterol with the cholesterol-specific cytolysins. PMID- 11279037 TI - Two WD repeat-containing TATA-binding protein-associated factors in fission yeast that suppress defects in the anaphase-promoting complex. AB - The general transcription factor IID consists of the TATA-binding protein (TBP) and multiple TBP-associated factors (TAFs). Here we report the isolation of two related TAF genes from the fission yeast Schizosaccharomyces pombe as multicopy suppressors of a temperature-sensitive mutation in the ubiquitin-conjugating enzyme gene ubcP4(+). The ubcP4(ts) mutation causes cell cycle arrest in mitosis, probably due to defects in ubiquitination mediated by the anaphase-promoting complex/cyclosome. One multicopy suppressor is the previously reported gene taf72(+), whereas the other is a previously unidentified gene named taf73(+). We show that the taf73(+) gene, like taf72(+), is essential for cell viability. The taf72(+) and taf73(+) genes encode proteins homologous to WD repeat-containing TAFs such as human TAF100, Drosophila TAF80/85, and Saccharomyces cerevisiae TAF90. We demonstrate that TAF72 and TAF73 proteins are present in the same complex with TBP and other TAFs and that TAF72, but not TAF73, is associated with the putative histone acetylase Gcn5. We also show that overexpression of TAF72 or TAF73 suppresses the cell cycle arrest in mitosis caused by a mutation in the anaphase-promoting complex/cyclosome subunit gene cut9(+). These results suggest that TAF72 and TAF73 may regulate the expression of genes involved in ubiquitin dependent proteolysis during mitosis. Our study thus provides evidence for a possible role of WD repeat-containing TAFs in the expression of genes involved in progression through the M phase of the cell cycle. PMID- 11279038 TI - Interactions between the Werner syndrome helicase and DNA polymerase delta specifically facilitate copying of tetraplex and hairpin structures of the d(CGG)n trinucleotide repeat sequence. AB - Werner syndrome (WS) is an inherited disorder characterized by premature aging and genomic instability. The protein encoded by the WS gene, WRN, possesses intrinsic 3' --> 5' DNA helicase and 3' --> 5' DNA exonuclease activities. WRN helicase resolves alternate DNA structures including tetraplex and triplex DNA, and Holliday junctions. Thus, one function of WRN may be to unwind secondary structures that impede cellular DNA transactions. We report here that hairpin and G'2 bimolecular tetraplex structures of the fragile X expanded sequence, d(CGG)(n), effectively impede synthesis by three eukaryotic replicative DNA polymerases (pol): pol alpha, pol delta, and pol epsilon. The constraints imposed on pol delta-catalyzed synthesis are relieved, however, by WRN; WRN facilitates pol delta to traverse these template secondary structures to synthesize full length DNA products. The alleviatory effect of WRN is limited to pol delta; neither pol alpha nor pol epsilon can traverse template d(CGG)(n) hairpin and tetraplex structures in the presence of WRN. Alleviation of pausing by pol delta is observed with Escherichia coli RecQ but not with UvrD helicase, suggesting a concerted action of RecQ helicases and pol delta. Our findings suggest a possible role of WRN in rescuing pol delta-mediated replication at forks stalled by unusual DNA secondary structures. PMID- 11279039 TI - RhoA inhibits the nerve growth factor-induced Rac1 activation through Rho associated kinase-dependent pathway. AB - The Rho family of small GTPases has been shown to be involved in the regulation of neuronal morphology, and Rac and Rho exert antagonistic actions in neurite formation. In this study, we have examined the cross-talk between Rac and Rho in relation to the nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells. NGF induced a rapid activation of Rac1 and suppression of RhoA activity. Constitutively active RhoA, RhoA(V14), or constitutively active Galpha(12) induced endogenous RhoA activation inhibited the NGF-induced Rac1 activation without any effect on the NGF-induced extracellular signal-regulated kinase activation. Moreover, Y-27632, an inhibitor of Rho-associated kinase, completely abolished the RhoA-induced down-regulation of the NGF-induced Rac1 activation. We also revealed that NGF induced a rapid recruitment of Rac1 to the cell surface protrusion sites and formed filamentous actin-rich protrusions. Activation of RhoA and Rho-associated kinase formed a thick ringlike structure of cortical actin filaments at the cell periphery and then inhibited the NGF-induced recruitment of Rac1 to protrusions. These results indicate that RhoA down regulates the NGF- induced Rac1 activation through Rho-associated kinase, inhibiting the neurite formation. PMID- 11279040 TI - Generation of sequence-specific, high affinity anti-DNA antibodies. AB - By taking advantage of the extreme stability of a protein-DNA complex, we have obtained two highly specific monoclonal antibodies against a predetermined palindromic DNA sequence corresponding to the binding site of the E2 transcriptional regulator of the human papillomavirus (HPV-16). The purified univalent antibody fragments bind to a double-stranded DNA oligonucleotide corresponding to the E2 binding site in solution with dissociation constants in the low and subnanomolar range. This affinity matches that of the natural DNA binding domain and is severalfold higher than the affinity of a homologous bovine E2 C-terminal domain (BPV-1) for the same DNA. These antibodies discriminate effectively among a number of double- and single-stranded synthetic DNAs with factors ranging from 125- to 20,000-fold the dissociation constant of the specific DNA sequence used in the immunogenic protein-DNA complex. Moreover, they are capable of fine specificity tuning, since they both bind less tightly to another HPV-16 E2 binding site, differing in only 1 base pair in a noncontact flexible region. Beyond the relevance of obtaining a specific anti-DNA response, these results provide a first glance at how DNA as an antigen is recognized specifically by an antibody. The accuracy of the spectroscopic method used for the binding analysis suggests that a detailed mechanistic analysis is attainable. PMID- 11279041 TI - The chromatin structure of the dual c-myc promoter P1/P2 is regulated by separate elements. AB - The proto-oncogene c-myc is transcribed from a dual promoter P1/P2, with transcription initiation sites 160 base pairs apart. Here we have studied the transcriptional activation of both promoters on chromatin templates. c-myc chromatin was reconstituted on stably transfected, episomal, Epstein-Barr virus derived vectors in a B cell line. Episomal P1 and P2 promoters showed only basal activity but were strongly inducible by histone deacetylase inhibitors. The effect of promoter mutations on c-myc activity, chromatin structure, and E2F binding was studied. The ME1a1 binding site between P1 and P2 was required for the maintenance of an open chromatin configuration of the dual c-myc promoters. Mutation of this site strongly reduced the sensitivity of the core promoter region of P1/P2 to micrococcal nuclease and prevented binding of polymerase II (pol II) at the P2 promoter. In contrast, mutation of the P2 TATA box also abolished binding of pol II at the P2 promoter but did not affect the chromatin structure of the P1/P2 core promoter region. The E2F binding site adjacent to ME1a1 is required for repression of the P2 promoter but not the P1 promoter, likely by recruitment of histone deacetylase activity. Chromatin precipitation experiments with E2F-specific antibodies revealed binding of E2F-1, E2F-2, and E2F-4 to the E2F site of the c-myc promoter in vivo if the E2F site was intact. Taken together, the analyses support a model with a functional hierarchy for regulatory elements in the c-myc promoter region; binding of proteins to the ME1a1 site provides a nucleosome-free region of chromatin near the P2 start site, binding of E2F results in transcriptional repression without affecting polymerase recruitment, and the TATA box is required for polymerase recruitment. PMID- 11279042 TI - The yeast hsp70 homologue Ssa is required for translation and interacts with Sis1 and Pab1 on translating ribosomes. AB - The 70-kDa heat shock proteins are molecular chaperones that participate in a variety of cellular functions. This chaperone function is stimulated by interaction with hsp40 proteins. The Saccharomyces cerevisiae gene encoding the essential hsp40 homologue, SIS1, appears to function in translation initiation. Mutations in ribosomal protein L39 (rpl39) complement loss-of-function mutations in SIS1 as well as PAB1 (poly(A)-binding protein), suggesting a functional interaction between these proteins. However, while a direct interaction between Sis1 and Pab1 is not detectable, both of these proteins physically interact with the essential Ssa (and not Ssb) family of hsp70 proteins. This interaction is mediated by the variable C-terminal domain of Ssa. Subcellular fractionations demonstrate that the binding of Ssa to ribosomes is dependent upon its C terminus and that its interaction with Sis1 and Pab1 occurs preferentially on translating ribosomes. Consistent with a function in translation, depletion of Ssa protein produces a general translational defect that appears similar to loss of Sis1 and Pab1 function. This translational effect of Ssa appears mediated, at least in part, by its affect on the interaction of Pab1 with the translation initiation factor, eIF4G, which is dramatically reduced in the absence of functional Ssa protein. PMID- 11279043 TI - Aphidicolin triggers a block to replication origin firing in Xenopus egg extracts. AB - DNA replication origins are located at random with respect to DNA sequence in Xenopus early embryos and on DNA replicated in Xenopus egg extracts. We have recently shown that origins fire throughout the S phase in Xenopus egg extracts. To study the temporal regulation of origin firing, we have analyzed origin activation in sperm nuclei treated with the DNA polymerase inhibitor aphidicolin. Sperm chromatin was incubated in Xenopus egg extracts in the presence of aphidicolin and transferred to a fresh extract, and digoxigenin-dUTP and biotin dUTP were added at various times after aphidicolin release to selectively label early and late replicating DNA. Molecular combing analysis of single DNA fibers showed that only a fraction of potential origins were able to initiate in the presence of aphidicolin. After release from aphidicolin, the remaining origins fired asynchronously throughout the S phase. Therefore, initiation during the S phase depends on the normal progression of replication forks assembled at earlier activated origins. Caffeine, an inhibitor of the checkpoint kinases ATR and ATM, did not relieve the aphidicolin-induced block to origin firing. We conclude that a caffeine-insensitive intra-S phase checkpoint regulates origin activation when DNA synthesis is inhibited in Xenopus egg extracts. PMID- 11279044 TI - Contractile activity-induced transcriptional activation of cytochrome C involves Sp1 and is proportional to mitochondrial ATP synthesis in C2C12 muscle cells. AB - Contractile activity induces adaptations in the expression of genes encoding skeletal muscle mitochondrial proteins; however, the putative signals responsible for these adaptations remain unknown. We used electrical stimulation (5 Hz, 65 V) of C2C12 muscle cells in culture to define some of the mechanisms involved in contractile activity-induced changes in cytochrome c gene expression. Chronic contractile activity (4 days, 3 h/day) augmented cytochrome c mRNA by 1.6-fold above control cells. This was likely mediated by increases in transcriptional activation, because cells transfected with full-length (-726 base pairs) or minimal (-66 base pairs) cytochrome c promoter/chloramphenicol acetyltransferase reporter constructs demonstrated contractile activity-induced 1.5-1.7-fold increases in the absence of contractile activity-induced increases in mRNA stability. Transcriptional activation of the -726 promoter was abolished when muscle contraction was inhibited at various subcellular locations by pretreatment with either the Na(+) channel blocker tetrodotoxin, the intracellular Ca(2+) chelator 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl) ester, or the myosin ATPase inhibitor 2,3-butanedione monoxime. It was further reduced in unstimulated cells when mitochondrial ATP synthesis was impaired using the uncoupler 2,4-dinitrophenol. Because the contractile activity-induced response was evident within the minimal promoter, electromobility shift assays performed within the first intron (+75 to +104 base pairs) containing Sp1 sites revealed an elevated DNA binding in response to contractile activity. This was paralleled by increases in Sp1 protein levels. Sp1 overexpression studies also led to increases in cytochrome c transactivation and mRNA levels. These data suggest that variations in the rate of mitochondrial ATP synthesis are important in determining cytochrome c gene expression in muscle cells and that this is mediated, in part, by Sp1-induced increases in cytochrome c transcription. PMID- 11279045 TI - The effect of the erg26-1 mutation on the regulation of lipid metabolism in Saccharomyces cerevisiae. AB - A temperature-sensitive Saccharomyces cerevisiae mutant harboring a lesion in the ERG26 gene has been isolated. ERG26 encodes 4alpha-carboxysterol-C3 dehydrogenase, one of three enzymatic activities required for the conversion of 4,4-dimethylzymosterol to zymosterol. Gas chromatography/mass spectrometry analyses of sterols in this mutant, designated erg26-1, revealed the aberrant accumulation of a 4-methyl-4-carboxy zymosterol intermediate, as well as a novel 4-carboxysterol. Neutral lipid radiolabeling studies showed that erg26-1 cells also harbored defects in the rate of biosynthesis and steady-state levels of mono , di-, and triglycerides. Phospholipid radiolabeling studies showed defects in the rate of biosynthesis of both phosphatidic acid and phosphatidylinositol. Biochemical studies revealed that microsomes isolated from erg26-1 cells contained greatly reduced 4alpha-carboxysterol-C3 dehydrogenase activity when compared with microsomes from wild type cells. Previous studies have shown that loss of function mutations in either of the fatty acid elongase genes SUR4/ELO3 or FEN1/GNS1/ELO2 can "bypass" the essentiality of certain ERG genes (Ladeveze, V., Marcireau, C., Delourme, D., and Karst, F. (1993) Lipids 28, 907-912; Silve, S., Leplatois, P., Josse, A., Dupuy, P. H., Lanau, C., Kaghad, M., Dhers, C., Picard, C., Rahier, A., Taton, M., Le Fur, G., Caput, D., Ferrara, P., and Loison, G. (1996) Mol. Cell. Biol. 16, 2719-2727). Studies presented here have shown that this sphingolipid-dependent "bypass" mechanism did not suppress the essential requirement for zymosterol biosynthesis. However, studies aimed at understanding the underlying physiology behind the temperature-sensitive growth defect of erg26-1 cells showed that the addition of several antifungal compounds to the growth media of erg26-1 cells could suppress the temperature-sensitive growth defect. Fluorescence microscopic analysis showed that GFP-Erg26p and GFP Erg27p fusion proteins were localized to the endoplasmic reticulum. Two-hybrid analysis indicated that Erg25p, Erg26p, and Erg27p, which are required for the biosynthesis of zymosterol, form a complex within the cell. PMID- 11279046 TI - Inhibition of interactions and interconversions of prion protein isoforms by peptide fragments from the C-terminal folded domain. AB - The formation of protease-resistant prion protein (PrP-res or PrP(Sc)) involves selective interactions between PrP-res and its normal protease-sensitive counterpart, PrP-sen or PrP(C). Previous studies have shown that synthetic peptide fragments of the PrP sequence corresponding to residues 119-136 of hamster PrP (Ha119-136) can selectively block PrP-res formation in cell-free systems and scrapie-infected tissue culture cells. Here we show that two other peptides corresponding to residues 166-179 (Ha166-179) and 200-223 (Ha200-223) also potently inhibit the PrP-res induced cell-free conversion of PrP-sen to the protease-resistant state. In contrast, Ha121-141, Ha180-199, and Ha218-232 were much less effective as inhibitors. Mechanistic analyses indicated that Ha166-179, Ha200-223, and peptides containing residues 119-136 inhibit primarily by binding to PrP-sen and blocking its binding to PrP-res. Circular dichroism analyses indicated that Ha117-141 and Ha200-223, but not non-inhibitory peptides, readily formed high beta-sheet structures when placed under the conditions of the conversion reaction. We conclude that these inhibitory peptides may mimic contact surfaces between PrP-res and PrP-sen and thereby serve as models of potential therapeutic agents for transmissible spongiform encephalopathies. PMID- 11279047 TI - Rotation of a complex of the gamma subunit and c ring of Escherichia coli ATP synthase. The rotor and stator are interchangeable. AB - ATP synthase (F0F1) transforms an electrochemical proton gradient into chemical energy (ATP) through the rotation of a subunit assembly. It has been suggested that a complex of the gamma subunit and c ring (c(10-14)) of F0F1 could rotate together during ATP hydrolysis and synthesis (Sambongi, Y., Iko, Y., Tanabe, M., Omote, H., Iwamoto-Kihara, A., Ueda, I., Yanagida, T., Wada, Y., and Futai, M. (1999) Science 286, 1722-1724). We observed that the rotation of the c ring with the cI28T mutation (c subunit cIle-28 replaced by Thr) was less sensitive to venturicidin than that of the wild type, consistent with the antibiotic effect on the cI28T mutant and wild-type ATPase activities (Fillingame, R. H., Oldenburg, M., and Fraga, D. (1991) J. Biol. Chem. 266, 20934-20939). Furthermore, we engineered F0F1 to see the alpha(3)beta(3) hexamer rotation; a biotin tag was introduced into the alpha or beta subunit, and a His tag was introduced into the c subunit. The engineered enzymes could be purified by metal affinity chromatography and density gradient centrifugation. They were immobilized on a glass surface through the c subunit, and an actin filament was connected to the alpha or beta subunit. The filament rotated upon the addition of ATP and generated essentially the same frictional torque as one connected to the c ring. These results indicate that the gammaepsilonc(10-14) complex is a mechanical unit of the enzyme and that it can be used as a rotor or a stator experimentally, depending on the subunit immobilized. PMID- 11279048 TI - Membrane insertion of the heptameric staphylococcal alpha-toxin pore. A domino like structural transition that is allosterically modulated by the target cell membrane. AB - Staphylococcal alpha-toxin forms heptameric pores on eukaryotic cells. After binding to the cell membrane in its monomeric form, the toxin first assembles into a heptameric pre-pore. Subsequently, the pre-pore transforms into the final pore by membrane insertion of an amphipathic beta-barrel, which comprises the "central loop" domains of all heptamer subunits. The process of membrane insertion was analyzed here using a set of functionally altered toxin mutants. The results show that insertion may be initiated within an individual protomer when its NH2 terminus activates its central loop. The activated state is then shared with the central loops of the residual heptamer subunits, which results in cooperative membrane penetration. This cooperation of the central loops commences while these are still remote from the lipid bilayer. Nevertheless, it is subject to modulation by the target membrane, which therefore acts across a distance much like an allosteric effector. However, while allosteric transitions usually are reversible, membrane insertion of alpha-toxin is an irreversible event, and we show here that it can proceed to completion in a domino-like fashion when triggered by as little as a single foreign atom within the entire heptamer. PMID- 11279049 TI - Tumor necrosis factor-alpha inhibits aquaporin 5 expression in mouse lung epithelial cells. AB - Aquaporin 5 (AQP5), the major water channel expressed in alveolar, tracheal, and upper bronchial epithelium, is significantly down-regulated during pulmonary inflammation and edema. The mechanisms that underlie this decrease in AQP5 levels are therefore of considerable interest. Here we show that AQP5 expression in cultured lung epithelial cells is decreased 2-fold at the mRNA level and 10-fold at the protein level by the proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha). Treatment of murine lung epithelial cells (MLE-12) with TNF-alpha results in a concentration- and time-dependent decrease in AQP5 mRNA and protein expression. Activation of the p55 TNF-alpha receptor (TNFR1) with an agonist antibody is sufficient to cause decreased AQP5 expression, demonstrating that the TNF-alpha effect is mediated through TNFR1. Inhibition of nuclear factor kappaB (NF-kappaB) translocation to the nucleus blocks the effect of TNF-alpha on AQP5 expression, indicating that activation of NF-kappaB is required, whereas inhibition of extracellular signal-regulated or p38 mitogen-activated protein kinases showed no effect. These data show that TNF-alpha decreases AQP5 mRNA and protein expression and that the molecular pathway for this effect involves TNFR1 and activated NF-kappaB. The ability of inflammatory cytokines to decrease aquaporin expression may help explain the connection between inflammation and edema. PMID- 11279050 TI - Circular permutation of 5-aminolevulinate synthase. Mapping the polypeptide chain to its function. AB - 5-Aminolevulinate synthase is the first enzyme of the heme biosynthetic pathway in non-plant eukaryotes and some prokaryotes. The enzyme functions as a homodimer and requires pyridoxal 5'-phosphate as a cofactor. Although the roles of defined amino acids in the active site and catalytic mechanism have been recently explored using site-directed mutagenesis, much less is known about the role of the 5-aminolevulinate synthase polypeptide chain arrangement in folding, structure, and ultimately, function. To assess the importance of the continuity of the polypeptide chain, circularly permuted 5-aminolevulinate synthase variants were constructed through either rational design or screening of an engineered random library. One percent of the random library clones were active, and a total of 21 active variants had sequences different from that of the wild type 5 aminolevulinate synthase. Out of these 21 variants, 9 displayed unique circular permutations of the 5-aminolevulinate synthase polypeptide chain. The new termini of the active variants disrupted secondary structure elements and loop regions and fell in 100 amino acid regions from each terminus. This indicates that the natural continuity of the 5-aminolevulinate synthase polypeptide chain and the sequential arrangement of the secondary structure elements are not requirements for proper folding, binding of the cofactor, or assembly of the two subunits. Furthermore, the order of two identified functional elements (i.e. the catalytic and the glycine-binding domains) is apparently irrelevant for proper functioning of the enzyme. Although the wild type 5-aminolevulinate synthase and the circularly permuted variants appear to have similar, predicted overall tertiary structures, they exhibit differences in the arrangement of the secondary structure elements and in the cofactor-binding site environment. Taken together, the data lead us to propose that the 5-aminolevulinate synthase overall structure can be reached through multiple or alternative folding pathways. PMID- 11279051 TI - Structural organization and regulation of the small proline-rich family of cornified envelope precursors suggest a role in adaptive barrier function. AB - The protective barrier provided by stratified squamous epithelia relies on the cornified cell envelope (CE), a structure synthesized at late stages of keratinocyte differentiation. It is composed of structural proteins, including involucrin, loricrin, and the small proline-rich (SPRR) proteins, all encoded by genes localized at human chromosome 1q21. The genetic characterization of the SPRR locus reveals that the various members of this multigene family can be classified into two distinct groups with separate evolutionary histories. Whereas group 1 genes have diverged in protein structure and are composed of three different classes (SPRR1 (2x), SPRR3, and SPRR4), an active process of gene conversion has counteracted diversification of the protein sequences of group 2 genes (SPRR2 class, seven genes). Contrasting with this homogenization process, all individual members of the SPRR gene family show specific in vivo and in vitro expression patterns and react selectively to UV irradiation. Apparently, creation of regulatory rather than structural diversity has been the driving force behind the evolution of the SPRR gene family. Differential regulation of highly homologous genes underlines the importance of SPRR protein dosage in providing optimal barrier function to different epithelia, while allowing adaptation to diverse external insults. PMID- 11279052 TI - The electrophilic and leaving group phosphates in the catalytic mechanism of yeast pyrophosphatase. AB - Binding of pyrophosphate or two phosphate molecules to the pyrophosphatase (PPase) active site occurs at two subsites, P1 and P2. Mutations at P2 subsite residues (Y93F and K56R) caused a much greater decrease in phosphate binding affinity of yeast PPase in the presence of Mn(2+) or Co(2+) than mutations at P1 subsite residues (R78K and K193R). Phosphate binding was estimated in these experiments from the inhibition of ATP hydrolysis at a sub-K(m) concentration of ATP. Tight phosphate binding required four Mn(2+) ions/active site. These data identify P2 as the high affinity subsite and P1 as the low affinity subsite, the difference in the affinities being at least 250-fold. The time course of five "isotopomers" of phosphate that have from zero to four (18)O during [(18)O]P(i) [(16)O]H(2)O oxygen exchange indicated that the phosphate containing added water is released after the leaving group phosphate during pyrophosphate hydrolysis. These findings provide support for the structure-based mechanism in which pyrophosphate hydrolysis involves water attack on the phosphorus atom located at the P2 subsite of PPase. PMID- 11279053 TI - Myelin-associated glycoprotein interacts with ganglioside GT1b. A mechanism for neurite outgrowth inhibition. AB - Myelin-associated glycoprotein (MAG) is expressed on myelinating glia and inhibits neurite outgrowth from post-natal neurons. MAG has a sialic acid binding site in its N-terminal domain and binds to specific sialylated glycans and gangliosides present on the surface of neurons, but the significance of these interactions in the effect of MAG on neurite outgrowth is unclear. Here we present evidence to suggest that recognition of sialylated glycans is essential for inhibition of neurite outgrowth by MAG. Arginine 118 on MAG is known to make a key contact with sialic acid. We show that mutation of this residue reduces the potency of MAG inhibitory activity but that residual activity is also a result of carbohydrate recognition. We then go on to investigate gangliosides GT1b and GD1a as candidate MAG receptors. We show that MAG specifically binds both gangliosides and that both are expressed on the surface of MAG-responsive neurons. Furthermore, antibody cross-linking of cell surface GT1b, but not GD1a, mimics the effect of MAG, in that neurite outgrowth is inhibited through activation of Rho kinase. These data strongly suggest that interaction with GT1b on the neuronal cell surface is a potential mechanism for inhibition of neurite outgrowth by MAG. PMID- 11279054 TI - Systematic evolution of a DNA aptamer binding to rat brain tumor microvessels. selective targeting of endothelial regulatory protein pigpen. AB - Tumor microvessels differ in structure and metabolic function from normal vasculature, and neoangiogenesis is associated with quantitative and qualitative changes in expression of endothelial proteins. Such molecules could serve as molecular addresses differentiating the tumor vasculature from those of the normal brain. We have applied Systematic Evolution of Ligands by EXponential enrichment (SELEX) against transformed endothelial cells as a complex target to select single-stranded DNA-ligands (aptamers) that function as histological markers to detect microvessels of rat experimental glioma, a fatal brain tumor that is highly vascularized. Both the SELEX selection procedure as well as subsequent deconvolution-SELEX were analyzed by fluorescence based methods (flow cytometry and fluorescence microscopy). Of 25 aptamers analyzed, one aptamer was selected that selectively bound microvessels of rat brain glioblastoma but not the vasculature of the normal rat brain including peritumoral areas. The molecular target protein of aptamer III.1 was isolated from endothelial cells by ligand-mediated magnetic DNA affinity purification. This protein was identified by mass spectrometry as rat homologue of mouse pigpen, a not widely known endothelial protein the expression of which parallels the transition from quiescent to angiogenic phenotypes in vitro. Because neoangiogenesis, the formation of new blood vessels, is a key feature of tumor development, the presented aptamer can be used as a probe to analyze pathological angiogenesis of glioblastoma. The presented data show that pigpen is highly expressed in tumor microvessels of experimental rat brain glioblastoma and may play an important role in warranting blood supply, thus growth of brain tumors. PMID- 11279055 TI - A diverse family of proteins containing tumor necrosis factor receptor-associated factor domains. AB - We have identified three new tumor necrosis factor-receptor associated factor (TRAF) domain-containing proteins in humans using bioinformatics approaches, including: MUL, the product of the causative gene in Mulibrey Nanism syndrome; USP7 (HAUSP), an ubiquitin protease; and SPOP, a POZ domain-containing protein. Unlike classical TRAF family proteins involved in TNF family receptor (TNFR) signaling, the TRAF domains (TDs) of MUL, USP7, and SPOP are located near the NH(2) termini or central region of these proteins, rather than carboxyl end. MUL and USP7 are capable of binding in vitro via their TDs to all of the previously identified TRAF family proteins (TRAF1, TRAF2, TRAF3, TRAF4, TRAF5, and TRAF6), whereas the TD of SPOP interacts weakly with TRAF1 and TRAF6 only. The TD of MUL also interacted with itself, whereas the TDs of USP7 and SPOP did not self associate. Analysis of various MUL and USP7 mutants by transient transfection assays indicated that the TDs of these proteins are necessary and sufficient for suppressing NF-kappaB induction by TRAF2 and TRAF6 as well as certain TRAF binding TNF family receptors. In contrast, the TD of SPOP did not inhibit NF kappaB induction. Immunofluorescence confocal microscopy indicated that MUL localizes to cytosolic bodies, with targeting to these structures mediated by a RBCC tripartite domain within the MUL protein. USP7 localized predominantly to the nucleus, in a TD-dependent manner. Data base searches revealed multiple proteins containing TDs homologous to those found in MUL, USP7, and SPOP throughout eukaryotes, including yeast, protists, plants, invertebrates, and mammals, suggesting that this branch of the TD family arose from an ancient gene. We propose the moniker TEFs (TD-encompassing factors) for this large family of proteins. PMID- 11279056 TI - Starvation promotes nuclear accumulation of the hsp70 Ssa4p in yeast cells. AB - Nuclear import of proteins that are too large to passively enter the nucleus requires soluble factors, energy, and a nuclear localization signal (NLS). Nuclear protein transport can be regulated, and different forms of stress affect nucleocytoplasmic trafficking. As such, import of proteins containing a classical NLS is inhibited in starving yeast cells. In contrast, the hsp70 Ssa4p concentrates in nuclei upon starvation. Nuclear concentration of Ssa4p in starving cells is reversible, and transfer of stationary phase cells to fresh medium induces Ssa4p nuclear export. This export reaction represents an active process that is sensitive to oxidative stress. In starving cells, the N-terminal domain of Ssa4p mediates Ssa4p nuclear accumulation, and a short hydrophobic sequence, termed Star (for starvation), is sufficient to localize the reporter proteins green fluorescent protein or beta-galactosidase to nuclei. To determine whether nuclear accumulation of Star-beta-galactosidase depends on a specific nuclear carrier, we have analyzed its distribution in mutant yeast strains that carry a deletion of a single beta-importin gene. With this assay we have identified Nmd5p as a beta-importin required to concentrate Star-beta galactosidase in nuclei when cells enter stationary phase. PMID- 11279057 TI - Identification of a novel chloride channel expressed in the endoplasmic reticulum, golgi apparatus, and nucleus. AB - MID-1 is a Saccharomyces cerevisiae gene encoding a stretch-activated channel. Using MID-1 as a molecular probe, we isolated rat cDNA encoding a protein with four putative transmembrane domains. This gene encoded a protein of 541 amino acids. We also cloned the human homologue, which encoded 551 amino acids. Messenger RNA for this gene was expressed abundantly in the testis and moderately in the spleen, liver, kidney, heart, brain, and lung. In the testis, immunoreactivity of the gene product was detected both in the cytoplasm and the nucleus. When expressed in Chinese hamster ovary cells, the gene product was located in intracellular compartments including endoplasmic reticulum and the Golgi apparatus. When microsome fraction obtained from the transfected cells, but not from mock-transfected cells, was incorporated into the lipid bilayer, an anion channel activity was detected. Unitary conductance was 70 picosiemens in symmetric 150 mm KCl solution. We designated this gene Mid-1-related chloride channel (MCLC). MCLC encodes a new class of chloride channel expressed in intracellular compartments. PMID- 11279058 TI - Reduced expression of the epithelial adhesion ligand laminin 5 in the skin causes intradermal tissue separation. AB - Laminin 5, the major keratinocyte adhesion ligand, is found in the lamina lucida subregion of the epidermal basement membrane of the skin, where it colocalizes with the anchoring filaments. Mutations in the genes encoding laminin 5 cause junctional epidermolysis bullosa, an inherited skin blistering disease characterized by abnormal hemidesmosomes and cleavage of the lamina lucida leading to epidermal detachment. In this work we describe the genetic basis of a new subtype of lethal inherited epidermolysis bullosa associated with reduced skin reactivity to laminin 5, presence of mature hemidesmosomes, and intradermal cleavage of the skin. The epidermolysis bullosa patients were heterozygous for a nonsense mutation (Q896X) and a splice site mutation (764-10T-->G) in the gene (LAMC2) for the gamma2 chain of laminin 5. The nonsense mutation causes accelerated decay of the corresponding mRNA, while the splice site mutation results in maturation of a cryptic wild-type gamma2 mRNA leading to reduced expression of wild-type laminin 5. In vitro studies using the probands' keratinocytes showed that secretion of reduced amounts of functional laminin 5 in the patient, although permitting formation of hemidesmosomes, fail to restore efficient cell adhesion. Our results provide the first evidence that laminin 5 contributes to the firm adhesion of the epithelial basement membrane to the underlying stroma. They also show that a low expression level of laminin 5 induces assembly of mature hemidesmosomes in vivo but fails to assure a stable cohesion of the dermal-epidermal junction. PMID- 11279060 TI - Chimeras between single-stranded DNA-binding proteins from Escherichia coli and Mycobacterium tuberculosis reveal that their C-terminal domains interact with uracil DNA glycosylases. AB - Uracil, a promutagenic base in DNA can arise by spontaneous deamination of cytosine or incorporation of dUMP by DNA polymerase. Uracil is removed from DNA by uracil DNA glycosylase (UDG), the first enzyme in the uracil excision repair pathway. We recently reported that the Escherichia coli single-stranded DNA binding protein (SSB) facilitated uracil excision from certain structured substrates by E. coli UDG (EcoUDG) and suggested the existence of interaction between SSB and UDG. In this study, we have made use of the chimeric proteins obtained by fusion of N- and C-terminal domains of SSBs from E. coli and Mycobacterium tuberculosis to investigate interactions between SSBs and UDGs. The EcoSSB or a chimera containing its C-terminal domain interacts with EcoUDG in a binary (SSB-UDG) or a ternary (DNA-SSB-UDG) complex. However, the chimera containing the N-terminal domain from EcoSSB showed no interactions with EcoUDG. Thus, the C-terminal domain (48 amino acids) of EcoSSB is necessary and sufficient for interaction with EcoUDG. The data also suggest that the C-terminal domain (34 amino acids) of MtuSSB is a predominant determinant for mediating its interaction with MtuUDG. The mechanism of how the interactions between SSB and UDG could be important in uracil excision repair pathway has been discussed. PMID- 11279059 TI - New splicing-site mutations in the SURF1 gene in Leigh syndrome patients. AB - The gene SURF1 encodes a factor involved in the biogenesis of cytochrome c oxidase, the last complex in the respiratory chain. Mutations of the SURF1 gene result in Leigh syndrome and severe cytochrome c oxidase deficiency. Analysis of seven unrelated patients with cytochrome c oxidase deficiency and typical Leigh syndrome revealed different SURF1 mutations in four of them. Only these four cases had associated demyelinating neuropathy. Three mutations were novel splicing-site mutations that lead to the excision of exon 6. Two different novel heterozygous mutations were found at the same guanine residue at the donor splice site of intron 6; one was a deletion, whereas the other was a transition [588+1G>A]. The third novel splicing-site mutation was a homozygous [516-2_516 1delAG] in intron 5. One patient only had a homozygous polymorphism in the middle of the intron 8 [835+25C>T]. Western blot analysis showed that Surf1 protein was absent in all four patients harboring mutations. Our studies confirm that the SURF1 gene is an important nuclear gene involved in the cytochrome c oxidase deficiency. We also show that Surf1 protein is not implicated in the assembly of other respiratory chain complexes or the pyruvate dehydrogenase complex. PMID- 11279061 TI - Death domain mutagenesis of KILLER/DR5 reveals residues critical for apoptotic signaling. AB - The Fas/tumor necrosis factor (TNF)/TRAIL receptors signal death through a cytoplasmic death domain (DD) containing six alpha-helices with positively charged helix 2 interacting with negatively charged helix 3 of another DD. DD mutation occurs in head/neck and lung cancer (TRAIL receptor KILLER/DR5) and in lpr mice (Fas). We examined the apoptotic potential of known KILLER/DR5 lung tumor-derived mutants (n = 6) and DD mutants (n = 18) generated based on conservation with DR4, Fas, Fas-associated death domain (FADD), and tumor necrosis factor receptor 1 (TNFR1). With the exception of Arg-330 required in Fas or FADD for aggregation or for TNFR1 cytotoxicity, surprisingly major loss-of function KILLER/DR5 alleles (W325A, L334A (lpr-like), I339A, and W360A) contained hydrophobic residues. Loss-of-function of I339A (highly conserved) has not been reported in DDs. Charged residue mutagenesis revealed the following points. 1) E326A, conserved in DR4, is dispensable for death; the homologous residue is positively charged in Fas, TNFR1, and FADD and is critical for DD interactions. 2) K331A, D336A, E338A, K340A, K343A, and D351A have partial loss-of-function suggesting multiple charges stabilize receptor-adapter interactions. Analysis of the tumor-derived KILLER/DR5 mutants revealed the following. 1) L334F has partial loss-of-function versus L334A, whereas E338K has major loss-of-function versus E338A, examples where alanine and tumor-specific substitutions have divergent phenotypes. 2) Unexpectedly, S324F, E326K, K386N, and D407Y have no loss-of function with tumor-specific or alanine substitutions. Loss-of-function KILLER/DR5 mutants were deficient in recruitment of FADD and caspase 8 to TRAIL death-inducing signaling complexes. The results reveal determinants within KILLER/DR5 for death signaling and drug design. PMID- 11279062 TI - Residue Gly1326 of the N-type calcium channel alpha 1B subunit controls reversibility of omega-conotoxin GVIA and MVIIA block. AB - We recently reported that amino acid residues contained within a putative EF hand motif in the domain III S5-H5 region of the alpha(1B) subunit affected the relative barium:calcium permeability of N-type calcium channels (Feng, Z. P., Hamid, J., Doering, C., Jarvis, S. E., Bosey, G. M., Bourinet, E., Snutch, T. P., and Zamponi, G. W. (2001) J. Biol. Chem. 276, 5726-5730). Since this region partially overlaps with residues previously implicated in block of the channel by omega-conotoxin GVIA, we assessed the effects of mutations in the putative EF hand domain on channel block by omega-conotoxin GVIA and the structurally related omega-conotoxin MVIIA. Both of the toxins irreversibly block the activity of wild type alpha(1B) N-type channels. We find that in addition to previously identified amino acid residues, residues in positions 1326 and 1332 are important determinants of omega-conotoxin GVIA blockade. Substitution of residue Glu(1332) to arginine slows the time course of development of block. Point mutations in position Gly(1326) to either arginine, glutamic acid, or proline dramatically decrease the time constant for development of the block. Additionally, in the G1326P mutant channel activity was almost completely recovered following washout. A qualitatively similar result was obtained with omega-conotoxin MVIIA, suggesting that common molecular determinants underlie block by these two toxins. Taken together the data suggest that residue Gly(1326) may form a barrier, which controls the access of peptide toxins to their blocking site within the outer vestibule of the channel pore and also stabilizes the toxin-channel interaction. PMID- 11279063 TI - Defining the drug-binding site in the human multidrug resistance P-glycoprotein using a methanethiosulfonate analog of verapamil, MTS-verapamil. AB - Defining the residues involved in the binding of a substrate provides insight into how the human multidrug resistance P-glycoprotein (P-gp) can transport a wide range of structurally diverse compounds out of the cell. Because verapamil is the most potent stimulator of P-gp ATPase activity, we synthesized a thiol reactive analog of verapamil (MTS-verapamil) and used it with cysteine-scanning mutagenesis to identify the reactive residues within the drug-binding domain of P gp. MTS-verapamil stimulated the ATPase activity of Cys-less P-gp and had a K(m) value (25 microM) that was similar to that of verapamil. 252 P-gp mutants containing a single cysteine within the predicted transmembrane (TM) segments were expressed in HEK 293 cells and purified by nickel-chelate chromatography and assayed for inhibition by MTS-verapamil. The activities of 15 mutants, Y118C (TM2), V125C (TM2), S222C (TM4), L339C (TM6), A342C (TM6), A729C (TM7), A841C (TM9), N842C (TM9), I868C (TM10), A871C (TM10), F942C (TM11), T945C (TM11), V982C (TM12), G984C (TM12), and A985C (TM12), were inhibited by MTS-verapamil. Four mutants, S222C (TM4), L339C (TM6), A342C (TM6), and G984C (TM12), were significantly protected from inhibition by MTS-verapamil by pretreatment with verapamil. Less protection was observed in mutants I868C (TM10), F942C (TM11) and T945C (TM11). These results indicate that residues in TMs 4, 6, 10, 11, and 12 must contribute to the binding of verapamil. PMID- 11279064 TI - 14-3-3 Binding to Na+/H+ exchanger isoform-1 is associated with serum-dependent activation of Na+/H+ exchange. AB - Na(+)/H(+) exchanger isoform-1 (NHE1), the ubiquitous form of the Na(+)/H(+) exchanger, has increased activity in hypertensive patients and in animal models of hypertension. Furthermore, NHE1 is activated in cells stimulated with growth factors. We showed previously that activation of the exchanger is dependent on phosphorylation of serine 703 (Ser(P)(703)) by p90 ribosomal S6 kinase (RSK). Because the NHE1 sequence at Ser(P)(703) (RIGSDP) is similar to a consensus sequence (RSXSXP) specific for 14-3-3 ligands, we evaluated whether serum stimulated 14-3-3 binding to NHE1. Five different GST-NHE1 fusion proteins spanning amino acids 515-815 were phosphorylated by RSK and used as ligands in a far Western analysis; only those containing Ser(P)(703) exhibited high affinity 14-3-3 binding. In PS127A cells (NHE1-overexpressing Chinese hamster fibroblasts) stimulated with 20% serum, NHE1 co-precipitation with GST-14-3-3 fusion protein increased at 5 min (5.2 +/- 0.4-fold versus control; p < 0.01) and persisted at 40 min (3.9 +/- 0.3-fold; p < 0.01). We confirmed that binding occurs at the RIGSDP motif using PS120 (NHE1 null) cells transfected with S703A-NHE1 or P705A NHE1 (based on data indicating that 14-3-3 binding requires phosphoserine and +2 proline). Serum failed to stimulate association of 14-3-3 with these mutants. A GST-NHE1 fusion protein was phosphorylated by RSK and used as a ligand to assess the effect of 14-3-3 on protein phosphatase 1-mediated dephosphorylation of Ser(P)(703). GST-14-3-3 limited dephosphorylation (66% of initial state at 60 min) compared with GST alone (27% of initial state; p < 0.01). The protective effect of GST-14-3-3 was lost in the GST-NHE1 P705A mutant. Finally, the base line rate of pH recovery in acid-loaded cells was equal in unstimulated cells expressing wild-type or P705A-NHE1. However, activation of NHE1 by serum was dramatically inhibited in cells expressing P705A-NHE1 compared with wild-type (0.13 +/- 0.02 versus 0.48 +/- 0.06 mmol of H(+)/min/liter, p < 0.01). These data suggest that 14-3-3 binding to NHE1 participates in serum-stimulated exchanger activation, a new function for 14-3-3. PMID- 11279065 TI - p110beta and p110delta phosphatidylinositol 3-kinases up-regulate Fc(epsilon)RI activated Ca2+ influx by enhancing inositol 1,4,5-trisphosphate production. AB - Fc(epsilon)RI-induced Ca2+ signaling in mast cells is initiated by activation of cytosolic tyrosine kinases. Here, in vitro phospholipase assays establish that the phosphatidylinositol 3-kinase (PI 3-kinase) lipid product, phosphatidylinositol 3,4,5-triphosphate, further stimulates phospholipase Cgamma2 that has been activated by conformational changes associated with tyrosine phosphorylation or low pH. A microinjection approach is used to directly assess the consequences of inhibiting class IA PI 3-kinases on Ca2+ responses after Fc(epsilon)RI cross-linking in RBL-2H3 cells. Injection of antibodies to the p110beta or p110delta catalytic isoforms of PI 3-kinase, but not antibodies to p110alpha, lengthens the lag time to release of Ca2+ stores and blunts the sustained phase of the calcium response. Ca2+ responses are also inhibited in cells microinjected with recombinant inositol polyphosphate 5-phosphatase I, which degrades inositol 1,4,5-trisphosphate (Ins(1,4,5)P3), or heparin, a competitive inhibitor of the Ins(1,4,5)P3 receptor. This indicates a requirement for Ins(1,4,5)P3 to initiate and sustain Ca2+ responses even when PI 3-kinase is fully active. Antigen-induced cell ruffling, a calcium-independent event, is blocked by injection of p110beta and p110delta antibodies, but not by injection of 5-phosphatase I, heparin, or anti-p110alpha antibodies. These results suggest that the p110beta and p110delta isoforms of PI 3-kinase support Fc(epsilon)RI induced calcium signaling by modulating Ins(1,4,5)P3 production, not by directly regulating the Ca2+ influx channel. PMID- 11279066 TI - Domains of apolipoprotein E contributing to triglyceride and cholesterol homeostasis in vivo. Carboxyl-terminal region 203-299 promotes hepatic very low density lipoprotein-triglyceride secretion. AB - Apolipoprotein (apo) E has been implicated in cholesterol and triglyceride homeostasis in humans. At physiological concentration apoE promotes efficient clearance of apoE-containing lipoprotein remnants. However, high apoE plasma levels correlate with high plasma triglyceride levels. We have used adenovirus mediated gene transfer in apoE-deficient mice (E(-)/-) to define the domains of apoE required for cholesterol and triglyceride homeostasis in vivo. A dose of 2 x 10(9) plaque-forming units of apoE4-expressing adenovirus reduced slightly the cholesterol levels of E(-)/- mice and resulted in severe hypertriglyceridemia, due to accumulation of cholesterol and triglyceride-rich very low density lipoprotein particles in plasma. In contrast, the truncated form apoE4-202 resulted in a 90% reduction in the plasma cholesterol levels but did not alter plasma triglyceride levels in the E(-)/- mice. ApoE secretion by cell cultures, as well as the steady-state hepatic mRNA levels in individual mice expressing apoE4 or apoE4-202, were similar. In contrast, very low density lipoprotein triglyceride secretion in mice expressing apoE4, but not apoE4-202, was increased 10-fold, as compared with mice infected with a control adenovirus. The findings suggest that the amino-terminal 1-202 region of apoE4 contains the domains required for the in vivo clearance of lipoprotein remnants. Furthermore, the carboxyl-terminal 203-299 residues of apoE promote hepatic very low density lipoprotein-triglyceride secretion and contribute to apoE-induced hypertriglyceridemia. PMID- 11279067 TI - An extended conformation of the macrophage mannose receptor. AB - The macrophage mannose receptor mediates phagocytosis of pathogenic microorganisms and endocytosis of potentially harmful soluble glycoproteins by recognition of their defining carbohydrate structures. The mannose receptor is the prototype for a family of receptors each having an extracellular region consisting of 8-10 domains related to C-type carbohydrate recognition domains (CRDs), a fibronectin type II repeat and an N-terminal cysteine-rich domain. Hydrodynamic analysis and proteolysis experiments performed on fragments of the extracellular region of the receptor have been used to investigate its conformation. Size and shape parameters derived from sedimentation and diffusion coefficients indicate that the receptor is a monomeric, elongated and asymmetric molecule. Proteolysis experiments indicate the presence of close contacts between several pairs of domains and exposed linker regions separating CRDs 3 and 6 from their neighboring domains. Hydrodynamic coefficients predicted for modeled receptor conformations are consistent with an extended conformation with close contacts between three pairs of CRDs. The N-terminal cysteine-rich domain and the fibronectin type II repeat appear to increase the rigidity of the molecule. The rigid, extended conformation of the receptor places domains with different functions at distinct positions with respect to the membrane. PMID- 11279068 TI - Oxidative protein cross-linking reactions involving L-tyrosine in transforming growth factor-beta1-stimulated fibroblasts. AB - The mechanisms by which ligand-stimulated generation of reactive oxygen species in nonphagocytic cells mediate biologic effects are largely unknown. The profibrotic cytokine, transforming growth factor-beta1 (TGF-beta1), generates extracellular hydrogen peroxide (H2O2) in contrast to intracellular reactive oxygen species production by certain mitogenic growth factors in human lung fibroblasts. To determine whether tyrosine residues in fibroblast-derived extracellular matrix (ECM) proteins may be targets of H2O2-mediated dityrosine dependent cross-linking reactions in response to TGF-beta1, we utilized fluorophore-labeled tyramide, a structurally related phenolic compound that forms dimers with tyrosine, as a probe to detect such reactions under dynamic cell culture conditions. With this approach, a distinct pattern of fluorescent labeling that seems to target ECM proteins preferentially was observed in TGF beta1-treated cells but not in control cells. This reaction required the presence of a heme peroxidase and was inhibited by catalase or diphenyliodonium (a flavoenzyme inhibitor), similar to the effect on TGF-beta1-induced dityrosine formation. Exogenous addition of H2O2 to control cells that do not release extracellular H2O2 produced a similar fluorescent labeling reaction. These results support the concept that, in the presence of heme peroxidases in vivo, TGF-beta1-induced H2O2 production by fibroblasts may mediate oxidative dityrosine dependent cross-linking of ECM protein(s). This effect may be important in the pathogenesis of human fibrotic diseases characterized by overexpression/activation of TGF-beta1. PMID- 11279069 TI - Structural compensation for the deficit of rRNA with proteins in the mammalian mitochondrial ribosome. Systematic analysis of protein components of the large ribosomal subunit from mammalian mitochondria. AB - The mammalian mitochondrial ribosome (mitoribosome) is a highly protein-rich particle in which almost half of the rRNA contained in the bacterial ribosome is replaced with proteins. It is known that mitochondrial translation factors can function on both mitochondrial and Escherichia coli ribosomes, indicating that protein components in the mitoribosome compensate the reduced rRNA chain to make a bacteria-type ribosome. To elucidate the molecular basis of this compensation, we analyzed bovine mitoribosomal large subunit proteins; 31 proteins were identified including 15 newly identified proteins with their cDNA sequences from human and mouse. The results showed that the proteins with binding sites on rRNA shortened or lost in the mitoribosome were enlarged when compared with the E. coli counterparts; this suggests the structural compensation of the rRNA deficit by the enlarged proteins in the mitoribosome. PMID- 11279070 TI - Molecular patterning of the oikoplastic epithelium of the larvacean tunicate Oikopleura dioica. AB - Appendicularia are protochordates that rely on a complex mucous secretion, the house, to filter food particles from seawater. A monolayer of cells covering the trunk of the animal, the oikoplastic epithelium, secretes the house. This epithelium contains a fixed number of cells arranged in characteristic patterns with distinct sizes and nuclear morphologies. Certain house structures appear to be spatially related to defined, underlying groups of cells in the epithelium. We show that the house is composed of at least 20 polypeptides, a number of which are highly glycosylated, with glycosidase treatments resulting in molecular mass shifts exceeding 100 kDa. Nanoelectrospray tandem mass spectrometric microsequencing of house polypeptides was used to design oligonucleotides to screen an adult Oikopleura dioica cDNA library. This resulted in the isolation of cDNAs coding for three different proteins, oikosin 1, oikosin 2, and oikosin 3. The latter two are novel proteins unrelated to any known data base entries. Oikosin 1 has 13 repeats of a Cys domain, previously identified as a subunit of repeating sequences in some vertebrate mucins. We also find one repeat of this Cys domain in human cartilage intermediate layer protein but find no evidence of this domain in any invertebrate species, including those for which entire genomes have been sequenced. The three oikosins show distinct and complementary expression patterns restricted to the oikoplastic epithelium. This easily accessible epithelium, with differential gene expression patterns in readily identifiable groups of cells with distinctive nuclear morphologies, is a highly attractive model system for molecular studies of pattern formation. PMID- 11279071 TI - Activation of Galpha s mediates induction of tissue-type plasminogen activator gene transcription by epoxyeicosatrienoic acids. AB - The epoxyeicosatrienoic acids (EETs) are products of cytochrome P450 (CYP) epoxygenases that have vasodilatory and anti-inflammatory properties. Here we report that EETs have additional fibrinolytic properties. In vascular endothelial cells, physiological concentrations of EETs, particularly 11,12-EET, or overexpression of the endothelial epoxygenase, CYP2J2, increased tissue plasminogen activator (t-PA) expression by 2.5-fold without affecting plasminogen activator inhibitor-1 expression. This increase in t-PA expression correlated with a 4-fold induction in t-PA gene transcription and a 3-fold increase in t-PA fibrinolytic activity and was blocked by the CYP inhibitor, SKF525A, but not by the calcium-activated potassium channel blocker, charybdotoxin, indicating a mechanism that does not involve endothelial cell hyperpolarization. The t-PA promoter is cAMP-responsive, and induction of t-PA gene transcription by EETs correlated with increases in intracellular cAMP levels and, functionally, with cAMP-driven promoter activity. To determine whether increases in intracellular cAMP levels were due to modulation of guanine nucleotide-binding proteins, we assessed the effects of EETs on Galpha(s) and Galpha(i2). Treatment with EETs increased Galpha(s), but not Galpha(i2), GTP-binding activity by 3.5-fold. These findings indicate that EETs possess fibrinolytic properties through the induction of t-PA and suggest that endothelial CYP2J2 may play an important role in regulating vascular hemostasis. PMID- 11279072 TI - Characterization of an activation protein-1-binding site in the murine interleukin-12 p40 promoter. Demonstration of novel functional elements by a reductionist approach. AB - Interleukin (IL)-12 is a heterodimeric cytokine produced by macrophages in response to intracellular pathogens and provides an obligatory signal for the differentiation of T-helper-1 cells. We previously reported an analysis of the IL 12 p40 promoter in RAW264.7 macrophages. Multiple control elements were involved in activation of transcription by bacterial products. A critical control element, located between -96 and -88, interacts with C/EBP family members. In this study, using a strategy to demonstrate functional activity in a minimal promoter context, three novel cis-acting elements are found to have an important role in IL-12 p40 promoter activation by lipopolysaccharide. One of these elements is characterized in detail. Mutations from -79 to -74 in the murine IL-12 p40 promoter significantly reduce lipopolysaccharide-induced promoter activity. Electrophoretic mobility shift assays demonstrate binding of AP-1 family members to this region. Spacing between the C/EBP and AP-1 site is important for promoter activation, suggesting cooperativity between these elements. c-Jun and a mutant c Jun molecule activate the IL-12 p40 promoter and synergistically activate the promoter when co-expressed with C/EBPbeta. Finally, this region of the promoter is demonstrated to be a target for mitogen-activated protein kinase and toll-like receptor signaling pathways. PMID- 11279073 TI - Calcineurin enhances MAPK phosphatase-1 expression and p38 MAPK inactivation in cardiac myocytes. AB - Multiple intracellular signaling pathways have been shown to regulate the hypertrophic growth of cardiac myocytes including mitogen-activated protein kinase (MAPK) and calcineurin-nuclear factor of activated T-cells. However, it is uncertain if individual regulatory pathways operate in isolation or if interconnectivity between unrelated pathways is required for the orchestration of the entire hypertrophic response. To this end, we investigated the interconnectivity between calcineurin-mediated cardiac myocyte hypertrophy and p38 MAPK signaling in vitro and in vivo. We show that calcineurin promotes down regulation of p38 MAPK activity and enhances expression of the dual specificity phosphatase MAPK phosphatase-1 (MKP-1). Transgenic mice expressing activated calcineurin in the heart were characterized by inactivation of p38 and increased MKP-1 expression during early postnatal development, before the onset of cardiac hypertrophy. In vitro, cultured neonatal cardiomyocytes infected with a calcineurin-expressing adenovirus and stimulated with phenylephrine demonstrated reduced p38 phosphorylation and increased MKP-1 protein levels. Activation of endogenous calcineurin with the calcium ionophore decreased p38 phosphorylation and increased MKP-1 protein levels. Inhibition of endogenous calcineurin with cyclosporin A decreased MKP-1 protein levels and increased p38 activation in response to agonist stimulation. To further investigate potential cross-talk between calcineurin and p38 through alteration in MKP-1 expression, the MKP-1 promoter was characterized and determined to be calcineurin-responsive. These data suggest that calcineurin enhances MKP-1 expression in cardiac myocytes, which is associated with p38 inactivation. PMID- 11279075 TI - Effects of genetic background on thermoregulation and fatty acid-induced uncoupling of mitochondria in UCP1-deficient mice. AB - An interaction between free fatty acids and UCP1 (uncoupling protein-1) leading to de-energization of mitochondria was assumed to be a key event for triggering heat production in brown fat. Recently, Matthias et al., finding indistinguishable de-energization of isolated brown fat mitochondria by fatty acids in UCP1-deficient mice and control mice, challenged this assumption (Matthias, A., Jacobsson, A., Cannon, B., and Nedergaard, J. (1999) J. Biol. Chem. 274, 28150-28160). Since their results were obtained using UCP1-deficient and control mice on an undefined genetic background, we wanted to determine unambiguously the phenotype of UCP1 deficiency with the targeted Ucp1 allele on congenic C57BL/6J and 129/SvImJ backgrounds. UCP1-deficient congenic mice have a very pronounced cold-sensitive phenotype; however, deficient mice on the F1 hybrid background were resistant to cold. We propose that heterosis provides a mechanism to compensate for UCP1 deficiency. Contrary to the results of Matthias et al., we found a significant loss of fatty acid-induced de-energization, as reflected by membrane potential and oxygen consumption, in brown fat mitochondria from UCP1-deficient mice. Unlike cold sensitivity, fatty acid-induced uncoupling of mitochondria was independent of the genetic background of UCP1-deficient mice. We propose that intracellular free fatty acids directly regulate uncoupling activity of UCP1 in a manner consistent with models described in the literature. PMID- 11279074 TI - Mutations in sialidosis impair sialidase binding to the lysosomal multienzyme complex. AB - Sialidosis is an autosomal recessive disease caused by the genetic deficiency of lysosomal sialidase, which catalyzes the catabolism of sialoglycoconjugates. The disease is associated with progressive impaired vision, macular cherry-red spots, and myoclonus (sialidosis type I) or with skeletal dysplasia, Hurler-like phenotype, dysostosis multiplex, mental retardation, and hepatosplenomegaly (sialidosis type II). We analyzed the effect of the missense mutations G68V, S182G, G227R, F260Y, L270F, A298V, G328S, and L363P, which are identified in the sialidosis type I and sialidosis type II patients, on the activity, stability, and intracellular distribution of sialidase. We found that three mutations, F260Y, L270F, and A298V, which are clustered in the same region on the surface of the sialidase molecule, dramatically reduced the enzyme activity and caused a rapid intralysosomal degradation of the expressed protein. We suggested that this region might be involved in sialidase binding with lysosomal cathepsin A and/or beta-galactosidase in the multienzyme lysosomal complex required for the expression of sialidase activity. Transgenic expression of mutants followed by density gradient centrifugation of cellular extracts confirmed this hypothesis and showed that sialidase deficiency in some sialidosis patients results from disruption of the lysosomal multienzyme complex. PMID- 11279076 TI - ADP-dependent DNA strand exchange by the mutant [P67G/E68A] RecA protein. AB - We have prepared a mutant RecA protein in which proline 67 and glutamic acid 68 in the NTP binding site were replaced by a glycine and alanine residue, respectively. The [P67G/E68A]RecA protein catalyzes the single-stranded DNA dependent hydrolysis of ATP and is able to promote the standard ATP-dependent three-strand exchange reaction between a circular bacteriophage phiX174 (phiX) single-stranded DNA molecule and a homologous linear phiX double-stranded (ds) DNA molecule (5.4 kilobase pairs). The strand exchange activity differs from that of the wild type RecA protein, however, in that it is (i) completely inhibited by an ATP regeneration system, and (ii) strongly stimulated by the addition of high concentrations of ADP to the reaction solution. These results indicate that the strand exchange activity of the [P67G/E68A]RecA protein is dependent on the presence of both ATP and ADP. The ADP dependence of the reaction is reduced or eliminated when (i) a shorter linear phiX dsDNA fragment (1.1 kilobase pairs) is substituted for the full-length linear phiX dsDNA substrate, or (ii) the Mg(2+) concentration is reduced to a level just sufficient to complex the ATP present in the reaction solution. These results indicate that it is the branch migration phase (and not the initial pairing step) of the [P67G/E68A]RecA protein-promoted strand exchange reaction that is dependent on ADP. It is likely that the [P67G/E68A]RecA mutation has revealed a requirement for ADP that also exists (but is not as readily apparent) in the strand exchange reaction of the wild type RecA protein. PMID- 11279078 TI - Chromatin is permissive to TATA-binding protein (TBP)-mediated transcription initiation. AB - Preinitiation complex assembly is nucleated by the binding of TFIID to the promoters of protein coding genes transcribed by RNA polymerase II. TFIID is comprised of the TATA-binding protein (TBP) and TBP-associated factors (TAF(II)s). We investigated the transcription properties of TBP and TFIID on chromatin templates. On naked templates both TBP and purified TFIID are able to initiate basal transcription. However, on chromatin templates only TBP mediates transcription initiation in a heat-treated extract, whereas TFIID does not. Moreover, TBP-mediated chromatin transcription is blocked in a nontreated extract. These observations suggest that a chromatin-targeted repressor is present in crude extracts and that chromatin per se is not refractory to transcription mediated by TBP. As TBP can function through TAF(II)-independent and TAF(II)-dependent pathways, the repression of TBP-mediated basal transcription may be an additional level to the control of Pol II transcription initiation on chromatin. PMID- 11279079 TI - p53 Latency. C-terminal domain prevents binding of p53 core to target but not to nonspecific DNA sequences. AB - The p53 transcription factor is either latent or activated through multi-site phosphorylation and acetylation of the negative regulatory region in its C terminal domain (CTD). How CTD modifications activate p53 binding to target DNA sequences via its core domain is still unknown. It has been proposed that nonmodified CTD interacts either with the core domain or with DNA preventing binding of the core domain to DNA and that the fragments of the CTD regulatory region activate p53 by interfering with these interactions. We here characterized the sequence and target specificity of p53 activation by CTD fragments, interaction of activating peptides with p53 and target DNA, and interactions of "latent" p53 with DNA by a band shift assay and by fluorescence correlation spectroscopy. In addition to CTD fragments, several long basic peptides activated p53 and also transcription factor YY1. These peptides and CTD aggregated target DNA but apparently did not interact with p53. The potency to aggregate DNA correlated with the ability to activate p53, suggesting that p53 binds to target sequences upon interactions with tightly packed DNA in aggregates. Latent full length p53 dissociated DNA aggregates via its core and CTD, and this effect was potentiated by GTP. Latent p53 also formed complexes via both its core and CTD with long nontarget DNA molecules. Such p53-DNA interactions may occur if latent p53 binding to DNA via CTD prevents the interaction of the core domain with target DNA sites but not with nonspecific DNA sequences. PMID- 11279080 TI - Erbin is a protein concentrated at postsynaptic membranes that interacts with PSD 95. AB - Neuregulin is a factor essential for synapse-specific transcription of acetylcholine receptor genes at the neuromuscular junction. Its receptors, ErbB receptor tyrosine kinases, are localized at the postjunctional membrane presumably to ensure localized signaling. However, the molecular mechanisms underlying synaptic localization of ErbBs are unknown. Our recent studies indicate that ErbB4 interacts with postsynaptic density (PSD)-95 (SAP90), a PDZ domain-containing protein that does not interact with ErbB2 or ErbB3. Using as bait the ErbB2 C terminus, we identified Erbin, another PDZ domain-containing protein that interacts specifically with ErbB2. Erbin is concentrated in postsynaptic membranes at the neuromuscular junction and in the central nervous system, where ErbB2 is concentrated. Expression of Erbin increases the amount of ErbB2 labeled by biotin in transfected cells, suggesting that Erbin is able to increase ErbB2 surface expression. Furthermore, we provide evidence that Erbin interacts with PSD-95 in both transfected cells and synaptosomes. Thus ErbB proteins can interact with a network of PDZ domain-containing proteins. This interaction may play an important role in regulation of neuregulin signaling and/or subcellular localization of ErbB proteins. PMID- 11279081 TI - Chemical synthesis and biological activity of bromohydrin pyrophosphate, a potent stimulator of human gamma delta T cells. AB - Small phosphorylated metabolites from mycobacteria stimulate human gammadelta T lymphocytes. Although such phosphoantigens could prove useful in the composition of vaccines involving gammadelta T cell-mediated immunity, their very low abundance in natural sources limits such applications. Here, we describe the chemical production, purification, and bioactivity of a phosphorylated bromohydrin (BrHPP) analogue that mimics the biological properties of natural phosphoantigens. This compound can be obtained in gram amounts, is easy to detect, and is of high stability in aqueous solutions. Whereas unspecific binding of BrHPP to a wide panel of cell surface receptors is not detected even at micromolar concentrations, nanomolar concentrations specifically trigger effector responses of human gammadelta T lymphocytes. Thus, BrHPP is a novel molecule enabling potent immunostimulation of human gammadelta T lymphocytes. PMID- 11279082 TI - Effects of histone acetylation on the solubility and folding of the chromatin fiber. AB - The folding ability of chromatin fractions containing approximately identical nucleosome numbers and the same linker histone composition, but with different extents of core histone acetylation, were analyzed by analytical ultracentrifugation. It was found that the acetylated fractions consistently exhibited a relatively small but significantly lower extent of compaction than that of their native nonacetylated counterparts. This was regardless of the extent of the size distribution heterogeneity of the fractions analyzed. Furthermore the acetylated chromatin fibers exhibited an enhanced solubility in both NaCl and MgCl(2), which is neither the result of a differential binding affinity of the linker histones to chromatin nor of an alteration in the relative amounts of the histone H1 variants. PMID- 11279084 TI - Biophysical characterization of the DNA binding domain of gpNu1, a viral DNA packaging protein. AB - Terminase enzymes are common to double-stranded DNA viruses. These enzymes "package" the viral genome into a pre-formed capsid. Terminase from bacteriophage lambda is composed of gpA (72.4 kDa) and gpNu1 (20.4 kDa) subunits. We have described the expression and biochemical characterization of gpNu1DeltaK100, a construct comprising the N-terminal 100 amino acids of gpNu1 (Yang, Q., de Beer, T., Woods, L., Meyer, J., Manning, M., Overduin, M., and Catalano, C. E. (1999) Biochemistry 38, 465-477). Here we present a biophysical characterization of this construct. Thermally induced loss of secondary and tertiary structures is fully reversible. Surprisingly, although loss of tertiary structure is cooperative, loss of secondary structure is non-cooperative. NMR and limited proteolysis data suggest that approximately 30 amino acids of gpNu1DeltaK100 are solvent-exposed and highly flexible. We therefore constructed gpNu1DeltaE68, a protein consisting of the N-terminal 68 residues of gpNu1. gpNu1DeltaE68 is a dimer with no evidence of dissociation or further aggregation. Thermally induced unfolding of gpNu1DeltaE68 is reversible, with concomitant loss of both secondary and tertiary structure. The melting temperature increases with increasing protein concentration, suggesting that dimerization and folding are, at least in part, coupled. The data suggest that gpNu1DeltaE68 represents the minimal DNA binding domain of gpNu1. We further suggest that the C-terminal approximately 30 residues in gpNu1DeltaK100 adopt a pseudo-stable alpha-helix that extends from the folded core of the protein. A model describing the role of this helix in the assembly of the packaging apparatus is discussed. PMID- 11279083 TI - Differential interactions of nucleotides at the two nucleotide binding domains of the cystic fibrosis transmembrane conductance regulator. AB - After phosphorylation by protein kinase A, gating of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel is regulated by the interaction of ATP with its nucleotide binding domains (NBDs). Models of this gating regulation have proposed that ATP hydrolysis at NBD1 and NBD2 may drive channel opening and closing, respectively (reviewed in Nagel, G. (1999) Biochim. Biophys. Acta 1461, 263-274). However, as yet there has been little biochemical confirmation of the predictions of these models. We have employed photoaffinity labeling with 8-azido-ATP, which supports channel gating as effectively as ATP to evaluate interactions with each NBD in intact membrane-bound CFTR. Mutagenesis of Walker A lysine residues crucial for azido-ATP hydrolysis to generate the azido ADP that is trapped by vanadate indicated a greater role of NBD1 than NBD2. Separation of the domains by limited trypsin digestion and enrichment by immunoprecipitation confirmed greater and more stable nucleotide trapping at NBD1. This asymmetry of the two domains in interactions with nucleotides was reflected most emphatically in the response to the nonhydrolyzable ATP analogue, 5'-adenylyl-beta,gamma-imidodiphosphate (AMP-PNP), which in the gating models was proposed to bind with high affinity to NBD2 causing inhibition of ATP hydrolysis there postulated to drive channel closing. Instead we found a strong competitive inhibition of nucleotide hydrolysis and trapping at NBD1 and a simultaneous enhancement at NBD2. This argues strongly that AMP-PNP does not inhibit ATP hydrolysis at NBD2 and thereby questions the relevance of hydrolysis at that domain to channel closing. PMID- 11279085 TI - Activation of Na+/H+ exchanger-directed protein kinases in the ischemic and ischemic-reperfused rat myocardium. AB - The activity of the Na(+)/H(+) exchanger has been implicated as an important contributing factor in damage to the myocardium that occurs during ischemia and reperfusion. We examined regulation of the protein in ischemic and reperfused isolated hearts and isolated ventricular myocytes. In isolated myocytes, extracellular signal-regulated kinases were important in regulating activity of the exchanger after recovery from ischemia. Ischemia followed by reperfusion caused a strong inhibitory effect on NHE1 activity that abated with continued reperfusion. Four major protein kinases of size 90, 55, 44, and 40 kDa phosphorylated the Na(+)/H(+) exchanger. The Na(+)/H(+) exchanger-directed kinases demonstrated dramatic increases in activity of 2-10-fold that was induced by 3 different models of ischemia and reperfusion in intact hearts and isolated myocytes. p90(rsk) was identified as the 90-kDa protein kinase activated by ischemia and reperfusion while ERK1/2 was identified as accounting for some of the 44-kDa protein kinase phosphorylating the Na(+)/H(+) exchanger. The results demonstrate that MAPK-dependent pathways including p90(rsk) and ERK1/2 and are important in regulating the Na(+)/H(+) exchanger and show their dramatic increase in activity toward the Na(+)/H(+) exchanger during ischemia and reperfusion of the myocardium. They also show that ischemia followed by reperfusion have important inhibitory effects on Na(+)/H(+) exchanger activity. PMID- 11279086 TI - Identification, characterization, and intracellular processing of ADAM-TS12, a novel human disintegrin with a complex structural organization involving multiple thrombospondin-1 repeats. AB - We have identified and cloned a human fetal lung cDNA encoding a new protein of the ADAM-TS family (a disintegrin and metalloproteinase domain, with thrombospondin type-1 modules) that has been called ADAM-TS12. This protein exhibits a domain organization similar to the remaining family members including a propeptide and metalloproteinase-like, disintegrin-like, and cysteine-rich domains. However, the number and organization of the TS repeats is unique with respect to other human ADAM-TSs. A total of eight TS-1 repeats arranged in three groups are present in this novel ADAM-TS. Analysis of intracellular processing of ADAM-TS12 revealed that it is synthesized as a precursor molecule that is first activated by cleavage of the prodomain in a furin-mediated process and subsequently processed into two fragments of different size: a 120-kDa N-terminal proteolytically active fragment containing the metalloproteinase and disintegrin domains, and a 83-kDa C-terminal fragment containing most of the TS-1 repeats. Somatic cell hybrid and radiation hybrid mapping experiments showed that the human ADAM-TS12 gene maps to 5q35, a location that differs from all ADAM genes mapped to date. Northern blot analysis of RNAs from human adult and fetal tissues demonstrated that ADAM-TS12 transcripts are only detected at significant levels in fetal lung but not in any other analyzed tissues. In addition, ADAM-TS12 transcripts were detected in gastric carcinomas and in tumor cell lines from diverse sources, being induced by transforming growth factor-beta in KMST human fibroblasts. These data suggest that ADAM-TS12 may play roles in pulmonary cells during fetal development or in tumor processes through its proteolytic activity or as a molecule potentially involved in regulation of cell adhesion. PMID- 11279087 TI - Porcine carbonyl reductase. structural basis for a functional monomer in short chain dehydrogenases/reductases. AB - Porcine testicular carbonyl reductase (PTCR) belongs to the short chain dehydrogenases/reductases (SDR) superfamily and catalyzes the NADPH-dependent reduction of ketones on steroids and prostaglandins. The enzyme shares nearly 85% sequence identity with the NADPH-dependent human 15-hydroxyprostaglandin dehydrogenase/carbonyl reductase. The tertiary structure of the enzyme at 2.3 A reveals a fold characteristic of the SDR superfamily that uses a Tyr-Lys-Ser triad as catalytic residues, but exhibits neither the functional homotetramer nor the homodimer that distinguish all SDRs. It is the first known monomeric structure in the SDR superfamily. In PTCR, which is also active as a monomer, a 41-residue insertion immediately before the catalytic Tyr describes an all-helix subdomain that packs against interfacial helices, eliminating the four-helix bundle interface conserved in the superfamily. An additional anti-parallel strand in the PTCR structure also blocks the other strand-mediated interface. These novel structural features provide the basis for the scaffolding of one catalytic site within a single molecule of the enzyme. PMID- 11279089 TI - Na+-Ca2+ exchanger remodeling in pressure overload cardiac hypertrophy. AB - Perturbations of Ca(2+) metabolism are central to the pathogenesis of cardiac hypertrophy. The electrogenic Na(+)-Ca(2+) exchanger mediates a substantial component of transmembrane Ca(2+) movement in cardiac myocytes and is up regulated in heart failure. However, the role of the exchanger in the pathogenesis of cardiac hypertrophy is poorly understood. Thoracic aortic banding in mice induced 50-60% increases in heart mass and cardiomyocyte size. Despite the absence of myocardial dysfunction, steady-state NCX1 transcript and protein levels were increased to an extent similar to that reported in heart failure. As recent studies indicate that calcineurin is critical to the expression of Na(+) Ca(2+) exchanger genes, we inhibited calcineurin with cyclosporin. Calcineurin inhibition blunted the increases in NCX1 transcript and protein levels and eliminated the increases in heart mass and cell volume normally associated with pressure overload. To examine the functional significance of these changes, we measured Na(+)-Ca(2+) exchanger current in two independent ways. Surprisingly, exchanger current density was decreased in hypertrophied myocytes, and this down regulation was eliminated by calcineurin inhibition. Together, these data reveal a role for Na(+)-Ca(2+) exchanger current in the electrical remodeling of hypertrophy and implicate calcineurin signaling therein. In addition, these data suggest the Na(+)-Ca(2+) exchanger is functionally regulated in hypertrophy. PMID- 11279088 TI - Molecular mechanism of aminoglycoside antibiotic kinase APH(3')-IIIa: roles of conserved active site residues. AB - The aminoglycoside antibiotic kinases (APHs) constitute a clinically important group of antibiotic resistance enzymes. APHs share structural and functional homology with Ser/Thr and Tyr kinases, yet only five amino acids are invariant between the two groups of enzymes and these residues are all located within the nucleotide binding regions of the proteins. We have performed site-directed mutagenesis on all five conserved residues in the aminoglycoside kinase APH(3') IIIa: Lys(44) and Glu(60) involved in ATP capture, a putative active site base required for deprotonating the incoming aminoglycoside hydroxyl group Asp(190), and the Mg(2+) ligands Asn(195) and Glu(208), which coordinate two Mg(2+) ions, Mg1 and Mg2. Previous structural and mutagenesis evidence have demonstrated that Lys(44) interacts directly with the phosphate groups of ATP; mutagenesis of invariant Glu(60), which forms a salt bridge with the epsilon-amino group of Lys(44), demonstrated that this residue does not play a critical role in ATP recognition or catalysis. Results of mutagenesis of Asp(190) were consistent with a role in proper positioning of the aminoglycoside hydroxyl during phosphoryl transfer but not as a general base. The Mg1 and Mg2 ligand Asp(208) was found to be absolutely required for enzyme activity and the Mg2 ligand Asn(195) is important for Mg.ATP recognition. The mutagenesis results together with solvent isotope, solvent viscosity, and divalent cation requirements are consistent with a dissociative mechanism of phosphoryl transfer where initial substrate deprotonation is not essential for phosphate transfer and where Mg2 and Asp(208) likely play a critical role in stabilization of a metaphosphate-like transition state. These results lay the foundation for the synthesis of transition state mimics that could reverse aminoglycoside antibiotic resistance in vivo. PMID- 11279090 TI - Role of Arg-166 in yeast cytochrome C1. AB - A systematic screen for dominant-negative mutations of the CYT1 gene, which encodes cytochrome c(1), revealed seven mutants after testing approximately 10(4) Saccharomyces cerevisiae strains transformed with a library of mutagenized multicopy plasmids. DNA sequence analysis revealed multiple nucleotide substitutions with six of the seven altered Cyt1p having a common R166G replacement, either by itself or accompanied with other amino acid replacements. A single R166G replacement produced by site-directed mutagenesis demonstrated that this change produced a nearly nonfunctional cytochrome c(1), with diminished growth on glycerol medium and diminished respiration but with the normal or near normal level of cytochrome c(1) having an attached heme group. In contrast, R166K, R166M, or R166L replacements resulted in normal or near normal function. Arg-166 is conserved in all cytochromes c(1) and lies on the surface of Cyt1p in close proximity to the heme group but does not seem to interact directly with any of the physiological partners of the cytochrome bc(1) complex. Thus, the large size of the side chain at position 166 is critical for the function of cytochrome c(1) but not for its assembly in the cytochrome bc(1) complex. PMID- 11279091 TI - Role of glutathione S-transferases in protection against lipid peroxidation. Overexpression of hGSTA2-2 in K562 cells protects against hydrogen peroxide induced apoptosis and inhibits JNK and caspase 3 activation. AB - The physiological significance of the selenium-independent glutathione peroxidase (GPx) activity of glutathione S-transferases (GSTs), associated with the major Alpha class isoenzymes hGSTA1-1 and hGSTA2-2, is not known. In the present studies we demonstrate that these isoenzymes show high GPx activity toward phospholipid hydroperoxides (PL-OOH) and they can catalyze GSH-dependent reduction of PL-OOH in situ in biological membranes. A major portion of GPx activity of human liver and testis toward phosphatidylcholine hydroperoxide (PC OOH) is contributed by the Alpha class GSTs. Overexpression of hGSTA2-2 in K562 cells attenuates lipid peroxidation under normal conditions as well as during the oxidative stress and confers about 1.5-fold resistance to these cells from H(2)O(2) cytotoxicity. Treatment with 30 microm H(2)O(2) for 48 h or 40 microm PC OOH for 8 h causes apoptosis in control cells, whereas hGSTA2-2-overexpressing cells are protected from apoptosis under these conditions. In control cells, H(2)O(2) treatment causes an early (within 2 h), robust, and persistent (at least 24 h) activation of JNK, whereas in hGSTA2-2-overexpressing cells, only a slight activation of JNK activity is observed at 6 h which declines to basal levels within 24 h. Caspase 3-mediated poly(ADP-ribose) polymerase cleavage is also inhibited in cells overexpressing hGSTA2-2. hGSTA2 transfection does not affect the function of antioxidant enzymes including GPx activity toward H(2)O(2) suggesting that the Alpha class GSTs play an important role in regulation of the intracellular concentrations of the lipid peroxidation products that may be involved in the signaling mechanisms of apoptosis. PMID- 11279092 TI - Recombinant human antibodies specific for the Pfs48/45 protein of the malaria parasite Plasmodium falciparum. AB - We report the first construction of two combinatorial human phage display libraries derived from malaria-immune patients. Specific single-chain Fv fragments (scFv) against Pfs48/45, a gamete surface protein of the sexual stages of Plasmodium falciparum, were selected and analyzed extensively. The selected scFv reacted with the surface of extracellular sexual forms of the parasite and showed Pfs48/45 reactivity on immunoblot. The scFv inhibit binding of human malaria sera to native Pfs48/45 from gametocytes. Moreover, the scFv bind to target epitopes of Pfs48/45 exposed in natural infections. Sequence analysis of eight scFv clones specific for epitope III of Pfs48/45 revealed that these clones could be divided into one V(H) family-derived germ-line gene (V(H)1) and two V(L) family segments (V(L)2 and V(K)I). PMID- 11279093 TI - Regulation and activity of the human ABCA1 gene in transgenic mice. AB - The ABCA1 transporter is one of the limiting steps in cellular cholesterol efflux. To study the expression and activity of the human ABCA1 gene in vivo we have examined mice containing two human BAC transgenes with different 5' ends. Mice containing a 255-kilobase (kb) BAC transgene, including 70 kb upstream of the previously defined exon 1, demonstrated a pattern of tissue-specific expression mimicking that of the endogenous mouse gene. Compared with macrophages from control mice, macrophages from these transgenics had increases in apoA-I cholesterol efflux heightened in response to increases in cell cholesterol content. The observed increase in macrophage apoA-I-mediated cholesterol efflux was not accompanied by alterations in plasma high density lipoprotein in the transgenics. Although mice containing a smaller 171-kb human BAC transgene, lacking the previously described exon 1 and ABCA1 promoter, did not express human ABCA1 in macrophages, they did express the human transgene in liver at levels comparable with those of the orthologous mouse gene. Analysis by 5' rapid amplification of cDNA ends of liver mRNA from these animals revealed a new ABCA1 exon 1 (exon 1A) and a previously unrecognized promoter. Analysis of human tissue revealed that exon 1A containing transcripts accounted for a high proportion of the ABCA1 mRNAs present in human liver. This analysis of ABCA1 transgenics showed that the expression of human ABCA1 transgenes can result in increased cholesterol efflux from macrophages, unaccompanied by changes in plasma high density lipoprotein, and identified a new ABCA1 promoter in humans. PMID- 11279094 TI - Negative regulatory role of Sp1 in metal responsive element-mediated transcriptional activation. AB - Transcription of mammalian metallothionein (MT) genes is activated by heavy metals via multiple copies of a cis-acting DNA element, the metal-responsive element (MRE). Our previous studies have shown that certain MREs of the human MT IIA gene (MREb, MREc, MREd, and MREf) are less active than the others (MREa, MREe, and MREg). Gel shift analysis of HeLa cell nuclear proteins revealed that whereas the active MREs strongly bind the transcription factor MTF-1 essential for metal regulation, the less active MREs bind another distinct protein, MREb BF. This protein recognizes the GC-rich region of MREb rather than the MRE core required for MTF-1 binding. All the MREs recognized by MREb-BF contain the CGCCC and/or CACCC motif, suggesting that the MREb-BF.MRE complex contains Sp1 or related proteins. Supershift analysis using antibodies against Sp1 family proteins as well as gel shift analysis using the recombinant Sp1 demonstrated that Sp1 represents the majority of MREb-BF activity. An MREb mutant with reduced affinity to Sp1 mediated zinc-inducible transcription much more actively than the wild-type MREb. Furthermore, when placed in the native promoter, this mutant MREb raised the overall promoter activity. These results strongly suggest that Sp1 acts as a negative regulator of transcription mediated by specific MREs. PMID- 11279095 TI - Unique disulfide bond structures found in ST8Sia IV polysialyltransferase are required for its activity. AB - NCAM polysialylation plays a critical role in neuronal development and regeneration. Polysialylation of the neural cell adhesion molecule (NCAM) is catalyzed by two polysialyltransferases, ST8Sia II (STX) and ST8Sia IV (PST), which contain sialylmotifs L and S conserved in all members of the sialyltransferases. The members of the ST8Sia gene family, including ST8Sia II and ST8Sia IV are unique in having three cysteines in sialylmotif L, one cysteine in sialylmotif S, and one cysteine at the COOH terminus. However, structural information, including how disulfide bonds are formed, has not been determined for any of the sialyltransferases. To obtain insight into the structure/function of ST8Sia IV, we expressed human ST8Sia IV in insect cells, Trichoplusia ni, and found that the enzyme produced in the insect cells catalyzes NCAM polysialylation, although it cannot polysialylate itself ("autopolysialylation"). We also found that ST8Sia IV does not form a dimer through disulfide bonds. By using the same enzyme preparation and performing mass spectrometric analysis, we found that the first cysteine in sialylmotif L and the cysteine in sialylmotif S form a disulfide bridge, whereas the second cysteine in sialylmotif L and the cysteine at the COOH terminus form a second disulfide bridge. Site-directed mutagenesis demonstrated that mutation at cysteine residues involved in the disulfide bridges completely inactivated the enzyme. Moreover, changes in the position of the COOH-terminal cysteine abolished its activity. By contrast, the addition of green fluorescence protein at the COOH terminus of ST8Sia IV did not render the enzyme inactive. These results combined indicate that the sterical structure formed by intramolecular disulfide bonds, which bring the sialylmotifs and the COOH terminus within close proximity, is critical for the catalytic activity of ST8Sia IV. PMID- 11279096 TI - Alanine-scanning mutagenesis of plasmatocyte spreading peptide identifies critical residues for biological activity. AB - Plasmatocyte spreading peptide (PSP) is a 23-amino acid cytokine that induces a class of insect immune cells called plasmatocytes to spread on foreign surfaces. The structure of PSP consists of a disordered N terminus (residues 1-6) and a well-defined core (residues 7-23) stabilized by a disulfide bridge between Cys(7) and Cys(19), hydrophobic interactions, and a short beta-hairpin. Structural comparisons also indicate that the core region of PSP adopts an epidermal growth factor (EGF)-like fold very similar to the C-terminal subdomain of EGF-like module 5 of thrombomodulin. To identify residues important for plasmatocyte spreading activity, we bioassayed PSP mutants in which amino acids were either replaced with alanine or deleted. Within the well-defined core of PSP, alanine replacement of Cys(7) and Cys(19) (C7.19A) eliminated all activity. Alanine replacement of Arg(13) reduced activity approximately 1000-fold in comparison to wild-type PSP, whereas replacement of the other charged residues (Asp(16), Arg(18), Lys(20)) surrounding Cys(19) diminished activity to a lesser degree. The point mutants Y11A, T14A, T22A, and F23A had activity identical or only slightly reduced to that of wild-type PSP. The mutant PSP-(7-23) lacked the entire unstructured domain of PSP and was found to have no plasmatocyte spreading activity. Surprisingly, E1A and N2A had higher activity than wild-type PSP, but F3A had almost no activity. We thus concluded that the lack of activity for PSP (7-23) was largely due to the critical importance of Phe(3). To determine whether reductions in activity correlated with alterations in tertiary structure, we compared the C7.19A, R13A, R18A, and F3A mutants to wild-type PSP by NMR spectroscopy. As expected, the simultaneous replacement of Cys(7) and Cys(19) profoundly affected tertiary structure, but the R13A, R18A, and F3A mutants did not differ from wild-type PSP. Collectively, these results indicate that residues in both the unstructured and structured domains of PSP are required for plasmatocyte-spreading activity. PMID- 11279097 TI - Molecular structure, processing, and tissue distribution of matrilin-4. AB - Matrilin-4 is the most recently identified member of the matrilin family of von Willebrand factor A-like domain containing extracellular matrix adapter proteins. Full-length matrilin-4 was expressed in 293-EBNA cells, purified using affinity tags, and subjected to biochemical characterization. The largest oligomeric form of recombinantly expressed full-length matrilin-4 is a trimer as shown by electron microscopy, SDS-polyacrylamide gel electrophoresis, and mass spectrometry. Proteolytically processed matrilin-4 species were also detected. The cleavage occurs in the short linker region between the second von Willebrand factor A-like domain and the coiled-coil domain leading to the release of large fragments and the formation of dimers and monomers of intact subunits still containing a trimeric coiled-coil. In immunoblots of calvaria extracts similar degradation products could be detected, indicating that a related proteolytic processing occurs in vivo. Matrilin-4 was first observed at day 7.5 post-coitum in mouse embryos. Affinity-purified antibodies detect a broad expression in dense and loose connective tissue, bone, cartilage, central and peripheral nervous systems and in association with basement membranes. In the matrix formed by cultured primary embryonic fibroblasts, matrilin-4 is found in a filamentous network connecting individual cells. PMID- 11279098 TI - Importance of homodimerization for the in vivo function of yeast RNA triphosphatase. AB - Saccharomyces cerevisiae RNA triphosphatase Cet1 is an essential component of the yeast mRNA capping apparatus. The active site of Cet1 resides within a topologically closed hydrophilic beta-barrel (the triphosphate tunnel) that is supported by a globular hydrophobic core. The homodimeric quaternary structure of Cet1 is formed by a network of contacts between the partner protomers. By studying the effects of alanine-cluster mutations, we highlight the contributions of two separate facets of the crystallographic dimer interface to Cet1 function in vivo. One essential facet of the interface entails hydrophobic cross-dimer interactions of Cys(330) and Val(331) and a cross-dimer hydrogen bond of Asp(280) with the backbone amide of Gln(329). The second functionally relevant dimer interface involves hydrophobic side-chain interactions of Phe(272) and Leu(273). Ala-cluster mutations involving these residues elicited lethal or severe temperature-sensitive phenotypes that were suppressed completely by fusion of the mutated triphosphatases to the guanylyltransferase domain of mammalian capping enzyme. The recombinant D279A-D280A and F272A-L273A proteins retained phosphohydrolase activity but sedimented as monomers. These results indicate that a disruption of the dimer interface is uniquely deleterious when the yeast RNA triphosphatase must function in concert with the endogenous yeast guanylyltransferase. We also identify key residue pairs in the hydrophobic core of the Cet1 protomer that support the active site tunnel and stabilize the triphosphatase in vivo. PMID- 11279099 TI - Mechanism of beta clamp opening by the delta subunit of Escherichia coli DNA polymerase III holoenzyme. AB - The beta sliding clamp encircles the primer-template and tethers DNA polymerase III holoenzyme to DNA for processive replication of the Escherichia coli genome. The clamp is formed via hydrophobic and ionic interactions between two semicircular beta monomers. This report demonstrates that the beta dimer is a stable closed ring and is not monomerized when the gamma complex clamp loader (gamma(3)delta(1)delta(1)chi(1)psi(1)) assembles the beta ring around DNA. delta is the subunit of the gamma complex that binds beta and opens the ring; it also does not appear to monomerize beta. Point mutations were introduced at the beta dimer interface to test its structural integrity and gain insight into its interaction with delta. Mutation of two residues at the dimer interface of beta, I272A/L273A, yields a stable beta monomer. We find that delta binds the beta monomer mutant at least 50-fold tighter than the beta dimer. These findings suggest that when delta interacts with the beta clamp, it binds one beta subunit with high affinity and utilizes some of that binding energy to perform work on the dimeric clamp, probably cracking one dimer interface open. PMID- 11279100 TI - The amino-terminal heptad repeats of the coiled-coil neck domain of pulmonary surfactant protein d are necessary for the assembly of trimeric subunits and dodecamers. AB - Pulmonary surfactant protein D (SP-D), a lung host defense protein, is assembled as multimers of trimeric subunits. Trimerization of SP-D monomers is required for high affinity saccharide binding, and the oligomerization of trimers is required for many of its functions. A peptide containing the alpha-helical neck region can spontaneously trimerize in vitro. However, it is not known whether this sequence is necessary for the complete cellular assembly of disulfide-cross-linked, trimeric subunits and dodecamers. For the present studies, we synthesized mutant cDNAs with deletions or site-directed substitutions in the neck domain of rat SP D, and examined the assembly of the newly synthesized proteins after transfection of CHO-K1 cells. The neck domain contains three "classical" heptad repeat motifs with leucine residues at the "d position," and a distinctive C-terminal repeat previously suggested to drive trimeric chain association. Deletion of the highly conserved core of the latter repeat (FSRYLKK) did not interfere with the secretion of dodecamers with lectin activity. By contrast, deletion of the entire neck domain or deletion of one or two amino-terminal repeats resulted in defective molecular assembly. The secreted proteins eluted in the position of monomers by gel filtration under nondenaturing conditions. In addition, the neck + carbohydrate recognition domain of SP-D was necessary and sufficient for the trimerization of a heterologous collagen sequence located amino-terminal to the trimeric coiled-coil. These studies provide strong evidence that the amino terminal heptad repeats of the neck domain are necessary for the intracellular, trimeric association of SP-D monomers and for the assembly and secretion of functional dodecamers. PMID- 11279101 TI - Association of the membrane proximal regions of the alpha and beta subunit cytoplasmic domains constrains an integrin in the inactive state. AB - The adhesiveness of integrins is regulated through a process termed "inside-out" signaling. To understand the molecular mechanism of integrin inside-out signaling, we generated K562 stable cell lines that expressed LFA-1 (alpha(L)beta(2)) or Mac-1 (alpha(M)beta(2)) with mutations in the cytoplasmic domain. Complete truncation of the beta(2) cytoplasmic domain, but not a truncation that retained the membrane proximal eight residues, resulted in constitutive activation of alpha(L)beta(2) and alpha(M)beta(2), demonstrating the importance of this membrane proximal region in the regulation of integrin adhesive function. Furthermore, replacement of the alpha(L) and beta(2) cytoplasmic domains with acidic and basic peptides that form an alpha-helical coiled coil caused inactivation of alpha(L)beta(2). Association of these artificial cytoplasmic domains was directly demonstrated. By contrast, replacement of the alpha(L) and beta(2) cytoplasmic domains with two basic peptides that do not form an alpha-helical coiled coil activated alpha(L)beta(2). Induction of ligand binding by the activating cytoplasmic domain mutations correlated with the induction of activation epitopes in the extracellular domain. Our data demonstrate that cytoplasmic, membrane proximal association between integrin alpha and beta subunits, constrains an integrin in the inactive conformation. PMID- 11279103 TI - Contextual equilibrium effects in DNA molecules. AB - The thermodynamic parameters of DNA triplex formation between oligonucleotides and double-stranded DNA segments containing adenine runs (A-tracts) were investigated to explore equilibrium structural effects exerted by flanking segments upon the A-tracts. Results obtained from isothermal titration calorimetry, temperature-dependent circular dichroism (CD), and UV melting experiments indicate that A-tracts, considered as a uniquely robust and inflexible DNA motif, can be structurally perturbed by neighboring sequences in a way that significantly affects the propensity of this motif to interact with triplex-forming oligonucleotides. These contextual equilibrium effects, which depend upon the composition and location of the flanking sequences, are likely to apply not only to the interaction of A-tracts with single-stranded DNA molecules but also to interactions with drugs and proteins. As such, the current results refine the guidelines for the design of triplex-forming oligonucleotides used for antigene strategies. More generally, they substantiate the notion that significant data might be encoded by structural DNA parameters. PMID- 11279102 TI - Sorting nexin 6, a novel SNX, interacts with the transforming growth factor-beta family of receptor serine-threonine kinases. AB - Sorting nexins (SNX) comprise a family of proteins with homology to several yeast proteins, including Vps5p and Mvp1p, that are required for the sorting of proteins to the yeast vacuole. Human SNX1, -2, and -4 have been proposed to play a role in receptor trafficking and have been shown to bind to several receptor tyrosine kinases, including receptors for epidermal growth factor, platelet derived growth factor, and insulin as well as the long form of the leptin receptor, a glycoprotein 130-associated receptor. We now describe a novel member of this family, SNX6, which interacts with members of the transforming growth factor-beta family of receptor serine-threonine kinases. These receptors belong to two classes: type II receptors that bind ligand, and type I receptors that are subsequently recruited to transduce the signal. Of the type II receptors, SNX6 was found to interact strongly with ActRIIB and more moderately with wild type and kinase-defective mutants of TbetaRII. Of the type I receptors, SNX6 was found to interact only with inactivated TbetaRI. SNXs 1-4 also interacted with the transforming growth factor-beta receptor family, showing different receptor preferences. Conversely, SNX6 behaved similarly to the other SNX proteins in its interactions with receptor tyrosine kinases. Strong heteromeric interactions were also seen among SNX1, -2, -4, and -6, suggesting the formation in vivo of oligomeric complexes. These findings are the first evidence for the association of the SNX family of molecules with receptor serine-threonine kinases. PMID- 11279104 TI - Two active molecular phenotypes of the tachykinin NK1 receptor revealed by G protein fusions and mutagenesis. AB - The NK1 neurokinin receptor presents two non-ideal binding phenomena, two component binding curves for all agonists and significant differences between agonist affinity determined by homologous versus heterologous competition binding. Whole cell binding with fusion proteins constructed between either Galpha(s) or Galpha(q) and the NK1 receptor with a truncated tail, which secured non-promiscuous G-protein interaction, demonstrated monocomponent agonist binding closely corresponding to either of the two affinity states found in the wild-type receptor. High affinity binding of both substance P and neurokinin A was observed in the tail-truncated Galpha(s) fusion construct, whereas the lower affinity component was displayed by the tail-truncated Galpha(q) fusion. The elusive difference between the affinity determined in heterologous versus homologous binding assays for substance P and especially for neurokinin A was eliminated in the G-protein fusions. An NK1 receptor mutant with a single substitution at the extracellular end of TM-III-(F111S), which totally uncoupled the receptor from Galpha(s) signaling, showed binding properties that were monocomponent and otherwise very similar to those observed in the tail-truncated Galpha(q) fusion construct. Thus, the heterogenous pharmacological phenotype displayed by the NK1 receptor is a reflection of the occurrence of two active conformations or molecular phenotypes representing complexes with the Galpha(s) and Galpha(q) species, respectively. We propose that these molecular forms do not interchange readily, conceivably because of the occurrence of microdomains or "signal transductosomes" within the cell membrane. PMID- 11279105 TI - Excision of 3' termini by the Trex1 and TREX2 3'-->5' exonucleases. Characterization of the recombinant proteins. AB - The excision of nucleotides from DNA 3' termini is an important step in DNA replication, repair, and recombination pathways to generate correctly base paired termini for subsequent processing. The mammalian TREX1 and TREX2 proteins contain potent 3'-->5' exonucleases capable of functioning in this capacity. To study the activities of these exonucleases we have developed strategies to express and purify the recombinant mouse Trex1 and human TREX2 proteins in Escherichia coli in quantities sufficient for biochemical characterization. The Trex1 and TREX2 proteins are homodimers that exhibit robust 3' excision activities with very similar preferred reaction conditions and preferences for specific DNA substrates. In a steady-state kinetic analysis, oligonucleotide substrates were used to measure 3' nucleotide excision by Trex1 and TREX2. The Michaelis constants derived from these data indicate similar apparent kcat values of 22 s( 1) for Trex1 and 16 s(-1) for TREX2 using single-stranded oligonucleotides. The apparent KM values of 19 nm for Trex1 and 190 nm for TREX2 suggest relatively high affinities for DNA for both Trex1 and TREX2. An exonuclease competition assay was designed using heparin as a nonsubstrate inhibitor with a series of partial duplex DNAs to delineate the substrate structure preferences for 3' nucleotide excision by Trex1 and TREX2. The catalytic properties of the TREX proteins suggest roles for these enzymes in the 3' end-trimming processes necessary for producing correctly base paired 3' termini. PMID- 11279107 TI - Cloning of an immunoglobulin family adhesion molecule selectively expressed by endothelial cells. AB - To gain fundamental information regarding the molecular basis of endothelial cell adhesive interactions during vascular formation, we have cloned and characterized a unique cell adhesion molecule. This molecule, named endothelial cell-selective adhesion molecule (ESAM), is a new member of the immunoglobulin superfamily. The conceptual protein encoded by cDNA clones consists of V-type and C2-type immunoglobulin domains as well as a hydrophobic signal sequence, a single transmembrane region, and a cytoplasmic domain. Northern blot analysis showed ESAM to be selectively expressed in cultured human and murine vascular endothelial cells and revealed high level expression in lung and heart and low level expression in kidney and skin. In situ hybridization analysis indicated that ESAM is primarily expressed in the developing vasculature of the embryo in an endothelial cell-restricted pattern. Epitope-tagged ESAM was shown to co localize with cadherins and catenins in cell-cell junctions. In aggregation assays employing ESAM-expressing Chinese hamster ovary cells, this novel molecule was shown to mediate cell-cell adhesion through homophilic interactions. The endothelial cell-selective expression of this immunoglobulin-like adhesion molecule coupled with its in vitro functional profile strongly suggests a role in cell-cell interactions that is critical for vascular development or function. PMID- 11279106 TI - Peroxidase self-inactivation in prostaglandin H synthase-1 pretreated with cyclooxygenase inhibitors or substituted with mangano protoporphyrin IX. AB - Self-inactivation imposes an upper limit on bioactive prostanoid synthesis by prostaglandin H synthase (PGHS). Inactivation of PGHS peroxidase activity has been found to begin with Intermediate II, which contains a tyrosyl radical. The structure of this radical is altered by cyclooxygenase inhibitors, such as indomethacin and flurbiprofen, and by replacement of heme by manganese protoporphyrin IX (forming MnPGHS-1). Peroxidase self-inactivation in inhibitor treated PGHS-1 and MnPGHS-1 was characterized by stopped-flow spectroscopic techniques and by chromatographic and mass spectrometric analysis of the metalloporphyrin. The rate of peroxidase inactivation was about 0.3 s(-)1 in inhibitor-treated PGHS-1 and much slower in MnPGHS-1 (0.05 s(-)1); as with PGHS-1 itself, the peroxidase inactivation rates were independent of peroxide concentration and structure, consistent with an inactivation process beginning with Intermediate II. The changes in metalloporphyrin absorbance spectra during inactivation of inhibitor-treated PGHS-1 were similar to those observed with PGHS 1 but were rather distinct in MnPGHS-1; the kinetics of the spectral transition from Intermediate II to the next species were comparable to the inactivation kinetics in each case. In contrast to the situation with PGHS-1 itself, significant amounts of heme degradation occurred during inactivation of inhibitor treated PGHS-1, producing iron chlorin and heme-protein adduct species. Structural perturbations at the peroxidase site (MnPGHS-1) or at the cyclooxygenase site (inhibitor-treated PGHS-1) thus can influence markedly the kinetics and the chemistry of PGHS-1 peroxidase inactivation. PMID- 11279108 TI - Binding of serum response factor to CArG box sequences is necessary but not sufficient to restrict gene expression to arterial smooth muscle cells. AB - Serum response factor (SRF) plays an important role in regulating smooth muscle cell (SMC) development and differentiation. To understand the molecular mechanisms underlying the activity of SRF in SMCs, the two CArG box-containing elements in the arterial SMC-specific SM22alpha promoter, SME-1 and SME-4, were functionally and biochemically characterized. Mutations that abolish binding of SRF to the SM22alpha promoter totally abolish promoter activity in transgenic mice. Moreover, a multimerized copy of either SME-1 or SME-4 subcloned 5' of the minimal SM22alpha promoter (base pairs -90 to +41) is necessary and sufficient to restrict transgene expression to arterial SMCs in transgenic mice. In contrast, a multimerized copy of the c-fos SRE is totally inactive in arterial SMCs and substitution of the c-fos SRE for the CArG motifs within the SM22alpha promoter inactivates the 441-base pair SM22alpha promoter in transgenic mice. Deletion analysis revealed that the SME-4 CArG box alone is insufficient to activate transcription in SMCs and additional 5'-flanking nucleotides are required. Nuclear protein binding assays revealed that SME-4 binds SRF, YY1, and four additional SMC nuclear proteins. Taken together, these data demonstrate that binding of SRF to specific CArG boxes is necessary, but not sufficient, to restrict transgene expression to SMCs in vivo. PMID- 11279109 TI - CRP modulates fis transcription by alternate formation of activating and repressing nucleoprotein complexes. AB - The DNA architectural proteins FIS and CRP are global regulators of transcription in Escherichia coli involved in the adjustment of cellular metabolism to varying growth conditions. We have previously demonstrated that FIS modulates the expression of the crp gene by functioning as its transcriptional repressor. Here we show that in turn, CRP is required to maintain the growth phase pattern of fis expression. We demonstrate the existence of a divergent promoter in the fis regulatory region, which reduces transcription of the fis promoter. In the absence of FIS, CRP activates fis transcription, thereby displacing the polymerase from the divergent promoter, whereas together FIS and CRP synergistically repress fis gene expression. These results provide evidence for a direct cross-talk between global regulators of cellular transcription during the growth phase. This cross-talk is manifested in alternate formation of functional nucleoprotein complexes exerting either activating or repressing effects on transcription. PMID- 11279110 TI - Requirement for HDM2 activity in the rapid degradation of p53 in neuroblastoma. AB - The wild type p53 tumor suppressor protein is rapidly degraded in normal cells by MDM2, the ubiquitin ligase that serves as the key regulator of p53 function by modulating protein stability. Cellular exposure to genotoxic stress triggers the stabilization of p53 by multiple pathways that converge upon interference with MDM2 function. In this study, we first investigated the ability of HDM2 (MDM2 human homologue) to degrade endogenous p53 in neuroblastoma (NB). Although the p53 protein in NB has been reported to be constitutively stabilized, we find that HDM2 in NB is functional and facilitates the rapid turnover of p53 in nonstressed cells via the proteasome pathway. Second, we examined the relationship between p53 and HDM2 in the adriamycin-mediated stabilization of p53 in NB. We demonstrate that while p53 stabilization depends neither upon the phosphorylation of specific N-terminal sites nor upon dissociation from HDM2, it requires inactivation of functional HDM2. In support of this notion, p53 stabilization following adriamycin resulted in an inhibition of both p53 ubiquitination and HDM2 ligase activity. Taken together, these data implicate a requirement for enzymatic inactivation of HDM2 as a novel mechanism for p53 stabilization in the DNA damage response pathway. PMID- 11279111 TI - Molecular mechanisms regulating the differential association of kainate receptor subunits with SAP90/PSD-95 and SAP97. AB - Recent studies have demonstrated that kainate receptors are associated with members of the SAP90/PSD-95 family (synapse-associated proteins (SAPs)) in neurons and that SAP90 can cluster and modify the electrophysiological properties of GluR6/KA2 kainate receptors when co-expressed in transfected cells. In vivo, SAP90 tightly binds kainate receptor subunits, while SAP97 is only weakly associated, suggesting that this glutamate receptor differentially associates with SAP90/PSD-95 family members. Here, green fluorescent protein (GFP)-tagged chimeras and deletion mutants of SAP97 and SAP90 were employed to define the molecular mechanism underlying their differential association with kainate receptors. Our results show that a weak interaction between GluR6 and the PDZ1 domain of SAP97 can account for the weak association of GluR6 with the full length SAP97 observed in vivo. Expression studies in HEK293 cells and in vitro binding studies further show that although the individual Src homology 3 and guanylate kinase domains in SAP97 can interact with the C-terminal tail of KA2 subunit, specific intramolecular interactions in SAP97 (e.g. the SAP97 N terminus (S97N) binding to the Src homology 3 domain) interfere with KA2 binding to the full-length molecule. Because receptor subunits are known to segregate to different parts of the neuron, our results imply that differential association of kainate receptors with SAP family proteins may be one mechanism of subcellular localization. PMID- 11279112 TI - Bcl-2 prevents Bax oligomerization in the mitochondrial outer membrane. AB - ATP depletion results in Bax translocation from cytosol to mitochondria and release of cytochrome c from mitochondria into cytosol in cultured kidney cells. Overexpression of Bcl-2 prevents cytochrome c release, without ameliorating ATP depletion or Bax translocation, with little or no association between Bcl-2 and Bax as demonstrated by immunoprecipitation (Saikumar, P., Dong, Z., Patel, Y., Hall, K., Hopfer, U., Weinberg, J. M., and Venkatachalam, M. A. (1998) Oncogene 17, 3401-3415). Now we show that translocated Bax forms homo-oligomeric structures, stabilized as chemical adducts by bifunctional cross-linkers in ATP depleted wild type cells, but remains monomeric in Bcl-2-overexpressing cells. The protective effects of Bcl-2 did not require Bcl-2/Bax association, at least to a degree of proximity or affinity that was stable to conditions of immunoprecipitation or adduct formation by eight cross-linkers of diverse spacer lengths and chemical reactivities. On the other hand, nonionic detergents readily induced homodimers and heterodimers of Bax and Bcl-2. Moreover, associations between translocated Bax and the voltage-dependent anion channel protein or the adenine nucleotide translocator protein could not be demonstrated by immunoprecipitation of Bax, or by using bifunctional cross-linkers. Our data suggest that the in vivo actions of Bax are at least in part dependent on the formation of homo-oligomers without requiring associations with other molecules and that Bcl-2 cytoprotection involves mechanisms that prevent Bax oligomerization. PMID- 11279113 TI - Characterization of a cluster of human high/ultrahigh sulfur keratin-associated protein genes embedded in the type I keratin gene domain on chromosome 17q12-21. AB - Low stringency screening of a human P1 artificial chromosome library using a human hair keratin-associated protein (hKAP1.1A) gene probe resulted in the isolation of six P1 artificial chromosome clones. End sequencing and EMBO/GenBank(TM) data base analysis showed these clones to be contained in four previously sequenced human bacterial artificial chromosome clones present on chromosome 17q12-21 and arrayed into two large contigs of 290 and 225 kilobase pairs (kb) in size. A fifth, partially sequenced human bacterial artificial chromosome clone data base sequence overlapped and closed both of these contigs. One end of this 600-kb cluster harbored six gene loci for previously described human type I hair keratin genes. The other end of this cluster contained the human type I cytokeratin K20 and K12 gene loci. The center of the cluster, starting 35 kb downstream of the hHa3-I hair keratin gene, contained 37 genes for high/ultrahigh sulfur hair keratin-associated proteins (KAPs), which could be divided into a total of 7 KAP multigene families based on amino acid homology comparisons with previously identified sheep, mouse, and rabbit KAPs. To date, 26 human KAP cDNA clones have been isolated through screening of an arrayed human scalp cDNA library by means of specific 3'-noncoding region polymerase chain reaction probes derived from the identified KAP gene sequences. This screening also yielded four additional cDNA sequences whose genes were not present on this gene cluster but belonged to specific KAP gene families present on this contig. Hair follicle in situ hybridization data for single members of five different KAP multigene families all showed localization of the respective mRNAs to the upper cortex of the hair shaft. PMID- 11279114 TI - Analysis of the phthiocerol dimycocerosate locus of Mycobacterium tuberculosis. Evidence that this lipid is involved in the cell wall permeability barrier. AB - Among the few characterized genes that have products involved in the pathogenicity of Mycobacterium tuberculosis, the etiological agent of tuberculosis, are those of the phthiocerol dimycocerosate (DIM) locus. Genes involved in biosynthesis of these compounds are grouped on a 50-kilobase fragment of the chromosome containing 13 genes. Analysis of mRNA produced from this 50 kilobase fragment in the wild type strain showed that this region is subdivided into three transcriptional units. Biochemical characterization of five mutants with transposon insertions in this region demonstrated that (i) the complete DIM molecules are synthesized in the cytoplasm of M. tuberculosis before being translocated into the cell wall; (ii) the genes fadD26 and fadD28 are directly involved in their biosynthesis; and (iii) both the drrC and mmpL7 genes are necessary for the proper localization of DIMs. Insertional mutants unable to synthesize or translocate DIMs exhibit higher cell wall permeability and are more sensitive to detergent than the wild type strain, indicating for the first time that, in addition to being important virulence factors, extractable lipids of M. tuberculosis play a role in the cell envelope architecture and permeability. This function may represent one of the molecular mechanisms by which DIMs are involved in the virulence of M. tuberculosis. PMID- 11279115 TI - Physical interaction and functional synergy between glucocorticoid receptor and Ets2 proteins for transcription activation of the rat cytochrome P-450c27 promoter. AB - We demonstrate that dexamethasone-mediated transcription activation of the cytochrome P-450c27 promoter involves a physical interaction and functional synergy between glucocorticoid receptor (GR) and Ets2 factor. Ets2 protein binding to a "weak" Ets-like site of the promoter is dependent on GR bound to the adjacent cryptic glucocorticoid response element. Coimmunoprecipitation and chemical cross-linking experiments show physical interaction between GR and Ets2 proteins. Mutational analyses show synergistic effects of Ets2 and GR in dexamethasone-mediated activation of the cytochrome P-450c27 promoter. The DNA binding domain of GR, lacking the transcription activation and ligand-binding domains, was fully active in synergistic activation of the promoter with intact Ets2. The DNA-binding domain of Ets2 lacking the transcription activation domain showed a dominant negative effect on the transcription activity. Finally, a fusion protein consisting of the GR DNA-binding domain and the transcription activation domain of Ets2 fully supported the transcription activity, suggesting a novel synergy between the two proteins, which does not require the transactivation domain of GR. Our results also provide new insights on the role of putative weak consensus Ets sites in transcription activation, possibly through synergistic interaction with other gene-specific transcription activators. PMID- 11279116 TI - DNA binding and transcriptional activation by a PDX1.PBX1b.MEIS2b trimer and cooperation with a pancreas-specific basic helix-loop-helix complex. AB - In pancreatic acinar cells, the HOX-like factor PDX1 acts as part of a trimeric complex with two TALE class homeodomain factors, PBX1b and MEIS2b. The complex binds to overlapping half-sites for PDX1 and PBX. The trimeric complex activates transcription in cells to a level about an order of magnitude greater than PDX1 alone. The N-terminal PDX1 activation domain is required for detectable transcriptional activity of the complex, even though PDX1 truncations bearing only the PDX1 C-terminal homeodomain and pentapeptide motifs can still participate in forming the trimeric complex. The conserved N-terminal PBC-B domain of PBX, as well as its homeodomain, is required for both complex formation and transcriptional activity. Only the N-terminal region of MEIS2, including the conserved MEIS domains, is required for formation of a trimer on DNA and transcriptional activity: the MEIS homeodomain is dispensable. The activity of the pancreas-specific ELA1 enhancer requires the cooperation of the trimer binding element and a nearby element that binds the pancreatic transcription factor PTF1. We show that the PDX1. PBX1b.MEIS2b complex cooperates with the PTF1 basic helix-loop-helix complex to activate an ELA1 minienhancer in HeLa cells and that this cooperation requires all three homeoprotein subunits, including the PDX1 activation domain. PMID- 11279117 TI - Dysregulated cannabinoid signaling disrupts uterine receptivity for embryo implantation. AB - The mechanisms by which synchronized embryonic development to the blastocyst stage, preparation of the uterus for the receptive state, and reciprocal embryo uterine interactions for implantation are coordinated are still unclear. We show in this study that preimplantation embryo development became asynchronous in mice that are deficient in brain-type (CB1) and/or spleen-type (CB2) cannabinoid receptor genes. Furthermore, whereas the levels of uterine anandamide (endocannabinoid) and blastocyst CB1 are coordinately down-regulated with the onset of uterine receptivity and blastocyst activation prior to implantation, these levels remained high in the nonreceptive uterus and in dormant blastocysts during delayed implantation and in pregnant, leukemia inhibitory factor (LIF) deficient mice with implantation failure. These results suggest that a tight regulation of endocannabinoid signaling is important for synchronizing embryo development with uterine receptivity for implantation. Indeed this is consistent with our finding that while an experimentally induced, sustained level of an exogenously administered, natural cannabinoid inhibited implantation in wild-type mice, it failed to do so in CB1(-/-)/CB2(-/-) double mutant mice. The present study is clinically important because of the widely debated medicinal use of cannabinoids and their reported adverse effects on pregnancy. PMID- 11279118 TI - Regulation of stress-responsive mitogen-activated protein (MAP) kinase pathways by TAO2. AB - Previous studies demonstrated that in vitro the protein kinase TAO2 activates MAP/ERK kinases (MEKs) 3, 4, and 6 toward their substrates p38 MAP kinase and c Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK). In this study, we examined the ability of TAO2 to activate stress-sensitive MAP kinase pathways in cells and the relationship between activation of TAO2 and potential downstream pathways. Over-expression of TAO2 activated endogenous JNK/SAPK and p38 but not ERK1/2. Cotransfection experiments suggested that TAO2 selectively activates MEK3 and MEK6 but not MEKs 1, 4, or 7. Coimmunoprecipitation demonstrated that endogenous TAO2 specifically associates with MEK3 and MEK6 providing one mechanism for preferential recognition of MEKs upstream of p38. Sorbitol, and to a lesser extent, sodium chloride, Taxol, and nocodazole increased TAO2 activity toward itself and kinase-dead MEKs 3 and 6. Activation of endogenous TAO2 during differentiation of C2C12 myoblasts paralleled activation of p38 but not JNK/SAPK, consistent with the idea that TAO2 is a physiological regulator of p38 under certain circumstances. PMID- 11279119 TI - Adjacent basic amino acid residues recognized by the COP I complex and ubiquitination govern endoplasmic reticulum to cell surface trafficking of the nicotinic acetylcholine receptor alpha-Subunit. AB - The nicotinic acetylcholine receptor in muscle is a ligand-gated ion channel with an ordered subunit arrangement of alpha-gamma-alpha-delta-beta. The subunits are sequestered in the endoplasmic reticulum (ER) and assembled into the pentameric arrangement prior to their exit to the cell surface. Mutating the Arg(313) Lys(314) sequence in the large cytoplasmic loop of the alpha-subunit to K314Q promotes the trafficking of the mutant unassembled alpha-subunit from the ER to the Golgi in transfected HEK cells, identifying an important determinant that modulates the ER to Golgi trafficking of the subunit. The association of the K314Q alpha-subunit with gamma-COP, a component of COP I coats implicated in Golgi to ER anterograde transport, is diminished to a level comparable to that observed for wild-type alpha-subunits when co-expressed with the beta-, delta-, and gamma-subunits. This suggests that the Arg(313)-Lys(314) sequence is masked when the subunits assemble, thereby enabling ER to Golgi trafficking of the alpha subunit. Although unassembled K314Q alpha-subunits accumulate in the Golgi, they are not detected at the cell surface, suggesting that a second post-Golgi level of capture exists. Expressing the K314Q alpha-subunit in the absence of the other subunits in ubiquitinating deficient cells (ts20) results in detecting this subunit at the cell surface, indicating that ubiquitination functions as a post Golgi modulator of trafficking. Taken together, our findings support the hypothesis that subunit assembly sterically occludes the trafficking signals and ubiquitination at specific sites. Following the masking of these signals, the assembled ion channel expresses at the cell surface. PMID- 11279120 TI - Tyrosine-phosphorylated caveolin is a physiological substrate of the low M(r) protein-tyrosine phosphatase. AB - Low M(r) phosphotyrosine-protein phosphatase is involved in the regulation of several tyrosine kinase growth factor receptors. The best characterized action of this enzyme is on the signaling pathways activated by platelet-derived growth factor, where it plays multiple roles. In this study we identify tyrosine phosphorylated caveolin as a new potential substrate for low M(r) phosphotyrosine protein phosphatase. Caveolin is tyrosine-phosphorylated in vivo by Src kinases, recruits into caveolae, and hence regulates the activities of several proteins involved in cellular signaling cascades. Our results demonstrate that caveolin and low M(r) phosphotyrosine-protein phosphatase coimmunoprecipitate from cell lysates, and that a fraction of the enzyme localizes in caveolae. Furthermore, in a cell line sensitive to insulin, the overexpression of the C12S dominant negative mutant of low M(r) phosphotyrosine-protein phosphatase (a form lacking activity but able to bind substrates) causes the enhancement of tyrosine phosphorylated caveolin. Insulin stimulation of these cells induces a strong increase of caveolin phosphorylation. The localization of low M(r) phosphotyrosine-protein phosphatase in caveolae, the in vivo interaction between this enzyme and caveolin, and the capacity of this enzyme to rapidly dephosphorylate phosphocaveolin, all indicate that tyrosine-phosphorylated caveolin is a relevant substrate for this phosphatase. PMID- 11279121 TI - Structural properties of carnation mottle virus p7 movement protein and its RNA binding domain. AB - Plant viral movement proteins (MPs) participate actively in the intra- and intercellular movement of RNA plant viruses to such an extent that MP dysfunction impairs viral infection. However, the molecular mechanism(s) of their interaction with cognate nucleic acids are not well understood, partly due to the lack of structural information. In this work, a protein dissection approach was used to gain information on the structural and RNA-binding properties of this class of proteins, as exemplified by the 61-amino acid residue p7 MP from carnation mottle virus (CarMV). Circular dichroism spectroscopy showed that CarMV p7 is an alpha/beta RNA-binding soluble protein. Using synthetic peptides derived from the p7 sequence, we have identified three distinct putative domains within the protein. EMSA showed that the central region, from residue 17 to 35 (represented by peptide p7(17-35)), is responsible for the RNA binding properties of CarMV p7. This binding peptide populates a nascent alpha-helix in water solution that is further stabilized in the presence of either secondary structure inducers, such as trifluoroethanol and monomeric SDS, or RNA (which also changes its conformation upon binding to the peptide). Thus, the RNA recognition appears to occur via an "adaptive binding" mechanism. Interestingly, the amino acid sequence and structural properties of the RNA-binding domain of p7 seem to be conserved among carmoviruses and some other RNA-binding proteins and peptides. The low conserved N terminus of p7 (peptide p7(1-16)) is unstructured in solution. In contrast, the highly conserved C terminus motif (peptide p7(40-61)) adopts a beta sheet conformation in aqueous solution. Alanine scanning mutagenesis of the RNA binding motif showed how selected positive charged amino acids are more relevant than others in the RNA binding process and how hydrophobic amino acid side chains would participate in the stabilization of the protein-RNA complex. PMID- 11279122 TI - Early-onset amyloid deposition and cognitive deficits in transgenic mice expressing a double mutant form of amyloid precursor protein 695. AB - We have created early-onset transgenic (Tg) models by exploiting the synergistic effects of familial Alzheimer's disease mutations on amyloid beta-peptide (Abeta) biogenesis. TgCRND8 mice encode a double mutant form of amyloid precursor protein 695 (KM670/671NL+V717F) under the control of the PrP gene promoter. Thioflavine S positive Abeta amyloid deposits are present at 3 months, with dense-cored plaques and neuritic pathology evident from 5 months of age. TgCRND8 mice exhibit 3,200 4,600 pmol of Abeta42 per g brain at age 6 months, with an excess of Abeta42 over Abeta40. High level production of the pathogenic Abeta42 form of Abeta peptide was associated with an early impairment in TgCRND8 mice in acquisition and learning reversal in the reference memory version of the Morris water maze, present by 3 months of age. Notably, learning impairment in young mice was offset by immunization against Abeta42 (Janus, C., Pearson, J., McLaurin, J., Mathews, P. M., Jiang, Y., Schmidt, S. D., Chishti, M. A., Horne, P., Heslin, D., French, J., Mount, H. T. J., Nixon, R. A., Mercken, M., Bergeron, C., Fraser, P. E., St. George-Hyslop, P., and Westaway, D. (2000) Nature 408, 979-982). Amyloid deposition in TgCRND8 mice was enhanced by the expression of presenilin 1 transgenes including familial Alzheimer's disease mutations; for mice also expressing a M146L+L286V presenilin 1 transgene, amyloid deposits were apparent by 1 month of age. The Tg mice described here suggest a potential to investigate aspects of Alzheimer's disease pathogenesis, prophylaxis, and therapy within short time frames. PMID- 11279123 TI - The small subunit of the mammalian mitochondrial ribosome. Identification of the full complement of ribosomal proteins present. AB - Identification of all the protein components of the small subunit (28 S) of the mammalian mitochondrial ribosome has been achieved by carrying out proteolytic digestions of whole 28 S subunits followed by analysis of the resultant peptides by liquid chromatography and tandem mass spectrometry (LC/MS/MS). Peptide sequence information was used to search the human EST data bases and complete coding sequences of the proteins were assembled. The human mitochondrial ribosome has 29 distinct proteins in the small subunit. Fourteen of this group of proteins are homologs of the Escherichia coli 30 S ribosomal proteins S2, S5, S6, S7, S9, S10, S11, S12, S14, S15, S16, S17, S18, and S21. All of these proteins have homologs in Drosophila melanogaster, Caenorhabditis elegans, and Saccharomyces cerevisiae mitochondrial ribosomes. Surprisingly, three variants of ribosomal protein S18 are found in the mammalian and D. melanogaster mitochondrial ribosomes while C. elegans has two S18 homologs. The S18 homologs tend to be more closely related to chloroplast S18s than to prokaryotic S18s. No mitochondrial homologs to prokaryotic ribosomal proteins S1, S3, S4, S8, S13, S19, and S20 could be found in the peptides obtained from the whole 28 S subunit digests or by analysis of the available data bases. The remaining 15 proteins present in mammalian mitochondrial 28 S subunits (MRP-S22 through MRP-S36) are specific to mitochondrial ribosomes. Proteins in this group have no apparent homologs in bacterial, chloroplast, archaebacterial, or cytosolic ribosomes. All but two of these proteins have a clear homolog in D. melanogaster while all but three can be found in the genome of C. elegans. Five of the mitochondrial specific ribosomal proteins have homologs in S. cerevisiae. PMID- 11279124 TI - Inhibition of the G2 DNA damage checkpoint and of protein kinases Chk1 and Chk2 by the marine sponge alkaloid debromohymenialdisine. AB - Cells can respond to DNA damage by activating checkpoints that delay cell cycle progression and allow time for DNA repair. Chemical inhibitors of the G(2) phase DNA damage checkpoint may be used as tools to understand better how the checkpoint is regulated and may be used to sensitize cancer cells to DNA-damaging therapies. However, few inhibitors are known. We used a cell-based assay to screen natural extracts for G(2) checkpoint inhibitors and identified debromohymenialdisine (DBH) from a marine sponge. DBH is distinct structurally from previously known G(2) checkpoint inhibitors. It inhibited the G(2) checkpoint with an IC(50) of 8 micrometer and showed moderate cytotoxicity (IC(50) = 25 micrometer) toward MCF-7 cells. DBH inhibited the checkpoint kinases Chk1 (IC(50) = 3 micrometer) and Chk2 (IC(50) = 3.5 micrometer) but not ataxia telangiectasia mutated (ATM), ATM-Rad3-related protein, or DNA-dependent protein kinase in vitro, indicating that it blocks two major branches of the checkpoint pathway downstream of ATM. It did not cause the activation or inhibition of different signal transduction proteins, as determined by mobility shift analysis in Western blots, suggesting that it inhibits a narrow range of protein kinases in vivo. PMID- 11279125 TI - Calcium is a key signaling molecule in beta-lapachone-mediated cell death. AB - beta-Lapachone (beta-Lap) triggers apoptosis in a number of human breast and prostate cancer cell lines through a unique apoptotic pathway that is dependent upon NQO1, a two-electron reductase. Downstream signaling pathway(s) that initiate apoptosis following treatment with beta-Lap have not been elucidated. Since calpain activation was suspected in beta-Lap-mediated apoptosis, we examined alterations in Ca(2+) homeostasis using NQO1-expressing MCF-7 cells. beta-Lap-exposed MCF-7 cells exhibited an early increase in intracellular cytosolic Ca(2+), from endoplasmic reticulum Ca(2+) stores, comparable to thapsigargin exposures. 1,2-Bis-(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester, an intracellular Ca(2+) chelator, blocked early increases in Ca(2+) levels and inhibited beta-Lap-mediated mitochondrial membrane depolarization, intracellular ATP depletion, specific and unique substrate proteolysis, and apoptosis. The extracellular Ca(2+) chelator, EGTA, inhibited later apoptotic end points (observed >8 h, e.g. substrate proteolysis and DNA fragmentation), suggesting that later execution events were triggered by Ca(2+) influxes from the extracellular milieu. Collectively, these data suggest a critical, but not sole, role for Ca(2+) in the NQO1-dependent cell death pathway initiated by beta-Lap. Use of beta-Lap to trigger an apparently novel, calpain like-mediated apoptotic cell death could be useful for breast and prostate cancer therapy. PMID- 11279126 TI - Follistatin: essential role for the N-terminal domain in activin binding and neutralization. AB - Follistatin is recognized to be an important regulator of cellular differentiation and secretion through its potent ability to bind and bioneutralize activin with which it is colocalized in many tissue systems. The 288-residue follistatin molecule is comprised of a 63-residue N-terminal segment followed by three repeating 10-cysteine "follistatin domains" also represented in several extracellular matrix proteins. We have used chemical modifications and mutational analyses to define structural requirements for follistatin bioactivity that previously have not been investigated systematically. Mutant follistatins were stably expressed from Chinese hamster ovary cell cultures and assayed for activin binding in a solid-phase competition assay. Biological activities were determined by inhibition of activin-mediated transcriptional activity and by suppression of follicle-stimulating hormone secretion by cultured anterior pituitary cells. Deletion of the entire N-terminal domain, disruption of N terminal disulfides, and deletion of the first two residues each reduced activin binding to <5 % of expressed wild-type follistatin and abolished the ability of the respective mutants to suppress activin-mediated responses in both bioassay systems. Hence, the three follistatin domains inherently lack the ability to bind or neutralize activin. Activin binding was impaired after oxidation of at least one tryptophan, at position 4, in FS-288. Mutation of Trp to Ala or Asp at either positions 4 or 36 eliminated activin binding and bioactivity. Mutation of a third hydrophobic residue, Phe-52, reduced binding to 20%, whereas substitutions for the individual Lys and Arg residues in the N-terminal region were tolerated. These results establish that hydrophobic residues within the N-terminal domain constitute essential activin-binding determinants in the follistatin molecule. The correlation among the effects of mutation on activin binding, activin transcriptional responses, and follicle-stimulating hormone secretion substantiates the concept that, at least in the pituitary, the biological activity of follistatin is attributable to its ability to bind and bioneutralize activin. PMID- 11279127 TI - Identification of novel TGF-beta /Smad gene targets in dermal fibroblasts using a combined cDNA microarray/promoter transactivation approach. AB - Despite major advances in the understanding of the intimate mechanisms of transforming growth factor-beta (TGF-beta) signaling through the Smad pathway, little progress has been made in the identification of direct target genes. In this report, using cDNA microarrays, we have focussed our attention on the characterization of extracellular matrix-related genes rapidly induced by TGF beta in human dermal fibroblasts and attempted to identify the ones whose up regulation by TGF-beta is Smad-mediated. For a gene to qualify as a direct Smad target, we postulated that it had to meet the following criteria: (1) rapid (30 min) and significant (at least 2-fold) elevation of steady-state mRNA levels upon TGF-beta stimulation, (2) activation of the promoter by both exogenous TGF-beta and co-transfected Smad3 expression vector, (3) up-regulation of promoter activity by TGF-beta blocked by both dominant-negative Smad3 and inhibitory Smad7 expression vectors, and (4) promoter transactivation by TGF-beta not possible in Smad3(-/-) mouse embryo fibroblasts. Using this stringent approach, we have identified COL1A2, COL3A1, COL6A1, COL6A3, and tissue inhibitor of metalloproteases-1 as definite TGF-beta/Smad3 targets. Extrapolation of this approach to other extracellular matrix-related gene promoters also identified COL1A1 and COL5A2, but not COL6A2, as novel Smad targets. Together, these results represent a significant step toward the identification of novel, early-induced Smad-dependent TGF-beta target genes in fibroblasts. PMID- 11279128 TI - The binding of Ku antigen to homeodomain proteins promotes their phosphorylation by DNA-dependent protein kinase. AB - The Ku antigen (70- and 80-kDa subunits) is a regulatory subunit of DNA-dependent protein kinase (DNA-PK) that promotes the recruitment of the catalytic subunit of DNA-PK (DNA-PKcs) to DNA ends and to specific DNA sequences from which the kinase is activated. Ku and DNA-PKcs plays essential roles in double-stranded DNA break repair and V(D)J recombination and have been implicated in the regulation of specific gene transcription. In a yeast two-hybrid screen of a Jurkat T cell cDNA library, we have identified a specific interaction between the 70-kDa subunit of Ku heterodimer and the homeodomain of HOXC4, a homeodomain protein expressed in the hematopoietic system. Unexpectedly, a similar interaction with Ku was observed for several additional homeodomain proteins including octamer transcription factors 1 and 2 and Dlx2, suggesting that specific binding to Ku may be a property shared by many homeodomain proteins. Ku-homeodomain binding was mediated through the extreme C terminus of Ku70 and was abrogated by amino acid substitutions at Lys595/Lys596. Ku binding allowed the recruitment of the homeodomain to DNA ends and dramatically enhanced the phosphorylation of homeodomain-containing proteins by DNA-PK. These results suggest that Ku functions as a substrate docking protein for signaling by DNA-PK to homeodomain proteins from DNA ends. PMID- 11279129 TI - Arabidopsis thaliana flavoprotein AtHAL3a catalyzes the decarboxylation of 4' Phosphopantothenoylcysteine to 4'-phosphopantetheine, a key step in coenzyme A biosynthesis. AB - The Arabidopsis thaliana flavoprotein AtHAL3a is related to plant growth and salt and osmotic tolerance. AtHAL3a shows sequence homology to the bacterial flavoproteins EpiD and Dfp. EpiD, Dfp, and AtHAL3a are members of the homo oligomeric flavin-containing Cys decarboxylase (HFCD) protein family. We demonstrate that AtHAL3a catalyzes the decarboxylation of (R)-4'-phospho-N pantothenoylcysteine to 4'-phosphopantetheine. This key step in coenzyme A biosynthesis is catalyzed in bacteria by the Dfp proteins. Exchange of His-90 of AtHAL3a for Asn led to complete inactivation of the enzyme. Dfp and AtHAL3a are characterized by a shortened substrate binding clamp compared with EpiD. Exchange of the cysteine residue of the conserved ACGD motif of this binding clamp resulted in loss of (R)-4'-phospho-N-pantothenoylcysteine decarboxylase activity. Based on the crystal structures of EpiD H67N with bound substrate peptide and of AtHAL3a, we present a model for the binding of (R)-4'-phospho-N pantothenoylcysteine to AtHAL3a. PMID- 11279130 TI - "Weak toxin" from Naja kaouthia is a nontoxic antagonist of alpha 7 and muscle type nicotinic acetylcholine receptors. AB - A novel "weak toxin" (WTX) from Naja kaouthia snake venom competes with [(125)I]alpha-bungarotoxin for binding to the membrane-bound Torpedo californica acetylcholine receptor (AChR), with an IC(50) of approximately 2.2 microm. In this respect, it is approximately 300 times less potent than neurotoxin II from Naja oxiana and alpha-cobratoxin from N. kaouthia, representing short-type and long-type alpha-neurotoxins, respectively. WTX and alpha-cobratoxin displaced [(125)I]alpha-bungarotoxin from the Escherichia coli-expressed fusion protein containing the rat alpha7 AChR N-terminal domain 1-208 preceded by glutathione S transferase with IC(50) values of 4.3 and 9.1 microm, respectively, whereas for neurotoxin II the IC(50) value was >100 microm. Micromolar concentrations of WTX inhibited acetylcholine-activated currents in Xenopus oocyte-expressed rat muscle AChR and human and rat alpha7 AChRs, inhibiting the latter most efficiently (IC(50) of approximately 8.3 microm). Thus, a virtually nontoxic "three-fingered" protein WTX, although differing from alpha-neurotoxins by an additional disulfide in the N-terminal loop, can be classified as a weak alpha-neurotoxin. It differs from the short chain alpha-neurotoxins, which potently block the muscle-type but not the alpha7 AChRs, and is closer to the long alpha-neurotoxins, which have comparable potency against the above-mentioned AChR types. PMID- 11279131 TI - The beta-amyloid precursor protein APP is tyrosine-phosphorylated in cells expressing a constitutively active form of the Abl protoncogene. AB - The cytosolic domain of the beta-amyloid precursor protein APP interacts with three PTB (phosphotyrosine binding domain)-containing adaptor proteins, Fe65, X11, and mDab1. Through these adaptors, other molecules can be recruited at the cytodomain of APP; one of them is Mena, that binds to the WW domain (a protein module with two conserved tryptophans) of Fe65. The enabled and disabled genes of Drosophila, homologues of the mammalian Mena and mDab1 genes, respectively, are genetic modulators of the phenotype observed in flies null for the Abl tyrosine kinase gene. The involvement of Mena and mDab1 in the APP-centered protein protein interaction network suggests the possibility that Abl plays a role in APP biology. We show that Fe65, through its WW domain, binds in vitro and in vivo the active form of Abl. Furthermore, in cells expressing the active form of Abl, APP is tyrosine-phosphorylated. Phosphopeptide analysis and site-directed mutagenesis support the hypothesis that Tyr(682) of APP(695) is the target of this phosphorylation. Co-immunoprecipitation experiments demonstrate that active Abl and tyrosine-phosphorylated APP also form a stable complex, which could result from the interaction of the pYENP motif of the APP cytodomain with the SH2 domain of Abl. These results suggest that Abl, Mena, and mDab1 are involved in a common molecular machinery and that APP can play a role in tyrosine kinase-mediated signaling. PMID- 11279132 TI - Linear relationships between the ligand binding energy and the activation energy of time-dependent inhibition of steroid 5alpha-reductase by delta 1-4 azasteroids. AB - The inhibition of steroid 5alpha-reductase (5AR) by Delta(1)-4-azasteroids is characterized by a two-step time-dependent kinetic mechanism where inhibitor combines with enzyme in a fast equilibrium, defined by the inhibition constant K(i), to form an initial reversible enzyme-inhibitor complex, which subsequently undergoes a time-dependent chemical rearrangement, defined by the rate constant k(3), leading to the formation of an apparently irreversible, tight-binding enzyme-inhibitor complex (Tian, G., Mook, R. A., Jr., Moss, M. L., and Frye, S. V. (1995) Biochemistry 34, 13453-13459). A detailed kinetic analysis of this process with a series of Delta(1)-4-azasteroids having different C-17 substituents was performed to understand the relationships between the rate of time-dependent inhibition and the affinity of the time-dependent inhibitors for the enzyme. A linear correlation was observed between ln(1/K(i)), which is proportional to the ligand binding energy for the formation of the enzyme inhibitor complex, and ln(1/(k(3)/K(i))), which is proportional to the activation energy for the inhibition reaction under the second order reaction condition, which leads to the formation of the irreversible, tight-binding enzyme-inhibitor complex. The coefficient of the correlation was -0.88 +/- 0.07 for type 1 5AR and -1.0 +/- 0.2 for type 2 5AR. In comparison, there was no obvious correlation between ln(1/K(i)) and ln(1/k(3)), which is proportional to the activation energy of the second, time-dependent step of the inhibition reaction. These data are consistent with a model where ligand binding energies provided at C-17 of Delta(1)-4-azasteroids is fully expressed to lower the activation energy of k(3)/K(i) with little perturbation of the energy barrier of the second, time dependent step. PMID- 11279133 TI - Cyclin D1 represses STAT3 activation through a Cdk4-independent mechanism. AB - STAT3 transcription factors are cytoplasmic proteins that induce gene activation in response to cytokine receptor stimulation. Following tyrosine phosphorylation, STAT3 proteins dimerize, translocate into the nucleus, and activate specific target genes. Activation is transient, and down-regulation of STAT3 signaling occurs within a few hours. In this study, we show that cyclin D1 inhibits STAT3 activation. In co-immunoprecipitation and pull-down assays, cyclin D1 was found to associate with the activation domain of STAT3 upon interleukin-6 stimulation. Overexpression of cyclin D1 inhibited transcriptional activation by STAT3 proteins. This effect was not shared by cyclin E, was independent of association with Cdk4, and was unaffected by inhibitors of Cdk4. Whereas cyclin D1 had no effect on the steady-state level of STAT3 proteins in the cytoplasm, it was found to reduce the STAT3 nuclear level in HepG2 cells. These results suggest a model by which cyclin D1 is part of a feedback network controlling the down-regulation of STAT3 activity and highlight a new activity for this cell cycle regulatory protein. PMID- 11279134 TI - A role for C/EBPbeta in regulating peroxisome proliferator-activated receptor gamma activity during adipogenesis in 3T3-L1 preadipocytes. AB - The differentiation of 3T3-L1 preadipocytes is regulated in part by a cascade of transcriptional events involving activation of the CCAAT/enhancer-binding proteins (C/EBPs) and peroxisome proliferator-activated receptor gamma (PPARgamma) by dexamethasone (DEX), 3-isobutyl-1-methylxanthine (MIX), and insulin. In this study, we demonstrate that exposure of 3T3-L1 preadipocytes to DEX and insulin fails to induce adipogenesis as indicated by a lack of C/EBPalpha, PPARgamma2, and adipose protein 2/fatty acid-binding protein expression; however, PPARgamma1 is expressed. Treatment of these MIX-deficient cells with a PPARgamma ligand, troglitazone, induces C/EBPalpha expression and rescues the block in adipogenesis. In this regard, we also show that induction of C/EBPalpha gene expression by troglitazone in C3H10T1/2 cells ectopically expressing PPARgamma occurs in the absence of ongoing protein synthesis, suggesting a direct transactivation of the C/EBPalpha gene by PPARgamma. Furthermore, ectopic expression of a dominant negative isoform of C/EBPbeta (liver-enriched transcriptional inhibitory protein (LIP)) inhibits the induction of C/EBPalpha, PPARgamma2, and adipose protein 2/fatty acid-binding protein by DEX, MIX, and insulin in 3T3-L1 cells without affecting the induction of PPARgamma1 by DEX. Exposure of LIP-expressing preadipocytes to troglitazone along with DEX, MIX, and insulin induces differentiation into adipocytes. Additionally, we show that sustained expression of C/EBPalpha in these LIP-expressing adipocytes requires constant exposure to troglitazone. Taken together, these observations suggest that inhibition of C/EBPbeta activity not only blocks C/EBPalpha and PPARgamma2 expression, but it also renders the preadipocytes dependent on an exogenous PPARgamma ligand for their differentiation into adipocytes. We propose, therefore, an additional role for C/EBPbeta in regulating PPARgamma activity during adipogenesis, and we suggest an alternative means of inducing preadipocyte differentiation that relies on the dexamethasone-associated induction of PPARgamma1 expression. PMID- 11279135 TI - Direct acetylation of the estrogen receptor alpha hinge region by p300 regulates transactivation and hormone sensitivity. AB - Regulation of nuclear receptor gene expression involves dynamic and coordinated interactions with histone acetyl transferase (HAT) and deacetylase complexes. The estrogen receptor (ERalpha) contains two transactivation domains regulating ligand-independent and -dependent gene transcription (AF-1 and AF-2 (activation functions 1 and 2)). ERalpha-regulated gene expression involves interactions with cointegrators (e.g. p300/CBP, P/CAF) that have the capacity to modify core histone acetyl groups. Here we show that the ERalpha is acetylated in vivo. p300, but not P/CAF, selectively and directly acetylated the ERalpha at lysine residues within the ERalpha hinge/ligand binding domain. Substitution of these residues with charged or polar residues dramatically enhanced ERalpha hormone sensitivity without affecting induction by MAPK signaling, suggesting that direct ERalpha acetylation normally suppresses ligand sensitivity. These ERalpha lysine residues also regulated transcriptional activation by histone deacetylase inhibitors and p300. The conservation of the ERalpha acetylation motif in a phylogenetic subset of nuclear receptors suggests that direct acetylation of nuclear receptors may contribute to additional signaling pathways involved in metabolism and development. PMID- 11279136 TI - Interferon-gamma sensitizes human myeloid leukemia cells to death receptor mediated apoptosis by a pleiotropic mechanism. AB - The role of interferon (IFN)-gamma as a sensitizing agent in apoptosis induced by ligation of death receptors has been evaluated in human myeloid leukemia cells. Incubation of U937 cells with IFN-gamma sensitized these cells to apoptosis induced by tumor necrosis factor-alpha, agonistic CD95 antibody, and tumor necrosis factor-related apoptosis-inducing ligand. Other human myeloid leukemic cells were also sensitized by IFN-gamma to death receptor-mediated apoptosis. Treatment of U937 cells with IFN-gamma up-regulated the expression of caspase-8 and potently synergized with death receptor ligation in the processing of caspase 8 and BID cleavage. Concomitantly, a marked down-regulation of BCL-2 protein was also observed in cells incubated with IFN-gamma. Furthermore, the caspase dependent generation of a 23-kDa fragment of BCL-2 protein, the release of cytochrome c from mitochondria and the activation of caspase-9 were also enhanced upon death receptor ligation in IFN-gamma-treated cells. Ectopically expressed Bcl-2 protein inhibited IFN-gamma-induced sensitization to apoptosis. In summary, these results indicate that IFN-gamma sensitizes human myeloid leukemic cells to a death receptor-induced, mitochondria-mediated pathway of apoptosis. PMID- 11279138 TI - The conformation of the glucocorticoid receptor af1/tau1 domain induced by osmolyte binds co-regulatory proteins. AB - The activation domains of many transcription factors appear to exist naturally in an unfolded or only partially folded state. This seems to be the case for AF1/tau1, the major transactivation domain of the human glucocorticoid receptor. We show here that in buffers containing the natural osmolyte trimethylamine N oxide (TMAO), recombinant AF1 folds into more a compact structure, as evidenced by altered fluorescence emission, circular dichroism spectra, and ultracentrifugal analysis. This conformational transition is cooperative, a characteristic of proteins folding to natural structures. The structure resulting from incubation in TMAO causes the peptide to resist proteolysis by trypsin, chymotrypsin, endoproteinase Arg-C and endoproteinase Gluc-C. Ultracentrifugation studies indicate that AF1/tau1 exists as a monomer in aqueous solution and that the presence of TMAO does not lead to oligomerization or aggregation. It has been suggested that recombinant AF1 binds both the ubiquitous coactivator CBP and the TATA box-binding protein, TBP. Interactions with both of these are greatly enhanced in the presence of TMAO. Co-immunoadsorption experiments indicate that in TMAO each of these and the coactivator SRC-1 are found complexed with AF1. These data indicate that TMAO induces a conformation in AF1/tau1 that is important for its interaction with certain co-regulatory proteins. PMID- 11279137 TI - A constitutive cytoprotective pathway protects endothelial cells from lipopolysaccharide-induced apoptosis. AB - Lipopolysaccharide (LPS) has been implicated as the bacterial component responsible for much of the endothelial cell injury/dysfunction associated with Gram-negative bacterial infections. Protein synthesis inhibition is required to sensitize the endothelium to lipopolysaccharide-induced apoptosis, suggesting that a constitutive or inducible cytoprotective protein(s) is required for endothelial survival. We have identified two known endothelial anti-apoptotic proteins, c-FLIP and Mcl-1, the expression of which is decreased markedly in the presence of cycloheximide. Decreased expression of both proteins preceded apoptosis evoked by lipopolysaccharide + cycloheximide. Caspase inhibition protected against apoptosis, but not the decreased expression of c-FLIP and Mcl 1, suggesting that they exert protection upstream of caspase activation. Inhibition of the degradation of these two proteins with the proteasome inhibitor, lactacystin, prevented lipopolysaccharide + cycloheximide-induced apoptosis. Similarly, lactacystin protected against endothelial apoptosis induced by either tumor necrosis factor-alpha or interleukin-1beta in the presence of cycloheximide. That apoptosis could be blocked in the absence of new protein synthesis by inhibition of the proteasome degradative pathway implicates the requisite involvement of a constitutively expressed protein(s) in the endothelial cytoprotective pathway. Finally, reduction of FLIP expression with antisense oligonucleotides sensitized endothelial cells to LPS killing, demonstrating a definitive role for FLIP in the protection of endothelial cells from LPS-induced apoptosis. PMID- 11279139 TI - Directed mutagenesis of apecific active site residues on Fibrobacter succinogenes 1,3-1,4-beta -D-glucanase significantly affects catalysis and enzyme structural stability. AB - The functional and structural significance of amino acid residues Met(39), Glu(56), Asp(58), Glu(60), and Gly(63) of Fibrobacter succinogenes 1,3-1,4-beta-d glucanase was explored by the approach of site-directed mutagenesis, initial rate kinetics, fluorescence spectroscopy, and CD spectrometry. Glu(56), Asp(58), Glu(60), and Gly(63) residues are conserved among known primary sequences of the bacterial and fungal enzymes. Kinetic analyses revealed that 240-, 540-, 570-, and 880-fold decreases in k(cat) were observed for the E56D, E60D, D58N, and D58E mutant enzymes, respectively, with a similar substrate affinity relative to the wild type enzyme. In contrast, no detectable enzymatic activity was observed for the E56A, E56Q, D58A, E60A, and E60Q mutants. These results indicated that the carboxyl side chain at positions 56 and 60 is mandatory for enzyme catalysis. M39F, unlike the other mutants, exhibited a 5-fold increase in K(m) value. Lower thermostability was found with the G63A mutant when compared with wild type or other mutant forms of F. succinogenes 1,3-1,4-beta-d-glucanase. Denatured wild type and mutant enzymes were, however, recoverable as active enzymes when 8 m urea was employed as the denaturant. Structural modeling and kinetic studies suggest that Glu(56), Asp(58), and Glu(60) residues apparently play important role(s) in the catalysis of F. succinogenes 1,3-1,4-beta-d-glucanase. PMID- 11279141 TI - Novel NEMO/IkappaB kinase and NF-kappa B target genes at the pre-B to immature B cell transition. AB - The IkappaB kinase (IKK) signaling complex is responsible for activating NF kappaB-dependent gene expression programs. Even though NF-kappaB-responsive genes are known to orchestrate stress-like responses, critical gaps in our knowledge remain about the global effects of NF-kappaB activation on cellular physiology. DNA microarrays were used to compare gene expression programs in a model system of 70Z/3 murine pre-B cells versus their IKK signaling-defective 1.3E2 variant with lipopolysaccharide (LPS), interleukin-1 (IL-1), or a combination of LPS + phorbol 12-myristate 13-acetate under brief (2 h) or long term (12 h) stimulation. 70Z/3-1.3E2 cells lack expression of NEMO/IKKgamma/IKKAP-1/FIP-3, an essential positive effector of the IKK complex. Some stimulated hits were known NF-kappaB target genes, but remarkably, the vast majority of the up-modulated genes and an unexpected class of repressed genes were all novel targets of this signaling pathway, encoding transcription factors, receptors, extracellular ligands, and intracellular signaling factors. Thirteen stimulated (B-ATF, Pim-2, MyD118, Pea-15/MAT1, CD82, CD40L, Wnt10a, Notch 1, R-ras, Rgs-16, PAC-1, ISG15, and CD36) and five repressed (CCR2, VpreB, lambda5, SLPI, and CMAP/Cystatin7) genes, respectively, were bona fide NF-kappaB targets by virtue of their response to a transdominant IkappaBalphaSR (super repressor). MyD118 and ISG15, although directly induced by LPS stimulation, were unaffected by IL-1, revealing the existence of direct NF-kappaB target genes, which are not co-induced by the LPS and IL-1 Toll-like receptors. PMID- 11279140 TI - Pkd1 unusual DNA conformations are recognized by nucleotide excision repair. AB - The 2.5-kilobase pair poly(purine.pyrimidine) (poly(R.Y)) tract present in intron 21 of the polycystic kidney disease 1 (PKD1) gene has been proposed to contribute to the high mutation frequency of the gene. To evaluate this hypothesis, we investigated the growth rates of 11 Escherichia coli strains, with mutations in the nucleotide excision repair, SOS, and topoisomerase I and/or gyrase genes, harboring plasmids containing the full-length tract, six 5'-truncations of the tract, and a control plasmid (pSPL3). The full-length poly(R.Y) tract induced dramatic losses of cell viability during the first few hours of growth and lengthened the doubling times of the populations in strains with an inducible SOS response. The extent of cell loss was correlated with the length of the poly(R.Y) tract and the levels of negative supercoiling as modulated by the genotype of the strains or drugs that specifically inhibited DNA gyrase or bound to DNA directly, thereby affecting conformations at specific loci. We conclude that the unusual DNA conformations formed by the PKD1 poly(R.Y) tract under the influence of negative supercoiling induced the SOS response pathway, and they were recognized as lesions by the nucleotide excision repair system and were cleaved, causing delays in cell division and loss of the plasmid. These data support a role for this sequence in the mutation of the PKD1 gene by stimulating repair and/or recombination functions. PMID- 11279142 TI - ATP induces a conformational change in lipid-bound cytochrome c. AB - Resonance energy transfer studies using a pyrene-labeled phospholipid derivative 1-palmitoyl-2-[10-(pyren-1-yl)decanoyl]-sn-glycero-3-phosphoglycerol (donor) and the heme (acceptor) of cytochrome c (cyt c) have indicated that ATP causes changes in the conformation of the lipid-bound protein (Rytomaa, M., Mustonen, P., and Kinnunen, P. K. J. (1992) J. Biol. Chem. 267, 22243-22248). Accordingly, after binding cyt c via its so called C-site to neat phosphatidylglycerol liposomes (mole fraction of PG = 1.0) has commenced, further quenching of donor fluorescence is caused by ATP, saturating at 2 mm nucleotide. ATP-induced conformational changes in liposome-associated cyt c could be directly demonstrated by CD in the Soret band region (380-460 nm). The latter data were further supported by time-resolved spectroscopy using the fluorescent cyt c analog with a Zn(2+)-substituted heme moiety. A high affinity ATP-binding site has been demonstrated in cyt c (Craig, D. B., and Wallace, C. J. A. (1993) Protein Sci. 2, 966-976) that is compromised by replacing the invariant Arg(91) to norleucine. Although no major effects on conformation and function of cyt c were concluded due to the modification, a significantly reduced effect by ATP on the lipid-bound [Nle(91)]cyt c was evident, implying that this modulation is mediated via the Arg(91)-containing binding site. PMID- 11279143 TI - Centrosome protein centrin 2/caltractin 1 is part of the xeroderma pigmentosum group C complex that initiates global genome nucleotide excision repair. AB - Nucleotide excision repair (NER) is carried out by xeroderma pigmentosum (XP) factors. Before the excision reaction, DNA damage is recognized by a complex originally thought to contain the XP group C responsible gene product (XPC) and the human homologue of Rad23 B (HR23B). Here, we show that centrin 2/caltractin 1 (CEN2) is also a component of the XPC repair complex. We demonstrate that nearly all XPC complexes contain CEN2, that CEN2 interacts directly with XPC, and that CEN2, in cooperation with HR23B, stabilizes XPC, which stimulates XPC NER activity in vitro. CEN2 has been shown to play an important role in centrosome duplication. Thus, those findings suggest that the XPC-CEN2 interaction may reflect coupling of cell division and NER. PMID- 11279144 TI - FKBP12 binding modulates ryanodine receptor channel gating. AB - The ryanodine receptor (RyR1)/calcium release channel on the sarcoplasmic reticulum of skeletal muscle is comprised of four 565,000-dalton RyR1s, each of which binds one FK506 binding protein (FKBP12). RyR1 is required for excitation contraction coupling in skeletal muscle. FKBP12, a cis-trans peptidyl-prolyl isomerase, is required for the normal gating of the RyR1 channel. In the absence of FKBP12, RyR1 channels exhibit increased gating frequency, suggesting that FKBP12 "stabilizes" the channel in the open and closed states. We now show that substitution of a Gly, Glu, or Ile for Val2461 in RyR1 prevents FKBP12 binding to RyR1, resulting in channels with increased gating frequency. In the case of the V2461I mutant RyR1, normal channel function can be restored by adding FKBP12.6, an isoform of FKBP12. These data identify Val2461 as a critical residue required for FKBP12 binding to RyR1 and demonstrate the functional role for FKBP12 in the RyR1 channel complex. PMID- 11279146 TI - Positive regulatory domain I binding factor 1 silences class II transactivator expression in multiple myeloma cells. AB - The major histocompatibility complex (MHC) class II transactivator (CIITA) acts as a master switch to activate expression of the genes required for MHC-II antigen presentation. During B-cell to plasma cell differentiation, MHC-II expression is actively silenced, but the mechanism has been unknown. In plasma cell tumors such as multiple myeloma the repression of MHC-II is associated with the loss of CIITA. We have identified that positive regulatory domain I binding factor 1 (PRDI-BF1), a transcriptional repressor, inhibits CIITA expression in multiple myeloma cell lines. Repression of CIITA depends on the DNA binding activity of PRDI-BF1 and its specific binding site in the CIITA promoter. Deletion of a histone deacetylase recruitment domain in PRDI-BF1 does not inhibit repression of CIITA nor does blocking histone deacetylase activity. This is in contrast to PRDI-BF1 repression of the c-myc promoter. Repression of CIITA requires either the N-terminal acidic and conserved PR motif or the proline-rich domain. PRDI-BF1 has been shown to be a key regulator of B-cell and macrophage differentiation. These findings now indicate that PRDI-BF1 has at least two mechanisms of repression whose function is dependent on the nature of the target promoter. Importantly, PRDI-BF1 is defined as the key molecule in silencing CIITA and thus MHC-II in multiple myeloma cells. PMID- 11279145 TI - Exploring the acceptor substrate recognition of the human beta-galactoside alpha 2,6-sialyltransferase. AB - Human beta1,4-galactoside alpha2,6-sialyltransferase I (ST6GalI) recognition of glycoprotein acceptors has been investigated using various soluble forms of the enzyme deleted to a variable extent in the N-terminal half of the polypeptide. Full-length and truncated forms of the enzyme have been investigated with respect to their specificity for a variety of desialylated glycoproteins of known complex glycans as well as related proteins with different carbohydrate chains. Differences in transfer efficiency have been observed between membrane and soluble enzymatic forms, indicating that deletion of the transmembrane fragment induces loss of acceptor preference. No difference in substrate recognition could be observed when soluble enzymes of similar peptide sequence were produced in yeast or mammalian cells, confirming that removal of the membrane anchor and heterologous expression do not alter enzyme folding and activity. When tested on free oligosaccharides, soluble ST6GalI displayed full ability to sialylate free N glycans as well as various N-acetyllactosaminyl substrates. Progressive truncation of the N terminus demonstrated that the catalytic domain can proceed with sialic acid transfer with increased efficiency until 80 amino acids are deleted. Fusion of the ST6GalI catalytic domain to the N-terminal half of an unrelated transferase (core 2 beta1,6-N-acetylglucosaminyltransferase) further showed that a chimeric form of broad acceptor specificity and high activity could also be engineered in vivo. These findings therefore delineate a peptide region of approximately 50 amino acids within the ST6GalI stem region that governs both the preference for glycoprotein acceptors and catalytic activity, thereby suggesting that it may exert a steric control on the catalytic domain. PMID- 11279147 TI - Chorismate synthase from the hyperthermophile Thermotoga maritima combines thermostability and increased rigidity with catalytic and spectral properties similar to mesophilic counterparts. AB - Chorismate synthase, the last enzyme in the shikimate pathway, catalyzes the transformation of 5-enolpyruvylshikimate 3-phosphate to chorismate, a biochemically unique reaction in that it requires reduced FMN as a cofactor. Here we report on the cloning, expression, and characterization of the protein for the first time from an extremophilic organism Thermotoga maritima which is also one of the oldest and most slowly evolving eubacteria. The protein is monofunctional in that it does not have an intrinsic ability to reduce the FMN cofactor and thereby reflecting the nature of the ancestral enzyme. Circular dichroism studies indicate that the melting temperature of the T. maritima protein is above 92 degrees C compared with 54 degrees C for the homologous Escherichia coli protein while analytical ultracentrifugation showed that both proteins have the same quaternary structure. Interestingly, UV-visible spectral studies revealed that the dissociation constants for both oxidized FMN and 5-enolpyruvylshikimate 3 phosphate decrease 46- and 10-fold, respectively, upon heat treatment of the T. maritima protein. The heat treatment also results in the trapping of the flavin cofactor in an apolar environment, a feature which is enhanced by the presence of the substrate 5-enolpyruvylshikimate 3-phosphate. Nevertheless, stopped-flow spectrophotometric evidence suggests that the mechanism of the T. maritima protein is similar to that of the E. coli protein. In essence, the study shows that T. maritima chorismate synthase exhibits considerably higher rigidity and thermostability while it has conserved features relevant to its catalytic function. PMID- 11279148 TI - Interaction between the Btk PH domain and phosphatidylinositol-3,4,5 trisphosphate directly regulates Btk. AB - Bruton's tyrosine kinase (Btk) binds to phosphatidylinositol-3,4,5-trisphosphate (PtdIns-3,4,5-P(3)) through the Btk pleckstrin homology (PH) domain, an interaction thought to be required for Btk membrane translocation during B cell receptor signaling. Here, we report that interaction of PtdIns-3,4,5-P(3) with the PH domain of Btk directly induces Btk enzymatic activation in an in vitro kinase assay. A point mutation that reduces interaction of PtdIns-3,4,5-P(3) with the Btk PH domain blocks in vitro PtdIns-3,4,5-P(3)-dependent Btk activation, whereas the PH domain deletion enhances Btk basal activity but eliminates the PtdIns-3,4,5-P(3)-dependent stimulation. Btk kinase activity and the Btk activation loop phosphorylation site are both required for the PtdIns-3,4,5-P(3) mediated stimulation of Btk kinase activity. Together, these results suggest that the Btk PH domain is positioned such that it normally suppresses both Btk kinase activity and access to substrates; when interacting with PtdIns-3,4,5-P(3), this suppression is relieved, producing apparent Btk activation. In addition, using Src family kinase inhibitors and Btk catalytically inactive mutants, we demonstrate that in vivo, the activation of Btk is due to both Lyn phosphorylation and PtdIns-3,4,5-P(3)-mediated direct activation. Thus, the Btk PtdIns-3,4,5-P(3) interaction serves to translocate Btk to the membrane and directly regulate its signaling function. PMID- 11279149 TI - Oxidized alkyl phospholipids are specific, high affinity peroxisome proliferator activated receptor gamma ligands and agonists. AB - Synthetic high affinity peroxisome proliferator-activated receptor (PPAR) agonists are known, but biologic ligands are of low affinity. Oxidized low density lipoprotein (oxLDL) is inflammatory and signals through PPARs. We showed, by phospholipase A(1) digestion, that PPARgamma agonists in oxLDL arise from the small pool of alkyl phosphatidylcholines in LDL. We identified an abundant oxidatively fragmented alkyl phospholipid in oxLDL, hexadecyl azelaoyl phosphatidylcholine (azPC), as a high affinity ligand and agonist for PPARgamma. [(3)H]azPC bound recombinant PPARgamma with an affinity (K(d)((app)) approximately 40 nm) that was equivalent to rosiglitazone (BRL49653), and competition with rosiglitazone showed that binding occurred in the ligand-binding pocket. azPC induced PPRE reporter gene expression, as did rosiglitazone, with a half-maximal effect at 100 nm. Overexpression of PPARalpha or PPARgamma revealed that azPC was a specific PPARgamma agonist. The scavenger receptor CD36 is encoded by a PPRE-responsive gene, and azPC enhanced expression of CD36 in primary human monocytes. We found that anti-CD36 inhibited azPC uptake, and it inhibited PPRE reporter induction. Results with a small molecule phospholipid flippase mimetic suggest azPC acts intracellularly and that cellular azPC accumulation was efficient. Thus, certain alkyl phospholipid oxidation products in oxLDL are specific, high affinity extracellular ligands and agonists for PPARgamma that induce PPAR-responsive genes. PMID- 11279150 TI - Cloning, Golgi localization, and enzyme activity of the full-length heparin/heparan sulfate-glucuronic acid C5-epimerase. AB - While studying the cellular localization and activity of enzymes involved in heparan sulfate biosynthesis, we discovered that the published sequence for the glucuronic acid C5-epimerase responsible for the interconversion of d-glucuronic acid and l-iduronic acid residues encodes a truncated protein. Genome analysis and 5'-rapid amplification of cDNA ends was used to clone the full-length cDNA from a mouse mastocytoma cell line. The extended cDNA encodes for an additional 174 amino acids at the amino terminus of the protein. The murine sequence is 95% identical to the human epimerase identified from genomic sequences and fits with the general size and structure of the gene from Drosophila melanogaster and Caenorhabditis elegans. Full-length epimerase is predicted to have a type II transmembrane topology with a 17-amino acid transmembrane domain and an 11-amino acid cytoplasmic tail. An assay with increased sensitivity was devised that detects enzyme activity in extracts prepared from cultured cells and in recombinant proteins. Unlike other enzymes involved in glycosaminoglycan biosynthesis, the addition of a c-myc tag or green fluorescent protein to the highly conserved COOH-terminal portion of the protein inhibits its activity. The amino-terminally truncated epimerase does not localize to any cellular compartment, whereas the full-length enzyme is in the Golgi, where heparan sulfate synthesis is thought to occur. PMID- 11279151 TI - Substrate hydrolysis by matrix metalloproteinase-9. AB - The catalytic clefts of all matrix metalloproteinases (MMPs) have a similar architecture, raising questions about the redundancy in substrate recognition across the protein family. In the present study, an unbiased phage display strategy was applied to define the substrate recognition profile of MMP-9. Three groups of substrates were identified, each occupying a distinct set of subsites within the catalytic pocket. The most prevalent motif contains the sequence Pro-X X-Hy-(Ser/Thr) at P(3) through P(2'). This sequence is similar to the MMP cleavage sites within the collagens and is homologous to substrates the have been selected for other MMPs. Despite this similarity, most of the substrates identified here are selective for MMP-9 over MMP-7 and MMP-13. This observation indicates that substrate selectivity is conferred by key subsite interactions at positions other than P(3) and P(1'). This study shows that MMP-9 has a unique preference for Arg at both P(2) and P(1), and a preference for Ser/Thr at P(2'). Substrates containing the consensus MMP-9 recognition motif were used to query the protein data bases. A surprisingly limited list of putative physiologic substrates was identified. The functional implications of these proteins lead to testable hypotheses regarding physiologic substrates for MMP-9. PMID- 11279152 TI - Binding of steroidogenic acute regulatory protein to synthetic membranes suggests an active molten globule. AB - Steroidogenic acute regulatory protein (StAR) mediates cholesterol transport from the outer to the inner mitochondrial membrane during steroid biosynthesis. The mechanism of StAR's action is not established. To address mechanistic issues, we assessed the binding of StAR to artificial membranes by fluorescence resonance energy transfer using endogenous StAR tryptophan residues as the donor and dansyl phosphatidylethanolamine in the bilayer as the acceptor. Mixing StAR with dansyl labeled vesicles composed of phosphatidylcholine increased the fluorescence intensity of dansyl emission excited at 280 nm by 10-40%. This interaction was dependent on pH, with a maximum at pH 3.0-3.5 and essentially no change above pH 5. Binding experiments at different temperatures and various combinations of phosphatidylcholine, phosphatidylglycerol, cardiolipin, and cholesterol showed that binding involves an electrostatic step and one or more other steps. Although binding prefers a thermodynamically ordered bilayer, the rate-limiting step occurs either when the bilayer is in a fluid state or when there is cholesterol induced membrane heterogeneity. Experiments with fluorescence and light scattering indicate that StAR binding promotes ordering and aggregation of anionic membranes. The inactive StAR mutant R182L had lower affinity for the membrane, and the partially active mutant L275P had intermediate affinity. Far-UV CD spectroscopy of StAR in PC membranes show more beta-structure than in aqueous buffers, and the presence of cardiolipin or cholesterol in the membrane fosters a molten globule state. Our data suggest that StAR binds to membranes in a partially unfolded molten globule state that is relevant to the activity of the protein. PMID- 11279153 TI - Accumulation of 55Fe-labeled precursors of the iron-molybdenum cofactor of nitrogenase on NifH and NifX of Azotobacter vinelandii. AB - Iron-molybdenum cofactor (FeMo-co) biosynthesis involves the participation of several proteins. We have used (55)Fe-labeled NifB-co, the specific iron and sulfur donor to FeMo-co, to investigate the accumulation of protein-bound precursors of FeMo-co. The (55)Fe label from radiolabeled NifB-co became associated with two major protein bands when the in vitro FeMo-co synthesis reaction was carried out with the extract of an Azotobacter vinelandii mutant lacking apodinitrogenase. One of the bands, termed (55)Fe-labeled upper band, was purified and shown to be NifH by immunoblot analysis. The (55)Fe-labeled lower band was identified as NifX by N-terminal sequencing. NifX purified from an A. vinelandii nifB strain showed a different electrophoretic mobility on anoxic native gels than did NifX with the FeMo-co precursor bound. PMID- 11279154 TI - Role of intronic E- and N-box motifs in the transcriptional induction of the acetylcholinesterase gene during myogenic differentiation. AB - In this study, we examined whether an intronic N-box motif is involved in the expression of acetylcholinesterase (AChE) during myogenesis. We determined that AChE transcripts are barely detectable in cultured myoblasts and that their levels increase dramatically in myotubes. Nuclear run-on assays revealed that this increase was accompanied by a parallel induction in the transcriptional activity of the AChE gene. These changes in transcription were also observed in transfection experiments using AChE promoter-reporter gene constructs. Mutation of the intronic N-box at position +755 base pairs (bp) reduced by more than 70% expression of the reporter gene in myotubes. Disruption of an adjacent E-box, at position +767 bp, also reduced expression of the reporter gene following myogenic differentiation. Co-transfection experiments using AChE promoter-reporter gene constructs and a myogenin expression vector showed that expression of this regulatory factor increased expression of the reporter gene in myotubes. Although the AChE promoter contains multiple E-boxes, mutation of this intronic one was sufficient to prevent the myogenin-induced increase in reporter gene expression. Together, these results indicate that changes in AChE gene transcription occur during myogenesis and highlight the contribution of the intronic N- and E-box motifs in the developmental regulation of the AChE gene in skeletal muscle. PMID- 11279155 TI - Epidermal growth factor (EGF) receptor kinase-independent signaling by EGF. AB - The ErbB family of receptors, which includes the epidermal growth factor receptor (EGFR), ErbB2, ErbB3, and ErbB4, mediate signaling by EGF-like polypeptides. To better understand the role of the EGFR tyrosine kinase, we analyzed signaling by a kinase-inactive EGFR (K721M) in ErbB-devoid 32D cells. K721M alone exhibited no detectable signaling capacity, whereas coexpression of K721M with ErbB2, but not ErbB3 or ErbB4, resulted in EGF-dependent mitogen-activated protein kinase (MAPK) activation. The kinase activity, but not tyrosine phosphorylation, of ErbB2 was required for EGF-induced MAPK activation. The presence of tyrosine phosphorylation sites in K721M was not a requisite for signaling, indicating that transphosphorylation of K721M by ErbB2 was not an essential mechanism of receptor activation. Conversely, the mutated kinase domain of K721M (residues 648-973) and tyrosine phosphorylation of at least one of the receptors were necessary. EGF was found to activate the pro-survival protein kinase Akt in stable cell lines expressing K721M and ErbB2 but, unlike cells expressing wild-type EGFR, was incapable of activating signal transducers and activators of transcription (STAT) or driving cell proliferation. These results demonstrate that EGFR-ErbB2 oligomers are potent activators of MAPK and Akt, and this signaling does not require EGFR kinase activity. PMID- 11279156 TI - Solution structure, backbone dynamics, and stability of a double mutant single chain monellin. structural origin of sweetness. AB - Single-chain monellin (SCM), which is an engineered 94-residue polypeptide, has been characterized as being as sweet as native two-chain monellin. Data from gel filtration high performance liquid chromatography and NMR has proven that SCM exists as a monomer in aqueous solution. In order to determine the structural origin of the taste of sweetness, we engineered several mutant SCM proteins by mutating Glu(2), Asp(7), and Arg(39) residues, which are responsible for sweetness. In this study, we present the solution structure, backbone dynamics, and stability of mutant SCM proteins using circular dichroism, fluorescence, and NMR spectroscopy. Based on the NMR data, a stable alpha-helix and five-stranded antiparallel beta-sheet were identified for double mutant SCM. Strands beta1 and beta2 are connected by a small bulge, and the disruption of the first beta-strand were observed with SCM(DR) comprising residues of Ile(38)-Cys(41). The dynamical and folding characteristics from circular dichroism, fluorescence, and backbone dynamics studies revealed that both wild type and mutant proteins showed distinct dynamical as well as stability differences, suggesting the important role of mutated residues in the sweet taste of SCM. Our results will provide an insight into the structural origin of sweet taste as well as the mutational effect in the stability of the engineered sweet protein SCM. PMID- 11279157 TI - p300-mediated acetylation of human transcriptional coactivator PC4 is inhibited by phosphorylation. AB - The human positive coactivator 4 (PC4) acts as a general coactivator for activator-dependent transcription, the activity of which is regulated negatively by phosphorylation. We report here that PC4 can be acetylated specifically by another coactivator, p300. Interestingly, phosphorylation of PC4 by casein kinase II inhibits the p300-mediated acetylation. Mass spectral analysis revealed that there are at least two lysine residues acetylated in PC4, as a result of which its DNA binding activity is stimulated. PMID- 11279158 TI - A defect in coenzyme Q biosynthesis is responsible for the respiratory deficiency in Saccharomyces cerevisiae abc1 mutants. AB - Ubiquinone (coenzyme Q or Q) is an essential component of the mitochondrial respiratory chain in eukaryotic cells. There are eight complementation groups of Q-deficient Saccharomyces cerevisiae mutants designated coq1-coq8. Here we report that COQ8 is ABC1 (for Activity of bc(1) complex), which was originally isolated as a multicopy suppressor of a cytochrome b mRNA translation defect (Bousquet, I., Dujardin, G., and Slonimski, P. P. (1991) EMBO J. 10, 2023-2031). Previous studies of abc1 mutants suggested that the mitochondrial respiratory complexes were thermosensitive and function inefficiently. Although initial characterization of the abc1 mutants revealed characteristics of Q-deficient mutants, levels of Q were reported to be similar to wild type. The suggested function of Abc1p was that it acts as a chaperone-like protein essential for the proper conformation and functioning of the bc(1) and its neighboring complexes (Brasseur, G., Tron, P., Dujardin, G., Slonimski, P. P. (1997) Eur. J. Biochem. 246, 103-111). Studies presented here indicate that abc1/coq8 null mutants are defective in Q biosynthesis and accumulate 3-hexaprenyl-4-hydroxybenzoic acid as the predominant intermediate. As observed in other yeast coq mutants, supplementation of growth media with Q(6) rescues the abc1/coq8 null mutants for growth on nonfermentable carbon sources. Such supplementation also partially restores succinate-cytochrome c reductase activity in the abc1/coq8 null mutants. Abc1/Coq8p localizes to the mitochondria, and is proteolytically processed upon import. The findings presented here indicate that the previously reported thermosensitivity of the respiratory complexes of abc1/coq8 mutants results from the lack of Q and a general deficiency in respiration, rather than a specific phenotype due to dysfunction of the Abc1 polypeptide. These results indicate that ABC1/COQ8 is essential for Q-biosynthesis and that the critical defect of abc1/coq8 mutants is a lack of Q. PMID- 11279159 TI - Estrogen receptor beta-selective transcriptional activity and recruitment of coregulators by phytoestrogens. AB - Estrogens used in hormone replacement therapy regimens may increase the risk of developing breast cancer. Paradoxically, high consumption of plant-derived phytoestrogens, particularly soybean isoflavones, is associated with a low incidence of breast cancer. To explore the molecular basis for these potential different clinical outcomes, we investigated whether soybean isoflavones elicit distinct transcriptional actions from estrogens. Our results demonstrate that the estrogen 17beta-estradiol effectively triggers the transcriptional activation and repression pathways with both estrogen receptors (ERs) ERalpha and ERbeta. In contrast, soybean isoflavones (genistein, daidzein, and biochanin A) are ERbeta selective agonists of transcriptional repression and activation at physiological levels. The molecular mechanism for ERbeta selectivity by isoflavones involves their capacity to create an activation function-2 surface of ERbeta that has a greater affinity for coregulators than ERalpha. Phytoestrogens may act as natural selective estrogen receptor modulators that elicit distinct clinical effects from estrogens used for hormone replacement by selectively recruiting coregulatory proteins to ERbeta that trigger transcriptional pathways. PMID- 11279160 TI - Biochemical characterization of the penta-EF-hand protein grancalcin and identification of L-plastin as a binding partner. AB - Grancalcin is a recently described Ca(2+)-binding protein especially abundant in human neutrophils. Grancalcin belongs to the penta-EF-hand subfamily of EF-hand proteins, which also comprises calpain, sorcin, peflin, and ALG-2. Penta-EF-hand members are typified by two novel types of EF-hands: one that binds Ca(2+) although it has an unusual Ca(2+) coordination loop and one that does not bind Ca(2+) but is directly involved in homodimerization. We have developed a novel method for purification of native grancalcin and found that the N terminus of wild-type grancalcin is acetylated. This posttranslational modification does not affect the secondary structure or conformation of the protein. We found that both native and recombinant grancalcin always exists as a homodimer, regardless of the Ca(2+) load. Flow dialysis showed that recombinant grancalcin binds two Ca(2+) per subunit with positive cooperativity and moderate affinity ([Ca(2+)](0.5) of 25 and 83 microm in the presence and absence of octyl glycoside, respectively) and that the sites are of the Ca(2+)-specific type. Furthermore, we showed, by several independent methods, that grancalcin undergoes important conformational changes upon binding of Ca(2+) and subsequently exposes hydrophobic amino acid residues, which direct the protein to hydrophobic surfaces. By affinity chromatography of solubilized human neutrophils on immobilized grancalcin, L plastin, a leukocyte-specific actin-bundling protein, was found to interact with grancalcin in a negative Ca(2+)-dependent manner. This was substantiated by co immunoprecipitation of grancalcin by anti-L-plastin antibodies and vice versa. PMID- 11279161 TI - Structure-function analysis of the active site tunnel of yeast RNA triphosphatase. AB - Cet1, the RNA triphosphatase component of the yeast mRNA capping apparatus, catalyzes metal-dependent gamma phosphate hydrolysis within the hydrophilic interior of a topologically closed 8-strand beta barrel (the "triphosphate tunnel"). We used structure-guided alanine scanning to identify 6 side chains within the triphosphate tunnel that are essential for phosphohydrolase activity in vitro and in vivo: Arg393, Glu433, Arg458, Arg469, Asp471 and Thr473. Alanine substitutions at two positions, Asp377 and Lys409, resulted in partial catalytic defects and a thermosensitive growth phenotype. Structure-function relationships were clarified by introducing conservative substitutions. Five residues were found to be nonessential: Lys309, Ser395, Asp397, Lys427 Asn431, and Lys474. The present findings, together with earlier mutational analyses, reveal an unusually complex active site in which 15 individual side chains in the tunnel cavity are important for catalysis, and each of the 8 strands of the beta barrel contributes at least one functional constituent. The active site residues fall into three classes: (i) those that participate directly in catalysis via coordination of the gamma phosphate or the metal; (ii) those that make critical water-mediated contacts with the gamma phosphate or the metal; and (iii) those that function indirectly via interactions with other essential side chains or by stabilization of the tunnel structure. PMID- 11279162 TI - Cloning of a human type II phosphatidylinositol 4-kinase reveals a novel lipid kinase family. AB - Phosphoinositide lipids regulate numerous cellular processes in all eukaryotes. The versatility of this phospholipid is provided by combinations of phosphorylation on the 3', 4', and 5' positions of the inositol head group. Two distinct structural families of phosphoinositide (PI) kinases have so far been identified and named after their prototypic members, the PI 3-kinase and phosphatidylinositol (PtdIns) phosphate kinase families, both of which have been found to contain structural homologues possessing PI 4-kinase activity. Nevertheless, the prevalent PtdIns 4-kinase activity in many mammalian cell types is conferred by the widespread type II PtdIns 4-kinase, which has so far resisted molecular characterization. We have partially purified the human type II isoform from plasma membrane rafts of human A431 epidermoid carcinoma cells and obtained peptide mass and sequence data. The results allowed the cDNA containing the full open reading frame to be cloned. The predicted amino acid sequence revealed that the type II enzyme is the prototypic member of a novel, third family of PI kinases. We have named the purified protein type IIalpha and a second human isoform, type IIbeta. The type IIalpha mRNA appears to be expressed ubiquitously in human tissues, and homologues appear to be expressed in all eukaryotes. PMID- 11279163 TI - Src family kinases are required for integrin-mediated but not for G protein coupled receptor stimulation of focal adhesion kinase autophosphorylation at Tyr 397. AB - Plating suspended Swiss 3T3 cells onto fibronectin-coated dishes promoted phosphorylation of endogenous focal adhesion kinase (FAK) at Tyr-397, the major autophosphorylation site, and at Tyr-577, located in the activation loop, as revealed by site-specific antibodies that recognize the phosphorylated form of these residues. Treatment with the selective Src family kinase inhibitor pyrazolopyrimidine 2 (PP-2) markedly reduced the phosphorylation of both Tyr-397 and Tyr-577 induced by fibronectin. Furthermore, fibronectin-mediated FAK phosphorylation at Tyr-397 was dramatically reduced in SYF cells (deficient in Src, Yes, and Fyn expression). Stimulation of Swiss 3T3 cells with bombesin also induced a rapid increase in the phosphorylation of endogenous FAK at Tyr-397. In contrast to the results obtained with fibronectin, PP-2 did not prevent FAK Tyr 397 phosphorylation stimulated by bombesin at a concentration (10 micrometer) that suppressed bombesin-induced FAK Tyr-577 phosphorylation. Similarly, PP-2 did not prevent Tyr-397 phosphorylation in Swiss 3T3 cells stimulated with other G protein-coupled receptor agonists including vasopressin, bradykinin, endothelin, and lysophosphatidic acid. Lysophosphatidic acid also induced FAK phosphorylation at Tyr-397 in SYF cells. Our results identify, for first time, the existence of Src-dependent and Src-independent pathways leading to FAK autophosphorylation at Tyr-397 stimulated by adhesion-dependent signals and G protein-coupled receptor agonists in the same cell. PMID- 11279164 TI - Limited accumulation of damaged proteins in l-isoaspartyl (D-aspartyl) O methyltransferase-deficient mice. AB - l-Isoaspartyl (d-aspartyl) O-methyltransferase (PCMT1) can initiate the conversion of damaged aspartyl and asparaginyl residues to normal l-aspartyl residues. Mice lacking this enzyme (Pcmt1-/- mice) have elevated levels of damaged residues and die at a mean age of 42 days from massive tonic-clonic seizures. To extend the lives of the knockout mice so that the long term effects of damaged residue accumulation could be investigated, we produced transgenic mice with a mouse Pcmt1 cDNA under the control of a neuron-specific promoter. Pcmt1 transgenic mice that were homozygous for the endogenous Pcmt1 knockout mutation ("transgenic Pcmt1-/- mice") had brain PCMT1 activity levels that were 6.5-13% those of wild-type mice but had little or no activity in other tissues. The transgenic Pcmt1-/- mice lived, on average, 5-fold longer than nontransgenic Pcmt1-/- mice and accumulated only half as many damaged aspartyl residues in their brain proteins. The concentration of damaged residues in heart, testis, and brain proteins in transgenic Pcmt1-/- mice initially increased with age but unexpectedly reached a plateau by 100 days of age. Urine from Pcmt1-/- mice contained increased amounts of peptides with damaged aspartyl residues, apparently enough to account for proteins that were not repaired intracellularly. In the absence of PCMT1, proteolysis may limit the intracellular accumulation of damaged proteins but less efficiently than in wild-type mice having PCMT1 mediated repair. PMID- 11279165 TI - Crystal structure of activated CheY. Comparison with other activated receiver domains. AB - The crystal structure of BeF(3)(-)-activated CheY, with manganese in the magnesium binding site, was determined at 2.4-A resolution. BeF(3)(-) bonds to Asp(57), the normal site of phosphorylation, forming a hydrogen bond and salt bridge with Thr(87) and Lys(109), respectively. The six coordination sites for manganese are satisfied by a fluorine of BeF(3)(-), the side chain oxygens of Asp(13) and Asp(57), the carbonyl oxygen of Asn(59), and two water molecules. All of the active site interactions seen for BeF(3)(-)-CheY are also observed in P Spo0A(r). Thus, BeF(3)(-) activates CheY as well as other receiver domains by mimicking both the tetrahedral geometry and electrostatic potential of a phosphoryl group. The aromatic ring of Tyr(106) is found buried within a hydrophobic pocket formed by beta-strand beta4 and helix H4. The tyrosine side chain is stabilized in this conformation by a hydrogen bond between the hydroxyl group and the backbone carbonyl oxygen of Glu(89). This hydrogen bond appears to stabilize the active conformation of the beta4/H4 loop. Comparison of the backbone coordinates for the active and inactive states of CheY reveals that only modest changes occur upon activation, except in the loops, with the largest changes occurring in the beta4/H4 loop. This region is known to be conformationally flexible in inactive CheY and is part of the surface used by activated CheY for binding its target, FliM. The pattern of activation-induced backbone coordinate changes is similar to that seen in FixJ(r). A common feature in the active sites of BeF(3)(-)-CheY, P-Spo0A(r), P-FixJ(r), and phosphono-CheY is a salt bridge between Lys(109) Nzeta and the phosphate or its equivalent, beryllofluoride. This suggests that, in addition to the concerted movements of Thr(87) and Tyr(106) (Thr-Tyr coupling), formation of the Lys(109)-PO(3)(-) salt bridge is directly involved in the activation of receiver domains generally. PMID- 11279166 TI - Insulin Induction of SOCS-2 and SOCS-3 mRNA expression in C2C12 Skeletal Muscle Cells Is Mediated by Stat5*. AB - Previously, by a yeast 2-hybrid screen, we identified signal transducer and activator of transcription 5b (Stat5b) as a substrate of the insulin receptor (IR). We demonstrated that refeeding of fasted mice leads to rapid activation of Stat5 proteins in liver, skeletal muscle, and fat, suggesting that Stat5b is a physiological target of insulin. Here, we show that injection of glucose or insulin into fasted mice leads to robust activation of both Stat5a and Stat5b in skeletal muscle. In C2C12 myotubes, we find that insulin stimulates tyrosine phosphorylation of Stat5a and Stat5b by 3-5-fold. This degree of Stat5 activation in vitro is significantly lower than what we observe in vivo and inversely correlates with IRS-1/2 levels. We can recapitulate robust insulin activation of Stat5 in C2C12 cells by stable overexpression of the human IR (hIR). To identify insulin-activated genes that are Stat5 targets, we also overexpressed an IR mutant (LA-hIR) that signals normally for mitogen-activated protein kinase- and phosphatidylinositol 3-kinase-dependent pathways but is deficient in Stat5 signaling in response to insulin. We demonstrate that insulin induces the expression of SOCS-2 mRNA in the wild type hIR but not in the LA-hIR overexpressing cells. The induction of SOCS-3 by insulin is reduced but not lost in the LA-hIR cells. Therefore, our results suggest that insulin induction of SOCS-2, and in part SOCS-3 mRNA expression, is mediated by Stat5 and can be independent of mitogen-activated protein kinase and phosphatidylinositol 3-kinase signaling pathways. PMID- 11279168 TI - The mannose/N-acetylgalactosamine-4-SO4 receptor displays greater specificity for multivalent than monovalent ligands. AB - Recognition of carbohydrates on glycosylated molecules typically requires multivalent interactions with receptors. Monovalent forms of terminal saccharides engaged by the receptor binding sites typically display weak affinities in the mm range and poor specificity. In contrast, multivalent forms of the same saccharides are bound with strong affinity (10(-7)-10(-9) m) and significantly greater specificity. Although multivalency can readily account for increased affinity, the molecular basis for enhanced specificity is not well understood. We have examined the specificity of the cysteine-rich domain of the mannose/GalNAc-4 SO4 receptor using monovalent and multivalent forms of the trisaccharide GalNAcbeta1,4GlcNAcbeta1,2Manalpha (GGnM) sulfated at either the C4 (S4GGnM) or C3 (S3GGnM) hydroxyl of the terminal GalNAc. Monovalent S4GGnM and S3GGnM have K(i) values of 25.8 and 16.2 microm, respectively. Multivalent conjugates of the same GalNAc-4-SO4- and GalNAc-3-SO4-bearing trisaccharides (6.7 mol of trisaccharide/mol of bovine serum albumin) have K(i) values of 0.013 and 0.170 microm, respectively. The 2000-fold versus 95-fold change in affinity seen for the multivalent forms of these 4-sulfated and 3-sulfated trisaccharides reflects a difference in the impact of conformational entropy. A large fraction of the SO4 3-GalNAc structures exists in a form that is not favorable for binding to the Cys rich domain. This reduces the effective concentration of SO4-3-GalNAc as compared with SO4-4-GalNAc under the same conditions and results in a markedly lower association rate. This difference in association rate accounts for the 12-fold difference in the rate of clearance from the blood seen with S4GGnM-BSA and S3GGnM-BSA in vivo. PMID- 11279167 TI - rDrak1, a novel kinase related to apoptosis, is strongly expressed in active osteoclasts and induces apoptosis. AB - This is the first report of a novel serine/threonine kinase, rabbit death associated protein (DAP) kinase-related apoptosis-inducing protein kinase 1 (rDRAK1), involved in osteoclast apoptosis. We searched for osteoclast-specific genes from a cDNA library of highly enriched rabbit osteoclasts cultured on ivory. One of the cloned genes has a high homology with human DRAK1 (hDRAK1), which belongs to the DAP kinase subfamily of serine/threonine kinases. By screening a rabbit osteoclast cDNA library and 5'-RACE (rapid amplification of cDNA ends), we obtained a full length of this cDNA, termed rDRAK1. The sequencing data indicated that rDRAK1 has 88.0, 44.6, 38.7, and 42.3% identity with hDRAK1, DAP kinase, DRP-1, and ZIP (zipper-interacting protein) kinase, respectively. To clarify the role of DRAK1 in osteoclasts, we examined the effect of three osteoclast survival factors (interleukin-1, macrophage colony-stimulating factor, and osteoclast differentiation-inducing factor) on rDRAK1 mRNA expression and the effect of rDRAK1 overexpression on osteoclast apoptosis. The results suggested that these three survival factors were proved to inhibit rDRAK1 expression in rabbit osteoclasts. After transfection of a rDRAK1 expression vector into cultured osteoclasts, overexpressed rDRAK1 was localized exclusively to the nuclei and induced apoptosis. Hence, rDRAK1 may play an important role in the core apoptosis program in osteoclast. PMID- 11279169 TI - Tyrosine sulfation of glycoprotein I(b)alpha. Role of electrostatic interactions in von Willebrand factor binding. AB - Glycoprotein I(b)alpha (GP I(b)alpha), the ligand binding subunit of the platelet glycoprotein Ib-IX-V complex, is sulfated on three tyrosine residues (Tyr-276, Tyr-278, and Tyr-279). This posttranslational modification is known to be critical for von Willebrand factor (vWF) binding; yet it remains unclear whether it provides a specific structure or merely contributes negative charges. To investigate this issue, we constructed cell lines expressing GP I(b)alpha polypeptides with the three tyrosine residues converted to either Glu or Phe and studied the ability of these mutants to bind vWF in the presence of modulators or shear stress. The mutants were expressed normally on the cell surface as GP Ib-IX complexes, with the conformation of the ligand-binding domain preserved, as judged by the binding of conformation-sensitive monoclonal antibodies. In contrast to their normal expression, both mutants were functionally abnormal. Cells expressing the Phe mutant failed to bind vWF in the presence of either ristocetin or botrocetin. These cells adhered to and rolled on immobilized vWF only when their surface receptor density was increased to twice the level that supported adhesion of cells expressing the wild-type receptor and even then only 20% as many rolled and rolled significantly faster than wild-type cells. Cells expressing the Glu mutant, on the other hand, were normal with respect to ristocetin-induced vWF binding and adhesion to immobilized vWF but were markedly defective in botrocetin-induced vWF binding. These results indicate that GP I(b)alpha tyrosine sulfation influences the interaction of this polypeptide with vWF primarily by contributing negative charges under physiological conditions and when the interaction is induced by ristocetin but contributes a specific structure to the botrocetin-induced interaction. PMID- 11279170 TI - Functional uncoupling of T-cell receptor engagement and Lck activation in anergic human thymic CD4+ T cells. AB - Human thymic CD1a-CD4+ T cells in the final stage of thymic maturation are susceptible to anergy induced by a superantigen, toxic shock syndrome toxin-1 (TSST-1). Thymic CD4+ T-cell blasts, established by stimulating human thymic CD1a CD4+ T cells with TSST-1 in vitro, produce a low level of interleukin-2 after restimulation with TSST-1, whereas TSST-1-induced adult peripheral blood (APB) CD4+ T-cell blasts produce high levels of interleukin-2. The extent of tyrosine phosphorylation of the T-cell receptor zeta chain induced after restimulation with TSST-1 was 2-4-fold higher in APB CD4+ T-cell blasts than in thymic CD4+ T cell blasts. The tyrosine kinase activity of Lck was low in both thymic and APB CD4+ T-cell blasts before restimulation with TSST-1. After restimulation, the Lck kinase activity increased in APB CD4+ T-cell blasts but not in thymic CD4+ T-cell blasts. Surprisingly, Lck was highly tyrosine-phosphorylated in both thymic and APB CD4+ T-cell blasts before restimulation with TSST-1. After restimulation, it was markedly dephosphorylated in APB CD4+ T-cell blasts but not in thymic CD4+ T cell blasts. Lck from APB CD4+ T-cell blasts bound the peptide containing the phosphotyrosine at the negative regulatory site of Lck-505 indicating that the site of dephosphorylation in TSST-1-activated T-cell blasts is Tyr-505. Confocal microscopy demonstrated that colocalization of Lck and CD45 was induced after restimulation with TSST-1 in APB CD4+ T-cell blasts but not in thymic CD4+ T-cell blasts. Further, remarkable accumulation of Lck in the membrane raft was observed in restimulated APB CD4+ T-cell blasts but not in thymic CD4+ T-cell blasts. These data indicate that interaction between Lck and CD45 is suppressed physically in thymic CD4+ T-cell blasts and plays a critical role in sustaining an anergic state. PMID- 11279171 TI - Acyl-CoA esters antagonize the effects of ligands on peroxisome proliferator activated receptor alpha conformation, DNA binding, and interaction with Co factors. AB - The peroxisome proliferator-activated receptor alpha (PPARalpha) is a ligand activated transcription factor and a key regulator of lipid homeostasis. Numerous fatty acids and eicosanoids serve as ligands and activators for PPARalpha. Here we demonstrate that S-hexadecyl-CoA, a nonhydrolyzable palmitoyl-CoA analog, antagonizes the effects of agonists on PPARalpha conformation and function in vitro. In electrophoretic mobility shift assays, S-hexadecyl-CoA prevented agonist-induced binding of the PPARalpha-retinoid X receptor alpha heterodimer to the acyl-CoA oxidase peroxisome proliferator response element. PPARalpha bound specifically to immobilized palmitoyl-CoA and Wy14643, but not BRL49653, abolished binding. S-Hexadecyl-CoA increased in a dose-dependent and reversible manner the sensitivity of PPARalpha to chymotrypsin digestion, and the S hexadecyl-CoA-induced sensitivity required a functional PPARalpha ligand-binding pocket. S-Hexadecyl-CoA prevented ligand-induced interaction between the co activator SRC-1 and PPARalpha but increased recruitment of the nuclear receptor co-repressor NCoR. In cells, the concentration of free acyl-CoA esters is kept in the low nanomolar range due to the buffering effect of high affinity acyl-CoA binding proteins, especially the acyl-CoA-binding protein. By using PPARalpha expressed in Sf21 cells for electrophoretic mobility shift assays, we demonstrate that S-hexadecyl-CoA was able to increase the mobility of the PPARalpha containing heterodimer even in the presence of a molar excess of acyl-CoA-binding protein, mimicking the conditions found in vivo. PMID- 11279173 TI - Flotillin-1-enriched lipid raft domains accumulate on maturing phagosomes. AB - Flotillin-1 was recently shown to be enriched on detergent-resistant domains of the plasma membrane called lipid rafts. These rafts, enriched in sphingolipids and cholesterol, sequester certain proteins while excluding others. Lipid rafts have been implicated in numerous cellular processes including signal transduction, membrane trafficking, and molecular sorting. In this study, we demonstrate both morphologically and biochemically that lipid rafts are present on phagosomes. These structures are enriched in flotillin-1 and devoid of the main phagosomes membrane protein lysosomal-associated membrane protein (LAMP1). The flotillin-1 present on phagosomes does not originate from the plasma membrane during phagocytosis but accumulates gradually on maturing phagosomes. Treatment with bafilomycin A1, a compound that inhibits the proton pump ATPase and prevents the fusion of phagosomes with late endocytic organelles, prevents the acquisition of flotillin-1 by phagosomes, indicating that this protein might be recruited on phagosomes from endosomal organelles. A proteomic characterization of the lipid rafts of phagosomes indicates that actin, the alpha- and beta-subunits of heterotrimeric G proteins, as well as subunits of the proton pump V-ATPase are among the constituents of these domains. Remarkably, the intracellular parasite Leishmania donovani can actively inhibit the acquisition of flotillin-1-enriched lipid rafts by phagosomes and the maturation of these organelles. These results indicate that specialized functions required for phagolysosome biogenesis may occur at focal points on the phagosome membrane, and therefore represent a potential target of intracellular pathogens. PMID- 11279172 TI - MKK6/3 and p38 MAPK pathway activation is not necessary for insulin-induced glucose uptake but regulates glucose transporter expression. AB - p38 mitogen-activated protein kinase (MAPK), which is situated downstream of MAPK kinase (MKK) 6 and MKK3, is activated by mitogenic or stress-inducing stimuli, as well as by insulin. To clarify the role of the MKK6/3-p38 MAPK pathway in the regulation of glucose transport, dominant negative p38 MAPK and MKK6 mutants and constitutively active MKK6 and MKK3 mutants were overexpressed in 3T3-L1 adipocytes and L6 myotubes using an adenovirus-mediated transfection procedure. Constitutively active MKK6/3 mutants up-regulated GLUT1 expression and down regulated GLUT4 expression, thereby significantly increasing basal glucose transport but diminishing transport induced by insulin. Similar effects were elicited by chronic (24 h) exposure to tumor necrosis factor alpha, interleukin 1beta, or 200 mm sorbitol, all activate the MKK6/3-p38 MAPK pathway. SB203580, a specific p38 MAPK inhibitor, attenuated these effects, further confirming that both MMK6 and MMK3 act via p38 MAPK, whereas they had no effect on the increase in glucose transport induced by a constitutively active MAPK kinase 1 (MEK1) mutant or by myristoylated Akt. In addition, suppression of p38 MAPK activation by overexpression of a dominant negative p38 MAPK or MKK6 mutant did not diminish insulin-induced glucose uptake by 3T3-L1 adipocytes. It is thus apparent that activation of p38 MAPK is not essential for insulin-induced increases in glucose uptake. Rather, p38 MAPK activation leads to a marked down-regulation of insulin induced glucose uptake via GLUT4, which may underlie cellular stress-induced insulin resistance caused by tumor necrosis factor alpha and other factors. PMID- 11279175 TI - Regulation of a human chloride channel. a paradigm for integrating input from calcium, type ii calmodulin-dependent protein kinase, and inositol 3,4,5,6 tetrakisphosphate. AB - We have studied the regulation of Ca(2+)-dependent chloride (Cl(Ca)) channels in a human pancreatoma epithelial cell line (CFPAC-1), which does not express functional cAMP-dependent cystic fibrosis transmembrane conductance regulator chloride channels. In cell-free patches from these cells, physiological Ca(2+) concentrations activated a single class of 1-picosiemens Cl(-)-selective channels. The same channels were also stimulated by a purified type II calmodulin dependent protein kinase (CaMKII), and in cell-attached patches by purinergic agonists. In whole-cell recordings, both Ca(2+)- and CaMKII-dependent mechanisms contributed to chloride channel stimulation by Ca(2+), but the CaMKII-dependent pathway was selectively inhibited by inositol 3,4,5,6-tetrakisphosphate (Ins(3,4,5,6)P(4)). This inhibitory effect of Ins(3,4,5,6)P(4) on Cl(Ca) channel stimulation by CaMKII was reduced by raising [Ca(2+)] and prevented by inhibition of protein phosphatase activity with 100 nm okadaic acid. These data provide a new context for understanding the physiological relevance of Ins(3,4,5,6)P(4) in the longer term regulation of Ca(2+)-dependent Cl(-) fluxes in epithelial cells. PMID- 11279176 TI - Identification of an ATP-binding cassette transporter for export of the O-antigen across the inner membrane in Rhizobium etli based on the genetic, functional, and structural analysis of an lps mutant deficient in O-antigen. AB - For O-antigen lipopolysaccharide (LPS) synthesis in bacteria, transmembrane migration of undecaprenyl pyrophosphate-bound O-antigen oligosaccharide subunits or polysaccharide occurs before ligation to the core region of the LPS molecule. In this study, we identified by mutagenesis an ATP-binding cassette transporter in Rhizobium etli CE3 that is likely responsible for the translocation of the O antigen across the inner plasma membrane. Mutant FAJ1200 LPS lacks largely the O antigen, as shown by SDS-polyacrylamide gel electrophoresis and confirmed by immunoblot analysis. Furthermore, LPS isolated from FAJ1200 is totally devoid of any O-chain glycosyl residues and contains only those glycosyl residues that can be expected for the inner core region. The membrane component and the cytoplasmic ATP-binding component of the ATP-binding cassette transporter are encoded by wzm and wzt, respectively. The Tn5 transposon in mutant FAJ1200 is inserted in the wzm gene. This mutation resulted in an Inf- phenotype in bean plants. PMID- 11279178 TI - Molecular cloning and characterization of spiggin. An androgen-regulated extraorganismal adhesive with structural similarities to von Willebrand Factor related proteins. AB - One of the most definitive examples of a vertebrate extraorganismal structural protein can be found in three-spined sticklebacks (Gasterosteus aculeatus). In the breeding male the kidney hypertrophies and synthesizes an adhesive protein called "spiggin," which is secreted into the urinary bladder from where it is employed as a structural thread for nest building. This paper describes the first molecular characterization of spiggin and demonstrates that this adhesive is a protein complex assembled from a potential of three distinct subunits (alpha, beta, and gamma). These subunits arise by alternative splicing, and 11 ketoandrogens induce their expression in stickleback kidneys. Analysis of the predicted amino acid sequence of each subunit reveals a modular organization whose structural elements display a similarity to the multimerization domains found within von Willebrand Factor-related proteins. These results implicate that spiggin utilizes a conserved multimerization mechanism for the formation of a viscous agglutinate from its constituent subunits in the urinary bladders of male sticklebacks. This novel extraorganismal structural protein is therefore ideally suited to its function as an adhesive thread. PMID- 11279177 TI - Arachidonic acid modulates the spatiotemporal characteristics of agonist-evoked Ca2+ waves in mouse pancreatic acinar cells. AB - In pancreatic acinar cells analysis of the propagation speed of secretagogue evoked Ca2+ waves can be used to examine coupling of hormone receptors to intracellular signal cascades that cause activation of protein kinase C or production of arachidonic acid (AA). In the present study we have investigated the role of cytosolic phospholipase A2 (cPLA2) and AA in acetylcholine (ACh)- and bombesin-induced Ca2+ signaling. Inhibition of cPLA2 caused acceleration of ACh induced Ca2+ waves, whereas bombesin-evoked Ca2+ waves were unaffected. When enzymatic metabolization of AA was prevented with the cyclooxygenase inhibitor indomethacin or the lipoxygenase inhibitor nordihydroguaiaretic acid, ACh-induced Ca2+ waves were slowed down. Agonist-induced activation of cPLA2 involves mitogen activated protein kinase (MAPK) activation. An increase in phosphorylation of p38(MAPK) and p42/44(MAPK) within 10 s after stimulation could be demonstrated for ACh but was absent for bombesin. Rapid phosphorylation of p38(MAPK) and p42/44(MAPK) could also be observed in the presence of cholecystokinin (CCK), which also causes activation of cPLA2. ACh-and CCK-induced Ca2+ waves were slowed down when p38(MAPK) was inhibited with SB 203580, whereas inhibition of p42/44(MAPK) with PD 98059 caused acceleration of ACh- and CCK-induced Ca2+ waves. The spreading of bombesin-evoked Ca2+ waves was affected neither by PD 98059 nor by SB 203580. Our data indicate that in mouse pancreatic acinar cells both ACh and CCK receptors couple to the cPLA2 pathway. cPLA2 activation occurs within 1-2 s after hormone application and is promoted by p42/44(MAPK) and inhibited by p38(MAPK). Furthermore, the data demonstrate that secondary (Ca2+ induced) Ca2+ release, which supports Ca2+ wave spreading, is inhibited by AA itself and not by a metabolite of AA. PMID- 11279180 TI - A chaperone for ribosome maturation. AB - The nascent pre-rRNA of eukaryotic ribosomes is fully transcribed and assembled into an 80-90 S nucleolar particle before being cleaved into mature ribosomal RNA. The interdependence of steps in the processing of this precursor RNA indicates that RNA processing, at least in part, acts as a quality control mechanism that helps ensure that only functional RNA is incorporated into mature ribosomes. In search of structural components that underlie this interdependence using the Schizosaccharomyces pombe internal transcribed spacer 1 (ITS) as a ligand for affinity chromatography of ITS1-specific proteins, we have isolated a large spliceosome-like protein complex, a ribosome assembly chaperone (RAC) of 20 or more polypeptides (Lalev, A. I., Abeyrathne, P. D., and Nazar, R. N. (2000) J. Mol. Biol. 302, 65-77). When the ITS2 spacer was used in the present study to isolate ITS2-specific proteins, the same proteins were identified consistent with a complex containing multiple specific binding sites. Subsequent competition binding studies indicated that the protein complex actually contains independent binding sites for all four of the transcribed spacers in the pre-rRNA. Because disruption of protein-binding sites in these spacer RNAs is known to severely affect rRNA processing, taken together these results suggest that the RAC complex is a chaperone for ribosome maturation acting as a "rack" on which critical structure is organized. PMID- 11279179 TI - Human epidermal growth factor (EGF) module-containing mucin-like hormone receptor 3 is a new member of the EGF-TM7 family that recognizes a ligand on human macrophages and activated neutrophils. AB - The epidermal growth factor (EGF)-TM7 subgroup of G-protein-coupled receptors is composed predominantly of leukocyte-restricted glycoproteins defined by their unique hybrid structure, in which extracellular EGF-like domains are coupled to a seven-span transmembrane moiety via a mucin-like stalk. The EGF-TM7 group comprises mouse F4/80, human EGF module-containing mucin-like hormone receptor (EMR) 1, human EMR2, and human and mouse CD97, the genes for which map to human chromosome 19p13 and the syntenic regions of the mouse genome. In this study we describe the cloning and characterization of EMR3, a novel human EGF-TM7 molecule, and show the existence of its cellular ligand. The EMR3 gene maps closely to the existing members of the EGF-TM7 family on human chromosome 19p13.1 and, in common with other EGF-TM7 genes, is capable of generating different protein isoforms through alternative splicing. Two alternative splice forms have been isolated: one encoding a 652-amino acid cell surface protein consisting of two EGF-like domains, a mucin stalk, and a putative G-protein-coupled receptor domain and the other encoding a truncated soluble form containing only two EGF like domains. As with other members of the EGF-TM7 family, EMR3 mRNA displays a predominantly leukocyte-restricted expression pattern, with highest levels in neutrophils, monocytes, and macrophages. Through the use of soluble EMR3 multivalent probes we have shown the presence of a ligand at the surface of monocyte-derived macrophages and activated human neutrophils. These interactions suggest a potential role for EMR3 in myeloid-myeloid interactions during immune and inflammatory responses. PMID- 11279181 TI - Regulation of myogenic terminal differentiation by the hairy-related transcription factor CHF2. AB - We have recently cloned a novel basic helix-loop-helix factor, CHF2, that functions as a transcriptional repressor. To address its role in the regulation of myogenic terminal differentiation, we analyzed its expression pattern during C2C12 mouse myotube formation. In undifferentiated myoblasts, CHF2 is expressed at high levels. After induction of myotube formation in low serum, CHF2 expression is barely detectable at 3 days after induction. Myogenin expression, in contrast, peaks at 3 days. In transiently transfected 10T1/2 embryonic fibroblasts, CHF2 inhibited MyoD-dependent activation of the myogenin promoter in a dose-dependent fashion. Electrophoretic mobility shift analysis indicated that CHF2 inhibits the binding of the MyoD.E47 heterodimer to the E-box binding site. CHF2 also inhibited myogenic conversion of 10T1/2 cells by MyoD, as measured by skeletal myosin heavy chain protein expression. Coimmunoprecipitation analysis indicated that CHF2 forms a protein complex with MyoD. Mutational analysis of CHF2 indicated that the repression activity for both transcription and myogenic conversion mapped to a hydrophobic carboxyl-terminal region and did not require either the basic helix-loop-helix or YRPW motifs. Our data indicate that CHF2 functions as a transcriptional repressor of myogenesis by formation of an inactive heterodimeric complex with MyoD and likely plays an important role in muscle development. PMID- 11279182 TI - A-kinase-anchoring protein AKAP95 is targeted to the nuclear matrix and associates with p68 RNA helicase. AB - The cell nucleus is structurally and functionally organized by the nuclear matrix. We have examined whether the nuclear cAMP-dependent protein kinase anchoring protein AKAP95 contains specific signals for targeting to the subnuclear compartment and for interaction with other proteins. AKAP95 was expressed in mammalian cells and found to localize exclusively to the nuclear matrix. Mutational analysis was used to identify determinants for nuclear localization and nuclear matrix targeting of AKAP95. These sites were found to be distinct from previously identified DNA and protein kinase A binding domains. The nuclear matrix-targeting site is unique but conserved among members of the AKAP95 family. Direct binding of AKAP95 to isolated nuclear matrix was demonstrated in situ and found to be dependent on the nuclear matrix-targeting site. Moreover, Far Western blot analysis identified at least three AKAP95-binding proteins in nuclear matrix isolated from rat brain. Yeast two-hybrid cloning identified one binding partner as p68 RNA helicase. The helicase and AKAP95 co-localized in the nuclear matrix of mammalian cells, associated in vitro, and were precipitated as a complex from solubilized cell extracts. The results define novel protein protein interactions among nuclear matrix proteins and suggest a potential role of AKAP95 as a scaffold for coordinating assembly of hormonally responsive transcription complexes. PMID- 11279183 TI - Chemical probing shows that the intron-encoded endonuclease I-SceI distorts DNA through binding in monomeric form to its homing site. AB - Despite its small size (27.6 kDa), the group I intron-encoded I-SceI endonuclease initiates intron homing by recognizing and specifically cleaving a large intronless DNA sequence. Here, we used gel shift assays and footprinting experiments to analyze the interaction between I-SceI and its target. I-SceI was found to bind to its substrate in monomeric form. Footprinting using DNase I, hydroxyl radical, phenanthroline copper complexes, UV/DH-MePyPs photosensitizer, and base-modifying reagents revealed the asymmetric nature of the interaction and provided a first glimpse into the architecture of the complex. The protein interacts in the minor and major grooves and distorts DNA at three distinct sites: one at the intron insertion site and the other two, respectively, downstream (-8, -9) and upstream (+9, +10) from this site. The protein appears to stabilize the DNA curved around it by bridging the minor groove on one face of the helix. The scissile phosphates would lie on the outside of the bend, facing in the same direction relative to the DNA helical axis, as expected for an endonuclease that generates 3' overhangs. An internally consistent model is proposed in which the protein would take advantage of the concerted flexibility of the DNA sequence to induce a synergistic binding/kinking process, resulting in the correct positioning of the enzyme active site. PMID- 11279184 TI - Novel members of the human oxysterol-binding protein family bind phospholipids and regulate vesicle transport. AB - Oxysterol-binding proteins (OSBPs) are a family of eukaryotic intracellular lipid receptors. Mammalian OSBP1 binds oxygenated derivatives of cholesterol and mediates sterol and phospholipid synthesis through as yet poorly undefined mechanisms. The precise cellular roles for the remaining members of the oxysterol binding protein family remain to be elucidated. In yeast, a family of OSBPs has been identified based on primary sequence similarity to the ligand binding domain of mammalian OSBP1. Yeast Kes1p, an oxysterol-binding protein family member that consists of only the ligand binding domain, has been demonstrated to regulate the Sec14p pathway for Golgi-derived vesicle transport. Specifically, inactivation of the KES1 gene resulted in the ability of yeast to survive in the absence of Sec14p, a phosphatidylinositol/phosphatidylcholine transfer protein that is normally required for cell viability due to its essential requirement in transporting vesicles from the Golgi. We cloned the two human members of the OSBP family, ORP1 and ORP2, with the highest degree of similarity to yeast Kes1p. We expressed ORP1 and ORP2 in yeast lacking Sec14p and Kes1p function and found that ORP1 complemented Kes1p function with respect to cell growth and Golgi vesicle transport, whereas ORP2 was unable to do so. Phenotypes associated with overexpression of ORP2 in yeast were a dramatic decrease in cell growth and a block in Golgi-derived vesicle transport distinct from that of ORP1. Purification of ORP1 and ORP2 for ligand binding studies demonstrated ORP1 and ORP2 did not bind 25-hydroxycholesterol but instead bound phospholipids with both proteins exhibiting strong binding to phosphatidic acid and weak binding to phosphatidylinositol 3-phosphate. In Chinese hamster ovary cells, ORP1 localized to a cytosolic location, whereas ORP2 was associated with the Golgi apparatus, consistent with our vesicle transport studies that indicated ORP1 and ORP2 function at different steps in the regulation of vesicle transport. PMID- 11279185 TI - CD95 signaling via ceramide-rich membrane rafts. AB - Clustering seems to be employed by many receptors for transmembrane signaling. Here, we show that acid sphingomyelinase (ASM)-released ceramide is essential for clustering of CD95. In vitro and in vivo, extracellularly orientated ceramide, released upon CD95-triggered translocation of ASM to the plasma membrane outer surface, enabled clustering of CD95 in sphingolipid-rich membrane rafts and apoptosis induction. Whereas ASM deficiency, destruction of rafts, or neutralization of surface ceramide prevented CD95 clustering and apoptosis, natural ceramide only rescued ASM-deficient cells. The data suggest CD95-mediated clustering by ceramide is prerequisite for signaling and death. PMID- 11279186 TI - Different recognition of DNA modified by aatitumor cisplatin and its clinically ineffective trans isomer by tumor suppressor protein p53. AB - The p53 gene encodes a nuclear phosphoprotein that is biologically activated in response to genotoxic stresses including treatment with anticancer platinum drugs. The DNA binding activity of p53 protein is crucial for its tumor suppressor function. DNA interactions of active wild-type human p53 protein with DNA fragments and oligodeoxyribonucleotide duplexes modified by antitumor cisplatin and its clinically ineffective trans isomer (transplatin) were investigated by using a gel mobility shift assay. It was found that DNA adducts of cisplatin reduced binding affinity of the consensus DNA sequence to p53, whereas transplatin adducts did not. This result was interpreted to mean that the precise steric fit required for the formation and stability of the tetrameric complex of p53 with the consensus sequence cannot be attained, as a consequence of severe conformational perturbations induced in DNA by cisplatin adducts. The results also demonstrate an increase of the binding affinity of p53 to DNA lacking the consensus sequence and modified by cisplatin but not by transplatin. In addition, only major 1,2-GG intrastrand cross-links of cisplatin are responsible for this enhanced binding affinity of p53. The data base on structures of various DNA adducts of cisplatin and transplatin reveals distinctive structural features of 1,2-intrastrand cross-links of cisplatin, suggesting a unique role for this adduct in the binding of p53 to DNA lacking the consensus sequence. The results support the hypothesis that the mechanism of antitumor activity of cisplatin may also be associated with its efficiency to affect the binding affinity of platinated DNA to active p53 protein. PMID- 11279187 TI - Regulatory elements governing transcription in specialized myofiber subtypes. AB - Skeletal myofibers of vertebrates acquire specialized metabolic and physiological properties as a consequence of developmental cues in the embryo and different patterns of contractile activity in the adult. The myoglobin gene is regulated stringently in muscle fibers, such that high myoglobin expression is observed in mitochondria-rich, oxidative myofibers (Types I and IIa) compared with glycolytic fibers (Type IIb). Using germ-line transgenesis and somatic cell gene transfer methods, we defined discrete regions of the murine and human genes encoding myoglobin that are sufficient to confer muscle- and fiber type-specific expression to reporter genes. Mutational analysis confirms the importance of A/T rich, MEF2-binding motifs in myoglobin gene regulation, as suggested by previous studies using different experimental approaches. In addition, we demonstrated a previously unsuspected role for an intragenic E-box motif as a negative regulatory element contributing to the tightly regulated variation in myoglobin gene expression among particular myofiber subtypes. PMID- 11279188 TI - Functional geometry of the permeation pathway of Ca2+-activated Cl-channels inferred from analysis of voltage-dependent block. AB - We examined the voltage-dependent block of Ca(2+)-activated Cl(-) channels by anthacene-9-carboxylic acid (A9C), diphenylamine-2-carboxylic acid (DPC), 4,4' diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), and niflumic acid (NFA) in excised inside-out and outside-out patches from Xenopus oocytes. The fraction of the voltage field (delta) experienced by the blocking drug was determined from the voltage dependence of block. All the drugs blocked by entering the channel from the outside. delta was 0.6 for A9C, 0.3 for DPC and DIDS, and <0.1 for NFA. Because the voltage dependence of the drugs differed, the order of potency was also voltage-dependent. At +100 mV the order of potency was NFA > A9C > DIDS > DPC (K(i) (microm) = 10.1, 18.3, 48, and 111, respectively). Because the drugs are hydrophobic, they can cross the bilayer when applied from the inside and block the channel from the outside. The equilibrium geometries of the blockers were determined by molecular modeling and compared with their blocking positions (delta). This analysis suggests that the channel is an elliptical cone with the largest opening facing the extracellular space. The selectivity filter has an apparent size of 0.33 x 0.75 nm, because C(CN)(3)-, which has these dimensions, permeates. The external opening is at least 0.60 x 0.94 nm, because DPC has these dimensions and penetrates the channel approximately 30%. PMID- 11279189 TI - Mutations leading to X-linked hypohidrotic ectodermal dysplasia affect three major functional domains in the tumor necrosis factor family member ectodysplasin A. AB - Mutations in the epithelial morphogen ectodysplasin-A (EDA), a member of the tumor necrosis factor (TNF) family, are responsible for the human disorder X linked hypohidrotic ectodermal dysplasia (XLHED) characterized by impaired development of hair, eccrine sweat glands, and teeth. EDA-A1 and EDA-A2 are two splice variants of EDA, which bind distinct EDA-A1 and X-linked EDA-A2 receptors. We identified a series of novel EDA mutations in families with XLHED, allowing the identification of the following three functionally important regions in EDA: a C-terminal TNF homology domain, a collagen domain, and a furin protease recognition sequence. Mutations in the TNF homology domain impair binding of both splice variants to their receptors. Mutations in the collagen domain can inhibit multimerization of the TNF homology region, whereas those in the consensus furin recognition sequence prevent proteolytic cleavage of EDA. Finally, a mutation affecting an intron splice donor site is predicted to eliminate specifically the EDA-A1 but not the EDA-A2 splice variant. Thus a proteolytically processed, oligomeric form of EDA-A1 is required in vivo for proper morphogenesis. PMID- 11279190 TI - Mutation of Cys-67 alters the thermodynamic stability of the human leukocyte antigen HLA0-B*2705. AB - The B pocket of the class I major histocompatibility complex-encoded protein HLA B*2705 has recently been suggested to be responsible for the misfolding of this HLA haplotype and thus to induce susceptibility to autoimmune inflammatory diseases. Four mutants of the B*2705 heavy chain were refolded in the presence of three control peptides. The monitoring of the thermal unfolding of the B*2705 peptide complexes by circular dichroism spectroscopy showed that all heterotrimeric mutants were markedly less stable than the corresponding complexes with the wild-type heavy chain. Among the four heavy chain mutations, the C67S change was investigated for unfolding and peptide binding properties because this position may mediate disulfide pair bridging and alter T-cell recognition of HLA B*2705. Wild-type heterotrimers completely unfold in a single transition at mild acidic pH whereas increase of the pH to mild basic conditions induce only a partial biphasic unfolding. Cys-67 seems to play a crucial role in controlling the thermodynamic stability of the B*2705-peptide complexes as the C67S mutant unfolds faster and with a single transition, independent of pH. Fluorescence polarization and size exclusion chromatography of unfolding intermediates suggest that the peculiar unfolding of the B*2705 wild-type heavy chain cannot be explained by modified peptide binding properties but more likely by the formation of high molecular weight species. PMID- 11279191 TI - Dendritic cell-specific MHC class II transactivator contains a caspase recruitment domain that confers potent transactivation activity. AB - The MHC class II transactivator (CIITA) is a critical transcription factor that regulates genes involved in antigen presentation function. At least three functional forms of CIITA gene products are transcribed from three different promoters. The CIITA gene expressed in dendritic cells (DC-CIITA) has a unique first exon encoding an extended N-terminal region of CIITA. Here, we show that the N terminus of DC-CIITA has high homology to a caspase recruitment domain (CARD) found in components of apoptosis and nuclear factor-kappaB signaling pathways. However, DC-CIITA does not regulate cell death, nor does it induce nuclear factor-kappaB activity. Instead, DC-CIITA is transcriptionally a more potent activator of the MHC class II gene than the form expressed in B cells. A single amino acid substitution in the CARD of DC-CIITA, predicted to disrupt CARD CARD interactions, diminished the transactivation potential of DC-CIITA. These results indicate that the CARD in the context of CIITA serves as a regulatory domain for transcriptional activity and may function to selectively enhance MHC class II gene expression in dendritic cells. PMID- 11279192 TI - p53-dependent transcriptional regulation of the APC promoter in colon cancer cells treated with DNA alkylating agents. AB - The APC (adenomatous polyposis coli) gene product is involved in cell cycle arrest and in apoptosis. The loss of APC function is associated with the development of colorectal carcinogenesis. In previous studies, we have shown that the APC gene is inducible and that the DNA damage-induced level of APC mRNA requires p53. In the present study, we examined the role of p53 in the transcriptional regulation of APC promoter and characterized two p53-binding sites on the cloned APC promoter (pAPCP). Results of electrophoretic mobility shift assay showed specific interactions of p53 protein with p53-binding site oligonucleotides. The DNA-protein complex formed in electrophoretic mobility shift assay was competed with unlabeled excess of p53-binding site oligonucleotide, unaffected with p53-binding site mutant or Sp1-binding site oligonucleotides, and supershifted with anti-p53 antibodies. In a transient transfection assay, the pAPCP promoter activity was lower in HCT-116(p53(+/+)) cells versus HCT-116(p53(-/-)) cells. p53-dependent down-regulation was further confirmed after co-transfection of pAPCP plasmid with pCMV-p53 into HCT-116(p53( /-)) and SAOS-2 (p53-negative) cells. However, the treatment of cells with DNA alkylating agents methylmethane sulfonate and N-methyl-N'-nitro-N nitrosoguanidine, which cause phosphorylation of p53 at Ser(15) and Ser(392), induced pAPCP promoter activity in HCT-116(p53(+/+)) cells. Other than p53 binding sites, using deletion mutation constructs, we have shown that N-methyl-N' nitro-N-nitrosoguanidine-induced transcriptional activation of the pAPCP promoter in HCT-116(p53(+/+)) cells depended upon the Sp1-binding site and the E-box B site. From these results, we conclude that unphosphorylated p53 can down-regulate and phosphorylated p53 can up-regulate the pAPCP promoter activity involving the p53, Sp1, or E-box B elements. These studies are important to understanding the role of p53 and APC in DNA damage-induced cell cycle arrest and/or apoptosis of cancer cells. PMID- 11279193 TI - Molecular basis for severe epimerase deficiency galactosemia. X-ray structure of the human V94m-substituted UDP-galactose 4-epimerase. AB - Galactosemia is an inherited disorder characterized by an inability to metabolize galactose. Although classical galactosemia results from impairment of the second enzyme of the Leloir pathway, namely galactose-1-phosphate uridylyltransferase, alternate forms of the disorder can occur due to either galactokinase or UDP galactose 4-epimerase deficiencies. One of the more severe cases of epimerase deficiency galactosemia arises from an amino acid substitution at position 94. It has been previously demonstrated that the V94M protein is impaired relative to the wild-type enzyme predominantly at the level of V(max) rather than K(m). To address the molecular consequences the mutation imparts on the three-dimensional architecture of the enzyme, we have solved the structures of the V94M-substituted human epimerase complexed with NADH and UDP-glucose, UDP-galactose, UDP-GlcNAc, or UDP-GalNAc. In the wild-type enzyme, the hydrophobic side chain of Val(94) packs near the aromatic group of the catalytic Tyr(157) and serves as a molecular "fence" to limit the rotation of the glycosyl portions of the UDP-sugar substrates within the active site. The net effect of the V94M substitution is an opening up of the Ala(93) to Glu(96) surface loop, which allows free rotation of the sugars into nonproductive binding modes. PMID- 11279195 TI - Phosphorylation of centrin during the cell cycle and its role in centriole separation preceding centrosome duplication. AB - Once during each cell cycle, mitotic spindle poles arise by separation of newly duplicated centrosomes. We report here the involvement of phosphorylation of the centrosomal protein centrin in this process. We show that centrin is phosphorylated at serine residue 170 during the G(2)/M phase of the cell cycle. Indirect immunofluorescence staining of HeLa cells using a phosphocentrin specific antibody reveals intense labeling of mitotic spindle poles during prophase and metaphase of the cell division cycle, with diminished staining of anaphase and no staining of telophase and interphase centrosomes. Cultured cells undergo a dramatic increase in centrin phosphorylation following the experimental elevation of PKA activity, suggesting that this kinase can phosphorylate centrin in vivo. Surprisingly, elevated PKA activity also resulted intense phosphocentrin antibody labeling of interphase centrosomes and in the concurrent movement of individual centrioles apart from one another. Taken together, these results suggest that centrin phosphorylation signals the separation of centrosomes at prophase and implicates centrin phosphorylation in centriole separation that normally precedes centrosome duplication. PMID- 11279194 TI - Molecular cloning and characterization of a novel (Na+,K+)/H+ exchanger localized to the trans-Golgi network. AB - The luminal pH of organelles along the secretory and endocytic pathways of mammalian cells is acidic and tightly regulated, with the [H+] varying up to 100 fold between compartments. Steady-state organellar pH is thought to reflect a balance between the rates of H+ pumping by the vacuolar-type H+-ATPase and H+ efflux through ill-defined pathways. Here, we describe the cloning of a novel gene (NHE7) in humans that is homologous to Na+/H+ exchangers, is ubiquitously expressed, and localizes predominantly to the trans-Golgi network. Significantly, NHE7 mediates the influx of Na+ or K+ in exchange for H+. The activity of NHE7 was also found to be relatively insensitive to inhibition by amiloride but could be antagonized by the analogue benzamil and the unrelated compound quinine. Thus, NHE7 displays unique functional and pharmacological properties and may play an important role in maintaining cation homeostasis of this important organelle. PMID- 11279196 TI - A novel p75TNF receptor isoform mediating NFkappa B activation. AB - We report the identification of a novel p75TNF receptor isoform termed icp75TNFR, which is generated by the use of an alternative transcriptional start site within the p75TNFR gene and characterized by regulated intracellular expression. The icp75TNFR protein has an apparent molecular mass of approximately 50 kDa and is recognized by antibodies generated against the transmembrane form of p75TNFR. The icp75TNFR binds the tumor necrosis factor(TNF) and mediates intracellular signaling. Overexpression of the icp75TNFR cDNA results in TNF-induced activation of NFkappaB in a TNF receptor-associated factor 2 (TRAF2)-dependent manner. Thus, our results provide an example for intracellular cytokine receptor activation. PMID- 11279197 TI - Glutathione S-transferase P1-1 (GSTP1-1) inhibits c-Jun N-terminal kinase (JNK1) signaling through interaction with the C terminus. AB - c-Jun N-terminal kinase (JNK)-mediated cell signaling pathways are regulated endogenously in part by protein-protein interactions with glutathione S transferase P1-1 (GSTP1-1) (). Using purified recombinant proteins, combined with fluorescence resonance energy transfer technology, we have found that the C terminus of JNK is critical to the interaction with GSTP1-1. The apparent K(d) for full-length JNK was 188 nm and for a C-terminal fragment (residues 200-424) 217 nm. An N-terminal fragment (residues 1-206) did not bind to GSTP1-1. Increased expression of the C-terminal JNK fragment in a tetracycline-inducible transfected NIH3T3 cell line produced a concentration-dependent increase in the kinase activity of JNK under normal, unstressed growth conditions indicating a dominant-negative effect. This suggests that the fragment can compete with endogenous full-length functional JNK resulting in dissociation of the GSTP1-1 JNK interaction and concomitant JNK enzyme activation. By using an antibody to hemagglutinin-tagged C-JNK, a concentration-dependent co-immunoprecipitation of GSTP1-1 was achieved. These data provide evidence for direct interactions between the C-terminal of JNK and GSTP1-1 and a rationale for considering GSTP1-1 as a critical ligand-binding protein with a role in regulating kinase pathways. PMID- 11279198 TI - The heterogeneous nuclear ribonucleoproteins I and K interact with a subset of the ro ribonucleoprotein-associated Y RNAs in vitro and in vivo. AB - The hY RNAs are a group of four small cytoplasmic RNAs of unknown function that are stably associated with at least two proteins, Ro60 and La, to form Ro ribonucleoprotein complexes. Here we show that the heterogeneous nuclear ribonucleoproteins (hnRNP) I and K are able to associate with a subset of hY RNAs in vitro and demonstrate these interactions to occur also in vivo in a yeast three-hybrid system. Experiments performed in vitro and in vivo with deletion mutants of hY1 RNA revealed its pyrimidine-rich central loop to be involved in interactions with both hnRNP I and K and clearly showed their binding sites to be different from the Ro60 binding site. Both hY1 and hY3 RNAs coprecipitated with hnRNP I in immunoprecipitation experiments performed with HeLa S100 extracts and cell extracts from COS-1 cells transiently transfected with VSV-G-tagged hnRNP-I, respectively. Furthermore, both anti-Ro60 and anti-La antibodies coprecipitated hnRNP I, whereas coprecipitation of hnRNP K was not observed. Taken together, these data strongly suggest that hnRNP I is a stable component of a subpopulation of Ro RNPs, whereas hnRNP K may be transiently bound or interact only with (rare) Y RNAs that are devoid of Ro60 and La. Given that functions related to translation regulation have been assigned to both proteins and also to La, our findings may provide novel clues toward understanding the role of Y RNAs and their respective RNP complexes. PMID- 11279200 TI - Identification of molecular determinants that are important in the assembly of N methyl-D-aspartate receptors. AB - To determine which domains of the N-methyl-d-aspartate (NMDA) receptor are important for the assembly of functional receptors, a number of N- and C-terminal truncations of the NR1a subunit have been produced. Truncations containing a complete ligand binding domain bound glycine antagonist and gave binding constants similar to those of the native subunit, suggesting they were folding to form antagonist binding sites. Since NR2A is not transported to the cell surface unless it is associated with NR1 (McIlhinney, R. A. J., Le Bourdelles, B., Tricuad, N., Molnar, E., Streit, P., and Whiting, P. J. (1998) Neuropharmacology 37, 1355-1367), surface expression of NR2A can be used to monitor the association of the subunits. There was progressive loss of NR2A cell surface expression as the N terminus of NR1a was shortened, with complete loss when truncated beyond residue 380. Removal of the C terminus and/or the last transmembrane domain did not affect NR2A surface expression. Similar results were obtained in co immunoprecipitation experiments. The oligomerization status of the co-expressed NR1a constructs and NR2A subunits was investigated using a non-denaturing gel electrophoresis system (blue native-polyacrylamide gel electrophoresis) and sucrose density gradient centrifugation. The blue native-polyacrylamide gel electrophoresis system also showed that the NR1a subunits could form a homodimer, which was confirmed using soluble constructs of the NR1a subunit. Together these results suggest the residues N-terminal of residue 380 are important for the association of NR2A with NR1a and that the complete N-terminal domain of the NR1a subunit is required for oligomerization with NR2A. PMID- 11279199 TI - The interaction of Src and RACK1 is enhanced by activation of protein kinase C and tyrosine phosphorylation of RACK1. AB - RACK1 is an intracellular receptor for the serine/ threonine protein kinase C. Previously, we demonstrated that RACK1 also interacts with the Src protein tyrosine kinase. RACK1, via its association with these protein kinases, may play a key role in signal transduction. To further characterize the Src-RACK1 interaction and to analyze mechanisms by which cross-talk occurs between the two RACK1-linked signaling kinases, we identified sites on Src and RACK1 that mediate their binding, and factors that regulate their interaction. We found that the interaction of Src and RACK1 is mediated, in part, by the SH2 domain of Src and by phosphotyrosines in the sixth WD repeat of RACK1, and is enhanced by serum or platelet-derived growth factor stimulation, protein kinase C activation, and tyrosine phosphorylation of RACK1. To the best of our knowledge, this is the first report of tyrosine phosphorylation of a member of the WD repeat family of proteins. We think that tyrosine phosphorylation of these proteins is an important mechanism of signal transduction in cells. PMID- 11279201 TI - Identification of reelin-induced sites of tyrosyl phosphorylation on disabled 1. AB - The study of mice with spontaneous and targeted mutations has uncovered a signaling pathway that controls neuronal positioning during mammalian brain development. Mice with disruptions in reelin, dab1, or both vldlr and apoER2 are ataxic, and they exhibit severe lamination defects within several brain structures. Reelin is a secreted extracellular protein that binds to the very low density lipoprotein receptor and the apolipoprotein E receptor 2 on the surface of neurons. Disabled-1 (Dab1), an intracellular adapter protein containing a PTB (phosphotyrosine binding) domain, is tyrosyl-phosphorylated during embryogenesis, but it accumulates in a hypophosphorylated form in mice lacking Reelin or both very low density lipoprotein receptor and apolipoprotein E receptor 2. Dab1 is rapidly phosphorylated when neurons isolated from embryonic brains are stimulated with Reelin, and several tyrosines have been implicated in this response. Mice with phenylalanine substitutions of all five tyrosines (Tyr(185), Tyr(198), Tyr(200), Tyr(220), and Tyr(232)) exhibit a reeler phenotype, implying that tyrosine phosphorylation is critical for Dab1 function. Here we report that, although Src can phosphorylate all five tyrosines in vitro, Tyr(198) and Tyr(220) represent the major sites of Reelin-induced Dab1 phosphorylation in embryonic neurons. PMID- 11279202 TI - Protein kinase A phosphorylates hepatocyte nuclear factor-6 and stimulates glucose-6-phosphatase catalytic subunit gene transcription. AB - Glucose-6-phosphatase is a multicomponent system that catalyzes the terminal step in gluconeogenesis. To examine the effect of the cAMP signal transduction pathway on expression of the gene encoding the mouse glucose-6-phosphatase catalytic subunit (G6Pase), the liver-derived HepG2 cell line was transiently co transfected with a series of G6Pase-chloramphenicol acetyltransferase fusion genes and an expression vector encoding the catalytic subunit of cAMP-dependent protein kinase A (PKA). PKA markedly stimulated G6Pase-chloramphenicol acetyltransferase fusion gene expression, and mutational analysis of the G6Pase promoter revealed that multiple cis-acting elements were required for this response. One of these elements was mapped to the G6Pase promoter region between 114 and -99, and this sequence was shown to bind hepatocyte nuclear factor (HNF) 6. This HNF-6 binding site was able to confer a stimulatory effect of PKA on the expression of a heterologous fusion gene; a mutation that abolished HNF-6 binding also abolished the stimulatory effect of PKA. Further investigation revealed that PKA phosphorylated HNF-6 in vitro. Site-directed mutation of three consensus PKA phosphorylation sites in the HNF-6 carboxyl terminus markedly reduced this phosphorylation. These results suggest that the stimulatory effect of PKA on G6Pase fusion gene transcription in HepG2 cells may be mediated in part by the phosphorylation of HNF-6. PMID- 11279203 TI - Molecular determinants underlying the formation of stable intracellular G protein coupled receptor-beta-arrestin complexes after receptor endocytosis*. AB - beta-Arrestins bind agonist-activated G protein-coupled receptors (GPCRs) and mediate their desensitization and internalization. Although beta-arrestins dissociate from some receptors at the plasma membrane, such as the beta2 adrenergic receptor, they remain associated with other GPCRs and internalize with them into endocytic vesicles. Formation of stable receptor-beta-arrestin complexes that persist inside the cell impedes receptor resensitization, and the aberrant formation of these complexes may play a role in GPCR-based diseases (Barak, L. S., Oakley, R. H., Laporte, S. A., and Caron, M. G. (2001) Proc. Natl. Acad. Sci. U. S. A. 98, 93-98). Here, we investigate the molecular determinants responsible for sustained receptor/beta-arrestin interactions. We show in real time and in live human embryonic kidney (HEK-293) cells that a beta-arrestin-2 green fluorescent protein conjugate internalizes into endocytic vesicles with agonist-activated neurotensin-1 receptor, oxytocin receptor, angiotensin II type 1A receptor, and substance P receptor. Using receptor mutagenesis, we demonstrate that the ability of beta-arrestin to remain associated with these receptors is mediated by specific clusters of serine and threonine residues located in the receptor carboxyl-terminal tail. These clusters are remarkably conserved in their position within the carboxyl-terminal domain and serve as primary sites of agonist-dependent receptor phosphorylation. In addition, we identify a beta arrestin mutant with enhanced affinity for the agonist-activated beta2-adrenergic receptor that traffics into endocytic vesicles with receptors that lack serine/threonine clusters and normally dissociate from wild-type beta-arrestin at the plasma membrane. By identifying receptor and beta-arrestin residues critical for the formation of stable receptor-beta-arrestin complexes, these studies provide novel targets for regulating GPCR responsiveness and treating diseases resulting from abnormal GPCR/beta-arrestin interactions. PMID- 11279204 TI - Inhibitory effect of bovine milk lactoferrin on the interaction between a streptococcal surface protein antigen and human salivary agglutinin. AB - Human whole saliva induces aggregation of Streptococcus mutans cells via an interaction between a surface protein antigen (PAc) of the organism and salivary agglutinin. Bovine milk inhibits the saliva-induced aggregation of S. mutans. In this study, the milk component that possesses inhibitory activity against this aggregation was isolated and found to be lactoferrin. Surface plasmon resonance analysis indicated that bovine lactoferrin binds more strongly to salivary agglutinin, especially to high molecular mass glycoprotein, which is a component of the agglutinin, than to recombinant PAc. The binding of bovine lactoferrin to salivary agglutinin was thermostable, and the optimal pH for binding was 4.0. To identify the saliva-binding region of bovine lactoferrin, 11 truncated bovine lactoferrin fragments were constructed. A fragment corresponding to the C terminal half of the lactoferrin molecule had a strong inhibitory effect on the saliva-induced aggregation of S. mutans, whereas a fragment corresponding to the N-terminal half had a weak inhibitory effect. Seven shorter fragments corresponding to lactoferrin residues 473-538 also showed a high ability to inhibit the aggregation of S. mutans. These results suggest that residues 473-538 of bovine lactoferrin are important in the inhibition of saliva-induced aggregation of S. mutans. PMID- 11279205 TI - Substrate recognition by UDP-galactose and CMP-sialic acid transporters. Different sets of transmembrane helices are utilized for the specific recognition of UDP-galactose and CMP-sialic acid. AB - Human UDP-galactose transporter (hUGT1) and CMP-sialic acid transporter (hCST) are related Golgi membrane proteins with 10 transmembrane helices. We have constructed chimeras between these proteins in order to identify submolecular regions responsible for the determination of substrate specificity. To assess the UGT and CST activities, chimeric cDNAs were transiently expressed in either UGT deficient mutant Lec8 cells or CST-deficient mutant Lec2 cells, and the binding of plant lectins, GS-II or PNA, respectively, to these cells was examined. During the course of analysis of various chimeric transporters, we found that chimeras whose submolecular regions contained helices 1, 8, 9, and 10, and helices 2, 3, and 7 derived from hUGT1 and hCST sequences, respectively, exhibited both UGT and CST activities. The dual substrate specificity for UDP-galactose and CMP-sialic acid of one such representative chimera was directly confirmed by in vitro measurement of the nucleotide sugar transport activity using a heterologous expression system in the yeast Saccharomyces cerevisiae. These findings indicated that the regions which are critical for determining the substrate specificity of UGT and CST resided in different submolecular sites in the two transporters, and that these different determinants could be present within one protein without interfering with each other's function. PMID- 11279206 TI - PTEN 2, a Golgi-associated testis-specific homologue of the PTEN tumor suppressor lipid phosphatase. AB - The tumor suppressor PTEN is a phosphatidylinositol phospholipid phosphatase, which indirectly down-regulates the activity of the protein kinase B/Akt survival kinases. Examination of sequence data bases revealed the existence of a highly conserved homologue of PTEN. This homologue, termed PTEN 2, contained an extended amino-terminal domain having four potential transmembrane motifs, a lipid phosphatase domain, and a potential lipid-binding C2 domain. Transcript analysis demonstrated that PTEN 2 is expressed only in testis and specifically in secondary spermatocytes. In contrast to PTEN, PTEN 2 was localized to the Golgi apparatus via the amino-terminal membrane-spanning regions. Molecular modeling suggested that PTEN 2 is a phospholipid phosphatase with potential specificity for the phosphate at the 3 position of inositol phosphates. Enzymatic analysis of PTEN 2 revealed substrate specificity that is similar to PTEN, with a preference for the dephosphorylation of the phosphatidylinositol 3,5-phosphate phospholipid, a known mediator of vesicular trafficking. Together, these data suggest that PTEN 2 is a Golgi-localized, testis-specific phospholipid phosphatase, which may contribute to the terminal stages of spermatocyte differentiation. PMID- 11279207 TI - Involvement of hematopoietic progenitor kinase 1 in T cell receptor signaling. AB - Hematopoietic progenitor kinase 1 (HPK1), a mammalian Ste20-related serine/threonine protein kinase, is a hematopoietic-specific upstream activator of the c-Jun N-terminal kinase. Here, we provide evidence to demonstrate the involvement of HPK1 in T cell receptor (TCR) signaling. HPK1 was activated and tyrosine-phosphorylated with similar kinetics following TCR/CD3 or pervanadate stimulation. Co-expression of protein-tyrosine kinases, Lck and Zap70, with HPK1 led to HPK1 activation and tyrosine phosphorylation in transfected mammalian cells. Upon TCR/CD3 stimulation, HPK1 formed inducible complexes with the adapters Nck and Crk with different kinetics, whereas it constitutively interacted with the adapters Grb2 and CrkL in Jurkat T cells. Interestingly, HPK1 also inducibly associated with linker for activation of T cells (LAT) through its proline-rich motif and translocated into glycolipid-enriched microdomains (also called lipid rafts) following TCR/CD3 stimulation, suggesting a critical role for LAT in the regulation of HPK1. Together, these results identify HPK1 as a new component of TCR signaling. T cell-specific signaling molecules Lck, Zap70, and LAT play roles in the regulation of HPK1 during TCR signaling. Differential complex formation between HPK1 and adapters highlights the possible involvement of HPK1 in multiple signaling pathways in T cells. PMID- 11279208 TI - The yeast plasma membrane protein Alr1 controls Mg2+ homeostasis and is subject to Mg2+-dependent control of its synthesis and degradation. AB - The Saccharomyces cerevisiae ALR1 (YOL130w) gene product Alr1p is the first known candidate for a Mg(2+) transport system in eukaryotic cells and is distantly related to the bacterial CorA Mg(2+) transporter family. Here we provide the first experimental evidence for the location of Alr1p in the yeast plasma membrane and for the tight control of its expression and turnover by Mg(2+). Using well characterized npi1 and end3 mutants deficient in the endocytic pathway, we demonstrate that Alr1 protein turnover is dependent on ubiquitination and endocytosis. Furthermore, cells lacking the vacuolar protease Pep4p accumulated Alr1p in the vacuole. Mutants lacking Alr1p (Deltaalr1) showed a 60% reduction of total intracellular Mg(2+) compared with the wild type and failed to grow in standard media. When starved of Mg(2+), mutant and wild-type cells had similar low levels of intracellular Mg(2+); but upon addition of Mg(2+), wild type cells replenished the intracellular Mg(2+) pool within a few hours, whereas Deltaalr1 mutant cells did not. Expression of the bacterial Mg(2+) transporter CorA in the yeast Deltaalr1 mutant partially restored growth in standard media. The results are discussed in terms of Alr1p being a plasma membrane transporter with high selectivity for Mg(2+). PMID- 11279209 TI - A dynamic role for HDAC7 in MEF2-mediated muscle differentiation. AB - The overlapping expression profile of MEF2 and the class-II histone deacetylase, HDAC7, led us to investigate the functional interaction and relationship between these regulatory proteins. HDAC7 expression inhibits the activity of MEF2 (-A, C, and -D), and in contrast MyoD and Myogenin activities are not affected. Glutathione S-transferase pulldown and immunoprecipitation demonstrate that the repression mechanism involves direct interactions between MEF2 proteins and HDAC7 and is associated with the ability of MEF2 to interact with the N-terminal 121 amino acids of HDAC7 that encode repression domain 1. The MADS domain of MEF2 mediates the direct interaction of MEF2 with HDAC7. MEF2 inhibition by HDAC7 is dependent on the N-terminal repression domain and surprisingly does not involve the C-terminal deacetylase domain. HDAC7 interacts with CtBP and other class-I and -II HDACs suggesting that silencing of MEF2 activity involves corepressor recruitment. Furthermore, we show that induction of muscle differentiation by serum withdrawal leads to the translocation of HDAC7 from the nucleus into the cytoplasm. This work demonstrates that HDAC7 regulates the function of MEF2 proteins and suggests that this class-II HDAC regulates this important transcriptional (and pathophysiological) target in heart and muscle tissue. The nucleocytoplasmic trafficking of HDAC7 and other class-II HDACs during myogenesis provides an ideal mechanism for the regulation of HDAC targets during mammalian development and differentiation. PMID- 11279210 TI - Sequential involvement of p115, SNAREs, and Rab proteins in intra-Golgi protein transport. AB - Delivery of transport vesicles to their receptor compartment involves tethering, priming, and fusion. Soluble NSF attachment protein-alpha (alphaSNAP) mediates the disruption of SNAREs by N-ethylmaleimide sensitive factor (NSF) and was employed to determine the hierarchy of proteins responsible for intra-Golgi protein transport. The N-terminal 23 amino acids of alphaSNAP are necessary for SNARE binding. The antibody 2F10 recognizes this SNARE interaction domain of alphaSNAP and inhibits intra-Golgi protein transport reversibly. This antibody was applied to modify the transport assay to determine the protein requirements relative to the action of alphaSNAP and NSF. We found that 1) p115 acts independently of alphaSNAP and NSF, 2) SNAREs are required after tethering and interact selectively after activation by alphaSNAP and NSF, and 3) Rab proteins act after SNARE activation and before fusion. PMID- 11279211 TI - Biochemical, biophysical, and functional characterization of bacterially expressed and refolded receptor binding domain of Plasmodium vivax duffy-binding protein. AB - Invasion of erythrocytes by malaria parasites is mediated by specific molecular interactions. Plasmodium vivax is completely dependent on interaction with the Duffy blood group antigen to invade human erythrocytes. The P. vivax Duffy binding protein, which binds the Duffy antigen during invasion, belongs to a family of erythrocyte-binding proteins that also includes Plasmodium falciparum sialic acid binding protein and Plasmodium knowlesi Duffy binding protein. The receptor binding domains of these proteins lie in a conserved, N-terminal, cysteine-rich region, region II, found in each of these proteins. Here, we have expressed P. vivax region II (PvRII), the P. vivax Duffy binding domain, in Escherichia coli. Recombinant PvRII is incorrectly folded and accumulates in inclusion bodies. We have developed methods to refold and purify recombinant PvRII in its functional conformation. Biochemical, biophysical, and functional characterization confirms that recombinant PvRII is pure, homogeneous, and functionally active in that it binds Duffy-positive human erythrocytes with specificity. Refolded PvRII is highly immunogenic and elicits high titer antibodies that can inhibit binding of P. vivax Duffy-binding protein to erythrocytes, providing support for its development as a vaccine candidate for P. vivax malaria. Development of methods to produce functionally active recombinant PvRII is an important step for structural studies as well as vaccine development. PMID- 11279212 TI - A water-soluble homodimeric serine palmitoyltransferase from Sphingomonas paucimobilis EY2395T strain. Purification, characterization, cloning, and overproduction. AB - Serine palmitoyltransferase (SPT, EC ) is a key enzyme in sphingolipid biosynthesis and catalyzes the decarboxylative condensation of l-serine and palmitoyl-coenzyme A to 3-ketodihydrosphingosine. We found that the Gram-negative obligatory aerobic bacteria Sphingomonas paucimobilis EY2395(T) have significant SPT activity and purified SPT to homogeneity. This enzyme is a water-soluble homodimeric protein unlike eukaryotic enzymes, known as heterodimers composed of tightly membrane-bound subunits, named LCB1 and LCB2. The purified SPT shows an absorption spectrum characteristic of a pyridoxal 5'-phosphate-dependent enzyme. The substrate specificity of the Sphingomonas SPT is less strict than the SPT complex from Chinese hamster ovary cells. We isolated the SPT gene encoding 420 amino acid residues (M(r) 45,041) and succeeded in overproducing the SPT protein in Escherichia coli, in which the product amounted to about 10-20% of the total protein of the cell extract. Sphingomonas SPT shows about 30% homology with the enzymes of the alpha-oxamine synthase family, and amino acid residues supposed to be involved in catalysis are conserved. The recombinant SPT was catalytically and spectrophotometrically indistinguishable from the native enzyme. This is the first successful overproduction of an active enzyme in the sphingolipid biosynthetic pathway. Sphingomonas SPT is a prototype of the eukaryotic enzyme and would be a useful model to elucidate the reaction mechanism of SPT. PMID- 11279213 TI - Yersinia enterocolitica YopP-induced apoptosis of macrophages involves the apoptotic signaling cascade upstream of bid. AB - Yersinia enterocolitica induces apoptosis in macrophages by injecting the plasmid encoded YopP (YopJ in other Yersinia species). Recently it was reported that YopP/J is a member of an ubiquitin-like protein cysteine protease family and that the catalytic core of YopP/J is required for its inhibition of the MAPK and NF kappaB pathways. Here we analyzed the YopP/J-induced apoptotic signaling pathway. YopP-mediated cell death could be inhibited by addition of the zVAD caspase inhibitor, but not by DEVD or YVAD. Generation of truncated Bid (tBid) was the first apoptosis-related event that we observed. The subsequent translocation of tBid to the mitochondria induced the release of cytochrome c, leading to the activation of procaspase-9 and the executioner procaspases-3 and -7. Inhibition of the postmitochondrial executioner caspases-3 and -7 did not affect Bid cleavage. Bid cleavage could not be observed in a yopP-deficient Y. enterocolitica strain, showing that this event requires YopP. Disruption of the catalytic core of YopP abolished the rapid generation of tBid, thereby hampering induction of apoptosis by Y. enterocolitica. This finding supports the idea that YopP/J induces apoptosis by directly acting on cell death pathways, rather than being the mere consequence of gene induction inhibition in combination with microbial stimulation of the macrophage. PMID- 11279214 TI - Differential functions of members of the low density lipoprotein receptor family suggested by their distinct endocytosis rates. AB - The low density lipoprotein receptor (LDLR) family is composed of a class of cell surface endocytic receptors that recognize extracellular ligands and internalize them for degradation by lysosomes. In addition to LDLR, mammalian members of this family include the LDLR-related protein (LRP), the very low density lipoprotein receptor (VLDLR), the apolipoprotein E receptor-2 (apoER2), and megalin. Herein we have analyzed the endocytic functions of the cytoplasmic tails of these receptors using LRP minireceptors, its chimeric receptor constructs, and full length VLDLR and apoER2 stably expressed in LRP-null Chinese hamster ovary cells. We find that the initial endocytosis rates mediated by different cytoplasmic tails are significantly different, with half-times of ligand internalization ranging from less than 30 s to more than 8 min. The tail of LRP mediates the highest rate of endocytosis, whereas those of the VLDLR and apoER2 exhibit least endocytosis function. Compared with the tail of LRP, the tails of the LDLR and megalin display significantly lower levels of endocytosis rates. Ligand degradation analyses strongly support differential endocytosis rates initiated by these receptors. Interestingly apoER2, which has recently been shown to mediate intracellular signal transduction, exhibited the lowest level of ligand degradation efficiency. These results thus suggest that the endocytic functions of members of the LDLR family are distinct and that certain receptors in this family may play their main roles in areas other than receptor-mediated endocytosis. PMID- 11279215 TI - Production of human type I collagen in yeast reveals unexpected new insights into the molecular assembly of collagen trimers. AB - Substantial evidence supports the role of the procollagen C-propeptide in the initial association of procollagen polypeptides and for triple helix formation. To evaluate the role of the propeptide domains on triple helix formation, human recombinant type I procollagen, pN-collagen (procollagen without the C propeptides), pC-collagen (procollagen without the N-propeptides), and collagen (minus both propeptide domains) heterotrimers were expressed in Saccharomyces cerevisiae. Deletion of the N- or C-propeptide, or both propeptide domains, from both proalpha-chains resulted in correctly aligned triple helical type I collagen. Protease digestion assays demonstrated folding of the triple helix in the absence of the N- and C-propeptides from both proalpha-chains. This result suggests that sequences required for folding of the triple helix are located in the helical/telopeptide domains of the collagen molecule. Using a strain that does not contain prolyl hydroxylase, the same folding mechanism was shown to be operative in the absence of prolyl hydroxylase. Normal collagen fibrils were generated showing the characteristic banding pattern using this recombinant collagen. This system offers new opportunities for the study of collagen expression and maturation. PMID- 11279216 TI - The viral RNA polymerase H4L subunit is required for Vaccinia virus early gene transcription termination. AB - Vaccinia virus early gene transcription is catalyzed by a multisubunit virion form of RNA polymerase that possesses a unique subunit, H4L. Prior studies from this laboratory showed that the NH(2)-terminal domain of H4L, containing amino acids 1-195, interacts with the COOH-terminal end of nucleoside triphosphate phosphohydrolase I (NPH I), an ATPase that is employed in early gene transcription termination. Carboxyl-terminal deletion mutations of NPH I lose both the ability to mediate transcription termination and binding to H4L, providing evidence that the interaction between NPH I and H4L is required for termination. In order to test this model further, antibodies raised against segments of H4L were tested for their ability to inhibit transcription termination in vitro. A bead-bound template was employed in these studies, which permitted us to separate transcription initiation from elongation and termination. Antibodies raised against H4L amino acids 1-256 inhibited termination in an in vitro assay using virus-infected cell extracts lacking NPH I, but antibodies raised against H4L amino acids 568-795 did not. Preincubation of anti-H4L(1-256) antibodies with H4L fragments 1-256 or 1-195 prevented antibody inhibition of termination, demonstrating that inhibition was mediated by antibody binding to one or more epitopes in the NH(2)-terminal end of H4L. Antibody inhibition of termination is reduced in wild type virus-infected cell extracts containing NPH I. Furthermore, preincubation of a NPH I minus cell extract with NPH I prior to antibody addition, or readdition of NPH I to isolated ternary complexes prepared in the absence of NPH I, prevented antibody inhibition of transcription termination. These data show that NPH I and the inhibitory antibodies compete for a binding site(s) on H4L, providing further evidence that the H4L subunit of the vaccinia virus RNA polymerase plays a direct role in transcription termination. PMID- 11279217 TI - Cholesterol biosynthesis from lanosterol. A concerted role for Sp1 and NF-Y binding sites for sterol-mediated regulation of rat 7-dehydrocholesterol reductase gene expression. AB - The 7-dehydrocholesterol reductase (Dhcr7) is the terminal enzyme in the pathway of cholesterol biosynthesis. We have previously reported that sterol depletion in vivo caused a significant induction of both liver mRNA and enzyme activity of Dhcr7 (Bae, S.-H., Lee, J. N., Fitzky, B. U., Seong, J., and Paik, Y.-K. (1999) J. Biol. Chem. 274, 14624-14631). In this paper, we also observed liver cell specific sterol-mediated Dhcr7 gene induction in vitro by sterol depletion in rat hepatoma cells, suggesting the presence of sterol-mediated regulatory elements in the Dhcr7 gene. To understand the mechanisms responsible for regulating Dhcr7 expression, we have isolated the 5'-flanking region of the gene encoding rat Dhcr7 and have characterized the potential regulatory elements of the gene that are responsible for sterol-mediated regulation. The Dhcr7 promoter contains binding sites for Sp1 (at -177, -172, -125, and -20), NF-Y (at -88 and -51), and SREBP-1 or ADD1 (at -33). Deletion analysis of the Dhcr7 gene promoter ( 1053/+31), employing a nested series of Dhcr7-luciferase constructs, demonstrated that the -179 upstream region of the gene is necessary and sufficient for optimal efficient sterol-regulated transcription. DNase I footprinting and electrophoretic mobility shift assay showed that the SRE1/E box (-33/-22) involved in sterol response of many sterol-related enzyme genes was protected specifically by the overexpressed recombinant ADD1. Mutational analysis for the functional relationship between the identified cis-elements in this region indicate that one of the binding sites for Sp1 (GC box at -125) and NF-Y (CCAAT box at -88) plays a cooperative role in the sterol-mediated activation, in which the latter site also acts as a co-regulator for SREBP-activated Dhcr7 promoter activity. We believe that Dhcr7 is the first enzyme characterized with a sterol regulatory function in the post-lanosterol pathway. This may be important for understanding the coordinated control of cholesterol biosynthesis as well as the molecular mechanism of Smith-Lemli-Opitz syndrome-related protein in mammals. PMID- 11279218 TI - Cellular FLICE-inhibitory protein splice variants inhibit different steps of caspase-8 activation at the CD95 death-inducing signaling complex. AB - Upon stimulation, CD95 (APO-1/Fas) recruits the adapter molecule FADD/MORT1, procaspase-8, and the cellular FLICE-inhibitory proteins (c-FLIP) into the death inducing signaling complex (DISC). According to the induced proximity model, procaspase-8 is activated in the DISC in an autoproteolytic manner by two subsequent cleavage steps. c-FLIP proteins exist as a long (c-FLIP(L)) and a short (c-FLIP(S)) splice variant, both of them capable of protecting cells from death receptor-mediated apoptosis. In stably transfected BJAB cells, both c FLIP(S) and c-FLIP(L) block procaspase-8 activation at the DISC. However, cleavage is blocked at different steps. c-FLIP(L) allows the first cleavage step of procaspase-8, leading to the generation of the p10 subunit. In contrast, c FLIP(S) completely inhibits cleavage of procaspase-8. Interestingly, p43-c FLIP(L) lacking the p12 subunit also prevents cleavage of procaspase-8. In contrast, a nonprocessable mutant of c-FLIP(L) allows the first cleavage of procaspase-8. In conclusion, both c-FLIP proteins prevent caspase-8 activation at different levels of procaspase-8 processing at the DISC. Our results indicate that c-FLIP(L) induces a conformation of procaspase-8 that allows partial but not complete proteolytical processing, whereas in contrast c-FLIP(S) even prevents partial procaspase-8 activation at the DISC. PMID- 11279219 TI - A point mutation in the LIM domain of Lhx3 reduces activation of the glycoprotein hormone alpha-subunit promoter. AB - Lhx3, a member of the LIM homeodomain family of transcription factors, is required for development of the pituitary in mice. A recent report has described a point mutation in the human LHX3 gene that is associated with a combined pituitary hormone disorder. The mutation is predicted to lead to the replacement of a tyrosine residue with a cysteine in the second LIM domain of LHX3. We have characterized the effects of this point mutation (Y114C) when analyzed in the context of the mouse Lhx3 coding sequence. Mobility shift assays demonstrated that the Lhx3 Y114C mutant is capable of binding DNA, although a decrease in the formation of a specific complex was observed. Transfection assays using an expression vector for either full-length Lhx3 or a GAL4-Lhx3 LIM domain fusion provided evidence that the Lhx3 Y114C mutant has a decreased ability to stimulate transcription. In particular, a GAL4-Lhx3 Y114C LIM mutant was unable to support Ras responsiveness of a modified glycoprotein hormone alpha-subunit reporter gene. Protein interaction studies suggest that the Y114C mutation may modestly reduce binding to the POU transcription factor, Pit-1. Interestingly, the Y114C mutation essentially abrogated binding to the putative co-activator/adapter, selective LIM-binding protein. The findings provide insights into the mechanisms mediating transcriptional activation by Lhx3 and suggest that the observed phenotype of the human mutation probably involves reduced transcriptional activity of the mutant LHX3. PMID- 11279220 TI - Transcription factor Rho does not require a free end to act as an RNA-DNA helicase on an RNA. AB - Escherichia coli Rho factor is a ring-shaped, homohexameric protein that terminates synthesis of RNA through interactions with the nascent RNA transcript. Because its mechanism of action may involve translocation of the RNA transcript through the hole in its ring structure, its action could depend on the availability of a free 5' terminus. To determine whether Rho's activity is 5'-end dependent, its ability to bind to and function on a circular derivative of lambda cro mRNA was investigated. The circular derivative was made in vitro by action of RNA ligase on a derivative of lambda cro RNA containing an extra 10-nucleotide sequence near the 5'-end that was complementary to a sequence located near the 3' end. Rho bound nearly as tightly to the circular derivative RNA as to the standard cro transcript. Rho was also able to readily dissociate a DNA oligonucleotide from its helical complex with the circular RNA in an ATP dependent reaction. Thus, the action of Rho on a transcript does not depend on the availability of a free 5' terminus. PMID- 11279221 TI - A Chlamydia trachomatis UDP-N-acetylglucosamine acyltransferase selective for myristoyl-acyl carrier protein. Expression in Escherichia coli and formation of hybrid lipid A species. AB - Chlamydia trachomatis lipid A is unusual in that it is acylated with myristoyl chains at the glucosamine 3 and 3' positions. We have cloned and expressed the gene encoding UDP-N-acetylglucosamine 3-O-acyltransferase of C. trachomatis (CtlpxA), the first enzyme of lipid A biosynthesis. C. trachomatis LpxA displays approximately 20-fold selectivity for myristoyl-ACP over R/S-3-hydroxymyristoyl ACP under standard assay conditions, consistent with the proposed structure of C. trachomatis lipid A. CtLpxA is the first reported UDP-N-acetylglucosamine acyltransferase that prefers a non-hydroxylated acyl-ACP to a hydroxyacyl-ACP. When CtlpxA was expressed in RO138, a temperature-sensitive lpxA mutant of Escherichia coli, five new hybrid lipid A species were made in vivo after 2 h at 42 degrees C, in place of Escherichia coli lipid A. These compounds were purified and analyzed by matrix-assisted laser desorption ionization/time of flight mass spectrometry. In each case, a myristoyl chain replaced one or both of the ester linked 3-hydroxymyristoyl residues of E. coli lipid A. With prolonged growth at 42 degrees C, all the ester-linked 3-hydroxymyristoyl residues were replaced with myristate chains. Re-engineering the structure of E. coli lipid A should facilitate the microbiological production of novel agonists or antagonists of the innate immunity receptor TLR-4, with possible uses as adjuvants or anti inflammatory agents. PMID- 11279222 TI - Gbeta gamma mediate differentiation of vascular smooth muscle cells. AB - Proliferation and subsequent dedifferentiation of vascular smooth muscle (VSM) cells contribute to the pathogenesis of atherosclerosis and postangioplastic restenosis. The dedifferentiation of VSM cells in vivo or in cell culture is characterized by a loss of contractile proteins such as smooth muscle-specific alpha-actin and myosin heavy chain (SM-MHC). Serum increased the expression of contractile proteins in neonatal rat VSM cells, indicating a redifferentiation process. RNase protection assays defined thrombin as a serum component that increases the abundance of SM-MHC transcripts. Additionally, serum and thrombin transiently elevated cytosolic Ca(2+) concentrations, led to a biphasic extracellular signal-regulated kinase (ERK) phosphorylation, up-regulated a transfected SM-MHC promoter construct, and induced expression of the contractile proteins SM-MHC and alpha-actin. Pertussis toxin, N17-Ras/Raf, and PD98059 prevented both the serum- and thrombin-induced second phase ERK phosphorylation and SM-MHC promoter activation. Constitutively active Galpha(q), Galpha(i), Galpha(12), and Galpha(13) failed to up-regulate SM-MHC transcription, whereas Gbetagamma concentration-dependently increased the SM-MHC promoter activity. Furthermore, the Gbetagamma scavenger beta-adrenergic receptor kinase 1 C terminal peptide abolished the serum-mediated differentiation. We conclude that receptor-mediated differentiation of VSM cells requires Gbetagamma and an intact Ras/Raf/MEK/ERK signaling. PMID- 11279223 TI - Indirect role for COPI in the completion of FCgamma receptor-mediated phagocytosis. AB - Recent evidence suggests that extension of pseudopods during phagocytosis requires localized insertion of endomembrane vesicles. The nature of these vesicles and the processes mediating their release and insertion are unknown. COPI plays an essential role in the budding and traffic of membrane vesicles in intracellular compartments. We therefore assessed whether COPI is also involved in phagosome formation. We used ldlF cells, a mutant line derived from Chinese hamster ovary cells that express a temperature-sensitive form of epsilonCOP. To confer phagocytic ability to ldlF cells, they were stably transfected with Fc receptors type IIA (FcgammaRIIA). In the presence of functional COPI, FcgammaRIIA transfected ldlF cells effectively internalized opsonized particles. In contrast, phagocytosis was virtually eliminated after incubation at the restrictive temperature. Similar results were obtained impairing COPI function in macrophages using brefeldin A. Notably, loss of COPI function preceded complete inhibition of phagocytosis, suggesting that COPI is indirectly required for phagocytosis. Despite their inability to internalize particles, COPI-deficient cells nevertheless expressed normal levels of FcgammaRIIA, and signal transduction appeared unimpeded. The opsonized particles adhered normally to COPI-deficient cells and were often found on actin-rich pedestals, but they were not internalized due to the inability of the cells to extend pseudopods. The failure to extend pseudopods was attributed to the inability of COPI-deficient cells to mobilize endomembrane vesicles, including a VAMP3-containing compartment, in response to the phagocytic stimulus. PMID- 11279224 TI - CREB-binding protein and p300 in transcriptional regulation. PMID- 11279225 TI - Tracking the light environment by cyanobacteria and the dynamic nature of light harvesting. PMID- 11279226 TI - Blue light sensing in higher plants. PMID- 11279227 TI - The influence of macromolecular crowding and macromolecular confinement on biochemical reactions in physiological media. PMID- 11279228 TI - The phytochromes, a family of red/far-red absorbing photoreceptors. PMID- 11279230 TI - Hemocyanins and invertebrate evolution. PMID- 11279231 TI - Oxygen reduction by nitric-oxide synthases. PMID- 11279232 TI - Signal transduction pathways involved in phosphorylation and activation of p70S6K following exposure to UVA irradiation. AB - Ultraviolet light A (UVA) plays an important role in the etiology of human skin cancer, and UVA-induced signal transduction has a critical role in UVA-induced skin carcinogenesis. The upstream signaling pathways leading to p70(S6K) phosphorylation and activation are not well understood. Here, we observed that UVA induces phosphorylation and activation of p70(S6K). Further, UVA-stimulated p70(S6K) activity and phosphorylation at Thr(389) were blocked by wortmannin, rapamycin, PD98059, SB202190, and dominant negative mutants of phosphatidylinositol (PI) 3-kinase p85 subunit (DNM-Deltap85), ERK2 (DNM-ERK2), p38 kinase (DNM-p38), and JNK1 (DNM-JNK1) and were absent in Jnk1-/- or Jnk2-/- knockout cells. The p70(S6K) phosphorylation at Ser(411) and Thr(421)/Ser(424) was inhibited by rapamycin, PD98059, or DNM-ERK2 but not by wortmannin, SB202190, DNM-Deltap85, or DNM-p38. However, Ser(411), but not Thr(421)/Ser(424) phosphorylation, was suppressed in DNM-JNK1 and abrogated in Jnk1-/- or Jnk2-/- cells. In vitro assays indicated that Ser(411) on immunoprecipitated p70(S6K) proteins is phosphorylated by active JNKs and ERKs, but not p38 kinase, and Thr(421)/Ser(424) is phosphorylated by ERK1, but not ERK2, JNKs, or p38 kinase. Moreover, p70(S6K) co-immunoprecipitated with PI 3-kinase and possibly PDK1. The complex possibly possessed a partial basal level of phosphorylation, but not at MAPK sites, which was available for its activation by MAPKs in vitro. Thus, these results suggest that activation of MAPKs, like PI 3-kinase/mTOR, may be involved in UVA-induced phosphorylation and activation of p70(S6K). PMID- 11279233 TI - The role of the betaDELSEED motif of F1-ATPase: propagation of the inhibitory effect of the epsilon subunit. AB - In F(1)-ATPase, a rotary motor enzyme, the region of the conserved DELSEED motif in the beta subunit moves and contacts the rotor gamma subunit when the nucleotide fills the catalytic site, and the acidic nature of the motif was previously assumed to play a critical role in rotation. Our previous work, however, disproved the assumption (Hara, K. Y., Noji, H., Bald, D., Yasuda, R., Kinosita, K., Jr., and Yoshida, M. (2000) J. Biol. Chem. 275, 14260-14263), and the role of this motif remained unknown. Here, we found that the epsilon subunit, an intrinsic inhibitor, was unable to inhibit the ATPase activity of a mutant thermophilic F(1)-ATPase in which all of the five acidic residues in the DELSEED motif were replaced with alanines, although the epsilon subunit in the mutant F(1)-ATPase assumed the inhibitory form. In addition, the replacement of basic residues in the C-terminal region of the epsilon subunit by alanines caused a decrease of the inhibitory effect. Partial replacement of the acidic residues in the DELSEED motif of the beta subunit or of the basic residues in the C-terminal alpha-helix of the epsilon subunit induced a partial effect. We here conclude that the epsilon subunit exerts its inhibitory effect through the electrostatic interaction with the DELSEED motif of the beta subunit. PMID- 11279234 TI - Mechanism for the reduction of telomerase expression during muscle cell differentiation. AB - Telomerase, the reverse transcriptase that maintains telomere DNA, is usually undetectable in adult human tissues, but is positive in embryonic tissues and in cancers. However, in rodents, several organs of normal adult animals express substantial amounts of telomerase activity. To elucidate relevant control mechanisms operating on the tissue-specific expression of telomerase in rodents, we examined the transcriptional regulation of telomerase reverse transcriptase (mTERT) gene in muscle cell differentiation. Reverse transcriptase-polymerase chain reaction analysis showed that the reduction of telomerase activity was caused by the decrease of mTERT mRNA level during myogenesis. Transfections of mTERT promoter showed that the proximal 225-base pair region is the core promoter responsible for basal transcriptional activity and also participates in the reduced transcription after muscle differentiation. Electrophoretic mobility shift assays showed that this region contained the GC-boxes, which bind to Sp1 family proteins, and the E-box, which binds to c-Myc. Furthermore, DNA binding activities of Sp1, Sp3, and c-Myc were down-regulated during myogenesis. These data suggest that Sp1, Sp3, and c-Myc have critical roles of TERT transactivation in mouse, and the lack of these transcription factors cause down-regulation of mTERT gene expression in muscle cells differentiation. PMID- 11279235 TI - Isolation and characterization of a U-specific 3'-5'-exonuclease from mitochondria of Leishmania tarentolae. AB - We have purified a 3'-5'-exoribonuclease from mitochondrial extract of Leishmania tarentolae over 4000-fold through six column fractionations. This enzyme digested RNA in a distributive manner, showed a high level of specificity for 3'-terminal Us, and was blocked by a terminal dU; there was slight exonucleolytic activity on a 3'-terminal A or C but no activity on a 3'-terminal G residue. The enzyme preferred single-stranded 3'-oligo(U) overhangs and did not digest duplex RNA. Two other 3'-5'-exoribonuclease activities were also detected in the mitochondrial extract, one of which was stimulated by a 3'-phosphate and the other of which degraded RNAs with a 3'-OH to mononucleotides in a processive manner. The properties of the distributive U-specific 3'-5'-exoribonuclease suggest an involvement in the U-deletion RNA editing reaction that occurs in the mitochondrion of these cells. PMID- 11279236 TI - The phosphatidylinositol 3-kinase pathway selectively controls sIL-1RA not interleukin-1beta production in the septic leukocytes. AB - Microbial components such as bacterial endotoxin lipopolysaccharide (LPS) can trigger highly lethal septic shock. The cardinal features of septic leukocytes include the repressed production of inflammatory cytokines, such as interleukin-1 beta (IL-1beta), and elevated production of anti-inflammatory cytokines, such as secretory interleukin-1 receptor antagonist (sIL-1RA). Pro- and anti-inflammatory cytokine gene transcriptions are equally repressed in septic leukocytes due to disruption of the LPS signaling pathway at the level of interleukin-1 receptor associated kinase. The selective elevation of sIL-1RA protein in septic blood is caused by efficient translation of residual sIL-1RA message. In this study, we report that the LPS-inducible phosphatidylinositol 3-kinase (PI3-kinase) dependent signaling pathway contributes to the elevated translation of sIL-1RA in septic/LPS-adapted leukocytes. We also observe that this pathway is gene specific and does not affect the production of proinflammatory IL-1beta protein. PMID- 11279237 TI - Biosynthesis of nucleotide-activated D-glycero-D-manno-heptose. AB - The glycan chain repeats of the S-layer glycoprotein of Aneurinibacillus thermoaerophilus DSM 10155 contain d-glycero-d-manno-heptose, which has also been described as constituent of lipopolysaccharide cores of Gram-negative bacteria. The four genes required for biosynthesis of the nucleotide-activated form GDP-d glycero-d-manno-heptose were cloned, sequenced, and overexpressed in Escherichia coli, and the corresponding enzymes GmhA, GmhB, GmhC, and GmhD were purified to homogeneity. The isomerase GmhA catalyzed the conversion of d-sedoheptulose 7 phosphate to d-glycero-d-manno-heptose 7-phosphate, and the phosphokinase GmhB added a phosphate group to form d-glycero-d-manno-heptose 1,7-bisphosphate. The phosphatase GmhC removed the phosphate in the C-7 position, and the intermediate d-glycero-alpha-d-manno-heptose 1-phosphate was eventually activated with GTP by the pyrophosphorylase GmhD to yield the final product GDP-d-glycero-alpha-d-manno heptose. The intermediate and end products were analyzed by high performance liquid chromatography. Nuclear magnetic resonance spectroscopy was used to confirm the structure of these substances. This is the first report of the biosynthesis of GDP-d-glycero-alpha-d-manno-heptose in Gram-positive organisms. In addition, we propose a pathway for biosynthesis of the nucleotide-activated form of l-glycero-d-manno-heptose. PMID- 11279238 TI - Involvement of a unique carbohydrate-responsive factor in the glucose regulation of rat liver fatty-acid synthase gene transcription. AB - Refeeding carbohydrate to fasted rats induces the transcription of genes encoding enzymes of fatty acid biosynthesis, e.g. fatty-acid synthase (FAS). Part of this transcriptional induction is mediated by insulin. An insulin response element has been described for the fatty-acid synthase gene region of -600 to +65, but the 2 3-fold increase in fatty-acid synthase promoter activity attributable to this region is small compared with the 20-30-fold induction in fatty-acid synthase gene transcription observed in fasted rats refed carbohydrate. We have previously reported that the fatty-acid synthase gene region between -7382 and -6970 was essential for achieving high in vivo rates of gene transcription. The studies of the current report demonstrate that the region of -7382 to -6970 of the fatty acid synthase gene contains a carbohydrate response element (CHO-RE(FAS)) with a palindrome sequence (CATGTGn(5)GGCGTG) that is nearly identical to the CHO-RE of the l-type pyruvate kinase and S(14) genes. The glucose responsiveness imparted by CHO-RE(FAS) was independent of insulin. Moreover, CHO-RE(FAS) conferred glucose responsiveness to a heterologous promoter (i.e. l-type pyruvate kinase). Electrophoretic mobility shift assays demonstrated that CHO-RE(FAS) readily bound a unique hepatic ChoRF and that CHO-RE(FAS) competed with the CHO-RE of the l type pyruvate kinase and S(14) genes for ChoRF binding. In vivo footprinting revealed that fasting reduced and refeeding increased ChoRF binding to CHO RE(FAS). Thus, carbohydrate responsiveness of rat liver fatty-acid synthase appears to require both insulin and glucose signaling pathways. More importantly, a unique hepatic ChoRF has now been shown to recognize glucose responsive sequences that are common to three different genes: fatty-acid synthase, l-type pyruvate kinase, and S(14). PMID- 11279239 TI - Interactions of CCCH zinc finger proteins with mRNA: tristetraprolin-mediated AU rich element-dependent mRNA degradation can occur in the absence of a poly(A) tail. AB - The CCCH family of tandem zinc finger proteins has recently been shown to promote the turnover of certain mRNAs containing class II AU-rich elements (AREs). In the case of one member of this family, tristetraprolin (TTP), absence of the protein in knockout mice leads to stabilization of two mRNAs containing AREs of this type, those encoding tumor necrosis factor alpha (TNFalpha) and granulocyte macrophage colony-stimulating factor. To begin to decipher the mechanism by which these zinc finger proteins stimulate the breakdown of this class of mRNAs, we co transfected TTP and its related CCCH proteins into 293 cells with vectors encoding full-length TNFalpha, granulocyte-macrophage colony-stimulating factor, and interleukin-3 mRNAs. Co-expression of the CCCH proteins caused the rapid turnover of these ARE-containing mRNAs and also promoted the accumulation of stable breakdown intermediates that were truncated at the 3'-end of the mRNA, even further 5' than the 5'-end of the poly(A) tail. To determine whether an intact poly(A) tail was necessary for TTP to promote this type of mRNA degradation, we inserted the TNFalpha ARE into a nonpolyadenylated histone mRNA and also attached a histone 3'-end-processing sequence to the 3'-end of nonpolyadenylated interleukin-3 and TNFalpha mRNAs. In all three cases, TTP stimulated the turnover of the ARE-containing mRNAs, despite the demonstrated absence of a poly(A) tail. These studies indicate that members of this class of CCCH proteins can promote class II ARE-containing mRNA turnover even in the absence of a poly(A) tail, suggesting that the processive removal of the poly(A) tail may not be required for this type of CCCH protein-stimulated mRNA turnover. PMID- 11279240 TI - TatB and TatC form a functional and structural unit of the twin-arginine translocase from Escherichia coli. AB - In Escherichia coli, a subset of periplasmic proteins is exported via the twin arginine translocation (Tat) pathway. In the present study, we have purified the Tat complex from E. coli, and we show that it contains only TatA, TatB, and TatC. Within the purified complex, TatB and TatC are present in a strict 1:1 ratio, suggesting a functional association. This has been confirmed by expression of a translational fusion between TatB and TatC. This Tat(BC) chimera supports efficient Tat-dependent export, indicating that TatB and TatC act as a unit in both structural and functional terms. The purified Tat complex contains varying levels of TatA, suggesting a gradual loss during isolation and a looser association. The molecular mass of the complex is approximately 600 kDa, demonstrating the presence of multiple copies of TatA, B, and C. Co immunoprecipitation experiments show that TatC is required for the interaction of TatA with TatB, suggesting that TatA may interact with the complex via binding to TatC. PMID- 11279241 TI - Nuclear factor kappa B-inducing kinase and Ikappa B kinase-alpha signal skeletal muscle cell differentiation. AB - Nuclear factor kappaB (NF-kappaB)-inducing kinase (NIK), IkappaB kinase (IKK) alpha and -beta, and IkappaBalpha are common elements that signal NF-kappaB activation in response to diverse stimuli. In this study, we analyzed the role of this pathway during insulin-like growth factor II (IGF-II)-induced myoblast differentiation. L6E9 myoblasts differentiated with IGF-II showed an induction of NF-kappaB DNA-binding activity that correlated in time with the activation of IKKalpha, IKKbeta, and NIK. Moreover, the activation of IKKalpha, IKKbeta, and NIK by IGF-II was dependent on phosphatidylinositol 3-kinase, a key regulator of myogenesis. Adenoviral transduction with the IkappaBalpha(S32A/S36A) mutant severely impaired both IGF-II-dependent NF-kappaB activation and myoblast differentiation, indicating that phosphorylation of IkappaBalpha at Ser-32 and Ser-36 is an essential myogenic step. Adenoviral transfer of wild-type or kinase deficient forms of IKKalpha or IKKbeta revealed that IKKalpha is required for IGF II-dependent myoblast differentiation, whereas IKKbeta is not essential for this process. Finally, overexpression of kinase-proficient wild-type NIK showed that the activation of NIK is sufficient to generate signals that trigger myogenin expression and multinucleated myotube formation in the absence of IGF-II. PMID- 11279242 TI - The human homologue of the yeast DNA repair and TFIIH regulator MMS19 is an AF-1 specific coactivator of estrogen receptor. AB - Steroid/nuclear hormone receptors are ligand-dependent transcriptional regulators that control gene expression in a wide array of biological processes. The transcriptional activity of the receptors is mediated by an N-terminal ligand independent transcriptional activation function AF-1 and a C-terminal ligand dependent transcriptional activation function AF-2. The nuclear receptor coactivator RAC3 (also known as AIB1/ACTR/pCIP/TRAM-1/SRC-3) is amplified in breast cancer cells, where it forms a complex with estrogen receptor (ER) and enhances AF-2 activity of the receptor. Here, we identify a putative human homologue of the yeast DNA repair and transcriptional regulator MMS19 as a RAC3 interacting protein. The human MMS19 interacts with the N-terminal PAS-A/B domain of RAC3 in vivo and in vitro through a conserved C-terminal domain. Interestingly, the human MMS19 also interacts with estrogen receptors in a ligand independent manner but not with retinoic acid receptor or thyroid hormone receptor. Overexpression of the interacting domain of hMMS19 strongly inhibits ER mediated transcriptional activation, indicating a dominant negative activity. In contrast, over expression of the full-length hMMS19 enhances ER-mediated transcriptional activation. We find that hMMS19 stimulates the AF-1 activity of ERalpha, but not the AF-2 activity, suggesting that hMMS19 may be an AF-1 specific transcriptional coactivator of estrogen receptor. PMID- 11279243 TI - Involvement of ADP-ribosylation factor 1 in cholera toxin-induced morphological changes of Chinese hamster ovary cells. AB - ADP-ribosylation factor 1 (ARF1) was originally found as a cofactor in CT catalyzed ADP-ribosylation of Galpha(s) but is now known to participate in vesicle trafficking. We asked whether ARF1 function in vesicular trafficking is necessary for CT-induced morphological changes in Chinese hamster ovary (CHO) cells, which result from increased intracellular cAMP. Brefeldin A treatment of cells suppressed CT action, confirming a requirement for Golgi integrity. Overexpression of a GFP-ARF1 fusion protein did not affect the morphological changes induced by CT, but changes were reduced in cells overexpressing guanine nucleotide exchange-defective ARF1(T31N) or GTP hydrolysis-deficient ARF1(Q71L) mutants. In cells expressing these mutants, 8-bromo-cAMP induced changes similar to those seen in cells transfected with ARF1 or vector. Inhibition of CT action was specific for mutants of ARF1 and not reproduced by analogous mutants of ARF5 or ARF6. ARF1(Q71L) was mostly colocalized with betaCOP, but ARF5(Q71L) less so. ARF6(Q67L) did not colocalize with betaCOP and was partially associated with the plasma membrane. These data are consistent with the conclusion that ARF1 influenced CT action in cells by its specific function in the vesicular transport pathway used by CT to travel from plasma membrane to Golgi to ER. PMID- 11279244 TI - Characterization of an evolutionarily conserved far-upstream enhancer in the human alpha 2(I) collagen (COL1A2) gene. AB - We have examined the chromatin structure around and upstream of the transcriptional start site of the human alpha2(I) collagen (COL1A2) gene. Four strong DNase I-hypersensitive sites (HS2-5) were only detected in fibroblasts, and a weaker one (HS1) was identified in type I collagen-negative cells. Another hypersensitive site potentially involved in COL1A2 silencing was found in intron 1 (HS(In)). HS1 and HS2 were mapped within conserved promoter sequences and at locations comparable to the mouse gene. HS3, HS4, and HS5 were likewise mapped approximately 20 kilobases upstream of COL1A2 at about the same position as the mouse far-upstream enhancer and within a remarkably homologous genomic segment. DNase I footprinting identified twelve areas of nuclease protection in the far upstream region (FU1-12) and within stretches nearly identical to the mouse sequence. The region containing HS3-5 was found to confer high and tissue specific expression in transgenic mice to the otherwise minimally active COL1A2 promoter. Characterization of the human element documented functional differences with the mouse counterpart. Enhancer activity substantially decreased without the segment containing FU1-7 and HS5, and inclusion of AluI repeats located 3' of HS3 augmented position-independent expression of the transgene. Hence, subtle differences may characterize the regulation of mammalian alpha2(I) collagen genes by evolutionarily conserved sequences. PMID- 11279246 TI - Virally induced lytic cell death elicits the release of immunogenic GRP94/gp96. AB - Necrotic cell death yields the release of cellular components that can function in the initiation of cellular immune responses. Given the established capacity of the endoplasmic reticulum chaperone GRP94 (gp96) to elicit CD8(+) T cell activation, we have investigated the cellular fate and antigenicity of GRP94 in differing scenarios of cell death. Virally induced cell death or mechanical cell death, elicited by freeze/thaw treatment of cell suspensions, yielded GRP94 release into the extracellular space; apoptotic cell death occurring in response to serum deprivation did not elicit GRP94 release. To assess the antigenicity of GRP94 released following virally induced cell death (lethal infection of cells with rVV ES-OVA(Met258-265), a recombinant, ovalbumin epitope-expressing vaccinia virus) or mechanical cell death (freeze/thaw of ovalbumin-expressing cells), tissue culture supernatant fractions were pulsed onto antigen-presenting cells, and antigen re-presentation was assayed as activation of an ovalbumin-specific T cell hybridoma. For both cell death scenarios, released GRP94 elicited a dose dependent, ovalbumin-specific, hybridoma activation. In contrast, calreticulin derived from rVV ES-OVA(Met258-265)-infected cell extracts did not stimulate B3Z activity. These data identify GRP94 as an antigenic component released upon pathological, but not apoptotic, cell death and provide an assay system for the identification of cellular components of related activity. PMID- 11279245 TI - A complex of the bacteriophage T7 primase-helicase and DNA polymerase directs primer utilization. AB - The lagging strand of the replication fork is initially copied as short Okazaki fragments produced by the coupled activities of two template-dependent enzymes, a primase that synthesizes RNA primers and a DNA polymerase that elongates them. Gene 4 of bacteriophage T7 encodes a bifunctional primase-helicase that assembles into a ring-shaped hexamer with both DNA unwinding and primer synthesis activities. The primase is also required for the utilization of RNA primers by T7 DNA polymerase. It is not known how many subunits of the primase-helicase hexamer participate directly in the priming of DNA synthesis. In order to determine the minimal requirements for RNA primer utilization by T7 DNA polymerase, we created an altered gene 4 protein that does not form functional hexamers and consequently lacks detectable DNA unwinding activity. Remarkably, this monomeric primase readily primes DNA synthesis by T7 DNA polymerase on single-stranded templates. The monomeric gene 4 protein forms a specific and stable complex with T7 DNA polymerase and thereby delivers the RNA primer to the polymerase for the onset of DNA synthesis. These results show that a single subunit of the primase-helicase hexamer contains all of the residues required for primer synthesis and for utilization of primers by T7 DNA polymerase. PMID- 11279247 TI - Formation pathways for lysine-arginine cross-links derived from hexoses and pentoses by Maillard processes: unraveling the structure of a pentosidine precursor. AB - Covalently cross-linked proteins are among the major modifications caused by the advanced Maillard reaction. So far, the chemical nature of these aggregates and their formation pathways are largely unknown. Synthesis and unequivocal structural characterization are reported for the lysine-arginine cross-links N(6) (2-([(4S)-4-ammonio-5-oxido-5-oxopentyl]amino)-5-[(2S,3R)-2,3,4- trihydroxybutyl] 3,5-dihydro-4H-imidazol-4-ylidene)-l-lysinate (DOGDIC 12), N(6)-(2-([(4S)-4 ammonio-5-oxido-5-oxopentyl]amino)-5-[(2S)-2,3-dihydroxypropyl]-3,5-dihydro-4H imidazol-4-ylidene)-l-lysinate (DOPDIC 13), and 6-((6S)-2-([(4S)-4-ammonio-5 oxido-5-oxopentyl] amino)-6-hydroxy-5,6,7,7a-tetrahydro-4H-imidazo[4,5-b] pyridin 4-yl)-l-norleucinate (pentosinane 10). For these compounds, as well as for glucosepane 9 and pentosidine 11, the formation pathways could be established by starting from native carbohydrates, Amadori products, and 3-deoxyosones, respectively. Pentosinane 10 was unequivocally proven to be an important precursor of pentosidine 11, which is a well established fluorescent indicator for advanced glycation processes in vivo. The Amadori products are shown to be the pivots in the formation of the various cross-links 9-13. The bicyclic structures 9-11 are directly derived from aminoketoses, whereas 12 and 13 stem from reaction with the 3-deoxyosones. All products 9-13 were identified and quantified from incubations of bovine serum albumin with the respective 3 deoxyosone or carbohydrate. From these results it seems fully justified to expect both glucosepane 9 and DOGDIC 12 to constitute important in vivo cross-links. PMID- 11279248 TI - Purine but not pyrimidine nucleotides support rotation of F(1)-ATPase. AB - The binding change model for the F(1)-ATPase predicts that its rotation is intimately correlated with the changes in the affinities of the three catalytic sites for nucleotides. If so, subtle differences in the nucleotide structure may have pronounced effects on rotation. Here we show by single-molecule imaging that purine nucleotides ATP, GTP, and ITP support rotation but pyrimidine nucleotides UTP and CTP do not, suggesting that the extra ring in purine is indispensable for proper operation of this molecular motor. Although the three purine nucleotides were bound to the enzyme at different rates, all showed similar rotational characteristics: counterclockwise rotation, 120 degrees steps each driven by hydrolysis of one nucleotide molecule, occasional back steps, rotary torque of approximately 40 piconewtons (pN).nm, and mechanical work done in a step of approximately 80 pN.nm. These latter characteristics are likely to be determined by the rotational mechanism built in the protein structure, which purine nucleotides can energize. With ATP and GTP, rotation was observed even when the free energy of hydrolysis was -80 pN.nm/molecule, indicating approximately 100% efficiency. Reconstituted F(o)F(1)-ATPase actively translocated protons by hydrolyzing ATP, GTP, and ITP, but CTP and UTP were not even hydrolyzed. Isolated F(1) very slowly hydrolyzed UTP (but not CTP), suggesting possible uncoupling from rotation. PMID- 11279249 TI - Conformation, localization, and integrin binding of talin depend on its interaction with phosphoinositides. AB - Talin is a structural component of focal adhesion sites and is thought to be engaged in multiple protein interactions at the cytoplasmic face of cell/matrix contacts. Talin is a major link between integrin and the actin cytoskeleton and was shown to play an important role in focal adhesion assembly. Consistent with the view that talin must be activated at these sites, we found that phosphatidylinositol 4-monophosphate and phosphatidylinositol 4,5-bisphosphate (PI4,5P(2)) bound to talin in cells in suspension or at early stages of adhesion, respectively. When phosphoinositides were associated with phospholipid bilayer, talin/phosphoinositide association was restricted to PI4,5P(2). This association led to a conformational change of the protein. Moreover, the interaction between integrin and talin was greatly enhanced by PI4,5P(2)-induced talin activation. Finally, sequestration of PI4,5P(2) by a specific pleckstrin homology domain confirms that PI4,5P(2) is necessary for proper membrane localization of talin and that this localization is essential for the maintenance of focal adhesions. Our results support a model in which PI4,5P(2) exposes the integrin-binding site on talin. We propose that PI4,5P(2)-dependent signaling modulates assembly of focal adhesions by regulating integrin-talin complexes. These results demonstrate that activation of the integrin-binding activity of talin requires not only integrin engagement to the extracellular matrix but also the binding of PI4,5P(2) to talin, suggesting a possible role of lipid metabolism in organizing the sequential assembly of focal adhesion components. PMID- 11279250 TI - Angiotensin II-induced stimulation of p21-activated kinase and c-Jun NH2-terminal kinase is mediated by Rac1 and Nck. AB - p21-activated kinase (PAK) has been shown to be an upstream mediator of JNK in angiotensin II (AngII) signaling. Little is known regarding other signaling molecules involved in activation of PAK and JNK by AngII. Rho family GTPases Rac and Cdc42 have been shown to enhance PAK activity by binding to p21-binding domain of PAK (PAK-PBD). In vascular smooth muscle cells (VSMC) AngII stimulated Rac1 binding to GST-PAK-PBD fusion protein. Pretreatment of VSMC by genistein inhibited AngII-induced Rac1 activation, whereas Src inhibitor PP1 had no effect. Inhibition of protein kinase C by phorbol 12,13-dibutyrate pretreatment also decreased AngII-mediated activation of Rac1. The adaptor molecule Nck has been shown previously to mediate PAK activation by facilitating translocation of PAK to the plasma membrane. In VSMC AngII stimulated translocation of Nck and PAK to the membrane fraction. Overexpression of dominant-negative Nck in Chinese hamster ovary (CHO) cells, stably expressing the AngII type I receptor (CHO-AT1), inhibited both PAK and JNK activation by AngII, whereas it did not affect ERK1/2. Finally, dominant-negative Nck inhibited AngII-induced DNA synthesis in CHO-AT1 cells. Our data provide evidence for Rac1 and Nck as upstream mediators of PAK and JNK in AngII signaling and implicate JNK in AngII-induced growth responses. PMID- 11279251 TI - ATP-dependent adenophostin activation of inositol 1,4,5-trisphosphate receptor channel gating: kinetic implications for the durations of calcium puffs in cells. AB - The inositol 1,4,5-trisphosphate (InsP(3)) receptor (InsP(3)R) is a ligand-gated intracellular Ca(2+) release channel that plays a central role in modulating cytoplasmic free Ca(2+) concentration ([Ca(2+)](i)). The fungal metabolite adenophostin A (AdA) is a potent agonist of the InsP(3)R that is structurally different from InsP(3) and elicits distinct calcium signals in cells. We have investigated the effects of AdA and its analogues on single-channel activities of the InsP(3)R in the outer membrane of isolated Xenopus laevis oocyte nuclei. InsP(3)R activated by either AdA or InsP(3) have identical channel conductance properties. Furthermore, AdA, like InsP(3), activates the channel by tuning Ca(2+) inhibition of gating. However, gating of the AdA-liganded InsP(3)R has a critical dependence on cytoplasmic ATP free acid concentration not observed for InsP(3)-liganded channels. Channel gating activated by AdA is indistinguishable from that elicited by InsP(3) in the presence of 0.5 mM ATP, although the functional affinity of the channel is 60-fold higher for AdA. However, in the absence of ATP, gating kinetics of AdA-liganded InsP(3)R were very different. Channel open time was reduced by 50%, resulting in substantially lower maximum open probability than channels activated by AdA in the presence of ATP, or by InsP(3) in the presence or absence of ATP. Also, the higher functional affinity of InsP(3)R for AdA than for InsP(3) is nearly abolished in the absence of ATP. Low affinity AdA analogues furanophostin and ribophostin activated InsP(3)R channels with gating properties similar to those of AdA. These results provide novel insights for interpretations of observed effects of AdA on calcium signaling, including the mechanisms that determine the durations of elementary Ca(2+) release events in cells. Comparisons of single-channel gating kinetics of the InsP(3)R activated by InsP(3), AdA, and its analogues also identify molecular elements in InsP(3)R ligands that contribute to binding and activation of channel gating. PMID- 11279252 TI - Voltage dependence of the apparent affinity for external Na(+) of the backward running sodium pump. AB - The steady-state voltage and [Na(+)](o) dependence of the electrogenic sodium pump was investigated in voltage-clamped internally dialyzed giant axons of the squid, Loligo pealei, under conditions that promote the backward-running mode (K(+)-free seawater; ATP- and Na(+)-free internal solution containing ADP and orthophosphate). The ratio of pump-mediated (42)K(+) efflux to reverse pump current, I(pump) (both defined by sensitivity to dihydrodigitoxigenin, H(2)DTG), scaled by Faraday's constant, was -1.5 +/- 0.4 (n = 5; expected ratio for 2 K(+)/3 Na(+) stoichiometry is -2.0). Steady-state reverse pump current-voltage (I(pump)-V) relationships were obtained either from the shifts in holding current after repeated exposures of an axon clamped at various V(m) to H(2)DTG or from the difference between membrane I-V relationships obtained by imposing V(m) staircases in the presence or absence of H(2)DTG. With the second method, we also investigated the influence of [Na(+)](o) (up to 800 mM, for which hypertonic solutions were used) on the steady-state reverse I(pump)-V relationship. The reverse I(pump)-V relationship is sigmoid, I(pump) saturating at large negative V(m), and each doubling of [Na(+)](o) causes a fixed (29 mV) rightward parallel shift along the voltage axis of this Boltzmann partition function (apparent valence z = 0.80). These characteristics mirror those of steady-state (22)Na(+) efflux during electroneutral Na(+)/Na(+) exchange, and follow without additional postulates from the same simple high field access channel model (Gadsby, D.C., R.F. Rakowski, and P. De Weer, 1993. Science. 260:100-103). This model predicts valence z = nlambda, where n (1.33 +/- 0.05) is the Hill coefficient of Na binding, and lambda (0.61 +/- 0.03) is the fraction of the membrane electric field traversed by Na ions reaching their binding site. More elaborate alternative models can accommodate all the steady-state features of the reverse pumping and electroneutral Na(+)/Na(+) exchange modes only with additional assumptions that render them less likely. PMID- 11279253 TI - Regulation of organelle acidity. AB - Intracellular organelles have characteristic pH ranges that are set and maintained by a balance between ion pumps, leaks, and internal ionic equilibria. Previously, a thermodynamic study by Rybak et al. (Rybak, S., F. Lanni, and R. Murphy. 1997. Biophys. J. 73:674-687) identified the key elements involved in pH regulation; however, recent experiments show that cellular compartments are not in thermodynamic equilibrium. We present here a nonequilibrium model of lumenal acidification based on the interplay of ion pumps and channels, the physical properties of the lumenal matrix, and the organelle geometry. The model successfully predicts experimentally measured steady-state and transient pH values and membrane potentials. We conclude that morphological differences among organelles are insufficient to explain the wide range of pHs present in the cell. Using sensitivity analysis, we quantified the influence of pH regulatory elements on the dynamics of acidification. We found that V-ATPase proton pump and proton leak densities are the two parameters that most strongly influence resting pH. Additionally, we modeled the pH response of the Golgi complex to varying external solutions, and our findings suggest that the membrane is permeable to more than one dominant counter ion. From this data, we determined a Golgi complex proton permeability of 8.1 x 10(-6) cm/s. Furthermore, we analyzed the early-to-late transition in the endosomal pathway where Na,K-ATPases have been shown to limit acidification by an entire pH unit. Our model supports the role of the Na,K ATPase in regulating endosomal pH by affecting the membrane potential. However, experimental data can only be reproduced by (1) positing the existence of a hypothetical voltage-gated chloride channel or (2) that newly formed vesicles have especially high potassium concentrations and small chloride conductance. PMID- 11279255 TI - Transduction gain in light adaptation of rod photoreceptors. PMID- 11279254 TI - Structural similarities between glutamate receptor channels and K(+) channels examined by scanning mutagenesis. AB - The pores of glutamate receptors and K(+) channels share sequence homology, suggesting a conserved secondary structure. Scanning mutagenesis with substitution of alanine and tryptophan in GluR6 channels was performed based on the structure of KcsA. Our assay used disruption of voltage-dependent polyamine block to test for changes in the packing of pore-forming regions. Alanine scanning from D567 to R603 revealed reduced rectification resulting from channel block in two regions. A periodic pattern from F575 to M589 aligned with the pore helix in KcsA, whereas a cluster of sensitive positions around Q590, a site regulated by RNA editing, mapped to the selectivity filter in KcsA. Tryptophan scanning from D567 to R603 revealed similar patterns, but with a complete disruption of spermine block for 7 out of the 37 positions and a pM dissociation constant for Q590W. Molecular modeling with KcsA coordinates showed that GluR6 pore helix mutants disrupting polyamine block pack against M1 and M2, and are not exposed in the ion channel pore. In the selectivity filter, tryptophan creates an aromatic cage consistent with the pM dissociation constant for Q590W. A scan with glutamate substitution was used to map the cytoplasmic entrance to the pore based on charge neutralization experiments, which established that E594 was uniquely required for high affinity polyamine block. In E594Q mutants, introduction of glutamate at positions S593-L600 restored polyamine block at positions corresponding to surface-exposed residues in KcsA. Our results reinforce proposals that the pore region of glutamate receptors contains a helix and pore loop analogous to that found in K(+) channels. At the cytoplasmic entrance of the channel, a negatively charged amino acid, located in an extended loop with solvent-exposed side chains, is required for high affinity polyamine block and probably attracts cations via a through space electrostatic mechanism. PMID- 11279256 TI - Neuregulin-1 proteins in rat brain and transfected cells are localized to lipid rafts. AB - Neuregulin-1 proteins and their receptors, which are members of the ErbB subfamily of receptor tyrosine kinases, play essential roles in the development of the nervous system and heart. Most neuregulin-1 isoforms are synthesized as transmembrane proproteins that are proteolytically processed to yield an N terminal fragment containing the bioactive EGF-like domain. In this study we investigated whether neuregulins are found in lipid rafts, membrane microdomains hypothesized to have important roles in signal transduction, protein trafficking, and proteolytic processing. We found that 45% of a 140-kDa neuregulin protein in rat brain synaptosomal plasma membrane fractions was insoluble in 1% Triton X 100. Flotation gradient analysis demonstrated the presence of the brain 140 kDa neuregulin protein in low-density fractions enriched in PSD-95, a known lipid raft protein. In transfected cells expressing the neuregulin I-beta 1a or the III beta 1a isoform, most of the neuregulin proprotein was insoluble in 1% Triton X 100, and neuregulin proproteins and C-terminal fragments were detected in lipid raft fractions. In contrast, the III-beta 1a N-terminal fragment was detected only in the detergent-soluble fraction. These results suggest that localization of neuregulins to lipid rafts may play a role in neuregulin signaling within the nervous system. PMID- 11279257 TI - Overactivation of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate and N methyl-D-aspartate but not kainate receptors inhibits phosphatidylcholine synthesis before excitotoxic neuronal death. AB - Glutamate receptor overactivation induces excitotoxic neuronal death, but the contribution of glutamate receptor subtypes to this excitotoxicity is unclear. We have previously shown that excitotoxicity by NMDA receptor overactivation is associated with choline release and inhibition of phosphatidylcholine synthesis. We have now investigated whether the ability of non-NMDA ionotropic glutamate receptor subtypes to induce excitotoxicity is related to the ability to inhibit phosphatidylcholine synthesis. alpha-Amino-3-hydroxy-5-methylisoxazole-4 propionate (AMPA)-induced a concentration-dependent increase in extracellular choline and inhibited phosphatidylcholine synthesis when receptor desensitization was prevented. Kainate released choline and inhibited phosphatidylcholine synthesis by an action at AMPA receptors, because these effects of kainate were blocked by the AMPA receptor antagonist LY300164. Selective activation of kainate receptors failed to release choline, even when kainate receptor desensitization was prevented. The inhibition of phosphatidylcholine synthesis evoked by activation of non-desensitizing AMPA receptors was followed by neuronal death. In contrast, specific kainate receptor activation, which did not inhibit phosphatidylcholine synthesis, did not produce neuronal death. Choline release and inhibition of phosphatidylcholine synthesis were induced by AMPA at non desensitizing AMPA receptors well before excitotoxicity. Furthermore, choline release by AMPA required the entry of Ca(2+) through the receptor channel. Our results show that AMPA, but not kainate, receptor overactivation induces excitotoxic cell death, and that this effect is directly related to the ability to inhibit phosphatidylcholine synthesis. Moreover, these results indicate that inhibition of phosphatidylcholine synthesis is an early event of the excitotoxic process, downstream of glutamate receptor-mediated Ca(2+) overload. PMID- 11279258 TI - Distribution of an NMDA receptor:GFP fusion protein in sensory neurons is altered by a C-terminal construct. AB - The NMDA receptor plays an important role in mediating sensory input to the spinal cord. Domains within the C-terminus of the NMDA receptor bind to cytoskeletal proteins and facilitate membrane targeting and synaptic clustering, and may participate in regulation of receptor function. One strategy to manipulate NMDA receptor function is to express C-terminal constructs in neurons to disrupt synaptic clustering via competition for binding motifs in cytoskeletal proteins and postsynaptic densities. Biolistic particle-mediated gene transfer was used to deliver plasmid DNA into organotypic cultures of dorsal root ganglia (DRG). Fusion proteins consisting of recombinant (r)NMDA receptor subunit 1-1 (rNR1-1) deletion constructs and enhanced green fluorescent protein (GFP) were expressed in sensory neurons and demonstrated unique distribution patterns within the cell. Expression of the full length rNR1-1:GFP construct was cytosolic and localized to membranous patches similar to endogenous NR1-1 protein expression in sensory neurons. Expression of a construct containing only the C-terminus, GFP:C0C1C2, demonstrated nuclear and membranous localization. When the GFP:C0C1C2 construct was co-expressed with rNR1-1 in sensory neurons, membranous localization of rNR1-1 was disrupted. In contrast, co-expression of a C-terminal cassette lacking the C1 exon cassette, GFP:C0C2, with rNR1-1 did not alter the membranous distribution of rNR1-1. This observation verifies the utility of a gene transfer strategy to diminish membranous NR1-1 content by expressing a construct containing the C1 exon cassette. PMID- 11279259 TI - Inhibition of vesicular glutamate storage and exocytotic release by Rose Bengal. AB - It had been thought that quantal size in synaptic transmission is invariable. Evidence has been emerging, however, that quantal size can be varied under certain conditions. We present evidence that alteration in vesicular [(3)H]L glutamate (Glu) content within the synaptosome (a pinched-off nerve ending preparation) leads to a change in the amount of exocytotically released [(3)H]Glu. We found that Rose Bengal, a polyhalogenated fluorescein derivative, is a quite potent membrane-permeant inhibitor (K(i) = 19 nM) of glutamate uptake into isolated synaptic vesicles. This vesicular Glu uptake inhibition was achieved largely without affecting H(+)-pump ATPase. We show that various degrees of reduction elicited by Rose Bengal in [(3)H]Glu in synaptic vesicles inside the synaptosome result in a corresponding decrease in the amount of [(3)H]Glu released in a depolarization- (induced by 4-aminopyridine) and Ca(2+)-dependent manner. In contrast, fluorescein, the halogen-free analog of Rose Bengal, which is devoid of inhibitory activity on vesicular [(3)H]Glu uptake, failed to change the amount of exocytotically released [(3)H]Glu. These observations suggest that glutamate synaptic transmission could be altered by pharmacological intervention of glutamate uptake into synaptic vesicles in the nerve terminal, a new mode of synaptic manipulation for glutamate transmission. PMID- 11279261 TI - Neurotrophic actions of novel compounds designed from cyclopentenone prostaglandins. AB - Previously we found that some cyclopentenone prostaglandin derivatives promoted neurite outgrowth from PC12 cells and dorsal root ganglia explants in the presence of nerve growth factor; and so we referred to them as neurite outgrowth promoting prostaglandins (NEPPs). In this study, NEPPs protected HT22 cells against oxidative glutamate toxicity. NEPP6, one of the most effective promoters of neurite outgrowth in PC12 cells, protected the cells most potently among NEPPs 1--10. Several derivatives, NEPPs 11--19, were newly synthesized based on the chemical structure of NEPP6. NEPP11 had a more potent neuroprotective effect than NEPP6. NEPP11 also prevented the death of cortical neurons induced by various stimuli and reduced ischemic brain damage in mice. Biotinylated compounds of NEPPs were synthesized to investigate their cellular accumulation. NEPP6-biotin protected the cells and emitted potent signals from the cells. In contrast, biotinylated non-neuroprotective derivatives emitted much weaker signals. These results suggest that NEPPs are novel types of neurotrophic compounds characterized by their dual biological activities of promoting neurite outgrowth and preventing neuronal death and that their accumulation in the cells is closely associated with their neuroprotective actions. PMID- 11279260 TI - Complement anaphylatoxin C5a neuroprotects through mitogen-activated protein kinase-dependent inhibition of caspase 3. AB - We previously reported that pretreatment of murine cortico-hippocampal neuronal cultures with the complement-derived anaphylatoxin C5a, protects against glutamate neurotoxicity. In this study we explored the potential mechanisms involved in C5a-mediated neuroprotection. We found that C5a neuroprotects in vitro through inhibition of apoptotic death because pretreatment with human recombinant (hr)C5a prevented nuclear DNA fragmentation coincidental to inhibition of the pro-apoptotic caspase 3 activity mediated by glutamate treatment. Also, hrC5a-mediated responses appeared to be receptor-mediated because pretreatment of cultures with the specific C5a receptor antagonist C177, prevented hrC5a-mediated neuroprotection. Based on this evidence, we further explored possible signaling pathways involved in hrC5a inhibition of caspase 3 activation and apoptotic neuronal death. We found that treatment of cultures with the mitogen-activated protein kinase (MAPK) pathway inhibitor PD98059 prevented hrC5a-mediated inhibition of caspase 3 and apoptotic neuron death. MAPK pathways, whose activation by hrC5a is inhibited by PD98059 and C177, include the extracellular signal-regulated kinase (ERK)2 and, to a lesser extent, ERK1. The study suggests that C5a may protect against glutamate-induced apoptosis in neurons through MAPK-mediated regulation of caspase cascades. PMID- 11279262 TI - Tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) by oxidant stress in cerebellar granule neurons: modulation by N-methyl-D-aspartate through calcineurin activity. AB - Insulin receptor-substrate-1 (IRS-1) is a docking protein for several tyrosine kinase receptors. Upon tyrosine phosphorylation, IRS-1 binds to signaling molecules that express Src homology 2 (SH-2) binding domains, including phosphatidylinositol 3-kinase (PI 3-kinase), phosphotyrosine phosphatase SHP-2 (Syp), Nck, Crk and Grb-2. Hydrogen peroxide (H(2)O(2)) induces tyrosine phosphorylation of key signaling mediators presumably by inhibition of tyrosine phosphatases. In many cell types, the activation of extracellular signal-related kinases (e.g. MAPK) and other protein kinases by H(2)O(2) leads to transcriptional activation. In the current study, we examined the effect of H(2)O(2) on IRS-1 tyrosine phosphorylation in primary cultured rat cerebellar granule neurons. H(2)O(2) stimulated the rapid tyrosine phosphorylation of IRS-1 and p42/p44 MAP kinase, and induced its association with PI 3-kinase. H(2)O(2) induced IRS-1 phosphorylation was rapidly reversible (5 min) whereas MAPK phosphorylation persisted for up to 1 h. NMDA reversed H(2)O(2)-mediated tyrosine phosphorylation of IRS-1 and its association with PI 3-kinase. The dephosphorylation of IRS-1 by NMDA was calcium-dependent and was inhibited by the calcineurin inhibitor cyclosporine. Calmodulin-dependent tyrosine phosphatase activity of calcineurin was observed in vitro using both immunoprecipitated and recombinant tyrosine-phosphorylated IRS-1 as substrates. These data highlight the role of multiple phosphatases in the regulation of IRS-1 tyrosine phosphorylation and identify a novel functional property of calcineurin. PMID- 11279263 TI - Induction of brain-derived neurotrophic factor by convulsant drugs in the rat brain: involvement of region-specific voltage-dependent calcium channels. AB - A high level of hippocampal brain-derived neurotrophic factor (BDNF) in normally aged as compared with young rats suggests that it is important to maintain a considerable level of hippocampal BDNF during aging in order to keep normal hippocampal functions. To elucidate possible mechanisms of endogenous BDNF increase, changes in levels of BDNF were studied in the rat brain following systemic administration of various convulsant agents; excitotoxic glutamate agonists, NMDA, kainic acid and (+/-)-alpha-amino-3-hydroxy-5-methylisoxazole-4 propionic acid (AMPA); GABA receptor antagonists, picrotoxin, pentylenetetrazole (PTZ) and lindane (gamma-hexachlorocyclohexane); and L-type voltage-dependent calcium channel agonist, BAY-K 8644. Kainic acid and AMPA, but not NMDA, caused remarkable increases in BDNF protein in the rat hippocampus and entorhinal cortex. Picrotoxin, PTZ and lindane stimulated BDNF production in the entorhinal cortex and also in the hippocampus of rats showing very severe convulsions. On the other hand, BAY-K 8644 treatment increased BDNF levels in the neocortex and entorhinal cortex. Maximal levels of BDNF protein were observed at 12--24 h, 8- 16 h and 6 h following administration of kainic acid, PTZ and BAY-K 8644, respectively. Kainic acid stimulated BDNF synthesis in presynaptic hippocampal granule neurons, but not in postsynaptic neurons with its receptors, while PTZ and BAY-K 8644 produced the same effects in postsynaptic neurons in the entorhinal cortex (in granule neurons in the hippocampus) and in the whole cortex, respectively. Nifedipine inhibited almost completely BAY-K 8644, but not PTZ, effects. omega-Conotoxin GVIA and DCG-IV partially blocked kainic acid induced enhancement of BDNF, indicating involvement of L-type and N-type voltage dependent calcium channels, respectively. In addition, BDNF levels in the hippocampus of mice deficient in D-myo-inositol-1,4,5-triphosphate receptor gene were scarcely different from those in the same region of controls, suggesting little involvement of intracellular calcium increase through this receptor. BAY-K 8644, but not kainic acid or PTZ, stimulated the phosphorylation of cyclic AMP responsive element binding protein. Our results indicate convulsant-dependent stimulation of BDNF production and involvement of region-specific voltage dependent calcium channels. PMID- 11279264 TI - Single GABAergic synaptic terminals from rat midbrain exhibit functional P2X and dinucleotide receptors, able to induce GABA secretion. AB - GABAergic terminals from rat midbrain characterized by immunolocalization of glutamic acid decarboxylase and/or the vesicular inhibitory amino acid transporter respond to ATP or P(1),P(5)-di(adenosine-5') pentaphosphate (Ap(5)A) with an increase in the intrasynaptosomal calcium concentration measured by a microfluorimetric technique in single synaptic terminals. The ATP response is mediated through the activation of P2X receptors with an abundant presence of P2X(3) subunits. Ap(5)A, however, exerts its effects by acting through a different receptor termed the dinucleotide receptor. Both receptors, once activated in the presence of extrasynaptosomal calcium, induce a concentration dependent GABA release from synaptosomal populations with EC(50) values of 16 and 20 microM for ATP and Ap(5)A, respectively. Specific inhibition of GABA release is obtained with pyridoxal phosphate-6-azophenyl-2',4'-disulphonic acid (80 microM) on the ATP effect and with P(1),P(5)-di(inosine-5') pentaphosphate (100 nM) on the dinucleotide receptor. PMID- 11279265 TI - Selective small-molecule inhibitors of glycogen synthase kinase-3 activity protect primary neurones from death. AB - The phosphatidylinositol 3-kinase (PI 3-kinase)/protein kinase B (PKB; also known as Akt) signalling pathway is recognized as playing a central role in the survival of diverse cell types. Glycogen synthase kinase-3 (GSK-3) is a ubiquitously expressed serine/threonine protein kinase that is one of several known substrates of PKB. PKB phosphorylates GSK-3 in response to insulin and growth factors, which inhibits GSK-3 activity and leads to the modulation of multiple GSK-3 regulated cellular processes. We show that the novel potent and selective small-molecule inhibitors of GSK-3; SB-415286 and SB-216763, protect both central and peripheral nervous system neurones in culture from death induced by reduced PI 3-kinase pathway activity. The inhibition of neuronal death mediated by these compounds correlated with inhibition of GSK-3 activity and modulation of GSK-3 substrates tau and beta-catenin. Thus, in addition to the previously assigned roles of GSK-3, our data provide clear pharmacological and biochemical evidence that selective inhibition of the endogenous pool of GSK-3 activity in primary neurones is sufficient to prevent death, implicating GSK-3 as a physiologically relevant principal regulatory target of the PI 3-kinase/PKB neuronal survival pathway. PMID- 11279266 TI - Overexpression of BETA2/NeuroD induces neurite outgrowth in F11 neuroblastoma cells. AB - BETA2/NeuroD, a basic helix-loop-helix (bHLH) transcription factor, has been shown to play important roles in the development of the nervous system and the maintenance and formation of pancreatic and enteroendocrine cells. The gain of function of BETA2/NeuroD in neurogenesis has been shown in Xenopus embryos. In this study, we investigated the neurogenic potential of BETA2/NeuroD using neuroblastoma cell line, F11, which could be induced to differentiate into neurons in the presence of cAMP. To induce or block the expression of BETA2/NeuroD, expression vectors for the full-length and a C-terminal deletion mutant of BETA2 were constructed and their transactivation potential was verified using reporter genes containing the insulin promoter sequences. Overexpression of BETA2 with full-length construct induced neurite outgrowth in F11 cells in the absence of cAMP. In contrast, the C-terminal deletion mutant, BETA2(1--233), which has dominant negative activity, inhibited neurite outgrowth induced by cAMP in F11 cells. These results indicate that BETA2/NeuroD plays an important role in terminal differentiation of neuroblastoma cells. They also imply that BETA2/NeuroD or related bHLH factors plays an essential role for differentiation of F11 neuroblastoma cells. PMID- 11279267 TI - Enrichment of N-methyl-D-aspartate NR1 splice variants and synaptic proteins in rat postsynaptic densities. AB - Previous studies have suggested that the localization of the NMDA receptor NR1 subunit may be determined by the splice variant form of NR1 present. Functional studies have also supported selective targeting of NR2A and NR2B to synaptic and extrasynaptic populations, respectively. We set out to determine whether rat cortical and cerebellar NR1 splice variants and NR2 subunits are differentially localized to the postsynaptic density. Using western blot techniques, we measured the percentage of NR1 containing each cassette and the enrichment of the different cassettes and other proteins in the preparations. The results indicate that: (1) no single cassette of NR1 is differentially enriched in the postsynaptic densities and (2) the NR2A and NR2B subunits are similarly enriched at the synapse. The enrichment profiles of postsynaptic density-associated proteins demonstrated similar enrichment levels for postsynaptic density (PSD) 95, the NMDA receptor subunits, chapsyn-110, and the CaMKII alpha subunit. However, synaptophysin, SAP-102, and the GABA(A) receptor beta subunit exhibited lower enrichment levels compared to PSD-95. Additionally, cerebellar but not cortical PSDs exhibited significantly lower enrichment of alpha-amino-3-hydroxy-5 methyl-4-isoxazolepropionic acid (AMPA) GluR1. Thus, although postsynaptic densities are highly enriched in synaptic proteins, there appears to be no selective incorporation of specific NR1 splice variants or NR2 subunits into this structure. PMID- 11279268 TI - Identity of nuclear high-mobility-group protein, HMG-1, and sulfoglucuronyl carbohydrate-binding protein, SBP-1, in brain. AB - High-mobility-group (HMG) proteins are a family of non-histone chromosomal proteins which bind to DNA. They have been implicated in multiple aspects of gene regulation and cellular differentiation. Sulfoglucuronyl carbohydrate binding protein, SBP-1, which is also localized in the neuronal nuclei, was shown to be required for neurite outgrowth and neuronal migration during development of the nervous system. In order to establish relationship between SBP-1 and HMG family proteins, two HMG proteins were isolated and purified from developing rat cerebellum by heparin-sepharose and sulfatide-octyl-sepharose affinity column chromatography and their biochemical and biological properties were compared with those of SBP-1. Characterization by high performance liquid chromatography--mass spectrometry (HPLC-MS), partial peptide sequencing and western blot analysis showed the isolated HMG proteins to be HMG-1 and HMG-2. Isoelectric focusing, HPLC-MS and peptide sequencing data also suggested that HMG-1 and SBP-1 were identical. Similar to SBP-1, both HMG proteins bound specifically to sulfated glycolipids, sulfoglucuronylglycolipids (SGGLs), sulfatide and seminolipid in HPTLC-immuno-overlay and solid-phase binding assays. The HMG proteins promoted neurite outgrowth in dissociated cerebellar cells, which was inhibited by SGGLs, anti-Leu7 hybridoma (HNK-1) and anti-SBP-1 peptide antibodies, similar to SBP-1. The proteins also promoted neurite outgrowth in explant cultures of cerebellum. The results showed that the cerebellar HMG-1 and -2 proteins have similar biochemical and biological properties and HMG-1 is most likely identical to SBP 1. PMID- 11279269 TI - Differential expression of 14 genes in amyotrophic lateral sclerosis spinal cord detected using gridded cDNA arrays. AB - In order to obtain insight into the aetiology and pathogenesis of amyotrophic lateral sclerosis (ALS), high-density gene discovery arrays (GDA human version 1.2) containing 18 400 non-redundant EST cDNAs pooled from different tissue libraries have been used to monitor gene expression in lumbar spinal cord from ALS cases compared with controls. Quantitative filter analysis revealed differential expression of cDNAs normalized to internal standards. These candidates have been further investigated and their expression in spinal cord characterized in a panel of ALS and control subjects. Significant differential expression was obtained for 14 genes, 13 being elevated (up to six-fold) and one decreased (by 80%) in ALS. Amongst those elevated in ALS were thioredoxin and glial fibrilary acid protein, which have already been shown to be up-regulated in ALS, thus supporting the reliability of this approach. The other differentially regulated transcripts confirmed in the expression studies represent potential candidates in ALS pathogenesis being involved in antioxidant systems, neuroinflammation, the regulation of motor neurone function, lipid metabolism, protease inhibition and protection against apoptosis. The use of the GDA system has greatly facilitated the screening and retrieval of sequence information and has generated useful information on the cascade of molecular events occurring in ALS and potentially may highlight new candidates playing a role in the aetiology and progression of this disease. PMID- 11279270 TI - Effects of R- and S-apomorphine on MPTP-induced nigro-striatal dopamine neuronal loss. AB - In order to establish whether the antioxidant and iron-chelating activities of R apomorphine (R-APO), a D(1)-D(2) receptor agonist, may contribute to its neuroprotective property, its S-isomer, which is not a dopamine agonist, was studied. The neuroprotective property of R- and S-APO has been studied in the MPTP model of Parkinson's disease (PD). Both S-APO (0.5-1 mg/kg, subcutaneous) and R-APO (10 mg/kg) pretreatment of C57-BL mice, protected against MPTP (24 mg/kg, intraperitoneally) induced dopamine (DA) depletion and reduction in tyrosine hydroxylase (TH) activity. However, only R-APO prevented nigro-striatal neuronal cell degeneration, as indicated by the immunohistochemistry of TH positive neurones in substantia nigra and by western analysis of striatal TH content. R-APO prevented the reduction of striatal-GSH and the increase in the ratio of GSSG over total glutathione, caused by MPTP treatment. In vitro both R APO and S-APO inhibited monoamine oxidase A and B activity at relatively high concentrations (100 and 300 micromol/L, respectively). The elevated activity of TH induced by the two enantiomers may contribute to the maintenance of normal DA levels, suggesting that one of the targets of these molecules may involve upregulation of TH activity. It is suggested that the antioxidant and iron chelating properties, possible monoamine oxidase inhibitory actions, together with activation of DA receptors, may participate in the mechanism of neuroprotection by APO enantiomers against MPTP. PMID- 11279271 TI - beta-Amyloid-induced neuronal apoptosis requires c-Jun N-terminal kinase activation. AB - beta-Amyloid (A beta) has been strongly implicated in the pathophysiology of Alzheimer's disease (AD), but the means by which the aggregated form of this molecule induces neuronal death have not been fully defined. Here, we examine the role of the c-Jun N-terminal kinases (JNKs) and of their substrate, c-Jun, in the death of cultured neuronal PC12 cells and sympathetic neurons evoked by exposure to aggregated A beta. The activities of JNK family members increased in neuronal PC12 cells within 2 h of A beta treatment and reached 3--4-fold elevation by 6 h. To test the role of these changes in death caused by A beta, we examined the effects of CEP-1347 (KT7515), an indolocarbazole that selectively blocks JNK activation. Inclusion of CEP-1347 (100--300 nM) in the culture medium effectively blocked the increases in cellular JNK activity caused by A beta and, at similar concentrations, protected both PC12 cells and sympathetic neurons from A beta evoked-death. Effective protection required addition of CEP-1347 within 2 h of A beta treatment, indicating that the JNK pathway acts relatively proximally and as a trigger in the death mechanism. A dominant-negative c-Jun construct also conferred protection from A beta-evoked death, supporting a model in which JNK activation contributes to death via activation of c-Jun. Finally, CEP-1347 blocked A beta-stimulated activation of caspase-2 and -3, placing these downstream of JNK activation. These observations implicate the JNK pathway as a required element in death evoked by A beta and hence identify it as a potential therapeutic target in AD. PMID- 11279272 TI - Distinct molecular mechanisms lead to diminished myelin basic protein and 2',3' cyclic nucleotide 3'-phosphodiesterase in qk(v) dysmyelination. AB - The genetic lesion of quakingviable (qk(v)) causes diminished expression of the QKI RNA-binding protein in myelin producing cells. Consequently, several structural myelin proteins are severely reduced. Among these affected proteins, the reduction of the myelin basic protein (MBP) results from post-transcriptional abnormalities of the MBP mRNA, presumably due to the lack of interactions with QKI. However, whether this is the common mechanism for reduced expression of other myelin proteins in qk(v) dysmyelination remains unclear. Here we report that distinct molecular mechanisms underlie the reduction of MBP and the 2',3' cyclic nucleotide 3'-phosphodiesterase (CNP) in qk(v) dysmyelination. MBP transcripts bind QKI strongly and are markedly reduced in the qk(v)/qk(v) oligodendrocytes in which QKI is almost completely lost. In contrast, CNP transcripts bind QKI weakly and are only slightly affected by the lack of QKI. None the less, CNP proteins are severely reduced in the qk(v)/qk(v) brain. Since CNP transcripts are predominantly associated with translating polyribosomes, diminished CNP expression in qk(v) dysmyelination is unlikely to be due to translational failures, but more likely results from accelerated protein degradation. PMID- 11279273 TI - DNase I disinhibition is predominantly associated with actin hyperpolymerization after traumatic brain injury. AB - To elucidate a role for the cytoskeletal protein actin in post-traumatic apoptotic cell death, the ability of actin-containing tissue extracts to inhibit exogenous DNase I was evaluated. In addition, cortical, hippocampal and thalamic extracts were examined for caspase-mediated actin cleavage and changes in actin polymerization state. Rats were anesthetized, subjected to lateral fluid percussion brain injury of moderate severity, and euthanized at 1 h, 6 h, 24 h, 1 week or 3 weeks post-injury (n = 3 per time-point). Tissue extracts from all brain regions of sham (uninjured) animals inhibited exogenous DNase I activity to a significant extent. However, inhibition of DNase I was significantly reduced at 1 h and 6 h in the injured hippocampus, and at 1 h, 6 h and 3 weeks in the thalamus. DNase I in cortical extracts of all injured animals was inhibited to a similar extent as that in uninjured animals. Actin fragments consistent with caspase-mediated proteolysis were observed in immunoblots of the injured hippocampus and thalamus at 1 h and 24 h, respectively, and were present up to 3 weeks post-injury. Transient actin hyperpolymerization was observed at 1 and 6 h post-injury in the thalamus and hippocampus, while actin depolymerization was observed at 1 and 3 weeks in the cortex and thalamus. Collectively our data suggest that DNase I disinhibition following brain trauma is associated predominantly with actin hyperpolymerization but also with actin depolymerization and concomitant caspase-mediated actin proteolysis. PMID- 11279274 TI - Molecular consequences of activated microglia in the brain: overactivation induces apoptosis. AB - Microglia, the resident immune cells in the brain, play a pivotal role in immune surveillance, host defense, and tissue repair in the CNS. In response to immunological challenges, microglia readily become activated as characterized by morphological changes, expression of surface antigens, and production of immune modulators that impact on neurons to induce neurodegeneration. However, little is known concerning the fate of activated microglia. In the present study, stimulation of cultured rat primary microglia with 1 ng/mL of the inflammagen lipopolysaccharide (LPS) resulted in a maximal activation as measured by the release of tumor necrosis factor alpha (TNF alpha). However, treatment with higher concentrations of LPS resulted in significantly lower quantities of detectable TNF alpha. Further analysis revealed that overactivation of microglia with higher concentrations of LPS (> 1 ng/mL) resulted in a time- and dose dependent apoptotic death of microglia as defined by DNA strand breaks, surface expression of apoptosis-specific markers (phosphatidylserine), and activation of caspase-3. In contrast, astrocytes were insensitive to LPS-induced cytotoxicity. In light of the importance of microglia and the limited replenishment mechanism, depletion of microglia from the brain may severely hamper its capacity for combating inflammatory challenges and tissue repair. Furthermore, overactivation induced apoptosis of microglia may be a fundamental self-regulatory mechanism devised to limit bystander killing of vulnerable neurons. PMID- 11279275 TI - Characterization of 'non-N-methyl-D-Aspartate' binding sites for gacyclidine enantiomers in the rat cerebellar and telencephalic structures. AB - Gacyclidine is a non-competitive NMDA receptor antagonist with potent neuroprotective properties. However, we have previously demonstrated that gacyclidine enantiomers [(-) and (+)GK11] interact with other ('non-NMDA') binding sites which may play a role in the lower self-neurotoxicity of this compound relative to the other NMDA receptor antagonists. Evidence for these binding sites has been obtained from autoradiographic and membrane binding experiments. They were found to be expressed at high levels in the molecular layer of the cerebellum, although they can also been seen in the granular layer and in telencephalic regions. The present study was designed to further characterize these gacyclidine 'non-NMDA' binding sites. The pharmacological profiles obtained on cerebellar and telencephalic membrane homogenates showed that they could not be linked directly to the main receptors or uptake complexes of the central nervous system (CNS). However, the comparison of (-) and (+)[(3)H]GK11 binding distribution in different mutant animals bearing specific cellular deficits in the cerebellum has demonstrated that the gacyclidine 'non NMDA' binding sites are associated with the dendritic trees of Purkinje cells. Interestingly, our study also shows that the radioligand binding to both cerebellar and telencephalic structures could be modulated by endogenous factors which can be removed by a stringent prewashing procedure. PMID- 11279276 TI - Probing the mechanism of rhodopsin-catalyzed transducin activation. AB - An agonist-bound G protein-coupled receptor (GPCR) induces a GDP/GTP exchange on the G protein alpha-subunit (G alpha) followed by the release of G alpha GTP and G beta gamma which, subsequently, activate their targets. The C-terminal regions of G alpha subunits constitute a major receptor recognition domain. In this study, we tested the hypothesis that the GPCR-induced conformational change is communicated from the G alpha C-terminus, via the alpha 5 helix, to the nucleotide-binding beta 6/alpha 5 loop causing GDP release. Mutants of the visual G protein, transducin, with a modified junction of the C-terminus were generated and analyzed for interaction with photoexcited rhodopsin (R*). A flexible linker composed of five glycine residues or a rigid three-turn alpha-helical segment was inserted between the 11 C-terminal residues and the alpha 5 helix of G alpha(t) like chimeric G alpha, G alpha(ti). The mutant G alpha subunits with the Gly-loop (G alpha(ti)L) and the extended alpha 5 helix (G alpha(ti)H) retained intact interactions with G beta gamma(t), and displayed modestly reduced binding to R*. G alpha(ti)H was capable of efficient activation by R*. In contrast, R* failed to activate G alpha(ti)L, suggesting that the Gly-loop absorbs a conformational change at the C-terminus and blocks G protein activation. Our results provide evidence for the role of G alpha C-terminus/alpha 5 helix/beta 6/alpha 5 loop route as a dominant channel for transmission of the GPCR-induced conformational change leading to G protein activation. PMID- 11279277 TI - Antisense suppression of GABA(A) receptor beta subunit levels in cultured cerebellar granule neurons demonstrates their importance in receptor expression. AB - GABA(A) receptors in the CNS are pentameric molecules composed of alpha, beta, gamma, delta, epsilon and theta subunits. Studies on transfected cells have shown that GABA(A) receptor beta subunit isoforms can direct alpha1 subunit localization within the cell. To examine the role of selected subunits in governing GABA(A) receptor expression in neurons, cultures of rat cerebellar granule cells were grown with antisense or sense oligodeoxynucleotides (ODNs) specific for the alpha 1, beta 2 or gamma 2 subunits. These subunits are all expressed in granule neurons where they are thought to contribute to an abundant receptor type. Following ODN treatment, subunit expression and distribution were examined by western blotting, immunocytochemistry and RT-PCR. Treatment of the cultures with the antisense, but not the corresponding sense, ODNs reduced the levels of the targeted subunit polypeptides. In addition, the beta 2 antisense ODN reduced the level of the alpha1 subunit polypeptide without altering the level of its mRNA. In contrast, treatment with the beta 2 subunit antisense ODN did not alter gamma 2 subunit polypeptide expression, distribution or mRNA level. These findings suggest that the alpha1 subunit requires a beta subunit for assembly into GABA(A) receptors in cerebellar granule neurons. PMID- 11279278 TI - A synthetic inhibitor of p53 protects neurons against death induced by ischemic and excitotoxic insults, and amyloid beta-peptide. AB - The tumor suppressor protein p53 is essential for neuronal death in several experimental settings and may participate in human neurodegenerative disorders. Based upon recent studies characterizing chemical inhibitors of p53 in preclinical studies in the cancer therapy field, we synthesized the compound pifithrin-alpha and evaluated its potential neuroprotective properties in experimental models relevant to the pathogenesis of stroke and neurodegenerative disorders. Pifithrin-alpha protected neurons against apoptosis induced by DNA damaging agents, amyloid beta-peptide and glutamate. Protection by pifithrin alpha was correlated with decreased p53 DNA-binding activity, decreased expression of the p53 target gene BAX and suppression of mitochondrial dysfunction and caspase activation. Mice given pifithrin-alpha exhibited increased resistance of cortical and striatal neurons to focal ischemic injury and of hippocampal neurons to excitotoxic damage. These preclinical studies demonstrate the efficacy of a p53 inhibitor in models of stroke and neurodegenerative disorders, and suggest that drugs that inhibit p53 may reduce the extent of brain damage in related human neurodegenerative conditions. PMID- 11279279 TI - ARPP-16/ARPP-19: a highly conserved family of cAMP-regulated phosphoproteins. AB - ARPP-16 and ARPP-19 are closely related cAMP-regulated phosphoproteins that were initially discovered in mammalian brain as in vitro substrates for protein kinase A (PKA). ARPP-16 is enriched in dopamine-responsive medium spiny neurons in the striatum, while ARPP-19 is ubiquitously expressed. ARPP-19 is highly homologous to alpha-endosulfine and database searches allowed the identification of novel related proteins in D. melanogaster, C. elegans, S. mansoni and yeast genomes. Using isoform-specific antibodies, we now show that ARPP-19 is composed of at least two differentially expressed isoforms (termed ARPP-19 and ARPP 19e/endosulfine). All ARPP-16/19 family members contain a conserved consensus site for phosphorylation by PKA (RKPSLVA in mammalian ARPP-16 and ARPP-19), and this site was shown to be efficiently phosphorylated in vitro by PKA. An antibody that specifically recognized the phosphorylated form of ARPP-16/19/19e was used to examine the phosphorylation of ARPP-16/19 family members in intact cells. In striatal slices, the phosphorylation of ARPP-16 was increased in response to activation of D(1)-type dopamine receptors, and decreased in response to activation of D(2)-type dopamine receptors. In non-neuronal cells, ARPP-19 was highly phosphorylated in response to activation of PKA. These results establish that ARPP-16/19 proteins constitute a family of PKA-dependent intracellular messengers that function in all cells. The high levels of ARPP-16 in striatal neurons and its bi-directional regulation by dopamine suggest a specific role in dopamine-dependent signal transduction. The conservation of this protein family through evolution suggests that it subserves an important cellular function that is regulated by PKA. PMID- 11279280 TI - alpha-Synuclein forms a complex with transcription factor Elk-1. AB - alpha-Synuclein has been identified as a component of Lewy bodies in Parkinson's disease and diffuse Lewy body disease, and glial cytoplasmic inclusions (GCIs) in multiple system atrophy (MSA). To explore the role of alpha-synuclein in the pathogenesis, we searched for molecules interacting with alpha-synuclein and discovered that GCIs are stained by anti-Elk-1 antibody. To seek the role of Elk 1 in synucleinopathies, we cotransfected alpha-synuclein and Elk-1 to cultured cells, and found small granular structure complexes where the two molecules colocalized. Moreover, alpha-synuclein and Elk-1 were co-immunoprecipitated from the cell lysates. For formation of the complex, the presence of both ETS and B box domains of Elk-1 was required. Although there was no evidence of direct binding between alpha-synuclein and Elk-1, we discovered that alpha-synuclein and Elk-1 both bind to ERK-2, a MAP kinase. The effect of alpha-synuclein on the MAP kinase pathway was assessed using the Pathdetect system, which showed prominent attenuation of Elk-1 phosphorylation with alpha-synuclein, and especially A53T mutant. Our results suggest that alpha-synuclein reacts with the MAP kinase pathway, which might cause dysfunction of neurons and oligodendrocytes and lead to neurodegeneration in Parkinson's disease and MSA. PMID- 11279281 TI - Neurotrophins differentially enhance acetylcholine release, acetylcholine content and choline acetyltransferase activity in basal forebrain neurons. AB - Several lines of evidence indicate that nerve growth factor is important for the development and maintenance of the basal forebrain cholinergic phenotype. In the present study, using rat primary embryonic basal forebrain cultures, we demonstrate the differential regulation of functional cholinergic markers by nerve growth factor treatment (24--96 h). Following a 96-h treatment, nerve growth factor (1--100 ng/mL) increased choline acetyltransferase activity (168- 339% of control), acetylcholine content (141--185%), as well as constitutive (148 -283%) and K(+)-stimulated (162--399%) acetylcholine release, but increased release was not accompanied by increased high-affinity choline uptake. Enhancement of ACh release was attenuated by vesamicol (1 microM), suggesting a vesicular source, and was abolished under choline-free conditions, emphasizing the importance of extracellular choline as the primary source for acetylcholine synthesized for release. A greater proportion of acetylcholine released from nerve growth factor-treated cultures than from nerve growth factor-naive cultures was blocked by voltage-gated Ca(2+) channel antagonists, suggesting that nerve growth factor modified this parameter of neurotransmitter release. Cotreatment of NGF (20 ng/mL) with K252a (200 nM) abolished increases in ChAT activity and prevented enhancement of K(+)-stimulated ACh release beyond the level associated with K252a, suggesting the involvement of TrkA receptor signaling. Also, neurotrophin-3, neurotrophin-4 and brain-derived neurotrophic factor (all at 5- 200 ng/mL) increased acetylcholine release, although they were not as potent as nerve growth factor and higher concentrations were required. High brain-derived neurotrophic factor concentrations (100 and 200 ng/mL) did, however, increase release to a level similar to nerve growth factor. In summary, long-term exposure (days) of basal forebrain cholinergic neurons to nerve growth factor, and in a less-potent fashion the other neurotrophins, enhanced the release of acetylcholine, which was dependent upon a vesicular pool and the availability of extracellular choline. PMID- 11279282 TI - Rescue from death but not from functional impairment: caspase inhibition protects dopaminergic cells against 6-hydroxydopamine-induced apoptosis but not against the loss of their terminals. AB - Despite the identification of several mutations in familial Parkinson's disease (PD), the underlying mechanisms of dopaminergic neuronal loss in idiopathic PD are still unknown. To study whether caspase-dependent apoptosis may play a role in the pathogenesis of PD, we examined 6-hydroxydopamine (6-OHDA) toxicity in dopaminergic SH-SY5Y cells and in embryonic dopaminergic mesencephalic cultures. 6-OHDA induced activation of caspases 3, 6 and 9, chromatin condensation and cell death in SH-SY5Y cells. The caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-(O methyl)fluoromethylketone (zVAD-fmk) or adenovirally mediated ectopic expression of the X-chromosomal inhibitor of apoptosis protein (XIAP) blocked caspase activation and prevented death of SH-SY5Y cells. Similarly, zVAD-fmk provided protection from 6-OHDA-induced loss of tyrosine hydroxylase-positive neurones in mesencephalic cultures. In contrast, zVAD-fmk failed to protect mesencephalic dopaminergic neurones from 6-OHDA-induced loss of neurites and reduction of [(3)H]dopamine uptake. These data suggest that, although caspase inhibition provides protection from 6-OHDA-induced death of dopaminergic neurones, the neurones may remain functionally impaired. PMID- 11279283 TI - 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) causes Akt phosphorylation and morphological changes in intracellular organellae in cultured rat astrocytes. AB - 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) is widely used for cell viability and cytotoxicity assays, but cell biological effects of MTT itself have not been investigated. In this paper we show that MTT induces a morphological change in an intracellular membranous compartment labeled with anti Rab5 antibody, dissociation of early endosomal auto-antigen (EEA1) from the membrane fraction, and phosphorylation of Akt probably through a phosphatidylinositol-3-OH kinase [PI(3)K] pathway in cultured rat astrocytes. These findings suggest that MTT affects cellular functions and conditions to some extent, and such effects of MTT may cause some discrepancies of measurement of cell viability using MTT assay and other assays. That is, the effects of MTT on cells could influence the results of cell viability assay. Moreover, MTT or other tetrazolium salts could be used as interesting activators of Akt to investigate the mechanism by which Akt or PI(3)K is activated. PMID- 11279284 TI - Isolation of 2000-kDa complexes of N-methyl-D-aspartate receptor and postsynaptic density 95 from mouse brain. AB - Neurotransmitter receptors in vivo are linked to intracellular adaptor proteins and signalling molecules driving downstream pathways. Methods for physical isolation are essential to answer fundamental questions about the size, structure and composition of in vivo complexes and complement the widely used yeast 2 hybrid method. The N-methyl-D-aspartate receptor (NMDAR) binds postsynaptic density 95 (PSD-95) protein; both are required for synaptic plasticity and learning and participate in other important pathophysiological functions. Here we describe the development and optimization of novel methods for large-scale isolation of NMDAR--PSD-95 complexes from mouse brain including immunoaffinity, immunoprecipitation, ligand-affinity and immobilized PSD-95 binding peptides. Short PDZ binding peptides modelled on NMDAR subunits were shown to isolate NMDAR complexes. Gel filtration indicated the native NMDAR--PSD-95 complexes were 2000 kDa, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) revealed a complexity suggesting a huge network of both structural components and signalling enzymes. These methods can be used to define the structure of the complexes at different synapses and in mice carrying gene mutations as well as new tools for drug discovery. PMID- 11279285 TI - Aging alters the multichemical networking profile of the human brain: an in vivo (1)H-MRS study of young versus middle-aged subjects. AB - In our most recent study of normal aging, we found decreased concentration of multiple chemicals in the brain of middle-aged subjects, as compared with younger subjects using in vivo proton magnetic resonance spectroscopy ((1)H-MRS). We hypothesized that these age-dependent differences in brain chemistry changes might be a reflection of the multichemical-networking-profile (MCNP) changes during aging. Using (1)H-MRS and correlation analysis, we examined the patterns of regional chemical levels and MCNP within and across multiple brain regions for all nine chemicals of (1)H-MR spectra. The brain chemistry changes and MCNP patterns were compared between 21 young (19--31-year-old) and 31 middle-aged (40- 52-year-old) normal volunteers. Middle-aged subjects demonstrated a significant decrease of chemical levels in the prefrontal cortex and sensorimotor cortex (SMC), as compared with the young age group. Of these, neurotransmitters GABA and glutamate in the dorsolateral prefrontal cortex (DLPFC) were altered the most. We also found a significant increase of overall chemical correlation strength in MCNP within and across all studied brain regions with increased age. These changes were caused by alterations in the pattern of negative chemical connectivity across brain regions, which become weaker (less negative) in middle aged subjects. The interregional chemical connectivity for the cingulate cortex, SMC and the thalamus was changed the most with increased age. Increased levels of chemical correlation strength across brain regions in aging were found for most chemicals studied (including neurotransmitters GABA and glutamate), and not for N acetyl aspartate. These age-related differences in the connectivity of neurotransmitters were not region dependent. The results suggest that aging is associated with changes of the regional brain chemistry and the brain MCNP. The latter process may reflect an adaptive or compensatory response (possibly related to the elongation of dendrites with aging) to reduced levels of regional brain chemicals. The (1)H-MRS approach proposed here can be used as a valuable tool in the study of the brain chemistry, MCNP and their relationships in normal and abnormal aging. PMID- 11279286 TI - Amyloid-beta peptide fragments p3 and p4 induce pro-inflammatory cytokine and chemokine production in vitro and in vivo. AB - Alzheimer's disease (AD) pathology is characterized by senile plaques containing amyloid-beta (A beta) peptide, a protein with neurotoxic and glial immune activating potential. In addition to the highly amyloidogenic peptides A beta(1- 40/42), plaques contain amino-terminal truncated A beta peptides including the alpha secretase-generated p3 fragments A beta(17--40/42). In the present study, A beta(17--40/42), A beta(1--40/42), A beta(1--16), and A beta(25--35) aged in different solvents exhibited varying capacity to activate the murine microglia cell line MG-7 depending on solvent, peptide 'aging', and peptide sequence that did not strictly correlate with beta-sheet formation. A beta(17--40/42) or A beta(1--42) stimulated production of the pro-inflammatory cytokines interleukin (IL)-1 alpha, IL-1 beta, IL-6 and tumor necrosis factor-alpha (TNF-alpha), and the chemokine MCP-1 from differentiated human monocytes (THP-1) while little or no stimulation was observed with the other A beta fragments. MG7 cells also produced these five pro-inflammatory proteins in response to A beta(1-42) whereas A beta(17--40/42) elicited mainly TNF-alpha and MCP-1. Murine and human astrocyte cell lines (D30 and U373, respectively) were generally less responsive to A beta fragments producing mainly IL-6 and MCP-1 in response to A beta(1--42) or A beta(17--40/42) fragments. In mice, an intracerebroventricular infusion of A beta(1--42) significantly increased IL-1 alpha, IL-1 beta, IL-6 and MCP-1 while A beta(17--40/42) increased MCP-1 and A beta(17--40) increased IL-1 beta. These results demonstrate that p3 and p4 A beta fragments are pro-inflammatory glial modulators and thus may play a role in development of the immunopathology observed in AD. PMID- 11279287 TI - Plasminogen expression in the neonatal and adult mouse brain. AB - Tissue-type plasminogen activator (tPA) has been implicated in a variety of types of neural plasticity, including cell migration, occlusion-induced visual system plasticity, and learning. In the periphery, plasminogen serves as tPA's primary substrate; however, studies attempting to identify plasminogen in the central nervous system have produced mixed results. We have performed a comprehensive, multitechnique study examining plasminogen expression in the neonatal and adult mouse brain. Reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization reveal plasminogen mRNA in the cortex, hippocampus and cerebellum of both neonatal and adult C57BL/6 mice. Immunocytochemistry reveals plasminogen protein expression in these same brain regions. Notably, plasminogen expression in the cerebellum occurs in the granule cell and the Purkinje cell layers. tPA activity in these same regions is involved in granule cell migration during development and motor learning in adulthood. Therefore, these findings demonstrate that plasminogen is present in the central nervous system and localized to areas where it could serve as a substrate for plasticity-related increases in tPA activity. PMID- 11279288 TI - Examination of the involvement of protein kinase A in D2 dopamine receptor antagonist-induced immediate early gene expression. AB - Immediate early genes (IEGs) are induced by different signaling pathways. It has been proposed that D2 dopamine receptor blockade induces IEG expression through activation of protein kinase A (PKA), although few studies have examined this issue in vivo. We infused the PKA inhibitor H-89 into the striatum of male rats, followed 30 min later by systemic administration of eticlopride. Eticlopride induced c-fos and zif268 mRNA expression in striatum was not blocked by H-89. In addition, eticlopride did not produce measurable levels of PKA activity in striatum, whereas the cAMP activator Sp-8-Br-cAMPs increased levels of activated PKA. Neither the adenosine A2a receptor agonist CGS 21680 nor the phosphodiesterase-4 inhibitor rolipram, each of which should increase PKA activation, potentiated eticlopride-induced IEG expression. To test whether other signaling pathways are involved in eticlopride-mediated gene induction, we also infused inhibitors of the mitogen-activated and calcium/calmodulin-dependent protein kinases into animals and then treated them with eticlopride. The data suggest that eticlopride-induced IEG expression is not solely dependent on these kinases either. These data suggest that PKA activation may not be necessary for induction of IEGs by D2 dopamine receptor antagonists and that other intracellular signaling pathways may be involved. PMID- 11279289 TI - Neuronal nicotinic acetylcholine receptor alpha3 subunit protein in rat brain and sympathetic ganglion measured using a subunit-specific antibody: regional and ontogenic expression. AB - A synthetic peptide corresponding to the C-terminus of the alpha 3 subunit of the rat neuronal nicotinic acetylcholine receptor (nAChR) was used to generate a rabbit polyclonal alpha 3 antibody. The specificity of this antibody was characterized by immunoblotting, immunohistochemical and immunoprecipitation techniques. Using this antibody, the relative densities of the alpha 3 subunit were quantitatively determined in different brain regions and in superior cervical ganglion (SCG). Among these regions, SCG, interpeduncular nucleus (IPN) and pineal gland showed the highest levels of alpha 3 protein expression. Habenula and superior colliculi had intermediate levels of expression. Low levels were found in cerebral cortex, hippocampus and cerebellum. The ontogenic profile of the alpha 3 subunit in the SCG was also determined. The alpha 3 protein level is low at postnatal day (P 1), but increases rapidly during the first seven postnatal days. This level then plateaus and remains stable through postnatal day 35. These findings suggest that neuronal nAChRs containing the alpha 3 subunit participate in important roles in specific regions of the rat brain and the SCG. PMID- 11279290 TI - Direct determination of the N-acetyl-L-aspartate synthesis rate in the human brain by (13)C MRS and [1-(13)C]glucose infusion. AB - A non-invasive (13)C magnetic resonance spectroscopy (MRS) technique is described for the determination of the N-acetyl-L-aspartate (NAA) synthesis rate, V(NAA), in the human brain in vivo. In controls, the mean V(NAA) was 9.2 +/- 3.9 nmol/min/g. In Canavan disease, where [NAA] is increased (p < 0.001) and [aspartate] is deceased (p < 0.001), V(NAA) was significantly reduced to 3.6 +/- 0.1 nmol/min/g (p < 0.001). These rates are in close agreement with the activity of the biosynthetic enzyme measured in vitro in animals, and with the rate of urinary excretion of NAA in human subjects with Canavan disease. The present result is consistent with the regulation of NAA synthesis by the activity of a single enzyme, L-aspartate-N-acetyltransferase, in vivo, and with its control in Canavan disease by limited substrate supply and/or product inhibition. The (13)C MRS technique provides the means for further determination of abnormal rates of neuronal NAA synthesis among neurological disorders in which low cerebral [NAA] has been identified. PMID- 11279292 TI - Do fertilin beta and cyritestin play a major role in mammalian sperm--oolemma interactions? A critical re-evaluation of the use of peptide mimics in identifying specific oocyte recognition protiens. AB - Integrins have been proposed to play a role in mammalian sperm-oocyte interactions for many years. To a large extent this hypothesis stems from the ability of short synthetic peptides, based on the disintegrin-like domains of two sperm surface integral membrane proteins, fertilin beta and cyritestin, to inhibit sperm--oocyte binding and fusion in vitro. Here we argue that such peptide mimics lack specificity in these simple IVF assay systems. Hence, whilst not precluding a role for fertilin beta and cyritestin in sperm-oolemma interactions, this lack of specificity indicates the need for considerable caution when interpreting results obtained using this approach. PMID- 11279293 TI - Gonadotrophins inhibit and activin induces expression of inhibin/activin beta(B) subunit mRNA in cultured human granulosa-luteal cells. AB - During the human menstrual cycle, serum inhibin concentrations fluctuate in a cyclic fashion. To examine the regulation of inhibin/activin beta(B) subunit gene expression in ovarian granulosa-luteal cells, the levels of beta(B) subunit mRNA were determined in primary cultures of human granulosa-luteal cells treated with gonadotrophins and protein kinase modulators. Granulosa cells were obtained from women undergoing an IVF programme. The cells were enzymatically dispersed, separated from red blood cells, and maintained in culture for 5--10 days before addition of different agents. Northern blot analysis with specific oligonucleotide probes was performed to study inhibin/activin beta(B) subunit mRNA levels. Both LH and FSH reduced the accumulation of beta(B) subunit mRNA in a dose-dependent manner. The protein kinase A activator, (Bu)(2)cAMP, and the protein kinase inhibitor staurosporine also inhibited beta(B) subunit mRNA expression dose-dependently. Activin A increased dose-dependently beta(B) subunit mRNA expression. Our study suggests that activin-induced and gonadotrophin inhibited beta(B) subunit expression in granulosa cells might be key factors in the transition from inhibin B to inhibin A dominance during the menstrual cycle. PMID- 11279294 TI - The balance between MMP-9 and MMP-2 and their tissue inhibitor (TIMP)-1 in luteinized granulosa cells: comparison between women with PCOS and normal ovulatory women. AB - Polycystic ovarian syndrome (PCOS) involves follicular atresia, formation of multiple ovarian cysts and is frequently associated with a higher abortion rate. Follicular development, ovulation, formation of corpus luteum and its regression involve extensive tissue remodelling. Mammalian ovaries express a number of matrix metalloproteinases (MMP) and their tissue inhibitors (TIMP). We assessed the differences in production of MMP-2, MMP-9 and TIMP-1 by cultured luteinized granulosa cells from women with PCOS and normal ovulatory women after ovarian stimulation for IVF treatment. In follicular fluid from women with PCOS, levels of MMP-9 and MMP-2 were higher than the normal group, as was the basal production of these proteins by cultured cells. Basal production of TIMP-1 by cultured cells was not different between PCOS and normal groups. A time-dependent increase in the production of MMP-9 was observed in cells from both normal and PCOS women, although the increase was more pronounced in the latter. Thus the MMP-TIMP balance is shifted toward greater MMP activity in luteinized granulosa cells from women with PCOS. PMID- 11279295 TI - Interleukin-1 receptor accessory protein is constitutively expressed in human endometrium throughout the menstrual cycle. AB - Interleukin-1 (IL-1) is one of the principal cytokines that participate in endocrine and local regulation of many endometrial and reproductive functions. The cellular response to IL-1 principally implicates receptor type 1 (IL-1R tI) and, according to recent data, an accessory protein (IL-1R-AcP) that seems to play an essential function in signal transduction. In the present study, we examined the expression of IL-1R-AcP in the endometrium of 39 normal fertile women throughout the menstrual cycle. As studied by immunohistochemistry, IL-1R AcP was detected across endometrial tissue, but more noticeably in the glands and luminal epithelium. The intensity of IL-1R-AcP immunostaining was consistently high throughout the menstrual cycle, and this was confirmed by Western blot analysis of the protein and corroborated by reverse transcription-polymerase chain reaction analysis of the mRNA. To our knowledge, this is the first report showing that IL-1R-AcP is expressed in endometrial tissue, and without any noticeable variation throughout the menstrual cycle. This suggests that the accessory protein, whose co-expression is critical for IL-1R tI-mediated cell activation, is, in contrast to the functional receptor, constitutively expressed and not subject to similar cycle-dependent regulation. PMID- 11279296 TI - The inhibitory effect of dienogest, a synthetic steroid, on the growth of human endometrial stromal cells in vitro. AB - Dienogest is a synthetic steroid that has been used as a progestogen in contraceptive pills and is currently being studied for its possible clinical use in the treatment of endometriosis. In this study, we investigated the direct effects of dienogest in differentiation and proliferation of human endometrial stromal cells (ESC) in vitro. After 12 days in the presence of oestradiol (10(-8) mol/l) plus dienogest (10(-6) mol/l), cultured ESC underwent morphological differentiation and produced prolactin, a typical marker for decidualization. By using Northern blot analysis and radioimmunoassay, it was shown that treatment of ESC with oestradiol (10(-8) mol/l) plus dienogest (10(-9) to10(-6) mol/l) led to an increase in the levels of prolactin mRNA and prolactin production in a dose dependent manner. Additionally, RU-486, a progesterone receptor antagonist, almost completely inhibited dienogest-induced prolactin production. As shown by the thymidine uptake method, there was a dose-dependent inhibition of ESC proliferation with dienogest (P < 0.01, control versus concentrations >10(-7) mol/l). The significant inhibition of ESC proliferation by dienogest (10(-7) mol/l) was partially reversed by RU-486 (10(-6) mol/l). In summary, dienogest directly acts on endometrial tissue in progestogenic response, such as decidualization, increased prolactin production and growth retardation. These data imply that dienogest exerts direct effect in suppressing growth of endometriotic implants. PMID- 11279297 TI - Cellular localization of human relaxin-like factor in the cyclic endometrium and placenta. AB - We have studied the cellular localization of the relaxin-like factor (RLF) in the histologically normal cyclic endometrium collected from days 3--26 of the menstrual cycle. RLF transcripts and protein were detected in the luminal and glandular epithelium and in stromal cells at all stages of the cyclic endometrium. Increased expression of RLF was observed in endometrial tissues in the proliferative as compared to the secretory phase, suggesting that oestrogens affect RLF gene activity in the human endometrium. The cellular localization of RLF transcripts and protein was also determined in first trimester placental tissues obtained from normal and ectopic tubal implantation sites and in third trimester placentae of normal and pre-eclamptic pregnancies. In first trimester placenta, weaker expression of RLF was observed in the syncytiotrophoblast as compared to the underlying cytotrophoblast. Extravillous trophoblast cells constitutively expressed RLF. Trophoblast cells were the main source of RLF in the human placenta and trophoblastic RLF gene activity was unaffected by either the site of implantation or the invasive properties of the cytotrophoblast as demonstrated by samples from patients with tubal implantation and pre-eclampsia respectively. Decidual cells weakly expressed RLF. The presence of unprocessed and cleaved immunoreactive RLF in term placenta was determined by Western analysis. The above results suggest a functional role for both RLF isoforms within normal placental tissue. PMID- 11279298 TI - Expression of 11 beta-hydroxysteroid dehydrogenase isozymes and corticosteroid hormone receptors in primary cultures of human trophoblast and placental bed biopsies. AB - Interconversion of active and inactive glucocorticoids, e.g. cortisol (F) and cortisone (E) is catalysed by 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) which exists as two isoforms. We have used human placental bed biopsies and an in vitro cytotrophoblast cell culture system to examine the expression and activity of the 11 beta-HSD isoforms along with that of the glucocorticoid and mineralocorticoid receptors (GR and MR). Immunohistochemistry localized 11 beta HSD1 to decidualized stromal cells and 11 beta-HSD2 to villous cytotrophoblast, syncytiotrophoblasts and trophoblast cells invading the placental bed and maternal vasculature. In primary cultures of human cytotrophoblast, 11 beta-HSD2, GR and MR mRNA were expressed. Low levels of 11 beta-HSD1 mRNA were noted in these cultured cells, but could be explained on the basis of contaminating, vimentin-positive decidual stromal cells (< or =5%). Enzyme activity studies confirmed the presence of a high-affinity, NAD-dependent dehydrogenase activity (K(m) 137 nmol/l and V(max) 128 pmol E/h/mg protein), indicative of the 11 beta HSD2 isoform. No reductase activity was observed. The presence of functional MR and GR was determined using Scatchard analyses of dexamethasone and aldosterone binding (MR K(d) 1.4 nmol/l B(max) 3.0; GR K(d) 6.6 nmol/l B(max) 16.2 fmol/ng protein). The expression of 11 beta-HSD1 in maternal decidua and 11 beta-HSD2 in adjacent trophoblast suggests an important role for glucocorticoids in determining trophoblast invasion. The presence of the MR within trophoblast indicates that some of the effects of cortisol could be MR- rather than GR mediated. PMID- 11279299 TI - Immunohistochemical localization of insulin-like growth factors I and II at the primary implantation site in the Rhesus monkey. AB - There are various cellular mediators which can affect the process of blastocyst implantation by regulating the proliferation and differentiation of conceptus and maternal endometrial cells. Insulin-like growth factors I (IGF-I) and II (IGF-II) are potent mitogenic and differentiation-promoting growth factors. However, the role of IGF peptides at implantation in primate species is not well understood. The objective of the present study was to immunohistochemically localize IGF-I and IGF-II peptides in trophoblast cells and maternal endometrial cells during lacunar and villous stages of placentation in the Rhesus monkey. Female animals (n = 10) were laparotomized on estimated days 13-16 after fertilization to collect primary implantation sites which were subjected to immunohistochemical staining for IGF-I and IGF-II peptides. Cell-type specificity for IGF-I and IGF II was evident with a very low level of IGF-I peptide immunolocalized in trophoblast cells lining lacunae, and primary and secondary villi, while moderate to high amounts of IGF-II peptide were detected in lamellar syncytiotrophoblast cells lining lacunae, early villi and cell columns, as well as in migrating trophoblast cells in the extravillous compartment and in endovascular trophoblast cells. The observed presence of IGF-II peptide in differentiated lamellar syncytiotrophoblast cells during the very early stages of implantation and placentation in the Rhesus monkey may be important in their transition to this differentiated cell population. Maternal endometrial cells showed similar distribution profiles for IGF-I and IGF-II. In conclusion, we report differential distribution of IGF-I and IGF-II peptides in trophoblast cell populations at the feto-maternal interface during lacunar and villous stages of gestation in the Rhesus monkey. PMID- 11279300 TI - The HLA-G genotype is potentially associated with idiopathic recurrent spontaneous abortion. AB - The causes for recurrent spontaneous abortion (RSA) remain unknown in a large proportion of the cases. Human leukocyte antigen (HLA)-G and HLA-E are expressed on invasive trophoblast cells, and are supposed to confer to materno-fetal tolerance. A total of 14 different nucleotide sequences have been described for HLA-G, including one dysfunctional null allele (HLA-G*0105N), while five different sequences have been described for HLA-E. In this study, 78 RSA couples and 52 fertile controls were typed for HLA-G and HLA-E by direct sequencing or single strand conformational polymorphism (SSCP) respectively. The overall analysis showed no significant difference in allele frequencies for either HLA-G or HLA-E between the two groups. However, HLA-G allele frequencies in women who had suffered from five or more RSA differed significantly from fertile controls (P: = 0.001), and from women who had undergone three or four RSA (P: = 0.027). Detailed analysis demonstrated a significant increase in the proportion of the HLA-G alleles *01013 and *0105N in the whole group of RSA women compared with fertile controls (P: = 0.007). When studying the prognostic value of HLA genotyping for pregnancy outcome (n = 41), 31 patients (76%) gave birth to a living child without performing immunotherapy. Seven out of 10 (70%) couples suffering from a further RSA carried the HLA-G*01013 or *0105N allele, compared with 10 out 31 (32%) couples giving birth (P: = 0.06). This study suggests that the HLA-G genotype may be a contributing factor in RSA. PMID- 11279301 TI - Expression of galanin in human placenta. AB - The neuropeptide galanin was originally implicated in the regulation of feeding behaviour. Today, galanin is implicated in several physiological functions including reproduction and feeding. Many hypothalamic neurohormones of the hypothalamo--pituitary axis (HPA) are also expressed in the placenta where the specialized topological compartments of the HPA are missing and where paracrine and autocrine regulatory mechanisms consequently prevail. Since galanin influences gonadotrophin-releasing hormone secretion in the HPA, we argued that a similar regulatory role for galanin might exist in human placenta. Since the presence of galanin in human placenta had not been previously reported, we analysed galanin expression in the human placenta by immunohistochemistry and quantitative polymerase chain reaction (PCR) throughout gestation. We found that the peptide hormone localizes to the syncytio- and cytotrophoblast layers; its RNA could be detected. By quantitative PCR we observed that throughout gestation, there is a loss of galanin mRNA which parallels the fall in signal intensity from immunohistochemical detection of the galanin oligopeptide. Furthermore, we detected secretion of galanin from isolated trophoblastic cells. We conclude that galanin may be an important and novel regulator of placental function. PMID- 11279302 TI - EP(4) receptors mediate prostaglandin E(2)-stimulated glycosaminoglycan synthesis in human cervical fibroblasts in culture. AB - The aim of this study was to determine the prostaglandin E (EP) receptors and second messengers implicated in glycosaminoglycan (GAG) synthesis by human cervical fibroblasts in culture. Human cervical fibroblasts were obtained from cervical biopsies in pre-menopausal, cycling women. Cultured cells were incubated with prostaglandin E(2) (PGE(2)) and an array of agonists and antagonists. Glycosaminoglycan synthesis was assayed after extraction by measuring the [(3)H]glucosamine and [(35)S]sulphate incorporated into GAG and cAMP production was determined by radioimmunoassay. PGE(2) significantly stimulated GAG synthesis. Neither 17-phenyl-trinor-PGE(2), the EP(1) selective agonist, nor sulprostone, an EP(3) agonist, had any effect on GAG production. Butaprost, the EP(2) selective agonist, also failed to increase GAG synthesis. AH6809, an EP(2) antagonist, had no effect on PGE(2)-stimulated GAG production. AH23848, an EP(4) antagonist, inhibited the GAG synthesis provoked by PGE(2). PGE(2) and butaprost significantly increased cAMP production. Both AH6809 and AH23848 inhibited the PGE(2)-stimulated cAMP production. H89, a cAMP-dependent protein kinase (PKA) inhibitor, did not inhibit PGE(2)-stimulated GAG synthesis and Sp-cAMPS, a selective PKA activator, failed to increase GAG production. In conclusion, both EP(4) and EP(2) receptors are present and functional in human cervical fibroblasts. Only EP(4) receptors mediate PGE(2) stimulated GAG synthesis in a PKA-independent pathway. PMID- 11279303 TI - Cellular distribution of platelet-derived growth factor, transforming growth factor-beta, basic fibroblast growth factor, and their receptors in normal bone marrow. AB - We performed an immunohistochemical study on 5 normal marrow samples and 3 fibrotic marrow samples to investigate the cellular distribution of various isoforms of platelet-derived growth factor (PDGF), transforming growth factor beta (TGF-beta), basic fibroblast growth factor (bFGF), and their corresponding receptors. Immature hematopoietic precursors strongly expressed PDGF-B, all the TGF-beta isoforms, PDGFRbeta, TGFbetaRI and II, and FGFR1, 3, and 4. Megakaryocytes stained primarily for PDGF-B, TGF-beta2-3, and PDGFRbeta. Histiocytes displayed intense TGF-beta1, bFGF, and FGFR2 expression. Fibroblasts and endothelial cells carried receptors for all the aforementioned cytokines. The last 2 cell types also expressed the ligand cytokines to varied degrees. PMID- 11279304 TI - Brain natriuretic peptide is a predictor of anthracycline-induced cardiotoxicity. AB - Anthracyclines are effective antineoplastic drugs, but they frequently cause dose related cardiotoxicity. The cardiotoxicity of conventional anthracycline therapy highlights a need to search for methods that are highly sensitive and capable of predicting cardiac dysfunction. We measured the plasma level of brain natriuretic peptide (BNP) to determine whether BNP might serve as a simple diagnostic indicator of anthracycline-induced cardiotoxicity in patients with acute leukemia treated with a daunorubicin (DNR)-containing regimen. Thirteen patients with acute leukemia were treated with a DNR-containing regimen. Cardiac functions were evaluated with radionuclide angiography before chemotherapies. The plasma levels of atrial natriuretic peptide (ANP) and BNP were measured at the time of radionuclide angiography. Three patients developed congestive heart failure after the completion of chemotherapy. Five patients were diagnosed as having subclinical heart failure after the completion of chemotherapy. The plasma levels of BNP in all the patients with clinical and subclinical heart failure increased above the normal limit (40 pg/ml) before the detection of clinical or subclinical heart failure by radionuclide angiography. On the other hand, BNP did not increase in the patients without heart failure given DNR, even at more than 700 mg/m(2). The plasma level of ANP did not always increase in all the patients with clinical and subclinical heart failure. These preliminary results suggest that BNP may be useful as an early and sensitive indicator of anthracycline-induced cardiotoxicity. PMID- 11279305 TI - Alpha-2-macroglobulin and interleukin-6 levels in steady-state sickle cell disease patients. AB - Endothelial activation and subclinical microvascular occlusions are an ongoing process during steady-state sickle cell disease, leading to interleukin production and an acute-phase response. Alpha-2-macroglobulin (alpha2M) is an acute-phase protein mainly regulated by interleukin-6 (IL-6). On the other hand, alpha2M acts as a carrier protein for IL-6 during inflammatory stress. The purpose of this study is to further assess the interactions between IL-6 and alpha2M as potent modulators of inflammatory reactions during the steady state of sickle cell disease. We measured alpha2M and IL-6 levels in 21 patients (12 male, 9 female; age range 12-44 years) in the steady state of sickle cell disease. Four patients had homozygous sickle cell anaemia and 17 had double heterozygous sickle cell/beta-thalassaemia. Diagnostic quantification of alpha2M was performed by rate nephelometry. Commercial enzyme immunoassay test kits were used for the quantitative measurement of IL-6. The alpha2M and IL-6 levels were compared to the values obtained from healthy volunteers. Mean values (+/- SD) of alpha2M and IL-6 were found to be significantly increased (p < 0.0005) in the patients (alpha2M: 337.2 +/- 104 mg/dl; IL-6: 4 +/- 2.1 pg/ml) compared to the healthy controls (alpha2M: 204.2 +/- 45.8 mg/dl; IL-6: 1.15 +/- 2.5 pg/ml). IL-6 values were positively correlated with alpha2M levels (r = 0.61, p < 0.01). We observed increased alpha2M and IL-6 levels in steady-state sickle cell disease and a positive correlation between these two inflammatory mediators. We suggest that alpha2M is a potent modulator of the inflammatory reaction and tissue repair mechanism during steady-state microvascular occlusions. Elucidating the role of alpha2M in sickle cell disease could lead to the development of novel strategies and therapies for preventing the harmful systemic or local effects of excess cytokine production. PMID- 11279306 TI - Increased prevalence of GSTM(1) null genotype in patients with myelodysplastic syndrome: a case-control study. AB - BACKGROUND AND OBJECTIVE: Myelodysplastic syndromes (MDS) are clonal disorders of bone marrow stem cells characterized by ineffective hematopoiesis leading to blood cytopenia; they often progress to acute myeloid leukemia (AML). The glutathione S-transferases (GST) detoxify various agents, including those implicated in MDS. Both GSTM(1) and GSTT(1) genes have "null" alleles and are polymorphic. We studied the impact of GTM(1) and GSTT(1) null genotypes on the MDS susceptibility, disease severity and laboratory indices with prognostic value for the syndrome. MATERIAL AND METHODS: In a hospital-based case-control study we analyzed lymphocyte DNA samples from 54 patients with MDS and 60 cancer-free controls matched for age, sex, smoking habits and origin. A multiplex polymerase chain reaction was used to genotype both GSTM(1) and GSTT(1) simultaneously. The chi(2) test was used for statistical evaluation of the data and the odds ratios and attributable risk and population attributable risk were also calculated. RESULTS: A significantly increased frequency of GSTM(1) null genotype was found among MDS patients (57.4%) compared to controls (33.3%) (p < 0.01), while the frequency of GSTT(1) null genotype was not significantly higher in MDS patients (11.1% vs. 6.66%). Neither GSTM(1) and GSTT(1) null genotype was associated with a particular category of the French-American-British (FAB) classification in the patients studied. Additionally, GSTM(1) null genotype was associated with a significant decrease in the absolute number of neutrophils among the MDS patients. CONCLUSIONS: Individuals with GSTM(1) null genotype may have increased susceptibility to MDS. Null genotypes do not seem to have be associated with FAB classification while they may be associated with putative prognostic factors. PMID- 11279307 TI - Expression of functional markers in acute nonlymphoblastic leukemia. AB - Multidrug resistance parameters, tissue infiltration parameters, receptors for colony-stimulating factors (CSFr) and cell cycle parameters were analyzed using flow cytometry in 145, 109 initial and 36 relapsed or refractory, acute nonlymphoblastic leukemia (ANLL) patients to find out clinically more reliable functional parameters. Lung resistance-associated protein (LRP) was most frequently expressed in ANLL (44.1%) followed by P-glycoprotein (PGP) (35.9%) and multidrug resistance-associated protein (MRP) (8.3%). LRP and PGP were expressed more frequently in relapsed or refractory ANLL than initial ANLL cases. Complete remission rate after standard chemotherapy falls in PGP-positive cases (p = 0.001). CD44-positive ANLL cases relapsed more frequently. The organ tropism is different depending on the infiltration parameters, vascular cell adhesion molecule to splenomegaly, matrix metalloprotease-2 to hepatomegaly and to extramedullary infiltration other than spleen, liver or lymph node. The percentage of the granulocyte-macrophage-CSFr expression was high in M4 and M5, and granulocyte-CSFr-positive ANLL showed less extramedullary infiltration (p = 0.007) and more PGP expression. Ki-67 was expressed significantly less in refractory ANLL than initial ANLL and DNA topisomerase IIalpha was expressed significantly more in the surviving patients group. In conclusion, analysis of these new functional parameters could help to predict and overcome the clinical behavior of each ANLL at the time of diagnosis. PMID- 11279308 TI - Mutational analysis of beta-thalassemia cases from the Aegean region of Turkey using an allele-specific oligonucleotide hybridization technique. AB - The aim of the present study was to evaluate the point mutations of beta thalassemia patients from the Aegean region of Turkey by using an allele-specific oligonucleotide hybridization technique. DNA isolated from peripheral blood samples of 75 children with beta-thalassemia major or intermedia was analyzed using a Bio-Rad mD(x)(TM)-Be Tha Gene 1 kit. We determined mutations in 56 (74.6%) patients. The allelic frequency of mutations in 150 chromosomes was as follows: IVS-I-110 (G-A) 44.1%, IVS-I-1 (G-A) 28.2%, IVS-I-6 (T-C) 13.3%, IVS-II 745 (C-G) 9.3%, IVS-II-1 (G-A) 2.7%, Cd 39 (C-T) 2.4%, -87 (C-G) 0% and Cd 6 (-A) 0%. The distribution of the mutation types was consistent with the findings of other research groups. PMID- 11279309 TI - Additional stem cell therapy for graft failure after allogeneic bone marrow transplantation. AB - In this study we retrospectively evaluated the effect and outcome of a boost dose of donor stem cells without additional chemotherapy or total body irradiation. Between March 1983 and August 1999, 20 of 788 (2.5%) patients receiving allogeneic bone marrow transplantation (BMT) were treated with an additional boost dose of donor cells. The reasons for the use of the boost treatment were primary graft failure (early rejection; n = 7), secondary graft failure including late rejection (n = 10), refractory pure red cell aplasia caused by the remaining recipient cells producing anti-erythrocyte antibodies (n = 2), and donor lymphocyte infusion induced pancytopenia (n = 1). The patients were aged from 17 to 48 years (median age 31 years). The underlying diseases of the patients were severe aplastic anemia in 12 patients, acute myelogenous leukemia in 3, acute lymphocytic leukemia in 3, and chronic myelogenous leukemia in 2. The donors were human leukocyte antigen-identical siblings in 18 cases, 1 mismatched related donor, and 1 unrelated donor. The cell source was bone marrow in 6 cases and peripheral blood progenitor cells in 14. The median interval between BMT and the boost treatment was 7 weeks (range 1-124). No conditioning regimen was given prior to the boost treatment for 11 patients, while 4 received total nodal irradiation (TNI) plus antithymocyte globulin (ATG), 3 ATG alone, and 2 TNI plus steroid. The median infused booster mononuclear cell dose was 2.55 x 10(8)/kg (range 0.28-37.0). Fifteen (75%) patients achieved a hematological recovery. After the boost treatment, 6 of 20 (30%) patients developed acute graft-versus host disease (GVHD) > or = grade II, 3 of whom had had prior GVHD. Five (31.3%) of the evaluable 16 patients developed chronic GVHD. The GVHDs were easily controlled using immunosuppressive agents except in the case of 1 patient. Five patients died after the boost treatment; 2 within 30 days, 2 within 60 days, and 1 after 32 months. The causes of death were: 3 engraftment failures, 1 late rejection, and 1 infection following GVHD. With a median follow-up of 31.5 months (range 6-92), the Kaplan-Meier method estimated that the overall survival rate 1 and 3 years after the boost treatment was 80 and 71%, respectively. The survival of patients with primary graft failure was determined to be significantly lower compared to that of patients with secondary graft failure, using the log rank test (p = 0.0176). Disease category, stem cell source, conditioning prior to a boost treatment, and year of boost treatment did not have an influence on survival. We conclude that the reinfusion of donor stem cells is frequently successful in achieving engraftment with rare occurrence of fatal GVHD. Furthermore, relatively good long-term survival was demonstrated. PMID- 11279310 TI - Increased expression of Fas (APO-1, CD95) on CD4+ and CD8+ T lymphocytes during total body irradiation. AB - Fas/APO-1 (CD95) is a cell surface molecule that can transduce apoptotic signals into cells. We examined the expression of Fas antigen on CD4+ and CD8+ T cells of patients who received total body irradiation (TBI) as a preparative regimen for allogeneic bone marrow transplantation. Numbers of peripheral blood lymphocytes were significantly reduced after TBI. Cytofluorometric analysis revealed a significantly higher expression of Fas on CD4+ and CD8+ T cells after TBI. Serum soluble Fas concentrations were significantly elevated after TBI. Changes in the Fas system were therefore accompanied by TBI-induced lymphocytopenia, suggesting that Fas plays a role in irradiation-induced apoptosis in vivo. PMID- 11279311 TI - Dup(12)(q13-q22) and 13q14 deletion in a case of B-cell chronic lymphocytic leukemia. AB - Cases with partial trisomy 12 have rarely been found in B-cell chronic lymphocytic leukemia (CLL). We report our clinical, cytogenetic and fluorescence in situ hybridization (FISH) findings in a CLL patient with a duplication of part of the long arm of chromosome 12 between bands q13-q22. This patient was the only case with this duplication among the 112 cases (0.9%) of CLL cytogenetically analyzed in our laboratory. FISH studies using unique-sequence specific probes for the RB-1 (retinoblastoma) gene and the D13S319 locus at the 13q14 band showed a monoallelic loss for the D13S319 locus (20% of cells) with a diploid RB-1 gene. Our case showed an atypical morphology (35% prolymphocytes), a high proliferation rate and progression of the disease, indicating that the duplication of this region may be equivalent to complete trisomy 12 in CLL patients. PMID- 11279312 TI - Infant leukemia suggestive of natural killer cell precursor origin followed an unusual clinical course. AB - We report a patient with infant leukemia showing the rare phenotypes of CD2+, CD4+, CD7+, CD56+, CD3-, CD5- who followed an unusual clinical course. The patient was a 3-month-old girl who was admitted because of unusual purpura looking like a black ring. On admission WBC count was 85.8 x 10(9)/l and bone marrow aspiration revealed a nucleated cell count of 112.0 x 10(9)/l with 70.8% atypical lymphocytes. On the 3rd hospital day, the WBC count decreased by about 1/5 without chemotherapy and partial remission was obtained. But about 3 weeks later, the WBC count increased again and she died. Based on surface marker analysis, genotypic analysis and autopsy, we diagnosed infant leukemia suggestive of natural killer (NK) cell precursor origin. PMID- 11279313 TI - Evolution to acute myeloblastic leukemia from chronic neutrophilic leukemia with dysplastic features in granulocytic lineage. AB - We experienced the case of an 82-year-old man with chronic neutrophilic leukemia (CNL) with dysplastic features in the granulocytic lineage which subsequently progressed to acute myeloblastic leukemia (AML) with myelofibrosis. The patient had hepatosplenomegaly, but there was no evident cause of neutrophilic leukocytosis. The cytogenetic study showed that he had a normal karyotype. Concentrations of the serum granulocyte colony-stimulating factor (G-CSF) were not detectable. Two years after the diagnosis of CNL, blastic transformation to AML occurred with myelofibrosis and significant morphological abnormalities in neutrophils. The blasts were positive for myeloperoxidase, CD33, CD34, and HLA DR, and the presence of dysplasia within the granulocytic lineage suggested that he had an abnormality at the level of the granulocyte-committed progenitors. Heterogeneous origins of CNL might lead to various clinicopathological features in each case. PMID- 11279314 TI - Bilateral interstitial pneumonic shadows caused by perivascular fibrosis and extramedullary megakaryopoiesis of the lung in a case of advanced agnogenic myeloid metaplasia and myelofibrosis. AB - A 59-year-old man with progressive and advanced agnogenic myeloid metaplasia, also called idiopathic myelofibrosis, had complications showing bilateral interstitial pneumonic shadows. Pathological assessment of transbronchial biopsy revealed pulmonary perivascular fibrosis and infiltration of megakaryocytes. Autopsy 3 months later showed extramedullary megakaryopoiesis and fibrosis in lung, pleura, kidney, liver and spleen. Histopathological analysis for platelet derived growth factor (PDGF) and PDGF-receptor revealed an abnormally high expression of the PDGF-receptor-beta gene in pulmonary fibroblasts. This is the first description of an association between pulmonary fibrosis and PDGF in idiopathic myelofibrosis. PMID- 11279315 TI - Colchicine: an effective treatment for refractory malignant pericardial effusion. AB - We report a patient with AL amyloidosis and chemo-resistant light-chain multiple myeloma who developed a progressive malignant pericardial effusion leading to cardiac tamponade. Despite pericardiocentesis and surgical intervention, the pericardial effusion failed to resolve. The administration of oral colchicine produced symptomatic relief within 5 days, with the resolution of pericardial effusion after 14 days of treatment. As the administration of colchicine is simple and the side-effect is usually minimal, further studies are warranted to establish the feasibility of using colchicine to treat malignant pericardial effusion. PMID- 11279316 TI - Comparative study on complete remission rate and overall survival in three groups classified based on the serum interleukin-18 level in non-Hodgkin's lymphoma patients. PMID- 11279317 TI - A technical approach for optimizing surveillance of patients with unstable coronary syndromes: continuous vectorcardiography ischemic monitoring. AB - Continuous vectorcardiography ischemic monitoring (VCG) is a noninvasive technique which has contributed much to our understanding of the dynamics of acute coronary syndromes. For risk assessment of patients with unstable angina pectoris or non-Q-wave myocardial infarction, VCG shows important prognostic power which is further enhanced by its combination with the measurement of highly sensitive and specific biochemical markers of myocardial injury. VCG surveillance after revascularization therapy of Q-wave myocardial infarction is widely accepted because of its potential of assessing vessel patency, of estimating infarct size, and of revealing residual myocardial ischemia. Valuable prognostic information is available in real time with VCG, and treatment strategies may be based upon more reliable grounds. In this survey, the VCG technique is presented together with the most frequently used vectorcardiographic variables. Further, the use of the method for risk stratification of patients with unstable coronary syndromes is also described. PMID- 11279318 TI - Chronic use of continuous dosing of long-term nitrates does not prevent cardiac events in patients with severe acute myocardial infarction. AB - This study was performed to investigate the effects of the chronic use of continuous, not eccentric, dosing of nitrates on cardiac events in patients with severe acute myocardial infarction. A total of 1,303 patients with healed myocardial infarction were divided into two groups: treatment with nitrates or nontreatment. Primary end points were nonfatal and fatal recurrent myocardial infarction, death from congestive heart failure and sudden death. Among the 725 patients treated with nitrates, 45 patients (6.2%) experienced cardiac events during the observation period, whereas only 17 of the 578 patients treated without nitrates (2.9%) had cardiac events. This difference was statistically significant (p < 0.01; odds ratio 2.18, 95% confidence interval 1.24-3.86). In Killip class II-IV patients, the incidence of cardiac events in patients with nitrates was significantly greater than in patients without nitrates (18.8 vs. 3.5%, p < 0.05). The chronic use of continuous, not eccentric, dosing of nitrates did not prevent cardiac events in patients with severe acute myocardial infarction. PMID- 11279319 TI - Reduced heart rate variability in ischemic heart disease is only partially caused by ischemia. An HRV study before and after PTCA. AB - BACKGROUND: Reduced heart rate variability (HRV) after acute myocardial infarction (AMI) indicates poor prognosis. HRV in patients with uncomplicated coronary artery disease is reduced, and an association with poor prognosis has been suggested. The mechanism of the HRV reduction is not known, but ischemia is a possibility. AIM: To evaluate, in angina patients with no prior AMI, no other disease and drug-free, if complete revascularization and thus important reduction of ischemia by means of PTCA influences HRV. PATIENTS AND METHODS: Twenty-four hour Holter recordings were performed at baseline prior to PTCA in 48 patients with angina and in 41 age-matched healthy control subjects. The recording was repeated 1 and 6 months after complete revascularization. In addition, HRV was registered during controlled respiration in the supine and standing positions and during cold pressure test at baseline in all angina patients and controls and in 17 consecutive angina patients 6 months after PTCA. RESULTS: Compared to controls, angina patients had a significantly reduced mean RR interval (p = 0.02), SD (p = 0.003), rMSSD (p = 0.03), pNN50 (p = 0.03), total power (p = 0.003), low- (p = 0.004) and high-frequency peak (p = 0.04), but normal SDNN, SDANN and LF/HF. One and 6 months after PTCA, 42/46 and 32/40 follow-up patients, respectively, were free of angina. Six months after PTCA, there was a significant recovery of vagal modulation seen in the frequency domain during controlled respiration, but only nonsignificant trends in HRV parameters analyzed over 24 h. CONCLUSION: Patients with uncomplicated angina had reduced HRV, mainly affecting vagal activity, but normal low frequency variability associated with mortality. Complete revascularization caused a partial normalization of vagal modulation indicating that ischemia may be one of but not the only mechanism of the HRV reduction in uncomplicated chronic coronary artery disease. PMID- 11279320 TI - Randomized, double-blind, placebo-controlled, multicenter study of sibutramine in obese hypertensive patients. AB - OBJECTIVES: To compare weight loss efficacy, safety and tolerability of sibutramine and placebo in mildly to moderately obese hypertensive subjects; to assess the effect of weight loss on blood pressure. DESIGN: Randomized, double blind, parallel-group; 3-week placebo run-in and 12-week treatment phase. SETTING: Nine hospital outpatient clinics and general practices in the Netherlands. PARTICIPANTS: 127 men and women, 18-65 years old, with body mass indices (BMI) ranging from 27 to 40 kg/m(2) and stabilized hypertension - mean resting diastolic blood pressure of 90-120 mm Hg - with or without antihypertensive medication. INTERVENTIONS: Sibutramine 10 mg once daily; placebo. MAIN OUTCOME MEASURES: Body weight, blood pressure, routine laboratory and clinical safety monitoring. RESULTS: Of 113 evaluable patients, 54 received sibutramine and 59 placebo. Weight reduction was significantly greater with sibutramine from week 2 onwards (last observation carried forward): mean, 4.4 kg with sibutramine and 2.2 kg with placebo (p = 0.002); mean percentage weight reduction, 4.7 and 2.3%, respectively (p < 0.001); mean BMI reduction, 1.6 and 0.8 kg/m(2), respectively (p < 0.01). Reduction in excessive body weight was associated with a reduction in blood pressure in both groups, although the mean reduction in supine diastolic blood pressure was numerically, but not statistically significantly, greater in the placebo group (5.7 mm Hg) compared with the sibutramine group (4.0 mm Hg; p = 0.21). Similar reductions were seen in supine systolic blood pressure. Both treatments were well tolerated. CONCLUSIONS: Sibutramine 10 mg once daily is a useful, effective therapy for obesity in the presence of stable hypertension. PMID- 11279321 TI - Which lead for Q-T interval measurements? AB - There are several Q-T interval measures (individual lead, mean Q-T interval or Q T dispersion) from a 12-lead ECG. Which should be used? As the ECG provides twelve different measures of the heart's "true" Q-T interval, and as, "a priori", no one measure is any better or worse than any other measure at estimating the "true" Q-T interval, the best measure is the average of these twelve measures, i.e., the mean Q-T interval. The best single lead to measure the Q-T interval is the lead that relates most closely to the mean Q-T interval which in 49 subjects with cardiac diseases was lead V(3), then lead II. The longest lead Q-T interval relates poorly to the mean and to the individual lead Q-T intervals and therefore carried information different to and quite unique from these measures. This unique information is Q-T dispersion which relates well to the longest lead Q-T interval (r = 0.71, p < 0.001). PMID- 11279322 TI - False-negative and false-positive ECG diagnoses of Q wave myocardial infarction in the presence of right bundle-branch block. AB - Right bundle-branch block (RBBB) has not traditionally been seen as an obstacle to ECG diagnosis of Q wave myocardial infarction (MI)--in clinical electrocardiography and vectorcardiography--because this conduction disturbance is not believed to cause significant alterations in the spatial orientation of initial excitation wavefronts. In the era of large-scale clinical trials, however, where serial ECG analysis is among the major diagnostic tools in MI classification, both false-positive and false-negative diagnoses of MI in the presence of RBBB have become increasingly evident. Because of the limited detectability of Q wave MI by ECG in the presence of RBBB, the electrocardiographic finding of Q wave MI should not be regarded as an independent diagnostic tool. It is best to utilize independent corroboration to establish the diagnosis of transmural infarction when RBBB is present. Further investigations are warranted to better delineate sensitivity, specificity, and predictive value of Q wave MI in the presence of RBBB. PMID- 11279323 TI - Changes in myocardial perfusion after transmyocardial laser revascularization in patients with end-stage angina pectoris. AB - The purpose of the present study was to analyze the effects of transmyocardial laser revascularization (TMLR) on myocardial perfusion. The value of (99m)Tc-MIBI scintigraphy in the detection of changes in perfusion of the lased and nonlased segments was assessed as well. In 15 patients before TMLR and then 3 and 6 months afterwards, MIBI scintigraphy and a stress test were carried out. At the beginning of the study, all patients were classified as having angina pectoris class III or IV (according to the criteria of the Canadian Cardiac Society); their ejection fraction was >30%. The parameters of the stress test increased significantly in 70% of the patients. Cardiac scintigraphy revealed improved perfusion of 33.7% of the transient defects within 3 months after TMLR which persisted at 6 months with a clear trend towards further improvement in the lased segments. TMLR has been found to be particularly beneficial in patients in whom other invasive methods of treatment cannot be applied. PMID- 11279324 TI - Is the tissue affinity of ACE inhibitors of relevance for the remodeling of the left ventricular wall following myocardial infarction? Estimations with cine magnetic resonance imaging. AB - BACKGROUND: Angiotensin-converting enzyme (ACE) inhibitors have been shown to be of value in the treatment of postinfarction remodeling. The question whether substances with a greater tissue affinity are associated with advantages for the acute and the chronic course is, however, still unclear. AIM: The aim of the present study was to investigate the influence of ACE inhibitors with differing tissue affinities on the remodeling of the left ventricular wall in patients recovering from myocardial infarction. METHODS: 52 patients (17 women, aged 38-73 years) suffering their first acute myocardial infarction were randomized to receive a daily dose of either 25-75 mg captopril or 10-20 mg fosinopril, beginning on the 7th postinfarction day. 28 patients had an anterior wall infarction and 24 patients an inferior wall infarction. The size of the infarct was determined using the creatine kinase integral method. 50 patients were investigated by cine magnetic resonance imaging 1 and 26 weeks after the infarction. The following parameters were determined: infarct weight and diastolic diameter of the infarcted zone, systolic wall stress, muscle mass, diastolic and systolic diameters, systolic wall thickening, and motility of the noninfarcted myocardium. RESULTS: The infarct weight increased under captopril by 5.7% (p < 0.05) and under fosinopril by 6.1% (p < 0.05). The diastolic diameter of the infarcted zone decreased by 12% under captopril (p < 0.001) and by 11% under fosinopril (p < 0.001). The systolic wall thickness increased by 12.1% (p < 0.001) and the muscle mass by 12.7% (p < 0.001) under captopril and by 15.4% (p < 0.001) and 9.6% (p < 0.01), respectively, under fosinopril. Under captopril, the diastolic diameter increased by 2.3% (p < 0.05) and the systolic diameter by 17.8% (p < 0.01) and under fosinopril by 2.8% (n.s.) and 17.5% (p < 0.001), respectively. The systolic wall thickening increased by 73.9% under captopril (p < 0.001) and by 129.4% under fosinopril (p < 0.001). The motility decreased by 13.8% (p < 0.05) under captopril and by 6.0% (n.s.) under fosinopril. For all parameters, the results seen in anterior wall infarction were appreciably poorer than those seen in inferior wall infarction. All the differences between captopril and fosinopril were not significant. CONCLUSIONS: Captopril and fosinopril show no major differences in their influence on left ventricular wall remodeling following myocardial infarction. On the basis of the present results, the tissue affinity of an ACE inhibitor does not appear to be of a significant relevance for postinfarction treatment. PMID- 11279325 TI - Is the morning peak of acute myocardial infarction's onset due to sleep-related breathing disorders? A prospective study. AB - Many studies have shown that the risk of experiencing a myocardial infarction (MI) is increased during the first hours of the morning. Sleep apnea syndrome (SAS) is associated with an enhanced adrenergic activity, prolonged a few hours after awakening. We aimed at assessing whether sleep breathing disorders could be a culprit for the morning excess rate of MI. We studied 40 middle-aged men admitted for an acute MI. An overnight polysomnographic study was performed 37.4 +/- 9.4 days after the MI. The prevalence of SAS was high (30%). The prevalence of SAS was significantly higher in patients with the MI onset during the morning. The circadian pattern was significantly different in patients with or without SAS: those with SAS presented an important peak of MI onset during the period between 06.00 and 11.59 h. None of them had their MI during the period between 24.00 and 05.59 h. This different nyctohemeral pattern underlines the potential role of sleep breathing disorders as a trigger of MI. PMID- 11279327 TI - Intraventricular isovolumic relaxation flow patterns improve the predicting power of Doppler echocardiography for the left ventricular filling pressure in patients with anterior wall myocardial infarction. AB - BACKGROUND AND PURPOSE: Previous studies have shown that left ventricular systolic asynchrony affects both the relaxation and filling phases of diastole. The purpose of this study was to delinate how the anterior wall dyssynergy influenced the intraventricular flow redistribution patterns during the isovolumic relaxation (IVR) period, which delineated the changes in diastolic suction performance and, therefore, determined the significant Doppler flow variables for predicting left ventricular filling pressure. METHODS: Seventy three patients with anterior wall myocardial infarction and dyssynergy were enrolled. Those who exhibited the whole IVR intraventricular flow redistributing toward the mitral apparatus, which indicated the reverse physiologic intraventricular pressure gradient in early diastole, were classified as group B, otherwise, as group A. The Doppler echocardiographic variables of mitral inflow were correlated with the left ventricular end-diastolic pressures (LVEDP). RESULTS: With lower ejection fraction rate and more apical dyssynergy, the group B patients had much slower mitral flow propagation. For group A patients, the independent determinants for LVEDP were the ratio of mitral flow propagation rate to peak velocity in early diastole, the early mitral flow deceleration time and the IVR time, all occurring in early diastole. In contrast, the only independent determinant for LVEDP in group B patients was the ratio of mitral peak flow velocity in early diastole to that in late diastole. CONCLUSIONS: The intraventricular IVR flow patterns could delineate how the left ventricular systolic dyssynergy influenced the diastolic process, and determine which echocardiographic variables were more useful for predicting LVEDP in patients with anterior wall myocardial infarction. PMID- 11279326 TI - Technetium-99m sestamibi/tetrofosmin myocardial perfusion scanning in cardiac and noncardiac sarcoidosis. AB - Left ventricular (LV) and right ventricular (RV) involvement in sarcoidosis must be firmly confirmed to determine patients' prognosis. We examined whether myocardial perfusion images using technetium-99m single photon emission computed tomography (SPECT) have a diagnostic benefit in the evaluation of biventricular involvement. Sixteen patients with sarcoidosis, aged 21-78 (54 +/- 12) years old, 5 males and 11 females, complicated with cardiac disease (cardiac sarcoidosis, n = 6) including tachyarrhythmias of ventricular origin (n = 5), atrioventricular block (n = 4), and congestive heart failure (NYHA > or = II, n = 1), were enrolled in this study. Myocardial SPECT using technetium-99m sestamibi or tetrofosmin was performed and semiquantitatively scored for comparison with 25 control subjects. Perfusion abnormalities were more frequently recognized in sarcoidosis (LV 5/16, 31% and RV 14/16, 88% vs. LV 0/25, 0% and RV 8/25, 32% in controls). LV involvement had a close correlation with atrioventricular block and with congestive heart failure, and multiple sites of RV involvement correlated with ventricular tachyarrhythmia of RV origin. Total number of defect segments were highest in cardiac sarcoidosis (18/30, 60% vs. 19/60, 32% in noncardiac sarcoidosis, and 11/150, 7% in controls, p = 0.0001), and semiquantitatively evaluated total LV and RV scores (ranging from 0 to 18) were higher than those of controls (15.1 +/- 1.8 vs. 11.4 +/- 3.0 in noncardiac sarcoidosis, and 9.0 +/- 5.0 in cardiac sarcoidosis) and exhibited a significant positive linear correlation with the RV ejection fraction (y = 19.8 + 1.83x, r = 0.786, p = 0.001). Biventricular SPECT using technetium-99m is clinically useful for the noninvasive evaluation of both ventricular involvements in sarcoidosis. PMID- 11279328 TI - Ventricular fibrillation refractory to automatic internal cardiac defibrillator in Fabry's disease. Review of cardiovascular manifestations. AB - Fabry's disease is a disorder of glycosphingolipid metabolism leading to alpha galactosidase deficiency with systemic sequelae. Clinical cardiac manifestations include dysrhythmias, structural abnormalities apparent on echocardiography, and histologic changes secondary to glycosphingolipid deposition. The introduction of automated internal cardiac defibrillators (AICD) has been shown to decrease the incidence of circulatory collapse in individuals with known terminal arrhythmias. We present a patient with Fabry's disease, who underwent coronary angiography without finding of obstructive disease. He returned after aborted sudden cardiac death necessitating the placement of an AICD. He again presented after an episode of ventricular fibrillation refractory to internal defibrillation necessitating advanced life support, and subsequently expired. We review the electrocardiographic, cardiovascular structural, and histologic manifestations of Fabry's disease. PMID- 11279329 TI - Neuropsychiatric toxicity of antiviral treatment in chronic hepatitis C. AB - Dose-dependent, reversible neuropsychiatric toxicity is reported in up to 30-40% of chronic hepatitis C patients treated with 6-12 months of interferon-alpha or interferon-alpha plus ribavirin combination therapy. Although risk factors remain poorly defined, neuropsychiatric side effects may be severe and dose-limiting in as many as 10-20% of treated patients. Diagnosis relies upon the detection of clinically apparent neuropsychiatric symptoms and the emerging use of self administered mood inventories and questionnaires. The current stepwise approach to management includes evaluation and treatment of systemic side effects, early use of adjuvant medications, and interferon-alpha and ribavirin dose reduction or cessation with psychiatric referral in selected cases. Although the cellular basis of the neuropsychiatric toxicity of interferon-alpha remains unknown, several hypothesis involving changes in central adrenergic, seratonergic, opioid and neuroendocrine pathways have been proposed. Recognition and management of the neuropsychiatric side effects of antiviral therapy will be of growing clinical importance as additional patients with chronic hepatitis C are treated and longer durations of therapy are utilized. PMID- 11279330 TI - Critical review of acid suppression in nonvariceal, acute, uppergastrointestinal bleeding. AB - Nonvariceal, upper gastrointestinal (GI) bleeding is a very common source of morbidity and mortality. The concept of ulcer clot dissolution being facilitated by a low gastric pH has allowed us to better understand the pathophysiology of nonvariceal upper GI bleeding. Placebo-controlled trials have shown the benefit of oral proton pump inhibitor administration in contrast to H(2) receptor antagonists. Furthermore, our recent experience with intravenous proton pump inhibitors has reinforced these observations. PMID- 11279331 TI - Selection of patients for laparoscopic antireflux surgery. AB - Since the first laparoscopic fundoplication was performed, the frequency of antireflux surgery has increased rapidly with some centers now having an experience of about 1,000 procedures. The question arises whether this increase is due to a change in indications for the surgical treatment of gastrointestinal reflux disease (GERD) despite the simultaneous appearance of powerful antisecretory medications. Adequate knowledge of the pathophysiology of GERD is necessary in order to establish selection criteria for patients suitable for laparoscopic antireflux surgery. In this article, we review the epidemiology and pathophysiology, and provide a rationale for medical and surgical treatment. We also offer an approach to patient selection for antireflux surgery. PMID- 11279332 TI - Update on noncardiac chest pain. AB - Patients with recurrent angina-like chest pain with normal coronary vessels are deemed to have the syndrome of noncardiac chest pain (NCCP). These patients, despite having significant cardiac disease ruled out, often spend a restricted lifestyle believing they have cardiac disease. These recurrent episodes of chest pain may be related to gastroesophageal reflux disease (GERD), spastic motility disorders of the esophagus and esophageal (visceral) hyperalgesia. These disease entities are often difficult to diagnose and treat except for GERD and achalasia. Recent prospective double-blind studies have shown that about 44% of these patients may have underlying GERD. There is now more evidence to support the practice of empiric use of proton pump inhibitors (PPIs) as the first step in therapy. Newer modalities for diagnosis like endoscopic ultrasonography (EUS) showed that this group of patients had sustained muscular contractions of longer that 68 s during chest pain. These sustained contractions noted on EUS were secondary to isometric contraction of the circular muscle which did not cause luminal constriction nor was related to contraction of the longitudinal muscles which cannot be recorded by pressure manometry. Treatment is difficult if patients do not respond to high-dose PPIs. Other medications which are known to alter visceral hyperalgesia in low doses, such as tricyclic antidepressants like imipramine and desyrel, can be tried. Psychological intervention may be useful in the management of some of these patie PMID- 11279333 TI - Clostridium difficile infection: risk factors, medical and surgical management. AB - BACKGROUND: Clostridium difficile has become recognized as a cause of nosocomial infection which may progress to a fulminant disease. METHODS: Literature review using electronic literature research back to 1966 utilizing Medline and Current Contents. All publications on antibiotic-associated diarrhea, antibiotic associated colitis, and pseudomembranous colitis as well as C. difficile infection were included. We addressed established and potential risk factors for C. difficile disease such as an impaired immune system and cost benefits of different diagnostic tests. An algorithm is outlined for diagnosis and both medical and surgical management of mild, moderate and severe C. difficile disease. RESULTS: Diagnosis of C. difficile infection should be suspected in patients with diarrhea, who have received antibiotics within 2 months or whose symptoms started after hospitalization. A stool specimen should be tested for the presence of leukocytes and C. difficile toxins. If this is negative and symptoms persist, stool should be tested with 'rapid' enzyme immunoabsorbent and stool cytotoxin assays, which are the most cost-effective tests. Endoscopy and other imaging studies are reserved for severe and rapidly progressive courses. Oral metronidazole or vancomycin are the antibiotics of choice. Surgery is rarely required for selected patients refractory to medical treatment. The threshold for surgery in severe cases with risk factors including an impaired immune system should be low. CONCLUSION: C. difficile infection has been recognized with increased frequency as a nosocomial infection. Early diagnosis with immunoassays of the stool and prompt medical therapy have a high cure rate. Metronidazole has supplanted oral vancomycin as the drug of first choice for treating C. difficile infections. PMID- 11279334 TI - Role of thrombotic vascular risk factors in inflammatory bowel disease. AB - Thromboembolic disease is a significant cause of morbidity and mortality in patients with inflammatory bowel disease (IBD). It is recognized that a hypercoagulable state exists in IBD which involves all components of the clotting system. It has been suggested that this hypercoagulable state is closely linked to the disease pathogenesis. Recent studies have shown that genetic defects such as factor V Leiden mutation and C677T methylenetetrahydrofolate reductase polymorphism associated with hyperhomocysteinemia seem to interfere in the thrombotic manifestations of IBD. Acquired factors such as antiphospholipid antibodies could also participate in the development of the thrombotic process. Deficiencies of other anticoagulant factors play a less important role in the thrombosis, and therefore it is not surprising that the results on these factors in IBD are contradictory. In conclusion the resultant gene-gene and gene environment interactions between risk factors are the key to the understanding of why an IBD patient develops thrombosis at a specific point in time. PMID- 11279335 TI - Influence of chronic alcohol abuse on hepatitis C virus replication. AB - BACKGROUND: Patients with alcoholic liver disease have a high prevalence of hepatitis C virus (HCV) infection. Several workers have shown that HCV-infected alcoholics have more severe biochemical and histological evidence of liver disease than anti-HCV-negative patients. One possible mechanism for the increased liver damage is that alcohol may have a stimulatory effect on HCV replication. The present study was carried out to examine this issue in detail. METHODS: Sixty eight HCV-infected patients, comprising of 50 chronic alcoholics, consuming 80 g or more of alcohol daily for at least 5 years, and 18 completely abstinent subjects were included in the study. Quantitative HCV-RNA was performed by the branched chain DNA (bDNA) technique. RESULTS: There was no significant difference in the mean serum HCV titers in chronic alcoholics compared to nonalcoholic subjects. Linear regression analysis showed no correlation between the daily ethanol consumption and HCV titers. Seven of the chronic alcoholics, 4 of whom were continuing to drink and 3 who had become abstinent, were retested after 6 months. There was no definite trend in the viral titers, either in abstinent individuals or in those who continued to drink. CONCLUSIONS: These findings suggest that chronic alcohol abuse does not influence the HCV load in the serum. Therefore, the observation that alcoholics with HCV infection have more severe liver damage requires some other explanation than increased HCV viral titers. PMID- 11279336 TI - Manometric components of the lower esophageal double hump. AB - BACKGROUND/AIMS: The lower esophageal sphincter manometry of patients with hiatal hernia often displays a double hump configuration. It seems that this is due to gastric herniation above the high-pressure zone of the crura. This study examines this manometric phenomenon in patients with hiatal hernia and relates it to the lower esophageal antireflux barrier. METHODS: Manometric and 24-hour pH studies of 68 consecutive patients with suspected gastroesophageal reflux disease were analyzed to obtain information regarding the double hump and acid reflux. RESULTS: The findings of a manometric double hump correlated well with the presence of a hiatal hernia of >5 cm. The overall length of the sphincter complex was greater in patients with a double hump, but the length below the respiratory inversion point was constant. Resting pressures at the respiratory inversion point were significantly lower than those measured at either high-pressure zone. The location of the respiratory inversion point was seen most commonly at the superior margin of the distal high-pressure zone. Double hump patients with a negative acid reflux score were found to have higher pressures in the distal high pressure zone than those patients with acid reflux. CONCLUSIONS: The two high pressure zones comprising the manometric double hump represent the crural and muscular components of the lower esophageal sphincter. Descriptive information regarding the double hump phenomenon is given, and the importance of the crural component of the lower esophageal sphincter in preventing acid reflux is stressed. PMID- 11279337 TI - Prevalence of gastroesophageal reflux disease in patients with extraesophageal symptoms referred from otolaryngology, allergy, and cardiology practices: a prospective study. AB - AIM: To investigate the prevalence of gastroesophageal reflux disease (GERD) as well as the clinical, endoscopic, and manometric characteristics in 57 adult patients with otolaryngeal symptoms, asthma, or noncardiac chest pain referred from specialized services. METHODS: The following evaluations were performed: (1) upper endoscopy, (2) 24-hour ambulatory esophageal pH monitoring, and (3) esophageal manometry. The prevalence of GERD was determined, and demographic, clinical, endoscopic, and manometric characteristics of patients with or without GERD were evaluated. RESULTS: Thirty-four out of 57 patients (60%) had GERD. The 95% confidence interval ranged from 48 to 72%. There was no statistical difference between patients with or without GERD regarding gender, age, or time of evolution of symptoms. Cough was more frequent in the subjects with GERD (75 vs. 25%, p<0.05). Nevertheless, cough was observed in only 53% of the patients with GERD. Patients suffering from laryngitis had a greater proximal and distal esophageal acid exposure time than those without. CONCLUSIONS: The prevalence of GERD was 60%. There is not a definite demographic or clinical profile that permits us to distinguish between patients with and without GERD among those with ear, nose, and throat and pulmonary symptoms or chest pain. PMID- 11279338 TI - Gastric metaplasia of the proximal esophagus associated with esophageal adenocarcinoma and Barrett's esophagus: what is the connection? Inlet patch revisited. AB - BACKGROUND/AIM: The inlet patch is an area of heterotopic gastric mucosa found in the proximal esophagus at the level of the upper esophageal sphincter. Limited data are available regarding this form of gastric metaplasia and its incidence, significance, and possible association with other esophageal diseases. We report our observations of such gastric metaplasias in patients with esophageal adenocarcinoma or Barrett's esophagus and high-grade dysplasia. METHODS: All patients having Barrett's esophagus and adenocarcinoma referred for photodynamic therapy were included in this study. The patients were prospectively evaluated endoscopically for the presence of gastric metaplasia of the proximal esophagus (salmon-colored area of a least 5 mm in diameter with cardia-type gastric metaplasia on biopsy). RESULTS: A total of 36 patients were included in this study: 11 patients with dysplastic Barrett's esophagus (8 males, mean age 79 years) and 25 adenocarcinoma patients (18 males, mean age 71 years). At endoscopy prior to photodynamic therapy, 11 patients (31%; 8 adenocarcinoma, 3 dysplastic Barrett's esophagus) were noted to have an area of gastric mucosa in the proximal esophagus. In each patient, there was at least 5 cm of normal squamous mucosa between gastric metaplasia and distal esophageal pathology. CONCLUSIONS: In this selected group of patients with high-grade dysplastic Barrett's esophagus or adenocarcinoma referred for photodynamic therapy, gastric metaplasia of the proximal esophagus was found in nearly one third. Prospective studies are under way to test more widely for this association and to determine whether this is a marker of disease severity and the result of similar pathogenetic mechanisms. PMID- 11279339 TI - Congenital esophageal stenosis. PMID- 11279340 TI - Jejunal diverticuli complicated by bacterial overgrowh syndrome. PMID- 11279342 TI - Oligoclonality of early lesions of the urothelium as determined by microdissection-supported genetic analysis. AB - AIM: To contribute to the ongoing discussion of clonality of human urothelial cancer it was considered a valuable approach to analyze multiple areas from cystectomy specimens for deletions of chromosomes known to be involved early in bladder cancer development. MATERIAL AND METHODS: Thus, in 86 biopsies of 4 human cystectomies with different histological findings (maximal diagnosis: pT1G2, pTaG3, pT2G2, normal) loss of heterozygosity (LOH) was investigated as a deletion marker using markers of chromosomes 8p, 9p, 9q and 17p. Findings were compared to histology of the lesion. RESULTS: Findings indicate: (1) no changes in the markers investigated in the bladder with histologically normal urothelium in contrast to detection of LOH in normal urothelium of tumour-bearing bladders; (2) an accumulation of the number of LOH with increasing malignancy of lesions within one bladder, and (3) indications of oligoclonal neoplastic lesions in two of the urinary bladders investigated. CONCLUSIONS: The investigation of multiple lesions within one bladder presents a snapshot of genetic changes in differently advanced tumour stages. The hypotheses of tumour evolution and oligoclonality as derived from our LOH data need to be supported by deletion-independent clonality studies as X-chromosomal inactivation analysis. PMID- 11279343 TI - Detection of gene amplification in intraductal and infiltrating breast cancer by laser-assisted microdissection and quantitative real-time PCR. AB - Gene amplification is one essential mechanism leading to oncogene activation which is supposed to play a major role in the pathogenesis of invasive breast cancer. However, using standard methodologies the detection of gene amplifications has been limited especially in small-sized lesions, like pre invasive precursor lesions. The combination of two novel technologies, laser based microdissection and quantitative real-time PCR, facilitates the detection of low-level amplifications in morphologically defined lesions. As a model system we investigated in situ breast cancer (ductal carcinoma in situ, DCIS) classified according to the morphology-based Van Nuys grading system for amplification of growth-regulatory genes. In this study 83 formalin-fixed, paraffin-embedded archival DCIS specimens were examined after laser-based microdissection by quantitative real-time PCR using the TaqMan detection system for amplification of the c-erbB2, topoisomerase IIalpha, c-myc and cyclinD1 gene. In a subset of 17 DCIS with adjacent infiltrating tumour components we compared intraductal and invasive tumour components in parallel for differences in amplification status. The combination of these new techniques represents an excellent tool to gain new insights into carcinogenesis by analyzing genetic alterations in morphologically identified heterogeneous lesions in breast cancer progression within the very same specimen or even tissue slide. PMID- 11279344 TI - Laser microdissection and microsatellite analyses of breast cancer reveal a high degree of tumor heterogeneity. AB - Carcinomas with productive fibrosis are the most common forms of breast cancer. Analysis of tumor-specific genomic alterations can be compromised by the presence of normal cells, demanding microdissection of small tumor areas to detect loss of heterozygosity (LOH) and microsatellite instability (MSI). The aim of this study was to evaluate the importance of precise laser microdissection for microsatellite analyses and investigation of tumor heterogeneity in breast cancer. 39 primary breast tumor samples were analyzed for MSI and LOH by PCR followed by polyacrylamide gel electrophoresis and silver staining using 15 microsatellite markers. Different tumor areas were processed separately in 30 patients. Both intraductal and invasive breast cancer regions were investigated in 11 patients. The following results were obtained: (1) accurate microdissection revealed MSI in 3 or more of the investigated markers (> or =20%) in 33% of the patients, a higher frequency than reported previously; (2) laser microdissection was 43% more sensitive in detection of LOH compared to manual microdissection due to a reduction of contamination by normal cells, and (3) 29 of 30 investigated tumors showed heterogeneity of genetic alterations in different tumor regions. Laser-based microdissection is a valuable tool in genetic analysis of desmoplastic tumors and allows an accurate determination of genetic alterations in histologically different tumor regions. PMID- 11279345 TI - Cell type-specific mRNA quantitation in non-neoplastic tissues after laser assisted cell picking. AB - OBJECTIVE: Cell type-specific mRNA quantitation can be reliably performed after harvesting less than 20 cell profiles from haemalaun-stained cryosections by laser-assisted cell picking. Up to now it has been unclear to what extent these techniques can be used to analyze differential gene expression in complex tissues. METHODS: Using a rat model of experimental endotoxin priming of the lung various pulmonary cell types were microdissected from isolated perfused and ventilated rat lungs after aerosol lipopolysaccharide/interferon-gamma stimulation. RESULTS: Porphobilinogen deaminase housekeeping gene (PBGD) and nitric oxide synthase II (NOSII) mRNA in arterial endothelial cells (AEC), bronchiolar epithelial cells (BEC), alveolar septum containing monocytes/macrophages (AS+), alveolar septum without monocytes/macrophages (AS-) and intraluminar alveolar macrophages (AM) could be quantified by real-time RT PCR. The strongest upregulation of NOSII mRNA occurred in AM, while minimal NOSII expression was detected in BEC, AS+ and AS-. In AEC NOSII mRNA was not detectable. CONCLUSION: The combination of laser microdissection and real-time RT PCR is a valuable tool for the quantitative in situ characterization of differential gene expression within complex tissues. PMID- 11279346 TI - Laser-assisted microdissection and short tandem repeat PCR for the investigation of graft chimerism after solid organ transplantation. AB - The detection of donor-derived cells in the blood and tissues of graft recipients after solid organ transplantation is a readily observed phenomenon called microchimerism. Yet very little is known about the persistence and integration of recipient-derived cells in the transplanted organ, indicating a form of intragraft chimerism. To further study this phenomenon and its possible influence on graft acceptance or rejection, we developed the following novel approach. Immunohistochemically labeled cells were isolated by means of laser-based microdissection and subsequent laser pressure catapulting from paraffine-embedded posttransplantation biopsies. The following use of a highly sensitive PCR assay analyzing one polymorphic short tandem repeat (STR) marker enabled us to clearly identify the genotypes in samples containing as little as 10 isolated cells. The combination of laser-based microdissection and STR-PCR thus provides a powerful tool for the genotyping of even very few cells isolated from routinely processed biopsies after solid organ transplantation. PMID- 11279347 TI - Quantitative molecular analysis of laser-microdissected paraffin-embedded human tissues. AB - Laser microdissection enables the contamination-free isolation of morphologically defined pure cell populations from archival formalin-fixed paraffin-embedded tissue specimens. Cells isolated by this method have been characterized by a wide variety of qualitative molecular assays, e.g. loss of heterozygosity, point mutations, clonality and lineage origin. The recently introduced real-time PCR technology renders the reliable quantification of very small amounts of nucleic acids possible. Several groups including our own showed that this technique can be successfully applied for the quantification of DNA and RNA isolated from microdissected archival tissue sections, even after immunohistochemical staining. The exact analysis of quantitative changes of nucleic acids during the course of pathological alterations has thus become possible. In many situations these quantitative changes can be expected to be more important than qualitative changes. The new technology for the quantification of structural genomic alterations and changes in the gene expression pattern in conjunction with microdissection have equipped morphologists with a powerful tool to study reactive and neoplastic changes of tissues. PMID- 11279348 TI - Laser capture microdissection: methodical aspects and applications with emphasis on immuno-laser capture microdissection. AB - Laser capture microdissection (LCM) is an easy, extremely fast and versatile method for the isolation of morphologically defined cell populations from complex primary tissues for molecular analyses. However, the optical resolution is limited due to the use of dried sections without coverslip necessary for tissue capture, and routine stains such as hematoxylin and eosin are sometimes insufficient for precise microdissection, especially in tissues with diffuse intermingling of different cell types and lack of easily discernible architectural features. Therefore, several groups have adapted immunohistochemical staining techniques for LCM. In addition to providing high contrast targets for microdissection, immunostaining allows selection of cells not only according to morphological, but also phenotypical and functional criteria. In order to allow reliable tissue transfer on one hand and preserve the integrity of the target of analysis such as DNA, RNA and proteins on the other hand, immunostaining protocols have to be modified for the purposes of LCM. The following review gives an overview of immuno-LCM and describes some applications, e.g. in the field of hematopathology. PMID- 11279349 TI - Recovering DNA and optimizing PCR conditions from microdissected formalin-fixed and paraffin-embedded materials. AB - Microdissection is a powerful technique in molecular pathology and genetic investigations. To detect genetic alterations such as gene mutation or deletion from tumor specimen, the purity of target cells is extremely critical. Unwanted cell contamination will dramatically dilute the detectable level of the abnormality. The major obstacle in clinical research is to obtain sufficient and qualified DNA from a small amount of formalin-fixed and paraffin-embedded materials. We have successfully modified our previous protocols and overcome the difficulties of recovery of DNA. After these modifications, almost every single formalin-fixed and paraffin-embedded specimen has been successfully amplified in the required DNA region. PMID- 11279350 TI - Diagnosis of papillary thyroid carcinoma is facilitated by using an RT-PCR approach on laser-microdissected archival material to detect RET oncogene activation. AB - OBJECTIVE: The purpose of this study was to investigate the value of the expression of the RET oncogene (rearranged during transfection) in papillary thyroid carcinomas (PTC) and its variants in the differential diagnosis of thyroid neoplasias. According to the literature RET oncogene activation by chromosomal rearrangements has been exclusively implicated in PTCs. METHODS: To establish the incidence of RET activation in PTCs we used 5- to 10-microm sections from archival paraffin blocks. Either parts of the tissue slices were manually dissected or a few distinct cells were microdissected by laser-mediated manipulation with the Robot-MicroBeam system. RNA was extracted from paraffin embedded thyroid tumors and the corresponding normal tissue. RT and nested PCR were performed using primers for RET/PTC1, PTC2 and PTC3, or for RET exons 12 and 13. PCR products were resolved by gel electrophoresis. RESULTS: We detected RET transcription in approximately 85% of the PTCs including follicular variants and in isolated cells of the same tissues, but not in nonmalignant thyroid tissue. CONCLUSIONS: Our method may serve as an additional diagnostic tool to characterize ambiguous neoplasias and to identify especially nonpapillary, i.e. follicular tumors, as papillary carcinomas. Additionally, this study has demonstrated that expressed genes can be analyzed from routine histopathological tissue slides or pooled single cells. Large retrospective studies can also be performed with this method. PMID- 11279351 TI - Microsatellite instability in tumor and nonneoplastic colorectal cells from hereditary non-polyposis colorectal cancer and sporadic high microsatellite instable tumor patients. AB - Genetic alterations such as loss of heterozygosity (LOH) and microsatellite instability (MSI) have been frequently studied in various tumor types. Genetic heterogeneity of nonneoplastic cells has not yet been sufficiently investigated. However, genomic instability in normal cells could be a potentially important issue, in particular when these cells are used as reference in LOH and MSI analyses of tumor samples. In order to investigate possible genetic abnormalities in normal colorectal cells of tumor patients, MSI analyses of normal colonic mucosa were performed. Up to 15 different laser-microdissected normal regions containing 50-150 cells were investigated in each of 15 individual microsatellite stable, sporadic high microsatellite-instable (MSI-H) and hereditary non polyposis coli cancer (HNPCC) colorectal cancer patients. Frequent MSI and heterogeneity in the MSI pattern were found both in normal and tumor cells from 10 HNPCC and sporadic MSI-H tumor patients whose tumors had defect mismatch repair protein expressions. This observation shows that MSI can also occur in nonneoplastic cells which has to be considered in MSI analyses for molecular HNPCC screening. In addition, considerable genetic heterogeneity was detected in all MSI-H (sporadic and HNPCC) tumors when analyzing five different regions with less than 150 cells, respectively. These differences were not detectable in larger tumor regions containing about 10,000 cells. Thus, heterogeneity of the MSI pattern (e.g. intratumoral MSI) is an important feature of tumors with the MSI-H phenotype. PMID- 11279352 TI - Laser microdissection as a new approach to prefertilization genetic diagnosis. AB - The genetic status of oocytes can be determined by polar body (PB) analysis. Following PB extraction, a genetic evaluation is performed. As each PB contains the complementary genetic material of the oocyte, PB analysis reveals information about its genetic status. Genetically altered oocytes may then be excluded from in vitro fertilization. The aim of our study was to evaluate laser microdissection as a tool for PB extraction purposes. Compared to the PB extraction with a sharp-ending pipette only, we could show that laser microdissection of the zona pellucida (laser zona drilling) with a UV-A laser and subsequent extraction with a blunt-ending pipette decreases the degeneration rate of oocytes. It is shown that laser pressure catapulting of extracted PB enables their contact-free transfer into tubes, thus decreasing the risk of contamination for further analysis. PMID- 11279353 TI - Forty-five years of stereotactic surgery for Parkinson's disease: a review. AB - An overview of a large experience of the surgical management of parkinsonism from 1954 to 1999 is outlined--from the original open surgery of Fenelon to the sophisticated, motor-driven electrodes of stereotactic surgery using depth microelectrode recording. The definition of the best target sites for the relief of tremor, rigidity and bradykinesia evolved progressively as accuracy in siting of lesions developed. The significance of these targets in understanding the pathophysiology of the disease and the neurotransmitters involved gradually became clearer and a way forward in future management was suggested. PMID- 11279354 TI - Effects of anterodorsal pallidal stimulation on gait freezing (Kinesia paradoxa) in Parkinson's disease. AB - The results of a double-blind evaluation of the effects of internal globus pallidus (GPi) stimulation in 7 patients with advanced Parkinson's disease are summarized. The evaluation was performed 6-8 months after surgery while the patients were on medication with an optimal dose and schedule. The stimulation was turned off for at least 12 h. It was turned on in the morning (or maintained turned off), and the best and worst scores during their daytime activity were recorded as the on-period and off-period scores, respectively. A significant reduction in the total score on part III of the Unified Parkinson's Disease Rating Scale was induced by GPi stimulation at the off-period (-57%) as well as the on-period (-36%). Clinically important improvement was also achieved in severe gait freezing (kinesia paradoxa) in 2 patients when stimulation was applied to the anterodorsal portion of the GPi. Such an effect was observed during unilateral stimulation of the right side alone. PMID- 11279355 TI - Acoustic neuroma--surgery or radiosurgery? AB - This paper discusses surgery and radiosurgery from the author's personal experience with both techniques. Multi-discipline assessment will allow the patient to make an informed decision regarding preferred management. PMID- 11279356 TI - Generalized seizures induced by an epileptic focus in the mesencephalic reticular formation: impact on the understanding of the generalizing mechanism. AB - The mode of seizure propagation was studied using a generalized seizure model induced by microinjection of kainic acid (KA) into a unilateral mesencephalic reticular formation (MRF) in cats and rats. Stereotactic surgery was performed under pentobarbital anesthesia; an injection cannula was placed into a unilateral MRF, and bipolar electrodes were implanted into the MRF and the thalamus. Microinjection of KA induced generalized seizures. Focal electrical seizures were elicited in the injected site of the MRF starting 30 min after the injection. The initial clinical change during each seizure was behavioral arrest. These seizures immediately developed to generalized seizures, which were characterized by generalized tonic convulsions with short-term clonic convulsions. On EEG, each generalized seizure started at the same time in all the sites of the brain recorded. Autoradiographic study using a rat model demonstrated high glucose utilization in the MRF, bilateral thalamus, forebrain and bilateral cerebral cortices. The results demonstrated an active participation of MRF in the mechanism of generalized seizures. PMID- 11279357 TI - Microvascular decompression in trigeminal neuralgia: a correlation of three dimensional time-of-flight magnetic resonance angiography and surgical findings. AB - We analyzed 104 patients with trigeminal neuralgia who underwent microvascular decompression and who were followed up for more than 12 months during the period from January 1992 to June 1998. In this recent series, we utilized three dimensional time-of-flight magnetic resonance angiography (3D-TOF MRA) for all patients with trigeminal neuralgia. A 3D-TOF MRA was beneficial in treatment planning and in predicting surgical outcome by demonstrating cranial nerve compression as well excluding other etiologies such as tumor or vascular lesions. The patients were followed up for 1-7 years (mean 5.7 +/- 1.2 years). Initial pain relief was complete in 89 patients (85.6%) and partial in 12 patients (11.5%). There were three primary failures (2.9%). The acceptable pain relief rate (complete relief: 79.8%, partial relief: 11.5%) was determined in the long term follow-up of surgical results. Pain recurred in 6 patients (5.8%). The mean time to recurrence was 48 months (36-93 months). There were no serious or annoying complications such as anesthesia dolorosa. PMID- 11279358 TI - Multimodality program involving stereotactic surgery in brain tumor management. AB - Stereotactic and image-guided surgery is becoming increasingly important in the management of brain tumors. Although there are several stereotactic modalities that have been reported to be of value, it is the combination of techniques in a multimodality approach that seems to show the most promise. Both frame-based and frameless guidance may facilitate glioma resection, allowing the optimal amount of resection while permitting avoidance of surrounding eloquent areas. Not only does this optimize resection, but leaving a minimal amount of gross tumor may provide a better bed for intracavitary chemotherapy. Deep tumors may be localized and approached through a small channel, and surgical exposure may be minimized to protect uninvolved areas of the brain. There is increasing evidence that patients operated with imaging guidance have a more benign course and more rapid discharge, perhaps with a lower incidence of adverse neurological sequelae. Stereotactic conformal radiotherapy allows a higher tumor dose while sparing uninvolved brain from radiation more efficiently than conventional techniques, and residual tumor may be treated with a boost of stereotactic radiotherapy. Stereotactic instillation of radioisotopes may be used to treat cystic tumors. Stereotactic insertion of cannulae or radioisotope seeds permit efficient brachytherapy. Stereotactic surgery has moved beyond a subspeciality, so that every neurosurgeon might benefit from using stereotactic techniques in brain tumor management. PMID- 11279359 TI - The idea of stereotaxy toward minimally invasive neurosurgery. AB - The idea of stereotaxy in modern neurosurgery is reviewed. Stereotactic surgery has been one of the particular neurosurgical techniques mainly used for functional disorders. Nowadays, it is widely used in the field of general neurosurgery owing to the rapid development of computer-assisted 3D brain images. Functional neurosurgery itself is also changing in the sense that many alternative surgical procedures are now available due to the progress in neuroscience. The original premise of stereotaxy is exactly the same as that of minimally invasive neurosurgery today. PMID- 11279360 TI - Computer-aided/image-guided and video-endoscopic resection of pituitary tumors. AB - Endoscopic pituitary surgery is a minimally invasive method which allows simple transnasal transphenoidal surgical access without retractors, and has been used by our team in 536 patients from 1993 to 1999. In 326 of these patients, computer aided image-guided techniques (CAN) were used in conjunction with endoscopy. We found that CAN techniques and endoscopy allow accurate and precise minimal access to small, deep/lateral microadenomas and enable a more complete resection of large pituitary tumors especially those which are firm/fibrous in nature. Overall results were better than conventional pituitary surgery with a lower morbidity and shorter hospital stays. However, special training and expertise is needed for surgeons to carry out these procedures with acceptable results. PMID- 11279361 TI - Neurosurgery for spasticity. AB - Spasticity is usually a useful substitute for deficiency of motor strength. However not infrequently, it may become harmful leading to an aggravation of motor disability. When excessive spasticity is not sufficiently controlled by physical therapy and pharmacological treatment, patients can have recourse to neurosurgery: neurostimulation, intrathecal baclofen or selective ablative procedures. Because excessive hypertonia has to be reduced without suppression of the useful muscular tone or impairment in the residual motor and sensory functions, neuroablative procedures must be as selective as possible. These selective lesions can be performed at the level of peripheral nerves, spinal roots, spinal cord, or the dorsal root entry zone (DREZ lesions). PMID- 11279362 TI - Expression of nerve growth factor and its receptors in the somatosensory-motor cortex of Macaca nemestrina. AB - The present study was designed to examine the nerve growth factor (NGF) system (ligand and receptor-expressing neurons) in the somatosensory (areas 1, 3a, and 3b) and motor (area 4) cortices of the mature macaque. Light and electron microscope immunohistochemistry was used to assess the distribution and identity of NGF-, p75-, and trk-expressing elements. In each cortical area examined, NGF positive neuronal somata were distributed through all laminae; most immunolabeled neurons were in layers II, III, and V. Based upon light microscope criteria (e.g., the morphology of proximal dendrites), both pyramidal and stellate neurons expressed NGF. Of the identifiable NGF- immunoreactive cells, 92% were pyramidal neurons and the remainder was stellate neurons. The electron microscope study showed that most (88%) NGF-positive somata formed symmetric synapses, whereas the others formed both symmetric and asymmetric synapses. As the somata of pyramidal neurons form only symmetric synapses and those of inhibitory stellate neurons form both symmetric and asymmetric somatic synapses, the ultrastructural data support the light microscopic analyses. In contrast, neurotrophin receptors, p75 and trk, were expressed chiefly by the cell bodies of layer V pyramidal neurons and the supragranular neuropil. At the ultrastructural level, receptor-positive profiles were post-synaptic elements (e.g., dendritic shafts and spines) and the concentration of immunoreactivity was greatest in the vicinity of post-synaptic densities. Thus, NGF regulatory systems parallel excitatory and inhibitory neurotransmitter systems. Cortex contains the morphological framework by which pyramidal and/or inhibitory stellate neurons can affect the activity of post synaptic pyramidal neurons via anterograde and autocrine/paracrine NGF systems. PMID- 11279363 TI - Gecko vision-visual cells, evolution, and ecological constraints. AB - Geckos comprise both nocturnal and diurnal genera, and between these categories there are several transitions. As all geckos depend on their visual sense for prey capture, they are promising subjects for comparison of morphological modifications of visual cells adapted to very different photic environments. Retinae of 22 species belonging to 15 genera with different activity periods are examined electron microscopically. Scotopic and photopic vision in geckos is not divided between "classical" rods and cones, respectively; both are performed by one basic visual cell type. Independent of the activity periods of the individual species, the visual cells of geckos exhibit characteristics of cones at all levels of their ultrastructure. Thus, gecko retinae have to be classified as cone retinae. Only the large size and the shape of the photoreceptor outer segments in nocturnal geckos are reminiscent of rods; the outer segments are up to 60 microm in length and up to 10 microm in diameter. The visual cells of diurnal geckos have considerably smaller outer segments with lengths ranging from 6 to 12 microm and diameters ranging from 1.3 to 2.1 microm. Nocturnal and diurnal species differ in the structure of their ellipsoids. One type of visual cell in nocturnal geckos has modified mitochondria with either rudimentary cristae or no cristae at all, and one type of visual cell in diurnal geckos possesses an oil droplet. The visual cells of Phelsuma guentheri and Rhoptropus barnardi are intermediate between those of nocturnal and diurnal species. PMID- 11279364 TI - Distribution of ras guanyl releasing protein (RasGRP) mRNA in the adult rat central nervous system. AB - In the nervous system, Ras signal transduction pathways are involved in cellular differentiation, neuronal survival and synaptic plasticity. These pathways can be modulated by Ras guanyl nucleotide exchange factors (Ras GEFs), which activate Ras protein by catalyzing the exchange of GDP for GTP. RasGRP, a recently discovered Ras GEF is expressed in brain as well as in T cells. In addition to the catalytic domain which catalyzes dissociation of Ras-GDP, RasGRP has a pair of calcium-binding EF hands and a diacylglycerol binding domain. The structure of RasGRP suggests that it serves to link calcium and lipid messengers to Ras signaling pathways. We have used an RNase protection assay to detect RasGRP mRNA in various regions of the rat brain and we have determined the cellular distribution of RasGRP mRNA by in situ hybridization. RasGRP mRNA is widely distributed and is present in both interneurons and projection neurons but not confined to any neuronal type or neurotransmitter phenotype. The presence of RasGRP mRNA in archicortical neurons suggests that this pathway may be important in phylogenetically older regions of the CNS. The restriction of RasGRP mRNA to subsets of neurons suggests that activation of Ras by RasGRP has a specific function in certain neuronal types. We did not detect RasGRP in glial cells. PMID- 11279365 TI - Ipsilateral, ventral corticospinal tract of the adult rat: ultrastructure, myelination and synaptic connections. AB - The corticospinal tract (CST) of the rat is a widely used model system in developmental, physiological, and regeneration studies. The CST of the rat consists of a main tract, that runs in the dorsomedial funiculus and several minor components. We have shown earlier that one of the minor components, the ipsilateral, ventral CST, projects all the way down the spinal cord in the adult rat and single fibers form large terminal arbors in their spinal target areas. Here we investigated its ultrastructure and compared it to that of CST fibers of the main tract. By the use of anterograde axonal tracing with biotin dextran amine (BDA) and pre-embedding avidin-peroxidase histochemistry we investigated axon diameters and myelination using electron microscopy. Ipsilateral, ventral CST fibers were found to run in the ventral funiculus close to the midline. They were intermingled with heavily myelinated large diameter axons, presumably reticulospinal, vestibulospinal, or tectospinal fibers. Ipsilateral, ventral CST fibers were of small diameter (0.68 microm, +/-0.04) and about [frac34] of them were moderately myelinated (9.64 +/- 0.7 layers of myelin). Co-localization of a rhodamine-dextrane anterograde tracer with the presynaptic marker synaptophysin using confocal microscopy and electron microscopy revealed varicosities on terminal arborisations to be presynaptic boutons and clearly demonstrated contacts to neurons in intermediate laminae of the spinal cord at lumbar spinal levels. This study extends our earlier work indicating that the ipsilateral, ventral CST component of the adult rat is a morphologically complete CST component and may perform similar functions to the main CST component. PMID- 11279366 TI - Ionotropic glutamate receptor subunits are differentially regulated in the motoneuronal pools of the rat hypoglossal nucleus in response to axotomy. AB - Unilateral hypoglossal nerve axotomy was used as a model to analyse immunohistochemically the expression of the GluR1, GluR2, GluR3, and GluR4 glutamate receptor subunits of the alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionate (AMPA) subtype and the NR1 subunit of the N-methyl-D aspartate (NMDA) subtype in the different morphofunctional hypoglossal pools from 1 to 45 days postaxotomy. Following hypoglossal nerve axotomy, the percentage of motoneurons that were GluR1-immunopositive and the labeling intensity for this subunit was increased in some hypoglossal pools. Immunolabeling for the GluR2 subunit was undetectable. These results contrast with the unchanged pattern for these two subunits after sciatic nerve axotomy previously described. Image analysis showed a significant decrease in the intensity of immunohistochemical labeling for the GluR2/3 and GluR4 subunits in motoneurons, although most motoneurons were still immunopositive for these 2 subunits after axotomy. The intensity of immunolabeling for the NR1 subunit was slightly decreased postlesion, whereas the percentage of NR1-immunopositive motoneurons increased. Immunoreactivity returned to basal levels 45 days postlesion. These findings show that in axotomized hypoglossal motoneurons, i) AMPA and NMDA receptor subunits are still expressed, ii) the composition of the ionotropic glutamate receptor subunit pool is subjected to continuous changes during the regeneration process, iii) AMPA receptors, if functional, would have physiological properties different to those in intact motoneurons, and iv) the various AMPA receptor subunits are differentially regulated. The present results also suggest a faster recovery of basal levels of immunoreactivity for caudally localised groups of motoneurons which could reflect a caudo-rostral sequential functional recovery in the hypoglossal nucleus. PMID- 11279368 TI - UPCOMING SOUNDINGS ARTICLES. PMID- 11279369 TI - SURVEY ALERT. PMID- 11279367 TI - Response of spiral ganglion neurones to cochlear hair cell destruction in the guinea pig. AB - Loss of ganglion cells after hair cell destruction in the mammalian cochlea continues to occur over a long period of time, with the possibility of more than one factor contributing to this process. Despite the absence of hair cells, some ganglion cells are, however, able to survive for considerable periods of time. Because functional ganglion cells are crucial to the successful use of cochlear implants, a better understanding of the response of these cells to injury is required so that their loss can be prevented or ameliorated. Quantitative light microscopy, electron microscopy and immunocytochemical techniques were used to examine the response of type I spiral ganglion neurones to hair cell destruction, in guinea pigs at 1, 3, 6 and 30 weeks survival following intracochlear injection with gentamicin. The time course of ganglion cell loss was determined, while a closer examination of those cells able to survive was carried out. A significant early loss of large numbers of ganglion cells was followed by a further significant loss of these cells by 30-week survival. At the same time a decrease in the numbers of central fibres was also observed. Surviving ganglion cells have little or no perikaryal myelin, an appearance resembling that of type I ganglion cells at 55 days gestation. Ganglion cells surviving the initial loss were also found to have a significantly larger soma size than controls although this was not maintained and at 30 weeks survival the few remaining cells were similar in size to that of controls. The growth associated protein GAP 43 was upregulated in surviving ganglion cells at 3 weeks survival, but appeared diminished by 6 weeks survival. These features may indicate a survival response in ganglion cells and may provide a basis on which to develop appropriate means to prevent their loss. PMID- 11279370 TI - [Project for the development of the SIAARTI site (www.siaarti.org)]. PMID- 11279372 TI - [Epidemiology, costs and ethics in intensive care departments]. PMID- 11279371 TI - [Minerva anestesiologica and impact factor]. PMID- 11279373 TI - From infection to sepsis and organ(s) dysfunction. The surgical point of view. PMID- 11279375 TI - [Latex allergy in pediatric age: an interdisciplinary perioperative management and case reports]. AB - BACKGROUND: Latex intraoperative allergy is more and more frequent, especially in at groups risk (patients with spina bifida or congenital genitourinary abnormalities, pluri-operated patients, atopic subjects) and in pediatric age. The main problem of this allergy consists in the necessity of a strict collaboration of many specialists, in order to identify and safeguard the patient. METHODS: Our experience has pointed out an interdisciplinary perioperative management able to: 1. identify patients affected by latex allergy; 2. submit them to a latex-safe perioperative proceeding; 3. check their conditions with periodical tests. Selecting patients through a history and a list of questions, identifying profiles of typical risk patients, organizing the operating room with latex-free materials and equipment were the most important issues. Since November 1997 to December 1999 eighteen latex-safe perioperative proceedings have been carried out on 8 subjects (2 with esophageal atresia, 4 with bladder exstrophy and 2 with cloacal exstrophy); 2 of them were emergency cases. RESULTS: No allergic reactions and no proceeding-linked complications have been registered. Operators have always been satisfied by materials and equipment. Anesthesiological and surgical times resulted equal to those without latex-safe management. CONCLUSIONS: This perioperative management of potential or verified latex allergic patients turned out to be valid, safe and easy in practical application. PMID- 11279374 TI - [Adiuvants in the axillary brachial plexus blockade. Comparison between clonidine, sufentanil and tramadol]. AB - BACKGROUND: Evaluated effects of tramadol used as adiuvant in brachial plexus block and compared with clonidine and sufentanil. METHODS: Randomized, prospectic study with 80 patients, ASA status I-II, undergoing carpal tunnel release performed under axillary plexus block with ropivacaine 0.75% 20 ml divided in 4 study groups: tramadolo 100 mg (Group T), clonidina 1.5 g/kg (Group C), sufentanil 20 g (Group S) in 5 ml. of NaCl. Control (Group F) NaCl 5 ml. Adeguacy of the block was evaluated using pinprick test ( three points scale) and a temperature test. Onset time, duration of analgesia and anesthesia were recorded. Also during the surgery the sedation score on a five-point scale was evaluated and were recorded episodes of hypotension, bradycardia, Sp02<90% and other side effects. Results are reported as median+/-SD. For statistical analysis ANOVA test, Bonferroni test and c2 test were used. RESULTS: Onset time of anesthesia showed significant difference between three study groups than control group, while no significant differences was recordered between groups S, C and T (S: 11+/-7 min; C: 12+/-4 min; T: 14+/-8 min; F: 20+/-11 min.). Same results were obtained among duration of anesthesia and analgesia, that were lower in F group. An adeguate quality of block for surgery was obtained in 79 patients. Only one patients of F group needed surgical infiltration. A significant difference was recordered among quality of anesthesia between group F and other three study groups. Highest sedation score was noted in C and S groups. Significantly highest incidence of bradycardia and hypotension episodes were observed in group C. CONCLUSIONS: The use of tramadol as adiuvant provides a significative redution of onset time. Also provides a prolongation of anesthesia and analgesia with a quality of block similar that obtained with clonidine and sufentanil and a lower incidence of side effects of clonidine (sedation, bradycardia and hypotension) and sufentanil(itch and sedation). We conclude that tramadol may be a useful alternative, as adiuvant in periferic block, with same effects of other drugs commonly used and a lower incidence of side effects. PMID- 11279376 TI - [Cost of ICU in Italy. Results from an empirical study on a sample of 12 hospitals]. AB - BACKGROUND: The Italian hospital payment system based on DRG doesn t properly include Intensive Care Units (ICU) costs. Since great emphasis has been recently given to rationing health care resources, assessing ICU costs seems to be dramatically relevant. Aim of the study was to assess the average yearly cost and the cost per diem of a sample of Italian multispecialistic ICU wards. METHODS: In September 1995, a questionnaire concerning data on variable and fixed cost was sent to 25 Italian ICU wards, 11 NHS hospital-based (Northern Italy: 5; Central Italy: 4; Southern Italy: 2) and 14 school of medicine-based (Northern Italy: 7; Central Italy: 5; Southern Italy: 2). Variable cost data included: disposable, drugs, blood and blood-derived products, physical tests, chemical and microbiological routines, instrumental diagnostic procedures and physiotherapy. Concerning fixed costs, data on personnel and equipment were requested. In addition, some hospital overheads data (utilities; power; heating; maintenance; cleaning; laundry; accounting; waste disposal; cafeteria) were collected. RESULTS: On the basis of the 12 questionnaires returned (Northern Italy: 9; Central Italy: 3; Southern Italy: 0), the yearly cost of an ICU ward is Liras 4,580,032,000 (range 2,739,277,000-7,704,292,000), whereas the average cost per diem is Liras 1,802,000 (range 1,234,000-3,179,000). Cost of personnel is about 61% of the above mentioned costs. CONCLUSIONS: Despite the lack of questionnaires from Southern Italy and the unavailability of some data concerning both the cost of equipment and the overheads, the remarkable average cost values obtained could support further research. PMID- 11279377 TI - [Epidemiologic analysis of patients afferent to the ICU]. AB - BACKGROUND: To understand in- and out-patients flow to and from an ICU during a year (1998). The setting of the study was an 8-beds Intensive Care Unit of a 480 beds General Hospital with an Emergency Department. METHODS: Retrospective analysis by a specific designed software of all patient data extrapolated from the hospital database, in order to: 1) Divide all ICU patients in four groups, according to the first admission Department; 2) Classify all ICU patients into 3 subgroups: a) medical; b) surgical; c) trauma; 3) Evaluate the different needs of ICU resources in these different patient populations. RESULTS: Two hundred and fifty-four patients were admitted to our ICU during the study period (1.2% of all admissions). The mean duration of ICU stay was 10.4 days. Thirty-five per cent of ICU admissions came from the Emergency Department, 61% of ICU patients were discharged to another hospital ward, while the remaining 7% had to be transferred to a different hospital; 2.8% of our patients had ICU re-admissions. The overall mortality rate was 32%. CONCLUSIONS: Compared with previously reported data, a lower re-admission rate (3%), a longer mean stay in the ICU (>10 days) and a higher occupancy rate (91.4%) were observed. These data suggest that a large part of the available resources for the intensive care in our hospital are devoted to the in-hospital patient care. The hypothesis is suggested that this could be mainly due to the lack of sub-critical care areas. PMID- 11279378 TI - Deaths with acute cerebral lesion and heart-beating potential organ donors in the Veneto region. AB - BACKGROUND: The study was aimed at describing the clinical characteristics of dead patients with acute cerebral lesion and analyzing reasons of the shortage of heart-beating potential organ donors in the Intensive Care Units (ICUs) in the Veneto Region. METHODS: Data have been prospectively recorded in 23 ICUs over six months for deceased patients with acute cerebral lesion (clinical data, death diagnosis) and for any potential organ donor (medical suitability, family interview, organ retrieval). RESULTS: In the ICUs of the Veneto Region in 1998 deceased patients with acute cerebral lesion were 187 per million population (p.m.p.); 317 cases have been studied. Median age was 64 years (range 7-93). Heart-beating death was legally confirmed only in 98/317 cases (31%) against a clinical diagnosis of brain death in 203/317 (64%). Only 82/317 (26%) were considered eligible donors and 48/317 (15%) became real donors (22.8 p.m.p.). Among the remaining 235 cadavers, 105 were over 70 years old. In the group of 130 under 70 years absolute contraindications were present only in 30 and problematical clinical situations were reported in 100. CONCLUSIONS: The number of deaths with acute cerebral lesion represents a sensible index and a key factor for evaluating the potential organ donor pool in small regions and in the single intensive care unit. Collected data demonstrate that in the Veneto Region the efficiency of solid organ retrieval can be improved and that organ donor shortage may depend, beyond family refusal, on clinical and cultural factors that hamper stabilized heart-beating deaths. Most potential donors with age over 70 or problematical clinical situations are preventively excluded by ICUs physicians. To improve organ donation all the patients who die in spite of neuro-intensive treatment should be prevented from circulatory arrest to permit legal declaration of death. Thus more potential organ donors without absolute contraindications could be recovered and time would exist for discussing any problematical situation with experts in organ procurement, particularly in respect to existing urgencies in the waiting list. PMID- 11279379 TI - Norepinephrine can be useful for the treatment of right ventricular failure combined with acute pulmonary hypertension and systemic hypotension. A case report. AB - A 48-year-old woman who underwent emergency cardiac surgery for removal of a thrombus partially occluding the mitral valve, developed pulmonary hypertension right ventricular failure and systemic hypotension, in the immediate postoperative period, a clinical condition not well controlled by high doses of epinephrine and dobutamine. The addition of a continuous infusion of norepinephrine in incremental dosages, caused the rise in cardiac index accompanied before by the reduction in the pulmonary pressure and the stability in the systemic pressure, than by the further reduction in the pulmonary pressure and the increase in the systemic pressure. The conclusion is drawn that norepinephrine is useful in the treatment of right ventricular failure which follows a condition of acute pulmonary hypertension, because the improvement of cardiac performance established without adverse effects on the pulmonary pressures whose values on the contrary progressively declined. PMID- 11279380 TI - ["Talking point" replaces "final page"]. PMID- 11279381 TI - [Atrial fibrillation and thromboembolic events prevention. State of the art]. AB - Atrial Fibrillation (AF) is a common cardiac arrhythmia and stroke is its most devasting complication. The rate of ischemic stroke among people with AF is approximately six times that of people without AF and varies importantely with coexistent cardiovascular diseases; therefore stratification of AF patients into those at high and low risk of thromboembolism has become a crucial determinant of optimal antithrombotic prophylaxis. Multivaria-te analyses of prospective studies consistently show prior TIA/stroke, diabetes, age, heart failure to be independently predictive of stroke; left ventricular dysfunction is also strongly associated with stroke risk. Several randomized clinical trials demonstrated that treatment with adjusted-dose warfarin reduces the risk of stroke in AF patients by about two thirds. The efficacy of aspirin for prevention of stroke is controversial, but supported by pooled results of 3 placebo-controlled trials yelding a 21% reduction in stroke. The inherent risk of stroke should be considered in selection of AF patients for lifelong anticoagulation. Patients with AF and a recent stroke or TIA or multiple risk factors for stroke are likely to benefit from anticoagulation therapy; at present a target INR 2,5 appears optimal for most patients, although INR closer to 2.0 may be safer for patients at increased risk for bleeding events. The addition of aspirin to low- dose warfarin regimen does not provide any significant benefits and should be avoided. Therapy with aspirin is appropriate for patients who are at low risk of stroke or are unable to receive anticoagulants. AF patients treated with aspirin, should be periodically evaluated for development of high-risk features favoring anticoagulation. PMID- 11279382 TI - Mid-term results of endovascular repair for abdominal aortic aneurysm, with loco regional anesthesia, in high-risk patients. AB - BACKGROUND: Aim of this study was to evaluate the results of endovascular repair of abdominal aortic aneurysm (AAA) in patients considered not suitable for traditional open surgical repair because of the high anesthesiological risk. METHODS: We have retrospectively evaluated the result of the endovascular treatment of 11 patients with AAA of more than 6 cm diameter and high surgical risk due to cardiac hypokinesia and/or respiratory insufficiency. Patients were selected by a team composed of vascular surgeons and vascular radiologists who decided to implant the graft according to anatomical features of the AAA and of the iliac arteries. The treatment was performed in loco-regional anesthesia. The main end-points were: implantation success, mortality, morbidity, the absence of endoleak during the follow up that lasted two years. RESULTS: All the grafts were successfully implanted. There were no complications caused by anesthesiological manouvres. We had a minor intra-operative vascular complication and we performed three adjunctive endovascular procedures. A patient died of acute myocardial infarction, in the post operative period. Mean stay was six days. Pre-discharge scan showed 3 endoleaks (type I), two of these healed spontaneously and one sealed by percutaneous endovascular treatment. During follow-up (3-24 months) no patient died. One endoleak (type II) still persists. CONCLUSIONS The use of loco regional anaesthesia allows us to treat high risk patients. Following strictly the criteria of patient selection, the surgical high risk seems not to influence significantly the mid term results that are almost equal to the ones obtained in low-risk patients. PMID- 11279383 TI - [Doppler tissue imaging in myocardial infarction]. AB - BACKGROUND: The Doppler Tissue Imaging (DTI) is a recent Doppler method that allows to measure the velocities of myocardial walls. Thus, DTI may analyse myocardial contraction and give a quantitative evaluation of systolic and diastolic function. The aim of the study is to appraise the myocardial contraction of the left ventricle in patients with a recent myocardial infarction (MI) comparing data of standard echocardiography with those of DTI. METHODS: Fifteen patients with recent uncomplicated MI (22+/-6 days from the study) and 10 normal subjects have been studied. All population studied underwent bidimensional echocardiography with DTI analysis of different myocardial segments. RESULTS: In the infarcted patients, myocardial velocities were significantly reduced in comparison with the normal subjects in systole and in diastole. In patients with MI the picks of systolic velocities of normokinetic segments were significantly higher than those of akinetic/diskinetic segments (p<0.05). CONCLUSION: In myocardial infarction, the contraction of left ventricle is altered and it can be analysed and quantified through of the new indexes of systolic and diastolic myocardial function furnished by the DTI. PMID- 11279384 TI - Surgical treatment of aorto-enteric fistulas. AB - BACKGROUND: Our series of secondary aorto prosthetic fistulas (PEF) to identify if and how different surgical treatment affect outcome is reviewed. METHODS: Between 1982 and December 1999, in the authors department, 42 patients were investigated for a secondary PEF. Mean age was 65 years: the mean time interval since the primitive aortic procedure was 49 months. Twenty patients were treated in emergency surgery: 29 presented evidence of gastrointestinal bleeding. The preoperative work-up included esophagogastroduodenoscopy, CT scan, and aortography. The vast majority of PEF were in a duodenal location. Surgical procedure carried out was graft excision, bowel suture or bowel resection, aortic stump closure and axillofemoral (AXF) bypass (11), new in situ revascularization by synthetic prosthesis (5), simple suture (9), graft excision without revascularization (1), in situ revascularization using arterial homograft (13). RESULTS: The mean surgery duration was 4 hours and 53 minutes, the mean blood loss was 1845+/-1132. Two patients died shortly after proximal aortic control was obtained. Early overall mortality was 50%, the early overall bypass occlusion rate was 12.5%, the early overall amputation rate was 10%, and the early new PEF rate was 12.5%. Late overall mortality was 22.5%, the late overall bypass occlusion rate was 20%, the late overall amputation rate was 7.5%, and the late new PEF rate was 10%. CONCLUSIONS: Bleeding of the gastrointestinal tract in patients with a history of intra-abdominal reconstructive vascular surgery must raise severe suspicion as to the certainty of existence of a PEF unless the diagnostic procedure excludes this possibility. All treatment methods resulted in catastrophic failure, related to recurrent PEF or septic complication. Perhaps, in the presence of PEF extra-anatomical bypass associated with aortic ligature remains an interesting surgical solution. PMID- 11279385 TI - [Ventricular pre-excitation: electrophysiopathology, criteria for interpretation and clinical diagnosis. References for geriatrics]. AB - The authors review the state-of-the-art on ventricular pre-excitation in medical and arrhythmological literature in order to facilitate the recognition of the various clinical forms, like classic and occult Wolff Parkinson withe syndrome and Lown Ganong Levine syndrome. A historical introduction reviews our electrophysiopathological knowledge of the electrical activation and conduction of ventricular pre-excitation compared to normal, starting from the anatomic discovery of conduction pathways to the possible use of transesophageal electrostimulation and endocavity mapping to study electric potentials. Avantgarde technologies have also been developed to eliminate anomalous pathways firstly by using a direct current dirscharge and secondly radiofrequency. Atrioventricular electric activation has been widely illustrated in normal subjects in order to create a model for comparison with pathological ventricular pre-excitation: the upper left portion of the septum is no longer the first zone to trigger the kinetic mechanism compared to the early fascicular fraying of the homonymous branch. Instead the upper right part of the septum is activated earlier owing to the anomalous fascia connected on this side to the right branch through their septal arborisations. As a result, this new conduction pathway activates the ventricular masses earlier through an anomalous route, given that there is no further contact with the atrioventricular nodes which act as a control. A similar situation is found in the left branch block where the ventriculogram is late with a normal PR, unlike pre-excitation when an early delta wave is present with a short PR. Electric conduction is also illustrated based on new knowledge of the circuit structures and the rings theory. Orthodromic tachycardia is distinguished from the antidromic form, double accessory pathway tachycardia, ectopic reciprocant atrial fibrillation tachycardia and occult bundle tachycardia which is studied using transesophageal stimulation with a time threshold of 70 ms for ventricular-atrial retrograde activation. The stimulation techniques using single or repeated extrastimulus are explained for this purpose, as well as those with serial extrastimulation and the physical characteristics of the circuit based on the ratio between voltage and resistance. The authors also report the practical aims of electrostimulation for determining the electric threshold of the anomalous circuit in terms of refractoriness, electric atrial stability, reciprocity and the occurrence of the macro re-entry. Lastly, the authors describe electric conduction by anomalous pathways based on the criterion of conduction and activation, both of which are analysed and compared on the basis of the intrinsicoid deflection morphology on the surface ECG: the aberrant qRs usually suggests an antidromic ventricular activation and retrograde conduction between atrium and ventricle, while normal intrinsicoid deflection demonstrates that the activation is orthodromic and the conduction anterograde, namely ventricle-atrial. Having been reproduced in a synoptic synthesis, these manifestations show a regular electrophysiological pattern because they are dissimilar from the behaviour of the monophasic bioelectric potential of the cardiac cells specialised in the conduction of the stimulus, whether they represent a normal or pathological electric pathway. The study is rounded off by the analysis of the reciprocant tachycardias and their re entry varieties related to the type of the anomalous bundles. A number of types of re-entry can be identified: sinusal re-entry (micro re-entry), atrial re entry, re-entry in the atrio-ventricular node, re-entry tachycardia and the so called triggered type. The discussion of the electrophysiopathological aspects of pre-excitation is followed by the clinical forms of ventricular pre-excitation that can be divided into 3 main types. The different ECG clinical pictures are set out in the summary table, together with the type of shunt and activation and possible variants, following Rosenbaum s classic paint: the common type B, the rare type A and a last variant, the C type. This section also describes the positional peculiarities of the Kent-Paladino bundle, the left ventricular, septal (anterior and posterior) and the multiple-bundle ones. The authors also illustrate the criterion and meaning of endocavity mapping in the search for anomalous bioelectric potentials that identify the pathway or the location of the endocardiac bundle and/or foci to be eliminated. A new echocardiographic technique is described with a conventional M mode, digitalised 2D and tissular Doppler which has a comparable ability to identify the anomalous pathways of electric conduction using a non-invasive method. (ABSTRACT TRUNCATED) PMID- 11279386 TI - Quadricuspid aortic valve, parossistyc supraventricular tachycardia and double right kidney: an uncommon association. AB - Fourteen years after surgery for replacement of the aortic valve, an interesting case previously unreported was brought to our attention. The female patient came to our OP Dept for a routine follow-up: she had been found at surgery to have a quadricuspid aortic valve. Operation dated October 1985. At a careful appraisal of the world-wide literature, we noticed that such an association of pathologies had never been reported before: quadricuspid aortic valve, paroxystical supraventricular tachycardia and right double kidney with double renal pelvis and double proximal ureter. Other anomalies associated with the quadricuspid valve, available in the literature are: patent duct, subvalvular fixed aortic stenosis, ventricular septal defect, hypoplastic anterior mitral leaflet and pulmonary stenosis. The pathologic findings at autopsy of this congenital malformation vary between 0.008% and 0.033%; attention must be turned to the fact that the incidence can be underestimated if not expressly searched for. The first quadricuspid aortic valve was described in 1862 by Balington in an autoptic report and sixty other cases have been reported since. In rare cases this pathology has been diagnosed at angiography. At follow up our patient remains in SR with rare transitory episodes of supraventricular tachycardia. PMID- 11279387 TI - Persistent ST segment elevation in a patient with metastatic involvement of the heart. AB - Electrocardiographic abnormalities are commonly seen with tumor invasion of the heart, but usually these abnormalities are not specific. Pronounced and prolonged lateral ST segment elevation in the absence of myocardial infarction occurred in a patient with epidermoid carcinoma of the left lung. Computer tomography showed the presence of tumor invasion of the heart. Prolonged ST segment elevation in the absence of Q waves seems to be a pathognomonic sign for tumor invasion of the heart. PMID- 11279388 TI - Mycotic aneurysm of the palmar artery associated with infective endocarditis. Case report and review of the literature. AB - A 26-year-old man was diagnosed with mycotic aneurysm of the left hand associated with active infective endocarditis. Preoperative arteriography of the hand revealed aneurysm of the radial side of the deep arch of the palmar artery. We approached the aneurysm from the dorsal side of the hand in order to avoid damage to the collateral vascular supply of the superficial arch of the palmar artery and neurological structures. As a result, the aneurysm was excised simply by proximal and distal ligation of the vessel. During follow-up over 14 months, no evidence of recurrent aneurysm formation or ischemia of the fingers has been obtained. PMID- 11279389 TI - Left sub costal minilaparotomy in aortic surgery. AB - Aim of this work is to present our surgical technique, i.e. a left sub costal transperitoneal minilaparotomy, used in 40 patients operated on in the last year for atherosclerotic aorto-iliac occlusive disease (aortofemoral bypass) and aortic or aorto-iliac aneurysm (aorto-aortic graft or aorto-iliac bifurcated graft sutured on the common iliac arteries). The patients are placed in a dorsal decubitus. The cutaneous incision of 10 to 15 cm, depending on the abdominal size, is parallel to the condro-costal edge and spreads from the linea alba to the edge of the rectus muscle. The linea alba is usually incised; the oblique and the transverse muscles are not touched. The bowel is maintained within the abdominal cavity. Usually we do not use self-retaining retractors. The abdominal wall and the bowel are retracted with moistened towels maintained by blade intestinal retractors. When the abdominal cavity is gained, conventional dissection of the aorta and iliac arteries is carried out. These manoeuvres and the following surgical procedure are performed as usually with standard vascular instruments. Nasogastric suction and drains are not used routinely. In our series, this minilaparotomy technique, joined to <>, and to an intensive postoperative training, allows a better outcome of the patient and a discharge home from 3rd to 5th postoperative day. So we think that this technique, not so expensive as endovascular repair or laparoscopic and video assisted surgery, nevertheless retains all the proven benefits of a minimally invasive surgery. PMID- 11279390 TI - [Sonographic characterization, Doppler ultrasonography and tumor markers in the diagnosis of malignancy of ovarian masses]. AB - BACKGROUND: The study analyses the diagnostic possibilities regarding ovarian neoplasms offered by different clinical approaches: B-mode morphological ultrasonographic examination, colour Doppler and Doppler pulsed ultrasonography, and lastly the assay of a number of tumour markers. METHODS: A prospective study was carried out in 125 selected patients attending the Ultrasonography unit of the Obstetrics and Gynecology Clinic at Parma University between June 1997 and June 1999 who presented an adnexal mass . All patients underwent transvaginal ultrasonography (multifrequency vaginal probe 5.0-6.5 MHz, Esaote Idea, Genova) to characterise the mass, applying 5 different ultrasonographic scores: Granberg, Sassone, Di Priest, Lerner, Ferrazzi. Colour Doppler imaging was then performed to analyse the vascularisation of the mass, also using pulsed Doppler to study a number of velocimetric parameters: pulsatility index, index of resistance, systolic and diastolic peak velocity, mean velocity. All the patients underwent surgery using laparotomy or video laparoscopy, accompanied by histological analysis. A number of different tumour markers were assayed prior to surgery: Cal25, CA19-9, CEA, beta-HCG, alpha-fetoprotein. RESULTS: Out of 127 pelvic masses examined, histological analysis showed that 19 were malignant and 108 benign. The diagnostic accuracy of malignancy was comparable for the 5 scores studied, with a minimum of 57.48% for Lerner and a maximum of 77.16% for Di Priest. The central importance of vascularisation was the only significant parameter among those analysed using colour Doppler which was useful for the diagnosis of a malignant neoplasm, with a diagnostic accuracy of 82.95%. No indicator obtained using pulsed Doppler was useful for diagnostic purposes. CA125 was the only tumour marker that revealed a statistically significant difference emerged between the benign (21.6 U/ml) and malignant (220.8 U/ml) masses. Its diagnostic accuracy was 75.58%. CONCLUSIONS: This study confirmed that the three methods analysed do not differentiate substantially in their overall diagnostic capacity of malignant ovarian neoplasms. The best performances for ecographic scores (Di Priest) did not exceed a sensitivity of 89.47% with a 21.25% incidence of false positives; this was comparable to CA125 with a sensitivity of 85.71% and false positives in 22.09%. In relation to the central importance of vascularisation, colour Doppler achieved a lower sensitivity (55.55%), but this was confirmed by a low incidence of false positives (7.95%). This revealed its importance as a useful method, especially for excluding the presence of malignant tumours. PMID- 11279391 TI - [Intraoperative complications of 697 consecutive operative hysteroscopies]. AB - BACKGROUND: Complications due to hysteroscopy are relatively rare events. They occur more frequently with operative hysteroscopy than with diagnostic hysteroscopy. Exact complications rates are difficult to determine owing to the natural tendency to report successes but not complications. Recognition of these situations will lead to prevention; in fact, all the most serious complications of operative hysteroscopy can be avoided when proper precautions are taken and close communication is maintained among gynecologic surgeon, the anesthesiologist and nursing staff. The more clinically significant complications are: uterine perforation, haemorrhage and electrolyte imbalance. METHODS: Between January 1993 and December 1998, 697 women underwent operative hysteroscopy in our Department. Operative hysteroscopy was performed with continuous flow, high frequency resectoscope. Under general anesthesia the cervix was dilated to 10 mm and the uterine cavity was distended with 1.5% glycine solution or mannitol under 80 to 120 mmHg pressure. Resection with electrocoagulation was completed. The patients were submitted to the following procedures: 354 endometrial polypectomies (50.7%), 160 myomectomies (23%), 114 endometrial ablations (16.4%) and 69 hysteroscopic metroplasties (9.9%). RESULTS: In our series complications occurred in 95 out of 697 patients (13.6%). The most important complications were: 12 (1.7%) uterine perforations, 48 (6.9%) intraoperative haemorrhages and 35 (5%) excessive hypotonic fluid absorptions. Four out of 12 perforations occurred during the dilation of the cervical channel. Since the distention of the uterine cavity could not be achieved, the procedures were stopped. No signs of vaginal or intraperitoneal haemorrhage were observed; 8 out of 12 perforations were due to the tip of the electrical source. The operative hysteroscopies were immediately stopped and the consequences were: 6 diagnostic laparoscopies, 1 laparotic hysterectomy (hemorrhage) and 1 laparotomy for thermal bowel injury. In 48 patients intraoperative bleeding could not be controlled with electrocautery. In these cases in the operating room a Foley catheter was inserted into the uterine cavity and the bulb inflated with 10 to 30 mL of liquid to tamponade the bleeding. The catheters were removed 12 to 24 hours later. No patients required blood transfusion. Excessive intravasation of electrolyte-free fluid occurred in 35 patients. Hyponatremia and hypokalemia (hypo-osmolarity result) were never serious. Headaches, nausea and vomiting were the most frequent symptoms of our patients. No cardiac arrhythmia, cerebral edema, brain herniation occurred. In our series, hemorrhage was the most common complication; intravasation and uterine perforation were at the second and the third place. Complications rates decreased progressively du to a better major training and experience of the surgeons. Also the curves of each complication show a significant decrease. Myomectomy in our hands has been the most dangerous procedure. However, serious sequelae were rare mainly for two reasons: we prefer stop the intervention rather than continue when a deficit of 1.000 mL is reached. Consequently, it is very important to discuss the possibility of incomplete resection of the endouterine lesion with the patient preoperatively; a protocol for fluid management in the operating room must be used for all the procedures (also the easiest) by all the surgeons and the nurses. CONCLUSIONS: Our relatively high prevalence of intraoperative complications and distribution of the different types do not differ from the findings of published reports. In personal experience operative hysteroscopy is a safe surgical procedure for the treatment of endouterine abnormalities. PMID- 11279392 TI - [Observations on vaginal delivery in women with previous cesarean section]. AB - BACKGROUND: The significant increase in cesarean sections both before (52.73%) and during labour (89.82%) observed in the 1990s compared to the period 1970-1980 prompted the authors to review the cases of women admitted to Department B of the Gynecology and Obstetrics Clinic at the University of Turin and to study the number of women with previous cesarean sections undergoing labour. The aim of this study was to throw light on this complex question and to reduce the incidence of surgical births wherever possible. METHODS: Two groups of pregnant women with previous cesarean sections were studied in Department B of the Gynecology and Obstetrics Clinic at the University of Turin: one group included women undergoing cesarean sections between 1990-1998, and the other included women undergoing cesarean sections between 1970-1980. The authors analysed the indications for repeat cesarean section and the percentage of vaginal births. RESULTS: The results show that during 1970-1980 the percentage of vaginal births was 24.34% (259 vaginal births out of 1593 patients), whereas between 1990-1998 the percentage of vaginal births fell to 10.18% (51 vaginal births out of 1060 patients). CONCLUSIONS: The high percentage of repeat cesarean sections found in the 1990s is not only due to strictly medical reasons, but also to ethical and political motives, and above all the maternal desire not to undergo natural labour. PMID- 11279393 TI - [Birth in water. A comparative study after 555 births in water]. AB - BACKGROUND: The object of our study is to research into the quality of the different delivery positions, offered in our hospital with special focus on the advantages for birth in water. METHODS: From February 1997 to 1 October 2000 we do research retrospectively on data of 555 deliveries in water, 320 on the traditional bed and 125 on the delivery stool give us the possibility to investigate about duration of birth, rate of episiotomies and perineum lacerations, consumption of painkillers, arterial umbilical cord pH and haemoglobin postpartum. RESULTS: In our comparing analysis of the duration of birth we could show a relevant reduction especially for primiparae which had delivered in water. The reduction is only significant for the first part of labor (360 minutes in the pool, 445 minutes on the traditional bed and 420 minutes on the stool) whereas there is no difference for the second part of labor. The significant reduction on episiotomies (1%) in comparison to the one on the traditional bed (20%) or on the stool (10%) for primiparae in water doesn t mean an increase at perineum lacerations. (each 25%). In water we saw no lacerations/injuries of the perineum for 58% of primiparae, on the traditional bed 36% and on the stool 43%. No woman in labour needed a painkiller in the pool. There was no difference found between the three groups referring to the arterial umbilical cord pH or the haemoglobin postpartum. CONCLUSIONS: Our study shows relevant medical advantages for a delivery in water: and a significant reduction of the duration of the first part of labour, significant less episiotomies and perineum lacerations and no need for painkillers. The security of the neonate is guaranteed under attention to the known contraindications. PMID- 11279394 TI - [Maternal morbidity in multiple pregnancies]. AB - BACKGROUND: This study aimed to outline the clinical physiognomy of maternal morbidity in multiple pregnancies in order to improve maternal and feto-neonatal. METHODS: We reviewed the admissions to Department B of the Gynecology and Obstetrics Clinic at Turin University during the decade 1989-1998. Out of 17,445 pregnancies, we noted a 205 multiple pregnancies (1.17%), including 199 sets of twins, 5 triplets and 1 quadruplets. The percentages for the incidence of the various forms of maternal morbidity were compared to a control group of 1000 single births. RESULTS: Of 205 multiple pregnancies, 169 (82.43%) presented complications of varying severity and associated with statistically significant increase (always over 50%). These took the form of: premature birth (75.12%), PROM (28.29%), threat of premature birth (14.63%), phlebectasia (9.75%), anemia (8.78%), hyperemesis in the first quarter (8.29%), abortion (4.89%), polyhydramnios (4.39%), urinary tract infection (1.95%), detachment of the placenta (1.95%), liver pathology (1.46%), placenta previa (0.97%). CONCLUSIONS: The pathological picture revealed by this survey may seriously jeopardise the normal evolution of pregnancy, with severe repercussions for mother and fetus neonate. These high-risk pregnancies should be included in a preventive programme of medical-social-outpatient and home assistance to guarantee early hospitalisation. PMID- 11279395 TI - [Induction of monofollicular cycles]. AB - The therapy of anovulatory infertility is not meant to obtain a pregnancy at any cost, but to restore an ovulation as physiological as possible. This involves the use of drugs and therapeutical protocols to obtain monofollicular cycles. Monofollicularity reduces the two main risks of induction of ovulation: ovarian hyperstimulation syndrome and multiple pregnancy. The aim of this study is a review of the Literature on ovulation induction and a comparison with the data of our Sterility Service. The importance of the question will be examined together with the most used ovulation induction drugs: clomiphene citrate, gonadotrophins and pulsatile GnRH. The parameters considered are: the number of follicles, single or multiple pregnancies and ovarian hyperstimulation. After a review about ovarian stimulation, the results of our Sterility Service are presented: 364 cycles of ovulation induction with clomiphene citrate, low-dose gonadotrophins or pulsatile GnRH were monitored; monofollicularity was obtained in 58,48% of ovulatory cycles. Differences between drugs will be described in the text. The therapy of anovulatory infertility aims to restore a physiological ovulation and to obtain a single pregnancy, not a pregnancy at any cost. PMID- 11279397 TI - [Controversial categories in cytopathology of the uterine cervix. II. AGUS: atypical glandular cells of undetermined significance]. AB - AGUS (Atypical Glandular Cells of Undetermined Significance), or AGCUS, is a category for reporting doubtful or suspicious glandular changes of the uterine cervix. Glandular lesions are not well known by the cytopathologist and their cytologic criteria are not completely reproducible. Only with the introduction of The Bethesda System (TBS) in 1988, the presence of endocervical cells is considered essential to correctly evaluate a cervical specimen. The origin of atypical glandular cells, endometrial or endocervical, should be distinguished. Moreover, endocervical AGUS should be further qualified as favor reactive or favor neoplastic or Adeno-carcinoma in situ (AIS) . Recently, it has been proposed to classify endocervical AGUS in a) AIS; and b) AGUS that cannot rule out AIS when incomplete criteria of AIS are present. Moreover, the origin of AGUS is sometimes impossible to know. In these cases, the diagnosis is AGUS not otherwise specified (NOS). The clinical management of AGUS presents different options depending on its origin or its further qualification: cytologic follow up, colposcopy and eventual biopsy, endocervical or endometrial curettage, hysteroscopy, human papillomavirus typing, etc. including conization and hysterectomy. In conclusion, an appropriate clinical management is needed to detect glandular or squamous lesions that can be frequently identified in the AGUS follow-up. PMID- 11279396 TI - [Controversial categories in cytopathology of the uterine cervix. I. ASCUS: atypical squamous cells of undetermined significance]. AB - ASCUS (Atypical Squamous Cells of Undetermined Significance) is a class for reporting cervical cytopathologic diagnoses. The Bethesda System (TBS) introduced ASCUS in 1988 and then defined this cytologic class further. Since the initial subclassification in probably reactive or probably neoplastic , TBS proposed different subgroups for a correct clinical management. At present, the subgroups are the following: ASCUS a) due to compromised specimen (poor processing or obscuring material); b) with mature intermediate-type cytoplasm; c) in postmenopausal women; d) atypical metaplasia; and e) with orangeophilic cytoplasm. Generally, clinical management of ASCUS presents 3 options: 1) cytologic follow-up (colposcopy only in a persistent diagnosis of ASCUS); 2) colposcopy; and 3) both Human papillomavirus testing and Pap-test. These options may be adopted in the ASCUS type b) and e) whereas in the type c) only in postmenopausal women receiving hormone replacement therapy. In the ASCUS type a), Pap-test should be immediately repeated (processing defects) or after therapy (excessive inflammation). In postmenopausal women not receiving hormone replacement therapy, the Pap-test has to be repeated after topical estrogen therapy. In ASCUS type d), a more aggressive follow-up is needed, such as colposcopy and eventual biopsy. Therefore, in the ASCUS diagnosis an effective communication between cytopathologist and clinician is needed for a correct clinical management. PMID- 11279398 TI - [Balneotherapy with arsenical-ferruginous water in chronic cervico-vaginitis. A case-control study]. AB - BACKGROUND: The aim of this study was to evaluate the efficacy and tolerability of vaginal irrigations with arsenical-ferruginose water from the spa at Terme di Levico in chronic cervico-vaginitis in order to assess the validity of balneotherapy in improving the symptoms and quality of life of patients. METHODS: An open comparative study was performed in 30 patients with symptoms attributable to aspecific chronic vaginitis (in child-bearing age) or vulvovaginal dystrophy (perimenopausal age). Twenty patients (Group A) received balneotherapy and 10 (Group B) were treated with placebo vaginal suppositories. All patients were required to fill in a questionnaire on symptoms. A thorough gynecological examination was performed, together with a Pap-test and vaginal secretion sample for bacterioscopic and microbiological tests. The same tests were repeated at the end of treatment. RESULTS: Post-treatment results showed a general reduction in the extent of gynecological symptoms reported by patients. A statistically significant reduction was only observed in patients receiving balneotherapy for the following symptoms: vaginal burning , vulvar burning , vaginal itch , vulvar itch , leukorrhea . An analysis of the cytological tests performed in patients enrolled in the study highlighted a high prevalence of phlogistic type findings, often in association with varying degrees of atrophy in postmenopausal patients. Those patients suffering from chronic vaginitis undergoing balneotherapy showed a reduction in the prevalence of phlogistic findings after treatment. CONCLUSIONS: Treatment with arsenical-ferruginose water led lo a marked reduction in the subjective symptoms reports by the study population (particular evident in patients with chronic leukorrhea), as was confirmed by objective signs (clinical, cytological and microbiological) of phlogosis. This was accompanied by excellent tolerability. These results justify the use of balneotherapy, according to the classic techniques and methods, in chronic cervicovaginal phlogistic processes. PMID- 11279399 TI - [New techniques in childbirth assistance. Experience and results of theoretical and practical training]. AB - In 1995, the Course on Integrated Obstetrical and Gynaecological Techniques was added to the training program of the Obstetrics and Gynaecology Clinic and to the Midwife Diploma School, at the Faculty of Medicine of the A. Avogadro University of East Piemonte. This addition was due to the demand to create a service to train young medical doctors and student midwives on the basis of the requirements of the World Health Organisation, concerning a more natural way of giving birth. In this paper the results obtained after a four years practical application of these clinical techniques are presented. The factors considered were the type of assistance offered in correlation with different outcomes for both mother and child. The study demonstrates a general improvement in the quality of assistance and a decrease of costs for the National Health Service. The data have been compared with those of the neighbour Hospital Division, where deliveries are assisted with traditional techniques. PMID- 11279400 TI - Skeletonized internal thoracic artery grafts and wound complications. PMID- 11279401 TI - Congestive heart failure: treat the disease, not the symptom--return to normalcy. PMID- 11279402 TI - Postoperative atrial fibrillation: an old problem crying for new solutions. PMID- 11279403 TI - Concomitant type I thyroplasty and thoracic operations for lung cancer: preventing respiratory complications associated with vagus or recurrent laryngeal nerve injury. AB - OBJECTIVES: We sought to prevent postoperative swallowing disorder, aspiration, and sputum retention in cases of recurrent laryngeal or vagus nerve section occurring during lung cancer resection. METHODS: In 14 of 25 consecutive patients, type I thyroplasty and thoracic operations were performed during the same period of anesthesia. All patients had a preoperative laryngeal computed tomographic scan providing us with indispensable measurements for vocal fold medialization under general anesthesia (ie, without intraoperative phonatory control). Nine remaining patients had a type I thyroplasty delayed from thoracic operations because of intraoperative doubt about laryngeal innervation injury, and 2 did not need a laryngeal operation. Main postoperative records consisted of swallowing ability, respiratory complications, and quality of voice. RESULTS: No swallowing disorder, aspiration, or sputum retention occurred in cases of concomitant laryngeal and thoracic operations. Of these 14 patients, a single case (7%) of major complication (vocal fold overmedialization) occurred and required an early and successful revision thyroplasty; one case of cervical hematoma that did not require surgical drainage was considered a minor complication (7%). Twelve (86%) patients who underwent the concomitant association of both operations were fully satisfied with their quality of voice. CONCLUSIONS: Type I thyroplasty and thoracic operation can be advantageously associated in case of injury to laryngeal motor innervation to prevent postoperative swallowing disability and dramatic respiratory complications. PMID- 11279404 TI - Chest wall invasion in non-small cell lung carcinoma: a rationale for en bloc resection. AB - OBJECTIVE: The choice of surgical approach to non-small cell lung cancer invading the chest wall, extrapleural resection versus en bloc chest wall resection, is much more related to the experience of the surgeon than to objective criteria. The aim of the present study is to help to establish a rationale for en bloc chest wall resection in lung cancer invading the chest wall. METHODS: From January 1990 to June 1999, of 1855 patients having major pulmonary resections for non-small cell lung carcinoma, 104 (5.6%) patients with neoplasms involving the chest wall underwent en bloc chest wall and lung resection plus radical mediastinal lymphadenectomy. RESULTS: All patients underwent complete resection with microscopically disease-free tissue margins. Depth of invasion was into the parietal pleura only in 28 (26.92%), into the pleura and soft tissue in 36 (34.62%), and into the pleura, soft tissue, and bone in 40 (38.46%). No operative mortality was reported. Follow-up was completed in 96 patients. One patient had a local recurrence. The overall 5-year estimated survival was 61.4%. Survival in the subsets T3 N0 and T3 N2 were, respectively, 67.3% and 17.9% (P =.007). The 5 year survival was 79.1% in involvement of parietal pleura only and 54.0% in involvement of soft tissue with or without bone invasion (P =.014). Five-year survival was 53.0% in adenocarcinoma versus 71.8% in squamous cell carcinoma (P =.329) and 74.1% in patients who did undergo radiation therapy versus 46.7% in patients who did not undergo radiation therapy (P =.023). CONCLUSIONS: En bloc resection of the chest wall and lung is the procedure of choice to obtain complete resection in lung carcinoma invading the chest wall. Survival is highly dependent on the completeness of resection, nodal involvement, and depth of chest wall invasion. PMID- 11279405 TI - Sequential thoracic metastasectomy prolongs survival by re-establishing local control within the chest. AB - OBJECTIVE: The value of sequential thoracic metastasectomies is unknown. We evaluate repeat metastasectomy for limited recurrences within the thorax. METHODS: From July 1988 to September 1998, 54 patients underwent 2 to 6 separate sequential procedures to excise metastases after recurrence isolated to the thorax. Kaplan-Meier survival and Cox modeling determined prognostic variables. RESULTS: Thirty-three men and 21 women, 22 to 76 years underwent 2 (100%, n = 54), 3 (50%), 4 (22%), or 5 to 6 (11%) metastasectomies. Fifty-four percent of patients had carcinoma, 35% sarcoma, 9% germ cell, and 2% melanoma. There were no operative deaths; all late deaths occurred from cancer. Median follow-up was 48 months. Cumulative 5-year survival from the second procedure was 57%. After the second, third, fourth, and fifth procedures, respectively, permanent control was achieved in 15 (27%) of 54 patients, 5 (19%) of 27, 1 (8%) of 12, and 0 of 7. Recurrence amenable to additional surgery occurred in 27 (50%) of 54, 12 (44%) of 27, 6 (50%) of 12, and 1 (17%) of 6. Mean hazard for the development of unresectable recurrence increased from 0.21 after the second procedure to 0.91 after the fifth procedure. The 5-year survival for the 27 patients undergoing only 2 metastasectomies was 60% (median not yet reached), 33% for the 15 patients undergoing only 3 metastasectomies (median 34.7 months), and 38% for the 12 patients undergoing 4 or more (median 45.6 months). From the time a recurrence was declared unresectable, patients had a 19% 2-year survival (median 8 months). CONCLUSIONS: Multiple attempts to re-establish intrathoracic control of metastatic disease is justified in carefully selected patients, but the magnitude of benefit decays with each subsequent attempt. PMID- 11279406 TI - Bilateral skeletonized internal thoracic artery grafts in patients with diabetes mellitus. AB - OBJECTIVE: Increased risk of deep sternal infections has prohibited routine bilateral internal thoracic artery grafting in diabetic patients. The technique for harvesting the skeletonized internal thoracic artery provides the potential to minimize this risk. The purpose of this study was to compare the outcome of bypass grafting with bilateral skeletonized internal thoracic arteries in diabetic and nondiabetic patients. METHODS: From May 1996 to April 1998, 231 consecutive diabetic and 534 nondiabetic patients underwent bilateral skeletonized internal thoracic artery grafting. Mean age was 66 years. Compared with the nondiabetic group, the diabetic group comprised more women (29% vs 18%, P =.001), had a greater prevalence of hypertension (53% vs 44%, P =.019) and congestive heart failure (20% vs 14%, P =.016), but a lower prevalence of preoperative acute myocardial infarction (26% vs 34%, P =.027). RESULTS: Operative mortality of diabetic patients was comparable with that of nondiabetic patients (3% vs 2.6%). The two groups also had similar occurrences of deep sternal infection (2.6% vs 1.7%, respectively, P =.40). Deep sternal infection was significantly more prevalent in obese, diabetic women (3/20 = 15%) than in diabetic patients without this combination of risk factors (3/211 = 1.4%, P <.0001) (odds ratio 11.1, confidence interval 2.1-59.4). Diabetic patients also had a higher incidence of stroke (3.5% vs 0.9%, P =.014). Three-year actuarial survival of diabetic patients was lower (91.3% vs 94.7%, P =.083). CONCLUSIONS: Bilateral skeletonized internal thoracic artery grafting is a good surgical revascularization option in diabetic patients. Operative mortality and prevalence of sternal infection are comparable with those of nondiabetic patients. However, the risk of sternal infection in obese diabetic women is high, and for them we advocate the use of a single artery instead of bilateral internal thoracic arteries. PMID- 11279407 TI - Repair of dyskinetic or akinetic left ventricular aneurysm: results obtained with a modified linear closure. AB - OBJECTIVE: In patients with a dyskinetic or akinetic area of the left ventricle, controversy exists over who will benefit from resection. This study evaluates results achieved with a modified linear closure in 193 of 196 consecutive cases. Preoperative cases (n = 160 [83%]) were in functional class III or IV with congestive heart failure (n = 115 [60%]), angina (n = 108 [56%]), and syncope (n = 67 [35%]). The ejection fraction was 25% +/- 8%, and echocardiography showed significant mitral regurgitation in 86 (45%) patients. In patients with detailed wall motion analysis, 50 (57%) were akinetic, and 37 (43%) were dyskinetic. METHODS: Repair was completed on the beating heart to minimize ischemia and allow assessment of wall function and viability to guide resection and repair. Additional procedures included coronary artery bypass grafting (n = 175 [91%]), septoplasty (n = 24 [12%]), and arrhythmia ablation (n = 77 [40%]). Ventricular and mitral valve function were assessed by means of preoperative and/or postoperative gated acquisition scans in 171 (90%) patients and Doppler echocardiograms in 170 (88%) patients. RESULTS: Hospital mortality was low (5/193 [2.6%]), although 34 (18%) patients needed perioperative intra-aortic balloon pump support. Actuarial survival at 1 and 5 years was 91% and 84%. Most late deaths were due to congestive heart failure. Seven patients required transplantation (interval, 36 +/- 32 months). As determined by multivariable analysis, factors predicting poor outcome at 5 years were preoperative mitral regurgitation of 2+ or greater, congestive heart failure, and ventricular tachycardia. Among survivors, 126 (80%) of 157 were in functional class I or II, and the average increase in ejection fraction postoperatively was 9.1% +/- 10.0%. Postoperative echocardiograms in 70 patients with significant mitral regurgitation preoperatively showed improved valve function in 40 (57%) of 70 patients. CONCLUSIONS: We conclude that repair of dyskinetic or akinetic aneurysms by means of a modified linear closure plus septoplasty in selected patients can be accomplished in the beating heart with low operative mortality, provides good symptomatic relief and long-term survival, and is associated with objective evidence of improved left ventricular and mitral valve function. PMID- 11279408 TI - Did the introduction of a minimally invasive technique change the incidence of atrial fibrillation after single internal thoracic artery-left anterior descending artery grafting? AB - OBJECTIVE: Atrial fibrillation after coronary artery bypass operations remains frequent and increases morbidity, as well as resource use. Its cause remains unclear. The introduction of a minimally invasive technique provides an opportunity to evaluate the effect of intraoperative factors, such as cardiopulmonary bypass, global myocardial ischemia, and myocardial protection technique, on the occurrence of this arrhythmia. METHODS: All the patients undergoing isolated left internal thoracic artery-left anterior descending artery grafting between January 1994 and December 1999 were reviewed. Twenty possible risk factors for postoperative atrial fibrillation, including the choice of operative technique--minimally invasive technique was introduced in January 1997- were entered into univariate and multivariable logistic regression analysis. RESULTS: Postoperative atrial fibrillation occurred in 36 (20%) of 183 patients. On univariate analysis, age (P <.001) and a history of supraventricular arrhythmia (P <.001) were found to be risk factors. In particular, 15 (22%) of 69 patients operated on with the minimally invasive technique had postoperative atrial fibrillation versus 21 (18%) of 114 in the standard group (P =.58). On multivariable analysis, including the operative technique, the same variables (P =.001 and.01, respectively) were identified as independent risk factors. CONCLUSIONS: The introduction of a minimally invasive technique for coronary artery bypass operations did not reduce the occurrence of postoperative atrial fibrillation in this study population. This suggests that prophylactic measures to reduce this arrhythmia should be focused on factors unrelated to cardiopulmonary bypass or myocardial preservation technique. PMID- 11279409 TI - Reduced postoperative blood loss and transfusion requirement after beating-heart coronary operations: a prospective randomized study. AB - OBJECTIVE: Coronary artery bypass grafting on the beating heart through median sternotomy is a relatively new treatment, which allows multiple revascularization without the use of cardiopulmonary bypass. A prospective randomized study was designed to investigate the effect of coronary bypass with or without cardiopulmonary bypass on postoperative blood loss and transfusion requirement. METHODS: Two hundred patients with coronary artery disease were prospectively randomized to (1) on-pump treatment with conventional cardiopulmonary bypass and cardioplegic arrest and (2) off-pump treatment on the beating heart. Postoperative blood loss identified as total chest tube drainage, transfusion requirement, and related costs together with hematologic indices and clotting profiles were analyzed. RESULTS: There was no difference between the groups with respect to preoperative and intraoperative patient variables. The mean ratio of postoperative blood loss and 95% confidence interval between groups was 1.64 and 1.39 to 1.94, respectively, suggesting on average a postoperative blood loss 1.6 times higher in the on-pump group compared with the off-pump group. Seventy-seven patients in the off-pump group required no blood transfusion compared with only 48 in the on-pump group (P <.01). Furthermore, less than 5% of patients in the on pump group required fresh frozen plasma and platelet transfusion compared with 30% and 25%, respectively, in the on-pump group (both P <.05). Mean transfusion cost per patient was higher in the on-pump compared with that in the off-pump group ($184.8 +/- $35.2 vs $21.47 +/- $6.9, P <.01). CONCLUSIONS: Coronary artery bypass grafting on the beating heart is associated with a significant reduction in postoperative blood loss, transfusion requirement, and transfusion-related cost when compared with conventional revascularization with cardiopulmonary bypass and cardioplegic arrest. PMID- 11279410 TI - Heart valves from pigs and the porcine endogenous retrovirus: experimental and clinical data to assess the probability of porcine endogenous retrovirus infection in human subjects. AB - OBJECTIVE: Replacement of heart valves in human subjects has become a routine procedure in cardiac operations. We sought to investigate whether commercially available glutaraldehyde-fixed porcine heart valve prostheses cause porcine endogenous retrovirus infection in human subjects because recent studies revealed that human cells can be infected with porcine endogenous retrovirus. METHODS: Blood samples of 18 patients who underwent aortic or mitral valve replacement with porcine heart valves were collected 6 months to 3 years after operation and tested for porcine endogenous retrovirus by means of polymerase chain reaction and reverse transcriptase-polymerase chain reaction. In addition, we tried to trace porcine endogenous retrovirus in 3 commercially available, glutaraldehyde fixed, porcine heart valves. RESULTS: Porcine endogenous retrovirus can be easily detected in native porcine heart valves and degrades completely within 1 week of fixation in glutaraldehyde. In all 3 commercially available porcine heart valves, no traces of porcine endogenous retrovirus were found. All blood samples showed negative test results for the porcine endogenous retrovirus genome. CONCLUSION: Our results indicate that glutaraldehyde fixation of porcine heart valves reliably prevents cross-species transmission of porcine endogenous retrovirus. PMID- 11279411 TI - Ministernotomy versus complete sternotomy for coronary bypass operations: no difference in postoperative pulmonary function. AB - OBJECTIVES: Less-invasive approaches in cardiac operations offer certain cosmetic advantages, but it is unclear whether there are additional positive effects with regard to the postoperative recovery of patients. The aim of this prospective and randomized study was to ascertain whether partial inferior midline sternotomy can improve pulmonary function, one of the best quantifiable parameters of postoperative recovery, after coronary artery bypass operations when compared with the standard full midline approach. METHODS: One hundred patients scheduled for elective coronary artery bypass grafting were randomized either for a full median sternotomy (standard sternotomy group, n = 50) or for a partial inferior sternotomy (ministernotomy group, n = 50). The following pulmonary features were assessed: vital capacity, forced expiratory volume, percentage of forced expiratory volume from vital capacity, total lung capacity, residual volume, maximum inspiratory pressure, and maximum expiratory pressure. Tests were performed preoperatively and on the fourth and tenth postoperative days. RESULTS: On the fourth postoperative day, both groups had a significant decrease in vital capacity (percentage of predicted values) when compared with preoperative values (preoperative vs fourth day: standard sternotomy group, 87.8% +/- 14.3% vs 42.1% +/- 10.2% [P <.0001]; ministernotomy group, 84.5% +/- 14.3% vs 41.5% +/- 11.8% [P <.0001]), with a significant tendency for recovery from the fourth to the tenth postoperative day (fourth vs tenth postoperative day: standard sternotomy group, 42.1% +/- 10.2% vs 66.3% +/- 12.3% [P =.001]; ministernotomy group, 41.5% +/- 11.8% vs 61.3% +/- 13.1 % [P =.002]). There were no differences in any test results between the groups on either the fourth or the tenth postoperative day. CONCLUSION: A less-invasive approach for coronary artery bypass operations with a partial inferior sternotomy does not improve early postoperative pulmonary function when compared with the conventional approach with a full sternotomy. PMID- 11279412 TI - Minimally invasive versus sternotomy approaches for mitral reconstruction: comparison of intermediate-term results. AB - BACKGROUND: This study compares intermediate-term outcomes of mitral valve reconstruction after either the standard sternotomy approach or the new minimally invasive approach. Although minimally invasive mitral valve operations appear to offer certain advantages, such as reduced postoperative discomfort and decreased postoperative recovery time, the intermediate-term functional and echocardiographic efficacy has not yet been documented. METHODS: From May 1996 to February 1999, 100 consecutive patients underwent primary mitral reconstruction through a minimally invasive right anterior thoracotomy and peripheral cardiopulmonary bypass and Port-Access technology (Heartport, Inc, Redwood City, Calif). Outcomes were compared with those for our previous 100 patients undergoing primary mitral repair who were operated on with the standard sternotomy approach. RESULTS: Although patients were similar in age, the patients undergoing the minimally invasive approach had a lower preoperative New York Heart Association classification (2.1 +/- 0.5 vs 2.6 +/- 0.6, P <.001). There was one (1.0%) hospital mortality with the sternotomy approach and no such case with the minimally invasive approach. Follow-up revealed that residual mitral insufficiency was similar between the minimally invasive and sternotomy approaches (0.79 +/- 0.06 vs 0.77 +/- 0.06, P =.89, 0- to 3-point scale); likewise, the cumulative freedom from reoperation was not significantly different (94.4% vs 96.8%, P =.38). Follow-up New York Heart Association functional class was significantly better in the patients undergoing the minimally invasive approach (1.5 +/- 0.05 vs 1.2 +/- 0.05, P <.01). CONCLUSIONS: These findings demonstrate comparable 1-year follow-up results after minimally invasive mitral valve reconstruction. Both echocardiographic results and New York Heart Association functional improvements were compatible with results achieved with the standard sternotomy approach. The minimally invasive approach for mitral valve reconstruction provides equally durable results with marked advantages for the patient and should be more widely adopted. PMID- 11279413 TI - Long-term stabilization of vein graft wall architecture and prolonged resistance to experimental atherosclerosis after E2F decoy oligonucleotide gene therapy. AB - OBJECTIVE: We tested the hypothesis that a single intraoperative transfection of rabbit vein grafts with a decoy oligonucleotide that blocks cell-cycle gene transactivation by the transcription factor E2F induces long-term stable adaptation that involves medial hypertrophy and a resistance to neointimal hyperplasia and atherosclerosis. METHODS: Jugular vein to carotid artery interposition vein grafts in hypercholesterolemic rabbits were treated, using pressure-mediated delivery, with either E2F decoy oligonucleotide, scrambled oligonucleotide, or vehicle alone. E2F decoy inhibition of cell-cycle gene expression was determined by measuring proliferating cell nuclear antigen upregulation and bromodeoxyuridine incorporation in vascular smooth muscle cells. Neointimal hyperplasia and atherosclerosis were compared between groups at 6 months after operation. Wall stress was derived from the ratio of luminal radius to wall thickness. Normal rabbits exposed to 6 weeks of diet-induced hypercholesterolemia starting 6 months after operation were analyzed in the same manner. RESULTS: The E2F decoy oligonucleotide, but not scrambled oligonucleotide or vehicle alone, inhibited proliferating cell nuclear antigen expression and smooth muscle cell proliferation. Furthermore, this manipulation of cell-cycle gene expression yielded an inhibition of neointimal hyperplasia and atherosclerotic plaque formation throughout the 6 months of cholesterol feeding. In normocholesterolemic rabbits, vehicle-treated and scrambled oligonucleotide treated vein grafts remain susceptible to diet-induced atherosclerosis as well, whereas resistance to this disease induction remained stable in genetically engineered grafts. CONCLUSION: A single intraoperative pressure-mediated delivery of E2F decoy effectively provides vein grafts with long-term resistance to neointimal hyperplasia and atherosclerosis. These findings suggest that long-term reduction in human vein graft failure rates may be feasible with this ex vivo gene therapy approach. PMID- 11279414 TI - Standard versus hemodynamic plus 19-mm St Jude Medical aortic valves. AB - OBJECTIVE: We reviewed our experience with aortic valve replacement using 19-mm St Jude Medical prostheses (St Jude Medical, Inc, St Paul, Minn) in 119 patients, among which 68 (group A) had a Standard model and 51 (group B) had a Hemodynamic Plus model. METHODS: Comparison between the 2 models included analysis of early and late mortality and all valve-related complications. Postoperative echocardiography was performed to evaluate the hemodynamic performance of both prosthetic models. Laboratory tests were performed to evaluate the amount of red blood cell damage caused by the transprosthetic turbulent flow. RESULTS: Average body surface area was 1.66 +/- 0.14 m(2) in group A and 1.65 +/- 0.16 m(2) in group B (P =.72). There was no statistically significant difference between the 2 groups in terms of preoperative variables (sex, cardiac rhythm, body surface area, preoperative gradients, and New York Heart Association class). Five-year follow-up was 100% complete. Although group A patients had significantly higher postoperative peak and mean gradients (P =.0001) and a lower effective orifice area (P =.0001), no statistical differences were found in terms of late (5-year) survival (P =.6) and postoperative complications (P =.09). Moreover, postoperative left ventricular mass was found to be similar in the 2 groups (P =.18). Hematologic evaluation did not show any significant difference between the 2 groups as to incidence of hemolysis. CONCLUSIONS: Aortic valve replacement with 19-mm aortic prostheses in patients with a body surface area of less than 1.7 m(2) allows good results. Although Hemodynamic Plus models have better hemodynamic results, no significant difference was found in terms of clinical results and clinical hemolysis. PMID- 11279415 TI - Chymase-dependent angiotensin II formation in the saphenous vein versus the internal thoracic artery. AB - OBJECTIVES: The great saphenous vein graft is known to be less patent than the internal thoracic artery graft. Recently, we reported that chymase-dependent angiotensin II formation plays an important role in the development of intimal hyperplasia in dog grafted veins. In this study we investigated the levels of angiotensin II-forming enzymes, angiotensin-converting enzyme, and chymase in human saphenous veins and internal thoracic arteries. METHODS: The saphenous vein and internal thoracic artery specimens were obtained from coronary artery bypass grafts of patients during surgical procedures (saphenous vein, n = 16; internal thoracic artery, n = 16). Activities of angiotensin-converting enzyme and chymase were determined by using the extract from the saphenous vein or internal thoracic artery. Sections of the saphenous vein or internal thoracic artery were stained with van Gieson's elastin stain and were immunostained with anti-human chymase antibody. RESULTS: The activities of angiotensin-converting enzyme in the saphenous vein and internal thoracic artery were 0.34 +/- 0.12 and 0.32 +/- 0.17 mU/mg protein, respectively, and the difference was not significant. The chymase activity in the saphenous vein was significantly higher than that in the internal thoracic artery (saphenous vein, 10.1 +/- 0.81 mU/mg protein; internal thoracic artery, 6.21 +/- 1.86 mU/mg protein). Chymase-positive cells in the saphenous vein were located in both the media and adventitia, and those in the internal thoracic artery were located only in the adventitia. The number of chymase positive cells in the saphenous vein was about 2.6 times that in the internal thoracic artery. CONCLUSION: The chymase activity, but not the angiotensin converting enzyme activity, was significantly higher in the saphenous vein, suggesting that the high levels of chymase activity may be related to the poorer performance of the saphenous vein for use as a bypass conduit. PMID- 11279417 TI - Neuropsychologic impairment after coronary bypass surgery: effect of gaseous microemboli during perfusionist interventions. AB - OBJECTIVE: Neuropsychologic impairment is a common complication of coronary bypass surgery. Cerebral microemboli during cardiopulmonary bypass are the principal cause of cognitive deficits after coronary bypass grafting. We have previously demonstrated that the majority of cerebral emboli occur during perfusionist interventions (ie, during the injection of air into the venous side of the cardiopulmonary bypass circuit). The purpose of this study was to determine whether an increase in perfusionist interventions is associated with an increased risk of postoperative cognitive impairment. METHODS: Patients undergoing elective coronary artery bypass grafting (n = 83) underwent a battery of neuropsychologic tests preoperatively and 3 months postoperatively. Patients were divided into 2 groups according to the median value of perfusionist interventions during cardiopulmonary bypass. Group 1 patients (n = 42) had fewer than 10 perfusionist interventions, and group 2 patients (n = 41) had 10 or more interventions. RESULTS: The 2 groups of patients were similar for all preoperative, intraoperative, and postoperative variables, with the exception of longer cardiopulmonary bypass times in group 2 patients (P <.001). Group 2 patients had lower mean scores on 9 of 10 neuropsychologic tests, with 3 (Rey Auditory Verbal Learning, Digit Span, and Visual Span) being statistically significant. Group 2 patients had worse cognitive test scores, even when controlling for increased bypass times. Group 2 patients had a nonsignificant trend toward an increased prevalence of neuropsychologic impairment 3 months postoperatively. CONCLUSIONS: Introduction of air into the cardiopulmonary bypass circuit by perfusionists, resulting in cerebral microembolization, may contribute to postoperative cognitive impairment. PMID- 11279416 TI - Expression of vascular endothelial growth factor and its receptors is increased, but microvascular relaxation is impaired in patients after acute myocardial ischemia. AB - BACKGROUND: Vascular endothelial growth factor, a specific endothelial mitogen, plays an important role in myocardial angiogenesis. Previous work has demonstrated increased expression of vascular endothelial growth factor and its receptors in a rat myocardial infarction model, as well as in a pig model of chronic ischemia. The expression of vascular endothelial growth factor and other growth factors after acute myocardial ischemia in patients has not been examined. In this study we examined the expression of vascular endothelial growth factor and its receptors and the responsiveness of human atrial microvessels to vascular endothelial growth factor before and after acute ischemia. METHODS: Paired specimens of human atrial tissue were harvested before and after atrial devascularization (ligation) in 16 patients undergoing coronary bypass operations. RESULTS: The messenger RNA (reverse transcriptase-polymerase chain reaction) level of vascular endothelial growth factor and vascular endothelial growth factor receptor 1 were increased by 22.2% +/- 4.2% and 30.7% +/- 7.6%, respectively (P <.05), in the ischemic specimens as compared with the control specimens. Protein expression (Western blotting) of vascular endothelial growth factor and that of vascular endothelial growth factor receptor 1 also were increased significantly by 71.7% +/- 27.8% and 68.2% +/- 27.6%, respectively (P <.05). However, both RNA and protein expressions of another vascular endothelial growth factor receptor, vascular endothelial growth factor receptor 2, and fibroblast growth factor and fibroblast growth factor receptor 1 were unchanged. Reactivity of precontracted atrial vessels was examined with video microscopy. Vascular endothelial growth factor-induced (33.9% +/- 2.4% vs 18.3% +/- 2.8% in control and ischemic vessels, respectively; P <.05), fibroblast growth factor induced (31.6% +/- 3.2% vs 15.8% +/- 4.1%, P <.05), and substance P-induced (84.5% +/- 3.7% vs 54.3% +/- 9.0%, P <.05) microvascular relaxations were decreased in ischemic samples and in the presence of N (G)nitro-L -arginine, whereas responses to sodium nitroprusside were unchanged (90.9% +/- 2.2% vs 91.2% +/- 2.0%). CONCLUSIONS: This study suggests that acute myocardial ischemia in patients results in increased expression of vascular endothelial growth factor but not fibroblast growth factor and that the functional activity of vascular endothelial growth factor receptors and that of other growth factors may be impaired. PMID- 11279419 TI - Aspects of the spinal cord circulation as assessed by intrathecal oxygen tension monitoring during various arterial interruptions in the pig. AB - OBJECTIVE: We sought to study the effect of various modes of interruption of the spinal cord blood supply on intrathecal oxygenation. METHODS: In 24 pigs intrathecal PO (2), PCO (2), and pH were continuously monitored with a multiparameter catheter (Paratrend 7, Biomedical Sensors; Diametrics Medical, Inc, St Paul, Minn) during and after aortic crossclamping or selective interruption of segmental arteries and proximal collateral circulation. RESULTS: Proximal aortic clamping (n = 6) produced complete ischemia, whereas a second clamp close to the celiac trunk (n = 4) partly protected against spinal cord ischemia. This is explained by prevention of the steal phenomenon in the excluded part of the aorta. Adding clamps to the subclavian arteries (n = 6) created complete spinal ischemia as the collateral circulation was interrupted. In another group (n = 4) all segmental arteries below T5 were occluded with no reaction in the intrathecal variables. Additional selective clamping of supreme intercostal arteries (n = 4) showed the relative importance of the subclavian and vertebral collateral pathways. CONCLUSIONS: Continuous intrathecal PO (2) was monitored during various modes of interruption of the spinal cord blood supply. This provided insight into the ischemia mechanisms and relative importance of the segmental contribution and proximal collateral pathways of the spinal cord circulation in pigs. A short literature review is given, and aspects of comparative anatomy are discussed. PMID- 11279418 TI - Left ventricular aneurysm repair in rats: structural, functional, and molecular consequences. AB - OBJECTIVES: This study examined the effects of aneurysm repair in a rat model of myocardial infarction on functional indices and on the spatiotemporal distribution of cardiac contractile protein and natriuretic peptide messenger RNA. METHODS: In a rat infarct model, expanded left ventricular aneurysms were plicated 4 weeks after infarction. At 30 weeks, transverse heart sections were taken at 4 levels (apex [level 1] through base [level 4]) and assessed by in situ hybridization histochemistry to determine regional messenger RNA levels of pre pro-atrial natriuretic peptide, cardiac alpha-actin, skeletal alpha-actin, myosin light chain-2v, and beta-myosin heavy chain. RESULTS: Rats with plicated left ventricular aneurysms had reduced left ventricular endocardial circumference (19%, P <.005), lower heart weight ratio (31%, P <.05), left ventricular end diastolic pressures (51%, P <.05), and increased +/-dP/dt (34%-38%, P <.05). Cardiac messenger RNA levels of pre-pro-atrial natriuretic peptide were reduced in the septum (levels 2 and 3), and skeletal alpha-actin levels were reduced in the septum and left ventricular free wall of plicated rats (level 3). beta-Myosin heavy chain levels were markedly reduced in peri-infarct regions of the left ventricular free wall, septum, and right ventricle in plicated rats at level 4, whereas myosin light chain-2v levels were reduced at levels 2 and 4 in the left ventricular free wall and at level 4 in the right ventricle. CONCLUSIONS: Plication of left ventricular aneurysm after infarction in the rat significantly reduced cardiac hypertrophy, improved cardiac function, and reduced the upregulation of pre-pro-atrial natriuretic peptide and both fetal and adult contractile protein isoforms associated with cardiac hypertrophy. PMID- 11279420 TI - Inactivation of the MEK/ERK pathway in the myocardium during cardiopulmonary bypass. AB - OBJECTIVES: A general pro-inflammatory response after cardiopulmonary bypass (CPB) may involve changes in signal transduction and in part be responsible for arrhythmias and myocardial dysfunction after cardiac surgery. The MEK/ERK (mitogen-activated protein kinase kinase/extracellular regulated kinase) pathway is common to many stimuli and may play a pivotal role in morbidity associated with CPB. We investigated the changes in MEK/ERK pathway and related enzymes after CPB in pigs. METHODS: We examined ventricular and atrial tissue from pigs before 90 minutes of normothermic CPB and after 90 minutes of post-CPB perfusion. The activities and protein levels of kinases MEK1/2, ERK1/2, a cellular tyrosine kinase (c-Src), protein kinase B (Akt), and the protein levels of mitogen activated protein kinase phosphatase (MKP-1) were studied by immunoblotting ventricular and atrial myocardium lysates and labeling sections with antibodies that recognize the activated forms of the kinases and the phosphatase. Control pigs were subjected to sternotomy and heparinization but not CPB. RESULTS: We found a consistent inactivation of MEK/ERK pathway in both ventricular and atrial myocardium with an increase in MKP-1, a negative regulator of ERK1/2. The activities and protein levels of c-Src and Akt were not significantly modified before or after CPB, suggesting a certain degree of specificity for the MEK/ERK pathway. Such changes were not observed in controls. The decrease of ERK1/2 and MEK1/2 phosphorylation 90 minutes after termination of CPB (as well as the increase of nuclear MKP-1 protein levels) was also apparent by confocal microscopy. CONCLUSIONS: These results collectively reveal a prevalence of inhibitory mechanisms in the MEK/ERK signal transduction machinery in myocardium subjected to CPB. PMID- 11279421 TI - Pediatric heart transplantation: improving results in high-risk patients. AB - OBJECTIVES: Our institutional experience with 73 pediatric patients undergoing cardiac transplantation between January 1, 1990, and December 31, 1999, was reviewed to determine the impact of unconventional donor and recipient management protocols implemented to extend the availability of this therapy. METHODS AND RESULTS: The introduction of donor blood cardioplegic solution with added insulin was associated with a significant improvement in patient and graft survival (hazard ratio [Cox] = 0.25, P =.08), despite significantly longer ischemic times with this protocol compared with the use of crystalloid-based donor procurement techniques (P <.01). Eleven patients underwent intentional transplantation of ABO incompatible donor hearts with the aid of a protocol of plasma exchange on bypass. In this subgroup, there were 2 early deaths caused by nonspecific graft failure (n = 1) and respiratory complications with mild vascular rejection (n = 1), and there was 1 late death caused by lymphoma. ABO-incompatible transplantation was not a risk factor for death by multivariate analysis. The postoperative course in these patients suggests minimal reactivity directed against incompatible grafts on the basis of low anti-donor blood group antibody production, in association with a favorable rejection profile. Ten of 13 patients requiring preoperative support with an extracorporeal membrane oxygenator survived transplantation; there were 3 additional late deaths in this subgroup (hazard ratio = 2.88, P =.05). CONCLUSIONS: The results with pediatric cardiac transplantation continue to improve as a result of changes in both surgical and medical protocols permitting successful treatment of patients conventionally considered at high risk or unsuitable for transplantation. PMID- 11279422 TI - Surgical intervention for anomalous origin of the left coronary artery from the pulmonary artery: the Tokyo experience. AB - BACKGROUND: Few studies after surgical repair of the anomalous origin of the left coronary artery have reported the importance of the mitral annuloplasty or the long-term results. METHODS: Between January 1982 and March 2000, 29 patients with anomalous origin underwent surgical intervention at our institution (direct aortic reimplantation in 19 and Takeuchi procedure in 10). Age at the time of operation ranged from 2 months to 24 years (median, 29.3 months), and 9 patients were infants. Twenty-four patients had varying degrees of mitral incompetence. Simultaneous mitral annuloplasty at the anterolateral commissure was performed in all 24 patients with incompetence. RESULTS: There were 2 hospital deaths among the infants, and no late deaths. Mean follow-up was 100 +/- 57 months, and the actuarial survival was 93.1% at 10 years (70% confidence limits, 87-99). Cardiothoracic ratio at discharge was not decreasing significantly (P =.35); however, this value 5 years after the operation showed the significant decrease (P =.003) versus preoperative value. Preoperative mitral incompetence decreased in all but one of the operative survivors with mitral annuloplasty at the last follow-up. The left ventricular fractional shortening z-score was not normalized at discharge but was normalized in the late period. CONCLUSION: These data demonstrate that impaired left ventricular function normalized in the long term (even if it was below normal immediately after operation) after 2-coronary repair. We recommend that the simultaneous mitral annuloplasty should be performed at the time of operation for patients who have mitral incompetence with anomalous origin of the left coronary artery. PMID- 11279423 TI - Prenatal diagnosis of congenital heart disease affects preoperative acidosis in the newborn patient. AB - OBJECTIVES: Congenital heart disease is the leading cause of death in the first year after birth. Prenatal diagnosis of the disease can optimize the preoperative condition of the patient and may help in the prevention of acidosis. In this retrospective study we compared the occurrence of metabolic acidosis in patients with and without prenatal diagnosis of a congenital heart disease. METHODS: Data of 408 patients who needed an operation for congenital heart disease within 31 days of life were analyzed retrospectively. Arterial blood gases at fixed time intervals and worst blood gas of 81 patients with and 327 patients without a prenatal diagnosis were compared, categorizing the patients on ductus dependency, anticipated univentricular or biventricular repair, and left-sided, right-sided, or no heart obstruction. RESULTS: In the overall group significant differences in lowest pH, lowest base excess, and highest lactate level were found, with metabolic acidosis more common among the patients with a postnatal diagnosis. In the group of patients with ductus-dependent congenital heart disease, the difference between patients receiving a prenatal and those receiving a postnatal diagnosis was more significant than in the group with non-ductus-dependent lesions. Analyzing patients with right-sided, left-sided, and no obstruction separately, significant differences were found in the group with left-sided heart obstruction for lowest pH and base excess and in the group with right-sided heart obstruction for lowest base excess. CONCLUSIONS: Prenatal diagnosis of congenital heart disease minimizes metabolic acidosis in patients with congenital heart disease and may be associated with improved long-term outcome and prevention of cerebral damage among this fragile group of patients, although no significant effect on direct surgical outcome was encountered. PMID- 11279424 TI - Epicardial pacemaker implantation and follow-up in patients with a single ventricle after the Fontan operation. AB - OBJECTIVES: There is an increasing incidence of sinus node dysfunction after the Fontan procedure. Inability to maintain atrioventricular synchrony after the Fontan operation has been associated with an adverse late outcome. Although pacing may be helpful as a primary or adjunct modality after the Fontan procedure, the effects of performing a late thoracotomy or sternotomy for epicardial pacemaker implantation are unknown. In addition, little is known about the long-term effectiveness of epicardial leads in patients with single ventricles. The purpose of this study was to compare the hospital course and follow-up of epicardial pacing lead implantation in patients with Fontan physiology and patients with 2-ventricle physiology. METHODS: We retrospectively reviewed all isolated epicardial pacemaker implantations and outpatient evaluations performed between January 1983 and June 2000. RESULTS: There was no difference in the perioperative course for the 31 Fontan patients (27 atrial and 41 ventricular leads [68 total]) compared with the 56 non-Fontan subjects (9 atrial and 61 ventricular leads [70 total]). The median length of stay in Fontan and non-Fontan patients was 3 and 4 days, respectively. There was no early mortality in either group. Pleural drainage for 5 days or longer was reported in 4% of the Fontan cohort and 3% of the non-Fontan group. Late pleural effusions were identified in only 2 patients in the Fontan group and 2 patients in the non Fontan group. There was no significant difference in epicardial lead survival between the Fontan group and the non-Fontan group (1 year, 96%; 2 years, 90%; 5 years, 70%). The overall incidence of lead failure was 17% (24/138). CONCLUSIONS: Epicardial leads can be safely placed in Fontan patients at no additional risk compared to patients with biventricular physiology. Sensing and pacing qualities were relatively constant in both the Fontan and non-Fontan groups over the first 2 years after implantation. PMID- 11279425 TI - Tricuspid valvectomy for right ventricular outflow cannula occlusion with the Thoratec ventricular assist device. PMID- 11279426 TI - Pleuroperitoneal shunts and tumor seeding. AB - A 76-year-old man with malignant mesothelioma of the left pleura was referred for surgical palliation. He was dyspneic at rest and had anterior chest pain and a persistent cough. Chest x-ray film revealed an extensive left pleural effusion. A thoracoscopy was performed, and 3L of pleural fluid was drained. Both the pleural surfaces and rhe diaphragm were studded with tumors. On maximal inflation of the lung, the parietal and visceral pleura did not oppose, and therefore a Denver shunt was inserted. At 6 weeks follow-up, the shunt was performing satisfactorily. At follow-up 9 weeks postoperatively, the subcutaneous tunnel was infiltrated by mesothelioma over a distance of some 15 cm. PMID- 11279427 TI - Massive pulmonary embolus complicating left atrial myxoma. AB - Atrial myxomas may present with a classic triad of constitutional symptoms, embolic events, and intracardiac obstruction (1). We report a case of a massive pulmonary thromboembolus complicating a left atrial myxoma in the absence of an atrial or ventricular septal defect. PMID- 11279428 TI - Alternate explanation for the increasing oxygen consumption and lactatemia after surgery with hypothermic cardiopulmonary bypass. PMID- 11279429 TI - Alkalosis induced by alpha-stat management: cause of neuronal injury after deep hypothermic perfusion. PMID- 11279430 TI - Stentless bioprostheses should be properly adjusted according to the disposition of patient coronary ostia at both the inflow and outflow insertion levels. PMID- 11279431 TI - Congenital Horner syndrome. PMID- 11279433 TI - Vascular endothelial cell is a stem cell for neointimal formation after injury. PMID- 11279435 TI - Resection of lung cancer invading the diaphragm. PMID- 11279436 TI - Developing academic cardiothoracic surgeons. PMID- 11279437 TI - How to get your paper published. AB - Getting published depends heavily on starting with a well-designed, well-executed research question that is accurately described, and submitting it to a journal with an appropriate audience. PMID- 11279438 TI - Building a clinical cardiothoracic surgical program: a multi-institutional model. AB - Building a multi-institutional cardiothoracic surgical program has the same guiding principles and values as a traditional single institutional program: ensuring high-quality patient care, training and fostering residents, recruiting and retaining quality faculty, and contributing to basic and clinical research. With a well-designed infrastructure and support system, this more complicated type of organization may permit academic cardiothoracic surgical programs to compete effectively and grow in a constantly changing economic and political environment. PMID- 11279439 TI - Building a clinical program in a single institution. AB - A successful clinical program within either a single institution or a multi institution complex requires the recruitment and retention of excellent faculty, a strong residency program, a successful, recognized research program, and leaders with administrative, organizational, and leadership skills. PMID- 11279440 TI - Getting promoted. AB - Promotion is an active process. At the beginning of his or her academic career, the surgeon should begin planning for this process. Surgeons need to understand the promotion documents at their institutions and to have a timetable for achieving tenure at the appropriate time. PMID- 11279441 TI - Becoming a division chief. AB - Leading a division or department of cardiothoracic surgery is both a tremendous honor and a significant responsibility. Key to such a position of leadership are committed, functional, and loyal teams focused on the end points of success, and the ability of the leader to develop a strategic vision and to implement a functional operating system. PMID- 11279442 TI - On medical management. AB - To be successful, academic medical centers must exhibit leadership, a strong foundation in science and education, wide-ranging clinical experience, continuous innovation, exemplary service, and an earned reputation for consistently good results. PMID- 11279443 TI - Developing new technology. AB - The specialty of cardiothoracic surgery should be one of constant innovation. Albert Einstein was a man of vision who said, "Imagination is more important than knowledge." With most technology expected to be obsolete in 5-7 years, thoracic surgeons need to draw from their imaginations to find innovative surgical solutions for the future. PMID- 11279444 TI - Influencing the political process for cardiothoracic surgeons. AB - Medicare was established by congressional legislation to provide excellent health care for all older US citizens. The economic viability of Medicare is in question, however, and the possibility of reduced services or program curtailments has been raised. To ensure appropriate support for our professional goals, as thoracic surgeons we must articulate our professional needs and expectations for continued medical progress. PMID- 11279445 TI - Problems with the president. PMID- 11279446 TI - Fossil-fuelled feuds. PMID- 11279447 TI - Delays allowed foot-and-mouth epidemic to sweep across Britain. PMID- 11279448 TI - Americans perplexed by GM food. PMID- 11279449 TI - Publishers challenged over access to papers. PMID- 11279450 TI - Butchery lay behind CJD cluster. PMID- 11279451 TI - Critics claim 'sight-saving' rice is over-rated. PMID- 11279453 TI - Race is on to win Australian funding for big science projects. PMID- 11279452 TI - Plans to reduce acceptable arsenic limit put on hold. PMID- 11279454 TI - Japan's academics get green light to make their fortunes. PMID- 11279455 TI - Sonar system offered special dispensation. PMID- 11279456 TI - Regulator rebuked over cannabis. PMID- 11279458 TI - The battle of Tugen Hills. PMID- 11279459 TI - Think like a bee. PMID- 11279461 TI - Faculty start-ups offer temptation to breach academic rules. PMID- 11279462 TI - Faculty start-ups offer temptation to breach academic rules . . . but Syngenta deal is a boon to Berkeley. PMID- 11279463 TI - 'Art' was a load of fluff. PMID- 11279464 TI - Managing foot-and-mouth. PMID- 11279470 TI - The good and the bad. PMID- 11279469 TI - Unwritten knowledge. PMID- 11279471 TI - Magnetic explosions in space. PMID- 11279472 TI - Palaeoanthropology Our newest oldest ancestor? PMID- 11279473 TI - Neurobiology Cannabinoids act backwards. PMID- 11279475 TI - Nanotechnology: Dragging single electrons. PMID- 11279474 TI - Plant biology: night moves of pregnant moths. PMID- 11279476 TI - Cognitive neuroscience: Colour my i's blue. PMID- 11279478 TI - Palaeontology: Chinese salamanders tell tales. PMID- 11279479 TI - Cancer: Chromosome defects in the colon. PMID- 11279482 TI - Pattern and intensity of physical activity. PMID- 11279483 TI - Forensic palaeontology: The Archaeoraptor forgery. AB - The Archaeoraptor fossil was announced as a 'missing link' and purported to be possibly the best evidence since Archaeopteryx that birds did, in fact, evolve from certain types of carnivorous dinosaur. It reportedly came from Early Cretaceous beds of China that have produced other spectacular fossils transitional between birds and extinct non-avian dinosaurs. But Archaeoraptor was revealed to be a forgery in which bones of a primitive bird and a non-flying dromaeosaurid dinosaur had been combined. Here we use high-resolution X-ray computed tomography (CT) to determine the nature and extent of the forgery, as well as how it was built, by imaging the fracture pattern and distribution of materials through the entire specimen. PMID- 11279484 TI - Liquid crystalline spinning of spider silk. AB - Spider silk has outstanding mechanical properties despite being spun at close to ambient temperatures and pressures using water as the solvent. The spider achieves this feat of benign fibre processing by judiciously controlling the folding and crystallization of the main protein constituents, and by adding auxiliary compounds, to create a composite material of defined hierarchical structure. Because the 'spinning dope' (the material from which silk is spun) is liquid crystalline, spiders can draw it during extrusion into a hardened fibre using minimal forces. This process involves an unusual internal drawdown within the spider's spinneret that is not seen in industrial fibre processing, followed by a conventional external drawdown after the dope has left the spinneret. Successful copying of the spider's internal processing and precise control over protein folding, combined with knowledge of the gene sequences of its spinning dopes, could permit industrial production of silk-based fibres with unique properties under benign conditions. PMID- 11279486 TI - Production of iron nanoparticles by laser irradiation in a simulation of lunar like space weathering. AB - 'Space weathering' is the term applied to the darkening and reddening of planetary surface materials with time, along with the changes to the depths of absorption bands in their optical spectra. It has been invoked to explain the mismatched spectra of lunar rocks and regolith, and between those of asteroids and meteorites. The formation of nanophase iron particles on regolith grains as a result of micrometeorite impacts or irradiation by the solar wind has been proposed as the main cause of the change in the optical properties. But laboratory simulations have not revealed the presence of these particles, although nano-second-pulse laser irradiation did reproduce the optical changes. Here we report observations by transmission electron microscopy of olivine samples subjected to pulse laser irradiation. We find within the amorphous vapour deposited rims of olivine grains nanophase iron particles similar to those observed in the rims of space-weathered lunar regolith grains. Reduction by hydrogen atoms implanted by the solar wind is therefore not necessary to form the particles. Moreover, the results support the idea that ordinary chondrites came from S-type asteroids, and thereby provides some constraints on the surface exposure ages of those asteroids. PMID- 11279485 TI - Essential role of the mitochondrial apoptosis-inducing factor in programmed cell death. AB - Programmed cell death is a fundamental requirement for embryogenesis, organ metamorphosis and tissue homeostasis. In mammals, release of mitochondrial cytochrome c leads to the cytosolic assembly of the apoptosome-a caspase activation complex involving Apaf1 and caspase-9 that induces hallmarks of apoptosis. There are, however, mitochondrially regulated cell death pathways that are independent of Apaf1/caspase-9. We have previously cloned a molecule associated with programmed cell death called apoptosis-inducing factor (AIF). Like cytochrome c, AIF is localized to mitochondria and released in response to death stimuli. Here we show that genetic inactivation of AIF renders embryonic stem cells resistant to cell death after serum deprivation. Moreover, AIF is essential for programmed cell death during cavitation of embryoid bodies-the very first wave of cell death indispensable for mouse morphogenesis. AIF-dependent cell death displays structural features of apoptosis, and can be genetically uncoupled from Apaf1 and caspase-9 expression. Our data provide genetic evidence for a caspase-independent pathway of programmed cell death that controls early morphogenesis. PMID- 11279487 TI - Rapid magnetic reconnection in the Earth's magnetosphere mediated by whistler waves. AB - Magnetic reconnection has a crucial role in a variety of plasma environments in providing a mechanism for the fast release of stored magnetic energy. During reconnection the plasma forms a 'magnetic nozzle', like the nozzle of a hose, and the rate is controlled by how fast plasma can flow out of the nozzle. But the traditional picture of reconnection has been unable to explain satisfactorily the short timescales associated with the energy release, because the flow is mediated by heavy ions with a slow resultant velocity. Recent theoretical work has suggested that the energy release is instead mediated by electrons in waves called 'whistlers', which move much faster for a given perturbation of the magnetic field because of their smaller mass. Moreover, the whistler velocity and associated plasma velocity both increase as the 'nozzle' becomes narrower. A narrower nozzle therefore no longer reduces the total plasma flow-the outflow is independent of the size of the nozzle. Here we report observations demonstrating that reconnection in the magnetosphere is driven by whistlers, in good agreement with the theoretical predictions. PMID- 11279488 TI - Manipulation of elementary charge in a silicon charge-coupled device. AB - The ultimate limit in the operation of an electronic device is the manipulation of a single charge. Such a limit has been achieved in single-electron tunnelling devices. However, these devices are based on multiple tunnel barriers and conductive islands, which are complex structures to fabricate. Here we demonstrate another type of device that can also manipulate elementary charge, but which is more suitable for large-scale integration. The device consists of two closely packed silicon wire-MOSFETs, which are commonly used building blocks of electronic circuits. We have developed a scheme to generate and store holes in the channels of either of these MOSFETs. Subsequently, holes can be transferred between the two MOSFETs at the level of an elementary charge, and their exact position can be monitored. This single-charge transfer device, which is operated at 25 K, is in effect a charge-coupled device. This is also the first realization of a silicon-based device that manipulates elementary charge. PMID- 11279489 TI - Vortex dynamics in superconducting MgB2 and prospects for applications. AB - The recently discovered superconductor magnesium diboride, MgB2, has a transition temperature, Tc, approaching 40 K, placing it intermediate between the families of low- and high-temperature superconductors. In practical applications, superconductors are permeated by quantized vortices of magnetic flux. When a supercurrent flows, there is dissipation of energy unless these vortices are 'pinned' in some way, and so inhibited from moving under the influence of the Lorentz force. Such vortex motion ultimately determines the critical current density, Jc, which the superconductor can support. Vortex behaviour has proved to be more complicated in high-temperature superconductors than in low-temperature superconductors and, although this has stimulated extensive theoretical and experimental research, it has also impeded applications. Here we describe the vortex behaviour in MgB2, as reflected in Jc and in the vortex creep rate, S, the latter being a measure of how fast the 'persistent' supercurrents decay. Our results show that naturally occurring grain boundaries are highly transparent to supercurrents, a desirable property which contrasts with the behaviour of the high-temperature superconductors. On the other hand, we observe a steep, practically deleterious decline in Jc with increasing magnetic field, which is likely to reflect the high degree of crystalline perfection in our samples, and hence a low vortex pinning energy. PMID- 11279490 TI - Extreme damping in composite materials with negative-stiffness inclusions. AB - When a force deforms an elastic object, practical experience suggests that the resulting displacement will be in the same direction as the force. This property is known as positive stiffness. Less familiar is the concept of negative stiffness, where the deforming force and the resulting displacement are in opposite directions. (Negative stiffness is distinct from negative Poisson's ratio, which refers to the occurrence of lateral expansion upon stretching an object.) Negative stiffness can occur, for example, when the deforming object has stored (or is supplied with) energy. This property is usually unstable, but it has been shown theoretically that inclusions of negative stiffness can be stabilized within a positive-stiffness matrix. Here we describe the experimental realization of this composite approach by embedding negative-stiffness inclusions of ferroelastic vanadium dioxide in a pure tin matrix. The resulting composites exhibit extreme mechanical damping and large anomalies in stiffness, as a consequence of the high local strains that result from the inclusions deforming more than the composite as a whole. Moreover, for certain temperature ranges, the negative-stiffness inclusions are more effective than diamond inclusions for increasing the overall composite stiffness. We expect that such composites could be useful as high damping materials, as stiff structural elements or for actuator type applications. PMID- 11279491 TI - Climate variability 50,000 years ago in mid-latitude Chile as reconstructed from tree rings. AB - High-resolution proxies of past climate are essential for a better understanding of the climate system. Tree rings are routinely used to reconstruct Holocene climate variations at high temporal resolution, but only rarely have they offered insight into climate variability during earlier periods. Fitzroya cupressoides-a South American conifer which attains ages up to 3,600 years-has been shown to record summer temperatures in northern Patagonia during the past few millennia. Here we report a floating 1,229-year chronology developed from subfossil stumps of F. cupressoides in southern Chile that dates back to approximately 50,000 14C years before present. We use this chronology to calculate the spectral characteristics of climate variability in this time, which was probably an interstadial (relatively warm) period. Growth oscillations at periods of 150-250, 87-94, 45.5, 24.1, 17.8, 9.3 and 2.7-5.3 years are identified in the annual subfossil record. A comparison with the power spectra of chronologies derived from living F. cupressoides trees shows strong similarities with the 50,000-year old chronology, indicating that similar growth forcing factors operated in this glacial interstadial phase as in the current interglacial conditions. PMID- 11279492 TI - Simulating the amplification of orbital forcing by ocean feedbacks in the last glaciation. AB - According to Milankovitch theory, the lower summer insolation at high latitudes about 115,000 years ago allowed winter snow to persist throughout summer, leading to ice-sheet build-up and glaciation. But attempts to simulate the last glaciation using global atmospheric models have failed to produce this outcome when forced by insolation changes only. These results point towards the importance of feedback effects-for example, through changes in vegetation or the ocean circulation-for the amplification of solar forcing. Here we present a fully coupled ocean-atmosphere model of the last glaciation that produces a build-up of perennial snow cover at known locations of ice sheets during this period. We show that ocean feedbacks lead to a cooling of the high northern latitudes, along with an increase in atmospheric moisture transport from the Equator to the poles. These changes agree with available geological data and, together, they lead to an increased delivery of snow to high northern latitudes. The mechanism we present explains the onset of glaciation-which would be amplified by changes in vegetation-in response to weak orbital forcing. PMID- 11279493 TI - Late Jurassic salamanders from northern China. AB - With ten extant families, salamanders (urodeles) are one of the three major groups of modern amphibians (lissamphibians). Extant salamanders are often used as a model system to assess fundamental issues of developmental, morphological and biogeographical evolution. Unfortunately, our understanding of these issues has been hampered by the paucity of fossil evidence available to assess the early history of the group. Here we report the discovery of an extraordinary sample of salamander fossils, some with rare soft-tissue impressions, from the Upper Jurassic of China. With over 500 articulated specimens, this assemblage documents the morphological diversity of early urodeles and includes larvae and adults of both neotenic and metamorphosed taxa. Phylogenetic analysis confirms that these salamanders are primitive, and reveals that all basal salamander clades have Asian distributions. This is compelling evidence for an Asian origin of Recent salamanders, as well as for an extensive and early radiation of several major lineages. These discoveries show that the evolution of salamanders has involved phylogenetic and ecological diversification around a body plan that has remained fundamentally stable for over 150 million years. PMID- 11279494 TI - Caterpillar-induced nocturnal plant volatiles repel conspecific females. AB - Plants respond to insect herbivory by synthesizing and releasing complex blends of volatile compounds, which provide important host-location cues for insects that are natural enemies of herbivores. The effects of these volatile blends on herbivore behaviour have been investigated to only a limited extent, in part because of the assumption that herbivore-induced volatile emissions occur mainly during the light phase of the photoperiod. Because many moths-whose larvae are some of the most important insect herbivores-are nocturnal, herbivore-induced plant volatiles have not hitherto been considered to be temporally available as host-location cues for ovipositing females. Here we present chemical and behavioural assays showing that tobacco plants (Nicotiana tabacum) release herbivore-induced volatiles during both night and day. Moreover, several volatile compounds are released exclusively at night and are highly repellent to female moths (Heliothis virescens). The demonstration that tobacco plants release temporally different volatile blends and that lepidopteran herbivores use induced plant signals released during the dark phase to choose sites for oviposition adds a new dimension to our understanding of the role of chemical cues in mediating tritrophic interactions. PMID- 11279495 TI - Unconscious priming eliminates automatic binding of colour and alphanumeric form in synaesthesia. AB - Synaesthesia is an unusual perceptual phenomenon in which events in one sensory modality induce vivid sensations in another. Individuals may 'taste' shapes, 'hear' colours, or 'feel' sounds. Synaesthesia was first described over a century ago, but little is known about its underlying causes or its effects on cognition. Most reports have been anecdotal or have focused on isolated unusual cases. Here we report an investigation of 15 individuals with colour-graphemic synaesthesia, each of whom experiences idiosyncratic but highly consistent colours for letters and digits. Using a colour-form interference paradigm, we show that induced synaesthetic experiences cannot be consciously suppressed even when detrimental to task performance. In contrast, if letters and digits are presented briefly and masked, so that they are processed but unavailable for overt report, the synaesthesia is eliminated. These results show that synaesthetic experiences can be prevented despite substantial processing of the sensory stimuli that otherwise trigger them. We conclude that automatic binding of colour and alphanumeric form in synaesthesia arises after initial processes of letter and digit recognition are complete. PMID- 11279496 TI - Vertical interactions across ten parallel, stacked representations in the mammalian retina. AB - The mammalian visual system analyses the world through a set of separate spatio temporal channels. The organization of these channels begins in the retina, where the precise laminations of both the axon terminals of bipolar cells and the dendritic arborizations of ganglion cells suggests the presence of a vertical stack of neural strata at the inner plexiform layer (IPL). Conversely, many inhibitory amacrine cell classes are multiply or diffusely stratified, indicating that they might convey information between strata. On the basis of the diverse stratification and physiological properties of ganglion cells, it was suggested that the IPL contains a parallel set of representations of the visual world embodied in the strata and conveyed to higher centres by the classes of ganglion cells whose dendrites ramify at that stratum. Here we show that each stratum receives unique and substantively different excitatory and inhibitory neural inputs that are integrated to form at least ten different, parallel space-time spiking outputs. The response properties of these strata are ordered in the time domain. Inhibition through GABAC receptors extracts spatial edges in neural representations and seems to separate the functional properties of the strata. We describe a new form of neuronal interaction that we call 'vertical inhibition' that acts not laterally, but between strata. PMID- 11279497 TI - Endogenous cannabinoids mediate retrograde signalling at hippocampal synapses. AB - Marijuana affects brain function primarily by activating the G-protein-coupled cannabinoid receptor-1 (CB1), which is expressed throughout the brain at high levels. Two endogenous lipids, anandamide and 2-arachidonylglycerol (2-AG), have been identified as CB1 ligands. Depolarized hippocampal neurons rapidly release both anandamide and 2-AG in a Ca2+-dependent manner. In the hippocampus, CB1 is expressed mainly by GABA (gamma-aminobutyric acid)-mediated inhibitory interneurons, where CB1 clusters on the axon terminal. A synthetic CB1 agonist depresses GABA release from hippocampal slices. These findings indicate that the function of endogenous cannabinoids released by depolarized hippocampal neurons might be to downregulate GABA release. Here we show that the transient suppression of GABA-mediated transmission that follows depolarization of hippocampal pyramidal neurons is mediated by retrograde signalling through release of endogenous cannabinoids. Signalling by the endocannabinoid system thus represents a mechanism by which neurons can communicate backwards across synapses to modulate their inputs. PMID- 11279498 TI - Ca2+ signalling between single L-type Ca2+ channels and ryanodine receptors in heart cells. AB - Ca2+-induced Ca2+ release is a general mechanism that most cells use to amplify Ca2+ signals. In heart cells, this mechanism is operated between voltage-gated L type Ca2+ channels (LCCs) in the plasma membrane and Ca2+ release channels, commonly known as ryanodine receptors, in the sarcoplasmic reticulum. The Ca2+ influx through LCCs traverses a cleft of roughly 12 nm formed by the cell surface and the sarcoplasmic reticulum membrane, and activates adjacent ryanodine receptors to release Ca2+ in the form of Ca2+ sparks. Here we determine the kinetics, fidelity and stoichiometry of coupling between LCCs and ryanodine receptors. We show that the local Ca2+ signal produced by a single opening of an LCC, named a 'Ca2+ sparklet', can trigger about 4-6 ryanodine receptors to generate a Ca2+ spark. The coupling between LCCs and ryanodine receptors is stochastic, as judged by the exponential distribution of the coupling latency. The fraction of sparklets that successfully triggers a spark is less than unity and declines in a use-dependent manner. This optical analysis of single-channel communication affords a powerful means for elucidating Ca2+-signalling mechanisms at the molecular level. PMID- 11279499 TI - Drought-induced guard cell signal transduction involves sphingosine-1-phosphate. AB - Stomata form pores on leaf surfaces that regulate the uptake of CO2 for photosynthesis and the loss of water vapour during transpiration. An increase in the cytosolic concentration of free calcium ions ([Ca2+]cyt) is a common intermediate in many of the pathways leading to either opening or closure of the stomatal pore. This observation has prompted investigations into how specificity is controlled in calcium-based signalling systems in plants. One possible explanation is that each stimulus generates a unique increase in [Ca2+]cyt, or 'calcium signature', that dictates the outcome of the final response. It has been suggested that the key to generating a calcium signature, and hence to understanding how specificity is controlled, is the ability to access differentially the cellular machinery controlling calcium influx and release from internal stores. Here we report that sphingosine-1-phosphate is a new calcium mobilizing molecule in plants. We show that after drought treatment sphingosine-1 phosphate levels increase, and we present evidence that this molecule is involved in the signal-transduction pathway linking the perception of abscisic acid to reductions in guard cell turgor. PMID- 11279500 TI - Hedgehog acts as a somatic stem cell factor in the Drosophila ovary. AB - Secreted signalling molecules of the Hedgehog (Hh) family have many essential patterning roles during development of diverse organisms including Drosophila and humans. Although Hedgehog proteins most commonly affect cell fate, they can also stimulate cell proliferation. In humans several distinctive cancers, including basal-cell carcinoma, result from mutations that aberrantly activate Hh signal transduction. In Drosophila, Hh directly stimulates proliferation of ovarian somatic cells. Here we show that Hh acts specifically on stem cells in the Drosophila ovary. These cells cannot proliferate as stem cells in the absence of Hh signalling, whereas excessive Hh signalling produces supernumerary stem cells. We deduce that Hh is a stem-cell factor and suggest that human cancers due to excessive Hh signalling might result from aberrant expansion of stem cell pools. PMID- 11279501 TI - Structural basis for co-stimulation by the human CTLA-4/B7-2 complex. AB - Regulation of T-cell activity is dependent on antigen-independent co-stimulatory signals provided by the disulphide-linked homodimeric T-cell surface receptors, CD28 and CTLA-4 (ref. 1). Engagement of CD28 with B7-1 and B7-2 ligands on antigen-presenting cells (APCs) provides a stimulatory signal for T-cell activation, whereas subsequent engagement of CTLA-4 with these same ligands results in attenuation of the response. Given their central function in immune modulation, CTLA-4- and CD28-associated signalling pathways are primary therapeutic targets for preventing autoimmune disease, graft versus host disease, graft rejection and promoting tumour immunity. However, little is known about the cell-surface organization of these receptor/ligand complexes and the structural basis for signal transduction. Here we report the 3.2-A resolution structure of the complex between the disulphide-linked homodimer of human CTLA-4 and the receptor-binding domain of human B7-2. The unusual dimerization properties of both CTLA-4 and B7-2 place their respective ligand-binding sites distal to the dimer interface in each molecule and promote the formation of an alternating arrangement of bivalent CTLA-4 and B7-2 dimers that extends throughout the crystal. Direct observation of this CTLA-4/B7-2 network provides a model for the periodic organization of these molecules within the immunological synapse and suggests a distinct mechanism for signalling by dimeric cell-surface receptors. PMID- 11279506 TI - All hands on deck at dawn. PMID- 11279507 TI - How does the mouse get its trunk? PMID- 11279502 TI - Crystal structure of the B7-1/CTLA-4 complex that inhibits human immune responses. AB - Optimal immune responses require both an antigen-specific and a co-stimulatory signal. The shared ligands B7-1 and B7-2 on antigen-presenting cells deliver the co-stimulatory signal through CD28 and CTLA-4 on T cells. Signalling through CD28 augments the T-cell response, whereas CTLA-4 signalling attenuates it. Numerous animal studies and recent clinical trials indicate that manipulating these interactions holds considerable promise for immunotherapy. With the consequences of these signals well established, and details of the downstream signalling events emerging, understanding the molecular nature of these extracellular interactions becomes crucial. Here we report the crystal structure of the human CTLA-4/B7-1 co-stimulatory complex at 3.0 A resolution. In contrast to other interacting cell-surface molecules, the relatively small CTLA-4/B7-1 binding interface exhibits an unusually high degree of shape complementarity. CTLA-4 forms homodimers through a newly defined interface of highly conserved residues. In the crystal lattice, CTLA-4 and B7-1 pack in a strikingly periodic arrangement in which bivalent CTLA-4 homodimers bridge bivalent B7-1 homodimers. This zipper like oligomerization provides the structural basis for forming unusually stable signalling complexes at the T-cell surface, underscoring the importance of potent inhibitory signalling in human immune responses. PMID- 11279508 TI - Fanconi anemia and breast cancer: what's the connection? PMID- 11279509 TI - A particular GAP in mind. PMID- 11279510 TI - The pancreas and its heartless beginnings. PMID- 11279513 TI - Retrotransposons as epigenetic mediators of phenotypic variation in mammals. AB - Phenotypic variation in mammals is frequently attributed to the action of quantitative trait loci (QTL) or the environment, but may also be epigenetic in origin. Here we consider a mechanism for phenotypic variation based on interference of transcription by somatically active retrotransposons. Transcriptionally competent retrotransposons may number in the tens of thousands in mammalian genomes. We propose that silencing of retrotransposons occurs by cosuppression during early embryogenesis, but that this process is imperfect and produces a mosaic pattern of retrotransposon expression in somatic cells. Transcriptional interference by active retrotransposons perturbs expression of neighboring genes in somatic cells, in a mosaic pattern corresponding to activity of each retrotransposon. The epigenotype of retrotransposon activity is reset in each generation, but incomplete resetting can lead to heritable epigenetic effects. The stochastic nature of retrotransposon activity, and the very large number of genes that may be affected, produce subtle phenotypic variations even between genetically identical individuals, which may affect disease risk and be heritable in a non-mendelian fashion. PMID- 11279514 TI - The callipyge mutation enhances the expression of coregulated imprinted genes in cis without affecting their imprinting status. AB - The callipyge (CLPG) phenotype (from kappa(alpha)lambda(iota), "beautiful," and pi(iota)gamma(epsilon), "buttocks") described in sheep is an inherited muscular hypertrophy that is subject to an unusual parent-of-origin effect referred to as polar overdominance: only heterozygous individuals having inherited the CLPG mutation from their sire exhibit the muscular hypertrophy. The callipyge (clpg) locus was mapped to a chromosome segment of approximately 400 kb (refs. 2-4), which was shown to contain four genes (DLK1, GTL2, PEG11 and MEG8) that are preferentially expressed in skeletal muscle and subject to parental imprinting in this tissue. Here we describe the effect of the CLPG mutation on the expression of these four genes, and demonstrate that callipyge individuals have a unique expression profile that may account for the observed polar overdominance. PMID- 11279515 TI - Mutations in SIP1, encoding Smad interacting protein-1, cause a form of Hirschsprung disease. AB - Hirschsprung disease (HSCR) is sometimes associated with a set of characteristics including mental retardation, microcephaly, and distinct facial features, but the gene mutated in this condition has not yet been identified. Here we report that mutations in SIP1, encoding Smad interacting protein-1, cause disease in a series of cases. SIP1 is located in the deleted segment at 2q22 from a patient with a de novo t(2;13)(q22;q22) translocation. SIP1 seems to have crucial roles in normal embryonic neural and neural crest development. PMID- 11279516 TI - Single-nucleotide polymorphisms in the public domain: how useful are they? AB - There is a concerted effort by a number of public and private groups to identify a large set of human single-nucleotide polymorphisms (SNPs). As of March 2001, 2.84 million SNPs have been deposited in the public database, dbSNP, at the National Center for Biotechnology Information (http://www.ncbi.nlm.nih.gov/SNP/). The 2.84 million SNPs can be grouped into 1.65 million non-redundant SNPs. As part of the International SNP Map Working Group, we recently published a high density SNP map of the human genome consisting of 1.42 million SNPs (ref. 3). In addition, numerous SNPs are maintained in proprietary databases. Our survey of more than 1,200 SNPs indicates that more than 80% of TSC and Washington University candidate SNPs are polymorphic and that approximately 50% of the candidate SNPs from these two sources are common SNPs (with minor allele frequency of > or =20%) in any given population. PMID- 11279517 TI - Genetic linkage of childhood atopic dermatitis to psoriasis susceptibility loci. AB - We have carried out a genome screen for atopic dermatitis (AD) and have identified linkage to AD on chromosomes 1q21, 17q25 and 20p. These regions correspond closely with known psoriasis loci, as does a previously identified AD locus on chromosome 3q21. The results indicate that AD is influenced by genes with general effects on dermal inflammation and immunity. PMID- 11279518 TI - Hepatocyte nuclear factor-1alpha is an essential regulator of bile acid and plasma cholesterol metabolism. AB - Maturity-onset diabetes of the young type 3 (MODY3) is caused by haploinsufficiency of hepatocyte nuclear factor-1alpha (encoded by TCF1). Tcf1-/- mice have type 2 diabetes, dwarfism, renal Fanconi syndrome, hepatic dysfunction and hypercholestrolemia. Here we explore the molecular basis for the hypercholesterolemia using oligonucleotide microchip expression analysis. We demonstrate that Tcf1-/- mice have a defect in bile acid transport, increased bile acid and liver cholesterol synthesis, and impaired HDL metabolism. Tcf1-/- liver has decreased expression of the basolateral membrane bile acid transporters Slc10a1, Slc21a3 and Slc21a5, leading to impaired portal bile acid uptake and elevated plasma bile acid concentrations. In intestine and kidneys, Tcf1-/- mice lack expression of the ileal bile acid transporter (Slc10a2), resulting in increased fecal and urinary bile acid excretion. The Tcf1 protein (also known as HNF-1alpha) also regulates transcription of the gene (Nr1h4) encoding the farnesoid X receptor-1 (Fxr-1), thereby leading to reduced expression of small heterodimer partner-1 (Shp-1) and repression of Cyp7a1, the rate-limiting enzyme in the classic bile acid biosynthesis pathway. In addition, hepatocyte bile acid storage protein is absent from Tcf1-/- mice. Increased plasma cholesterol of Tcf1 /- mice resides predominantly in large, buoyant, high-density lipoprotein (HDL) particles. This is most likely due to reduced activity of the HDL-catabolic enzyme hepatic lipase (Lipc) and increased expression of HDL-cholesterol esterifying enzyme lecithin:cholesterol acyl transferase (Lcat). Our studies demonstrate that Tcf1, in addition to being an important regulator of insulin secretion, is an essential transcriptional regulator of bile acid and HDL cholesterol metabolism. PMID- 11279519 TI - Sequence diversity in CYP3A promoters and characterization of the genetic basis of polymorphic CYP3A5 expression. AB - Variation in the CYP3A enzymes, which act in drug metabolism, influences circulating steroid levels and responses to half of all oxidatively metabolized drugs. CYP3A activity is the sum activity of the family of CYP3A genes, including CYP3A5, which is polymorphically expressed at high levels in a minority of Americans of European descent and Europeans (hereafter collectively referred to as 'Caucasians'). Only people with at least one CYP3A5*1 allele express large amounts of CYP3A5. Our findings show that single-nucleotide polymorphisms (SNPs) in CYP3A5*3 and CYP3A5*6 that cause alternative splicing and protein truncation result in the absence of CYP3A5 from tissues of some people. CYP3A5 was more frequently expressed in livers of African Americans (60%) than in those of Caucasians (33%). Because CYP3A5 represents at least 50% of the total hepatic CYP3A content in people polymorphically expressing CYP3A5, CYP3A5 may be the most important genetic contributor to interindividual and interracial differences in CYP3A-dependent drug clearance and in responses to many medicines. PMID- 11279520 TI - Mutational and functional analyses reveal that ST7 is a highly conserved tumor suppressor gene on human chromosome 7q31. AB - Loss of heterozygosity (LOH) of markers on human chromosome 7q31 is frequently encountered in a variety of human neoplasias, indicating the presence of a tumor suppressor gene (TSG). By a combination of microcell-fusion and deletion-mapping studies, we previously established that this TSG resides within a critical region flanked by the genetic markers D7S522 and D7S677. Using a positional cloning strategy and aided by the availability of near-complete sequence of this genomic interval, we have identified a TSG within 7q31, named ST7 (for suppression of tumorigenicity 7; this same gene was recently reported in another context and called RAY1). ST7 is ubiquitously expressed in human tissues. Analysis of a series of cell lines derived from breast tumors and primary colon carcinomas revealed the presence of mutations in ST7. Introduction of the ST7 cDNA into the prostate-cancer-derived cell line PC3 had no effect on the in vitro proliferation of the cells, but abrogated their in vivo tumorigenicity. Our data indicate that ST7 is a TSG within chromosome 7q31 and may have an important role in the development of some types of human cancer. PMID- 11279521 TI - Learning deficits, but normal development and tumor predisposition, in mice lacking exon 23a of Nf1. AB - Neurofibromatosis type 1 (NF1) is a commonly inherited autosomal dominant disorder. Previous studies indicated that mice homozygous for a null mutation in Nf1 exhibit mid-gestation lethality, whereas heterozygous mice have an increased predisposition to tumors and learning impairments. Here we show that mice lacking the alternatively spliced exon 23a, which modifies the GTPase-activating protein (GAP) domain of Nf1, are viable and physically normal, and do not have an increased tumor predisposition, but show specific learning impairments. Our findings have implications for the development of a treatment for the learning disabilities associated with NF1 and indicate that the GAP domain of NF1 modulates learning and memory. PMID- 11279522 TI - Trinucleotide expansion in haploid germ cells by gap repair. AB - Huntington disease (HD) is one of eight progressive neurodegenerative disorders in which the underlying mutation is a CAG expansion encoding a polyglutamine tract. The mechanism of trinucleotide expansion is poorly understood. Expansion is mediated by misaligned pairing of repeats and the inappropriate formation of DNA secondary structure as the duplex unpairs. It has never been clear, however, whether duplex unpairing occurs during mitotic replication or during strand-break repair. In simple organisms, trinucleotide expansion arises by replication slippage on either the leading or the lagging strand, homologous recombination, gene conversion, double-strand break repair and base excision repair; it is not clear which of these mechanisms is used in mammalian cells in vivo. We have followed heritable changes in CAG length in male transgenic mice. In germ cells, expansion is limited to the post-meiotic, haploid cell and therefore cannot involve mitotic replication or recombination between a homologous chromosome or a sister chromatid. Our data support a model in which expansion in the germ cells arises by gap repair and depends on a complex containing Msh2. Expansion occurs during gap-filling synthesis when DNA loops comprising the CAG trinucleotide repeats are sealed into the DNA strand. PMID- 11279523 TI - The amnionless gene, essential for mouse gastrulation, encodes a visceral endoderm-specific protein with an extracellular cysteine-rich domain. AB - Fate-mapping experiments in the mouse have revealed that the primitive streak can be divided into three functional regions: the proximal region gives rise to germ cells and the extra-embryonic mesoderm of the yolk sac; the distal region generates cardiac mesoderm and node-derived axial mesendoderm; and the middle streak region produces the paraxial, intermediate and lateral plate mesoderm of the trunk. To gain insight into the mechanisms that mediate the assembly of the primitive streak into these functional regions, we have cloned and functionally identified the gene disrupted in the amnionless (amn) mouse, which has a recessive, embryonic lethal mutation that interferes specifically with the formation and/or specification of the middle primitive streak region during gastrulation. Here we report that the gene Amn encodes a novel type I transmembrane protein that is expressed exclusively in the extra-embryonic visceral endoderm layer during gastrulation. The extracellular region of the Amn protein contains a cysteine-rich domain with similarity to bone morphogenetic protein (BMP)-binding cysteine-rich domains in chordin, its Drosophila melanogaster homolog (Short gastrulation) and procollagen IIA (ref. 3). Our findings indicate that Amn may direct the production of trunk mesoderm derived from the middle streak by acting in the underlying visceral endoderm to modulate a BMP signaling pathway. PMID- 11279524 TI - An alternative mode of translation permits production of a variant NBS1 protein from the common Nijmegen breakage syndrome allele. AB - Nijmegen breakage syndrome (NBS) is a rare chromosomal-instability syndrome associated with cancer predisposition, radiosensitivity and radioresistant DNA synthesis-S phase checkpoint deficiency, which results in the failure to suppress DNA replication origins following DNA damage. Approximately 90% of NBS patients are homozygous for the 657del5 allele, a truncating mutation of NBS1 that causes premature termination at codon 219. Because null mutations in MRE11 and RAD50, which encode binding partners of NBS1, are lethal in vertebrates, and mouse Nbs1 null mutants are inviable, we tested the hypothesis that the NBS1 657del5 mutation was a hypomorphic defect. We showed that NBS cells contain the predicted 26-kD amino-terminal protein fragment, NBS1p26, and a 70-kD NBS1 protein (NBS1p70) lacking the native N terminus. The NBSp26 protein is not physically associated with the MRE11 complex, whereas the p70 species is physically associated with it. NBS1p70 is produced by internal translation initiation within the NBS1 mRNA using an open reading frame generated by the 657del5 frameshift. We propose that the common NBS1 allele encodes a partially functional protein that diminishes the severity of the NBS phenotype. PMID- 11279525 TI - An abundance of X-linked genes expressed in spermatogonia. AB - Spermatogonia are the self-renewing, mitotic germ cells of the testis from which sperm arise by means of the differentiation pathway known as spermatogenesis. By contrast with hematopoietic and other mammalian stem-cell populations, which have been subjects of intense molecular genetic investigation, spermatogonia have remained largely unexplored at the molecular level. Here we describe a systematic search for genes expressed in mouse spermatogonia, but not in somatic tissues. We identified 25 genes (19 of which are novel) that are expressed in only male germ cells. Of the 25 genes, 3 are Y-linked and 10 are X-linked. If these genes had been distributed randomly in the genome, one would have expected zero to two of the genes to be X-linked. Our findings indicate that the X chromosome has a predominant role in pre-meiotic stages of mammalian spermatogenesis. We hypothesize that the X chromosome acquired this prominent role in male germ-cell development as it evolved from an ordinary, unspecialized autosome. PMID- 11279526 TI - TSLC1 is a tumor-suppressor gene in human non-small-cell lung cancer. AB - The existence of tumor-suppressor genes was originally demonstrated by functional complementation through whole-cell and microcell fusion. Transfer of chromosome 11 into a human non-small-cell lung cancer (NSCLC) cell line, A549, suppresses tumorigenicity. Loss of heterozygosity (LOH) on the long arm of chromosome 11 has been reported in NSCLC and other cancers. Several independent studies indicate that multiple tumor-suppressor genes are found in this region, including the gene PPP2R1B at 11q23-24 (ref. 7). Linkage studies of NSCLC are precluded because no hereditary forms are known. We previously identified a region of 700 kb on 11q23.2 that completely suppresses tumorigenicity of A549 human NSCLC cells. Most of this tumor-suppressor activity localizes to a 100-kb segment by functional complementation. Here we report that this region contains a single confirmed gene, TSLC1, whose expression is reduced or absent in A549 and several other NSCLC, hepatocellular carcinoma (HCC) and pancreatic cancer (PaC) cell lines. TSLC1 expression or suppression is correlated with promoter methylation state in these cell lines. Restoration of TSLC1 expression to normal or higher levels suppresses tumor formation by A549 cells in nude mice. Only 2 inactivating mutations of TSLC1 were discovered in 161 tumors and tumor cell lines, both among the 20 primary tumors with LOH for 11q23.2. Promoter methylation was observed in 15 of the other 18 primary NSCLC, HCC and PaC tumors with LOH for 11q23.2. Thus, attenuation of TSLC1 expression occurred in 85% of primary tumors with LOH. Hypermethylation of the TSLC1 promoter would seem to represent the 'second hit' in NSCLC with LOH. PMID- 11279527 TI - Dyssegmental dysplasia, Silverman-Handmaker type, is caused by functional null mutations of the perlecan gene. AB - Perlecan is a large heparan sulfate (HS) proteoglycan present in all basement membranes and in some other tissues such as cartilage, and is implicated in cell growth and differentiation. Mice lacking the perlecan gene (Hspg2) have a severe chondrodysplasia with dyssegmental ossification of the spine and show radiographic, clinical and chondro-osseous morphology similar to a lethal autosomal recessive disorder in humans termed dyssegmental dysplasia, Silverman Handmaker type (DDSH; MIM 224410). Here we report a homozygous, 89-bp duplication in exon 34 of HSPG2 in a pair of siblings with DDSH born to consanguineous parents, and heterozygous point mutations in the 5' donor site of intron 52 and in the middle of exon 73 in a third, unrelated patient, causing skipping of the entire exons 52 and 73 of the HSPG2 transcript, respectively. These mutations are predicted to cause a frameshift, resulting in a truncated protein core. The cartilage matrix from these patients stained poorly with antibody specific for perlecan, but there was staining of intracellular inclusion bodies. Biochemically, truncated perlecan was not secreted by the patient fibroblasts, but was degraded to smaller fragments within the cells. Thus, DDSH is caused by a functional null mutation of HSPG2. Our findings demonstrate the critical role of perlecan in cartilage development. PMID- 11279528 TI - Screening a large reference sample to identify very low frequency sequence variants: comparisons between two genes. AB - Most human sequence variation is in the form of single-nucleotide polymorphisms (SNPs). It has been proposed that coding-region SNPs (cSNPs) be used for direct association studies to determine the genetic basis of complex traits. The success of such studies depends on the frequency of disease-associated alleles, and their distribution in different ethnic populations. If disease-associated alleles are frequent in most populations, then direct genotyping of candidate variants could show robust associations in manageable study samples. This approach is less feasible if the genetic risk from a given candidate gene is due to many infrequent alleles. Previous studies of several genes demonstrated that most variants are relatively infrequent (<0.05). These surveys genotyped small samples (n<75) and thus had limited ability to identify rare alleles. Here we evaluate the prevalence and distribution of such rare alleles by genotyping an ethnically diverse reference sample that is more than six times larger than those used in previous studies (n=450). We screened for variants in the complete coding sequence and intron-exon junctions of two candidate genes for neuropsychiatric phenotypes: SLC6A4, encoding the serotonin transporter; and SLC18A2, encoding the vesicular monoamine transporter. Both genes have unique roles in neuronal transmission, and variants in either gene might be associated with neurobehavioral phenotypes. PMID- 11279529 TI - A major susceptibility locus for leprosy in India maps to chromosome 10p13. AB - Leprosy, a chronic infectious disease caused by Mycobacterium leprae, is prevalent in India, where about half of the world's estimated 800,000 cases occur. A role for the genetics of the host in variable susceptibility to leprosy has been indicated by familial clustering, twin studies, complex segregation analyses and human leukocyte antigen (HLA) association studies. We report here a genetic linkage scan of the genomes of 224 families from South India, containing 245 independent affected sibpairs with leprosy, mainly of the paucibacillary type. In a two-stage genome screen using 396 microsatellite markers, we found significant linkage (maximum lod score (MLS) = 4.09, P < 2x10-5) on chromosome 10p13 for a series of neighboring microsatellite markers, providing evidence for a major locus for this prevalent infectious disease. Thus, despite the polygenic nature of infectious disease susceptibility, some major, non-HLA-linked loci exist that may be mapped through obtainable numbers of affected sibling pairs. PMID- 11279530 TI - Neurodegeneration: diseases of the cytoskeleton? PMID- 11279531 TI - Inhibition of IAP's protection by Diablo/Smac: new therapeutic opportunities? PMID- 11279532 TI - The role of p53 in neuronal cell death. AB - The p53 tumor suppressor gene is a sequence-specific transcription factor that activates the expression of genes engaged in promoting growth arrest or cell death in response to genotoxic stress. A possible role for p53-related modulation of neuronal viability has been suggested by the finding that p53 expression is elevated in damaged neurons in acute models of injury such as ischemia and epilepsy and in brain tissue samples derived from patients with chronic neurodegenerative diseases. Moreover, the absence of p53 has been shown to protect neurons from a wide variety of acute toxic insults. Signal transduction pathways associated with p53-induced cell death are being unraveled and suggest that intervention may prove fruitful in maintaining neuronal viability and restoring function following cytopathic insults. PMID- 11279533 TI - Neuronal life and death: an essential role for the p53 family. AB - Recent evidence indicates that the p53 tumor suppressor protein, and its related family member, p73, play an essential role in regulating neuronal apoptosis in both the developing and injured, mature nervous system. In the developing nervous system, they do so by regulating naturally-occurring cell death in neural progenitor cells and in postmitotic neurons, acting to ensure the apoptosis of cells that either do not appropriately undergo the progenitor to postmitotic neuron transition, or that fail to compete for sufficient quantities of trophic support. Somewhat surprisingly, in developing postmitotic neurons, p53 plays a proapoptotic role, while a naturally-occurring, truncated form of p73, DeltaNp73, antagonizes p53 and plays an anti-apoptotic role. In the mature nervous system, numerous studies indicate that p53 is essential for the neuronal death in response to a variety of insults, including DNA damage, ischemia and excitotoxicity. It is likely that all of these insults culminate in DNA damage, which may well be a common trigger for neuronal apoptosis. In this regard, the signaling pathways that are responsible for triggering p53-dependent neuronal apoptosis are starting to be elucidated, and involve cell cycle deregulation and activation of the JNK pathway. Finally, accumulating evidence indicates that p53 is perturbed in the CNS in a number of neurodegenerative disorders, leading to the hypothesis that longterm oxidative damage and/or excitotoxicity ultimately trigger p53-dependent apoptosis in the chronically degenerating nervous system. PMID- 11279534 TI - Pathogenesis of prion diseases: a progress report. AB - Almost 20 years have passed since Stanley Prusiner proposed that the agent causing transmissible spongiform encephalopathies consists exclusively of a protein and termed it prion. A mixed balance can be drawn from the enormous research efforts that have gone into prion research during this time. On the negative side, the protein-only hypothesis has not been conclusively proven yet. On the positive side, our understanding of spongiform encephalopathies has experienced tremendous advances, mostly through human genetics, mouse transgenetics, and biophysical methods. Perhaps the most astonishing development is the realization that many human neurodegenerative diseases for which transmissibility has been more or less stringently excluded, may follow pathogenetic principles similar to those of prion diseases. Also, the hypothesis that prion-like phenomena may underlie certain non-genetic traits observed in yeast has resulted in the surprising recognition that the instructional self propagating changes in protein conformation may be much more prevalent in nature than previously thought. The latter developments have been astonishingly successful, and one could now argue that the prion principle is much more solidly established in yeast than in mammals. PMID- 11279535 TI - Bcl-2 prevents mitochondrial permeability transition and cytochrome c release via maintenance of reduced pyridine nucleotides. AB - Digitonin-permeabilized PC12 and GT1-7 neural cells exhibited a cyclosporin A sensitive decrease in mitochondrial membrane potential, increased volume, and release of the pro-apoptotic factor cytochrome c in the presence of Ca2+ and the mitochondrial permeability transition (MPT) inducers t-butyl hydroperoxide (t bOOH) or phenylarsine oxide (PhAsO). Although the concentration of PhAsO required to induce the MPT was similar for Bcl-2 negative and Bcl-2 overexpressing transfected cells (Bcl-2(+)), the level of t-bOOH necessary for triggering the MPT was much higher for Bcl-2(+) cells. A higher concentration of t-bOOH was also necessary for promoting the oxidation of mitochondrial pyridine nucleotides in Bcl-2(+) cells. The sensitivity of Bcl-2(- ) cell mitochondria to t-bOOH but not PhAsO could be overcome by the use of conditions that protect the pyridine nucleotides against oxidation. We conclude that the increased ability of Bcl-2(+) cells to maintain mitochondrial pyridine nucleotides in a reduced redox state is a sufficient explanation for their resistance to MPT under conditions of oxidative stress induced by Ca2+ plus t-bOOH. PMID- 11279536 TI - Treatment with annexin V increases immunogenicity of apoptotic human T-cells in Balb/c mice. AB - Exposure of phosphatidylserine on the outer leaflet of the cytoplasmic membrane is an early event during apoptotic cell death and serves as a recognition signal for phagocytes. Usually the clearance of apoptotic cells does not initiate inflammation or immune response. We investigated the immune response in Balb/c mice towards apoptotic human T-cells. Animals injected with apoptotic cells showed significantly reduced humoral immune responses, especially Th1-dependent IgG2a titres, compared to controls immunised with viable cells. However, treatment of apoptotic cells with annexin V (AxV) significantly increased the humoral immune response. AxV binds with high affinity to anionic phospholipids and as a result interferes with the phosphatidylserine recognition by phagocytes. Our results indicate that AxV treatment may be used to increase the efficiency of apoptotic cell-based vaccines, e.g. some tumour vaccines. PMID- 11279538 TI - Role of NAD(P)H oxidase in the tamoxifen-induced generation of reactive oxygen species and apoptosis in HepG2 human hepatoblastoma cells. AB - Previously, tamoxifen (TAM) has been shown to induce apoptosis through elevation of intracellular Ca2+ in HepG2 human hepatoblastoma cells. In this study we investigated the role of reactive oxygen species (ROS) in the TAM-induced apoptosis, and interrelationship between intracellular Ca2+ and ROS. TAM induced a slow and sustained increase in intracellular ROS level. An antioxidant, N acetylcysteine significantly inhibited both ROS production and apoptosis induced by TAM, suggesting that ROS may play an essential role in the TAM-induced apoptosis. In a time frame ROS generation followed intracellular Ca2+ increase, and the extracellular and intracellular Ca2+ chelation with EGTA and BAPTA/AM, respectively, completely inhibited the TAM-induced ROS production, indicating that intracellular Ca2+ may mediate the ROS generation. Inhibitors of NAD(P)H oxidase, diphenylene iodonium, phenylarsine oxide and neopterine, significantly blocked the TAM-induced ROS generation and apoptosis, implying that this oxidase may act as a source enzyme for the production of ROS. These results suggest that non-phagocytic NAD(P)H oxidase may play a novel role as a mediator of the apoptosis associated with intracellular Ca2+ in HepG2 cells. PMID- 11279537 TI - Erk-dependent cytosolic phospholipase A2 activity is induced by CD95 ligand cross linking in the mouse derived Sertoli cell line TM4 and is required to trigger apoptosis in CD95 bearing cells. AB - In the present study we demonstrated that CD95L cross-linking generated reverse signalling in the mouse derived Sertoli cell line TM4. Treatment of TM4 cells with mAb anti-CD95L induced activation of the cytosolic phospholipase A2 (cPLA2). Cytosolic PLA2 activation was controlled by the MAPK pathway as indicated by the ability of the specific MEK inhibitor, PD098059, to abolish cPLA2 activation. In addition, Western blot experiments showed a rapid increase in phosphorylated Erk1/2 following CD95L cross-linking, while no effect on the phosphorylation of other MAPK, p38 or JNK, was observed. CD95L cross-linking by mAb increased the levels of soluble CD95L and apoptotic activity of TM4 cell supernatants, which was blocked by co-incubation with the PLA2 inhibitor, AACOCF3 or PD098059. Finally, pre-treatment of TM4 cells with AACOCF3 or PD098059 completely abolished TM4-induced apoptosis of Jurkat T cells, thus indicating that the Erk/cPLA2 pathway is required for CD95L-induced apoptosis. PMID- 11279539 TI - Differential susceptibility to CD95 (Apo-1/Fas) and MHC class II-induced apoptosis during murine dendritic cell development. AB - Disappearance of antigen presenting cells (APC) from the lymph node occurs following antigen specific interactions with T cells. We have investigated the regulation of CD95 (Apo-1/Fas) induced apoptosis during murine dendritic cell (DC) development. Consistent with the moderate levels of CD95 surface expression and low, or absent, MHC class II expression, immature DC in bone marrow cultures were highly sensitive to CD95 induced apoptosis, but insensitive to class II mediated apoptosis. In contrast, mature splenic, epidermal and bone marrow derived DC were fully resistant to CD95 induced cell death, but sensitive to class II induced apoptosis. Although caspase 3 and 8 activation was detected in immature DC undergoing CD95L-induced apoptosis, the pan-caspase inhibitor zVAD fmk did not inhibit the early events of CD95-induced mitochondrial depolarisation or phosphatidyl serine exposure and only partially inhibited the killing of immature DC. In contrast, zVAD-fmk was completely effective in preventing CD95L mediated death of murine thymocytes. Collectively, these data do not support a major role of CD95: CD95L ligation in apoptosis of mature DC, but rather emphasise the existence of distinct pathways for the elimination of DC at different stages of maturation. PMID- 11279540 TI - Potential mechanisms of resistance to TRAIL/Apo2L-induced apoptosis in human promyelocytic leukemia HL-60 cells during granulocytic differentiation. AB - Human promyelocytic leukemia HL-60 cells are well known to differentiate into granulocytes or monocytes in the presence of some agents such as DMSO or PMA, respectively. Differentiated HL-60 cells become resistant to some apoptotic stimuli including anticancer drugs or irradiation though undifferentiated cells significantly respond to these stimuli. TRAIL (TNF-related apoptosis-inducing ligand) which is also known as Apo2 ligand (Apo2L), a new member of TNF family, can induce apoptosis in some tumor cells but not in many normal cells. We show here that apoptosis is well induced in HL-60 cells by TRAIL, but susceptibility to TRAIL is reduced during granulocytic differentiation by DMSO. We also suggest some possible mechanisms by which granulocytic differentiated cells become resistant to TRAIL-induced apoptosis. First, in granulocytic differentiated cells, expression of antagonistic decoy receptors for TRAIL (TRAIL R3/TRID/DcR1/LIT and TRAIL-R4/TRUNDD/DcR2) were enhanced. In addition, expression of Toso, a cell surface apoptosis regulator, seemed to block activation of caspase-8 by TRAIL via enhanced expression of FLIPL in granulocytic differentiated cells. These findings suggest that differentiated cells are resistant using plural mechanisms against various apoptosis-inducing stimuli rather than undifferentiated cells. PMID- 11279541 TI - Failure of Bcl-2 family members to interact with Apaf-1 in normal and apoptotic cells. AB - CED-9 blocks programmed cell death (apoptosis) in the nematode C. elegans by binding to and neutralizing CED-4, an essential activator of the aspartate directed cysteine protease (caspase) CED-3. In mammals, the CED-9 homologs Bcl-2 and Bcl-xL also block apoptosis by interfering with the activation of CED-3-like caspases. However, it is unknown whether this occurs by binding to the CED-4 homolog Apaf-1. Whilst two groups previously detected an interaction between Bcl xL and Apaf-1 in immunoprecipitates,1,2 another group found no interaction between Apaf-1 and any of ten individual members of the Bcl-2 family using the same experimental approach.3 In this study, we aimed to resolve this discrepancy by monitoring the binding of Apaf-1 to three Bcl-2 family members within cells. Using immunofluorescence and Western blot analysis, we show that whilst Apaf-1 is a predominantly cytoplasmic protein, Bcl-2, Bcl-xL and Bax mostly reside on nuclear/ER and mitochondrial membranes. This pattern of localization is maintained when the proteins are co-expressed in both normal and apoptotic cells, suggesting that Bcl-2, Bcl-xL or Bax do not significantly sequester cytoplasmic Apaf-1 to intracellular membranes. In addition, we confirm that Apaf-1 does not interact with Bcl-2 and Bcl-xL in immunoprecipitates. Based on these data, we propose that Apaf-1 is not a direct, physiological target of Bcl-2, Bcl-xL or Bax. PMID- 11279543 TI - Cytokine regulation of apoptosis and Bcl-2 expression in lymphocytes of patients with systemic lupus erythematosus. AB - Both faulty regulation of apoptosis and the inappropriate expression of several interleukins have been considered important defects of lymphocytes in the human autoimmune disease systemic lupus erythematosus (SLE). We therefore tested the in vitro effect of recombinant interleukin (IL-)-2, 4, 7, and 15 on peripheral blood mononuclear cells from patients with SLE and from healthy volunteers. Intracellular Bcl-2 and Bax expression was measured by fluorocytometry and the rate of apoptosis was determined by the TUNEL technique and propidium iodide staining. IL-2, IL-4, IL-7 and IL-15 led to a significant increase in Bcl-2 and a reduction in cell death rates, which was even more pronounced in SLE. Bax levels remained unchanged. Interestingly, the high ex vivo Bcl-2 content of lymphocytes from some SLE patients was maintained after growth factor withdrawal. Anti apoptotic cytokine signaling may significantly influence the deregulation of cell death in SLE lymphocytes. PMID- 11279542 TI - TNF down-regulation of receptor tyrosine kinase-dependent mitogenic signal pathways as an important step in cytostasis induction and commitment to apoptosis of Kym-1 rhabdomyosarcoma cells. AB - Growth of Kym-1 rhabdomyosarcoma cells depends on endogenous receptor tyrosine kinase signals activated by insulin and insulin-like growth factors (IGF), as revealed from enhancement of proliferation by insulin and IGF-1 and cytostatic action of inhibitors of IR/IGFR kinases. Depending on the presence or absence of the caspase inhibitor z-VAD-fmk, TNF induced full growth arrest or apoptosis, respectively, indicating dominance of TNF over mitogenic signal pathways in Kym-1 cells. In accordance with a caspase-independent cytostatic action, TNF downregulated IR kinase activity and caused a profound inhibition of downstream mitogenic signals including the MAPK cascade and STAT5, key pathways of proliferation and cell survival. Removal of z-VAD-fmk after 24 h induced rapid cell death in the absence of TNF. The inhibition of survival signals concomitant with persisting proapoptotic signals may tip the balance towards an irreversible commitment of the cell to apoptosis that becomes apparent upon relief of suppression of effector caspases. PMID- 11279544 TI - Dependence of granzyme B-mediated cell death on a pathway regulated by Bcl-2 or its viral homolog, BHRF1. AB - The molecular pathways responsible for apoptosis in response to granzyme B have remained unresolved. Here we present data supporting the notion that granzyme B mediated cell death is largely dependent on a pathway that is inhibitable by Bcl 2 or its viral analog BHRF1. We used a panel of stably transfected FDC-P1 mouse myeloid cell lines to show that overexpression of functional, wild-type Bcl-2 or BHRF1 rescued cells from granzyme B-mediated apoptosis, whereas mutated (Gly145- >Glu) Bcl-2, or wild-type Bcl-2 directed to the plasma membrane conferred no protection. Overexpression of Bcl-2 resulted in inhibition of multiple parameters of apoptosis in response to purified perforin and granzyme B, including DNA fragmentation, changes in light scatter profile indicating cell shrinkage and increased refractivity, loss of mitochondrial membrane potential and inhibited colony formation in clonogenic assays. Nevertheless, when exposed to cytotoxic lymphocytes, FDC-P1 and YAC-1 cells overexpressing Bcl-2 remained susceptible to death imparted by cytolytic granules, irrespective of whether the granules contained granzyme B. Thus, alternative granzyme B-independent pathways can be activated by intact lymphocytes to overcome Bcl-2-like inhibitors of apoptosis, enabling CTLs to overcome potential viral blocks to granzyme B-mediated cell death. PMID- 11279545 TI - Caspase 6 activity initiates caspase 3 activation in cerebellar granule cell apoptosis. AB - Using a well documented ex vivo system consisting of rodent cerebellar granule cells (CGCs) the activation of caspases 3 and 6 during apoptosis induced by withdrawal of trophic support was analyzed. At the time of deprivation, the addition of the irreversible, broad-spectrum caspase inhibitor zVADfmk or the cell permeable, caspase 6 inhibitor CP-VEID-cho can transiently suppress the appearance of apoptosis, including the early appearance of DNA fragmentation. Using immunoblotting and fluorogenic peptide assays we observe deprivation induced activation of caspases 3 and 6, but not caspase 9. Furthermore, active caspase 6 is capable of processing and activating procaspase 3 in cellular extracts prepared from non-apoptotic CGCs, whereas caspase 3 failed to activate caspase 6. In consonant with this, the cell permeable caspase 6 inhibitor prevented deprivation-induced caspase 3 activation whereas a cell permeable caspase 3 inhibitor, CP-DEVD-cho, had no effect on caspase 6 activation. This would indicate that caspase 6 is a significant inducer of the early caspase 3 activity in apoptotic CGCs. PMID- 11279548 TI - Differentiate or die: the view from Montreal. PMID- 11279547 TI - Apoptosis induced by dithiothreitol in HL-60 cells shows early activation of caspase 3 and is independent of mitochondria. AB - Previous studies have shown that under certain conditions some thiol-containing compounds can cause apoptosis in a number of different cell lines. Herein, we investigated the apoptotic pathways in HL-60 cells triggered by dithiothreitol (DTT), used as a model thiol compound, and tested the hypothesis that thiols cause apoptosis via production of hydrogen peroxide (H2O2) during thiol oxidation. The results show that, unlike H2O2, DTT does not induce apoptosis via a mitochondrial pathway. This is demonstrated by the absence of early cytochrome c release from mitochondria into the cytosol, the lack of mitochondrial membrane depolarization at early times, and the minor role of caspase 9 in DTT-induced apoptosis. The first caspase activity detectable in DTT-treated cells is caspase 3, which is increased significantly 1 - 2 h after the start of DTT treatment. This was shown by following the cleavage of both a natural substrate, DFF 45/ICAD, and a synthetic fluorescent substrate, z-DEVD-AFC. Cleavage of substrates of caspases 2 and 8, known as initiator caspases, does not start until 3 - 4 h after DTT exposure, well after caspase 3 has become active and at a time when apoptosis is in late stages, as shown by the occurrence of DNA fragmentation to oligonucleosomal-sized pieces. Although oxidizing DTT can produce H2O2, data presented here indicate that DTT-induced apoptosis is not mediated by production of H2O2 and occurs via a novel pathway that involves activation of caspase 3 at early stages, prior to activation of the common 'initiator' caspases 2, 8 and 9. PMID- 11279546 TI - 28S ribosome degradation in lymphoid cell apoptosis: evidence for caspase and Bcl 2-dependent and -independent pathways. AB - Apoptosis, a physiological form of cell death, is characterized by the activation of a program that kills cells and recycles their constituents. We have used thymoma cell lines to examine the role of Bcl-2 and caspases in ribosomal destruction during apoptosis. Glucocorticoid- and calcium ionophore (A23187) induced apoptosis of S49 Neo cells resulted in both 28S rRNA and DNA degradation. Interestingly, anisomycin, a potent protein synthesis inhibitor, also induced 28S rRNA and DNA fragmentation suggesting that the responsible nucleases are present in the viable cells and become activated during apoptosis. The anti-apoptotic protein, Bcl-2, inhibited both glucocorticoid- and anisomycin-induced DNA and 28S rRNA degradation but could not protect against A23187-induced nucleic acid degradation. We next examined the role of caspase activation in the generation of 28S rRNA degradation through the use of ZVAD, a general caspase inhibitor. Under conditions where ZVAD substantially decreased 28S rRNA degradation induced by glucocorticoid or anisomycin, no decrease was observed when A23187 was used to induce apoptosis. Surprisingly, RNA degradation, like DNA degradation, occurs exclusively in shrunken lymphocytes but not those with normal cell volume despite equivalent exposure of the cells to the apoptotic signals. Together, these findings indicate the ribosome is a specific target for death effectors during apoptosis and that a caspase/Bcl-2-independent pathway exists to activate its destruction. PMID- 11279549 TI - Nitric oxide-mediated redox reactions in pathology, biochemistry and medicine. PMID- 11279550 TI - Alcohol as a risk factor for HIV transmission among American Indian and Alaska Native drug users. AB - Quantitative alcohol interviews conducted as part of the National Institute on Drug Abuse (NIDA) Native American Supplement revealed very high rates of alcohol use among American Indian and Alaska Native active crack and injection drug users (IDUs). Of 147 respondents who completed the alcohol questionnaire, 100& percent had drunk alcohol within the past month, almost 42& percent reported that they drank every day, and 50& percent drank until they were drunk one-half of the time or more. Injection drug users (IDUs) demonstrated the highest frequency and quantity of alcohol use in the past 30 days. A significant positive association was also found between crack and alcohol use in the past 48 hours (c(2)=5.30, p<.05). Finally, those claiming more episodes of using alcohol before or during sex, reported significantly more events of unprotected sexual intercourse. Qualitative data from all four sites corroborated these quantitative findings. Many individuals also reported episodes of blacking out while drinking, and learned later that they had had unprotected sex with complete strangers or individuals they would not otherwise accept as partners. Implications of these findings for HIV/AIDS prevention efforts are addressed. PMID- 11279551 TI - Unemployment, drug use, and HIV risk among American Indian and Alaska Native drug users. AB - American Indians and Alaska Natives have had low employment in recent history. Drug users also have low employment due to cycles of drug use and relapse,and the impact of the type of drug abused on levels of functioning. Drug use is associated with increased HIV risk through injection drug use, frequency of injection, and needle sharing. Data from three sites of the NIDA Cooperative Agreement for Community Based-Outreach/Intervention Research were analyzed to determine the relationship among race/ethnicity, age, and level of educational attainment on employment and unemployment at intake interview and six-month follow-up. HIV risk for those employed and unemployed was then assessed. American Indian and Alaska Native drug users were younger, less educated, and less likely to have a paid job at both intake and follow-up than non-Native drug users. Those participants who were unemployed at baseline interview who were American Indian/Alaska Native were less likely to transition to employment at six-month follow-up than other race/ethnicity groups in the cohort. However, all participants showed low levels of employment at follow-up. Individuals who were employed at baseline and those who transitioned to employment had lower levels of injection drug use and needle sharing than those who were unemployed at both baseline and follow-up. American Indian and Alaska Native drug users may be at risk for acquisition of HIV due to drug risk behaviors that appear to be associated with unemployment. PMID- 11279552 TI - HIV drug and sex risk behaviors among American Indian and Alaska Native drug users: gender and site differences. AB - Little research has been conducted on HIV drug and sex risk behaviors of American Indians and Alaska Natives who use illicit drugs. Data from studies conducted with other ethnic groups indicates differences in HIV drug and sex risk behaviors of men and women and between drug users from different regions, cities, communities, and intervention sites. This study examines whether these differences in HIV drug and sex risk behaviors also exist for American Indians and Alaska Natives. Results indicate that risk behaviors of American Indians and Alaska Natives do differ like that of other ethnic groups. In particular American Indian and Alaska Native women reported engaging in significantly greater levels of some drug and many sex risk behaviors than men. Significant differences between intervention sites were also found for intensity of use of various drugs and for some HIV drug risk behaviors. PMID- 11279553 TI - Alaska Native drug users and sexually transmitted disease: results of a five-year study. AB - Although Alaska has one of the highest rates of alcohol consumption in the U.S., there are very few reports of other drug use in Alaska. This five-year NIDA funded study sampled out-of-treatment injection drug users (IDUs) and crack cocaine smokers in Anchorage, Alaska. This paper is a summary of results comparing risk behavior for HIV and sexually transmitted disease infection among Alaska Natives (n=216) to non-Natives (primarily Blacks n=394 and Whites n=479) from this study. IDUs and crack cocaine smokers were recruited using a targeted sampling plan. All subjects tested positive to cocaine metabolites, or morphine, using urinalysis, or had visible track marks. Several analyses of this database have indicated that Alaska Native women are at high risk for gonorrhea infection. They are also at risk for HIV infection due to high rates of behavior related to blood-borne disease transmission. We have also found that White men who have sex with both White and Alaska Native women are significantly less likely to use condoms with the Alaska Native women. HIV preventive education efforts aimed at Alaska Native women need to be implemented on a major scale. PMID- 11279554 TI - HIV and AIDS among American Indians and Alaska Natives. PMID- 11279555 TI - Patterns and predictors of HIV risk among urban American Indians. AB - A preliminary survey of HIV risk and service preferences among American Indians residing in the New York metropolitan area included 68 women and 32 men (M age=35.8 years). Overall, the sample was knowledgeable about the mechanisms of HIV transmission, and 58 percent reported having taken an HIV test. However, of the 63 percent who reported sexual activity in the last six months, 73 percent reported engaging in vaginal or anal sex without a condom with at least 1 partner, and 52 percent used condoms none of the time during vaginal and anal sex. Almost half (43 percent) reported alcohol or other drug (AOD) use for non ceremonial purposes in the last six months. Alarmingly, 44 percent reported lifetime trauma, including domestic violence (20 percent) and physical (29 percent) or sexual (26 percent) assault by a family member or stranger. Bivariate and multivariate analyses indicated trauma and drug use were factors that may place respondents at risk for sexual transmission of HIV. Trauma variables were better predictors of HIV risk behaviors than social cognitive variables providing preliminary support for the use of a postcolonial framework in American Indian HIV studies. PMID- 11279556 TI - American Indians with HIV/AIDS: health and social service needs, barriers to care, and satisfaction with services among a Western tribe. AB - This study investigated the health care and social service needs, barriers to care and satisfaction with services among American Indians with HIV/AIDS in a western tribe. Individual interviews were conducted with 28 respondents, which constituted nearly the entire population obtaining HIV/AIDS medical services from the IHS in the target area. The survey found that expressed need for services in this frontier rural area were lower than urban counterparts, but that access to needed services was lower. Common unmet medical needs include mental health services, eye and dental care, traditional Native medicine, and substance abuse treatment. Common unmet social service needs include housing assistance, help obtaining food and clothing, and transportation. Limited access to essential services impedes the ability of American Indians with HIV/AIDS to maintain effective medical regimens. PMID- 11279557 TI - The Ahalaya case-management program for HIV-infected American Indians, Alaska Natives, and Native Hawaiians: quantitative and qualitative evaluation of impacts. AB - The Ahalaya case management model was designed to provide culturally sensitive services to HIV-positive American Indians (AI), Alaska Natives (AN), and Native Hawaiians (NH). This program started in 1991 and expanded across the country in 1994. The evaluation plan included a client satisfaction survey, along with focus groups and key informant interviews. Of the 389 active clients enrolled, 132 responded to the anonymous 35-item questionnaire. Responses were favorable regarding benefits of the programs. Self-reported quality of life changes after enrollment also were significantly improved (Wilcoxon Signed Rank Test: T=6.87, p=.000; n=131). Qualitative data highlighted other important issues. Social relationships-with staff, community, and family-were critical to client welfare, as a source of both strength and fear. While AI/AN/NH case management programs have been shown effective, services need to expand, and they have to facilitate resolutions to problems in clients social relationships. PMID- 11279558 TI - Unmasking Dashkayah: storytelling and HIV prevention. PMID- 11279559 TI - Respondent bias in the collection of alcohol and tobacco data in American Indians: the Strong Heart Study. AB - This study addresses the impact of assessment method (interviewer-administered questionnaire vs. self-administered questionnaire) and interviewers demographic characteristics (gender, ethnicity, and residency) on responses to alcohol and tobacco questions. The study population included 1,522 men and women aged 45 to 74 from the Dakota Center of the Strong Heart Study (SHS), a multi-center study of cardiovascular disease in American Indians. Assessment method effects were greater for alcohol than tobacco but did not differ by interviewer characteristics. PMID- 11279560 TI - Recruitment of American Indians in epidemiologic research: the Strong Heart Study. AB - This paper describes the methods used to recruit American Indian (AI) populations for the Strong Heart Study (SHS), a community-based study of cardiovascular disease (CVD) and its risk factors in AI men and women. Recruitment strategies included personal contact by recruiters and drivers/recruiters in remote areas, SHS staff participation in community activities, and mass media. A total of 4,549 participants aged 45-74 years were recruited from 13 American Indian tribes and communities. Overall participation rates were 72&percnt, 55&percnt, and 62&percnt, respectively, for the three study centers (Arizona, the Dakotas, and Oklahoma). Participant feedback and educational material related to risk factor reduction and promoting a healthy lifestyle were emphasized. Participants were likely to be female, young, and nonsmokers. Barriers to recruitment included lack of telephones in a large proportion of households, conflicting beliefs about health/health care/research, fears, taboos, and occasional rumors about study examination procedures. Participants were referred for follow-up of health problems detected by the study. The strong commitment of the participating communities helped to insure the success of the SHS, which can be considered a model for recruitment in future American Indian population-based studies. Success was facilitated by the use of a variety of recruitment techniques. PMID- 11279561 TI - Prenatal alcohol use among urban American Indian/Alaska Native women. AB - This paper examines prenatal drinking among American Indian/Alaska Native women using the 1988 Urban Indian Over-sample for the National Maternal and Infant Health Survey. Using univariate, bivariate, and multivariate analyses, alcohol consumption during pregnancy was examined by demographic and behavioral variables. Although one out of every five American Indian/Alaska Native women consumed some amount of alcohol during pregnancy, those who used alcohol drank less than one drink per month. PMID- 11279562 TI - Bicultural resynthesis: Tailoring an effectiveness trial for a group of urban American Indian women. AB - The purpose of this qualitative study of a 6-week effectiveness trial was to describe among a group of urban American Indian women, the process of successful traditionalism in the form of bicultural resynthesis. Bicultural resynthesis represents a major current attempt on the part of the participants to integrate traditional and contemporary demands in a positive, culturally-consistent manner. The themes of shame and isolation, adapting to survive, deculturation, ethnic switching/renewal, and bicultural resynthesis are discussed. Further support is achieved for retraditionalization of American Indian women s roles as an effective means of achieving American Indian self-determination and as a potential way of helping women overcome problems. PMID- 11279563 TI - Hand transplantation. PMID- 11279564 TI - Different reference frames can lead to different hand transplantation decisions by patients and physicians. AB - Different frames of reference can affect one's assessment of the value of hand transplantation. This can result in different yet rational decisions by different groups of individuals, especially patients and physicians. In addition, factors other than frames of reference can affect one's evaluation of hand transplantation, which can result in different decisions. PMID- 11279565 TI - Surgical management of delayed union and nonunion of distal radius fractures. AB - Ten patients with malaligned fractures of the distal radius that demonstrated either delayed healing or the development of an atrophic or synovial nonunion on standard radiographs were treated with surgical realignment, stable internal fixation, and autogenous iliac crest bone grafting. All 10 fractures healed with acceptable radiologic alignment within 3 months of the index procedure. After an average follow-up period of 3 years 6 months (range, 2 years to 8 years 6 months) patients had an average of 105 degrees wrist flexion and extension, 145 degrees forearm rotation, and 73% grip strength compared with the opposite limb. In the treatment of malaligned, ununited fractures of the distal radius, specific techniques and implants must be tailored to the deformity of the distal radius and the shape of the distal fragment. A stable, well-aligned wrist with preservation of at least 50 degrees mobility in flexion and extension was achieved in every patient, but the final result was compromised by associated problems in 3 patients. PMID- 11279566 TI - Ilizarov hybrid external fixation for fractures of the distal radius: Part I. Feasibility of transfixion wire placement. AB - The advantages of Ilizarov external fixation, allowing early motion of adjacent joints during fixation of periarticular fractures, have not yet been applied to distal radius fractures. A magnetic resonance imaging study of 10 normal volunteers evaluated the safety of passing percutaneous transfixion pins across the distal radius in 3 forearm positions. Even in the optimal forearm position, the safe zones between the transfixion pin, vessel, nerve, or tendon was small, suggesting that open placement would be required. A cadaver study in 8 specimens demonstrated that the pins could be placed with an open technique using an aiming device and that the pins could be placed without limiting forearm rotation. The proximity of vital structures to transfixion pins dictates open placement to safely apply Ilizarov fixation to distal radius fractures. PMID- 11279567 TI - Ilizarov hybrid external fixation for fractures of the distal radius: Part II. Internal fixation versus Ilizarov hybrid external fixation: Stability as assessed by cadaveric simulated motion testing. AB - The in vitro stability of an Ilizarov hybrid external fixator was compared with that of a dorsal 3.5-mm AO T-plate in 8 unpaired, fresh-frozen upper extremities. A specially designed testing device that used computer-controlled pneumatic actuators was used to simulate active finger, wrist, and forearm motions by applying loads to relevant tendons. A comminuted extra-articular distal radius fracture was modelled using a dorsally based wedge osteotomy. Fracture stability was assessed using an electromagnetic tracking device to measure motion across the fracture site after randomized application of the plate and the hybrid fixator. During simulated finger and wrist motions with the forearm pronated or supinated, motion of the distal fragment with the hybrid fixator applied was comparable to or statistically less than with the AO plate applied. During simulated forearm rotation, the stability provided by the 2 fixation types was similar, although the plate allowed statistically less radial-ulnar deviation of the fragment. In this model of a 2-part extra-articular distal radius fracture, the clinically meaningful stability of the Ilizarov hybrid external fixator was comparable to that of the dorsal AO plate. PMID- 11279568 TI - Galeazzi fracture-dislocation: a new treatment-oriented classification. AB - Forty patients with Galeazzi fracture-dislocations were treated with open reduction and internal fixation of the radial shaft fracture. Intraoperative distal radioulnar joint (DRUJ) instability after anatomic reduction was managed with supplemental wire transfixion of the DRUJ (10 patients) or open reduction and triangular fibrocartilage complex repair (3 patients). Two patterns of fracture-dislocation were identified based on the location of the radial shaft fracture. Twenty-two type I fractures were in the distal third of the radius within 7.5 cm of the midarticular surface of the distal radius; 12 of these cases were associated with intraoperative DRUJ instability. Eighteen type II fractures were in the middle third of the radial shaft more than 7.5 cm from the midarticular surface of the distal radius. Only one of these fractures had intraoperative DRUJ instability after open reduction and internal fixation of the radial shaft fracture. A high index of suspicion, early recognition, and acute treatment of DRUJ instability will avoid chronic problems in this complex injury. PMID- 11279569 TI - The radioulnar ratio: a new method of quantifying distal radioulnar joint subluxation. AB - Computed tomography was used to image the distal radioulnar joint (DRUJ) for instability. Four methods were used to quantify subluxation of the DRUJ: the Mino criteria, the epicenter method, the congruency method, and a new method called the radioulnar ratio (RUR). Validity of the various methods was evaluated in clinical and laboratory situations. Rheumatoid patients with symptomatic DRUJ pathology had significantly more abnormal RUR values (100% vs 73% [epicenter method] and 88% [Mino criteria]). The RUR detected instability sooner in a progressive laboratory-induced instability model. The intraobserver and interobserver reliability of the RUR was high, with intraclass correlation coefficients of 0.89 and 0.87, respectively. The RUR demonstrated superior performance in the diagnosis of DRUJ subluxation. PMID- 11279570 TI - Evaluation of the sigmoid notch with computed tomography following intra articular distal radius fracture. AB - A classification system for disruption patterns of the sigmoid notch of the radius associated with distal radius fractures has not been established. Using plain x-rays and corresponding computed tomography (CT) scans we characterized and quantified the types of sigmoid notch involvement in 20 consecutive distal radius fractures with radiocarpal joint extension. Plain radiographs revealed fracture extension into the sigmoid notch in only 7 cases (35%) and the CT scans demonstrated fracture extension into the sigmoid notch in 13 cases (65%). Of the 13 fractures with sigmoid notch involvement, 9 (69%) were displaced and 4 (31%) were nondisplaced. Sigmoid notch articular step-off (n = 7) and gapping (n = 9) were detectable on the CT scans but not on the x-rays. Plain x-rays appear to underestimate sigmoid notch involvement following distal radius fractures. In addition, CT appears to be a superior diagnostic modality for quantifying sigmoid notch fracture step-off and articular gapping. PMID- 11279571 TI - The surgical treatment of Kienbock's disease by radius and ulna metaphyseal core decompression. AB - We present a new surgical procedure, metaphyseal core decompression, to treat Kienbock's disease. Twenty-two patients were treated between 1976 and 1988 and were retrospectively reviewed. Sixteen were male and 6 female with an average age of 36 years (range, 18-64 years). The surgical technique involved curettage of the distal radius and ulna metaphyseal bone through small cortical windows. The average follow-up period was 10 years (range, 6-16 years). No postoperative complications were noted and no patient underwent additional surgical procedure. Sixteen patients were pain-free; 4 noted occasional pain. Twenty returned to their prior occupation. One patient had moderate pain and altered his occupation. Another had increasing pain and had x-ray evidence of advanced intercarpal arthritis. The average arc of wrist extension and flexion was 77% of the opposite wrist and the average grip strength was 75%. Long-term follow-up monitoring indicates that the metaphyseal core decompression produces results at least as good as other surgical procedures without any postoperative complications. PMID- 11279572 TI - Minnaar type 1 symptomatic lunotriquetral coalition: a report of nine patients. AB - Coalition of carpal bones is relatively common. It most frequently involves the lunate and triquetrum. This finding is almost always coincidental and asymptomatic. The fusion also can be incomplete, however, resembling a pseudarthrosis, and these patients can become symptomatic. To the best of our knowledge only 6 patients with a symptomatic Minnaar type 1 lunotriquetral coalition have been described in the literature. We present an additional nine symptomatic patients (12 wrists) with this type of coalition. A lunotriquetral arthrodesis was performed in 5 cases. In 3 cases a proximal row carpectomy was performed. PMID- 11279573 TI - Open reduction and internal fixation of acute displaced scaphoid waist fractures. AB - Fourteen consecutive patients with acute displaced scaphoid waist fractures were treated with open reduction and internal fixation. The operative technique consisted of anatomic reduction of the displaced scaphoid waist fracture, correction of carpal instability, radial bone grafting for comminution, and internal fixation with K-wires or Herbert screw. The patients were evaluated an average of 26 months (range, 4-48 months) after surgery. Thirteen of the 14 (93%) fractures united. The average time to union was 11.5 weeks (range, 8-20 weeks). Fracture union was confirmed with trispiral tomography. Final radiographic assessment consistently revealed a healed scaphoid fracture, restored intrascaphoid alignment, and no evidence of carpal instability. All patients regained functional wrist range of motion (wrist extension, 57 degrees; wrist flexion, 52 degrees ) and grip strength. Open reduction and internal fixation of acute displaced scaphoid waist fractures restores scaphoid alignment and leads to predictable union. Early operative intervention avoids malunion and carpal instability that often occurs with closed management of these complex fractures. PMID- 11279575 TI - The microvasculature of the nail bed, nail matrix, and nail fold of a normal human fingertip. AB - The organization of the microvasculature of the dorsal human fingertip based on a vascular corrosion cast was examined using a stereoscopic microscope. The variations of the superficial capillary network of the 3 specialized areas of skin of the dorsal fingertip (the nail bed, the nail matrix, and the nail fold) are described. In the nail bed numerous capillary loops were observed arising from a deeper regular arrangement of sagittally aligned, parallel rows of vessels. The size and direction of inclination of the capillary loops varied, getting longer and more inclined to the nail bed distally, with the longest capillary loops seen at the hyponychium. There were no capillary loops at the nail matrix region, but there was a single, layered, rectangular plexus of capillaries in the plane of the nail matrix. This extended distally to sagittally stretched coils of vessels that straightened out as the nail matrix enters the nail bed region. At the edge of the proximal nail fold the capillary loops looked like fine bristles and were approximately 3 times shorter than those found on the nail bed and hyponychium. This study provides a baseline for future work in understanding the changes in the microvasculature of the dorsal fingertip due to injury or pathology. PMID- 11279574 TI - Limits and indications of the dorsal transposition flap: critical evaluation of 15 cases. AB - The dorsal transposition flap was used in 15 cases of distal fingertip amputation. The amputations were either oblique ulnar, oblique radial, oblique palmar, or transverse. Two flaps developed partial necrosis. Flat nail growth always occurred, but the nail was considered short in 4 cases due to a proximal amputation through the nail bed. The mean 2-point discrimination test result was 8 mm. Distal interphalangeal joint range of motion was normal in 5 cases. Four cases lacked 10 degrees from full extension, 9 cases lacked up to 20 degrees from full flexion, and 1 case lacked 35 degrees. Even if this procedure is simple, reliable, and fast, due to its limited size and arc of rotation, as well as its poor sensibility, this flap may only be indicated for oblique ulnar fingertip amputations. PMID- 11279576 TI - An unusual presentation of a linear epidermal nevus. AB - We report an unusual presentation of a linear epidermal nevus in an adolescent male. Epidermal nevi most commonly appear in infancy and early childhood. They often are found in association with other organ system anomalies. We describe a palmar linear epidermal nevus that caused impairment of the patient's use of his hand. Therapeutic management involved surgical excision of the nevus and reconstruction of the area with a full-thickness skin graft. PMID- 11279577 TI - Treatment of recurrent compressive neuropathy of peripheral nerves in the upper extremity with an autologous vein insulator. AB - The treatment of entrapment neuropathy in the upper extremity with surgical decompression has generally provided good results. Recurrence of symptoms, however, is not uncommon and its management is both challenging and difficult. Nineteen patients with recurrent carpal tunnel and cubital tunnel syndrome were treated with the vein wrapping technique using the autogenous saphenous vein. The average number of surgeries before vein wrapping was 3.3. The mean patient age was 53 years (range, 28-75 years) and the mean follow-up period was 43 months (range, 24-78 months). All patients reported reduction in pain and the sensory disturbances secondary to the compression of the median or ulnar nerve. Two-point discrimination and electrodiagnostic findings also improved. PMID- 11279578 TI - Results of nerve transfer techniques for restoration of shoulder and elbow function in the context of a meta-analysis of the English literature. AB - We report the results of 15 patients who underwent nerve transfer for restoration of shoulder and elbow function at our institution for traumatic brachial plexus palsy. We present these results in the context of a meta-analysis of the English literature, designed to quantitatively assess the efficacy of individual nerve transfers for restoration of elbow and shoulder function in a large number of patients. One thousand eighty-eight nerve transfers from 27 studies met the inclusion criteria of the analysis. Seventy-two percent of direct intercostal to musculocutaneous transfers (without interposition nerve grafts) achieved biceps strength > or =M3 versus 47% using interposition grafts. Direct intercostal transfers to the musculocutaneous nerve had a better ability to achieve > or =M4 elbow strength than transfers from the spinal accessory nerve (41% vs 29%). The suprascapular nerve fared significantly better than the axillary nerve in obtaining > or =M3 shoulder abduction (92% vs 69%). At our institution 90% of intercostal to musculocutaneous transfers (n = 10) achieved > or =M3 bicep strength and 70% achieved > or =M4 strength. Four of seven patients achieved > or =M3 shoulder abduction with a single nerve transfer and 6 of 7 regained > or =M3 strength with a dual nerve transfer. This study suggests that interposition nerve grafts should be avoided when possible when performing nerve transfers. Better results for restoration of elbow flexion have been attained with intercostal to musculocutaneous transfers than with spinal accessory nerve transfers and spinal accessory to suprascapular transfers appear to have the best outcomes for return of shoulder abduction. We conclude that nerve transfer is an effective means to restore elbow and shoulder function in brachial plexus paralysis. PMID- 11279579 TI - Using intact nerve to bridge peripheral nerve defects: an alternative to the use of nerve grafts. AB - This preliminary study was conducted to determine whether a regenerating peripheral nerve in a rat model can use the epineurium of an intact nerve to bridge a nerve gap defect. To create the intact nerve bridge a 1-cm segment of the peroneal nerve is resected leaving a gap defect. The proximal and distal peroneal nerve stumps are sutured 1-cm apart, in an end-to-side fashion, to the epineurium of the intact tibial nerve. The following experimental groups were used (n = 12): group A, immediate primary repair of resected segment; group B, intact nerve bridge technique; group C, nerve autograft; and group D, gap in situ control. Evaluation 12 weeks after surgery included measurement of the tibialis anterior muscle contraction force, axonal counting, wet weight of the tibialis anterior muscle, and histologic examination. The results of this animal study support 3 main conclusions: regenerating axons can use the epineurium of an intact nerve to bridge a gap in nerve continuity; when using functional recovery to assess regeneration, there is no significant difference between standard nerve autografts and the intact nerve bridge technique; and based on histologic examination, the intact nerve bridge technique does not injure the intact tibial nerve used to bridge the gap defect. Taken together, the results of this preliminary animal study suggest that the intact nerve bridge technique may be a potential alternative to standard nerve autografts in appropriate circumstances. Further investigation in a higher animal model is warranted before considering clinical application of the intact nerve bridge technique. PMID- 11279580 TI - Comparative study of different surgical procedures using sensory nerves or neurons for delaying atrophy of denervated skeletal muscle. AB - To observe the effect of sensory components on muscle atrophy, 4 surgical procedures applying sensory nerves or neurons to the denervated muscle were conducted in a rat model: sensory nerve implantation (group B), sensory motor nerve crossover (group C), dorsal root ganglia implantation (group D), and implantation of preganglionically avulsed sensory nerve (group E). Rats with complete denervation served as controls (group A). The degree of muscle atrophy was evaluated after surgery by fibrillation potential amplitude, muscle wet weight, muscle fiber cross-sectional area, collagen content, and protein content. Compared with group A, the results in groups B, D, and E were superior 1 month after surgery and the results in groups B, C, and E were superior 3 months after surgery. Implantation of normal or preganglionically avulsed sensory nerve delayed atrophy. Dorsal root ganglia implantation only had a short-term trophic influence. In the animal model setting, sensory motor nerve crossover is effective only after it is maintained for at least 3 months. PMID- 11279581 TI - Effects of human amniotic fluid on peritendinous adhesion formation and tendon healing after flexor tendon surgery in rabbits. AB - The effect of the topical application of human amniotic fluid (HAF) on peritendinous adhesion formation and tendon healing was investigated in 32 New Zealand adult rabbits. The long flexor tendons of the digits of each hind paw were completely divided and repaired with a modified Kessler technique. The rabbits were randomly divided into 4 experimental groups according to the type of repair used: sheath excision, sheath excision and local HAF application, sheath repair, and sheath repair and local HAF application. The extent of adhesions and the healing status of the tendons were macroscopically and histologically evaluated at 12 weeks. Tensile strength of the repaired tendons was measured biomechanically at 20 weeks. The least adhesion and the best healing were observed in tendons treated with sheath repair and HAF application. Tendons treated with HAF had significantly higher tensile load values. Topical application of HAF immediately after tenorrhaphy is significantly effective in preventing peritendinous adhesion formation without impairment of tendon healing in this rabbit model. PMID- 11279582 TI - Effect of suture locking and suture caliber on fatigue strength of flexor tendon repairs. AB - The objectives of this cadaveric study were 2-fold: to determine the effect of different locking configurations on the cyclical fatigue strength of flexor tendon repairs and to assess the differences between each repair when a 3-0 or 4 0 suture is used. One hundred twenty flexor digitorum profundus tendons were cut and repaired using nonlocked, simple locked, and cross-stitch locked variations of 2- and 4-strand flexor tendon repairs. Using an incremental cyclical loading protocol we performed 10 trials of each repair with both 3-0 and 4-0 sutures and analyzed the number of Newton-cycles to failure using a 3-way ANOVA. The use of a 3-0 suture led to a 2- to 3-fold increase in fatigue strength in all repairs tested and the fatigue strength of the 4-strand repairs was significantly greater than the 2-strand repairs. All repairs performed with 4-0 suture failed by suture rupture. Of the 3-0 suture repairs, the three 2-strand repairs and the 4-strand cross-stitch locked repair failed by suture rupture. In contrast, 6 of 10 of the 4-strand simple locked and nonlocked repairs failed by suture pullout. There was no significant difference in fatigue strength between the 2 locked and the nonlocked 2-strand repairs using either 3-0 or 4-0 suture. There also was no significant difference in holding capacity or fatigue strength between the simple locked or nonlocked 4-strand repairs. However, the 4-strand cross-stitch locked repair with a 3-0 suture had significantly improved fatigue strength and holding capacity compared with the other repairs tested. Based on the consistently inferior biomechanical performance of 4-0 suture, we recommend that 3-0 suture be considered for 2- or 4-strand tendon repairs when early active motion is planned. The orientation of the transverse and longitudinal components of simple locked repairs did not significantly influence their holding capacity or fatigue strength. The cross-stitch type of locked repair provides better holding capacity and fatigue strength compared with simple locked or nonlocked 4-stranded flexor tendon repairs. PMID- 11279583 TI - Effect of pulley integrity on excursions and work of flexion in healing flexor tendons. AB - We investigated the effect of incision of a single critical pulley on excursions and work of flexion in healing flexor tendons. Forty-two long toes from 21 white leghorn chickens were used as the experimental model. Gliding excursions of the flexor digitorum profundus tendons and work of flexion of the long toes were studied 8 weeks after tendon repair to determine the functions of the healed tendons in intact, incised, or enlarged A2 pulleys. Eleven additional chickens (22 long toes) were used to obtain tendon excursion measurements in normal chicken toes. At 8 weeks, gliding excursions were statistically smaller in the intact pulley group than in the incised or enlarged pulley groups; the excursions were 73% +/- 4% for the intact pulley group, 88% +/- 9% for the incised pulley group, and 91% +/- 8% for the enlarged sheath group compared with the normal group. Work of flexion of the toes in the intact pulley group was statistically greater than that in the incised or enlarged pulley groups. Excursion efficiency of the flexor tendons was not statistically different among the toes receiving different treatments in the pulley. The results of this study demonstrate that release of a single pulley after repair of the tendons in this area improved gliding excursions of the tendons and reduced resistance to motion of the repaired tendons, and provide support for partial A2 pulley incision after repair of the tendons in the area of the pulley. PMID- 11279584 TI - Biomechanical evaluation of thumb opposition transfer insertion sites. AB - The optimal location for insertion of the transferred tendon in opposition transfer is controversial. The purpose of this study was to examine 4 commonly used insertion sites into the thumb and determine which maximizes thumb opposition. The flexor digitorum superficialis of the ring finger was used as a donor tendon and was attached in random order to the abductor pollicis brevis (APB) tendon, the APB and extensor pollicis longus, the flexor pollicis brevis (FPB) and dorsal radial extensor hood, and the ulnar extensor hood at the base of the proximal phalanx. As normal opposition was simulated, the minimum distance between the thumb and little finger and the pinch force were measured. The FPB and radial dorsal extensor hood site resulted in the statistically highest pinch force. The FPB and radial dorsal extensor hood and the APB sites had statistically smaller minimum distances between the thumb and little finger than the ulnar extensor hood site. A subjective evaluation of the 3-dimensional thumb path of motion revealed that the FPB and radial dorsal extensor hood site and the APB insertion site allowed the closest approximation of normal thumb opposition. This biomechanical study supports the use of the FPB and radial dorsal extensor hood insertion site or APB insertion site for opposition transfers. PMID- 11279585 TI - Quantitative abductor pollicis brevis strength testing. PMID- 11279587 TI - The recurrence of deformity after surgical centralization for radial clubhand. PMID- 11279590 TI - The treatment of retinoblastoma: a case study. AB - Research in the treatment of retinoblastoma and multidrug resistance has led to new treatment protocols for children. This case study introduces information regarding a clinical trial for the treatment of intraocular retinoblastoma. It also highlights important nursing issues in the care of these children and their families. PMID- 11279591 TI - Effects of prior experience with dehydration and water on the time course of dehydration-induced drinking in weanling rats. AB - Although cellular dehydration increases oral responding and swallowing of orally infused water in rats as young as 2 days old, it is not until well after the time of weaning that dehydration stimulates immediate water-seeking and initiation of drinking in situations where the water source must be approached voluntarily. Recent work has shown that the goal-directed appetitive sequence for drinking orienting, approaching, and initiating contact with water-matures much later than the more precocial oral licking and swallowing behaviors, and normally comes to be elicited by dehydration only after post-weaning experience with dry food. In the current experiments we evaluate some critical features of post-weaning experience with dehydration and drinking, and find that prior experience with initiating drinking while dehydrated, but not experience with dehydration nor water per se, alters the time course of water intake during a subsequent hydrational challenge. The effects of experience are manifested as an increased proportion of water consumed in the early portion of the test, rather than a general increase in total consumption. These findings are consistent with the interpretation that prior experience is necessary for the coordination of water oriented appetitive behaviors that lead to the initiation and maintenance of drinking bouts, and provide further evidence for an associative learning account of the acquisition of dehydration-induced drinking. PMID- 11279592 TI - Effects of the uterine environment and neocortical ectopias upon behavior of BXSB Yaa+ mice. AB - Between 40-60% of BXSB-Yaa+mice have neocortical ectopias, a genetically based brain anomaly. The presence of ectopias is known to affect several cognitive measures. A second way to affect cognition is by transferring embryos into foreign uteri. These variables were jointly investigated in three experiments. BXSB-Yaa+mouse embryos were transferred into same-strain uteri, or into uteri of hybrid mice. At birth, pups were cross-fostered to hybrid mothers or were reared by their birth mothers. When adult, the mice were given a series of behavioral tests with primary emphasis upon cognitive competence. Across all three studies, mice transferred into hybrid uteri were superior in the Morris maze and the Lashley III maze, and performed more effectively in shuttlebox avoidance learning. They were less effective in the simple water escape task, and the uterus groups did not differ in discrimination learning. Thus, development within a foreign uterus enhanced spatial learning and fear-based conditioning. Ectopic mice were superior to non-ectopics in learning the Morris maze, a finding consistent with prior research using the congenic BXSB-Yaa strain. There were Uterus x Ectopia interactions on a few measures, indicating that, under certain condition, whether the presence of ectopias is beneficial or detrimental is contingent upon the uterine environment within which the organism develops. PMID- 11279593 TI - Cholecystokinin receptor antagonists increase the rat pup's preference toward maternal-odor and rug texture. AB - The role of the cholecystokinin (CCK) system in mediating the infant's natural preferences toward maternal-related stimuli was examined by peripheral administration of selective CCK(A) and CCK(B) receptor antagonists (Devazepide and L-365,260, respectively) to 11-12-day-old rats and presenting them with a 3 minute preference test. In Experiment 1, the choice was between two floor textures, rug and plywood; the time spent on the relatively preferred side (rug) was measured. In Experiment 2, the odor of maternal faces emanated from one end of the test arena; time spent near that end was measured. These sensory stimuli were chosen as they represent olfactory and tactile aspects of the dam and nest. Compared to controls, both CCK receptor antagonists selectively increased the time spent on the preferred side, in both experiments, without affecting axillary temperature or locomotor activity. The results suggest that CCK may mediate and attenuate the infant's attraction toward naturally preferred stimuli. PMID- 11279595 TI - Early maternal rejection and later social anxiety in juvenile and adult Japanese macaques. AB - This study investigated the relationships between early maternal style and subsequent juvenile and adult behavior of offspring in Japanese macaques (Macaca fuscata). Early maternal style had no effect on baseline behavior of offspring when adult. In contrast, early maternal style was correlated with the response of adult offspring to stressful social interactions, and particularly with their response to actual or potential aggression. Infants whose mothers encouraged more independence showing high rates of rejection were less fearful and did cope better with stressful situations when adult. Although based on correlational data, these results suggest that in macaques maternal rejection can promote offspring independence and the development of a less anxious personality. PMID- 11279594 TI - Infants' long-term memory for a serial list: recognition and reactivation. AB - Serial lists contain information about item identity and item order. Using a task designed for nonverbal animals, we previously found that 3- and 6-month-olds exhibited a primacy effect after 24 hr, remembering both item identity and item order. Presently, we examined their memory of list information after longer delays. In Experiment 1, the serial-position curve reverted to a U-shape after 1 week at both ages, revealing that the common practice of attributing primacy and recency effects to long- and short-term memory, respectively, is flawed. In Experiment 2, a precuing procedure confirmed that 6-month-olds' memory still contained order information after 1 week, but 3-month-olds' reactivated memory contained none. Experiments 3A and 3B confirmed that increasing the complexity of information that was learned shortened the delay after which it could be retrieved. Testing infants after delays longer than have previously been used with animals or human adults sheds new light on an old phenomenon. PMID- 11279599 TI - Sensitive detection of rare cancer cells in sputum and peripheral blood samples of patients with lung cancer by preproGRP-specific RT-PCR. AB - RT-PCR-based amplification of transcripts expressed in cancer but not in normal non-neoplastic cells is increasingly used for the sensitive detection of rare disseminated or exfoliated cancer cells to improve cancer staging and early detection protocols. However, these assays are frequently hampered by false positive test results due to low-level transcription of the marker genes in normal cells. To overcome these limitations, target transcripts have to be identified that are tightly suppressed in normal non-neoplastic tissues, whereas they should be actively transcribed in the respective cancer cells. Here, we tested RT-PCR assays for 7 neuroendocrine marker transcripts including NCAM, PGP 9.5, gastrin, gastrin receptor, synaptophysin, preprogastrin-releasing peptide (preproGRP) and GRP-receptor to detect rare exfoliated tumor cells in peripheral venous blood and sputum samples from patients with lung cancer. Among these preproGRP RT-PCR was the only assay with which illegitimate transcription in blood or sputum samples from healthy donors or patients with unrelated diseases did not interfere. However, it reproducibly detected up to 10 small-cell lung cancer cells diluted in either 10 ml blood or 5 ml sputum samples. Single blood and sputum samples were collected directly before diagnostic bronchoscopy from 175 patients suspected to have lung cancer. Twenty-six of these had small-cell lung cancer (SCLC). Thereof, 13 patients (50%) tested positive in the blood sample and 5 of 23 patients (22%) tested positive in the sputum sample. Moreover, among 92 patients with non-small-cell lung cancer (NSCLC) 25 patients (27%) had disseminated cancer cells in peripheral blood. Amplification of preproGRP transcripts from clinical samples is a sensitive and specific assay to detect disseminated or exfoliated lung cancer cells either in peripheral blood or sputum samples. PMID- 11279600 TI - Detection of integrated papillomavirus sequences by ligation-mediated PCR (DIPS PCR) and molecular characterization in cervical cancer cells. AB - Human papillomavirus (HPV) genomes usually persist as episomal molecules in HPV associated preneoplastic lesions whereas they are frequently integrated into the host cell genome in HPV-related cancers cells. This suggests that malignant conversion of HPV-infected epithelia is linked to recombination of cellular and viral sequences. Due to technical limitations, precise sequence information on viral-cellular junctions were obtained only for few cell lines and primary lesions. In order to facilitate the molecular analysis of genomic HPV integration, we established a ligation-mediated PCR assay for the detection of integrated papillomavirus sequences (DIPS-PCR). DIPS-PCR was initially used to amplify genomic viral-cellular junctions from HPV-associated cervical cancer cell lines (C4-I, C4-II, SW756, and HeLa) and HPV-immortalized keratinocyte lines (HPKIA, HPKII). In addition to junctions already reported in public data bases, various new fusion fragments were identified. Subsequently, 22 different viral cellular junctions were amplified from 17 cervical carcinomas and 1 vulval intraepithelial neoplasia (VIN III). Sequence analysis of each junction revealed that the viral E1 open reading frame (ORF) was fused to cellular sequences in 20 of 22 (91%) cases. Chromosomal integration loci mapped to chromosomes 1 (2n), 2 (3n), 7 (2n), 8 (3n), 10 (1n), 14 (5n), 16 (1n), 17 (2n), and mitochondrial DNA (1n), suggesting random distribution of chromosomal integration sites. Precise sequence information obtained by DIPS-PCR was further used to monitor the monoclonal origin of 4 cervical cancers, 1 case of recurrent premalignant lesions and 1 lymph node metastasis. Therefore, DIPS-PCR might allow efficient therapy control and prediction of relapse in patients with HPV-associated anogenital cancers. PMID- 11279601 TI - Lung carcinogenesis: resveratrol modulates the expression of genes involved in the metabolism of PAH in human bronchial epithelial cells. AB - Studies suggest that resveratrol (trans-3,4',5-trihydroxystilbene), which is a diphenolic antioxidant found in plants and foods, has cancer chemopreventive and chemotherapeutic potential. A lower risk of lung cancer among consumers of wine compared with consumers of other beverages has been observed, which may be partly attributed to the high content of resveratrol particularly in red wine. We have studied the effect of resveratrol on the expression of genes involved in the metabolism of polycyclic aromatic hydrocarbons in the human bronchial epithelial cell line BEP2D. Expression of the cytochrome P450 1A1 (CYP1A1) and 1B1 (CYP1B1), microsomal epoxide hydrolase (mEH), and glutathione S-transferase P1 (GSTP1) genes was measured by quantitative reverse transcriptase-polymerase chain reaction. The cells were treated either with benzo[a]pyrene or 2,3,7,8 tetrachlorodibenzo-p-dioxin in the presence or absence of resveratrol. Resveratrol inhibited both the constitutive and the induced expression of CYP1A1 and CYP1B1 in a dose-dependent manner. In contrast, the expression of the mEH gene was increased in response to resveratrol and no change in the expression of GSTP1 was found. The altered gene expression in response to resveratrol was reflected in a reduced overall level of benzo[a]pyrene metabolism. These data indicate that resveratrol may exert lung cancer chemopreventive activity through altering the expression of genes involved in the metabolism of polycyclic aromatic hydrocarbons, resulting in altered formation of carcinogenic benzo[a]pyrene metabolites in human bronchial epithelial cells. PMID- 11279602 TI - Endothelial cell apoptosis is inhibited by a soluble factor secreted by human colon cancer cells. AB - In contrast to normal tissues, tumors are exposed to adverse environmental conditions, such as hypoxia and acidity. Despite this caustic environment, tumor cells and supporting vascular structures survive the latter, providing nutrients and oxygen to facilitate tumor growth. Therefore, we hypothesized that cancer cells express factors that protect endothelial cells from apoptosis. Human umbilical vein endothelial cells were grown in serum-free medium or in medium conditioned by either human colon cancer cells or non-malignant cells. Endothelial cells grown in medium conditioned by colon cancer cells demonstrated a decrease in apoptosis, whereas endothelial cells grown in medium conditioned by non-malignant cells did not. Erk-1/2 phosphorylation increased when endothelial cells were incubated in medium conditioned by colon cancer cells but not when cells were incubated in medium conditioned by non-malignant cells. Protein fractions from medium conditioned by colon cancer cells that were < 3 kDa protected endothelial cells from apoptosis and activated Erk-1/2. Heat inactivated conditioned media did not protect endothelial cells from apoptosis and did not activate Erk-1/2. Human colon cancer cells secrete a soluble factor or factors that inhibit endothelial cell apoptosis. This factor is likely to be a protein or protein fragment because it loses its activity after heat inactivation. These studies support the hypothesis that tumor cells can alter the phenotype of endothelial cells. PMID- 11279603 TI - Amyloid beta protein precursor is involved in the growth of human colon carcinoma cell in vitro and in vivo. AB - Amyloid beta protein precursor (APP) is a membrane-bound protein ubiquitously expressed in a variety of types of cells. However, its biological functions remain largely uncertain, particularly in non-neural cells and tumors. Our previous studies revealed that a secreted form of APP having a Kunitz-type inhibitor domain is a major serine proteinase inhibitor secreted by human colon carcinoma cells. In our study, we used an antisense RNA strategy to selectively inhibit the expression of APP in the human colon carcinoma cell line SW837. A vector capable of expressing an antisense mRNA complementary to 911 bases of the 5' end of APP mRNA was transfected into SW837 cells. After selection, 2 stably transfected antisense clones were obtained in which both the APP protein and mRNA were significantly suppressed. The proliferative potential and colony-forming efficiency of the antisense clones in vitro were markedly suppressed compared with the parent and mock-transfected clones. The addition of the conditioned medium of parent cells or purified secretory APP enabled these antisense effects to be overcome in vitro. The suppressed growth was also observed in vivo when the cells were injected subcutaneously into nude mice. Histologically, formation of tubular structures appeared to be suppressed in the antisense clones in vivo. These observations suggest potentially important roles of APP in cellular proliferation and differentiation of colon carcinoma cells. PMID- 11279604 TI - The alphaVbeta6 integrin regulates its own expression with cell crowding: implications for tumour progression. AB - Expression of the growth-promoting integrin alphavbeta6 in colon cancer cells induces gelatinase B secretion and activation, the inhibition of which abolishes alphavbeta6-mediated tumour cell growth within a collagen matrix. Herein, we show that high cell density selectively enhances alphavbeta6 expression in a protein kinase C (PKC)-dependent manner in preference to other beta integrin subunits, resulting in a marked increase in gelatinase B secretion as cells reach confluence. Moreover, PKC activity increases with cell confluence, and the rise in PKC activity is much greater for alphavbeta6-expressing cells than for colon cancer cells which lack alphavbeta6. We propose a self-perpetuating system of colon cancer progression in which the integrin alphavbeta6 provides a means of sustaining tumour cell proliferation. In this model, alphavbeta6 regulates its own expression via a PKC-mediated signalling pathway as tumour cells become crowded and quiescent. The alphavbeta6-mediated induction of gelatinase B secretion facilitates peri-cellular matrix degradation, which helps overcome crowding and restores cell proliferation. PMID- 11279605 TI - Aminopeptidase N regulated by zinc in human prostate participates in tumor cell invasion. AB - Aminopeptidase N (AP-N) degrades collagen type IV and is proposed to play a role in tumor invasion. However, the precise functions of AP-N in tumor cells and the relationship of AP-N to prostate cancer remains unclear. In our study, we examined a possible role for zinc in the regulation of AP-N enzymatic activity in relation to tumor cell invasion in human prostate. AP-N purified from human prostate was irreversibly inhibited by low concentrations of zinc (Ki = 11.2 microM) and bestatin. AP-N, which has zinc in the active center, was also inhibited by the chelating agents, EDTA, o-phenanthroline and EGTA. EDTA was shown to remove zinc from the enzyme. When the effects of zinc and bestatin on invasion of PC-3 cells were investigated in vitro using a Transwell cell-culture chamber, zinc and bestatin effectively suppressed cell invasion into Matrigel at the concentration range of 50-100 microM. These results strongly suggest that the suppression of PC-3 cell invasion by zinc is based on the inhibition of AP-N activity by zinc. We also evaluated the expression of AP-N to investigate the relationship with the progression of prostate disease in human cancerous prostate. AP-N was found to be located at the cytoplasmic membranes of prostate gland epithelial cells and to be expressed more in prostate cancer, while the expression of prostate-specific antigen (PSA), which is a useful marker for prostate cancer, was shown in normal and cancer tissues, suggesting that AP-N is potentially a good histological marker of prostate cancer. Thus, highly expressed AP-N in human cancerous prostate probably plays an important role in the invasion and metastasis of prostate cancer cells. PMID- 11279607 TI - Cytostatic effect of polyethylene glycol on human colonic adenocarcinoma cells. AB - Polyethylene glycol (PEG 8000) is a potent cancer chemopreventive agent. This osmotic laxative polymer markedly suppresses colon cancer in rats. To explain the mechanism, we have tested the in vitro effect of PEG on four human cell lines. Two poorly differentiated adenocarcinoma lines (HT29 and COLO205), a fetal mucosa line (FHC) and a differentiated line (post-confluent Caco-2) were incubated with various PEG concentrations for 2-5 days. Results show that PEG markedly and dose dependently inhibited HT29 and COLO205 cell growth. This cytostatic effect was associated with a blocking of the cell cycle in G0/G1 phase. In addition, PEG decreased the viability of HT29 and COLO205 adenocarcinoma cells. In contrast, post-confluent intestinal-like Caco-2 cells and normal FHC cells were, respectively, not or little affected by PEG. Moreover, the lactate concentration increased twofold in the medium of PEG-treated HT29 cells compared with untreated cells. Microscopic observations showed that PEG induced cell shrinking, membrane blebbing and the condensation of nuclear chromatin. However, because no DNA ladder and no annexin staining were detected, we presume that PEG did not induce apoptosis. PEG increased the osmotic pressure of the culture medium. Hyperosmotic media with added NaCl or sorbitol also inhibited HT29 cell growth, and increased lactate release. These results suggest that PEG may be selectively cytostatic for proliferating cancer cells. This growth inhibition may be due to the high osmotic pressure induced by PEG in vitro. Because the osmotic pressure is high in feces of PEG-fed rats, it may explain the suppression of colon carcinogenesis by PEG. PMID- 11279606 TI - The MEK1-ERK map kinase pathway and the PI 3-kinase-Akt pathway independently mediate anti-apoptotic signals in HepG2 liver cancer cells. AB - Primary liver cancers, which are generally hypervascular in nature, depend highly on blood supply. So far there are few reports on apoptosis of liver cancer cells upon deprivation of serum-derived survival factors. The aim of our study is to clarify molecular mechanisms by which liver cancer cells survive with the aid of serum. In HepG2 liver cancer cells, serum deprivation induced time-dependent increase in the number of apoptotic cells, which was detected by fragmentation of genomic DNA and fluorescent nuclear staining. The activity of extracellular signal-regulated kinase (ERK) did not decrease considerably after serum deprivation, although it increased after serum stimulation. However, we found that the MEK1 inhibitor PD98059, but not the p38 kinase inhibitor SB203580, potently induced apoptosis of the liver cancer cells in the presence of serum, indicating that the MEK-ERK signaling pathway is required for serum-dependent survival of HepG2 cells. In agreement with this notion, transient expression of active MEK1 prevented apoptosis in serum-deprived condition. We also found that the protective effect of serum against apoptosis was totally abrogated by LY294002 or wortmannin, which are the inhibitors of phosphatidylinositol (PI) 3 kinase. The activity of Akt, the target of PI 3-kinase, decreased gradually after deprivation of serum, whereas it was rapidly reactivated upon serum stimulation. These data indicate that survival of HepG2 liver cancer cells depends upon serum and that both the MEK1-ERK- and the PI 3-kinase-Akt- pathways are required for survival signaling to the nucleus. PMID- 11279608 TI - Mutation analysis of NTRK2 and NTRK3, encoding 2 tyrosine kinase receptors, in sporadic human medullary thyroid carcinoma reveals novel sequence variants. AB - Somatic mutations in the proto-oncogene RET are found in 25% to 80% of sporadic medullary thyroid carcinomas (MTCs). The significance of somatic RET mutation in MTC initiation and progression, however, remains unknown. Like RET, TRK is a neurotrophic receptor tyrosine kinase. Immunostaining has shown that only a subset of normal C cells expresses Trk family receptors, but in C-cell hyperplasia, they consistently express NTRK2, with variable expression of NTRK1 and NTRK3. In later stages of MTC, NTRK2 expression was reduced while NTRK3 expression was increased. In the context of these data, we sought to determine whether sequence variants in NTRK2 and NTRK3 are responsible for these differences in protein expression. We determined the genomic structure of NTRK2 and found that it consists of at least 17 exons varying in size from 36 to 306 bp. Mutation analysis of sporadic MTC did not reveal any sequence variants in NTRK2 but did reveal 3 variants in NTRK3, c.573C >T (N191N, exon 5), c.678T > C (N226N, exon 6) and c.1488C > G (A496A, exon 12) occurring among 19 chromosomes (31%), 1 chromosome (2%) and 24 chromosomes (39%), respectively. Corresponding germline also harbored these variants. There was a trend toward excess association of the NTRK3 variant c.1488C > G (A496A) in cases (24/62 chromosomes, 39%) compared to controls (18/62, 29%), but this difference did not reach significance (p > 0.05). The remaining 2 NTRK3 variants occurred with similar frequencies between MTC cases and population-matched controls (19 vs. 17 and 1 vs. 0, p > 0.05). We conclude that sequence variants in NTRK2 and NTRK3 are not likely to be responsible for large differences in expression at the protein level, but we cannot exclude very low penetrance effects. PMID- 11279609 TI - Expression of the recombination activating genes (RAG1 and RAG2) is not detectable in Epstein-Barr virus-associated human lymphomas. AB - The expression of the recombination activating genes (RAG1 and RAG2) is largely restricted to immature lymphoid cells. Previous studies have suggested that Epstein-Barr virus (EBV) infection may lead to a re-induction of RAG expression in mature B lymphocytes. To assess the significance of this mechanism for the pathogenesis of malignant lymphomas, we have examined the expression of RAG genes in 11 cases of EBV-associated endemic Burkitt's lymphoma (BL), 25 cases of Hodgkin's disease (HD, 17 EBV(+), 8 EBV(-)) and 10 cases of follicular non Hodgkin's lymphoma (NHL). Using in situ hybridization, expression of the RAGs was detected in cortical thymocytes in normal thymus and in the tumor cells of 2 of 3 lymphoblastic NHL. By contrast, there was no detectable RAG expression in the BL, HD and follicular NHL cases. Our results indicate that re-induction of RAG expression does not occur in human lymphomas in vivo. Thus, it is unlikely to play a role in the development of translocations involving immunoglobulin gene loci which are characteristically found in BL and follicular NHL. Moreover, our study shows that in situ hybridization is a suitable method for the analysis of RAG expression in human tissue sections. PMID- 11279610 TI - Immunogenicity of recombinant GA733-2E antigen (CO17-1A, EGP, KS1-4, KSA, Ep-CAM) in gastro-intestinal carcinoma patients. AB - Targeting the GA733 antigen (also known as CO17-1A, EGP, KS1-4, KSA, Ep-CAM) by monoclonal antibody CO17-1A or anti-idiotypic antibodies mimicking the CO17-1A or GA733 epitope has induced prolonged survival and specific immune responses to the antigen, respectively, in colorectal cancer (CRC) patients. In pre-clinical studies in mice and rabbits, recombinant baculovirus-derived GA733-2E antigen was superior to anti-idiotypic antibodies at modulating specific immune responses. Our aim was to evaluate the immunogenicity and potential toxicity of alum precipitated GA733-2E in a phase I trial in patients with resected CRC or pancreatic cancer. Six patients with advanced pancreatic carcinoma and 6 with CRC Dukes' stage A, B or C received between 4 and 7 doses of alum-precipitated GA733 2E at 50, 200 or 800 microg/dose at monthly intervals. Antibody binding to GA733 2E or antigen-positive CRC cells was determined, as were antigen-specific proliferative, cytolytic T-lymphocyte and delayed-type hypersensitivity responses. Six of the 12 patients developed antigen-specific humoral immune responses after immunotherapy, and 8 developed cellular immune responses. The overall immune response rate, including patients with humoral and/or cellular immune responses, was 83%. Median overall survival of the CRC and pancreatic cancer patients was 39.8 and 11.2 months, respectively. Following 18 years of single-epitope targeting of the GA733 antigen, immunization of patients against multiple epitopes of the antigen frequently induces an immune response in the absence of significant toxicity, despite relatively widespread expression of this antigen on normal epithelial cells. PMID- 11279611 TI - Murine monoclonal anti-idiotypic antibody as a surrogate antigen for human Her 2/neu. AB - The anti-idiotypic (Id) monoclonal antibody (MAb) 520C9-6b (IgG1k), raised in syngenic mice against the murine anti-Her2/neu MAb 520C9 (Ab1), functionally mimics a human Her2/neu epitope and serves as a surrogate for the protein antigen. Immunization of allogeneic C57BL/6 mice and rabbits with 520C9-6b (Ab2) induced anti-Her2/neu-specific antibodies that react with antigen-positive SKBr3 cells by ELISA and FACS analysis. The immune sera inhibited binding between Ab1 and Ab2 and vice versa (binding of Ab2 to Ab1), indicating that it was a true anti-anti-Id (Ab3) antibody. The Ab3 sera or purified Ab3 specifically lysed Her2/neu-positive SKBr3 cells, but no significant lysis was observed in antigen negative LS174T cells in an antibody-dependent cellular cytotoxicity assay. An Id specific cellular immune response was also demonstrated in an in vitro lymphocyte proliferation assay. Furthermore, a panel of tumor tissues and tumor cells was screened for the presence of the Her2/neu epitope by its reactivity with Ab1 and Ab3 using immunohistochemistry and FACS analysis. Identical results were obtained using either Ab1 or Ab3 (Ab1'). The data indicated that anti-Id 520C9-6b can induce Her2/neu-specific antibody in experimental animals and may serve as a potential network antigen for the treatment of patients bearing Her2/neu-positive tumors. PMID- 11279613 TI - Influence of the route of administration on targeting of ovarian cancer with the chimeric monoclonal antibody MOv18: i.v. vs. i.p. AB - MOv18 antibody binds the membrane folate receptor highly expressed on ovarian carcinoma cells. Since ovarian cancer is mainly limited to the peritoneal cavity, locoregional delivery of therapeutics can be an option. The same patient was injected i.v. and i.p. with c-MOv18 IgG labeled with different radionuclides. To study the kinetics of c-MOv18, patients were divided into 2 groups. Fifteen patients received c-MOv18 labeled with (131)I, (125)I and (123)I (for imaging). Seven patients were operated 2 days, 7 patients 6 days and 1 patient 3 days post injection. Radioactivity was determined in blood, ascites and biopsies of tumor and of several normal tissues. No adverse events occurred. No anti-MOv18 responses were observed. The area under the blood activity vs. time curve (AUC) was significantly lower after i.p. injection for 2 and 6 days post-injection (p = 0.01 and p = 0.02, respectively). At 2 days post-injection, a significant difference in tumor uptake was found in favor of the i.v. route of administration (4.9% and 2.4%ID/kg for i.v. and i.p., respectively; p < 0.0001). Uptake in solid tumor tissue in ovarian cancer patients operated 6 days post-injection was not significantly different (p = 0.79) for both routes (3.8% and 3.9%ID/kg for i.v. and i.p., respectively). In conclusion, no advantage could be demonstrated for the i.p. route with respect to tumor uptake. The i.p. route could be advantageous with respect to bone marrow toxicity since the AUC was significantly lower for the i.p. route. However, within 2 days post-injection, the blood clearance followed the same pattern for both routes. PMID- 11279612 TI - Enrichment of memory T cells and other profound immunological changes in the bone marrow from untreated breast cancer patients. AB - Previous studies with animal tumors showed that bone marrow (BM) is a privileged site where potentially lethal tumor cells are controlled in a dormant state by the immune system. Here, we investigated BM of breast cancer patients with respect to tumor cell content, immune activation status and memory T-cell content. BM-derived cells from primary operated breast cancer patients (n = 90) were compared with those from healthy donors (n = 10) and also with cells from respective blood samples. Cytokeratin 19-positive tumor cells were detected by nested polymerase chain reaction. Three-color flow cytometry was used to identify numbers and activation state of T cells, natural killer (NK) cells, monocytes/macrophages and subsets by a panel of monoclonal antibodies (mAbs). The proportion of memory T cells among the CD4 and CD8 T cells was much higher in BM of cancer patients than in healthy donors (p < 0.001). The extent of memory T cell increase was related to the size of the primary tumor. Patient-derived BM memory CD8 T cells could be shown to contain specific HLA-A2/Her-2/neu(369-377) tetramer binding cells. Patients with disseminated tumor cells in their BM had more memory CD4 T cells and more CD56(+) CD8(+) cells than patients with tumor cell-negative BM. Only some of the immunological changes seen in BM samples of cancer patients were also detectable in peripheral blood samples. Our hypothesis that BM is a special compartment for immunological memory and tumor dormancy is supported by the above findings. The overall results reveal that BM is a valuable additional compartment for immune diagnosis in pathological conditions and possibly for follow-up treatment strategies. PMID- 11279614 TI - Overcoming multi-drug resistance using an intracellular anti-MDR1 sFv. AB - We made an intracellular single-chain variable fragment (sFv) from the C219 monoclonal antibody that recognized the intracellular domain of the multidrug resistance (MDR) gene product, P-glycoprotein (P-gp). Immuno-cytochemistry using the FITC conjugated anti-C-myc tag antibody showed that the sFv protein was expressed in the cytoplasm of the cells. Although transfection of the sFv did not result in the down-regulation of P-gp expression in P-gp positive MDR cells as determined by flow cytometry analysis, Adriamycin (ADM) uptake and Rhodamine123 (Rh123) retention were increased by the C219 intra-cellular sFv transfection. The transfected cells exhibited a higher sensitivity to ADM using a 10-day colony formation assay. The conventional 3-day MTT assay showed the drug resistant tendency in C219 sFv transfected cell we tested. The growth rate of C219 sFv transfected cells was delayed in all non-MDR and MDR cells that might be the reason why C219 transfected cells exhibited the drug resistant tendency in the MTT assay. Despite this unexpected effect of C219 sFv on growth rate, our data suggest that the intra-cellular sFv technique could knockout MDR functionally and may offer a means of increasing the effectiveness of tumor chemotherapy. PMID- 11279615 TI - Acquired resistance of melanoma cells to the antineoplastic agent fotemustine is caused by reactivation of the DNA repair gene MGMT. AB - Acquired resistance to antineoplastic agents is a frequent obstacle in tumor therapy. Malignant melanoma cells are particularly well known for their unresponsiveness to chemotherapy; only about 30% of tumors exhibit a transient clinical response to treatment. In our study, we investigated the molecular mechanism of acquired resistance of melanoma cells (MeWo) to the chloroethylating drug fotemustine. Determination of O(6)-methylguanine-DNA methyltransferase (MGMT) activity showed that MeWo cells that acquired resistance to fotemustine upon repeated treatment with the drug display high MGMT activity, whereas the parental cell line had no detectable MGMT. The resistant cell lines exhibit cross resistance to other O(6)-alkylating agents, such as N-methyl-N'-nitro-N nitrosoguanidine. Acquired resistance to fotemustine was alleviated by treatment with the MGMT inhibitor O(6)-benzylguanine demonstrating that reactivation of MGMT is the main underlying cause of acquired alkylating drug resistance. As compared with control cells, both MGMT mRNA and MGMT protein were expressed at a high level in fotemustine resistant cells. Southern blot analysis proved that the MGMT gene was not amplified. There was also only an insignificant difference in the CpG methylation pattern of the MGMT promoter whereas a clear hypermethylation in the body of the gene was observed in fotemustine resistant cells. The conclusion that hypermethylation is responsible for reactivation of the MGMT gene gained support by the finding that MGMT activity significantly declined and cells reverted (partially) to the parental sensitive phenotype upon treatment with 5 azacytidine. This is the first report of acquired resistance to a chloroethylating antineoplastic drug of melanoma cells due to gene hypermethylation. PMID- 11279617 TI - On the pharmacokinetics of topically applied 5-aminolevulinic acid and two of its esters. AB - The kinetics of protoporphyrin IX (PpIX) production in normal tissues and WiDr tumors of mice were studied after topical application of 5-aminolevulinic acid (ALA) and its methyl ester and hexyl ester. ALA and ALA esters were applied on a spot of 1.0 cm diameter on normal skin and on skin overlaying tumors. PpIX production was studied by fluorescence measurements. ALA induced PpIX not only on the spot of application but also on remote skin areas. This was not found for the ALA esters. They produced PpIX only on the spot of application. Thus, ALA, but neither its esters nor PpIX, is passing into the circulation. The time needed for ALA to enter the circulation through normal skin was about 5 hr. Even when looking normal, the skin overlaying tumors was more permeable to ALA than normal skin. Thus, when applied on the tumor, ALA induced PpIX on remote skin areas without any lag phase. Mainly, PpIX was found in all tissues although small amounts of a porphyrin with an excitation peak at about 400 nm, supposedly uroporphyrin and/or coproporphyrin, were found, notably in remote skin areas. An altered stratum corneum of the skin overlaying tumors probably contributes to the tumor-selectivity, although in the present tumor system less PpIX was found in tumors than in muscles. This is probably related to biochemical and physiological conditions in this particular tumor, since i.p. injection of ALA also leads to less PpIX formation in the tumor than in skin/muscle tissue. Nevertheless, it seems evident that ALA can diffuse more easily from the skin surface and down to the vasculature in the tumor than in the normal tissue and that this leads to a higher concentration of PpIX in the tumor than would have been found if the physiological factors relevant for drug diffusion were the same for tumors as for skin/muscles. PMID- 11279616 TI - Connexin 43 (cx43) enhances chemotherapy-induced apoptosis in human glioblastoma cells. AB - Stable re-expression of connexin 43 (cx43) in human glioblastoma suppresses transformation and tumorigenicity. The present study was designed to examine the role of cx43 in chemotherapy-induced apoptosis. Expression of cx43 in human glioblastoma cells significantly increased sensitivity to several common chemotherapeutic agents, including etoposide, paclitaxel (Taxol) and doxorubicin, compared with control-transfected cells. The increased sensitivity to chemotherapeutic agents resulted from apoptosis as evidenced by Hoechst dye staining, TUNEL assay and annexin V assay. These cx43-mediated effects were coupled with decreased expression of the specific apoptosis inhibitor bcl-2. Over expression of bcl-2 in cx43-transfected cells partially confers the resistance to apoptosis induced by etoposide, suggesting that the cx43-mediated apoptosis to chemotherapeutic agents is regulated in part through the down-regulation of bcl-2 expression. Furthermore, the cx43-mediated apoptosis in response to chemotherapeutic drugs may not be linked to increased gap junctional communication in cx43-transfected cells. Our results demonstrate a new role of cx43 in the mediation of apoptosis during chemotherapy. PMID- 11279618 TI - Modification of cancer risks in offspring by sibling and parental cancers from 2,112,616 nuclear families. AB - Comparisons of cancer risks in persons by sibling cancers and those by parental cancers are informative of elucidating the potential genetic modes in the etiology of the cancers. The Swedish Family-Cancer Database was used to systematically estimate the effects of parental and sibling cancers on the cancer risks in the individuals born after 1934 (offspring). The study population included 5,520,756 offspring and their parents from 2,112,616 nuclear families. Standardized incidence ratios (SIRs) were calculated to analyze the risks for cancers in offspring by parental cancers (offspring risk) and by sibling cancers (sibling risk). For 20 concordant sites, all offspring and sibling risks were significantly increased except for sibling risks for squamous cell carcinoma of the skin and myeloma. Apart from breast cancer, the SIRs were more than 10 when offspring had both an affected parent and an affected sib at the concordant site. The ratio for the sibling to offspring risk was around 2.0 or more for gastric, renal, non-thyroid endocrine, urinary bladder, colon, testicular and prostate cancers and leukemia. For discordant sites, many reported across-site associations were confirmed and several consistent novel associations (rectum skin, breast-endocrine and lung-endocrine) were found only among sibs. Our findings suggested that low-penetrance polygenic dominant effects or dominant genes of high penetrance but low mutant allele frequency in the population may be involved in the observed familial cancers at many sites. Recessive or X-linked effects may contribute particularly to gastric, renal, non-thyroid endocrine, bladder, colon, testicular and prostate cancers and leukemia. The search for pleiotropic recessive/X-linked susceptibility genes should be well motivated based on our results. PMID- 11279619 TI - Measuring interval cancers in population-based screening using different assays of fecal occult blood testing: the District of Florence experience. AB - The fecal occult blood test (FOBT) has demonstrated its efficacy in reducing mortality from colorectal cancer (CRC). The guaiac-based FOBT has been criticized for its low sensitivity. In this study, two different assays for FOBT (guaiac or an immunochemical test based on reversed passive hemagglutination [RPHA]) were tested for comparison within a population-based screening program for colorectal cancer in the province of Florence (Italy). The proportional incidence method was used to calculate sensitivity for both FOBTs, according to rank of screening (first or repeat), age at entry (two groups of 50 to 59 and 60 to 70 years old) and lesion site (colon or rectum). When comparing FOBTs, the sensitivity multivariate Poisson regression was used to adjust for other variables. The sensitivity after the first 2 years was 50% (95% confidence interval [CI] 34% to 63%) for the guaiac test versus 82% (95% CI 67% to 92%) for RPHA. At multivariate analysis the risk of developing an interval cancer after a guaiac test is almost 3 times that after RPHA (rate ratio = 2.64; 95% CI 1.3 to 5.4). Our study confirms that RPHA is more sensitive than the guaiac test. The assumption that FOBT screening for CRC has to be based on a guaiac test should be reconsidered, and RPHA should be recommended as the standard FOBT for screening purposes. PMID- 11279620 TI - Physical activity and risk of renal cell cancer. AB - The relation between physical activity and renal cell cancer is unclear. High occupational physical activity has been associated with a decreased risk of renal cell cancer among men-but not among women-in two previous studies, while no association has been found for leisure time physical activity. Our aim was to investigate the association between occupational and leisure time physical activity in a prospective cohort of 17,241 Swedish twins. Information on physical activity and a wide range of potential confounding factors was obtained through a mailed questionnaire. During follow-up from 1967 through 1997 we identified 102 cases of renal cell cancer. We found no evidence of an inverse association between either occupational or leisure time physical activity and risk of renal cell cancer in this prospective cohort. PMID- 11279621 TI - Trends in non-Hodgkin lymphoma (NHL) and HIV-associated NHL deaths in the United States. AB - Since a significant number of lymphomas have been associated with the human immunodeficiency virus (HIV), the purpose of this study was to describe the impact of HIV infection on non-Hodgkin's lymphoma (NHL) mortality trends and demographics. Multiple-cause-of-death data for the United States from 1979 through 1996 were obtained from the National Center for Health Statistics, Centers for Disease Control and Prevention. Annual NHL deaths rates for the United States were calculated as the number of NHL deaths per 100,000 persons, based on estimates of the U.S. resident population. The time periods 1979-1982, 1986-1989, and 1993-1996 were examined for changes over time. To describe NHL and HIV infection mortality, the characteristics of NHL deaths with HIV infection listed anywhere on the death records were examined beginning in 1987. This study found that despite reports of a lower incidence rate of NHL among blacks with HIV/AIDS, death rates from lymphomas associated with HIV/AIDS have markedly increased in black males and females over time. It was also noted that in agreement with other studies, this study documented a decrease in NHL/HIV mortality in 1996. PMID- 11279622 TI - Bone marrow cytogenetic abnormalities of aplastic anemia. AB - Cytogenetic abnormalities in association with aplastic anemia have been reported fairly infrequently. Clonal cytogenetic abnormalities at initial diagnosis are uncommon. A retrospective study was performed of the cytogenetic findings in patients with typical morphological and clinical features of severe aplastic anemia from a single institution for the years 1988 through 1998. A total of 30 cases of aplastic anemia, 16 men and 14 women, were identified. The median age was 60 with females being significantly older (67.5 years) in comparison to males (44 years). Bone marrow specimens failed to yield metaphases in 16 cases and normal karyotypes were detected in 11 cases. Cytogenetic abnormalities were detected in 3 cases. Clonal abnormalities, as defined, occurred in only 2 cases (6.7%). A review of the literature identified a total of 24 cases of aplastic anemia with abnormal cytogenetic findings. Overall, the most common chromosome abnormalities are trisomies of 6 and 8 and loss of chromosome 7. Trisomy 6 is more common at diagnosis while loss of chromosome 7 is more common after therapy. PMID- 11279623 TI - A pilot study of rhuIL-11 treatment of refractory ITP. AB - The objective of this research was to determine whether rhuIL-11 is an effective treatment in patients with refractory immune thrombocytopenic purpura (ITP). Platelet production is decreased in certain cases of refractory ITP. IL-11 stimulates megakaryocytopoiesis in vitro and was licensed for its clinical effects to ameliorate chemotherapy-induced thrombocytopenia. A pilot study was initiated, intending to enroll 12 patients with ITP. These patients were to receive rhuIL-11 (Neumega) at a dose of 50 microg/kg subcutaneously daily for 21 consecutive days and be observed afterward for 21 additional days. CBC with platelets were obtained twice weekly with visits and physical examinations weekly. The study was terminated after 7 patients were enrolled because of toxicity and lack of efficacy. All 7 patients had had ITP for >9 years and had failed splenectomy, intravenous gammaglobulin, corticosteroids, and a variety of other treatments. The patients at entry all had platelet counts <20,000/microl; 5 of 7 had counts <10,000/microl. The maximal median increase for any day of the study was 6,000/microl. No patient achieved a count of 30,000/microl, and only 3 patients achieved (once each) a platelet count >20,000/microl. Substantial toxicity was seen. The nadir hemoglobin decrease was a mean of 2 g/dl. rhuIL-11 was not effective at increasing the platelet count in any of these patients with refractory ITP. Toxicity was substantial. The lack of platelet response to rhuIL 11 in this study does not exclude the possibility of better effects at other doses and/or in less refractory patients. PMID- 11279624 TI - Oral combination chemotherapy in conjunction with filgrastim (G-CSF) in the treatment of AIDS-related non-Hodgkin's lymphoma: evaluation of the role of G CSF; quality-of-life analysis and long-term follow-up. AB - In 1993 we reported the efficacy and toxicity profile of an oral combination regimen administered to 18 patients with AIDS-related lymphoma (NHL-1 study). We observed a 61% response rate; 39% one-year survival rate; nearly two-thirds of patients developed > or = grade 3 leukopenia; and 28% of cycles were associated with febrile neutropenia. These results prompted us to shorten the duration of therapy and to add G-CSF to ameliorate the myelosuppression. Twenty patients with biopsy-proven AIDS-related lymphoma were treated with three 6-week cycles of oral chemotherapy consisting of lomustine (CCNU) 100 mg/m2 on day 1, cycles no. 1 and 3; etoposide 200 mg/m2 days 1-3; cyclophosphamide and procarbazine both 100 mg/m2 days 22-31; and G-CSF 5 microg/kg subcutaneously days 5-21 and days 33-42 (NHL-2 study). The following analyses were undertaken: (1) evaluation of toxicity and efficacy parameters for patients in the current (NHL-2) study; (2) analysis of the clinical role of G-CSF by (historical) comparison with the NHL-1 study of the same regimen without G-CSF; (3) quality-of-life assessments using the Functional Living Index-Cancer (FLIC) and Brief Symptom Inventory (BSI) instruments for all 38 patients (NHL-1+2); and (4) long-term follow-up for all 38 patients. In the current study the overall objective response using ECOG criteria was 70% (95% CI, 50-90%) with 6 CRs (30%) and 8 PRs (40%). The median survival duration was 7.3 months (range: 0.5-51+ months). One patient developed CNS relapse. There were no significant differences with respect to demographics or prognostic factors between the patient populations of the NHL-1 study and the current study (P > 0.2 for each factor). Myelosuppression was the major toxicity in both studies. In the current study versus the NHL-1 study, although the lower incidences of grade 3/4 myelosuppression (51% vs. 64%) and febrile neutropenia (17% vs. 28%) on a per cycle basis were not statistically significant, fewer patients (40% vs. 60%) were affected. However, the severity of myelotoxicity was lessened with the addition of G-CSF, measured in terms of the discontinuation of therapy, myelotoxic deaths, and freedom from grade 3/4 myelotoxicity ( P < 0.02). The number of hospitalizations for febrile neutropenia (7 in the NHL-2 study vs. 13 in the NHL 1 study) was also significantly different (P < 0.05). Quality-of-life analysis confirmed no significant functional or psychological deterioration during therapy except for patients experiencing febrile neutropenia, whose functional capacity deteriorated (P < 0.04). The 1-year, 18-month, and 2-year survival rates for the combined studies (38 patients) were 32%, 21%, and 13%, respectively. At time of death 49% of patients were free from progression of their lymphoma. Administration of the oral regimen has resulted in 13% of patients surviving two years, and half of patients surviving free from progression of their lymphoma. This regimen is efficacious and considerate of patient quality-of-life issues. The addition of G-CSF to the regimen decreases the frequency of hospitalization for febrile neutropenia. PMID- 11279625 TI - Rare adult acute lymphocytic leukemia with CD56 expression in the ECOG experience shows unexpected phenotypic and genotypic heterogeneity. AB - Expression of CD56, a marker of natural killer (NK) cells, in acute lymphocytic leukemia (ALL) is rare and, to date, has been described only in non-B lineage ALL. Among 194 patients with CD56 analysis on the ongoing Eastern Cooperative Oncology Group (ECOG) ALL trial, E2993, 6 cases of CD56+ ALL were found (3.1%) with a median of 95% of blast cells expressing CD56, compared with a median of 1% of blast cells in CD56- ALL (P = 0.0001). FAB-L2 characteristics dominated, without granulation. Blast cells from four CD56+ patients expressed T-cell antigens at variable levels of maturation. A clonal rearrangement of the T-cell receptor beta (TCRbeta) gene was detected only in one patient. TCRbeta variable gene usage studies in this and one other CD56+ ALL patient demonstrated a significantly perturbed usage pattern in both patients when compared with control lymphocytes. The two remaining cases typed as early pre-B ALL (CD19+, CD10+), with one case co-expressing CD7. Cytogenetically, 4 patients were normal, 1 complex abnormal, and 1 Philadelphia chromosome positive. Epstein-Barr virus (EBV) sequences were detected in one T- and both B-lymphoid cases. Our data suggest that CD56 is expressed at a precursor stage common to the T- and the B cell lineage. PMID- 11279626 TI - Response to high-dose intravenous immune globulin as a valuable factor predicting the effect of splenectomy in chronic idiopathic thrombocytopenic purpura patients. AB - This study was conducted to verify whether the response to high-dose intravenous immune globulin (IVIG) was related to the effect of splenectomy in chronic idiopathic thrombocytopenic purpura (ITP) patients. A total of 79 patients over 16 years of age were enrolled in this study. The response to the treatment was classified on the basis of the platelet count as no response (NR, <50 x 10(9)/l), incomplete response (IR, (50-150) x 10(9)/l), and complete response (CR, >150 x 10(9)/l). The response was evaluated after the infusion of high-dose IVIG, within 2 weeks after splenectomy (immediate response), and during a follow-up period of more than 6 months after splenectomy (sustained response), respectively. 58 patients (73.4%) showed responses (CR or IR) to high-dose IVIG. After splenectomy, immediate responses were observed in 73 patients (92%). The response to high-dose IVIG had no relationship with the immediate response to splenectomy (P = 0.333). A follow-up evaluation was possible with 58 patients; 6 patients with NR in immediate responses did not show any response during the follow-up period, and 17 patients relapsed within 6 months after immediate responses, so 35 patients (60.3%) had sustained responses. Responders to IVIG had significantly higher sustained response rates to splenectomy than non-responders (62% vs. 38%, P = 0.001). These results indicate that the response to high-dose IVIG could be a valuable factor predicting the sustained response to splenectomy in chronic ITP patients. PMID- 11279627 TI - High-grade T-cell lymphoma complicating B-cell chronic lymphocytic leukemia: an unusual manifestation of "Richter's syndrome". AB - Approximately 3% of patients with B-cell chronic lymphocytic leukemia (CLL) develop a high-grade large-cell lymphoma consistent with Richter's Syndrome. In most cases, these lymphomas are of B-cell origin and are believed to arise by clonal evolution from the CLL cells. We present a case of a patient with a 10 year history of B-CLL who developed an aggressive large-cell lymphoma, confirmed by immunophenotype to be of T-cell origin. We suggest that in patients with CLL, immunodysregulation can result in the proliferation of T cells, which may mutate and result in the development of a new malignant clone. PMID- 11279628 TI - Acute monoblastic leukemia with t(8;16): a distinct clinicopathologic entity; report of a case and review of the literature. AB - We report a case of acute monoblastic leukemia with t(8;16) in a 71-year-old man who had rapid rise of leukocyte counts from 20.3 x 10(9)/l to 62.7 x 10(9)/l in two weeks. The peripheral blood showed many granular promonocytes that led to the consideration of acute promyelocytic leukemia of the hypogranular variant. The bone marrow, however, revealed mainly monoblasts with erythrophagocytosis. Cytogenetic study finally confirmed the diagnosis of acute monoblastic leukemia with t(8;16). The patient died three days after admission. The demonstration of these two characteristic features of this subtype, granular promonocytes and erythrophagocytosis by monoblasts, separately, in the peripheral blood and bone marrow is unusual and misleading. This cytogenetic abnormality can be demonstrated only in M5 and M4 with characteristic clinical features of disseminated intravascular coagulation, extramedullary involvement, and poor prognosis. Although it is not a common disease, this specific subtype of acute myelogenous leukemia is consistently associated with a specific cytogenetic marker, thus it should be considered a distinct clinicopathologic entity. PMID- 11279629 TI - Neutropenic enterocolitis: a rare presenting complication of acute leukemia. AB - Neutropenic enterocolitis is a necrotizing inflammatory process with intramural infection that occurs predominantly in neutropenic patients. This syndrome is most frequently observed after chemotherapy for hematologic and solid tissue malignancies, but it can also be observed in a number of other clinical settings as well. Neutropenic enterocolitis can be a rare presenting complication of acute leukemia. We report a case of acute lymphoblastic leukemia that presented with abdominal pain due to neutropenic enterocolitis. The diagnostic and treatment challenges associated with this manner of presentation are discussed. PMID- 11279630 TI - Anti-lymphoma effect of naproxen and indomethacin in a patient with relapsed diffuse large B-cell lymphoma. AB - A 77-year-old man with relapsed non-Hodgkin's lymphoma, diffuse large B-cell type, was treated with naproxen, a nonsteroidal anti-inflammatory drug (NSAID), for paraneoplastic fever. A dramatic disappearance of not only the fever but also generalized lymphadenopathy was observed. Naproxen was continued, and he maintained remission for 10 months. When relapse of lymphoma occurred in spite of continuous naproxen administration, indomethacin, another type of NSAID, was tried. Surprisingly, rapid regression of lymphoma occurred again and was maintained for almost 1 year. These results indicate that NSAIDs are effective in some patients with non-Hodgkin's lymphoma. PMID- 11279631 TI - Recombinant human erythropoietin use in hemolytic anemia due to prosthetic heart valves: a promising treatment. AB - Two patients are reported with hemolytic anemia due to red blood cell fragmentation in association with prosthetic heart vales. They were treated with erythropoietin which eliminated the need for packed red blood cell transfusion. PMID- 11279632 TI - Effects of HIV infection on age and cause of death for persons with hemophilia A in the United States. AB - Because of changes in factor replacement therapy and in treatment of human immunodeficiency virus (HIV) infection, we examined death record data for persons with hemophilia A in the United States to evaluate effects of HIV infection on age and causes of death. Multiple cause-of-death data from 1968 through 1998 were examined to assess death rates for persons with hemophilia A. ICD-9 coded causes of death from 1979 through 1998 were examined to assess long-term trends. From 1979 through 1998, 4,781 deaths among persons with hemophilia A were reported, of which 2,254 (47%) had HIV-related disease listed as a cause of death. In the late 1980s, mortality among persons with hemophilia A increased markedly, and the age adjusted death rate peaked at 1.5 per 1,000,000 population in 1992. Median age at death decreased from 55 years in 1979-1982 to 40.5 years in 1987-1990, and increased to 46 years in 1995-1998. In the period 1995-1998, the median age of hemophilia A decedents with HIV-related disease was 33 years, compared to 72 years for those without HIV-related disease; the most frequently listed causes of death for those without HIV-related disease were hemorrhagic and circulatory phenomena; the most frequently listed for those with HIV-related disease were diseases of liver and the respiratory system. From 1995 to 1998, hemophilia A associated deaths decreased by 41%, with a 78% decrease among those who had HIV related disease. Although HIV infection has adversely effected mortality for persons with hemophilia A, the marked recent decrease in the death rate among persons with hemophilia A appears to reflect advances in care for those with HIV related disease and is consistent with a decline in HIV mortality observed in the general population. PMID- 11279633 TI - Results of an outpatient-based stem cell allotransplant program using nonmyeloablative conditioning regimens. AB - Using nonmyeloablative, immunosuppressive, fludarabine (FLU)-based conditioning regimens, we have performed allogeneic peripheral blood stem cell transplants in 26 patients (8 with chronic myelogenous leukemia, 6 with acute myelogenous leukemia, 10 with acute lymphoblastic leukemia, 1 with myelodysplasia, and 1 with thalassemia major). Conditioning consisted of FLU/busulphan/cyclophosphamide/cyclosporin-A (CyA)/methotrexate, or FLU/melphalan/CyA/methotrexate. The median granulocyte recovery time to 0.5 x 10(9)/l was 11 days, whereas the median platelet recovery time to 20 x 10(9)/l was 12 days. Twelve patients did not need red blood cell transfusions, and 8 did not need platelet transfusions. In 21 individuals (81%), the procedure could be completed fully on an outpatient basis. Follow-up times range between 30 and 600 days: one patient failed to engraft and recovered endogenous hemopoiesis; six out of 26 patients developed acute graft-versus-host disease (GVHD) whereas 7/22 developed chronic GVHD. Twelve patients (46%) have died, nine of them with a relapsing disease and three with GVHD; median post-transplant survival (SV) was 300 days, whereas the 12-month SV was 42%. The 100-day mortality was 3.8% and the transplant-related mortality was 11.5%. This procedure is substantially less costly than its counterpart, using in-hospital myeloablative conditioning regimens, and it may represent another approach in the management of patients requiring an allogeneic stem cell transplant. PMID- 11279634 TI - Pharmacodynamics and pharmacokinetics of single doses of subcutaneous pegylated human G-CSF mutant (Ro 25-8315) in healthy volunteers: comparison with single and multiple daily doses of filgrastim. AB - Ro 25-8315 is produced by conjugation of rhG-CSF mutant with polyethylene glycol (PEG). The purpose of this study was to examine the pharmacodynamics and pharmacokinetics of Ro 25-8315 in comparison with Filgrastim (rhG-CSF). Subjects received single subcutaneous doses of Ro 25-8315 ranging from 10 to 150 microg/kg using a double-blind, randomized, placebo-controlled design. Filgrastim was administered as a single dose (5 or 10 microg/kg) and, following a 14-day washout period, daily for 7 days. Ro 25-8315 increased absolute neutrophil count (ANC) by 6- to 8-fold and CD34+ cell count more than 30-fold at the highest doses tested. Single doses (60-150 microg/kg) of Ro 25-8315 and multiple doses of Filgrastim had similar effects on ANC and CD34+, although Ro 25-8315 had a greater effect on CFU-GM. The pharmacokinetics of Ro 25-8315 were dose-dependent, with peak concentrations and area under the serum concentration-time curve (AUC) increasing 100-fold over the range of doses studied. Time to reach peak concentration (T(max)) and half-life of Ro 25-8315 averaged 20-30 hr at all doses, approximately three times longer than with Filgrastim. Adverse events were not serious and occurred with similar frequency with both products. Pegylation of rhG CSF mutant results in more desirable pharmacokinetic properties and a longer duration of action with effective increases in ANC and measures of peripheral blood progenitor cell mobilization for at least 1 week. PMID- 11279635 TI - Racial variation in fasting and random homocysteine levels. AB - Homocysteine (Hcy) levels have been shown to be a predeterminant of thrombotic diseases. We measured the Hcy levels of 50 blacks and 50 whites equally divided by gender to determine if there is a significant racial difference in either fasting or random Hcy levels. Dietary, medication, smoking, alcohol, past medical, educational, and occupational histories were obtained, and the body mass index calculated. Total serum fasting and random Hcy levels, B12, folate, BUN, creatinine, and lipid profiles were drawn from each participant. Analysis of the results showed that white males have the highest fasting Hcy levels, 10.5 microM/l, whereas random Hcy levels were not significantly different. Correlation between fasting and random Hcy levels was poor (R = 0.61). B12 levels in black subjects were significantly higher, 490.8 pg/ml, compared to whites, 382.8 pg/ml, P = 0.001, but contributed little to total Hcy levels (R(2) = 0.08). Folic acid levels, all within normal range, were not significantly different between the two racial groups and also did not appear to greatly affect Hcy levels (R(2) = 0.06). Our study demonstrates that, despite the genetic diversity of these two racial groups in the U.S., white males in this age group have higher fasting Hcy levels than black males, and white males, but not black males, have higher fasting homocysteine levels than females. This discrepancy in Hcy levels may reflect methylene-tetrahydrofolate reductase (MTHFR) enzyme polymorphisms, known to be higher in whites, rather than socioeconomic influences. PMID- 11279636 TI - Invasive pulmonary aspergillosis in neutropenic patients during hospital construction: before and after chemoprophylaxis and institution of HEPA filters. AB - Between September 1993 and December 1993, during extensive hospital construction and indoor renovation, a nosocomial outbreak of invasive pulmonary aspergillosis occurred in acute leukemia patients treated in a regular ward that has only natural ventilation. The observed infection rate was 50%. Chemoprophylaxis with intravenous continuous low-dose amphotericin B was then instituted as a preventive measure. During the next 18 months invasive pulmonary aspergillosis developed in 43% of acute leukemia patients. After that period a new hematology ward was opened with an air filtration system through high-efficiency particulate air filtration (HEPA) filters, and a bone marrow transplantation program was started on the hematology service. During the following three years, none of the acute leukemia or bone marrow transplantation patients who were hospitalized exclusively in the hematology ward developed invasive pulmonary aspergillosis, although 29% of acute leukemia patients who were housed in a regular ward, because of shortage of space in the new facility, still contracted invasive pulmonary aspergillosis. Overall, 31 patients were diagnosed with invasive pulmonary aspergillosis during almost five years: 74% of patients recovered from invasive pulmonary aspergillosis, and 42% are long-term survivors; 26% of patients died of resistant leukemia with aspergillosis, but no one died of invasive pulmonary aspergillosis alone. In conclusion, during an on-going construction period, an extremely high incidence rate of invasive pulmonary aspergillosis in acute leukemia patients undergoing intensive chemotherapy was observed. Institution of low-dose intravenous amphotericin B prophylaxis marginally reduced the incidence rate of invasive pulmonary aspergillosis. Keeping patients in a special ward with air filtration through a HEPA system eliminated invasive pulmonary aspergillosis completely. Among patients who developed invasive pulmonary aspergillosis, early diagnosis and treatment are probably the explanation for the favorable outcome. PMID- 11279637 TI - Frequency of the 677 C-->T mutation of the methylenetetrahydrofolate reductase gene among Kuwaiti sickle cell disease patients. AB - Sickle cell disease (SCD) is relatively mild among Kuwaiti Arabs. However, an atypical subset of patients exists with frequent, severe vaso-occlusive crisis and osteonecrosis. The thermolabile variant of MTHFR, resulting from a C-->T mutation at nucleotide 677, has been shown to be associated with hyperhomocysteinemia, which is an important risk factor for premature vascular disease. We have screened an unselected group of 41 Kuwaiti SCD patients (33 SS and 8 Sbeta(0)-thal) attending the Hematology Clinic of Kuwait University Teaching Hospital for the MTHFR mutation, using a PCR-RFLP method. The patients were aged 2-41 years (mean of 12.8 +/- 8.6). One (2.4%) individual was homozygous for the mutation while 15 (36.6%) were heterozygous, giving an allele frequency of 20.7%. Twenty-one patients (14 SS and 7 Sbeta(0)-thal) were screened for osteonecrosis using MRI of the hip (spin-echo T1- and T2-weighted images). Seven (33.3%) had varying degrees of osteonecrosis, among whom the frequency of the 677 C-->T allele was 21.4%. The frequency was identical among those without osteonecrosis. Although the allele frequency is higher among our patients compared to American SS patients, our results do not suggest an association with osteonecrosis. PMID- 11279638 TI - Lactate dehydrogenase production and release in a newly established human myeloma cell line. AB - Aggressive multiple myeloma with high serum lactate dehydrogenase (LDH) often has unusual clinical features and is considered to be a distinct clinical entity of multiple myeloma. A myeloma cell line, designated Maska-98, was established from the bone marrow of a patient with aggressive myeloma with extremely high serum LDH that was resistant to conventional chemotherapy. Maska-98 cells had morphological features of immature plasma cells, and immunophenotypic analysis showed that the cells expressed the plasma cell-associated surface antigens including CD38, 49d, and 56, but no T- or B-cell antigens, such as CD2, 3, 4, 8, 19, and 20. Maska-98 cells contained cytoplasmic immunoglobulin (IgG lambda). By utilizing this cell line we demonstrated that the myeloma cells produce and release a large amount of LDH, since (i) abundant LDH was found in the culture supernatant of Maska-98, (ii) immunocytochemical analysis showed that cytoplasm of the cells was strongly stained with anti-LDH monoclonal antibody, and (iii) Maska-98 cells expressed a greater amount of LDH mRNA than the T-cell line TALL 1, as shown by reverse transcription-polymerase chain reaction. As far as we know, there is no report of a myeloma cell line producing excess LDH. Therefore, Maska-98 would provide a novel source for further studies of the pathogenesis of aggressive multiple myeloma with high serum LDH. PMID- 11279639 TI - Arsenic trioxide- and idarubicin-induced remissions in relapsed acute promyelocytic leukaemia: clinicopathological and molecular features of a pilot study. AB - Arsenic trioxide (As2O3) effectively induces remissions in relapsed acute promyelocytic leukaemia (APL), but the safety of its long-term administration is unknown. The anthracycline idarubicin is highly active alone or in combination chemotherapy for the treatment of APL. To minimize arsenic exposure and based on the high sensitivity of APL cells to anthracyclines, we conducted a prospective study to evaluate induction with As2O3 followed by consolidation with idarubicin in the treatment of APL in relapse. Eight patients were treated with As2O3 at a daily dose of 10 mg until remission, followed by three monthly courses of idarubicin, at 6 mg/m(2)/day for 5 days in the first course and 6 mg/m(2)/day for 2 days in the subsequent two courses. All patients achieved morphological but not molecular remission after As2O3 treatment. During As2O3 therapy, an increase in white cell count peaking at a median of 17 days occurred in all the cases. Serial flow cytometric analysis of apoptosis, with mitochondrial APO2.7 antigen expression and the sub-G1 cell fraction on DNA histogram as markers, showed induction of apoptosis of APL cells in vivo. With both qualitative and real-time quantitative polymerase chain reaction, all patients were shown to attain molecular remission after subsequent idarubicin treatment. With a median follow up of 13 months, seven of eight patients have remained in complete clinical remission, with six patients in molecular remission as well. One patient who was in third remission became PCR-positive after being transiently negative. One patient died from an intracranial extramedullary relapse after achieving marrow molecular remission. We conclude that As2O3 induction followed by idarubicin consolidation is an effective therapy for APL in relapse. This regimen avoids the possible long-term toxicities of As2O3 and mutagenicity of combination chemotherapy, a strategy that might be suitable for this potentially curable leukaemia. PMID- 11279640 TI - Retrospective review of the management of elective surgery with desmopressin and clotting factor concentrates in patients with von Willebrand disease. AB - Limited data are available regarding optimal treatment with desmopressin (DDAVP) or intermediate-purity FVIII concentrates rich in VWF (CFCs) in patients with von Willebrand disease (VWD) who undergo planned surgery. We undertook a retrospective review over 10 years (1988-1997) and identified 27 patients treated with DDAVP for 35 surgical events and 38 patients who received CFCs for 68 elective surgical events. Tranexamic acid was usually added for mucosal surgery. The FVIII:C levels and the severity of surgery were used to determine the frequency and the doses of postoperative treatment. For major surgery the median pre- and post-operative doses of CFCs were 54 and 43 IU/kg, respectively, and for minor surgery the median doses varied between 34 and 52 IU/kg preoperatively and between 23 and 37 IU/kg postoperatively. The effectiveness of haemostasis was excellent in 32 events (91%) treated with DDAVP and in 56 events (82%) treated with CFCs. It is concluded that patients with VWD do not carry an increased operative risk if appropriate therapy is given. PMID- 11279641 TI - Recurrent leg ulcers and arterial thrombosis in a 33-year-old homozygous variant of antithrombin. AB - We report here a homozygous variant case of antithrombin (AT) associated with arterial thrombosis and recurrent leg ulcers. The deep vein thrombosis was recognized by the venogram of his pelvic veins. His leg ulcers were scattered around his left ankle and accompanied by lipodermatosclerosis, which was evident in venous insufficiency. The propositus had developed cerebral infarction 12 years prior to his leg ulcers. Coagulation study showed low heparin cofactor activity of his AT with a normal level of immunoreactive AT. Nucleotide sequence analysis of the exon 2 of his AT gene showed Arg47-Cys mutation, leading to the lack of affinity of AT for heparin. The propositus is a homozygote for this abnormality. PMID- 11279642 TI - Successful treatment with recombinant factor VIIa of therapy-resistant severe bleeding in a patient with acquired von Willebrand disease. AB - We describe an elderly man who presented with life-threatening hematuria and gastrointestinal bleeding caused by acquired von Willebrand disease associated with monoclonal gammopathy of undetermined significance (MGUS). Standard therapy with desmopressin, von Willebrand factor-containing factor VIII concentrate, tranexamic acid, and immunoglobulin failed to achieve adequate hemostasis. However, treatment with recombinant activated factor VII (rFVIIa) arrested the bleeding completely. Since acquired von Willebrand disease can lead to life threatening hemorrhage, clinicians should consider rFVIIa as an effective treatment option if standard therapy fails. PMID- 11279643 TI - Paris-Trousseau syndrome platelets in a child with Jacobsen's syndrome. AB - The thrombocytopenia in an infant with clinical features of Jacobsen's syndrome characterized by multiple congenital anomalies, cardiac defects, psychomotor retardation, and deletion of chromosome 11 at 11q23.3 has been evaluated. Study of his platelets in the electron microscope revealed giant alpha granules in his cells identical in appearance to those reported in the family with Paris Trousseau syndrome. As a result, the Paris-Trousseau syndrome appears to be a variant of the Jacobsen syndrome, and the thrombocytopenia observed in all cases of chromosome 11q23.3 deletion due to dysmegakaryopoieses. Giant alpha granules are frequently observed in normal platelets during long-term storage and may form in Jacobsen and Paris-Trousseau platelets during prolonged residence in the bone marrow. PMID- 11279644 TI - Thalidomide-associated hepatitis: a case report. AB - We report a case of hepatitis in a 58-year-old woman being treated with thalidomide for end-stage plasma cell leukemia. The patient had a medical history including chronic stable hepatitis C infection. At diagnosis there was a severe anemia, thrombocytopenia, hypercalcemia, IgG paraproteinemia, peripheral blood myeloma cells, and a marrow plasmacytosis with lytic bony lesion. The disease was refractory to standard chemotherapy, and she was treated with oral thalidomide. Within 1 week she became jaundiced and developed a marked transaminitis. This promptly resolved upon cessation of thalidomide alone. Thalidomide has recently enjoyed renewed interest as a treatment in many disorders, including plasma cell leukemia. To our knowledge, this is the first reported case of thalidomide associated hepatotoxicity. Although the mechanism of its actions on the liver are uncertain, it is possible that thalidomide acts as a direct hepatotoxin or as an immuno-modulator, altering the activity of chronic viral hepatitis. We present this case to increase awareness of a new potential side effect of thalidomide as its clinical indications expand. PMID- 11279646 TI - Lenograstim and filgrastim effects on neutrophil motility in patients undergoing chemotherapy: evaluation by computer-assisted image analysis. PMID- 11279647 TI - Positive predictive values of imaging studies used before accessory splenectomy. PMID- 11279645 TI - Acute adrenal failure associated with fluconazole after administration of high dose cyclophosphamide. AB - A 63-year-old man received high-dose cyclophosphamide for peripheral blood stem cell (PBSC) harvest. He received 200 mg fluconazole. On day 3, atrial fibrillation developed with blood pressure declining to 78 mmHg. The rapid adrenocorticotropin (ACTH) test showed blunted adrenal responses. He was suspected as having adrenal failure, and fluconazole was discontinued. The rapid ACTH test became normal on Day 14, and PBSCs were successfully harvested. To clarify the association between adrenal failure and fluconazole, we resumed 400 mg fluconazole on Day 16 and repeated the test on Day 21, which showed blunted adrenal responses. This case demonstrates that prophylactic use of fluconazole can cause adrenal insufficiency. PMID- 11279648 TI - Symptomatic presentation of a sickle cell heterozygote: an evaluation of genetic factors. PMID- 11279649 TI - Clinical and pathologic findings in 52 consecutively autopsied cases with multiple myeloma. AB - We studied clinical features and pathologic findings in 52 consecutively autopsied patients with multiple myeloma in our center between 1979 and 1998. Distant extraosseous involvement was found in 33 patients (63.5%). Thirty-one patients (59.6%) were proven to have infection at autopsy, among which pneumonia was most common site of infection. Amyloidosis was shown in 8 patients. Second malignancies were observed in 4 cases. The three major causes of death were hemorrhage, infection, and renal failure, which accounted for death in approximately 70% of the patients. Advances in the anticancer and antimicrobial chemotherapies might have decreased deaths due to myeloma itself or infection. PMID- 11279650 TI - Cell fusion is not involved in the generation of giant cells in the Hodgkin-Reed Sternberg cell line L1236. AB - The mechanism of multinucleated cell formation in Hodgkin's disease has not yet been elucidated. We asked whether the giant multinucleated cells of the H-RS cell line L1236 develop via fusion of the predominant smaller cells. As a positive control for the fusion assay, human B cells from the B-cell lymphoma cell line BJA-B were split into two fractions, stained with the fluorochromes CMTMR and CMFDA, respectively, and fused using the polyethylene glycol 1500 cell hybridization protocol. Double-stained cells indicating fusion of BJA-B cells were detectable for up to 5 days. In parallel, L1236 cells were split into two fractions, stained with the fluorochromes, and mixed. No double-stained L1236 cells were detected. The same result was obtained when using FACS-sorted small mononuclear L1236 cells. It is thus concluded that the large multinucleated cells of the monoclonal H-RS cell line L1236 have emerged by endomitosis rather than by spontaneous cell fusion. PMID- 11279651 TI - Home treatment of deep venous thrombosis with low molecular weight heparin: Long term incidence of recurrent venous thromboembolism. AB - Outpatient treatment of deep venous thrombosis (DVT) with low molecular weight heparin (LMWH) seems as safe and effective as inpatient treatment with unfractionated heparin (UFH). However, most of the randomized trials comparing a LMWH with UFH described clinical outcomes within 3-6 months. The long-term incidence of recurrent VTE after treatment of DVT with LMWH remains to be established. The primary objective of this retrospective study was to document the long-term incidence of recurrent venous thromboembolism (VTE) in patients with DVT treated with a LMWH, nadroparin in an outpatient basis. The patients were evaluated 46 months after inclusion in two cohorts comparing home treatment with nadroparin (n = 130) with in-hospital treatment with intravenous UFH (n = 149). More than 60% of the patients in the nadroparin group could be treated at home, either entirely or after a short stay in hospital. The age-adjusted thrombosis-free survival was not statistically significant between nadroparin and UFH-treated patients (P = 0.084). There was a nonsignificant trend favoring nadroparin as compared with UFH. The hazard ratio (HR) for recurrent VTE in the nadroparin group with respect to the UFH group was 0.44 (95% confidence interval, 0.17-1.12). No significant differences were observed in overall mortality or major hemorrhage between the two treatment groups. Our study suggests that home treatment of DVT with LMWH is at least as effective and safe as in-hospital UFH after a long-term follow-up period. PMID- 11279652 TI - Hypercoagulability states in upper-extremity deep venous thrombosis. AB - Deep venous thrombosis of the upper extremity (DVTUE) is a rare thrombotic disorder that may occur spontaneously but is most often related to predisposing factors, such as an indwelling central venous catheter, malignancy, or exercise. The role of coagulation disorders, i.e., a hypercoagulable state in the pathogenesis of DVTUE is not well known. We have evaluated both genetic and acquired thrombophilia parameters in consecutive patients with DVTUE. A hypercoagulable state was found in 32% of the patients. The most frequent coagulation abnormality was the presence of lupus anticoagulant or anticardiolipin antibodies (27%). Factor V Leiden mutation was detected in two patients, antithrombin deficiency in one, and none of the patients had the prothrombin G20210A gene variant or protein C or S deficiency. The prevalence of coagulation abnormalities was not significantly different in a subgroup of patients with spontaneous DVTUE as compared to those with an obvious predisposing factor, such as an indwelling central venous catheter. We conclude that antiphospholipid antibodies are frequently found in patients with DVTUE. Factor V Leiden mutation, prothrombin 20210A gene variant, protein C deficiency, and protein S deficiency do not seem to play a major pathogenetic role in DVTUE. PMID- 11279653 TI - Proximal promoter of the murine syndecan-1 gene is not sufficient for the developmental pattern of syndecan expression in B lineage cells. AB - Syndecan-1 (CD138) is a cell membrane proteoglycan that binds extracellular matrix components and various growth factors. The role of syndecan-1 in the control of cell growth and morphology has been illustrated by its altered expression in hematological malignancies such as multiple myeloma as well as some solid tumors. It has been reported that the expression of syndecan-1 in cells of the B lineage is developmentally regulated such that pre-B cells and plasma cells express syndecan-1 while mature B cells do not. Thus, we investigated whether the proximal promoter region of the murine syndecan-1 promoter was able to confer the observed on-off-on expression of syndecan-1 in cells of the B lineage as they develop from pre-B cells to plasma cells. Experiments carried out using deletion mutants of the proximal promoter cloned upstream of the CAT reporter gene transfected into murine cell lines, representing the above stages of B-cell development, such as BA/F3 (pro-B cell), 70Z/3 (pre-B cell), 2PK3 (late mature B cell), and MPC-11 (plasma cell), showed detectable levels of CAT expression. The WEHI-231 (mature B cell) cell lines did not show detectable levels of CAT reporter activity. The strong levels of expression were observed with a fragment of the proximal promoter spanning the region from -365 to -95 (from the translation start point). However, Northern analysis of RNA obtained from the five murine B-cell lines, representing various stages of B-cell development, showed that the 70Z/3, MPC-11 but not BA/F3, and 2PK3 cells expressed detectable levels of syndecan-1 mRNA. By FACS analysis, using a rat anti mouse syndecan-1 antibody, syndecan-1 expression on the cell surface was found to correlate with the observed mRNA expression patterns in these cell lines. Our results indicate that the proximal promoter of the murine syndecan-1 promoter is not sufficient for the observed developmental pattern of syndecan expression in B cells. PMID- 11279654 TI - Do the acute platelet responses of patients with immune thrombocytopenic purpura (ITP) to IV anti-D and to IV gammaglobulin predict response to subsequent splenectomy? AB - The acute platelet response to Intravenous Gammaglobulin (IVIG) has been reported to predict response to subsequent splenectomy of patients with ITP. The current study was undertaken to determine if the platelet response to IV anti-D (Winrho SDF) predicts response to subsequent splenectomy. The 61 HIV-uninfected children and adults in this study had taken part in the pre-licensing studies of IV anti-D and were all those who not only had evaluable platelet responses to IV anti-D but also had undergone splenectomy and had information available describing its 1 year outcome. Results of treatment with IVIG were available in 38 of these 61 patients. Neither response to the initial infusion of IV anti-D, nor response to the initial or last IVIG, predicted the response in either children or adults to subsequent splenectomy. However, response to the last anti-D infusion in adults was strongly correlated (P = 0.003) to response to subsequent splenectomy as was hemolysis >/=2.0 gm/dl after IV anti-D (P = 0.03). There was no overall relationship between response to IV anti-D or IVIG, and response to subsequent splenectomy. However, a good platelet response in adults to the last IV anti-D and a hemoglobin decrease >/=2.0 gm/dl both appeared to predict response to subsequent splenectomy. PMID- 11279655 TI - CD34-positive acute promyelocytic leukemia is associated with leukocytosis, microgranular/hypogranular morphology, expression of CD2 and bcr3 isoform. AB - Acute promyelocytic leukemia (APL) has a favorable prognosis. Current therapy includes chemotherapy used in combination with all-trans-retinoic acid (ATRA). Although the differentiating effects of ATRA on promyelocytes have been well established, in vitro studies have shown that less-differentiated APL blasts (CD34(+)) demonstrate a variable responsiveness to ATRA. To assess the clinical relevance of this finding, we analyzed a cohort of 38 patients with t(15;17) and/or PML-RARalpha APL to determine the incidence and laboratory features of CD34(+) APL. Thirty-two percent (12/38) of cases were CD34(+). There was a difference in WBC at presentation between CD34(+) and CD34(-) cases (34.6 +/- 9.2, mean +/- standard error vs. 5.4 +/- 2.0, P = 0.009). Patients with CD34(+) APL demonstrated a micro/hypogranular phenotype (75%) (P = 0.001), co-expression of CD2(+) (83%) (P = 0.001), and the bcr3 isoform (100%) (P = 0.017). In contrast, CD34(-) cases demonstrated hypergranular morphology (65%), CD2(+) (15%), and the bcr1 isoform (50%). A high presenting WBC count (?G10 x 10(9)/L) was associated with an inferior overall survival (Log rank = 0.0047). Patients with CD34(+) APL demonstrated an incidence of early mortality of 50%. Despite a marked correlation between CD34 positivity and increased WBC count, overall survival of CD34(+) and CD34(-) cases did not differ significantly in our small cohort. Immunophenotypic analysis for CD34 expression should be included in future large APL trials to determine if detection of CD34(+) blasts represents an independent adverse prognostic factor. PMID- 11279656 TI - Correlation of hematopoietic progenitor cell count determined by the SE-automated hematology analyzer with CD34(+) cell count by flow cytometry in leukapheresis products. AB - The yield of stem cell collection after mobilization is crucial for autologous peripheral blood stem cell (PBSC) transplantation. Quantitative determinations of CD34(+) cells using flow cytometry or stem cell culture have been used, but these methods require much time, technical experience, and expensive reagents. The automated hematology analyzer (Sysmex SE-9000trade mark, TOA, Japan) equipped with the Immature Information (IMI) channel for immature myeloid cells can detect IMI(+) cells within 90 sec. Detection is made possible by the combination of a special reagent system and direct current/radiofrequency biosensors. We studied the relation of IMI(+) cells and variable cell counts with CD34(+) cell yield in autologous stem cell harvest. In a series of 32 patients (median age, 44 years; M:F = 11:21), 184 leukaphereses were performed after mobilization regimens with chemotherapy and G-CSF or G-CSF alone. Full blood cell counts were enumerated on peripheral blood (PB) samples taken prior to each leukapheresis. Mononuclear cell (MNC) and IMI(+) cell counts by automated hematology analyzer and flow cytometry based CD34(+) cell yield were measured on the harvested product. The relationship among PB white blood cells (WBC), PB monocytes, IMI(+) cells, MNC, and CD34(+) cell yield in a single leukapheresis was estimated by Pearson correlation analysis. PB WBC count showed no correlation with CD34(+) cell yield in a single leukapheresis (r = 0.02, P = 0.81). PB monocyte count showed a weak correlation (r = 0.21, P = 0.01) and MNC in harvest also showed a weak correlation (r = 0.36, P = 0.0001) with CD34(+) cell yield. In contrast, CD34(+) cell yield correlated well with IMI(+) cell count (r = 0.68, P = 0.0001), and data could be fitted by a linear regression equation, y = 0.330 + 0.974x. IMI(+) cell assay by the automated hematology analyzer correlated well with the CD34(+) cell yield in a mobilized autologous stem cell harvest. The IMI(+) cell count might be used as a simple and efficient indicator of blood stem cell mobilization and collection. PMID- 11279657 TI - Burkitt's lymphoma of stomach: A case report and review of literature. AB - Primary gastrointestinal non-Hodgkin's lymphoma is a distinct clinicopathological entity. This is the commonest site of all extranodal lymphomas. Non-endemic Burkitt's lymphoma (high-grade, small non-cleaved lymphocytic type) is rare in non-HIV adult population. We hereby report Burkitt's lymphoma of stomach in a non HIV adult. Gastric and lymph node biopsy confirmed the diagnosis. Patient was treated using a third generation chemotherapy protocol without any surgical resection. The patient relapsed within 3 months of completion of primary treatment and died of progressive disease. PMID- 11279659 TI - Excessive anticoagulation in patients with mild renal insufficiency receiving long-term therapeutic enoxaparin. AB - Low-molecular-weight heparins, such as enoxaparin, are increasingly being used for treatment of venous thromboembolism. We describe two patients who received therapeutic enoxaparin for several months. Although their serum creatinine values were normal, both had mild renal insufficiency (creatinine clearance 60-70 ml/min), and both accumulated the drug abnormally and experienced clinical bleeding. These results suggest that patients receiving enoxaparin (or other low molecular-weight heparins) in therapeutic doses for periods of more than 4 weeks should be considered for laboratory monitoring to avoid bleeding. PMID- 11279658 TI - Reversible sideroblastic anemia associated with the tetracycline analogue COL-3. AB - Eight of 35 patients with cancer receiving COL-3, a tetracycline derivative with antiangiogenic properties, developed anemia while on treatment. All of these patients were enrolled on an approved Phase I clinical trial at the National Cancer Institute. Three of these patients had bone marrow examinations that revealed ringed sideroblasts. This paper describes these cases. Am. J. Hematol. 67:51-53, 2001. Published 2001 Wiley-Liss, Inc. PMID- 11279660 TI - A novel beta-thalassemia intermedia phenotype containing Nt494+129T-->C and NT494+132C-->A mutations in cis and a Nt168C-->T (beta(o) 39 point) mutation in trans. PMID- 11279661 TI - Normal delta globin gene sequence in carrier of the silent-101 (C-T) beta thalassemia mutation with normal HbA2 level. PMID- 11279662 TI - Multiple complications of IVIG therapy in a patient with Guillain-Barre syndrome. PMID- 11279663 TI - Refractory autoimmune thrombocytopenic purpura treatment with Rituximab. PMID- 11279665 TI - Sensory and endocrine characteristics of the avian pineal organ. AB - The avian pineal organ represents a transitional type between a photosensory organ of lower vertebrates and the endocrine gland of mammals and shows remarkable changes in its innervation and structure during ontogeny. In the avian pineal organ the progressive reduction of the pinealofugal component and the spectacular increase in pinealopetal sympathetic innervation occur in parallel. In domestic fowl the number of intrapineal AChE-positive (afferent) neurons decreases rapidly during ontogenetic development, whereas the sympathetic innervation becomes more prominent. Furthermore, the end vesicle of the pineal organ is an anatomical entity fully separated from the brain in the adult domestic fowl, as observed in some mammalian pineals. The avian pineal organ contains several types of photoreceptors with different photopigments and the synthesis of melatonin, the pineal hormone, is controlled by light. Immunoreactivity for photopigments is reduced during the posthatching development of chicken, whereas neuron-specific enolase (NSE)-immunoreactive pinealocytes increase remarkably in number in the end-vesicle of the domestic fowl with age, followed by a gradual expansion toward the proximal portion. NSE is the most acidic isoenzyme of the glycolytic enzyme enolase and is useful as a cytoplasmic marker of neurons and neuroendocrine tissue. The above-mentioned findings reflect the sequence of changes leading from pineal sense organs to pineal gland. The demonstration of melatonin receptors in a variety of avian peripheral tissues suggest a possible direct action of melatonin on the physiological functions of different organ systems in response to internal and external stimuli. PMID- 11279666 TI - Comparative ultrastructure and cytochemistry of the avian pineal organ. AB - The breeding of birds is expected to solve problems of nourishment for the growing human population. The function of the pineal organ synchronizing sexual activity and environmental light periods is important for successful reproduction. Comparative morphology of the avian pineal completes data furnished by experiments on some frequently used laboratory animals about the functional organization of the organ. According to comparative histological data, the pineal of vertebrates is originally a double organ (the "third" and the "fourth eye"). One of them often lies extracranially, perceiving direct solar radiation, and the other, located intracranially, is supposed to measure diffuse brightness of the environment. Birds have only a single pineal, presumably originating from the intracranial pineal of lower vertebrates. Developing from the epithalamus, the avian pineal organ histologically seems not to be a simple gland ("pineal gland") but a complex part of the brain composed of various pinealocytes and neurons that are embedded in an ependymal/glial network. In contrast to organs of "directional view" that develop large photoreceptor outer segments (retina, parietal pineal eye of reptiles) in order to decode two-dimensional images of the environment, the "densitometer"-like pineal organ seems to increase their photoreceptor membrane content by multiplying the number of photoreceptor perikarya and developing follicle-like foldings of its wall during evolution ("folded retina"). Photoreceptor membranes of avian pinealocytes can be stained by antibodies against various photoreceptor-specific compounds, among others, opsins, including pineal opsins. Photoreceptors immunoreacting with antibodies to chicken pinopsin were also found in the reptilian pineal organ. Similar to cones and rods representing the first neurons of the retina in the lateral eye, pinealocytes of birds possess an axonal effector process which terminates on the vascular surface of the organ as a neurohormonal ending, or forms ribbon-containing synapses on pineal neurons. Serotonin is detectable immunocytochemically on the granular vesicles accumulated in neurohormonal terminals. Pinealocytic perikarya and axon terminals also bind immunocytochemically recognizable excitatory amino acids. Peripheral autonomic fibers entering the pineal organ through its meningeal cover terminate near blood vessels. Being vasomotor fibers, they presumably regulate the blood supply of the pineal tissue according to the different levels of light dependent pineal cell activity. PMID- 11279667 TI - Comparative view of pineal gland morphology of nocturnal and diurnal birds of tropical origin. AB - Although having a similar developmental pattern, the pineal gland of tropical birds varies in shape, size, and morphology, probably more than any other part of the avian brain. Following the old classification, we noted a solid follicular (transitional) type of the pineal gland in the nocturnal bird Athene brama, and a tubulo-follicular and elongated tubular types of pineal gland in diurnal birds Perdicula asiatica and Euroloncha punchulata, respectively. Detailed light (LM) and electron microscopic (EM) studies of the pineal gland from these tropical birds revealed the presence of a well-developed, functionally active gland in nocturnal birds (contrary to reports available until now). Unlike diurnal birds, the nocturnal bird A. brama has no deep pineal in the posterior region (near the habenular commissure). It could be that the deep encephalic receptors have no/fewer functions in nocturnal birds. At present, we were unable to define the significance of deep pineal in these tropical avian species. A notable difference in the proximodistal orientation of intrapineal follicles and parenchymatous cells was noted among these birds due to different habitats. Ultrastructurally, the pinealocytes exhibited great similarities in terms of secretory organelles, except for the presence of some peculiar membranous structure in E. punchulata. The pinealocytes have rudimentary photoreceptive features (e.g., outer segment) along with cytoplasmic organelles for secretory activity, suggesting both photosensory and photosecretory types of function. The present study also suggests more heterogenicity in pineal gland morphology (cellular architecture) among diurnal birds than the nocturnal one. PMID- 11279668 TI - Developmental potency of cultured pineal cells: an approach to pineal developmental biology. AB - The pineal organ is still an enigma in regard to its developmental and phylogenetic origin. Little is known of the mechanism involved in determination and differentiation of pineal cells and virtually no studies have been done on the induction and tissue interactions during pinealogenesis. Interest is also centered on the evolutional transformation in structure and function, which may be related to the developmental alterations in pineal morphogenesis between the lower and higher vertebrate species. For developmental studies, avian embryos have great advantages for various experimental manipulations, such as cell and organ culture, surgical operation, and in situ transfection of developmental genes. The present review describes our cell culture studies, which have been done on developing rat and quail pineal organs, in order to elucidate the developmental potency of pineal cells and the regulatory mechanism involved in the phenotypic expression of cell properties. A number of phenotypes including numerous neuron-specific substances are shown immunohistochemically to be expressed only under culture conditions, and not observed in the mature pineal organ. As development proceeds, some of the potencies for cell differentiation are lost; hence, in the mature pineal organs most neuronal phenotypes are not expressed. Numerous factors were discovered which affect phenotypic expression of cultured pineal cells in a cell-type-specific manner. These findings, together with immunohistochemical observations on developing pineal organs, reveal that the developing pineal organ is a unique and useful model system for developmental neurobiology and that cell culture techniques offer a powerful tool for the understanding of development and cell differentiation of this particular organ. PMID- 11279669 TI - Multiphotoreceptor and multioscillator system in avian circadian organization. AB - Photoperiodism and circadian rhythms have been studied intensively in birds because Aves are typical seasonal breeders and diurnal animals. Light is the most important environmental factor involved in entrainment of circadian rhythms and photoperiodism. The eyes and the extraocular photoreceptors, such as the pineal organ and hypothalamus, are reported to have an important function not only for photoreception but also for circadian organization in nonmammalian vertebrates, including birds. In this report, we review the roles of the eyes, pineal organ, and deep brain as the components of the multiphotoreceptor and multioscillator system in avian circadian organization. PMID- 11279670 TI - Circadian organization and the role of the pineal in birds. AB - All organisms exhibit significant daily rhythms in a myriad of functions from molecular levels to the level of the whole organism. Significantly, most of these rhythms will persist under constant conditions, showing that they are driven by an internal circadian clock. In birds the circadian system is composed of several interacting sites, each of which may contain a circadian clock. These sites include the pineal organ, the suprachiasmatic nucleus (SCN) of the hypothalamus, and, in some species, the eyes. Light is the most powerful entraining stimulus for circadian rhythms and, in birds, light can affect the system via three different pathways: the eyes, the pineal, and extraretinal photoreceptors located in the deep brain. Circadian pacemakers in the pineal and in the eyes of some avian species communicate with the hypothalamic pacemakers via the rhythmic synthesis and release of the hormone melatonin. Often the hypothalamic pacemakers are unable to sustain persistent rhythmicity in constant conditions in the absence of periodic melatonin input from the pineal (or eyes). It has also been proposed that pineal pacemakers may be unable to sustain rhythmicity in constant conditions without periodic neural input from the SCN. Significant variation can occur among birds in the relative roles that the pineal, the SCN, and the eyes play within the circadian system; for example, in the house sparrow pacemakers in the pineal play the predominant role, in the pigeon circadian pacemakers in both the pineal and eyes play a significant role, and in Japanese quail ocular pacemakers play the predominant role. PMID- 11279671 TI - Unraveling the enigma: the role of melatonin in seasonal processes in birds. AB - Birds, unlike mammals, do not use the annual profile of pineal melatonin secretion to coordinate their reproductive efforts with a favorable time of year. Melatonin in birds mediates the entrainment of circadian activity rhythms, and thus helps to time hatching of eggs and facilitate migration. However, the role of melatonin as a reliable indicator of day length for seasonal processes has remained equivocal for many years. Recently, the influence of melatonin on two physiological processes involved in aspects of seasonal reproduction has been identified in European starlings: 1) the regulation of seasonal changes in immune function, and 2) neuroplasticity in the song control system. Melatonin can enhance cell-mediated immune function and acts as an inhibitory hormone on the song control system. Melatonin receptor (MelR) density in a forebrain song control nucleus, Area X, is regulated as a function of reproductive state; there is marked downregulation of MelR in Area X during the breeding season in starlings. Seasonal regulation of immune function and neural plasticity within the song control system, and the efficacy of the action of melatonin on these two processes, appears to be modified by the same central, thyroid-dependent mechanism that controls the reproductive state of birds. These data indicate that the interaction of day length and hormones of different classes affects the ability of melatonin to affect seasonal processes in birds. The downstream consequences of MelR regulation within the song control system are discussed with regard to the cellular action of melatonin and its possible interaction with immediate-early genes and transcription factors. PMID- 11279672 TI - Photoreception and circadian clock system of the chicken pineal gland. AB - Chicken pinealocytes contain three major components of the circadian clock system: 1) a self-sustained oscillator, 2) a photic-input pathway to the oscillator, and 3) an overt output represented by the rhythmic production of melatonin. Even under cultured conditions of isolated pineal gland or dissociated pinealocytes, the input-oscillator-output functions are well maintained. Because of these experimental advantages, chicken pineal gland has been one of the best models for the study of the circadian clock system. Since the finding of a pineal specific photoreceptive molecule, pinopsin, we have characterized the endogenous phototransduction pathway in the pinealocytes. On the other hand, despite the long history of chick pineal research, the molecular mechanism underlying the pineal clock oscillation has been largely unknown. Our recent characterization of the chick pineal clock genes strongly suggests that they constitute a transcription/translation-based autoregulatory feedback loop, which is very similar to that generating circadian rhythmicity in mammalian SCN. PMID- 11279673 TI - Development of scoliosis following pinealectomy in young chickens is not the result of an artifact of the surgical procedure. AB - Pinealectomy in young chickens consistently results in scoliosis, which has many characteristics similar to those seen in adolescent idiopathic scoliosis. The mechanism underlying this phenomenon remains a mystery and it is not yet entirely clear whether some unidentified aspect of the extensive surgery is the major factor rather than the removal of the pineal gland. Four different types of pinealectomy surgery were performed on young chickens as well as deliberate damage to the cerebral cortex which simulated the extreme of any accidental damage that might occur during surgery. Scoliosis was assessed from weekly radiographs. No differences in incidence of scoliosis, degree of severity, or pattern of curve development were observed for any of the experimental groups when compared with controls. In all groups approximately 55% of the chickens developed scoliosis that progressed rapidly. Different pinealectomy procedures and deliberate damage to the cerebral cortex produce scoliosis in young chickens with the same incidence and characteristics. This suggests strongly that the mechanism behind the phenomenon is due to the removal of the pineal gland and not some artifact of the extensive surgery. The pinealectomy model in young chickens is proving to be a good model for studying AIS in humans. An understanding of the mechanism underlying this phenomenon has the potential to provide further insights into the etiology of AIS and can lead to the development of novel treatment methods. PMID- 11279674 TI - Contribution of cryofixation and freeze-substitution to analytical microscopy: a study of Tritrichomonas foetus hydrogenosomes. AB - The hydrogenosome, an organelle that produces molecular hydrogen and ATP from the oxidation of pyruvate or malate under anaerobic conditions, presents some characteristics common to mitochondria. It is found in several trichomonad species, protists living in oxygen-poor environments, as well as certain free living ciliates, rumen ciliates, and some fungi. We performed a comparative microanalytical study (energy dispersive X-ray analysis and electron spectroscopic imaging) of different fixation methods for electron microscopy analyzing hydrogenosomes of the bovine parasite Tritrichomonas foetus. The study included the elemental composition and the mapping of calcium, phosphorus, and oxygen. A preparation of T. foetus cells, based on cryoimmobilization by high pressure freezing and freeze-substitution, was compared to a second preparation based on chemical fixation followed by dehydration and routine processing. The ultrastructural preservation achieved by the cryotechnique was far superior to the chemical fixation, since it allowed the successful cryoimmobilization of intracellular ion contents. The detection of several cations (Al, Mg, Co, Ca, Fe) by X-ray microanalysis inside the peripheral vesicle of the hydrogenosome was only possible in cryofixed cells. The presence of aluminum and cobalt ion in the hydrogenosomal vesicle was established for the first time. Electron-spectroscopic images of calcium showed that this element, in addition to the vesicle compartment, is present in the hydrogenosome's membrane in varying concentrations, which might reflect changes in the physiology of this organelle. PMID- 11279675 TI - Antipsychotic medication for those with both schizophrenia and learning disability. AB - BACKGROUND: Antipsychotic medication is the standard treatment for people with learning disability and schizophrenia. OBJECTIVES: To determine the efficacy of any antipsychotic medication for treating people with a dual diagnosis of learning disability and schizophrenia. SEARCH STRATEGY: Electronic searching of Biological Abstracts, the Cochrane Schizophrenia Group's Register of trials (September 2000), the Cochrane Library, EMBASE, PsycLIT MEDLINE and National Research Register (Issue 3 2000). Unpublished data were sought from pharmaceutical companies. Both authors independently selected the relevant studies from the reports identified in this way. SELECTION CRITERIA: 1. All randomised controlled trials of antipsychotic medication, regardless of dosage, versus a placebo control, of longer than one month's duration. 2. Anyone over 18 years of age with both learning disability and schizophrenia. Learning disability was defined as a measured IQ of 70 or less. Any mode of diagnosis of schizophrenia was acceptable. DATA COLLECTION AND ANALYSIS: The two reviewers independently attempted to select and then extract data but it was not possible to do this with the single study that met the inclusion criteria. MAIN RESULTS: Only one relevant randomised trial was found by the searches. This study included four people with a dual diagnosis of schizophrenia and learning disability, but results were available for only two. The groups to which the other two people were allocated were unclear. In order to display the data, too many assumptions would have to have been made about these other two people and any results would be uninformative and potentially misleading. REVIEWER'S CONCLUSIONS: Using the methods described the reviewers found no randomised controlled trial evidence to guide the use of antipsychotic medication for those with both learning disability and schizophrenia. Until the urgent need for randomised controlled trials is met, clinical practice will continue to be guided by extrapolation of evidence from randomised controlled trials involving people with schizophrenia but without learning disability and non-randomised trials of those with learning disability and schizophrenia. PMID- 11279676 TI - Elective versus selective caesarean section for delivery of the small baby. AB - BACKGROUND: Elective caesarean delivery for women in preterm labour might reduce the chances of fetal or neonatal death, but it might also increase the risk of maternal morbidity. OBJECTIVES: To assess the effects of a policy of elective caesarean delivery versus selective caesarean delivery for women in preterm labour. SEARCH STRATEGY: The Cochrane Pregnancy and Childbirth Group trials register was searched. Date of last search: September 2000. SELECTION CRITERIA: Randomised trials comparing a policy of elective caesarean delivery versus expectant management with recourse to caesarean section. DATA COLLECTION AND ANALYSIS: One reviewer assessed eligibility and trial quality, and both contributed to the update. MAIN RESULTS: Six studies involving 122 women were included. All trials reported recruiting difficulties. No significant differences between elective and selective policies for caesarean delivery were found for fetal, neonatal or maternal outcomes. REVIEWER'S CONCLUSIONS: There is not enough evidence to evaluate the use of a policy for elective caesarean delivery for small babies. Randomised trials in this area are likely to continue to experience recruitment problems. However, it still may be possible to investigate elective caesarean delivery in preterm babies with cephalic presentations. PMID- 11279677 TI - Hospitalisation and bed rest for multiple pregnancy. AB - BACKGROUND: Bed rest used to be widely advised for women with a multiple pregnancy. OBJECTIVES: The objective was to assess the effect of bed rest in hospital for women with a multiple pregnancy for prevention of preterm birth and other fetal, neonatal and maternal outcomes. SEARCH STRATEGY: The Cochrane Pregnancy and Childbirth Group trials register, the Cochrane Controlled Trials Register and reference lists of relevant articles were searched. Date of last search: August 2000. SELECTION CRITERIA: Randomised trials which compare outcomes in women with a multiple pregnancy and their babies who were offered bed rest in hospital with women only admitted to hospital if complications occurred. DATA COLLECTION AND ANALYSIS: Assessment for inclusion and methodological quality of the trials was done by the reviewer. Data were extracted by the reviewer and double entered. All eligible trials were included in the initial analysis. Prespecified sensitivity analyses have been carried out to evaluate the effect of trial quality, the effects of hospitalisation for bed rest in women with an uncomplicated twin pregnancy, in women with a triplet pregnancy and in women with a twin pregnancy complicated by cervical effacement and dilatation prior to labour. MAIN RESULTS: Six trials were included which involved over 600 women and 1400 babies. (1) Analyses of all trials. Routine bed rest in hospital for multiple pregnancy did not reduce the risk of preterm birth, or perinatal mortality. There was a trend to a decreased number of low birth weight infants born to women in the routinely hospitalised group, which became significant when the trial using alternate allocation was excluded (odds ratio (OR) 0.79; 95% confidence interval (CI) 0.63-0.99). No differences were seen in the number of very low birth weight infants. No support for the policy was found in other neonatal outcomes. No information is available on developmental outcomes for infants in any of the trials. Women's views about the care they received were reported rarely. (2) Analyses of hospitalisation for bed rest in women with an uncomplicated twin pregnancy. The risk of preterm birth was not reduced. Indeed significantly more women gave birth very preterm (< 34 weeks gestation) (OR 1.84; 95% CI 1.01-3.34). No differences were seen in perinatal mortality, or in other neonatal outcomes. Women receiving hospitalisation for bed rest had a decreased risk of developing hypertension (OR 0.55; 95% CI 0.32-0.97), although this effect was no longer apparent when the trial using alternate allocation was excluded. (3) Analyses of hospitalisation for bed rest in women with a triplet pregnancy. Most of the comparisons made between the hospitalised and control groups suggest beneficial treatment effects from routine hospitalisation for bed rest. However all the differences observed between the experimental and control groups were compatible with chance variation. (4) Analyses of hospitalisation for bed rest in women with a twin pregnancy complicated by cervical effacement and dilatation prior to labour. No differences were seen in the risk of preterm birth, perinatal mortality, fetal growth or in other neonatal outcomes. REVIEWER'S CONCLUSIONS: There is currently not enough evidence to support a policy of routine hospitalisation for bed rest in multiple pregnancy. No reduction in the risk of preterm birth or perinatal death is evident, although there is a suggestion that fetal growth is improved. For women with an uncomplicated twin pregnancy the results of this review suggest that it may be harmful in that the risk of very preterm birth is increased. Until further evidence is available to the contrary, the policy cannot be recommended for routine clinical practice. PMID- 11279678 TI - Early light reduction for preventing retinopathy of prematurity in very low birth weight infants. AB - BACKGROUND: Retinopathy of prematurity (ROP) causes vision loss in many premature infants each year, despite the advances being made in treatment. In the search for ways to prevent the disease altogether, the exposure of the retina to bright ambient light following premature birth has been a natural hypothesis, since the premature infant normally would be in the dark in-utero environment. Several controlled studies have now addressed this theory. OBJECTIVES: To answer the question: "Among very low birth weight infants, what is the effect of reducing early environmental light exposure on the incidence of any "Acute ROP", or "Poor ROP Outcomes"? SEARCH STRATEGY: Searches were made of the Cochrane Neonatal Group Register of Controlled Trials, Medline, EMBASE, the Cochrane Library, previous reviews including cross references, abstracts, conference and symposia proceedings, and expert informants. The search terms used were [retrolental fibroplasia or retinopathy of prematurity] and [light or light/ae or lighting or lighting/ae or light/tu or lighting/st]. This search was updated as of November 2000. SELECTION CRITERIA: Randomized or quasi-randomized controlled trials that reduced light exposure to premature infants within the first 7 days following birth were considered for this review. DATA COLLECTION AND ANALYSIS: Data on clinical outcomes including any Acute ROP and Poor ROP Outcome were excerpted by both reviewers independently and consensus reached. Data analysis was conducted according to the standards of the Neonatal Cochrane Review Group. MAIN RESULTS: Data from four recent randomized trials, and one much older quasi-randomized trial failed to show any reduction in Acute ROP, or Poor ROP Outcome with the reduction of ambient light to premature infants' retinas. The number of infants studied to date allows 95% confidence that IF there were a true difference being missed, it would be smaller than 7 percentage points on a background of 54% of all infants under 2 kg developing ROP. REVIEWER'S CONCLUSIONS: Decreasing retinal ambient light exposure in premature infants is very unlikely to reduce the incidence of ROP. PMID- 11279679 TI - Nimodipine for primary degenerative, mixed and vascular dementia. AB - BACKGROUND: Dementia is an age-related condition in which Alzheimer's disease (AD) and cerebrovascular disease account for the bulk of cases. The role played by calcium in regulating brain functions is well known - the calcium ion links membrane excitation to subsequent intracellular enzymatic response. Change in calcium homeostasis is one important effect of aging with repercussions on higher cortical functions. Nimodipine is an isopropyl calcium channel blocker which can easily cross the blood brain barrier. Its primary action is to reduce the number of open channels, thus restricting influx of calcium ions into the cell. The usefulness of nimodipine in patients with Alzheimer's disease and vascular dementia and unspecified dementia is still controversial with mixed results. In spite of the uncertainties about its efficacy in dementia, nimodipine is currently a frequently prescribed drug for cognitive impairment and dementia in several European countries. This review will be conducted in two phases; the current review is based on evidence from published data only. The second phase will be based on individual-patient data analysed centrally and added to this review in due course. OBJECTIVES: To determine the clinical efficacy of nimodipine for the symptoms of dementia, either unclassified or according to the major subtypes - Alzheimer's disease, vascular, or mixed Alzheimer's and vascular dementia. SEARCH STRATEGY: The Cochrane Dementia Group Register of Clinical Trials was searched using the terms 'nimodipine' and 'isopropyl (2-methoxy-ethyl) 1,4-dihydro-2, 6-dimethyl-4-(3-nitrophenyl)-3, 5-pyridinedicarboxylate'. SELECTION CRITERIA: All unconfounded, double-blind, randomised trials in which treatment with nimodipine was administered for more than a day and compared to placebo in patients with dementia, either unclassified or according to the major subtypes - Alzheimer's disease, vascular, or mixed Alzheimer's and vascular dementia. DATA COLLECTION AND ANALYSIS: Data were extracted independently by the reviewers and the odds ratio (95%CI) or the average difference (95%CI) were estimated. Both intention-to-treat and on-treatment results were extracted. MAIN RESULTS: This review produced no clear results. Many of the data published were not capable of being sensibly pooled. The data were compatible with nimodipine producing improvement, no change or even harm for those with Alzheimer's disease, vascular dementia, or mixed Alzheimer's and vascular dementia. It was not possible to use many of the published results in a combined analysis. For measures of overall clinical improvement, the intention-to-treat analysis, based on one study only, failed to detect any difference between nimodipine and placebo (OR 0.53; 95%CI 0.25 - 1.13). An on-treatment analysis, based on one study only, produced a statistically significant difference in favour of nimodipine (SMD 4.4; 95%CI 3.9 - 5.0). For cognitive function, the effect of nimodipine was statistically significantly different from placebo for the Mini Mental State Examination score (0-30; high =good) (SMD 0.9; 95%CI 0.59 - 1.22) and there was a statistically significant effect in favour of treatment for the Wechsler Memory Scale (SMD 0.47; 95%CI 0.17 - 0.77). These analyses were based only on those who completed the study and not intention-to-treat analyses. There were no results presented in a form suitable for pooling for functional autonomy, behaviour, quality of life dependency (eg institutionalization), effect on carer, death, acceptability of treatment (as measured by withdrawal rate, safety (as measured by the incidence of adverse effects, including side effects, leading to withdrawal). REVIEWER'S CONCLUSIONS: This review provides no convincing evidence that nimodipine is a useful treatment for the symptoms of dementia, either unclassified or according to the major subtypes - Alzheimer's disease, vascular, or mixed Alzheimer's and vascular dementia. However, as so few of the trials presented data in a format suitable for pooling, the results of this review may be modified when further data from all relevant trials are included. There is an urgent need for the independent evaluation of the data already existing in the trials but not accessible through published or grouped data. An independent meta analysis of the individual patient data is required. Nimodipine cannot be currently recommended in patients with dementia. The results and conclusions of this update are unaltered by further searching as the additional studies do not add any further valid/eligible data. PMID- 11279680 TI - Planned Caesarean section for term breech delivery. AB - BACKGROUND: Routine use of caesarean section for breech presentation is widespread. However, poor outcomes after breech birth might be the result of underlying conditions causing breech presentation rather than damage during delivery. OBJECTIVES: The objective of this review was to assess the effects of planned caesarean section for breech presentation on measures of pregnancy outcome. SEARCH STRATEGY: We searched the Cochrane Pregnancy and Childbirth trials register and the Cochrane Controlled Trials register. Date of last search: November 2000. SELECTION CRITERIA: Randomised trials comparing planned caesarean section for breech presentation with planned vaginal delivery. DATA COLLECTION AND ANALYSIS: Reviewers assessed trial eligibility and quality. Data were entered onto and analysed using RevMan software. MAIN RESULTS: Caesarean delivery occurred in 550/1227 (45%) of those women allocated to a vaginal delivery protocol. Planned caesarean section was associated with modestly increased maternal morbidity (relative risk 1.29, 95% confidence interval 1.03 to 1.61). Perinatal and neonatal death (excluding fatal anomalies) were greatly reduced (0.29, 0.10 to 0.86). The reductions were similar for countries with low and high perinatal mortality rates. Perinatal/neonatal death or neonatal morbidity was also greatly reduced (0.31, 0.19 to 0.52). The difference was smaller for countries with a high national perinatal mortality rate. REVIEWER'S CONCLUSIONS: Planned caesarean section greatly reduces both perinatal/neonatal mortality and neonatal morbidity, at the expense of somewhat increased maternal morbidity. Cost, and future morbidity due to the caesarean section scar were not assessed. The option of external cephalic version is dealt with in separate reviews. The data from this review will help to inform individualised decision-making regarding breech delivery. PMID- 11279681 TI - Pre-operative traction for fractures of the proximal femur. AB - BACKGROUND: Pre-operative traction following an acute hip fracture remains standard practice in some hospitals. OBJECTIVES: To evaluate the effects of traction applied to the injured limb prior to surgery for a fractured hip. Different methods of applying traction (skin or skeletal) were considered. SEARCH STRATEGY: We searched the Cochrane Musculoskeletal Injuries Group trials register up to September 1999, MEDLINE (1966 to October 2000), CINAHL (1982 to August 2000), EMBASE (1980 to August 2000), CENTRAL (Issue 4, 2000 of The Cochrane Library), the National Research Register (Issue 3, 2000) and bibliographies of trial reports. Date of the most recent search: October 2000. SELECTION CRITERIA: All randomised or quasi-randomised trials comparing either skin or skeletal traction with no traction, or skin with skeletal traction for patients with an acute hip fracture prior to surgery. DATA COLLECTION AND ANALYSIS: Both reviewers independently assessed trial quality, using a nine item scale, and extracted data. Additional information was sought from all trialists. Wherever appropriate and possible, the data are presented graphically. MAIN RESULTS: Six randomised trials, mainly of moderate quality, involving a total of 938 predominantly elderly patients, were identified and included in the review. This review update includes a newly identified trial. The inclusion of this trial resulted in no important change in the results or conclusions. Five trials compared traction with no traction. The new study found a statistically significant reduction in rest pain in the traction group but did not indicate if this was clinically significant; there was no difference in analgesic use. The other four trials found no evidence of benefit from traction, either in the relief of pain, ease of fracture reduction or quality of fracture reduction at time of surgery. One of these trials included both skin and skeletal traction groups. This trial and one other which compared skeletal traction with skin traction found no important differences between these two methods, although the initial application of skeletal traction was noted as being more painful and most costly. REVIEWER'S CONCLUSIONS: From the evidence available, the routine use of traction (either skin or skeletal) prior to surgery for a hip fracture does not appear to have any benefit. Where a policy of general or selective application of traction exists, the choice of method must remain a decision based on evaluation of the individual patient. Further, high quality trials would be required to confirm or refute the absence of benefits of traction. PMID- 11279682 TI - Corticosteroids for preventing relapse following acute exacerbations of asthma. AB - BACKGROUND: Acute asthma is responsible for many emergency department visits annually. Between 12-16% will relapse to require additional interventions within two weeks of ED discharge. Treatment of acute asthma is based on rapid reversal of bronchospasm and reducing airway inflammation and this review examines the evidence for using systemic corticosteroids to improve outcomes after discharge from the ED. OBJECTIVES: To determine the benefit of corticosteroids (oral, intramuscular, or intravenous) for the treatment of asthmatic patients discharged from an acute care setting (i.e. usually the emergency department) after assessment and treatment of an acute asthmatic exacerbation. SEARCH STRATEGY: The Cochrane Airways Group "Asthma and Wheez* RCT" register was searched using the terms: a) Asthma OR Wheez* b) Glucocorticoid OR Steroid* AND c) Exacerbat* OR Relapse* OR Emerg*. In addition, authors of all included studies were contacted to determine if unpublished studies which met the inclusion criteria were available. Bibliographies from included studies, known reviews and texts were also searched for additional citations. SELECTION CRITERIA: Only randomized controlled trials were eligible for selection. Studies were included in this review if they dealt with the outpatient treatment of asthmatic exacerbations using glucocorticoids at discharge and reported either relapse rate or PFTs. Two independent reviewers first identified potentially relevant studies and then selected articles for inclusion. Methodological quality was assessed independently by two reviewers. Agreement was assessed using kappa (k) statistics. DATA COLLECTION AND ANALYSIS: Data were extracted independently by two reviewers; authors were contacted to verify the extracted data and clarify missing information. When author contact was unsuccessful, missing data were estimated from graphs where possible. Sensitivity, sub-group and overall analyses were performed using the Cochrane Review Manager. MAIN RESULTS: A search that yielded 229 references identified 169 (73%) original publications. Reviewers identified 8 studies for potential inclusion (k =0.76); 18 references were added by searching publication reference lists and contact with authors. Of these 26 articles, a total of 7 were included in the overview. Two studies used intramuscular corticosteroids, five studies used oral corticosteroids. Significantly fewer patients in the corticosteroid group relapsed to receive additional care in the first week (odds ratio (OR) 0.35; 95% confidence interval (CI): 0.17, 0.73). This favourable effect was maintained over the first 21 days (OR 0.33; 95% CI: 0.13, 0.82). Patients receiving corticosteroids had less need for beta-agonists (weighted mean difference (WMD) -3.3 activations/day; 95% CI: 5.5, -1.0). Changes in pulmonary function tests (SMD 0.045; 95% CI: -0.47, 0.56) and side effects (SMD 0.03; 95% CI : -0.38, 0.44) in the first 7-10 days, while rarely reported, showed no differences between the treatment groups. Statistically significant heterogeneity was identified for the side effect results; all other outcomes were homogeneous. It appears that IM corticosteroids are similarly efficacious to a 7-10 day tapering course of oral agents. From these results, as few as 13 patients need to be treated to prevent relapse to additional care after an exacerbation of asthma. REVIEWER'S CONCLUSIONS: A short course of corticosteroids following assessment for an acute exacerbation of asthma significantly reduces the number of relapses to additional care and decreases beta-agonist use without an apparent increase in side effects. Intramuscular corticosteroids appear as effective as oral agents. PMID- 11279683 TI - Calcium channel blockers for neuroleptic-induced tardive dyskinesia. AB - BACKGROUND: Tardive dyskinesia (TD) is a potentially disfiguring movement disorder of the orofacial region often caused by use of neuroleptic drugs. A wide range of strategies has been used to help manage TD and, for those who are unable to have their antipsychotic medication stopped or substantially changed, the calcium-channel blocking group of drugs (diltiazem, nifedipine, nimodipine, verapamil) has been suggested as a useful adjunctive treatment. OBJECTIVES: To determine the effects of calcium-channel blocker drugs (diltiazem, nifedipine, nimodipine, verapamil) for treatment of neuroleptic-induced TD in people with schizophrenia, schizoaffective disorder or other chronic mental illnesses. SEARCH STRATEGY: Electronic searches of Biological Abstracts (1982-2000), Cochrane Library (Issue 4, 2000), Cochrane Schizophrenia Group's Register of trials (November 2000), EMBASE (1980-2000), LILACS (1982-2000), MEDLINE (1966-2000), PsycLIT (1974-2000), and SCISEARCH were undertaken. References of all identified studies were searched for relevant citations. Principal authors of trials were contacted. SELECTION CRITERIA: Randomised clinical trials comparing calcium channel blockers to placebo or no intervention for people with both TD and schizophrenia or serious mental illness were reliably selected. DATA COLLECTION AND ANALYSIS: Data were to have been independently extracted and analysed on an intention-to-treat basis. The relative risk (RR) and 95% confidence intervals (CI) of homogeneous dichotomous data were to have been calculated using a random effects model, and, where possible, the number needed to treat calculated. Weighted mean differences (WMD) were to have been calculated for continuous data. MAIN RESULTS: No trials were included. Seven studies were excluded; five were not randomised and two small randomised crossover studies provided no usable data. Two more small randomised controlled trials await assessment. The authors have been contacted for relevant information. REVIEWER'S CONCLUSIONS: Based on currently available information, no confident statement can be made about the effectiveness of calcium-channel blockers for treating people with neuroleptic induced tardive dyskinesia. Before evaluation of these drugs in larger randomised controlled trials, clinicians should carefully weigh up the possible benefits against their potential adverse effects. PMID- 11279684 TI - Repeated lumbar or ventricular punctures in newborns with intraventricular hemorrhage. AB - BACKGROUND: Although it has been possible to reduce the percentage of premature infants suffering intraventricular hemorrhage, posthemorrhagic hydrocephalus remains a serious problem without a good treatment. There is a high rate of cerebral palsy, and ventriculoperitoneal shunt surgery makes the child permanently dependent on the valve and catheter system. Shunt surgery cannot be carried out early because of the blood in the cerebrospinal fluid (CSF) and the brain may be subjected to periods of raised pressure. Early tapping of CSF by lumbar puncture or ventricular tap was suggested as a way of temporarily reducing pressure and removing blood and protein and thereby avoiding permanent hydrocephalus. OBJECTIVES: To determine whether repeated CSF tapping, by lumbar puncture or ventricular tap, reduced the risk of permanent shunt dependence, neurodevelopmental disability or death in neonates at risk of, or actually developing, post-hemorrhagic hydrocephalus (PHH). This form of treatment was based on the hypothesis that repeated tapping removed protein and blood from the CSF, thus clearing obstruction from the channels of CSF absorption. SEARCH STRATEGY: Pediatric, Neurosurgical and General Medical Journals were handsearched from 1976 up to October 2000, as well as the Medline database (via PubMed) and the Cochrane Controlled Trials Register. Personal contacts were used. SELECTION CRITERIA: Four controlled trials ( with five published papers) were identified, three being randomised and the fourth using alternative allocation. Two trials evaluated repeated lumbar punctures in neonates with intraventricular hemorrhage (IVH) and two trials evaluated repeated CSF tapping infants with IVH followed by progressive ventricular dilatation. DATA COLLECTION AND ANALYSIS: In addition to details of the patient selection and patient allocation, the interventions were extracted. The end-points examined were: ventriculoperitoneal shunt, death, disability, multiple disability and death or disability. MAIN RESULTS: The studies were sufficiently similar in the question they were asking and the interventions were sufficiently in common that they could be combined when assessing the effect of the intervention. When repeated CSF tapping was compared to conservative treatment, the relative risks for shunt placement, death, disability and multiple disability were very close to 1.0 with no statistically significant effect. There is also evidence that this form of treatment increased the risk of CSF infection. REVIEWER'S CONCLUSIONS: Early repeated CSF tapping cannot be recommended for neonates at risk of, or actually developing, post hemorrhagic hydrocephalus. PMID- 11279685 TI - Vitamin D and vitamin D analogues for preventing fractures associated with involutional and post-menopausal osteoporosis. AB - BACKGROUND: Due to their known effects on bone metabolism, vitamin D and related compounds have been proposed for the prevention of osteoporosis and fractures. OBJECTIVES: To determine the effects of supplementation with Vitamin D or a Vitamin D analogue in the prevention of fractures of the axial and appendicular skeleton in elderly men or women with involutional or post-menopausal osteoporosis. SEARCH STRATEGY: We searched MEDLINE, EMBASE, CINAHL, LILACS, CABNAR, BIOSIS, HEALTHSTAR, Current Contents, The Cochrane Database of Systematic Reviews, the Cochrane Musculoskeletal Injuries Group trials register, and bibliographies of identified trials and reviews. Date of the most recent search: September 2000. SELECTION CRITERIA: Any randomised or quasi-randomised trial which compared vitamin D or a vitamin D analogue, either alone or in combination with calcium supplementation, with a placebo, no intervention, or the administration of calcium supplements, with eligible fracture outcomes, in elderly men or women with involutional or post-menopausal osteoporosis. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality, by use of a nine item scale, and extracted data. Additional information was sought from trialists. Where possible the data were pooled. Pooling of data, where it was admissible, used pooled relative risk and fixed effects model. MAIN RESULTS: Almost all estimates of treatment effects are based on single studies. Administration of vitamin D3 alone without calcium co-supplementation was not associated with any reduction in incidence of hip fracture (relative risk (RR) 1.20, 95% confidence interval (CI) 0.83, 1.75) or other non-vertebral fracture. Administration of vitamin D3 with calcium co-supplementation to frail elderly people in sheltered accommodation was associated with a reduction in incidence of hip fracture (RR 0.74, 95% CI 0.60, 0.91). In healthy younger, ambulant participants the effect on hip fracture is unknown (RR 0.36, 95% CI 0.01, 8.78), although there appears to be a significant overall effect on non-vertebral fracture incidence in this group ( RR 0.46, 95% CI 0.23,0.90). Calcitriol (1,25 dihdyroxy vitamin D) was effective in reducing the incidence of vertebral deformity (RR 0.49, 95% CI 0.25, 0.95). Calcitriol was more effective than calcium in reducing the frequency of new vertebral deformities during the third year of treatment (RR 0.28, 95% CI 0.15, 0.52). 1-alpha-hydroxy vitamin D was effective in reducing the incidence of non-vertebral fractures in a single small study of elderly people whose mobility was impaired by neurological disease (RR 0.12, 95% CI 0.02, 0.95). No statistically significant effects were found for other comparisons of vitamin D or its analogues against each other, with and without calcium supplementation. REVIEWER'S CONCLUSIONS: Uncertainty remains about the efficacy of regimens which include vitamin D or its analogues in fracture prevention. Particularly if co-supplementation of calcium is required, significant cost differences are likely to exist between regimens. Further large randomised trials are currently being conducted to clarify the effectiveness of community fracture prevention programmes employing vitamin D supplementation. PMID- 11279686 TI - Vitamin K prior to preterm birth for preventing neonatal periventricular haemorrhage. AB - BACKGROUND: Preterm infants are at risk of periventricular haemorrhage. This can be a sign of brain damage that might lead to neurodevelopmental abnormalities, including cerebral palsy. It has been suggested that vitamin K might improve coagulation in preterm infants. OBJECTIVES: The objective of this review was to assess the effects of vitamin K administered to women at risk of imminent very preterm birth to prevent periventricular haemorrhage and associated neurological injury in the infant. SEARCH STRATEGY: We searched the Cochrane Pregnancy and Childbirth Group trials register, Cochrane Controlled Trials Register, and bibliographies up to September 2000. SELECTION CRITERIA: Randomised or quasi randomised trials of vitamin K administered parenterally or orally to women at risk of imminent preterm birth. The primary outcomes were neonatal mortality, neonatal neurological morbidity, as measured by the presence of periventricular haemorrhage (PVH) on ultrasound during the first week of life, and long term neurodevelopment. Secondary outcomes included other neonatal morbidity and any maternal side effects. DATA COLLECTION AND ANALYSIS: Eligibility, trial quality assessment and data extraction were done independently by two reviewers. MAIN RESULTS: Five trials were included, involving more than 420 women. The trials were of variable quality. Antenatal vitamin K was associated with a non significant trend to a reduction in all grades of periventricular haemorrhage (relative risk (RR) 0.82, 95% confidence interval (CI) 0.67-1.00) and in severe PVH (grades 3 and 4) (RR 0.75, 95% CI 0.45-1.25) for babies receiving prenatal vitamin K compared with control babies. This trend disappeared when poorer quality trials were excluded. Information on neurodevelopment was only given for a small sample of children in one trial with discrepancy in results given in the two reports. REVIEWER'S CONCLUSIONS: Vitamin K administered to women prior to very preterm birth has not been shown to significantly prevent periventricular haemorrhages in preterm infants. PMID- 11279687 TI - Antibiotic prophylaxis for surgery for proximal femoral and other closed long bone fractures. AB - BACKGROUND: Wound infection and other hospital-acquired infections cause significant morbidity after internal fixation of fractures (osteosynthesis). The administration of antimicrobial agents (antibiotics) may reduce the frequency of infections. OBJECTIVES: To determine whether the prophylactic administration of antibiotics in patients undergoing surgical management of hip or other long bone fractures reduces the incidence of wound and other hospital acquired infections. SEARCH STRATEGY: We searched the The Cochrane Library, Issue 3 2000; MEDLINE, EMBASE, LILACS, Current Contents, Dissertation Abstracts, and Index to UK Theses to August 2000. Bibliographies of identified articles were screened for further relevant trials. No language restriction was applied. SELECTION CRITERIA: Randomised or quasi-randomised controlled trials involving - Participants: Any patients with a hip or other closed long bone fracture undergoing surgery for internal fixation or replacement arthroplasty. INTERVENTIONS: Any regimen of systemic antibiotic prophylaxis administered at the time of surgery. OUTCOME MEASURES: Wound infection (deep and superficial), urinary tract infection, respiratory tract infection, adverse effects of prophylaxis, economic evaluations. DATA COLLECTION AND ANALYSIS: Two reviewers independently screened papers for inclusion, assessed trial quality using an eight item scale, and extracted data. Additional information was sought from two trialists. Pooled data are presented graphically. MAIN RESULTS: Data from 8307 participants in 22 studies were analysed. In patients undergoing surgery for closed fracture fixation, single dose antibiotic prophylaxis significantly reduced deep wound infection (relative risk 0.40, 95%CI 0.24, 0.67) superficial wound infections, urinary infections, and respiratory tract infections. Multiple dose prophylaxis had an effect of similar size on deep wound infection (relative risk 0.40, 95%CI 0.24, 0.67), but significant effects on urinary and respiratory infections were not confirmed. Economic modelling using data from one large trial indicates that single dose prophylaxis with ceftriaxone is a cost-effective intervention. There are limited data for the incidence of adverse effects, but as expected they appear to be more common in those receiving antibiotics, compared with placebo or no prophylaxis. REVIEWER'S CONCLUSIONS: Antibiotic prophylaxis should be offered to those undergoing surgery for closed fracture fixation. On ethical grounds, further placebo controlled randomised trials of the effectiveness of antibiotic prophylaxis in closed fracture surgery are unlikely to be justified. Trials addressing the cost-effectiveness of different effective antibiotic regimens would need to be very large. PMID- 11279688 TI - Interventions to implement prevention in primary care. AB - BACKGROUND: Primary care physicians hold a strategic position in delivering preventive services. However discrepancies exist between evidence based guidelines and practice. OBJECTIVES: To assess the effects of interventions to improve the delivery of preventive services in primary care. SEARCH STRATEGY: We searched the Cochrane Effective Practice and Organisation of Care Group specialised register (November 1995; August 1999), MEDLINE (1980 to 1995) and hand searched relevant journals. SELECTION CRITERIA: Randomised trials, controlled before and after studies, and interrupted time series analyses of interventions to improve preventive services by primary care professionals responsible for patient care. DATA COLLECTION AND ANALYSIS: Two researchers independently extracted data and assessed study quality. MAIN RESULTS: Fifty-five studies were included, involving more than 2000 health professionals and 99,000 people, with 83 comparisons between intervention and control groups. Post intervention differences between intervention and control groups varied widely within and across categories of interventions. Most interventions were found to be effective in some studies, but not in others. Five comparisons of group education versus no intervention showed absolute change of preventive services varying between -4% and +31%. Nine comparisons of physician reminders versus no intervention showed absolute change of preventive services varying between 5% and 24%. Fourteen comparisons of multifaceted interventions versus no intervention showed absolute change of preventive services varying between -3% and +64%. Six comparisons of multifaceted interventions versus group education reported absolute changes varying between -31% and +28%. All these comparisons used randomised groups. Ten comparisons of multifaceted interventions versus no intervention used non-randomised groups and showed absolute change of preventive services varying between -5% and +21%. The remaining planned comparisons within categories of interventions contained less than five comparisons. REVIEWER'S CONCLUSIONS: There is currently no solid basis for assuming that a particular intervention or package of interventions will work. Effective interventions to increase preventive activities in primary care exist, but there is considerable variation in the level of change achieved, with effect sizes usually small or moderate. Tailoring interventions to address specific barriers to change in a particular setting is probably important. Multifaceted interventions may be more effective than single interventions, because more barriers to change can be addressed. Future research should analyse barriers to change and interventions to implement preventive services in more detail, to clarify how interventions relate to specific barriers. Since more complex interventions are likely to be more effective but also more costly, economic evaluations should also be included. PMID- 11279689 TI - Multidisciplinary team interventions for delirium in patients with chronic cognitive impairment. AB - BACKGROUND: Delirium is common in hospitalized elderly people. In the frail elderly, delirium may occur in 60% of those hospitalized. In the cognitively impaired, 45% have been shown to develop delirium and these patients have longer lengths of hospital stay and a higher rate of complications which, with other factors, together contribute to an increase in cost of care. The combination of being elderly and chronically cognitively impaired leads to a high risk of delirium with the associated increased risk of prolonged hospital stay, complications, and poor outcomes. The management of delirium has commonly been multifaceted - the primary emphasis has always been on the diagnosis and therapy of the precipitating factors, but as these may not be immediately resolved, symptomatic and supportive care may become of major importance. OBJECTIVES: The objective of this review is to assess the available evidence for the effectiveness, if any, of multidisciplinary team interventions in the coordinated care of patients with delirium superimposed on an underlying chronic cognitive impairment compared with the usual care of older cognitively impaired patients. SEARCH STRATEGY: A search of all available databases and sources of references was carried out in July 2000; this comprised the CDCIG specialised register in addition to the Reviewer's files and bibliographic sources. SELECTION CRITERIA: Selection for possible inclusion in this review is made on the basis of the research methodology - controlled trials whose participants are reported as having chronic cognitive impairment, and who then developed incident delirium and were randomly assigned to either coordinated multidisciplinary care or usual care. DATA COLLECTION AND ANALYSIS: Nine (9) controlled trials were identified for possible inclusion in the review, none of which meets the inclusion criteria, for reasons which are recorded in the table of excluded studies, and no data, therefore, were extracted for cross study analysis or synthesis. MAIN RESULTS: No studies focused on patients with prior cognitive impairment, so management of delirium in this group could not be assessed. There is very little information on the management of delirium in the literature but there is an increasing body of information about the incidence, risks and prognosis of the disorder in the elderly population. REVIEWER'S CONCLUSIONS: The management of delirium needs to be studied in a more clearly defined way before evidence-based guidelines can be developed. Insufficient data are available for the development of evidence based guidelines on diagnosis or management. There is scope for research in all areas - from basic pathophysiology and epidemiology to prevention and management. Though much recent research has focused on the problem of delirium, the evidence is still difficult to utilize in management programmes. Research needs to be undertaken targeting specific groups known to be at high risk of developing delirium, for example the cognitively impaired and the frail elderly. As has been highlighted by Inouye 1999, delirium has very important economic and health policy implications and is a clinical problem which affects all aspects of care of the elderly. Delirium, though a frequent problem in the hospitalised elderly patient, is still managed empirically and there is no evidence in the literature to support change to current practice at this time. PMID- 11279690 TI - Enteral antibiotics for preventing necrotizing enterocolitis in low birthweight or preterm infants. AB - BACKGROUND: Necrotizing enterocolitis continues to be a problem, particularly in preterm neonates. There have been reports published suggesting that the use of enteral antibiotics may be effective as prophylaxis. This systematic review was undertaken to clarify the issue. OBJECTIVES: To evaluate the benefits and harms of enteral antibiotic prophylaxis for necrotizing enterocolitis in low birth weight and preterm infants. SEARCH STRATEGY: Searches were made of the Oxford Database of Perinatal trials, MEDLINE (1966 - June 2000; search terms: necrotizing enterocolitis, antibiotics; limits: newborn infant), previous reviews with cross references, abstracts, conference and symposia proceedings, expert informants and journal hand searching in the fields of neonatal pediatrics and microbiology. SELECTION CRITERIA: All randomized or quasi-randomized controlled trials where enteral antibiotics were used as prophylaxis against NEC in LBW (<2500g) and/or preterm (<37 weeks gestation) infants. DATA COLLECTION AND ANALYSIS: The standard method of the Cochrane Collaboration and its Neonatal Review Group was used. The methodological quality of each trial was reviewed by the second author who was blinded to the trial authors and institutions. Each author extracted data separately before comparison and resolution of differences. Relative risk (RR), risk difference (RD), and number needed to treat were used in the analysis. MAIN RESULTS: Five eligible trials involving 456 infants were included. The administration of prophylactic enteral antibiotics resulted in a statistically significant reduction in NEC [RR 0.47 (0.28, 0.78); RD -0.10 ( 0.16, -0.04); NNT 10 (6, 25)]. There was a statistically significant reduction in NEC-related deaths [RR 0.32 (0.10, 0.96); RD -0.07 (-0.13, 0.01); NNT 14 (8, 100)]. There was a trend towards a reduction in all deaths which was not significant [RR 0.67(0.34, 1.32)]. There were no significant differences in NEC like enteropathies (one trial only). One study found a statistically significant increase in the incidence of colonization with resistant bacteria and the summary analysis of three trials gave an increase which was just significant [RR 1.73 (1.00, 2.97); RD 0.07 (0.00, 0.13)]. REVIEWER'S CONCLUSIONS: Evidence suggests that oral antibiotics reduce the incidence of NEC in low birth weight infants. However concerns about adverse outcomes persist, particularly related to the development of resistant bacteria. To address this question further, a large trial would be required with a sample size sufficient to examine all the important benefits and harms. Adverse outcomes associated with infection should be evaluated, and microbiological studies looking for the development of resistant bacteria should be undertaken PMID- 11279691 TI - Interventions for treating proximal humeral fractures in adults. AB - BACKGROUND: Proximal humeral fractures are common yet management varies widely. In particular, the role and timing of any surgical intervention have not been clearly defined. OBJECTIVES: To collate and evaluate the scientific evidence supporting the various methods used for treating proximal humeral fractures. SEARCH STRATEGY: We searched the Cochrane Musculoskeletal Injuries Group trials register, MEDLINE, PubMed, the Cochrane Controlled Trials Register, CINAHL, the National Research Register and bibliographies of trial reports. The search was completed in July 2000. SELECTION CRITERIA: All randomised studies pertinent to the treatment of proximal humeral fractures were selected. DATA COLLECTION AND ANALYSIS: Independent quality assessment and data extraction were performed by two reviewers. Although quantitative data from trials are presented, trial heterogeneity prevented pooling of results. MAIN RESULTS: Nine randomised trials were included. All were small trials; the largest study involved only 85 patients. Bias in these trials could not be ruled out. Six trials evaluated conservative treatment, two compared surgery with conservative treatment and one compared two surgical techniques. In the 'conservative' group there was very limited evidence indicating that the type of bandage used made any difference in terms of time to fracture union and the functional end result. However, an arm sling was generally more comfortable than a body bandage. There was some evidence that mobilisation at one week instead of three weeks alleviated pain in the short term without compromising long term outcome. Two trials provided some evidence that patients, when given sufficient instruction to pursue an adequate physiotherapy programme, could generally achieve a satisfactory outcome if allowed to exercise without supervision. Operative reduction improved fracture alignment in two trials. However, in one trial, surgery was associated with a greater risk of complication, and did not result in improved shoulder function. Fracture fixation of severe injuries was associated with a high rate of re operation in one trial, comparing tension-band wiring fixation with hemi arthroplasty. REVIEWER'S CONCLUSIONS: Only tentative conclusions can be drawn from the available randomised trials, which do not provide robust evidence for many of the decisions which need to be made in contemporary fracture management. It is unclear whether operative intervention, even for specific fracture types, will produce consistently better long term outcomes. There is a need for good quality evidence for the management of these fractures. PMID- 11279692 TI - Synchronized mechanical ventilation for respiratory support in newborn infants. AB - BACKGROUND: During synchronized mechanical ventilation, positive airway pressure and spontaneous inspiration coincide. Thus, if synchronous ventilation is provoked, it is likely that adequate gas exchange should be achieved at lower peak airway pressures, reducing barotrauma and hence airleak and chronic lung disease. Synchronous ventilation can be achieved by manipulation of rate and inspiratory time during conventional ventilation and employment of patient assisted ventilation. OBJECTIVES: To compare (i) the efficacy of synchronized mechanical ventilation, delivered as high frequency positive pressure ventilation or triggered ventilation (patient triggered ventilation (PTV) or synchronous intermittent mandatory ventilation (SIMV)) with conventional ventilation (ii) different types of triggered ventilation SEARCH STRATEGY: Searches were made from 1985-2000 of the Oxford Database of Perinatal Trials, Medline (MeSH terms: mechanical ventilation; triggered ventilation; newborn infant); previous reviews, abstracts, symposia proceedings, hand searching of journals in the English language and contacting expert informants. SELECTION CRITERIA: Randomized or quasi randomized clinical trials comparing synchronized ventilation delivered as high frequency positive pressure ventilation (HFPPV) or triggered ventilation (PTV/SIMV) to conventional mechanical ventilation (CMV) in neonates. Randomized trials comparing different triggered ventilation modes (PTV and SIMV) in neonates. DATA COLLECTION AND ANALYSIS: Data regarding clinical outcomes including mortality, airleaks (pneumothorax or pulmonary interstitial emphysema (PIE)), severe intracerebral haemorrhage (grades 3 and 4), chronic lung disease (oxygen dependency beyond 28 days) and duration of weaning/ventilation. Data subdivided into three groups: (i) HFPPV vs CMV; (ii) PTV/SIMV vs CMV; (iii) PTV vs SIMV. Data analysis was conducted according to the standards of the Neonatal Cochrane Review Group. MAIN RESULTS: The meta-analysis demonstrates that HFPPV compared to CMV was associated with a reduction in the risk of airleak (typical relative risk for pneumothorax was 0.69 (95% CI 0.51, 0.93). PTV/SIMV compared to CMV was associated with a shorter duration of ventilation (weighted mean difference -31.8 hours, 95% CI -54.1, -9.6). PTV compared to SIMV was associated with a trend to a shorter duration of weaning (weighted mean difference -42.4 hours, 95% CI -94.4, 9.6). No disadvantage to HFPPV or triggered ventilation was noted regarding other outcomes but neither ventilatory mode was associated with a significant reduction in the incidence of chronic lung disease. REVIEWER'S CONCLUSIONS: Compared to conventional ventilation, benefit is demonstrated for both HFPPV and triggered ventilation with regard to a reduction in airleak and a shorter duration of ventilation respectively. In none of the trials was complex respiratory monitoring undertaken and thus it is not possible to conclude that the mechanism of producing those benefits is by provocation of synchronized ventilation. Further trials are needed to determine whether synchronized ventilation is associated with other benefits but optimization of trigger and ventilator design with respect to respiratory diagnosis is encouraged before embarking on further trials. PMID- 11279693 TI - Surgery or embolisation for varicocele in subfertile men. AB - BACKGROUND: A varicocele is an, almost exclusively left-sided, varicosity of the pampiniform plexus of the spermatic cord, forming a tangle of distended blood vessels in the scrotum. Although the concept that varicocele causes male subfertility and therefore varicocelectomy cures male subfertility has been around for almost fifty years now, the mechanisms by which varicocele would affect fertility have not yet been satisfactorily explained, and neither have the mechanisms by which varicocelectomy would resolve subfertility. Furthermore, it has been questioned whether a causal relation exists at all between the distension of the pampiniform plexus and impairment of fertility. OBJECTIVES: To evaluate the effect of varicocele treatment on pregnancy rate in subfertile couples. SEARCH STRATEGY: Relevant trials were identified in the Cochrane Menstrual Disorders and Subfertility Group's specialised register of controlled trials. A MEDLINE search, using the group's search strategy, was performed for the period 1966-2000. Also, hand searching was performed of 22 specialist journals in the field from their first issue till 2000. Cross references and references from review articles were checked. SELECTION CRITERIA: RCTs were included if they were relevant to the clinical question posed, if they reported pregnancy rates as an outcome measure, and if they reported data in treated (surgical ligation or radiological embolization of the internal spermatic vein) and untreated groups. DATA COLLECTION AND ANALYSIS: Six studies met the inclusion criteria for this review. One (Nieschlag 1995/1998) was an extension of a previously published study (Nieschlag 1995/1998), which left five studies for analysis (Nilsson 1979; Breznik 1993; Madgar 1995; Yamamoto 1996; Nieschlag 1995/1998). The results of a WHO megatrial are awaited but as yet are unavailable. The WHO data will be added if and when they will have become available. All five only included men from couples with subfertility problems, one (Madgar 1995) excluded men with sperm counts <5 mill/mL, three (Nilsson 1979; Breznik 1993; Yamamoto 1996) also included men with normal semen analysis. One study (Yamamoto 1996) specifically addressed only men with subclinical varicoceles as diagnosed by thermography. Potentially relevant trials were screened independently by two authors (JE and JC). Any differences of opinion were resolved by consensus meeting (none occurred for this review). Studies were excluded from meta-analysis if they made comparisons other than those specified above. MAIN RESULTS: One trial (Madgar 1995) reported a statistically significant improvement in pregnancy rate following high ligation of the left spermatic vein. None of the other four studies showed individually a significant effect on pregnancy rates of varicocele treatment over no-treatment (Nilsson 1979; Breznik 1993; Yamamoto 1996), or over counseling only (Nieschlag 1995/1998). The combined RR (Relative Risk; random effects method) of the five studies is 1.06 (95%CI 0.57 1.94), the Peto OR (Odds Ratio) is 1.15 (95%CI 0.73-1.83). REVIEWER'S CONCLUSIONS: Insufficient evidence exists that treatment of varicocele in men from couples with otherwise unexplained subfertility does improve the couple's spontaneous pregnancy chances. PMID- 11279694 TI - Tamoxifen for early breast cancer. AB - BACKGROUND: There have been many randomised trials of adjuvant tamoxifen among women with early breast cancer, and an updated overview of their results is presented. OBJECTIVES: In this report, the Early Breast Cancer Trialists' Collaborative Group present their third 5-yearly systematic overview (meta analysis) of treatment with tamoxifen. SEARCH STRATEGY: Trial identification procedures for the EBCTCG overviews have been described elsewhere. See under "EBCTCG" in the Breast Cancer Collaborative Review Group module. SELECTION CRITERIA: All randomised trials that began before 1990 and compared adjuvant tamoxifen for any duration versus no such treatment for women with early breast cancer. DATA COLLECTION AND ANALYSIS: In 1995, information was sought on each woman in any randomised trial that began before 1990 of adjuvant tamoxifen versus no tamoxifen before recurrence. Information was obtained and analysed centrally on each of 37,000 women in 55 such trials, comprising about 87% of the worldwide evidence. Compared with the previous such overview, this approximately doubles the amount of evidence from trials of about 5 years of tamoxifen and, taking all trials together, on events occurring more than 5 years after randomisation. MAIN RESULTS: Nearly 8000 of the women had a low, or zero, level of the oestrogen receptor protein (ER) measured in their primary tumour. Among them, the overall effects of tamoxifen appeared to be small, and subsequent analyses of recurrence and total mortality are restricted to the remaining women (18,000 with ER positive tumours, plus nearly 12,000 more with untested tumours, of which an estimated 8000 would have been ER-positive). For trials of 1 year, 2 years, and about 5 years of adjuvant tamoxifen, the proportional recurrence reductions produced among these 30,000 women during about 10 years of follow-up were 21% (SD 3), 29% (SD 2), and 47% (SD 3), respectively, with a highly significant trend towards greater effect with longer treatment (2p<0.00001). The corresponding proportional mortality reductions were 12% (SD 3), 17% (SD 3), and 26% (SD 4), respectively, and again the test for trend was significant (2p=0.003). The absolute improvement in recurrence was greater during the first 5 years, whereas the improvement in survival grew steadily larger throughout the first 10 years. The proportional mortality reductions were similar for women with node-positive and node-negative disease, but the absolute mortality reductions were greater in node-positive women. In the trials of about 5 years of adjuvant tamoxifen the absolute improvements in 10-year survival were 10.9% (SD 2.5) for node-positive (61.4% vs 50.5% survival, 2p<0.00001) and 5.6% (SD 1.3) for node-negative (78.9% vs 73.3% survival, 2p<0.00001). These benefits appeared to be largely irrespective of age, menopausal status, daily tamoxifen dose (which was generally 20 mg), and of whether chemotherapy had been given to both groups. In terms of other outcomes among all women studied (ie, including those with "ER-poor" tumours), the proportional reductions in contralateral breast cancer were 13% (SD 13), 26% (SD 9), and 47% (SD 9) in the trials of 1, 2, or about 5 years of adjuvant tamoxifen. The incidence of endometrial cancer was approximately doubled in trials of 1 or 2 years of tamoxifen and approximately quadrupled in trials of 5 years of tamoxifen (although the number of cases was small and these ratios were not significantly different from each other). The absolute decrease in contralateral breast cancer was about twice as large as the absolute increase in the incidence of endometrial cancer. Tamoxifen had no apparent effect on the incidence of colorectal cancer or, after exclusion of deaths from breast or endometrial cancer, on any of the other main categories of cause of death (total nearly 2000 such deaths; overall relative risk 0.99 [SD 0.05]). REVIEWER'S CONCLUSIONS: For women with tumours that have been reliably shown to be ER negative, adjuvant tamoxifen remains a matter for research. However, some years of adjuvant tamoxifen treatment substantially improves the 10-year survival of women with ER-positive tumours and of women whose tumours are of unknown ER status, with the proportional reductions in breast cancer recurrence and in mortality appearing to be largely unaffected by other patient characteristics or treatments. PMID- 11279695 TI - Intraventricular streptokinase after intraventricular hemorrhage in newborn infants. AB - BACKGROUND: Hydrocephalus following intraventricular hemorrhage (IVH) is still one of the most serious complications of premature birth. Ventriculoperitoneal shunt surgery cannot be carried out early and permanent dependence on a shunt is associated with several serious complications. OBJECTIVES: To determine whether intraventricular streptokinase after intraventricular hemorrhage reduces the risk of permanent shunt dependence, neurodevelopmental disability or death in neonates at risk of, or actually developing, post-hemorrhagic hydrocephalus (PHH). This form of therapy is based on the hypothesis that multiple blood clots in the CSF are the initial cause of post-hemorrhagic ventricular dilatation and lysis of clots could reopen the pathways of circulation and re-absorption of CSF. SEARCH STRATEGY: Pediatric, Neurosurgical and General Medical Journals were handsearched from 1976 until October 2000, as well as the Medline database (via PubMed) and the Cochrane Controlled Trials Registry up to October 2000. Personal contacts were used. SELECTION CRITERIA: One randomized trial evaluated intraventricular streptokinase in infants developing post-hemorrhagic ventricular dilatation. DATA COLLECTION AND ANALYSIS: Details of patient selection, patient allocation and the interventions were extracted. The end-points examined were: ventriculoperitoneal shunt, death, meningitis, and secondary hemorrhage. MAIN RESULTS: When intraventricular streptokinase was compared with conservative management of post hemorrhagic ventricular dilatation, the numbers of deaths and babies with shunt dependence were identical in both groups. No information on the effect of intraventricular streptokinase on disability is available. There is cause for concern about meningitis and secondary intraventricular hemorrhage but numbers are insufficient to quantify the risks. REVIEWER'S CONCLUSIONS: Intraventricular fibrinolytic therapy with streptokinase, given when post-hemorrhagic ventricular dilatation is established, cannot be recommended for neonates following IVH. A conservative approach with CSF drainage applied only to symptomatic raised intracranial pressure seems appropriate. PMID- 11279696 TI - Endotracheal intubation at birth for preventing morbidity and mortality in vigorous, meconium-stained infants born at term. AB - BACKGROUND: On the basis of evidence from non-randomised studies, it has been recommended that all babies born through thick meconium should have their tracheas intubated so that suctioning of their airways can be performed. The aim is to reduce the incidence and severity of meconium aspiration syndrome. However, for term babies who are vigorous at birth endotracheal intubation may be both difficult and unnecessary. OBJECTIVES: To determine if endotracheal intubation and suction of the airways at birth in vigorous term meconium-stained babies is more beneficial than routine resuscitation including aspiration of the oro pharynx. SEARCH STRATEGY: The search was made from Oxford Database of Perinatal Trials, Cochrane Controlled Trials Register, MEDLINE and information obtained from knowledgeable practising neonatologists. SELECTION CRITERIA: Randomised trials which compared a policy of routine vs no (or selective) use of endotracheal intubation and aspiration in the immediate management of vigorous term meconium-stained babies at birth. DATA COLLECTION AND ANALYSIS: Data regarding clinical outcomes including mortality, meconium aspiration syndrome, other respiratory conditions, pneumothorax, need for oxygen supplementation, stridor, convulsions and hypoxic-ischaemic encephalopathy were abstracted and analysed using Revman 4.1. MAIN RESULTS: Four randomised controlled trials of endotracheal intubation at birth in vigorous term meconium-stained babies were identified. Meta-analysis of these trials does not support routine use of endotracheal intubation at birth in vigorous meconium-stained babies to reduce mortality, meconium aspiration syndrome, other respiratory symptoms or disorders, pneumothorax, oxygen need, stridor, HIE and convulsions. However, the event rates of many of these outcomes is low in the reported trials making reliable estimates of treatment effect impossible. REVIEWER'S CONCLUSIONS: Routine endotracheal intubation at birth in vigorous term meconium-stained babies has not been shown to be superior to routine resuscitation including oro-pharyngeal suction. This procedure cannot be recommended for vigorous infants until more research is available. PMID- 11279697 TI - Nebulized racemic epinephrine for extubation of newborn infants. AB - BACKGROUND: Following a period of mechanical ventilation, post-extubation upper airway obstruction can occur in newborn infants, especially after prolonged, traumatic or multiple intubations. The subsequent increase in upper airway resistance may lead to respiratory insufficiency and failure of extubation. The vasoconstrictive properties of epinephrine, and its proven efficacy in the treatment of croup in infants, has led to the routine use of inhaled nebulised epinephrine immediately post-extubation in some neonatal units. It is also recommended for neonates with post-extubation tracheal obstruction and stridor in neonatal and respiratory textbooks and reviews. OBJECTIVES: The primary objective was to assess whether nebulised epinephrine administered immediately after extubation in neonates weaned from IPPV decreases the need for subsequent additional respiratory support. SEARCH STRATEGY: Searches were made using MeSH search terms 'epinephrine' and 'exp infant, newborn'. Databases searched included: MEDLINE from1966 to September 2000; CINAHL from 1982 to September 2000; Current Contents from 1994 to September 2000; and the Cochrane Controlled Trials Register, The Cochrane Library 2000 Issue 3. Previous searches up to March 1999 have included the Oxford Database of Perinatal trials, expert informants and journal hand searching mainly in the English language, expert informant searches in the Japanese language by Prof. Ogawa, previous reviews including cross references, abstracts, and conference and symposia proceedings. SELECTION CRITERIA: All randomised and quasi-randomised control trials in which nebulised epinephrine was compared with placebo immediately post-extubation in newborn infants who have been weaned from IPPV and extubated, with regard to clinically important outcomes (i.e. need for additional respiratory support, increase in oxygen requirement, respiratory distress, stridor or the occurrence of side effects). DATA COLLECTION AND ANALYSIS: No studies met our criteria for inclusion in this review. MAIN RESULTS: No studies were identified which looked at the effect of inhaled nebulised epinephrine on clinically important outcomes in infants being extubated. REVIEWER'S CONCLUSIONS: IMPLICATIONS FOR PRACTICE: There is no evidence either supporting or refuting the use of inhaled nebulised racemic epinephrine in newborn infants. IMPLICATIONS FOR RESEARCH: randomised controlled trials are needed comparing inhaled nebulised racemic epinephrine with placebo in neonates post-extubation. This should be looked at both as a routine treatment post-extubation and as specific treatment for post-extubation upper airway obstruction. Study populations should include the group of infants at highest risk for upper airway obstruction from mucosal swelling because of their small glottic and sub-glottic diameters (ie those infants with birthweights less than 1000 grams). PMID- 11279698 TI - Vaginal misoprostol for cervical ripening and induction of labour. AB - BACKGROUND: Misoprostol (Cytotec, Searle) is a prostaglandin E1 analogue marketed for use in the prevention and treatment of peptic ulcer disease. It is inexpensive, easily stored at room temperature and has few systemic side effects. It is rapidly absorbed orally and vaginally. Although not registered for such use, misoprostol has been widely used for obstetric and gynaecological indications, such as induction of abortion and of labour. This is one of a series of reviews of methods of cervical ripening and labour induction using standardised methodology. OBJECTIVES: To determine the effects of vaginal misoprostol for third trimester cervical ripening or induction of labour. SEARCH STRATEGY: The Cochrane Pregnancy and Childbirth Group trials register, the Cochrane Controlled Trials Register and bibliographies of relevant papers. Date of last search: October 2000. SELECTION CRITERIA: The criteria for inclusion included the following: (1) clinical trials comparing vaginal misoprostol used for third trimester cervical ripening or labour induction with placebo/no treatment or other methods listed above it on a predefined list of labour induction methods; (2) random allocation to the treatment or control group; (3) adequate allocation concealment; (4) violations of allocated management not sufficient to materially affect conclusions; (5) clinically meaningful outcome measures reported; (6) data available for analysis according to the random allocation; (7) missing data insufficient to materially affect the conclusions. DATA COLLECTION AND ANALYSIS: A strategy has been developed to deal with the large volume and complexity of trial data relating to labour induction. This involves a two-stage method of data extraction. The initial data extraction is done centrally, and incorporated into a series of primary reviews arranged by methods of induction of labour, following a standardised methodology. The data will then be extracted from the primary reviews into a series of secondary reviews, arranged by category of woman. To avoid duplication of data in the primary reviews, the labour induction methods have been listed in a specific order, from one to 25. Each primary review includes comparisons between one of the methods (from two to 25) with only those methods above it on the list. MAIN RESULTS: Compared to placebo, misoprostol was associated with increased cervical ripening (relative risk of unfavourable or unchanged cervix after 12 to 24 hours with misoprostol 0.09, 95% confidence interval 0.03 to 0.24). It was also associated with a reduced need for oxytocin (relative risk 0.52, 95% confidence interval 0.41 to 0.68). Misoprostol was more effective than prostaglandin E2 vaginally for labour induction (relative risk of failure to achieve vaginal delivery in 24 hours 0.70, 95% confidence interval 0.61 to 0.81). Oxytocin augmentation was used less often with misoprostol than with prostaglandin E2 vaginally (relative risk 0.65, 95% confidence interval 0.60 to 0.71). Uterine hyperstimulation and meconium stained liquor were more common with misoprostol than with prostaglandin E2. Lower doses of misoprostol compared to higher doses did not show significant differences except for more need for oxytocin augmentation and less uterine hyperstimulation without fetal heart rate changes. Information on women's views is conspiciously lacking. REVIEWER'S CONCLUSIONS: Vaginal misoprostol appears to be more effective in inducing labour than conventional methods of cervical ripening and labour induction. The apparent increase in uterine hyperstimulation is of concern. The studies were not large enough to exclude the possibility of rare but serious adverse effects, particularly uterine rupture, which has been reported following misoprostol use in women with and without previous caesarean section. The authors request information on cases of uterine rupture known to readers. Further research is needed to establish safety. PMID- 11279699 TI - Orthodontic treatment for posterior crossbites. AB - BACKGROUND: 'Posterior crossbite' occurs when the top back teeth bite inside the bottom back teeth. When it affects one side of the mouth the lower jaw may have to move to one side to allow the back teeth to meet together. It is unclear what causes posterior crossbites and they may develop or improve at any time from when the baby teeth come into the mouth to when the adult teeth come through. Several treatments have been recommended to correct this problem. Some treatments widen the upper teeth whilst others are directed at treating the cause of the posterior crossbite e.g. breathing problems or sucking habits. Most treatments have been used at each stage of dental development. OBJECTIVES: The aim of this review was to evaluate orthodontic treatments used to expand the maxillary dentition and correct posterior crossbites. SEARCH STRATEGY: All randomised and controlled clinical trials identified from the Cochrane Controlled Trials Register according to the Oral Health Group Search Strategy and stored in the Cochrane Collaboration Oral Health Group Database of Clinical Trials, a MEDLINE search using the Mesh term Palatal Expansion Technique and relevant free text words, hand searching the British, European and American journals of orthodontics and Angle Orthodontist, and the bibliographies of papers and review articles which reported the outcome of orthodontic treatment to expand the maxillary dentition and/or correct a posterior crossbite that were published as abstracts or papers between 1970 and 1999. SELECTION CRITERIA: All randomised and controlled clinical trials published as full papers or abstracts which reported quantitative data on the outcomes crossbite correction, molar and/or canine expansion, signs and symptoms of temporomandibular joint dysfunction or respiratory disease. DATA COLLECTION AND ANALYSIS: Data were extracted without blinding to the authors, treatments used or results obtained. The first named authors of randomised and controlled clinical trials were written to in an attempt to establish the method of randomisation / allocation and identify unpublished studies. Odds ratio, relative risk, relative risk reduction, absolute risk reduction, the number need to treat and corresponding 95% confidence intervals, were calculated for event data. The weighted mean difference and 95% confidence intervals were calculated for continuous data. MAIN RESULTS: Using the search strategy seven randomised and five controlled clinical trials were identified but following correspondence with the authors, three of the randomised and one of the controlled clinical trials were reclassified giving five randomised and seven controlled clinical trials for inclusion in the review. For the update an additional CCT was found giving five RCTs and eight CCTs for inclusion in this update. Trials comparing occlusal grinding in the primary dentition with/without an upper removable expansion appliance in the mixed dentition versus no treatment, banded versus bonded and two point versus four point rapid maxillary expansion, banded versus bonded slow maxillary expansion, transpalatal arch with/without buccal root torque, an upper removable expansion appliance versus quad-helix were identified. Occlusal grinding in the primary dentition with/without the addition of an upper removable expansion plate, in the mixed dentition for those children who did not respond to grinding, was shown to be effective in preventing a posterior crossbite in the primary dentition from being perpetuated to the mixed and permanent dentitions. No evidence of a difference in treatment effect (molar and canine expansion) between the test and control intervention was found in the trials which compared banded versus bonded and two point versus four point rapid maxillary expansion, banded versus bonded slow maxillary expansion, transpalatal arch with/without buccal root torque, or upper removable expansion appliance versus quad-helix. Insufficient data were provided in the paper comparing two point versus four point rapid maxillary expansion to allow a formal analysis. REVIEWER'S CONCLUSIONS: The evidence from the trials reported by Lindner (1989); Thilander (1984) suggests that removal of premature contacts of the baby teeth is effective in preventing a posterior crossbite from being perpetuated to the mixed dentition and adult teeth. When grinding alone is not effective, using an upper removable expansion plate to expand the top teeth will decrease the risk of a posterior crossbite from being perpetuated to the permanent dentition. The comparisons of treatments made in the trials reported by Asanza (1997); Sandikcioglu (1997); Mossaz-Joelson (1989); Ingervall (1995); Schneidman (1990) were inconclusive so recommendations for clinical practice can not be made based on the results of these trials. (ABSTRACT TRUNCATED) PMID- 11279700 TI - Buflomedil for intermittent claudication. AB - BACKGROUND: Intermittent claudication is pain, caused by chronic occlusive arterial disease, that develops in a limb during exercise and is relieved with rest. Buflomedil is a vasoactive agent claimed to have beneficial effects on the microcirculation. It is used chiefly to treat peripheral vascular disease and to a lesser extent for cerebrovascular arterial disease. However, its clinical efficacy for intermittent claudication has not yet been critically examined. OBJECTIVES: To evaluate the available evidence on the efficacy of buflomedil for intermittent claudication. SEARCH STRATEGY: We searched Medline, International Pharmaceutical Abstracts (IPA) and the Cochrane Controlled Trials Register. Abbott Laboratories, the distributor of buflomedil, was asked to provide reports of controlled clinical trials. Reference lists of retrieved articles were checked, and enquiries sent to authors of known trials, to identify additional trials. Finally, we conducted a Science Citation Index search. SELECTION CRITERIA: Trial reports had to be double-blinded, randomized, and conformed to our PIO-criteria (Patients, Intervention, Outcome) to be considered for inclusion. Patients were required to have proven intermittent claudication (Fontaine stage II); the intervention was to be oral administration of buflomedil compared to placebo; and outcomes had to include pain-free walking distance (PFWD) and maximum walking distance (MWD) analysed by standardized exercise test. DATA COLLECTION AND ANALYSIS: Searches of bibliographic databases yielded three eligible randomized controlled trials (RCTs) and a meta-analysis referring to nine eligible trials. Two of these nine trials had already been identified; two had been published in journals not referenced in traditional bibliographic indexes; and five were unpublished. Despite multiple requests, only one of the five unpublished trials was provided by the author of the meta-analysis, the other four could not be retrieved. Four of the six eligible trials retrieved were subsequently excluded after quality evaluation. Data on walking distances were extracted from the two remaining trials. Differences in incremental gain between active and placebo groups for PFWD and MWD with their confidence intervals were calculated. MAIN RESULTS: Both RCTs showed moderate improvements in PFWD for patients on buflomedil. In one trial this improvement (75 m, 95% CI 37-114) was statistically significant, but in the other, with a wholly diabetic population, it was non-significant (81m, 95% CI -9-170) compared to placebo. For both RCTs the gains in MWD were statistically significant, but with wide confidence intervals (81 m, 95% CI 30-131; and 171 m, 95% CI 27-316 respectively). Pooling of the data was not attempted. REVIEWER'S CONCLUSIONS: There is little evidence available to evaluate the efficacy of buflomedil for intermittent claudication. Most available trials are of poor quality and were excluded. The two trials included showed moderately positive results but these are undermined by publication bias since we know of another four unpublished, irretrievable, and inconclusive studies. There is a lack evidence for the efficacy of buflomedil in intermittent claudication. PMID- 11279701 TI - Manual therapy for asthma. AB - BACKGROUND: A variety of manual therapies with similar postulated biologic mechanisms of action are commonly used to treat patients with asthma. Manual therapy practitioners are also varied, including physiotherapists, respiratory therapists, chiropractic and osteopathic physicians. A systematic review across disciplines is warranted. OBJECTIVES: To evaluate the evidence for the effects of manual therapies for treatment of patients with bronchial asthma. SEARCH STRATEGY: Trials were searched in computerized general (EMBASE, CINAHL and MEDLINE) and specialized databases (Cochrane Complementary Medicine Field, Cochrane Rehabilitation Field, ICL, and MANTIS). In addition, bibliographies from included studies were assessed, and authors of known studies were contacted for additional information about published and unpublished trials. Date of most recent search: December 1998. SELECTION CRITERIA: Trials were included if they: (1) were randomised; (2) included asthmatic children or adults; (3) examined one or more types of manual therapy; and (4) included clinical outcomes. DATA COLLECTION AND ANALYSIS: All three reviewers independently extracted data and assessed trial quality using a standard form. MAIN RESULTS: From an initial 316 unique citations, 48 full text articles were retrieved and evaluated, which resulted in nine citations to five RCTs (290 patients) suitable for inclusion. Trials could not be pooled statistically because studies that addressed similar interventions used disparate patient groups or outcomes. The methodological quality of one of two trials examining chiropractic manipulation was good and neither trial found significant differences between chiropractic spinal manipulation and a sham manoeuvre on any of the outcomes measured. Quality of the remaining three trials was poor. One small trial compared massage therapy with a relaxation control group and found significant differences in many of the lung function measures obtained. However, this trial had poor reporting characteristics and the data have yet to be confirmed. One small trial compared chest physiotherapy to placebo and one small trial compared footzone therapy to a no treatment control. Neither trial found differences in lung function between groups. A further search conducted in July 2000 did not yield any more studies REVIEWER'S CONCLUSIONS: There is insufficient evidence to support the use of manual therapies for patients with asthma. There is a need to conduct adequately sized RCTs that examine the effects of manual therapies on clinically relevant outcomes. Future trials should maintain observer blinding for outcome assessments, and report on the costs of care and adverse events. Currently, there is insufficient evidence to support or refute the use of manual therapy for patients with asthma. PMID- 11279702 TI - Counselling for depression in primary care. AB - BACKGROUND: There is wide clinician and patient support for counselling in primary care, particularly in the UK. This review examines the effectiveness and cost effectiveness of counselling for psychological and psychosocial problems in the primary care setting. OBJECTIVES: To assess the effects of counselling in primary care by reviewing cost and outcome data for patients with psychological and psychosocial problems considered suitable for counselling. SEARCH STRATEGY: The search strategy included electronic searching of databases (including the CCDAN Register of RCTs and CCTs) along with handsearching of a specialist journal. Published and unpublished sources (clinical trials, books, dissertations, agency reports etc.) were searched, and their reference lists scanned. Contact was made with subject experts and CCDAN members. SELECTION CRITERIA: Randomised and controlled patient preference trials comparing counselling in primary care with usual general practitioner care for patients with psychological and psychosocial problems considered suitable for counselling. Trials completed before the end of April 1998 were included in the review. DATA COLLECTION AND ANALYSIS: Trials were independently assessed by at least two reviewers for appropriateness of inclusion and methdological quality. MAIN RESULTS: Four trials, involving 678 participants, of whom 487 were followed up, were included. Data for psychological symptom levels (four trials) were pooled statistically. Patients receiving counselling had significantly better psychological symptom levels post intervention than patients receiving usual general practitioner care (standardised mean difference -0.30, 95% CI, (-0.49 to 0.11). The effect remained statistically significant when the results from studies with less rigorous methodology were excluded in a sensitivity analysis. Patients who received counselling tended to be more satisfied with their treatment (three trials). Health service utilisation data were reported in all trials reviewed, but only one trial undertook a cost analysis. No clear cost advantage was associated with either counselling or usual general practice care. REVIEWER'S CONCLUSIONS: Patients who received counselling were more likely to have improved psychological symptom levels than those who did not receive counselling. Levels of satisfaction with counselling were high. There is limited information about the cost effectiveness of counselling, with one study reporting no clear cost advantage with either counselling or general practice care. The four trials included in this review were all pragmatic trials of counselling in primary care in the UK, which reflect the reality of clinical provision in this context. There were methdological weaknesses identified in the studies, which should be taken into account when considering the results. The evidence base will be extended by trials of counselling which are nearing completion. PMID- 11279703 TI - Tamoxifen for relapse of ovarian cancer. AB - BACKGROUND: Tamoxifen is an important drug for treating breast cancer. Ovarian cancer cells are known to possess receptors for hormones and may thus also respond to tamoxifen. OBJECTIVES: Tamoxifen is used to treat breast cancer in women whose tumours have oestrogen receptors. Since ovarian cancers also commonly have oestrogen receptors, it has been suggested that tamoxifen may be of some benefit. The objective of this review was to assess the effects of tamoxifen in women with relapsed ovarian cancer. SEARCH STRATEGY: We searched the Cochrane Gynaecological Cancer Group trials register and references from relevant articles. We also contacted researchers and drug companies. SELECTION CRITERIA: Randomised and non-randomised studies of tamoxifen in women with ovarian cancer who have not responded to conventional chemotherapy. Only trials involving 10 or more patients were included. DATA COLLECTION AND ANALYSIS: One reviewer assessed eligibility and extracted data from non-randomised studies. Two reviewers were to have independently assessed the quality and extracted data from any randomised trials found. MAIN RESULTS: Eleven non-randomised series, one non-randomised phase two study and two randomised trials were included. Only observational data from women treated with tamoxifen are reported. Sixty of 623 women (9.6%) treated with tamoxifen achieved an objective response to treatment. However this varied from 0% to 56% in different studies. Stable disease, for variable periods of four weeks or more, was observed in 131 of 411 (31.9%) women from eight studies. There were not enough data to assess duration of response, survival, or the palliative effect of tamoxifen on symptom control or quality of life. REVIEWER'S CONCLUSIONS: There is some evidence from observational studies that tamoxifen may produce a response in a modest proportion of women with relapsed ovarian cancer. However, there are no reliable data from randomised controlled trials. PMID- 11279704 TI - D-Penicillamine for preventing retinopathy of prematurity in preterm infants. AB - BACKGROUND: Retinopathy of prematurity remains a common problem. A low rate of this disorder was unexpectedly observed among infants treated with intravenous d penicillamine to prevent hyperbilirubinemia. This observation led to the investigation of its use to prevent retinopathy of prematurity. OBJECTIVES: To answer the question: Among very low birth weight infants, what is the effect of prophylactic administration of d-penicillamine on the incidence of acute ROP or severe ROP, and side effects including death? SEARCH STRATEGY: Searches were made of multiple electronic databases, previous reviews including cross references, abstracts, conference/symposia proceedings, and expert informants. The search was updated to November 2000. SELECTION CRITERIA: Randomized or quasi-randomized controlled trials that administered d-penicillamine to infants less than 2000g birth weight within the day following birth were considered relevant to this review. Additional case series were examined for potential side effects. DATA COLLECTION AND ANALYSIS: Data on clinical outcomes were excerpted by 3 reviewers independently, and consensus reached. Data analysis was conducted according to the standards of the Neonatal Cochrane Review Group. MAIN RESULTS: Two randomized trials on the effects on ROP were identified. When combined, they showed a significantly lower incidence of acute ROP in the treated infants, relative risk of 0.09, 95% CI [0.01,0.71]. Severe stages of ROP could not be analyzed. There was no effect on death rates, relative risk 0.99 95% CI [0.70,1.39]. No side effects were reported, and follow up at one year revealed no significant differences in spasticity or developmental delay, although there were more rehospitalizations among the controls. In other reports of using d-penicillamine in over 140 infants for hyperbilirubinemia, skin rashes were reported in 2 infants and one had vomiting that may have been related. REVIEWER'S CONCLUSIONS: D-penicillamine is unlikely to affect survival, and may reduce the incidence of acute ROP among survivors. Studies to date justify further investigation of this drug in a broader population; careful attention to possible side effects is needed. PMID- 11279705 TI - Moderately early (7-14 days) postnatal corticosteroids for preventing chronic lung disease in preterm infants. AB - BACKGROUND: Corticosteroids have been used late in the neonatal period to treat chronic lung disease (CLD) in preterm babies and early to try to prevent it. CLD is likely to be the result of persisting inflammation in the lung and the use of powerful anti-inflammatory drugs like dexamethasone has some rationale. Early use tends to be associated with increased adverse effects so that studies of moderately early treatment (7-14 days postnatal) might have the dual benefits of fewer side effects and onset of action before chronic inflammation is established. OBJECTIVES: To determine if moderately early (7-14 days) postnatal corticosteroid treatment vs control (placebo or nothing) is of benefit in the prevention and/or treatment of early chronic lung disease in the preterm infant. SEARCH STRATEGY: Randomised controlled trials of postnatal corticosteroid therapy were sought from the Oxford Database of Perinatal Trials, Cochrane Database of Controlled Trials, Medline, hand searching paediatric and perinatal journals, examining previous review articles and information received from practising neonatologists. SELECTION CRITERIA: Randomised controlled trials of postnatal corticosteroid treatment from 7-14 days of birth in high risk preterm infants were selected for this review. DATA COLLECTION AND ANALYSIS: Data regarding clinical outcomes including mortality up to 28 days and before discharge, failure to extubate, mortality or chronic lung disease at 28 days and 36 weeks, CLD at 28 days and 36 weeks, infection, hyperglycaemia, hypertension, severe retinopathy of prematurity (ROP), pneumothorax, necrotizing enterocolitis (NEC), gastrointestinal bleeding, severe intraventricular haemorrhage (IVH), hypertrophic cardiomyopathy, late rescue with dexamethasone and abnormal neurological examination at follow-up were abstracted and analysed using RevMan 4.1. MAIN RESULTS: Moderately early steroid treatment (vs placebo or nothing) reduced mortality by 28 days, chronic lung disease at 28 days and 36 weeks, and death or chronic lung disease at 28 days or 36 weeks. Earlier extubation was facilitated. There was no significant effect on the rates of pneumothorax, severe ROP, and NEC. Adverse effects included hypertension, hyperglycaemia, gastrointestinal bleeding, hypertrophic cardiomyopathy and infection. Steroid treated infants were less likely to need late rescue with dexamethasone. There was only one small study of longterm follow-up and it did not show any increase in adverse neurological outcome. REVIEWER'S CONCLUSIONS: Moderately early corticosteroid therapy (started at 7-14 days) reduces neonatal mortality and CLD, but at the cost of important short term adverse effects. No reliable evidence concerning long term effects is provided by the trials included in this review. In view of the evidence of an important increase in cerebral palsy and other adverse neurodevelopmental outcomes from trials in which postnatal steroids were begun either earlier or later than 7-14 days, there are reasonable grounds for extending this concern to moderately early initiation of steroid therapy. More research is urgently needed, including long term follow-up of survivors included in previous and any future trials, before the benefits and risks of postnatal steroid treatment, including initiation at 7-14 days, can be reliably assessed (See DART study; ~~ Doyle 2000~~). PMID- 11279707 TI - Mannitol for acute stroke. AB - BACKGROUND: Mannitol is an osmotic agent and a free radical scavenger so it might decrease oedema and tissue damage in stroke. OBJECTIVES: To test whether treatment with mannitol reduces short and long-term case fatality and dependency after acute ischaemic stroke or cerebral parenchymal haemorrhage. SEARCH STRATEGY: We searched the Cochrane Stroke Group Specialised Trials Register. In addition to this, supplementary MEDLINE searches were performed. The Chinese Stroke Trials Register was checked and the Latin-American databank LILACS was searched with the search term MANNITOL and its variations in the Portuguese and Spanish languages. A search was performed of Masters and Ph.D. degree theses in the databank of Sao Paulo University, and in abstracts of medical congresses on neurology and neurosurgery from 1965 to 1997 in Brazil. SELECTION CRITERIA: Truly randomised unconfounded clinical trials comparing the effect of mannitol with placebo or open control in patients with acute ischaemic stroke or parenchymal haemorrhage were eligible for inclusion. DATA COLLECTION AND ANALYSIS: Two reviewers independently selected the trials to be included in the review. After reaching an agreement on which trials to include, two of the reviewers extracted data from the trials and performed the data analysis. Accuracy of data extraction was checked by comparing the results. Included trials were tabulated for methodological quality including the method of randomisation and blinding, and stating if CT was performed, if patients were lost to follow-up and if intention to-treat analysis was performed. Data synthesis and analysis was performed using the Cochrane Review Manager software. MAIN RESULTS: Only one trial fulfilled the inclusion criteria. The number of included patients was small (36 treated and 41 controls) and the follow up was short. Neither beneficial nor harmful effects of mannitol could be proved. Case fatality, the proportion of dependent patients at the end of the follow up and side effects were not reported and were not available from the investigators. The planned outcome analyses and sensitivity analyses could not be performed due to lack of appropriate trials. REVIEWER'S CONCLUSIONS: There is currently not enough evidence to decide whether the routine use of mannitol in acute stroke would result in any beneficial or harmful effect. The routine use of mannitol in all patients with acute stroke is not supported by any evidence from randomised controlled clinical trials. Further trials are needed to confirm or refute the routine use of mannitol in acute stroke. PMID- 11279706 TI - Early postnatal (<96 hours) corticosteroids for preventing chronic lung disease in preterm infants. AB - BACKGROUND: Chronic lung disease (CLD) remains a major problem in neonatal intensive care units. Persistent inflammation in the lungs is the most likely underlying pathogenesis. Corticosteroids have been used to either prevent or treat CLD because of their potent anti-inflammatory effects. OBJECTIVES: To determine if postnatal corticosteroid treatment is of benefit in the prevention of chronic lung disease (CLD) in the preterm infant. This review examines the outcome of trials where preterm infants at risk of CLD were given postnatal steroids within 96 hours after birth. SEARCH STRATEGY: Randomised controlled trials of postnatal corticosteroid therapy were sought from the Oxford Database of Perinatal Trials, the Cochrane Controlled Trials Register, Medline, hand searching paediatric and perinatal journals, examining previous review articles and information received from practising neonatologists. SELECTION CRITERIA: Randomised controlled trials of postnatal corticosteroid treatment within 96 hours of birth (early) in high risk preterm infants were selected for this review. DATA COLLECTION AND ANALYSIS: Data regarding clinical outcomes including mortality, failure to extubate, pulmonary air leak, survival without chronic lung disease, CLD defined at 28 days postnatal age and 36 weeks post menstrual age, patent ductus arteriosus (PDA), severe intraventricular haemorrhage (IVH), periventricular leucomalacia (PVL), infection, hyperglycaemia, hypertension, severe retinopathy of prematurity (ROP), necrotising enterocolitis (NEC), gastrointestinal bleeding, intestinal perforation and long-term outcomes were abstracted and analysed using RevMan 4.1. MAIN RESULTS: Nineteen randomised controlled trials of early postnatal corticosteroid treatment of preterm babies at risk of developing CLD were identified. A meta-analysis of these trials demonstrates benefits as regards earlier extubation, decreased risks of CLD at both 28 days and 36 weeks, death or CLD at 28 days, PDA and pulmonary air leak. There were no differences in the rates of neonatal mortality, infection, severe ROP, severe IVH, PVL, NEC and pulmonary haemorrhage. Gastrointestinal bleeding and intestinal perforation were important adverse effects and the risks of hyperglycaemia and hypertension were also increased. In the two trials which have reported late outcomes, several adverse neurological effects were found at follow up examinations of survivors treated with early steroids: abnormal neurological examination, cerebral palsy and developmental delay. REVIEWER'S CONCLUSIONS: The benefits of early postnatal corticosteroid treatment (< 96 hours) may not outweigh the known or potential adverse effects of this treatment. Adverse gastrointestinal effects early in the neonatal period and adverse neurological outcomes seen at follow-up mean that current use of early postnatal steroids needs to be reconsidered. There is a compelling need for the long term follow-up and reporting of late outcomes, especially neurological and developmental outcomes, among surviving infants who participated in all randomised trials of early postnatal corticosteroid treatment. The role of inhaled steroids remains to be elucidated. PMID- 11279708 TI - Nerve blocks (subcostal, lateral cutaneous, femoral, triple, psoas) for hip fractures. AB - BACKGROUND: Various nerve blocks using local anaesthetic agents have been used in order to reduce pain after hip fracture. OBJECTIVES: To determine the effects of nerve blocks (inserted either pre-operatively, operatively or post-operatively) as part of the treatment for a hip fracture. SEARCH STRATEGY: The Cochrane Musculoskeletal Injuries Group specialised trials register, MEDLINE, and bibliographies of trial reports were searched. Date of the most recent search: October 2000. SELECTION CRITERIA: Randomised and quasi-randomised trials involving the use of nerve blocks as part of the care of a hip fracture patient. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality, by use of a nine item scale, and extracted data. Wherever appropriate, results of outcome measures were pooled. MAIN RESULTS: Seven randomised or quasi-randomised trials involving 269 patients were included. Two trials related to insertion of a nerve block pre-operatively and the remaining five to peri-operative insertion. Nerve blocks resulted in a reduction of the quantity of parenteral or oral analgesia administered to control pain from the fracture/operation or during surgery and a reduction in reported pain levels. It was not possible to demonstrate if this reduction in analgesia use was associated with any other clinical benefit. REVIEWER'S CONCLUSIONS: Because of the small number of patients included in this review and the differing type of nerve blocks and timing of insertion, it is not possible to determine if nerve blocks confer any significant benefit when compared with other analgesic methods as part of the treatment of a hip fracture. Further trials with larger numbers of patients and full reporting of clinical outcomes would be justified. PMID- 11279709 TI - Mycobacterium vaccae immunotherapy for treating tuberculosis. AB - BACKGROUND: Some authorities have advocated Mycobacterium vaccae immunotherapy for treating tuberculosis and other infections caused by mycobacteria. OBJECTIVES: To assess the effects of Mycobacterium vaccae as an adjunct to chemotherapy for treating tuberculosis. SEARCH STRATEGY: We searched the Cochrane Infectious Diseases Group trials register, the Cochrane Controlled Trials register, Medline, Embase and reference lists of articles. We also contacted organisations and individuals working in the field. SELECTION CRITERIA: Randomised and quasi-randomised trials using whole, killed Mycobacterium vaccae for patients with tuberculosis. DATA COLLECTION AND ANALYSIS: One reviewer assessed trial quality and extracted data. MAIN RESULTS: Six trials met the inclusion criteria. There was no effect on mortality (three trials, OR 1.01, 95% CI 0.51 to 1.99). No consistent effect on sputum negativity or sputum culture was shown. Most immunotherapy recipients experienced local adverse reactions (two trials, OR 18.2, 95% CI 9 to 37), some of which progressed to ulceration and scarring. REVIEWER'S CONCLUSIONS: Immunotherapy with Mycobacterium vaccae does not appear to benefit patients with tuberculosis. PMID- 11279710 TI - Acupuncture for idiopathic headache. AB - BACKGROUND: Acupuncture is widely used for the treatment of headache, but its effectiveness is controversial. OBJECTIVES: To determine whether acupuncture is: more effective than no treatment - more effective than 'sham' (placebo) acupuncture - as effective as other interventions used to treat idiopathic (primary) headaches. SEARCH STRATEGY: Electronic searches were performed in MEDLINE, EMBASE, the Cochrane Controlled Trials Register, and the database of the Cochrane Field for Complementary Medicine. We also contacted researchers in the field and checked the bibliographies of all articles obtained. SELECTION CRITERIA: Randomized or quasi-randomized clinical trials comparing acupuncture with any type of control intervention for the treatment of idiopathic (primary) headaches were included. DATA COLLECTION AND ANALYSIS: Information on patients, interventions, methods, and results was extracted by at least two independent reviewers using a pre-tested standard form. Results on headache frequency and intensity were summarized descriptively. Responder rate ratios (responder rate in treatment group/responder rate in control group) were calculated as a crude indicator of results for sham-acupuncture-controlled trials. Quantitative meta analysis was not possible due to trial heterogeneity and insufficient reporting. MAIN RESULTS: Twenty-six trials including a total of 1151 patients (median, 37; range, 10-150) met the inclusion criteria. Sixteen trials were conducted among patients with migraine, six among patients with tension-type headache, and four among patients with various types of headaches. The majority of trials had methodological and/or reporting shortcomings. In eight of the 16 trials comparing true and sham (placebo) acupuncture in migraine and tension-type headache patients, true acupuncture was reported to be significantly superior; in four trials there was a trend in favor of true acupuncture; and in two trials there was no difference between the two interventions. (Two trials were uninterpretable.) The 10 trials comparing acupuncture with other forms of treatment yielded contradictory results. REVIEWER'S CONCLUSIONS: Overall, the existing evidence supports the value of acupuncture for the treatment of idiopathic headaches. However, the quality and amount of evidence are not fully convincing. There is an urgent need for well-planned, large-scale studies to assess the effectiveness and cost-effectiveness of acupuncture under real-life conditions. PMID- 11279711 TI - Routine perineal shaving on admission in labour. AB - BACKGROUND: Pubic or perineal shaving is a procedure performed before birth in order to lessen the risk of infection if there is a spontaneous perineal tear or if an episiotomy is performed. OBJECTIVES: To assess the effects of routine perineal shaving on admission in labour on maternal and neonatal outcomes, according to the best available evidence. SEARCH STRATEGY: The register of clinical trials maintained and updated by the Cochrane Pregnancy and Childbirth Group. In addition, the Cochrane Controlled Trials Register was searched. Date of last search: July 2000. SELECTION CRITERIA: All controlled trials (including quasi randomised) which compared perineal shaving versus no perineal shaving were included in the review. DATA COLLECTION AND ANALYSIS: Trials under consideration were evaluated for methodological quality and appropriateness for inclusion, without consideration of their results. MAIN RESULTS: Only two trials fulfilled the prespecified criteria. In the earlier trial, 389 women were alternately allocated to receive either skin preparation and perineal shaving (control) or clipping of vulval hair only (experimental). In the second trial, which included 150 participants, perineal shaving was compared with the cutting of long hairs for procedures only. The primary outcome for both trials was maternal febrile morbidity. No differences were found (combined odds ratio (OR) 1.26, 95% confidence interval (CI) 0.75, 2.12). In the smaller trial, fewer women who had not been shaved had gram negative bacterial colonisation compared with women who had been shaved (OR 0.43, 95% CI 0.20, 0.92). REVIEWER'S CONCLUSIONS: There is insufficient evidence to recommend perineal shaving for women on admission in labour. PMID- 11279712 TI - Mucolytics for bronchiectasis. AB - BACKGROUND: Bronchiectasis is predominantly an acquired disease process representing the end stage of a variety of unrelated pulmonary insults. It is defined as a persistent irreversible dilatation and distortion of medium-sized bronchi. Patients diagnosed with bronchiectasis frequently have difficulty exporating the infected sputum. Mucolytic agents target hyper-secretion or changed physiochemical properties of sputum to make it easier to clear. One drug, recombinant human DNase, breaks down the DNA that is released at the site of infection by neutrophils. OBJECTIVES: The objective of this review was to assess the effects of ingested or inhaled mucolytics in people with bronchiectasis. SEARCH STRATEGY: We searched the Cochrane Airways Group trials register, reference lists of relevant articles. We also contacted experts in the field and drug companies. SELECTION CRITERIA: Randomised trials of mucolytic treatment in people with bronchiectasis but not cystic fibrosis. DATA COLLECTION AND ANALYSIS: Data extraction was performed independently by two reviewers. Study authors were contacted for confirmation. MAIN RESULTS: Three trials were included, but none of their data could be aggregated in a meta analysis. Compared to placebo, high doses of bromhexine with antibiotics eased difficulty in expectoration (weighted mean difference -0.53, 95% confidence interval -0.81 to -0.25 at 16 days). There was also a reduction in sputum production with bromhexine (weighted mean difference -21.5%, 95% confidence interval -38.9 to -4.1 % at day 16). Compared to placebo, recombinant human DNase showed no difference in forced expiratory volume or forced vital capacity in one study and was reported to have a significant negative effect on forced expiratory volume in another study. Adverse effects, including influenza-like symptoms, were more common in the group receiving recombinant human DNase. REVIEWER'S CONCLUSIONS: There is not enough evidence to evaluate the routine use of mucolytics for bronchiectasis. High doses of bromhexine coupled with antibiotics may help with sputum production and clearance. PMID- 11279713 TI - Interventions for educating children who have attended the emergency room for asthma. AB - BACKGROUND: Asthma is one of the most common reasons for paediatric admissions to hospital, with substantial cost to the community. There is some evidence to suggest that many hospital admissions could be prevented with effective education about asthma and its management. OBJECTIVES: To conduct a systematic review of the literature in order to identify whether asthma education leads to improved health outcomes in children who have attended the emergency department for asthma. SEARCH STRATEGY: We searched the Cochrane Airways Group trials register, including Medline, Embase, and Cinahl databases, and reference lists of trials and review articles. SELECTION CRITERIA: Randomised controlled trials or controlled clinical trials of asthma education for children who had attended the emergency department for asthma, with or without hospitalisation, within the previous 12 months. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality and extracted data. Study authors were contacted for additional information. MAIN RESULTS: Eight trials involving 1407 patients were included, in all the education was provided by nurses or researchers. Compared to control (usual care or lower intensity education) education did not reduce subsequent emergency department (ED) visits [4 trials; relative risk (RR)= 0.87, 95% confidence interval (CI) 0.37 to 2.08], hospital admissions [5 trials; RR=0.74, 95% CI 0.38 to 1.46] and unscheduled doctor visits [5 trials; RR= 0.74, 95% CI 0.49 to 1.12). Each analysis showed evidence of heterogeneity among the studies (P<0.01). Subgroup analyses by the overall difference in scale of intervention between treatment and control groups (comprehensive programme versus information only) or the timing of the intervention/recruitment (early versus delayed) gave similar results to the main analysis and still revealed significant heterogeneity between trials. REVIEWER'S CONCLUSIONS: On the basis of the published trials, there is no firm evidence to support the use of asthma education for children who have attended the emergency department for asthma as a means of reducing subsequent ED visits, hospital admissions or unscheduled doctor visits. Some trials appeared to show clear evidence of benefit, but reasons for differences between these and the negative studies is not clear. More research is required PMID- 11279714 TI - Antibiotic prophylaxis for intrauterine contraceptive device insertion. AB - BACKGROUND: Concern about the risk of upper genital tract infection (pelvic inflammatory disease) often limits use of the IUD, a highly effective contraceptive. Prophylactic antibiotic administration around the time of induced abortion significantly reduces the risk of postoperative endometritis.(Sawaya, 1996) Since the risk of IUD-related infection is limited to the first few weeks to months after insertion,(Lee, 1983; Farley, 1992) contamination of the endometrial cavity at the time of insertion(Mishell, 1966) appears to be the mechanism, rather than the IUD or string itself. Thus, antibiotic administration before IUD insertion might reduce the risk of upper genital tract infection from passive introduction of bacteria at insertion. OBJECTIVES: To assess the effectiveness of prophylactic antibiotic administration before IUD insertion in reducing IUD-related complications and discontinuations within three months of insertion. The primary outcome was pelvic inflammatory disease (four reports) or early removals of the device (two reports). SEARCH STRATEGY: We searched both MEDLINE and EMBASE, handsearches of journals through CENTRAL, and lists of references. We also wrote to international experts in the field to identify unpublished studies. SELECTION CRITERIA: We included randomized controlled trials using any antibiotic compared with a placebo. We found four such trials; two had pilot study data available. DATA COLLECTION AND ANALYSIS: DATA EXTRACTION: We used searches of MEDLINE, EMBASE, and handsearches of journals available through CENTRAL. We also reviewed lists of references in original research and in review articles. We wrote to experts to identify unpublished trials and made telephone calls to authors to supply missing information. Two independent reviewers abstracted data. We assessed the validity of each study using methods suggested in the Cochrane Handbook. DATA SYNTHESIS: We generated 2x2 tables for the principal outcome measures. We used the Peto modified Mantel-Haenszel technique to calculate odds ratios and assessed statistical heterogeneity between studies. MAIN RESULTS: The odds ratios for pelvic inflammatory disease associated with use of prophylactic doxycycline or azithromycin compared with placebo or no treatment was 0.89 (95%CI 0.53-1.51). Use of prophylaxis was associated with a small reduction in unscheduled vists to the provider (OR 0.82; 95% CI 0.70-0.98). Use of doxycycline or azithromycin had little effect on the likelihood of removal of the IUD within 90 days of insertion (OR 1.05; 95% CI 0.68-1.63). Significant heterogeneity did not exist between studies. REVIEWER'S CONCLUSIONS: Use of either doxycycline 200 mg or azithromycin 500 mg by mouth before IUD insertion confers little benefit. While the reduction in unscheduled visits to the provider was marginally significant, the cost-effectiveness of routine prophylaxis remains questionable. A uniform finding in these trials was the low risk of IUD associated infection, with or without use of antibiotic prophylaxis. PMID- 11279715 TI - Dietary advice given by a dietitian versus other health professional or self-help resources to reduce blood cholesterol. AB - BACKGROUND: The average level of blood cholesterol is an important determinant of the risk of coronary heart disease. Blood cholesterol can be reduced by dietary means. Although dietitians are trained to provide dietary advice, for practical reasons it is also given by other health professionals and occasionally through the use of self-help resources. OBJECTIVES: To assess the effects of dietary advice given by a dietitian compared with another health professional, or the use of self-help resources, in reducing blood cholesterol in adults. SEARCH STRATEGY: We searched The Cochrane Library (to Issue 2 1999), MEDLINE (1966 to January 1999), EMBASE (1980 to December 1998), Cinahl (1982 to December 1998), Human Nutrition (1991 to 1998), Science Citation Index, Social Sciences Citation Index, hand searched conference proceedings on nutrition and heart disease, and contacted experts in the field. SELECTION CRITERIA: Randomised trials of dietary advice given by a dietitian compared with another health professional or self help resources. The main outcome was difference in blood cholesterol between dietitian groups compared with other intervention groups. DATA COLLECTION AND ANALYSIS: Two reviewers independently extracted data and assessed study quality. MAIN RESULTS: Eleven studies with 12 comparisons were included, involving 704 people receiving advice from dietitians, 486 from other health professionals and 551 people using self-help leaflets. Four studies compared dietitian with doctor, seven with self-help resources, and one compared dietitian with nurse. Participants receiving advice from dietitians experienced a greater reduction in blood cholesterol than those receiving advice only from doctors (-0.25 mmol/L (95% CI -0.37, -0.12 mmol/L)). There was no statistically significant difference in change in blood cholesterol between dietitians and self-help resources (-0.10 mmol/L (95% CI -0.22, 0.03 mmol/L)). No statistically significant differences were detected for secondary outcome measures between any of the comparisons with the exception of dietitian versus nurse for HDLc, where the dietitian groups showed a greater reduction (-0.06 mmol/L (95% CI -0.11, -0.01)). No significant heterogeneity between the studies was detected. REVIEWER'S CONCLUSIONS: Dietitians were better than doctors at lowering blood cholesterol in the short to medium term, but there was no evidence that they were better than self-help resources. The results should be interpreted with caution as the studies were not of good quality and the analysis was based on a limited number of trials. More evidence is required to assess whether change can be maintained in the longer term. There was no evidence that dietitians provided better outcomes than nurses. PMID- 11279716 TI - Pelvic floor muscle training for urinary incontinence in women. AB - BACKGROUND: Pelvic floor muscle training is the most commonly recommended physical therapy treatment for women with stress leakage of urine. It is also used in the treatment of women with mixed incontinence, and less commonly for urge incontinence. Adjuncts, such as biofeedback or electrical stimulation, are also commonly used with pelvic floor muscle training. The content of pelvic floor muscle training programmes is highly variable. OBJECTIVES: To determine the effects of pelvic floor muscle training for women with symptoms or urodynamic diagnoses of stress, urge and mixed incontinence, in comparison to no treatment or other treatment options. SEARCH STRATEGY: Search strategy: We searched the Cochrane Incontinence Group trials register (May 2000), Medline (1980 to 1998), Embase (1980 to 1998), the database of the Dutch National Institute of Allied Health Professions (to 1998), the database of the Cochrane Rehabilitation and Related Therapies Field (to 1998), Physiotherapy Index (to 1998) and the reference lists of relevant articles. We handsearched the proceedings of the International Continence Society (1980 to 2000). We contacted investigators in the field to locate studies. Date of the most recent searches: May 2000. SELECTION CRITERIA: Randomised trials in women with symptoms or urodynamic diagnoses of stress, urge or mixed incontinence that included pelvic floor muscle training in at least one arm of the trial. DATA COLLECTION AND ANALYSIS: Two reviewers assessed all trials for inclusion/exclusion and methodological quality. Data were extracted by the lead reviewer onto a standard form and cross checked by another. Disagreements were resolved by discussion. Data were processed as described in the Cochrane Handbook. Sensitivity analysis on the basis of diagnosis was planned and undertaken where appropriate. MAIN RESULTS: Forty-three trials met the inclusion criteria. The primary or only reference for 15 of these was a conference abstract. The pelvic floor muscle training programs, and comparison interventions, varied markedly. Outcome measures differed between trials, and methods of data reporting varied, making the data difficult to combine. Many of the trials were small. Allocation concealment was adequate in five trials, and nine trials used assessors masked to group allocation. Thirteen trials reported that there were no losses to follow up, seven trials had dropout rates of less than 10%, but in the remaining trials the proportion of dropouts ranged from 12% to 41%. Pelvic floor muscle training was better than no treatment or placebo treatments for women with stress or mixed incontinence. 'Intensive' appeared to be better than 'standard' pelvic floor muscle training. PFMT may be more effective than some types of electrical stimulation but there were problems in combining the data from these trials. There is insufficient evidence to determine if pelvic floor muscle training is better or worse than other treatments. The effect of adding pelvic floor muscle training to other treatments (e.g. electrical stimulation, behavioural training) is not clear due to the limited amount of evidence available. Evidence of the effect of adding other adjunctive treatments to PFMT (e.g. vaginal cones, intravaginal resistance) is equally limited. The effectiveness of biofeedback assisted PFMT is not clear, but on the basis of the evidence available there did not appear to be any benefit over PFMT alone at post treatment assessment. Long-term outcomes of pelvic floor muscle training are unclear. Side effects of pelvic floor muscle training were uncommon and reversible. A number of the formal comparisons should be viewed with caution due to statistical heterogeneity, lack of statistical independence, and the possibility of spurious confidence intervals in some instances. REVIEWER'S CONCLUSIONS: Pelvic floor muscle training appeared to be an effective treatment for adult women with stress or mixed incontinence. Pelvic floor muscle training was better than no treatment or placebo treatments. The limitations of the evidence available mean that is difficult to judge if pelvic floor muscle training was better or worse than other treatments. Most trials to date have studied the effect of treatment in younger, premenopausal women. The role of pelvic floor muscle training for women with urge incontinence alone remains unclear. Many of the trials were small with poor reporting of allocation concealment and masking of outcome assessors. In addition there was a lack of consistency in the choice and reporting of outcome measures that made data difficult to combine. Methodological problems limit the confidence that can be placed in the findings of the review. Further, large, high quality trials are necessary. PMID- 11279717 TI - Interventions to improve the management of diabetes mellitus in primary care, outpatient and community settings. AB - BACKGROUND: Diabetes is a common chronic disease that is increasingly managed in primary care. Different systems have been proposed to manage diabetes care. OBJECTIVES: To assess the effects of different interventions, targeted at health professionals or the structure in which they deliver care, on the management of patients with diabetes in primary care, outpatient and community settings. SEARCH STRATEGY: We searched the Cochrane Effective Practice and Organisation of Care Group specialised register, the Cochrane Controlled Trials Register (Issue 4 1999), MEDLINE (1966-1999), EMBASE (1980-1999), Cinahl (1982-1999), and reference lists of articles. SELECTION CRITERIA: Randomised trials (RCTs), controlled clinical trials (CCTs), controlled before and after studies (CBAs) and interrupted time series (ITS) analyses of professional, financial and organisational strategies aimed at improving care for people with Type 1 or Type 2 diabetes. The participants were health care professionals, including physicians, nurses and pharmacists. The outcomes included objectively measured health professional performance or patient outcomes, and self-report measures with known validity and reliability. DATA COLLECTION AND ANALYSIS: Two reviewers independently extracted data and assessed study quality. MAIN RESULTS: Forty-one studies were included involving more than 200 practices and 48,000 patients. Twenty-seven studies were RCTs, 12 were CBAs, and two were ITS. The studies were heterogeneous in terms of interventions, participants, settings and outcomes. The methodological quality of the studies was often poor. In all studies the intervention strategy was multifaceted. In 12 studies the interventions were targeted at health professionals, in nine they were targeted at the organisation of care, and 20 studies targeted both. In 15 studies patient education was added to the professional and organisational interventions. A combination of professional interventions improved process outcomes. The effect on patient outcomes remained less clear as these were rarely assessed. Arrangements for follow-up (organisational intervention) also showed a favourable effect on process outcomes. Multiple interventions in which patient education was added or in which the role of the nurse was enhanced also reported favourable effects on patients' health outcomes. REVIEWER'S CONCLUSIONS: Multifaceted professional interventions can enhance the performance of health professionals in managing patients with diabetes. Organisational interventions that improve regular prompted recall and review of patients (central computerised tracking systems or nurses who regularly contact the patient) can also improve diabetes management. The addition of patient-oriented interventions can lead to improved patient health outcomes. Nurses can play an important role in patient-oriented interventions, through patient education or facilitating adherence to treatment. PMID- 11279718 TI - Ropinirole for levodopa-induced complications in Parkinson's disease. AB - BACKGROUND: Long-term levodopa therapy for Parkinson's disease is complicated by the development of motor fluctuations and abnormal involuntary movements. One approach is to add a dopamine agonist at this stage of the disease to reduce the time the patient spends immobile or off and to reduce the dose of levodopa in the hope of reducing such problems in the future. OBJECTIVES: To compare the efficacy and safety of adjuvant ropinirole therapy versus placebo in patients with Parkinson's disease already established on levodopa therapy and suffering from motor complications. SEARCH STRATEGY: Electronic searches of MEDLINE, EMBASE and the Cochrane Controlled Trials Register. Handsearching of the neurology literature as part of the Cochrane Movement Disorders Group's strategy. Examination of the reference lists of identified studies and other reviews. Contact with SmithKline Beecham. SELECTION CRITERIA: Randomised controlled trials of ropinirole versus placebo in patients with a clinical diagnosis of idiopathic Parkinson's disease and long-term complications of levodopa therapy. DATA COLLECTION AND ANALYSIS: Data was abstracted independently by the authors and differences settled by discussion. The outcome measures used included Parkinson's disease rating scales, levodopa dosage, 'off' time measurements and the frequency of withdrawals and adverse events. MAIN RESULTS: Three double-blind, parallel group, randomised, controlled trials have been conducted on 263 patients. The two phase II studies were relatively small, were conducted over the short term (12 weeks), and used relatively low doses of ropinirole (mean administered doses 3.3 and 3.5 mg/d) in a twice daily regime. In view of this clinical heterogeneity and some statistical heterogeneity, the results of these trials have not been included in a meta-analysis. The conclusions of this review are based on the evidence from a single phase III study which was medium term (26 weeks) and used ropinirole doses in line with the current UK licensed maximum in a thrice daily regime. In view of difficulties in assessing changes in off time in ~~ Leiberman 98~~, caused by the initial imbalance between the arms of the trial, it is unsafe to draw any firm conclusion about the effect of ropinirole on off time. However, as an adverse event, dyskinesia was significantly increased in those who received ropinirole (~~ Leiberman 98~~; odds ratio 2.90; 1.36, 6.19 95% CI; Table 8). Measurements of motor impairments and disability were poor in this study with incomplete information available. Levodopa dose could be reduced in ~~ Leiberman 98~~ with a significantly larger reduction on ropinirole than on placebo (weighted mean difference 180 mg/d; 106, 253 95% CI; Table 2). No significant differences in the frequency of adverse event reports were noted between ropinirole and placebo apart from the increase in dyskinesia with ropinirole. There was a trend towards fewer withdrawals from ropinirole in ~~ Leiberman 98~~ but this did not reach statistical significance. REVIEWER'S CONCLUSIONS: Ropinirole therapy can reduce levodopa dose but at the expense of increased dyskinetic adverse events. No clear effect on off time reduction was found but this may have been due to the under-powering of the single evaluable trial. Inadequate data on motor impairments and disability was collected to assess these outcomes. These conclusions apply to short and medium term treatment, up to 26 weeks. Further longer term trials are required, with measurements of effectiveness, and also studies to compare the newer with the older dopamine agonists. PMID- 11279719 TI - Ropinirole versus bromocriptine for levodopa-induced complications in Parkinson's disease. AB - BACKGROUND: Long-term levodopa therapy for Parkinson's disease is complicated by the development of motor fluctuations and abnormal involuntary movements. One approach is to add a dopamine agonist at this stage of the disease to reduce the time the patient spends immobile or off and to reduce the dose of levodopa in the hope of reducing such problems in the future. OBJECTIVES: To compare the efficacy and safety of adjuvant ropinirole therapy with bromocriptine in patients with Parkinson's disease already established on levodopa therapy and suffering from motor complications. SEARCH STRATEGY: Electronic searches of MEDLINE, EMBASE and the Cochrane Controlled Trials Register. Handsearching of the neurology literature as part of the Cochrane Movement Disorders Group's strategy. Examination of the reference lists of identified studies and other reviews. Contact with SmithKline Beecham. SELECTION CRITERIA: Randomised controlled trials of ropinirole versus bromocriptine in patients with a clinical diagnosis of idiopathic Parkinson's disease and long-term complications of levodopa therapy. DATA COLLECTION AND ANALYSIS: Data was abstracted independently by the authors and differences settled by discussion. The outcome measures used included Parkinson's disease rating scales, levodopa dosage, 'off' time measurements and the frequency of withdrawals and adverse events. MAIN RESULTS: In the 3 trials identified, no significant differences between ropinirole and bromocriptine were found in off time reduction, dyskinesia as an adverse event, motor impairment and disability, or levodopa dose reduction. Withdrawal rates and adverse event frequency were similar with the two agents apart from significantly less nausea with ropinirole (odds ratio 0.50; 0.29, 0.84 95% CI; p =0.01). REVIEWER'S CONCLUSIONS: Ropinirole is at least as good as bromocriptine in patients with Parkinson's disease with motor complications in terms of improving off time and reducing levodopa dose, without increasing adverse events including dyskinesia. However, these comparitor studies may have been underpowered to detect clinically meaningful differences between the agonists. Further, much larger, phase IV studies are required to examine the efficacy, effectiveness, and safety of all of the dopamine agonists as adjuvant therapy in Parkinson's disease. PMID- 11279720 TI - Cabergoline for levodopa-induced complications in Parkinson's disease. AB - BACKGROUND: Long term levodopa therapy in Parkinson's disease is associated with the development of motor complications including abnormal involuntary movements and a shortening response to each dose (wearing off phenomenon). It is thought that dopamine agonists can reduce the duration of immobile off periods and the need for levodopa therapy whilst maintaining or improving motor impairments and only minimally increasing dopaminergic adverse events. OBJECTIVES: To compare the efficacy and safety of adjuvant cabergoline therapy versus placebo in patients with Parkinson's disease, already established on levodopa and suffering from motor complications. SEARCH STRATEGY: Electronic searches of MEDLINE, EMBASE and the Cochrane Controlled Trials Register. Handsearching of the neurology literature as part of the Cochrane Movement Disorders Group's strategy. Examination of the reference lists of identified studies and other reviews. Contact with Pharmacia Upjohn Limited. SELECTION CRITERIA: Randomised controlled trials of cabergoline versus placebo in patients with a clinical diagnosis of idiopathic Parkinson's disease and long-term complications of levodopa therapy. DATA COLLECTION AND ANALYSIS: Data was abstracted independently by the authors and differences settled by discussion. The outcome measures used included Parkinson's disease rating scales, levodopa dosage, off time measurements and the frequency of withdrawals and adverse events. MAIN RESULTS: Cabergoline has been compared with placebo in two phase II (6 - 12 weeks) and one phase III randomised controlled trials (24 weeks). These were double-blind, parallel group, multicentre studies including 268 patients with Parkinson's disease and motor complications. The reduction of 1.14 hours (WMD; 95% CI -0.06, 2.33; p = 0.06) in off time in favour of cabergoline was not statistically significant. Inadequate data on dyskinesia was collected either on rating scales or as adverse event reporting to allow a conclusion to be drawn. A small but statistically significant advantage of cabergoline over placebo was seen in one study for UPDRS ADL (part II) score and UPDRS motor score. No such advantage was seen in one other study due to small numbers of patients and the comparatively low doses of cabergoline used. No significant differences in Schwab and England scale were seen in two studies. Levodopa dose reduction was significantly greater with cabergoline (WMD 149.6 mg/d; 95% CI 94.1, 205.1; p < 0.00001). There was a trend towards more dopaminergic adverse events with cabergoline but this did not reach statistical significance at the p < 0.01 level. However, there was a trend towards fewer withdrawals from cabergoline. REVIEWER'S CONCLUSIONS: In the management of the motor complications seen in Parkinson's disease, cabergoline can be used to reduce levodopa dose and modestly improve motor impairment and disability with an acceptable adverse event profile. These conclusions are based on, at best, medium term evidence. PMID- 11279721 TI - Cabergoline versus bromocriptine for levodopa-induced complications in Parkinson's disease. AB - BACKGROUND: Long term levodopa therapy in Parkinson's disease is associated with the development of motor complications including abnormal involuntary movements and a shortening response to each dose (wearing off phenomenon). It is thought that dopamine agonists can reduce the duration of immobile off periods and the need for levodopa therapy whilst maintaining or improving motor impairments and only minimally increasing dopaminergic adverse events. OBJECTIVES: To compare the efficacy and safety of adjuvant cabergoline therapy versus bromocriptine in patients with Parkinson's disease, already established on levodopa and suffering from motor complications. SEARCH STRATEGY: Electronic searches of MEDLINE, EMBASE and the Cochrane Controlled Trials Register. Handsearching of the neurology literature as part of the Cochrane Movement Disorders Group's strategy. Examination of the reference lists of identified studies and other reviews. Contact with Pharmacia Upjohn Limited. SELECTION CRITERIA: Randomised controlled trials of cabergoline versus bromocriptine in patients with a clinical diagnosis of idiopathic Parkinson's disease and long-term complications of levodopa therapy. DATA COLLECTION AND ANALYSIS: Data were abstracted independently by the authors and differences settled by discussion. The outcome measures used included Parkinson's disease rating scales, levodopa dosage, off time measurements and the frequency of withdrawals and adverse events. MAIN RESULTS: Cabergoline has been compared with bromocriptine in five randomised, double-blind, parallel group studies including 1071 patients. Only one of the phase II studies was medium term (36 weeks), the others all being short term (12 -15 weeks). The non-significant difference in off time reduction produced by cabergoline compared with bromocriptine was 0.29 hours/day in favour of the former (weighted mean difference; 95% CI -0.10, 0.68; p = 0.15). Dyskinesia reported as an adverse event was significantly increased with cabergoline compared with bromocriptine (Peto odds ratio 1.57; 95% CI 1.05, 2.35; p = 0.03). Motor impairment and disability were measured in four of the studies using the UPDRS rating scale but the small differences in UPDRS ADL (part II) and motor (part III) scores were not statistically significant in any study. Similarly, no significant difference in Schwab and England score was seen. The number of patients rated as much or very much improved on a clinician's global impression scale was similar with both agonists. Levodopa dose reduction was no different between cabergoline and bromocriptine. There was more confusion with cabergoline (Peto odds ratio 2.02; 95% CI 1.09, 3.76; p = 0.03). Otherwise, dopaminergic adverse events were comparable with these agonists and no significant difference in all cause withdrawal rate was found. REVIEWER'S CONCLUSIONS: Cabergoline produces similar benefits to bromocriptine in off time reduction, motor impairment and disability ratings, and levodopa dose reduction over the first three months of therapy. Dyskinesia and confusion were increased with cabergoline but otherwise the frequency of adverse events and withdrawals from treatment were similar with the two agonists. PMID- 11279722 TI - Melatonin for preventing and treating jet lag. AB - BACKGROUND: Jet-lag commonly affects air travellers who cross several time zones. It results from the body's internal rhythms being out of step with the day-night cycle at the destination. Melatonin is a pineal hormone that plays a central part in regulating bodily rhythms and has been used as a drug to re-align them with the outside world. OBJECTIVES: To assess the effectiveness of oral melatonin taken in different dosage regimens for alleviating jet-lag after air travel across several time zones. SEARCH STRATEGY: We searched the Cochrane Controlled Trials Register, MEDLINE, EMBASE, PsychLit and Science Citation Index electronically, and the journals 'Aviation, Space and Environmental Medicine' and 'Sleep' by hand. We searched citation lists of relevant studies for other relevant trials. We asked principal authors of relevant studies to tell us about unpublished trials. Reports of adverse events linked to melatonin use outside randomised trials were searched for systematically in 'Side Effects of Drugs' (SED) and SED Annuals, 'Reactions Weekly', MEDLINE, and the adverse drug reactions databases of the WHO Uppsala Monitoring Centre (UMC) and the US Food & Drug Administration. SELECTION CRITERIA: Randomised trials in airline passengers, airline staff or military personnel given oral melatonin, compared with placebo or other medication. Outcome measures should consist of subjective rating of jet lag or related components, such as subjective wellbeing, daytime tiredness, onset and quality of sleep, psychological functioning, duration of return to normal, or indicators of circadian rhythms. DATA COLLECTION AND ANALYSIS: : Ten trials met the inclusion criteria. All compared melatonin with placebo; one in addition compared it with a hypnotic, zolpidem. Nine of the trials were of adequate quality to contribute to the assessment, one had a design fault and could not be used in the assessment. Reports of adverse events outside trials were found through MEDLINE, 'Reactions Weekly', and in the WHO UMC database. MAIN RESULTS: : Nine of the ten trials found that melatonin, taken close to the target bedtime at the destination (10pm to midnight), decreased jet-lag from flights crossing five or more time zones. Daily doses of melatonin between 0.5 and 5mg are similarly effective, except that people fall asleep faster and sleep better after 5mg than 0.5mg. Doses above 5mg appear to be no more effective. The relative ineffectiveness of 2mg slow-release melatonin suggests that a short-lived higher peak concentration of melatonin works better. Based on the review, the number needed to treat (NNT) is 2. The benefit is likely to be greater the more time zones are crossed, and less for westward flights. The timing of the melatonin dose is important: if it is taken at the wrong time, early in the day, it is liable to cause sleepiness and delay adaptation to local time. The incidence of other side effects is low. Case reports suggest that people with epilepsy, and patients taking warfarin may come to harm from melatonin. REVIEWER'S CONCLUSIONS: Melatonin is remarkably effective in preventing or reducing jet-lag, and occasional short-term use appears to be safe. It should be recommended to adult travellers flying across five or more time zones, particularly in an easterly direction, and especially if they have experienced jet-lag on previous journeys. Travellers crossing 2-4 time zones can also use it if need be. The pharmacology and toxicology of melatonin needs systematic study, and routine pharmaceutical quality control of melatonin products must be established. The effects of melatonin in people with epilepsy, and a possible interaction with warfarin, need investigation. PMID- 11279723 TI - Preoperative chemotherapy for resectable thoracic esophageal cancer. AB - BACKGROUND: Carcinoma of the esophagus is a relatively uncommon but lethal cancer that continues to kill over 90% of its victims within 5 years. Surgery is the treatment of choice for most localized esophageal cancer patients. However, despite curative resection, the 5-year survival rate ranges from 15% to 39%. The failure of surgery to cure clinically localized esophageal cancer is because of the advanced state of the disease before symptoms occur, high frequency of lymph node involvement, and the common occurrence of submucosal spread and extension to surrounding structures. Preoperative chemotherapy has been used in an attempt to decrease tumour activity, increase resectability, and improve disease-free and overall survival. A number of studies have investigated whether preoperative chemotherapy followed by surgery leads to an improvement in cure rates, but the individual reports have not been encouraging. The role of preoperative chemotherapy in the treatment of resectable thoracic esophageal cancer remains undefined. OBJECTIVES: The objective of this review is to determine the role of preoperative chemotherapy on overall survival and/or quality-of-life for patients with resectable thoracic esophageal carcinoma. SEARCH STRATEGY: Trials were identified by searching the Cochrane Controlled Trials Register (Issue 2 - 2000), MEDLINE (1966 - 2000), EMBASE (1988 - 2000) and CancerLit (1993 - 2000). The references of all identified studies, review articles, and standard textbooks were examined. Members of the Cochrane UGPD Group and experts in the oncology field were contacted and asked to supply details of any outstanding clinical trials and relevant unpublished materials. There were no language restrictions. The searches were updated in June 2000. The clinical trial registers of the National Cancer Institute and the Radiation Therapy Oncology Group were consulted for ongoing trials. SELECTION CRITERIA: Types of studies Studies (published or unpublished) that randomised patients with potentially resectable carcinoma of the esophagus (of any histologic type) to chemotherapy or no chemotherapy before surgery were included in this review. Studies were excluded if they were not truly randomised (phase I or II trials), earlier versions of updated trials, if other treatment modalities (e.g. radiotherapy, hyperthermia) were used, or if there was not a surgery alone control arm. Types of participants The participants consisted of patients with potentially resectable thoracic esophageal carcinoma (of any histologic type). Trials involving patients with carcinoma of the cervical esophagus were excluded. Types of interventions Trials that compared chemotherapy before surgery (esophagectomy) with surgical resection alone (esophagectomy). Types of outcome measures The primary outcome was death at yearly intervals. Morbidity (complications), and quality-of-life were secondary outcomes. DATA COLLECTION AND ANALYSIS: Overall mortality at yearly intervals was determined by extracting the total number of patients randomised to the treatment and control groups and the number of deaths in each group. All analyses were carried out on intention-to-treat that is patients were analyzed according to their allocated treatment, irrespective of whether they received that treatment. Mortality at 1, 2, 3, 4 and 5 years were used as endpoints of clinical relevance. If survival numbers at the specified time intervals were not given, they were estimated from the published survival curves. The number of deaths in the treatment groups (preoperative chemotherapy plus surgery) was compared to the number of deaths in the control groups (surgery alone). Treatment modalities as well as patient demographics and characteristics and side-effects were also recorded. Trials meeting the inclusion criteria were evaluated by two independent reviewers using the Jadad method MAIN RESULTS: A total of 14 randomised controlled trials and 1 meta-analysis of preoperative chemotherapy versus surgery alone for esophageal carcinoma were identified to be potentially eligible for review. This review is based on 7 randomised trials and 1653 patients. At 1 year the Peto odds ratio based on the fixed-effects models showed no difference in mortality between preoperative chemotherapy and surgery alone (OR = 1.03). At 2 years there was a 20% significant decrease in mortality for preoperative chemotherapy (OR = 0.80; 95% C.I. 0.65 to 0.99) but the results were not robust. The results at 3, 4, and 5 years found odds ratios tending to favour preoperative chemotherapy, but wide confidence intervals that included 1. None of the published trials reported on quality-of-life outcomes. There appeared to be an increased risk of morbidity with chemotherapy. REVIEWER'S CONCLUSIONS: The results of this review suggest that there is no strong evidence to recommend preoperative chemotherapy in the treatment of surgically resectable carcinomas of the thoracic esophagus. (ABSTRACT TRUNCATED) PMID- 11279724 TI - Postnatal phenobarbitone for the prevention of intraventricular hemorrhage in preterm infants. AB - BACKGROUND: Intraventricular hemorrhage (IVH) is a major complication of preterm birth. Large hemorrhages are associated with a high risk of disability and hydrocephalus. Instability of blood pressure and cerebral blood flow are postulated as causative factors. Another mechanism may involve reperfusion damage from oxygen free radicals. Phenobarbitone has been suggested as a safe treatment which stabilises blood pressure and may protect against free radicals. OBJECTIVES: To determine whether postnatal administration of phenobarbitone to preterm infants reduces the risk of intraventricular hemorrhage (IVH), neurodevelopmental impairment or death. SEARCH STRATEGY: See the Search Strategy of the Neonatal Collaborative Review Group. The reviewer has been a active trialist in this area and has personal contact with many groups in this field. Journals handsearched from 1976 (when cranial CT scanning started) to October 2000 include: Pediatrics, J Pediatrics, Archives of Disease in Childhood, Pediatric Research, Developmental Medicine and Child Neurology, Acta Paediatrica, European J of Pediatrics, Neuropediatrics, New England J of Medicine, Lancet and British Medical J. The National Library of Medicine (USA) database (via PubMed) and the Cochrane Controlled Trials Register were searched through to October 2000 using the MeSH terms intraventricular hemorrhage, newborn infants, premature infant, intracranial hemorrhage, phenobarbitone, phenobarbital. The searches were not limited to the English language, as long as the article included an English abstract. Promising articles were read in the original language or translated. SELECTION CRITERIA: Included were randomized or quasi-randomized controlled trials in which phenobarbitone was given to preterm infants identified as being at risk of IVH because of gestational age below 34 weeks, birthweight below 1500 g, or respiratory failure. Adequate determination of IVH by ultrasound or CT was also required. DATA COLLECTION AND ANALYSIS: In addition to details of patient selection and control of bias, the details of the administration of phenobarbitone were extracted. The end-points searched for included: IVH ( with grading), posthemorrhagic ventricular dilatation or hydrocephalus, neurodevelopmental impairment and death. In addition, possible adverse effects of phenobarbitone such as hypotension, mechanical ventilation, pneumothorax, hypercapnia, and acidosis were searched for. MAIN RESULTS: Nine controlled trials were included with 740 infants recruited. There was heterogeneity between trials for the outcome IVH, with one trial finding a significant decrease in IVH and another trial finding an increase in IVH in the group receiving phenobarbitone. Meta-analysis showed no difference between the phenobarbitone treated group and the control group in either IVH (typical relative risk 1.04, CI 0.87, 1.25), severe IVH (typical relative risk 0.91, CI 0.66, 1.27), posthemorrhagic ventricular dilatation (typical relative risk 0.89, CI 0.38, 2.08), severe neurodevelopmental impairment (typical relative risk 1.44, CI 0.41, 5.04) or death before hospital discharge (typical relative risk 0.88, CI 0.64, 1.21) There was a consistent trend in the trials towards increased use of mechanical ventilation in the phenobarbitone treated group, which was supported by the meta analysis (typical relative risk 1.18, CI 1.06, 1.32; typical risk difference 0.129, CI 0.045, 0.213), but there was no significant difference in pneumothorax, acidosis or hypercapnia. REVIEWER'S CONCLUSIONS: Postnatal administration of phenobarbitone cannot be recommended as prophylaxis to prevent IVH in preterm infants and is associated with an increased need for mechanical ventilation. PMID- 11279725 TI - Beta2-agonists for acute bronchitis. AB - BACKGROUND: The optimal treatment for acute bronchitis is not clear. Because many patients with acute bronchitis have airflow limitation as well as cough, beta2 agonists may be useful. OBJECTIVES: To determine whether beta2-agonists improve the symptoms of acute bronchitis in patients who do not have underlying pulmonary disease. SEARCH STRATEGY: Cochrane Library, MEDLINE, EMBASE, and Conference Proceedings using "bronchodilator (exp)", "adrenergic beta-agonist (exp)", or "sympathomimetics (exp)" and "bronchitis" or "cough"; Science Citation Index for referenced publications; and letters to manufacturers of beta2-agonists. SELECTION CRITERIA: Trials in which patients (adults or children age > 2 years) without known pulmonary disease who were diagnosed with acute bronchitis or acute cough without other cause were randomized to beta2-agonist vs. placebo, no treatment, or alternative treatment. DATA COLLECTION AND ANALYSIS: Three reviewers independently first selected outcomes and evaluated trial quality while blinded to study results, and then extracted data. Trials in children and in adults were analyzed separately. MAIN RESULTS: Two trials in children with acute cough and no evidence of airway obstruction did not find any benefits from beta2 agonists. Five trials in adults with acute cough or acute bronchitis had mixed results, but overall summary statistics did not reveal any significant benefits from beta2-agonists. Subgroups of patients with evidence of airflow limitation had lower symptom scores if given beta2-agonists in one trial; and the trials that did note quicker resolution of cough in patients given beta2-agonists were those that had a higher proportion of patients with wheezing at baseline. Patients given beta2-agonists (whether oral or inhaled) were more likely to report tremor, shakiness, or nervousness than patients in the control groups (NNH for children 9, 95% CI 5 to 100; NNH for adults 2.3, 95% CI 2 to 3). REVIEWER'S CONCLUSIONS: There is no evidence to support using beta2-agonists in children with acute cough who do not have evidence of airflow obstruction. There is also little evidence that the routine use of beta2-agonists for adults with acute cough is helpful. These agents may reduce symptoms, including cough, in patients with evidence of airflow obstruction; but this potential benefit is not well supported by the available data and must be weighed against the adverse effects associated with beta2-agonists. PMID- 11279726 TI - Corticosteroids for acute severe asthma in hospitalised patients. AB - BACKGROUND: Corticosteroids are currently used routinely in the management of acute severe asthma. The optimal dose and route of administration continues to be debated. Some investigators have reported a greater benefit of higher doses of corticosteroids in the management of severe asthma, while others have not. OBJECTIVES: To determine whether higher doses of systemic corticosteroids (oral, intravenous or intramuscular) are more effective than lower doses in the management of patients with acute severe asthma requiring hospital admission. SEARCH STRATEGY: Randomised controlled trials were identified from the Cochrane Airways Group Asthma Register. In addition, primary authors and content experts were contacted to identify eligible studies. Bibliographies from included studies, known reviews and texts were also searched. SELECTION CRITERIA: Studies were selected for inclusion in the review if they met the following broad inclusion criteria: described as randomised controlled trials, included patients with acute severe asthma, compared different doses of corticosteroids (any route) in 2 or more treatment arms, and had a minimum period of follow up of 24 hours. Two reviewers independently assessed the studies for inclusion and disagreement was resolved by third party adjudication. DATA COLLECTION AND ANALYSIS: Data were extracted independently by two reviewers if the authors were unable to verify the validity of information. Missing data were obtained from authors or calculated from other data presented in the paper. The data were analysed as weighted mean differences (WMD) for primary pulmonary function outcomes using a fixed effects model. For the purposes of the review, three broad categories of corticosteroid dose (equivalent dose of methylprednisolone in 24 hours) were defined in advance: low dose (< or = 80 mg), medium dose (> 80 mg and < or = 360 mg) and high dose (> 360 mg). There were thus 3 main comparison groups: low versus medium dose, medium versus high dose and low versus high dose. MAIN RESULTS: Nine trials were included; a total of 344 adult patients have been studied (96 with low dose, 85 with medium dose and 163 with high dose corticosteroids). Only 6 trials provided sufficient data for the meta-analysis. There were no clinically or statistically significant differences detected in % predicted FEV1 among comparison groups after 24, 48 or 72 hours. At 48 hours, the weighted mean difference was -3.3% predicted (95% confidence interval -12.4 to + 5.8) for the low vs medium dose comparison, -1.9% predicted (95% CI -8.1 to + 4.3) for the medium vs high dose comparison and + 0.5% predicted (95% CI - 7.8 to + 8.8) for the low vs high dose comparison. There appeared to be no significant differences in side effects or rates of respiratory failure among the varying doses of corticosteroids. A further search was conducted on 3rd August 2000. No new trials were identified. REVIEWER'S CONCLUSIONS: No differences were identified among the different doses of corticosteroids in acute asthma requiring hospital admission. Low dose corticosteroids (< or = 80 mg/day of methylprednisolone or < or = 400 mg/day of hydrocortisone) appear to be adequate in the initial management of these adult patients. Higher doses do not appear to offer a therapeutic advantage. PMID- 11279727 TI - Galantamine for Alzheimer's disease. AB - BACKGROUND: Galantamine (also called galanthamine, marketed as Reminyl (Janssen)) can be isolated from several plants, including daffodil bulbs, and now synthesized. Galantamine is a specific, competitive, and reversible acetylcholinesterase inhibitor. It is also an allosteric modulator at nicotinic cholinergic receptor sites potentiating cholinergic nicotinic neurotransmission. A small number of early studies showed mild cognitive and global benefits for patients with Alzheimer's disease, and recently several multicentre clinical trials have been published with positive findings. Galantamine has received regulatory approval in Sweden, is available in Austria, and awaits marketing approval in the United States, Europe, and other countries. OBJECTIVES: The objective of this review is to assess the clinical effects of galantamine in patients with probable Alzheimer's disease, and to investigate potential moderators of an effect. SEARCH STRATEGY: The Cochrane Dementia Group specialized register of clinical trials was searched using the terms 'galantamine,' and 'galanthamine' (15 February 2000) as was the Cochrane Controlled Trials Register (2000, Issue 2). These terms were also used to search the following databases: EMBASE, MEDLINE, PsychLit; Combined Health Information Database, NRR (National Research Register), ADEAR (Alzheimer's Disease Education and Referral Centre clinical database, BIOMED (Biomedicine and Health), Glaxo-Wellcome Clinical Trials Register, National Institutes of Health Clinical Trials Databases, Current Controlled Trials, Dissertation Abstracts (mainly North American dissertations) 1961-1994, Index to UK Theses (British dissertations) 1970-1994. Published reviews were inspected for further sources. Additional information was collected from an unpublished investigational brochure for galantamine. SELECTION CRITERIA: Trials selected were randomized, double-blind, parallel-group, and unconfounded comparisons of galantamine with placebo for a treatment duration of greater than 4 weeks in people with Alzheimer's disease. DATA COLLECTION AND ANALYSIS: Data were extracted independently by the reviewers and pooled where appropriate and possible. The pooled odds ratios (95%CI) or the average differences (95%CI) were estimated. Intention-to-treat and observed cases data were both reported, if the data were available to be reported. -Outcomes of interest include the Alzheimer's Disease Assessment Scale-cognitive subscale (ADAS-cog), clinical global impression of change (CIBIC-plus or CGIC), Alzheimer's Disease Cooperative Study/Activities of Daily Living (ADCS-ADL), Disability Assessment for Dementia scale (DAD) and Neuropsychiatric Inventory (NPI). - Potential moderating variables of a treatment effect included trial duration and dose. MAIN RESULTS: Seven trials were identified that met criteria for entry, with 6 being Phase II or III industry-sponsored multicentre trials. One was of 12 weeks duration; one of 5 months; one of 29 weeks; and the rest of 6 months duration. Trials of 5 months or more were aggregated in the analyses as '6 months'. Overall, galantamine showed significant treatment effects at daily doses of 16-32 mg/d for trials of 3- to 6-months duration. For global ratings, trials of 3 months duration with doses of 24-32mg/d (Odds Ratio (OR) 2.2; 95%CI 1.4 to 3.7) and 36mg/d (OR 3.3; 95%CI 1.2 to 9.3) were statistically significant in favour of treatment. For trials of 6 months duration (5-months to 29 weeks), only doses of 8mg/d failed to be statistically significant (24mg: OR 2.0; 95%CI 1.5 to2.5; 32mg: OR 1.9; 95%CI 1.4 to 2.5). For cognitive function over 6 months duration: at a 24mg/d, improvements measured -3.5 points (k=3; 95%CI -4.3 to -2.8) on weighted mean difference on the ADAS-Cog scale, and -4.0 points at 32mg/d (k=2; 95%CI -5.0 to -3.0). Both observed cases (WMD 3.8; 95%CI 0.3 to 7.3) and intention to treat analyses using the Disability Assessment of Dementia gave statistically significant results in favour of treatment for daily doses of 32mg for 6 months duration. The small number of trials available for analysis, however, limited the power of analyses to detect differences. Galantamine consistently failed to show statistically significant treatment effects at doses of 8mg/day. Galantamine's adverse effects appear similar to those of other cholinesterase inhibitors, in that it tends to produce gastrointestinal effects acutely and with dosage increases. Overall, people treated with galantamine at doses of 24-32 mg/d were more likely to discontinue participation in most trials than were people treated with lower doses or placebo, but in the one trial with a slower rate of titration the discontinuation rate was not significantly greater than placebo for the 16 mg/day dose. (ABSTRACT TRUNCATED) PMID- 11279728 TI - Anterior vaginal repair for urinary incontinence in women. AB - BACKGROUND: Anterior vaginal repair (anterior colporrhaphy) is an operation traditionally used for moderate or severe stress urinary incontinence in women. About a third of adult women experience urinary incontinence. OBJECTIVES: To determine the effects of anterior vaginal repair (anterior colporrhaphy) on stress or mixed urinary incontinence in comparison with other management options. SEARCH STRATEGY: We searched the Cochrane Incontinence Group's trials register, and the reference lists of relevant articles. Date of most recent search: September 2000. SELECTION CRITERIA: Randomised or quasi-randomised trials that included anterior vaginal repair for the treatment of urinary incontinence. DATA COLLECTION AND ANALYSIS: Both reviewers independently extracted data and assessed trial quality. Two trial investigators were contacted for additional information. MAIN RESULTS: Nine trials were identified which included 333 women having an anterior vaginal repair and 599 who received comparison interventions. A single small trial provided insufficient evidence to assess anterior vaginal repair in comparison with physical therapy. The performance of anterior repair in comparison with needle suspension appeared similar but clinically important differences could not be confidently ruled out. No trials compared anterior repair with suburethral sling operations or laparoscopic retropubic suspensions, or compared alternative vaginal operations. Anterior vaginal repair was less effective than open abdominal retropubic suspension based on patient-reported cure rates in eight trials both in the short-term (failure rate within first year after anterior repair 82/279, 29% vs 50/346, 14%; RR 1.89, 95% CI 1.39 to 2.59) and long-term (after first year, 132/322, 41% vs 68/395, 17%; RR 2.50, 95% CI 1.92 to 3.26). There was evidence from three of these trials that this was reflected in more repeat operations for incontinence (25/107, 23% vs 4/164, 2%; RR 8.87, 95% CI 3.28 to 23.94). These findings held irrespective of the co existence of prolapse (pelvic relaxation). Later prolapse operation appeared to be equally common after vaginal (3%) or abdominal (4%) operation. In respect of the type of open abdominal retropubic suspension, most data related to comparisons of anterior vaginal repair with Burch colposuspension. The few data describing comparison of anterior repair with the Marshall-Marchetti-Krantz procedure were consistent with those for Burch colposuspension. REVIEWER'S CONCLUSIONS: There were not enough data to allow comparison of anterior vaginal repair with physical therapy or needle suspension for primary urinary stress incontinence in women. Open abdominal retropubic suspension appeared to be better than anterior vaginal repair judged on subjective cure rates in six trials, even in women who had prolapse in addition to stress incontinence (four trials). The need for repeat incontinence surgery was also less after the abdominal operation. However, there was not enough information about post-operative complications and morbidity. PMID- 11279729 TI - Aerobic exercise interventions for people with HIV/AIDS. AB - BACKGROUND: The profile of HIV infection is constantly changing. Although once viewed as an illness progressing to death, HIV infection now presents as a chronic infection characterized by unpredictable cycles of wellness and illness. Thus, the needs of this population have increasingly included management of impairments, disabilities and handicaps. Exercise is a key management strategy employed by rehabilitation professionals to address these issues. Exercise has been shown to improve strength, cardiovascular function, and psychological status in seronegative populations (see Eds., Bouchard, C., Shephard, R.J., & Stephens, T. (1993). Physical Activity, Fitness, and Health. Champaign, IL: Human Kinetics Publishers.) But what are the effects of exercise for people living with HIV infection? If the risks and benefits of exercise for people living with HIV infection are better understood, appropriate exercise prescription may be practiced by health care providers. Improved exercise prescription may enhance the effectiveness of HIV management, thus improving overall outcomes for people living with HIV infection. OBJECTIVES: To examine the effect of aerobic exercise interventions on cardiopulmonary, immunologic and psychological parameters in adults living with HIV infection. SEARCH STRATEGY: Studies were identified using MEDLINE, EMBASE, SCIENCE CITATION INDEX, AIDSLINE CINAHL, HEALTHSTAR, PSYCHLIT, SOCIOFILE, SCI, SSCI, ERIC, DIA and abstracts from international AIDS meetings, ICAAC, and other major meetings. Reference lists from pertinent articles and books and personal contact with authors were also used, as were Collaborative Review Group databases and results of hand searching of targeted journals. All languages were included. Searches covered the period from 1980 to July 1999. SELECTION CRITERIA: For inclusion, studies had to be randomized control trials involving HIV+ adults 18 years of age or older and had to include at least one group randomized to receive aerobic exercise performed at least three times/week for at least four weeks. DATA COLLECTION AND ANALYSIS: Data on study design, participants, interventions, and outcomes were extracted from the reports onto specifically designed data collection forms by at least two reviewers. MAIN RESULTS: Six studies satisfied the eligibility criteria. The main results indicated that performing constant or interval aerobic exercise for at least 20 minutes, at least three times per week for four weeks may lead to increased CD4 count, improved cardiopulmonary fitness, and improved psychological status. These findings are limited to those who continued to exercise and for whom there was adequate follow-up data. REVIEWER'S CONCLUSIONS: Aerobic exercise appears to be safe and may be beneficial for adults living with HIV/AIDS. These findings are limited by the small sample sizes and large drop-out rates of the included studies. Further research is required to determine the optimal parameters of aerobic exercise and stage of disease in which aerobic exercise may be most beneficial for adults living with HIV infection. PMID- 11279730 TI - Exercise-based rehabilitation for coronary heart disease. AB - BACKGROUND: The burden of cardiovascular disease world-wide is one of great concern to patients and health care agencies alike. Cardiac rehabilitation aims to restore patients with heart disease to health through exercise only based rehabilitation or comprehensive cardiac rehabilitation. OBJECTIVES: To determine the effectiveness of exercise only or exercise as part of a comprehensive cardiac rehabilitation programme on the mortality, morbidity, health-related quality of life (HRQoL) and modifiable cardiac risk factors of patients with coronary heart disease. SEARCH STRATEGY: Electronic databases were searched for randomised controlled trials, using standardised trial filters, from the earliest date available to December 31st 1998. SELECTION CRITERIA: Men and women of all ages, in hospital or community settings, who have had myocardial infarction, coronary artery bypass graft or percutaneous transluminal coronary angioplasty, or who have angina pectoris or coronary artery disease defined by angiography. DATA COLLECTION AND ANALYSIS: Studies were selected independently by two reviewers, and data extracted independently. Authors were contacted where possible to obtain missing information. MAIN RESULTS: This systematic review has allowed analysis of an increased number of patients from approximately 4500 in earlier meta-analyses to 8440 (7683 contributing to the total mortality outcome). The pooled effect estimate for total mortality for the exercise only intervention shows a 27% reduction in all cause mortality (random effects model OR 0.73 (0.54, 0.98)). Comprehensive cardiac rehabilitation reduced all cause mortality, but to a lesser degree (OR 0.87 (0.71, 1.05)). Total cardiac mortality was reduced by 31% (random effects model OR 0.69 (0.51, 0.94)) and 26% (random effects model OR 0.74 (0.57, 0.96)) in the exercise only and comprehensive cardiac rehabilitation groups respectively. Neither intervention had any effect on the occurrence of non-fatal myocardial infarction. There was a significant net reduction in total cholesterol (pooled WMD random effects model -0.57 mmol/l (-0.83, -0.31)) and LDL (pooled WMD random effects model -0.51 mmol/l (-0.82, -0.19) in the comprehensive cardiac rehabilitation group. REVIEWER'S CONCLUSIONS: Exercise-based cardiac rehabilitation is effective in reducing cardiac deaths. It is not clear from this review whether exercise only or a comprehensive cardiac rehabilitation intervention is more beneficial. The population studied in this review is still predominantly male, middle aged and low risk. Identification of the ethnic origin of the participants was seldom reported. It is possible that patients who would have benefited most from the intervention were excluded from the trials on the grounds of age, sex or co-morbidity. PMID- 11279731 TI - Continuous nasogastric milk feeding versus intermittent bolus milk feeding for premature infants less than 1500 grams. AB - BACKGROUND: Most premature infants less than 1500 grams birth weight must be fed initially by tube because of their inability to suck effectively, or to coordinate sucking, swallowing and breathing. Milk feedings can be given by tube either intermittently, typically over 10-20 minutes every two or three hours, or continuously, using an infusion pump. Although theoretical benefits and risks of each method have been proposed, effects on clinically important outcomes remain uncertain. OBJECTIVES: To examine the evidence from randomized trials regarding the effectiveness of continuous versus intermittent bolus nasogastric milk feeding in premature infants less than 1500 grams. The primary outcomes reviewed included feeding tolerance, days to reach full enteral feeding, somatic growth, days to discharge and incidence of necrotizing enterocolitis (NEC). SEARCH STRATEGY: Searches were performed of MEDLINE, CINAHL, HealthSTAR, and the Cochrane Controlled Trials Register. As well, studies identified from abstracts and conference proceedings and references from relevant publications were retrieved. SELECTION CRITERIA: Randomized and quasi-randomized clinical trials that met the following criteria for relevance: a) Enrollment of infants < 1500 grams birth weight with no major congenital anomalies which might interfere with feeding tolerance b) Comparison of continuous nasogastric versus intermittent bolus tube feedings using breastmilk or formula c) Assessment of relevant outcomes including feeding tolerance, days to full feeds, somatic growth, days to discharge, and complications such as NEC or apnea DATA COLLECTION AND ANALYSIS: All articles retrieved from the complete search were assessed independently by the two reviewers for relevance (see selection criteria), and for methodologic quality using the following criteria: blinding of randomization, blinding of intervention, complete follow-up and blinding of outcome measurement. Only those articles judged by both reviewers to be relevant and to have appropriate methodologic quality were included in the analysis. Differences were resolved through discussion and consensus of the reviewers. MAIN RESULTS: Infants fed by continuous tube feeding method took longer to reach full enteral feeds (weighted mean difference 3.0 days; 95% CI 0.7, 5.2). Although there was no evidence of a difference in the days to discharge overall, one study suggested a trend toward earlier discharge for infants less than 1000 grams birth weight fed by the continuous tube feeding method (mean difference (MD) -11days; 95% CI -21.8, 0.2). Overall, there was no evidence of a difference in somatic growth (weight, length, head circumference or skinfold thickness) between the two groups, but subgroup analyses in one study suggested that infants less than 1000 grams and 1000 - 1250 grams birthweight gained weight faster when fed by the continuous tube feeding method (MD 2.0 g/day; 95% CI 0.5, 3.5; MD 2.0 g/day; 95% CI 0.2, 3.8, respectively). There was no evidence of a difference in the incidence of NEC. One study showed a trend toward more apneas during the study period in infants fed by the continuous tube feeding method (MD 14.0 apneas during study period; 95% CI -0.2, 28.2). REVIEWER'S CONCLUSIONS: Infants fed by the continuous tube feeding method took longer to reach full feeds, but there was no difference in somatic growth, days to discharge, or the incidence of NEC for infants fed by continuous versus intermittent bolus tube feeds. Small sample sizes, methodologic limitations and conflicting results of the studies to date, together with inconsistencies in controlling variables that may affect outcomes, make it difficult to make universal recommendations regarding the best tube feeding method for premature infants less than 1500 grams. The clinical benefits and risks of continuous versus intermittent nasogastric tube milk feeding cannot be reliably discerned from the limited information available from randomized trials to date. PMID- 11279732 TI - Laser resurfacing for facial acne scars. AB - BACKGROUND: Most people have acne at some stage during their life, with about one per cent being left with permanent acne scars. Recent laser techniques are thought to be more effective than chemical peels and dermabrasion. OBJECTIVES: To assess the effects of laser resurfacing for treating facial acne scars. SEARCH STRATEGY: We searched MEDLINE (1966 to April 1999), EMBASE (1980 to April 1999), Science Citation Index (1981 to April 1999), the Cochrane Controlled Trials Register (April 1999), DARE (April 1999), INAHTA (April 1999), NHS HTA Internet site (April 1999). Dermatological Surgery (1995 to March 1999) and the British Journal of Dermatology (1995 to September 1999) were handsearched. We searched the reference lists of relevant articles and contacted experts and commercial laser manufacturers. SELECTION CRITERIA: Randomised controlled trials which compare different laser resurfacing techniques for treating patients with facial acne scars, or compare laser resurfacing with other resurfacing techniques or no treatment. DATA COLLECTION AND ANALYSIS: Two reviewers independently selected studies, assessed the quality of studies and extracted data. MAIN RESULTS: No randomised controlled trials where laser treatment was compared to either placebo or a different type of laser were found. Most of the 27 studies uncovered were poor quality case series with small numbers of acne-scarred patients. REVIEWER'S CONCLUSIONS: The lack of good quality evidence does not enable any conclusions to be drawn about the effectiveness of lasers for treating atrophic or ice-pick acne scars. Well designed randomised controlled comparisons of carbon dioxide versus Erbium:YAG laser are urgently needed. PMID- 11279733 TI - Interventions for preventing obesity in children. AB - BACKGROUND: The prevalence of obesity and overweight is increasing world-wide. Obesity in children impacts on their health in both the short and longer term. Obesity prevention strategies are poorly understood. OBJECTIVES: To assess the effectiveness of interventions designed to prevent obesity in childhood. SEARCH STRATEGY: Electronic databases were searched from 1985 to October 1999. SELECTION CRITERIA: Data from RCTs and non-randomised trials with concurrent control group were included. studies with follow up of one year minimum were selected, A priori, this was subsequently amended to include studies with minimum follow up of three months. The possible susceptibility of post hoc questions to bias is addressed. DATA COLLECTION AND ANALYSIS: Two reviewers independently extracted data and assessed study quality. MAIN RESULTS: Seven studies were included, three long-term and four short- term. The studies included were diverse in terms of study design and quality, target population, theoretical underpinning of intervention approach, and outcome measures. As such, it was not possible to combine study findings using statistical methods. REVIEWER'S CONCLUSIONS: The findings of the review suggest that currently there is limited quality data on the effectiveness of obesity prevention programs and as such no generalisable conclusions can be drawn. The need for well-designed studies which examine a range of interventions remains a priority. PMID- 11279734 TI - Betahistine for Meniere's disease or syndrome. AB - BACKGROUND: Meniere's disease is characterised by attacks of hearing loss, tinnitus and disabling vertigo. Betahistine is used by many people to reduce the frequency and severity of these attacks but there is conflicting evidence relating to its effects. OBJECTIVES: The objective of this review was to assess the effects of betahistine in people with Meniere's disease. SEARCH STRATEGY: We searched the Cochrane Controlled Trials Register (The Cochrane Library issue 4,1999), MEDLINE (January 1966 to December 1999), EMBASE (January 1985 to December 1999) and Index Medicus (1962 to 1966). We checked reference lists of articles and contacted pharmaceutical companies for further studies. SELECTION CRITERIA: Randomised controlled studies of betahistine versus placebo in Meniere's disease. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality and extracted data. Study authors were contacted for further information. MAIN RESULTS: Six trials involving 162 patients were included. No trial met the highest quality standard set by the review because of inadequate diagnostic criteria or methods, and none assessed the effect of betahistine on vertigo adequately. Most trials suggested a reduction of vertigo with betahistine and some suggested a reduction in tinnitus but all these effects may have been caused by bias in the methods. One trial with good methods showed no effect of betahistine on tinnitus compared with placebo in 35 patients. None of the trials showed any effect of betahistine on hearing loss. No adverse effects were found with betahistine. REVIEWER'S CONCLUSIONS: There is insufficient evidence to say whether betahistine has any effect on Meniere's disease. PMID- 11279735 TI - Anti-fibrinolytic use for minimising perioperative allogeneic blood transfusion. AB - BACKGROUND: Concerns regarding the safety of transfused blood have prompted re consideration of the use of allogeneic (blood from an unrelated donor) blood transfusion. OBJECTIVES: To assess the effects of the anti-fibrinolytic drugs aprotinin, tranexamic acid, and epsilon aminocaproic acid, on peri-operative red blood cell (RBC) transfusion. SEARCH STRATEGY: We searched MEDLINE (to May 1998), EMBASE (to December 1997), web sites of international health technology assessment agencies (to May 1998). References in identified trials and review articles were checked and authors contacted to identify any additional studies. SELECTION CRITERIA: Randomised controlled trials of anti-fibrinolytic drugs in adults scheduled for non-urgent surgery. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality and extracted data. MAIN RESULTS: We found 61 trials of aprotinin (7027 participants). Aprotinin reduced the rate of RBC transfusion by a relative 30% (RR=0.70: 95%CI: 0.64 to 0.76). The average absolute risk reduction (ARR) was 20.4% (95%CI: 15.6% to 25.3%). On average, aprotinin use saved 1.1 units of RBC (95%CI: 0.69 to 1.47) in those requiring transfusion. Aprotinin also significantly reduced the need for re-operation due to bleeding (RR=0.40: 95%CI: 0.25 to 0.66). We found 18 trials of tranexamic acid (TXA) (1,342 participants). TXA reduced the rate of RBC transfusion by a relative 34% (RR=0.66: 95%CI: 0.54 to 0.81). This represented an ARR of 17.2% (95%CI: 8.7% to 25.7%). TXA use resulted in a saving of 1.03 units of RBC (95%CI: 0.67 to 1.39) in those requiring transfusion. We found four trials of epsilon aminocaproic acid (EACA) (208 participants). EACA use resulted in a statistically non-significant reduction in RBC transfusion (RR=0.48: 95%CI: 0.19 to 1.19). Comparisons between agents Eight trials made 'head-to-head' comparisons between TXA and aprotinin. There was no significant difference between the two drugs in the rate of RBC transfusion: RR=1.21 (95%CI: 0.83 to 1.76) for TXA compared to aprotinin. Adverse Effects Aprotinin did not seem to be associated with an excess risk of adverse effects, including thrombo-embolic events (thrombosis RR=0.64: 95%CI: 0.31 to 1.31) and renal failure (RR=1.19: 95%CI: 0.79 to 1.79). Fewer data were available for TXA and EACA. REVIEWER'S CONCLUSIONS: From this review it appears that aprotinin reduces the need for red cell transfusion, and the need for re-operation due to bleeding, without serious adverse effects. However, there was significant heterogeneity in trial outcomes, and some evidence of publication bias. Similar trends were seen with TXA and EACA, although the data were rather sparse. The poor evaluation of these latter drugs is unfortunate as results suggest they may be equally as effective as aprotinin, but are significantly cheaper. The evidence reviewed here supports the use of aprotinin in cardiac surgery. Further small trials of this drug are not warranted. Future trials should be large enough to compare the efficacy and cost-effectiveness of aprotinin with that of TXA and EACA. PMID- 11279736 TI - Topical negative pressure for treating chronic wounds. AB - BACKGROUND: Chronic wounds mainly affect the elderly and those with multiple health problems. Despite the use of modern dressings, some of these wounds take a long time to heal, fail to heal, or recur, causing significant pain and discomfort to the person and cost to health services. Topical negative pressure is used to promote healing of surgical wounds by using suction to drain excess fluid from wounds. OBJECTIVES: To assess the effectiveness of topical negative pressure (TNP) in treating people with chronic wounds and to identify an optimum TNP regimen. SEARCH STRATEGY: The Cochrane Wounds Group Specialised Trials Register was searched until July 2000. Experts in the field and relevant companies were contacted to enquire about ongoing and recently completed relevant trials. In addition citations within obtained papers were scrutinised to identify additional studies. SELECTION CRITERIA: All randomised controlled trials which evaluated the effectiveness of TNP in treating chronic wounds were considered. DATA COLLECTION AND ANALYSIS: Eligibility for inclusion, data extraction and details of trial quality was conducted by two reviewers independently. A narrative synthesis of results was undertaken as only two small trials fulfilled the selection criteria and they used different outcome measures. MAIN RESULTS: Two small trials with a total of 34 participants evaluated the effectiveness of TNP on chronic wound healing. Trial 1 considered patients with any type of chronic wound; Trial 2 considered patients with diabetic foot ulcers only. The trials compared TNP (as open cell foam dressing with continuous suction) for the first 48 hours with saline gauze dressings. Trial 1 reported a statistically significant reduction in wound volume at 6 weeks in favour of TNP. Trial 2 (continuous suction, followed by intermittent suction after 48 hours) reported a reduction in the number of days to healing and a reduction in wound surface area at 2 weeks in favour of TNP, - although no statistical analysis was reported. REVIEWER'S CONCLUSIONS: The two small trials provide weak evidence suggesting that TNP may be superior to saline gauze dressings in healing chronic human wounds. However, due to the small sample sizes and methodological limitations of these trials, the findings must be interpreted with extreme caution. The effect of TNP on cost, quality of life, pain and comfort was not reported. It was not possible to determine which was the optimum TNP regimen. PMID- 11279737 TI - Levetiracetam add-on for drug-resistant localization related (partial) epilepsy. AB - BACKGROUND: The majority of patients with epilepsy have a good prognosis and their seizures are well controlled by a single antiepileptic drug. However, up to 30% develop refractory seizures, particularly those with partial seizures. In this review, we summarise the current evidence regarding a new antiepileptic drug, levetiracetam, when used as an add-on treatment for drug-resistant localization related (partial) epilepsy. OBJECTIVES: To evaluate the effects of levetiracetam on seizures, side effects, quality of life and cognition, when used as an add-on treatment for patients with a drug-resistant localization related (partial) epilepsy. SEARCH STRATEGY: We searched the Cochrane Epilepsy Group trials register, the Cochrane Controlled Trials Register (Cochrane Library Issue 2, 2000). In addition, we contacted UCB SA (makers of levetiracetam) and experts in the field to seek any ongoing studies or unpublished studies. SELECTION CRITERIA: Randomized placebo controlled add-on trials of levetiracetam in patients with a drug-resistant localization related (partial) epilepsy. DATA COLLECTION AND ANALYSIS: Two reviewers independently selected trials for inclusion and extracted relevant data. The following outcomes were assessed: (a) 50% or greater reduction in total seizure frequency; (b) treatment withdrawal (any reason); (c) side effects; (d) cognitive effects; (e) quality of life. Primary analyses were intention to treat. Sensitivity best and worst case analyses were also undertaken. Summary odds ratios (ORs) were estimated for each outcome. Dose response was evaluated in regression models. MAIN RESULTS: Four trials (1023 patients) were included. All four trials had data for treatment withdrawal and side effect outcomes. Three trials (904 patients) had data for 50% or greater reduction in seizure frequency. Three trials (595 patients) had data for quality of life and cognitive outcomes. The overall Odds Ratio (OR) (95% Confidence Interval (CI)) for 50% or greater reduction in total seizure frequency outcome was 3.81 (2.78,5.22). Dose regression analysis shows clear evidence that levetiracetam reduces seizure frequency with an increase in efficacy with increasing dose of levetiracetam. Approximately 15% of patients taking 1000 mg and 20-30% of patients taking 3000 mg levetiracetam per day have a 50% or greater reduction in seizure frequency. Patients were not significantly more likely to have levetiracetam withdrawn, OR (95% CI) 1.25 (0.87,1.80). The following side effects were significantly associated with levetiracetam: dizziness 2.36 (1.21, 4.61) and infection 1.82 (1.05, 3.14) whereas accidental injury was significantly associated with placebo 0.55 (0.32, 0.93). Quality of life and cognitive effect outcomes suggest that levetiracetam has a positive effect on cognition and some aspects of quality of life. REVIEWER'S CONCLUSIONS: Levetiracetam reduces seizure frequency when used as an add-on treatment for patients with a drug-resistant localization related (partial) epilepsy, and seems well tolerated. Minimum effective and maximum tolerated doses have not been identified. The trials reviewed were of 16-24 weeks duration and results cannot be used to confirm longer term effects. Our results cannot be extrapolated to monotherapy or to patients with other seizure types or epilepsy syndromes. Great care should also be taken with any attempt to apply these results to children. PMID- 11279738 TI - Specialist epilepsy nurses for treating epilepsy. AB - BACKGROUND: Epilepsy is the most common serious neurological condition after stroke, with a 0.5 per cent prevalence, and a two to three per cent life time risk of being given a diagnosis of epilepsy in the developed world. As a result of the perceived deficiencies and suggestions to improve the quality of care offered to people with epilepsy, two models of service provision have been suggested by researchers: specialist epilepsy out-patient clinics (as opposed to the management of patients in general neurology clinics or general medical clinics) and nurse-based liaison services between primary (GP) and secondary/tertiary (hospital based) care. OBJECTIVES: The aim of this review is to overview the evidence from controlled trials investigating the effectiveness of specialist epilepsy nurses compared to routine care. SEARCH STRATEGY: The following databases were searched: The Cochrane Controlled Trials Register (The Cochrane Library, Issue 4, 1999), MEDLINE, GEARS, BIDS (EMBASE=Excepta Medica), ECRI, Effectiveness Healthcare Bulletin, Effectiveness Matters, Bandolier, Evidence Based Purchasing, National Research Register, Vignettes and expert panels from Standing Group on Health Technology Assessment, PsycLit database, World Wide Web sites and reference lists of articles. SELECTION CRITERIA: All randomized controlled and quasi-randomized trials which considered specialist epilepsy nurse interventions with standard or alternative care were included in this review. DATA COLLECTION AND ANALYSIS: Two reviewers independently selected trials for inclusion and extracted the relevant data. The following outcomes were assessed: (a) seizure frequency (b) appropriateness of medication prescribed (c) social or psychological functioning scores (d) knowledge about epilepsy scores (e) objective measures of general health status/quality of life (f) patients' reports of information received (g) number of days spent on sick leave/missing school and employment status (h) costs of care (i) adverse effects. MAIN RESULTS: Three trials were included, two based in general practice and one in a neurology centre. The population of patients differed between trials, for example one study excluded patients with learning disabilities, and one only recruited patients with a new diagnosis. In view of this heterogeneity we decided not to pool results in a meta-analysis. As yet, there is no convincing evidence that specialist epilepsy nurses improve outcomes for people with epilepsy overall. Important outcomes (e.g. seizure frequency, psychosocial functioning, knowledge of epilepsy, general health status, work days lost, depression and anxiety scores) show no significant improvement. There is some evidence that those patients who have not had an epileptic seizure in the last six months are less at risk for depression. There is also evidence that newly diagnosed patients whose knowledge about epilepsy is poor may improve their epilepsy knowledge scores after nurse intervention. REVIEWER'S CONCLUSIONS: It is clearly plausible that specialist epilepsy nurses could improve quality in epilepsy care. However, there is as yet little evidence to support this assumption as the present research base is small. Further research is needed to investigate the effectiveness of specialist epilepsy nurses before such recommendations can be made. PMID- 11279739 TI - Epilepsy clinics versus general neurology or medical clinics. AB - BACKGROUND: Epilepsy is the most common serious neurological condition after stroke, with a 0.5 per cent prevalence, and a two to three per cent life time risk of being given a diagnosis of epilepsy in the developed world. As a result of perceived deficiencies of the quality of care offered to people with epilepsy, two models of service provision have been suggested by researchers: specialist epilepsy out-patient clinics (as opposed to the management of patients in general neurology clinics or general medical clinics) and nurse-based liaison services between primary (GP) and secondary/tertiary (hospital based) care. OBJECTIVES: The aim of this review was to overview the evidence from controlled trials investigating the effectiveness of specialist epilepsy clinics compared to routine care. A second similar review investigating the effectiveness of specialist epilepsy nurses is also underway. SEARCH STRATEGY: We searched the Cochrane Epilepsy Group trials register, the Cochrane Controlled Trials Register (Cochrane Library Issue 4, 1999), MEDLINE (January 1966 to December 1999), GEARS, BIDS (EMBASE=Excepta Medica(1998-99)), ECRI, Effectiveness Healthcare Bulletin, Effectiveness Matters, Bandolier, Evidence Based Purchasing, National Research Register, Vignettes and expert panels from Standing Group on Health Technology Assessment, PsycLit database, World Wide Web sites and reference lists of articles. In addition, we contacted experts in the field. SELECTION CRITERIA: All randomized controlled and quasi-randomized trials which considered specialist epilepsy clinic interventions with standard or alternative care were included in this review. DATA COLLECTION AND ANALYSIS: No controlled trials of suitable quality were identified for inclusion in the review. MAIN RESULTS: No controlled trials of suitable quality were identified for inclusion in the review. REVIEWER'S CONCLUSIONS: It is not known whether specialist epilepsy clinics improve outcomes for people with epilepsy. As yet, there is no high quality evidence which describes their effectiveness in improving care for people with epilepsy. PMID- 11279740 TI - Eversion versus conventional carotid endarterectomy for preventing stroke. AB - BACKGROUND: Carotid endarterectomy is conventionally undertaken by a longitudinal arteriotomy. Eversion carotid endarterectomy (CEA), which employs a transverse arteriotomy and reimplantation of the carotid artery, is reported to be associated with low perioperative stroke and restenosis rates but an increased risk of complications associated with a distal intimal flap. OBJECTIVES: The objective of this review was to determine whether eversion CEA was safe and more effective than conventional CEA. The null-hypothesis was that there was no difference between the eversion and the conventional CEA techniques (performed either with primary closure or patch angioplasty). SEARCH STRATEGY: The reviewers searched MEDLINE and the Cochrane Stroke Group Trials Register (last searched: December 1999), and hand searched eight surgical journals and conference proceedings. Researchers were contacted to identify additional published and unpublished studies. SELECTION CRITERIA: All randomised trials comparing eversion to conventional techniques in patients undergoing carotid endarterectomy were examined in this review. Outcomes were stroke and death, carotid restenosis/occlusion and local complications. DATA COLLECTION AND ANALYSIS: Data were extracted independently by two reviewers to assess eligibility and describe trial characteristics, and by one reviewer for the meta-analyses. Discrepancies were resolved by discussion. When possible, unpublished data were obtained from investigators. MAIN RESULTS: Five trials were included for a total of 2465 patients and 2590 arteries. Three trials included bilateral carotid endarterectomies. In one trial, arteries rather than patients were randomised so that it was not clear how many patients had been randomised in each group, therefore, information on the risk of stroke and death from this study were considered in a separate analysis. There were no significant differences in the rate of perioperative stroke and/or death (1.7% vs 2.6%, odds ratio [OR] 0.44, 95% confidence interval [CI] 0.10-1.82) and stroke during follow-up (1.4% vs 1.7%, OR: 0.84, 95% CI: 0.43-1.64) between eversion and conventional CEA techniques. Eversion CEA was associated with a significantly lower rate of restenosis >50% during follow-up (2.5% vs 5.2%, OR: 0.48, 95% CI: 0.32 -0.72). However, there was no evidence that the eversion technique for CEA was associated with a lower rate of neurological events when compared to conventional CEA. There were no statistically significant differences in local complications between the eversion and conventional group. No data were available to define the cost benefit of eversion CEA technique. REVIEWER'S CONCLUSIONS: Eversion CEA may be associated with low risk of arterial occlusion and restenosis. However, numbers are too small to definitively assess benefits or harms. Reduced restenosis rates did not appear to be associated with clinical benefit in terms of reduced stroke risk, either perioperatively or later. Until further evidence is available, the choice of the CEA technique should depend on the experience and familiarity of the individual surgeon. PMID- 11279742 TI - Chinese medicinal herbs for chronic hepatitis B. AB - BACKGROUND: Hepatitis B virus infection is a serious health problem worldwide. Traditional Chinese medicinal herbs have been widely used to treat chronic liver diseases, and many controlled trials have been done to investigate their efficacy. OBJECTIVES: To assess the efficacy and safety of traditional Chinese medicinal herbs for chronic hepatitis B infection. SEARCH STRATEGY: Searches were applied to the following electronic databases: the CHBG Trials Register, the Cochrane Complementary Medicine Field Trials Register, the Cochrane Library, MEDLINE, EMBASE and BIOSIS. Five Chinese journals and conference proceedings were handsearched. No language restriction was used. SELECTION CRITERIA: Randomised or quasi-randomised trials with at least three months follow-up. Trials of Chinese medicinal herbs (single or compound) compared with placebo, no intervention, general non-specific treatment or interferon treatment were included. Trials of Chinese medicinal herbs plus interferon versus interferon alone were also included. Trials could be double-blind, single-blind or not blinded. DATA COLLECTION AND ANALYSIS: Data were extracted independently by two reviewers. The methodological quality of trials was evaluated using the Jadad-scale plus allocation concealment. Intention-to-treat analyses were performed. MAIN RESULTS: Nine randomised trials, including 936 patients, met the inclusion criteria. Methodological quality was considered adequate in only one trial. There was a significant funnel plot asymmetry (regression coefficient=3.37, standard error 1.40, P=0.047). Ten different medicinal herbs were tested in the nine trials. Compared to non-specific treatment or placebo, Fuzheng Jiedu Tang (compound of herbs) showed significantly positive effects on clearance of serum HBsAg, HBeAg, and HBV DNA; Polyporus umbellatus polysaccharide on serum HBeAg and HBV DNA; Phyllanthus amarus on serum HBeAg. Phyllanthus compound and kurorinone showed no significant effect on clearance of serum HBeAg and HBV DNA and on alanine aminotransferase normalisation compared to interferon treatment. There were no significant effects of the other examined herbs. REVIEWER'S CONCLUSIONS: Some Chinese medicinal herbs may work in chronic hepatitis B. However, the evidence is too weak to recommend any single herb. Rigorously designed, randomised, double blind, placebo-controlled trials are required. PMID- 11279741 TI - Anticoagulants or antiplatelet therapy for non-rheumatic atrial fibrillation and flutter. AB - BACKGROUND: Atrial fibrillation (AF) carries a high risk of stroke and other thromboembolic events. Appropriate use of drugs to prevent thromboembolism in patients with AF involves comparing the patient's risk of stroke to the risk of hemorrhage from medication use. OBJECTIVES: To quantify risk of stroke, major hemorrhage and death from using medications that have been rigorously evaluated for prevention of thromboembolism in AF. SEARCH STRATEGY: Articles were identified through the Cochrane Collaboration's CENTRAL database and MEDLINE until December 1999. SELECTION CRITERIA: Included Randomized controlled trials of drugs to prevent thromboembolism in adults with non-postoperative AF. Excluded RCTS of patients with rheumatic valvular disease. DATA COLLECTION AND ANALYSIS: Data were abstracted by two reviewers. Odds ratios from all qualitatively similar studies were combined, with weighting by study size, to yield aggregate odds ratios for stroke, major hemorrhage, and death for each drug. MAIN RESULTS: Fourteen articles were included in this review. Warfarin was more efficacious than placebo for primary stroke prevention [aggregate odds ratio (OR) of stroke=0.30 [95% Confidence Interval (C.I.) 0.19,0.48]], with moderate evidence of more major bleeding [ OR= 1.90 [95% C.I. 0.89,4.04].]. Aspirin was inconclusively more efficacious than placebo for stroke prevention [OR=0.68 [95% C.I. 0.29,1.57]], with inconclusive evidence regarding more major bleeds [OR=0.81[95% C.I. 0.37,1.78]]. For primary prevention, assuming a baseline risk of 45 strokes per 1000 patient-years, warfarin could prevent 30 strokes at the expense of only 6 additional major bleeds. Aspirin could prevent 17 strokes, without increasing major hemorrhage. In direct comparison, there was moderate evidence for fewer strokes among patients on warfarin than on aspirin [aggregate OR=0.64[95% C.I. 0.43,0.96]], with only suggestive evidence for more major hemorrhage [OR =1.58 [95% C.I. 0.76,3.27]]. However, in younger patients, with a mean age of 65 years, the absolute reduction in stroke rate with warfarin compared to aspirin was low (5.5 per 1000 person-years) compared to an older group (15 per 1000 person-years). Low-dose warfarin or low-dose warfarin with aspirin was less efficacious for stroke prevention than adjusted-dose warfarin. REVIEWER'S CONCLUSIONS: The evidence strongly supports warfarin in AF for patients at average or greater risk of stroke, although clearly there is a risk of hemorrhage. Although not definitively supported by the evidence, aspirin may prove to be useful for stroke prevention in sub-groups with a low risk of stroke, with less risk of hemorrhage than with warfarin. Further studies are needed of low- molecular weight heparin and aspirin in lower risk patients. PMID- 11279743 TI - Superoxide dismutase for preventing chronic lung disease in mechanically ventilated preterm infants. AB - BACKGROUND: Free oxygen radicals have been implicated in the pathogenesis of chronic lung disease in preterm infants. Superoxide dismutase is a naturally occurring enzyme which provides a defence against such oxidant injury. Exogenously administered superoxide dismutase has been tested in clinical trials to prevent chronic lung disease in preterm infants. OBJECTIVES: To determine if exogenously administered superoxide dismutase is efficacious in the prevention of chronic lung disease in preterm infants who are mechanically ventilated, and efficacious in decreasing the following outcomes: bronchopulmonary dysplasia, intraventricular hemorrhage, periventricular leukomalacia, retinopathy of prematurity, necrotizing enterocolitis, patent ductus arteriosus and mortality. To determine the frequency and nature of adverse effects of superoxide dismutase. SEARCH STRATEGY: We searched Medline (1966 - 2000) and the Cochrane Controlled Trials Register (CCTR) using the following keywords: [bronchopulmonary dysplasia OR chronic lung disease] AND superoxide dismutase, limited to human studies in newborn infants (infant, newborn). We hand searched the reference lists of articles located and the abstracts of the Society for Pediatric Research (USA) (published in Pediatric Research) from 1980 - 2000. SELECTION CRITERIA: Randomized controlled trials where subjects were preterm infants who had developed or were at risk of developing respiratory distress syndrome requiring assisted ventilation and who were randomly allocated to receive either superoxide dismutase (in any form, by any route) or placebo or no treatment. We included studies which reported any of the following outcomes: chronic lung disease, bronchopulmonary dysplasia, any intraventricular hemorrhage, intraventricular hemorrhage grades III/IV, patent ductus arteriosus, periventricular leukomalacia, retinopathy of prematurity, necrotizing enterocolitis, neonatal mortality, death prior to discharge and neurodevelopmental outcome. DATA COLLECTION AND ANALYSIS: We extracted and assessed separately all data for each study and entered final data into RevMan. We did not perform subgroup analyses (which were originally planned) because only two studies were eligible for inclusion. We assessed the methodological quality of the studies by assessing the risk for bias. We pooled the outcomes of infants who had developed bronchopulmonary dysplasia at 28 days with those who had died at 28 days to derive the combined outcome of bronchopulmonary dysplasia or death at 28 days. Similarly we pooled the outcomes of infants who had respiratory problems after discharge with those who had died prior to discharge to derive the combined outcome of respiratory problems after discharge or death. We used the standard method of the Cochrane Neonatal Review Group for statistical analysis, using a fixed effect model. MAIN RESULTS: Two randomized controlled trials were included for analysis. No differences were found in either study or in the pooled data in death prior to discharge, oxygen dependency at 36 weeks corrected age, oxygen dependency at 28 days of life or in other outcomes. In one study (Rosenfeld 1984), survivors who had been treated with superoxide dismutase had a shorter duration of continuous positive airway pressure (4.9 vs 9.7 days), a lower frequency of respiratory problems after discharge (relative risk 0.33, 95% confidence limits 0.11, 0.96) and a lower frequency of chest radiograph abnormalities (relative risk 0.30, 95% confidence limits 0.11, 0.87) compared to survivors who received placebo. A third study was available only in abstract form and will be evaluated for inclusion after publication. REVIEWER'S CONCLUSIONS: Based on currently available published trials, there is insufficient evidence to draw firm conclusions about the efficacy of superoxide dismutase in preventing chronic lung disease of prematurity. Data from a small number of treated infants suggest that it is well tolerated and has no serious adverse effects. PMID- 11279744 TI - Surgical procedures to evacuate incomplete abortion. AB - BACKGROUND: Incomplete abortion is a major problem that should be effectively managed with safe and appropriate procedures. Surgical evacuation of the uterus for management of incomplete abortion usually involves vacuum aspiration or sharp curettage. OBJECTIVES: To compare the safety and effectiveness of surgical uterine evacuation methods for management of incomplete abortion. SEARCH STRATEGY: We searched the Cochrane Pregnancy and Childbirth Group trials register, Medline from 1966, Popline from 1970, and the Cochrane Controlled Trials Register. Trials were also identified from reference lists of reviews. Date of last search: October 2000. SELECTION CRITERIA: Randomized trials where different surgical methods were used to manage incomplete abortion were eligible for inclusion. DATA COLLECTION AND ANALYSIS: We extracted population characteristics, settings, and exclusion criteria, in addition to outcomes such as complications of the procedure, duration, need for re-evacuation, blood transfusion, and analgesia/anesthesia. MAIN RESULTS: Two trials were included. Vacuum aspiration was associated with statistically significantly decreased blood loss (-17 mls weighted mean difference, 95% confidence interval (CI) -24 to -10 mls), less pain (relative risk (RR): 0.74, 95% CI 0.61, 0.90), and shorter duration of procedure (-1.2 minutes weighted mean difference, 95% CI -1.5 to 0.87 minutes), than sharp curettage, in the single study that evaluated these outcomes. Serious complications such as uterine perforation and other morbidity were rare and the sample sizes of the trials were not large enough to evaluate small or moderate differences. REVIEWER'S CONCLUSIONS: Vacuum aspiration is safe, quick to perform, and less painful than sharp curettage, and should be recommended for use in the management of incomplete abortion. Analgesia and sedation should be provided as necessary for the procedure. PMID- 11279745 TI - Single versus combination intravenous antibiotic therapy for people with cystic fibrosis. AB - BACKGROUND: Choice of antibiotic, and the use of single or combined therapy are controversial areas in the treatment of respiratory infection in Cystic Fibrosis (CF). Advantages of combination therapy include wider range of modes of action, possible synergy and reduction of resistant organisms; advantages of monotherapy include lower cost, ease of administration and reduction of drug related toxicity. Current evidence does not provide a clear answer and therefore the use of intravenous antibiotic therapy in CF requires further evaluation. OBJECTIVES: To assess the effectiveness of single compared to combination intravenous antibiotic therapy in the treatment of patients with CF. SEARCH STRATEGY: The Cochrane CF and Genetic Disorders Group Specialised Register of Controlled Trials and the abstract books of the three major CF conferences were searched to identify randomised controlled trials. The register was compiled by conducting detailed computer searches of Medline from 1966-present and Embase 1974-1995. SELECTION CRITERIA: Randomised controlled trials comparing a single intravenous antibiotic with a combination of that antibiotic plus a second antibiotic in patients with CF. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality and extracted data. MAIN RESULTS: A total of nine studies including 386 patients compared a single agent to a combination of the same antibiotic and one other. There was a wide variation in the individual antibiotics used in each study. In total, the studies included eight comparisons of a beta-lactam antibiotic (penicillin-related or third generation cephalosporin) with a beta-lactam-aminoglycoside combination and three comparisons of an aminoglycoside with a beta-lactam-aminoglycoside combination. These two groups of studies were analysed as separate subgroups. There was considerable heterogeneity amongst these trials which led to difficulties in performing the review and interpreting the results. The meta-analysis did not demonstrate any significant differences between monotherapy and combination therapy, in terms of lung function, symptom scores and adverse effects. Single therapy was associated with an increase in the number of patients with resistant strains of Ps. aeruginosa at two to eight weeks follow-up. This is an important preliminary finding which needs further clarification with a good quality long term study. These results should be interpreted with caution. All but two of the included trials were published between 1977 and 1988; these were single centre studies with flaws in the randomisation process and small sample size. Overall, the methodological quality was poor. REVIEWER'S CONCLUSIONS: The results of this systematic review of monotherapy versus combination therapy for pulmonary exacerbations in CF are inconclusive. The review raises important methodological issues. There is a need for a randomised controlled trial which needs to be well designed in terms of adequate randomisation allocation, blinding, power and long term follow up. Results need to be standardised to a consistent method of reporting, in order to validate the pooling of results from multiple studies. PMID- 11279746 TI - Opioid antagonists under sedation or anaesthesia for opioid withdrawal. AB - BACKGROUND: Withdrawal (detoxification) is necessary prior to drug-free treatment. It may also represent the end point of long-term treatment such as methadone maintenance. The availability of managed withdrawal is essential to an effective treatment system. OBJECTIVES: To assess the effectiveness of interventions involving the administration of opioid antagonists to induce opioid withdrawal with concomitant heavy sedation or anaesthesia. SEARCH STRATEGY: Multiple electronic databases (including MEDLINE, EMBASE, PsycLIT, Australian Medical Index, Cochrane Clinical Trials Register, and CINAHL) were systematically searched. Reference lists of retrieved studies, reviews and conference abstracts were handsearched. SELECTION CRITERIA: Randomised or quasi-randomised controlled trials that compared the administration of opioid antagonists under sedation or anaesthesia with another form of treatment. DATA COLLECTION AND ANALYSIS: One reviewer assessed studied for inclusion and undertook data extraction. Inclusion decisions and the overall process were confirmed by consultation between all three reviewers. MAIN RESULTS: As yet, no studies have been published comparing treatment regimes involving the administration of opioid antagonists under sedation or anaesthesia with other approaches to detoxification. Treatment regimes for the studies considered for this review varied in the opioid antagonist used, the dose and mode of administration, the anaesthetic agent, duration of anaesthesia and adjunct medications employed. More detailed monitoring of withdrawal is required before any conclusions can be drawn as to what comprises a typical pattern of withdrawal and what factors might influence the pattern. There is only very limited information on referral to ongoing treatment, and relapse to opioid use. Together with the lack of adequate comparisons, this makes it impossible to draw any conclusions about the long-term effectiveness, or the cost-effectiveness, of withdrawal induced by opioid antagonists under sedation or anaesthesia. REVIEWER'S CONCLUSIONS: Considerably more research evidence will be needed before any conclusions can be drawn regarding the effectiveness of managing withdrawal by administration of opioid antagonists under sedation or anaesthesia. The risk of vomiting during sedation and respiratory depression point to the approach being limited to facilities equipped for intubation and assisted ventilation, and with the capacity to respond to adverse events that might occur. The approach must be regarded as experimental with both risks and benefits remaining uncertain. PMID- 11279747 TI - Alpha2 adrenergic agonists for the management of opioid withdrawal. AB - BACKGROUND: Withdrawal (detoxification) is necessary prior to drug-free treatment. It may also represent the end point of long-term treatment such as methadone maintenance. The availability of managed withdrawal is essential to an effective treatment system. OBJECTIVES: To assess the effectiveness of interventions involving the use of alpha2 adrenergic agonists (clonidine, lofexidine, guanfacine, guanabenz acetate) to manage the acute phase of opioid withdrawal. SEARCH STRATEGY: Multiple electronic databases were systematically searched. Reference lists of retrieved studies, reviews and conference abstracts were handsearched and relevant pharmaceutical companies contacted. SELECTION CRITERIA: Randomised or quasi-randomised controlled trials that compared alpha2 adrenergic agonists with another form of treatment (or placebo) to modify the signs and symptoms of withdrawal in participants who were primarily opioid dependent. DATA COLLECTION AND ANALYSIS: One reviewer assessed studies for inclusion and undertook data extraction. Inclusion decisions and the overall process were confirmed by consultation between all four reviewers. MAIN RESULTS: Twenty-four studies, involving 1956 participants, were included. Nineteen were randomised controlled trials; in two allocation was by participant choice, one used alternate allocation and in two the method of allocation was unclear. Ten studies compared a treatment regime based on an alpha2 adrenergic agonist with one based on reducing doses of methadone. Diversity in study design, assessment and reporting of outcomes limited the extent of quantitative analysis. From the comparison of alpha2 adrenergic agonist regimes with reducing doses of methadone, withdrawal intensity is similar to, or marginally greater with alpha2 adrenergic agonists, but signs and symptoms of withdrawal occur and resolve earlier in treatment. Participants stay in treatment longer with methadone. The likelihood of completing withdrawal is similar, or slightly less, with clonidine or lofexidine. Clonidine is associated with more adverse effects (low blood pressure, dizziness, dry mouth, lack of energy) than reducing doses of methadone. Lofexidine does not reduce blood pressure to the same extent as clonidine, but is otherwise similar to clonidine. REVIEWER'S CONCLUSIONS: Treatment regimes based on the alpha2 adrenergic agonists clonidine and lofexidine, and those based on reducing doses of methadone over a period of around 10 days, have similar efficacy in the management of withdrawal from heroin or methadone. Participants stay in treatment longer with methadone regimes and experience less adverse effects. The lower incidence of hypotension makes lofexidine more suited to use in outpatient settings than lofexidine. There are insufficient data available to support a conclusion on the efficacy of guanfacine. PMID- 11279748 TI - Fibreoptic phototherapy for neonatal jaundice. AB - BACKGROUND: Phototherapy is used to treat newborn infants with hyperbilirubinaemia. Fibreoptic phototherapy is a new mode of phototherapy which is reported to lower serum bilirubin (SBR) while minimising disruption of normal infant care. OBJECTIVES: To evaluate the efficacy of fibreoptic phototherapy. SEARCH STRATEGY: The standard search strategy of the Cochrane Collaboration was used including searches of the Cochrane Controlled Trials Register, MEDLINE, EMBASE and discussion with experts in the field. SELECTION CRITERIA: Randomised or quasi-randomised controlled trials evaluating the efficacy of fibreoptic phototherapy in the management of newborn infants with hyperbilirubinaemia. DATA COLLECTION AND ANALYSIS: Thirty-one studies were identified of which 24 met inclusion criteria. They evaluated the efficacy of fibreoptic phototherapy in a number of different clinical situations and patient populations. MAIN RESULTS: Fibreoptic phototherapy was more effective at lowering SBR than no treatment but less effective than conventional phototherapy (percentage change in SBR after 24 hours of treatment: WMD -10.7%, 95%CI -18.14, -3.26 and WMD 3.59%, 95%CI 1.27, 5.92 respectively). Fibreoptic phototherapy was equally as effective as conventional phototherapy in preterm infants and when two fibreoptic devices were used simultaneously (change in SBR after 24 hours of treatment: WMD 1.7%, 95%CI 2.65, 6.05 and change in SBR per day over whole treatment period: WMD 2.82%, 95%CI -1.84, 7.48 respectively). A combination of fibreoptic and conventional phototherapy was more effective than conventional phototherapy alone (duration of phototherapy: WMD -12.51 hr, 95%CI -16.00, -9.02, meta-analysis affected by heterogeneity). No conclusion can be made on the superiority of one fibreoptic device over another as the two studies comparing them (one favouring BiliBlanket, the other finding no difference) did not contain a common outcome measure. REVIEWER'S CONCLUSIONS: Fibreoptic phototherapy has a place in the management of neonatal hyperbilirubinaemia. It is probably a safe alternative to conventional phototherapy in term infants with physiological jaundice. No trials have been identified which support the widely-held view that fibreoptic devices interfere less with infant care or impact less on parent-child bonding. PMID- 11279749 TI - Interventions for the treatment of twin-twin transfusion syndrome. AB - BACKGROUND: Twin-twin transfusion syndrome, a condition affecting monochorionic twin pregnancies, is associated with a high risk of perinatal mortality and morbidity. A number of treatments have been introduced to treat the condition but it is unclear which intervention improves maternal and fetal outcome. OBJECTIVES: The objective of this review was to evaluate the impact of treatment modalities in twin-twin transfusion syndrome. SEARCH STRATEGY: We searched The Cochrane Pregnancy and Childbirth Trials Register and Cochrane Controlled Trials Register. We also searched conference proceedings and made personal contact with experts active in the area of the review. Date of last search: August 2000. SELECTION CRITERIA: Randomised and quasi-randomised studies of amnioreduction versus laser coagulation, septostomy versus laser coagulation or septostomy versus amnioreduction. DATA COLLECTION AND ANALYSIS: Eligibility was assessed by one reviewer. Study authors were contacted for additional information. MAIN RESULTS: No studies were included. REVIEWER'S CONCLUSIONS: There is no current evidence from randomised trials to influence practice. Three ongoing randomised studies have been identified. PMID- 11279750 TI - Interventions for chronic abacterial prostatitis. AB - BACKGROUND: Chronic abacterial prostatitis is a common disabling but enigmatic condition with a symptom complex of pelvic area pain and lower urinary tract symptoms. The scope of treatments recommended for chronic abacterial prostatitis is a testament to how little is known about what causes the condition and how to treat it. As a result, chronic abacterial prostatitis often causes physician frustration, patient confusion and dissatisfaction, variable thresholds for referral, and potentially inappropriate antibiotic use. OBJECTIVES: Examine the evidence regarding the effectiveness of therapies for chronic abacterial prostatitis. SEARCH STRATEGY: Studies were identified through a search of MEDLINE (1966-2000), the Cochrane Library, bibliographies of identified articles and reviews, and contact with an expert. SELECTION CRITERIA: Studies were eligible if they: (1) are randomized controlled trials (RCTs) or controlled clinical trials (CCTs) (2) involve men with chronic abacterial prostatitis (3) control group receives placebo, sham intervention, active pharmacologic or device therapy for chronic abacterial prostatitis and (4) outcomes data are provided. Eligibility was assessed by at least two independent observers. DATA COLLECTION AND ANALYSIS: Study information on patients, interventions, and outcomes was extracted independently by 2 reviewers. The main outcome was the efficacy of treatment for chronic abacterial prostatitis vs. control in improving urologic symptom scale scores or global report of urinary tract symptoms. Secondary outcomes included changes in the prostate examination, uroflowmetry, urodynamics, analysis of urine, expressed prostatic secretions and seminal fluid, and prostate ultrasonography. MAIN RESULTS: The 15 treatment trials involved: medications used to treat benign prostatic hyperplasia (n=4 trials); anti-inflammatory medications (n=2 trials); antibiotics (n=1 trial); thermotherapy (n=5 trials); and miscellaneous medications (n=3 trials). The disparity between studies did not permit quantitative analysis. There were a total of 600 enrollees (age range 38 45). All but one of the trials were done outside the United States. REVIEWER'S CONCLUSIONS: The treatment trials are few, weak methodologically, and involve small sample sizes. The routine use of antibiotics and alpha blockers for chronic abacterial prostatitis is not supported by the existing evidence. The small studies examining thermal therapy appear to demonstrate benefit of clinical significance and merit further evaluation. Additional treatment trials are required and they should report important patient characteristics (e.g., race), study design details and utilize clinically relevant and validated assessment measures. PMID- 11279751 TI - Eradication of Helicobacter pylori for non-ulcer dyspepsia. AB - BACKGROUND: Helicobacter pylori (H pylori) is the main cause of peptic ulcer disease. The role of H pylori in non-ulcer dyspepsia is less clear. OBJECTIVES: To determine the effect of H pylori eradication on dyspepsia symptoms and quality of life scores in patients with non-ulcer dyspepsia. SEARCH STRATEGY: Trials were identified through electronic searches of the Cochrane Controlled Trials Register (CCTR), MEDLINE, EMBASE, CINAHL and SIGLE, using appropriate subject headings and keywords, searching bibliographies of retrieved articles, and through contacts with experts in the fields of dyspepsia and with pharmaceutical companies. SELECTION CRITERIA: All parallel group randomised controlled trials (RCTs) comparing drugs to eradicate H pylori with placebo or other drugs known not to eradicate H pylori for patients with non-ulcer dyspepsia. DATA COLLECTION AND ANALYSIS: Data were collected on individual and global dyspeptic symptom scores, quality of life measures and adverse effects. Dyspepsia outcomes were dichotomised into minimal/resolved versus same/worse symptoms. MAIN RESULTS: Twelve randomised controlled trials were included in the systematic review. Ten trials compared antisecretory dual or triple therapy with placebo antibiotics +/- antisecretory therapy, and evaluated dyspepsia at 3-12 months. Nine of these trials gave results as dichotomous outcomes evaluating 2,541 patients and there was no significant heterogeneity between the studies. There was a 9% relative risk reduction in the H pylori eradication group (95% CI = 4% to 14%) compared to placebo. The number needed to treat to cure one case of dyspepsia = 15 (95% CI = 10 to 31). A further two trials compared Bismuth based H pylori eradication with an alternative pharmacological agent. These trials were smaller and had a shorter follow-up but suggested H pylori eradication was more effective than either H2 receptor antagonists or sucralfate in treating non-ulcer dyspepsia. REVIEWER'S CONCLUSIONS: H pylori eradication therapy has a small but statistically significant effect in H pylori positive non-ulcer dyspepsia. An economic model suggests this modest benefit may still be cost-effective but more research is needed. PMID- 11279752 TI - Hypothermia to reduce neurological damage following coronary artery bypass surgery. AB - BACKGROUND: Coronary artery bypass surgery (CABG) may be life saving, but known side effects include neurological damage and cognitive impairment. The temperature used during cardiopulmonary bypass (CPB) may be important with regard to these adverse outcomes, where hypothermia is used as a means of neuroprotection. OBJECTIVES: To assess the effectiveness of hypothermia during CABG in reducing neurological damage and subsequent cognitive deficits. SEARCH STRATEGY: The Cochrane Controlled Trials Register was searched for randomised controlled trials (RCT) and this was updated by searching MEDLINE and EMBASE to December 1999 using database specific RCT filters. Reference lists of retrieved articles were searched and experts in the field were contacted. SELECTION CRITERIA: Only RCTs were considered. All patients undergoing CABG, either first time or revisions, elective or emergency procedures, were included. Any hypothermia protocol was considered. Only trials reporting neurological outcomes were included. DATA COLLECTION AND ANALYSIS: Studies were selected independently and data were extracted from the source papers independently by two reviewers. Authors were contacted for further information. Studies were combined with meta analysis where appropriate, and meta-regression was used to explore heterogeneity. MAIN RESULTS: There was a trend towards a reduction in the incidence of non fatal strokes in the hypothermic group (OR 0.68 (0.43, 1.05)). Conversely, there was a trend for the number of non stroke related perioperative deaths to be higher in the hypothermic group (OR 1.46 (0.9, 2.37)). Hypothermia had no effect on the incidence of non fatal myocardial infarction (OR 1.05 (0.81, 1.37)), but the incidence of another marker of myocardial damage, low output syndrome, was higher in the hypothermic group (OR 1.21 (0.99, 1.48). When pooling all "bad" outcomes (stroke, perioperative death, myocardial infarction, low output syndrome, intra aortic balloon pump use) there was no significant advantage of either hypothermia or normothermia (OR 1.07 (0.92, 1.24)). Only 4 of 17 trials reported neuropsychological function as an outcome. REVIEWER'S CONCLUSIONS: This review could find no definite advantage of hypothermia over normothermia in the incidence of clinical events. Hypothermia was associated with a reduced stroke rate, but this is off set by a trend towards an increase in non stroke related perioperative mortality and myocardial damage. There is insufficient data to date to draw any conclusions about the use of mild hypothermia. Similarly, there is insufficient data to date to comment on the effect of temperature during CPB on subtle neurological deficits, and further trials are needed in these areas. PMID- 11279753 TI - Terlipressin for acute esophageal variceal hemorrhage. AB - BACKGROUND: Terlipressin (triglycyl lysine vasopressin) is a synthetic analogue of vasopressin, which has been used in the treatment of acute variceal hemorrhage. In contrast to vasopressin, terlipressin can be administered as intermittent injections instead of continuous intravenous infusion and it has a safer adverse reactions profile. However, its effectiveness remains uncertain. OBJECTIVES: To determine if treatment with terlipressin improves outcome in acute esophageal variceal hemorrhage and is safe. SEARCH STRATEGY: Randomized clinical trials were identified by searching the following databases: MEDLINE, EMBASE, the Cochrane Controlled Trials Register, the Cochrane Hepato-Biliary Group Controlled Trials Register, Biosis, and Current Contents. The bibliographies of identified publications were checked. Experts in the field and the manufacturers of terlipressin were contacted. SELECTION CRITERIA: All randomized clinical trials which compared terlipressin with: (a) placebo or no treatment, (b) balloon tamponade, (c) endoscopic treatment, (d) octreotide, (e) somatostatin and (f) vasopressin, in the setting of acute variceal hemorrhage. DATA COLLECTION AND ANALYSIS: Eligibility, trial quality assessment and data extraction were done independently by two reviewers. The primary outcome measure was mortality. Secondary outcomes were failure of initial hemostasis, rebleeding, procedures required for uncontrolled bleeding or rebleeding, transfusion requirements and length of hospitalization. MAIN RESULTS: Twenty studies were identified for all the comparison groups, involving 1609 patients. There were seven studies (with 443 patients) comparing terlipressin to placebo, five of which were considered to be high quality studies based on the Jadad scale. The meta-analysis indicates that terlipressin was associated with a statistically significant reduction in all cause mortality compared to placebo (relative risk 0.66, 95% confidence interval 0.49 to 0.88). Three studies (with 302 patients) were identified comparing terlipressin to somatostatin, two of which were high quality studies; only one high quality study (219 patients) comparing terlipressin to endoscopic treatment was identified. Within the limited power provided by these small numbers of patients, no statistically significant difference was demonstrated between terlipressin and either somatostatin or endoscopic treatment in any of the outcomes. For the remaining comparison groups (terlipressin versus balloon tamponade, terlipressin versus octreotide and terlipressin versus vasopressin) only small, low quality studies were identified and no difference was demonstrated in any of the major outcomes. There was no difference between the terlipressin group and any of the comparison groups in the number of adverse events that caused death or withdrawal of medication. REVIEWER'S CONCLUSIONS: On the basis of a 34% relative risk reduction in mortality, terlipressin should be considered to be effective in the treatment of acute variceal hemorrhage. Further, since no other vasoactive agent has been shown to reduce mortality in single studies or meta-analyses, terlipressin might be the vasoactive agent of choice in acute variceal bleeding. PMID- 11279754 TI - Inhaled vs oral steroids for adults with chronic asthma. AB - OBJECTIVES: To determine therapeutically equivalent doses of inhaled versus oral steroids for adults with chronic asthma. SEARCH STRATEGY: The Cochrane Airways Group trials register was searched using the terms: (drug delivery systems OR ((nebuli* OR inhal* OR MDI) AND oral*)) AND ( steroid* OR corticosteroid* OR glucocorticoid* OR beclomethasone OR betamethasone OR fluticasone OR cortisone OR dexamethasone OR hydrocortisone OR prednisolone OR prednisone OR triamcinolone). SELECTION CRITERIA: Randomised controlled trials were selected of at least 4 weeks duration and included patients over the age of 15 years with chronic asthma. Trials compared inhaled steroids and oral prednisolone or prednisone; where the maximum dose for inhaled steroids was 2000 mcg/day and prednisolone 60 mg (on alternate days). DATA COLLECTION AND ANALYSIS: Two independent reviewers screened 1285 titles and abstracts from the electronic search, bibliography searches and other contacts. Of these, 10 trials met previously defined inclusion criteria. Two reviewers independently extracted study characteristics, and outcome measures. MAIN RESULTS: All trials were small and no data could be pooled. Carry-over effects were present in at least one cross-over trial. Data from six trials produced the same pattern, in which prednisolone 7.5-12 mg/day appeared to be as effective as inhaled steroid 300-2000 mcg/day. In two trials, inhaled steroid 300-400 mcg/day was more effective than prednisolone 5 mg/day. All doses of inhaled steroid appeared to be more effective than alternate day doses of prednisolone up to 60 mg on alternate days. Side-effect data were reported too variably to permit comparisons. A 30% incidence was reported in one study in patients receiving prednisolone 5 mg/day, none were reported in patients on inhaled steroids. A further search was conducted in October 2000 which yielded no further trials. REVIEWER'S CONCLUSIONS: A daily dose of prednisolone 7.5-10 mg/day appears to be equivalent to moderate-high dose inhaled corticosteroids. Side-effects may be present on low doses, so if there is no alternative to oral steroids, the lowest effective dose should be prescribed. PMID- 11279755 TI - Methyl-xanthines for exacerbations of chronic obstructive pulmonary disease. AB - BACKGROUND: International guidelines currently recommend the use of methyl xanthines for exacerbations of chronic obstructive pulmonary disease (COPD) for patients who have incomplete responses to bronchodilators. However, available clinical trials are small and underpowered to evaluate the benefits and risks of methyl-xanthines in this acute setting. OBJECTIVES: To determine the benefit of methyl-xanthines compared to standard care for COPD exacerbations. SEARCH STRATEGY: Randomised controlled trials (RCTs) were identified from the Cochrane Airways Review Group COPD Register which is a compilation of systematic searches of CINAHL, EMBASE, MEDLINE and CENTRAL and hand searching of 20 respiratory journals. In addition, primary authors and content experts were contacted to identify eligible studies. Bibliographies from included studies, known reviews and texts were also searched. SELECTION CRITERIA: Only RCTs were eligible for inclusion. Studies were included if patients presented with acute COPD exacerbations and were treated with either methyl-xanthines (oral or intravenous) or placebo (with or without standard care) early in the acute treatment. Studies also needed to report either pulmonary function or admission results. Two reviewers independently selected potentially relevant articles and selected articles for inclusion. Methodological quality was independently assessed by two reviewers. DATA COLLECTION AND ANALYSIS: Data were extracted independently by two reviewers if the authors were unable to verify the validity of information. Missing data were obtained from authors or calculated from other data presented in the paper. The data were analysed using the Cochrane Review Manager 4.0.4 Studies were pooled to yield weighted mean differences (WMD) or odds ratios (OR) and reported using 95% confidence intervals (95%CI). MAIN RESULTS: From 28 identified references, 4 RCTs met inclusion criteria (172 patients). Mean change in forced expiratory volume in one second (FEV1) at 2 hours was similar in methyl xanthine and placebo groups (FEV1 WMD: -8 ml; 95% CI: -85 to 69 ml). The only study to report hospitalization rates showed a non-significant reduction with methyl-xanthines (OR: 0.3; 95% CI: 0.1 to 1.8) among 39 patients. Patients receiving methyl-xanthines had similar improvements in symptom scores, but reported more gastrointestinal side effects (OR: 5.3; 95% CI: 1.3 to 21.0) than patients receiving placebo. REVIEWER'S CONCLUSIONS: There is no evidence to support the routine use of methyl-xanthines for COPD exacerbations. Methyl xanthines do not appreciably improve FEV1 during COPD exacerbations and cause adverse effects; evidence of their effect on admissions is limited. PMID- 11279756 TI - Early emergency department treatment of acute asthma with systemic corticosteroids. AB - BACKGROUND: The airway edema and secretions associated with acute asthma are most effectively treated with anti-inflammatories such as corticosteroids delivered by inhaled, oral, intravenous or intra-muscular routes. There is an unresolved debate about the use of systemic corticorticoids in the early treatment of acute asthma for emergency department patients. OBJECTIVES: To determine the benefit of treating patients with acute asthma with systemic corticosteroids within an hour of presenting to the emergency department (ED). SEARCH STRATEGY: Randomised controlled trials were identified from the Cochrane Airways Group Asthma Register. Primary authors and content experts were contacted to identify eligible studies. Bibliographies from included studies and known reviews were searched. SELECTION CRITERIA: Only randomised controlled trials (RCTs) or quasi-randomised trials were eligible for inclusion. Studies were included if patients presenting to the ED with acute asthma were treated with IV/IM or oral corticosteroids (CS) vs. placebo within 1 hour of arrival and either admission rate or pulmonary function results were reported. DATA COLLECTION AND ANALYSIS: Trial selection, data extraction and quality assessment were carried out independently by two reviewers, and confirmed with corresponding authors. MAIN RESULTS: Twelve studies involving 863 patients (435 corticosteroids; 428 placebo) were included. Early use of CS for acute asthma in the ED significantly reduced admission rates (N = 11; pooled OR: 0.40, 95% CI: 0.21 to 0.78). This would correspond with a number needed to treat of 8 (95% CI: 5 to 21). This benefit was more pronounced for those not receiving systemic CS prior to ED presentation (N = 7; OR: 0.37, 95% CI: 0.19 to 0.70) and those with more severe asthma (N = 7; OR: 0.35, 95% CI: 0.21 to 0.59). Oral CS therapy in children was particularly effective (N = 3; OR: 0.24, 95% CI: 0.11 to 0.53); no trials in adults used the oral route. Side effects were not significantly different between corticosteroid treatments and placebo. A further search was conducted in September 2000 which did not yield any further trials. REVIEWER'S CONCLUSIONS: Use of corticosteroids within 1 hour of presentation to an ED significantly reduces the need for hospital admission in patients with acute asthma. Benefits appear greatest in patients with more severe asthma, and those not currently receiving steroids. Children appear to respond well to oral steroids. PMID- 11279757 TI - Angiotensin converting enzyme inhibitors in normotensive diabetic patients with microalbuminuria. AB - OBJECTIVES: To examine whether the progression of early diabetic renal disease to end-stage renal failure may be slowed by the use of angiotensin converting enzyme inhibitors for reasons other than their antihypertensive properties, so that they have value in the treatment of normotensive diabetics with microalbuminuria. SEARCH STRATEGY: Medline was searched for English language reviews and randomised controlled trials. Personal reference lists, and reference lists of retrieved studies were also used. SELECTION CRITERIA: Randomised controlled trials with separate identifiable results for initially normotensive diabetic patients, who received angiotensin converting enzyme inhibitors for at least one year and were compared with controls. DATA COLLECTION AND ANALYSIS: Meta-analyses were performed on the results of 12 randomised controlled trials with a variety of patient inclusion and exclusion criteria. One further study met all conditions for inclusion but did not provide data in useable form for meta-analyses. MAIN RESULTS: Albumin excretion rate fell for patients on angiotensin converting enzyme inhibition in 12 of the 13 studies but did so for only two of the 13 groups on placebo. Treatment provided a significant reduction in albumin excretion rate in both insulin dependent diabetes mellitus and non insulin dependent diabetes mellitus. Treatment with either captopril, enalapril or lisinopril reduced albumin excretion rate in comparison with control patients. A significantly greater lowering of blood pressure was experienced by initially normotensive patients in the angiotensin converting enzyme inhibitor than in the placebo group. Average glycosylated haemoglobin fell a little in the treated patients and rose in the controls, the difference being just significant. The difference in changes in glomerular filtration rate did not reach statistical significance. REVIEWER'S CONCLUSIONS: Inhibition of angiotensin converting enzyme can arrest or reduce the albumin excretion rate in microalbuminuric normotensive diabetics, as well as reduce or prevent an increase in blood pressure. But, given the drop in blood pressure in patients on angiotensin converting enzyme inhibitors, it is not certain that the reduction of albumin excretion rate is due to a separate renal effect. A direct link with postponement of end-stage renal failure has not been demonstrated. There appear to be no substantial side effects. PMID- 11279758 TI - Media-based behavioural treatments for behavioural disorders in children. AB - BACKGROUND: Prevalence studies show that behaviour problems in children are quite common (10-15% in preschoolers). These problems may manifest as, for example anxiety, sadness, over-activity and tantrums. Some studies have shown that these problems can be persistent, and that they lead to a range of problems in adolescence and adulthood. Many approaches are used to address behavioural problems such as medication, or more usually, psychological treatments either directly with the child and/or his/her family. Behavioural interventions have been shown to be highly effective but access to these treatments is limited due to factors such as time and expense. Presenting the information parents need in order to manage these behaviour problems in booklet or other media-based format would reduce the cost and thus increase access to these interventions. In the adult population it seems that media-based interventions such as these can be moderately effective. Given that the cost of media-based treatment is so low it is useful to know how effective they are when given to parents. It was hypothesised that media-based treatments would be less effective than conventional psychological treatments and that efficacy would improve with increasing amounts of therapist intervention. OBJECTIVES: To review the effects of media-based behavioural therapies (definitions below), for any young person with a behavioural disorder (however diagnosed) compared to standard care and no treatment controls. SEARCH STRATEGY: A range of electronic databases were systematically searched using a specified search strategy. Individual journals of interest were hand-searched where necessary, references in all selected trials were checked for other trials and, where it was thought to be of possible use, individual authors were contacted for additional information. SELECTION CRITERIA: Randomised controlled trials of behavioural media-based treatments for behaviour problems in children. DATA COLLECTION AND ANALYSIS: Abstracts and titles of studies identified from searches of electronic databases were read to determine whether they met the inclusion criteria. Full copies of those possibly meeting these criteria from electronic or other searches were assessed by the reviewer and queries were resolved by discussion with an independent rater. Data were analysed using Revman. MAIN RESULTS: In general, media-based therapies for behavioural disorders in children had a moderate effect when compared with both no-treatment controls and with standard care. Significant improvements were often made with the addition of up to 2 hours of therapist time. REVIEWER'S CONCLUSIONS: These formats of delivering behavioural interventions for carers of children are possibly worth considering in clinical practice. For straightforward cases media-based interventions may be enough to make clinically significant changes in a child's behaviour, and reduce the amount of time primary care workers have to devote to each case. Consequently this would increase the number of families who could possibly benefit from this type of intervention. Media based therapies would therefore appear to have both clinical and economic implications as regards the treatment of children with behavioural problems. PMID- 11279759 TI - Interprofessional education: effects on professional practice and health care outcomes. AB - BACKGROUND: As patient care becomes more complex, effective collaboration between health and social care professionals is required. However, evidence suggests that these professionals do not collaborate well together. Interprofessional education (IPE) offers a possible way forward in this area. OBJECTIVES: To assess the usefulness of IPE interventions compared to education in which the same professions were learning separately from one another. SEARCH STRATEGY: We searched the Cochrane Effective Practice and Organisation of Care Group specialised register, MEDLINE (1968 to 1998) and Cinahl (1982 to 1998). We also hand searched the Journal of Interprofessional Care (1992 to 1998), the Centre for the Advancement of Interprofessional Education Bulletin (1987 to 1998), conference proceedings, the 'grey literature' held by relevant organisations, and reference lists of articles. SELECTION CRITERIA: Randomised trials, controlled before and after studies and interrupted time series studies of IPE interventions designed to improve collaborative practice between health/social care practitioners and/or the health/well being of patients/clients. The participants included chiropodists/podiatrists, complementary therapists, dentists, dietitians, doctors/physicians, hygienists, psychologists, psychotherapists, midwives, nurses, pharmacists, physiotherapists, occupational therapists, radiographers, speech therapists and/or social workers. The outcomes included objectively measured or self reported (validated instrument) patient/client outcomes and reliable (objective or validated subjective) health care process measures. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed the eligibility of potentially relevant studies. MAIN RESULTS: The total yield from the search strategy was 1042, of which 89 were retained for further consideration. However none of these studies met the inclusion criteria. REVIEWER'S CONCLUSIONS: Despite finding a large body of literature on the evaluation of IPE, these studies lacked the methodological rigour needed to begin to convincingly understand the impact of IPE on professional practice and/or health care outcomes. PMID- 11279760 TI - Ivermectin for onchocercal eye disease (river blindness). AB - BACKGROUND: It is believed that ivermectin (a microfilaricide) could prevent blindness due to onchocerciasis. However, when given to everyone in communities where onchocerciasis is common, the effects of ivermectin on lesions affecting the eye are uncertain and data on whether the drug prevents visual loss is unclear. OBJECTIVES: The aim of this review is to assess the effectiveness of ivermectin in preventing visual acuity and visual field loss in onchocercal eye disease. The secondary aim is to assess the effects of ivermectin on lesions affecting the eye in onchocerciasis. SEARCH STRATEGY: We searched the Cochrane Eyes and Vision Group specialised register, the Cochrane Controlled Trials Register - CENTRAL, MEDLINE, EMBASE, the reference lists of identified trials, the Science Citation Index and we contacted investigators, experts and pharmaceutical companies to identify additional trials. SELECTION CRITERIA: We included randomised controlled trials with at least one year follow up, comparing ivermectin at a dose of 150 micrograms per kilogram of body weight with either placebo or no treatment. Participants were people normally resident in endemic onchocercal communities with or without one or more characteristic signs of ocular onchocerciasis. DATA COLLECTION AND ANALYSIS: Two reviewers independently extracted data and assessed trial quality. Study authors were contacted for additional information. Trials varied in design and setting, so no meta-analysis was done. MAIN RESULTS: This review includes five trials with data from 3810 participants. All the trials compared ivermectin with placebo and were judged to be of moderate risk of bias in terms of methodological quality. No statistically significant difference was observed in any trial (reporting visual acuity outcome) between ivermectin and placebo groups for visual acuity loss. REVIEWER'S CONCLUSIONS: Questions about the effectiveness of ivermectin in preventing visual acuity loss have not been answered by best available evidence. PMID- 11279761 TI - Timing and volume of fluid administration for patients with bleeding following trauma. AB - BACKGROUND: Treatment of haemorrhagic shock involves maintaining blood pressure and tissue perfusion until bleeding is controlled. Different resuscitation strategies have been used to maintain the blood pressure in trauma patients until bleeding is controlled. However, while maintaining blood pressure may prevent shock, it may worsen bleeding. OBJECTIVES: To assess the effects of early versus delayed, and larger versus smaller volume of fluid administration in trauma patients with bleeding. SEARCH STRATEGY: We searched the Cochrane Controlled Trials Register, the specialised register of the Injuries Group, MEDLINE, EMBASE, the National Research Register and the Science Citation Index. We checked reference lists of identified articles and contacted authors and experts in the field. SELECTION CRITERIA: Randomised trials of the timing and volume of intravenous fluid administration in trauma patients with bleeding. Trials in which different types of intravenous fluid were compared were excluded. DATA COLLECTION AND ANALYSIS: Two reviewers independently extracted data and assessed trial quality. MAIN RESULTS: We did not combine the results quantitatively because the interventions and patient populations were so diverse. Early versus delayed fluid administration: Three trials reported mortality and two coagulation data. In the first trial (n=598) relative risk (RR) for death with early fluid administration was 1.26 (95% confidence interval of 1.00-1.58). The weighted mean differences (WMD) for prothrombin time and partial thromboplastin time were 2.7 (95% CI 0.9-4.5) and 4.3 (95% CI 1.74-6.9) seconds respectively. In the second trial (n=50) RR for death with early blood transfusion was 5.4 (95% CI 0.3 107.1). The WMD for partial thromboplastin time was 7.0 (95% CI 6.0-8.0) seconds. In the third trial (n=1309) RR for death with early fluid administration was 1.06 (95% CI 0.77-1.47). Larger versus smaller volume of fluid administration: Three trials reported mortality and one coagulation data. In the first trial (n=36) RR for death with a larger volume of fluid resuscitation was 0.80 (95% CI 0.28 22.29). Prothrombin time and Partial thromboplastin time were 14.8 and 47.3 seconds in those who received a larger volume of fluid as compared to 13.9 and 35.1 seconds in the comparison group. In the second trial (n=99) RR for death with a high (100 mm Hg) compared to low (70 mm Hg) systolic blood pressure resuscitation target was 1.02 (95% CI 0.27-3.85). In the third trial (n=25) there were no deaths. REVIEWER'S CONCLUSIONS: We found no evidence from randomised controlled trials to support early or larger volume of intravenous fluid administration in uncontrolled haemorrhage. There is continuing uncertainty about the best fluid administration strategy in bleeding trauma patients. Further randomised controlled trials are needed to establish the most effective fluid resuscitation strategy. PMID- 11279762 TI - Clotiapine for acute psychotic illnesses. AB - BACKGROUND: Acute psychotic illness, especially when associated with agitated or violent behaviour, requires urgent pharmacological tranquillisation or sedation. Clotiapine, a dibenzothiazepine neuroleptic, is being used for this purpose in several countries. OBJECTIVES: To estimate the effects of clotiapine when compared to other 'standard' or 'non-standard' treatments of acute psychotic illness in controlling disturbed behaviour and reducing psychotic symptoms. SEARCH STRATEGY: The Cochrane Controlled Trials Register (Issue 2, 2000), The Cochrane Schizophrenia Group's Register (May 2000), EMBASE (1980-2000), MEDLINE (1966-2000), PASCAL (1973-2000) and PsycLIT (1970-2000) were methodically searched. This was supplemented by hand searching reference lists, contacting industry and relevant authors. SELECTION CRITERIA: Randomised clinical trials comparing clotiapine to any treatment, for people with acute psychotic illnesses such as in schizophrenia, schizoaffective disorder, mixed affective disorders, manic phase of bipolar disorder, brief psychotic episode or organic psychosis following substance abuse. DATA COLLECTION AND ANALYSIS: Studies were reliably selected, quality assessed and data extracted. Data were excluded where more than 50% of participants in any group were lost to follow up. For binary outcomes a standard estimation of the risk ratio (RR) and its 95% confidence interval (CI) was calculated. Where possible, the weighted number needed to treat statistic (NNT), and its 95% confidence interval (CI), was also calculated. If heterogeneity was found, a random effects model was used. For continuous outcomes, endpoint data were preferred to change data. Non-skewed data from valid scales were summated using a weighted mean difference (WMD). Again, if heterogeneity was found a random effects model was used. A Mantel-Haenszel chi square test was used to investigate the possibility of heterogeneity. MAIN RESULTS: Five trials were included. None compared clotiapine with placebo, but control drugs were either antipsychotics (chlorpromazine, perphenazine, trifluoperazine and zuclopenthixol acetate) or benzodiazepines (lorazepam). Versus antipsychotics: results for global clinical outcome were heterogeneous (p=0.09) but did not suggest clotiapine to be superior, or inferior, to chlorpromazine, perphenazine, or trifluoperazine (total randomised = 83). Use of clotiapine did change the proportion of people ready for hospital discharge by the end of the study in one small trial (n=49, RR 1.04 95%CI 0.96 to 2.12). Overall, attrition rates were low. No significant difference was found for those allocated to clotiapine compared with people randomised to other antipsychotics (n=121, RR 2.26 95%CI 0.40 to 13). Weak data suggests that clotiapine may result in less need for antiparkinsonian treatment compared with zuclopenthixol acetate (n=38, RR 0.43 95%CI 0.02 to 0.98). Versus lorazepam: when used to control aggressive/violent outbursts for people already treated with haloperidol, clotiapine did not significantly improve mental state compared to lorazepam (WMD 3.36 95%CI -8.09 to 1.37). Much data could not be pooled due to skew or inadequate presentation of results. Economic outcomes and satisfaction with care were not addressed. REVIEWER'S CONCLUSIONS: We found no significant evidence to support the use of clotiapine rather than other 'standard' or 'non-standard' treatments for the management of acute psychotic illness. The trials included in this review all present important methodological flaws. We do not wish to discourage clinicians from using clotiapine in the psychiatric emergency, we would just like to point out the fact that good quality controlled trials are needed on this subject. PMID- 11279763 TI - Early use of inhaled corticosteroids in the emergency department treatment of acute asthma. AB - BACKGROUND: Systemic corticosteroid therapy is central to the management of acute asthma The use of ICS may also be beneficial in this setting. OBJECTIVES: To determine the benefit of ICS for the treatment of patients with acute asthma managed in the emergency department (ED). SEARCH STRATEGY: Randomised controlled trials (RCTs) were identified from the Cochrane Airways Review Group register. Bibliographies from included studies, known reviews, and texts also were searched. SELECTION CRITERIA: Only RCTs or quasi-randomised trials were eligible for inclusion. Studies were included if patients presented with acute asthma to the ED or its equivalent, and were treated with ICS or placebo, in addition to standard therapy. Two reviewers independently selected potentially relevant articles, and then independently selected articles for inclusion. Methodological quality was independently assessed by two reviewers. DATA COLLECTION AND ANALYSIS: Data were extracted independently by two reviewers if the authors were unable to verify the validity of extracted information. Missing data were obtained from the authors or calculated from other data presented in the paper. MAIN RESULTS: Seven trials were selected for inclusion, but data were not available for one of them. In the six usable trials, (4 adult, 2 paediatric), a total of 352 patients were studied (179 ICS, 173 non-ICS treated). Patients treated with ICS were less likely to be admitted to hospital (OR: 0.33; 95% CI: 0.17, 0.64). This benefit was confined to patients not receiving concomitant systemic steroids (CS). Patients receiving concomitant CS showed a similar, but non-significant, trend towards reduced admissions compared to placebo treatment (OR 0.45; 95% CI: 0.18, 1.14). Patients receiving ICS also demonstrated small, significant improvements in peak expiratory flows (PEFR WMD: 8%; 95% CI: 3, 13 %) and forced expiratory volumes (FEV1 WMD: 5%; 95% CI: 0.4, 10 %). The treatment was well tolerated, with few reported adverse side effects. A secondary analysis compared ICS alone vs CS alone; in the four trials included, there was significant heterogeneity between the study results for admission rates which precluded meaningful pooling of the study results. REVIEWER'S CONCLUSIONS: Inhaled steroids reduced admission rates in patients with acute asthma, but it is unclear if there is a benefit of ICS when used in addition to systemic corticosteroids. There is insufficient evidence that ICS therapy results in clinically important changes in pulmonary function or clinical scores when used in acute asthma. Similarly, there is insufficient evidence that ICS alone is as effective as CS. Further research is needed to clarify if there is a benefit of ICS when used in addition to CS. PMID- 11279764 TI - Oral methyl-xanthines for bronchiectasis. AB - BACKGROUND: Bronchiectasis is characterised by chronic sputum production,bronchial wall dilation,recurrent infection and airflow limitation. Methylxanthines are used in the management of airflow limitation associated with asthma and COPD, where they are also purported to have anti-inflammatory properties. In theory they may be of use in bronchiectasis. OBJECTIVES: To determine the efficacy of methylxanthines in the treatment of bronchiectasis. SEARCH STRATEGY: The Cochrane Airways Group clinical trials register derived from MEDLINE,EMBASE and hand searches using the terms bronchiectasis, aminophylline, theophylline and methyl- xanthine SELECTION CRITERIA: Only randomised controlled trials were to be considered. DATA COLLECTION AND ANALYSIS: The results of the searches were reviewed by two authors. Searches yielded seven trials none of which met the inclusion criteria. MAIN RESULTS: No randomised controlled trials were identified. REVIEWER'S CONCLUSIONS: Further research is required to establish if the methylxanthines have a role in the treatment of bronchiectasis. PMID- 11279765 TI - A comparison of physiotherapy techniques for patients with Parkinson's disease. AB - BACKGROUND: Despite optimal medical and surgical therapies for Parkinson's disease, patients develop progressive disability. The role of the physiotherapist is to maximise functional ability and minimise secondary complications through movement rehabilitation within a context of education and support for the whole person. What form of physiotherapy is most effective in the treatment of Parkinson's disease remains unclear. OBJECTIVES: 1. To compare the efficacy and effectiveness of novel physiotherapy techniques versus 'standard' physiotherapy in patients with Parkinson's disease. Standard physiotherapy is defined as the type of therapy that the physiotherapist would usually use to treat Parkinson's disease. 2. To compare the efficacy and effectiveness of one physiotherapy technique versus a second form of physiotherapy. SEARCH STRATEGY: Relevant trials were identified by electronic searches of MEDLINE, EMBASE, CINAHL, ISI-SCI, AMED, MANTIS, REHABDATA, REHADAT, GEROLIT, Pascal, LILACS, MedCarib, JICST-EPlus, AIM, IMEMR, SIGLE, ISI-ISTP, DISSABS, Conference Papers Index, Aslib Index to Theses, the Cochrane Controlled Trials Register, the CentreWatch Clinical Trials listing service, the metaRegister of Controlled Trials, ClinicalTrials.gov, CRISP, PEDro, NIDRR and NRR; and examination of the reference lists of identified studies and other reviews. SELECTION CRITERIA: Only randomised controlled trials (RCT) were included. DATA COLLECTION AND ANALYSIS: Data was abstracted independently by KD and CEH and differences settled by discussion. MAIN RESULTS: Seven trials were identified with 142 patients. All used small numbers of patients and the method of randomisation and concealment of allocation was poor or not statedin all of the trials. These methodological problems could potentially lead to bias from a number of sources. The methods of physiotherapy varied so widely that the data could not be combined. REVIEWER'S CONCLUSIONS: Considering the small number of patients examined, the methodological flaws in many of the studies and the possibility of publication bias, there is insufficient evidence to support or refute the efficacy of any given form of physiotherapy over another in Parkinson's disease. Another Cochrane review, Physiotherapy for patients with Parkinson's Disease, found that there was insufficient evidence to support or refute the efficacy of physiotherapy compared to no physiotherapy in Parkinson's disease. A wide range of physiotherapy approaches were used in these studies and a survey of UK physiotherapists confirmed that they also use an eclectic combination of techniques in the treatment of Parkinson's disease (Plant 1999). Therefore a consensus must be found as to 'best practice' physiotherapy for Parkinson's disease. The efficacy of 'standard' physiotherapy should be proved first before examining variations in physiotherapy methods. Therefore large well designed randomised controlled trials are needed to judge the effect of physiotherapy in Parkinson's disease. After this large RCTs are needed to demonstrate the most effective form of physiotherapy in Parkinson's disease. Outcome measures with particular relevance to patients, carers, physiotherapists and physicians should be chosen and the patients monitored for at least 6 months to determine the duration of any effect. The trials should be reported according to CONSORT guidelines (CONSORT 1996). PMID- 11279766 TI - Non-pharmacological therapies for dysphagia in Parkinson's disease. AB - BACKGROUND: Dysphagia occurs frequently in Parkinson's disease although patients themselves may be unaware of swallowing difficulties. Speech and language therapists in conjunction with nurses and dietitians use techniques that aim to improve swallowing and reduce the risk of choking, aspiration and chest infections. OBJECTIVES: ~Bullet~To compare the efficacy and effectiveness of non pharmacological swallowing therapy for dysphagia versus placebo or no intervention in patients with Parkinson's disease. ~Bullet~To compare one form of non-pharmacological swallowing therapy for dysphagia with another in patients with Parkinson's disease. SEARCH STRATEGY: Relevant trials were identified by electronic searches of MEDLINE, EMBASE, CINAHL, ISI-SCI, AMED, MANTIS, REHABDATA, REHADAT, GEROLIT, Pascal, LILACS, MedCarib, JICST-EPlus, AIM, IMEMR, SIGLE, ISI ISTP, DISSABS, Conference Papers Index, Aslib Index to Theses, the Cochrane Controlled Trials Register, the CentreWatch Clinical Trials listing service, the metaRegister of Controlled Trials, ClinicalTrials.gov, CRISP, PEDro, NIDRR and NRR; and examination of the reference lists of identified studies and other reviews. SELECTION CRITERIA: Only randomised controlled trials (RCT) were included. We did not examine any trials using drugs or surgery to treat the dysphagia. We did not examine any trials where part of the therapist's advice was to insert a nasogastric or percutaneous gastrostomy tube. DATA COLLECTION AND ANALYSIS: Not applicable. MAIN RESULTS: No randomised controlled trials or controlled trials were found that examined the efficacy of non-pharmacological swallowing therapy for the treatment of dysphagia in Parkinson's disease. However there is one large RCT currently recruiting patients that will compare 'chin down' posture with thickened liquids in the treatment of dysphagia. The main outcomes will be the rates of aspiration and pneumonia. REVIEWER'S CONCLUSIONS: There is currently no evidence to support or refute the efficacy of non pharmacological swallowing therapy for dysphagia in Parkinson's disease. Large well designed placebo-controlled RCTs are required to assess the effectiveness of swallowing therapy for dysphagia in Parkinson's disease and reported according to CONSORT guidelines. Suitable outcome measures should be chosen so that the efficacy and effectiveness of non-pharmacological swallowing therapy can be assessed and an economic analysis performed. Outcomes which have meaning to patients and carers should be used wherever possible since they need to know the value of this therapy in practical terms. The patients should be followed for at least 6 months to determine the duration of any improvement. PMID- 11279767 TI - Oral versus intra-vaginal imidazole and triazole anti-fungal treatment of uncomplicated vulvovaginal candidiasis (thrush). AB - BACKGROUND: Anti-fungals are available for oral and intra-vaginal treatment of vulvovaginal candidiasis (thrush). OBJECTIVES: The primary objective of this review was to assess the relative effectiveness of oral versus intra-vaginal anti fungals for the treatment of uncomplicated vulvovaginal candidiasis. The secondary objectives of the review were to assess the cost-effectiveness, safety and patient preference of oral versus intra-vaginal anti-fungals. SEARCH STRATEGY: The following sources were searched: The Cochrane Library (Issue 4, 1999), MEDLINE (January 1985 to May 2000), EMBASE (January 1980 to January 2000) and the Cochrane Collaboration Sexually Transmitted Disease Group Specialised Register of Controlled Trials. The reference lists of retrieved articles were reviewed manually. The manufacturers of anti-fungals available in the UK were contacted. SELECTION CRITERIA: ~Bullet~Randomised controlled trials published in any language. ~Bullet~Trials had to compare at least one oral anti-fungal with one intra-vaginal anti-fungal. ~Bullet~Women (aged 16 years or over) with uncomplicated vulvovaginal candidiasis. ~Bullet~The diagnosis of vulvovaginal candidiasis to be made mycologically (i.e. a positive culture and / or microscopy for yeast). ~Bullet~Trials were excluded if they solely involved subjects who were HIV positive, immunocompromised, pregnant, breastfeeding or diabetic. ~Bullet~The primary outcome measure was clinical cure. DATA COLLECTION AND ANALYSIS: Duplicate scrutiny was performed of the titles and abstracts of the electronic search results. Full article formats of all selected abstracts were retrieved and independently assessed by two reviewers. Independent duplicate abstraction was performed by four reviewers. Disagreements regarding trial inclusion or data abstraction were resolved by discussion between the reviewers. Odds ratios were pooled using the random effects model. Chi-squared tests with a p-value of less than 0.1 indicated heterogeneity in the results. MAIN RESULTS: Seventeen trials are included in the review, reporting 19 oral versus intra vaginal anti-fungal comparisons. No statistically significant differences were shown between oral and intra-vaginal anti-fungal treatment for clinical cure at short term (OR 1.00 (95% CI, 0.72 to 1.40)) and long term (OR 1.03 (95% CI, 0.72 to 1.49)) follow-up. No statistically significant differences for mycological cure were observed between oral and intra-vaginal treatment at short term (OR 1.20(95% CI, 0.87 to 1.65)) or long term follow-up (OR 1.30 (95% CI, 0.99 to 1.71)). Two trials each reported one withdrawal from treatment due to an adverse reaction. Treatment preference data were poorly reported. REVIEWER'S CONCLUSIONS: No differences exist in terms of the relative effectiveness (measured as clinical and mycological cure) of anti-fungals administered by the oral and intra-vaginal routes for the treatment of uncomplicated vaginal candidiasis. No definitive conclusion can be made regarding the relative safety of oral and intra-vaginal anti-fungals for uncomplicated vaginal candidiasis. The oral route of administration is the preferred route for anti-fungals for the treatment of vulvovaginal candidiasis. The decision to prescribe or recommend the purchase of an anti-fungal for oral or intra-vaginal administration should take into consideration: safety, cost and treatment preference. Unless there is a previous history of adverse reaction to one route of administration or contraindications: if women are purchasing their own treatment, they should be given full information about the characteristics and costs of treatment to make their own decision. If health services are paying the treatment cost, decision-makers should consider whether the higher cost of oral anti-fungal administration is worth the gain in convenience, if this is the patient's preference. PMID- 11279768 TI - Lifestyle modification for obstructive sleep apnoea. AB - BACKGROUND: Obstructive sleep apnoeas are due to transient closure of the upper airway during sleep and merge into hypopnoeas in which the airway narrows, but some airflow continues. They are due to the forces compressing the airway overcoming those which stabilise its patency. The commonest association is obesity in which fatty tissue is deposited around the airway. Exercise has been recommended as a method of losing weight, but other techniques which achieve this are also thought to improve symptoms due to sleep apnoeas. Sleep hygiene may alter the sleep structure and the control of the upper airway during sleep and thus promote its patency. OBJECTIVES: The objectives of this review are to determine whether weight loss, sleep hygiene and exercise are effective in the treatment of obstructive sleep apnoeas. SEARCH STRATEGY: The Cochrane Airways Group Trials Register, MEDLINE, EMBASE, CINAHL and reference lists of review articles have been searched. SELECTION CRITERIA: Randomised, single or double blind placebo controlled, either parallel group or crossover design studies of any of these interventions were to have been included. DATA COLLECTION AND ANALYSIS: No completed trials have been identified. MAIN RESULTS: No randomised trial data were available for analysis. REVIEWER'S CONCLUSIONS: There is a need for randomised controlled trials of these commonly used treatments in obstructive sleep apnoeas. These should identify which sub groups of patients with sleep apnoeas benefit most from each type of treatment and they should have clear and standardised outcome measures. PMID- 11279769 TI - Beclomethasone at different doses for chronic asthma (review). AB - BACKGROUND: Beclomethasone dipropionate (BDP) is available in a wide range of daily doses for the treatment of long-term asthma. OBJECTIVES: To assess the evidence for a dose response relationship for BDP in the treatment of long-term asthma. SEARCH STRATEGY: We searched the Cochrane Airways Group trial register, Cochrane Controlled Trials Register (The Cochrane Library issue 1 1999) and references lists of articles. Authors and Glaxo Wellcome UK were contacted to identify eligible studies. We also hand searched the proceeding from relevant respiratory society meetings, the British Journal of Clinical Research and the European Journal of Clinical Research for studies. SELECTION CRITERIA: Prospective, randomised trials comparing two or more daily doses of BDP in patients over the age of two years with long-term asthma. DATA COLLECTION AND ANALYSIS: Trials were selected for inclusion and scored for quality by two reviewers. Data were extracted by one reviewer. Authors were contacted to clarify details of study design and retrieve missing data. MAIN RESULTS: 11 trials involving 1614 subjects were included. Methodological quality was variable. Studies rarely gave a clear indication of the degree of asthma control at baseline. Less than two-fold to five-fold dose differences were assessed by different studies. The results are reported as weighted mean differences (WMD) with 95% confidence limits (95% CI). The number of trials (N) contributing to each outcome is stated. In non-oral steroid treated asthmatics a small advantage of BDP 800 mcg/d over 400 mcg/d was apparent for improvement in morning peak expiratory flow rate (PEFR) compared to baseline, WMD 11 L/min (95% CI 4 to 19 L/min) N=2; improvement in forced expired volume in one second (FEV1) compared to baseline, WMD 9 ml (95% CI 3 to 140) N=1; and reduction in night-time symptom score compared to baseline, WMD 0.13 (95% CI 0.04 to 0.22) N=1. Studies that assessed BDP 1000 v 500 mcg/d and BDP 1600 v 400 mcg/d demonstrated significant advantage of higher dose over lower dose for histamine bronchial hyper responsiveness (BHR) and percentage improvement in FEV1 compared to baseline. No differences between higher and lower daily doses of BDP were apparent for daytime symptoms, withdrawals due to asthma exacerbation, oropharyngeal side effects or measures of hypothalamo-pituitary-adrenal (HPA) function. No difference in prednisolone sparing effect was apparent when comparing high dose and low dose BDP in oral corticosteroid (OCS) dependent patients. REVIEWER'S CONCLUSIONS: BDP appears to demonstrate a shallow dose response effect in long-term asthma for a small number of efficacy outcomes over range of daily doses from 400 mcg/d to 1600 mcg/d, although the clinical significance of the improvements afforded by higher doses is questionable. PMID- 11279770 TI - Carbonic anhydrase inhibitors for hypercapnic ventilatory failure in chronic obstructive pulmonary disease. AB - BACKGROUND: Carbonic anhydrase inhibitors such as acetazolamide cause a mild metabolic acidosis and may stimulate breathing. Some patients with severe chronic obstructive pulmonary disease (COPD) develop chronic hypercapnic ventilatory failure. In theory, they may benefit from use of these drugs with a fall in arterial carbon dioxide level (PCO2) and a rise in arterial oxygen (PO2). OBJECTIVES: To determine the effectiveness and safety of acetazolamide in the treatment of hypercapnic ventilatory failure due to COPD SEARCH STRATEGY: The Cochrane Register of Controlled Clinical Trials was searched along with Medline, Embase, Central and CINAHL for relevant randomised control trials. SELECTION CRITERIA: Trials were included in the review provided they were placebo controlled, carried out in patients with stable chronic ventilatory failure due to COPD. DATA COLLECTION AND ANALYSIS: Data were extracted and analysed by two reviewers (PJ and MG) and agreement was reached by consensus. Where data could be aggregated they were analysed using a fixed efefcts model and reported as a weighted mean difference (WMD) and its associated 95% confidence interval (95% CI). MAIN RESULTS: Four trials were included in the review. Of these, two were randomised parallel studies, one was a crossover study and the other had a sequential design. A total of 84 patients were involved. Study quality was mixed and the studies were short (typically two weeks). All studies showed a similar direction and size of effect. In the randomised parallel studies, acetazolamide caused a metabolic acidosis and produced a non-significant fall in PCO2 (WMD 0.41 kPa; 95% CI -0.91, 0.09; N=2) and a significant rise in PO2 (WMD 1.54 kPa; 95% CI 0.97, 2.11; N=2). One study reported an improvement in sleep but there were no data concerning outcomes such as health status, symptoms, exacerbation rate, hospital admissions or deaths. Side effects were reported infrequently. REVIEWER'S CONCLUSIONS: Acetazolamide can produce a small increase in arterial PO2 and fall in PCO2. These conclusions are drawn from a few small short studies that were not all of high quality. It is not known whether this physiological improvement is associated with clinical benefit. PMID- 11279771 TI - Helium-oxygen mixture for nonintubated acute asthma patients. AB - BACKGROUND: Helium and oxygen mixtures (heliox), have been used sporadically in respiratory medicine for decades. Their use in acute respiratory emergencies such as asthma has been the subject of considerable debate. Despite the lapse of more than 60 years since it was first proposed, the role of heliox in treating patients with acute severe asthma is unclear. OBJECTIVES: To determine the effect of the addition of heliox to standard medical care on the course of acute asthma, as measured by pulmonary function testing and clinical endpoints. SEARCH STRATEGY: Randomized controlled trials were identified from the Cochrane Airways Group Asthma Register which is a compilation of systematic searches of CINAHL, EMBASE, MEDLINE, and CENTRAL and hand searching of the 20 most productive respiratory care journals. In addition, primary authors and experts were contacted to identify eligible studies. References from included studies, known reviews and texts were also searched. SELECTION CRITERIA: Inclusion criteria were: 1) randomized, single or double blind, controlled trials; 2) children or adults with a clinical diagnosis of acute asthma seen in emergency departments or equivalent acute care settings; 3) compared treatment with inhaled heliox compared with a control (oxygen or air). Two reviewers independently assessed the studies for inclusion and quality assessment; disagreement was resolved by a third reviewer and consensus. DATA COLLECTION AND ANALYSIS: Data from all included trials were combined using weighted mean differences (WMD), with 95% confidence intervals (95% CI) in a random effects model. Homogeneity of effect sizes were tested with the Dersimonian and Laird method with p<0.1 as the cut point for significance. Sensitivity analyses were performed on age (adults vs. children), different helium-oxygen mixtures and methodological quality. MAIN RESULTS: A total of 4 randomized controlled trials were selected for inclusion with a total of 288 acute asthma patients. Three studies involved adults and one study dealt solely with children. Two were of low quality. The main outcome variable was spirometric measurements (PFT % predicted) in all trials. Pooling the four trials showed no significant differences (WMD = -1.61; 95% CI: -6.64 to 3.41). All pulmonary function tests were recorded during heliox administration (15 to 60 min). There was no evidence of heterogeneity between studies. There were no significant differences between groups when adults vs. children, high vs. low quality, and high vs. low heliox dose studies were compared. The findings were the with a fixed effects model. REVIEWER'S CONCLUSIONS: The existing evidence does not provide support for the administration of helium-oxygen mixtures to patients presenting to the emergency department with moderate to severe acute asthma. Heliox treatment does not have a role in the initial treatment of patients with acute asthma. These conclusions are based upon between group comparisons and small studies. Additional research in this setting may be warranted. PMID- 11279772 TI - Computerised advice on drug dosage to improve prescribing practice. AB - BACKGROUND: Maintaining therapeutic concentrations of toxic drugs is a complex task. Several computer systems have been designed to help doctors determine optimum drug dosage. Significant improvements in health could be achieved if computer advice was shown to be beneficial. OBJECTIVES: To assess whether computer support for drug dosage benefits patients and hence whether it should be more widely available. SEARCH STRATEGY: We searched the Cochrane Effective Practice and Organisation of Care Group specialised register (June 1996), MEDLINE (1966 to June 1996), EMBASE (1980 to June 1996), hand searched the journal Therapeutic Drug Monitoring (1979 to June 1996), reference lists of articles and contacted experts in the field. SELECTION CRITERIA: Randomised trials, interrupted time series and controlled before and after studies of computerised advice on drug dosage. The participants were health professionals responsible for patient care. The outcomes were: any objectively measured change in the behaviour of the health care provider (such as changes in the dose of drug used); any change in the health of patients, resulting from computer support (such as adverse reactions to drugs). DATA COLLECTION AND ANALYSIS: Two reviewers independently extracted data and assessed study quality. MAIN RESULTS: Fifteen trials involving 1229 patients were included. The drugs studied were theophylline, warfarin, heparin, aminoglycosides, nitroprusside, lignocaine, oxytocin, fentanyl and midazolam. Interventions usually targeted doctors although some studies attempted to influence prescribing by pharmacists and nurses. All included studies took place on acute medical conditions in hospital settings. Although all studies used reliable outcome measures, sample size was often small and only two studies reported a sample size calculation. Computer support for drug dosage gave significant benefits reducing: 1. The time to achieve therapeutic control (standardised mean difference -0.44, 95% CI -0.70 to -0.17); 2. Toxic drug levels (risk difference -0.12, 95% CI -0.24 to -0.01); 3. Adverse reactions (risk difference -0.06, 95% CI -0.12 to 0.00); 4. Length of hospital stay (standardised mean difference -0.32, 95% CI -0.60 to -0.04). There was a tendency for computer support to result in higher doses of drugs, although this did not reach statistical significance. REVIEWER'S CONCLUSIONS: This systematic review provides evidence to support the use of computer assistance in determining drug dosage. Further clinical trials are necessary to determine whether the benefits seen in specialist applications can be realised in general use. PMID- 11279773 TI - Intra-operative mitomycin C for glaucoma surgery. AB - BACKGROUND: Trabeculectomy is performed as a treatment for many types of glaucoma in an attempt to lower the intra-ocular pressure. Mitomycin C is an antimetabolite applied between the sclera and conjunctiva during the initial stages of a trabeculectomy to prevent excessive post-operative scarring and thus reduce the risk of failure. OBJECTIVES: The objective of this review is to assess the effects of intra-operative application of mitomycin C in eyes of people undergoing trabeculectomy. SEARCH STRATEGY: We searched the Cochrane Eyes and Vision Group specialised register, The Cochrane Controlled Trials Register - CENTRAL, MEDLINE, EMBASE and the reference lists of relevant articles. We used the Science Citation Index to search for articles that cited the included studies. We contacted investigators and experts for details of additional relevant trials. SELECTION CRITERIA: We included randomised trials of intra operative mitomycin C compared to placebo in trabeculectomy. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality and extracted data. We contacted trial investigators for missing information. Data were summarised using relative risk, odds ratio and weighted mean difference. MAIN RESULTS: This review includes 11 trials involving a total of 698 participants. The trials included three types of participants (those at high risk of failure, those undergoing trabeculectomy combined with cataract surgery, and those with no previous surgical intervention). Mitomycin C appears to be effective in reducing the relative risk of failure of trabeculectomy both in eyes at high risk of failure (relative risk 0.32, 95% confidence interval 0.20 to 0.53) and those undergoing surgery for the first time (relative risk 0.29, 95% confidence interval 0.16 to 0.53). No significant effect on failure was noted in the group undergoing trabeculectomy combined with cataract extraction. Mean intra-ocular pressure was significantly reduced at 12 months in all three participant groups receiving mitomycin C compared to placebo. No significant increase in permanent sight threatening complications was detected. Some evidence exists that mitomycin C increases the risk of cataract. The quality of trial reporting is poor in eight trials. Repeat analysis with three trials rated as low risk of bias did not yield different results. REVIEWER'S CONCLUSIONS: Intra-operative mitomycin C reduces the risk of surgical failure in eyes that have undergone no previous surgery and in eyes at high risk of failure. Compared to placebo it reduces mean intra-ocular pressure at 12 months in all groups of participants in this review. Apart from an increase in cataract formation following mitomycin C, no demonstrable significant increase in other side effects was detected. PMID- 11279774 TI - Interventions for herpes simplex virus epithelial keratitis. AB - BACKGROUND: Many clinical trials have been performed on the acute treatment of dendritic epithelial keratitis. Surveys of antiviral pharmacology and of herpes simplex virus eye disease have evaluated different commercially available agents, but a systematic review of all comparative clinical studies has not previously been undertaken. OBJECTIVES: The objective of this review is to compare the effects of various treatments for dendritic or geographic herpes simplex virus epithelial keratitis. SEARCH STRATEGY: Sources searched for relevant studies were the Cochrane Eyes and Vision Group specialized register, The Cochrane Controlled Trials Register - CENTRAL, MEDLINE, EMBASE, Index Medicus, Excerpta Medica Ophthalmology, reference lists of primary reports, review articles, and corneal textbooks and conference proceedings pertaining to ocular virology. SELECTION CRITERIA: This review includes comparative clinical trials that assessed oral or topical ophthalmic antiviral agents, or physical or chemical debridement in people with active epithelial keratitis. DATA COLLECTION AND ANALYSIS: The reviewer extracted data and assessed trial quality. Interventions were compared by the proportions of participants healed at seven days and at fourteen days after trial enrollment. MAIN RESULTS: This review includes data from 96 trials which randomised a total of 4991 participants. Compared to idoxuridine, the topical application of vidarabine, trifluridine, or acyclovir generally resulted in a significantly greater proportion of participants healing within one week of treatment. Among these three antiviral agents, no treatment emerged as significantly better for the therapy of dendritic epithelial keratitis. Insufficient placebo-controlled studies were available to assess debridement and other physical and physicochemical methods of treatment. Interferon monotherapy had a slight beneficial effect on dendritic epithelial keratitis, but not better than other antiviral agents, and was useful with debridement. REVIEWER'S CONCLUSIONS: Currently available and investigational antiviral agents are effective and nearly equivalent, but the combination of an antiviral nucleoside and interferon seems to speed healing. Future trials of the acute treatment of herpes simplex virus epithelial keratitis must aim to achieve adequate statistical power for assessing the primary outcome and should consider the effect of lesion size and other characteristics on treatment response. PMID- 11279775 TI - Digitalis for treatment of congestive heart failure in patients in sinus rhythm. AB - BACKGROUND: Digitalis glycosides have been in clinical use in the treatment of congestive heart failure for more than 200 years. In recent years several trials have been conducted to address concerns about efficacy and toxicity. Although a systematic review of the literature was recently published, an update is required to include more current trials. OBJECTIVES: To examine the effectiveness of digitalis glycosides in treating congestive heart failure in patients with normal sinus rhythm. To examine the effect of digitalis in patients taking diuretics, ACE inhibitors, and beta blockers; patients with varying severity and duration of disease; patients with prior exposure to digitalis vs. no prior exposure; and patients with diastolic vs. systolic dysfunction. SEARCH STRATEGY: Electronic databases were searched between 1966 and 2000. Dissertation Abstracts and annual meeting abstracts of the American Heart Association, American College of Cardiology, and European Society of Cardiology were searched from 1996-2000. In addition, reference lists provided by the pharmaceutical industry (Glaxo Wellcome Inc.) were searched. SELECTION CRITERIA: Included were randomized placebo controlled trials of 20 or more adult patients of either sex with symptomatic congestive heart failure who were studied for seven weeks or more. Excluded were trials in which the prevalence of atrial fibrillation was 2% or greater, or in which any arrhythmia that might compromise cardiac function or any potentially reversible cause of heart failure such as acute ischemic heart disease or myocarditis was present. DATA COLLECTION AND ANALYSIS: Articles selected from the searches described above were reviewed by one of the coauthors, and validated by staff from the central office of the Heart Collaborative Review Group in Bristol, UK. MAIN RESULTS: Eleven articles meeting the defined criteria were identified, and major endpoints of mortality, hospitalization, and clinical status, based respectively upon on 8, 4, and 10 of these selected studies, were recorded and analyzed. The data show that there is no difference in mortality between treatment and control groups, whereas digitalis therapy is associated with a lower rate of hospitalization and of clinical deterioration. REVIEWER'S CONCLUSIONS: The literature indicates that digitalis has a useful role in the treatment of patients with congestive heart failure who are in normal sinus rhythm. PMID- 11279776 TI - Alarm interventions for nocturnal enuresis in children. AB - BACKGROUND: Enuresis (bedwetting) is a socially disruptive and stressful condition which affects around 15-20% of five year olds, and up to 2% of young adults. Although there is a high rate of spontaneous remission, the social, emotional and psychological costs to the children can be great. OBJECTIVES: To assess the effects of alarm interventions on nocturnal enuresis in children, and to compare alarms with other interventions. SEARCH STRATEGY: The following electronic databases were searched: MEDLINE to June 1997; AMED; ASSIA; BIDS; BIOSIS Previews (1985-1996); CINAHL; DHSS Data; EMBASE (1974 to June 1997); PsycLIT and SIGLE. Organisations, manufacturers, researchers and health professionals concerned with enuresis were contacted for information. The reference sections of obtained studies were also checked for further trials. Date of the most recent search: July 1997. SELECTION CRITERIA: All randomised trials of alarm interventions for nocturnal enuresis in children were included in the review. Trials were eligible for inclusion if: children were randomised to alarm treatment compared with controls, other behavioural methods or drugs for nocturnal bedwetting; participants with organic causes for their bedwetting were excluded; and baseline assessments of the level of bedwetting were reported. Trials focused solely on daytime wetting were excluded. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed the quality of the eligible trials, and extracted data. MAIN RESULTS: Twenty two randomised trials met the inclusion criteria, involving 1125 children who received treatment with alarms. The quality of many of the trials was poor, and many comparisons were addressed only by single trials. Children treated with alarms were significantly more likely than untreated controls to become dry during treatment (RR for failing to achieve 14 dry nights 0.27, 95% CI 0.19 to 0.39) and failing to remain dry (RR 0.58, 95%CI 0.46 to 0.74). There was insufficient evidence to judge whether one type of alarm is better than another and whether alarms alone were as good as or better than other behavioural interventions alone or as a supplement to alarm treatment. Desmopressin or tricyclics seem as effective as alarms while on treatment. There was limited evidence to suggest that the relapse rate might be lower after stopping alarm treatment than after desmopressin (RR 0.11, 95% CI 0.02 to 0.78). REVIEWER'S CONCLUSIONS: Alarm interventions are an effective treatment for nocturnal bedwetting in children. Desmopressin and tricyclics appeared as effective while on treatment, but this effect was not sustained after treatment stopped, and alarms may be more effective in the long term. Comparisons between drug and behavioural treatments are needed, and should include relapse rates after treatment is finished. PMID- 11279778 TI - Electromagnetic therapy for the treatment of pressure sores. AB - BACKGROUND: Electromagnetic therapy is used with the aim of improving the healing of chronic wounds such as pressure sores and venous leg ulcers OBJECTIVES: To assess the effectiveness of electromagnetic therapy in the treatment of pressure sores SEARCH STRATEGY: The Cochrane Wounds Group search strategy was used (see Scope) to search for randomised controlled trials (RCTs) of electromagnetic therapy for the treatment of pressure sores SELECTION CRITERIA: Randomised controlled trials comparing electromagnetic therapy with sham electromagnetic therapy, or other (standard) treatment DATA COLLECTION AND ANALYSIS: Results of searches were scrutinised by one reviewer (and checked by a second) to identify possible RCTs and full reports of these were obtained. Details of eligible studies were extracted and summarised using a data extraction sheet. Attempts were made to obtain missing data by contacting authors. Data extraction was checked by a second reviewer. MAIN RESULTS: A total of two eligible RCTs were identified for inclusion in this review. The first of these studies (Comorosan 1993) was a three armed study comparing electromagnetic therapy, electromagnetic therapy in combination with standard therapy, and standard therapy alone. The second study (Salzburg 1995) was a comparison between electromagnetic therapy and sham therapy on 30 male patients with a spinal cord injury and a grade two or grade three pressure sore. Neither study found a statistically significant difference between the healing rates of electromagnetic therapy treated and control group patients. REVIEWER'S CONCLUSIONS: The results suggest no evidence of a benefit in using electromagnetic therapy to treat pressure sores. However the possibility of a beneficial or harmful effect cannot be ruled out due to the fact there were only two trials with methodological limitations and small numbers of patients. PMID- 11279777 TI - Budesonide for maintenance of remission in Crohn's disease. AB - OBJECTIVES: The primary objective was to precisely derive an estimate of the efficacy of oral budesonide for the maintenance of remission in Crohn's disease. SEARCH STRATEGY: Medline 1966-September 2000 was searched using the text and key words "oral budesonide", "Crohn's disease", "Crohn disease", and "inflammatory bowel disease". Proceedings from the American Gastroenterology Association conference (1980-1998) were hand searched. Additionally, the Cochrane Controlled Trials Register and the Inflammatory Bowel Disease Review Group Trials Registers were also searched. The manufacturer of oral budesonide was also contacted, as were some of the local trialists involved in the oral budesonide trials. Relevant articles were retrieved, and their reference lists were also reviewed. SELECTION CRITERIA: Randomized controlled trials of oral budesonide in which patients at entry were in remission with a Crohn's Disease Activity Index (CDAI) <= 150, and had disease restricted to the ileum and colon. DATA COLLECTION AND ANALYSIS: The primary outcome was the relative risk (RR) of relapse (and 95% confidence interval [CI]) during the 12 months of treatment, as defined by the number of patients who relapsed, and the number of patients who entered the trial. The numbers needed to treat were also derived. MAIN RESULTS: Three randomized controlled trials of oral budesonide (controlled ileal release preparation) 6 mg/day, 3mg/day and placebo for a one year period were included in the analysis. Budesonide 6 mg/day was not effective at preventing relapse of Crohn's disease over the 12 months of treatment. The relative risk of relapse was 0.89 (95% CI: 0.71-1.13) comparing budesonide 6 mg/day and placebo. Similar results were also observed in the comparison of budesonide 6 mg/day with budesonide 3 mg/day; the relative risk of relapse was 0.89 (95% CI: 0.70-1.11). The 3 mg/day dose was similarly found to be ineffective at preventing relapse during the 12 months of treatment; the relative risk of relapse was 1.00 (95% CI: 0.80-1.24). The results of the analysis of relative risk of relapse were supported by the analysis of withdrawal due to treatment failure. The relative risks for withdrawal due to treatment failure for budesonide 6 mg/day compared with placebo was 0.85 (95% CI: 0.65-1.10), for budesonide 3 mg/day compared with placebo was 0.94 (95% CI: 0.73 1.20), and for budesonide 6 mg/day compared with budesonide 3 mg/day was 0.89 (95% CI: 0.68-1.17). REVIEWER'S CONCLUSIONS: Oral budesonide therapy at 6 mg/day is not effective in preventing relapses of Crohn's disease. PMID- 11279779 TI - Electromagnetic therapy for the treatment of venous leg ulcers. AB - BACKGROUND: Electromagnetic therapy is used with the aim of improving the healing of chronic wounds such as pressure sores and venous leg ulcers OBJECTIVES: To assess the effectiveness of electromagnetic therapy in the treatment of venous leg ulcers SEARCH STRATEGY: The Cochrane Wounds group search strategy was used (see Scope) to search for randomised controlled trials (RCTs) of electromagnetic therapy for the treatment of venous leg ulcers SELECTION CRITERIA: Randomised controlled trials comparing electromagnetic therapy with sham electromagnetic therapy or other (standard) treatment DATA COLLECTION AND ANALYSIS: Results of searches were scrutinised by one reviewer (and checked by a second) to identify possible RCTs and full reports of these were obtained. Details of eligible studies were extracted and summarised using a data extraction sheet. Attempts were made to obtain missing data by contacting authors. Data extraction was checked by a second reviewer. MAIN RESULTS: A total of three eligible RCTs were identified. Two trials compared the use of electromagnetic therapy to sham (Ieran 1990; Kenkre 1996) and one trial (Stiller 1992) compared it with standard topical treatments. One of the trials found a difference in healing rates of borderline statistical significance between electromagnetic therapy and sham, although the direction of treatment effect was consistently in favour of electromagnetic therapy the difference was not statistically significant. REVIEWER'S CONCLUSIONS: There is currently no reliable evidence of benefit of electromagnetic therapy in the healing of venous leg ulcers. PMID- 11279780 TI - Radical radiotherapy for stage I/II non-small cell lung cancer in patients not sufficiently fit for or declining surgery (medically inoperable). AB - BACKGROUND: In general, surgery is believed to offer the best prospects for cure for early stage non-small cell lung cancer (NSCLC). In spite of the intention to consider all patients with stage I-II disease for surgery, there are those who, although technically operable, either refuse surgery or are considered inoperable because of insufficient respiratory reserve, cardiovascular disease or general frailty. This group may therefore be considered "medically inoperable". Some respiratory physicians refer these patients for radical radiotherapy whilst others believe that radiotherapy has little to offer and adopt a watch policy, referring patients for palliative radiotherapy only when they become symptomatic. Although there is little evidence from randomised trials to support the use of radical radiotherapy for stage I/II NSCLC, it is the perception of most clinical oncologists (radiotherapists) that patients should receive radical, as opposed to palliative, treatment (COIN 1999). OBJECTIVES: To determine the effectiveness and the morbidity of radical radiotherapy for medically inoperable NSCLC. SEARCH STRATEGY: Randomised trials were sought by electronic searching the Cochrane Clinical Trials Register and both randomised and non-randomised trials sought by searching Medline and Excerpta Medica (Embase). Further studies were identified from references cited in those papers already identified by electronic searching. SELECTION CRITERIA: Studies of patients of any age with stage I/II NSCLC receiving radiotherapy at a dose greater than 40Gy in 20 fractions over four weeks or its radiobiological equivalent. DATA COLLECTION AND ANALYSIS: Two randomised and thirty-five non-randomised studies were identified. One randomised and nine non-randomised studies did not meet the selection criteria and were not included in the review. MAIN RESULTS: In the randomised trial comparing two radiotherapy schedules, two-year survival was superior following continuous hyperfractionated accelerated radiotherapy (CHART; 37%) compared to 60Gy in 30 fractions over six weeks (24%). There were 26 non-randomised retrospective studies including an estimated 2003 patients, in which overall survival results varied between 33-72% at two years, 17-55% at three years and 0-42% at five years. The proportion of deaths not due to cancer was 11-43%. Cancer-specific survival was between 54-93% at two years, 22-56% at three years and 13-39% at five years. Complete response rates were 33-61% and local failure rates between 6 70%. Distant metastases developed in approximately 25% of patients. Better response rates and survival were seen in those with smaller tumours and in those receiving higher doses though the reasons for prescribing higher doses were not clearly stated. Worse outcome was seen in those with prior weight loss or poor performance status. Assessment of treatment-related morbidity and effects on quality of life and symptom control were inconclusive because of the lack of prospective evaluation and paucity of data. REVIEWER'S CONCLUSIONS: There were no randomised trials that compared a policy of immediate radical radiotherapy with palliative radiotherapy given when patients develop symptoms. In the absence of such trials, radical radiotherapy appears to result in a better survival than might be expected had treatment not been given. A substantial, though variable, proportion of patients died during follow-up from causes other than cancer. The optimal radiation dose and treatment technique (particularly with respect to mediastinal irradiation) remain uncertain. PMID- 11279781 TI - Strategies for increasing women participation in community breast cancer screening. AB - BACKGROUND: Strategies for reducing breast cancer mortality in western countries have focused on screening, at least for women aged 50 to 69 years. One of the requirements of any community screening program is to achieve a high participation rate, which is related to methods of invitation. Therefore, it was decided to systematically review the scientific evidence on the different strategies aimed at improving women's participation in breast cancer screening programs and activities. OBJECTIVES: To assess the effectiveness of different strategies for increasing the participation rate of women invited to community (population-based) breast cancer screening activities or mammography programs. SEARCH STRATEGY: MEDLINE (1966-2000), CENTRAL (2000), and EMBASE (1998-1999) searches for 1966 to 1999 were supplemented by reports and letters to the European Screening Breast Cancer Programs (Euref Network). SELECTION CRITERIA: Both published and unpublished trials were eligible for inclusion, provided the women had been invited to a community breast screening activity or program and had been randomised to an intervention group or a control group with no active intervention. DATA COLLECTION AND ANALYSIS: We identified 151 articles, which were reviewed independently by two people. The discrepancies were resolved by a third reviewer in order to reach consensus. Thirty-four studies were excluded because they lacked a control group; 58 of the other 117 articles were considered as opportunistic and not community-based; 59 articles, which reported 70 community-based randomised controlled trials or clinical controlled trials, were accepted. In 24 of these, the control group had not been exposed to any active intervention, but 8 of the 24 had to be excluded because the denominator for estimating attendance was unknown. At the end, 16 studies constituted the material for this review, although two studies were further excluded because their groups were not comparable at baseline. Data from all but one study were based on or converted to an intention-to-treat analysis. Attendance in response to the mammogram invitation was the main outcome measure. MAIN RESULTS: The evidence favoured five active strategies for inviting women into community breast cancer screening services: letter of invitation (OR 1.66, 95% CI 1.43 to 1.92), mailed educational material (OR 2.81, 95% CI 1.96 to 4.02), letter of invitation plus phone call (OR 2.53, 95% CI 2.02 to 3.18), phone call (OR 1.94, 95% CI 1.70 to 2.23), and training activities plus direct reminders for the women (OR 2.46, 95% CI 1.72 to 3.50). Home visits did not prove to be effective (OR 1.06, 95 % CI 0.80 to 1.40) and letters of invitation to multiple examinations plus educational material favoured the control group (OR 0.62, 95 % CI 0.32 to 1.20). REVIEWER'S CONCLUSIONS: Most active recruitment strategies for breast cancer screening programs examined in this review were more effective than no intervention. Combinations of effective interventions can have an important effect. Some costly strategies, as a home visit and a letter of invitation to multiple screening examinations plus educational material, were not effective. Further reviews comparing the effective interventions and studies that include cost effectiveness, women's satisfaction and equity issues are needed. PMID- 11279782 TI - Glucosamine therapy for treating osteoarthritis. AB - BACKGROUND: Osteoarthritis (OA) is the most common form of arthritis, and it is often associated with significant disability and an impaired quality of life. OBJECTIVES: To review all randomized controlled trials (RCTs) evaluating the effectiveness and toxicity of glucosamine in osteoarthritis (OA). SEARCH STRATEGY: We searched MEDLINE, Embase, and Current Contents up to November 1999, and the Cochrane Controlled Trials Register. We also wrote letters to content experts, and hand searched reference lists of identified RCTs and pertinent review articles. SELECTION CRITERIA: Relevant studies met the following criteria: 1) RCTs evaluating the effectiveness and safety of glucosamine in OA, 2) Both placebo based and comparative studies were eligible, 3) Both single blinded and double-blinded studies were eligible. DATA COLLECTION AND ANALYSIS: Data abstraction was performed independently by two investigators and the results were compared for degree of agreement. Gotzsche's method and a validated tool (Jadad 1995) were used to score the quality of the RCTs. Continuous outcome measures were pooled using standardized mean differences. Dichotomous outcome measures were pooled using Peto Odds Ratios. MAIN RESULTS: Collectively, the 16 identified RCTs provided evidence that glucosamine is both effective and safe in OA. In the 13 RCTs in which glucosamine was compared to placebo, glucosamine was found to be superior in all RCTs, except one. In the four RCTs in which glucosamine was compared to an NSAID, glucosamine was superior in two, and equivalent in two. REVIEWER'S CONCLUSIONS: Further research is necessary to confirm the long term effectiveness and toxicity of glucosamine therapy in OA. Most of the trials reviewed only evaluated the Rotta preparation of glucosamine sulfate. It is not known whether different glucosamine preparations prepared by different manufacturers are equally effective in the therapy of OA. PMID- 11279783 TI - Herbal therapy for treating osteoarthritis. AB - BACKGROUND: The increasing popularity of complementary and alternative medicine appears to be particularly evident amongst people with chronic disease. In the treatment of osteoarthritis, one therapy that has been identified as having potential benefit, is herbal medicine (phytotherapy). OBJECTIVES: To determine the effectiveness of herbal therapies in treating osteoarthritis. SEARCH STRATEGY: Databases for mainstream and complementary medicine were searched using terms to include all forms of arthritis combined with herbal medicine. We searched the following electronic databases: MEDLINE, EMBASE, CISCOM, AMED, CINAHL, Cochrane Controlled Trials Register (CCTR), Cochrane Musculoskeletal specialized register, Dissertation Abstracts, BIDS ISI and the Cochrane Complementary Medicine Fields Specialized Register. We also searched the reference lists from retrieved trials. SELECTION CRITERIA: All randomized trials of herbal interventions in osteoarthritis, compared to placebo. Studies were included according to the a priori protocol and agreement between two reviewers who independently read each selected paper for content and for assessment of quality. Papers of any language were included. DATA COLLECTION AND ANALYSIS: Data were extracted independently by the same two reviewers. MAIN RESULTS: Five studies (four different herbal interventions) met the review criteria. Two studies were suitable for data pooling. It was not possible to draw firm conclusions from the single studies but two studies of avocado/soybean unsaponifiables showed beneficial effects on functional index, pain, intake of NSAIDs and global evaluation. No serious side effects were reported. REVIEWER'S CONCLUSIONS: The evidence for avocado-soybean unsaponifiables in the treatment of osteoarthritis is convincing but evidence for the other herbal interventions is insufficient to either recommend or discourage their use. PMID- 11279784 TI - Herbal therapy for treating rheumatoid arthritis. AB - BACKGROUND: The increasing popularity of the use of complementary and alternative interventions or treatments appears to be particularly evident amongst people with chronic disease. In the treatment of rheumatoid arthritis, one therapy that has been identified as having potential benefit, is herbal medicine (phytotherapy). OBJECTIVES: To assess the effectiveness of herbal therapies in the treatment of rheumatoid arthritis. SEARCH STRATEGY: We developed a search strategy using terms to include all forms of arthritis combined with herbal medicine. We searched the following electronic databases from 1966 to 2000: MEDLINE, EMBASE, CISCOM, AMED, CINAHL, Cochrane Controlled Trials Register (CCTR), Cochrane Musculoskeletal specialized register, Dissertation Abstracts, BIDS ISI and the Cochrane Complementary Medicine Fields Specialized Register. This was supplemented by searching the reference lists from retrieved trials. SELECTION CRITERIA: All randomized trials of herbal interventions in rheumatoid arthritis, compared to placebo. Two reviewers independently read and selected each potential study according to the criteria published in an a priori protocol. Papers of any language were included. DATA COLLECTION AND ANALYSIS: Data were extracted independently by the same two reviewers and an assessment of methodological quality was conducted. MAIN RESULTS: Eleven studies met the inclusion criteria. Seven of the studies compared gamma-linolenic acid (GLA) to placebo although three of these were not suitable for data pooling. The remaining studies considered four different herbal interventions and were assessed individually. All of the GLA studies found some improvement in clinical outcomes but methodology and study quality was variable, making it difficult to draw conclusive results. However, the better quality studies suggest potential relief of pain, morning stiffness and joint tenderness. With the exception of one intervention (Tripterygium wilfordii hook F), no serious side effects were reported. REVIEWER'S CONCLUSIONS: There appears to be some potential benefit for the use of GLA in rheumatoid arthritis although further studies are required to establish optimum dosage and duration of treatment. The single studies are inconclusive. PMID- 11279785 TI - Drugs for discoid lupus erythematosus. AB - BACKGROUND: Discoid lupus erythematosus is a chronic form of cutaneous (skin) lupus which can cause permanent scarring if treatment is inadequate. Many drugs have been used to treat this disease and some of these are potentially very toxic. OBJECTIVES: To assess the effects of drugs for discoid lupus erythematosus. SEARCH STRATEGY: We searched the Cochrane Clinical Trials Register (December 1999), MEDLINE (January 1966 to December 1999), EMBASE (January 1980 to January 2000), and the reference lists of relevant reviews. Index Medicus (1956 to 1966) was handsearched and 7 experts in the field were approached for information about unpublished trials. SELECTION CRITERIA: Randomised trials of drugs to treat people with discoid lupus erythematosus. Drugs included in the search were azathioprine, chloroquine, clofazimine, corticosteroids, (oral and topical), dapsone, gold, interferon alpha-2a, methotrexate, phenytoin, retinoids, sulphasalazine and thalidomide. DATA COLLECTION AND ANALYSIS: Two reviewers independently examined each retrieved study for eligibility. MAIN RESULTS: Two trials involving 136 participants were included. In a cross-over study of twelve weeks duration fluocinonide 0.05% cream (a potent topical corticosteroid), appeared to be markedly better than hydrocortisone 1% cream ( a mild corticosteroid). Clearing or excellent improvement was seen in 27% of people using fluocinonide and in 10% of those using hydrocortisone, giving a 17% absolute benefit in favour of fluocinonide (95% CI 4.5 to 29.5% and NNT 6). In the second trial, hydroxychloroquine was compared with acitretin in 58 people. There was marked improvement or clearing in 46% of people using acitretin and in 50% of those on hydroxychloroquine, a nonsignificant 4% absolute gain with hydroxychloroquine (95%CI -23% to 30%). The adverse effects were more frequent and more severe in the acitretin group. REVIEWER'S CONCLUSIONS: Fluocinonide cream may be more effective than hydrocortisone in treating people with discoid lupus erythematosus. Hydroxychloroquine and acitretin appear to be of equal efficacy, although adverse effects are more frequent and more severe with acitretin. There is not enough reliable evidence about other drugs used to treat discoid lupus erythematosus. PMID- 11279786 TI - Magnesium sulphate versus lytic cocktail for eclampsia. AB - BACKGROUND: Eclampsia, the occurrence of a seizure in association with pre eclampsia, is a rare but serious complication of pregnancy. A number of different anticonvulsants are used to control eclamptic fits and to prevent further seizures. OBJECTIVES: The aim of this review was to compare the effects of magnesium sulphate with those of lytic cocktail when used for the care of women with eclampsia. SEARCH STRATEGY: The register of trials held by the Cochrane Pregnancy and Childbirth Group was searched for relevant trials. The Cochrane Controlled Trials Register in The Cochrane Library Issue 2, 2000 was also searched. SELECTION CRITERIA: Randomised trials recruiting women with eclampsia, and comparing any use of magnesium sulphate with any use of lytic cocktail. DATA COLLECTION AND ANALYSIS: Data were extracted from each report without any blinding of the results or of the treatments which women received. MAIN RESULTS: Two trials with 199 women were included in the review. These were both small and of average quality. Magnesium sulphate was better than lytic cocktail at preventing further fits (relative risk (RR) 0.09, 95% confidence interval (CI) 0.03-0.24; risk difference (RD) 0.43, 95% CI -0.53, -0.34; number needed to treat (NNT) 3, 95% CI 2-3) and was associated with less respiratory depression (RR 0.12, 95% CI 0.02-0.91). Magnesium sulphate was also associated with fewer maternal deaths than lytic cocktail, but the difference was not statistically significant (RR 0.25, 95% CI 0.04-1.43). REVIEWER'S CONCLUSIONS: Magnesium sulphate is the anticonvulsant of choice for women with eclampsia. Lytic cocktail should be abandoned. PMID- 11279787 TI - Acupuncture for induction of labour. AB - BACKGROUND: This is one of a series of reviews of methods of cervical ripening and labour induction using standardised methodology. The use of complementary therapies is rising and some women look to complementary therapies during pregnancy and childbirth to be used alongside conventional medical practice. Acupuncture involves the insertion of very fine needles into specific points of the body. The limited observational studies to date suggest acupuncture for induction of labour appears safe, has no known teratogenic effects, and may be effective. The evidence regarding the clinical effectiveness of this technique is limited. OBJECTIVES: To determine the effects of acupuncture for third trimester cervical ripening or induction of labour. SEARCH STRATEGY: The Cochrane Pregnancy and Childbirth Group trials register, the Cochrane Controlled Trials Register and bibliographies of relevant papers. SELECTION CRITERIA: The criteria for inclusion included the following: (1) clinical trials comparing acupuncture used for third trimester cervical ripening or labour induction with placebo/no treatment or other methods listed above it on a predefined list of labour induction methods; (2) random allocation to the treatment or control group; (3) adequate allocation concealment; (4) violations of allocated management not sufficient to materially affect conclusions; (5) clinically meaningful outcome measures reported; (6) data available for analysis according to the random allocation; (7) missing data insufficient to materially affect the conclusions. DATA COLLECTION AND ANALYSIS: A strategy has been developed to deal with the large volume and complexity of trial data relating to labour induction. This involves a two-stage method of data extraction. The initial data extraction is done centrally, and incorporated into a series of primary reviews arranged by methods of induction of labour, following a standardised methodology. The data will then be extracted from the primary reviews into a series of secondary reviews, arranged by category of woman. To avoid duplication of data in the primary reviews, the labour induction methods have been listed in a specific order, from one to 25. Each primary review includes comparisons between one of the methods (from two to 25) with only those methods above it on the list. MAIN RESULTS: No trials met the inclusion criteria for the systematic review. REVIEWER'S CONCLUSIONS: There is a need for a well designed randomised controlled trial to evaluate the role of acupuncture to induce labour. PMID- 11279788 TI - Fetal vibroacoustic stimulation for facilitation of tests of fetal wellbeing. AB - BACKGROUND: Acoustic stimulation of the fetus has been suggested to improve the efficiency of antepartum fetal heart rate testing. OBJECTIVES: The objective of this review was to assess the merits or adverse effects of the use of fetal vibroacoustic stimulation in conjunction with tests of fetal wellbeing. SEARCH STRATEGY: We searched the Cochrane Pregnancy and Childbirth Group trials register. Date of last search: October 2000. SELECTION CRITERIA: All published and unpublished randomized controlled trials assessing the merits of the use of fetal vibroacoustic stimulation in conjunction with tests of fetal wellbeing. DATA COLLECTION AND ANALYSIS: Both reviewers independently extracted data and assessed trial quality. Authors of published and unpublished trials were contacted for further information. MAIN RESULTS: A total of seven trials with a total of 4325 participants were included. Fetal vibroacoustic stimulation reduced the incidence of non-reactive antenatal cardiotocography test (odds ratio (OR) 0.61, 95% confidence interval (CI) 0.49-0.75) and reduced the overall mean cardiotocography testing time (weighted mean difference (WMD) -4.55 minutes, 95% CI -5.96 minutes to -3.14 minutes). Vibroacoustic stimulation evoked more than mock stimulation when used in conjunction with fetal heart rate testing (OR 0.08, 95% CI 0.06-0.12). REVIEWER'S CONCLUSIONS: Vibroacoustic stimulation offers benefits by decreasing the incidence of non-reactive cardiotocography and reducing the testing time. Further randomized trials should be encouraged to determine not only the optimum intensity, frequency, duration and position of the vibroacoustic stimulation, but also to evaluate the efficacy, predictive reliability, safety and perinatal outcome of these stimuli with cardiotocography and other tests of fetal wellbeing. PMID- 11279789 TI - Communicating with children and adolescents about their cancer. AB - BACKGROUND: Communication with children and adolescents with cancer about their disease and treatment and the implications of these is an important aspect of good quality care. It is often poorly performed in practice. Various interventions have been developed that aim to enhance communication involving children or adolescents with cancer. OBJECTIVES: To examine the effects of interventions to enhance communication with children and/or adolescents about their cancer, its treatment and their implications. SEARCH STRATEGY: The following electronic databases were searched: Cochrane Library; Medline; PsycLit; Cinahl; Cancerlit; EMBASE; Sociofile; Health Management Information Consortium; ASSIA; LISA; ERIC; PAIS; Information Science Abstracts; Dissertation Abstracts; JICST; Pascal; Linguistics and Language Behavior Abstracts; Mental Health Abstracts; AMED; MANTIS. Bibliographies of identified studies were also checked and contact made with experts in the field. SELECTION CRITERIA: Randomised and non-randomised controlled trials and before and after studies that evaluated the effects of interventions to enhance communication with children and/or adolescents about their cancer, treatment and related issues. DATA COLLECTION AND ANALYSIS: Data relating to the interventions, populations and outcomes studied and the design and methodological quality of included studies were extracted by one reviewer and checked by another reviewer. A narrative summary of the results is presented. MAIN RESULTS: Six studies met the criteria for inclusion. They were diverse in terms of the interventions evaluated, study designs used, types of people who participated and the outcomes measured. One study of a computer assisted education programme reported improvements in knowledge and understanding about blood counts and cancer symptoms. Two out of two studies of school reintegration programs reported improvements in some aspects of psychosocial wellbeing (one in anxiety and one in depression), social wellbeing (two in social competence and one in social support) and behavioural problems; and one reported improvements in physical competence. REVIEWER'S CONCLUSIONS: Interventions to enhance communication involving children and adolescents with cancer have not been widely or rigorously assessed. The weak evidence that exists suggests that some children and adolescents with cancer may derive some benefit from specific information-giving programs and from interventions that aim to facilitate their reintegration into school and social activities. More research is needed to investigate the effects of these and other related interventions. PMID- 11279790 TI - Rescue high frequency oscillatory ventilation vs conventional ventilation for infants with severe pulmonary dysfunction born at or near term. AB - BACKGROUND: Pulmonary disease is a major cause of mortality and morbidity in term and near term infants. Conventional ventilation (CV) has been used for many years but may lead to lung injury, require the subsequent use of more invasive treatment such as extra corporeal membrane oxygenation (ECMO), or result in death. There are some studies indicating that high frequency oscillatory ventilation (HFOV) may be more effective in these infants as compared to CV. OBJECTIVES: The objective of this review is to determine if HFOV, as compared to conventional ventilation, reduces mortality and morbidity in term or near term infants with intractable lung disease without an increase in adverse effects. SEARCH STRATEGY: Standard search methods of the Cochrane Neonatal Review group were used. These included searches of the Oxford Database of Perinatal Trials, MEDLINE, EMBASE, previous reviews including cross references, abstracts, conferences and symposia proceedings, expert informants, and journal hand searching by the Cochrane Collaboration. SELECTION CRITERIA: Randomized or quasi randomized trials comparing HFOV and CV in term or near term infants with intractable respiratory failure were included in this review. DATA COLLECTION AND ANALYSIS: The standard methods of the Cochrane Neonatal Review Group were used. The investigators separately extracted, assessed and coded all data for each study. Any disagreement was resolved by discussion. Data were synthesized using relative risk (RR) and risk difference (RD). MAIN RESULTS: Only one trial met the inclusion criteria. This rescue trial of 81 infants showed no evidence of a reduction in mortality at 28 days [RR 0.51 (0.05, 5.43)] or in failed therapy on the assigned mode of ventilation requiring cross-over to the other mode [RR 0.73 (0.47, 1.13)]. There were no significant differences in the numbers of patients requiring extracorporeal membrane oxygenation [RR 2.05 (0.85, 4.92)], days on a ventilator, days in oxygen or days in hospital. REVIEWER'S CONCLUSIONS: There are no data from randomized controlled trials supporting the routine use of rescue HFOV in term or near term infants with severe pulmonary dysfunction. The area is complicated by diverse pathology in such infants and by the occurrence of other interventions (surfactant, inhaled nitric oxide, inotropes). Randomized controlled trials are needed to establish the role of rescue HFOV in near term and term infants with pulmonary dysfunction before widespread use of this mode of ventilation in such infants. PMID- 11279791 TI - Oral oestrogen replacement therapy versus placebo for hot flushes. AB - BACKGROUND: Hot flushes and night sweats are common symptoms experienced by menopausal women. Hormone replacement therapy (HRT), containing oestrogens alone or oestrogens together with progestogens in a cyclic or continuous regimen, is often recommended for their alleviation. OBJECTIVES: To examine the effect of oral HRT compared to placebo on these vasomotor symptoms and the risk of early onset side-effects. SEARCH STRATEGY: As developed by the Menstrual Disorders Group and Subfertility group of the Cochrane Collaboration. SELECTION CRITERIA: Double-blind, randomised, placebo-controlled trials of oral HRT therapy for at least three months duration. DATA COLLECTION AND ANALYSIS: Study quality and outcome data were assessed independently. Random effects models were considered appropriate due to the variety of trial methodologies. The meta-analyses were explored for sensitivity to trial quality and therapy duration. Symptom frequency and severity were assessed separately, together with withdrawals and side effects. Frequency data were analysed using the Weighted Mean Difference (WMD) between treatment and placebo outcomes. For severity data, odds ratios were estimated from the proportional odds model. From 99 references originally identified, 21 trials meeting the selection criteria were included in the review. Study participants totalled 2,511. Trial duration ranged from three months to three years. MAIN RESULTS: There was a significant reduction in the weekly hot flush frequency for HRT compared to placebo (WMD -17.46, 95% CI -24.72, -10.21). This was equivalent to a 77% reduction in frequency (95% CI 58.2, 87.5) for HRT relative to placebo. Symptom severity was also significantly reduced compared to placebo (OR 0.13, 95% CI 0.08, 0.22). Withdrawal for lack of efficacy occurred significantly more often on placebo therapy (OR 17.25, 95% CI 8.23, 36.15). Withdrawal for adverse events, commonly breast tenderness, oedema, joint pain and psychological symptoms, was not significantly increased for HRT therapy (OR 1.38, 95% CI 0.87, 2.21). In women who were randomised to placebo treatment, a 50.8% (95% CI 41.7, 58.5) reduction in hot flushes was observed between baseline and end of study. REVIEWER'S CONCLUSIONS: Oral HRT is highly effective in alleviating hot flushes and night sweats. Therapies purported to reduce such symptoms must be assessed in blinded trials against a placebo or a validated therapy. Withdrawals due to side-effects were only marginally increased in the HRT groups despite the inability to tailor HRT in these fixed dose trials. Comparisons of hormonal doses, product types or regimens require analysis of trials with these specific "within study" comparisons. PMID- 11279792 TI - Imaging multidrug resistance with radiolabeled substrates for P-glycoprotein and multidrug resistance protein. PMID- 11279793 TI - Positron-emitting radiopharmaceuticals: how safe are they? PMID- 11279794 TI - Technetium-99m-tetrofosmin would be a substrate for multidrug resistance associated protein (MRP): comparison between a leukemia cell line with high MRP gene expression and its parental cell line. AB - The kinetics of cellular accumulation and retention of technetium-99m-tetrofosmin (99mTc-TF) were investigated in wild type HL60/WT cell line and in its doxorubicin-resistant HL60/DOX cell line with multidrug resistance-associated protein (MRP), but without P-gp overexpression, to determine whether 99mTc-TF is a substrate for MRP. METHODS: The accumulation and washout of 99mTc-TF were observed in both cell lines at 37 degrees C. The effect of verapamil on the kinetics was also assessed. RESULTS: 99mTc-TF net accumulation was significantly lower in HL60/DOX (1.35 +/- 0.23%) than in HL60/WT (12.79 +/- 0.47%) at 60 min (P < 0.001). Three minutes after exchanging the incubation solution to the tracer free medium, only 18.20 +/- 0.34% of 99mTc-TF remained in HL60/DOX, whereas 84.74 +/- 0.65% did in HL60/WT (P < 0.001). In the presence of 10 microM verapamil, 99mTc-TF net accumulation in HL60/DOX was 302% of the control and the washout was significantly delayed. CONCLUSION: 99mTc-TF would be a substrate for MRP and 99mTc-TF may be used as a functional imaging agent of MRP in vivo. PMID- 11279795 TI - Numerical selection of optimal tumor imaging agents with application to engineered antibodies. AB - Three analytic indicators were used to compare five members of a monoclonal antibody (Mab) family. The cognates consisted of the genetically engineered intact chimeric IgGI (cT84.66) and related engineered fragments [scFv, diabody, minibody, F(ab')2] reactive against the same epitope of carcinoembryonic antigen (CEA). All analyses were based on radioiodinated Mabs targeting to colorectal xenografts of LS174T tumors in nude mice. Affinity constants were evaluated initially. A second indicator was the imaging figure of merit (IFOM) which determines how rapidly a statistically significant tumor image can be acquired. Finally, deconvolution was used to determine tumor temporal response to an arterial bolus. This last analysis gave the possible tumor accumulation in the absence of normal tissue sequestration. Affinities were all in excess of 10(8) M 1 and were highest for the divalent Mabs. Using the IFOM criterion, an 131I label was best suited as a radiolabel for the intact (IgG) T84.66, while an 123I label indicated optimal imaging with either minibody or F(ab')2. Deconvolution analyses showed that divalent members behaved similarly while the univalent member (scFv) had a tumor residence time smaller by an order of magnitude. The diabody had the largest impulse response function, but renal uptake may limit its present usefulness. PMID- 11279796 TI - Role of repeated F-18 fluorodeoxyglucose imaging in management of patients with bone and soft tissue sarcoma. AB - AIM OF THE STUDY: To assess the impact of repeated F-18 FDG studies on the management of patients with bone and soft tissue (B&S) sarcomas. MATERIAL AND METHODS: Twenty patients with B&S tissue tumors (11 M and 9 F age 17-72 years) had 52 F-18 FDG Dual Head Coincidence Imaging (DHCI) studies. 7 patients were followed for 6 months to 2 years clinically after removal of the primary tumor. Thirteen patients were evaluated for suspected recurrences. Patient's preparation, F-18 FDG injection and imaging procedure were done according to department protocol. Attenuation corrected images were interpreted visually by 3 trained physicians. Tumor to background ratios were calculated for all lesions. RESULTS: In 13 patients having both studies, baseline FDG and CT/MRI were concordant in 8 patients, FDG detected more lesions in 3 patients but it did not detect 4 metastatic pulmonary nodules in 2 patients. Follow up studies showed stable disease in 10 patients while 6 patients who showed worsening disease needed to change their chemotherapy. Surgery was avoided in 2 patients and 2 patients showed improved response. CONCLUSION: Repeated F-18 FDG DHCI examinations proved to have an impact on the clinical management of patients with malignant bone and soft tissue sarcoma. It helps to differentiate postoperative changes from local recurrence. PMID- 11279797 TI - Treatment of kidney cancer with autologous tumor cell vaccines of short-term cell lines derived from renal cell carcinoma. AB - BACKGROUND: We established short-term cultures of autologous tumors from patients with renal carcinoma for use as active specific immunotherapy (i.e., autologous vaccine). METHODS: Between 9/91 and 9/99 the cell biology laboratory of the Hoag Cancer Center received 69 kidney tumor samples that had been surgically excised, including 43 primary tumors and 26 metastatic lesions. Efforts were made to establish short-term tumor cell cultures to use as autologous tumor cell vaccines. Prior to treatment, patients underwent a baseline skin test for delayed tumor hypersensitivity (DTH) and then received s.c. injections of 10 million irradiated tumor cells that were given with various adjuvants weekly x3 and then monthly x5. RESULTS: Cell lines were established for 55/69 patients (80%) including 36/43 (84%) from primary tumors and 19/26 (73%) from distant metastases. Vaccines were prepared for 41 patients; 27 were treated. At the time of this analysis, follow up data was available for 26 patients with a median follow up > 5 years. Treatment was well-tolerated. Of 10 patients who had no evident disease at the time of treatment, nine were alive 1-8 years later; 5/8 had conversion of their DTH test from negative to positive. For 16 patients with measurable metastatic disease at the time of treatment, there were no objective tumor responses; their median survival was 5.0 months. Among these 16 patients, only 1/8 DTH tests converted, but three had a positive baseline DTH test; one was previously treated with interleukin-2 and tumor infiltrating lymphocytes and two others were previously treated with autolymphocyte therapy. CONCLUSIONS: Vaccine therapy with short-term cultures of autologous tumor cells is feasible, well tolerated and associated with conversion of DTH and long-term survival in patients who are free of disease at the time treatment is initiated. However, significant anti-tumor responses were not seen in patients with measurable disease at the time vaccine treatment was initiated. PMID- 11279798 TI - An overview on anticancer activities of the Viscum album extract Isorel. AB - The activity principle of the mistletoe (Viscum album L.) phytotherapeutics could be considered as combined cytotoxic and "biological response modifying" activities (increasing host defense against cancer) that result from the activities of the plant lectins and the other biologically relevant substances. We found before that the aqueous extract Isorel, produced by Novipharm GmbH (Portschach, Austria) from the entire plant (planta tota) of fresh mistletoe under standardized conditions with bioassay validated batch consistency, can be valuable in experimental adjuvant cancer therapy increasing efficiency of cyclophosphamide chemotherapy. In current study we found that Isorel increases the reactivity of the tumor-bearing mice lymphocytes to the mitogens (ConA and LPS) in vitro, thus indicating its immune stimulating effects for the cancer immunosuppressed lymphocytes. Moreover, Isorel inhibited the incorporation of 3H labelled amino acids (protein synthesis) in various malignant cell lines. For the growth inhibition mostly higher MW components were responsible, although even less than 500 Da components were also active. We further analyzed the effects of drug application in vicinity of tumor (murine mammary carcinoma) and compared it with systemic effects. The animals carried mammary carcinoma in both hind limbs and were also injected with tumor cells i.v. to develop artificial lung metastases. Isorel was applied only at the right side (in the limb distal from the tumor) and caused persistent and almost complete inhibition of the tumor growth for 2/7 animals. Anticancer effects were less pronounced on the contralateral side tumors, although tumor growth rate was transiently reduced for some mice. Histology revealed that Isorel treatment, both at the side of tumor and systemically, increased the incidence of apoptosis and necrosis in the tumors, while reduction of mitosis was noticed only for the tumors in vicinity of the tumor exposed to Isorel. Finally, animals treated with Isorel had, on the average, three times less lung metastases than the controls. Thus, we conclude that both local and systemic effects of the application of Isorel could be of benefit for the tumor-bearing organism resulting in immunomodulation combined with tumor growth inhibition and reduction of metastases. According to the in vitro results, antitumorous effects could be the result not only of the mistletoe lectins and the other high MW factors, but also of the very low MW (< 500 Da) substances that deserve further analyses. PMID- 11279799 TI - Cell cycle control in cellular homeostasis during the immune response: interactions between TH1, TH2 cytokines, and Bcl2 and p53 molecules. AB - Cytokine regulation of lymphocyte survival may play an important role in the control of the cell cycle during the immune response both in health and disease. Expression of the Bcl2 gene promotes cell survival by countering apoptosis stimuli. The p53 protein has been implicated in the control of the cell cycle, in the synthesis and repair of DNA and in programmed cell death. TH1 and TH2 cytokines exert a mutual cross-regulation on the precursors of TH1- or TH2-type effector cells which are important mediators in directing the immune system towards the appropriate response. TH1 and TH2 cytokines have also been implicated in the modulation of the expression of cell cycle regulator genes. Therefore, the study of the relationships between TH1 and TH2 cytokines and Bcl2 and p53 molecules in healthy subjects could lead to a better understanding of the physiological regulation of the immune response and identify markers for prognostic and diagnostic indices and biotherapeutic treatment. We determined the serum levels of cytokines (IL2, IFN gamma, IL4, IL10, IL5, IL6, IL1 beta, TNF alpha, IL8), soluble receptors (sIL2R, sIL6R), Bcl2-protein and p53-antibody in a group of healthy subjects. Multivariate statistical analyses were used to study the cytokine network relationships with Bcl2-protein and p53-antibody, as they allow a simultaneous evaluation of all variables which reflects the physiological situation. Our overall results suggest that relationships exist between TH1 and TH2 cytokines and the Bcl2-protein and p53-antibody in physiological conditions. This information could now be used in experimental studies to create diagnostic and prognostic indices for the monitoring of health and disease. PMID- 11279800 TI - In vivo and in vitro measurement of apoptosis in breast cancer cells using 99mTc EC-annexin V. AB - OBJECTIVE: The purpose of this study was to develop an imaging technique to measure and monitor tumor cells undergoing programmed death caused by radiation and chemotherapy using 99mTc-EC-annexin V. Annexin V has been used to measure programmed cell death both in vitro and in vivo. Assessment of apoptosis would be useful to evaluate the efficacy and mechanisms of therapy and disease progression or regression. METHODS: Ethylenedicysteine (EC) was conjugated to annexin V using sulfo-N-hydroxysuccinimide and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide-HCl as coupling agents. The yield of EC-annexin V was 100%. In vitro cellular uptake, pre- and post-radiation (10-30 Gy) and paclitaxel treatment, was quantified using 99mTc-EC-annexin V. Tissue distribution and planar imaging of 99mTc-EC-annexin V were determined in breast tumor-bearing rats at 0.5, 2, and 4 hrs. To demonstrate in vivo cell apoptosis that occurred during chemotherapy, a group of rats was treated with paclitaxel and planar imaging studies were conducted at 0.5-4 hrs. Computer outlined region of interest (ROI) was used to quantify tumor uptake on day 3 and day 5 post-treatment. RESULTS: In vitro cellular uptake showed that there was significantly increased uptake of 99mTc-EC-annexin V after irradiation (10-30 Gy) and paclitaxel treatment. In vivo biodistribution of 99mTc-EC-annexin in breast tumor-bearing rats showed increased tumor-to-blood, tumor-to-lung and tumor-to-muscle count density ratios as a function of time. Conversely, tumor-to blood count density ratios showed a time-dependent decrease with 99mTc-EC in the same time period. Planar images confirmed that the tumors could be visualized clearly with 99mTc-EC-annexin. There was a significant difference of ROI ratios between pre- and post-paclitaxel treatment groups at 2 and 4 hrs post injection. CONCLUSION: The results indicate that apoptosis can be quantified using 99mTc-EC annexin and that it is feasible to use 99mTc-EC-annexin to image tumor apoptosis. PMID- 11279801 TI - CD40 Ligand--an anti-apoptotic molecule in Hodgkin's disease. AB - The expression of CD40L was investigated in HD involved lymph nodes by flow cytometry (FCM) and reverse transcriptase polymerase chain reaction (RT-PCR). Also an investigation of the role of CD40L in upregulation of the anti-apoptotic gene BclxL in a Hodgkin's disease (HD) derived cell line was undertaken. HD patients (n = 18) had significantly higher numbers of activated CD4+ and CD8+ T cells in the tumor microenvironment as compared to controls (n = 8). HD patients also demonstrated higher numbers of CD4+, CD8+ and CD19+ lymphocytes co expressing CD40L as compared to controls. The CD40L signal was consistently and significantly upregulated in HD patients (n = 5) as compared to controls (n = 3) at the mRNA level. RT-PCR and FCM analysis revealed that soluble CD40L upregulated BclxL levels in the Fas-sensitive HD cell line HDLM2. We conclude that CD40L can act as an important anti-apoptotic molecule by upregulating BclxL expression in Reed-Sternberg cells of HD and may be partly responsible for their survival 'in-vivo'. PMID- 11279802 TI - Radioimmunotherapy of solid tumors: from fairytale to reality. PMID- 11279804 TI - An ambivalent alliance. Hostile and benevolent sexism as complementary justifications for gender inequality. AB - The equation of prejudice with antipathy is challenged by recent research on sexism. Benevolent sexism (a subjectively favorable, chivalrous ideology that offers protection and affection to women who embrace conventional roles) coexists with hostile sexism (antipathy toward women who are viewed as usurping men's power). The Ambivalent Sexism Inventory, first validated in U.S. samples, has been administered to over 15,000 men and women in 19 nations. Hostile and benevolent sexism are complementary, cross-culturally prevalent ideologies, both of which predict gender inequality. Women, as compared with men, consistently reject hostile sexism but often endorse benevolent sexism (especially in the most sexist cultures). By rewarding women for conforming to a patriarchal status quo, benevolent sexism inhibits gender equality. More generally, affect toward minority groups is often ambivalent, but subjectively positive stereotypes are not necessarily benign. PMID- 11279803 TI - Phase I clinical evaluation of a neutralizing monoclonal antibody against epidermal growth factor receptor. AB - Ior egf/r3, a neutralizing monoclonal antibody (mAb) against Epidermal Growth Factor Receptor (EGFR) was generated at the Cuban Institute of Oncology. Immunoscintigraphic studies in 148 patients with this 99-m Technetium (99Tc) labeled mAb, showed a high sensitivity and specificity for in vivo detection of epithelial tumors. To study safety, pharmacokinetic and immunogenicity of ior egf/r3 at high doses, a phase I clinical trial was conducted. Nineteen patients with advanced epithelial tumors received 4 mAb intravenous infusions at 6 dose levels: from 50 to 500 mg. Previously, immunoscintigraphic images using the same mAb labeled with 99Tc were acquired. Blood samples were collected for pharmacokinetic analysis and HAMA response. After mAb therapy, objective response was classified according to WHO criteria. Ior egf/r3 was well tolerated in spite of the high-administered doses. Only a severe adverse reaction consisting of hypotension and lethargy was observed. In 13 patients, selective accumulation of 99Tc-labeled mAb was observed at the site of the primary tumor or the metastasis. Pharmacokinetic analysis revealed that elimination half-life and the area under the time-concentration curve increased linearly with dose. HAMA response was detected in 17 patients. After 6 months of mAb therapy, 4 patients had stable disease. One patient had a tumor partial remission after 3 cycles of ior egf/r3. PMID- 11279805 TI - Single-participant research design. Bringing science to managed care. AB - The ongoing transition to managed health care continues to have repercussions for health care providers, perhaps the most important of which is an emphasis on accountability for demonstrating the usefulness of clinical interventions. This requirement places a premium on intervention research and highlights the historically strained relationship between psychological research and professional practice. In the midst of this challenge, researchers have increasingly criticized the logic and practice of traditional null hypothesis significance testing. This article describes the history, epistemology, and advantages of single-participant research designs for behavioral scientists and professionals in clinical settings. Although its lack of correspondence with the Fisherian tradition has precluded widespread adoption, the single-participant alternative features a design power and flexibility well suited to both basic science and applied research. PMID- 11279806 TI - Psychological testing and psychological assessment. A review of evidence and issues. AB - This article summarizes evidence and issues associated with psychological assessment. Data from more than 125 meta-analyses on test validity and 800 samples examining multimethod assessment suggest 4 general conclusions: (a) Psychological test validity is strong and compelling, (b) psychological test validity is comparable to medical test validity, (c) distinct assessment methods provide unique sources of information, and (d) clinicians who rely exclusively on interviews are prone to incomplete understandings. Following principles for optimal nomothetic research, the authors suggest that a multimethod assessment battery provides a structured means for skilled clinicians to maximize the validity of individualized assessments. Future investigations should move beyond an examination of test scales to focus more on the role of psychologists who use tests as helpful tools to furnish patients and referral sources with professional consultation. PMID- 11279807 TI - The nurture assumption persists. PMID- 11279808 TI - Behavior genetics and parenting theory. PMID- 11279809 TI - Models of parenting and adolescent drinking. PMID- 11279810 TI - Are behavioral genetic and socialization research compatible? PMID- 11279811 TI - Toward nature with nurture. PMID- 11279812 TI - A marriage of opposites: self-help and the health care system. PMID- 11279813 TI - Growing challenges for the protection of global health in the 21st century. PMID- 11279814 TI - Infectious disease surveillance in North Rhine-Westphalia: first steps in the development of an early warning system. AB - It is often difficult to detect warning signals on infectious disease outbreaks from raw surveillance data. Data need to be used for a timely generation and distribution of information on the current state of infectious diseases. This offers the opportunity to detect outbreaks and to initiate preventive measures. To improve the surveillance system in North Rhine-Westphalia we have introduced an infectious disease barometer, a simple tool based on weekly notification data. The aim of this tool is not to provide in depth data analysis, but to help to detect clusters or outbreaks. This can be the first step in the development of an early warning system and can support epidemiological investigation and policy making. PMID- 11279815 TI - Hospital infestation by the cluster fly, Pollenia rudis sensu stricto Fabricius 1794 (Diptera: Calliphoridae), and its possible role in transmission of bacterial pathogens in Germany. AB - The potential of the cluster fly, Pollenia rudis sensu stricto, to transmit bacterial pathogens was investigated during a mass infestation that took place in a German hospital. Cluster flies were individually examined for mesophilic bacteria carried on the exoskeleton. Bacterial growth could only be detected by using the enrichment culture technique to increase sensitivity, but not by direct intoculation of fly samples to agar plates. All 50 cluster fly samples that were tested carried opportunistic aerobic mesophilic Bacillus spp., whereas 41 fly samples were positive for Erwinia spp., 16 samples for Erwinia amylovara, 24 samples for Stenotrophomonas maltophilia, and 4 samples for Flavobacterium odoratum. Staphylococcus lugdunensis and Pseudomonas aeruginosa were found in 5 samples. No bacteriologically sterile cluster fly samples were obtained. The whole bacterial pattern found on P. rudis s. s. is known for its potential to cause opportunistic and/or nosocomial infections in humans. The results obtained led to the assumption that mass infestations of cluster flies occurring in sensitive areas, especially in hospitals, may cause a low, but not neglectable health threat due to mechanical transmission of bacterial pathogens. PMID- 11279816 TI - Prevalence and growth of pathogens on salad vegetables, fruits and sprouts. AB - A total of 120 samples, comprising different types of raw vegetables (seven), fruits (three) and sprouts (three) obtained from street vendors, were tested for aerobic plate count, coliform count and various food-borne pathogens. Average aerobic plate counts for salad vegetables, fruits and sprouts were greater than 10(10) cfu/g and 10(9) cfu/g respectively. Pathogens isolated were S. aureus, E. coli, Enterobacter sp., Klebsiella sp., S. typhi, Serratia sp., Providencia sp. and P. aeruginosa. The antibiotic resistant patterns of the isolates revealed P. aeruginosa to be the most antibiotic resistant, E. coli, Salmonella, Enterobacter and P. aeruginosa also showed the presence of plasmids. The model development phase of this study involved 27 growth curves conducted under 9 combinations of temperature and pH in the Brain Heart Infusion Broth. Models for specific growth rate and lag period were developed by response surface modelling using multiple linear regression analysis. The model provides an estimate of bacterial growth in response to any combination of the variables studied within specified ranges. Growth patterns of organisms on vegetable and fruits were also studied at room temperature (32 degrees C) to assess the growth in the actual food environment. Cucumber and watermelon supports the growth of S. aureus and S. typhi, carrot retarded their growth while pineapple did not support the growth. PMID- 11279817 TI - Distribution of free-living amoebae (FLA) during preparation and supply of drinking water. AB - Free-living amoebae (FLA) are widely distributed in aquatic environments with increasing importance in hygienic, medical and ecological relationship to man. Only few data are available about abundances of these protozoa in the treatment of drinking water and standards in the management of water quality are not suitable for detection of FLA. Prevalence of FLA were investigated within six selected German drinking water treatment plants in the course of the purification process of surface water and in a subsequent drinking water supply. The data give a short survey about the prevalence and reduction of FLA in processing and supply of drinking water. PMID- 11279818 TI - High concentrations of fluoride and boron in drinking water wells in the Muenster region--results of a preliminary investigation. AB - In 1998, two cases of severe dental fluorosis in schoolchildren occurred in the Muenster region. These cases took place in one household, where fluoridated toothpaste, fluoridated salt, and fluoride tablets were consumed. Furthermore, the family used drinking water from its private well only. Analyses of the well water ordered by local health officials revealed very high amounts of fluoride, boron, and other electrolytes. This unusual combination of high amounts of fluoride and boron could also be found in the water of a great number of other private wells that are the only source for drinking water in this rural region of the Muensterland. Anthropogenic sources could be excluded. Because of this, the results of the water samples were collated to the specific geological situation in this area. In the Muenster region there are marl layers of the chalk era covered with quarternary sediments. The quarternary sediments are up to 10 to 20 metres thick and they usually conduct the groundwater. The marl contains high concentrations of fluoride and boron. In some places the groundwater has contact with these layers. To check the amount of fluoride and boron in the groundwater, indicator values were sought, which can give a hint of high contents of these trace elements. In this study the conductivity and acidity were identified as possible indicators of a high amount of fluoride and boron in the drinking water in this specific region. To work economically and efficiently, the drinking water should be checked for fluoride and boron on a regular basis only when these values are extraordinarily high. In the case of high concentrations, especially of fluoride, in the drinking water the persons concerned should be informed about their potential health risk, giving them the opportunity to optimise the total daily intake of fluoride. PMID- 11279819 TI - A geographical information system (GIS) as a tool for microbial risk assessment in catchment areas of drinking water reservoirs. AB - The main tributaries of three drinking water reservoirs of Northrhine-Westfalia (Germany) were monitored within a 14-month period mainly for bacterial and parasitic contamination. In this context a detailed geo-ecological characterisation within the differing catchment areas was carried out to reveal a reliable informational basis for tracing back the origin of microbial loads present in the watercourses. To realise a microbial risk assessing geo-ecological information system (MRA-GIS), a Geographical Information System (GIS) has been implemented for the study areas. The results of the microbiological investigations of the watercourses showed an input of pathogens into all three of the tributaries. It could be demonstrated that the use of MRA-GIS database and some GIS-techniques substantially support the spatial analysis of the microbial contamination patterns. From the hygienic point of view, it is of the utmost importance to protect catchment areas of surface water reservoirs from microbial contamination stemming from human activities and animal sources. This constitutes essential part of the multi-barrier concept which stresses the importance of reducing diffuse and point pollution in catchment areas of water resources intended for human consumption. MRA-GIS proves to be helpful to manage multi barrier water protection in catchment areas and ideally assists the application of the HACCP concept on drinking water production. PMID- 11279820 TI - Pulmonary function and urban air pollution in preschool children. AB - Epidemiologic evidence indicates that respiratory disease in infancy and childhood has respiratory health consequences in later life. Pulmonary function is considered a good index of early effects. This study assessed the relationship of pulmonary function in preschool children in Leipzig, Germany, and exposure to high levels of air pollution during early childhood. Spirometric measures were taken of 235 preschoolers (126 boys, 109 girls, mean age 5.1 +/- 1.3 years) attending 16 randomly selected daycare centres, using the 'Bosch Spiro 501' spirometer. The results showed decrements in the average FVC (85.5% predicted [pred]) and FEV1 (90.2% pred) differing with spatial variations in the ambient air pollution burden of the children's residential area. Exposure to a pollution profile of heavy traffic and/or domestic heating showed markedly lower FVC (78.9% and 85.5% pred, respectively) and FEV1 (82.4% and 88.5% pred). Miller's Diagnostic Quadrant Model of Disease Classification, categorizing pulmonary function data for preliminary diagnostic purposes, assessed the lung function values (FVC% pred/relative FEV1 [FEV1/FVC]%) of a significant number of children as 'restrictions' (n = 52; 22.1%). Summarizing: variations in spirometric indices were observed across exposure groups with a significant number of children showing signs of a restrictive ventilatory function. These cross-sectional data, however, do not permit to conclude with any degree of certainty that this is indicative of an early sign of a functional deficit. PMID- 11279821 TI - Comparison of two sampling methods for the detection of gram-positive and gram negative bacteria in the environment: moistened swabs versus Rodac plates. AB - The objective of this study was to evaluate the moistened swab technique vs. Rodac plates for detecting Gram-positive and Gram-negative bacteria in the inanimate environment. Over a period of 22 months, the environment of 190 patients infected or colonized with MRSA, VRE or multiresistant Gram-negative bacteria was sampled in turn. MRSA and VRE could be detected with either method in 33 out of 54 (61.1%) patient rooms in 174 out of 706 (24.6%) environmental samples. However, multiresistant Gram-negative bacteria were found in 42 out of 136 (30.9%) rooms with a very low frequency of 89 out of 1827 (4.9%) environmental samples (p < 0.0001). The sensitivity of the swab technique for Gram-positive cocci was 54% (94/174) vs. 69.5% (121/174) for the Rodac plates, ([CI95%], 47-61% vs. 62-76%, p < 0.05). In contrast, the sensitivity of the swab technique for Gram-negative bacteria was 74.2% (66/89) vs. 42.7% (38/89) for the Rodac plates, ([CI95%], 64-83% vs. 32-54%, p < 0.05). In conclusion, environmental contamination with Gram positive cocci is detected more often than with Gram-negative bacteria. For the detection of Gram-positive cocci, Rodac plates are superior to the swab technique; whereas Gram-negative rods can be detected more often by the swab technique. All these results proved to be statistically significant. PMID- 11279822 TI - Genotoxicity of arsenical compounds. AB - With respect to global human health hazard, arsenic (As) is one of the most important environmental single substance toxicants. Currently, millions of people all over the world are exposed to the ubiquitous element in exposure levels leading to long-term toxicity, in particular cancer. Unfortunately, it has not been elucidated up to now how As mechanistically leads to the induction of neoplasia. Besides its tumorigenic potential, As has been shown to be genotoxic in a wide variety of different experimental set-ups and biological endpoints. In vitro, the element was shown to induce chromosomal mutagenicity like micronuclei, chromosome aberrations, and sister chromatid exchanges. It mainly acts clastogenic but also has an aneugenic potential. Instead, its potential to induce point mutations is very low in bacterial as well as in mammalian cell systems. However, in combined exposure with point mutagens in vitro, As was shown to enhance the frequency of chemical mutations in a synergistic manner. Additionally, As was shown to induce chromosome aberrations and micronuclei in vivo in experiments with mice. After long-term exposure to As-contaminated drinking water, the great majority of human biomonitoring studies found elevated frequencies of DNA lesions like micronuclei or chromosome aberrations. Respective occupational studies are few. Like it is the case for As carcinogenicity, it is not known through which mechanism the genotoxicity of As is mediated, although the data available indicate that As may act indirectly on DNA, i.e. via mechanisms like interference of regulation of DNA repair or integrity. Because of the indirect mode of action, it has been discussed as well that As's genotoxicity may underlie a sublinear dose-response relationship. However, various problems like non-standardized test systems and experimental variability make it impossible to prove such statement. Basically, to be able to improve risk assessment, it is of crucial importance to scientifically approach the mechanistic way of induction of As's genotoxicity and carcinogenicity. PMID- 11279824 TI - Sensitive indoor air monitoring of formaldehyde and other carbonyl compounds using the 2,4-dinitrophenylhydrazine method. AB - The toxic potential of formaldehyde and other aliphatic/aromatic carbonyl compounds requires the determination of even low amounts of these compounds in indoor air. The existing DFG-method for workplace monitoring using adsorption at 2,4-dinitrophenylhydrazine (DNPH)-coated sorbents followed by HPLC-UV/DAD analysis of the extract was modified in order to decrease detection limits. The improvement included an increase in volume and rate of the air sampling, testing applicability of different adsorption materials and a decrease of the extraction volume of the hydrazones. 13 DNPH-derivatives could be separated well on a RP18 column followed by UV/DAD-detection at 365 nm. Recovery rates of 70-100% were determined (apart from acetone with 19%) using dynamically produced artifical carbonyl atmospheres. Detection limits of 0.05-0.4 microgram/m3 were reached by this method which are sufficient for indoor air monitoring. PMID- 11279823 TI - Comparison of different digestive tract models for estimating bioaccessibility of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/F) from red slag 'Kieselrot'. AB - 'Kieselrot' (red slag), a highly PCDD/F-contaminated leaching residue from a copper production process, has been used as surface layer for more than 1,000 sports fields, playgrounds and pavements in Germany and neighbouring countries. Children can ingest this material directly by hand-to-mouth activities or soil pica behaviour. Furthermore secondary contamination of farm land or kitchen gardens by drift of red slag dust may lead to an enrichment of PCDD/F within the food-chain. PCDD/F can be mobilized from contaminated materials by digestive juices and thus become bioaccessible for intestinal absorption. Two different digestive tract models were used to estimate the bioaccessibility of PCDD/F from red slag and to study the influence of food material on the mobilization of the contaminants. The bioaccessibility of PCDD/F from red slag depends on the charge of red slag material used, the bile content of the intestinal juice and on the presence of lipophilic foodstuffs. A low bioaccessibility of less than 5% was found when using a digestive tract model with a low bile content and in absence of food material. The bioaccessibility was estimated to be more than 60% when using a model with a higher bile content and in the presence of whole milk powder. A low bioaccessibility of PCDD/F from red slag in general--as assumed until now and mentioned in legal provision--was not confirmed by our study. Considering observations for the different homologue groups it is obvious that bioaccessibility is the first of several important steps to estimate human health risks arising from contaminated materials. In case red slag contaminated with PCDD/F their absorption rate in the digestive tract and/or metabolism might be at least just like important. PMID- 11279825 TI - Improved method for the fluorimetric detection of beta-D-galactosidase in water. AB - A very convenient method to quantify coliform bacteria in water can probably be designed via the determination of the activity of the enzyme beta-D galactosidase, whose natural occurrence is, apart from less frequently occurring aeromonads mainly restricted to this type of microorganisms. 4-methylumbelliferyl beta-D-galactoside is used as substrate, which is hydrolyzed during the enzymatic reaction; the released 4-methylumbelliferone can be quantified fluorimetrically. In the present study the influence of various physical and chemical parameters on the determination is investigated and the experimental conditions are optimized. Most important entities are the pH value during hydrolysis, the presence of nutrients and co-factors in the sample, and the modification of the substrate. Statistical evaluation of the results obtained by changing single or multiple parameters reflects clearly their positive or negative influence on the enzyme activity. Thus, deliberate addition of surfactants, specific nutrients, salts and co-enzymes results in a significantly increased activity of beta-D-galactosidase towards the substrate, which can be advantageously exploited to increase the sensitivity of the analytical method together with a decrease of the detection limit. The influence of the parameters and the optimized conditions of the improved analytical methods are presented. PMID- 11279826 TI - Inversion polymorphisms and nucleotide variability in Drosophila. PMID- 11279827 TI - Quantitative trait locus mapping of fitness-related traits in Drosophila melanogaster. AB - We examined the genetic architecture of four fitness-related traits (reproductive success, ovariole number, body size and early fecundity) in a panel of 98 Oregon R x 2b3 recombinant inbred lines (RILs). Highly significant genetic variation was observed in this population for female, but not male, reproductive success. The cross-sex genetic correlation for reproductive success was 0.20, which is not significantly different from zero. There was significant genetic variation segregating in this cross for ovariole number, but not for body size or early fecundity. The RILs were genotyped for cytological insertion sites of roo transposable elements, yielding 76 informative markers with an average spacing of 3.2 cM. Quantitative trait loci (QTL) affecting female reproductive success and ovariole number were mapped using a composite interval mapping procedure. QTL for female reproductive success were located at the tip of the X chromosome between markers at cytological locations 1B and 3E; and on the left arm of chromosome 2 in the 30D-38A cytological region. Ovariole number QTL mapped to cytological intervals 62D-69D and 98A-98E, both on the third chromosome. The regions harbouring QTL for female reproductive success and ovariole number were also identified as QTL for longevity in previous studies with these lines. PMID- 11279828 TI - The 62E early-late puff of Drosophila contains D-spinophilin, an ecdysone inducible PDZ-domain protein dynamically expressed during metamorphosis. AB - At the onset of metamorphosis in Drosophila melanogaster, the steroid hormone 20 OH ecdysone induces a small number of early and early-late puffs in the polytene chromosomes of the third-instar larval salivary gland whose activity is required for regulating the activity of a larger set of late puffs. Most of the corresponding early and early-late genes have been found to encode transcription factors that regulate a much larger set of late genes. In contrast, we describe here the identification of an ecdysone-regulated gene in the 62E early-late puff, denoted D-spinophilin, that encodes a protein similar to the mammalian protein spinophilin/neurabin II. The D-spinophilin protein is predicted to contain a highly conserved PP1-binding domain and adjacent PDZ domain, as well as a coiled coil domain and SAM domain, and belongs to a family of related proteins from diverse organisms. Transcription of D-spinophilin is correlated with 62E puff activity during the early stages of metamorphosis and is ecdysone-dependent, making this the first member of this gene family shown to be regulated by a steroid hormone. Examination of the dynamic patterns of D-spinophilin expression during the early stages of metamorphosis are consistent with a role in central nervous system metamorphosis as well as a more general role in other tissues. As D-spinophilin appears to be the only member of this gene family in Drosophila, its study provides an excellent opportunity to elucidate the role of an important adaptor protein in a genetic model organism. PMID- 11279829 TI - Clinal analysis of a chromosomal hybrid zone in the house mouse. AB - These studies centre on the 'Barcelona' karyotypic race of the western house mouse (Mus musculus domesticus), first described by Adolph & Klein (1981). This is one of many races within M. m. domesticus characterized by metacentric chromosomes that have originated by repeated Robertsonian fusions, with perhaps further modification by whole-arm reciprocal translocations. Data on 111 mice from 20 sites show that the race is centred 24 km to the west of Barcelona city and has a homozygous metacentric karyotype of 2n = 28 (3.8, 4.14, 5.15, 6.10, 9.11, 12.13). The race has a small range, and mice with the standard 40 acrocentric karyotype were caught only 30 km from the race centre. Throughout the area of occurrence of metacentrics there is polymorphism (i.e. presence of acrocentrics in the population), although all six metacentrics approach fixation close to the race centre. Thus, there is a hybrid zone between the Barcelona and standard races. The centres and widths of all clines (except 3.8) were determined. Likelihood ratio tests showed that most of the cline centres differed significantly in position (i.e. the clines were staggered) and the clines for metacentrics 6.10 and 9.11 were significantly narrower than those for 4.14, 5.15 and 12.13. Overall, the clines tended to be wider the further they were from the race centre. There are various possible explanations for this hybrid zone structure and further data are needed to distinguish between them. PMID- 11279830 TI - Rapid mutational declines of viability in Drosophila. AB - High rates of mildly deleterious mutation could cause the extinction of small populations, reduce neutral genetic variation and provide an evolutionary advantage for sex. In the first attempts to estimate the rate of mildly deleterious mutation, Mukai and Ohnishi allowed spontaneous mutations to accumulate on D. melanogaster second chromosomes shielded from recombination and selection. Viability of the shielded chromosomes appeared to decline rapidly, implying a deleterious mutation rate on the order of one per zygote per generation. These results have been challenged, however; at issue is whether Mukai and Ohnishi may have confounded viability declines caused by mutation with declines resulting from environmental changes or other extraneous factors. Here, using a method not sensitive to non-mutational viability changes, I reanalyse the previous mutation-accumulation (MA) experiments, and report the results of a new one. I show that in each of four experiments, including Mukai's two experiments, viability declines due to mildly deleterious mutations were rapid. The results give no support for the view that Mukai overestimated the declines. Although there is substantial variation in estimates of genomic mutation rates from the experiments, this variation is probably due to some combination of sampling error, strain differences and differences in assay conditions, rather than to failure to distinguish mutational and non-mutational viability changes. PMID- 11279831 TI - Linkage disequilibrium mapping of the bolting gene in sea beet using AFLP markers. AB - The possibility of using linkage disequilibrium mapping in natural plant populations was assessed. In studying linkage disequilibrium among 137 mapped AFLP markers in four populations of sea beet (Beta vulgaris ssp. maritima (L.) Arcang.) it was shown that tightly linked loci could be detected by screening for associations. It was hypothesized that the short distances spanned by linkage disequilibrium enable markers that are very tightly linked to a target gene to be identified. The hypothesis was tested by whole-genome screening of AFLP markers for association with the gene for the annual growth habit, the B gene, in a sample of 106 sea beets. Despite the dominant nature of AFLP, two markers showing significant linkage disequilibrium with the B gene were detected. The results indicate the potential use of linkage disequilibrium for gene mapping in natural plant populations. PMID- 11279832 TI - Two-locus identity probabilities and identity disequilibrium in a partially selfing subdivided population. AB - Measures of association of genes at different loci (linkage disequilibrium) are widely used to determine whether the structure of natural populations is clonal or not, to map genes from population data, or to test for the homogeneity of response of molecular markers to background selection, for example. However, the usual definitions of parameters for gametic associations may not be suitable for all these purposes. In this paper, we derive the recursion equations for one- and two-locus identity probabilities in an infinite island model. We study the role of drift, gene flow, partial selfing and mutation model on the expected association of genes across loci. We define the 'within-subpopulation identity disequilibrium' as the difference between the joint two-locus probability of identity in state and the expected product of one-locus identity probabilities. We evaluate this parameter as a function of recombination rate, effective size, gene flow and selfing rate. Within-subpopulation identity disequilibrium attains maximum values for intermediate immigration rates, whatever the selfing rate. Moreover, identity disequilibrium may be very small, even for high selfing rates. We discuss the implications of these findings for the analysis of data from natural populations. PMID- 11279833 TI - Inferring the trajectory of genetic variance in the course of artificial selection. AB - A method is proposed to infer genetic parameters within a cohort, using data from all individuals in an experiment. An application is the study of changes in additive genetic variance over generations, employing data from all generations. Inferences about the genetic variance in a given generation are based on its marginal posterior distribution, estimated via Markov chain Monte Carlo methods. As defined, the additive genetic variance within the group is directly related to the amount of selection response to be expected if parents are chosen within the group. Results from a simulated selection experiment are used to illustrate properties of the method. Four sets of data are analysed: directional selection with and without environmental trend, and random selection, with and without environmental trend. In all cases, posterior credibility intervals of size 95% assign relatively high density to values of the additive genetic variance and heritability in the neighbourhood of the true values. Properties and generalizations of the method are discussed. PMID- 11279834 TI - Masking and purging mutations following EMS treatment in haploid, diploid and tetraploid yeast (Saccharomyces cerevisiae). AB - The yeast, Saccharomyces cerevisiae, was used as a model to investigate theories of ploidy evolution. Mutagenesis experiments using the alkylating agent EMS (ethane methyl sulphonate) were conducted to assess the relative importance that masking of deleterious mutations has on response to and recovery from DNA damage. In particular, we tested whether cells with higher ploidy levels have relatively higher fitnesses after mutagenesis, whether the advantages of masking are more pronounced in tetraploids than in diploids, and whether purging of mutations allows more rapid recovery of haploid cells than cells with higher ploidy levels. Separate experiments were performed on asexually propagating stationary phase cells using (1) prototrophic haploid (MAT alpha) and diploid (MATa/alpha) strains and (2) isogenic haploid, diploid and tetraploid strains lacking a functional mating type locus. In both sets of experiments, haploids showed a more pronounced decrease in apparent growth rate than diploids, but both haploids and diploids appeared to recover very rapidly. Tetraploids did not show increased benefits of masking compared with diploids but volume measurements and FACScan analyses on the auxotrophic strains indicated that all treated tetraploid strains decreased in ploidy level and that some of the treated haploid lines increased in ploidy level. Results from these experiments confirm that while masking deleterious mutations provides an immediate advantage to higher ploidy levels in the presence of mutagens, selection is extremely efficient at removing induced mutations, leading growth rates to increase rapidly over time at all ploidy levels. Furthermore, ploidy level is itself a mutable trait in the presence of EMS, with both haploids and tetraploids often evolving towards diploidy (the ancestral state of S. cerevisiae) during the course of the experiment. PMID- 11279835 TI - Multi-trait QTL mapping in barley using multivariate regression. AB - Many studies of QTL locations record several different traits on the same population, but most analyses look at this information on a trait-by-trait basis. In this paper we show how the regression approach to QTL mapping of Haley & Knott (1992) may be extended to a multi-trait analysis via multivariate regression, easily programmed in statistical packages. A procedure for identifying QTL locations using forward selection and bootstrapping is proposed. The method is applied to examine the locations for QTLs for six yield characters (the number of fertile stems, the grain number of the main stem, the main stem grain weight, the single plant yield, the plot yield and the thousand grain weight) in a doubled haploid population of spring barley. Several chromosomal locations with effects on more than one trait are found. The method is also suitable for examining a single trait measured in different years or environments, and is used here to examine data on heading date, a highly heritable trait, and plot yield, a trait with moderate heritability and showing QTL-environment interactions. PMID- 11279836 TI - Evaluation of the effects of exercise and a mild hypocaloric diet on cardiovascular risk factors in obese subjects. AB - A weight reduction program to improve cardiovascular risk factors was implemented in obese subjects. The program consisted of exercise training corresponding to the anaerobic threshold (AT) and a mild hypocaloric diet for 12 weeks. In this program, we evaluated the effects of a combination of exercise training and a diet on cardiovascular risk factors such as obesity, dyslipidemia, and poor exercise performance in obese subjects. In addition, we also evaluated the independent effects of exercise training and dietary modification. For this purpose, we adopted a relative training time and a diet score. A relative training time was calculated as the number of times that the subject performed exercises divided by all of the training sessions scheduled, and the diet score was calculated from information which each subject provided on a self-assessment questionnaire. Twenty three obese subjects (Age: 24-54 years old, 19 men and 4 women, body mass index (BMI) > 26 kg/m2) participated in this study. After the 12 week intervention, the mean reductions in body weight, body mass index and body fat were 4.7 kg, 1.7 kg/m2 and 2.9%, respectively (P < 0.0001). The % change in body weight was significantly associated with the diet score and with the relative training time. The mean reductions in total cholesterol, triglyceride and low density lipoprotein cholesterol were 21 mg/dl (P < 0.002), 34 mg/dl (P < 0.01) and 15.9 mg/dl (P < 0.01), respectively, and the % change in triglyceride was significantly associated with the diet score (P = 0.0056) and tended to correlate with the relative training time (P = 0.0596). Oxygen uptake at AT and at peak exercise were increased from 14.1 +/- 1.6 to 16.0 +/- 3.1 ml/min/kg (P < 0.005) and from 26.3 +/- 4.8 to 28.4 +/- 4.9 ml/min/kg (P < 0.002), respectively. A combination of aerobic exercise and a mild hypocaloric diet significantly contributed not only to weight loss but also to the improvement of dyslipidemia and exercise performance, but either hypocaloric diet or mild exercise independently did less. The diet score and the relative training time were useful for evaluating separately dietary modification and the quantity of exercise. PMID- 11279837 TI - Occupational factors contributing to low self-esteem in registered nurses and licensed practical nurses: a multivariate analysis. AB - The present study examines job-related factors leading to low self-esteem in nurses. The lowering of self-esteem suggests that such nurses had difficulty in fully accepting themselves and their circumstances. Subjects were registered nurses (RN) and licensed practical nurses (LPN) at hospitals, and unemployed registered nurses (UEN) seeking employment. Questionnaires were provided at 53 hospitals and a Nurse Bank in Kanagawa Prefecture. The responses of 552 RN, 146 LPN and 433 UEN were analyzed. Questions were asked about personal life, past or present nursing experience, working conditions, nursing skills, satisfaction with work performance and self-esteem. Factors giving rise to low self-esteem were determined using logistic regression analysis and logistic discriminant analysis. Employment status and qualifications were determined to be the most important factors determining the self-esteem of nurses. The next most important factors were 'a limited number of years of experience (less than five years)' and 'dissatisfaction with discretion and responsibility as a nurse' (P < 0.01). Adjusted odds ratio for a reduction in self-esteem for LPN was 4.07 times higher than for UEN, and 2.2 times higher than for RN by logistic regression analysis. LPN are treated as unskilled workers, and thus significant differences were apparent in their performance of certain job tasks. These differences were analyzed using discriminant analysis, and were referred to as follows, 1: Advanced assessment skills, 2: Advanced technical skills, 3: Advanced communication skills, and 4: Nursing plan and documentation (positive discrimination rate was 70.8%). Job dissatisfaction is closely associated with the level of professional training. Continuous education and a feedback system for various levels of nurses are needed. PMID- 11279838 TI - Enantioselective metabolism of propanolol in isolated hepatocytes prepared from untreated, PB- or 3-MC-pretreated rats. AB - The present paper describes the metabolism of a chiral drug propranolol (PL) using isolated hepatocytes freshly prepared from untreated, PB- or 3-MC pretreated rats. In order to examine not only the existence of enantioselectivity but also the effect of enzyme inducer (PB or 3-MC) on PL metabolism, 500 microM PL (RS-PL, R(+)-PL or S(-)-PL) was incubated at 37 degrees C using 8 x 10(6) cells/ml isolated rat hepatocytes. Then, the elimination amount of PL and the formation amounts of eight kinds of the metabolites including ring hydroxylated metabolites (4-OH-, 5-OH- and 7-OH-PL) and side chain metabolites (NDP, AcNDP, PGL, NLA and NAA) were simultaneously determined by HPLC. By 3-MC- and PB pretreatment, a significant increase was noticed in PL elimination and also in the formation of PL metabolites, NDP, NLA and NAA. Furthermore, the presence of enantioselectivity was observed, i.e. the substrate R(+)-PL was always eliminated faster than the substrate S(-)-PL. Regarding the metabolite formation, NDP, AcNDP and NAA were dominantly produced from R(+)-PL, and NLA, PGL and the ring hydroxylated metabolites from S(-)-PL. In all cases of PL elimination and the metabolite formation, the amounts of the metabolites derived from RS-PL indicated the mean values of the respective amounts derived from R(+)-PL and S (-)-PL. Using the three kinds of isolated rat hepatocytes mentioned above, the kinetic parameters of NDP-formation at 37 degrees C for 10 min were calculated using RS PL, R(+)-PL or S(-)-PL as a substrate. From the pseudo values of V'max/K'm (microliter/min.8 x 10(6) cells), the easiest formation of NDP from R(+)-PL was observed in the rat hepatocyte system pretreated with 3-MC. PMID- 11279839 TI - Comparison of three in vitro assay systems used for assessing cytotoxic effect of heavy metals on cultured human keratinocytes. AB - The cell viability assay using cultured cells is of great advantage to elucidate the biological effect of potentially toxic substances. Recently, a novel assay system, Tetracolor One cell proliferation assay (Seikagaku Co., Tokyo, Japan), has been developed. In this report, we compare the results of the Tetracolor One assay regarding the cytotoxic effect of three heavy metal salts on cultured adult keratinocytes to those of the neutral red dye uptake assay and the MTT eluted stain assay. In this study, these three methods showed almost similar results. Compared to the other two methods, however, the Tetracolor One assay, which requires only one-step procedure before spectrophotometric measurement, is easier to use, and errors in measurement, which may be produced through the multistep procedure, are much less in this assay. Therefore, we believe that the Tetracolor One assay system is useful for assessing the cytotoxic effect of heavy metals on cultured human keratinocytes. PMID- 11279840 TI - [Treatment and results of interstitial lung diseases in video assisted thoracoscopic lung biopsy]. AB - In order to establish treatment of interstitial lung diseases in video assisted thoracoscopic lung biopsy, we retrospectively reviewed our experiences. The present study included 7 patients with a mean age of 46.4, range from 24 to 61, who were treated at our department from 1996 through 1999. They were 5 men and 2 women. The pathologic diagnosis was nonspecific interstitial pneumonia in 3 patients who responded to steroid therapy. Three other patients had usual interstitial pneumonia. One patient had lymphocytic interstitial pneumonia. No complications occurred. The results indicate that video assisted thoracoscopic lung biopsy is an effective and safe way to diagnose interstitial lung diseases. PMID- 11279841 TI - [Long-term survival of hormone-refractory prostate cancer: a case report]. AB - Although prostate cancer initially responds well to endocrine therapy, it becomes resistant to the therapy a few years later, and is called hormone-refractory cancer. In general, hormone-refractory prostate cancer is resistant to all kinds of therapy and the prognosis is extremely poor. Here we report an unusual case of a person with hormone-refractory prostate cancer, who has been surviving for more than 5 years after being diagnosed as having this type of cancer. A 75-year-old man was diagnosed with prostate cancer (poorly differentiated adenocarcinoma, T3 N0 M1, stage D2) and initial endocrine therapy combined with castration and estrogen was effective. Four years later, the tumor marker of prostate-specific antigen (PSA) increased and the cancer was thought to be hormone-independent, refractory state. Alteration of antiandrogen from chlormadinone acetate to flutamide was effective and PSA was kept at low levels for 6 months. When PSA rose again, we started oral chemotherapy with tegafur.uracil. PSA decreased to normal range (complete response) and remained stable for 10 months. After that, a rapid increase of PSA was controlled for 7 months by oral chemotherapy with estramustine phosphate sodium and VP-16. This case indicates that alteration of antiandrogens or oral chemotherapy may be useful in some cases with hormone refractory prostate cancer. PMID- 11279843 TI - [Occupationally induced nitric acid and sulfuric acid burns: an analysis of 2 patients from the aspect of occupational health]. AB - We report two patients who suffered from acid burns while working in chemical factories. Case 1: a 44-year-old man who received burn induced by nitric acid on the face and extremities. Despite his protecting facial mask, he was exposed to nitric acid on his face through a gap between the mask and skin surface. Nitric acid was also sprinkled on his scalp which was not covered by a helmet or a protecting device. In addition, he suffered from acid burn on the right scapular region, the right upper arm, and the lower extremities through the work clothes. Case 2: a 26-year-old man who suffered from sulfuric acid burn on the forearms. Both patients were accidentally exposed to acids while they filled tanks with the acids through a hose. Following the manual of the factories, they washed the exposed skin with water for more than 15 minutes after the exposure. Although they recovered without any serious sequel, there remained partial deep tissue destruction of the skin. We reviewed these two cases from the aspect of industrial medicine, and proposed the following three points for improvement in the workplace to prevent accidental acid burns. 1) re-education or enlightenment activities for the well-experienced workers to avoid negligence to the danger of strong acid. 2) recommendation to take a complete shower to avoid overlooking of unaware acid injury. 3) improvement in the protecting facial mask. In addition, clinicians who examine acid-burn patients should not pass over the presence of deep ulcers lying behind the thick crust on the injured area. PMID- 11279842 TI - [A case of pemphigus foliaceus associated with bullous impetigo successfully treated with tetracycline and nicotinamide]. AB - A 50-year-old Japanese woman visited our clinic, complaining of generalized erythema with painful erosions and bullae. The histopathological findings of the skin lesion suggested development of impetigo. Gentamycin-resistant Staphylococcus aureus was detected by the bacterial culture examination from the impetiginous bullae. A direct immunofluorescence study of the lesion showed an intercellular deposition of IgG and C3 in the upper epidermis. We diagnosed this case as pemphigus foliaceus associated with bullous impetigo. A combined oral administration of tetracycline (200 mg/day) and nicotinamide (1200 mg/day) for 3 weeks was successful. In Japan, patients with moderate to severe symptoms of pemphigus foliaceus are usually treated with oral steroid therapy. To our knowledge, however, there is no reported pemphigus case which has been successfully treated only with tetracycline and nicotinamide. PMID- 11279844 TI - [Report from the 8th UOEH Meeting of Gastrointestinal Image Diagnosis]. PMID- 11279845 TI - [Report from 11th UOEH Meeting of Gastrointestinal Image Diagnosis]. PMID- 11279846 TI - Drugs, vitamins, and gender. PMID- 11279847 TI - The value of a legal perspective on medicine. PMID- 11279848 TI - Gender and dermatology. AB - Women are the primary focus of dermatologic care in the United States. The awareness of differences in cutaneous biology between the sexes is vital to appropriate diagnosis and treatment of patients. Although research directed at women's health has become a greater priority in recent years, there has been very little gender-specific research in dermatology. This paper reviews the available data pertaining to dissimilarities in male and female cutaneous biology. PMID- 11279849 TI - Sex differences and the HPA axis: implications for psychiatric disease. AB - Depression is twice as common in women as in men. Given that stress plays a major role in the onset of this disorder, sex differences in the response to stress could influence women's vulnerability to depression. The data presented suggest that greater stress responsiveness in women may be influenced by ovarian steroids. This greater stress responsiveness and resistance to glucocorticoid feedback may be one biological factor contributing to the greater incidence of depression in women. PMID- 11279850 TI - Gender differences in pain sensitivity and responses to analgesia. AB - Variations in pain sensitivity between men and women have been reported in numerous studies. Women have more severe and longer lasting pain than men. The goal of this review is to summarize the current state of knowledge with respect to the sexual dimorphism of surgical pain and its management. This assessment excludes gender-specific pain conditions that are exclusively found in women (e.g., labor pain and surgical pain related to gynecologic procedures), focusing instead on those pain conditions that are common to men and women. PMID- 11279851 TI - Multiple ovarian hormone effects on brain structure and function. AB - Ovarian steroids have numerous effects on the brain throughout the life span, beginning during gestation and continuing into senescence. Ovarian steroids have widespread effects on catecholaminergic neurons and serotonergic pathways and on the basal forebrain cholinergic system. Another important action of ovarian hormones is the regulation of synapse turnover in the hippocampus as exhibited during the fourth or fifth day of a female rat's estrus cycle. In the human brain, estrogen replacement therapy is associated with improvements in episodic and declarative memory, which depend on the hippocampus, and is involved in the processing of emotional information. Because of their widespread influences on neuronal systems, ovarian steroids are important factors to consider in the treatment of depressive illness. PMID- 11279852 TI - Depression and health care policy. PMID- 11279853 TI - Some thoughts about women's health and its evolution. PMID- 11279854 TI - An affirmation of research and health care for women in the 21st century. PMID- 11279855 TI - Report of the Task Force on Research on Women's Health for the 21st Century: a personal overview. PMID- 11279856 TI - New health care approaches influenced by the women's health movement. PMID- 11279857 TI - Thoughts on traveling to Israel: attending the Bi-National Israel-USA conference. Promoting women's health across generations. PMID- 11279858 TI - Congressional trends and gender-specific health issues. PMID- 11279859 TI - Gender-specific prescribing: medications and the menstrual cycle. PMID- 11279860 TI - Genetic diagnosis and the abortion paradigm. PMID- 11279861 TI - Acute respiratory failure: a gender-based outcomes analysis. AB - OBJECTIVE: To determine the influence of gender on patient outcomes and the delivery of medical care during mechanical ventilation for acute respiratory failure. DESIGN: A prospective cohort study. SETTING: Medical intensive care unit of a university-affiliated urban teaching hospital. PATIENTS: Three hundred twenty-one patients with acute respiratory failure requiring mechanical ventilation. INTERVENTIONS: Prospective patient surveillance and data collection. MEASUREMENTS AND RESULTS: One hundred six patients (33.0%) died during hospitalization. No statistically significant difference in mortality rates was observed between male and female patients (36.1% versus 30.1%; P = .252). The duration of mechanical ventilation, intensive care unit length of stay, and hospital length of stay were also similar between men and women with acute respiratory failure. The presence of the acute respiratory distress syndrome (ARDS; adjusted odds ratio [AOR] = 3.65; 95% confidence interval [CI] = 2.34 to 5.71; P = .004), the number of acquired organ system derangements (AOR = 1.36; 95% CI = 1.20 to 1.53; P = .013), the predicted hospital mortality using acute physiology and chronic health evaluation (APACHE) II scores (AOR = 1.05; 95% CI = 1.04 to 1.05; P < .001), and patient age (AOR = 1.02; 95% CI = 1.01 to 1.03; P = .016) were found to be independent determinants of hospital mortality using multiple logistic regression analysis. CONCLUSIONS: These data suggest that important gender-specific differences in outcome do not occur among patients with respiratory failure requiring mechanical ventilation. Severity of illness, patient age, and acquired organ system derangements appear to be the most important determinants of mortality for patients with acute respiratory failure, regardless of patient gender. PMID- 11279862 TI - Anxiety disorders and the emergence of sex differences in major depression. AB - OBJECTIVE: To examine the role of anxiety disorders in the development of sex differences in the risk of major depression. DESIGN: An epidemiologic study. PATIENTS: This study surveyed 1007 young adults, randomly selected from a large HMO in southeastern Michigan. METHOD: The National Institute of Mental Health's Diagnostic Interview Schedule was administered to measure major depression and specific anxiety disorders (as per Diagnostic and Statistical Manual of Mental Disorders, third edition revised, guidelines). A composite variable--"any anxiety"--was used, and age of onset was defined as the age at which the earliest anxiety disorder began. RESULTS: Prior anxiety signaled an increased risk for major depression in both sexes. Women were not more vulnerable than men to becoming depressed after an anxiety disorder. Prior anxiety disorders accounted for a considerable part of the sex differences in major depression. Controlling for prior substance use disorder did not alter the results. CONCLUSION: Women's higher rates of anxiety disorders might play a role in their higher risk of depression. Substance use disorder is not men's counterpart of anxiety in terms of its potential role in the onset of depression. Future research should address the question of women's greater risk for anxiety disorders. Intervention trials to examine whether effective treatments of anxiety disorders might reduce the risk of depression would shed light on the mechanisms that link anxiety and depression. PMID- 11279863 TI - Gender similarities and differences in the course of depression. AB - This article provides a gender-based overview of the current literature on depression. Characteristics common to both genders include depressive recurrence, chronic depression associated with an increased risk for suicide, and comorbid disorder complications; differences include substance abuse or dependence, completed suicides, rapid cycling, and seasonality. There are several inconsistencies regarding gender differences in the course of major depression- the time to recovery from episodes, the time to recurrence, the likelihood of becoming chronically depressed, and the severity of episodes. PMID- 11279864 TI - Gender and drug abuse research. PMID- 11279865 TI - Thoughts on genetic engineering and the fountain of youth. PMID- 11279866 TI - A review of gender-specific legislation, 1999. PMID- 11279867 TI - Lessons (re)learned from cystic fibrosis carrier screening. PMID- 11279868 TI - Oral contraceptive therapy in women: drug interactions and unwanted outcomes. AB - Oral contraceptives are highly effective contraceptive agents used worldwide that have a relatively low incidence of adverse side effects. Drug interactions that decrease the efficacy of contraception or that limit the efficacy of concomitant medications can occur. The most frequent clinically encountered interactions involve antibiotics or anticonvulsants. The incidence of adverse thrombotic events is low and can be further minimized by the use of low estrogen doses; avoidance of smoking or not prescribing oral contraceptives in smokers; and avoidance of exposure in genotypes at high risk for thrombosis (Leiden Factor V mutation). PMID- 11279869 TI - Gonadotropin modulation of interleukin-1 secretion. AB - OBJECTIVE: To determine the influence of pituitary gonadotropins, which increase dramatically in concentration at ovulation and in the early years of the postmenopausal transition, on inflammatory cytokine production. DESIGN: Cross sectional population sampling, in vitro experimentation. PARTICIPANTS: Healthy subjects, including six men, five women between the ages of 18 and 35 years, and four women who were four to 20 years past menopause. METHOD: Isolated peripheral blood mononuclear cells were incubated with physiological concentrations of follicle-stimulating hormone (FSH) and luteinizing hormone (LH). The concentrations of interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6) secreted into the supernatants were measured by enzyme-linked immunosorbent assays. RESULTS: Under basal conditions, FSH stimulated IL-1 beta secretion by cells isolated from women in the follicular phase. Under conditions of cellular activation with bacterial lipopolysaccharide, FSH and LH interacted to inhibit IL 1 beta secretion by cells isolated from all groups. The gonadotropins had no significant influence on IL-6 secretion regardless of donor group or cellular activation state. CONCLUSION: The results of this study support the concept that gonadotropins may contribute to the changes in IL-1 beta secretion that occur at the periovulatory and postmenopausal periods. PMID- 11279870 TI - Optimal management of dyslipidemia in women and men. AB - Relatively few studies have examined the clinical effectiveness of cholesterol lowering drugs in women as compared with men. Most clinical trials do indicate that cholesterol lowering reduces clinical events equally effectively in both genders. However, high-density lipoprotein cholesterol and triglyceride levels may have more prognostic significance in women. The recent finding of a higher risk of death in younger women as compared with younger men who sustain a heart attack, combined with the high proportion of morbidity and mortality due to CHD in older women, emphasizes the need for a better understanding of heart disease in women. This review explores the similarities and differences between women and men regarding the management of dyslipidemia. PMID- 11279871 TI - The carcinogenic and toxic effects of tobacco smoke: are women particularly susceptible? AB - Smoking is the leading preventable cause of premature death and disability in Canadian women. Lung cancer, ischemic heart disease, chronic obstructive lung disease, and stroke account for two-thirds or more of the smoking-attributable deaths in women. Lung cancer now exceeds breast cancer as the leading cause of cancer death in women, and both incidence and mortality rates continue to climb. Strong evidence suggests that for the same number of cigarettes smoked, women are more susceptible than men to the carcinogenic effects on their lungs. Evidence also is growing that lung function in women is more adversely affected by smoking and that smoking may be a stronger risk factor for myocardial infarction in women than it is in men. More research into the mechanisms underlying these gender related susceptibilities is needed. Policies and programs to prevent girls from starting to smoke and to facilitate quitting in women of all ages must be public health priorities. PMID- 11279873 TI - Emerging manifestations of HIV/AIDS: is it the virus or the antiviral therapy? PMID- 11279872 TI - Psychosocial causes of depression. AB - Research has consistently determined that women have higher rates of depression than men. It is possible that the sex ratio of depression incidence varies according to time and place, and that this variation is a reflection of women's variable social circumstances. However, further research is needed to determine a convincing explanation for women's greater susceptibility to depression. PMID- 11279874 TI - Issues in antiretroviral therapy: when to start? PMID- 11279875 TI - Treatment adherence improves outcomes and manages costs. AB - Our experience with other chronic diseases, such as hypertension, diabetes, and asthma, has shown that adherence to treatment over time is about 50%. In HIV treatment, a significantly higher rate of adherence (i.e., 95% or greater) is required to achieve good outcomes. HAART is effective and cost-effective. Even with the high cost of antiretroviral drugs, the decrease in hospital utilization in addition to improved quality of life with HAART more than offsets the increased cost of drugs. This cost shifting from hospital utilization has been shown to result in a decrease of total monthly costs of care in many settings. In addition to decreased mortality and cost savings from decreased hospital utilization associated with HAART, the appropriate use of expensive antiretroviral drugs and the resultant reduction in antiretroviral resistance can save lives and money over the long term. However, we know that the performance of drugs in clinical trials is not always borne out in today's real world of ambulatory HIV care, underscoring the need for treatment adherence strategies in the HAART era. Our understanding of what improves adherence to antiretroviral treatment is still incomplete. However, there are a number of approaches that address the patient, the provider/multidisciplinary team, and the treatment regimen itself. The dedicated TAC, while not the only solution, has been shown to be an effective team member and a solution worth considering in managed care settings. When added to the costs of today's care, this team member should still prove cost-effective in the final analysis. PMID- 11279876 TI - Cost-effectiveness of voluntary prenatal and routine newborn HIV screening. PMID- 11279877 TI - No adverse effects in infants following short-course zidovudine. PMID- 11279878 TI - Adverse effects of HIV postexposure prophylaxis are common. PMID- 11279880 TI - Initiate HAART during primary infection for better immune reconstitution. PMID- 11279879 TI - Bristol-Myers warns of AIDS drugs' use. PMID- 11279881 TI - Use of p24 antigen screening for HIV in primary care. PMID- 11279882 TI - Gene therapy trial results released. PMID- 11279883 TI - Drug abuse aspects of HIV/AIDS and other infections. PMID- 11279884 TI - Pediatric clinical trials program for AIDS. PMID- 11279885 TI - Trial will add immunotherapeutic agent to first cycle of antiviral cocktails. PMID- 11279886 TI - Phase III hepatitis C program begins. PMID- 11279887 TI - Phase III trial in hepatitis B prevention. PMID- 11279888 TI - Use of HIV protease inhibitors as pharmacoenhancers. AB - Short- and long-term therapy with many protease inhibitors (PIs) at standard dosing may be limited by inconvenient dosing regimens, high tablet volumes, and variable drug exposure. Booster agents, such as low (or "baby") doses of ritonavir, and codosing with the nonnucleoside analogue delavirdine are increasingly used to maintain high PI trough exposures. Combinations of HIV PIs, either alone or coadministered with nucleoside analogues, have demonstrated substantial virologic and immunologic responses sustained over long periods of follow-up. This approach is rapidly becoming the standard of care with PI use. The advantages of the boosted PI approach include raising trough drug concentrations, diminishing interpatient variability, prolonging drug half-life to allow twice-daily and possibly once-daily dosing, and diminishing food requirements and tablet volume. In addition, increases in drug exposure may potentially enable inhibition of the virus in the presence of reduced sensitivity to PIs. Using a boosted PI, in the absence of comparative data on resistance profiles of boosted regimens, should be considered differently from dual-PI combinations, where there is exposure to 2 active PIs. Some physicians are now using a booster agent plus 2 PIs in the salvage therapy setting in an attempt to achieve effective, yet less toxic, concentrations of 2 PIs to overcome different resistant populations in a patient's viral quasispecies. PMID- 11279889 TI - Structured treatment interruption in HIV infection. AB - Structured (and unstructured) treatment interruptions have been evaluated during well-controlled acute and chronic HIV infection and before multidrug salvage therapy. In the first 2 instances, the rationale for this strategy is to stimulate or preserve HIV-specific CD4 T cells and broadly directed cytotoxic T lymphocyte responses. Before salvage therapy, treatment interruptions have led to the reemergence of drug-susceptible virus, at least in blood plasma. Although some evidence suggests beneficial immune stimulation with successive interruptions of therapy begun during acute infection, the long-term benefits of this strategy remain unproved in any clinical setting. The potential dangers of interrupting treatment--recrudescent acute retroviral syndrome, emergence of drug resistant virus, substantial declines in CD4 T cells, and new or recurrent opportunistic infections--are not theoretic. PMID- 11279890 TI - [Molecular biology in the study of hepatitis C virus]. PMID- 11279892 TI - [Immune cellular response in hepatitis C]. PMID- 11279891 TI - [Immune lesion of hepatitis B]. PMID- 11279893 TI - [Current treatment of hepatitis B]. PMID- 11279894 TI - [Current treatment of chronic hepatitis B]. PMID- 11279895 TI - [Treatment of pretransplant and posttransplant viral cirrhosis]. PMID- 11279896 TI - [Hepatic transplant in alcoholic hepatopathy]. PMID- 11279897 TI - [Laboratory diagnosis of hepatitis B]. PMID- 11279898 TI - [Retransplant in recurrence of hepatopathy caused by HCV]. PMID- 11279899 TI - [Hepatic transplant in primary and metastatic tumors of the liver except hepatocarcinoma]. PMID- 11279900 TI - Consensus statement on improving migraine management. PMID- 11279901 TI - Sumatriptan injection and tablets in clinical practice: results of a survey of 707 migraineurs. AB - This report from the Gothenburg Migraine Clinic in Sweden describes the results of a telephone survey of over 700 patients using open-label sumatriptan tablets or sumatriptan injection for the treatment of migraine. The information is based on the patients' cumulative experience of more than 76,000 subcutaneous injections, 56,000 tablets 100 mg, and 20,000 tablets 50 mg. The results demonstrate that sumatriptan tablets were preferred by a greater proportion of patients than sumatriptan injection. However, sumatriptan injection was perceived as the most effective dosing form by the greatest proportion of patients. Among patients who found sumatriptan injection to be the most effective dosing form, the most frequently cited reasons were efficacy and speed of action. Among patients who found sumatriptan tablets to be the most effective dosing form, the most frequently cited reasons were fewer side effects and lack of experience with other dosing forms. Approximately half of sumatriptan users indicated that sumatriptan use was associated with decreases in the numbers of days with migraine, days at work with migraine, and days off work/normal activities due to migraine. When they worked during a migraine attack, patients estimated that they were 76% effective after taking sumatriptan compared with 47% effective with other medications and 49% effective with no medication. In general, side effects were reported less frequently among sumatriptan tablet users compared with injection users. Asked to compare sumatriptan with the best migraine treatment that they had used before, 94% of patients indicated that sumatriptan was better or much better than their previous therapies. These data provide important information from the clinical practice setting to complement the data from sumatriptan clinical trials. PMID- 11279902 TI - John Graham Senior Clinicians Award Lecture. Posttraumatic migraine. AB - The term posttraumatic migraine has been used in several contexts. The purpose of this discussion is to outline the different circumstances in which migraine may follow trauma. Cases from the literature and from the Headache Unit of Montefiore Medical Center are reviewed. Although trauma may be one of many triggers of migraine, trauma is sometimes the sole or predominant precipitating factor; e.g., footballer's migraine. In the posttraumatic syndrome, some exacerbations of headache upon a background of chronic daily headache often fulfill the criteria of migraine. Trauma may trigger the first attack of migraine in a susceptible individual. Biochemical and epidemiologic studies suggest that trauma may be the main etiologic factor of migraine in some cases. Migraine may also follow trauma on the basis of chance. Differentiating the different types of posttraumatic migraine has diagnostic, therapeutic, and legal implications. PMID- 11279903 TI - Frequency of factor V Leiden in juvenile migraine with aura. AB - Patients with migraine are known to be at risk for stroke. It has been reported that in a group of patients with cerebral ischemia and the Leiden mutation of factor V, 67% had classical migraine. We have studied the frequency of this mutation in a group of Italian children and adolescents affected by migraine with aura. The Leiden mutation was detected in 2 (3.5%) of 57 patients and in 8 (3.7%) of 219 controls. The 2 patients carrying the mutation had no peculiar characteristics as compared with the rest of the migrainous population. In our study, the frequency of the Leiden mutation in patients was not different from that of controls. These data contrast with those collected in the Finnish population and in a group of northwestern Italian adult patients, but agree with results previously reported from The Netherlands. PMID- 11279904 TI - Headache associated with pituitary adenomas. AB - The objectives of this study were to analyze the characteristics of headache in patients with pituitary adenoma and to investigate the mechanisms involved. Fifty one patients (27 females and 24 males) with pituitary adenoma were examined. Nineteen (37.3%) of these patients (13 females and 6 males) had headache preoperatively. Most commonly, the headache was generalized (42.1%); overall headache was more frequent in the anterior half of the head (84.2%). Seventeen (89.5%) patients had bilateral headache. Headache was usually described as head heaviness (57.9%) and continuous (57.9%). Pulsating headache and dull pain were only reported by the female patients and were mostly intermittent. The mean age of patients with headache was younger than that of those without headache. Headache was more prevalent in patients with a prolactin-secreting adenoma (57.1%). There were no correlations between visual disturbances, hypopituitarism, tumor size, or cavernous sinus invasion and headache. Hemorrhagic pituitary adenoma in 4 (57.1%) of 7 patients did not always contribute to headache. The headache was improved after surgery in 14 (73.6%) of the 19 patients. In the male patients who survived postoperatively (5 of 6), headache was improved. PMID- 11279906 TI - Hemicrania continua with onset at an early age. AB - The authors describe the case of an adolescent who had suffered from unilateral continuous headache since she was 8 years old. Treatment with indomethacin 50 mg/day promptly relieved the pain which had affected her for the previous 9 years. To the best of our knowledge, this is the earliest onset of hemicrania continua reported. PMID- 11279905 TI - Chronic paroxysmal hemicrania presenting as otalgia with a sensation of external acoustic meatus obstruction: two cases and a pathophysiologic hypothesis. AB - OBJECTIVE: To describe two cases of chronic paroxysmal hemicrania manifested by otalgia with a sensation of external acoustic meatus obstruction and to suggest that the trigeminal-autonomic reflex is a mechanism for the sensation of ear blockage. BACKGROUND: Maximum pain in chronic paroxysmal hemicrania is most often in the ocular, temporal, maxillary, and frontal regions. It is less often located in the nuchal, occipital, and retro-orbital areas. Review of the literature on chronic paroxysmal hemicrania found no reports of pain primarily localized to the ear and associated with a sensation of external acoustic meatus obstruction. METHODS: The history, physical examination, imaging studies, and successful treatment plan in two patients with otalgia and ear fullness and a subsequent diagnosis of chronic paroxysmal hemicrania are summarized. RESULTS: The first patient was a 42-year-old woman with a 10-year history of unilateral, severe, paroxysmal otalgia occurring five times a day with a duration of 2 to 60 minutes. During an attack, the ear became erythematous and the external acoustic meatus felt obstructed. There were no other associated autonomic signs. The second patient was a 49-year-old woman with a 3-year history of unilateral, severe, paroxysmal otalgia occurring 4 to 15 times a day with a duration of 3 to 10 minutes. During an attack, her ear felt obstructed, and she noted ipsilateral eyelid edema and ptosis. Both patients quickly became pain-free after taking indomethacin and required its continued use to prevent headache recurrence. CONCLUSIONS: Chronic paroxysmal hemicrania may be manifested by otalgia with a sensation of external ear obstruction. When the otalgia is paroxysmal, unilateral, severe, frequent, and associated with autonomic signs, one should consider the diagnosis of chronic paroxysmal hemicrania, especially because of the prompt response to indomethacin. The most important feature to consider when making the diagnosis of chronic paroxysmal hemicrania is the frequent periodicity of discrete, brief attacks of unilateral cephalgia separated by pain-free intervals. It is hypothesized that the sensation of ear obstruction in these patients is due to swelling of the external acoustic meatus mediated through increased blood flow by the trigeminal-autonomic reflex. PMID- 11279908 TI - Crying migraine. AB - Eighty-five percent of migraineurs report triggers which include a diverse array of internal and external factors. Crying as a trigger has been reported in two women, without details, in only one prior study. In the present report, the clinical history of two women (aged 38 and 41 years, respectively) with migraines triggered by crying are detailed. In both women, the migraines were triggered by crying associated with sadness or emotional upset. Crying when happy or due to cutting onions was not a trigger. Only in the second patient was crying during a sad movie or theatrical production also a trigger. Crying may be a common underrecognized migraine trigger. PMID- 11279907 TI - Extratrigeminal episodic paroxysmal hemicrania. Further clinical evidence of functionally relevant brain stem connections. AB - A woman, aged 59 years, developed a constant, left, occipital headache associated with episodes of discrete exacerbations occurring three to five times daily for 3 days, each lasting 15 to 20 minutes, and associated with left ptosis, conjunctival injection, and redness of the left ear. Pain-free remissions, which usually lasted 2 weeks, ceased after a mild neck injury, but the headaches responded promptly to indomethacin. This case, illustrating a transition from an occipital episodic to chronic paroxysmal hemicrania, is discussed as a variation of the trigeminal-autonomic cephalalgias. PMID- 11279909 TI - Cerebral metastasis presenting as Valsalva-induced migraine with aura. PMID- 11279910 TI - Circumstances of onset of chronic headache in patients attending a specialty practice. AB - In a data base of 1013 patients with chronic headache, 44.1% reported the onset of headaches before the age of 20 (preadult-onset headache) and 55.9% at or after that age (adult-onset headache). Circumstances of headache onset were mentioned significantly more frequently in the latter than in the former group (8.5% versus 38.2%, P < .001). The 10 most common circumstances of headache onset in adult onset chronic headache, accounting for 87.9% of all circumstances mentioned, are further discussed. The 3 most common factors, accounting for 58.4% of all circumstances, were head, neck, or back injury; stress; and pregnancy. Illnesses, influenzalike and otherwise, and estrogen administration for birth control or hormone replacement shared the fourth place. It is concluded that in the etiology of chronic headache, other than genetic factors should be considered as well. PMID- 11279911 TI - Neuroimaging diagnosis of Tolosa-Hunt syndrome: MRI contribution. AB - OBJECTIVE: To present our experience in the neuroradiological diagnosis of six patients with Tolosa-Hunt syndrome. METHODS: Computerized tomograms and MRI, with and without contrast enhancement, of the cranium and orbits of patients fulfilling IHS criteria for the diagnosis of Tolosa-Hunt syndrome were analyzed. RESULTS: Standard CT scan, with and without contrast enhancement, disclosed an enlarged cavernous sinus in one patient and was normal in the remaining five. In comparison, MRI was clearly abnormal in the four patients on whom it was performed, showing a convex enlargement of the symptomatic cavernous sinus by an abnormal tissue isointense with gray matter on short TR/TE images and iso hypointense on long TR/TE scans. This abnormal tissue markedly increased in signal intensity after contrast injection and, in two patients, extended into contiguous regions, mainly the orbital apex and subtemporal fossa ipsilaterally. One patient had follow-up studies after successful treatment with corticosteroids. Although diminished in size, the abnormal tissue was still visible on MRI after 3 months of treatment and only disappeared after 6 months of treatment. CONCLUSIONS: These MRI findings help in the differential diagnosis of the Tolosa-Hunt syndrome from conditions such as meningioma, lymphoma, and sarcoidosis, as well as confirming the similarities of the Tolosa-Hunt syndrome and orbital pseudotumor. In the presence of painful ophthalmoplegia, the finding by MRI of cavernous sinus enlargement, with the herein described signal and extension characteristics and slow resolution with corticosteroid treatment, is highly suggestive of the Tolosa-Hunt syndrome. PMID- 11279912 TI - Effect of guided imagery on quality of life for patients with chronic tension type headache. AB - OBJECTIVE: To determine the effect of adjuvant guided imagery on patients with chronic tension-type headache. BACKGROUND: Management of chronic tension-type headache often requires a combination of pharmacological and nonpharmacological therapies. Guided imagery is a relaxation technique based on visualizing pleasant images and body awareness. METHODS: One hundred twenty-nine patients with chronic tension-type headache completed the Headache Disability Inventory and the Medical Outcomes Study Short Form (SF-36) at their initial visit to a specialty headache center and again 1 month after the visit. In addition to individualized headache therapy, patients listened to a guided imagery audiocassette tape daily for the month. One hundred thirty-one control subjects received individualized therapy without guided imagery. RESULTS: Controls and the patients who listened to the guided imagery tape improved in headache frequency, headache severity, patient global assessment, quality of life, and disability caused by headache. More guided imagery patients (21.7%) than controls (7.6%) reported that their headaches were much better (P = .004). The guided imagery patients had significantly more improvement than the controls in three of the SF-36 domains: bodily pain (95% CI; guided imagery patients 11.0, controls 0.2), vitality (95% CI; guided imagery patients 10.9, controls 1.7), and mental health (95% CI; guided imagery patients 7.8, controls 0.4). CONCLUSIONS: Guided imagery is an effective adjunct therapy for the management of chronic tension-type headache. PMID- 11279913 TI - Precipitating factors of headache. A prospective study in a national control matched survey in migraineurs and nonmigraineurs. AB - Prospective studies of precipitating factors in migraine are rare. Mig Access is a national control-matched survey conducted to evaluate the access of migraineurs to health care in France. This study allowed us to screen prospectively some precipitating factors of headache in migraineurs and in nonmigraineurs. Three hundred eighty-five migraineurs (group 1) and 313 nonmigraineurs (group 2) kept a diary for a 3-month period (a total of 35,805 day in group 1 and 29,109 days in group 2). Precipitating factors were reported for each headache period. Headache intensity was self-assessed during each headache period using a visual analog scale of 0 to 100. Headache was reported on 4274 days (12%) in group 1 and on 602 days (2%) in group 2. Headache intensity was greater in group 1 (39 +/- 20 versus 32 +/- 19, P < .05). The most frequent precipitating factors (reported at least once by more than 10% of subjects [range 18% to 80%] in both groups) were fatigue and/or sleep, stress, food and/or drinks, menstruation, heat/cold/weather, and infections in both groups. All these factors except infections were reported to cause headache more frequently in migraineurs than in nonmigraineurs. Mean intensity of headache related to fatigue and/or sleep, stress, food and/or drinks, hot/cold weather, and menstruation varied from 37 to 43 in migraineurs and from 29 to 35 in nonmigraineurs. Headache with the highest mean intensity was due to infections in the two groups (47 +/- 20 in group 1, 45 +/- 23 in group 2). Our results support that endogenous factors are the most frequent triggers of headache in migraineurs. The most frequent precipitating factors of headache appear identical in migraineurs and in nonmigraineurs. Our results suggest that similar triggers could precipitate headache of different type in these two populations. PMID- 11279914 TI - Sudden onset of severe headache associated with polycythemia: hyperdense middle cerebral arteries demonstrated by cranial computed tomography. PMID- 11279915 TI - Comparison of rizatriptan and sumatriptan: a reply to O'Quinn et al. PMID- 11279916 TI - Late-life (migrainous) scintillating zigzags without headache: one person's 27 year experience. AB - BACKGROUND: Episodes of typical scintillating zigzags similar to the visual aura of migraine may occur without headache, especially after the age of 50. There is no record of one individual's long-term experience. PRESENT MATERIAL: This paper is a personal account of all 41 episodes of scintillating zigzags that occurred between the ages of 59 and 85. Observations were made on several aspects of the visual event itself--evolution, pattern, scotoma, coloration, duration, lateralization, etc. The chronological distribution of spells, circumstances of occurrence, time of day, the season, etc were noted. The study was observational in nature. Documentation of this experience could be of value clinically and scientifically. RESULTS: The spells occurred irregularly, unrelated to season, time of day, activity at onset, diet, and temperamental state. The characteristic appearance was a flickering zigzag line that began centrally and migrated to the periphery. The display, which was stereotyped, was achromatic. The average duration was 15 minutes. Both visual fields were equally involved, never at the same time. The details of the scintillating zigzag line are actually complex and an account of the observations of others and of the author is included here. PMID- 11279917 TI - Weekend headache: a possible role of work and life-style. AB - The purpose of our study was to determine whether or not patients reporting weekend headache exhibit distinctive features in their work habits, family life, and leisure on workdays and on weekends as compared to other headache sufferers, and whether or not they are inclined to change their living habits at the weekend. The study was done on an initial sample of 50 patients referred to the University of Parma Headache Centre between October 1996 and April 1997. These patients completed a specially designed questionnaire which, in addition to demographics, contained specific questions relevant to the subject matter being investigated. They were also given a diary which they had to complete for 8 consecutive weeks in order to determine the actual frequency of headache attacks over different days of the week. The questionnaire data were only analyzed for the 38 women in the sample, because there were too few male controls for an accurate comparison with weekend headache sufferers. Among the women with weekend headache, work habits, family life, and leisure were such as to suggest a possible increase in stress and frustration on weekends, which might have made them perceive the headaches occurring on Saturdays and Sundays as more severe. No changes were found in the intake of substances such as coffee and alcohol, nor in cigarette smoking over the different days of the week. Finally, analysis of the diaries showed an increased frequency of headache attacks on weekends only among the men, which seems to corroborate the hypothesis of weekend headache as a disorder typically affecting men. PMID- 11279918 TI - Headache and cardiovascular risk factors: positive association with hypertension. AB - The study analyzes the prevalence of cardiovascular risk factors in 1343 patients with severe headache (399 men and 944 women), aged 15 to 64 years; analyses were controlled for sex, age, and type and frequency of headache. Prevalence of various forms of headache was different between men and women. Age and days per year with headache were significantly different among various forms of headache. For men and women with headache, age directly related to prevalence of hypertension, hypercholesterolemia, and obesity. Due to low prevalence, analyses by age were not done for diabetes mellitus. For cigarette smoking, prevalence was not related to age in men, but was inversely related to age in women. With control for age, prevalence of cardiovascular risk factors was not significantly different among patients with different forms of headache, except for cluster headache. Among men with cluster headache, prevalence was high for cigarette smoking, but low for hypercholesterolemia. With control for age, days per year with headache did not relate to prevalence of cardiovascular risk factors except for cigarette smoking in men. Compared to data for a population sample used as control, patients with headache had higher prevalence of hypertension in both sexes, independent of age (odds ratio 1.51, 95% confidence interval 1.28 to 1.80); the difference between patients with headache and the control population was lower with increasing age. The high prevalence of hypertension among patients with headache was not due to overweight. The data indicate that headache is significantly associated with hypertension, but not with other cardiovascular risk factors. PMID- 11279919 TI - Tension-type headache: pain, fatigue, tension, and EMG responses to mental activation. AB - Twenty patients with tension-type headache (14 chronic and 6 episodic) and 20 group-matched controls were selected for this study. They participated in a 1 hour, complex, two-choice, reaction-time test, as well as 5-minute pretest and 20 minute posttest periods. Subjects reported any pain in the forehead, temples, neck, and shoulders, as well as any feelings of fatigue and tension during the pretest, and every 10 minutes during the test and posttest by visual analog scales. Superficial electromyography was recorded simultaneously from positions representing the frontal and temporal muscles, neck (mostly splenius), and trapezius muscles. The location of pain corresponded to the position of the electrodes, but extended over a larger area. The test provoked pain in the forehead, neck, and shoulders of patients, i.e., pain scores from these regions increased significantly during the test. The pain scores continued to increase posttest. In patients, the EMG response of the trapezius (first 10 minutes of the test) was elevated relative to pretest. In controls, only the frontal muscles showed an EMG test response. Patients showed significantly higher EMG responses than controls in the neck (whole test period) and trapezius (first 10 minutes of the test period). There were significant differences in pain and fatigue scoring between patients and controls in all three periods and in tension scoring posttest. Fatigue correlated with pain, with increasing significance for all locations examined, while tension was mainly associated with the neck pain. The meaning of the variables "tension" and "fatigue" in headache, and their association with recorded muscle activity in various regions is discussed. The EMG response of the trapezius muscle to the test is discussed in comparison with similar responses observed in patients with other pain syndromes. PMID- 11279920 TI - Alpha-dihydroergocryptine in the prophylaxis of migraine: a multicenter double blind study versus flunarizine. AB - This multicenter, double-blind, clinical study was designed to compare the efficacy and safety of alpha-dihydroergocryptine and flunarizine in the prophylaxis of migraine without aura. One hundred thirty-five patients fulfilling the diagnostic criteria of the International Headache Society were enrolled at five neurologic centers. The study design included a 1-month pretreatment phase with placebo; a 6-month, double-blind, double-dummy treatment phase with alpha dihydroergocryptine (10 mg twice daily) or flunarizine (5 mg once daily); a further 3-month follow-up phase without treatment. Efficacy was assessed using the patient's diary. Laboratory tests, vital signs, and adverse events were monitored. Analysis of covariance for repeated measures was performed on the intent-to-treat sample. Both treatments led to a significant reduction in the frequency of migraine, days with headache, and use of relief medication. Overall, 51% of those treated with alpha-dihydroergocryptine and 49% of those treated with flunarizine were responders (50% or greater reduction in attack frequency), the average percentage of reduction being 64% with alpha-dihydroergocryptine and 51% with flunarizine. There was no significant difference between the two groups in terms of incidence of adverse events; dizziness and weight gain were the most frequent observed adverse events with alpha-dihydroergocryptine and flunarizine, respectively. Based on the overall improvement in migraine parameters, alpha dihydroergocryptine can be recommended for use in migraine prophylaxis. PMID- 11279921 TI - Primary chronic daily headache: clinical and pharmacological aspects. A clinic based study in Oman. AB - This study on primary chronic daily headache was based on the 1996 proposed revision of the diagnostic criteria of the International Headache Society (IHS). To investigate the relative frequency, clinical characteristics, and associated features of primary chronic daily headache in Omani patients, 171 patients visiting the Neurology Clinic at Sultan Qaboos University Hospital were evaluated. Forty-five percent was diagnosed as suffering from primary chronic daily headache (female to male ratio, 1.7:1). Sixty-two percent suffered from transformed migraine and 34% from chronic tension-type headache. The average age across sexes was 32.3 +/- 12.3 years. A dull heavy feeling in the head was reported by 58% of patients and was associated in less than one third with associated features characteristic of migraine. All headache types shared the same trigger factors. All patients were taking medication, predominantly analgesics, at the time of their first visit. We concluded that primary chronic daily headache is very common with the relative frequency of transformed migraine being similar to that found in Mediterranean studies. Also in Oman, chronic use/overuse of analgesics and nonsteroidal anti-inflammatory drugs is a problem that coexists with primary chronic daily headache. Finally, the proposed revised IHS criteria are highly recommended as a standard classification system for this type of headache. PMID- 11279923 TI - Intranasal lidocaine to prevent headache following migraine aura. AB - OBJECTIVE: To report the consistent effect of intranasal lidocaine 4% on preventing headache following aura in one individual. BACKGROUND: A treatment that could prevent the headache which follows an aura would be an important advance in the treatment of migraine. No migraine abortive treatment has been shown to have such an effect. METHODS: A 15-year-old adolescent boy with a history of recurrent headache since aged 2, fulfilling the criteria for migraine with aura, was seen in consultation. Intranasal lidocaine 4% was used during the aura phase to prevent the headaches. RESULTS: Before using intranasal lidocaine, the patient invariably experienced a migraine following a typical visual aura. The episodes occurred approximately weekly, with a stable pattern for several years. When given during the aura, intranasal lidocaine prevented the headache following the aura, and remained successful on all but two occasions over 1 1/2 years of use (approximately 75 episodes). There was no effect on the duration of the aura itself. CONCLUSIONS: Intranasal lidocaine consistently prevented the development of headache symptoms following aura in this individual. Such an effect suggests a role for the sphenopalatine ganglion in the development of migraine pain. PMID- 11279922 TI - Coexistence of chronic paroxysmal hemicrania and benign cough headache. AB - We report on the coexistence of both chronic paroxysmal hemicrania and cough headache in a middle-aged woman. Typical chronic paroxysmal hemicrania and cough headache episodes appeared independently and responded to indomethacin. The possible pathophysiological significance of this concurrence of both types of indomethacin-responsive pain is discussed. PMID- 11279924 TI - Dramatic headache relief after sumatriptan in a patient with a pituitary macroadenoma. AB - We are reporting an interesting case of pituitary macroadenoma. The patient presented with sudden, bifrontal, pulsating headache; photophobia; and an abducens nerve palsy, due to extension of the tumor into the cavernous sinus region. The headache resolved completely after a subcutaneous injection of sumatriptan. PMID- 11279925 TI - Spinal puncture falsely blamed for postoperative headache. PMID- 11279926 TI - Tc 99m ECD-SPECT during migraine aura without headache. PMID- 11279927 TI - Denial of hospitalization for headache. PMID- 11279928 TI - Home use of Demerol for migraine. PMID- 11279929 TI - The occipital cortex is hyperexcitable in migraine: experimental evidence. AB - OBJECTIVES: Threshold for generation of magnetophosphenes has been reported to be lower in migraine. We compared the threshold for eliciting phosphenes by transcranial magnetic stimulation and the ability to visually trigger headache in a select group of individuals with migraine with and without aura to normal controls. METHODS: Transcranial magnetic stimulation was performed using the Cadwell MES-10 stimulator. A circular coil, 9.5 cm in diameter, was applied to the occipital scalp (7 cm above the inion). Stimulator intensity was increased in 10% increments until subjects reported visual phenomena or 100% intensity was reached. Stimulator intensity was then fine-tuned to determine the threshold at which phosphenes were seen. In the same subjects, visual stimulation was given in 3.0 T MRI and if a headache occurred the response was recorded. RESULTS: Fifteen subjects with migraine were compared to 8 controls. A significant proportion of the migraineurs (86.7%) developed phosphenes compared to the controls (25%) (P = .006). The probability of triggering a headache was also higher in the migraineurs (53%); no headache was triggered in the controls (P = .019). A significant correlation was found between the threshold for phosphenes on transcranial magnetic stimulation and visually triggered headache (P = .002). When only migraine was considered, there was again a significant trend (P = .084). CONCLUSIONS: There is a difference in threshold for excitability of occipital cortex in migraineurs and controls. The hyperexcitable visual cortex in migraine is predisposed to visually triggered headache. PMID- 11279930 TI - Psychiatric comorbidity is related to headache induced by chronic substance use in migraineurs. AB - Headache centers have to deal with patients suffering from headache induced by chronic substance use which is a well-recognized complication of migraine treatment. The objective of this study was to compare psychiatric comorbidity between migraineurs with and without chronic substance use: 34 migrainous inpatients with chronic substance use were compared with 34 sex-matched noncomplicated migraineurs in a case-control study. The results showed a significantly higher prevalence of major depressive disorder, panic disorder, and social phobia in the patients with a history of chronic substance use. Consistently, anxious and depressive dimensions were significantly higher in these patients. Therefore, psychiatric morbidity may be linked to chronic substance use in migraineurs. This stresses the importance of psychiatric assessment and the need for appropriate treatment in such patients. PMID- 11279931 TI - Diagnosis of migraine in children attending a pediatric headache clinic. AB - The International Headache Society (IHS) criteria for migraine are not sufficient to diagnose migraine in children. Specifically, the duration and localization of the headache are different in children and adults with migraine. This study compared the formal IHS criteria with pediatric-amended IHS criteria and IHS criteria with the duration factor removed in children younger than 18 years. In addition, the older criteria by Vahlquist and by Prensky and Sommer were also compared. Finally, clinical diagnosis of migraine was compared with IHS criteria with the duration factor removed. The study showed that many children with a shorter duration headache have migraine and also that a number of children with a very long duration of headaches still fit the diagnosis of migraine. Unilateral headache is quite uncommon. The majority of children with migraine complained of bilateral headaches. It is concluded that the IHS criteria for pediatric migraine should be revised. We suggest making the duration factor a minor criteria for migraine in children or to exclude headaches lasting longer than 72 hours only in children younger than 15 years. PMID- 11279932 TI - Prothrombotic genetic risk factors in patients with coexisting migraine and ischemic cerebrovascular disease. AB - The role of hemostatic elements in stroke has been clearly defined but several prothrombotic polymorphisms of hemostatic factors, important for other thromboembolic disorders, seem not to be very significant in stroke. Recently, the high prevalence of factor V Leiden in patients with stroke and a history of migraine has suggested an association between migraine and prothrombotic genetic risk factors. Stroke being a multifactorial disease, the aim of this study was to test whether prothrombotic tendencies increase the risk of stroke in patients with migraine. We determined the prevalence of four prothrombotic genetic risk factors (factor V R/Q 506, factor II 20210 G/A, decanucleotide insertion/deletion in the factor VII promoter, and the platelet HPA-1 alloantigen system) in 17 patients with coexisting ischemic cerebrovascular disease and migraine, 107 patients with ischemic cerebrovascular disease, 106 patients with migraine, and 202 control subjects. Genotyping for all polymorphisms analyzed in our study were performed after specific genomic polymerase chain reaction, and confirmed by single-strain conformation polymorphism analysis. In the group of patients with coexisting ischemic cerebrovascular disease and migraine, the prevalence of prothrombotic genotypes (factor V Leiden, 5.8%; factor II 20210 A, 0%; factor VII A1, 70.6%; and HPA-1b, 35.3%) was similar to that obtained in all other groups. We can conclude that the studied polymorphisms do not seem to be associated with the development of ischemic cerebrovascular disease in those patients with migraine. PMID- 11279933 TI - The first-of-Ramadan headache. AB - This study was designed to estimate the frequency and characteristics of headaches occurring on the first day of Ramadan (Moslems' fasting month) and to determine possible causes. One hundred fifty copies of a specially designed questionnaire were distributed on the second day of fasting to a random sample of hospital staff. Completed questionnaires were obtained from 116 subjects (77%). Headaches were reported by 37 (41%) of the 91 persons who had fasted as compared to 2 (8%) of those 25 who did not fast (P = .002). The headache was of tension type in 78% of the cases. Headache frequency increased with the duration of fasting and affected mainly those prone to have headaches, more particularly of the tension type and the most important exogenous-associated factor was caffeine withdrawal. Other factors such as lack of sleep, hypoglycemia, and dehydration may have been contributory in a small number of cases. A progressive reduction of caffeine consumption in the weeks preceding the month of Ramadan and a cup of strong coffee just before the start of the fast may prevent the occurrence of first-of-Ramadan headache. PMID- 11279934 TI - Epidemiology of migraine among students from randomly selected secondary schools in Lodz. AB - The purpose of the present study was to determine the prevalence of migraine among 2351 secondary school students aged 15 to 19 years. Six hundred fifty-nine students (120 males and 539 females) complained of migraine, including 148 with additional tension-type headache (mixed headache). Migraine with aura was diagnosed in 213 students (49 with mixed headache). The remaining 446 students (99 with mixed headache) had migraine without aura. In 83 students (16 with mixed headache), headaches were developing into migrainous states. In 237 students (56 with mixed headache), headaches were accompanied by dizziness. In 128 females (25 with mixed headache), interrelation between migraine and menstruation was found. Familial factors affecting the occurrence of migraine were noted in 536 students (127 with mixed headache). It was found that 28% of secondary school students aged 15 to 19 years suffer from migraine. Nine percent of them have migraine with aura and 19%, migraine without aura. The prevalence of migraine among secondary school students is about three times higher in females than in males. Migraine with tension-type headache differs from pure migraine in respect of more numerous attacks within 1 year among females, and of more frequent occurrence of migraine with sensory aura among males. PMID- 11279935 TI - Use of percutaneous electrical nerve stimulation (PENS) for treating ECT-induced headaches. AB - Five patients who experienced migrainelike attacks associated with electroconvulsive therapy (ECT) were treated using a novel nonpharmacologic therapy known as percutaneous electrical nerve stimulation (PENS). In this sham controlled preliminary evaluation, PENS therapy proved to be a useful alternative to opioid analgesics for the acute treatment and/or prevention of ECT-induced headache. PMID- 11279936 TI - Naratriptan in the prophylaxis of transformed migraine. AB - We report three patients with transformed migraine, previously refractory to a wide variety of traditional preventive pharmacologic and nonpharmacologic interventions. Naratriptan 2.5 mg given each morning, with a second tablet allowed for breakthrough headache, at least 4 hours later, demonstrated a remarkable reduction in frequency and intensity of daily headache. In addition, a subjective improvement in quality of life and restoration of functioning including a decrease in missed workdays was noted. All three patients had previously experienced good responses to sumatriptan or zolmitriptan, but were limited in frequency of use by the authors. The patients were not experiencing rebound phenomena at the onset of treatment with naratriptan. Clinical responses were noted within 3 to 7 days of initiation of treatment. Traditional risk factor analysis and screening were performed. Naratriptan was extremely well tolerated, with no cardiovascular adverse events reported or observed. Possible mechanisms of action are discussed. PMID- 11279937 TI - Fatal cardiac arrhythmia after oral sumatriptan. AB - Typically, 3% to 5% of patients experience sensations of heaviness, pressure, and tightness in the chest after administration of sumatriptan, but there is little ECG evidence of ischemia. The serious cardiovascular incidents after consuming sumatriptan have been associated mostly with the subcutaneous dosage form of this drug and with patients with underlying cardiovascular risk factors. We report a case of fatal cardiac arrhythmia in an otherwise perfectly healthy patient with migraine after consuming a single 100-mg dose of oral sumatriptan. PMID- 11279938 TI - Migrainous stroke causing bilateral anterior cerebral artery territory infarction. AB - A 38-year-old man developed bilateral anterior cerebral artery territory infarction during the course of a migraine. Magnetic resonance imaging showed bilateral ischemic lesions involving the cortex of the paramedian region of the frontal and parietal lobes, more prominent on the right. Cerebral angiography was normal. To our knowledge, this is the first report of bilateral anterior cerebral artery territory infarction from migraine. PMID- 11279939 TI - Therapeutic gain: a critique. PMID- 11279940 TI - Comprehensive/tertiary care for headache. PMID- 11279941 TI - Subacute angle-closure glaucoma as a cause of headache. PMID- 11279942 TI - Musculoskeletal abnormalities in chronic headache. PMID- 11279943 TI - Teaching aids for primary care. PMID- 11279944 TI - Headache associated with pituitary adenomas. PMID- 11279945 TI - Attenuation by butalbital of capsaicin-induced c-fos-like immunoreactivity in trigeminal nucleus caudalis. AB - We examined the effects of butalbital (30, 100, and 1000 micrograms/kg) on the number of cells expressing c-fos-like immunoreactivity (c-fos-LI), a marker of neuronal activation, within lamina I, IIo of the trigeminal nucleus caudalis and the nucleus of the solitary tract 2 hours after the intracisternal injection of capsaicin (0.1 mL; 15.25 mg/mL) or vehicle in urethane-anesthetized guinea pigs (N = 45). Robust c-fos-LI was observed within nuclei of cells in the trigeminal nucleus caudalis after capsaicin (329 +/- 35). Butalbital dose-dependently reduced the number of labeled cells to a maximum of 66% (1000 micrograms/kg intraperitoneally [i.p.], P < .01) in lamina I, IIo but not within area postrema, medial reticular nucleus, or the nucleus of the solitary tract. Pretreatment with bicuculline (30 micrograms/kg i.p.) blocked the effect of butalbital, thereby suggesting the importance of the GABAA receptor to activation involved in the transmission of nociceptive information. Our studies suggest the possibility that GABAA receptors might provide an important therapeutic target in migraine and related headache disorders. PMID- 11279946 TI - Effects of visual stimuli and a stressor on head pain. AB - This study sought to experimentally validate two reported precipitants of chronic headaches, namely, negative affect (anxiety, anger, depression) and visual disturbance (flicker, glare, eyestrain), and to investigate whether they triggered common or different physiological mechanisms. Twenty-two male and 68 female subjects (46 with migraine, 29 with tension-type headache, and 15 controls) were submitted to antecedent challenges in the laboratory which induced negative affect or visual disturbance and to a control challenge. The results demonstrated that negative affect and visual disturbance can indeed precipitate headaches, and that the physiological responses associated with these antecedents differ, but the findings were not conclusive as to whether one or more physiological mechanisms are operative. Follow-up revealed that the antecedent challenges had significant effects on headache activity 48 to 72 hours after termination. PMID- 11279947 TI - Fluoxetine for migraine prophylaxis: a double-blind trial. AB - Selective serotonin reuptake inhibitors have recently been used in the treatment of migraine. OBJECTIVE: We studied the safety and efficacy of fluoxetine in the prevention of migraine. PATIENTS: Between February 1997 and December 1997, we examined 52 patients (33 women) at the Headache Diagnosis and Therapy Service of the Second University of Naples. Ages ranged from 18 to 65 years, and all patients suffered from migraine without aura according to IHS 1988 criteria. The sample was divided into two groups: group A included 32 patients (19 women; mean age, 36.8 years [SD 12.4]) who received fluoxetine at a dosage of 20 mg per day; group B included 20 patients (14 women; mean age, 38.8 years [SD 15.6]) who received placebo. METHODS: Our study was a single-center, randomized, double blind, parallel study of fluoxetine for the prophylactic control of migraine and consisted of two phases: 30 days of pharmacological wash out and 6 months of therapy with monthly follow-up. Patients were randomly assigned to two groups: A, fluoxetine or B, placebo. At the first visit, patients provided a detailed history and underwent neurological evaluation and a Zung test for depression. No pathological values were revealed. In order to monitor symptomatology, all patients received a form for the calculation of the total pain index at monthly follow-up. RESULTS: A comparison of the total pain index between basal values (calculated during the period of wash out) and monthly follow-up (calculated monthly during the period of 6 months of the therapy) showed significant reduction (P < .05) beginning from the third month of treatment in the fluoxetine group and no significant reduction in the placebo group. CONCLUSION: Even if preliminary and to be confirmed, these data seem to support the use of fluoxetine in the treatment of migraine. PMID- 11279948 TI - An examination of the validity of the IHS classification system for migraine and tension-type headache in the college student population. AB - The validity of the International Headache Society (IHS) classification system for college-aged students with headache was examined using cluster analysis. Undergraduate college student volunteers (N = 369) underwent a structured diagnostic interview for headaches, and the sample was divided into two subsamples for purposes of replication. A hierarchical cluster analysis (Ward's method) of the headache characteristics reported by the first subsample suggested a statistically distinct three-cluster solution, and the solution was replicated using the second subsample. It appeared that one cluster was tensionlike, while the other two were migrainelike. Nonhierarchical cluster analyses (K-means) of the cases from each subsample revealed a similar pattern of a tensionlike and two migrainelike clusters. Identical three-cluster solutions were found for the second subsample both by using cluster centers from the first subsample and by clustering the cases independently, suggesting that the cluster solution was not a random finding. The IHS classification system appears to lack adequate specificity and sensitivity for college-aged students with headache who report migrainelike symptoms. Thus, the generalizability of research results using college-aged students with headache to the adult population may be questionable. PMID- 11279949 TI - Prevalence of migraine in schoolchildren and some clinical comparisons between migraine with and without aura. AB - OBJECTIVES: To determine the prevalence of migraine and its association with age, gender, and social class and to find out whether or not the headache and nonheadache characteristics differ between children with migraine, with and without aura, using the diagnostic criteria of the International Headache Society for childhood migraine. DESIGN: Population-based study in two stages comprising an initial screening questionnaire followed by telephone interviews of children with symptoms. SETTING: Eighteen kindergarten and 39 primary and secondary schools in Thessaloniki and its semiurban areas. SUBJECTS: Four thousand children, aged 4 to 15 years, representing a random sample of 5% of schoolchildren in Thessaloniki and its semiurban areas. MAIN OUTCOME MEASURES: (1) The prevalence of migraine, (2) the connection of migraine with social class, (3) differences in the occurrence of individual symptoms between migraine with and without aura. RESULTS: The results of the present study show that migraine prevalence was 6.2% (95% confidence interval [CI], 5.4 to 7.0). The estimated prevalences of migraine with and without aura were 2.8% (95% CI, 2.3 to 3.4) and 3.4% (CI, 2.8 to 4.0), respectively. The prevalence of migraine increased with age and it was found to be almost equal in boys and girls aged 7 to 9 years or younger, but in older age groups the prevalence was higher in girls than in boys. The data showed no evidence that connected migraine with social class. It also showed that except for the aura, the headache (e.g., frequency, duration, location, quality, and severity) and nonheadache (e.g., nausea, vomiting, phonophobia, and photophobia) characteristics were no different between children with migraine, with and without aura. In conclusion, our findings indicate that migraine is a common underdiagnosed cause of severe recurrent headache in children. The findings show that childhood migraine is not connected with social class and varies with age and gender, and that except for the aura, both migraine with and without aura are so similar in their headache and nonheadache clinical characteristics that a common pathogenesis is plausible. PMID- 11279950 TI - Impact of the International Headache Society criteria on the use of neuroimaging for headache diagnosis in a headache clinic. AB - OBJECTIVE: To evaluate the impact of the introduction of the International Headache Society (IHS) criteria on the use of neuroimaging for headache diagnosis in a specialist outpatient center. BACKGROUND: The general indications for neuroimaging in headache are a matter of debate. International Headache Society criteria should improve diagnostic accuracy, consequently reducing the use of expensive diagnostic procedures such as brain CT or MRI scan. METHODS: We reviewed the medical records of all 2739 new patients seen in our center from 1984 to 1996, analyzing the records of those patients who underwent neuroimaging before or after the introduction of the IHS criteria in 1988. RESULTS: There were no differences in the number of CT scans ordered in the period before (6.04%) or after (6.06%) the introduction of the IHS criteria. Only 12 scans revealed significant abnormalities, probably unrelated to headache. CONCLUSIONS: These results suggest that the yield of CT scanning patients seen in a headache clinic is very low, even when alarm signs are present notwithstanding strict adherence to dignostic criteria. An improved definition of secondary forms of headache might help to reduce this adjunctive cost to the care of patients with headache in a headache clinic. PMID- 11279951 TI - Demographics of attendees at public education seminars. AB - PURPOSE: The demographics of patients who attend public awareness seminars relating to headache have not been studied. In order to improve the presentations at these meetings, it was felt that the meeting planners should know as much as possible about the audience. METHODS: Attendees at a public awareness seminar entitled Help for Headaches were asked to respond to a series of questions using an audience response system. RESULTS: The majority of the 212 responders were women (90%) and were over the age of 50 (53%). Most felt that they had more than one type of headache (64%), were treated by a family practitioner or internist (58%), and had been denied important diagnostic or therapeutic modalities (42%). The majority were dissatisfied with current treatment (87%) and attended the meeting to find out more about new treatments (64%). Fifty-eight percent felt their physicians did not know enough about headaches. Forty-nine percent felt they had rebound headaches. CONCLUSIONS: Public awareness seminars for headache should be designed with the realization that most attendees will be women who have more than one type of headache. The majority will be interested in new and alternative therapies, and approximately half will have rebound headaches. PMID- 11279952 TI - Chronic daily bilateral headache responsive to indomethacin. AB - Indomethacin is known to be specifically effective for chronic paroxysmal hemicrania, episodic paroxysmal hemicrania, and hemicrania continua. Different forms of idiopathic stabbing headaches have also been responsive to indomethacin, but less consistently than the others. Two cases of indomethacin-responsive headache are reported. One patient presented with what appeared to be new-onset, chronic, daily, bilateral headache aggravated by coughing. Both the chronic daily headache and the exacerbations induced by coughing were suppressed with indomethacin therapy. The second patient experienced hemicrania continua responsive to indomethacin, and the response persisted even when the headache evolved into bilateral continuous pain. There may be other idiopathic primary headache disorders that are peculiarly responsive to indomethacin. When any primary headache disorder does not respond to standard therapy, a brief therapeutic trial of indomethacin is warranted. PMID- 11279953 TI - Severe acne as a side effect of propranolol and nadolol in a migraineur. AB - Beta-blockers have proven effective in the treatment of migraine. Dermatologic side effects are extremely rare. We report a patient with migraine who developed an acnelike dermatitis with two different beta-blockers with complete resolution of the acne upon discontinuation of each drug. PMID- 11279954 TI - Relief of anesthesia dolorosa with gabapentin. PMID- 11279955 TI - Droperidol analgesia in organic headache. PMID- 11279956 TI - Cardiovascular responses to pain and stress in migraine. AB - OBJECTIVE: The purpose of this study was to investigate how migrainous subjects and controls differ in their cardiovascular reactivity and recovery to a cognitive and an acute pain laboratory stressor. BACKGROUND: Prior research suggests that individuals subject to migraine may respond physiologically to pain and stress differently than controls. METHODS: Fifty-two women (26 with migraine and 26 controls) participated in a single laboratory session. Multiple cardiovascular responses to a cognitive (mental arithmetic) and an acute pain (cold pressor task) stressor were recorded with cardiac impedance methods. The cardiovascular responses measured included systolic and diastolic blood pressure, heart rate, cardiac output, stroke volume, and total peripheral resistance. RESULTS: Results indicated that the migrainous participants displayed different cardiovascular recovery patterns for total peripheral resistance, cardiac output, and stroke volume following the termination of the mental arithmetic task. CONCLUSIONS: These results support the hypothesis that individuals with migraine respond physiologically to stress differently than control individuals. PMID- 11279957 TI - Recurrent headache in adolescents: nonreferred versus clinic population. AB - OBJECTIVE: To compare headache activity, psychosocial measures, and cold pressor response between referred and nonreferred adolescents with frequent headache. DESIGN: Thirteen boys and 19 girls with a mean age of 13.4 +/- 0.9 years who had been referred to a hospital-based behavioral treatment program for recurrent headache were compared with an age- and sex-matched school-based population of nonreferred students consisting of 31 adolescents with frequent headaches and 32 adolescents with infrequent or no headaches. All subjects completed the Spielberger State-Trait Anxiety Inventory/Trait form, the Children's Depression Inventory, the Childhood Somatization Inventory, and measures of headache activity and related functional disability. Additionally, all subjects reported interval discomfort scores on a 40-second cold pressor test with arm immersion in a 10 degrees +/- 1 degree C cold water bath. RESULTS: Subjects from both headache groups reported significantly more anxiety than those with infrequent or no headaches. The school-based nonreferred adolescents reported more depressive symptoms than the clinic-based referred subjects. In addition, the latter group reported headaches of longer duration and more school days missed due to headaches than both other groups. Whereas school-based subjects and those with infrequent or no headaches reported relatively low initial cold pressor test scores and gradually reported increasing scores with time, clinic-based subjects rated their discomfort as high at the initial interval report and maintained high levels throughout the test. No differences in somatization were found among groups. CONCLUSION: Although adolescents who seek behavioral treatment for recurrent headache do not report more psychological symptoms than nonreferred adolescents with frequent headaches, they report headaches of longer duration, miss more school days due to headache, and report higher initial sustained discomfort scores to a standardized noxious stimulus. PMID- 11279958 TI - Longitudinal prospective study of headache during pregnancy and postpartum. AB - Chronic headache fluctuates in response to changes in hormonal levels. Headache generally improves with rising estrogen levels, and worsens with falling levels. Headache should, therefore, predictably improve with pregnancy and worsen postpartum. Several retrospective studies have confirmed this pattern. In this study, 49 pregnant women with chronic headache (18 with migraine, 16 with tension type, and 15 with combined migraine and tension-type) were followed prospectively. Headache activity was recorded daily throughout pregnancy and for 3 months postpartum. Overall, there was a 30% improvement in headache between the second and third trimesters for the entire sample. This was not statistically significant. Headache improved significantly for 41% of the women, with a slightly greater tendency for headache to improve in women with migraine compared to those with tension-type or combined migraine and tension-type headaches. Headache activity was not influenced by history of menstrual migraine, history of headache change with prior pregnancies, parity, or breast-feeding. In general, women reporting headache at the end of their first trimester continued to report headache throughout pregnancy and postpartum. PMID- 11279959 TI - Amino acids in the saliva of patients with migraine. AB - There are a number of hypotheses concerning the pathogenesis of migraine, but they are frequently conflicting. In addition to the vascular hypothesis, clinical data are available that excitatory amino acids may play an important role in the development of the disease. In this study, free amino acid concentrations were measured by RP-HPLC in the saliva of 23 migraineurs without aura, 14 migraineurs with aura, and 20 healthy subjects. Significantly higher concentrations of glutamic acid, serine, glycine, arginine, and tyrosine were found in the saliva samples of both groups of migraineurs relative to the control group. It is suggested that amino acids causing hyperexcitability in the central nervous system may be linked to the pathogenesis of migraine. PMID- 11279960 TI - Management of chronic daily headache utilizing a uniform treatment pathway. AB - OBJECTIVE: To determine if the use of a uniform treatment pathway might be effective in treating patients with primary chronic daily headache. METHODS: Thirty-three consecutive patients with primary chronic daily headache were managed according to a treatment pathway which involved sequential administration of divalproex sodium, amitriptyline, amitriptyline plus phenelzine, or methadone. RESULTS: Twenty-two patients (67%) reported a 50% or greater reduction in headache days per month following initiation of treatment. Most positive treatment responses (17 [77%] of 22) were attributed to divalproex sodium. CONCLUSION: Implementation of a uniform treatment pathway may result in significant clinical improvement in a sizable proportion of patients with chronic daily headache. PMID- 11279961 TI - Objective behavior associated with an "ordinary" mild headache: a surprising failure of pain onset to signal self-protective or self-regulatory behavior. AB - OBJECTIVE: The goal of this study was to determine whether onset of an "ordinary" headache initiated self-protective behavior or self-regulation, as indexed by a reduction in effort expenditure. METHODS: A nonclinical sample was employed. The ambition and performance accuracy of a headache-developing group (n = 23) and a sex-matched, headache-free group (n = 23) was compared during a series of mental arithmetic problems. Embedded within the series of math problems was a task involving recall of a stressor previously found to induce headache in many subjects. RESULTS: Onset of mild head pain did not lead to effort conservation; instead, heightened ambition appeared to characterize the headache-developing participants before as well as after headache onset. Headache-developing subjects also displayed a performance accuracy deficit. CONCLUSIONS: The data suggest unusually ambitious, effortful task engagement may contribute to the onset of mild "ordinary" headache. This possibility requires further examination under other controlled conditions as well as in the natural environment. PMID- 11279962 TI - Improvement of tension-type headache when treating wrinkles with botulinum toxin A injections. AB - Botulinum toxin A has been used to treat a spectrum of neuromuscular diseases. In recent years, it has become an accepted treatment for dynamic facial wrinkles. Following treatment of glabella and forehead wrinkles with botulinum toxin A, 9 of 134 patients coincidentally reported improvement of tension-type headache. We have retrospectively studied this group of patients in whom improvement of facial wrinkles closely paralleled improvement of tension-type headache. This observation suggests a role for muscle action in tension-type headache and a novel treatment. PMID- 11279963 TI - Central nervous system superficial siderosis, headache, and epilepsy. AB - Almost 95 cases of superficial siderosis of the central nervous system have been reported in the literature. These patients showed a clinical syndrome characterized by ataxia, deafness, pyramidal system involvement, and mental deterioration with xanthochromic cerebrospinal fluid and neuroradiological findings of hemosiderin deposits. About 30% of the patients had headache as an accompanying symptom. In the present case report, we describe a 33-year-old man with the typical clinical features of superficial siderosis, who complained, since aged 8, of a severe recurrent frontal headache often associated with loss of consciousness occurring after at least 2 hours of pain. The MRI and CSF findings were consistent with subarachnoid bleeding. In our patient, headache due to meningeal irritation by subarachnoid blood induced seizures as a probable reflex of extreme pain. Carbamazepine and nimodipine prophylaxis dramatically reduced the frequency of headaches and seizures. PMID- 11279964 TI - Ophthalmoplegic migraine with unusual features. AB - We report the details of a patient with an unusual form of ophthalmoplegic migraine resulting in permanent vertical misalignment of the affected eye. The presentation, history, and ophthalmologic examination are reported as well as disease course and follow-up complications. We review the literature on ophthalmoplegic migraine with discussion regarding typical presentation, methods of diagnosis, and other diseases which may cause diagnostic confusion. In light of current case reports on ophthalmoplegic migraine, this is the first documented example with a permanent deficit. PMID- 11279965 TI - Estrogen replacement and migraine aura. AB - OBJECTIVE: To assess the association between estrogen replacement therapy and migraine aura. BACKGROUND: Estrogen replacement therapy is increasingly used by perimenopausal and postmenopausal women for management of menopausal symptoms and for long-term protection against osteoporosis and arterial disease. There are few reports about the effects of estrogen replacement therapy on migraine. METHOD: Case reports were collected from women developing migraine aura related to use of estrogen replacement therapy. RESULTS: Four patients who developed migraine aura associated with the use of estrogen replacement therapy are described. In all cases, reducing the dose of estrogen or changing the route of delivery was associated with loss of aura. CONCLUSION: These findings suggest that high levels of estrogen in women using replacement therapy can trigger migraine aura. PMID- 11279966 TI - Warning: the excedrin migraine warning label is inadequate to warn consumers of the risk of medication rebound headache. PMID- 11279968 TI - Hearing the headache person: the dagwood sandwich. PMID- 11279967 TI - Supraorbital neuralgia. PMID- 11279969 TI - Intranasal lidocaine for migraine: a randomized trial and open-label follow-up. AB - OBJECTIVE: To study the efficacy of intranasal lidocaine for the treatment of migraine when administered by subjects in a nonclinic setting. DESIGN: A 1-month, randomized, controlled, double-blind trial, followed by a 6-month open-label follow-up. SETTING: Ambulatory subjects treating themselves outside of a medical setting. SUBJECTS: One hundred thirty-one adult subjects with migraine, diagnosed according to International Headache Society criteria, were enrolled in the study: 113 treated at least one headache in the controlled trial, and 74 treated at least one headache in the open-label phase. All subjects were members of the Kaiser Permanente Southern California Medical Care Program and were recruited at two urban medical centers. INTERVENTION: Intranasal lidocaine 4% or saline placebo 0.5 mL was dropped into the nostril on the side of the headache, or bilaterally for bilateral headache, according to study protocol. MAIN OUTCOME MEASURES: Trial: percent of headaches relieved to mild or none at 15 minutes and relapse of headache within 24 hours. Open-label: percent of headaches relieved to mild or none at 15 and 30 minutes and relapse within 24 hours. RESULTS: In the controlled trial, headache was relieved within 15 minutes in 34 (35.8%) of 95 subjects treated with 4% intranasal lidocaine compared with 8 (7.4%) of 108 subjects receiving placebo (P < .001). Headaches relapsed in 7 (20.6%) of 34 subjects treated with 4% intranasal lidocaine compared to 0 of 8 placebo subjects (P = .312). In the open-label follow-up, headaches were relieved in 129 (41.2%) of 313 episodes within 15 minutes and in 141 (57.6%) of 245 episodes after 30 minutes. Headaches relapsed in 28 (19.9%) of 140. The response did not diminish over time: 32 (62.8%) of 51 first headaches were relieved at 30 minutes and 10 (71.4%) of 14 seventh headaches were relieved. Relapse occurred in 28 (20%) [corrected] of 129 headaches at a mean time (+/- SD) of 7.4 (+/- 6.6) hours. CONCLUSION: Intranasal lidocaine 4% provides rapid relief of migraine symptoms. For those subjects who do respond, the effect does not diminish over 6-month follow-up. PMID- 11279970 TI - Sensitivity to various stimuli in primary headaches: a questionnaire study. AB - Questions about discomfort or pain produced by various stimuli (e.g., light, sound, exercise, neck movements) are currently used to differentiate between various primary headache disorders. In order to evaluate the usefulness of differences in sensitivity to physical stimuli in headache diagnosis, the answers to a questionnaire about sensitivity to various stimuli were compared in 68 patients with migraine, 45 with tension-type headache, 46 with cluster headache, and 23 patients with cervicogenic headache, and in 71 controls. Even among controls, a high proportion reported that many of these stimuli could elicit some degree of discomfort or pain. Without headache, migraineurs differed from the other patients with headache and controls mainly in their increased sensitivity to light. With headache, patients with tension-type headache were the least sensitive and migraineurs were the most sensitive to all stimuli, except for stimuli stemming from neck movements, to which patients with cervicogenic headache were most sensitive. Migraineurs also reported the highest degree of sensitivity regarding aggravation and provocation of headache. However, the most striking finding was that all patient groups, cluster headache in particular, became significantly more sensitive with headache than without headache to almost all stimulus categories. This may indicate that these headaches share important pathogenetic mechanisms. The fact that no headache had a very specific sensitivity profile may point to weaknesses of present headache classification systems. PMID- 11279971 TI - Changing pattern of headache pointing to cerebral venous thrombosis after lumbar puncture and intravenous high-dose corticosteroids. AB - OBJECTIVE: To emphasize the diagnostic importance of change in the headache pattern which pointed to cerebral venous thrombosis in two patients after lumbar puncture and high-dose intravenous methylprednisolone for suspected multiple sclerosis. RESULTS: Both patients had a diagnostic lumbar puncture for suspected multiple sclerosis and were treated with high-dose intravenous methylprednisolone. Both developed a postlumbar puncture headache that was initially postural, typical of low cerebrospinal fluid pressure. Three days later, the headache became constant, lost its postural component, and was associated with bilateral papilledema. Magnetic resonance imaging of the brain disclosed superior sagittal and lateral sinuses thrombosis. The diagnostic difficulties of such cases and the potential role of lumbar puncture and corticosteroids as risk factors for cerebral venous thrombosis are discussed. CONCLUSIONS: When a typical postdural puncture headache loses its postural component, investigations should be performed to rule out cerebral venous thrombosis, particularly in the presence of other risk factors. PMID- 11279972 TI - Cluster headache. Premonitory symptoms. AB - The emergence of symptoms which may precede by days the onset of a series of painful attacks of cluster headache is not often reported in the medical literature. In this report, four patients who described these premonitory symptoms are presented. The importance of premonitory symptoms is emphasized, for they provide a means to institute an early prophylactic therapy and the possibility of clarifying the physiopathology of this primary headache. PMID- 11279973 TI - Treatment of migraine with pulsing electromagnetic fields: a double-blind, placebo-controlled study. AB - The effect of exposure to pulsing electromagnetic fields on migraine activity was evaluated by having 42 subjects (34 women and 8 men), who met the International Headache Society's criteria for migraine, participate in a double-blind, placebo controlled study. Each subject kept a 1-month, pretreatment, baseline log of headache activity prior to being randomized to having either actual or placebo pulsing electromagnetic fields applied to their inner thighs for 1 hour per day, 5 days per week, for 2 weeks. After exposure, all subjects kept the log for at least 1 follow-up month. During the first month of follow-up, 73% of those receiving actual exposure reported decreased headaches (45% good decrease, 14% excellent decrease) compared to half of those receiving the placebo (15% worse, 20% good, 0% excellent). Ten of the 22 subjects who had actual exposure received 2 additional weeks of actual exposure after their initial 1-month follow-up. All showed decreased headache activity (50% good, 38% excellent). Thirteen subjects from the actual exposure group elected not to receive additional exposure. Twelve of them showed decreased headache activity by the second month (29% good, 43% excellent). Eight of the subjects in the placebo group elected to receive 2 weeks of actual exposure after the initial 1-month follow-up with 75% showing decreased headache activity (38% good, 38% excellent). In conclusion, exposure of the inner thighs to pulsing electromagnetic fields for at least 3 weeks is an effective, short-term intervention for migraine, but not tension headaches. PMID- 11279975 TI - MMPI changes associated with therapeutic intervention: a migraine control study. AB - The investigation of personality traits of migraineurs with the Minnesota Multiphasic Personality Inventory (MMPI) is important research, but so far has led to diverse conclusions. This study aimed to investigate the influences of treatment intervention on the personality of migraineurs. Twenty-three Chinese patients (5 men, 18 women) with migraine (2 with aura, 21 without aura) were given the Chinese edition of the MMPI, before and after treatment, and were compared with 30 nonheadache healthy control subjects (6 men, 24 women). Statistical analyses were made among the three groups. The results revealed that patients in the pretreatment group with migraine had significantly higher scores on subtests of neuroticism (hypochondriasis, depression, hysteria) and schizophrenia. After treatment, the scores on subtests of hysteria, psychasthenia, and schizophrenia were remarkably lower (P < .05); the MMPI profile of the posttreatment group was within the reference range, but the scores of the neurotic scales were still higher than those of the healthy control group (P < .01). These results suggest that after treatment, disturbances in thinking, sentiment, and behavior were eliminated, and anxiety symptoms remarkably reduced, but some "migraine personality" characteristics remained and could influence the long-term results of treatment to some extent. It is suggested that management of migraine should include psychological intervention. PMID- 11279974 TI - Histamine as a therapeutic alternative in migraine prophylaxis: a randomized, placebo-controlled, double-blind study. AB - This study was undertaken to test the efficacy of the subcutaneous administration (twice a week) of consecutively increasing doses of histamine (0.1 to 1 ng) in the prophylaxis of migraine, compared to placebo, under a controlled, double blind, clinical trial for 12 weeks. Sixty patients were selected, under criteria established by the International Headache Society (both sexes, 18 to 65 years of age, a migraine history of more than 1 year, one to six headache attacks per month), having no additional neurological or cardiovascular pathologies, and after a complete clinical and laboratory examination including computer-assisted tomography. Comparison between the groups treated with placebo (n = 30) and histamine (n = 30), on data collected for the 4th, 8th, and 12th weeks of treatment, revealed that histamine exerted a significantly (P < .0001) greater reduction (compared to placebo) in the frequency, intensity, and duration of migraine attacks, as well as on the use of rescue medication. No significant (P > .05) adverse experiences or side effects in either group developed to impede the blinding of the study or the planned order of events. We conclude that the subcutaneous administration of histamine (at very low doses) constitutes a novel and effective therapeutic approach in migraine prophylaxis, aimed at limiting excessive inflammatory responses involved in the pathophysiology of migraine through the activation of H3 receptors. PMID- 11279976 TI - Treatment of cluster headache with mirtazapine. PMID- 11279977 TI - Angiotensin II plasma levels are unaltered during spontaneous migraine attacks. PMID- 11279978 TI - Exertional vibrating hemicrania. PMID- 11279979 TI - Ten commandments of headache medicine. PMID- 11279980 TI - [Hypoglycemia risk factors. Long term complications of poorly controlled diabetics]. PMID- 11279981 TI - [New drugs for diabetes treatment. Fast-acting insulin analogues]. PMID- 11279982 TI - [Better blood sugar control in diabetics. Insulin glargin--a long-acting insulin analogue]. PMID- 11279984 TI - [Away with the needle. Noninvasive administration routes for insulin: improved quality of life for diabetics?]. PMID- 11279983 TI - [A new aspect in diabetes treatment. Results of an orally administered insulin mimetic]. PMID- 11279985 TI - [Counseling in the pharmacy. Drug consumer problems with insulin administration]. PMID- 11279986 TI - [Better quality for less money? No utopia but new future solutions in diabetes management]. PMID- 11279987 TI - A pilgrim's progress. PMID- 11279988 TI - You've been stuck. What do you do? PMID- 11279989 TI - Developmental care of children. Are nurses avoiding the issue? PMID- 11279990 TI - Adherence concerns. PMID- 11279991 TI - The lidocaine patch. PMID- 11279992 TI - Lobbying efforts gain momentum. PMID- 11279993 TI - Improved rates of compliance with hand antisepsis guidelines: a three-phase observational study. PMID- 11279994 TI - Understanding bulimia. Signs, symptoms and the human experience. PMID- 11279995 TI - Emergency. Amniotic fluid embolism. PMID- 11279996 TI - Reflections. Of moose and maggots. PMID- 11279998 TI - All for one and one for all. Using state legislation to secure federal protections. PMID- 11279997 TI - Adult pediatric patients. PMID- 11279999 TI - Professional development. Use the media. PMID- 11280000 TI - Hand stand. PMID- 11280001 TI - Kids on the move. Preventing obesity among urban children. PMID- 11280002 TI - Navigating the world of alternative medicine. Worthwhile Web sites. PMID- 11280003 TI - Know your rights. PMID- 11280004 TI - Mutational origin of new mating type specificities in flowering plants. AB - Many hermaphroditic plants avoid self-fertilization by rejecting pollen that express genetically-determined specificities in common with the pistil. Self incompatibility systems typically show extremely high genetic diversity, some maintaining hundreds of specificities. This article addresses the genetic and evolutionary mechanisms through which new mating specificities arise. Recent investigations of the genetic and physiological basis of self-incompatibility are reviewed. Two evolutionary pathways are considered: one which requires full expression of self-incompatibility in all intermediates and one in which new mating specificities arise through episodes of partial breakdown and restoration of self-incompatibility. PMID- 11280005 TI - The exceptional mitochondrial DNA system of the mussel family Mytilidae. AB - Species of the families Mytilidae (sea mussels) and Unionidae (fresh water mussels) contain two types of mitochondrial DNA (mtDNA), the F that behaves as the standard animal mtDNA and the M that is transmitted through the sperm and establishes itself only in the male gonad. The two molecules have, therefore, separate transmission routes, one through the female and the other through the male lineage. The system has been named doubly uniparental inheritance (DUI). Another important feature of sea mussels is that the sex ratio among offspring of a pair mating is determined by the female parent only. The mechanism of DUI remains unknown. One hypothesis that is consistent with all observations is that the standard maternal inheritance was modified in mussels via the evolution of a suppressor gene that is expressed during oogenesis and has two alleles, the inactive and the active allele. In the presence of the active allele in the mother's genotype the egg is supplied with a substance that interferes and the normal mechanism of elimination of sperm mitochondria. This will explain why half of mussels have the father's mtDNA and half do not, but would not explain why presence/absence of paternal mtDNA is linked with the male and female gender, respectively. To provide an explanation for this linkage, one would have to assume that there is a causal relationship between retention of paternal mtDNA and sex determination. PMID- 11280006 TI - Different domains of Sgs1 are required for mitotic and meiotic functions. AB - The SGS1 of Saccharomyces cerevisiae is a homologue for human Bloom's syndrome, Werner's syndrome, and Rothmund-Thomson's syndrome causative genes. Disruptants of SGS1 show high sensitivity to methyl methanesulfonate (MMS) and hydroxyurea, and hyper recombination phenotypes including interchromosomal homologous recombination in mitotic growth. In addition, sgs1 disruptants show poor sporulation and a reduced level of meiotic recombination as assayed by return-to growth. We examined domains of Sgs1 required for mitotic and meiotic functions of Sgs1 by transfecting variously mutated SGS1 into sgs1 disruptants. The N-terminal 1-401 amino acid region was required for complementation of MMS sensitivity and suppression of hyper heteroallelic recombinations of sgs1 disruptants in mitotic growth and for complementation of poor sporulation and of reduced meiotic recombination. Although the N-terminal 1-125 amino acid region was absolutely required for the complementation of MMS sensitivity and suppression of hyper heteroallelic recombinations in mitotic growth, it was dispensable for the meiotic functions. In contrast, the highly acidic region (400-596 amino acid) was dispensable for the mitotic functions but a deletion of this region affected the meiotic functions. The C-terminal 1271-1350 amino acid region containing a HRDC (helicase and RNaseD C-terminal) domain was dispensable for the mitotic and meiotic functions. Although DNA helicase activity of Sgs1 was not required for Sgs1 to complement the meiotic functions, a deletion of helicase motifs III-IV (842-1046 amino acid) abolished the complementing activity of Sgs1, indicating that a structurally intact helicase domain is necessary for Sgs1 to fulfill its meiotic functions. PMID- 11280007 TI - Tnr8, a foldback transposable element from rice. AB - An insertion sequence 418 bp in length was found in one member of rice retroposon p-SINE1 in Oryza glaberrima. This sequence had long terminal inverted repeats (TIRs) and is flanked by direct repeats of a 9-bp sequence at the target site, indicative that the insertion sequence is a rice transposable element, which we named Tnr8. Interestingly, each TIR sequence consisted of a unique 9-bp terminal sequence and six tandem repeats of a sequence about 30 bp in length, like the foldback transposable element first identified in Drosophila. A homology search of databases and analysis by PCR revealed that a large number of Tnr8 members with sequence variations were present in the rice genome. Some of these members were not present at given loci in several rice species with the AA genome. These findings suggest that the Tnr8 family members transposed long ago, but some appear to have mobilized after rice strains with the AA genome diverged. The Tnr8 members are thought to be involved in rearrangements of the rice genome. PMID- 11280008 TI - Differentiation within the genus Leptocarabus (excl. L. kurilensis) in the Japanese Islands as deduced from mitochondrial ND5 gene sequences (Coleoptera, Carabidae). AB - The phylogenetic trees have been constructed for the mitochondrial ND5 gene sequences from the Japanese Leptocarabus ground beetles, which contain 101 specimens collected from nearly the complete distribution ranges of them consisting of five morphological species, i.e., Leptocarabus procerulus, L. kumagaii, L. hiurai, L. kyushuensis and L. arboreus. On the trees, there are recognized two major lineages, each of which is further divided into two or more sublineages. The phylogenetic lines are geographically linked. Two or more species occur in a single lineage, and the same species appear in different lines. We suggest that transformation from one type of morphology to another took place in parallel in various periods of evolution of the Japanese Leptocarabus. From the phylogenetic tree and the dating from the nucleotide substitution rate and the geohistorical data it is inferred that the ancestry of all the Japanese Leptocarabus species was derived from a protoform of L. kyushuensis inhabited the ancient Japan area, followed by separation into two lineages after split of the Japanese Islands from the Eurasian Continent. They then propagated distribution to occupy their own habitat ranges, during which the morphological transformation took place in some lineages. PMID- 11280009 TI - Mapping of rDNA on the chromosomes of Eleusine species by fluorescence in situ hybridization. AB - Mapping of rDNA sites on the chromosomes of four diploid and two tetraploid species of Eleusine has provided valuable information on genome relationship between the species. Presence of 18S-5.8S-26S rDNA on the largest pair of the chromosomes, location of 5S rDNA at four sites on two pairs of chromosomes and presence of 18S-5.8S-26S and 5S rDNA at same location on one pair of chromosomes have clearly differentiated E. multiflora from rest of the species of Eleusine. The two tetraploid species, E. coracana and E. africana have the same number of 18S-5.8S-26S and 5S rDNA sites and located at similar position on the chromosomes. Diploid species, E. indica, E. floccifolia and E. tristachya have the same 18S-5.8S-26S sites and location on the chromosomes which also resembled with the two pairs of 18S-5.8S-26S rDNA locations in tetraploid species, E. coracana and E. africana. The 5S rDNA sites on chromosomes of E. indica and E. floccifolia were also comparable to the 5S rDNA sites of E. africana and E. coracana. The similarity of the rDNA sites and their location on chromosomes in the three diploid and two polyploid species also supports the view that genome donors to tetraploid species may be from these diploid species. PMID- 11280010 TI - Highly repetitive elements from Chinese bitterlings (genus Rhodeus, Cyprinidae). AB - We have isolated and characterized several highly repetitive DNA elements from two species of Chinese bitterlings, Rhodeus atremius suigensis and R. ocellatus ocellatus. They comprise a partly interspersed and partly tandem repetitive family of about 1.0 to 1.3 kb in length. Individual elements showed considerable length variation, but genomic Southern blotting revealed two major length groups. Their restricted presence of these elements among related species and relative copy number differences indicated rapid change of genome structure in this group of fish. The isolated elements may be useful landmarks for further chromosomal studies. PMID- 11280011 TI - The role of childhood friendships in psychological adjustment. PMID- 11280012 TI - Friendship and peer rejection as predictors of adult adjustment. PMID- 11280013 TI - Children's friendship experiences and psychological adjustment: theory and research. PMID- 11280014 TI - Children with attention-deficit/hyperactivity disorder: peer relationships and peer-oriented interventions. PMID- 11280015 TI - Peer group dynamics associated with iatrogenic effects in group interventions with high-risk young adolescents. PMID- 11280016 TI - Friendship and the worlds of childhood. PMID- 11280017 TI - Update on preanalytic variables in common tests for coagulation. PMID- 11280018 TI - Natural alternatives for the common cold: hype or hope? PMID- 11280019 TI - Use of dobutamine stress echocardiography in determination of myocardial viability. AB - Echocardiography is a versatile, accurate and readily available method for the evaluation of cardiac anatomy and function. Used in conjunction with exercise or pharmacological stress protocols, modern 2-dimensional echocardiographic techniques can readily assess regional, as well as global, left ventricular (LV) function, and provide information regarding the presence, severity and distribution of coronary artery disease (CAD). In contrast to the coronary vasodilators adenosine and dipyridamole, dobutamine infusion simulates exercise by raising myocardial oxygen consumption through its inotropic and chronotropic effects. Dobutamine is a beta 1 receptor agonist that exerts positive inotropic activity without significant alpha or beta 2 effect. This results in little effect on peripheral vascular tone and considerably less chronotropic and arrhythmogenic activity than dopamine, norepinephrine or isoproterenol. Ischemic end-points during DSE include the development of abnormalities in regional wall motion or thickening of previously normal segments or worsening in areas of baseline hypokinesis. Augmentation of wall motion in hypokinetic segments has been reported by several studies during low-dose dobutamine infusion and has been suggested as a marker of myocardial viability. PMID- 11280020 TI - Identification of genes involved in cell senescence and immortalization: potential implications for tissue ageing. AB - The limited proliferative potential of normal cells in culture, cell replicative senescence, is an accepted model for ageing at the cellular level. Tumour derived, or viral- or carcinogen-transformed cells have escaped senescence and proliferate without control (immortal). We and others have found that fusion of normal with immortal human cells yields hybrids that have regained growth control and cease division. This demonstrates that the phenotype of replicative senescence is dominant and that cells immortalize because of defects in senescence-related genes. We exploited the recessive nature of immortality and by fusing different immortal cell lines with each other identified four complementation groups for indefinite division. Immortal parental cell lines with similar senescence gene defects when fused yielded hybrids with unlimited division potential and were assigned to the same complementation group. Fusion of immortal cell lines with different gene defects resulted in complementation in the hybrids, which had limited division capability. These parental cell lines were assigned to different complementation groups. Using microcell-mediated chromosome transfer, we then demonstrated that introduction of a normal human chromosome 4 induced senescence only in immortal cell lines assigned to complementation group B. We have now cloned the gene on chromosome 4, MORF4 (mortality factor on chromosome 4). It is a member of a family of seven genes and only MORF4 and the MORF-related genes MRG15 and MRGX are expressed. The predicted protein motifs strongly suggest this is a novel family of transcription factors. We have identified interacting proteins, some of which are also novel. These genes have the potential to modulate expression of a large number of genes by chromatin remodelling. They, therefore, also have the potential to affect tissue function due to changes in expression activity during ageing. PMID- 11280021 TI - Do alterations in glutathione and iron levels contribute to pathology associated with Parkinson's disease? AB - A growing body of evidence has implicated oxidative stress as an important factor in the neuropathology associated with Parkinson's disease. Dopaminergic nigrostriatal neurons, the predominant cells lost in Parkinson's, are believed to be highly prone to oxidative damage due to the propensity for dopamine to auto oxidize and thereby produce elevated levels of hydrogen peroxide and catecholamine quinones. Hydrogen peroxide formed during this process can either be converted by iron to form highly reactive hydroxyl radicals or removed through reduction by glutathione. Glutathione can also conjugate with quinones formed during dopamine oxidation preventing them from facilitating the release of iron from the iron-storage molecule ferritin. Alterations in both iron and glutathione levels in the substantia nigra have been correlated with the neuronal degeneration accompanying Parkinson's disease but a direct causative role for either has yet to be definitively proved. We will discuss the use of genetically engineered cell and mouse lines generated in our laboratory as models to examine the role that alterations in iron and glutathione levels may play in neurodegeneration of dopaminergic neurons of the substantia nigra associated with Parkinson's disease, and how these two parameters may interact with one another to bring this about. PMID- 11280022 TI - Ageing and cancer: the telomere and telomerase connection. AB - Telomeres are repetitive DNA sequences at the ends of linear chromosomes. Telomerase, a cellular reverse transcriptase, helps stabilize telomere length in human stem, reproductive and cancer cells by adding TTAGGG repeats onto the telomeres. Each time a telomerase-negative cell divides some telomeric sequences are lost. When telomeres are short, cells enter an irreversible growth arrest state called replicative senescence. In most instances cells become senescent before they can become cancerous, thus the growth arrest induced by short telomeres may be a potent anti-cancer mechanism. Since most cancers express telomerase, maintenance of telomere stability is likely to be required for the long-term viability of tumours. Inhibition of telomerase results in gradual erosion of telomeres followed by cessation of proliferation or apoptosis, and thus may be a promising target for cancer therapy. Introduction of the telomerase catalytic protein component into telomerase-silent cells is sufficient to restore telomerase activity and extend cellular life span. However, cells with introduced telomerase are not cancer cells since they have not accumulated the other changes needed to become cancerous. This indicates that telomerase-induced telomere length manipulations may have utility for tissue engineering and for dissecting the molecular mechanisms underlying genetic diseases including cancer. PMID- 11280023 TI - Ageing and the immune system. AB - Immune system alterations during ageing are complex and pleiotropic, suggestive of remodelling or altered regulation, rather than simple immune deficiency. The most dramatic changes with age occur within the T cell compartment, the arm of the immune system that protects against pathogens and tumours, consistent with the increased incidence and severity of infection and cancer in the elderly. Indeed, autopsy studies confirm infection as the major cause of death in the very old. Increased serum levels of inflammatory mediators are another hallmark of ageing, suggestive of either regulatory defects or an ongoing attack on sub clinical neoplastic disease or infection. Qualitative changes in antibody production, including those secreted by the gut mucosal immune compartment, affect responses to foreign antigens as well as to prophylactic vaccines. Innate immunity, the first line of defence that precedes the antigen-specific T and B cell responses, also undergoes changes with age. Some of the immune effects associated with ageing are secondary to overall organismic changes, such as alterations in the viscosity of cell membranes and proteolytic cellular machinery. Evidence suggesting that immune system changes may be involved in some major age-related pathologies, such as atherosclerosis and Alzheimer's disease, will be discussed. PMID- 11280024 TI - Mechanisms of age-related bone loss. AB - The human skeleton is formed and modelled during childhood and youth through the influence of hormones and daily mechanical usage. Around the age of 20-25 years, the skeleton achieves its maximum mass and strength. Thereafter, and throughout adult life, bone is lost at an almost constant rate due to the dynamic bone turnover process: the remodelling process. During this process, small packets of bone are renewed by teams of bone cells coupled together in time and space. In an adult human skeleton there will be 1-2 million active remodelling sites at any time point. The vast number of turnover units combined with a slightly negative balance at the completion of each process leads to the age-related loss of bone mass mentioned above and, concomitantly, to loss of structural continuity and strength. The magnitude of this loss will be determined by hormonal factors, nutrition and mechanical usage. As a consequence of the remodelling process, the bone tissue of the skeleton will always be younger than the age of the individual. However, as a consequence of the remodelling process, osteopenia and osteoporotic fractures will also occur. In this article, the remodelling-induced changes in the human spine will be used as an example of ageing bone. PMID- 11280025 TI - The old heart: operating on the edge. AB - Excitation of cardiac cells is accompanied by Ca2+ influx which triggers a transient increase in cytosolic [Ca2+], (Cai), and contraction. While the amplitudes of the Cai transient and contraction increase with the extent of cell Ca2+ loading, excess Ca2+ loading leads to dysregulation of Ca2+ homeostasis, impaired contraction, arrhythmia and cell death. The cell Ca2+ load is determined by membrane structure and permeability characteristics, the intensity of stimuli that modulate Ca2+ influx or efflux via regulatory function of proteins within membranes, and reactive oxygen species (ROS), which affect both membrane structure and function. Cardiocytes of senescent hearts exhibit a reduced threshold for pathologic manifestations of excess Ca2+ loading during stimulation (physiologic or pharmacologic) that increases Ca2+ influx, e.g. in response to neurotransmitters, post-ischaemic reperfusion, or oxidative stress. Cell 'remodelling' is one cause of the relative Ca2+ intolerance of cardiocytes in the senescent heart; cells increase in size and changes occur in the amounts of proteins that regulate Ca2+ handling due, in part, to altered gene expression; another cause is a change in the composition of membranes in which Ca2+ regulatory proteins reside, e.g. an increase in membrane omega 6:omega 3 polyunsaturated fatty acids (PUFA); a third cause is an enhanced likelihood for intracellular generation of ROS. Each class of these determinants changes with ageing and reduces the threshold for Ca2+ overload to occur with the older heart. The risk of excess Ca2+ loading within the senescent heart can potentially be reduced by gene therapy to restore Ca2+ regulatory proteins, by diet to reverse the membrane omega 6:omega 3 PUFA imbalance, or by antioxidants. PMID- 11280026 TI - Pharmaceutical intervention of advanced glycation endproducts. AB - Recent studies have revealed that reducing sugars, such as glucose, react with proteins through non-enzymatic glycosylation to form irreversible, covalently cross-linked proteins known as advanced glycation endproducts (AGEs). Furthermore, it has been demonstrated that this naturally occurring process, accelerated in diabetics due to hyperglycaemia, impairs biological functions leading to cardiovascular disorders, as well as diabetic and age-related complications. Pharmaceutical intervention to prevent or reverse these complications have focused on inhibiting the formation of AGEs by compounds such as dimethyl-3-phenacylthiazolium chloride or breaking the glucose derived cross links by selective cleavage. Intervention targeted at AGE cross-links in vivo offers a way to interfere with age-related changes of tissues. PMID- 11280027 TI - The anti-ageing action of dietary restriction. AB - Over 60 years ago, McCay's laboratory showed that dietary or calorie-restriction dramatically increased the lifespan of rats. Since then, numerous laboratories with a variety of strains of rats and mice have confirmed this initial observation and have shown that reducing calorie intake (without malnutrition) significantly increases both the mean and maximum survival of rodents. Currently, dietary restriction is the only experimental manipulation that has been shown to retard ageing of mammals. Although mechanism whereby dietary restriction retards ageing is currently unknown, much of the emerging data suggest that the calorie restricted rodents live longer and age more slowly because they are more resistant to stress and have an enhanced ability to protect cells against damaging agents. PMID- 11280028 TI - Mitochondrial DNA mutations in disease and ageing. AB - The chronological accumulation of mitochondrial DNA mutations has been proposed as a potential mechanism in the physiological processes of ageing and age-related disease. We discuss the evidence behind this theory and relate some of the ageing mitochondrial changes to mitochondrial DNA disorders. In particular, we describe the aggregation of cytochrome c oxidase-deficient cells in both skeletal muscle and the CNS in normal ageing as seen in the mitochondrial DNA disorders. These mitochondrial enzyme-deficient cells have been shown to occur in significant quantities in both muscle and CNS in patients with mitochondrial DNA disorders. In both ageing and mtDNA disorder muscle these cytochrome c-deficient fibres contain high levels of a single mutant strain of mitochondrial DNA. Whether these mutations are a primary or secondary event in the physiology of ageing remains to be determined. PMID- 11280029 TI - A mitochondrial paradigm for degenerative diseases and ageing. AB - A variety of degenerative diseases have now been shown to be caused by mutations in mitochondrial genes encoded by the mitochondrial DNA (mtDNA) or the nuclear DNA (nDNA). The mitochondria generate most of the cellular energy by oxidative phosphorylation (OXPHOS), and produce most of the toxic reactive oxygen species (ROS) as a by-product. Genetic defects that inhibit OXPHOS also cause the redirection of OXPHOS electrons into ROS production, thus increasing oxidative stress. A decline in mitochondrial energy production and an increase in oxidative stress can impinge on the mitochondrial permeability transition pore (mtPTP) to initiate programmed cell death (apoptosis). The interaction of these three factors appear to play a major role on the pathophysiology of degenerative diseases. Inherited diseases can result from mtDNA base substitution and rearrangement mutations and can affect the CNS, heart and skeletal muscle, and renal, endocrine and haematological systems. In addition, somatic mtDNA mutations accumulate with age in post-mitotic tissues in association with the age-related decline in mitochondrial function and are thought to be an important factor in ageing and senescence. The importance of mitochondrial defects in degenerative diseases and ageing has been demonstrated using mouse models of mitochondrial disease. An mtDNA mutation imparting chloramphenical resistance (CAPR) to mitochondrial protein synthesis has been transferred into mice and resulted in growth retardation and cardiomyopathy. A nDNA mutation which inactivates the heart-muscle isoform of the adenine nucleotide translocator (Ant1) results in mitochondrial myopathy and cardiomyopathy; induction of ROS production; the compensatory up-regulation of energy, antioxidant, and apoptosis gene expression; and an increase in the mtDNA somatic mutation rate. Finally, a nDNA mutation which inactivates the mitochondrial Mn superoxide dismutase (MnSOD) results in death in about 8 days due to dilated cardiomyopathy, which can be ameliorated by treatment with catalytic anti-oxidants. A partial MnSOD deficiency chronically increases oxidative stress, decreases OXPHOS function, and stimulates apoptosis. Thus, the decline of mitochondrial energy production resulting in increased oxidative stress and apoptosis does play a significant role in degenerative diseases and ageing. PMID- 11280030 TI - Specific metal-catalysed protein oxidation reactions in chronic degenerative disorders of ageing: focus on Alzheimer's disease and age-related cataracts. AB - Abnormalities of protein aggregation and deposition may play an important role in the pathophysiology of a diverse set of chronically progressive degenerative disorders including Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease and age-related cataracts. We propose that aberrant metalloprotein reactions may be a common denominator in these diseases. In these instances, an abnormal reaction between a protein and redox active metal ions (especially copper or iron) promotes the generation of reactive oxygen species, and possibly, protein radicalization. These products then lead to chemical modification of the protein, alterations in protein structure and solubility, and oxidative damage to surrounding tissue. In this review, we explore these ideas by focusing on two common diseases of ageing, Alzheimer's disease and age-related cataracts. Understanding the metalloprotein biochemistry in both diseases may lead to a better understanding of the underlying pathophysiology in both disorders and suggest novel targets for therapeutic agents. PMID- 11280031 TI - The priority of basic research on ageing vulnerability in a comprehensive research agenda on ageing for the 21st century. AB - The prospects for individual and population ageing as we enter a new century pose some of the greatest social, economic and humanitarian challenges humankind as a whole has ever faced. The basic biological mechanisms that control human ageing remain ill understood but it is clear that for many individuals exhibiting predisposition to risk factors for certain chronic diseases, such as coronary heart disease, diabetes, osteoporosis, certain cancers and Alzheimer's disease, such predisposition is mediated through genetic processes that operate at a most fundamental biomolecular level interacting with nongenetic attributes. The prospect of improved understanding of the fundamental processes underlying the pathogenesis of common age-related diseases that may lead to identification of interventions that are effective in preventing, delaying or ameliorating the diseases and their consequences is compelling. It is this prospect that provides the prime justification for giving high priority to research on ageing vulnerability in a comprehensive research agenda on ageing for the 21st century. PMID- 11280033 TI - Multiple substrates of late-onset dementia: implications for brain protection. AB - Age is the single most important risk factor for progressive dementia in populations worldwide. In developed countries the prevalence of dementia is estimated to be 3-5% at age 65 years and expected to double every decade thereafter. Although there is ageing-related attrition of neural tissue accompanied by profound changes in brain glia, marked neuronal loss and severe cognitive impairment are associated with pathological changes. Accelerated somatic ageing of the vasculature comprising endothelial and smooth muscle cells and slowed glial replacement are also likely to pre-dispose to degenerative processes. Approximately 90% of patients with late-onset dementia have neuropathological features of Alzheimer's disease (AD), dementia with Lewy bodies (DLB), or vascular dementia (VaD), alone or in combination. Both AD and DLB reveal extensive amyloid beta deposition within senile plaques. Neurofibrillary tangles evident as tau pathology are much reduced in DLB where symptoms may be more related to cholinergic transmitter abnormalities than structural pathology. Depletion of brain acetylcholine is also encountered in VaD, which like AD and DLB may respond to cholinergic therapy. Cerebrovascular pathology, ischaemic brain damage and neurovascular instability resulting in cerebral hypoperfusion appears fundamental in the pathogenesis of late-onset dementia. The apolipoprotein E epsilon 4 allele, a major genetic susceptibility factor for AD also associated with cardiovascular pathology, may contribute to neurodegenerative changes through vascular mechanisms. The interrelationships of these multiple substrates of late-onset dementia have major implications for neuroprotective and disease slowing therapies. Measures that improve cardiovascular function and increase brain perfusion would be useful to attenuate cognitive decline. PMID- 11280034 TI - Ageing of murine small intestinal stem cells. AB - Most organs of the body comprise populations of cells that are committed to specialized functions and that are renewed from small numbers of uncommitted progenitor or 'stem' cells. Stem cells are of central importance in the study of ageing because any senescent decline in the number or functional competence of stem cells will impair the capacity for renewal and turnover of committed cells, with potentially serious consequences for tissue homeostasis. The intestinal epithelium represents an excellent model system for the study of stem cells. Its spatial and hierarchical organisation allows the study of the function or characteristic of a given cell according to its position within the crypt. Hence, the stem cells which are located at the 4th-5th cell position from the bottom can be studied together with their daughter cells, as they divide and differentiate while migrating along the crypt-villus axis. The ability of the stem cells to undergo apoptosis and the capacity to regenerate the epithelium following injury were investigated in mice of different ages. Stem cells from older animals showed an increased apoptotic response following exposure to low doses of ionising radiation. The regenerative capacity was estimated by measuring the crypt survival levels and the growth rate of surviving crypts after high doses of irradiation. Surviving crypts in the older mice, suggesting an impairment in the damage recognition/response mechanisms, were both fewer and smaller than in young mice. The growth rate of surviving crypts was determined by measuring the crypt area and the number of cells/crypt at various times after 14 Gy irradiation. There was a growth delay of between half and one day in the older mice, and they subsequently grew more slowly. The number of cells susceptible to regenerate a crypt was also estimated. Surprisingly, they appear to be more numerous in the older mice. These studies indicate important age-related alterations in the capacity of the stem cells to regenerate the crypts after radiation-induced damage. The molecular bases of these changes are currently being investigated. Preliminary data showed alteration in the level of p53 and p21 expression, suggesting an age-related defect in the capacity to recognize damage and initiate apoptosis or repair. PMID- 11280035 TI - Haematopoietic stem cell ageing. AB - The question of whether haematopoietic stem cells age has raised considerable controversy, and has been re-opened recently, as a result of the growing interest in stem cells for transplantation and gene therapy. Studies have focused on the generation of different blood cell elements and the capacity for self-renewal; properties that characterize stem cells. Taken together, it appears that basal haematopoiesis is maintained throughout life, yet, the capacity to cope with haematological stress is decreased in advanced age. In principle, stem cells derived from aged donors can be used for autologous transplantation, when needed to recover basic haematopoiesis. However, patterns of T cell development are altered in ageing, and intervention to augment T cell response still needs to be considered. Current methods for expansion and maintenance of stem cells in vitro enable examination of stem cell potential for long-term expansion and function. A critical evaluation of the possible risks of replicative senescence and developmental changes in stem cells has become feasible. Ageing effects may relate to cell replication, cell migration and lymphoid differentiation. Understanding of the mechanisms underlying these processes will enable the fidelity of stem cell expansion and maintenance of their potential for long-term function. PMID- 11280036 TI - [Kinetics of potassium transfer during hemodialysis]. AB - INTRODUCTION: In order to study whether the removal of potassium in haemodialysis patients could be increased, we analyzed the kinetics of potassium transfer in the dialyzer. METHOD: 40 patients were included in the study. We studied: a) in vitro potassium exchanges between erythrocytes and plasma; b) plasma and erythrocyte potassium concentrations at dialyzer input and output; c) potassium transfers into the dialysate, using plasma clearance and direct measurement in the collected dialysate and d) erythrocyte potassium concentrations at the beginning and the end of dialysis. RESULTS: In vitro, there is virtually no potassium transfer between erythrocytes and plasma. In vivo, erythrocyte potassium concentration is not affected by the dialyzer (98.7 +/- 6.4 mmol/l to 97.7 +/- 7.5 mmol/l, p = NS). Potassium transfer levels determined by calculated plasma clearance were similar to values obtained by measuring potassium in dialysate (0.71 +/- 0.10 mmol/min vs 0.68 +/- 0.10 mmol/min, p = NS). These results suggest that erythrocytes do not participate in potassium exchange in the dialyzer. This was confirmed by measured erythrocyte potassium concentrations, which were the same at the beginning and the end of dialysis (104.0 +/- 5.6 mmol/l vs 104.2 +/- 5.0 mmol/l, p = NS). PMID- 11280037 TI - [Actinomycosis after renal transplantation: apropos of 1 case and review of the literature]. AB - Actinomycosis is a suppurative infection usually due to a facultative anaerobic bacteria, actinomyces israelii. This rare infection has been reported in immunocompetent individuals, with buccal or pharyngeal mucosal erosions. Paradoxically, few cases have been observed after solid organ transplantation: 2 cases after lung, 1 case after heart-lung transplantation and 1 case after renal transplantation. We report on a renal transplant recipient who developed a tongue and oropharynx suppurative abscess, looking like an epithelioma. Histological examination showed granulomatous inflammation with an angiofibroblastic reaction; few colonies of actinomyces were also observed by the pathologist. This lesion disappeared easily and totally after tetracycline treatment. PMID- 11280038 TI - [Refractory ascites in hemodialysis: treatment by paracentesis- reinjection during dialysis]. AB - Two hemodialysis patients, one male and one female, aged 46 and 54 years, were treated with preceed respectively for refractory ascites secondary to hepatic cirrhosis and for large polycystic liver. Preceed was decided because of the rapid reappearance of effusion following repeated puncture and albumin infusion, the poor tolerance to ultrafiltration (UF) and the poor nutritional status of the patients, with severe hypoalbuminemia. Abdominal paracentesis was performed on initiation of the dialysis session. Reinjection of the ascites fluid was made into the arterial line, allowing its UF and control of its flow. The procedure was performed whenever necessary, i.e., when inter-dialysis weight gain and ascites volume were high. In both cases, improvement was quickly obtained, with less rapid and less severe reappearance of the effusion and correction of albuminemia. Dialysis sessions with UF were better tolerated. No notable side effect was observed. The first patient was treated for 2 months, when he died of an unrelated cause. The other patient was treated for 6 months and then could be transferred to a dialysis center near her home. Twenty five months after start of dialysis treatment, kidney and liver transplantation were performed in this same patient. After transplantation, reappearance of moderate ascites and oedema is attributed to e degradation of renal function, without liver dysfunction. Five weeks after transplantation, improvement of renal function and ascites regression were noted. Preceed is an effective method of treating refractory ascites in the hemodialysis patient. Compared to classical paracentesis, it has the advantage of good tolerance, patient comfort and moderate cost. PMID- 11280039 TI - [Obstetrical acute renal failure: a public health problem in developing countries]. PMID- 11280041 TI - [Polygamy in Atriplex halimus L. (Chenopodiaceae)]. AB - It appears that up to now the inflorescence and flower morphologies of Atriplex halimus have been described incompletely. This species has been classified as monoecious or dioecious. Numerous observations and ontogenic studies have pointed to types of flowers morphologically and functionally hermaphroditic, never described until now. One specimen of this species presents both unisexual, male and female flowers and bisexual flowers, so Atriplex halimus is polygam and more precisely trimonoecious. Observation of inflorescences reveals a structure based on the spike and the dichasium. The sex distribution along the inflorescence axis was studied and the existence of a physiological gradient controlling its expression is discussed. PMID- 11280040 TI - [Plasminogen activator inhibitor type 1: physiology and role in renal physiopathology]. AB - Plasminogen activator inhibitor type 1 plays a prominent part in the regulation of extra and intra-vascular fibrinolysis through the inhibition of plasmin formation. In addition to its role in the resolution of blood clots, PAI-1 is involved in a variety of other biological processes including extracellular remodeling, cellular mobility, embryo implantation, development and tumoral proliferation. Moreover, PAI-1 is also implicated in various pathological processes such as thromboembolic diseases, atherosclerosis and fibrosis formation, particularly in the kidney and the lung. Inhibition of PAI-1 activity or of PAI-1 synthesis by specific antibodies, peptidic antagonists, antisens oligonucleotides or decoy oligonucleotides has been obtained in vitro but need to be evaluated in vivo. All these findings may have new therapeutical implications, explaining the importance of studies on PAI-1 production and regulation. PMID- 11280042 TI - Influence of gangliosides or LPS-like gangliosides on the tumoricidal activity of adherent leukocytes. AB - We previously showed that highly metastatic clones derived from the poorly metastatic human melanoma cell line M4Be are very radiosensitive provided that they are deficient in complex gangliosides. Here, we report that the highly metastatic clone 4 appears more sensitive to activated adherent leukocytes than M4Be via a transmembrane TNF-alpha-dependent mechanism. Adherent leukocytes (AL) were freshly isolated from different blood donors and were activated with Esherichia coli lipopolysaccharide (LPS). These AL contain 80% (73-93%) monocytes, 15% (6-20%) B lymphocytes and 5% (1-8%) T lymphocytes. The tumour cell survival following contact with AL was estimated with a clonogenic assay where isolated tumour cells were plated for 14 days with AL. We show on the one hand that either exogenous bovine brain GM1 gangliosides or Campylobacter jejuni LPS with GM1-like structure (LPS-like GM1) significantly decrease the hypersensitivity of clone 4 to AL. On the other hand, the cleaving with neuraminidase of more than 50% of the sialic residues bound to endogenous gangliosides in resistant M4Be cells significantly increases their sensitivity to AL. Thus, our highly metastatic cells appear both very sensitive to activated AL when they are deficient in complex gangliosides and resistant to AL when they are transiently exposed to exogenous gangliosides or LPS-like gangliosides. These in vitro data may reflect the paradoxidal behaviour of highly metastatic cells in vivo which appear both very sensitive to physiological stresses and able to survive to form secondary tumours. PMID- 11280043 TI - Identification of parasitoses in a child burial from Adak Island (Central Aleutian Islands, Alaska). AB - Bothriocephalid (Diphyllobothrium pacificum) and Ascarid (Ascaris lumbricoides) eggs have been identified in a sample taken in the abdominal cavity of a child skeleton found in Zeto Point (ADK-011), an archaeological site on Adak Island in the Central Aleutian Islands (Alaska). PMID- 11280044 TI - A study of French centenarians: are ACE and APOE associated with longevity? AB - Association study is the method of choice to identify genes involved in complex processes that result from the interaction of environmental and genetic factors. However, because of biases that increase the risk of false positive reports, preliminary positive conclusions have to be reproduced on other populations to be validated as firm conclusions. In 1994, certain alleles of two genes, APOE (Apolipoprotein E) and ACE (angiotensin converting enzyme), were reported to be more frequent in French centenarians, suggesting an association with such a complex polyfactorial process as longevity. Enlargement of the French centenarian cohort allows a new assessment of this hypothesis on 563 centenarians. In contrast to APOE, the ACE association was not confirmed. Retrospective analysis of the initial study revealed discrepancies that may in part explain this observation. Risk of reporting false positive associations is discussed and recommendations to set up a rigorous experimental design are proposed. PMID- 11280045 TI - [Expression of sexual dimorphism in the fetal pelvic girdle]. AB - The objective of this study was to analyse the development of the foetal pelvis in order to define normal anatomic reference values as a function of gender and gestational age. The study population included 500 stillborn foetuses between the gestational ages of 18 and 41 weeks. Those foetuses without known demographic histories were strictly excluded. For each case studied, an AP radiograph was performed with the following parameters measured by two independent observers: pelvic width, inter-iliac width, inter-sciatic nodes, inter-pubic width and bi ischial width. The correlation between these radiographic measurements and the gestational age as well as the gender was analysed. The result indicated that the inter-ischiatic distance is significantly greater in the female foetus after the 26-27th week of gestation (P < 0.0062). Standard growth for the female and the male foetal pelvis is proposed with potential application in the study of normal and pathological development of the foetus. PMID- 11280046 TI - Interspecific competition between freshwater snails of medical importance: a Venezuelan example. AB - Lake Valencia is located in the centre of the endemic area of the intestinal schistosimiasis in Venezuela. The dominance of two pulmonate species, Biomphalaria glabrata and B. prona., was observed in the lake. Both species are strongly associated with two distinct types of habitats suggesting that competition is occurring between these two species. B. glabrata and B. prona play the role of intermediate hosts of schistosomes in Venezuela. At the present time, parasite transmission is not occurring in the lake but the planning of important development programmes represents a risk of creation of active schistosomiasis foci. The knowledge of the importance and distribution of the snail host populations is therefore essential and must be taken into account for developing future control strategies. PMID- 11280047 TI - Frequency and viability of diploid and haploid male offspring of mated females of solitary endoparasitoid Diadromus pulchellus. AB - Sex determination in the order Hymenoptera is based on arrhenotoky, hymenopteran males are usually haploid and females diploid. Males of the Ichneumonidae Diadromus pulchellus, solitary endoparasitoid of A Acrolepiopsis assectella pupae, are normally haploid, but diploid males are present in a natural population and can be obtained in an experimental population. The future of an ovocyte laid by mated females of the solitary endoparasitoid D. pulchellus was characterised by six probabilities related to the sex and development of the ovocyte. The probabilities of fertilisation of female ovocyte (k1) or non fertilisation (k3) showed that an inseminated female functioned as a unmated female for half of the time (since k1 = 0.492 and k3 = 0.455) with the probability of fertilisation of male ovocyte, k2, equal to 0.053. The survival probabilities of each type of ovocyte showed that an ovocyte had a high probability of developing up to the adult stage, although the difference between the calculated sex ratios at laying (males/females = 1.032) and at emergence (0.90) revealed a slight reduction in the number of haploid sons. The probabilities of fertilisation and of viability of all the ovocytes laid by each of the 33 mated females were analysed by an ascending hierarchical classification of Euclidean distances and by an analysis of their principal components. The 33 mothers were distributed into four distinct sub-groups characterised by a sex ratio varying from an exclusive presence of females to an exclusive presence of males. Our hypothesis was that this distribution in four sub-sets could not simply result from the random nature of the sample. PMID- 11280048 TI - [Maples at the sub-Alpine vegetation belt: a long history]. AB - Pollen analysis was carried out on lacustrine sediment of a small hollow (15 m x 25 m) at the treeless sub-Alpine belt (202 m) of the inner Maurienne valley in the northern French Alps. A 2,500-year-long maple settlement was demonstrared. Three AMS dates of terrestrial plant macroremains support the chronology. First, Betula and Salix spread prior to 9,000 C14 BP. The first pollen grains of Acer, Abies and Pinus cembra are quoted at 8,600 C14 BP. High frequencies of Alnus glutinosa/incana (20%) and Acer (10%) show that mixed communities of Acer and Alnus persisted above the mountainous Abies forest between 7,490 and 5,850 C14 BP. After 5,850 C14 BP, the decrease in Acer stands could be attributed to fire as suggested by the strong increase in Betula and by the delayed expansion of Pinus cembra. PMID- 11280049 TI - Impact of land use in catchment and human activities on water, sediment and vegetation of Mediterranean temporary pools. AB - The vegetation and physical and chemical characteristics of the water and sediment in ten temporary pools submitted to various anthropogenic disturbance were studied in Morocco over two hydrological cycles (1997-1998 and 1998-1999). Results of multivariate and parametric analyses show that disturbance has a significant impact on water and sediment. Agriculture in the catchment resulted in higher levels of N and the use of detergent to higher levels of phosphorus in both water and sediment. Mineral extraction resulted in higher depth and longer duration of flooding. Vegetation characteristics were better correlated to hydrology (water depth, duration of flooding) than to nutrient variables. These results suggest that although agriculture in the catchment contribute in modifying the sediment characteristics, the impact on vegetation and its conservation value is limited. However, care should be taken of the long-term effects of agriculture through cumulative effects and of the possible consequences of changing the agricultural practices. PMID- 11280050 TI - Chemo-defence system. AB - By analogy with the immune defence system, the existence is suggested of a chemo defence system protecting living organisms against toxic substances, whether natural or man-made, that are present in the environment. This paper deals first with the various facets of such a system: mechanisms involving, among others, lipophilic compounds, hydrophilic compounds, oxidants, acidosis, genotoxics and metals; second, with the biological characteristics of the system and a comparison with the immune defence system: partial immaturity of the young, inducibility, non-specificity and specificity, and saturability; we will show that the two systems share many common features; third, with the evolution of the system, which demonstrates that the system is very old and suggesting that it came into existence before the immune defence system; and fourth, with some of its consequences: estimation of the 'toxic effects' of low doses, hormesis, impact of a vegetable diet on health. Finally, it could be emphasised that life is well protected against chemicals by its chemo-defence system, which appeared very early with the first living organisms on the earth. PMID- 11280051 TI - Associations of peak shifts in age--prevalence for human malarias with bednet coverage. AB - Effects of bednet coverage (C) on prevalence of malaria were analysed using data from 1990-92 from 9 Papua New Guinean villages. Effects of coverage varied by age, resulting in a shift in age of peak prevalence from 4.7 (C = 0%) to 11.6 (C = 100%) years for Plasmodium falciparum, from 3.4 to 4.9 years for P. vivax and from 11.0 to 16.8 years for P. malariae. In small areas with no bednets the age distribution of P. falciparum parasitaemia was like that of a holoendemic area. Where coverage was complete the pattern corresponded to mesoendemicity. Thus, protracted use of bednets can result in profound changes in the endemicity of malaria even when coverage is incomplete and without insecticide treatment. Average entomological inoculation rates (EIRs) estimated from indoor landing rates on individuals without bednets were 35, 12 and 10 infectious bites per person per annum for P. falciparum, P. vivax and P. malariae, respectively. Logistic regression analyses indicated that the EIR estimate for P. falciparum was related to prevalence of this species independently of effects of bednet coverage. However, the recent EIR still accounted for much less variation than did the bednets. A similar pattern was seen for P. malariae, while there were no significant relationships between the recent EIR and the parasite positivity for P. vivax. It is concluded that short-term variations in inoculation rate are not important determinants of parasite prevalence in this population. PMID- 11280052 TI - Evaluation under field conditions of the colourimetric DELI-microtest for the assessment of Plasmodium falciparum drug resistance. AB - It has been frequently stressed that improved methods are needed to monitor the fast spread of drug-resistant Plasmodium falciparum parasites in endemic areas. We recently developed a colourimetric microtest, double-site enzyme-linked lactate dehydrogenase enzyme immunodetection assay (DELI), to assess drug resistance in vitro. This method, which proved highly effective under laboratory conditions, was evaluated under field conditions in 2 African areas (in Senegal and Burkina Faso) in 1997 and 1998, respectively. The sensitivities of isolates from symptomatic (n = 50) and asymptomatic individuals (n = 26) infected with P. falciparum were assessed in parallel by the new DELI-microtest and the isotopic microtest. IC50 values of the isolates determined for chloroquine, quinine, amodiaquine and mefloquine were well correlated (r = 0.79, P < 0.001). The proportions of sensitive and resistant isolates determined using the 2 methods were similar. The DELI-microtest proved to be faster to implement than the isotopic-microtest, easier to perform, and did not require sophisticated equipment. Moreover, a larger number of isolates can be tested since parasitaemias as low as 0.005% could be reliably measured with the DELI microtest. These initial field studies thus support the value of the DELI microtest for large-scale drug-sensitivity monitoring. PMID- 11280053 TI - Molecular tracking of infections by Leishmania infantum. AB - Leishmania infantum is a major opportunistic parasite in patients with acquired immune deficiency syndrome and is very variable in these subjects. Isoenzyme characterization is not able to explain this variability, since half of the stocks isolated from patients co-infected with human immunodeficiency virus and Leishmania belong to zymodeme MON-1. Amplification of L. infantum minicircles by the polymerase chain reaction (PCR) and digestion of the amplified product to reveal restriction fragment length polymorphisms (RFLP) has proved very useful in distinguishing between relapses and reinfections in co-infected, treated patients. We have confirmed the existence of a leishmaniasis outbreak among intravenous drug users in north-east Spain, previously detected by isoenzymatic analysis. We have documented persistence of the same strain of Leishmania in 2 treated co-infected patients throughout several years, regardless of the theoretical rapid evolution ascribed to kinetoplast deoxyribonucleic acid minicircle sequences. We suggest using this PCR-RFLP technique to detect reinfections in treated co-infected subjects. PMID- 11280054 TI - Identification of Simbu, California and Bunyamwera serogroup bunyaviruses by nested RT-PCR. AB - We describe a reverse transcription-polymerase chain reaction (RT-PCR) with primers that anneal to the 5' and 3' ends and amplify the Bunyavirus S RNA segments. The RT-PCR was done on the fluids of C6/36 cells infected with each of 21 bunyaviruses. The bunyaviruses studied, with the exception of Catu virus, produced amplicons having 700 to 1300 base pairs and probably contained the whole S RNA segment sequence. A nested PCR performed with these amplicons distinguished California and most Bunyamwera serogroup viruses from other bunyaviruses by use of BBC specific internal primers for the S RNA segment, and distinguished Simbu serogroup viruses from others by use of BS specific internal primers. The nested PCR amplicons of Guaroa, Maguari, California encephalitis, Bunyamwera, and Oropouche viruses were sequenced. The sequences were aligned with previously known sequences of the S RNA segment of the same viruses, showing a high degree of homology and thus confirming the specific origin of these amplicons. The nested RT-PCR is suitable as a specific screening for most California and Bunyamwera serogroup and Simbu serogroup viruses depending on the use of BBC or BS internal primers. Oropouche virus is an important public health problem in Brazil and the nested PCR with BS primers could be used for the detection of this virus in tissue culture and mouse brain isolates as well as in clinical samples. PMID- 11280055 TI - Malaria in the highlands of Madagascar after five years of indoor house spraying of DDT. AB - The central region of Madagascar is a vast area of highlands (altitude 700-2000 m). Malaria transmission has re-established itself here since the last epidemic of 1985-90 and has caused the deaths of 40,000 persons according to the Minister of Health. To combat the main malaria vector in the region, Anopheles funestus, annual programmes of indoor house spraying of DDT were carried out between December 1993 and January 1998 in most rural areas at altitude 1000-1500 m. A parasitological and serological study was then conducted in the highland schools to evaluate the impact of the programme and set up a database on the region. Using a cluster-sampling method 2 independent selections were conducted (one of 130 sites, the other of 40 sites). During the study, 13,462 schoolchildren were examined, 71% living in sprayed villages. Parasite prevalence among schoolchildren declined as altitude increases, from 11% at 700-900 m to 0.4% at > 1500 m. Below 1500 m, the impact of the spraying on the prevalence of the parasite was very clear (an average decrease of from 20% to 2.7% below 1000 m and of from 4.5% without spraying to 0.8% at 1000-1500 m). Geographical analysis of the data showed that the marginal regions remained the most affected by malaria (especially outside spraying zones), and persistence of 'pockets of transmission' at 1000-1500 m, essentially in areas where spraying has never been used. In 9 schools, anti-Plasmodium antibodies were sought by indirect immunofluorescence on thick smears of parasitized red blood cells. The seroprevalence ranged from 22% to 63%, which suggests that the parasite is still circulating in the region. Even though our data show that vector control continues to be very successful in the Madagascan highlands, rapid reinfection could occur and must be monitored following spraying. To this end, the Minister for Health, with the support of the Italian Co-operation, has placed the region under epidemiological surveillance since 1997. An alert system for the timely detection of the sources of epidemics and the targeting of the antivectoral campaign is also in operation. Our study suggests that this strategy should be reinforced by the spraying of DDT in the marginal zones in order to consolidate the results obtained at higher altitudes. PMID- 11280056 TI - Risk factors for typhoid fever in the Mekong delta, southern Viet Nam: a case control study. AB - In order to identify risk factors for typhoid fever in a highly endemic area, we undertook a case-control study in the Mekong delta, Viet Nam. Cases were 144 consecutive patients admitted to hospital with blood culture-confirmed typhoid fever. Two controls (1 in the hospital and 1 in the community) were chosen for each case. Standardized interviews were conducted with questions regarding recent contact with a typhoid fever patient, eating habits, hygiene and socio-economic level. Cases were more likely to have been in contact with a patient with typhoid fever than hospital controls (adjusted odds ratio (OR) = 5.2, 95% confidence interval (95% CI) 1.7-15.9) or community controls (adjusted OR = 11.9, 95% CI 2.3 60.7); 11% and 14% of typhoid fever cases (compared to hospital or community controls, respectively) were attributable to recent contact with a patient with this disease. These findings suggest that strategies directed towards the persons in contact with a patient might reduce the incidence of secondary cases of typhoid fever. PMID- 11280057 TI - Further evidence for an exceptionally low prevalence of Helicobacter pylori infection among peptic ulcer patients in north-eastern peninsular Malaysia. AB - The Helicobacter pylori infection rate was determined in 124 consecutive patients with duodenal ulcers (DU), gastric ulcers (GU), duodenal erosions or gastric erosions diagnosed by endoscopy at a single institution in north-eastern peninsular Malaysia in 1996-97. Biopsies of the gastric antrum and body were subjected to the urease test, Gram staining of impression smears, culture and histopathological examination. Serology was undertaken on all patients using a locally validated commercial kit. Infection was defined as a positive result in at least one test. The infection rates were 20% (10/50), 21.2% (7/33), 16.7% (1/6) and 17.1% (6/35) in DU, GU, duodenal erosion and gastric erosion patients, respectively. The infection rate among Malays [7.0%, (6/86)] was lower than in non-Malays [47.4% (18/38)] (P < 0.001). There was a higher infection rate among males, who constituted 62.1% (77/124) of the sample. Seventy-eight patients (62.9%) were receiving non-steroidal anti-inflammatory drugs (NSAIDs) and 33 patients (26.6%) were neither receiving NSAIDs nor were infected with H. pylori. The H. pylori infection rate among peptic ulcer patients in this predominantly Malay rural population appears to be the lowest reported in the world thus far. Empirical H. pylori eradication therapy in peptic ulcer patients is clearly not indicated in this community. The possible reasons for the low prevalence of H. pylori infection are discussed. PMID- 11280058 TI - Presence of Lutzomyia evansi, a vector of American visceral leishmaniasis, in an urban area of the Colombian Caribbean coast. PMID- 11280059 TI - Detection of Wuchereria bancrofti in mosquitoes by the polymerase chain reaction: a potentially useful tool for large-scale control programmes. AB - Focally endemic bancroftian filariasis is targeted for elimination in the Nile delta of Egypt. Improved methods are needed for identifying endemic villages to be included in the control programme and for monitoring its success. We have evaluated the performance of a polymerase chain reaction (PCR) assay in estimating Wuchereria bancrofti infection in pools of Culex pipiens (1-25 females) from 2 adjacent villages with high (El Qolzom, 10.8%) and low (Kafr Shorafa, 2.1%) prevalence rates of human filariasis. This assay detects a repeated sequence in W. bancrofti deoxyribonucleic acid (DNA). Mosquitoes resting within houses were captured by aspiration and pooled by house. Houses were classified as positive or negative for human filarial infection based on night blood examinations of residents. The assay detected parasite DNA in mosquitoes from 60% of 25 infected houses and 24% of 25 uninfected houses. PCR processing of mosquitoes caught within houses of unknown filariasis infection status (44 in El Qolzom, 37 in Kafr Shorafa) identified 31.8% and 8.1% of houses, respectively, as containing infected mosquitoes. These results support the validity of the PCR assay for evaluating filarial prevalence in different villages. C. pipiens collected outdoors in dry ice-baited traps and tested by PCR (266 in Qolzom, 82 in Kafr Shorafa) did not contain parasite DNA. Pools of female mosquitoes (296 in Qolzom, 240 in Kafr Shorafa) captured in oviposition traps were also negative. We concluded that the PCR based assay is a powerful epidemiological tool that can be used for evaluating W. bancrofti infection in villages in the Nile delta and for monitoring the application of control programmes in filariasis endemic areas. PMID- 11280060 TI - The fibre-web blood sampling technique applied to serological diagnosis of schistosomiasis mansoni. AB - The fibre-web technique for sampling, storing and transport of venous or capillary blood has been evaluated, in 84 schoolchildren from the Mwanza region of Tanzania, with regard to diagnostic efficacy for determination of the schistosome circulating anodic antigen (CAA) under conditions similar to those prevailing in the field. Although the average concentrations determined in fibre web eluates were only about half of those determined in serum, the prevalences of CAA-positive individuals for the 2 sample materials were approximately the same. The average coefficient of variation calculated on determination of CAA in venous blood fibre-web eluates amounted to 7%. The study shows that the fibre-web technique is well suited for use under field conditions. PMID- 11280061 TI - Detection of anti-cysticercus antibodies by ELISA using whole blood collected on filter paper. PMID- 11280062 TI - Fatal Pneumocystis carinii pneumonia in HIV-seropositive infants in Harare, Zimbabwe. AB - Lung biopsies taken post mortem from 24 HIV-seropositive children who died of pneumonia in Harare Hospital (Zimbabwe) during 1995 were examined for pathogens using histology, culture, microscopy and polymerase chain reaction (PCR). Pneumocystis carinii was detected in 16 (67%) children, in 5 of whom bacterial pathogens were also detected. There were 2 cases of cytomegalovirus infection. On the basis of histology and PCR, none of the children had tuberculosis. These data add to the evidence that P. carinii pneumonia may be a significant cause of death in HIV-infected children in southern Africa. Policies on treatment for severe pneumonia, and on prophylaxis for children born to HIV-seropositive mothers need to be re-examined. PMID- 11280063 TI - Pneumocystis carinii pneumonia in patients with AIDS in South Africa. PMID- 11280064 TI - Serum C-reactive protein and detection of tuberculosis in persons co-infected with the human immunodeficiency virus. PMID- 11280065 TI - Mutations in the pfmdr1, dhfr and dhps genes of Plasmodium falciparum are associated with in-vivo drug resistance in West Papua, Indonesia. AB - This study (conducted in 1996-99) examines the association of mutations in pfmdr1, dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) genes of Plasmodium falciparum with in-vivo drug resistance in West Papua, Indonesia. Initially, 85 patients infected with P. falciparum were treated with chloroquine, of whom 21 were cleared of parasites, 49 had parasitaemias classified as RI, RII or RIII resistance and 1 patient had recrudescent parasitaemia. Fansidar (pyrimethamine-sulfadoxine) was the second-line treatment and 18 patients were cleared of parasites and 31 had continuing infections classified as RI, RII or RIII resistance and 1 patient had recrudescent parasitaemia. The pfmdr1, dhfr and dhps genes were examined for mutations previously shown to be associated with resistance to these drugs. In this study, mutations in pfmdr1 were associated with chloroquine resistance and mutations in both dhfr and dhps were associated with Fansidar resistance in vivo. Interestingly, Gly-437 in dhps along with Arg 59/Asn-108 in dhfr were associated with RI, RII and RIII resistance whereas Glu 540 was highly associated with only RII and RIII Fansidar resistance. This finding supports the hypothesis that the molecular basis of RI, RII and RIII Fansidar resistance involves an accumulation of mutations in both dhfr and dhps. These results suggest that mutations in both dhfr and dhps genes are a good predictor of potential Fansidar treatment failure. PMID- 11280066 TI - The comparative efficacy of chloroquine and sulfadoxine-pyrimethamine for the treatment of uncomplicated falciparum malaria in Kampala, Uganda. AB - Chloroquine (CQ) remains the first-line treatment for uncomplicated malaria in much of Africa despite the growing problem of resistance to this drug. Sulfadoxine-pyrimethamine (SP) is often used after CQ treatment failure and has replaced CQ as the first-line treatment in parts of Africa. To compare the efficacy of these 2 regimens, we evaluated, in March-August 1999, clinical and parasitological responses over 28 days in 214 children and adults from Kampala, Uganda, with uncomplicated falciparum malaria. Compared to SP, significantly more patients treated with CQ developed early or late clinical failure (54% vs 11%, P < 0.001) and parasitological failure (72% vs 30%, P < 0.001) during 14 days of follow-up. The risk of treatment failure occurring after day 14 was similar between the 2 treatment groups. Among those treated with CQ, children aged < 5 years were at higher risk of clinical failure than older individuals (76% vs 28%, P < 0.001), an association not seen with SP (11% vs 10%, P = 0.91). Although early parasite clearance was significantly better in the SP group (P = 0.001), fever clearance at day 3 was the same (CQ 85%, SP 86%). These and other recent findings suggest that consideration be given to replacing CQ as the first-line therapy for uncomplicated malaria in Uganda, particularly in young children. PMID- 11280067 TI - Evaluation of a new sulfadoxine sensitivity assay in vitro for field isolates of Plasmodium falciparum. PMID- 11280068 TI - The impact of a school health programme on the prevalence and morbidity of urinary schistosomiasis in Mwera Division, Pangani District, Tanzania. AB - The prevalence of urinary schistosomiasis among schoolchildren in Pangani District (Tanzania) was assessed rapidly by a questionnaire approach. Based on the results, a strategy of selective treatment with praziquantel was adopted. Eleven primary schools in Mwera Division, Pangani District, with about 2500 schoolchildren were included in a control programme for urinary schistosomiasis. Macro- and microscopic haematuria diagnosed visually and with urine reagent strips was used as an indirect indicator of Schistosoma haematobium infection. Intensity of infection among children was monitored in class 5 (median age 14 years, range 11-17) by urine filtration techniques. Treatment was administered as 40 mg/kg praziquantel in a single dose at the beginning of the school year. The programme was implemented by schoolteachers and coordinated by the District Health Management Team in collaboration with the District Education Office. Teachers were responsible for carrying out all programme activities. Community participation was through collaboration with Teachers-Parents Associations and Village Health Committees. Coverage at yearly (1995-99) examination varied from 67.7% to 80.3%. Prevalence of haematuria decreased from 51.2% (range 22.2-89.5%) at baseline to 23.4% (range 5.8-56.7%) in 1999, a reduction of 54.3%. Macrohaematuria was 21.2% at baseline and 7.2% in 1999, a reduction of 66.0%. Prevalence of infection in class 5 was reduced by 71.4% and geometric mean intensity of positives reduced from 71 eggs/10 mL (95% confidence interval [CI] 52.5-97.7) to 28 eggs/10 mL (95% CI 25.7-55.0), a reduction of 60.6%. Teachers were highly committed, and secured community participation and a smooth implementation of the programme. The community accepted the introduction of a cost-recovery system, whereby parents pay for the treatment of children with episodes of visible haematuria during the school year. Communities also participated in the improvement of sanitary installations at the schools. PMID- 11280069 TI - The efficacy of praziquantel against Schistosoma mansoni infection in Ndombo, northern Senegal. PMID- 11280070 TI - Artemether administered together with haemin damages schistosomes in vitro. AB - We conducted experiments in vitro to assess the effect of artemether in combination with haemin on adult Schistosoma japonicum, S. mansoni and S. haematobium. When schistosomes were maintained in a medium containing artemether at concentrations of 20 micrograms/mL or less for 72 h, no apparent effect on the schistosomes was seen. When the medium contained 50 or 100 micrograms/mL haemin as well as artemether, the schistosomes showed decreased motor activity 2-24 h after exposure, which was followed by the staining of the whole worm body a reddish-yellow colour, dilatation of the intestine, and extensive vesiculation of the tegument. Some of the schistosomes died 24 h after exposure, and almost all died within 48-72 h. When schistosomes were exposed to the same concentrations of haemin alone, they were stained a light yellow colour but there was no apparent effect on their survival. Our findings suggest that artemether interacts with haemin to exert a toxic effect on the worms, which might be of importance in the further elucidation of the mechanism of action of artemether on schistosomes. PMID- 11280071 TI - Area effects of bednet use in a malaria-endemic area in Papua New Guinea. AB - Relationships between area coverage with insecticide-free bednets and prevalence of Plasmodium falciparum were investigated in 7 community-based surveys over a 33 month period in 1990-93 in 6 villages in the Wosera area of Papua New Guinea. Spatial patterns in circumsporozoite rates for P. falciparum, P. vivax isomorphs K210 and K247, and P. malariae, and the proportions of mosquito blood meals positive for specific human, goat, cat, dog and pig antigens were determined using ELISAs. P. falciparum prevalence in humans was better explained by bednet coverage in the immediate vicinity than by personal protection alone. Circumsporozoite rates for both P. falciparum and P. vivax were also inversely related to coverage with bednets. There was some increase in zoophagy in areas with high coverage, but relatively little effect on the human blood index or on overall mosquito densities. In this setting, protracted use of untreated bednets apparently reduces sporozoite rates, and the associated effects on prevalence are greater than can be accounted for by personal protection. Even at high bednet coverage most anophelines feed on human hosts, so the decreased sporozoite rates are likely to be largely due to reduction of mosquito survival. This finding highlights the importance of local vector ecology for outcomes of bednet programmes and suggests that area effects of untreated bednets should be reassessed in other settings. PMID- 11280072 TI - Assessing the risk of benzimidazole therapy during pregnancy. PMID- 11280073 TI - An open, randomized comparative trial of two antivenoms for the treatment of envenoming by Sri Lankan Russell's viper (Daboia russelii russelii). AB - Russell's viper (Daboia russelii russelii) is an important cause of morbidity and mortality in Sri Lanka. In a study in 1985, Haffkine equine polyspecific antivenom in doses up to 20 g proved ineffective in clearing antigenaemia and caused a high incidence of anaphylactoid reactions. A new, monospecific ovine Fabantivenom (Polonga TAb) has been developed against the venom of Sri Lankan Russell's viper and, to assess its safety and efficacy, we carried out (in 1997) an open, randomized comparison of this with the Haffkine antivenom currently in use in the country. Patients with systemic envenoming following Russell's viper bite were randomized to receive an initial intravenous dose of either 1 g of Polonga TAb (n = 23) or 10 g of Haffkine antivenom (n = 20). One dose of Polonga TAb permanently restored blood coagulability in only 9 (41%) of 22 patients and 13 needed repeated doses, whereas the majority (14/20; 70%) had restored coagulability after 1 dose of Haffkine antivenom. There was a tendency towards more rapid resolution of local swelling and systemic manifestations in the Haffkine group. Venom antigenaemia was eliminated more quickly in the Haffkine group and ovine Fab was cleared from the circulation more rapidly than equine F(ab')2. To evaluate safety, patients were closely observed for adverse reactions. Following a severe reaction with Haffkine antivenom all subsequent patients in this group were treated prophylactically with hydrocortisone and chlorpheniramine. Despite this, the incidence of adverse reactions was significantly higher in the Haffkine group compared with the PolongaTAb group (81% compared with 48%) and 4 patients had a severe anaphylactic reaction in the former group. In conclusion, the new antivenom is safer than Haffkine antivenom but, to avoid repeated doses, an initial dose higher than 1 g is needed in the treatment of Sri Lankan Russell's viper envenoming. The safety of this larger dose is the subject of further studies. PMID- 11280074 TI - Premunition against Plasmodium falciparum in a malaria hyperendemic village in Myanmar. AB - Premunition, naturally acquired protective immunity against Plasmodium falciparum, has been described in hyperendemic areas of Africa and Papua New Guinea. However, its occurrence in Asia is debatable. In order to elucidate this question, a longitudinal study was undertaken in Oo-Do, a malaria endemic village in Myanmar [Burma] in 1995-97. Only 2 species, Plasmodium falciparum and P. vivax, were detected, with the former predominating. Data from 116 subjects showed that all were infected at one time or another, over a period of 3 years, with a 38% reinfection rate after eradication of patent parasitaemia. The high rate of prevalence (90-100%) of parasite-specific antibodies in the indirect immunofluorescence antibody test and the presence of the primary vector (Anopheles minimus) and 15 other species of Anopheles throughout the year indicated a high level of transmission. The spleen rate was 70% in 5-9 years old children and was inversely related with age. The incidence of parasitaemia was maximal (49%) in children aged 2-4 years, and then declined marginally with age. There was a significant difference (P = 0.001) between the asymptomatic and febrile parasitaemia levels. Also, malarial episodes occurred more frequently in children than in adults (P = 0.001). Taken together, all these facts indicated that the inhabitants of Oo-Do had progressively developed non-sterile partial protective immunity against P. falciparum malaria, or premunition. To our knowledge, this is the first detailed clinico-epidemiological study to document the occurrence of premunition in Myanmar. PMID- 11280075 TI - Interleukin-13 in Iranian patients with visceral leishmaniasis: relationship to other Th2 and Th1 cytokines. AB - The role of interleukin (IL)-13, a Th2 cytokine sharing many of the features of IL-4, has not previously been examined in patients with visceral leishmaniasis (VL). We examined sera from Iranian patients with VL caused by Leishmania infantum. Serum IL-13 was detected in 50% (22/44) of patients with active primary disease. In comparison, IL-10 was detected in 79.5% (35/44), interferon gamma (IFN gamma) in 38.5% (17/44), and IL-4 in only 5% (2/44) of these patients. With few exceptions all 3 cytokines were undetectable after clinical recovery following antimony therapy. Five of 7 patients (71%) who failed antimony therapy and had relapsing disease had similar levels of IL-10 to patients with active primary disease. However, with only 1 exception, IL-13, IFN gamma and IL-4 were not detected in such patients. These data suggest that relapsing disease may result from defective cellular immunity, unrelated to immunosuppression mediated by IL-10. PMID- 11280076 TI - The immunopathology of actinomycetoma lesions caused by Streptomyces somaliensis. AB - The immune responses in actinomycetoma lesions caused by Streptomyces somaliensis in Sudan were characterized by immunohistochemistry during 1997-1998. In sections stained with haematoxylin and eosin, the inflammatory reaction around the grain was of 2 types. In type I there were 3 zones: a neutrophil zone immediately around the grain, an intermediate zone containing mainly macrophages, and a peripheral zone consisting of lymphocytes and plasma cells. Zone 1 stained positively for CD15 (neutrophils), zone 2 for CD68 (macrophages) and CD3 (T lymphocytes), and zone 3 for CD20 (B lymphocytes). In the type II reaction, there was no neutrophil zone, the grains being surrounded only by macrophages and giant cells. This was confirmed by immunohistochemistry, which demonstrated the presence of CD3 positive cells. Immunoglobulins G and M and complement were demonstrated on the surface of the grain and on filaments inside the grain. Neutrophils and macrophages were recruited into the lesion by complement and were involved in the fragmentation of the grain. The cytokine profile in the lesion and regional lymph nodes was of a dominant Th2 pattern (interleukins-10 and 4). PMID- 11280078 TI - A mini-exon multiplex polymerase chain reaction to distinguish the major groups of Trypanosoma cruzi and T. rangeli in the Brazilian Amazon. PMID- 11280077 TI - African Burkitt's lymphoma: a new perspective. AB - High titres of antibody to Epstein-Barr virus (EBV) late genes identify individuals at risk of developing endemic Burkitt's lymphoma (eBL). Viral lytic cycle early and intermediate-early gene expression in BL is associated with a favourable tumour response to chemotherapy. Our study investigated whether serological data identifying antibody expression to zta, a viral function that activates lytic-cycle gene expression, correlate with expression of its gene in tumours, and could have prognostic value. Studies on 10 Malawian patients, with presumed BL on clinical grounds, showed good correlations, suggesting that serum antibody responses might predict treatment responsiveness. The results with 1 patient were particularly striking. When admitted in January 1998, prognosis was poor as he was unable to walk, and had tumour cells, characteristic of stage IV disease, in his bone marrow. Laboratory investigations showed particularly high levels both of serum zta antibodies and of gene expression in his tumour. Follow up confirmed him alive 6 months after hospital discharge. Among the EBV-positive cases, 2 were ultimately diagnosed as rhabdomyosarcoma, a tumour not previously associated with this virus. The findings from this small study, if confirmed, should have value for future BL management in resource-poor parts of the world. PMID- 11280079 TI - Psychiatric genetics in the 21st century. PMID- 11280082 TI - Developmental disability training in Canadian psychiatry residency programs. AB - OBJECTIVE: To examine current training in developmental disabilities in Canadian psychiatry residency programs and to determine, from the programs' perspectives, how provinces across Canada are responding to the needs of persons with developmental disabilities and comorbid mental health disturbances (persons with a "dual diagnosis"). METHOD: A survey was completed by residency directors, or their designate, for all 16 psychiatry residency programs in Canada. RESULTS: Persons with developmental disabilities require psychiatric services throughout their lives, but inadequate training opportunities exist in many of the residency programs, particularly those involving adults and adolescents. While some didactic opportunities are usually available, supervised clinical opportunities are rare, and many of those reported are optional. CONCLUSIONS: Across Canada, there have been insufficient advances in clinical training and service developments to meet the needs of individuals with developmental disabilities and comorbid mental health disturbances. PMID- 11280083 TI - Global assessment of functioning following assertive community treatment in Edmonton, Alberta: a longitudinal study. AB - OBJECTIVE: To examine longitudinally the effects of Assertive Community Treatment (ACT) on Global Assessment of Functioning (GAF) scores in Edmonton, Alberta. METHODS: We acquired GAF scores for all clients at initial registration in the ACT program and at subsequent 18- and 36-month time points while in ACT. We analyzed both the entire ACT cohort and separate diagnostic groups. RESULTS: We obtained baseline and follow-up GAF scores for 411 clients, of whom the largest diagnostic group suffered from schizophrenia (n = 189), followed by bipolar disorder (n = 98). Collapsed across all groups, GAF scores significantly improved at both 18 (P < 0.0001) and 36 months (P < 0.0001). By group, at 18-month follow up, significant improvements were seen in patients with delusional disorder (P < 0.05), dysthymia (P < 0.05), schizoaffective disorder (P < 0.05), and schizophrenia (P < 0.001). This was also seen at 36-month follow-up, with the addition of significant improvements in those with bipolar disorder (P < 0.05). Those patients with major affective disorder or psychosis not otherwise specified (NOS) did not show significant improvements over time. Regardless of diagnosis, those clients with baseline GAF scores of < or = 40 significantly improved at both 18-month (P < 0.0001) and 36-month (P < 0.0001) follow-up, while those with baseline GAF scores above 40 did not show significant improvement. CONCLUSIONS: GAF scores improved at 18- and 36-month follow-up from enrolment in an ACT program. Groups with different diagnoses and levels of functioning at time of enrolment may not benefit to the same degree. PMID- 11280080 TI - Genetic counselling for schizophrenia in the era of molecular genetics. AB - OBJECTIVE: To review the role of genetic counselling for individuals with psychiatric illnesses. METHOD: Using schizophrenia as an example and including updated information about a genetic subtype (22q deletion syndrome), we discuss the value of the genetic counselling process in psychiatry, with support from the literature and our clinical experience. RESULTS: Genetic counselling, the process through which knowledge about the genetics of illnesses is shared, provides information on the inheritance of illnesses and their recurrence risks; addresses the concerns of patients, their families, and their health care providers; and supports patients and their families dealing with these illnesses. For comprehensive medical management, this service should be available to all individuals with schizophrenia and their families. CONCLUSIONS: New findings in the genetics of psychiatric illness may have important clinical implications for patients and their families. PMID- 11280084 TI - Posttraumatic stress disorder, trauma exposure, and the current health of Canadian bus drivers. AB - OBJECTIVE: Previous studies of veterans have linked posttraumatic stress disorder (PTSD) after combat-related trauma to increased reports of health problems. It is unclear whether this association between PTSD and increased health problems generalizes to civilians who are exposed to a broader array of traumatic events. We also do not know whether trauma exposure is associated with increased health problems in individuals who do not develop PTSD. Using a non-treatment-seeking civilian sample, we examined whether lifetime PTSD or trauma exposure by itself was associated with current health problems. METHODS: Using a cross-sectional design and self-report measures, we evaluated urban Canadian bus drivers (n = 342) on trauma exposure, lifetime PTSD, and current health problems. Based on their responses, we divided our sample into individuals who had never experienced trauma (n = 91), trauma-exposed individuals who had never developed PTSD (n = 218), and persons who developed PTSD at some point after trauma (n = 33). We compared these groups on health problems, treatment service use, and health assessment measures. RESULTS: The PTSD group reported increased health complaints, more frequent use of health treatments, and poorer health self ratings compared with the exposed non-PTSD and nonexposed groups. Trauma-exposed drivers without PTSD did not differ from unexposed drivers on any health measure. Controlling for sex and trauma frequency did not alter our findings. CONCLUSIONS: Trauma exposure that leads to PTSD is associated with increased health problems, while trauma exposure alone is not. Our results extend previous findings to a broader civilian context and clarify associations between trauma exposure and health. PMID- 11280081 TI - Genetic insights into schizophrenia. AB - OBJECTIVE: To outline new insights into the genetic etiology of schizophrenia. METHODS: We discuss several commonly held beliefs about the genetic issues in schizophrenia. RESULTS: The complex genetic nature of the illness poses a challenge for investigators seeking causative genetic mutations. Multiple independent research findings are, however converging to identify a relatively small number of chromosomal locations that appear to contain schizophrenia susceptibility genes. Also, a clinically relevant genetic subtype of schizophrenia (22qDS) has been identified. We are developing a better understanding of how schizophrenia relates to other psychiatric disorders. While investigations into the possible roles of dopaminergic and serotonergic systems continue, other approaches that do not require theories of the mechanism of illness are also being used to identify candidate susceptibility genes. CONCLUSIONS: Research to date suggests that our understanding of the pathophysiology of schizophrenia will soon be fundamentally altered by genetic approaches to this complex disease. PMID- 11280085 TI - Individual predictors of posttraumatic distress: a structural equation model. AB - OBJECTIVE: Recent research has called into question the "dose-effect" model of understanding response to trauma and has turned attention to the contribution of personality and environmental factors. This research seeks to model the interrelation of relational capacity (a component of personality), perceptions of social support, and posttraumatic distress. METHOD: A group of firefighters (n = 164) completed questionnaires that addressed exposure to traumatic events, social support, current level of distress, and relational capacity. Structural equation modelling was used to develop a framework for understanding traumatic reactions. RESULTS: The overall fit of the hypothesized model was excellent. Relational capacity had a significant negative effect on support, indicating that perceived social support decreased as disturbances in relational capacity increased. Perceived social support had a significant negative effect on level of distress. CONCLUSION: While some emotional response to disturbing events may be normal, the severity of symptoms covaries with the ability of the individual to develop and sustain supportive relationships to buffer the impact of events. PMID- 11280086 TI - Delirium in psychiatric inpatients: a case-control study. AB - OBJECTIVE: To investigate the clinical and pharmacoepidemiological determinants of delirium in a psychiatric inpatient population. METHOD: A case-control study design was used. Potential cases and potential controls were identified using hospital discharge data. The clinical record of each subject was reviewed using a validated protocol to confirm case and control status. Subsequently, exposure data were recorded from clinical records. RESULTS: Subjects admitted to hospital with delirium tended to be older, to have pre-existing cognitive deficits, and to have diagnoses of substance use disorders. Subjects who developed delirium after their admission to hospital were older than control subjects, more likely to have a history of cognitive impairment, and were significantly more likely to be treated during the hospitalization with lithium or anticholinergic antiparkinsonian medications. Antipsychotic medication exposures were also associated with delirium, but only at standard or above-standard dosage levels. Antidepressant and sedative-hypnotic medications were not associated with delirium. CONCLUSIONS: These findings indicate that using conservative dosages of antipsychotic medications and minimizing the use of anticholinergic medications for parkinsonian symptoms may help to prevent delirium in psychiatric inpatients. Anticonvulsant mood stabilizers may convey less delirium risk than lithium. Antidepressant medications and sedative-hypnotics were not important determinants of delirium in this population. PMID- 11280087 TI - Alcoholism: beliefs and attitudes among Canadian alcoholism treatment practitioners. AB - OBJECTIVES: To explore differences in views concerning adjunctive medications and theoretical orientation among Canadian practitioners from different professional backgrounds who treat alcoholism. METHODS: A survey of clinicians from different disciplines was conducted by mail. The response rate was 56%: 95 drug and alcohol counsellors, 46 social workers, 81 nonpsychiatrist addiction physicians, and 74 addiction psychiatrists. The number of items in the questionnaire was reduced using principal component analysis. Group differences were explored using analysis of variance with Bonferroni correction and Scheffe's posthoc comparisons. RESULTS: Physicians and nonphysicians differed in their views on the utility of medications in treating alcohol problems, the disease concept of alcohol problems, and the classification of alcohol abuse or dependence as psychiatric conditions. No group differences emerged on views regarding cognitive behavioural treatment, pharmacological-only interventions, combined treatment, and recovery without treatment. Psychopathology in the alcoholic was significantly more likely to be considered as secondary to the use of alcohol by nonpsychiatrist physicians. Nonphysician practitioners viewed alcoholic behaviour as self-medication. CONCLUSIONS: Groups differed on questionnaire items concerning medication use and the disease concept of alcoholism. Agreement on several areas may facilitate bridging the gap across disciplines. The implications of these results are discussed. PMID- 11280088 TI - Criminal harassment by patients with mental disorders. AB - OBJECTIVE: To assess whether there is a subgroup of persons with mental disorders who engage in criminal harassment and to determine whether substance abuse is a cofactor in this behaviour. METHOD: A cross-sectional casenote study was used to examine incidents of harassment by patients prior to admission to acute and forensic wards (n = 106) at a provincial psychiatric hospital. RESULTS: Of 106 patients, 8 (7.5%) were found to have engaged in behaviour defined as criminal harassment prior to admission. Only 1 was charged under Section 264 of the Criminal Code. Alcohol was a cofactor in only 1 case. CONCLUSIONS: The number of persons with a mental disorder who engage in criminal harassment prior to admission is relatively small. The behaviour is not usually identified at any stage of the admission as criminal harassment. Although substance-abuse problems were prevalent, substance use was not a concomitant risk factor for behaviour defined as criminal harassment. PMID- 11280089 TI - Treatment of posttraumatic stress disorder with olanzapine. PMID- 11280090 TI - Quetiapine-associated hypomania in a woman with schizoaffective disorder. PMID- 11280091 TI - Severe anorexia in an Amish Mennonite teenager. PMID- 11280092 TI - Quetiapine treatment in patients with Tourette syndrome. PMID- 11280093 TI - Antidepressant-induced sexual dysfunction in adolescence. PMID- 11280094 TI - A pilot study of the technical quality of telemedical consultations for remote trauma management. AB - The technical performance of a telemedical system when used for remote trauma management was compared with face-to-face consultation. Two rooms, 20 yards apart, were linked telemedically in the same Accident & Emergency Department. Two hundred patients, with 'minor' and 'moderate' injuries, underwent the two types of consultation. The Accident & Emergency consultant marked physical parameters using a five-point pre-determined Likert scale. The following parameters were thought to be of excellent quality when compared to face-to-face consultation: overhead fluorescent lighting for the background illumination, video lighting for a close-up view, sound quality after volume adjustment, echo-cancellation after adjustment and lip synchronization. However, the following parameters scored poorly: sound before volume adjustment, echo-cancellation before adjustment, fine and coarse movements. It can be concluded that the quality of lighting and image quality are good in telemedicine. Sound and movement still present some problems. This technology is likely to be used more frequently for remote trauma management. PMID- 11280095 TI - Video-based computer-assisted learning. AB - As a relevant exemplar of MPEG-1 and MPEG-2 digital video use, a multimedia computer-assisted learning (CAL) application dealing with Critical Communication Issues in medicine was developed. The application allowed the student to navigate through a series of high-quality digital video and audio clips that were delivered in near real-time from an Intranet server. This paper gives a brief background to the MPEG-2 video compression system and discusses the use of digital video in a CAL environment. PMID- 11280096 TI - The Kodak Awards to the institute of medical illustrators 2000. PMID- 11280097 TI - Superior temporal branch vein occlusion. PMID- 11280098 TI - The 'UCH album', circa 1916. PMID- 11280099 TI - Techniques for current awareness. Part 3: Using library and information services and bibliographic databases. PMID- 11280100 TI - Acromegaly. 1951. PMID- 11280101 TI - The comparative effectiveness of conventional and digital image libraries. AB - Before introducing a hospital-wide image database to improve access, navigation and retrieval speed, a comparative study between a conventional slide library and a matching image database was undertaken to assess its relative benefits. Paired time trials and personal questionnaires revealed faster retrieval rates, higher image quality, and easier viewing for the pilot digital image database. Analysis of confidentiality, copyright and data protection exposed similar issues for both systems, thus concluding that the digital image database is a more effective library system. The authors suggest that in the future, medical images will be stored on large, professionally administered, centrally located file servers, allowing specialist image libraries to be tailored locally for individual users. The further integration of the database with web technology will enable cheap and efficient remote access for a wide range of users. PMID- 11280103 TI - Healthcare resource utilisation and costs of treating NSAID-associated gastrointestinal toxicity. A multinational perspective. AB - OBJECTIVE: The aim of the study was to perform an economic analysis of a new therapy in 11 countries (Australia, Belgium, Finland, France, Germany, Italy, The Netherlands, Spain, Sweden, Switzerland and the UK) to assess the cost of treating the gastrointestinal (GI) events associated with the use of nonsteroidal anti-inflammatory drugs in patients with osteoarthritis and rheumatoid arthritis. METHODS: Estimates of GI event-related costs were based on the results of resource utilisation questionnaires. Resources required for the treatment and follow-up of GI events were identified and converted into costs from society and payer perspectives. RESULTS: From the perspective of society, the total per-event cost of managing GI-related events varies from $US51 to $US772 for GI discomfort, from $US108 to $US1100 for anaemia, from $US145 to $US1200 for ulcer and from $US1923 to $US5473 for serious GI events requiring hospitalisation. From the payer perspective, the total per-event cost varies from $US47 to $US680 for GI discomfort, from $US144 to $US762 for anaemia, from $US229 to $US795 for ulcer and from $US1787 to $US6729 for serious GI events requiring hospitalisation. The total cost is driven by hospital expenses for those events requiring hospital admission. For GI discomfort, physician consultations are generally the cost driver, whereas for ulcer and anaemia, cost is primarily driven by the rate of endoscopy. CONCLUSIONS: Costs associated with nonsteroidal anti-inflammatory drug related GI events differ significantly across countries as a result of variations in resources consumed and price/tariff policies. PMID- 11280102 TI - The burden of arthritis and nonsteroidal anti-inflammatory treatment. A European literature review. AB - The purpose of this literature review is to summarise data available from publications describing the burden of osteoarthritis and rheumatoid arthritis in Europe, and to highlight gaps in the literature. On the basis of extensive literature research, the epidemiology of arthritis, its treatment costs, and iatrogenic costs related to nonsteroidal anti-inflammatory drug (NSAID) treatments are described, differentiating results by country. The review shows that, as well as having a significant impact on healthcare budgets, arthritis also affects patients and caregivers. For those countries where data were available, indirect costs were found to be of comparable magnitude to direct costs. Additionally, it was found that the iatrogenic costs related to the treatment of NSAID-induced adverse events are a significant component of the total costs of arthritis. The number of publications on the burden of arthritis in Europe is rather small in comparison with what is available for the US. Comparison of national results shows wide variations between countries, which may be partly due to discrepancies in the methodology applied to estimate the burden of arthritis, the cost items included in the analysis, and the data sources used to gather cost information. Additionally, comparing the burden of arthritis by country across Europe is difficult because of the variety of ways in which results are presented, e.g. on a per-patient basis, or for the whole population. To better understand the burden of illness of arthritis in Europe, not only is more research required, but the methodology to be applied in burden-of-illness analyses must also be standardised. PMID- 11280104 TI - A framework for evaluating the clinical consequences of initial therapy with NSAIDs, NSAIDs plus gastroprotective agents, or celecoxib in the treatment of arthritis. AB - OBJECTIVE: The purpose of this study is to provide a framework for estimating the economic efficiency of nonselective nonsteroidal anti-inflammatory drugs (NSAIDs), concomitant gastroprotective agents (GPAs) to reduce the risk of NSAID toxicity, and celecoxib, a specific cyclo-oxygenase-2 inhibitor. Concomitant GPA therapies considered include one of the following: proton pump inhibitors (PPIs) plus NSAIDs, histamine H2 receptor antagonists (H2RAs) plus NSAIDs, misoprostol plus NSAIDs, and a single tablet formulation of diclofenac/misoprostol. DESIGN: The study employs a decision-tree framework to establish probabilities of upper gastrointestinal (GI) adverse events occurring over a 6-month time frame. Celecoxib clinical trial data are used to establish probabilities of upper GI events for celecoxib and NSAIDs, and published literature is used to predict upper GI events for the other concomitant therapies. Upper GI adverse events included in the decision-tree are as follows: GI discomfort, symptomatic ulcer, serious GI complications (with and without death), and anaemia with occult bleeding. MAIN OUTCOME MEASURES AND RESULTS: Clinical probabilities indicate celecoxib has significant tolerability and safety advantages compared with nonselective NSAIDs. Celecoxib also reduces the risk of GI adverse events to a similar or superior degree when compared with reductions observed with NSAIDs with concomitant GPAs. CONCLUSION: Use of celecoxib is expected to significantly reduce the economic costs of GI toxicity and its associated morbidity. PMID- 11280105 TI - An economic model for determining the costs and consequences of using various treatment alternatives for the management of arthritis in Canada. AB - OBJECTIVE: To construct a decision analytical model to compare the costs and clinical consequences of treating patients with celecoxib or various nonsteroidal anti-inflammatory drug (NSAID)/gastrointestinal (GI) co-therapy regimens for the management of osteoarthritis and rheumatoid arthritis. The model quantified the number of patients expected to experience any GI complication commonly associated with NSAID therapy. DESIGN: Resource use for the treatment of each GI complication in the model was estimated after consulting Canadian experts. Standard unit costs from Ontario were applied to resources to calculate the cost of each complication. MAIN OUTCOME MEASURES AND RESULTS: The model revealed that the NSAID-alone regimen was associated with the lowest cost [$262 Canadian dollars ($Can) per patient per 6 months] followed by the celecoxib regimen ($Can273), diclofenac/misoprostol ($Can365), NSAID + histamine H2 receptor antagonist ($Can413), NSAID + misoprostol ($Can421), and NSAID + proton pump inhibitor ($Can731). A break-even analysis showed that up to 80% of the study cohort could be treated with celecoxib instead of the NSAID-alone regimen without increasing the health system's overall budget. Celecoxib was associated with the fewest GI-related deaths, hospitalised events; symptomatic ulcers, and cases of anaemia. The celecoxib regimen was also associated with the fewest cases of upper GI distress. Sensitivity analyses revealed that the model was most sensitive to the distribution of GI risk in the population and to the ingredient costs of the treatment alternatives. CONCLUSIONS: This model indicates that the use of celecoxib could lead to the avoidance of a significant number of NSAID attributable GI adverse events, and the incremental cost of using celecoxib for arthritis patients > or = 65 years of age in place of current treatment alternatives would not impose an excessive incremental impact on a Canadian provincial healthcare budget. PMID- 11280106 TI - Economic evaluation of celecoxib, a new cyclo-oxygenase 2 specific inhibitor, in Switzerland. AB - OBJECTIVE: The aim of this study was to predict the cost effectiveness of celecoxib, a cyclo-oxygenase 2 (COX-2) specific inhibitor, in the treatment of arthritis patients in Switzerland. METHODS: We applied a decision analytical model to compare the effects of 6 months' treatment with the following: (i) celecoxib; (ii) nonsteroidal anti-inflammatory drug (NSAID) alone; NSAID protected with (iii) proton pump inhibitor (PPI), (iv) histamine H2 receptor antagonist (H2RA), or (v) misoprostol; and (vi) diclofenac/misoprostol. Treatment costs included drug acquisition and the management of gastrointestinal (GI) adverse effects, classified as GI discomfort, symptomatic ulcer, anaemia and serious GI events (requiring hospitalisation). Probabilities were derived from celecoxib clinical trials and the literature. Drug utilisation patterns and treatment costs reflecting Swiss practice were obtained from local sources. Analysis was from the public health insurers' perspective. A range of sensitivity analyses was performed. RESULTS: For the base case of patients at typical risk (0.56% per 6 months) of serious GI events, the total expected costs of 6 months' treatment were as follows: celecoxib 435 Swiss francs (SwF); NSAID alone SwF510; diclofenac/misoprostol SwF522; and other protected NSAID regimens between SwF1034 and SwF1415. Celecoxib remained the lowest costing treatment over all categories of GI risk. Celecoxib generated 115 expected adverse events per 1000 patients per 6 months, followed by NSAID + PPI (119), NSAID + H2RA (154), NSAID + misoprostol (202), diclofenac/misoprostol (203), and NSAID alone (220), again for the base case. The cost per adverse event averted for celecoxib compared with NSAIDs alone was estimated in a stochastic version of the model using Monte Carlo simulation. In 95% of 500 iterations, celecoxib was predicted to save both costs and adverse events, thus dominating NSAIDs alone; the maximum cost per adverse event averted was SwF440. CONCLUSIONS: Celecoxib is predicted to be the most cost effective of the treatments considered for managing arthritis patients in Switzerland. A policy of switching patients from NSAIDs to celecoxib is predicted to be cost saving for public health insurers, while reducing the burden of iatrogenic GI side effects. Greater cost savings would be realised when patients are switched from NSAIDs used with gastroprotective agents. Models such as this can provide a useful but simplified view of treatment outcomes and predicted results require prospective validation in clinical practice. PMID- 11280107 TI - Acute asthma. Emergency management in the community. AB - BACKGROUND: The prevalence of asthma in Australia is one of the highest in the world. Statistics suggest that two million or more Australians across all age groups have the disease. It is one of the top five medical problems referred to hospital and one of the commonest presentations seen in emergency departments. Asthma was the cause of death of 685 Australians in 1998. All general practitioners will have a cohort of asthmatics in their practice and consequently must be prepared to manage these people at any time, but especially at times of acute exacerbation. OBJECTIVE: To outline the features of acute asthma and its management in the community setting. DISCUSSION: General practitioners have a crucial role to play in the community and prehospital management of asthma. An awareness of the features and treatment of acute exacerbations will help them reduce an enormous burden on the health system and contribute to a reduction in morbidity and mortality in their patients. PMID- 11280108 TI - Epidemiology of asthma in children. Who gets asthma and why? AB - BACKGROUND: Asthma is a common disease in childhood which results in a substantial burden of illness in Australia. This is manifest by distressing symptoms, impact on quality of life and role performance and health service utilisation. OBJECTIVE: This article reviews the current literature on the epidemiology of asthma in children. It particularly focuses on the prevalence, trends in prevalence, risk factors and causes of asthma. The emphasis is on common questions that parents of children with asthma would like answered by their general practitioners. DISCUSSION: We know something about some of the risk factors for asthma and a little about the mechanisms for the development of asthma. Despite the paucity of knowledge about the causes of asthma, some positive steps can be taken to prevent it. Allergic or asthmatic parents, whose children have a high risk of developing asthma, can be advised to avoid smoking during pregnancy and avoid environmental tobacco smoke exposure after the child is born, to undertake house dust mite control strategies, to breastfeed their babies for at least three months and subsequently to provide their child with a nutritious, balanced diet. PMID- 11280109 TI - Management issues in adult asthma. AB - BACKGROUND: Asthma is common in the adult population and is one of the most frequently managed problems in general practice. It affects over two million Australians and costs about $600 million annually to the community in direct and indirect costs. OBJECTIVE: This article looks at the management of adult asthma including assessment, drug treatment, adherence issues and the importance of regular medical review. DISCUSSION: The burden of illness from asthma is concentrated in general practice. The general practitioner therefore, plays a crucial role in the direct management of asthma and in assisting patients to optimally self manage their asthma. General practitioners are ideally positioned to ensure that all their patients with asthma understand their disease, have written instructions about when and how to treat exacerbations (all people with asthma are at risk of exacerbations) and are prescribed optimal therapy to control symptoms. PMID- 11280110 TI - Moving towards organised care of chronic disease. The 3+ visit plan. AB - BACKGROUND: The health care system is better organised to react to acute rather than chronic presentations of disease, yet a significant proportion of health care expenditure is on chronic diseases and their complications. OBJECTIVE: This article seeks to argue the case for the organised care of chronic disease. Asthma is used as the demonstration disease because a model has already been developed and a pilot evaluation has been done. DISCUSSION: The National Asthma Campaign has been involved in the development, by general practitioners, of a system of asthma care organised in a way appropriate for general practice in Australia. The care is provided over a series of visits and ensures that all of the six steps of the Asthma Management Plan are covered. PMID- 11280111 TI - Proactive asthma care. The benefits of behavioural change. PMID- 11280112 TI - Newer antiplatelet therapies in stroke prevention. AB - BACKGROUND: Aspirin has been the mainstay of antiplatelet therapy in stroke prevention for 30 years. In the past decade, a number of new antiplatelet strategies have been shown in clinical trials to provide some benefits over aspirin therapy. These new compounds include ticlopidine, clopidogrel and the combination of aspirin with dipyridamole. OBJECTIVES: To review the efficacy and dosage of aspirin in stroke prevention, and to review the benefits and risks of the newer strategies, compared with aspirin. Based on the evidence from randomised, controlled clinical trials and systematic overviews, to present practical clinical guidelines for the use of aspirin and the newer antiplatelet drugs. DISCUSSION: For most patients aspirin monotherapy is still recommended as the first line antiplatelet strategy. However, some stroke clinicians are now recommending the combination of aspirin plus dipyridamole as a first line approach. For patients who are allergic to aspirin, clopidogrel is the drug of first choice and has largely replaced ticlopidine. For aspirin failures, either combined aspirin plus dipyridamole, or clopidogrel, are recommended. The combination of aspirin plus clopidogrel has theoretical appeal, is valuable in prevention of coronary stent thrombosis and is undergoing clinical trial in stroke prevention. Other novel approaches, such as oral platelet Gp IIb/IIIa antagonists are also being evaluated. PMID- 11280113 TI - Cooking and oxygen. An explosive recipe. AB - BACKGROUND: Home oxygen therapy is commonly prescribed for the treatment of chronic obstructive pulmonary disease (COPD). The risks of smoking while using this therapy have been well described. OBJECTIVE: To discuss the Royal Brisbane Hospital Burns Unit's experience and present case studies which illustrate the danger of alternative ignition sources while using home oxygen. DISCUSSION: The dangers of home oxygen therapy can be minimised by careful patient selection, education and ongoing monitoring. PMID- 11280115 TI - Practice tip. Improvised suppository inserter. PMID- 11280114 TI - Patient education. Home oxygen fact sheet. PMID- 11280116 TI - Herbs and metabolic/endocrine disease. From past to present. AB - This is the third article in this series. It covers a range of metabolic problems, some new and some old, in which herbal medicine has been traditionally suggested to have a place or where pharmaceuticals have herbal derivations. With the current 'epidemic' of diabetes and hyperlipidaemia, many patients are tempted by the media to consider herbal remedies. There is great concern that proven effective remedies may be ignored to the detriment of our patients' health. PMID- 11280117 TI - Herb-drug interaction guide. PMID- 11280118 TI - Casting acute fractures. Part 5--The gutter slab. AB - BACKGROUND: Punching injuries commonly present with displaced fractures of the distal fifth metacarpal. These are typically difficult to maintain in a reduced position. OBJECTIVE: The specific technique for the application of a strong slab likely to maintain reduction is demonstrated, the concept and use of moulding pads to safely undertake aggressive moulding is illustrated. DISCUSSION: Maintenance of reduction of these fractures is largely for cosmetic purposes. Immobilisation in a moulded cast improves the patient's comfort. PMID- 11280119 TI - Moderation in all things. PMID- 11280120 TI - General practice research. A window of opportunity. PMID- 11280121 TI - Urinary tract infections in men in a primary care population. AB - OBJECTIVE: There have been few studies on urinary tract infections (UTIs) in adult males. This study aimed to look at the clinical features of males presenting with urinary tract infections in a predominantly gay general practice population. METHOD: A retrospective audit and analysis was carried out of all male patients presenting with symptoms suggestive of a UTI and in whom a pathogen was cultured from urine. The subjects were drawn from two urban general practices in Melbourne. Subjects with known pre-existing urinary tract abnormalities, or in whom recent urinary tract instrumentation had been performed, were excluded, as were subjects in whom Neisseria gonorrhoeae was cultured from urine. RESULTS: Thirty-three subjects were identified, with a total of 47 presentations. Six subjects were excluded on the basis of pre-existing known factors which would predispose to UTIs, leaving 27 subjects, with 37 episodes of UTI. The mean age was 43 (range = 28-62) and 25 of the 27 identified as gay. Nine out of the 27 (33%) were HIV positive with a mean CD4 of 574/mm3 (range = 41-1812). The main presenting symptoms were dysuria/burning on micturition, urinary frequency, fever/sweats and haematuria. Urethral discharge occurred in two episodes. Multiple symptoms on presentation were common. The main organism cultured was E. coli. Fourteen subjects underwent further radiological investigation and two abnormalities were detected. CONCLUSION: Uncomplicated UTIs are uncommon in males presenting to general practice. The symptoms are similar to those in females with UTIs, though fever may be more common. Treatment with oral antibiotics in a primary care setting is generally curative. Further investigation of males with UTIs may be appropriate, but more studies are needed as to the cost-benefit of this. PMID- 11280122 TI - Integration from the Australian GP's perspective. AB - OBJECTIVE: To report on what general practitioners' perceptions are about their role in relation to activities that support integration and what they are doing. METHOD: General practitioner perceived integrative behaviour was measured using a survey containing 114 statements about, 'what constitutes a well integrated GP'. Four hundred and forty-eight GPs were randomly sampled from the Health Insurance Commission (HIC) Medicare billing database in 1996. A response rate of 47% was obtained, yielding 208 surveys for analysis. RESULTS: General practitioners reported integrative activities such as being accessible to patients and working within a multidisciplinary team as currently occurring optimally. Not occurring optimally were: hospital and community involvement; participation in local projects; student education; and payment for working with others. Rural practitioners reported significantly more hospital and community involvement compared with metropolitan practitioners. Less than one-third of GPs reported that they were linked to other services by computer and used a computer for storage/communication of patient information. DISCUSSION: There are many obstacles preventing integrative activities in daily general practice. Policy and attitudinal changes as well as financial incentives are required to enable GPs to practise in an integrated manner. Infrastructure support to encourage GP education, training and information technology are essential to improve GP integration. Many such initiatives are currently in progress, and will require future evaluation. Findings from this 1996 survey will provide some useful baseline information assisting with future evaluation studies. PMID- 11280123 TI - Application of the privacy principles to general practice. AB - BACKGROUND: There are escalating requirements for general practitioners to comply with recognised privacy principles. With amendments to the Commonwealth Privacy Act (1988) imminent, there is an urgent need to formulate methods for applying these principles to general practice. OBJECTIVE: The article provides an explanation of the origins of the privacy principles and a simple self audit which general practitioners can use to assess the extent to which their usual practices conform with them. DISCUSSION: A careful review of the principles indicates that new measures will be needed before most general practices will be able to approach required standards of conduct. Practical strategies for achieving best practice are discussed and challenges confronting general practices in applying the principles are canvassed. Ethics committees should be used more often to provide independent review of practice policies and proposals to use patient information in new ways. General practitioners can expect increasing scrutiny and debate concerning confidentiality. In order to maintain patient trust in GPs as responsible data custodians, the privacy principles can be seen as a quality improvement tool. PMID- 11280124 TI - Asthma care. An example of GPs taking the lead. PMID- 11280125 TI - Spinning dangerously in the 'top end'. PMID- 11280126 TI - Homeopathy. PMID- 11280127 TI - A better way for after hours care? PMID- 11280128 TI - Diagnosing pulmonary embolism in pregnancy--an ongoing debate. PMID- 11280129 TI - Of studies, syntheses, synopses, and systems: the "4S" evolution of services for finding current best evidence. PMID- 11280130 TI - Progress in the control of prostate cancer, Rhode Island, 1987-1998. PMID- 11280132 TI - Hospitals and the changing work environment: promoting gender equity and fair treatment for medical students. PMID- 11280133 TI - Bringing baby friendly to Rhode Island. PMID- 11280131 TI - Ethics, money & negative studies. PMID- 11280134 TI - Beta-blockers, diabetes, and hypoglycemia: risky business? PMID- 11280135 TI - Governor Lucius F.C. Garvin, MD--Rhode Island's Champion Dreamer. PMID- 11280136 TI - Boerhaave's syndrome. PMID- 11280137 TI - Invasive pneumococcal disease. PMID- 11280138 TI - Beta-blockers an important cause of depression: a medical myth without evidence. PMID- 11280139 TI - Atrial fibrillation and anticoagulation. PMID- 11280140 TI - Utilization of clinical preventive services among Rhode Island adults with and without health insurance coverage, 1999. PMID- 11280141 TI - Medicaid managed care. Good health care, or health care slavery? PMID- 11280142 TI - Oral health status of second grade school children in upstate New York. AB - This report summarizes the results of a survey of second grade children conducted in upstate New York. The survey was designed to monitor progress toward achieving Healthy People 2000 objectives and Maternal and Child Health Services Block Grant performance measures. Data on oral health status, use of preventive measures and insurance coverage were collected on 2,474 children from 76 schools. In addition to obtaining population estimates, disparities in oral health between poor and nonpoor children were assessed. The results showed that approximately 52% of second grade children had dental caries, and 35% had untreated disease. Approximately 43% received fluoridated water, and 44% of children living in non fluoridated areas used fluoride supplements on a regular basis. Only 25% of the children had dental sealants. The percentage of children covered by comprehensive and basic insurance plans was approximately 19% and 41%, respectively. Many of the national oral health objectives were not met primarily because of the higher rate of disease among the poor and their lower use of preventive services. These findings regarding oral health status and use of preventive services are similar to the national data. PMID- 11280143 TI - The future of dentistry 2001. PMID- 11280144 TI - Fluorides in the new millennium. AB - Water fluoridation is known to be the most successful public health measure of the 20th century. More than half a century later, we are still reaping the benefits of fluoridation. We now know that the most important mechanism of fluoride action occurs through daily low-dose exposures. The battle between demineralization and remineralization occurs constantly, and fluoride shifts the balance to the latter. This dynamic, daily process far supersedes the pre eruptive fluoride incorporation in importance. Fluoride, however, is now available in a variety of very different forms, useful in waging the war on caries. Nevertheless, opponents of this therapeutic agent maintain that its widespread use should be curtailed. Although the benefits of fluoride can no longer be disputed, fluoride supplementation must be supported and approached with consideration of total fluoride exposure. PMID- 11280145 TI - Community-based dental services for patients with special needs. AB - Deinstitutionalization of individuals with mental retardation and developmental disabilities has increased the demand for dental services for these patients by community practitioners. There are numerous difficulties associated with the delivery of care to this population with special needs. Nevertheless, a county-by county review for New York State indicates that in most instances, if all dentists are willing to help, there would be a relatively small number of these patients per dental practitioner. PMID- 11280146 TI - Out front for the children. PMID- 11280147 TI - OSHA moves to protect employees from work-related injuries. PMID- 11280148 TI - Implementation of the electronic medical record: the team approach. AB - The implementation of the electronic medical record (EMR) is a process that involves knowledge and skills of technology and group dynamics. The literature was reviewed to identify the most effective methodology for EMR implementation. Integral to the success of any EMR implementation is the 'buy-in' of the organization. To facilitate the implementation the use of a tiered team approach is advantageous. There must be an executive steering team whose role is to provide the vision, the approval, and the money for the project. The project steering team makes major policy decisions based on user needs and requirements, organization infrastructure, and general implementation strategies. The project work team has the responsibility of ensuring the implementation is satisfactory to the organization. This article discusses the membership, roles, and functions of the three EMR implementation teams. PMID- 11280149 TI - Evaluation of documentation before and after implementation of a nursing information system in an acute care hospital. AB - Economic pressures on healthcare systems have intensified the necessity of demonstrating the unique contribution of nursing care to patient outcomes. The use of nursing information systems (NIS) has increased completeness of some nursing documentation elements. This study's purpose was to evaluate differences in documentation completeness of nurse assessments of patient outcomes (NASSESS), achievement of patient outcomes (NGOAL), nursing interventions done (NQUAL), and routine assessments before and after implementation of an NIS in a 100-bed urban university hospital in west Tennessee and before and after retraining in NIS use and care planning. NIS implementation did not improve documentation within the first six months. However, retraining and continued NIS use did significantly improve NASSESS, NGOAL, NQUAL, and blood pressure documentation 18 months postimplementation. Nurses must evaluate documentation completeness before and periodically after NIS implementation, using results to improve patient record data validity for patient care decisions, quality improvement, and research. PMID- 11280150 TI - Hypermedia-assisted instruction: authoring with learning guidelines. AB - The use of computers in nursing education has evolved from elementary drill and practice tutorials to challenging presentations. Hypermedia technology has emerged as a powerful method of instruction that allows the user to experience new concepts, gain cognitive skill, and encounter realistic clinical situations in a no-harm environment. Hypermedia incorporates sound, video, graphics, text, and interaction. This article describes the process used to develop a CD-ROM for adult cardiovascular nursing that can be used as an interactive instructional program. A description of the use of Gagne's instructional design theory is incorporated as a framework to guide the project. Educators possess knowledge expertise and must become involved in the design of new teaching methodologies. If instructional design principles are followed when using authoring software, learners are motivated with real, challenging, life-like, nonthreatening scenarios to expand their critical-thinking and decision-making skills. PMID- 11280151 TI - Teaching pathophysiology to diverse students using an online discussion board. AB - Communication on the internet offers teachers a unique method for promoting learning among students who are challenged by numerous factors such as linguistic and cultural considerations. This article describes the use of out-of-class online discussions to promote active student learning among a diverse group of baccalaureate nursing students enrolled in a large lecture class. Student-to student and teacher-to-student asynchronous threaded discussions occurred out of class. Students earned class points by posting and responding to classroom topics. By the end of the 15-week semester, there were approximately 790 messages on the bulletin board. Eighty-five percent of the 64 students participated in this assignment. Of the 64 students, approximately 50% were moderate to high users of the activity and 14% did not participate at all. There were no statistical differences between native English-speaking and non-native English speaking students and their preference for study partners, interest, or experience with the internet. These initial results suggest that this assignment, based on electronic discussions, promoted greater participation and feedback among this diverse nursing student population. Despite the potential isolation of internet and e-mail-based communications, this computer-based technology may, ironically, facilitate an important shift from teacher-orchestrated to student centered learning. PMID- 11280152 TI - Use of and satisfaction with a browser-based nurse teaching tool in a surgical intensive care unit. AB - Our goal was to determine if a computerized teaching tool is an effective teaching method for nurses in a high-stress fast-paced intensive care unit. We also measured the level of satisfaction with this method of instruction. Thirty six surgical intensive care nurses used a Web-based Microsoft PowerPoint presentation located on the intranet at nursing stations located on the surgical intensive care unit (SICU). The presentation was designed to provide instruction regarding the methodology and use of APACHE III prior to its implementation. Paired t-tests were performed to compare the results of a pretest and posttest. The questions were divided into two types: methodology and use. After the nurses completed their training sessions, they were asked to complete a questionnaire. The questionnaire questions were rated one a 1 to 5 scale. The average scores were higher on the posttest compared to the pretest (63.2% vs. 69.1%, p = 0.03). The methodology scores were higher on the posttest (74.3% vs. 88.2%, p = 0.001), while the use scores remained the same at 78.1% vs. 75.0%. Our Web-based teaching tool is an effective way to train nurses to understand the APACHE III medical system. The tool was effective at conveying the APACHE III medical systems methodology but was not effective in explaining the usefulness of the system. Most important, the nurses thought the browser-based teaching tool was easily accessible and an effective way to communicate new material to the medical staff. PMID- 11280153 TI - Advances in the skeletal anatomy of the wrist. AB - Over the past decade there has been an increased interest in and study of the skeletal anatomy of the wrist. Different morphologic patterns of carpal anatomy have been identified and classified in the wrist. Certain morphologic types of carpal bones have been shown to have a correlation with the development of arthritis in joints of the wrist. Different shapes of carpal bones have also been shown to determine and affect the type of kinematics that they demonstrate. PMID- 11280154 TI - Biomechanics of pronation and supination of the forearm. AB - Pronation-supination, the rotation of the forearm around its longitudinal axis, is an important motion because it allows the hand to be oriented, allowing one to take food and carry it to the mouth, perform personal hygiene, and live autonomously. The motion depends on the integrity of two bones, the radius and the ulna, as well as joints, ligaments, and muscles. In every pathological case, as described in this article, the anatomical features must be restored for normal function. PMID- 11280155 TI - Vascularization of the thumb. Anatomy and surgical applications. AB - Microsurgical procedures require rapid and atraumatic dissection of vessels. The authors have tried to schematize the most common variations of the palmar arteries, dividing the thumb into three segments, delimited from the MCP and interphalangeal flexion crease. If a vascular anastomosis has to be carried out in the first segment, the arteries that intersect the palmar surface of such a region do not generally hold much interest for the surgeon as far as size and constancy are concerned. One therefore can avoid frustration while searching in vain for such an artery by starting dissection in the dorsal compartment, where there is a much better chance of finding an artery suitable in every aspect. In replanting a thumb in the second segment, the ulnar collateral artery should be the artery that is looked for first, because it is usually the biggest, the most superficial, and nearly always is composed of a single trunk (Fig. [figure: see text] 18). Let us not forget, however, that the contralateral artery can frequently have all the necessary requirements for an adequate anastomosis. In the third segment, the layout of the vessels is rather difficult to schematize; if it is true that the inverted Y or H shape can make a microanastomosis easier in cases of distal reimplantation (Fig. 19), it is also true that such a pattern is impossible to foresee, and, therefore, the interest of such classification is more academic than practical. Skin loss coverage at the thumb level greatly differs from the that of fingers. In the past few years, a multitude of useful new flaps has been presented by different authors. Their accomplishment (especially those of the last generation, who base their survival on extremely fine vascular axes) presupposes an adequate knowledge of surgical and microsurgical anatomy. Surgeons dealing with this type of pathology should be capable of performing all of the possible flaps because each may be indicated in specific situations. PMID- 11280156 TI - The trapeziometacarpal joint. Tenotomy of the accessory tendons in early osteoarthritis. AB - The TMC joint is an articulation with special articular surfaces adapted to produce simple (nonrotatory) and complex (rotatory) metacarpal movements. Its articular anatomy and biomechanics are closely related to the pathogenesis of osteoarthritis. The joint works under high transarticular compressive-shearing forces. In osteoarthritic thumbs, the articular forces are increased because of the constant presence of accessory APL tendons, almost exclusively of the digastric type. Other factors should be considered in the pathogenesis of TMC joint osteoarthritis, such as repetitive use of the thumb under unfavorable patterns of function) strong side-to-side pinch grips, thumb with the tendency to maintain in reposition), cartilage aging, hormonal disturbances in women, and general osteoarthritic disease. Osteoarthritic thumbs in stages I and II that have failed to respond to conservative treatment are candidates to unload the joint by tenotomy of the transarticular accessory tendons. Long-term results have been very satisfactory (97%), eliminating or substantially reducing pain and returning patients to their activity. The procedure is contraindicated in severe (stage III) TMC joint osteoarthritis and in primary articular instability. PMID- 11280157 TI - Anatomy of the radial branches of the palmar arch. Variations and surgical importance. AB - In this study, the authors present the results of the investigation of arteries that replace digital collaterals of the radial side of the hand when the superficial palmar arch (SPA) does not develop completely. The replacement occurs with the first interosseous dorsal and the first interosseous palmar arteries. The former takes place through the middle developed branch that issues the radial collateral of the thumb and the ulnar collateral of the little finger whereas the latter takes place in three cases: a) presence of a branch entering into the retroadductor space and finishing like the previously mentioned; b) the origin of the trunk of the ulnar collateral of the thumb finger and the radial collateral of the index finger; c) due to the origin of both collaterals of the thumb finger. In cases when the SPA does not issue the fourth palmar collateral, this one is replaced by the second palmar interosseous artery, a branch of the deep palmar arch. These three arteries are combined to form three different basic kinds of arterial replacement that are described, adding a fourth group of exceptions that does not fit into any of the categories mentioned previously. PMID- 11280158 TI - A new method of diagnosing metacarpophalangeal instabilities of the thumb. AB - With a plain anteroposterior stress radiograph of the metacarpophalangeal joint of the thumb and observation of the displacement of the sesamoid bones, it is possible to interpret the injury mechanism and accurately diagnose the anatomical site of the ligament lesion. Fresh anatomical specimens were used to systematically investigate and document the role played in joint stability by each of the anatomical elements. Each ligament was sectioned and instability documented. A separate group (8) was used to replicate the different mechanisms responsible for ligament lesions. A radiological study was performed on 20 patients who had no history of trauma in their metacarpophalangeal joint and on another group of 17 patients with disease. Clinical instability was radiologically studied and showed a pattern of displacement of the sesamoid bones with respect to the metacarpal head, closely related to the different types of ligament lesions. When trauma occurs at the flexed metacarpophalangeal joint, the metacarpophalangeal ligament alone is torn and parallelism between the sesamoid bones and the metacarpal is not lost. In contrast, when trauma occurs with the joint extended, both the metacarpophalangeal and sesamoid metacarpal ligaments tear and the normal anatomical relationship between the sesamoid bones and the metacarpal head is lost. Therefore, if the radiograph shows no parallelism between the sesamoid bones and the metacarpal head, the injury is total and complete and requires surgical reparation. This highly reliable method can be implemented in any emergency department at low cost. It allows the extent and seriousness of the ligament lesion to be determined in an emergency. PMID- 11280159 TI - Anatomy of the flexor retinaculum of the wrist and the flexor carpi radialis tunnel. AB - The flexor retinaculum forms a retinacular bridge over the carpal tunnel extending from ulnar to radial direction. Its main function is to protect the contained without a significant mechanical action in supporting the transverse carpal arch. The osteofibrous tunnel of flexor carpi radialis is independent and presents four sections. The palmaris longus tendon presents a distal insertion forming the superficial layer of the aponeurosis palmaris. The flexor carpi ulnaris tendon has four distal insertions. PMID- 11280160 TI - The dorsal ligaments of the wrist. AB - The anatomy and function of the dorsal ligaments of the wrist have received increased attention over the past decade. The anatomy, variability, osseous attachments, and function of the dorsal radiocarpal (DRC) and the dorsal intercarpal (DIC) ligaments have been studied. More detailed anatomic and mechanical studies of the subregions of the intercarpal ligaments have also been recently reported. The DRC and the DIC form a lateral V configuration that allows normal carpal kinematics while maintaining an indirect dorsal stabilizing effect on the scaphoid throughout the range of motion of the wrist. The DRC and the DIC together, in their lateral V configuration, act effectively as a dorsal radioscaphoid ligament that has the ability to vary its length three-fold by changing the angle between two arms of the V. PMID- 11280161 TI - Anatomy and histology of the scapholunate ligament. AB - The scapholunate ligament links the scaphoid to the lunate. It runs transversally at its posterior aspect and obliquely at its anterior aspect, allowing significant relative motion between the two bones. From the neutral position to the full extension position, the lunate rotates by 28 degrees and the scaphoid by 30 degrees; from the neutral position to the full flexion position, the lunate rotates by 30 degrees, whereas the scaphoid rotation is 60 degrees because of the motion of the scaphoid around the capitate. The ligament's dorsal part is shorter and more resistant than the anterior part, allowing a pseudodissociation during flexion. Kauer described an additional movement of the scapholunate pair attributable to differences in the shapes of the scaphoid and lunate proximal poles. The scaphoid curve is more important and the scaphoid needs to glide on the lunate to maintain radioscaphoid congruity. As a result, there is sagittal ligament torsion. This can be a partial explanation for failure of scapholunate arthrodesis. This description of the scapholunate ligament is of interest to understand the relative importance of the three parts of this ligament. It can, in particular, explain the failure of ligamentous reconstruction that considers the scapholigament as a homogeneous structure. In addition, the three parts do not have the same tensile strength. The posterior part is the most resistant to tear forces and needs more than a 300 N tensile stress to fail. The anterior part fails with 150 N stress and the intermediary portion can withstand only a 25 N to 50 N stress. In comparison, the triquetrolunate ligament (which is also divided in three parts--anterior intermediate, and posterior) has failure coefficients opposite those of the scapholunate ligament: The anterior part is more resistant (300 N) than the posterior (150 N); the intermediate part has the same tensile strength as the scapholunate intermediate part. These biomechanical studies demonstrate the importance of the scapholunate ligamentous posterior part in controlling flexion and extension motion and the anterior part for rotational control. Both parts of the ligament are necessary for an harmonious functioning of the scapholunate pair. PMID- 11280162 TI - Localized medial triquetral-hamate instability: anatomy and operative reconstruction-augmentation. AB - Localized medial triquetral-hamate instability (LTHMI) is a relatively uncommon form of wrist instability. After an initial traumatic lesion elongating the triquetral-hamate (TH) complex (TH medial ligament and floor of the extensor carpi ulnaris [ECR] sheath), pain occurs during professional activities demanding ulno-radial movements (polo, tennis, music playing). Imbrication of the TH complex and augmentation with the ECU tendon is suggested for LTHMI. PMID- 11280163 TI - Anatomy and biomechanics of the interosseous membrane: its importance in the longitudinal stability of the forearm. AB - The interosseous membrane (IOM) links the ulna and the radius and acts as an extrinsic ligament, assisting the proximal radioulnar joint (PRUJ) and distal radioulnar joint (DRUJ) ligaments. It checks dissociating forces, transmits forces from one forearm bone to the other, and coordinates loading forces. The anterior plane of descending fibers from the radius checks the proximal displacement of this bone. Intermediate descending fibers are the strongest. The posterior plane, with ascending fibers from the radius, checks its proximal displacement. An early repair of a torn IOM is feasible and should be performed. This can be associated with an augmentation procedure. PMID- 11280164 TI - Rise in free intracellular calcium in HeLa cells infected with aggregative Klebsiella pneumoniae strains isolated from cases of diarrhoea. AB - BACKGROUND & OBJECTIVES: Klebsiella pneumoniae strains occasionally cause diarrhoea in humans. This study was done to determine the involvement of calcium in the pathogenesis of aggregative K. pneumoniae strains. METHODS: A total of nine strains of K. pneumoniae were tested for adherence assay in HeLa cell line. A representative strain CO-1215 was used for [Ca2+]i study using Fura-2 fluorescence. RESULTS: Infection of cultured HeLa cells with aggregative K. pneumoniae strain resulted in five-fold elevation of intracellular level of free calcium ([Ca2+]i) with maximum Ca2+ influx at 3 h after bacterial infection. Chelation of extracellular Ca2+ with [ethylenebis(oxyethylenenitrile)] tetraacetic acid and suspension of cells in Ca2+ free buffer suggested that the rise of Ca2+ in aggregative K. pneumoniae infected HeLa cells was due to influx of Ca2+ from extracellular medium. INTERPRETATION & CONCLUSIONS: This study showed aggregative adherence in HeLa cells and this adherence leads to influx of extracellular Ca2+. The unrestricted passage of calcium ions across cell membranes could cause phosphorylation of proteins involved in ion transport across the membrane, which could result in secretory diarrhoea. Further work is in progress to study the enterotoxicity of these strains in an in vitro rabbit intestinal model. PMID- 11280165 TI - Inactivation of chloramphenicol by Staphylococcus aureus biotype C from humans & animals. AB - During an investigation, 55 biotype C (bovine and caprine biotype) Staphylococcus aureus isolated from 43 cows suffering from mastitis, and 20 biotype C Staph. aureus strains from the nares and the side of nail-tips of the right thumbs of 20 farm workers (milkers and animal attendants) on six small dairy farms in Assam and Meghalaya were isolated. Three strains from the former and six strains from the latter from among the isolates on two of the farms were found resistant to chloramphenicol, when tested with a routine susceptibility test. Test of the organisms by the agar dilution method indicated that the resistant strains had a minimal inhibitory concentration for chloramphenicol of 32 micrograms/ml or more, while, the chloramphenicol sensitive strains and two reference strains, Staph. aureus ATCC 25923 and Micrococcus luteus ATCC 9341, had < or = 8 micrograms/ml. Two bovine and five human chloramphenicol resistant strains showed positive results when tested by the Gots test. When these strains were tested by a standard method in broth, containing 30 micrograms chloramphenicol per ml, all showed evidence of inactivating chloramphenicol up to a non-detectable level within 36 h. Inactivation of chloramphenicol by Staph. aureus has clinical significance. PMID- 11280166 TI - Observations on amphotericin B treatment of kala-azar given in a rural set up in Bihar, India. AB - BACKGROUND & OBJECTIVES: In a kala-azar endemic area in rural Bihar with a large number of patients unresponsive to sodium antimony gluconate and pentamidine, treatment with amphotericin B was tried in a rural set-up with the objective to cure these patients, and to assess whether such a centre could be run successfully in a rural set-up. METHODS: After thorough clinical examination and biochemical investigations, parasitologically confirmed patients who had haemoglobin above 5 g/dl, electrolyte imbalance if any, corrected, and ECG changes suggestive of myocardial damage stabilised after 10 days of bed rest were treated. Amphotericin B deoxycholate was infused at a dose of 1 mg/kg body weight, daily for 20 days. The adverse events were closely monitored. RESULTS: All 7 (100%) untreated patients of kala-azar, 258 (97%) of the 266 antimony and pentamidine resistant patients, and 31 (86%) of the 36 patients who had relapsed after a low dose regimen of amphotericin B, were cured with 20 infusions of amphotericin B. Eight (3%) patients of the antimony and pentamidine resistant group and 5 (14%) patients who had relapsed after low dose amphotericin B regimen required 25 infusions of amphotericin B to achieve parasitological cure. INTERPRETATION & CONCLUSIONS: With some precautions and proper management, all patients of kala-azar could be cured with amphotericin B in a rural set-up. A significant (P < 0.05) percentage of patients of group C relapsing after a low dose amphotericin B regime requiring 25 infusions for cure suggests that an adequate dose regime of amphotericin B should be given during the first course of treatment to prevent emergence of drug resistance. PMID- 11280167 TI - Carrier detection in non-deletional Duchenne/Becker muscular dystrophy families using polymorphic dinucleotide (CA) repeat loci of dystrophin gene. AB - BACKGROUND & OBJECTIVES: Carrier detection and prenatal diagnosis is of great importance for families with one or more sons affected with Duchenne/Becker muscular dystrophy (D/BMD). In about 35-40 per cent of these patients, the causative mutation does not involve gross rearrangement in the structure of dystrophin gene. In these non-deletional families, genetic counselling can be provided only by linkage analysis. The aim of the present study was to determine the carrier status of female relatives in north Indian families with non deletional D/BMD using highly polymorphic intragenic dinucleotide (CA) repeat markers. METHODS: Six short tandem repeats (STRs) spanning 5' (1), central (4) and 3' regions of the dystrophin gene were used to analyse 14 unrelated families comprising 68 individuals with 12 female siblings at risk of being carriers. RESULTS: Five female siblings inherited at risk STR haplotype, six inherited normal haplotype and one had meiotic recombination. The intragenic recombinations were observed in three families at the central region STR loci and in one family between the proximal and central regions of the gene. INTERPRETATION & CONCLUSIONS: Our study suggested that at least 6 STR markers spanning 5', central and 3' regions of the dystrophin gene are essential to ascertain one or more informative loci and to rule out recombinations in non-deletional D/BMD families for carrier analysis. PMID- 11280168 TI - An experimental study of tolerance among alcohol dependent individuals. AB - BACKGROUND & OBJECTIVES: Continued alcohol use leads to tolerance, however, some dependent individuals lose tolerance despite continued alcohol consumption. The exact mechanism for this is not known. This study evaluated tolerance in alcohol dependent patients in a treatment centre using multiple measures. METHODS: Male patients with alcohol dependence (DSM III R criteria) were chosen and detoxified in an inpatient setting. On day 14 of detoxification, each subject was given ethanol (0.75 g/kg body wt) mixed in an equal amount of placebo (cola) drink once and an equivalent amount of placebo (cola) during the other occasion in a single blind, randomised, cross over manner. Assessment of each subject was made using multiple measures (clinical, neuro-psychological tests, scales for subjective effect and blood alcohol levels), 30 min after intake of each drink. RESULTS: The subjects (n = 26) did not very under the two conditions (alcohol/placebo) as regards subjective effects, tests on logical memory and Bender Gestalt test (BGT). Cognitive screening scores though different under the two conditions, were within the normative range. Of these 26 subjects, 50 per cent showed clinical signs of intoxication after consumption of alcohol. These two groups (impaired vs unimpaired) were comparable on all base-line clinical parameters, assessment of euphoria and sedation, and various neuropsychological tests except BGT under the two conditions (placebo/alcohol). The non-tolerant (impaired) group scored significantly (P < 0.05) worse on BGT after alcohol consumption. INTERPRETATION & CONCLUSIONS: The study suggests that clinical tests were more sensitive in detecting intoxication. Further studies are needed to understand the mechanism of loss of tolerance. PMID- 11280169 TI - Escherichia coli heat stable enterotoxin receptors & guanylyl cyclase activity in the intestinal brush border membrane of hamsters & guinea pigs. AB - BACKGROUND & OBJECTIVES: Although Escherichia coli heat stable enterotoxin (STa) causes diarrhoea in laboratory animals, no studies were done to find out the species specific variation of distribution of the STa receptors in laboratory animals. The present investigation evaluates the density of STa receptors and the guanylyl cyclase (GC) activity in the small intestinal epithelial cells of hamsters and guinea pigs. METHODS: Brush border membrane (BBM) was prepared from the small intestines of hamsters and guinea pigs. Receptor binding assay, GC assay and autoradiography were performed to determine the density of STa receptors, the GC activity and molecular weights of the STa binding proteins respectively. RESULTS: The receptor densities, per mg BBM protein at equilibrium, were found to be 4.1 x 10(9) and 1.5 x 10(12) in hamsters and guinea pigs respectively. The GC activity was found to be lower in STa treated hamster BBM compared to that of guinea pig. Scatchard analysis of the stoichiometric data showed a linear plot, and STa bound with association constants of 0.31 x 10(12) M 1 and 1.04 x 10(12) M-1 in hamsters and guinea pigs respectively. Autoradiographic analysis of the SDS-PAGE, revealed that 125I-STa bound apparently to a 45 kDa membrane protein in hamster and a 115 kDa membrane protein in guinea pig. INTERPRETATION & CONCLUSIONS: It appears that a lower density of STa receptor exists in hamsters compared to that in guinea pigs. STa binds with a single population of STa receptors in each species with different ligand binding affinities. Also, the molecular weights of the STa binding proteins differ in these species. Moreover, the GC activity was found to be lower in hamsters than in guinea pigs. PMID- 11280170 TI - New approaches to an old and vexing problem: improving the results of SEPS: an overview. AB - Lipodermatosclerosis and chronic ulceration have been longstanding and vexing problems caused by chronic venous insufficiency (CVI). While traditional approaches have been mainly medical with the use of compression, bedrest, and elevation; operative therapy for CVI has now been shown to cause earlier healing with fewer ulcer recurrences. The development of subfascial endoscopic surgery (SEPS) promises a more elegant approach applicable to outpatient or day surgery. However, in a recent trial, early results showed a 22% ulcer recurrence at 30 months, which did not compare favorably with traditional approaches. We have used extrafascial perforator interruption for SEPS recurrence and have now modified our SEPS approach particularly for low-lying ulcers. This overview suggests use of a combination of SEPS with an extrafascial perforator division when skin change relates to retro or submalleolar perforating veins. Several procedures, rather than one intervention may be required in CVI to prevent or divert transmission of venous hypertension to areas of affected skin, including saphenous stripping, staged valveplasty and treatment of iliac occlusions. PMID- 11280171 TI - Operative treatment of a unilateral bicondylar fracture of the femur. AB - A case of isolated tangential fracture of the dorsal part of both condyles of the femur (bicondylar Hoffa fracture) is described. After open reduction, four lag screws were introduced through the fractured condyles to fix the fragments. A plaster cast was applied for a period of four weeks. Full weight-bearing was authorized after six weeks. Full range of motion of the knee was observed as early as after three months, as well as after twenty-one months. The mechanism and therapeutic implications are discussed and the literature is reviewed. PMID- 11280172 TI - Renal artery occlusion following blunt abdominal trauma. PMID- 11280173 TI - The pathology of complex fistula in ano. AB - Anal fistulae do not heal spontaneously without surgery. Knowledge of anorectal anatomy and function, and diligent examination frequently requiring anaesthesia are prerequisites for adequate assessment, classification and treatment of the pathology. Fistulography, endoanal ultrasound and MRI of the anorectum may have additional diagnostic value with therapeutic impact in complex and recurrent fistulae. Parks' classification is most useful because it is the best guide for surgical therapy. In the modern treatment of trans- or suprasphincteric fistula in ano fast or slow division of any part of the striated perianal musculature is to be avoided in order to prevent anal incontinence. PMID- 11280174 TI - Assessment of complex perineal fistulas. PMID- 11280175 TI - Difficult fistulae. AB - Fistula in ano is a common disorder. The goals of treatment are to cure the fistula with minimal loss of sphincter function and with minimal healing time. Fortunately about 90% of fistulae are simple and obey Goodsall's rule. These fistulae are easily treated by the "lay-open" technique. Treatment can however become much more difficult with increasing complexity of fistula tracks, higher internal opening with major sphincter involvement, atypical and secondary tracks or at recurrence. Understanding of the anatomy and the pathogenesis of fistulae is mandatory to identify the fistula tracks and the internal opening and to tailor the treatment accordingly. Endoanal flap repairs and the use of setons are most widely accepted for the treatment of difficult fistulae but many other options exist. Underlying factors or associated diseases such as inflammatory bowel, AIDS and other sexually transmitted diseases, carcinoma, radiotherapy, hidroadenitis or other obscure infections may influence the final outcome and often demand a specific approach. PMID- 11280176 TI - Fistula in ano: the place of rectal advancement flap technique. AB - Classical surgical techniques for anal trans- or suprasphincteric fistulas imply the division of the sphincters likely to induce postoperative continence impairment. The rectal advancement flap technique achieves healing of the fistula in a significant number of patients, while avoiding any sphincter division, and therefore the development of further incontinence. PMID- 11280177 TI - Rectovaginal fistulas. AB - The treatment of rectovaginal fistulas is controversial. The choice of the technique used for repair depends on many factors. Therefore the classification, etiology and treatment are discussed, in order to help decision making in the management of this troublesome disease. PMID- 11280178 TI - Unilateral pudendal thigh flap in the treatment of complex rectovaginal fistula. PMID- 11280179 TI - [Problems at the end of life and euthanasia. Viewpoint of the Conseil de l'Ordre de Medecins]. PMID- 11280180 TI - Euthanasia as an example of conflict between ethical precepts and entitlement rights. A surgeon's viewpoint. PMID- 11280181 TI - [Future goals of the SRBC. Societe Royale Belge de Chirurgie]. PMID- 11280182 TI - Total mesorectal excision in the treatment of rectal cancer: a review. AB - Despite the improvements in preoperative staging, surgical technique and adjuvant therapy, local recurrence remains a significant problem in rectal cancer surgery. Several patient- and tumour-related risk factors for the development of local recurrence have been identified and are being addressed by regimens of pre- or postoperative adjuvant therapy. Total mesorectal excision (TME) recently has been shown to result in a low recurrence rate even without the use of adjuvant therapy. Nevertheless, conclusive evidence in the form of a prospective randomized trial is to date not available. This paper describes the technique of TME and reviews the clinical and pathological data supporting its use in rectal cancer surgery. PMID- 11280183 TI - Informed consent. PMID- 11280184 TI - The patient's informed consent. Recent evolution of the case law. The physician's point of view. PMID- 11280185 TI - Informed consent: ethical point of view. PMID- 11280186 TI - Some practical advice concerning a medical lawsuit. PMID- 11280187 TI - A basic understanding of the judicial procedure. PMID- 11280188 TI - Informed consent. PMID- 11280189 TI - Informed consent: principles and recent developments. PMID- 11280190 TI - New surgical techniques and informed consent--safety first. AB - Surgical techniques always imply uncertainty about risks and outcome. Many law systems explicitly recognise the patient's right with respect to informed decision-making. This right refers to information about the risks and benefits, the experience of the surgeon, and the performance of the surgical team. In practice, the informed consent is difficult to obtain, and patients have often difficulty to recall the adequate information. Especially when risk and benefit assessment has to be done with the focus on a new technology that is expected to prevent disease in the future, safety of this less invasive technique compared with the current strategy is important. Therefore, it is also of importance to look at the mid-term and long-term follow-up. Appropriate knowledge of the technology involved, follow-up data, and risk information provided by a professional safety-committee should be the input of the informed consent procedure. PMID- 11280191 TI - Governmental proposals concerning the rights of the patient. Pandora's box? PMID- 11280192 TI - Tuberculosis notifications in Australia, 1998. AB - Since the inception of the National Mycobacterial Surveillance System (NMSS) in 1991, annual crude notification rates for tuberculosis have remained stable at between 5 and 6 per 100,000 population. In 1998, there was a total of 923 TB notifications in Australia of which 884 were new TB cases, and 39 relapsed cases. The corresponding annual crude notification rate for new and relapsed TB was 4.72 and 0.21 per 100,000 respectively. Seventy-seven percent of notifications that had a country of birth reported were overseas born. In keeping with trends observed over recent reporting years, the populations for which notified TB rates are highest include the overseas born from high prevalence countries and Indigenous Australians. The lowest rates of disease have continued to be reported in the non-Indigenous, Australian born population. Surveillance reports over the last seven years indicate that the rate of disease in this population is gradually declining. PMID- 11280193 TI - Measles outbreak among young adults in Victoria. AB - An outbreak of laboratory confirmed measles involving 30 young adults and an infant is reported. Of the young adult cases, 17 (57%) were hospitalised. After a trip to India, the primary case returned to Sydney early in January 2001 and then visited Melbourne infecting several individuals. Secondary spread appears to have occurred at a nightclub. On the basis of RNA typing the measles strain involved is of genotype 'D'. PMID- 11280194 TI - Varicella vaccine in post-exposure prophylaxis. AB - Evidence concerning the effectiveness of Oka-based varicella vaccines when administered following exposure to varicella zoster virus in domestic and hospital settings is reviewed. The evidence appears to support post-exposure use of Oka-derived varicella vaccines in children within 3 days of rash onset in the index case. Despite vaccination, a small proportion will develop mild, but infectious, chickenpox, especially if they have been exposed in the household setting. Controlled studies of post-exposure prophylaxis in adults using both Varilrix and Varivax II are still needed. The applicability of this approach to disease control in health care facilities and in community settings warrants wider discussion. PMID- 11280195 TI - Onshore catering increases the risk of diarrhoeal illness amongst cruise ship passengers. AB - Of 134 Queensland passengers on a cruise, 91 (67.9%) people reported various illnesses including 41 (30.6%) who reported diarrhoeal symptoms. Queensland passengers who ate while onshore at non-Australian ports were significantly more at risk of developing diarrhoeal symptoms than those who did not. Passengers were particularly at risk when they ate onshore while undertaking a tour compared with those who did not undertake this tour. Travellers should be warned of the possibility of contracting diarrhoeal illness from onshore catering. PMID- 11280196 TI - Communicable diseases surveillance. Presentation of NNDSS data. PMID- 11280197 TI - A primary school outbreak of pharyngoconjunctival fever caused by adenovirus type 3. AB - High rates of absenteeism in a North Queensland primary school, due to eye irritation, fever, headache, and stomach pain, were reported to the Tropical Public Health Unit in October 2000. Subsequent investigation demonstrated that the symptoms were due to adenovirus infection. Symptoms were consistent with a diagnosis of pharyngoconjunctival fever. At the height of the outbreak, about 40 per cent of students were absent. There was a strong association between the development of symptoms, and having been swimming on a recent school camp. Adenovirus could not be isolated from swimming pool water from the resort where the camp had been held. However, when inspected the swimming pool was not adequately chlorinated or maintained. It is probable that adenovirus infection was transmitted via swimming pool water at the school camp, and the outbreak might have been avoided by higher standards of swimming pool maintenance. PMID- 11280198 TI - Highlights from the physician services market survey. PMID- 11280200 TI - Getting the most from office temporaries. AB - Practically every medical practice finds itself short-staffed at one time or another. In some cases, a temporary employee is all that is needed to keep the practice running smoothly. Because of this, many temporary employment agencies today gear themselves specifically to filling both administrative and clinical positions in medical practices. This articles explores when and how to hire a temporary employee. It suggests strategies for preparing for the temporary employee's arrival, orienting the employee to the practice and assigned tasks, and evaluating the temporary worker's performance. In addition, this article offers ideas about working well with temporary employment agencies, as well as likely policies one would encounter when trying to hire a temporary employee permanently. PMID- 11280201 TI - How to keep track of the important details. PMID- 11280199 TI - Improving access to clinical offices. AB - Optimal access to office care requires a detailed understanding of a practice's capacity to provide care and demand for services. Once capacity and demand are known, they can be effectively managed to provide care today for those needs that arise today. Such a system of "open access" benefits clinicians and patients alike. This article describes specific steps a practice can take to achieve open access. PMID- 11280202 TI - A rational solution to billing problems. AB - For many practices, billing is a back-office task to be completed when the patient leaves the office. Historically, medical offices have placed their least experienced personnel at the front desk while the experienced staff remain hidden behind the scenes handling claims. Today's emphasis on customer service, however, has pushed the pendulum so that seasoned personnel are among the first with whom patients interact. Billing is a process that begins at the front desk when the appointment is made. Understanding the billing pipeline and how every action in a practice can contribute to the successful submission of "clean" claims and improved cash flow can encourage the physician and staff to think "billing" during each step of a patient's visit. This article will take you through the steps involved in the billing pipeline from patient registration to final payment or collection agency referral. For each step, we will point out how the actions of the staff affect billing and what can be done to assure accurate, clean claims are submitted promptly. PMID- 11280203 TI - Coding for surgical procedures. PMID- 11280204 TI - U.S. Supreme Court rules on physicians' role under ERISA group health plans. PMID- 11280205 TI - Pen-based computing: applications in clinical medicine. AB - Handheld, pen-based computing is an affordable, empowering technology that has application in small practice settings as well as in hospital-wide environments. Although hardware standards are evolving, there is now a stable base of applications, wired and wireless connectivity, and other features that make pen based computing an attractive alternative to paper and pen patient data acquisition. When combined with centralized data synchronization, pen-based computers can also serve as an affordable medium for the dissemination of clinical guidelines, standard formulary, and other patient-related data. Wireless, and to a lesser extent, wired pen-PDAs hold the potential to make web based electronic medical records accessible from anywhere at any time. PMID- 11280206 TI - Efficiently finding information on the Internet. AB - The Internet: a vast frontier of endless information. Despite this gold mine of knowledge, most users remain lost in cyberspace with little knowledge of what is available, where or how to find these treasures. Popular communities such as America Online have emerged because they help to organize the content into manageable pieces. But once you venture outside these confines, you get lost. This article will enable you to efficiently find general information and medically-related content on the Internet. PMID- 11280207 TI - Mediation sans litigation in malpractice. AB - Malpractice litigation is felt to provide a standard for practice. It can be costly both in terms of settlement awards and detrimental impact on the physician. Mediation offers opportunities to bypass that stringent legal process yet allows a resolution of disputes and allows proper redress of grievances. This article reviews the various factors that prevent its widespread application. PMID- 11280208 TI - A physician's guide to disability insurance. AB - Many physicians purchase disability insurance to protect their families and financial resources against unforeseen illness or injury. This article discusses various issues frequently encountered in disability claims and suggests ways of maximizing coverage and avoiding pitfalls that may negatively impact benefit claims. PMID- 11280209 TI - Peer review: confidentiality and privilege. PMID- 11280210 TI - Overview of CRICO's (Controlled Risk Insurance Company) 1999 professional liability claims data. PMID- 11280211 TI - [Studies on the relationship between serum albumin concentration and lipid peroxidation in the erythrocyte membrane of maintenance hemodialysis patients]. AB - We conducted a study on the long-term prognosis of maintenance hemodialysis patients with and without diabetes mellitus(144 cases). As a result, significant differences were observed in the long-term prognosis over a 15-year period between those with a serum albumin concentration of more than 3.8 g/dl and those with a serum albumin concentration of less than 3.8 g/dl. The group with a serum albumin concentration of more than 3.8 g/dl showed a better prognosis than the group with a concentration of less than 3.8 g/dl. We investigated the levels of thiobarbituric acid-reactive substances (TBA-RS) in the erythrocyte membrane of maintenance hemodialysis patients(23 cases), pre-dialysis uremic phase patients(12 cases) and healthy controls(7 cases). TBA-RS in the erythrocyte membrane of pre-dialysis uremic phase patients was higher than that observed in the other groups. The TBA-RS in the erythrocyte membrane in the maintenance hemodialysis without diabetes mellitus group was lower than that in the pre dialysis uremic phase group. The TBA-RS in the erythrocyte membrane in the maintenance hemodialysis without diabetes mellitus group thus indicated an inverse relationship with the serum albumin concentration, Kt/V and erythrocyte tocopherol contents. These results suggested that serum albumin protects against erythrocyte membrane lipid peroxidation, and that this function is related to hemodialysis. We thus conclude that hypoalbuminemia appears to increase the oxidative damage and acceleration of arteriosclerosis, while it also worsens the long-term prognosis of maintenance hemodialysis patients. PMID- 11280212 TI - [A case report of chronic chyluria probably due to Bancroftian filariasis, which showed hypoproteinemia]. AB - Proteinuria is commonly observed in patients with chyluria due to Bancroftian filariasis. However, whether or not hypoalbuminemia is caused by chyluria alone is still a matter of debate. This is because various forms of glomerulonephritis are complicated in such patients. Herein, we report a case we have recently encountered. A 72-year-old male was admitted to our division for further evaluation of nephrotic syndrome. He was from the Southernmost part of Japan, where Bancroftian filariasis has been epidemic, and had developed persistent chyluria over a period of nearly 50 years. There was no other past history of illness except for diabetes mellitus(DM) pointed out 3 months prior to admission. The physical and laboratory examinations on admission fulfilled the diagnostic criteria for nephrotic syndrome. Lymphoscintigraphy showed an intense tracer accumulation in both kidneys. A renal biopsy was performed. At the light microscopic level, the glomeruli looked normal. Edema of the tubulointerstitium was noted. At the electron microscopic level, effacement of podocyte foot processes was not observed. Immunofluorescent study did not show glomerular deposition of immunoglobulins and complements. He also had persistent microscopic hematuria. Automated urinary sediment analysis by real-time confocal scanning laser microscopy revealed red blood cells of the non-glomerular type. Taken together, these findings strongly indicated that hypoalbuminemia of this patient was caused by chyluria alone. In conclusion, a report of the present case provides strong evidence that hypoalbuminemia of a patient with Bancroftian filariasis could be caused by chyluria alone. PMID- 11280213 TI - [A case of Sjogren's syndrome presenting with hypokalemic myopathy due to renal tubular acidosis]. AB - A 37-year-old woman was admitted to our university hospital because of severe flaccid quadriplegia. Her laboratory data, lip biopsy and muscle biopsy findings were compatible with hypokalemic myopathy due to renal tubular acidosis(RTA) type I associated with primary Sjogren's syndrome. Kidney biopsy revealed chronic tubulointerstitial nephritis(TIN), consisting of focal mononuclear cell infiltration with tubulitis, interstitial fibrosis and tubular atrophy. Immunohistochemical analysis of the renal biopsy specimens showed that the infiltrating mononuclear cells were predominantly CD8+T cells, and CD68+ cells(macrophages), whereas CD4+ T cells were fewer in number. Following potassium administration and alkali therapy, hypokalemia and metabolic acidosis were ameliorated and limb palsy gradually subsided. Finally, RTA improved with prednisolon and short term cyclophosphamide treatment without supplemental potassium and alkali therapy. PMID- 11280214 TI - [Effect of interferon therapy on hepatitis B virus related membranous nephropathy associated with focal segmental glomerulosclerosis]. AB - We experienced a 24-year-old Japanese man, who was a hepatitis B virus carrier with nephrotic syndrome. Liver biopsy showed that he was suffering from chronic hepatitis (activity 2, fibrosis 2). Renal biopsy revealed membranous nephropathy(MN) with focal segmental glomerulosclerosis(FGS). Immunofluorescentic findings revealed the presence of HBe antigen along the glomerular capillaries as well as HBe antigenemia in circulation. Therefore, we diagnosed this case as HB virus-related membranous nephropathy associated with FGS lesions. He was treated with interferon(IFN) alpha-2b for over a month and angiotensin converting enzyme inhibitor. These therapies reduced urinary protein excretion from 4-6 g/day to 1 2 g/day, in accordance with a decrease in the titer of HBV DNA polymerase. The second renal biopsy revealed that the histological change from MN to membranoproliferative glomerulonephritis Type III after IFN therapy. These results suggest that IFN therapy might be effective for HB virus-related MN associated with FGS. PMID- 11280215 TI - [A case of gastric antral vascular ectasia (GAVE) with chronic renal failure]. AB - A 76-year-old woman was admitted to our hospital complaining of tarry stool, general fatigue and marked anemia(Hb 5.2 g/dl). Gastric endoscopic findings showed longitudinal red stripes and diffuse erythematous spots, indicating dilated vascular vessels. They resembled the stripes of a watermelon at the gastric antrum. The marked anemia was caused by chronic blood loss from the abnormally dilated mucosal and submucosal capillary veins in the gastric antrum. She was diagnosed as having gastric antral vascular ectasia(GAVE) with chronic renal failure(CRF). The association of GAVE and CRF is considered to be rare according to previous reports in Japan. Endoscopic argon plasma coagulation therapy was performed three times. After therapy, capillary dilatation disappeared, and the marked anemia was greatly improved. Argon plasma coagulation therapy was found to be a safe and effective procedure for this disease. Although GAVE is essentially a benign gastric disease, endoscopic therapy should be the treatment of first choice for this disease. PMID- 11280216 TI - Blood glucose changes following anticholinesterase insecticide poisoning. PMID- 11280217 TI - Cost burden to diabetic patients with foot complications--a study from southern India. AB - AIM: To study the economic burden of management of diabetes in patients with foot complications, as a large number of them suffer from foot complications of varying severity. This study relates to direct cost to diabetic patients with foot complications. MATERIAL AND METHODS: An illustrative sample of 270 Type 2 diabetic subjects, 164 without foot complications (Control group, Group 1) and 106 with foot complications (Group 2) were studied. They were available for the study during a six month period from January to June 1998. Group 2 had two sub groups, i.e., those who needed out-patient (OP) treatment only (n = 23) and those who needed treatment in the hospital (HP) (n = 83). The study subjects were interviewed personally by the educator to collect demographic data and treatment expenditure. RESULTS: Total median expenditure incurred by the diabetic subjects without foot complications (Group 1) was Rs. 4373/- and by those with foot complications (Group 2) was Rs. 15,450/-. Patients who required hospitalised treatment incurred higher expenses than the OP patients, towards doctor's fees and hospitalisation (P < 0.0001). The percent of total income spent by the HP patients was higher than by the OP patients (P < 0.02). CONCLUSIONS: Diabetic subjects with foot problems incur very heavy expenditure in the treatment process. Most of the direct costs of diabetes treatment results from its complications. The hospitalisation costs for the complications of diabetes are particularly heavy. This underscores the need to reduce complications and also their economic burden. PMID- 11280218 TI - Spectrum of renal lesions in HIV patients. AB - BACKGROUND: A variety of renal lesions have been reported in HIV positive patients from western world however there is paucity of Indian data. METHODS: Over a four year period, all hospitalised HIV positive patients were screened for renal involvement. Screening was done with urinalysis. Those with abnormality in urine examination underwent further assessment with clinical, biochemical, immunological profile and renal biopsy. Renal histology was studied by light and electron microscopy. RESULTS: Twenty-five (17.6%) of the 142 patients screened, had proteinuria/abnormal urinary sediment however none of the patient had proteinuria in nephrotic range. Fourteen of these 25 patients were asymptomatic while others had AIDS. Renal biopsy was studied by light microscopy in all and by electron microscopy in 11 cases. On histology mesangioproliferative GN was encountered in eight, focal segmental glomerulosclerosis in four and collapsing GN in one patient. In two cases cryptococcal infiltration and in one lymphomatous deposits were seen in glomerulus and interstitium. In one patient interstitium showed granulomas and in other three mononuclear cell infiltration. Histology was normal in 8 (32%) patients. On EM visceral cell hyperplasia and vacuolisation was seen in all, two had collapse of glomerular basement membrane and in three cases tubuloreticular structures were seen. There was no co-relation of renal histology with duration or severity of the disease (p > 0.05). No deterioration of renal function was seen over a short follow up period of 4.2 months (1-20 months). CONCLUSION: This study highlights that HIV patients exhibiting abnormal urinary sediment usually have underlying renal lesion and at times unexpected opportunistic infections may be present. PMID- 11280220 TI - Seroprevalence of human immunodeficiency virus infection in an infectious disease hospital. AB - AIMS AND OBJECTIVES: To analyse clinicopathological features of HIV infected patients admitted in an infectious disease hospital in Mumbai. MATERIAL AND METHODS: Retrospective study of 501 patients admitted from 1st September, 1996 to 28th February, 1998 and screened for HIV status out of clinical suspicion was carried out. HIV seropositivity was established by double confirmation of spot test results with microwell Elisa test. RESULTS: HIV seropositivity was seen in 39.92%. Out of these 94% were adults with male preponderance and 96.5% had only HIV-1 infection. The significant clinical features in HIV positive patients were chronic diarrhea (51.5%), prolonged fever (41.5%) and history of exposure (34%). Pulmonary tuberculosis could be diagnosed in 19.5% of HIV positive patients. In patients with prolonged/recurrent jaundice HBsAg and HIV was noted as a coinfection within 10/42 cases tested. CONCLUSIONS: Increasing prevalence of HIV seropositivity was noted in patients admitted to an infectious disease hospital. Association of HIV infection with tuberculosis and in icteric cases with HBsAg was significant. The HIV screening should be carried out in patients with prolonged illness resistant to usual mode of treatment. PMID- 11280219 TI - Impact of hepatitis C virus infection on renal transplant outcome in India--a single centre study. AB - BACKGROUND: Hepatitis C virus (HCV) infection is currently the main cause of hepatotropic viral infection in renal transplant (RT) recipient throughout the world. Contrary reports are available as regard graft and patient survival and liver disease outcome in these patients. From India, outcome of HCV positive patients following RT has not been documented. Herewith, we present results of RT in HCV positive patients at our centre. METHODS: Study design was prospective case control with primary end point being graft and patient survival and the exposure being HCV infection. Between June 1995 till February 1998, 128 patients had RT at our hospital, of which, 37 (28.9%) were anti-HCV positive at the time of RT. All the patients were on triple immunosuppressive therapy. As a policy of unit, none of the donor had HBV and/or HCV infection. Anti-HCV positive patients formed the subjects (Gr. I), while anti-HCV negative patients severed as control (Gr. II). Anti HCV was done using 3rd generation ELISA tests kit. HCV-RNA could not be done due to non-availability. None of the positive patient was treated with anti-viral therapy. Acute rejection, serious infections, patient and graft survival and outcome of liver disease was compared in these patients. RESULTS: Mean age of the patients, number of males, number of pre-RT haemodialysis and blood transfusion, donor age and HLA-mismatch were comparable in both the groups. Mean follow-up in Gr. I was 28 +/- 9.4 months and in Gr. II 31.4 +/- 7.6 months. At the end of this follow-up, acute rejection was seen in 43% and 33.3% patient in Gr. I and II respectively. In Gr. I, serious infections were seen in 30% while the same in Gr. II was 11.8% (p < 0.01). There was no difference in graft survival in Gr. I and II (72% and 66%) and the patient survival were also similar (72% and 66%). Of the deaths in Gr. I, 80% died of sepsis and 20% died of liver cell failure related to one each of hepatitis B and hepatitis E. Of the deaths in Gr. II, 65% died of sepsis and 17% died of hepatic cell failure. But, there was no difference in causes of deaths in these two groups. In both the groups, none of liver related death was due to isolated HCV infection. There was no effect of donor age, HLA mismatch, number of haemodialysis and pre-RT blood transfusion on the survival of graft as well as patient. CONCLUSION: In conclusion, HCV infection is major problem in RT with us. In a short follow-up of nearly 30 months, graft and patient survival is same in HCV positive and negative patients. However, serious infections are significantly more common in HCV positive patients. PMID- 11280221 TI - Efficacy of noninvasive nasal mechanical ventilation in acute respiratory failure: monitored by breathing patterns. AB - OBJECTIVES: Till a decade back, the mainstay delivery of mechanical ventilation to patients with acute respiratory failure was through the endotracheal tube. To obviate the various complications of endotracheal intubation, noninvasive mechanical ventilation (NIMV) techniques were devised which could be used outside the confines of an ICU setting, and have been employed by several workers to achieve a high success rate. METHODS: Twenty patients of acute respiratory failure (ARF) were treated with NIMV. The latter was delivered by assist control mode of ventilator and nasal CPAP mask. Improvement of the illness was monitored by serial ABG analysis and breathing pattern parameters over one week. RESULTS: The application of NIMV was successful in reversing the illness in 17 patients (85%) and the other three patients had to be intubated. The pH rose from 7.267 +/ 0.087 at presentation to 7.411 +/- 0.032 (p < 0.00005), the PaCO2 dropped from 85.17 +/- 13.48 mmHg to 46.27 +/- 3.79 mmHg (p < 0.00005). The PaO2 improved from 52.36 +/- 11.14 mmHg to 63.60 +/- 7.55 mmHg (p < 0.005) on the seventh day. The respiratory rate (RR) decreased from 30.07 +/- 6.10 breaths/min to 22.05 +/- 4.05 breaths/min (p < 0.00005), the %RC (percent rib cage) also showed a significant reduction in rib cage contraction from 61.7 +/- 10.17 to 42.76 +/- 10.66 (p < 0.00005). A marked improvement during the initial four hours of application of NIMV was observed in all the 17 patients. CONCLUSIONS: It was concluded that NIMV resulted in marked improvement in both the breathing pattern and blood gas parameters in patients of ARF; and PaCO2, pH, RR, %RC served as the best indicators of improvement. NIMV was observed to be most useful in those patients who had CO2 retention. PMID- 11280222 TI - Prevalence of obesity in adults--an epidemiological study from Kashmir Valley of Indian Subcontinent. AB - OBJECTIVE: Obesity is a health problem in the majority of the developed countries and is emerging as a serious problem in the developing countries. In this study we examined 5083 Kashmiri adults to determine the prevalence of obesity. METHODS: In this epidemiological study, after multistage sampling procedure from all the six districts of Kashmir Valley, 5083 Kashmiri adults (males and non-pregnant females of > or = 40 years age) were examined for body mass index (BMI) and waist to hip ratio (WHR). RESULTS: Out of 5083 study subjects, 2496 were males and 2587 were females. BMI ranged between 14.6 to 38.5 kg/m2 in males and between 13.6 to 42.6 kg/m2 in females. BMI in females was comparatively more than that in males (23.88 +/- 3.94 Vs 22.30 +/- 3.11, p < 0.001). WHR ranged between 0.68 to 1.16 in males and between 0.65 to 1.16 in females. Again females had comparatively more WHR than their male counterparts (0.935 +/- 0.055 vs 0.926 +/- 0.055, P < 0.001). According to BMI, the overall prevalence of obesity in the study population was 15.01%; the prevalence of obesity in males was 7.01% and in females 23.69%. CONCLUSIONS: We conclude that obesity is a growing problem even in developing regions like ours. It is more common in females and in urban population. PMID- 11280223 TI - Prazosin therapy and scorpion envenomation. AB - BACKGROUND: 25-30% fatality due to acute pulmonary oedema in victims of Indian red scorpion (Mesobuthus tamulus) sting have been reported from Western Maharashtra, India. The advent of prazosin in recent years has revolutionized the management of severe scorpion sting cases. Majority of cases developed acute pulmonary oedema in 4-8 hours in a hospital setting irrespective of control of their arterial blood pressure with six hourly oral prazosin regimen, these cases recovered with extra dose of prazosin. We developed a standardised protocol for acute phase of treatment of these cases with the aim of preventing the development of pulmonary oedema. METHOD: We compared scorpion sting cases managed by non-protocol conventional (NPC) treatment and those by standardised protocol (SP) that included three hourly dose of oral prazosin. SP group included severe scorpion sting cases admitted to general hospital at Mahad in the year 1998 (Jan. Dec.). While those admitted in the year 1997 (Jan.-Dec.) before the SP was implicated were the NPC group. FINDING: Characteristics on arrival of severe scorpion sting patients SP (n-17) and NPC (n-15) groups were similar that more case 6 (35%) from SP group had several hypertension on arrival. On arrival two cases from SP group and one from NPC group had pulmonary oedema. 16 (94.11%) patients from SP group recovered uneventfully, compared with 8 (53.33%) in NPC group (p-0.05). 0% Vs 5 (38.46%) developed acute pulmonary oedema (p < 0.0001) from SP and NPC group respectively, three (one had on arrival two patients during hospitalization) from NPC group had massive pulmonary oedema recovered with i.v. nitroprusside drip (SNP). While from SP group one had massive pulmonary oedema on arrival recovered with i.v. SNP, other one had pulmonary oedema recovered with oral prazosin. Cool extremities (vasoconstriction) persisted 11.5 (5-20) VS 18 (12-26) hours in SP and NPC group respectively. INTERPRETATION: Compared with NPC management; development of acute pulmonary oedema prevented by standardised protocol regimen at rural setting. PMID- 11280224 TI - Recent advances in the pathobiology of septicemia and septic shock. PMID- 11280225 TI - Diabetes mellitus and oncology. PMID- 11280226 TI - Pictorial CME. Isospora belli and Cyclospora cayetanensis in a case of chronic diarrhoea in an immunocompromised host. PMID- 11280227 TI - The newer unconventional indications of permanent pacemakers. AB - In recent years, the indications for permanent pacemakers have expanded. The interest has focussed on hypertrophic cardiomyopathy, dilated cardiomyopathy and a new entity called hypertensive hypertrophy with cavity obliteration (HHCO). Pacemaker therapy is establishing itself for the prevention of atrial fibrillation. Pacing for neurocardiogenic syncope with newer pacing mode has encouraging datas. Pacemaker for long QT syndrome, after cardiac transplant and for haemodynamic improvement in occasional cases of first degree atrio ventricular block is getting attention. The AHA and ACC guidelines updated in 1998 for implantation of cardiac pacemakers, now include several of these newer indications. PMID- 11280228 TI - Glycosuria in organophosphate and carbamate poisoning. AB - OBJECTIVE: Puzzled by the observation of occurrence of transient glycosuria in several patients admitted with organophosphate and carbamate compound poisoning, we undertook a critical analysis of this observation. METHODS: Of the fifty-one consecutive patients admitted to the intensive care unit with organophosphate and carbamate compound poisoning, in 23 subjects the nature of the compound consumed was known; these were studied. The occurrence and duration of glycosuria, its magnitude, associated hyperglycemia if any, and correlation thereof were recorded. RESULTS: Sixteen out of the 23 subjects (69%) demonstrated transient glycosuria. There were 13 men and 10 women. 10 subjects had euglycemia associated with glycosuria; 6 subjects had transient glycosuria with hyperglycemia (defined as random glucose over 160 mg%). None of the subjects had diabetes mellitus and the pre-hospital-discharge blood sugars were in the normal range. CONCLUSION: Glycosuria (renal), albeit transient, was noted in a proportion of subjects admitted with organophosphate and carbamate poisoning. The exact etiology of this is unclear but in the light of recent literature, it is likely that oxidative stress at the renal tubular level leading to renal tubular damage may be the most likely explanation. Further, larger studies are needed to elucidate this is detail. PMID- 11280229 TI - Prevalence of hypertension amongst Mumbai executives. AB - OBJECTIVE: To find out prevalence of hypertension amongst Mumbai executives. METHODS: Data of annual medical check-up of 1653 executives was evaluated. Blood pressure was measured as per JNC VI/WHO guidelines. RESULTS: Overall prevalence of hypertension amongst Mumbai executives was 26.86%. 21.28% of executives who were hypertensive based on causal reading were later found to have normal or high normal blood pressure. CONCLUSION: For all epidemiological surveys, blood pressure must be recorded on at least two subsequent occasions after initial screening. PMID- 11280230 TI - Adrenoleukodystrophy presenting as Addison's disease in childhood. PMID- 11280231 TI - Empty sella syndrome presenting as galactorrhoea. AB - A prolactin secreting tumour is the commonest cause of the amenorrhoea galactorrhoea syndrome. Galactorrhoea is a rare presentation of an empty sella syndrome. The empty sella syndrome commonly presents with headache and visual impairment and occasionally with endocrine disturbances in hypertensive middle aged women. The authors present a case of hyperprolactinemia resulting in galactorrhoea in a middle aged lady associated with a primary empty sella syndrome. PMID- 11280232 TI - Disseminated pulmonary hydatid disease presenting as multiple cannon ball shadows in human immunodeficiency virus infection. PMID- 11280233 TI - Unilateral inter-nuclear ophthalmoplegia in systemic lupus erythematosus. AB - We report a case of a 14 year old girl with SLE who developed neurological involvement in the form of posterior internuclear ophthalmoplegia (pINO). An MRI showed lesion involving pons which corroborated with the pINO. PMID- 11280234 TI - Acute lymphoblastic leukemia presenting as breast mass. AB - We present here a 34 years female who presented with bilateral breast lumps as the initial manifestation of acute lymphoblastic leukemia. She was treated with consolidation chemotherapy and showed good response. PMID- 11280235 TI - Guillain Barre' syndrome--in an HIV seropositive subject. PMID- 11280236 TI - Benign intracranial hypertension: a rare manifestation of Behcet's syndrome. PMID- 11280237 TI - Obstructive jaundice due to a rare cause of biliary stricture. PMID- 11280238 TI - Pregnancy in a patient with prolactinoma off treatment. PMID- 11280239 TI - Post herpes zoster galactorrhoea. PMID- 11280240 TI - The venous hum. PMID- 11280241 TI - Pregnancy with cerebral malaria. PMID- 11280242 TI - Endothelin, nitric oxide and prazosin. PMID- 11280243 TI - CPC as a teaching module. PMID- 11280244 TI - Poetry and medicine: the other side. PMID- 11280245 TI - Alien hand syndrome. PMID- 11280246 TI - Young onset abdominal aortic aneurysm. PMID- 11280247 TI - Strategies for the management of 'difficult to treat' patients with tuberculosis. PMID- 11280248 TI - PHLS works towards effective interaction with primary care. AB - The PHLS primary care initiative was set up to develop and maintain links with primary care professionals, including researchers, GP research networks, and primary care groups in order to prioritize research and laboratory service developments. Such collaboration has spawned the development of antibiotic guidance, which has been posted on the Internet, and participation of the PHLS in other infectious disease guidance (for example, Prodigy). The implementation of the NHS information strategy will further facilitate the development of and access to local and national primary care guidance, laboratory reporting, and participation in surveillance. Primary care professionals identified several main research priorities in an exercise that facilitated collaboration in the development of research projects and grant applications. These priorities are the natural history of infections presenting to GPs, antibiotic resistance, the role of clinical scores and the laboratory in the diagnosis of infection, the importance of patient and professional attitudes in modifying antimicrobial prescribing, and the role of health professionals in community infection control. Current rapid changes in primary care in the new NHS--such as NHS Direct and the establishment of walk-in centres--may affect the surveillance of communicable disease by offering opportunities for improved data collection but also by posing threats to current systems of surveillance. PMID- 11280249 TI - Do health care workers need to wear masks when caring for patients with pulmonary tuberculosis? AB - Outbreaks of hospital acquired tuberculosis (TB) in the 1990s, some of which affected staff, highlight the fact that TB still poses a risk to health care workers (HCWs). This risk is best minimised by the primary controls of early identification and treatment, good infection control practice such as early isolation, patients covering mouths when coughing, and engineering controls such as negative pressure isolation. There is no direct evidence to prove that the use of respiratory protective devices has prevented HCWs from acquiring TB, but modelling has shown that environmental controls are not enough to prevent exposure. Masks that filter aerosols (including TB) and have at least 95% efficiency of filtering particles 0.3 micron in diameter can reduce exposure. Environmental controls in the United Kingdom (UK) do not (always) include a specified minimum number of air-changes per hour or negative pressure. Therefore it seems reasonable to advise that HCWs should use masks as advocated by national guidelines for high risk procedures. Masks are also of value when entering any body cavity or dissecting any viscus/organ in which TB is suspected, as in the necropsy room or operating theatre. This is recommended in current guidelines from the United States but not those from the UK. PMID- 11280250 TI - Audit of bloodborne virus infections in injecting drug users in general practice. AB - Two hundred and ninety patients attending a single general practice in Edinburgh were known to have used illegal drugs, 145 of whom were identified as past or present injectors. Data on bloodborne virus infections and immunisation against hepatitis B virus (HBV) were gathered during 1998, attempts were made to improve the level of testing for bloodborne viruses and immunisation against HBV, and follow up was carried out between October 1999 and February 2000. One hundred and fifteen patients were studied in detail. Evidence of previous HBV infection was found in 31 of 71 tested in 1998 (44%) and 40 of 99 tested at follow up (40%). In 1998 54 out of the 75 tested for hepatitis C antibodies (72%) were positive compared with 73 out of 108 (68%) at follow up. Twenty-six of the 80 tested for HIV antibodies were positive in 1998 (33%) and 26 of 105 at follow up (25%). Large numbers of injecting drug users in our study were found to be not immune to hepatitis B and required immunisation. An abbreviated protocol for immunisation was devised, including post vaccination checks and boosting as necessary. Hepatitis C testing was requested after counselling in most cases, resulting in important and positive interventions. Prevention opportunities for all three bloodborne viruses were identified. PMID- 11280251 TI - Knowledge and attitudes of hospital staff to occupational exposure to bloodborne viruses. AB - In order to assess awareness of occupational risk of exposure to bloodborne viruses a questionnaire was sent to 245 health care workers, representing a 10% sample of employees with patient contact in a large teaching hospital in Scotland, stratified by occupational group. One hundred and eight questionnaires (44%) were returned. Seventy per cent of respondents in laboratory and clinical areas described themselves as having sufficient knowledge for their own area of practice, but many gave incorrect answers or expressed uncertainty about the infectivity of HIV and hepatitis B and hepatitis C viruses. Ninety-four respondents were unaware that a regimen containing more than one antiretroviral drug is now recommended for post exposure prophylaxis of HIV infection, 37 thought they had been at risk of bloodborne viral infection and had contacted the occupational health department for advice, and 68 respondents disagreed with guidelines from the United Kingdom's General Medical Council on testing of patients for bloodborne viruses. The results indicate a need for educational initiatives for new and existing staff. PMID- 11280252 TI - Acute hepatitis B in two patients transmitted from an e antigen negative cardiothoracic surgeon. AB - Hepatitis B virus (HBV) infection was transmitted by a locum cardiothoracic surgeon to two patients during coronary artery bypass surgery. Both patients presented 12 weeks after surgery and developed serious clinical illness. The surgeon was known to be hepatitis B surface antigen (HBsAg) positive, hepatitis B e antigen (HBeAg) negative, and to have antibodies to HBeAg (anti-HBe). Sequences of regions of the HBV surface and core genes from the patients and surgeon were indistinguishable. An exercise was undertaken to notify all patients on whom the surgeon had operated while employed at the hospital where the transmissions occurred. One hundred and twenty-three out of 126 patients were tested. No evidence of transmission to any other patient was found. Revised recommendations by the UK Health Departments as to which health care workers should be permitted to perform exposure prone procedures have recently been published. PMID- 11280253 TI - Comparison between self-reported hepatitis B, hepatitis C, and HIV antibody status and oral fluid assay results in Irish prisoners. AB - Self-reported hepatitis B virus (HBV), hepatitis C virus (HCV), and HIV infection status was compared with the results of oral fluid assays of antibodies to these viruses in prisoners from nine of the 15 prisons in the Republic of Ireland. A total of 1205 out of 1366 prisoners completed a confidential questionnaire and 1193 provided analysable oral fluid specimens for testing for antibodies to HBV core antigen (anti-HBc), HCV (anti-HCV), and HIV (anti-HIV). The self-reported prevalence of hepatitis infection (hepatitis B: 5%; hepatitis C: 19%) was lower than that derived from oral fluid assays (anti-HBc: 9%; anti-HCV: 37%). The self reported prevalence of HIV infection was similar to that found by oral fluid assay (2%). Many discrepancies were found between self-reported results and the results of oral fluid assays. Of those who reported being positive for HBV, HCV, or HIV, 48%, 5%, and 58%, respectively, tested negative on the oral fluid assay. Of those who reported a previous negative test result for HBV, HCV, or HIV, 10%, 37%, and 2%, respectively, had positive oral fluid assays. Self-reports of hepatitis and HIV infection status are unreliable and should not be used as a basis for planning preventive and treatment services for prisoners. All prisoners should have the opportunity to be tested for HBV, HCV, and HIV infection. PMID- 11280254 TI - Presentation with influenza-like illness in general practice: implications for use of neuraminidase inhibitors. AB - General practitioners in the Midlands Research Practice Consortium (MidReC), combined list size 140,000, completed questionnaires about 918 patients in whom they had made working diagnoses of influenza-like illness during an outbreak of influenza A H3N2 from 1 December 1999 to 4 February 2000. Adults, more females than males consulted most, reflecting the age and sex distribution reported to the Royal College of General Practitioners Weekly Returns Service. Illness at presentation was considered severe in 4%, moderately severe in 49%, mild in 45%, and asymptomatic (for example, attended for certificates) in 1% of patients. In seven tenths of patients, the practitioner estimated that the likelihood of influenza was 70% or more and in just over half, 80% or more. Half of patients aged over 75 years were seen at home, but only 7% of those under 55 years. Less than a quarter of patients consulted within two days of having become ill, with the highest consultation frequency on the third and fourth days. Preschool children presented earliest: 75% were seen within two days, compared with only 17% of adults over 75 years. Four fifths of patients were seen on the same day as they contacted the practice, and 12% on the following day. Given the brief time window for effective antiviral treatment, only a small proportion of patients are likely to be prescribed these drugs unless consulting behaviour, especially in elderly people, changes considerably. PMID- 11280255 TI - Influenza surveillance in England and Wales: October 1999 to May 2000. AB - The period of increased influenza activity in England and Wales in the winter of 1999/2000 was associated with considerable morbidity and mortality and well publicized pressure on hospital services. The influenza activity coincided with the regular annual increase in respiratory syncytial virus infections and the Christmas and New Year holiday period. Consultation rates with general practitioners for influenza-like illness did not reach 'epidemic' levels but were higher than seen in many winters and comparable with those seen in two out of the previous three winters. In common with those winters, attack rates for influenza like illness and acute bronchitis were especially high in elderly people among whom complications of acute infection and hospital admissions increased. Excess mortality due to influenza during this period appeared to be substantial but was not as high as seen in the last epidemic (1989/90). PMID- 11280256 TI - Media management of a community outbreak of meningococcal meningitis. AB - Cases and, particularly, clusters of meningococcal meningitis often create high levels of public concern and attract the interest of news media. We describe below our experience of managing intense local and national media interest during a community outbreak of meningococcal disease in Derbyshire. Our learning is set out as ten key recommendations. Four of these are around managing the media, including using a proactive press release, providing detailed briefings, using a single spokesperson and coordination of the response by a press officer experienced in media management. Another four describe how to deliver an appropriate on-site response, often requested during community intervention programmes. The two final recommendations relate to ensuring good communication and supporting staff during what is an intensely stressful period. We hope our experiences may help others if faced with a similar problem. PMID- 11280257 TI - Survey of the prescribing of chemoprophylaxis to index cases of invasive meningococcal disease. AB - Chemoprophylaxis is given to contacts of cases of invasive meningococcal disease to reduce the risk of secondary cases by eradicating carriage. In the United Kingdom index cases are also recommended to receive chemoprophylaxis. This is usually undertaken by the clinical team managing the case. One hundred and fifty cases of probable meningococcal infection notified to the consultants in communicable disease control in a local health authority were reviewed to identify the proportion receiving chemoprophylaxis and to examine the final clinical outcome, in terms of diagnosis, of each case. Twenty-five per cent of notified cases (37) did not receive chemoprophylaxis and this proportion varied significantly between three local hospitals. We estimate that 15 of the 37 index cases who did not receive chemoprophylaxis were likely to have had invasive meningococcal disease. PMID- 11280258 TI - Family clusters of campylobacter infection. AB - Possible family clusters of campylobacter infection among isolates referred to a single, large public health laboratory were reviewed to determine the proportion of clusters associated with more than one strain of campylobacter. A total of 23 clusters, each made up of members of a single household infected within two months of each other, were identified between September 1996 and January 1999. These accounted for about 5% of all campylobacter infections confirmed during the study period. Thirteen of these families were infected with single strains of C. jejuni and one with C. coli. In ten families specimens were taken over intervals ranging from six to 56 days. In eight clusters more than one strain of C. jejuni was identified and in one family two patients were infected with C. jejuni and one with C. lari. These findings are consistent with the observation that potential sources of campylobacter infection may be contaminated with more than one strain. PMID- 11280259 TI - National assessment of prevalent diagnosed HIV infections. AB - HIV infection is associated with high treatment and care costs and subject to large differences in prevalence between health districts. Equitable distribution of resources requires information provided by an annual national survey of prevalent diagnosed HIV infections (SOPHID). This measures HIV caseloads by health district of residence throughout England, Wales, and Northern Ireland and is used to inform local public health professionals and to improve allocation of government funding for HIV prevention and care. Survey totals are adjusted by underreporting and non-attendance factors to produce a more accurate assessment of the total caseloads. On average the combined adjustments increase the reported caseload by 14.7% annually. Adjusted prevalence estimates ranged from 14,164 in 1995 to 18,460 in 1998, an increase of 30%. PMID- 11280260 TI - Epidemiology of chickenpox in Scotland: 1981 to 1998. AB - Scotland requires cases of chickenpox to be notified formally and maintains comprehensive data on general practice consultations, hospital admissions, abortions, laboratory reports, and mortality associated with the disease. These were used to investigate the age specific incidence of chickenpox for the years 1981 to 1998. The general trend was towards decreased age at infection: most infections now occur in the 1 to 4 year age group, rather than among schoolchildren. Hospital admissions for which a diagnosis of chickenpox was recorded increased, mainly in the under 5 and 25 to 34 year age groups. These data, which we believe to be among the most comprehensive available on current chickenpox epidemiology, may be used to inform preventative policy, particularly now that a live vaccine for the prevention of primary varicella infection is available. If vaccination against varicella is introduced in the United Kingdom, these data will provide a baseline against which to assess its impact on primary varicella infection. PMID- 11280261 TI - Infection of foot ulcers with Staphylococcus aureus associated with increased mortality in diabetic patients. AB - Diabetic patients with foot ulceration have a poorer prognosis than those without ulceration. The reason for this is unclear, but there is considerable interest in the putative links between infection and atherogenesis, and it is notable that diabetic foot ulcers (DFU) are often infected with Staphylococcus aureus and the main cause of death in DFU patients is ischaemic heart disease. We examined the 5 year survival of 71 diabetic patients who presented with foot ulcers that were newly infected (Sa group, n = 56) or not infected at all during the study period (non-Sa group, n = 15) with S. aureus. Twenty-nine patients (52%) infected with S. aureus died compared with three patients (20%) whose foot ulcers were not infected with S. aureus. The patients in the two groups were similar in age and duration of diabetes. The overall five year mortality rate was 10.4% per year for those infected, significantly higher than the average of 4.0% for patients without infection (p = 0.015). None of the patients was bacteraemic or died directly from sepsis. Infection of DFU by S. aureus may increase the risk of death in diabetic patients. PMID- 11280262 TI - Outcome of medical screening of Kosovan refugees in Ireland: 1999. AB - In March 1999 armed conflict broke out in Kosova and about 900,000 ethnic Albanians were displaced. We reviewed the health care offered to the 945 Kosovan refugees who arrived in Ireland in 1999, which included screening for tuberculosis (TB) and hepatitis B. On arrival in Ireland 540 refugees had already received oral polio vaccine (57%), 512 diphtheria, tetanus, and acellular pertussis or diphtheria and tetanus vaccine (54%), 310 BCG (33%), 207 measles, mumps, and rubella vaccine (22%) and 60 Haemophilus influenzae type b (6%). Twelve refugees were diagnosed with TB. Twenty-six refugees were HBsAg positive (3%) and 168 were anti-HBcAg positive (18%). Organised screening of Kosovan refugees on a voluntary basis (uptake > 95%) revealed low percentages who had been immunised and relatively high rates of TB and hepatitis B. The provision of optimum immunisation, screening, and treatment services to address these issues requires substantial staffing and financial resources. PMID- 11280263 TI - Management of occupational exposure to hepatitis C: survey of United Kingdom practice. AB - We conducted a survey to assess the action taken when health care workers suffered inoculation injuries from patients who might have hepatitis C virus (HCV) infection. A substantial proportion of health care workers may be receiving inadequate follow up and existing guidelines should now be implemented in full and extended to include guidance on treatment. PMID- 11280264 TI - Do all outbreaks really happen on a Friday afternoon? AB - People who work in the control of communicable diseases often claim that incidents such as disease outbreaks tend to happen on Friday afternoons. A survey of outbreaks of foodborne illness did not support this belief, which we suggest is a myth. PMID- 11280265 TI - Effectiveness of the meningococcal vaccination programme for British Armed Forces recruits. AB - After several clusters of meningococcal disease arose among recruits to the British Armed Forces, vaccination against A + C meningococcal disease was introduced for military recruits in 1992. From 1993 onwards, no further clusters of group C infection were reported and incidence of meningococcal disease among trained soldiers fell, but there was no significant reduction in the overall incidence of meningococcal disease in recruits. PMID- 11280267 TI - Timing of immunisation of premature infants on the neonatal unit and after discharge to the community. AB - Premature infants discharged to the community are having their primary immunisations delayed and this delay increases with subsequent doses. The importance and safety of immunising premature infants should be stressed to parents, health visitors, and general practitioners. PMID- 11280266 TI - Case control study of epidural catheter infections in a district general hospital. AB - Ten infections associated with the use of epidural catheters for post-operative pain relief were identified in patients of a district general hospital in 1997 and 1998. A case control study showed that the infections were commoner in the summer months and associated with analgesia infused by syringes rather than pumps. Infection rates and risk factors can be analysed regularly if pain teams maintain a database of epidural catheters inserted, which includes information about infections. PMID- 11280268 TI - Effects of infection control measures on skin of health care workers. AB - A review of patients presenting to a contact dermatitis investigation clinic in 1999 showed that almost half (60/124) of those with occupational hand dermatitis were health care workers. The use of fragrance-free products would be a simple intervention to reduce the prevalence of allergic contact dermatitis without affecting antimicrobial efficacy; other measures are discussed. PMID- 11280269 TI - Science, sense, and nonsense about HIV in Africa. PMID- 11280270 TI - Prevalence of drug injecting among prison inmates. PMID- 11280271 TI - Preventing transmission of bloodborne viruses in prisons. PMID- 11280272 TI - Recombinant protein arrays. PMID- 11280273 TI - Importance of staff job satisfaction in provision of a high quality microbiology service. AB - The practice of quality assurance is a vital aspect of clinical laboratory work. Much effort has been rightly put in to setting up and running external quality assurance programmes, internal quality assurance and control, gaining accreditation with Clinical Pathology Accreditation (UK) Ltd (CPA) and undertaking clinical audit. The contribution that biomedical scientists can make at all levels to quality assurance is very important, because a good pathology service depends on teamwork. The attitude of each individual member of staff can therefore affect the quality the of a laboratory's output. PMID- 11280274 TI - Occupational health strategy for Great Britain. AB - The Health and Safety Commission (HSC) recently launched a long term occupational health strategy for England, Scotland, and Wales. Set out in a document entitled Securing health together and on a dedicated website the ten year strategy aims to reduced occupational ill health by a series of five work programmes. This article looks at the background to the document, its content, and likely impact at work. PMID- 11280275 TI - [Asbestos: contemporary problems of industrial medicine and ecology]. AB - The article covers main steps of research work conducted by Ekaterinbourg Medical Research Center and determines contemporary problems of asbestos. The authors list priority directions of ecologic and hygienic research on the "Asbestos and Health" problem. PMID- 11280276 TI - [Children's health and environmental air pollution with dust containing asbestos]. AB - According to most parameters, children's health in area containing asbestos production is considered as typical one for industrial environment. Dust containing asbestos induces higher incidence of respiratory diseases and immune changes. Frequent respiratory diseases are characteristic for junior age groups of children. PMID- 11280277 TI - [Evaluation of the mineral fibers content of pulmonary tissue in people engaged in extraction and concentration of chrysotile asbestos and in those living near enterprises]. AB - The authors determined concentration of asbestos fibers in lung tissue samples obtained in hospital of Asbestos town in Sverdlovsk region. The samples were taken from 47 individuals who died with various causes. of Workers engaged into extraction and beneficiation of chrysotile asbestos at Bazhenovsky field, into production of chrysotile asbestos goods and those who reside in close proximity to the enterprises demonstrated no differences in general content of fibrous particles and of chrysotile asbestos fibers in lung tissue, if compared to workers of the same enterprises and industrial regions dwellers of Canada (being second, after Russia, for total chrysotile asbestos production). The study proved concentrations of amphibole asbestos fibers to be one order lower than those in Canada. PMID- 11280278 TI - [Clinico-roentgenological and occupational correlations as a method of revealing the dose-effect dependence in the development of chrysotile-asbestos fibrosis]. AB - For understanding up-to-date situation with fibrosis caused by chrysotile asbestos, the authors conducted X-ray and hygienic studies in 2 enterprises extracting and enriching asbestos--"Tuvaasbest" JSC and "Orenbourgasbest" JSC. Authors followed correlation between diffuse pulmonary changes and length of service, dust load and cumulative exposure dose of respirable fibers. The strongest correlation was demonstrated for dust load. Workers engaged into up-to date industry extracting and enriching asbestos are diagnosed as having indistinct and moderate roentgenologic types of fibrosis. Examples of dose-effect dependence prove dust load to be informative and expedient integral parameter for hygienic regulation and evaluation of asbestos-containing dust and industrial aerosols doses. PMID- 11280280 TI - [Occupational expedience of using the artificial substitutes of asbestos (literature review)]. PMID- 11280279 TI - [Health status of workers contacting chrysotile asbestos waste on railway objects]. AB - Asbestos dust associated with other factors induces more hazardous effects, than if acting solely. That is seen especially in women whose morbidity grows 1.5-2.0 times higher than in men of the same occupations. Further studies are recommended for better knowledge asbestos dust influence under provocative action of other hazards. PMID- 11280281 TI - [Clinical features of pulmonary diseases caused by chrysotile asbestos dust]. AB - The authors studied clinical, roentgenologic and functional signs of asbestosis and chronic dust bronchitis in 57 workers engaged into extraction and processing of asbestos. Occupational peripheral lung cancer was revealed in 4 cases. Chronic dust bronchitis appeared to be diagnosed with delay, usually at I-II stages. PMID- 11280282 TI - [Biological effects of chrysotile asbestos dust obtained by dry and moist enriching methods (experimental study)]. PMID- 11280283 TI - [Occupational evaluation of air in workplace area and populated zone at asbestos cardboard production]. PMID- 11280284 TI - [Environmental protection is one of the priorities of "Uralasbest" joint stock company]. PMID- 11280286 TI - [Ecological and occupational evaluation of chrysotile-asbestos fibers emitted during construction and exploitation by roof materials made of asbestos cement]. PMID- 11280285 TI - [Preliminary and periodic medical examinations of asbestos production workers]. AB - The article covers general principles of health care for workers in various industries associated with contact to dust containing asbestos. The authors present main requirements of legal papers basing concepts that determine role of employers, trade unions, workers and medical officers in creation and maintenance of safe work conditions, in establishment of prophylactic, therapeutic and rehabilitation measures. PMID- 11280287 TI - Proceedings of the 1st Conference on the EU Thematic Network Immunology and Ageing in Europe (ImAginE). Tubingen, Germany, 14-18 April 2000. PMID- 11280288 TI - Trial experience and problems of parental recollection of consent. PMID- 11280289 TI - Birth events and cerebral palsy: facts were not presented clearly. PMID- 11280290 TI - Reducing speed limit to 20 mph in urban areas. Evidence based principles should be applied to non-health sector interventions. PMID- 11280291 TI - Reducing speed limit to 20 mph in urban areas. Long term sequelae of road traffic accidents must not be underestimated. PMID- 11280292 TI - Reducing speed limit to 20 mph in urban areas. Health professionals should ensure that local authorities reduce speed limits. PMID- 11280293 TI - Reducing speed limit to 20 mph in urban areas. Both advisory and mandatory speed limits are being introduced in Edinburgh. PMID- 11280294 TI - Email health support service is already operating in Africa. PMID- 11280295 TI - Doesn't everyone know that purpose of advertising is to promote products. PMID- 11280296 TI - Support from pharmacies an help people stop smoking. PMID- 11280298 TI - Emergency medicine and bedside tests. PMID- 11280297 TI - Application of 2-octyl-cyanoacrylate controls persistent ascites fluid leak. PMID- 11280299 TI - Proceedings of the 6th European Society for Organ Transplantation Congress. Rodos, October 25-28, 1993. PMID- 11280301 TI - Cardiac hyperacute rejection: a new look or a skewed look at an old problem? PMID- 11280300 TI - Single-lumen subcutaneous ports inserted by interventional radiologists in patients undergoing chemotherapy: incidence of infection and outcome of attempted catheter salvage. AB - BACKGROUND: Subcutaneous ports are commonly used for vascular access in patients with cancer undergoing chemotherapy. OBJECTIVES: To determine the incidence of catheter-related infection and to assess the efficacy of catheter salvage in subcutaneous ports. METHODS: We retrospectively reviewed 300 subcutaneous single lumen chest ports inserted by interventional radiologists in 294 patients between December 1, 1995, and November 15, 1997, at the Cleveland Clinic Foundation, Cleveland, Ohio. The number of days that the catheter remained in situ, infection rate, treatment, and outcome of infection were determined. RESULTS: Two hundred ninety-four patients had a total of 79 748 catheter-days. Vascular access for chemotherapy was the indication for 95% of the subcutaneous ports placed. Seventeen catheters (5.7%) developed 20 episodes of noninfectious complications resulting in the removal of 6 ports. Seventeen patients (5.7%) developed catheter related infections (2.1/10 000 catheter-days) including 10 episodes of catheter related bacteremia (1.2/10 000 catheter-days). The most common organism isolated was Staphylococcus aureus. A total of 15 of the 17 infected catheters were removed. Salvage was attempted in 6 patients in whom 4 catheters were eventually removed due to recurrent bacteremia (2 patients) and persistent local infection (2 patients). One of the 10 patients with catheter-related bacteremia developed septic arthritis. There were no complications associated with attempted catheter salvage. CONCLUSIONS: Subcutaneous single-lumen ports inserted by interventional radiologists in patients undergoing chemotherapy have low complication rates but infections remain the leading cause of catheter loss. Antibiotic therapy without catheter removal is unlikely to eradicate catheter-related bacteremia. PMID- 11280302 TI - Vaccines and worldwide utilization. PMID- 11280303 TI - Tuberculosis-still the #1 killer in infectious diseases. PMID- 11280305 TI - Cardiovascular Radiation Therapy IV Symposium. February 16-18, 2000. Washington, DC, USA. Abstracts. PMID- 11280304 TI - ECR 2001. Proceedings of the 13th European Congress of Radiology. March 2-6, 2001. Vienna, Austria. PMID- 11280306 TI - Transcatheter Cardiovascular Therapeutics (TCT-11). September 22-26, 1999. Washington, DC, USA. Abstracts. PMID- 11280308 TI - Sensory processing of the aquatic environment. Proceedings of a symposium. 21-27 March 1999. Heron Island, Australia. PMID- 11280309 TI - From the Centers for Disease Control and Prevention. Evaluation of child sexual abuse prevention program--Vermont, 1995-1997. PMID- 11280307 TI - Neural mechanisms involved in the delay of gastric emptying and gastrointestinal transit of liquid after thoracic spinal cord transection in awake rats. AB - Spinal cord transection (SCT) delays gastric emptying (GE), and intestinal and gastrointestinal (GI) transit of liquid in awake rats. This study evaluates the neural mechanisms involved in this phenomenon. Male Wistar rats (N = 147) were fasted for 16 h and had the left jugular vein cannulated followed by laminectomy or laminectomy + complete SCT between T4 and T5 vertebrae. The next day, a test meal (1.5 ml of a phenol red solution, 0.5 mg/ml in 5% glucose) was administered by gavage feeding and 10 min later cervical dislocation was performed. Dye recovery in the stomach, and proximal, mid and distal small intestine was determined by spectrophotometry. SCT inhibited GE and GI transit since it increased gastric recovery by 71.3% and decreased mid small intestine recovery by 100% (P < 0.05). Subdiaphragmatic vagotomy, celiac ganglionectomy + section of the splanchnic nerves, i.v. hexamethonium (20 mg/kg) or yohimbine (3 mg/kg) prevented the development of the SCT effect on GE and GI transit. Pretreatment with i.v. naloxone (2 mg/kg), L-NAME (3 mg/kg) or propranolol (2 mg/kg) was ineffective. Bilateral adrenalectomy or guanethidine (10 mg/kg) increased the magnitude of the GE inhibition, while i.v. prazosin (1 mg/kg) or atropine (0.5 mg/kg) decreased the magnitude but did not abolish the GE inhibition. In summary, the inhibition of GI motility observed 1 day after thoracic SCT in awake rats seems to involve vagal and possibly splanchnic pathways. PMID- 11280310 TI - From the Centers for Disease Control and Prevention. Circulation of a type 2 vaccine-derived poliovirus--Egypt, 1982-1993. PMID- 11280311 TI - From the Centers for Disease Control and Prevention. Injection practices among nurses--Valcea, Romania, 1998. PMID- 11280312 TI - JAMA patient page. Heart emergencies caused by irregular heartbeat. PMID- 11280313 TI - Evidence for the role of sunlight exposure in the etiology of choroidal melanoma. PMID- 11280314 TI - Freiburg oncologist found guilty of scientific misconduct. PMID- 11280315 TI - Conflict-of-interest policies. PMID- 11280316 TI - Conflict-of-interest policies. PMID- 11280317 TI - Conflict-of-interest policies. PMID- 11280318 TI - To protect those who serve. PMID- 11280319 TI - Intrathecal methylprednisolone for postherpetic neuralgia. PMID- 11280320 TI - Intrathecal methylprednisolone for postherpetic neuralgia. PMID- 11280321 TI - Intrathecal methylprednisolone for postherpetic neuralgia. PMID- 11280322 TI - Intrathecal methylprednisolone for postherpetic neuralgia. PMID- 11280323 TI - Intrathecal methylprednisolone for postherpetic neuralgia. PMID- 11280324 TI - Intrathecal methylprednisolone for postherpetic neuralgia. PMID- 11280326 TI - The effect of fecal occult-blood screening on the incidence of colorectal cancer. PMID- 11280325 TI - The effect of fecal occult-blood screening on the incidence of colorectal cancer. PMID- 11280327 TI - Increased incidence of perforated appendixes in Hmong children in California. PMID- 11280328 TI - From the Centers for Disease Control and Prevention. Primary and secondary syphilis--United States, 1999. PMID- 11280329 TI - From the Centers for Disease Control and Prevention. Outbreak of syphilis among men who have sex with men--southern California, 2000. PMID- 11280330 TI - From the Centers for Disease Control and Prevention. Mortality from coronary heart disease and acute myocardial infarction--United States, 1998. PMID- 11280331 TI - JAMA patient page. Treating depression with electroconvulsive therapy. PMID- 11280332 TI - From the Centers for Disease Control and Prevention. Blood and hair mercury levels in young children and women of childbearing age--United States, 1999. PMID- 11280333 TI - From the Centers for Disease Control and Prevention. Risk for meningococcal disease associated with the Hajj 2001. PMID- 11280334 TI - From the Centers for Disease Control and Prevention. Outbreak of poliomyelitis- Dominican Republic and Haiti, 2000-2001. PMID- 11280335 TI - JAMA patient page. Benefits of physical activity for the heart. PMID- 11280336 TI - Universal cut-off BMI points for obesity are not appropriate. PMID- 11280337 TI - An olive oil-rich diet reduces scavenger receptor mRNA in murine macrophages. AB - During atherogenesis, a pathological accumulation of lipids occurs within aortic intimal macrophages through uptake of oxidised LDL via scavenger receptors. Here we investigated whether some of the anti-atherosclerotic effects ascribed to an olive oil rich-diet are mediated through effects on macrophage scavenger receptors (MSR). Male C57 Bl6 mice aged 6 weeks were fed for 12 weeks on a low fat diet (containing 25 g corn oil/kg) or on high-fat diets containing 200 g coconut oil, olive oil or safflower oil/kg. Thioglycollate-elicited peritoneal macrophages were analysed for fatty acid composition by GC and the levels of mRNA coding for three MSR (MSRA type I, MSRA type II and CD36) were measured by reverse-transcription polymerase chain reaction. Feeding mice diets enriched with different fats resulted in significant differences in the fatty acid profile of macrophages, which reflected the fatty acid compositions of the diets. These differences were accompanied by a lower level of mRNA for MSRA type I, MSRA type II and CD36 in macrophages from mice fed an olive-oil-enriched diet compared with the mice fed on the low-fat diet. These data suggest that part of the protective effect of olive oil against atherosclerosis might be via reducing macrophage uptake of oxidised LDL. Whether this effect is due to the downregulation of gene transcription directly by unsaturated fatty acids or is the result of the effect of monounsaturated fatty acids or other components of olive oil on LDL composition and oxidation remains to be ascertained. PMID- 11280338 TI - A cognitive neuroscience account of posttraumatic stress disorder and its treatment. AB - Recent research in the areas of animal conditioning, the neural systems underlying emotion and memory, and the effect of fear on these systems is reviewed. This evidence points to an important distinction between hippocampally dependent and non-hippocampally-dependent forms of memory that are differentially affected by extreme stress. The cognitive science perspective is related to a recent model of posttraumatic stress disorder, dual representation theory, that also posits separate memory systems underlying vivid reexperiencing versus ordinary autobiographical memories of trauma. This view is compared with other accounts in the literature of traumatic memory processes in PTSD, and the contrasting implications for therapy are discussed. PMID- 11280339 TI - The relationship between involuntary pelvic floor muscle activity, muscle awareness and experienced threat in women with and without vaginismus. AB - This study assessed the relationship between involuntary pelvic floor muscle activity, muscle awareness and experienced threat in women with and without vaginismus. Information about this relationship may help understand the mechanism of vaginismus. Twenty-two women with vaginismus and seven control women participated in the study. Women were exposed to four emotion-inducing film excerpts. Vaginal electromyography was recorded. Experienced threat was continuously monitored with the use of a lever. Women responded with increased pelvic floor muscle activity to the threatening and sexually-threatening film excerpt. No changes occurred during the neutral and erotic excerpt. The subjective experienced threat as indicated with the lever showed the same response pattern. However, awareness of changes in muscle activity showed a slightly different pattern. Individual data were inspected. In general, agreement was found between recorded changes in muscle activity and experienced threat. The results of the erotic excerpt showed that awareness of changes in muscle activity is not only determined by information from the pelvic floor muscles, but also by other factors like situational information and the expectations of the women. The data support the idea of a general defense reaction as a mechanism of involuntary pelvic floor muscle activity. PMID- 11280340 TI - Correlates of autobiographical memory specificity: the role of depression, anxiety and childhood trauma. AB - The present study examined the role of childhood trauma, major depressive disorder (MDD), and anxiety disorder (AD) in overgeneral autobiographical memory. Ninety-three outpatients and 24 healthy controls completed a childhood trauma questionnaire and an autobiographical memory test (AMT). Results showed that MDD diagnosis rather than trauma history predicted AMT-performance. Memory specificity was not related to AD diagnosis, recovered MDD, or self-rated depression severity. The present findings cast doubts on theories that emphasize the role of childhood trauma in overgeneral autobiographical memory. PMID- 11280341 TI - The treatment of hypochondriasis: exposure plus response prevention vs cognitive therapy. AB - In this study (1) exposure in vivo plus response prevention, (2) cognitive therapy and (3) a waiting-list control condition were compared on their efficacy on the treatment of hypochondriasis. Seventy-eight patients with a DSM-IV diagnosis of hypochondriasis were randomly assigned to one of these conditions. Patients in both active treatment conditions improved significantly on all the measures, whereas the patients in the waiting-list control condition did not improve. The improvements were clinically significant. Exposure in vivo plus response prevention and cognitive therapy were equally effective. The improvements were maintained at the 7 months follow up. PMID- 11280342 TI - Causal modeling of relations among learning history, anxiety sensitivity, and panic attacks. AB - We used structural equation modeling (SEM) to test the hypothesis that childhood instrumental and vicarious learning experiences influence frequency of panic attacks in young adulthood both directly, and indirectly through their effects on anxiety sensitivity (AS). A total of 478 university students participated in a retrospective assessment of their childhood learning experiences for arousal reactive sensations (e.g., nausea, racing heart, shortness of breath, dizziness) and arousal-non-reactive sensations (i.e., colds, aches and pains, and rashes). SEM revealed that learning history for arousal-reactive somatic symptoms directly influenced both AS levels and panic frequency; AS directly influenced panic frequency; and learning history for arousal-non-reactive symptoms directly influenced AS but did not directly influence panic frequency. These results are consistent with the findings of previous retrospective studies on the learning history origins of AS and panic attacks, and provide the first empirical evidence of a partial mediation effect of AS in explaining the relation between childhood learning experiences and panic attacks in young adulthood. Implications for understanding the etiology of panic disorder are discussed. PMID- 11280343 TI - The concealment of obsessions. AB - Patients' deliberate concealment from others of the content and frequency of their obsessions is a common and important aspect of obsessive-compulsive disorder (OCD). It is an overlooked manifestation of the safety behaviour that is believed to sustain OCD (e.g., neutralizing, thought suppression, avoidance behaviour, concealment). The phenomenon of concealment is understandable in terms of the cognitive theory of obsessions which states that obsessions are caused when the person attaches catastrophic personal significance to their unwanted intrusive thoughts. It is suggested that the selected, planned, suitable disclosure of obsessions can be therapeutic--presumably because it exposes the patient to alternative interpretations of the significance of the unwanted thoughts. PMID- 11280344 TI - The effect of UCS inflation and deflation procedures on 'fear' conditioning. AB - Davey (1992: Classical conditioning and the acquisition of human fears and phobias: a review and synthesis of the literature. Advances in Behaviour Research and Therapy, 14, 29-66) hypothesized that subjective revaluation of an unconditioned stimulus (UCS) would determine the strength of the autonomic conditioned response (CR) in the fear conditioning paradigm. The purpose of the present study was to examine the effect of UCS aversiveness on the CR strength in the fear conditioning paradigm. The UCS aversiveness was controlled by the UCS intensity; that is, the UCS intensity was increased for the inflation group or decreased for the deflation group. Thirty subjects were randomly assigned to the inflation or the deflation group, and they participated under both experimental and control conditions. All subjects went through the pretest, the acquisition of classical conditioning, the UCS intensity operation, and the test sessions. The indices of the CR were skin conductance responses (SCRs) and a subjective aversion to the conditioned stimulus (CS). The main results were as follows. (1) The CR strength measured by SCR was increased by the UCS inflation and decreased by the UCS deflation. (2) The subjective aversiveness of the CS was not sensitive to both manipulations of UCS intensity. These results suggested that the autonomic CR strength might be influenced by the subjective revaluation of UCS, as Davey (1992) described. The result from the test of the subjective aversiveness of the CS, however, could not support Davey's model. The difference between expressions of the SCR and the subjective aversiveness of the CS might be caused by different learning systems. PMID- 11280345 TI - Assessment of anxiety sensitivity in young American Indians and Alaska Natives. AB - In the present study, the Anxiety Sensitivity Index [ASI; Behav. Res. Ther. 24 (1986) 1] was administered to 282 American Indian and Alaska Native college students in a preliminary effort to: (a) evaluate the factor structure and internal consistency of the ASI in a sample of Native Americans; (b) examine whether this group would report greater levels of anxiety sensitivity and gender and age-matched college students from the majority (Caucasian) culture lesser such levels; and (c) explore whether gender differences in anxiety sensitivity dimensions varied by cultural group (Native American vs. Caucasian). Consistent with existing research, results of this investigation indicated that, among Native peoples, the ASI and its subscales had high levels of internal consistency, and a factor structure consisting of three lower-order factors (i.e. Physical, Psychological, and Social Concerns) that all loaded on a single higher order (global Anxiety Sensitivity) factor. We also found that these Native American college students reported significantly greater overall ASI scores as well as greater levels of Psychological and Social Concerns relative to counterparts from the majority (Caucasian) culture. There were no significant differences detected for ASI physical threat concerns. In regard to gender, we found significant differences between males and females in terms of total and Physical Threat ASI scores, with females reporting greater levels, and males lesser levels, of overall anxiety sensitivity and greater fear of physical sensations; no significant differences emerged between genders for the ASI Psychological and Social Concerns dimensions. These gender differences did not vary by cultural group, indicating they were evident among Caucasian and Native Americans alike. We discuss the results of this investigation in relation to the assessment of anxiety sensitivity in American Indians and Alaska Natives, and offer directions for future research with the ASI in Native peoples. PMID- 11280346 TI - The unreliable change of reliable change indices. AB - The classic method for assessment of reliable change, in 1991 re-introduced as Jacobson's RC, can be characterized as a confidence interval method. In recent years, several RC indices have been proposed using Kelley's (1947) (Kelley, T. L. (1947). Fundamentals of statistics. Cambridge: Harvard University Press) formula for estimating true change. In these proposals, interval estimation and confidence intervals are mixed up, which leads to unjustified probability statements. When Kelley's estimate is correctly expanded into a normal distributed statistic, the classic approach reveals itself as a large sample approximation of a properly constructed RCI based on Kelley's formula. Researchers should continue using the classic approach for the determination of reliable change. PMID- 11280347 TI - Fludarabine in the treatment of an active phase of a familial haemophagocytic lymphohistiocytosis. PMID- 11280348 TI - Visceral leishmaniasis: also beware of these deceptive microbes in non-endemic countries! PMID- 11280350 TI - Intestinal inflammation in cystic fibrosis: an alternative hypothesis. PMID- 11280349 TI - Intestinal inflammation in cystic fibrosis. PMID- 11280352 TI - Lumbar puncture should not be performed in meningococcal disease. PMID- 11280353 TI - Prophylaxis for respiratory syncytial virus infection: missing the target. PMID- 11280354 TI - Hajj and risk of blood borne infections. PMID- 11280355 TI - Bone graft substitutes: a comparative qualitative histologic review of current osteoconductive grafting materials. AB - This paper investigated the osteogenic potential of 6 osteoconductive grafting materials derived from human, bovine, and synthetic sources: HTR, BOP, Biogran, Laddec, Dembone, and Osteograf. Twenty-eight New Zealand rabbits were used in this study. The active group consisted of 24 animals and the control group consisted of 4 animals. The median condyle of each tibia was drilled with a 5-mm diameter bur to form 8 mm-deep cavities. A control group included 8 osseous cavities, with 1 hole in each tibia. These cavities were washed and left unfilled. In the active group, each grafting material filled 8 osseous cavities in 8 tibiae of different animals. Half of the active and control osseous cavities were investigated with decalcified hematoxylin and eosin-stained sections. The other half were studied with scanning electron microscopy. It was concluded that Laddec bovine bone granules possessed the best potential for an osteoconductive grafting material, followed by the bioglass crystals of Biogran and the hydroxyapatite particles of Osteograf, respectively. The least potential for rapid bone formation was demonstrated by the copolymers of HTR and BOP, and Dembone allograft bone particles did not reveal active bone healing. PMID- 11280357 TI - When contemplating treatment involving endosseous implants, what clinical and laboratory factors most significantly affect your choice of an implant system? PMID- 11280356 TI - Oral rehabilitation with osseointegrated implants in a patient with oromandibular dystonia with blepharospasm (Brueghel's syndrome): a patient history. AB - Oromandibular dystonia with blepharospasm (also known as Brueghel's syndrome, Meige's syndrome, or idiopathic orofacial dystonia) is characterized by intense and involuntary spasms of the orofacial muscles, with a frequent loss of teeth and occlusal alterations that worsen the dystonic manifestations and cause mucosal lesions that can lead to complete edentulism. The history of a patient with oromandibular dystonia who was rehabilitated with mandibular overdentures supported by endosteal implants is presented. Oral rehabilitation with implant supported overdentures improved the situation, despite serious problems with instability. Mandibular overdentures supported by endosteal implants were satisfactorily used to re-establish occlusion, ensuring prosthetic stability and improving the dynamics of the masticatory muscles. PMID- 11280358 TI - Dental students and dental implants: what's the right combination? PMID- 11280359 TI - Maxillary sinus augmentation with the xenograft Bio-Oss and autogenous intraoral bone for qualitative improvement of the implant site: a histologic and histomorphometric clinical study in humans. AB - The aim of the present clinical study was to determine, through histologic and histomorphometric investigations of human bone specimens, whether the addition of autogenous bone to the bone substitute material Bio-Oss can produce a high quality implant site. To improve vertical bone height, 13 sinus floor elevations were carried out in a total of 12 patients. Augmentation of the maxillary sinus floor was completed using a mixture of Bio-Oss and bone harvested intraorally from the mandibular symphysis, the retromolar space, or the tuberosity region. Following an average of 7.1 months of healing, 36 Branemark System implants were placed. During this surgical intervention, 23 cylinder-shaped bone biopsies were taken from the augmented maxillary region using trephine burs. Histologic analysis of the bone biopsies revealed that the Bio-Oss granulate was well integrated into the newly formed bone; 33.1% (+/- 12.4%) of the substitute material surface was in direct contact with bone. Histomorphometric analysis of the samples revealed an average percentage proportion of bone of 18.9% (+/- 6.4%). The bovine substitute material and soft tissue occupied, respectively, 29.6% (+/- 8.9%) and 51.5% (+/- 9.4%) of the measured surface. When the implants were uncovered after an average healing phase of 6 months, all 36 implants had become osseointegrated. The combination of osteoconductive Bio-Oss and osteoinductive autogenous bone thus proved to be a material suitable for application in sinus floor augmentation. PMID- 11280360 TI - The implant-supported telescopic prosthesis: a biomechanical analysis. AB - This in vitro project investigated load transfer through screw-retained telescopic prostheses. Three Branemark System implants incorporating strain gauges were embedded in an aluminum block. Telescopic prostheses that included 1 mesial and 1 distal cantilever were fabricated over 1 central EsthetiCone and 2 Ti-Adapt abutments. The buffering capacity of the cement in a combined screw retained/cemented prosthesis was studied. The degree of misfit of the prostheses could be adjusted by applying shims of various thicknesses under the EsthetiCone. Load distribution was measured while a 50-N load was applied in turn over each implant and each cantilever. The results showed that tightening the central prosthetic screw widened the load distribution. The cement accommodated misfits between the layers of the telescope, significantly reducing bending moments on some supporting implants. The system exhibited a degree of tolerance to misfit and can provide a versatile prosthodontic option. PMID- 11280361 TI - Vertical distraction osteogenesis of edentulous ridges for improvement of oral implant positioning: a clinical report of preliminary results. AB - This study examined the opportunities offered by intraoral distraction osteogenesis to vertically elongate insufficient alveolar ridges and thereby improve local anatomy for ideal implant placement. Eight patients presenting with vertically deficient edentulous ridges were treated by means of the distraction osteogenesis principle with an intraoral alveolar distractor. Two to 3 months after consolidation of the distracted segments, 26 implants were placed in the distracted areas. Four to 6 months later, abutments were connected and prosthetic loading of the implants was started. The mean follow-up after initial prosthetic loading was 14 months. In all patients, the desired bone gain was reached at the end of distraction (mean vertical bone gain of 8.5 mm). Probing depth, Bleeding Index, and Plaque Index around implants were evaluated, and Periotest values were also calculated. The cumulative success rate of implants was 100%. Radiographic examinations 12 months after functional loading of implants showed a significant increase in the density of the newly generated bone in the distracted areas. This technique seems to be reliable, and the regenerated bone has withstood the functional demands of implant loading. Success rates of implants, periodontal indices of peri-implant soft tissues, and Periotest values were consistent with those reported in the literature regarding implants placed in native bone. PMID- 11280362 TI - The self-tapping and ICE 3i implants: a prospective 3-year multicenter evaluation. AB - This multicenter prospective clinical evaluation was undertaken to determine the therapeutic success and marginal bone level stability of 3i's self-tapping and ICE implants after 3 years of prosthetic loading. Between July 1995 and June 1996, 189 completely or partially edentulous patients were treated with 614 machined-surface screw-type commercially pure titanium implants (self-tapping or ICE). Two hundred seventy-seven self-tapping implants were placed in 85 patients (average age of 56 years), and 337 ICE implants were placed in 104 patients (average age of 61 years). A total of 360 implants (58.6%) were placed in posterior segments. Easier placement was reported with the ICE implant in normal or dense bone. For the self-tapping implants, survival rates of 92.9% and 91.6% were noted after 1 and 3 years of prosthetic loading, respectively. Survival rates of 95.4% and 93.8% were obtained with the ICE implant for the same periods. Late failures (after loading) were more common than early failures (before loading) for both types of implants. The marginal bone level of 238 self-tapping implants (85.9%) and of 307 ICE implants (91%) was radiographically evaluated at 3 years. Marginal bone level was at the first thread for 95.1% of implants. A loss of marginal bone level of 2 to 4 threads was noted for 4.9% of the evaluated implants. No implant showed bone loss greater than the fourth thread. Overall survival rates of 94.3% and 92.9% were obtained after 1 and 3 years of prosthetic loading, respectively, for 596 and 588 implants. PMID- 11280363 TI - Influence of flap design on peri-implant interproximal crestal bone loss around single-tooth implants. AB - The anterior maxilla represents a therapeutic challenge for single-tooth replacement with implants. The surgical trauma delivered to soft and hard tissues during implant placement can influence the future esthetic result. The clinician should use surgical techniques that prevent esthetic complications, such as increased crown length or loss of interdental papillae, without compromising osseointegration. This prospective study investigated the interproximal crestal bone loss occurring after placement of single-tooth implants using 2 different flap designs: a widely mobilized flap design that included papillae, and a limited flap design that protected papillae. The interproximal crestal bone loss was of practical importance and statistically significantly less following the use of a limited flap design versus the widely mobilized flap procedure. PMID- 11280364 TI - Computed tomographic diagnosis and localization of bone canals in the mandibular interforaminal region for prevention of bleeding complications during implant surgery. AB - In this study, computed tomograms (CTs) of 70 patients were examined for visible vascular canals in the mandible as well as for their localization, incidence, diameter, and content. All patients examined showed at least 1 lingual perforating bone canal in the mandible. Since such vascular canals are encountered regularly, routine CT examination is recommended prior to implant surgery to help avoid severe bleeding complications during the placement of implants in the interforaminal region. PMID- 11280365 TI - Rehabilitation of patients with severely resorbed maxillae by means of implants with or without bone grafts: a 3- to 5-year follow-up clinical report. AB - Forty three patients with severely resorbed maxillae who had been referred for implant treatment were assigned to 1 of 3 treatment options: bone grafting and implant placement (graft group), modified implant placement with no bone grafting (trial group), or optimized complete dentures (no-implant group). Sixteen, 20, and 7 patients, respectively, were assigned to the 3 groups. The patients have been examined annually, and at the time of this report they had been followed for 3 to 5 years after treatment. At the 1-year follow-up, 10% (22 of 221) of the implants had been lost, and at the 2-year follow-up, 18% of the implants had been lost (40 of 221; 25% in the graft and 13% in the trial group); after that time, no further losses occurred. Life table analysis showed cumulative success rates of 82% in the graft group and 96% in the trial group after 1 year, and 74% and 87%, respectively, at the final examination after 3 to 5 years. The failure rate was higher in smokers than in non-smokers. A substantial reduction of the grafted bone, especially of onlay grafts, occurred early after grafting surgery in many patients. Mean marginal peri-implant bone loss was 0.6 mm during the period from prosthesis connection to the 1-year follow-up, and from the 1-year to the 3-year follow-up, average peri-implant bone loss was 0.3 mm in the graft group and 0.5 mm in the trial group. The results corroborated previous findings that patients with severely resorbed maxillae have an increased risk of implant failure in comparison to patients with good bone quantity and quality. However, in this investigation, practically all implant losses occurred during the first 2 years, whereupon a steady state seemed to follow for up to 5 years after loading. PMID- 11280366 TI - The radiographic assessment of implant patients: decision-making criteria. AB - Indications for the most frequently used imaging modalities in implant dentistry are proposed based on clinical need and biologic risk for the patient. To calculate the biologic risk, the authors carried out dose measurements. They demonstrated that the risk from a periapical radiograph is 20% of that from a panoramic radiograph. A panoramic radiograph and a series of 4 conventional tomographs of a single-tooth gap in the molar region carry 5% and 13% of the risk from computed tomography of the maxilla, respectively. Panoramic radiography is considered the standard radiographic examination for treatment planning of implant patients, because it imparts a low dose while giving the best radiographic survey. Periapical radiographs are used to elucidate details or to complete the findings obtained from the panoramic radiograph. Other radiographic methods, such as conventional film tomography or computed tomography, are applied only in special circumstances, film tomography being preferred for smaller regions of interest and computed tomography being justified for the complete maxilla or mandible when methods for dose reduction are followed. During follow up, intraoral radiography is considered the standard radiographic examination, particularly for implants in the anterior region of the maxilla or for scientific studies. In patients requiring more than 5 periapical images, panoramic radiography is preferred. PMID- 11280367 TI - Reconstruction of severely resorbed alveolar ridge crests with dental implants using a bovine bone mineral for augmentation. AB - This study reviews the outcome of implant placement in 61 patients after augmentation of severely atrophic alveolar bone with a bovine bone mineral, Bio Oss. Bone augmentation was performed at 4 different sites: alveolar crest width, alveolar crest height, antral cavity, or nasal cavity. After a mean healing time of 11.9 months, 231 implants were placed in Bio-Oss bone. The time of loading of the implants varied between 12 and 113 months. Calculated from the time of implant placement and irrespective of loading time, a survival rate of 80.5% for the individual implants was estimated. In most patients (73%), Bio-Oss was mixed with autogenous bone from the chin. However, the results indicated that autogenous bone may be excluded from the Bio-Oss graft. PMID- 11280369 TI - Resistance to Marek's disease herpesvirus-induced lymphoma is multiphasic and dependent on host genotype. AB - Genotype-dependent differences in Marek's disease (MD) susceptibility were identified using 14-day-old line N and 6(1) (resistant) and 151 and 7(2) (susceptible) inbred chickens infected with HPRS-16 MD virus (MDV). All line 72 chickens developed progressive MD. Line 15I had fluctuating MD-specific clinical signs and individuals recovered. A novel histologic scoring system enabled indices to be calculated for lymphocyte infiltration into nonlymphoid organs. All genotypes had increased mean lesion scores (MLSs) and mean total lesion scores after MDV infection. These differed quantitatively and qualitatively between the genotypes. Lines 6(1) and 7(2) had a similar MLS distribution in the cytolytic phase, although scores were greater in line 7(2). At the time lymphomas were visible in line 7(2), histologic lesions in line 6(1) were regressing. AV37+ cells were present in similar numbers in all genotypes in the cytolytic phase, suggesting that neoplastically transformed cells were present in all genotypes regardless of MD susceptibility. After the cytolytic phase, AV37+ cell numbers increased in lines 7(2) and 15I but decreased in lines 6(1) and N. In the cytolytic and latent phases, in all genotypes, most infiltrating cells were CD4+. After this time, line 7(2) and 15I lesions increased in size and most cells were CD4+; line 6(1) and N lesions decreased in size and most cells were CD8+. In all genotypes, AV37 immunostaining was weak in lesions with many CD8+ cells, suggesting that AV37 antigen expression or AV37+ cells were controlled by CD8+ cells. The rank order, determined by clinical signs and pathology, for MD susceptibility (highest to lowest) was 7(2) > 15I > 6(1) > N. PMID- 11280368 TI - The symphyseal single-tooth implant for anchorage of a mandibular complete denture in geriatric patients: a clinical report. AB - Little information exists to define the minimum number of implants required for sufficient anchorage of mandibular overdentures. To date, 2 implants placed in the interforaminal region have been considered the minimum. The aim of this study was to examine whether a single symphyseal implant would suffice for adequate anchorage of a mandibular complete denture in elderly patients (octogenarians), and whether this surgically, prosthetically, and financially simple concept would also satisfy patients needing replacement of the mandibular dentition. Nine patients with a mean age of 82.2 years underwent placement of a single symphyseal endosseous implant and anchorage of a complete denture using a ball attachment. Standardized recall examinations, including patient response and inspections of the peri-implant soft tissue and bone conditions, were carried out at 3- to 6 month intervals for a period of 1 1/2 years. It was found that anchorage with a single implant led to both a significant improvement in patients' subjective satisfaction (P < .01) and a significant reduction in reported symptoms (P < .01). During the observation phase, pocket depth and bone resorption initially increased around implants but stabilized after the sixth month. Denture management (placement and removal) also improved during the recall period (P < .01). The results of this study indicate that oral rehabilitation by mandibular complete dentures anchored on a single implant can be considered an economical therapeutic alternative to a conventional mandibular complete denture for very old (octogenarian) patients. PMID- 11280370 TI - In vitro evidence for effects of magnesium supplementation on quinolone-treated horse and dog chondrocytes. AB - Quinolones and magnesium deficiency cause similar lesions in joint cartilage of young animals. Chondrocytes cultivated in the presence of quinolones and in Mg free medium show severe alterations in cytoskeleton and decreased ability to adhere to the culture dish. We investigated whether Mg2+ supplementation can prevent quinolone-mediated effects on chondrocytes in vitro. Chondrocytes cultivated in Dulbecco's modified Eagle's medium/HAM's F-12 medium were treated with ciprofloxacin (80 and 160 microg/ml) and enrofloxacin (100 and 150 microg/ml). Mg2+ was added at a concentration of 0.0612 mg/ml (MgCl) and 0.0488 mg/ml (MgSO4) or a triple dose. In addition, cells were cultivated in Mg-free medium and accordingly treated with Mg2+ supplementation. After 5 days in culture, the number of adherent cells per milliliter was determined. The number of chondrocytes in quinolone-treated groups decreased to 12-36% that of the control group within the culture period. With Mg2+ supplementation, the number of attached cells increased to 40-70% that of control cells. The threefold dose of Mg2+ led to better results than did the single dose. Cell proliferation tested by immunohistochemical staining with Ki67 (clone MIB5) decreased from 70% in control groups to 55%, 48%, and 30% in enrofloxacin-treated groups in a concentration dependent manner (50, 100, and 150 microg/ml). Addition of Mg2+ did not increase the rate of cell proliferation. These results suggest that a great part of quinolone-induced damage is due to magnesium complex formation, as Mg2+ supplementation is able to reduce the effects in vitro. However, quinolone effects on cell proliferation seem to be an independent process that is not influenced by magnesium supplementation. PMID- 11280371 TI - Pathobiology of A/chicken/Hong Kong/220/97 (H5N1) avian influenza virus in seven gallinaceous species. AB - Direct bird-to-human transmission, with the production of severe respiratory disease and human mortality, is unique to the Hong Kong-origin H5N1 highly pathogenic avian influenza (HPAI) virus, which was originally isolated from a disease outbreak in chickens. The pathobiology of the A/chicken/Hong Kong/220/97 (H5N1) (HK/220) HPAI virus was investigated in chickens, turkeys, Japanese and Bobwhite quail, guinea fowl, pheasants, and partridges, where it produced 75-100% mortality within 10 days. Depression, mucoid diarrhea, and neurologic dysfunction were common clinical manifestations of disease. Grossly, the most severe and consistent lesions included splenomegaly, pulmonary edema and congestion, and hemorrhages in enteric lymphoid areas, on serosal surfaces, and in skeletal muscle. Histologic lesions were observed in multiple organs and were characterized by exudation, hemorrhage, necrosis, inflammation, or a combination of these features. The lung, heart, brain, spleen, and adrenal glands were the most consistently affected, and viral antigen was most often detected by immunohistochemistry in the parenchyma of these organs. The pathogenesis of infection with the HK/220 HPAI virus in these species was twofold. Early mortality occurring at 1-2 days postinoculation (DPI) corresponded to severe pulmonary edema and congestion and virus localization within the vascular endothelium. Mortality occurring after 2 DPI was related to systemic biochemical imbalance, multiorgan failure, or a combination of these factors. The pathobiologic features were analogous to those experimentally induced with other HPAI viruses in domestic poultry. PMID- 11280372 TI - Histopathology of experimental plague in cats. AB - Formalin-fixed paraffin-embedded archival tissues of seven adult cats of both sexes that died after being experimentally infected with Yersinia pestis were examined light microscopically to characterize the lesions. The cats were exposed in two groups using two routes of infection: ingestion of Y. pestis-infected rodent or a subcutaneous injection of Y. pestis to simulate a flea bite. Immunohistochemistry was performed on tissues from all organ systems from a representative cat from each group to determine the distribution of Y. pestis bacilli during infection. In all seven cats, bubonic plague lesions were seen. The lesions of pneumonic plague were present in two cats. Septicemic plague was confirmed in all seven cats by bacteriologic culture. Aggregations of bacteria were seen in lymphoid tissue in all cats and in lung tissues from the two cats with pneumonic plague. The most consistent histologic finding was necrosuppurative inflammation in the lymph nodes. Invariably, Y. pestis bacteria were present in large numbers at affected sites. Orally infected cats had more numerous lesions in the lymph nodes of the head and neck regions. These experimentally induced cases of feline plague document that cats are unique among carnivores in exhibiting bubonic, pneumonic, and septicemic plague following exposure to Y. pestis. The lesions of the orally infected cats were consistent with those previously described for naturally occurring Y. pestis infections in cats and corroborate the contention that cats most commonly contract plague by eating Y. pestis-infected rodents and not via flea bite. The histopathology of Y. pestis disease in these cats is comparable to that described for human plague. PMID- 11280373 TI - Comparison of gastritis and gastric epithelial proliferation in Helicobacter heilmannii-infected nude and BALB/c mice. AB - Gastric mucosal hypertrophy/nodular hyperplasia occurs as a consequence of Helicobacter infection in mice and humans. The pathogenesis of this hyperplastic response is not understood. To determine the role of host cellular immunity in gastric mucosal hypertrophy/hyperplasia, 6-8-week-old female euthymic BALB/c (n = 30) or NIH athymic nude (n = 40) mice were inoculated with H. heilmannii. Equal numbers of uninoculated mice of each strain served as controls. Mice from each group were euthanatized at 0.5, 6, 12, and 18 months postinoculation (PI). Lymphoplasmacytic gastritis and lymphoid follicle development were less severe in nude mice than in euthymic mice at <6 months PI. The prevalence of gastritis at 0.5, 6, 12, and 18 months PI was 0%, 17%, 67%, and 88%, respectively, in infected nude mice and 33%, 83%, 71%, and 100%, respectively, in infected BALB/c mice. CD4+ T cells in infected nude mice were evident at > or =6 months PI but were less numerous than in infected BALB/c mice at comparable time intervals. However, numbers of CD4+ T cells increased substantially throughout the experiment in infected BALB/c mice. The prevalence of nodular mucosal hyperplasia at 0.5, 6, 12, and 18 months PI was 0%, 0%, 33%, and 20%, respectively, in infected nude mice and 0%, 33%, 80%, and 80%, respectively, in infected BALB/c mice. Nodular hyperplasia occurred in association with the appearance of chronic lymphoplasmacytic inflammation and CD4+ T cells at 12 and 18 months PI in nude mice. H. heilmannii-associated gastritis and mucosal remodeling is reduced in immunodeficient mouse strains lacking normal CD4+ T cell numbers as compared with the response in immunocompetent mice. Additionally, CD4 immunocompetence is an integral aspect of the mucosal hypertrophy/nodular hyperplasia in experimental H. heilmannii-associated disease of mice. PMID- 11280374 TI - Uptake of Mycobacterium avium subsp. paratuberculosis through the distal small intestinal mucosa in goats: an ultrastructural study. AB - Various pathogens gain access to the intestinal wall via specialized cells, the M cells, found among the follicle-associated epithelial cells overlying the domes of the Peyer's patches. The present study was undertaken to examine the uptake of live Mycobacterium avium subsp. paratuberculosis in the distal small intestine of goat kids. Following laparotomy, distal small intestinal segments of five goats were ligated and injected with bacterial suspension. After 1 hour, the intestinal segments were excised and fixed for light and electron microscopic studies. M. a. paratuberculosis organisms were observed by transmission electron microscopy at locations in the intestinal wall, suggesting transcellular transportation through the M cells. The organisms were present both in the cytoplasm of the M cells and in the cytoplasm of intraepithelial leukocytes found in M-cell pockets. Intercellular bacteria between M cells were occasionally seen. Bacteria were not observed in association with the absorptive epithelium. This study indicates that in goat kids, M. a. paratuberculosis enters the intestinal wall primarily through the M cells in the follicle-associated epithelium of the Peyer's patches. PMID- 11280375 TI - Otitis interna is a frequent sequela to Streptococcus suis meningitis in pigs. AB - Twenty-eight histologically confirmed cases of porcine leptomeningitis were examined retrospectively, with focus on the pathology of the inner and middle ear, brain, and vestibulocochlear nerve. Tissues were evaluated by histology and immunohistochemistry for Streptococcus suis serotype 2 antigen, and the bacteriologic results were recorded. Exudative otitis interna was diagnosed in 20/28 pigs (71%). The lesions primarily affected the perilymphatic ducts, with consistent involvement of the scala tympani. Perineuritis of the vestibulocochlear nerve was seen in all but four of the ears affected with otitis interna. Immunohistochemically, S. suis serotype 2 antigen was demonstrated in the leptomeningeal, perineural, and labyrinthine exudates in 11 cases. Otitis media was diagnosed in 10/28 pigs (34%), but evidence of extension to the inner ear was not observed. The findings were highly similar to descriptions of meningogenic labyrinthitis in humans and in laboratory animal models. Otitis interna in pigs can also develop via the meningogenic route and is not always, as generally stated, tympanogenic. PMID- 11280376 TI - Feline vaccine-associated fibrosarcoma: an ultrastructural study of 20 tumors (1996-1999). AB - Twenty feline vaccine-associated sarcomas were examined by transmission electron microscopy. Tumors contained pleomorphic spindle cells, histiocytoid cells, and giant cells. Most tumors contained myofibroblasts, which had morphologic features similar to those of fibroblasts. These cells were further distinguished by subplasmalemmal dense plaques and thin cytoplasmic actin myofilaments organized as elongated bundles concentrated at irregular intervals forming characteristic dense bodies. Intracellular crystalline particulate material was found in 5 of the 20 tumors. Energy dispersive X-ray spectroscopy was used to identify the crystalline material within one tumor as aluminum-based. One tumor from a feline leukemia virus-infected cat contained budding and immature retroviral particles. PMID- 11280377 TI - Cutaneous DNA vaccination against Ebola virus by particle bombardment: histopathology and alteration of CD3-positive dendritic epidermal cells. AB - We analyzed the localization of gold particles, expression of immunogenic protein, and histopathologic changes after vaccinating guinea pigs and mice with a DNA vaccine to the Ebola virus glycoprotein administered by cutaneous particle bombardment. Gold particles were deposited in all layers of the epidermis and in the dermis. Those in the epidermis were lost as the damaged layers sloughed, while those in the dermis were phagocytized by macrophages. Glycoprotein was demonstrated by immunohistochemistry primarily in keratinocytes in the epidermis and hair follicle epithelium and less frequently in dermal macrophages, fibroblasts, sebocytes, and cells that appeared to be Langerhans cells. The number of cells that expressed glycoprotein increased between 4 and 8 hours postvaccination, then decreased to near zero by 48 hours. The vaccine sites were histologically divisible into three zones. The central portion, zone 1, contained the most gold particles in the dermis and epidermis and had extensive tissue damage, including full-thickness epidermal necrosis. Zone 2 contained fewer gold particles in the epidermis and dermis and had less extensive necrosis. The majority of cells in which glycoprotein was expressed were in zone 2. Zone 3 contained gold particles only in the epidermis and had necrosis of only a few scattered cells. Regeneration of the epidermis in damaged areas was evident at 24 hours postvaccination and was essentially complete by day 5 in the mice and day 10 in the guinea pigs. Inflammatory changes were characterized by hemorrhage, edema, and infiltrates of neutrophils initially and by infiltrates of lymphocytes and macrophages at later times. In zone 1, inflammation affected both the epidermis and dermis. Peripherally, inflammation was relatively limited to the epidermis. CD3-positive dendritic epidermal cells were demonstrated in the epidermis and superficial hair follicles of unvaccinated immunocompetent mice and beige mice but not of SCID mice. These cells disappeared from all but the most peripheral portions of the vaccine sites of vaccinated mice within 24 hours. They reappeared slowly, failing to reach numbers comparable with unvaccinated mice by 35 days postvaccination. The epidermis of control guinea pigs also had CD3 positive cells, but they did not have dendrites. These findings should contribute to a better understanding of the mechanisms operating in response to DNA vaccination by particle bombardment. PMID- 11280378 TI - Demonstration of Akabane virus antigen using immunohistochemistry in naturally infected newborn calves. AB - Eight newborn calves showing ataxia were necropsied and examined histologically. Six of seven cerebrospinal fluid samples collected from these animals had neutralizing antibody for Akabane virus (AKV). All examined calves had nonsuppurative encephalomyelitis, localized mainly in the midbrain and spinal cord. Corresponding to the encephalitic lesion, AKV antigen was demonstrated in neuroglial cells in the brain stem and neuronal cells in the ventral horn of the spinal cord. This is the first study to demonstrate AKV antigen by immunohistochemistry in naturally infected newborn calves. PMID- 11280379 TI - Diffuse leptomeningeal malignant histiocytosis in the brain and spinal cord of a Tibetan Terrier. AB - An 8-year-old male Tibetan Terrier showed prolonged astasia, complete paralysis, ticlike signs, and seizure and died 2 months after the onset of symptoms. Histopathologically, there was moderate to severe infiltration of pleomorphic histiocytic mononuclear cells bilaterally in the basiarachnoidal and ventricular areas of the brain. The spinal dura mater, arachnoidal space, and leptomeninges were also affected by infiltrative proliferation of these mononuclear cells. The infiltrating cells had the morphologic characteristics of histiocytes but exhibited moderate pleomorphism and atypia, with abundant mitotic figures. With immunohistochemistry and lectin histochemistry, most of the infiltrating cells were positive for lysozyme and lectin RCA-1 and negative for glial fibrillary acid protein, suggesting that they were of monocytic/histiocytic-origin. Positive proliferating cell nuclear antigen immunostaining demonstrated that most nuclei of the histiocytic cells were in the S phase of the cell cycle, consistent with a proliferating population of cells. Based on these findings, the case was diagnosed as diffuse leptomeningeal malignant histiocytosis. PMID- 11280380 TI - Small intestine large granular lymphoma in a horse. AB - A 12-year-old Appaloosa gelding was referred to the Texas Veterinary Medical Center with a history of chronic diarrhea and weight loss. At necropsy, numerous oval, craterlike ulcers were observed throughout the small intestine. Histologically, these lesions were composed of a neoplastic proliferation of round cells with intracytoplasmic phosphotungstic acid-hematoxylin-positive granules. The tumor cells stained positively for the CD3 antigen and negatively for a B-cell marker. A diagnosis of large granular lymphoma was based on the morphologic and immunohistochemical characteristics of the neoplasm. The postmortem presentation of this case depicted unusual multifocal, ulcerative lymphomatous lesions throughout the small intestine without involvement of the regional lymph nodes. The histologic and ultrastructural morphology of the neoplastic lymphocytes was similar to that in previously reported cases of abdominal equine large granular lymphomas, but in this case the neoplasm was restricted to the small intestine. PMID- 11280381 TI - Multiple epulides in 13 cats. AB - Epulides account for 0-7.8% of tumors in surveys of feline oral neoplasms. A review of the literature revealed no reports of multiple epulides in cats. Multiple, concurrent epulides were diagnosed microscopically in 13 cats. Fibromatous and ossifying epulides were diagnosed in 11 of 13 cats and fibromatous epulides were diagnosed in 2 of 13 cats. Microscopically, these epulides were nonencapsulated, well-vascularized, infiltrative, highly cellular neoplasms that expanded the gingiva and were composed of haphazardly arranged, spindle-shaped to stellate cells amid a dense, collagenous stroma. Osseous foci were a feature in the fibromatous and ossifying epulides. The mitotic rate was low and there was marked hyperplasia of the overlying gingiva with a prominent downgrowth of epithelial cords. These tumors recurred in 8 of 13 cats following surgical excision. While uncommon, multiple epulides in cats have a high incidence of recurrence but do not appear to have metastatic potential. PMID- 11280383 TI - Naturally occurring spirochetes in the colonic mucosa of raccoons (Procyon lotor). AB - The large intestines of 21 raccoons (Procyon lotor; 11 wild caught, 10 laboratory confined) were examined for the presence of intestinal spirochetes. Light microscopy of sections stained with hematoxylin and eosin and Warthin-Starry stain showed the presence of spiral shaped organisms deep within the lumina of intestinal glands at the ileocolonic junction of 16 raccoons (76% prevalence). All laboratory-confined, group-housed raccoons harbored the organisms, but only 6/11 (55% prevalence) live-trapped raccoons were positive for these spirochetes. The organisms were free in the glandular lumina, and there were no microscopic lesions. Two types of spirochetes were identified in the colonic glands: a slender spirochete 10-13 microm in length, 0.3 microm in diameter, and possessing long, thin tapered ends and a larger, regularly waved spiral organism (0.5 microm in diameter). The slender spirochete did not resemble any of the known spirochete genera and failed to grow on medium used to propagate oral treponemes and members of the genus Brachyspira. PMID- 11280382 TI - Septicemia and peritonitis due to Actinobacillus equuli infection in an adult horse. AB - Actinobacillus equuli is a rare cause of peritonitis in adult horses. Septicemia and peritonitis due to A. equuli were diagnosed at necropsy in an 8-year-old Saddlebred mare. The origin of the infection was not known; however, small necrotic colonic mucosal lesions presumed to have been caused by phenylbutazone treatment may have allowed bacterial invasion. A good response to antimicrobial treatment has been documented in the small numbers of previously reported acute cases of peritonitis. Because it is potentially treatable, it is important for pathologists and clinicians to identify horses with A. equuli peritonitis. PMID- 11280384 TI - Immunohistochemical detection of tumor suppressor gene p53 protein in feline injection site-associated sarcomas. AB - Sarcomas associated with injection sites are a rare but important problem in cats. Immunohistochemical detection of p53 protein may correlate to mutation of the p53 tumor suppressor gene, a gene known to be important in oncogenesis. The expression of nuclear p53 protein in 40 feline injection site-assocated sarcomas was examined by immunohistochemical staining. In 42.5% (17/40), tumor cell nuclei were stained darkly; in 20% (8/40), tumor cell nuclei were stained palely; and in 37.5% (15/40), tumor cell nuclei were unstained. Immunohistochemical detection of p53 protein in a proportion of injection site-associated sarcomas suggests that mutation of the p53 gene may play a role in the pathogenesis of these tumors. PMID- 11280385 TI - Aural-pharyngeal polyps associated with Cryptosporidium infection in three iguanas (Iguana iguana). AB - Cryptosporidium spp. infection was associated with aural-pharyngeal polyps in three iguanas (Iguana iguana). All iguanas were presented for masses protruding from the ear canal, and the disease was characterized by a chronic clinical course. The masses consisted of nests of cystic glands surrounded by abundant fibrous connective tissue and lined by hyperplastic cuboidal to pseudostratified columnar epithelium that was moderately to heavily colonized by cryptosporidial organisms. Electron microscopy revealed that the majority of organisms were trophozoites. PMID- 11280386 TI - Gastrointestinal stromal tumors of the equine cecum. AB - Ten cecal tumors were identified during the postmortem examination of seven horse carcasses at slaughter (one horse had three tumors). The multinodular and hemorrhagic tumors ranged from 1 to 10 cm in diameter and consisted of spindle cells arranged in thin, interconnected trabeculae that were often separated by sinuses filled with mucinous fluid, erythrocytes, and siderophages. Spindle cells of all tumors were immunopositive for vimentin, neuron-specific enolase, and c kit protein but lacked reactivity with antibodies to glial fibrillary acidic protein, S100 protein, and desmin. In one tumor, spindle cells diffusely bound antibodies to synaptophysin. Most tumors contained focal reactivity to smooth muscle actin antibodies; one tumor reacted diffusely. Ultrastructurally, tumor cells were connected by desmosome-like structures and exhibited extended cell processes; some contained dense core neurosecretory granules. These equine stromal tumors appeared to share some characteristics with human gastrointestinal stromal tumors. PMID- 11280388 TI - Reporting laws suggest need for abuse standards. PMID- 11280387 TI - BSE: could it happen here? Experts say probably not. PMID- 11280389 TI - Tapping a "tremendous resource"--allied group expertise. PMID- 11280390 TI - Clients can find a pet on the Net. PMID- 11280391 TI - Veterinarians can help ease financial, emotional worries. PMID- 11280392 TI - Opposes restoring "star" to veterinary corps. PMID- 11280393 TI - Disagrees with ECFVG process. PMID- 11280394 TI - Call for professional courtesy. PMID- 11280395 TI - Are we eroding the veterinarian-client relationship? PMID- 11280396 TI - 2000 Report of the AVMA Panel on Euthanasia. PMID- 11280397 TI - Vaccine-associated feline sarcomas. PMID- 11280398 TI - What is your diagnosis? Emphysematous cystitis, pneumaturia and lumbar spondylosis. PMID- 11280399 TI - Evaluation of raw food diets for dogs. PMID- 11280400 TI - Employment, starting salaries, and educational indebtedness of year-2000 graduates of US veterinary medical colleges. PMID- 11280401 TI - Some observations about Internet practice and veterinarians. PMID- 11280403 TI - Primary T-cell lymphoma of the central nervous system in a dog. AB - Primary T-cell lymphoma is a rare form of CNS neoplasia. Diagnosis may be aided by use of cytologic examination of CSF. Primary CNS T-cell lymphoma should be considered in a patient with multiple cranial nerve abnormalities, even if results of imaging studies are considered normal. PMID- 11280402 TI - Internet income: bane or bonanza? PMID- 11280404 TI - Use of pericardial patch graft reconstruction of the right atrium for treatment of hemangiosarcoma in a dog. AB - Atrial mass resection is possible in a limited number of dogs with hemangiosarcoma in which only the right atrial appendage or atrial free wall is involved, A pericardial patch graft can be used to reconstruct the right atrium after resection of large atrial tumors. Tumor-free margins can be obtained by use of this technique. PMID- 11280405 TI - Granulosa cell tumor in a guinea pig. AB - Abdominal distention is a common clinical sign in guinea pigs and may have many causes. Abdominal ultrasonography may be a useful diagnostic tool in differentiation of abdominal disorders in guinea pigs. Ovariohysterectomy is indicated for granulosa cell tumors and cystic rete ovarii in guinea pigs. PMID- 11280406 TI - Clinical features of dilated cardiomyopathy in Great Danes and results of a pedigree analysis: 17 cases (1990-2000). AB - OBJECTIVE: To determine clinical features of dilated cardiomyopathy (DCM) in Great Danes and to determine whether DCM is familial in this breed. DESIGN: Retrospective study. ANIMALS: 17 Great Danes with DCM. PROCEDURE: Medical records of Great Danes in which DCM was diagnosed on the basis of results of echocardiography (fractional shortening < 25%, end-systolic volume index > 30 ml/m2 of body surface area) were reviewed. Pedigrees were obtained for affected animals, as well as for other Great Danes in which DCM had been diagnosed. RESULTS: Dilated cardiomyopathy appeared to be familial and was characterized by ventricular dilatation, congestive heart failure (left-sided or biventricular), and atrial fibrillation. Pedigree analysis suggested that DCM was inherited as an X-linked recessive trait, but the mode of inheritance could not be definitively identified. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that DCM may be an X-linked recessive trait in Great Danes. Thus, dogs with DCM probably should not be used for breeding, and female offspring of affected dogs should be used cautiously. Male offspring of affected females are at an increased risk of developing DCM and should be evaluated periodically for early signs of disease. Results of pedigree analysis were preliminary and should be used only as a guide for counseling breeders, rather than as a basis for making breeding decisions. PMID- 11280407 TI - Nonhealing corneal ulcers in cats: 29 cases (1991-1999). AB - OBJECTIVE: To compare mean healing times after debridement, debridement with grid keratotomy, and superficial keratectomy in cats with nonhealing corneal ulcers. DESIGN: Retrospective study. ANIMALS: 29 cats with 36 nonhealing corneal ulcers. PROCEDURE: Medical records of cats with nonhealing corneal ulcers were reviewed. Signalment, duration of clinical signs, ophthalmic abnormalities, and response to various treatment protocols were recorded. RESULTS: Mean age of affected cats was 7 years, 8 months. Affected breeds included domestic shorthair (17 cats), Persian (9), Himalayan (2), and Siamese (1). Clinical signs were evident for approximately 2 weeks prior to referral. Both eyes were affected in 4 cats. Mean healing time of ulcers treated with superficial debridement was 30 days. Mean healing time of ulcers treated with superficial debridement and grid keratotomy was 42 days. Superficial keratectomy was performed on 2 eyes and resulted in a healing time of 2 weeks. Formation of a corneal sequestrum was evident in 2 of 21 eyes treated with superficial debridement. Formation of a corneal sequestrum was evident in 4 of 13 eyes treated with superficial debridement and grid keratotomy. CONCLUSIONS AND CLINICAL RELEVANCE: Brachycephalic cats appear to be predisposed to developing nonhealing corneal ulcers. The combination of superficial debridement and grid keratotomy did not decrease mean healing time of nonhealing ulcers, compared with superficial debridement alone. Grid keratotomy may predispose cats with corneal ulcers to develop a corneal sequestrum. PMID- 11280408 TI - Clinical outcome and diseases associated with extreme neutrophilic leukocytosis in cats: 104 cases (1991-1999). AB - OBJECTIVE: To describe diseases, prognosis, and clinical outcomes associated with extreme neutrophilic leukocytosis in cats. DESIGN: Retrospective study. ANIMALS: 104 cats with extreme neutrophilic leukocytosis. PROCEDURE: Medical records from 1991 to 1999 were examined to identify cats that had > or =50,000 WBC/microl with > or =50% neutrophils. Signalment, absolute and differential WBC counts, rectal temperature, clinical or pathologic diagnosis, duration and cost of hospitalization, and survival time were reviewed. RESULTS: Mean age of cats was 8.3 years, mean WBC count was 73,055 cells/microl, and mean absolute neutrophil count was 59,046 cells/microl. Mean duration of hospitalization was 5.9 days, and mean cost of hospitalization was $2,010. Twenty-nine (28%) cats were febrile, and 63 (61%) cats died. Overall median survival time was 30 days. Cats with neoplasia were nearly 14 times as likely to die unexpectedly as cats with other diseases. CONCLUSIONS AND CLINICAL RELEVANCE: Extreme neutrophilic leukocytosis was associated with a high mortality rate. The prognostic importance of extreme neutrophilic leukocytosis should not be overlooked. Cats and dogs have similar diseases, mortality rates, and treatment costs associated with extreme neutrophilic leukocytosis. PMID- 11280410 TI - Effect of deslorelin acetate on gonadotropin secretion and ovarian follicle development in cycling mares. AB - OBJECTIVE: To evaluate gonadotropin secretion and ovarian function after administration of deslorelin acetate to induce ovulation in mares. DESIGN: Randomized controlled trial. ANIMALS: 16 healthy mares with normal estrous cycles. PROCEDURE: 8 control mares were allowed to ovulate spontaneously, whereas 8 study mares received deslorelin to induce ovulation when an ovarian follicle > 35 mm in diameter was detected. Follicle development and serum concentrations of gonadotropins were monitored daily during 1 estrous cycle. Pituitary responsiveness to administration of gonadotropin-releasing hormone (GnRH) was evaluated 10 days after initial ovulation. RESULTS: Interovulatory intervals of mares treated with deslorelin (mean +/- SD, 25.6 +/- 2.6 days) were longer than those of control mares (22.9 +/- 1.8 days). Diameter of the largest follicle was significantly smaller during 2 days of the diestrous period after ovulation in deslorelin-treated mares than in control mares. Concentrations of follicle stimulating hormone (FSH) were lower in deslorelin-treated mares on days 5 through 14 than in control mares. Concentrations of luteinizing hormone were not different between groups during most of the cycle. Gonadotropin release in response to administration of GnRH was lower in mares treated with deslorelin acetate than in control mares. CONCLUSIONS AND CLINICAL RELEVANCE: Administration of deslorelin was associated with reduction in circulating concentrations of FSH and gonadotropin response to administration of GnRH during the estrous cycle. Low concentration of FSH in treated mares may lead to delayed follicular development and an increased interovulatory interval. PMID- 11280409 TI - Factors associated with Salmonella shedding among equine colic patients at a veterinary teaching hospital. AB - OBJECTIVE: To evaluate factors potentially associated with fecal Salmonella shedding among equine patients hospitalized for colic at a veterinary teaching hospital and to determine the effects of probiotic treatment on fecal Salmonella shedding and clinical signs. DESIGN: Longitudinal study and controlled trial. ANIMALS: 246 equine colic patients. PROCEDURE: History and medical information were obtained from patient records. Fecal and environmental samples were submitted for aerobic bacterial culture for Salmonella enterica. Fifty-one patients were treated with a commercially available probiotic; 46 were treated with a placebo. Logistic regression was used to evaluate data. RESULTS: Salmonella organisms were detected in feces from 23 (9%) patients at least once during hospitalization. Patients were more likely to shed Salmonella organisms if diarrhea was evident < or = 6 hours after hospitalization and duration of hospitalization exceeded 8 days (odds ratio [OR], 20.3), laminitis developed during hospitalization (OR, 12.0), results of nasogastric intubation were abnormal (OR, 4.9), leukopenia was evident < or =6 hours after hospitalization (OR, 4.6), or travel time to the teaching hospital exceeded 1 hour (OR, 3.5). Horses treated with the probiotic did not differ from control horses in regard to likelihood of fecal Salmonella shedding (OR, 1.5) or prevalence of clinical signs. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that certain risk factors are associated with fecal shedding of S enterica among equine patients hospitalized at a veterinary teaching hospital because of colic and that pathogen monitoring in patients and the hospital environment and use of barrier nursing precautions for equine colic patients are beneficial. PMID- 11280411 TI - Response to immunotherapy in six related horses with urticaria secondary to atopy. AB - Urticaria secondary to atopy may be a familial problem in some horses. Immunotherapy using a vaccine containing antigens selected on the basis of history and results of intradermal testing can be an effective method of managing atopy in horses; a response to therapy may be seen within 2 months. PMID- 11280412 TI - Sensitivity and specificity of serum copper determination for detection of copper deficiency in feeder calves. AB - OBJECTIVE: To determine the relationship between serum and liver copper concentrations and evaluate serum copper determination for diagnosis of copper deficiency in juvenile beef calves. DESIGN: Cross-sectional study. ANIMALS: 105 juvenile beef calves. PROCEDURE: Copper concentrations were measured in paired liver and serum samples from 6- to 9-month-old beef calves. Regression models that predicted liver copper concentration as a function of serum copper concentration were developed. Sensitivity and specificity of serum copper concentration for detection of low liver copper concentration were determined, using a range of serum copper concentrations as test endpoints. Positive and negative predictive values were calculated. RESULTS: The association between serum and liver copper concentrations was significant; however, regression models accounted for only a small portion of the variation in liver copper concentrations. For a serum copper concentration endpoint of 0.45 microg/g, sensitivity and specificity for detection of low liver copper concentration were 0.53 and 0.89, respectively. Positive and negative predictive values of serum copper concentration for detection of low liver copper concentration ranged from 0.37 to 0.85 and 0.63 to 0.94, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Regression models are inappropriate for predicting copper status as a function of serum copper concentration. Serum copper concentration is fairly specific for detection of low liver copper concentration but only marginally sensitive when serum copper concentration of 0.45 microg/g is used as a test endpoint. The value of serum copper concentration as a diagnostic indicator depends on prevalence of copper deficiency. PMID- 11280413 TI - Practitioner review: psychological sequelae of head injury in children and adolescents. AB - Many children suffer an injury to the head at some time, but relatively few of these cause major problems. However, in a few cases the sequelae can be far reaching. This review considers how to evaluate the significance of a head injury. Factors to consider are (1) injury variables: cause, severity and type of injury; (2) child variables: premorbid functioning, age, and developmental level of the child both at injury and at assessment; and (3) the cognitive, behavioural, and emotional problems that may arise. Parental beliefs and knowledge about the injury as well as the overall effect of an injury on the family as a whole are also considered. PMID- 11280414 TI - Malnutrition and mental development: is there a sensitive period? A nested case control study. AB - To examine the possibility that there is an early sensitive period for the effects of malnutrition on cognitive development, three groups of children (N = 197) were recruited from a birth cohort with known growth characteristics in south-west Ethiopia (N = 1,563). All had initial weights > or = 2,500 g. Early growth falterers dropped in weight below the third centile (z < -1.88) of the NCHS/WHO reference population in the first 4 months. Late growth falterers were children not in the first group whose weights were below the third centile at 10 and 12 months. Controls were a stratified random sample with weights above the third centile throughout the first year. All children were tested blind at 2 years using the Bayley Scales of Infant Development, adapted for use in Ethiopia. Mean (SD) scores on the psychomotor scale were 10.2 (3.7) in the controls, 6.6 (4.2) in the early growth falterers, and 8.5 (4.3) in the late growth falterers. For the mental scale they were 28.9 (5.8), 22.6 (6.2), and 26.6 (6.1) respectively. Both overall differences were statistically significant at p < .001, and planned comparisons between the control and the combined growth faltering groups, and between the early and later growth faltering groups, showed that each difference was statistically significant for both scales. However, early weight faltering was associated with weight at the time of testing (r = .33), which was associated with scores both on the psychomotor (r = .53) and the mental scale (r = .49). After taking weight at the time of testing into account there was no additional effect attributable to the timing of growth faltering. In this population, therefore, early malnutrition does not have specific adverse effect beyond the contribution that it makes to enduring malnutrition over the first 2 years. PMID- 11280415 TI - Genetic influences on language impairment and literacy problems in children: same or different? AB - Data from two twin studies are examined to assess genetic and environmental influences on literacy, and the etiological relationship between language and literacy. Study 1 used children from 86 families previously recruited for a study of the genetics of specific language impairment (see Bishop, North, & Donlan, 1995), who completed tests of single-word reading and spelling. Literacy problems in this sample were common, were strongly heritable, and showed a close genetic relationship with poor nonword repetition. Study 2 included two subsets of children: 37 twin pairs who had taken part in study 1, 3 to 4 years earlier, and 100 twin pairs recruited from the general population by Bishop et al. (1999). All children were given a standardised test of nonword reading. There was no genetic influence on nonword reading ability, either across the normal range, or at the lower extreme, though there were significant associations with some social variables. However, bivariate DeFries Fulker analysis suggested that in this study, as in study 1, there was shared genetic variance between poor nonword repetition and literacy deficits. It is concluded that poor nonword repetition, which is known to be highly heritable, puts the child at risk for literacy problems. However, in a general population sample, such as that included in study 2, poor nonword repetition is a relatively rare correlate of literacy problems, which are more likely to have an environmental origin. Thus the different pattern of results in the two studies can be explained if one assumes that genetic influences are substantial only when literacy problems are severe and/or accompanied by oral language difficulties. PMID- 11280416 TI - Psychological mechanisms in hyperactivity: I. Response inhibition deficit, working memory impairment, delay aversion, or something else? AB - This study tested the predictions of three different theories of hyperactivity: response inhibition deficit, working memory impairment, and delay aversion. A sample of 51 pervasively hyperactive children and 119 control children, identified by screening a general population sample of 1,316 twin pairs, were assessed on tests relating to each of these theories. The hyperactive group performed worse than the control group on the delay aversion measure and some of the working memory tasks. Controlling for IQ removed the significant group differences on the working memory measures, however. There were no significant group differences on the inhibition variables derived from the stop task. However, there was evidence of a pattern of responding on the stop task that was strongly characteristic of hyperactivity: hyperactive children were variable in their speed, generally slow and inaccurate in responding. This pattern of responses may indicate a nonoptimal effort/ activation state. Hyperactive girls were indistinguishable from hyperactive boys in their performance on the tasks. PMID- 11280417 TI - Psychological mechanisms in hyperactivity: II. The role of genetic factors. AB - The main aim of this study was to combine two research approaches to hyperactivity: the behaviour genetic approach and the testing of psychological theories of hyperactivity. For a sample of 268 twin pairs aged 7-11 years we obtained ratings on the Conners' scales from both teachers (CTRS-28) and parents (CPRS-48). Forty-six hyperactive twin pairs (pairs in which at least one twin was pervasively hyperactive) and 47 control twin pairs were assessed on a psychological test battery. Confirming findings from previous twin studies, a substantial proportion of the variance in hyperactivity considered as a dimension was due to genetic effects. There was significant evidence of genetic effects also on extreme hyperactivity, although the present group heritability estimates were somewhat lower than those reported in most previous studies. We investigated the possibility that the psychological mechanisms we reported to be associated with hyperactivity (Kuntsi, Oosterlaan, & Stevenson, 2001) share common genetic factors with hyperactive behaviour. The data produced significant evidence of such shared genetic effects only on hyperactivity and the variability of reaction times. Given that the high variability in speed of responding would indicate a state-regulation problem, this is the psychological mechanism that could possibly be the "link" between genetic effects and hyperactive behaviour. PMID- 11280418 TI - The development of a brief screening measure of emotional distress in children. AB - We report several studies developing a parent-rated measure of emotional distress for children in Singapore, with the key objectives being to derive a very brief valid measure of global distress. The refined item set comprised behaviourally expressed broad manifestations of emotional distress. Three developmental studies were undertaken, with the first two involving parental ratings on the measure for validation against clinician-rated distress levels, while also testing two rating options for the measure. We established clear comparative advantages to the rating anchors used in the Revised Rutter Scales. High inter-rater agreement was established across parental ratings, with the latter finding supporting objectives for the measure. Paternal scores correlated more strongly than maternal scores with clinician-generated distress scores. Additional properties of the measure were tested in a large community sample of nearly 2,000 Singapore schoolchildren in their last 2 years of primary school, allowing prevalence estimates and mean scores to be derived for each item. Here, girls and boys received identical total scores, scores were also independent of the number of children in the family and of ordinal position, and mothers returned higher scores than fathers. PMID- 11280419 TI - Development and current functioning in adolescents with Asperger syndrome: a comparative study. AB - Adolescents with Asperger syndrome (AS: without delay in speech development, diagnosed according to ICD-10 clinical criteria) were compared with a group with high-functioning autism (HFA: all with delayed speech development), and a group with conduct disorder (CD). Family and genetic studies suggest that Asperger syndrome and autism form part of the same spectrum, whereas the social impairments in conduct disorder are assumed to have different origins. The aims were to explore the relationships between early speech development and other aspects of functioning in autistic disorders, and to compare autistic and nonautistic social impairments. Early and current behaviour and IQ profiles were investigated. The CD group were clearly different from both the AS and HFA groups. The AS group tended to have less severe early behavioural abnormalities than the HFA group, and were unlikely to have speech abnormalities, but other communicative, social, and restricted/ stereotyped behavioural difficulties were largely of a similar pattern to the abnormalities in the HFA group. Eighty per cent of the AS group met criteria for autism on the diagnostic algorithm associated with the Autism Diagnostic Interview-Revised. By adolescence, the AS group were reported to be as abnormal as the HFA group but in structured 1:1 interaction their conversation was better. IQ profile in the AS group showed relative strength on verbal measures, unlike the HFA group, but relatively good performance on the Block Design subtest of the WISC/WAIS was a feature of both the AS and HFA groups. The results indicate closely similar behavioural manifestations may arise by adolescence despite differences in speech development. Follow-up studies and further family investigations will be required to clarify the origins of these and other patterns of autistic development. PMID- 11280420 TI - The "Reading the Mind in the Eyes" Test revised version: a study with normal adults, and adults with Asperger syndrome or high-functioning autism. AB - In 1997 in this Journal we published the "Reading the Mind in the Eyes" Test, as a measure of adult "mentalising". Whilst that test succeeded in discriminating a group of adults with Asperger syndrome (AS) or high-functioning autism (HFA) from controls, it suffered from several psychometric problems. In this paper these limitations are rectified by revising the test. The Revised Eyes Test was administered to a group of adults with AS or HFA (N = 15) and again discriminated these from a large number of normal controls (N = 239) drawn from different samples. In both the clinical and control groups the Eyes Test was inversely correlated with the Autism Spectrum Quotient (the AQ), a measure of autistic traits in adults of normal intelligence. The Revised Eyes Test has improved power to detect subtle individual differences in social sensitivity. PMID- 11280421 TI - A study of memory functioning in individuals with autism. AB - Memory tasks were administered to 14 high-functioning individuals with autism and 14 typically developing individuals matched on chronological age and verbal intelligence. The tasks consisted of free and cued recall of 15 semantically unrelated words in 3 encoding conditions: phonological encoding, semantic encoding, and a no encoding orientation. In both groups, semantic orientation led to better free recall than did orientation toward syllabic encoding or absence of orientation. In contrast, semantic cues at retrieval led to better cued recall than phonological cues in typically developing individuals, whereas both types of cue had the same effect in prompting cued recall for individuals with autism. These findings are incompatible with the hypothesis of an amnesic deficit and do not support the notion of executive or semantic deficits in the memory problems of autistic individuals, at least for those with a high level of functioning. It is proposed that these findings can be accounted for by enhanced phonological processing in autism. This interpretation is consistent with other findings of enhanced processing of low-level perceptual information in the visual and auditory modality in autism. PMID- 11280423 TI - Advancing advanced mind-reading tests: empathic accuracy in adults with a pervasive developmental disorder. AB - Research using advanced but static mind-reading tests with high-functioning adults with a pervasive developmental disorder (PDD) provided evidence for subtle social cognitive deficits. In the present study, adults with PDD were unimpaired on such tasks, relative to individually matched normal controls. Significant differences between the two groups were, however, found on a more naturalistic empathic accuracy task developed for this study. Participants viewed two videotaped interactions that both depicted a male and female stranger having an initial conversation and were asked to infer the unexpressed thoughts and feelings of the four targets. Subjects with PDD performed significantly worse on the second video. These findings suggest that the mind-reading deficit of a subgroup of able adults with PDD may only be apparent when a sufficiently complex naturalistic assessment method is being used. PMID- 11280422 TI - Executive functioning in high-functioning children with autism. AB - Executive functioning was investigated in 34 children (24 boys and 10 girls) with developmental language disorder (DLD) and 21 children (18 boys and 3 girls) with high-functioning autistic disorder (HAD) matched on Full Scale IQ, Nonverbal IQ, age (mean age 9 year, 1 month), and SES. The DLD group had a Verbal IQ that was 10 points higher than the HAD group. These children were given the Wisconsin Card Sorting Test (WCST), the Mazes subtest from the WISC-R, the Underlining test, and the Rapid Automatized Naming test. In addition, these children were given the Vineland Scales of Adaptive Functioning and the Wing Diagnostic Symptom Checklist in order to assess severity of autistic symptomatology. Results indicated that the only significant difference between the two groups on the cognitive tasks was perseverative errors on the WCST; there was no significant difference on total number of categories achieved or total number of errors on the WCST or on the other executive function measures. There was also significant overlap in the scores between the two groups and the difference in perseverative errors was no longer significant when Verbal IQ was partialled out. Executive functioning was strongly related to all IQ variables in the DLD group and particularly related to Verbal IQ in the HAD group. Although there was a relationship in the HAD group between executive functioning and adaptive functioning, as well as between executive functioning and autistic symptomatology, these relationships were generally no longer significant in the HAD group after the variance due to Verbal IQ was accounted for. The results are interpreted to indicate that although impaired executive functioning is a commonly associated feature of autism, it is not universal in autism and is unlikely to cause autistic behaviors or deficits in adaptive function. PMID- 11280424 TI - Let's clear the air of second hand smoke! PMID- 11280426 TI - Patterns of alcohol use and misuse among elderly rest home residents in Christchurch. AB - AIMS: To determine the prevalence of alcohol use and misuse among elderly rest home residents in Christchurch. METHODS: A cross-sectional prevalence survey was conducted among 175 residents aged 65 years and over, randomly selected from 30 rest homes in Christchurch, in 1998. Hazardous patterns of alcohol consumption in the past twelve months were determined by the Alcohol Use Disorders Identification Test (AUDIT) questionnaire, and alcohol dependence in the past 12 months and lifetime was determined by a strctured clinical interview using DSM-IV criteria. RESULTS: Of 246 eligible participants, 175 (71.1%) residents were interviewed, 115 women and 60 men, mean age, 82.6 years (SD=7.8) compared with 83.2 years (SD=6.3) for non-participants. The prevalence of hazardous patterns of alcohol consumption in the past twelve months by the AUDIT (cut-off score 8) was 5.1% (95% CI = 1.8-8.4). According to DSM-IV criteria, the prevalence of lifetime alcohol dependence was 20.5% (95% CI = 13.5-27.6) and for the past twelve months was 0.5% (95% CI = 0-1.7). The prevalence of lifetime alcohol dependence was significantly higher in men 36.7% (95% CI = 23.2-50.1) than women 12.2% (95% CI = 5.6-18.8) (p = 0.0001). CONCLUSIONS: In spite of advanced age, a small proportion of elderly rest home residents consumed quantities of alcohol that put them at risk of future damage to physical or mental health. Lifetime prevalence of alcohol dependence was comparable to the general population estimates and was higher in men than women. PMID- 11280425 TI - A serological survey of antibodies to rabbit haemorrhagic disease virus (rabbit calicivirus disease) in two rural Central Otago communities. AB - AIMS: To determine whether individuals from two rural communities with heavy exposure to the Rabbit Haemorrhagic Disease Virus (RHDV) developed antibodies to this virus. METHODS: Sera were assayed using competition ELISA (cELISA) and solid phase ELISA (spELISA). Exposure estimates were based on answers to an interviewer administered questionnaire. RESULTS: Of the 104 participants, 79 were considered to have experienced high or medium exposure, many of whom described specific exposures. There were 58 people who reported contact with RHDV infected bait, organ homogenate mixtures or rabbit body fluids. A one-way analysis of variance (Kruskal Wallis) found that human cELISA results were differently distributed from both strongly RHDV positive rabbits (chi2(1) = 27.37, p < 0.001) and weakly RHDV positive rabbits (chi2(1) = 27.35, p < 0.001). The distribution of assay results in each exposure group did not differ in either cELISA (chi2(2) = 2.49, p = 0.29) or spELISA (chi2(2) = 1.70, p = 0.43). Relatively fewer results were categorised as reactive (two 'barely' positive and two doubtful) than in a previous survey of 493 unexposed people. None of the five positive results categorised by the less specific spELISA occurred in people described as 'barely' positive or doubtful by cELISA. CONCLUSIONS. No serological evidence of infection with RHDV was found in a cohort including many heavily exposed individuals. PMID- 11280427 TI - The rapid whole blood agglutination d-dimer assay has poor sensitivity for use as an exclusion test in suspected deep vein thrombosis. AB - AIMS: Several clinical studies have proposed using d-dimer as an initial screening test to exclude thrombosis in cases of suspected (DVT). In published series, these assays have variable sensitivity, raising concerns that they may not be sufficiently robust for clinical practice. The aim of the study was to examine the sensitivity of two commercially available d-dimer assays and to assess their value and safety as initial screening tests in suspected DVT. METHODS: In this prospective study, blood samples were collected for d-dimer measurement (SimpliRED assay and IL test d-dimer) in all patients presenting to the emergency department over a twelve month period. All patients underwent compression ultrasound scanning as the primary diagnostic procedure. RESULTS: 235 patients were included in the study. 51(22.8%) had a DVT confirmed on ultrasound. The SimpliRED assay was positive in only 33 cases, with seventeen cases of confirmed DVT giving a negative result (six cases with proximal vein thrombosis). Assay sensitivity was 66%, with a negative predictive value of 88.9%. The IL test gave three false negatives (all below knee thromboses) giving assay sensitivity and negative predictive value of 94.1% and 96.8% respectively. CONCLUSIONS: The precise role of d-dimer testing in the diagnosis of venous thrombosis has yet to be established. From our results and a review of published series, we conclude that the SimpliRED assay is too insensitive to use as a reliable exclusion test in cases of suspected DVT, however, the more sensitive automated IL test d-dimer may have a role in the initial assessment. We propose that the IL d-dimer test is used in conjunction with a pre-test probability score to identify patients at low risk of DVT and recommend that this approach is tested in a clinical study before introduction into practice. PMID- 11280428 TI - The clinical significance of atypical squamous cells of undetermined significance: a laboratory audit of cervical reporting. AB - AIM: To determine the outcomes in women diagnosed with 'Atypical Squamous Cells of Undetermined Significance' (ASCUS) on cervical smears. METHODS: All diagnoses of ASCUS on cervical smears made at Southern Community Laboratories (SCL) in Christchurch in 1996 were retrieved from the SCL database and correlated with all available previous and subsequent smear and biopsy results from these patients. The outcome was reported as the most significant (highest grade) cervical smear or biopsy over the following two year period. RESULTS: 278 women had smear results of ASCUS in 1996, reflecting 2.3% of total cervical smear diagnoses at SCL (Christchurch) for that period. Follow-up was available for 260 (94%). 61% had benign (normal or inflammatory) changes, 6% had persistent ASCUS (smear only), 18% had a Low Grade Squamous Intraepithelial Lesion (LSIL), and 15% had a High Grade Squamous Intraepithelial Lesion (HSIL). All women with ASCUS who subsequently developed HSIL had persistent abnormal smears. CONCLUSIONS: An ASCUS smear result indicates a group of women who have an increased risk for detection of HSIL. The effectiveness of routine Pap smears for detection of cervical cytologic abnormality is confirmed. PMID- 11280429 TI - Getting the message across: sun protection information in media weather reports in New Zealand. AB - AIMS: To assess the public reach, awareness, understanding and response to the burn time and the Ultra Violet Index (UVI) in media weather reports in New Zealand. METHODS: Data from a representative sample (n = 396), ages 16-44 years, were gathered over four consecutive summer weeks of 1999 via a random digit dialling telephone survey. Items collected included sources of weather reports and their frequency of use; knowledge, understanding, perception and use of the burn time and the UVI; sun-related beliefs, attitudes and behaviours. RESULTS: Exposure to weekend weather bulletins was sustained, and occurred mainly via television (83%) and radio (50%). There was greater awareness of the burn time than the more recent UVI (89% vs 43%). The UVI was less often used to guide sun protection actions (49% vs 63%) but better understood (94% vs 66%) and more often recalled along with sun protective messages (56% vs 32%) than the burn time. Few could describe the burn time or the UVI for the past Sunday. Self-perceived understanding of the burn time was higher than its measured, sub-optimal, comprehension (96% vs 65%). CONCLUSIONS: Further efforts are needed to promote the UVI, particularly on TV1 and on radio, and to reach younger adults and less educated groups. For a transition period, presentation of the burn time should be restricted to complementing the UVI. Thereafter, the UVI should become the standard indicator of UV level in New Zealand. PMID- 11280430 TI - The future of high speed molecular biology in medicine. PMID- 11280431 TI - Obtaining consent for epidural analgesia for women in labour. PMID- 11280432 TI - The attitude of The Medical Association to medical services and the Social Security Act, 1938. PMID- 11280433 TI - Prognosis in children with melanoma. PMID- 11280434 TI - Continuity of patient care and safety. PMID- 11280435 TI - Effect of disciplinary complaints. PMID- 11280436 TI - ACE inhibitors for hypertension. PMID- 11280437 TI - Liposomal bleomycin: increased therapeutic activity and decreased pulmonary toxicity in mice. AB - Conventional and sterically stabilized liposomes derived from phosphatidylcholine or the antitumor agents, hexadecylphosphocholine and octadecyl-(1,1-dimethyl-4 piperidino-4-yl)-phosphate, as bilayer forming constituents, containing bleomycin, were developed and tested. Liposomal encapsulation of bleomycin enhanced strongly the antitumor activity against P388 leukemia and the Lewis lung carcinoma. This effect was clearly dependent on the size and lipid composition of the bleomycin-containing liposomes. The therapeutic effects were nearly equal for liposomal and free bleomycin in the B16 melanoma. The partial replacement of phosphatidylcholine by alkylphospholipids and the inclusion of polyethylene glycol modified lipids for sterical stabilization did not further improve the therapeutic efficacy but increased, in some cases, the toxicity of liposomes. Bleomycin-induced lung injury was not observed if liposomal bleomycin was administered. PMID- 11280438 TI - Cell culture and animal studies for intracerebral delivery of borocaptate in liposomal formulation. AB - The efficacy of a liposomal formulation for intracerebral delivery of borocaptate (BSH) to brain tumor cells has been investigated using cell culture to study BSH uptake and persistence and using tumor-bearing rats to determine BSH distribution in the brain. During a 16-hr incubation, cellular uptake of BSH solution or BSH liposomal formulation was similar. However, the cellular persistence of BSH greatly increased when BSH was present in liposome. The differences in cellular persistence for BSH solution and BSH-loaded liposomes were significant both in 12 hr and 24-hr incubation experiments (p < 0.05 and p < 0.01, respectively). For the studies involving tumor-bearing rats, BSH level in tumor tissue was significantly higher than that in normal brain tissue at 2 hr and 6 hr after intracerebral injection of BSH-loaded liposomes (p < 0.01). Our study indicated that the liposomal formulation enhanced cellular persistence of BSH in tumor cells and therefore favored the boron accumulation in the cells. With the prolonged physical retention of liposomes at the local injection site and the cellular retention of BSH enhanced by the liposomes, the intracerebral delivery of BSH using liposomal formulation may provide an effective boron delivery approach for boron neutron capture therapy of brain tumors. PMID- 11280439 TI - In vivo absorption studies of insulin from an oral delivery system. AB - Alginate microspheres prepared by an emulsion-based process were loaded with insulin by a remote loading process. We observed that the time of exposure, pH of the remote loading medium, and beta-cyclodextrin complexation of insulin influenced drug loading. In vivo absorption studies of insulin from optimized microspheres were carried out in diabetic albino rats. Serum sugar levels on administration of multiple oral doses of the microspheres and a radioimmunoassay for serum insulin indicated absorption of insulin from the gastrointestinal region. This process could be utilized for the development of an oral insulin delivery system. PMID- 11280440 TI - Formulation, characterization, and in vitro release of glyburide from proliposomal beads. AB - The objective of our study was to formulate and evaluate proliposomes in the form of enteric-coated beads using glyburide as a model drug. The beads were enteric coated with Eudragit L-100 by a fluidized bed coating process using triethyl citrate as plasticizer. Content uniformity of glyburide was estimated using HPLC analysis of beads dissolved in methanol. These proliposomal beads formed liposomes on disintegration in phosphate buffered saline (pH 7.4), which was confirmed by transmission electron microscopy. The dissolution study of enteric coated beads exhibited enhanced dissolution compared with pure drug and a marketed product. Liposomes can be successfully prepared for oral administration in the form of enteric-coated beads that may offer a stable system to produce liposomes for oral administration. PMID- 11280441 TI - Receptor-mediated gene delivery to HepG2 cells by ternary assemblies containing cationic liposomes and cationized asialoorosomucoid. AB - Unilamellar cationic liposomes have been prepared from an equimolar mixture of 3beta[N',N'-dimethylaminopropane)-carbomoyl] cholesterol (Chol-T), a higher homologue of 3beta[N',N'-dimethylaminoethane)-carbomoyl] cholesterol (DC-Chol), and dioleoylphosphatidyl-ethanolamine. The DNA binding capabilities of Chol-T and Chol-T/DOPE liposomes have been demonstrated in lipid impregnated paper-DNA binding assays and gel retardation experiments, respectively. These liposomes have been combined with pRSVL plasmid DNA and N-ethyl-N'-(3 trimethylpropylammonium) carbodiimide iodide modified asialoorosomucoid (Me+ CDI urea-AOM) to generate ternary electrostatic assemblies intended for selective entry into cells displaying the galactose-specific lectin. This effect has been evaluated in the human hepatocellular carcinoma cell line HepG2 in which high levels of luciferase activity were achieved (up to 1.84 x 10(7) relative light units/mg protein) after transfection with complexes containing liposomes (1-3 microg), Me+CDI urea-AOM (2 microg), and DNA (0.5 microg) in 0.5 mL culture medium. Transfections conducted in the presence of free asialoorosomucoid afforded much lower luciferase activity (up to 1.5 x 10(5) relative light units/mg protein) confirming that DNA uptake was predominantly via asialoorosomucoid receptor-mediated endocytosis. We concluded therefore that modular complexes used in our study display the carbohydrate moiety of the glycoprotein component prominently, thus permitting interaction of terminal galactose units with their cognate receptors on the cell membrane. PMID- 11280442 TI - Preparation and analgesic activity of Eudragit RS100 microparticles containing diflunisal. AB - Two different techniques, the quasi-emulsion solvent diffusion method and spray drying that provide polar and nonpolar preparation environments, were used to prepare microspheres from Eudragit RS100 (RS) (acrylic/methacrylic copolymer) incorporating the nonsteroidal anti-inflammatory drug diflunisal. The effects of pH on the preparation medium and drug/polymer ratio on production yield and drug incorporation, as well as on the in vitro drug release at pH 1.2 and 6.8 from tabletted microparticles, were evaluated. The drug-polymer interactions and the effect of diflunisal incorporation in the polymer matrix on drug crystallinity have been evaluated by using differential scanning calorimetry, IR and ultraviolet spectroscopy, x-ray diffraction, and microscopy analysis. A preliminary biological assay confirmed that diflunisal maintains its analgesic activity after intraperitoneal administration to rats. PMID- 11280443 TI - Fluid dynamics in man of an intraperitoneal drug delivery solution: 4% icodextrin. AB - Interest in targeting drugs into the peritoneal cavity for intra-abdominal cancers or infections is undergoing a revival as recent clinical trials have demonstrated, not only a regional advantage in concentration of the active agent, but also improved long-term outcomes. Solutions currently used for intraperitoneal (IP) drug delivery have short residence times, however, which can limit the exposure of all areas of the peritoneum to the active agent. Icodextrin 4% solution was compared with saline and a glucose-based peritoneal dialysis solution in a clinical study of IP residence time. The study was carried out during the fortnightly rest phase in 9 patients undergoing 5-fluorouracil (5-Fu) IP treatment for colorectal cancer. The volume remaining in the peritoneal cavity was measured at 0, 12, 24, 48, 72, and 96 hr after an instillation of 2 liters of each fluid. Saline (n = 3 dwells) and glucose (n = 3 dwells) peritoneal dialysis solutions were almost fully absorbed by 24 hr, and the patients experienced discomfort when using these solutions. In contrast, icodextrin 4% solution (n = 188 dwells) maintained its instilled volume for up to 48 hr, and half the instilled volume remained after 72 and 96 hr. This result would allow extensive and prolonged coverage of the peritoneal surface. Icodextrin 4% solution may be an effective vehicle to deliver therapeutic agents into the peritoneal cavity. PMID- 11280444 TI - The Berashis cell: a review--is it similar to the embryonic stem cell? PMID- 11280446 TI - Anesthesia for victims of nerve agents undergoing surgery: establishment of a management protocol. AB - Awareness of the effects of nerve agents still being stocked around the world has led to the establishment of protocols for rapid evacuation and decontamination of large civilian areas. Perioperative care protocol and procedures for the administration of anesthesia in lifesaving surgical intervention for combined multiple trauma and intoxicated casualties have not been established. There are also no guidelines for anesthetizing those casualties who had already been medicated on site. The only exception to anecdotal reports and isolated communications on these issues is the organophosphate poisoning database of substances which act similarly to the nerve agents or their antagonists. We gathered the few reports existing on the subjects in the literature and extrapolated the knowledge of the nature and action of various anesthetic drugs. We applied it to biological and physiological conditions that exist in combined chemical and multiple trauma casualties and consolidated the data into two charts that may serve as guidelines for preventing the avoidable hazardous outcomes. PMID- 11280445 TI - Increased levels of serum hepatocyte growth factor in patients with end-stage renal disease. AB - Blood levels of hepatocyte growth factor (HGF) have been found to be elevated in patients with chronic renal failure. The cause of the increase in this mitogen is unclear. We determined serum HGF levels in 34 patients on maintenance dialysis and ten healthy volunteers. Predialysis serum HGF levels were elevated in patients with end-stage renal disease as compared to control subjects (1.65 +/- 0.2 ng/mL vs 0.46 +/- 0.04 ng/mL, p<0.01). In addition, serum HGF levels were significantly higher in African-American dialysis patients compared to Caucasian patients (2.18 +/- 0.36ng/mL vs 1.18 +/- 0.12ng/mL, p<0.01). The observed differences could not be accounted for by variations in serum creatinine, serumalbumin, or blood pressure between the African-American and Caucasian patients. Serum HGF levels were elevated in patients with end-stage renal disease, and were higher in African-American than Caucasian patients, but the pathophysiology and significance of this finding remain unclear. PMID- 11280447 TI - Plasma levels of orally and subcutaneously administered pentoxifylline in rabbits. AB - In addition to its ability to improve microcirculation, pentoxifylline also has anti-tumor necrosis factor-alpha (TNF) actions which have prompted investigations into its potential efficacy in disease states involving elevated TNF levels. One such disease entity is AIDS where aberrant TNF seems to mediate axonal degeneration within thecentral nervous system. To this end, we have previously established a rabbit model of TNF-mediated axonal degeneration, and demonstrated that pentoxifylline attenuates this effect. Unfortunately, there has been to date only one limited report on the pharmacokinetics of pentoxifylline in rabbits. Therefore, the present report evaluates plasma levels of pentoxifylline and two of its primary metabolites through high-performance liquid chromatography after both the oral and subcutaneous administration of pentoxifylline. Our results indicate that in rabbits there is a very rapid absorption and metabolism of pentoxifylline after either oral or subcutaneous administration. In comparison with the mouse, the rabbit seems to absorb and metabolize pentoxifylline slower. In contrast to man, the rabbit had lower metabolite plasma levels than the parent drug itself. PMID- 11280448 TI - Regulation of dosage of conjugated equine estrogen is useful for add-back therapy. AB - In the hormonal treatment of uterine myomas, which are estrogen dependent, GnRH agonist (GnRHa) therapy has become widespread. However, the severe hypo estrogenic state induced by the GnRHa gives rise to annoying side effects. Although the risk of these side effects may be reportedly modified when GnRHa is combined with estrogen (add-back therapy), it is difficult to target serum estradiol (E2) concentration to stay within the therapeutic window (20 approximately 50 pg/mL) by administering 0.625 mg conjugated equine estrogen (CEE)/day. Also, there is great individual variation in the circulating E2 concentration by administering the same dosage of CEE. Therefore, the use of smaller quantities of CEE in different dosages may approximate more closely to the clinical situation. This article focuses on the methods of administration of CEE combined with GnRHa for women with symptomatic uterine myomas. PMID- 11280449 TI - Prognostic value of Hanifin and Rajka's feature sets in adult atopic dermatitis patients. AB - Prognostication of the course of atopic dermatitis (AD) is of special importance in prophylaxis of this disease. The aim of the study was to describe the AD minor feature sets, according to Hanifin and Rajka, on the basis of which one can predict the occurrence of extensive skin lesions, allergic rhinitis, atopic bronchial asthma, cataracts, hand and/or foot dermatitis, food intolerance, urticaria and elevated IgE serum in AD patients. Two-hundred-fifty-four patients were included in the study. Minor feature sets of prognostic value were selected using the MANOVA method in conjunction with the discriminant analysis. On the basis of the above sets, one can predict: involvement of more than 60% of body surface, occurrence of allergic rhinitis, elevated serum IgE, cataracts, atopic bronchial asthma, food intolerance and urticaria with the probability ratio of 90.8%, 82.8%, 79.9%, 78.4%, 77.3%, 77.1% and 76.1% respectively. In the examined group, no association was found between hand and/or foot dermatitis and the tendency towards skin infection. PMID- 11280450 TI - Prolonged survival in two cases of T-prolymphocytic leukemias with complex hypodiploid chromosomal abnormalities. AB - We encountered two cases of T-prolymphocytic leukemias (T-PLL) with complex hypodiploid chromosomal abnormalities. Both cases showed mild organomegaly and marked leukocytosis (144.5 x 10(9)/L, 102.6 x 10(9)/L, respectively). Although both cases developed into refractory progressive diseases at the terminal stage, the oral administration of dexamethasone was very effective for leukocytosis and thrombocytopenia in case 1 and oral cyclophosphamide was effective for reducing elevated leukocytes and the organomegaly in case 2. Despite the poor prognosis of T-PLL, our cases showed that less toxic therapies such as oral dexamethasone or cyclophosphamide may be the treatment of choice for patients with an indolent phase of T-PLL. Our study and previously reported findings suggest that complex hypodiploid chromosomal abnormalities are characteristic in T-PLL. PMID- 11280452 TI - The impact of myocardial viability as determined by rest-redistribution 201Tl single photon emission CT imaging and the choice of therapy on prognosis in patients with left ventricular dysfunction. AB - While thallium-201 (201Tl) single-photon emission CT (TSPECT) scintigraphy is a commonly used method for determining the viability of the myocardium, its value for predicting outcome is limited. The diagnosis of myocardial viability in patients with ischemic systolic left ventricular dysfunction might indicate which of them will benefit more from surgical revascularization. Forty patients (mean +/- SD aged 64.5 +/- 12 years, 33 males and 7 females) with impaired systolic left ventricular function (ejection fraction < or = 45%) underwent TSPECT examination. Twenty patients were surgically revascularized and 20 were treated medically. The patients were followed-up for a 34 +/- 10-month period and the cardiac long-term prognosis was evaluated. The significant viability percentage (SVP), defined as the percentage of the total number of segments showing a normal uptake of 201Tl redistribution divided by the total number of segments evaluated, was > or =55: this was observed in 18 patients. Among them, the cardiac event free survival was 100% in the surgical group versus 22% in the medical group. However, in patients with non-SVP, the survival was lower and not significantly different in the two treatment groups. Surgical revascularization is the preferred method of treatment in patients with ischemic systolic left ventricular dysfunction and myocardial viability as defined by TSPECT scintigraphy. PMID- 11280451 TI - First generation cephalosporins as therapy for uncomplicated pyelonephritis in children. A retrospective analysis. AB - There is no consensus opinion for the optimal management of pyelonephritis in children. We summarized our experience with first generation cephalosporins by a retrospective analysis of 129 pediatric patients with pyelonephritis who were treated either by first generation cephalosporins (97 patients, group 1) or broad spectrum antibiotics (32 patients, group 2). Group 1 patients were less likely to have reported previous urinary infections or anatomical urinary tract abnormalities (16.2% vs. 53.1%, p= 0.002) and pathogens other than E. coli (7.3% vs. 25%, p=0.02). Resistance to first generation cephalosporins was identified in 22.6% of pathogens cultured, however, only 7.5% of them had poor clinical responses and required alternative treatment replacement. Our findings show that first generation cephalosporins could be used in our region to treat pyelonephritis in an otherwise healthy child, and that they can provide therapeutic success even in the face of apparent bacterial resistance. PMID- 11280453 TI - Non-invasive diagnosis of a Schwannoma by ultrasonography: a case study. AB - A 52-year-old female came to this institution complaining of a right leg mass lesion. Ultrasonography showed a 16 x 12 x 20mm hypoechoic solid mass lesion in the right leg. The patient underwent surgery during the diagnosis of Schwannoma. Preoperative diagnosis of Schwannoma is difficult by examination. Computerized tomography and magnetic resonance imaging, showed the origin of Schwannoma. However, the lesion was noted to be in a direct continuity with the cord-like echogenic structure consistent with a nerve by echography. We prefer to conduct preoperative examination by ultrasonography in patients with Schwannoma. In conclusion, we have reported a case of Schwannoma diagnosed by non-invasive ultrasonography. PMID- 11280454 TI - Lyme disease--United States, 1999. AB - Lyme disease (LD) is caused by the tickborne spirochete Borrelia burgdorferi sensu lato and is the most common vectorborne disease in the United States. Surveillance for LD was initiated by CDC in 1982, and the Council of State and Territorial Epidemiologists designated it a nationally notifiable disease in January 1991. This report summarizes the number of LD cases reported to CDC during 1999. Although the number of cases decreased from 1998, the number of cases in 1999 was higher than the number reported during the early 1990s. LD can be prevented by avoiding tick-infested habitats, by using personal protective measures, by vaccination, by checking for and removing ticks attached to the body and clothes, and by reducing tick populations. PMID- 11280455 TI - Knowledge and use of folic acid among women of reproductive age--Michigan, 1998. AB - Neural tube defects (NTDs), which include spina bifida and anencephaly, are serious malformations that occur in the developing fetus during the first 17-30 days after conception. Consumption of supplements containing folic acid can reduce NTDs 50%-70%. In the United States, approximately 4000 pregnancies are affected by NTDs each year, including approximately 140 infants in Michigan. In 1992, the U.S. Public Health Service recommended that all women of childbearing age consume at least 400 microg of folic acid daily. In 1998, the Institute of Medicine reaffirmed that recommendation and added that women capable of becoming pregnant take 400 microg of synthetic folic acid daily from fortified foods and/or supplements and consume a balanced, healthy diet of folate-rich foods. This report summarizes findings from the 1998 Behavioral Risk Factor Surveillance System (BRFSS) about multivitamin use and folic acid knowledge among women of reproductive age in Michigan. The findings suggest that public health campaigns that promote the consumption of folic acid should target women who are young, unmarried, obese, smoke, eat few fruits and vegetables, and have a low level of education. PMID- 11280456 TI - Update on the supply of tetanus and diphtheria toxoids and of diphtheria and tetanus toxoids and acellular pertussis vaccine. AB - During the last quarter of 2000, the U.S. Public Health Service learned of a shortage of tetanus and diphtheria toxoids (Td) and tetanus toxoid (TT) resulting from decreased production of these vaccines by the two U.S. manufacturers. Previously published recommendations outlined priorities for use of the limited supply of Td and TT. The shortage was expected to be resolved by early 2001; however, on January 10, 2001, Wyeth Lederle (Pearl River, New York) announced it had stopped production of tetanus toxoid-containing products. Although a small amount of Td is produced by the University of Massachusetts for local distribution, Aventis Pasteur (Swiftwater, Pennsylvania) is now the sole nationwide distributor of Td and TT. Aventis Pasteur is shipping limited quantities of vaccine to assure a wide distribution of available doses. PMID- 11280458 TI - Frictional dermal melanosis (lifa disease) over bony prominences. AB - Seventy-one patients (70 females and one male) were studied. Their ages at presentation ranged from 16-43 years with a mean of 24.4 years. The duration of disease varied from three months to ten years with a median of 2.7 years. All patients were slim and had recorded using a washing agent (Lifa) during bathing with vigorous friction. Family history of the same disease was present in 11 (15.5%) patients. The pigmentation was brown-black with a rippled pattern and showed no contrast under Wood's light. The areas commonly affected were clavicles (71.8%), shins (36.6%), upper back (32.4%), Adam's apple (31%), lateral aspects of the arms (31%) and other areas. The histopathology of biopsies from 24 patients showed marked dermal melanosis. Amyloid deposits in the papillary dermis were present in one patient. This clinical and histopathological picture suggests that this common condition is related to frictional melanosis. PMID- 11280457 TI - Azithromycin monthly pulse vs daily doxycycline in the treatment of acne vulgaris. AB - Acne vulgaris is a common skin disease seen primarily in adolescent and young adults. As the treatment involves long term therapy with antibiotics, an agent with a long half life can be very useful in increasing the compliance. To evaluate the role of a monthly dose of azithromycin and compare it with daily doxycycline, we conducted this randomized comparative study. Sixty patients with moderate to severe acne were randomly assigned to two treatment groups, A & B. Patients in group A received 100 mg doxycycline daily in addition to topical 0.05% tretinoin cream, whereas patients in group B were given 500 mg azithromycin once a day for four days per month along with 0.05% topical tretinoin for a total of 12 weeks. Of the 60 patients, 22 in group A and 28 in group B were evaluated. The monthly dose of azithromycin was found to be as effective as daily doxycycline on a pure protocol basis and statistically significantly better than doxycycline by intention to treat analysis. PMID- 11280459 TI - A clinico-mycological study of dermatophytoses in Nepal. AB - A clinico-mycological study of 100 cases of dermatophytosis was done in the B.P. Koirala Institute of Health Sciences in the eastern region of Nepal. The incidence of dermatophytoses in this study was 4.54% with a M: F ratio of 2.5: 1. The commonest age group was 11-20 years old. A single clinical type was found in 68%; 32% had two or more clinical types. The study revealed tinea corporis (43%) as the most common clinical type followed by tinea cruris (33%) and tinea pedis (20%). Positive culture was obtained in 94% of cases. A total of eight different species of dermatophytes were isolated with T. rubrum (45.74%) as the most common species followed by T. mentagrophytes (26.6%), T. tonsurans (11.7%), M. audouinii (8.36%), E. floccosum (4.26%), T. schoenlenii (2.13%), T. violaceum (2.13%) and T. verrucosum (1.06%). PMID- 11280460 TI - Clinico-myocological profile of tinea capitis in North India and response to griseofulvin. AB - There is a paucity of literature on tinea capitis from North India. The response to griseofulvin has not been studied as well. We studied 153 consecutive patients of tinea capitis for clinical patterns, causative dermatophytic species, clinico etiological correlation, and response to griseofulvin. Culture and sensitivity were done on all patients. All patients were treated with griseofulvin for 6-8 weeks; non-responders were further treated with fluconazole. Ninety percent of the patients were less than 15 years of age, 75% belonged to poor socioeconomic groups and 19% had a family history of tinea capitis. The seborrheic variant was the commonest clinical pattern seen in 47.8% of patients, followed by grey patch, black dot, kerion, and alopecia-areata-like tinea capitis in 35.9%, 8.5%, 6.5% and 1.3% of patients, respectively. Only 66% of patients had a positive culture. T. violaceum was the commonest dermatophytic species isolated in 38% patients. M. audouinii, T. schoenleinii, T. tonsurans, M. gypseum, T. verrucosum and T. mentagrophytes were isolated in 34%, 10%, 9%, 3%, 3% and 3% of patients, respectively. Of the isolates 94% were susceptible to griseofulvin, and 100% were susceptible to fluconazole. By using griseofulvin for 6-8 weeks 97.4% of the patients were cured; nonresponders required therapy with fluconazole for cure. To conclude, tinea capitis is still a disease of younger people of poor socioeconomic status. T. violaceum and M. audouinii are the most common responsible dermatophytes. The response to griseofulvin was excellent, and it should be used as a first line therapy. PMID- 11280461 TI - Fulminant and relentless cutaneous necrosis with excruciating pain caused by calciphylaxis developing in a patient undergoing peritoneal dialysis. AB - A 50-year-old Japanese female with chronic renal failure who had been on continuous ambulatory peritoneal dialysis developed fulminant systemic cutaneous necrosis that began as painful livedo reticularis-like skin lesions on her thighs. Because of disseminated vascular calcification within the muscular layer of her lower limbs, we eventually diagnosed her with calciphylaxis. The skin necrosis progressed rapidly, and she died of sepsis and pneumonia on the 53rd hospital day. In addition to her long-lasting severe hyperparathyroidism and extremely elevated serum phosphorus and calcium levels, mechanical, frictional stimulation inflicted on the local skin and administration of corticosteroids were suspected to have precipitated the calciphylaxis. Our lack of awareness of this disease in its early stages resulted in our missing the chance to do a parathyroidectomy that might have changed the course. It is important to know the clinical features of this rare disease in order to make a diagnosis as early as possible. PMID- 11280462 TI - Leukocytoclastic vasculitis with IgA deposits in angioimmunoblastic T cell lymphoma. AB - Angioimmunoblastic T cell lymphoma (AILD) is a type of peripheral T cell lymphoma associated with fever and generalized lymphadenopathy. Cutaneous manifestations are seen in approximately 40% of the patients. We report herein a Japanese male patient with AILD associated with generalized purpura. The histology of the purpura included leukocytoclastic vasculitis with IgA deposits, which is rare in this disease. Using in situ hybridization and PCR methods, we showed that the involved lymph node was positive for Epstein-Barr virus and that the purpura was negative. PMID- 11280463 TI - An association of twenty-nail dystrophy with vitiligo. AB - A rare association of twenty-nail dystrophy with segmental vitiligo is described in two patients. Vitiligo preceded the nail dystrophy. In both cases, all twenty nails were uniformly affected with the nail plates showing longitudinal striations and loss of luster. Longitudinal nail biopsy revealed a histological picture suggestive of eczematous changes and lichen planus respectively. Intramatrix injections of triamcinolone acetonide into the proximal and lateral nail folds were administered with considerable improvement. PMID- 11280464 TI - Bullous lichen planus arising in the skin graft donor site of a psoriatic patient. AB - The coexistence of psoriasis vulgaris and bullous disorders, particularly bullous pemphigoid, has been described previously. We present an unusual case of bullous lichen planus arising in the skin graft donor site of a psoriatic patient. To our knowledge, such an association has not been reported to date. PMID- 11280465 TI - Erythematous swelling of the lip associated with Sjogren's syndrome and mimicking cheilitis granulomatosa. AB - A 64-year-old Japanese woman developed therapy-resistant erythematous swelling of her upper lip. Our tentative clinical diagnosis of cheilitis granulomatosa was ruled out later by the laboratory findings including increased levels of anti nuclear-antibody (ANA), anti-SSA/Ro antibody, and positive Schirmer test as well as by a histopathological picture showing a dense perivascular infiltration of plasma cells and mononuclear cells in the dermis instead of granulomatous changes. To the best of our knowledge, this is the first patient in whom annular erythema associated with S ogren's syndrome involved only the upper lip and produced clinical features simulating cheilitis granulomatosa. PMID- 11280466 TI - Rod-shaped bodies resembling birbeck granule-like structures in endothelial cells of dermal capillaries in generalized granuloma annulare. AB - A previous study demonstrated that, in generalized granuloma annulare in the epidermis, the Langerhans' cell section area and the number of Langerhans' cell granules or Birbeck granules per cell section were increased, suggesting an active state of these Langerhans' cells. Reexamination by transmission electron microscopy of the same tissue, but in samples also containing dermal tissue, from the same subjects revealed endothelial cells with rod-shaped bodies resembling Birbeck granules or Birbeck granule-like structures. This finding has not been previously described in blood vessels of human skin and is described here. PMID- 11280467 TI - Pityriasis rotunda mimicking tinea cruris/corporis and erythrasma in an Indian patient. AB - Pityriasis rotunda is a rare disease characterized by perfectly round to oval, sharply defined, scaly, hypo/hyperpigmented patches of variable number and size located mainly over the trunk and proximal extremities. More than 95% of the reported cases in medical literature are from three countries/ethnic populations, namely Japan, South Africa (Bantu), and Italy (Sardinian islanders). To the best of my knowledge, no patient with the characteristic clinico-pathologic features has been reported from the Indian subcontinent. I report a 44-year-old man with eighteen pityriasis rotunda patches, persistent for nearly 20 years. The lesions in the groin and axillae closely resembled erythrasma and tinea, and he had received treatment for these conditions several times in the past. Histopathology of the skin biopsy showed thinning of the epidermis with a thinned-out granular layer and a sparse lymphomononuclear infiltrate in the dermis. A review of literature suggests that there are two subsets of the disease. The type I subset is comprised of pityriasis rotunda associated with systemic illness and is seen in Black or Oriental patients with no family history of the disease. The lesions tend to subside on treatment of the underlying illness. The type II subset patients are Caucasians as well as Blacks and Orientals with no underlying systemic illness. Familial occurrence is possible; lesions tend to be persistent and unresponsive to therapy. PMID- 11280468 TI - Laugier-Hunziker syndrome. AB - Laugier-Hunziker syndrome is a benign pigmentary disorder which manifests as macular hyperpigmentation of the lips and buccal mucosa. Some patients have longitudinal pigmented bands of nails. The syndrome has no systemic associations. Two patients of this rare syndrome are reported. Disorders producing similar pigmentary changes which must be differentiated are discussed. PMID- 11280469 TI - Linear and whorled nevoid hypermelanosis. AB - Linear and whorled nevoid hypermelanosis (LWNHM) is a reticulate pigmentary disorder with a sporadic occurrence, representing genetic mosaicism. It is characterised by hyperpigmented macules in a reticulate pattern along Blaschko's lines, sparing the mucous membranes and stabilising after one to two years. It may be associated with various neurological abnormalities. The disorder may resemble incontinentia pigmenti, epidermal nevus, or zebra-like hyperpigmentation clinically. We report LWMNHM in a 15-year-old girl with progressively increasing streaks of reticulate hyperpigmented macules arranged in a whorled pattern over the trunk and extremities, which appeared soon after birth. There was no history of any preceding eruption or any associated systemic abnormality. Histopathological examination revealed basal cell hyperpigmentation without any pigmentary incontinence. CT scan of the brain was normal. PMID- 11280470 TI - Giant cafe-au-lait macule in neurofibromatosis type 1. PMID- 11280471 TI - Diabetic foot disorders: a clinical practice guideline. American College of Foot and Ankle Surgeons. AB - Foot ulcerations, infections, and Charcot neuropathic osteoarthropathy are three serious foot complications of diabetes mellitus that can too frequently lead to gangrene and lower limb amputation. Consequently, foot disorders are one of the leading causes of hospitalization for persons with diabetes and can account for expenditures in the billions of dollars annually in the U.S. alone. Although not all foot complications can be prevented, dramatic reductions in their frequency have been obtained through the implementation of a multidisciplinary team approach to patient management. Using this concept, the authors present a Clinical Practice Guideline for diabetic foot disorders based on currently available evidence. The underlying pathophysiology and treatment of diabetic foot ulcers, infections, and the diabetic Charcot foot are thoroughly reviewed. Although these guidelines cannot and should not dictate the standard of care for all affected patients, they are intended to provide evidence-based guidance for general patterns of practice. The goal of a major reduction in diabetic limb amputations is certainly possible if these concepts are embraced and incorporated into patient management protocols. PMID- 11280472 TI - Continuity and discontinuity of behavioral inhibition and exuberance: psychophysiological and behavioral influences across the first four years of life. AB - Four-month-old infants were screened (N = 433) for temperamental patterns thought to predict behavioral inhibition, including motor reactivity and the expression of negative affect. Those selected (N = 153) were assessed at multiple age points across the first 4 years of life for behavioral signs of inhibition as well as psychophysiological markers of frontal electroencephalogram (EEG) asymmetry. Four month temperament was modestly predictive of behavioral inhibition over the first 2 years of life and of behavioral reticence at age 4. Those infants who remained continuously inhibited displayed right frontal EEG asymmetry as early as 9 months of age while those who changed from inhibited to noninhibited did not. Change in behavioral inhibition was related to experience of nonparental care. A second group of infants, selected at 4 months of age for patterns of behavior thought to predict temperamental exuberance, displayed a high degree of continuity over time in these behaviors. PMID- 11280473 TI - Children's probability intuitions: understanding the expected value of complex gambles. AB - Two experiments used Information Integration Theory to study how children judge expected value of complex gambles in which alternative outcomes have different prizes. Six-year-olds, 9-year-olds and adults (N = 73 in Study 1, N = 28 in Study 2) saw chance games that involved shaking a marble in a bicolored tube. One prize was won if the marble stopped on blue, another if it stopped on yellow. Children judged how happy a puppet playing the game would be, with the prizes and probability of the blue and yellow outcomes varied factorially. Three main results appeared in both studies: First, participants in all age groups used the normatively prescribed multiplication rule for integrating probability and value of each individual outcome--a striking finding because multiplicative reasoning does not usually appear before 8 years of age in other domains. Second, all age groups based judgment of overall expected value meaningfully on both alternative outcomes, but there were individual differences--many participants deviated from the normative addition rule, showing risk seeking and risk averse patterns of judgment similar to the risk attitudes often found with adults. Third, even the youngest children took probability to be an abstract rather than physical property of the game. Overall, in contrast to the traditional view, the present results demonstrate functional understanding of probability and expected value in children as young as 5 or 6. These results contribute to the growing evidence on children's intuitive reasoning competence. This intuition can, on the one hand, support surprisingly precocious performance in young children, but it may also contribute to the biases evident in adults' judgment and decision. PMID- 11280474 TI - Testing a core emotion-regulation prediction: does early attentional persistence moderate the effect of infant negative emotionality on later development? AB - To test the hypothesis that early attentional persistence will moderate the effect of infant negative emotionality on social competence, problem behavior, and school readiness at age 3, data collected as part of the NICHD Study of Early Child Care were subject to structural equation modeling analyses (N = 1,038). Consistent with Eisenberg et al.'s data on older children, high levels of negative emotionality were associated with low levels of social competence only when attentional persistence was poor. No such moderating effects of attentional persistence emerged in the case of behavior problems. And in the case of school readiness, findings indicated that high levels of negative emotionality predicted high levels of school readiness when attentional persistence was high, a result opposite to that found with respect to the prediction of social competence. PMID- 11280475 TI - Chronicity and instability of children's peer victimization experiences as predictors of loneliness and social satisfaction trajectories. AB - The present investigation was conducted to predict children's loneliness and social satisfaction growth curves from changes in their peer victimization status. Toward this aim, 388 children (193 boys, 195 girls) were interviewed at five points: as children entered kindergarten (in the fall) and spring of kindergarten through third grade. At each assessment, data were gathered on the frequency of children's peer victimization and degree of loneliness and social satisfaction. Groups were formed on the basis of timing and duration of children's victimization status. Hierarchical linear modeling was used to test several hypotheses regarding the nature of victimized children's growth curves. For instance, consistent with the Onset Hypothesis, the trajectories that emerged for children who moved from nonvictim to victim classification showed increasing levels of loneliness and decreasing social satisfaction. In contrast, findings for the Cessation Hypothesis were mixed, which suggests that children moving from victim to nonvictim status do not necessarily evidence significant improvements in loneliness or social satisfaction. The somewhat disparate trajectories that emerged for loneliness and social satisfaction are discussed. PMID- 11280476 TI - Conceptions of ability as stable and self-evaluative processes: a longitudinal examination. AB - It has generally been taken for granted that conceiving of ability as stable leads to negative self-evaluative processes, particularly in the face of failure. Yet, a close examination of the empirical findings suggests that the picture may be more complex. In this research, a three-wave longitudinal design spanning 12 months was employed. Older elementary school children (N = 932) indicated their conceptions of academic and social ability as stable to external forces and to internal forces. They also provided information about the importance they place on academic and social competence, their knowledge about academic and social performance, their preference for academic challenge, their perceptions of academic and social competence, and their attributions for academic and social performance. Children's grades in school and their acceptance by peers were obtained as indicators of performance. Over time, conceiving of ability as stable to external forces, particularly in the academic domain, appeared to heighten the importance placed on competence, performance knowledge, preference for challenge, perceptions of competence, and self-enhancing attributions. In contrast, conceptions of ability as stable to internal forces, particularly in the academic domain, appeared to be fostered by placing little importance on competence, a lack of performance knowledge, avoidance of challenge, negative perceptions of competence, self-deprecating attributions, and poor performance. PMID- 11280477 TI - Children's social reasoning about inclusion and exclusion in gender and race peer group contexts. AB - This study investigated whether children's and adolescents' judgments about exclusion of peers from peer group activities on the basis of their gender and race would differ by both age level and the context in which the exclusion occurred. Individual interviews about exclusion in several different contexts were conducted with 130 middle-class, European American children and adolescents. Younger children were expected to reject exclusion, by using judgments based on moral reasoning, regardless of the potential cost to group functioning, whereas older children were expected to condone exclusion on the basis of group membership in cases in which the inclusion of these children might interrupt effective group functioning. On measures of judgments, justifications for those judgments, and ratings of the appropriateness of exclusion, the vast majority of children used moral reasoning and rejected exclusion in contexts in which only the presence of a stereotype justified it. As expected, however, older children (13 years) were more likely to allow exclusion than younger children (7 and 10 years) when group functioning was threatened, and they justified this exclusion by using appeals to effective group functioning. PMID- 11280478 TI - Self-efficacy beliefs as shapers of children's aspirations and career trajectories. AB - This prospective study tested with 272 children a structural model of the network of sociocognitive influences that shape children's career aspirations and trajectories. Familial socioeconomic status is linked to children's career trajectories only indirectly through its effects on parents' perceived efficacy and academic aspirations. The impact of parental self-efficacy and aspirations on their children's perceived career efficacy and choice is, in turn, entirely mediated through the children's perceived efficacy and academic aspirations. Children's perceived academic, social, and self-regulatory efficacy influence the types of occupational activities for which they judge themselves to be efficacious both directly and through their impact on academic aspirations. Perceived occupational self-efficacy gives direction to the kinds of career pursuits children seriously consider for their life's work and those they disfavor. Children's perceived efficacy rather than their actual academic achievement is the key determinant of their perceived occupational self-efficacy and preferred choice of worklife. Analyses of gender differences reveal that perceived occupational self-efficacy predicts traditionality of career choice. PMID- 11280479 TI - The vicissitudes of measurement: a confirmatory factor analysis of the Emotional Autonomy Scale. AB - This study examined the factor structure of the Emotional Autonomy Scale (EAS) as proposed by Steinberg and Silverberg. Participants were from three independent samples of adolescents in grades 6 (n = 1,842), 8 (n = 1,769), and 10 (n = 1,232), with each sample consisting of three ethnic groups: African American, European American, and Mexican American. None of the confirmatory factor analyses for these samples supported the factor structure proposed by Steinberg and Silverberg. From the three models tested, the EAS is best described by the four originally proposed factors, combined with two method factors, one consisting of the positively worded scale items and one consisting of the negatively worded scale items. Results show that the EAS exhibits poor construct validity and behaves quite differently for the different grade and ethnic groups. The strong impact of method variance on the factor structure is discussed. Although various alternative solutions to the psychometric problems in the EAS are proposed, the most credible solution may be to reexamine the conceptual foundations of emotional autonomy and develop better measures of those concepts for adolescents. PMID- 11280480 TI - Biases in young children's communication about spatial relations: containment versus proximity. AB - Four experiments examined 3- and 4-year-olds' ability to communicate about containment and proximity relations. One hundred twenty-eight children either described where a miniature mouse was hiding in a dollhouse or they searched for the mouse after the experimenter described where it was hiding. The mouse was always hidden with a small landmark that was either in or next to a large landmark. When describing where the mouse was hiding, children were more likely to successfully disambiguate the small landmark when it was in the large landmark (e.g., under the plant in the dresser) than when it was next to the large landmark (e.g., under the plant next to the dresser). When searching for the mouse, 3-year-olds were faster to initiate their searches when the small landmark was in the large landmark than when it was next to the large landmark. Together, these results suggest that there are informational biases in young children's spatial communication. PMID- 11280481 TI - Autonomy and adolescent social functioning: the moderating effect of risk. AB - This study examined the moderating effect of risk on the relation between autonomy processes and family and adolescent functioning. The present sample comprised 131 adolescents from either a low-risk or high-risk social context, their mothers, and their peers. Observational ratings of autonomy processes within the mother-adolescent dyad were obtained, along with adolescent reports of the quality of the mother-adolescent relationship, and both adolescent and peer reports of the adolescent's functioning. Consistent with past research, in low risk families, behavior undermining autonomy was negatively related to relationship quality, and adolescents' expressions of autonomy were linked with positive indices of social functioning. In high-risk families, however, undermining of autonomy was positively linked with mother-adolescent relationship quality, and adolescents' expressions of autonomy were linked with negative indices of social functioning. Results are interpreted as demonstrating the ways in which the developmental task of attaining autonomy in adolescence is systematically altered depending on the level of risk and challenge in the adolescent's social context. PMID- 11280482 TI - A "natural experiment" in childrearing ecologies and adolescents' attachment and separation representations. AB - Employing a quasi-experimental design, this study explored the long-term effects of different childrearing ecological contexts. Participants were 131 adolescents (aged 16-18) from four groups: some who lived in a city, some from a kibbutz familial setting, some from a kibbutz communal setting, and a transitional group that included adolescents raised in a communal setting as young children who moved to a familial sleeping arrangement before the age of six. Adolescents' state of mind with regard to attachment and representations regarding separation were examined. Participants were administered the Adult Attachment Interview, the Separation Anxiety Test, and background questionnaires. The group raised in a communal setting in the kibbutz showed a higher incidence of nonautonomous attachment representations and less competent coping with imagined separations than did the other groups. By contrast, the transitional group was comparable to the city and the kibbutz familial groups. These results are discussed in light of the plasticity and adaptability of children to changed circumstances. PMID- 11280483 TI - Parental sensitivity, infant affect, and affect regulation: predictors of later attachment. AB - This longitudinal study on 94 families examined the extent to which parent sensitivity, infant affect, and affect regulation at 4 months predicted mother infant and father-infant attachment classifications at 1 year. Parent sensitivity was rated from face-to-face interaction episodes; infant affect and regulatory behaviors were rated from mother-infant and father-infant still-face episodes at 4 months. Infants' attachment to mothers and fathers was rated from the Strange Situation at 12 and 13 months. MANOVAs indicated that 4-month parent and infant factors were associated with infant-mother but not infant-father attachment groups. Discriminant Function Analysis further indicated that two functions, "Affect Regulation" and "Maternal Sensitivity," discriminated infant-mother attachment groups; As and B1-B2s showed more affect regulation toward mothers and fathers than B3-B4s and Cs at 4 months, and mothers of both secure groups were more sensitive than mothers of Cs. Finally, the association between maternal sensitivity and infant-mother attachment was partially mediated by infant affect regulation. PMID- 11280484 TI - Sisters, brothers, and delinquency: evaluating social influence during early and middle adolescence. AB - Although a number of studies have shown that brothers are highly correlated for delinquent behavior, much less research has been conducted on sisters. We propose that sisters, like brothers, show notable similarity for delinquent behavior, and also promote each other's delinquency through direct interaction. We examined these issues in 164 brother and sister pairs studied over a 4-year period (from early to middle adolescence) in a study of intact families in the rural Midwest. Sibling similarity for self-reports of delinquent behavior were highly correlated for both brothers and sisters. Conditional effects of high levels of hostile coercive sibling relationships and older sibling delinquency predicted younger sibling delinquency in both brother and sister pairs. For brothers, conditional effects were also detected for high levels of warmth-support, in contrast to sisters. The conditional effects of older sibling delinquency and relationship quality were shown to predict change in younger sibling delinquency through adolescence. The results add to a growing literature on sibling effects as well as theoretical models that emphasize the role of social interaction between siblings as a risk factor for the development of delinquent activity in adolescence. PMID- 11280485 TI - Separation anxiety in parents of adolescents: theoretical significance and scale development. AB - Parents of adolescents commonly face separation-related issues associated with children's increasing independence and imminent leave-taking. The aims of this investigation were (1) to develop a reliable and valid measure of parental emotions associated with separation and (2) to validate the measure by relating it to other attributes (attachment relationship quality, parent-child communication, and parent-adolescent differentiation) assessed in mothers, fathers, and their adolescents. The newly constructed, 35-item Parents of Adolescents Separation Anxiety Scale (PASAS) was administered to 686 parents of teenagers in grades 6, 8, 10, and 12 or college-bound freshmen and seniors. Factor analyses supported formation of two subscales: Anxiety about Adolescent Distancing (AAD) and Comfort with Secure Base Role (CSBR); both subscales showed distinctive patterns of change with child age. Parents' reports indicated that healthy adult attachment styles were associated with lower AAD and higher CSBR scores; children of parents who had higher AAD scores reported lower quality of attachment to both mothers and fathers. PMID- 11280486 TI - Variation in teenage mothers' experiences of child care and other components of welfare reform: selection processes and developmental consequences. AB - Developmental evaluations of the current wave of welfare reform programs present challenges with regard to (1) assessing child outcomes; (2) accounting for heterogeneity among low-income families in both baseline characteristics and involvement in self-sufficiency activities and supports, and (3) development of alternatives to experimental approaches to causal inference. This study (N = 1,079) addresses these challenges by examining effects on 4- to 6-year-old children of different patterns of child care, self-sufficiency activities, and other service utilization indicators among experimental-group mothers in a 16 site welfare reform program. Outcomes in areas of cognitive ability and behavior problems were investigated. The study identified seven subgroups of participants engaging in different patterns of service utilization and activity involvement. A two-stage simultaneous equation methodology was used to account for selection, and effects on child cognitive ability of participation in specific patterns of services and activities were found. For example, children of mothers characterized by high levels of involvement in center-based child care, education, and job training showed higher levels of cognitive ability than children of mothers in groups characterized by high involvement in center-based care and education, or center-based care and job training. In addition, children of mothers in groups with high levels of involvement in any of these activities showed higher levels of cognitive ability than those with low levels of involvement. The bulk of selection effects occurred through site-level differences, rather than family-level socio-economic status or maternal depression indicators. Implications for welfare reform program and policy concerns are discussed. PMID- 11280487 TI - Work-based antipoverty programs for parents can enhance the school performance and social behavior of children. AB - We assess the impact of the New Hope Project, an antipoverty program tested in a random assignment experimental design, on family functioning and developmental outcomes for preschool- and school-aged children (N = 913). New Hope offered wage supplements sufficient to raise family income above the poverty threshold and subsidies for child care and health insurance to adults who worked full-time. New Hope had strong positive effects on boys' academic achievement, classroom behavior skills, positive social behavior, and problem behaviors, as reported by teachers, and on boys' own expectations for advanced education and occupational aspirations. There were not corresponding program effects for girls. The child outcomes may have resulted from a combination of the following: Children in New Hope families spent more time in formal child care programs and other structured activities away from home than did children in control families. New Hope parents were employed more, had more material resources, reported more social support, and expressed less stress and more optimism about achieving their goals than did parents in the control sample. The results suggest that an anti-poverty program that provides support for combining work and family responsibilities can have beneficial effects on the development of school-age children. PMID- 11280488 TI - Processing of rapid auditory stimuli in school-age children referred for evaluation of learning disorders. AB - Tallal hypothesized that reading disabled children have a domain-general deficit in processing rapidly occurring auditory stimuli that degrades speech perception, thereby limiting phonologic awareness and thus reading acquisition. She predicted they would be disproportionately affected by rapidly presented auditory stimuli. In this study, one hundred 7- to 11-year-old children with learning impairment (LI) and 243 non-learning impaired (NLI) children were evaluated on a two-tone auditory discrimination paradigm. LI committed more errors, but effects of timing were comparable. The same result was obtained for a subsample of good and poor readers. Task performance predicted reading, spelling, and calculation. Neural processes underlying perception of speech and other auditory stimuli may be less effective in poor readers; however, contrary to Tallal's hypothesis, rate may not be specifically affected. PMID- 11280489 TI - God's beliefs versus mother's: the development of nonhuman agent concepts. AB - Little research exists on how children understand the actions of nonhuman agents. Researchers often assume that children overgeneralize and attribute human properties such as false beliefs to nonhuman agents. In this study, three experiments were conducted to test this assumption. The experiments used 24 children in New York (aged 2,11-6,11 years), 52 children in Michigan (aged 3,5 6,11 years), and a second group of 45 children in Michigan (3,4-8,5 years) from Christian backgrounds. In the first two experiments, children participated in false-belief tests in which they were asked about human and various nonhuman agents including animals and God. Experiment 3 consisted of a modified perspective-taking task, also including nonhuman agents. The results of the study suggest that children do not consistently use human agent concepts but instead can use different agent concepts for some nonhuman agents like God and special animals. Children are not bound to anthropomorphize, but they often do. PMID- 11280490 TI - Children's judgements of psychological harm in normal and noncanonical situations. AB - This study investigated children's (3-, 5-, and 7-year-olds) and adults' (total N = 92) integration of information about intentions, acts, and outcomes in moral judgments of psychological harm. Behavioral and emotional predictions and judgments of act acceptability and punishment were made under normal and noncanonical causal conditions. Participants at all ages judged it wrong to inflict negative psychological reactions of fear or embarrassment on unwilling participants, even when these reactions were idiosyncratic or noncanonical. When assigning punishment, younger children tended to use an outcome rule, whereas older participants were more likely to use an intention rule or a conjunction rule (if outcome is negative and intention is negative, then punish). The results show that children as young as 3 years are able to take into account other people's idiosyncratic perspectives when making moral judgments of psychological harm. PMID- 11280491 TI - Thinking about the past: early knowledge about links between prior experience, thinking, and emotion. AB - In two studies the authors investigated the situations where 3- to 7-year-olds and adults (N = 152) will connect a person's current feelings to the past, especially to thinking or being reminded about a prior experience. Study 1 presented stories featuring a target character who felt sad, mad, or happy after an event in the past and who many days later felt that same negative or positive emotion upon seeing a cue related to the prior incident. For some story endings, the character's emotion upon seeing the cue matched, or was congruent, with the current situation, whereas for others, the emotion mismatched the present circumstances. Participants were asked to explain the cause of each character's current feelings. As a further comparison, children and adults listened to behavior cuing stories and provided explanations for characters' present actions. Study 2 presented emotional scenarios that varied by emotion-situation fit (whether the character's emotion matched the current situation), person-person fit (whether the character's emotion matched another person's), and past history information (whether information about the character's past was known). Results showed that although there were several significant developments with increasing age, even most 3-year-olds demonstrated some knowledge about connections between past events and present emotions and between thinking and feeling. Indeed, children 5 years and younger revealed strikingly cogent understanding about historical-mental influences in certain situations, especially where they had to explain why a person, who had experienced a negative event in the past, was currently feeling sad or mad in a positive situation. These findings help underwrite a more general account of the development of children's coherent understandings of life history, mind, and emotion. PMID- 11280492 TI - In-depth review effect of androgens on anemia and malnutrition in renal failure: implications for patients on peritoneal dialysis. AB - OBJECTIVE: To analyze the implications of the potential use of androgens in peritoneal dialysis patients, focusing on their effects on hematologic and nutritional parameters. This manuscript reviews the different compounds for clinical use, dosage schedules, adverse effects, and how therapy with androgens might be used to treat anemia and malnutrition in these dialysis patients. DATA SOURCES: Studies in the literature dealing with the effects of androgens on hematologic and nutritional parameters, and their role in uremic anemia and malnutrition. STUDY SELECTION: Studies in which uremic patients received androgens as therapy for anemia or malnutrition. DATA EXTRACTION: Data were abstracted from all of these studies. RESULTS: This review shows that androgens are anabolic substances that also have significant actions on erythropoiesis. A number of clinical studies in uremic patients have found that these compounds have beneficial effects on hematologic parameters and nutritional status, similarly to other therapies, such as recombinant human erythropoietin and recombinant human growth hormone, respectively. CONCLUSIONS: Androgens have been shown to have a beneficial effect on anemia due to renal disease and on nutritional status in uremic patients. Further studies need to be done with larger groups of patients. Objectives for additional research are suggested. PMID- 11280493 TI - Peritoneal dialysis: better than, equal to, or worse than hemodialysis? Data worth knowing before choosing a dialysis modality. AB - Technological advances such as those that allow the delivery of an adequate dialysis dose to a larger percentage of patients, minimization of peritoneal membrane damage with more biocompatible solutions, and lower peritonitis rates will undoubtedly improve retention of patients on peritoneal dialysis (PD) for longer periods. Currently, only 15% of the world dialysis population is managed by PD. Peritoneal dialysis has many advantages over hemodialysis, and if end stage renal disease (ESRD) patients are fully informed about them, the proportion of patients who would prefer this treatment would rise to 25%-30%. An integrated approach to the treatment of ESRD could start with PD in a large percentage of patients, especially those who will receive a kidney transplant within 2 - 3 years. With the present epidemic of ESRD, this approach could lead to a significant saving, relieve the pressure on dialysis units, and allow a larger number of ESRD patients to be treated. PMID- 11280494 TI - Blood pressure and left ventricular hypertrophy in patients on different peritoneal dialysis regimens. AB - OBJECTIVE: To examine the relation between the results of ambulatory 24-hour blood pressure monitoring (ABPM) and left ventricular mass index (LVMI), then to find the independent determinant for left ventricular hypertrophy (LVH) in peritoneal dialysis (PD) patients. Finally, to evaluate the differences in the clinical and cardiovascular characteristics between patients on continuous ambulatory PD (CAPD) and continuous cyclic PD (CCPD). DESIGN: An open, nonrandomized, cross-sectional study. SETTING: Divisions of nephrology and cardiology in a medical center. PATIENTS: Thirty-two uremic patients on maintenance PD therapy (22 patients on CAPD, and 10 on CCPD) without anatomical heart disease or history of receiving long-term hemodialysis. INTERVENTIONS: Home blood pressure (BP) and office BP were measured using the Korotkoff sound technique by sphygmomanometer. ABPM was employed for continuous measurement of BP. Echocardiography was performed for measurement of cardiac parameters and calculation of LVMI. MAIN OUTCOME MEASURES: Multivariate logistic regression analysis was performed for independent determinant of LVH in PD patients. The differences in clinical and cardiovascular characteristics between CAPD and CCPD patients were compared. RESULTS: Simple regression analysis showed positive correlations between LVMI and the duration of hypertension, ambulatory nighttime BP/BP load/BP load > 30%, serum phosphate, calcium-phosphate product, ultrafiltration (UF) volume, and percentage of UF volume during the nighttime. A negative correlation was noted between LVMI and dipping. In multiple regression analysis, the duration of hypertension was the only variable linked to LVMI. In multivariate logistic regression analysis, only ambulatory nighttime systolic BP load > 30% had an independent association with LVH. There were correlations between office/home BP and ambulatory 24-hour BP. In addition, CCPD patients had higher LVMI, UF volume during the nighttime, and percentage of UF volume during the nighttime than those of CAPD patients. CONCLUSIONS: In this study, ambulatory nighttime systolic BP load > 30% had an independent association with LVH. Office and home BP measurements were correlated with ABPM in PD patients. The result that CCPD patients had a higher LVMI than CAPD patients may be due to a relative volume overload during the daytime in CCPD patients. PMID- 11280495 TI - Amadori albumin and advanced glycation end-product formation in peritoneal dialysis using icodextrin. AB - OBJECTIVE: To study the influence of peritoneal dialysis (PD) solutions on the formation of early glycated products and advanced glycation end-products (AGEs). DESIGN AND PATIENTS: The formation of both Amadori albumin and AGEs in glucose- and icodextrin-based PD fluids was analyzed in vitro and in peritoneal effluents of continuous cyclic peritoneal dialysis (CCPD) patients. RESULTS: Albumin incubated with glucose-based PD fluids showed a time- and glucose concentration dependent formation of Amadori albumin and AGEs. Aminoguanidine completely inhibited AGE but not Amadori albumin formation. Albumin incubated in icodextrin resulted in the lowest levels of Amadori albumin and AGE. Amadori albumin levels in effluents of 24 CCPD patients (12 glucose and 12 icodextrin for their daytime dwells) were similar. Dialysate samples collected during a mass transfer area coefficient test in 16 CCPD patients (8 glucose, 8 icodextrin) showed an increase in Amadori albumin formation from baseline (p < 0.0001), without a difference between the groups. In the total group, there was a positive relationship between duration on PD and dialysate Amadori albumin concentration at 240 minutes (p = 0.03). The Amadori albumin dialysate-to-plasma (D/P) ratio at 240 minutes was 0.82+/-0.11, and its clearance amounted to 7.71+/-1.14 mL/min, while the albumin D/P ratio was 0.010+/-0.003 and its clearance was 0.089+/-0.017 mL/min. In a peritoneal biopsy of a CCPD patient, Amadori albumin was observed in the mesothelial layer and the endothelium of the peritoneum. CONCLUSIONS: Using icodextrin-based instead of glucose-based PD fluids can largely reduce the formation of Amadori albumin and AGEs. However, CCPD patients using icodextrin during daytime dwells do not have lower effluent levels of Amadori albumin and AGEs, probably due to the exposure to glucose during their nighttime exchanges. Kinetic studies suggest washout of locally produced Amadori albumin. PMID- 11280496 TI - Preservation of residual renal function in dialysis patients: effects of dialysis technique-related factors. AB - OBJECTIVES: Residual renal function (RRF) is of paramount importance to dialysis adequacy, morbidity, and mortality, particularly for long-term continuous ambulatory peritoneal dialysis (CAPD) patients. Residual renal function seems to be better preserved in patients on CAPD than in hemodialysis (HD) patients. We analyzed RRF in 45 patients with end-stage renal disease (ESRD), commencing either CAPD or HD, to prospectively define the time course of the decline in RRF, and to evaluate dialysis-technique-related factors such as cardiovascular stability and bioincompatibility. STUDY DESIGN: Single-center prospective investigation in parallel design with matched pairs. MATERIALS: Fifteen patients starting CAPD and 15 matched pairs of patients commencing HD were matched according to cause of renal failure and RRF. Hemodialysis patients were assigned to two dialyzer membranes differing markedly in their potential to activate complement and cells (bioincompatibility). Fifteen patients were treated exclusively with the cuprophane membrane (bioincompatible) and the other 15 patients received HD with the high-flux polysulfone membrane (biocompatible). MEASUREMENTS: Residual renal function was determined at initiation of dialytic therapy and after 6, 12, and 24 months. Dry weight (by chest x ray and diameter of the vena cava) was closely recorded throughout the study, and the number of hypotensive episodes counted. RESULTS: Residual renal function declined in both CAPD and HD patients, although this decline was faster in HD patients (2.8 mL/minute after 6 months and 3.7 mL/min after 12 months) than in CAPD patients (0.6 mL/min and 1.4 mL/min after 6 and 12 months respectively). It declined faster in patients with bioincompatible than with biocompatible HD membranes (3.6 mL/min vs 1.9 mL/min after 6 months). Eleven percent of the HD sessions were complicated by clinically relevant blood pressure reductions, but there were no differences between the two dialyzer membrane groups. None of the CAPD patients had documented hypotensive episodes. None of the study patients suffered severe illness or received nephrotoxic antibiotics or radiocontrast media. CONCLUSIONS: The better preservation of RRF in stable CAPD patients corresponded with greater cardiovascular stability compared to HD patients, independently of the membrane used. Furthermore, there was a significantly higher preservation of RRF in HD patients on polysulfone versus cuprophane membranes, indicating an additional effect of biocompatibility, such as less generation of nephrotoxic substances by the membrane. Thus, starting ESRD patients on HD prior to elective CAPD should be avoided for better preservation of RRF. PMID- 11280497 TI - Effect of fluid supplementation and modality on peritoneal permeability characteristics in a rat peritoneal dialysis model. AB - OBJECTIVE: Hemoconcentration may influence peritoneal permeability parameters in anesthetized animals without fluid supplementation. Therefore, the aim of this study was to investigate the effects of fluid supplementation on peritoneal permeability in an acute peritoneal dialysis model in anesthetized rats. DESIGN: To study the effect of fluid supplementation on peritoneal permeability characteristics, 24 anesthetized male Wistar rats were investigated in 3 groups during a 4-hour standardized peritoneal permeability analysis with a 3.86% glucose dialysis solution (SPARa). The groups included a control group with no fluid supplementation (None, n = 8), a group with continuous subcutaneous infusion of 0.9% NaCl 3 mL/hr (SC, n = 9), and a group with continuous intravenous infusion of 0.9% NaCl 3 mL/hr (IV, n = 7). Inflow, sampling, and outflow of the dialysate during the SPARa occurred via a cannula inserted intraperitoneally in the lower left quadrant of the abdomen. Blood was drawn at the end of the dwell. Baseline blood samples were obtained from six separate untreated rats. RESULTS: Plasma osmolality was significantly lower in the IV group (334+/-1.4 mOsm/kg) compared to the None group (348+/-0.7 mOsm/kg, p < 0.01), and not different from the SC group (335+/-6.4 mOsm/kg), but higher than baseline (314+/-5.3 mOsm/kg, p < 0.001). Urine production during the dwell was not different among the groups: None 10.6+/-5.3 mL; SC 8.0+/-6.0 mL; and IV 10.5+/-5.6 mL. Transcapillary ultrafiltration after 4 hours was significantly higher in the IV group (p < 0.05) compared to the other two groups. Net ultrafiltration and effective lymphatic absorption were similar in all groups. Mass transfer area coefficient of urea (MTACurea) was significantly greater in the IV group (155+/-23.2 microL/minute, p < 0.003), but not different between the None (118+/-16.2 microL/min) and SC (123+/-25.9 microL/min) groups. Correcting these for the baseline plasma concentration resulted in higher values, but the IV data remained greater than the SC and None groups (p < 0.01). The glucose absorption, albumin, and IgG clearances and the sieving of sodium were alike in all groups. CONCLUSION: It can be concluded that IV fluid supplementation is more effective in preventing dehydration than SC supplementation, and it enhanced some peritoneal permeability characteristics in anesthetized rats during a 4-hour investigation. It is therefore important to standardize fluid supplementation in experiments with anesthetized animals. PMID- 11280498 TI - Fluid and solute transport using different sodium concentrations in peritoneal dialysis solutions. AB - BACKGROUND: Fluid and sodium balance is important for the success of long-term peritoneal dialysis. Convective transport is the major determinant for sodium removal during peritoneal dialysis using conventional dialysis solutions. However, recent studies showed that lower sodium concentration in dialysate could significantly increase sodium removal by increasing the diffusion gradient, thereby increasing diffusive transport. In the present study, we investigated the influence of the sodium concentration gradient on the diffusive transport coefficient, K(BD) for sodium. METHODS: A 4-hour dwell study was done in Sprague Dawley rats using 25 mL 5% glucose (NS), 5% glucose + 0.3% NaCl (LS), 5% glucose + 0.6% NaCl (MS), or 5% glucose + 0.9% NaCl (HS), with frequent dialysate and blood sampling. Radiolabeled human albumin (RISA) was added to the solution as an intraperitoneal volume marker. The peritoneal fluid and sodium transport characteristics were evaluated. RESULTS: Significant ultrafiltration (both net ultrafiltration and transcapillary ultrafiltration) was observed in each group despite the osmolality of the 5% glucose solution being slightly lower than the plasma osmolality. There was no difference in peritoneal fluid absorption rate and direct lymphatic absorption among the four groups. With the sieving coefficient for sodium set to 0.55, a significantly higher K(BD) for sodium was found in the NS compared to the HS group. The K(BD) for sodium was 0.21+/-0.01, 0.20+/-0.01, 0.17+/-0.01, and 0.09+/-0.01 mL/min for the NS, LS, MS, and HS groups, respectively. The K(BD) values for glucose were significantly lower in the NS and LS groups compared to the MS and HS groups. CONCLUSIONS: Our results suggest that (1) sodium concentration may affect peritoneal sodium K(BD)--as the sodium concentration gradient increased, the K(BD) decreased; (2) 5% glucose solution could induce significant peritoneal ultrafiltration in normal rats despite its initial hypo-osmotic nature, this was due to the significantly lower glucose transport rate than sodium transport rate; and (3) a lower dialysate sodium concentration may decrease peritoneal glucose absorption. PMID- 11280499 TI - A physiological analysis of hyponatremia: implications for patients on peritoneal dialysis. AB - The basis for hyponatremia is a negative balance for sodium (Na+) plus potassium (K+) and/or a positive balance for water. In patients with normal renal function, vasopressin is needed to prevent the excretion of electrolyte-free water. Vasopressin is not important when there is little residual renal function. If hyponatremia is accompanied by a quantitatively appropriate gain in weight, this implies that a gain of electrolyte-free water was the basis for hyponatremia. In the absence of this weight gain, a loss of salts is to be suspected. If the extracellular fluid (ECF) volume is obviously low, hyponatremia is due to a deficit of NaCl, unless there is a deficit of K+. With a KCl deficit and a contracted ECF volume, there should also be a large shift of Na+ into cells, so metabolic alkalosis would not be an expected finding. In contrast, those patients with no change in weight who have a normal or expanded ECF volume are subdivided into those with a gain of solutes restricted to the ECF compartment (glucose, mannitol), or those with a deficit of solutes of intracellular fluid origin, which implies that a catabolic state (malnutrition) may be present. PMID- 11280500 TI - Hyponatremia in patients undergoing CAPD: role of water gain and/or malnutrition. AB - BACKGROUND: Hyponatremia has a number of different causes; some may have serious untoward implications for patients undergoing chronic ambulatory peritoneal dialysis (CAPD). OBJECTIVE: To determine the pathophysiology of hyponatremia in patients on CAPD. METHODS: A retrospective analysis was carried out on 210 patients on CAPD. We selected patients with 2-4 consecutive periods when the plasma sodium concentration was < or =130 mmol/L and again when it was > 133 mmol/L. Exclusion criteria included hyperglycemia, orthostatic hypotension, edema, and inadequate records. RESULTS: An electrolyte-free water gain appeared to be the main cause of hyponatremia in only 1 of 5 patients because this was the only patient with a significant increase in body weight. In 1 patient, there was weight loss in the hyponatremic period, suggesting tissue catabolism was present. In 3 patients, there was neither weight gain nor evidence for a contracted extracellular fluid volume in the hyponatremic period, suggesting that intracellular potassium and phosphate loss could be the major mechanism for their hyponatremia. CONCLUSION: When hyponatremia is due to a catabolic state, its management should aim to restore intracellular fluid composition (i.e., to correct malnutrition). PMID- 11280501 TI - CAPD: a control strategy to prevent spread of HCV infection in end-stage renal disease. PMID- 11280502 TI - A critical pathway for outpatient treatment of CAPD peritonitis. PMID- 11280503 TI - Effects of intraperitoneal cefazolin on mesothelial cells in noninfected CAPD patients. PMID- 11280504 TI - Peritoneal dialysis fluids induce the stress response in human mesothelial cells. PMID- 11280505 TI - Gastrointestinal motor function in children treated with peritoneal dialysis. PMID- 11280506 TI - Adsorption of erythropoietin and growth hormone to peritoneal dialysis bags and tubing. AB - OBJECTIVE: To study the adsorption of erythropoietin and growth hormone to dialysis bags and tubing. DESIGN: In vitro study in which radiolabeled erythropoietin and recombinant human growth hormone were added to small-volume (50- and 250-mL) dialysis bags. Recovery was measured after 15-minute dwells. Experiments were performed in triplicate. SETTING: University hospital. RESULTS: Adsorption of erythropoietin and growth hormone was less than 7%. CONCLUSION: Adsorption of erythropoietin and recombinant human growth hormone to dialysis bags and tubing is minimal. This finding provides another argument in favor of intraperitoneal therapy in pediatric peritoneal dialysis. PMID- 11280507 TI - Peritoneal metallothionein content in patients with end-stage renal disease on or not on peritoneal dialysis. PMID- 11280508 TI - Does a face mask prevent peritonitis? PMID- 11280509 TI - Wangiella dermatitidis peritonitis in a CAPD patient. PMID- 11280511 TI - Research & development methodology for recycling residues as building materials- a proposal. AB - This article presents a proposal of methodology for conducting such research and development. The data/statistics waste collection statistics phase must cover geographical distribution, seasonal variations on production rates, waste management practices, current applications and their related costs and revenues. Waste characterisation must be comprehensive with physical, environmental and chemical aspects, including waste variability and waste contamination from shipping, handling and storage activities. Based on the previous results a broad forecast of potential applications must be developed based on very simple rules like minimisation of transportation distances and energy consumption, etc. Marketing evaluation is a very important step, frequently neglected when choosing the best applications for a particular waste. Other steps are product development and performance evaluation. Environmental evaluation of the new technology is very important because not all recycling is environmentally sound. This evaluation must be based on the life cycle assessment (LCA) and has to consider the environmental benefit of avoiding landfill disposal of the waste, and could include leaching or other specific tests or simulations. Also, the technological transference phase must be carefully planned and developed. Each proposed step is discussed, examples are given and needs for further research emphasised. PMID- 11280510 TI - Bacillus licheniformis peritonitis in a CAPD patient. PMID- 11280512 TI - Leaching behaviour of synthetic aggregates. AB - In the framework of EU project "Utilising innovative kiln technology to recycle waste into synthetic aggregate" (BRST-CT98-5234), the leaching behaviour of synthetic aggregates has been studied to assess its environmental compatibility in the various stages of its use. Since the conditions are very different for the different uses, the assessment calls for a variety of different leaching conditions. The pH dependence test is used to cover important differences in pH environment to which the materials are exposed to as well as for an assessment of the buffering capacity of the material. Synthetic aggregate features a low buffer capacity, which makes it sensitive to externally imposed pH conditions. Utilisation and storage exposed to acidic conditions needs to be avoided. The results of the pH dependence test and column leaching test are mutually consistent. The CEN TC 154 method appears to provide systematically low values due to the arbitrary selection of test conditions. Synthetic aggregate studied to date will not adversely affect the concrete in its service life. The main issue for aggregate use is the recycling and the "end of life" condition, when the material becomes construction debris. Not metals, but oxyanions, such as Cr VI and Mo are most relevant under these conditions. A concise test has been applied to assess crucial aspects of leaching for different production mixes. PMID- 11280513 TI - Behaviour of cement-treated MSWI bottom ash. AB - MSWI bottom ash is the residue of combustion. The use of bottom ash in road construction is wide spread. French legislation forbids the disposal of resuable waste in special landfill from 2002. Moreover, "arrete du 9 mai 1994" provides environmental criteria (leaching threshold, etc.), and evaluates this material according to utilisation in road construction. In such application, bottom ash is often treated with binder to improve its mechanical features. Nevertheless, bottom ash is subject to chemical problems. These problems induce an expansion which brings about cracking and finally road destruction. Therefore, it is necessary to estimate the swelling potential of MSWI bottom ash prior utilisation. This is one of the aims of the European contract "Mashroad" (contract BRST CT97-5150). This study involved 4 years of work on cement-treated MSWI bottom ash. It examined different tests that show the importance of oxidation of aluminium in the swelling reaction and the efficiency of different treatments. Different binders were used in order to have different proportions of clinker. The kinetic aspects of aluminium-binder reaction were also studied. Finally, we present a special cell to measure the swelling pressure of these materials is presented. PMID- 11280514 TI - Self-cementing properties of crushed demolished concrete in unbound layers: results from triaxial tests and field tests. AB - A 2-year study is underway to evaluate the expected growth in stiffness in layers of crushed concrete from demolished structures. This growth is said to be a result of self-cementing properties. The study consists of repeated load triaxial tests on manufactured specimens after different storing time together with falling weight deflectometer, FWD, measurements on test sections. Results so far show a clear increase with time in resilient modulus and in back-calculated layer modulus for all concrete materials. The increase is the largest in the first months and then diminishes. The field measurements show a more considerable growth in stiffness than the laboratory tests, with a doubled value two years after construction. Comparative investigations on natural aggregates, mostly crushed granite do not show any growth in stiffness, neither in the laboratory nor in the field. Consequences for the choice of design modulus are discussed. PMID- 11280515 TI - Synthetic aggregates from combustion ashes using an innovative rotary kiln. AB - This paper describes the use of a number of different combustion ashes to manufacture synthetic aggregates using an innovative rotary 'Trefoil' kiln. Three types of combustion ash were used, namely: incinerated sewage sludge ash (ISSA); municipal solid waste incinerator bottom ash (MSWIBA-- referred to here as BA); and pulverised fuel ash (Pfa). The fine waste ash fractions listed above were combined with a binder to create a plastic mix that was capable of being formed into 'green pellets'. These pellets were then fired in a Trefoil kiln to sinter the ashes into hard fused aggregates that were then tested for use as a replacement for the natural coarse aggregate in concrete. Results up to 28 days showed that these synthetic aggregates were capable of producing concretes with compressive strengths ranging from 33 to 51 MPa, equivalent to between 73 and 112% of that of the control concrete made with natural aggregates. PMID- 11280516 TI - Immobilisation of PAH in waste materials. AB - This paper gives an overview of the results of a research project into the possibilities of immobilising polycyclic aromatic hydrocarbons (PAH), that are present in waste materials. The results show that with hydraulic binders the waste materials can be solidified. The PAH do still leach to a relatively high extent. However, this PAH leaching can be decreased by more than a factor 10 by means of the addition of a specific additive. The immobilisation product fulfils technological requirements for the use as a road base construction material, such as compressive strength. PMID- 11280517 TI - Environmental impact of ferrochrome slag in road construction. AB - Vargon Alloys in Western Sweden is one of the largest producers of ferrochrome slag in Europe. Ferrochrome slag is a by-product from the production of ferrochrome, an essential component in stainless steel. Extensive tests have been carried out on the physical properties of the ferrochrome slag from Vargon Alloys and it was found to be highly suitable as road construction material. The composition and leaching tests of the ferrochrome slag show that the chromium content is high, 1-3%, although leaching under normal conditions is very low. With the exception of potassium (K), which had a potential leaching capacity (availability test) of around 16%, the leaching of chromium, nickel, zinc and other elements was just a few per cent. However, all these tests were conducted in the laboratory. What happens out in the field, under the influence of acid rain and biological activity, and how does this compare with the laboratory results? To answer this question an investigation was carried out to study the environmental impact of ferrochrome slag in roads that were built in 1994. The investigation includes soil sampling (total content and leachable amounts of metals) and groundwater analysis (filtered and non-filtered samples). In addition, a new method involving the bio-uptake of chromium and other metals by the roots of the dandelion (Taraxacum officinale) was tested. The results show that there was a low migration of particles from the slag to the underlying soil and that the leaching into the groundwater was also low for all the elements analysed. However, there seemed to be a significant uptake of Cr by plants growing with their roots in the slag. An investigation of plant uptake was an important complement to laboratory leaching tests on alternative materials. PMID- 11280518 TI - Leaching behaviour of a chromium smelter waste heap. AB - This paper reports the results of geochemical sampling and modelling of leachates from a chromite ore processing residue (C.O.P.R.) pile under rainwater infiltration. The waste pile is located in the north of England and consists of 800,000 m3 of waste. The pH of fresh leachate is similar to that of a solution in equilibrium with portlandite Ca(OH)2, which is a major constituent of the waste. The in-gassing of CO2(g) causes the pH of the leachates to drop along the drainage ditch and calcite precipitation to occur. The extent of in-gassing is dependent upon the flow rate within the drainage ditch. The dissolution of solid solutions containing residual chromate is likely to control chromate concentrations within the leachate. PMID- 11280519 TI - Life-cycle impacts of the use of industrial by-products in road and earth construction. AB - A two-stage study "Life cycle analysis of road construction and earthworks" was part of a more extensive Finnish research project "Assessment of the applicability of secondary products in earthworks". In the first stage of this work a life-cycle impact assessment procedure for the comparison and evaluation of alternative road and earth constructions was developed. Additionally, a database containing the environmental burdens of the most significant construction materials and unit operations was constructed. In order to evaluate the applicability of the procedure, the use of coal ash, crushed concrete waste and granulated blast-furnace slag in road construction was evaluated in case studies. The use of these secondary products was also compared with the use of natural materials in corresponding applications. The aim of the second stage was to transfer the assembled data for utilisation as a practical model by creating an inventory analysis program to calculate and compare the life cycle impacts of the most common road constructions and foundation engineering methods. The data obtained in the first stage was also augmented to the extent necessary for this purpose. The results of case studies indicate that the production and transport of the materials used in road constructions produce the most significant environmental burdens. Production of the bitumen and cement, crushing of materials and transport of materials are the most energy consuming single life cycle stages of the construction. A large part of the emissions to atmosphere originates from energy production. In the expert assessment, consumption of natural materials and leaching behaviour were also regarded as being of great significance. PMID- 11280520 TI - Modelling the effects of waste components on cement hydration. AB - Ordinary Portland Cement (OPC) is often used for the solidification/stabilization (S/S) of waste containing heavy metals and salts. These waste components will precipitate in the form of insoluble compounds on to unreacted cement clinker grains preventing further hydration. In this study the long term effects of the presence of contaminants in solidified waste is examined by numerically simulating cement hydration after precipitation of metal salts on the surface of cement grains. A cement hydration model was extended in order to describe pore water composition and the effects of cement grain coating. Calculations were made and the strength development predicted by the model was found to agree qualitatively with experimental results found in literature. The complete model is useful in predicting the strength and leaching resistance of solidified products and developing solidification recipes based on cement. PMID- 11280521 TI - Products of steel slags an opportunity to save natural resources. AB - In Germany, and in the most industrial countries, the use of blast furnace and steel slags as an aggregate for civil engineering, for metallurgical use and as fertiliser has a very long tradition. Since the introduction of the basic oxygen steel making furnace (BOF) process and the electric arc furnace (EAF) process the German steel industry started extensive research on the development of fields of application for BOF and EAF slags. These investigations have been mainly performed by Forschungsgemeinschaft Eisenhuttenschlacken e. V. (FEhS), the Research Association for blast furnace and steel slags. Today steel slags are well characterised and long-term experienced materials mainly used as aggregates for road construction (e.g. asphaltic or unbound layers), as armour-stones for hydraulic engineering constructions (e.g. stabilisation of shores), and as fertiliser for agriculture purposes. These multifarious fields of application could only be achieved because the steelworks influence the quality of slags by a careful selection of raw materials and a suitable process route. Furthermore, subsequent procedures like a treatment of the liquid slag, an appropriate heat treatment and a suitable processing have been developed to ensure that the quality of steel slags is always adequate for the end use. Depending on the respective field of application, the suitability of steel slags has to be proven by determining the technical properties, as well as the environmental compatibility. For this reason test methods have been developed to evaluate the technical properties especially the volume stability and the environmental behaviour. To evaluate the volume stability a suitable test (steam test) has been developed and the results from laboratory tests were compared with the behaviour of steel slags under practical conditions, e.g. in a road. To determine the environmental behaviour leaching tests have been developed. In the meanwhile most of these test methods are drafted or already accepted as a CEN standard and are used for a continuous quality control. Usually the suitability of steel slags is stated by fulfilling the requirements of national and/or international standards and regulations. Based on these standards and regulations in Germany in 1998 about 97% of the produced steel slags have been used as aggregates for road construction (e.g. as surface layer, road base and sub base for high trafficked roads), ways, earthworks, and armourstones for hydraulic structures. Consistent to the successful long-term experience not only products of steel slags but also products of blast furnace slags have been eliminated from the European Waste Catalogue and the European Shipment of Waste Regulation of the European Community, as well as from the lists of OECD for transfrontier movements by the decision of the OECD-Council from 21 September, 1995. PMID- 11280522 TI - The building materials decree: an example of a Dutch regulation based on the potential impact of materials on the environment. AB - Some 20 years ago Dutch society became very aware of the problem of the increasing production of waste materials and of the need to find solutions. Technical solutions had to be developed and introduced into society. Due to the discovery of many soil pollution problems and other environmental calamities, people asked for clear criteria on which proper and safe solutions could be based. Therefore a large number of environmental aspects had to be taken into account in the developing field of environmental policy. The Dutch building materials decree is one of the steps on the road towards a sustainable society. The decree is based on the soil protection act and the surface water protection act. The decree gives quality criteria for the application and re-use of stony materials and earth used as building materials. No difference is made between primary materials, secondary materials and waste materials. The decree is applicable in case these materials are used in constructions where they are in contact with rain, surface water and ground water (e.g. in embankments, road building, outside walls of buildings, foundations and roofs). For implementation of the decree in the construction industry it was necessary to develop standards, methods for testing and certification schemes. In order to guarantee independent testing and judgement, laboratories had to meet quality standards put forward in an accreditation scheme. Although this legislation does not cover all environmental aspects, it has proved to be an important element in judging the environmental quality of construction materials in a direct or indirect way, and a contribution to the management of waste materials. The question of clear and fair criteria was easily put forward. Answering this question, however, turned out to be a complex task in which a great number of aspects had to be considered and a great many tools had to be developed, checked and thoroughly discussed. In some cases practical choices had to be made in order to prevent an unclear situation continuing too long. After an introduction period of three years, the decree came into full operation on 1 July 1999. PMID- 11280523 TI - Screening for varices in patients with cirrhosis: where do we stand? PMID- 11280524 TI - Barrett's esophagus: from process to outcomes. PMID- 11280525 TI - Progress in treatment for Crohn's disease. PMID- 11280526 TI - Incidental gallbladder cancer. PMID- 11280527 TI - Gastroenterology in the new millennium: where are we and where are we going? PMID- 11280528 TI - Management of Crohn's disease in adults. PMID- 11280529 TI - Composition of the postprandial refluxate in patients with gastroesophageal reflux disease. AB - OBJECTIVE: It is not known whether the characteristics of the postprandial refluxate in patients with gastroesophageal reflux disease (GERD) differ from those observed in normal subjects. The aim of this study was to characterize the postprandial refluxate in adult patients with GERD using combined intraluminal electrical impedance and pH measurements. METHODS: Postprandial gastroesophageal reflux was assessed in 16 patients with GERD and 15 controls. pH and intraluminal electrical impedance were used to identify acid and nonacid reflux of liquid, mixed (liquid + gas) or gas. RESULTS: Transient lower esophageal sphincter relaxations (TLESRs) and reflux of gastric contents were equally frequent in both groups. However, patients with GERD had more acid reflux [8 (4.7-10.5)/h vs 3.5 (2.6-6)/h, p < 0.05], and normal subjects had more nonacid reflux [5 (4.3-6.7)/h is 3 (1-3.5)/h, p < 0.05]. Gas reflux was less frequent in GERD than in controls (51% vs 68%; p < 0.05). Pure liquid reflux, however, was more frequent (40% vs 26%, p < 0.05) and twice as likely to be acid in GERD. During TLESRs, liquid acid reflux was more frequent in GERD than in controls. CONCLUSIONS: TLESRs and reflux of gastric contents are similarly frequent in patients with GERD and controls. However, patients with GERD have more acid reflux and less nonacid reflux. Differences in the air-liquid composition of the refluxate may contribute to the higher rate of acid reflux observed in these patients. PMID- 11280530 TI - Efficacy and safety of esomeprazole compared with omeprazole in GERD patients with erosive esophagitis: a randomized controlled trial. AB - OBJECTIVE: In patients with gastroesophageal reflux disease (GERD), esomeprazole, the S-isomer of omeprazole, has demonstrated pharmacological and clinical benefits beyond those seen with the racemic parent compound. This study was designed to further evaluate the efficacy and tolerability of esomeprazole relative to that of omeprazole in healing erosive esophagitis and resolving accompanying symptoms of GERD. METHODS: Esomeprazole 40 mg was compared with omeprazole 20 mg once daily in 2425 patients with erosive esophagitis (Helicobacter pylori negative by serology) in an 8-wk, multicenter, randomized, double-blind, parallel-group study conducted in 163 centers throughout the US. The primary efficacy endpoint was the proportion of patients with healed esophagitis at wk 8. Secondary endpoints were the proportion of patients healed at wk 4, resolution of heartburn at wk 4, time to first resolution and sustained resolution of heartburn, and proportion of heartburn-free days and nights. Safety and tolerability were also assessed. RESULTS: Significantly more patients were healed with esomeprazole versus omeprazole at wk 8 (93.7% vs 84.2%, p < 0.001; life table estimates, intention-to-treat analysis). Healing rates at wk 4 were 81.7% and 68.7%, respectively. Esomeprazole was superior to omeprazole for all secondary measures and had a similar safety profile. The most common adverse events in both treatment groups were headache, diarrhea, and nausea. CONCLUSIONS: Esomeprazole demonstrates significantly greater efficacy than omeprazole in the treatment of GERD patients with erosive esophagitis. The tolerability and safety of esomeprazole are comparable to that of omeprazole. (Am PMID- 11280531 TI - Geographic differences in the distribution of intestinal metaplasia in duodenal ulcer patients. AB - OBJECTIVE: A strong correlation exists between atrophic gastritis and the intestinal type of gastric carcinoma. Duodenal ulcer disease characteristically has an antral predominant gastritis and a lower risk for gastric cancer. The aim of this study was to investigate the extent and distribution of intestinal metaplasia in duodenal ulcer in countries differing in gastric cancer incidence. METHODS: Topographically mapped gastric biopsy specimens (median 11) were obtained from patients with duodenal ulcer in four countries (Korea, Colombia, USA, and South Africa). Sections were stained with a triple stain and evaluated for Helicobacter pylori (H. pylori), active inflammation, and intestinal metaplasia. RESULTS: One hundred and sixty-five patients with duodenal ulcer were examined (29 from Korea, 52 from Colombia, 62 from the USA, and 22 from South Africa). The percentage of biopsies with intestinal metaplasia was significantly greater in Korean patients (86%) compared with that in other countries (50%) (p = 0.0004). Intestinal metaplasia was most prevalent in the antrum lesser curve and greater curve, and the body lesser curve. Intestinal metaplasia was present in the gastric corpus of 38% of duodenal ulcer patients from Korea compared with an average of 10% elsewhere (p = 0.018). No differences were observed in the density or distribution of H. pylori infection or in the degree of active gastritis between countries. CONCLUSIONS: Although antral predominant gastritis is the prevalent pattern of gastritis in duodenal ulcer, intestinal metaplasia in the gastric corpus may be found with geographic differences. These findings suggest that duodenal ulcer and gastric cancer are not mutually exclusive diseases but are rather ends of the spectrum of H. pylori infection. PMID- 11280532 TI - Self-reported effectiveness and physician consultation rate in users of over-the counter histamine-2 receptor antagonists. AB - OBJECTIVE: Decreased physician visits for dyspepsia were predicted with the histamine-2 receptor antagonists (H2RA) release to over-the-counter (OTC) status. The aim of this study was to examine the presentation frequency for dyspeptic complaints before and after the OTC release of the H2RA and the self-reported effectiveness of OTC H2RA. METHODS: Two cross-sectional surveys were used in a community sample. The patients comprised a random age- and sex-stratified sample of 1600 ambulatory adults in 1993 and 1800 in 1997. Self-report, valid mail surveys gathered information on healthcare seeking and gastrointestinal symptoms in 1993 and 1997 and antisecretory use in 1997. RESULTS: Presentation frequency for dyspepsia was 22% in 1993 versus 23.5% in 1997. Only 16% of chronic users of the OTC H2RA obtained complete relief of symptomatic episodes. Use of an OTC H2RA was highly associated with presentation to a physician in the past year. CONCLUSIONS: OTC H2RA infrequently provided the complete relief desired by patients. Presentation frequency to physicians for dyspeptic complaints did not change with availability of H2RA OTC. PMID- 11280533 TI - Evaluation of the Helicobacter pylori stool antigen test (HpSA) for detection of Helicobacter pylori infection in children. AB - OBJECTIVE: Helicobacter pylori (H. pylori) infection is usually acquired in early childhood. Noninvasive methods for detection of H. pylori infection are required to study its incidence, transmission, and clearance. They should be easy to perform, inexpensive, and have a high diagnostic accuracy, especially in infants and toddlers. Both serology and the 13C-urea breath test (13C-UBT) do not fulfill all these requirements. The aim of this study was to evaluate a new enzyme immunoassay for detection of H. pylori antigen in stool (Premier Platinum HpSA, Meridian Diagnostics, Cincinnati, OH) in a large cohort of children and to compare it to invasive techniques and the 13C-UBT. METHODS: HpSA was performed in 310 stool samples of 274 children divided into three groups. Group A consisted of 145 children and adolescents (0.5-19.8 yr, 66/145 <6 yr) who underwent upper endoscopy for various gastrointestinal symptoms. H. pylori status was defined by histology, culture, and rapid urease test from biopsies of the antrum and corpus. A 13C-UBT was performed in 133 of 145 children. Group B consisted of 22 patients (5.7-16.1 yr) who were retested with both noninvasive tests 8 wk after anti-H. pylori triple therapy. Group C consisted of 129 healthy infants and toddlers (0.9 3.1 yr) who were tested with the 13C-UBT. Children with discrepant or positive test results were retested after 2 and 12 months. Results of the HpSA were read at 450/620 nm by spectrophotometry. An optical density <0.100 was defined as negative, >0.120 as positive, and values between 0.100 and 0.120 were considered as equivocal. RESULTS: In Group A, the HpSA gave false-negative results in five of 45 infected children and false-positive results in four of 100 noninfected children, whereas four patients (2.8%) showed equivocal results. In both infected and noninfected children, no relation between the optical density values and age was found. The 13C-UBT was correct in 132 of 133 children tested. In Group B, there was complete concordance between the HpSA and 13C-UBT: 19 children tested negative and three positive. In Group C, concordant results between the two noninvasive methods were found in 124 of 129 (96%) toddlers (122 negative and two positive). Retesting of five children with discrepant results revealed that, on initial testing, the HpSA was incorrect in two (one false-positive, one false negative), and the 13C-UBT was incorrect in three (always false-positive). CONCLUSIONS: In symptomatic children, the HpSA revealed a sensitivity of 88.9% (95% CI 77.3-96.3) and a specificity of 94.0% (88.1-97.7) compared to the 13C UBT, 100% (94.0-100) and 98.9% (94.7-100), respectively. However, in healthy toddlers, the HpSA performed as well as the 13C-UBT with excellent concordance between the two noninvasive tests. There was no age dependency of the stool test results, and changing the cutoff would not have improved accuracy. Thus, the HpSA test seems suitable to monitor the success of anti-H. pylori therapy. PMID- 11280534 TI - Similarities in cyclic vomiting syndrome across age groups. AB - OBJECTIVE: Cyclic vomiting syndrome is well recognized in children yet has poorly defined pathogenesis and treatment. Cyclic vomiting syndrome is occasionally diagnosed in older subjects, but little attempt has been made to determine if such cases represent a unique disorder. METHODS: We reviewed clinical data from 39 patients aged 1.8-75 yr with cyclic vomiting syndrome meeting published criteria for diagnosis. Clinical characteristics were compared between subjects with symptom onset in childhood (<12 yr, n = 18) and subjects with onset at an older age (> or =12 yr, n = 21; mean age at onset 34.8+/-3.8 yr). RESULTS: All patients had stereotypical episodes of vomiting separated by varying symptom-free intervals. The prevalence rates of prodromal symptoms, triggering events, alleviants, associated symptoms including abdominal pain and diarrhea, and past or family history of migraine were similar in the children and older subjects with the syndrome (p > 0.3 for each). Delay in diagnosis was greater in the older subset (3.1+/-0.8 yr vs 7.9+/-3.1 yr, p < 0.05). Interepisode intervals and total number of hospitalizations did not differ significantly between younger and older patients, but duration of episodes was significantly longer in the older group (2.0+/-0.5 days vs 3.8+/-0.4 days, p < 0.01). When subjects were further substratified by age of illness onset, duration of episodes progressively increased from infant/toddlers (1.8+/-0.4 days) through childhood (2.3+/-0.5 days) and adolescence (2.9+/-1.0 days) and into adulthood (3.9+/-0.5; p < 0.05 across groups). Episode duration did not lengthen further in subgroups >20 yr of age. CONCLUSIONS: Many characteristics of cyclic vomiting syndrome are similar irrespective of age at disorder onset, suggesting a uniform pathogenesis. Duration of episodes increases with age to age 20 yr. Increased awareness of the condition and a high index of suspicion may help decrease delay in diagnosis after symptom onset. PMID- 11280535 TI - Efficacy of cisapride and domperidone in functional (nonulcer) dyspepsia: a meta analysis. AB - BACKGROUND: The efficacy of prokinetic agents in functional (nonulcer) dyspepsia has been questioned based on recent trial results. We performed a meta-analysis to determine the efficacy of cisapride and domperidone in functional dyspepsia. METHODS: Computer and manual searching was used to identify placebo-controlled studies that included >20 patients. The statistical analysis focused on: global assessment by the investigator, epigastric pain, early satiety, abdominal distension and nausea (all rated on four-point scales). Results are reported as odds ratios (OR) in favor of treatment. Regression analysis was performed to evaluate possible effect modifiers. The relationship between improvement in gastric emptying and symptoms was also evaluated. RESULTS: For cisapride, 17 studies met the inclusion criteria, but varying numbers of studies had to be used for the different outcome measures. For all outcome measures, there was a statiscally significant benefit in favor of cisapride: global assessment of improvement by the investigator or patient (OR 2.9, 95% CI 1.5-5.8), epigastric pain (OR 0.19, 95% CI 0.05-0.7), early satiety (OR 0.18, 95% CI 0.9-0.4), abdominal distension (OR 0.32, 95% CI 0.1-0.7), and nausea (OR 0.26, 95% CI 0.1 0.5). Age of patient, year of publication, and country where study was performed had only small modifying effects. There were insufficient data to determine whether there is a relationship between improvement in gastric emptying and response to treatment. For domperidone, four of eight studies could be used for the analysis of global assessment of improvement by the investigator. This showed an OR of 7.0 (95% CI 3.6-16) in favor of domperidone. CONCLUSIONS: Both cisapride and domperidone seem to be efficacious in functional dyspepsia, although this conclusion is largely based on global assessment by the investigator, which may not be an optimal outcome measure. PMID- 11280536 TI - Serum amylase and lipase activities in normal pregnancy: a prospective case control study. AB - OBJECTIVE: The diagnosis of acute pancreatitis during pregnancy is usually based on the association of upper abdominal pain, nausea or vomiting, and elevated serum amylase or lipase activities. The changes in these enzymatic activities have not been clearly established during normal pregnancy. The aim of this study was therefore to evaluate serum amylase and lipase activities in healthy pregnant women. METHODS: Serum amylase and lipase activities were measured in 103 pregnant women (first trimester, n = 34; second trimester, n = 36; third trimester, n = 33) and in 103 nonpregnant women matched for age and not receiving oral contraception. RESULTS: Serum amylase activity was similar in pregnant women and nonpregnant women during all trimesters of pregnancy. Serum lipase activity was significantly lower during the first trimester of pregnancy compared to nonpregnant women (48.6+/-27.6 vs 59.2+/-29.3 IU/L, p < 0.05) and compared to the third trimester (48.6+/-27.6 vs 76.3+/-35.8 IU/L, p < 0.001). Serum lipase activity was not statistically different between pregnant and nonpregnant women during the second and third trimesters. CONCLUSION: An increase in serum amylase and lipase activities during pregnancy should be taken into account, as in nonpregnant women. PMID- 11280537 TI - Early detection of pancreatic cancer in patients with chronic pancreatitis: diagnostic utility of a K-ras point mutation in the pancreatic juice. AB - OBJECTIVE: Point mutations of the K-ras oncogene at codon 12 have been described several months before the onset of pancreatic cancer in isolated cases of chronic pancreatitis (CP). The aim of this study was to evaluate the interest of a prospective follow-up of patients with CP and K-ras mutations at codon 12 in the detection of early pancreatic cancer. METHODS: From February 1996 to March 1998, 36 patients (mean age 52.6 yr, 31 men, five women) with CP (alcoholic: 61.1%, pancreas divisum: 5.6%, autoimmune: 5.6%, unknown origin: 27.7%) were included and then prospectively monitored (median duration of 22 months) for detection of pancreatic carcinoma. K-ras point mutations were examined by two-step polymerase chain reaction combined with restriction enzyme digestion in pancreatic juice collected during endoscopic retrograde pancreatography. RESULTS: Ten patients (27.8%) were positive for K-ras mutation. Patients with and without the mutation were not different with respect to age and sex ratio. K-ras mutations were homogeneously distributed according to the etiology (alcoholic vs nonalcoholic) and morphological characteristics (ductal stricture or mass vs none) of CP. A pancreatic carcinoma was discovered at an invasive stage in two patients, respectively at 7 and 17 months after disclosure of a K-ras mutation, versus none in patients without the mutation (p < 0.02). CONCLUSIONS: Presence of a K-ras gene mutation is not rare in patients with CP and represents an increased risk of developing pancreatic cancer. However, its utility for the detection of early pancreatic cancer remains doubtful in clinical practice. PMID- 11280538 TI - Endoscopic ultrasound in idiopathic acute pancreatitis. AB - OBJECTIVE: The aim of this study was to determine the utility of endoscopic ultrasound (EUS) in patients with unexplained acute pancreatitis, and whether endoscopic retrograde cholangiopancreatography (ERCP) is subsequently needed. METHODS: Subjects who underwent EUS for assessment of idiopathic acute pancreatitis were identified, their medical records were reviewed, and they were contacted for a follow-up telephone interview. EUS diagnosis was compared with the final diagnosis and outcome. RESULTS: EUS revealed a cause of pancreatitis in 21 of the 31 subjects (68%), including microlithiasis in five (16%), chronic pancreatitis in 14 (45%), pancreas divisum in two (6.5%), pancreatic cancer in one (3.2%), and was not diagnostic in 10 (32%). During a mean follow-up period of 16 months, diagnosis changed in four subjects (13%), and nine subjects (29%) had ERCP because of persistent symptoms or recurrent pancreatitis. CONCLUSION: EUS, a less invasive test than ERCP, demonstrated an etiology in two-thirds of patients with idiopathic acute pancreatitis. Most patients did not require ERCP during the follow-up period. EUS can be an alternative to ERCP in patients with unexplained acute pancreatitis. PMID- 11280539 TI - Decreased efficacy of polyethylene glycol lavage solution (golytely) in the preparation of diabetic patients for outpatient colonoscopy: a prospective and blinded study. AB - OBJECTIVES: The aim of the present study was to compare, in a prospective and blinded fashion, the efficacy of 6 L of polyethylene glycol-based lavage solution (Golytely) administered on an outpatient basis in diabetic versus nondiabetic patients. METHODS: A total of 54 consecutive nondiabetic and 45 consecutive diabetic patients requiring outpatient colonoscopy underwent colonic cleansing by drinking 6 L of Golytely the evening before the procedure. The entire procedure, from rectum to cecum, was videotaped and coded for later review by the Chief of Endoscopy who was blinded to the identity and medical history of the patients. The primary outcome measure was the quality of the preparation score, numerically rated on a 14-point scale (0-13) based on the surface area of the mucosa that could be examined and the consistency of the residual stool. RESULTS: There was a significant difference in the quality ratings for the bowel preparations, with an overall superior preparation in the nondiabetic group (2.4+/-1.6 vs 5.4+/-3.1, p < 0.001). A total of 97% of the nondiabetic patients had a preparation rated as good or better, compared with only 62% of the diabetic patients (p < 0.001). Preparations rated as poor or futile, necessitating repeat colonoscopy, occurred in no nondiabetic but in 9% of diabetic patients (p < 0.01). Within the diabetic group, there was no significant difference in bowel preparation scores between those patients aged >70 yr and those <70 yr, those requiring and those not requiring insulin, those with Hb A1c values >8% and those with values <8%, and those with and without peripheral neuropathy. CONCLUSION: We conclude that diabetic patients (irrespective of insulin use, diabetic control, or diabetic neuropathy) have a significantly poorer response to a 6-L Golytely preparation than do nondiabetic patients. PMID- 11280540 TI - Association of bowel movement frequency and use of laxatives with the occurrence of symptomatic gallstone disease in a prospective study of women. AB - OBJECTIVES: The authors prospectively examined the association between bowel movement frequency (used as a proxy for intestinal transit), laxative use, and the risk of symptomatic gallstone disease. METHODS: A total of 79,829 women, aged 36-61 yr, without a history of symptomatic gallstone disease and free of cancer, responded to a mailed questionnaire in 1982 that assessed bowel movement frequency and use of laxatives. Between 1984 and 1996, 4,443 incident cases of symptomatic gallstone disease were documented. Relative risks (RRs) of symptomatic gallstone disease and 95% confidence intervals (CIs) were calculated using logistic regression. RESULTS: After controlling for age and established risk factors, the multivariate RRs were, compared to women with daily bowel movements, 0.97 (95% CI 0.86-1.08) for women with bowel movements every third day or less, and 1.00 (95% CI 0.91-11.1) for women with bowel movement more than once daily. No trend was evident. As compared to women who never used laxatives in 1982, a significant modest inverse association was seen for monthly laxative use, with a multivariate RR of 0.84 (95% CI 0.72-0.98), and weekly to daily laxative use was associated with a RR of 0.88 (95% CI 0.78-1.02). CONCLUSIONS: These findings do not support an association between infrequent bowel movements and risk of symptomatic gallstone disease in women, and indicate that simple questions directed at bowel movement frequency are unlikely to enhance our ability to predict risk of symptomatic gallstone disease. The slightly inverse association between use of laxatives and risk of symptomatic gallstone disease may be due to a mechanism that is not related to bowel movement frequency. PMID- 11280541 TI - Infliximab for Crohn's disease in clinical practice at the Mayo Clinic: the first 100 patients. AB - OBJECTIVE: The aim of this study was to report the clinical outcome and adverse events in the first 100 patients with refractory inflammatory and/or fistulizing Crohn's disease treated with infliximab at the Mayo Clinic. METHODS: Patient data was abstracted from medical records. Clinical response was classified as complete response, partial response, and nonresponse. RESULTS: Indications for infliximab therapy were: inflammatory disease (61 patients), fistulizing disease (26 patients), or both (13 patients). Patients received one to seven infusions of infliximab (5 mg/kg) for a total of 242 infusions. In all, 50 patients had complete response, 22 had partial response, and 28 had nonresponse. Median time to response was 7 days (range 1-21 days). Median duration of response was 10.3 weeks (range 3-25 wk). A total of 95 patients received concomitant treatment with immune modifiers. Steroid withdrawal was possible in 29/40 patients (73%). Median time of follow-up was 34 wk (range 14-48 wk). Clinically significant adverse events after infliximab included: abscess formation in two patients (perianal, peristomal), pneumonia in two patients, varicella zoster in three patients, candida esophagitis in one patient, and infusion-related reactions in 19 patients. A total of 23 patients were continued on infliximab as maintenance treatment. CONCLUSIONS: This study provides additional evidence that infliximab is safe and beneficial in clinical practice for refractory Crohn's disease. PMID- 11280542 TI - Diagnostic value of anti-Saccharomyces cerevisiae and antineutrophil cytoplasmic autoantibodies in inflammatory bowel disease. AB - OBJECTIVES: Correct diagnosis of inflammatory bowel disease (IBD), especially the differentiation between Crohn's disease (CD) and ulcerative colitis (UC), is highly important toward treatment and prognosis. Serological markers are noninvasive diagnostic tools that could be of value in differentiating CD from UC, in cases of indeterminate colitis, and in the identification of subgroups in IBD. The aim of this study was to evaluate the diagnostic accuracy of perinuclear antineutrophil cytoplasmic (pANCA) and anti-Saccharomyces cerevisiae antibodies (ASCA) for IBD. METHODS: ASCA and pANCA were studied in a large cohort of consecutive IBD patients (n = 582: 407 CD, 147 UC, and 28 indeterminate colitis), patients with non-IBD diarrheal illnesses (n = 74), and healthy controls (n = 157). An indirect immunofluorescence technique and a standardized ELISA were performed for detection of pANCA and ASCA, respectively. RESULTS: Prevalence of ASCA and pANCA was high in CD patients (59.7%) and UC (49.7%) patients, respectively. Positivity for both markers was significantly lower in healthy and non-IBD controls. Accuracy data (sensitivity, specificity, PPV, and NPV, respectively) for differentiating IBD from controls are as follows: ASCA+: 60% (243/407), 91% (345/378), 88% (243/276), and 68% (345/509); pANCA+: 50% (73/147), 95% (605/638), 69% (73/106), and 89% (605/679); ASCA+/pANCA-: 56% (229/407), 94% (355/378), 91% (229/252), and 67% (355/533); and pANCA+/ASCA-: 44% (65/147), 97% (620/638), 78% (65/83), and 88% (620/702). CONCLUSIONS: Specificity of serological markers for IBD is high, but low sensitivity makes them less useful as diagnostic tests. The combination of tests is probably more powerful, although, clinical subgroups still need to be defined. The usefulness of these markers in indeterminate colitis needs to be studied prospectively. PMID- 11280543 TI - Asymptomatic inflammatory bowel disease with special reference to ulcerative colitis in apparently healthy persons. AB - OBJECTIVE: We examined cases of asymptomatic inflammatory bowel diseases, particularly asymptomatic ulcerative colitis, which were found in apparently healthy Japanese persons who underwent general health screening. METHODS: Patients with positive immunological fecal occult blood test (IFOBT) among approximately 236,000 persons participating in the health screening program at the Aichi Prefectural Center for Health Care for the past 9 yr underwent total colonoscopy. In patients with ulcerative colitis, we investigated the sex and age distributions, extent of lesion, endoscopic activity, incidence rate, and clinical course. RESULTS: In all, 35 cases of inflammatory bowel disease were detected, and 274 cases of colorectal cancer (not discussed here) were detected in the same population. The 35 cases of inflammatory bowel disease consisted of 19 cases of ulcerative colitis (12 of asymptomatic and minimally symptomatic ulcerative colitis, and seven of symptomatic or with past history of ulcerative colitis); five of intestinal tuberculosis; two of Crohn's disease; two of amebic colitis; and seven of endoscopic colitis. The 12 patients with asymptomatic and minimally symptomatic ulcerative colitis consisted of 11 men and one woman aged 36-63 yr (mean 46.2 yr). We classified these cases into three grades of severity according to endoscopic findings: four cases were mild, eight moderate, and none severe. Of these 12 cases, three were found endoscopically because of positive IFOBT, although barium enema was normal. Anatomic types of colitis cases included three of total colitis, three left-sided colitis, two proctitis, and four right sided or segmental colitis. In one case, the disease extended proximally during the course of observation. CONCLUSIONS: We found 35 cases of inflammatory bowel disease because of positive IFOBT performed as part of a general health screening. Of these, 19 cases were ulcerative colitis. These included many asymptomatic and minimally symptomatic cases, which could be very important in helping to elucidate the natural history of ulcerative colitis; thus, long-term follow up is necessary. PMID- 11280545 TI - Vitamin B12 deficiency in untreated celiac disease. AB - OBJECTIVES: Iron and folate malabsorption are common in untreated celiac disease as the proximal small intestine is predominantly affected. Vitamin B12 deficiency is thought to be uncommon, as the terminal ileum is relatively spared. This study aims to investigate the prevalence of vitamin B12, deficiency in patients with untreated celiac disease. METHODS: Prospective study of 39 consecutive biopsy proven celiac disease patients (32 women, seven men; median age 48 yr, range 22 77 yr) between September 1997 and February 1999. The full blood count, serum vitamin B12, red blood cell folate, and celiac autoantibodies (IgA antigliadin and IgA antiendomysium antibodies) were measured before and after a median of 4 months (range 2-13 months) of treatment with a gluten-free diet. In vitamin B12 deficient patients, intrinsic factor antibodies and a Schilling test, part 1, were performed. RESULTS: A total of 16 (41%) patients were vitamin B12 deficient (<220 ng/L) and 16 (41%) patients (11 women and live men) were anemic. Concomitant folate deficiency was present in only 5/16 (31%) of the vitamin B12 patients. The Schilling test, performed in 10 of the vitamin B12-deficient patients, showed five low and five normal results. Although only five patients received parenteral vitamin B12, at follow-up the vitamin B12 results had normalized in all patients. Acral paraesthesia at presentation in three vitamin B12-deficient patients resolved after vitamin B12 replacement. CONCLUSIONS: Vitamin B12 deficiency is common in untreated celiac disease, and concentrations should be measured routinely before hematinic replacement. Vitamin B12 concentrations normalize on a gluten-free diet alone, but symptomatic patients may require supplementation. PMID- 11280544 TI - Outcome and management of patients with large rectoanal intussusception. AB - OBJECTIVES: Rectoanal intussusception is the funnel-shaped infolding of the rectum, which occurs during evacuation. The aims of this study were to evaluate the risk of full thickness rectal prolapse during follow-up of patients with large rectoanal intussusception, and whether therapy improved functional outcome. METHODS: Between September 1988 and July 1997, patients diagnosed with a large rectoanal intussusception by cinedefecography (intussusception > or = 10 mm, extending into the anal canal) were retrospectively evaluated. Patients with full thickness rectal prolapse on physical examination or cinedefecography were excluded, as were patients with colonic inertia or a history of surgery for rectal prolapse. The patients were divided into three groups according to the treatment received: group I, conservative dietary therapy; group II, biofeedback; and group III, surgery. Outcomes were obtained by postal questionnaires or telephone interviews. Parameters included age, gender, past medical and surgical history, change of bowel habits, fecal incontinence score, and development of full thickness rectal prolapse. RESULTS: Of the 63 patients, 18 were excluded (seven patients had confirmed full thickness rectal prolapse, four had previous surgery for rectal prolapse, three had colonic inertia, and four died). Follow-up data were obtained in 36 (80%) of the remaining 45 patients. The mean follow-up of this group was 45 months (range, 12-118 months). There were 34 women and two men, with a mean age of 72.4 yr (range, 37-91 yr). The mean size of the intussusception was 2.2 cm (range, 1.0-5.0 cm). The patients were classified as follows: group I, 13 patients (36.1%); group II, 13 patients (36.1%); and group III, 10 patients (27.8%). Subjectively, symptoms improved in five (38.5%), four (30.8%), and six (60.0%) patients in the three groups (p > 0.05). Among the patients with constipation, the decrease in numbers of assisted bowel movements per week (time of diagnosis to present) was significantly greater in group II compared to group 1 (8.1+/-2.8 vs 0.8+/-0.5, respectively, p = 0.004). Among the patients with incontinence, incontinence scores improved more in group II as compared to either group I or group III (time of diagnosis to present, 3.7+/-4.2 to 1.1+/-5.4 vs 1.4+/-2.2, respectively, p > 0.05). Six patients (two in group I, three in group II, and one in group III) had the sensation of rectal prolapse on evacuation; however, only one patient in group I developed full thickness rectal prolapse. CONCLUSIONS: This study demonstrated that the risk of full thickness rectal prolapse developing in patients medically treated for large intussusception is very small (1/26, 3.8%). Moreover, biofeedback is beneficial to improve the symptoms of both constipation and incontinence in these patients. Therefore, biofeedback should be considered as the initial therapy of choice for large rectoanal intussusception. PMID- 11280546 TI - Prevalence of thyroid disorders in untreated adult celiac disease patients and effect of gluten withdrawal: an Italian multicenter study. AB - OBJECTIVES: Many afflictions have been associated with celiac disease, but chance associations may exists. The aim of this study was to establish, by means of a multicenter prospective study, the prevalence of thyroid impairment among adult patients with newly diagnosed celiac disease and to evaluate the effect of a 1-yr gluten withdrawal on thyroid function. METHODS: A total of 241 consecutive untreated patients and 212 controls were enrolled. In 128 subjects a thorough assessment, including intestinal biopsy, was repeated within 1 yr of dietary treatment. Thyroid function was assayed by measuring the levels of TSH, free T3, free T4, thyroperoxidase, and thyroid microsome antibodies. RESULTS: Thyroid disease was 3-fold higher in patients than in controls (p < 0.0005). Hypothyroidism, diagnosed in 31 patients (12.9%) and nine controls (4.2%), was subclinical in 29 patients and of nonautoimmune origin in 21. There was no difference regarding hyperthyroidism, whereas autoimmune thyroid disease with euthyroidism was present in 39 patients (16.2%) and eight controls (3.8%). In most patients who strictly followed a 1-yr gluten withdrawal (as confirmed by intestinal mucosa recovery), there was a normalization of subclinical hypothyroidism. Twenty-five percent of patients with euthyroid autoimmune disease shifted toward either a subclinical hyperthyroidism or subclinical hypothyroidism; in these subjects, dietary compliance was poor. In addition, 5.5% of patients whose thyroid function was normal while untreated developed some degree of thyroid dysfunction 1 yr later. CONCLUSIONS: The greater frequency of thyroid disease among celiac disease patients justifies a thyroid functional assessment. In distinct cases, gluten withdrawal may single-handedly reverse the abnormality. PMID- 11280547 TI - Clinical utility of serodiagnostic testing in suspected pediatric inflammatory bowel disease. AB - OBJECTIVES: Confronted with nonspecific symptoms, accurate screening tests would be useful to clinicians to distinguish between functional childhood disorders and inflammatory bowel disease (IBD), thus avoiding invasive diagnostic testing. Traditional ulcerative colitis-specific perinuclear antineutrophil cytoplasmic antibody (pANCA) and Crohn's disease-specific anti-Saccharomyces cerevisiae antibody (ASCA) serodiagnostic assays have recently been modified, with ELISA cut off values recalculated to maximize sensitivity. The aim of this study was to determine whether the combination of these serodiagnostic tests could maximize diagnostic accuracy and minimize invasive investigations in pediatric patients presenting with nonspecific symptoms suggestive of IBD. METHODS: With investigators blinded to clinical diagnoses, ASCA, ANCA, and pANCA profiles were obtained prospectively from 128 patients undergoing complete diagnostic evaluation for IBD. In phase I, diagnostic accuracy and predictive values of the modified and traditional assays were compared for the IBD (n = 54) and non-IBD groups (n = 74). In phase II, the overall accuracy of a novel sequential diagnostic testing strategy was determined. Additionally, the potential number of invasive investigations avoided with the hypothetical application of this strategy to the cohort was determined. RESULTS: For phase I, the modified serodiagnostic assay was more sensitive (81 vs 69%), whereas the traditional assay had a higher specificity (96 vs 72%) for IBD (p < 0.05) For phase II, false positive diagnoses would have been reduced by 81%, yielding an overall sequential testing strategy accuracy of 84%. CONCLUSIONS: The incorporation of sequential noninvasive testing into a diagnostic strategy may avoid unnecessary and costly evaluations and facilitate clinical decision making when the diagnosis of IBD in children is initially uncertain. PMID- 11280548 TI - Medical diagnoses and procedures associated with clostridium difficile colitis. AB - OBJECTIVES: The aim of this study was to examine the associations of Clostridium difficile colitis with other comorbid conditions and procedural interventions among hospitalized patients. METHODS: The Patient Treatment File of the Department of Veterans Affairs contains the computerized records of all inpatients treated in 172 Veterans Affairs hospitals distributed throughout the United States. The computerized medical records of 15,091 cases with C. difficile colitis and 61,931 controls without the diagnosis were extracted from the annual files between 1993 and 1998. In a multivariable logistic regression, the occurrence of C. difficile colitis served as outcome variable, whereas the occurrences of other diagnoses or procedures served as predictor variables. RESULTS: The total numbers of diagnoses in the case and control group were 136,840 and 465,972, respectively. The numbers of procedures were 75,479 and 129,612, respectively. C. difficile colitis was significantly associated with HIV infection, candidiasis, malignant neoplasm and chemotherapy, malnutrition, pneumonia, aspiration pneumonitis, intestinal obstruction, diverticulitis, renal failure, urinary tract infection, decubitus, and osteomyelitis. Interventional procedures involving the respiratory tract, bone marrow biopsy, arterial and venous catheterization, urinary catheterization, dialysis, gastrostomy tube, and physical therapy were also frequently associated with the development of C. difficile colitis. CONCLUSIONS: These associations reflect the influence of causal relationships (such as the use of antibiotics and chemotherapy), an increased risk of exposure to C. difficile among immobilized bedridden patients with chronic disease states, or a general system failure in patients with end stage disease. Knowledge of such associations could help to alert physicians to an increased risk of C. difficile colitis among particular groups of susceptible patients. PMID- 11280549 TI - Prevalence of cytomegalovirus infection in severe refractory ulcerative and Crohn's colitis. AB - OBJECTIVES: Cytomegalovirus infection has been reported as a cause of refractory inflammatory bowel disease, but no data are available on its prevalence in severe colitis. The aim of this study was to evaluate the prevalence and outcome of cytomegalovirus infection in a consecutive series of patients with severe steroid refractory colitis admitted to our department from 1997 to 1999. METHODS: Among 62 patients with severe colitis, 55 with ulcerative colitis and seven with Crohn's disease, 19 (30%) were resistant to intravenous steroids and bowel rest. In all of them, rectal biopsies were examined for cytomegalovirus (the flexible proctoscopy being performed without air insufflation and limited to the first 10 cm). Buffy coat preparation on leukocytes was also performed to detect systemic infection. If cytomegalovirus was not detected, cyclosporine was started. RESULTS: In seven (five with ulcerative colitis and two with Crohn's disease) out of 19 (36%) patients with refractory disease, cytomegalovirus was diagnosed in the rectal specimens as well as by buffy coat preparation. Five patients went into remission after antiviral treatment (three with ganciclovir and two with foscarnet). One patient did not respond and was operated on. In one patient, cytomegalovirus was found in the surgical specimen. CONCLUSIONS: Cytomegalovirus infection is a frequent cause of severe refractory colitis. Rectal biopsy should always be performed in severe steroid-resistant colitis. PMID- 11280550 TI - Mean platelet volume: a useful marker of inflammatory bowel disease activity. AB - OBJECTIVES: We investigated whether the mean platelet volume would be a useful marker in the evaluation of inflammatory bowel disease activity. METHODS: Complete blood count, C-reactive protein, erythrocyte sedimentation rate, serum thrombopoietin and erythropoietin, plasma beta-thromboglobulin, and platelet factor 4 were measured in 93 patients with ulcerative colitis, 66 patients with Crohn's disease, and 38 healthy blood donors. Disease activity was assessed by the Clinical Colitis Activity Index in patients with ulcerative colitis and by the Crohn's Disease Activity Index in patients with Crohn's disease. RESULTS: Mean platelet count was increased in patients with active compared to inactive ulcerative colitis (p < 0.05), and in patients with active compared to inactive Crohn's disease (p = 0.0002) or healthy controls (p < 0.0001). On the other hand, mean platelet volume was significantly decreased in patients with active compared to inactive ulcerative colitis (p = 0.02) or healthy controls (p < 0.0001), and in patients with active compared to inactive Crohn's disease (p = 0.0005) or healthy controls (p < 0.0001). Mean platelet volume was inversely correlated with the white blood cell count (r = -0.17, p = 0.02), C-reactive protein (r = -0.46, p = 0.009) and erythrocyte sedimentation rate (r = -0.28, p = 0.008). No significant correlations were found between mean platelet volume and serum thrombopoietin or erythropoietin levels; however, a strong negative correlation between mean platelet volume and beta-thromboglobulin (r = -0.34, p < 0.0001) and platelet factor 4 (r = -0.30, p = 0.0002) was observed. CONCLUSIONS: Mean platelet volume is significantly reduced in active inflammatory bowel disease and is negatively correlated with the known inflammatory bowel disease activity markers and the platelet activation products. We propose that mean platelet volume provides a useful marker of activity in inflammatory bowel disease. PMID- 11280551 TI - Is mycophenolate mofetil an effective alternative in azathioprine-intolerant patients with chronic active Crohn's disease? AB - OBJECTIVES: Up to 50% of patients with Crohn's disease (CD) develop steroid dependent or refractory disease requiring immunosuppression. Azathioprine (AZA) is usually used for this purpose but must be withdrawn in up to 10% of patients because of adverse events. Mycophenolate mofetil (MMF) is of proven efficacy and safety in transplantation and in some autoimmune disorders. The aim of the present study was to investigate the effect of MMF, especially in AZA-intolerant patients with chronic active CD, in comparison to a matched control group treated with AZA. METHODS: In a retrospective study, 15 patients treated with MMF and 30 randomly chosen, matched patients treated with AZA for chronic active CD were compared over a period of 1 yr. Intolerance to AZA was the indication for MMF. Crohn's Disease Activity Index (CDAI), steroid demand, extraintestinal manifestations, and hematological and biochemical parameters were assessed at the start of therapy and 1, 2, 3, 6, 9, and 12 months thereafter. RESULTS: All patients who completed the 12 months of treatment (77% AZA, 60% MMF) achieved remission. Under MMF, the cumulative prednisolone dose could be reduced by 1 g in the first half year, whereas, under AZA, this reduction was possible only in the second half year. MMF patients had almost twice as many flare-ups (80% vs 47%). Adverse events prompted the withdrawal of AZA in five patients (17%) and of MMF in three (20%). CONCLUSIONS: Both drugs are effective in inducing remission. AZA seems to be more effective in maintaining remission. The onset of therapeutic effect is delayed less under MMF. Both drugs have steroid sparing potential, which is delayed under AZA. It seems that AZA still is the immunouppressant of choice in chronic active CD, but MMF is a reasonable alternative in patients who do not tolerate AZA. PMID- 11280552 TI - Colonic chicken skin mucosa: association with juvenile polyps in children. AB - OBJECTIVES: Chicken skin mucosa is a newly described endoscopic finding associated with colonic neoplasms in adults. Chicken skin mucosa was sought in children with juvenile polyps to determine the prevalence, endoscopic features, and location. An alternative theory is proposed for the pathogenic mechanism of this finding. METHODS: Children having colonoscopy and polypectomy were prospectively evaluated for the presence of chicken skin mucosa. The location of the polyps was determined at colonoscopy; the size of removed polyps was measured during processing of samples in pathology. Biopsies from colonic chicken skin mucosa were stained with hematoxylin and eosin and mucicarmine. RESULTS: Over a 1 yr period, 27 juvenile polyps were removed from 15 children at colonoscopy. Eleven of 15 children (73%) were found to have polyps with chicken skin mucosa; overall, 43% of the polyps had associated chicken skin mucosa. Chicken skin mucosa-positive polyps were larger than chicken skin mucosa-negative polyps and were only found in the rectosigmoid colon. Lipid-laden macrophages were found in all samples of chicken skin mucosa tested. CONCLUSIONS: Chicken skin mucosa is a common finding in children with juvenile polyps. It probably is the result of local mucosal trauma, rather than a preneoplastic lesion. PMID- 11280553 TI - Effect of ecabet sodium enema on mildly to moderately active ulcerative proctosigmoiditis: an open-label study. AB - OBJECTIVES: Ecabet sodium (ES), a nonabsorbable antigastric ulcer agent, has been shown to adhere to the region of an ulcer. It topically enhances gastric mucosal defensive factors such as the endogenous prostaglandins, capsaicin-sensitive sensory nerves, nitric oxide, and mucin. All of these mucosal defensive factors play an important role in maintaining the mucosal integrity of the colon and rectum. Therefore, we investigated the effect of ES in patients with mildly to moderately active ulcerative proctosigmoiditis. METHODS: In an open-label study, seven patients with mildly to moderately active ulcerative colitis (UC) who had an inflamed mucosa in the rectum and/or sigmoid and were resistant to 4-wk topical and systemic standard treatment were treated with an ES enema b.i.d. for 14 days. The enema consisted of ES (1 g) and tepid water (20 or 50 ml). These patients were assessed by the Clinical Activity Index, colonoscopically, and histologically before and after the ES therapy. The ES therapy was started after obtaining informed consent from the patients. RESULTS: Six of the seven patients responded to therapy and achieved clinical, endoscopic, and histological remissions. One patient was withdrawn because of increased stool frequency. All six patients who completed the study showed a significant change in the mean Clinical Activity Index score from 5.3+/-1.4 (mean +/- SD) to 0.5+/-0.8 (p < 0.05), in the colonoscopic score from 3.0+/-0.9 to 0.8+/-0.4 (p < 0.05), and in the histological score from 2.7+/-0.5 to 0.5+/-0.6 (p < 0.05), and achieved remission at the end of the study. There were no side effects attributable to the ES therapy. Five of the six patients are still in clinical remission after a median follow-up period of 5 months. CONCLUSIONS: The ES enemas proved to be a safe and potentially useful adjuvant therapy currently available for treating patients with mildly to moderately active ulcerative proctosigmoiditis. A controlled study is necessary to confirm our results. PMID- 11280554 TI - Relation between cigarette smoking and celiac disease: evidence from a case control study. AB - OBJECTIVES: It has been suggested that environmental factors other than gliadin might play a role in pathogenesis of celiac disease. Cigarette smoking was reported to exert a protective effect against the development of symptomatic celiac disease; however, this relationship was not confirmed. The aim of this study was to determine the effect of cigarette smoking on celiac disease. METHODS: A cohort of 87 consecutive celiac disease patients attending the clinic of Malabsorption and 174 age- and sex-matched individuals diagnosed with functional GI disorder were included in the study. Clinical information was obtained both at the time of diagnosis and at follow-up by reviewing the clinical history. Smoking information was obtained through an in-person interview using a questionnaire. RESULTS: Although 33% of controls were current smokers at the time of the study, only 16% of celiac patients were smokers at diagnosis (odds ratio, 0.39; 95% confidence interval 0.19-0.79; p < 0.006). The proportion of nonsmokers among patients (84%) was significantly greater than that among controls (67%; odds ratio, 2.54; 95% confidence interval 1.27-5.16; p < 0.007). Current smoker patients had a lower baseline BMI (p < 0.05) and body weight (p < 0.05) compared to former smokers. Compared with nonsmokers, control individuals who were active smokers at entry in the study were younger (p < 0.02) and had lower body weight (p < 0.03) and BMI (p < 0.03). Interestingly, positive lineal correlation was observed between age at diagnosis and daily cigarette consumption (r = 0.72; p < 0.004) in active smokers. We did not detect any relationship either between causes for cessation of smoking and clinical symptoms or between differences in the proportions of smoking habits when patients were stratified according to their clinical status at diagnosis (symptomatic vs subclinical/asymptomatic cases). CONCLUSIONS: This study provides evidence that, compared with control subjects, a significantly lower proportion of patients with celiac disease were current smokers at the time of diagnosis, and that cigarette smoking delayed diagnosis of celiac disease. Our study suggests that the nutritional compromise of patients with celiac disease who smoked resulted from the summation of the effect of celiac disease per se and that produced by the smoking habit. Further studies are necessary to identify whether the relationship between smoking and celiac disease is causal or incidental. PMID- 11280555 TI - Tolerability and safety of alosetron during long-term administration in female and male irritable bowel syndrome patients. AB - OBJECTIVES: Alosetron (Lotronex) is a new therapeutic agent for irritable bowel syndrome (IBS) in women with diarrhea-predominant IBS. This multicenter randomized, double-blind, placebo-controlled study assessed the safety and tolerability of alosetron during long-term (< or = 12 months) treatment. METHODS: A total of 859 subjects (637 female and 222 male) with IBS were enrolled from 130 sites in the United States and were randomized 3:1 to receive 1 mg alosetron or placebo b.i.d. for 48 wk; of the subjects, 649 (76%) were randomized to the alosetron group and 212 (24%) to the placebo group. Of the original group, 850 subjects received at least one dose of alosetron (n = 640) or placebo (n = 210). RESULTS: In all, 59% of the subjects completed the study. Safety data were similar in treatment groups and within age, sex, racial origin, and hormone use. Adverse events were reported by 83% (530/640) and 76% (159/210) of subjects in the alosetron and placebo groups, respectively, (p < 0.05) and were similar with the exception of constipation; 32% of subjects receiving alosetron reported constipation, compared to 5% in the placebo group (p < 0.001). Most reports (72%) of constipation were of mild or moderate severity, and 66% of subjects with constipation had single episode of 8 days median duration. Constipation occurred a median of 13 days after initiating treatment and resolved spontaneously, with laxative, or after a brief interruption of therapy. Of the subjects, 4% (11/210) in the alosetron and 5% (28/ 640) in the placebo group experienced serious adverse events. Two deaths occurred in subjects with pre-existing cardiovascular risk factors; neither death was attributed to the study drug. CONCLUSIONS: Alosetron 1 mg b.i.d. for 12 months was well tolerated. Constipation is the most frequent adverse event, with a higher incidence of transient constipation in alosetron-treated patients, typically occurring in the first month of treatment. PMID- 11280556 TI - Seasonal variation in acute intestinal vasculopathy mortality in France. AB - OBJECTIVES: Seasonal and circadian rhythms are observed in cardiovascular diseases. Seasonal variation in acute intestinal vasculopathy has never been investigated. This report describes the seasonal variation of acute intestinal vasculopathy mortality in the French population. METHODS: All deaths that occurred among French adults over the period 1987-1996 (N = 20,830) for acute intestinal vasculopathy (International Classification of Diseases, Ninth Revision code 557.0) were examined retrospectively. Cumulated monthly averages were expressed as the percentage above or below the average monthly value during the entire study period. RESULTS: Deaths for acute intestinal vasculopathy peaked in January (15% above the trend), and were lowest in July (11% below the trend), both in the overall population (Roger's test: p < 0.001) and in subgroups defined by age (>69 yr old) and sex. Compared to other subgroups, the >90 yr old individuals had a higher incidence of acute intestinal vasculopathy with a greater amplitude of seasonal variation (p < 0.001). CONCLUSIONS: Awareness of higher risk during winter would help to reduce the high mortality from acute intestinal vasculopathy. A better understanding of this seasonal pattern would allow practitioners to improve early diagnosis and treatment. PMID- 11280557 TI - Treatment responses in collagenous colitis. AB - OBJECTIVE: In the nearly 20 yr since collagenous colitis was first recognized, the results of therapies have not been systematically described in substantial numbers of patients. We have therefore conducted a retrospective analysis of 26 patients treated in this institution during the years 1991-1994. METHODS: Twenty nine cases of collagenous colitis were obtained by review of biopsy specimens collected between 1991 and 1994 at The Mount Sinai Hospital. Each chart was reviewed for patient demographics, symptoms, coexisting conditions, specific therapies, and therapeutic outcomes. Additional data were obtained from telephone calls to patients when deemed necessary. Three patients were exeluded from the study because of lack of follow-up. Therapeutic outcomes were defined as follows: Complete Remission (CR): normalization of bowel function; Partial Remission (PR): 50% reduction in frequency of bowel movements; Failure: <50% reduction in frequency of bowel movements; or Relapse: return of symptoms after cessation of treatment. Median follow-up was 58 wk from time of diagnosis, with a range of 22 376 wk. RESULTS: The 26 patients (25 women, one man) had a mean age of 62 yr (range, 22-85 yr) at diagnosis. Of 26 patients, 22 responded to some form of therapy and one had spontaneous remission. Six of the responders ultimately remained in CR with no therapy. Twelve are maintained on 5-aminosalicylic acid (5 ASA) and or antidiarrheals to control symptoms. An additional six required prednisone throughout the follow-up period to remain in CR or PR. Two patients failed all therapy. CONCLUSION: Collagenous colitis is a treatable condition in most patients. We recommend initial therapy with antidiarrheals, followed by a trial of 5-ASA agent. A trial of 5-ASA in combination with prednisone should be attempted in patients refractory to 5-ASA alone, with subsequent attempts in the responders to taper prednisone and maintain remission with no therapy, if possible, or with 5-ASA and/or antidiarrheal agents if necessary. PMID- 11280558 TI - VEGF, basic-FGF, and TGF-beta in Crohn's disease and ulcerative colitis: a novel mechanism of chronic intestinal inflammation. AB - OBJECTIVE: Inflammatory bowel disease (IBD), the precise etiology of which remains unknown, is comprised of two forms of chronic intestinal inflammation; ulcerative colitis (UC) and Crohn's disease (CD). Recent evidence increasingly suggests that IBD is the result of dysfunctional immunoregulation manifested by inappropriate production of mucosal cytokines. An abnormal microcirculatory system has also been implicated in its pathogenesis. To elucidate the mechanism of ischemic change in IBD, we assesse serum concentration levels of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (b-FGF), and plasma level of endothelin-1 (ET-1). We also investigated the expression of VEGF, b-FGF, and transforming growth factor-beta1,2,3 (TGF-beta1,2,3) in tissue by immunostaining. METHODS: Blood samples were obtained from 11 patients with UC, 11 patients with CD, and 10 patients as controls. Paraffin-embedded samples were used for an immunohistochemical study. RESULTS: The concentration levels (in picograms per milliliter) were as follows: for ET-1, UC: 127+/-47.0, CD: 167.3+/ 35.1, and controls (asthma: 38.5+/-23.8, p < 0.01; diverticulitis: 40.5+/-25.6, p < 0.01), for b-FGF, UC: 9.2+/-1.9, CD: 9.1+/-1.5, and controls (asthma: 5.0+/-0, p < 0.01; diverticulitis: 5.0+/-0, p < 0.01), for VEGF, UC: 659.8+/-181.0, CD: 740.0+/-182.3, and controls (asthma: 193.7+/-58.7, p < 0.01; diverticulitis: 199.6+/-59.7, p < 0.01). The levels of VEGF and b-FGF were significantly higher in active IBD than those in the controls. There was a significant positive correlation among the serum levels of VEGF and b-FGF and the plasma level of ET 1; that is, elevated VEGF, b-FGF, and ET-1 levels correlated well with each other. Immunohistochemical studies showed increased venula in the submucosa and lamina propria. Overexpression of VEGF and b-FGF in endothelial cells was revealed and TGF-beta2 and TGF-beta3 were found in inflammatory cells of active IBD, but no change was observed around the vessels in the controls. CONCLUSIONS: It is suggested that the reciprocal reaction of these cytokines may contribute to angiogenesis in IBD b inducing intestinal ischemia through vasoconstriction. PMID- 11280559 TI - Appendectomy is more frequent but not a risk factor in Crohn's disease while being protective in ulcerative colitis: a comparison of surgical procedures in inflammatory bowel disease. AB - OBJECTIVE: Appendectomy was shown to be protective in patients with ulcerative colitis (UC). There are fewer data in Crohn's disease (CD). Other operations were less studied. The aim of this study was to investigate the prevalence of appendectomy, cholecystectomy, and tonsillectomy, including their timing, in patients with inflammatory bowel disease in comparison to controls. METHODS: Two hundred seventy-one patients with UC and 260 with CD, 475 clinic controls, and 428 community controls were interviewed. RESULTS: Appendectomy was found in 5.5% patients with UC, in 11% of clinic controls (p < 0.05), and 7.7% of community controls (p = not significant). The differences were more significant for appendectomy before onset of disease. Appendectomy was performed in 19.2% of patients with CD, in 10.9% of clinic controls, and in 10.1% of community controls (p < 0.01). However, there were no significant differences when only appendectomy before onset of disease was considered. Cholecystectomy was found in 1.5% of patients with UC, in 6.1% of clinic controls (p < 0.01), and in 4.5% of community controls (p = not significant). The difference remained significant when confined to operations performed before disease onset. No such difference was found in patients with CD. No significant difference was found in the prevalence of tonsillectomy between patients and controls. CONCLUSIONS: Appendectomy is protective in UC; it is more frequent, but not a risk factor in CD. The role of cholecystectomy should be investigated further. PMID- 11280560 TI - The use of screening and preventive therapies for gastroesophageal varices in patients referred for evaluation of orthotopic liver transplantation. AB - OBJECTIVE: Screening for varices has been recommended in patients with cirrhosis to prevent variceal hemorrhage (primary prophylaxis). In addition, therapy is recommended after the initial episode of variceal bleeding to prevent recurrence (secondary prophylaxis). However, the degree of adherence to these recommendations remains unclear. The purpose of our study was to determine whether these recommendations are being followed in patients presenting for evaluation of orthotopic liver transplantation. METHODS: One hundred twenty-five patients referred for liver transplantation were evaluated. Data regarding demographics, clinical information, relevant time intervals (diagnosis of cirrhosis to screening, screening to initial variceal bleeding, variceal bleeding to referral, diagnosis of cirrhosis to referral), screening strategies used, and implementation of primary or secondary prophylaxis was obtained. The differences among quantitative variables were analyzed with Student's t test. Qualitative variables were evaluated with the Mantel-Haenzel chi2 test or Fisher's exact test. Statistical significance was designated at p < 0.05. RESULTS: Our study found that 46% of patients presenting for evaluation of liver transplantation had screening endoscopy or radiological studies to detect the presence of varices. On the contrary, secondary prophylaxis was performed in all patients with a prior history of variceal hemorrhage. Screening for varices displayed no regional differences. CONCLUSIONS: In our cohort, screening for varices is not being consistently performed, thus delaying the timely implementation of primary prophylaxis. Therefore, the adherence to currently available practice guidelines and the education of physicians to implement screening in this patient population is an important goal. PMID- 11280561 TI - The accuracy of SM-HCV rapid test for the detection of antibody to hepatitis C virus. AB - OBJECTIVE: Conventional tests for antibody to hepatitis C virus (HCV) require considerable time before results are available. The aim of this study is to examine the accuracy of a new quick test (SM-HCV Rapid Test) for the detection of antibody to hepatitis C virus with reference to the well-established third generation enzyme immunoblot assay (EIA-3; Abbott Laboratories, Chicago, IL). METHODS: A total of 290 subjects (100 patients with chronic hepatitis C infections, 95 patients with other chronic liver diseases, 95 healthy subjects) were recruited. Thirty microliters of serum was tested for anti-HCV by SM-HCV Rapid Test according to the manufacturer's instruction. Liver function tests and serum HCV RNA by polymerase chain reaction (PCR) were measured. RESULTS: In the 100 patients positive for anti-HCV by EIA-3, 98 of these patients were also positive for anti-HCV by SM-HCV Rapid Test. In the 95 patients with other chronic liver diseases, 94 samples were negative for anti-HCV by both EIA-3 and SM-HCV Rapid Test. The remaining one patient was positive for anti-HCV by the EIA-3 but negative by the SM-HCV Rapid Test. In the 95 controls, which were negative for anti-HCV by EIA-3, all were also negative for anti-HCV by SM-HCV Rapid Test and HCV RNA by PCR. Using EIA-3 as the gold standard screening test for anti-HCV, the sensitivity and the specificity of SM-HCV Rapid Test were 98% and 100%, respectively. The positive predictive value and negative predictive value of SM HCV Rapid Test were 100% and 97.9%, respectively. CONCLUSIONS: SM-HCV Rapid Test is a reliable test with high sensitivity and specificity. The anti-HCV result can be available within a very short period of time. It is a useful screening test for anti-HCV. PMID- 11280562 TI - Postoperative morbidity, mortality, costs, and long-term survival in severely obese patients undergoing orthotopic liver transplantation. AB - OBJECTIVE: Severely obese patients who undergo orthotopic liver transplantation are likely to have higher morbidity, mortality, costs, and a lower long-term survival. METHODS: This case-control study was done at a university hospital. One hundred twenty-one consecutive patients who underwent liver transplantation between 1994 and 1996 were studied. Severe obesity was defined as body mass index (BMI) more than 95th percentile (>32.3 for women and >31.1 for men), and moderate obesity was defined as BMI between 27.3 and 32.3 for women and 27.8 and 31.1 for men. The outcome measures were intraoperative complications, postoperative complications (wound infections, bile leak, vascular complications), length of hospital stay, costs of transplantation, and long-term survival RESULTS: The baseline characteristics, UNOS status, and cause of liver disease at the time of transplantation were similar in severely obese (n = 21, BMI = 37.4+/-4.8 kg/m2), obese (n = 36, BMI 28.7+/-0.9 kg/m2), and nonobese patients (n = 64, BMI 23.8+/ 2.5 kg/m2). The intraoperative complications and transfusion requirements were similar in all three groups. The postoperative complications such as respiratory failure (p = 0.009) and systemic vascular complications (p = 0.04) were significantly higher in severely obese patients. The overall perioperative complication rate was 0.61 (39 of 64 patients) in nonobese patients, 0.77 (28 of 36 patients) in obese patients, and 1.43 (30 of 21 patients) in severely obese patients (p = 0.01). Infections were the leading cause of death in all groups accounting for 57-66% of deaths. The length of hospital stay was significantly higher in obese patients. The hospital costs of transplantation were higher ($30,000-$40,000) in severely obese patients than in nonobese patients. The long term patient survival was similar between the groups (Kaplan-Meier analysis). CONCLUSIONS: Despite higher postoperative complications, severely obese patients have an acceptable long-term survival, which is comparable to nonobese patients. The cost of transplantation is higher among severely obese patients. There was no increased incidence of cardiovascular mortality among severely obese patients during the follow-up period. PMID- 11280564 TI - Hepatic manifestations of hemophagocytic syndrome: a study of 30 cases. AB - OBJECTIVE: Hemophagocytic syndrome has been defined as the combination of a proliferation of cytologically benign, actively phagocytic macrophages in bone marrow, spleen, or lymph nodes in association with fever, cytopenia, splenomegaly, and hypertriglyceridemia. Hepatic dysfunction is often present but the nature of the hepatic lesions and related manifestations have not been fully characterized. The aim of this study was to ascertain the features of hepatic involvement in hemophagocytic syndrome. METHODS: Thirty patients with hemophagocytic syndrome were retrospectively studied. Inclusion criteria included: 1) bone marrow with hemophagocytic histiocytosis, 2) clinical or biological signs of hepatic involvement, and 3) available liver specimen. RESULTS: The association of fever, jaundice, and hepatomegaly or splenomegaly was present in 50% of the patients. Median value for serum alanine transaminase activity was five times the upper limit of normal values (range, 0.3-125), for serum alkaline phosphatase activity 2.7 upper limit of normal values (range, 0.2 47.7), for total bilirubin 136 micromol/L (range, 4-681 micromol/L), and for factor V 70% (range, 19-145%). Sinusoidal dilatation with hemophagocytic histiocytosis were found in the biopsy specimen in all patients. An underlying condition potentially responsible for altered immune function (lymphoma, leukemia, liver transplantation) was identified in 29 patients. Liver biopsy was diagnostic for the underlying condition in 15 patients (including eight cases with nonspecific bone marrow biopsy findings). High serum bilirubin, elevated serum alkaline phosphatase activity, low factor V level, and lack of treatment for the underlying disease were associated with a poor prognosis. CONCLUSIONS: Hemophagocytic syndrome should be suspected in immunodeficient patients with fever, jaundice, and hepatosplenomegaly. Hepatic lesions are characterized by nonspecific sinusoidal dilatation with hemophagocytic histiocytosis and in 50% of the patients by alterations specific to the underlying condition. Liver biopsy is a useful diagnostic procedure in patients with this clinical presentation. PMID- 11280563 TI - Clinical significance of autoantibody to hepatocyte membrane antigen in type 1 autoimmune hepatitis. AB - OBJECTIVE: By using HepG2 as flow cytometry target, we have reported that autoantibody to hepatocyte membrane antigen (anti-HMA) was frequently found in autoimmune hepatitis (AIH) patients. In this study, we have examined this autoantibody in relation to clinical features in these patients. METHODS: HepG2 cells were incubated with diluted serum and subsequently with FITC-conjugated antihuman immunoglobulin. The results were expressed as relative fluorescence intensity. The prevalence of anti-HMA was estimated by setting the upper limit of mean +/- 3 SD obtained from healthy subjects. RESULTS: We found that the mean relative fluorescence intensity was 1.67+/-0.5 in AIH with low serum ALT level (group 1 AIH), 4.20+/-1.9 in AIH with high serum ALT level (group 2 AIH), and 1.92+/-0.9 in age-matched chronic hepatitis C virus-positive patients. Their positive rate was 37.5% (three of eight) in group 1 AIH, 95.0% (19 of 20) in group 2 AIH, and 33.3% (four of 12) in chronic hepatitis C patients. In 12 group 2 AIH patients, their mean relative fluorescence intensity was significantly decreased during immunosuppressive therapy. The association between serum ALT level and anti-HMA was confirmed by the facts that a significant direct quantitative relationship existed between these two levels and by serial studies of anti-HMA in four AIH patients. Anti-HMA was also detected in five non-B, non-C hepatitis patients having clinical features resembling those of AIH. CONCLUSIONS: The present results have shown that the anti-HMA was tightly associated with the degree of hepatocyte inflammation and that the measurement of anti-HMA may have some advantage in clinical evaluation of some of non-B, non-C hepatitis patients. PMID- 11280565 TI - Prevalence of hepatitis A virus and hepatitis B virus immunity in patients with polymerase chain reaction-confirmed hepatitis C: implications for vaccination strategy. AB - OBJECTIVES: Administration of vaccine for hepatitis A virus (HAV) and hepatitis B virus (HBV) is recommended for patients with chronic hepatitis C (CHC) because of the potential for increased severity of acute hepatitis superimposed on existing liver disease. The aim of this study is to determine the prevalence of antibodies directed against HAV and HBV in patients with CHC, analyze demographic and risk factors associated with this prevalence, and develop a cost-effective vaccination strategy. METHODS: We reviewed records from 1092 CHC patients. Demographics and information regarding risk factors were obtained by history and questionnaire administered to all patients. The costs of vaccination and antibody testing were determined, based on standard laboratory and clinic charges at our institution. HAV and HBV markers were correlated to race, age, and risk factors. RESULTS: Of the total population studied (n = 1092), 72% were African-Americans, 27% white, and 1% others. Of 671 CHC patients tested for anti-HAV IgG, 252 (38%) were positive. Of 743 CHC patients tested for HBV antibodies (anti-hepatitis B core IgG or anti-hepatitis B surface), 494 (67%) were positive. African-Americans are more likely to have antibodies to HAV and HBV (67% and 75%, respectively) compared to whites (27% and 20%). The prevalence of anti-HAV was 76% in patients >60 yr, 34% in the 40- to 60-yr-old age group, and 21% in patients <40 yr. The highest prevalence of HBV antibodies was found in patients between the ages of 40 60 yr. No HCV risk factors were associated with increased HAV risk. In CHC patients with HBV antibodies, however, illicit injection drug use was the predominant risk factor. CONCLUSIONS: The prevalence of anti-HAV in patients with CHC was found to be similar to that of the general population in the United States (33% according to recent Centers for Disease Control data), consistent with the hypothesis that the two infections do not share risk factors. Because the prevalence of HAV immunity is low in CHC patients <40 yr, empiric HAV vaccination is cost effective. If two doses of vaccine are to be given, however, antibody testing of all HCV patients is indicated. In the subset of patients >60 yr of age or who are African-American, where the prevalence of HAV exposure is considerably higher, it would be cost effective to check the antibody ($36.00), before vaccination ($97.00). The prevalence of HBV antibodies, however, is significantly increased in patients with CHC compared with the general population (5.3% per the Centers for Disease Control), likely as a result of exposure to similar parenteral risk factors. HBV antibody testing ($26.00 per test) should, therefore, be undertaken in all CHC patients who are hepatitis B surface antigen negative, as this approach is cost-effective compared to empiric HBV vaccination ($438.00 for a three injection course). PMID- 11280566 TI - Thiamine treatment of chronic hepatitis B infection. AB - OBJECTIVE: Chronic hepatitis B is an international health concern that causes cirrhosis, hepatocellular carcinoma, liver failure, and death. Current treatment options are expensive and associated with side effects; however, indirect evidence suggests a relationship between relative thiamine deficiency and chronic hepatitis B infection. METHODS: The authors present three case studies wherein multiple crossovers of daily thiamine administration were used to evaluate a hypothesized association between thiamine treatment and aminotransferase levels. RESULTS: In each case study, thiamine administration was associated with reduction in aminotransferase levels and the fall of HBV DNA to undetectable levels. Analyses by t test demonstrated a statistically significant reduction in aminotransferase levels in all three cases. CONCLUSIONS: The relationship between thiamine administration and chronic hepatitis B infection warrants further study. If proven effective in reducing liver damage or inducing remission of the hepatitis B virus in larger trials, thiamine will offer obvious advantages over the current treatments for chronic viral hepatitis B infection. PMID- 11280567 TI - Evidence of an increased nitric oxide production in primary biliary cirrhosis. AB - OBJECTIVE: Although possible implications of nitric oxide in the pathophysiology of liver cirrhosis have been extensively studied, until now few articles have addressed the assessment of nitric oxide production in primary biliary cirrhosis. This study was directed to evaluate circulating nitrosyl-hemoglobin levels as well as neutrophil elastase and soluble adhesion molecule concentrations in this condition, by assuming these parameters as possible markers of either inflammatory response or neutrophil activation. METHODS: Laboratory investigations were performed in 30 patients with primary biliary cirrhosis, in 13 patients with postviral and/or alcoholic cirrhosis, and in a group of eight subjects with chronic hepatitis. RESULTS: Although no difference was detected with respect to chronic hepatitis subjects, higher levels of nitrosyl-hemoglobin adducts were found in primary biliary cirrhosis patients than in postviral or alcoholic cirrhotics and in normal subjects (3.55+/-1.75 arbitrary units vs 1.95+/-0.57 and 0.84+/-0.34, p = 0.0004 and p < 0.0001, respectively). Similarly, more elevated concentrations of neutrophil elastase (213.7+/-192.0 microg/L vs 51.1+/-34.3 and 38.0+/-11.5, p < 0.0001 and p < 0.0001, respectively) as well as of soluble forms of intercellular adhesion molecule 1 and endothelial-leukocyte adhesion molecule 1 were shown in primary biliary cirrhosis patients than in subjects with cirrhosis of other etiologies and in controls. CONCLUSIONS: Highly enhanced nitric oxide production in primary biliary cirrhosis could be related to the development of strong inflammation and at least partially to neutrophil activation, thus suggesting a putative role of these cellular mediators in the development of liver damage owing to their ability to synthesize and release a wide variety of important factors, including elastase and nitric oxide. PMID- 11280568 TI - The quality of care in Barrett's esophagus: endoscopist and pathologist practices. AB - BACKGROUND: The diagnosis of Barrett's esophagus (BE) has important psychological and economic implications. Although accepted standards for endoscopic biopsy methods and pathological interpretation for BE exist, adherence to these standards as a measure of the quality of care in BE has not been evaluated. Our aim was to assess the quality of care in BE by evaluating the process of care and adherence to accepted standards of practice. METHODS: Explicit process-of-care criteria were developed using a systematic literature review and expert opinion in four domains of care: the quality of biopsy methods, the adequacy in identifying endoscopic landmarks, endoscopist-pathologist communication, and pathological interpretation and reporting. We reviewed all endoscopy and pathology reports of BE patients at two institutions from 1994-1997. An academic medical center (N = 237) with staff endoscopists and an academically affiliated community hospital (N = 100) with private-practice endoscopists were analyzed. RESULTS: Physicians showed the highest adherence to accepted standards of care in the "adequacy of identifying landmarks" and "endoscopist-pathologist communication" domains, with a > or =70% adherence rate in most criteria. Conversely, physicians demonstrated the poorest adherence with the "quality of biopsy methods" and "pathologist interpretation and reporting" domains, with adherence rates frequently <60%. Significantly, biopsies were taken in the presence of visible esophagitis 35% of the time. Performance on several of the quality indicators varied significantly by the practice setting. CONCLUSIONS: We have identified several opportunities for quality improvement efforts. In every domain, there is room for improvement, particularly in the quality of biopsy methods. As initiatives to screen the large population of gastroesophageal reflux disease patients for BE may be imminent, the time is now to define the critical process-of-care measures to minimize the risk of overdiagnosis and inadequate endoscopic surveillance. PMID- 11280569 TI - Gender differences in colorectal polyps and tumors. AB - OBJECTIVES: To use a national endoscopy database (CORI) to determine 1) whether gender differences are noted in the prevalence and location of polyps and tumors; 2) whether women have a higher rate of right-sided polyps or tumors; and 3) whether age influences these results. METHODS: CORI database from April 1, 1997 to February 19, 1999, captured in a computer-generated report, was analyzed. Polyps for this study were defined as sessile or pedunculated and as >9 mm. Tumors were defined as lesions characteristic of adenocarcinoma (mass, apple core). Pure right-sided colon (PRS) was defined as cecum, ascending, hepatic flexure; right-sided as PRS plus the transverse colon; and left-sided as the splenic flexure, descending, sigmoid and rectum. RESULTS: Men have a greater risk of polyps [odds ratio (OR), 1.5] and tumors (OR, 1.4) than women. The risk of finding polyps and tumors at colonoscopy increases with age, with the highest risk noted in those >69 yr of age relative to patients <50 yr of age (polyps, OR = 2.7; tumors, OR = 4.0). Right-side polyps and pure right-sided polyps as defined by the study design were noted to be more frequent than left-sided polyps in patients >60 yr of age. Women have a greater risk of developing pure right sided polyps (OR, 1.2), tumors (OR, 1.6) and right-sided tumors (OR, 1.5) than men. CONCLUSIONS: Men have a higher prevalence of colon polyps and tumors than women. A progressive risk of polyp or tumor formation is noted with aging. Women had a greater number of pure right-sided polyps and tumor development. Colonoscopy is needed to correctly diagnose an increasing prevalence of right sided pathology in the elderly. PMID- 11280570 TI - Differing attitudes toward virtual and conventional colonoscopy for colorectal cancer screening: surveys among primary care physicians and potential patients. AB - OBJECTIVES: To investigate attitudes among primary care physicians and potential patients concerning "virtual" and conventional colonoscopy for colorectal cancer screening. METHODS: We sent 1000 questionnaires to primary care physicians by electronic or postal mail and administered 400 to potential patients. Questionnaires contained progressively detailed information about the tests and asked for choices based on information presented. RESULTS: One hundred eight eight primary care physicians and 323 potential patients were included. Results indicated the following: 76.6% of potential patients and 47.3% of physicians initially preferred virtual colonoscopy because of its noninvasive nature; 23.6% of potential patients and 52.9% of physicians valued the ability of conventional colonoscopy to visualize the mucosa directly; and 67.4% of potential patients and 51.6% of physicians preferred virtual colonoscopy because it does not require sedation. Considering all information, most potential patients preferred virtual to conventional colonoscopy (60.2% vs 25.7%), whereas more physicians preferred conventional to virtual colonoscopy (44.9% vs 30.3%). Additionally, 82.3% of potential patients would comply more with recommendations for colorectal cancer screening, and 61.7% of physicians would refer more patients for screening, if virtual colonoscopy was available. CONCLUSIONS: Potential patients preferred virtual to conventional colonoscopy, whereas physicians favored conventional colonoscopy. Physicians placed more importance on the ability of conventional colonoscopy to visualize the mucosa directly, the opportunity for therapy, and cost. Potential patients were more encouraged than physicians by the availability of virtual colonoscopy for improving participation in colorectal cancer screening. PMID- 11280571 TI - Leech jar. PMID- 11280572 TI - Cytomegalovirus colitis complicating ulcerative colitis in the steroid-naive patient. AB - We report a case of cytomegalovirus (CMV) presenting as acute refractory colitis in a patient with a pre-existing 14-month history of ulcerative colitis (UC) who had never previously been treated with corticosteroids or immunosuppressants. A review of existing literature and previous cases of patients with coincident CMV and UC are examined, stratifying these cases based upon absence or presence of corticosteroid use. To date, only five previous case reports of CMV colitis in patients naive to corticosteroids have been described, and only one previous case has had UC diagnosed over 4 wk before the development of CMV colitis. We further discuss the relationship between these two diseases as well as the diagnosis, treatment, patient characteristics, and outcome of CMV infection of the colon in patients with underlying UC. We discuss the need to consider the diagnosis of CMV colitis in patients with refractory UC who are not receiving corticosteroid treatment as well as those who are refractory and are being treated with immunosuppressants and/or corticosteroids. PMID- 11280573 TI - Electrical nerve stimulation in the management of enterocutaneous low-output fistulas: a report of two cases. AB - Two patients with low-output enterocutaneous fistulas after surgery were treated with electrical nerve stimulation (ENS). Ultrasonography was useful for the application of this treatment method and for the charting of its progress. Fistula output diminished rapidly in both cases, and the closure of the track was achieved after several sessions of ENS. The procedure is simple and safe and is suggested as an option for the treatment of low-output enterocutaneous fistulas. PMID- 11280574 TI - Dieulafoy's lesion of the small bowel causing massive gastrointestinal bleeding: two case reports and literature review. AB - Dieulafoy's lesions are an often unrecognized cause of obscure, massive GI hemorrhage. Their diagnosis may elude conventional investigations, including upper and lower endoscopy, arteriography, and even laparotomy. In this paper, we report two cases of small-bowel Dieulafoy lesions. The first, a jejunal lesion, occurred in a young patient and was discovered at laparotomy. The second was an ileal Dieulafoy's malformation in an older patient. An intraoperative endoscopy with surgical guidance may be needed for definitive localization of this lesion. PMID- 11280575 TI - Plexosarcoma: endoscopic ultrasound and electron-microscopic characteristics of a stromal tumor. AB - Endoscopic ultrasound is useful for managing submucosal masses; however, some of these lesions can be difficult to classify except with full histological and electron microscopic evaluation. A 72-yr-old woman was seen with upper GI bleeding. Endoscopy showed a 1.7-cm sessile ulcerated submucosal mass in the duodenal bulb. Endoscopic ultrasound revealed an echolucent submucosal mass arising from the fourth echolayer, the muscularis propria of the duodenal wall. These findings suggested that the lesion was a leiomyoma. The patient eventually had the lesion resected because of recurrent bleeding. Histologically it was a spindle cell tumor that on electron microscopy showed neuronal elements consistent with a plexosarcoma, or gastrointestinal autonomic nerve tumor. These lesions account for some one third of all gastrointestinal stromal tumors. Despite their low grade malignant histologic appearance, local recurrence or hepatic metastases occur in about 70% of patients. PMID- 11280576 TI - LKM3 autoantibodies in hepatitis C cirrhosis: a further phenomenon of the HCV induced autoimmunity. AB - Chronic hepatitis C is frequently associated with laboratory markers-including LKM1 autoantibodies--of autoimmunity. A 62-yr-old woman with hepatitis C cirrhosis presented autoantibodies against liver and kidney microsomal proteins. By further evaluation of autoantibodies using ELISA and immunoblotting LKM1 and LKM3 autoantibodies could be revealed. The target antigen of LKM3 autoantibodies proved to be UGT-1.1 isoenzyme. In the absence of chronic hepatitis D infection or autoimmune hepatitis type 2, this is the first case that reports the occurrence of LKM3 autoantibodies in HCV-induced chronic liver disease. PMID- 11280577 TI - Upper endoscopy and glucagon: a new technique in the management of acute esophageal food impaction. PMID- 11280578 TI - NSAID colitis--resolving a diagnostic dilemma? PMID- 11280579 TI - In pursuit of doctor Whipple's nemesis. PMID- 11280580 TI - Albumin: a look at the evidence. PMID- 11280581 TI - Transportability and reproducibility of the AST/ALT ratio in chronic hepatitis C patients. PMID- 11280582 TI - Clinical utility of the AST/ALT ratio in chronic hepatitis C. PMID- 11280583 TI - Re: Koshy et al.--The use of propofol versus midazolam. PMID- 11280584 TI - Optimal strategy of treatment of Helicobacter pylori infection: renaissance of antisecretory drugs? PMID- 11280585 TI - Spontaneous elimination of serum HCV-RNA after total gastrectomy for early gastric cancer in a patient with chronic hepatitis C. PMID- 11280586 TI - Successful treatment of esophageal perforation with a removable self-expanding plastic stent. PMID- 11280587 TI - Periductal fibrosis of biliary atresia--is it not caused by fat-storing cells? PMID- 11280588 TI - Re: Brunt et. al.--Histological evaluation of iron in liver biopsies: relationship to HFE mutations. PMID- 11280589 TI - Fluticasone in eosinophilic corrugated ringed esophagus. PMID- 11280590 TI - Mast cells: do they really have a role in disturbed bowel habits of IBS patients? PMID- 11280591 TI - Endoscopic appendectomy for a carcinoid tumor of the appendix. PMID- 11280592 TI - Peptic ulcer, or should it be called Marshall's disease. PMID- 11280594 TI - Pellagra in an immunocompetent patient with cytomegalovirus colitis. PMID- 11280593 TI - Malarial pancreatitis: report of two cases and review of the literature. PMID- 11280595 TI - Haloperidol and benztropine interaction presenting as acute intestinal pseudo obstruction. PMID- 11280596 TI - Posttraumatic pancreatitis. PMID- 11280597 TI - Transnasal placement of percutaneous endoscopic gastrostomy with a pediatric endoscope in oropharyngeal obstruction. PMID- 11280598 TI - Hyalinising spindle cell tumour with giant rosettes: report of a case with unusual features including original histological and ultrastructural observations. AB - Hyalinising spindle cell tumour with giant rosettes (HSCTGR) is an uncommon, recently described low-grade sarcoma which shares many histological features with low-grade fibromyxoid sarcoma (LGFMS). We report a case of HSCTGR occurring in the deep soft tissues of the thigh of a 46-year-old woman, that presented as a slowly growing, painless mass. Microscopically the tumour was composed of spindled stromal cells amongst which were scattered so-called collagen rosettes. The distinctive feature of this case was the previously unreported finding of lymphoid cells of T-cell phenotype admixed with fibrohistioctyic cells in the cellular cuff surrounding the collagenous core of the rosettes. The case was further unusual in that it included focal areas of increased cellularity with a mitotic count of up to three per 10 high-power fields. While the latter feature has been associated with increased recurrences and metastases in LGFMS, it is not known whether the significance is similar in HSCTGR. The spindled stromal cells showed ultrastructural features of poorly differentiated fibroblasts, while those at the edges of the rosettes showed features of altered fibroblasts, some with a fibrohistiocytic appearance. These findings support the interpretation that HSCTGR forms part of the spectrum of sarcomas showing fibroblastic differentiation. PMID- 11280599 TI - Solid variant of alveolar rhabdomyosarcoma with unbalanced t(2;13) and hypotetraploidy, without MYCN amplification. AB - The histological subtype of alveolar rhabdomyosarcoma (AR) is characterised by the cytogenetic translocation t(2;13)(q35;q14) in approximately 70% of cases, a rearrangement rarely present in the embryonal rhabdomyosarcoma (ER) subtype. The MYCN gene is amplified in some cases of AR. We present a young man with an unusual pattern, namely solid variant of AR with hypotetraploidy and the t(2;13) in an unbalanced form. The MYCN gene was not amplified on FISH, but showed increased copy number, consistent with ploidy. PMID- 11280600 TI - CNS findings in iniencephaly: case report and literature review. AB - A male iniencephalic foetus of about 25 weeks gestation is described. The baby was born to a 22-year-old mother, who presented with abdominal pain of 1 day duration. An ultrasound scan at that time showed multiple foetal anomalies and the pregnancy was terminated. A stillborn baby was delivered. At autopsy, characteristic findings of iniencephaly were seen in the dysmorphic foetus as well as multiple structural abnormalities. The autopsy results, with emphasis on the neuropathological findings, are described and various hypotheses of the pathogenesis of iniencephaly are discussed with reference to theories of embryological development and other cases in the literature. PMID- 11280601 TI - Fine needle aspiration cytology of pulmonary lesions: a reliable diagnostic test. AB - The objective of this study was to determine the accuracy of image-guided fine needle aspiration cytology (FNAC) in the diagnosis of pulmonary lesions. A retrospective study was undertaken of 286 patients with 288 lesions, who underwent a total of 302 procedures. The FNAC diagnoses were reported as malignant, suspicious, atypical, benign or non-diagnostic. Subsequently the FNAC diagnoses were correlated with either the histological or clinical diagnoses. Of the 288 lesions, 64.6% were reported on FNAC as malignant, 2.1% suspicious, 2.4% atypical, 20.8% benign and 10.1% nondiagnostic. On review of the suspicious, atypical, selected benign cases and non-diagnostic FNAC by an independent pathologist there was agreement with the original FNAC diagnosis in all cases. All of 186 malignant FNAC diagnoses were confirmed malignant either clinically or on subsequent histology. Four of the six suspicious FNAC diagnoses had a malignant outcome, one patient had organising pneumonia on excision biopsy and one was lost to follow up. Six of the seven atypical FNAC diagnoses were confirmed on histology as malignant, while one lesion resolved spontaneously. Fifty-two of 60 benign FNAC diagnoses were confirmed benign either clinically or on histology. Seven of the lesions diagnosed as benign on FNAC were proven to be malignant. One patient with a benign FNAC diagnosis was lost to follow-up. Ten of the 29 non-diagnostic FNAC group were later shown on clinical or histological follow up to be malignant. This study shows that image guided FNAC for the diagnosis of malignant pulmonary lesions has a sensitivity of at least 92% and a specificity of at least 96%. It is a reliable diagnostic test although its accuracy is limited by technical difficulties in obtaining an adequate sample. PMID- 11280602 TI - Investigation of human papillomavirus in transitional cell carcinomas of the urinary bladder in South Africa. AB - AIM: To investigate the prevalence of human papillomavirus in transitional cell carcinoma of the urinary bladder in South Africa. METHODS: Ninety-one archival samples of bladder transitional cell carcinoma were subjected to polymerase chain reaction (PCR) and non-isotopic in situ hybridisation (NISH) for the detection of human papillomavirus 6, 11, 16, 18, 31, and 33 genotypes. RESULTS: HPV was detected in only one case with PCR. HPV was not detected in any of the cases subjected to the NISH system. CONCLUSION: This study shows that although HPV has been shown to be associated with uterine cervical and esophageal squamous cell carcinomas in South Africa, this virus is not present in the transitional cell carcinoma of the urinary bladder in this geographical location. It is suggested that other factors, including nitrosamine exposure, p53 mutation, and additional unknown chromosomal events, may play a role in the carcinogenesis of this neoplasm in the bladder. PMID- 11280603 TI - Immunohistochemical expression of hormone receptors in invasive breast carcinoma: correlation of results of H-score with pathological parameters. AB - The results of H-scores of oestrogen and progesterone receptor (ER and PR) expression in 150 invasive breast cancers were correlated with conventional pathological prognostic parameters: tumour size, histological grade and subtype, lymph node status, lymphovascular invasion, Nottingham Prognostic Index (NPI) and pathological stage. ER and PR status was determined by immunohistochemical staining of sections cut from archival paraffin-embedded tissue blocks. We defined positive receptor expression as a H-score of 50 and above. Our findings revealed ER and PR positivity in 98 (65%) and 52 (35%) cases, respectively. Fifty one (34%) ER-positive cases also showed PR expression, while 51 (34%) tumours were negative for both ER and PR. Positive expression for ER and PR was significantly correlated with histological grade (P < 0.0005), mitotic score (P < 0.05) and nuclear pleomorphism (P < 0.05). When we used the relatively simpler method of a cut off of at least 10% tumour cell nuclear staining of moderate or greater intensity as positive receptor status, we found that it agreed well with results of the H-score, a more quantitative method of assessment. PMID- 11280604 TI - Pelvic ependymoma arising from the small bowel. AB - A 37-year-old woman underwent resection of an abdominal tumour which was adherent to the wall of the ileum. The diagnosis of an ependymoma was supported by evidence of typical perivascular pseudorosettes which stained positive for glial fibrillary acidic protein and contained abundant intermediate filaments within the elongated processes by electron microscopy. Flow cytometric study showed a diploid population of tumour cells. This is the first case of an ependymoma arising from the small bowel without any connection to the genital tract, the omentum or with the sacroccygeal area. As is the case with other unusual and ectopic localisations of ependymomas, prognosis of this tumour is difficult to evaluate. PMID- 11280605 TI - Rapid radiation-induction of ATM protein levels in situ. AB - Ataxia-telangiectasia (A-T) is characterised by hypersensitivity to ionising radiation (IR), immunodeficiency, neurodegeneration and predisposition to malignancy. Mutations in the A-T gene (ATM) often result in reduced levels of ATM protein and/or compromise ATM function. IR induced DNA damage is known to rapidly upregulate ATM kinase activity/phosphorylation events in the control of cell cycle progression and other processes. Variable expression of ATM levels in different tissues and its upregulation during cellular proliferation indicate that the level of ATM is also regulated by mechanisms other than gene mutation. Here, we report on the IR induction of ATM protein levels within a number of different cell types and tissues. Induction had begun within 5 min and peaked within 2 h of exposure to 2 Gy of IR, suggesting a rapid post-translational mechanism. Low basal levels of ATM protein were more responsive to IR induction compared to high ATM levels in the same cell type. Irradiation of fresh skin biopsies led to an average three-fold increase in ATM levels while immunohistochemical analyses indicated "low expressing" cells within the basal layer with ten-fold increases in ATM levels following IR. ATM "high expressing" lymphoblastoid cell lines (LCLs) which were initially resistant to the radiation induction of ATM levels also became responsive to IR after ATM antisense expression was used to reduce the basal levels of the protein. These results demonstrate that ATM is present in variable amounts in different tissue/cell types and where basal levels are low ATM levels can be rapidly induced by IR to saturable levels specific for different cell types. ATM radiation-induction is a sensitive and rapid radioprotective response that complements the IR mediated activation of ATM. PMID- 11280606 TI - Urine laminin and kallikrein, markers of tubulointerstitial damage in experimental protein overload on pre-existing renal damage. AB - We studied the response of urinary protein overload on preexisting tubulointerstitial nephritis (TIN), which was induced in male Sprague Dawley rats by hexachloro-1,3-butadiene (HCBD). Five days after the development of TIN, puromycin aminonucleoside (PAN) was administered to induce urinary protein overload. Urinary laminin and kallikrein were measured. Urine specimens were collected daily for 14 days and on day 21; and tissue specimens were collected on days 1, 4, 7, 10, 14 and 21. Urinalysis was correlated with the renal pathology at the light microscopic level. Laminin excretion was increased on day 4; one day before total protein, indicating damage to the basement membrane. Kallikrein levels also fell early indicating distal tubular damage. There is clear evidence that urine protein overload in a previously damaged kidney with tubulointerstitial injury leads to accelerated and more severe renal damage. Laminin and kallikrein are early and sensitive markers of renal injury. PMID- 11280607 TI - Compound heterozygosity for haemochromatosis gene mutations and hepatic iron overload in allogeneic bone marrow transplant recipients. AB - Iron overload has been proposed as a cause of liver dysfunction after BMT Factors which could be relevant to iron overload include the number of red cell transfusions and mutations within the haemochromatosis gene (HFE). Two point mutations, Cys282Tyr and His63Asp, have been described within HFE. Cys282Tyr homozygosity is associated with haemochromatosis; the effect of compound heterozygosity, Cys282Tyr/His63Asp, on iron status is variable. We analysed HFE status in 52 allograft patients surviving more than 6 months. Compound heterozygosity was identified in three patients (Cases 1-3). Iron status and liver function were evaluated and, in Cases 1 and 2, liver histology and iron content as well. Case 3 who received 12 units of red cells had a normal ferritin and liver function. Cases 1 and 2 received 29 and 59 units, respectively, and had high serum ferritins and transferrin saturations, abnormal liver function and significant hepatic iron overload on biopsy. Iron overload in Case 1 patient progressed in the context of GVHD and in the absence of further transfusion, suggesting that liver GVHD may increase hepatic iron accumulation. These cases demonstrate the variable phenotypic expression of HFE compound heterozygosity in BMT recipients, which may be only partly explained by transfusional iron loading. Venesection or chelation therapy should be considered in patients with coexistent hepatic GVHD and iron overload. PMID- 11280608 TI - Editorial: the changing face of pathology. AB - As the journal advances into its 33rd volume of publication and into the new millennium, significant developments in information technology have allowed restructuring of the format, review and publishing procedures of the journal, Pathology. This in turn enables rapid publication of timely and significant articles of interest to both diagnostic and research pathologists. There is an emphasis to develop the educational and professional development aspects of the journal while maintaining a rigorous peer review process to ensure publication of high quality research and review articles. A number of changes that will facilitate these processes are outlined. The journal is also available on-line on the web in PDF format and there are considerations made to further exploit this avenue of communication for the publication of supplementary material. The Editor and Editorial Board encourages Fellows of the Royal College of Pathologists, readers and authors to communicate their views on these and other future developments with the Editorial Office. PMID- 11280609 TI - High level excretion of Norwalk-like virus following resolution of clinical illness. AB - We report the case of an elderly woman excreting high levels (about 5 x 10(5) virions per gram of faeces) of Norwalk-like virus (NLV) in the absence of any clinical symptoms of gastroenteritis. Analysis by reverse transcription, polymerase chain reaction and DNA sequencing was carried out on a 342-nucleotide region of open reading frame 1. This indicated that the NLV belonged to genogroup 2 and was more closely related to the Camberwell subgroup, the most common circulating in southeast Australia at present, than to the Norwalk and Mexico viruses. PMID- 11280610 TI - Multicentre study of the in vitro activity of cefepime, a broad-spectrum cephalosporin, compared to other broad-spectrum agents. AB - The in vitro activity of cefepime was compared to that of a range of other broad spectrum agents, using a gradient diffusion MIC method, against 995 recent clinical isolates of Enterobacteriaceae, Pseudomonas aeruginosa, other nonfermentative gram-negative bacilli, staphylococci (except oxacillin-resistant Staphylococcus aureus), streptococci, enterococci, and aerobic gram-positive bacilli. Cefepime had excellent activity against Enterobacteriaceae, including eight presumptive extended-spectrum beta-lactamase producers and 33 stably derepressed mutants of natural cephalosporinases. Activity against Pseudomonas aeruginosa was similar to ceftazidime and superior to cefpirome. Its activity against gram-positive cocci was also good, being more active against staphylococci and only slightly less active against streptococci than ceftriaxone. Cefepime maintained activity against bacteria resistant to aminoglycosides and ciprofloxacin. Enterococci, Bacillus species, Burkholderia cepacia and Stenotrophomonas maltophilia were predicably resistant. Cefepime has a spectrum of activity almost as broad as that of the carbapenems. PMID- 11280611 TI - Rapid identification of mycobacterium tuberculosis complex from Bactec 12B broth cultures originating from non-respiratory secretion specimens: comparison between Accuprobe and in-house polymerase chain reaction. AB - The rapid identification of Mycobacterium tuberculosis complex from 161 consecutive positive Bactec 12B cultures originated from non-respiratory secretion specimens was evaluated by comparing Accuprobe and an in-house polymerase chain reaction method. Using conventional biochemical method as the gold standard, 85.7% of the isolates (n = 138) were identified as Mycobacterium tuberculosis. The sensitivities of Accuprobe and the in-house-polymerase chain reaction method in identifying Mycobacterium tuberculosis complex were 92.0% (127/138) and 93.5% (129/138), respectively. Specificities of both methods being 87.0% (20/23 for both tests). Using a batch system of performing identification once weekly, the average turnaround time for Accuprobe, from the time of primary inoculation, was 27 days, which was comparable with that of in-house polymerase chain reaction, which was 29 days. The average time required for a confirmatory result based on conventional biochemical identification method was 84 days. The estimated reagent cost of in-house polymerase chain reaction was only one-fifth of that of Accuprobe. Therefore, the choice of methodology will depend on the individual laboratory's availability of resources and clinical demand. In the event of discrepant results between rapid identification methods and conventional methods, histological examination and clinical assessment can provide guidance as to the nature of the infection. PMID- 11280612 TI - Practical evaluation of molecular subtyping and phage typing in outbreaks of infection due to Salmonella enterica serotype typhimurium. AB - Identification and control of food-poisoning outbreaks due to salmonellosis depend on prompt microbiological diagnosis and subtyping to identify the causative strain. In Australia, Salmonella enterica subspecies enterica serotype typhimurium (S. typhimurium) is responsible for 40-70% of cases of human salmonellosis. Phage typing is the usual method of subtyping S. typhimurium, but on its own, has limitations. We compared it with three molecular subtyping methods using 100 isolates of S. typhimurium, representing four different phage types (PT 1, 9, 126 and 135) and comprising 74 isolates from three presumed outbreaks, 25 isolates from sporadic cases of salmonellosis and S. typhimurium ATCC 10428 (phage type 126). The isolates were divided into 11 subtypes by IS200 restriction fragment length polymorphism (RFLP) typing, four each by ribotyping and pulsed-field gel electrophoresis (PFGE) and 17 distinct strains using a combination of phage and molecular typing. Isolates from two presumed outbreaks were resolved into multiple strains, possibly explaining the failure to identify a common source for either during the original investigations. IS200 RFLP analysis was the most discriminatory and reproducible typing method. Several strains were identifiable within and shared between phage types 1, 9 and 126. Phage and IS200 RFLP typing together, would provide improved definition of S. typhimurium outbreaks. PMID- 11280613 TI - Timely topic: anaplastic lymphoma kinase (ALK) spreads its influence. AB - Anaplastic lymphoma kinase (ALK) is normally not expressed in human tissues except selected sites in the nervous system. Its expression and constitutive activation as a result of a chromosomal translocation involving 2p23 plays a pivotal role in the genesis of anaplastic large cell lymphoma. ALK expression has been instrumental in defining a homogeneous subset from the category of anaplastic large cell lymphoma, characterised by occurrence in young patients, primary systemic presentation, favorable prognosis, a broad morphological spectrum, nuclear and/or cytoplasmic immunostaining for ALK protein, and a number of possible fusion partner genes such as NPM, ATIC, TFG, TPM3 and CLTCL. Recently ALK has been implicated in the genesis of another tumour type, the inflammatory myofibroblastic tumours. The ALK-positive examples occur in children and young adults, involving a variety of sites, such as the abdomen, mesentery, liver, bladder, mediastinum, lung and bone. The partner genes identified in some cases are TPM3 (tropomyosin 3) and TPM4 (tropomyosin 4). These molecular findings also further support the neoplastic nature of at least a subset of inflammatory myofibroblastic tumours. PMID- 11280614 TI - The smad proteins and TGFbeta signalling: uncovering a pathway critical in cancer. AB - The critical role of TGFbeta in development and growth control is well established but the signalling pathway has only recently been elucidated. Identification of the Smads as TGFbeta's intracellular signalling mediators has led to an explosion of information on a novel signalling network that links the cell surface to the nucleus. Many cancers develop resistance to the growth inhibitory effects of TGFbeta and mutations in signalling pathway components have been discovered that may underly tumour progression. PMID- 11280615 TI - The role of TGFbeta in human cancers. AB - Transforming growth factor beta (TGFbeta) is secreted as a large latent precursor from both normal and transformed cells which needs to be activated for biological activity. The active TGFbeta binds either directly to TbetaR-II or indirectly by binding to beta-glycan which then presents the TGFbeta to TbetaR-II. Formation of the TGFbeta-TbetaR-II complex rapidly leads to phosphorylation of TbetaR-I. TbetaR-I, in turn, phosphorylates receptor-specific Smads and induces their translocation into the nucleus. TGFbeta is able to act as a growth stimulator or inhibitor and elicits a broad spectrum of biological effects on various cell types. However, these cells may lose their sensitivity and responsiveness to TGFbeta. Down-regulation or loss of functional receptors, aberrant signal transduction pathways due to Smad mutations, loss of the cell's ability to activate latent TGFbeta, loss of the peptide itself or functional genes that control the transcription and translation of TGFbeta may contribute to development of cancer. PMID- 11280616 TI - Statistics in the pathology laboratory: characteristics of diagnostic tests. AB - Sensitivity, specificity and receiver operating characteristic (ROC) curves all provide information about the ability of a diagnostic test to provide useful information in the assessment of disease. They are discussed in this review along with the importance of estimates of precision. PMID- 11280617 TI - Adenomyolipoma of the uterus: a case report. AB - Adenomyolipoma of the uterus is a rare, benign, polypoid lesion considered to be of hamartomatous origin or represent an unusual type of benign Mullerian mixed tumour with a heterologous element. The authors present a case of uterine adenomyolipoma and discuss its pathogenesis. A 62-year-old woman complained of lower abdominal pain and postmenopausal bleeding. Imaging techniques revealed a solid ovarian mass and a polypoid intrauterine lesion. The frozen section diagnosis of the ovarian mass was a thecoma. A total hysterectomy and bilateral salpingo-oophorectomy were performed. On gross examination a pedunculated, polypoid lesion of 7x4.5x3cm was found in the uterine cavity. Microscopically, the polypoid lesion contained both epithelial and mesenchymal elements. The epithelial elements were endometrial glands of various size, formed by proliferative endometrial cells. The mesenchymal elements were composed of endometrial stroma, smooth muscle and mature adipocytes. Both the epithelial and the mesenchymal elements showed a benign appearance, were intermingled with each other and periglandular stromal condensation was absent. The lesion had an irregular surface. Microscopic diagnosis was an adenomyolipoma. The peculiar shape and microscopic features of this lesion suggested that it was a variant of benign Mullerian mixed tumour. PMID- 11280618 TI - Partial liquid ventilation: is there a niche? PMID- 11280619 TI - Comparison of the cerebral effects of dopamine and norepinephrine in severely head-injured patients. AB - OBJECTIVE: To compare the cerebral effects of dopamine and norepinephrine after severe head injury. DESIGN: Prospective, clinical study. SETTING: Surgical intensive care unit in a university hospital. PATIENTS: Nineteen patients with severe head-injuries already requiring vasopressor therapy. Group 1: patients receiving dopamine (n = 9); group 2: patients receiving norepinephrine (n = 10). INTERVENTION: Vasopressor therapy was switched from dopamine to norepinephrine in group 1 and from norepinephrine to dopamine in group 2, maintaining the same mean arterial pressure (MAP). MEASUREMENTS AND RESULTS: MAP, intracranial pressure (ICP), jugular venous oxygen saturation (SjvO2), transcranial Doppler mean velocity in the middle cerebral artery (Vm), and transoesophagal Doppler aortic output (AO) were evaluated under dopamine and norepinephrine. Means for each group were compared with the paired Student's t-test. For the same MAP, ICP was significantly higher with dopamine than norepinephrine in both groups (respectively, group 1: 26 +/- 11 vs 23 +/- 11 mmHg, P < 0.005; group 2: 39 +/- 13 vs 31 +/- 9 mmHg, P < 0.005). SjvO2, Vm, and AO did not change significantly between treatments. The ICP variation between treatments was not correlated with the variation of any other measured parameter. The ICP variation between treatments was significantly higher in group 2 than group 1, which could be explained by autoregulation mechanisms. CONCLUSIONS: For the same MAP, ICP was significantly higher with dopamine than norepinephrine with no argument supporting an increase of cerebral blood flow. PMID- 11280620 TI - Evaluation of two processed EEG analyzers for assessment of sedation after coronary artery bypass grafting. AB - OBJECTIVES: Processed EEG monitoring has been suggested for sedation depth evaluation in intensive care unit (ICU) patients. The present study investigated the efficacy of two processed EEG monitors using SEF90% or SEF95% and BIS to differentiate between conscious (Ramsay score 4) and unconscious sedation (Ramsay score 6). DESIGN AND SETTING: Prospective, randomized trial in a surgical ICU of a university teaching hospital. PATIENTS: Patients recovering from elective coronary bypass grafting. INTERVENTION: One of two EEG analyzers was installed (A: Aspect A-1000 measuring SEF95% and BIS; D: Drager pEEG measuring SEF90%). At ICU admission unconscious sedation (Ramsay score 6), and at three 30-min intervals conscious sedation (Ramsay score 4) were investigated. MEASUREMENTS AND RESULTS: Fourteen patients were monitored by A and 14 by D. The interindividual variability (coefficient of variation 32-69 %) was large for all three processed EEG methods. SEF90% of analyzer D and BIS of analyzer A showed a statistically significant difference between unconscious and conscious sedation (11 +/- 3 and 17 +/- 6 Hz, p = 0.005; 74 +/- 10 and 83 +/- 10, p = 0.02). Positive and negative predictive values for SEF90% of analyzer D (0.57, 95% CI 0.34-0.77; and 0.92, 95% CI 0.64-0.99) and BIS of analyzer A (0.55, 95 % CI 0.32-0.76; and 0.87, 95 % CI 0.60-0.98) were too low for discrimination between conscious and unconscious sedation. CONCLUSIONS: The use of processed EEG monitoring cannot be recommended for assessing sedation depth after cardiac surgery. PMID- 11280621 TI - Pharmacokinetics of sequential intravenous and enteral fluconazole in critically ill surgical patients with invasive mycoses and compromised gastro-intestinal function. AB - OBJECTIVES: (1) To determine the pharmacokinetics of sequential intravenous and enteral fluconazole in the serum of surgical intensive care unit (ICU) patients with deep mycoses. (2) To determine the concentrations of fluconazole reached at the site of infection. (3) To determine if enteral administration of fluconazole, which has an important pharmaco-economic advantage, is justified in this specific patient group. DESIGN: Descriptive, sequential study as a part of a therapeutic drug monitoring programme. SETTING: Eighteen-bed surgical ICU in a referral centre. PATIENTS: Fourteen critically ill surgical patients with recent gastro intestinal (GI) surgery and deep mycosis caused by a fluconazole-susceptible fungus and a calculated creatinine clearance of more than 40 ml/min. INTERVENTIONS: Fluconazole dosage regimen: 400 mg i. v. every 24 h with an extra dose of 400 mg i.v. after 12 h on day 1. If the clinical condition allowed enteral administration on day 4, the content of two capsules of 200 mg was given via the feeding tube with concomitant enteral feeds. MEASUREMENTS AND MAIN RESULTS: Serum, exudate from the site of infection and urine samples collected at assumed steady state ( after > or = 5 doses). Fluconazole concentrations were determined by high-performance liquid chromatography (HPLC). The mean area under the concentration curve (AUC0-24 h) in serum after enteral administration did not significantly differ from the AUC0-24 h during intravenous treatment. The elimination half-life was longer compared to healthy volunteers. The mean (95% CI) estimated bioavailability was 124 (90-158)%. The mean (95% CI) area under the concentration time curves (AUCs) achieved in the exudate from the site of infection were 67 (55-79)% of the AUCs reached in serum for both regimens. CONCLUSIONS: In critically ill patients with recent GI surgery and/or peritonitis the bioavailability of enteral fluconazole was adequate. The concentrations of fluconazole reached in exudate were lower than those in serum for both regimens, but adequate to treat most cases of deep mycoses in this specific patient group. PMID- 11280623 TI - Impact of an electronic information system on physician workflow and data collection in the intensive care unit. AB - OBJECTIVE: To test the hypotheses that: (1) integrating information processing tasks using an electronic clinical information system (ECIS) decreases time to complete these tasks by hand; and (2) structured data entry encourages generation of more detailed records and capture of specific data elements even when entry is voluntary. DESIGN: Prospective observational time analysis during medical documentation tasks. Retrospective analysis of clinical documentation completed by hand or electronically. SETTING: Eleven bed pediatric intensive care unit within an academic medical center. PARTICIPANTS: Five pediatric intensive care medicine attending physicians. MEASUREMENTS: Compared handwritten and electronic documentation to determine: (1) time spent entering data or composing notes; (2) number of descriptors documenting patients' physical exams; (3) users' preferences for structured or unstructured data entry; (4) frequency of documenting specific data elements related to nutritional support. RESULTS: Documentation time varied by user but not charting method: it took 13 % less time to document using the ECIS but this was not significant. Electronic documents were more detailed than handwritten containing 50 % more descriptors (17.8 +/- 1.4 vs 11.6 +/- 1.4) overall and some data elements that were not handwritten: information related to nutritional supplementation was recorded in 13 % of electronic documents but in none of 89 handwritten documents. CONCLUSIONS: Electronic and handwritten documentation consumed equal amounts of time. Structured entry, compared to handwriting, may encourage recording of specific or otherwise unincorporated data elements resulting in a more detailed record. This suggests that user interfaces and decision support components may influence both the types and complexity of clinical data recorded by caregivers. PMID- 11280622 TI - Glutamine: a life-threatening deficiency in the critically ill? PMID- 11280624 TI - Daily classification of the level of care. A method to describe clinical course of illness, use of resources and quality of intensive care assistance. AB - OBJECTIVE: To develop a simple and comparable clinical method able to distinguish between higher and lower complexities of care in the ICU. DESIGN: Retrospective analysis. SETTING: Database of European ICUs Study I (Euricus-I: including 12,615 patients and 55,464 patient/days), prospectively collected in 89 ICUs of 12 European countries. METHODS AND RESULTS: A panel of experts developed the classification of the complexity of care. Six (in addition to monitoring, two levels of respiratory support--R and r--two levels of circulatory support--C and c--and dialysis) out of the nine items of Nine Equivalents of Nursing Manpower use Score (NEMS), a therapeutic index, were utilised. Two levels of care (LOCs) were defined according to a more (HT) and a less complex (LT) combination of common activities of care. The two LOCs were significantly related to mortality: higher in HT and they rose with increasing cumulative number of HT days. HT accounted for 31,976 NEMS days (57.7%) while 23,488 (42.3 %) were LT. Major respiratory and cardiovascular support accounted for about 80 % of the HT days. Respiratory assistance and monitoring were responsible for an equivalent percentage of LT days. The distribution of the clinical classification of LOCs coincided with that of the managerial scores of LOCs in the literature. CONCLUSIONS: The managerial instrument described uses simple and reliable clinical data. It is able to distinguish between patients with different severity and outcome, and shows that every additional consecutive day spent in ICU as HT increases the probability of death. Moreover, (1) it suggests the possibility of describing the clinical course of illness by relating the complexity/level of medical care to the available technology and staff; (2) using relevant markers of clinical activity, it might be useful to include in quality control programmes. PMID- 11280625 TI - Human errors in a multidisciplinary intensive care unit: a 1-year prospective study. AB - OBJECTIVES: To determine the incidence and identify risk factors of critical incidents in an ICU. DESIGN: Prospective observational study of consecutive patients admitted over 1 year to an ICU. Critical incidents were recorded using predefined criteria. Their causes and consequences were analysed. The causes were classified as technical failure, patient's underlying disease, or human errors (subclassified as planning, execution, or surveillance). The consequences were classified as lethal, leading to sequelae, prolonging the ICU stay, minor, or without consequences. The correlation between critical incidents and specific factors including patient's diagnosis and severity score, use of monitoring and therapeutic modalities was analysed by uni- and multivariate analysis. SETTING: An 11-bed multidisciplinary ICU in a non-university teaching hospital. PATIENTS: 1,024 consecutive patients admitted to the ICU. INTERVENTION: None. MEASUREMENTS AND MAIN RESULTS: The median length of ICU stay by the 1,024 patients was 1.9 days. Of the 777 critical incidents reported 2% were due to technical failure and 67 % to secondary to underlying disease. There were 241 human errors (31%) in 161 patients, evenly distributed among planning (n = 75), execution (n = 88), and surveillance (n = 78). One error was lethal, two led to sequelae, 26 % prolonged ICU stay, and 57 % were minor and 16 % without consequence. Errors with significant consequences were related mainly to planning. Human errors prolonged ICU stay by 425 patient-days, amounting to 15 % of ICU time. Readmitted patients had more frequent and more severe critical incidents than primarily admitted patients. CONCLUSIONS: Critical incidents add morbidity, workload, and financial burden. A substantial proportion of them are related to human factors with dire consequences. Efforts must focus on timely, appropriate care to avoid planning and execution mishaps at the beginning of the ICU stay; surveillance intensity must be maintained, specially after the fourth day. PMID- 11280626 TI - A cost-effectiveness analysis of stays in intensive care units. AB - OBJECTIVE: To evaluate patient outcome and the efficiency of stays in intensive care units (ICUs). DESIGN: Prospective study. SETTING: Seven ICUs of teaching hospitals in the Paris area. PATIENTS: Two hundred eleven stays including one in three consecutive patients admitted from September to November 1996. MEASUREMENTS AND MAIN RESULTS: For each patient, the following information was collected during the ICU stay: diagnosis, severity scores, organ failures, workload, cost and mortality. A cost-effectiveness ratio was computed for 176 stays with at least one organ failure, at hospital discharge and 6 months later. Quality of life was measured with EuroQol questionnaires 6 months after discharge in 64 patients representing 62 % of the patients contacted. The mean total ICU cost per stay was US$ 14,130 (+/- 6,550) (higher for non-survivors--US$ 19,060, median 10,590--than for survivors US$ 12,370, median 5,780). The incremental cost effectiveness ratio was US$ 1,150 per life-year saved and the incremental cost utility ratio was US$ 4,100 per quality-adjusted life-year (QALY) saved, without discounting. These results compare favourably with other health-care options. However substantial variations were observed according to age, severity, diagnosis, number of organ failures and discount rate. Intoxication had the lowest ratio (US$ 620/QALY) and acute renal insufficiency the highest (US$ 30,625/QALY). CONCLUSIONS: This work provides medical and economic information on ICU stays in teaching hospitals and enables comparisons with other health-care options. PMID- 11280627 TI - Changes in quality of life after medical intensive care. AB - OBJECTIVES: To determine outcome and changes in health-related quality of life (QOL) in medical intensive care patients. DESIGN AND SETTING: Prospective comparison of QOL before and 6 months after intensive care unit (ICU) admission in a 12-bed noncoronary medical ICU of a university hospital. PATIENTS: All 325 consecutively admitted adult patients who spent at least 24 h on the ICU were eligible. MEASUREMENTS AND RESULTS: QOL measurements were collected before and 6 months after ICU admission. Comorbidity classified by the Charlson index was 2.44 +/- 1.96. Mean stay in the ICU was 10.4 +/- 15.1 days, mean Acute Physiology and Chronic Health Evaluation II score was 23 +/- 10. Cumulative mortality was: ICU 24 %, hospital 34 %, 6 months 43 %. Relative to baseline, follow-up interviews of 185 survivors revealed no significant changes in the overall QOL score (p = 0.93). The subscales basic physiological activities (p = 0.07) and normal daily activities (p = 0.15) showed a nonsignificant deterioration. A significant improvement was noted for the domain emotional state (p = 0.013). CONCLUSIONS: Six months after admission to a medical ICU most survivors had regained their preadmission health-related QOL. Multivariate analysis showed that preadmission QOL, age, and severity of illness were most strongly associated with follow-up QOL. Of the survivors 86 % were living at home, and all but one of those previously in employment had returned to their former work. Most patients (94%) would undergo ICU treatment again if necessary. PMID- 11280628 TI - Selenium and the "free" electron. Selenium--a trace to be followed in septic or inflammatory ICU patients? PMID- 11280629 TI - Needs of relatives of critical care patients: perceptions of relatives, physicians and nurses. AB - OBJECTIVES: This study investigated differences between the perceptions of relatives, physicians and nurses concerning the needs of relatives of critical care patients. DESIGN AND PARTICIPANTS: Perceived needs were assessed in 200 relatives, 38 physicians, and 143 nurses using a 45-item questionnaire. SETTING: Data were gathered at the intensive care unit of the University Hospital Gasthuisberg. RESULTS AND CONCLUSIONS: The overall rankings of the needs by the three groups are very similar. Information emerges as most important factor, with considerably less importance attached to comfort and support. There were significant differences between the groups on all categories and on 24 individual needs. Regarding the need categories, both nurses and physicians underestimate the relatives' need for information and proximity to the patient. Physicians also underestimate the relatives' need for assurance. On the individual need items, relatives' needs are generally underestimated by the staff, but in some cases overestimations are found. PMID- 11280631 TI - Health informatics. AB - Health informatics is the development and assessment of methods and systems for the acquisition, processing and interpretation of patient data with the help of knowledge from scientific research. This definition implies that health informatics is not tied to the application of computers but more generally to the entire management of information in healthcare. The focus is the patient and the process of care. The apparent information overload and the imperfection of medical decision making motivate the use of information systems for medical decision support. Health informatics provides tools to control processes in healthcare, acquire medical knowledge and communicate information between all people and organisations involved with healthcare. Although the development of medical information systems may often lag behind the available possibilities, the technological state of the current medical information systems is better than it is generally held to be. Health informatics should help healthcare professionals to provide better and more cost-effective care and enable healthcare systems to be more efficient and to adapt better to our patients' needs. Health informatics may reshape the way we deliver care to meet the demands of the future. PMID- 11280630 TI - International Consensus Conferences in Intensive Care Medicine: non-invasive positive pressure ventilation in acute respiratory failure. Organised jointly by the American Thoracic Society, the European Respiratory Society, the European Society of Intensive Care Medicine, and the Societe de Reanimation de Langue Francaise, and approved by the ATS Board of Directors, December 2000. PMID- 11280632 TI - Intracatheter nitroglycerin infusion fails to prevent catheter-related venous thrombosis: a randomized, controlled trial. AB - OBJECTIVE: Catheter-related thrombosis is a common problem in the pediatric intensive care unit. Strategies that reduce the incidence of thrombosis may have significant clinical advantage. Nitroglycerin (NTG) infusions release nitric oxide (NO). NO is responsible for much of the vasodilating and antithrombotic properties of the vasculature. We hypothesized that an intracatheter NTG infusion would reduce the incidence of catheter-related thrombosis. DESIGN: Prospective, randomized, controlled trial. SETTING: Pediatric intensive care unit. PATIENTS AND PARTICIPANTS: Children of 6 years or less with femoral venous catheters who were not on antithrombotic therapy. INTERVENTIONS: Subjects were randomly assigned to NTG or control groups. NTG group patients received NTG at 0.1 mcg x kg x min in 5 % dextrose; control group patients received only 5 % dextrose. Infusions were delivered continuously through the catheter until the catheter was removed. Demographic data, physical and laboratory findings, catheter insertion attempts and infusate composition were recorded. Clinical evidence of vascular thrombosis or catheter malfunction was noted. Ultrasound examinations were performed within 2 days of catheter insertion and within 2 days after removal. MEASUREMENTS AND RESULTS: Forty-four patients (age 12.0 +/- 2.6 months) completed the study, 21 in the NTG group and 23 in the control group. Duration of catheter placement was 7.5 +/- 0.7 days. Twelve of 44 patients (27 %) had thrombi: 7/21 in the NTG group; 5/23 in the control group (p = NS). There were no significant differences between children with and without thrombi in age, gender, number of insertion attempts, duration of catheter placement, clinical signs of thrombosis or infections. CONCLUSIONS: Catheter-related thrombosis is common after placement of femoral venous catheters in children. Low dose intracatheter NTG infusion does not protect against catheter-related venous thrombosis in children. PMID- 11280633 TI - The endothelial response to oxygen deprivation: biology and clinical implications. AB - This review provides the theoretical background of phenotypic and gene-based changes in the vessel wall triggered by acute hypoxia. Only in the last few decades has the endothelium been ascribed a prominent role as a modulator of vascular homeostasis under both physiological and pathological conditions. Molecular mechanisms leading to endothelial activation are being rapidly elucidated and their contribution to vascular dysfunction during hypoxia becoming better understood. New insights gained from hypoxic cell culture and ischaemic organ models may ultimately lead to new treatment strategies. If nothing else, insights gained from vascular research will lead to a more complete understanding of the inflammatory processes in blood vessels and how they impact on human disease. PMID- 11280634 TI - Capillary leak syndrome in children who undergo cardiopulmonary bypass: clinical outcome in comparison with complement activation and C1 inhibitor. AB - OBJECTIVES: Capillary leak syndrome (CLS) is associated with significantly increased morbidity and occurs after cardiopulmonary bypass in children with congenital heart disease. We investigated the early clinical parameters that predict the development of CLS and examined the relationship between the presence of CLS and complement and contact activation and C1 esterase inhibitor (C1-INH) during and after bypass. DESIGN: In this prospective study we took serial serological measurements of the complement and contact system and C1-INH in a cohort of 27 infants before, during, and up to 96 h after open-heart surgery. RESULTS: Complement and contact activation and a decrease in C1-INH were measured in all infants during and after CPB. Ten infants developed CLS postoperatively. Younger age and longer bypass time were strongly correlated to the development of CLS. No relationship was found between the degree of hypothermia, weight, gender, or cross-clamp time. C1-INH concentration and activity were lower peri- and postoperatively in the CLS group. Infants with CLS had a more pronounced postoperative increase in the C5a and C3a levels, higher postoperative level of factor XIIa, and lower prekallikrein activity than those without CLS. CONCLUSION: Contact and complement activation occurs during cardiopulmonary bypass and contributes to CLS more frequently in infants of a younger age and with a prolonged bypass time. This activation and decrease in C1-INH was strongly expressed in the CLS group, and therefore early substitution of C1-INH may prevent CLS after open-heart surgery in high-risk infants. PMID- 11280635 TI - Are transoesophageal Doppler parameters a reliable guide to paediatric haemodynamic status and fluid management? AB - OBJECTIVE: Transoesophageal Doppler (TOD) has been used in adults to optimise left ventricular filling on the basis of the waveform parameters. We wished to see if a similar relationship exists in children, specifically: (a) whether change in thermodilution stroke volume (SV) following a fluid bolus corresponded to change in Doppler stroke distance, Doppler corrected flow time (FTc), or central venous pressure (CVP); (b) whether a response to fluid challenge (defined as an increase in SV of greater than 10%) can be predicted on the basis of an absolute value for FTc or CVP prior to fluid bolus; and (c) the relationship between FTc and systemic vascular resistance index. DESIGN: Prospective, comparison study. SETTING: Sixteen-bed paediatric intensive care unit of a university hospital. PATIENTS: Ninety-four ventilated children were studied, median (range) age 25 months (4 days- 16 years). Diagnoses included: post-cardiac surgery (n = 58), sepsis/multi-organ failure (n = 29), respiratory disease (n = 5), and other (n = 2). INTERVENTIONS: A 4-MHz, 5.5-mm diameter, flexible TOD probe was placed when patients were haemodynamically stable. Five consecutive measurements of stroke distance and FTc were made and averaged, concurrently with five SV measurements by femoral artery thermodilution. SV was then augmented by administration of fluid (10 ml/kg), and haemodynamic recordings were repeated. MEASUREMENTS AND MAIN RESULTS: The median (range) SV was 17 ml (2-64 ml). The median coefficients of variation were 3.9 % for SV, 3.5 % for stroke distance, and 3.1% for FTc. Changes in SV were accurately tracked by changes in stroke distance (mean bias 1.8 %, limits of agreement +/- 17%), but not by FTc or CVP. FTc was weakly inversely correlated with systemic vascular resistance (r = -0.15, P < 0.05). Among non-cardiac patients (n = 36), the optimal FTc that predicted an improvement in SV following fluid bolus was 0.394 s (area under ROC curve 0.756), giving a sensitivity of 90 %, specificity of 62 %, positive predictive value of 47 %, and a negative predictive value of 94 %. CVP was a poor predictor for all patient groups. CONCLUSIONS: TOD stroke distance is able to follow changes in SV following fluid bolus amongst ventilated children, and can predict when further volume loading is unlikely to improve SV amongst general, but not cardiac ICU patients. CVP is a poor discriminator of volume status in this group of patients. PMID- 11280636 TI - Low-dose dopamine in neonatal and pediatric intensive care: a systematic review. AB - OBJECTIVES: To assess the current use of low-dose dopamine (< 5 microg/kg per minute) to improve renal function and urine volume (UV) in neonatal (NICU) and pediatric (PICU) intensive care units, and to assess the available evidence to support this practice. DESIGN: A written survey was used to assess the current use of low-dose dopamine among all 19 NICUs and PICUs in the Netherlands. In addition, a review of the literature of clinical intervention studies in which low-dose dopamine was administered to improve renal function and UV was performed. METHODS: The clinical intervention studies focused on preterm neonates, critically ill infants and children, and those who underwent cardiac surgery. Either creatinine clearance or glomerular filtration rate and increase in UV were used to measure renal function improvement. RESULTS: Our survey showed that among the 19 NICUs and PICUs, dopamine is regularly used either to improve renal function (n = 7) or to enhance UV (n = 13). The literature review identified seven clinical studies. Of these only one was a randomized controlled trial in preterm neonates, and this showed no positive correlation between renal function and UV. The other studies were uncontrolled experiments in preterm infants that claimed positive effects on UV (n = 5) and creatinine clearance (n = 2). CONCLUSIONS: The widespread use today of low-dose dopamine in Dutch NICUs and PICUs is not supported in the literature. Evidence from well performed clinical studies to support the use of low-dose dopamine for improving renal function and UV in critically ill neonates and children is largely insufficient. In view of adverse effects, the use of low-dose dopamine in neonatal and pediatric intensive care patients should be reconsidered. PMID- 11280637 TI - Comparison of procalcitonin with C-reactive protein and serum amyloid for the early diagnosis of bacterial sepsis in critically ill neonates and children. AB - OBJECTIVES: To evaluate procalcitonin (PCT) as a diagnostic marker of bacterial sepsis in critically ill neonates and children and to compare the results of PCT with those of C-reactive protein (CRP) and serum amyloid (SAA). DESIGN AND SETTING: Prospective, observational study in neonatal and pediatric intensive care units. PATIENTS: A total of 116 divided into four groups according to age and diagnosis: neonates (aged 3-30 days) with sepsis (n = 20), neonates without sepsis (n = 26), children (aged 2-12 years) with sepsis (n = 32), and children without sepsis (n = 38). INTERVENTIONS: Serum PCT, CRP, and SAA were measured on admission or when a bacterial sepsis was suspected. Area under the receiver operating characteristic (ROC) curve, optimum predictive values, and optimum diagnostic cut off values were evaluated. RESULTS: Admission PCT was significantly higher in neonates and children with sepsis than in the other groups. In the neonates the area under the ROC curve was 0.99 for PCT, 0.95 for CRP, and 0.98 for SAA; in the children it was 1 for PCT, 0.93 for CRP, and 0.96 for SAA. Cutoff concentrations for optimum prediction of sepsis in the neonates were PCT > 6.1 ng/ml (diagnostic efficiency: 93.8%), CRP > 23.0 mg/l (89.7%), and SAA > 41.3 mg/l (95.3%); in the children they were PCT > 8.1 ng/ml (100%), CRP > 22.1 mg/l (89.8%), and SAA > 67.2 mg/l (94.4%). CONCLUSION: In critically ill children PCT concentration is a better diagnostic marker of sepsis than CRP and SAA. In critically ill neonates, however, PCT, CRP, and SAA are similar diagnostic markers of sepsis. A PCT concentration higher than 8.1 ng/ml identified all children with bacterial sepsis. PMID- 11280638 TI - Xanthine oxidase activity and blood glutathione redox ratio in infants and children with septic shock syndrome. AB - OBJECTIVES: The possible role of xanthine oxidase (XO) activation in the signal transduction process during the septic shock syndrome was examined. The XO activity index after caffeine intake was assessed simultaneously with the blood glutathione redox ratio, a known parameter of oxidative stress. DESIGN AND SETTING: An investigational clinical study in a nine-bed pediatric intensive care unit. PATIENTS: Critically ill infants and children (n = 34) with systemic inflammatory response syndrome following infection, trauma or major surgery. Biochemical investigations (n = 54) were performed at various stages of the shock syndrome, characterized by pediatric risk of mortality and organ dysmetabolic scores. Controls consisted of 30 healthy children. MEASUREMENTS AND RESULTS: The in vivo XO activity index was measured as the urinary ratio of two metabolites of caffeine: 1-methyluric acid and 1-methylxanthine. The blood concentrations of oxidized (GSSG) and reduced glutathione (GSH) were determined. The XO activity index and redox ratio GSSG/GSH were highly increased in patients in shock dominated by the clinical symptoms of a proinflammatory response. A significantly lower XO activity index was found with an increased GSSG/ GSH in patients whose stage of shock was characteristic of an excessive anti-inflammatory response. The XO activity index and GSSG/ GSH were correlated closely with each other (r = 0.624, n = 54; p < 0.001), and were also related to the daily severity scores. CONCLUSION: Potent and simultaneous activation of the two redox systems strongly indicates a definite role of free radicals from XO in the overspill of the acute proinflammatory reaction of the shock syndrome, followed by a significant downregulation. PMID- 11280639 TI - The value of capillary whole blood lactate for blood transfusion requirements in anaemia of prematurity. AB - OBJECTIVE: To evaluate the usefulness of blood lactate as an indication for blood transfusion in anaemia of prematurity by means of a study protocol which considers the site of blood sampling and the repeatability of lactate measurements. DESIGN: Prospective clinical study. SETTING: Multidisciplinary, neonatalpaediatric intensive care unit of a non-university, teaching children's hospital. PATIENTS AND METHODS: Comparison of pre- and 48-h post-transfusion capillary whole blood lactate in 18 anaemic premature babies. In 30 neonates the agreement between capillary and arterial lactate was analysed by using the Bland Altman plot. In 30 stable premature infants four capillary lactate measurements were carried out within 24 h and analysed with regard to variability (coefficient of variation (CV); association between SD and mean) and to establish normal values. RESULTS: In the transfused infants, haematocrit increased from 23 (SD 3)% to 37 (SD 3)%. Mean lactate decreased from 2.5 (SD 1.0) to 1.7 (SD 0.5) mmol/l (p = 0.003). Pretransfusion lactate did not correlate with pre-transfusion haematocrit, heart rate, respiratory rate, number of apnoeas/bradycardias and weight gain (multiple regression). The mean difference between capillary and arterial lactate was 0.17 (SD 0.24) mmol/l and the 95 % confidence interval (CI) was -0.31 to 0.65 mmol/l. The CV of repetitive measurements was 19.8 (SD 9.8)% and SD correlated positively with mean lactate values (p = 0.001); the 95 % CI (normal range for premature infants) was 1.56-1.90 mmol/l. CONCLUSIONS: Capillary whole blood lactate measurements in newborn babies agree excellently with arterial values. Lactate measurements add little information to the decision whether to transfuse or not, considering the variability of this parameter in stable premature infants and the lack of correlation with other possible clinical indicators of compromised oxygen delivery. PMID- 11280640 TI - In vitro validation of a metabolic monitor for gas exchange measurements in ventilated neonates. AB - OBJECTIVE: To evaluate the Datex Deltatrac II for measurements in neonates requiring mechanical ventilation. DESIGN: Prospective laboratory evaluation, using a ventilated lung model and gas injection. During simulation of 79 neonatal respiratory settings, assessment of oxygen consumption (VO2), carbon dioxide production (VCO2) and respiratory quotient (RQ) was compared to a reference method (mass spectrometry, wet gas spirometry) using the statistical method of Bland and Altman. INTERVENTIONS: Respiratory variables, which may influence the accuracy and precision of gas exchange measurements, were varied within the following ranges: inspired oxygen fraction (FIO2): 0.21-0.8, expired carbon dioxide fraction (FECO2) and inspiratory-expiratory oxygen fraction (DFO2): 0.0032-0.0256, expiratory flow rate: 1.0-2.5 l/min, inspiratory pressure: 10-55 mbar, respiratory rate 25-60/min, constant RQ of 1. This resulted in 79 tests with VCO2 and VO2 ranging from 8-64 ml/min. MEASUREMENTS AND RESULTS: The coefficient of repeatability for ten single subsequent Deltatrac measurements was 8.09 ml/min for VO2 and 9.17 ml/min for VCO2 compared to 2.02 ml/min and 0.90 ml/min for VO2 and VCO2 with repeated reference measurements. The coefficient of repeatability of the Deltatrac measurements improved considerably when means of subsequent 5 min intervals were compared: 0.68 ml/min for VO2 and 0.28 ml/ min for VCO2. The difference between the two methods (Deltatrac-reference) was -3.8 % (2 s: 11.4%) for VO2, 13.2% (2s: 7.9%) for VCO2 and 17.6% (2s: 16.7%) for RQ. The agreement between methods deteriorated with smaller (FECO2) or DFO2 and increasing FIO2. CONCLUSIONS: Considering limits of agreement of less than +/- 20% as clinically acceptable, results for VO2 assessment indicate acceptable accuracy and precision whereas VCO2 and RQ assessments exceed this limit. Limited accuracy and precision result from detection of CO2 following dilution of expiratory gases and increased sensitivity to error propagation by Haldane equations due to the small differences between inspiratory and expiratory gas fractions. PMID- 11280641 TI - Survey of the use of intracranial pressure monitoring in children in the United Kingdom. AB - OBJECTIVE: To establish current practice for the monitoring and management of acute intracranial hypertension in children in United Kingdom intensive care units (ICUs). DESIGN: Postal questionnaire, targetted by prior telephone survey, to all ICUs admitting five or more children per annum with acute neurological illness. RESULTS: Of the units contacted 70 % responded, approximately one-half of which reported the use of intracranial pressure (ICP) monitoring. Only data from these units are presented. Nearly all of these units consider monitoring following serious head injury, but its use in non-traumatic brain injury is less widespread. The decision to institute ICP monitoring is based mainly upon neuroimaging appearances and Glasgow Coma Scale score. ICP and cerebral perfusion pressure targets differ markedly between centres, with only 46 % and 65 % of units, respectively, setting age-dependent parameters. Mannitol and varying degrees of hyperventilation are employed by all units to lower ICP. The majority also use barbiturates, diuretics, and fluid restriction. Controlled hypothermia is used in 52 % of units. Paediatric units are more likely to employ age dependent cerebral perfusion pressure targets. Specific therapies employed to lower ICP are similar to those used in adult centres. CONCLUSION: Faced with a lack of both evidence and consensus, the management of acute intracranial hypertension in childhood varies widely. National or international guidelines for the management of children with raised intracranial pressure are needed. These should incorporate the physiological differences between children of different ages. PMID- 11280642 TI - Misplacement of a femoral venous catheter into the ascending lumbar vein: repositioning using ultrasonographic guidance. AB - A 5-week-old infant with congenital chylothorax required long-term intravenous access for parenteral nutrition. Cannulation of the inferior vena cava via the left femoral vein was attempted, but the catheter was misplaced into the left ascending lumbar vein. Catheter removal is advised when such malposition is identified. We were able successfully to redirect the catheter into the inferior vena cava using ultrasonographic guidance. This procedure has not been described previously in children. We propose that repositioning of incorrectly placed vascular catheters can be achieved using ultrasound guidance at the bedside. PMID- 11280643 TI - Pulmonary administration of prostacyclin (PGI2) during partial liquid ventilation in an oleic acid-induced lung injury: inhalation of aerosol or intratracheal instillation? AB - OBJECTIVE: The purpose of this study was to investigate the effects of aerosolized prostacyclin (A-PGI2) and intratracheally instilled prostacyclin (I PGI2) during partial liquid ventilation (PLV) on gas exchange and pulmonary circulation in rabbits with acute respiratory distress. DESIGN: Prospective control study. SETTING: A research laboratory at a university medical centre. SUBJECTS: Sixty-nine Japanese white rabbits. INTERVENTION: Lung injury was induced by oleic acid and the animals were divided into five groups of ten each: a mechanical gas ventilation (GV) group, an A-PGI2 group, a PLV group, an A PGI2+PLV group and an I-PGI2+PLV group. PLV, A-PGI2+PLV and I-PGI2+PLV groups received 15 ml/ kg perflubron intratracheally while receiving mechanical GV. A PGI2 and A-PGI2+PLV groups received aerosolized PGI2 (50 ng/kg/min) in combination with GV or PLV, respectively. The I-PGI2+PLV group was instilled 50 ng/kg/min PGI2 intratracheally in combination with PLV. RESULT: After lung injury, all animals developed hypoxia, hypercarbia and pulmonary hypertension. The improvement of partial pressure of arterial oxygen (PaO2) in the A-PGI2 and PLV groups was transient, whereas the A-PGI2+PLV and I-PGI2+PLV groups showed consistent improvement throughout the experiment. The PaO2 values of the I PGI2+PLV group were significantly higher than those of the other groups 120 min after treatment. The mean pulmonary artery pressure (PAP) significantly decreased after treatment in the A-PGI2, APGI2+PLV and I-PGI2+PLV groups. CONCLUSIONS: The results suggest that both aerosolized and intratracheally instilled PGI2 improve oxygenation and reduce PAP during PLV in oleic acid lung injury. PMID- 11280644 TI - Inhibition of lung phosphodiesterase improves responsiveness to inhaled nitric oxide in isolated-perfused lungs from rats challenged with endotoxin. AB - OBJECTIVES: To investigate the ability of phosphodiesterase (PDE) selective inhibitors to improve responsiveness to inhaled nitric oxide (NO) in isolated perfused lungs of rats pretreated with endotoxin/lipopolysaccharide (LPS). DESIGN AND SETTING: Prospective, controlled animal study in the animal research facility of a university hospital. INTERVENTIONS: Sixteen hours after adult Sprague-Dawley rats were injected intraperitoneally with 0.4 mg/ kg E. coli 0111:B4 LPS administration, lungs were isolated and perfused, and the thromboxane mimetic U46619 was employed to increase the mean pulmonary artery pressure by 5-7 mmHg. The lungs were then ventilated with or without 0.4 ppm NO, and erythro-9-(2 hydroxy-3-nonyl) adenine (EHNA; PDE type 2 inhibitor), milrinone (PDE type 3 inhibitor), or zaprinast (inhibitor of PDE types 5 and 9) were added to the perfusate. MEASUREMENTS AND RESULTS: In the presence of EHNA (12.5, 25, 50 microM) the vasodilator response to inhaled NO was not greater than in its absence (0.25 +/- 0.25, 0.5 +/- 0.25, 0.75 +/- 0.25 mmHg vs. 0.25 +/- 0.25, 0.5 +/- 0.25, 0.75 +/- 0.25 mmHg, respectively). In the presence of milrinone (125, 250, 500 nM), the vasodilator response to inhaled NO was also not improved. In contrast, zaprinast (3.7, 7.4, 14.8 microM) augmented the pulmonary vasodilatory effect of inhaled NO in lungs from LPS-pretreated rats from 0.25 +/- 0.25, 0.5 +/ 0.25, 0.75 +/- 0.25 mmHg to 0.75 +/- 0.25, 1.5 +/- 0.5, 1.75 +/- 0.75 mmHg, respectively (p < 0.05). CONCLUSIONS: Our results demonstrate that inhibition of pulmonary PDE enzyme activity with zaprinast increases vasodilator responsiveness to inhaled NO in lungs obtained from rats 16 h after LPS challenge. PMID- 11280645 TI - Reduction in intestinal leukocyte adherence in rat experimental endotoxemia by treatment with the 21-aminosteroid U-74389G. AB - OBJECTIVES: To investigate leukocyte adherence in intestinal venules in experimental endotoxemia after treatment with the 21-aminosteroid U-74389G. DESIGN AND SETTING: Prospective, randomized, controlled animal study in an experimental laboratory. SUBJECTS: Twenty-one male Wistar rats weighing 190 +/- 40 g. INTERVENTIONS: The rats were divided equally into three groups: (a) control group, (b) endotoxemia (5 mg/kg lipopolysacharide from Escherichia coli O55:B5), and (c) endotoxemia and U-74389G administration 30 min before (3 mg/kg) and 60 min after endotoxin challenge (1.5 mg/ kg). MEASUREMENTS AND MAIN RESULTS: The distal small intestine of the animals was examined using intravital fluorescence videomicroscopy 2 h after endotoxin challenge. Leukocytes were stained in vivo by means of rhodamine 6G. In the endotoxemic animals we observed a fourfold increase in the count of firmly adherent leukocytes in submucosal post-capillary and collecting venules. Treatment with the 21-aminosteroid U-74389G significantly attenuated the count of sticking leukocytes in the collecting venules (control, 61 +/- 10 cells/mm2; lipopolysaccharide, 237 +/- 42 cells/mm2; U-74389G 125 +/- 9 cells/mm2; p < 0.05). In these venules leukocyte rolling behavior was comparable to that in the control group without endotoxin challenge. CONCLUSIONS: Administration of U-74389G, which has radical scavenging properties, attenuates leukocyte adherence in selected populations of intestinal venules which is found increased during endotoxemia. Thus, 21-aminosteroids may have an impact in the treatment of endotoxin-induced intestinal injury. PMID- 11280646 TI - Effects of magnesium sulfate on tissue lactate and malondialdehyde levels in experimental head trauma. AB - OBJECTIVE: To determine the effects of magnesium sulfate (MgSO4) on tissue lactate and malondialdehyde (MDA) levels in rabbit brain after experimental head trauma. DESIGN: Prospective, randomized trial. SUBJECTS: Thirty New Zealand rabbits. INTERVENTIONS: Group 1 (n = 10) was the sham operated group. Group 2 (n = 10) (untreated group) and group 3 (n = 10) received head trauma with the weight drop method. MgSO4 was administered 100 mg/kg (15 %) i. v. immediately after the head trauma to group 3. Trauma was applied to one side. The non-contused side was named as "a" and the contused side as "b". MEASUREMENTS: One hour after trauma, brain cortices were resected and the concentrations of lactate and MDA were determined using the spectrophotometric enzymatic and thiobarbituric acid methods. One-way ANOVA and Tukey's HSD tests were used for the evaluation of the results. P < 0.05 was considered as significant. Pearson's correlation test was used between lactate and MDA levels (P < 0.001). RESULTS: There were significant differences between MDA and lactate levels of group 1 and all other groups; non contused (a) and contused (b) sides of groups 2 and 3; groups 2b-3a, 2b-3b (P < 0.05). The difference in MDA levels was significant between groups 2a-3b (P < 0.05). Correlation between lactate and MDA was very good in group 1, and excellent in groups 2a, 2b, 3a, and 3b. CONCLUSIONS: These results demonstrate that head trauma leads to an increase in brain tissue lactate and MDA levels, and MgSO4 suppresses the rise in contused tissue when given after head trauma. PMID- 11280647 TI - The effect of hypoxemic reperfusion on cerebral protection after a severe global ischemic brain insult. AB - BACKGROUND AND PURPOSE: Reactive oxygen species contribute to membrane lipid peroxidation and neuronal death and have been implicated in anoxic encephalopathy. We tested whether hypoxemic reperfusion (HR) after global cerebral ischemia would improve neurological recovery. METHODS: Two groups of pigs (n = 11 in each group) were subjected to a model of a 10-min global cerebral and systemic ischemia to compare the effect of hypoxemic reperfusion (group HR) with the classical hyperoxemic control (group C). A third group not subjected to ischemia served as control to the control group (n = 6, group CC), but received hyperoxygenation at the respective period of reperfusion. The outcome was evaluated by means of neurological assessment and the extent of lipid peroxidation measuring the plasma malonaldehyde (MDA) together with hydroxyalkenals (HALK). RESULTS: Animals of group HR exhibited a significantly superior neurological outcome compared with those of group C at all three consecutive assessments after reperfusion (post-resuscitation P = 0.006, at 8 h P = 0.003, and at 24 h P = 0.007). The levels of MDA and HALK are lower in the HR group than in group C (P = 0.029). Additionally, in the CC group these molecules increased significantly early at hyperoxygenation (P = 0.02). A faster lactate metabolism in the HR group was observed during reperfusion, though non significant. CONCLUSIONS: Hypoxemic reperfusion during resuscitation from a severe global ischemic cerebral insult improves the neurological outcome compared with classic hyperoxemic reperfusion. This is additionally confirmed by the decreased production of the molecules of lipid peroxidation. In the absence of preceding ischemia, these molecules may increase by simple over-oxygenation. PMID- 11280648 TI - Epinephrine, norepinephrine and dopamine infusions decrease propofol concentrations during continuous propofol infusion in an ovine model. AB - OBJECTIVE: To determine the effects of exogenous ramped infusions of epinephrine, norepinephrine and dopamine on arterial and effluent brain blood concentrations of propofol under steady state intravenous anesthesia. DESIGN: Prospective, randomized animal study. SETTING: University research laboratory. SUBJECTS: Five adult female merino sheep. INTERVENTIONS: Induction (5 mg/kg) and continuous infusion of propofol (15 mg/min) with controlled mechanical ventilation to maintain PaCO2 40 mmHg. After 1 h of continuous anesthesia, each animal randomly received ramped infusions of epinephrine, norepinephrine (10, 20, 40 microg/min) and dopamine (10, 20, 40 microg x kg x min) in 3 x 5 min intervals followed by a 30-min washout period. MEASUREMENTS: Arterial and sagittal sinus whole blood for determination of propofol concentrations using high-pressure liquid chromatography. Cardiac output using a thermodilution method. Level of consciousness using an observational scale. MAIN RESULTS: All three drugs significantly and transiently increased cardiac output in a dose-dependent fashion to a maximum of 146-169% of baseline. Baseline arterial and sagittal sinus propofol concentrations were not statistically different prior to catecholamine infusions. All three drugs significantly reduced mean arterial propofol concentrations (95 % CI, p < 0.05): epinephrine to 41.8% of baseline (11.4-72), norepinephrine to 63 % (27-99) and dopamine to 52.9 % (18.5-87.3). There were parallel reductions of concentrations in sagittal sinus blood leaving the brain. The lowest blood concentrations were associated with emergence from anesthesia. Arterial concentrations were inversely related to the simultaneously determined cardiac output (r2 = 0.74, p < 0.0001). Comparison of the data with the predictions of a previously developed recirculatory model of propofol disposition in sheep showed the data were consistent with a mechanism based on increased first pass dilution and clearance of propofol secondary to the increased cardiac output. CONCLUSIONS: Catecholamines produced circulatory changes that reversed propofol anesthesia. These observations have potential clinical implications for the use of propofol in hyperdynamic circulatory conditions, either induced by exogenous catecholamine infusions or pathological states. PMID- 11280649 TI - Response of neonatal platelets to nitric oxide in vitro. AB - OBJECTIVES: Several studies have demonstrated altered platelet function during nitric oxide inhalation (iNO) in adults and neonates. In vitro NO inhibits activation of fibrinogen receptor glycoprotein (GP) IIb/IIIa in a dose-dependent manner. In neonates GPIIb/IIIa response to stimulation is physiologically attenuated during the first days after birth in comparison to adults; the effects of NO on GPIIb/IIIa in neonates, however, are less established. We investigated the response of platelets from neonates, their mothers, and nonpregnant controls to the NO donor SIN-1 in vitro. DESIGN: Umbilical cord and venous (mother, controls) platelet-rich plasma was stimulated in vitro with 10 microM ADP or 0.05 U/ml thrombin in the presence or absence of 10 microM SIN-1. GPIIb/IIIa activation was determined by two-color flow cytometry. SETTING: Delivery department of an university hospital. PATIENTS AND PARTICIPANTS: Ten healthy term neonates, their mothers and nonpregnant controls. MEASUREMENTS AND RESULTS: NO significantly reduced GPIIb/IIIa activation in thrombin- and ADP-stimulated platelets in all groups (p < 0.001). Neonatal platelets were significantly hyporeactive to stimulation (p < 0.05), but the relative response to SIN-1 was similar in all three groups (70 +/- 5 %). CONCLUSIONS: The relative amount of NO induced inhibition of GPIIb/ IIIa activation in neonates is thus similar to that of adults. However, due to the intrinsic hyporesponsiveness of neonatal platelets and NO-synergistic pharmacodynamic profiles of other drugs (e.g., prostacyclin), possible adverse effects of iNO must be considered. PMID- 11280650 TI - Infusion pump performance with vertical displacement: effect of syringe pump and assembly type. AB - OBJECTIVE: To evaluate the effect of different infusion pump models on continuity of drug delivery during vertical displacement of syringe pumps. DESIGN: Zero-drug delivery time (ZDDT), retrograde aspiration volume, and infusion bolus were recorded using the same syringe in three different models of syringe pump after lowering and elevating the pump. Compliance of each infusion assembly was measured using the occlusion release technique at 38 mmHg. RESULTS: Lowering the pump by 50 cm at an infusion rate of 1 ml/h resulted in ZDDT values ranging from 2.78 +/- 0.29 to 5.99 +/- 1.09 min. Elevating the syringe pump to its original position caused infusion boluses between 44.1 +/- 3.2 and 77.1 +/- 5.1 microl. The results demonstrated that there are large differences between syringe pump models (F = 66.8, df = 2/33, p < 0.0001) and between pumps of the same model (F = 21.3, df = 1/34, p < 0.0001). A similar pattern was found in retrograde aspiration volume and infusion bolus. CONCLUSION: All tested pumps led to clinically relevant flow irregularities during vertical displacement of the syringe pump. Thus, vertical displacement of any syringe pump connected to an infusion line delivering highly potent drugs at low infusion rates should be avoided. The variability across syringe pumps indicates that syringe pump design remains an area of potential further improvement for reducing the risk of adverse patient events. PMID- 11280651 TI - Comparative clinical trial of progressive dilatational and forceps dilatational tracheostomy. AB - OBJECTIVE: To compare the safety and early complications of progressive dilatational tracheostomy (PDT) and forceps dilatational tracheostomy (FDT). DESIGN: Prospective randomized trial. SETTING: Three-centre university and public teaching hospitals. PATIENTS: One hundred and twenty-seven consecutive patients were randomized to undergo PDT or FDT. RESULTS: The duration of the procedure was 7.0 +/- 3.5 min in the PDT group and 6.4 +/- 4.9 min in the FDT group, which was not significantly different. Intraprocedural complications occurred in 14 % with PDT and 31% with FDT (p = 0.03), and were usually minor. Difficult or false insertion of the cannula in eight patients after FDTwas the most common complication. CONCLUSIONS: Both percutaneous tracheostomy techniques are safe. The early complication rate of PDT appeared to be lower than FDT, but the early complication rate of FDT may be decreased significantly with small modifications to the set. PMID- 11280653 TI - Aerosolized beta2-agonists in the intensive care unit: just do it. PMID- 11280652 TI - Changing patterns of airway accidents in intubated ICU patients. AB - OBJECTIVE: To document the changes in patterns of airway accidents in intubated patients. DESIGN: Prospective recording of all airway accidents over two periods: 1994-1997 and 1998-1999. PATIENTS: Ventilated patients (5,046) intubated for 9,289 days over 4 years (1994-1997) and 2,932 ventilated patients intubated for 6,339 days over 2 years (1998-1999). MEASUREMENTS: The incidence and pattern of airway accidents over a 2-year period were compared to an earlier similar analysis done in the previous 4 years. RESULTS: The total accident rate in the 1994-1997 period was 36 in 5,046 patients over 9,289 intubated-patient days. The total accident rate in the period 1998-1999 was 20 in 2,932 patients over 6,339 intubated-patient days. The frequency of blocked tracheal tube increased to equal that of unplanned extubation (UE) of endotracheal tube (ETT) as the commonest airway accident. There were nine episodes of blocked tracheal tube in the two current years compared to four in the previous 4 years and there were nine episodes of UE in the two current years compared to 15 in the previous 4 years. There were a total of 18 ETT accidents in 2,930 patients over 5,309 ETT days compared to a total of two tracheostomy accidents in 67 patients over 1,030 tracheostomy days. CONCLUSIONS: We noted a change of the pattern of airway accidents. We noted an increasing trend in the incidence of blocked tracheal tubes, associated with an increased duration of heat and moisture exchanger filters use. We also noted that the incidence of tracheostomy tube accidents was similar to that of ETT accidents in the current study, unlike the earlier study where tracheostomy tube accidents were more frequent than ETT accidents. This was due to the elimination of tracheostomy tube displacements during the later study period. We associated this with the use of adjustable tracheostomy length tubes. PMID- 11280654 TI - The Internal jugular veins are asymmetric. Usefulness of ultrasound before catheterization. AB - OBJECTIVE: To demonstrate an asymmetry of the internal jugular veins, a finding which will have consequences for catheterization. DESIGN: Prospective study. SETTING: The medical ICU of a university-affiliated teaching hospital. PATIENTS: Eighty critically ill consecutive patients. INTERVENTION: Measurement of the cross-sectional area of the internal jugular veins. Search for an asymmetry, defined as an area at least twice that of the contralateral vein. RESULTS: An asymmetry was noted in 62.5% of the patients. The dominant vein was the right in only 68 % of these cases. In addition, 23% of the 160 jugular internal veins had an area of 0.4 cm2 or less. CONCLUSIONS: Using a simple technique, ultrasound identifies the dominant internal jugular vein, thus indicating the safer side before blind catheterization. PMID- 11280655 TI - Peripartum cardiomyopathy: a case series. AB - OBJECTIVES: We describe our experience with peripartum cardiomyopathy. DESIGN AND SETTING: A case series in intensive care units (ICU) of a district hospital and a referral center. PATIENTS: Six patients who required admission to an ICU after the onset of peripartum cardiomyopathy. RESULTS: Five of the six patients survived, with total recovery of ventricular function. After 1 year of follow-up all five survivors were symptom free with a normal ventricular function. CONCLUSIONS: There is a low rate of ICU admissions for peripartum cardiomyopathy, which has a potentially fatal prognosis. However, this disease can be detected by echocardiography among patients without the semiology. PMID- 11280656 TI - Delayed diagnosis of disseminated strongyloidiasis. AB - We report the case of a 51-year-old woman who presented with a confusing spectrum of systemic symptoms after starting steroid therapy for a rheumatological disorder. The diagnosis of disseminated strongyloidiasis was made after a delay of 2 weeks. This paper outlines the symptom complex with which this critically ill woman presented, the course of her disease and the treatment of her disseminated strongyloidiasis. PMID- 11280657 TI - Acute hepatic steatosis complicating massive insulin overdose and excessive glucose administration. AB - OBJECTIVE: To describe a case of acute hepatic steatosis due to excessive administration of glucose in the setting of massive insulin overdose, a complication which is rapidly and completely reversible if glucose infusion is rapidly tapered. DESIGN: Case report, clinical. SETTING: Intensive care unit, university hospital. PATIENT: A single patient admitted to the ICU. INTERVENTION: Intravenous glucose after insulin overdose. MEASUREMENTS AND MAIN RESULTS: On the 3rd day, increases in transaminase (ASAT 420 IU/l, ALAT 610 IU/l), bilirubin (147 mmol/l) and lactate (6.8 mmol/l), a decrease in arterial pH (7.32) and slightly increased liver size on ultrasound examination suggested acute hepatic steatosis. Clinical and laboratory abnormalities resolved rapidly after discontinuation of excessive glucose infusions (1,400 g/day for 3 days). CONCLUSIONS: Very large amounts of glucose after massive insulin overdose are potentially dangerous. Even though the fear of hypoglycemia-induced neurologic damage should be a constant preoccupation in this situation, glucose administration should be titrated on closely monitored blood glucose levels. PMID- 11280658 TI - Gastric tonometry. PMID- 11280659 TI - Intermittent positive pressure ventilation for the crushed chest: an epic in intensive care. PMID- 11280660 TI - About the rationale of the insertion of a Heimlich valve in the thoracostomy circuit in newborn infants and small children. PMID- 11280661 TI - Use of the Blue Rhino tracheostomy set for emergency airway management. PMID- 11280662 TI - Right-to-left interatrial shunt secondary to right ventricular myocardial infarction: a novel therapeutic approach. PMID- 11280663 TI - Acute respiratory failure after re-expansion pulmonary oedema localised to a lobe. PMID- 11280664 TI - Lethal heart failure caused by aluminium phosphide poisoning. PMID- 11280665 TI - Reversible myocardial dysfunction after exertional heat stroke. PMID- 11280666 TI - Belching and the development of Boerhaave's syndrome. PMID- 11280668 TI - An intermediate syndrome after parathion poisoning. PMID- 11280667 TI - An idiopathic total gastric necrosis. PMID- 11280669 TI - Clearance of arsenic by haemodialysis after acute poisoning with arsenic trioxide. PMID- 11280670 TI - Partial pressure of oxygen and partial pressure of carbon dioxide of perfluorocarbon liquid during partial liquid ventilation: their regional difference and their dependence on tidal volume and positive end-expiratory pressure level. AB - OBJECTIVE: To investigate the regional partial pressure of oxygen (PO2) and partial pressure of carbon dioxide (PCO2) of the perfluorocarbon liquid (PpfcO2, PpfcCO2) during partial liquid ventilation (PLV). DESIGN: Prospective, controlled study. SETTING: A research laboratory at a university medical center. SUBJECTS: Thirteen Japanese white rabbits. INTERVENTIONS: After the tracheostomy, PLV was started with perflubron (15 ml/kg) following saline lung lavage. Fractional inspired oxygen (FIO2) was 1.0, respiratory rate was 30 bpm and tidal volume (VT) was 30 ml. Two epidural catheters (18 gauge) were inserted from the rubber diaphragm interposed in the respiratory circuit to sample perflubron. One catheter was inserted into the left lower lobe bronchus and placed at 5-6 cm distal from the carina (DISTAL). The other one was inserted at the tip of the endotracheal tube (PROXIMAL). Then the effect of the larger VT (50 ml) or positive end-expiratory pressure (PEEP; 10 cmH2O) to the gas tension in perflubron was examined. MEASUREMENTS AND MAIN RESULTS: (1) In the lower VT (30 ml) with 0 cmH20 PEEP, DISTAL PpfcO2 was significantly lower than PROXIMAL PpfcO2 (265 72 vs 386 +/- 47 mmHg, p < 0.0001), and DISTAL PpfcCO2 was significantly higher than PROXIMAL PpfcCO2 (51.1 +/- 14.4 vs 42.4 +/- 11.8 mmHg (p = 0.0007)), (2) the higher VT setting increased PpfcO2 (p = 0.0001) and decreased PpfcCO2 (p < 0.0001), although the gas tension gradient was significant, (3) 10 cmH2O PEEP increased PpfcO2 (p = 0.0004) and decreased PpfcCO2 (p = 0.0089) in the DISTAL sample. CONCLUSION: There was a difference in gas tension in perflubron between the central airway and the peripheral dependent lung region, and gas tension in perflubron was affected by the VT and the PEEP level. PMID- 11280671 TI - Effect of inspiratory flow rate on beta2-agonist induced bronchodilation in mechanically ventilated COPD patients. AB - OBJECTIVES: To test the effect of two different inspiratory flow rates on the bronchodilation induced by beta2-agonists administered by metered dose inhaler (MDI). PATIENTS: Ten patients with acute exacerbation of chronic obstructive pulmonary disease and receiving mechanical ventilation with constant inspiratory flow (V'I). DESIGN: Patients received four puffs of salbutamol (100 microg/puff) with either low V'I (0.6 l/s) or high V'I (1.2 l/s) administered with an MDI adapted to inspiratory limb of the ventilator circuit using an aerosol cloud enhance spacer. After a 6-h washout patients were crossed-over to receive the drug by the alternative mode of administration. MEASUREMENTS AND RESULTS: Static and dynamic airway pressures, intrinsic positive end-expiratory pressure, and minimum and maximum inspiratory resistance values showed a significant decrease after salbutamol. These changes were not affected by the inspiratory flow rate and were evident 15, 30, and 60 min after administration. Heart rate, static end inspiratory respiratory system compliance, and the difference between minimum and maximum inspiratory resistance were unchanged after salbutamol. CONCLUSIONS: Salbutamol delivered by MDI and spacer device induces significant bronchodilation in mechanically ventilated patients with chronic obstructive pulmonary disease, but the magnitude of the effect is not affected by the inspiratory flow rate. These results do not support flow rate manipulations when bronchodilators are administered during controlled mechanical ventilation. PMID- 11280672 TI - Effects of patient-triggered automatic switching between mandatory and supported ventilation in the postoperative weaning period. AB - OBJECTIVE: To compare two ventilator settings in the postoperative weaning period. Patient-triggered automatic switching between controlled ventilation and supported spontaneous breathing (Automode, AM) was compared to synchronised intermittent mandatory ventilation (SIMV) with stepwise manual adjustment of mandatory frequency according to the breathing activity. DESIGN: Prospective clinical investigation. SETTING: Eighteen-bed intensive care unit in a university hospital. PATIENTS: Forty postoperative patients with healthy lungs who had undergone brain tumour surgery. INTERVENTIONS: Randomisation either to the AM or SIMV weaning procedure after entering the ICU. MEASUREMENTS AND RESULTS: Total weaning time and number of manipulations on the ventilator were observed. Cardiocirculatory and respiratory parameters were measured consecutively at five points during the weaning period. No significant differences were seen for cardiocirculatory parameters, airway pressures and oxygenation between the two groups. There was a trend to shorter weaning times with AM (136 +/- 46 min vs 169 +/- 68 min, n.s.), the average number of manipulations on the ventilator was lower (0.55 +/- 0.69 vs 5.05 +/- 1.19,p < 0.001) and arterial partial pressure of carbon dioxide (PaCO2) levels showed fewer variations in the late phase of the weaning period (39.5 +/- 3.1 vs 38.3 +/- 7.2, p < 0.001 for differences in variance). CONCLUSIONS: Automatic, patient-triggered switching between controlled and supported mode of ventilation can be used for postoperative weaning of neurosurgical patients with healthy lungs. Compared to a SIMV weaning procedure, fewer manipulations on the ventilator are necessary and individual adaptation of ventilation seems to be more accurate. PMID- 11280673 TI - Pattern of spontaneous breathing: potential marker for weaning outcome. Spontaneous breathing pattern and weaning from mechanical ventilation. AB - OBJECTIVE: To quantitatively assess the spontaneous breathing (SB) pattern, during minimal ventilatory support, of patients who pass or fail weaning trials from mechanical ventilation. DESIGN: A prospective, clinical trial. SETTING: Intensive care unit of a university teaching hospital. PATIENTS: Fifty-two tracheally intubated and hemodynamically stable patients who were judged clinically ready for extubation. METHODS: Using a computerized respiratory profile monitor, continuous respiratory parameters were obtained while patients were receiving four or less synchronized intermittent mandatory (SIMV) breaths and during CPAP trials. Coefficients of variation (CV) of spontaneous tidal volumes and flows during SIMV trials as well as the entropies and dimensions of the breathing patterns during CPAP trials were used to assess the dynamical breathing behaviors of the patients who passed or failed weaning trials. MEASUREMENTS AND RESULTS: Thirty-nine extubations were successful and 13 were not. The CV of the spontaneous tidal volumes (VT) and the spontaneous peak inspiratory flows (PF), the Kolmogorov entropy and the dimension of the SB patterns were compared in the two groups. The CV of VT (9.13 +/- 4.11 vs 26.07 +/ 6.94), the CV of PF (11.63 +/- 4.18 vs 29.88 +/- 12.07), the Kolmogorov entropy (0.09 +/- 0.03 bits/cycle vs 0.39 +/- 0.09 bits/cycle), and the dimension of the SB pattern (1.33 +/- 0.07 vs 3.93 +/- 0.47) were all significantly smaller (P < 0.05) in the successfully extubated group versus the group that failed extubation. CONCLUSION: The spontaneous breathing pattern during minimal mechanical ventilatory support is more chaotic in patients who failed extubation trials compared to patients who passed extubation trials. Thus, we speculate that characterizing the SB pattern during minimal ventilatory support might be a useful tool in differentiating between extubation success and failure. PMID- 11280674 TI - Chronic obstructive pulmonary disease patients with invasive pulmonary aspergillosis: benefits of intensive care? AB - OBJECTIVES: Invasive pulmonary aspergillosis (IPA) is increasingly recognized as a cause of acute respiratory failure in patients with chronic obstructive pulmonary disease (COPD) treated with corticosteroids. For these patients admission in intensive care unit (ICU) is often required for life-support and mechanical ventilation. Whether this approach improves outcome is unknown. DESIGN AND SETTING: Retrospective study in a university hospital intensive care unit. PATIENTS: Between November 1993 and December 1997, 23 COPD patients were admitted in our ICU and received antifungal agents for possible IPA. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: The clinical features and the outcome were reviewed. Diagnosis of IPA was classified as confirmed (positive lung tissue biopsy and/or autopsy) or probable (repeated isolation of Aspergillus from the airways with consistent clinical and radiological findings). Among the 23 patients treated for Aspergillus, 16 fulfilling these criteria for IPA were studied. Steroids had been administered at home to all patients but one and were increased during hospitalization in all. Twelve patients suffered a worsening of their bronchospasm precipitating acute respiratory failure. During ICU stay all patients required mechanical ventilation for acute respiratory failure. Although amphotericin B deoxycholate was started when IPA was suspected (0.5-1.5 mg/kg per day), all patients died in septic shock (n = 5) or in multiple-organ failure. CONCLUSIONS: The poor prognosis of intubated COPD patients with IPA, in spite of antifungal treatment suggests that further studies are required to define the limits and indications for ICU management of these patients. PMID- 11280676 TI - Prevalence of Helicobacter pylori infection in stress-induced gastric mucosal injury. AB - OBJECTIVE: To determine the role of Helicobacter pylori infection in critically ill patients admitted to the intensive care unit in the formation of gastric and duodenal mucosal injury in these patients. DESIGN AND SETTING: Prospective cohort analysis in an 18-bed mixed medical and surgical closed format ICU in a teaching hospital. PATIENTS: Fifty consecutive patients admitted to the intensive care unit for emergency reasons and requiring mechanical ventilation were included. INTERVENTIONS: H. pylori infection was detected by the laser-assisted ratio analyzer [13C]urea breath test (UBT). Gastric and duodenal mucosal lesions were assessed by upper gastrointestinal endoscopy and classified as minor (up to five erosions or submucosal hemorrhages) or major (more than five erosions or submucosal hemorrhages) mucosal injury. MEASUREMENTS AND MAIN RESULTS: Six patients were not eligible because the UBT could not be processed. Of the 44 eligible patients 22 were H. pylori positive by UBT and 22 H. pylori negative. Either minor or major gastric mucosal injury was found on endoscopy in 66 %. Of the 29 patients with minor mucosal injury 10 (34.5 %) were infected with H. pylori as indicated by positive LARA 13C-UBT. In contrast, of the 15 patients with major mucosal injury 12 (80%) were infected with H. pylori (p = 0.004). H. pylori was the only risk factor significantly associated with major mucosal injury in a multiple regression analysis (p = 0.019). CONCLUSION: The severity of gastric and duodenal mucosal injury in critically ill patients during mechanical ventilation is significantly correlated with the presence of H. pylori infection. PMID- 11280675 TI - Lactate: may I have your votes please? PMID- 11280677 TI - Base excess and lactate as prognostic indicators for patients admitted to intensive care. AB - OBJECTIVE: To examine whether values of arterial base excess or lactate taken on admission to a general intensive care unit indicate prognosis, and whether this can be used as a screening tool for future intensive care admissions. DESIGN: Observational study. SETTING: University teaching hospital general adult intensive care unit. PATIENTS: 148 consecutive patients admitted to the intensive care unit. INTERVENTIONS: Arterial blood samples were obtained on admission to the intensive care unit and 24 h following admission. MEASUREMENTS AND RESULTS: Arterial base excess and lactate concentrations were measured from the blood samples. Both base excess and arterial lactate samples on admission have good prognostic abilities (area under the curve on receiver operator characteristic analysis of 0.73, 0.78, respectively). The value of base excess on admission with the best predictive ability was a base excess more negative than -4 mmol/l, and the corresponding value for lactate was greater than 1.5 mmol/l. The combination of these two markers on admission to the intensive care unit led to a sensitivity of 80.3 % and a specificity of 58.7 % for mortality. The achievement of this combination was associated with an increased mortality (50.6 % vs. 15 %, p < 0.0001), older age (70 vs. 61.5 years, p < 0.05), a greater requirement for inotropic support (30.9 % vs. 4.5%, p < 0.0001) and higher organ failure scores both on admission and for the subsequent 24 h. CONCLUSIONS: Both base excess and lactate, or the combination of the two, can be used to predict outcome in patients admitted to the intensive care unit. These variables could be utilized to identify patients who have a high risk for mortality and thus who should be admitted to the intensive care unit. PMID- 11280678 TI - Plasma glutamine depletion and patient outcome in acute ICU admissions. AB - OBJECTIVE: To evaluate whether low plasma glutamine (PG) is related to severity of illness, and actual and predicted hospital mortality. DESIGN: Prospective cohort study. SETTING: 18-bed closed format general intensive care unit (ICU) of a teaching hospital. PATIENTS: Cohort of 80 seriously ill patients non-electively admitted to the ICU. INTERVENTIONS: Blood sampling for the determination of PG at ICU admission. MEASUREMENTS AND RESULTS: Severity of illness and predicted mortality were calculated using the locally validated APACHE II, SAPS II, and MPM II 0 and 24 systems. Illness scores, and actual and predicted hospital mortality were compared between patients with total PG < 0.420 mmol/l ("low PG") and patients with PG > or = 0.420 mmol/l. Mean total PG was 0.523 mmol/l, range 0.220 1.780 mmol/l. Low PG (n = 25) was associated with higher age (P = 0.03), shock as primary diagnosis, and higher actual hospital mortality (60 % vs 29 %, P = 0.01). Normal to high PG was associated with high plasma creatine phosphokinase (P = 0.007) There was a nonsignificant trend towards higher severity of illness scores and predicted mortality rates in the low PG group. The presence of low PG significantly improved mortality prediction when added as a factor to the APACHE II predicted mortality rate (P = 0.02). CONCLUSIONS: Low PG at acute ICU admission is related to higher age, shock as primary diagnosis, and higher hospital mortality. Low PG represents a risk of poor outcome, not fully reflected in the presently used mortality prediction systems. PMID- 11280680 TI - Is there a need for blood substitutes in the new millennium and what should we expect in the way of safety and efficacy? A research perspective. PMID- 11280679 TI - Influence of selenium supplements on the post-traumatic alterations of the thyroid axis: a placebo-controlled trial. AB - OBJECTIVE: To investigate whether early selenium (Se) supplementation can modify the post-traumatic alterations of thyroid hormone metabolism, since the first week after trauma is characterised by low plasma Se and negative Se balances. DESIGN: Prospective, placebo-controlled randomised supplementation trial. SETTING: Surgical ICU in a tertiary university hospital. PATIENTS: Thirty-one critically ill trauma patients aged 42 +/- 16 years (mean +/- SD), with severe multiple injury (Injury Severity Score 30 +/- 7). INTERVENTION: Supplementation during the first 5 days after injury with either Se or placebo. The selenium group was further randomised to receive daily 500 microg Se, with or without 150 mg alpha-tocopherol (AT) and 13 mg zinc supplements. The placebo group received the vehicle. Circulating Se, AT, zinc, and thyroid hormones were determined on D0 (= day 0, admission), D1, D2, D5, D10, and D20. RESULTS: Plasma Se, low on D0, normalised from D1 in the selenium group; total T4 and T3 increased more and faster after D2 (P = 0.04 and 0.08), reverse T3 rising less between D0 and D2 (P = 0.05). CONCLUSIONS: Selenium supplements increased the circulating Se levels. Supplementation was associated with modest changes in thyroid hormones, with an earlier normalisation of T4 and reverse T3 plasma levels. The addition of AT and zinc did not produce any additional change. PMID- 11280681 TI - Vasoconstrictive response of rat mesenteric arterioles following infusion of cross-linked, polymerized, and conjugated hemoglobin solutions. AB - Infusion of hemoglobin-based oxygen-carrying solutions (HBOCs) produce an immediate rise in blood pressure with most solutions, both in animals and humans, as a result of systemic and pulmonary vasoconstriction. Autoregulation of the O2 supply by the microvasculature has been proposed as a phenomenon involved in the vasoconstriction elicited by HBOCs. Nevertheless, little is known about the ability of various HBOCs to induce constriction in the microcirculation according to their specific physicochemical properties (viscosity, molecular weight, P50, etc.). This study was therefore designed to assess the effects of three HBOCs, that is, bis(3.5-dibromosalicyl) fumarate-crosslinked hemoglobin (alphaalpha-Hb), dextran-benzene-tetracarboxylate-conjugated hemoglobin (Hb-Dex-BTC) and o raffinose-oligomerized hemoglobin (o-raffinose-Hb), on the vascular tone of rat mesenteric arterioles (diameter, 15-25 microm) viewed microscopically in moderate hemodilution conditions. The effects of HBOCs were compared to those elicited by a reference solution of hydroxyethyl starch (HES-200) infused in the same conditions. In each experimental group, a fall in arteriolar diameter was observed 2 min and 5 min after infusion of the solution. The maximum changes were observed in Hb-Dex-BTC and o-raffinose-Hb groups, in which diameter decreased from 6.9 +/- 0.5% and 5.2 +/- 0.7%, respectively, 2 min after infusion. The changes in arteriolar diameter induced by Hb-Dex-BTC and o-raffinose-Hb were significantly higher than those elicited by HES-200 and alphaalpha-Hb. In conclusion, our data indicate that moderate hemodilution with HBOCs induces instantaneous constriction in rat mesenteric arterioles, with amplitudes depending on both pharmacological and physicochemical properties of the hemoglobin solution infused. PMID- 11280682 TI - A reverse microemulsion polymerization method for preparation of bioadhesive polyacrylic acid nanoparticles for mucosal drug delivery: loading and release of timolol maleate. AB - Polyacrylic acid nanoparticles were successfully synthesized using a reverse microemulsion polymerization process. They had a narrow size range, averaging approximately 50 nm, and were stable in buffer. The particles were isolated and lyophilized in dry powder form, and were redispersible as individual particles in buffer. The drug timolol maleate was loaded into the nanoparticles from aqueous drug solutions and, when the drug-loaded particles were dispersed in a phosphate buffer solution, the drug slowly released over several hours from the nanoparticles. PMID- 11280684 TI - Direct measurements of hemoglobin interactions with liposomes using EPR spectroscopy. AB - Electron paramagnetic resonance (EPR) spectroscopy was used to compare the rates of autoxidation at 37 degrees C of acellular and liposome-encapsulated hemoglobin (LEH) crosslinked between alpha chains with bis (3,5-dibromosalicyl) fumarate (alphaalphaHb). This method avoids the difficulties inherent in using conventional ultraviolet-visible (UV-vis) spectroscopy caused by the high turbidity of liposome suspensions. Rate constants of 0.039/h and 0.065/h were obtained for the alphaalphaHb and LEH samples, respectively. Similar oxidation measurements with alphaalphaHb using UV-vis spectroscopy gave a rate constant comparable to that obtained with EPR spectroscopy. Indirect measurement of the oxidation kinetics of LEH utilizing extraction of alphaalphaHb with chloroform from partially oxidized LEH samples was unreliable because the amount of extractable hemoglobin was inversely proportional to the degree of oxidation. EPR measurements showed a shift in the g value and substantial enhancement in the intensity of the bis-histidine low-spin B complex for the encapsulated hemoglobin, indicating a perturbation of this low-spin complex. We suggest that lipid-associated perturbations are responsible for the enhancement of the oxidation observed with the LEH samples compared to the unencapsulated material. PMID- 11280683 TI - Enhancement of microalgal growth by using perfluorocarbon as oxygen carrier. AB - The contribution of green microalgae has been recognized in the production of useful products such as chemicals, fatty acids, proteins, carotenoids, pigments, polysaccharides, and pharmaceuticals. One of the challenges to the development of profitable bioproduct markets is the design, development, modeling, and evaluation of cost-effective production systems. The photobioreactors for microalgae can be either open or closed systems in large-scale production. In various situations, it has been reported that closed photobioreactors offer many advantages over open systems. These advantages include lower contamination, higher biomass densities, and better process control. Nevertheless, for the scale up of enclosed tubular photobioreactors, the accumulation of oxygen as a photosynthetic byproduct has been a major limitation because it severely inhibits growth of microalgae. In this study, we use the distinctive feature of liquid perfluorocarbons (PFCs) as a carrier that efficiently removes the accumulated oxygen from algal medium phase in the spinner culture flasks. The results show the growth kinetics of microalgae cultivated by this approach is significantly improved. PMID- 11280685 TI - A system establishment compatibility profiles for artificial oxygen carriers and other substances. AB - Worldwide, great efforts are being made to develop a clinically useful artificial oxygen carrier. Toxicological and immunological compatibility is generally tested using animal experiments but inflammatory parameters in particular show large species-specific differences. Therefore, we developed an in vitro system using human components to establish a compatibility profile of unknown compounds. The test system comprises induction of hemolysis, activation of complement (C3a), induction/suppression of cytokine production, influence on cell proliferation, direct toxicity on peripheral leukocytes, and phagocytosis of the material under test and of microbes. The test system will be described, along with results of various perfluorocarbon emulsions. When testing lecithin-based perfluorodecalin (PFD) emulsions, and comparing them to Pluronic-based PFD emulsions, we could show that Pluronic-based emulsions were virtually untoxic to peripheral human leukocytes. They neither inhibited cell proliferation nor caused any hemolysis, but caused mild to moderate inhibition of endotoxin-induced cytokine production. At the same time, lecithin-based PFD emulsion caused substantial cytotoxicity in phagocytic cells like monocytes (60-100% after 24 h incubation) and granulocytes (10-20% after 24 h incubation). They also suppressed endotoxin-induced cytokine production in monocytes to more than 98% and inhibited cell proliferation of an endothelial (ECV 304) and a monocytic cell line (MonoMac6) to more than 95%. PMID- 11280686 TI - Semifluorinated symmetrical diethers for intraocular use: synthesis, characterization, and in vitro biocompatibility. AB - Semifluorinated symmetrical diethers were synthesized using the William ether synthesis. These diethers should have similar properties to perfluorocarbons as are chemical inertness and high oxygen solubility, but in contrast a considerably lower density. With their lower density the damaging of the choroidal tissue of the eye observed with perfluorocarbons should be avoided. The synthesized diethers are inert compounds being stable against nucleophiles, oxidiziers and strong bases. Their density is in the range of 1.1-1.2 g/cm3. Besides the physical and chemical tests we conducted several in vitro biocompatibility tests. The tests comprised induction of hemolysis, the generation of C3a complement, the influence on the production of interleukin1beta, the influence on cell proliferation of a Raji and a Hela cell line (3H-Thymidine uptake) and finally the direct cytotoxic effect on these cell lines. All tested symmetrical diethers were positive in one or more tests and can be expected to be incompatible in vivo. Especially the "short" semifluorinated diethers [(CF3CH2O)2(CH2)3-6] showed a nearly total inhibition of cell proliferation or interleukin1beta release. Further variation of the compounds will be necessary to generate better biocompatible derivates. PMID- 11280687 TI - Changes to the Pharmaceutical Benefits Advisory Committee. PMID- 11280688 TI - Time to move beyond clinical practice guidelines? PMID- 11280689 TI - Awareness during general anaesthesia: is it worth worrying about? PMID- 11280690 TI - Evidence-based medicine--time for a reality check. PMID- 11280691 TI - Randomised controlled trial to change the hospital management of unstable angina. AB - OBJECTIVES: To examine the benefits of a guideline-based educational program to improve management of unstable angina pectoris (UAP) in hospital patients. DESIGN: Randomised controlled trial. SETTING: 37 public hospitals across New South Wales. PATIENTS: 1,872 patients admitted with a diagnosis of UAP between 1 February and 30 June 1996 (baseline survey), and 1,368 patients with the same diagnosis admitted between 1 July and 31 December 1998 (follow-up survey). INTERVENTION: Educational sessions run by local opinion leaders, presenting guidelines on management of UAP from the National Health and Medical Research Council and feedback on local practice using data from the baseline survey. Sessions were run between March and June 1998. MAIN OUTCOME MEASURES: Use of evidence-based practice, identified by review of medical records. RESULTS: Use of beta-blockers increased in intervention and control hospitals, although the increase was significant only in the former. Use of calcium-channel blockers decreased significantly in both intervention and control hospitals. However, the change in drug use between baseline and follow-up did not differ significantly between intervention and control hospitals. CONCLUSIONS: Despite some appropriate changes in drug use for UAP management between 1996 and 1998, there was no evidence that a guideline-based educational program was of benefit in changing management. This reaffirms the difficulty of changing doctors' behaviour through practice guidelines. Alternative methods of encouraging evidence-based practice should be considered. PMID- 11280692 TI - Training general practitioners to recognise and respond to psychological distress and suicidal ideation in young people. AB - OBJECTIVE: To determine the effectiveness of a training program for general practitioners in recognising and responding to psychological distress and suicidal ideation in young people. DESIGN AND SETTING: The study, conducted in general practice surgeries in Tasmania, Victoria and Western Australia in 1996 and 1997, used a pre-/posttest design to audit consecutive young patients presenting in the six weeks before and the six weeks after the GPs' participation in the training program. PARTICIPANTS: Consisted of 23 GPs who attended a youth suicide prevention workshop and 423 patients aged 15-24 years who presented to the GPs' surgeries (203 pre-workshop and 220 post-workshop). INTERVENTION: GPs attended a one-day training workshop designed to enhance their ability to recognise, assess and manage young patients at risk of suicide. MAIN OUTCOME MEASURES: Scores on three patient self-report inventories (General Health Questionnaire-12 [GHQ-12], Center for Epidemiological Studies Depression Scale [CES-D] and Depressive Symptom Inventory--Suicidality Subscale [DSI-SS]); a GP completed form for each patient summarising presenting complaint(s), psychological assessment and proposed management plan. RESULTS: After training, GPs demonstrated increased recognition rates of psychologically distressed patients scoring above the cut-offs of the GHQ-12 (48% increase; odds ratio [OR], 1.748; 95% CI, 0.904-03.381) and CES-D (39.5% increase; OR, 2.067; 95% CI, 1.031 4.143); enquiry about suicidal ideation increased by 32.5% (OR, 1.483; 95% CI, 0.929-2.366); and identification of suicidal patients (determined by DSI-SS score) increased by 130% (OR, 3.949; 95% CI, 1.577-9.888). Training did not lead to any significant change in GPs' patient management strategies. CONCLUSIONS: A one-day training course can significantly enhance GP detection rates of psychological distress and suicidal ideation in young patients, but higher recognition rates do not necessarily lead to changes in patient management. PMID- 11280693 TI - Adverse events associated with rush hymenoptera venom immunotherapy. AB - OBJECTIVES: To determine the incidence and nature of adverse events associated with the induction of rush Hymenoptera venom immunotherapy. DESIGN: Retrospective descriptive case study. SETTING: The asthma and allergy unit at a major metropolitan teaching hospital, between 1 January 1989 and 30 June 1999. PATIENTS: All patients with anaphylaxis to stings of Hymenoptera insects who received rush venom immunotherapy as inpatients. OUTCOME MEASURES: Hypersensitivity reactions to venom administration, including angioedema, skin rashes, hypotension and asthma, as well as any other adverse events related to the inpatient stay. RESULTS: 68 venom-allergic patients received 73 courses of rush immunotherapy; 89% were desensitised to honey bee venom, 10% to yellow jacket wasp venom, and one to paper wasp venom. Hypersensitivity reactions occurred after 36 subcutaneous injections (3.8% of all injections given) in 26 patients (38%). CONCLUSION: In our cohort, immunotherapy was accompanied by a high incidence of adverse systemic events during the induction phase. Immunotherapy should only be given by experienced staff in centres where there are facilities for resuscitation. PMID- 11280695 TI - Non-valvular atrial fibrillation and stroke prevention. National Blood Pressure Advisory Committee of the National Heart Foundation. AB - Atrial fibrillation (AF) affects 5% of people older than 65 years. Among patients with AF, the risk of stroke averages about 5% per year. The risk of stroke increases cumulatively with increasing age, previous transient ischaemic attack or stroke, hypertension, diabetes, impaired left ventricular function and a large left atrium. Management aims to identify and treat the underlying cause, control the ventricular rate, restore and maintain sinus rhythm, and minimise the risk of stroke. Warfarin reduces the risk of stroke by about two-thirds, and aspirin by about one-fifth. The risk of anticoagulant-associated haemorrhage increases with serious concomitant disease, and with poorly controlled hypertension and poorly controlled anticoagulation. All patients with chronic AF should be considered for oral anticoagulant therapy, and the decision based on the balance between the risks of thromboembolism and bleeding. The recommended INR (international normalised ratio) is 2.0-3.0. Treating 1,000 "average" AF patients (ie, those with a 5% per year risk of stroke) with warfarin prevents about 30 strokes and causes at least two episodes of major haemorrhage each year. Treating 1,000 PMID- 11280694 TI - Firecracker injuries to the hand. AB - Between September 1999 and April 2000, the Hand Unit at St George Hospital, Sydney, treated three young men with severe injuries caused by holding a lighted firecracker. These cases illustrate the typical injuries seen with this mechanism of injury. They highlight the dangers of these explosive devices and the potential to improve the laws relating to fireworks. PMID- 11280696 TI - Disease control in the information era. AB - As a result of advances in information technology, there is now a new capacity to manage, interpret and apply data for the benefit not only of individual patients but of the population as a whole. Population health information systems are currently inadequate to meet the needs of disease control. In a rapidly changing world, effective public health action requires timely and efficient data about what is happening in the whole population. As the national effort to harness information technology to the needs of individual patient care begins, it is desirable that the electronic patient record also becomes the building block for public health research and monitoring. Individual healthcare and population healthcare should be two sides of the one coin. Ownership, privacy and access to the contents of the electronic health record should now be addressed in the context that disease control in the whole population will increasingly depend upon an efficient "real time" information system. PMID- 11280697 TI - Trichloroethylene and cancer: a carcinogen on trial. AB - The organic solvent trichloroethylene has been used in dry cleaning, as an industrial degreasing agent and as a solvent for oils and resins; large numbers of workers have been exposed to trichloroethylene, mainly by inhalation. Trichloroethylene has been categorised as a Group 2A carcinogen (probably carcinogenic to humans) by the International Agency for Research on Cancer (World Health Organization) and a Category 2 carcinogen (to be regarded as carcinogenic to humans) by the Australian National Industrial Chemicals Notification and Assessment Scheme. The Administrative Appeals Tribunal was asked to determine the validity of classifying trichloroethylene as a Category 2 rather than a Category 3 (data inadequate for making a satisfactory assessment) carcinogen. In the AAT's determination, relevant epidemiological evidence was not taken into account because such evidence concerned tumour sites apart from the kidney (the site of tumour induction by trichloroethylene in rats). This mode of evaluation is fundamentally different from that used by the International Agency for Research on Cancer. The precedent set by the consideration of carcinogenicity data in this case could have significant implications for classification of other putative carcinogens PMID- 11280698 TI - Evidence-based medicine: useful tools for decision making. AB - Evidence-based medicine (EBM) integrates clinical experience and patient values with the best available research information. There are four steps in incorporating the best available research evidence in decision making: asking answerable questions; accessing the best information; appraising the information for validity and relevance; and applying the information to patient care. Applying EBM to individual patients requires drawing up a balance sheet of benefits and harms based on research and individual patient data. The most realistic and efficient use of EBM by clinicians at the point of care involves accessing and applying valid and relevant summaries of research evidence (evidence-based guidelines and systematic reviews). The future holds promise for improved primary research, better EBM summaries, greater access to these summaries, and better implementation systems for evidence-based practice. Computer-assisted decision support tools for clinicians facilitate integration of individual patient data with the best available research data. PMID- 11280699 TI - Wasp sting mortality in Australia: one further case. PMID- 11280700 TI - Use of the term "indigenous". PMID- 11280701 TI - The new health insurance rebate. PMID- 11280702 TI - Psychotropic drugs and preschoolers. PMID- 11280703 TI - Adhesion between cells and extracellular matrix with special reference to hepatic stellate cell adhesion to three-dimensional collagen fibers. AB - Hepatic stellate cells are located in the perisinusoidal space (space of Disse), and extend their dendritic, thin membranous processes and fine fibrillar processes into this space. The stellate cells coexist with a three-dimensional extracellular matrix (ECM) in the perisinusoidal space. In turn the three dimensional structure of the ECM regulates the proliferation, morphology, and functions of the stellate cell. In this review, the morphology of sites of adhesion between hepatic stellate cells and extracellular matrix is described. Hepatic stellate cells cultured in polystyrene dishes spread well, whereas the cells cultured on or in type I collagen gel become slender and elongate their long cellular processes which adhere directly to the collagen fibers. Cells in type I collagen gel form a large number of adhesive structures, each adhesive area forming a face but not a point. Adhesion molecules, integrins, for the ECM are localized on the cell surface. Elongation of the cellular processes occurs via integrin-binding to type I collagen fibers. The signal transduction mechanism, including protein and phosphatidylinositol phosphorylation, is critical to induce and sustain the cellular processes. Information on the three dimensional structures of ECM is transmitted via three-dimensional adhesive structures containing the integrins. PMID- 11280704 TI - Ultrastructural, immunocytochemical and flow cytometry study of mouse peritoneal cells stimulated with carrageenan. AB - In the present paper we performed a morphological characterization of mouse peritoneal cells stimulated in vivo for 24 h with carrageenan (CAR) and lipopolysaccharide (LPS) by ultrastructural and flow cytometry analysis. In all samples, the flow cytometry studies showed the presence of three major populations consisting of monocytes, macrophages and lymphocytes. A special recruitment of monocytes was detected in CAR-injected mice. Macrophages and monocytes from CAR-treated mice displayed a characteristic phenotype, with a larger number of cytoplasmic vacuoles and numerous membrane projections, as compared to the cells collected from LPS- and PBS-injected mice. The induction of vacuolization was also confirmed upon in vitro treatment with CAR for 15 min to 24 h. The in vivo CAR-induced vacuoles were not related to lipid storage as judged by the lack of lipidic labeling after imidazole treatment at the ultrastructural level. In order to investigate the acidic nature of the vacuoles we used acidothropic probes, Lysotracker Yellow (LY) and Acridine Orange (AO). CAR injection activated the ability of peritoneal cells to incorporate LY around 2-5 times higher than control cells. However, the AO incorporation was 10-fold lower in CAR-stimulated cells than in LPS-stimulated ones. It is possible that the increase in intracellular vacuolization observed in CAR-stimulated cells could be related to exocytosis, since in most vacuoles the inflammatory protein MRP-14 was immunolocalized. The presence of MRP-14 in the culture supernatant of adherent peritoneal cells from CAR-injected mice was further comfirmed by ELISA, suggesting the discharge of MRP-14 enriched vacuole contents in the extracellular medium. We concluded that the morphological characteristics of activated monocytes and macrophages may depend on the nature of the triggering stimuli. Our observations reflect different functional phenotypes of monocytes/macrophages after in vivo stimulation with inflammatory agents such as CAR and LPS. PMID- 11280706 TI - Atomic force microscopic evidence for Z-band as a rigid disc fixing the sarcomere structure of skeletal muscle. AB - Atomic force microscopic images of single skeletal myofibrils showed periodical broad filamentous bands interspaced with narrow rigid bands corresponding to the sarcomere structures of skeletal muscle (Yoshikawa, Y., Yasuike, T., Yagi, A., and Yamada, T. 1999. Biochem. Biophys. Res. Comm., 256: 13-19). In order to identify the narrow rigid bands, comparative studies were made for intact single myofibrils and those treated with calcium-activated neutral protease by use of atomic force microscopy. It was found that (a) the periodical narrow rigid bands present in intact myofibrils were completely absent in myofibrils treated with calcium-activated neutral protease, and that (b) myofibrils treated with calcium activated neutral protease were very fragile compared with intact myofibrils. As calcium-activated neutral protease selectively removes Z-bands of myofibrils (Reddy, M. K., Etlinger, J. D., Rabinowitz, M., Fischman, D. A., and Zak, R. 1975. J. Biol. Chem., 250: 4278-4284), these results clearly indicate that (a) the narrow rigid bands are the Z-bands, and that (b) the Z-bands are the essential disc supporting the sarcomere structure of skeletal muscle. PMID- 11280705 TI - Scraped-wounding causes activation and association of C-Src tyrosine kinase with microtubules in cultured keratinocytes. AB - In order to elucidate the function of c-Src in keratinocytes, we studied the intracellular distribution of its active and inactive form in cultured normal human keratinocyte, using anti-c-Src monoclonal antibody clone 28, which recognizes the active form of c-Src (dephosphorylated at COOH-terminal residue Tyr 530), and monoclonal antibody clone 327 which recognizes both active and inactive forms. Since c-Src has been suggested to be involved in the control of cell adhesion in other cells, we produced a dynamic condition of cell migration by cutting culture cell colonies into squares to form a mesh pattern with a blade (culture wound model). Before cutting, the active form was expressed in cells located only at the periphery of colonies or isolated migrating cells, and was associated with microtubules. Wounding the colony generated a dramatic and rapid activation of c-Src in a few rows of cells along the cut edges, which were made even at the middle of colony, resulting in the association of the active form with microtubules. This increase of the active form was also detected by immunoblotting of cell extracts. These reactions were inhibited by 1 mM sodium orthovanadate, a protein-tyrosine phosphatase inhibitor. ST 638, a potent Src family tyrosine kinase inhibitor, inhibited the migration of keratinocytes in the culture wound healing model. These results suggest that wounding the culture causes activation of c-Src in keratinocytes, and thus activated c-Src may play a role in the function of microtubules during cell migration, especially at an early stage of wound healing. PMID- 11280707 TI - Leptin mediates a proliferative response in human gastric mucosa cells with functional receptor. AB - BACKGROUND: Recently, the rat stomach was reported as a source of leptin, a hormone mainly secreted by adipocytes. Also Helicobacter pylori-induced gastritis in humans was associated with locally elevated leptin levels. In addition, it was suggested that gastric leptin adjusts the function of the intestinal tract in parallel to the function of hypothalamic satiety centers. AIMS: Here we examined the synthesis and potential physiologic role of leptin in the human stomach. METHODS: RT-PCR was employed to detect leptin mRNA in the human stomach and the human gastric carcinoma cell line AGS while immunogold staining and electron microscopy were used to detect leptin protein. The in vitro effects of leptin on cell proliferation were examined in the AGS cell line. RESULTS: No leptin mRNA could be detected by RT-PCR, yet immunogold labeling and electron microscopy allowed visualization of leptin protein in the human gastric mucosa. At concentrations of 100 nM, leptin led to a significantly increased BrdU-uptake in AGS cells (+27%, p < 0.017). The MAP-kinase-1-specific inhibitor U0126 blocked the leptin-induced cell proliferation in a dose-dependent fashion. CONCLUSIONS: Leptin protein may not be produced but rather stored in human gastric cells. Leptin-induced increases in the proliferation of gastric mucosa cells suggests that leptin might contribute to mucosal integrity and gastroprotection. PMID- 11280708 TI - Aging and calcitriol regulation of IP3 production in rat skeletal muscle and intestine. AB - We previously reported that calcitriol [1,25(OH)2-vitamin D3] in rat skeletal muscle and duodenum stimulates the hydrolysis of polyphosphoinositides by phospholipase C (PLC), generating the second messengers inositol trisphosphate (IP3) and diacylglycerol (DAG), and that this mechanism is altered in old animals. As previously reported in muscle, we show in the present study that GTPgammaS (100 microM, 15 s), the non-hydrolyzable analogue of GTP, increased IP3 release from young rats duodenum to the same extent as 1 nM calcitriol (+ 100%), while GDPbetaS (100 microM) suppressed hormone-dependent IP3 production. Similarly to calcitriol, GTPgammaS response was diminished in old rats. Contrary to muscle, pretreatment with Bordetella pertussis toxin did not modify calcitriol dependent IP3 in duodenum. The antibody, anti-G alpha q/11 (1:200) and anti-G alpha i (1:200) blocked calcitriol-dependent IP3 release in muscle from young rats, indicating that the hormone activates an isoform of PLC coupled to the alpha subunit of Gq/11 and possibly the betagamma subunits of Gi. The aged muscle was insensitive to anti G alpha i. In rat duodenum the hormone effects were suppressed by anti-Gq/11 both in young and aged animals. In 24-month-old rats, Gq/11 and Gi protein levels were greatly reduced both in muscle and duodenum, suggesting that a deficiency in G protein expression with aging may have important consequences for correct receptor/effector coupling and could explain age-related declines in the function of second messenger systems linked to G proteins. PMID- 11280709 TI - Inhibition of T3 production in levothyroxine-treated female mice by the root extract of Convolvulus pluricaulis. AB - An investigation was made to evaluate the role of Convolvulus pluricaulis root extract in the regulation of hyperthyroidism in female mice. Its possible site of action was also studied. L-Thyroxine treatment for 30 days increased serum concentrations of thyroxine (T4) and triodothyronine (T3). The activity of hepatic 5'-monodeiodinase (5'-DI) and glucose-6-phosphatase (G-6-Pase) was also enhanced. On the other hand, administration of the plant extract either alone or with L-T4, decreased serum T3 concentration and the activity of hepatic 5'-DI and G-6-phase, without marked alteration in hepatic lipid peroxidation, indicating the possible regulation of hyperthyroidism by the plant extract. It appears that the action of the plant extract on thyroid function is primarily mediated through the inhibition of 5'-DI enzyme activity. PMID- 11280710 TI - Adrenomedullin and the blood-brain barrier. AB - Adrenomedullin (ADM) is present both in the periphery and brain. In addition to its peripheral effects, this peptide can exert central effects such as decreasing food ingestion. We used multiple-time regression analysis to determine that labeled ADM can cross from blood to brain with an apparent influx constant (K(I)) of 5.83 +/- 1.44 x 10(-4) ml/g-min, much faster than that of albumin, the vascular control. HPLC showed that almost all of the injected 125I-ADM in the brain was intact, and capillary depletion showed that it could reach the parenchyma of the brain. However, more 125I-ADM was reversibly associated with the brain vasculature than we have seen with any other peptide tested by these methods. After intracerebroventricular injection, 125I-ADM exited the brain with the bulk reabsorption of cerebrospinal fluid at an efflux rate comparable to that of albumin. Although there was no blood-to-brain saturation, in situ brain perfusion of 125I-ADM in blood-free physiological buffer showed self-inhibition by excess unlabeled ADM. This, along with evidence of the lack of protein binding shown by capillary zone electrophoresis, indicated competition for the binding site of ADM at the BBB. The low lipophilicity of ADM determined by the octanol/buffer partition coefficient was also consistent with the prominent reversible association of ADM with the vasculature of the BBB. This suggests a function for ADM at the cerebral blood vessels, such as altering cerebral blood flow and perfusion, without disruption of the BBB. PMID- 11280711 TI - The decrease of rat postprandial plasma triacylglycerol concentration after multiple cycles of starvation-refeeding. AB - The effect of multiple cycles of starvation-refeeding on rat body weight and on plasma lipid concentration was studied. After 1 cycle of starvation-refeeding, the rat body weight did not change significantly; however the postprandial plasma triacylglycerol concentration decreased approximately 2-fold as compared to rats fed ad libitum. After 8 cycles of starvation-refeeding, both rat body weight and plasma triacylglycerols concentration decreased. In contrast, the plasma cholesterol (both total and HDL cholesterol) concentration did not change appreciably either after 1 or 8 cycles of starvation-refeeding as compared to control. Although the postprandial plasma triacylglycerol concentration decreased in both groups (i.e. after 1 and 8 cycles of starvation-refeeding), this phenomenon appears to last longer after 8 cycles of starvation-refeeding. The epididymal white adipose tissue weight decreased after both 1 and 8 cycles of starvation-refeeding. After 1 cycle of starvation-refeeding followed by 3, 6 and 9 days of ad libitum feeding, the epididymal white adipose tissue weight increased progressively, reaching the control value at day 9. In contrast, after 8 cycles of starvation-refeeding followed by 9 days of ad libitum feeding, the epididymal white adipose tissue weight did not reach the control value. These results suggest that dieting is associated with body and adipose tissue weight loss as well as with the decrease of plasma triacylglycerol concentration. Furthermore, our results suggest that better maintenance of low adipose tissue weight and low plasma triacylglycerol concentration may be achieved after multiple cycles of starvation-refeeding. PMID- 11280712 TI - Influence of hepatic cells on allogeneic islet transplantation in rats without immunosuppressive drugs. AB - With the hypothesis of a possible helpful effect of the liver on islets transplanted into it, we have performed experiments that suggest some effect of hepatic cells for islet tolerance. We have studied 6 groups of Wistar rats made diabetic with streptozotocin and transplanted in sham conditions and with a mixture of islets and hepatic cells (allo-co-transplantation) in several conditions, all of them via the portal vein, and observed them over 30 days. Groups were as follows: Group A had a sham transplantation with saline. Group B was transplanted with hepatic cells alone. Group C was transplanted with islets alone without hepatic cells. Group D was co-transplanted with cultured islets and fresh hepatic cells (ratio 1:100). Group E was as group D with a ratio of 1:200. Group F also had co-transplantation, but after co-culture of islets and hepatic cells for 24 hours. Results show reversion of diabetes in group D for 4-5 days, and thereafter, a fall of blood glucose during the period observed. The effect was less marked in group F after co-culture of islets and hepatic cells. Paradoxically, when the ratio of islets and hepatic cell were 1:200, the results were not so good. These results suggest that hepatic cells have some helpful effect on islets when co-transplanted in the liver via the portal vein. More studies are needed to clarify if this effect can be related to some hepatic cell subpopulation; also if the effect is a membrane one, cell-to-cell contact, or through some secreted product. PMID- 11280713 TI - High antigenicity of intraperitoneal insulin infusion via implantable devices: preliminary rat studies. AB - Intraperitoneal insulin infusion of Genapol stabilized insulin via implantable devices significantly improves diabetes control and hypoglycemia frequency in type 1 diabetes while it increases insulin antibody levels. Causes for this particular antigenicity remain unknown. The role of insulin modifications occurring in the reservoir on the antigenicity observed was assessed by comparing the antigenicities of the insulin coming from the vial or from the pump reservoir. Rats were injected intraperitoneally with insulin sampled either from a vial (group 1) or from a pump reservoir during a refill of a clinical trial (group 2). Two control groups, one without insulin, the second one receiving a mixture of silicone and insulin were also studied. Human insulin antibody levels were assessed by RIA 10 days after 4 weekly immunizations. AIA levels were higher in group 1 compared to group 2 (P = 0.003 for the first experiment, P = 0.04 in the second experiment). The increased antigenicity of the insulin sampled from the implanted pump might be due to the insulin modifications occurring during the storage in the device. Insulin aggregates could be involved in this antigenicity since they are known to be antigenic and their concentration was shown to be related to the amplitude of the antigenic response. PMID- 11280714 TI - The pituitary response to ovine corticotropin-releasing hormone is enhanced in obese men and correlates with insulin resistance. AB - Hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis in central obesity has been demonstrated in women. We studied the corticotropin (ACTH) and cortisol response to ovine corticotropin releasing hormone (oCRH) and its association to parameters of adiposity and insulin resistance in a group of 19 healthy obese (BMI > 25 kg/m2) and 9 non-obese men. Relative insulin resistance was assessed by the homeostatic model assessment (HOMA IR). Baseline ACTH was similar, while cortisol was lower in the obese group. The ACTH response to oCRH was significantly higher in the obese group. ACTH incremental area under the curve (iAUC) correlated with age, HOMA IR, and sagittal diameter but not with leptin. In multiple regression analysis, only HOMA IR was an independent predictor of ACTH iAUC. In conclusion, obese men have hyperactivity of the HPA axis at the pituitary level, which appears to be linked to insulin resistance. PMID- 11280715 TI - Relationships of plasma C-peptide and gender to the urinary excretion of inositols in older people. AB - PURPOSE: The urinary excretions of myo-inositol and D-chiro-inositol are elevated in diabetes, and have been suggested as possible markers or effectors of insulin action. The aim of the present study was to measure the urinary excretion of these compounds, and to assess possible relationships with the metabolic control of glucose, in older, non-diabetic men and women. SUBJECTS: 32 older (age range 54-71 yrs), moderately overweight (body mass index 29.1 +/- 0.4 kg/m2, mean +/- SEM), non-diabetic men (n = 17) and women (n = 15). METHODS: 75 g oral glucose tolerance testing was done the day after all subjects had consumed nutrient defined menus for five days. Plasma samples were analyzed for the concentrations of glucose, insulin, and C-peptide, and the 180-minute area under the curve (AUC) for each of these compounds was calculated. Samples from 24-hour urine collections were analyzed for the concentrations of myo-inositol, D-chiro inositol, L-chiro-inositol, and pinitol. RESULTS: The fasting glucose, insulin, and C-peptide, and the AUC for glucose and insulin, were not different between men and women. C-peptide AUC was greater in the men versus the women (p < 0.001). The median urinary excretions (micromol/g creatinine) of myo-inositol (p < 0.001), D-chiro-inositol (p < 0.001), L-chiro-inositol (p < 0.05), and pinitol (p < 0.001) were higher, and the myo-inositol:D-chiro-inositol ratio was lower (p < 0.001), in the men versus women. For all subjects combined, C-peptide AUC was positively correlated with the urinary excretion of each of the measured inositols, as well as the myo-inositol:D-chiro-inositol ratio. The correlations between C-peptide AUC and these inositols were strongly influenced by the co linear relationship between C-peptide AUC and gender. CONCLUSIONS: Collectively, these data show that older, moderately overweight, non-diabetic men and women with gender-related differences in glucose-stimulated C-peptide AUC, an indirect indicator of insulin secretion, also display differences in the urinary excretion of myo-inositol, D-chiro-inositol, L-chiro-inositol, and pinitol. The gender related difference in the myo-inositol:D-chiro-inositol ratio suggests that, while the urinary excretion of all of the inositols measured were higher in the men than the women, the difference was more pronounced for D-chiro-inositol. PMID- 11280716 TI - A large family with hereditary MTC: role of RET genetic analysis in differential diagnosis between MEN 2A and FMTC. AB - Germline mutations of the RET proto-oncogene cause three different cancer syndromes: multiple endocrine neoplasia type 2A (MEN 2A), multiple endocrine neoplasia type 2B (MEN 2B) and familial medullary thyroid carcinoma (FMTC). In the absence of biochemical and/or clinical evidence of pheochromocytoma and hyperparathyroidism, patients with MEN 2A disease display the same phenotype of FMTC disease, although prognosis and clinical management in both affected and unaffected familial members are quite different. We studied a family with hereditary MTC, whose proband was referred to us because of enlarged cervical nodes and increased calcitonin serum levels 28 years after the total thyroidectomy for MTC. Cervical node dissection was carried out and subsequently the presence of MTC metastasis was histologically confirmed. A RET genomic mutation at codon 634 (TGC-->TTC) was identified in the proband and in seven out of 19 familial members studied. Accordingly, a hereditary disease was suggested. However, the strong association of RET mutation at codon 634 with the presence of pheochromocytoma in MEN 2 disease suggested a more rigorous management in all gene carriers. Indeed, during the follow-up pheochromocytoma was subsequently identified in the proband. This finding suggests that all families with a pedigree suggestive of FMTC should be regarded at risk from MEN 2A disease, at least when a critical mutation in the RET cysteine domain is detected. PMID- 11280717 TI - Nutrient regulation of post-heparin lipoprotein lipase activity in obese subjects. AB - This study examines the immediate effect of ingestion of oral carbohydrate and fat on lipoprotein lipase (LPL) activity post-heparin in six lean and six obese age-matched women. Subjects were given, on two separate occasions, 340 kcal carbohydrate or an equicaloric amount of fat, both in 300 ml of water. Post heparin LPL activity (10,000 U) was measured on each occasion 120 minutes after ingestion of the meal. Following oral carbohydrate postprandial plasma insulin levels were significantly higher in obese subjects than in lean (p < 0.01). Impaired glucose tolerance was seen in the obese group. GIP secretion was similar in lean and obese subjects both during oral fat and carbohydrate ingestion. GLP-1 secretion post-carbohydrate was lower in obese subjects. Total LPL activity unadjusted for body weight was similar in the two groups after carbohydrate administration but was significantly lower when adjusted per kg body weight. Total LPL activity was lower in the lean group at 130 minutes after fat administration (p < 0.02). Fasting serum triglycerides were higher in the obese group and were inversely related to the post-carbohydrate LPL activity (r = - 0.65, p < 0.02). Intraluminal lipoprotein lipase activity is not increased in established obesity. Fat and carbohydrate nutrients may affect LPL activity differently in lean and obese subjects. PMID- 11280718 TI - Evidence of sex hormone binding globulin binding sites in the medial preoptic area and hypothalamus. AB - We have demonstrated a high density of both radiolabeled progesterone and estradiol conjugated to bovine serum albumin binding sites in the medial preoptic area and hypothalamus. Infusions of sex hormone binding globulin into the medial preoptic area of rats increased their female sexual receptivity similarly to the effect of estradiol conjugated to bovine serum albumin, suggesting sex hormone binding globulin acts at binding sites for estradiol conjugated to bovine serum albumin. In this study sex hormone binding globulin was used to displace radiolabeled progesterone conjugated to bovine serum albumin from plasma membrane fractions from the medial preoptic area-anterior hypothalamus and medial basal hypothalamus of ovariectomized rats injected with either 5 microg estradiol benzoate or sesame oil vehicle. We found that sex hormone binding displaced radiolabeled progesterone conjugated to bovine serum albumin in both areas and that in vivo estradiol treatment greatly increased the relative displacement by sex hormone binding globulin in the medial preoptic area-anterior hypothalamus. We interpret these data as indicating the presence of sex hormone binding globulin receptors in brain plasma membranes and further suggest that endogenous steroid conditions may alter these receptors. PMID- 11280719 TI - Breast cancer metastatic potential correlates with a breakdown in homospecific and heterospecific gap junctional intercellular communication. AB - Breast cancer progresses toward increasingly malignant behavior in tumorigenic and metastatic stages. In the series of events in the metastatic stage, tumor cells leave the primary tumor in breast and travel to distant sites where they establish secondary tumors, or metastases. In this report, we demonstrate that cell-cell communication via gap junctions is restored in the metastatic human breast carcinoma cell line MDA-MB-435 when it is transfected with breast metastasis suppressor 1 (BRMS1) cDNA. Furthermore, the expression profile of connexins (Cxs), the protein subunits of gap junctions, changes. Specifically, the expression of BRMS1 in MDA-MB-435 cells increases Cx43 expression and reduces Cx32 expression, resulting in a gap junction phenotype more similar to normal breast tissue. Taken together, these results suggest that gap junctional communication and the Cx expression profile may contribute to the metastatic potential of these breast cancer cells. PMID- 11280720 TI - The 104-123 amino acid sequence of the beta-domain of von Hippel-Lindau gene product is sufficient to inhibit renal tumor growth and invasion. AB - The von Hippel-Lindau (VHL) tumor suppressor gene is mutated in patients with VHL disease and in the majority of patients with sporadic renal cell carcinomas (RCCs). RCCs are dependent on insulin-like growth factor-1 receptor-mediated signaling for tumor growth and invasion in vivo. Reintroduction of the VHL gene product (pVHL) can inhibit on insulin-like growth factor-I receptor-mediated signaling in RCC cells in vitro through interaction with protein kinase C delta and is mediated by a specific amino acid sequence (104-123) in the beta-domain of the pVHL. In the present study, the amino acid sequence (104-123) of the pVHL was conjugated to the protein transduction domain of HIV-TAT protein (TATFLAGVHL peptide) to facilitate entry into cells, and we demonstrate that this amino acid region of VHL is sufficient to block proliferation and invasion of 786-O renal cancer cells in vitro. Furthermore, daily i.p. injections with the TATFLAGVHL peptide retarded and, in some cases, caused partial regression of renal tumors that were implanted in the dorsal flank of nude mice. Treatment with this peptide also inhibits the invasiveness of renal tumors. A 56% decrease in the proliferative index in tumors treated with the TATFLAGVHL-peptide versus control peptide-treated mice was observed. Taken together, these results show the novel importance of a 20-amino acid sequence of the beta-domain of the VHL gene product capable of inhibiting tumor growth and invasion. These results lay the foundation for a unique approach toward treating RCCs using this small-molecular-weight peptide fused to the TAT-sequence, which may, in the future, be used alone or in conjunction with other therapies. PMID- 11280721 TI - Suppression of a squamous cell carcinoma (SCC)-related serpin, SCC antigen, inhibits tumor growth with increased intratumor infiltration of natural killer cells. AB - Squamous cell carcinoma (SCC) antigen (SCCA), a member of the ovalbumin serine proteinase inhibitor family, serves as a circulating marker of squamous cell carcinoma (SC). One of the SCCAs, SCCA1, has been suggested to play a role in the attenuation of apoptosis in vitro and in the augmentation of tumor growth in vivo. In the present study, the infection of a SCC cell line (SKG IIIa) with recombinant retrovirus that expressed the antisense SCCA mRNA suppressed expression of SCCA in vitro. Local administration of this retrovirus into tumors by inoculation in nude mice suppressed tumor growth. Treatment of tumor tissue in vivo is also associated with increased numbers of apoptotic tumor cells and large mononuclear cells in the tumor. To test the possible role of SCCA in the infiltration of large mononuclear cells, we analyzed the effect of SCCA1 on migration of natural killer (NK) cells induced by monocyte-chemoattractant protein-1 in vitro. SCCA1 suppressed migration of NK cells completely, and this inhibitory effect was lost by mutation of the reactive site loop of SCCA1. These results suggest that antisense SCCA may suppress the growth of SCC in vivo not only by the augmentation of intracellular apoptosis but also by the increased infiltration of NK cells into the tumor. PMID- 11280722 TI - Noninvasive imaging of alpha(v)beta3 integrin expression using 18F-labeled RGD containing glycopeptide and positron emission tomography. AB - The alpha(v)beta3 integrin is an important cell adhesion receptor involved in tumor-induced angiogenesis and tumor metastasis. Here we describe the 18F labeling of the RGD-containing glycopeptide cyclo(-Arg-Gly-Asp-D-Phe-Lys(sugar amino acid)-) with 4-nitrophenyl 2-[18F]fluoropropionate and the evaluation of this compound in vitro and in tumor mouse models. Binding assays with isolated immobilized alpha(v)beta3, alpha(v)beta5, and alpha(IIb)beta3 as well as in vivo studies using alpha(v)beta3-positive and -negative murine and xenotransplanted human tumors demonstrated receptor-specific binding of the radiolabeled glycopeptide yielding high tumor:background ratios (e.g., 120 min postinjection: tumor:blood, 27.5; tumor:muscle, 10.2). First imaging results using a small animal positron emission tomograph suggest that this compound is suitable for noninvasive determination of the alpha(v)beta3 integrin status and therapy monitoring. PMID- 11280723 TI - Vascular endothelial growth factor C promotes tumor lymphangiogenesis and intralymphatic tumor growth. AB - Many solid tumors produce vascular endothelial growth factor C (VEGF-C), and its receptor, VEGFR-3, is expressed in tumor blood vessels. To study the role of VEGF C in tumorigenesis, we implanted MCF-7 human breast carcinoma cells overexpressing recombinant VEGF-C orthotopically into severe combined immunodeficient mice. VEGF-C increased tumor growth, but unlike VEGF, it had little effect on tumor angiogenesis. Instead, VEGF-C strongly promoted the growth of tumor-associated lymphatic vessels, which in the tumor periphery were commonly infiltrated with the tumor cells. These effects of VEGF-C were inhibited by a soluble VEGFR-3 fusion protein. Our data suggest that VEGF-C facilitates tumor metastasis via the lymphatic vessels and that tumor spread can be inhibited by blocking the interaction between VEGF-C and its receptor. PMID- 11280725 TI - Adenovirus expressing RIZ1 in tumor suppressor gene therapy of microsatellite unstable colorectal cancers. AB - Viral vector-mediated delivery of tumor suppressor genes represents a promising strategy of cancer therapy. Several best-studied tumor suppressor genes, such as p53 and retinoblastoma (Rb), have been evaluated for gene therapy of tumors that carry mutations in these genes. However, these genes may not be applicable to microsatellite instability positive [MSI(+)] tumors because they are rarely mutated in these tumors. The Rb-interacting zinc finger gene RIZ1 is commonly mutated in MSI(+) colorectal, gastric, and endometrial cancers and has demonstrated a capacity to induce cell cycle arrest and apoptosis. Here, we found that RIZ1 expression through adenovirus vectors suppressed growth of MSI(+) HCT116 colorectal xenograft tumors that carry RIZ1 mutations. Malignant cells in the established tumors were efficiently transduced by RIZ1 adenoviruses and underwent apoptosis in response to RIZ1 expression. In comparison, a recombinant p53 adenovirus did not induce apoptosis and tumor suppression. These results suggest that RIZ1 may be useful in gene therapy of MSI(+) colorectal cancers. PMID- 11280724 TI - Silica-induced activation of c-Jun-NH2-terminal amino kinases, protracted expression of the activator protein-1 proto-oncogene, fra-1, and S-phase alterations are mediated via oxidative stress. AB - Crystalline silica has been classified as a group 1 human carcinogen in the lung. However, its mechanisms of action on pulmonary epithelial cells which give rise to lung cancers are unclear. Using a nontransformed alveolar type II epithelial cell line (C10), we show that alpha-quartz silica causes persistent dose-related increases in phosphorylation of c-Jun-NH2-terminal amino kinases (JNKs) that are inhibited by antioxidants (P < or = 0.05). Increases in activator protein-1 (AP 1) binding to DNA and transactivation of AP-1-dependent gene expression by silica were accompanied by increases in steady-state mRNA levels of the AP-1 family members, c-jun, junB, fra-1, and c-fos at 8 h and elevated mRNA levels of fra-1 at 24 h (P < or = 0.05). Addition of tetramethylthiourea inhibited silica associated increases infra-1 and proportions of cells in S-phase (P < or = .05). Our findings indicate that silica induces JNK activity, AP-1-dependent gene expression, ie., fra-1, and DNA synthesis via oxidative stress. Moreover, they suggest that silica may act mechanistically as a mitogen or tumor promoter, rather than a genotoxic carcinogen, in the development of lung cancers. PMID- 11280726 TI - Geldanamycin and its analogue 17-allylamino-17-demethoxygeldanamycin lowers Bcr Abl levels and induces apoptosis and differentiation of Bcr-Abl-positive human leukemic blasts. AB - HL-60/Bcr-Abl cells, with ectopic expression of p185 Bcr-Abl tyrosine kinase (TK), and K562 cells, with endogenous expression of p210 Bcr-Abl TK, display a high degree of resistance against antileukemic drug-induced apoptosis (G. Fang et al., Blood, 96: 2246-2256, 2000). Present studies demonstrate that treatment with ansamycin antibiotic geldanamycin (GA), or its less toxic analogue 17-allylamino 17-demethoxygeldanamycin (17-AAG), induces cytosolic accumulation of cytochrome c and cleavage and activities of caspase-9 and caspase-3, triggering apoptosis of HL-60/Bcr-Abl and K562 cells. GA or 17-AAG down-regulated intracellular Bcr-Abl and c-Raf protein levels, as well as reduced Akt kinase activity. Similar to Raf 1, v-Src, and Her-2-neu, Bcr-Abl TK has chaperone association with heat shock protein 90 (Hsp90). By binding and inhibiting Hsp90, GA or 17-AAG treatment shifted the binding of Bcr-Abl from Hsp90 to Hsp70 and induced the proteasomal degradation of Bcr-Abl, because cotreatment with proteasome inhibitor PSC341 reduced both GA (or 17-AAG)-mediated down-regulation of Bcr-Abl levels and inhibited apoptosis of HL-60/Bcr-Abl and K562 cells. These data establish the in vitro activity of GA and 17-AAG against Bcr-Abl-positive leukemic cells and support the in vivo investigation of 17-AAG against Bcr-Abl-positive leukemias. PMID- 11280727 TI - Protease pretreatment increases the efficacy of adenovirus-mediated gene therapy for the treatment of an experimental glioblastoma model. AB - Effective virus-mediated gene therapy for cancer will be facilitated by procedures that enhance the low level of gene transfer mediated by replication deficient, recombinant viral vectors. We found recently that protease pretreatment of solid tumors is a useful strategy for enhancing virus-mediated gene transduction in vivo. In this study, we examined the potential of protease pretreatment to improve the efficacy of a gene therapy strategy for prodrug activation that depends on infection with a recombinant adenovirus encoding herpes simplex virus thymidine kinase (Ad-HSV-tk). Trypsin or a dissolved mixture of collagenase/dispase was inoculated into xenografts derived from the human glioblastoma multiforme-derived cell lines, U87 or U251. Ad-HSV-tk was administered 24 h after protease pretreatment, and animals were then treated for 10 days with ganciclovir (GCV). We found that protease pretreatment increased the efficacy of adenovirus mediated HSV-tk/GCV gene therapy in these experimental tumor models. Mice receiving Ad-HSV-tk/GCV after protease pretreatment demonstrated a significantly greater regression of tumors compared with those treated with Ad-HSV-tk/GCV alone. No adverse effects of protease pretreatment were observed. No signs of metastasis were seen either by histological inspection of lymph nodes or by a PCR-based analysis of selected mouse tissues to detect human tumor cells. Our findings indicate that protease pretreatment may be a useful strategy to enhance the efficacy of virus-mediated cancer gene therapy. PMID- 11280728 TI - Proline oxidase, encoded by p53-induced gene-6, catalyzes the generation of proline-dependent reactive oxygen species. AB - The p53-dependent initiation of apoptosis is accompanied by the induction of proline oxidase (POX), a mitochondrial enzyme catalyzing the conversion of proline to pyrroline-5-carboxylate with the concomitant transfer of electrons to cytochrome c. However, the contribution of increased POX activity to apoptosis, if any, remains unknown. Using Adriamycin to initiate p53-dependent apoptosis, we showed that the expression of POX is up-regulated in a time- and dose-dependent manner in a human colon cancer cell line (LoVo). In cells expressing POX, the addition of proline increases reactive oxygen species (ROS) generation in a concentration-dependent manner; glutamate, a downstream product of proline oxidation, had no effect. Induction of POX was dependent on the p53 status of the cell. In the conditionally immortalized murine colonic epithelial cell line YAMC, where the p53 phenotype can be modulated by temperature, proline oxidase expression and ROS production could only be induced when the cells were phenotypically p53-positive. To confirm that the observed ROS production was not secondary to some other effect of p53, we also conditionally expressed POX in a p53-negative colon cancer line. Again, we found a proline-dependent ROS increase with POX expression. We hypothesize that proline oxidation supports the generation of ROS by donating reducing potential to an electron transport chain altered either by p53-dependent mechanisms or by overexpression of POX. PMID- 11280729 TI - Tumorigenic effect of nonfunctional p53 or p21 in mice mutant in the Werner syndrome helicase. AB - Werner syndrome is an autosomal recessive disorder characterized by genomic instability and by the premature onset of a number of age-related diseases, including malignancy. To assess a potential collaboration between p21 or p53 cell cycle regulators and Wrn proteins, Wrn mutant mice were created and mated with p21 or p53 null mice to generate double mutants. The p21 null/Wrn mutant mice did not show an acceleration of tumorigenesis during the first year of life, suggesting that the p53-dependent G1-S cell cycle checkpoint (which operates via p21) is not involved in Wrn-abetted tumor suppression. In contrast, the p53 null/Wrn mutant mice were particularly remarkable with respect to the rapidity with which they developed tumors. These mice were also distinguished by the variety of tumors they developed compared to those that developed in p53 null mice. Such data suggest a genetic interaction between p53 and Wrn in which loss of Wrn provokes a more variable p53 response unrelated to its role in the G1-S cell cycle checkpoint. PMID- 11280730 TI - Elevated breast cancer risk in irradiated BALB/c mice associates with unique functional polymorphism of the Prkdc (DNA-dependent protein kinase catalytic subunit) gene. AB - Female BALB/c mice are unusually radiosensitive and more susceptible than C57BL/6 and other tested inbred mice to ionizing radiation (IR)-induced mammary tumors. This breast cancer susceptibility is correlated with elevated susceptibility for mammary cell transformation and genomic instability following irradiation. In this study, we report the identification of two BALB/c strain-specific polymorphisms in the coding region of Prkdc, the gene encoding the DNA-dependent protein kinase catalytic subunit, which is known to be involved in DNA double stranded break repair and post-IR signal transduction. First, we identified an A -> G transition at base 11530 resulting in a Met --> Val conversion at codon 3844 (M3844V) in the phosphatidylinositol 3-kinase domain upstream of the scid mutation (Y4046X). Second, we identified a C --> T transition at base 6418 resulting in an Arg --> Cys conversion at codon 2140 (R2140C) downstream of the putative leucine zipper domain. This unique PrkdcBALB variant gene is shown to be associated with decreased DNA-dependent protein kinase catalytic subunit activity and with increased susceptibility to IR-induced genomic instability in primary mammary epithelial cells. The data provide the first evidence that naturally arising allelic variation in a mouse DNA damage response gene may associate with IR response and breast cancer risk. PMID- 11280731 TI - Gene expression profiling with oligonucleotide microarrays distinguishes World Health Organization grade of oligodendrogliomas. AB - Oligodendrogliomas are the second most common type of glial neoplasm with distinct prognostic and therapeutic implications. Although refinements have led to improved clinical stratification, current grading schemes are still limited by subjective histopathological criteria. In this report, we have used oligonucleotide array technology to perform expression profiling in morphologically classic oligodendrogliomas. Expression information from approximately 1100 genes divided tumors into two molecularly distinct groups that corresponded exactly to their previously assigned histological grades. Subsequent gene clustering identified a subset of 196 transcripts showing a common, differential expression pattern between tumor grades. A number of these genes have been associated with the maintenance of cytoarchitecture, cellular differentiation and maturation, immunogenicity, and chemotherapeutic resistance. These results demonstrate the utility of gene expression profiling as an objective, ancillary tool for grading oligodendrogliomas and a potential approach for classifying diffuse gliomas where histological assessment may be difficult or ambiguous. PMID- 11280732 TI - Hypoxia inducible factor (HIF-1a and HIF-2a) expression in early esophageal cancer and response to photodynamic therapy and radiotherapy. AB - Hypoxia inducible factor 1a and 2a (HIF-1a and HIF-2a) are key proteins regulating cellular response to hypoxia. Because the efficacy of photodynamic therapy (PDT) is dependent on the presence of oxygen, the assessment of HIF-1a and HIF-2a expression may be of value in predicting clinical response to PDT. Using recently produced MoAbs, we examined the expression of HIF1a and HIF2a in a series of 37 early-stage esophageal cancers treated with PDT and with additional radiotherapy in case of incomplete response after PDT. Strong expression of the HIF1a and of HIF2a proteins in all optical fields examined was noted in 51% and in 13% of cases, respectively. High expression was associated with a low complete response (CR) rate and with the absence of bcl-2 protein expression. On the contrary, bcl-2 expression was associated with a high CR rate. Combined analysis of HIF1a and bcl-2 protein expression revealed that of 16 cases with high HIF1a expression and the absence of bcl-2 reactivity, only 1 (7%) responded completely to PDT (P = 0.007). Bivariate analysis showed that HIF1a expression was independently related to response to PDT (P = 0.04; t ratio = 2.8), whereas bcl-2 approached significance (P = 0.07; t-ratio = 1.8). The final response to radiotherapy was high (70%) and independent of the HIF and bcl-2 status, which may be a result of reoxygenation after cellular depletion mediated by PDT. The present study suggests that assessment of HIF and of bcl-2 expression are important predictors of in vivo sensitivity to PDT. Modulation of PDT response with bioreductive drugs and/or drugs targeting bcl-2 (i.e., taxanes) may prove of significant therapeutic importance in a subgroup of patients with high HIF expression. PMID- 11280733 TI - Invasion-specific genes in malignancy: serial analysis of gene expression comparisons of primary and passaged cancers. AB - The invasive growth of malignant cells induces an admixture of host reactions including desmoplasia, angiogenesis, and immune reactions Pancreatic cancer has a prominent and characteristic host reaction at the site of primary invasion. To obtain new insights into the process of tumor invasion, we studied global patterns of gene expression using serial analysis of gene expression in pancreatic cancer, with extension to other tumor types. Here we report a cluster of invasion-specific genes in pancreatic and other cancers. This cluster contains genes that derive from distinct components of the host reaction, including some that may be useful as diagnostic markers and therapeutic targets. PMID- 11280734 TI - Aberrant expression of HDMX proteins in tumor cells correlates with wild-type p53. AB - It has been shown that the Hdmx gene is amplified in a subset of gliomas, but thus far, no data are available on HDMX protein expression in tumor cells. We now report that a significant fraction of tumor cell lines expresses increased HDMX levels compared with normal cells; in general, HDMX expression in these tumor cell lines correlates with the presence of wild-type p53. Analysis of tumor material showed that high HDMX expression is not a result of cell line establishment. Interestingly, several cell lines express alternative, shorter HDMX proteins. These results suggest that deregulated expression of HDMX plays a role in carcinogenesis as an alternative way to inactivate p53. PMID- 11280735 TI - Somatic mutation of mitochondrial DNA in cancerous and noncancerous liver tissue in individuals with hepatocellular carcinoma. AB - Unlike other types of cancer, hepatocellular carcinoma (HCC) is usually preceded by chronic inflammation caused by viral infection. The mutation of mitochondrial DNA (mtDNA) in hepatocarcinogenesis associated with viral infection was investigated. Compared with control liver tissue, the frequency of mtDNA mutations was markedly increased in both noncancerous and cancerous liver specimens from individuals with HCC. The accumulation of mtDNA mutations in HCC tissue reflected the degree of malignancy. The frequency of mtDNA mutations in HCC tissue was also greater than that described previously for other types of tumors. These observations suggest that the repeated destruction and regeneration of liver tissue associated with chronic viral hepatitis lead to the accumulation of mtDNA mutations. The genetic instability that results in the high rate of mtDNA mutation in cancerous liver tissue is also consistent with the multicentric hepatocarcinogenesis detected clinically. PMID- 11280736 TI - Target cell-restricted triggering of the CD95 (APO-1/Fas) death receptor with bispecific antibody fragments. AB - Like many other cell surface receptors, the CD95 (APO-1/Fas) molecule needs to be cross-linked by its physiological ligand or by immobilized or multimeric antibodies to mediate biological activity, that is, induction of apoptotic cell death. Monomeric CD95 antibodies of the IgG2a or IgG1 subtype block rather than induce apoptosis. We report here that such antibodies, hybridized to a second antibody directed against a different target antigen on the same cell, effectively induce apoptosis of the cells if the expression of the target antigen exceeds a certain threshold level. It appears that this effect is due to bicellular binding of bispecific antibodies resulting in mutual cross-linking of the CD95 death receptor and the target antigen. Using bispecific reagents, it may therefore be possible to restrict the activation of death receptors to a given target site, e.g., a tumor. In general terms, our findings illustrate a principle according to which the triggering of a cell surface receptor may be confined to a given target cell using bispecific reagents with target X cell surface receptor specificity. PMID- 11280737 TI - Increased oxidative stress in ataxia telangiectasia evidenced by alterations in redox state of brains from Atm-deficient mice. AB - Ataxia-telangiectasia (A-T) is a genetic disorder caused by mutational inactivation of the ATM gene. A-T patients display a pleiotropic phenotype and suffer primarily from progressive ataxia caused by degeneration of cerebellar Purkinje and granule neurons. Disruption of the mouse Atm locus creates a murine model of A-T that exhibits most of the clinical features of the human disease. We previously hypothesized that some aspects of A-T, such as the preferential loss of certain neurons, could result from a continuous state of increased oxidative stress (G. Rotman and Y. Shiloh, Cancer Surv., 29: 285-304, 1997; G. Rotman and Y. Shiloh, BioEssays, 19: 911-917, 1997). The present work tests this hypothesis by analyzing markers of redox state in brains of Atm-deficient mice. We found alterations in the levels of thiol-containing compounds in Atm (-/-) brains, as well as significant changes in the activities of thioredoxin, catalase, and manganese superoxide dismutase in Atm (-/-) cerebella. These changes are indicative of increased levels of reactive oxygen species, which are seen primarily in the cerebellum of Atm-deficient mice. Our findings support the hypothesis that the absence of functional ATM results in oxidative stress, which may be an important cause of the degeneration of cerebellar neurons in A-T. PMID- 11280739 TI - XIAP regulates Akt activity and caspase-3-dependent cleavage during cisplatin induced apoptosis in human ovarian epithelial cancer cells. AB - Chemoresistance is a major hurdle for successful cancer therapy. Although multiple mechanisms have been implicated to be involved in cisplatin resistance, recent evidence has suggested that X-linked inhibitor of apoptosis protein (XIAP) may be a key determinant in chemosensitivity in ovarian cancer. Cell fate is determined by a balance between cell survival and apoptotic signaling. Whereas phosphatidylinositol 3-kinase (PI 3-K) and XIAP are believed to be important cell survival factors in human ovarian surface epithelial cancer cells, if and how they interact to confer resistance to chemotherapy is not known. In the present study, we have investigated the role of XIAP in the regulation of the PI 3-K/Akt survival pathway in chemosensitive (A2780-s, OV2008, and OVCAR-3) and resistant (A2780-cp) ovarian cancer cell lines and the nature of this interaction in cell death/survival signaling. Cisplatin decreased XIAP protein levels and induced Akt cleavage and apoptosis in chemosensitive, but not in resistant, ovarian cancer cells. Cisplatin also induced cleavage of caspase-9 and caspase-3, a process blocked by XIAP overexpression. Pretreatment of ovarian cancer cells and their whole cell lysate with tetrapeptide inhibitors of caspases in vitro significantly decreased Akt cleavage induced by cisplatin and exogenous active caspase-3. Adenoviral sense XIAP cDNA expression increased XIAP protein levels and increased Akt phosphorylation, indicative of activation of Akt and, likely, of PI 3-K. This was associated with a decrease in cisplatin-induced apoptosis. In a cell line (OVCAR-3) where basal phosphorylated Akt levels were high, XIAP overexpression failed to increase further the level of this phosphoprotein. XIAP down-regulation induced Akt cleavage and apoptosis, and treatment of whole cell lysate with human recombinant active caspase-3 resulted in a similar pattern of Akt cleavage. In the presence of the PI 3-K inhibitor (LY294002), XIAP overexpression failed to block cisplatin-induced apoptosis and to induce Akt phosphorylation, suggesting that the site of action of XIAP is upstream of Akt in this cell survival pathway. Taken together, the results indicate that XIAP prevents apoptosis through a PI 3 K-dependent inhibition of the caspase cascade. These results demonstrate a novel mechanism by which XIAP regulates apoptosis and the possible involvement of the PI 3-K/Akt survival pathway in XIAP-mediated chemoresistance of ovarian cancer cells. PMID- 11280738 TI - Phosphatidylinositol 3-kinase activity in epidermal growth factor-stimulated matrix metalloproteinase-9 production and cell surface association. AB - Activation of the epidermal growth factor (EGF) receptor regulates many processes associated with metastasis, including modulation of cell:cell and cell:substrate interactions, production of matrix-degrading proteinases, and cellular migration. We have demonstrated previously that EGF stimulates migration and matrix metalloproteinase (MMP)-9-dependent invasion of ovarian cancer cells. In this study, we compare the roles of EGF-induced phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) activities in regulation of cellular responses associated with ovarian tumor cell metastasis. Inhibition of PI3K and MAPK activity impairs EGF-stimulated cell migration, in vitro invasion, and MMP-9 production. PI3K activity is not required for growth factor disruption of cell:cell junctions, whereas inhibitors of extracellular signal-regulated kinase (ERK)1/ERK2 activation and p38 MAPK activity block EGF-dependent junction dissolution. EGF promotes pro-MMP-9 binding to the cell surface through a mechanism that is independent of extracellular enzyme concentration. Interestingly, inhibition of PI3K activity abolishes EGF-induced cell surface association of pro-MMP-9, whereas inhibitors of MAPKs only partially block the response. These data suggest that EGF receptor activation promotes a PI3K dependent induction of a cell surface pro-MMP-9 binding component that may facilitate gelatinase-mediated cellular invasion and supports an expanded role for elevated PI3K activity in cellular responses associated with ovarian tumor metastasis. In addition, our findings support the hypothesis that divergent kinase activities regulate distinct cellular events associated with growth factor induced invasion of ovarian cancer cells. PMID- 11280740 TI - Rapid release of intracellular galectin-3 from breast carcinoma cells by fetuin. AB - Galectin-3, a beta-galactoside binding protein, plays a significant role in cell to extracellular matrix interactions. Despite its extracellular expression, the precise physiological mechanisms that trigger its release from the intracellular milieu have not been characterized. The present analyses were, therefore, done to identify the extracellular matrix proteins with propensity to induce the release of intracellular galectin-3 from breast carcinoma cells. Our studies demonstrate that fetuin, a serum glycoprotein that is abundant in the fetal serum, is capable of inducing the rapid release (approximately 1 min) of intracellular galectin-3 from the cells. The mechanism by which galectin-3 is rapidly released appears to be novel and does not depend on changes in intracellular calcium levels. We also report that galectin-3-expressing breast carcinoma cells in serumless medium adhere and spread well on microtiter wells in the presence of fetuin and divalent ions in a carbohydrate-dependent manner. The data suggest that fetuin is a natural modulator of galectin-3 secretion/release and that the secreted galectin 3 modulates the activity of cell surface receptors for extracellular matrix proteins. PMID- 11280741 TI - Sequential analysis of morphological and biological properties of beta-catenin accumulated crypts, provable premalignant lesions independent of aberrant crypt foci in rat colon carcinogenesis. AB - Our previous study (Cancer Res., 60: 3323-3327, 2000) showed that frequent beta catenin gene mutations are present in beta-catenin-accumulated crypts, which occur early in rodent colonic carcinogenesis, with a lack of the appearance of aberrant crypt foci (ACF). To clarify the nature of such lesions, we performed a sequential analysis of the morphological and biological properties of beta catenin-accumulated crypts. Azoxymethane was administered s.c. to male F344 rats (15 mg/kg body weight) once a week for 3 weeks, and the animals were sacrificed at 5, 10, and 20 weeks after the carcinogen treatment. Both the number of crypts/lesion and the diameter of beta-catenin-accumulated crypts were significantly increased with time courses of 5, 10, and 20 weeks from carcinogen exposure (P < 0.01). Likewise, the histological abnormality in those crypts, assessed by semiquantitative analyses, was also increased with time (P < 0.01). Conversely, ACF did not show any increase in histological abnormality during the time course and maintained a monotonous histology throughout the experiment. The histological abnormality score for beta-catenin-accumulated crypts was significantly higher than for ACF at every time point (P < 0.001). The number of AgNOR/nucleus in beta-catenin-accumulated crypts was significantly higher than in ACF (P < 0.001). Beta-catenin-accumulated crypts were accompanied frequently by Paneth cells and had decreased hexosaminidase activity. Such data, together with the results in our previous report, strongly suggest that beta-catenin accumulated crypts, which are independent of ACF, are truly premalignant lesions for colon cancer. PMID- 11280742 TI - A clonal growth model: time-course simulations of liver foci growth following penta- or hexachlorobenzene treatment in a medium-term bioassay. AB - A combination of experimental and simulation approaches were used to analyze the clonal growth of preneoplastic, enzyme-altered foci during liver carcinogenesis in an initiation-promotion regimen. Male Fisher 344 rats, 8 weeks of age, were initiated with a single dose (200 mg/kg, i.p.) of diethylnitrosamine (DEN). Beginning 2 weeks later, animals were exposed to daily gavage consisting of 0.1 mmol/kg pentachlorobenzene (PECB) or hexachlorobenzene (HCB) in corn oil vehicle for 6 weeks. Partial hepatectomy was performed 3 weeks after initiation. Experimental data including liver weight, hepatocyte density (number of hepatocytes/unit volume), 5-bromo-2'-deoxyuridine-labeling index for analysis of cell division rate, and number and volume of glutathione-S-transferase pi positive foci were collected 23, 26, 28, 47, or 56 days after initiation. Model parameters describing liver growth were obtained directly from the experimental data. The probability of mutation/division of normal cells and the growth rate of initiated cells were inferred by a comparison of model outcomes with the observed time courses of foci development. To describe the time-dependent increases in foci volume and the concomitant reduction of foci number observed in all treatment groups, the calibrated model for the DEN controls incorporated the hypothesis of two initiated cell populations (referred to as A and B cells) within the framework of the two-stage model. The B cells are initiated cells that have a selective growth advantage under conditions that inhibit the growth of A cells and normal hepatocytes. The parameter values defined in the DEN controls were used to evaluate experiments involving the administration of PECB or HCB. Both PECB and HCB caused a significant increase in foci volume compared with the DEN controls. HCB treatments resulted in increased proliferation of normal hepatocytes, which was not observed for PECB under the same treatment regimen. The best description of the data resulted from the model incorporating the hypothesis that PECB and HCB promoted the growth of foci via increased net growth rates of B cells. We present here a biologically based clonal growth simulation platform to describe the growth of preneoplastic foci under experimental manipulations of initiation-promotion studies. This simulation work is an example of quantitative approaches that could be useful for the analysis of other initiation-promotion studies. PMID- 11280743 TI - ErbB2 overexpression on occult metastatic cells in bone marrow predicts poor clinical outcome of stage I-III breast cancer patients. AB - Occult hematogenous micrometastases are the major cause for metastatic relapse and cancer-related death in patients with operable primary breast cancer. Although sensitive immunocytochemical and molecular methods allow detection of individual breast cancer cells in bone marrow (BM), a major site of metastatic relapse, current detection techniques cannot discriminate between nonviable shed tumor cells and seminal metastatic cells. To address this problem, we analyzed the relevance of erbB2 overexpression on disseminated cytokeratin-18-positive breast cancer cells in the BM of 52 patients with locoregionally restricted primary breast cancer using immunocytochemical double labeling with monoclonal antibody 9G6 to the p185erbB2 oncoprotein. Expression of p185erbB2 on BM micrometastases was detected in 31 of 52 (60%) patients independent of established risk factors such as lymph node involvement, primary tumor size, differentiation grade, or expression of p185erbB2 on primary tumor cells. After a median follow-up of 64 months, patients with p185erbB2-positive BM micrometastases had developed fatal metastatic relapses more frequently than patients with p185erbB2-negative micrometastases (21 versus 7 events; P = 0.032). In multivariate analysis, the presence of p185erbB2-positive micrometastases was an independent prognostic factor with a hazard ratio of 2.78 (95% confidence interval, 1.11-6.96) for overall survival (P = 0.029). We therefore conclude that erbB2 overexpression characterizes a clinically relevant subset of breast cancer micrometastases. PMID- 11280744 TI - Heterogeneous nuclear ribonucleoprotein B1 as early cancer biomarker for occult cancer of human lungs and bronchial dysplasia. AB - Heterogeneous nuclear ribonucleoprotein (hnRNP) B1 is a RNA-binding protein of Mr 37,000. We previously reported that hnRNP B1 was specifically overexpressed in the nuclei of human lung cancer cells, particularly in squamous cell carcinoma (E. Sueoka et al., Cancer Res., 59: 1404-1407, 1999). We extended this study to determine whether hnRNP BL was overexpressed in roentgenographically occult cancers of the lungs and premalignant lesions of squamous cell carcinomas, such as bronchial dysplasia. The additional object of our study was to examine the usefulness of hnRNP B1 as a potential diagnostic marker for squamous cell carcinoma of various organs, such as the oral cavity and esophagus in humans. Surgically resected specimens of bronchial dysplasia, lung cancers, and various human squamous cell carcinomas, collected at two hospitals in Japan, were subjected to immunohistochemical staining with anti-hnRNP B1 antibody. Overexpression of hnRNP B1 protein was observed in 100% of stage I lung cancer tissues, but it was not found in normal bronchial epithelium. Squamous cell carcinoma of the lungs showed stronger staining than other histological types, and elevation of hnRNP B1 was found in both roentgenographically occult lung cancers and bronchial dysplasia. Furthermore, cytological examination with anti hnRNP B1 antibody detected cancer cells in sputum, suggesting the potential of hnRNP B1 protein as a new biomarker for the very early stage of lung cancer in humans. Because strong staining of hnRNP B1 was also observed in various squamous cell carcinomas of oral and esophageal tissues as shown in our recent reports, overexpression of hnRNP B1 seems to be a common event in the carcinogenic processes of squamous cell carcinoma. These results suggest that hnRNP B1 protein could be a useful diagnostic biomarker for both the very early stages of lung cancer and various squamous cell carcinomas in humans. PMID- 11280745 TI - Key importance of the Helicobacter pylori adherence factor blood group antigen binding adhesin during chronic gastric inflammation. AB - Helicobacter pylori has been assigned as a class I carcinogen because of its relation to gastric adenocarcinoma. Chronic H. pylori infection may lead to severe gastritis, glandular atrophy (AT), and intestinal metaplasia (IM). Strains secreting the vacuolating toxin VacA and producing the cytotoxin-associated antigen CagA (type 1 strains), as well as the blood group antigen binding adhesin (BabA) targeting Lewis(b) antigens, have been associated previously with distal gastric adenocarcinoma (M. Gerhard et al., Proc. Natl. Acad. Sci. USA, 96: 12778 12783, 1999) and may therefore also be related to lesions preceding gastric cancer. Antral and corpus biopsies were collected from 451 patients; 151 were H. pylori positive, as determined by PCR. Gastric biopsies were histologically evaluated for activity of gastritis (G0-G3, granulocyte infiltration), chronicity of gastritis (L1-L3, lymphocyte infiltration), and the presence of IM and/or AT according to the Sydney classification. Simultaneously, the presence of bacterial genes encoding virulence and adherence factors (racAs1/s2, cagA, and babA2) was determined by PCR. The presence of cagA+ and vacAs1 (alone or combined) both correlated with activity and chronicity of gastritis (P < 0.05); however, the overall prevalence of these genes was 60 or 72%, respectively, and was thus relatively frequent. The babA2 gene, encoding the adhesin BabA, was detected in 38% of infected patients and was correlated with the activity of gastritis in antrum and corpus (P < 0.005). cagA+/vacAs1+ strains (suggesting the presence of type 1 strains) that were also babA2 positive were detected more frequently in patients with severe histological alterations (such as G3, IM, or AT) compared with subjects without these changes (P < 0.01). cagA+/vacAs1+ strains that were babA2 negative, however, lacked a significant correlation with severe histological changes, activity, or chronicity of gastritis in antrum and corpus. Adherence of H. pylori via BabA appears to be of importance for efficient delivery of VacA and CagA and may play a special role in the pathogenesis of severe histological changes. PMID- 11280746 TI - Overexpression of aromatase leads to hyperplasia and changes in the expression of genes involved in apoptosis, cell cycle, growth, and tumor suppressor functions in the mammary glands of transgenic mice. AB - Our previous studies have shown that overexpression of aromatase results in increased tissue estrogenic activity and induction of hyperplastic and dysplastic lesions in aromatase transgenic mammary glands. In this study, we have examined the effects of aromatase overexpression on biochemical changes in the aromatase transgenic mice. Our results show an increase in the expression of both estrogen and progesterone receptors, and their expression is maintained in the transgenic mammary tissue even without circulating ovarian estrogens. Our results also show an increase in the expression of several growth factors and cell cycle genes in the aromatase transgenic mammary glands, which is consistent with the observed increase in proliferating cell nuclear antigen levels and cellular proliferation. Interestingly, we have also observed a decrease in the expression of epidermal growth factor receptor and its ligands, epidermal growth factor and transforming growth factor alpha, as well as several tumor suppressor genes such as p53 and retinoblastoma. This study presents novel and interesting findings that are consistent with the current models of aromatase influence and the complex interactions of biochemical pathways leading to mammary tumorigenesis. PMID- 11280747 TI - Evaluation of androgen, estrogen (ER alpha and ER beta), and progesterone receptor expression in human prostate cancer by real-time quantitative reverse transcription-polymerase chain reaction assays. AB - Steroid hormones can have profound effects on prostate tumor development making it important to define steroid receptor expression in prostate tissues. For this purpose, androgen receptor (AR) and estrogen receptor (ER alpha and ER beta) expression was quantified in 12 clinically localized and 11 hormone-refractory sporadic prostate tumors, using real-time quantitative reverse transcription-PCR assays. To gain more insight into hormone-responsiveness, estrogen-regulated progesterone receptor (PGR) and androgen-regulated prostatic acid phosphatase (PAP) mRNA levels were also quantified. There is a decrease in expression of ER beta in both clinically localized and hormone-refractory tumors relative to normal prostate tissues. Moreover, hormone-refractory tumors display a decreased expression of ER alpha and an increased expression of AR. There is a positive association between ER alpha, ER beta, and PGR expression (P < 0.0001) and a negative association between AR and the androgen-regulated gene PAP expression in hormone-refractory tumors. Taken together, these data indicate that, although increased expression of the AR gene might play a key role in endocrine treatment failure, it cannot be considered as the sole actor of this unresolved dilemma, and abnormalities in ER alpha and/or ER beta expression may also modulate the growth response of prostate cancer to hormone withdrawal. Our results also suggest that ER alpha and ER beta expression status could be used to identify advanced prostate tumor patients who may respond to antiestrogen therapy. PMID- 11280748 TI - Modulation of experimental colon tumorigenesis by types and amounts of dietary fatty acids. AB - Epidemiological studies and laboratory animal model assays suggest that a high intake of dietary fat promotes colorectal cancer. Several in vivo and in vitro studies support the hypothesis that omega-6 fatty acids promote colon tumorigenesis, whereas omega-3 fatty acids lack promoting activity. Fat intake in the United States traditionally includes high amounts (30% of total caloric intake) of saturated fat rather than omega-6 fatty acids. Therefore, the present study was designed to compare the modulatory effects of a high-fat diet containing mixed lipids (HFML), a diet rich in saturated fatty acids (the average American diet), a diet with fish oil (HFFO) that is rich in omega-3 fatty acids, and a low-fat corn oil diet (LFCO) on the formation of chemically induced colonic aberrant crypt foci (ACF) and tumors, cyclooxygenase (COX)-2 activity, and apoptosis during experimental colon carcinogenesis. At 5 weeks of age, groups of male F344 rats were fed a 5% corn oil diet (LFCO). At 7 weeks of age, rats intended for carcinogen treatment received s.c. injections of azoxymethane at a dose level of 15 mg/kg of body weight once weekly for 2 weeks. Beginning 1 day after the carcinogen treatment, groups of rats were then maintained on experimental diets containing 20% HFML or 20% HFFO. Rats were killed at 8, 23, or 38 weeks after azoxymethane treatment. Colonic ACF and tumors were evaluated histopathologically, and apoptosis was evaluated by the terminal deoxynucleotidyl transferase-mediated nick end labeling method. Colonic mucosae and tumor samples harvested at week 38 were analyzed for COX-2 synthetic activity and expression. The rats fed the HFML diet showed significantly increased total colonic ACF (P < 0.001-0.0001) with a multiplicity of > or = 4 aberrant crypts/focus (P < 0.0001) compared with the effects of the HFFO or LFCO diets at week 8, 23, and 38. Interestingly, there was a 2- to 3-fold increase (> or = 4) in multicrypt foci in rats given the HFML diet as compared with such foci in rats fed the HFFO or LFCO diets. By week 23, the HFML diet had significantly increased the incidence of colonic tumors (30-60%) and their multiplicity (100-141%) when compared with the effects of the LFCO or HFFO diets. At week 38, the HFML diet had induced 100% colon tumor incidence and a 4-fold multiplicity of adenocarcinomas compared with the LFCO and HFFO diets. At weeks 23 and 38, a significantly lower percentage of apoptotic colonic epithelial cells were observed in the tumors of animals fed the HFML diet as compared with those fed the HFFO diet. The HFML diet caused significantly increased levels of COX-2 activity in colon tumors (P < 0.05-0.01), and these tumors had enhanced levels of COX-2 expression as compared with those in assays with LFCO or HFFO diets. These observations demonstrate for the first time that HFML diets containing high levels of saturated fatty acids (such as those in Western diets) promote colon carcinogenesis. Although the mechanisms involved in colon tumor promotion by a HFML diet are not fully known, our results indicate that the modulation of eicosanoid production via the influence on COX activity and the suppression of apoptosis may play a key role in HFML diet induced colon tumorigenesis. PMID- 11280750 TI - Localization of a human reduced folate carrier protein in the mitochondrial as well as the cell membrane of leukemia cells. AB - IgG polyclonal antiserum was generated in New Zealand White rabbits immunized with a 16-mer peptide consisting of a specific amino acid sequence at residues corresponding to the sixth to seventh predicted transmembrane domain of the human reduced folate carrier (RFC). Using Western immunoblotting to examine the cytosolic and membrane fractions of the human CCRF-CEM T-cell lymphoblastic leukemia cell line, polyclonal antihuman RFC antiserum recognized two bands in the cytosolic fraction (approximately 60 kDa and approximately 70 kDa) on 10% polyacrylamide gels. In the membrane fraction, an approximately 60-kDa protein was identified. Comparative studies of a panel of human tumor cell lines including the HT1080 fibrosarcoma, 8805 malignant fibrous histiocytoma, and the MCF breast cancer cell lines revealed similar findings. Likewise, a recombinant approximately 60-kDa membrane protein was identified after expression of baculovirus-infected Sf9 insect cells containing cDNA of the human RFC. In the CEM-7A cell line, a variant of the CCRF-CEM cell line that overexpresses the RFC, 21-fold overexpression of the approximately 60-kDa membrane protein (RFC) was shown by Western analysis. To characterize further the cellular distribution of the human RFC, immunohistochemical analyses were performed in CCRF-CEM T-cell lymphoblastic leukemia cells. Predominantly membrane localization of the antibody reacting sites was detected; however, a cytoplasmic component was noted as well. By confocal microscopy and by immunogold electron microscopy, the cytoplasmic expression was found to be largely of mitochondrial origin. These findings were corroborated by Western immunoblotting of mitochondrial membrane isolates from the CCRF-CEM cell line, which demonstrate an approximately 60-kDa protein. The localization of the human RFC to the mitochondrial membrane is a novel finding, and it suggests a role for the mitochondrial membrane in the transport of folates. PMID- 11280749 TI - Human papillomavirus 16 and 18 L1 serology compared across anogenital cancer sites. AB - Human papillomavirus (HPV) DNA has been detected in the great majority of cancers of the uterine cervix and anus, whereas the association of HPV DNA with cancer at other anogenital sites has produced less consistent results. This study was designed to compare HPV exposure among anogenital cancer cases and matched controls. Cases (1782) of anogenital cancer diagnosed in the Seattle area from 1978 to 1998 were identified and interviewed. Their responses were compared with those of 2383 age- and sex-matched controls. Blood was drawn at interview from both cases and controls and tested for antibodies to HPV-16 and HPV-18. Tissue blocks were tested for HPV DNA for 649 cases. Serum antibodies to HPV-16 were associated with in situ and invasive cancer at all sites among men and women with the exception of in situ penile cancer. Anti-HPV-18 antibodies were associated with cancers at all sites among women. The increased risk of cancer associated with HPV-16 seropositivity ranged from odds ratio = 1.8 (95% confidence interval, 1.4-2.5) for adenocarcinoma of the cervix to odds ratio = 5.9 (95% confidence interval, 3.4-10.3) for anal cancer in men. Associations between seroprevalence and cancers were stronger when analyses were restricted to HPV-16- or HPV-18 DNA positive cases. HPV DNA was detected in >80% of cancers from all sites tested. HPV-16 DNA was the type most frequently detected at all sites (range, 40.9 82.2%). HPV-18 DNA was detected in 44.7% of adenocarcinomas of the cervix but detected much less often (2.6-18.1%) at other sites. These findings support an important role for HPV infection in anogenital cancer at all sites. Differences in the proportion of seropositives among HPV-16 DNA-positive cases by site suggest either that the immune response varies by site or that cancer development may lead to changes in antibody responses in a site-specific fashion. PMID- 11280751 TI - Vascular attack by 5,6-dimethylxanthenone-4-acetic acid combined with B7.1 (CD80) mediated immunotherapy overcomes immune resistance and leads to the eradication of large tumors and multiple tumor foci. AB - The promise of cancer immunotherapy is that it will not only eradicate primary tumors but will generate systemic antitumor immunity capable of destroying distant metastases. A major problem that must first be surmounted relates to the immune resistance of large tumors. Here we reveal that immune resistance can be overcome by combining immunotherapy with a concerted attack on the tumor vasculature. The functionally related antitumor drugs 5,6-dimethylxanthenone-4 acetic acid (DMXAA) and flavone acetic acid (FAA), which cause tumor vasculature collapse and tumor necrosis, were used to attack the tumor vasculature, whereas the T-cell costimulator B7.1 (CD80), which costimulates T-cell proliferation via the CD28 pathway, was used to stimulate antitumor immunity. The injection of cDNA (60-180 microg) encoding B7.1 into large EL-4 tumors (0.8 cm in diameter) established in C57BL/6 mice, followed 24 h later by i.p. administration of either DMXAA (25 mg/kg) or FAA (300 mg/kg), resulted in complete tumor eradication within 2-6 weeks. In contrast, monotherapies were ineffective. Both vascular attack and B7.1 immunotherapy led to up-regulation of heat shock protein 70 on stressed and dying tumor cells, potentially augmenting immunotherapy. Remarkably, large tumors took on the appearance of a wound that rapidly ameliorated, leaving perfectly healed skin. Combined therapy was mediated by CD8+ T cells and natural killer cells, accompanied by heightened and prolonged antitumor cytolytic activity (P < 0.001), and by a marked increase in tumor cell apoptosis. Cured animals completely rejected a challenge of 1 x 10(7) parental EL-4 tumor cells but not a challenge of 1 x 10(4) Lewis lung carcinoma cells, demonstrating that antitumor immunity was tumor specific. Adoptive transfer of 2 x 10(8) splenocytes from treated mice into recipients bearing established (0.8 cm in diameter) tumors resulted in rapid and complete tumor rejection within 3 weeks. Although DMXAA and B7.1 monotherapies are complicated by a narrow range of effective doses, combined therapy was less dosage dependent. Thus, a broad range of amounts of B7.1 cDNA were effective in combination with 25 mg/kg DMXAA. In contrast, DMXAA, which has a very narrow range of high active doses, was effective at a low dose (18 mg/kg) when administered with a large amount (180 microg) of B7.1 cDNA. Importantly, combinational therapy generated heightened antitumor immunity, such that gene transfer of B7.1 into one tumor, followed by systemic DMXAA treatment, led to the complete rejection of multiple untreated tumor nodules established in the opposing flank. These findings have important implications for the future direction and utility of cancer immunotherapies aimed at harnessing patients' immune responses to their own tumors. PMID- 11280752 TI - p53 effects both the duration of G2/M arrest and the fate of temozolomide-treated human glioblastoma cells. AB - Temozolomide (TMZ) is a DNA-methylating agent that has recently been introduced into Phase II and III trials for the treatment of gliomas. TMZ produces O6 methylguanine in DNA, which mispairs with thymine during the next cycle of DNA replication. Subsequent futile cycles of DNA mismatch repair can lead to a p53 associated apoptotic cell death, although this mechanism has been described mostly in hematopoietic neoplasms. We studied the action of TMZ in gliomas and the role p53 might play by using U87 glioma cells that were either p53-wild-type or p53-deficient (by virtue of expression of the viral oncoprotein E6). LN-Z308 cells, in which p53 gene is deleted, were also used. p53-proficient U87 MG cells underwent a prolonged, p53- and p21(Waf1/Cip1)-associated G2-M arrest beginning 2 days after TMZ treatment. Although very few of these cells underwent apoptosis, most underwent senescence over a 10-day period. p53-deficient (E6-transfected U87 and LN-Z308) cells similarly underwent G2-M arrest in response to TMZ, but this arrest was accompanied by only minor changes in p53 or p21(Waf1/Cip1) and was reversed within 7 days of TMZ treatment in association with the appearance of cells with either 8n or subG1 DNA content. These results suggest that glioma cells respond to TMZ by undergoing G2-M arrest. p53 is not necessary for this G2 M arrest to occur but is important in the duration of G2-M arrest and in the ultimate fate of TMZ-treated cells. Therefore, the integrity of the G2-M cell cycle checkpoint may be important in the cytotoxicity of TMZ in glioma cells. PMID- 11280753 TI - Characterization of a novel topoisomerase I mutation from a camptothecin resistant human prostate cancer cell line. AB - In this study, we characterized the structure and function of topoisomerase I (top1) protein in the camptothecin (CPT)-resistant prostate cancer cell lines, DU 145/RC0.1 and DU-145/RC1 (RC0.1 and RC1, respectively). Both of the cell lines were previously selected by continuous exposure to 9-nitro-CPT. The RC0.1 and RC1 cells have high cross-resistance to CPT derivatives including SN-38 and topotecan, but are not cross-resistant to the non-top1 inhibitors etoposide, doxorubicin, and vincristine. Although the top1 protein levels were not decreased in the resistant cells compared with the parental cells, CPT-induced DNA cleavage was markedly reduced in the RC0.1 and RC1 nuclear extracts. The resistant-cell line nuclear extracts also demonstrated top1 catalytic activity and resistance to CPT, in in vitro assays. Reverse transcription-PCR products from the resistant cell lines were sequenced, and revealed a point mutation resulting in a R364H mutation in the top1 of both RC0.1 and RC1. No wild-type top1 RNA or genomic DNA was detected in the resistant cell lines. Using a purified recombinant R364H top1, we found that the R364H mutant top1 was CPT resistant and fully active. In the published top1 crystal structure, the R364H mutation is close to the catalytic tyrosine and other well-known mutations leading to CPT resistance. PMID- 11280754 TI - Targeting of aluminum (III) phthalocyanine tetrasulfonate by use of internalizing monoclonal antibodies: improved efficacy in photodynamic therapy. AB - The use of monoclonal antibodies (MAbs) directed against tumor-associated antigens for targeting of photosensitizers is an interesting option to improve the selectivity of photodynamic therapy (PDT). Hydrophilic photosensitizers are most suitable for conjugation to MAbs because of their water solubility. The photosensitizer aluminum (III) phthalocyanine tetrasulfonate [AlPc(SO3H)4] has many ideal photochemical properties; however, because of its hydrophilicity, the free form of this sensitizer does not readily reach the critical intracellular target and, therefore, is ineffective in PDT. On the basis of our previous studies, we hypothesized that AlPc(SO3H)4 might be suitable for PDT when coupled to internalizing tumor-selective MAbs. In this study, a reproducible procedure is presented for coupling of AlPc(SO3H)4 to MAbs via the tetra-glycine derivative AlPc(SO2Ngly)4. Conjugation was performed to chimeric MAb (cMAb) U36 and murine MAbs (mMAb) E48 and 425 using a labile ester. Conjugates showed preservation of integrity and immunoreactivity and full stability in serum in vitro. At molar ratios >4, the solubility of the conjugates decreased. Data on the in vitro efficacy of PDT showed that in the chosen experimental setup the internalizing AlPc(SO2Ngly)4-mMAb 425 conjugate was about 7500 times more toxic to A431 cells than the free sensitizer (IC50s, 0.12 nM versus 900 nM). The AlPc(SO2Ngly)4-mMAb 425 conjugate was also more toxic than meta-tetrahydroxyphenylchlorin-mMAb 425 conjugates and free meta-tetrahydroxyphenylchlorin that had been tested previously (M. B. Vrouenraets et al., Cancer Res., 59: 1505-1513, 1999) in the same system (IC50s, 7.3 nm and 2.0 nM, respectively). Biodistribution analysis of AlPc(SO2Ngly)4-125I-labeled cMAb U36 conjugates with different sensitizer:MAb ratios in squamous cell carcinoma-bearing nude mice revealed selective accumulation in the tumor, although to a lesser extent than for the unconjugated 125I-labeled cMAb U36, whereas tumor:blood ratios were similar. These findings indicate that AlPc(SO3H)4 has high potential for use in PDT when coupled to internalizing tumor-selective MAbs. PMID- 11280755 TI - T-cell activation by recombinant receptors: CD28 costimulation is required for interleukin 2 secretion and receptor-mediated T-cell proliferation but does not affect receptor-mediated target cell lysis. AB - Recombinant T-cell receptors with antibody-like specificity are successfully used to direct CTLs toward a MHC-independent immune response against target cells. Here we monitored the specific activation of receptor grafted CTLs in the context of CD28 costimulation. Peripheral blood T cells were retrovirally engrafted with recombinant anti-CD30 and anti-carcinoembryonic antigen receptors, respectively, that harbor either the Fc epsilonRI-gamma or the CD3-zeta intracellular signaling domain. Cross-linking of recombinant receptors by solid-phase bound ligand, i.e., CD30 and a carcinoembryonic antigen receptor-specific anti-idiotypic antibody, respectively, induces IFN-gamma secretion that is further enhanced by CD28 costimulation of grafted T cells. Induction of interleukin (IL)-2 secretion, in contrast, requires CD28 costimulation in addition to receptor cross-linking, irrespective of T-cell preactivation by anti-CD3 monoclonal antibody plus IL-2 or by anti-CD3 monoclonal antibody plus anti-CD28 monoclonal antibody. Accordingly, induction of IL-2 secretion upon receptor cross-linking by membrane-bound antigen requires CD28/B7 costimulation whereas IFN-gamma secretion and cell proliferation does not. The efficiency of cytolysis by receptor-grafted CTLs does not depend on and is not affected by CD28 costimulation. The data demonstrate that CTL proliferation, cytokine secretion, and cytolysis upon receptor cross-linking are differentially modulated by CD28 costimulation and that cytolysis does not require B7 expression on target cells. PMID- 11280756 TI - Pharmacological and toxicological aspects of 4-demethoxy-3'-deamino-3'-aziridinyl 4'-methylsulphonyl-daunorubicin (PNU-159548): a novel antineoplastic agent. AB - 4-demethoxy-3'-deamino-3'-aziridinyl-4'-methylsulphonyl-daunorubicin (PNU-159548) belongs to a novel class of antitumor compounds (termed alkycyclines) and is currently undergoing Phase II clinical trial. In the present study, we investigated the in vitro and in vivo antitumor activity, the pharmacokinetics, and the toxicological profile of this compound. PNU-159548 showed good cytotoxic activity in murine and human cancer cells growing in vitro, with an average concentration for 50% growth inhibition of 15.8 ng/ml. The drug showed strong antitumor efficacy in vivo after i.v. and p.o. administration against rapidly proliferating murine leukemias and slowly growing transplantable human xenografts. At non-toxic doses, PNU-159548 produced complete regression and cures in ovarian, breast, and human small cell lung carcinomas. Fourteen of 16 models studied, including colon, pancreatic, gastric, and renal carcinomas, astrocytoma and melanoma, were found to be sensitive to PNU-159548. In addition, PNU-159548 was effective against intracranially implanted tumors. Toxicological studies revealed myelosuppression as the main toxicity in both mice and dogs. The maximum tolerated doses, after a single administration, were 2.5 mg/kg of body weight in mice, 1.6 mg/kg in rats, and 0.3 mg/kg in dogs. In the cyclic studies, the maximum tolerated doses were 0.18 mg/kg/day (cumulative dose/cycle: 0.54 mg/kg) in rats and 0.05 mg/kg/day (cumulative dose/cycle: 0.15 mg/kg) in dogs. PNU 159548 showed minimal cardiotoxicity, when compared with doxorubicin in the chronic rat model at a dose level inducing similar myelotoxicity. Animal pharmacokinetics, carried out in mice, rats, and dogs, was characterized by high volumes of distribution, plasma clearance of the same order of the hepatic blood flow, and short terminal half-life. These findings support the conclusion that PNU-159548 is an excellent candidate for clinical trials in the treatment of cancer. PMID- 11280757 TI - 4-Demethoxy-3'-deamino-3'-aziridinyl-4'-methylsulphonyl-daunorubicin (PNU 159548), a novel anticancer agent active against tumor cell lines with different resistance mechanisms. AB - The activity of 4-demethoxy-3'-deamino-3'-aziridinyl-4'-methylsulphonyl daunorubicin (PNU-159548), a new alkycycline with high antitumor activity against a broad range of cancer cells, was evaluated in vitro and in vivo in cells selected for resistance to different anticancer agents. Both in vitro and in vivo, PNU-159548 did retain its activity in cells expressing the multidrug resistance (MDR) phenotype, associated to MIDR-1 gene overexpression or with an alteration in the topoisomerase II gene (altered MDR), independently on the drug used for the selection of the resistant cell line. According to these data, the intracellular uptake of PNU-159548 is not influenced by the presence of MDR-1. PNU-159548 was also active, both in vitro and in vivo, against cells showing resistance to various alkylating agents iincluding cisplatin, cyclophosphamide, and melphalan) and topoisomerase I-inhibitors. Cells defective in nucleotide excision repair, which did show hypersensitivity to treatment with UV irradiation and alkylating agents, showed only a marginally increased sensitivity to PNU 159548. Similarly, the activity of the drug was not influenced by the mismatch repair system, as assessed in two different cellular systems deficient in hMLH1 expression and in which hMLH1 activity was restored by chromosome 3 transfer. The results obtained clearly indicate that the new anticancer agent PNU-159548 is able to overcome the classical mechanisms of resistance emerging after treatment with the most clinically used anticancer agents, and it could represent an alternate choice in the treatment of those tumors refractory to conventional therapy. PMID- 11280758 TI - Experimental chronotherapy of mouse mammary adenocarcinoma MA13/C with docetaxel and doxorubicin as single agents and in combination. AB - The therapeutic index of docetaxel, doxorubicin and their combination may be improved by an adequate selection of the circadian time of administration. The present study constitutes a prerequisite to testing the clinical relevance of chronotherapy in human breast cancer. Three experiments were performed in C3H/HeN mice. Each treatment modality was administered i.v. once a week for 3 weeks at one of six circadian stages, during the light span, when the mice were resting: 3, 7, and 11 h after light onset (HALO), or during darkness, when the mice were active: 15, 19, and 23 HALO. The circadian time dependency of single agent tolerability was investigated in healthy mice using four dose levels for docetaxel (38.8, 23.3, 14, and 8.4 mg/kg/injection) and for doxorubicin (13.8, 8.3, 5 and 3 mg/kg/injection; experiment 1). The circadian time dependency of each single agent efficacy was studied in MA13/C-bearing mice, using two dose levels of docetaxel (38.8 or 23.3 mg/kg/injection) or doxorubicin (8.3 or 5 mg/kg/injection; experiment 2). The toxicity and the efficacy of the simultaneous docetaxel-doxorubicin combination were assessed as a function of dosing time in experiment 3. Two combinations were tested (A, 16.3 mg/kg/injection of docetaxel and 2.5 mg/kg/injection of doxorubicin; and B, 11.6 and 3.5 mg/kg/injection, respectively) at each of the above six circadian times. Mortality, body weight change, and tumor size were recorded for 60-70 days in each experiment. Single agent docetaxel or doxorubicin was significantly best tolerated near the middle of the rest span (7 HALO) and most toxic in the middle of the activity phase (19 HALO). Docetaxel or doxorubicin as a single drug were also most effective at 7 HALO, irrespective of dose. Treatment at 7 HALO produced highest rates of complete tumor inhibition (81% versus 11% at 3 HALO for docetaxel, p from chi2 <0.001, and 69% versus 44% at 11 HALO for doxorubicin, not significant) and highest day 60 survival rate (100% versus 28% at 3 HALO for docetaxel, p from chi2 <0.001 and 89% versus 69% at 15 HALO for doxorubicin, not significant). Docetaxel-doxorubicin combinations were most effective following dosing in the beginning of the rest span or short after the onset of the activity span, with regard to the rates of both complete tumor inhibitions (45% at 3 HALO versus 15% at 19 HALO) and day 70 survival rates (85% and 80% at 3 and 7 HALO respectively, versus 20% at 19 HALO). The efficacy of single agent docetaxel or doxorubicin and that of their combination varied largely as a function of circadian dosing time. Single agent docetaxel at 7 HALO was the best treatment option in this model with regard to both tolerability and efficacy. This timing may correspond to the middle of the night in cancer patients. PMID- 11280759 TI - Effects of chemotherapy by 1,3-bis(2-chloroethyl)-1-nitrosourea on single-quantum and triple-quantum-filtered 23Na and 31P nuclear magnetic resonance of the subcutaneously implanted 9L glioma. AB - The effects of chemotherapy [25 mg/kg 1,3-bis(2-chloroethyl)-1-nitrosourea administered with a single i.p. injection] on cellular energetics by 31P nuclear magnetic resonance (NMR) spectroscopy, total tissue sodium by single-quantum (SQ) 23Na NMR spectroscopy, and intracellular sodium by triple-quantum-filtered (TQF) 23Na NMR spectroscopy were studied in the s.c. 9L glioma. Animals were studied by NMR 2 days before therapy and 1 and 5 days after therapy. Destructive chemical analysis was also performed 5 days after therapy to validate the origin of changes in SQ and TQF 23Na signals. One day after treatment, there was no significant difference between control and treated tumors in terms of tumor size or 23Na and 31P spectral data. Five days after therapy, treated tumors had 28 +/- 16% (P < 0.1) lower SQ 23Na signal intensity, 46 +/- 20% (P < 0.05) lower TQF 23Na signal intensity, 125 +/- 51% (P < 0.05) higher ATP:Pi ratio, 186 +/- 69% (P < 0.05) higher phosphocreatine:Pi ratio, and 0.17 +/- 0.06 pH units (P < 0.05) higher intracellular pH compared with control tumors. No significant differences in TQF 23Na relaxation times were seen between control and treated tumors at any time point. Destructive chemical analysis showed that the relative extracellular space of control and treated tumors was identical, but the treated tumors had 21 +/- 8% (P < 0.05) lower total tissue Na+ concentration and 60 +/- 24% (P < 0.05) lower intracellular Na+ concentration compared with the controls. The higher phosphocreatine:Pi and ATP:Pi ratios after 1,3-bis(2-chloroethyl)-1-nitrosourea treatment indicate improved bioenergetic status in the surviving tumor cells. The decrease in SQ and multiple-quantum-filtered 23Na signal intensity was largely attributable to a decrease in Na(i)+ because the treatment did not change the relative extracellular space. The improved energy metabolism could decrease the intracellular concentration of Na+ by increasing the activity of Na+-K+-ATPase and decreasing the activity of Na+/H+. Although both 23Na and 31P spectra were consistent with improved cellular metabolism in treated tumors, the 23Na methods may be better suited for monitoring response to therapy because of higher signal:noise ratio and ease of imaging the single 23Na resonance. PMID- 11280760 TI - Response of LNCaP spheroids after treatment with an alpha-particle emitter (213Bi)-labeled anti-prostate-specific membrane antigen antibody (J591). AB - A theoretical drawback to alpha-particle therapy with 213Bi is the short range of the particle track coupled with the short half-life of the radionuclide, thereby potentially limiting effective cytotoxicity to rapidly accessible, disseminated individual tumor cells (e.g., as in leukemia). In this work, a prostate carcinoma spheroid model was used to evaluate the feasibility of targeting micrometastatic clusters of tumor cells using 213Bi-labeled anti-prostate-specific membrane antigen (PSMA) antibody, J591. In prostate cancer, vascular dissemination of tumor cells or tumor cell clusters to the marrow constitutes an important step in the progression of this disease to widespread skeletal involvement, an incurable state. Such prevascularized clusters are ideal targets for radiolabeled antibodies because the barriers to antibody penetration that are associated with the capillary basal lamina have not yet formed. Beta- and gamma-emitting radionuclides such as 131I, which are widely used in radioimmunotherapy, are not expected to be effective when targeting single cells or small cell clusters. This is because the range of the emissions is one to two orders of magnitude greater than the target size, and the energy deposited per traversal is insufficient to produce any significant radiobiological effect. Spheroids of the prostate cancer cell line, LNCaP-LN3, were used as a model of prevascularized micrometastases; their response to an anti-PSMA antibody, J591, radiolabeled with the alpha particle emitter 213Bi (T(1/2), 45.6 min.) has been measured. The time course of spheroid volume reductions was found to be sensitive to the initial spheroid volume. J591 labeled with 0.9 MBq/ml 213Bi resulted in a 3-log reduction in spheroid volume on day 33, relative to control, for spheroids with an initial diameter of 130 microm; 1.8 MBq/ml were required to achieve a similar response for spheroids with an initial diameter of 180 microm. Equivalent spheroid responses were observed after 12 Gy of acute external beam photon irradiation. Monte Carlo-based microdosimetric analyses of the 213Bi decay distribution in individual spheroids of 130-microm diameter yielded an average alpha-particle dose of 3.7 Gy to the spheroids, resulting in a relative biological effectiveness factor of 3.2 over photon irradiation. The activity concentrations used in the experiments were clinically relevant, and this work supports the possibility of using 213Bi-labeled antibodies not only for disseminated single tumor cells, as found in patients with leukemia, but also for micrometastatic tumor deposits up to 180 microm in diameter (1200 cells). PMID- 11280761 TI - Vascular endothelial growth factor effects on nuclear factor-kappaB activation in hematopoietic progenitor cells. AB - Vascular endothelial growth factor (VEGF) inhibits of the activation of transcription factor nuclear factor-kappaB (NF-kappaB) in hematopoietic progenitor cells (HPCs), and this is associated with alterations in the development of multiple lineages of hematopoietic cells and defective immune induction in tumor-bearing animals. Antibodies to VEGF have been shown to abrogate this effect. The mechanism by which VEGF antagonizes the induction of NF kappaB was investigated in this study. Using supershift electrophoretic mobility shift analysis, we found that although tumor necrosis factor alpha (TNF-alpha) induced the nuclear translocation and DNA binding of p65-containing complexes, VEGF alone induced nuclear translocation and DNA binding of the complexes containing RelB. These results were confirmed by immunofluorescence confocal microscopy. VEGF effectively blocked TNF-alpha-induced NF-kappaB activation in HPCs from RelB-/- mice, however, similar to the effect observed in HPCs obtained from RelB+/- and RelB+/+ mice. This suggests that RelB is not required for VEGF to inhibit NF-kappaB activation. However, although TNF-alpha induced rapid activation of IkappaB kinase (IKK) as expected, this activity was substantially reduced in the presence of VEGF. This decreased IKK activation correlated with the inhibition of IkappaB alpha phosphorylation and degradation of IkappaB alpha and IkappaB epsilon in HPCs. VEGF alone, however, did not have any effect on phosphorylation of IkappaB alpha or degradation of IkappaB alpha and other inhibitory molecules IkappaB beta, IkappaB epsilon, or Bcl-3. SU5416, a potent inhibitor of the VEGF receptor I (VEGFR1) and VEGFR2 receptor tyrosine kinases, did not abolish the inhibitory effect of VEGF, indicating that the VEGF effect is mediated by a mechanism unrelated to VEGFR1 or VEGFR2 tyrosine kinase activity. Thus, VEGF appears to inhibit TNF-alpha-induced NF-kappaB activation by VEGFR kinase-independent inhibition of IKK. Therapeutic strategies aimed at overcoming VEGF-mediated defects in immune induction in tumor-bearing hosts will need to target this kinase-independent pathway. PMID- 11280762 TI - Stimulation of beta1 integrin down-regulates ICAM-1 expression and ICAM-1 dependent adhesion of lung cancer cells through focal adhesion kinase. AB - Adhesion molecules are involved in intracellular signaling in various physiological and pathological processes including metastasis and growth of tumor cells. Tumor cells interact with various host cells as well as with extracellular matrices through certain adhesion molecules such as integrins. We here propose that stimulation of beta1 integrin reduces intercellular adhesion molecule (ICAM) 1-mediated interaction of lung cancer cells with CTLs. This concept is based on the following findings: (a) engagement of beta1 integrins on certain lung cancer cells by a specific antibody or by ligand matrices such as fibronectin and collagen markedly reduced ICAM-1 expression on the cell surface and induced sICAM 1; (b) down-regulation of ICAM-1 by stimulation of beta1 integrins was abrogated by tyrosine kinase inhibitors or by transfection of dominant negative truncations of focal adhesion kinase (FAK); (c) engagement of beta1 integrins also reduced ICAM-1-dependent adhesion of lung cancer cells to T cells, a process completely inhibited by tyrosine kinase inhibitors and by transfection of dominant negative forms of FAK; and (d) stimulation of beta1 integrins prevented killing of lung cancer cells by autologous CTLs. In malignant tumors, cancer cells, including lung cancer cells, are surrounded by extracellular matrix proteins such as fibronectin and collagen. This suggests that the engagement of beta1 integrins by matrix proteins potentially occurs in cancer cells in vivo and that continuous stimulation via beta1 integrins reduces ICAM-1-expression, ICAM-1-mediated adhesion of cancer cells to CTLs and their killing by CTLs. Our results suggest that such processes can lead to the escape of lung cancer cells in vivo from immunological surveillance. PMID- 11280763 TI - Divergent effects of 4-1BB antibodies on antitumor immunity and on tumor-reactive T-cell generation. AB - 4-1BB is an inducible receptor-like protein expressed rapidly by both CD4 and CD8 T-cells after activation. 4-1BB cross-linking, either by binding to 4-1BBL or by antibody ligation, delivers a costimulatory signal to enhance T-cell activation and proliferation. Previous studies have demonstrated that the administration of 4-1BB monoclonal antibodies (mAbs) induces antitumor immune responses. In the current study using several murine tumors, we examined the systemic effects of 4 1BB mAb on the growth of s.c., intracranial (i.c.), and pulmonary metastases. In addition, the effects of 4-1BB mAb on the generation of antitumor effector T cells were examined. Treatment of 3-day i.c. MCA 205 sarcoma and GL261 glioma with the antibody resulted in prolongation of survival and cure of disease in some mice, whereas only minimal therapeutic effects were observed in established s.c. and pulmonary tumors. No antitumor effects against the poorly immunogenic B16/D5 melanoma were observed. Interestingly, successful treatment of i.c. tumors induced concomitant regression of s.c. tumors. Experiments using severe combined immunodeficient mice and mice depleted of either CD4 or CD8 T cells demonstrated T-cell dependence of the antitumor effects. For generation of effector T cells in the tumor-draining lymph nodes (LNs), administration of 4-1BB mAb had adverse effects, despite the apparent hypertrophy of the LNs. During in vitro activation of tumor-draining LN T cells with anti-CD3 and interleukin 2, the 4-1BB mAb augmented proliferation, resulting in an increase in CD8 T cells. However, they were less therapeutic than not treated LN cells. In adoptive immunotherapy, the coadministration of 4-1BB mAb enhanced the therapeutic efficacy. These results thus demonstrate the limits and potential advantages of 4-1BB antibody interactions with antitumor T cells in vivo and in vitro and suggest that therapeutic interactions of the antibody may be used in a variety of immunotherapeutic approaches. PMID- 11280764 TI - Molecular basis of T cell-mediated recognition of pancreatic cancer cells. AB - Pancreatic cancer continues to be a major unsolved health problem in the world. The prognosis of pancreatic cancer is extremely poor with a median survival of 3 4 months and the 5-year survival being 1-4%. This poor prognosis is primarily because of a lack of effective therapies, and thus development of new treatment modalities is needed. One of these treatments could involve specific immunotherapy, for which elucidation off the molecular basis of T cell-mediated recognition of cancer cells is required. We report here six different genes and 19 immunogenic epitopes from pancreatic adenocarcinoma cells and T-cell receptor beta usage of HLA-A2-restricted CTL clones reacting to some of these epitopes. Sixteen of 19 epitopes were found to possess the ability to induce HLA-A2 restricted CTL activity in the peripheral blood lymphocytes of patients with pancreatic and also colon adenocarcinomas. These results should provide a scientific basis for the development of specific immunotherapy for pancreatic and colon cancer patients. PMID- 11280765 TI - Melanoma patients respond to a cytotoxic T lymphocyte-defined self-peptide with diverse and nonoverlapping T-cell receptor repertoires. AB - HLA-A2+ melanoma patients develop naturally a strong CD8+ T cell response to a self-peptide derived from Melan-A. Here, we have used HLA-A2/peptide tetramers to isolate Melan-A-specific T cells from tumor-infiltrated lymph nodes of two HLA A2+ melanoma patients and analyzed their TCR beta chain V segment and complementarity determining region 3 length and sequence. We found a broad diversity in Melan-A-specific immune T-cell receptor (TCR) repertoires in terms of both TCR beta chain variable gene segment usage and clonal composition. In addition, immune TCR repertoires selected in the patients were not overlapping. In contrast to previously characterized CD8+ T-cell responses to viral infections, this study provides evidence against usage of highly restricted TCR repertoire in the natural response to a self-differentiation tumor antigen. PMID- 11280766 TI - Identification of a tissue-specific putative transcription factor in breast tissue by serological screening of a breast cancer library. AB - Application of SEREX (serological analysis of recombinant tumor cDNA expression libraries) to different tumor types has led to the identification of several categories of human tumor antigens. In this study, the analysis of a breast cancer library with autologous patient serum led to the isolation of seven genes, designated NY-BR-1 through NY-BR-7. NY-BR-1, representing 6 of 14 clones isolated, showed tissue-restricted mRNA expression in breast and testis but not in 13 other normal tissues tested. Among tumor specimens, NY-BR-1 mRNA expression was found in 21 of 25 breast cancers but in only 2 of 82 nonmammary tumors. Structural analysis of NY-BR-1 cDNA and the corresponding genomic sequences in the recently released working draft of human genome indicated that NY-BR-1 is composed of 37 exons and has an open reading frame of 4.0-4.2 kb, encoding a peptide of Mr 150,000-160,000. A bipartite nuclear localization signal motif indicates a nuclear site for NY-BR-1, and the presence of a bZIP site (DNA binding site followed by leucine zipper motif) suggests that NY-BR-1 is a transcription factor. Additional structural features include five tandem ankyrin repeats, implying a role for NY-BR-1 in protein-protein interactions. NY-BR-1 thus represents a breast tissue-specific putative transcription factor with autoimmunogenicity in breast cancer patients. In addition to NY-BR-1, a homologous gene, NY-BR-1.1, was identified in this study. NY-BR-1.1 shares 54% amino acid homology with NY-BR-1 and also shows tissue-restricted mRNA expression. However, unlike NY-BR-1, NY-BR-1.1 mRNA is expressed in brain, in addition to breast and testis. The exon structure of NY-BR-1.1 remains to be defined. Using human genome database, NY-BR-1 was localized to chromosome 10p11 p12, and NY-BR-1.1 was tentatively localized to chromosome 9. PMID- 11280767 TI - T cell-dependent antitumor immunity mediated by secondary lymphoid tissue chemokine: augmentation of dendritic cell-based immunotherapy. AB - Secondary lymphoid tissue chemokine (SLC) is a CC chemokine that is selective in its recruitment of naive T cells and dendritic cells (DCs). In the lymph node, SLC is believed to play an important role in the initiation of an immune response by colocalizing naive T cells with DC-presenting antigen. Here, we used SLC as a treatment for tumors established from the poorly immunogenic B16 melanoma. Intratumoral injections of SLC inhibited tumor growth in a CD8+, T cell-dependent manner. SLC elicited a substantial infiltration of DCs and T cells into the tumor, coincident with the antitumor response. We next used SLC gene-modified DCs as a treatment of established tumors. Intratumoral injections of SLC-expressing DCs resulted in tumor growth inhibition that was significantly better than either control DCs or SLC alone. Distal site immunization of tumor-bearing mice with SLC gene-modified DCs pulsed with tumor lysate elicited an antitumor response whereas control DCs did not. We also found that s.c. injection of lysate-pulsed DCs expressing SLC promoted the migration of T cells to the immunization site. This report demonstrates that SLC can both induce antitumor responses and enhance the antitumor immunity elicited by DCs. PMID- 11280768 TI - Regulation of growth and tumorigenicity of breast cancer cells by the low molecular weight GTPase Rad and nm23. AB - Rad is the prototypic member of a family of novel Ras-related GTPases that is normally expressed in heart, skeletal muscle, and lung and that has been shown to exhibit a novel form of bi-directional interaction with the nm23 metastasis suppressor. In the present study, we have investigated the expression of Rad in normal and neoplastic breast tissues by Western blot and immunohistochemistry and the functional effect of altered Rad expression in breast cancer cell lines. We found that, although Rad is frequently expressed in normal breast tissue (23/30 Rad+ve), expression is usually lost in adjacent invasive carcinoma (8/30 Rad+ve; P < 0.0001). However, where Rad expression persists in a small proportion of tumors, it is associated with higher grade, larger size, and extensive axillary nodal involvement (n = 48; P = 0.035, P = 0.016, P = 0.022, respectively). Furthermore, Rad is also highly expressed in a breast cancer cell line with high tumorigenic and metastatic potential (MDA-MB231). To further examine the role of Rad in breast cancer, we stably transfected a Rad-ve breast cancer cell line (MDA MB435). We observed an increase in growth and marked increased colony formation in soft agar in vitro (P < 0.05) and an increase in tumor growth rate in nude mice (P < 0.05). Moreover, coexpression of nm23 with wild-type Rad inhibited the effect of Rad on growth of these cells in culture and markedly inhibited tumor growth in vivo. Additional transfection studies with mutated Rad cDNAs revealed that the growth-promoting effects of Rad appeared to be mediated through its NH2- and COOH-terminal regions, rather than its GTPase domain, and might involve acceleration of cell cycle transition. These findings suggest that Rad may act as an oncogenic protein in breast tissues and demonstrate a potential mechanism by which interaction between Rad and nm23 may regulate growth and tumorigenicity of breast cancer. PMID- 11280769 TI - Oct-2 and Bob-1 deficiency in Hodgkin and Reed Sternberg cells. AB - Hodgkin and Reed Sternberg (H-RS) cells represent the malignant cells in classical Hodgkin's disease. Although derived from germinal center B cells, they do not express surface immunoglobulin. This has been explained by the presence of crippling mutations within the immunoglobulin genes in numerous cases of Hodgkin's disease. As immunoglobulin gene expression in B cells requires an interaction between octamer sites and the transactivating factors Oct-2 and Bob 1, this study addresses the expression of the transcription factors Oct-2 and Bob 1 in H-RS cells. In Hodgkin's disease-derived cell lines, low levels of Oct-2 transcripts but no Oct-2 protein were detected. Transcripts of Bob-1, a B-cell specific co-factor of Oct-2, could not be observed in these cell lines. Absence of Oct-2 and Bob-1 protein expression in primary H-RS cells was demonstrated by performing immunohistochemistry in 20 cases of classical Hodgkin's disease. H-RS cells stained negative for both proteins in all of the cases analyzed. In conclusion, absence of functional Oct-2 and Bob-1 cells represents a novel mechanism for immunoglobulin gene deregulation in H-RS cells. Lack of Oct-2 and Bob-1 points to a defect in transcription machinery in H-RS cells and is associated with lack of immunoglobulin gene expression in these cells. PMID- 11280770 TI - Activation of beta-catenin during hepatocarcinogenesis in transgenic mouse models: relationship to phenotype and tumor grade. AB - Mutations affecting phosphorylation sites in the beta-catenin gene have been implicated in the development of human and rodent hepatocellular carcinomas (HCCs). To further investigate the involvement of this gene in hepatocarcinogenesis, we used several transgenic mouse models of hepatic tumors induced by overexpression of c-myc in the liver either alone or in combination with transforming growth factor (TGF) alpha or TGF-beta1. Activation of beta catenin, as judged by the presence of mutations and/or nuclear translocation of the protein, was most frequent in liver tumors from c-myc (4/17; 23.5%) and c myc/TGF-beta1 (6/18; 33.3%) transgenic mice. However, it was very rare in faster growing and histologically more aggressive HCCs developed in c-myc/TGF-alpha mice (1/20; 5%). Administration of diethylnitrosamine, phenobarbital, or 2-amino-3,8 diethylimidazo[4,5-f]quinoxaline did not significantly affect the occurrence of beta-catenin mutations. Notably, nuclear accumulation of beta-catenin was observed only in adenomas and highly differentiated carcinomas with eosinophilic phenotype. Furthermore, preneoplastic lesions with eosinophilic phenotype frequently displayed focal nuclear positivity, colocalized with areas of high proliferation. In contrast, basophilic and clear-cell foci, as well as pseudo glandular and poorly differentiated HCCs, exhibited a normal or reduced membranous immunoreactivity for beta-catenin. These studies suggest that nuclear translocation of beta-catenin and activation of Wingless/Wnt signaling may represent an early event in liver carcinogenesis, providing a growth advantage in a subset of hepatic tumors with a more differentiated phenotype. PMID- 11280771 TI - Increased loss of chromosome 9p21 but not p16 inactivation in primary non-small cell lung cancer from smokers. AB - Epidemiological studies have demonstrated a causal association between tobacco use and carcinoma of the lung, and some genetic targets of the carcinogens in cigarette smoke have been defined recently. We further examined the effect of cigarette smoking on the frequency of allelic losses on chromosome 9p21 and the incidence of p16 inactivation. Chromosomal loss at 9p21-24 was determined by microsatellite analysis using 14 markers in 47 patients with non-small cell lung cancer. In addition, p16 gene inactivation was determined by DNA sequence analysis, methylation-specific PCR, and immunohistochemistry. Tumors from a group of nonsmokers (n = 14) were compared with tumors from a group of smokers (n = 33) matched for cell type, tumor stage, and gender. Allelic loss encompassing the p16 locus was present significantly (P = 0.01) more often in smokers (23 of 33 smokers, 70%) than in nonsmokers (4 of 14 nonsmokers, 28%). No significant differences in the frequency of p16 inactivation were observed between smokers and nonsmokers (45% versus 36%). However, homozygous deletion of the p16 gene locus and point mutation of p16 gene were only observed in tumors from smokers, whereas the p16 gene was inactivated in tumors from nonsmokers only through promoter hypermethylation. Thus, inactivation of the p16 gene is a common event in all non-small cell lung cancer, but the mechanism of gene alteration differs between smokers and nonsmokers. The significant link between tobacco and loss of the p16 locus identifies additional genetic targets of smoking in the pathogenesis of lung cancer. PMID- 11280772 TI - Stabilized beta-catenin immortalizes colonic epithelial cells. AB - The majority of colonic neoplasias contain mutations in either the adenomatous polyposis coli or the beta-catenin (beta-cat) gene, both of which result in elevated levels of cytoplasmic beta-cat. The oncogenic activity of beta-cat has been explored in vivo and in vitro with conflicting results. We tested the hypothesis that beta-cat is capable of immortalizing and transforming cultured epithelial cells that represent the precursors to colon cancer. A truncated form of beta-cat (deltaN89) was stably expressed in murine colonic epithelial cells that were conditionally immortalized by temperature-sensitive T antigen expression and contained a mutant ApcMin allele [Immorto-Min colonic epithelium (IMCE)]. IMCE cells, grown under nonpermissive conditions, were immortalized by expression of the truncated beta-cat protein as determined by sustained growth in culture and escape from senescence as measured by endogenous beta-galactosidase activity. IMCE neo cells at nonpermissive conditions underwent extensive apoptosis, an effect that was blocked by the expression of deltaN89 beta-catenin. IMCE beta-cat cells had significantly lower p19 and p53 protein levels compared to IMCE neo cells, suggesting that alterations in these two key genes may mediate the effects of beta-cat on both cellular senescence and apoptosis. IMCE beta-cat cells were also transformed as determined by growth in the absence of serum, anchorage-independent growth, and sustained tumor growth in nude mice. Stable beta-cat-expressing populations could not be generated in conditionally immortalized colonic epithelia cells with a wild-type Apc background. These studies demonstrated the immortalizing activity of stabilized beta-cat for the first time and extend the transforming ability of mutated beta-cat to a cell line representing a precursor to colorectal cancer. PMID- 11280773 TI - p27Kip1 is required for PTEN-induced G1 growth arrest. AB - The tumor suppressor PTEN is one of the most commonly inactivated genes in human cancer. Glioblastoma multiforme cells harboring mutant PTEN have abnormally high levels of 3' phosphoinositides and elevated protein kinase B activity. Expression of wild-type PTEN in glioma cells, containing endogenous mutant PTEN, reduces 3' phosphoinositides levels, inhibits PKB activity, and induces G1 cell cycle arrest. We investigated the mechanism of the PTEN-induced growth arrest in glioma cell lines. Expression of PTEN is associated with increased expression of p27Kip1, decreased expression of cyclins A and D3, inhibition of cdk2 activity, and dephosphorylation of pRb. Inactivation of p53, by the human papilloma virus E6 oncoprotein, does not prevent PTEN-induced G1 arrest, implying that p53 is not required for G1 arrest. In contrast, p27Kip1 antisense oligonucleotides abrogated the growth arrest induced by PTEN. Furthermore, blocking p27Kip1 expression prevented the PTEN-induced reduction of cyclin-dependent kinase 2 activity, indicating that p27Kip1 functions upstream of cyclin-dependent kinase 2 in the PTEN regulatory cascade. These results implicate p27Kip1 as a critical mediator of PTEN-induced G1 arrest. PMID- 11280774 TI - SMAD3 represses androgen receptor-mediated transcription. AB - The androgen-signaling pathway is important in the growth and progression of prostate cancer. Androgen ablation therapy, which may result in programmed cell death, is often used to treat advanced prostate cancer. The growth-promoting effects of androgen are mediated mostly through the androgen receptor (AR). Transforming growth factor beta (TGF-beta) plays critical roles in controlling prostate cell proliferation, differentiation, and apoptosis. Normal transcripts and proteins of TGF-beta receptors are frequently lost in prostate cancer cells, especially in advanced stages of the disease. However, the mechanisms by which TGF-beta inhibits proliferation and induces apoptosis in prostate cancer cells is not clear. We investigated the molecular mechanism by which TGF-beta inhibits transcriptional activation mediated by AR. Using transient transfection systems, we demonstrated that Smad3 specifically represses transcriptional activation mediated by AR on two natural androgen-responsive promoters. This repression is transmitted through TGF-beta signaling and can be regulated by other Smad proteins. A protein-protein interaction between AR and Smad3 was identified in vitro and in vivo, and the transcription activation domain of AR and the MH2 of Smad3 were identified as being responsible for binding. Additional functional experiments showed that the repression of AR by Smad3 is mediated solely through the MH2 domain. These results provide fresh insight for understanding the mechanism by which TGF-beta regulates the androgen-signaling pathway in prostate cancer cells. PMID- 11280775 TI - Mapping the sites of putative tumor suppressor genes at 6p25 and 6p21.3 in cervical carcinoma: occurrence of allelic deletions in precancerous lesions. AB - Allelic deletions on the short arm of chromosome 6 (6p) are one of the common, possibly early, genetic changes that occur in the pathogenesis of cervical carcinoma (CC). Previous loss of heterozygosity (LOH) studies in CC identified a number of critical regions of deletions on 6p. However, the precise location of minimally deleted regions and their role in precancerous lesions have not been well characterized. To address these questions, we first performed a detailed LOH analysis on 6p in 59 cases of invasive CC. The pattern of LOH identified two minimal regions of deletions, one spanning a 5 cM genetic distance at 6p25 and a second site of 10.3 cM deletion mapping to 6p21.3. The 6p21.3 minimal deletion spans HLA class I genes. To understand the role of 6p genetic alterations in the development of CC, we also investigated 12 high-grade and 4 low-grade cases of cervical intraepithelial neoplasia (CIN) for LOH after laser microdissection. The high-grade CINs exhibited 91.7% LOH, and low-grade CINs had 50% LOH. These findings implicate the presence of at least two tumor suppressor genes on 6p relevant to CC and suggest that these genetic alterations occur very early in CC development. This study should therefore facilitate the identification of tumor suppressor genes on 6p and may identify which CINs are at high risk of progressing to invasive CC. PMID- 11280776 TI - Microsatellite instability occurs in distinct subtypes of pediatric but not adult central nervous system tumors. AB - Length alterations in microsatellite repeats, termed microsatellite instability (MSI), are found in 10-15% of sporadic colon, endometrial, and gastric cancers harboring defects in DNA mismatch repair (MMR) genes We used the microsatellite markers Big Adenine Tract (BAT) 26 and BAT-25 from the reference panel of five markers recommended by the National Cancer Institute to evaluate the incidence of MSI in 206 central nervous system tumors. We screened 102 pediatric and 104 adult cases representing 165 astrocytic and 41 nonastrocytic tumors. The overall incidence of MSI was 8% (16 of 206). All 16 tumors with MSI were found in pediatric rather than adult patients. MSI was associated with two distinct subtypes of pediatric tumors occurring in 27% (12 of 45) of WHO grade III and grade IV astrocytomas and 24% (4 of 17) of gangliogliomas We evaluated the difference in clinicopathological and genetic features among 45 high-grade pediatric astrocytomas by MSI status. The median survival for pediatric patients with MSI (n = 12) was 8 months compared with 15 months for those patients without MSI (n = 33; P = 0.18). The frequency of p53 gene mutations was 13% for pediatric patients with MSI (n = 8) compared with 47% for those patients without MSI (n = 19; P = 0.19). These results revealed a trend between MSI status and prog nosis and MSI status and frequency of p53 gene mutations. Our data suggest that pediatric high-grade astrocytomas can be attributed to two different genetic pathways: a MMR-deficient pathway and a MMR proficient pathway. PMID- 11280777 TI - Genome-wide analysis of gene expression in human hepatocellular carcinomas using cDNA microarray: identification of genes involved in viral carcinogenesis and tumor progression. AB - To disclose detailed genetic mechanisms in hepatocellular carcinoma (HCC) with a view toward development of novel therapeutic targets, we analyzed expression profiles of 20 primary HCCs and their corresponding noncancerous tissues by means of cDNA microarrays consisting of 23,040 genes. Up-regulation of mitosis promoting genes was observed in the majority of the tumors examined. Some genes showed expression patterns in hepatitis B virus-positive HCCs that were different from those in hepatitis C virus-positive HCCs; most of them encoded enzymes that metabolize carcinogens and/or anticancer agents. Furthermore, we identified a number of genes associated with malignant histological type or invasive phenotype. Accumulation of such data will make it possible to define the nature of individual tumors, to provide clues for identifying new therapeutic targets, and ultimately to optimize treatment of each patient. PMID- 11280778 TI - Expression of vesicular monoamine transporters, synaptosomal-associated protein 25 and syntaxin1: a signature of human small cell lung carcinoma. AB - Vesicular monoamine transporters (VMATs) are a prerequisite for the uptake of biogenic amines into intracellular storage organelles, whereas soluble N ethylmaleimide-sensitive factor attachment protein receptors (SNAREs; such as SNAP-25 and syntaxin1) are essential for exocytosis of biogenic amines by neurons and endocrine cells. In this study, we examined whether these proteins exist in high-grade malignant small cell lung carcinomas (SCLCs), large cell carcinomas, adenocarcinomas, and squamous cell carcinomas of the lung. We analyzed two established human SCLC cell lines, one adenocarcinoma cell line, paraffin embedded tumors (SCLC, n = 25; large cell carcinoma, n = 10; adenocarcinoma, n = 10; squamous cell carcinoma, n = 10), and snap-frozen SCLC samples (n = 2). Using immunocytochemistry, Western blotting, Northern blotting, RT-PCR, and sequencing, we identified VMAT1, VMAT2, SNAP-25, and syntaxin1 in cultured SCLC cells. Immunohistochemistry carried out on paraffin sections revealed that all SCLC tumors express VMAT1, VMAT2, SNAP-25, and syntaxin1. The presence of SNAP-25 and syntaxin1 in SCLC was confirmed by RT-PCR performed with material extracted from paraffin sections. Western blot analysis and RT-PCR carried out with snap-frozen SCLC tumors revealed the presence of SNAREs and VMATs. Immunohistochemistry showed that non-SCLC tumors were negative for SNAREs and VMATs, with the exception of immunostaining for SNAP-25 and syntaxin1 in 3 of 10 adenocarcinomas. Our findings indicate that SCLC cells are endowed with transporters necessary for intracellular storage of biogenic amines and with proteins required for exocytosis of secretory products. These proteins may be used as markers of differentiation of human lung tumors. Moreover, the presence of VMATs provides the basis for a diagnostic application of biogenic amine-derived tracers in positron emission tomography of SCLC tumors. PMID- 11280779 TI - Angiopoietin-2 is related to tumor angiogenesis in gastric carcinoma: possible in vivo regulation via induction of proteases. AB - Tumor angiogenesis progresses by a dynamic balance between tumor vascular regression and growth. Angiopoietin (Ang)-2 (the natural antagonist for the angiogenic Tie-2 receptor) and vascular endothelial growth factor (VEGF) are thought to be critical regulators in this process; therefore, these may play a critical role in cancer aggressiveness. The aim of this study was to clarify the clinical and biological significance of the expression of Ang-2 in human gastric cancers and to investigate the relationship between Ang-2 together with VEGF and the induction of proteases such as matrix metalloproteinases (MMPs) in the process of tumor development. Eighty-five individuals with gastric cancer, who had undergone surgery without preoperative treatment, were studied. A stable transfectant of the human MKN-7 gastric cancer cell lines with an Ang-2 expression vector was used for the experimental study. First, we examined the relationship between the mRNA expression of Angs by Northern blot analysis and clinicopathological features. High Ang-2-expression cases showed more frequent vascular involvement and more advanced stages of disease compared with low Ang-2 expression cases (P < 0.05). With regard to prognosis, the survival time for patients in the high-Ang-2 mRNA group was significantly shorter (P < 0.05). When we examined the localization of Ang-2 in human gastric cancers, immunohistochemical analysis revealed that this protein was expressed predominantly in cancer tissues when compared with normal tissues. Interestingly it was expressed not only in endothelia cells (ECs) but also in cancer cells. Second, Ang-2-transfected cells were implanted in vivo into the gastric walls of nude mice. Ang-2-transfectant mice developed highly metastatic tumors with hypervascularity as compared with MKN-7 or control vector-transfectant tumors. There was a significant correlation between Ang-2 mRNA expression and lower grade of vessel maturation. Third, on the basis of the in vivo data, we focused on production of proteases such as MMPs to investigate possible mechanisms in these processes. MMP-1, MMP-9, and urokinase-type plasminogen activator in ECs were strongly up-regulated by Ang-2 in the presence of VEGF in vitro. These data suggest that production of Ang-2 is implicated in tumor development in human gastric cancers. Its production may contribute to tumor angiogenesis by induction of proteases in ECs, which may be enhanced in the presence of VEGF. PMID- 11280781 TI - The gene for the axonal cell adhesion molecule TAX-1 is amplified and aberrantly expressed in malignant gliomas. AB - The human TAX-1 gene encodes a Mr 135,000 glycoprotein that is transiently expressed on the surface of a subset of neurons during development and is involved in neurite outgrowth. The TAX-1 gene has been mapped to a region on chromosome 1 that has been implicated in microcephaly and the Van der Woude syndrome. Using restriction landmark genome scanning to search for amplified genes in gliomas, we found TAX-1 to be amplified in 2 high-grade gliomas among a group of 26 gliomas investigated. Real-time reverse transcription-quantitative PCR analysis detected high levels of TAX-1 mRNA in glial tumors, even in the absence of TAX-1 gene amplification. Immunohistochemical analysis revealed abundant levels of TAX-1 in neoplastic glial cells of glioblastoma multiforme tumors. Because glial tumors are highly invasive and in view of the role of TAX-1 in neurite outgrowth, we investigated the potential role of TAX-1 in glioma cell migration. Using an in vitro assay, we found that the migration of glioma tumor cells is profoundly reduced in the presence of either an anti-TAX-1 antibody or a TAX-1 antisense oligonucleotide. Our findings suggest that TAX-1 plays a role in glial tumorigenesis and may provide a potential target for therapeutic intervention. PMID- 11280780 TI - HuR, a RNA stability factor, is expressed in malignant brain tumors and binds to adenine- and uridine-rich elements within the 3' untranslated regions of cytokine and angiogenic factor mRNAs. AB - Tumors of the central nervous system (CNS) often have sustained expression of labile genes, including angiogenic growth factors and immunosuppressive cytokines, which promote tumor progression. Stabilization of the RNA transcripts for these genes, such as vascular endothelial growth factor (VEGF), is an important molecular pathway for this up-regulation. HuR, a member of the Elav family of RNA-binding proteins, has been implicated in this pathway through its binding to adenine and uridine (AU)-rich stability elements (ARE) located in the 3' untranslated regions (3'-UTRs) of the mRNA. Whereas three of the Elav family members (Hel-N1, HuC, and HuD) are restricted to young and mature neurons, HuR is more broadly expressed, including proliferating cells of the developing CNS. Because RNA stabilization of labile genes may promote tumor growth, we analyzed and compared the expression pattern of HuR in 35 freshly resected and cultured CNS tumors to determine whether there was any correlation with tumor grade or histological type. We found that HuR mRNA was consistently expressed in all of the tumors, regardless of cell origin or degree of malignancy. Using a novel HuR specific polyclonal antibody, we found that strong HuR protein expression was limited to high-grade malignancies (glioblastoma multiforme and medulloblastoma). Within the glioblastoma multiforme, prominent HuR expression was also detected in perinecrotic areas in which angiogenic growth factors are up-regulated. To further define its role as a potential RNA stabilizer, we analyzed whether HuR could bind to the stability motifs within the 3'-UTRs of cytokines and growth factors linked to brain tumor progression. We used a novel ELISA-based RNA binding assay and focused on the 3'-UTRs of angiogenic factors VEGF, COX-2, and (interleukin) IL-8 as well as the immunomodulating factors IL-6, transforming growth factor (TGF)-beta and tumor necrosis factor (TNF)-alpha as potential RNA ligands. Our results indicated overall a very high binding affinity to these RNA targets. A comparison of these ligands revealed a hierarchy of binding affinities with the angiogenic factors, and TGF-beta showing the highest (Kd of 1.8-3.4 nM), and TNF-alpha the lowest (Kd of 18.3 nM). The expression pattern of HuR, coupled with the RNA binding data, strongly suggests a role for this protein in the posttranscriptional regulation of these genes in CNS tumors. PMID- 11280782 TI - Expression of erbB-4/HER-4 growth factor receptor isoforms in ovarian cancer. AB - Immunohistochemical expression of erbB4 protein was identified in 93% (49 of 53) of ovarian cancers using the HFR-1 antibody (targeted to the cytoplasmic domain of the erbB4 receptor) and in 89% (47 of 53) of ovarian cancers using the H4.77.16 antibody (targeted to the extracellular domain). Tumors of serous histology were more likely to express a higher level of erbB4 than endometrioid tumors, and for stage III serous tumors, long-term survival was associated with moderate to high coexpression of erbB4 and erbB2. Within ovarian cancer cell lines, high erbB4 expression was associated with cisplatin resistance. Using reverse transcription-PCR, the presence of multiple isoforms of erbB4 mRNA was identified in both ovarian primary tumors and cell lines. Splice variants in the juxtamembrane (JM-a and JM-d) and cytoplasmic (CT-a and CT-b) regions were identified in mRNA of both cell lines and primary tumors. The use of an anti erbB4 blocking antibody suggested that erbB4 was not the mediator of the growth stimulatory effects of neuregulin in ovarian cancer cells and indeed could potentially antagonize this effect. PMID- 11280783 TI - Establishment of a novel species- and tissue-specific metastasis model of human prostate cancer in humanized non-obese diabetic/severe combined immunodeficient mice engrafted with human adult lung and bone. AB - Bone is the most common site of metastasis in prostate cancer (PC), and to generate an animal model to investigate the basis of the unique organ tropism of PC cells for bone, we engrafted humanized non-obese diabetic/severe combined immunodeficient (NOD/SCID-hu) mice with human adult bone (HAB) and lung (HAL). Human PC cell lines LNCaP (1 x 10(7)) and PC-3 (5 x 10(6)) were injected into male NOD/SCID-hu mice via the lateral tail vein at 3-4 weeks after implantation. At 8 weeks after the injection, LNCaP and PC-3 cells had metastasized specifically to HAB in 35 and 65%, respectively, of the mice. The tumors formed by LNCaP appeared to be the osteoblastic type, whereas the PC-3 tumors consisted of osteolytic lesions without any surrounding osteogenic response. A feature of experimental metastasis of PC in NOD/SCID-hu mice was its specificity for HAB tissue. Human PC cells had no or very low metastatic potential in regard to implanted HAL, mouse bone, or native mouse bone. These findings indicate that metastasis of PC cells to HAB is both species and tissue specific. The availability of this small animal model could provide a useful tool for identifying and analyzing important features of the human PC metastatic process that cannot be addressed in conventional metastasis models. PMID- 11280784 TI - Up-Regulation of Bcl-2 in microvascular endothelial cells enhances intratumoral angiogenesis and accelerates tumor growth. AB - Vascular endothelial growth factor (VEGF) has been shown to be a potent mediator of angiogenesis that functions as a survival factor for endothelial cells by up regulating Bcl-2 expression. We have recently reported that human dermal microvascular endothelial cells (HDMECs) seeded in biodegradable sponges and implanted into severe combined immunodeficient (SCID) mice organize into functional human microvessels that transport mouse blood cells. In this study, we implanted sponges seeded with OSCC-3 (oral squamous cell carcinoma) or SLK (Kaposi's sarcoma) together with endothelial cells into SCID mice to generate human tumors vascularized with human microvessels. This model system was used to examine the role of both endothelial cell Bcl-2 and the proangiogenic chemokine interleukin-8 (IL-8) on tumor growth and intratumoral microvascular density. Coimplantation of HDMECs overexpressing Bcl-2 (HDMEC-Bcl-2) and tumor cells resulted in a 3-fold enhancement of tumor growth when compared with the coimplantation of control HDMECs and tumor cells. This was associated with increased intratumoral microvascular density and enhanced endothelial cell survival. To determine whether the enhanced neovascularization mediated by Bcl-2 overexpression in endothelial cells was influenced by the synthesis of endogenous mediators of angiogenesis, we screened these cells for expression of VEGF, basic fibroblast growth factor (bFGF), and IL-8 by ELISA. HDMEC-Bcl-2 cells and VEGF treated HDMECs exhibited a 15-fold and 4-fold increase, respectively, in the expression of the proangiogenic chemokine IL-8 in vitro, whereas the expression of VEGF and bFGF remained unchanged. Transfection of antisense Bcl-2 into HDMECs blocked VEGF-mediated induction of IL-8. Conditioned media from HDMEC-Bcl-2 induced proliferation and sprouting of endothelial cells in vitro and neovascularization in rat corneas. Anti-IL-8 antibody added to HDMEC-Bcl-2 conditioned media markedly reduced the potency of these responses. SCID mice bearing VEGF-producing tumor implants that were treated with anti-lL-8 antibody exhibited a 43% reduction in microvessel density and a 50% reduction in tumor weight compared with treatment with a nonspecific antibody. These results demonstrate that the up-regulation of Bcl-2 expression in endothelial cells that constitute tumor microvessels enhances intratumoral microvascular survival and density and accelerates tumor growth. Furthermore, endothelial cells that overexpress Bcl-2 have more angiogenic potential than control cells, and IL-8 neutralizing antibodies attenuate their angiogenic activity in vitro and in vivo. PMID- 11280785 TI - High cancer cell death in syngeneic tumors developed in host mice deficient for the stromelysin-3 matrix metalloproteinase. AB - Matrix metalloproteinases (MMPs) are extracellular enzymes. Some of them are known to be involved in tumor development and/or progression. Several cellular functions have been proposed for MMPs during malignant processes. Notably, they may be involved in tissue-remodeling processes through their ability to digest matrix components or to participate in tumor neoangiogenesis and, subsequently, in cancer cell proliferation. One of these MMPs, stromelysin-3 (ST3/MMP11), although devoid of enzymatic activity against the matrix components, is associated with human tumor progression and poor patient clinical outcome. Using several in vivo experimental models, it has been demonstrated that ST3 expression by the fibroblastic cells surrounding malignant epithelial cells promotes tumorigenesis in a paracrine manner. The present study was devoted to the identification of the cellular function underlying this ST3-induced tumor promotion using a syngeneic tumorigenesis model in mice. Our results show that ST3 exhibits a new and unexpected role for a MMP, because ST3-increased tumorigenesis does not result from increased neoangiogenesis or cancer cell proliferation but from decreased cancer cell death through apoptosis and necrosis. Thus, during malignancy, the cellular function of ST3 is to favor cancer cell survival in the stromal environment. PMID- 11280786 TI - Role of phosphatidylinositol 3-kinase-Akt pathway in nucleophosmin/anaplastic lymphoma kinase-mediated lymphomagenesis. AB - The NPM/ALK fusion gene, formed by the t(2;5) translocation in a subset of anaplastic large cell lymphomas, encodes a Mr 75,000 hybrid protein that contains the NH2-terminal portion of the nucleolar phosphoprotein nucleophosmin (NPM) joined to the entire cytoplasmic portion of the receptor tyrosine kinase anaplastic lymphoma kinase (ALK). NPM/ALK encodes a constitutively activated tyrosine kinase that belongs to the family of tyrosine kinases activated by chromosomal translocations. Our studies showed that NPM/ALK, similar to other members of this family, activates phosphatidylinositol 3-kinase (PI3K) and its downstream effector, serine/threonine kinase (Akt). PI3K was found in complex with NPM/ALK. Both PI3K and Akt kinase were permanently activated in NPM/ALK transfected BaF3 murine hematopoietic cells and in NPM/ALK-positive, but not in NPM/ALK-negative, patient-derived anaplastic large cell lymphoma cell lines. In addition, Akt was phosphorylated/activated in protein samples isolated from four patients diagnosed with ALK-positive T/null-cell lymphomas. The PI3K inhibitors wortmannin and LY294002 induced apoptosis in NPM/ALK+ cells but exerted only minor effects on the control BaF3 parental cells and peripheral blood mononuclear cells stimulated by growth factors. Furthermore, retroviral infection of NPM/ALK+ BaF3 cells with a dominant-negative PI3K mutant (delta p85) or a dominant negative Akt mutant (K179M) inhibited proliferation and clonogenic properties of the infected cells. Finally, the Akt mutant (K179M) suppressed the tumorigenicity of NPM/ALK-transfected BaF3 cells injected into syngeneic mice. In conclusion, our data indicate that NPM/ALK constitutively activates the PI3K-Akt pathway and that this pathway plays an important role in the NPM/ALK-mediated malignant transformation. PMID- 11280787 TI - Aberrant activation of c-kit protects colon carcinoma cells against apoptosis and enhances their invasive potential. AB - Multiple genetic aberrations contribute to the development of biologically aggressive, clinically malignant colorectal carcinomas (CRCs). Some of these have been linked to inappropriate signaling through the tyrosine kinase moieties of growth factor receptors. We have described previously (G. Bellone et al., J. Cell. Physiol., 172: 1-11, 1997) that human CRCs overexpress both the receptor tyrosine kinase c-kit and its ligand, stem cell factor (SCF), relative to normal mucosa cells, thus establishing an autocrine c-kit-mediated loop. In addition, we noted that exogenous SCF contributes to anchorage-independent growth of HT-29 colon carcinoma cells in semisolid medium. Here, we investigated possible roles of the c-kit/SCF autocrine/paracrine system in survival and invasive capacity of DLD-1 colon carcinoma cells. We report that SCF was required for migration and invasion of DLD-1 cells through reconstituted basement membranes (Matrigel) and up-regulated gelatinase (matrix metalloproteinase-9) activity in DLD-1 cells. Furthermore, we describe that SCF supported survival of DLD-1 cells in growth factor-deprived conditions. These results suggest multiple roles of c-kit activation in support of the malignant phenotype of DLD-1 cells related to growth, survival, migration, and invasive potential. PMID- 11280788 TI - Radiation therapy to a primary tumor accelerates metastatic growth in mice. AB - The surgical removal of a primary tumor can result in the rapid growth of metastases. The production of angiogenesis inhibitors by the primary tumor is one mechanism for the inhibition of metastatic tumor growth. The effect of curative radiotherapy to a primary tumor known to make an inhibitor of angiogenesis and the effects on distant metastases has not been studied. We here show that the eradication of a primary Lewis lung carcinoma (LLC-LM), which is known to generate angiostatin, is followed by the rapid growth of metastases that kill the animal within 18 days after the completion of radiation therapy. The right thighs of C57BL/6 mice (n = 25) were injected s.c. with 1 x 10(6) LLC-LM cells. Animals were randomized to one of five groups: no irradiation, 40 Gy in one fraction, 30 Gy in one fraction, 40 Gy in two 20 Gy fractions, or 50 Gy in five 10 Gy fractions. Tumors were clinically eradicated in each treatment group. All of the surviving animals became dyspneic and were killed within 14-18 days after the completion of radiation therapy. Examination of their lungs revealed >46 (range, 46-62) surface metastases in the treated animals compared with 5 (range, 2-8) in the untreated animals. The lung weights had increased from 0.2 g (range, 0.19 0.22 g) in the controls to 0.58 g (range 0.44-0.84) in the experimental animals. The most effective dose regimen was 10 Gy per fraction for five fractions, and serial experiments were conducted with this fractionation scheme. Complete response of the primary tumor was seen in 25 of 35 (71%) mice. The average weight of the lungs in the nonirradiated animals was 0.22 g (range, 0.19-0.24 g) and in the irradiated animals was 0.66 g (range, 0.61-0.70 g). The average number of surface metastases increased from five per lung (range, 2-13) in the control animals to 53 per lung (range, 46-62) in the irradiated animals. Both differences were statistically significant with P < 0.001. If the nontumor-bearing leg was irradiated or the animals were sham-irradiated, no difference in the number of surface metastases or lung weights was observed between the control group and the treated group. Urinary levels of matrix metalloproteinase 2, the enzyme responsible for angiostatin processing in this tumor model, were measured and correlated with the viability and size of the primary tumor. Administration of recombinant angiostatin prevented the growth of the metastases after the treatment of the primary tumor. In this model, the use of radiation to eradicate a primary LLC-LM tumor results in the growth of previously dormant lung metastases and suggests that combining angiogenesis inhibitors with radiation therapy may control distant metastases. PMID- 11280789 TI - Centrosome defects can account for cellular and genetic changes that characterize prostate cancer progression. AB - Factors that determine the biological and clinical behavior of prostate cancer are largely unknown. Prostate tumor progression is characterized by changes in cellular architecture, glandular organization, and genomic composition. These features are reflected in the Gleason grade of the tumor and in the development of aneuploidy. Cellular architecture and genomic stability are controlled in part by centrosomes, organelles that organize microtubule arrays including mitotic spindles. Here we demonstrate that centrosomes are structurally and numerically abnormal in the majority of prostate carcinomas. Centrosome abnormalities increase with increasing Gleason grade and with increasing levels of genomic instability. Selective induction of centrosome abnormalities by elevating levels of the centrosome protein pericentrin in prostate epithelial cell lines reproduces many of the phenotypic characteristics of high-grade prostate carcinoma. Cells that transiently or permanently express pericentrin exhibit severe centrosome and spindle defects, cellular disorganization, genomic instability, and enhanced growth in soft agar. On the basis of these observations, we propose a model in which centrosome dysfunction contributes to the progressive loss of cellular and glandular architecture and increasing genomic instability that accompany prostate cancer progression, dissemination, and lethality. PMID- 11280790 TI - Altered expression of Ape1/ref-1 in germ cell tumors and overexpression in NT2 cells confers resistance to bleomycin and radiation. AB - The human AP endonuclease (Ape1 or ref-1) DNA base excision repair (BER) enzyme is a multifunctional protein that has an impact on a wide variety of important cellular functions including oxidative signaling, transcription factor regulation, and cell cycle control. It acts on mutagenic AP (baseless) sites in DNA as a critical member of the DNA BER repair pathway. Moreover, Ape1/ref-1 stimulates the DNA-binding activity of transcription factors (Fos-Jun, nuclear factor-kappaB, Myb, ATF/cyclic AMP-responsive element binding protein family, HIF 1alpha, HLF, PAX, and p53) through a redox mechanism and thus represents a novel component of signal transduction processes that regulate eukaryotic gene expression. Ape1/ref-1 has also been shown to be closely linked to apoptosis associated with thioredoxin, and altered levels of Ape1/ref-1 have been found in some cancers. In a pilot study, we have examined Ape1/ref-1 expression by immunohistochemistry in sections of germ cell tumors (GCTs) from 10 patients with testicular cancer of various histologies including seminomas, yolk sac tumors, and malignant teratomas. Ape1/ref-1 was expressed at relatively high levels in the tumor cells of nearly all sections. We hypothesized that elevated expression of Ape1/ref-1 is responsible in part for the resistance to therapeutic agents. To answer this hypothesis, we overexpressed the Ape1/ref-1 cDNA in the GCT cell line NT2/D1 using retroviral gene transduction with the vector LAPESN. Using an oligonucleotide cleavage assay and immunohistochemistry to assess Ape1/ref-1 repair activity and expression, respectively, we found that the repair activity and relative Ape1/ref-1 expression in GCT cell lines are directly related. NT2/D1 cells transduced with Ape1/ref-1 exhibited 2-fold higher AP endonuclease activity in the oligonucleotide cleavage assay, and this was reflected in a 2-3-fold increase in protection against bleomycin. Lesser protection was observed with gamma-irradiation. We conclude that: (a) Ape1/ref-1 is expressed at relatively high levels in some GCTs; (b) elevated expression of Ape1/ref-1 in testicular cancer cell lines results in resistance to certain therapeutic agents; and (c) Ape1/ref-1 expression in GCT cell lines determined by immunohistochemistry and repair activity assays parallels the level of protection from bleomycin. We further hypothesize that elevated Ape1/ref-1 levels observed in human testicular cancer may be related to their relative resistance to therapy and may serve as a diagnostic marker for refractory disease. To our knowledge, this is the first example of overexpressing Ape1/ref-1 in a mammalian system resulting in enhanced protection to DNA-damaging agents. PMID- 11280791 TI - Clinicopathological significance of core 2 beta1,6-N acetylglucosaminyltransferase messenger RNA expressed in the pulmonary adenocarcinoma determined by in situ hybridization. AB - Cell surface carbohydrates of epithelial cells play important roles in tumor progression. Previously, we have shown that expression of core 2 branched O glycans in colorectal cancer is closely correlated with the vessel invasion and depth of invasion (K. Shimodaira et al., Cancer Res., 57: 5201-5206, 1997). To test whether this is also the case in human lung cancer, we have examined the expression pattern of core 2 beta1,6-N-acetylglucosaminyltransferase (C2GnT) mRNA responsible for the biosynthesis of core 2 branched O-glycans in 41 cases of lung cancer. Using in situ hybridization, C2GnT mRNA was detected in 73.2% of the lung cancer cells, irrespective of the histopathological type; whereas in normal lung tissues, its expression was restricted to the basal cells of bronchial mucosa. These results indicate that the expression level of C2GnT mRNA was significantly enhanced in association with malignant transformation. Statistical analysis between the C2GnT mRNA expressed in pulmonary adenocarcinoma and clinicopathological variables revealed that the expression of C2GnT was correlated with vessel invasion and lymph node metastasis with significant difference (P < 0.05), but expression of sialyl Le(x), which is frequently expressed in the adenocarcinoma, was not significantly correlated with lymph node metastasis. These results indicate that C2GnT mRNA detected by in situ hybridization reflects the malignant potentials of pulmonary adenocarcinoma, because lymph node metastasis is the most affecting factor to the patients' prognosis. PMID- 11280792 TI - Inhibition of angiogenesis and tumor growth by SCH221153, a dual alpha(v)beta3 and alpha(v)beta5 integrin receptor antagonist. AB - New blood vessel formation is essential for tumor growth and metastatic spread. Integrins alpha(v)beta3 and alpha(v)beta5 are arginine-glycine-aspartic acid dependent adhesion receptors that play a critical role in angiogenesis. Hence, selective dual alpha(v)beta3 and alpha(v)beta5 antagonists may represent a novel class of angiogenesis and tumor-growth inhibitors. Here, an arginine-glycine aspartic acid-based peptidomimetic library was screened to identify alpha(v)beta3 antagonists. Selected compounds were then modified to generate potent and selective dual inhibitors of alpha(v)beta3 and alpha(v)beta5 receptors. One of these compounds, SCH 221153, inhibited the binding of echistatin to alpha(v)beta3 (IC50 = 3.2 nM) and alpha(v)beta5 (IC50 = 1.7 nM) with similar potency. Its IC50 values for related alpha(IIb)beta3 and alpha5beta1 receptors were 1294 nM and 421 nM, respectively, indicating that SCH 221153 is highly selective for alpha(v)beta3 and alpha(v)beta5 receptors. In cell-based assays, SCH 221153 inhibited the binding of echistatin to alpha(v)beta3- and alpha(v)beta5 expressing 293 cells and blocked the adhesion of endothelial cells to immobilized vitronectin and fibroblast growth factor 2 (FGF2). SCH 221153, but not the inactive analogue SCH 216687, was effective in inhibiting FGF2 and vascular endothelial growth factor-induced endothelial cell proliferation in vitro with an IC50 equal to 3-10 microM. Angiogenesis induced by FGF2 in the chick chorioallantoic membrane assay was also inhibited by SCH 221153. Finally, SCH 221153 exerted a significant inhibition on tumor growth induced by intradermal or s.c. injection of human melanoma LOX cells in severe combined immunodeficient mice. PMID- 11280793 TI - A probasin-large T antigen transgenic mouse line develops prostate adenocarcinoma and neuroendocrine carcinoma with metastatic potential. AB - Neuroendocrine (NE) cells may be involved not only in growth and differentiation of the normal prostate but also in carcinogenesis and progression of prostate adenocarcinoma (Pca), including development of androgen resistance. However, the exact pathophysiology of NE cells in Pca remains poorly understood. Here we describe a transgenic model of Pca with progressive NE differentiation. Seven lines of transgenic mice with the rat prostate-specific large probasin promoter linked to the SV40-large T antigen (Tag) that develop prostatic neoplasia have been established. In this study, one of the seven lines (12T-10) was characterized by examination of 52 mice aged from 2-12 months. With advancing age, low-grade prostatic intraepithelial neoplasia, high-grade prostatic intraepithelial neoplasia, microinvasion, invasive carcinoma, and poorly or undifferentiated carcinoma with NE differentiation appeared in the prostates in sequential order. Whereas Tag is expressed uniformly in prostate epithelium, only an increasing subset of cells in prostatic intraepithelial neoplasia showed NE differentiation by chromogranin immunostaining. Frankly invasive carcinoma developing subsequently showed occasional definitive glandular differentiation (adenocarcinoma) and particularly undifferentiated carcinoma with NE histological features similar to those observed in NE carcinomas in humans. The NE carcinomas occurred in the dorsolateral and ventral lobes and were generally androgen receptor negative. Twenty-one of 32 (66%) mice aged > or = 6 months and 15 of 17 (88%) mice aged > or = 9 months developed metastatic tumors, as confirmed by histology and/or Tag immunohistochemistry. Metastases occurred at the later time points, with metastasis to regional lymph nodes, liver, and lung being particularly common. Metastases showed histological features of NE differentiation, as confirmed by chromogranin immunostaining and electron microscopy. An athymic nude mouse that received a s.c. implant of a primary NE tumor developed Tag-positive metastatic tumors with similar NE differentiation. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry identified identical protein profiles between the primary NE tumor and lesions in the extraprostatic organs. Hence, in the 12T-10 large probasin promoter-Tag mouse, high-grade prostatic intraepithelial neoplasia develops progressively greater NE differentiation and progresses to invasive adenocarcinoma and NE carcinoma, with a high percentage of metastases. The predictable progression through these stages will allow testing of therapeutic interventions as well as possible further delineation of the role of NE cells in Pca progression. PMID- 11280794 TI - Tumor necrosis factor alpha and interleukin 11 secreted by malignant breast epithelial cells inhibit adipocyte differentiation by selectively down-regulating CCAAT/enhancer binding protein alpha and peroxisome proliferator-activated receptor gamma: mechanism of desmoplastic reaction. AB - The dense layer of fibroblasts that accumulate around malignant breast epithelial cells (i.e., desmoplastic reaction) arises from the breast adipose tissue and provides structural and biochemical support for breast cancer. We report herein a number of epithelial-stromal interactions responsible for desmoplastic reaction in breast cancer using cultured 3T3-L1 murine fibroblasts and human adipose fibroblasts, which can be activated with a mixture of hormones to differentiate to mature adipocytes. Adipocyte differentiation was inhibited by coculturing fibroblasts with various breast cancer cell lines (T47D, MCF-7, SSC202, SSC78, and SSC30) completely or by breast cancer cell conditioned media in a dose dependent manner; on the other hand, adipocyte differentiation was not inhibited by coculturing with normal human primary mammary epithelial cell conditioned medium. This tumor effect was eliminated using neutralizing antibodies against tumor necrosis factor (TNF)-alpha or interleukin (IL)-11. TNF-alpha and IL-11 levels were 2.5-3 times higher in T47D conditioned medium compared with control medium, and TNF-alpha transcripts were detectable in T47D but not in 3T3-L1 cells in culture, indicating that the malignant epithelial cell is the major site of cytokine production. This was confirmed in vivo in mastectomy specimens, where immunoreactive TNF-alpha and IL-11 were readily detectable in malignant epithelial cells but not in the majority of the surrounding fibroblasts. Adipocyte differentiation is mediated by the expression of a cascade of adipogenic transcription factors, including CCAAT/enhancer binding protein (C/EBP)beta, C/EBPdelta, peroxisome proliferator-activated receptor (PPAR)gamma and C/EBPalpha. C/EBPalpha and PPARgamma are essential for this process. We demonstrated by Northern analysis that exposure of activated 3T3-L1 cells to T47D cell conditioned medium strikingly decreased the levels of PPARgamma and C/EBPalpha transcripts and increased the levels of C/EBPbeta and C/EBPdelta transcripts. In these 3T3-L1 cells, inhibition of differentiation was also confirmed by markedly suppressed levels of aP2 mRNA, which is an adipocyte specific gene. These in vitro observations were confirmed in sections of human malignant breast tumors, where immunoreactive C/EBPalpha was readily detectable in adipose flbroblasts distant to the tumor but not in intratumoral fibroblasts. Treatment of 3T3-L1 cells with T47D cell conditioned medium or TNF-alpha changed neither the numbers of cells in G0-G1, S, and G2 phases nor the rate of [3H]thymidine incorporation, thus ruling out a proliferative effect of malignant cells on the surrounding fibroblasts. In summary, desmoplastic reaction primarily occurs via the action of cytokines (TNF-alpha and IL-11) secreted by the malignant epithelial cells to inhibit differentiation of adipose fibroblasts to mature adipocytes. This tumor-induced block in adipocyte differentiation is mediated by the selective inhibition of expression of the essential adipogenic transcription factors, i.e., PPARgamma and C/EBPalpha. PMID- 11280795 TI - The expression of vascular endothelial growth factor correlates with mutant p53 and poor prognosis in human breast cancer. AB - Wild-type p53 protein has been shown to inhibit angiogenesis through thrombospondin in the preclinical setting. Here, we determined the associations between the expression of the angiogenic factor vascular endothelial growth factor (VEGF) and the p53 status, including different mutation sites and types, in primary breast cancer. Cytosols from 224 primary breast cancer patients were analyzed with an enzyme immunoassay for determination of human VEGF165 protein content. p53 status was determined by cDNA-based sequencing of the entire coding region, by immunohistochemistry (IHC), and by a p53 luminometric immunoassay (LIA) method. Statistically significant associations was found between higher VEGF content and non-wild-type p53 status for all methods; sequence-based data (P = 0.0019), IHC data (P = 0.0068), and the LIA method (r = 0.427; P > 0.001). Highest VEGF values were detected in tumors with p53 insertions, deletions, and stop codon mutations (P = 0.0043). Combining p53 status and VEGF content resulted in additional prognostic information, relapse-free survival (RFS; P = 0.0377), overall survival (OS; P = 0.0319), and breast cancer corrected survival (BCCS; P = 0.0292). In multivariate analysis, the relative hazard increased when the VEGF data were added to the p53 status, with a relative hazard of 1.7 for RFS and 3.0 for BCCS, compared with 1.1 for RFS and 1.4 for BCCS among the patients with either high VEGF content or p53 mutation. Higher VEGF content was statistically significantly correlated with a worse outcome for patients with estrogen receptor positive tumors receiving adjuvant tamoxifen: RFS (P = 0.0471), OS (P = 0.0134), BCCS (P = 0.0064), as well as in multivariate analysis with point estimates of 3.4 and 2.1 for BCCS and RFS, respectively. VEGF expression is related to p53 status in human breast cancer patients. Combining VEGF with p53 status resulted in better prognostic prediction. PMID- 11280796 TI - Impaired alpha-interferon signaling in transitional cell carcinoma: lack of p48 expression in 5637 cells. AB - The limited success of IFN-alpha therapy for clinical treatment of transitional cell carcinoma (TCC) has prompted us to investigate the responsiveness of TCC lines to IFN-alpha. The response to IFN-alpha in terms of 561 gene induction, an IFN-stimulated response element-containing IFN-alpha/beta-inducible gene, and IFN stimulated gene factor 3 (ISGF3) formation was normal in primary human urothelial cells. We tested the antiproliferative effects of IFN-alpha in three TCC lines as a measure of IFN-alpha responsiveness, and variable patterns of growth inhibition were observed in three TCC lines. More than 90% growth inhibition was noted in TCCSUP cells, whereas only 40% and 10% inhibition by IFN-alpha was observed in 5637 and HT1197 cells, respectively. IFN-alpha treatment formed extremely low levels of ISGF3 in electrophoretic mobility shift assays in these later two relatively insensitive cells. In addition, expression of the 561 gene was significantly reduced in these two TCC lines by Northern blots. We have further identified a low expression level of Tyk2 in HT1197 cells compared with two other TCCs. This suggests that an extremely low ISGF3 level after IFN-alpha treatment may be due to low Tyk2 expression or other unidentified defects. In 5637 cells, p48 protein expression was undetectable. This undetectable p48 expression is not due to a deletion in the coding region because the correct size protein is detected following IFN-gamma treatment. Consequently, the ISGF3 complex formation and 561 gene induction were restored by IFN-gamma pretreatment plus IFN-alpha treatment. Introduction of p48 expressing plasmid into 5637 cells was sufficient to form the ISGF3 complex by IFN-alpha treatment, suggesting the defect lies in the expression of p48 protein in 5637 cells. Detailed mechanistic understanding of the action of IFNs in bladder cancer cell lines may explain the abrogated therapeutic response of IFN-alpha in the clinical treatment of TCCs. PMID- 11280797 TI - Induction of ETS-1 and ETS-2 transcription factors is required for thyroid cell transformation. AB - The proteins of the Ets family are transcription factors involved in signal transduction, cell cycle progression, and differentiation. In this study, we report that thyroid cell neoplastic transformation is associated with a dramatic increase in ETS transcriptional activity, which is dependent on the accumulation of Ets-1, Ets-2, and other Ets-related proteins. Inhibition of ETS transactivation activity by the Ets-dominant negative construct (Ets-Z) induced programmed cell death in human thyroid carcinoma cell lines but not in normal thyroid cells. Apoptotic cell death induced by Ets-Z was dependent on the reduction of c-MYC protein levels, because it was prevented by overexpression of c-myc. Taken together, these data indicate that the induction of Ets-1 and Ets-2 transcription factors plays a pivotal role in thyroid cell neoplastic transformation. PMID- 11280798 TI - Regulation of MMP-1 and MMP-2 production through CD147/extracellular matrix metalloproteinase inducer interactions. AB - Extracellular matrix metalloproteinase inducer (EMMPRIN; CD147) is a heavily glycosylated protein containing two immunoglobulin superfamily domains. It is enriched on the surface of tumor cells and stimulates the production of matrix metalloproteinases (MMPs) by adjacent stromal cells. Here we use CD147 transfectants and immobilized recombinant CD147-Fc fusion protein to show that CD147/FMMPRIN engages in a homophilic interaction, predominantly through the first immunoglobulin domain. Anti-CD147 antibody 8G6 and recombinant CD147-Fc fusion protein markedly inhibited not only homophilic interaction, but also the production of secreted MMP-2 by breast cancer cell line MDA-435 and the MMP-2 dependent invasion of MDA-435 cells through reconstituted basement-membrane Matrigel. Purified native CD147 induced the production of secreted MMP not only by dermal fibroblasts (MMP-1) but also by MDA-435 cells themselves (MMP-2), suggesting homophilic CD147-binding may occur in the context of both heterotypic and homotypic cell-cell interactions. Purified deglycosylated CD147 failed to induce MMP-1 or MMP-2, but instead antagonized the MMP-1-inducing activity of purified native CD147. Our results suggest that homophilic CD147 interactions may play a key role in MMP-2 production and tumor cell invasion, and that perturbation of this molecule may have potential therapeutic uses in the prevention of MMP-2 and MMP-1-dependent cancer metastasis. PMID- 11280799 TI - Parathyroid hormone-related protein induces interleukin 8 production by prostate cancer cells via a novel intracrine mechanism not mediated by its classical nuclear localization sequence. AB - PTHrP (parathyroid hormone-related protein) overexpression by prostate carcinoma cells has been implicated in tumor progression. Although the biological effects of PTHrP can be mediated by the G-protein-coupled PTH/PTHrP receptor, PTHrP also has intracrine actions mediated by a nuclear localization sequence at residues 87 107. We investigated the effect of PTHrP transfection and treatment on production by prostate carcinoma cells of IL (interleukin)-8, which can regulate prostate cancer growth by angiogenic activity and growth-promoting effects. Six prostate cancer cell lines exhibited constitutive expression of PTHrP and IL-8 that were significantly correlated (r = 0.93; P < 0.01). We transfected wild-type and mutant PTHrP into these cells. Wild-type PTHrP1-173 and PTHrP33-173 lacking the PTH/PTHrP receptor-binding domain induced a 3-fold stimulation of IL-8 production but not production of another angiogenic factor, vascular endothelial growth factor. Transfection of the COOH-terminal truncation mutant PTHrP1-87 induced a 5 fold simulation of IL-8 and a 3-fold increase in IL-8 mRNA. Cells transfected with PTHrP1-87 and 1-173 also showed increased cell proliferation. In contrast, exogenous PTHrP1-34 and 1-86 peptides did not significantly affect IL-8 production; moreover, PTHrP-neutralizing antibodies did not inhibit the production of IL-8 by transfected PTHrP. Additional transfection studies with progressively COOH-terminally truncated PTHrP1-87 defined a 23-amino acid sequence, PTHrP65-87, required for PTHrP1-87 to robustly stimulate IL-8 in prostate cancer cells. Confocal microscopy and immunoassay demonstrated PTHrP1-87 nuclear localization. Our results demonstrate that PTHrP acts to induce IL-8 production in prostate cancer cells via an intracrine pathway independent of its classical nuclear localization sequence. This novel pathway could mediate the effects of PTHrP on the progression of prostate cancer. PMID- 11280800 TI - The farnesyltransferase inhibitor L744,832 reduces hypoxia in tumors expressing activated H-ras. AB - Many tumors contain extensive regions of hypoxia. Because hypoxic cells are markedly more resistant to killing by radiation, repeated attempts have been made to improve the oxygenation of tumors to enhance radiotherapy. We have studied the oxygenation of tumor xenografts in nude mice after treatment with the farnesyltransferase inhibitor L744,832. Hypoxia was assessed by measuring the binding of the hypoxic cell marker pentafluorinated 2-nitroimidazole. We show that xenografts from two tumor cell lines with mutations in H-ras had markedly improved oxygenation after farnesyltransferase treatment. In contrast, xenografts from two tumors without ras mutations had equivalent hypoxia regardless of treatment. The effect on tumor oxygenation could be detected at 3 days and remained after 7 days of treatment. These results indicate that treatment with farnesyltransferase inhibitors can alter the oxygenation of certain tumors and suggest that such treatment might be useful in the radiosensitization of these tumors. PMID- 11280801 TI - Cystemustine induces redifferentiation of primary tumors and confers protection against secondary tumor growth in a melanoma murine model. AB - N'-(2-Chloroethyl)-N-(2-(methylsulfonyl)-ethyl)-N'-nitrosourea (cystemustine) is a chloroethylnitrosourea that has been used in the treatment of human melanoma. Its main antitumor effect is DNA damage to malignant melanocytes. Although unreported at present, other effects may also account for its cytotoxicity, some of them could be more or less delayed with respect to its administration. In this report, we have developed a model of secondary tumor with B16 melanoma in syngeneic C57B16 recipients to investigate the impact of cystemustine treatment of primary B16 melanoma tumors on the fate of secondary implanted untreated tumors. The data presented in this report indicate that cystemustine-treated cells or the administration of cystemustine provoke an important growth delay of primary melanoma tumors, together with an increase in cell pigmentation and cell morphology changes. Data also show that prime treatment induces a dramatic decrease in tumor weight of secondary untreated tumors accompanied by an increase in melanin content and an alteration of cell morphology. Finally, 1H-NMR spectroscopy was performed on treated B16 cells, showing an alteration in the phospholipid derivatives of melanocytes, suggesting subsequent modifications of membrane phospholipid composition. In conclusion, the data highlight two important findings: (a) cystemustine produces modifications other than DNA damage, i.e., cell morphology changes, pigmentation, and phospholipid metabolism alterations, indicating an interference with cell cycle, cell redifferentiation, and proliferation programs; and (b) cystemustine-treated tumors appear to confer a protective effect against the development of secondary untreated tumors that may be mediated by cytokines or an immune response. PMID- 11280802 TI - EphA2 overexpression causes tumorigenesis of mammary epithelial cells. AB - Elevated levels of protein tyrosine phosphorylation contribute to a malignant phenotype, although the tyrosine kinases that are responsible for this signaling remain largely unknown. Here we report increased levels of the EphA2 (ECK) protein tyrosine kinase in clinical specimens and cell models of breast cancer. We also show that EphA2 overexpression is sufficient to confer malignant transformation and tumorigenic potential on nontransformed (MCF-10A) mammary epithelial cells. The transforming capacity of EphA2 is related to the failure of EphA2 to interact with its cell-attached ligands. Interestingly, stimulation of EphA2 reverses the malignant growth and invasiveness of EphA2-transformed cells. Taken together, these results identify EphA2 as a powerful oncoprotein in breast cancer. PMID- 11280804 TI - Up- and down-regulation of granulocyte/macrophage-colony stimulating factor activity in murine skin increase susceptibility to skin carcinogenesis by independent mechanisms. AB - The role of granulocyte-macrophage colony-stimulating factor (GM-CSF) in tumorigenesis is complex. On the one hand, GM-CSF can promote tumor cell growth, survival, and even metastasis. On the other hand, it can stimulate tumor cell rejection. In skin, it is early expressed after topic application of tumor promoting agents and therefore may be responsible for changes that correlate with skin tumor promotion (e.g., epidermal hyperproliferation and inflammation). To analyze GM-CSF function in skin tumorigenesis, we generated transgenic mice epidermally overexpressing either GM-CSF or a GM-CSF antagonist. Both types of transgenic mice exhibited significantly increased numbers of benign tumors in a two-step skin carcinogenesis experiment using 7',12'-dimethylbenz[a]anthracene (DMBA) as initiator and 12-O-tetradecanoylphorbol-CSF displayed a significantly elevated carcinoma burden following a single-step carcinogenesis protocol consisting of tumor initiation only. Therefore, endogenous promotion is responsible for elevated tumor development in GM-CSF-overexpressing mice. In antagonist transgenic animals, an increased tumorigenicity of modified B16 tumor cells after cutaneous transplantation as compared with nontransgenic or GM-CSF transgenic mice was observed. Thus, the antitumor activity leading to the repression of tumor cell growth in control mice is GM-CSF dependent and is compromised in mice expressing the antagonist. We suggest that both, up regulation and down-regulation of GM-CSF activity in skin, increase the incidence and growth of tumors via two independent mechanisms: endogenous tumor promotion in the case of increased GM-CSF activity and compromised tumor cell rejection in the case of decreased GM-CSF activity. PMID- 11280803 TI - Distribution and functional consequences of nucleotide polymorphisms in the 3' untranslated region of the human Sep15 gene. AB - Selenium has been shown to prevent cancer in a variety of animal model systems. Both epidemiological studies and supplementation trials have supported its efficacy in humans. However, the mechanism by which selenium suppresses tumor development remains unknown. Selenium is present in known human selenoproteins as the amino acid selenocysteine (Sec). Sec is inserted cotranslationally in response to UGA codons within selenoprotein mRNAs in a process requiring a sequence within the 3'-untranslated region (UTR), referred to as a Sec insertion sequence (SECIS) element. Recently, a human Mr 15,000 selenoprotein (Sep15) was identified that contains an in-frame UGA codon and a SECIS element in the 3'-UTR. Examination of the available cDNA sequences for this protein revealed two polymorphisms located at position 811 (C/T) and at position 1125 (G/A) located within the 3'-UTR. Here, we demonstrate significant differences in Sep15 allele frequencies by ethnicity and that the identity of the nucleotides at the polymorphic sites influences SECIS function in a selenium-dependent manner. This, together with genetic data indicating loss of heterozygosity at the Sep15 locus in certain human tumor types, suggests that Sep15 may be involved in cancer development, risk, or both. PMID- 11280805 TI - Elevation of breast carcinoma Nm23-H1 metastasis suppressor gene expression and reduced motility by DNA methylation inhibition. AB - We hypothesize that elevation of Nm23-H1 expression in micrometastatic breast cancer cells may inhibit their metastatic colonization and further invasion, and induce differentiation, thus resulting in a clinical benefit. The current study investigated the possible contribution of DNA methylation to the regulation of Nm23-H1 expression, based on the observation that two CpG islands are present in its promoter. 5-Aza-2'-deoxycytidine (5-Aza-CdR), a DNA methylation inhibitor, increased the Nm23-H1 expression of 5 of 11 human breast carcinoma cell lines in vitro, including 3 of 3 metastatically competent lines. Increased Nm23-H1 expression was accompanied by a reduction in motility in vitro, with minimal effect on proliferation. Both increased Nm23-H1 expression and decreased motility were observed using low (75 nM) concentrations of 5-Aza-CdR. Array analysis of MDA-MB-231 breast carcinoma cells treated with 5-Aza-CdR confirmed the elevation of nm23-H1 mRNA, whereas relatively few other genes exhibited altered expression. Bisulfite sequencing of the two CpG islands in a panel of cell lines and in 20 infiltrating ductal carcinomas revealed that one island (-3090 bp to -3922 bp) exhibited infrequent differential methylation. The data indicate that DNA methylation inhibitors can directly or indirectly cause both elevation of Nm23-H1 expression and decreased function in one aspect of metastasis, motility. PMID- 11280806 TI - Malignant breast epithelial cells stimulate aromatase expression via promoter II in human adipose fibroblasts: an epithelial-stromal interaction in breast tumors mediated by CCAAT/enhancer binding protein beta. AB - Expression of aromatase P450 (P450arom), which catalyzes the formation of estrogens, is aberrantly increased in adipose fibroblasts surrounding breast carcinomas, giving rise to proliferation of malignant cells. Aromatase in human adipose tissue is primarily expressed in undifferentiated fibroblasts under the control of several distinct and alternatively used P450arom promoters. In tumor free breast adipose tissue, P450arom is usually expressed at low levels via a distal promoter (I.4), whereas in the breast adipose tissue bearing a tumor, P450arom is increased through the activation of two proximal promoters, II and I.3. Because the in vivo activation of P450arom promoter II is a key event responsible for aberrantly high P450arom expression in breast tumors, we studied the molecular basis for the enhancement of P450arom promoter II using human adipose fibroblasts (HAFs) in primary culture treated with T47D breast cancer cell-conditioned medium (TCM) as a model system. Upon treatment with TCM, HAFs displayed a striking induction of P450arom mRNA levels via promoter II usage. This effect appeared to be specific for malignant breast epithelial cells, because conditioned media from breast cancer cell lines T47D and MCF-7 induced promoter II activity, whereas normal breast epithelial cells or liver or prostate cancer cell lines did not produce such an effect. Although treatment with a cyclic AMP analogue also caused a switch in the promoter use from I.4 to II in cultured HAFs, TCM-induced promoter II use was found to be mediated via a cyclic AMP-independent pathway. Use of serial deletion mutants of the promoter II 5' flanking sequence revealed the presence of critical cis-acting elements in the 517/-278 bp region, which regulate the baseline activity. TCM caused a 5.7-fold induction of the -517-bp promoter II construct, whereas site-directed mutagenesis of a CCAAT/enhancer binding protein (C/EBP) binding site (-317/-304 bp) abolished both baseline and TCM-induced activities. Ectopic expressions of C/EBPalpha and C/EBPbeta, but not C/EBPdelta, significantly induced promoter II activity. Moreover, we demonstrated the presence of both C/EBPbeta and C/EBPdelta but not C/EBPalpha in a DNA-protein complex formed by the nuclear extract from TCM treated HAFs and a probe containing this critical C/EBP binding element (-317/ 304 bp). Finally, treatment of HAFs with TCM strikingly induced C/EBPbeta expression, whereas this did not affect the levels of C/EBPalpha or C/EBPdelta transcripts. In conclusion, malignant breast epithelial cells secrete factors, which induce aromatase expression in adipose fibroblasts via promoter II. This is, at least in part, mediated by a TCM-induced up-regulation and enhanced binding of C/EBPbeta to a promoter II regulatory element. PMID- 11280807 TI - Correspondence re: O. J. Arola et al., acute doxorubicin cardiotoxicity involves cardiomyocyte apoptosis. Cancer Res., 60: 1789-1792, 2000. PMID- 11280808 TI - HIV sinusitis: rationale for a treatment algorithm. AB - Over 12 million individuals worldwide are infected with the human immunodeficiency virus (HIV). Up to 60% of these persons may suffer from disease of the paranasal sinuses. Numerous differences exist between sinusitis in HIV and non-HIV infected patients. Some of the differences include HIV sinusitis pathogenesis, bacteriology, and management. This paper addresses these issues so that physicians may adequately prevent, diagnose, and treat persons suffering from HIV sinus infection. PMID- 11280809 TI - Radiology case of the month. Tearing pain and swelling in the left chest wall. Grade III muscle strain of the pectoralis major and minor. PMID- 11280810 TI - The journal 150 years ago. January 1849. PMID- 11280811 TI - Surgical management of the paralyzed eye. AB - Blindness is the most dreaded complication of an untreated paralyzed eyelid following injury to the facial nerve. Injuries to the facial nerve are mainly postsurgical. Assessment of neural injury using serial testing is important to be able to differentiate between temporary and permanent paralysis. In the former case, medical management could be sufficient and, in the latter case, a surgical procedure is required. The physician has the choice among several procedures to repair upper eyelid paralysis with the gold standard being the gold weight implant. Lower eyelid ectropion can be repaired using a lateral or medial canthal tightening procedure. If the lower eyelid ectropion is severe, a cartilage implant may be required. PMID- 11280812 TI - Plasmodium falciparum malaria. AB - A 13-year-old adolescent daughter of a missionary presented with fever and jaundice 1 week after returning from Africa. Examination of peripheral blood film revealed the diagnosis of Plasmodium falciparum infection. Therapy with oral quinine and doxycycline was curative. Diagnosis requires a travel history and a high index of suspicion. Because of the frequency of international travel, United States physicians need to be familiar with the presentation and management of imported P falciparum. Preparation for such travel must include careful counseling and optimal use of chemoprophylaxis. PMID- 11280813 TI - Soft tissue sarcoma. AB - Soft tissue sarcomas comprise approximately 1% of all adult malignancies. These mesenchymal neoplasms require carefully planned diagnostic and therapeutic strategies to optimize postextirpative functional preservation. Soft tissue sarcomas most frequently occur in the lower extremity and typically require a multidisciplinary team approach with surgical resection, radiation therapy, and possibly chemotherapy for limb salvage. PMID- 11280814 TI - Mortality experience for Louisiana chemical workers. AB - A previous mortality study for Louisiana workers of The Dow Chemical Company identified greater than expected deaths due to leukemia and aleukemia and cancer of the brain and central nervous system. The current study updated the mortality experience through 1992. No new deaths due to leukemia and aleukemia and only one new death due to cancer of the brain and central nervous system were observed. No causes of death were statistically significantly elevated. The number of observed deaths was statistically significantly less than expected for a number of disease categories. Mortality rates were not shown to be higher among Louisiana chemical employees when compared to the United States, state, and regional populations. PMID- 11280815 TI - Are our children less deserving of the freedoms we've enjoyed? PMID- 11280816 TI - ECG of the month. Dual duel? Atrial rhythm. PMID- 11280817 TI - [Surgical treatment of ulcerative rectocolitis: technical aspects of total restorative proctocolitis with S pouch]. AB - Restorative proctocolectomy with an ileal-pouch-anal anastomosis seems to be the treatment of choice for ulcerative colitis. The aim of this study was to discuss a number of technical and functional aspects of the procedure that still appear to be controversial such as the shape of the pouch, the mucosectomy and the type of anastomosis. The authors report on their experience with the surgical treatment of ulcerative colitis with an "S" pouch. The technique reported, however, differs from the original method proposed by Parks and Nicholls in 1978 and the reasons for this surgical choice are discussed. A six-year experience (1993-1999) regarding 35 patients undergoing this approach is reported. No perioperative deaths were observed. The early and long-term complication rates were 8.5% and 11.4%, respectively. The average number of daily evacuations was 4. Mucosectomy affords complete resolution of the disease, while the particular shape of the pouch guarantees good functional results. PMID- 11280818 TI - [Postoperative pain in inguinal hernioplasty: does it exist?]. AB - Tension-free hernioplasty in the Day Hospital or Short Stay Surgery setting is now considered the gold standard for hernia treatment, but early discharge is not synonymous with a rapid recovery, and pain in the inguinal region may persist for months after the operation. The authors examined 100 tension-free hernioplasties, performed in 1999. 72 Lichtenstein and 28 Trabucco hernioplasties were performed in 97 men and 3 women (min age 18 yrs., max. 90 yrs., mean 55 yrs.). Three months after hernioplasty residual inguinal pain was present in 10 patients (mild in 9, moderate in 1). Hypo-dysaesthesia in the inguino-crural region was also present in 8 patients (mild in 7, moderate in 1). Operative technique, emergency surgery and the surgeon himself were not correlated with results; occupational status was significantly associated with residual pain, which affected active workers more than retired patients. PMID- 11280819 TI - Long-term results after a low anterior resection with mucosectomy and colo-anal sleeve anastomosis for a diffuse cavernous haemangioma of the rectum. AB - Diffuse cavernous hemangioma of the rectum is an unusual lesion. We reporting the case of an 18-year-old man with a rectal cavernous hemangioma in whom recurrent rectal bleeding and marked anemia were thought to be caused by his co-existing internal hemorrhoids. This resulted in a 2-year delay in reaching the correct diagnosis. Digital rectal examination revealed a walnut-sized, wide-based, elastic, soft mass 3 cm proximal to the anal verge. Colonoscopy revealed a bluish, submucosal lesion with superficial capillary dilatation at the same site. Arteriography demonstrated vascular tumours in the territory of the right hypogastric artery and the superior rectal artery. In 1972, Parks and co-workers described resection and colo-anal sleeve anastomosis as an alternative operation in the treatment of this rare malformation. We will describe the clinical presentation, diagnosis, and long-term results in a patient with this condition managed with this surgical technique. The patient has done well without any recurrence of rectal bleeding for over 10 years since his operation. Resection with a colo-anal sleeve anastomosis offers major advantages such as a lower risk of intraoperative bleeding, no risk of damaging the pelvic nerves, sparing of continence and avoidance of a permanent colostomy. It should therefore be considered the treatment of choice for this uncommon condition. PMID- 11280820 TI - [Middle lobectomy for bronchiectasis: clinical case and review of the literature]. AB - Over the past few years bronchiectasis, among the chronic lung diseases, has been the second most important after tuberculosis in terms of frequency and mortality. Although the incidence of the disease has been decreasing in recent years, the illness is currently of great surgical interest because of an upsurge of cases among people considered to be below the bread line. The authors present the clinical case and surgical treatment of a young adult with middle lobe bronchiectasis, with a 10-year primary IgG deficiency and severe bronchopneumonia requiring hospitalisation. Medical treatment, long regarded as the treatment of choice in this condition, has reduced the short-term morbidity of patients suffering from the disease, without affecting its ultimate mortality which is still very high today. The policy in the past to reserve surgery only for the most complicated cases or for patients not responding to medical treatment can now be considered obsolete, due to the reduced surgical risks (less than 1%) and to faster patient recovery. Further surgical indications are mono- or bilaterally located forms of the disease and failure to respond to medical treatment for more than 2 years. A review of the literature enables the authors to affirm that in the absence of randomised trials on the effectiveness of surgical vs medical treatment, it seems clear that surgical therapy is the best option, being curative and safe, with a high percentage of complete remission of disease and very low operative risks and mortality. It can therefore guarantee good quality of life, radically changing the prognosis which otherwise is fatal in 1/3 of patients suffering from this orphan disease. PMID- 11280821 TI - [Intraductal mucinous tumor of the pancreas: report of a clinical case]. AB - A 49-year-old diabetic patient with abdominal pain was found at ultrasonography and computed tomography to have a cystic mass in the head of the pancreas with dilatation of the main pancreatic duct. The head of the pancreas and the duodenum were removed surgically. Examination of the operative specimen showed chronic pancreatitis, dilatation of the main pancreatic duct, and impacted mucus in the secondary ducts with villous proliferation of the ductal epithelium, thus allowing a diagnosis of intraductal adenomatosis. There was no evidence of malignancy. The resection margin was involved, and consequently the remainder of the pancreas was removed six months after the initial surgical procedure. A review of the literature showed that intraductal adenomatosis tends to spread and carries a high risk of malignant transformation. Surgery is required because of the risk of pancreatic duct obstruction and pancreatic cancer. Intraductal papillary tumour of the pancreas shares many characteristic with other adenomatous proliferation of the gastrointestinal tract (colorectal villous adenoma, bile duct adenomatosis) including the presence of villous structures with increased mucus production, a tendency to spread massively, and a high risk of malignant transformation. PMID- 11280822 TI - [Multiple primary malignant neoplasms. Report of a rare case with 5 metachronous tumors]. AB - The Authors report on a patient admitted several times for the occurrence of five multiple metachronous primary malignancies (laryngeal carcinoma, endometrial adenocarcinoma, rectal cancerous polyp, ampulla of Vater carcinoma and transverse colon cancer). All five carcinomas were independent primary cancers, and the lengthy time intervals between the onsets of the individual tumours supports their independent non-metastatic origin. Classification, pathogenesis, genetic and environmental interactions of multiple tumours are discussed. In the case reported, a family history of colon cancer was present, while no genetic marker abnormalities or chronic exposure to carcinogens were found. The case report shows that an aggressive, appropriate surgical approach together with thorough follow-up monitoring offers a chance of long-term survival for patients with metachronous malignant primary tumours. PMID- 11280823 TI - [Secondary hyperparathyroidism]. AB - We reviewed the results of twenty-four years' experience of parathyroidectomy for secondary hyperparathyroidism. Over the period from 1976 to 2000, we performed 171 parathyroidectomies for secondary hyperparathyroidism (91 males, 80 females; median age: 57.4). Autotransplantation was performed in 45. Of the 171 patients undergoing parathyroidectomy, 14 (8.1%) were treated for relapse. Twenty-four of the 45 patients treated with autotransplantation (53.3%) were followed up for 82 +/- 37 months. Hypoparathyroidism was observed in 33.3% of these (worse after 6 months in 21% and stable in 46%). Five patients (11.1%) experienced a relapse of hyperparathyroidism. We believe that the treatment of choice for secondary hyperparathyroidism is subtotal parathyroidectomy, the indications for autotransplantation in parathyroid surgery are rare. PMID- 11280824 TI - [Radioimmuno-guided surgery (RIGS) in breast disease]. AB - Radioimmunoguided surgery is a new technology capable of detecting minimal neoplastic lesions using radiocolloids. We used this technique in two fields: to detect sentinel lymph nodes in breast cancer and to remove non-palpable breast lesions. Radioimmunoguided surgery was employed in 135 women; in 32 for sentinel lymph nodes and in 103 for radioguided occult lesion localization using a radioactive tracer (Technetium Tc99m) injected subdermally for sentinel nodes, or near to the non-palpable lesions under US guidance. In our experience these two applications of radioimmunoguided surgery are useful and accurate for determining the nature of lesions and for providing definitive treatment in a single surgical intervention. PMID- 11280825 TI - [Does locoregional chemotherapy improve survival in patients with non-resectable pancreatic carcinoma? Results of an open controlled study]. AB - About 90% of patients suffering from pancreatic carcinoma are diagnosed with disease that is not amenable to surgical intervention due to local infiltration or the presence of hepatic metastases. Palliative intra-arterial chemotherapy was developed to improve the response in these patients by increasing the antiblastic dose and minimizing the side effects. The aim of this study is to evaluate the efficacy of this treatment comparison to a control group. From December 1994 to February 1997, 135 patients with ductal carcinoma, in whom 68 were stage III and 67 stage IV, with a median age of 63.3 years (range 38.4-79), were enrolled in an open study. Sixty four patients were subjected to a median of 3.5 cycles, according to intra-arterial FLEC protocol. Four patients had a partial response (6.3%), 27 enjoyed a stabilization of their disease (42.2%) and 13 showed disease progression (20.3%). The toxicity was mild. The overall survival was 8.3 months, better in the treated group (9.6 months) in respect to the control one (7.1 months), although this was not statistically significant. The treatment reported here, therefore, does not seem to change the prognosis of patients affected by no resectable pancreatic carcinoma, but it may demonstrate good tolerability and minimal toxicity. PMID- 11280826 TI - [Prognostic value of C reactive protein in acute pancreatitis]. AB - Controversy still exists regarding the clinical features of acute pancreatitis: it is not known whether this is a disease which progresses from mild to severe forms or which arises immediately as severe acute pancreatitis. An early diagnosis, however, is regarded as mandatory for successful treatment. Over the years many Authors have proposed different scoring systems for the early assessment of the clinical evolution of acute pancreatitis. The most widely used scoring systems (Ranson, Osborne, Apache II) are often cumbersome and difficult to use in clinical practice because of their multifactorial nature. Thus, a number of unifactorial prognostic indices have been employed in routine hospital practice, such as C-reactive protein, serum amylase and serum lipase. These serum enzymes are easy to obtain in normal clinical practice and many authors consider them as reliable as multifactorial scoring systems. One hundred and five patients affected by acute pancreatitis have been hospitalised in the Surgical Department of San Giacomo Hospital (Rome) over an nine-year period. All patients underwent C reactive protein, amylase, and lipase serum assays on days 1, 3 and 5 after admission. The results show that C-reactive protein assay is highly sensitive in detecting necrotic forms of acute pancreatitis. The authors conclude that C reactive protein, together with both serum amylase and serum lipase, often provides a precise picture of the clinical situation in patients with acute pancreatitis. On this basis the best therapeutic option can be chosen. PMID- 11280827 TI - [Effect of probiotic administration on colic anastomosis healing]. AB - Colic anastomoses are still affected by a high incidence of leakage. We speculate that a supply of fibres and probiotic bacteria improves the healing of colic anastomoses due to a higher production of short-chain fatty acids. These are known to improve the anastomotic healing of colic sutures. Sixty Lewis rats, weighing from 250 g to 350 g, were divided into 6 groups. Groups A + A1 were fed with a low-fibre diet (less than 0.1%), Groups B + B1 with normal rat chow and groups C + C1 with normal rat chow + Lactobacillus plantarum 299v. Transections and re-anastomosis of the distal colon were performed. Groups A1, B1 and C1 were sacrificed after 3 days, and groups A, B, and C after 7 days. The bursting pressure of colic anastomoses was measured. All data are expressed as mean (+/- S.D.). The pH of the colon contents was evaluated by means of a fine needle plastic electrode only in groups A1, B1 and C1. The results were studied by analysis of variance followed by the Student Newman Keuls test for multiple comparisons (significance level P < 0.05). Three days postoperatively, the pH of the colic lumen was lower in animals fed with a normal diet (pH 7.1 +/- 0.3 without Lp supplementation, 6.5 +/- 0.2 with Lp supplementation) than in animals fed with a low-fibre diet (pH 8.0 +/- 0.3). Bursting pressures were significantly higher in the groups fed with fibre and fibre + Lactobacilli than in animals on a low-fibre diet, both on day 3 and day 7. On the basis of these data there seems to be no support for the belief that a supply of fibre-rich food might impair healing and promote development of anastomotic leakage. On the contrary, short chain fatty acids and fibres would seem to facilitate the healing of colic anastomoses. PMID- 11280828 TI - [Intestinal infarction. Retrospective clinical study]. AB - Acute mesenteric ischaemia is the result of inadequate blood flow to all or part of the small intestine and the right half of the colon. Irrespective of the cause of the ischaemic insult, the end results are similar, namely, a spectrum of bowel injury ranging from completely reversible alterations of bowel function to transmural haemorrhagic necrosis of the intestinal wall. Depending on the degree of ischaemia and the length of bowel involved, a wide variety of clinical presentations are observed. Mesenteric infarction is a pathology which is encountered fairly often in elderly patients where the concomitance of other diseases make its prognosis more severe, especially since the diagnosis is usually late. The pessimism expressed more than 70 years ago concerning this disease is still shared by many physicians today. The authors report on their experience with 37 cases of mesenteric infarction. The median age of the patients was 77 years (range: 66-91). The mortality rate was 67.5% (25 deaths) which is in line with the results in the literature. The median hospital stay was 17 days (range: 10-71). The authors emphasise the difficulty of diagnosing and treating this entity, also in view of the fact that, in most centres, it is impossible to perform emergency selective angiography of the superior mesenteric artery. The need for an early specific diagnosis is stressed, because the therapeutic options may vary widely in relation to the different causes of acute intestinal ischaemia. PMID- 11280829 TI - Intestinal infarction: report of 98 cases. AB - The Authors conducted a retrospective study on 98 patients with intestinal infarction observed from 1987 to 1999 in the Emergency Care Unit of the Loreto Hospital, Naples. In our hospital there are over 20,000 admissions, 3,900 of whom in the Emergency Care Unit. Intestinal infarction accounts for 0.049% of all admissions and 0.45% of emergency surgery admissions. About 500 laparotomies are performed annually, 1% of which for intestinal infarction. All patients in this series were operated on within 10 hours of admission. The following procedures were performed: 31 jejuno-ileal resections; 26 right hemicolectomies associated with small intestine resection; 5 upper mesenteric artery embolectomies plus wide gut resections (3 also underwent second-look operations within 36 hours of the initial surgery with further gut resection); 1 Hartmann's and 5 Volkmann's operations (all of these patients had colonic gangrene); 30 (30.5%) underwent exploratory laparotomy due to massive infarction. The prognosis of intestinal infarction is still ominous. Our mortality rate is 68%. Both clinical and laboratory data are non-specific and delayed diagnosis is the main cause of this mortality rate. Abdominal CT is an accurate and sensitive diagnostic tool. TPN enables us to achieve good nutritional support even for wider resections. PMID- 11280830 TI - [Gabexate mesilate vs gabexate mesilate combined with octreotide in the prevention of postoperative complications of pancreatic surgery: preliminary results]. AB - To date, gabexate mesilate, a synthetic protease inhibitor, has been used in the prophylaxis and treatment of acute pancreatitis, but has yet to be tested in preventing the postoperative complications of pancreatic surgery. For this purpose we planned a pilot study based on two treatment groups, each numbering 25 patients, submitted to high-risk pancreatic resection. In the first group, all patients received a continuous infusion of gabexate mesilate 1 g/day up to postoperative day 4; the second group of patients received the same treatment plus octreotide 0.1 mg every 8 hours for 5 days after surgery. All patients were followed until discharge with clinical and instrumental investigations to detect the onset of postoperative complications. The overall incidences of an uneventful course were 40% (10/25) and 32% (8/25), respectively. We found 12 complications closely related to pancreatic surgery in the former and 8 in the latter group. In the combined treatment group therefore we observe a 33% reduction in the incidence of related abdominal complications (12 vs 8). This favourable trend, however, needs to be confirmed in a larger multicentre trial. PMID- 11280831 TI - [Complex duodenopancreatic injuries]. AB - Injuries of the duodenopancreatic region are rare and difficult to diagnose and treat. The related high mortality is mainly due to the presence of associated lesions. Complex traumas (AAST grade IV and V lesions) require difficult surgical treatment with high postoperative morbidity and mortality rates. In a review of 200 pancreaticoduodenectomies performed for pancreatic head traumas the postoperative mortality was 31%. The authors present 6 cases of complex duodenopancreatic traumas, treated from 1995 to 1999. The aetiology was blunt trauma in 5 cases (83%) and a shotgun wound in 1 case (17%). In 3 cases, with a grade V lesion of the pancreatic head, a pancreaticoduodenectomy was performed. A case of a grade IV lesion of the tail of the pancreas was treated with distal splenopancreatectomy. Two cases of grade IV lesions of the third part of the duodenum were submitted to duodenal resection with direct anastomosis. One postoperative death was observed in a patient treated with duodenal resection. The overall mortality was 16%. A pancreatic fistula, which healed spontaneously, was observed in a case of pancreaticoduodenectomy. PMID- 11280832 TI - [Cranial and neck nerve injuries following carotid endarterectomy intervention. Review of the literature]. AB - The aim of the study was to establish the operative techniques and findings that can influence the reported incidence of cranial and cervical nerve injuries. Eight main studies comprising 1,616 carotid endarterectomies and published over the period from 1990 to October 2000 were reviewed. There were no statistically significant differences between neck incision (vertical or transverse) and number of injuries. In one study, multiple deficits were observed most frequently in patients treated by the eversion technique (P = 0.2). Additional prospective trials are needed in large numbers of patients to assess the incidence of cranial and cervical nerve injuries. Most injuries are transient and involve the vagus and hypoglossal nerves. A number of factors related to the operation, such as general anaesthesia, eversion technique and the surgeon's experience, may influence the incidence of such injuries. Repeat endarterectomy is associated with a high incidence of cranial and/or cervical nerve injuries. This is extremely important for establishing the real advantage of endovascular angioplasty or stenting of the carotid artery. PMID- 11280833 TI - [Myogenic stromal tumors of the stomach: personal experience]. AB - Myogenic gastric tumours are a rare pathology and present difficulties in terms of nosographic classification, which in most cases can be overcome thanks to improvements in imaging and immunohistochemical techniques. Over the period 1995 1999 we observed 5 patients with aspecific dyspeptic symptoms and occasional epigastric pain, suffering from non-epithelial gastric tumours, associated, in one case, with a carcinoma of the stomach. Histological examination of endoscopic biopsies was inconclusive for a definite histopathological diagnosis, while intraoperative biopsies showed the myogenic origin and the absence of morphostructural abnormalities. In the light of these data, we performed three wedge resections, one distal gastric resection and, in the patient with advanced gastric cancer, a D3 total gastrectomy. Histological examination, immunohistochemistry and cytofluorometry enabled us to diagnose stromal tumours with a low risk of malignancy in all cases. At follow-up after 9-54 months all patients are still alive and free of disease. Though the preoperative diagnosis of stromal tumours is possible with endosonography and CT, only histology, immunohistochemistry and cytofluorometry enable us to define the condition nosographically and establish a prognosis with sufficient accuracy to allow correct surgical treatment. A prolonged follow-up is always necessary to identify eventual relapses and/or metastases, which are particularly frequent in the borderline group or in cases with a high risk of malignancy. PMID- 11280834 TI - [Surgical treatment of redundant colon after retrosternal esophagocolonoplasty for caustic esophageal stenosis]. AB - The Authors report two cases of transposed colon redundancy occurring after surgical treatment in 37 patients with caustic oesophageal strictures by retrosternal oesophagoplasty. Surgical management was required because of persistent dysphagia and weight loss in both patients. The technique performed was a resection of the redundant loop with a termino-terminal colo-colonic anastomosis via a right thoracic approach. PMID- 11280835 TI - [Treatment of hemorrhoids with circular mechanic stapler]. AB - In a pilot study undertaken in collaboration with the Department of Surgery of "San Carlo di Nancy" Hospital in Rome, over the period form January 1998 to February 2000, 128 patients with haemorrhoidal disease underwent surgery using a circular stapler to "lift" the mucous-haemorrhoidal prolapse, according to the pathogenetic theory discussed here below. We compared the results of our series with those of a retrospective series of 80 patients that undergoing traditional surgery (Khubchandani 45, Milligan-Morgan 30, Whitehead 5), evaluating length of operation, postoperative pain and complications. Our preliminary data show that the technique requires only a short learning period, reduces the length of the operation, reduces the medium- and long-term pain and allows mables the patient to resume full working activity earlier. PMID- 11280836 TI - ECG of the month. To lump or not to lump? Inferolateral myocardial infarction. PMID- 11280837 TI - Fractures of the maxilla. AB - A basic understanding of midface fractures is essential for those involved in the initial evaluation, emergency and general management, diagnosis, specialty consultation, and maxillofacial surgery of patients with fractures of the maxilla. To achieve the goal of restoring proper form and function to the upper jaw and face, one must be able to recognize, diagnose, and treat maxillary fractures. This requires knowledge of the anatomy and physiology of the midface, as well as modalities of evaluation and treatment. Each of these topics are summarized in this paper. PMID- 11280838 TI - Radiology case of the month. Right clavicle lesion. Lytic lesion of the right clavicle. PMID- 11280839 TI - The Journal 150 years ago. February 1849 Asiatic cholera. PMID- 11280840 TI - Irritable bowel syndrome: overview of diagnosis and treatment. AB - Irritable bowel syndrome (IBS) is common in primary care practice and in gastroenterology clinics and is occasionally seen in psychiatric clinics. The symptoms include abdominal cramping, bloating, and pain, as well as diarrhea, constipation, or both. Treatment includes patient education and reassurance, dietary modification, medications if necessary, and consideration of psychological interventions. The etiology of IBS is poorly understood. Research suggests a role for bowel dysmotility, altered pain perception, history of physical and sexual abuse, and psychiatric disturbance, though none of these factors alone has been proven to cause IBS. PMID- 11280841 TI - Transient visual loss due to severe anemia in a patient with AIDS. AB - We present a case of a patient with AIDS who developed a profound anemia caused by zidovudine, an important antiretroviral drug. In the setting of concurrent cytomegalovirus retinitis, the anemia produced transient visual loss that resolved with transfusion of red blood cells. Withdrawal of zidovudine resulted in a stable hemoglobin. This case describes an unusual manifestation of severe anemia. Anemia itself is a very common complication of treatment with zidovudine, one of the most commonly used agents in the treatment of AIDS. The relationship of profound anemia to transient visual loss and the role played by zidovudine in anemia in AIDS patients are discussed. PMID- 11280842 TI - Effect of diabetes education on glucose control. AB - After diabetes education, 39 adult diabetic patients were randomized to either an education group or control group. The two groups received identical medical care and follow-up, except that the education group met with their diabetes educator on at least a quarterly basis. Neither group showed any statistically significant change in their glycosylated hemoglobin values, although the education group did have a 4% drop after initial education compared to a 6% rise in the control group. The education group had a lower attrition rate and a better improvement in self-rated dietary compliance. Education remains the cornerstone of diabetes management. Our team identified some trends between the two groups as well as some ideas to improve motivating and developing a stronger and more effective relationship with our patients. PMID- 11280843 TI - Chronic non-cancer pain: an overview of assessment and contemporary management. AB - Pain is one of the foremost reasons for which people seek healthcare. The cost of pain to the American economy approximates $85 billion-$90 billion annually. Approximately one-third of Americans have some element of chronic pain. Acute and chronic pain are different entities requiring different approaches to treatment. The ability to assess chronic pain is fundamental to its management. The use of various subjective testing modalities, combined with a thorough history and physical examination and a review of pertinent laboratory data, enables the clinician to devise a management strategy. The management of chronic non-cancer pain syndrome often involves a concerted multidisciplinary endeavor that utilizes nerve block, pharmacological, psychological, surgical, and physical therapies. Trigger-point injections and sympathetic, epidural, subarachnoid, interpleural, intravenous, regional, and peripheral nerve blocks are utilized as indicated. Pharmacological management entails the use of numerous agents including nonsteroidal anti-inflammatory drugs, opioids, antidepressants, and anticonvulsants. Newer agents hold promise for facilitating the care of these patients. PMID- 11280844 TI - Doctors' dirty little secrets: the dark side of medical privacy. PMID- 11280846 TI - Marginal likelihood estimation for proportional odds models with right censored data. AB - One major aspect in medical research is to relate the survival times of patients with the relevant covariates or explanatory variables. The proportional hazards model has been used extensively in the past decades with the assumption that the covariate effects act multiplicatively on the hazard function, independent of time. If the patients become more homogeneous over time, say the treatment effects decrease with time or fade out eventually, then a proportional odds model may be more appropriate. In the proportional odds model, the odds ratio between patients can be expressed as a function of their corresponding covariate vectors, in which, the hazard ratio between individuals converges to unity in the long run. In this paper, we consider the estimation of the regression parameter for a semiparametric proportional odds model at which the baseline odds function is an arbitrary, non-decreasing function but is left unspecified. Instead of using the exact survival times, only the rank order information among patients is used. A Monte Carlo method is used to approximate the marginal likelihood function of the rank invariant transformation of the survival times which preserves the information about the regression parameter. The method can be applied to other transformation models with censored data such as the proportional hazards model, the generalized probit model or others. The proposed method is applied to the Veteran's Administration lung cancer trial data. PMID- 11280845 TI - Duration of accrual and follow-up for two-stage clinical trials. AB - Group sequential trials with time to event end points can be complicated to design. Not only are there unlimited choices for the number of events required at each stage, but for each of these choices, there are unlimited combinations of accrual and follow-up at each stage that provide the required events. Methods are presented for determining optimal combinations of accrual and follow-up for two stage clinical trials with time to event end points. Optimization is based on minimizing the expected total study length as a function of the expected accrual duration or sample size while providing an appropriate overall size and power. Optimal values of expected accrual duration and minimum expected total study length are given assuming an exponential proportional hazards model comparing two treatment groups. The expected total study length can be substantially decreased by including a follow-up period during which accrual is suspended. Conditions that warrant an interim follow-up period are considered, and the gain in efficiency achieved by including an interim follow-up period is quantified. The gain in efficiency should be weighed against the practical difficulties in implementing such designs. An example is given to illustrate the use of these techniques in designing a clinical trial to compare two chemotherapy regimens for lung cancer. Practical considerations of including an interim follow-up period are discussed. PMID- 11280847 TI - Comparing the survival of two groups with an intermediate clinical event. AB - Consider a subject entered on a clinical trial in which the major endpoint is a time metric such as death or time to reach a well defined event. During the observational period the subject may experience an intermediate clinical event. The intermediate clinical event may induce a change in the survival distribution. We consider models for the one and two sample problem. The model for the one sample problem enables one to test if the occurrence of the intermediate event changed the survival distribution. This models provides a way of carrying out non randomized clinical trial to determine if a therapy has benefit. The two sample problem considers testing if the probability distributions, with and without an intermediate event, are the same. Statistical tests are derived using a semi Markov or a time dependent mixture model. Simulation studies are carried out to compare these new procedures with the log rank, stratified log rank and landmark tests. The new tests appear to have uniformly greater power than these competitor tests. The methods are applied to a randomized clinical trial carried out by the Aids Clinical Trial Group (ACTG) which compared low versus high doses of zidovudine (AZT). PMID- 11280849 TI - Data-based modeling of the failure rate of repairable equipment. AB - A database of failures of many types of medical equipment was analysed, to study the dependence of failure rate on equipment age and on time since repair. The intention was to use this large dataset to assess the validity of some widely used models of failure rate, such as the power-law and loglinear Poisson processes, and so to recommend simple and adequate models to those practitioners having little data to discriminate between rival models. The aim is also to illustrate a methodology for computing policy costs from failure databases. The power-law process model was found to fit slightly better overall than did the loglinear and linear processes. Some related models were created to fit an observed peaking of failure rate. The data showed a decreasing hazard of (first) failure after repair for some equipment types. This can be due to imperfect or hazardous repair, and also to differing failure rates among a population of machines. Two simple models of imperfect repair were used to fit the data, and an Empirical Bayes method was used to fit a model of variable failure rate between machines. Neglect of such variation can lead to an over-estimate of the hazardousness of repair. PMID- 11280848 TI - Using frailties in the accelerated failure time model. AB - The accelerated failure time (AFT) model is an important alternative to the Cox proportional hazards model (PHM) in survival analysis. For multivariate failure time data we propose to use frailties to explicitly account for possible correlations (and heterogeneity) among failure times. An EM-like algorithm analogous to that in the frailty model for the Cox model is adapted. Through simulation it is shown that its performance compares favorably with that of the marginal independence approach. For illustration we reanalyze a real data set. PMID- 11280850 TI - Estimation in degradation models with explanatory variables. AB - Influence of covariates on degradation is modelled. Models which include dependence of the intensity of the process of traumatic events on the degradation level are also discussed. Estimation of reliability and degradation characteristics from data with covariates is considered in dynamic environments. PMID- 11280851 TI - The undescended testicle. AB - The cryptorchid testis is a common pediatric condition, usually diagnosed by the primary physician. The diagnosis, classification, and treatment options of the cryptorchid testis are discussed in hopes of clarifying some of the controversy surrounding this common problem. PMID- 11280852 TI - Clinical and radiographic evaluation of bone tumors. AB - Timely diagnosis of osseous tumors is essential in providing proper management. Appropriate imaging studies are essential to this process, however, if inconclusive, they can be superceded by information obtained through the patient history and physical examination. PMID- 11280853 TI - Partnering in CHF disease management. AB - Congestive heart failure mortality is increasing while medications are underprescribed. Disease management programs inform physicians, educate patients, increase monitoring, and facilitate compliance. Improved outcomes include decreased hospitalization and emergency room visits, and improved quality of life. PMID- 11280854 TI - Theoretical basis for the scleral expansion band procedure for surgical reversal of presbyopia [SRP]. AB - A new technique, the scleral expansion band procedure, has been developed for the surgical reversal of presbyopia. An understanding of demonstrable clinical effects of the scleral expansion band procedure, based on Schachar's theory, requires a revision of historically held views concerning the mechanism of accommodation. PMID- 11280855 TI - Cardiac risk assessment for noncardiac surgery: current concepts. AB - Strategies for perioperative risk assessment in patients undergoing noncardiac surgery vary among physicians and are aimed to estimate the risk and minimize complications. We propose simplistic guidelines for assessing and modifying risk for patients undergoing a wide variety of procedures. PMID- 11280856 TI - A review of diabetic gastropathy. AB - Diabetes mellitus affects various organs, including the gastrointestinal tract. The stomach is commonly affected, and symptoms related to the upper GI tract are frequently reported. Management of diabetic gastropathy involves dietary modifications, pharmacological agents, and occasionally, alternative feeding methods. PMID- 11280857 TI - Diagnosis and prevention of rabies. AB - Over a million Americans are bitten by animals every year. Since the rabies vaccine is uniformly effective and the disease is uniformly fatal when the vaccine is not given, management decisions must be made promptly. PMID- 11280858 TI - Menstrual cycle effects on common medical conditions. AB - Menstrual cycle-related exacerbation of common medical conditions such as migraine, epilepsy, asthma, irritable bowel syndrome, and diabetes, is a well recognized phenomenon. Accurate documentation of symptoms on a menstrual calendar allows identification of women with cyclic alterations in disease activity. PMID- 11280859 TI - Oral anticoagulant treatment in very elderly patients with atrial fibrillation. AB - Many factors can influence the final decision to treat nonrheumatic atrial fibrillation in the very elderly patient with anticoagulants. Therefore, a systemic approach in which the thromboembolic and hemorrhagic risk profiles are taken into account is suggested. PMID- 11280860 TI - Utility of prognostic stratification in adults with community-acquired bacterial meningitis. AB - Prognostic stratification uses baseline clinical features to subdivide patients into subgroups with different risks for a particular outcome. We review the importance of prognostic stratification in internal medicine, in infectious diseases, and in adults with community-acquired bacterial meningitis. PMID- 11280861 TI - [Precancerous lesions of the bronchi: endoscopic detection and treatment]. PMID- 11280862 TI - [Lymphatic involvement in non-small cell bronchial cancer: surgical and prognostic implications]. PMID- 11280863 TI - [Positron-emission tomography (PET) and non-small cell bronchial cancer]. PMID- 11280864 TI - [Radiochemotherapy of non-resectable stage III non-small cell bronchial cancer ]. PMID- 11280865 TI - [Chemotherapy of stage IV non-small cell bronchial cancer]. PMID- 11280866 TI - [Treatment of cerebral metastasis of bronchial cancer]. PMID- 11280867 TI - [What is new at the ASCO?]. PMID- 11280868 TI - [Management of small-cell bronchial cancer]. PMID- 11280869 TI - [Neoadjuvant treatment of early stages]. PMID- 11280870 TI - [Drug-induced abnormalities of the cornea]. PMID- 11280871 TI - [An immunopathological study of autoimmune keratitis in nude mice with embryonic rat thymus grafts]. AB - PURPOSE: The purpose of the present study was to examine the keratitis detected in these mice after the xenogeneic thymus gland transplantation. METHODS: The thymus glands were collected from F 344 rat embryos and transplanted under the renal capsules of BALB/c nude mice (rat TG nude mice). Histological and immunohistological examinations and a transfer experiment of keratitis was also conducted. RESULTS: The histological image was characterized by the infiltration of mononuclear cells and neutrophils and the marked angiogenesis. The autoantibodies reacting specifically to corneal epithelial cells and corneal stroma were recognized in the blood. The deposits of immunoglobulins and complements were simultaneously observed in the corneal stroma and around the basement membrane. Corneal lesions of rat TG nude mice were therefore successfully transferred into naive nude mice by host splenic CD 4 positive cells. CONCLUSION: The present study demonstrated that autoimmunity was involved in the development of keratitis in rat TG nude mice and that these mice were the first animal models to develop autoimmune keratitis spontaneously. PMID- 11280872 TI - [Adverse effects of beta-blocker eye drops on the ocular surface]. AB - PURPOSE: We investigated the adverse effects of beta-blocker eye drops on tears and ocular surface epithelium. METHODS: We studied twenty-three eyes of twenty three glaucoma patients [10 males, 13 females: 53.8 +/- 12.2 (yrs; mean +/- standard deviation)] treated with beta-blocker eye drops for more than three months and thirty two control subjects (16 males, 16 females: 50.4 +/- 10.9). The parameters described below were compared between the groups: 1. the radius of tear meniscus curvature, 2. grades for tear lipid layer interference patterns, 3. non-invasive breakup time (N-BUT), 4. cotton thread value, 5. scores of fluorescein staining, 6. fluorescein breakup time (F-BUT), 7. scores of rose bengal staining, 8. and Schirmer I value. RESULTS: The glaucoma group showed a significant decrease in the radius of tear meniscus curvature (p = 0.0007), a significantly lower distribution in the grades for tear lipid layer interference patterns (p = 0.0270), a significant difference in the scores of fluorescein staining (p < 0.0001), a significant shortening in F-BUT (p = 0.0050), a significantly higher distribution in the scores of rose Bengal staining (p = 0.0010), and a significantly smaller value in Schirmer I value (p = 0.0042). However, there was no significant difference in N-BUT and cotton thread value. CONCLUSIONS: These results clearly demonstrate that the ocular surface in glaucoma patients treated with beta-blocker eye drops show dry-eye-like changes in terms of tears and ocular surface epithelium. PMID- 11280873 TI - [The characteristics of the optically calculated axial length and the predicted refraction when using the intraoperative refraction]. AB - PURPOSE: To understand the effect of the error in corneal refractive power on the optically calculated axial length (AXO) and the postoperative predicted refraction (Q'gl) calculated from the aphakic refraction (Qaph). METHOD: We used the Gullstrand's schematic eye, in which the intraocular lens (+20 D) was inserted at a depth of 5 mm, and calculated Qaph and postoperative refraction (Qgl) geometrically when the corneal radius of the anterior surface (Rfc) changed from 7 mm to 9 mm. AXO was calculated using the calculation formula from a previous report, and then Q'gl was calculated from the AXO and the true axial length (AXT) using the theoretical calculation formula (regarding the fictitious corneal refractive index as 1.3315). RESULTS: When the measured corneal radius of the anterior surface (K) was equal to Rfc, the error of the AXO was largest (AXO/AXT = 101.512) when K was 9 mm. The error in power prediction (Qgl-Q'gl) of the AXT was 7.7 times larger than that of AXO. CONCLUSION: If K is exact, AXO is useful to predict the intraocular lens depth because the error of AXO is small. The error in power prediction of AXO is smaller than that of AXT. PMID- 11280874 TI - [Grading of light perception and hand movement utilizing a novel device called the low vision evaluator]. AB - PURPOSE: There has been no device for grading very low visual function expressed as light perception or hand movement up to the present. It is necessary to evaluate even slight effects of modern and classical therapies during the follow up period. The goal of this study was to invent a novel device for grading such low visual function. METHODS: Sixty-two patients with a visual acuity of counting fingers or worse due to various diseases were examined. The device, which we call the Low Vision Evaluator, has a pair of goggles equipped with two white light emitting diodes. Nine variations of stimulus for light intensity (1, 10, 100 cd/m2 or 10, 100, 1,000 cd/m2) and duration (0.1, 0.3, 1 second) can be delivered in a random sequence. Patients were asked to push a button whenever they saw a stimulus. Response of patients to the stimuli was recorded. RESULTS AND CONCLUSIONS: The visual function of light perception and hand movement could be reproducibly assessed by this method. The method represents a valuable new tool for evaluating low visual function. PMID- 11280875 TI - [Shortening the duration of prone positioning after macular hole surgery- comparison between one week and one day]. AB - PURPOSE: We shortened the duration of prone positioning after macular hole surgery from one week to one day and evaluated the initial hole closure rate. SUBJECT AND METHOD: The first group included 34 eyes of 33 patients who underwent surgery between April 1998 and August 1999. All 33 patients were instructed to maintain the prone position for one week after surgery (one week group). The second group included 21 eyes of 21 patients who underwent surgery between September 1999 and March 2000. These 21 patients were told to maintain the prone position for 24 hours after surgery (one day group). The indication for surgery was eyes with the reported onset of symptoms within 6 months and without long standing macular holes. Eyes which underwent retinal pigment epithelium (RPE) or internal limiting membrane (ILM) removal were excluded from the study. Phacoemulsification and intraocular lens implantation were combined in patients with phakic eyes. In the one day group, the patients were instructed to avoid the face up position for one week. RESULTS: There were no significant differences between the two groups in terms of preoperative factors, and initial hole closure rates (i.e., 91.2% in the one week and 90.5% in the one day groups). CONCLUSION: These results suggests that the duration of prone positioning after macular hole surgery can be reduced to as little as 24 hours without RPE or ILM removal in eyes without long-standing holes. PMID- 11280876 TI - [Measurement of apparent accommodation with a 20/20 near vision optotype]. AB - PURPOSE: The value of apparent accommodation varies with methods of measurement. To discuss the details of apparent accommodation, it is appropriate to measure it with the smallest possible near vision optotype. In the present study, we used a 20/20 near vision optotype for the measurement of apparent accommodation. SUBJECTS AND METHODS: Forty-six eyes of thirty-eight patients (45-84 years old) who had undergone cataract surgery and intraocular lens implantation, and had at least 20/20 best corrected visual acuity at near and far distances, were used in this study. After the eyes were corrected by glasses to gain the best corrected long distance visual acuity, they were forced to watch a 20/20 near vision optotype. Then we gradually added plus lenses until they could recognize the optotype. The value of apparent accommodation was recorded by subtracting the value of plus lens by which the eye could first recognize the 20/20 near vision optotype from three diopters. RESULTS: The value of apparent accommodation was 0.00-3.00 D (medium 0.50 D). Two eyes had three diopters of apparent accommodation. CONCLUSION: In the present study with correction of astigmatism and small near vision optotype, most eyes showed smaller apparent accommodation than those in previous studies. Despite that, patients with three diopters of apparent accommodation do exist. To analyze high quality visual functions, we should use the smallest possible near vision optotype for the measurement of apparent accommodation. PMID- 11280877 TI - [Blood flow in retinal vessels of normal-tension glaucoma with or without a history of optic disc hemorrhages]. AB - PURPOSE: To evaluate blood flow in retinal vessels of normal-tension glaucoma (NTG) with or without a history of optic disc hemorrhages (DH) and compare it with that in non-glaucomatous eyes using scanning laser fluorescein video angiography. METHODS: We enrolled 14 eyes of 14 NTG patients with a history of DH (DH (+) group), 12 eyes of 12 NTG patients without history of DH (DH (-) group), and 10 eyes from 10 non-glaucomatous patients matched for age, intraocular pressure, and systemic blood pressure. No statistically significant difference was observed between the DH (+) and DH (-) groups of NTG in the global indices of the Humphrey visual field. Fluorescein angiography was performed using a scanning laser ophthalmoscope with an argon blue laser. A series of approximately 100 consecutive video images at 1/2 second intervals from just before the dye appearance in the central retinal artery was loaded into an external personal computer system. Based on this acquired image series, we obtained fluorescein filling curves for 10 x 10 pixel measuring areas placed on each of the superior temporal and inferior-temporal branch retinal arteries and veins at 1/5 papillary diameter from the disc edge. In each vessel, time to the highest fluorescein intensity (peak time, sec) and the time constant of the filling curve (tau, sec) were obtained. Time difference between the peak times in vein and artery (peak time difference) was also calculated. RESULTS: Statistically significant differences were observed among the three groups in the peak time of inferior temporal artery and vein, and superior-temporal vein (ANOVA, p < 0.01). Also there were statistically significant differences in the tau of all vessels (ANOVA, p < 0.05). No statistically significant differences were observed in the peak time differences. By multivariate analysis, the DH (+) and DH (-) groups of NTG showed significantly longer peak times and tau s than did the non glaucomatous eyes (p < 0.05). However, no statistically significant differences were observed in any parameters between the DH (+) and DH (-) groups of NTG. CONCLUSIONS: In NTG, dye filling rate in both the central retinal arteries and veins seems to be delayed. However, this delay does not differ between DH (+) and DH (-) groups. PMID- 11280878 TI - [Clinical features of epidemic nosocomial keratoconjunctivitis in 41 patients]. AB - PURPOSE: To evaluate the clinical features of nosocomial epidemic keratoconjunctivitis (EKC) occurring in the ophthalmology ward of Tokyo Medical University Hospital. MATERIALS AND METHODS: We studied the symptoms and clinical course of 41 patients who had EKC caused by nosocomial infections in our hospital. We attempted to detect adenovirus antigen and viral DNA from conjunctival swabs and also to isolate the virus. RESULTS: The clinical symptoms of EKC, including postoperative cases, were not severe. In some cases, patients' complaints, for example, increase of lacrimation or appearance of a foreign body sensation, were contributory to diagnosis. Among the 41 patients, 31 out of 34 (91.2%) EKC patients who had undergone ophthalmic surgery had EKC in the operated eye. In cases receiving bilateral operations, EKC occurred first in the initially operated eye. The sensitivity of Adeno-check was 76.9%. Adenovirus type 19 was isolated from conjunctival swabs in 26 cases. CONCLUSIONS: The early diagnosis of EKC is extremely important to prevent the spread of nosocomial infections. Careful observation of operated eyes and close attention to patient complaints may aid in the early detection of EKC. PMID- 11280879 TI - [A case of suspected drug-induced ocular pemphigoid]. AB - BACKGROUND: Lacrimal obstruction can occur as a complication of ocular cicatricial pemphigoid. We report a patient who was diagnosed as having drug induced ocular cicatricial pemphigoid associated with acquired lacrimal canalicular obstruction in spite of relatively mild subconjunctival scar formation. CASE: The patient was a 69-year-old woman. She had been treated for glaucoma, blepharoconjunctivitis, dacryocystitis, and lacrimal canalicular obstruction for two years with topical administration including 0.5% timolol maleate, 0.1% dipivefrine hydrocholoride, 0.1% fluorometholone, and 0.3% ofloxacin. The patient had moderate conjunctival hyperemia without shortening of the inferior conjunctival sac, corneal ulceration, neovascularization, and keratinization. Lacrimal canalicular obstruction progressed further as topical ocular medications were continued. Topical anti-glaucoma medications were stopped after dacryocystorhinostomy. Although the blepharoconjunctivitis was improved, left inferior conjunctival symblepharon, and medical canthal keratinization was progressive despite the use of topical corticosteroids. The conjunctival biopsy specimen showed the lacrimal punctum covered with proliferated conjunctival epithelium. There was a moderate stromal infiltration of small lymphocytes and plasma cells. We diagnosed this patient as having drug-induced ocular cicatricial pemphigoid caused by topical anti-glaucoma medications. CONCLUSION: Lacrimal canalicular obstructions may occur with the topical anti-glaucoma medications even when subconjunctival scarring is mild. PMID- 11280880 TI - [The formation and involution of the optociliary vein during the course of central retinal vein occlusion]. AB - BACKGROUND: Formation of collateral pathways by veins on the optic nerve head is sometimes observed during the resolution of a central retinal vein occlusion (CRVO). However, no cases have been reported which documented the formation and involution of these collateral pathways during the resolution of a CRVO. CASE: A 56-year-old man with a non-ischemic CRVO was followed over a 10-year period by ophthalmoscopy, fluorescein angiography (FA), and indocyanine green angiography (IA). Initial examination revealed only mild arteriosclerosis and absence of the optociliary vein (OCV) in the right eye. Approximately 9 years later, the patient returned with non-ishemic CRVO, and FA demonstrated a prolongation of the retinal circulation time to 12.1 seconds (normal < 11.0 seconds). Monitoring of this patient during treatment disclosed development of an OCV on the optic nerve head and gradual decrease in the retinal hemorrhage. In FA, the retinal circulation time was shortened and returned to normal. IA verified an outflow through the OCV into choroidal veins. Later, ophthalmoscopy showed a narrowing of the diameter of the OCV. CONCLUSION: We suggest that the OCV developed to drain blood from retinal veins to the vortex veins through choroidal veins to compensate for the slowing of retinal blood outflow. The reperfusion of the central retinal vein was accompanied by the narrowing of the OCV. PMID- 11280881 TI - Pheochromocytoma of the urinary bladder: a case report. AB - We present a 66-year old female patient with pheochromocytoma of the urinary bladder. We performed transabdominal needle biopsy of the tumor without suspicion of pheochromocytoma because of her well-controlled blood pressure and no characteristic symptoms following administration of antihypertensive medication. Hypertensive crisis (260/130 mmHg) occurred just after the needle insertion. The diagnosis was pheochromocytoma. The norepinephrine level in the serum and her blood pressure normalized without antihypertensive medication after partial cystectomy. Pheochromocytoma should be suspected in cases of intramural bladder tumors, especially in a normotensive patients receiving antihypertensive medication. PMID- 11280882 TI - [Vesicorectal fistula due to pelvic foreign body: a case report]. AB - We report a case of vesicorectal fistula caused by a pelvic foreign body. An 84 year-old woman presented with urinary tract infection and bladder stone. During transurethral lithotripsy, a foreign body was observed in the stone. CT and colonoscopy revealed a vesicorectal fistula due to a foreign body. After continuous bladder washout over a period of one month, resection of the foreign body, fistulectomy, and sigmoidostomy were performed. The foreign body was suspected to be a medical mesh from a sling operation. After the surgery, the patient's course was uneventful. This is the second patient with vesicointestinal fistula due to a foreign body in the Japanese literature. PMID- 11280883 TI - [Small cell carcinoma of the prostate: a case report]. AB - A 81-year-old man was admitted to our department with the chief complaints of pollakisuria and difficulty in voiding. He presented with increased serum PSA level (over 100 ng/ml). We performed biopsy of the prostate and found a moderately differentiated adenocarcinoma. Various urological examinations showed metastases to paraaortic lymph nodes and systemic bones. He was started-on hormonal therapy. Nine months from the start of hormonal therapy, this therapy was effective and the serum PSA level was decreased to 14 ng/ml. Thereafter, the serum PSA level and the tumor volume were increased and he died 29 months from the start of treatment. The autopsy revealed small cell carcinoma with adenocarcinoma of the prostate. PMID- 11280884 TI - [Testicular pure teratoma: a case report]. AB - A 46-year-old man visited our hospital, complaining of fever and painful swelling of the right scrotum contents. The symptoms and signs suggested epididymitis, but testicular tumor could not be excluded. Therefore, high inguinal orchiectomy was performed. Macroscopic findings were compatible with the testicular teratoma, containing hair, and epididymitis. Histological findings revealed that the tumor was composed of mature epidermis with skin appendages, cartilage, hair, bone and adipose tissue, and that many leucocytes infiltrated in the epididymis, resulting in the diagnosis of pure mature teratoma with epididymitis. For 6 months after the operation, no evidence of recurrence has been observed. Despite its histologically benign appearance, primary pure teratoma of the testis has a metastatic and recurrent potential. Therefore, primary pure teratoma should be man aged in the same way as other nonseminomatous germ cell tumors. PMID- 11280885 TI - [A case of intratubular germ cell tumor giving rise to seminoma in a subfertile man]. AB - A 34-year-old man visited our hospital with the complaint of right scrotal swelling. Right high orchiectomy was performed under the diagnosis of testicular tumor. Pathological examination was seminoma, pT2pN0pM0 and 3 courses of chemotherapy (cisplatin, vinblastine, bleomycin) were performed. The patient had undergone testicular biopsy for infertility at another hospital 6 years before this visit. Re-examination of the biopsy specimen revealed a intratubular germ cell tumor. No evidence of recurrence or metastasis was found 2 years after surgery. PMID- 11280886 TI - [Statistics on the operations at the Department of Urology, Fuji City General Hospital during a 10-year period (January 1989-December 1998)]. AB - A 10-year statistic survey was carried out on the operations performed at the Department of Urology, Fuji City General Hospital. The total number of operations was 1,862 cases during 10 years. The number of operations for malignant tumors has increased since 1994 year after year. Transurethral resection of prostate for benign prostatic hyperplasia has decreased since 1992, with the advent of the alpha 1-blocker. The patients with urolithiasis are being cured by extracorporeal shock wave lithotripsy on an outpatient basis, at our hospital, except in special cases. It was suggested to be safe, so the complications needing admission appeared in only 1.2% of the cases. PMID- 11280887 TI - [Diagnosis and treatment of prostate cancer--it's diversity and future prospects]. PMID- 11280888 TI - [Laparoscopic adrenalectomy at Shinshu University School of Medicine]. AB - We report our experience with transperitoneal laparoscopic adrenalectomy in 26 cases (mean age 45 years). We experienced primary aldosteronism in 19 cases, Cushing syndrome in 6 cases and non-functioning tumor in one case. There was no significant difference in the operation time between right and left, men and women, primary aldosteronism and Cushing syndrome. The blood loss decreased with training. There were no severe complications during and after the operation. The weight of the resected adrenal glands increased. The blood loss decreased significantly compared with the open surgery. Transperitoneal laparoscopic adrenalectomy is becoming the safe and standard surgery for the adrenal gland tumor, and the number of suitable cases for this procedure is expected to increase in the future. PMID- 11280889 TI - [Experience with partial nephrectomy for renal cell carcinoma]. AB - From Sept. 1991 to Jan. 1999, we performed partial nephrectomy on 7 patients with renal cell carcinoma. The indication was imperative for 3 patients, and elective for 4 patients. The 3 imperative cases consisted of bilateral renal cell carcinomas, a polycystic kidney disease and a contralateral atrophic kidney. All 4 patients with elective indication revealed renal cell carcinoma with a normal functioning contralateral kidney. The tumor size ranged from 1.3 cm to 6.0 cm (2.7 cm on average). The mean clamping time of renal artery was 22 minutes and mean blood loss was 400 ml. The pathological stage was pT1a in 6 patients and pT1b in one patient. Postoperative follow-up ranged from 4 months to 92 months (mean: 43 months). One patient with bilateral renal cell carcinoma died of metastases to the lungs and brain at 25 months postoperatively. The remaining 6 patients are alive without recurrence and metastasis. We obtained a good postoperative course in our selected patients with low stage. Thus it was considered that partial nephrectomy is effective against small renal cell carcinoma. PMID- 11280890 TI - [A randomized comparative study assessing once versus twice a day treatment of benign prostatic hyperplasia with terazosin]. AB - We compared the efficacy of once a day administration of terazosin hydrochloride with that of twice a day administration for benign prostatic hyperplasia (BPH) patients. Forty-two patients with BPH were randomly assigned to receive a maximum dose of 2 mg terazosin either once (n = 21) or twice (n = 21) a day. International prostate symptom score (IPSS), uroflowmetry and side effect profile were determined before and 4 weeks after randomization. Both groups were similar with respect to patient age, baseline IPSS and prostate volume. After 4 weeks of treatment with terazosin, significant improvement in IPSS, maximum flow rate and mean flow rate were observed in both groups. However, these improvements did not differ significantly between them. In addition, there were no differences in side effects between the groups. In conclusion, once a day administration of terazosin hydrochloride is as effective and safe as twice a day administration in patients with BPH. PMID- 11280891 TI - [The voiding after the suburethral sling operation, obstructive or non obstructive?]. AB - To examine whether or not the suburethral sling operation produces obstruction during voiding, seven females who underwent the sling operation using synthetic material (Vesica Sling Kit) were studied postoperatively urodynamically. Uroflowmetry and residual urine measurement showed no overt voiding difficulties in any cases. However, in one case, a pressure flow study indicated equivocal and the position of the sling material was judged to be too proximal by fluoroscopic monitoring. In all other 6 cases pressure flow was shown to be non-obstructive. Fluoroscopic finding also demonstrated in these 6 cases an appropriate bladder neck opening at the time of voiding and the sling was positioned just from the bladder neck to mid-urethra. Thus, it is concluded that the suburethral sling operation produces no obstruction as long as the position of the sling material is carefully determined from bladder neck to mid-urethra and excessive tension is avoided. PMID- 11280892 TI - [A case of adrenal pheochromocytoma with contralateral adrenocortical adenoma]. AB - A case of a pheochromocytoma in the right adrenal gland and adrenocortical adenoma in the left adrenal gland of a 58-year-old male is reported. The patient was incidentally found to have a right adrenal tumor by ultrasonographic study. A computerized tomographic (CT) study and magnetic resonance image (MRI) study revealed bilateral adrenal tumors. The sizes of the right tumor and left tumor were 2.5 x 3.5 cm and 1.2 x 1.0 cm, respectively. The intensity of each tumor was different on T2-weighted MRI. 131I-MIBG scintigram showed the uptake of right adrenal gland. The existence of pheochromocytoma was confirmed by the elevated levels of catecholamines. We performed venous sampling to be certain whether the patient had unilateral or bilateral pheochromocytoma. As a result, bilateral adrenal pheochromocytoma was diagnosed. Therefore, we performed bilateral adrenalectomy. However, histopathological examination revealed right pheochromocytoma and left non-functioning adrenocortical adenoma. PMID- 11280893 TI - [A case of malignant fibrous histiocytoma arising from the renal capsule]. AB - A 62-year-old man was admitted with a chief complaint of general malaise. Computed tomography showed a large mass adjacent to the parenchyma of the left kidney. The mass was 17 x 13 x 12 cm in size. Preoperative diagnosis was left renal cell carcinoma and left radical nephrectomy was performed. Histopathologically, the tumor was diagnosed as malignant fibrous histiocytoma (MFH), and the tumor was considered to have arisen from the renal capsule. There has been no recurrence for 7 months postoperatively. We review 40 cases of MFH arising from the kidney or the renal capsule in the literature. PMID- 11280894 TI - [A case of transitional cell carcinoma of the bladder in a juvenile patient]. AB - A 15-year-old male was referred to our hospital with the chief complaint of gross hematuria. Cystoscopic examination revealed a papillary tumor on the posterior wall. Transurethral resection of the bladder tumor was performed. Histological examination of the excised tumor showed transitional cell carcinoma, grade 1, pTa. Recurrence has not been observed for about 2 years after the operation. We investigated 54 previously reported Japanese cases of bladder cancer before age twenty including the present case. PMID- 11280895 TI - [Recent advances of models for scleroderma with special reference to tight skin mouse]. PMID- 11280896 TI - Effects of oral administration of Echinacea purpurea (American herb) on incidence of spontaneous leukemia caused by recombinant leukemia viruses in AKR/J mice. AB - Four-week-old female AKR/J mice were given oral doses of powdered leaves from Echinacea purpurea three times weekly for 8 weeks (7.5 mg/mouse/week): controls received phosphate-buffered saline. Mean survival age of experimental AKR/J mice treated with the E. purpurea preparation was significantly prolonged and enlargement of thymic lymphoma in experimental mice was significantly suppressed compared with controls. In normal 3-week-old female AKR/J mice, mortality from thymic lymphoma was delayed markedly after injection into the thymus of cell-free extract of thymus from the experimental 28-week-old female AKR/J mice that received the oral E. purpurea preparation was injected directly into the thymus. Proliferation of endogenous recombinant murine leukemia viruses (MuLV) in the thymus was markedly inhibited after the first oral administration of the E. purpurea preparation as compared with untreated controls (final age, 28 weeks). Production of endogenous interferon (IFN)-gamma in AKR/J mice was also effectively augmented by the oral treatment with the E. purpurea preparation, however, the production of other cytokines such as tumor necrosis factor (TNF) alpha and interleukin (IL)-12 was minimal. These results suggest that this suppressive effects on spontaneously occurring leukemia caused by endogenous recombinant MuLV in female AKR/J mice may depend on enhancement of nonspecific immune or cellular immune systems (or of both) by the E. purpurea preparation. PMID- 11280897 TI - [A case of sarcoidosis characterized by a nasopharyngeal tumor and neurological lesions]. AB - A 20-year-old man was admitted to a hospital complaining a slight fever lasting for 3 months associated with a dull headache and weight loss. A tumor was found in the nasopharynx of which biopsy specimen revealed granulomas with Langhans' giant cells. He was given antituberculous agents without symptomatic improvement, and transferred to our hospital. Serum levels of soluble IL-2 receptor and lysozyme were increased, and a significant uptake was observed by Ga scintigraphy at the nasopharynx and bilateral hilar lymphnodes. Furthermore, spinal fluid contained increased number of mononuclear cells, and T2-weighted MRI scans showed an enhanced lesion at the pituitary stalk. The specimen of both TBLB and repeated biopsy of the nasopharyngeal tumor showed granulomas without caseous necrosis. Taken together with these findings, a diagnosis of sarcoidosis with CNS involvement was finally made, and he made a favorable progress by treatment with prednisolone. This is an unique case which emphasizes importance of differential diagnosis of nasopharyngeal tumors with neurological manifestations in the clinicalsetting of rheumatology. PMID- 11280899 TI - [Ventilatory failure due to the limitation of chest movement in a case of FPS]. AB - A 72-year-old woman presented with cervicothoracal skin lesions mimicked to scleroderma and muscular atrophy in 1996. Because of the elevation of serum creatinine kinase (CK), muscular biopsy was performed at another institution. Under the diagnosis of polymyositis, she was treated with corticosteroid. Despite of the decrease in serum CK levels by corticosteroid therapy, skin lesions and mascular dystrophy gradually worsened to extend to the regions of major pectoral, paravertebral, and femoral muscles. In 1997, she was admitted to our hospital because of dyspnea. On admission, the limitation of the chest movement was obvious and she developed respiratory arrest due to CO2 narcosis. The femoral magnetic resonance image (MRI) showed increased signal intensity of subcutaneous tissues and fascia on T2-weighted image. The block biopsy specimens obtained from the cervical lesion revealed fibrotic thickness and chronic inflammation of subcutaneous septa, fascia, and perimysium. She was treated by mechanical ventilation and cimetidine and weekly methotrexate were added to the corticosteroid therapy because of the diagnosis of FPS. Thereafter, the skin and muscular lesions as well as the MRI findings were improved. The concept of FPS was proposed by Naschitz et al. This condition is pathologically characterized by cicatrizing fascitis, septal and lobular panniculitis, and perimysial fibrosis and peripheral blood and tissue eosinophilia is not important for diagnosis. FPS includes classical eosinophilic fascitis but is also associated with several disorders such as malignancy. This case is suggestive of the therapeutic consideration of FPS in terms of the response to cimetidine and MTX. PMID- 11280898 TI - [Cytomegalovirus antigenemia assay as a useful tool for early diagnosis and therapy for cytomegalovirus infections in three cases with collagen diseases]. AB - We report here three cases of collagen diseases with cytomegalovirus infections. (1) A 21-year-old female, who had been diagnosed as systemic lupus erythematosus, lupus nephritis and lupus peritonitis, had fever. Cytomegalovirus antigenemia (CMV-Ag) assay was 10/8 positive. (2) A 33-year-old female, who had been diagnosed as Wegener glanulomatosis, had fever and liver dysfunction. CMV-Ag assay was 933/896 positive. (3) A 64-year-old female, who had been diagnosed as microscopic polyangitis, had fever, liver dysfunction and pneumonia. CMV-Ag assay was 6/2 positive. They were considered to be complicated with CMV infections. We could make early diagnoses of CMV infection by using CMV-Ag assay and treat them with anti-CMV therapy effectively. PMID- 11280900 TI - [Brain abscess developed five years after splenectomy in a patient with idiopathic thrombocytopenic purpura]. AB - The increased susceptibility to infection following splenectomy has been well documented. We report here, a case of brain abscess developed five years after splenectomy in a patient with idiopathic thrombocytopenic purpura (ITP). A 65 year-old woman was admitted to our hospital because of fever, mental disorientation, and weakness in August, 1999. She had been followed with diagnoses of essential hypertension and type 2 diabetes mellitus (DM) since 1988. The patient was diagnosed as having ITP in 1992, and then underwent splenectomy in 1994. Monoclonal gammopathy of undetermined significance (MGUS) of IgG, lambda type was found in February 1999. Though high grade fever persisted after admission, blood cultures were negative. Antibiotics were given without a satisfactory effect. Right hemiparesis and worsening of consciousness developed subsequently. Contrast enhanced cranial computed tomography (CT) disclosed a ringed enhanced low density mass in the left parieto-occipital lobe compatible with a diagnosis of brain abscess. Surgical drainage was performed and 20 ml of pus was obtained. No primary infectious focus of the brain abscess was detected with intensive examinations. PMID- 11280901 TI - [A boy diagnosed SLE after Staphylococcus aureus infection over a long period]. AB - We encountered a 13-year-old boy with SLE who showed specific pathophysiology. The affected child was hospitalized because of long-standing cutaneous empyesis considered to be Staphylococcus aureus infection, followed by manifestation of meningoencephalitis-like symptoms. On a close check up, the patient was diagnosed as having SLE complicated with interstitial lupus nephritis and verrucosis of the left ventricle. Besides the findings, the blastogenesis of the patient's lymphocyte was low against stimulation of sac-1 which connects with the Fc portion of lgG, one of the constituent proteins of Staphylococcus. Moreover, anti phospholipid antibodies turned positive during immunosuppressive therapy and subcutaneous abscess due to Pseudomonas aeruginosae developed concurrently at about the same time, which posed difficulties in the treatment. The affected child had had Staphylococcus aureus infections over a long period of time before diagnosis of SLE and was susceptible to bacterial infections due to Pseudomonas aeruginosae during the treatment. The clinical course of this case was considered important in presuming the complex immunologically abnormal condition of SLE in childhood. PMID- 11280902 TI - [The occurrence of Churg-Strauss syndrome in two patients with remitted asthma]. AB - We report two patients with Churg-Strauss syndrome (CSS) which occurred on the remission stage of bronchial asthma. Case 1 of a 64-year-old woman suffered from asthma in June, 1997, and got relief with treatment. In February, 1998, dysesthesia, pain and severe muscle weakness occurred in the extremities and erythematous rashes appeared on the extremities and back. She was transferred to our hospital on March 3. Peripheral blood eosinophilia was observed and a diagnosis of CSS was made. Eosinophilic tissue infiltration and vasculitis was found in the skin biopsy specimen. She was treated with prednisolone (60 mg/day) with moderate improvement. But the dysesthesia in the extremities, bilateral foot drop and the weakness of the left hand grasping power continued. Case 2 of a 62 year-old woman suffered from asthma in 1995, which improved by treatment. In March 1998, dysesthesia and pain in the lower extremities occurred and progressed. Erythematous rashes appeared on the feet and she was admitted to our hospital in May. Peripheral blood eosinophilia and eosinophilic tissue infiltration in the skin biopsy specimen were observed and a diagnosis of CSS was made. The treatment with prednisolone (60 mg/day) improved the pain but the dysesthesia continued. It is important to know the occurrence of CSS on the remission stage of bronchial asthma. PMID- 11280903 TI - [Surgery for hilar cholangiocarcinoma--result and limit of surgical treatment]. PMID- 11280904 TI - [Biliary endoprosthesis for hilar tumors]. PMID- 11280905 TI - [Clinicopathological study with regard to ulcer in the cancer lesion of the depressed type intramucosal cancer of the stomach]. AB - We histologically investigated 207 cases with the depressed type intramucosal cancer of the stomach, as to the risk factor of ulcer formation in the cancer (Ul.), tumor size of less than and not less than 10 mm, histological type of differentiated carcinoma (Diff.) and undifferentiated carcinoma (Undiff.), tumor location classified by the usually occurring region of the peptic ulcer (Usual portion and Unusual portion) and the intramucosal propagations of extensively vertical type (EV) and nonextensively vertical type (NV). RESULT: 1) Tumor size, histological type, tumor location and intramucosal propagation type were significantly related to Ul., as the result of multivariate analysis. 2) The frequency of Ul. at the lesions of these cases, regardless of histological type, was higher at the lesion of Usual portion than at Unusual portion. And the frequency of Ul., independently of tumor location, was higher in Undiff. than in Diff.. 3) The frequency of Ul. at the lesions of Unusual portion in Undiff. was higher in EV than in NV. CONCLUSION: We suggested that extensively vertical propagation type of intramucosa, as well as large tumor size, Undiff. and at the lesion of Usual portion, was a significant risk factor for Ul. of the depressed type intramucosal cancer of the stomach. 2) We concluded that the intramucosal propagation type of the cancer was particularly concerned with Ul. at the intramucosal lesion of Unusual portion in Undiff. PMID- 11280906 TI - [A long-term survival case of gastric carcinoid with multiple liver metastasis]. PMID- 11280907 TI - [Intramural hematoma of the large intestine caused by cytomegalovirus vasculitis in a patient with SLE]. PMID- 11280908 TI - [A case of ulcerative colitis (proctitis type) complicated with deep venous thrombosis and treated with heparin successfully]. PMID- 11280909 TI - [A case of multiple liver metastasis of gastric cancer which was effectively treated by arterial infusion chemotherapy with hyperthermia and 10 years survival]. PMID- 11280910 TI - [A case of autoimmune pancreatitis observed from local enlargement to the diffuse swelling of the pancreas in its long clinical course]. PMID- 11280911 TI - [A case of pancreatic arteriovenous malformation treated by transcatheter arterial embolization and transjugular intrahepatic portosystemic shunt]. PMID- 11280912 TI - [Successful treatment of massive pancreatic ascites with peritoneal drainages and a somatostatin analogue. Report of a case]. PMID- 11280913 TI - [A case with long-term survival after undergoing total pancreatectomy for intraductal papillary-mucinous carcinoma]. PMID- 11280914 TI - [Hepatocellular carcinoma with A-V shunt formation after US-guided biopsy, PEIT, and PMCT]. PMID- 11280915 TI - [A point mutation at Arg169 (CGG-->TGG) in hereditary protein C deficiency]. AB - We investigated a 56-year-old Japanese man with protein C deficiency, who was referred to our hospital because of venous sinus thrombosis and pulmonary thromboembolism. Protein C (PC) activity and the corresponding antigen level in plasma were 66% and 106% of the normal values, respectively. Both the activity and antigen levels of protein C were reduced by approximately 50% in plasma from the patient's mother. All exons and their flanking intron regions were amplified by PCR from genomic DNA. Sequencing analysis of the PCR fragments revealed that the patient was heterozygous for a C to T substitution at nucleotide position 6218, resulting in a single amino acid substitution of arginine (CGG) by tryptophan (TGG) at codon 169 of the heavy chain. We analyzed the patient, his mother, and normal controls by a Sac II digestion study of exon 7 and found that the patient and his mother had the same C to T point mutation at base 6218. This mutation could have been responsible for the defective activation of the molecule and the resulting thrombotic disorder. The patient is now being treated with warfarin, and so far no further clinical thrombotic episode has occurred. PMID- 11280916 TI - [Complex translocation (8;15;21) (q22;p12;q22) in a child with AML-M2 showing de novo appearance of the short form of AML1-MTG8 chimeric mRNA during the course]. AB - A 4-year-old boy admitted with exophthalmos was diagnosed as having acute myeloblastic leukemia with maturation (AML-M2). Chromosomal analysis (G-banding) showed t(8;15;21). Fluorescence in situ hybridization (FISH), reverse transcriptase-polymerase chain reaction (RT-PCR), spectral karyotyping (SKY), and nucleolar organizer region (NOR) staining suggested that this complex translocation might have resulted from stepwise translocation, namely an initial translocation between chromosomes 8 and 21, followed by a second translocation between der(21) and chromosome 15, rather than the other possibility of clockwise translocation. During chemotherapy, RT-PCR demonstrated the short form of AML1 MTG8 mRNA, in addition to chimeric mRNA of the usual length. Sequence analysis revealed that this shorter chimeric mRNA had resulted from deletion of a 250-bp sequence at the 5' end of MTG8. A literature search failed to reveal any similar cases of t(8;21) AML-M2 associated with this deletion of chimeric mRNA. PMID- 11280917 TI - [Philadelphia chromosome-positive acute lymphoblastic leukemia with monosomy 7 successfully treated with intermediate- and high-dose ara-C]. AB - A 52-year-old man was admitted for treatment of acute lymphoblastic leukemia (ALL). The bone marrow was hypercellular with 67.2% blasts, which were negative for peroxidase, and expressed CD13, CD33, CD34, CD10 and CD7. Cytogenetic and molecular studies revealed t(9;22) and -7(Ph/-7) with major BCR/ABL rearrangement. The patient was treated with the L-AdVP regimen, but failed to achieve complete remission (CR). He then received two courses of chemotherapy consisting of intermediate- and high-dose cytarabine (ara-C), resulting in CR. This case suggests that Ph/-7 ALL with major BCR/ABL gene rearrangement showing coexpression of myeloid antigens may be sensitive to intermediate- and high-dose ara-C. PMID- 11280918 TI - [Medicine in 21st century: contribution of hematology to advanced medicine]. PMID- 11280919 TI - [Paroxysmal nocturnal hemoglobinuria: pathogenesis and diversity of clinical manifestations]. PMID- 11280920 TI - [Upshaw-Schulman syndrome: its pathogenesis and therapy]. PMID- 11280921 TI - [Role of ultrasonography in diagnosis of neutropenic enteritis: a study of 4 cases]. AB - Neutropenic enteritis is a septic or inflammatory disease of the colon. It is usually encountered in patients with hematological malignancy who have undergone chemotherapy, and it presents as fever, diarrhea, and abdominal pain, although the symptoms are not always specific. The diagnostic features of neutropenic enteritis revealed by barium enema, CT and ultrasonography have been reported previously. Here we report 4 cases of neutropenic enteritis in which ultrasound was used for diagnosis, and also for monitoring the clinical course of the disease. Because neutropenic enteritis is rapidly progressive, early diagnosis and therapeutic intervention are required. We believe that ultrasonography is a useful method for examining patients with neutropenic enteritis, being noninvasive, mobile, and providing rapid results in real time, thus aiding early diagnosis and clinical follow-up. PMID- 11280922 TI - [Sequential analysis of p210- and p190-bcr-abl by RT-PCR after allogeneic bone marrow transplantation for p210/p190-bcr-abl double positive acute lymphoblastic leukemia]. AB - In patients with both p210-bcr-abl (p210) and p190-bcr-abl (p190)-positive acute lymphoblastic leukemia, the number of p190 transcripts is lower than that of p210 transcripts. It is speculated that the p190 transcript occurs as a consequence of alternative splicing or missplicing events in the BCR gene. Four patients with both p210- and p190-positive acute lymphoblastic leukemia were studied for expression of p210 and p190 by RT-PCR before and after allogeneic bone marrow transplantation. p190 negativity was documented in all four patients, followed by p210 negativity one to two months later in three patients. These results suggest that negativity for p190 indicates an ongoing decrease in the small number of residual leukemic cells. In one patient p190 appeared transiently in spite of prolonged negativity for p210 18 months after bone marrow transplantation. We conclude that analysis of p210 and p190 is useful for following up patients with both p210- and p190-positive acute lymphoblastic leukemia. PMID- 11280923 TI - [Transitional pre-B-cell ALL]. AB - An 18-month-old girl was referred to our hospital because of fever and pancytopenia. On admission, her bone marrow nuclear cell count was 45,000/microliter, being mostly blasts with cleaved nuclei. The leukemic cells were negative for peroxidase staining, expressed CD10, CD19, CD34 and sIg mu, and did not express sIg kappa and lambda, corresponding to a minor subpopulation of B cells known as transitional pre-B-cells (TPBs). Since TPB-ALL has been reported only infrequently, the incidence and clinical picture of this rare type of ALL are still uncertain. Extensive immunophenotypic studies to determine the expression of sIg mu, sIg kappa and lambda will provide accurate diagnosis, which is essential for effective management of this condition. PMID- 11280924 TI - [Therapy-related myeloid leukemia following platinum-based chemotherapy for ovarian cancer]. AB - A 40-year-old woman, who had suffered from AML (M1) in 1983, developed ovarian cancer (stage IIIc) in December 1996 after long-term remission. She underwent surgical resection of the cancer, 10 courses of standard chemotherapy and tandem PBSCT (total dose: CBDCA 6,750 mg, CDDP 200 mg, CPA 16,000 mg, THP-ADR 450 mg). After receiving the last course of chemotherapy in June 1998, she was referred to our hospital in September 1998 because of pancytopenia. Laboratory findings showed pancytopenia with 34% leukemic cells, which were positive for alpha NBE and negative for POX and CAE. Surface-marker analysis of the leukemic cells showed positivity for CD11c, CD33, CD56, and DR, and chromosome analysis revealed 47, XX, +8. The patient was diagnosed as having AML (M5a), and received induction therapy consisting of IDR and Ara-C, which led to complete remission. As she had not received etoposide, this case was thought to have been therapy-related leukemia due to the platinum agents used for treating the ovarian cancer. PMID- 11280925 TI - [The microglial activation and the expression of heat shock protein 27 through the propagation pathway of kainic acid-induced hippocampal seizure in the rat]. AB - We studied activation of microglia and expression of the 27 kDa heat shock protein (HSP27) in the brain during kainic acid-induced acute hippocampal seizures in rats. The microglial activation was observed at 6 hrs after seizure induction, but the expression of HSP27 was delayed until 3 days after seizure induction. The gross anatomical distributions of the two phenomena in the brain structures were almost identical, being localized not only in the primary focus at the dorsal hippocampus ipsilateral to the kainic acid injection, but also in selected remote brain structures that was highly consistent with the propagation pathways of the hippocampal seizure as detected previously by metabolic mapping. These structures included the hippocampus, amygdala, entorhinal cortex, piriform cortex, sensorimotor cortex, hypothalamus and thalamus. A close observation, however, revealed a difference in distribution of the two phenomena in the layers of the contralateral hippocampus: The HSP27 expression showed a layer-specific distribution, being localized selectively in the molecular layer and hilus of the dentate gyrus, and the radiatum and molecular layers of the CA-3 subfield suggesting the expression in the neuropil. On the other hand, the distribution of the microglial activation was non-specific to the layers, being scattered in the whole regions of the dorsal hippocampus. There were no apparent morphological changes in the neurons in these structures except for the ipsilateral dorsal hippocampus, by light microscopic examinations with hematoxylin-eosin staining. These findings thus indicate that activation of microglial cells and expression of HSP27 occur transsynaptically by epileptic activities through the propagation pathways of hippocampal seizure and suggest that these phenomena may reflect a part of early microenvironmental alterations in epileptic brain. PMID- 11280926 TI - Reward deficiency syndrome: a biogenetic model for the diagnosis and treatment of impulsive, addictive, and compulsive behaviors. AB - The dopaminergic system, and in particular the dopamine D2 receptor, has been implicated in reward mechanisms. The net effect of neurotransmitter interaction at the mesolimbic brain region induces "reward" when dopamine (DA) is released from the neuron at the nucleus accumbens and interacts with a dopamine D2 receptor. "The reward cascade" involves the release of serotonin, which in turn at the hypothalmus stimulates enkephalin, which in turn inhibits GABA at the substania nigra, which in turn fine tunes the amount of DA released at the nucleus accumbens or "reward site." It is well known that under normal conditions in the reward site DA works to maintain our normal drives. In fact, DA has become to be known as the "pleasure molecule" and/or the "antistress molecule." When DA is released into the synapse, it stimulates a number a DA receptors (D1-D5) which results in increased feelings of well-being and stress reduction. A consensus of the literature suggests that when there is a dysfunction in the brain reward cascade, which could be caused by certain genetic variants (polygenic), especially in the DA system causing a hypodopaminergic trait, the brain of that person requires a DA fix to feel good. This trait leads to multiple drug-seeking behavior. This is so because alcohol, cocaine, heroin, marijuana, nicotine, and glucose all cause activation and neuronal release of brain DA, which could heal the abnormal cravings. Certainly after ten years of study we could say with confidence that carriers of the DAD2 receptor A1 allele have compromised D2 receptors. Therefore lack of D2 receptors causes individuals to have a high risk for multiple addictive, impulsive and compulsive behavioral propensities, such as severe alcoholism, cocaine, heroin, marijuana and nicotine use, glucose bingeing, pathological gambling, sex addiction, ADHD, Tourette's Syndrome, autism, chronic violence, posttraumatic stress disorder, schizoid/avoidant cluster, conduct disorder and antisocial behavior. In order to explain the breakdown of the reward cascade due to both multiple genes and environmental stimuli (pleiotropism) and resultant aberrant behaviors, Blum united this hypodopaminergic trait under the rubric of a reward deficiency syndrome. PMID- 11280929 TI - Design and implementation of an introductory course for computer applications in molecular genetics. A case study. AB - Formal training in computational biology was initiated at Wayne State University in 1990 to meet the needs of the faculty. This was still at a time when the molecular databases and analysis tools could be housed in what is now equivalent to a modern but dated desktop computer. In 1995 the course was expanded to include graduate students to provide these senior students with a foundation in computational biology. This course has armed our students with a requisite set of basic skills that are necessary for a successful career in molecular genetics. It is now an integral component of the graduate program of the Center for Molecular Medicine and Genetics and our experiences in course delivery have been detailed (BioInformatics Methods and Protocols, S. Misener and S. A. Krawetz, eds., Humana Press, Totowa, NJ, 2000.). The course was expanded to a campus-wide unlimited enrollment program for the summer of 2000 to address the needs of our student body. In this review we present our experience with delivering a multidisciplinary campus-wide computational biology course to a new and widely diverse student body. PMID- 11280927 TI - Modifications of the histone N-terminal domains. Evidence for an "epigenetic code"? AB - A multicellular organism is made up of a variety of different cell types and tissues. This organization is accomplished by a well-concerted action of different regulatory molecules, which--in a very hierarchical manner--influence the expression of certain cell-specific genes. Many of those regulators are transcription factors, which directly influence the expression of the controlled gene by binding to a specific DNA sequence within its promoter or enhancer region. This binding then leads to an enhancement or a decrease in the rate of transcription of that particular gene and eventually regulates the production of the corresponding polypeptide. One major obstacle to the binding of these transcription factors is the fact that DNA is not readily accessible in the eukaryotic nucleus. It is associated with a class of very basic proteins called histones. This complex of histones and DNA is called chromatin. PMID- 11280930 TI - Vaccine trials. AB - This article reviews some of the issues involved in evaluating vaccines in humans. Vaccine trials are required for licensure and are essential for demonstrating a vaccine's safety and protective efficacy. The formal framework of phase I, II, and III trials is described, with particular emphasis on the choice of hypotheses, trial design, and biases that arise in the context of vaccine trials. However, some aspects of a vaccine's performance cannot be evaluated in clinical trials owing to their relatively small size. Thus, vaccine evaluation must continue after licensure, for example, to evaluate the vaccine with respect to rare reactions, duration of protection, and ecological effects. The article reviews some of the methods commonly used for post-licensure studies of vaccine efficacy and safety. PMID- 11280928 TI - Cowpea mosaic virus as a vaccine carrier of heterologous antigens. AB - The plant virus, cowpea mosaic virus (CPMV), has been developed as an expression and presentation system to display antigenic epitopes derived from a number of vaccine targets including infectious disease agents and tumors. These chimeric virus particles (CVPs) could represent a cost-effective and safe alternative to live replicating virus and bacterial vaccines. A number of CVPs have now been generated and their immunogenicity examined in a number of animal species. This review details the humoral and cellular immune responses generated by these CVPs following both parenteral and mucosal delivery and highlights the potential of CVPs to elicit protective immunity from both viral and bacterial infection. PMID- 11280931 TI - Rapid extraction and purification of environmental DNA for molecular cloning applications and molecular diversity studies. AB - A rapid method for the extraction and purification of DNA from environmental samples for molecular cloning applications was developed. The indigenous cells from plant debris, organic materials, sediments, and soils were lysed directly by using DAS-IZ solution and the nucleic acids were precipitated with isopropanol. A simple purification step using DAS-IIZ solution without binding matrix produced highly pure, colorless and undegraded DNA with molecular weight of more than 20 kb. The superiority of this method was tested for wide applications in molecular cloning, i.e., construction of genomic library by using Lambda DASHII Vector and GigapackIII XL, plasmid library, cloning of gene encoding protease, and molecular microbial diversity analysis. An additional advantage of this method is that only 0.1 g of sample is required, if analysis of many samples in short time should be done. To extract large amounts of environmental DNA for molecular cloning lasts only 30 min and to purify it less than 1 h. PMID- 11280932 TI - Expression of green or red fluorescent protein (GFP or DsRed) linked proteins in nonmuscle and muscle cells. AB - The introduction of the green fluorescent protein (GFP) plasmids that allow proteins and peptides to be expressed with a fluorescent tag has had a major impact on the field of cell biology. It has enabled the dynamics of a wide variety of proteins to be analyzed that could not otherwise be detected in live cells. Transient transfections of muscle and nonmuscle cells with plasmids encoding various cytoskeletal proteins ligated to green fluorescent protein or Ds red protein allow changes in the cytoskeletal network to be studied in the same cell for time periods up to several days. With this approach, proteins that could not be purified and directly labeled with fluorescent dyes and microinjected into cells can now be expressed and visualized in a wide variety of cells. Procedures are presented for transfection of the nonmuscle cell, PtK2, and primary cultures of embryonic chick myocytes, and for studying the live transfected cells. PMID- 11280933 TI - Advantages and disadvantages of using PCR techniques to characterize transgenic plants. AB - The polymerase chain reaction (PCR) revolutionized molecular biology to a similar extent as the discovery of plasmids and restriction endonucleases. However, there are some limitations to the use of PCR. Transgenic plants containing potato spindle tuber viroid (PSTVd) cDNA constructs, demonstrated to become de novo methylated upon PSTVd infection, represent a good example to illustrate the advantages of PCR. PSTVd is a 359 nt long autonomously replicating plant pathogenic RNA where all of its enzymatic requirements are entirely provided by the host cell. In addition, viroids that propagate without a DNA intermediate barely tolerate nucleotide substitutions of their RNA genome without losing infectivity. PCR is the method of choice to characterize the sequence context of genome-integrated viroid cDNA or of reverse transcribed PSTVd RNA, and can hardly be replaced by any alternative procedure. Furthermore, the precise examination of DNA methylation patterns (genomic sequencing) is entirely dependent on PCR. In contrast, the use of PCR is critical for the determination of copy number and arrangement of transgene constructs. Here, the advantages and disadvantages of PCR are discussed and protocols for PCR amplification of cDNA, genomic DNA, and bisulfite-treated DNA from transgenic plants are presented. PMID- 11280934 TI - Genotyping methodologies. AB - With current advances in genetics it is now possible to routinely screen the entire genome of multiple affected individuals in the search for disease predisposition genes. Such a large-scale undertaking requires some careful management of both samples and data in order to make best use of all available information. Here we have detailed the two main approaches to a genome-wide search and the best ways we have found of storing, transforming, and analyzing the subsequently produced data, as well as some general considerations to enhance the chances of success. PMID- 11280935 TI - Affective style, psychopathology, and resilience: brain mechanisms and plasticity. AB - The brain circuitry underlying emotion includes several territories of the prefrontal cortex (PFC), the amygdala, hippocampus, anterior cingulate, and related structures. In general, the PFC represents emotion in the absence of immediately present incentives and thus plays a crucial role in the anticipation of the future affective consequences of action, as well as in the persistence of emotion following the offset of an elicitor. The functions of the other structures in this circuit are also considered. Individual differences in this circuitry are reviewed, with an emphasis on asymmetries within the PFC and activation of the amygdala as 2 key components of affective style. These individual differences are related to both behavioral and biological variables associated with affective style and emotion regulation. Plasticity in this circuitry and its implications for transforming emotion and cultivating positive affect and resilience are considered. PMID- 11280936 TI - Making a good decision: value from fit. AB - The classic answer to what makes a decision good concerns outcomes. A good decision has high outcome benefits (it is worthwhile) and low outcome costs (it is worth it). I propose that, independent of outcomes or value from worth, people experience a regulatory fit when they use goal pursuit means that fit their regulatory orientation, and this regulatory fit increases the value of what they are doing. The following postulates of this value from fit proposal are examined: (a) People will be more inclined toward goal means that have higher regulatory fit, (b) people's motivation during goal pursuit will be stronger when regulatory fit is higher, (c) people's (prospective) feelings about a choice they might make will be more positive for a desirable choice and more negative for an undesirable choice when regulatory fit is higher, (d) people's (retrospective) evaluations of past decisions or goal pursuits will be more positive when regulatory fit was higher, and (e) people will assign higher value to an object that was chosen with higher regulatory fit. Studies testing each of these postulates support the value from-fit proposal. How value from fit can enhance or diminish the value of goal pursuits and the quality of life itself is discussed. PMID- 11280937 TI - Core knowledge. AB - Compex cognitive skills such as reading and calculation and complex cognitive achievements such as formal science and mathematics may depend on a set of building block systems that emerge early in human ontogeny and phylogeny. These core knowledge systems show characteristic limits of domain and task specificity: Each serves to represent a particular class of entities for a particular set of purposes. By combining representations from these systems, however, human cognition may achieve extraordinary flexibility. Studies of cognition in human infants and in nonhuman primates therefore may contribute to understanding unique features of human knowledge. PMID- 11280938 TI - Unraveling the mysteries of anxiety and its disorders from the perspective of emotion theory. AB - The ascendance of emotion theory, recent advances in cognitive science and neuroscience, and increasingly important findings from developmental psychology and learning make possible an integrative account of the nature and etiology of anxiety and its disorders. This model specifies an integrated set of triple vulnerabilities: a generalized biological (heritable) vulnerability, a generalized psychological vulnerability based on early experiences in developing a sense of control over salient events, and a more specific psychological vulnerability in which one learns to focus anxiety on specific objects or situations. The author recounts the development of anxiety and related disorders based on these triple vulnerabilities and discusses implications for the classification of emotional disorders. PMID- 11280939 TI - Behavioral health care dot-com and beyond: computer-mediated communications in mental health and substance abuse treatment. AB - Computer-mediated communications (CMC) such as e-mail, websites, and CD-ROM and DVD programs, both on and off the Internet, will play an ever larger role in the future of behavioral health care. This address describes the current rapid expansion of CMC and some profound changes that are likely in the future. The reasons for using such CMC programs are addressed, and an example of a new multimedia version of the Addiction Severity Index is described. Issues and concerns about future uses of CMC are raised, as are possible implications for psychology. PMID- 11280940 TI - Reflections on receiving an award. AB - Reflecting on the events that culminated in her receiving the Award for Distinguished Contributions to Applied Psychology as a Professional Practice, Canter, an independent practitioner, discusses the road she traveled to become a clinical psychologist and to become involved in professional organizational activities. She believes that this award was given to her because of her contributions to psychology over her lifetime as an effective and hardworking leader, mentor, and role model in her home state of Arizona and nationally. She addresses some of her ideas about effective leadership and mentoring in the American Psychological Association (APA), providing many examples from which she has learned. Canter also shares some thoughts about APA's position as a leader in the development and enforcement of professional ethics. PMID- 11280941 TI - An ethnopolitical approach to working with people of color. AB - North Americans have been expected to abdicate their ethnic backgrounds and blend into a single homogeneous identity. However, the United States President's Initiative on Race (1998) concluded that the greatest challenge facing North Americans is to accept and take pride in defining themselves as a multiracial democracy. With an ethnopolitical approach, the author studies effects of oppression, racism, and political repression on individuals, groups, and societies. She concludes that psychologists can help ameliorate racism in society by taking an antiracist stance, promoting a safe society where racial-social equity and justice prevail, and helping to formulate a collective identity that affords freedom to all members of society. PMID- 11280942 TI - Blame, shame, and community: justice responses to violence against women. AB - Justice processing for crimes against women is reviewed. The data reveal conviction rates for partner violence and rape by known acquaintances are miniscule; mandatory arrest, protection orders, and diversion programs inadequately deter rebattering; few losses are compensated; and the adversarial justice process is retraumatizing, exacerbating survivor's self-blame. To better address crimes against women, several nations and tribal communities use communitarian approaches, forms of restorative justice. The offense is framed to include the perpetrator, victim, and community. The process forgoes incarceration to have family, peers, and advocates design perpetrator rehabilitation, victim restoration, and social reintegration of both victim and perpetrator. Evaluations suggest communitarian justice may increase victim satisfaction, raise the social costs of offending, multiply social control and support resources, and open a new avenue to targeted prevention. PMID- 11280943 TI - Crafting usable knowledge. AB - Current emphasis on evaluating interventions does not address the problems of dissemination and utilization of these interventions, particularly in complex settings such as schools. Research on interventions is of value, but its generalizability to specific contexts is limited. Further, little is known about actual use of empirically supported interventions in practice settings. These concerns suggest the following: (a) There is a need to examine the dissemination process, including practitioner education and the development of a consumer information mindset by researchers; (b) guidance about selecting interventions would benefit from a systematic problem-solving orientation; and (c) research training and methodology need to be augmented with strategies and techniques suitable for developing an empirical approach to practice. These issues are addressed with specific examples drawn from school-based practice. PMID- 11280944 TI - Improving the lives of millions of school children. AB - R. Fulghum (1993) contended that all he really needed to know, he learned in kindergarten. This finding does not generalize to predoctoral-postdoctoral education and school-based action researchers. After 20 years, the author's current aspiration is to collaborate with others to disseminate research-based social and emotional learning programs that improve the lives of millions of school children. High-quality graduate and postdoctoral training inspired this commitment. However, realistically speaking, ongoing early- and midcareer training are needed to achieve it. This author describes 2 sets of advanced learning experiences that strongly affected his current activities. He draws implications from these experiences to encourage more support for the early- and midcareer training of school-based action researchers. The address also highlights several priorities that midcareer scientists should address to advance the positive impact that psychology has on the lives of children. PMID- 11280945 TI - Establishing linkages through international psychology: dealing with universalities and uniquenesses. AB - This article chronicles one individual's personal odyssey toward becoming an international psychologist--and hopefully a citizen of a professional world that has no geographic borders. It is written as a narrative that is divided into chronological stages and several domains of professional activities. From each period, the main observations and lessons learned are extrapolated. It is hoped that readers who are interested in teaching, conducting workshops, and doing clinical work or research in other countries will find some of the personal reflections and professional ideas presented to be thought provoking and useful in the development of guiding principles in international psychology. PMID- 11280946 TI - A diagnostic approach to identifying submicroscopic 7p21 deletions in Saethre Chotzen syndrome: fluorescence in situ hybridization and dosage-sensitive Southern blot analysis. AB - PURPOSE: To report on the use of fluorescence in situ hybridization (FISH) and dosage-sensitive Southern blot analysis in the molecular diagnosis of patients with Saethre-Chotzen syndrome. METHODS: FISH and dosage-sensitive Southern blot analysis utilizing TWIST gene probes were performed on patients with Saethre Chotzen syndrome but without an identifiable TWIST sequence variation. RESULTS: Four unrelated patients with a deletion of the TWIST gene were identified by Southern blot; one of them had a complex chromosomal rearrangement involving 7p21 and no apparent deletion by FISH, suggesting a smaller deletion in the region including the TWIST gene. A fifth patient had an abnormal TWIST gene fragment on Southern blot analysis that segregated with the disease in the family; FISH was normal in this patient, suggesting a partial deletion or rearrangement in or near the gene. CONCLUSION: FISH and dosage-sensitive Southern blot analysis are useful diagnostic tools in Saethre-Chotzen syndrome without TWIST sequence variation. PMID- 11280947 TI - Hemochromatosis-associated morbidity in the United States: an analysis of the National Hospital Discharge Survey, 1979-1997. AB - PURPOSE: The recent discovery of the HFE gene and its association with hereditary hemochromatosis has renewed the attention directed to iron-overload diseases. Population screening for hereditary hemochromatosis is under debate, and population-based estimates of morbidity associated with hereditary hemochromatosis are needed. The purpose of this study is to estimate the number of hemochromatosis-associated hospitalizations in the United States using a population-based dataset. METHODS: National Hospital Discharge Survey and census data were used to estimate hemochromatosis-associated hospitalization rates for persons 18 years of age and over. RESULTS: From 1979 through 1997, the rate of hemochromatosis-associated hospitalizations was 2.3 per 100,000 persons in the United States. The rate among persons 60 years of age and over increased more than 60% during this time period. CONCLUSION: The increase in the rate of hereditary hemochromatosis-associated hospitalizations among older persons is consistent with recent trends in mortality data and may reflect the rising awareness of iron-overload disorders in the United States. PMID- 11280948 TI - Genetic testing for breast cancer: where are health care providers in the decision process? AB - PURPOSE: To identify BRCA1/2 knowledge, genetic testing intentions, and communication patterns in breast cancer survivors (survivors). METHODS: A population-based survey was conducted of 276 female survivors diagnosed between the ages of 40 and 49 and living 5 to 10 years postdiagnosis. RESULTS: Of the 79% who responded, 8% spoke with health care providers and 53% spoke with relatives about testing. Few (26%) correctly answered over half the BRCA knowledge questions. Intention to obtain testing varied (26-67%), depending on insurance coverage. CONCLUSION: Health care providers and survivors seldom discuss BRCA testing. Providing information to survivors would increase their ability to make informed testing decisions. PMID- 11280949 TI - Oncologists' opinions on genetic testing for breast and ovarian cancer. AB - PURPOSE: To determine oncologists' practices and beliefs about genetic testing for hereditary breast and ovarian cancer and the extent to which oncologists are utilizing clinical genetics services. METHODS: A survey was mailed to oncologists who treat adult patients in Washington, Oregon, Idaho, or Alaska. RESULTS: Most oncologists (79%) had discussed genetic tests with their patients, and 76% indicated they would like patients considering genetic testing to consult with a genetic counselor. Yet few (19%) indicated their medical practice had the necessary services and staff to offer genetic testing, and only 11% had made referrals to medical genetics or genetic counselors. CONCLUSION: Most respondents support the use of genetic services, but few have made referrals to genetic counselors. Increased communication between oncologists and genetic counselors may enhance collaboration between these two disciplines. PMID- 11280950 TI - Partial trisomy 7p defined by analysis of a complex chromosome rearrangement using a BAC clone panel. AB - PURPOSE: To illustrate the use of bacterial artificial chromosome (BAC) clone panels for molecular cytogenetic analysis of complex chromosome rearrangements (CCRs). METHODS: High resolution cytogenetics followed by fluorescence in situ hybridization (FISH) analysis using chromosome band-specific BAC probes, in addition to commercially available probes. RESULTS: High resolution cytogenetics in conjunction with FISH using commercially available probes proved inadequate to resolve problems in characterizing a balanced CCR in the mother of a patient who had inherited an unbalanced form of the CCR. Accurate interpretation of the CCR and the unbalanced rearrangement in the patient as trisomy 7p12.2-->p21.3 was accomplished only through use of the BAC clone panel. CONCLUSION: Use of BAC clone panels can enhance the power of FISH analysis in defining chromosome rearrangements that cannot be resolved by high resolution chromosome analysis. PMID- 11280952 TI - Laboratory standards and guidelines for population-based cystic fibrosis carrier screening. PMID- 11280953 TI - Recommendations of core competencies in genetics essential for all health professionals. PMID- 11280951 TI - American College of Medical Genetics consensus statement on factor V Leiden mutation testing. PMID- 11280954 TI - Cystic fibrosis population carrier screening: here at last--are we ready? PMID- 11280957 TI - An update on obsessive-compulsive disorder. AB - It is now recognized that obsessive-compulsive disorder (OCD) affects almost 3% of the world's population and is a major worldwide health problem. Much has been learned in the past 2 decades about the treatment of these disorders. Recent developments in neuroimaging techniques have led to a better understanding of the biology of OCD and the brain circuits that may be involved in the production of symptoms. The most effective treatment approaches, based on controlled data, are behavior therapy consisting of exposure and response prevention and specific medications. Although controversial, more invasive neurosurgical and neurostimulation approaches may hold some promise for severely disabled patients. Despite impressive research efforts, a small minority of patients remain refractory to treatment. Future clinical research should focus on this refractory group. PMID- 11280956 TI - Treatment-resistant bipolar disorder. AB - Over the last few years, the number of potential pharmacotherapies for bipolar disorder has greatly expanded. Yet the database for virtually all these newer treatments consists of case reports and case series. Among these newer treatments, recently released anticonvulsants are most promising. Lamotrigine has already shown efficacy for treating bipolar depression, while gabapentin's efficacy has yet to be documented in a controlled study. Alone among its medication class, topiramate, another anticonvulsant, is associated with weight loss. Novel antipsychotics are effective in treating acute mania. With the exception of clozapine, their efficacy as true mood stabilizers is still unknown. Utilizing combinations of mood stabilizers is common and appropriate but demands knowledge of potential pharmacokinetic interactions. Other approaches for treatment resistant bipolar disorder include high-dose thyroid hormones, calcium channel blockers, electroconvulsive therapy, and omega-3 fatty acids. Finally, the efficacy of adjunctive psychosocial strategies is a topic of active investigation. PMID- 11280955 TI - Down syndrome congenital heart disease: a narrowed region and a candidate gene. AB - PURPOSE: Down syndrome (DS) is a major cause of congenital heart disease (CHD) and the most frequent known cause of atrioventricular septal defects (AVSDs). Molecular studies of rare individuals with CHD and partial duplications of chromosome 21 established a candidate region that included D21S55 through the telomere. We now report human molecular and cardiac data that narrow the DS-CHD region, excluding two candidate regions, and propose DSCAM (Down syndrome cell adhesion molecule) as a candidate gene. METHODS: A panel of 19 individuals with partial trisomy 21 was evaluated using quantitative Southern blot dosage analysis and fluorescence in situ hybridization (FISH) with subsets of 32 BACs spanning the region defined by D21S16 (21q11.2) through the telomere. These BACs span the molecular markers D21S55, ERG, ETS2, MX1/2, collagen XVIII and collagen VI A1/A2. Fourteen individuals are duplicated for the candidate region, of whom eight (57%) have the characteristic spectrum of DS-CHD. RESULTS: Combining the results from these eight individuals suggests the candidate region for DS-CHD is demarcated by D21S3 (defined by ventricular septal defect), through PFKL (defined by tetralogy of Fallot). CONCLUSIONS: These data suggest that the presence of three copies of gene(s) from the region is sufficient for the production of subsets of DS-CHD. This region does not include genes located near D21S55, previously proposed as a "DS critical region," or the genes encoding collagens VI and XVIII. Of the potential gene candidates in the narrowed DS-CHD region, DSCAM is notable in that it encodes a cell adhesion molecule, spans more than 840 kb of the candidate region, and is expressed in the heart during cardiac development. Given these properties, we propose DSCAM as a candidate for DS-CHD. PMID- 11280958 TI - Psychodynamic psychotherapy of borderline personality disorder: a contemporary approach. AB - Recent trends in the economics of mental health care threaten to undermine the use of psychodynamic psychotherapy for the treatment of patients with borderline personality disorder. These trends are driven in part by the assumption that such treatment of these challenging patients is very expensive. The author highlights empirical research that supports both the usefulness and the cost-effectiveness of this treatment approach. He also reviews some effective clinical strategies with borderline patients. PMID- 11280959 TI - Cognitive biases in anxiety disorders and their effect on cognitive-behavioral treatment. AB - Cognitive theorists hypothesize that cognitive biases are a major component in the development and maintenance of anxiety disorders. These include attentional biases toward threat-related information, distorted judgments of risk, and selective memory processing. The empirical evidence for these cognitive biases in anxiety disorder populations is reviewed. Potential deleterious effects of these biases on the process of cognitive-behavioral therapy are also discussed, as are possible ways of overriding those effects and maximizing treatment efficacy. PMID- 11280960 TI - Virtual reality: using the virtual world to improve quality of life in the real world. AB - Mental health professionals are increasingly integrating advances in technology to improve the health of those in their care (American Psychological Association, 2000). The authors describe the immersive properties of virtual reality and its importance for clinical purposes and then review the literature describing current clinical applications of virtual reality (VR) and research documenting its efficacy. Virtual reality has been used in the treatment of specific phobias, posttraumatic stress disorder, eating disorders, and pain management. PMID- 11280961 TI - A relational-cultural model: healing through mutual empathy. AB - Relational-cultural theory offers an alternative to traditional theories of psychological development. Whereas traditional theories view mature functioning as characterized by movement from dependence to independence, relational-cultural theory suggests that maturity involves growth toward connection and relationship throughout the life span. After contrasting these two theoretical perspectives, the author describes a therapeutic approach based on the relational-cultural model, which involves mutual empathy and working with shame. A case example illustrates this approach. The author suggests that the relational-cultural model has applications at both the personal and societal levels. PMID- 11280962 TI - Developmental and lesion effects in brain activation during sentence comprehension and mental rotation. AB - The development of neurocognitive networks was examined in 2 cognitive paradigms: auditory sentence comprehension and mental rotation of alphanumeric stimuli. Patterns of brain activation were measured with whole brain echoplanar functional magnetic resonance imaging at 3 Tesla in 5 adults (20-28 years old), 7 children (9-12 years old), and 6 pediatric patients (9-12 years old) with perinatal strokes or periventricular hemorrhages. Healthy children and adults activated similar neurocognitive networks, but there were developmental differences in the distribution of activity across these networks. In the sentence task, children showed more activation in the inferior visual area suggesting an imagery strategy rather than a linguistic strategy for sentence processing. Furthermore, consistent use of a sentence comprehension strategy, whether correct or incorrect as compared to chance performance, was associated with greater activation in the inferior frontal area (Broca's) in both children and pediatric patients. In the mental rotation task, healthy adults showed more activation in the superior parietal and middle frontal areas and less activation in the supramarginal gyrus, suggesting adults were primarily engaged in visual-spatial manipulation and less engaged in the recognition of noncanonical views of stimuli. The pediatric patients showed patterns of activation consistent with organization of cognitive processing into homologous areas of the contralateral hemisphere. PMID- 11280963 TI - Bone lead levels and language processing performance. AB - The relation between bone lead absorption and language processing abilities in 156 randomly selected 11- to 14-year-old boys who were asymptomatic for lead toxicity is examined. Tibial lead concentrations were measured by X-ray fluorescence spectroscopy. The language processing outcome variables consisted of the least and most difficult subtests from the Nonword Repetition Task, Competing Language Processing Task, and the Revised Token Test. Participants were classified by quartiles according to bone lead concentrations, and analysis of variance and analysis of covariance measured the impact on language processing scores. Results showed that children in the highest bone lead quartile displayed decreased language processing performance on the most difficult language processing tasks but not on the easier tasks. PMID- 11280964 TI - An event-related potential study of older children with an early history of failure to thrive. AB - Elementary and junior high school children (n = 13), who were diagnosed with nonorganic failure to thrive (FTT) as infants and toddlers, were compared with a normal control group (n = 14) on visual event-related potentials (ERPs) elicited during a primed lexical decision task. Positive stimuli were real words that were identical to the priming stimuli; negative stimuli were nonpronounceable letter strings. Although the groups did not differ in word-list reading level, the former FTT group had slower reaction (decision) times and did not show ERP evidence of priming in the N400 epoch. Anterior sites yielded better separation of the real words and letter strings than posterior sites. A late anterior component between 500 msec to 650 msec poststimulus onset showed the largest condition effect for both groups. The control group had a larger negative going late anterior component to words than the FTT group. The combined reaction time and ERP findings point to less automatized word recognition in the FTT group. PMID- 11280965 TI - Drawing abilities in Williams syndrome: a case study. AB - Children with Williams syndrome (WS) have been reported to exhibit an unusual cognitive profile characterized by marked preservation of linguistic abilities and poor visuospatial abilities against a backdrop of generalized mental retardation. Much of the data documenting this profile come from studies of older children and adults with WS. Very few studies have reported findings from the preschool and early school-age period. As a result, little is known about the early development of cognitive processes in children with WS. Capirci, Sabbadini, and Volterra (1996) reported data from a longitudinal case study of early language development in a young child with WS. This article presents the longitudinal profile of visuospatial abilities in this same child. Data on copying and free drawing collected over a period extending from late preschool to early school age are reported. It is clear from these data that this child does indeed exhibit deficits in visuospatial abilities. Her performance clearly improved with age, but deficits persist. PMID- 11280966 TI - Neural plasticity and cognitive development. AB - It has been well documented that the effects of early occurring brain injury are often attenuated relative to later occurring injury. The traditional neuropsychological account of these observations is that, although the developing neural system normally proceeds along a well-specified maturational course, it has a transient capacity for plastic reorganization that can be recruited in the wake of injury. This characterization of early neural plasticity is limited and fails to capture the much more pervasive role of plasticity in development. This article examines the role of neural plasticity in development and learning. Data from both animal and human studies show that plasticity plays a central role in the normal development of neural systems allowing for adaptation and response to both exogenous and endogenous input. The capacity for reorganization and change is a critical feature of neural development, particularly in the postnatal period. Subtractive processes play a major role in the shaping and sculpting of neural organization. However, plasticity is neither transient nor unique to developing organisms. With development, neural systems stabilize and optimal patterns of functioning are achieved. Stabilization reduces, but does not eliminate, the capacity of the system to adapt. As the system stabilizes, plasticity becomes a less prominent feature of neural functioning, but it is not absent from the adult system. The implications of this broader view of plasticity for our understanding of development following early brain damage are discussed. PMID- 11280967 TI - Associations between altered immune function and organochlorine contamination in young Caspian terns (Sterna caspia) from Lake Huron, 1997-1999. AB - Previous studies of laboratory animals and wildlife species have demonstrated the immunotoxicity of organochlorines. This study confirmed that associations between organochlorines and suppressed T cell function and enhanced antibody production in young Caspian terns from the Great Lakes, first observed in the early 1990s, continued into the late 1990s. These associations were based on measurement of organochlorines in plasma of individuals and pooled egg samples. During 1997-99, immune function, hematological variables, and organochlorine contamination were measured in prefledgling Caspian terns at two Lake Huron colonies: Channel Shelter Island (Confined Disposal Facility) at the mouth of the Saginaw River in southern Saginaw Bay and Elm Island in the North Channel. Elevated organochlorine exposure, reproductive effects, and decreased recruitment have been documented previously in the Saginaw Bay colony. Concentrations of polychlorinated biphenyls (PCBs) in eggs and plasma and 1,1-dichloro-2,2-bis(p)chlorophenyl)ethylene (DDE) in plasma were consistently higher in Saginaw Bay compared to the North Channel. The mean phytohemagglutinin (PHA) skin test, a measure of T lymphocyte function, was 42% lower in Saginaw Bay. Regression analyses showed strong negative associations between the PHA response and plasma PCBs and, to a slightly lesser degree, DDE. Despite interyear differences, total antibody titers following immunization with sheep red blood cells were higher in Saginaw Bay than the North Channel. Titers were positively associated with plasma PCBs and DDE. Plasma PCBs and DDE were negatively correlated with the percentage of monocytes and positively correlated with the percentage of basophils. PMID- 11280968 TI - DNA fingerprint comparison of rainbow trout and RTG-2 cell line using random amplified polymorphic DNA. AB - The detection of genotoxic effects using in vitro cell systems can be extremely useful in risk assessment procedures. However, care should be taken in the extrapolation of in vitro results since, amongst other factors, established cell lines may deviate from the genetic characteristics of their species. In this work, the genetic similarities between the RTG-2 cell line and rainbow trout individuals (Oncorhynchus mykiss) from several fish farms have been studied by the RAPD technique. Results show a significant analogy in the band patterns obtained for both systems, up to 73% of the bands composing the fingerprint of the RTG-2 cell line were found in all the individuals analysed. The inter population similarity index (Lynch, 1990), considering the RTG-2 cell line as a population, gives a value of 0.931 between both systems. The dendrogram constructed from all the individuals, considering the RTG-2 cell line as just another individual of a single population, showed that the genetic structure of the cell line was not different from those of the other individuals tested. The strong genetic similarity of both systems, together with the previously proven capability of the RAPD technique to detect genetic alterations caused in vitro by genotoxic agents, can be very useful in genetic ecotoxicological studies. PMID- 11280969 TI - Aflatoxin contamination in supplemental and wild foods of northern bobwhite. AB - Esophagi were removed from northern bobwhite (Colinus virginianus) that were killed by hunters during the 1996-1997 (n = 39) and 1997-1998 (n = 27) hunting seasons in Wheeler County, Texas and Roger Mills County, Oklahoma to determine if they were exposed to aflatoxin (AF) by consuming either wild or supplemental food. Esophagi were segregated into three categories based upon their contents: all wild seeds (n = 11), all supplemental foods (n = 21), and mixed foods (n = 18). Contents of esophagi were then analyzed for AF concentration. Mean (+/- SE) AF concentration (ppb) of wild seeds was 2.44 +/- 0.54; supplemental foods, 0.12 +/- 0.41; and mixed foods, 0.53 +/- 0.40. Wild seeds had higher (P = 0.004) AF concentrations than either the supplemental or mixed categories, although these levels are below those found to cause damage to northern bobwhite. This information suggests that northern bobwhite may consume contaminated food much more often than previously thought. PMID- 11280970 TI - Cyanide-induced alterations to the biophysical conformations of the isolated fish liver. AB - The purpose of this study is to examine the applicability of an infared spectroscopic methodology for the study of an environmental problem. The effect of cyanide concentrations on the biophysical conformation of the fish liver homogenate was determined by using an attenuated total reflectance (ATR)/Fourier transform infrared (FT-IR) microspectroscopy. Alive male model fish, Tilapia Zillii, was used. The liver from fish was isolated and homogenized in pH 8.0 Tris buffer solution. The results indicate that the IR peak intensity increased markedly in the C-H stretching range (3000-2800 cm-1), ester C = O stretching of lipids (1743 cm-1) and carbohydrate bands (1195-950 cm-1), but decreased in the amide I at 1649 cm-1 and the free asymmetric stretching band of phosphate at 1261 cm-1 with the increase of KCN concentrations. The marked release of hepatic enzymes and glutathione into homogenate induced by cyanide might account for the higher IR spectral peak intensity of fish liver tissue after treatment with KCN. The cyanide was also found to induce the protein structure of fish liver homogenate from alpha-helical conformation to beta-conformation. PMID- 11280971 TI - Exposure of California quail to organophosphorus insecticides in apple orchards in the Okanagan Valley, British Columbia. AB - We studied the exposure and effect of the organophosphate insecticides azinphos methyl and diazinon on adult California quail (Callipepla californica) in an apple orchard in the Okanagan Valley, British Columbia. Cholinesterase activity was measured in plasma samples (n = 65) collected from 54 individuals either prior to spraying, immediately (< 24 hours) or 10 days after three spray events. Mean plasma cholinesterase levels declined significantly (P < 0.05, n = 12) to 61% of pre-spray mean activity (controls) immediately following the first spray event, but by ten days had recovered to 86% of mean control activity. Subsequent spray events caused no significant declines in mean plasma cholinesterase activity. Four of the 26 quail sampled within 24 h of a spray event exhibited plasma-ChE inhibition exceeding 50% inhibition. Radio-tagged quail (n = 25) were monitored throughout the breeding season to determine use of orchards and detect changes in use patterns resulting from the spraying of insecticides. Use of orchards by quail varied over the summer, with the highest use occurring in May, declining to very low use by July. Quail exhibited a diurnal pattern, roosting in sparsely forested uplands at night, travelling to orchard areas to feed early each morning and returning to roosts at dusk. Orchard use by quail differed during spray events compared to non-spray times. During the three hour period immediately after spraying (0530-0800), 14-20% of observed quail were in the orchard, after which use declined to < 4%, and returned to 12% by the next day. During non-spray times, 3-13% of radio-tagged the quail were observed in orchard habitat, with the heaviest use (13%) occurring later in the day (0830-1700 h). Seven radio-tagged quail were predated during the study period. However, no deaths could be attributed to insecticide poisoning as carcasses were not in suitable condition for testing. It was concluded that adult quail using orchard habitat early in the summer may be acutely poisoned by anti-cholinesterase insecticides, but the risk of exposure declined over the summer. PMID- 11280972 TI - Combined effects of food concentration and the herbicide 2,4 dichlorophenoxyacetic acid on the population dynamics of Brachionus patulus (Rotifera). AB - Herbicides are important in crop protection and management. A number of them including 2,4-D (2,4-dichlorophenoxyacetic acid), however, may reach water bodies and eventually affect the non-target organisms such as rotifers. In the present work, we studied the influence of 6 concentrations viz. 0 (control), 100, 200, 300, 400 and 500 mg l-1 of 2,4-D on the population growth of the rotifer Brachionus patulus under two algal (Chlorella) food levels (0.5 x 10(6) and 1.5 x 10(6) cells ml-1). Regardless of herbicide concentration, the population growth of B. patulus was dependent on the algal food levels, in that an increase in algal food level supported a better population growth. Similarly at any Chlorella density, the herbicide had a negative influence on the population growth of B. patulus. Herbicide level of 500 mg l-1 inhibited population growth of B. patulus beyond 5 days. Rotifers grown under low food density and high herbicide concentration (300 mg l-1 or above) were completely eliminated after day 15. The rate of population increase (r) (mean +/- standard error) in the controls varied from 0.46 +/- 0.002 and 0.55 +/- 0.004 under 0.5 x 10(6) and 1.5 x 10(6) cells ml 1 of Chlorella, respectively. The r values became negative under both, low and high food levels, at or beyond 300 mg l-1 of 2,4-D. The maximal population abundance (ind. ml-1) in controls varied from 294 +/- 9 to 503 +/- 21 under low and high food levels of Chlorella, respectively. The role of algae in mitigating adverse effects of high herbicide concentrations to rotifers has been discussed. PMID- 11280973 TI - [Treatment of locally advanced or metastatic non-small cell bronchial cancer]. PMID- 11280974 TI - [Treatment of small-cell bronchial cancer]. PMID- 11280975 TI - [Pleural mesothelioma]. PMID- 11280976 TI - [Summary of the congress]. PMID- 11280977 TI - [Platinum derivatives in development]. PMID- 11280978 TI - [New targets in the treatment of lung cancer. Role of EGF-R tyrosine kinase inhibitors]. PMID- 11280979 TI - [Screening, prevention, and tobacco]. PMID- 11280980 TI - [Pre- and postoperative chemotherapy of non-small cell bronchial cancer]. PMID- 11280981 TI - [Chemoradiotherapy combination for non-small cell bronchial cancer]. PMID- 11280982 TI - Bench-scale studies on the simultaneous formation of PCBs and PCDD/Fs from combustion systems. AB - The presence of endocrine disrupting chemicals (EDCs) in the environment has wide ranging potential ecological and health impacts on animals and humans. A significant amount of experimental and theoretical work has been performed the examining formation and control of polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDD/Fs), which account for only part of the EDCs being emitted from combustion devices. Generally accepted mechanistic theories for PCDD/F formation propose heterogeneous reactions in the cooler regions of the combustor involving gas-phase organic precursors (such as chlorobenzenes or chlorophenols), a chlorine donor [such as hydrogen chloride (HCl)], and a flyash bound metallic catalyst (such as copper chloride). There is evidence that some other proposed EDCs, including polychlorinated biphenyls (PCBs), are formed through a similar mechanistic pathway as PCDD/Fs. In addition, there is evidence that certain important steps in the catalytic reaction between the copper catalyst and the organic precursors may suggest a common rate limiting step for the heterogeneous formation of the previously mentioned EDCs. This paper reports on a bench-scale experimental study to characterize a newly built reactor system that was built to: produce levels and distributions of PCDD/F production similar to those achieved by previous researchers; verify similar responses to changes in independent variables; examine the hypothesis that PCB formation rates exhibit trends similar to PCDD/F formation rates as reactor variables are changed; and begin to explore the dependence of PCB formation on temperature and precursor type. The reactor system has been built, and initial reactor characterization studies have been performed. Initial experiments yielded results that support the hypothesis of a similar formation mechanism of PCBs and PCDD/Fs in combustors. Initial experiments uncovered potential deficiencies with the reactor system and the experimental procedures and have suggested corrective action to improve the experimental system. PMID- 11280983 TI - Fundamental study of the behavior of chlorine during the combustion of single RDF. AB - A fundamental study of the combustion characteristics and the de-HCl behavior of a single refuse-derived fuel (RDF) pellet was carried out to explain the de-HCl phenomena of RDF during fluidized bed combustion and to provide data for the development of high efficiency power generation technology using RDF. In this research, combustion and pyrolysis experiments were carried out in an electrical furnace using a series of model and actual RDF samples. The de-HCl capability of Ca(OH)2 in RDF was evaluated by measuring the emission fraction of HCl in the flue gas and the capture fraction of Cl in the residue. It was found that the capture fraction of Cl components in the residue increased from 0 to nearly 70% when the molar ratio of Ca/Cl was changed from 0 to around 13. Apparently, the capture fraction also decreased with increasing oxygen concentration in the feed gas. The devolatilization process of RDF was confirmed to be a very important part of de-HCl process. The effect of temperature profile of the RDF pellet on the de-HCl process, as it varies with the heating rate of RDF and the oxygen concentration in the vicinity of the sample, is discussed. PMID- 11280984 TI - Simultaneous capture of metal, sulfur and chlorine by sorbents during fluidized bed incineration. AB - Metal capture experiments were carried out in an atmospheric fluidized bed incinerator to investigate the effect of sulfur and chlorine on metal capture efficiency and the potential for simultaneous capture of metal, sulfur and chlorine by sorbents. In addition to experimental investigation, the effect of sulfur and chlorine on the metal capture process was also theoretically investigated through performing equilibrium calculations based on the minimization of system free energy. The observed results have indicated that, in general, the existence of sulfur and chlorine enhances the efficiency of metal capture especially at low to medium combustion temperatures. The capture mechanisms appear to include particulate scrubbing and chemisorption depending on the type of sorbents. Among the three sorbents tested, calcined limestone is capable of capturing all the three air pollutants simultaneously. The results also indicate that a mixture of the three sorbents, in general, captures more metals than a single sorbent during the process. In addition, the existence of sulfur and chlorine apparently enhances the metal capture process. PMID- 11280985 TI - Melting and stone production using MSW incinerated ash. AB - Most of the municipal solid waste (MSW) in Japan is incinerated and the generated ash is landfilled. However, environmental pollution problems have increased and Japan has decreased final disposal sites for landfills. With the application of a melting system, the volume of incinerated ash can be reduced and the effective use of melted slag is being developed for use in civil engineering works. However, the low strength of melted slag as a vitreous structure has limited its effective use. As a solution for this deficiency, a technology to crystallize melted slag into higher strength produced stones was developed. With the joint cooperation of Chiba Prefecture and Kamagaya City, a demonstration plant for melting and stone production with a capacity of 4.8 tons of incinerator ash per day was constructed. The demonstration test was conducted from May 1998 to June 1999 with satisfactory results stated below. Long-term stable operation and performance of the plant have been confirmed and effective applications of produced stones have been demonstrated on a commercial scale. The results are as follows. 1. A stable, continuous operation and good quality produced stones have been confirmed by treating more than 750 tons of MSW incinerated ash. 2. More than 99.9% of dioxins contained in the incinerated ash were decomposed, and the concentration of dioxins in produced stones were less than the detection limit set by Japanese environmental standards. 3. Leaching values of hazardous heavy metals of produced stones sufficiently met the environmental standard on soil pollution of the Environment Agency with superior leaching behavior for the vitreous slag, thus confirming their safe applications. 4. The effective application of produced stones for aggregate was tested based on Japanese Industrial Standards and every figure of test results met the Japanese standard values. The use of produced stones as raw materials for permeable pavement blocks has been confirmed in commercial construction for a park in Chiba Prefecture. Asphalt use was also demonstrated by paving a commercial roadway in Kamagaya City. PMID- 11280986 TI - Evaluation of waste pyrolysis characteristics in a pressurized fluidized bed reactor. AB - To obtain the distribution of fuel components to gas, tar and char in a pressurized fluidized bed waste pyrolyzer, experiments were conducted with a laboratory scale fluidized bed reactor. Waste samples were fed batchwise from the top of the reactor into the fluidized bed of silica sand and pyrolyzed by nitrogen/nitrogen-O2 gas and the effects of pressure, particle size, heating rate and oxygen addition were investigated. In the case of rubber, the char yield tended to increase a little and the tar yield decrease over the pressure of 304 709 kPa. In comparison with the thermogravimetry data it was clearly demonstrated that the char yield from fluidized bed pyrolysis is much lower. A small amount of oxygen addition decreased both tar and char yields but its further increase did not affect them very much. PMID- 11280987 TI - Effects of flue gas composition on the catalytic destruction of chlorinated aromatic compounds with a V-oxide catalyst. AB - When using catalytic flue gas cleaning, several flue gas compounds may influence oxidation reactions of hazardous volatile organic compounds, possibly leading to lower reaction rates and, thus, to an incomplete destruction. Experimental investigations were performed with regard to the influence of selected flue gas compounds, like hydrogen chloride, sulfur dioxide, oxygen, and water vapour, on the catalytic destruction behavior of chlorobenzenes under flue gas cleaning conditions of an incineration plant. For this purpose, a metal oxide catalyst was operated at different temperatures at a space velocity of 3600 h-1 in a laboratory-scale fixed bed reactor with model flue gases, and with real flue gases generated from the TAMARA waste incineration plant. The results obtained from the studies with model flue gas were analyzed with respect to reaction kinetics. These kinetics were applied for comparison with the experimental data gained in the real flue gas. PMID- 11280988 TI - Method development and implementation for co-planar polychlorinated biphenyls (PCBs). AB - The Emission Measurement Center (EMC) in the Environmental Protection Agency's Office of Air Quality Planning and Standards was directed to conduct an emissions test program at a sewage sludge incinerator in support of a Maximum Achievable Control Technology (MACT) standard. One pollutant category of concern at these facilities was polychlorinated biphenyls, or PCBs. An objective of the test program was to measure co-planar PCBs in the incinerator emissions, the sewage sludge introduced to the incinerator, and the scrubber water effluent used in controlling the incinerator emissions. Co-planar PCB congeners are those having four or more chlorine atoms with only a few substitutions in the ortho positions, i.e. positions designated 2, 2', 6, and 6'. Thirteen PCB compounds are sometimes referred to as the "WHO PCBs," because the World Health Organization (WHO) has derived toxic equivalency factors for these congeners. Studies have shown that these dioxin-like compounds can react with the aryl hydrocarbon receptor. This same reaction is believed to initiate adverse health effects for dioxin and furan congeners. In order to conduct the co-planer PCB testing, the EMC had to develop analytical methods that could measure the 13 co-planar PCBs. The purpose of the test program was to develop, evaluate, and implement analytical test methods capable of measuring co-planar PCBs in three matrices: incinerator stack gases, sewage sludge, and scrubber water effluents. The paper summarizes the initial development work that was performed in preparation of analytical test protocols that could measure co-planar PCBs in air, water, and sludge matrices. Following the method development, a MACT emissions test program was conducted at a sewage sludge facility in July 1999 and these data are also summarized in the paper. PMID- 11280989 TI - Hazardous waste incineration emissions in perspective. AB - There has been increasing concern over the stack emissions of toxic substances from hazardous waste incinerators, and with improved sampling and analytical technology, measurements are being made at lower and lower levels to support risk assessment and various types of decision-making. However, it is generally difficult to visualize these levels of emissions, which span many orders of magnitude. Data on stack emissions were compiled from various research and compliance testing programs, and representative examples of various types of emissions were plotted on a series of graphs that spans the entire range of concentrations that may be encountered. The result is an illustrative tool for communication as to what emissions from hazardous waste incinerators are actually like. PMID- 11280990 TI - Development and evaluation of a mass spectrometer-based continuous emission monitor for volatile organic compound emissions from combustion devices. AB - A mass spectrometer-based continuous emission monitor (MS-CEM) for organic compound emissions from combustion devices was developed and evaluated at the Louisiana State University (LSU) pilot-scale rotary kiln incinerator (RKI). The MS-CEM consists of a stack probe, heat-traced sampling line, vacuum pump, particulate filter, Nafion@ dryer and mass spectrometer. The mass spectrometer includes a computer that controls and optimizes the operation of the unit. The MS CEM is capable of continuously analyzing up to 40 different volatile organic compounds on a real-time basis. The MS-CEM is capable of analyzing, computing and recording the analytical results for each and up to 40 different organic compounds in less than 0.3 s. Four different volatile organic compounds were mixed together and injected into the baghouse inlet while simultaneously analyzing each organic component exiting the RKI stack gas. The results obtained from MS-CEM were compared with the material balance values. The system response time (including the MS-CEM) varies from 1.1 to 1.5 min. PMID- 11280991 TI - [A new melanoma antigen-encoding gene subfamily in human chromosome X]. AB - A lot of studies have been focused on the tumor-related genes. We have cloned a new melanoma antigen-encoding gene (MAGE) from human fetal liver of third trimester and subsequently found that it represented a new MAGE gene subfamily, named MAGE-D. The MAGE-D subfamily contained three orthologs including human MAGE D1, rat SNERG-1 and mouse DLXIN-1, and two paralogs including human MAGE-D and human KIAA1114. All human members of MAGE-D subfamily are located in human chromosome Xp11.21-p11.23. Moreover, MAGE-D subfamily stands out from other known subfamilies MAGE-A, -B and -C of MAGE family in view of typical features such as exon/intron organization, absence of tumorspecific antigens, evolutionarily divergent in sequences. The identification of MAGE-D subfamily will be helpful in understanding the genesis of tumor. PMID- 11280993 TI - [Actin is located in the nucleus and nuclear matrix of HeLa cells]. AB - HeLa cell nuclei were isolated and the nuclear matrix specimens were prepared. After labelled with an anti-actin antibody and FITC-conjugated secondary antibody, both the nuclei and nuclear matrix specimens were observed to emanate specific yellow-green fluorescence. The fluorescent signals in the nuclear matrix were much stronger than that in the nuclei, and the signals in former were widespread throughout the whole structure while that in the latter were mainly defined to their peripheral regions. A 43 kD band was revealed in nuclei and nuclear matrix specimens by SDS-PAGE, and was then proved to be actin by Western blot, confirming the immuno-fluorescence observations. When stained with TRICT conjugated phalloidin, both the nuclei and nuclear matrix specimens were found to give off specific, red fluorescent signals which represent the location of F actin (filamentous actin), and the pattern of TRITC signals distribution in the nuclei and nuclear matrix showed similarity with that of FITC signals. The existence and significance of actin and F-actin in the nuclei and nuclear matrix were discussed. PMID- 11280992 TI - [Molecular evolution of the Thr-Gly region of the period gene in Drosophila and some dipterans]. AB - In this study, the molecular evolution of the Thr-Gly region of the period gene was characterized, using dipteran groups with close, medium, and long distance phylogenetic relationship. No sexual selection or other positive selection was found to be acting on the Thr-Gly region. The evolutionary rate of the Thr-Gly region in nasuta subgroup was 10.4 x 10(-9) synonymous substitution/site/year. The divergence time of the nasuta subgroup of Drosophila was estimated to be 1-3 mya. A phylogenetic tree of Drosophila genus was reconstructed, which is well supported by evidences from archaebiological and biogeographical studies, The molecular evolutionary pattern of Thr-Gly region was discussed. PMID- 11280994 TI - [Protein databank for several tissues derived from five instar of silkworm]. AB - We attempted to construct protein databank of silkworm (Bombyx mori L.) for clarifying the gene expression and post-translational modification. Proteins from silkworm body wall, fat body and middle intestines were separated by two dimensional electrophoresis. The N-terminal amino acid sequences of 40 proteins and their homology to proteins from other organisms were determined. 58.5% proteins were high homologous to the already reported proteins in D. melanogaster and 36.5% to those in other organisms. Only 5% of the proteins were homologous to the known protein in silkworm. The N-terminal sequences of 27 proteins were first found in silkworm, and all these data were registered in SWISS-PROT through the Internet. The results suggested that the information of gene expression and post translational modification could be obtained by the information of protein databank. PMID- 11280995 TI - [PCR-SSCP and sequencing analysis on 5' terminal region of mtDNA control region in sheep]. AB - PCR-SSCP and sequencing of the left domain in ovine mitochondrial control region revealed that all 202 sheep from Small Tail Han sheep, Wuzhumuqin sheep, Hu Yang, Suffol, Sharolai and hybrids between Dorset (Male) and Small Tail Han sheep (Female) can be classified into two types: common type and mutation type, this result suggested that modern domestic sheep originate from two main ancestors. PMID- 11280996 TI - New rice mutants lacking glutelin alpha-1 subunit. AB - Four rice (Oryza sativa L.) mutant lines lacking glutelin alpha-1 subunit were obtained by screening the progenies of fertilized egg treatment with MNU. SDS PAGE and IEF analyses showed that the mutants without pI6.82 polypeptide in common while forming/increasing other polypeptide indicating that the mutants were controlled by structural genes. IEF analysis also suggested that the polypeptides of pI6.82 and pI8.58 derived from the same glutelin precursor. The mutants are considered to be useful material for glutelin quality improvement and for genetic regulatory mechanism study of glutelin biosynthesis. PMID- 11280997 TI - [Construction of a bacterial artificial chromosome (BAC) contig encompassing the bacterial blight resistance gene Xa4 locus in rice]. AB - The gene Xa4 confers dominantly resistance to rice bacterial blight, which has been finely mapped between RFLP markers G181 and L1044, and co-segregated with the resistance gene homologues sequence marker RS13. The three markers were used to screen a rice Bacterial Artificial Chromosome (BAC) library constructed from IRBB56, a Xa4-harborring indica variety, resulting in the detection of totally 128 positive clones. Of the 18 positive clones picked out by RS13, 4 and 6 clones were simultaneously detected by G181 and L1044, respectively. Based on their HindIII restriction patterns, 12 clones were selected out to construct a contig that spanned about 420 kb covering the Xa4 locus, which is a solid base for the isolation of Xa4 gene. PMID- 11280998 TI - [Analyses of population genetic structure by using dominant markers]. AB - Dominant markers tend to under-estimate the amount of genetic diversity relative to codominant systems when applying in population genetics. In order to compare various existing methods for analysis of genetic structure, RAPD markers were used to detect genetic variability of 5 populations of Oryza granulata from China. The results indicated that both Shannon index of diversity and Nei gene diversity were superior to percentage of polymorphic bands (PPB) because the latter lacked of ability to describe frequency difference of polymorphic bands. Mantel test showed significant relation (r > 0.95, t > t0.01) among matrice of 17 different genetic similarities, which indicated that all of them could be used in analysis of genetic relations of individuals of Oryza granulata. Both AMOVA analysis based on phi st distance and analysis of Nei's distance showed consistent results in defining relationship among the 5 populations, and Lynch Milligan pruning should be used to improve the estimation of population parameters. All of AMOVA, Gst and Shannon diversity analyses obtained similar results with majority of genetic variation occurring between Yunnan and Hainan, and low levels of genetic diversity resided within regions and populations. PMID- 11280999 TI - [Relationship between differential expression patterns of multigene families and heterosis in a wheat diallel crosses]. AB - In order to understand molecular basis of heterosis, the patterns of differential gene expression of multigene families between wheat hybrids and their parents in seedling leaves were analyzed by using mRNA differential display. Relationships between differential gene expression patterns, heterosis and F1 hybrid performance were determined. Four patterns of differential gene expression were observed, which include: (1) bands observed in both parents but not in the F1; (2) bands occurring in only one parent but not in the F1 or the other parent; (3) bands detected in only the F1 but neither of the parents; (4) bands present in one parent and F1 but absent in the other parent. The analysis showed that patterns of differential gene expression were not correlated with the F1 hybrid performance for all the eight agronomic traits. However, differentially expressed fragments that occurred only in the F1 but neither of the parents were found to be positively correlated with heterosis. On the contrary, fragments observed in both parents but not in the F1 were negatively correlated with heterosis. It is concluded that the differential expression of regulatory genes plays an important role in heterosis. PMID- 11281000 TI - [Development and identification of T. aestivum-S. cereale-H. villosa double translocation line 1RS.1BL, 6VS.6AL via chromosome C-banding and dual color FISH]. AB - In order to enhance the disease resistance of wheat-rye 1BL.1RS translocation line and broaden its genetic bases, a 1RS.1BL line was crossed to 6VS.6AL and from its F2 progenies, a double translocation line designated as 1RS.1BL, 6VS.6AL (2n = 42) was identified by (root tip cell) RTC chromosome C-banding. Its mean chromosome configuration at PMC MI was 19.14II + 1.86[symbol: see text] II indicating genetic stability. Rye and H. villosa genomic DNA labelled by biotin- and digoxingenin-11-dUTP respectively were used as probes simultaneously for dual color FISH identification. The results confirmed the C-banding results. Rye and H. villosa chromatin after FISH expressed green or red signals respectively in both RTC and PMC of the double translocation line. This line showed normal fertility, desirable agronomic traits and resistance to powdery mildew, and might be of interest for wheat improvement. PMID- 11281001 TI - [Development of laboratory sequence analysis software based on WWW and UNIX]. AB - Sequence analysis tools based on WWW and UNIX were developed in our laboratory to meet the needs of molecular genetics research in our laboratory. General principles of computer analysis of DNA and protein sequences were also briefly discussed in this paper. PMID- 11281002 TI - [Effects of Pro229-->Ser and Glu243-->Gly on the characters of thermostable catechol 2, 3-dioxygenase]. AB - In order to investigate the effects of amino acid replacement on the characters of thermostable catechol 2, 3-dioxygenase, two mutants (Pro229Ser and Glu243Gly) of this enzyme were obtained by using the method of PCR random mutagenesis. The wild type thermostable catechol 2, 3-dioxygenase and these two mutants (Pro229Ser, Glu243Gly) were over expressed in E. coli TG1 and purified. The enzymatic characters and thermostability of the wild type enzyme and the two mutants (Pro229Ser, Glu243Gly) were analyzed. The results revealed that the optimum enzymatic temperature of the two mutants were the same as that of the wild type enzyme (60 degrees C) and the Kcat/Km value of Pro229Ser and Glu243Gly (4.89 +/- 0.01 x 10(6) mol-1 s-1 and 5.88 +/- 0.01 x 10(6) mol-1 s-1, respectively) were reduced compared with the wild type enzyme (6.97 +/- 0.01 x 10(6) mol-1 s-1). However, the thermostability of Pro229Ser extremely decreased 10.2 degrees C and the thermostability of Glu243Gly slightly increased 1.5 degrees C. It was proposed that Pro229 played an important role on the thermostability of thermostable catechol 2, 3-dioxygenase. PMID- 11281003 TI - [Cloning and expression of nisZ gene in Lactococcus lactis]. AB - The gene encoding the precursor of nisin was amplified by PCR using the lambda HJ 3 DNA as the template, which contained the entire nisin biosynthesis gene cluster from Lactococcus lactis AL2 with high yield of nisin, and was cloned into pMG36e. The recombinant plasmid pHJ201 was introduced into Lactococcus lactis NZ9800 by electroporation. pHJ201 is very stable in L. lactis NZ9800. Antimicrobial activity test and Tricine-SDS-PAGE analysis revealed that L. lactis NZ9800 harbouring pHJ201 restored ability of nisin production, but the production level was markedly lower than L. lactis AL2. The result of DNA sequence analysis indicated that Nisin Z is produced by L. lactis AL2. PMID- 11281004 TI - Belling the cat: clinical investigation in vulnerable populations (a good idea, but who's going to volunteer?). PMID- 11281005 TI - A significant day for women's health and the FDA. PMID- 11281006 TI - Antipsychotic medications and ethnicity. PMID- 11281007 TI - Does exclusion or inclusion better protect women? PMID- 11281008 TI - Estrogen effects on the brain. AB - This article reviews available data on the effects of estrogen on the brain. Such information is derived from basic laboratory studies, inferences from comparisons of gender differences, observational epidemiologic studies, and clinical trials of estrogen in humans. Findings suggest the potentially favorable influences of estrogen on mood and cognition in older women and augment current knowledge about physical risks and benefits of estrogen use. PMID- 11281009 TI - Gender differences in the genetic epidemiology of major depression. AB - This article reviews the current state of knowledge about gender differences in the genetic epidemiology of major depression. In particular, this article seeks to answer two central questions: (1) Is the magnitude of the genetic influences on the liability to major depression similar in men and women? and (2) Are the genetic factors that predispose women to major depression the same as those that predispose men to major depression? PMID- 11281010 TI - Antepartum and postpartum depression. AB - Psychiatric mood disorders can and do occur in pregnant women. Women with antepartum depression have a risk of poor nutrition, substance abuse, and prenatal noncompliance. Careful assessment of risk and benefits to the pregnant woman and to the unborn child must be made before pharmacologic therapy is initiated. The three postpartum mood disorders--postpartum "blues," postpartum depression, and postpartum psychosis--are common, and education is an important instrument in the treatment of these disorders. PMID- 11281011 TI - Medical communication and gender: a summary of research. AB - Communication patterns between physicians and patients during the medical visit reveal behavioral gender differences. Studies suggest varying satisfaction levels depending on physician gender. The communication style of female physicians often includes slightly more focus on the patient's emotional and psychosocial concerns, more positively toned communications, and a more egalitarian style reflected in increased levels of patient participation. However, because patients' satisfaction with female physicians does not typically exceed that for male physicians, other factors may influence satisfaction ratings. PMID- 11281012 TI - Women's health in Israel and the United States--a cross-cultural perspective. PMID- 11281013 TI - Urinary incontinence: a family affair. PMID- 11281014 TI - The evolution of women's health resources. PMID- 11281015 TI - Does the photochemical conversion of colchicine into lumicolchicines involve triplet transients? A solvent dependence study. AB - beta- and gamma-lumicolchicines are photoproducts formed by the cycloisomerization of the tropolone ring of colchicine (COL) alkaloids. The mechanism of the photoconversion, suggested to involve the triplet state, is examined here by studying the effect of the solvent polarity on the lumicolchicine photoisomer ratio. Triplet COL, detected by laser flash photolysis, is quenched by oxygen, but not by transtilbene or 1-methylnaphtalene. Neither the quantum yield of conversion of COL nor the photoproduct ratio was altered by the presence of oxygen. Likewise, energy transfer to COL from triplet acetone produced by either isobutanal/horseradish peroxidase system or tetramethyldioxetane thermolysis failed to provoke photoreaction of COL. Our data argue against the intermediacy of a COL triplet state in the photoisomerization and stress on the role of specific solvent-solute interactions in determining the partitioning of excited singlet state into the beta- and gamma-isomer formation. PMID- 11281016 TI - Transient absorption from the 1Bu+ state of all-trans-beta-carotene newly identified in the near-infrared region. AB - We have attempted subpicosecond time-resolved absorption spectroscopy of all trans-beta-carotene in organic solvents in the 820-1060 nm region and found novel transient absorption features which lived in subpicosecond time scales. A first component that appeared immediately after excitation showed a lifetime of 190 +/- 10 fs in n-hexane in agreement with the 1Bu+ lifetime that had been determined by fluorescence upconversion spectroscopy (195 +/- 10 fs). (Kandori et al. [1994] J. Am. Chem. Soc. 116, 2671-2672.) Therefore, this component is assigned to a transient absorption from the 1Bu+ state. PMID- 11281017 TI - Nucleotide oxidation mediated by naphthalimide excited states with covalently attached viologen cosensitizers. AB - The ground- and excited-state interactions of polymethylene-linked 1,8 naphthalimide-viologen dyads with calf-thymus DNA have been investigated. By virtue of the covalently attached viologen, the compounds represent the first example of linked chromophore/cosensitizer systems in the photooxidation of duplex DNA. The compounds associate strongly with DNA. Analysis of ground-state spectral changes yield binding constants of 0.7-2.5 x 10(6) M-1. Upon 355 nm pulsed irradiation of the compounds in the presence of calf-thymus DNA, reduced viologen is observed within the laser pulse. Photoproducts are not observed on this time scale in the absence of DNA. Since ground-state bleaching of the naphthalimide was not observed, the results suggest that DNA nucleobases are the species being oxidized. The quantum efficiency of radical production increases with the extent of binding to DNA. Under conditions where the compounds are bound predominantly to DNA, the quantum efficiencies were found to range from 0.02 to 0.03. Although small, the values represent a substantial increase in charge separation yield compared to 1,8-naphthalimide compounds that lack the covalently attached viologen. The mechanism of radical production and effect of number of intervening methylenes are discussed. PMID- 11281018 TI - Analysis of fluoroquinolone-mediated photosensitization of 2'-deoxyguanosine, calf thymus and cellular DNA: determination of type-I, type-II and triplet triplet energy transfer mechanism contribution. AB - Fluoroquinolone (FQ) antibacterials are known to exhibit photosensitization properties leading to the formation of oxidative damage to DNA. In addition, photoexcited lomefloxacin (Lome) was recently shown to induce the formation of cyclobutane pyrimidine dimers via triplet-triplet energy transfer. The present study is aimed at gaining further insights into the photosensitization mechanisms of several FQ including enoxacin (Enox), Lome, norfloxacin (Norflo) and ofloxacin (Oflo). This was achieved by monitoring the formation of DNA base degradation products upon UVA-mediated photosensitization of 2'-deoxyguanosine, isolated and cellular DNA. Oflo and Norflo act mainly via a Type-II mechanism whereas Lome and, to a lesser extent, Enox behave more like Type-I photosensitizers. However, the extent of oxidative damage was found to be relatively low. In contrast, it was found that cyclobutane thymine dimers represent the major class of damage induced by Enox, Lome and Norflo within isolated and cellular DNA upon UVA irradiation. This striking observation confirms that FQ are able to promote efficient triplet energy transfer to DNA. The levels of photosensitized formation of strand breaks, alkali-labile sites and oxidative damage to cellular DNA, as measured by the comet assay, were confirmed to be rather low. Therefore, we propose that the phototoxic effects of FQ are mostly accounted for energy transfer mechanism rather than by Type-I or -II photosensitization processes. PMID- 11281019 TI - cis-Urocanic acid does not induce the expression of immunosuppressive cytokines in murine keratinocytes. AB - trans-Urocanic acid (UCA) acts as a chromophore for UV radiation in the epidermis and isomerizes to cis-UCA which then initiates some of the changes leading to UV induced immunosuppression. The mechanism of the immunomodulation by cis-UCA is unknown at present, but one possibility is that the interaction between cis-UCA and keratinocytes causes the release of immunosuppressive cytokines locally. To test this hypothesis, PAM-212 cells, a murine keratinocyte cell line, were incubated with 0.10-100 micrograms/mL trans- and cis-UCA for 6 or 24 h, respectively. The expression of interleukin (IL)-10, transforming growth factor (TGF)-beta and tumor necrosis factor (TNF)-alpha messenger RNA (mRNA) was then measured by reverse transcription-polymerase chain reaction in comparison with the mRNA for the house-keeping gene, beta-actin. No change or significant induction of any of the cytokine messages occurred. However, the expression of IL 10 messenger RNA (mRNA) was induced 24 h after UVB irradiation (300 J/m2) and that of TNF-alpha mRNA occurred 6 h after treatment with phorbol myristate acetate. The expression of IL-10 protein was also examined by immunostaining in both PAM-212 cells and B16-F10 murine melanoma cells between 3 and 48 h after incubation with 10 and 100 micrograms/mL cis- and trans-UCA. No alteration was seen with either isomer at either concentration. In contrast, UVB irradiation of both cell lines resulted in a marked increase in intracellular IL-10 protein at 24 and 48 h. Therefore the upregulation of the immunosuppressive cytokines, IL 10, TNF-alpha and TGF-beta, in keratinocytes is unlikely to be the mechanism by which cis-UCA induces immunosuppression in mice. PMID- 11281020 TI - Application of molecular orbital calculations to interpret the chlorophyll spectral forms in pea photosystem II. AB - The energy and oscillator strength of electronic transitions of chlorophyll (Chl) amino acid complexes were calculated by using molecular orbital methods. The energies varied widely with coordinated amino acids and the difference between the maximum and minimum energy was about 830 cm-1. This energy difference was comparable with the spreading of absorption bands for light-harvesting Chl protein complexes of photosystem II (LHC II) of green plants. The feature of the Qy band for pea LHC II was interpreted with the aid of the calculated energies and oscillator strengths. Four spectral components of the band were assigned to individual Chl-amino acid complexes. PMID- 11281021 TI - Evidence from action and fluorescence spectra that UV-induced violet-blue-green fluorescence enhances leaf photosynthesis. AB - We assessed the contribution of UV-induced violet-blue-green leaf fluorescence to photosynthesis in Poa annua, Sorghum halepense and Nerium oleander by measuring UV-induced fluorescence spectra (280-380 nm excitation, 400-550 nm emission) from leaf surfaces and determining the monochromatic UV action spectra for leaf photosynthetic O2-evolution. Peak fluorescence emission wavelengths from leaf surfaces ranged from violet (408 nm) to blue (448 nm), while excitation peaks for these maxima ranged from 333 to 344 nm. Action spectra were developed by supplementing monochromatic radiation from 280 to 440 nm, in 20 nm increments, to a visible nonsaturating background of 500 mumol m-2 s-1 photosynthetically active radiation and measuring photosynthetic O2-evolution rates. Photosynthetic rates tended to be higher with the 340 nm supplement than with higher or lower wavelength UV supplements. Comparing photosynthetic rates with the 340 nm supplement to those with the 400 nm supplement, the percentage enhancement in photosynthetic rates at 340 nm ranged from 7.8 to 9.8%. We suspect that 340 nm UV improves photosynthetic rates via fluorescence that provides violet-blue-green photons for photosynthetic energy conversion because (1) the peak excitation wavelength (340 nm) for violet-blue-green fluorescence from leaves was also the most effective UV wavelength at enhancing photosynthetic rates, and (2) the magnitude of photosynthetic enhancements attributable to supplemental 340 nm UV was well correlated (R2 = 0.90) with the apparent intensity of 340 nm UV-induced violet-blue-green fluorescence emission from leaves. PMID- 11281022 TI - Antivascular treatment of solid melanoma tumors with bacteriochlorophyll-serine based photodynamic therapy. AB - We describe here a strategy for photodynamic eradication of solid melanoma tumors that is based on photo-induced vascular destruction. The suggested protocol relies on synchronizing illumination with maximal circulating drug concentration in the tumor vasculature attained within the first minute after administrating the sensitizer. This differs from conventional photodynamic therapy (PDT) of tumors where illumination coincides with a maximal concentration differential of sensitizer in favor of the tumor, relative to the normal surrounding tissue. This time window is often achieved after a delay (3-48 h) following sensitizer administration. We used a novel photosensitizer, bacteriochlorophyll-serine (Bchl Ser), which is water soluble, highly toxic upon illumination in the near-infrared (lambda max 765-780 nm) and clears from the circulation in less than 24 h. Nude CD1 mice bearing malignant M2R melanotic melanoma xenografts (76-212 mm3) received a single complete treatment session. Massive vascular damage was already apparent 1 h after treatment. Changes in vascular permeability were observed in vivo using contrast-enhanced magnetic resonance imaging (MRI), with the contrast reagent Gd-DTPA, by shortening spin-spin relaxation time because of hemorrhage formation and by determination of vascular macromolecular leakage. Twenty-four hours after treatment a complete arrest of vascular perfusion was observed by Gd DTPA-enhanced MRI. Histopathology performed at the same time confirmed primary vascular damage with occlusive thrombi, hemorrhage and tumor necrosis. The success rate of cure of over 80% with Bchl-Ser indicates the benefits of the short and effective treatment protocol. Combining the sensitizer administration and illumination steps into one treatment session (30 min) suggests a clear advantage for future PDT of solid tumors. PMID- 11281024 TI - The application of a compact multispectral imaging system with integrated excitation source to in vivo monitoring of fluorescence during topical photodynamic therapy of superficial skin cancers. AB - A novel, compact and low-cost multispectral fluorescence imaging system with an integrated excitation light source is described. Data are presented demonstrating the application of this method to in vivo monitoring of fluorescence before, during and after topical 5-aminolevulinic acid photodynamic therapy of superficial skin cancers. The excitation source comprised a fluorescent tube with the phosphor selected to emit broadband violet light centered at 394 nm. The camera system simultaneously captured spectrally specific images of the fluorescence of the photosensitizer, protoporphyrin IX, the illumination profile and the skin autofluorescence. Real-time processing enabled images to be manipulated to create a composite image of high contrast. The application and validation of this method will allow further detailed studies of the characteristics and time-course of protoporphyrin IX fluorescence, during topical photodynamic therapy in human skin in vivo. PMID- 11281023 TI - Influence of the substitution of 3-vinyl by 3-formyl group on the photodynamic properties of chlorin P6: molecular, cellular and in vivo studies. AB - Molecular in vitro and in vivo properties of 3-devinyl-3-formylchlorin p6 (FCp6) were examined in order to characterize this derivative as a new prospective photosensitizer. The long-wavelength absorption maximum of FCp6 was 690-696 nm (depending on environment). FCp6 was found to bind readily to membranous structures and form complexes with some proteins. The dye was associated with the plasmalemma and distributed rather diffusely along the cytoplasm with ca a three fold higher accumulation within mitochondria in A549 human adenocarcinoma cells. The spectral analysis revealed that the major part of FCp6 was bound to membranes within cells. The membrane-bound FCp6 was shown to generate singlet oxygen efficiently. The average cytoplasmic concentration of FCp6 in A549 cells achieved ca 80% of its extracellular concentration in complete medium. The dye was characterized by a very fast efflux (16-fold decrease in 2 h). The ex vivo analysis of FCp6 fluorescence in mice revealed that the maximal dye content in blood, tissues, organs and tumor was achieved in less than 1 h after injection, followed by a considerable (ca six-fold) decrease during the next 23 h and a long term persistence at low level. A preferential accumulation of FCp6 in subcutaneously implanted Ehrlich carcinoma along with its higher retention level comparing to the surrounding skin and muscles were observed in mice treated with different dye doses. In vitro cytotoxic assays with A549 and Raji B-cell lymphoma cells as well as in vivo analyses using Ehrlich carcinoma in mice revealed the very low toxicity of FCp6 without light irradiation and the significant photodynamic activity of this compound. PMID- 11281025 TI - Photodamage induced by Zinc(II)-phthalocyanine to microtubules, actin, alpha actinin and keratin of HeLa cells. AB - We have studied the photosensitizing effects of zinc(II)-phthalocyanine (ZnPc) on the cytoskeleton of HeLa cells using sublethal (10(-7) M, followed by 1 or 3 min of red light to induce 20%, LD20, or 60%, LD60, cell death, respectively) or lethal (5 x 10(-6) M and 15 min of irradiation, LD100) experimental conditions. The immunofluorescent analysis of the cytoskeleton showed a variable photodamage to microtubules (MT), actin microfilaments (AF) and intermediate filaments of keratin (KF), as well as on alpha-actinin, which was dependent on treatment conditions. Both sublethal treatments induced deep alterations on interphase and mitotic MT. The mitotic index increased with time with the maximum at 18 h (12%) or 24 h (14%) after LD20 or LD60, respectively. The alterations on AF and alpha actinin were much more severe than those observed on KF at any evaluated time. With the exception of the KF, which remained partially organized, the MT and AF network was severely damaged by the lethal treatment. Western blot analysis for alpha-tubulin, G-actin and alpha-actinin from soluble and insoluble fractions confirmed the results observed by immunofluorescence, thus indicating that these cytoskeletal components are involved in cell damage and death by ZnPc photosensitization. PMID- 11281026 TI - Purpurin-18 in combination with light leads to apoptosis or necrosis in HL60 leukemia cells. AB - Photodynamic therapy (PDT), a cancer treatment using a photosensitizer and visible light, has been shown to induce apoptosis or necrosis. We report here that Purpurin-18 (Pu18) in combination with light induces rapid apoptotic cell death in the human leukemia cell line (HL60) at low doses and necrosis at higher concentrations. Cells treated with Pu18 and light under apoptotic conditions exhibited DNA laddering and an increase in both cellular content of subdiploid DNA and externalization of phosphatidylserine (PS), indicating DNA fragmentation and loss of membrane phospholipid asymmetry. In the absence of light activation, Pu18 at nanomolar concentrations had no detectable cytotoxic effect. Caspase-3 activity was increased even after 1 h from treatment with low doses of Pu18 and light. The PS exposure and nuclear features of apoptosis were prevented by treatment of cells before illumination with caspase inhibitors benzyloxycarbonyl Val-Ala-Asp-fluoromethylketone (Z-VAD-FMK) and benzyloxycarbonyl-Asp-Glu-Val-Asp fluoromethylketone (Z-DEVD-FMK). Conversely, the caspase-1 inhibitor, acetyl-Tyr Val-Ala-Asp-aldehyde (Ac-YVAD-CHO) failed to suppress the apoptosis. No protective effect of the three caspase inhibitors was observed when the cells were exposed to necrotic concentrations of Pu18 and light. Our results show that caspase-3, but not caspase-1, is involved in the signaling of apoptotic events in PDT with Pu18-induced apoptosis of HL60 cells. Moreover, both the time course of PS exposure and the effect of caspase inhibitors on it indicate that it is regulated in the same manner as DNA fragmentation. PMID- 11281028 TI - Photoinduced inactivation of T7 phage sensitized by symmetrically and asymmetrically substituted tetraphenyl porphyrin: comparison of efficiency and mechanism of action. AB - We investigated the efficiency and the mechanism of action of two--one symmetrically and one asymmetrically substituted--glycoconjugated tetraphenyl porphyrins in their photoreaction with T7 phage as a model of nucleoprotein (NP) complexes. A correlation was found between the dark inactivation of T7 and the binding of porphyrins determined by fluorescence spectroscopy. Both types of porphyrin sensitized the photoinactivation of T7, but the slopes of inactivation kinetics were markedly different. There was no correlation between the dark binding and the photosensitizing efficacy of the two derivatives. Inactivation was moderated by 1,3-diphenylisobenzofuran and 1,3-dimethyl-2-thiourea; however, neither of them inhibited T7 inactivation completely. This result suggests that both Type-I and Type-II reactions play a role in the virus inactivation. Optical melting studies revealed structural changes in the protein part but not in the DNA of the photochemically treated NP complex. Polymerase chain reaction analysis of a 555 bp segment of gene 1 and a 3826 bp segment of genes 3 and 4 failed to demonstrate any DNA damage. PMID- 11281027 TI - Effects of fluence rate on cell survival and photobleaching in meta-tetra (hydroxyphenyl)chlorin-photosensitized Colo 26 multicell tumor spheroids. AB - We report the influence of fluence rate on the photobleaching and cell survival in Colo 26 multicell spheroids photosensitized by meta-tetra (hydroxyphenyl)chlorin (mTHPC). Photosensitizer degradation and therapeutic efficacy increased dramatically and progressively when the fluence rate was reduced over the range from 90 to 5 mW cm-2. These experimental results were compared to a mathematical model of photobleaching based on self-sensitized singlet oxygen reactions with the photosensitizer ground state. This model incorporates photophysical parameters obtained from microelectrode measurements of oxygen depletion at the surface of mTHPC-sensitized spheroids and was refined by including the inhomogeneous distribution of mTHPC in spheroids and oxygen depletion in the bulk medium. Since the model is consistent with the experimental data we conclude that the fluence rate dependence of the cell survival and of mTHPC photobleaching is due to photochemical oxygen consumption and a predominantly singlet oxygen-mediated mechanism of mTHPC photobleaching. The threshold dose of reacting singlet oxygen was calculated to be 7.9 +/- 2.2 mM in this system. PMID- 11281029 TI - Is delta-aminolevulinic acid dehydratase rate limiting in heme biosynthesis following exposure of cells to delta-aminolevulinic acid? AB - Understanding the regulation and control of heme/porphyrin biosynthesis is critical for the optimization of the delta-aminolevulinic-acid (ALA)-mediated photodynamic therapy of cancer, in which endogenously produced protoporphyrin IX (PPIX) is the photosensitizer. The human breast cancer cell line MCF-7, the rat mammary adenocarcinoma cell line R3230AC, the mouse mammary tumor cell line EMT-6 and the human mesothelioma cell line H-MESO-1 were used to study ALA-induced PPIX levels and their relationship to delta-aminolevulinic acid dehydratase (ALA-D) activity in vitro. Incubation of these cell lines with 0.5 mM ALA for 3 h resulted in a significant increase in PPIX accumulation, compared with control cells, but there was no significant change in ALA-D activity. Exposure of cells incubated with ALA to 30 mJ/cm2 of fluorescent light, a dose that would cause a 50% reduction in cell proliferation, did not significantly alter the activity of ALA-D. Increasing the activity of porphobilinogen deaminase (PBGD), the enzyme immediately subsequent to ALA-D, by four- to seven-fold via transfection of cells with PBGD complementary DNA did not alter the activity of ALA-D. However, incubation of cells with various concentrations of succinyl acetone, a potent inhibitor of ALA-D, caused a concomitant decline in both PPIX accumulation and ALA-D activity. These data imply that when cells are exposed to exogenous ALA, ALA-D is an important early-control step in heme/porphyrin biosynthesis and that regulation of PPIX synthesis by this dehydratase may impact the effectiveness of ALA-mediated photosensitization. PMID- 11281030 TI - Urokinase activity in corneal fibroblasts may be modulated by DNA damage and secreted proteins. AB - Proteases like urokinase-type plasminogen activator (uPA) play an important role in tumor invasion. Cells derived from ultraviolet radiation (UVR)-induced corneal sarcomas of Monodelphis domestica produce relatively high levels of uPA compared to the untransformed keratocytes suggesting a mechanism for their invasiveness. Because UVR is known to stimulate uPA production in many cell types, UVR exposure may further increase uPA expression in corneal tumor cells, thus enhancing their ability to infiltrate. We investigated control of basal uPA levels and the induction of uPA by UVR in transformed and untransformed corneal keratocytes from Monodelphis. These studies took advantage of the fact that Monodelphis possesses an active photolyase that can be stimulated to remove UVR-induced pyrimidine dimers by exposure to long-wavelength visible photoreactivating light (PRL). Our studies showed that significant induction of uPA occurred in response to 200 J/m2 UVR. This induction was partially blocked by treatment with PRL, indicating that DNA damage, the pyrimidine dimer in particular, played a role in uPA induction. In untransformed cultured corneal fibroblasts, the heparin-binding protein inhibitor, suramin, reduced basal uPA levels, UVR-induced uPA production and cell proliferation. Basic fibroblast growth factor, a heparin-binding growth factor known to be UVR-inducible in mesenchymal cells, stimulated uPA production and cell proliferation; however, anti-bFGF antibodies did not significantly decrease proliferation or basal uPA production. These findings suggested that basal levels of uPA secretion were modulated in response to heparin-binding growth factors and that these growth factors may also have mediated the effect of UVR on uPA levels. PMID- 11281031 TI - Light irradiation induces fragmentation of the plasmodium, a novel photomorphogenesis in the true slime mold Physarum polycephalum: action spectra and evidence for involvement of the phytochrome. AB - A new photomorphogenesis was found in the plasmodium of the true slime mold Physarum polycephalum: the plasmodium broke temporarily into equal-sized spherical pieces, each containing about eight nuclei, about 5 h after irradiation with light. Action spectroscopic study showed that UVA, blue and far-red lights were effective, while red light inhibited the far-red-induced fragmentation. Difference absorption spectra of both the living plasmodium and the plasmodial homogenate after alternate irradiation with far-red and red light gave two extremes at 750 and 680 nm, which agreed with those for the induction and inhibition of the fragmentation, respectively. A kinetic model similar to that of phytochrome action explained quantitatively the fluence rate-response curves of the fragmentation. Our results indicate that one of the photoreceptors for the plasmodial fragmentation is a phytochrome. PMID- 11281032 TI - Psychometric evaluation of trauma and posttraumatic stress disorder assessments in persons with severe mental illness. AB - Interrater reliability, internal consistency, test-retest reliability, and convergent validity were examined for the Trauma History Questionnaire (THQ), the Clinician-Administered Posttraumatic Stress Disorder (PTSD) Scale (CAPS), and the PTSD Checklist (PCL) in 30 clients with severe mental illnesses. Interrater reliability for the THQ and CAPS was high, as was internal consistency of CAPS and PCL subscales. The test-retest reliability of the THQ was moderate to high for different traumas. PTSD diagnoses on the CAPS and PCL showed moderate test retest reliability. Lower levels of test-retest reliability for PTSD diagnoses were related to psychosis diagnoses and symptoms. However, when more stringent criteria for PTSD were used on the CAPS, it had excellent test-retest reliability across all clients. CAPS and PCL diagnoses of PTSD showed moderate convergent validity. The results support the reliability of trauma and PTSD assessments in clients with severe mental illness. PMID- 11281033 TI - Sensitivity and specificity of the phallometric test for pedophilia in nonadmitting sex offenders. AB - The specificity of phallometric testing for pedophilia has been calculated using sex offenders against adult women. Does the offender's actual number of prior sexual contacts with women affect such estimates? To answer this, the authors' studied 82 male sex offenders against adult women, 172 offenders against unrelated children, and 70 offenders against their own biological children or stepchildren. Phallometric testing included visual and auditory depictions of prepubescent, pubescent, and adult males and females. The results for offenders against women showed that those who had had sexual contact with the greatest number of women (consenting or nonconsenting) had the lowest probability of being diagnosed as pedophilic. Specificity, calculated for those who had sexual contact with the most women and thus the most evidence of attraction to them, was 96%. Sensitivity, calculated analogously for men with the most offenses against children, was 61%. PMID- 11281034 TI - Examining symptom expression as a function of symptom severity: item performance on the Hamilton Rating Scale for Depression. AB - Despite widespread use of the Hamilton Rating Scale for Depression (HRSD; M. Hamilton, 1960), questions have been raised concerning its psychometric properties. A nonparametric item response model was used to examine how the probability of observing a specific symptom of depression changes with increases in depressive severity in a sample of depressed adults. Results showed that options from a number of items on the HRSD did not vary as a function of severity and therefore should not be viewed as indicators of depressive severity. The extent to which symptoms are expressed as a function of depressive severity carries important implications for the use of the HRSD as a measure of severity and for the debate concerning construction of depression as a continuum. Results argue against viewing depression as a simple continuum. PMID- 11281035 TI - Using item response theory to understand comorbidity among anxiety and unipolar mood disorders. AB - The authors hypothesized that anxiety and unipolar mood disorders are often comorbid because each disorder indicates a broad, higher order factor. In a clinical subsample of the nationally representative National Comorbidity Survey participants (N = 251), a one-factor model fit the correlations among 7 dichotomous anxiety and unipolar mood diagnoses. Following the lead provided by literature on the structure of emotional and behavioral problems in children, we labeled this factor internalizing. Item response theory was used to explore how each diagnosis mapped onto the internalizing factor. The test information function derived from the 7 diagnoses suggested that they measure primarily the higher end of the factor. In addition, very high scores on internalizing (meeting criteria for 6-7 disorders) were associated with increased social costs, a phenomenon not well captured by the "comorbidity" concept. The results underscore the need to develop clinical assessment instruments that span the full range of the internalizing factor and measure both the shared and distinctive features of anxiety and unipolar mood disorders in a graded, continuous fashion. PMID- 11281036 TI - Introduction to the special section on clinical applications of analogue behavioral observation. AB - This Special Section addresses methods, validity, and utility of analogue behavioral observation. Separate contributions to the Special Section cover analogue behavioral observation of marital interaction, child behavior problems, parent-child interaction, and adult social functioning. Additional articles address psychometric foundations of analogue behavioral observations, general issues, and future directions in the development and evaluation of this assessment method. Many published studies were reviewed in detail and issues of validity, clinical assessment utility, and sources of variance in obtained measures are addressed. PMID- 11281037 TI - Analogue assessment of child behavior problems. AB - The psychometric properties of analogue assessment measures of child behavior problems are reviewed. Analogue assessment refers to an observational measure of targeted behaviors that are elicited by simulated experimental conditions, which, in turn, are devised to approximate natural circumstances. For the most part, this assessment approach has been used sporadically in the clinical setting with children who are behaviorally disturbed. Lack of standardization of measures and inconsistent findings of ecological validity are among several concerns noted. The paucity of available data limits conclusions that can be drawn at this time about the role of analogue assessment in the evaluation and treatment of child behavior problems. PMID- 11281038 TI - Clinic observations of structured parent-child interaction designed to evaluate externalizing disorders. AB - Observing structured parent-child interaction in clinic analogs has been a tradition in child clinical assessment since the 1960s. The clinic analog is designed to re-create important conditions from natural contexts such that dysfunctional parent-child patterns can be observed and modified. To those basic goals, the modern clinician would add diagnosis and treatment evaluation. Three classes of parent-child clinic analogs designed to evaluate preadolescent children with externalizing disorders were reviewed: free play, parent-directed play, and parent-directed chores. Free-play analogs and parent-directed chore analogs were found to have merit, but remain psychometrically underdeveloped. Practitioner reliance on questionnaires and interviews appears likely to continue until observational analogs have attained sufficient psychometric qualities to facilitate routine clinical decisions. PMID- 11281041 TI - Clinical applications of analogue behavioral observation: dimensions of psychometric evaluation. AB - Clinical assessment applications of analogue behavioral observation are discussed in the context of psychometric principles. Analogue behavioral observation involves the measurement of a client's overt behavior in a contrived situation that is analogous to situations that the client is likely to encounter in his or her natural environment. The goal of analogue behavioral observation is to derive valid estimates of the client's behavior in a current or future natural environment. Analogue behavioral observation instruments are often developed with insufficient attention to their psychometric properties, particularly content validity. Psychometric evaluative dimensions vary in their importance, as a function of the goals of the assessment. Although analogue behavioral observation instruments can be sensitive to change, their validity can erode over time and is affected by numerous sources of variance. Analogue behavioral observation assessment may be especially useful in detecting important functional relations in clinical assessment. PMID- 11281040 TI - Analogue observational methods in the assessment of social functioning in adults. AB - This article provides a clinically oriented overview of analogue observational methods used in the assessment of problematic social functioning, specifically skill deficits and social anxiety. This article emphasizes role-play assessment methods, the predominant method used in clinical settings. An examination of the psychometric characteristics of analogue assessment methods is presented, followed by a review of procedural and structural considerations that may impact the quality of assessment data. Of special concern are the potential impacts of instructional variables, structured versus ideographic role-played situations, confederate characteristics and behavior, molar and molecular levels of assessment, self-ratings versus clinician ratings of functioning, and physical attractiveness. Finally, published and empirically evaluated analogue observation tests are critically reviewed with an emphasis on features that may impact their utility in clinical practice. PMID- 11281039 TI - Observation of couple conflicts: clinical assessment applications, stubborn truths, and shaky foundations. AB - The purpose of this review is to provide a balanced examination of the published research involving the observation of couples, with special attention toward the use of observation for clinical assessment. All published articles that (a) used an observational coding system and (b) relate to the validity of the coding system are summarized in a table. The psychometric properties of observational systems and the use of observation in clinical practice are discussed. Although advances have been made in understanding couple conflict through the use of observation, the review concludes with an appeal to the field to develop constructs in a psychometrically and theoretically sound manner. PMID- 11281042 TI - Exporting analogue behavioral observation from research to clinical practice: useful or cost-defective? AB - The special section on analogue behavioral observation (ABO) provided an in-depth review of various ABO assessment procedures. Despite their availability, however, these procedures are rarely used in clinical practice. This may result in part from the traditions on which most ABO assessments are based, from distinctions between clinical and research assessment environments, and from the need for more information about the cost-effectiveness of ABO strategies for meeting specific needs of clinicians in applied settings. Suggestions for bridging the research clinical gap involve investigating more thoroughly when ABO does and does not provide useful information for various purposes in applied settings and increasing accessibility and cost-effectiveness of ABO procedures for practitioners. PMID- 11281043 TI - Measuring cancer patients' psychological distress and well-being: a factor analytic assessment of the Mental Health Inventory. AB - This study examined the psychometric structure of the Mental Health Inventory (MHI) in 433 cancer patients. Using structural equation modeling, confirmatory factor analyses (CFAs) were conducted. Next, exploratory factor analysis (EFA) was used to explore an alternative MHI factor structure with a randomly chosen subsample. Finally, CFAs were conducted on 6 MHI models with the second subsample. Convergent validity was examined by administering the Positive and Negative Affect Schedule (PANAS) and the Dyadic Adjustment Scale (DAS). The CFAs with the original MHI factor structure indicated inadequate fit, supporting the need to conduct an EFA. Results of the EFA indicated support for a 5-factor solution but numerous differences in item factor loadings. The CFA indicated that the 5-factor correlated model was the best fitting model. Correlations between the PANAS and the DAS with the MHI provided preliminary support for the convergent validity of the MHI. Together, these results indicate that the original MHI factor structure may require modification for use in patients with cancer. PMID- 11281044 TI - Current status of inhaled nitric oxide therapy in the intensive care units. PMID- 11281045 TI - Sore throat following tracheal intubation. AB - Post-operative sore throat is a common minor complication after anesthesia. This paper reviews the factors which influence post-operative sore throat in intubated patients. Two hundred and sixty six intubated patients were investigated to find the incidence of sore throat after elective anesthesia in a middle eastern population. The overall incidence of sore throat was 63.9%. There was no significant difference in the incidence of sore throat between males and females, and in the age groups studied. Anesthetic factors including the use of relaxants, the experience of the anesthesiologist, the number of intubation attempts and lubrication of the tracheal tube did not significantly alter the incidence of sore throat. Duration of anesthesia of greater than 90 minutes was associated with significant increase in sore throat (p < 0.001). Surgical factors including type of surgery, the use of throat packs and early oral intake did not alter the incidence of sore throat. Nasogastric tube insertion was associated with a significantly increased incidence of sore throat (p < 0.01). PMID- 11281046 TI - Reversal of mivacurium-induced neuromuscular block by neostigmine. PMID- 11281047 TI - The use of low-dose rocuronium to facilitate laryngeal mask airway insertion. AB - In this two-phase study, the efficacy of low-dose rocuronium to facilitate laryngeal mask airway (LMA) insertion was evaluated. First, the onset time of 100, 150 and 300 micrograms.kg-1 rocuronium was determined using mechanomyography in three groups of patients (n = 10 in each) anaesthetized with propofol-fentanyl nitrous oxide. In the second part, 100, 150 or 300 micrograms.kg-1 rocuronium or placebo was administered randomly to four groups of patients (n = 50 in each) in a double-blind manner. Following this, anaesthesia was induced with propofol 2.5 mg.kg-1. Patients in the group of 300 micrograms.kg-1 rocuronium or placebo received propofol after 1.5 min. Patients in the group of 100 or 150 micrograms.kg-1 rocuronium received propofol after 3 min. The LMA was inserted 90 sec later. Immediately before the induction of anaesthesia, patients were questioned about the symptoms of neuromuscular block. In phase 1, onset times were 180 sec (SD 41), 191 sec (59) and 89 sec (34), respectively. In phase 2, insertion of the LMA was graded as easy in 90.6% of patients receiving rocuronium, compared with 42% of patients who had only propofol (P < 0.05). Rocuronium improved the overall ease of LMA insertion. LMA insertion was graded easy in 80% of patients who received 100 micrograms.kg-1 rocuronium. The incidence of unpleasant effects was greatest with 300 micrograms.kg-1 rocuronium. The optimal dose needed to facilitate LMA insertion with minimal unpleasant effects appeared to be 100 micrograms.kg-1 rocuronium. PMID- 11281048 TI - An experimental study on the relationship of intra-abdominal pressure and renal ischemia. AB - This study was undertaken in order to determine whether or not the increased intra-abdominal pressure during laparoscopic procedures causes renal ischaemia and parenchymal pathology. Fifteen adult New Zealand rabbits were used in the study. Anaesthesia was maintained by 2% isoflurane, 50% O2 in air. Heart rate (HR), mean arterial pressure (MAP), central venous pressure (CVP), end-tidal carbon dioxide (PETCO2), airway pressure (Paw) and blood gases were monitored. Rabbits in control group (group C, n = 7) and study group (group S, n = 8) had a Veress needle placed supraumbilically. Group S was insufflated with CO2 sequentially at 5, 10 and 15 mmHg of intra-abdominal pressures (IAP); each pressure level was maintained for 20 minutes. At the end of the study, laparotomy was performed and blood was withdrawn from renal vein for measurements of renin and angiotensin I levels, and the other kidney was removed simultaneously for pathological evaluation. Haemodynamic and respiratory measurements were stable in group C and were variable in group S. The renin level was 7.27 +/- 0.34 ng.mL-1 and angiotensin I was 5.01 +/- 0.32 ng.mL-1 in group C. In group S, levels of renin and angiotensin I were 26.2 +/- 5.9 ng.mL-1 and 39.4 +/- 12.1 ng.mL-1 respectively, being significantly higher than group C (p < 0.05). Pathological scores were 0.02 +/- 0.008 in group C and 0.82 +/- 0.124 in group S (p < 0.05). There were significant histological changes in group S compared with group C. During prolonged laparoscopic operations high intra-abdominal pressures may result in intra-abdominal organ ischaemia. PMID- 11281050 TI - Apoptosis: regulating death for a better life. PMID- 11281049 TI - The protective effects of high dose ascorbic acid and diltiazem on myocardial ischaemia-reperfusion injury. AB - In this study, we aimed to compare the myocardial protective effects of high dose ascorbic acid with the effects obtained by adding diltiazem to high dose ascorbic acid. We studied 30 elective cardiac surgery patients prospectively. In ascorbic acid group (group AA), ascorbic acid was given after induction and just before aortic declamping, 50 mg.kg-1 each time. In ascorbic acid + diltiazem group (group AA + D), diltiazem was added to ascorbic acid (0.3 mg.kg-1, i.v. after induction and then 2 micrograms.kg-1 min-1 i.v. infusion until declamping). Group C was the control group. There was no significant difference between groups in terms of cardiac enzyme levels. After declamping, the arterial and coronary sinus malondialdehyde levels, measured as a marker of lipid peroxidation, were increased significantly in the group C while remained stable in the other two groups. Ventricular fibrillation (VF) after declamping was positive in 3, 1 and 6 patients in the groups AA, AA + D and C respectively. In this study, we observed the prevention of lipid peroxidation in the group AA and group AA + D. The only positive result obtained by addition of diltiazem to high dose ascorbic acid was the decrease in the frequency of VF after declamping. We concluded that the prevention of lipid peroxidation in the groups AA and AA + D provided no measurable protection over myocardial ischaemia-reperfusion injury. PMID- 11281051 TI - Second dose thiopentone attenuates the haemodynamic response to laryngoscopy and intubation. AB - The study was undertaken to evaluate the effectiveness of large (divided) thiopentone dosage on the peripheral haemodynamic response to laryngoscopy and intubation. Seventy-six (76) patient aged 18 to 67 years were sequentially assigned to either the second dose thiopentone group (n = 36) or the control group (n = 40). The first group had 4 mg.kg-1 thiopentone for induction of anaesthesia, then 1.5 mg.kg-1 suxamethonium chloride for muscle relaxation and a second dose of thiropentone (4 mg.kg-1) just before laryngoscopy and intubation. The control group had thiopentone 4 mg.kg-1 for induction of anaesthesia, suxamethonium 1.5 mg.kg-1 for muscle relaxation and then laryngoscopy and intubation. The heart rate (HR), systolic arterial pressure (SAP), diastolic arterial pressure (DAP), mean arterial pressure (MAP), and rate pressure product (RPP) were measured before induction of anaesthesia and after laryngoscopy and intubation. The difference in the values represented the haemodynamic response to laryngoscopy and intubation. The second dose thiopentone technique compared with the control group, significantly attenuated the post-intubation rise in HR (19.7 vs. 30.9), SAP (18.0 vs. 37.5) and RPP (4795.4 vs. 8440.0). The post intubation rise in DAP (33.9 vs. 42.5) and MAP (31.9 vs. 42.0) didn't show significant difference between the two groups. PMID- 11281052 TI - Combination of granulocyte colony-stimulating factor and antibacterial drugs for the treatment of ventilatory associated nosocomial pneumonia. AB - In this prospective study, we aimed to investigate the role of Granulocyte Colony Stimulating Factor (rhG-CSF) supplement to antibiotherapy, for the treatment of ventilator-associated nosocomial pneumonia (VAP) in patients intubated due to acute respiratory failure. In Emergency Intensive Care Unit (EICU), 28 patients on mechanical ventilation are randomised into two groups as rhG-CSF and control, after they are diagnosed to have VAP. The first group received 5 micrograms/kg/day subcutaneous rhG-CSF as a supplement to antibiotherapy while in the second group the sole treatment was antibiotherapy. For each patient studied, the chart is reviewed at the first day of mechanical ventilation and for 8 days after VAP for the following parameters: erythrocyte, leucocyte, granulocyte and platelet counts; SGOT, SGPT, blood urea, creatinine; microbiological analyses of transtracheal aspirate, hemocultures and infiltrations shown on chest x-ray. APACHE II scores of patients are also recorded. Statistical comparisons among groups are performed with Mann-Whitney U test. The groups didn't differ significantly in erythrocyte, platelet counts and blood urea, creatinine, SGOT, SGPT (p > 0.05). The difference is found to be much more significant according to leucocyte and granulocyte counts in rhG-CSF group, when compared to control group (p < 0.001). We conclude, that combination of antibacterial agents and rhG-CSF may be beneficial for the treatment of VAP. PMID- 11281053 TI - [Contrast agent improves diagnostic value of dobutamine stress echocardiography]. AB - OBJECTIVES: Suboptimal endocardial definition reduces the diagnostic value of stress echocardiography for coronary artery disease, but intravenous infusion of a left ventricular contrast agent (Albunex) may enhance endocardial border delineation and improve the diagnostic value of dobutamine stress echocardiography. METHODS: Fifty-six patients, 38 with myocardial infarction, 16 with angina pectoris and two normal subjects, were enrolled in this study. Dobutamine was infused in scalar doses of 5 to 40 micrograms/kg/min. Intravenous infusion of Albunex (0.15 ml/kg) was administered at rest and during peak dobutamine stress during monitoring of the apical four-chamber view. The left ventricle in the apical four-chamber view was divided into six segments and an endocardial delineation score of 0 to 3 (none to excellent visualization) was given to each segment. RESULTS: Endocardial delineation score was increased after Albunex infusion from 2.0 to 2.3 in the basal-septal, 2.0 to 2.4 in the mid septal, 1.1 to 1.8 in the apical-septal, 0.7 to 1.2 in the apical-lateral, 0.9 to 1.6 in the mid-lateral, and 1.2 to 1.9 in the basal-lateral segments during peak dobutamine administration. Endocardial border resolution in the lateral wall showed greater improvement than in the septal wall after Albunex infusion. Diagnostic values in the left anterior descending artery territory failed to improve with Albunex infusion (sensitivity 82% to 89%, specificity 94% to 89%, and accuracy 86% to 89%), whereas a higher diagnostic accuracy was noted in the left circumflex artery territory with Albunex compared to without Albunex (sensitivity 63% to 81%, specificity 88% to 98%, and accuracy 80% to 93%, p < 0.05). CONCLUSIONS: Contrast agent improves the diagnostic accuracy of dobutamine stress echocardiography in the left circumflex artery territory. PMID- 11281054 TI - [Efficacy of low-dose mutant tissue-type plasminogen activator followed by planned rescue percutaneous transluminal coronary angioplasty as reperfusion therapy for acute myocardial infarction]. AB - OBJECTIVES: The efficacy of injection of a low-dose mutant tissue-type plasminogen activator (mt-PA), monteplase, followed by planned rescue percutaneous transluminal coronary angioplasty (PTCA) was compared with that of primary PTCA. METHODS: A total of 164 patients with acute myocardial infarction within 12 hr from onset were randomly assigned to a treatment with 80 x 10(4) U bolus of monteplase (Group M) or no administration (Group P) by the envelope method, followed by immediate angiography with angioplasty in patients with Thombolysis in Myocardial Infarction (TIMI) flow grade 0, 1 or 2. RESULTS: There were no differences in baseline characteristics between the two groups. Initial angiography showed a higher reperfusion rate (TIMI 2 + 3: 21% + 38% vs 13% + 9%, p < 0.001) and the median time to TIMI 3 was shorter (63 vs 78 min, p < 0.005) in Group M than in Group P, but the final TIMI 3 rate was similar (93% vs 96%). Peak creatine kinase was lower, and predischarge left ventricular ejection fraction measured in 70% of all patients was higher (59 +/- 9% vs 54 +/- 14%, p = 0.02) in Group M than in Group P. Recurrent ischemia with ST elevation occurred in three patients in Group M, but death, re-acute myocardial infarction or stroke did not occur in either group and the rate of bleeding complication was similar (4.9% vs 3.7%). PTCA was performed less frequently in Group M, but medical expenses were comparable in both groups. CONCLUSIONS: Low-dose mt-PA followed by rescue PTCA is effective for early recanalization and preservation of left ventricular function without increases in bleeding complications or medical expenses. These results suggest that low-dose mt-PA should be given to all patients with acute myocardial infarction who are scheduled to undergo primary PTCA. PMID- 11281055 TI - [Outcome of patients with significant coronary stenosis but without ischemic evidence on exercise myocardial perfusion scintigraphy]. AB - OBJECTIVES: The rates of cardiac events and coronary revascularization were evaluated in patients with significant coronary stenosis of more than 75% by the American Heart Association (AHA) classification but no ischemic evidence by exercise myocardial perfusion scintigraphy. METHODS: Subjects were 171 patients (113 males, 58 females, mean age 66 +/- 9 years) undergoing coronary angiography and without scintigraphic evidence of myocardial ischemia. They were divided into two groups according to the severity of coronary artery stenosis based on AHA classification. Group A was composed of 139 patients with more than 75% stenosis (101 patients with 75% stenosis and 38 patients with more than 90% stenosis), and Group B was composed of 32 patients with 50% stenosis. Cardiac events including angina pectoris (n = 63), myocardial infarction (n = 1), heart failure (n = 2) and cardiac death (n = 0), coronary revascularization and predictive factors were evaluated during follow-up of 34 +/- 21 months. Furthermore, the interval between coronary revascularization and exercise myocardial perfusion scintigraphy was estimated. RESULTS: The rates of cardiac events (45%) and coronary revascularization (29%) in Group A were significantly higher than the rate of cardiac events (9%, p < 0.05) and coronary revascularization (6%, p < 0.05) in Group B. Only percentage stenosis and the number of diseased vessels affected the rates of cardiac event and coronary revascularization. CONCLUSIONS: Patients with significant coronary stenosis, but without ischemic evidence by exercise myocardial perfusion scintigraphy, have a relatively high rate of cardiac event and coronary revascularization, especially in patients with severe stenosis or multivessel disease. However, coronary revascularization should not be performed in all patients with significant coronary stenosis. PMID- 11281056 TI - [Usefulness of right ventricular Doppler index for predicting outcome in patients with dilated cardiomyopathy]. AB - OBJECTIVES: Left ventricular Doppler index (LVDI) is believed to be a useful echocardiographic index of systolic and diastolic ventricular function. However, the usefulness of right ventricular Doppler index (RVDI) remains uncertain, especially in dilated cardiomyopathy. The predictive value of RVDI for estimating long-term cardiac events, including cardiac death, was investigated. METHODS: Fifty-nine consecutive patients with dilated cardiomyopathy (41 males and 18 females, mean age 52 +/- 15 years) were enrolled in this follow-up study. RVDI and LVDI were calculated as follows: DI = (isovolumic contraction time + isovolumic relaxation time)/ejection time. RESULTS: During a follow-up period of 3.7 +/- 3.0 years, 27 (46%) of the patients exhibited cardiac events, including cardiac death (n = 9), heart failure (n = 16) and tachyarrhythmias (n = 2) requiring in-hospital treatment. Patients with these cardiac events had higher LVDI and RVDI at the initial follow-up examination, and RVDI had a significant linear correlation with LVDI (LVDI = 0.550 + 0.452 x RVDI, r = 0.530, p = 0.0001). The 6-year survival rate was significantly lower in patients with both LVDI > or = 0.78 and RVDI > or = 0.49 than in other patients (50% vs 75%, respectively, p = 0.009). Cox proportional hazards model analysis showed that RVDI > or = 0.49 was the independent predictor of cardiac events (p = 0.0153) and cardiac death (p = 0.0003). CONCLUSIONS: RVDI is clinically useful for estimating the outcome of patients with dilated cardiomyopathy. PMID- 11281057 TI - [Incidence and severity of coronary artery disease in patients with acute aortic dissection: comparison with abdominal aortic aneurysm and arteriosclerosis obliterans]. AB - OBJECTIVES: The incidence and severity of coronary artery disease were studied in patients with acute aortic dissection (AAD), and compared with coronary artery disease in patients with abdominal aortic aneurysm (AAA) or arteriosclerosis obliterans (ASO). METHODS: A total of 71 patients(42 males, 29 females, mean age 61.4 +/- 10.0 years) with AAD, undergoing coronary angiography between 1988 and 1999, were studied including 38 patients with open communication type and 33 patients with thrombosed type. According to the Stanford classification, 18 patients were type A and 53 patients were type B. Patients with AAD following Marfan syndrome or chest trauma were excluded from the study. Selective coronary angiography was performed in age- and sex-matched patients with AAA(n = 57; 42 males, 15 females, mean age 63.9 +/- 4.6 years) or ASO (n = 95; 66 males, 29 females, mean age 62.4 +/- 9.4 years). Coronary artery disease was defined as > or = 75% stenosis (left main trunk lesion of > or = 50% stenosis) by multidirectional imaging. RESULTS: Significant coronary artery disease was demonstrated in 14 patients with AAD (19.7%), 25 patients with AAA (43.9%), and 49 patients with ASO (51.5%). The incidence of coronary artery disease was significantly lower in the AAD group than in the other two groups (p < 0.05). One vessel disease was present in approximately 70% of the patients with AAD and coronary artery disease. In contrast, multivessel disease was observed in approximately 50% of patients with AAA and ASO. Classification of the patients with AAD according to the blood flow in the false lumen showed coronary artery disease was more highly associated with the thrombosed type [10 (30.3%) of 33 patients] than the open communication type [4 (10.5%) of 38 patients]. Multivariate logistic regression analysis of the patients with AAD showed coronary artery disease was associated with a high serum total cholesterol level (p = 0.025) and the thrombosed type (p = 0.043). CONCLUSIONS: The incidence of coronary artery disease was significantly lower among patients with AAD than among age- and sex-matched patients with AAA or ASO. Coronary artery disease developed in 30% of the patients with the thrombosed type of AAD, although the prognosis seemed to be good. PMID- 11281058 TI - [A 74-year-old woman with patent ductus arteriosus complaining of dyspnea]. PMID- 11281059 TI - [A quarter century ago: a JCC member looks back. No. 32]. PMID- 11281060 TI - [Quantitative estimation by ELISA of IgG anti-D (RH1) antibodies in immunoglobulin preparations and in the sera of immunized donors]. AB - Immunoglobulin preparations of anti-D (RH1) are injected to prevent haemolytic disease of the newborn. Such preparations are obtained by the fractionation of plasma from immunized donors. Measurement of the concentration of IgG anti-D is required to estimate the potency of anti-D preparations and sera from immunized donors. We have developed an ELISA method for the quantification of IgG anti-D. This method included the following steps, sensitization of red cells by anti-D, solubilization of red cell membranes by Triton, and eventually, measurement of IgG anti-D concentration by ELISA. The international reference preparation of anti-D (68/419) was used as a reference. With this method, we measured IgG anti-D concentrations in 5 immunoglobulin preparations of anti-D and in the sera of 10 donors immunized by D antigen. The ELISA results were compared with those obtained by automated hemagglutination. A mean anti-D concentration of 56.2 micrograms/mL was found by ELISA in immunoglobulin preparations. Similar results were obtained by automated hemagglutination (mean 52 micrograms/mL). In the sera of 10 D-immunized donors, anti-D IgG concentration varied from 2.2 to 59.8 micrograms/mL. A good correlation between ELISA and automated hemagglutination was observed in these sera (r = 0.98, p < 10(-7)). In conclusion, the ELISA technique offers an alternative to automated hemagglutination. It requires only the standard equipment necessary for immuno-enzymatic methods. PMID- 11281061 TI - [Regional program to improve the quality of blood transfusions in hospitals: experience of the Poitou-Charentes region]. AB - In the Poitou-Charentes area, a regional pilot program was implemented over a two year-period to improve transfusion safety in public and private hospitals. This program consisted in: (i) an evaluation of the transfusion chain in hospitals; (ii) a regional program to guide hospitals in improving the quality process. Five workgroups were set up. Three persons in each hospital should participate in the workgroup: one representing the administration, one the medical staff and one the nursing staff. After a six months follow-up several hospitals were prompted to implement corrective and preventive measures to improve transfusion safety; (iii) a letter was regularly published to contribute to set-up a regional haemovigilance network. Such a quality improvement program revealed to be a relevant method to steer the changing blood transfusion process in hospitals. PMID- 11281062 TI - [Methodology for the development of a program for following and maintaining the competency of human resources]. AB - The progressive introduction of a management program for the maintenance and assessment of staff competence has also focussed attention on the human factor, a major consideration in risk management and quality control. This article has examined the relevant tools and practical means of application, and proposes a methodology combining a methodical analysis of processes with the determination of the minimal knowledge required for participation in the practical and theoretical training programs that provide a means of objective evaluation. The results obtained in terms of technical, organizational and cultural impact have also been analyzed. PMID- 11281063 TI - [Evaluation of the analytic performance of blood collection tubes (BD Vacutainer SST) for the screening of anti-HIV, anti-HTLV, anti-HCV, anti-HBc, anti-CMV antibodies, and of HBs, P24 HIV antigens, and of alanine aminotransferase]. AB - The Laboratory of Viral Diseases Immunology (Laboratoire d'Immunologie des Maladies Virales) of the Northern Region Blood Bank (Etablissement Francais du Sang Nord de France) performs between 180.000 and 200.000 viral blood qualifications per year. The use of a serum gel separator evacuated tube should contribute to improve the quality of the pre-analytical phase. However, it must not impact negatively the analytical performances. We evaluated such tube within our specific environment and with the various reagents used in routine. The open study compared the BD Vacutainer plain tube (7 mL, non siliconised) with the BD Vacutainer SST tube (6 mL siliconised with serum gel separator) against the anti HIV, anti-HTLV, anti-HCV, anti-HBc, anti-HBs, anti-CMV antibodies, the HBs, HIV P24 antigen and the alanine aminotransferase. The study objectives were to find potential gel interference; to verify the diagnostic sensitivity, reagents specificity, and reproducibility. The results analysis show: equivalent performances with the anti-HIV Ab (Anti HIV 1/2 recombinant--Biotest et Genscreen HIV 1/2--Sanofi), anti HIV WB Ab (New Lav Blot 1--Sanofi), anti-HBs Ab (Enzygnost anti-HBs micro--Behring), anti-HBc Ab (HBc Elisa Test System--Ortho), anti-CMV Ab (Enzygnost anti-CMV IgG + M--Behring) kits; lower performances with: The Vironostika HIV Uni Form II plus 0--Organon kit with a -3.5% signal decrease around the ratio R = 2.7 for positive anti-HIV Ab. The Elisa test System 3 Ag HBs Ortho kit with an increase of the mean ratio of the negative Ag HBs samples; better performances with: the Vironostika HIV 1 Antigen--Organon kit with a +10% signal increase around the threshold ratio R = 1 for positive Ag HIV samples. This deserves further study to verify that the specificity is maintained. The HTLV Type 1 et 2 EIA--Ortho kit with +8% signal increase around the ratio R = 2 for positive anti-HTLV Ab samples without change of the specificity. The Ortho HCV 3.0 Elisa Test System and HTLV Type 1 et 2 EIA kits with a clear and significant improvement of the reproducibility of the anti-HCV and anti-HTLV Ab screenings. The results of this evaluation, together with the intrinsic BD SST tube characteristics, lead to the conclusion that its use would contribute to improve the quality. Because of the specificities of each laboratory, a change of tube type, as with any other material or reagent, request a close monitoring of the first results to confirm the absence of negative effects. PMID- 11281064 TI - [Blood donation. Job description of the function of a physicion in a district subdivision]. AB - Following the 1995 national reorganization of the transfusion system, the Nord Pas de Calais blood transfusion center has modified its blood collection organization, with the creation of four districts divided into three or four subdivisions. This change was part of Quality Assurance implementation in the center. This article provides the job description for a physician in charge of such a subdivision as well as the method chosen to design this description (inspired by the diagram of Hijman). This job was conceived as an association of human skills knowledge with the strategy of the blood center. In addition to his medical activity, the district subdivision physician becomes a manager and an organizer. The method used highlights the participation of all disciplines involved in blood donation. PMID- 11281065 TI - [Hematopoietic differentiation of embryonic stem cells in mice: a model to study the biology of hematopoiesis]. AB - The manipulation of embryonic stem (ES) cells allows to generate mice with specific alteration in any gene. This is therefore an invaluable tool for studying gene function. A number of genes involved in the regulation of hematopoiesis have been inactivated, including genes that encode transcription factors, cytokines and their receptors as well as those encoding for intracellular signalling proteins. Alternatively, ES cells are able to differentiate towards myeloid, lymphoid and endothelial lineages under specific culture conditions. The role of master genes controlling hematopoiesis can be investigated by substituting the in vitro hematopoietic differentiation model of ES cells to mice fabrication. This method can be applied for studying effects of gene inactivation or overexpression of normal or abnormal gene. Interestingly, in vitro differentiation of ES cells recapitulates some aspects of embryonic development, including the emergence of the hemangioblast, the common precursor of hematopoietic and endothelial lineages. Thus, hematopoietic differentiation of ES cells constitutes a model for studying effects of gene manipulation on both hematopoiesis and emergence and commitment of the more hematopoietic primitive cell, the hemangioblast, during embryogenesis. In our studies, we used ES cells inactivated for the c-mpl gene, the thrombopoietin receptor, for dissecting the functions of various intracytoplasmic domain of c-mpl in the response of ES cell derived hematopoietic cells to TPO. PMID- 11281066 TI - [Sleep in fibromyalgia: review of clinical and polysomnographic data]. AB - Fibromyalgia syndrome is a common chronic pain syndrome that is often associated with sleep disturbances characterized by subjective experience of non-restorative sleep. The complaints of sleep disturbances are correlated with polysomnographic features showing clear abnormalities in the continuity of sleep as well as in the sleep architecture. Sleep-recording abnormalities are characterized by a reduced sleep efficiency with increased number of awakenings, a reduced amount of slow wave sleep and an abnormal alpha wave intrusion in non rapid eye movement, termed alpha-delta sleep. These data were confirmed by spectral analysis of sleep showing an increased EEG power density in the higher frequency band and a reduced EEG power density in the lower frequency bands. Moreover, other microstructural aspects of sleep were modified with high frequency of arousals and alpha-K complex reported, both indicators of fragmented sleep. The fibromyalgia symptoms may relate to a non-restorative sleep disorder associated with the alpha-EEG sleep anomalies. However, alpha-EEG sleep anomaly is non-specific for fibrositis, also seen in normal controls during stage 4 sleep deprivation. Moreover, fibromyalgia patients may also experience primary sleep disorder such as sleep apnea or periodic leg movements. The etiology of this common condition is incompletely understood and the existence of a specific entity of fibromyalgia is still a matter of debate. However, several studies have found abnormal brain metabolism of substances such as serotonin implicated in sleep arousal and pain mechanisms and administration of tricyclic antidepressants and selective serotonin reuptake inhibitors may be useful in fibromyalgia. Pain, poor sleep quality and anxiety may contribute to the clinical picture. Several factors such as psychological, environmental, genetic factor, altered serotonin metabolism and altered sleep physiology are involved in the pathogenesis of fibromyalgia. PMID- 11281067 TI - [Myoclonus in the adult: diagnostic approach]. AB - Myoclonus, defined as shock-like involuntary movement, may be physiological or caused by a very wide variety of hereditary and acquired conditions. Because myoclonus can originate from different disorders and lesions affecting quite varied levels of the central and peripheral nervous systems, it represents from many points of view a diagnostic challenge. Moreover, new entities have been recently individualized, such as cortical tremor, which deserve renewed attention. The aim of this review is to propose a rationale for a diagnostic approach based on clinical and electrophysiological grounds. In this setting, we successively address 1) the clinical features allowing a positive diagnosis of myoclonus; 2) the clinical clues to the etiology; 3) the relevance of the clinical context to the diagnosis; and 4) the contribution of neurophysiology. Differentiating myoclonus from tics, spasm, chorea and dystonia can be difficult, and a careful reappraisal of clinical features allowing precise identification is presented. Moreover, the topographical distribution of myoclonus, the temporal pattern of muscle recruitment, the condition of occurrence and the rhythm of the event, may provide clinical clues relevant to the diagnosis. Myoclonus without associated epilepsy, myoclonus with epilepsy, myoclonus with encephalopathy, parkinsonism and/or dementia represent overlapping clinical categories, although they remain useful for the diagnostic approach. Using electrophysiology (including back-averaging EEG, MEG, SEP, C-reflex studies) to determine the origin of myoclonus may not allow us to focus on the underlying condition. Indeed, in many instances, the myoclonus is cortical in origin, but the pathology is found elsewhere. PMID- 11281068 TI - Nap polygraphic recordings after partial sleep deprivation in patients with suspected epileptic seizures. AB - A review of the literature shows that nap recordings make a significant contribution to epilepsy studies, providing evidence of specific EEG findings in patients suspected of having epilepsy. In addition, sleep deprivation can cause paroxysmal EEG activity and clinical seizures. We studied retrospectively 686 patients, 51.8% males and 48.2% females, who had experienced at least one episode classified from the clinical point of view as epileptic in origin. They were divided into six age groups. Patients underwent a two-hour (1 P.M.-3 P.M.) nap video-polygraphic recording (EEG 13 channels using the standard 10-20 system, EOG, ECG, EMG and respiration), following a partial sleep deprivation (1 to 3 h) the night before. A second recording was made in 40 patients. In 35.3% of patients, a complete sleep cycle was obtained; in 64.6% sufficient light and deep NREM sleep was obtained, but not REM stage; in 9.3%, we only observed drowsiness and stage 1 of sleep, and this group was excluded from the analysis. Interictal and/or ictal epileptic discharges were observed during the first nap recording in 245 patients (40.4% of the sample). In addition, in 40 patients (11%) with normal or inconclusive first nap EEG, a second recording was able to demonstrate epileptic abnormalities in 35% of cases. Because of its good cost/benefit ratio and availability in most western laboratories, we consider the 'nap plus partial sleep deprivation' method as advantageous over other activation procedures. PMID- 11281069 TI - Middle-latency auditory evoked potentials in children at high risk for alcoholism. AB - PURPOSE: In the course of a high-risk study for alcoholism, the middle-latency auditory evoked potentials (MAEPs) of children of alcoholics were explored. MATERIAL AND METHODS: A series of auditory clicks (0.1 ms, 60 dB SL, 1.1/s) were used to record the Pa and Pb peaks of the MAEPs in 15 children of alcoholics with a multigenerational family history of alcoholism, and 17 control subjects, ranging from 10 to 14 years of age. RESULTS: The latency of Pb was shorter in the high-risk than in the control group, and there was also a significant risk group by age interaction on Pa latency. The amplitude of Pa was smaller in the children of alcoholics. CONCLUSIONS: The characteristics of the MAEPs of the high-risk subjects did not match the pattern of abnormalities previously observed in chronic alcoholics, which are supposed to be a consequence of the neurotoxic effects of ethanol. Nonetheless, the results showed significant differences in MAEPs between children of alcoholics and controls, pointing to an anomalous pattern of information transmission from thalamus to cortex that should be further analyzed using larger samples in a broader age range. PMID- 11281070 TI - Short latency facilitation between pairs of threshold magnetic stimuli studied in amyotrophic lateral sclerosis. AB - The aim of the present study was to investigate the effects of short latency facilitation between pairs of threshold magnetic stimuli on the motor cortex of amyotrophic lateral sclerosis (ALS) patients, at an interstimulus interval (ISI) of 1-5 ms. As compared to controls, the facilitatory effects normally recorded at ISI 1 and 3 ms were considerably reduced. This suggested that the neuronal circuitry responsible for these effects, which is supposed to be the same as those generating I-waves, is impaired in ALS. PMID- 11281071 TI - Bipolar disorder in children. AB - This article presents an overview of bipolar disorder (BPD) in children, a condition that only recently has been recognized as a legitimate diagnosis. Bipolar disorder in children is underrecognized for many reasons including lack of awareness, diagnostic confusion, and the different clinical picture in children. Available data strongly suggest that prepubertal childhood BPD is a non episodic, chronic, rapid cycling, mixed manic state. It may be comorbid with attention-deficit/hyperactivity disorder (ADHD) and conduct disorder (CD) or it may demonstrate features of ADHD and CD, further complicating recognition and subsequent treatment. Treatment issues are discussed, and some reasons for the urgency of early recognition and treatment are explained. PMID- 11281072 TI - Seclusion & restraint. Understanding recent changes. AB - 1. Seclusion and restraint must be a last resort, emergency response to a crisis situation that appears to present imminent risk of harm to the patient, staff, or others. 2. JCAHO-accredited hospitals billing for Medicare or Medicaid must meet HCFA requirements to maintain compliance. 3. Policy writers and administrators must carefully scrutinize all applicable regulatory standards to ensure all requirements are met. PMID- 11281073 TI - Least to most restrictive interventions. A continuum for mental health care facilities. AB - While this article does not propose to provide answers to the many questions prompted by the new Patient's Rights CoP (1999) or the newly revised JCAHO standards on seclusion and restraint (JCAHO, 2000), a prudent plan of action for any organization serving patients with a behavioral component to their treatment would be to adopt a policy of least restrictive intervention prior to consideration of seclusion and restraint in response to emergency patient care needs. It was the authors' intent to present a representative sample of the available literature and from that to develop a continuum of intervention possibilities ranging from least to most restrictive. In addition, suggestions found in the literature for implementation of least to most restrictive programs have been provided. PMID- 11281074 TI - Stigma. Shades of visibility. PMID- 11281075 TI - Computer addiction. When monitor becomes control center. AB - Computer addiction is a newly recognized problem. While controversy exists about whether computer addiction should be considered a primary psychiatric disorder, clinicians are treating increasing numbers of clients experiencing problems caused by excessive computer use. Case studies are provided that include typical histories and symptoms. Behavioral cognitive therapy is discussed as a treatment approach. The stages of change theory is recommended as a strategy to help clients plan and implement change. PMID- 11281076 TI - New approaches to lung cancer. Changing role for family physicians. PMID- 11281077 TI - A new way of thinking about health care systems? PMID- 11281078 TI - General practice in rural Tanzania. PMID- 11281080 TI - Ophthaproblem. Herpes zoster. PMID- 11281079 TI - Discontinuing antidepressants and benzodiazepines upon becoming pregnant. Beware of the risks of abrupt discontinuation. AB - QUESTION: Two of my patients are planning to become pregnant. One is taking paroxetine and the other lorazepam. We have discussed what to do when they become pregnant and have decided they should stop taking these drugs as soon as pregnancy is confirmed. Is this the right decision? ANSWER: The decision to discontinue these drugs during pregnancy should be based on scientific evidence rather than "hearsay" that women should not take psychotropic medications during pregnancy. Recent epidemiologic studies have documented the relative safety of these drugs, so women should not feel compelled to stop taking them when they become pregnant. If, after receiving appropriate evidence-based information, a woman decides to stop taking the drugs, they should be gradually tapered off to avoid abrupt discontinuation syndrome. PMID- 11281081 TI - Medicolegal file. Right to confidentiality versus duty to disclose. PMID- 11281082 TI - Just the berries. Management of heart failure. PMID- 11281083 TI - Alendronate and male osteoporosis. PMID- 11281084 TI - [Evaluation of the physician-patient relationship competence. Development and validation of an assessment instrument]. AB - OBJECTIVE: To develop and validate the design of a grid that assesses doctor patient relationship skills. DESIGN: Evaluation study of an assessment instrument. SETTING: Private practices and family practice units. PARTICIPANTS: From a sample of volunteers, 100 family physicians either in private practice or in a family practice unit completed the proposed grid independently. MAIN OUTCOME MEASURES: The Cronbach alpha coefficient was used to analyze internal consistency. Factorial analysis was used to determine whether the grid's anticipated dimensions were in fact present. RESULTS: The Cronbach alpha coefficient had a very high value (0.92), indicating that the items in the grid were highly homogeneous. Two key factors emerged from the factorial analysis; the first factor alone (understanding patients' experience) accounted for almost 42% of the variance. CONCLUSION: The proposed grid presents some interesting metrologic qualities. It is short and relatively simple to use to assess relationship skills of future and practising family physicians. The grid must now be further validated using a variety of cases. PMID- 11281085 TI - Prescribing practices and attitudes toward giving children antibiotics. AB - OBJECTIVE: To investigate whether overprescribing is common in treatment of pediatric upper respiratory infections and to examine factors that influence prescribing antibiotics for children. DESIGN: A random, stratified sample of practising family physicians was surveyed with a mailed questionnaire. Initial nonresponders were mailed a second questionnaire. SETTING: British Columbia. PARTICIPANTS: A total of 608 general and family physicians. Response rate was 64%; 392/612 surveys were completed. MAIN OUTCOME MEASURES: Physicians' self reported prescribing practices and knowledge of and attitudes toward using antibiotics for children's upper respiratory tract infections. RESULTS: Relative to treatment guidelines developed for the study, most physicians responded appropriately to the cough (94%) and lobar pneumonia (99.1%) vignettes. More than half the physicians (56.5%) reported they would immediately prescribe antibiotics for tympanic membrane dysfunction, and 79.4% indicated they would prescribe antibiotics for pharyngitis without obtaining a laboratory culture. Approximately 25% of physicians in the study did not believe that prior antibiotic use increased personal risk for acquiring drug-resistant infection, and 23.1% did not believe that antibiotic use was an important factor in promoting resistance in their communities. CONCLUSION: Education in current treatment of pediatric upper respiratory tract illnesses and antimicrobial drug resistance is required. The high response to the questionnaire (64%) and the many requests from physicians to receive the project's educational materials (45%) indicate a high level of interest in this subject. PMID- 11281086 TI - Proton pump inhibitors. Compliance with a mandated step-up program. AB - OBJECTIVE: To assess compliance with a step-up approach to proton pump inhibitor (PPI) therapy before implementation of a new provincial policy to promote histamine-type 2 receptor antagonist (H2RA) use before PPI therapy. DESIGN: Population-based, retrospective, open cohort study using prescribing and medical procedure data from January 1, 1995, to April 30, 1999. SETTING: Health administration databases for the universal health care system in Ontario. PARTICIPANTS: Approximately 1.4 million residents of Ontario older than 65 years. MAIN OUTCOME MEASURES: Proportion of patients who received a trial of H2RA therapy or gastrointestinal diagnostic testing 12 months before starting PPI therapy in 1996. RESULTS: Among the 25,870 patients who met study criteria in 1996, about 63% had received H2RAs 12 months before starting PPI therapy and 73% had had a trial of H2RAs or gastrointestinal diagnostic testing. Repeat analysis for January through April 1999, following the new policy implementation, showed that about 72% of patients had had a trial of H2RAs within 12 months of starting PPI therapy. CONCLUSION: A modest gain (9%) in compliance with using H2RA therapy within 12 months before starting PPI therapy was seen following introduction of the step-up intervention. In future, costs and benefits of potential interventions should be carefully considered before implementing new policies. PMID- 11281088 TI - Case report: acupuncture for carpal tunnel syndrome. Ultrasound assessment of adjunct therapy. PMID- 11281087 TI - Early detection for lung cancer. New tools for casefinding. AB - OBJECTIVE: To review data from published population trials and clinical practice guidelines on screening for lung cancer to provide a recommendation for early detection of lung cancer. QUALITY OF EVIDENCE: Literature was searched via MEDLINE using the MeSH headings "lung neoplasm," "mass screening," "thoracic radiography," and "sputum." Only prospective randomized controlled trials with large numbers of subjects were selected. MAIN MESSAGE: Risk of lung cancer among long-term heavy smokers continues even years after stopping smoking. Risk is highest in smokers with chronic obstructive pulmonary disease. Canadian clinical practice guidelines currently recommend that sputum cytology examination and chest radiography (CXR) not be used for lung cancer screening. This guideline was deducted from four randomized population trials in the 1970s that have serious limitations and applies to asymptomatic adults only. A CXR and sputum cytology examination are indicated in symptomatic current and former smokers older than 45 years with a smoking history of 30 pack-years or more and airflow obstruction defined as a forced expiratory volume in 1 second/forced vital capacity (FEV1/FVC) of 70% or less and a FEV1 lower than 70%. Curative treatment is available for early lung cancer. Substantial advances in innovative technologies for early detection using low-dose spiral CT and newer sputum tests have been made in the last three decades. Additional studies are under way to evaluate these new technologies. CONCLUSION: Primary care physicians have an important role in identifying people at risk of developing lung cancer and in supporting research to evaluate new screening technology. PMID- 11281089 TI - Case report: severe neurologic reaction to ciprofloxacin. PMID- 11281090 TI - Small-group CME using e-mail discussions. Can it work? AB - PROBLEM BEING ADDRESSED: Traditional continuing medical education (CME) approaches do not work well in changing physicians' behaviour, but some promising strategies and technologies might help. Our program sought to meld small-group learning with an Internet e-mail approach. OBJECTIVE OF PROGRAM: In 1994, the Family medicine Education and Research Network (FERN) was developed to support on line discussion among London, Ont, and area family physicians. To support educational, moderated case discussions using e-mail, FERN Dissemination (FERN-D) was introduced to a subgroup of participants. We hoped to increase awareness and use of evidence-based research in clinical practice and to increase use of Internet-based resources for CME. The target group was family physicians in the London area. MAIN COMPONENTS OF PROGRAM: Forty volunteers were recruited and were e-mailed one case every 2 weeks; 34 completed the study. Each case was followed by further postings and, at the end of 2 weeks, by a summary of the group's discussion. Background material for each case was researched and was evidence based. Evaluation was conducted using preintervention and postintervention mailed surveys combined with an e-mail feedback questionnaire and a modified focus group. CONCLUSION: On-line case-based discussion is a promising strategy for encouraging family physicians to access current research. More research is needed to determine whether it can be effectively used to change physicians' practice. PMID- 11281091 TI - Finding the right information at the right time. Part 2: MEDLINE, medical journals, and websites. PMID- 11281093 TI - Residents' page. How to keep up-to-date with a click. PMID- 11281092 TI - When patients have cancer. The Canadian Cancer Society is there to help. PMID- 11281094 TI - Hypothesis: the research page. Odds ratios and relative risks. PMID- 11281095 TI - Use of interreflection and shadow for surface contact. AB - The interaction of light with surfaces results in a number of lighting effects that may serve as valuable visual cues. Previous research on shadows has shown them to be effective in determining the three-dimensional (3-D) layout of a scene, but interreflections have been ignored as cues for spatial layout. Interreflections as well as shadows may help to disambiguate the 3-D layout of objects by providing information about an object's contact with a surface. We generated computer images of a box on an extended textured ground plane that was either in contact with the ground or was slightly above the ground. Images were rendered for four conditions: (1) no shadow + no interreflection, (2) shadow only, (3) interreflection only, and (4) shadow + interreflection. A photometrically incorrect condition was also included. The participants rated the degree of contact for each image on a scale, which was used to generate receiver operating characteristic (ROC) curves and a measure of sensitivity. In the images with no shadows or interreflections, the participants performed at chance levels. Interreflections, shadows, and a combination of interreflections and shadows all resulted in high sensitivity for judging object contact. More important, information from shadows and interreflections can be combined, resulting in near perfect judgment of surface contact. Interreflections and shadows can be effective cues for object contact. PMID- 11281096 TI - Sequential priming of 3-D perceptual organization. AB - In four experiments, the effects of sequential priming on the perceptual organization of complex three-dimensional (3-D) displays were examined. Observers were asked to view stereoscopic arrays and to search an embedded subset of items for an odd-colored target while 3-D orientation of the stimuli was varied randomly between trials. Search times decreased reliably when 3-D stimulus orientation was unchanged on consecutive trials, indicating substantial sequential priming by 3-D stimulus layout. The priming was nonsensory and was independent of priming by additional stimulus characteristics. Finally, priming by 3-D layout was unaffected by observers' foreknowledge of display orientation. Results indicate that perceptual organization of 3-D stimuli is guided by a short term trace of 3-D spatial relationships between stimuli. PMID- 11281097 TI - Eye movements during change detection: implications for search constraints, memory limitations, and scanning strategies. AB - Search, memory, and strategy constraints on change detection were analyzed in terms of oculomotor variables. Observers viewed a repeating sequence of three displays (Scene 1-->Mask-->Scene 2-->Mask...) and indicated the presence-absence of a changing object between Scenes 1 and 2. Scenes depicted real-world objects arranged on a surface. Manipulations included set size (one, three, or nine items) and the orientation of the changing objects (similar or different). Eye movements increased with the number of potentially changing objects in the scene, with this set size effect suggesting a relationship between change detection and search. A preferential fixation analysis determined that memory constraints are better described by the operation comparing the pre- and postchange objects than as a capacity limitation, and a scanpath analysis revealed a change detection strategy relying on the peripheral encoding and comparison of display items. These findings support a signal-in-noise interpretation of change detection in which the signal varies with the similarity of the changing objects and the noise is determined by the distractor objects and scene background. PMID- 11281098 TI - Direction repulsion in unfiltered and ring-filtered Julesz textures. AB - Perceived directions of motion were measured for each of two superposed two dimensional dynamic random patterns consisting of unfiltered or ring-filtered dense random-check (Julesz) textures. One pattern always moved in a cardinal direction (up, down, left, or right), and the other texture always moved in an oblique direction separated from the cardinal component by 20 degrees-80 degrees. Several cardinal/oblique speed ratios were tested. In Experiment 1, the textures were unfiltered. In Experiment 2, the textures were ring filtered and had the same center frequency (1, 2, or 4 cpd). In Experiment 3, a 1-cpd ring-filtered texture was paired with a 2-, 4-, or 8-cpd texture. Subjects consistently misperceived the directions of component motion in these experiments; the angular separation of movement of the two textures was perceptually exaggerated, a phenomenon referred to as direction repulsion (Marshak & Sekuler, 1979). The results show that (1) direction repulsion occurs across at least a fourfold range of spatial frequencies and a sixfold range of speed ratios, (2) direction repulsion varies systematically with speed ratio, and (3) across most conditions, direction repulsion is anisotropic--direction repulsion is more evident in the oblique directions than in the cardinal directions. These findings suggest that the spatiotemporal range of inhibitory interactions involved in motion transparency is much greater than previously appreciated. PMID- 11281099 TI - A variable mapping task produces symmetrical interference between global information and local information. AB - When processing global and local aspects of compound visual figures, a robust finding is that global targets are detected faster and more accurately than local targets. Moreover, unidirectional interference is often observed. Despite the convincing evidence that global information and local information are available together, when attention is focused on the global level, items from the local level often have very little, if any, effect on behavior. If local information is available with global information, then why is global dominance so often observed under such a wide range of conditions? This paper is concerned with the fate of the ignored, and apparently ineffective, local distractors. In our experiments, at least one critical factor was stimulus-response (S-R) mapping. We compared a consistent S-R task, which facilitated a speed advantage for global, with a variable S-R task, which required a higher degree of semantic analysis for each stimulus. The two tasks produced large differences in behavior, showing unidirectional interference in the consistent S-R task, and strong bidirectional interference in the variable S-R task. Thus, the identity of ignored local distractors was available, even under conditions that favored focused attention to global information. The results provide support for a model in which global processing proceeds more quickly at early perceptual stages and in which local processing can catch up if processing demands are increased at later stages. PMID- 11281100 TI - What are the units of visual short-term memory, objects or spatial locations? AB - We investigated whether the capacity of visual short-term memory (VSTM) is defined by number of objects or number of spatial locations. Previous work is consistent with either alternative. To distinguish these factors, we used overlapping stimuli that allowed us to independently manipulate the number of spatial locations while holding constant the number of objects and features to be encoded (Duncan, 1984; Vecera & Farah, 1994). In Experiment 1, the number of spatial locations had no effect on VSTM, suggesting that VSTM is object based. Experiments 2 and 3 ruled out alternative explanations based on perceptual segregation difficulty or decision noise factors. Our results provide additional support to Luck and Vogel's (1997) demonstration that integrated objects form the units of VSTM capacity. PMID- 11281101 TI - How to produce an absent-advantage in visual search. AB - In a series of four experiments, we investigated the conditions under which target-absent responses are faster than target-present responses in visual search. Previous experiments have shown that such an absent-advantage occurs mainly for homogeneous distractors arranged in a regular pattern. From these results, it has been concluded that the absent-advantage is due to perceptual processes, such as grouping by similarity. Our data show that such processes are not sufficient. Rather, the absent-advantage is the result of interactions between perceptual and decisional processes. Certain perceptual conditions, such as randomizing stimulus patterns, lead to specific criteria settings that produce an absent-advantage. That such an account can explain our main results is demonstrated by modeling our data with a modified version of the Guided Search 2 model. PMID- 11281102 TI - Search for multiple targets: remember the targets, forget the search. AB - Models of visual search performance typically assume that search proceeds by sampling without replacement. This requires memory for each deployment of attention. We tested this assumption of memory-driven search using a multiple target search paradigm. We held total set size constant, varied the number of targets in the display, and asked subjects to report whether or not there were at least n targets present, where n was varied by block. This allowed us to measure the time to find each subsequent target. Memory-driven search predicts that reaction time should be a linear function of n. The alternative memory-free search hypothesis predicts an accelerating function. The data falsify the memory driven hypothesis. They were consistent with the memory-free search hypothesis but would also be consistent with memory for a small number of previously attended locations. PMID- 11281103 TI - Attentional capture by color without any relevant attentional set. AB - The aim of the present study was to investigate mechanisms underlying attentional capture by color. Previous work has shown that a color singleton is able to summon attention only in the presence of a relevant attentional set, whereas when a color singleton is not useful for a task, evidence for purely stimulus-driven attentional capture is controversial. Three visual search experiments (T-L task) were conducted using a method different from that based on set sizes, consisting of monitoring target-singleton distance in a unique display size. In Experiment 1, we demonstrated that attention can be summoned in a real stimulus-driven manner by an irrelevant color singleton. Experiment 2A extended this observation, showing that the color singleton attracted attention even when capture was detrimental. However, Experiment 2B showed that such capture can be strategically prevented. Finally, in Experiment 3, we examined whether such a capture was due to a spatial shift or to a filtering cost, providing evidence supporting the shift hypothesis. Stimulus-driven capture was observed when color was neither the defining nor the reported target attribute (Yantis, 1993) and when subjects naive of visual search tasks were used. The present results give experimental support to many contemporary models of visual attention. PMID- 11281104 TI - Contingent attentional capture or delayed allocation of attention? AB - Under certain circumstances, external stimuli will elicit an involuntary shift of spatial attention, referred to as attentional capture. According to the contingent involuntary orienting account (Folk, Remington, & Johnston, 1992), capture is conditioned by top-down factors that set attention to respond involuntarily to stimulus properties relevant to one's behavioral goals. Evidence for this comes from spatial cuing studies showing that a spatial cuing effect is observed only when cues have goal-relevant properties. Here, we examine alternative, decision-level explanations of the spatial cuing effect that attribute evidence of capture to postpresentation delays in the voluntary allocation of attention, rather than to on-line involuntary shifts in direct response to the cue. In three spatial cuing experiments, delayed-allocation accounts were tested by examining whether items at the cued location were preferentially processed. The experiments provide evidence that costs and benefits in spatial cuing experiments do reflect the on-line capture of attention. The implications of these results for models of attentional control are discussed. PMID- 11281105 TI - Attending to the parts of a single object: part-based selection limitations. AB - Studies of object-based attention have demonstrated poorer performance in dividing attention between two objects in a scene than in focusing attention on a single object. However, objects often are composed of several parts, and parts are central to theories of object recognition. Are parts also important for visual attention? That is, can attention be limited in the number of parts processed simultaneously? We addressed this question in four experiments. In Experiments 1 and 2, participants reported two attributes that appeared on the same part or on different parts of a single multipart object. Participants were more accurate in reporting the attributes on the same part than attributes on different parts. This part-based effect was not influenced by the spatial distance between the parts, ruling out a simple spatial attention interpretation of our results. A control study demonstrated that our spatial manipulation was sufficient to observe shifts of spatial attention. This study revealed an effect of spatial distance, indicating that our spatial manipulation was adequate for observing spatial attention. The absence of a distance effect in Experiments 1 and 2 suggests that part-based attention may not rely entirely on simple shifts of spatial attention. Finally, in Experiment 4 we found evidence for part-based attention, using stimuli controlled for the distance between the parts of an object. The results of these experiments indicate that visual attention can selectively process the parts of an object. We discuss the relationship between parts and objects and the locus of part-based attentional selection. PMID- 11281106 TI - The midstream order deficit. AB - The relative order of an auditory sequence can be more difficult to apprehend when it is presented repeatedly without pause (i.e., cycling) than when it is presented only once (Warren, Obusek, Farmer, & Warren, 1969). We find that this phenomenon, referred to as the midstream order deficit (MOD), can also occur with visual stimuli. The stimuli need not form separate perceptual "streams," and the effect can occur with presentation rates as slow as five items per second, even though the identification of individual letters is very accurate at this rate. However, if the first item of the sequence is visually very distinct from the preceding items, relative order reports can be as accurate in the cycling condition as in the single-presentation condition. Our results suggest that the MOD is not due to masking, attentional blink, repetition blindness, Reeves and Sperling's (1986) order illusion, memory limitations, or decision criteria. The MOD may reflect an attentional cost to the initiation of order encoding, which is distinct from the allocation of attention is required in order to detect and identify individual items. To initiate order encoding successfully, one's attention must be set for, or captured by, an initial salient event. PMID- 11281107 TI - The cost of expecting events in the wrong sensory modality. AB - We examined the effects of modality expectancy on human performance. Participants judged azimuth (left vs. right location) for an unpredictable sequence of auditory, visual, and tactile targets. In some blocks, equal numbers of targets were presented in each modality. In others, the majority (75%) of the targets were presented in just one expected modality. Reaction times (RTs) for targets in an unexpected modality were slower than when that modality was expected or when no expectancy applied. RT costs associated with shifting attention from the tactile modality were greater than those for shifts from either audition or vision. Any RT benefits for the most likely modality were due to priming from an event in the same modality on the previous trial, not to the expectancy per se. These results show that stimulus-driven and expectancy-driven effects must be distinguished in studies of attending to different sensory modalities. PMID- 11281108 TI - A comparison of tactile spatial sensitivity on the palm and fingerpad. AB - Studies of tactile spatial pattern perception have, for the most part, been carried out using the fingerpad. On the basis of these studies, models have been developed linking spatial pattern identification and resolution with underlying neural structures. It has been suggested that with appropriate scaling, these models would apply to the processing of spatial patterns presented to other sites on the body. Spatial sensitivity was examined on another site on the body, the palm, using two measures, letter identification and grating orientation. The results from these measures were compared with results from similar studies conducted on the fingerpad and with estimates of the density of innervation of the fingerpad and palm. To produce levels of performance similar to those on the fingerpad required letters on the palm 50 mm in height, seven to nine times larger than those used on the fingerpad. Gratings had to be six to more than seven times larger on the palm to produce the same levels of performance achieved on the fingerpad. For the two types of receptor systems sensitive to spatial information, the ratio of density of innervation between the fingerpad and the palm is estimated to be 5.7:1 and 8.8:1. Performance of spatial tasks on the palm can be predicted quantitatively from fingerpad data with a moderate degree of accuracy. Qualitative comparisons between the palm and fingerpad data indicate that spatial patterns are processed similarly at the two sites. PMID- 11281109 TI - The depth and selectivity of suppression in binocular rivalry. AB - Binocular rivalry occurs when the two eyes are presented with incompatible stimuli and the perceived image alternates between the two stimuli. The aim of this study was to find out whether the periodic perceptual loss of a monocular stimulus during binocular rivalry is mirrored by a comparable loss of contrast sensitivity. We presented brief test stimuli to one eye while its conditioning stimulus was dominant or suppressed. The test stimuli were varied widely across four stimulus domains--namely, the relative stimulation of medium- and long wavelength-sensitive cones, duration, spatial frequency, and grating orientation. The result in each case was the same. Suppression depended slightly or not at all on the type of test stimulus, and contrast sensitivity during suppression was around 64% of that during dominance. The effect of suppression on sensitivity is therefore very weak, relative to its effect on the perceived image. Furthermore, suppression was largely independent of the similarity between the conditioning and the test stimuli, indicating that our results are better explained by eye suppression than by stimulus suppression. A model is presented to account for the small, monocular sensitivity loss during suppression: It assumes that test detection precedes conditioning stimulus perception in the visual pathway. PMID- 11281110 TI - Category discriminability, base-rate, and payoff effects in perceptual categorization. AB - The optimality of perceptual categorization performance under manipulations of category discriminability (i.e., d' level), base rates, and payoffs was examined. Base-rate and payoff manipulations across two category discriminabilities allowed a test of the hypothesis that the steepness of the objective reward function affects performance (i.e., the flat-maxima hypothesis), as well as the hypothesis that observers combine base-rate and payoff information independently. Performance was (1) closer to optimal for the steeper objective reward function, in line with the flat-maxima hypothesis, (2) closer to optimal in base-rate conditions than in payoff conditions, and (3) in partial support of the hypothesis that base-rate and payoff knowledge is combined independently. Implications for current theories of base-rate and payoff learning are discussed. PMID- 11281111 TI - Opposing views of revaccination strategies. PMID- 11281112 TI - Beware of backfire. PMID- 11281113 TI - Customer service. Enjoying an e-visit. PMID- 11281114 TI - HIPAA. A CEO's guide to regulations and compliance. AB - American hospitals are expected to spend up to $24 billion to comply with the Health Insurance Portability and Accountability Act, but the new law may help them finally realize the efficiencies and reductions in administrative costs long promised by new technology. This foldout section provides checklists to help you measure your readiness. PMID- 11281115 TI - Leriche Memorial Lecture at 24th World Congress 'Takayasu's arteritis in Asia'. PMID- 11281116 TI - Risk management and infection control--time to get our priorities right in the United Kingdom. PMID- 11281117 TI - Clinical governance and infection control in the United Kingdom. AB - The recent organizational changes in the NHS have at their core the concept of clinical governance. Although initially poorly defined and understood this term has now taken on a clear identity, placing quality alongside fiscal probity and corporate governance at the top of NHS priorities. Integral to clinical governance are the basic elements of clear national standards for services and treatments that are to be locally delivered through assured, monitored, high quality healthcare. It is within this framework that workers in infection control must develop their own methods of applying clinical governance. This review explores the implications that the strategy of clinical governance holds for the speciality of infection control, emphasizing the benefits its active adoption can bring and highlighting the key relevance of clinical risk management in this setting. It illustrates clinical governance as a tool to engage colleagues on a multi-disciplinary front, most particularly the crucial link to senior Trust management. PMID- 11281118 TI - The organization of infection control in Belgium. AB - The authors describe the organization of infection control in Belgium with respect to official regulations, the tasks and the training of the infection control doctor and of the infection control nurse, functioning of the infection control committee, the financing and the availability of guidelines. PMID- 11281119 TI - Ninth International Symposium on Staphylococci and Staphylococcal Infection (ISSSI), Kolding, Denmark 14-17 June 2000. PMID- 11281120 TI - First report of Neisseria meningitidis intermediately resistant to penicillin in Croatia. PMID- 11281121 TI - Bifunctional enzyme 6'-N-aminoglycoside acetyltransferase-2"-O-aminoglycoside phosphotransferase in Lactobacillus and Pediococcus isolates of animal origin. PMID- 11281122 TI - Discrepant analysis is still at large. PMID- 11281124 TI - Re: prior history and its impact on criminal recidivism. PMID- 11281125 TI - Addiction professionals' attitudes regarding treatment of nicotine dependence. PMID- 11281126 TI - 'It's like everything--you just roll with it.' Interview by Marc Peyser. PMID- 11281123 TI - Modulation of Kv3 potassium channels expressed in CHO cells by a nitric oxide activated phosphatase. AB - 1.Voltage-gated K+ channels containing Kv3 subunits play specific roles in the repolarization of action potentials. Kv3 channels are expressed in selective populations of CNS neurons and are thought to be important in facilitating sustained and/or repetitive high frequency firing. Regulation of the activity of Kv3 channels by neurotransmitters could have profound effects on the repetitive firing characteristics of those neurons. 2.Kv3 channels are found in several neuronal populations in the CNS that express nitric oxide synthases (NOSs). We therefore investigated whether Kv3 channels are modulated by the signalling gas nitric oxide (NO). 3. We found that Kv3.1 and Kv3.2 currents are potentially suppressed by D-NONOate and other NO donors. The effects of NO on these currents are mediated by the activation of guanylyl cyclase (GC), since they are prevented by Methylene Blue, an inhibitor of GC, and by ODQ, a specific inhibitor of the soluble form of GC. Moreover, application of 8-Br-cGMP, a permeant analogue of cGMP, also blocked Kv3.1 and Kv3.2 currents. 4.KT5283, a cGMP-dependent protein kinase (PKG) blocker, prevented the inhibition of Kv3.1 and Kv3.2 currents by D NONOate and 8-Br-cGMP. This indicates that activation of PKG as a result of the increase in intracellular cGMP levels produced by D-NONOate or 8-Br-cGMP is necessary for channel block. 5. Although the effects of NO on Kv3.1 and Kv3.2 channels require PKG activity, two observations suggest that they are not mediated by phosphorylation of channel proteins: (a) the reagents affect both Kv3.2 and Kv3.1 channels, although only Kv3.2 proteins have a putative PKA-PKG phosphorylation site, and (b) mutation of the PKA-PKG phosphorylation site in Kv3.2 does not interfere with the effects of NO or cGMP. 6. The inhibitory effects of NO and cGMP on Kv3.1 and Kv3.2 currents appear to be mediated by the activation of serine-threonine phosphatase, since they are blocked by low doses of okadaic acid. Furthermore, direct intracellular application of the catalytic subunit of protein phosphatase 2A inhibited Kv3.2 currents, indicating that activity of PKG-induced phosphatase is necessary and sufficient to inhibit these channels. 7. The results suggest that basal phosphorylation of Kv3 channel proteins is required for proper channel function. Activation of phosphatases via NO or other signals that increase cGMP might be a potent mechanism to regulate Kv3 channel activity in neurons. PMID- 11281127 TI - [Meta-analyses and systematic errors in publications]. PMID- 11281128 TI - L-Tyrosine beta-naphthylamide is a potent competitive inhibitor of tyramine N (hydroxycinnamoyl)transferase in vitro. AB - L-Tyrosine beta-naphthylamide, a synthetic substrate designed to measure tyrosine aminopeptidase activity, is a potent inhibitor of hydroxycinnamoyl-CoA:tyramine N (hydroxycinnamoyl)transferase (THT) purified from elicited tobacco cell suspension cultures. The inhibition is competitive, with the inhibitor binding reversibly to the tyramine binding site of the enzyme. Similar results were obtained with THT extracted from elicited potato cell-suspension cultures. Ki values were found to be 0.66 microM for the enzyme from tobacco and 0.3 microM for the enzyme from potato. L-Tyrosine 7-amido-4-methylcoumarin, a fluorogenic substrate for tyrosine aminopeptidases, the structure of which is close to that of L-tyrosine beta-naphthylamide. was also a powerful inhibitor, but slightly less effective with Ki values of 0.72 and 0.42 microM for tobacco and potato THT, respectively. L-Tyrosine beta-naphthylamide was rapidly hydrolysed when fed in vivo to tobacco or potato cell cultures or when incubated in crude enzymic extracts prepared from these cultures. This hydrolysis, which is presumably catalysed by aminopeptidases, precludes the use of L-tyrosine amides as inhibitors of THT in vivo. PMID- 11281129 TI - Interaction of metal ions with lupin seed conglutin gamma. AB - Various metal ions were capable of aggregating and precipitating conglutin gamma, an oligomeric glycoprotein purified from Lupinus albus seeds, at neutral pH values. The most effective metal ions, at 60-fold molar excess to the protein, were Zn2+, Hg2+ and Cu2+; a lower influence on the physical status of conglutin gamma was observed with Cr3+, Fe3+, Co2+, Ni2+, Cd2+, Sn2+, and Pb2+, while Mg2+, Ca2+ and Mn2+ had no effect at all. The insolubilisation of the protein with Zn2+, which is fully reversible, strictly depended on both metal concentration and pH. with middle points of the sharp transitions at three-fold molar excess and pH 6.5, respectively. Conglutin gamma is also fully retained on a metal affinity chromatography column at which Zn2+ and Ni2+ were complexed. A drop of pH below 6.0 and the use of chelating agents, such as EDTA and imidazole, fully desorbed the protein. A slightly lower binding to immobilised Cu2+ and Co2+ and no binding with Mg2+, Cd2+ and Mn2+ were observed. The role of the numerous histidine residues of conglutin gamma in the binding of Zn2+ is discussed. PMID- 11281130 TI - Patterns of phenylpropanoids in non-inoculated and potato virus Y-inoculated leaves of transgenic tobacco plants expressing yeast-derived invertase. AB - The patterns of secondary metabolites in leaves of yeast invertase-transgenic tobacco plants (Nicotiana tabacum L. cv. Samsun NN) were analyzed. Plants expressing cytosolic yeast-derived invertase (cytInv) or apoplastic (cell wall associated) yeast invertase (cwInv) showed a characteristic phytochemical phenotype compared to untransformed controls (wild-type plants). The level of phenylpropanoids decreased in the cytInv plants but increased in the cwInv plants, which showed an induced de novo synthesis of a caffeic acid amide, i.e. N caffeoylputrescine. In addition, the level of the coumarin glucoside scopolin was markedly enhanced. Increased accumulation of scopolin in the cwInv plants is possibly correlated with the induction of defense reactions and the appearance of necrotic lesions similar to the hypersensitive response caused by avirulent pathogens. This is consistent with results from potato virus Y-infected plants. Whereas there was no additional increase in the coumarins in leaves following infection in cwInv plants, wild-type plants showed a slight increase and cytInc a marked increase. PMID- 11281131 TI - Predicting quantitative phytochemical markers in single Echinacea plants or clones from their DNA fingerprints. AB - Amplified restricted fragment length polymorphism (AFLP) data analysis was found to be a statistically significant predictor of phytochemical markers in cultivated Echinacea purpurea germplasm and some related wild species. Over 50 accessions grown under greenhouse conditions were subjected to AFLP analysis and the same assessed for content of tetraene and cichoric acid by high pressure liquid chromatography. The first and second canonical correlation of DNA variables and the phytochemical variables were significant. Individual regressions of cichoric acid and dodeca-2E, 4E, 8Z, 10E/Z-tetraenoic acid isobutyl amide predicted by DNA polymorphism analysis against actual HPLC determined values were nearly linear. Mantel's test showed that there was a weak correlation but a strong association of values of the phytochemical variables and the DNA polymorphism data. PMID- 11281132 TI - Biosynthesis of the bicycloalternarenes, mixed terpenoids of Alternaria alternata. AB - By incorporation of [2-13C]-mevalonate, [1-13C]-acetate and [1-13C]-glucose we could reveal that the phytopathogenic fungus Alternaria alternata biosynthesized the mixed terpenoids bicycloalternarenes via the classic mevalonate pathway. The polyketid pathway does not participate in the biosynthesis of bicycloalternarenes, because there is no incorporation of [13C]-acetate into the C-ring of these compounds. The labelling pattern in this nonterpenoid part of bicycloalternarenes after feeding with [1-13C]-glucose and [U-13C6]-glucose, respectively, allows the assumption that metabolites of the shikimate pathway are involved. PMID- 11281133 TI - Distribution of 8-oxygenated leaf-surface flavones in the genus Ocimum. AB - Nine species of Ocimum (Lamiaceae) were surveyed for leaf-surface flavonoids by means of HPLC with diode array detection and atmospheric pressure chemical ionisation (APCI) mass spectrometry. The analysis revealed the presence of 23 different flavones, most of which were identified by comparing their UV and mass spectra with those of standards. Almost all taxa investigated contained flavones methoxylated in the 6- and 8-positions, such as nevadensin, xanthomicrol and gardenin B. The same taxa also produced flavones methoxylated in the 6-position but hydroxylated in the 8-position, including isothymusin (5,8,4'-trihydroxy-6,7 dimethoxyflavone), pedunculin (5,8-dihydroxy-6,7,4'-trimethoxyflavone) and a new flavone, 5,7,8-trihydroxy-6,4'-dimethoxyflavone, which was given the trivial name pilosin. This compound was isolated from O. americanum var. pilosum and also detected as a minor constituent in O. x citriodorum leaf extracts. Its molecular structure was elucidated by means of NMR spectroscopy. 8-Oxygenated flavones were absent only from O. lamiifolium. APCI mass spectrometry of the flavonoids revealed that the product ions formed by collision induced dissociation of the protonated molecule provided structural information about the substitution pattern of the A-ring. The chemotaxonomic and biogenetic implications of the results are discussed. PMID- 11281134 TI - Phytoecdysteroids in the genus Asparagus (Asparagaceae). AB - Phytoecdysteroids, plant steroids which are analogues of invertebrate steroid hormones, probably contribute to the deterrence of phytophagous invertebrate predators. They also seem to possess antimicrobial activity and several pharmaceutical and medicinal benefits have been ascribed to them. Here. we present a survey of seeds of 16 species of the genus Asparagus (Asparagaceae), including the crop species A. officinalis, for ecdysteroid agonists (including phytoecdysteroids) and antagonists. Seven species were found to contain ecdysteroids with levels ranging from just detectable (A. racemosus and A. sarmentosus) to relatively high (A. laricinus). RP-HPLC/RIA/bioassay has been used to separate positive extracts of four species (A. falcatus, A. laricinus, A. ramosissimus and A. scandens) and analyse the ecdysteroid profiles. The identities of the major ecdysteroids were confirmed by NP-HPLC. Seeds of A. officinalis do not contain detectable levels of ecdysteroids, but leaves, stems and roots contain low levels (detectable by RIA). This indicates that A. officinalis retains the genetic capacity to synthesise ecdysteroids and that future strategies could be developed for enhanced protection of asparagus spears through elevated ecdysteroid levels. PMID- 11281136 TI - A computer-assisted approach for chemotaxonomic studies--diterpenes in Lamiaceae. AB - This paper describes a new computer approach for chemotaxonomic studies. The methodology employed enables the search for chemical substructures as taxonomic descriptors using an expert system built with plant natural products. The operation of the system was tested with diterpenes as taxonomic markers in Lamiaceae. PMID- 11281135 TI - A species-selective allelopathic substance from germinating sunflower (Helianthus annuus L.) seeds. AB - From the exudate of germinating sunflower (Helianthus annuus L.) seeds was isolated a stereoisomer of diversifolide, 4, 15-dinor-3-hydroxy-1(5)-xanthene 12,8-olide (designated sundiversifolide) as determined by analysis of its IR, APCI-, ESI- and HR-MS and 13C and 1H NMR spectra. This substance inhibited shoot and root growth of cat's-eyes by about 50% at a concentration of 30 ppm. It also showed species-selective activity on the shoot and root growth of tested plants. When cat's-eyes seeds were incubated together with sunflower seeds, the cat's eyes growth was inhibited. Furthermore, it was detected from an extract of river sand when sunflower seeds were incubated on the sand. These results indicate that sundiversifolide has an allelopathic function in sunflower plants. PMID- 11281137 TI - Effects of temperature on polyunsaturation in cytostatic lipids of Haslea ostrearia. AB - Unusual chemicals produced by the-'blue oyster' diatom, Haslea ostrearia, include the water-soluble blue pigment marennine and numerous polyunsaturated sesterterpene oils or haslenes. Aqueous extracts of the alga exhibit in vitro and in vivo activities against human lung cancer cells and anti-HIV effects. Here we report that three haslenes also demonstrate in vitro cytostatic action against a human lung cancer cell line. The most active haslene is the most unsaturated and unsaturation in the haslenes increases with increasing algal growth temperature. PMID- 11281138 TI - Membrane disruption and enzyme inhibition by naturally-occurring and modified chacotriose-containing Solanum steroidal glycoalkaloids. AB - Naturally-occurring 3beta-O-chacotriosides of solasodine (solamargine), of its 22S, 25S isomer tomatidenol (beta-solamarine), and of solanidine (chaconine), as well as ring E- and F-modified derivatives of solamargine were prepared and assayed in order to assess the relevance of aglycone structural features to membrane-disruption and enzyme-inhibitory activities of the related glycoalkaloids. A ring E-opened dihydro-derivative of solasodine (the chacotrioside of dihydrosolasodine A) did not bind to cholesterol, stigmasterol or ergosterol in vitro, disrupt PC/cholesterol liposomes or mammalian erythrocytes. or inhibit acetylcholinesterase in vitro. It did not synergise with the solatrioside of dihydrosolasodine A or solasonine (nor did solamargine with dihydrosolasodine A solatrioside) in haemolysis tests. The ring F modified derivative, N-nitrososolamargine, did not inhibit acetylcholinesterase in vitro, but lysed liposomes at > or = 150 microM and pH 7. Increasing the pH to 8 (but not 9) further enhanced disruption. The combination of N-nitrososolamargine and solasonine did not cause any disruption of liposomes. Beta-solamarine showed no anti-acetylcholinesterase activity in vitro at up to 100 microM, but disrupted liposomes at 75 and 150 microM, although not to the extent caused by solamargine or chaconine. In combination with both the (inactive) solatriosides, solasonine and solanine, 75 microM beta-solamarine produced synergistic effects, with liposome disruption greater than 150 microM beta-solamarine alone. Beta solamarine, solamargine and chaconine showed similar haemolytic activity. Beta solamarine synergised with the solatriosides solasonine and solanine in disrupting erythrocytes. Preliminary structure-activity relationships were evaluated for the active chacotriosides in an attempt to define the scope and limitations of this model study. PMID- 11281139 TI - Furanocoumarins from Dorstenia gigas. AB - A series of linear and angular prenylated furanocoumarins and a benzofuran derivative were isolated from leaves and twigs of Dorstenia gigas (Moraceae), a plant occurring endemically on Socotra Island (Yemen). The structures were elucidated by spectroscopic methods (NMR, MS, UV) and chemical derivatization. PMID- 11281140 TI - Tetrahydroisoquinoline-monoterpene and iridoid glycosides from Alangium lamarckii. AB - From the water soluble fraction of the dried fruits of Alangium lamrckii, four tetrahydroisoquinoline-monoterpene glycosides, 6-O-methyl-N-deacetylisoipecosidic acid, 7-O-methyl-N-deacetylisoipecosidic acid, 6,7-di-O-methyl-N deacetylisoipecosidic acid and 6"-O-alpha-D-glucopyranosyl-6-O-methyl-N deacetylisoipecosidic acid, and an iridoid glycoside, 6'-O-alpha-D glucopyranosylloganic acid, were isolated, together with six known compounds. The structures of the previously unknown compounds were determined by spectroscopic and chemical means. The significance of these glucosides in the biogenesis of Alangium alkaloids is discussed; 6-O-methyl-N-deacetylisoipecosidic acid was also chemically converted into 10-O-demethylprotoemetine and dihydroisoalangine. PMID- 11281141 TI - Flavonoids and phenylpropanoid derivatives from Campanula barbata. AB - Four new phenylpropanoid derivatives, barbatosides A-D, and a new catechin, barbatoflavan, were isolated from the whole plant of Campanula barbata L. (Campanulaceae) and identified as wahlenbergioside-3'-O-glucoside, wahlenbergioside-3'-O-(2'''-(p-methoxycinnamoyl))-glucoside, wahlenbergioside-3' O-(4'''-(trans-p-coumaroyl))-glucoside, wahlenbergioside-3'-O-(4"'-(cis-p coumaroyl))-glucoside and 3-acetyl-5-methoxy-7,3',4'-trihydroxy-8-O-glucoside flavan-3-ol, respectively, by spectroscopic methods. In addition, four flavonols were isolated and identified as kaempferol-3-O-glucoside, kaempferol-3-O rutinoside, quercetin-3-O-glucoside and quercetin-3-O-rutinoside. Barbatoflavan demonstrated scavenging properties towards the DPPH radical. PMID- 11281142 TI - Alkaloids from Crinum moorei. AB - Thirteen alkaloids were isolated from Crinum moorei two of which are new. These are 3-[4'-(8'-aminoethyl)phenoxy] bulbispermine and mooreine. The structures of the new alkaloids were determined by spectroscopic methods. PMID- 11281143 TI - Perfusion-related parameters in intravoxel incoherent motion MR imaging compared with CBV and CBF measured by dynamic susceptibility-contrast MR technique. AB - OBJECTIVE: Perfusion-related parameters obtained by intravoxel incoherent motion (IVIM) MR imaging (MRI) were compared with cerebral blood volume and flow (CBV and CBF), retrieved by dynamic susceptibility-contrast (DSC) MRI. MATERIAL AND METHODS: Twenty-eight volunteers (average age 68.5 years) were investigated. Spin echo echo-planar imaging with IVIM-encoding gradients was employed (36 different b values, 0-1200 s/mm2). The perfusion fraction and the pseudo-diffusion coefficient were calculated for regions in thalamus gray matter and frontal white matter, using asymptotic and full fitting. In DSC-MRI, a Gd-DTPA-BMA contrast agent bolus was monitored using simultaneous-dual FLASH. Deconvolution of the measured tissue concentration-versus-time curve with an arterial input function from the carotid artery was applied, and maps of CBV and CBF were calculated. RESULTS: The correlation between the perfusion fraction and CBV was r=0.56 (p<0.0000006) using asymptotic fitting, and r=0.35 (p<0.0004) when full fitting was applied. Average CBF was 41.5 ml/(min 100 g), to be compared with the IVIM based value of 63.6 ml/(min 100 g), obtained from the median value of the pseudo diffusion coefficient in combination with assumptions about capillary network structure. CONCLUSION: The IVIM concept provided results that agreed reasonably with conventional CBV and CBF. The non-linear fitting to noisy signal data was problematic, in accordance with previously presented simulations. PMID- 11281144 TI - Re: Gelig Thurfjell M., et al. Local breast cancer recurrence caused by mammographically guided punctures. Acta Radiol. 41 (2000), 435. PMID- 11281145 TI - 1975: the ADA comes to Chicago. PMID- 11281146 TI - Surface X-ray absorption spectroscopy: principles and some examples of applications. AB - The application of X-ray spectroscopy to surface problems has started almost at the same time as that in materials science. While the theoretical bases are obviously the same, SEXAFS has some experimental peculiarities which are reviewed here. Some examples of this technique will then be given. PMID- 11281147 TI - Study of the local atomic strain field in a Zr-doped TiAl intermetallic alloy by EXAFS and ab initio FLAPW calculations. AB - The mechanical properties of gamma-TiAl-based intermetallic alloys are strongly influenced by Ti d-Ti d and Al p-Ti d hybridizations. These directional bonds supply good mechanical properties at high temperature but they are also associated with a low ductility at room temperature. Small amounts (about 3%) of additional elements can improve this behaviour noticeably and modify the mechanical properties. In the present case, the addition of zirconium in the gamma-TiAl-based alloys has been reported to increase their ductility at low temperature. Beyond the modifications of the directional bonds which come directly from the band structure, the different atomic volumes of the solute atoms induce local deformation fields which interact with the dislocations. We present ab initio calculations (FLAPW method) compared with an EXAFS study which the aim is to determine the site preference of zirconium in the TiAl lattice and the deformation fields around the solute atoms. PMID- 11281148 TI - ELNES investigations of the oxygen K-edge in spinels. AB - The results of a systematic study of the oxygen K-edge electron energy-loss spectroscopy (ELNES) from a series of aluminium- and chromium-containing spinels are presented. Extra fine structure in the region up to 10 eV above the edge onset is observed for the chromium-containing compounds and is assigned to transitions to states created by mixing of oxygen 2p and metal 3d orbitals. The experimental data has been simulated using the multiple scattering code, FEFF8. Good agreement was obtained in the case of magnesium aluminate, but relatively poor agreement was obtained in the case of the chromites. The possible fingerprints in the oxygen K-edge ELNES corresponding to a high degree of inversion the spinel structure and to a tetragonal distortion of the cubic structure are discussed. PMID- 11281149 TI - Observation of oxygen vacancy ordering and segregation in perovskite oxides. AB - Oxygen vacancies are known to dominate the overall electrical behavior of perovskite oxides, which are used in many applications. Although theories have been developed to explain the effect of these vacancies and the defect chemistry of perovskites, there has yet to be incontrovertible evidence of the fundamental origins of the structure-property relationship. However, recently developed technologies in scanning transmission electron microscopy, such as Z-contrast imaging and EELS combined with in-situ heating experiments, provide a new opportunity to address vacancy characteristics and defect chemistry on the basic atomic level. In this paper we discuss the practical aspects of these techniques and demonstrate their application to the characterization of defect chemistry and vacancies in ordered micro-domains, at domain boundaries and at grain boundaries. PMID- 11281150 TI - Valence electron energy loss study of Fe-doped SrTiO3 and a sigma13 boundary: electronic structure and dispersion forces. AB - Valence electron energy loss spectroscopy in a dedicated scanning transmission electron microscope has been used to obtain the interband transition strength of a sigma13 tilt grain boundary in SrTiO3. In a first step the electronic structure of bulk SrTiO3 has been analysed quantitatively by comparing VEELS spectra with vacuum ultraviolet spectra and with ab initio density of states calculations. The electronic structure of a near sigma13 grain boundary and the corresponding dispersion forces were then determined by spatially resolved VEELS. Also the effects of delocalization of the inelastic scattering processes were estimated and compared with results from the literature. PMID- 11281151 TI - Success and limits of common final-state approximations. AB - Several current models for the treatment of final-state effects in the calculation of electron energy loss near edge structures are studied. Ab-initio calculations for the corresponding electronic reference configurations were carried out for the atoms Li to Kr. Edge onset energies can be directly obtained as energy differences between ground and excited state only if the excitation is explicitly accounted for; implicit schemes such as the Z + 1 or Slater's transition state approximation yield satisfactory results only for the K-edge of atoms up to phosphorous. Concerning the intensity modulation of the edge, however, the Z + 1 approximation exhibits the closest overall agreement with the exact K-shell excitation. Other approaches model the electron-hole interaction more effectively for the d-elements, but show pronounced, non-continuous discrepancies for the noble gases and the early p-elements. PMID- 11281152 TI - Density-functional modelling of core-hole effects in electron energy-loss near edge spectra. AB - A series of (MgO)n supercells (n = 1, 4, 8, 16, 32) with three-dimensional periodic boundary conditions is investigated by density-functional band-structure calculations. The influence of supercell size and shape on calculated electron energy-loss near-edge spectra is assessed quantitatively, employing the Z + 1 approximation for the representation of final-state effects. Relevant convergence criteria are the length scale set by the spatial extension of the valence electron screening cloud around the core hole and the interaction energy of neighbouring core-hole centres. A sufficient supercell size provided, the Z + 1 approximation yields a highly satisfactory description of excitations from the 1s shell of light elements, such as Mg and O, compared to experimental data. For comparison, pseudopotentials for excited states were generated for Mg, both with a large core (1s, 2s, 2p orbitals) and a small core (1s orbital only) included into the pseudopotential. The corresponding calculations with frozen core holes lead to very good agreement with the results from the Z + 1 calculation for the 1s excitations. The explicit treatment of the subvalence shell (2s, 2p), however, is mandatory for the proper modelling of excitations from orbitals higher than 1s. This indicates that the core polarisability plays an important role in excitations from more extended shells. PMID- 11281153 TI - Core-hole effect in the ELNES of alpha-Al2O3: experiment and theory. AB - The occurrence of the core-hole effect at the Al-K and Al-L1 edge in alpha-Al2O3 was studied by comparing experimental electron energy-loss near-edge structures (ELNES) and results of band-structure calculations with and without accounting for core-hole effects by the Z + 1 approximation. It was found that the theoretically calculated unoccupied p-like projected densities of states (PDOS) without Z + 1 approximation matches better to the experimental Al-L1 ELNES, whereas the PDOS with Z + 1 approximation matches better to the experimental Al-K ELNES. We conclude that the localisation of the initial state is an important prerequisite for the observability of the core-hole effect in the ELNES. PMID- 11281154 TI - Orientation dependence of ionization edges in EELS. AB - Anisotropy in the density of unoccupied states can be detected in the fine structure of ionization edges in angle-resolved EELS. It is shown that in a crystal an interference term occurs in the inelastic signal, and how it relates to electron channeling and site selection. The combination of orientation and site selection induces subtle variations in the ELNES. It is shown how this technique can be used to analyze local anisotropy related to the point group of the target atom. A second example shows how to extract non-dipole transitions at small scattering angles. PMID- 11281155 TI - Core-hole effects on energy-loss near-edge structure. AB - We present first-principles electron energy-loss near-edge structure calculations that incorporate electron-hole interactions and are in excellent agreement with experimental data obtained with X-ray absorption spectroscopy (XAS) and electron energy-loss spectroscopy (EELS). The superior energy resolution in XAS spectra and the new calculations make a compelling case that core-hole effects dominate core-excitation edges of the materials investigated: Si, SiO2, MgO, and SiC. These materials differ widely in the dielectric constant leading to the conclusion that core-hole effects dominate all core-electron excitation spectra in semiconductors and insulators. The implications of the importance of core holes for simulations of core-electron excitation spectra at interfaces will be discussed. PMID- 11281156 TI - Electron-energy-loss near edge structures of six-fold-coordinated Zn in MgO. AB - Electron-energy-loss near edge structures (ELNES) at the Zn-L(2,3) edge and the O K edge have been measured for 10 mol% ZnO-doped MgO, and were compared with spectra from reference materials. In order to interpret the spectra, first principles molecular orbital calculations were made using model clusters composed of 125 and 153 atoms. Photoabsorption cross sections (PACS) were computed at the Slater's transition state in which a half-filled core hole was included in the self-consistent calculations. The difference in the coordination numbers of Zn was found well distinguishable by the Zn-L(2,3)-edge ELNES. The experimental spectra in the first 25 eV were well reproduced by the theoretical PACS. In this energy region, the Zn-L(2,3)-edge ELNES from four-fold coordinated Zn showed four sets of peaks, whereas the six-fold coordinated Zn exhibits three sets of peaks. The origin of these peaks can be explained by the point symmetry within the first coordination unit. A small shift toward the lower energy side was observed in the O-K edge ELNES of the ZnO-doped MgO as compared with pure MgO. This can be ascribed to the lower energy of the Zn-4s orbital as compared with the Mg-3s orbital, which is the common mechanism to the difference in the band gap between MgO and ZnO. PMID- 11281157 TI - Simulation of thermal diffuse scattering including a detailed phonon dispersion curve. AB - Thermal vibration of the atoms in a crystal give rise to a diffuse background in the diffraction pattern (in between the normal allowed Bragg reflections). The Einstein model for phonon vibrations in a crystal leads to Gaussian statistics for the phonons. However, the Einstein model ignores the possibility of correlation between the atoms. An accurate model of the phonon dispersion curves for silicon is used to generate a set of more accurate random atomic displacements. These displacements are used in a multislice-style simulation to gauge the validity of the Einstein approximation. The phonon dispersion curve yields a small additional oscillatory structure in the thermal diffuse scattering (TDS) pattern. This does not produce significant changes in the annular dark field scanning transmission electron microscope (ADF-STEM) image signal, but could have a large impact on convergent beam measurements of bond charges. PMID- 11281158 TI - Treatment with parathyroid peptides and estrogen replacement for severe postmenopausal vertebral osteoporosis: prediction of long-term responses in spine and femur. AB - Fifteen women with severe vertebral osteoporosis were treated with daily parathyroid peptide (hPTH) plus hormone-replacement co-therapy (HRT) for 1 year. Eight other patients were randomized to HRT alone. Co-therapy with hPTH and HRT resulted in an impressive mean treatment response at the spine (dual-energy X-ray absorptiometry DXA) 15% above baseline; P < 0.015 compared with the HRT group) at 2 years, while at the proximal femur and radius there were smaller increases. hPTH co-therapy led to a significantly positive metabolic calcium balance at 1 year (by 2.13 mmol Ca/day, equivalent to a 5% annual increment in total body calcium; P = 0.015). The magnitude of the lumbar spine DXA response at 2 years depended statistically on the increase in bone formation rate, measured with 85Sr (r2 adjusted 0.48; P < 0.005) and patients with a large spine DXA response had larger calcium balance improvements (P < 0.03). Plasma osteocalcin changes tracked closely with increases in bone formation rate (r2 = 0.87). In seven patients treated throughout with HRT alone, and in eight hPTH-treated patients (three of whom switched to bisphosphonate therapy at year 4). DXA spine changes seen in years 3-5 were minimal, with no evidence of a statistically significant difference between groups. It is concluded that hPTH or comparable PTH receptor activators remain the most promising anabolic treatment for osteoporosis currently under clinical evaluation and a 6- or 12-month measurement of bone formation or a marker predicts the 2-5 year bone density outcome. Post-hPTH treatment, loss of bone appeared preventable with anti-resorptive therapy. PMID- 11281159 TI - Prevalence of vertebral fractures in a population-based sample in Japan. AB - In order to compare the prevalence of vertebral fractures in caucasian American and Japanese women, we examined the occurrence of vertebral fractures in a population of 1092 female residents of Yamanashi prefecture who participated in a screening program for osteoporosis. The relationship between vertebral fractures and bone mineral density (BMD) of the second metacarpal bone was also studied. Our findings regarding the prevalence of vertebral fractures in Yamanashi agreed strongly with the findings of previous reports in Japan. The prevalence of vertebral fractures was equal or lower among women in Japan compared with caucasians in Minnesota aged from 50 to 60 years. A comparison of the fracture and non-fracture groups revealed significant differences in age, height, and metacarpal BMD. However, after correction for age, no significant difference was found between the two groups. PMID- 11281160 TI - A possible role for leptin in normo- or hypoparathyroid uremic bone in postmenopausal dialysis women. AB - Leptin has been proposed to be a key molecule involved in energy regulation. Based on the generally acknowledged concept that a heavier person has a higher bone density, leptin is thought to be potentially involved in bone metabolism. Serum leptin, various bone markers, and bone mineral density (BMD) were studied in 51 dialysis patients (26 men and 25 women). The serum concentrations of leptin in dialysis patients ranged from 0.7 to 10.4 ng/ml (mean 3.2 +/- 2.1 ng/ml) for males and from 1.4 to 44.6 ng/ml (mean 11.8 +/- 10.4 ng/ml) for females. There was a good correlation between the body mass index (BMI) and leptin concentration in both male and female patients (P < 0.05). Serum leptin levels also correlated well with the age-adjusted z-score for BMD (P < 0.02) and were inversely correlated with levels of the carboxy-terminal propeptide of type I procollagen (P < 0.05) in females patients, but not in male patients. In conclusion, these results suggest an actual contribution of serum leptin in maintaining bone density in postmenopausal female dialysis patients. PMID- 11281161 TI - A longitudinal study for femoral neck bone mineral density increases in premenopausal caddies using dual-energy X-ray absorptiometry. AB - This longitudinal study examined whether bone mineral density (BMD) of the lumbar spine and proximal femur is maintained in premenopausal caddies (n = 6; mean age 37.8 years) in comparison with desk workers (n = 6; mean age 40.8 years) at the same golf club. BMD was followed for 12 months using dual-energy X-ray absorptiometry (DXA) and bone metabolic markers and athletic ability were also examined. Longitudinally, for caddies, the change per year in BMD of the lumbar spine was +0.009 g/cm2, while that of the proximal femur was +0.022 g/cm2, showing significant differences (P < 0.05 by signed-rank test). Their athletic ability, in terms of leg-press power, also significantly increased, whereas bone metabolic markers, such as serum alkaline phosphatase, 1,25-(OH), vitamin D3, parathyroid hormone and the deoxypyridiniline/creatinine ratio, did not show significant changes. For desk workers, the change per year in BMD of the lumbar spine was +0.011 g/cm2, while that of the proximal femur was -0.006 g/cm2. Their BMD, athletic ability and bone metabolic markers did not show significant changes. These findings support the results of our previous study, that premenopausal women can achieve continuous gain in femoral neck BMD by regular intense athletic activity, and suggest that this is also true by the continuous extensive walking of golf caddies. PMID- 11281162 TI - Raloxifene. AB - Raloxifene is a selective estrogen receptor modulator, a compound that has estrogen agonist activity at some sites and antagonist activity at others. In investigations in animals and in rigorously conducted trials in humans, raloxifene treatment is associated with a 30%-40% reduction in risk of one or more spine fractures using the 60 mg dose. This reduction in risk is found in women with or without baseline fractures, in women with bone mineral density (BMD) in the lower, middle, or upper third of the low range (all had BMD reduced by more than 2.5 SD) and in women aged less than 65 years, between 65-70 years, and greater than 70 years. A reduction in ankle fractures, but not hip or wrist fractures, was found. Raloxifene treatment also is associated with a 60%-70% reduction in risk for breast cancer and is associated with reduced total and LDL cholesterol, lower fibrinogen, and no rise in triglyceride. Reduced aortic wall cholesterol content is reported in animal studies. These are surrogate endpoints of cardioprotection. There is no evidence that raloxifene reduces the incidence of myocardial or cerebrovascular events. Raloxifene does not induce breast tenderness, endometrial hyperplasia, menstrual bleeding, or endometrial cancer, but may be associated with an increased risk of thromboembolic disease (1/1000 cases per year), leg cramps in 2%-4% of cases and hot flushes in 4%-6% of cases, usually in first 6 months. PMID- 11281163 TI - Mineral density and histomorphometric assessment of bone changes in the proximal tibia early after induction of type II collagen-induced arthritis in growing and mature rats. AB - Bone changes in both actively growing (6-week-old) and mature (6-month-old) rats with collagen-induced arthritis (CIA) were investigated in order to clarify the mechanisms of osteoporosis near inflamed joints in patients with early rheumatoid arthritis (RA) and juvenile RA. In female Sprague-Dawley rats, the proximal tibiae from the CIA and control groups early after immunization, when any influence of immobilization due to joint pain and swelling is minimal, were studied using dual X-ray absorptiometry and histomorphometry after double labeling with tetracycline. Arthritis developed within 10-14 days after immunization in both growing and mature rats. Physical activity, growth, and body weight continued to resemble that of the control group for at least 10 days. The bone mineral density in the proximal tibia did not differ significantly between the CIA and control groups. In growing rats, a highly significant increase in bone resorption, and decreases in bone formation and trabecular bone volume became evident histomorphometrically before visible signs of arthritis had developed. In mature rats, bone formation was markedly decreased without an increase in bone resorption. The differences in the reaction between growing and mature rats reflected a difference in the number of remodeling sites (units) and an uncoupling between osteoblasts and osteoclasts. We conclude that osteoporosis near inflamed joints results from an imbalance between bone resorption and formation caused by immune reactions in the CIA rats. Moreover, a decrease in bone formation may, in part, precede the clinical onset of arthritis. PMID- 11281164 TI - Effect of 1alpha-hydroxyvitamin D3 and egg-shell calcium on bone metabolism in ovariectomized osteoporotic model rats. AB - Egg-shell calcium (Ca) is one of the effective Ca sources for bone metabolism. In the present study, we investigated whether egg-shell Ca had similar effects compared with calcium carbonate (CaCO3) when vitamin D3 (1alpha(OH)D3) treatment was given to an osteoporotic rat model. In both 1alpha(OH)D3-supplemented and unsupplemented rats, the bone mineral density (BMD) of the lumber spine in the vitamin-supplemented group increased significantly compared with the unsupplemented group. In a Ca balance study, there were also significant differences in intestinal Ca absorption, urinary Ca and fecal Ca between the vitamin-supplemented and -unsupplemented groups. These results show that egg shell Ca could have similar effects to CaCO3 on bone metabolism. In contrast with CaCO3, vitamin D3 supplementation did not significantly increase serum Ca levels in the egg-shell Ca group; however, the mechanism of Ca absorption is still unclear. Our results suggest that egg-shell Ca may be an effective nutrient in Ca metabolism for people treated with vitamin D3. PMID- 11281165 TI - Involvement of interleukin-6 and prostaglandin E2 in periarticular osteoporosis of postmenopausal women with rheumatoid arthritis. AB - In experimental arthritis, blocking of the receptor activator of nuclear factor kappaB ligand (RANKL) by osteoprotegerin (OPG) treatment prevents bone loss but not inflammation, suggesting that there are inflammation-related factors that regulate RANKL and OPG. However, it is not known which factors regulate RANKL and OPG in human inflammation-induced bone loss. To clarify the inflammation-related factors that play a role in periarticular osteoporosis in patients with rheumatoid arthritis (RA), the synovial fluid and synovium of the knee joint, and the periarticular cancellous bone of the femoral condyle were collected at surgery from postmenopausal women with RA or osteoarthritis (OA). All patients with RA had radiologic bone loss on the femoral condyles, while such a loss was not observed in patients with OA. The present study examined: (i) tumor necrosis factor alpha (TNFalpha), interleukin (IL)-1beta and IL-6 levels in synovial fluid: (ii) TNFalpha, IL-1beta and IL-6 messenger RNA (mRNA) expression in the synovium and the cancellous bone that contained bone marrow; and (iii) IL-6 and prostaglandin E2 (PGE2) production in cultured osteoblast-lineage cells derived from collagenase-treated cancellous bone fragments. Inflammation of the knee joints in patients with RA was confirmed by significantly higher proinflammatory cytokine levels in the synovial fluid and the synovium than those seen in patients with OA. In patients with RA, mRNA expression of IL-6, but not TNFalpha and IL-1beta, in the cancellous bone and IL-6 and PGE2 production in the osteoblast-lineage cells were significantly higher than in patients with OA. These findings suggest, for the first time, that IL-6 is involved in periarticular osteoporosis in postmenopausal women with RA. IL-6 and PGE2 released from osteoblast-lineage cells could be, at least in part, responsible for human inflammation-induced bone loss. PMID- 11281166 TI - Effects of sodium bicarbonate supplementation on axial and peripheral bone mass in rats on strenuous treadmill training exercise. AB - We observed the effects of sodium bicarbonate supplement on bone mass in rats on strenuous treadmill training. Sixty female Wistar rats (93-days-old; mean initial weight 261 +/- 16 g) were studied. One group of 15 rats was killed at the beginning of the experiments (basal control group), while another group of 15 rats was not manipulated (Exer-NaB-). Another group of 15 rats was exercised but did not receive sodium bicarbonate (Exer+NaB-), while the final group of 15 rats exercised and received sodium bicarbonate (Exer+NaB+) at a dose of 0.05 mg/kg/day, administered by esophageal catheter on exercise days. These rats were killed at the end of 11 weeks. Femoral and vertebral length, weight, and bone mineral content (BMC) and density (BMD) were measured. According to ANOVA with the Tukey-Kramer test, femur length and weight, vertebral weight, femur BMC and BMD, vertebral BMC and BMD and the ratio between femur and vertebral BMC and final body weight, and plasma bicarbonate were lower in the basal control and Exer+NaB- groups than in the two other groups (P < 0.005-0.0001). Overall, there was a positive correlation between femur and vertebral BMC and femur BMC and length (P < 0.0001 for all). Only in the Exer+NaB- group was there a positive association between plasma bicarbonate levels and femur length (r = 0.78; P < 0.0005). Our study demonstrates the adverse effects of strenuous exercise on bone, and the usefulness of sodium bicarbonate supplements in preventing and minimized these effects. PMID- 11281167 TI - Slowing of gastrointestinal transit by oleic acid: a preliminary report of a novel, nutrient-based treatment in humans. AB - Chronic diarrhea may occur when gastrointestinal transit is abnormally rapid. We hypothesized that oleic acid given prior to a meal would slow gastrointestinal transit and reduce diarrhea by activating nutrient-triggered inhibitory feedback mechanisms in the small intestine. Transit time was measured in eight normal subjects following ingestion of a control emulsion (0 ml oleic acid), and in 45 patients with chronic diarrhea following ingestion of emulsions containing 0, 1.6, and 3.2 ml oleic acid. Stool volume and frequency on and off treatment were compared. Transit time in normal subjects was 102.4 +/- 11.2 min (mean +/- SE). Transit times in patients was shorter at 29.3 +/- 2.8 min with the 0-ml dose (P < 0.001), but increased to 57.2 +/- 4.5 min with the 1.6-ml dose and to 83.3 +/- 5.2 min with the 3.2-ml dose (P < 0.001). In the 18 patients who provided stool records, frequency of bowel movements decreased from 6.9 +/- 0.8 to 5.4 +/- 0.9 bowel movements/24 hr (P < 0.05) and stool volume decreased from 1829.0 +/- 368.6 to 1322.5 +/- 256.9 ml/24 hr with treatment (P < 0.05). An emulsion containing oleic acid slowed gastrointestinal transit and reduced diarrhea by activating nutrient-triggered inhibitory feedback mechanisms in the small intestine. PMID- 11281168 TI - Impaired gut contractility following hemorrhagic shock is accompaied by IL-6 and G-CSF production and neutrophil infiltration. AB - Recovery from hemorrhagic shock (HS) is frequently accompanied by bowel stasis. The aim of this study was to examine whether or not HS initiates an inflammatory response that includes production of cytokines, specifically G-CSF and interleukin-6 (IL-6), and recruitment of leukocytes within the intestinal muscularis which contribute to impaired muscle contractility. Sprague-Dawley rats were subjected to HS (MAP 40 mm Hg for 156 min) followed by resuscitation, and then they were killed at 4 hr. Shock animals demonstrated accumulation of PMNs in the jejunal muscularis and decreased spontaneous and bethanechol-stimulated muscle contractility. Semiquantitative RT-PCR demonstrated elevated levels of IL 6 and G-CSF mRNA in shock animals in full-thickness jejunum and in mucosa and muscularis layers compared to sham controls. Immunostaining demonstrated increased IL-6 protein production within the muscularis externa and submucosa. In situ hybridization studies localized G-CSF mRNA production to the submucosa. Gel shift assays revealed increased NF-kappaB and Stat3 activity in full-thickness jejunum and jejunal layers of shock animals. Activation of Stat3 also was demonstrated in normal muscularis tissue exposed to IL-6 and G-CSF in vitro. IL-6 and G-CSF are produced in the muscularis and mucosa layers of the gut in HS where they may contribute to PMN recruitment and smooth muscle dysfunction. PMID- 11281169 TI - Impaired postprandial gastric myoelectrical activity in Chinese patients with nonulcer dyspepsia. AB - Using a homemade electrogastrography (EGG) system, we studied the characteristics of the myoelectrical rhythm in Chinese patients with nonulcer dyspepsia (NUD). Based on short-term Fourier transformation, recorded slow waves could be automatically analyzed to obtain the following parameters: dominant frequency/power, percent of normal frequency (2-4 cpm), power ratio, etc. EGG parameters, Helicobacter pylori status, histological examination of gastric mucosa, and dyspeptic symptoms were recorded in 27 NUD patients. Compared to 32 healthy controls, the Chinese NUD patients had abnormal postprandial EGG parameters including a lower percentage of regular 2-4 cpm slow waves (70.10 +/- 2.97% vs 79.08 +/- 2.95%, P < 0.05), a lower level of increment of dominant power (0.62, +/- 0.91 vs 3.76 +/- 0.58 dB, P < 0.05), lower power ratio (1.42 +/- 0.28 vs 2.79 +/- 0.39, P < 0.05) and a higher instability coefficient (0.36 +/- 0.03 vs 0.26 +/- 0.03, P < 0.05). However, Helicobacter pylori infection and its associated gastritis did not influence any EGG parameters in NUD patients. Six main dyspeptic symptoms and total symptom score had no correlation with any EGG parameters. In conclusion, Chinese NUD patients may have abnormal postprandial stomach myoelectrical activity, but these EGG abnormalities are not a direct result of Helicobacter pylori infection and its related gastritis and do not contribution to the dyspeptic symptoms. PMID- 11281171 TI - Disturbance of purine nucleotide metabolism: a possible early key event in development of intestinal damage induced by chemotherapy. AB - Protective strategies to minimize the hematological toxicity in connection with bone marrow transplantation (BMT) have been successful, but toxicity to the gastrointestinal tract prevents further dose escalation and therefore limits the application of the treatment. As it is known that chemotherapy leads to disruption of the intestinal barrier and morphological changes of mitochondria in enterocytes, this study was conducted in order to investigate intestinal energy metabolism and permeability after intensive cytotoxic therapy in rats. Intestinal damage was produced by intraperitoneal administration of the cytostatic etoposide. Intestinal permeability was assessed by a [51Cr]EDTA absorption test and intestinal purine nucleotide content by a high-performance liquid chromatography (HPLC) technique. Four hours after the administration of etoposide, and the next 48 hr, there was a significant increase in the intestinal permeability (P < 0.05) and a significant reduction of the purine nucleotide content in the intestinal epithelial cells (P < 0.01) as compared to control animals. This early disturbance in enterocyte energy metabolism may be a key event in the development of the intestinal damage, induced by chemotherapy, and an explanation for the early disruption of the intestinal barrier demonstrable before morphological changes are evident. PMID- 11281170 TI - Mechanisms for contractile effect of Dai-kenchu-to in isolated guinea pig ileum. AB - The mechanisms by which Dai-kenchu-to (TJ-100), a kampo medicine, enhances gastrointestinal motility was investigated using isolated guinea pig ileum. TJ 100 induced contractions accompanied by autonomous contraction at a concentration of more than 3 x 10(-4) g/ml in a dose-related manner. The TJ-100-induced ileal contraction was suppressed by atropine and tetrodotoxin, but not by hexamethonium. This effect was partially suppressed in the presence of high concentrations of ICS 205-930, a serotonin 4 (5-HT4) receptor antagonist. In addition, TJ-100 showed an acetylcholine (ACh)-releasing action in the smooth muscle tissues of ileum. These results suggest that contractile response induced by TJ-100 is partially mediated by ACh released from the cholinergic nerve endings and that 5-HT4 receptors would be involved in the effect of TJ-100. PMID- 11281172 TI - Expression of melanoma antigen-encoding gene-1 predicts lymph node involvement in early gastric carcinomas. AB - Our previous study suggested that melanoma antigen-encoding gene-1 may be activated in gastric carcinomas when the tumors develop or invade. In the present work we studied the gene as a preoperative indicator of lymph node involvement in early gastric carcinoma. We used a reverse transcription-polymerase chain reaction to analyze expression of the gene in 47 endoscopic biopsy specimens from early gastric carcinomas. Relationships between gene expression and histopathologic findings were analyzed statistically. Eight of 47 tumor samples (17.0%) expressed the gene. The gene was not expressed in adjacent nontumor samples from carcinoma patients or in tissues from patients with benign gastric diseases. Gene expression correlated with infiltrative tumor growth in contrast to tumors with expansive growth patterns (P = 0.018). Gene expression also correlated with expression of platelet-derived growth factor-A (P < 0.001). MAGE 1 gene expression in biopsy specimens was a preoperative indicator of nodal involvement (P = 0.013). In conclusion, melanoma antigen-encoding gene-1 expression in biopsy specimens from early gastric carcinoma may serve preoperatively as an indicator of lymph node involvement. PMID- 11281173 TI - Adjuvant radiotherapy for gastrointestinal stromal tumor of the rectum. PMID- 11281174 TI - Anal squamous cell carcinoma in a patient with Peutz-Jeghers syndrome. PMID- 11281175 TI - Hybrid classification of sphincter of Oddi dysfunction based on simplified Milwaukee criteria: effect of marginal serum liver and pancreas test elevations. AB - To date, when using the Milwaukee classification for sphincter of Oddi dysfunction (SOD), one cannot accurately classify patients with marginal elevations in laboratory tests; ie, < 1.5 x the upper limit of normal (ULN). Since subsequent treatment may depend on how they are classified, we sought to determine whether these patients should be considered as type II or type III. Between January 1993 and October 1996, 113 consecutive patients (82 females and 31 males; ages 12-87 years) without prior sphincterotomy were referred to consider a diagnosis of SOD type II or III. SOD II patients had pancreaticobiliary-type pain and laboratory elevations >1.5 x ULN or dilated ducts, while SOD III patients had pain only. Hybrid patients had pain and marginal laboratory elevations <1.5 x ULN, with normal duct diameters. Drainage times, frequency, duration, and propagation were not assessed. Sphincter of Oddi manometry (SOM) was performed in each case, and the frequency of abnormal biliary and/or pancreatic basal sphincter pressure was compared, with respect to type II, III, and hybrid SOD. Successful SOM was obtained in 113/114 patients: Abnormal basal sphincter pressure was found in 65, 89, and 43% of type II, hybrid, and type III SOD, respectively. We found no statistical difference between type II and hybrid patients. In contrast, there was statistical difference between types II and III patients and between type III and hybrid patients. In conclusion, there was no significant difference in the frequency of elevated basal sphincter pressure in SOD type II versus hybrid, and thus they should be considered as one group. PMID- 11281176 TI - Which is a less invasive pancreatic head resection: PD, PPPD, or DPPHR? AB - Less invasive pancreatic head resection, such as pylorus preserving pancreatoduodenectomy (PPPD) and duodenum preserving pancreatic head resection (DPPHR) has been introduced for the treatment of pancreatoduodenal lesions, especially for benign conditions, in consideration of postoperative quality of life. Surgical stress and exocrine and endocrine function of the residual pancreas were examined in 44 patients with PPPD, 10 with conventional pancreatoduodenectomy (PD) and six with DPPHR. Clinical findings including serum levels of C reactive protein (CRP), fasting blood sugar, a 120-min value of the 75-g oral glucose tolerance test (OGTT), N-benzol-L-tyrosyl-p-aminobenzoic acid (BT-PABA) excretion value (a pancreatic exocrine function test), and volume of postoperative pancreatic juice drainage were compared among the three different variants of pancreatectomy. Operation time and operative blood loss in PD were largest of the three, followed by PPPD and DPPHR. Postoperative elevation of serum CRP on postoperative day (POD) 2 or 3 was similar among the three different types of operation. Fasting blood sugar concentrations were not different among the three groups at short- and long-term after the operation, while the 120-min value of the GTT showed a marked elevation at long-term only after PPPD. The volume of pancreatic juice drainage increased up to POD 4 and became constant thereafter. The total amount of pancreatic juice drainage from POD 4 to 13 was smallest in PD (637 ml) followed by PPPD (1,255 ml) and DPPHR (1,431 ml). The BT PABA value declined after PD (-20.3%, P = 0.0437) and PPPD (-20.2%, P = 0.0239) at short term, but not after DPPHR (8.2%). These findings suggest that the early impairment of the pancreatic exocrine function after PD and PPPD but not after DPPHR may indicate that the invasiveness of pancreatic head resection to the pancreatic functions is greater in PD and PPPD than in DPPHR. PMID- 11281178 TI - Gallbladder vasculitis associated with type-1 cryoglobulinemia. AB - A patient with type I cryoglobulinemia and monoclonal gammopathy of uncertain significance was found to have acute gallbladder vasculitis. The most prominent manifestation was upper abdominal pain in the setting of normal liver tests. An abdominal ultrasound demonstrated a thickened gallbladder wall, along with gallstones. HIDA scanning showed a nonfunctioning gallbladder with an edematous and thickened wall. There was characteristic leukocytoclastic vasculitis affecting the gallbladder. The patient recovered uneventfully subsequent to cholecystectomy. Gallbladder vasculitis should be considered in patients with unexplained upper abdominal pain and systemic vasculitis. PMID- 11281177 TI - Role of prostaglandins in regulation of pancreatic enzyme secretion by various diets. AB - We have demonstrated by the use of isolated rat pancreatic acini that exogenous prostaglandins of the E type inhibit secretagogue-stimulated amylase secretion. We here studied whether the pancreas is a source of prostaglandin synthesis and whether prostaglandins mediate regulation of pancreatic enzyme secretion by various diets. Prostaglandin E2 was measured by enzyme immunoassay in pancreatic acini from either normal animals or after 10 days of feeding with different diets. Acini were prepared by collagenase digestion. Amylase secretion was measured after stimulation with cholecystokinin in the presence or absence of indomethacin, an inhibitor of prostaglandin synthesis. Prostaglandin E2 concentration in pancreatic acini was comparable to other organs such as kidney and liver. Feeding a diet enriched in proteins caused an increase of cholecystokinin-stimulated maximal amylase secretion and a decrease of prostaglandin E2 concentration. Incubation of acini with indomethacin caused a decrease in prostaglandin E2 concentration and an increase in cholecystokinin stimulated amylase secretion. We conclude that regulation of pancreatic enzyme secretion by diets may be mediated by prostaglandins. PMID- 11281179 TI - Attributable risk of H. pylori in peptic ulcer disease: does declining prevalence of infection in general population explain increasing frequency of non-H. pylori ulcers? AB - Recent reports in the United States have found that fewer peptic ulcers are due to Helicobacter pylori than previously believed. The aim of this study is to determine if the declining prevalence of H. pylori infection in the general population can account for the apparent increase in the frequency of non-H. pylori ulcers. A total of 396 patients with peptic ulcer or ulcer scar were enrolled in this study. The pre-1950 population consisted of 149 patients with gastric ulcers and with 44 duodenal ulcers. The post-1950 population consisted of 96 patients with gastric ulcers and 107 with duodenal ulcers. The frequency of H. pylori-negative gastric ulcers was 5.4% in patients born before 1950 and 4.2% in patients born after 1950, and the frequency of H. pylori-negative duodenal ulcers was 0% and 1.9%, respectively. There are no statistical differences between the two populations in gastric and duodenal ulcers. H. pylori seropositivity was 74.9% in asymptomatic volunteers born before 1950 and 20.7% in those born after 1950 (P < 0.01) in the general population. The attributable risk of H. pylori infection in peptic ulcer diseases was not affected by the prevalence of H. pylori infection in the general population in Japan. This suggests that the apparent increase in frequency of non-H. pylori ulcers in the United States is not simply due to the declining prevalence of infection. Other explanations for non-H. pylori ulcers should be sought. PMID- 11281180 TI - Expression of gene for EIIIA- and EIIIB- fibronectin, fetal types of fibronectin, during gastric ulcer healing in rats. AB - Fibronectin (FN) is important for wound healing via cell proliferation, adhesion, differentiation, and matrix formation. Fetal types of FN mRNA, which include the region of EIIIA or EIIIB, are well expressed during embryonic development and wound healing. This study was done to investigate the mRNA expression of full length FN, EIIIA- and EIIIB-FN, and the localization of FN in the gastric tissues during ulcer healing with northern blot analysis and immunohistochemical technique. Gastric ulcers in rats were produced by acetic acid. EIIIA- and EIIIB FN mRNA were not detected in normal gastric tissues, but were expressed in the ulcerated tissues throughout the healing phase. However, on day 60 (in the scarred phase), the EIIIA- and EIIIB-FN mRNA had disappeared. The levels of full length FN mRNA were increased from day 3 to 32 compared with the control levels, and decreased to the control levels on day 60. Full-length FN was predominantly localized at the mesenchyme around the infiltrating inflammatory cells in the granulation tissues and the basement membranes of the nonproliferating epithelial cells, which were regenerated at the ulcer margin. Thus, fetal gene transcripts of FN suggest the important role of fetal FN in gastric ulcer healing, mainly via the migration of mesenchymal and epithelial cells. PMID- 11281181 TI - Protective effect of famotidine, omeprazole, and melatonin against acetylsalicylic acid-induced gastric damage in rats. AB - It has been reported that both omeprazole and famotidine have a protective effect against gastric mucosal damage induced by acetylsalicylic acid (ASA) and other nonsteroidal anti-inflammatory drugs. Since active oxygen species and lipid peroxidation were reported to play a role in the pathogenesis induced by ASA, we aimed to study if omeprazole and famotidine have any antioxidant effect by comparing their protective effect with that of melatonin, an effective antioxidant and free radical scavenger. Mucosal damage was evaluated by macroscopic examination, histological analysis and by measurement of lipid peroxidation (LPO), glutathione (GSH), and myeloperoxidase (MPO) activity. Omeprazole (20 micromol/kg per os), famotidine (3 mg/kg per os), and melatonin (10 mg/kg intraperitoneally) significantly prevented gastric ulcerogenesis induced by ASA (200 mg/kg per os) and decreased the ulcer index. Gastric LPO and MPO activity that were increased significantly by ASA were decreased after treatment with omeprazole, famotidine, and melatonin. ASA treatment decreased significantly the gastric GSH levels, and pretreatment with omeprazole, famotidine, or melatonin increased it significantly. Famotidine and omeprazole decreased the gastric acidity, which was increased by ASA, whereas melatonin had no effect on this parameter. These findings suggest that active oxygene species and LPO have an important role in the pathogenesis of gastric mucosal damage induced by ASA and that both famotidine and omeprazole may be protective against this damage, although they were not as efficient as melatonin as an antioxidant. On the other hand, the antisecretory effect of omeprazole and famotidine may also be contributing to their antiulcer effect. PMID- 11281182 TI - 1,8-cineol, a food flavoring agent, prevents ethanol-induced gastric injury in rats. AB - This study investigated the gastroptrotective effect of 1,8-cineole (cineole) on ethanol-induced gastric mucosal damage in rats and the possible mechanisms involved. 1,8-Cineole (50-200 mg/kg), given orally 1 hr before administration of 1 ml of absolute ethanol significantly attenuated the ethanol-induced gastric injury in a manner similar to nordihydroguairetic acid, a known lipoxygenase inhibitor. 1,8-Cineole showed a tendency to restore the ethanol-associated decreases in nonprotein sulfhydryls, suggesting a possible antioxidant effect. In gastric secretion studies, 1,8-cineole, similar to cimetidine, a known histamine 2 receptor antagonist, demonstrated significant inhibitions of both gastric juice volume as well as total acid output. The protection offered by 1,8-cineole was found to be unaltered by 8-phenyltheophylline or L-NAME, indicating that its effect is not mediated by endogenous adenosine or nitric oxide. These results, taken together with the earlier reports, suggest that the antioxidant and lipoxygenase inhibitory actions of 1,8-cineole are of prime importance in affording gastroprotection against ethanol injury in the rat. PMID- 11281183 TI - Diclofenac-induced gastric mucosal fluorescence in rats. AB - We previously reported that the gastric mucosa emits fluorescence of porphyrins at the onset of gastric lesions induced by hemorrhagic shock. In this study, we investigated whether the fluorescent substance concerns with the gastric mucosal injuries induced by diflofenac, a nonsteroidal antiinflammatory drug (NSAID). In the gastric mucosa treated with diclofenac, lesions were generated and myeloperoxidase activity increased. Diclofenac administration also increased thiobarbituric acid-reactive substances, a index of tissue peroxidation. After diclofenac treatment, the gastric mucosal fluorescence intensities rose. HPLC analysis demonstrated that the fluorescent substances were mesoporphyrin and protoporphyrin, which were the same as found in hemorrhagic shock. Pretreatment of the tissue with radical scavenging substances, catalase and troxipide, restrained the increase of mucosal fluorescence intensity, tissue peroxidation, and lesion formation. These findings indicate that diclofenac treatment induced the generation of porphyrins as well as tissue peroxidation in gastric mucosal tissue. This study suggests that autofluorescence observation is a useful tool to identify diclofenac-induced gastric injury. PMID- 11281184 TI - Similar T-cell oligoclonality in antimitochondrial antibody-positive and negative primary biliary cirrhosis. AB - Approximately 5% of patients with clinical and histological features suggestive of primary biliary cirrhosis do not have anti-mitochondrial antibodies that can be detected by current methodologies. Although the role of these autoantibodies in the pathogenesis of liver disease is uncertain, T lymphocytes within the portal tracts are felt to be important mediators of bile duct destruction. In order to investigate the hypothesis that a similar T-cell process may be involved in both antimitochondrial antibody-positive and -negative primary biliary cirrhosis, we characterized the oligoclonally expanded T cells in both types of patients by analysis of complementarity determining region 3 length in peripheral blood mononuclear cells. The distribution of oligoclonally expanded T cells was similar in both groups. This finding does not support a distinct T-cell-mediated pathogenesis for anti-mitochondrial antibody-positive and -negative primary biliary cirrhosis but rather suggests that similar processes may be involved in the immunopathogenesis of both. PMID- 11281185 TI - Antipyrine clearance and metabolite formation in primary biliary cirrhosis. AB - The disposition of antipyrine is altered and may be a prognostic factor in the presence of various types of hepatic dysfunction. The object of the present study was to investigate whether antipyrine clearance and metabolite formation are useful to detect altered metabolic function in primary biliary cirrhosis. Saliva clearance of antipyrine and the formation clearance of antipyrine metabolites (hydroxymethylantipyrine, HMA; norantipyrine, NORA; and 4-hydroxyantipyrine, OHA) were investigated in a group of 34 women with biopsy-proven PBC (mean age 60 years; range 39-87 years) and in 15 healthy control women (mean age 62 years; range 46-78 years). Parameters of antipyrine clearance of patients in stage I and II were similar to those observed in healthy subjects. When compared to patients in stage I, patients in advanced stages showed a reduction in antipyrine clearance (-29% and -44% in stages III and IV, respectively) and increases in antipyrine half-life (+24% and +75% in stages III and IV, respectively). The reduction in antipyrine clearance was due to a reduction in the formation of all three antipyrine metabolites, with the formation clearance of both HMA and NORA decreasing to a slightly greater extent than that of OHA. Antipyrine clearance correlated significantly with serum bilirubin (P < 0.017) and the Mayo risk score (P < 0.001). Logistic regression analysis indicated that antipyrine clearance was an independent predictor of the histological stage of the disease (P < 0.001). Antipyrine clearance and metabolite formation is a sensitive parameter for assessing hepatic metabolic function in primary biliary cirrhosis. PMID- 11281187 TI - Primary calcified gastrinoma of the liver. PMID- 11281186 TI - Effects of ethanol administration on hepatocellular ultrastructure of regenerating liver induced by partial hepatectomy. AB - Acute ethanol administration partially inhibits DNA and protein syntheses during liver regeneration (LR) induced by partial hepatectomy (PH) in rats. Previous findings that the magnitude of ethanol's deleterious effects on LR are related to the route and timing of its administration led us to perform studies at the ultrastructural level, comparing ethanol effects on PH-induced LR, as a consequence of its administration route. PH promoted alterations on the endoplasmic reticulum and mitochondria, accompanied by decreased glycogen and increased lipid content in cytoplasm. Structural nuclear and nucleolar activities were also evident. Intragastric ethanol administration practically abolished the adaptative changes found in PH-promoted regenerating hepatocytes, whereas its administration through the intraperitoneal route induced later ultrastructural modifications, indicating cellular proliferation. These results suggest that ethanol, under certain conditions, could stimulate liver proliferation triggered by PH. The mechanism underlying this surprising effect of ethanol on LR remains to be elucidated. However, it is suggested that an altered ethanol metabolism by rats subjected to PH could be involved. PMID- 11281188 TI - Myositis, microvesicular hepatitis, and progression to cirrhosis from troglitazone added to simvastatin. AB - A 68-year-old woman, with type 2 diabetes mellitus, hypercholesterolemia, and prior long-term simvastatin therapy, self-resumed troglitazone after running out of metformin. She developed an acute severe hepatitis with microvesicular steatosis and mysositis. There was subsequent resolution of the myositis but progression of the hepatitis to symptomatic cirrhosis over a period of 12 weeks. Both troglitazone and simvastatin are metabolized by cytochrome P-450 3A4. Troglitazone typically induces metabolism of drugs metabolized by this cytochrome so that simple simvastatin toxicity seems less likely to have been involved. The association with myositis, the severity of the hepatitis with progression to cirrhosis, and the presence of microvesicular steatosis suggests altered mitochondrial metabolism, which has been described with each agent, as the underlying pathogenic mechanism. Although troglitazone (Rezulin) has been withdrawn from the market, other similar agents are available for therapy of type 2 diabetes mellitus. Increased awareness of a potential interaction between these two classes of drugs is warranted. PMID- 11281189 TI - Intestinal growth adaptation and glucagon-like peptide 2 in rats with ileal- jejunal transposition or small bowel resection. AB - Glucagon-like peptide 2 (GLP-2), produced by enteroendocrine L-cells, regulates intestinal growth. This study investigates circulating and intestinal GLP-2 levels in conditions with altered L-cell exposure to nutrients. Rats were allocated to the following experimental groups: ileal-jejunal transposition, resection of the proximal or distal half of the small intestine, and appropriate sham-operated controls. After two weeks, ileal-jejunal transposition led to pronounced growth of the transposed segment and also of the remaining intestinal segments. Plasma GLP-2 levels increased twofold, whereas GLP-2 levels in the intestinal segments were unchanged. In resected rats with reduced intestinal capacity, adaptive small bowel growth was more pronounced following proximal resection than distal small bowel resection. Circulating GLP-2 levels increased threefold in proximally resected animals, and twofold in the distally resected group. Tissue GLP-2 levels were unchanged in resected rats. The data indicate that transposition of a distal part of the small intestine, and thereby exposure of L cells to a more nutrient-rich chyme, leads to intestinal growth. The adaptive intestinal growth is associated with increased plasma levels of GLP-2, and GLP-2 seems to act in an endocrine as well as a paracrine manner. PMID- 11281191 TI - Gut mucosal secretion of interleukin 1beta and interleukin-8 predicts relapse in clinically inactive Crohn's disease. AB - Trials of maintenance therapy in Crohn's disease are often underpowered, and there is need for objective markers of relapse. We assessed the relationship of whole gut lavage fluid cytokines to relapse in inactive Crohn's disease. Fifty four patients with inactive Crohn's disease were prospectively assessed. Inactivity was determined as a Crohn's disease activity index of <150 and whole gut lavage fluid immunoglobulin G <10 microg/ml. All patients underwent whole gut lavage with analysis of IL-1beta and IL-8. Follow up was for one year. Patients with elevated whole gut lavage fluid IL-1beta (P < 0.004) and IL-8 (P < 0.02) had greater chance of relapse. Young age, short disease duration, and fistulating disease also relapsed more frequently. Multiple regression identified IL-1beta as an independent variable. In conclusion, an elevated whole gut lavage fluid IL 1beta in inactive Crohn's disease identifies patients at high risk of relapse. PMID- 11281190 TI - Slow transit constipation: a disorder of pelvic autonomic nerves? AB - Slow transit constipation (STC) is a severe motility disorder, which in the majority of cases is of unknown etiology. In some, symptoms arise de novo in childhood, but a proportion of patients present in later life, including after pelvic surgery or childbirth. Our aims were: (1) to describe our current knowledge of the anatomy and function of the pelvic autonomic nerves with respect to colonic motility (experimental and observational studies); (2) to discuss evidence for pelvic nerve injury in STC arising after pelvic surgery or childbirth; and (3), on the basis that such patients are clinically indistinguishable from patients with chronic idiopathic STC, to evaluate whether there is evidence that pelvic autonomic neuropathy has an etiologic role in patients with chronic idiopathic STC. The outcome was as follows: (1) The clear importance of the pelvic autonomic nerves in colonic motor function is documented. (2) While there is an association between pelvic surgery and childbirth, and the onset of STC, there is little direct anatomical evidence that pelvic denervation occurs in these patients. However the phenotype of these patients is similar to results of experimental and observational studies. (3) Clinical, physiological, and histological similarities exist between patients whose symptoms arose following pelvic intervention and those whose symptoms arise de novo (idiopathic). We further present evidence for possible pathogenetic mechanisms underlying pelvic autonomic neuropathy in chronic idiopathic STC. PMID- 11281192 TI - Multiple accumulation of Vdelta2+ gammadelta T-cell clonotypes in intestinal mucosa from patients with Crohn's disease. AB - The gammadeltaT cells have been known to play an important role in the regulation of the mucosal immune system, but the relationship between these cells and the pathogenesis of Crohn's disease (CD) has remained obscure. We now demonstrate the T-cell receptor (TCR) Vdelta2 gene transcripts characterize antigenic immune response in the intestinal mucosa from patients with CD. TCR Vdelta2 gene transcripts of six patients with CD and six controls were subcloned and 20 randomly selected clones from each sample were subjected to nucleotide sequencing. Sequence analysis demonstrated that the different clones in the intestinal mucosa of CD were significantly increased (11.833 +/- 0.946) compared to controls (7.167 +/- 1.470) (P = 0.0374). The presence of intraindividual dominance and oligoclonality of TCR Vdelta2 gene transcripts in normal controls appears reflect positive selection and expansion of specific gammadelta T cells in normal controls. By contrast TCR Vdelta2 gene transcripts in the intestinal mucosa of CD can express different clonotypes. We conclude that accumulation of multiple Vdelta2+ gammadelta T-cell clonotypes are involved in the pathogenesis of CD. PMID- 11281193 TI - Alteration of culture regime modifies antioxidant defenses independent of intracellular reactive oxygen levels and resistance to severe oxidative stress within confluent Caco-2 "intestinal cells". AB - Reactive oxygen species are implicated in the development of gastrointestinal pathologies. Caco-2 monolayers are routinely used to study intestinal oxidative stress and its potential amelioration by pharmacological agents or dietary micronutrients. Little is known of the plasticity of Caco-2 antioxidant defenses with changes in culture conditions. We examined whether the frequency of culture media renewal alters the antioxidant-prooxidant status and integrity of Caco-2 monolayers. In comparison to monolayers subject to daily media renewal, increasing periods between media exchange resulted in varying degrees of suppression of catalase, glutathione reductase, and glutathione-S-transferase activity. No significant changes to superoxide dismutase activity, total glutathione, or intracellular ROS profiles were observed. Alkaline phosphatase activity, as a marker of differentiation, and mean monolayer cell population size were also unaffected. We suggest that Caco-2 antioxidant enzyme activities are differentially sensitive to changes in culture conditions. Studies employing this cell line for antioxidant-oxidative stress interactions will need to evaluate responses with respect to culture regime. PMID- 11281194 TI - Immunohistochemical analysis of distribution of RON receptor tyrosine kinase in human digestive organs. AB - The immunohistochemical distribution of RON receptor tyrosine kinase in digestive organs of both human fetus and adult, including the esophagus, stomach, duodenum, small intestine, colon, rectum, liver, gallbladder, pancreas, and spleen, was investigated semiquantitively using an affinity-purified rabbit polyclonal antibody. RON was observed to be widely distributed throughout various digestive organs and cell types in humans. The immunoreactivity for RON was observed in the epithelium of the esophagus, small intestine, colon, hepatocytes, Kupffer cells, and splenic macrophages both in the adult and the fetus, suggesting that the MSP/RON signaling pathway possesses the proper biological properties to possibly be involved in morphogenesis or differentiation of cells in these organs and cell types. Several organs differed in immunoreactivity between adult and fetus. No immunoreactive cells were found in the pancreas of adults; however, immunoreactivity was observed in acinar cells and in some of the duct or ductular cells and endocrine cells of the islet of the fetus. Similarly, immunoreactivity was not observed in gastric mucosa except in the intestinal metaplastic cells in adults; however, immunoreactivity was found in the foveolar epithelium of the stomach of the fetus. Although the biological significance of RON in malignancy is unclear, the presence of RON immunoreactivity in the fetus and it lack in the adult may indicate that RON is a oncofetal substance in human pancreas and stomach. PMID- 11281195 TI - Prevention of deleterious effects of reperfusion injury using one-week high-dose allopurinol. AB - Allopurinol has been widely used to reduce the severity of the reperfusion injury. However, conflicting data have been reported regarding the dosage, the duration of the timing, and the administrative regimen of the drug. The aim of this study was, therefore, to evaluate the effects of short versus long periods of allopurinol pretreatment on the anastomotic healing of intestines, directly after being subjected to ischemia-reperfusion (IR) stress. Furthermore, the effects of an allopurinol pretreatment on the survival rate following IR stress, was also assessed. One hundred thirty-seven male Wistar rats with a median weight of 235 (range, 180-275) g used in the study. In group I (control group, N = 20) superior mesenteric artery (SMA) and collateral vessels were isolated but not occluded. In group II, the profound IR group (PIR, N = 42), the SMA was occluded immediately distal to the aorta with collateral interruption using an atraumatic arterial clip for 30 min. In group III [two days of allopurinol (ALL) pretreatment group, 2ALL, N = 38], allopurinol (100 mg/kg body wt) was given intraperitoneally on a daily basis for two days prior to the experiment. In group IV (seven days of allopurinol pretreatment group, 7ALL, N = 37), the same pretreatment and the allopurinol schedule was performed for seven days before surgery. All animals underwent 3 cm of ileal resection and primary anastomosis, 10 cm proximal to ileocecal valve. Within each group, animals were anesthetized either on the third or seventh postoperative days. Abdominal wound healing, intraabdominal adhesions, anastomotic complications, anastomotic bursting pressure measurements, and bursting site were recorded as were the histopathologic evaluation. No rats in group I, 20 rats in group II, 18 rats in group III, and 7 rats in group IV died (P = 0.0003). Anastomotic dehiscence was found in one of 20 group I, in 11 of 22 in group II, in 9 of 20 in group III, and in 3 of 30 in group IV (P = 0.0003). On the third and seventh days, the median bursting pressures of the anastomosis were determined: 42 and 235 mm Hg in group I, 17 and 105 mm in Hg in group II, 22 and 183 mm Hg in group III, and 36 and 214 mm Hg in group IV (P < 0.0001). The burst occurred at the anastomoses in all animals tested on the third postoperative day, one in group I, six in group II, four in group III and one in group IV on the seventh postoperative day (P < 0.01). All deleterious effects of reperfusion injury on intestinal anastomosis healing, including survival rates and the histopathological parameters, were significantly prevented by seven days, but not two days, of high-dose allopurinol pretreatment. PMID- 11281196 TI - Isolation and preliminary characterization of the medium-chain fatty acid:CoA ligase responsible for activation of short- and medium-chain fatty acids in colonic mucosa from swine. AB - The ATP-dependent activation of short- and medium-chain fatty acids to their respective CoA thioester adducts was investigated in the colonic mucosa from swine. Subcellular fractionation of a homogenate of the mucosa from the entire length of the colon revealed a predominantly mitochondrial localization for activity toward fatty acids ranging from propionate through laurate. These activities could be released from mitochondria in soluble form by freeze-thaw lysis. Purification of these activities revealed that they all appeared to reside with a single enzyme. This suggests that the entire colon contains a single form of medium-chain fatty acid:CoA ligase (MCFA:CoA ligase). The ligase also had activity toward benzoate and salicylate, although this activity was significantly lower than activity toward medium-chain fatty acids. The enzyme had the highest activity at Vmax with butyrate as substrate and had the lowest Km for octanoate. Butyrate and octanoate were mutually inhibitory. Activity toward both substrates was also efficiently inhibited by cyclohexane carboxylate. The molecular weight of the enzyme was estimated by gel filtration chromatography to be ca. 46,500. These data indicate that the colonic MCFA:CoA ligase is significantly different from the hepatic and kidney MCFA:CoA ligases. PMID- 11281198 TI - The 2000 NCRP Lauriston S. Taylor lecture--administered radioactivity: unde venimus quoque imus. National Council on Radiation Protection and Measurements. AB - In this lecture, which celebrates Lauriston S. Taylor, a pioneer in radiation protection and founding president of the NCRP, I review some features of medically administered radioactivity through the past century and attempt to forecast some aspects of its future. I have used as my guide NCRP Report No. 70, Nuclear Medicine--Factors Influencing the Choice and Use of Radionuclides in Diagnosis and Therapy. The following topics are addressed: (1) decision-making considerations in the choice of radiopharmaceutical drug products, (2) factors in choosing an instrument for nuclear medicine procedures, (3) radiation dose, (4) evaluation of radionuclide procedures and their clinical utility, and (5) guidelines for performing nuclear medicine procedures. PMID- 11281199 TI - The new millennium: values, perceptions of risk, and the key roles of science and technology. AB - Radiation protection and management of radioactive waste streams and products are certain to be important areas of public policy, worker education, and technology development in the new millennium. Overriding values of freedom, sustainability, transparency, and public participation in decision making about technology's benefits and risks will shape the public policy agenda. Early engagement of stakeholders in the identification and assessment of risks and in communications about risk management will be beneficial in most cases. Putting specific environmental problems into broader public health and ecologic context will be helpful to all parties and will improve decisions about how best to utilize precious resources and enhance public confidence in the process and the outcomes. PMID- 11281197 TI - Blood purine and energy status in rats with colitis. AB - Colitis reduces the blood and tissue levels of adenosine deaminase and adenylate deaminase. Whether this has any effect on blood purines remains to be determined. The aim of this study was to measure the adenylate pool, substrates of the above enzymes, and energy status in blood from rats with colitis. Colitis was induced by intrarectal administration of acetic acid and followed over a period of seven days. The levels of ATP, ADP, AMP, adenosine, inosine, and uric acid were analyzed by HPLC, and energy status was estimated. Myeloperoxidase was used as a marker of colitis. Concentrations of ATP, ADP, AMP and adenosine decreased during days 1-5, whereas energy status decreased on day 2. The concentrations of inosine, uric acid, and hemoglobin remained unaltered, whereas colonic myeloperoxidase activity increased. These, findings demonstrate colitis-induced reduction of the circulating purines, which may be due to their enhanced usage for the repair of the inflamed colon. PMID- 11281200 TI - What views and uses of radiation sources in the 21st century? AB - Considering that in 1899 neither biotechnology nor the electronic revolution were foreseen, some humility might be advisable when one looks into the crystal ball for the future role of radiation sources. In the past 50 years, nuclear medicine, nuclear weapons, and nuclear power have had a huge impact in the world. In the next 50 years, nuclear weapons may be phased out, nuclear power revived, and nuclear medicine may continue, especially for diagnostic purposes. Conflicts between great powers and blocks will no longer be about territorial or ideological domination but about trade, finance, information, and the environment and the weapons used will not be bombs but investments, credits, and control of information. Nuclear power-still based on fission-will be relaunched and get more uses, e.g., to propel ships, to produce heat for industry and for space heating, and perhaps to desalinate water. The public will be more at ease with radiation as it is better educated, as nuclear safety continuously improves and new types of nuclear power plants emerge, as waste sites fail to cause any problems, and as no other energy source is found to deliver so much energy at reasonable cost with negligible impact on climate and environment. One kilogram of oil corresponds to 4 kWh of electricity. One kilogram of uranium fuel corresponds to 50,000 kWh, and 1 kg of plutonium 6,000,000 kWh! In nuclear medicine, radiation may give way to other treatments as the understanding of cancer advances. On the other hand, the extreme ease with which sources of radiation can be identified is unmatched and likely to make them useful tools as tracers and markers in medicine-and other fields-for a long time. For certain uses--perhaps food irradiation--radiation sources, such as cobalt, may be replaced by accelerators which may be switched on and off at will. As more sources are used, registration and control of them must be made very effective around the whole world. Very high natural emissions of radon will continue to call for cautionary measures, but many other nonradiating substances will be identified as hazardous to health and call for vigorous intervention. PMID- 11281201 TI - Deterministic effects. AB - Deterministic effects are distinguished from stochastic effects for radiation protection purposes by the following characteristics: both incidence and severity increase as a function of dose after a threshold dose is reached. Cell killing is central to all deterministic effects with the exception of radiation-induced cataracts. The understanding of radiation-induced killing of cells has increased greatly in the last decade with an extraordinarily intense interest in apoptosis. Programmed cell death has long been known to developmental biologists and the importance of cell death has been recognized and quantified by tumor biologists and students of cell kinetics but the coining of a new name and the increase of understanding of the molecular aspects of cell death has stimulated interest. Some cells appear to be very sensitive to radiation and undergo apoptosis, whereas others such as fibroblasts do not with equal frequency. This characteristic, like many others, underlines the genetic differences among cell types. We are reaching a time that there are techniques and the knowledge to apply them to clinical and radiation protection problems. In radiotherapy, success depends on the differential effect between tumor and normal tissues that is obtained. To design the optimum therapy, a profile of both the tumor cells and the cells of the normal tissues that may be at risk would help. The profile would characterize the radiosensitivity and the underlying factors, which could help in the choice of adjunct therapy for tumor and normal tissue. Fibrosis, a common unwanted late effect, appears to be influenced by genetic factors, at least in experimental animals. Techniques are available for treating people as individuals more than ever before, and that must be a good thing to do. Protection against deterministic effects would seem an easy matter but we are uncomfortably ignorant of the precise effect of protracted low-dose irradiation on tissues, such as the bone marrow and the testis, important features of risk in space. Entering the new century, it may be timely to classify radiation effects, as Radiation Effects Research Foundation (RERF) has done, into cancer, genetic effects, and noncancer effects. The recognition in the atomic-bomb survivors of noncancer effects at doses on the order of 0.5 Sv (half the dose level considered a threshold in earlier studies) should stimulate interest in deterministic effects. PMID- 11281202 TI - Resolving the molecular mechanisms of radiation tumorigenesis: past problems and future prospects. AB - Major goals in experimental radiobiology continue to be the resolution of the fundamental mechanisms of cellular radiation response and to gain detailed understanding of the contribution made by these responses to the complex in vivo processes of tumor development. The coupling of this framework of fundamental knowledge to epidemiological measures of risk greatly strengthens the biological basis of radiological protection. A selective overview is provided of recent advances in the understanding of the mechanisms and genetics of radiation tumorigenesis and their possible implications. It is suggested that rapid technical and academic advances in biology greatly expand the opportunities to further refine scientific knowledge on tumor risk after radiation. PMID- 11281203 TI - Radiation risk estimates in the beginning of the 21st century. AB - In the early years of the 21st century, results from a number of epidemiologic studies of populations with specific ionizing radiation exposures will become available. These include populations with accidental exposures in the former Soviet Union and elsewhere and populations with occupational exposures from routine operations of nuclear power plants. The strengths and limitations of these studies are reviewed together with the radiation protection questions they may answer. Many of these studies will provide specific information to complement the atomic-bomb survivor studies, particularly the effects of dose-rate and exposure protraction, modifiers of radiation risks (both environmental and host factors), and different types of radiation. These studies will therefore be important as a test of the adequacy of the current scientific bases for the radiation protection of workers and the general public. An example is thyroid cancer risk in young children following the Chernobyl accident, which has brought attention to a very high sensitivity of very young children that was difficult to assess on the basis of atomic-bomb data alone. Radiation protection will also benefit from formal comparisons and combined analyses of data from populations with different exposure patterns and exposures. Finally, future epidemiological studies will be most valuable if they are well focused, designed specifically to answer outstanding radiation protection questions. An integrated approach based on epidemiology and mechanistic studies, in which epidemiologic studies are designed to test specific mechanistic hypotheses and realistic mechanistic models are used for the analysis of epidemiological data, will probably be the most fruitful for radiation protection. PMID- 11281204 TI - Estimation of the hereditary risks of exposure to ionizing radiation: history, current status, and emerging perspectives. AB - This paper provides a brief overview of the advances in the field of the estimation of the genetic risks of exposure of human populations to ionizing radiation from the early 1950's to the present and of the developments that are anticipated in the coming years. The latter are based on the view that the insights gained from human genetics, especially human molecular genetics, will be increasingly applied to address problems in risk estimation. Owing to the paucity of human data on radiation-induced mutations, mouse data on radiation-induced mutations are used to predict the risk of genetic diseases in humans using the doubling dose method. With this method, the risk per unit dose is expressed as a product of three quantities, i.e., P x 1/DD x MC where P is the baseline frequency of genetic diseases, 1/DD (the relative mutation risk per unit dose; DD refers to the doubling dose, i.e., the radiation dose required to produce as many mutations as those that occur spontaneously in a generation) and MC is the disease class-specific mutation component (a measure of the relative increase in disease frequency per unit relative increase in mutation rate). The five important changes that are now introduced in genetic risk estimation include (1) an upward revision of the baseline frequency of Mendelian diseases to 2.4% (from 1.25% used until the early 1990's); (2) a reversion to the conceptual basis for DD calculations used in the 1972 BEIR report of the U.S. National Academy of Sciences, namely, the use of human data on spontaneous mutation rates and mouse data on induced mutation rates (instead of the use of mouse data for both these rates as has been the case from mid-1970's until the early 1990's); (3) the fuller development and use of the MC concept for predicting the responsiveness of Mendelian and multifactorial diseases to increases in mutation rate; (4) the introduction of a new disease-class-specific quantity called the "potential recoverability correction factor" or PRCF in the risk equation to bridge the gap between the rates of induced mutations in mice and the risk of inducible genetic diseases in humans; and (5) the introduction of the concept that multisystem developmental abnormalities are likely to be among the principal phenotypes of radiation induced genetic damage in humans. All these advances now permit, for the first time in 40 y, the estimation of risks for all classes of genetic diseases. For a population exposed to low-LET, chronic or low-dose irradiation, the risks predicted for the first generation progeny are the following (all estimates are per million live born progeny per gray of parental irradiation): autosomal dominant and x-linked diseases, approximately 750 to 1,500 cases; autosomal recessive, nearly zero; chronic multifactorial diseases, approximately 250 to 1,200 cases; and congenital abnormalities, approximately 2000 cases. The total risk per gray is of the order of approximately 3,000 to 4,700 cases, which represent approximately 0.4 to 0.6% of the baseline frequency of these diseases (738,000 per million) in the population. The advances anticipated in the coming years are likely to permit the estimation of genetic risks of radiation with greater precision than is now possible. PMID- 11281206 TI - Rebuilding trust in established institutions. AB - Established institutions responsible for the setting and enforcement of radiation protection standards are facing increased demands from the public to be more accountable. In the past, the National Council on Radiation Protection and Measurements (NCRP) and others within the radiation protection community have relied chiefly on professional relationships to build trust among government agencies, health care providers, and industry. During the last four decades, the trust of the public in society's institutions has eroded significantly. This diminishment of faith in recognized experts and the unparalleled characteristics of the production, testing, and use of nuclear weapons pose a serious challenge to those within the radiation protection establishment. When other factors such as the limits of scientific knowledge are considered, the issue of trust takes on added significance. This presentation will explore how radiation standard-setters and regulators can build trust by respecting the demands of a skeptical populace for meaningful participation in the decision-making process. PMID- 11281205 TI - Public involvement in science and decision making. AB - Members of the public are becoming increasingly interested in understanding risks associated with their exposure to radionuclides and chemicals in the environment. They also want to be more involved in decision making about future exposures to risks. This paper reviews one community's involvement in decisions about technical methods to calculate soil cleanup criteria for the Rocky Flats Environmental Technology Site near Denver, Colorado. The public anticipated that much of the site would be available for their use following cleanup. Final decisions regarding the future use of the site have yet to be made; however, the soil action levels were developed for this eventuality. When the public expressed considerable concern about cleanup standards for the site in 1996, a community group met to focus efforts on reviewing the cleanup standards. Later, the U.S. Department of Energy officially established the community panel to oversee an independent calculation of radionuclide soil action levels that would be used as the basis for cleanup at Rocky Flats. The primary radionuclide of concern was 239+240Pu. The Radionuclide Soil Action Level Oversight Panel (Panel) was substantively involved in all aspects of the work, from selecting the contractor, approving the computer code that formed the basis of the calculation, and assisting in developing the exposure scenarios, to selecting the values for the numerous input parameters. Communicating the uncertainties to the public, which was a major component of the analysis of soil action levels, presented a unique challenge. Over the course of the 18-mo project, the Panel and interested members of the public gained an understanding of the technical elements of the calculation and the sensitivities of the different parameters. This project serves as an excellent model of the effectiveness of public involvement in science and decision making for the future. It also illustrates the public expectations, difficulties, and time commitments encountered when making scientific decisions in a public forum. Although the process was time consuming for the scientists responsible for the calculations, a more technically defensible as well as publicly acceptable soil action level emerged. The technical approach developed during the project has been recommended for use as a decision-making tool for cleanup of the site. PMID- 11281207 TI - Scientists, policy makers, and the public: a needed dialogue. AB - Effective incorporation of scientific knowledge into public policy requires effective dialogue among scientists, policy makers, and the general public. How can this be accomplished so that all three groups have confidence in the processes leading to policies? What are the appropriate roles for scientists? What are the appropriate uses of science? Suggested answers will be proposed. PMID- 11281208 TI - Harmonizing controls for chemicals and radionuclides. AB - Risk harmonization means more than harmonizing the way that the risk from ionizing radiation is managed by regulatory agencies. It also means bringing a common language and approach to the management of all environmental carcinogens. If this goal is accomplished, it can result in better communication between radiation and chemical risk managers and a better informed public. If the public understands that there is consistency and coherence in how risks are managed, they are more likely to accept risk management decisions. PMID- 11281209 TI - Control of low-level radiation exposure: what is the problem and how can it be solved? AB - The carcinogenic risks of exposure to low-level ionizing radiation used by ICRP have been challenged as being, at the same time, both too high and too low. This paper explains that the epidemiological evidence will always be limited at low doses, so that understanding the cellular mechanisms of carcinogenesis is increasingly important to assess the biological risks. An analysis is then given of the reasons why the challenges to ICRP, especially about the linear nonthreshold response model, have arisen. As a result of considering the issues, the Main Commission of ICRP is now consulting on a revised, simpler approach based on an individual oriented philosophy. This represents a potential shift by the Commission from the past emphasis on societal-oriented criteria. These proposals have been promulgated through IRPA, and an open literature publication was published in the Journal of Radiological Protection in June 1999. On the basis of comments received and the observations presented at the IRPA 10 Conference, the Commission will begin to develop the outline of the next recommendations. It is now more than 10 y since ICRP distributed, for comment, a draft of what was to become the publication of the 1990 recommendations. The Commission plans to develop its new recommendations on a time scale of the next 4 or 5 y. In this paper, many of the issues that will need to be addressed in the development of the recommendations will be identified. These issues will cover biological effects, dosimetric quantities, and the establishment of those levels of dose at which different protection requirements will be put into place. Concepts of exclusion and exemption will need to be clarified as well as the meaning of how to achieve what the proposal identifies as "as low as reasonably practicable." Finally, the Commission has decided to develop an environmental radiation protection philosophy that will need to be developed as part of the new recommendations. PMID- 11281210 TI - Alternative goals and policy mechanisms for radiation protection. AB - For decades, radiation protection philosophy and practice have been the domain of professionals trained in the physical and medical sciences. Radiation protection policy was formulated and implemented largely without consultation with or participation by other professional disciplines or the general public. Although, from a public health standpoint, past radiation protection efforts have worked well, there are growing signs that the public has lost confidence in the radiation risk management system as currently configured. This paper draws on perspectives of political science, social psychology, economics, and decision analysis to analyze the limitations of the current radiation protection regime and to suggest alternatives for enhancing public trust and improving efficacy. PMID- 11281211 TI - The depleted uranium controversy. PMID- 11281212 TI - A genome-wide family-based linkage study of coeliac disease. AB - The susceptibility to develop coeliac disease (CD) has a strong genetic component, which is not entirely explained by HLA associations. Two previous genome wide linkage studies have been performed to identify additional loci outside this region. These studies both used a sib-pair design and produced conflicting results. Our aim is to identify non-MHC genetic loci contributing to coeliac disease using a family based linkage study. We performed a genome wide search in 16 highly informative multiply affected pedigrees using 400 microsatellite markers with an average spacing of 10 cM. Linkage analysis was performed using lod score and model free methods. We identified two new potential susceptibility loci with lod scores of 1.9, at 10q23.1, and 16q23.3. Significant, but lower lod scores were found for 6q14 (1.2), 11p11 (1.5), and 19q13.4 (0.9), areas implicated in a previous genome wide study. Lod scores of 0.9 were obtained for both D78507, which lies 1 cM from the gammaT-cell receptor gene, and for D2S364, which lies 12 cM from the CTLA4 gene. PMID- 11281213 TI - Diversity of mtDNA lineages in Portugal: not a genetic edge of European variation. AB - The analysis of the hypervariable regions I and II of mitochondrial DNA in Portugal showed that this Iberian population presents a higher level of diversity than some neighbouring populations. The classification of the different sequences into haplogroups revealed the presence of all the most important European haplogroups, including those that expanded through Europe in the Palaeolithic, and those whose expansion has occurred during the Neolithic. Additionally a rather distinct African influence was detected in this Portuguese survey, as signalled by the distributions of haplogroups U6 and L, present at higher frequencies than those usually reported in Iberian populations. The geographical distributions of both haplogroups were quite different, with U6 being restricted to North Portugal whereas L was widespread all over the country. This seems to point to different population movements as the main contributors for the two haplogroup introductions. We hypothesise that the recent Black African slave trade could have been the mediator of most of the L sequence inputs, while the population movement associated with the Muslim rule of Iberia has predominantly introduced U6 lineages. PMID- 11281214 TI - Allelic association between the NRAMP1 gene and susceptibility to tuberculosis in Guinea-Conakry. AB - Forty four families from Guinea-Conakry were analysed to test for association between NRAMP1 (Natural Resistance Associated Macrophage Protein 1) polymorphisms and tuberculosis. Each family included at least one affected sib and one parent. Healthy sibs were also analysed and on average the families included four members. A total of 160 individuals were included in the final dataset. The analysis of association was performed using an extended TDT test, TRANSMlT, to allow for missing information in the parental genotypes. Three polymorphisms in the NRAMP1 gene were typed: a microsatellite (CA) repeat, a 4 bp deletion in the 3' untranslated region and a single nucleotide change in intron 4. The single base change in intron 4 was significantly associated (p = 0.036) with tuberculosis. Our results therefore confirm, using a family-based approach on a newly studied population, the previously reported association between this polymorphism and tuberculosis in a population-based study of West Africans. PMID- 11281215 TI - Allelic association in the FRAX region. AB - The sex chromosomes enable direct determination of haplotypes, and the analysis of 8 microsatellite markers from the FRAX region on Xq27-q28 contributes 7219 independent haplotypes from our study in Wessex. Allelic association increases with frequency of alleles, and is less for trinucleotide than dinucleotide repeats. The estimate of epsilon, the exponential decline of association with distance in kb, is 0.0023. The swept radius 1/epsilon estimates the distance at which disequilibrium falls to e(-1) approximately .37 of its initial value. The current study estimates the swept radius of association to be 433 kb, which is surprisingly close to estimates for SNPs, and suggests that a marker density of 1/100 kb should be powerful in regions such as FRAX. An explanation for these results is offered, and some speculations made about what will be found when SNPs are subjected to an equally intensive study in multiple regions. PMID- 11281216 TI - Monte Carlo methods for linkage analysis of two-locus disease models. AB - Parametric linkage analysis of simultaneous mapping of the two disease loci of a qualitative trait governed by a two-locus model has been shown to provide greater power in detecting linkage than standard lod-score analysis that maps a single disease locus. Despite its great potential for power gains, two-locus parametric analysis has not been used routinely in disease gene mapping. due to the computational intensity of currently available methods and programs. In this paper, we propose a Markov chain Monte Carlo (MCMC) method for performing lod score analysis of qualitative traits governed by two-locus models. This method obtains lod-score estimates that can be arbitrarily close to their corresponding exact values. The algorithm implementing this MCMC method is linear in the number of markers. This feature enables us to perform two-locus analysis mapping each trait to a set of markers, instead of just to a single marker. We analyzed an alcohol dependence dataset composed of 105 pedigrees with various sizes and various degrees of missingness in the observed marker and disease data. The estimates from our MCMC procedure match up well with the lod scores from exact analysis, but it took much less time for the MCMC procedure to obtain the results. We also performed a simulation study to investigate power gains with additional markers. Our results indicate that an additional marker on each map can provide a great deal more information for linkage measured in terms of the magnitude of lod scores. PMID- 11281217 TI - The consistency of the posterior probability of linkage. AB - When searching for trait loci along the genome, properly incorporating prior genomic information into the analysis will almost certainly increase the chance of success. Recently, we devised a method that utilizes such prior information in the mapping of trait genes for complex disorders (Vieland, 1998; Wang el al. 1999; Vieland et al. 2000). This method uses the posterior probability of linkage (PPL) based on the admixture model as a measure of linkage information. In this paper, we study the consistency of the PPL. It is shown that, as the number of pedigrees increases, the PPL converges in probability to 1 when there is linkage between the marker and a trait locus, and converges to 0 otherwise. This conclusion is shown to be true for general pedigrees and trait models, even, when the likelihood functions are based on misspecified trait models. As part of the effort to prove this conclusion, it is shown that when there is no linkage, the maximum likelihood estimator of the recombination fraction in the admixture model is asymptotically 0.5, even when the admixture model misrepresents the true model. PMID- 11281218 TI - Transmission/disequilibrium tests for quantitative traits. AB - The transmission/disequilibrium test (TDT) is a powerful method of locating disease genes. The TDT was originally proposed for use in studies of qualitative traits in families with both parents available. Recently, the TDT has been extended to studies of qualitative traits in sibships without parents available and in families with one parent available. It has also been extended for use in studies of quantitative traits in families with both parents available and in sibships with multiple offspring. In this paper, we first propose a new class of TDT-type tests for linkage in the presence of linkage disequilibrium for use in studies of families with both parents available. The TDT of Spielman et al. (1993) for qualitative traits and the TDT of Rabinowitz (1997) for quantitative traits are special cases of the new tests. Second, we propose a new class of TDT type tests for linkage for use in studies of families with one parent available. Third, we study the validity and the power of the tests using simulations. Finally, we propose a method of combining data from different types of families. The combined test is valuable and allows researchers full use of the available data in detecting linkage between a marker locus and an unobservable quantitative trait locus. An important feature of the tests proposed in this paper is that no assumptions on the distribution of the quantitative traits are needed. PMID- 11281219 TI - Simple approximations for the maximal transmission/disequilibrium test with a multi-allelic marker. AB - Spielman et al. (1993) popularized the transmission/disequilibrium test (TDT) to test for linkage between disease and marker loci that show a population association. Several authors have proposed extensions to the TDT for multi allelic markers. Many of these approaches exhibit a 'swamping' effect in which a marker with a strong effect is not detected by a global test that includes many markers with no effect. To avoid this effect, Schaid (1996) proposed using the maximum of the bi-allelic TDT statistics computed for each allele versus all others combined. The maximal TDT statistic, however, no longer follows a chi square distribution. Here, a refinement to Bonferroni's correction for multiple testing provided by Worsley (1982) based on maximal spanning trees is applied to calculate accurate upper bounds for the type I error and p-values for the maximal TDT. In addition, an accurate lower Bonferroni bound is applied to calculate power. This approach does not require any simulation-based analysis and is less conservative than the standard Bonferroni correction. The bounds are given for both the exact probability calculations and for those based on the normal approximation. The results are assessed through simulations. PMID- 11281220 TI - Should ambiguous trios for the TDT be discarded? PMID- 11281222 TI - The effects of children's presence on woman abuse. AB - The association between the presence of children and woman abuse was investigated. Data were collected from 419 women who had called the police because of an abusive incident involving their male partner. Minor children were present in the home in 3/4 of the cases and were frequent witnesses to the abusive incident. In more than 1/2 of the cases, children had witnessed the assault according to the victims; 2/3 of the victims reported that children had seen the police when they arrived. There was almost no association between the presence of children in the home and assault on women: the presence of children was not associated with cumulative incidence of abuse, severity of abuse, degree of injury, or the victim's decision-making process in calling the police. However, police were more likely to provide information and referrals to shelters when children were present. PMID- 11281221 TI - The "Medea complex" among men: the instrumental abuse of children to injure wives. AB - It has been previously documented that wife and child abuse often co-occur. The present study tested competing hypotheses about the reasons for this co occurrence, specifically trait versus instrumental theories of aggression within families. Three commonly cited catalysts (unemployment, drinking, and life-stress events) for men's abuse of family members were analyzed to determine whether they equally predict partner or child abuse. Interviews were conducted with 363 women and children about spousal and paternal abuse, and women were interviewed about sociodemographics and the stressors described above. Families were oversampled for the presence of spousal violence. Logistic regressions revealed that heavy drinking (log-odds ratio 4.86) and life stress events (log-odds ratio 1.6) predicted men's abuse of their partners. These risk factors were unrelated to child abuse. Wife battering, however, placed children at heightened risk (log odds ratio = 2.77). Children of battered women stood a 42% chance of receiving escalated abuse from their fathers. It is proposed that men's abuse of children is in many instances instrumental in order to coerce or retaliate against women, echoing the Greek myth of Medea who killed her own children to spite their father. PMID- 11281223 TI - Adult somatic preoccupation and its relationship to childhood trauma. AB - Somatic preoccupation has been associated with a variety of comorbid psychiatric conditions including childhood trauma, personality disorder, and depression. The current study was undertaken to simultaneously explore the inter-relationship of these psychiatric variables as conceptualized in a path model. Participants (N = 120), both men and women, seen for nonemergent health care in a resident-staffed internal medicine clinic, were given questionnaires exploring the presence of childhood trauma, borderline personality symptomatology, current depression, worry, and somatic preoccupation. With one exception, all simple correlation coefficients among study variables were relatively substantial. By sequencing variables into an a priori model and using a path analytic approach, several indirect and direct relationships among variables were evident. Most important, childhood trauma exhibited a direct effect on somatic preoccupation as well as indirect effects through borderline personality disturbance and current depression. These data suggest that childhood trauma may be a precursor for somatic preoccupation during adulthood. PMID- 11281224 TI - The prevalence and correlates of psychological distress following physical and sexual assault in a young adult cohort. AB - Among a birth cohort of New Zealand's 21-year-olds, 41% experienced physical or sexual assault in the previous 12 months. The level of psychological distress experienced by the 374 victims was determined in interviews assessing for symptoms indicative of posttraumatic stress disorder and ratings of impairment in activities of daily living. Of the 141 women victims, 32.6% were identified as experiencing psychological distress as were 9.9% of the 233 men. For men, bivariate analyses showed psychological distress was significantly associated with factors indicative of increased assault severity, and for women an increased likelihood of distress was associated with the location of assault and the relationship to the assailant. Positive indicators of social support were not significantly associated with less adverse psychological outcomes. However, for both men and women, resisting the assailant was associated with a reduced likelihood of psychological distress. Multivariate analyses revealed that for both women and men, unemployment uniquely predicted variance in distress, over and above that accounted for by characteristics of the assault. PMID- 11281225 TI - Psychological abuse and posttraumatic stress disorder in battered women: examining the roles of shame and guilt. AB - Psychological abuse among battered women has been relatively understudied. However, battered women's reports in the existing qualitative and quantitative research suggest that the effects of psychological abuse can be even more damaging than the effects of physical abuse. The current study attempted to clarify the relationship between psychological abuse and posttraumatic stress disorder (PTSD) within a sample of battered women by statistically controlling for the effects of physical abuse. This study also explored the affective experiences of shame and guilt as important variables in the development of PTSD in battered women. This investigation replicated previous work suggesting that battered women are very much at risk for a diagnosis of PTSD and suggests that clinicians and researchers may need to focus on psychological abuse as a predictor of PTSD symptomatology. The current findings encourage attention to shame reactions in battered women and suggest new directions in the study of PTSD for other traumatized populations. PMID- 11281226 TI - The epidemiology of physical attack and rape among crack-using women. AB - This prospective study examines the epidemiology of physical attack and rape among a sample of 171 not-in-treatment, crack-cocaine using women. Since initiating crack use, 62% of the women reported suffering a physical attack. The annual rate of victimization by physical attack was 45%. Overall, more than half of the victims sought medical care subsequent to an attack. The prevalence of rape since crack use was initiated was 32%, and the annual rate was 11%. Among those women having been raped since they initiated crack use, 83% reported they were high on crack when the crime occurred as were an estimated 57% of the perpetrators. Logistic regression analyses showed that duration of crack use, arrest for prostitution, and some college education were predictors of having experienced a physical attack. Duration of crack use and a history of prostitution were predictors of suffering a rape. Drug abuse treatment programs must be sensitive to high levels of violence victimization experienced by crack cocaine using women. Screening women for victimization, and treating the problems that emanate from it, may help make drug abuse treatment more effective. PMID- 11281227 TI - Domestic violence laws: the voices of battered women. AB - This article reports the findings from an exploratory survey of battered women's views about mandatory arrest, mandatory reporting by doctors and nurses, no-drop policies, confidentiality laws, privilege laws, court-victim advocate programs, and specialized domestic violence courts. Although there was general support for the adoption of these laws, some variation based on demographic and situational circumstance was found. These findings raise questions about the universalistic nature of polices developed to address the problem of domestic violence. Battered women are not a homogeneous group, and public policy may be better designed to accommodate the individual needs of victims. PMID- 11281228 TI - Pulse pressure in normotensives: a marker of cardiovascular disease. AB - The purpose of the present study was to evaluate the relation of the systemic arterial pulse pressure and other parameters derived from the 24-h arterial blood pressure (BP) monitoring to the severity of coronary artery disease, carotid lesions, and left ventricular (LV) mass index in patients without arterial hypertension. One hundred ten patients with known coronary artery disease underwent coronary arteriography, 24-h arterial BP monitoring, and ultrasound imaging of the carotid arteries and the myocardium. Measurements of 24-h arterial BP monitoring (systolic, diastolic, and average BP, pulse pressure, abnormal values of systolic and diastolic BP, and heart rate), the severity of coronary heart disease (Gensini score), intima-media thickness (IMT) of the common carotid artery and LV mass index were determined in all patients. By univariate analysis, only 24-h pulse pressure was significantly related to the severity of coronary artery disease (P < .01), carotid IMT(P < .01), and LV mass index (P < .01). In a multivariate analysis, 24-h pulse pressure was also the best predictor of the severity of coronary lesions (P = .009), carotid IMT (P = .003), and LV mass index (P = .009). Gensini score was related (P < .01) to LV mass index and not to carotid IMT. In conclusion, systemic arterial pulse pressure derived from 24-h arterial BP monitoring is related to coronary artery disease, carotid IMT, and LV mass index independently of age or any other derivative of 24-h arterial BP monitoring, indicating that this parameter could be a marker of global cardiovascular risk. PMID- 11281229 TI - Blood pressure and arterial compliance in young adults: the Minnesota Children's Blood Pressure Study. AB - The aim of this study was to assess the relation between blood pressure (BP) and arterial compliance in a healthy sample of young adults. School children (aged 10 to 14 years at entry) were surveyed in 1977 to 1978, and 1,207 were followed once to twice yearly until age 23 years. Arterial compliance was measured in 179 adults at the last follow-up visit. The sample included individuals in the upper tertile of systolic BP during the last three follow-up visits and race- and sex matched individuals in the lower two tertiles. We obtained radial artery waveforms using a calibrated tonometer device and characterized waveform morphology to determine large artery (C1) and oscillatory (C2) compliance. Blood pressure was measured using random zero sphygmomanometers. The mean and standard deviation of C1 was 2.13 +/- 0.59 mL/mm Hg and of C2 was 0.083 +/- 0.02 mL/mm Hg. Systolic BP was inversely related to C1 (P < .001) and C2 (P < .01) after adjustment for gender, height, weight, insulin, and HDL and LDL cholesterol. After adjustment, a 1 SD change in systolic BP was associated with a -0.30 mL/mm Hg change in C1 and a -.008 mL/mm Hg change in C2. Data from the Minnesota Children's Blood Pressure Study indicate that systolic BP is inversely related to arterial compliance, particularly C1 (the large artery, or capacitive compliance). PMID- 11281230 TI - Hormone replacement therapy increases plasma level of angiotensin II in postmenopausal hypertensive women. AB - The renin-angiotensin-aldosterone system plays a major role in the pathogenesis of hypertension by enhancing the production or the activity of angiotensin II (ANG II). We evaluated the effects of hormone replacement therapy (HRT) on the renin-angiotensin-aldosterone system and on bradykinin in postmenopausal women (PMW) who were hypertensive or normotensive. Subjects included 28 PMW whose elevated blood pressure (BP) was well controlled on antihypertensive agents excluding diuretics, angiotensin-converting enzyme (ACE) inhibitors, and ANG II receptor antagonists. As controls, we evaluated 16 normotensive PMW. All subjects received oral HRT daily for 6 months. The plasma levels of angiotensin I (ANG I), ANG II, and bradykinin as well as plasma renin activity (PRA) showed a significant increase in HRT in the hypertensive group, but not in the normotensive group. The serum ACE activity showed a significant decrease in both groups, but the plasma level of aldosterone was unchanged. Despite the decrease in serum ACE activity, there was an increase in the plasma ANG II level. Hormone replacement therapy increased the level of ANG II in the hypertensive women, but their BP was unaffected. The increase in plasma bradykinin level may maintain homeostasis in the presence of an increase in plasma ANG II, which is a risk factor for cardiovascular disease. Hormone replacement therapy was associated with a decrease in serum ACE and an increase in plasma bradykinin in hypertensive PMW. Accordingly, the protective effect of HRT against cardiovascular disease in PMW can be provided by a decrease in ACE activity and an increase in bradykinin. PMID- 11281231 TI - Plasma renin activity and insulin resistance in African American and white children: the Bogalusa Heart Study. AB - Recent studies have suggested that the renin-angiotensin system is a feature of the insulin resistance syndrome. However, whether such a relationship occurs in childhood and in both African Americans and whites is not clear. We examined this issue in a sample of 264 African American and white children aged 7 to 16 years who participated in a cross-sectional survey of the Bogalusa Heart Study (n = 3,524). Children were selected using a stratified random sampling procedure based on race-, age-, and sex-specific percentiles of diastolic blood pressure. Whites had higher plasma renin activity than African Americans (7.1 +/- 3.6 ng/mL/h v 5.3 +/- 3.5 ng/mL/h, P < .01). Renin activity correlated with blood pressure (BP) (r = 0.21, P < .05) and insulin resistance index defined by post-glucose 1-h insulin X 1-h glucose (r = 0.19, P < .05) only in white children. Other components of insulin resistance syndrome (percent body fat, systolic blood pressure, high-density lipoprotein cholesterol, and triglycerides) showed no relation to renin in both races using univariate analyses. The distribution of insulin resistance index and renin activity among children with elevated BP (above 90th percentile) showed that the percentage of children with both high insulin resistance index and renin values was significantly greater in whites than in African Americans (45.6% v 23.3%, P < .05). A multivariate factor analysis of risk variables of insulin resistance syndrome resulted in clusters of BP/adiposity (factor 1), lipids/adiposity (factor 2), and insulin resistance/renin/adiposity (factor 3) in white children, with adiposity linking the three factors. However, a different pattern emerged in African American children for factor 2 and factor 3, and renin was not part of the cluster in any of the three factors. These observations suggest that renin may be a component of insulin resistance syndrome detectable in early life only in whites. PMID- 11281232 TI - Familial factors in the antihypertensive response to lisinopril. AB - BACKGROUND: Although it is widely recognized that there are familial elements in the pathogenesis of hypertension, remarkably little is known about the influence of family history on response to specific antihypertensive agents. METHODS: This study was designed to address that issue by comparing the depressor response to lisinopril in a dose range of 10 to 40 mg in 74 patients enrolled as sibling pairs. Because all patients were treated with lisinopril, ambulatory blood pressure monitoring (ABPM), an objective measure not influenced by the investigators, was used to assess the primary blood pressure (BP) outcome variable. RESULTS: Diastolic BP was highly correlated between sibling pairs at baseline (r = 0.476; P < .03) and on treatment (r = 0.524; P = .0021). Ethnicity/race had a striking influence on lisinopril dose and response rate. Among African American patients, 23 of 28 reached the top dose of 40 mg/day, whereas only 14 of 36 Caucasian patients reached that dose level. Among Caucasians, 92% responded, and only 48% of African Americans. Responders were characterized by being younger and heavier, having significantly lower microalbuminuria at baseline, higher baseline renal plasma flow (RPF), and higher urinary kallikrein. CONCLUSION: Among Caucasians, the presence of a hypertensive sibling predicts a striking therapeutic response to angiotensin converting enzyme inhibition. PMID- 11281233 TI - Urinary adrenomedullin is related to ET-1 and salt intake in patients with mild essential hypertension. Salt Sensitivity Group of Italian Society of Hypertension. AB - Adrenomedullin (ADM) infusion increases salt excretion in the rat. However, there is no evidence that this substance is related to changes in salt intake in humans. In this study we sought whether the urinary excretion rate of this autacoid is related to salt intake and by the expected changes in arterial pressure in patients with mild essential hypertension. The influence of salt intake on the renal excretion of ADM was investigated in 55 hypertensive patients in a double blind, randomized and crossover study comparing a 2-week 50 mmol/day salt intake period with a 150 mmol/day salt intake period. Twenty-four-hour ADM and endothelin-1 (ET-1) excretion rate were measured by radioimmunoassay on preextracted urinary samples (intraassay confidence variable <8%). The antibodies used in these assays had minimal ADM-ET-1 cross-reactivity (<1%). Twenty-four hour microalbuminuria was measured by nephelometry. On univariate analysis changes in urinary ADM were significantly related to those in salt excretion (r = 0.33, P = .01) as well as to changes in urinary ET-1 (r = 0.56, P = .0001). Furthermore, changes in urinary albumin excretion were related to those in urinary ET-1 (r = 0.26, P = .05), but were independent of those in urinary ADM (P = .19). In a multiple regression model including age, sex, body mass index, and changes in systolic pressure, plasma renin activity and plasma aldosterone and urine volume, salt excretion resulted as the stronger independent predictor of urinary ADM (r = 0.33, P = .01). However, changes in urinary salt lost prediction power (P = .11) for urinary ADM when urinary ET-1 was introduced into the model. In this model (multiple r = 0.31) urinary ET-1 resulted to be the only independent predictor of urinary ADM (beta = 0.56, P = .0001). This study is the first to show that the renal excretion of ADM is related to changes in salt intake and that it is tightly linked to that of ET-1. The data support the notion that these autacoids play a role in the regulation of sodium metabolism in patients with mild hypertension. The intercorrelations between ET-1, ADM, and microalbuminuria are compatible with the hypothesis that ET-1 is involved in a salt-induced increase in glomerular pressure and suggest that ADM may act as a counterregulatory factor in this situation. PMID- 11281234 TI - Comparison of two calcium blockers on hemodynamics, left ventricular mass, and coronary vasodilatory in advanced hypertension. AB - Dihydropyridine and nondihydropyridine calcium channel blockers (CCB) differ in pharmacologic characteristics. Few clinical studies distinguish effects of CCB as monotherapy. We conducted a comprehensive comparison of two CCB on patients with moderate to severe hypertension. Thirty patients with pretreatment diastolic blood pressures > or = 100 mm Hg were randomly assigned to either nifedipine-GITS or verapamil-SR. Dose titration achieved a diastolic blood pressure of < or = 95 mm Hg or a decrease of > or = 15 mm Hg over 4 weeks. Clinic blood pressure (BP), 24-h ambulatory BP, exercise BP, left ventricular mass, systolic and diastolic function by echocardiography, and coronary flow reserve by split-dose thallium 201 imaging with adenosine were assessed at baseline, end of titration, 3 months and 6 months of treatment. Plasma renin activity, atrial natriuretic peptide, norepinephrine, and epinephrine were assayed. Both drugs caused similar reductions in clinic and 24-h ambulatory BP and similar reductions in left ventricular mass index. Compared to nifedipine-GITS, verapamil-SR produced a significantly lower resting and peak exercise heart rate. Nifedipine-GITS elicited a lower peak exercise systolic BP. At end titration nifedipine-GITS produced lower plasma atrial natriuretic peptide levels, no longer apparent by 6 months. Plasma norepinephrine was lower with verapamil-SR, also at end titration and at 3 months, but not at 6 months. Plasma epinephrine and plasma renin activity were unchanged by either drug. There was no difference for systolic or diastolic left ventricular function or coronary flow reserve between the two treatments. Once daily nifedipine-GITS and verapamil-SR are equally effective for reduction of arterial pressure in moderate to severe hypertension. Differences in their hemodynamic profiles and neurohormonal responses are consistent with preclinical pharmacologic characteristics. The clinical implications of their similarities and differences remain to be fully evaluated in outcome studies. PMID- 11281235 TI - ACE inhibitors, beta-blockers, calcium blockers, and diuretics for the control of systolic hypertension. AB - The objective of this study was to determine which of the common groups of antihypertensive drugs is most effective at lowering systolic blood pressure (SBP) in elderly patients with previously untreated hypertension and the percentage of patients controlled with single or sequential monotherapy. Subjects were recruited from patients attending other outpatient clinics and entered into the study if their SBP was more than 150 mm Hg after three visits. Patients were given a low and high dose of each of the main classes of drugs or placebo for 1 month each. The study was a balanced, randomized crossover design with five periods: placebo; angiotensin converting enzyme inhibitors; beta-blocking drugs; calcium-blocking drugs; and thiazide diuretics. Blood pressure (BP) was measured 24 to 26 h after the previous dose. A questionnaire for side effects was administered at each visit. Seventy-four patients entered the study. beta Blockers could not be used in 15 patients because of asthma or bronchospasm and these had two placebo periods. There were 9 of 66 patients on P, 9 of 46 on beta blockers, 4 of 65 on calcium-blocking drugs, 4 of 65 on diuretic, and 1 of 62 patients on ACE inhibitors who did not progress to the higher dose because of side effects. Decreases in SBP compared to randomized placebo were calcium blocking drugs 15 mm Hg = diuretic 13 mm Hg > ACE inhibitors 8 mm Hg = beta blockers 5 mm Hg. Blood pressure decrease correlated with placebo BP (P < .0005, r = 0.53 to 0.70). When corrected for placebo, target SBP (<140 mm Hg) was reached in between 6% to 15% of patients on monotherapy. Sequential monotherapy achieved target in 29%. Angiotensin converting enzyme inhibitors, calcium blocking drugs, and diuretics had no more side effects than placebo. Patients on beta-blockers had more side effects and the well-being score was reduced. Diuretics and calcium-blocking drugs are more effective in elderly patients at lowering SBP pressure. beta-Blockers were relatively ineffective, were frequently contraindicated, and had more side effects. Monotherapy achieved control in only a small number of patients. In elderly people with essential hypertension, therapy should be instituted with diuretics or calcium-blocking drugs, but combination therapy will usually be required to achieve goal. PMID- 11281237 TI - Alterations in calcium and magnesium content of red cell membranes in patients with primary hypertension. AB - A cellular calcium-magnesium antagonism seems to be involved in the pathogenesis of primary hypertension. Total plasma, intracellular, and membranous calcium (Ca) and magnesium (Mg) contents were determined in 39 untreated patients with essential hypertension (EH) and 40 normotensive healthy subjects (NT). Membranous and intracellular measurements were performed in erythrocytes. Ca and Mg contents were measured by atomic absorption spectroscopy and membrane protein was determined according to Bradford's method as a membranous reference. There was no significant difference in plasma Ca (NT: 2.60 +/- 0.15 v EH: 2.64 +/- 0.17 mmol/L) and Mg concentrations (NT: 0.83 +/- 0.12 v EH: 0.87 +/- 0.14 mmol/L) in the studied groups. Intracellular Mg (NT: 1.72 +/- 0.15 mmol/L v EH: 1.64 +/- 0.19 mmol/L) and Ca (NT: 2.06 +/- 0.20 mmol/L v EH: 2.10 +/- 0.24 mmol/L) contents were also not significantly different between groups. Membrane Ca content was significantly increased in the EH group (2.23 +/- 0.32 micromol/g membranous protein) compared to controls (1.05 +/- 0.30 micromol/g membranous protein, P < .01). On the contrary, membranous Mg content was significantly decreased compared to controls (0.31 +/- 0.09 v 0.50 +/- 0.10 mmol/g membranous protein content, P < .01). The Ca/Mg ratio in membranes was significantly increased in EH as compared to healthy subjects (P < .01) and correlated to mean arterial blood pressure values (r = 0.47, P < .01). We conclude that the membranous alterations of Ca and Mg metabolism, shown by increased Ca/Mg ratio in red cell membranes of hypertensive subjects, may play a role in the pathogenesis of primary hypertension. PMID- 11281236 TI - Inhibitors of arachidonic acid metabolism have variable effects on calcium signaling pathways. AB - The metabolic pathways of arachidonic acid (AA) have been shown to be important in the regulation of cellular function. Several studies have demonstrated both direct and indirect effects of products of these pathways in the regulation of vascular actions, and in particular on signaling pathways. Because intracellular calcium concentration is a significant mediator of stimulus-coupled signal transduction, we investigated the effects of AA pathway inhibitors on angiotensin II (Ang II)-induced calcium mobilization in cultured rat vascular smooth muscle cells (VSMC). Thus, specific calcium pools were examined for differential effects resulting from inhibitors of the three major pathways of AA metabolism. Inhibition of lipoxygenase (LO) with 2.5 micromol/L of 5,8,11 eicosatriynoic acid (ETI), cyclooxygenase (CO) with 2 micromol/L of ibuprofen (IBU), and cytochrome P 450 (P450) with 1 micromol/L of 7-ethoxyresorufin (7ER) all reduced total Ang II induced intracellular calcium transients ([Ca2+]i) in fura 2-loaded cultured rat VSMC. However, the sites of action affected were unique for each inhibitor. Pretreatment of VSMC with either ETI or IBU inhibited thapsigargin (TG) (1 micromol/L)-sensitive calcium increments (control; 118.0 +/- 33.1 nmol/L, n = 9, ETI; 34.7 +/- 4.8 nmol/L, n = 9, IBU; 40.3 +/- 8.8 nmol/L, n = 8, P < .05 v control). Both caffeine (CAF) and ryanodine (RY) treatment attenuated Ang II induced [Ca2+]i; however, pretreatment with ETI, IBU, or 7ER did not alter this effect. In other studies, the LO inhibitor ETI attenuated Ang II-induced Ca2+ influx, whereas inhibitors of CO and P450 pathways had no effect. These data show that 1) E PMID- 11281239 TI - Doxazosin, an alpha1-adrenergic antihypertensive agent, decreases serum oxidized LDL. AB - Oxidized low-density lipoprotein (ox-LDL) is well known to play an important role in atherogenesis through the recruitment of monocytes into vessel walls and the deposition of cholesterol ester in the macrophages, which leads to the formation of lipid-rich plaque. It was assumed that only trace amounts of ox-LDL were present in plasma because the half-life of ox-LDL was only a few minutes. Recently, through the use of a monoclonal antibody against ox-LDL, a quantitative method to measure serum ox-LDL concentration has been developed. Metabolites of doxazosin, an alpha1-adrenergic antihypertensive agent, have been reported to inhibit oxidation of LDL in vitro. In this study, we investigated the in vivo effect of doxazosin on LDL oxidation using this new method to measure serum ox LDL concentration. After the administration of doxazosin for 1 to 2 months, serum concentration of ox-LDL decreased significantly (P < .05). Although the reduction of ox-LDL concentration does not strictly indicate doxazosin's antiatherosclerotic effect, it may constitute one of doxazosin's additional weapons beside lowering blood pressure and serum lipid values in the prevention of atherosclerosis. PMID- 11281238 TI - Soluble E-selectin in essential hypertension: a correlate of vascular structural changes. AB - BACKGROUND: Increased expression of the endothelial leukocyte adhesion molecule E selectin is implicated in vascular disease and may accompany the development of hypertension. We evaluated plasma soluble (s) E-selectin to assess its relationship with endothelium-dependent and endothelium-independent vasodilation in patients with hypertension. METHODS: Thirty-one previously untreated and uncomplicated essential hypertensive patients were compared with 16 normotensive controls for changes in forearm blood flow (by strain-gauge plethysmography) in response to brachial artery infusion of the endothelium-dependent vasodilator acetylcholine, and of the endothelium-independent vasodilator sodium nitroprusside. As an index of structural changes, minimal forearm vascular resistances were calculated as the ratio between maximal vasodilation after 13 min of ischemia and mean blood pressure. RESULTS: Responses to acetylcholine were significantly lower and minimal forearm vascular resistances higher in hypertensives versus controls, whereas responses to nitroprusside were comparable. Baseline sE-selectin concentrations were (mean +/- SEM) 37.4 +/- 1.8 ng/mL in hypertensives and 27.8 +/- 0.7 ng/mL in normotensives (P < .001). In essential hypertensive patients, a significant (P < .01) correlation with the response to nitroprusside (r = -0.47) was found, but not with the response to acetylcholine or minimal forearm vascular resistances. sE-selectin was also positively correlated with age and LDL cholesterol. At multivariate analysis, sE selectin remained significantly correlated with nitroprusside responses and LDL cholesterol. CONCLUSIONS: In patients with essential hypertension, plasma levels of sE-selectin are higher than in normotensive controls and mostly related to structural vascular changes. PMID- 11281240 TI - Left ventricular function impairment in pregnancy-induced hypertension. AB - The changes induced by transient hypertension on cardiac structure and function are unclear. Pregnancy-induced hypertension offers a natural and spontaneous model of this condition. To assess the potential of echocardiographic Doppler to unmask left ventricular function impairment, we studied 28 women aged 26.4 +/- 7.2 years with pregnancy-induced hypertension defined as blood pressure higher than 140/90 mm Hg in the third trimester of pregnancy without a history of hypertension. Twenty normal pregnant women, aged 27.5 +/- 6.4 years, were the controls. Left ventricular diastolic diameter, fractional shortening, E velocity, A velocity, E/A ratio, isovolumetric relaxation time (IRT), isovolumetric contraction time (ICT), ejection time (ET), and the combined index of myocardial performance (Tei index = IRT + ICT/ET), were calculated by echocardiography Doppler 2 to 4 days postpartum. There were statistically significant differences between groups in the following parameters: E/A ratio: 1.3 +/- 0.3 in pregnancy induced hypertension v 1.5 +/- 0.3 in normal pregnant women (P < .05), IRT: 104 +/- 14 msec v 84 +/- 7 msec (P < .000), and the Tei index: 0.51 +/- 0.15 v 0.35 +/- 0.04 (P < .00), respectively. According to this data pregnancy-induced hypertension evaluated 2 to 4 days after delivery showed left ventricular dysfunction, mainly diastolic. The IRT and the Tei index are the most useful echocardiographic parameters to unmask left ventricular dysfunction in pregnancy induced hypertension. PMID- 11281242 TI - Low-dose combination therapy: an important first-line treatment in the management of hypertension. PMID- 11281241 TI - The role of nitric oxide and the renin-angiotensin system in salt-restricted Dahl rats. AB - To elucidate the role of nitric oxide (NO) and renin-angiotensin system (RAS) in the development of salt-sensitive hypertension, we investigated the pressor responses and renal histologic changes after long-term inhibition of endogenous NO synthesis in Dahl-Iwai salt-sensitive (DS) and salt-resistant (DR) rats under salt-re-stricted conditions that exaggerate RAS activation. Male DS and DR rats (6 weeks old) were fed with a low-salt (0.3%) diet for 5 weeks. NG-nitro-L arginine (L-NA; dissolved in 60 mg/L deionized water), an arginine analogue acting as a NO-inhibitor, was also administered for 5 weeks. L-NA administration induced a gradual increase in systolic blood pressure (SBP) in both strains, and the pressor response in DS rats was apparently more enhanced relative to that in DR rats. Urinary nitrate plus nitrite (u-NOx) excretion was decreased by L-NA, with a significant negative correlation between SBP and u-NOx excretion in DS rats but not in DR rats. Plasma renin activity and urinary aldosterone level were significantly increased in L-NA-treated DS rats on week 5. Marked histologic changes with glomerular sclerosis and increased proteinuria and urinary N-acetyl beta-glucosaminidase excretion were found in L-NA-treated DS rats but not DR rats. Competitive RT-PCR of mRNA extracted from the glomeruli revealed that angiotensin II type 1 receptor (AT1R) mRNA level was significantly lower in DS rats than in DR rats at week 2, and that L-NA administration significantly reduced glomerular AT1R level of DS rats at week 5, possibly because of downregulation. Our results showed that, even under sodium restriction, the pressor response and renal injury induced by chronic NO inhibition were markedly more enhanced in DS rats than in DR rats, which indicates that depletion of NO participates in both the development of hypertension and glomerular injury in DS rats through a potential activation of RAS irrespective of sodium loading. These data suggest that endogenous NO is an essential determinant of salt-sensitive hypertension in DS rats. PMID- 11281243 TI - Combination therapy as the initial drug treatment for hypertension: when is it appropriate? PMID- 11281244 TI - Laragh's lessons in pathophysiology and clinical pearls for treating hypertension. PMID- 11281245 TI - Hypertension due to perinephric compression: the "Page" kidney. PMID- 11281246 TI - The insulin resistance epidemic in India: fetal origins, later lifestyle, or both? AB - In India there is a rapidly escalating epidemic of insulin resistance syndrome (diabetes and coronary heart disease). Contribution of genes and environment is under debate. Small size at birth coupled with subsequent obesity increases risk for insulin resistance syndrome in later life. The tendency of Indians to have higher body fat and central adiposity compared with other races may be programmed in utero. The adipose tissue releases not only fatty acids but also a number of proinflammatory cytokines, which increase insulin resistance and cause endothelial dysfunction. Crowding, infections, and environmental pollution in Indian cities may increase cardiovascular risk by stimulating fat cells. Prevention of diabetes and coronary heart disease in India will have to be approached throughout the life cycle. PMID- 11281247 TI - Rationale for antioxidant therapy in premature infants to prevent bronchopulmonary dysplasia. AB - Bronchopulmonary dysplasia (BPD) is a chronic lung disease common in premature infants that can cause severe complications. BPD's pathogenesis is multifactorial but oxidative processes during the first week of life are thought to play a key role in the development of the disease. Prevention of this oxidation through antioxidant therapy is therefore of interest. Unfortunately, this therapy has not proven effective, most likely owing to the nonspecific strategy used. This review focuses on the challenges facing researchers and clinicians in improving the antioxidant status of premature infants in order to prevent or lessen the severity of BPD. This review will focus on the particular oxidations that may lead to BPD and the specific therapies that can be used to counter these processes. PMID- 11281249 TI - Plant sterols and their derivatives: the current spread of results. AB - Whereas the cholesterol-lowering action of plant sterols and their derivatives has been well established, more recent results have produced new information concerning the similar effects of various mixtures and dose-response relationships. Moreover, although these materials have generally been viewed as safe for long-term use in almost all sectors of the population, some concerns remain regarding the impact of phytosterol consumption on other lipid-soluble nutrients, which need to be further addressed. Further work is also required to define the relationship between plant sterol consumption, circulating sterol levels, and risk of certain cancers. PMID- 11281248 TI - Does dieting during lactation put infant growth at risk? AB - For some women, postpartum retention of weight gained during pregnancy may contribute to obesity. A recent 10-week randomized intervention showed that infants of initially overweight, lactating mothers who exercised and dieted to lose an average of 0.5 kg/week grew normally. The findings of this study support the Institute of Medicine guidelines for weight loss in overweight women who are exclusively breast-feeding their child. PMID- 11281250 TI - Fruits and vegetables and the risk of stroke. AB - Combining data from two large-scale prospective observational studies in women and men, investigators link the consumption of fruits and vegetables to the risk of ischemic stroke. Of interest is their breakdown of these foods into cruciferous vegetables and citrus fruit juice, which confer the greatest protection. PMID- 11281251 TI - Analysis of pesticide runoff from mid-Texas estuaries and risk assessment implications for marine phytoplankton. AB - During 1993, estuarine surface water samples were collected from the mid-Texas coast (Corpus Christi to Port Lavaca, TX). Agricultural watershed areas as well as tidal creeks immediately downstream were chosen as sampling sites along with adjoining bay sampling stations. Collections were made throughout the growing season (February to October 1993) before and after periods of significant (> 1.25 cm) rainfall. All samples were initially screened for the presence of pesticides using enzyme-linked immunosorbent assay (ELISA) test kits (EnviroGard) for triazine herbicides and carbamate insecticides. All samples were extracted and then analyzed using gas chromatography (GC) for quantification of atrazine. Only samples testing positive for carbamate insecticides via ELISA were further extracted for GC analysis to quantify aldicarb and carbofuran. Additionally, laboratory toxicity tests using phytoplankton were examined from published, peer reviewed literature and compared with the atrazine field levels found in Texas. Results of ELISA screening indicated the presence of triazine herbicides in nearly all samples (>93%). GC analysis further confirmed the presence of atrazine concentrations ranging from <0.01-62.5 microg/L. Screening tests also found detectable levels of carbamate insecticides (aldicarb and carbofuran) that were also confirmed and quantified by GC. Comparison of measured concentrations of atrazine compared with published toxicity tests results indicated that there was a potential environmental risk for marine/estuarine phytoplankton in surface waters of Texas estuaries, particularly when the chronic nature of atrazine exposure is considered. PMID- 11281252 TI - Organochlorine pesticides and heavy metals in muscle and ovaries of Gulf coast striped bass (Morone saxatilis) from the Apalachicola River, Florida, USA. AB - Eight female Gulf coast striped bass (Morone saxatilis) broodfish collected for induced spawning from the Apalachicola River below the Jim Woodruff Lock and Dam were analyzed for organochlorine pesticides (OCs) and metals in muscle and ovarian tissues. Chemical analyses revealed that muscle and ovaries contained detectable amounts of OCs and metals. Concentrations of p,p'-DDE, a derivative of the pesticide DDT, in muscle and ovary (0.54 and 0.65 microg/g, respectively) were significantly higher than alpha-chlordane, dieldrin, and p,p'-DDD. The presence of p,p'-DDE, an antiandrogenic compound, in females suggests that the compound also may be present in male striped bass. Concentrations of Cr, Hg, Mg, and Mn were higher in muscle than in ovarian tissues. Concentrations of Hg have almost doubled in muscle tissues (0.85 microg/g) and tripled in ovaries (0. 15 microg/g) in our samples, compared with the data from 1986 to 1989. Organochlorine pesticides and metal contaminants were present in muscle and ovarian tissues of adult females and may have been retarding development of eggs leading to low hatching rates. PMID- 11281253 TI - Influence of paraquat, glyphosate, and cadmium on the activity of some serum enzymes and protein electrophoretic behavior (in vitro). AB - In vitro study for the determination of the toxicity of some pesticides (glyphospate and paraquat) and cadmium chloride (CdCl2) on the activities of serum acetylcholinesterase (AChE), lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (AlP), and acid phosphatase (AcP) is described. Changes in electrophoretic patterns of serum proteins were also tested. Results revealed that glyphosate was effective on all enzymes except AcP. Its IC50 values (the concentration of compound that inhibits 50% of the enzyme activity in 1 h at 37 degrees C) were 714.3, 750, 54.2, 270.8, and 71.4 mM for AChE, LDH, AST, ALT, and AlP, respectively. The inhibitory effect of paraquat varied markedly among all enzymes. The IC50 values of paraquat were 321.4 and 750 mM for AST and ALT, respectively. It had mild effect on AChE and LDH; and no effect on the activities of AlP and AcP. The effect of CdCl2 was pronounced with AChE, ALT, AlP, and AcP, and no effect on LDH and AST was found. The corresponding IC50 values were 77.7, 22.2, 33.3, and 83.3 mM for AChE, ALT, AlP, and AcP, respectively. Polyacrylamide gel electrophoretic patterns of serum proteins showed marked differences with glyphosate and CdCl2 but not with paraquat. The results suggest that the in vitro enzyme-activity test seems to have a potential for the assessment of pesticide and heavy metal toxicity. PMID- 11281254 TI - Population growth of Euchlanis dilatata (Rotifera): combined effects of methyl parathion and food (Chlorella vulgaris). AB - In the present work, the combined impact of four concentrations (0, 0.0625, 0.125, and 0.25 mg/L) of methyl parathion and three densities (0.5 x 10(6), 1.0 x 10(6), and 2.0 x 10(6) cells/mL) of the green alga Chlorella vulgaris on the population growth of Euchlanis dilatata was studied. In general, regardless of the food level, an increase in the concentration of methyl parathion resulted in a significant reduction of the maximal population density and rate of population increase. The population growth rate in the controls ranged from 0.248 to 0.298; rates were lower in the presence of the pesticide. At any toxicant concentration, rotifers fed higher algal density showed significantly higher population growth compared with those at lower food levels. An interaction between toxicant and food level was evident on the population growth of E. dilatata. Results have been discussed in light of the protective role of algal density on the toxic effects of insecticides on rotifers and the differences in susceptibility to toxicants between planktonic and littoral rotifers. PMID- 11281255 TI - Effects of thiobencarb herbicide to an alga (Nannochloris oculata) and the cladoceran (Daphnia magna). AB - Chronic toxicity studies were conducted with an algae (Nannochloris oculata) and the cladoceran (Daphnia magna) to determine their relative sensitivities to the thiocarbamate herbicide thiobencarb (S-4-chlorobenzyl diethylthiocarbamate). Most of the algal populations were initially affected by exposure to the herbicide. Thiobencarb concentrations higher than 0.5 mg/L significantly reduced algal densities after 24-h exposure. The 24-h static EC50 in D. magna was 3.01 mg/L. The sublethal effects of 0.3, 0.37, 0.5, 0.75, and 1.5 mg/L of thiobencarb concentrations on the survival, reproduction, and growth of D. magna were monitored for 21 days. The parameters used to determined the effect of the herbicide on D. magna were mean total young per female; mean brood size; days to first brood; intrinsic rate of natural increase (r); growth; and survival. Reproduction was significantly reduced at thiobencarb concentrations of 0.30 mg/L and higher while survival was affected after exposure to 0.75 and 1.5 mg/L of the pesticide. The r value decreased with increasing concentrations of thiobencarb. Growth, as measured by body length, was depressed significantly after exposure to all herbicide concentrations tested. PMID- 11281256 TI - Chronic toxicity tests with Daphnia magna for examination of river water quality. AB - Chronic toxicity tests with Daphnia magna were applied for examination of river water quality. Water was sampled from the Maioka River in Yokohama City on May 14, 20, and 27, 1999, and used for the test after solid-phase extraction. The chronic test was carried out according to the OECD method. The duration was 21 days and the total number of live offspring produced per parent animal was counted. The results of the tests showed, survival rates of 100% using river water sampled on May 14 and 20 and the total numbers of live offspring produced per parent animal did not differ from the control. However, the survival rate of the sample collected on May 27 was 0% and the pesticides, fenitrothion, and thiobencarb were detected in the water. In addition to the river water samples, reconstituted water (Elendt M7) with additions of fenitrothion and thiobencarb was prepared to investigate mortality. When the reconstituted water with thiobencarb was applied to the test, the total number of live offspring produced per parent animal did not differ from the control. In contrast, when reconstituted water with fenitrothion was applied to the test, most parents were alive, but the total number of live offspring produced per parent animal was apparently different. The results of the above tests indicate that D. magna was affected not only by fenitrothion in the river water collected on May 27, but also by other factors that were not clarified in this study. PMID- 11281257 TI - The histopathological effects of thiodan on the liver and gut of mosquitofish, Gambusia affinis. AB - Thiodan (33.7% endosulfan), a polychlorinated cyclodiene insecticide, was evaluated for its histopathological effects on mosquitofish, Gambusia affinis, by light microscopy. Fish were exposed to doses of 0.00 (control), 1.00, 2.50, and 5.00 microg/L on days 7, 14, 21, and 30. No histopathological effects were apparent at control group. The histopathological alterations were characterized as oedema, degeneration, accumulation of lymphocytes in the lamina propria, disintegration of villuses, pycnotic state of nuclei, and necrosis in gut; degeneration, hypertrophy, sinusoids enlargement, hemorrhage, pycnosis position of nuclei, vacuolization of cell cytoplasm, infiltration of mononuclear lymphocyte, and congestion in liver. These alterations were time- and dose dependent. PMID- 11281258 TI - Optimizing microwave-assisted solvent extraction (MASE) of pesticides from soil. AB - Microwave-assisted solvent extraction (MASE) was investigated as an alternative for extraction of parathion (O,O-diethyl O-4-nitrophenyl phosphorothioate), methyl parathion (O,O-dimethyl O-4-nitrophenyl phosphorothioate), p,p'-DDE [1,1' dichloro-2,2-bis(4-chlorophenyl)ethane], hexachlorobenzene (HCB), simazine (6 chloro-N2,N4-diethyl- 1 ,3,5-triazine-2,4-diamine) and paraquat dichoride (1,1' dimethyl-4,4'-bipyridinium) from two different soils and from an earthworm growing substrate. The matrices were fortified with 14C-radiolabeled pesticides and extracted with various solvent systems under different microwave conditions. Recoveries of more than 80% could be obtained depending on the used microwave conditions and solvent, except for paraquat whose recovery was generally less efficient. Thus, MASE can be successfully used to extract pesticides from environmental and biological samples and could be a viable alternative to conventional extraction methods. The technique uses smaller amounts of organic solvents, thereby minimizing the costs of the analysis and the disposal of waste solvent. PMID- 11281259 TI - Adsorption of methabenzthiazuron on six allophanic and nonallophanic soils: effect of organic matter amendment. AB - This article reports on methabenzthiazuron [1-(1,3-benzothiazol-2-yl)-1,3 dimethylurea] (MBT) adsorption process on six agricultural allophanic and nonallophanic soils. The effect of amendment with exogenous organic matter was also studied. Adsorption kinetic fits an hyperbolic model. MBT adsorption reached an apparent equilibrium within 2 h and followed a second-order reaction. The maximum adsorbed amounts for natural soils ranged from 32 to 145 microg g(-1). Rate constants were considered relatively low (0.27-1.5 x 10(-4) [microg g(-1)](1 n) s-1); the slow process was attributed to a combined effect of difussion and adsorption. MBT adsorption fits the Freundlich model with r values > or =0.998 at P < or = 0.001 significance levels. Kf and Freundlich exponents (l/n) ranged from 5.3 to 82.1 cm3 g(-1) and from 0.66 to 0.73, respectively. Kf values for soils with a low organic matter content were lower than that obtained from the only typical allophanic soil derived from volcanic ash under study. Lineal regression analysis between Kf and organic matter content of nonallophanic soils gave a correlation coefficient of 0.980 (P = 0.02). Dispersion of Kd values together with close values of K(OM) indicate that organic matter (OM) was the principal component responsible for MBT adsorption in unamended soils. Addition of peat decreased soil pH and increased adsorption capacity for allophanic and nonallophanic soils. Kinetic experiments showed enhancements of Xmax values and lower rate constants. PMID- 11281260 TI - Conserved cassette structure of vertebrate Mr 300 kDa mannose 6-phosphate receptors: partial cDNA sequence of fish MPR 300. AB - The existence of two homologous mannose 6-phosphate receptors (MPRs) with overlapping, but distinct functions has raised the question of at what stage in the phylogenetic tree the two receptors have occurred for the first time. In this paper, we present a partial cDNA sequence of Mr 300 kDa MPR (MPR 300) from poeciliid fish (Xiphophorus). It contains a 5'-untranslated region followed by the initiator ATG, and an open reading frame that corresponds to cassettes 1-5 and part of cassette 6 of mammalian MPR 300. The size of the mRNA transcript for fish MPR 300 was comparable with that of other vertebrates. The amino acid sequence of fish MPR 300 displays 48-52% similarity with mammalian and chicken MPR 300. In particular, all the cysteine residues involved in disulfide bonding and an arginine residue, which is considered to be part of the mannose 6 phosphate binding site in cassette 3 of mammalian MPR 300, are conserved. Sequence similarities were significantly higher within cassette 3 and within cassette 5, to which a ligand-binding function has not yet been ascribed. Sequence similarities of the internal cassettes of MPR 300 are discussed with regard to the multifunctional nature of MPR 300. PMID- 11281261 TI - Hydrolysis of carnosine and related compounds by mammalian carnosinases. AB - Comparative study of hydrolysis of carnosine and a number of its natural derivatives by human serum and rat kidney carnosinase was carried out. The rate of carnosine hydrolysis was 3-4-fold higher then for anserine and ophidine. The rate of homocarnosine, N-acetylcarnosine and carcinine hydrolysis was negligible by either of the enzymes used. Our data show that methylation, decarboxylation or acetylation of carnosine increases resistance of the molecule toward enzymatic hydrolysis. Thus, metabolic modification of carnosine may increase its half-life in the tissues. PMID- 11281262 TI - Molecular cloning and sequence analysis of the Danio rerio catalase gene. AB - Catalase is an antioxidant enzyme that plays a central role in the protection against oxidative stress through the metabolism of hydrogen peroxide. Catalase has been well studied in plants, bacteria, and mammals, but little work has been done in other vertebrate species. We have cloned the zebrafish (Danio rerio) catalase cDNA containing the complete coding region and analyzed expression by both reverse transcription polymerase chain reaction and western blot. The deduced amino acid sequence predicts a protein of 526 amino acids with both the primary DNA and amino acid sequences highly conserved among vertebrate species. The major protein-heme contact points in the catalase enzyme complex are also well conserved, although several amino acids associated with the second and third levels of the major substrate channel are not, suggesting potential differences in substrate access or specificity. The 3' flanking region of the cDNA contains a dinucleotide repeat near the termination codon consisting of a near perfect CA array that is polymorphic. The rat and mouse catalase genes also contain a CA repeat sequence in the 3' untranslated region, which, along with an adjacent 5' stem-loop structure, has previously been shown to be a site for mRNA protein binding (Clerch, 1995, Arch. Biochem. Biophys. 317 (1995) 267-274). A stem-loop structure is also predicted adjacent to the zebrafish CA repeat, suggesting a similar role in catalase gene regulation. PMID- 11281263 TI - Ontogenetic changes in biochemical composition during larval and early postlarval development of Lepidophthalmus louisianensis, a ghost shrimp with abbreviated development. AB - Changes in growth and biochemical composition during the transition from egg through zoea to decapodid in the ghost shrimp, Lepidophthalmus louisianensis (Schmitt, 1935), were documented in terms of dry weight, lipid classes, fatty acid composition, and carbon to nitrogen (C:N) ratios. Larvae of the ghost shrimp were mass-reared in the laboratory (28 degrees C; 20% S) from hatching to the decapodid stage. latroscan lipid class analysis revealed that major lipid classes in recently produced eggs were phospholipids (80.8 +/- 1.3%) and triglycerides (16.0 +/- 1.1%), which decreased during the incubation period. Polar lipids (zoea 1: 77.4 +/- 1.7%; zoea II: 77.5 +/- 2.1%; decapodid: 80.0 +/- 1.7%) and neutral lipids, of which free fatty acids (zoea I: 10.5 +/- 2.7%; zoea II: 13.1 +/- 5.2%; decapodid: 7.8 +/- 2.1%) were dominant, represented the major lipid classes in the zoeal and decapodid stages. Triglycerides were present in small amounts. The predominant fatty acids of L. louisianensis eggs, zoeae and decapodids were palmitic (16:0), stearic (18:0), eicosapentaenoic (20:5omega3), oleic (18:1omega9), and arachidonic (20:4omega6). Elemental composition of eggs, larvae, and the decapodid stage revealed conspicuous changes in the C:N ratio, with N being relatively stable during larval development but C decreasing during the decapodid stage. These data suggest independence of newly hatched L. louisianensis on external energy resources. This combined with the ability to incorporate saturated fatty acids into polar lipids provides a selective advantage for fast development of new tissue and growth, characteristic of decapod crustacean larvae with lecithotrophic development. PMID- 11281265 TI - The amino acid sequence of pancreatic alpha-amylase from the ostrich, Struthio camelus. AB - The amino-acid sequence of alpha-amylase isolated from the pancreas of the ostrich, Struthio camelus was determined. The alpha-amylase (OPA) consisted of 497 amino acid residues with pyroglutamic acid at the N-terminus and no oligosaccharide. Amino acid identity between OPA and chicken, porcine and human pancreatic alpha-amylases individually, was found to be 88, 82 and 86%, respectively. PMID- 11281264 TI - Metabolic adaptations to environmental changes in Caenorhabditis elegans. AB - Metabolic adaptations to environmental changes were studied in Caenorhabditis elegans. To assess adjustments in enzyme function, maximum activities of key enzymes of main metabolic pathways were determined. After a 12 h incubation at varying temperatures (10, 20 degrees C) and oxygen supplies (normoxia or anoxia), the activities of the following enzymes were determined at two measuring temperatures in tissue extracts: lactate dehydrogenase (LDH; anaerobic glycolysis), 3-hydroxyacyl-CoA-dehydrogenase (HCDH; fatty acid oxidation), isocitrate dehydrogenases (NAD-IDH, NADP-IDH; tricarboxylic acid cycle) and isocitrate lyase (ICL; glyoxylate cycle). Incubation at 20 degrees C induced a strong increase in maximum LDH activity. Anoxic incubation caused maximum HCDH and NADP-IDH activities and, at 10 degrees C incubation, LDH activity to increase. Maximum NAD-IDH and ICL activities were not influenced by any type of incubation. In order to study the time course of metabolic adaptations to varying oxygen supplies, relative quantities of free and protein-bound NADH were determined in living C. elegans using time-resolved fluorescence spectroscopy. During several hours of anoxia, free and protein-bound NADH showed different time courses. One main result was that just at the moment when the protein-bound NADH had reached a constant level, and the free NADH started to increase rapidly, the worms fell into a rigor state. The data on enzyme activity and NADH fluorescence can be interpreted on the basis of a two-stage model of anaerobiosis. PMID- 11281266 TI - Intermediate filament proteins in echinoderm coelomocytes. AB - The presence and organization of intermediate filament (IF) proteins in petaloid coelomocytes from two species of echinoderms, the sea urchin Strongylocentrotus droehachiensis and the sea cucumber Cucumaria frondosa, were studied. Two monoclonal antibodies (IFA and Ah6) and one polyclonal antibody (W3-1) that together recognize invertebrate as well as vertebrate IF proteins were used to probe coelomocytes by immunofluorescence and immunoblotting methods. All three antibodies cross-reacted with a single Mr 68,000 sea urchin lamin, as well as two putative lamin isoforms of approximately Mr 70,000 and 68,000 in sea cucumber coelomocytes. Both IFA and Ah6 labeled granular material in the cytoplasm of sea urchin coelomocytes; by contrast, IFA labeling revealed a striking network of reticular material irregularly arrayed within the central regions of the sea cucumber coelomocyte cytoplasm. In addition, foci of Ah6-positive material were present in coelomocyte nuclei from both species. Comparison of immunoblotting patterns among whole cell and isolated nuclear preparations suggest that the cytoplasmic IF-like material is composed of Mr 46,000 and 58,000 polypeptides, while Mr 215,000 and 185,000 proteins are candidates for the immunoreactive nuclear foci. Further study of the functions of these non-filamentous arrays of IF proteins may furnish valuable insights into the evolution of IF function within vertebrate cells, particularly with respect to certain cytoplasmic and nuclear regulatory functions with which IF proteins have been speculated to be involved. PMID- 11281267 TI - Gastropod arginine kinases from Cellana grata and Aplysia kurodai. Isolation and cDNA-derived amino acid sequences. AB - Arginine kinase (AK) was isolated from the radular muscle of the gastropod molluscs Cellana grata (subclass Prosobranchia) and Aplysia kurodai (subclass Opisthobranchia), respectively, by ammonium sulfate fractionation, Sephadex G-75 gel filtration and DEAE-ion exchange chromatography. The denatured relative molecular mass values were estimated to be 40 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The isolated enzyme from Aplysia gave a Km value of 0.6 mM for arginine and a Vmax value of 13 micromole Pi min(-1) mg protein(-1) for the forward reaction. These values are comparable to other molluscan AKs. The cDNAs encoding Cellana and Aplysia AKs were amplified by polymerase chain reaction, and the nucleotide sequences of 1,608 and 1,239 bp, respectively, were determined. The open reading frame for Cellana AK is 1044 nucleotides in length and encodes a protein with 347 amino acid residues, and that for A. kurodai is 1077 nucleotides and 354 residues. The cDNA-derived amino acid sequences were validated by chemical sequencing of internal lysyl endopeptidase peptides. The amino acid sequences of Cellana and Aplysia AKs showed the highest percent identity (66-73%) with those of the abalone Nordotis and turbanshell Battilus belonging to the same class Gastropoda. These AK sequences still have a strong homology (63-71%) with that of the chiton Liolophura (class Polyplacophora), which is believed to be one of the most primitive molluscs. On the other hand, these AK sequences are less homologous (55 57%) with that of the clam Pseudocardium (class Bivalvia), suggesting that the biological position of the class Polyplacophora should be reconsidered. PMID- 11281268 TI - Gene structure of two-domain arginine kinases from Anthopleura japonicus and Pseudocardium sachalinensis. AB - Unusual two-domain arginine kinases (AKs) arose independently at least two times during molecular evolution of phosphagen kinases: AKs from the primitive sea anemone Anthopleurura japonicus and from the clam Pseudocardium sachalinensis. To elucidate its unusual evolution, the structures of Anthopleura and Pseudocardium AK genes have been determined. The Anthopleura gene consisted of 4 exons and 3 introns: two domains are linked by a bridge intron, and each domain contains one intron in different positions. On the other hand, the Pseudocardium gene consisted of 10 exons and 9 introns: two domains are also linked by a bridge intron, and domains 1 and 2 contains 3 and 5 introns, respectively, of which 3 introns are located in exactly same positions. Since the two domains of Pseudocardium AK are estimated to have diverged about 290 million years ago, the 3 introns have been conserved at least for this long. Comparison of intron positions in Anthopleura, Pseudocardium and C. elegans AK genes indicates that there is no intron conserved through the three AK lineages, in sharp contrast to relatively conservative intron positions in creatine kinase (CK) gene family. PMID- 11281269 TI - Heterogeneity of the ribosome-inactivating protein trichosanthin in Trichosanthes kirilowii tubers. AB - The ribosome-inactivating protein trichosanthin isolated from the tubers of Trichosanthes kirilowii, the Chinese drug Tianhuafen, has a molecular mass of approximately 26 kDa. We show here that T. kirilowii tubers also contain ribosome inactivating proteins with a small extent of structural variation from and a larger molecular mass than trichosanthin. PMID- 11281270 TI - Characterization of Japanese eel immunoglobulin M and its level in serum. AB - Japanese eel immunoglobulin M (IgM) was purified from the sera of Anguilla japonica immunized with Edwardsiella tarda FPU 347 and characterized. Analysis of the purified IgM on sodium dodecyl sulfate-polyacrylamide gels (SDS-PAGE) under reducing and non-reducing conditions revealed that the eel IgM was a tetrameric protein with a molecular weight of 790,000; it contained an equimolar heavy chain and light chain with molecular weights of 72,000 and 25,000, respectively. While the N-terminal sequence of the heavy chain, VELTQPGSMVLKPGQSLTI, showed similarity to the variable regions of those of teleost fishes Igs, the N-terminal sequence of the light chain, DIVLTQSPAVQSVQLGDT, was similar to the variable regions of chondrostei and mammalian kappa chains. Lectin-binding assays showed that the binding of concanavalin A (Con A) to the Japanese eel IgM heavy chain was competitively inhibited by D-mannose and could be abolished by alpha mannosidase treatment indicating the presence on the heavy chain of oligosaccharides, whose terminal were a bound mannoses. The average IgM concentration in the sera of the healthy eels was 3.4 mg ml(-1); it amounted to 10.3% of the total serum protein. PMID- 11281271 TI - The effects of 5-HT on sensory, central and motor neurons driving the abdominal superficial flexor muscles in the crayfish. AB - Serotonin (5-HT) induces a variety of physiological and behavioral effects in crustaceans. However, the mechanisms employed by 5-HT to effect behavioral changes are not fully understood. Among the mechanisms by which these changes might occur are alterations in synaptic drive and efficacy of sensory, interneurons and motor neurons, as well as direct effects on muscles. We investigated these aspects with the use of a defined sensory-motor system, which is entirely contained within a single abdominal segment and consists of a 'cuticular sensory neurons segmental ganglia abdominal superficial flexor motor neurons-muscles' circuit. Our studies address the role of 5-HT in altering (1) the activity of motor neurons induced by sensory stimulation; (2) the inherent excitability of superficial flexor motor neurons; (3) transmitter release properties of the motor nerve terminal and (4) input resistance of the muscle. Using en passant recordings from the motor nerve, with and without sensory stimulation, and intracellular recordings from the muscle, we show that 5-HT enhances sensory drive and output from the ventral nerve cord resulting in an increase in the firing frequency of the motor neurons. Also, 5-HT increases transmitter release at the neuromuscular junction, and alters input resistance of the muscle fibers. PMID- 11281272 TI - Interaction of actin with the capping protein, CapZ from sea bass (Dicentrarchus labrax) white skeletal muscle. AB - We have compared the functional properties of CapZ from fish white skeletal muscle with those of CapZ from chicken muscle. CapZ is a heterodimer, which enhances actin nucleation and inhibits the depolymerization process by binding to the barbed ends of microfilaments. Here, we report the interaction of CapZ not only with F-actin, but also with monomeric actin. The affinity of sea bass CapZ for G-actin estimated by enzyme-linked immunosorbent assay (ELISA) was in the microM range. This association was PIP2 dependent. Binding contacts with the barbed end of actin were delimited by both ELISA and fluorescence approaches. One site (actin sequence 338-348) was located in a helical region of the subdomain 1, region already implicated in the interaction with other actin binding proteins such as gelsolin. Another site implicates the C-terminal region (sequence 360 372) of actin. Finally, the partial competition of antibodies directed against CapZ alpha or beta-subunits towards CapZ interaction with actin filaments suggests both subunits participate in the complex with actin. PMID- 11281273 TI - Partial purification and characterization of 12-lipoxygenase in bullfrog erythrocytes. AB - 12-Lipoxygenase (12-LO) in bullfrog (Rana catesbeiana) erythrocytes was purified partially by ion exchange chromatography and affinity chromatography. Bullfrog 12 LO was a single chain protein with a pI of 7.1-7.8 and MW of 7.77 kDa. This enzyme did not show typical Michaelis Menten type kinetics. At low substrate concentrations, it had a lag phase and at higher substrate concentrations, the activity was inhibited. The product of linoleic acid (LA), 13-hydroperoxy-9, 11 octadecadienoic acid (13-HpODE), was an activator for the enzyme. When arachidonic acid (AA) was used as substrate, 13-HpODE also affected the Km of bullfrog 12-LO towards AA. The affinity of LA towards bullfrog 12-LO was higher than the affinity of AA. Suicide inactivation was much more rapid than that of any mammalian 12-LO reported. Hemoglobin (Hb) inhibited the activity of 12-LO partially and removing Hb eliminated this inhibition. Both Hb and Met-Hb inhibited the 12-LO activity but did not denatured completely the Hb, suggesting that the inhibition was a direct interaction between 12-LO and Hb protein chain and was not due to competition between 12-LO and Hb for oxygen. This study characterizes bullfrog 12-LO with respect to stability, optimal pH, suicide inactivation and interaction with Hb and provides important evolutionary information about this enzyme. PMID- 11281274 TI - Characterisation of red and white muscle myosin heavy chain gene coding sequences from antarctic and tropical fish. AB - To understand molecular adaptation for locomotion at different environmental temperatures, we have studied the myosin heavy chain genes as these encode the molecular motors involved. For this purpose, cDNA libraries from white (fast) and red (slow) myotomal muscle of an Antarctic and a tropical fish were constructed and from these different myosin heavy chain cDNAs were isolated. Northern and in situ hybridisation confirmed in which type of muscle these isoform genes are expressed. The cDNAs were sequenced and the structure of the ATPase sites compared. There was a marked similarity between the tropical fast myosin and the Antarctic slow myosin in the loop 1 region, which has similar amino acid side chains, charge distribution and conformation. These findings help to explain why the myofibrils isolated from white muscle of tropical fish show a lower specific ATPase activity than the white muscle of Antarctic fish but a similar activity to the Antarctic red (slow) muscle. It also provides insight into the way molecular motors in Antarctic fish have evolved to produce more power and thus ensure effective swimming at near zero temperatures by the substitution or addition of a few residues in strategic regions, which include the ATPase site. PMID- 11281275 TI - A branch site mutation leading to aberrant splicing of the human tyrosine hydroxylase gene in a child with a severe extrapyramidal movement disorder. AB - We report a branch site mutation in the gene of the enzyme tyrosine hydroxylase (TH): a -24t > a substitution two bases upstream of the adenosine in the branchpoint sequence (BPS) of intron 11. As normal lariat formation is therefore prevented, alternative splicing takes place; use of the BPS of intron 12 results in skipping of exon 12, whereas the use of a cryptic branch site in intron 11 leads to partial retention of this intron in the mRNA. This leads in both cases to an aberrant protein product. In the one case, skipping of exon 12 results in the absence of 32 amino acids. In the other, retention of 36 nucleotides of intron 11 in the mRNA results in the incorporation of twelve additional amino acids. The functional consequences of this mutation for the patient, who is also heterozygous for another previously identified mutation, become apparent in a severe clinical phenotype. PMID- 11281276 TI - The use of long PCR to confirm three common alleles at the CYP2A6 locus and the relationship between genotype and smoking habit. AB - Long PCR followed by nested PCR has previously been used to determine CYP2A6 160H alleles, but the method proved unreliable. We have optimized this approach in a DNA bank of 1032 subjects (age range 59-74 years) to give reliable results, yielding indirect molecular evidence and very strong statistical evidence of hitherto unrecognized common alleles (designated O) recalcitrant to the long PCR. Coding three alleles (160L, 160H and O) and an approach to association analysis originally developed to deal with null alleles implicit in ABO blood group phenotyping, the contribution of 160H (functionally null) to reduced smoking habit has been clearly measured for the first time, unconfounded by alleles null to the long PCR. The most significant findings (p < 0.01) are that the possession of a 160H allele, compared with not possessing a 160H allele, is associated with a mean age of starting regular smoking 3 years later (95% CI +/- 1.93 years, average start age 20-21 years rather than 17-18 years); and that the average likelihood of quitting smoking at any time is 1.75 fold (95% CI, 1.17-2.61) for those possessing an 160H allele compared with those who have no 160H allele. This suggests that a smoking subject with a genotype predicted to confer 50% of the ability to eliminate nicotine via the CYP2A6 pathway has almost twice the likelihood of quitting smoking. PMID- 11281277 TI - Association of the G-2548A polymorphism in the 5' region of the LEP gene with overweight. AB - Mutations in the translated part of the leptin gene (LEP) have been found in only two families. Nevertheless DNA polymorphisms in the LEP region are linked to extreme obesity. We previously found in the 5' region of LEP a polymorphism, G 2548A, associated with a differerce in BMI reduction following a low calorie diet in overweight women. Recently, this polymorphism was associated with extreme obesity in women. In this work, we genotyped a new sample from the general population including 314 normal weight (BMI < 27 kg/m2) and 109 overweight subjects (BMI > or = 27 kg/m2). The genotype and allele frequencies were significantly different between groups, with the G-2548 allele being more frequent in the overweight subjects (p < 0.01). In men, carriers of this allele had lower leptin concentrations adjusted for fat mass (p = 0.05). Our results indicate that variations at the leptin locus are associated with common obesity phenotypes, and not only with extreme obesity or the rare mendelian obesity syndromes. PMID- 11281278 TI - Patterns of male-specific inter-population divergence in Europe, West Asia and North Africa. AB - We typed 1801 males from 55 locations for the Y-specific binary markers YAP, DYZ3, SRY10831 and the (CA)n microsatellites YCAII and DYS413. Phylogenetic relationships of chromosomes with the same binary haplotype were condensed in seven large one-step networks, which accounted for 95% of all chromosomes. Their coalescence ages were estimated based on microsatellite diversity. The three largest and oldest networks undergo sharp frequency changes in three areas. The more recent network 3.1A clearly discriminates between Western and Eastern European populations. Pairwise Fst showed an overall increment with increasing geographic distance but with a slope greatly reduced when compared to previous reports. By sectioning the entire data set according to geographic and linguistic criteria, we found higher Fst-on-distance slopes within Europe than in West Asia or across the two continents. PMID- 11281279 TI - Selecting SNPs in two-stage analysis of disease association data: a model-free approach. AB - For large numbers of marker loci in a genomic scan for disease loci, we propose a novel 2-stage approach for linkage or association analysis. The two stages are (1) selection of a subset of markers that are 'important' for the trait studied, and (2) modelling interactions among markers and between markers and trait. Here we focus on stage 1 and develop a selection method based on a 2-level nested bootstrap procedure. The method is applied to single nucleotide polymorphisms (SNPs) data in a cohort study of heart disease patients. Out of the 89 original SNPs the method selects 11 markers as being 'important'. Conventional backward stepwise logistic regression on the 89 SNPs selects 7 markers, which are a subset of the 11 markers chosen by our method. PMID- 11281280 TI - Evaluating linkage and linkage disequilibrium: use of excess sharing and transmission disequilibrium methods in affected sib pairs. AB - Two popular and robust approaches to analysing affected sib pair (ASP) data for linkage are the traditional excess sharing methods and the transmission/disequilibrium test (TDT). Here we derive an overall test of linkage for multi-allelic ASP marker data which comprises two component tests: one for excess sharing and one for transmission disequilibrium. This method has several advantages. Firstly the overall test of linkage is often more powerful than either of the two component tests. Secondly the method makes it possible to determine the contribution of linkage disequilibrium (LD), in addition to linkage, to an overall positive linkage result. This is useful because the presence of LD in addition to linkage may suggest that the marker locus is in very close proximity to a disease susceptibility gene. Thirdly the method provides estimates of the risk associated with transmission of the different marker alleles. PMID- 11281282 TI - On power and efficiency robust linkage tests for affected sibs. AB - For diseases that do not follow a clear Mendelian pattern of inheritance nonparametric tests applied to affected sibs have been shown to be robust to the inherent uncertainty about the precise underlying genetic model. It is known that the weights optimizing the power of tests using IBD alleles shared by affected sib pairs or triples depend on the underlying model. We show how efficiency robustness techniques, used in other areas of statistics, provide a systematic approach for constructing a robust linear combination of the statistics that are optimal for the individual members of a family of plausible genetic models. The method depends on the correlation matrix of the optimal tests as these correlations reflect how different the models are. When the minimal correlation is less than 0.5, an alternate robust procedure is proposed. The methods apply to combining data from sibships of different sizes. PMID- 11281281 TI - The triangle test statistic (TTS): a test of genetic homogeneity using departure from the triangle constraints in IBD distribution among affected sib-pairs. AB - The proportions of affected sibs sharing 2, 1 or 0 identical by descent parental marker alleles have been shown to conform to the 'triangle constraints' (Suarez, 1978; Holmans, 1993). It has also been shown (Dudoit & Speed, 1999) that the constraints are verified provided certain assumptions hold. In this study we explore a realistic situation in which the constraints fail due to the presence of a factor in which the sibs differ, a factor on which penetrance depends. This factor may be a characteristic of the trait (severe vs. mild form), or the presence/absence of an associated trait or an environmental factor. We show that under such situations, using the triangle constraints may lead to important loss of power to detect linkage by the MLS test. We propose here an alternative approach in order to detect both linkage and heterogeneity. PMID- 11281283 TI - Phenylarsine oxide inhibits nitric oxide synthase in pulmonary artery endothelial cells. AB - The role of protein tyrosine phosphorylation during regulation of NO synthase (eNOS) activity in endothelial cells is poorly understood. Studies to define this role have used inhibitors of tyrosine kinase or tyrosine phosphatase (TP). Phenylarsine oxide (PAO), an inhibitor of TP, has been reported to bind thiol groups, and recent work from our laboratory demonstrates that eNOS activity depends on thiol groups at its catalytic site. Therefore, we hypothesized that PAO may have a direct effect on eNOS activity. To test this, we measured (i) TP and eNOS activities both in total membrane fractions and in purified eNOS prepared from porcine pulmonary artery endothelial cells and (ii) sulfhydryl content and eNOS activity in purified bovine aortic eNOS expressed in Escherichia coli. High TP activity was detected in total membrane fractions, but no TP activity was detected in purified eNOS fractions. PAO caused a dose-dependent decrease in eNOS activity in total membrane and in purified eNOS fractions from porcine pulmonary artery endothelial cells, even though the latter had no detectable TP activity. PAO also caused a decrease in sulfhydryl content and eNOS activity in purified bovine eNOS. The reduction in eNOS sulfhydryl content and the inhibitory effect of PAO on eNOS activity were prevented by dithiothreitol, a disulfide-reducing agent. These results indicate that (i) PAO directly inhibits eNOS activity in endothelial cells by binding to thiol groups in the eNOS protein and (ii) results of studies using PAO to assess the role of protein tyrosine phosphorylation in regulating eNOS activity must be interpreted with great caution. PMID- 11281284 TI - Nitric oxide stimulates tyrosine phosphorylation of focal adhesion kinase, Src kinase, and mitogen-activated protein kinases in murine fibroblasts. AB - Nitric oxide (NO) can participate in cellular signaling. In this study, monoclonal antibodies against proteins from the growth factor-mediated signalling pathway were used to identify a set of 126-, 56-, 43-, and 40-kDa proteins phosphorylated on tyrosine at NO stimulation of murine fibroblasts overexpressing the human epidermal growth factor receptor. The band corresponding to the 126-kDa protein was FAK. The 56-kDa protein was Src kinase, and the doublet 43- and 40 kDa protein corresponded to the extracellular-regulated MAP kinases (ERK1/ERK2). The effects of NO on focal adhesion complexes were also investigated. FAK was constitutively associated with the adapter protein Grb2 in HER14 cells. Treatment of the cells with the NO donor, sodium nitroprusside, or with EGF did not change this association. We also detected a basal constitutive association of Src kinase with FAK in HER14 cells. In NO-treated cells, this association was stimulated. The doublet 43/40-kDa protein was identical to the ERK1/ERK2 MAP kinases. NO stimulated an increase in ERK1/ERK2 phosphorylation as assessed by a shift in its eletrophoretic mobility and by increased phosphotyrosine immunoreactivity. Furthermore, NO-dependent activation of ERK1/ERK2 depended on the intracellular redox status. Inhibition of glutathione synthesis was necessary to promote activation of the kinases. PMID- 11281285 TI - The biosynthesis of erythroascorbate in Saccharomyces cerevisiae and its role as an antioxidant. AB - This study investigated the ability of the yeast Saccharomyces cerevisiae to synthesize ascorbate and its 5-carbon analogue erythroascorbate from a variety of precursors, and their importance as antioxidants in this organism. Studies of ascorbate and analogues in micro-organisms have been reported previously, but their function as antioxidants have been largely ignored. Ascorbate and erythroascorbate concentrations in yeast extracts were measured spectrophotometrically, and their levels and identity were checked using liquid chromatography-electrospray mass spectrometry. The yeast was readily able to synthesize ascorbate from L-galactono-1,4-lactone or erythroascorbate from D arabinose and D-arabino-1,4-lactone, whereas L-gulono-1,4-lactone was a much poorer substrate for ascorbate biosynthesis. In untreated cells, the concentration of ascorbate-like compounds was below the level of detection of the methods of analysis used in this study (approximately 0.1 mM). Intracellular ascorbate and erythroascorbate were oxidized at high concentrations of tert butylhydroperoxide, but not hydrogen peroxide. Their synthesis was not increased in response to low levels of stress, however, and preloading with erythroascorbate did not protect glutathione levels during oxidative stress. This study provides new information on the metabolism of ascorbate and erythroascorbate in S. cerevisiae, and suggests that erythroascorbate is of limited importance as an antioxidant in S. cerevisiae. PMID- 11281286 TI - Increased oxidative modification of albumin when illuminated in vitro in the presence of a common sunscreen ingredient: protection by nitroxide radicals. AB - We previously reported on the ability of dibenzoylmethane (DBM) and a relative, Parsol 1789, used as a ultraviolet A (UVA)-absorbing sunscreen, to generate free radicals upon illumination, and as a consequence, to inflict strand breaks in plasmid DNA in vitro. This study has now been extended to determine the effects of Parsol 1789 and DBM on proteins, under UVA illumination, with the sole purpose of gaining more knowledge on the photobiological effects of sunscreen chemicals. Parsol 1789 (100 microM) caused a 2-fold increase in protein carbonyl formation (an index of oxidative damage) in bovine serum albumin (BSA) when exposed to illumination, and this damage was both concentration- and time-dependent. The degree of protein damage was markedly reduced by the presence of free radical scavengers, namely piperidinic and indolinonic nitroxide radicals, in accordance with our previous study. Vitamin E had no effect under the conditions used. The results obtained corroborate the fact that Parsol 1789 generates free radicals upon illumination and that these are, most probably, responsible for the protein damage observed under the conditions used in our system. However, at present, we cannot extrapolate from these results the relevance to human use of sunscreens; therefore, further studies should be necessary to determine the efficacy at the molecular and cellular level of this UVA-absorber in order to ascertain protection against photocarcinogenic risk. PMID- 11281287 TI - Alpha-tocopherol downregulates the expression of GPIIb promoter in HEL cells. AB - Alpha-tocopherol is known to inhibit platelet aggregation but the mechanism responsible for this has not been elucidated. Glycoprotein IIb (GPIIb) is the alpha-subunit of the platelet membrane protein GPIIb/IIIa, which functions as a specific receptor for platelet aggregation. Human erythroleukemia (HEL) cells are megakaryocytelike and express the megakaryocyte-specific GPIIb gene. To understand the molecular mechanism of alpha-tocopherol on antiaggregation, we analyzed the effect of physiologically relevant concentrations of alpha tocopherol on the expression of human GPIIb promoter in HEL cells. The enhancement of tetradecanoylphoerbol-12,13-acetate (TPA)-mediated transient and optimal expression of plasmids was achieved by adding 10(-7)-M TPA in the medium 24 h before lipofection. Transient expression of GPIIb promoter was determined in transfected cells pretreated with various concentrations of alpha-tocopherol. Our data shows that the GPIIb promoter activity was downregulated to 55, 23, 27, 20, and 15% in the presence of 10, 20, 40, 80, and 120 microg/ml of alpha-tocopherol, respectively, as compared with that in the absence of alpha-tocopherol. The downregulation of alpha-tocopherol on the TPA-mediated GPIIb promoter activity may result in a reduction of GPIIb protein expression and thus contribute to antiplatelet aggregation. PMID- 11281288 TI - Lysosomal destabilization during macrophage damage induced by cholesterol oxidation products. AB - We have previously shown that oxidized low-density lipoprotein (LDL) induces damage to the macrophage lysosomal membranes, with ensuing leakage of lysosomal contents and macrophage cell death. Cholesterol oxidation products (ChOx) have been reported to be the major cytotoxic components of oxidized LDL/LDL- and also to stimulate cholesterol accumulation in vascular cells. In the present study, we characterized the initial events during macrophage damage induced by cholesterol oxidation products (ChOx). Within 24 h of exposure, ChOx caused lysosomal destabilization, release to the cytosol of the lysosomal marker-enzyme cathepsin D, apoptosis, and postapoptotic necrosis. Enhanced autophagocytosis and chromatin margination was found 12 h after the exposure to ChOx, whereas apoptosis and postapoptotic necrosis was pronounced 24 and 48 h after the exposure. Some lysosomal vacuoles were then filled with degraded cellular organelles, indicating phagocytosis of apoptotic bodies by surviving cells. Because caspase-3 activation was detected in the ChOx-exposed cells, lysosomal destabilization may associate with the leakage of lysosomal enzymes, and activation of the caspase cascade. MnSOD mRNA levels were markedly increased after 24 h of exposure to ChOx, suggesting associated induction of mitochondrial protection repair or turnover. We conclude that ChOx-induced damage to lysosomes and mitochondria are sequelae to the cascade of oxysterol cytotoxic events. The early disruption of lysosomes induced by ChOx, with resultant autophagocytosis may be a critical event in apoptosis and/or necrosis of macrophages/foam cells during the development of atherosclerotic lesions. PMID- 11281289 TI - Pine bark extract pycnogenol downregulates IFN-gamma-induced adhesion of T cells to human keratinocytes by inhibiting inducible ICAM-1 expression. AB - Expression of intercellular adhesion molecule-1 (ICAM-1) is necessary for leukocyte/keratinocyte interactions. Upregulation of ICAM-1 expression in keratinocytes has been observed in several inflammatory dermatoses, such as psoriasis, atopic dermatitis, and lupus erythematosus. Inflammatory cytokines, such as interferon-gamma (IFN-gamma), upregulate ICAM-1 expression in keratinocytes. Because of potent antioxidant and anti-inflammatory properties of the French maritime pine bark extract, Pycnogenol (Horphag Research, Geneva, Switzerland), its effects were investigated on the interaction of T cells with keratinocytes after activation with IFN-gamma and the molecular mechanisms involved in such interactions. Studies were performed using a human keratinocyte cell line, HaCaT. Cell adhesion in the presence of IFN-gamma was studied using a coculture assay. Treatment of HaCaT cells with 20 U/ml IFN-gamma for 24 h markedly induced adherence of Jurkat T cells to HaCaT cells. PYC pretreatment (50 microg/ml, 12 h) significantly inhibited IFN-gamma induced adherence of T cells to HaCaT cells (p < .01). ICAM-1 plays a major role in the IFN-gamma-induced adherence of T cells to keratinocytes. Thus, the effect of PYC on IFN-gamma induced ICAM-1 expression was investigated as well. Pretreatment of HaCaT cells with PYC significantly inhibited IFN-gamma-induced expression of ICAM-1 expression in HaCaT cells. The downregulation of inducible ICAM-1 expression by PYC was both dose and time dependent. A 50 microg/ml dose of PYC and a 12 h pretreatment time (i.e., before activation with IFN-gamma) provided maximal (approximately 70%) inhibition of inducible ICAM-1 expression in HaCaT cells. Gamma-activated sequence present on the ICAM-1 gene confers IFN-gamma responsiveness in selected cells of epithelial origin (e.g., keratinocytes) that are known to express ICAM-1 on activation with IFN-gamma. Gel-shift assays revealed that PYC inhibits IFN-gamma-mediated activation of Stat1, thus suggesting a transcriptional regulation of inducible ICAM-1 expression by PYC. These results indicate the therapeutic potential of PYC in patients with inflammatory skin disorders. PMID- 11281290 TI - Beta-carotene antagonizes the effects of eicosapentaenoic acid on cell growth and lipid peroxidation in WiDr adenocarcinoma cells. AB - The effects of combinations between eicosapentaenoic acid (EPA) and beta-carotene on cell growth and lipid peroxidation were investigated in human WiDr colon adenocarcinoma cells. EPA alone was able to inhibit the growth of WiDr cells in a dose- and time-dependent manner. Such an inhibition involved fatty acid peroxidation, as shown by the remarkable increase in the levels of Malondialdehyde (MDA) in EPA-treated cells. Beta-carotene was capable of reducing the growth inhibitory effects of EPA and the levels of MDA in a dose- and a time dependent manner. In addition, EPA increased beta-carotene consumption in WiDr cells. This study provides evidence that beta-carotene can antagonize the effects of EPA on colon cancer cell growth and lipid peroxidation. PMID- 11281291 TI - Increased mitochondrial superoxide generation in neurons from trisomy 16 mice: a model of Down's syndrome. AB - Increased neuronal cell death in neurodegenerative diseases has been suggested to result from an increased mitochondrial generation of radical oxygen species (ROS). To test this hypothesis, we investigated superoxide formation in cultured hippocampal neurons from diploid and trisomy 16 mice (Ts16), a model of Down's syndrome. Microflurometric techniques were used to measure superoxide-induced oxidation rate of hydroethidine (HEt) to ethidium and reduced nicotinamide adenine dinucleotide (NADH) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) autofluorescence signal to monitor changes in neuronal energy metabolism. We found an increase in superoxide formation by more than 50% in Ts16 neurons in comparison with diploid control neurons. In the presence of the mitochondrial respiratory chain complex I inhibitor rotenone superoxide production was blocked in diploid neurons, but the increased superoxide generation in Ts16 neurons remained. Uncoupling of mitochondrial oxidative phosphorylation using carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) caused irreversible deficiency in the energy metabolism, monitored by NAD(P)H autofluorescence in Ts16 neurons, but not in diploid control neurons. These results suggest an increased basal generation of superoxide in Ts16 neurons, probably caused by a deficient complex I of mitochondrial electron transport chain, which leads to an impaired mitochondrial energy metabolism and finally neuronal cell death. PMID- 11281292 TI - Oxidative stress and adenine nucleotide control of mitochondrial permeability transition. AB - Mitochondria can initiate apoptosis by releasing cytochrome c after undergoing a calcium-dependent permeability transition (MPT). Although the MPT is enhanced by oxidative stress and prevented by adenine nucleotides such as adenosine 5' diphosphate (ADP), the hypothesis has not been tested that oxidants regulate the effects of exogenous adenine nucleotides on the MPT and cytochrome c release. We found that cytochrome c release from intact rat liver mitochondria depended strictly on pore opening and not on membrane potential, and that MPT-enhancing oxidative stress also augmented cytochrome c release. At low oxidative stress, micromolar (ADP) and low adenosine 5'-triphosphate (ATP)/ADP ratio inhibited the MPT and cytochrome c release, whereas ATP or high ATP/ADP had only a slight effect. In freshly isolated mitochondria, the time to half-maximal MPT was related to the log of the ATP/ADP ratio. This function was shifted to shorter times by oxidative stress which decreased ADP protection and caused ATP to accelerate the calcium-dependent MPT. By comparison, mitochondria treated with reducing agents and those isolated from septic rats were protected from the MPT by both nucleotides. These results indicate that oxidation-sensitive site(s) in the membrane regulate the effects of adenine nucleotides on the MPT. The oxidant based differences in the effects of ADP and ATP on the pore support the novel hypothesis that failure of the cell to consume ATP and provide adequate ADP at the adenine nucleotide transporter during oxidative stress predisposes to cytochrome c release and initiation of apoptosis. PMID- 11281293 TI - Isolation and identification of a phenolic antioxidant from Aloe barbadensis. AB - A potent antioxidative compound has been isolated from a methanolic extract of Aloe barbadensis Miller using a combination of column and thin-layer chromatography. The antioxidant activity of this substance was similar to that of alpha-tocopherol as assessed in vitro using rat brain homogenates. On the basis of electrospray ionization and electron-impact ionization mass spectra in combination with reversed-phase, high-performance liquid chromatographic behavior, this compound has been identified as 8-C-beta-D-[2-O-(E) coumaroyl]glucopyranosyl-2-[2-hydroxy]-propyl-7-methoxy-5-methylchromone. PMID- 11281294 TI - Isolation, purification, and characterization of a rat liver mitochondrial protein disulfide isomerase. AB - The isolation and purification to electrophoretical homogeneity and characterization of a protein disulfide isomerase from rat liver mitochondria is reported. The purified enzyme exhibits a single band on sodium dodecylsulfatepolyacrylamide gel electrophoresis with an apparent molecular weight of approximately 54 kDa. Comparatively, the microsomal form shows an apparent molecular weight of 57 kDa indicating that the two forms are slightly different. The antibody raised against the microsomal isoform does not recognize the mitochondrial enzyme. To characterize the enzyme, different classical methodologies utilized for protein disulfide isomerase estimation have been adopted. The isolated enzyme is active with all of them, indicating that it comprises all the features of a typical protein disulfide isomerase. At the mitochondrial level the enzyme appears mostly localized at the membrane level. Its potential involvement in mitochondrial membrane permeability control is also discussed. PMID- 11281295 TI - Mitochondrial permeability transition induced by 1-hydroxyethyl radical. AB - Impairment of mitochondrial functions has been found in ethanol-induced liver injury. Ethanol can be oxidized to the 1-hydroxyethyl radical (HER) by rat liver microsomal systems. Experiments were carried out to evaluate the ability of HER to cause mitochondrial swelling as an indicator of the mitochondrial permeability transition (MPT). Electron spin resonance (ESR) spectroscopy was used to detect HER and to study its interaction with mitochondria. The ESR signal intensity of the spin adduct formed from alpha-(4-pyridyl-1-oxide) N-tert-butylnitrone (POBN) and HER generated from either a thermic decomposition of 1,1'-dihydroxyazoethane (DHAE) or a Fenton reaction system containing ethanol was markedly diminished by the addition of mitochondria, indicating an interaction between HER and mitochondria. Exposure of rat liver mitochondria to HER generated from either system caused swelling, as reflected by a decrease in absorbance at 540 nm, in a HER concentration-dependent and a cyclosporin A-sensitive manner. Mitochondrial swelling was also induced in the Fenton reaction system without ethanol. The DHAE dependent generation of HER in mitochondrial suspension resulted in a decrease of membrane protein thiols and collapse of the membrane potential (delta psi). The swelling induced by HER was prevented by glutathione and vitamin E, but not by superoxide dismutase. Catalase did not prevent the swelling caused by the acetaldehyde/hydroxylamine O-sulfonate (HOS) system, but was inhibitory in the Fenton reaction system with or without ethanol. These results indicate that HER, as well as hydroxyl radical, can induce the MPT, and suggest the possibility that the collapse of delta psi caused by HER may, at least in part, contribute to impairment of mitochondrial function caused by ethanol and in ethanol-induced liver injury. PMID- 11281296 TI - Polynucleotide degradation during early stage response to oxidative stress is specific to mitochondria. AB - Oxidative stress is known to modulate RNA expression in both prokaryotic and eukaryotic cells. We have previously determined that a preferential and calcium dependent downregulation of mitochondrial RNA occurs in HA-1 hamster fibroblasts in response to hydrogen peroxide, and that this is accompanied by the degradation of mitochondrial genomic DNA. Here we extended these studies to determine whether downregulation is specific to transcripts derived from mitochondrial-encoded genes; to determine whether genomic DNA degradation occurs in the nucleus; and to compare overall polynucleotide stress response with cellular growth arrest and apoptosis. We observed that nuclear genome-encoded mRNAs whose protein products are targeted for the electron transport chain of mitochondria were not degraded. Furthermore, early stage degradation of genomic DNA, assessed within the first 5 h of peroxide exposure, was specific to mitochondria, as nuclear genomic DNA was not degraded under the same treatment conditions. These differential degradations occurred under conditions where extensive growth-arrest and moderate apoptosis were observed, and were accompanied by significant induction of the growth arrest mRNAs gadd45, gadd153, and adapt15/gadd7. Combined, these results indicate that there is a general degradation of mitochondrial- but not nuclear-polynucleotides during early stage response of HA-1 fibroblasts to oxidative stress. PMID- 11281298 TI - Oxygen '99 shatters attendance records: 650 attend New Orleans meeting. PMID- 11281297 TI - The heme synthesis and degradation pathways: role in oxidant sensitivity. Heme oxygenase has both pro- and antioxidant properties. AB - The heme biosynthetic and catabolic pathways generate pro- and antioxidant compounds, and consequently, influence cellular sensitivity to oxidants. Heme precursors (delta-aminolevulinic acid, porphyrins) generate reactive oxygen species (ROS), from autoxidation and photochemical reactions, respectively. Heme, an essential iron chelate, serves in respiration, oxygen transport, detoxification, and signal transduction processes. The potential toxicity of heme and hemoproteins points to a critical role for heme degradation in cellular metabolism. The heme oxygenases (HOs) provide this function and participate in cellular defense. This hypothesis emerges from the observation that the activation of HO-1 is an ubiquitous cellular response to oxidative stress. The reaction products of HO activity, biliverdin, and its subsequent metabolite bilirubin, have antioxidant properties. Furthermore, iron released from HO activity stimulates ferritin synthesis, which ultimately provides an iron detoxification mechanism that may account for long-term cytoprotection observed after HO induction. However, such models have overlooked potential pro-oxidant consequences of HO activity. The HO reaction releases iron, which could be involved in deleterious reactions that compete with iron reutilization and sequestration pathways. Indeed, the induction of HO activity may have both pro- and antioxidant sequelae depending on cellular redox potential, and the metabolic fate of the heme iron. PMID- 11281299 TI - Role of cytokines in the pathogenesis of restenosis after percutaneous transluminal coronary angioplasty. AB - BACKGROUND: Inflammatory cytokines play an important role in mediating inflammatory/proliferative responses including atherosclerosis. However, their role in the pathogenesis of restenosis after percutaneous transluminal coronary angioplasty (PTCA) remains to be clarified. OBJECTIVE: To determine plasma levels of inflammatory cytokines as well as cytokine-generation capacities of monocytes before PTCA and after the follow-up period. METHODS: Plasma levels of cytokines in 34 consecutive patients before and 3-6 months after PTCA were measured by enzyme-linked immunosorbent assay. We measured the plasma levels of macrophage colony-stimulating factor (MCSF) and transforming growth factor-beta. Cytokine generation capacities of monocytes were also measured by a whole-blood induction method with lipopolysaccharide. The levels of cytokines measured for assessment of the capacities included those of interleukin-1alpha, interleukin-1beta, interleukin-6, granulocyte-colony-stimulating factor, tumor necrosis factor-alpha and interferon-gamma. RESULTS: Plasma levels of MCSF in patients without restenosis (n = 20) decreased significantly (from 1460+/-138 microg/ml before PTCA to 1039+/-125 microg/ml after the follow-up period, P < 0.01), whereas those in patients with restenosis (n = 14) increased significantly (from 1107+/-105 microg/ml before PTCA to 1039+/-125 microg/ml after the follow-up period, P < 0.05). We noted a positive correlation between the increase in plasma levels of MCSF and the extent of loss of lumen by restenosis. Cytokine-generation capacities of monocytes for interleukin-1alpha and interleukin-1beta of patients with restenosis significantly increased but those of patients without restenosis did not. Furthermore, plasma levels of C-reactive protein decreased significantly only in patients without restenosis after the follow-up period. CONCLUSIONS: These results suggest that inflammatory changes mediated by cytokines may be involved in the pathogenesis of restenosis after PTCA. PMID- 11281300 TI - On the wide spectrum of abnormalities in the coronary arteries of Whipple's disease. AB - BACKGROUND: There is growing interest in the role of microbes in the pathogenesis of coronary atherosclerosis but most of the evidence has been seroepidemiologic. It would be useful to know more about the cytology and histology of coronary lesions containing clearly depicted microbes. OBJECTIVE: To define carefully the assorted abnormalities apparent in the coronary arteries of individuals dying with Whipple's disease. METHODS: Myocardial tissue from 12 cases of Whipple's disease was studied by light microscopy. Slides were stained routinely (in sequence) with either the periodic-acid-Schiff (PAS) or Goldner-trichrome method and some with Gomori methenimine silver. Cardiac slides with PAS-positive bacilli were compared to lesions in jejunal lamina propria. RESULTS: There were abundant sites of coronary arterial damage associated with presence of Whipple bacilli, more in the tunica media than in intima and adventitia. Bacilli in the arterial lesions were identical to those in lamina propria. Medial lesions were often associated with a fibroproliferative 'atheroma'. Both intracellular and extracellular bacilli were found. Most lesions were devoid of inflammation, but some sites exhibited either florid arteritis or dense scarring. Arteries that were scarred or inflamed exhibited only a few bacilli. There was an apparent affinity of bacilli for the nuclei in medial smooth muscle cells and in nearby ventricular myocytes. Apoptosis (TUNEL-positive) was present in medial smooth muscle cells, endothelial cells, and ventricular myocytes. CONCLUSIONS: There is a wide spectrum of coronary abnormalities in Whipple's disease. It would be useful to know how often the Whipple bacillus is a part of the total pathogen burden in coronary disease. PMID- 11281301 TI - Quantitative assessment of diffuse coronary artery disease using a three dimensional reconstruction method compared with intravascular ultrasound images. AB - BACKGROUND: It can be difficult to estimate the degree of stenosis in patients with diffuse coronary artery disease (CAD), because of the lack of a normal reference segment. If the size of normal coronary lumen has a direct relation to size of distal myocardial bed, it could be used to estimate the 'normal' cross sectional area of coronary lumen. Accordingly, we could estimate the degree of stenosis of coronary arteries with diffuse disease by comparing them with calculated 'normal' areas of lumen. OBJECTIVE: To assess the validity of the above hypothesis. METHOD: Fourteen subjects without coronary atherosclerosis (group A) and 16 patients with CAD (group B) underwent simultaneous bidirectional coronary arteriography. Using these coronary arteriograms, we determined the relationship between cross-sectional area of coronary lumen measured by using a computerized edge-detection system and summed distal branch length calculated by using our computerized three-dimensional reconstruction method. RESULTS: For group A, we found a close correlation between area of lumen and branch length (r= 0.948). However, for group B, there were some segments for which the measured area of lumen was clearly smaller than that expected from the relationship for group A. From this relationship for group A, we calculated the stenosis ratios of 22 segments and, to confirm their accuracy, we compared the stenotic ratios with those measured on intravascular ultrasound images. The stenotic ratio of each segment of stenotic coronary artery calculated by our method agreed significantly well with the results obtained from the ultrasound measurements (r= 0.980). CONCLUSIONS: These observations validate a novel approach to quantifying diffuse CAD using clinical arteriograms. PMID- 11281302 TI - Impact of abciximab and coronary stenting on outcomes and costs of percutaneous coronary interventions in a community hospital. AB - OBJECTIVE: To assess costs and outcomes of coronary stenting and balloon angioplasty with and without adjunctive treatment with abciximab for 3758 consecutive elective percutaneous coronary interventions at a single community center over the 2.5-year period between 1 January 1995 and 30 June 1997. RESULTS: Abciximab was more common among patients who had recently suffered myocardial infarction, patients with unstable angina, and patients with more complex coronary lesions. Use of abciximab in conjunction with balloon angioplasty or stenting and stenting alone was associated with significant reductions in incidence of major adverse cardiovascular events in hospital. Multivariate analysis indicated that use of abciximab and stenting were associated with significant independent effects on risk of an event. Hospital costs were increased for patients administered abciximab, treated with stenting, or both. Total costs and costs inclusive of those incurred in catheterization laboratory and pharmacy increased significantly with increasing complexity of lesions. Multivariate regression analysis (baseline cost US$5621) identified death (US$16098), emergency revascularization (US$13678), usage of multiple stents (US$1423 for each stent), and use of abciximab (US$1269) as independent predictors of a greater cost. One-year follow-up revealed significant differences among treatment strategies in terms of risk of need for subsequent revascularization procedures. Lack of stenting but not use of abciximab was identified as a significant predictor of need for repeat revascularization procedures. CONCLUSIONS: Our findings are in general agreement with cost analyses of use of abciximab for populations in clinical trials and suggest that improvements of early clinical outcome with abciximab treatment and stenting justify the incremental cost of treatment in a community hospital setting. PMID- 11281303 TI - The influence of short-term treatment with simvastatin on the inflammatory profile of patients with hypercholesterolaemia. AB - BACKGROUND: Treatment with 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase inhibitors can reduce cardiovascular mortality of patients with atherosclerosis. This effect is probably due not only to a decrease in concentration of cholesterol, but also to non-lipid-involving mechanisms elicited by the action of statin drugs. OBJECTIVE: To investigate the influence of short term therapy with simvastatin on markers of inflammation and oxidation processes in patients with hypercholesterolaemia. DESIGN: We administered 20mg simvastatin daily for 12 weeks to 19 patients with hypercholesterolaemia (250-400 mg/dl). Peripheral blood samples for evaluation of plasma concentrations of thiobarbituric acid reactive substances (malonaldehyde), stable metabolites of nitric oxide (NOx) and interleukin 6 (11-6) were taken before and after the therapy. RESULTS: Plasma levels of malonaldehyde decreased significantly (from 4.533+/-0.428 versus 3.690+/-0.310 micromol/l, P = 0.04) during the study period. Similarly, there was a significant decrease in the plasma concentrations of NOx (from 33.477+/-4.352 micromol/l versus 25.919+/-2.561 micromol/l, P = 0.02). There were significant positive correlations between concentrations of total cholesterol and NOx in plasma (r = 0.4397, P = 0.008) and of low-density lipoprotein and NOx (r = 0.3987, P = 0.02). The plasma level of interleukin 6 remained unchanged by the intervention (1.837+/-0.200 versus 1.820+/-0.169 pg/ml, P = 0.54). CONCLUSIONS: Short-term therapy with simvastatin decreases the plasma concentrations of markers of peroxidation of lipids and of stable metabolites of nitric oxide in hypercholesterolaemic patients, but leaves levels of interleukin 6 unaffected. PMID- 11281304 TI - Diabetes and coronary artery disease--therapy and outcomes. PMID- 11281305 TI - Bibliography current world literature. PMID- 11281306 TI - Homozygosity for the C677-->T mutation of 5,10-methylenetetrahydrofolate reductase and total plasma homocyst(e) ine are not associated with greater than normal risk of a first myocardial infarction in northern Sweden. AB - BACKGROUND: Results of several case-control studies have shown elevated total plasma homocyst(e)ine (TPH) and homozygosity for the point mutation C677-->T in the gene for 5,10-methylenetetrahydrofolate reductase (MTHFR) to be associated with a greater than normal risk of atherosclerotic vascular disease. However, there have been few epidemiologic studies and the interpretation of the results is not clear-cut. OBJECTIVE: To elucidate whether homozygosity for the point mutation C677-->T in the gene for MTHFR, and TPH are risk factors for a first myocardial infarction. DESIGN: A prospective nested case-control study in Northern Sweden. METHODS: Among more than 36000 persons screened, 78 cases satisfied the inclusion criterion of having developed, after sampling, a first myocardial infarction. For each case, two controls matched for sex and age were randomly selected. RESULTS: We found no statistically significant difference among the prevalences of the three possible MTHFR genotypes -/- (no mutation), +/+ (both alleles have the mutation), and +/- among cases and controls in univariate conditional logistic regression analysis. Mean levels of TPH in patients and controls were 12.2+/-4.9 and 12.2+/-3.5 micromol/l (means +/- SD), respectively (NS). CONCLUSIONS: In this study neither homozygosity for the point mutation C677-->T in the gene for MTHFR nor TPH was related to a greater than normal risk of a first myocardial infarction for members of the population of northern Sweden. Further research is needed in order to show whether TPH is an independent risk factor for a first myocardial infarction. PMID- 11281307 TI - Vascular remodeling in atherosclerotic coronary arteries is affected by plaque composition. AB - BACKGROUND: Narrowing of lumen in atherosclerotic lesions is determined not solely by accumulation of plaque but also by constrictive or expansive vascular remodeling. Underlying mechanisms and determinants of these bidirectional processes are not known. OBJECTIVES: To elucidate the response of vascular remodeling to progressive atherosclerosis by analyzing its potential association with composition of plaque. METHODS: Seventy patients with 77 de-novo coronary artery lesions underwent intravascular ultrasound imaging before coronary intervention. Target lesions were defined as soft, fibrous/mixed, and calcified plaques. Quantitative measurements of area of lumen (A(L)), total area of vessel (A(TV)) and area of plaque (A(P) = A(TV)-A(L)) were performed at the lesion site and at the proximal and distal reference sites. Remodeling was determined by using a remodeling index [I(R) = (stenosis of A(TV)/mean reference A(TV)) x 100]. RESULTS: Overall vascular remodeling was balanced with a mean remodeling index of 100.2+/-19.3% and a high interlesion range (60.2-152.4%). The remodeling index for soft lesions was significantly higher than those for fibrous/mixed and calcified lesions (110+/-18.8 versus 96.2+/-14.4 and 85.9+/-15.1%, P < 0.01). Calcified lesions exhibited lower remodeling indexes than did uncalcified lesions (85.9+/-15.1 versus 104.6+/-18.4%, P < 0.01). CONCLUSIONS: Processes involved in vascular remodeling are affected by composition of plaque insofar as there is a higher prevalence of constrictive remodeling among calcified plaques and a higher prevalence of expansive remodeling among soft lesions. These findings indicate that constrictive remodeling is a late manifestation in atherogenesis. Future studies are warranted in order to enhance the understanding of progression of atherosclerosis, and of mechanisms of vascular remodeling and their impacts on interventional therapy. PMID- 11281308 TI - Determinants of the severity and extent of coronary artery disease in patients with type-2 diabetes and in nondiabetic subjects. AB - BACKGROUND: Factors predicting the anatomic distribution and the severity and extent of coronary atherosclerosis in patients with clinically manifest coronary artery disease (CAD) for type-2 diabetic patients could be different than those for nondiabetic patients. OBJECTIVE: To study the determinants of severity and extent of CAD in consecutive patients with type 2 diabetes mellitus, compared with those for matched nondiabetic patients, undergoing clinically indicated coronary angiography. METHODS: Coronary angiograms of 48 men and seven women with type-2 diabetes and an equal number of nondiabetic subjects were analyzed quantitatively. Scores reflecting severity and extent of CAD were compared with potential risk factors using univariate correlation analyses and multivariate regression models. RESULTS: For the diabetics, a global coronary atheroma burden index was independently and directly related to age (P = 0.022) and to level of intermediate-density lipoprotein cholesterol (P = 0.055), and inversely to level of particles of a subtype of high-density lipoprotein (P = 0.022). Several angiographic indexes were related to the duration of diabetes and control of glycemia. For the nondiabetic group, global atheroma burden was independently related to age (P = 0.028), a history of hypertension (P = 0.028), and concentration of low-density lipoprotein (P = 0.013), and inversely to level of apolipoprotein A-I (P = 0.008). The duration of coronary disease and a history of smoking were also predictive of severe coronary atherosclerosis among nondiabetic patients. CONCLUSIONS: Classical risk factors are strong predictors of the severity and extent of coronary atherosclerosis in nondiabetic patients, but the most important determinants for type-2 diabetic patients are levels of triglyceride-rich lipoproteins and apolipoprotein A-I-containing particles of high-density lipoprotein, and factors directly related to diabetes. PMID- 11281309 TI - Memory testing in dementia: how much is enough? AB - Analyses of eight widely used memory measures (Word List Acquisition and Recall used in the Alzheimer's Disease Assessment Scale and the Consortium to Establish a Registry for Alzheimer's Disease neuropsychology battery, Wechsler Memory Scale Revised [WMS-R] Logical Memory I and II, WMS-R Visual Reproduction I and II, the memory scores from the Neurobehavioral Cognitive Status Examination [NCSE], memory scores from the Mini-Mental State Examination [MMSE]), and the MMSE total score showed each to have moderate predictive power in differentiating between patients with mild dementia and healthy normal controls. When these instruments were combined in a logistic regression analysis, three of them had substantial predictive power. Together, the Word List Acquisition, WMS-R Logical Memory II, and WMS-R Visual Reproduction II were 97.26% accurate (100% sensitive and 94.59% specific) in distinguishing these two groups. The Word List Acquisition is a brief test that alone had high accuracy (92%). These memory tests are highly useful in the diagnosis of mild dementia. PMID- 11281310 TI - Apathy, anhedonia, and psychomotor retardation in elderly psychiatric patients and healthy elderly individuals. AB - Normal aging of the brain affects the basal ganglia-thalamocortical circuits. These circuits are implicated in several neuropsychiatric disorders. Normal aging may therefore influence the symptomatology of psychiatric disorders in the elderly. We investigated motivational behavior that is associated with the function of these circuits, such as apathy, anhedonia, and psychomotor retardation in healthy elderly subjects and psychiatric inpatients (age > or = 60 yr). Apathy, anhedonia, and psychomotor retardation were assessed with the Apathy Evaluation Scale, the Snaith-Hamilton Pleasure Scale, and the Widlocher Retardation Rating Scale. Other measurements included the Comprehensive Psychopathological Rating Scale, the Mini-Mental State Examination, and the assessment of vascular risk factors. We found some evidence for age-related changes in motivational behavior. In the healthy elderly group (n = 64), increasing age was associated with anhedonia, and in the patient group (n = 62), increasing age was associated with psychomotor retardation. Motivational disturbances could be the effect of an interaction between brain aging and the neuropathology of psychiatric disorders in the elderly. PMID- 11281311 TI - Worsening of post-traumatic stress disorder symptoms with cognitive decline: case series. AB - We present three cases of post-traumatic stress disorder (PTSD) symptoms associated with cognitive decline. Patient age ranged from 57 to 70 years old and all patients had war-related PTSD. In each case, the patient had a history of PTSD that was under fairly good control until the onset of cognitive impairment due to Alzheimer's disease or vascular or alcohol-related dementia. These cases suggest that neurodegeneration of memory pathways may disinhibit symptoms of PTSD. PMID- 11281312 TI - Functional status and clinical correlates in cognitively impaired community living older people. AB - We describe the prevalence of cognitive impairment in a population of community living older people, its association with functional decline, and degree of comorbidity. In addition, we examined the relationship between different levels of cognitive impairment and mortality. We conducted an observational study of 1787 patients aged 65 years and above with any degree of cognitive impairment. Patient data were collected with the Minimum Data Set for Home Care. More than 50% of patients had some level of cognitive impairment, which correlates with the degree of physical frailty. On the contrary, patients with cognitive impairment appear to have fewer comorbid conditions and are less likely to receive medications than patients with normal cognitive status. In particular, hypertension, congestive heart failure, chronic obstructive pulmonary disease, cancer, diabetes mellitus, and osteoporosis are found more frequently among patients with normal mental status compared with those showing some level of cognitive defects. Yet, more severe cognitive impairment is associated with a higher mortality rate. Demented patients are characterized by a high prevalence of functional disability and by increased mortality. This increased morbidity and mortality rate is associated with a lower prevalence of comorbid clinical conditions and drug use, relative to patients with normal cognitive performance. The present findings support the possibility that severe cognitive impairment has an independent effect on survival. PMID- 11281313 TI - Nortriptyline in geriatric depression resistant to serotonin reuptake inhibitors: case series. AB - Research on treatment-resistant depression in the elderly has been limited, and recommendations for clinical management have often been extrapolated from studies using nongeriatric patients. This report describes a series of 10 elderly patients with refractory depression who were treated with nortriptyline after failing to respond to an adequate trial of a serotonin reuptake inhibitor. Seven (70%) of the patients responded to the addition or substitution of nortriptyline. All seven of the responders have remained on nortriptyline for maintenance therapy, none of whom have experienced recurrence of their depression after an average treatment duration of 1 year. Response to nortriptyline occurred in about 4 weeks in most patients. The mean daily dose of nortriptyline was 54 mg, and the mean plasma level was 97 ng/mL. Minor side effects occurred in three patients. No patients developed significant electrocardiogram changes. Nortriptyline, possibly due to its different mechanism of action, may be effective as either an adjunctive or replacement antidepressant in some cases of geriatric depression that are resistant to serotonin reuptake inhibitors. PMID- 11281314 TI - Performance on the Mattis Dementia Rating Scale in patients with vascular dementia: relationships to neuroimaging findings. AB - Impairment on screening measures such as the Mattis Dementia Rating Scale (MDRS) provides evidence of dementia in patients with cerebrovascular disease. However, the relationships between neuroimaging findings and performance on the MDRS in vascular dementia (VD) have not been determined. In the present study, we examined the relationships between subcortical hyperintensity (SH) volume and whole brain volume (WBV) on the subscales and total score of the MDRS. Results revealed that SH accounted for a significant amount of variance on the Initiation/Perseveration and Construction subscales, whereas WBV accounted for a significant amount of variance on the Memory subscale. The total score on the MDRS was found to be significantly related to WBV but not SH. These results suggest that subcortical damage and brain volume account for different aspects of cognitive decline in VD and that overall cognitive impairment may reflect cortical and subcortical involvement. PMID- 11281315 TI - Feasibility and effectiveness of treatments for post-stroke depression in elderly inpatients: systematic review. AB - To determine the feasibility and effectiveness of antidepressive treatments for post-stroke depression in elderly medical inpatients, MEDLINE was searched for potentially relevant articles published from January 1987 to August 1997 using the keywords "depression or depressive disorder" (exploded) and "aged." Thirteen reports met the following inclusion criteria: (1) published in English or French; (2) minimum age criterion of 55 and over or mean age 65 and over; (3) post-stroke subjects admitted to a medical, geriatric, or rehabilitation service; (4) used accepted criteria for depression; (5) examined treatment(s) for depression; and (6) reported outcomes as a depression diagnosis and/or symptom level. Data were abstracted independently from each article by two reviewers. The limited evidence suggests contraindications to treatment of 83% of a group to receive a heterocyclic antidepressant compared with 11% of a group to receive a selective serotonin reuptake inhibitor (SSRI); rates of discontinuation and study completion are similar for heterocyclics, SSRIs and psychostimulants. All of the treatments appear to be at least modestly effective in the short term. PMID- 11281316 TI - Single photon emission computed tomography and apolipoprotein E in Alzheimer's disease: impact of the epsilon4 allele on regional cerebral blood flow. AB - The aim of this study was to examine the impact of the apolipoprotein E (APOE) epsilon4 allele on semiquantitative regional cerebral blood flow (rCBF) in Alzheimer's disease. Single photon emission computed tomography technetium (SPECT) with (99m)Tc d,l-hexamethyl propylenamine oxine was used to determine rCBF in 41 consecutive patients (18 males/23 females) with probable Alzheimer's disease according to the National Institute of Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association criteria (mean age 71.0 years; range 54-85). The mean Mini-Mental State Examination (MMSE) score was 20.4 (range 10-30). After normalization of CBF to mean blood flow in the cerebellum, values for rCBF in several cortical regions of interest, side-to-side asymmetry indices, and anterior-posterior ratios were calculated. Determination of the APOE genotype from blood samples was performed using restriction enzyme polymerase chain reaction technique. Multivariate regression analyses and the Wilcoxon rank-sum test for unpaired data (Mann Whitney) were used for statistical analysis. The patients comprised 27APOE epsilon4-positive and 14APOE epsilon4-negative individuals. Five patients were APOE epsilon4 homozygotes. APOE epsilon4-positive patients had significantly reduced rCBF in the right frontal and left occipital lobes. On nonparametric analysis, the most prominent differences between epsilon4-negative and epsilon4 positive patients were demonstrated in subregions representing the frontal association cortex (Mann-Whitney, P < .01). Age-stratified analysis suggested that these findings could be demonstrated predominantly in the elderly patients. The results of this study suggest that the APOE genotype in itself may have an impact on the pattern of rCBF deficits in Alzheimer's disease. The more pronounced reduction of rCBF in frontal association cortex observed in elderly APOE epsilon4-positive patients might predict clinical progression. PMID- 11281317 TI - Anxiety and Alzheimer's disease. AB - This study investigated symptoms of anxiety in two samples of clinic outpatients diagnosed with Alzheimer's disease (AD). Clinician and caregiver reports were obtained using standardized measures to characterize a broad array of anxiety symptoms. Anxiety symptoms were reported for a substantial proportion of subjects, regardless of whether clinician or caregiver ratings were used. Anxious or worried appearance was most common (68% to 71%), followed by fearfulness, tension, restlessness, and fidgeting (37% to 57%). Sleep disturbance and various somatic symptoms were less common (8% to 34%). Although anxiety symptoms were prevalent, only 5% to 6% of subjects met Diagnostic and Statistical Manual of Mental Disorders criteria for the diagnosis of generalized anxiety disorder. In both samples, anxiety symptoms were associated with depression, behavioral disturbances, and increased cognitive impairment. Study findings support a high occurrence of anxiety in patients with dementia, and treatments for anxiety might therefore be helpful in reducing the psychiatric burden of AD. PMID- 11281318 TI - Unrecognized dementia in elderly patients admitted to hospital with psychiatric symptoms. AB - In this study, we hypothesized that elderly patients with first admission to a psychiatric hospital commonly suffer from dementia but did not have such a diagnosis on admission. Over a period of 5 years, we surveyed all medical journals from patients treated for the first time at the Department of Psychogeriatrics. By selecting all inpatients treated at the only regional psychiatric hospital within a defined geographic area, the selected patients became representative of this geographic area. After a diagnostic work-up at the hospital, 72 of 239 patients satisfied clinical criteria of dementia. Of these patients, 7 had Alzheimer's disease and 7 had evidence of vascular dementia. We identified 51 of 72 patients (71%) with an endpoint diagnosis of dementia made at the hospital without any clinical information suggesting dementia at the time of hospitalization. Nonspecific psychosis (35%), depression (15%), and behavioral disturbances (8%) represented the most common diagnoses proposed by the referring doctor. In conclusion, dementia may be a difficult diagnosis in elderly patients with psychiatric symptoms. This study reminds clinicians that dementia should be considered in these patients. PMID- 11281319 TI - Biodiversity and horizontal gene transfer in culturable bacteria isolated from activated sludge enriched in nonylphenol ethoxylates. AB - One hundred and twenty bacterial isolates, from activated sludge of a treatment plant collecting wastes enriched in ethoxylated nonylphenols, were studied. Sixty isolates were selected on rich medium and 60 on mineral medium containing two nonylphenol ethoxylates as the sole carbon source. Analysis of biodiversity at the species level was performed by comparing the AluI restriction patterns of the 16S ribosomal DNA amplified by PCR from 120 isolates. The rDNA restriction analysis enabled us to cluster the isolates into 15 groups, five of which represented nearly 77% of the community. Phylogenetic analysis of five strains belonging to these main groups made it possible to assign four of them to the genera Acinetobacter, Aeromonas and Shewanella and one to the Proteus group. The analysis of plasmid content showed a high variability and suggested that horizontal gene transfer had taken place at the intraspecific, interspecific and intergeneric levels. PMID- 11281320 TI - Second-order selection in bacterial evolution: selection acting on mutation and recombination rates in the course of adaptation. AB - The increase in genetic variability of a population can be selected during adaptation, as demonstrated by the selection of mutator alleles. The dynamics of this phenomenon, named second-order selection, can result in an improved adaptability of bacteria through regulation of all facets of mutation and recombination processes. PMID- 11281321 TI - Survival of Escherichia coli during long-term starvation: effects of aeration, NaCl, and the rpoS and osmC gene products. AB - The survival of Escherichia coli was investigated during long-term starvation in rich media. In aerated cultures, E. coli lost the ability to form colonies earlier in NaCl-free Luria broth than in LB medium containing NaCl. Improved survival at low aeration and the sensitivity to hydrogen peroxide in aging cultures indicated a major role for oxidative stress in cell mortality. Mutants in rpoS, lacking the sigmaS subunit of RNA polymerase, showed altered survival in salt-containing media. However, in the absence of NaCl, although these mutants exhibited a massive loss of viability during the first 2 days, this was followed by a stabilization of the number of survivors. The starved culture contained survivors until at least day 9, long after a wild-type strain had completely lost viability. This peculiar behavior suggests that, in rich media of low osmotic pressure, sigmaS helps in short-term survival but hampers long-term survival. Mutants in osmC, a member of the rpoS regulon, also exhibited reduced survival and increased sensitivity to oxidative stress. The biochemical function of the envelope protein OsmC remains unknown, but present data indicated that it participates, directly or indirectly, in the defense against oxidative compounds. PMID- 11281323 TI - Cell division protein FtsZ: running rings around bacteria, chloroplasts and mitochondria. AB - Of all the proteins involved in prokaryotic cell division FtsZ is one of the earliest acting and most widely distributed, being found in all but a few species. We discuss several recent discoveries of FtsZ in eukaryotic cells and the protein's role in the division of chloroplasts and mitochondria, organelles that are of bacterial origin. PMID- 11281322 TI - Sup35p yeast prion-like protein as an adapter for production of the Gag-p55 antigen of HIV-1 and the L-chain of botulinum neurotoxin in Saccharomyces cerevisiae. AB - Effective expression of the HIV-1 core protein Gag-p55 was obtained in Saccharomyces cerevisiae under control of the inducible UASgal/CYC1 promoter as a translational fusion with the prion-forming NM domain of the translation terminator Sup35p (eRF3) of S. cerevisiae. where only poor expression of the original-type Gag-p55 was observed. A deletion within the Sup35NM prion-forming domain altering Sup35-associated [PSI] inheritance did not compromise expression of the Sup35NM Gag-p55 fusion protein. Therefore, either the mechanism of this phenomenon is not directly related to the effect of Sup35p prion-formation or the modified protein maintains residual prion-forming abilities. The recombinant Sup35p-Gag-p55 protein was quite stable under boiling in an alkali/sodium dodecyl sulfate (SDS) solution and completely retained its antigenic properties. Moreover, 10-min boiling of the native yeast cells in this solution allowed immediate inhibition of lysosomal and other yeast proteases, responsible for autolysis of many natural and recombinant proteins. The use of this method of preliminary enrichment for the recombinant fusion protein Sup35p-Gag-p55 with the SDS-alkaline extraction could be useful for yeast heterologous expression and purification of other of insoluble and unstable proteins. A translational fusion with the NM domain of Sup35p was also used to produce another poorly soluble protein, the L-chain of botulinum exotoxin A, in S. cerevisiae. When the Sup35p fragment was removed from the recombinant construct encoding a fused Sup35/BoNT protein, a dramatic drop in both transformation efficiency and growth rate of transformants was shown. PMID- 11281324 TI - Detection and quantification of the iap gene of Listeria monocytogenes and Listeria innocua by a new real-time quantitative PCR assay. AB - A real-time quantitative polymerase chain reaction (PCR) assay for direct detection and enumeration of Listeria monocytogenes and Listeria innocua was developed and applied to artificially contaminated milk samples. The iap gene present in both species was used as a target for amplification of a 175-bp (L. monocytogenes) and a 309-bp (L. innocua) fragment. To ensure that L. monocytogenes and L. innocua are specifically detectable, tests were carried out using 42 L. monocytogenes strains and 33 L. innocua strains belonging to different serovars. Specificity was also confirmed using 22 bacterial strains not belonging to the genus Listeria, including closely related bacteria. In addition to specificity, the reported assay is characterized by a wide dynamic range of quantification and a high sensitivity, as we could detect as few as six copies of the iap gene per PCR using purified DNA as template. When applied to direct detection and quantification of L. monocytogenes in milk, the more rapid real time quantitative PCR assay was as sensitive as the traditional plate count method, but real-time quantitative PCR-derived iap gene copy numbers were one to two logs higher than colony-forming units obtained by the plate count method. PMID- 11281325 TI - Computer identification of Shigella species by rRNA gene restriction patterns. AB - We describe a MluI ribotyping scheme for Shigella which approaches correlation with serotyping. One hundred and seventeen reference strains and previously serotyped clinical isolates representing the 57 Shigella serotypes and biotypes were included in this study. A total of 51 distinct ribotypes were obtained and a database was built with them. The number of bands composing each ribotype varied from 9 to 15. The fragments ranged in size from 1.6 to 18.8 kbp. One hundred and eleven clinical isolates were successfully identified in a double blind study with standard biochemical/serologic methods, by automatic comparison of their ribotypes with our database using the software Taxotron. PMID- 11281326 TI - Interactions between bovine endothelial cells and Pasteurella multocida: association and invasion. AB - We investigated the association and the invasion of a bovine aortic endothelial cell (BAEC) line by Pasteurella multocida to study the potential role of internalized bacteria and possible intracellular survival during Pasteurella infections. Our data indicate that P. multocida is able to adhere to and to invade BAECs. The density of the bacterial population plays a defined role for an optimal mechanism of interaction between bacteria and cells, as does the incubation period of association and invasion. The optimal bacteria/cells ratio was found to be 100/1, while the optimal infection time was approximately 4 h of incubation. Bacterial internalization was dependent on microfilament and microtubule stability. The invasion ability of P. multocida in the presence of cytochalasin D was reduced by 60%; in the presence of colchicine it was reduced by 97% and in the presence of nocodazole it was reduced by 95%. Our data show that internalized P. multocida did not induce mortality of invaded endothelial cells. Some Pasteurella cells were able to survive and undergo exocytosis. PMID- 11281327 TI - Identification by in situ hybridization of segmented filamentous bacteria in the intestine of diarrheic rainbow trout (Oncorhynchus mykiss). AB - Nonculturable segmented filamentous bacteria (SFB) have been described in the gut of rats, mice and chickens, and 16S rRNA sequences for these organisms are available. These organisms, peripherically related to Clostridium phylogenetic group I, have been provisionally named 'Candidatus Arthromitus'. This work reports the observation of similar bacteria in the intestinal content of the distal intestine, preferentially, in the adult rainbow trout (Oncorhynchus mykiss) that exhibited episodic acute diarrhea, usually during the summer. Abdominal distension, intestinal fluid-mucus content and epithelium detachment were observed in trout. The demonstration that the observed microorganisms are bacteria and belong in the 'Candidatus Arthromitus' group was achieved by in situ hybridization with, respectively, a eubacterial probe and an oligonucleotide probe designed to react specifically with SFB 16S rRNA (encoded by the rrs gene) sequences. The sequenced rrs gene was compared with published sequences and found to be closely related to (although distinct from) other SFB sequences. Implication of these bacteria in trout diarrheic illness remains hypothetical. PMID- 11281328 TI - eae-negative attaching and effacing Escherichia coli from piglets with diarrhea. AB - One hundred and ninety strains of Escherichia coli that were isolated from pigs with diarrhea in the state of Sao Paulo, Brazil, and that were negative for enterotoxins and cytotoxins were investigated. Strains which adhered to HeLa cells were examined for fluorescence actin staining (FAS), the ability to induce attaching and effacing (A/E) lesions on HEp-2 cells detectable by transmission electron microscopy and the presence of eae gene sequences detected by PCR. Intimin production was detected by western blot and serogrouping was performed. Forty-seven isolates adhered to HeLa cells in several patterns, but none adhered in a localized adherence pattern. However, seven of the 47 adherent strains were positive for the FAS reaction, although the reactions were usually weak or atypical. One FAS-negative and three FAS-positive strains, which were examined for their ability to induce A/E lesions, were all positive. Subsequently, testing of these strains for the eae gene showed that they all lacked this gene. These findings, along with earlier reports of eae-negative A/E E. coli, suggest that higher quantities of E. coli in this category might be detected if more reliance were placed on phenotypic tests rather than on gene detection tests alone. PMID- 11281329 TI - Biodegradation of hydroxylated and methoxylated benzoic, phenylacetic and phenylpropenoic acids present in olive mill wastewaters by two bacterial strains. AB - Two aerobic bacterial strains, a chlorophenol-degrading bacterium characterized in this work as a Ralstonia sp. LD35 on the basis of the sequence of the gene encoding for 16S ribosomal RNA, and Pseudomonas putida DSM 1868, capable of metabolizing 4-methoxybenzoic acid, were tested for their capacity to degrade monocyclic aromatic acids responsible for the toxicity of olive mill wastewaters (OMWs). Both strains possess interesting and complementary degradation capabilities in resting cell conditions: Ralstonia sp. LD35 was found to metabolize 4-hydroxybenzoic, 4-hydroxyphenylacetic, 3,4-dihydroxycinnamic and cinnamic acid, whereas DSM 1868 was capable of metabolizing 4-hydroxy-3 methoxybenzoic, 3,4-dimethoxybenzoic and 4-hydroxy-3,5-dimethoxybenzoic acid, as well as 4-hydroxybenzoic and 4-hydroxyphenylacetic acid. The kinetic parameters describing the growth of the two strains on the same compounds were determined in growing-cell batch conditions, and showed that both strains presented high affinity and high specific growth rates towards all assayed substrates. In addition, the two strains were capable of growing on and extensively biodegrading a mixture of monocyclic aromatic acids commonly found at high concentrations in OMWs, and of growing on a 20% dilution of a natural OMW. All these features make the two strains attractive candidates for the development of a biotechnological process for the biodegradation of aromatic compounds found in OMWs. PMID- 11281330 TI - Improvement in the RFLP procedure for studying the diversity of nifH genes in communities of nitrogen fixers in soil. AB - Several specific primers for the nifH gene were tested with different pure telluric N2-fixing strains. A PolF/PolR primer set provided successful amplification of 19 representative N2-fixing strains. Three restriction enzymes, HaeIII, NdeII and MnlI, chosen for restriction fragment length polymorphism (RFLP) analyses, were the most discriminating for the study of nifH gene diversity as they resulted in differences between strains at the species level. Amplification by selected primers and RFLP were applied to assess the genetic diversity of the nifH gene pool in soil. Pair soils, one under cultivation, the second under permanent pasture, were found to harbor a contrasting diversity of nifH genes. Pure strain profiles could not be recognized in the nifH soil patterns. Using the simple procedure described, it was shown that the structure of nitrogen fixers in soil was influenced by soil functioning. PMID- 11281331 TI - Walking, chair rising, and stair climbing after total knee arthroplasty: patellar resurfacing versus nonresurfacing. AB - During the past decade, the technology and design of knee joint prostheses has progressed considerably. However, there is still much controversy on whether resurfacing the patella during routine total knee arthroplasty (TKA) is necessary. This study compares the biomechanics of the lower limb in patients after TKA with and without patellar resurfacing during level walking, stair climbing, and chair rising. Eighteen patients who underwent TKA by two different surgeons using the same prosthesis were studied after full rehabilitation while walking, stair climbing, and chair rising. Patients were divided between those who were resurfaced and those who were not resurfaced. An aged-matched control population was recruited for comparison. The Hospital for Special Surgery Knee Rating Scale was used to gather clinical information. Kinematic and kinetic parameters were collected using a 5-camera Motion Analysis System and an AMTI OR6 5 force platform. For level walking, patients were asked to walk at a self selected speed down an 8-m walkway. For stair climbing, patients were asked to climb a 4-step staircase without handrail support and for chair rising, patients were asked to rise from a chair that was positioned at the height of their knee joint line. Five trials for each side were recorded for averaging and statistical analysis. Temporal-spatial parameters and kinematic and kinetic variables at the knee joint were tested for significance using the repeated measures analysis of variance (ANOVA). There were no significant differences in the biomechanics of walking, stair climbing, or chair rising between patients after TKA with and without a resurfaced patella. PMID- 11281332 TI - Reconstructive treatment of posterolateral rotatory instability of the knee: a biomechanical study. AB - Twelve cadaveric knees were tested to determine effective reconstructive treatment for severe chronic posterolateral rotatory knee instability accompanied by excessive varus and posterior laxity. Posterolateral, varus, and posterior laxity were measured, first with the ligaments intact, then after complete sectioning of the posterior cruciate ligament (PCL) and posterolateral structures, and finally after reconstruction of these structures in different orders. The increases in those laxities were produced following the sectioning of all of the structures and disappeared throughout the flexion range after combined reconstruction of the PCL, lateral collateral ligament (LCL), and popliteus tendon. However, some residual increase in the laxity was always observed if any of the three structures were excluded from reconstruction. Therefore, combined reconstruction of the PCL, LCL, and popliteus tendon is essential and adequate for treating severe chronic posterolateral rotatory instability. PMID- 11281333 TI - Localized pigmented villonodular synovitis involving the fat pad of the knee. PMID- 11281334 TI - Operative treatment of combined anterior and posterior cruciate ligament injuries in complex knee trauma: can the cruciate ligaments be preserved? AB - A retrospective study was performed focusing on operative treatment after combined anterior cruciate ligament (ACL)/posterior cruciate ligament (PCL) injuries. The operative treatment included the preservation of one or both cruciate ligaments. Twenty-eight patients, average age 30 years (range: 12-55 years), were evaluated 5.4 years (range: 1-14 years) postoperatively. Twenty-two operations were performed in patients with acute injuries (<30 days after trauma) and 6 operations in patients with chronic instabilities (>30 days after trauma). Both cruciate ligaments were preserved by suture or refixation in 16 patients. Suture of one and reconstruction of the other cruciate ligament with autologous tendon graft was performed in 12 cases. In addition, 61 procedures (meniscal suture/resection, medial/lateral reconstruction, tendon suture, and open reduction and internal fixation were performed. Postoperative treatment included continuous passive motion and protected weight bearing. Eleven (27% acute, 83% chronic) patients required revision (ACL/PCL reconstruction, osteotomy, and meniscal repair). At follow-up, 43% of the patients were very satisfied and 46% were satisfied. Seventy-one percent (89% preinjury) of the patients were able to maintain intensive and moderate International Knee Documentation Committee (IKDC) activity levels. The IKDC evaluation of the patients (acute %/chronic cases %) was graded for symptoms: A 39% (45/17), B 35% (27/67), C 15% (18/0), and D 11% (9/17); for range of motion: A 42% (36/67), B 42% (50/17), C 16% (14/17), and D 0%; and for ligaments: A 21% (18/17), B 33% (45/0), C 42% (32/83), and D 4% (5/0). Radiographic findings were A 18%, B 41%, and C 41%. Primary repair of acute injuries was superior to the delayed repair of chronic instabilities. Preservation of cruciate ligaments in acute combined ACL/PCL tears results in a satisfying knee function despite distinct residual ligament instability. Although suture of the cruciate ligaments in open technique is a therapeutic option in acute multiligamentous knee injuries, it is not recommended for the treatment of chronic instabilities. PMID- 11281335 TI - Radiographic and computed tomographic analysis of the position of the tibial tubercle in recurrent dislocation and subluxation of the patella. AB - Among the factors that influence the patellofemoral alignment and stability, the position of the tibial tubercle was evaluated using plain radiographs and computed tomography (CT). Radiographs of the Q-angle with the knee in full extension and quadriceps muscle relaxed showed that mean Q-angles in the dislocation or subluxation group and control group were 13.8 degrees and 14.3 degrees, respectively. The major reason for this deceptive measurement was a patella positioned laterally in the dislocation or subluxation group. Better estimation was made with the modified Q-angle measurement by using the most concave point of the intercondylar notch as a reference point instead of the center of the patella. By overshadowing CT images of the levels of the patellofemoral joint and the tibial tubercle, it was possible to evaluate the location of the tibial tubercle in relation to the femoral trochlea. Both the angular measurement (lateral deviation angle [LDA]) and the distance measurement (lateral deviation index [LDI]) showed that the tibial tubercle in the dislocation or subluxation group was rotated externally and shifted laterally (LDA, 36.3+/-7.0 degrees and LDI, 30.1+/-5.6) compared with the control group (LDA, 20.2+/-7.1 degrees and LDI, 15.1+/-5.6). Further study is warranted to elucidate the exact mechanism of recurrent patellar dislocation and subluxation. PMID- 11281336 TI - Distal femoral varus osteotomy in the valgus osteoarthritic knee. AB - The results of 18 distal femoral varus osteotomies performed in 18 patients between 1982 and 1993 were evaluated. All patients had degenerative arthritis of the lateral compartment of the knee associated with a valgus deformity. At surgery, the average patient age was 54 years (range: 38-75 years). The average follow-up was 9 years (range: 5-16 years). The average tibiofemoral angle was 17.5 degrees of valgus preoperatively and 6 degrees postoperatively. Seventeen patients (1 patient died from an unrelated cause) were evaluated at follow-up according to the Knee Society rating system. At follow-up, 13 (77%) were rated as good or excellent by the Knee Society rating system. The Knee score improved from 54 points preoperatively to 89 points postoperatively. The functional score improved from 65 points preoperatively to 86 points postoperatively. One knee required a subsequent total knee arthroplasty (TKA) 5 years after osteotomy due to severe and persistent pain. No patient had infection or nonunion. Varus osteotomy of the distal femur is a reliable and effective surgical procedure for the treatment of gonarthrosis associated with valgus deformity in both young and older active patients, where it can be an alternative to TKA. PMID- 11281337 TI - A comparison of crushed ice and continuous flow cold therapy. AB - Crushed ice was compared to continuous flow cold therapy for control of postoperative pain after arthroscopic patellar tendon autograft anterior cruciate ligament (ACL) reconstruction. With all other variables held constant, cold was administered by either continuous flow (group 1) or crushed ice (group 2). The cold therapy was constant for 3 days, then as needed in days 4 through 7. Data were collected by investigator evaluations and patient diaries. Pain was assessed by visual analog scale (VAS) and categorical pain scale (Likert). Eighty-seven patients were included (52 continuous flow and 35 crushed ice). Continuous passive motion averaged 54 hours for group 1 and 43 hours for group 2 (P<.05). Knee motion at one week averaged more (5 degrees/88 degrees) for group 1 (flexion range: 48 degrees-155 degrees) than for group 2 (6 degrees/77 degrees) (flexion range: 25 degrees-125 degrees) (P=.03). Likert pain scores for group 2 patients were always statistically greater than group 1 patients from the first hour through postoperative day 6 (P<.01). The average VAS pain was always greater for group 2 and statistically greater for postoperative day 1 (P<.01). Hydrocodone bitartrate with acetaminophen use in group 2 was greater than in group 1 for postoperative days 1 (P<.001) and 2 (P=.035). The respective cold modality VAS measured performance was 9.1 for group 1 and 7.8 for group 2 (P<.01). During postoperative days 4 through 6, group 1 patients applied their cold modality for 47.9 hours but group 2 patients applied their cold modality for 29.5 hours (P<.01). Compared to crushed ice, continuous flow cold therapy lowered VAS and Likert pain scores more, reduced hydrocodone bitartrate with acetaminophen use, was used more often, increased continuous passive motion, increased 1-week knee flexion, and was given significantly higher performance ratings by patients. Continuous flow cold is superior to crushed ice for outpatient ACL reconstruction pain and should not be considered an equivalent modality. PMID- 11281338 TI - Verapamil versus amlodipine in proteinuric non-diabetic nephropathies treated with trandolapril (VVANNTT study): design of a prospective randomized multicenter trial. AB - Angiotensin converting enzyme inhibitors (ACEI) are the most effective antiproteinuric agents and should be used as first-line drugs in both diabetic and non-diabetic proteinuric nephropathies. The role of calcium channel blockers (CCB) is much more controversial. In diabetic patients verapamil and diltiazem seem more effective than dihydropyridines in reducing urinary protein excretion, and have additive effects with ACEI, but little is available on chronic treatment of non-diabetic nephropathies for non-dihydropyridine CCBs. To test whether the combination of verapamil 180 mg or amlodipine 5 mg with trandolapril 2 mg reduces urinary protein excretion more than trandolapril 2 mg alone, we planned a prospective, randomized, double-blind, multicenter trial. The secondary aims are to evaluate the effects of both treatments on the selectivity of proteinuria and check their safety. Consecutive patients aged between 18 and 70 years with non diabetic proteinuria > or =2 g/24 h and plasma creatinine < 3 mg/dl or creatinine clearance > or = 20 ml/min are asked to participate. After a four-week run-in during which previous antihypertensive therapy is withdrawn, a single dose of trandolapril 2 mg is given once a day in open conditions for four weeks. At the end of this period patients are randomly assigned to receive once a day, in a double blind fashion, either trandolapril 2 mg and verapamil 180 mg [plus a placebo], or trandolapril 2 mg plus amlodipine 5 mg. They are monitored after one, two, five and eight months. PMID- 11281339 TI - Long term effect of nifedipine GITS and lisinopril on subclinical organ damage in patients with essential hypertension. AB - BACKGROUND: Preventing subclinical organ damage is currently a major issue in the management of patients with essential hypertension. Antihypertensive drugs which act through different pathophysiological mechanisms might confer specific target organ protection beyond what is already provided by their blood pressure lowering effect. METHODS: Thirty-one patients with essential hypertension were randomized to receive long-term treatment with either a calcium channel blocker (nifedipine GITS, 90 mg/day) or an ACE-inhibitor (lisinopril, 20 mg/day). Blood pressure, left ventricular mass, carotid wall thickness and timed urinary albumin excretion were measured at baseline and over the course of 24 months of treatment. RESULTS: Both regimens significantly lowered mean blood pressure over the 24 months (from 124+/-2 to 103+/-2 mmHg in the lisinopril group and from 122+/-2 to 104+/-1 in the nifedipine group). Overall, end-organ damage improved with persistent blood pressure control. However, the two treatments had different specific effects. Lisinopril induced a more pronounced reduction of the left ventricular mass index (from 56+/-3 to 52+/-2 g/m2.7, P< 0.05) and urinary albumin excretion (from 34+/ 15 to 9+/-2 microg/min, P< 0.01), while nifedipine achieved a greater reduction of carotid intima plus media thickness (from 0.8+/-0.06 to 0.6+/-0.06 mm, P< 0.01). CONCLUSIONS: Blood pressure control does help reduce the severity of organ damage in patients with essential hypertension. Different antihypertensive treatments may confer additional specific cardiorenal and vascular protection regardless of blood pressure control. These data could be useful when devising individualized therapeutic strategies in high-risk hypertensive patients. PMID- 11281340 TI - Plasma sulfate concentration and hyperhomocysteinemia in hemodialysis patients. AB - BACKGROUND: Severe hyperhomocysteinemia is common in hemodialysis patients, who also present a dramatic increase in plasma concentrations of sulfate, one of the main products of methionine and cysteine catabolism. The aim of this study was to verify the relationship between high plasma sulfate levels and cysteine or homocysteine concentrations in hemodialysis patients. METHODS: Plasma sulfate, cysteine and homocysteine concentrations and some renal efficiency parameters were determined in 18 patients with end-stage renal failure, all undergoing 4h hemodialysis three times a week. The pattern of post-dialysis rises on plasma concentrations of sulfate, cysteine and homocysteine was established. RESULTS: Plasma sulfate, cysteine and homocysteine levels were significantly higher in patients than in normal controls. Plasma sulfate concentrations positively correlated with cysteinemia (p = 0.031; r = 0.482) which, in turn correlated with homocysteinemia (p = 0.042; r = 0.460). Sulfate levels also correlated with blood creatinine (p = 0.004; r = 0.630), nitrogen (p = 0.000; r = 0.899), protein (p = 0.014; r = 0.555), and albumin (p = 0.003; r = 0.642). Post-dialysis rises in sulfate and cysteine were detected some hours before homocysteine. CONCLUSION: The results suggest that high sulfate levels, due mainly to impaired renal function, are involved in the altered metabolism of homocysteine in hemodialysis patients. PMID- 11281341 TI - Cigarette smoking and kidney involvement. AB - Cigarette smoking has adverse effects on health causing ischemic heart disease, stroke, chronic obstructive lung disease and cancers of the respiratory and upper digestive tract, pancreas, kidney and urinary tract. Smoking causes an acute increase in mean arterial pressure and heart rate mediated by catecholamines and beta-adrenergic mechanisms. Chronic cigarette smoking reduces renal plasma flow, probably increasing synthesis of the vasoconstrictor endothelin and reducing generation of the vasodilatory endothelial nitric oxide. There is clinical evidence that cigarette smoking has important adverse effects on renal outcome in primary hypertension, diabetic nephropathy, primary glomerular diseases, systemic diseases involving the kidney and in patients on chronic hemodialysis, or after renal transplantation. PMID- 11281342 TI - Impact of high-flux/high-efficiency dialysis on folate and homocysteine metabolism. AB - High-flux/high-efficiency (HF/HE) dialysis may have detrimental effects on micro nutrients and water-soluble vitamins, such as vitamin B6, whose levels are lowered. Folate deficiency may increase cardiovascular risk through an increase in homocysteine (Hcy) serum levels. We therefore investigated the effects of dialysis with a high-flux (HF) membrane on folate and Hcy metabolism. Twelve patients without any folate supplementation, receiving dialysis with a low-flux membrane prior to the study (TO), were switched to dialysis using a HF triacetate membrane for four months (T1, T2, T3, T4) and received an oral daily folate supplementation during the two last months (T3, T4). Mean predialysis plasma folate levels fell dramatically after one month of HF dialysis (T1) and remained significantly lower than the initial level (p<0.05) at T2. Hcy concentrations were high in all patients at TO (mean 47.3 +/- 17.6 microM, normal range 5 to 15 microM). They did not change during the first two months of the study but dropped steeply after the beginning of oral folate supplementation. Folate supplementation should be used in HF/HE dialysis to avoid folate depletion. The combination of folate supplementation and HF/HE may lower Hcy levels and reduce cardiovascular morbidity and mortality in these patients. PMID- 11281343 TI - Hyperhomocysteinemia in renal transplant patients: an independent factor of cardiovascular disease. AB - Hyperhomocysteinemia (Hcy) is an independent factor of cardiovascular disease, which is the main cause of morbidity and mortality both in uremic and kidney transplant patients. The aim of the study was to determine Hcy, plasminogen activator inhibitor (PAI-1) and lipoprotein (a) (Lp(a)) serum levels in 70 patients with a well functioning renal transplant. We also verified whether these levels were modified by a multivitamin therapy. The genetic polymorphism of the methylenetetrahydrofolate reductase (MTHFR) enzyme which plays a main role in Hcy metabolism, was studied as well. We found Hcy, PAI-1 and Lp(a) levels significantly elevated with respect to healthy control subjects. The thermolabile form of the MTHFR enzyme was linked to higher Hcy levels. After a short time on therapy with B6, B12 and folic acid vitamins, Hcy and PAI-1 decreased to normal levels. The authors conclude that high Hcy levels could be a relevant covariate for cardiovascular disease in transplant patients and they suggest that vitamin supplementation be recommended as a part of therapy. PMID- 11281344 TI - Gitelman's syndrome (familial hypokalemia-hypomagnesemia). AB - Gitelman's syndrome (GS) is a heritable renal disorder characterized by hypomagnesemia, hypokalemia and hypocalciuria, and distinct from Bartter's syndrome (BS). As compared to those with BS, patients with GS present at an older age, and they have a milder clinical picture, normal or slightly decreased concentrating ability, reduced urinary excretion of calcium, and permanently decreased serum magnesium level. GS is caused by defective NaCl transport in the distal convoluted tubule, and linked to the gene encoding the thiazide sensitive Na-Cl-cotransporter located on chromosome 16q. Patients with BS, on the other hand, have mutations in the transporters in the thick ascending loop of Henle (NKCC2, ROMK, and C1C-Kb). Treatment of GS consists of magnesium salt replacement. Long term prognosis in terms of maintaining growth, preserving renal function and life expectancy is excellent. PMID- 11281345 TI - Healing of diabetes and vascular ulcers on switching from peritoneal dialysis to hemodialysis. AB - Vascular complications are the main problem in diabetic patients and can be worsened by continuous ambulatorial peritoneal dialysis (CAPD). A 46-year old woman with a family history of diabetes progressively developed hyperglycemia and subsequently lower limb ulcers after beginning CAPD. Hypertonic bags were required to control fluid balance. On account of the severe and painful ulcers, the patient was changed to hemodialysis. Within a few weeks her diabetes improved and the vascular ulcers healed completely. PMID- 11281346 TI - Hypertension-induced renal failure. PMID- 11281347 TI - The renoprotective effect of combined antihypertensive drugs. AB - In recent years, a target blood pressure (BP) of at least 125/75 mmHg has been sought in order to reduce the rate of progression of chronic renal failure (CRF) and cardiovascular mortality. Some antihypertensive agents, such as ACE inhibitors, calcium channel blockers and angiotensin receptor antagonists (and perhaps endothelin converting enzyme inhibitors and endothelin antagonists), may also reduce CRF progression because they block some of the pathogenetic mechanisms involved in renal damage. Although this effect seems to be partially independent of BP reduction, it is still not clear whether these drugs are really superior to other antihypertensive agents when low BP is achieved. However, the possibility that combination treatments with some of these drugs may confer additive or even synergistic renoprotective effects other than BP control is not only fascinating, but also important because multidrug antihypertensive regimens are required anyway to manage BP adequately in the majority of patients with CRF. PMID- 11281348 TI - Inaugural Ormrod Lecture. PMID- 11281350 TI - Basic knowledge of Internet in forensic medicine: logging on, fetching files and information. AB - The Internet stands at the forefront of telecommunications in medicine, including forensic medicine, since information technology (IT) has been able to revolutionize medical and scientific practices. Today, forensic physicians and professionals need to be familiar with the use of computers and the key applications of information technology: multimedia and the Internet. From the office, the forensic physician can communicate with other physicians by means of e-mail, take part in discussion groups, obtain information on meetings and get information from public libraries and various databases by means of file transfer protocol. The search among the huge amount of information is facilitated by the 'click and play' use of the Internet, by its increased ease and availability of access and by faster communications to an increasing number of accessible technical, scientific and biomedical resources. Therefore, it is useful to introduce some of the most frequent concepts encountered when exploring the Internet, to give simple references for any forensic physician exploring the Internet and to present typical Internet aspects by illustrations saved from worldwide websites and linked to the accompanying text. Some of the main forensic websites and basic Internet procedures are described, explaining how to search and exchange information in the domain of forensic medicine and sciences. Search engines and search procedures on the worldwide web are briefly explained. The aim of this paper is to give forensic physicians, scientists and law professionals some basic tools and references to access the vast possibilities of the Internet. PMID- 11281349 TI - Mononuclear cells in subcutaneous haemorrhage with special consideration of myeloid percursor cells. AB - Various hematogenous markers were used to differentiate and quantify the types of mononuclear cells present in subcutaneous haemorrhages. Fifty samples of subcutaneous bleeding with a survival time of a few minutes to more than 48 hours were studied. The various cell types were detected using the following stains: Naphthol AS-D chloracetate esterase for myeloid cells, including mast cells; (alpha1-antichymotrypsin for monocytes/macrophages; UCHL1 for T-lymphocytes; and L26 for B lymphocytes. The percentage of monocytes/macrophages was found to increase in dependence on survival time, whereas T-lymphocytes declined. Within minutes of injury neutrophilic granulocytes had emigrated into the surrounding tissue and mast cell degranulation had occurred within the haemorrhagic zone. Esterase-positive mononuclear cells, namely metamyelocytes, were detected within minutes after injury and were still present after survival times exceeding 48 hours; however, no dependence on survival time or cause of death was found. Although the increasing number of monocytes/ macrophages and T-lymphocytes was expected, the sometimes high percentage of myeloid precursor cells within the wound were surprising. Possible explanations for this phenomenon are discussed. PMID- 11281351 TI - The sexual habits of males who molest. AB - To gain better understanding into the causes of molesting behaviour in males, we examined a prison cohort of molesters and compared them with a control group comprising thieves. The molesters were older, had attained a higher educational level and were more likely to be married. They had their initial sexual intercourse at an older age but tended to have multiple sexual relationships and were less likely to report having viewed pornographic material. Three months prior to committing their offences, molesters seemed to engage in less sexual activity. The implications are that molesters had a need for more explicit sexual contact but were unable to sustain ongoing, stable, intimate sexual relationships. PMID- 11281352 TI - The identification and treatment of opiate users in police custody. AB - All detainees admitted to seven London police stations were observed over a six month period (n=2,947). Four per cent were identified as opiate users, although the actual percentage is likely to be much higher. Compared to the general population of detainees there were significantly more women among known opiate users and this group also contained a higher percentage of white detainees and people born in the British Isles. People born in continental European countries were also over-represented. A little more than half of known users did not reveal their use on arrival at the police station. At least 60% of known opiate users remained well throughout their detention, 30% were intoxicated through drugs at the time of their arrest, but only 13% displayed signs or symptoms of withdrawal during their detention. Overall, 65% of the known opiate users were seen by a police surgeon and of these 52% were given medication. All of those withdrawing were given drug treatment, but most of those who were intoxicated by opiates, or who remained well throughout their detention, received no medication. Of those given medication 86% received an opiate, dihydrocodeine being the commonest preparation, usually in association with a benzodiazepine. Despite the adoption of differing management paradigms among police surgeons, the actual medical treatment of opiate-using detainees was pragmatic and determined by individual need. PMID- 11281353 TI - Section 5(2) of the Mental Health Act 1983 in practice: an audit and a review. AB - All applications of s.5(2) of the Mental Health Act 1983 (MHA) between January 1997 and December 1998 were examined to assess variables likely to affect outcome and to compare these findings to other similar published studies. Of the 154 applications (7% of all admissions), 56 were converted to s.3 and 39 to s.2 of the MHA. We found that the time of application, grade of doctor making the application and the day of application were the best predictors of outcome of s.5(2). Apart from a few exceptions, our findings were generally in keeping with previous published results. These findings suggest a national trend in the clinical use of s.5(2) and may provide a useful guide for those considering reform of this part of the MHA. PMID- 11281354 TI - Formal justifications for compulsory psychiatric detention. AB - The formal justifications for all detentions under s.2 of the Mental Health Act 1983 within an inner-city mental health trust were examined over a 12-month period. The study explored: the nature of the justifications for detention; the extent to which these were associated with patient characteristics; and the extent to which the two medical practitioners involved in each case agreed on the justifications. The justifications reflected a greater emphasis on the protection of the individual concerned rather than the protection of others. A content analysis of the textual justifications revealed five broad themes: the nature of the risk posed by the patient; the patient's capacity to provide informed consent; their need for hospitalization; their lack of consent to informal admission; and their reliability or likely compliance. There was a significant association between patients' sex, ethnic group, diagnosis and the nature of risk indicated in the documentation, but further research is needed to clarify the nature of this association. The study found that in nearly a quarter of cases, the two professionals did not agree about whether or not the patient presented a danger to others. This lack of agreement was not associated with any patient or professional characteristics, and may reflect the complexity of this area of risk assessment. The authors suggest that the issue of 'risk' needs to be addressed in a more sophisticated manner within the Mental Health Act. Specifically, further guidance is needed as to the nature and levels of risk that constitute grounds for detention. Further guidance is also needed regarding the issues that need to be recorded on the legal documentation for detention. PMID- 11281355 TI - Institutional behaviour and time in treatment among psychopaths admitted to a prison-based therapeutic community. AB - Previous research has indicated that psychopathy is associated with high levels of institutional misconduct and high attrition rates in therapeutic communities. However, most of this work has been conducted with North American populations. In the present study, data is presented on institutional behaviour and time in therapy for 104 inmates admitted to Grendon Therapeutic Prison. The results indicated that high scores on the Hare Psychopathy Checklist Revised were significantly associated with failure to progress from the assessment unit onto a therapy wing and the number of adjudications and security information reports. A trend was found for psychopathy to be associated with shorter periods in therapy, but this just failed to reach significance. The results confirm previous research on the association between psychopathy and institutional misconduct and provide some support for the association between psychopathy and early termination of treatment in therapeutic communities. A number of methodological issues are discussed. PMID- 11281356 TI - Use of the Mental Health Act criteria in the decision-making process for compulsory admissions: a study of psychiatrists in South London. AB - This study investigates the decision-making process for admitting patients compulsorily under the Mental Health Act 1983 of England and Wales. We used three case-vignettes describing different clinical situations: 1) a man with depression and psychotic symptoms; 2) a woman with a possible first episode of psychosis; and 3) a man with a history of substance abuse and bipolar disorder. The vignettes were administered to a group of psychiatrists working at the Bethlem Royal and Maudsley Hospitals in South London. The psychiatrists were asked to rate 11 factors from the most to the least important in their decision to admit the individual compulsorily. Three factors resembled the criteria considered in the Mental Health Act 1983 for compulsory admission: current mental state; severity of the disease; and dangerousness to self or others. Three were other clinical features of the patient: diagnosis; psychiatric history; and likely response of the mental state to the medical treatment. The remaining five were sociodemographic features of the patient: age and gender; owning a home; occupational status; and social support available. The psychiatrists were not given the option that the patient would accept a voluntary admission to hospital. We obtained responses from 42 psychiatrists (25 males and 17 females). The most important factor for deciding to detain a patient compulsorily was perceived dangerousness to self and others. The current mental state of the patient and the severity of the illness were the next two important factors. Our results confirm that the criteria recommended by the Mental Health Act 1983 of England and Wales were applied consistently in three different hypothetical situations. The need for protection of the patient or others may take precedence over the current mental state of the patient or the severity of the illness, a finding that warrants further investigation in view of the current debate on the responsibility of psychiatrists in deciding to detain subjects affected by personality disorder. PMID- 11281357 TI - What happens to special hospital patients admitted to medium security? AB - This article describes the outcome of all the Special Hospital patients admitted to and discharged from the Norvic Clinic over a period of ten years since its opening in 1984. This study identified 23 patients. Of those transferred, 56% had a legal classification of mental illness. A much higher percentage (91%) were restricted under the powers of s.41 of the Mental Health Act 1983, compared to 69% in the West Midlands study (Cope and Ward, 1993). There appears to be a significant difference between the mental disorder groups for age on admission to Special Hospital when considering all patients (males and females). This difference is not maintained when males and females are looked at separately, although the result for males is nearly significant (p = 0.0606). The number of female patients is small (five) and so the tests will hardly be reliable for the female sample. No significant differences were demonstrated for any of the other variables considered. Like the West Midlands study, minor reconvictions were completely absent. This was an unusual finding within the West Midlands study and not replicated from any of the previous Special Hospital studies. In this study, failure constituted an unsuccessful transfer with return to Special Hospital and reconviction. The percentage of those returned to Special Hospital without reaching the community was slightly lower than the West Midlands study, for both the mentally ill and those classified as suffering from psychopathic disorder. PMID- 11281358 TI - Aorto-oesophageal fistula and intestinal infarction secondary to volvulus following ingestion of foreign bodies in a schizophrenic patient. AB - We present a case of death caused by voluntary ingestion of non-organic foreign bodies observed at the Institute of Legal Medicine of the University of Trieste. The victim was a young schizophrenic patient whose death, at first giving rise to suspicions of murder by another psychiatric patient, was found to be caused by an uncommon chronic permanence of foreign bodies at different locations of the digestive tract which suddenly evolved into a series of simultaneous lethal complications as yet never described. The case also raised the issue of possible responsibilities of the subject's healthcare providers. PMID- 11281359 TI - Full-fat cream. PMID- 11281360 TI - Cellular immune-endocrine interaction in adrenocortical tissues. AB - BACKGROUND: The detection of important immunocompetence-like features on endocrine steroid cells raises questions about direct intercellular communication between the adrenal and immune systems. This article summarizes our recent work and new data on immune-adrenal interactions. MATERIALS AND METHODS: RT-PCR and immunohistochemistry were performed to examine MHC class II (HLA-DR) expression in adrenocortical tumours. Coculture systems of NCI-H295 adrenocortical carcinoma cells and HLA-matched lymphocytes were used to examine effects on steroid production and survival of lymphocytes. RESULTS: HLA-DR m-RNA is found in both benign and malignant adrenals, except the NCI-H295 cell line. Under direct coculture conditions with NCI-H295 cell line, spontaneous apoptosis of immune cells was reduced. Synthesis of cortisol and especially of dehydroepiandrosterone production of tumour cells was markedly increased. Differences by separating CD4- and CD8- T cells were not detected. CONCLUSIONS: Direct cellular contact between lymphocytes and adrenocortical cells seems to be involved in the peripheral regulation of androgen synthesis in the adrenal. The molecular basis of this interaction is not known. With regard to normal adrenals, ligation of MHC class II antigens could be a potential mechanism for a peripheral regulation of androgen secretion. PMID- 11281361 TI - Differentiation and regulation of the corticotropic pituitary cell. AB - A number of primary diseases of the pituitary with adrenocorticotropin dysregulation have been recognized. A few genetic defects have been identified as causes of secondary adrenocortical insufficiency. Much less is known about the ontogeny of corticotrophic tumours leading to a hypercorticolaemic state. To improve the diagnosis and treatment of these disorders, a better understanding of the mechanisms of corticotrophic pituitary cell differentiation and regulation is of clear interest. Studies using molecular tools have enhanced our knowledge over recent years, and a few reports of considerable relevance are summarized in this review. PMID- 11281362 TI - Transcription factors as regulators of steroidogenic P-450 enzymes. AB - Steroid hormones are essential for normal sexual development, accommodation to stress, and regulation of fluid and electrolyte balance. Biosynthesis of these different classes of steroids and its appropriate regulation requires the precisely controlled expression of six different cytochrome P-450 enzymes and two hydroxysteroid dehydrogenases in different tissues. The molecular mechanism of the tissue-specific and pituitary hormone-regulated expression of the genes encoding P-450 enzymes in the steroidogenic tissues is the central problem of long-term regulation of steroidogenesis. Orphan members of the nuclear receptor superfamily play an important role in mediating transcriptional regulation of several steroid hydroxylase genes. Two of these transcription factors, steroidogenic factor-1 (SF-1) and DAX-1, will be reviewed here in detail. PMID- 11281363 TI - 11Beta-hydroxysteroid-dehydrogenase isoforms: tissue distribution and implications for clinical medicine. AB - 11Beta-hydroxylation is essential for glucocorticoid and mineralocorticoid activity of a steroid. The enzyme catalyzing this reaction is termed 11beta hydroxysteroid-dehydrogenase (11beta-HSD). Two isoenzymes of 11beta-HSD have been characterized in human tissues. Whereas 11beta-HSD-I works mainly as a reductase, 11beta-HSD-II only functions as an oxidizing (inactivating) enzyme for physiological glucocorticoids. Thus, the tissue distribution of both enzymes plays a crucial role for the specific glucocorticoid status of an organ. This review summarizes our knowledge of tissue distribution of both 11beta-HSD isoenzymes, their physiological function and pathophysiological role in certain clinical abnormalities, and their relevance to the metabolism of synthetic glucocorticoid and mineralocorticoid compounds. PMID- 11281364 TI - Intra-adrenal regulation of androgen synthesis. AB - The adrenal cortex encloses the neuroendocrine medulla and is itself subdivided into three distinct zones, each having a specific function and regulation. While the glomerulosa and the fasciculata control vital systems of mineral and energy supply, which are stringently regulated by higher control factors, the function of the reticularis is less clear, beyond supplying a pool of weak androgens, and consequently we do not understand its redundant regulation. The following questions need to be answered: 1. How is the formation of dehydroepiandrosterone (DHEA) and androstenedione differently regulated from glucocorticoid synthesis in normally functioning adrenals? 2. How might growth factors, which increase prepubertally, prime the adrenarche? 3. The regulation of the 17/20-lyase enzyme activity is one of the key factors of adrenal androgen secretion (review [2]). How can the two activities of the P450c17 enzyme be differently regulated in the same cell in a developmentally dependent fashion? This review focuses on the intra-adrenal growth factor system and on the role of 17/20-lyase regulation, as well as on their possible interactions. The increase of activity of the 17/20 lyase enzymatic activity is necessary for the rise of C19 steroids, while the relative increase of formation of DHEA is only possible in the presence of a low 3beta-hydroxysteroid-dehydrogenase (3betaHSD) activity. PMID- 11281365 TI - Intracrinology and the local enzymatic control of hormone distribution and metabolism: dehydroepiandrosterone does not just act as a prohormone for androgens and estrogens. AB - The study of subcellular environments for the interaction of biomolecules and observations of certain features of hormone actions have nourished the concept of 'intracrinology', which describes hormone actions within a singular cell, in contrast to the well-described autocrine, paracrine and endocrine fashion. Synthesis and metabolism of DHEA make it a likely candidate for intracrine actions in target tissues. Recent experimental findings are reviewed in this context. PMID- 11281366 TI - Leptin and the adrenal gland. AB - BACKGROUND: Leptin is involved in the maintenance of energy balance acting on food intake, thermogenesis and energy expenditure. Via its receptor in the hypothalamus, leptin modulates the functioning of the hypothalamic-pituitary adrenal axis and the systemic sympathetic/adrenomedullary system, which are closely linked to the regulation of energy balance and body weight. In regard of potential interactions of leptin and adrenal hormones this study intended to characterize the role of leptin in the human adrenal gland. MATERIALS AND METHODS: A novel technique of laser capture microdissection was used to separate cortical and chromaffin cells for mRNA expression studies of leptin receptor isoforms and leptin mRNA in adrenal tissue and cell line NCI-H295. Immunostaining was used to localize leptin receptor in human adrenal slices. The influence of leptin on basal and ACTH-stimulated steroid hormone secretion and enzyme expression was assessed. The effect of leptin on proliferation and viability of adrenal cells in primary culture and of the NCI-H295 cell line was studied by the WST-1 assay and by 3H-thymidine test. RESULTS: Our data demonstrate that leptin can regulate the human adrenal function directly, via its receptors on adrenocortical cells. Leptin decreased the corticotropin-stimulated release of steroid hormones in vitro without any effect on cell proliferation. Leptin did not significantly affect the expression of cytochrome P450 scc m RNA in humans, but decreased the ACTH stimulated expression of the cytochrome P450 17alpha mRNA [corrected]. CONCLUSIONS: The adipo-adrenal interaction mediated by leptin further underscores the close link of metabolism and stress regulation in humans. PMID- 11281367 TI - Dehydroepiandrosterone--a neurosteroid. AB - Dehydroepiandrosteone (DHEA) and its sulfate ester (DHEAS) are the major secretory products of the human adrenal glands and serve as precursors for both androgenic and estrogenic steroids. DHEA/S concentrations are particularly high in the brain, and DHEA/S and related steroids can be synthesized de novo in brain glial cells. Therefore, the term 'neurosteroids' has been coined for these compounds. This review summarizes findings in neurosteroid physiology on a cellular and molecular level, and outlines current concepts of how these compounds modulate physiological functions of the brain. Today, despite promising preclinical and human data the present clinical studies provide only weak evidence, if any, in favour of a DHEA replacement therapy. PMID- 11281368 TI - Nongenomically initiated steroid actions. AB - The classical theory of steroid hormone action comprises binding to an intracellular receptor followed by modulation of transcriptional and translational events. These cumbersome model explains the characteristic latency of these genomic steroid effects. Over the past two decades, increasing evidence for rapid nongenomic effects of steroids, incompatible with the traditional model, has accumulated. These alternative steroid effects have been described for all classes of steroids and a multitude of species and tissues with different mechanisms of action. PMID- 11281370 TI - Histopathological classification of adrenal tumours. AB - Tumours in the adrenals originate from the adrenal cortex, the adrenal medulla or as metastases from extra-adrenal primaries. Differentiating between these three groups is the first task a pathologist has to tackle when dealing with specimens from the adrenal region. Whereas this is possible in every case with total removal of the adrenal tumour it may be impossible in fine needle biopsies of such tumours. The second great problem is the dignity of adrenal tumours, which cannot be determined in many adrenomedullary and some adrenocortical tumours. Immunostainings are helpful but the basic method remains the histopathological examination of paraffin sections. This review gives an update of pathological findings in several adrenal tumour entities, and provides guidelines for the diagnosis of these tumours in the light of recently published data. PMID- 11281369 TI - How hydrocortisone substitution influences the quality of life and the bone metabolism of patients with secondary hypocortisolism. AB - BACKGROUND: Even in the setting of chronic glucocorticoid substitution in hypocortisolaemic patients, severe side-effects will eventually occur when the dosage is inappropriately high. This study evaluates the effect of usual hydrocortisone substitution dosages on the well-being of the patients and on parameters of the bone metabolism to establish an optimum substitution dosage. DESIGN: In a double blind study nine patients with secondary hypocortisolism, being divided in three groups of three, received different doages of hydrocortisone (15, 20, 30 mg per day). Well-being was assessed using three different, validated questionnaires. Markers of bone metabolism were measured in blood and urine. RESULTS: The patients' quality of life was not impaired even at low dosages of hydrocortisone (15 or 20 mg per day). Of all laboratory parameters only osteocalcin significantly changed, decreasing at higher dosages. CONCLUSIONS: Our study shows that a risk of bone loss may be avoided with a substitution dosage of 20 mg or even 15 mg hydrocortisone per day, without influencing the well-being of the patient. PMID- 11281372 TI - Molecular adrenocortical tumourigenesis. AB - Adrenocortical neoplasms are the most frequent abnormality of the adrenal cortex. Most of these lesions are clinically silent and are detected incidentally by ultrasound or computed tomography. The prevalence of these so-called 'incidentalomas' in the general population is around 1%, increasing with age and reaching 6% among those in the age range 60-70 years. In contrast, primary adrenocortical carcinoma, a highly malignant tumour, is rare, having an incidence of one case per million per year. Recent progress has been achieved in the understanding of adrenocortical tumourigenesis by mapping and identification of genes responsible for hereditary tumours that involve the adrenal gland. Investigation of the clonal composition of adrenal tumours demonstrates that adrenal carcinomas are monoclonal, whereas adrenal adenoma may be polyclonal in approximately 25-40% of cases. Oncogenes and tumour-suppressor genes involved in adrenal carcinomas include mutations in the p53 tumour-suppressor gene and rearrangements of the chromosomal locus 11p15.5 associated with IGF II hyperexpression. Constitutive activation of the ACTH receptor-G protein-cAMP signal cascade does not play a role in adrenal tumour formation. Conversely, deletions of the ACTH receptor gene have recently been found in undifferentiated adenomas and in aggressive adrenocortical carcinomas, and, more recently, confirmed in a larger series of tumours. The available literature indicates that the signalling pathways of adrenocortical tumours are different from those of other endocrine neoplasms, such as pituitary and thyroid adenomas. PMID- 11281371 TI - Molecular mechanisms of dissociative glucocorticoid activity. AB - BACKGROUND: Glucocorticoids mediate their effects on target cells via transactivation and transrepression of certain target genes. While conventional glucocorticoids do not distinguish between transactivation and transrepression, new glucocoticoids should be able to dissociate these effects, thus lowering the potential of unwanted side-effects of glucocorticoids in clinical use. In this study, we developed a new experimental system to test potentially selective glucocorticoids in normal lymphocytes. MATERIALS AND METHODS: Following pretreatment with phytohaemagglutinin, normal lymphocytes were transfected, using electroporation, with pGL3 luciferase reporter vectors under the control: (1) of the human IL-2 promoter; and (2) of a glucocorticoid response element (GRE). Luciferase activity was measured in response to various steroid compounds, including the potentially dissociative glucocorticoid medroxyprogesterone acetate (MPA). RESULTS: The IL-2 promoter was induced 267.2 +/- 27.5-fold (mean +/- SD) by phorbol ester and ionomycin. In these cells, hydrocortisone and dexamethasone caused a 22.9 +/- 3.6% and a 38.4 +/- 10% reduction in luciferase activity, respectively. Under GRE control, hydrocortisone stimulated luciferase activity 6.4 +/- 0.50-fold and dexamethasone 8.2 +/- 0.4-fold. MPA-induced transrepression was 73.3 +/- 7.2% for the IL-2 promoter, and transactivation was 2.4 +/- 0.4-fold with the GRE-driven construct. The natural progestin progesterone did not have significant effects on either construct. CONCLUSIONS: This is the first system that allows efficient analysis of glucocorticoid-dependent transactivation and transrepression in normal human lymphocytes. Compared to conventional glucocorticoids, MPA can be referred to as a dissociative glucocorticoid, its transrepression/transactivation ratio being 6.6 (transrepression 1.91/transactivation 0.29), with dexamethasone being the standard (transrepression 1/transactivation 1). We conclude that MPA is a highly promising substance for the treatment of autoimmune/inflammatory diseases. PMID- 11281373 TI - The role of the insulin-like growth factor system in adrenocortical tumourigenesis. AB - BACKGROUND: The insulin-like growth factor (IGF) system plays a central role in the mechanism of transformation and tumourigenesis. Elevated levels of IGF-II and IGF-I have been found in adrenocortical carcinomas. MATERIAL AND METHODS: We examined binding characteristics and concentrations of both IGF-receptors in normal adult human adrenocortical glands, and compared them with the IGF-I receptor binding in adrenocortical rumours of various origins. The human IGF-I receptor was overexpressed in the mouse adrenocortical tumour cell line Y1, and growth studied in response to IGF stimulation. The influence of IGF-II on adrenal morphology and function was assessed in transgenic mice that postnatally overexpress IGF-II. RESULTS: While the abundance of the IGF-I receptor in adrenocortical hyperplasias and adenomas was similar to normal tissue, a strong overexpression of the intact IGF-I receptor was found in three out of four adrenocortical carcinomas. Y1 cells overexpressing the human IGF-I receptor respond to IGF-I with an increase in thymidine incorporation by 140%. Furthermore, the antiproliferative effect of ACTH is blunted. In transgenic mice postnatally overexpressing IGF-II, adrenal weight is increased, mainly due to a 50% increase in the number of zona fasciculata cells. Plasma corticosterone levels in these mice are twofold higher than in controls, in contrast to similar plasma ACTH levels, thus indicating a direct effect of IGF-II on adrenal cell hyperplasia and function. CONCLUSION: There is substantial evidence that the IGF system is involved in adrenal growth and tumourigenesis. High local levels of IGF II in combination with elevated IGF-I receptor concentrations would represent a significant growth advantage of the adrenocortical carcinoma cell and could contribute to a highly malignant phenotype. IGF-II overexpression alone seems not to be sufficient for malignant transformation. PMID- 11281374 TI - New mechanisms of adrenostatic compounds in a human adrenocortical cancer cell line. AB - BACKGROUND: Adrenostatic compounds are frequently used in the treatment of patients with Cushing's syndrome and act via direct inhibition of steroidogenic enzymes. However, additional mechanisms may be involved in the blockade of adrenal steroid secretion. We therefore investigated the effects of aminoglutethimide (AG), metyrapone (MTP) and etomidate (ETO) in the human NCI h295 adrenocortical carcinoma cell line. MATERIALS AND METHODS: Cells were incubated with increasing doses of the adrenostatic compounds. Steroid hormone secretion (cortisol, 17-OH-progesterone, DHEA-S) and cAMP synthesis were determined and Northern blot analysis and cell proliferation experiments were performed. RESULTS: ETO was the most potent adrenostatic compound inhibiting P450c11 hydroxylase at low concentrations (IC50 15 nM), and also blocking P450 side-chain cleavage (scc) enzyme (IC50 400 nM) at higher concentrations. The pattern of enzyme inhibition was similar for MTP with an IC50 of 3-5 microM for P450c11 and 17 microM for P450scc, while AG blocked P450scc with an IC50 of 10 microM. AG significantly suppressed the baseline ACTH-R mRNA expression in a dose dependent fashion (300 microM AG: 5% +/- 1%; 30 microM AG: 64% +/- 1%; 3 microM AG: 108% +/- 19% compared with control cells: 100% +/- 11%) but increased glucocorticoid receptor mRNA. The reduced ACTH-R mRNA was paralleled by low ACTH induced cAMP accumulation indicating reduced expression of ACTH-R protein. The simultaneous incubation of hydrocortisone together with AG reversed the inhibitory effect of AG on the ACTH-R expression. AG and ETO inhibited cell proliferation in the NCI-h295 cells, but ETO was much more potent and showed antiproliferative effects at concentrations of 6 microM. The growth inhibition was not reversed by administration of hydrocortisone. CONCLUSIONS: Our data demonstrate that adrenostatic compounds not only act by suppression of steroidogenic enzymes but can also influence both ACTH-R expression and cell proliferation in adrenal cells. As these effects occur in vitro at concentrations that are reached during treatment with these drugs in patients, they are probably also of clinical relevance. Particularly the antiproliferative activity of ETO may be useful in Cushing's syndrome due to adrenocortical cancer. The interaction of steroidogenesis, ACTH-R and glucocorticoid receptor expression as well as cell proliferation provides a new concept of the intra-adrenal response to stress in humans. PMID- 11281375 TI - Metabolism of glucocorticoids and mineralocorticoids in patients with adrenal incidentalomas. AB - BACKGROUND: Adrenal incidentalomas are mostly nonfunctioning adrenocortical adenomas (NFI). However, in 5%-12% of the patients a preclinical Cushing's syndrome (PCS) with autonomous cortisol production by the tumour is present. Since urinary free cortisol excretion is not sensitive enough to determine subclinical hypercortisolism, in the present study more sensitive indicators of daily cortisol production were measured. DESIGN: (1) Tetrahydrocortisol, tetrahydrocortisone, urinary free cortisone together with urinary free cortisol were measured in 35 patients with adrenal incidentalomas (29 NFI and six PCS) and in 35 healthy controls. (2) Since little is known about daily aldosterone production, aldosterone metabolite excretions were measured. (3) As recently reported, ACTH stimulation revealed an increased response of precursors of the glucocorticoid and mineralocorticoid pathway. To find out which steroidogenic enzymes have altered activities, a 1-24ACTH stimulation test (250 microg i.v.) was carried out in 25 patients and 18 healthy controls with determination of multiple steroids. (4) Finally, since it was assumed that 21-hydroxylase deficiency or even 11b-hydroxylase deficiency may be involved in adrenal tumourigenesis, the prevalence of germline CYP21B and CYP11B1 mutations were studied in the same patients, who had underwent the ACTH stimulation test. RESULTS: (1) Glucocorticoid metabolites were within the normal range in all but three patients with NFI. As a group, the patients had subtle alterations in cortisol metabolism. Tetrahydrocortisol excretion was elevated in NFI and PCS compared with normal subjects (2.1 +/- 0.2 and 2.5 +/- 0.5 vs. 1.5 +/- 0.1 mg 24 h(-1); P < 0.05). Accordingly, the twofold elevation of the tetrahydrocortisol/free cortisol ratio indicates an increased 5beta-reduction of cortisol in the liver. (2) Tetrahydroaldosterone and aldosterone-18-glucuronide excretions were not different to controls. (3) In patients with incidentalomas an increased response to ACTH was seen for 17-hydroxyprogesterone (595 +/- 133 vs. 160 +/- 25 ng dL(-1)), 21-desoxycortisol (105 +/- 25 vs. 29 +/- 9 ng dL(-1)) and 11-desoxycortisol (401 +/- 40 vs. 293 +/- 17 ng dL(-1)). (4) In only one of 25 patients, a heterozygous deletion in exon 3 of the CYP21B gene was detected. CONCLUSIONS: (1) In conclusion, even the excretion of the main glucocorticoid metabolites is not a marker sensitive enough to distinguish between NFI and PCS. However, it is also possible that alterations in cortisol secretion are qualitative rather than quantitative. (2) Zona glomerulosa function is not influenced. (3) The elevation of 21-desoxycortisol argues against an impairment of 11beta-hydroxylase and favours a decreased activity of 21-hydroxylase. All others had wild-type sequences of both genes. (4) In conclusion, neither 21 hydroxylase deficiency nor 11beta-hydroxylase deficiency are predisposing factors for adrenal tumourigenesis. PMID- 11281376 TI - Molecular cues for the development of adrenal chromaffin cells and their preganglionic innervation. AB - Based on recent evidence from in vitro and gene knockout/insertion studies, this short review summarizes the molecular scenario underlying the development of adrenal chromaffin cells and their preganglionic innervation. During migration of neural crest cells from the dorsal surface of the neural tube to their destinations in the sympathetic primordia and adrenal glands, precursors of the so-called sympathoadrenal (SA) cell lineage are exposed to signals from the notochord and ventral neural tube probably including the protein, Sonic hedgehog. These, and signals in the region of the dorsal aorta (members of the family of bone morphogentic proteins), where SA progenitor cells subsequently assemble, are essential for the induction of the adrenergic phenotype. SA progenitor cells subsequently differentiate into paravertebral and prevertebral sympathetic neurones, intra- and extra-adrenal chromaffin cells and intermediate SIF (small intensely fluorescent) cells. Based on in vitro studies with isolated SA and chromaffin progenitor cells, glucocortiocids have been claimed as essential for suppressing neuronal commitment and for channelling SA cells towards the chromaffin phenotype. However, mice deficient for a functional glucocorticoid receptor possess the full complement of adrenal chromaffin cells at birth, suggesting that signals other than glucocorticoid hormones may be important in triggering chromaffin cell differentiation. The cholinergic neurones that are preganglionic to adrenal chromaffin cells have their cell bodies located in the intermediolateral column (IML) of the spinal cord. For their normal development, these neurones require signals from the adrenal medulla, which include neurotrophin-4, a major neurotrophic factor of adrenal chromaffin cells. Taken together, these data provide a more complete picture of molecular signalling in the development of one of the most important neuroendocrine tissues in vertebrates. PMID- 11281377 TI - The role of interleukin-6 in the human adrenal gland. AB - Interleukin (IL)-6 is a potent activator of the hypothalamic-pituitary-adrenal (HPA) axis on all levels in humans, and appears to play a pathogenic role in conditions related to chronic stress and physiological ageing; with physiological ageing showing a similar hormonal and immunological pattern to chronic stress. IL 6 and its receptor IL-6R are co-expressed at similar sites in the human adrenal gland, which seems to be an important source of IL-6 production. In vitro, in primary cultures of adrenal gland cells, chronic exposure to IL-6 stimulates adrenocortical steroid release in a time- and dose-dependent manner. This explains the high systemic cortisol levels in the absence of adequate plasma concentrations of corticotropin (ACTH) observed in patients after long-term treatment with IL-6. It could therefore be concluded that in situations of prolonged stress, when corticotropin-releasing hormone and ACTH release are suppressed by feedback inhibition due to circulating glucocorticoids, IL-6 maintains the elevated glucocorticoid levels by direct stimulation of adrenocortical steroidogenesis via autocrine/paracrine mechanisms. PMID- 11281378 TI - High-dose mitoxantrone and cyclophosphamide without stem cell support in high risk and advanced solid tumors: a phase I trial. AB - This phase I study was designed to develop a high-dose combination of two cycles of mitoxantrone and cyclophosphamide in patients with solid tumors, as an alternative to single-cycle high-dose regimens that use only alkylating agents. Treatment was delivered with granulocyte colony-stimulating factor (G-CSF), but without stem cell support, in order to avoid potential tumor cell reinfusion. Thirty-one patients with advanced solid tumors received two cycles of high-dose mitoxantrone (20-30 mg/m2) plus high-dose cyclophosphamide (3000-4000 mg/m2). All patients received G-CSF until hematologic recovery. Dose-escalation was performed when less than 50% of cycles per level had dose-limiting toxicity (DLT). The maximum tolerated dose (MTD) achieved was mitoxantrone 25 mg/m2 and cyclophosphamide 4000 mg/m2. Main dose-limiting toxicities (DLTs) were hematological: grade IV neutropenia lasting more than 7 days and thrombopenia below 20 x 10(9)/l requiring more than one platelet transfusion. Non hematological DLT consisted predominantly of grade III emesis and asthenia. Follow-up after each cycle was performed in an outpatient setting and there were no toxic deaths. In conclusion, the administration of two cycles of high-dose mitoxantrone and cyclophosphamide with G-CSF support is safe and feasible. MTD was mitoxantrone 25 mg/m2 and cyclophosphamide 4000 mg/m2. Evaluation of this regimen is being done in a phase II trial. PMID- 11281379 TI - A fludarabine-based conditioning regimen for severe aplastic anemia. AB - Graft rejection is a common problem after alternative donor transplantation for patients with refractory severe aplastic anemia (SAA). Intensification of the conditioning regimen, with the inclusion of irradiation, has often been advocated to combat this problem. With this approach engraftment rate improved, but the incidence of transplant-related complications is also increased, resulting in little change in the overall outcome. We investigated the use of the combination of fludarabine, cyclophosphamide and anti-thymocyte globulin as the conditioning regimen in five multiply-transfused SAA patients. Three patients received an HLA one-antigen disparate related donor transplant, while two patients were given marrow from matched, unrelated donors. The regimen was well tolerated, with only grade I toxicity encountered. With a median follow-up of 9 months, all patients are alive with complete donor chimerism. We conclude that fludarabine may be used in place of irradiation to augment the conditioning regimen of cyclophosphamide and anti-thymocyte globulin for alternative donor transplantation in children with SAA. PMID- 11281380 TI - Osteopetrosis: a single centre experience of stem cell tranisplantation and prenatal diagnosis. AB - Malignant osteopetrosis (MOP) is an autosomal recessive disease in which osteoclast dysfunction results in excessive bone deposition and early infant death. Thirteen children suffering from MOP from four related families all belonging to one Bedouin tribe, were studied. The disease was diagnosed as early as at a few days postnatal to 5 months. Nine children underwent BMT, four of whom are still alive; one is blind and two have markedly reduced vision. Four children who did not undergo BMT died between 4 and 6 months of age. Recently, the gene for MOP has been mapped for this Bedouin tribe allowing prenatal diagnosis. Seven pregnancies were subsequently prenatally diagnosed and two fetuses were found to be affected. Pregnancy was electively terminated in one case. In the other case the parents refused and after establishing the diagnosis, the newborn was transplanted at the age of 7 days. PMID- 11281381 TI - Cord blood collection before and after placental delivery: levels of nucleated cells, haematopoietic progenitor cells, leukocyte subpopulations and macroscopic clots. AB - The number of nucleated cells infused into the recipient of a cord blood (CB) transplant has emerged as the most important factor affecting the probability and speed of engraftment. At present, there is no international consensus on the procedure of CB collection in the maternity ward. In order to maximise the yield of viable cells in a CB unit, we aimed to investigate the efficiency of CB collection, with respect to the time of delivery of the placenta. We analysed stem and progenitor cells in terms of CD34+ cell content and colony-forming activities, lymphocyte subpopulations and the presence of macroscopic clots in 93 paired CB samples, collected before and after the delivery of the placenta. Our results demonstrated that the median concentrations of nucleated cells and total colony-forming unit (CFU) were significantly lower in CB collected after placenta delivery by 9.5% (P < 0.001) and 11.6% (P = 0.015), respectively, when compared to their counterparts collected before placental delivery. A reduction of granulocytes (P < 0.001), monocytes (P < 0.001) and CD19+ B lymphocytes (P = 0.031) was observed, with no significant change in the proportion of T cell subsets (CD4+, CD8+ cells) or activated T cells (CD25+, CD45RO+ cells) in samples collected after placenta delivery. The incidence of macroscopic clots was also higher in these samples (31% vs 1%, P < 0.001). The reduction of stem and progenitor cells correlated significantly with that of major cell populations, indicating a general cell loss, possibly due to clotting activities developed with time. Our study has documented strong evidence for recommending the collection of CB before the delivery of the placenta. PMID- 11281382 TI - Bone marrow transplantation from matched siblings in patients with fanconi anemia utilizing low-dose cyclophosphamide, thoracoabdominal radiation and antithymocyte globulin. AB - Nineteen patients with Fanconi anemia (FA) and bone marrow failure underwent bone marrow transplantation (BMT) from matched siblings. Median age at BMT was 8.7 years. Conditioning consisted of low-dose cyclophosphamide (CY 5 mg/kg x 4 days) and thoracoabdominal irradiation (TAI 400 cGy). Graft-versus-host disease (GVHD) prophylaxis was cyclosporin A (CsA) in 13 patients and CsA plus methotrexate in 6 patients. Antithymocyte globulin (ATG) was added in the pretransplant as well as the post-transplant period. All patients received high-dose acyclovir from day 2 pre-BMT to day 28 post BMT, and intravenous immunoglobulins (IVIG), 500 mg/kg weekly from day 7 pre-BMT to day 90 post BMT. No fungal prophylaxis was given. All patients engrafted, (median, 14 days for an absolute neutrophil count > or =0.5 x 10(9)/l; median, 37 days for platelet count > or =20 x 10(9)/l). Fourteen (74%) patients are alive with sustained engraftment and are transfusion independent. Three (16.6%) patients developed acute GVHD; none developed chronic GVHD. Five (26%) patients developed invasive fungal infections, and two (10%) developed fatal CMV disease. We believe the addition of ATG may have contributed to the increased incidence of severe life-threatening fungal and viral infections in our series. PMID- 11281383 TI - Autologous bone marrow transplantation for childhood acute lymphoblastic leukemia: a novel combined approach consisting of ex vivo marrow purging, modulation of multi-drug resistance, induction of autograft vs leukemia effect, and post-transplant immuno- and chemotherapy (PTIC). AB - In an attempt to reduce the high relapse rate associated with ABMT, five children with high-risk first CR and 19 in second or subsequent CR lacking matched family allogeneic donors underwent ABMT with chemopurged bone marrow utilizing verapamil (VPL), vincristine, and VP-16. Patients were conditioned with TBI, VPL bolus and infusion with VP-16 and cyclophosphamide. The first cohort of patients (n = 4) received only cyclosporin A (CsA). The second cohort (n = 7) received CsA and alpha interferon (total = 11 with post-transplant immunotherapy alone.) The third cohort (n = 13) received CsA and six alternating cycles of alphaIFN and chemotherapy and six additional cycles of chemotherapy (vincristine, VP-16, Ara C, prednisone) followed by G-CSF (post-transplant immune chemotherapy (PTIC)). The 2-year DFS is 42+/-10% (90% confidence interval (CI) is 26.5-58.5%) and 2 year overall survival is 54+/-10% (90% CI is 37.5-70.5%). Furthermore, patients receiving PTIC (n = 13) vs immunotherapy alone (CsA+/-aIFN) (n = 11) had a substantially better 2 year DFS and OS: 69+/-13% vs 13+/-12% and 85+/-10% vs 25+/ 15% (P = 0.008 and P = 0.06, respectively). These results suggest that the use of ABMT with chemopurging, combined with PTIC is well tolerated and may be an alternative new approach in the treatment of a subset of children with high-risk first CR or > or = second CR ALL who lack closely matched family-related allogeneic donors. PMID- 11281384 TI - Allogeneic bone marrow transplantation in children failing prior autologous bone marrow transplantation. AB - Twenty-three children with de novo acute myelogenous leukemia (AML) (n = 20), secondary AML (n = 1), or non-Hodgkin's lymphoma (NHL) (n = 2) underwent allogeneic bone marrow transplantation (alloBMT) for graft failure (n = 1) or recurrent malignancy (n = 22) between February 1992 and August 1999 following autologous BMT (ABMT). Induction chemotherapy was given to 14 patients and nine patients went directly to alloBMT. Five received marrow from matched siblings, 14 from matched unrelated donors and four from mismatched family members. Conditioning regimens included cyclophosphamide, cytarabine, and total body irradiation. Nine patients are alive disease-free between 627 and 2433 days (1.7 6.7 years) post BMT resulting in a 4-year DFS of 39%. Eight patients relapsed at a median of 206 days (range, 35-669 days) post alloBMT and all eventually died. Eight patients (two of whom also relapsed) died of RRT. Although RRT and relapse remain significant problems, a significant percentage of pediatric patients failing ABMT may be cured with alloBMT. PMID- 11281385 TI - Pancreatic insufficiency in patients with chronic graft-versus-host disease. AB - Diarrhea is a difficult diagnostic problem in patients with chronic graft-versus host disease (cGVHD) because there are many causes of it. Although intestinal involvement has been reported in early studies of untreated cGVHD, this is now a very rare presentation of the disease. In addition to other etiologies, pancreatic insufficiency should also be considered in patients with cGVHD who demonstrate malabsorption. The pathogenesis of pancreatic insufficiency in these patients is unknown. Pancreatic enzyme supplements can be very effective in treating this rare condition. PMID- 11281386 TI - Allogeneic peripheral blood stem cell transplantation results in less alteration of early T cell compartment homeostasis than bone marrow transplantation. AB - Since low T cell counts evaluated 1 month after allogeneic bone marrow transplantation (BMT) are associated with an increased risk of leukemia relapse (Powles et al., Blood 1998; 91: 3481-3486), we compared, in a randomized multicentric clinical study, the peripheral blood cells obtained 30 days after allogeneic BMT vs allogeneic G-CSF-mobilized peripheral blood stem cell transplantation (BCT) in an HLA-identical setting. T cell counts were higher 30 days after BCT (718+/-142 cells/microl, n = 20) than after BMT (271+/-53 cells/microl, n = 26, P = 0.006). However, T cells were less activated after BCT than after BMT, as demonstrated by a lower expression level of CD25 and a lower percentage of HLA-DR+ and CD95+ T cells. Furthermore, CD4+, CD8+ and CD45RA+ post BCT T cell counts correlated with the number of cells infused with the PBSC graft, while such a correlation was not observed between post-BMT counts and BM graft cell numbers, suggesting that the intensity of post-transplant peripheral lymphoid expansion and/or deletion differed between BCT and BMT. A comparison of the input of T cells expressing different CD45 isoforms with the post-transplant cell recovery further confirmed that, within the CD4+ T cell subset, post transplant expansions occurred at a higher level after BMT than after BCT, affecting mainly the CD4+ CD45RO+ subset. Altogether, our data demonstrate for the first time in a randomized setting that homeostasis of the T cell pool is less altered early after BCT than after BMT. This may have a strong impact on the graft-versus-leukemia (GVL) effect and subsequent relapse rate. PMID- 11281387 TI - Radiation-associated pneumonitis following autologous stem cell transplantation: predictive factors, disease characteristics and treatment outcomes. AB - High-dose therapy followed by autologous stem cell transplantation (ASCT) prolongs survival in patients with multiple myeloma and is relatively safe with treatment-related mortality rates of only 1-5%. Interstitial pneumonitis (IP) is normally an infrequent complication of ASCT with a reported incidence of 0-16%. Between 1992 and 1998, 94 myeloma patients at our center underwent ASCT using a high-dose regimen of etoposide (60 mg/kg), melphalan (160 mg/m2) and fractionated TBI 12 Gy. An unusually high incidence of IP (29/94 (31%)) was noted. Mortality in the IP patients was high at 45%. Patients developing IP were more frequently anemic than those who did not have pulmonary complications (hemoglobin <100 g/l) prior to transplant (P = 0.03) but no other pre-transplant factors were predictive (ie age, gender, smoking history, CMV status, pulmonary function, creatinine, beta2-microglobulin or C-reactive protein, prior cumulative chemotherapy or chest irradiation). A significantly lower IP rate was noted in 32 contemporaneous myeloma control patients conditioned with BU-CY without TBI at our center (3/32 (9%); P=0.03) and in 32 lymphoma control patients conditioned with the same melphalan and etoposide regimen minus the TBI (2/32 (6%); P = 0.003). In contrast, when using the same TBI-containing regimen in 32 concurrently treated lymphoma patients, an increase in IP similar to that seen in our myeloma cohort (7/32 (22%); P = 0.3) was noted. This strongly suggests that TBI is the predominant factor contributing to lung toxicity. We conclude that radiation-associated pneumonitis cannot be easily predicted by pretransplant variables. Therefore surveillance, early recognition and prompt therapy are recommended. PMID- 11281388 TI - Lack of known hepatitis virus in hepatitis-associated aplastic anemia and outcome after bone marrow transplantation. AB - Viral infection has been shown to induce aplastic anemia, unidentified types of hepatitis being the most common cause for aplastic anemia-associated viral hepatitis. The survival rate for this group of patients after bone marrow transplantation with stem cells from an HLA-matched sibling is not well known. The aim of this study was to determine the prevalence of hepatitis G virus (HGV) and transfusion transmitted virus (TTV) infection in non-A, non-B, non-C hepatitis associated-aplastic anemia (HAAA) patients, and to define the role of bone marrow transplantation (BMT) as a therapeutic modality for this disease. Sixty-eight patients (43 males and 25 females) with aplastic anemia, underwent allogeneic BMT at the Hadassah University Hospital between 1981 and 1997. Onset of hepatitis was defined as jaundice and elevated alanine aminotransaminase (ALT) levels. Onset of aplastic anemia was defined as the first date on which varying degrees of pancytopenia occurred: hemoglobin level below 10 g/dl, WBC below 2 x 10(9)/l and low platelet count 10 x 10(10)/l. Serial serum samples from HAAA patients were assayed for virological and/or serological markers of hepatitis A, B, C, D, E, G viruses, TTV and parvovirus B19. Seventeen of the 68 patients with aplastic anemia (25%) suffered from hepatitis, 12 males and five females, ages 5 to 36 years. The mean interval between onset of hepatitis and first indication of aplastic anemia was 62 days (range 14-225 days). The development of aplastic anemia was unrelated to age, sex or severity of hepatitis. Ten of the 17 patients (59%) achieved complete ALT recovery prior to the diagnosis of aplastic anemia. Serum samples were available for 15 patients; none had evidence of acute or active hepatitis A, B, C, D, E, G and TTV virus infection at the time of diagnosis. Parvovirus B19 DNA sequences were not detectable in 10 of 12 tested cases; two positive results were detected in serum samples obtained after blood transfusion, making the analysis of these positive results difficult. All 17 patients underwent BMT. The mean post-BMT follow-up period was 38 months (range 1 day-123 months), five patients (30%) died 1 to 160 days post BMT, and 12 (70%) are alive 31 to 123 months after BMT. Relapsing hepatitis was not observed in any of the patients. In conclusion, HAAA is a disease of the young and the etiologic agent associated with HAAA remains unknown. HGV, TTV and parvovirus B19 sequences were not detected in any of the HAAA cases. The survival rate after BMT with stem cells from an HLA-matched sibling is similar to that for patients with non hepatitis-associated aplastic anemia. PMID- 11281389 TI - Yield of bronchoalveolar lavage in ventilated and non-ventilated children after bone marrow transplantation. AB - A study was undertaken to retrospectively evaluate the yield of bronchoalveolar lavage (BAL) in a single-institution series of children after bone marrow transplantation (BMT) and to compare the yield of BAL between the ventilated and nonventilated patients. We reviewed charts of 52 consecutive children after BMT who underwent BAL. Thirty patients (41 BALs) were nonventilated (group 1) and 33 patients (45 BALs) were ventilated for respiratory failure (group 2). Eleven patients were included in both groups. BAL was performed a median of 255 and 28.5 days after BMT in groups 1 and 2, respectively (P < 0.001). Group 1:17 pathogens were isolated from 13 BALs; a single pathogen from 10 BALs. Group 2:15 pathogens were isolated from 14 BALs (31.1% positive). Viruses were isolated from 13 BALs in group 2. A severe complication of BAL occurred in only one patient from group 1 (1.1%). Open lung biopsies were performed in one patient in group 1 and eight patients in group 2. The histological findings correlated with the BAL findings in 66.7%. In conclusion, there was no difference in the yield of BAL between the groups. Therapy was changed in one third of the patients dictated by the BAL findings. The risk of severe complications was relatively low. A good correlation between open lung biopsy (OLB) and BAL was found. PMID- 11281390 TI - Urinary trypsin inhibitor concentration can predict the immunological insult of chemotherapy and complications after bone marrow transplantation. AB - Urinary trypsin inhibitor has attracted attention as an index of the systemic inflammatory response syndrome. In this study, the urine concentration of trypsin inhibitor was measured to compare the immunological insult of conventional chemotherapy and conditioning chemotherapy for bone marrow transplantation. We also investigated whether urinary trypsin inhibitor was a useful index of the complications and outcome of bone marrow transplantation. Urinary trypsin inhibitor concentration was determined before chemotherapy, on the day after finishing chemotherapy (day 0 of transplantation), and during recovery of the white cell count, in 17 patients (seven receiving conventional chemotherapy and 10 receiving conditioning for bone marrow transplantation). Urinary trypsin inhibitor concentrations were significantly higher after conditioning for bone marrow transplantation than after conventional chemotherapy (P < 0.001), indicating that conditioning was more invasive. After bone marrow transplantation, the incidence of severe complications and the mortality rate were higher in patients whose urinary trypsin inhibitor concentrations rose during recovery of the white cell count. Comparison of urinary trypsin inhibitor concentrations suggested that conditioning for bone marrow transplantation was more invasive than conventional chemotherapy. This study also suggested that the urine concentration of trypsin inhibitor could be useful for predicting the risk of complications and outcome of bone marrow transplantation. PMID- 11281391 TI - Aerobic bacterial and fungal infections in peripheral blood stem cell transplants. AB - Allogeneic and autologous peripheral blood stem cell transplants are frequently complicated by infections. This study was performed to evaluate early and late infections in 74 patients who underwent peripheral blood stem cell transplantation (PBSCT). Fifty-eight patients received allogeneic and 16 autologous PBSCT. All patients received fluconazole, ciprofloxacin and acyclovir prophylaxis. 93.1% of alloPBSCT patients and 87.5% of autoPBSCT patients developed fever. Febrile episodes were commonly seen in the week of transplantation (66%). There was a median of 3 days with fever in alloPBSCT, and 2 days in autoPBSCT. Period of neutropenia was 15 days for AlloPBSCT and 12 days for AutoPBSCT. The microbiological identification rate was 47% (32/68). Gram positive infections dominated the early period (50%) and Gram-negative bacterial infections dominated the late period (50%). All our patients had Hickman-type catheters and 26 infections involving catheters were seen. Sixteen occurred in the early, and 10 in the late period. Ten of 14 (71.4%) late bacterial infections were catheter-related. The dominance of Gram-positive infections and high rates of methicillin resistance warranted the use of vancomycin extensively. Surveillance cultures were found to be useful in selected patients. Although slime factor is an important virulence factor, there was no difference between slime factor positive and negative coagulase-negative staphylococci isolated during infections. In conclusion, febrile episodes are the most frequent complication of PBSCT and Gram-positive microorganisms remain the main pathogen in these patients because of catheter use, mucositis and ciprofloxacin prophylaxis. Methicillin resistance is increasing and glycopeptides remain the only choice for treating such infections. Although the infection rate is high, measures taken to prevent and treat infections result in very low rates of mortality from infection in PBSCT patients. PMID- 11281392 TI - Opsonophagocytic activity against Streptococcus pneumoniae type 19F in allogeneic BMT recipients before and after vaccination with pneumococcal polysaccharide vaccine. AB - Opsonophagocytic activity (OPA) of pneumococcal polysaccharide (Pnc PS) antibodies in vitro, a measure of antibody functional activity, usually correlates with specific IgG antibody concentrations measured by an EIA method. In order to investigate the functional activity of specific Pnc PS antibodies, we determined IgG antibodies to Pnc PS type 19F by an EIA method and OPA against Pnc type 19F by a killing assay in a randomized study, where 23 adult allogeneic BMT recipients were vaccinated with Pnc PS vaccine at 8 months (early group) and 21 recipients at 20 months (late group) after transplantation. Serum samples drawn before BMT, before vaccination, and at 1 and 16 months after vaccination were available from 27, 35, 34 and 30 patients, respectively. The geometric mean anti Pnc 19F concentrations were 4.3, 1.3, 1.6 and 1.3 microg/ml in the early group and 3.8, 0.9, 0.6 and 0.6 microg/ml in the late group before transplantation, before vaccination, and at 1 and 16 months after vaccination, respectively. OPA (titre > or = 8) was found in 10/27, 5/35, 5/34 and 6/30 patients before BMT, before vaccination, and at 1 and 16 months after vaccination, respectively. The specific IgG antibody concentration and OPA correlated with each other before BMT, and in the early group patients before and at 1 month after vaccination. The results demonstrate that after Pnc PS vaccination allogeneic BMT recipients have antibodies with low functional activity to a Pnc PS antigen associated with low specific IgG responses. There is a need to study new Pnc conjugate vaccines in multi-dose schedules for their capacity to elicit higher specific antibody concentrations with high OPA in BMT recipients. PMID- 11281393 TI - Bone marrow transplantation in a case of severe, type II congenital dyserythropoietic anaemia (CDA II). AB - Type II congenital dyserythropoietic anaemia (CDA-II or HEMPAS) is an autosomal recessive disorder, representing the most frequent form of congenital dyserythropoiesis. It is characterised by normocytic anaemia, variable jaundice and hepato-splenomegaly. Gallbladder disease and secondary haemochromatosis are frequent complications. We report a case characterised by severe transfusion dependent anaemia. The proband inherited CDA-II in association with beta thalassaemia trait. Splenectomy did not abolish the transfusion dependence and this, in association with poor compliance to iron-chelation therapy, prompted us to consider bone marrow transplantation (BMT) from his HLA-identical sibling. The preparative regimen included busulfan, thiotepa and fludarabine, and graft-versus host disease prophylaxis consisted of cyclosporin A and short-term methotrexate. Engraftment of donor cells was prompt and the post-transplant course uncomplicated. The patient is alive and transfusion-independent 36 months after allograft. This is the first case of severe CDA-II to undergo BMT. Analysis of this pedigree suggests that interaction with beta-thalassaemia enhanced the clinical severity of CDA-II, making BMT an attractive therapy for patients with transfusion dependence. PMID- 11281394 TI - Fatal sepsis due to mycobacterium tuberculosis after allogeneic bone marrow transplantation. AB - Mycobacterium tuberculosis is a serious, but rare infectious complication after allogeneic bone marrow transplantation. We describe a case of fatal sepsis due to Mycobacterium tuberculosis after allogeneic bone marrow transplantation for Philadelphia chromosome-positive ALL. The diagnosis was made after BAL. Although broad-spectrum antituberculous therapy was started immediately after diagnosis, blood cultures became positive for Mycobacterium tuberculosis. The patient developed severe pyrexias and finally died of multi-organ failure. Rapid progression of mycobacterial infection should be considered in patients post BMT with unexplained fever, particularly in patients from endemic areas. PMID- 11281395 TI - Porphyria cutanea tarda in a patient with post-transplant MDS. AB - We report a case of porphyria cutanea tarda associated with myelodysplastic syndrome in a patient after high-dose chemotherapy and peripheral blood stem cell transplantation for recurrent non-Hodgkin's lymphoma. PMID- 11281396 TI - Graft failure in a patient with systemic lupus erythematosus (SLE) treated with high-dose immunosuppression and autologous stem cell rescue. AB - A 32-year-old female with WHO grade IV, dialysis dependent, lupus nephritis was treated with high-dose immunosuppression and autologous stem cell rescue. Stem cells were mobilized with cyclophosphamide (CY) and G-CSF, and 4.07 x 10(6) CD34+ cells/kg were obtained after CD34+ cell selection using the CellPro column. The preparative regimen consisted of CY, and antithymocyte globulin (ATG), with methylprednisolone. After apparent primary engraftment of neutrophils on day 9, the patient developed recurrent neutropenia on day 19. She showed no evidence of engraftment by day 35, and back-up unmanipulated stem cells were given without effect. Subsequently, she received unmanipulated peripheral stem cells (2 x 10(6) CD34+ cells/kg) from an HLA-identical sibling. The patient remained pancytopenic and expired on day 62 from disseminated fungal infection. An autopsy revealed no evidence of hematopoietic recovery. Progenitor cell assays were performed with the patient's stem cells, which were collected prior to transplantation, and serum collected day 27. Morphologic examination of the patient's cell colonies grown in the presence of her serum revealed abnormal shapes and non-adherent cells. There were significantly fewer BFU-e colonies and a trend toward fewer CFU GM colonies with the patient's cells and serum compared to normal donor cells. We concluded that a substance present in her serum mediated graft failure and prevented engraftment after additional stem cell infusions. PMID- 11281397 TI - Rituximab can be useful as treatment for minimal residual disease in bcr-abl positive acute lymphoblastic leukemia. AB - We report the results of administering CD20 monoclonal antibody (MoAb) in a 32 year-old man with bcr-abl-positive acute lymphoblastic leukemia. Morphological complete remission was achieved after two lines of chemotherapy with persistence of blast cells (2%) in flow cytometric analysis of marrow cells. Since no HLA matched donor for allogeneic bone marrow transplantation (BMT) was found, anti CD20 MoAb therapy was administered for in vivo marrow purging, prior to autologous peripheral blood stem cell (PBSC) harvest and transplantation. After MoAb therapy <0.1% of blast cells were observed and the molecular abnormality (bcr-abl gene rearrangement) disappeared. PMID- 11281398 TI - Severe cutaneous ulceration following treatment with thalidomide for GVHD. AB - We report two cases of severe leg ulcerations in patients being treated with thalidomide for graft-versus-host disease following bone marrow transplantation. Local wound care and debridement were attempted, but one patient required skin grafting to ensure healing. We propose that this complication may be due to the antiangiogenic properties of thalidomide and urge careful attention to skin breakdown in patients being treated with this compound. PMID- 11281400 TI - Comparative institutional response to economic policy managed competition and governmentality. AB - This article provides a comparative conceptual framework for understanding why so many governments found economic policies based on managed competition attractive and yet dangerous to implement. The framework conceptualizes governments as a kind of organizational complex and thus governments as an international population of organizations, each embedded in a state that tries to harness and direct behaviour through what Foucault called "governmentality". This nascent concept is made more robust here and joined with Fligstein's historical research on the response of leading organizations when fundamental change threatens a population of organizations, by embracing a new conception of control that allows them to re-establish their control and pre-eminence. Fligstein studied corporations, but his model can be fruitfully extended to governments. Economic sociology has not to date been able to do much comparative research on institutional responses to economic policy; but this set of case studies and conceptual framework provide such an opportunity. PMID- 11281399 TI - Myositis, polyserositis with a large pericardial effusion and constrictive pericarditis as manifestations of chronic graft-versus-host disease after non myeloablative peripheral stem cell transplantation and subsequent donor lymphocyte infusion. AB - The clinical features of chronic graft-versus-host disease (cGVHD) following a non-myeloablative peripheral blood stem cell (PBSC) transplant may differ from those that occur after a conventional allograft. We describe a man with Hodgkin's disease refractory to chemotherapy and radiotherapy who was transplanted from an HLA-identical brother, who developed cGVHD characterised, in particular, by polymyositis, polyserositis with a large pericardial effusion and constrictive pericarditis, 1 month after donor lymphocyte infusion for relapsed disease. Constrictive pericarditis has not been previously reported after a conventional allograft, and none of these features have been reported after a non myeloablative transplant. The course of cGVHD necessitated potent immunosuppression leading to the presumed loss of graft-versus-lymphoma (GVL) effect. PMID- 11281401 TI - Managed competition, governmentality and institutional response in the United Kingdom. AB - This article traces the use of managed competition policy to transform the NHS from an administered public service to a set of interlocking markets and contracts. It reviews the overlooked origins of managed competition in the new managerialism and explains the relationship between managed competition and the cost crisis of the NHS by extending Foucault's concept of governmentality to revise the concept of the state. The paper then describes how the government structured health care markets, using managed competition as an instrument of governmentality. It summarises institutional responses by health authorities, hospital trusts, and GP fundholders. The terms "master institution", "dictated competition" and "coercive partnering" are introduced as new concepts for economic sociology and as strategies of governmentality. Implementation, however, led to resistance, opposition and eventual abandonment of managed competition as too disruptive and costly. Yet, this analysis contends, managed competition has left an enduring legacy of accountability to purchasers in economic terms such as efficiency, transaction costs, and cost effectiveness. The policies of the new government are based on coercive partnering and doctor-based "commissioning". This and the Internet imply revolutionary changes for the health professions and the delivery of health care services through networks of moebius-strip organisations interacting in flexible sequences and subject to communitarian pressures. PMID- 11281402 TI - The marginal success of regulated competition policy in the Netherlands. AB - In the second half of the 1980s the government in the Netherlands adopted a regulated competition policy as part of a comprehensive programme designed to restructure the health care system. The programme was a product of its social and political context, promoted by a group of political entrepreneurs and created to improve efficiency. Despite the initial political support and a long political debate the government had to acknowledge by 1992 that the restructuring would not take place. But changes fostered limited competition between sickness funds and more extensive competition in the small market for supplementary policies. This, however, has not led to sickness funds becoming powerful purchasers that forced hospitals and doctors to improve their efficiency. Rather, they compete for subscribers, become part of large insurance conglomerates, and market more supplementary options. Culturally, health care institutions have become more entrepreneurial, taken up more business concepts, and made the language of markets, products and consumer sovereignty more common. The impact of these changes on the health care system is still unknown, but they create pressure for more health care services, leaving the government with problems that equal those of the 1980s. PMID- 11281403 TI - Cost containment, solidarity and cautious experimentation: Swedish dilemmas. AB - This paper uses secondary data analysis and a literature review to explore a "Swedish Dilemma": Can Sweden continue to provide a high level of comprehensive health services for all regardless of ability to pay--a policy emphasizing "solidarity"--or must it decide to impose increasing constraints on health services spending and service delivery--a policy emphasizing "cost containment?" It examines recent policies and longer term trends including: changes in health personnel and facilities; integration of health and social services for older persons; introduction of competition among providers; cost sharing for patients; dismantling of dental insurance; decentralization of government responsibility; priority settings for treatment; and encouragement of the private sector. It is apparent that the Swedes have had considerable success in attaining cost containment--not primarily through "market mechanisms" but through government budget controls and service reduction. Further, it appears that equal access to care, or solidarity, may be adversely affected by some of the system changes. PMID- 11281404 TI - Adopting and adapting managed competition: health care reform in Southern Europe. AB - A new paradigm appeared in Europe in the early 1990 s regarding the reform of health care systems. This paradigm has come to be known as the managed competition paradigm, among other terms. First introduced in Great Britain, it entails the separation of the financing/purchasing and providing functions, so that competition among providers is enhanced, while maintaining universal access and public financing, at least in principle. This article explores to what extent such paradigm has been emulated within the Greek, Italian, Portuguese and Spanish health care systems. Reform in the direction of managed competition may be ascertained in all four countries. However, each country has emphasized different aspects of the paradigm, and the degree and rhythm of implementation of reform has varied. The article considers the circumstances under which the new paradigm was born, and its main characteristics; analyzes actual reforms in Southern European countries; and provides a tentative explanation of the diffusion mechanisms. It concludes that the crucial factor explaining the different paths of policy adoption and adaptation is the character of the initial health care system. PMID- 11281405 TI - Implementing managed competition in Israel. AB - As of January 1, 1995, Israel's National Health Insurance (NHI) Law laid the foundations for regulating competition among the country's four private, not-for profit sick funds. Prior to NHI the sick funds (SFs) had competed without governmental control. Extensive research on NHI implementation and the behavior of the sick funds (SFs) after passage of NHI reveals a paradoxical development: The NHI bill drew on the rhetoric of managed competition and did indeed establish a legal and structural framework for regulating competition among the SFs. Nevertheless, in practice, SF autonomy was constrained and competition over provision of statutory care was limited. Rather than fostering competition, the main thrust of the NHI reforms was to enhance central government's control over SF expenses in order to constrain government expenditures. The NHI reforms did encourage the SFs to cut costs and make visible service improvements. However, the reforms did not lead the SFs to reorganize, expand the scope of their services, or improve clinical quality, as the reformers had hoped. Nor did the reforms help eliminate the SF's operating deficits or insure financial stability for the whole health system. Furthermore, the reforms had unanticipated and undesired outcomes, including aggressive and illegal marketing by SFs and collaboration among SFs to restrict the extent of care provided under compulsory insurance. The Israeli case suggests that the theory of managed competition contains unrealistic assumptions about the types of competitive behavior that result from exposure to managed competition and the capacity of government and health providers to monitor quality. In addition to stemming from universal limitations to the managed competition model, the implementation pattern in Israel reflects local, historical forces and the interplay of Israel's powerful health system actors. PMID- 11281406 TI - Transforming health sectors: new logics of organizing in the New Zealand health system. AB - This paper develops a relational analysis (drawing on the insights of historical institutionalism and economic sociology) of the ongoing process of radical health sector restructuring in New Zealand. The original 'reforms', based on a 'purchaser provider' split, are outlined so as to emphasize their politically consequential ambiguity: was restructuring about revitalizing an essentially public health system or about creating the basis for an eventually private health system with a residual state role? The actual process of restructuring is then traced, emphasizing the responses it has evoked from differently situated actors within the health sector as this is entwined with the political system. The focus is on explaining the largely unintended consequences that have resulted, including the abandonment or significant modification of most of the originally enacted forms of organization together with the emergence of new organizational forms, initiated by providers, and largely unanticipated by the restructurers. PMID- 11281407 TI - Managed care in Latin America: the new common sense in health policy reform. AB - This article presents the results of the comparative research project, "Managed Care in Latin America: Its Role in Health System Reform." Conducted by teams in Argentina, Brazil, Chile, Ecuador, and the United States, the study focused on the exportation of managed care, especially from the United States, and its adoption in Latin American countries. Our research methods included qualitative and quantitative techniques. The adoption of managed care reflects the process of transnationalization in the health sector. Our findings demonstrate the entrance of the main multinational corporations of finance capital into the private sector of insurance and health services, and these corporations' intention to assume administrative responsibilities for state institutions and to secure access to medical social security funds. International lending agencies, especially the World Bank, support the corporatization and privatization of health care services, as a condition of further loans to Latin American countries. We conclude that this process of change, which involves the gradual adoption of managed care as an officially favored policy, reflects ideologically based discourses that accept the inexorable nature of managed care reforms. PMID- 11281408 TI - Biofantasies: genetics and medicine in the print news media. AB - The contemporary news media is an important site for exploring the diverse and complex cultural images of genetics and its medical possibilities, and of the mechanisms by which these images are (re) produced and sustained. This article investigates how the print news media 'frames' stories on genetics and medicine. It is based on a discourse analysis of articles appearing in three Australian newspapers in the late 1990s. Gene stories were found to be prominent in each of the newspapers, and to emphasise the medical benefits of genetic research. Stories frequently cite and quote scientists, who explain the nature and significance of the research and/or its implications for treatment or prevention. Many stories focus on new genetic discoveries, and portray genetic researchers as involved in a quest to unlock nature's secrets. Stories of hope, and depictions of geneticists as warriors or heroes, appear regularly. The positive vision of genetics is supported by the use of particular metaphors, accompanying illustrative material, 'human interest' stories, and reference to credible sources. There is rarely mention of the influence of non-genetic factors and 'multifactorial' interactions on disorders, or questioning of the goals, direction, methods, or value of genetic research. Scientists made extensive use of the media in their efforts to maintain a positive image of research in the face of public concerns about scientists 'going too far', following the announcement of the cloning of Dolly. Boundaries were drawn between 'therapeutic cloning'--implicitly defined as 'good', useful, and legitimate--and 'reproductive cloning'--seen as 'bad', dangerous, and illegitimate. By framing news stories as they do, the print news media are likely to exert a powerful influence on public responses to health problems. With new genetic technologies becoming more integrated in preventive medicine and public health, it is important to investigate how news stories help shape the agenda for public debate. PMID- 11281409 TI - Does childhood health affect chronic morbidity in later life? AB - Our analysis examines whether childhood health has long-term and enduring consequences for chronic morbidity. As a part of this analysis, we address two methodological issues of concern in the literature. Is adult height a surrogate for childhood health experiences in modeling chronic disease in later life? And, are the effects of adult socioeconomic status on chronic disease overestimated when childhood health is not accounted for? The analysis is based on a topical module to the third wave of the Health and Retirement Study, a representative survey of Americans aged 55-65 in 1996. Our results support the hypothesis that poor childhood health increases morbidity in later life. This association was found for cancer, lung disease, cardiovascular conditions, and arthritis/rheumatism. The associations were highly persistent in the face of statistical controls for both adult and childhood socioeconomic status. No support was found for using adult height as a proxy for the effects of childhood health experiences. Further, the effects of adult socioeconomic status were not overestimated when childhood health was excluded from the explanatory models. Our results point to the importance of an integrated health care policy based on the premise of maximizing health over the entire life cycle. PMID- 11281410 TI - Ethnic and class differences in health in relation to British South Asians: using the new National Statistics Socio-Economic Classification. AB - The paper examines the use of the new measure of social class in the UK, the National Statistics Socio-Economic Classification (NS-SEC) and other socio economic variables in explaining differences in health between British South Asians and the majority White population. There are a number of hypotheses which try to explain ethnic differences in health and yet there have been relatively few empirical studies which test the explanatory value of these hypotheses. Cross sectional data from the fourth National Survey of Ethnic Minorities (1993-1994) with 2860 white, 1268 Indian and 1771 Pakistani and Bangladeshi adult respondents are analysed. The associations of self-rated health with ethnicity, social class, local area deprivation and standard of living are analysed. Pakistani and Bangladeshi respondents have the poorest self-rated health, followed by Indians. Differences in self-rated health between ethnic groups reduce to non-significance after adjusting for social class, local area deprivation and standard of living. There is some evidence of social class differences in the health of Indians and not much evidence for Pakistanis and Bangladeshis. The NS-SEC is useful in explaining ethnic differences in health. The poorer health of Indians, Pakistanis and Bangladeshis compared to Whites may be largely understood in terms of factors related to occupational social class, material living conditions and local area deprivation. PMID- 11281411 TI - Socioeconomic status and health among older adults in Thailand: an examination using multiple indicators. AB - A survey of 14,000 Thais, 50 and older, is utilized to test hypotheses about the association between socioeconomic status (SES) and health previously derived through observation of Western populations. Central among hypotheses is the notion that an inverse association can be uniformly detected across a matrix of SES indicators and health outcomes. Indicators of SES in this study include the traditional education, occupation and income measures, and a measure of household possessions. This later indicator may be particularly useful in a non-Western setting such as Thailand. Health outcomes were derived in order to represent a subjective, a social, and a medical conceptualization of health and included measures for self-assessed health, functional disorders and chronic health conditions. All health indicators were dichotomously coded as existence or non existence. Also tested are associations throughout ordered categories of SES. Results generally support hypotheses. Unadjusted for other SES covariates, all four indicators of status strongly related to two of the three health measures, when controlling for other important covariates. When adjusted for each other, a number of the associations remained strong. There were also several notable exceptions. Little association existed between SES and chronic health disorders. The results may serve to highlight the distinct social environment acting upon the association. PMID- 11281412 TI - X-linked juvenile retinoschisis: mutations at the retinoschisis and Norrie disease gene loci? AB - Juvenile retinoschisis (RS) and Norrie disease (ND) are X-linked recessive retinal disorders. Both disorders, in the majority of cases, are monogenic and are caused by mutations in the RS and ND genes, respectively. Here we report the identification of a family in which mutations in both the RS and ND genes are segregating with RS pathology. Although the mutations identified in this report were not functionally characterized with regard to their pathogenicity, it is likely that both of them are involved in RS pathology in the family analyzed. This suggests the complexity and digenic nature of monogenic human disorders in some cases. If this proves to be a widespread problem, it will complicate the strategies used to identify the genes involved in diseases and to develop methods for intervention. PMID- 11281413 TI - Amino-acid substitutions in the IKAP gene product significantly increase risk for bronchial asthma in children. AB - The complex etiology of bronchial asthma (BA), one of the most common inflammatory diseases throughout the world, involves a combination of various genetic and environmental factors. A number of investigators have undertaken linkage and association studies to shed light on the genetic background of BA, but the genetic aspects of this disease are still poorly understood. In the course of a project to screen the entire human genome for single nucleotide polymorphisms (SNPs) that might represent useful markers for large-scale association analyses of common diseases and pharmacogenetic traits, we identified six SNPs within the gene encoding I-kappaB-associated protein (IKAP), a regulator of the NF-kappaB signal pathway. Most of these SNPs were in linkage disequilibrium with each other. We observed a strong allelic association between BA in childhood and two of the SNP sites, T3214A (Cys1072Ser) and C3473T (Pro1158Leu); P = 0.000004 for T3214A and P = 0.0009 for C3473T. T3214A was also associated with BA in adult patients (P = 0.000002), but C3473T was not (P = 0.056). To confirm the above results, we compared estimated frequencies of haplotypes of the six SNPs between BA patients and controls. We found a strong association between BA in childhood and a specific haplotype, TGAAAT, that involved two amino-acid substitutions (819T, 2295G, 2446A, 2490A, 3214A, and 3473T; P = 0.00004, odds ratio, 2.94; 95% confidence interval [CI], 2.48-3.4). On the other hand, haplotype TACGTC, which differed from the TGAAAT haplotype in the last five nucleotides, was inversely correlated with the BA phenotype (P = 0.002; odds ratio, 9.83; 95% CI, 8.35-11.31). These results indicated that specific variants of the IKAP gene, or a variant in linkage disequilibrium with the TGAAAT haplotype, might be associated with mechanisms responsible for early-onset BA. PMID- 11281414 TI - Phylogenetic analysis of mtDNA haplogroup TJ in a Finnish population. AB - An association between mitochondrial DNA (mtDNA) mutations 11778G>A and 14484T>C and mtDNA haplogroup J suggests that this haplogroup harbors substitutions capable of modifying the phenotype of Leber's disease. Our knowledge of the compilation of substitutions in haplogroup J is based on only a small number of complete mtDNA sequences, however. We constructed phylogenetic networks for mtDNA haplogroup TJ that were based on the sequence of the complete coding region and the hypervariable segment I, respectively, in 28 Finnish samples. The networks revealed a subdivision of the haplogroup into subclusters T1, T2, J1, and J2, while comparison of the two networks suggested nine fast evolving nucleotide sites in the hypervariable segment I. Genotypes of patients harboring 11778G>A or 14484T>C were obtained from the literature and were then placed in the network. Only four substitutions were found to be common to the patients, but none of these was unique to haplogroup J. If increased penetrance of the 11778G>A and 14484T>C mutations in patients belonging to haplogroup J is assumed, combinations of ancient substitutions must be implicated. PMID- 11281415 TI - Molecular cloning, tissue expression, and chromosomal assignment of a novel gene encoding a subunit of the human signal-recognition particle. AB - Human cancers derived from breast, esophageal, or ovarian tissues frequently show allelic losses on chromosome band 17q25. Moreover, a locus responsible for hereditary focal nonepidermolytic palmoplantar keratoderma, a condition associated with esophageal cancer (TOC; tylosis with oesophageal cancer), has been mapped to the same band. During efforts to sequence, by shotgun methods, a 1 Mb target region that we had defined as the DNA segment harboring the putative tumor suppressor gene(s) involved in these events, we identified a novel cDNA. The full-length cDNA is 2495bp long and is expressed predominantly in skeletal muscle, heart, kidney, and placenta. The predicted product, a 627-amino-acid protein, exhibited significant sequence homology to the canine 68-kd subunit of the signal recognition particle that has been implicated in the transport of secreted and membrane proteins to the endoplasmic reticulum for proper processing. We confirmed the location of this gene at chromosome 17q25.1 by radiation-hybrid mapping and by fluorescence in situ hybridization. PMID- 11281416 TI - Analysis for microdeletions of Y chromosome in a single spermatozoon from a man with severe oligozoospermia. AB - Since the association between Y chromosome deletions and spermatogenic failure was demonstrated in 1976, there have been many reports of Y chromosome microdeletions. Peripheral blood lymphocytes (PBLs) have been used for the analysis because the method is convenient, materials are easy to obtain, and PBL genomic DNA is similar to that of germ cells such as spermatozoa. However, PBLs originate from somatic tissue, not from germ cells. In this study, we analyzed 30 spermatozoa in semen ejaculated by an infertile male with Y chromosome microdeletions, while 50 spermatozoa from a normal fertile male were used as a control. The same Y chromosome microdeletion as that found in PBL was identified in each of the 12 spermatozoa which contained the Y chromosome in the infertile patient. These results indicated that spermatozoa (germ cells) had the same Y chromosome microdeletion as PBL (somatic cells). This supports the conjecture that microdeletions are transmitted to the next generation via the treatment of infertility by intracytoplasmic sperm injection. PMID- 11281417 TI - Y chromosome compound haplotypes with the microsatellite markers DXYS265, DXYS266, and DXYS241. AB - Two newly developed microsatellite markers on Yp11 (DXYS265) and Yq11.21 (DXYS266) and our previously reported marker, on Yp11 (DXYS241), were typed by triplex polymerase chain reaction (PCR) in 102 Japanese, 18 white American, and 17 black American males. The DXYS265 locus revealed three alleles, the DXYS266 locus showed two alleles, while the DXYS241 locus showed five alleles. Nine different compound haplotypes were observed among the males. Of these, two haplotypes were common to all groups, while four were limited to Japanese. Pedigree analysis of 61 Japanese families revealed no mutations of these loci. The triplex PCR developed in this study, as well as the new loci, are useful for tracing paternal lineages in human migration studies and population analysis, in addition to Y chromosome evolutionary studies. PMID- 11281418 TI - A sequence change (Arg158Gln) in the leucine zipper-like motif region of the MYOC/TIGR protein. AB - The myocilin/trabecular meshwork-inducible glucocorticoid response (MYOC/TIGR) gene was identified as a gene that caused open angle glaucoma (OAG). Single strand conformation polymorphism analysis and subsequent sequence analysis were performed for the MYOC/TIGR gene in 120 unrelated Japanese OAG patients with increased intraocular pressure (IOP), 116 unrelated OAG patients without increased IOP, and 106 unrelated control subjects without glaucoma. An Arg158Gln sequence change in the leucine zipper-like motif (LZM) region in the myosin homology domain was found in 2 OAG patients with or without increased IOP, and in a 56-year-old control subject without glaucoma. This is the first report of missense sequence change in the LZM region of the MYOC/TIGR protein in subjects showing various phenotypes, including a control subject. These findings suggest that Arg158Gln in the LZM region is probably a rare nondisease-causing polymorphism, despite its important role in this region, because it was found in a control subject, although Arg158Gln was previously reported as a probable disease-causing mutation. PMID- 11281419 TI - Identification of DMC1, a novel gene in the TOC region on 17q25.1 that shows loss of expression in multiple human cancers. AB - Frequent allelic losses within chromosomal band 17q25.1 in a variety of human cancers have suggested the presence of one or more tumor suppressor genes in this region. Furthermore, a genetic locus responsible for familial focal nonepidermolytic palmoplantar keratoderma, a condition associated with cancer of the esophagus, lies in the same region. This esophageal-cancer susceptibility locus, TOC (tylosis with oesophageal cancer), might be a target of deletions at 17q25.1 in multiple types of malignancy. Using the reverse transcriptase polymerase chain reaction (RT-PCR) to examine cancer cell lines for alterations in the expression of transcripts from this portion of 17q, we identified a novel gene that we designated DMC1 (downregulated in multiple cancer-1). The full length cDNA is 3293bp long. Its putative product is an integral membrane protein of 788 amino acids, belonging to the class of so-called 'inside-out" membrane proteins; it lacks a signal sequence but contains an N-terminal cytoplasmic domain, a single transmembrane peptide, and a C-terminal extracellular domain. We documented loss of expression of DMC1 in 2 of 10 breast-cancer cell lines, in 7 of 10 cervical-cancer lines, in 7 of 13 hepatocellular-cancer lines, in 3 of 7 lung-cancer lines, in 3 of 6 thyroid-cancer lines, in 2 of 6 gastric-cancer lines, and in 2 of 4 renal cell-cancer lines. Our results suggest that loss of expression of the DMC1 gene at 17q25.1 may play an important role in the development of cancers in a broad range of human tissues. PMID- 11281420 TI - Overestimated frequency of a possible emphysema-susceptibility allele when microsomal epoxide hydrolase is genotyped by the conventional polymerase chain reaction-based method. AB - A recent association study suggested that the His113 variant of microsomal epoxide hydrolase (mEPHX) may confer a risk for development of emphysema, presumably by increasing susceptibility to smoking injury. Before considering a possible role of this enzyme in pulmonary disease, we attempted to characterize the genetic polymorphism further. The Tyr/His113 polymorphism within exon 3 of mEPHX was initially examined in 62 healthy individuals by conventional methods involving polymerase chain reaction (PCR)-based determination of a restriction fragment length polymorphism (RFLP). Genomic nucleotide sequences, including the polymorphic site and the downstream primer sequence, were further analyzed in 95 unrelated, healthy Japanese volunteers by single-stranded conformation polymorphism (SSCP) analysis and direct sequencing. Genotyping by the first method (PCR-RFLP) revealed that the allelic distribution in our test population apparently deviated from Hardy-Weinberg equilibrium. Sequence analysis showed that a synonymous nucleotide substitution, AAG to AAA (Lys119), was located just within the published primer site. The AAA at codon 119 was present only in alleles with Tyr113, and its frequency reached 0.31 in our panel of 190 Japanese alleles. This substitution potentially hampered PCR amplification because of the nucleotide mismatch, with the result that the frequency of the Tyr113 variation was underestimated. The frequency of His113, a possible emphysema susceptibility allele of the mEPHX gene, was thus overestimated when human DNA samples were genotyped in the conventional way. Depending on the population(s) tested, this anomaly could represent a pitfall for PCR-based association studies. PMID- 11281421 TI - JSEM: a framework for identifying and evaluating indicators. AB - There are two issues in indicator development that have not been adequately addressed: (1) how to select an optimal combination of potentially redundant indicators that together best represent an endpoint, given cost constraints; (2) how to identify and evaluate indicators when the endpoint is unmeasured. This paper presents an approach to identifying and evaluating combinations of indicators when the mathematical relationships between the indicators and an endpoint may not be quantified, a limitation common to many ecological assessments. The approach uses the framework of Structural Equation Modeling (SEM), which combines path analysis with measurement models, to formalize available information about potential indicators and to evaluate their potential adequacy for representing an endpoint. Unlike traditional applications of SEM which require data on all variables, our approach---judgement-based SEM (JSEM)- can utilize expert judgement regarding the strengths and shapes of indicator endpoint relationships. JSEM is applied in two stages. First, a conceptual model that relates variables in a network of direct and indirect linkages is developed, and is used to identify indicators relevant to an endpoint. Second, an index of indicator strength--i.e., the strength of the relationship between the endpoint and a set of indicators--is calculated from estimates of correlation between the modeled variables, and is used to compare alternative sets of indicators. The second stage is most appropriate for large, long-term assessments. Although JSEM is not a statistical technique, basing JSEM on SEM provides a structure for validating the conceptual model and for relining the index of indicator strength as data become available. Our main objective is to contribute to a rigorous and consistent selection of indicators even when knowledge about the ability of indicators to represent an endpoint is limited to expert judgement. PMID- 11281422 TI - Long-term and seasonal changes in chemical composition of surface waters in Latvia. AB - The changes of the chemical composition in Latvian inland waters over the last twenty years were determined. based on data of the National Monitoring Programme. Relationships between the water composition were studied. The chemical composition and seasonal change pattern depend a lot on hydrological factors which differed between eastern and western Latvia. Different pattern of long-term changes is found for substances which have different sources and sinks, at first for nutrients (decrease only in the last few years) and inorganic ingredients (commonly increasing trend). An attempt was made to identify the main sources of the major water ingredients and link changes of their concentrations with changes in loading to waters. PMID- 11281424 TI - Comparative studies on the usefulness of seven ecological indices for the marine coastal monitoring close to the shore on the Swedish East coast. AB - The simultaneous behaviour of seven ecological indices (Hurlbert's, Margalef's, Menhinick's, Shannon's, species number, Jaccard's and saprobic index) was studied based on phytoplankton data close to the shore on the East coast of Sweden during the summer 1998. The sampling stations had a similar eutrophication level and were located in bays. Standard phytoplankton databases were used in calculating the indices, which were later compared using cluster analysis. Hurlbert's, Margalef's, Menhinick's, Shannon's and species number indices, as measure of community diversity, produced similar trends which often differed from those based on Jaccard's index of similarity. However, the simultaneous use of these indices was found meaningful as a possible part of the monitoring close to the shore. The application of a saprobic index lead to erroneous conclusions in the studied case. PMID- 11281423 TI - Pesticides and trace metals residue in grape and home made wine in Jordan. AB - Sixty home made wine and sixty-four grape samples were collected from five territories in Jordan, where grapes and wine are mostly producted. The collected samples were analyzed for the most used organochlorine pesticides (OCP) and organophosphorous pesticides (OPP) in Jordan, as well as for four heavy metals (Ni, Cu, Zn and Pb). The results showed that OCPs residues were detected in 73% of the wine samples but no OPPs residue were detected which is due to generally shorter half life of the later pesticide. Grapes showed higher incident of contamination than wine, however, OCPs and OPPs with both short and long half lives were detected. The OPPs were detected in only 8.3% of the analyzed grape samples. Heavy metals showed higher values in grapes than in the wine samples and it was attributed to removal of solids during wine preparation processes or through contamination of wine during storage. Most of the samples were below toxic limit. PMID- 11281425 TI - For what applications can probability and non-probability sampling be used? AB - Almost any type of sample has some utility when estimating population quantities. The focus in this paper is to indicate what type or combination of types of sampling can be used in various situations ranging from a sample designed to establish cause-effect or legal challenge to one involving a simple subjective judgment. Several of these methods have little or no utility in the scientific area but even in the best of circumstances, particularly complex ones, both probabilistic and non-probabilistic procedures have to be used because of lack of knowledge and cost. We illustrate this with a marbled murrelet example. PMID- 11281426 TI - The environmental implications of soil erosion in the United States. AB - Soil erosion has both on-farm and off-farm impacts. Reduction of soil depth can impair the land's productivity, and the transport of sediments can degrade streams, lakes, and estuaries. Since 1933, soil conservation policies have existed in the United States. Originally they focused on the on-farm benefits of keeping soil on the land and increasing net farm income. Beginning in the 1980s, however, policy goals increasingly included reductions in off-site impacts of erosion. As a consequence of conservation efforts associated with explicit U.S. government policies, total soil erosion between 1982 and 1992 was reduced by 32% and the sheet and rill erosion rate fell from an average of 4.1 tons per acre per year in 1982 to 3.1 tons per acre in 1992 while the wind erosion rate fell from an average of 3.3 tons per acre per year to 2.4 tons per acre per year over the same period. Still, soil erosion is imposing substantial social costs. These costs are estimated to be about $37.6 billion annually. To further reduce soil erosion and thereby mitigate its social costs, there are a number of policy options available to induce farmers to adopt conservation practices including education and technical assistance. financial assistance, research and development, land retirement, and regulation and taxes. PMID- 11281427 TI - Changes in patient perception and behavior following Mohs micrographic surgery. AB - BACKGROUND: Mohs micrographic surgery (MMS) is used for the removal of difficult or recurrent skin cancer. Little is known about the behavioral changes of patients who have undergone this procedure. OBJECTIVE: This study was conducted to document patients' long-term psychological and behavioral changes following MMS. METHODS: A survey was mailed to a sample of 260 persons who underwent the Mohs procedure in 1997. It included questions on sunscreen use, level of anxiety about cancer, patient confidence in MMS, and changes in high-risk habits. RESULTS: The study included 214 patients who responded to our mailed questionnaire. Trends showed an increase in some but not all preventative measures taken to avoid skin cancer recurrence post-MMS. Other healthy lifestyle changes, such as decreased cigarette smoking, were not noted. CONCLUSION: Mohs micrographic surgery has an impact on some aspects of patients' health-related behavior, especially skin cancer prevention. Other aspects which are not affected may be targets for extra patient education. PMID- 11281428 TI - Anchorage-independent growth of p53-knockout dermal fibroblasts is reversed by wild-type p53. AB - BACKGROUND: p53 is a 393-residue nuclear phosphoprotein. Mutation of p53 occurs in over half of all human cancers and thus is a crucial step in the process of cell transformation and tumorigenesis. Since tumorigenesis is a multistep process, it generally requires the mutation of certain key oncogenes and/or tumor suppressor genes. Using p53-deficient mice, we can investigate the p53-dependent mechanisms leading to tumorigenesis. OBJECTIVE: To examine the unique anchorage independent growth characteristics of dermal fibroblasts isolated from p53 deficient mice. METHODS: The growth characteristics of highly confluent cultured dermal fibroblasts from wild-type (p53+/+) and p53-deficient (p53-/-) mice were compared by DNA fragmentation assay, colony formation in soft agar, and overexpression of a wild-type p53 transgene in p53-deficient cells. RESULTS: p53 /- fibroblasts have a growth rate dramatically higher than p53+/+ cells and detach from plastic cultureware at high density. The detachment of p53-/- cells is not due to apoptosis. Furthermore, these cells have the capacity to grow in soft agar-a hallmark of cell transformation-and this anchorage-independent growth can be reversed by the introduction of a wild-type p53 transgene. CONCLUSION: Dermal fibroblasts isolated from p53-deficient mice show anchorage-independent growth. Therefore, the absence of p53 is sufficient for the initiation of cell transformation in this cell type and establishes this model system as an excellent tool to dissect the molecular steps involved in oncogenesis. PMID- 11281429 TI - Effects of a sun protection program targeting elementary school children and their parents. AB - BACKGROUND: Excessive sun exposure in childhood is considered a risk factor for later development of skin cancer, so sun awareness programs targeting children have been developed. OBJECTIVE: To assess the benefits of involving parents at home in the sun protection program received by their children at school. METHOD: The existing "Sun and the Skin" program was enhanced in two ways. Parents were educated both about their child's program and with supplemental information. Also, sunscreen was distributed to each child. RESULTS: Certain methods of sun protection, particularly the use of sunscreen, are being practiced by the majority of children, while others, such as protective clothing, have not been readily adopted. The enhanced group of students showed improvement over control and standard groups in their attitude toward tanning. There is a need for teachers to remind their students to practice protective measures. CONCLUSIONS: While a sun-awareness curriculum has been shown to be beneficial for elementary school children, the adjunct of parental and school involvement in this process can improve the results and ultimately decrease the risk of skin cancer in the children. PMID- 11281430 TI - Olanzapine may be an effective adjunctive therapy in the management of acne excoriee: a case report. AB - BACKGROUND: The self-inflicted dermatoses such as acne excoriee and neurotic excoriations are often chronic, recurring, and resistant to standard dermatologic therapies. OBJECTIVE: We present a 28-year-old woman with longstanding acne excoriee, whose acne started at age 14 years and was followed by acne excoriee at age 16 years. The patient reported that her acne and self-excoriative behavior were exacerbated by psychological stress. The previously treatment-resistant acne excoriee responded favorably to treatment with the atypical antipsychotic agent olanzapine. METHODS: The patient was started on olanzapine 2.5 mg at bedtime. RESULTS: After 4 weeks of therapy with olanzapine she reported a significant decline in her self-excoriative behavior which was associated with an improvement in her acne excoriee. The patient used the olanzapine 2.5 mg for 6 months, during which time she also entered psychotherapy in order to deal with some psychosocial stressors that were exacerbating her self-excoriative behavior. The patient has not experienced a recurrence in her self-excoriative behavior or acne excoriee for 4 months after discontinuing the olanzapine. CONCLUSION: Olanzapine may prove to be a useful adjunctive therapy in some self-induced dermatoses including acne excoriee. PMID- 11281431 TI - A case of cutaneous Mycobacterium chelonae abscessus infection in a renal transplant patient. AB - BACKGROUND: Mycobacterium chelonae is an atypical "fast-growing Mycobacteria" that is a rare cause of human infection. There have been several reports of cutaneous infection among immunosuppressed patients, as well as in immunocompetent individuals following trauma. Most cases to date seem to have occurred among renal transplant recipients, raising the possibility that there is something inherent to the renal transplant patient that increases their susceptibility more than other immunocompromised patients. OBJECTIVE: The differential diagnosis of subcutaneous nodules distributed in a sporotrichoid pattern is extensive, particularly in an immunocompromised host. Although several cases of cutaneous M. chelonae abscessus infection have been reported among both immunosuppressed and immunocompetent patients, the clinical presentation has varied, and few cases have reported the appearance of lesions in a sporotrichoid pattern. We present a case of a renal transplant patient with a reported history of trauma to the lower extremities, who presents with subcutaneous nodules distributed in a sporotrichoid pattern. The patient is found to have M. chelonae abscessus infection, fails several treatment regimens, and presents with a recurrence. The literature of M. chelonae infection is reviewed, and the various treatment options are discussed. METHODS: An initial skin biopsy was stained with Hematoxylin and Eosin and revealed deep dermal abscesses with acid-fast bacilli in clusters. The culture became positive for Mycobacterium chelonae abscesses in four days and was found to be sensitive to multiple antibiotics. The patient underwent surgical excision of 14 nodules, which revealed findings consistent with the skin biopsy, and was subsequently treated with the appropriate antibiotics. RESULTS: Despite treatment with a full course of an organism sensitive antibiotic regimen, the patient returned with persistent and recurrent nodules six weeks later. The patient was then treated as an inpatient with a seven-week course of intravenous antibiotics and was discharged home on a combined intravenous and oral regimen. CONCLUSION: Although M. chelonae abscessus is an extremely rare cause of infection among humans, there seems to be a predominance of cases reported among renal transplant patients. The explanation for this is not entirely clear; however, the organism must be considered as a cause of infection in any renal transplant recipient who presents with subcutaneous nodules. Eradication of the organism presents a tremendous challenge to the clinician, and, as presented here, even with appropriate antibiotics, there is a high rate of recurrence. PMID- 11281432 TI - Eosinophilic fasciitis occurring four weeks after the onset of dialysis in a renal failure patient. AB - BACKGROUND: Eosinophilic fasciitis is a rare, scleroderma-like disease that usually affects the extremities of young to middle-aged males. The disease may cause flexion contractures and limit joint mobility and is associated with peripheral eosinophilia. The fascia, by definition, is infiltrated with mononuclear cells and typically with eosinophils. Eosinophilic fasciitis may be separated from another sclerodermatous disorder, linear scleroderma, by its response to systemic corticosteroids. The etiology is unclear but eosinophilic fasciitis has numerous disease associations. However, it has not previously been associated with renal failure and hemodialysis. OBJECTIVE: This article reports a case of eosinophilic fasciitis occurring four weeks following the onset of hemodialysis. METHODS: The clinical and histologic features confirmed the diagnosis of eosinophilic fasciitis. He was treated with systemic corticosteroids with good response. CONCLUSION: This is the first reported patient who developed eosinophilic fasciitis in close temporal relationship with the start of hemodialysis. While eosinophilic fasciitis may be coincidental with a common disorder, namely, renal failure, it is interesting to note that hemodialysis patients often have immune-regulation abnormalities and peripheral eosinophilia. PMID- 11281433 TI - The efficacy and safety of a combination benzoyl peroxide/clindamycin topical gel compared with benzoyl peroxide alone and a benzoyl peroxide/erythromycin combination product. AB - BACKGROUND: Topical clindamycin and benzoyl peroxide have each demonstrated clinical efficacy in the treatment of acne vulgaris. When used in tandem, they promise greater efficacy than either individual agent through their antibacterial and anti-inflammatory effects. OBJECTIVE: To determine the efficacy and safety of combination benzoyl peroxide/ clindamycin compared with benzoyl peroxide or benzoyl peroxide/erythromycin in the treatment of acne. METHODS: In this randomized, 10-week, multicenter, single-blind trial, 492 patients with moderate to moderately severe acne were treated twice daily with 5% benzoyl peroxide/1% clindamycin, 5% benzoyl peroxide, or 5% benzoyl peroxide/3% erythromycin and assessed every 2 weeks. RESULTS: Compared with benzoyl peroxide, benzoyl peroxide/clindamycin demonstrated significantly greater reductions in inflammatory lesions (p = 0.04) and significantly greater overall improvement as assessed by physicians (p < or = 0.04) and patients (p < 0.001). Benzoyl peroxide/clindamycin demonstrated a nonsignificant trend for greater efficacy compared to benzoyl peroxide/erythromycin. Dry skin was the most frequent (< or = 7.3%) adverse event with all three therapies. CONCLUSION: Benzoyl peroxide/clindamycin demonstrated improved efficacy and similar tolerability; to benzoyl peroxide used alone and was similar to benzoyl peroxide/ erythromycin, making this combination product an effective alternative antimicrobial therapy for acne. PMID- 11281434 TI - Human papillomavirus infections: epidemiology, pathogenesis, and therapy. AB - BACKGROUND: Human papillomaviruses (HPV) are common human pathogens and are classified into more than 80 different types. These viruses produce benign warts in many cases and aggressive squamous cell carcinomas in other cases. OBJECTIVE: The goal of this review is to update the reader on the epidemiology, pathogenesis, and therapy of HPV infections. Nonanogenital warts are transmitted by skin-to-skin contact while anogenital warts are usually transmitted sexually. Both types of warts produce much morbidity but rarely undergo malignant transformation. They are commonly treated with surgical or cytodestructive therapy, but immunomodulatory agents, such as imiquimod, have been proven to be very effective in anogenital warts and are being evaluated in nonanogenital warts. Other types of HPV have marked oncogenic potential such that over 99% of all cervical cancers and over 50% of other anogenital cancers are due to infection with oncogenic HPV. Many cofactors, such as cigarette smoking, genetics, and helper viruses, have potential roles in HPV oncogenesis, but their relative contributions are poorly understood. Other control measures for warts and HPV-associated cancers are under study, but the greatest future potential may be from the development of prophylactic and therapeutic vaccines. CONCLUSIONS: Infection with HPV is very prevalent as are the clinical manifestations of this family of pathogens. Improved therapies for warts (e.g., imiquimod) have recently become available. Vaccines for HPV offer hope for future interventions for warts as well as for prevention of anogenital malignancies. PMID- 11281435 TI - Photodynamic therapy with 5-aminolevulinic acid induces apoptosis and caspase activation in malignant T cells. AB - BACKGROUND: Preliminary studies have suggested that photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA) can improve psoriasis and mycosis fungoides, two diseases where normal or malignant T cells play a central role. OBJECTIVES: To determine if ALA-PDT induces apoptosis and caspase activation in Jurkat cells, a malignant T-cell line. METHODS: Jurkat cells were incubated with ALA in the presence of [14C]-thymidine followed by red light exposure. DNA fragmentation was measured 24 hours later with a DNA elution assay. The influence on DNA fragmentation of ALA concentration, time between ALA addition and light exposure, as well as light fluence were studied. The occurrence of oligonucleosome-sized DNA fragmentation was also studied with DNA electrophoresis. Caspase-3-like activity was monitored by measuring Ac-DEVD-AMC hydrolysis. RESULTS: DNA fragmentation as high as 88% was observed 24 hours after ALA-PDT. The percentage of DNA fragmentation increased with increasing doses of ALA, red light fluence, as well as longer incubation time with ALA. DNA fragmentation was observed as early as 3 hours after ALA-PDT. The presence of apoptosis after ALA-PDT was confirmed by DNA electrophoresis. An increase in caspase-3-like activities was detected following ALA-PDT. CONCLUSION: ALA-PDT induces apoptosis and caspase-3 like activation in Jurkat cells. PMID- 11281436 TI - Helicobacter pylori infection and its treatment in Singapore. PMID- 11281437 TI - Prevalence of Helicobacter pylori in peptic ulcer disease in a Singapore hospital. AB - INTRODUCTION: High Helicobacter pylori (H. pylori) prevalence is well documented among peptic ulcer patients. However, there have been recent reports of declining H. pylori infection rates in developed countries. Based on previous local data in a different hospital, H. pylori prevalence was 66% in gastric ulcer, 86% in duodenal ulcer and 75% in combined gastric and duodenal ulcers. Our present study aims to review the recent H. pylori prevalence in peptic ulcer patients in a Singapore hospital. H. pylori infection rate in relation to sex, age, non steroidal anti-inflammatory drug (NSAID) use and race were also examined. METHODS: Over a 6-month period, patients diagnosed with peptic ulcer by oesophagogastroduodenoscopy were selected. Relevant information was obtained from case notes retrieved from the Medical Records Office. H. pylori status was assessed by rapid urease test (CLOtest) and/or histology. Exclusion criteria were(1) history of peptic ulcer(2), active bleeding(3), cancer or(4) recent use of antibiotics or proton pump inhibitors. RESULTS: 107 peptic ulcer patients were selected; 53 gastric ulcer, 47 duodenal ulcer and 7 with combined gastric and duodenal ulcers. Overall H. pylori prevalence was 67.9%(36/53), 85.1%(40/47) and 85.7%(6/7) respectively. Except for the gender variable in gastric ulcer group, age, race and NSAID use was not found to influence H. pylori prevalence. CONCLUSIONS: The prevalence of H. pylori among our peptic ulcer patients is slightly lower compared to overseas studies but the local trend,when compared indirectly to another previous local study did not appear to have changed much. Reasons for the lower H. pylori infection rate in comparison to overseas studies are discussed. The lower H. pylori prevalence among female gastric ulcer patients may be due to the higher prevalence of NSAID use. PMID- 11281438 TI - A randomised trial of amoxycillin versus clarithromycin in combination with omeprazole for eradication of Helicobacter pylori infection in Singapore. AB - Dual therapy has been reported to produce H.pylori eradication rate of 75-80%. This study is designed to determine the efficacy of omeprazole 20 mg bd in combination with amoxycillin 500 mg tid (Group A), amoxycillin 750 mg tds (Group B) and clarithromycin 500 mg tid (Group C) in Singapore. One hundred and forty eight patients with H. pylori positive duodenal ulcers between ages of 22 and 69 were enrolled from two centres. There were 48 patients in Group A, 50 patients in Group B and 50 patients in Group C. The medication was given for 14 days. The patients were re-evaluated with an upper GI endoscope 4 weeks after cessation of treatment Successful eradication was defined as H.pylori negative on histology and culture. Based on intention to treat analysis, the eradication rate was 47.8% in Group A, 68% in Group B and 66% in Group C. The difference between GroupA and B were statistically significant (p = 0.04). Based on all patient treated analysis, the eradication rate was 57.5% in Group A, 70.7% in Group B and 75% in Group C. The difference in eradication rates was not statistically significant. Adverse events were reported in 21% of all patients with no difference in the adverse event rate between all groups. The eradication rate achieved with dual therapy in this study was similar to that attained in Western population. Higher dose amoxycillin regime gives a significantly higher eradication than a lower dose amoxycillin. PMID- 11281439 TI - Lumbar puncture refusal in febrile convulsion. AB - A descriptive study was carried out on patients admitted for febrile convulsion over a two-year period to determine rate of lumbar puncture (LP) refusal, factors associated with LP refusal and outcome of such patients. From 77 patients indicated and requested for LP, 19 (25%) patients refused the procedure. Refusal of LP was significantly more common among the Malay ethnic group (p = 0.01) but not significantly associated with age,gender or whether the patient was admitted for a first or recurrent febrile convulsion. Half of the patients who refused LP had to be started empirically on antibiotics for meningitis. Patients who refused LP were also 8.5 times more likely to discharge themselves "at own risk" (AOR), compared to other patients with febrile convulsion (p = 0.004). In conclusion, LP refusal is a common problem in the local setting and is a hindrance to the proper management of patients with fever and seizure. Appropriate measures must be carried out to educate the public, particularly those from the Malay ethnic group on the safety and usefulness of the procedure. Reasons for patients discharging AOR following LP refusal also need to be addressed and problems rectified. PMID- 11281441 TI - Rapid prenatal diagnosis of chromosome abnormalities. AB - The aim of the present work was to examine the efficacy of using FISH for the rapid prenatal diagnosis of common chromosome aneuploidies. A total of 100 analyses over a six month period were included in the study. Diagnosis was possible in all cases. A mosaic for trisomy 21 proved, by comparison with an extensive analysis of long term cultures, to be an apparent false positive. Otherwise the technique was reliable, accurate and relatively straightforward to perform. Results could be available within 24 hrs. In most cases an additional long term full analysis was also done, so as to exclude rarer aneuploidies and structural rearrangements. This methodology is seen as a useful addition to the prenatal diagnostic repertoire. PMID- 11281440 TI - Pyogenic liver abscess--a tropical centre's experience in management with review of current literature. AB - AIM OF STUDY: To perform a retrospective study, with the help of literature review, of the management of patients with pyogenic liver abscess in a general hospital. METHOD: A retrospective study of 73 consecutive patients treated atTanTock Seng Hospital between January 1994 and December 1997 was conducted to determine the demographic, clinical, laboratory, radiological and microbiological characteristics of these patients, as well as the management strategies employed. RESULTS: Liver abscess was more common in males, occurring more frequently in the right hepatic lobe. Most patients presented with non-specific clinical and biochemical features. A raised alkaline phosphatase level was the most common biochemical abnormality found in about two-thirds of patients. Ultrasonography was not as sensitive as computed tomographic scans in detecting abscesses. Klebsiella pneumoniae was the most common etiological agent detected in cultures of blood and abscess aspirates. All patients were treated with intravenous antibiotics. Twenty-two (30%) needed percutaneous catheter drainage and five (7%) required surgical management. There was no hospital mortality in our series. Prolonged hospitalisation was associated with advanced age, degree of loculation within the abscess, concomitant diabetes mellitus and Klebsiella septicaemia. CONCLUSION: Pyogenic liver abscesses require a high index of suspicion for early diagnosis. When appropriate therapy in the form of antibiotics in combination with percutaenous drainage or surgery is administered, mortality is very low. However, significant morbidity is still a problem, particularly in the elderly, diabetic patient. PMID- 11281442 TI - Obsessive compulsive disorder following left middle cerebral artery infarct. AB - A case of a 37-year-old male who developed compulsive washing after a massive infarct involving the left middle cerebral artery, is described. Although an obsessive compulsive disorder can occur following neurological disorders,there have been no previous reports of it occurring following a middle cerebral artery infarct. PMID- 11281443 TI - Immunoglobulin D multiple myeloma in our hospital--a rare occurrence. AB - Immunoglobulin (Ig) D multiple myeloma is a rare presentation, usually with an aggressive course and a poorer prognosis. It accounts for about 1-2% of newly diagnosed mulitple myeloma patients. Due to its rarity, reports on Ig D multiple myeloma are limited in the literature. We therefore present 4 cases of Ig D multiple myeloma in our hospital over a period of 8 years between 1990 to 1998. The average age of presentation of our patients was 44 years old with a female preponderance. Common presenting symptoms were appetite and weight loss and bone pain. Two patients presented with neurological symptoms and 2 with renal impairment. Three patients had an associated lambda paraproteinaemia and the fourth had a kappa paraproteinaemia. A common finding in Ig D myeloma is a small or no spike seen on serum electrophoresis together with heavy Bence Jones proteinuria. A review of the literature on Ig D myeloma is also presented. PMID- 11281444 TI - Golf buggy related head injuries. AB - Our department has recently managed three cases of serious head injuries resulting from falls from golf buggies. One of them sustained moderate head injury with a small cerebral contusion and skull fracture. Two of them sustained severe head injury with extensive cerebral contusions, extradural haematoma requiring craniotomy. Of the three patients, two made good recoveries whereas the third remained vegetative. We feel that instruction on the safe use of golf buggies is inadequate and should be intensified. PMID- 11281445 TI - Inhaled foreign bodies in children--anaesthetic considerations. AB - Many anaesthetic problems and hazards exist in children with inhaled foreign bodies. Careful, stepwise and detailed consideration of clinical history, examination and investigations cannot be over-emphasised. Special consideration has to be given to the fragile paediatric cardiopulmonary status especially in the presence of airway foreign body. The small paediatric airway is frequently shared for anaesthesia and endoscopy. Complete cooperation and good communication between the endoscopist and the anaesthetist is paramount in achieving an optimal outcome. PMID- 11281446 TI - Clinics in diagnostic imaging (54). AB - A 29-year-old man sustained a pelvic fracture and haemorrhagic shock after a road traffic accident. He underwent an exploratory laparotomy open cystostomy, and iliac artery embolisation. Subsequent antegrade cystography and computed tomography showed a complete traumatic posterior urethral rupture with a "high riding" bladder. Delayed repair of the posterior urethra was performed 6 months later with good functional outcome. The risk of urethral injury in pelvic fractures, the mechanism of injury, and the role of imaging in the diagnosis of possible urethral injury in pelvic fractures are discussed. PMID- 11281447 TI - Lack of association between alpha2-macroglobulin polymorphisms and Alzheimer's disease. AB - This study was undertaken to investigate the role of two alpha2-macroglobulin (A2M) polymorphisms, an intronic 5-bp deletion and Ile1000Val, in the development of Alzheimer's disease (AD) and to evaluate the interaction between the apolipoprotein E (APOE) and A2M polymorphisms. The A2M polymorphisms were screened by using polymerase-chain-reaction-based assays in 555 white late-onset AD cases and 446 controls. The gentoype distributions of the 5-bp deletion and Ile1000Val polymorphisms were comparable between cases and controls (P = 0.158 and P = 0.148, respectively). Likewise, there was no significant difference in allele frequencies of each polymorphism among cases and controls (P = 0.361 and P = 0.062, respectively). The stratification of data by APOE*4 status also did not yield any significant association. In conclusion, we observed no association between either the intronic deletion polymorphism or the Ile1000Val polymorphism of A2M and AD in our case-control cohort. PMID- 11281449 TI - Clonal maintenance of imprinted expression of SNRPN and IPW in normal lymphocytes: correlation with allele-specific methylation of SNRPN intron 1 but not intron 7. AB - DNA methylation is a heritable and reversible modification to CpG sites in the mammalian genome. Parental allele-specific methylation is hypothesized to be important in the establishment and maintenance of imprinted gene expression; however, dynamic changes in allele-specific patterns have been observed. The upstream regulatory region of the small nuclear riboprotein N gene (SNRPN) is an important imprinting control region (ICR) for establishing and maintaining the methylation imprint in the locus on 15q11-13 associated with Prader-Willi and Angelman syndromes (PWS). To compare directly the role of allele-specific methylation patterns and the maintenance of imprinted expression in the PWS region, clonal populations of normal T lymphocytes were cultured for 22-25 generations. A novel long-range semi-nested polymerase chain reaction (PCR) strategy was utilized in order to span two different methylation sites, and a polymorphism within SNRPN was used so that allele-specific methylation of both sites could be determined. Reverse transcription/PCR followed by polymorphism analysis was also performed in order to determine parental allele-specific transcription. Exclusive paternal expression at both SNRPN and IPW was maintained in all T cell clones and correlated with maternal methylation of the intron 1 NotI site. In contrast, biallelic methylation was observed in all clones at the previously described paternally methylated HpaII site in intron 7. These results demonstrate that the maintenance of paternal expression of SNRPN and IPW correlates with a strict clonal maintenance of allele-specific methylation at the CpG-dense 5' end of SNRPN. Differential maintenance of methylation sites within imprinted genes may depend on the density and chromatin organization of surrounding CpG sites. PMID- 11281448 TI - Hybrids monosomal for human chromosome 5 reveal the presence of a spinal muscular atrophy (SMA) carrier with two SMN1 copies on one chromosome. AB - We have analyzed the survival motor neuron gene (SMN1) dosage in 100 parents of children with homozygous SMN1 deletions. Of these parents, 96 (96%) demonstrated the expected one-copy SMN1 carrier genotype. However, four parents (4%) were observed to have a normal two-copy SMN1 dosage. The presence of two intact SMN1 genes in the parent of an affected child indicates either the occurrence of a de novo mutation event or a situation in which one chromosome has two copies of SMN1, whereas the other is null. We have separated individual chromosomes from two of these parents with two-copy SMN1 dosage by somatic cell hybridization and have employed a modified quantitative dosage assay to provide direct evidence that one parent is a two-copy/ zero-copy SMN1 carrier, whereas the other parent had an affected child as the result of a de novo mutation. These findings are important for assessing the recurrence risk of parents of children with spinal muscular atrophy and for providing accurate family counseling. PMID- 11281450 TI - Quantitation of fetal DNA in maternal serum in normal and aneuploid prenancies. AB - We investigated whether the amount of circulating cell-free fetal DNA in maternal serum is influenced by fetal karyotype, using real-time quantitative polymerase chain reaction assay. Serum samples were obtained from pregnant women at gestational ages ranging from 15 to 17 weeks, prior to their undergoing amniocentesis. In total, we examined 70 samples consisting of 55 cases of pregnancy with 46,XY, 5 cases with 47,XY,+21, 3 cases with 47,XY,+18, a single case with 46,XY,dup(1) and 2 cases with twins of 46,XY, and 4 cases with 46,XX which were used as negative controls. We measured the concentration of the SRY sequence as a molecular marker for fetal DNA. The SRY sequence was detectable and measurable when the fetuses were male except for one case with 47,XY,+18. This case showed fetal growth retardation and bradycardia. No amplification signals of the SRY sequence were detected when the fetuses were female. The mean concentration of fetal DNA in maternal serum was 31.5 copies/ml in the pregnancy with 46,XY, 23.5 copies/ml in the pregnancies with 47,XY,+21 and 21.5 copies/ml in the pregnancies with 46,XY,+18. There were no significant differences in the concentration of fetal DNA between pregnancies with fetuses of normal karyotype and those with fetuses of abnormal karyotype. PMID- 11281451 TI - Polymorphism in promoter region of Fcalpha receptor gene in patients with IgA nephropathy. AB - We have recently identified two novel polymorphisms (-114T/C and +56T/C relative to the major transcription start site) in the functional promoter region of the Fcalpha receptor (FcalphaR) gene. Since altered FcalphaR expression may be associated with IgA nephropathy, we examined these promoter polymorphisms in this disease. Patients with IgA nephropathy (n = 90), patients with other primary glomerulonephritis (n = 50), and healthy adults (n = 83) were studied. Genotyping was performed by the polymerase chain reaction (PCR) followed by direct sequence analysis and was subsequently confirmed by PCR with mismatched primers followed by restriction fragment length polymorphism analysis. The study demonstrated that the frequency of the +56C allele in patients with IgA nephropathy (0.511) was significantly (P < 0.01) higher than that in patients with other primary glomerulonephritis (0.350) and healthy adults (0.337). In addition, a significant increase in the frequency of the +56CC genotype was observed in patients with IgA nephropathy (27.8% vs 10.0% in other GN and vs 9.6% in healthy adults). In contrast, no significant difference in the frequencies of the +56CC genotype and +56C allele was observed between other primary glomerulonephritis patients and healthy adults. The frequency of the -114CC genotype in patients with IgA nephropathy was significantly increased compared with those in both control groups (15.6% vs 4.0% in other GN and vs 2.4% in healthy adults). Polymorphisms of the FcalphaR promoter region therefore appear to be associated with susceptibility to IgA nephropathy, suggesting the importance of the FcalphaR gene and its expression in this disease. PMID- 11281452 TI - Numerical chromosome abnormalities in the spermatozoa of the fathers of children with trisomy 21 of paternal origin: generalised tendency to meiotic non disjunction. AB - The purpose of this study was the evaluation of aneuploidy frequencies in the spermatozoa of two fathers (DP-4 and DP-5) who had children with Down syndrome (DS) of paternal origin and in whom a previous sperm analysis by fluoresence in situ hybridisation (FISH) had suggested a generalised tendency to meiotic non disjunction. Sperm samples were simultaneously hybridised with FISH probes for chromosomes 4, 13 and 22. Disomy frequencies for each of the chromosomes and diploidy frequencies were compared with data obtained from nine control donors. Both DS fathers had a statistically significant increase in the frequency of disomy for chromosomes 13 and 22. DP-5 also had an increased frequency of diploid spermatozoa. Our data suggest that the two DS fathers have a generalised susceptibility to meiotic non-disjunction and that acrocentric chromosomes seem to be more sensitive to such disturbance in the meiotic process. PMID- 11281453 TI - Isolation and characterization of the UBASH3A gene on 21q22.3 encoding a potential nuclear protein with a novel combination of domains. AB - In order to identify candidate genes for Down syndrome phenotypes or monogenic disorders that map to human chromosome 21q22.3, we have used genomic sequence and expressed sequence tags mapping to an autosomal recessive deafness (DFNB10) critical region to isolate a novel 2.5-kb cDNA that maps between TFF1 and D21S49. A semi-quantitative reverse transcription/polymerase chain reaction method revealed that UBASH3A gene expression is limited to only a few tissues, with its highest expression in spleen, peripheral blood leukocytes, and bone marrow. The putative 661-amino-acid protein shows considerable homology to a hypothetical protein from Drosophila melanogaster but only domain homologies to other organisms. Both the human and D. melanogaster proteins contain protein-protein interaction domains, viz., SH3 and ubiquitin-associated (UBA) domains, in addition to a novel domain also containing a nuclear localization signal. This is the first protein described containing both UBA and SH3 domains. The gene, thus called UBASH3A, spans 40 kb and is divided into 15 exons. Mutation analysis excluded UBASH3A as being responsible for DFNB10. PMID- 11281454 TI - Characterization of new mutations in the coding sequence and 5'-untranslated region of the human prostacylcin synthase gene (CYP8A1). AB - Inheritable interindividual differences in prostacyclin production may be implicated in the pathogenesis of several human vascular diseases. Using a polymerase chain reaction/single-strand conformation polymorphism strategy, we screened for mutations in the gene encoding cytochrome P450 prostacyclin synthase (CYP8A1). DNA samples from healthy French volunteers (n = 130) of Caucasian origin were examined. Five mutations, comprising two previously reported silent mutations and three novel rare missense mutations (P38L, S118R, and R379S), were identified in the coding sequence of the gene. In the 5'-proximal region, we also found a variable number of tandem repeats (VNTR) polymorphism that consisted of four different alleles with 4-6 tandem repeats of a 9-bp unit containing a putative Spl transcriptional factor binding site. One of these (R6), a frequent allele (23.6% of alleles tested) harboring six repeats, is novel, whereas the other three are known. In vitro analysis of the effect of each VNTR allele on promoter activity of a reporter gene was performed by a transient transfection assay. Data confirmed the modulator effect of the VNTR polymorphism on reporter gene transcription. Furthermore, the data demonstrate that allele R6 has the most potent inducing effects in the A549 cell line and, after IL-6 stimulation, in human pulmonary artery endothelial cells. Overall, the data demonstrate that CYP8A1 is polymorphic in Caucasians, and that a polymorphism affecting the 5' proximal region may result in interindividual differences in CYP8A1 transcriptional regulation in vivo. Additional factors, such as the presence of inflammatory mediators, may be required to modulate transcription of the CYP8A1 gene. PMID- 11281455 TI - TSC1 and TSC2 deletions differ in size, preference for recombinatorial sequences, and location within the gene. AB - Large TSC gene rearrangements are not rare findings in tuberous sclerosis. Interestingly, all deletions, duplications and inversions so far described involve TSC2, none being associated with TSC1. In order to shed light on the structural basis of the preferential DNA rearrangements in TSC2 over TSC1 and to assess, in an unselected patient population, the prevalence of large re arrangements in both TSC loci, we screened 202 tuberous sclerosis patients consecutively referred at our center. Southern blot analysis on EcoRI+HindIII double-digested DNA identified 19 partial or full-length gene deletions: three involved TSC1 and sixteen TSC2. The breakpoint sequence of seven internal deletions, three in TSC1 and four in TSC2, allowed us to speculate on the mechanism favoring TSC2 unequal recombinations and to identify a deletion hot spot that lies in TSC1 and that may be relevant in the routine genetic testing of tuberous sclerosis. Briefly, three major features appear to distinguish TSC1 from TSC2 deletions: (1) deletion size: all TSC1 deletions are within the transcriptional unit, whereas 12 of the 16 TSC2 deletions have at least one external breakpoint; (2) location within the gene: all TSC1 deletions are confined to the 3'end of the gene (all three 5' breakpoints being located in intron 20) thus resulting in the same frameshift mutation following amino acid K875, whereas the TSC2 internal breakpoints appear to be scattered along the gene; (3) preference for recombinatorial sequences: six out of eight internal TSC2 breakpoints map within Alu repeats, whereas none of the three TSC1 deletions appear to be Alu-mediated. Indeed, in the latter gene, unique structural features (a purine-rich tract flanked by pyrimidine-rich segments) surrounding one of the two identified breakpoint cluster regions might play a role in promoting inappropriate recombinations. PMID- 11281456 TI - Back mutation can produce phenotype reversion in Bloom syndrome somatic cells. AB - A unique and constant feature of Bloom syndrome (BS) cells is an excessive rate of sister-chromatid exchange (SCE). However, in approximately 20% of persons with typical BS, mosaicism is observed in which a proportion of lymphocytes (usually a small one) exhibits a low-SCE rate. Persons with such mosaicism predominantly are genetic compounds for mutation at BLM, and the low-SCE lymphocytes are the progeny of a precursor cell in which intragenic recombination between the two sites of BLM mutation had generated a normal allele. Very exceptionally, however, persons with BS who exhibit mosaicism are homozygous for the causative mutation. In two such exceptional homozygous persons studied here, back mutation has been demonstrated: one person constitutionally was homozygous for the mutation 1544insA and the other for the mutation 2702G-->A. Revertant (low-SCE) lymphoblastoid cells in each person were heterozygous for their mutations, i.e., a normal allele was now present. The normal alleles must have arisen by back mutation in a precursor cell, in one person by the deletion of an A base and, in the other, the nucleotide substitution of a G base for an A base. Thus, back mutation now becomes, together with intragenic recombination, an important genetic mechanism to consider when explaining examples of a reversion of somatic cells to "normal" in persons with a genetically determined abnormal phenotype. PMID- 11281457 TI - Neonatal presentation of adult-onset type II citrullinemia. AB - Adult-onset type II citrullinemia (CTLN2) is characterized by a liver-specific argininosuccinate synthetase deficiency caused by a deficiency of the citrin protein encoded by the SLC25A13 gene. Until now, however, no SLC25A13 mutations have been reported in children with liver diseases. We described three infants who presented as neonates with intrahepatic cholestasis associated with hypermethioninemia or hypergalactosemia detected by neonatal mass screening. DNA analyses of SLC25A13 revealed that one patient was a compound heterozygote for the 851de14 and IVS11+IG-->A mutations and two patients (siblings) were homozygotes for the IVS11+lG-->A mutation. These results suggested that there may be a variety of liver diseases related to CTLN2 in children. PMID- 11281458 TI - A summary of 20 CACNA1F mutations identified in 36 families with incomplete X linked congenital stationary night blindness, and characterization of splice variants. AB - Incomplete X-linked congenital stationary night blindness (CSNB) is a recessive, non-progressive eye disorder characterized by abnormal electroretinogram and psychophysical testing and can include impaired night vision, decreased visual acuity, myopia, nystagmus, and strabismus. Including the 20 families previously reported (Bech-Hansen et al. 1998b), we have now analyzed patients from a total of 36 families with incomplete CSNB and identified 20 different mutations in the calcium channel gene CACNA1F. Three of the mutations account for incomplete CSNB in two or more families, and a founder effect is clearly demonstrable for one of these mutations. Of the 20 mutations identified, 14 (70%) are predicted to cause premature protein truncation and six (30%) to cause amino acid substitutions or deletions at conserved positions in the alpha1F protein. In characterizing transcripts of CACNA1F we have identified several splice variants and defined a prototypical sequence based on the location of mutations in splice variants and comparison with the mouse orthologue, Cacnalf. PMID- 11281459 TI - Marked differences in unilateral isolated retinoblastomas from young and older children studied by comparative genomic hybridization. AB - Although it is established that the loss of function of both alleles of the RB1 gene is a prerequisite for the development of retinoblastoma, little is known about the genetic events that are required for tumor progression. We used comparative genomic hybridization (CGH) to search for DNA copy number changes in isolated unilateral retinoblastomas. From a series of 66 patients with retinoblastomas with somatic mutations in both RB1 alleles, tumor samples from 13 children with the youngest (2.0-9.8 months) and 13 with the oldest (36.2-84.1 months) age at operation were studied. Loss at 13q14, the location of RB1, was demonstrated in two tumors only. Recurring chromosome imbalances included gains at 6p (11/26), 1q (10/26), 2p (4/26), and 17q (4/26), gains of the entire chromosome 19 (3/26), and losses at 16q (9/26). A commonly gained region at 1q32 was identified. Increased dosage of GAC1, a candidate oncogene located in 1q32, was found in two of four tumors by Southern blot analysis. Comparison of the CGH findings revealed that retinoblastomas from children with an older age at operation showed significantly more frequent (13/13 cases vs 4/13 cases; P = 0.0005) and more complex genetic abnormalities (median, 5 changes/abnormal tumor vs median, 1.5 changes/abnormal tumor; P = 0.003) than retinoblastomas from children with a young age at operation. Gains at 1q, 2p, 17q, of the entire chromosome 19 and losses of 16q were restricted to the older age group. Our results suggest that the progression of retinoblastomas from older patients follows mutational pathways different from those of younger patients. PMID- 11281460 TI - Qualitative research in obstetrics and gynaecology. PMID- 11281461 TI - A comparison of oral and vaginal misoprostol tablets in induction of labour at term. AB - OBJECTIVE: To compare the efficacy of equivalent doses of orally administered with vaginally administered misoprostol in induction of labour at term. DESIGN: A non-blinded randomised controlled trial. SETTING: Induction and labour wards of a UK teaching hospital. PARTICIPANTS: Two hundred and forty-five pregnant women at term, with medical or obstetric indications for labour induction and unfavourable cervices. METHODS: The women were randomly assigned to receive 50 microgm of misoprostol orally or vaginally four hourly to a maximum of five doses. MAIN OUTCOME MEASURES: Interval from induction to vaginal delivery, mode of delivery, oxytocic and analgesic requirements in labour, neonatal outcome, patient satisfaction and acceptability. RESULTS: The mean induction to vaginal delivery interval was significantly shorter in the vaginal group compared with the oral group (17.8h vs 27.9h: mean difference 10.1 hrs, 95% CI 5.8-14.4). More women were delivered within 24 hours (80% vs 46.3%; RR 1.7, 95% CI 1.3-2.1) and fewer women needed oxytocin augmentation (39% vs 58.2%; RR 0.67, 95% CI 0.5-0.9) in the vaginal group. There was no difference in the mode of delivery, analgesic requirements or neonatal outcomes in the two groups. There was a higher incidence of uterine hyperstimulation in the vaginal group (4.9% vs 0.8%, RR 6, 95% CI 0.07 48.7) and more caesarean sections were performed for fetal distress in this group (13% Vs 2.4%, RR 5.3, 95% CI 1.6-17.7), although the overall operative delivery rate was similar in the two groups. CONCLUSION: Misoprostol effectively induces labour, with the vaginal route of administration having a faster action than the oral route in equivalent doses. However, the more frequent occurrence of hyperstimulation and the higher intervention rate for fetal distress in the vaginal group could mean that the preferred route might be oral. More trials are needed to find the right oral dosage that combines efficacy with safety. PMID- 11281462 TI - Blood selenium and glutathione peroxidases in miscarriage. AB - OBJECTIVE: To investigate the selenium glutathione and glutathione peroxidase (glutathione-Px) levels in blood in women who experience miscarriage. DESIGN: An observational study. PARTICIPANTS: Forty women with miscarriage occurring in the first and second trimesters. METHODS: Concentrations were measured in whole blood and plasma. Glutathione was measured in red cells and glutathione-Px activity was measured in the red cells and plasma. The results were compared with 36 women in the same period of viable pregnancy and 28 age-matched, healthy, non-pregnant controls. RESULTS: The selenium concentrations in whole blood and plasma of women following abortion were the same as in viable pregnancy, but were significantly lower compared with controls. The glutathione levels were significantly higher in women with miscarriage compared with viable pregnancies and with non-pregnant women. Red cell and plasma glutathione-Px activities of women who had had a miscarriage were significantly lower than in both normal pregnancies and the control group. CONCLUSION: The decreased activities of the antioxidant enzymes, red cell and plasma glutathione-Px, may play an important role in the aetrology of spontaneous abortion. PMID- 11281463 TI - The development of high venous velocity at the fetal umbilical ring during gestational weeks 11-19. AB - OBJECTIVE: To determine the occurrence of high venous velocities at the umbilical ring in the normal early second trimester, based on the assumption that a narrow umbilical ring may cause obstruction and increased venous blood velocity at the abdominal wall. DESIGN: Cross-sectional study. SETTING: Hospital antenatal clinic. POPULATION: One hundred and one low risk singleton pregnancies specifically recruited for the study. METHODS: Ultrasound was used at 11-19 weeks to determine the diameter and velocity in the umbilical vein at the fetal end of the cord and at the inlet through the abdominal wall. Outcome measures 10th, 50th and 90th centiles were estimated for the time-averaged maximum velocity in the cord and at the abdominal inlet. The increase of velocity as the blood entered the abdominal wall was calculated in percent of the velocity in the cord. RESULTS: During weeks 11-12 there was hardly any difference between blood velocity in the umbilical vein at the umbilical ring and that in the cord. From week 13 onwards it was increasingly common to find blood acceleration at the umbilical ring of 50-500%. Velocity increment >50% was found in 0/12 fetuses (0%) at 11-12 weeks, 5/20 (25%) at 13-14 weeks, and in 21/28 (75%) at 17-19 weeks. CONCLUSIONS: Blood velocity is higher in the umbilical vein at the abdominal wall than the cord, particularly after 13 weeks of gestation. If acceleration of blood velocity at the umbilical ring is a sign of a narrow inlet, it seems that a progressive tightening occurs during the second trimester. PMID- 11281464 TI - Maternal plasma glucose at high altitude. AB - OBJECTIVE: To compare plasma glucose in pregnant women living at very high altitude; pregnant women living at sea level; non-pregnant women living at very high altitude; and non-pregnant women living at sea level. DESIGN: Cross sectional study. PARTICIPANTS: Ninety-four pregnant women attending for routine antenatal care at 8-42 weeks of gestation in Cerro de Pasco, Peru which is situated 4370 m above sea level; 122 pregnant women in Lima, which is at sea level; 22 non-pregnant women in Cerro de Pasco; and 31 non-pregnant women in Lima. METHODS: Plasma glucose concentrations were measured in samples obtained from the antecubital vein between 8 am and 10 am after an overnight period of fasting for at least 10 hours. RESULTS: Fasting plasma glucose was lower in women at high altitude than in those at sea level, and in both groups the level was lower in pregnant women than in non-pregnant controls. The body mass index was not significantly different between all four groups, and it did not have a significant independent contribution in explaining the variance in fasting plasma glucose. CONCLUSION: Women native at high altitude have lower plasma glucose concentrations before and during pregnancy than those at sea level. PMID- 11281465 TI - Biochemical markers of maternal bone turnover are elevated in pre-eclampsia. AB - OBJECTIVES: To investigate the hypothesis that bone turnover is reduced in pre eclampsia compared with normal pregnancy. DESIGN: A prospective cross-sectional study. SETTING: Obstetric unit at the Chelsea and Westminster Hospital, London. METHODS: Third trimester maternal plasma levels of the cross-linked carboxyl terminal telo-peptide of type I collagen (ICTP), (a marker of bone resorption) and the carboxyl terminal pro-peptide of type I pro-collagen (PICP), (a marker of bone formation) were compared in 25 women with pre-eclampsia and in 24 normal pregnant controls. The subjects were matched for maternal age, booking body mass index and gestational age. PICP and ICTP levels were measured by radio immunoassay. RESULTS: ICTP and PICP levels were significantly increased in women with pre-eclampsia compared with the normal pregnant controls (P = 0.0001 and P = 0.004, for ICTP and PICP respectively, Wilcoxon signed ranked test). There was no significant correlation between either of the markers and booking body mass index, blood pressure, serum uric acid levels or platelet count. CONCLUSIONS: These data suggest that bone turnover is increased in established pre-eclampsia compared with normal pregnancy. Further studies are required to investigate whether this precedes the onset of the disease. PMID- 11281466 TI - Fetal and maternal lactate increase during active second stage of labour. AB - OBJECTIVE: To determine longitudinally fetal and maternal blood lactate concentrations during the second stage of labour. DESIGN: Prospective, observational study of randomly selected labours. SETTING: Labour ward, Sultanah Aminah General Hospital, Johore Bahru, Malaysia. MAIN OUTCOME MEASURES: Fetal scalp and maternal venous blood lactate, umbilical arterial and vein lactate and acid-base balance at delivery. RESULTS: Sixty-nine women and their infants were monitored in the second stage of labour. Mean maternal venous lactate by the end of the first stage was 2.6 +/- 1.0 (+/- S.D.) mmol/L and increased to 3.6 +/- 1.4, 4.2 +/- 1.7. 4.8 +/- 1.6, 5.4 +/- 2.1 and 4.3 +/- 0.9 mmol/L, respectively, for every 15 minute of bearing down. Corresponding values for fetal scalp blood lactate were 2.4 +/- 1.1, 3.1 +/- 1.6, 3.2 +/- 1.8, 4.2 +/- 2.4, 4.9 +/- 2.8 and 5.8 +/- 1.9 mmol/L. The mean slope of maternal lactate increase was 0.070 mmol/L per minute (95% CI 0.050, 0.090) and for fetal lactate increase 0.032 mmol/L per minute (95% C.I.: 0.018, 0.045). The duration of active second stage was significantly associated with fetal lactate (P < 0.001) and maternal lactate (P = 0.03) at the time of crowning of the fetal head, and lactate in umbilical arterial and vein blood at delivery (P < 0.001). Expulsion time > or = 45 minutes, compared with shorter active second stage, and acidaemia at birth implied larger arterial-venous lactate differences (P < 0.001). Fetal lactate at crowning was also significantly associated with the umbilical arterial-veonus lactate difference (P = 0.03). CONCLUSIONS: Maternal and fetal lactate concentrations increase significantly with duration of the active second stage of labour, more rapidly in the mother. It is likely that fetal anaerobic metabolism is the main source for the fetal lactate increase. PMID- 11281467 TI - Differences in hormone replacement therapy use by social class, region and psychological symptoms. AB - OBJECTIVE: To describe the relationship between socio-demographic factors, heart disease risk factors, psychological symptoms and the use of hormone replacement therapy by English women. DESIGN: Cross-sectional analysis of a population-based survey. SETTING: England. POPULATION: 13,214 women aged 40-69 years who participated in the nurse-administered schedule of the Health Survey for England between 1993 and 1996. OUTCOME: Current hormone replacement therapy use. RESULTS: Women from social classes II and I and women who live in the south of England were more likely to use hormone replacement therapy independently of a range of socio-demographic factors including education. The adjusted odds ratio for social classes II and I compared with social classes IV and V was 1.51 (95% CI 1.20 to 1.91) and for women in the South of England was 1.38 (95% CI 1.18 to 1.62). Women with a history of heart disease and those with high cholesterol levels were less likely to use hormone replacement therapy. Women with psychological symptoms were more likely to be prescribed hormone replacement therapy, as were those who had recently seen a doctor. CONCLUSION: There is marked socio-demographic inequity in use of hormone replacement therapy. This may accentuate existing inequalities in health and reduce any potential benefits of Hormone Replacement Therapy for public health. The relationship between psychological symptoms, use of medical services and use of hormone replacement therapy suggests that hormone replacement therapy is prescribed for the management of psychological symptoms. PMID- 11281468 TI - A comparison of isosorbide mononitrate, misoprostol and combination therapy for first trimester pre-operative cervical ripening: a randomised controlled trial. AB - OBJECTIVE: To determine whether a combined therapy with isosorbide mononitrate (40mg) and misoprostol (400 microg) for pre-operative cervical ripening in the first trimester would result in improved clinical effectiveness, and fewer side effects compared with each agent used alone. DESIGN: Randomised controlled trial. SETTING: Glasgow Royal Infirmary. POPULATION: Sixty-six primigravid women scheduled for suction termination of pregnancy. METHODS: Women were randomly assigned to receive before surgery, per vaginam, isosorbide mononitrate 40 mg (n = 22), misoprostol 400 microg (n = 22) or both agents together [isosorbide mononitrate 40 mg and misoprostol 400 microg] (n = 22). MAIN OUTCOME MEASURES: 1. To assess the cumulative force required to dilate the cervix to 8 mm; 2. the onset of new symptoms before termination of pregnancy. RESULTS: The cervical resistance following combination therapy with isosorbide mononitrate and misoprostol was not significantly different than following misoprostol alone [24.5N vs 18.5N; median difference (95% CI) 19N (-22 to 49)]. Pre-treatment with misoprostol used alone resulted in a lower cervical resistance than isosorbide mononitrate alone (18.5N vs 39N, P = 0.04, Mann-Whitney U test). There was no difference in the number of women remaining asymptomatic following either isosorbide mononitrate or misoprostol or combination therapy [14/22 (64%) vs 11/21 (52%) vs 11/22 (50%), Fisher's exact test]. CONCLUSIONS: We have not shown any advantage of combining misoprostol with the nitric oxide donor isosorbide mononitrate compared with misoprostol alone for pre-operative cervical ripening in the first trimester. PMID- 11281469 TI - Psychosocial and other characteristics of women complaining of menorrhagia, with and without actual increased menstrual blood loss. AB - OBJECTIVE: To discover whether psychosocial factors can explain why many women with normal menstrual blood loss seek care for menorrhagia. DESIGN: Cross sectional comparative study of women referred for menorrhagia. SETTING: Gynaecology departments of all five university teaching hospitals in Finland. SAMPLE: Two hundred and twenty-six women aged 35-49 years complaining of menorrhagia. MAIN OUTCOME MEASURES: Several psychosocial factors, seeking medical attention, menstrual blood loss. RESULTS: Twenty-nine percent of the women had their menstrual blood loss in the normal range (menstrual blood loss <60 mL). By univariate analysis, unemployment, anxiety, perceived inconvenience, abdominal pain, haemoglobin level and serum ferritin concentration distinguished this group of women from those with true menorrhagia. Unemployment, perceived inconvenience, abdominal pain and serum ferritin remained significant variables by multivariate analysis. CONCLUSIONS: A significant proportion of women with complaints of menorrhagia have their measured menstrual blood loss within the normal range. Psychosocial factors can have an impact on their seeking health care. Better understanding of the factors, which explain complaints of menorrhagia in women with normal bleeding could improve both medical outcomes and reduce the cost of treatment for menorrhagia. PMID- 11281470 TI - Exploration of cyclical changes in memory and mood in postmenopausal women taking sequential combined oestrogen and progestogen preparations. AB - OBJECTIVE: To investigate the effect of progesterone on cognitive function, mood, sleep quality and libido when added to oestrogen in sequential combined hormonal replacement therapy regimens. DESIGN: Observational study over three hormonal replacement therapy cycles. SETTING: Menopause Centre of Ospedale Maternita, Bologna, Italy. POPULATION: Twenty-three postmenopausal women with an average of 70 months of amenorrhoea (range 12 to 234 months) on different sequential combined hormonal replacement therapy regimen for an average of 15 months (range 3-48) months. METHODS: Psychological testing for memory, mood, sleep quality and libido during the oestrogen only part of the cycle compared with the oestrogen progestogen part of the cycle. RESULTS: Twenty women completed the six visits of the trial. The addition of progestogens to oestrogen appeared to benefit memory (P < 0.01) but worsened mood (P < 0.005). There was no evidence of change in other parameters such as sleep quality or libido. CONCLUSION: The addition of progestogens improved memory above what was obtained with oestrogen alone. This effect did not depend on an improvement of mood since the latter worsened during the progestogenic phase of an hormonal replacement therapy cycle. Progestogen added to oestrogen did not significantly influence sleep or libido. PMID- 11281471 TI - Age at menarche and age at menopause in relation to hepatocellular carcinoma in women. AB - OBJECTIVES: To assess whether age at menarche, age at menopause, parity, and selected blood hormones are associated with risk of hepatocellular carcinoma among women. DESIGN: Case-control. SAMPLE: and setting Data collected from 50 cases of hepatocellular carcinoma among women and 62 female controls with minor trauma or surgical conditions who attended one of three hospitals in Athens, Greece between 1995 and 1998. METHODS: Researchers collected information on Reproductive variables and assayed sera samples for blood hormone levels and for chronic infection with Hepatitis B and C viruses. RESULTS Individuals with hepatocellular carcinoma had a lower mean age at menarche and a significantly higher mean age at menopause. After adjusting for potential confounding, age at menopause remained an important and significant predictor, increasing the risk of hepatocellular carcinoma 24% for each later year of menopause (P < 0.001). For each year that menarche was delayed, risk of hepatocellular carcinoma declined 21% (P = 0.100). Mean levels of insulin-like growth factor-1 and its binding protein were significantly reduced in cases compared with controls, while levels of oestradiol, testosterone and sex hormone binding globulin were somewhat higher among the cases. CONCLUSIONS: This study provides indirect, but converging evidence that steroid hormones in general, and oestrogens in particular, play an important role in the aetiology of hepatocellular carcinoma among women. PMID- 11281472 TI - The effect of stopping smoking on cervical Langerhans' cells and lymphocytes. AB - OBJECTIVE: To investigate the effects of stopping smoking on cervical Langerhans' cells and lymphocytes. DESIGN: Prospective intervention study. SETTING: A large family planning clinic in central London. POPULATION: Women volunteers prepared to attempt to give up smoking for six months. Their most recent cervical smear showed no abnormality greater than mild dyskaryosis. METHODS: The women were seen at three-month intervals for six months. Reduction in smoking was assessed by self-reporting and validated by salivary cotinine concentrations. Colposcopy and a biopsy of a normal area were performed at the first and last visits. Any area of abnormality was also biopsied at the final visit. Langerhans' cells and lymphocytes were counted. MAIN OUTCOME MEASURES: Proportional changes in counts of Langerhans' cells and lymphocytes with reduction in smoking. RESULTS: Reduction in smoking by 20 to 40 cigarettes per day was significantly associated with a reduction of between 6% and 16% in counts of Langerhans cells, CD8 and total lymphocytes. Heavy smoking was significantly associated (P = 0.02) with an increased chance of persistent human papillomavirus infection. The presence of candida was associated with significantly higher counts of between 41% and 47% in total lymphocytes and CD8 lymphocytes. In contrast, the presence of anaerobic vaginosis was associated with significantly lower counts of between 16% and 30% in Langerhans cells, CD4 and CD8 lymphocytes. CONCLUSIONS: This large intervention study has demonstrated a clear relationship between reduction in smoking and changes in cervical immune cell counts. Future studies need to take into account cytokine interactions, which recent studies suggest may be significant in the immune response to both human papillomavirus and cervical intraepithelial neoplasia and the ever-increasing complexity of the cell-mediated immune system of the cervix. PMID- 11281473 TI - Return of fertility in nulliparous women after discontinuation of the intrauterine device: comparison with women discontinuing other methods of contraception. AB - OBJECTIVE: To clarify the effect of the using the intrauterine device on fertility in nulliparous women. DESIGN: Prospective cohort study of two groups of nulliparous women, one recruited while using an intrauterine device and the other while using an oral contraceptive. SETTING: Seventeen family planning clinics in England and Scotland. SAMPLE: 1,071 nulliparous, married women, aged 18-40 years, 558 of whom contributed information to the main objective of the study. METHODS: The women were recruited between 1982 and 1985 and followed up annually to 1994. Dates and reasons for any contraceptive method changes (which were most frequently to barrier methods) were recorded, together with the outcome of any pregnancies, at each follow up. MAIN OUTCOME MEASURES: The number of nulliparous women giving birth at term after stopping contraception (oral contraceptive, intrauterine device or barrier method) in order to conceive. RESULTS: Women who stopped using a barrier method to achieve a planned pregnancy conceived most quickly: 54% were delivered after one year vs 39% of intrauterine device and 32% of oral contraceptive users (log rank P = 0.002). There was no association between fertility and duration of oral contraceptive use: However, short term intrauterine device users (< 42 months) showed a fertility pattern more favourable than seen in those discontinuing oral contraceptives, with increasing duration of intrauterine device use being associated with decreasing fertility (linear trend P = 0.005); the fertility of women who had used the intrauterine device for 78 + months was the most impaired (28% were delivered by 12 months vs 46% of short term users; at 36 months the corresponding figures were 79% vs 91%). This association remained after adjusting for potential confounding factors, including maternal age, husband's social class, and history of gynaecological illnesses, factors which themselves had independent associations with fertility. CONCLUSIONS: Long term intrauterine device use in nulliparous women appears to be associated with an increased risk of fertility impairment. PMID- 11281474 TI - Inadequate repeatability of the one-hour pad test: the need for a new incontinence outcome measure. AB - OBJECTIVE: To assess the reproducibility of two one-hour pad tests performed within one week using serial ultrasound scanning to obtain identical bladder volumes, and to measure the effect of patient anxiety upon test reproducibility. DESIGN: Prospective observational study. SETTING: Tertiary urogynaecological unit. SAMPLE: Fifty-six incontinent women undergoing 112 pad tests. METHOD: Two one-hour pad tests were performed with natural diuresis one week apart prior to treatment. At the second test, serial ultrasound scans were performed until bladder volume reached that of the first test, followed by identical provocation. MAIN OUTCOME MEASURES: One-hour pad loss, bladder volumes (Vol1, Vol2), anxiety VAS questionnaire. RESULTS: Despite serial scanning, bladder volumes differed significantly. Median volume before second pad test was 541 mL, compared with 433 mls before first test (P < 0.001). The second pad test was also significantly larger than first (median 16g vs 4g, P = 0.017), and 13/56 (23%) women were dry on the first test but incontinent on the second. In 26 women (46%) both bladder volumes were similar, but the second pad loss was still significantly greater (median 14g vs 4g, P = 0.037). The mean difference between tests was 10g and the limits of agreement were wide (ranging from -44 to +66 g difference for the test result). Women were more anxious about leaking during the first test (Median VAS during the first test was 2.8cm, compared with 0.6cm during the second test, P = 0.008). 42.5% found the second test to be more typical. CONCLUSION: In women with similar bladder volumes, the test-retest reliability of the one-hour pad test was judged to be clinically inadequate, as the first and second pad test could differ by -44 to +66g. Lower anxiety levels at the second test may account for this finding. The one-hour pad test is a useful baseline measure of incontinence, but the poor repeatability suggests that is not an optimal measure of post-treatment change. PMID- 11281475 TI - Bladder neck mobility in continent nulliparous women. AB - OBJECTIVE: To evaluate the mobility of the vesical neck during coughing and valsalva in healthy nulliparous volunteers and to test the reliability of the technique applied. DESIGN: Clinical observational study. SETTING: Department of Obstetrics and Gynaecology, Cantonal Hospital Lucerne, Switzerland. POPULATION: Thirty-nine nulliparous volunteers. METHODS: Vesical neck motion was assessed with perineal ultrasound. Intra-abdominal pressure was controlled for with an intrarectal probe. Intra-rater reliability was evaluated. RESULTS: Vesical neck mobility was significantly lower during coughing (8 mm, SD 4 mm) than during valsalva (15 mm, SD 10 mm) (P < 0.005). Between individuals mobility varied from 4 mm to 32 mm during coughing and from 2 mm to 31 mm during valsalva. Test-retest studies showed a maximum difference between to tests during coughing of 4 mm and during valsalva of 5 mm. CONCLUSION: The bladder neck is mobile in normal continent women and bladder neck mobility is lower during coughing than during Valsalva. PMID- 11281476 TI - Can a cyclo-oxygenase type-2 selective tocolytic agent avoid the fetal side effects of indomethacin? AB - We evaluated the efficacy and safety of nimesulide (100 mg orally twice daily for > 48 hours) in a pilot series of five women (two with twin pregnancies) at 24(+6) weeks (range 21(+3) - 27(+2)) in preterm labour which was unresponsive to intravenous ritodrine. Nimesulide therapy was continued for eight days (5-16) and was associated with a prolongation of pregnancy of 27 days (6-69). Oligohydramnios occurred in all seven fetuses after three to nine days of therapy, and in the five pregnancies that continued after discontinuation of nimesulide, it resolved within four days (2-7). None of the babies manifested permanent renal damage. PMID- 11281477 TI - Thromboembolism in pregnant women with mechanical prosthetic heart valves anticoagulated with low molecular weight heparin. PMID- 11281478 TI - An endometrioid tumour of the ovary presenting with hyperandrogenism, secondary polycythaemia and hypertension. PMID- 11281479 TI - Osseous metaplasia of cervical epithelium. PMID- 11281480 TI - Prenatal diagnosis of the Wolf-Parkinson-White-syndrome by fetal magnetocardiography. PMID- 11281481 TI - Fibroid embolisation: a technique not without significant complications. PMID- 11281482 TI - Leptomkeningocoele: a rare complication of ventouse delivery. PMID- 11281483 TI - Maternal height and external pelvimetry to predict cephalo-pelvic disproportion in nulliparous African women. PMID- 11281484 TI - The misoprostal third stage study: a randomised controlled comparison between orally administered misoprostol and standard management. A double-blind placebo controlled randomised trial of misoprostol and oxytocin in the management of the third stage labour. PMID- 11281485 TI - Esophageal carcinoma in Southern Thailand. AB - BACKGROUND: Southern Thailand is an area with a high frequency of esophageal carcinoma. This paper presents basic data regarding esophageal carcinoma patients from this region. METHODS: Patients with histopathological confirmed esophageal carcinoma were retrospectively reviewed. The age, sex, location of tumor and resectability were studied. RESULTS: A total of 813 cases of esophageal carcinoma were reviewed, male:female ratio was 3.54:1 (634:179). Average age in males was 64.62 years and 64.30 years in females. The peak age-incidence was 51-70 years. Squamous cell carcinoma was most commonly found in the mid thoracic portion of the esophagus with 369 cases (45.39%), 70 cases (8.61%) were found in the cervical portion of the esophagus. Adenocarcinoma cancer was found at the esophagogastric junction in 47 cases (5.78%). Only 293 cases (36.04%) were operable. Respiratory tract involvement was noted in 49 cases. CONCLUSION: The most common type of esophageal cancer in Southern Thailand is squamous cell carcinoma, as in other countries in Asia. The status of the patients, advanced age and locally advanced tumor were major factors of our low operable rate. PMID- 11281487 TI - Actinomycosis of the urinary bladder. AB - The case of a 49 year-old female patient, with frequency of urination for 2 years, having a mass at the anterior bladder wall and at the anterior abdominal wall is reported. Cystoscopy found an impression of the anterior bladder wall and hyperremic edematous bladder mucosa. Pre-operative computerized tomography suspected bladder tumor. Laparotomy revealed an inflammatory firm mass at the anterior bladder wall and another mass at the anterior abdominal wall. Partial cystectomy and excision of the mass at the anterior abdominal wall were performed. After the pathological examination confirmed actinomycosis, the patient was treated post operatively with penicillin. She recovered well. PMID- 11281486 TI - A reevaluation of antibiotic prophylaxis in laparoscopic cholecystectomy: a randomized controlled trial. AB - BACKGROUND: To assess the result of antibiotic prophylaxis in low-risk patients undergoing elective laparoscopic cholecystectomy with respect to the postoperative septic complications. METHOD: One hundred and two low-risk patients were randomized into 1 of 2 treatment arms (1) cefazolin 1 g intravenously after induction of anesthesia (PA group) and (2) no prophylactic antibiotics (NONE group). Laparoscopic cholecystectomy was attempted in all cases. The patients were followed-up for postoperative septic complications for at least 30 days at the out-patient clinic or by telephone contact. In both groups, sex, age, weight, American Society of Anesthesiologists patient classification score, operative time, surgical techniques, number of port sites, intraoperative cholangiograms, intraoperative gallbladder rupture, postoperative hospital stay, and postoperative septic complications were compared. The statistical analysis of data performed by computer program SPSS 10.0 for Windows was based on the Independent-Samples T Test or the Pearson Chi-Square (2-sided). RESULTS: There was only one minor problem of superficial wound infection in the NONE group. Comparison of data showed no statistically significant difference between the groups. CONCLUSION: Antibiotic Prophylaxis may not be necessary in low-risk patients undergoing elective laparoscopic cholecystectomy. PMID- 11281488 TI - Idiopathic sudden sensorineural hearing loss. AB - BACKGROUND: Sudden sensorineural hearing loss is one of the most controversial unsolved mysteries in Otolaryngology. Lack of a universally accepted definition of sudden sensorineural hearing loss, insufficient knowledge of pathogenesis, lack of a standard method for evaluating the patients, in addition to a high spontaneous recovery rate, all complicate the study of sensorineural hearing loss and the investigation of different treatment modalities. OBJECTIVE: To study the clinical manifestation and prognostic factors, which influence the recovery of hearing in sudden sensorineural hearing loss. PATIENTS AND METHOD: Patients with idiopathic sudden sensorineural hearing loss who were admitted to Srinagarind Hospital from January 1994 to December 1998 were included. The clinical manifestations, audiograms and investigations of these patients were analysed. RESULTS: Of the fifty-six patients, who met the criterion, 34 were females and 22 males. The average age of onset was 43.7 years (SD = 13.46, range = 13-66 years). The onset of hearing loss was sudden in 50 per cent of cases, whereas, 46.4 per cent of cases were noted on awakening in the morning and the remainder had rapidly progressive hearing loss. The hearing loss was unilateral in 92.9 per cent of cases. 96.4 per cent of the patients had tinnitus and 66.1 per cent of the patients had vertigo. 64.3 per cent of the patients had some degree of recovery (complete recovery in 28.6% and partial recovery in 35.7%). The severity of hearing loss significantly influenced the outcome of the patients. CONCLUSION: Approximately two-thirds of the patients with idiopathic sudden hearing loss had some degree of recovery. Among contributing factors, only the severity of hearing loss significantly influenced the prognosis. PMID- 11281489 TI - The long term outcome of thirty eight post-transfusion hepatitis C. AB - Thirty-eight cases of post-transfustion HCV hepatitis have been followed for 5-24 years. Cirrhosis and hepatocellular carcinoma were found in 44.7 per cent and 13.1 per cent respectively. Cirrhosis was recognised by pathological evidence as early as one and a half years after transfusion and the clinical evidences of decompensated cirrhosis were noted in the fifth year post-transfusion onward. Hepatocellular carcinoma was first recognised in year ten and thereafter. Nine patients died of liver failure or hepatocellular carcinoma during years 8-16 of the follow-up. Therefore, it is of utmost importance to screen out the HCV infected blood donors and to treat the HCV patients as early and as effectively as possible. PMID- 11281490 TI - Needle stick injuries during medical training among Thai pre-clinical year medical students of the Faculty of Medicine, Chulalongkorn University. AB - OBJECTIVE: To study episodes of needle stick injuries during medical training among pre-clinical year medical students of the Faculty of Medicine, Chulalongkorn University. SETTING: Faculty of Medicine, Chulalongkorn University. DESIGN: Retrospective descriptive study. SUBJECTS: 375 pre-clinical year medical students of the Faculty of Medicine, Chulalongkorn University during academic year 1998-1999. METHOD: Questionnaire survey and interviewing. RESULTS: In this study only 5 students revealed that they had ever had accidental exposure during venipuncture training. Each episode of exposure was different in the details. Most of the accidents occurred after venipuncture practice (80%). CONCLUSION: Although the universal precautions are taught in many subjects, we also found that accidental exposures still occur. Not only prevention of accidents but also post-exposure management should be frequently reiterated to the medical students at every level. Any medical practice of students should be under supervisor control. PMID- 11281491 TI - Syringomyelia as a complication of tuberculous meningitis. AB - Tuberculous meningitis (TBM) is a common manifestation of extrapulmonary tuberculosis. Syringomyelia is a rare complication of TBM. We report a case of syringomyelia due to TBM. A 25 year old Thai male was admitted with a history of progressive paraparesis and loss of body sensation. He had a history of TBM in the previous year, and was treated with antituberculous drugs. Physical examination revealed a temperature of 37 degrees C. Motor power was grade 3/5 with generalized hyperreflexia. He had bilateral loss of pain, temperature and vibratory sensation below the T7 level. A magnetic resonance imaging of the spine demonstrated a long segment of syrinx from C4 to the conus medullaris region. A T12-L1 laminectomy and syringosubarachnoid shunt were done. His clinical symptoms improved after surgery. PMID- 11281492 TI - Pulmonary sparganosis: a case report with five years follow-up. AB - Sparganosis has a world wide distribution, but only a few patients have pulmonary involvement. The term sparganosis is defined as an infection by the larva of parasitic tapeworms of Spirometra species. We present here-in a patient, who was infected by this parasite and had pulmonary symptoms. The chest roentgenography revealed diffuse multiple nodular infiltration with cavitations. Bronchoscopy with a transbronchial lung biopsy was nondiagnostic. Finally, open lung biopsy was performed, and the histologic examination revealed plerocercoid larva of sparganum. The patient was treated with mebendazole 40 mg/kg/day for 6 months and his symptoms and pulmonary function improved. In the 5th year of follow-up, he presented with more progressive dyspnea and developed cor pulmonale, and finally died from pneumonia with sepsis. The objective of this report was to present a rare manifestation of sparganosis and it's clinical course. Currently, there is no known effective treatment for this disease. PMID- 11281493 TI - Insulinoma in childhood. AB - A 9-year-old boy with convulsions is herein described. He was diagnosed and treated for epilepsy and insufficient adrenal function for four years with no response. Hypoglycemia from hyperinsulinism was found and the source of the hyperinsulinism was a tumor of the tail of the pancreas--located by computerized tomographic scan and magnetic resonance imaging. Distal pancreatectomy was performed with good results. Histology of the tumor showed islet cell tumors with capsular invasion. For this type of patient, long-term follow-up should include: prevention of metastasis or recurrence, and testing for multiple endocrine neoplasia type 1. PMID- 11281494 TI - Prenatal diagnosis of VACTERL association: a case report. AB - A prenatal diagnosis of VACTERL association, a combination of vertebral (V), anal (A), cardiac (C), tracheoesophageal (TE), renal (R) and limb (L) anomalies was made at 30 weeks of gestation, based on the sonographic demonstration of vertebral defects, bilateral renal agenesis, and left lower limb defects. Additionally, severe oligohydramnios and fetal growth restriction were also documented. After proper counseling, elective termination of pregnancy was done, resulting in a stillborn infant with multiple malformations compatible with the VACTERL association. The postnatal X-ray and autopsy revealed verterbral defects, anorectal atresia with undetermined sex, cardiac defect of ventricular septal defect, tracheal agenesis with distal atresia of esophagus, bilateral renal agenesis, and limbs defects. The chromosomal study revealed normal, 46,XY. This report emphasizes the important role of prenatal ultrasound in the diagnosis and management of this disorder. PMID- 11281495 TI - Aerobic microbiological study in term pregnant women with premature rupture of the membranes: a case-control study. AB - To determine the aerobic microorganisms related to premature rupture of the membranes (PROM) in term pregnant women, a case-controlled study was performed on pregnant women delivered at Rajavithi Hospital between November 1, 1996 and July 30, 1997. Two hundred and twenty pregnant women with PROM and 220 pregnant women without PROM were recruited by simple random sampling. The diagnosis of rupture of the membrane was made by history and by positive microscopic ferning and pH testing performed during speculum examination. The demographic characteristics were not statistically significantly different between both groups. We could not isolate any organisms (35.9% in the study group and 49.5% in the control group). Candida albicans and Klebsiella pneumoniae were the only two significant differences demonstrated between the study and control group (p<0.05). Candida albicans, the most prevalent organism in the study group, demonstrated significant difference between the study and control group (14.5% and 7.7% respectively) (p<0.05). Klebsiella pneumoniae demonstrated significant difference between the study and control group (7.30% and 4.10% respectively) (p<0.05). Gardnerella vaginalis, the most prevalent organism in the control group, showed no significant difference between the control and study group (16.40% and 14.10% respectively) (p=0.547). PMID- 11281496 TI - Drug addicts treatment for ten years in Thanyarak Hospital (1989-1998). AB - The problem of drug addicts has increased over the past 3-4 years. A retrospective descriptive study of inpatient drug addicts at Thanyarak Hospital was done. Data from the record pool from October 1989 to September 1998 was reviewed for demographic information, diagnosis, HIV infection and the outcome of treatment. The study showed that the total number of patients increased from 7,595 cases in 1989 to 10,661 cases in 1995, but decreased in the next three years to 7,633 cases in 1998. Males constituted more than 91.5 per cent in each year. Mean ages decreased from 31.1+/-8.8 to 26.5+/-9.3 years. Most of the patients were labourers or were unemployed. The number of students also increased. Initially, students constituted only 1.3 per cent but in the last 3 years this increased to 4.0, 8.0 and 17.1 per cent, respectively. The education level has gradually increased. Heroin addiction was approximately 80.6-92.4 per cent in the first 8 years but markedly decreased to 38.0 per cent in the last year. Opium addiction decreased from 3.8 per cent to 1.0 per cent. Methamphetamine addiction markedly increased from 0.4 per cent to 51.5 per cent. The outcome of the treatments showed that patients who attended the 2-3 weeks detoxification treatment program was 30.9-43.5 per cent but patients who joined the therapeutic community rehabilitation program was only 0.8-4.2 per cent. The mortality rate increased from 2.1 per thousand to 5.2 per thousand and more HIV infected patients died than non-HIV infected patients. This rate varied from 1.7 times in 1989 to 8 times in 1995 and 4.5 times in 1996. We conclude from this study that drug addicts changed from heroin to Methamphetamine especially among young students in the last 2-3 years. HIV infection was still high in old cases (about 40%). PMID- 11281497 TI - Sonographic morphology scores (SMS) for differentiation between benign and malignant ovarian tumor. AB - The objectives of this cross-sectional descriptive analysis are to determine the sensitivity and specificity of sonographic morphology scores (SMS) in distinguishing between benign and malignant ovarian tumors and to determine the best cut-off score. The study was conducted at the Department of Obstetrics and Gynecology, Faculty of Medicine, Chiang Mai University. Two hundred and forty eight nonpregnant patients scheduled for elective surgery for ovarian tumors between July, 1996 and March, 1998 were recruited into the study and were sonographically examined in 24 hours of surgery by the same sonographer to evaluate inner wall structure, wall thickness, septum, echogenicity and score of the tumors. The final diagnosis was pathologically confirmed as the gold standard. It was found that the score of 9 from reciever operating characteristic curve was the best cut-off score, giving the sensitivity of 93.1 per cent and specificity of 75.6 per cent. In conclusion, the SMS system is probably useful in distinguishing ovarian malignancy from benign ovarian tumor. PMID- 11281498 TI - The long term results of internal fixation of displaced intra-articular calcaneal fractures. AB - One hundred and fourteen displaced intra-articular fractures of the calcaneus (47 tongue type and 67 joint depression type) were treated by open reduction and internal fixation with multiple H plate and were followed-up for an average of 6.75 years. All the fractures healed radiographically in 3 months. Average time for bone healing was 2.25 months for the tongue type and 2.42 months for the joint depression type (p > 0.05). Average post operative Bohler tuber angle was 25.05 and 22.71 degrees in the tongue type and joint depression type respectively (p > 0.05). All patients could return to work in 8 months. The average time for returning to work was 3.42 months in the tongue type and 4.16 months in the joint depression type (p < 0.05). Three calcanei (2.6%) had wound infection. There were significant differences between the two types in degree of residual pain, walking activities and range of subtalar joint movement. There were no significant differences in the ability to work, change of footwear and hind foot swelling. The end functional result was rated as excellent in sixty one (53.5%), good in twenty (17.5%), fair in fourteen (12.3%) and poor in nineteen (16.7%) calcanei. There was significant difference in the satisfactory result (excellent to fair) between the tongue type (91.5%) and joint depression type (77.6%). PMID- 11281499 TI - The metabolic and bone density effects of continuous combined 17-beta estradiol and noresthisterone acetate treatments in Thai postmenopausal women: a double blind placebo-controlled trial. AB - OBJECTIVE: To compare the changes of lipid parameters, liver function tests, fasting plasma glucose and bone density in Thai postmenopausal women who received this combined hormonal treatment and placebo. STUDY DESIGN: Double-blinded, randomized controlled trial study. MATERIAL AND METHOD: Sixty postmenopausal women attending the menopause clinic at Chulalongkorn Hospital from July, 1996 to December, 1996, were enrolled in the study. The patients were randomized to receive the placebo or drug (17 beta-estradiol 2 mg and norethisterone acetate 1 mg) continuously. Patient characteristics, physical examination, liver function tests, fasting plasma glucose, lipid parameters (fasting total cholesterol, low density lipoprotein cholesterol (LDL), high density lipoprotein (HDL) and triglyceride level) and bone densitometry were performed before beginning the study. The lipid parameters were repeated at 3, 6 and 12 months. Fasting plasma glucose, liver function tests and bone densitometry were repeated at 12 months. RESULTS: In the drug group, there were significant changes in the cholesterol at 3, 6 and 12 months when compared to the baseline. There were significant differences at 3, 6 and 12 months when compared between groups. The HDL values were not significantly different within groups. The LDL values at 3, 6, 12 months were significantly lower than the baseline in the drug group when compared within groups and at 6, 12 months in the placebo group. The triglyceride values were not significantly different between groups and within groups. There was no significant change between groups and within groups of fasting plasma glucose, total bilirubin, direct bilirubin, AST, ALT, albumin and globulin. The alkaline phosphatase values were significantly decreased at 12 months in the drug group. The bone density of total BMD and T-score at the spine of the drug group increased significantly at 12 months. The per cent change per year was +5.1. In contrast, the values in the placebo group decreased significantly, the per cent change per year was -0.9. The same pattern was also found in the bone density of the total hip. But when focused to the femoral neck, we found no significant change in both groups. CONCLUSION: This continuous combined treatment resulted in beneficial changes of bone density and lipid profiles. The therapy prevented bone loss and the changes in serum lipoprotein were concordant with a lipid profile associated with a decreased risk of coronary heart disease in Thai postmenopausal women. PMID- 11281500 TI - An open, baseline controlled evaluation of sertraline safety and efficacy in the treatment of depression in Thai patients. AB - An open, baseline controlled study of sertraline in depressed patients was conducted in 6 treatment sites. Eighty-two patients between 20-82 years of age with DSM III-R diagnosis of a depressive illness received sertraline 50-200 mg/day. Among evaluable patients, there was a significant reduction in depressive symptoms at the final visit. A statistically significant change from baseline in Montgomery Asberg Depression Rating Scale (MADRS), Hospital Anxiety Depression Rating Scale (HAD), and Clinical Global Impression Severity of Illness Scale (CGI S) scores was demonstrated. On the basis of MADRS criterion, 96.0 per cent of patients responded and on the basis of CGI-S criterion, 86.6 per cent of patients responded. In 73.2 per cent of patients the final sertraline dosage was 50 mg. All-cause adverse events were recorded in 35 patients (42.7%), whereas 22 (26.8%) had adverse events that were judged treatment-related. The most frequently reported events were nausea and headache. Overall, the patients tolerated sertraline very well. The results of the study suggest that sertraline is an effective, well-tolerated and safe treatment for depression in Thai patients. PMID- 11281501 TI - Stricture of the male urethra: 29 years experience of 323 cases. AB - The authors reported 323 cases of male urethral stricture managed at Ramathibodi Hospital from 1969 to 1998 (29 years period). Etiology included traumatic causes 237 (73%) and post infection 54 cases (16%). The managements were urethroplasties 281 cases (87%), urethrotomy 21 cases (6%) and dilatation 21 cases (6%). The over all successful rate of urethroplasty was 89 per cent. The mean follow-up time was 2.5 years (0.5-15 years). PMID- 11281502 TI - Clinical features of septic arthritis of sternoclavicular joint. AB - We studied 21 patients with septic arthritis of the sternoclavicular joint at Chulalongkorn University Hospital between January 1987 and January 1997. There were 15 males (71.4%) and 6 females (28.6%). The mean age was 47.4 years with a range of 16 to 69. More than half of the patients (57.1%) were aged more than 50 years and most had associated diseases including diabetes mellitus and cirrhosis. Almost all of the younger age group had a history of intravenous drug abuse. All of the patients had fever and sternoclavicular joint pain. Most of the patients (66.7%) had monoarticular arthritis, whereas, the others had oligoarticular arthritis. Staphylococcus aureus was the most commonly or identified organism in the patients. Retrosternal abscess was seen by computerized tomography in 6 patients (28.6%). All patients received parenteral antibiotics, and 5 patients (23.8%) required surgical drainage of a retrosternal abscess. Eighteen patients recovered but there were 3 (14.3%) deaths. All of these had retrosternal abscesses. The major cause of death was septic shock. Septic arthritis of the sternoclavicular joint is an uncommon disease in Thai clinical practice. Although uncommon, retrosternal abscess is a life threatening complication. PMID- 11281503 TI - Chlamydia pneumoniae in community-acquired pneumonia. AB - Chlamydia pneumoniae has been established recently as an important human respiratory pathogen. The aim of this study was to define the prevalence of C. pneumoniae in community-acquired pneumonia. We prospectively investigated adult patients who were treated as inpatients and outpatients. Acute and convalescent serum samples were obtained from each patient. Serological diagnosis of C. pneumoniae infection was determined by enzyme-linked immunosorbent assay (ELISA). Eighty paired sera were tested for C. pneumoniae-specific IgM, IgG and IgA. Twenty-one patients (26.2%) had serological results compatible with acute C. pneumoniae infection. Eighteen (85.7%) of these infected patients were C. pneumoniae-specific IgM positive, three had a seroconversion of IgA and two had a four-fold or greater increase in C. pneumoniae-specific IgG antibody titer. The most common clinical manifestations of community-acquired pneumonia due to C. pneumoniae were fever (100%), cough (100%), chest pain (47.6%) and shortness of breath (42.9%). Physical examination revealed crackle in 85.7 per cent of the cases. These findings suggest that C. pneumoniae is a common cause of community acquired pneumonia in Thailand. PMID- 11281504 TI - Hepatic arterial collaterals after transcatheter oily chemoembolization of hepatocellular carcinoma. AB - From July 1989 to June 1999, 100 patients, 76 male and 24 female, were admitted for treatment of hepatocellular carcinoma (HCC) with transcatheter oily chemoembolization (TOCE) using lipiodol 10 ml mixed with an anticancer drug (mitomycin C 20 ml) and gelfoam particles, described by Nakamura H et al. The periodic follow-up angiogram showed hepatic collaterals which developed according to the mode of embolization. For peripheral hepatic arterial embolization such as segmental or lobar arterial embolization, the intrahepatic collaterals were commonly demonstrated. However, for more proximal hepatic arterial embolization of the tumor feeder arteries such as the proper hepatic artery and the common hepatic artery, the extrahepatic collaterals were commonly demonstrated with fine, small tortuous vasculatures, rendering a repeat TOCE more difficult. The hepatic collaterals are presented. PMID- 11281505 TI - Effect of long-term intake of Asian food with different glycemic indices on diabetic control and protein conservation in type 2 diabetic patients. AB - The study was carried out in 10 females with type 2 diabetes aged 32-60 yrs. All of them were receiving weight-maintaining diets composed of 12 per cent protein, 30 per cent fat and 58 per cent carbohydrate. The only difference among all study diets was the types of complex carbohydrate used. High-glycemic diet (HG) or low glycemic diet (LG) consisted mainly of glutinous rice or mungbean noodles and the intermediate-glycemic diet (DM) was solely white rice. After the metabolic evaluation of the baseline diet (BL), each subject was placed on DM and followed randomly by HG and LG or vice versa for 4 weeks each. The diurnal plasma glucose levels tended to be lowest after LG. The integrated plasma glucose levels among all diets were not different. The integrated insulin levels after DM and LG did not differ but they were lower than HG and BL. Long-term ingestion of all test diets spilt less urinary glucose than BL, the lowest was LG. HbA1 levels and nitrogen balance after all diets were better than BL, the best was LG. It was concluded that in addition to strict dietary control, ingestion of mungbean noodles (a low glycemic diet) without increasing fiber intake, can improve diabetic control and protein conservation in type 2 diabetes. PMID- 11281507 TI - Towards the modal ECT treatment. PMID- 11281506 TI - The efficacy of terbutaline and magnesium sulfate in the management of preterm labor. AB - Ninety-six patients with preterm labor at 28 weeks to 35 weeks gestation were randomized to terbutaline or magnesium sulfate untill 36 weeks gestation, 25 patients were excluded from the study. Of the remaining 71 patients, 35 patients received terbutaline and 36 patients received magnesium sulfate. The result of the study showed that, there were no significant differences (P > 0.05) regarding time to stop, mean gestational age at delivery, time gained, failure rate, time to recurrent labor and readmission for recurrent labor, birth weight, apgar score and fetal survival. Serious maternal side effects were not observed with terbutaline or magnesium sulfate, although the majority of women also received dexamethasone. Neither drug caused serious adverse neonatal effects. PMID- 11281508 TI - Apolipoprotein E polymorphism and response to electroconvulsive therapy. AB - The aim of this study was to determine whether the apolipoprotein E genotype differs in patients who respond or do not respond to electroconvulsive therapy (ECT). Inpatients, out-patients, and day-treatment patients who had received ECT comprised the study group. The 34 patients included met DSM-III-R criteria for affective or schizoaffective disorder. Responder or nonresponder status was assessed using the Clinical Global Inventory and Montgomery Asberg Depression Rating Scale. Blood samples were taken and coded when the patients entered the study. DNA extraction and apolipoprotein E genotyping were performed with no knowledge of the clinical classification of the patients. A significant difference in E4 genotype distribution was found between ECT responders and nonresponders (p < 0.02); psychosis was significantly less frequent in this group (p = 0.046), and there was a trend toward older onset of depression among these persons (p = 0.10). Only the E3/3 genotype was found in the patients with early onset depression. The E4 genotype appears to define a subgroup of patients with late-onset depression who respond to ECT. If confirmed in prospective studies, this may provide a useful marker in the treatment decision-making process for late-onset depression. PMID- 11281509 TI - Changes in regional cerebral blood flow after electroconvulsive therapy for depression. AB - Fifteen patients with major depression and normal results of magnetic resonance imaging or computed tomographic studies were treated by electroconvulsive therapy (ECT). The regional cerebral blood flow (rCBF) of these patients was imaged using Tc-99m hexamethylpropylene amineoxime single-photon emission computed tomography before and after treatment, and their images were compared with a population of 11 healthy volunteers. Before ECT treatment, the patients had hypoperfusion of the frontal region compared with the controls, and they had multiple areas of altered perfusion throughout the brain. Five of the patients had an excellent clinical response to ECT; these patients also showed changes toward normal in rCBF. The remaining patients had minimal to moderate clinical response and showed no significant change in rCBF. These results indicate that improvement in clinical status as a result of ECT is correlated with a change toward normal in rCBF. PMID- 11281510 TI - Effectiveness of ECT combined with risperidone against aggression in schizophrenia. AB - Aggressive behavior in schizophrenic patients can often be problematic not only for the patients themselves, but for their families and others. This study examined the effect of electroconvulsive therapy (ECT) in combination with risperidone in an open trial in 10 male schizophrenic patients with significant aggressive behaviors. Patients were given bilateral ECT five times a week in combination with risperidone. The mean total number of times of ECT was 6.6 (range 5-9). The aggressive behavior in five of the six patients, who showed positive symptoms, was rapidly ameliorated within 12 days. The ECT/risperidone regimen also eliminated aggressive behavior in four patients showing no positive symptoms within 10 days. These treatment effects lasted for at least 6 months in 9 (of the 10) patients. The results suggest that ECT, combined with risperidone, produce a rapid and effective elimination of aggressive behaviors in schizophrenic patients. In addition, there was a resolution of aggression in four patients with no positive symptoms. This suggests that aggression in some schizophrenic patients develops as a primary symptom of schizophrenia and is not related to other positive symptoms of the disease or the patient's personality traits. PMID- 11281511 TI - ECS-Induced mossy fiber sprouting and BDNF expression are attenuated by ketamine pretreatment. AB - Recent evidence suggests hippocampal and possibly cortical atrophy is associated with major depression. Chronic electroconvulsive seizures (ECS) induce brain derived neurotrophic factor (BDNF) expression and sprouting of the mossy fiber pathway in the hippocampus, effects that may be related to electroconvulsive therapy's (ECT) mechanism of action. The objective of this study was to investigate the role of NMDA (N-methyl-D-aspartate) receptor in mediating the ECS induced mossy fiber sprouting and BDNF expression. Timm histochemistry and in situ hybridization methodologies were used to determine the effect of pretreatment with ketamine, an NMDA antagonist, on ECS-induced sprouting and BDNF expression. The results demonstrate the ability of ketamine pretreatment to attenuate ECS-induced sprouting in the dentate gyrus and BDNF expression in the medial prefrontal cortex and the dentate gyrus. In addition, we found a significant decrease in seizure duration with ketamine pretreatment. These data suggest that NMDA receptor activation contributes to both the regulation of neurotrophic factor expression and the morphological changes associated with seizure activity. However, other effects resulting from shortened seizure duration and seizure intensity cannot be excluded. These findings are of increasing interest, as they relate to the use of ECT in the treatment of depression, and the specific anesthetic agents that are used. PMID- 11281512 TI - Nicardipine improves the antidepressant action of ECT but does not improve cognition. AB - INTRODUCTION: Cognitive impairment, the most important adverse effect of electroconvulsive therapy (ECT), may involve elevated intracellular calcium ion signaling. Animal research suggests that calcium channel-blocking agents, which attenuate excessive intracellular calcium activity, may reduce cognitive dysfunction caused by ECT. METHOD: The lipid-soluble calcium channel-blocking drug nicardipine or matching placebo were randomly assigned to 26 patients with major depressive disorder receiving ECT. A rater blind to the experimental condition administered the Hamilton Depression Rating Scale, the Montgomery Asberg Depression Rating Scale, the Beck Depression Inventory, the Mini-Mental State Examination and a comprehensive battery of neuropsychological tests prior to ECT, at the completion of ECT, and 6 months after ECT completion. RESULTS: Compared with patients receiving placebo, patients taking nicardipine had significantly lower scores on the Hamilton and Montgomery-Asberg but not the Beck Depression rating scale scores at the completion of ECT. There were no differences between placebo and nicardipine groups in depression scores 6 months after ECT. Cognitive function declined over the course of ECT and improved over the next 6 months in both groups, but changes were statistically significant for only two subtests on the neuropsychological battery. Changes in Mini-Mental State Examination scores were small and were not significant at any point. There were no significant differences between nicardipine and placebo treated groups in any assessment of cognition. DISCUSSION: Standard approaches to ECT in younger patients without preexisting neurological impairment do not produce cognitive side effects of sufficient severity for calcium channel-blocking agents to reduce these side effects demonstrably. Studies of treatments for cognitive impairment should be conducted in patients with risk factors for more severe cognitive impairment such as geriatric patients or patients with a history of interictal delirium during previous treatment with ECT. A possible effect of nicardipine in enhancing the antidepressant action of ECT requires further investigation in a study designed to test this action. PMID- 11281513 TI - The effects of ECT on brain glucose: a pilot FDG PET study. AB - BACKGROUND: Regional brain activity was measured before and after electroconvulsive therapy (ECT) using [18F]-fluorodeoxyglucose (FDG) positron emission tomography (PET). METHODS: 6 patients (4 females) with major depression were free of psychotropic medications for at least 2 weeks prior to baseline FDG scans. Patients were treated with bifrontotemporal ECT, and posttreatment scans were obtained after the last treatment. RESULTS: A region of interest (ROI) analysis of absolute metabolic rate showed a decrease in CMRglu after ECT in all 61 regions examined. In 17 of the 61 regions, the decrease was significant at the p < 0.05 level. In the right parietal lobe, and the right anterior and left posterior frontal lobes, the decrease in CMRglu significantly correlated with the decrease in Hamilton Depression Rating Scale (HDRS) scores (r = 0.83, 0.82, and 0.84, respectively). The analysis of CMRglu normalized to global metabolic rate showed significant increases in 8 of 61 regions, including basal ganglia, upper brainstem, and occipital lobe. DISCUSSION: The decreases in global glucose metabolism and correlation of changes in frontal metabolism with decreases in HDRS are consistent with earlier brain imaging studies of ECT. The relative increases in CMRglu observed in regions with known dopaminergic innervation (caudate and upper brainstem) have not been previously reported. PMID- 11281514 TI - Nitroprusside and ECS-induced retrograde amnesia. AB - Previous research found that the administration of verapamil and felodipine immediately before electroconvulsive shocks (ECS) attenuated ECS-induced retrograde amnesia. This study examined whether sodium nitroprusside, an antihypertensive drug that does not affect calcium channels, has a similar action. Adult male Sprague-Dawley rats received nitroprusside (0.5 mg/kg ip) or saline 3 minutes before each of three once-daily true or sham ECS. Retention of pre-ECS learning was studied 1 day after ECS using a passive avoidance task. Nitroprusside was associated with increased seizure duration in ECS-treated rats, and with enhanced recall in both true and sham ECS groups. The latter finding suggests that nitroprusside nonspecifically improves cognitive functions, and does not support the hypothesis that ECS-induced cognitive impairment is a result of blood-brain barrier breach. Nitric oxide mechanisms may underlie the benefits purveyed by nitroprusside. PMID- 11281515 TI - Severity of subcortical gray matter hyperintensity predicts ECT response in geriatric depression. AB - OBJECTIVE: To determine the effect of subcortical white and gray matter lesions on ECT outcome. METHOD: 41 geriatric psychiatric inpatients underwent an MRI scan during their ECT work-up. Periventricular, deep white matter, and subcortical gray matter hyperintensities were graded. The associations of low versus high hyperintensity ratings and symptom scores, Clinical Global Impression severity (CGS) ratings, Montgomery-Asberg Depression Scale score, and number of treatments were examined using t-tests and repeated measures ANOVA. RESULTS: Patients with more severe subcortical gray hyperintensities (SCG) had significantly less improvement as measured by CGS ratings. CONCLUSIONS: SCG severity may limit the improvement of patients receiving ECT. Further studies are needed to examine differences based on electrode placement and to determine whether patients with severe SCG may require more ECT treatments in an index course. PMID- 11281516 TI - The outcome of 369 ECT consultations. AB - OBJECTIVE: The components of a pre-ECT consultation have been well-described, but the outcome has not been described. We describe the outcome of 369 consecutive ECT consultations. METHODS: We performed a retrospective review of ECT consultations performed at Wake Forest University School of Medicine between January 23, 1992, and October 22. 1998. Each consultation was coded as either recommending against ECT, unenthusiastic about ECT, or recommending ECT. RESULTS: Thirteen percent of the patients at their first consultation needed clarification of their capacity to consent to ECT. Additional testing was recommended in 34%, and additional medical consultation was recommended for 11% of the patients. The ECT consultation recommended against ECT for 4% of patients, was unenthusiastic for an additional 15% of patients, and was enthusiastic for 81%. Likelihood of receiving ECT was strongly influenced by the consulting physician's level of enthusiasm for ECT. Enthusiasm for ECT, in turn, was highly related to diagnosis. CONCLUSIONS: The recommendations from an ECT consultation appeared influential in the likelihood of receipt of ECT. The consultation's enthusiasm for ECT, in turn, was related to the patient's diagnosis. Furthermore, the consultation revealed the need for additional testing, medical consultation, or clarification of capacity to consent in a substantial number of patients. PMID- 11281517 TI - Electroconvulsive therapy and Friedreich's ataxia. AB - Friedreich's ataxia is commonly associated with depression. Treatment of the depression can be difficult due to numerous morbid medical conditions. ECT is a safe and effective treatment option. PMID- 11281518 TI - ECT for prolonged catatonia. AB - OBJECTIVE AND BACKGROUND: Electroconvulsive therapy (ECT) is highly effective for acute catatonia but its use in prolonged catatonia is not well established. We report three cases of prolonged catatonia with medical complications or comorbidities treated by ECT. METHOD: Case reports. RESULTS: A 24 year-old woman developed fever and autonomic instability after parenteral neuroleptics. Catatonia and autonomic signs persisted for 14 weeks. After minimal improvement from lorazepam, 15 bilateral ECTs led to resolution. A 26-year-old woman with a history of lupus erythematosus, complicated by lupus cerebritis with lesions in the cortex and basal ganglia and a communicating hydrocephalus, was catatonic for 9 weeks. Lorazepam produced marginal improvement. A series of 14 bilateral ECTs led to improved mobility, speech, and interaction, but the response was less robust than Case 1. A 40-year-old man with mental retardation and intermittent psychosis developed severe neuroleptic malignant syndrome and remained catatonic for 4 months. After lorazepam produced minimal improvement, his catatonia resolved with 20 bilateral ECTs. CONCLUSIONS: ECT may improve prolonged catatonia with complex medical comorbidities, but may require many treatment sessions. Gross cerebral pathology may predict a less robust response. As for acute catatonia, ECT may resolve prolonged catatonia after benzodiazepines have failed. PMID- 11281519 TI - Extended continuation and maintenance ECT for long-lasting episodes of major depression. AB - While most major depressive episodes are relatively short lasting, a substantial minority of patients with severe mood disorders respond poorly to available treatments and remain ill for extended periods of time. The possible role of continuation and maintenance electroconvulsive therapy (ECT) in the management of such patients has not been clearly established. Three patients are reported whose long-lasting, relatively medication refractory episodes of major depression were treated with ECT for both acute and continuation and maintenance purposes. Although relapses occurred despite treatment, the frequency, severity, and duration of relapses were apparently diminished by ECT. Relapses occurred with diminishing frequency over the course of individual episodes, and were typically reversed by three or four closely spaced ECT treatments. No medical complications occurred. While complaints of memory difficulty were common, Mini-Mental Status Examinations 6 weeks after ECT were unimpaired. Continuation and maintenance ECT appears to be a safe and effective treatment modality for long-lasting episodes of major depression. Recommendations for clinical management are proposed. PMID- 11281520 TI - ECT for major depression and mania with advanced dementia. AB - Two patients with advanced dementia and severe affective disorders were successfully treated with electroconvulsive therapy (ECT) without significant adverse effects. These reports illustrate that ECT can be effective for depression and mania even when complicated by moderate or severe dementia. PMID- 11281521 TI - Electroconvulsive therapy-responsive depression in a patient with progressive supranuclear palsy. AB - We report the case of a 68-year-old woman with progressive supranuclear palsy whose depression was successfully treated with electroconvulsive therapy. She tolerated the treatments well and showed neither improvement nor decline in the neurologic symptoms of her illness. PMID- 11281522 TI - Successful ECT in a patient with hydrocephalus, shunt, hypopituitarism, and paraplegia. AB - A 25-year-old patient with paraplegia, hypopituitarism, hydrocephalus, and a ventriculoperitoneal shunt was successfully treated with a course of bilateral electroconvulsive therapy (ECT) for major depression. Brain imaging studies and neurology/ endocrinology consultations were obtained prior to the use of ECT. Throughout the course of ECT, his replacement hormonal therapy continued. Prior to each ECT, additional parenteral hydrocortisone was also administered. Consistent with the previously published reports, the patient did not experience any neurological deterioration. A brief review of the literature on the use of ECT in patients with panhypopituitarism, spinal cord injury, and hydrocephalus is presented. PMID- 11281523 TI - Quantifying the ECT dose: the right unit remains elusive. PMID- 11281524 TI - Invited editorial: A window of opportunity for radiation research. PMID- 11281525 TI - Preliminary studies to develop a personal dosemeter for use by aircraft crew. AB - This paper describes preliminary work to develop a cosmic-radiation dosemeter for use by military aircraft crew. The dosemeter is based on a combination of CR-39 etched-track detectors and TLD-700 thermoluminescent detectors. It is intended that the CR-39 be used to assess the neutron dose, while the TLD-700 is used to assess the photon and charged particle dose. The sensitivity of CR-39 to the neutron component of cosmic radiation was estimated by irradiating samples of the plastic at the CERN-CEC High Energy Reference Field Facility. This facility produced a radiation field with a neutron spectrum resembling that of the neutron component of cosmic radiation at typical airflight altitudes. The response of the CR-39 was linear over the range of doses studied (0.2-6.0 mSv) and there was no significant fading in the 6-month period after irradiation. The TLD-700 component of the dosemeter was calibrated using 137Cs gamma rays. The response of the TLD 700 was linear over the range of doses studied (0.01-5.0 mSv) with no significant fading in the 6-month period after irradiation. It was concluded that a combination of CR-39 and TLD-700 detectors would provide an effective cosmic radiation dosemeter for use by military aircraft crew. PMID- 11281526 TI - Transfer across the human gut of environmental technetium in lobsters (Homarus gammarus L.) from the Irish Sea. AB - Few data are available on the uptake by the human gut of the element technetium. Of current radiological interest in connection with discharges of technetium-99 in liquid discharges from BNFL, Sellafield, is uptake from European lobsters (Homarus gammarus), whose edible parts are known to concentrate technetium. In this study, a group of eight adult volunteers (six males and two females) ate samples of edible flesh from lobsters caught off the west Cumbrian coast and provided 24 h samples of urine and faeces for analysis. Detection of uptake from the gut by difference between intake and faecal measurements proved insensitive, suggesting a low value of the gut transfer factor (f1 value) of up to 0.1 with a maximum (two standard deviations) level of about 0.3. In urine, technetium was detectable at a relatively low level compared with the intakes, consistent with a low absorption across the gut. Values for f1 were derived with the aid of literature data for excretion following intravenous administration of technetium 95 m as pertechnetate, and gave averaged data for f1 in the range 0.046 to 0.23. These results are in broad conformity with those derived from the faecal measurements, and suggest a lower value than the 0.5 used by ICRP. PMID- 11281527 TI - Kinetics of systemic ruthenium in human blood using a stable tracer. AB - The biokinetics of ruthenium after oral and intravenous administration has been investigated in two human subjects using the stable isotope 101Ru as a tracer. Tracer concentrations in blood plasma have been determined using activation analysis with protons. The results presented here prove that the stable tracer technique is a valuable tool for obtaining relevant information about the biokinetics of ruthenium in humans. From these pilot studies, it may be argued that the clearance of systemic ruthenium from plasma is significantly slower than the predictions of the biokinetic model currently recommended by the International Commission on Radiological Protection (ICRP). The experimental data for the orally administered tracer, which reflect the gastrointestinal absorption process, differ from the curve derived from the ICRP model, suggesting that the uptake into the systemic circulation may be lower than predicted. On the basis of these preliminary data, investigations on a larger number of subjects with improvements in the experimental design are scheduled. PMID- 11281528 TI - The evasive plausibility. AB - A frequent practical problem is to assess the probability of a potential single event (X). The event probability P(X) is usually conditional to some assumption (A) and is then written P(X / A). Whether the assumption is valid or not is an unknown fact, but its validity may be assigned a weight psi(A) indicating the observer's belief in its validity. The weighted quantity would be the unconditional probability of X if the weighting factor were a true probability. However, it is not a stochastic quantity even though in Bayesian statistics it would be treated as a probability. It follows that the weighted quantity, i.e. psi (A) x P(X / A), is not an unconditional probability in the usual sense. It is suggested that it be given a special name, for example 'plausibility'. PMID- 11281529 TI - Implementation of an electronic personal dosimetry system (EPD) at Oldbury-on Severn power station. AB - This article presents the implementation of an electronic personal dosemeter (EPD) as a film badge replacement at Oldbury-on-Severn power station, which is the first major site to use an approval issued by the UK Health and Safety Executive (HSE) for dose measurement by an EPD. The practicalities and history behind the introduction of an EPD for personal dosimetry are described. PMID- 11281530 TI - Invited editorial: Radiation exposures of aircrew in high altitude flight. PMID- 11281531 TI - ICRP and UNSCEAR: some distant memories. International Commission on Radiological Protection. United Nations Scientific Committee on the Effects of Atomic Radiation. AB - The operations of ICRP and UNSCEAR have been partly complementary during the past half century, and there has been considerable overlapping of membership of the two bodies. From its inception in 1955 UNSCEAR collected imformation upon which estimates of radiation risk have been based; these have been used by ICRP as a background to its recommendations during recent decades. Over the years there have been many interesting incidents connected with the work of these two bodies, in which a number of memorable characters participated. Some of these are recalled. PMID- 11281532 TI - ICRP: preserving a valuable asset. International Commission on Radiological Protection. PMID- 11281533 TI - Radon exposure and the risk of lung cancer. PMID- 11281534 TI - Bone cancer risk. PMID- 11281535 TI - Healthy worker effect. PMID- 11281537 TI - Contaminated watches on sale in France. PMID- 11281536 TI - Depleted uranium. PMID- 11281538 TI - Radiological Protection Institute of Ireland examination of storage of liquid high-level waste at Sellafield. PMID- 11281539 TI - UNSCEAR report 2000: sources and effects of ionizing radiation. United Nations Scientific Comittee on the Effects of Atomic Radiation. PMID- 11281540 TI - A critical review of the system of radiation protection. First reflections of the OECD Nuclear Energy Agency's Committee on Radiation Protection and Public Health (OECD/NEA, 2000). PMID- 11281541 TI - Invited editorial: Gut transfer and doses from environmental technetium. PMID- 11281544 TI - Workshop on Comparative Radiobiology and Protection of the Environment. PMID- 11281545 TI - Effect of tauroursodeoxycholate and S-adenosyl-L-methionine on 17beta-estradiol glucuronide-induced cholestasis. AB - BACKGROUND/AIMS: S-adenosyl-L-methionine (SAMe) and tauroursodeoxycholate (TUDC) exert an additive ameliorating effect on taurolithocholate (TLC)-induced cholestasis. The aims were to investigate the protective effect of SAMe on 17beta estradiol-glucuronide (17betaEG) cholestasis and to find out whether SAMe and TUDC may exert an additive, ameliorating effect. METHODS: Hepatocyte couplet function was assessed by canalicular vacuolar accumulation (cVA) of cholyllysylfluorescein (CLF). Cells were co-treated with 17betaEG and SAMe, TUDC, or both (protection study), or treated with 17betaEG and then with SAMe, TUDC or both (reversion study) before CLF uptake. Couplets were also co-treated with SAMe and dehydroepiandrosterone (DHEA), a competitive substrate for the sulfotransferase involved in 17betaEG detoxification. The effects of 17betaEG, SAMe and TUDC were also examined on intracellular distribution of F-actin. RESULTS: Both SAMe and TUDC significantly protected against, and reversed, 17betaEG-induced cholestasis, but their effects were not additive. DHEA abolished the protective effect of SAMe. 17BetaEG did not affect the uptake of CLF into hepatocytes at the concentrations used, and also, it did not affect the intracellular distribution of F-actin. CONCLUSIONS: 17BetaEG does not affect the uptake of CLF into hepatocytes. SAMe and TUDC protect and reverse 17betaEG induced cholestasis, but without an additive effect. Protection by SAMe may involve facilitating the sulfation of 17betaEG. PMID- 11281546 TI - Short-chain ceramide regulates hepatic methionine adenosyltransferase expression. AB - BACKGROUND: The metabolism of methionine plays an important role in regulating hepatic cellular function. Methionine adenosyltransferase (MAT) is the enzyme that catalyses the biosynthesis of S-adenosylmethionine (AdoMet) from ATP and methionine. Liver-specific MAT I/III levels are down-regulated in the regenerating rat liver after partial hepatectomy. Tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) are two cytokines fundamental for liver regeneration. TNF-alpha stimulates sphingomyelin metabolism and ceramide generation in a variety of cell systems. AIMS: The role of exogenous cell permeable ceramide in modifying MAT I/III mRNA levels and its association with TNF-alpha and IL-6 actions were investigated in rat hepatocytes and H35 hepatoma cells. RESULTS: C2-ceramide (N-acetylsphingosine) at 1-10 microM decreased MAT I/III expression. The effect was maximum after 2 h of treatment and it was maintained up to 24 h. MAT I/III protein levels also decreased. IL-6 (1-10 ng/ml) potentiated C2-ceramide effects in cultured hepatocytes while decreasing by itself MAT I/III levels with a similar time-response curve in both cell types. C2 ceramide actions were not associated with an increase in cell death. TNF-alpha was also a potent antagonist for MAT I/III expression, at 1-20 ng/ml decreased MAT I/III levels and induced endogenous ceramide generation. The decrease of MAT I/III mRNA levels (in all the cases) was not due to a decrease in mRNA half-life which suggests a regulation at the transcriptional level. Finally, the decrease in MAT I/III mRNA levels correlated to a decrease in MAT activity. CONCLUSION: This work demonstrates that short-chain ceramide can be used as a novel exogenous agonist that can modulate hepatic methionine metabolism in association with cytokines. PMID- 11281547 TI - Dietary cholesterol does not normalize low plasma cholesterol levels but induces hyperbilirubinemia and hypercholanemia in Mdr2 P-glycoprotein-deficient mice. AB - BACKGROUND/AIMS: Mdr2 P-glycoprotein deficiency in mice (Mdr2(-/-) leads to formation of cholesterol/cholesterol-depleted bile and reduced plasma HDL cholesterol. We addressed the questions: (1) does HDL in Mdr2(-/-) mice normalize upon phospholipid and/or cholesterol feeding, and (2): is the Mdr2(-/-) liver capable of handling excess dietary cholesterol. METHODS: Male and female Mdr2(-/ ) and Mdr2(+/+) mice were fed diets with or without additional phosphatidylcholine and/or cholesterol. Plasma, hepatic and biliary lipids as well as liver function parameters and expression of transport proteins involved in bile formation were analyzed. RESULTS: Feeding excess phospholipids and/or cholesterol did not affect lipoprotein levels in Mdr2(+/+) or Mdr2(-/+) mice. Dietary cholesterol caused hyperbilirubinemia (male +100%; female +500%) and elevated plasma bile salts (male +200%; female +1250%) in Mdr2(-/-) mice only, independent of phospholipids. Bile flow nor biliary bile salt and bilirubin secretion were affected in cholesterol-fed Mdr2(-/-) mice. Elevated plasma bile salts may be related to cholesterol-induced reduction of hepatic Na+-taurocholate cotransporting protein expression in Mdr2(-/-) mice. CONCLUSION: Excess dietary phospholipids and cholesterol do not normalize low HDL associated with Mdr2 P glycoprotein-deficiency. Induction of hyperbilirubinemia and hypercholanemia by dietary cholesterol in Mdr2(-/-) mice delineates the important role of biliary lipid secretion in normal hepatic functioning. PMID- 11281548 TI - Differential regulation of UDP-GlcUA transport in endoplasmic reticulum and in Golgi membranes. AB - BACKGROUND: In the endoplasmic reticulum (ER), the stimulation of UDP glucuronosyltransferase (UGT) by UDP-GlcNAc is based on the interaction of transport across the ER membrane of UDP-GlcUA with UDP-GlcNAc. Intramicrosomal UDP-GlcNAc stimulates influx of UDP-GlcUA and thereby enhances delivery of UDP GlcUA to the catalytic center of UGT in the ER lumen. AIM: The aim of this study is to investigate whether the interactions between nucleotide sugars for transport across the ER membrane also occur in the Golgi apparatus, and thereby affect UGT activity in Golgi membranes. RESULTS: We found that Golgi membrane preparations display UGT activity which, unlike in ER membranes, is not stimulated by UDP-GlcNAc. Efflux of intravesicular UDP-GlcNAc and UDP-Xyl marginally enhanced uptake of UDP-GlcUA in Golgi vesicles; such trans-stimulation was much more pronounced in the ER. Efflux of intravesicular UDP-GlcNAc was strongly trans-stimulated by cytosolic UDP-GlcUA in ER-derived vesicles but less so in Golgi-derived vesicles. CONCLUSION: The interaction between transport of UDP-GlcUA and transport of UDP-GlcNAc or UDP-Xyl is different in Golgi vesicles compared with ER vesicles. This finding is consistent with the different effects of UDP-GlcNAc on glucuronidation in Golgi and ER. PMID- 11281549 TI - Ascitic fluid carcinoembryonic antigen and alkaline phosphatase levels for the differentiation of primary from secondary bacterial peritonitis with intestinal perforation. AB - BACKGROUND/AIMS: In cirrhotic patients, spontaneous bacterial peritonitis (SBP) may be difficult to distinguish from secondary peritonitis with occult intestinal perforation; Runyon's criteria (based on ascitic fluid glucose, protein and lactate dehydrogenase levels) are sensitive but not specific. Ascitic fluid carcinoembryonic antigen (CEA) and alkaline phosphatase (AP) are potential markers for secondary peritonitis. METHODS: Ascitic fluid CEA and AP levels were prospectively compared among three subject groups--cirrhotic patients with sterile ascites, cirrhotic patients with SBP, and patients (cirrhotic and non cirrhotic) with perforation-related secondary peritonitis. RESULTS: The secondary peritonitis group (n = 38 including 11 cirrhotic patients) had significantly higher mean CEA and AP levels than the SBP (n = 34) and sterile ascites patients (n = 63). Of secondary peritonitis patients, 92% fulfilled predetermined criteria (either CEA >5 ng/ml or AP >240 units/l) versus only 12% of SBP patients; sensitivity was 92% and specificity 88% for differentiating secondary peritonitis from SBP. Runyon's criteria had a sensitivity of 97% and specificity of 56%. Stratification of secondary peritonitis patients by the presence or absence of cirrhosis did not alter our results. CONCLUSIONS: Ascitic fluid CEA or AP elevations appear to be sensitive and specific markers for perforation-related secondary peritonitis in cirrhotic as well as non-cirrhotic patients. PMID- 11281550 TI - Human hepatic stellate cells secrete adrenomedullin: potential autocrine factor in the regulation of cell contractility. AB - BACKGROUND/AIMS: Hepatic stellate cells (HSCs) are perisinusoidal pericytes which have receptors for vasoactive factors, such as endothelin-1, which can regulate cell contractility in an autocrine manner. It is unknown whether human HSCs have receptors for and are able to synthesize the vasodilator peptide adrenomedullin (ADM), a peptide produced by most contractile cells. METHODS AND RESULTS: Stimulation of HSCs with ADM resulted in a dose-dependent raise in cAMP concentration (radioimmunoassay) and markedly blunted the endothelin-induced increase in [Ca2+]i and cell contraction, as assessed in cells loaded with fura-2 using a morphometric method. The existence of the receptor CRLR for ADM and their associated proteins RAMP-1 and RAMP-2 was demonstrated by reverse transcriptase polymerase chain reaction (RT-PCR). Moreover, activated human HSCs spontaneously secreted ADM in the culture medium in a time-dependent manner. ADM secretion was markedly enhanced by tumour necrosis factor-alpha and interleukin-1beta. Specific mRNA for ADM (RT-PCR and Northern blot) was detected in HSCs and increased after incubation of cells with cytokines. CONCLUSIONS: Human HSCs have functional receptors for ADM, the stimulation of which blunts the contractile effect of endothelin-1. Cultured human HSCs secrete ADM in baseline conditions. This secretion is markedly increased by cytokines. These results suggest that ADM can regulate HSCs' contractility in an autocrine manner. PMID- 11281551 TI - Effect of aldosterone on collagen steady state levels in primary and subcultured rat hepatic stellate cells. AB - BACKGROUND/AIMS: Activation of the renin-angiotensin-aldosterone system can lead to collagen accumulation and reactive myocardial fibrosis. This study aims at evaluating the effect of aldosterone on extracellular matrix synthesis by rat hepatic stellate cells. METHODS: Cultured cells were treated with different concentrations of aldosterone (10(-6)-10(-10) M) and metabolically labeled with 35S-methionine/35S-cysteine. Procollagen types I, III and IV, laminin and fibronectin were specifically immunoprecipitated and quantified by phosphor imaging. Using the reverse transcription-polymerase chain reaction, we investigated the expression of the mineralocorticoid receptor in hepatic stellate cells. RESULTS: Quantitation showed that 10(-6) M aldosterone induced procollagen type I synthesis significantly, whereas procollagen type IV expression was significantly affected by 10(-9) and 10(-10) M aldosterone, both in primary hepatic stellate cells. RT-PCR experiments clearly demonstrated a lack of expression of the mineralocorticoid receptor in hepatic stellate cells. CONCLUSION: We demonstrated that aldosterone altered moderately procollagen type I and IV synthesis by primary hepatic stellate cells, but not by activated stellate cells which are the principal cellular sources of extracellular matrix proteins in chronic liver disease. Moreover, hepatic stellate cells do not express the mineralocorticoid receptor, suggesting that the observed modest changes of extracellular matrix synthesis are probably due to mineralocorticoid receptor unrelated mechanisms. PMID- 11281552 TI - Matrix metalloproteinase (MMP)-2, MMP-7, and tissue inhibitor of metalloproteinase-1 are closely related to the fibroproliferative process in the liver during chronic hepatitis C. AB - BACKGROUND/AIMS: To study whether expression of matrix metalloproteinases and their inhibitors correlate with ongoing fibrogenesis, we measured hepatic mRNA levels of matrix metalloproteinase (MMP) -2, MMP-7, and MMP-9 as well as tissue inhibitor of metalloproteinase (TIMP) -1, TIMP-2, and TIMP-3 and compared it to histology, procollagen IV alpha-1 chain mRNA levels, and biochemical parameters in patients with chronic active hepatitis C (CAH). METHODS: Quantitative reverse transcription-polymerase chain reaction/enzyme-linked immunossorbent assay using in vitro transcribed competitor and standard RNA were performed from ten normal livers (N), 29 CAH liver biopsies and seven samples with hepatitis C virus (HCV) induced end-stage cirrhosis (Ci). RESULTS: From N to Ci both TIMP and MMP RNA expression increased. However, none of the RNA levels differed significantly between CAH patients with and without fibrosis. Non-parametric correlation analysis and receiver operating characteristics curves show that MMP-2, MMP-7, and TIMP-1 provide the best discrimination between cirrhosis and pre-cirrhotic stages. They also correlate with histologic and biochemical inflammatory activity and with procollagen IV mRNA. CONCLUSION: Hepatic fibroproliferation is associated with alterations of hepatic TIMP and MMP expression. The relation of hepatic TIMP and MMP mRNA levels to disease stage and inflammatory activity underlines their potential as diagnostic markers in chronic liver disease. PMID- 11281553 TI - Hepatocyte apoptosis is a pathologic feature of human alcoholic hepatitis. AB - BACKGROUND/AIMS: The pathogenesis of alcoholic hepatitis (AH) remains poorly understood. Although apoptosis is now recognized as a mechanism of liver injury, the extent and mechanisms of apoptosis in human AH remain unknown. Thus, our aims were to quantify hepatocyte apoptosis in patients with AH, correlate it with disease severity, and identify the mechanisms of apoptosis induction. METHODS: Hepatocyte apoptosis was assessed in 26 patients with AH and 27 controls without liver disease using the TUNEL assay and immunohistochemistry for activated caspase 3. Liver specimens were also graded for disease severity. The expression of the death receptors, Fas and tumor necrosis factor-alpha receptor 1 (TNF-R1), was assessed by immunohistochemistry. RESULTS: In contrast to normal livers, TUNEL- and caspase 3-positive hepatocytes were readily observed in the livers of patients with AH. In the AH group, hepatocyte apoptosis was significantly higher in patients with a serum bilirubin of > 3 mg/dl. Apoptosis was also greater in grade 4 steatohepatitis. The Fas receptor was strongly expressed in hepatocytes in AH, but not in normal livers; the TNF-R1 expression was comparable in both groups. CONCLUSIONS: The present results demonstrate that hepatocyte apoptosis is significantly increased in human AH and justify therapeutic strategies aimed at inhibiting apoptosis in this disease. PMID- 11281554 TI - Clinical and biological relevance of hepatocyte apoptosis in alcoholic hepatitis. AB - BACKGROUND/AIMS: Although human and experimental studies have shown that apoptosis plays a role in hepatocyte death in alcoholic liver disease, its clinical and biological significance has not been investigated in alcoholic hepatitis (AH). The aim of this study was to quantify hepatocyte apoptosis in AH and to attempt to relate it to the clinical and biological severity of the disease. METHODS: The hepatocyte apoptotic index was determined using a double in situ transferase-mediated dUTP nick end (TUNEL) and CD15 (neutrophils) labelling on 35 liver biopsies from patients with AH lesions of different severities. The specificity of TUNEL labelling for apoptosis was monitored both by morphology and fractin (a caspase actin cleavage site) immunostaining. RESULTS: The hepatocyte apoptotic index ranged from 0.3 to 28% and was related to the severity of alcoholic hepatitis as measured by the Maddrey score (P < 0.05; Mann-Whitney test) while ballooning (which reflects hepatocytes potentially undergoing necrosis) and neutrophil indexes were not. CONCLUSIONS: This suggests that hepatocyte apoptosis could be a therapeutic target to treat or to prevent alcoholic hepatitis in cirrhotic patients. Co-localization of apoptotic hepatocytes with neutrophils and the strong quantitative correlation would suggest an apoptosis dependent transmigration of neutrophils. PMID- 11281555 TI - Mechanisms for experimental buprenorphine hepatotoxicity: major role of mitochondrial dysfunction versus metabolic activation. AB - BACKGROUND/AIMS: Although sublingual buprenorphine is safely used as a substitution drug in heroin addicts, large overdoses or intravenous misuse may cause hepatitis. Buprenorphine is N-dealkylated to norbuprenorphine by CYP3A. METHODS: We investigated the mitochondrial effects and metabolic activation of buprenorphine in isolated rat liver mitochondria and microsomes, and its toxicity in isolated rat hepatocytes and treated mice. RESULTS: Whereas norbuprenorphine had few mitochondrial effects, buprenorphine (25-200 microM) concentrated in mitochondria, collapsed the membrane potential, inhibited beta-oxidation, and both uncoupled and inhibited respiration in rat liver mitochondria. Both buprenorphine and norbuprenorphine (200 microM) underwent CYP3A-mediated covalent binding to rat liver microsomal proteins and both caused moderate glutathione depletion and increased cell calcium in isolated rat hepatocytes, but only buprenorphine also depleted cell adenosine triphosphate (ATP) and caused necrotic cell death. Four hours after buprenorphine administration to mice (100 nmol/g body weight), hepatic glutathione was unchanged, while ATP was decreased and serum transaminase increased. This transaminase increase was attenuated by a CYP3A inducer and aggravated by a CYP3A inhibitor. CONCLUSIONS: Both buprenorphine and norbuprenorphine undergo metabolic activation, but only buprenorphine impairs mitochondrial respiration and ATP formation. The hepatotoxicity of high concentrations or doses of buprenorphine is mainly related to its mitochondrial effects. PMID- 11281556 TI - Eosinophil-induced liver injury: an experimental model using IL-5 transgenic mice. AB - BACKGROUND/AIMS: In certain liver diseases, activated eosinophils are considered to be important effector cells in addition to T-cell-mediated cytotoxicity. No experimental model, however, has been developed for in vivo analysis of the cytotoxic mechanisms. METHODS: Interleukin-5 (IL-5) transgenic mice (C3H/HeN TgN(IL-5)Imeg), which exhibit marked eosinophilia without liver injury, were injected once with 25 microg of lipopolysaccharide (LPS) intraperitoneally. The mice were sacrificed weekly and eosinophilic injuries were assessed microscopically. To clarify the role of Kupffer cells and tumor necrosis factor alpha (TNF-alpha) in the liver injury, gadolinium chloride (GdCl3) and anti-TNF alpha neutralizing antibody were administrated before the LPS injection. RESULTS: Two weeks after injection, transgenic mice exhibited marked infiltration of eosinophils and extensive lobular necrosis. Transmigration of eosinophils through vascular endothelium and degranulation of eosinophil cytotoxic granules in inflamed areas were observed. These eosinophilic injuries were transient, but liver-specific. Pre-administration of GdCl3 and anti-TNF-alpha markedly reduced the hepatic inflammation, suggesting that LPS-activated Kupffer cells play a key role in producing the cytotoxicity of eosinophils by releasing TNF-alpha. CONCLUSIONS: We have established an experimental model of eosinophil-induced liver injury using IL-5 transgenic mice. Since this model is simple and highly reproducible, it will be useful for analysis of in vivo cytotoxic mechanisms of eosinophils. PMID- 11281557 TI - Prednisolone suppresses ischemia-reperfusion injury of the rat liver by reducing cytokine production and calpain mu activation. AB - BACKGROUND: We investigated the effects of prednisolone on cytokine production and calpain mu activation during hepatic ischemia-reperfusion (IR) injury. METHODS: The hilar area of the left lateral and median lobes of rat liver was clamped for 60 min. Prednisolone was administered at 1.0, 3.0, or 10 mg/kg at 30 min before ischemia. In addition to biochemical and microscopic analyses, IL-beta and TNF-alpha production was evaluated by RT-PCR. Calpain mu activation and talin degradation were determined by Western blotting, using specific antibodies. RESULTS: In the control and prednisolone (1.0 mg/kg) groups, serum AST and ALT levels were elevated, and cell membrane bleb formation was observed after 2 h of reperfusion. Moreover, calpain mu activation, talin degradation, and overexpression of IL-beta and TNF-alpha mRNAs were detected. Infusion of prednisolone at 3.0 or 10 mg/kg significantly suppressed biochemical and microscopic changes. At 10 mg/kg, prednisolone markedly suppressed IL-beta and TNF-alpha transcription and calpain mu activation and talin degradation, consistent with the improved 7-day survival after total hepatic ischemia (75% vs. 25% in control group, P = 0.039). CONCLUSIONS: Cytoprotective effect of prednisolone in hepatic IR injury was closely associated with suppression of IL beta/TNF-alpha production and calpain mu activation. PMID- 11281558 TI - Gastrin inhibits cholangiocarcinoma growth through increased apoptosis by activation of Ca2+-dependent protein kinase C-alpha. AB - BACKGROUND/AIMS: We determined the role of gastrin in the regulation of cholangiocarcinoma growth. METHODS: We evaluated for the functional presence of cholecystokinin (CCK)-B/gastrin receptors in the cholangiocarcinoma cell lines, Mz-ChA-1, HuH-28 and TFK-1. We determined the effect of gastrin on the growth of Mz-ChA-1, HuH-28 and TFK-1 cells. We evaluated the effect of gastrin on growth and apoptosis of Mz-ChA-1 in the absence or presence of inhibitors for CCK-A (L 364, 718) and CCK-B/gastrin (L-365, 260) receptors, the intracellular Ca2+ chelator (BAPTA/AM), and the protein kinase C (PKC)-alpha inhibitor, H7. We evaluated if gastrin effects on Mz-ChA-1 growth and apoptosis are associated with membrane translocation of PKC-alpha. RESULTS: Gastrin inhibited DNA synthesis of Mz-ChA-1, HuH-28 and TFK-1 cells in a dose- and time-dependent fashion. The antiproliferative effect of gastrin on Mz-ChA-1 cells was inhibited by L-365, 260, H7 and BAPTA/AM but not L-364, 718. Gastrin induced membrane translocation of PKC-alpha. The inhibition of growth of Mz-ChA-1 cells by gastrin was associated with increased apoptosis through a PKC-dependent mechanism. CONCLUSIONS: Gastrin inhibits the growth of Mz-ChA-1, HuH-28 and TFK-1 cells. Gastrin inhibits growth and induces apoptosis in Mz-ChA-1 cells through the Ca2+ dependent PKC-alpha. The data suggest a therapeutic role for gastrin in the modulation of cholangiocarcinoma growth. PMID- 11281559 TI - A controlled trial of calcitonin therapy for the prevention of post-liver transplantation atraumatic fractures in patients with primary biliary cirrhosis and primary sclerosing cholangitis. AB - BACKGROUND/AIMS: Accelerated bone loss occurs early after liver transplantation (OLT) and, in cholestatic patients with pre-existing osteopenia, causes spontaneous fracturing. This study aimed to investigate the efficacy of calcitonin, a powerful inhibitor of bone resorption, in preventing or reducing the accelerated rate of bone loss and fracturing which occurs in patients with primary biliary cirrhosis and primary sclerosing cholangitis early after OLT. METHODS: Sixty-three patients undergoing OLT for primary biliary cirrhosis (n = 26) and primary sclerosing cholarigitis (n = 37) were randomized to receive: (a), 100 IU/day of salmon calcitonin subcutaneously for the first 6 months posttransplant; or (b), no therapy. At pretransplant, and at 4 and 12 months after OLT, patients were investigated clinically, biochemically, by bone mineral density of the lumbar spine, and by radiographs of the thoracolumbar spine, chest and site of any bone pain. RESULTS: The bone mineral density of the lumbar spine fell equally at 4 months in both groups, from 0.85 to 0.81 g/cm2 in calcitonin treated patients (n = 29) and from 0.88 to 0.82 g/cm2 in controls (n = 34); at 12 months, both groups had stabilized to 0.83 g/cm2. Fracturing was the same in both groups. CONCLUSIONS: Calcitonin therapy for the first 6 months after OLT is unable to prevent or reduce accelerated bone loss or spontaneous fractures which occur in the first posttransplant year. PMID- 11281560 TI - Transdermal oestrogen therapy protects postmenopausal liver transplant women from osteoporosis. A 2-year follow-up study. AB - BACKGROUND/AIMS: Hormone replacement therapy (HRT) prevents osteoporosis in postmenopausal women by inhibiting bone resorption, but the benefits of oestrogen therapy in liver transplant patients have not been studied. METHODS: The effect of transdermal HRT was studied in 33 postmenopausal liver transplant women. The main outcome measure was the change in bone mineral density (BMD) which was measured annually for 2 years. The effect on bone turnover was studied by assessment of the serum aminoterminal propeptide of type I procollagen (PINP). RESULTS: The mean lumbar BMD increased from 0.816 at baseline to 0.858 and to 0.878 g/cm2 (P < 0.001) after 1 and 2 years of therapy, respectively. The BMD of the femoral neck increased from 0.665 to 0.690 g/cm2 (P < 0.006). During the first and second years, the mean BMD of the lumbar spine increased by 5.3 and 1.2%, while that of the femoral neck increased by 3.3 and 1.2%. After 2 years of HRT, only one-fifth of the patients had osteoporosis, whereas over half of the women had osteoporosis at baseline. The median serum PINP decreased by 47% at 1 year and remained decreased at 2 years compared with baseline levels. CONCLUSION: Transdermal HRT decreased the turnover rate of mineralized bone matrix. Transplant women responded with increased BMD, just like healthy postmenopausal women. PMID- 11281561 TI - The long-term outcome of interferon-alpha treated and untreated patients with HBeAg-negative chronic hepatitis B. AB - BACKGROUND/AIMS: This study aimed to evaluate the effect of interferon-alpha therapy on the long-term outcome of HBeAg-negative chronic hepatitis B. METHODS: A cohort of 209 interferon-alpha treated and 195 untreated patients with histologically documented HBeAg-negative chronic hepatitis B were closely followed for a mean of 6 (1-13.5) years. Patients with decompensated liver disease and/or hepatocellular carcinoma at presentation were excluded. RESULTS: Survival and complication (liver decompensation and/or hepatocellular carcinoma) free survival were significantly worse in patients with compared to those without baseline cirrhosis and in patients older compared to those younger than 45 years (P < 10(-4)). Sustained biochemical remission was achieved in 57 treated patients. Sustained responders had significantly better survival and complication free survival than non-sustained responders (P = 0.027 and P = 0.019, respectively) or than untreated patients (P = 0.048 and P = 0.012, respectively). Multivariate analysis showed that absence of baseline cirrhosis, younger age, and sustained biochemical remission were independently associated with an improved outcome. CONCLUSION: In patients with HBeAg-negative chronic hepatitis B, sustained biochemical remission induced by interferon-alpha is associated with improved long-term outcome, even in the presence of cirrhosis and old age, both known factors associated with worse survival. Therefore, long-term biochemical remission appears to represent a satisfactory therapeutic target in this setting. PMID- 11281562 TI - Steatosis and chronic hepatitis C: analysis of fibrosis and stellate cell activation. AB - BACKGROUND/AIMS: Steatosis is a frequent histological finding in chronic hepatitis C and is associated with increased hepatic fibrosis. METHODS: We studied 80 patients with untreated chronic hepatitis C to determine whether steatosis contributes to fibrosis through a steatohepatitis-like pathway. RESULTS: Fine sinusoidal and/or central vein fibrosis was present in 52 patients (65%). This was typically located in acinar zone 3 and had a chicken-wire appearance similar to that seen in steatohepatitis. A statistically significant relationship was found between subsinusoidal fibrosis and age (r(s) = 0.33, P = 0.003) and grade of steatosis (r(s) = 0.35, P = 0.001). Mean body mass index was higher in patients with focal (28.4 +/- 4.7 kg/m2) or extensive (29.6 +/- 5.9 kg/m2) subsinusoidal fibrosis than in those patients with no subsinusoidal fibrosis (25.5 +/- 3.7 kg/m2). The extent of alpha-smooth muscle actin staining (as a marker of stellate cell activation) correlated with the degree of portal inflammation and the stage of portal fibrosis, but not with the grade of hepatic steatosis. CONCLUSIONS: These findings suggest that in hepatitis C infection, host factors, particularly adiposity, contribute to both steatosis and acinar fibrosis. The implication of these observations is that weight reduction may provide an important therapeutic strategy for patients with chronic hepatitis C. PMID- 11281563 TI - Characterization of an immunologically conserved epitope from hepatitis C virus E2 glycoprotein recognized by HLA-A2 restricted cytotoxic T lymphocytes. AB - BACKGROUND/AIMS: Identification of epitopes recognized by cytotoxic T lymphocytes (CTL) in hepatitis C virus (HCV) proteins is of importance because they can be used for vaccination, treatment of infection or monitoring of immune responses. Our purpose was to characterize new CTL epitopes in HCV structural proteins. METHODS: Peptides were synthesized and tested in HLA-A2 binding assays. Binder peptides were used to stimulate peripheral blood mononuclear cells from HCV+ patients and controls, and activity measured in chromium release and ELISPOT assays. RESULTS: Twenty binder peptides were found, and stimulation of HCV+ patient cells with nine peptides showing high binding ability led to the growth of CD8+ CTL recognizing peptide E2(614-622) in association with HLA-A2. Peptide E2(614-622) was recognized by 30% of HLA-A2+ patients with chronic HCV infection, but no responses were observed in control groups. Five peptides derived from region E2(614-622) from 26 different viral isolates bound to HLA-A2 molecules, and all of them but one, containing Phe at position 622, were recognized by E2(614-622) specific CTL. CONCLUSIONS: These results show that peptide E2(614 622) belongs to a highly conserved region of HCV E2, and might be a good candidate to induce anti-HCV CTL responses in HLA-A2+ subjects. PMID- 11281564 TI - Apoptosis in alcoholic hepatitis: a novel therapeutic target? PMID- 11281565 TI - Mitochondria: important target for drug toxicity? PMID- 11281566 TI - Post-liver transplantation osteoporosis. PMID- 11281568 TI - Quality of reporting of meta-analyses: the QUOROM statement. Will it help? PMID- 11281567 TI - Cholesterol and cholestasis: a lesson from the Mdr2 (-/-) mouse. PMID- 11281570 TI - Images in hepatology. Postoperative biliary fistula treated with percutaneous microcoil embolization. PMID- 11281569 TI - Hepatitis after intravenous buprenorphine misuse in heroin addicts. AB - BACKGROUND: Sublingual buprenorphine is used as a substitution drug in heroin addicts. Although buprenorphine inhibits mitochondrial function at high concentrations in experimental animals, these effects should not occur after therapeutic sublingual doses, which give very low plasma concentrations. CASE REPORTS: We report four cases of former heroin addicts infected with hepatitis C virus and placed on substitution therapy with buprenorphine. These patients exhibited a marked increase in serum alanine amino transferase (30-, 37-, 13- and 50-times the upper limit of normal, respectively) after injecting buprenorphine intravenously and three of them also became jaundiced. Interruption of buprenorphine injections was associated with prompt recovery, even though two of these patients continued buprenorphine by the sublingual route. A fifth patient carrying the hepatitis C and human immunodeficiency viruses, developed jaundice and asterixis with panlobular liver necrosis and microvesicular steatosis after using sublingual buprenorphine and small doses of paracetamol and aspirin. CONCLUSIONS: Although buprenorphine hepatitis is most uncommon even after intravenous misuse, addicts placed on buprenorphine substitution should be repeatedly warned not to use it intravenously. Higher drug concentrations could trigger hepatitis in a few intravenous users, possibly those whose mitochondrial function is already impaired by viral infections and other factors. PMID- 11281571 TI - Ursodeooxycholic acid for primary biliary cirrhosis. PMID- 11281572 TI - Duration of immunosuppressive therapy in autoimmune hepatitis. PMID- 11281573 TI - Intratumoral production of interleukin-5 leading to paraneoplastic peripheral eosinophilia in hepatocellular carcinoma. PMID- 11281574 TI - Stiffness optimisation of cement and stem materials in total hip replacement. AB - It is acknowledged that bone resorption and fatigue fracture of cement in total hip replacement may cause feature problems. The solution is frequently sought associated with the stiffness of cement and stem. The purpose of this paper is firstly to describe the effect of changes in modulus of elasticity of the cement material for the implanted prosthesis on the fatigue notch factor (Kf). The paper further describes a method of numerical optimisation to determine the optimal stiffness characteristics of cement and stem materials, which minimises the probability of fatigue fracture of cement at all interfaces with the stem and the bone, while limiting the amount of bone resorbed. The parameters describing the elastic moduli of cement and stem were considered as design variables. The method was applied to an equivalent 2D finite element model of femoral hip replacement in combination with an optimisation procedure using the ANSYS program. The results of the first study suggest that lower modulus of elasticity of cement material decreases Kf in the cement at all interfaces and proximal bone while higher values increase Kf. For the second aim, Young's moduli of about 0.6 and 22 GPa are optimal for cement and stem materials, respectively. These characteristics decreased the probability of fatigue fracture of cement at all interfaces with the stem and the bone as a result of decreasing Kf in cement at all interfaces, while limiting the amount of bone resorbed as a result of increasing Kf in the proximal bone. PMID- 11281575 TI - Effects of ceramic component on cephalexin release from bioactive bone cement consisting of Bis-GMA/TEGDMA resin and bioactive glass ceramics. AB - The purpose of this study was to elucidate the effect of amount of ceramic cement powder on drug release from bioactive bone cement. The associated bone-bonding strength was also investigated. The bioactive bone cement under investigation consisted of bisphenol-alpha-glycidyl methacrylate (Bis-GMA), triethylene-glycol dimethacrylate (TEGDMA) resin and a combination of apatite- and wollastonite containing glass-ceramic (A-W GC) powder. A-W GC powder (50%, 70% and 80% w/w) containing 5% cephalexin (CEX) powder hardened within 5 min after mixing with Bis GMA/TEGDMA resin. The compressive strength of the cement with or without drug increased with increasing the amount of ceramic powder. The compressive strength of the 80% ceramic cement without the incorporation of cephalexin was 194 MPa. This compressive strength was about 3 times higher than that for polymethylmethacrylate cement. After the cement was implanted in the proximal metaphysis of the tibiae of male rabbits, the failure load for the cement was found to increase with increasing of the amount of ceramic powder. This finding suggested that the cement formed a bonding with bone. In vitro CEX release from bioactive bone cement pellets in a simulated body fluid at pH 7.25 and 37 degrees C continued for more than 2 weeks. Drug release profile followed the Higuchi equation initially, but not at later stages. The drug release rate increased with increasing amount of ceramic powder in the mixture. Since the pore volume of the cement increased with increasing of amount of ceramic powder, the drug diffused in the pores between the ceramics particle and polymer matrix. As hydroxyapatite precipitated on the cement surface, the drug release rate decreased, as observed at the later release stage. These results suggest that varying the amount of ceramic powder in the cement system could control the drug release rate from bioactive bone cement. PMID- 11281576 TI - Comparative wear and wear debris under three different counterface conditions of crosslinked and non-crosslinked ultra high molecular weight polyethylene. AB - The wear debris generated from ultra high molecular weight polyethylene (UHMWPE) have been recognised as one of the major causes of failure in total hip replacements (THR). It is essential to reduce the wear debris generated from UHMWPE acetabular cups in order to minimise this problem. Debris in the submicron size range is believed to have greater osteolytic potential. It is now known that crosslinked UHMWPE acetabular cups have reduced volumetric wear rates but little is known about the influence of crosslinking on the size and morphology of the wear debris. In this study, the wear of grade GUR 1020 crosslinked (vacuum gamma irradiated), GUR 1120 crosslinked (acetylene enhanced irradiated) and non cross linked (ethylene oxide sterilised) GUR 1020 UHMWPE was compared in multidirectional pin-on-plate wear tests under three different counterface conditions (smooth, isotropically rough and scratched counterfaces). Multidirectional motion was chosen because this motion was closer to the relative motion in the natural hip. From this study, better wear resistance of crosslinked UHMWPE compared with non-crosslinked UHMWPE was demonstrated for the smooth counterface conditions. However, in the rough and scratched counterface conditions, the vacuum gamma irradiated crosslinked material produced significantly higher wear rates than the non-crosslinked material. The analysis of the wear debris showed that the majority of the volume of the acetylene enhanced crosslinked UHMWPE wear debris was in the most biologically active size range (0.1 to 0.5 microm). In contrast, the non-crosslinked material and the vacuum gamma irradiated crosslinked material had a greater proportion of the volume of the debris in the larger size ranges which are less biologically active. This has important implications for its osteolytic potential. PMID- 11281577 TI - A depth profile of oxidation and gel fraction in gamma-irradiated silane crosslinked and ultra high molecular weight polyethylenes. AB - The depth profile of oxidation index and gel fraction has been measured for two silane crosslinked poly(ethylene) (SXLPE) acetabular cups (one gamma irradiated in air, and one non-irradiated, both with a shelf-life of 13 years) and for two UHMWPE components (one gamma irradiated in air and one non-irradiated, with shelf lives of 13 and 7 years, respectively). Only the irradiated UHMWPE exhibited any variation in these properties with depth. The oxidation profile (maximum 1 mm below surface) has been explained to result from reduced levels of diffused oxygen with depth, giving rise to a balance of alkyl and peroxyl radicals (and hence maximum carbonyl production) just below the surface. The gel fraction profile (maximum 4 mm below surface) is also attributed to the lower levels of diffused oxygen with depth, causing crosslinking to dominate in the bulk and chain scission to dominate at the surface. The resistance to oxidative degradation in the non-irradiated SXLPE was attributed to the use of antioxidants in the polymer processing. PMID- 11281579 TI - Effect of oligofucose derivatives on acetic acid-induced gastric ulcer in rats. AB - This study attempted to enhance the anti-ulcer activity of fucoidan from Cladosiphon okamuranus TOKIDA by chemical modification with a hydrophobic group. The suitable number of fucose residues in the effective compound was also clarified to obtain a compound of constant quality. Degraded fucoidans were coupled with several hydrophobic groups via Schiff bases, and their anti-ulcer activities were determined by acetic acid-induced ulcer models in rats. Size fractionated oligofucose was also modified and assayed for anti-ulcer activity. Among the modified oligofucoses, only the oligofucose-dodecylaniline combination (OFDA) significantly promoted ulcer healing. The effective dose was 0.2 mg/kg/d. The most suitable number of fucose residues in the compound for the anti-ulcer activity was determined to be around 12. We succeeded in enhancing the anti-ulcer activity of Cladosiphon fucoidan by modification with dodecylaniline. The activity of this compound was comparable or greater than that of typical anti ulcer agents. By determination of the optimal OF chain length for the anti-ulcer activity of OFDA, it became possible to obtain OFDA of constant quality. PMID- 11281578 TI - Effect of calcium on the surface properties of phospholipid monolayers with respect to surfactant formulations in respiratory distress syndrome. AB - The effects of calcium, in the form of calcium chloride, at concentrations of 5 and 20 mM, were studied on the surface properties of physiologic relevance to specialised biomaterials which replace lung surfactant in Respiratory Distress Syndrome. The dynamic surface pressure, re-spreading ratio, compressibility, hysteresis area and recruitment index of pure films of the main phospholipids of pulmonary surfactant namely dipalmitoyl phosphatidylcholine, phosphatidylglycerol, phosphatidylethanolamine and of binary mixed films of these phospholipids in the ratio of 2:3 were studied both in the presence and absence of calcium by in vitro analysis using a Wilhelmy balance. Surface excess films, of each of the surfactant systems, with initial concentration 15 A2 molecule were compressed at the rate of 50 seconds/cycle past collapse till a compression ratio of 4:1. The presence of 5 mM calcium caused a significant decrease in compressibility (p < 0.05 Mann-Whitney U test) of all the surfactant monolayer films. No further benefit was obtained by adding 20 mM calcium over that of 5 mM calcium. A significant beneficial effect of calcium (p < 0.05 Mann-Whitney U test) on film stability was observed when analysing the materials in a pulsating bubble surfactometer, in which liposomal suspensions of 1% concentration in the presence of 5 mM calcium were pulsated at a high frequency of 40 cycles per minute, corresponding to the respiratory frequency of neonates. The ultrastructure of the liposomal suspensions were also studied using cryogenic scanning electron microscopy and longitudinal micro-tubular structures were found on addition of 5 mM calcium, which could have resulted in the improved performance of the exogenous surfactants with respect to compressibility and stability. PMID- 11281580 TI - Push-out strength of hydroxyapatite coated by sputtering technique in bone. AB - Hydroxyapatite was coated with 1 microm thickness on titanium columns of a length of 10 mm, an outer diameter of 4.0 mm by radio frequency magnetron sputtering. The hydroxyapatite coating titanium columns were implanted in the diaphysis of femora of 3 adult dogs, and push-out test was carried out after 2, 4 and 12 weeks of implantation using a testing machine. The interface of bone/column was observed histologically after the test. At 12 weeks the push-out strengths of coating and non-coating columns were 3.5 and 1 MPa, respectively. Histological observation indicated a formation of thin connective tissue with 5-30 microm thickness at the interface of the bone/column. No inflammation was observed during the implantation periods. PMID- 11281581 TI - Surgical correction of blepharoptosis in patients with myasthenia gravis. AB - PURPOSE: To describe the results of surgical correction of blepharoptosis in a series of patients with myasthenia gravis (MG). METHODS: In this retrospective case series, we reviewed the medical records of all patients with MG who did not respond to medical therapy and underwent surgical correction for blepharoptosis at the Mayo Clinic between 1985 and 1999. The primary outcome measure was change in interpalpebral eyelid fissure height. RESULTS: Sixteen blepharoptosis procedures were performed on 10 patients with MG. Eight of the 10 patients had ocular MG. Two of the 10 patients had systemic MG. Of the 16 procedures performed, 9 were external levator advancements (ELA), six were frontalis slings, and one was a tarsomyectomy. Patients were followed postoperatively for an average of 34 months (range, 14-126 months). The amount of ptosis was quantified pre- and postoperatively for seven of the nine eyelids that underwent ELA. For these seven eyelids (five patients), there was a statistically significant improvement in the mean interpalpebral eyelid fissure height from 3.7 mm preoperatively to 7.8 mm postoperatively, with a mean difference of 4.1 mm (95% confidence interval 1.9 mm to 6.25 mm, p = 0.0038). Postoperative complications included worsened diplopia in one patient with ELA and exposure keratopathy in one patient with frontalis sling. Two of the ELA eyelids developed recurrent ptosis requiring additional surgery more than 2 years after the initial procedure. CONCLUSIONS: Blepharoptosis surgery can achieve eyelid elevation in patients who have failed to respond to medical therapy for MG. Potential complications include worsened diplopia and exposure keratopathy. PMID- 11281582 TI - Dacryocystosclerotherapy. AB - PURPOSE: To introduce a less invasive therapeutic method in selected patients with primary nasolacrimal duct obstruction. METHODS: Noncomparative interventional case series. Seven patients with primary acquired nasolacrimal duct obstruction who were poor candidates for dacryocystorhinostomy because of underlying medical conditions and two symptomatic patients with previous incomplete dacryocystectomy were included. Ethanolamine oleate 5% was slowly injected through the puncta into the patent portion of the lacrimal drainage apparatus. RESULTS: After a mean follow-up period of 26 months, the lacrimal pathway was permanently obliterated in 6 cases (66.7%). In two cases, the procedure was repeated successfully and in one case, the patient refused to repeat the procedure after the initial failure. CONCLUSIONS: In selected patients, ethanolamine oleate dacryocystosclerotherapy appears to offer a simple, low-cost, less invasive alternative to dacryocystectomy. PMID- 11281583 TI - Intraoperative mitomycin C in dacryocystorhinostomy. AB - PURPOSE: To assess the efficacy of intraoperative mitomycin C (MMC) in external dacryocystorhinostomy (EXT-DCR). METHODS: Forty-six cases (50 lacrimal drainage systems [LDS]) with nasolacrimal duct obstruction were randomized into three groups. In the control group, a standard EXT-DCR procedure was performed. In the two MMC groups, a piece of cotton soaked with 0.2 mg/ml MMC (group 1) or 0.5 mg/ml MMC (group 2) was applied to the nasal mucosa and the mucosa of the lacrimal sac in the osteotomy site for 5 minutes. RESULTS: The dacryocystorhinostomy in all patients was patent by irrigation 2 to 3 weeks postoperatively. After a mean follow-up interval of 35.2 +/- 5.3 months, the dacryocystorhinostomy was patent in 15 (83%) of 18 LDS in the control group, 16 (100%) of 16 LDS in group 1, and 15 (94%) of 16 LDS in group 2. The mean ostium sizes were 22.2 +/- 5.0 mm2 in group 1, 20.6 +/- 4.5 mm2 in group 2, and 13.2 +/- 2.7 mm2 in group 3 at the final follow-up visit; the difference between the patients treated with MMC and the control group was statistically significant. There was no statistically significant difference between the two MMC groups, however. No surgical complications occurred. CONCLUSIONS: Intraoperative MMC in DCR is a safe and effective adjuvant that helps achieve favorable long-term success rates. PMID- 11281584 TI - Supernumerary orbital muscle in congenital eyelid retraction. AB - PURPOSE: Although several reports of supernumerary orbital muscles related to the levator palpebrae superioris have been published, no case has been associated with congenital eyelid retraction. This report describes an apparent causal relationship between an accessory levator muscle slip and congenital eyelid retraction. METHODS: Case report and literature review. RESULTS: Release of the anomalous muscle's attachment from the superior tarsal border alone resulted in resolution of the eyelid retraction. CONCLUSIONS: Eyelid muscle anomalies may be a cause of congenital eyelid retraction. Ophthalmologists who treat eyelid disorders should be aware of this possibility when evaluating and operating on patients with congenital eyelid retraction. PMID- 11281585 TI - Destructive eyelid lesions in sarcoidosis. AB - PURPOSE: To report the clinical and histopathologic findings of a patient with sarcoidosis causing bilateral destruction of the lower eyelids. METHODS: Case report. RESULTS: Bilateral destructive lower eyelid lesions and cicatricial entropion developed in a 43-year-old man with systemic sarcoidosis. Histopathology was consistent with sarcoid granulomas. Disease progression was arrested with systemic prednisone and methotrexate before eyelid reconstruction was performed. CONCLUSIONS: Sarcoidosis very rarely can cause destruction of full thickness eyelid architecture. Active inflammation should be controlled before reconstruction. PMID- 11281586 TI - Delayed retrobulbar hematoma after blepharoplasty. AB - PURPOSE: To describe a case of late orbital hematoma after blepharoplasty and review the literature on this subject. METHODS: A healthy woman developed an orbital hematoma and loss of vision 7 days after an elective blepharoplasty. Computed tomography and magnetic resonance images of the orbits were obtained 1 day after the vascular accident. RESULTS: Computed tomography and magnetic resonance images clearly demonstrated that the site of the hemorrhage was the superior nasal fat pad. Blood from this region molded the globe and accumulated in the intraconal space behind the posterior sclera. The patient was successfully treated with conservative measures only. CONCLUSIONS: Orbital hematoma, which is the main cause of loss of vision after blepharoplasty, can be a late postoperative complication. PMID- 11281587 TI - Lacrimal gland ductal cyst abscess. AB - PURPOSE: To describe a case of lacrimal gland ductal cyst complicated by secondary infection. METHODS: Case report. RESULTS: A 51-year-old woman presented acutely with an enlarging, painful mass in the superotemporal fornix. Clinical examination, echography, and surgical evaluation revealed a lacrimal gland ductal cyst with abscess formation. The lacrimal gland cyst was treated with oral antibiotics in combination with incision, drainage, and marsupialization. CONCLUSIONS: Lacrimal gland ductal cysts are rare but must be considered in the differential diagnosis of lacrimal gland and upper eyelid mass lesions. Typically, lacrimal gland ductal cysts develop after chronic inflammation, infection, or trauma. We describe a patient who presented acutely with a lacrimal gland ductal cyst associated with a rare complication of abscess formation. PMID- 11281588 TI - Rhinoorbitocerebral actinomycosis. AB - PURPOSE: To report a case of actinomycotic orbital abscess with subdural empyema and pansinusitis, an unusual presentation of a rarely seen infection. METHODS: Case report. RESULTS: A 35-year-old man sought treatment for signs and symptoms of an orbital abscess 22 days after a dental extraction. Computed tomography demonstrated a left orbital abscess with left pansinusitis and a large subdural empyema. Surgical clearance of all purulent material was done followed by prolonged penicillin therapy. Culture of pus from all sources yielded Actinomycosis israelii. At the time of discharge and 1-month follow-up, the patient had 20/20 vision with no neurologic deficits. CONCLUSIONS: In orbital infections with atypical presentations, unusual pathogens should be considered as the causative agents. PMID- 11281589 TI - Recurrent orbital inflammation from metastatic orbital carcinoid tumor. AB - PURPOSE: To report an unusual clinical presentation for carcinoid tumor metastatic to the orbit. METHODS: Two adult patients with metastatic carcinoid tumor and unilateral orbital masses are described. RESULTS: Both patients sought treatment for acute unilateral orbital inflammation. Neuroimaging revealed orbital metastases adjacent to the inflamed adnexal tissue. Based on each patient's description of similar, prior, untreated episodes, minimal medical management was initiated. Resolution of the inflammatory signs was spontaneous in one case and assisted by pulsed, systemic corticosteroids in the second case. Compressive neuropathic vision loss occurred 11 months later in the second case. CONCLUSIONS: Carcinoid tumor metastatic to the orbit may manifest as recurrent orbital and ocular adnexal inflammation. These signs differ from systemic carcinoid syndrome in that they are unilateral, limited to only the orbital and ocular adnexal soft tissues, and resolve over days. Clinicians must carefully differentiate this manifestation from that of tumor necrosis, adnexal infection, or orbital outlet obstruction. This presentation may result from the spontaneous release of local inflammatory mediators intrinsic to the orbital tumor. PMID- 11281590 TI - Unsuspected recurrent pituitary adenoma presenting as an orbital mass. AB - PURPOSE: Orbital invasion of pituitary tumors is rare and usually accompanied by optic nerve head pallor and visual loss. We describe a case of unilateral massive orbital invasion by a recurrent pituitary tumor with preserved visual acuity and normal optic nerve appearance. METHODS: Case report. RESULTS: Progressive proptosis developed 15 years after transphenoidal removal of a pituitary tumor. Based on the radiological appearance and the clinical history, the patient was suspected to have a sphenoid wing meningioma secondary to previous radiation treatment. A combined neurosurgical and orbital approach was used to remove the intraorbital mass, which extended from the cranial cavity through the superior orbital fissure and the optic canal. Histopathologic examination demonstrated a recurrent nonsecreting pituitary adenoma. CONCLUSIONS: Orbital extension of a recurrent pituitary adenoma should be considered in the differential diagnosis of progressive proptosis even in the absence of significant optic neuropathy. PMID- 11281591 TI - Orbital nodular fasciitis simulating a dermoid cyst in an 8-month-old child. Case report and review of the literature. AB - PURPOSE: To report a clinicopathologic correlation of orbital nodular fasciitis that simulated a dermoid cyst in an infant and to review the literature on orbital nodular fasciitis. METHOD: Case report; literature review. RESULTS: A progressively enlarging subcutaneous mass superotemporal to the right eye that clinically resembled a dermoid cyst developed in an 8-month-old boy. Magnetic resonance imaging disclosed an enhancing, circumscribed, solid, soft-tissue mass in the anterior part of the orbit superotemporally. The tumor was removed intact by a superolateral orbitotomy. Histopathologic and immunohistochemical studies revealed findings consistent with nodular fasciitis, a pseudoneoplastic lesion marked by a proliferation of myofibroblasts. CONCLUSIONS: Although nodular fasciitis in the ocular region usually occurs in adults and older children, it should also be considered in the differential diagnosis of orbital or adnexal masses in infants. PMID- 11281592 TI - Orbital involvement in allergic fungal sinusitis. PMID- 11281593 TI - Digital photography for the ophthalmic plastic surgeon. PMID- 11281594 TI - More information on Alloderm. PMID- 11281595 TI - Five things oculoplastic surgeons should know about business administration. PMID- 11281597 TI - Glabellar rhytids in thyroid-associated orbitopathy. AB - PURPOSE: To correlate the presence and severity of glabellar rhytids with disease severity in patients with thyroid-associated orbitopathy (TAO). METHODS: Retrospective, noncomparative medical record review of 113 consecutive new patient visits with the clinical diagnosis of TAO. Ocular examination features studied included assessment of visual acuity, optic nerve appearance and function, proptosis, strabismus, eyelid position, and exposure keratopathy. These features were correlated with graded clinical photographs evaluating rhytid severity using the Fisher exact text. RESULTS: Eighty of 97 patients included in the study (82.5%) had glabellar rhytids. Eyelid retraction was present in 95 of the 97 patients (98%). Patients with diplopia had more severe rhytids (p < 0.05). Patients with at least three abnormal clinical findings tended to have more severe rhytids. CONCLUSIONS: Glabellar rhytids are a common finding in patients with TAO, and tend to be present in patients with diplopia and multiple stigmata of the disease. PMID- 11281598 TI - The maximal Hughes procedure. AB - PURPOSE: To describe the use of oblique medial and lateral periosteal flaps with the Hughes tarsoconjunctival flap for the repair of maximal defects of the lower eyelid. METHODS: A small prospective case series of eight patients requiring lower eyelid reconstruction following with maximal defect of the lower eyelid. The patients underwent a Hughes tarsoconjunctival advancement combined with oblique medial and lateral periosteal flaps, and were assessed for aesthetic outcome and surgical complications. RESULTS: All patients had uncomplicated surgery. Outcomes assessed included corneal protection, eye closure, lower eyelid retraction, complications, and patient satisfaction. Eyelid contour and protection was excellent in all patients. Postoperatively, one patient had mild lower eyelid retraction, and in a second patient, medial ectropion with mild lower eyelid retraction developed that required subsequent revision. CONCLUSIONS: The maximal Hughes procedure is a safe and effective procedure that may be performed with patients under local anesthesia and may avoid the need for more extensive techniques for surgical repair of maximal defects of the lower eyelid. PMID- 11281596 TI - Systemic corticosteroid use in orbital lymphangioma. AB - PURPOSE: To describe the clinical results of systemic corticosteroid use in a series of patients with orbital lymphangioma. METHODS: Four patients (two adults and two children) were treated with corticosteroids using intravenous, oral, or both routes for 2 days to a month. Corticosteroids were used with and without other therapies for symptomatic exacerbations. RESULTS: The adults showed more improvement with pain than with swelling, whereas the children had improvement with both the signs and symptoms. There were no complications in any patient. CONCLUSIONS: Systemic corticosteroids are a useful therapeutic option for patients with orbital lymphangioma and can be used as an adjuvant treatment to surgery and other modalities. Resolution of symptoms with corticosteroids was expedited compared with the natural history of the disease in the patients studied. PMID- 11281599 TI - Electron capture dissociation of gaseous multiply charged ions by Fourier transform ion cyclotron resonance. AB - Fourier-transform ion cyclotron resonance instrumentation is uniquely applicable to an unusual new ion chemistry, electron capture dissociation (ECD). This causes nonergodic dissociation of far larger molecules (42 kDa) than previously observed (<1 kDa), with the resulting unimolecular ion chemistry also unique because it involves radical site reactions for similarly larger ions. ECD is highly complementary to the well known energetic methods for multiply charged ion dissociation, providing much more extensive protein sequence information, including the direct identification of N- versus C-terminal fragment ions. Because ECD only excites the molecule near the cleavage site, accompanying rearrangements are minimized. Counterintuitively, cleavage of backbone covalent bonds of protein ions is favored over that of noncovalent bonds; larger (>10 kDa) ions give far more extensive ECD if they are first thermally activated. This high specificity for covalent bond cleavage also makes ECD promising for studying the secondary and tertiary structure of gaseous protein ions caused by noncovalent bonding. PMID- 11281601 TI - A Fourier-transform ion cyclotron resonance study of the 3,5-didehydrophenyl cation. AB - The 3,5-didehydrophenyl cation has been generated in good purity via sustained off-resonance irradiation for collision-activated dissociation of 3,5 dinitrobenzoyl chloride in a Fourier-transform ion cyclotron resonance mass spectrometer. Differences in the ion-molecule reactivity of this species from that of its cyclic and acyclic isomers allowed isomeric distinction to be achieved. This study represents the first definitive identification of this fundamentally interesting, doubly aromatic ion. However, the formation of the 3,5 didehydrophenyl cation was found to be the exception rather than the rule, with most 1,3,5-substituted benzenes yielding mainly acyclic C6H3+ isomers under electron ionization conditions. This mixed ion population was attributed to isomerization of fragmentation intermediates rather than any intrinsic instability of the 3,5-didehydrophenyl cation. PMID- 11281600 TI - Gas-phase reactions of hydrated alkaline earth metal ions, M2+ (H2O)n (M = Mg, Ca, Sr, Ba and n = 4-7), with benzene. AB - Gas-phase reactions of hydrated divalent alkaline earth metal ions and benzene were investigated by electrospray ionization Fourier-transform mass spectrometry. Rate constants for solvent-exchange reactions were determined as a function of hydration extent for Mg2+, Ca2+, Sr2+, and Ba2+ clusters containing four to seven water molecules each. All of the strontium and barium clusters react quickly with benzene. Barium reacts slightly faster than the corresponding strontium cluster with the same number of water molecules attached. For calcium, clusters with four and five water molecules react quickly, whereas those with six and seven water molecules do not. Magnesium with four water molecules reacts quickly, but not when five through seven water molecules are attached. The slow reactivity observed for some of these clusters indicates that the cation-pi interaction between the metal ion and benzene is partially screened by the surrounding water molecules. The reactivity of magnesium with seven water molecules is intermediate that of the hexa- and pentahydrate and the tetrahydrate. This result is consistent with the seventh water molecule being in the outer shell and much more weakly bound. The unusual trend in reactivity observed for magnesium may be due to the presence of mixed shell structures observed previously. These results are the first to provide information about the relative importance of cation-pi interactions in divalent metal ions as a function of metal hydration extent. Such studies should also provide a model and some insight into the relative binding affinities of divalent metal ions to aromatic residues on peptides and proteins. PMID- 11281602 TI - Gas-phase hydrogen/deuterium exchange of positively charged mononucleotides by use of Fourier-transform ion cyclotron resonance mass spectrometry. AB - The gas-phase structures of protonated (deoxy)nucleoside-5'- and 3' monophosphates (mononucleotides) have been examined by the use of gas-phase hydrogen/deuterium (H/D) exchange and high-field Fourier-transform ion cyclotron resonance mass spectrometry. These nucleotides were reacted with three different deuterating reagents: ND3, D2O, and D2S, of which ND3 was the most effective. All mononucleotides fully exchanged their labile hydrogen for deuterium with ND3 with the exception of deoxycytidine-3'-monophosphate, deoxyadenosine-5'-monophosphate, adenosine-5'-monophosphate, and adenosine-3'-monophosphate. Semiempirical calculations demonstrate the presence of hydrogen bonding upon protonation of the purine mononucleotides which may lead to incomplete H/D exchange. H/D exchange rates differed between the deoxymononucleotides and the ribomononucleotides, suggesting that the 2'-OH group plays an important role in the exchange process. Reactions of nucleosides and mononucleotides with D2O demonstrate that a structure-specific long-lived ion-molecule complex between D2O and the mononucleotide involving the phosphate group is necessary for exchange to overcome the high-energy activation barrier. In contrast, a structure-specific long-lived ion-molecule complex between the mononucleotides and ND3 is not required for exchange to occur. PMID- 11281603 TI - Chiral recognition in gas-phase cyclodextrin: amino acid complexes--is the three point interaction still valid in the gas phase? AB - The validity of the "three-point interaction" model is examined in the guest exchange reaction involving complexes of cyclodextrins and amino acids. The amino acid guest is exchanged in the gas phase in the presence of a gaseous alkyl amine. The net reaction is proton transfer between the protonated amino acid and the alkyl amine. The amino acid is lost as a neutral species. This reaction is sensitive to the chirality of the amino acid. Several amino acids are examined as well as the respective methyl esters to determine the role of the three interacting groups (ammonium, carboxylic acid, and side chain) in enantioselectivity. We find that the three-point interaction model is indeed valid in the gas phase. Enantioselectivity is optimal when two points of attraction and one repulsion is present in the gas-phase complex. The results are supported by molecular modeling calculations. A mechanism for the exchange is proposed. PMID- 11281605 TI - A gated trapping strategy with a two-time constant and a delay for catching in field generated ions that range over three decades in mass-to-charge and two decades in velocity in Fourier-transform mass spectrometry. AB - In-field, matrix-assisted laser desorption/ionization (MALDI) may provide a means to keep part of the original promise of Fourier-transform mass spectrometry (FTMS) to give high performance and versatile mass spectrometry from a mechanically simple instrument. Gated trapping has been employed as a means of catching MALDI-produced ions in the FTMS trap. This approach is important for both in-field and externally produced ions. Even with improvements, gated trapping has not yet been able to catch ions over wide ranges of mass-to-charge and velocity. A design of a "two-time constant with a delay" gated trapping strategy using "idealized" potentials in a normalized system is given as an example to establish that in principle gated trapping strategies can capture ions that range over three decades of m/z and two decades in velocity. A procedure for calculating a physical system from the normalized system is given. The design is tolerant of variations in the physical parameters used to define the physical system from the normalized system. PMID- 11281604 TI - Gas-phase cleavage of PTC-derivatized electrosprayed tryptic peptides in an FT ICR trapped-ion cell: mass-based protein identification without liquid chromatographic separation. AB - Condensed phase protein sequencing typically relies on N-terminal labeling with phenylisothiocyanate ("Edman" reagent), followed by cleavage of the N-terminal amino acid. Similar Edman degradation has been observed in the gas phase by collision-activated dissociation of the N-terminal phenyl thiocarbamoyl protonated peptide [1] to yield complementary b1 and y(n-1) fragments, identifying the N-terminal amino acid. By use of infrared multiphoton (rather than collisional) activation, and Fourier transform ion cyclotron resonance (rather than quadrupole) mass analysis, we extend the method to direct analysis of a mixture of tryptic peptides. We validate the approach with bradykinin as a test peptide, and go on to analyze a mixture of 25 peptides produced by tryptic digestion of apomyoglobin. A b1+ ion is observed for three of the Edman derivatized peptides, thereby identifying their N-terminal amino-acids. Search of the SWISS-PROT database gave a single hit (myoglobin, from the correct biological species), based on accurate-mass FT-ICR MS for as few as one Edman-derivatized tryptic peptide. The method is robust-it succeeds even with partial tryptic digestion, partial Edman derivatization, and partial MS/MS IRMPD cleavage. Improved efficiency and automation should be straightforward. PMID- 11281606 TI - Qualitative comparison between the quantum calculations and electrospray mass spectra of complexes of polyammonium macrotricyclic ligands with dicarboxylic acids. AB - The host-guest interactions play a very important role in chemical and biological processes. It is therefore important to be able to characterize these complexes. Electrospray mass spectrometry can be used to characterize the complex formation. It provides information on the mass and the charge of these ionic complexes. In this article, we show that the use of ab initio and semiempirical calculations, in addition to the results obtained by electrospray mass spectrometry, reveal to be a promising tool for the study of these noncovalent complexes. In this article, host-guest complexes formed by macropolycyclic polyammonium host molecules and dicarboxylic acids are studied. PMID- 11281607 TI - The methanol-induced conformational transitions of beta-lactoglobulin, cytochrome c, and ubiquitin at low pH: a study by electrospray ionization mass spectrometry. AB - The methanol-induced conformational transitions under acidic conditions for beta lactoglobulin, cytochrome c, and ubiquitin, representing three different classes of proteins with beta-sheets, alpha-helices, and both alpha-helices and beta sheets, respectively, are studied under equilibrium conditions by electrospray ionization mass spectrometry (ESI-MS). The folding states of proteins in solution are monitored by the charge state distributions that they produce during ESI and by hydrogen/deuterium (H/D) exchange followed by ESI-MS. The changes in charge state distributions are correlated with earlier studies by optical and other methods which have shown that, in methanol, these proteins form partially unfolded intermediates with induced alpha-helix structure. Intermediate states formed at about 35% methanol concentration are found to give bimodal charge state distributions. The same rate of H/D exchange is shown by the two contributions to the bimodal distributions. This suggests the intermediates are highly flexible and may consist of a mixture of two or more rapidly interconverting conformers. H/D exchange of proteins followed by ESI-MS shows that helical denatured states, populated at around 50% methanol concentration, transform into more protected structures with further increases in methanol concentration, consistent with previous circular dicroism studies. These more protected structures still produce high charge states in ESI, similar to those of the fully denatured proteins. PMID- 11281608 TI - Metastable ion formation and disparate charge separation in the gas-phase dissection of protein assemblies studied by orthogonal time-of-flight mass spectrometry. AB - The dissection of specific and nonspecific protein complexes in the gas phase is studied by collisionally activated decomposition. In particular, the gas phase dissection of multiple protonated homodimeric Human Galectin I, E. Coli Glyoxalase I, horse heart cytochrome c, and Hen egg Lysozyme have been investigated. Both the Human Galectin I and E. Coli Glyoxalase I enzymes are biologically active as a dimer, exhibiting molecular weights of approximately 30 kDa. Cytochrome c and Lysozyme are monomers, but may aggregate to some extent at high protein concentrations. The gas phase dissociation of these multiple protonated dimer assemblies does lead to the formation of monomers. The charge distribution over the two concomitant monomers following the dissociation of these multiple protonated dimers is found to be highly dissimilar. There is no evident correlation between the solution phase stability of the dimeric proteins and their gas-phase dissociation pattern. Additionally, in the collisionally activated decomposition spectra diffuse ion signals are observed, which are attributed to monomer ions formed via slow decay of the collisionally activated dimer ions inside the reflectron time-of-flight. Although, the formation of these diffuse metastable ions may complicate the interpretation of collisionally activated decomposition mass spectra, especially when studying noncovalent protein complexes, a simple mathematical equation may be used to reveal their origin and pathway of formation. PMID- 11281609 TI - Gln-Gly cleavage: correlation between collision-induced dissociation and biological degradation. AB - Tryptic digestion of the 150-residue human acidic salivary proline-rich protein 1 (PRP-1) generated eight peptides, two of which corresponded to the N-terminal 30 residue segment. In each of the other six tryptic peptides, a consensus repeat with the structure PQGPPQQGG was present. A facile Gln-Gly cleavage between the second and the third residues of the repeat was observed during collision-induced dissociation experiments. We postulate possible mechanisms to account for this reactivity, involving attack on the peptidyl carbonyl group by the Gln sidechain. Significantly, the Gln-Gly cleavage has been shown to be biologically important in the bacterial degradation of PRPs in saliva, generating bacteria-binding Pro Gln C-termini. We suggest a link between the gas-phase chemistry and the biochemical degradation of these molecules. PMID- 11281610 TI - Physical/chemical separations in the break-up of highly charged droplets from electrosprays. AB - Highly-charged droplets, as formed by an electrospray process, are known to undergo asymmetric fission to form smaller droplets. We have observed a chemical and physical separation phenomenon that occurs in the droplet break-up process and is related to a compound's surface activity in solution. Two experimental approaches demonstrated that the smaller satellite droplets and the progeny droplets generated by the spray formation and asymmetric fission processes to be surfactant-enriched. These smaller droplets were also effectively separated from the larger primary and residual droplets because of their smaller inertia and high surface charge density, and a region attributed to the initially formed smaller satellite droplets was found to be strikingly confined in a narrow periphery region of the electrospray. The phenomenon may have utility for chemical separations and have significant implications for the sensitivity and selectivity of electrospray ionization-mass spectrometry. PMID- 11281611 TI - Acute effect of physical exercise on serum insulin-like growth factor-binding protein 2 and 3 in healthy men: role of exercise-linked growth hormone secretion. AB - The purpose of this study was to delineate the role of GH on serum IGF-I, IGFBP-2 and -3 responses to exercise. Hormones were evaluated in six trained male subjects before (-30, -15, 0), during (+15) and after (+30, +45, + 60, +90 min) a thirty-minutes treadmill exercise (60% VO2max), both after a single administration of a somatostatin analog (i.e., octreotide, 0.1 mg sc) and after saline. The same evaluations were performed without exercise with similar treatments. The results showed that: 1) octreotide significantly inhibited the GH response to exercise, 2) exercise increased IGFBP-3 concentration (+37.4% at +90, p < 0.05), whereas no modification of IGFBP-2 and of IGF-I/ IGFBP-2 and IGFBP 3/IGFBP-2 ratios were observed, 3) octreotide amplified the IGFBP-3 increase after exercise (p < 0.01 vs. exercise, from + 30 to + 60, or octreotide alone) and, without exercise, slightly increased IGFBP-3 (+15% at +75, p < 0.05) and decreased IGF-I (-14.8% at +75, p < 0.01). We concluded that GH has a reduced role, as a stimulating factor, in the serum acute IGFBP-3 increase after exercise and that octreotide is probably able to directly amplify this response. Unfortunately, we can only speculate on the physiological pathways involved. PMID- 11281612 TI - Anaerobic power and achievement of VO2 plateau in pre-pubertal boys. AB - A low anaerobic power has been proposed as a factor that may be limiting the achievement of a plateau in VO2 of children who perform maximal aerobic power tests. This study examined the frequency of plateau achievement in pre-pubertal children and compared VO2max, peak (PP) and mean (MP) anaerobic power in subjects who either achieved a plateau (PLAT) or did not (NO PLAT). Eighteen healthy pre pubertal (Tanner Stage, pubic hair = 1) males (age = 9.1 1.6 yrs, ht = 134.4 +/- 9.7cm, wt = 33.3 +/- 9.2kg, VO2max = 40.0 +/- 6.7 ml x kg(-1) min(-1)) were tested. All subjects completed a 30 sec Wingate Anaerobic Test and a McMaster aerobic protocol to volitional fatigue on a cycle ergometer. Only 33% of the subjects met the PLAT criterion. No differences were found for PP or MP between those who achieved a plateau and those who did not (PLAT: PP= 6.3 +/- 0.8W/kg and MP = 5.2 +/- 0.7W/kg; NO PLAT: PP= 6.3 +/- 1.2 W/kg and MP = 5.2 +/- 1.3 W/kg). We conclude that anaerobic power is not a factor limiting the achievement of a plateau in VO2 of pre-pubertal boys who perform maximal aerobic power tests. PMID- 11281613 TI - Validation of a rating scale of perceived exertion in young children. AB - The aim of this study was to develop and validate a scale rating the perceived exertion of young children (RPE-C) who do not read. This scale presents seven pictures showing a man who becomes progressively fatigued. In order to evaluate this scale, we examined its reliability and sensitivity. Thirteen children (aged 5 to 6 years) completed two identical incremental maximal running trials during a period of one week. A group of 12 adolescents and a group of 12 adults also tested the RPE-C by performing one trial under the same experimental conditions as the child group. RPE-C was recorded at the end of each load level. Statistical analysis revealed significant effects for velocity (F = 23.98, p < 0.0001) with non-significant effects for the trials. Intraclass correlation coefficient (ICC) revealed an acceptable reliability of RPE-C at low and high exercise intensities but also showed a low reliability at intermediate load levels (0.17 < ICC < 0.77). A significant correlation between RPE-C and heart rate (r2 = 0.61, p < 0.0001) was observed in children. However, the r2 level was lower than that observed for the adolescent or adult groups. This finding demonstrates the sensitivity of RPE-C to discriminate the effects of age on perceived exertion. RPE-C seems to be a good tool which can be used in young children to monitor exercise performed at low and high intensities. PMID- 11281614 TI - Attenuation of increase in circulating cortisol and enhancement of the acute phase protein response in vitamin C-supplemented ultramarathoners. AB - Supplementary vitamin C (2 x 500 mg tablets daily) or a matched placebo was administered to 10 and 6 ultramarathon athletes respectively for 7 days prior to participation in a 90 kilometer running event, as well as on the day of the race and for 2 days after its completion. Circulating concentrations of vitamins A, C and E, as well as those of leukocytes and platelets, myeloperoxidase, C-reactive protein (CRP), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF), cortisol, and creatine kinase were measured 16 hours before the race and at 30 min, 24 hours, and 48 hours after completion. Pre-race vitamin C concentrations in the supplemented group were unchanged after the race (118.2 +/- 15.9 and 115.9 +/- 11.9 micromol/l) while an increase was observed in the placebo group immediately post-race (85.8 +/- 11.9 to 107.4 +/- 18.8 micromol), with a return to pre-race values after 24 hours. Immediately on completion of the race transient elevations occurred in the concentrations of circulating neutrophils, monocytes and platelets, IL-6, cortisol, CRP, and creatine kinase in both groups. In the supplemented group the concentrations of CRP were significantly higher (p < 0.01) at each of the post-race time-points while those of cortisol were 30% lower immediately post-race. These observations provide evidence that supplementation with vitamin C may blunt the adaptive mobilization of this vitamin from the adrenals during exercise-induced oxidative stress and may be associated with an enhancement of the acute phase protein response and attenuation of the exercise induced increase in serum cortisol. PMID- 11281615 TI - Mucosal IgA response to repeated wingate tests in females. AB - The purpose of this study was to examine the effect of repeated bouts of brief, intermittent, maximal effort exercise on the concentration of secretory IgA (S IgA) and incidence of upper respiratory tract infection (URTI) in females. Twenty six non-smoking, active, adult females participated in this study. The exercise intervention consisted of three consecutive, all-out 30-second Wingate leg cycling tests (0.075 kg/kg body mass), which were separated by three minutes of recovery. Unstimulated saliva was collected prior to (PRE) and five minutes after completion (POST) of the exercise testing protocol. IgA, protein and osmolality were measured, and secretory IgA (SIgA), IgA:Protein, and IgA: Osmolality were calculated. In addition, subjects completed standard logs indicating signs and symptoms of upper respiratory tract infection (URTI) for three weeks following the test. Saliva flow rates (478.5 +/- 50; 345.4 +/- 50 microl/min), S-IgA (55.8 +/- 4.7; 35.4 +/- 3.6 microg/min), and IgA: Protein ratio (30.7 +/- 3.0; 17.5 +/- 1.8 microg IgA/mg protein) were significantly lower POST compared to PRE (P < 0.05). No significant differences were observed for raw IgA, protein, osmolality, and IgA: osmolality. The results of this study show that brief, intermittent, maximal effort exercise results in an acute decrease in S-IgA in healthy adult females in less than eight minutes. However, this exercise induced transient decrease did not appear to be associated with an increase in clinical symptoms of URTI in the weeks following the exercise test intervention. PMID- 11281616 TI - The effect of moderate aerobic exercise and relaxation on secretory immunoglobulin A. AB - A deficiency in secretory immunoglobulin A (sIgA) is associated with recurrent upper respiratory tract infections both in the general community and in elite athletes. The aim of this paper was to investigate the effect of aerobic exercise and relaxation on various indices of sIgA in 12 male and 8 female adults who varied in levels of recreational activity. Salivary samples were obtained before, immediately after and 30 minutes after an incremental cycle ergometer test to fatigue, after 30 minutes of cycling at 30% or 60% of maximum heart rate, and after 30 minutes of relaxation with guided imagery. Each session was run on a separate day. When expressed in relation to changes in salivary flow rate, sIgA did not change after exercise. However, both the absolute concentration and secretion rate of sIgA increased during relaxation (167 +/- 179 microg x ml(-1), p < 0.001; and 37 +/- 71 microg x min(-1), p < 0.05 respectively). Nonspecific protein increased more than sIgA during incremental exercise to fatigue (decrease in the sIgA/protein ratio 92 +/- 181 microg x mg protein(-1), p < 0.05), but sIgA relative to protein did not change during relaxation. Our findings suggest that sIgA secretion rate is a more appropriate measure of sIgA than sIgA relative to protein, both for exercise and relaxation. These data suggest the possibility of using relaxation to counteract the negative effects of intense exercise on sIgA levels. PMID- 11281617 TI - Growth changes in the elastic properties of human tendon structures. AB - The purpose of this study was to investigate the growth changes in the elastic properties of human tendon structures. 9 younger boys (age 10.8 +/- 0.9 years, YBG), 9 elder boys (14.8 +/- 0.3 years, EBG), and 14 young adult men (24.7 +/- 1.6 years, ADG) volunteered to take part in the present study. Using a B-mode ultrasonic apparatus, the elongation of tendon and aponeurosis of vastus lateralis muscle (VL) was noninvasively measured in vivo, while subjects performed the extension of knee joint isometrically with force production levels from zero (relax) to maximal voluntary contraction (MVC) within 5 seconds. A curvilinear relationship was found between elongation of tendon structures (dL) and muscle force (Fm). This relationship consisted of two components, a steep initial change in length followed by a linear-region. The relationship between dL and Fm was fitted to a linear regression, and then the dL and dFm within 50 to 100% MVC was defined as compliance of tendon structures. The MVC force was the greatest in ADG and the lowest in YBG among the three groups. Significant age related differences were found in compliance; 4.1 +/- 0.9 x 10(-2) mm/N for YBG, 2.9 +/- 1.1 x 10(-2) mm/N for EBG and 1.8 +/- 0.3 x 10(-2) mm/N for ADG. The dL/thigh length (TL) was significantly greater in YBG than in the other two groups above 0.35 MPa of Fm per muscle cross-sectional area (muscle stress). However, there was no significant difference between EBG and ADG in the relationship between dL/TL and muscle stress. The ratio of fascicle length to TL in YBG was significantly lower than those in the other two groups. These results suggest that the tendon structures in younger boys are more compliant than those in older boys and young men. The observed properties of tendon structures in the younger boys may play a role in protecting younger boys from athletic injuries associated with immature muscle-tendon complex. PMID- 11281618 TI - Proprioception in anterior cruciate ligament reconstruction. Endoscopic versus open two-tunnel technique. A prospective study. AB - ACL-reconstruction with patellar tendon autograft is a standard procedure which can be performed arthroscopically with a femoral half tunnel drilled from the joint or using the two-tunnel technique with medial miniarthrotomy and additional femoral approach. The arthroscopic procedure with single incision was hypothesized to improve proprioception and to provide earlier rehabilitation. Twenty-nine patients with chronic ACL-deficiency were included in the prospective study. Fifteen patients were operated endoscopically, 14 patients using the two tunnel technique. Proprioception, Lysholm and Tegner scores as well as stability (KT-1000) were assessed preoperatively, 3 and 6 months postoperatively as well as after 3.9 +/- 0.4 years. A significant deficit of proprioception was assessed in both groups preoperatively. Six months postoperatively, both groups showed a restitution of proprioception near full extension and full flexion of the knee. In the mid-range position, the proprioception could not be restored. At the final examination after 3.9 years, the deficit documented in the mid-range position still persisted. There were no differences in proprioception, clinical results and stability between the arthroscopic and the open technique. PMID- 11281619 TI - Shock absorption capacities of mouthguards in different types and thicknesses. AB - Although sports mouthguards provide protection against trauma, dentoalveolar injuries can still occur with the mouthguards in place. This study examined the effect of mouthguard protection in an in vitro model. A simulated maxilla, out of a polymethylmethacrylate (PMMA) arch, containing replaceable resin teeth, was used to assess the performance of different mouthguard designs. "Boil and bite" and custom-fitted mouthguards (ethylene vinyl acetate [EVA]) laminated with hard (poly-vinyl chloride [PVC]) or soft labial intermediate EVA layers were fabricated according to manufacturers' instructions. A steel ram was dropped onto the mouthguards at the maxillary incisor region. Changes in voltage, which were induced by a strain gauge at the back of the upper left incisor, were measured with an amplified voltmeter. Data were analysed by ANOVA at a significance level of 0.05. "Boil and bite" and mouthguards layered with silicone or with small hard PVC inserts of 1.5 mm thickness demonstrated less absorption and differed significantly from the other mouthguard systems (p < 0.05). Bilaminated mouthguards with hard PVC inserts of 0.8 mm, 1.5 mm or 2 mm thickness showed no significant differences to those with 1.5 mm thick (EVA) inserts. The absorption rates amounted to 33 % compared with the unprotected tooth. PMID- 11281620 TI - Motor performance of the foot after Achilles rupture repair. AB - BACKGROUND: The purpose of this study was to examine some motor performance aspects of the unloaded lower extremity after Achilles tendon rupture repair. The measured motor performance aspects were simple reaction time, choice reaction time, speed of movement, foot tapping speed, and coordination. METHODS: Ninety patients (76 men and 14 women) with total closed Achilles tendon rupture had been operated on a mean of 3.1 years before the measurements. Ninety age and gender matched control subjects were drawn from a larger reference group from the local population. Age and gender-matched pairs were set up, and the results were compared. The measurements were made with the HPM/BEP-system, which is a multifunctional system designed to measure different motor aspects of the feet, including reaction time, movement speed, tapping speed, and coordination. RESULTS: There were no statistically significant differences in the results between the operated and non-operated lower extremities in the patient group a mean of 3.1 years after the operation. When the results were compared between the patient and control groups, no statistically significant differences were found. CONCLUSION: Based on the results, it seems that the measured motor performance functions of the unloaded lower extremity had fully recovered after the Achilles tendon rupture repair in the above mentioned aspects, and the operated patients do not have an increased risk to get reinjury of Achilles tendon because of the lower performance of these motor functions of the lower extremity. PMID- 11281621 TI - A descriptive epidemiological study of shoulder injury in top level English male volleyball players. AB - The aims of this study were to estimate the prevalence and incidence of shoulder sports injuries, to discover the main shoulder injury, and to survey outcome of treatment or injuries in top level male volleyball athletes. Furthermore, the actions which most commonly cause injuries and the differences of physical characteristics between injured and healthy players were also investigated. Fifty nine English Volleyball Federation division one athletes were recruited in the 1997/98 and 1998/99 seasons. All subjects completed two different questionnaires; a First recruitment and monthly Follow-up questionnaire throughout the period in question. Twenty-seven of the fifty-nine athletes had a history of shoulder sports injury, with a total of 29 injuries reported. The results of the First recruitment showed that overuse type injuries (19/29) were the main shoulder injuries. Cuff muscle tendinitis was predominant in these injuries (14/29). Furthermore, spiking was the major action during which a shoulder injury (23/29) first occurred. In the follow-up phase the incidences of shoulder chronic injury (or pain), re-injury, and new injury in these twenty-seven players were 3.0, 9.3 and 1.0 injuries/1,000 hours of exposure respectively. The mean duration of chronic injury or pain was 2.3 +/- 1.3 (+/- SD) months. The distribution of history of regular training, between injured and healthy subject groups, was significantly different (p = 0.008). This study has identified rotator cuff muscle/tendon injuries or involved lesions as the main shoulder injuries in top level English male volleyball athletes. These injuries result in prolonged shoulder pain symptoms. PMID- 11281622 TI - Respiratory effects of a single dive to 50 meters in sport divers with asymptomatic respiratory atopy. AB - Increasing popularity of sports diving makes it likely that subjects with allergic respiratory diseases will be involved in diving with self contained underwater breathing apparatus (scuba). The present study evaluated the effects of a single scuba-dive on pulmonary function in subjects with respiratory atopy. Specific airways conductance (sGaw), residual volume (RV), forced vital capacity (FVC), forced expiratory volume in 1 sec (FEV1), mid expiratory flow at 50% of FVC (MEF50), and transfer factor for carbon monoxide (TLCO) were measured in 9 sport divers with a history of hay fever and 9 matched healthy sport divers (control) before, 3 hours and 24 hours after a wet hyperbaric chamber dive to a depth of 50 m. Airway hyperresponsiveness (AHR) was assessed by methacholine challenge 4 weeks after the dive. Atopic subjects and controls did not differ with respect to anthropometric data, diving experience, and predive lung function. A 3% reduction in FVC was found 24h after the dive (p < 0.05) in both groups, whereas sGaw decreased by 15% 24 h after the dive (p < 0.05) in the subjects with respiratory atopy only. Postdive changes in RV, FEV1, MEF50, and TLCO did not reach level of statistical significance. AHR was obtained in 8/9 subjects with respiratory atopy. We conclude that subjects with atopic sensitization and asymptomatic AHR may be more susceptible to effects of diving on pulmonary function. PMID- 11281623 TI - Effect of aerobic training and detraining on left ventricular dimensions and diastolic function in prepubertal boys and girls. AB - PURPOSE: The purpose of this study was to determine the effect of an aerobic training program on the left ventricular (LV) cardiac morphology and function of prepubertal children. METHODS: Twenty-nine 10-11 year old boys and girls (TG) participated in a 13-week running program (3 x 1 h/week, intensity: > 80% HRmax), 26, of the same age, served as a control group (CG). M-mode, 2-dimensional and pulsed-wave Doppler analyses were performed, during resting conditions, before and after the training period (T) as well as, for TG only, after 2 months of detraining (D). RESULTS: LV internal chamber dimension increased (+ 4.6 %, p < 0.01) while wall thicknesses concomitantly decreased (-10.7%, p < 0.05) as a result of T. All cardiac morphological parameters returned to pretraining values after D. Doppler-derived measurements of LV diastolic filling performance were also significantly altered by Tand D. A significant enhancement in the early diastolic passive LV filling with a concomitant reduction in the late diastolic active LV filling were in fact obtained after T. The training-induced bradycardia (-7 beats x min(-1), p < 0.01) was probably responsible for the changes in the late characteristics of the diastolic active filling. All diastolic filling indexes returned to pretraining values after D. Systolic function indexes were not modified after either T or D. No changes were obtained for the overall LV morphological and functional variables after 13 weeks of normal life for CG. CONCLUSION: These findings indicate that cardiac morphological adaptations can occur in prepubertal children after several months of aerobic training. These alterations differ however, in some areas, to those classically reported in adults following endurance training programs where both an increase in LV size and mass exist. Our data likewise demonstrate that endurance training is able to induce favourable LV diastolic filling modifications, directed principally towards an enhancement in the early rapid filling inflow and a corresponding reduction in the atrial contribution to the total diastolic inflow. PMID- 11281624 TI - Cutaneous vascular response and thermoregulation in individuals with paraplegia during sustained arm-cranking exercise. AB - This study investigated whether a 60-minute arm-cranking exercise at 50% of the individual maximal power output would increase lower limb skin blood flow (laser Doppler flowmetry) in individuals with high-level (T5-T9; n = 6) and low-level paraplegia (T10-T12; n = 6), compared to 6 able-bodied controls. Significant (P < 0.05) group by time interactions (two-way repeated measures ANOVA) were found for leg cutaneous vascular conductance, leg skin temperature and esophageal temperature. Cutaneous vascular conductance increased to a peak of approximately 180% of pre-exercise rest in both paraplegic groups and to -436% in the control group, with differences after 15, 30, 45 and 60 minutes of exercise. Leg skin temperature increased by approximately 0.3 C in individuals with paraplegia and decreased by approximately 2.0 C in able-bodied. Esophageal temperature increases at the end of exercise were higher in individuals with paraplegia (approximately 0.9 C) than in able-bodied subjects (approximately 0.5 C). Heart rate was higher in the paraplegic groups than in able-bodied, whilst stroke volume and cardiac output were not different (impedance cardiography). The data suggest that lesion level had no influence on the results. These findings indicate that there is no excessive shunting of blood to the skin of the lower limbs of individuals with paraplegia during sustained exercise. PMID- 11281625 TI - Inhibition of leukocyte chemiluminescence by platelets: role of platelet-bound fibrinogen. AB - Tethering of PMNL by platelets via CD62P has been shown to cause PMNL activation. Co-incubation of purified PMNL with platelets that were activated with thrombin and then fixed and washed, resulted in the formation of platelet-PMNL conjugates as well as in a generation of reactive oxygen species that were measured as luminol-enhanced chemiluminescence. When platelets were thrombin activated in the presence of RGDS to prevent binding of fibrinogen to membrane receptors, they had a reduced capacity to adhere to PMNL, but ROS generation was enhanced. In samples of citrated whole blood RGDS as well as the more specific platelet fibrinogen receptor antagonist GR144053F or a dissociation of the platelet glycoprotein IIb/IIIa complex markedly enhanced ROS generation that was induced by stirring the samples for 10 min at 1000 rpm, by 175%, 95% and 138%, respectively. Removal of platelets from the whole blood samples also resulted in an enhancement of stirring-induced ROS generation, which was inversely correlated to the platelet count. These data provide some evidence that platelets are capable of inhibiting ROS generation in PMNL by a mechanism that involves platelet-bound fibrinogen and probably depends on fibrinogen-mediated platelet-PMNL contact. PMID- 11281626 TI - Changes in platelet glycoprotein receptors after smoking--a flow cytometric study. AB - Cigarette smoking is accepted to be one of the major factors which increase the risk of coronary artery disease, peripheral vascular disease and stroke. A number of studies have been carried out on the acute and chronic effects of tobacco smoking on platelet activation. An enhancing effect of high nicotine cigarette smoking on platelet aggregation has been reported. Since platelet receptors are involved in the final stage of platelet aggregation, the intention of this study was to investigate platelet receptors in acute and chronic smokers before and after smoking. Nineteen chronic smokers, 18 acute smokers and 18 healthy non smoking controls were included in the present study. Platelet aggregation was carried out using ristocetin, adenosine diphosphate (ADP) and collagen both before and after smoking in acute and chronic smokers. Flow cytometric studies of platelets were carried out utilizing fluorescein isothiocyanate (FITC)-labelled anti-human fibrinogen antibody in unstimulated and ADP-stimulated platelets, FITC labelled anti-GP IIb/IIa antibody, FITC-labelled anti-GP Ib/IX antibody and FITC labelled P-selectin antibody. The intensity of fluorescence was graded into three groups and expressed in arbitrary units. The interesting data generated in the present work is that in vivo platelet activation occurs immediately after smoking a cigarette which is detected by using FITC-labelled anti-human fibrinogen antibody binding to platelet and by P-selectin expression. It is also quite evident from the present study that a significant number of circulating platelets are in the activated state in chronic smokers. Therefore this study suggests that smoking-induced platelet activation may be an important contributory mechanism for acute coronary events in smokers. PMID- 11281627 TI - A comparison of von Willebrand Factor antigen with platelet activity in vitro in normal and venous occlusion blood. AB - Von Willebrand Factor (vWF) is essential for normal haemostasis involving platelet aggregation induced by high shear forces. In vitro a functional test of platelet aggregation using the filterometer is abnormal in von Willebrand's disease. However in normal people there is no significant correlation between the antigenic assay of vWF and the filter results. To study this discrepancy normal blood before and during venous occlusion, and blood before and after infusion of 1 deamino-(8-D-arginine) vasopressin was studied. During venous occlusion (VO) the increase in vWF due to the release of large multimers correlated precisely with the increase in the filterometer results. That this was due to the plasma vWF and not to any change induced in the platelets was shown as follows: The methodology was altered so that a small amount of the donor's platelet-poor plasma (PPP) was added to homologous normal substrate blood. The effect of the added donor's PPP was then shown to be closely correlated to the increase in the antigenic assay. Analysis of vWF multimer size showed during VO an increase in large multimers. We conclude that the effect of vWF on normal blood may be obscured by variation in platelet aggregability. In the filterometer system as elsewhere the large active multimers probably play a major part in causing platelet adhesion, aggregation and filter blocking. The filterometer test is influenced by the amount of vWF antigen, by the molecular size and activity of the vWF and by platelet sensitivity. Clinically this is a useful global test. PMID- 11281628 TI - Platelet aggregatory responses to low-dose collagen are maintained in hirudin anticoagulated whole blood for 24 h when stored at room temperature. AB - Whole blood from 15 volunteers was anticoagulated with hirudin (200U/l) and the response to a known submaximal concentration of collagen (0.6 microg/ml) was tested by impedance aggregometry. In 8 volunteers platelet counts were also taken before and after the maximum aggregatory response. These tests were repeated when the samples had rested for 24 h at room temperature. The median [interquartile range] aggregatory response immediately after sampling was 17.3 [16.7-18.4] ohms. At 24 h it was 17.7 [15.8-19.3] ohms (p = 0.88) although variance was increased (p = 0.006). The immediate platelet count before collagen exposure was 438 [381 510] x 10(9)/l and 258 [227-297] x 10(9)/l post-collagen. At 24 h the platelet count was 448 [443-473] x 10(9)/l (p = 0.224 versus immediate count) but variance was not increased (p = 0.215). After full aggregation the count fell to 284 [234 304] x 10(9)/l (p = 0.592 versus early post-collagen). Variances were similar (p = 0.558). Aggregate response ratios increased non-significantly after 24 h from 0.59 [0.53-0.62] to 0.64 [0.51-0.68] although variance was increased (p = 0.021). Full macroaggregatory responses by impedance aggregometry were seen after 24h storage of whole blood with hirudin at room temperature. This suggests both that distant assessment of platelet function using a standardized method is possible and a potential role of thrombin inhibition for platelet storage. PMID- 11281629 TI - Soluble P-selectin is influenced by cancer chemotherapy. PMID- 11281630 TI - Acquired platelet dysfunction with eosinophilia in children in the south of Thailand. AB - One hundred and sixty-eight children aged 13 months to 12.6 years with acquired platelet dysfunction with eosinophilia (APDE) were studied. The male to female ratio was 1.15:1. All of the children were in good health and no history of any drug ingestion was detected. All of the children had widespread spontaneous bruising on the extremities, body and face off and on. Severe bleeding symptoms were detected in 8% of these patients. The number of platelets in these children was within the normal range but the platelet morphology was abnormal in all of them. Eosinophilia was detected in 86% of these children. Prolonged bleeding time was detected in 53% of these patients. Abnormal platelet adhesiveness was found in 33% of cases. Abnormal platelet aggregation induced by collagen was the most sensitive test in these patients. Abnormal ADP release from the platelets was detected in these patients by the absence of a second wave of aggregation during stimulation of PRP by ADP or epinephrine. Abnormal or no ATP secretion from the platelets during stimulation by ADP, epinephrine or collagen was detected in these patients. Ristocetin-induced platelet aggregation was normal in these children. Decreased or absence of platelet dense granules by TEM study was detected in some patients. These changes in platelet functions and morphology may be due to acquired storage pool deficiency of the platelet. Parasitic infection was detected in 56% of these children. About 83% of these children with APDE had serum total IgE higher than 100 IU/ml. There was no correlation between the number of eosinophils and serum total IgE and the severity of bleeding symptoms. The majority of children with APDE did not receive any treatment except those who had severe bleeding symptoms which required platelet concentrate to stop bleeding. In more than 90% of the patients, the bruising or ecchymosis disappeared within 6 months and the abnormal platelet functions returned to normal within 4 months. Recurrence of these bleeding syndromes was detected in 7% of the children. PMID- 11281631 TI - Use of glucosamine and chondroitin sulfate in the management of osteoarthritis. AB - The goals of osteoarthritis therapy are to decrease pain and to maintain or improve joint function. The pharmacologic treatment of this condition has included the use of aspirin, acetaminophen, and nonsteroidal anti-inflammatory drugs. More recently, numerous studies have investigated the potential role of chondroprotective agents in repairing articular cartilage and decelerating the degenerative process. The reports of limited clinical experience with two of these agents, glucosamine and chondroitin sulfate, as well as the accompanying publicity in the popular media, have generated controversy. Advocates of these alternative modalities cite reports of progressive and gradual decline of joint pain and tenderness, improved mobility, sustained improvement after drug withdrawal, and a lack of significant toxicity associated with short-term use of these agents. Critics point out that in the great majority of the relevant clinical trials, sample sizes were small and follow-up was short-term. PMID- 11281632 TI - Protrusio acetabuli: diagnosis and treatment. AB - Idiopathic protrusio acetabuli is an uncommon disease process with both primary idiopathic and secondary forms. It is important to consider all etiologic possibilities before evaluating treatment options. Diagnosis is made on the basis of an anteroposterior radiograph of the pelvis that demonstrates a center-edge angle greater than 40 degrees and medialization of the medial wall of the acetabulum past the ilioischial line. For the skeletally immature patient, triradiate fusion (occasionally combined with intertrochanteric osteotomy) provides good results. For the young adult, valgus intertrochanteric proximal femoral osteotomy is recommended. In the older adult, this procedure may provide an acceptable result if there is minimal arthritis. For patients with more advanced arthritis, total hip arthroplasty with lateralization of the cup to a normal position provides a predictable long-term solution. PMID- 11281633 TI - The limping child: evaluation and diagnosis. AB - A limp is a common reason for a child to present to the orthopaedist. Because of the long list of potential diagnoses, some of which demand urgent treatment, an organized approach to evaluation is required. With an understanding of normal and abnormal gait, a directed history and physical examination, and the development of a differential diagnosis based on the type of limp, the patient's age, and the anatomic site that is most likely affected, the orthopaedist can take a selective approach to diagnostic testing. Laboratory tests are indicated when infection, inflammatory arthritis, or a malignant condition is in the differential diagnosis. The C-reactive protein assay is the most sensitive early test for musculoskeletal infections; an abnormal value rapidly returns to normal with effective treatment. Imaging should begin with plain radiography. Ultrasonography is particularly valuable in assessing the irritable hip and guiding aspiration, if necessary. PMID- 11281634 TI - Medial elbow problems in the overhead-throwing athlete. AB - The elbow is subjected to enormous valgus stresses during the throwing motion, which places the overhead-throwing athlete at considerable risk for injury. Injuries involving the structures of the medial elbow occur in distinct patterns. Although acute injuries of the medial elbow can occur, the majority are overuse injuries as a result of the repetitive forces imparted to the elbow by throwing. Injury to the ulnar collateral ligament complex results in valgus instability. Valgus extension overload leads to diffuse osseous changes within the elbow joint and secondary posteromedial impingement. Overuse of the flexor-pronator musculature may result in medial epicondylitis and occasional muscle tears and ruptures. Ulnar neuropathy is a common finding that may be due to a variety of factors, including traction, friction, and compression of the ulnar nerve. Advances in nonoperative and operative treatment regimens specific to each injury pattern have resulted in the restoration of elbow function and the successful return of most injured overhead athletes to competitive activities. With further insight into the relevant anatomy, biomechanics, and pathophysiology involved in overhead activities and their associated injuries, significant contributions can continue to be made toward prevention and treatment of these injuries. PMID- 11281635 TI - Talus fractures: evaluation and treatment. AB - Fractures of the talus are uncommon. The relative infrequency of these injuries in part accounts for the lack of useful and objective data to guide treatment. The integrity of the talus is critical to normal function of the ankle, subtalar, and transverse tarsal joints. Injuries to the head, neck, or body of the talus can interfere with normal coupled motion of these joints and result in permanent pain, loss of motion, and deformity. Outcomes vary widely and are related to the degree of initial fracture displacement. Nondisplaced fractures have a favorable outcome in most cases. Failure to recognize fracture displacement (even when minimal) can lead to undertreatment and poor outcomes. The accuracy of closed reduction of displaced talar neck fractures can be very difficult to assess. Operative treatment should, therefore, be considered for all displaced fractures. Osteonecrosis and malunion are common complications, and prompt and accurate reduction minimizes their incidence and severity. The use of titanium screws for fixation permits magnetic resonance imaging, which may allow earlier assessment of osteonecrosis; however, further investigation is necessary to determine the clinical utility of this information. Unrecognized medial talar neck comminution can lead to varus malunion and a supination deformity with decreased range of motion of the subtalar joint. Combined anteromedial and anterolateral exposure of talar neck fractures can help ensure anatomic reduction. Posttraumatic hindfoot arthrosis has been reported to occur in more than 90% of patients with displaced talus fractures. Salvage can be difficult and often necessitates extended arthrodesis procedures. PMID- 11281637 TI - Neurologic complications after lumbar spine surgery. AB - With the increasing complexity and number of lumbar spine operations being performed, the potential number of patients who will sustain perioperative complications, including those that involve neural structures, has also increased. Neurologic complications after lumbar spine surgery can be categorized by the perioperative time period during which they occur and by their mechanism of injury. Although the overall incidence of neurologic complications after lumbar surgery is low, the severity of these injuries mandates careful preoperative planning, awareness of risk, and meticulous attention to perioperative details. PMID- 11281636 TI - Kienbock's disease: diagnosis and treatment. AB - Kienbock's disease, or osteonecrosis of the lunate, can lead to chronic, debilitating wrist pain. Etiologic factors include vascular and skeletal variations combined with trauma or repetitive loading. In stage I Kienbock's disease, plain radiographs appear normal, and bone scintigraphy or magnetic resonance imaging is required for diagnosis. Initial treatment is nonoperative. In stage II, sclerosis of the lunate, compression fracture, and/or early collapse of the radial border of the lunate may appear. In stage IIIA, there is more severe lunate collapse. Because the remainder of the carpus is still uninvolved, treatment in stages II and IIIA involves attempts at revascularization of the lunate-either directly (with vascularized bone grafting) or indirectly (by unloading the lunate). Radial shortening in wrists with negative ulnar variance and capitate shortening or radial-wedge osteotomy in wrists with neutral or positive ulnar variance can be performed alone or with vascularized bone grafting. In stage IIIB, palmar rotation of the scaphoid and proximal migration of the capitate occur, and treatment addresses the carpal collapse. Surgical options include scaphotrapeziotrapezoid or scaphocapitate arthrodesis to correct scaphoid hyperflexion. In stage IV, degenerative changes are present at the midcarpal joint, the radiocarpal joint, or both. Treatment options include proximal-row carpectomy and wrist arthrodesis. PMID- 11281638 TI - A nuclear protein tyrosine phosphatase induces shortening of G1 phase and increase in c-Myc protein level. AB - PTP-S2 is a ubiquitously expressed nuclear protein tyrosine phosphatase which shows increased expression upon mitogenic stimulation in a variety of cells in vitro and in vivo. In order to understand the role of this enzyme in cell cycle progression, tetracycline-regulated HeLa clones expressing PTP-S2 were isolated and characterized. Tetracycline-controlled expression of PTP-S2 increased the rate of cell proliferation. An analysis of the distribution of cells in various phases of the cell cycle in an exponentially growing cell population showed that there was a large decrease in the percentage of cells in G1 phase in a PTP-S2 expressing population of cells compared to nonexpressing cells. This decrease in the percentage of cells in G1 was dependent on the level of PTP-S2 expression. There was a corresponding increase in the percentage of cells in G2/M but no significant increase in the percentage of cells in S phase. An analysis of the time course of cell cycle progression after release from double thymidine block showed that the duration of G1 phase was significantly shortened in cells induced to express exogenous PTP-S2. However, the duration of S phase was not significantly altered and the duration of G2 phase was increased to some extent. Induction of PTP-S2 expression was associated with an increase in c-Myc protein levels, although the c-Myc mRNA level was not changed. Our results suggest that overexpression of PTP-S2 promotes progression of cells through G1 to S phase and is associated with increased level of c-Myc protein through a posttranscriptional mechanism. PMID- 11281639 TI - Apoptotic cell debris and phosphatidylserine-containing lipid vesicles induce apolipoprotein J (clusterin) gene expression in vital fibroblasts. AB - The molecular events in cells undergoing programmed cell death (apoptosis) are well studied; however, the response of the surviving neighbor cells to local cell death is largely uncharacterized. Apolipoprotein J (clusterin) is an 80-kDa glycoprotein that has been implied in cytoprotection of the vital cells, presumably by assisting in the clearance of apoptotic vesicles and membrane remnants. Its mRNA is specifically up-regulated in the vital cells of apoptotic tissues. The molecular mechanisms, however, leading to this response are not known. We here show that exposure of vital fibroblasts to apoptotic vesicles, disrupted vital cells, and trypsin-treated membrane remnants induces apoJ mRNA. Moreover, lipid vesicles consisting of phosphatidylserine (PtSer) and dimyristoylphosphatidylcholine (PC), but not liposomes with PC alone nor with dimyristoylphosphatidylethanolamine or phosphatidic acid, did elevate apoJ mRNA level. These results suggest that, apart from mediating the endocytic uptake of the apoptotic vesicles, PtSer also serves as a trigger to stimulate the expression of genes that might be involved in the cellular clearance process. PMID- 11281640 TI - A functional role for ERK in gene induction, but not in neurite outgrowth in differentiating neuroblastoma cells. AB - The human neuroblastoma cell line SH-SY5Y can differentiate into a functional sympathetic neuronal phenotype when treated with low concentrations of the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) in the presence of serum or defined growth factors. When TrkA is introduced into the cells, NGF also induces differentiation. In both cases, protein kinase C (PKC) is pivotal for induction and maintenance of the differentiated phenotype. We have recently shown that PKC activity is needed to enable the MAPK ERK to accumulate in the nucleus of SH-SY5Y cells and hence activate transcription. To find out whether this could be one reason for the PKC dependency in the differentiation process we have investigated the role of ERK during neuronal differentiation of these cells. The results show that ERK was needed for full upregulation of the neuronal marker genes NPY and GAP-43. However, ERK activity was not necessary for TPA-induced neurite formation. Neither was activation of ERK sufficient to promote neurite outgrowth. The results clearly show that there was no correlation between nuclear ERK activity, measured as SRE transactivation, and neurite formation in TPA differentiated SH-SY5Y neuroblastoma cells. PMID- 11281641 TI - Nuclear accumulation of exogenous DNA fragments in viable cells mediated by FGF-2 and DNA release upon cellular injury. AB - We and others have previously shown that basic fibroblast growth factor (FGF-2 or bFGF) can be used as a targeting molecule to help carry plasmid DNA into cells when the growth factor molecule is physically coupled to the DNA molecule being delivered. Herein we report our observations on the FGF-mediated uptake of exogenous labeled DNA into cultured cells in a manner that is representative of that which may occur under physiological conditions at sites of wounded tissue. Cellular debris at such sites contains nucleic acid fragments released from dead cells, as well as growth factors such as FGF-2 that function early in the wound repair process. Using a cell culture model designed to mimic the local environment of a wound with respect to the presence of soluble FGF-2 and DNA fragments, we have shown that FGF-2 is able to direct the cellular uptake and nuclear localization of fragments of exogenous DNA via the FGF receptor into intact and healthy cells. Furthermore, we can monitor and quantitate this type of FGF-mediated DNA delivery by using indirect immunofluorescence of bromodeoxyuridine-labeled exogenous DNA. Our results suggest that this type of FGF-mediated DNA fragment uptake could allow for the transduction of viable nearest neighbor cells at sites of injury in vivo. Such a phenomenon may lead to mutational aberrations in the recipient cells and enhance the probability of wound carcinogenesis. PMID- 11281642 TI - Overexpression of components of the Frizzled-Dishevelled cascade results in apoptotic cell death, mediated by beta-catenin. AB - Frizzled (fz) functions as a 7-transmembrane receptor in the Frizzled-Dishevelled signal transduction cascade. It is involved in architectural control of development in species as divergent as Drosophila and vertebrates. Regulation of multicellular architecture requires control of cell alignment, but also involves an equilibrium among cell proliferation, differentiation, and apoptosis. Recently, modulation of the Frizzled-Dishevelled (Dvl) cascade has been related to apoptosis. However, the role of beta-catenin, a second messenger in the Frizzled-Dishevelled cascade, in programmed cell death is a matter of debate. To elucidate the role of this cascade in apoptosis, we studied the effect of overexpression of fz1, fz2, dvl1, and beta-catenin. The signal transduction pathway and the involvement of beta-catenin were further investigated by using different inhibitors. These experiments were performed in different cell types: COS7, 293, and PC12. Overexpression of fz1, fz2, and dvl1 induced apoptosis in COS7 and 293 cells. beta-Catenin appears to be the mediator for this process since beta-catenin overexpression as well as lithium and valproate induced apoptosis. In contrast, lithium treatment did not result in apoptosis in PC12 cells. We conclude that different components of the Frizzled-Dishevelled cascade can induce apoptosis, but that this effect is dependent on the cell type. PMID- 11281643 TI - The fibronectin-derived antiadhesive peptides suppress the myofibroblastic conversion of rat hepatic stellate cells. AB - We previously found that fibronectin (FN) had a functional site (YTIYVIAL sequence in the 14th type III module) suppressing the integrin-mediated cell adhesion to extracellular matrix. FN-derived peptides containing this antiadhesive site were also shown to regulate cellular processes such as proliferation, differentiation, and apoptosis. The present study shows that the FN-derived antiadhesive peptides suppress the myofibroblastic conversion of rat hepatic stellate cells (HSC). Freshly isolated HSC underwent myofibroblastic conversion during culture in the presence of FBS, as evaluated by indices representing the phenotypic activation of HSC, including increased proliferation, consumption of vitamin A-enriched lipid droplets, and expression of alpha-smooth muscle actin. However, appearance of these myofibroblastic characters was suppressed by coculturing HSC with the FN-derived antiadhesive peptides. On the other hand, the activated HSC, which had already acquired the myofibroblastic phenotype through repeated subculture, secreted FN and then stimulated matrix assembly of ED-A (+) cellular FN as well as plasma FN, while the FN-derived antiadhesive peptides inhibited them. Furthermore, the FN-derived antiadhesive peptides suppressed the integrin-mediated adhesion of the primary HSC to plasma FN and ED-A (+) cellular FN substrates. These results suggested that the FN derived antiadhesive peptides down-regulated the myofibroblastic conversion of HSC in an indirect manner by inhibiting the integrin-mediated adhesive interaction of HSC with ED-A (+) cellular FN. PMID- 11281644 TI - Direct inhibition of Indian hedgehog expression by parathyroid hormone (PTH)/PTH related peptide and up-regulation by retinoic acid in growth plate chondrocyte cultures. AB - Indian hedgehog (Ihh) is highly expressed in prehypertrophic chondrocytes in vivo and has been proposed to regulate the proliferation and maturation of chondrocytes and bone collar formation in the growth plate. In high-density cultures of rabbit growth-plate chondrocytes, Ihh mRNA was also expressed at the highest level in the prehypertrophic stage. To explore endogenous factors that regulate Ihh expression in chondrocytes, we examined the effects of various growth factors on Ihh mRNA expression in this system. Retinoic acid (RA) and bone morphogenetic protein-2 enhanced Ihh mRNA expression, whereas PTH/PTH-related peptide (PTHrP) markedly suppressed Ihh expression. RA at more than 10(-8) M induced the expression of Ihh and Patched 1 (Ptc1) within 3 h, before it increased the type X collagen mRNA level at 6-24 h. Cycloheximide blocked the up regulation of Ihh by RA, indicating the requirement of de novo protein synthesis for this stimulation. These findings suggest that RA is involved in the up regulation of Ihh during endochondral bone formation. In contrast to RA, PTH (1 84) at 10(-7) M abolished the mRNA expression of Ihh and Ptc1 within 2-4 h, before it suppressed the expression of type X collagen at 12-24 h. The inhibition of Ihh expression by PTH (1-84) did not require de novo protein synthesis. PTH (1 34), PTHrP (1-34), and (Bu)(2)cAMP also suppressed Ihh expression. On the other hand, Ihh has been reported to induce PTHrP synthesis in the perichondrium. Consequently, the direct inhibitory action of PTH/PTHrP on Ihh appears to be a negative feedback mechanism that prevents excess PTHrP accumulation in cartilage. PMID- 11281645 TI - Cyclic AMP- and IL6-signaling cross talk: comodulation of proliferation and apoptosis in the 7TD1 B cell hybridoma. AB - Proliferation of the 7TD1 B cell hybridoma is dependent on the survival factor interleukin-6 (IL6). IL6 inhibits physiological cell death and allows expansion of populations of serum-stimulated cells. In this report, we demonstrate that cyclic AMP (cAMP)- and IL6-dependent signaling pathways can interact, controlling proliferation of 7TD1 cells through modulation of apoptosis. Cyclic AMP analogues inhibited proliferation, as well as other treatments that increased intracellular cAMP. The cAMP-induced inhibition could be reversed after 24 h by the removal of dibutyryl-cAMP from the culture medium and readdition of IL6. In the absence of IL6, cAMP induced a slow loss of viable cells. This decrease in viable cells in the presence of cAMP was accompanied by a marked increase in apoptosis. The increase in apoptotic cells after 48 h was preceded at 24 h by a parallel increase in DEVD-caspase activity after treatment with cell-permeable cAMP analogues. Increased DEVD-caspase activity and subsequent apoptosis could both be blocked by the addition of IL6. These coregulating actions may represent a cross talk signaling mechanism modulating cytokine activation of cellular proliferation and survival. PMID- 11281646 TI - The requirement of tyrosines 579 and 581 for maximal ligand-dependent activation of the betaPDGFR is influenced by noncytoplasmic regions of the receptor. AB - Mutating tyrosines 579 and 581 of the beta platelet-derived growth factor receptor (betaPDGFR) tyrosine kinase to phenylalanines (the F2 mutation) impair activation of the receptor in response to ligand, but mutation of the analogous tyrosines in the alphaPDGFR has no effect on ligand-dependent receptor activation. We have found that the F2 mutation has only a modest effect on ligand dependent activation of a chimeric PDGFR composed of the extracellular and transmembrane domains of the alphaPDGFR and the cytoplasmic domain of the betaPDGFR by three measures: (1) the ability to phosphorylate endogenous and exogenous protein substrates in vitro, (2) phosphorylation of tyrosine 857, and (3) binding of the effector proteins PLCgamma, RasGAP, and SHP-2. Conversely, the F2 mutation substantially impairs ligand-dependent activation of chimeric PDGFRs that consist of either the extracellular domain alone or the extracellular and transmembrane domains of the betaPDGFR and all remaining sequence from the alphaPDGFR by two measures: (1) phosphorylation of endogenous protein substrates in vitro and (2) binding of PLCgamma and SHP-2. Our results indicate that the requirement of tyrosines 579 and 581 for maximal activation of the betaPDGFR in response to ligand is primarily determined by noncytoplasmic regions of the receptor. PMID- 11281648 TI - Targeting of calsequestrin to the sarcoplasmic reticulum of skeletal muscle upon deletion of its glycosylation site. AB - The glycoprotein calsequestrin (CS) is segregated to the junctional sarcoplasmic reticulum (jSR) and is responsible for intraluminal Ca(2+) binding. A chimeric CS hemoagglutinin 1 (HA1), obtained by adding the nine amino acid viral epitope hemoagglutinin to the carboxy terminal of CS and shown to be correctly segregated to skeletal muscle jSR [A. Nori, K. A. Nadalini, A. Martini, R. Rizzuto, A. Villa, and P. Volpe (1997). Chimeric calsequestrin and its targeting to the junctional sarcoplasmic reticulum of skeletal muscle. Am. J. Physiol. 272, C1420 C1428] lends itself as a molecular tool to investigate the targeting domains of CS. A putative targeting mechanism of CS to jSR implies glycosylation-dependent steps in the endoplasmic reticulum (ER) and Golgi complex. To test this hypothesis, CS-HA1DeltaGly, a mutant in which the unique N-glycosylation site Asn316 was changed to Ile, was engineered by site-directed mutagenesis. The mutant cDNA was transiently transfected in either HeLa cells, myoblasts of rat skeletal muscle primary cultures, or regenerating soleus muscle fibers of adult rats. The expression and intracellular localization of CS-HA1DeltaGly was studied by double-labeling epifluorescence by means of antibodies against either CS, HA1, or the ryanodine receptor calcium release channel. CS-HA1DeltaGly was expressed and retained to ER and ER/sarcoplasmic reticulum of HeLa cells and myotubes, respectively, and expressed, sorted, and correctly segregated to jSR of regenerating soleus muscle fibers. Thus, the targeting mechanism of CS in vivo appears not to be affected by glycosylation-that is, the sorting, docking, and segregation of CS are independent of cotranslational and posttranslational glycosylation or glycosylations. PMID- 11281647 TI - dSIR2 and dHDAC6: two novel, inhibitor-resistant deacetylases in Drosophila melanogaster. AB - We have identified new members of the histone deacetylase enzyme family in Drosophila melanogaster. dHDAC6 is a class II deacetylase with two active sites, and dSIR2 is an NAD-dependent histone deacetylase. These proteins, together with two class I histone deacetylases, dHDAC1 and dHDAC3, have been expressed and characterized as epitope-tagged recombinant proteins in Schneider SL2 cells. All these proteins have in vitro deacetylase activity and are able to deacetylate core histone H4 at all four acetylatable lysine residues (5, 8, 12, and 16). Recombinant dHDAC6 and dSIR2 are both insensitive to TSA and HC toxin and resistant, relative to dHDAC1 and dHDAC3, to inhibition by sodium butyrate. Indirect immunofluorescence microscopy of stably transfected SL2 lines reveals that dHDAC1 and dSIR2 are nuclear, dHDAC6 is cytosolic, and dHDAC3 is detectable in both cytosol and nucleus. dHDAC6 and dSIR2 elute from Superose 6 columns with apparent molecular weights of 90 and 200 kDa, respectively. In contrast, dHDAC1 and dHDAC3elute at 800 and 700 kDa, respectively, suggesting that they are components of multiprotein complexes. Consistent with this, recombinant dHDAC1 coimmunoprecipitates with components of the Drosophila NuRD complex and dHDAC3 with an as yet unknown 45-kDa protein. PMID- 11281649 TI - c-JUN gene induction and AP-1 activity is regulated by a JNK-dependent pathway in hypoxic HepG2 cells. AB - Hypoxia is an important pathophysiological stress that occurs during blood vessel injuries and tumor growth. It is now well documented that hypoxia leads to the activation of several transcription factors which participate in the adaptive response of the cells to hypoxia. Among these transcription factors, AP-1 is rapidly activated by hypoxia and triggers bFGF, VEGF, and tyrosine hydroxylase gene expression. However, the mechanisms of AP-1 activation by hypoxia are not well understood. In this report, we studied the events leading to AP-1 activation in hypoxia. We found that c-jun protein accumulates in hypoxic HepG2 cells. This overexpression is concomitant with c-jun phosphorylation and JNK activation. Moreover, we showed that AP-1 is transcriptionally active. We also observed that AP-1 transcriptional activity is inhibited by a MEKK1 dominant negative mutant. Moreover, the MEKK1 dominant negative mutant as well as deletion of the AP-1 binding sites within the c-jun promoter inhibited the c-jun promoter activation by hypoxia. All together, these results indicate that, in hypoxic HepG2 cells, AP 1 is activated through a JNK-dependent pathway and that it is involved in the regulation of the c-jun promoter, inducing a positive feedback loop on AP-1 activation via c-jun overexpression. PMID- 11281651 TI - Assembly of laminin polymers is dependent on beta1-integrins. AB - Recent reports suggest that laminin deposition is controlled by the cell via specific receptors, one of which is dystroglycan. In this study, the involvement of beta1-integrins in this process was investigated by comparing beta1-integrin deficient cells of different phenotypes with their normal counterparts. Normal embryonic stem (ES) cells and embryoid bodies (EBs) derived from them were found to deposit cell-associated laminin into fibrillar networks, and in the EBs a basement membrane was assembled under the primitive endoderm. beta1-deficient ES cells and their EBs formed only small amounts of dot-like laminin deposits. Skeletal myotubes formed after prolonged differentiation in EBs were found to be surrounded by laminin, nidogen, and perlecan by immunofluorescent staining irrespective of the presence of beta1-integrins on the myotubes. However, at the electron microscope level only very thin sheet-like structures were detected close to the beta1-deficient myotubes, while the wt myotubes formed thick basement membranes. An epithelial cell line, GE11, derived from the beta1 integrin-deficient ES cells was also unable to assemble laminin on the cell surface, while transfection of the cells with the integrin beta1 subunit resulted in formation of a dense laminin network. Taken together, these results suggest that dystroglycan and beta1-integrins can both contribute to the recruitment of laminin to cell surfaces and that integrins are required at a subsequent step in the formation of basement membranes. PMID- 11281650 TI - Effects of simian virus 40 T-antigens on normal human mammary epithelial cells reveal evidence for spontaneous alterations in addition to loss of p16(INK4a) expression. AB - Under standard culture conditions, normal human mammary epithelial cells (HMECs) divide a limited number of times before proliferation ceases in a growth-arrested state referred to as selection. Cells that have undergone spontaneous loss of p16(INK4a) expression due to hypermethylation of the p16(INK4a) CpG island emerge from selection and proliferate for an extended, but limited, period before senescence. Here we show, as expected, that selection was bypassed by expression of SV40 large T-antigen proteins containing an intact pRb-binding domain in preselection cells. These cells were immortalized with high efficiency (seven of nine separate cultures). Also as expected, postselection cells were immortalized by expression of the human papillomavirus-16 E6 oncoprotein (four of four cultures), which inactivates p53 protein. In contrast, we found that expression of SV40 large T-antigen protein, which also inactivates p53, was poorly maintained in postselection cultures due to its growth-suppressive effects; consequently, these cells became immortalized at low efficiency (one of 11 cultures). Reexpression of p16(INK4a) in postselection HMECs by the demethylating agent, 5-azacytidine, or transfection of a p16(INK4a) expression plasmid did not restore the ability of these cells to undergo SV40-induced transformation. Postselection HMECs are a widely used in vitro model system, but these observations indicate they have undergone changes in gene expression in addition to loss of p16(INK4a) expression. PMID- 11281652 TI - Caspase-dependent cytosolic release of cytochrome c and membrane translocation of Bax in p53-induced apoptosis. AB - Activation of p53 induces apoptosis in various cell types. However, the mechanism by which p53 induces apoptosis is still unclear. We reported previously that the activation of a temperature-sensitive mutant p53 (p53(138Val)) induced activation of caspase 3 and apoptosis in Jurkat cells. To elucidate the pathway linking p53 and downstream caspases, we examined the activation of caspases 8 and 9 in apoptotic cells. The results showed that both caspases were activated during apoptosis as judged by the appearance of cleavage products from procaspases and the caspase activities to cleave specific fluorogenic substrates. The significant inhibition of apoptosis by a tetrapeptide inhibitor of caspase 8 and caspase 9 suggested that both caspases are required for apoptosis induction. In addition, the membrane translocation of Bax and cytosolic release of cytochrome c, but not loss of mitochondrial membrane potential, were detected at an early stage of apoptosis. Moreover, Bax translocation, cytochrome c release, and caspase 9 activation were blocked by the broad-spectrum caspase inhibitor, Z-VAD-fmk and the caspase 8-preferential inhibitor, Ac-IETD-CHO, suggesting that the mitochondria might participate in apoptosis by amplifying the upstream death signals. In conclusion, our results indicated that activation of caspase 8 or other caspase(s) by p53 triggered the membrane translocation of Bax and cytosolic release of cytochrome c, which might amplify the apoptotic signal by activating caspase 9 and its downstream caspases. PMID- 11281653 TI - Mechanisms of cell cycle arrest in response to TGF-beta in progestin-dependent and -independent growth of mammary tumors. AB - TGF-beta1 modulation of cell cycle components was assessed in an experimental model in which the synthetic progestin medroxyprogesterone acetate (MPA) induced mammary tumors in Balb/c mice. TGF-beta1 inhibited both MPA-induced proliferation of progestin-dependent C4HD epithelial cells and proliferation of the progestin independent variant cell type C4HI, arresting cells in G(1) phase of the cell cycle. Progestin-independent 60 epithelial cells evidenced reduced response to TGF-beta1 antiproliferative effects. TGF-beta1 inhibition of cyclins D1 and A expression and up-regulation of p21(CIP1) levels were the common findings in all three cell types. In addition, a significant content reduction of cyclin D1/cdk4 and cyclin A/cdk2 complexes was found after TGF-beta1 inhibition of MPA-dependent and -independent proliferation. TGF-beta1 inhibited cyclin D2 expression and up regulated p27(KIP1) levels only when acting as inhibitor of MPA-induced proliferation of C4HD cells. Regulation of these two cell cycle components resulted in decreased cyclin D2/cdk2 complex and in increased p27(KIP1) association with cdk2 in C4HD cells treated with TGF-beta1. These two molecular mechanisms, unobserved in progestin-independent growth of C4HI or 60 cells, were associated with a significantly higher degree of inhibition of cdk2 kinase activity in C4HD cells compared to that found in TGF-beta-treated C4HI or 60 cells. Reduced sensitivity of 60 cells to the growth-inhibitory effects of TGF beta1 correlated with significantly lower levels of p15(INK4B), p21(CIP1), and p27(KIP1) expressed in these cells, compared to the levels present in C4HD or C4HI cells, and correlated as well with lack of expression of p16(INK4). Thus, common targets were found to exist in TGF-beta1 inhibitory action on breast cancer cells, but regulation of specific targets was found when TGF-beta1 inhibited proliferation driven by the progesterone receptor. PMID- 11281655 TI - Involvement of polyamines in B cell receptor-mediated apoptosis: spermine functions as a negative modulator. AB - The B cell lymphoma WEHI231 has been used as a model for studying clonal deletion of B cells on the basis of its ability to undergo growth arrest and apoptosis by B cell antigen receptor (BCR) cross-linking. To comprehensively analyze the genes involved in BCR-mediated apoptosis, we applied the technique of serial analysis of gene expression (SAGE) to WEHI231. Comparison of expression patterns revealed that BCR cross-linking caused coordinate changes in the expression of genes involved in polyamine metabolism. Polyamines are ubiquitous compounds required for cell proliferation and homeostasis. The coordinate expression of the polyamine-related genes was confirmed by semiquantitative reverse transcriptase polymerase chain reaction analysis. During apoptosis, the genes involved in polyamine biosynthesis were downregulated, whereas those involved in polyamine catabolism were upregulated, suggesting that intracellular polyamines play a role in BCR-mediated apoptosis. Levels of intracellular putrescine, spermidine, and spermine were reduced after BCR cross-linking. These effects were prevented by concurrent CD40 stimulation, which blocked BCR-mediated apoptosis. Furthermore, addition of spermine could repress the BCR-mediated apoptosis by attenuating the mitochondrial membrane potential (Deltapsim) loss and activation of caspase-7 induced by BCR signaling. These findings strongly suggest that polyamine regulation is involved in apoptosis during B cell clonal deletion. PMID- 11281654 TI - BAG-1 p50 isoform interacts with the vitamin D receptor and its cellular overexpression inhibits the vitamin D pathway. AB - Human BAG-1 is an anti-apoptotic protein with four protein isoforms (BAG-1 p50, p46, p33, and p29). BAG-1 p46 was originally isolated in a screen for proteins binding to the glucocorticoid receptor; it binds and modulates the action of several members of the nuclear steroid hormone receptor superfamily. The vitamin D receptor (VDR) is another member of this superfamily, and the vitamin D pathway is important for prevention and therapy of osteoporosis, renal failure, cancer, and psoriasis. Therefore, we investigated the effect of the recently isolated BAG 1 p50 on the vitamin D pathway. By use of Far Western blot analysis and glutathione S-transferase BAG-1 p50 binding assays, BAG-1 p50 was demonstrated to interact with the VDR, and the BAG-1 p50 N-terminus was required. In U87 cells that were stably transfected with BAG-1 p50, binding of the VDR to its response element in electrophoretic mobility shift assays was blocked, enhancement of transcriptional activation was inhibited, cell growth rate was enhanced, cell growth inhibition induced by 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] was blocked, and 1,25(OH)2D3-mediated VDR induction was inhibited. These results suggest that BAG-1 p50 is a novel regulator of the vitamin D signaling pathway, and its overexpression may lead to cellular resistance to 1,25(OH)2D3 therapy. PMID- 11281657 TI - The histochemical structure of the deep fascia and its structural response to surgery. AB - The histochemical structure of the deep fascia and its interface with the underlying muscle was examined in ten pigs. This structure was also evaluated after it had been raised as a fascial flap and in another site after the underlying muscle surface had been disrupted. The deep fascial is a simple structure of densely-packed collagen bundles and elastin fibres, and has hyaluronic acid concentrated on its inner surface, which is in contact with the underlying muscle. There is no specialised lining of this surface of the fascia to account for its gliding properties. The post-surgical specimens demonstrated preservation of the structure of the interface between fascia and muscle, including the retention of the hyaluronic acid lining, if the epimysium was intact. However, if the epimysium was disrupted, the structure of the interface was obliterated. PMID- 11281658 TI - Vascular blood supply of the dorsal side of the thumb, first web and index finger: anatomical study. AB - Twenty-nine hands were dissected in order to define the dorsal blood supply of the thumb, the first web and the index finger. The main objective was to determine if it is possible to create a local osteocutaneous flap to cover partial and complex tissue defects in the distal part of the thumb. We found that the thumb metacarpal could be used as a donor site in these situations, with either a radiodorsal pedicle or an ulnadorsal pedicle. PMID- 11281659 TI - Replantation of very thumb distal amputations with pre-osteosynthesis interpositional vein graft. AB - Replantation of distal thumb amputations is difficult because of the anatomic orientation of the thumb and the location and size of vessels. A simplified technique is presented, in which interposition vein grafts are anastomosed to a terminal branch of the digital artery and a subcutaneous vein in the amputated thumb tip. This technique is convenient, relatively easy and reliable. PMID- 11281660 TI - Treatment of phalangeal fractures in severely injured hands. AB - This retrospective study was performed to assess the incidence of complications of operative treatment of phalangeal fractures. Risk factors for the development of complications were also investigated. Records and radiographs of 350 patients with 666 operatively treated phalangeal fractures were studied. Minimum follow-up was 1 year. A total of 176 fractured fingers were amputated primarily or secondarily, leaving 490 fractures for follow-up. Ninety-three fractures were treated conservatively. Nonunion necessitating reoperation developed in 6% (31/490) of fractures, malunion in 9% (44/490) and infection in 2% (8/490). Infection, segmental bone loss and (neuro)vascular injury predisposed to nonunion and replantation predisposed to malunion. There was a statistical correlation between the use of external fixation and malunion. Nonunion, malunion, and infection rates were similar to other studies. PMID- 11281661 TI - Phalangeal neck fractures in children: classification and outcome in 66 cases. AB - A series of 66 children with 67 phalangeal neck fractures in the hand is presented. Young children (1-3 years of age) made up 44% of the series. The mechanism of injury was entrapment of the digit in a closing door or a swing in almost all cases. Type I fractures (n = 13) were undisplaced and treatment with a splint resulted in excellent results in almost all cases. Type II fractures were defined as displaced fractures with some bone-to-bone contact. There were 47 Type II fractures and the outcome was significantly affected by the method of initial management. Fractures treated without K-wire fixation had a significantly worse outcome than those who underwent K-wire fixation. Type III fractures (n = 7) were displaced with no bone-to-bone contact. If inadequately treated, these fractures did not unite. PMID- 11281662 TI - Dynamic external fixation for pilon fractures of the interphalangeal joints. AB - Eight consecutive pilon fractures of the finger proximal interphalangeal joint and one of the interphalangeal joint of the thumb were treated by closed reduction and application of a new dynamic external fixator. The average range of movement achieved was 12 degrees -88 degrees and there were no serious complications. The technique described offers an effective and simple solution for treatment of pilon fractures of the interphalangeal joint. PMID- 11281663 TI - Hand blood supply in radial forearm flap donor extremities: a qualitative analysis using doppler examination. AB - This retrospective study evaluated the contribution of the forearm arteries to the blood supply of the hand after radial forearm flap surgery. Doppler ultrasound examinations of the radial, ulnar, anterior interosseous and posterior interosseous arteries were performed in the distal forearm using a continuous emission directional Doppler and a modified Allen test. Twenty-seven patients were included in this investigation which demonstrated a significant contribution of the anterior interosseous artery to hand vascularity after radial forearm flap surgery. PMID- 11281664 TI - Nerve repair: a neurobiologist's view. PMID- 11281665 TI - Surgical anatomy of spinal accessory nerve: is trapezius functional deficit inevitable after division of the nerve? AB - The course of spinal accessory nerve in the posterior triangle, the innervation of the sternocleidomastoid and trapezius muscles and the contributions from the cervical plexus were studied in 20 cadaveric dissections. The nerve was most vulnerable to iatrogenic injuries after leaving the sternocleidomastoid. Direct innervation of trapezius by cervical plexus branches was noted in five dissections, whereas connections between the cervical plexus and the spinal accessory nerve were observed in 19 dissections. These were usually under the sternocleidomastoid (proximal to the level of division of the nerve in nerve transfer procedures). Although the contribution from the cervical plexus to trapezius innervation is considered minimal, trapezius function can be protected in neurotization procedures by transecting the spinal accessory nerve distal to its branches to the upper position of trapezius. PMID- 11281666 TI - The ulnar nerve at the elbow and its local branching: an anatomic study. AB - Thirty nine cadaver elbows were dissected and the branching of the ulnar nerve, as well as the cubital tunnel and adjacent potential sites of nerve compression were studied. An arcade of Struthers was present in 26 specimens and Osborne's ligament was present in all specimens. A discrete flexor pronator aponeurosis overlying the ulnar nerve was present in 17 specimens. An average of one (range, 0-3) capsular nerve branches were noted. These originated an average 7 mm proximal (range, 45 mm proximal to 24 mm distal) to the medial epicondyle. An average of three (range, 1-6) motor branches to the flexor carpi ulnaris muscle were noted, and one of these originated proximal to the medial epicondyle in two specimens. Significant variation was noted in the capsular and motor branching of the ulnar nerve. Care must be taken to identify the motor branches of the ulnar nerve when performing a transposition. PMID- 11281667 TI - Trapeziometacarpal instability treated with modified Brunelli ligamentoplasty. AB - Seventeen cases of instability of the trapeziometacarpal joint were treated surgically using either the original Brunelli technique or one of two modifications. Six patients achieved very good, nine achieved good and two achieved poor results. The results of the three surgical techniques were similar and we recommend the use of the palmaris longus tendon for the ligament reconstruction, as it is the easiest to perform. PMID- 11281668 TI - Can the outcome of carpal tunnel release be predicted? AB - Between 1994 and 1996 we performed a prospective study on the effect of carpal tunnel release on the health status of 96 patients. The Nottingham Health Profile, a validated global scoring system, was used to assess quality of life before, and at 4 months after surgery. Carpal tunnel syndrome had a significant impact on the health status of our patients. There were significant improvements in the scores for pain, energy and sleep. Patients who were dissatisfied following surgery had significantly higher pre-operative scores, indicating poor perceived health status. Our findings show that outcome assessment tools have predictive value in identifying patients who may not benefit from surgery, or in whom a poor result might be anticipated. PMID- 11281669 TI - The sequelae of reflex sympathetic dystrophy. AB - This paper presents the results of a retrospective analysis of 94 patients who were assessed at a mean of 11 months after successful treatment of reflex sympathetic dystrophy (RSD) of the hand. Fifty-four percent still complained of pain related to the weather, and many complained of cold intolerance (44%), slight pain after use (34%), nail and hair growth changes (34%), sensory disturbances (34%) and stiffness of fingers in the morning (28%). There were also complaints of reduced finger extension, pain and loss of movement in the shoulder joint and hand swelling after use, and 78% of patients had significantly reduced grip strength. These results suggest that, in spite of resolution of the acute RSD problem, significant long term sequelae of RSD continue to impair function of the hand in a proportion of patients. PMID- 11281670 TI - Intraneural median nerve pressure in carpal tunnel syndrome. AB - In order to determine whether endoscopic carpal tunnel release decompresses the median nerve, we measured the intraneural median nerve pressure pre- and postoperatively in 55 hands. The median nerve pressure was significantly reduced postoperatively. PMID- 11281671 TI - Absorbable versus non-absorbable suture in carpal tunnel decompression. AB - This randomised prospective clinical study compared the use of an absorbable suture (subcuticular 4:0 polyglactin 910) and a non-absorbable suture (5:0 monofilament polypropylene) for elective carpal tunnel decompression wound closure. An increased perception of pain was reported by the patients in the polypropylene (Prolene) group. At the 6-week assessment, there was a higher level of residual wound inflammation in the polyglactin 910 (Vicryl) group. PMID- 11281672 TI - Adverse effect of porcine collagen interposition after trapeziectomy: a comparative study. AB - Twenty-six hands in 26 adults with osteoarthrosis of the thumb trapeziometacarpal joint were randomised to undergo either trapeziectomy alone (control) or with the interposition of porcine dermal collagen xenograft (Permacol). The study was terminated prematurely because of apparent reactions to the implants in six of 13 patients. The collagen interposition group required more frequent review on clinical grounds and were discharged later after surgery. Three of the implants have been removed and histology revealed foreign body reactions in all. There was no difference in thumb movement or power after surgery between the two groups. However, improved grip strength was observed and improved function were reported only in the control group. Permacol patients reported greater pain and were less satisfied with their operations than control patients. We conclude that interposition of Permacol is detrimental to the results of trapeziectomy. PMID- 11281673 TI - Treatment of congenital swan neck deformity with dynamic tenodesis of proximal interphalangeal joint. AB - Congenital swan neck deformities in seven fingers of two patients were treated by transfer of the flexor digitorum superficialis tendon to a tendon graft which was attached the extensor aponeurosis over the middle phalanx. The tendon transfer is protected for at least 2 months by a modified Murphy splint. PMID- 11281674 TI - Carpal tunnel syndrome due to an intraneural perineurioma in a 2-year-old child. AB - We report a 2-year-old girl with carpal tunnel syndrome due to a large intraneural perineurioma that required resection and nerve reconstruction. PMID- 11281675 TI - Dactylitis in a patient with brucellosis. AB - Dactylitis is an important feature of inflammatory arthritis and unusual complication of osteoarticular brucellosis. We report a case of dactylitis of the index finger in a female patient with brucellosis. PMID- 11281676 TI - Scoring system for brachial plexus palsy. PMID- 11281679 TI - Analysis of High-Resolution Spectra of (18)O(3). AB - Using a Fourier transform spectrometer, we have recorded the spectra of the (18)O(3) species of ozone in the region 1300-3100 cm(-1), with a resolution of 0.003 cm(-1). The large product pathlength x pressure enable us to record 18 bands, 14 for the first time. The analysis has been performed using effective Hamiltonians for polyads of strongly interacting states for ozone, accounting for Coriolis and anharmonic resonances. The spectral parameters are derived for 16 vibrational states, including the two "dark" states (040) and (130). Various resonances are studied through the mixing coefficients of rovibrational wavefunctions. Systematic intensity measurements allow determination of transition moment parameters for 16 bands. Finally, a complete list of all transitions from 1300 to 3100 cm(-1), with cutoffs 10(-26) cm(-1)/mol cm(-2) (296 K), is calculated. Copyright 2001 Academic Press. PMID- 11281680 TI - Analysis of High-Resolution Spectra of (18)O(3). AB - The analysis of high-resolution spectra of (18)O(3) is reported here for the range 3100-5000 cm(-1). Eight sets of polyads, corresponding to eight newly observed bands and accounting for resonance perturbations with eight other "dark" states, are analyzed for the first time. These analyses lead to eight sets of spectroscopic parameters, band centers, and rotational and centrifugal distortion constants, as well as transition moment parameters. In addition, various resonances are studied and compared to the similar ones for (16)O(3). This study allows us to check the work on (16)O(3) and to confirm that the accidentally strong high-order Deltav=8 anharmonic resonance for bands 5 nu(3)<- >3nu(1)+nu(2)+nu(3) was an exceptional case of the major isotopic species. Copyright 2001 Academic Press. PMID- 11281681 TI - Fourier Transform Emission Spectroscopy of CuCl. AB - The electronic spectra of CuCl were observed in the 18 000 cm(-1) to 25 000 cm( 1) spectral region using a Bruker IFS 120 HR Fourier transform spectrometer (FTS) and with the FTS associated with the McMath-Pierce Solar Telescope at Kitt Peak. On the basis of ab initio calculations, the labels for the electronic states were revised, and the a(3)Sigma(+)(1)-X(1)Sigma(+) 0-0 band, the b(3)Pi(0) X(1)Sigma(+) 0-0, 1-0, and 0-1 bands, the b(3)Pi(1)-X(1)Sigma(+) 0-0, 1-0, and 0 1 bands, the A(1)Pi-X(1)Sigma(+) 0-0, 1-0, and 0-2 bands, and the B(1)Sigma(+) X(1)Sigma(+) 0-0 and 1-0 bands were measured. Improved spectroscopic constants were obtained for the excited and ground states. Copyright 2001 Academic Press. PMID- 11281682 TI - Microwave Spectrum, Structure, and Hyperfine Constants of Kr-AgCl: Formation of a Weak Kr-Ag Covalent Bond. AB - The pure rotational spectrum of the complex Kr-AgCl has been measured between 8 15 GHz using a cavity pulsed-jet Fourier transform microwave spectrometer. The complex was found to be linear and relatively rigid, with a Kr-Ag bond length of approximately 2.641 A. The Kr-Ag stretching frequency was estimated to be 117 cm( 1). Ab initio calculations performed at the MP2 level of theory gave the geometry, vibration frequencies, Kr-Ag bond dissociation energy, and orbital populations. The Kr-Ag bond dissociation energy was estimated to be approximately 28 kJ mol(-1). The Kr-Ag force constant and dissociation energy are greater than those of Ar-Ag in Ar-AgCl. The chlorine nuclear quadrupole coupling constants show slight changes on complex formation. Ab initio orbital population analysis shows a small shift in sigma-electron density from Kr to Ag on complex formation. The combined experimental and ab initio results are consistent with the presence of a weak Kr-Ag covalent bond. Copyright 2001 Academic Press. PMID- 11281683 TI - Frequency Measurement of Pure Rotational Transitions of D(2)O from 0.5 to 5 THz. AB - Frequencies of pure rotational transitions of D(2)O were measured in the region 0.5-5 THz with a high-precision far-infrared spectrometer using a tunable radiation source. Measured frequencies of about 150 spectral lines, 30 of them being newly measured lines, provide an excellent frequency standard for the far infrared region together with our previous measurements on H(2)(16)O, H(2)(17)O, and H(2)(18)O. Molecular parameters of Watson's A-reduced Hamiltonian have been obtained to reproduce the observed frequencies. Copyright 2001 Academic Press. PMID- 11281684 TI - High-Temperature Infrared Emission Measurements on HNC. AB - The emission spectrum of HNC has been measured from 400 to 4100 cm(-1). The HNC was observed as an equilibrium mixture of HCN and HNC in a fused quartz cell heated to 1370 K. The three fundamental bands and many hot bands of HNC were measured with resolutions ranging from 0.006 cm(-1) for the lowest fundamental to 0.033 cm(-1) for the other two. High rotational levels up to J=62 were observed as well as vibrational levels up to v(2)=5. Now all the quadratic contributions to the vibrational and rotational term values have been determined, as well as some higher order terms. Copyright 2001 Academic Press. PMID- 11281685 TI - Improvement of the Spectroscopic Constants of the PF(A(3)Pi) State and Assignment of the PF(A-X) Transition Dipole Function. AB - High-resolution laser-excitation spectra were acquired for the v'=3-7 levels of the PF(A(3)Pi(0,1,2)<--X(3)Sigma(-)) transition from PF(X(3)Sigma(-)) molecules generated in a discharge flow reactor. These results were combined with lower resolution excitation spectra for the v'=8-11 levels and with existing high resolution data in the literature for v'=0 and 1 to assign improved spectroscopic constants for PF(A(3)Pi(0,1,2)). The abnormal vibrational energy level spacings for all spin components of the PF(A(3)Pi) state are evidence for a homogeneous interaction with another (3)Pi state. The Lambda-doublet separation in the PF((3)Pi(0)) substate increases with vibrational level, which is taken as evidence for interaction with the PF(b(1)Sigma(+)) state. Laser-induced fluorescence spectra from individual v'=0-5 levels were used to obtain vibrational band intensities of the A-X transition. The strong dependence of the transition dipole on the r-centroid is consistent with the reduction in the radiative lifetimes with increasing v' level. The similarity between the isovalent PF(A(3)Pi) and SO(A(3)Pi) states is noted and the bond dissociation energy of PF(X(3)Sigma(-)) is discussed. Copyright 2001 Academic Press. PMID- 11281686 TI - Fourier Transform Spectroscopy of the BaI Molecule: Simultaneous Analysis of Seven Electronic States Including the D(2)Sigma(+) and the G(2)Sigma(+) States. AB - In this work, the BaI D(2)Sigma(+) and G(2)Sigma(+) electronic states were investigated using laser-induced fluorescence (LIF) and Fourier transform spectroscopy (FTS). The LIF visible spectra were obtained by using the second harmonic of the Ti:sapphire single-mode laser and the Ar(+) and Kr(+) multimode lasers as excitation sources. Previously recorded data, taken from C. A. Leach, A. A. Tsekouras, and R. N. Zare (1992, J. Mol. Spectrosc. 153, 59-72) and from R. F. Gutterres, J. Verges, and C. Amiot (1999, J. Mol. Spectrosc. 196, 29-44; 2000, J. Mol. Spectrosc. 200, 253-260; and 2000, J. Mol. Spectrosc. 201, 326-327) were combined with the present data. Accurate and improved molecular constants for the X(2)Sigma(+), B(2)Sigma(+), A('2)Delta, A(2)Pi, C(2)Pi, and D(2)Sigma(+) states and 16 term values of the G(2)Sigma(+) state were derived from a simultaneous treatment of the whole data set (12 684 transitions) with a standard deviation of 3.26x10(-3) cm(-1). Copyright 2001 Academic Press. PMID- 11281687 TI - Jet-Cooled Multiphoton Ionization Spectroscopy of the iso-Butanal 3s<--n Rydberg Transition. AB - We report the results of a study on the vibrational structure in the (n, 3s) Rydberg state of iso-butanal, carried out via 2+1 multiphoton ionization spectroscopy in a supersonic expansion. Spectra were recorded for both the normal species, (CH(3))(2)CHCHO, and the deuterated isotopomer, (CH(3))(2)CDCHO. The spectra show a complicated vibrational pattern attributable to the gauche (C(1)) conformer which is known to be 250 cm(-1) lower in energy than the more symmetric trans (C(s)) conformer. The spectra are dominated by the low-frequency formyl torsion which appears in combination with the two methyl torsions (in-phase and out-of-phase) and with several skeletal modes. Vibrational assignments have been aided by ab initio calculations of the (n, 3s) Rydberg state. Copyright 2001 Academic Press. PMID- 11281688 TI - High-Lying Rotational Levels of Water: An Analysis of the Energy Levels of the Five First Vibrational States. AB - As a continuation of the work carried out on the ground and (010) vibrational states of water (R. Lanquetin, L. H. Coudert, and C. Camy-Peyret, 1999, J. Mol. Spectrosc. 195, 54-57), rotational energy levels for these two states are revisited here and new accurate rotational energy levels are considered for the three next vibrational states, that is, the (020), (100), and (001) states. Experimental rotational energies, along with their uncertainties, are retrieved through analyses of already published data sets and of discharge and flame emission spectra. The maximum value of J for the obtained levels is 25 for the ground state, 21 for the (010) state, and 20 for the three next states. Based on the bending-rotation Hamiltonian approach (L. H. Coudert, 1997, J. Mol. Spectrosc. 181, 246-273), a new theoretical approach is proposed to calculate rotational energies in the five interacting vibrational states under consideration and is used to carry out an analysis of the experimental energies. Comparisons with other existing energy level data sets are also presented. Copyright 2001 Academic Press. PMID- 11281689 TI - H(2)-Broadening Coefficients in the nu(4) Band of NH(3). AB - H(2)-broadening coefficients have been measured for 66 rovibrational lines of NH(3) at room temperature in the (P)P and (R)P branches of the nu(4) band in the range 1470-1600 cm(-1), using a high-resolution Fourier transform spectrometer. The collisional widths are obtained by fitting Voigt profiles to the measured shapes of the lines. The broadening coefficients are found to decrease on the whole as J increases and they increase with K for a given J value. The results are compared with those calculated from a semiclassical model in which the inversion vibration of NH(3) and collision-induced transitions with DeltaK = 0 and DeltaK = +/- 3 are taken into account. The intermolecular potential used includes electrostatic, induction, and dispersion energy contributions. The calculations performed by considering only DeltaK = 0 transitions provide significantly lower broadenings but with a satisfactory J and K dependence. The same trends are obtained for the broadening coefficients in inversion-rotation transitions and in the Q branch of the nu(1) parallel band of NH(3). Copyright 2001 Academic Press. PMID- 11281690 TI - Absolute Intensities of the nu(1) and nu(3) Bands of (16)O(3). AB - New experimental data on the nu(1) and nu(3) bands of (16)O(3) improving the value of absolute line intensities have been obtained. The intensities of 295 lines have been measured with an average accuracy between 2.5% and 3% and the rotational expansion of the transition moment operators for the nu(1) and nu(3) bands has been deduced. Finally, a complete listing of line intensities has been computed with an intensity cutoff of 1x10(-25) cm(-1)/molecule cm(-2). Copyright 2001 Academic Press. PMID- 11281691 TI - Carbon Monoxide Triplet Rydberg Series in the f Complex Region. AB - Using a multistep state-selective excitation scheme, fluorescence-dip spectra of carbon monoxide have been recorded in the energy region between 106 100 and 112 200 cm(-1). Prominent groups of bands were tentatively identified as transitions into the 6f em leader10f (v=0) and 9psigma em leader11psigma (v=0) triplet Rydberg series. For the f complex states, molecular constants have been calculated. The triplet f levels show an unusually large rotational constant and are located at higher energies than their singlet counterparts. Copyright 2001 Academic Press. PMID- 11281692 TI - Fourier Transform Infrared Spectra of CH(2)DF: The nu(5) and nu(6) Bands. AB - Results of a high-resolution infrared study of the spectroscopy of monodeuterated methyl fluoride, CH(2)DF, are reported for the first time. Spectra ranging from 500 to 3300 cm(-1) have been obtained and cover all the fundamental bands at resolutions down to 0.005 cm(-1). The two lowest energy fundamentals, the nu(5) and nu(6) bands, have been analyzed in detail. Since the molecule has C(s) symmetry, in principle both these bands are AB hybrids, since they belong to the irreducible representation A'. However, it was found that both are almost pure A type bands. A total of 597 A-type lines of the nu(5) band and 619 A-type lines of the nu(6) band have been assigned. Vibrational and rotational spectroscopic constants have been determined by least-squares fitting to the data. An improved band center for nu(7) is also reported. Copyright 2001 Academic Press. PMID- 11281693 TI - A Hund's Case (a) Analysis of the AB - We have recently observed a weak electronic subband near 513 nm in the electronic spectrum of cobalt monofluoride. A rotational analysis has led to its identification as a (3)Phi(3)-X(3)Phi(4) subband where DeltaSigma=-1 and DeltaOmega=-1. This critical datum has been used in combination with our previously published data (A. G. Adam et al., 1994, Chem. Phys. Lett. 230, 82) to obtain a Hund's case (a) analysis for the [18.8](3)Phi(i)-X(3)Phi(i) transition of CoF. The spectroscopic constants and electronic states of CoF are compared to those of CoH and Co(+). Two distinct excited (3)Phi electronic state vibrational progressions have also been identified in the CoF spectra. The band positions and rotational constants have been used to calculate equilibrium constants for the excited (3)Phi states. The two electronic transitions are identified as the K(3)Phi(i)-X(3)Phi(i) and L(3)Phi(i)-X(3)Phi(i) transitions based on comparisons with CoH. Copyright 2001 Academic Press. PMID- 11281694 TI - Absolute Line Intensities and Self-Broadening Coefficients in the nu(1) Band of (35)Cl(12)C(14)N. AB - The self-broadening coefficients and the intensities of 29 lines in the nu(1) band of cyanogen chloride ((35)Cl(12)C(14)N) have been measured at high resolution in the range 699-736 cm(-1), using a tunable diode-laser spectrometer. The collisional widths and most of the intensities are obtained by fitting Voigt and Rautian profiles to the measured shapes of the lines. From the analysis of the line intensities we determine the absolute strength as well as the Herman Wallis factors for the nu(1) band. A semiclassical calculation of the self broadening coefficients, performed by considering the main electrostatic interactions only, has provided larger results than the experimental data. Copyright 2001 Academic Press. PMID- 11281695 TI - Rovibrational and Rotational Spectroscopy of Levels of Propyne around 1000 cm( 1). AB - Four vibrational levels in the energy region around 1000 cm(-1) were studied. These were the v(5)=1 and v(8)=1 fundamental levels, both components of the v(9)=v(10)=1 combination level (l(9)=l(10)=+/-1 and l(9)=-l(10)=+/-1), and both components of the v(10)=3 overtone level (l(10)=+/-1 and +/-3). New FTIR spectra with a synchrotron radiation source were recorded in the region of the "superhot" v(10)=3<--2 bands, which made possible the first assignment of levels of the v(10)=3(+/-1) sublevel. More than 330 new rotational transitions in the combination and overtone levels were measured by millimeter-wave spectroscopy betwen 50 and 360 GHz. The new data were analyzed simultaneously together with the previously assigned rovibrational data for the fundamental and combination levels and rotational data for the fundamental levels using a global model with all anharmonic, Coriolis, l-type, and alpha-resonances. Significant improvement of data reproduction and very good consistency with the Hamiltonian parameters of the lower vibrational levels v(9)=1 and v(10)=1, 2 were achieved. A strong dependence of the A(v) constant on the l(10) quantum number is found for propyne: this is shown to be characteristic of skeleton C-C identical withC or C-C identical withN bending modes in H(3)CCCH, H(3)CCN, and their fully deuterated species. Copyright 2001 Academic Press. PMID- 11281696 TI - The Visible Emission Spectrum of the HgGa(2) Excimer. AB - A spectroscopic study of the Ga-Hg excimer has been carried out for the first time. The vapor mixture excited by a RF discharge shows emission bands with an intensity maximum at 5020 A. The density of Hg in this study was in the range of 10(18)-10(19) atoms/cc and that of Ga about 10(11) atoms/cc. High-resolution spectra with a dispersion of 1.2 A/mm were obtained, and vibrational structures were resolved and analyzed on the basis of a triatomic species HgGa(2). Copyright 2001 Academic Press. PMID- 11281697 TI - High-Resolution Laser Spectroscopy of YbCl: The B(2)Sigma(+)-X(2)Sigma(+) Transition. AB - High-resolution laser excitation spectra have been obtained for the 0-0, 1-1, and 0-1 bands of the B(2)Sigma(+)-X(2)Sigma(+) transition of YbCl and a rotational analysis has been performed on the (174)Yb(35)Cl and (172)Yb(35)Cl isotopomers. Comparison of the spin-rotation constant, gamma, for the B(2)Sigma(+) state with the lambda-doubling constant of the A(2)Pi(1/2) state (1) shows that the two excited states form a unique perturber pair arising from the 6psigma and 6ppi orbitals centered on the Yb(+) ion. The principal results for the B(2)Sigma(+) state are B(e)=0.097552(5) cm(-1), R(e)=2.43623(6) A, gamma(e)=-2.1655(6)x10(-4) cm(-1), and DeltaG(1/2)=313.111(2) cm(-1). Copyright 2001 Academic Press. PMID- 11281698 TI - Fourth-Order Rotational Corrections to the Effective Dipole Moments of Nonrigid Asymmetric Rotors. AB - An expression for the fourth-order rotational correction terms of the effective dipole moments of nonrigid asymmetric rotors has been derived using the method of contact transformation. The treatment takes into account the large-amplitude bending motion. The correction terms have been calculated for the different bending states of the H(2)O molecule. A poor convergence of the rotational series for this molecule has been obtained and some different nonpolynomial forms for these series, obtained by different summation methods, have been proposed and tested. Copyright 2001 Academic Press. PMID- 11281699 TI - High-Resolution Spectroscopy and Analysis of the nu(3) and nu(4) Fundamentals of Monoisotopic (70)GeF(4). AB - The first high-resolution study on germanium tetrafluoride is reported. We used a monoisotopic sample of (70)GeF(4). The FTIR spectra of the two infrared active fundamentals, namely the nu(4) (bending) and nu(3) (stretching) modes, were recorded at a temperature of ca. 210 K and a resolution (1/maximum optical path difference) of 0.0031 and 0.0023 cm(-1), respectively. These spectra were analyzed using the STDS software developed in Dijon. In both cases, we obtained a fit with a root mean square better than 1x10(-3) cm(-1). Both bands show very regular structures with no detectable perturbation. Copyright 2001 Academic Press. PMID- 11281700 TI - Experimental and Theoretical Study of the Electronic States and Spectra of TeH and TeLi. AB - Gas-phase emission spectra of the hitherto unknown free radical TeLi have been measured in the NIR range with a Fourier-transform spectrometer. The emissions were observed from a fast flow system in which tellurium vapor in argon carrier gas was passed through a microwave discharge and mixed with lithium vapor in an observation tube. Two systems of blue-degraded bands were measured at high spectral resolution in the ranges 8000-9000 and 5700-6700 cm(-1) and vibrational and rotational analyses were performed. In order to aid in the analysis of the experimental data, a series of relativistic configuration interaction calculations has been carried out to obtain potential curves for the low-lying states of TeLi and the isovalent TeH and also electric dipole transition moments connecting them. As in the TeH system, the ground state of TeLi is found to be X(2)Pi(i), but with a remarkably smaller spin-orbit splitting. The TeLi calculations indicate a strongly bound A(2)Sigma(+) state, while in TeH the analogous state is computed to lie significantly higher at approximately 32 000 cm(-1), and it is strongly predissociated. Based on the theoretical analysis, the observed TeLi band systems are assigned to the transitions A(2)Sigma(+)(A1/2)- >X(1)(2)Pi(3/2)(X(1)3/2) and A(2)Sigma(+)(A1/2)-->X(2)(2)Pi(1/2)(X(2)1/2). Analysis of the spectra has yielded the molecular constants (in cm(-1)) X(1)(2)Pi(3/2):omega(e)=457.49(3), omega(e)x(e)=2.482(9), B(0)=0.408908(8); X(2)(2)Pi(1/2): T(e)=2353.44(3), omega(e)=456.28(4), omega(e)x(e)=2.635(8), B(0)=0.414954(8), p(0)=1.00637(4); A(2)Sigma(+): T(e)=8574.64(2), omega(e)=437.81(3), omega(e)x(e)=2.581(8), B(0)=0.423903(8), p(0)=-0.19915(2), where the numbers in parentheses are the standard deviations of the parameters. Comparison of the isovalent TeLi and TeH systems emphasizes that the difference in bonding character (ionic in TeLi vs covalent in TeH) is responsible for qualitative differences in the electronic spectra of these two molecules. Copyright 2001 Academic Press. PMID- 11281701 TI - Laser-Induced Fluorescence Spectrum of the Orbitally Forbidden &Btilde;'(2)Delta &Xtilde;(2)Sigma(+) Transition of SrCCH. PMID- 11281702 TI - Plasmodium falciparum: immunogenicity of alum-adsorbed clinical-grade TBV25-28, a yeast-secreted malaria transmission-blocking vaccine candidate. AB - Gozar, M. M. G., Muratova, O., Keister, D. B., Kensil, C. R., Price, V. L., and Kaslow, D. C. 2001. Plasmodium falciparum: Immunogenicity of alum-adsorbed clinical-grade TBV25-28, a yeast-secreted malaria transmission-blocking vaccine candidate. Experimental Parasitology 97, 61-69. The fusion of Pfs25 and Pfs28, two major surface antigens on zygotes and ookinetes of Plasmodium falciparum, as a single recombinant protein (TBV25-28) was previously shown to elicit potent transmission-blocking antibodies in mice. Clinical-grade TBV25-28 was subsequently manufactured and its potency was evaluated in rabbits. Rabbits received three doses of either clinical-grade TBV25H or clinical-grade TBV25-28 adsorbed to alum with or without QS-21. As measured in a standard membrane feeding assay, addition of QS-21 to the formulations appeared to enhance transmission-blocking potency of rabbit sera after two vaccinations but not after three vaccinations. Surprisingly, TBV25H elicited more potent transmission blocking antibodies than did TBV25-28, a result strikingly different from those of previous mouse experiments using research-grade TBV25-28. The apparent decrease in potency of clinical-grade TBV25-28 in rabbits appears to reflect an enhancement in potency of clinical-grade TBV25H in a new formulation rather than simply a species difference in immunogenicity of TBV25-28. PMID- 11281703 TI - Ancylostoma caninum: the finger cell neurons mediate thermotactic behavior by infective larvae of the dog hookworm. AB - Bhopale, V. M., Kupprion, E. K., Ashton, F. T., Boston, R., and Schad, G. A. 2001. Ancylostoma caninum: The finger cell neurons mediate thermotactic behavior by infective larvae of the dog hookworm. Experimental Parasitology 97, 70-76. In the amphids (anteriorly positioned, paired sensilla) of the free-living nematode Caenorhabditis elegans, the so-called finger cells (AFD), a pair of neurons, each of which ends in a cluster of microvilli-like projections, are known to be the primary thermoreceptors. A similar neuron pair in the amphids of the parasitic nematode Haemonchus contortus is also known to be thermoreceptive. The hookworm of dogs, Ancylostoma caninum, has apparent structural homologs of finger cells in its amphids. The neuroanatomy of the amphids of A. caninum and H. contortus is strikingly similar, and the amphidial cell bodies in the lateral ganglia of the latter nematode have been identified and mapped. When the lateral ganglia of first-stage larvae (L1) of A. caninum are examined with differential interference contrast microscopy, positional homologs of the recognized amphidial cell bodies in the lateral ganglia of H. contortus L1 are readily identified in A. caninum. The amphidial neurons in A. caninum were consequently given the same names as those of their apparent homologs in H. contortus. It was hypothesized that the finger cell neurons (AFD) might mediate thermotaxis by the skin-penetrating infective larvae (L3) of A. caninum. Laser microbeam ablation experiments with A. caninum were conducted, using the H. contortus L1 neuronal map as a guide. A. caninum L1 were anesthetized and the paired AFD class neurons were ablated. The larvae were then cultured to L3 and assayed for thermotaxis on a thermal gradient. L3 with ablated AFD-class neuron pairs showed significantly reduced thermotaxis compared to control groups. The thermoreceptive function of the AFD class neurons associates this neuron pair with the host-finding process of the A. caninum infective larva and shows functional homology with the neurons of class AFD in C. elegans and in H. contortus. PMID- 11281705 TI - Tick-Theileria interactions in response to immune activation of the vector. AB - Watt, D. M., Walker, A. R., Lamza, K. A., and Ambrose, N. C. 2001. Tick-Theileria interactions in response to immune activation of the vector. Experimental Parasitology 97, 89-94. Immune mechanisms towards the haemoprotozoan parasite Theileria parva were investigated in their tick vector, Rhipicephalus appendiculatus. The exoskeletons of adult ticks were initially pierced with bacteria-coated, saline-coated, or sterile dry glass needles. Haemolymph was extracted from the ticks at 6, 24, 48, and 72 h postinjection and applied to bacterial plates to measure the growth inhibition effects. The inhibition zones were larger with all the injected groups compared to uninjected controls. The largest inhibition zones were seen 24 h after injection with bacteria-coated needles. An experiment was carried out to investigate whether antibacterial immune responses were relevant to the parasite/tick relationship and, if so, which parasite form was most vulnerable. R. appendiculatus nymphs were infected with T. parva by feeding on an infected calf and were then injected with needles on days 7, 13, 15, and 17 throughout their moult in an attempt to induce tick immune responses at the same time as different lifecycle forms of T. parva would be present. Salivary glands from the moulted adult ticks in the control and different treatment groups were dissected out and examined for the presence of T. parva sporoblasts. No difference in infection levels was seen in any of the treatment groups compared with the controls, suggesting that immune responses in R. appendiculatus, induced by bacterial injection, do not affect T. parva infections. The fecundity of injected ticks was compared with that of uninjected controls to ensure that the injection procedure itself was not detrimental to the ticks. Injected females had higher engorgement masses than controls but reduced levels of egg hatching. PMID- 11281704 TI - Toxoplasma gondii: molecular cloning and characterization of a novel 18-kDa secretory antigen, TgMIC10. AB - Hoff, E. F., Cook, S. H., Sherman, G. D., Harper, J. M., Ferguson, D. J. P., Dubremetz, J. F., and Carruthers, V. B. 2001. Toxoplasma gondii: Molecular cloning and characterization of a novel 18-kDa secretory antigen, TgMIC10. Experimental Parasitology, 97, 77-88. During host cell invasion, Toxoplasma gondii secretes proteins from specialized organelles (micronemes and rhoptries) located at the apical end of the parasite. The contents of the micronemes appear to be crucial to T. gondii invasion, as inhibition of microneme secretion prevents parasite entry into host cells. Here we describe a new T. gondii microneme protein, TgMIC10. Molecular characterization of a full-length TgMIC10 cDNA revealed that TgMIC10 lacks homology to any previously characterized proteins, although a homologue, NcMIC10, was identified in a closely related parasite, Neospora caninum. TgMIC10 has an unusually long secretory leader sequence of 58 amino acids; the mature TgMIC10 is 18 kDa, possesses nine diglutamic acid repeats and an imperfect repeat sequence (RK(R/Y)HEEL), and is entirely devoid of cysteines. Antibodies raised against recombinant TgMIC10 recognized the native TgMIC10 and localized the protein to the micronemes in indirect immunofluorescence and immunoEM experiments. Comparison of immunofluorescence images indicates that TgMIC10 expression is higher in T. gondii tachyzoites, which are responsible for active infection, than in bradyzoites, which are responsible for latent infection. PMID- 11281706 TI - Effects of dietary polyamine deficiency on Trypanosoma gambiense infection in rats. AB - Nishimura, K., Araki, N., Ohnishi, Y., and Kozaki, S. 2001. Effects of dietary polyamine deficiency on Trypanosoma gambiense infection in rats. Experimental Parasitology 97, 95-101. A diet deficient in polyamines decreases the availability of dietary polyamines. We used rats infected with the Wellcome strain of Trypanosoma gambiense to examine the effects of polyamine-deficient chow (PDC) on trypanosome proliferation and symptoms of infection. Rats fed PDC showed limited increase of trypanosome and symptoms of infection and limited loss of body weight and anemia. Survival in these rats was prolonged. Before infection, the heparinized plasma concentration of spermidine in the PDC-fed rats was lower than that in control rats fed with standard chow. After infection, the content of spermidine in red blood cells increased in the control rats, but was only slightly increased in PDC-fed rats. The content of spermidine in the trypanosomes after infection was low in the PDC-fed rats. Decreases in the polyamine content of trypanosomes limited their increase. These observations suggest that a reduction in dietary polyamines may help in the regulation of trypanosome infection. PMID- 11281708 TI - Biomagnetic separation of contaminating host leukocytes from plasmodium-infected erythrocytes. AB - Carlton, J. M-R., Yowell, C. A., Sturrock, K. A., and Dame, J. B. 2001. Biomagnetic separation of contaminating host leukocytes from Plasmodium-infected erythrocytes. Experimental Parasitology 97, 111-114. PMID- 11281707 TI - Loss of multiple hydrogenosomal proteins associated with organelle metabolism and high-level drug resistance in trichomonads. AB - Land, K. M., Clemens, D. L., and Johnson, P. J. 2001. Loss of multiple hydrogenosomal proteins associated with organelle metabolism and high-level drug resistance in trichomonads. Experimental Parasitology 97, 102-110. In trichomonads, metronidazole is activated to its cytotoxic form in a specialized energy-producing organelle called the hydrogenosome. Electron transport components in the organelle, pyruvate:ferredoxin oxidoreductase and ferredoxin, donate a single electron to the drug, converting it to a cytotoxic free radical. Previous biochemical analyses of enzyme activities of highly resistant strains of both Trichomonas vaginalis and Tritrichomonas foetus reveal undetectable activity for pyruvate:ferredoxin oxidoreductase and another hydrogenosomal enzyme, hydrogenase. We have chosen to analyze a highly drug-resistant strain of T. foetus and its parental drug-sensitive strain from which it was derived to study the molecular basis for these enzyme defects. Quantitation of pyruvate:ferredoxin oxidoreductase and ferredoxin levels in sensitive and resistant cells shows a marked reduction of these proteins in the resistant strain. RNA analysis reveals an approximately 60% reduction in pyruvate:ferredoxin oxidoreductase mRNA and 90 98% reduction in mRNA levels encoding hydrogenosomal proteins hydrogenase, ferredoxin, and malic enzyme. We have measured the levels of transcription of these genes and observed 60% reduction of pyruvate:ferredoxin oxidoreductase gene transcription and 85% reduction in malic enzyme gene transcription in the resistant strain. The reduction or absence of these organellar proteins is likely to reduce or eliminate the ability of the cell to activate the drug, giving rise to the highly resistant phenotype. Ultrastructural analysis of thin sections revealed that resistant cells are 20% smaller in size and hydrogenosomes in resistant cells are approximately one-third the size of those in the drug sensitive parental strain. These data suggest that altered gene expression of multiple hydrogenosomal proteins results in the modification of the organelle and leads to drug resistance. PMID- 11281709 TI - Plasmodium falciparum: gelatin enrichment selects for parasites with full-length chromosome 2. implications for cytoadhesion assays. AB - Waterkeyn, J. G., Cowman, A. F., and Cooke, B. M. 2001. Plasmodium falciparum: Gelatin enrichment selects for parasites with full-length chromosome 2. Implications for cytoadhesion assays. Experimental Parasitology 97, 115-118. PMID- 11281711 TI - Gag-derived proteins of HIV-1 isolates from Indian patients: cloning, expression, and purification of p17 of B- and C-subtypes. AB - A simple and efficient method for expression in Escherichia coli and purification of matrix protein, p17, of human immunodeficiency virus type 1 (HIV-1) of both B- and C-subtypes is described. DNA sequences encoding p17 of B- and C-subtype were cloned from respective gag sequences. The gag sequences were obtained by PCR amplification using DNA extracted from peripheral blood lymphocytes of an HIV-1 infected patient from India. A T7-promoter-based expression system was optimized for expression of p17 in soluble form. p17 (B- and C-subtype) was purified to near homogeneity using conventional chromatographic techniques. Purification of p17 (C-subtype) is described for the first time with yield of 7.7 mg from a 1 liter culture. The yield of p17 (B-subtype) is 14.7 mg from a 1-liter culture, which is severalfold better than that reported earlier. N-terminal sequencing and CD spectra of the purified proteins, p17B and p17C, show that the proteins are properly processed and well-folded. The immunoreactivity of both types of p17 to sera from HIV-infected individuals is comparable. PMID- 11281710 TI - Solution studies of recombinant human stromal-cell-derived factor-1. AB - Stromal-cell-derived factor-1 (SDF-1alpha) is an 8-kDa chemokine that is constitutively expressed in bone-marrow-derived stromal cells and has been identified as a ligand for the CXCR4 receptor. We produced the chemokine recombinantly as methionine-SDF-1alpha in Escherichia coli without the leader peptide sequence. The protein was denatured, refolded, and further purified by reversed-phase HPLC. SDF-1alpha was shown to be >95% pure as judged by SDS-PAGE. The final yield of purified and refolded SDF-1alpha was 1-2 mg per gram of wet cell paste. The refolded protein is a ligand for the CXCR4 receptor and has been used to block HIV-mediated cell fusion and downmodulates the CXCR4 receptor. Our ability to purify hundreds of milligrams of refolded protein allowed us to conduct detailed studies of the biophysical properties of the protein. We have used a combination of biophysical techniques to study the solution properties of SDF-1alpha. The average mass of SDF-1alpha, as determined by static light scattering, gave us the first indications that the chemokine may self-associate. Further investigation with sedimentation velocity ultracentrifugation confirmed the existence of two species. The measured s(20, W) values defined two masses corresponding to monomer and dimer. Finally, sedimentation equilibrium ultracentrifugation and dynamic light scattering yielded a composite value of 150 +/- 30 microM for the dimerization constant. We conclude that SDF-1alpha exists in a monomer-dimer equilibrium. PMID- 11281712 TI - Expression in Pichia pastoris of Candida antarctica lipase B and lipase B fused to a cellulose-binding domain. AB - Candida antarctica lipase B (CALB) and C. antarctica lipase B fused to a cellulose-binding domain (CBD-CALB) were expressed functionally in the methylotrophic yeast Pichia pastoris. The cellulose-binding domain originates from cellulase A of the anaerobic rumen fungus Neocallimastix patriciarum. The genes were fused to the alpha-factor secretion signal sequence of Saccharomyces cerevisiae and placed under the control of the alcohol oxidase gene (AOX1) promoter. The recombinant proteins were secreted into the culture medium reaching levels of approximately 25 mg/L. The proteins were purified using hydrophobic interaction chromatography and gel filtration with an overall yield of 69%. Results from endoglycosidase H digestion of the proteins showed that CALB and CBD CALB were N-glycosylated. The specific hydrolytic activities of recombinant CALB and CBD-CALB were identical to that reported for CALB isolated from its native source. The fusion of the CBD to the lipase resulted in a greatly enhanced binding toward cellulose for CBD-CALB compared with that for CALB. PMID- 11281713 TI - The cell wall and cell division gene cluster in the Mra operon of Pseudomonas aeruginosa: cloning, production, and purification of active enzymes. AB - We have cloned the Pseudomonas aeruginosa cell wall biosynthesis and cell division gene cluster that corresponds to the mra operon in the 2-min region of the Escherichia coli chromosome. The organization of the two chromosomal regions in P. aeruginosa and E. coli is remarkably similar with the following gene order: pbp3/pbpB, murE, murF, mraY, murD, ftsW, murG, murC, ddlB, ftsQ, ftsA, ftsZ, and envA/LpxC. All of the above P. aeruginosa genes are transcribed from the same strand of DNA with very small, if any, intragenic regions, indicating that these genes may constitute a single operon. All five amino acid ligases, MurC, MurD, MurE, MurF, and DdlB, in addition to MurG and MraY were cloned in expression vectors. The four recombinant P. aeruginosa Mur ligases, MurC, MurD, MurE, and MurF were overproduced in E. coli and purified as active enzymes. PMID- 11281714 TI - Expression, purification, and biophysical characterization of the BRCT domain of human DNA ligase IIIalpha. AB - The C-terminal regions of several DNA repair and cell cycle checkpoint proteins are homologous to the breast-cancer-associated BRCA-1 protein C-terminal region. These regions, known as BRCT domains, have been found to mediate important protein-protein interactions. We produced the BRCT domain of DNA ligase IIIalpha (L3[86]) for biophysical and structural characterization. A glutathione S transferase (GST) fusion with the L3[86] domain (residues 837-922 of ligase IIIalpha) was expressed in Escherichia coli and purified by glutathione affinity chromatography. The GST fusion protein was removed by thrombin digestion and further purification steps. Using this method, (15)N-labeled and (13)C/(15)N double-labeled L3[86] proteins were prepared to enable a full determination of structure and dynamics using heteronuclear NMR spectroscopy. To obtain evidence of binding activity to the distal BRCT of the repair protein XRCC1 (X1BRCTb), as well as to provide insight into the interaction between these two BRCT binding partners, the corresponding BRCT heterocomplexes were also prepared and studied. Changes in the secondary structures (amount of helix and sheet components) of the two constituents were not observed upon complex formation. However, the melting temperature of the complex was significantly higher relative to the values obtained for the L3[86] or X1BRCTb proteins alone. This increased thermostability imparted by the interaction between the two BRCT domains may explain why cells require XRCC1 to maintain ligase IIIalpha activity. PMID- 11281715 TI - Expression, purification, and crystallization of the RGS-like domain from the Rho nucleotide exchange factor, PDZ-RhoGEF, using the surface entropy reduction approach. AB - Lsc-homology domains are found in several eukaryotic nucleotide exchange factors which act on Rho-family GTPases. They show limited amino acid sequence similarity to RGS proteins, which down-regulate the cellular signaling by the alpha-subunits of trimeric G-proteins and have been shown to interact with Galpha12 and Galpha13. It is believed that the RGS-like (RGSL) domain constitutes the functional link between G-protein-coupled receptors and cytosolic Rho-GTPases. We report here the expression, purification, and crystallization of the RGSL domain from the PDZ-RhoGEF. To obtain X-ray-grade crystals we have used the recently proposed approach of crystallization by mutational surface entropy reduction, in which selected Lys --> Ala, Glu --> Ala, and/or combined point mutations are introduced into the target protein to reduce the cumulative conformational entropy of surface residues. Of the five mutants that were designed and prepared, the second one tried (K463A, E465A, E466A) yielded crystals suitable for further analysis and diffracted X-rays to 2.8 A resolution on a home source. The crystals exhibit hexagonal symmetry, space group P6(1) 22 or P6(5) 22, with unit cell parameters a = b = 63.1 A, c = 202.1 A, and contain one molecule in the asymmetric unit. PMID- 11281716 TI - A family 2a carbohydrate-binding module suitable as an affinity tag for proteins produced in Pichia pastoris. AB - The family 2a carbohydrate-binding module (CBM), Cel5ACBM2a, from the C-terminus of Cel5A from Cellulomonas fimi, and Xyn10ACBM2a, the family 2a CBM from the C terminus of Xyn10A from C. fimi, were compared as fusion partners for proteins produced in the methylotrophic yeast Pichia pastoris. Gene fusions of murine stem cell factor (SCF) with both CBMs were expressed in P. pastoris. The secreted SCF Xyn10ACBM2a polypeptides were highly glycosylated and bound poorly to cellulose. In contrast, fusion of SCF to Cel5ACBM2a, which lacks potential N-linked glycosylation sites, resulted in the production of polypeptides which bound tightly to cellulose. Cloning and expression of these CBM2a in P. pastoris without a fusion partner confirmed that N-linked glycosylation at several sites was responsible for the poor cellulose binding. The nonglycosylated CBMs produced in E. coli had very similar cellulose-binding properties. PMID- 11281717 TI - Production of recombinant bovine lactoferrin N-lobe in insect cells and its antimicrobial activity. AB - Lactoferrin is a multifunctional, iron-binding glycoprotein found in physiological fluids of mammals. In the present study, a gene encoding the N terminal half (N-lobe) of bovine lactoferrin was cloned and expressed in cultured insect cells using a baculovirus expression system. One mutation was found in the lactoferrin N-lobe gene, but it resulted in no amino acid substitution. The recombinant lactoferrin N-lobe was secreted into the culture medium and partially purified by means of an immobilized heparin column. The recombinant lactoferrin N lobe secreted was not glycosylated, but it possessed antimicrobial activity toward Escherichia coli O111. The recombinant product synthesized and accumulated in the host cells exhibited greater electrophoretic mobility on SDS-PAGE than the secreted product and showed no potency to inhibit the growth of bacteria. It is thought that the product accumulated intracellularly lacks antimicrobial ability due to its degradation in the host cells or due to disruption of the active conformation. PMID- 11281718 TI - Overexpression and purification of Rhizobium etli glutaminase A by recombinant and conventional procedures. AB - Rhizobium etli glutaminase A was purified to homogeneity by conventional procedures that included ammonium sulfate differential precipitation, ion exchange chromatography, hydrophobic interaction chromatography, gel filtration, and dye-ligand chromatography. Alternatively, the structural glsA gene that codifies for glutaminase A was amplified by PCR and cloned in the expression vector pTrcHis. The recombinant protein was purified to homogeneity by affinity chromatography. This protein showed the same kinetic properties as native glutaminase A (K(m) for glutamine of 1.5 mM and V(max) of 80 micromol ammonium min(-1) mg protein(-1)). Physicochemical and biochemical properties of native and recombinant glutaminase were identical. The molecular mass of recombinant glutaminase A (M(r) 106.8 kDa) and the molecular mass of the subunits (M(r) 26.9 kDa) were estimated by mass spectrometry. These results suggest that R. etli glutaminase A is composed of four identical subunits. The high-level production of recombinant glutaminase A elevates the possibilities for determination of its three-dimensional structure through X-ray crystallography. PMID- 11281719 TI - Low-temperature increases the yield of biologically active herring antifreeze protein in Pichia pastoris. AB - Antifreeze proteins and antifreeze glycoproteins are structurally diverse molecules that share a common property in binding to ice crystals and inhibiting ice crystal growth. Type II fish antifreeze protein of Atlantic herring (Clupea harengus harengus) is unique in its requirement of Ca(2+) for antifreeze activity. In this study, we utilized the secretion vector pGAPZalpha A to express recombinant herring antifreeze protein (WT) and a fusion protein with a C terminal six-histidine tag (WT-6H) in yeast Pichia pastoris wild-type strain X-33 or protease-deficient strain SMD1168H. Both recombinant proteins were secreted into the culture medium and properly folded and functioned as the native herring antifreeze protein. Furthermore, our studies demonstrated that expression at a lower temperature increased the yield of the recombinant protein dramatically, which might be due to the enhanced protein folding pathway, as well as increased cell viability at lower temperature. These data suggested that P. pastoris is a useful system for the production of soluble and biologically active herring antifreeze protein required for structural and functional studies. PMID- 11281720 TI - Design and expression of polymeric immunoglobulin fusion proteins: a strategy for targeting low-affinity Fcgamma receptors. AB - We have developed a family of cloning vectors that direct expression of fusion proteins that mimic aggregated immunoglobulin (IgG) (AIG) and immune complex function with respect to their interactions with FcgammaR and that allow for the inclusion and targeting of a second protein domain to cells expressing FcgammaR. This was accomplished by expressing multiple linear copies of the hinge and CH2 domains (HCH2) of human IgG(1) fused to the framework region of human IgG(1). Convenient restriction sites allow for the facile introduction of additional amino-terminal domains. The resulting molecule is tripartite. The carboxyl-IgG(1) framework domain provides stability and permits dimerization, and the intervening polymer provides increased effector function and targeting to FcgammaR while the amino-terminal domain can deliver an additional signal to cells expressing FcgammaR. To demonstrate the utility of the vectors, the extracellular domain of human CD8alpha was expressed as a HCH2 polymer fusion protein. The fusion proteins were secreted in useful amounts from polyclonal cell lines established in Sf9 cells following transfection and selection with G418. The biological activity of the recombinant CD8alpha-HCH2 polymers was determined and compared to those of AIG and an anti-CD16 monoclonal antibody using an in vitro assay. The activity of the fusion proteins positively correlates to the number of HCH2 units. The largest polymer tested was severalfold more potent than AIG at similar concentrations. The HCH2 polymers described here represent a new strategy in the design of recombinant proteins for the therapeutic targeting of FcgammaR in autoimmune disorders. PMID- 11281721 TI - Cell-free production of biologically active polypeptides: application to the synthesis of antibacterial peptide cecropin. AB - An approach to preparative production of polypeptides, including uneasily testable, degradable, and toxic ones, is proposed on the basis of in vitro expression systems of last generation, such as continuous-exchange cell-free and continuous-flow cell-free transcription-translation systems. The approach implies that a polypeptide of interest is synthesized as a fusion protein with the polypeptide linked to green fluorescent protein (GFP) through a cleavable spacer. The GFP moiety provides direct visualization and quantitative monitoring of the polypeptide synthesis, as well as solubility and stability of the product. The synthesis of functionally active antibacterial polypeptide cecropin P1 (31 amino acid residues) has been demonstrated. PMID- 11281722 TI - The cochaperone murine stress-inducible protein 1: overexpression, purification, and characterization. AB - Murine stress-inducible protein 1 (mSTI1) is a cochaperone that is homologous with the human heat shock cognate protein 70 (Hsc70)/heat shock protein 90 (Hsp90)-organizing protein (Hop). To analyze the biochemical properties of mSTI1 and the stoichiometry of the Hsc70.mSTI1.Hsp90 association, recombinant mSTI1 was produced in untagged, histidine (His)-tagged, and glutathione S-transferase (GST) tagged forms. His-mSTI1 was detected either as a dimer during size-exclusion-high performance liquid chromatography (SE-HPLC) or as a monomer during Superdex 200 gel filtration chromatography. SE-HPLC on GST-mSTI1 and untagged mSTI1 suggested that mSTI1 existed as a monomer. Cross-linking of His-mSTI1 detected a compact monomeric species and a dimeric species. Gel filtration on the association of bovine STI1 or His-mSTI1 with Hsc70 detected species of molecular mass consistent with a dimeric STI1 species or a 1:1 complex of STI1 and Hsc70. Our data and that of others suggest that mSTI1 and its homologues exist as either a monomer or a dimer and that this facilitates its proposed function as an Hsc70/Hsp90 organizing protein. PMID- 11281723 TI - Identification, cloning, and expression of a functional phenylalanyl-tRNA synthetase (pheRS) from Staphylococcus aureus. AB - Phenylalanyl-tRNA synthetase (pheRS) is unique among aminoacyl tRNA synthetases in that it is a heterotetrameric enzyme composed of two alpha-subunits and two larger beta-subunits. In prokaryotes, the alpha- and beta-subunits of pheRS are encoded by the genes pheS and pheT, respectively. In this report we describe the isolation of a DNA fragment (3.52 kb) containing the pheS and pheT genes from a Staphylococcus aureus (WCUH29) genomic DNA library. Both genes, found as a part of transcriptional operon, were predicted to encode polypeptides which showed strong primary and structural similarity to prokaryotic phenylalanyl-tRNA synthetase alpha- and beta- subunits. We describe the high-level overexpression and purification of recombinant S. aureus pheRS using pheS and pheT genes as part of an artificial operon in Escherichia coli. For comparative analysis we also report a procedure for the purification of native pheRS from S. aureus (Oxford Strain) and demonstrate that Michaelis-Menten parameters for both recombinant and native enzyme, at least for phenylalanine tRNA aminoacylation are comparable. PMID- 11281724 TI - Expression of cloned cDNA for the human mitochondrial RNA polymerase in Escherichia coli and purification. AB - A full-length cDNA for the mature, mitochondrial form of human mitochondrial RNA polymerase was cloned and expressed under the control of T5 or tac promoter in Escherichia coli. The cDNA was efficiently expressed at 37 degrees C, but virtually all the polymerase produced was insoluble, and renaturation of the inclusion bodies was unsuccessful. When the cells were grown at 25 degrees C, however, a portion of approximately 10% was soluble and active. The protein was purified 100-fold from the soluble lysates to homogeneity by two-step chromatography using Ni-nitrilotriacetic acid-Sepharose and heparin-agarose columns, as an N-terminal histidine tag attached and as the tag cleaved away. The purified polymerases with and without the histidine tag were both active in RNA polymerization in vitro as measured with poly(dA-dT) template, and specific activity was 140,000 units/mg. The purified enzyme has the same biochemical properties as the polymerase fraction partially purified from the human mitochondria, except for the promoter-specific activity that was not observed with the purified polymerase in the presence of mitochondrial transcription factor A. Additional factor(s) and/or mammalian-specific or regulatory modification(s) of the polymerase should be necessary for promoter-specific transcription. PMID- 11281725 TI - Expression and purification of recombinant mouse Ets-1 transcription factor. AB - Ets-1 is a transcription factor which belongs to the ETS family. Its mRNA is expressed in the embryo during normal development and also in tumors. In order to sort out functional Ets-1-binding sites among those present in gene promoters, we constructed an expression vector and designed a purification protocol for the production of the 440-amino-acid form of mouse Ets-1, based on heparin-Sepharose affinity and anion-exchange chromatographies. This protocol allows the purification of large amounts of pure recombinant protein as assessed by SDS PAGE, C18 reverse-phase HPLC, amino-terminal sequencing, and mass spectrometry. The purified protein is recognized by specific anti-Ets-1 antibodies and binds to DNA ETS-binding sites. PMID- 11281726 TI - Heterologous expression of soluble, active proteins in Escherichia coli: the human estrogen receptor hormone-binding domain as paradigm. AB - The human estrogen receptor ligand-binding domain (hER-E/F), including the distal F domain, has been expressed to high levels in a soluble, active form in Escherichia coli in order to facilitate biophysical studies. The ability of a series of vectors incorporating strong transcriptional and translational signals to provide an efficient expression system for hER-E/F was investigated. High level expression was obtained from all of the vectors used in the study. Although the majority of hER-E/F protein was produced in insoluble form under standard bacterial culture conditions, hER-E/F could be produced in soluble, biologically active form by altering the sequence of the expressed protein and by varying the host culture conditions. Several parameters, including the presence of a His tag, growth temperature, and addition of ethanol and 17beta-estradiol to the growth medium were shown to have a positive effect on production of soluble hER-E/F. An optimized expression system capable of producing from 25 to 35 mg of biologically active hER-E/F in 1 liter of cell culture was designed, and a simple, rapid purification protocol for hER-E/F produced in this system was developed. Characterization of purified hER-E/F by Edman degradation and mass spectrometry verified the identity of the protein. The K(D) for 17beta-estradiol binding to purified hER-E/F was determined to be 0.6 +/- 0.1 nM. The parameters controlling soluble, heterologous protein production observed in this study may be generally applicable to the expression of other heterologous proteins in E. coli. PMID- 11281727 TI - Interactions between phosphatidylinositol 3-kinase and nitric oxide: explaining the paradox. AB - Nitric oxide (NO) and the many derivatives and reactive oxygen intermediates thereof are all molecules that are utilised by mammalian cells in the war against microbial pathogens and tumours. They are potentially toxic molecules and, with damage control being crucial, the production and metabolism of nitric oxide is a tightly regulated process. The duality of NO is well documented. On the one hand, beneficial effects include normal healing in the skin and intestinal mucosa, killing of certain bacteria, regulating T cell proliferation and differentiation (Th1 vs Th2), and regulating leukocyte recruitment, by affecting adhesion molecule expression. On the other hand, persistent high levels of NO can lead to the production of toxic metabolites (peroxynitrite and hydroxyls), which can have detrimental effects, such as increased microvascular and epithelial permeability, increased oxidative stress (which can damage DNA), and damage to iron-sulphur proteins in mitochondria. NO has been reported to modulate its own production and the mechanisms involved in this self-regulation are being hotly pursued. The purpose of this review is to update recent intriguing advances in our understanding of the interaction of the phosphatidylinositol (PI) 3-kinase dependent signal transduction pathway in regulating the activity of the enzymes that generate NO, namely, the nitric oxide synthases. PMID- 11281728 TI - Leptin signaling in human peripheral blood mononuclear cells, activation of p38 and p42/44 mitogen-activated protein (MAP) kinase and p70 S6 kinase. AB - The adipocyte-derived hormone leptin plays an important role as a relayer of nutritional status to several organ systems. Evidence is accumulating that leptin plays an important role in the adequate functioning and maintenance of the immune system. Here we show that leptin induces sustained phosphorylation of p38 MAP kinase in human peripheral blood mononuclear cells (PBMCs). We show furthermore that leptin induces two routes to phosphorylation of the 40S ribosomal protein S6, one is activation of the p90 ribosomal S6 kinase (RSK) via the MEK/p42/p44 MAP kinase pathway, the other is via activation of p70 S6 kinase. Thus, these results give new insight in the mechanism that underlies the immunomodulatory effects of leptin. PMID- 11281729 TI - Distinct mechanisms of inhibition of interleukin-6-induced Stat3 signaling by TGF beta and alpha-thrombin in CCL39 cells. AB - We previously demonstrated that exposure of CCL39 lung fibroblast cells to alpha thrombin inhibits interleukin-6 (IL-6)-induced tyrosine phosphorylation of Stat3 (signal transducers and activators of transcription-3) protein via a mitogen activated protein (MAP)-kinase dependent mechanism. In the present study, we investigated the mechanism of regulation of IL-6-induced signaling by transforming growth factor-beta (TGF-beta) and compared this to alpha-thrombin mediated inhibition. We demonstrate that exposure of CCL39 cells to TGF-beta completely inhibits IL-6-induced Stat3 tyrosine phosphorylation and gp130 gene expression. However, in contrast to alpha-thrombin, TGF-beta-mediated inhibition did not require activation of the MAP kinase pathway. Also, unlike alpha thrombin, TGF-beta-mediated inhibition requires synthesis of new proteins. Interestingly, TGF-beta and alpha-thrombin both inhibit IL-6-induced expression of gp130 mRNA levels. These results demonstrate that although the end effects are the same, alpha-thrombin and TGF-beta utilize distinct mechanisms to inhibit IL-6 induced Stat3 signaling. PMID- 11281730 TI - PD98059 attenuates hydrogen peroxide-induced cell death through inhibition of Jun N-Terminal Kinase in HT29 cells. AB - We have investigated the effects of hydrogen peroxide (H(2)O(2)), a potent naturally occurring oxidant on cell signaling and viability in the pluripotent HT29 intestinal cell line. There was a dose-dependent reduction in cell viability upon exposure to H(2)O(2) as measured by the XTT assay. Features of apoptosis were indicated by the findings of PARP and caspase 3 cleavage, as well as changes in cell morphology using phase contrast and nuclear fragmentation using fluorescence microscopy. There was a dose-dependent increase in the activation of p45-JNK, p42/p44-ERK, and p38-HOG. Surprisingly, oxidant-induced cell injury could be attenuated by preincubation with PD98059 to 50% of untreated control cells (P = 0.002). This and UO126, another MEK inhibitor were ably to reproducibly inhibit p45-JNK activation induced by hydrogen peroxide. Transfection with kinase-inactive constructs of JNK and ERK revealed that the improvement in cell viability was due to inhibition of JNK and not ERK. Transient transfections with AP-1 and NF-kappaB luciferase reporter constructs did not reveal any transcriptional activation due to hydrogen peroxide exposure however, in both cases the basal levels of transcriptional activity were suppressed in the presence of PD98059. It is concluded that JNK mediates H(2)O(2)-induced cellular injury in the HT29 cell line, and additionally, we report for the first time that JNK activation can be inhibited by both PD98059 and UO126 at conventional doses used to inhibit MEK. PMID- 11281731 TI - IGF1 activates PKC alpha-dependent protein synthesis in adult rat cardiomyocytes. AB - Acute exposure to interleukin 1 beta (IL1beta) or insulin-like growth factor 1 (IGF1) promoted the translocation of PKC alpha from the cytosol to the membrane of adult rat cardiomyocytes. Western analysis demonstrated that membranal localization of PKC alpha was increased from 23% in the control to 49% after exposure to IGF1, and it was increased to 42% after exposure to IL1beta. Activation of Erk1/Erk2 by IGF1 and IL1beta was studied using a phosphorylation specific antibody. IGF1-induced activation of p44/p42 MAP kinase was blocked by preincubation with the PKC inhibitors, bisindolylmaleimide and Go6976, as well as the tyrosine kinase inhibitor, genistein. IGF1 increased the rate of protein synthesis, indicated by the increase in L-[(14)C(U)] phenylalanine incorporation over time, and this effect was inhibited by Go6976. PMID- 11281732 TI - Using neural networks for prediction of subcellular location of prokaryotic and eukaryotic proteins. AB - T. Kohonen's self-organization model, a typical neural network model, was applied to predict the subcellular location of proteins from their amino acid composition. The Reinhardt and Hubbard database was used to examine the performance of the neural network method. The rates of correct prediction for the three possible subcellular location of prokaryotic proteins were 96.1% by the self-consistency test and 84.4% by the jackknife test. The rates of correct prediction for the four possible subcellular location of eukaryotic proteins were 95.6% by the self-consistency test and 70.6% by the jackknife test. PMID- 11281733 TI - Comparative effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on MCF-7, RL95-2, and LNCaP cells: role of target steroid hormones in cellular responsiveness to CYP1A1 induction. AB - A study was conducted to investigate whether target hormones affect 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD)-inducible gene expression, using as an experimental model system three human cancer cell lines, breast (MCF-7), uterine (RL95-2), and prostate (LNCaP). Exposure to TCDD induced the CYP1A1 gene in all three cell lines. MCF-7 and RL95-2 cells showed more than 15- and 10-fold induction of EROD (7-ethoxyresorufin O-deethylase) activity, respectively, compared with the less responsive LNCaP cells. Surprisingly, however, TCDD induced reporter gene activity driven by a single XRE element was similar in RL95 2 and LNCaP cells. The steady-state levels of expression of aryl hydrocarbon receptor (AhR) and aryl hydrocarbon receptor nuclear translocator (ARNT) were similar in all three cell lines. Expression of the CYP1B1 and PAI-2 genes was induced by TCDD in MCF-7 and RL95-2, but not in LNCaP, cells. Transient coexpression of estradiol receptor-alpha (ER-alpha) with a TCDD-responsive reporter plasmid and subsequent TCDD treatment increased responsiveness to TCDD in RL95-2 and LNCaP cells. Treatment with AZA-C, a DNA methyltransferase inhibitor, enhanced responsiveness to TCDD, in terms of EROD activity in LNCaP cells, but not in MCF-7 and RL95-2 cells, suggesting that DNA methylation in the CpG dinucleotide within the XRE core sequence is another factor involved in silencing of CYP1A1 in LNCaP cells. TCDD markedly inhibited E(2)- or testosterone induced reporter gene activities in all three cell lines. Conversely, these target hormones inhibited TCDD-induced EROD activity in the three cell lines. These findings suggest that TCDD and the target steroid hormones negatively regulate each other's activity. PMID- 11281734 TI - Overexpression of protein kinase C delta represses expression of proliferin in NIH3T3 cells that regulates cell proliferation. AB - Protein kinase C (PKC) delta is known to inhibit proliferation of many cell types. In this study we found that overexpression of PKCdelta reduced proliferation of NIH3T3 cells. To identify specific genes regulated by PKCdelta in regulation of cell proliferation, we used differential display-polymerase chain reaction in PKCdelta-overexpressing NIH3T3 cells and found that the expression of proliferin, a secreted protein known to stimulate cell proliferation, was significantly repressed. Transient transfection study indicated that the repression of proliferin expression was inversely proportional to the expression levels of PKCdelta. Addition of an anti-proliferin antibody to culture medium to neutralize the secreted proliferin decreased cell proliferation in a dose-dependent manner. Our results, therefore, suggest that overexpression of PKCdelta induces transcriptional repression of proliferin, thereby resulting in inhibition of cell proliferation. PMID- 11281735 TI - Cloning and expression of the mouse PIP49 (Pancreatitis Induced Protein 49) mRNA which encodes a new putative transmembrane protein activated in the pancreas with acute pancreatitis. AB - We have used a microarray-based strategy to characterize, at the molecular level, the pancreatic emergency program set up by the pancreatic cells in response to pancreatitis. In this strategy, the phenotype of the pancreatitis-affected pancreas is established by characterization of a large number of its transcripts using a high-density mouse cDNA microarray. This method allows identification of transcripts differentially expressed during pancreatitis. We describe here the cloning, sequencing, and expression analysis of a new gene, named PIP49 (Pancreatitis Induced Protein 49). Its very strong expression is specific of acinar cells and occurs rapidly after initiation of the acute phase of pancreatitis. Analysis of its primary and secondary structures strongly suggests that PIP49 encodes a putative transmembrane protein. PMID- 11281736 TI - The association of bone marrow lesions with pain in knee osteoarthritis. AB - BACKGROUND: The cause of pain in osteoarthritis is unknown. Bone has pain fibers, and marrow lesions, which are thought to represent edema, have been noted in osteoarthritis. OBJECTIVE: To determine whether bone marrow lesions on magnetic resonance imaging (MRI) are associated with pain in knee osteoarthritis. DESIGN: Cross-sectional observational study. SETTING: Veterans Affairs Medical Center. PATIENTS: 401 persons (mean age, 66.8 years) with knee osteoarthritis on radiography who were drawn from clinics in the Veterans Administration health care system and from the community. Of these persons, 351 had knee pain and 50 had no knee pain. MEASUREMENTS: Knee radiography and MRI of one knee were performed in all participants. Those with knee pain quantified the severity of their pain. On MRI, coronal T(2)-weighted fat-saturated images were used to score the size of bone marrow lesions, and each knee was characterized as having any lesion or any large lesion. The prevalence of lesions and large lesions in persons with and without knee pain was compared; in participants with knee pain, the presence of lesions was correlated with severity of pain. RESULTS: Bone marrow lesions were found in 272 of 351 (77.5%) persons with painful knees compared with 15 of 50 (30%) persons with no knee pain (P < 0.001). Large lesions were present almost exclusively in persons with knee pain (35.9% vs. 2%; P < 0.001). After adjustment for severity of radiographic disease, effusion, age, and sex, lesions and large lesions remained associated with the occurrence of knee pain. Among persons with knee pain, bone marrow lesions were not associated with pain severity. CONCLUSIONS: Bone marrow lesions on MRI are strongly associated with the presence of pain in knee osteoarthritis. PMID- 11281737 TI - Beta-blockers in congestive heart failure. A Bayesian meta-analysis. AB - PURPOSE: Congestive heart failure is an important cause of patient morbidity and mortality. Although several randomized clinical trials have compared beta blockers with placebo for treatment of congestive heart failure, a meta-analysis quantifying the effect on mortality and morbidity has not been performed recently. DATA SOURCES: The MEDLINE, Cochrane, and Web of Science electronic databases were searched from 1966 to July 2000. References were also identified from bibliographies of pertinent articles. STUDY SELECTION: All randomized clinical trials of beta-blockers versus placebo in chronic stable congestive heart failure were included. DATA EXTRACTION: A specified protocol was followed to extract data on patient characteristics, beta-blocker used, overall mortality, hospitalizations for congestive heart failure, and study quality. DATA SYNTHESIS: A hierarchical random-effects model was used to synthesize the results. A total of 22 trials involving 10 135 patients were identified. There were 624 deaths among 4862 patients randomly assigned to placebo and 444 deaths among 5273 patients assigned to beta-blocker therapy. In these groups, 754 and 540 patients, respectively, required hospitalization for congestive heart failure. The probability that beta-blocker therapy reduced total mortality and hospitalizations for congestive heart failure was almost 100%. The best estimates of these advantages are 3.8 lives saved and 4 fewer hospitalizations per 100 patients treated in the first year after therapy. The probability that these benefits are clinically significant (>2 lives saved or >2 fewer hospitalizations per 100 patients treated) is 99%. Both selective and nonselective agents produced these salutary effects. The results are robust to any reasonable publication bias. CONCLUSIONS: beta-Blocker therapy is associated with clinically meaningful reductions in mortality and morbidity in patients with stable congestive heart failure and should be routinely offered to all patients similar to those included in trials. PMID- 11281739 TI - Confounding by contraindication in a nationwide cohort study of risk for death in patients taking ibopamine. AB - BACKGROUND: Outcomes may differ in treated and untreated patients because of a contraindication for treatment in the latter that is independently associated with the outcome of interest. OBJECTIVE: To evaluate the effects of confounding by contraindication on risk factors for death in patients taking ibopamine after its use was restricted in early September 1995. DESIGN: Retrospective cohort study. SETTING: The Netherlands. PATIENTS: 1146 patients with congestive heart failure who were prescribed ibopamine at least once and for whom medication history and medical data were available. MEASUREMENTS: Cardiovascular risk factors, clinical characteristics, and medication use. Each patient was assigned an index date (the date of death, or a random date for patients still alive at the end of the study). RESULTS: In univariate analyses comparing patients with an index date before and those with an index date after 8 September 1995, the relative risk for death associated with current use of ibopamine was 3.02 (95% CI, 2.12 to 4.30) compared with 0.71 (CI, 0.53 to 0.96), respectively. In multivariate analyses, the risk for death was 2.62 (CI, 1.76 to 3.90) and 0.93 (CI, 0.84 to 1.02), respectively. CONCLUSION: The marked inversion of the relative risk estimate can be considered a practical example of confounding by contraindication. PMID- 11281740 TI - Changes in DNA methylation in neoplasia: pathophysiology and therapeutic implications. AB - Methylation of DNA is a biochemical modification that can influence gene expression and is involved in inactivating one of the two X chromosomes in women. Evidence that has accumulated in the past 10 years suggests that cancer cells usurp this physiologic mechanism and use it to their benefit by inactivating tumor suppressor genes and related proteins. However, the primary structure of the affected proteins remains intact; reversal of abnormalities in DNA methylation may therefore restore the tumor-suppressive function of these genes and provide a novel approach to cancer therapy. Two demethylating drugs, 5 azacytidine and 5-aza-deoxycytidine, are currently being tested in clinical trials, and several others are in preclinical development. In this article, the biological rationale for targeting aberrant methylation in cancer therapy is reviewed and completed phase I and II trials of this approach, some of which show promise for treatment of hematologic malignancies, are summarized. PMID- 11281741 TI - Pseudoaccountability. AB - The public has long entrusted the medical profession to regulate its own practices, but our efforts to do so have been uneven. In place of rigorous, enforceable standards, we have sometimes reverted to pseudoaccountability: weak regulations that only give the appearance that we have been responsible in setting and enforcing high standards. We have failed to deal effectively with substandard practitioners, lagged in preventing medical errors, inadequately documented patient care in the medical record, slipped up in protecting patients in clinical research projects, and accepted financial arrangements with industry that may affect our judgment. In response, government has intervened, and threatens to intervene further with regulations that sometimes are excessively intrusive and cumbersome. To preserve our remaining autonomy, we must show that we are serious about protecting the public. Professional membership organizations must stop promulgating weak guidelines and offering evaluative methods to assess and regulate their own coveted members. Instead, they must turn over assessment of their members to arm's-length, disinterested groups. Regulatory programs must have provisions with impeccable standards, surveillance processes, and methods to deal with infractions. In planning new self-regulatory approaches, we should invite open discussion and genuine involvement by independent members of the public or the government to help ensure that our decisions about standard setting are not self-serving. Our profession has already lost much authority. We can ill afford to lose more. PMID- 11281742 TI - Edema of the bone marrow can cause pain in osteoarthritis and other diseases of bone and joints. PMID- 11281744 TI - Evidence base for management of acute exacerbations of chronic obstructive pulmonary disease. PMID- 11281746 TI - Nobility. PMID- 11281747 TI - Fathers, doctors, and time. PMID- 11281745 TI - Management of acute exacerbations of chronic obstructive pulmonary disease: a summary and appraisal of published evidence. AB - PURPOSE: To review critically the available data on diagnostic evaluation, risk stratification, and therapeutic management of patients with acute exacerbations of chronic obstructive pulmonary disease (COPD). DATA SOURCES: English-language articles were identified by searching MEDLINE (1966 to 2000, week 5), EMBASE (1974 to 2000, week 18), HealthStar (1975 to June 2000), and the Cochrane Controlled Trials Register (2000, Issue 1). STUDY SELECTION: The best available evidence on each subtopic was selected for analysis. Randomized trials, sometimes buttressed by cohort studies, were used to evaluate therapeutic interventions. Cohort studies were used to evaluate diagnostic tests and risk stratification. DATA EXTRACTION: Study design and results were summarized in evidence tables. Individual studies were rated by internal validity, external validity, and quality of design. Statistical analyses of combined data were not performed. DATA SYNTHESIS: Data on the utility of most diagnostic tests are limited. However, chest radiography and arterial blood gas sampling seem useful while acute spirometry does not. Identifiable clinical variables are associated with risk for relapse and risk for death after hospitalization for an acute exacerbation. Evidence of efficacy was found for bronchodilators, corticosteroids, and noninvasive positive-pressure ventilation. There is also support for the use of antibiotics in patients with more severe exacerbations. On the basis of limited data, mucolytics and chest physiotherapy do not seem to be of benefit, and oxygen supplementation seems to increase the risk for respiratory failure only in an identifiable subgroup of patients. CONCLUSIONS: Although suggestions for appropriate management can be made on the basis of available evidence, the supporting literature is scarce and further high-quality research is necessary. Such research will require an improved, generally acceptable, and transportable definition of acute exacerbation of COPD, as well as improved methods for observing and measuring outcomes. PMID- 11281748 TI - Adherence to protease inhibitors. PMID- 11281750 TI - Cost-utility analysis. PMID- 11281752 TI - Giant-cell arteritis of the female genital tract. PMID- 11281755 TI - Synthesis and characterization of coronanes: multicyclopropane-fused macrocyclic arrays. AB - Stepwise macrocyclization of the all syn-trans-1,15-quinquecyclopropanedimethanol (4) with iso- and terephthaloyl chlorides and 4,4'-methanediyl-dibenzoic acid (28) gave the corresponding coronanes 22, 23, and 32. The same protocol was used with all syn-trans-1,21-septecyclopropanedimethanol (5) and 2,3 naphthalenedicarboxylic acid to obtain the macrolide 27. Direct macrocyclization of diol 4 and 1,10-phenanthroline-2,9-dicarbonyl chloride (33) and 2,2' bipyridine-4,4'-dicarbonyl chloride (35) gave the coronanes 34 and 36, respectively. Ring closing metathesis (RCM) of the diene 42 using Cl2(Cy3P)2Ru=CHPh (48) (Grubbs's catalyst) gave the macrocyclic lactone 45. The structures of coronanes 22, 23, 32, 34, 36, and 45 were confirmed by X-ray crystallographic studies which showed the cyclopropyl chain to adopt very differing conformations throughout the series. Several of the macrocycles have significant free pathways through their ring centers, and in the case of compound 34 there is a water molecule hydrogen bonded within the ring. This latter compound has the potential to act as a chiral ligand to metal centers. PMID- 11281756 TI - Photoreaction of 2-halo-N-pyridinylbenzamide: intramolecular cyclization mechanism of phenyl radical assisted with n-complexation of chlorine radical. AB - The photochemical behavior of 2-halo-N-pyridinylbenzamide (1-4 in Chart 1) was studied. The photoreaction of 2-chloro-N-pyridinylbenzamides 1a, 2a, 3a, and 4 afforded photocyclized products, benzo[c]naphthyridinones (6-9 and 16), in high yield, whereas the bromo analogues 1b, 2b, and 3b produced extensively photoreduced products, N-pyridinylbenzamides (1c, 10, and 11), with minor photocyclized product. Since the photocyclization reaction of 2-chloro-N pyridinylbenzamide is retarded by the presence of oxygen and sensitized by the presence of a triplet sensitizer, acetone or acetophenone, a triplet state of the chloro analogue is involved in the reaction. Since several radical intermediates, particularly n-complexes of chlorine radical, are identified in the laser flash photolysis of 2-chloro-N-pyridinylbenzamide, an intramolecular cyclization mechanism of phenyl radical assisted with n-complexation of chlorine radical for the cyclization reaction is proposed: the triplet state (78 kcal/mol) of the chloro analogue (1a), which is populated by the excitation of 1a undergoes a homolytic cleavage of the C-Cl bond to give phenyl and chlorine radicals; while chlorine radical holds the neighbor pyridinyl ring with its n-complexation, the intramolecular arylation of the phenyl radical with the pyridinyl ring proceeds to produce a conjugated 2,3-dihydropyridinyl radical and then the conjugated radical aromatizes to afford a cyclized product, benzo[c]naphthyridinone by ejecting a hydrogen. The photoreduction product can be formed by hydrogen atom abstraction of the phenyl sigma radical from the environment. PMID- 11281757 TI - Synthesis, chemical properties, and biological evaluation of CC-1065 and duocarmycin analogues incorporating the 5-methoxycarbonyl-1,2,9,9a tetrahydrocyclopropa. AB - The synthesis of 5-methoxycarbonyl-1,2,9,9a-tetrahydrocyclopropa[c]benz[e]indol-4 one (C5-CO2Me-CBI), a substituted CBI derivative bearing a C5 methoxycarbonyl group, and its corresponding 5-hydroxymethyl derivative are described in efforts to establish substituent electronic effects on the agents' functional reactivity and the resulting effect this has on their rate of DNA alkylation. Resolution of an immediate C5-CO2Me-CBI precursor and its incorporation into both enantiomers of 16 and 17, analogues of the duocarmycins, are also detailed. A study of the solvolysis reactivity and regioselectivity of N-BOC-C5-CO2Me-CBI (12) revealed that the introduction of a C5 methyl ester modestly slowed the rate of solvolysis (1.8x, pH 3) without altering the inherent reaction regioselectivity (>20:1). The comparison of the X-ray structures of the N-CO2Me derivatives of C5-CO2Me-CBI and CBI revealed correlations with the reaction regioselectivity and the relative reactivity of the compounds. The latter correlated well with the less reactive C5 CO2Me-CBI exhibiting a shortened N2-C2a bond length (1.386 vs 1.390 A) and smaller chi1 dihedral angle (8.1 degrees vs 21.2 degrees ) indicative of greater vinylogous amide conjugation and was accompanied by a diminished (cross conjugated) cyclopropane conjugation (shorter bond lengths). Establishment of the DNA alkyation properties revealed that C5-CO2Me-CBI-based agents retained the identical alkylation selectivity of the natural products. More importantly, the C5 methyl ester was found to decrease the rate (0.77x) of DNA alkylation relative to CBI, consistent with its inherent lower reactivity. These results indicate that the previously observed increase in the rate of DNA alkylation for C7 substituted CBI analogues including CCBI (7-cyano-CBI) is contrary to expectations based on their inherent reactivities. Unlike 17, in which the C5 methyl ester does not bind in the minor groove, the C7 substituent lies in the minor groove extending the rigid length of the agents, further enhancing the DNA binding-induced conformational change responsible for activation toward nucleophilic attack and catalysis of the DNA alkylation reaction. PMID- 11281758 TI - Conformational analysis of indole alkaloids corynantheine and dihydrocorynantheine by dynamic 1H NMR spectroscopy and computational methods: steric effects of ethyl vs vinyl group. AB - 1H NMR (400 MHz) spectra of the indole alkaloid dihydrocorynantheine recorded at room temperature show the presence of two conformers near coalescence. Low temperature 1H NMR allowed characterization of the conformational equilibrium, which involves rotation of the 3-methoxypropenoate side chain. Line-shape analysis yielded enthalpy of activation DeltaH(double dagger) = 71 +/- 6 kJ/mol, and entropy of activation DeltaS(double dagger) = 33 +/- 6 J/mol.K. The major and minor conformation contains the methyl ether group above and below the plane of the ring, respectively, as determined by low-temperature NOESY spectra, with free energy difference DeltaG degrees = 1.1 kJ/mol at -40 degrees C. In contrast to dihydrocorynantheine, the corresponding rotamers of corynantheine are in the fast exchange limit at room temperature. The activation parameters determined for corynantheine were DeltaH(double dagger) = 60 +/- 6 kJ/mol and DeltaS(double dagger) = 24 +/- 6 J/mol.K, with DeltaG degrees = 1.3 kJ/mol at -45 degrees C. The difference in the exchange rates of the rotamers of corynantheine and dihydrocorynantheine (respectively, 350 s(-1) and 9 s(-1) at 0 degrees C) reflects the difference in the steric bulk of the vinyl and the ethyl group. The conformational equilibria involving the side chain rotation as well as inversion of the bridgehead nitrogen in corynantheine and dihydrocorynantheine was studied by force-field (Amber and MMFF) and ab initio (density-functional theory at the B3LYP/6-31G level) computational methods, the results of which were in good agreement with the 1H NMR data. However, the calculations identified the rotamers as essentially isoenergetic, the experimental energy differences being to small to be reproduced exactly by the theory. Comparison of density-functional and force-field calculations with experimental results identified Amber as giving the most accurate results in the present case. PMID- 11281759 TI - Efficient synthesis of gamma-alkylidenetetronic esters by sequential Lewis acid catalyzed. AB - A new approach for the synthesis of gamma-alkylidenetetronic acids and esters is reported which involves Me3SiOTf-catalyzed, regio- and stereoselective cyclization of 4-alkoxy-1,3-bis(trimethylsilyloxy)-1,3-butadienes with oxalyl chloride. The alpha-hydroxy group of the butenolides is efficiently functionalized by palladium-catalyzed cross-coupling reactions via the corresponding enol triflates. PMID- 11281760 TI - An unusually acidic methyl group directly bound to acridinium cation. AB - 9,10-Dimethylacridinium chloride (1: X = Cl) exhibited strong acidity of pH 3.90 (3.4 x 10(-3) M, 20 degrees C) in an aqueous solution. H-D exchange reaction of 1 in D2O indicated that protons in the 9-methyl group dissociated to generated H+ ions. This is a unique example of a methyl proton functioning as an acid. The acidity derives from the wider pi face in acridinium capable of delocalizing the newly formed negative charge upon proton dissociation. PM3 calculations provided stabilization factors (deltaDeltaH(f)o) between proton dissociated and undissociated forms of several N-heterocycles and also confirmed the acidity observed in acridinium. PMID- 11281761 TI - N(1)-C(5')-linked dimer hydrates of 5-substituted uracils produced by anodic oxidation in aqueous solution. AB - Electrochemical dimerization reactivity has been studied for 5-substituted uracils (5XU) including thymine (1a: X = Me) and 5-halouracil derivatives (1b: X = F; 1c: X = Cl; 1d: X = Br; 1e: X = I). Upon galvanostatic electrolysis of Ar saturated aqueous solution 1a underwent anodic oxidation to produce N(1)-C(5')- and N(1)-C(6')-linked dimer hydrates, 1-(6'-hydroxy-5',6'-dihydrothymin-5' yl)thymine (5a) and 1-(5'-hydroxy-5',6'-dihydrothymin-6'-yl)thymine (6a), as the major products. These N-C-linked dimerizations were accompanied by the formation of novel stereoisomeric C(5)-C(5')-linked dimers (meso isomer: 13a[meso]; racemic isomer: 13a[rac]) with a condensed tetrahydrofuran ring skeleton. Similar electrolyses of 5-fluorouracil (1b) and 5-chlorouracil (1c) also afforded the corresponding N(1)-C(5')-linked dimer hydrates, 1-(5'-fluoro-6'-hydroxy-5',6' dihydrouracil-5'-yl)-5-fluorouracil (5b) and 1-(5'-chloro-6'-hydroxy-5',6' dihydrouracil-5'-yl)-5-chlorouracil (5c), respectively, while resulting in neither N(1)-C(6')-linked dimer analogues nor C(5)-C(5')-linked dimers, unlike the reactivity of 1a. In contrast to 1a-c, no dimeric products were obtained from 5-bromouracil (1d) and 5-iodouracil (1e). The present electrochemical method was applicable to the cross-dimerization into N(1)-C(5')-linked heterodimer hydrates composed of binary 5-substituted uracils that occurred in competition with the formation of homodimer hydrates. A mechanism of the N(1)-C(5')-linked dimerization of 1a-c has been proposed, by which allyl-type radical intermediates with limiting mesomeric forms of N(1)-centered and C(5)-centered pyrimidine radicals (2a-c [N(1)]/2a-c [C(5)]) are generated via anodic one-electron oxidation and subsequent deprotonation at N(1) and undergo a head-to-tail coupling. PMID- 11281762 TI - The preparation of a new "safety catch" ester linker for solid-phase synthesis. AB - A new "safety catch" linker for esters has been synthesized on polystyrene resin. This 2-tert-butoxyphenol resin 10 may be acylated to give a relatively stable ester that will allow nucleophilic chemistry without reaction at the linking ester group. Removal of the tert-butyl group with acid unmasks a highly reactive 2-hydroxyphenyl ester that reacts readily with nucleophiles to cause release of the product from the resin. This sequence has been exemplified by acylating the resin with various bromo acids, carrying out nucleophilic displacements with thiols, phenols, or amines, activating the ester with trifluoroacetic acid and cleaving from the resin with amines to give the (nucleophile) substituted carboxamides in high yield and purity. Kinetic studies with a model ester revealed half-lives for reaction with morpholine of 119 h for the tert butoxyphenyl ester and 1 min for the corresponding phenol. PMID- 11281763 TI - Site selectivity in the synthesis of O-methylated hydroxamic acids with diazomethane. AB - In this paper we report the results obtained by treating some selected hydroxamic acids with diazomethane in ethereal media. The multitask reagent diazomethane was used either as a base to induce deprotonation of the chosen hydroxamic acids or as conjugated acid which undergoes one-pot methylation processes of the generated anions. Product distributions clearly showed that a high site selectivity is expressed by the different deprotonated species in the alkylation processes. Under the adopted conditions, the prevalent site of methylation is in all the cases the oxygen of the hydroxamic acid. While in aliphatic hydroxamic acids only O-alkylation is observed, in the aromatic substrates, the NH group competes with the OH function as the nucleophilic site, although the OH reactivity still dominates. PMID- 11281765 TI - The tricarbonylchromium template for stereocontrol in radical reactions of arenes. AB - Chromium tricarbonyl complexed aryl aldeyhydes and ketones underwent Sm(II) promoted radical lactone formation in the presence of alpha,beta-unsaturated esters to produce diastereomerically pure lactones in good yields. The completely diastereoselective lactone formation involves capture of the benzylic ketyl radical by the ester anti to the chromium tricarbonyl moiety. The relative stereochemistry of the lactone and chromium tricarbonyl moieties was proven by X ray crystallography and supports the proposed mechanism. Enantiopure chromium tricarbonyl complexed arenes afforded single enantiomers when subjected to Sm(II) promoted radical lactone formation condiditions. The enantio- and diastereomerically pure chromium tricarbonyl complexed lactones were subsequently treated with BF3.OEt2 to generate a mixture of diastereomers via Lewis acid promoted chromium tricabonyl directed cationic rearrangement. The diastereomers were separated and individually decomplexed with I2 to afford both of the corresponding chromium-free enantiomerically pure lactones starting from a single enantiomerically pure chromium tricarbonyl complex. PMID- 11281764 TI - C(5)-C(5a)-modified bicyclomycins: synthesis, structure, and biochemical and biological properties. AB - Bicyclomycin (1) is a novel antibiotic that targets rho transcription termination factor in Escherichia coli. We have demonstrated that retention of the C(5)-C(5a) exomethylene unit in 1 is not essential for inhibition. In a recent paper we proposed a working model for 1 and rho function and suggested that 1 binds in a cleft with the C(5)-C(5a) exomethylene unit directed toward the dimeric interface of two rho monomers. This report examines the bicyclomycin C(5)-C(5a) structural constraints necessary for retention of rho inhibitory activity. Three classes of C(5)-C(5a)-modified bicyclomycins have been prepared and their inhibitory activities evaluated in the poly C-dependent ATPase and filter disk antimicrobial assays. The first series consisted of 12 analogues (8-19) that contained a C(5a) unsaturated substituent and possessed C(5E)-geometry. The second set were a pair of C(5a)-substituted C(5E)- and C(5Z)-geometrical isomers (21 and 23). The final group of compounds consisted of six C(5)-C(5a)-dihydrobicyclomycins (24-28, 34) where the terminal substituent was systematically varied. We find that extending the C(5)-C(5a) double bond with unsaturated substituents provides bicyclomycin derivatives with excellent inhibitory activities in the biochemical assay, and that enhanced inhibitory activity is observed for the C(5E) geometrical isomer compared with its C(5Z) counterpart. Finally, C(5a)-substituted dihydrobicyclomycin inhibitory activity appears to be tightly regulated by the nature and spatial placement of the C(5a)-terminal substituent with respect to the [4.2.2]-bicyclic ring system. The observed biochemical activities for the C(5a)-extended conjugated bicyclomycin derivatives and the (5E) and (5Z) isomers were correlated with a structural model for the 1-rho complex. PMID- 11281766 TI - Chemo- and diastereoselectivity in the dimethyldioxirane oxidation of 2,3-dihydro 4H-1-benzothiopyran-4-ones and 4H-1-benzothiopyran-4-ones. Unusual reactivity of 4H-1-benzothiopyran-4-one 1-oxides. AB - The oxidation of the 1-thiochromanones 1-3 by dimethyldioxirane (DMD) produced the corresponding sulfoxides 4-6 or sulfones 7-9; their relative amounts depended on the amount of oxidant used. A low diastereoselectivity was observed in the sulfoxidation of the 2-substituted 1-thiochromanones 2 and 3, due to the small steric differentiation during the DMD attack. An unusual reactivity pattern was found in the DMD oxidation of the 1-thiochromones 10-12, in that the sulfoxides 13-15 were more reactive toward the electrophilic oxidizing agent than the corresponding sulfides. The observed anomaly may be explained in terms of transannular stabilization of the transition structure (TS) for the sulfone formation, promoted through favorable conformational effects in the sulfoxide. Higher sulfoxide/sulfone ratios were found in solvents of greater hydrogen bond donor capacity, which is in accordance with the postulated stabilizing effect. PMID- 11281767 TI - Novel[1n]-meta-heterophane frameworks with a bis-betaine nature. AB - A convergent "5 + 1" and "5 + 3" synthetic strategy allowed the synthesis of the first examples of bis-betaines 2 and 3, a prototype of phanes that incorporate heterocyclic betaines. The structure of the quadrupolar macrocyclic systems 2 and 3 together with the dicationic [1(6)]- and [1(8)] meta-heterophane precursors 5*2X and 6*2X were examined by spectroscopy using 1H and 13C NMR techniques together with 1H-DNMR studies and electrospray ionization mass spectrometry. PMID- 11281768 TI - Polynucleating open-chain systems with imidazole and proton-ionizable 1,2,4 triazole structural motifs. AB - A multistep route for obtaining the polynucleating open-chain systems 3-5 is reported. These advanced intermediates required elaborate processes that proceeded for the pentanuclear protophanes 3 in seven steps, whereas the trinuclear compounds 4 and 5 were obtained in six steps PMID- 11281769 TI - Deoxysugars via microbial reduction of 5-acyl-isoxazolines: application to the synthesis of 3-deoxy-D-fructose and derivatives. AB - 5-Acylisoxazolines 3a-d were obtained by 1,3-dipolar cycloaddition from acetoxymethyl vinyl ketone and nitro precursors. Compounds 3a-d were biotransformed by Aspergillus niger into a 1:1 mixture of stereomers of 5 dihydroxyethyl isoxazolines (+)-4a-d (anti) and (-)-5a-d (syn). Both stereomers were obtained in good yields and with high optical purities. Carbonyl reduction by Aspergillus niger produces alcohols of R-configuration thus giving an access to D-sugar analogues: Compound (+)-4d was converted to 3-deoxy-D-erythro-hexulose and several protected derivatives. Total synthesis of 3-deoxy-D-fructose-6 phosphate was also achieved in two steps and 64% overall yield from (+)-4d. PMID- 11281770 TI - A Study of factors affecting alpha-(N-carbamoyl)alkylcuprate chemistry. AB - The effect of Cu(I) salt (i.e., CuCN, CuCN.2LiCl, CuI), cuprate reagent, sec butyllithium quality, solvent, and temperature upon the chemical yields obtained in the reactions of alpha-(N-carbamoyl)alkylcuprates [i.e., N-Boc-protected alpha aminoalkylcuprates] with (E)1-iodo-1-hexene, 5,5-dimethyl-2-cyclohexenone, methylvinyl ketone, crotonate esters, and an acid chloride has been examined. Cuprate conjugate addition and vinylation reactions can succeed with low-quality sec-butyllithium, presumably containing insoluble lithium hydride and lithium alkoxide impurities, although yields are significantly lower than those obtained with high-quality s-BuLi. alpha-(N-Carbamoyl)alkylcuprates prepared from high quality sec-butyllithium are thermally stable for 2-3 h at room temperature and are equally effective when prepared from either insoluble CuCN or THF-soluble CuCN.2LiCl. Use of the latter reagent permits rapid cuprate formation at -78 degrees C, thereby avoiding the higher temperatures required for cuprate formation from THF-insoluble CuCN that are problematic with solutions containing thermally unstable alpha-lithiocarbamates. PMID- 11281772 TI - Design and synthesis of conformationally constrained glycosylated amino acids. AB - To probe the conformational requirements of O-linked glycoproteins for binding to various enzymes and receptors, two conformationally constrained glycosylated amino acids, 2 and 3, were designed. The analogues were found to represent two potential low energy conformations of the parent conjugate, 1, by molecular modeling. A convergent synthesis of both 2 and 3 from D-galactose and L methionine is presented. PMID- 11281771 TI - A new class of glycosidase inhibitor: synthesis of salacinol and its stereoisomers. AB - Salacinol (4) is one of the active principles in the aqueous extracts of Salacia reticulata that are traditionally used in Sri Lanka and India for the treatment of diabetes. The syntheses of salacinol (4), the enantiomer of salacinol (5), and a diastereomer (7) are described. The synthetic strategy relies on the selective nucleophilic attack of 2,3,5-tri-O-benzyl-1,4-anhydro-4-thio-D- or L-arabinitol at C-1 of 2,4-O-benzylidene D- or L-erythritol-1,3-cyclic sulfate. The work serves to resolve the ambiguity about the exact structure of salacinol and establishes conclusively the structure of the natural product. PMID- 11281774 TI - Enantioselective synthesis of (-)-wikstromol using a new approach via malic acid. AB - The total synthesis of (-)-wikstromol, a bioactive alpha-hydroxylated lactone lignan, from natural malic acid using a consecutive alkylation strategy is presented. First, alkylation of a malic acid ester provided the monobenzyl derivative, which was then converted to an alpha-substituted dioxolanone. This derivative was reacted in a second alkylation step to a double benzylated dioxolanone, which was transformed to bis-O-benzyl-protected (-)-wikstromol and subsequently to the natural product. Only six steps were required to produce wikstromol in 30% overall yield. A second approach from malic acid, the double alkylation of dienolates from 5-oxo-1,3-dioxolan-4-yl acetic acid derivatives, was not successful. No reaction conditions were found to afford the dienolates. Instead, rapid fragmentation of the dioxolanones to fumaric acid derivatives and pivalaldehyde occurred even at -105 degrees C, and aldol reaction products with good stereoselectivity were formed. The relative configuration of the major isomer was determined by X-ray structure analysis. By comparison of NMR data it is shown that a previous assignment of the configuration of one of the described aldol products was incorrect. PMID- 11281773 TI - Solid-phase synthesis of O-linked glycopeptide analogues of enkephalin. AB - The synthesis of 18 N-alpha-FMOC-amino acid glycosides for solid-phase glycopeptide assembly is reported. The glycosides were synthesized either from the corresponding O'Donnell Schiff bases or from N-alpha-FMOC-amino protected serine or threonine and the appropriate glycosyl bromide using Hanessian's modification of the Koenigs-Knorr reaction. Reaction rates of D-glycosyl bromides (e.g., acetobromoglucose) with the L- and D-forms of serine and threonine are distinctly different and can be rationalized in terms of the steric interactions within the two types of diastereomeric transition states for the D/L and D/D reactant pairs. The N-alpha-FMOC-protected glycosides [monosaccharides Xyl, Glc, Gal, Man, GlcNAc, and GalNAc; disaccharides Gal-beta(1-4)-Glc (lactose), Glc beta(1-4)-Glc (cellobiose), and Gal-alpha(1-6)-Glc (melibiose)] were incorporated into 22 enkephalin glycopeptide analogues. These peptide opiates bearing the pharmacophore H-Tyr-c[DCys-Gly-Phe-DCys]- were designed to probe the significance of the glycoside moiety and the carbohydrate-peptide linkage region in blood brain barrier (BBB) transport, opiate receptor binding, and analgesia. PMID- 11281775 TI - A novel route to the marasmane skeleton via a tandem rearrangement cyclopropanation reaction. total synthesis of (+)-isovelleral. AB - A general and efficient route to the marasmane skeleton is described. Total syntheses of two simple marasmanes (35 and 37) in racemic form were achieved using a MgI2-catalyzed rearrangement-cyclopropanation reaction of trimethylsilyl enol ether 31 derived from naphthalenone 30. The reaction proceeds in high yield with complete diastereoselectivity and does not require the use of special cyclopropanation reagents. Application of this novel route to the marasmane framework was extended to the synthesis of naturally occurring (+)-isovelleral (41). PMID- 11281776 TI - Optically active dendrimers with a binaphthyl core and phenylene dendrons: light harvesting and enantioselective fluorescent sensing. AB - Optically active dendrimers containing a 1,1'-binaphthyl core and cross conjugated phenylene dendrons were synthesized and characterized. The chiral optical properties of these phenylene-based dendrimers are different from the previously reported phenyleneethynylene-based dendrimers probably because of the increased steric interaction between the adjacent phenylene units. UV and fluorescence spectroscopic studies demonstrate that the energy harvested by the periphery of the dendrimers can be efficiently transferred to the more conjugated core, generating much enhanced fluorescence signal at higher generation. The fluorescence of these dendrimers can be quenched both efficiently and enantioselectively by chiral amino alcohols. The energy migration and light harvesting effects of the dendrimers make the higher generation dendrimer more sensitive to fluorescent quenchers than the lower ones. Thus, the dendritic structure provides a signal amplification mechanism. These materials are potentially useful in the enantioselective recognition of chiral organic molecules. PMID- 11281777 TI - Ab initio analysis of lithium dimethylaminoborohydride. AB - Ab initio calculations were used to determine the equilibrium geometries and energies of lithium dimethylaminoborohydride. Relative energies of the monomeric and dimeric species were calculated in the gas phase and for the dimethyl ether microsolvated molecules. The most stable structure was a dimer in which the lithium and boron atoms were bridged by two hydrogen atoms, similar to the three center two-electron bonds in diborane. This hydrogen bridging was maintained in the lithium dimethylaminoborohydride bis(dimethyl ether) microsolvate. PMID- 11281778 TI - Conformations of allylic fluorides and stereoselectivities of their diels-alder cycloadditions. AB - The preparations of new allylic fluorides from the corresponding alcohols are reported. Conformational analysis is achieved by comparison of experimental NMR measurements with theoretical (B3LYP) calculations of relative energies of conformers and J(H,H) and J(H,F) coupling constants. The Diels-Alder reactions of allylic fluorides are investigated experimentally and theoretically. The stereoselectivities of the reactions were determined by NMR analysis and, in one case, by X-ray crystallography. Theoretical predictions of stereoselectivity based upon transition state modeling provided good agreement with experiment. Theoretical models for allylic fluorides and transition state conformations are reported. PMID- 11281779 TI - Total synthesis of the epidermal growth factor inhibitor (-)-reveromycin B. AB - The total synthesis of the epidermal growth factor inhibitor reveromycin B (2) in 25 linear steps from chiral methylene pyran 13 is described. The key steps involved an inverse electron demand hetero-Diels-Alder reaction between dienophile 13 and diene 12 to construct the 6,6-spiroketal 11 which upon oxidation with dimethyldioxirane and acid catalyzed rearrangement gave the 5,6 spiroketal aldehyde 9. Lithium acetylide addition followed by oxidation/reduction and protective group manipulation provided the reveromycin B spiroketal core 8 which was converted into the reveromycin A (1) derivative 6 in order to confirm the stereochemistry of the spiroketal segment. Introduction of the C1-C10 side chain began with sequential Wittig reactions to form the C8-C9 and C7-C6 bonds, and a tin mediated asymmetric aldol reaction installed the C4 and C5 stereocenters. The final key steps to the target molecule 2 involved a Stille coupling to introduce the C21-C22 bond, succinoylation, selective deprotection, oxidation, and Wittig condensation to form the final C2-C3 bond. Deprotection was effected by TBAF in DMF to afford reveromycin B (2) in 72% yield. PMID- 11281780 TI - A novel approach to the taxane BC ring system through formation of alpha-ketol by oxidative removal of the phenylsulfonyl group with subsequent in situ oxidation. AB - Cis-fused bicyclo[6.4.0]dodecene 11 was converted into taxane BC ring system 21 in three steps; transformation of the phenylsulfonyl group to an alpha-hydroxy carbonyl group by the treatment with potassium hexamethyldisilazide (KHMDS) and triethyl phosphite under oxygen atmosphere, followed by reductive elimination of the hydroxyl group of alpha-ketol moiety, and inversion of ring juncture. Epimerization of the sulfonyl group of 11 was indispensable for the first oxidation process (17 --> 18) and the second oxidation of 12 leading to hydroxylation at the alpha-position of the carbonyl group proceeded with high regio- and stereoselectivity to give 13. On the other hand, reaction of the cross conjugated compound 5 with KHMDS at 0 degrees C brought about a complete reorganization of molecular framework to provide the compound 7 in which the five membered ring and the conjugated seven-membered ring were connected through a single bond. PMID- 11281781 TI - Building a small polypropionate library. Synthesis of all possible stereotetrads (building blocks for polyketide synthesis) from furan. AB - The all diastereomeric stereotetrads (polypropionate fragments) were synthesized in a stereodivergent fashion, starting from the Diels-Alder adducts of furan with acrylic acid and (-)-2-camphanoxyacrylonitrile, respectively. The key intermediates of these sequences were the oxanorbornenic sulfones 7, (-)-53 and (+)-54. Regio- and stereocontrolled alkylative ring opening of these intermediates afforded the cyclohexenyl sulfones 14, (+)-55 and (-)-58 which were transformed into the desired stereotetrads 30, 31, 40, 41, (+)-62, (+)-63, (-) 66, and (+)-69. PMID- 11281782 TI - An approach toward isoindolobenzazepines using the ammonium ylide/Stevens. AB - Ammonium ylides derived from the Cu(II)-catalyzed decomposition of alpha-diazo carbonyls tethered to tertiary amines underwent a benzylic Stevens [1,2] rearrangement to give tetrahydroisoquinolines or benzazepines containing fused five-membered rings, a feature found in the cephalotaxus alkaloids. Model studies were also carried out toward the synthesis of lennoxamine, a member of the isoindolobenzazepine family of alkaloids. The approach utilized is based on the Rh(II)-catalyzed reaction of an alpha-diazo carbonyl compound containing an amido group in the gamma-position. Treatment of several N,N-dialkyl-substituted amido diazo-esters with Rh2(OAc)4 in benzene at 80 degrees C in the presence of several dienophiles gave [4 + 2]-cycloadducts derived from the Diels-Alder reaction of a transient alpha-amino isobenzofuran intermediate. In the absence of an external trapping agent, no rearranged product derived from an ammonium ylide intermediate could be detected in the crude reaction mixture. In contrast to this result, reaction of the related diazo dihydroisoquinoline amide 46 with Rh2(OAc)4 afforded the isoindolobenzazepine ring system in high yield. Formation of the 5,7 fused skeleton was rationalized in terms of a spirocyclic ammonium ylide that underwent a preferential Stevens [1,2]-shift of the benzylic carbon atom. While we were ultimately thwarted in using the ammonium ylide/rearrangement cascade for a lennoxamine synthesis by an uncooperative diazo transfer reaction, the cascade sequence was shown to be useful for the preparation of various isoindolobenzazepines. PMID- 11281783 TI - Correlation of ionization potentials and HOMO energies versus relative reactivities of Cl2, of Br2, and of I2 with representative acyclic alkenes. comparison with other additions to alkenes. AB - Plots of log k(rel) versus IP or versus HOMO for the title reactions are presented; similarities and differences among the reactions are discussed. The Cl2 and Br2 data each show a single line of correlation with positive slope for all alkenes regardless of the steric requirements; increasing substitution at the double bond increases the reaction rate, indicating an electrophilic reaction. Each plot of the I2 data calculated for adsorption exhibits a natural separation into groups of similarly substituted alkenes, in which increased substitution reduces the rate. Within each group, a good-to-excellent correlation is observed, with a lower IP generally corresponding to a higher relative rate. The results indicate that the relative magnitude of the steric requirements about the double bond is similar to that of the electronic effects in iodination. Plot shapes for iodination are compared to those of other reactions, such as hydroboration, oxymercuration, complexation with Ag+, and complexation with MeHg+. PMID- 11281784 TI - Selenium-catalyzed oxidations with aqueous hydrogen peroxide. 2. Baeyer-Villiger reactions in homogeneous solution. AB - Several diselenides were tested for catalytic activity in Baeyer-Villiger reactions with 60% aqueous hydrogen peroxide. Bis[3,5-bis(trifluoromethyl)phenyl] diselenide forms the corresponding 3,5-bis(trifluoromethyl)benzene seleninic acid in situ, which is a highly reactive and selective catalyst for the oxidation of carbonyl compounds in 1,1,1,3,3,3-hexafluoro-2-propanol, 2,2,2-trifluoroethanol, or dichloromethane. PMID- 11281785 TI - Facile and efficient sulfenylation method using quinone mono-O,S-acetals under mild conditions. AB - A novel sulfenylation method induced by aromatization of quinone mono-O,S-acetals is described. These sulfenylation reagents readily react with silyl enolethers or electron rich aromatic compounds to give sulfenylation products under mild conditions. In particular, O,S-acetal 2j, which possesses a pentafluorophenylthio function, is the most effective reagent from the standpoint of the adaptability for various substrates. PMID- 11281786 TI - Efficient preparation of natural and synthetic galactosides with a recombinant beta-1,4-galactosyltransferase-/UDP-4'-gal epimerase fusion protein. AB - The numerous biological roles of LacNAc-based oligosaccharides have led to an increased demand for these structures for biological studies. In this report, an efficient route for the synthesis of beta-galactosides using a bacterial beta-4 galactosyltransferase/-UDP-4'-gal-epimerase fusion protein is described. The lgtB gene from Neisseria meningitidis and the galE gene from Streptococcus thermophilus were fused and cloned into an expression vector pCW. The fusion protein transfers galactose to a variety of different glucose- and glucosamine containing acceptors, and utilizes either UDP-galactose or UDP-glucose as donor substrates. A crude lysate from Escherichia coli expressing the fusion protein is demonstrated to be sufficient for the efficient preparation of galactosylated oligosaccharides from inexpensive UDP-glucose in a multigram scale. Lysates containing the fusion protein are also found to be useful in the production of more complex oligosaccharides in coupled reaction mixtures, e.g., in the preparation of sialosides from N-acetylglucosamine. Thus, bacterially expressed fusion protein is well suited for the facile and economic preparation of natural oligosaccharides and synthetic derivatives based on the lactosamine core. PMID- 11281787 TI - Modeling the 1,3-dipolar cycloaddition of nitrones to vinylboranes in competition with boration, cyclization, and oxidation reactions. AB - Structures and energetics of reactants, reactant complexes, concerted transition structures, and products of the cycloaddition of the prototypical nitrone with vinylborane have been produced and discussed. Structure optimizations have been performed at the B3LYP/6-31G(d) and B3LYP/AUG-cc-pVDZ levels of approximation, and single-point calculations on the B3LYP geometries have been carried out at the MP4(SDTQ) level with the same basis sets. Kinetic contributions to standard enthalpies, entropies, and free enthalpies have been computed at the same levels of geometry optimizations. The effects of methyl and chloro substitution on the BH2 group and of methyl substitution on the vinyl moiety has been also explicitly considered. The most striking theoretical features of this cycloaddition are (i) the formation of reactant complexes where the nitrone oxygen is strictly bound up to the boron atom (B...O interactions), (ii) their persistence in the endo/exo transition structures, and (iii) energy profiles suggesting very high reaction rates, regiospecificity (5-borylisoxazolidines) and complete endo stereoselectivity. The BH2 (BX2) substituent appears to induce a sort of intramolecular catalysis which is also largely selective in favor of the endo reaction path. Possible competitive reaction paths such as cyclization, organoboration, and oxidation have equally been investigated, on the same grounds, both with prototypical reagents and with dimethylvinylborane, dichlorovinylborane, 2-methyl-1-propenylborane, and 2-methyl-1 propenyldichloroborane. The transition structures for these reaction paths are significantly higher in energy than those of the corresponding 1,3-dipolar cycloadditions in the sequence oxidation >> cyclization > boration > cycloaddition, whereas the resulting reaction products show the reversed sequence. Polar solvents appear to increase the competition of boration although maintaining its character of secondary reaction. As expected, the reaction rate of 1,3-dipolar cycloaddition is lowered by dimethyl substitution on the vinyl CH2 reacting center (i.e., for the reaction of 2-methyl-1-propenylborane and 2-methyl 1-propenyldichloroborane) whereas the reaction rate of boration is increased, the boration results being significantly competitive even in the gas phase. Experiments for the control of the above predictions are not yet available. PMID- 11281788 TI - Suzuki reaction of vinyl triflates from six- and seven-membered N-alkoxycarbonyl lactams with boronic acids and esters. AB - The Pd(0)-catalyzed reaction of vinyl triflates from N-alkoxycarbonyl lactams with different boron compounds has been studied. The coupling reaction of alkenylboronates and arylboronic acids with six- and seven-membered lactam derived N-alkoxycarbonyl vinyl triflates was feasible under very mild conditions in THF-water employing (Ph3P)2PdCl2 as a catalyst and Na2CO3 as a base, which provided in high yields the corresponding 6- or 7-substituted N-alkoxycarbonyl 3,4-dihydro-2H-pyridines and N-alkoxycarbonyl-2,3,4,5-tetrahydroazepines. Allylboronates reacted slower but, with vinyl triflates from delta-valerolactam, still gave acceptable yields of the coupling product. Alkylboronic acids required different reaction conditions, in particular the presence of Ag2O together with a base in anhydrous toluene and (dppf)PdCl2 as a catalyst, affording the corresponding 6-alkyl-N-alkoxycarbonyl-3,4-dihydro-2H-pyridines in high yields. PMID- 11281789 TI - Regio- and stereochemically controlled formation of hydroxamic acid containing anti- or syn-1,4-cycloalkenols from acylnitroso-derived Diels-Alder adducts. AB - Treatment of acylnitroso hetero Diels-Alder cycloadducts 2 with iron(III) or copper(II) in an alcohol solvent induces ring opening to afford predominantly monocyclic anti-1,4-hydroxamic acids 3. However, treatment of cycloadducts 2 with copper(II) in toluene reverses the stereoselectivity of the ring opening to afford syn-1,4-hydroxamic acids 4. These regio- and stereoselective processes separately provide anti-1,4- and syn-1,4-disubstituted cyclopentenes while regenerating a hydroxamic acid moiety, thus enhancing the chemical versatility of the Diels-Alder cycloadducts. PMID- 11281790 TI - Synthesis, properties, and redox behavior of di(1-azulenyl)(2- and 3 thienyl)methyl cations and dications composed of two di(1-azulenyl)methylium units connected with 2,5-thiophenediyl and 2,5-thienothiophenediyl spacers. AB - The titled stable monocations, di(1-azulenyl)(2- and 3-thienyl)methyl cations 7a,b and 8a,b and dications composed of two di(1-azulenyl)methylium units connected with 2,5-thiophenediyl and 2,5-thieno[3,2-b]thiophenediyl spacers 9a,b and 10a,b were prepared by hydride abstraction of the corresponding methane derivatives. These mono- and dications 7a,b, 8a,b, 9a,b, and 10a,b showed high stability with large pK(R)+ values. The values of monocations 7a,b and 8a,b were 11.2-11.8 +/- 0.1 and 11.4-12.4 +/- 0.1, respectively. Two cation units in dications 9a,b and 10a,b were neutralized via one step at the pH of 11.1-11.7 +/- 0.1, which corresponds to the average of the pK(R)+ values of the dications and half-neutralized monocations. Electrochemical behavior of 7a,b, 8a,b, 9a,b, and 10a,b was examined by cyclic voltammetry (CV). Formation of the thienoquinoid products 18a,b and 19a,b from 9a,b and 10a,b was characterized by UV-vis spectroscopy under electrochemical reduction conditions. Chemical reduction of 9a,b and 10a,b with Zn powder in acetonitrile afforded 18a,b and 19a,b as deep colored crystals, which exhibited rather high electron-donating ability. PMID- 11281791 TI - Nitrile reduction in the presence of Boc-protected amino groups by catalytic hydrogenation over palladium-activated Raney-nickel. PMID- 11281792 TI - Efficient stereoselective synthesis of alpha-hydroxy aldehydes with (R)-piperidin 3-ol as a new chiral auxiliary. PMID- 11281793 TI - Concise formation of 4-benzyl piperidines and related derivatives using a Suzuki protocol. PMID- 11281794 TI - A novel and expeditious approach to thiophene-3-carboxylates. PMID- 11281795 TI - Palladium-catalyzed formation of aryl tert-butyl ethers from unactivated aryl halides. PMID- 11281796 TI - Do the anti-selectivities of 2,3-endo,endo-dimethylnorbornan-7-one and the corresponding diethyl analog obey the cieplak model? An ab initio MO investigation and application of the cation complexation model. PMID- 11281797 TI - Synthesis of cyclic ketones by electrochemical reduction of S-(2 methoxycarbonyl)phenyl thiolesters. PMID- 11281798 TI - Concise synthesis of (+)-2-C-methyl-D-erythritol-4-phosphate. PMID- 11281799 TI - Polystyrene-supported benzenesulfonyl azide: a diazo transfer reagent that is both efficient and safe. PMID- 11281800 TI - An efficient asymmetric synthesis of tarchonanthuslactone. PMID- 11281802 TI - (Cyanomethyl)trialkylphosphonium iodides: efficient reagents for the intermolecular alkylation of amines with alcohols in solution and on solid phase. PMID- 11281801 TI - Unsymmetrical ozonolysis of a Diels-Alder adduct: practical preparation of a key intermediate for heme total synthesis. PMID- 11281803 TI - Highly regioselective Vilsmeier-Haack acylation of hexahydropyrroloindolizine. PMID- 11281804 TI - Asymmetric 1,3-dipolar cycloaddition of a (Z)-alkene dipolarophile. Synthesis of (3S,4R) ethyl 1-azabicyclo[2.2.1]heptane-3-carboxylate. PMID- 11281805 TI - Synthesis and absolute stereochemistry of a constitutionally new spiroacetal from an insect. PMID- 11281806 TI - An easy and convenient synthesis of Weinreb amides and hydroxamates. PMID- 11281807 TI - History, celebrations, and the transmission of hope: the American Music Therapy Association, 1950-2000. PMID- 11281808 TI - The effect of parent training in music and multimodal stimulation on parent neonate interactions in the neonatal intensive care unit. AB - This study examined the effects of parent training in music and multimodal stimulation on the quantity and quality of parent-neonate interactions and the weight gain and length of hospitalization of premature and low birthweight (LBW) infants in a Neonatal Intensive Care Unit (NICU). Twenty sets of parents and premature LBW infants participated in the study. Parents in the experimental group (n = 10) received approximately one hour of instruction in appropriate uses of music, multimodal stimulation including massage techniques, and signs of infant overstimulation and techniques for its avoidance. Parent-neonate interactions, specifically parent actions and responses and infant stress and nonstress behaviors, were observed for subjects in both groups. Infant stress behaviors were significantly fewer and appropriateness of parent actions and responses were significantly greater for experimental infants and parents than for control subjects. Parents in the experimental group also self-reported spending significantly more time visiting in the NICU than did parents of control infants. In addition, length of hospitalization was shorter and average daily weight gain was greater for infants whose parents received training, although these differences were not significant. A one month, postdischarge follow-up showed little difference between experimental and control group parent-infant interactions in the home. PMID- 11281809 TI - Improvisation in music therapy: human communication in sound. PMID- 11281810 TI - Music therapy practicum practices: a survey of music therapy educators. AB - Music therapy program directors were surveyed about various aspects of their music therapy practica. The survey addressed various aspects of the structure of the practicum, on-campus clinics, faculty supervision, videotaped supervision, and evaluation of practicum students. Responses were received from 38 educators from undergraduate programs. Results indicate that there is wide variety in the number of practica required and in the amount of time spent in practica. Several populations are included in the practicum experiences offered by every university, although the means of achieving this variety varies. Results also showed variations in the way that practicum experiences are set up so that in some cases students observe and assist, sometimes eventually assuming full responsibility for the session, while in others the students are responsible for the entire session from the beginning. The content of classes that accompany the practica, when such classes exist, is similar from program to program. There are also wide variations in practicum supervision and faculty compensation for this supervision. PMID- 11281811 TI - Beta-amyloid therapies in Alzheimer's disease. AB - Neurones in the brain produce beta-amyloid fragments from a larger precursor molecule termed the amyloid precursor protein (APP). When released from the cell, these protein fragments may accumulate in extracellular amyloid plaques and consequently hasten the onset and progression of Alzheimer's disease (AD). A beta fragments are generated through the action of specific proteases within the cell. Two of these enzymes, beta- and gamma-secretase, are particularly important in the formation of A beta as they cleave within the APP protein to give rise to the N-terminal and C-terminal ends of the A beta fragment, respectively. Consequently, many researchers are investigating therapeutic approaches that inhibit either beta- or gamma-secretase activity, with the ultimate goal of limiting A beta; production. An alternative AD therapeutic approach that is being investigated is to employ anti-A beta antibodies to dissolve plaques that have already formed. Both of these approaches focus on the possibility that accrual of A beta leads to neuronal degeneration and cognitive impairment characterised by AD and test the hypothesis that limiting A beta deposition in neuritic plaques may be an effective treatment for AD. PMID- 11281812 TI - Experimental approaches and drugs in development for the treatment of dementia. AB - Treatment of dementia can be divided as symptomatic treatment of cognitive or non cognitive symptoms and the treatment of underlying pathology. In the last decade the thrust of symptomatic treatment of Alzheimer's disease (AD) has been enhancement of cholinergic transmission. Besides the acetycholinesterase inhibitors (AChE-I) currently in use, cholinergic agonists and enhancers are in development. Other therapeutic approaches directed towards neurotransmitter substitution or modulation include serotoninergic, noradrenergic substances, neuropeptides and those acting via excitatory amino acid receptors, such as ampakines or NMDA antagonists. Introduction of atypical neuroleptics represents the most recent development in the treatment of behavioural symptoms. Efforts to treat the underlying pathology are based on modulation of APP processing in order to decrease the accumulation of beta-amyloid, those to decrease tau hyperphosphorylation, use of nerve growth factors and those based on Apo-E modulation. Potential use of oestrogens and NSAIDs are also under investigation. Recently, vaccination with amyloid-beta peptide has been reported to be effective in an animal model of AD, this putative vaccine is now in clinical trials. Likewise, recent studies suggest that some statins may have a prophylactic effect. PMID- 11281813 TI - Adenosine in the treatment of stroke: yes, maybe, or absolutely not? AB - Agonist stimulation of adenosine A(1) receptors has been consistently shown to result in reduction of brain damage following experimentally induced global and focal brain ischaemia in animals. Unsurprisingly, the use of adenosine A(1) receptors as targets for the development of clinical therapeutics suitable for treatment of ischaemic brain disorders has been suggested by many authors. The latest studies of adenosine and its receptors indicate that adenosine-mediated actions might be far more complex than originally anticipated, casting some doubt about the rapid development of stroke treatment based on adenosine. This review discusses the possible role of adenosine receptor subtypes (A(1), A(2) and A(3)) in the context of their potential as therapeutics in stroke. PMID- 11281814 TI - Kynurenic acid antagonists and kynurenine pathway inhibitors. AB - The kynurenine pathway accounts for the metabolism of around 80% of non-protein tryptophan metabolism. It includes both an agonist (quinolinic acid) at NMDA receptors and an antagonist (kynurenic acid). Since their discovery, there has been a major development of kynurenic acid analogues as neuroprotectants for the treatment of stroke and neurodegenerative disease. Several prodrugs of kynurenic acid or its analogues that can be hydrolysed within the CNS are also available. More recently, the pathway itself has proved to be a valuable drug target, affected by agents which reduce the synthesis of quinolinic acid and increase the formation of kynurenic acid. The change in the balance of these, away from the excitotoxin and towards the neuroprotectant, has anticonvulsant and neuroprotective properties. PMID- 11281815 TI - Therapeutic potential of CRF receptor antagonists: a gut-brain perspective. AB - Activation of the corticotropin-releasing factor (CRF) family of neuropeptide receptors in the brain and periphery appears to mediate stress-related changes in a variety of physiological and functional domains. Comparative pharmacology of CRF receptor agonists suggests that CRF, urocortin, sauvagine and urotensin consistently mimic, and conversely peptide CRF receptor antagonists lessen, the functional consequences of stressor exposure. Together with the development of novel non-peptide CRF receptor antagonists, a growing number of CRF receptor selective ligands are available to elucidate the neurobiology and physiological role of CRF systems. The present review considers available preclinical evidence as well as results from one Phase II clinical trial which address the hypothesis that CRF receptor antagonists may represent a new option for pharmacotherapy of stress-related disorders. PMID- 11281817 TI - Substance P receptor antagonists in the therapy of migraine. AB - Clinical observations, the vascular component of migraine pain, its pulsating or throbbing pain character, have focused attention on the trigeminal innervation of pain-sensitive intracranial structures, such as the dura mater and large vessels. These intracranial structures are innervated by the ophthalmic branch of the trigeminal nerve, which is marked by the presence of vasoactive peptides, such as substance P and calcitonin gene-related peptide. Substance P is a mediator of the sterile inflammation of the dura mater, which has been considered to be the source of migraine pain. Modern antimigraine drugs, such as 5-HT(1B/D) agonists (triptans), block this dural neurogenic inflammation dose-dependently in an animal model but their vasoconstrictor effects have led to a search for non vasoconstrictor approaches. One such approach has been substance P (neurokinin-1) antagonists. These are highly effective in animal models of dural inflammation and have no significant vasoconstrictive effect. However, several NK(1) antagonists failed to demonstrate any effect in acute migraine. Current clinical and experimental evidence therefore supports the view that NK(1) receptor antagonists may have no significant antimigraine properties. PMID- 11281816 TI - Novel treatments for bipolar disorder. AB - Treatments other than lithium have recently emerged as equally important in the management of bipolar disorder. The spectrum of efficacy of newer treatments differs from lithium and among the novel drug treatments valproate, generally used as the better tolerated divalproex form, principally benefits manic symptomatology both acutely and in prophylaxis. Atypical antipsychotic drugs have demonstrated efficacy in reducing acute manic symptoms. No controlled evidence of efficacy in prophylaxis has been published. Lamotrigine has demonstrated efficacy in both acute bipolar depression and maintenance efficacy in rapid cycling bipolar patients, especially those patients with bipolar II disorder, which is principally manifested as depression. Randomised, double-blind, placebo- controlled studies provide good evidence that regimens of risperidone or olanzapine in combination with lithium or valproate provide greater improvement in acute mania than the mood stabilisers alone. Similarly, valproate combined with antipsychotics provided greater improvement in mania than antipsychotic medication alone and resulted in lower dosage of the antipsychotic medication. A positive but unclear placebo-controlled study of omega-3 fatty acids added to lithium in bipolar disorder needs confirmation in standard clinical trial paradigms. Several other drugs that were reported as beneficial in various facets of bipolar disorder in open trials have not differed from placebo when studied in randomised, placebo-controlled trials. PMID- 11281818 TI - Therapeutic potential of prostaglandin analogues in glaucoma. AB - One of the most recent contributions to the therapeutic arsenal available for the treatment of glaucoma is the prostaglandin (PG) analogues. They represent a new class of ocular hypotensive drugs, targeting the uveoscleral outflow of ocular aqueous humour. Two drugs, latanoprost and unoprostone, are presently commercially available. In terms of intraocular pressure (IOP) reduction, latanoprost is the most powerful drug in clinical use today. The once daily dosing promotes compliance. Additional effect is achieved in combination with other hypotensive drugs, including those that increase trabecular outflow facility. The most frequent side effect is increased iris pigmentation that seems to be irreversible. A low frequency of cystoid macular oedema has been reported, predominantly in patients whose blood-retinal barrier (BRB) is compromised. Systemic side effects are rare. The experience with unoprostone is still much less than that with latanoprost. The ocular hypotensive mechanism of action of unoprostone is not well documented but an increase in uveoscleral outflow may be at least a part of its mode of action. Systemic side effects are rare and the ocular side effects seem to be mild. The ocular hypotensive effect is less than that of latanoprost and may not be suitable for monotherapy. It is widely accepted that the IOP alone is not responsible for the development of glaucomatous visual defects. It remains to be seen if this class of drugs will preserve vision in glaucoma patients better than other classes. More PG analogues are under development for potential clinical use. PMID- 11281819 TI - Novel approaches to treat muscular dystrophies. AB - Muscular dystrophies (MD) are a clinically and genetically heterogeneous group of skeletal muscle-wasting diseases. Mutations in the dystrophin gene result in dystrophin deficiency, which constitutes the pathogenic basis of Duchenne and Becker MD (DMD and BMD). Several MD are caused by mutations in other recently identified genes coding for proteins linked to the sarcolemma, the nuclear envelope or the contractile apparatus. In addition, several MD have been mapped to different chromosomal loci and for most of them, the identification of the molecular defect is underway. The immediate result is an ongoing reclassification of the MD into disorders defined not by clinical characteristics but specific genetic mutations. At present, therapy of MD is based on symptomatic treatment and supportive care. Convincing evidence for clinical efficacy is only available for corticosteroids that also suffer from frequent and severe side effects. Up to now, curative therapy is not available, although promising new molecular therapies are under investigation in animal models of MD. Current treatment strategies are discussed and a perspective for effective molecular therapy is given. PMID- 11281820 TI - LHRH analogues as anticancer agents: pituitary and extrapituitary sites of action. AB - Two classes of luteinising hormone-releasing hormone (LHRH) analogues have been developed so far to be used for oncological therapies: LHRH agonists and antagonists. LHRH agonists are widely and successfully used for the management of steroid-dependent malignancies. Chronic administrations of these compounds result in downregulation and desensitisation of pituitary LHRH receptors and, therefore, in a complete suppression of gonadal function. LHRH agonist administration is effective, safe and reversible, suffering only from the 'flare-up' phenomenon at the beginning of treatment. LHRH antagonists suppress the pituitary-gonadal function by competing with native LHRH for binding to its pituitary receptor but without giving rise to the intracellular cascade of events evoked by the natural hormone or LHRH agonists. Synthetic peptides belonging to the last generations of LHRH antagonists have already been successful in clinical trials. They are completely devoid of the 'flare-up' phenomenon and seem to be free of side effects, such as histamine release. Recently, the expression of LHRH and LHRH receptors has been reported in a number of hormone-responsive tumours. In contrast with the pituitary LHRH receptor which is coupled to the Gq/11-PLC intracellular system of events, stimulation of the tumour LHRH receptor by LHRH is followed by the activation of a Gi protein and a decrease in cAMP levels. This intracellular pathway mediates the inhibitory action of the autocrine/paracrine LHRH system on tumour cell proliferation. The activation of LHRH receptors at tumour level may then represent an additional and more direct mechanism of action for the antitumoural activity of LHRH agonists. Surprisingly, LHRH antagonists also exert a marked antimitogenic activity on a number of hormone-responsive cancer cell lines, indicating that these compounds might behave as antagonists at pituitary level and as agonists at the level of the tumour. The observation that the inhibitory LHRH autocrine system is also present in some steroid-unresponsive cancer cell lines might suggest a possible clinical utility of LHRH analogues also for those tumours that have escaped the initial phase of hormone dependency. PMID- 11281821 TI - Bimatoprost: a member of a new class of agents, the prostamides, for glaucoma management. AB - Bimatoprost, a synthetic analogue of endogenous prostamides, is in development as a topical ocular hypotensive agent for the treatment of glaucoma and ocular hypertension. Prostamides are a newly discovered class of compounds that have been shown to have potent ocular hypotensive activity in the laboratory. Bimatoprost mimics the endogenous prostamides by lowering intraocular pressure (IOP). Bimatoprost provides outstanding control of IOP throughout the day, and a high percentage of patients receiving bimatoprost achieve the low target pressures important for clinical success. In controlled clinical trials, bimatoprost 0.03% given once daily has displayed efficacy superior to timolol 0.5% given twice daily, the current standard for therapy. Analysis of pooled six month data from two large Phase III trials demonstrated that mean IOP was consistently 2 - 3 mmHg lower with bimatoprost q.d. than with timolol b.i.d. Bimatoprost 0.03% q.d. has also been shown to provide significantly better diurnal IOP control than latanoprost 0.005% q.d., probably the most efficacious topical medication currently available. Patients receiving bimatoprost q.d. were more likely than timolol or latanoprost patients to achieve low target pressures. In all clinical evaluations, bimatoprost q.d. has been demonstrated to be safe and well-tolerated. Bimatoprost will likely be available for clinical use in 2001 and it has great potential to be superior to all other medications in IOP lowering efficacy. It is anticipated that bimatoprost will have an important role in therapy for glaucoma and ocular hypertension. PMID- 11281822 TI - Tiotropium bromide. AB - Tiotropium bromide is a new long-lasting anticholinergic drug which, like ipratropium bromide, is a quaternary ammonium derivative. It binds with high affinity to muscarinic receptors but dissociates very slowly from M(1)- and M(3) muscarinic receptors. Pharmacology studies have demonstrated a prolonged protective effect against cholinergic agonists and cholinergic nerve stimulation in animal and human airways. In Phase II studies single inhaled doses of tiotropium bromide have a bronchodilator and bronchoprotective effect in asthmatic and chronic obstructive pulmonary disease (COPD) patients of over 24 h. In Phase III studies, once daily inhaled tiotropium is an effective bronchodilator in COPD patients, giving great improvement in lung function and reduction in symptoms than ipratropium bromide given four times daily. The drug is well-tolerated and the only side effect of note is dryness of the mouth which occurs in approximately 10% of patients. Since, anticholinergics are the bronchodilators of choice in COPD it is likely that tiotropium bromide will become the most widely used bronchodilator for COPD patients in the future. PMID- 11281823 TI - The status of ongoing trials for mild cognitive impairment. AB - Mild cognitive impairment (MCI) is a term used to describe memory decline or other specific cognitive impairment in individuals who do not have dementia or significant impairment of other cognitive functions beyond that expected for their age or education. It has been suggested that as much as 38% of the elderly population would meet criteria for MCI and although the associated memory deficits are mild, the fact that up to 15% of MCI patients, particularly those with a particular type of memory impairment, convert to Alzheimer's disease (AD) annually has prompted serious attention. Despite the high conversion rate, MCI cannot be used synonymously with early or mild AD, as patients with AD are impaired not only in memory performance but in other cognitive domains as well; they meet diagnostic criteria for dementia. However, since there is a high conversion rate from MCI to AD, it is likely many with MCI have the underlying neuropathology of AD, though they do not yet meet clinical diagnostic criteria. Therefore, treatment strategies developed for AD, specifically acetylcholinesterase inhibitors and Cox-2 inhibitors, have been among the first employed to treat MCI. It is hoped that by impeding the progression of MCI in this manner, fewer patients will convert to AD. This article will give a brief overview of the condition of mild cognitive impairment and an account of trial methodology and current treatment strategies being employed for MCI. PMID- 11281824 TI - Neuroprotective agents in traumatic brain injury. AB - The role of neuroprotection in traumatic brain injury (TBI) is reviewed. Basic research and experimental investigations have identified many different compounds with potential neuroprotective effect. However, none of the Phase III trials performed in TBI have been successful in convincingly demonstrating efficacy in the overall population. A common misconception is that consequently these agents are ineffective. The negative results as reported in the overall population may in part be caused by specific aspects of the head injury population as well as by aspects of clinical trial design and analysis. The heterogeneity of the TBI population causes specific problems, such as a risk of imbalances between placebo and treated groups but also causes problems when a possible treatment effect is evaluated in relation to the prognostic effect present. Trials of neuroprotective agents should be targeted first of all to a population in which the mechanism at which the agent is directed is likely to be present and secondly to a population in which the chances of demonstrating efficacy are realistic, e.g., to patients with an intermediate prognosis. The possibilities for concomitant or sequential administration of different neuroprotective agents at different times deserve consideration. The potential for neuroprotection in TBI remains high and we should not be discouraged by recent failures obtained up until now. Rather, prior to initiating new trials, careful consideration of experimental evidence is required in order to optimise chances for mechanistic targeting and lessons learned from previous experience need to be taken to heart in the design of future studies. PMID- 11281825 TI - The multicomponent reactions and their libraries for natural and preparative chemistry. AB - It was recently recognized that three different types of multi-component reactions (MCRs) exist. In preparative chemistry, the MCRs of type II form their products particularly efficiently. These reactions correspond to equilibria of educts and intermediate products, whose final products are formed practically irreversibly. In recent years, the four component reaction of the isocyanides (U 4CR) of type II and their unions with various reactions and MCRs have become an important industrial process for preparing products and their libraries. It has been demonstrated that all conceivable collections of U-4CR educts can be converted into the corresponding products. In the usual chemical reactions, only the substituents of the products can be varied, whereas the U-4CR and related reactions can also produce skeletally different types of products with diverse substituents. The preparative advantages of forming products by the one-pot MCRs and the great variety of the possible products are illustrated in this review. PMID- 11281826 TI - Dynamic combinatorial chemistry. AB - Dynamic combinatorial chemistry is based on the reversible combination of initial building blocks to form dynamic combinatorial libraries. It has recently emerged as an efficient strategy to detect and to evaluate affinity between the library products and a target molecule. In this review, examples from various fields of chemistry and biochemistry are presented and extensively discussed. The last section deals with the practical aspects for implementing this approach. PMID- 11281827 TI - Antigenic and immunogenic phage displayed mimotopes as substitute antigens: applications and limitations. AB - The most exciting potential of phage displayed peptide libraries is to obtain small peptide molecules that mimic an antigen, at least with respect to a particular epitope. In addition to their interest as research tools, such mimotopes could in principle be useful as diagnostic tools or for eliciting antibodies to a predefined epitope. However, the reduction of the phage insert sequence to a short peptide that can compete with the antigenic and in particular with the immunogenic properties of the natural antigen faces considerable difficulties. This review assesses critically the antigenicity of phage displayed peptides as free peptides and in different molecular environments. The difficulties to use mimotopes to induce antibodies that bind to the natural antigen (crossreactive immunogenicity) and the considerable discrepancy between antigenicity and immunogenicity of phage-derived peptides are discussed. Peptides selected with antibodies from phage displayed random peptide libraries have raised considerable expectations as low molecular weight substitutes of the natural antigen. This review will focus on the results of phage displayed random peptide libraries screened with antibodies specific for proteins, carbohydrates and nucleic acids and critically examine how the above expectations have been met. PMID- 11281828 TI - Solid phase synthesis of 4-hydroxycinnamic acid and its derivatives for potential use in combinatorial chemistry: a novel route for the synthesis of 4 hydroxycinnamoyl coenzyme A and NMDA receptor antagonists. AB - Synthesis of 4-hydroxycinnamic acid 6 and its N-hydroxysuccinimide ester 8 has been carried out in high yield on solid support. Further development allowed the synthesis of 4-hydroxycinnamoyl CoA 1 in excellent overall yield. The utility of solid phase as a method for the synthesis of 4-hydroxycinnamic acid derivatives was demonstrated by the synthesis of a number of compounds including the NMDA receptor antagonists, N-(phenylalkyl)cinnamides 9 and 10. PMID- 11281830 TI - Shotgun phage display cloning. AB - Shotgun phage display cloning is a useful tool for studying interactions between bacterial and host proteins. Libraries are constructed by cloning randomly fragmented prokaryotic DNA into phage mid-vectors. Theoretically, these libraries will consist of phages that together display all proteins encoded by the bacterial genome. Selecting a gene III-based library, made from Staphylococcus aureus DNA, against IgG and fibronectin resulted in 20-40% positive clones after two pannings. Increasing the number of fusion proteins per phage particle by using gene VIII-based display, increased the frequency of correct clones to 75 100%. PMID- 11281829 TI - Alternative bacteriophage display systems. AB - Filamentous phage has been extensively used to implement various aspects of phage display technology. The success of these organisms as vectors to present foreign peptides and to link them to their coding sequences is a consequence of their structural and biological characteristics. Some of these properties, however, represent a limitation when one attempts to display proteins that cannot be efficiently exported through the bacterial membrane or do not fold properly in the periplasm. Thus, the desirability of developing alternative display systems was recognised recently and led to the development of a different class of display vectors that assemble their capsid in the cytoplasm and are released via cell lysis. This review describes and compares the properties of these alternative display systems. PMID- 11281831 TI - The powerful combination of phage surface display of cDNA libraries and high throughput screening. AB - Phage surface display of cDNA libraries facilitates cloning, expression and rapid selection of functional gene products physically linked to their genetic information through gene product-ligand interactions. Efficient screening technologies based on selective enrichment of clones expressing desired gene products allows, within a short time, the isolation of all ligand-specific clones that are present in a library. Manual identification of clones by restriction analysis and random sequencing is unlike to be successful for the isolation of gene products derived from rare mRNA species resulting from selection of the libraries using polyvalent ligands like serum from patients. Here we describe rapid handling of large numbers of individual clones selected from molecular libraries displayed on phage surface using the power of robotics-based high throughput screening. The potential of the combination of cDNA-phage surface display, with selection for specific interactions by functional screening and robotic technology is illustrated by the isolation of more sequences potentially encoding IgE-binding proteins than postulated from Western blot analyses using extracts derived from raw material of complex allergenic sources. The subsequent application of functional enrichment and robotics-based screening will facilitate the rapid generation of information about the repertoire of protein structures involved in allergic diseases. PMID- 11281832 TI - Peptide display in functional genomics. AB - The completion of the human genome project has opened novel scientific avenues in functional genomics, structural genomics and proteomics. These areas have a common goal: the identification of all the proteins acting and cross-talking in a single cell at a defined moment of its lifecycle. The expansion of these areas in bioscience has been facilitated by the rapid development of high throughput screening (HTS) methods which has, in turn, attracted the business community to make investments in this novel business segment of biotechnology. By using these HTS methods, the hope is that novel targets will be validated much more rapidly speeding up the development of novel drugs. Numerous techniques and tools have emerged over the past decade for the identification of small target-specific molecular ligands that exploit a common feature: the exploration of molecular diversity using combinatorial methods. While chemists developed new methods for rapidly and efficiently synthesising and screening large collections of small molecules, biologists used recombinant DNA techniques for selecting displayed repertoires. To this end, the discovery of new low molecular weight peptides is becoming increasingly important, not only as molecular tools for the understanding of protein-protein interactions but also for the generation of lead compounds. PMID- 11281833 TI - Surface display on gram positive bacteria. AB - Heterologous surface display on Gram-positive bacteria was first described almost a decade ago and has since then developed into an active research area. Gram positive bacterial surface display has today found a range of applications, in immunology, microbiology and biotechnology. Live bacterial vaccine delivery vehicles are being developed through the surface display of selected foreign antigens on the bacterial surfaces. In this field, "second generation" vaccine delivery vehicles are at present being generated by the addition of mucosal targeting signals through co-display of adhesins, in order to achieve targeting of the live bacteria to immunoreactive sites to thereby increase immune responses. Engineered Gram-positive bacteria are further being evaluated as novel microbial biocatalysts with heterologous enzymes immobilized as surface exposed on the bacterial cell surface. A discussion has started whether bacteria can find use as new types of whole-cell diagnostic devices since single-chain antibodies and other variants of tailor-made binding proteins can be displayed on bacteria. Bacteria with increased binding capacity for certain metal ions can be created and potential environmental or biosensor applications for such recombinant bacteria as biosorbents are being discussed. This article explains the basis of Gram-positive bacterial surface display, and discusses current uses and possible future trends of this emerging technology. PMID- 11281834 TI - The baculovirus expression system as a tool for generating diversity by viral surface display. AB - It has become a major goal of molecular biologists, biochemists, and immunologists to be able to modulate the structure of proteins, in order to increase their antigenicity, alter their biological properties and/or explore their function. Based on the concept of bacterial phage display, by which proteins are being selected and analyzed in conjunction with their genetic information, eukaryotic systems have been investigated for their use in generating biomolecular diversity. The advantage of posttranslational modification and the possible harbouring of structural complex proteins has lead scientists to include eukaryotic systems in the wide field of molecular design. The ideal expression vectors for surface display are eukaryotic viruses, that allow large gene insertions, efficiently present foreign proteins on the particle surface, are easy to propagate and, if possible, not pathogenic to humans. By inserting peptides into a native virus coat protein or by expressing foreign proteins as coat protein fusion proteins or linked to specific anchor domains it becomes possible to display polypeptides of interest on the surface of replicating particles. A variety of different strategies are currently under investigation in order to utilize the baculovirus insect cell expression system for efficient display on the surface of virus particles as well as on the surface of virally infected insect cells. Increasing the transfection efficiency, optimizing cloning procedures, and establishing applicable selection methods have lead to the development of a powerful tool for drug screening and ligand screening. PMID- 11281835 TI - High-throughput screening of surface displayed gene products. AB - With the human genome project approaching completion, there is a growing interest in functional analysis of gene products. The characterization of large numbers of proteins, their expression patterns and in vivo localisations, demands the use of automated technology that maintains a logistic link to the encoding genes. As a complementary approach, phage display is used for recombinant protein expression and the selection of interacting (binding) molecules. Cloning of libraries in filamentous bacteriophage or phage mid vectors provides a physical link between the expressed protein and its encoding DNA sequence. High-throughput technology for automated library handling and phage display selection has been developed using picking-spotting robots and a module for pin-based magnetic particle handling. This system enables simultaneous interaction screening of libraries and the selection of binders to different target molecules at high throughput. Target molecules are either displayed on high-density filter membranes (protein filters) or tag-bound to magnetic particles and can be handled as native ligands. Binding activity is confirmed by magnetic particle ELISA in the microtitre format. The whole procedure from immobilisation of target molecules to confirmed clones of binders is automatable. Using this technology, we have selected human scFv antibody fragments against expression products of human cDNA libraries. PMID- 11281836 TI - Peptidomics: the comprehensive analysis of peptides in complex biological mixtures. AB - Progress in the sequencing of genomes has resulted in an increasing demand for a functional analysis of gene products in order to understand the underlying physiology. Proteomics has established itself as a highly valuable technology for producing functionally related data in an unparalleled fashion, but is methodologically restricted to the analysis of proteins with higher molecular masses (>10 kDa). The development of a technology which covers peptides with low molecular weight and small proteins (0.5 to 15 kDa) was necessary, since peptides, amongst them families of hormones, cytokines and growth factors, play a central role in many biological processes. To summarise the technologies used for this approach the term "peptidomics" is introduced. In this article, we present the rationale and first results of a novel, universal peptide display approach for the analysis and visualisation of peptides and small proteins from biological samples. Special attention is given to samples derived from extracellular fluids such as blood plasma and cerebrospinal fluid. Additionally, a high throughput identification procedure for the analysis of peptides in their native and processed molecular form is outlined. PMID- 11281837 TI - Recent developments in sequence selective minor groove DNA effectors. AB - DNA is a well characterized intracellular target but its large size and sequential nature make it an elusive target for selective drug action. Binding of low molecular weight ligands to DNA causes a wide variety of potential biological responses. In this respect the main consideration is given to recent developments in DNA sequence selective binding agents bearing conjugated effectors because of their potential application in diagnosis and treatment of cancers as well as in molecular biology. Recent progress in the development of cross linked lexitropsin oligopeptides and hairpins, which bind selectively to the minor groove of duplex DNA, is discussed. Bis-distamycins and related lexitropsins show inhibitory activity against HIV-1 and HIV-2 integrases at low nanomolar concentrations. Benzoyl nitrogen mustard analogs of lexitropsins are active against a variety of tumor models. Certain of the bis-benzimidazoles show altered DNA sequence preference and bind to DNA at 5'CG and TG sequences rather than at the preferred AT sites of the parent drug. A comparison of bifunctional bizelesin with monoalkylating adozelesin shows that it appears to have an increased sequence selectivity such that monoalkylating compounds react at more than one site but bizelesin reacts only at sites where there are two suitably positioned alkylation sites. Adozelesin, bizelesin and carzelesin are far more potent as cytotoxic agents than cisplatin or doxorubicin. A new class of 1,2,9,9a-tetrahydrocyclo propa[c]benz[e]indole-4-one (CBI) analogs i.e., CBI-lexitropsin conjugates arising from the latter leads are also discussed.A number of cyclopropylpyrroloindole (CPI) and CBI-lexitropsin conjugates related to CC-1065 alkylate at the N3 position of adenine in the minor groove of DNA in a sequence specific manner, and also show cytotoxicities in the femtomolar range. The cross linking efficiency of PBD dimers is much greater than that of other cross linkers including cisplatin, and melphalan. A new class of PBD-lexitropsin conjugates is also discussed. Certain functional models of the bleomycins (BLMs) show outstanding DNA cleavage activity comparable with that of and positionally distinct from natural BLM. PMID- 11281838 TI - Sequence recognition of DNA by lexitropsins. AB - Lexitropsins are modular polyamide molecules that are designed to "read" the base sequence of DNA. Lexitropsins constructed of three types of subunits--pyrrole, imidazole and hydroxypyrrole--allow full recognition of DNA base sequences. Structural studies have revealed the atomic basis of this specificity. Theoretical studies have explored the effectiveness of lexitropsins in targeting a given sequence within a genome, and have been used to analyze and improve lexitropsin design. PMID- 11281839 TI - Selection of an unnatural peptide library for dsDNA binding. AB - More and more, nucleic acids have become prime targets in the development of new compounds, able to control gene expression. For the development of new sequence selective dsDNA binding ligands, one can learn a lot from existing models such as, lexitropsins, combilexins and actinomycin D. This analysis, together with the knowledge of the details on protein-DNA interactions, has inspired the assembly of unnatural amino acids in a combinatorial way to generate a dsDNA recognition library. The first selection round has led to the selection of new DNA binding molecules, which may lead on the long run to the discovery of new DNA binding motifs. PMID- 11281840 TI - DNA minor groove alkylating agents. AB - Recent work on a number of different classes of anticancer agents that alkylate DNA in the minor groove is reviewed. There has been much work with nitrogen mustards, where attachment of the mustard unit to carrier molecules can change the normal patterns of both regio- and sequence-selectivity, from reaction primarily at most guanine N7 sites in the major groove to a few adenine N3 sites at the 3'-end of poly(A/T) sequences in the minor groove. Carrier molecules discussed for mustards are intercalators, polypyrroles, polyimidazoles, bis(benzimidazoles), polybenzamides and anilinoquinolinium salts. In contrast, similar targeting of pyrrolizidine alkylators by a variety of carriers has little effect of their patterns of alkylation (at the 2-amino group of guanine). Recent work on the pyrrolobenzodiazepine and cyclopropaindolone classes of natural product minor groove binders is also reviewed. PMID- 11281841 TI - Targeting double stranded DNA with peptide nucleic acid (PNA). AB - PNA (peptide nucleic acid) is a DNA mimic with a pseudopeptide (polyamide) backbone which can be used to target double stranded DNA with high sequence specificity. PNA therefore has great potential in the development of biomolecular tools for manipulation of DNA as well as for the development of DNA targeted gene therapeutic drugs. The status of this field is discussed in terms of PNA binding modes and mechanism as well as applications. PMID- 11281842 TI - Sequence specific recognition of ligand-DNA complexes studied by NMR. AB - The last few years have represented an accelerated accumulation in detailed information about ligand-DNA interactions. A collected view of literature information is essential for advancing our understanding of the principles of ligand-DNA recognition, utilizing this valuable information for construction of a modeling database, and eventually the rational design of DNA-binding ligands possessing desired properties. This review is concentrated on structure-based information on ligand-oligodeoxyribonucleotide (DON) complexes published since 1995, especially focusing on the results obtained from NMR structure elucidation. The discussions emphasize the sequence specific recognition of novel binding motifs or binding modules of ligand molecules rather than specific atomic details. A comprehensive list of DNA binding ligands are discussed in the text and are also summarized in a table. The DNA sequences that are recognized by specific ligand molecules as studied by NMR are annotated in a figure to provide a clear view of target selection. This review also briefly describes NMR methods for characterization and structure elucidation of ligand-DNA complexes. PMID- 11281843 TI - Bioactive sesquiterpenes produced by fungi: are they useful for humans as well? AB - Higher fungi are characterised by the production of macroscopic fruiting bodies to generate and to distribute their spores. These fruiting bodies are under constant threat of other organisms feeding on them. As a consequence these organisms developed a number of strategies for protection, one of them is the production of toxins. The fungal subdivision Basidiomycotina produce toxic sesquiterpenes many of them are derived from the protoilludane skeleton. This skeleton is transformed and rearranged to a large number of compounds. Some of these sesquiterpenes show interesting biological properties which may be attractive for medicinal chemistry. The overview describes the different types of bioactive fungal sesquiterpenes derived from humulene known to date in Basidiomycotina and their formation. The metabolites are discussed according to their sesquiterpene skeleton and the different metabolites are compared. Where available biological activities concerning antifungal, antibacterial, cytotoxic and enzyme inhibition data are given. Special attention was paid for the different activities of these metabolites and the attempts made to use them in medicinal chemistry. The question whether metabolites produced for the self protection of fungi can be used for pharmaceutical applications for humans will be addressed and discussed. PMID- 11281844 TI - Ligand design for alpha(1) adrenoceptors. AB - An area of continuing interest in medicinal chemistry is the design, synthesis and pharmacological evaluation of ligands which bind at adrenoceptor subtypes, which include alpha(1A), alpha(1B), alpha(1D); alpha(2A), alpha(2B), alpha(2C); beta(1), beta(2), beta(3) and possibly beta(4) subtypes. The selective blockade or stimulation of these receptor subtypes is of on-going pharmacological and medicinal interest. However, the design principles for ligand differentiation at these subtypes still need further development. This review focuses on alpha(1) adrenoceptors with a concentration on literature over the past five years. Structural, physiological and therapeutic aspects of the alpha(1A), alpha(1B) and alpha(1D) subtypes are discussed together with ligands binding to these receptor subtypes. Approaches to alpha(1) adrenoceptor ligand design based on known ligands and on receptor docking are evaluated. A new combined approach using pharmacophores and receptor docking affords possibilities for deeper insights into achieving small molecule binding selectivity. PMID- 11281845 TI - Reactions of morphine derivatives with phenyliodo(III)diacetate (PIDA): synthesis of new morphine analogues. AB - The reactions of morphine and its derivatives with phenyliodo(III)diacetate (PIDA) have been studied. This methodology has not been introduced to morphine alkaloids, despite the fact that such a strategy would ensure dearomatization of the electrophilic aromatic ring of morphine derivatives leading to nucleophilic ortho-quinoidal structures with potential pharmacological interest. The products, formed in regio- and diastereoselective or diastereospecific reactions, carry mixed-acetal or 1,3-dioxolane moieties. At low concentrations 6a has mu-opioid agonist character but in higher concentrations showed a non receptorial antagonist effect on isolated mouse vas deferens. PMID- 11281846 TI - Recent advances in NMR: expanding its role in rational drug design. AB - The technology of nuclear magnetic resonance spectroscopy continues to advance at a rapid pace. Recent improvements in gradient technology and the coupling of NMR to various chromatography methods will provide new opportunities in drug discovery. These opportunities include rapid high throughput screening to determine the receptor-bound conformations of small organic ligands. NMR coupled to liquid chromatography has opened a new door to the quantitative and qualitative analysis of complex mixtures including metabolites extracted from body fluids and extracts containing various natural products. This review will focus on the following four advances in NMR technology: 1) pulse-field gradient (PFG) NMR, 2) SAR (structure activity relationship) by NMR, 3) LC-NMR (liquid chromatography), and 4) application of membrane models for the study of neuropeptides. The information content available to medicinal chemists from each experiment will be discussed. PMID- 11281847 TI - Agonists at the alpha4beta2 nicotinic acetylcholine receptors: structure-activity relationships and molecular modelling. AB - Agonists of the alpha4beta2 nicotinic acetylcholine receptors have been synthesised as potential drugs for treatment of a variety of diseases. In this review, the published nicotinic agonists are presented and, on the basis of the molecular structure, the compounds are divided into three compound classes, nicotinoids (structurally close to nicotine), bicyclic compounds (structurally close to epibatidine and anatoxin-a), and analogues of imidacloprid (structurally close to the insecticide imidacloprid). The structure-activity relationships are discussed within and in between the classes. On the basis of computational studies of ligands for the nicotinic acetylcholine receptors the structure activity relationships are discussed and a possible binding mode suggested. The binding mode encompasses: (A) an interaction between an anionic site in the receptor and the protonated nitrogen atom in the ligand, (B) a hydrogen bond between a hydrogen bond donor in the receptor and a hydrogen bond acceptor in the ligand, (C) an interaction between a pi-system (heteroaromatic ring, carbonyl bond) in the ligand and another pi-system or a positively charged amino acid residue in the binding site, (D) a pi-cation interaction between aromatic residues in the receptor binding site and the protonated nitrogen atom in the ligand, and (E) steric interactions of positive and negative character around the aliphatic and the heteroaromatic part of the ligand. PMID- 11281848 TI - Development of response-selective agonists of human C5a anaphylatoxin: conformational, biological, and therapeutic considerations. AB - Numerous studies on the relationship between the structure and function of peptide agonists derived from the biologically active, C-terminal region of human C5a anaphylatoxin have been reported over the past decade. These studies have been performed with the objective of parlaying this structure-function information into the design of peptide/peptidomimetic modulators of C5a receptor (C5aR)-mediated function. In this review, we describe a rational approach for the development of conformationally biased, decapeptide agonists of C5a and described how these stabilized and specific conformational features relate to the expression of specific C5a-like activities in vitro and in vivo. The therapeutic potential of such response-selective C5a agonists is discussed and underscored by the results of one such response-selective C5a agonist that was used in vivo as an effective molecular adjuvant capable of generating antigen-specific humoral and cellular immune responses. Finally, we describe the synthesis of a new generation of highly response-selective, conformationally biased C5a agonist and discuss the in vitro and in vivo biologic results that so indicate this biologic selectivity. PMID- 11281849 TI - Structure-activity relationships of multidrug resistance reversers. AB - Multidrug resistance, MDR, is a major obstacle in the chemotherapeutic treatment of cancer. MDR can be reversed by drugs that vary widely in their chemical structure and main biological action. Many efforts are directed to find out the relationships between the structure and MDR reversal effect of these drugs. In this review we try to summarize the results of a variety of studies on identification of structure-activity relationships, SARs, and quantitative SARs, QSARs, of different MDR reversing drugs. As any reasonable (Q)SAR study relies on a real or putative presentation about the mechanism of action of the studied compounds, the most significant MDR mechanisms revealed till now are shortly discussed. Special attention is paid to P-glycoprotein, P-gp, related MDR as the most experimentally and clinically tested form of drug resistance. The currently proposed models of P-gp functioning and mechanisms of MDR modulation are presented. Problems that can arise in (Q)SARs studies are discussed in advance to allow the reader to judge on possible pitfalls. The physicochemical and structural properties of MDR modulators as found by different research groups are commented and summarized. From the discussed studies it can be concluded that the careful selection of relevant structural and biological data processed with appropriate QSAR and especially 3D-QSAR methods, is a promising approach to structure-activity studies of MDR reversers. PMID- 11281850 TI - The 2-chlorotrityl resin: a worthy addition to the medicinal chemist's toolbox. AB - The polystyrene-based 2-chlorotrityl resin was originally used in the synthesis of peptides using an Fmoc-amino acid/carboxyl-linked protocol. While traditionally employed to prepare a number of biologically active peptides, the resin has received increasing attention as a support for the synthesis of pseudopeptide and non-peptide molecules recently. This review focuses on 2 chlorotrityl resin-supported synthesis of small molecules that collectively display a broad range of biological activities. PMID- 11281851 TI - Current therapies and emerging targets for the treatment of diabetes. AB - Concurrent with the spread of the western lifestyle, the prevalence of all types of diabetes is on the rise in the world's population. The number of diabetics is increasing by 4-5% per year with an estimated 40-45% of individual's over the age of 65 years having either type II diabetes or impaired glucose tolerance. Since the signs of diabetes are not immediately obvious, diagnosis can be preceded by an extended period of impaired glucose tolerance resulting in the prevalence of beta-cell dysfunction and macrovascular complications. In addition to increased medical vigilance, diabetes is being combatted through aggressive treatment directed at lowering circulating blood glucose and inhibiting postprandial hyperglycemic spikes. Current strategies to treat diabetes include reducing insulin resistance using glitazones, supplementing insulin supplies with exogenous insulin, increasing endogenous insulin production with sulfonylureas and meglitinides, reducing hepatic glucose production through biguanides, and limiting postprandial glucose absorption with alpha-glucosidase inhibitors. In all of these areas, new generations of small molecules are being investigated which exhibit improved efficacy and safety profiles. Promising biological targets are also emerging such as (1) insulin sensitizers including protein tyrosine phosphatase-1B (PTP-1B) and glycogen synthase kinase 3 (GSK3), (2) inhibitors of gluconeogenesis like pyruvate dehydrogenase kinase (PDH) inhibitors, (3) lipolysis inhibitors, (4) fat oxidation including carnitine palmitoyltransferase (CPT) I and II inhibitors, and (5) energy expenditure by means of beta 3 adrenoceptor agonists. Also important are alternative routes of glucose disposal such as Na+-glucose cotransporter (SGLT) inhibitors, combination therapies, and the treatment of diabetic complications (eg. retinopathy, nephropathy, and neuropathy). With may new opportunities for drug discovery, the prospects are excellent for development of innovative therapies to effectively manage diabetes and prevent its long term complications. This review highlights recent (1997 2000) advances in diabetes therapy and research with an emphasis on small molecule drug design (275 references). PMID- 11281852 TI - The immunomodulatory effects of anti-thyroid drugs are mediated via actions on thyroid cells, affecting thyrocyte-immunocyte signalling: a review. AB - The mechanism of action of the immunosuppressive effects of antithyroid drugs has remained a matter of controversy, despite our earlier contention that such effects in vivo were indirect; ie., it was our view that the drugs were acting on the thyroid cells, reducing their thyroid hormone production and other activities, with a consequent reduction in thyrocyte-immunocyte signalling. The reduction in the activation of CD4+ cells,the increased number and activation of CD8+ (and CD8+CDllb+) cells, and the reduction of soluble interleukin-2 receptors, thought once to be direct effects of the medication, are now shown to be due to amelioration of the hyperthyroidism. Thus the reduction in thyroid hormone production induced by the drugs is central to these actions. In addition, the iodination of thyroglobulin is inhibited by these agents, which may affect antigen presentation by the thyrocyte. Furthermore, there is now evidence that the thionamides interfere with thyrocyte expression of such molecules as Class I antigen, interleukin-1, interleukin-6, prostaglandin E2, and heat shock protein. The expression of thyrocyte Class II antigen is probably not inhibited by these drugs, although one group has shown that lectin-stimulated thyrocyte Class II expression is diminished by this treatment; this group postulated that this effect might be mediated by reduced interferon gamma production by T lymphocytes, but in vitro experiments do not corroborate this proposal. In any event, the actions as described of the effects of antithyroid drugs on the thyroid cells (particularly normalization of thyroid function) would certainly suffice to explain the diminution of thyroid antibodies (including thyroid stimulating antibody), the reduced immunological response, and the increased remission rate in Graves disease as a consequence of antithyroid drug therapy, without the need to invoke a direct immunosuppressive effect. PMID- 11281853 TI - Prostaglandins and lipid modification. AB - Postaglandins(PG) and low-density lipoproteins (LDL) both are playing a key role in atherogenesis. Their interaction at the local vascular level is of central relevance in plaque formation and progression. Details of these complex actions however, still need to be elucidated. Lipoproteins are influencing the PG production of arterial wall cells and platelets, while PGs in turn have been shown to regulate lipoprotein receptor binding and entry into the arterial wall. Modification of LDL severely influences arterial wall trapping and foam cell formation. During LDL-modification, isoprostanes, a new family of compounds generated by free radical catalysed action, independent of cyclooxygenase, are formed. 8-epi PGF(2alpha) the most well known member exerts a great variety of proatherogenic actions, among them vasoconstriction and platelet activation; it also serves as a mitogen and stimulator of endothelin release. The influence of various eicosanoids on lipoprotein modification, however, has not been assessed yet. PMID- 11281854 TI - Pharmacology of phosphoinositides, regulators of multiple cellular functions. AB - Inositol phospholipids represent a small fraction of the phospholipids present in all cellular membranes with remarkable importance in regulating various cell functions. They are synthesized from phosphatidylinositol by sequential phosphorylations on the several hydroxyls of the inositol ring to create polyphosphoinositides that function either as docking sites to promote formation of molecular signaling complexes, or serve as precursors for soluble inositol polyphosphates that act as diffusible intracellular messengers. Phosphoinositides are involved in the control of many processes, including membrane traffic, endo- and exocytosis, mitogenesis and apoptosis. Pharmacological tools have helped to clarify many details of phosphoinositide metabolism and have unveiled the roles of these lipids in the control of specific signaling pathways. However, because of their pleiotropic functions it has been questionable whether pharmacological manipulation of inositide formation and metabolism can be of therapeutic value. This review briefly summarizes the means by which inositide functions have been pharmacologically manipulated, and discusses possibilities for specifically targeting certain aspects of their regulatory functions. PMID- 11281856 TI - Editorial. PMID- 11281855 TI - Characterization of protein glycoforms with N-linked neutral and phosphorylated oligosaccharides: studies on the glycosylation of endoglucanase 1 (Cel7B) from Trichoderma reesei. AB - Using anion-exchange chromatography the catalyticdomain of endoglucanase 1 (Cel7B) from Trichoderma reesei was resolved in multiple fractions with different isoelectric points, presumably related to different glycoforms of the enzyme. The protein fractions were analysed using lectins and electrospray MS. Isolated N glycans were analysed by fluorophore-assisted carbohydrate electrophoresis and amine-adsorption HPLC. The results show that this particular preparation contained at least 14 different glycoforms. The major isoform contained only one GlcNAc, presumably N-linked, and one mannose, most probably O-linked to serine/threonine at a separate site. Except for a small population containing Man(5)GlcNAc(2)+1-2 Man, the rest of the protein had negatively charged phosphate containing N-glycans. All glycoforms contained at least one O-linked mannose residue. The increased negative charge of the protein, introduced by oligosaccharide phosphorylation, is the most probable reason for the different isoelectric points and the occurrence of multiple peaks during purification. PMID- 11281857 TI - Expectations of health care: promoted, managed or shared? AB - Volume, costs and content of medical care depend on professional and public expectations. The UK National Health Service (NHS) removed price barriers to access, so depressed expectations became an important factor in cost control. In USA, professional control of care business inflated expectations and costs. Managed care in the NHS failed to rationalize care because managers seem even less trustworthy than clinicians as arbiters of rational expectations in contexts of underfunding. Rational expectations depend on restored trust, mutual and managerial respect for expertise of both clinicians and patients, and transcendence of the provider-consumer model for value production in medical care. PMID- 11281858 TI - The use of evidence by health care user organizations. AB - OBJECTIVES: To explore the use of research evidence by consumer and patient organizations and the extent to which their goals and activities are consistent with evidence-based health care and patient-centred care. DESIGN: A mailed survey, telephone and face-to-face interviews of leaders of organizations representing health care users. SETTING: Norway. PARTICIPANTS: Sixty-nine of 109 questionnaires that were mailed were included in our analysis and approximately 20 interviews were conducted with representatives of general consumer and patient advocacy groups and interest groups that focus on particular diseases or disabilities. MEASUREMENTS: Information was collected on the goals of the organizations, the nature of their everyday work, the extent to which research information is required in this work and how research information is accessed and appraised. RESULTS: An important focus of many user groups is peer support. They tend to emphasize experience-based knowledge. A total of 82% of the respondents said that they often or sometimes had use for research results in their work. Research-based information is most often obtained indirectly through physicians or researchers. CONCLUSIONS: Norwegian health care user organizations do not appear to promote evidence-based health care. To the extent that they help to disseminate scientific information, they appear to do so uncritically, relying on few sources and traditional authorities. PMID- 11281859 TI - Development of a patient decision aid for choice of surgical treatment for breast cancer. AB - PURPOSE: A patient decision aid for the surgical treatment of early stage breast cancer was developed and evaluated. The rationale for its development was the knowledge that breast conserving therapy (lumpectomy followed by breast radiation) and mastectomy produce equivalent outcomes, and the current general agreement that the decision for the type of surgery should rest with the patient. METHODS: A decision aid was developed and evaluated in sequential pilot studies of 18 and 10 women with newly diagnosed breast cancer who were facing a decision for breast conserving therapy or mastectomy. Both qualitative (general reaction, self-reported anxiety, clarity, satisfaction) and quantitative (knowledge and decisional conflict) measures were assessed. RESULTS: The decision aid consists of an audiotape and workbook and takes 36 min to complete. Based on qualitative comments and satisfaction ratings, 17 of 18 women reported a positive reaction to the decision aid, and all 18 reported that it helped clarify information given by the surgeon. Women did not report an increase in anxiety and 17 of 18 women were either satisfied or very satisfied with the decision aid. CONCLUSION: This pilot study supports the hypothesis that this decision aid may be a helpful adjunct in the decision for surgical management of early stage breast cancer. We are currently conducting a randomized trial of the decision aid versus a simple educational pamphlet to evaluate its efficacy as measured by knowledge, decisional conflict, anxiety and post-decisional regret. PMID- 11281861 TI - A randomized controlled trial of information-giving to patients referred for coronary angiography: effects on outcomes of care. AB - OBJECTIVE: To assess the impact of providing an educational videotape, 'Treatment Choices for Ischaemic Heart Disease: a Shared Decision-Making Program Videotape,' to patients referred for coronary angiography compared with standard patient physician decision making (usual care). STUDY DESIGN: Randomized controlled clinical trial. SETTING: University Hospital and Veterans Affairs Hospital. PATIENTS: A consecutive sample of 217 patients referred for coronary angiography were randomized to receive 'usual care' or to receive the videotape in addition to standard patient physician decision making (videotape): 109 completed the study (50% completion rate). MAIN OUTCOME MEASURES: Knowledge of coronary artery disease, satisfaction, self-reported physical and mental health functioning, and the proportion of patients who were referred for coronary revascularization. RESULTS: Compared with patients who received 'usual care,' those who received the videotape were more knowledgeable (mean score 83 vs. 58%; P < 0.0001) but less satisfied with their treatment (79 vs. 88%; P = 0.038). There were no significant differences between the videotape and 'usual care' groups with respect to satisfaction with the decision making process (mean score 73 vs. 77%; P = 0.37), satisfaction with the decision made (mean score 73 vs. 78%; P = 0.28), physical functioning (38 vs. 38%; P = 0.76), mental health functioning (49 vs. 49%; P = 0.94), or in referral for coronary revascularization (OR 0.60; 95% CI 0.22-1.65; P = 0.33). CONCLUSION: Although the educational videotape increased patients' knowledge level, it was associated with a decrease in their level of satisfaction with treatment. Before there is wide-spread dissemination of this technology, advocates should demonstrate its effectiveness in everyday practice. PMID- 11281860 TI - What can patients do to improve health care? AB - OBJECTIVE: To give an overview of the value of different interventions for increasing the role of individual patients in improving the quality of care provision. SEARCH strategy: Medline searches and manual searches in medical journals covering the period from 1980 until June 1997. INCLUSION CRITERIA: Studies reporting descriptions and evaluations of seven types of interventions that aim to help integrate the needs and preferences of individual patients into health care provision. DATA EXTRACTION: The following information was extracted: assumptions underlying the interventions; resources needed for development and implementation; and acceptability to clinicians. MAIN RESULTS: Several interventions for increasing patients' roles in health care could be successful in clinical practice, such as feeding forward patient data to clinicians, interactive patient education and feedback to health care providers about patients' evaluations of care. The available research focuses on feedback methods. Insights into the benefits and limitations of the use of the different interventions for improving care are limited. CONCLUSION: The active role that patients' views play in the contact with a care provider is often neglected. Promising interventions for the empowerment of individual patients require further development and evaluation. PMID- 11281862 TI - User involvement and the NHS reforms. AB - The policy of 'user involvement' in the UK National Health Service emerged during the 1990s along with the reforms that created an internal market. Despite the official rhetoric, progress has been limited. Critics suggest that, not only was the policy flawed in its conception by the construction of service users as consumers and the conflation of consumerism with empowerment, but collaborative models of involvement have tended to legitimate rather than challenge existing provision. Some commentators have questioned the value of user involvement initiatives and proposed that alternative approaches, such as a strengthening of procedural rights or alignment with broader political campaigns, would be more appropriate. The low prominence given in the recent Government White Paper The New NHS1 to the contribution of service users, however, represents less of an ideological shift than a concentration on other, in the Government's view, more pressing priorities: namely, a concern to address the problems of public legitimacy and low staff morale by engaging in greater public participation and giving health professionals a more central role. The result has been a weakening of the users' voice by a conflation of user involvement with public participation and giving health professionals the authority to define users' needs for them. Service users risk, not only having their contribution devalued, but losing the right to an independent and distinctive voice. There is a real danger that the issues of user involvement will not be included on local agendas and the disparities between provision and need and between professionals' and users' views will increase. PMID- 11281864 TI - A survey of women's preferences regarding alternative surgical treatments for menorrhagia. AB - BACKGROUND: Menorrhagia represents a major health burden to a large number of women in the UK. A range of medical and surgical treatments is now available for the condition, but each has its own advantages and limitations. METHODS: A postal survey was undertaken of women with apparently uncomplicated menorrhagia referred to one of two NHS hospitals. The aim was to elicit women's preferences for the characteristics of surgical treatment and, specifically, to assess how they traded-off the characteristics of hysterectomy and minimal access surgery. RESULTS: A total of 221 women returned their questionnaire and were included in the study (59% of those sent out). The characteristics most frequently rated as 'very important' were getting back to usual activities as soon as possible (57%), experiencing the least pain and discomfort (46%), spending a short time in hospital (46%) and stopping periods for good (43%). The single characteristic most frequently rated as the most important by women was stopping periods for good (28%). Similar proportions of women preferred abdominal hysterectomy (43%) and endometrial resection (41%) when these were described to them. When asked to think about specific treatments of which they were aware, strong positive (43%) and negative (42%) preferences were indicated. Women's main information source was their general practitioner (70%), but only 44% considered themselves well informed about menorrhagia and its treatment. CONCLUSIONS: Many women referred to hospital with menorrhagia have conflicting objectives for treatment, feel under informed about therapies and have not formed treatment preferences. It is important to develop well-structured and accessible sources of information for women and methods to elicit their treatment preferences, and to evaluate the impact of such tools on treatment selection and outcomes. PMID- 11281865 TI - Communication and decision-making in labour: do birth plans make a difference? AB - OBJECTIVES: To assess usage of birth plans, and examine differences in social and obstetric characteristics, and intrapartum experiences of women who did and did not use a birth plan. DESIGN: Population-based survey distributed by hospitals and home birth practitioners, 6-7 months post-natally. SETTING AND PARTICIPANTS: Women who gave birth in Victoria, Australia over a 2-week period in September, 1993, excluding those who had a stillbirth or neonatal death. MAIN OUTCOME MEASURES: Use of a written birth plan; perceived helpfulness, advantages and disadvantages of birth plans; relationship between use of birth plans and overall rating of intrapartum care, and involvement in decision-making. RESULTS: Twenty per cent of women (270/1336) had prepared a written birth plan and discussed it with caregivers. Women who made use of a birth plan were more likely to be satisfied with pain relief (OR = 1.74[1.3-2.3]), but did not differ from women not completing a birth plan in terms of their overall rating of intrapartum care, or involvement in making decisions about their care. CONCLUSIONS: The lack of association between use of a written birth plan and variables assessing women's views of intrapartum care suggest there are insufficient grounds for continuing to advocate a policy of encouraging pregnant women to complete written birth plans, unless it is within the context of a well-designed randomized trial able to provide further evidence regarding their effectiveness. PMID- 11281863 TI - Quality of care from the patients' perspective: from theoretical concept to a new measuring instrument. AB - INTRODUCTION: Patient views on quality of care are of paramount importance with respect to the implementation of quality assurance (QA) and improvement (QI) programmes. However, the relevance of patient satisfaction studies is often questioned because of conceptual and methodological problems. Here, it is our belief that a different strategy is necessary. OBJECTIVE: To develop a conceptual framework for measuring quality of care seen through the patients' eyes, based on the existing literature on consumer satisfaction in health care and business research. RESULTS: Patient or consumer satisfaction is regarded as a multidimensional concept, based on a relationship between experiences and expectations. However, where most health care researchers tend to concentrate on the result, patient (dis)satisfaction, a more fruitful approach is to look at the basic components of the concept: expectations (or 'needs') and experiences. A conceptual framework - based on the sequence performance, importance, impact - and quality judgements of different categories of patients derived from importance and performance scores of different health care aspects, is elaborated upon and illustrated with empirical evidence. CONCLUSIONS: The new conceptual model, with quality of care indices derived from importance and performance scores, can serve as a framework for QA and QI programmes from the patients' perspective. For selecting quality of care aspects, a category-specific approach is recommended including the use of focus group discussions. PMID- 11281866 TI - Using conjoint analysis to elicit the views of health service users: an application to the patient health card. AB - OBJECTIVE: To demonstrate the application of conjoint analysis (CA) for eliciting the views of health service users. METHODS: A CA study was conducted alongside a randomized controlled trial evaluating the introduction of a patient health card (PHC). The PHC was evaluated with respect to three other aspects of general practice: number of days between making a non-urgent appointment and seeing a doctor; waiting time in reception between the time of the appointment and seeing a doctor; and whether the patient is usually seen by the doctor of their choice. A postal questionnaire was sent to 100 individuals from a general practice in Inverurie, Scotland. RESULTS: Seventy-five individuals returned the questionnaire, of whom 51 answered the CA section. The PHC was the least important of the attributes considered. The number of days between making a non urgent appointment and seeing a doctor was considered to be the most important. A 1-day reduction in the number of days to appointment was four and a half times more important than having a PHC; a 1-minute reduction in waiting time in the reception area was three and a half times more important than having a PHC; and seeing a doctor of choice was over three times more important than having a PHC. Satisfaction or utility scores for different ways of providing a general practice service also indicated that priority should be given to reducing waiting time to see a doctor or reducing waiting time in reception. CONCLUSIONS: While the PHC is a significant and positive predictor of satisfaction in general practice, it is less important than the other three attributes considered. More generally, CA appears to be a potentially useful instrument for eliciting the views of health service users. PMID- 11281867 TI - Why DISCERN? PMID- 11281868 TI - DISCERN in practice. PMID- 11281869 TI - Editorial. PMID- 11281870 TI - Who are you, and who are we? Looking through some key words. AB - The terminology used to describe individuals who come into contact with health services is problematic. Many of the most commonly used words, for example, patient, consumer, user, carry overtones or imply characteristics, which may be misleading or unacceptable to those to whom they are applied. PMID- 11281871 TI - Minority ethnic community participation in needs assessment and service development in primary care: perceptions of Pakistani and Bangladeshi people about psychological distress. AB - OBJECTIVES: To promote community participation in exploring perceptions of psychological distress amongst Pakistani and Bangladeshi people, in order to develop appropriate services. DESIGN: Training and facilitation of resident community members (as community project workers), to define and conduct qualitative research involving semistructured interviews in their own communities, informing primary care led commissioning and service decision making. Setting A socio-economically disadvantaged inner-city locality in the UK. Participants One-hundred and four South Asian people (49 of Pakistani and 55 of Bangladeshi origin), interviewed by 13 resident community members. RESULTS: All community project workers completed training leading to a National Vocational Qualification, and successfully executed the research. Most study respondents located their main sources of stress within pervasive experience of racism and socio-economic disadvantage. They were positive about 'talking' and neutral listening as helpful, but sought strategies beyond non-directive counselling services that embraced practical welfare advice and social support. The roles of primary health care professionals were believed to be restricted to physical ill health rather than personal distress. The importance of professionals' sex, age, ethnicity and social status were emphasized as affecting open communication. Practical recommendations for the re-orientation and provision of services were generated and implemented in response to the findings, through dialogue with a primary care commissioning group, Health and Local Authority, and voluntary agencies. CONCLUSIONS: The work illustrates the feasibility and value of a community participation approach to research and service development in addressing a challenging and neglected area of minority ethnic health need. It offers one model for generating responsive service change in the context of current health policy in the UK, whilst also imparting skills and empowering community members. The study findings emphasize the need to recognize the social contexts in which distress is experienced and have implications for effective responses. PMID- 11281872 TI - The effects of an 'explicit' values clarification exercise in a woman's decision aid regarding postmenopausal hormone therapy. AB - OBJECTIVE: To evaluate the incremental effect of a graphic weigh-scale values clarification exercise to explicitly consider the personal importance of the benefits versus the risks in a woman's decision aid regarding postmenopausal hormone therapy. DESIGN: Randomized controlled trial. Intervention Decision aid including information on options, benefits and risks, and their probabilities either followed by: (1) a graphic weigh-scale values clarification exercise to explicitly consider the personal importance of each benefit and risk; or (2) a summary of the main benefits and risks to implicitly consider benefits versus the risks. SAMPLE: Two-hundred and one women aged 50-69 years from Ottawa, Canada, who had never used hormone therapy. OUTCOME: Perceived clarity of values, a sub scale of the decisional conflict scale; congruence between personal values of benefits and risks (measured on 0-10 importance rating scale) and choices (accept, decline, unsure regarding preventive hormone therapy [HRT]) using discriminant function analysis. RESULTS: There were no statistically significant differences between interventions in perceived clarity of values and overall congruence between values and choices. Amongst those choosing HRT, there was a trend in those exposed to the graphic weigh-scale exercise to have better congruence between values and choices compared to implicit values clarification (P = 0.06). CONCLUSION: The use of the graphic weigh-scale exercise in a decision aid conveys no overall short-term benefit. Further study is needed to specifically determine effects in those changing the status quo and on the quality of patient-practitioner communication and persistence with decisions. PMID- 11281873 TI - Attitudes to randomized clinical trials amongst out-patients attending a medical oncology clinic. AB - OBJECTIVE: To assess the understanding of and attitudes towards randomized clinical trials amongst patients attending oncology out-patient clinics. DESIGN: Cross-sectional survey. SUBJECTS: Patients attending medical oncology out-patient clinics at a Sydney teaching hospital. MAIN OUTCOME: Patients' willingness to participate in a randomized clinical trial. RESULTS: Sixty consecutive patients were surveyed. The mean age was 55.2 (SD 14) years. Eighty-eight per cent of respondents thought that patients should be asked to participate in trials testing new treatments, however, only a third would consider participating in a randomized trial themselves. If a trial was endorsed by an independent cancer information service such as the NSW Cancer Council, 72% of respondents would be more likely to participate. Knowledge about randomized trials was not high. Respondents scored a median of 3 out of 7 (interquartile range, 2-4) correct answers to a series of questions about randomized trials. Patients willing to participate in a randomized trial were more likely to perceive the doctor favourably (P = 0.05), less likely to perceive trials as experimental (P = 0.05) and less likely to perceive trials as representing an inconvenience or loss of control (P = 0.09). CONCLUSIONS: Understanding amongst patients of the need for and mechanisms of randomized clinical trials is not good. This may contribute to the difficulties investigators face in seeking consent for clinical trials. Evaluation of new strategies to educate the public and patients about randomized trials is needed. Involvement of consumers in the design and conduct of clinical trials and evaluation of strategies to improve doctors' communication of clinical trial information is also required. PMID- 11281874 TI - Italian forum of Europa Donna: a survey of breast cancer associations. AB - Europa Donna is the first European woman's movement against breast cancer. It is a coalition of breast cancer associations and individual women and is active in 20 nations. Europa Donna is not intended to replace existing organizations. Rather, it provides a focus for the exchange of information and experiences between members and serves as a moving force for combined action. It promotes public awareness of breast cancer, advances in research and good clinical practice. In Italy there are more than 200 active breast cancer associations. The Italian forum of Europa Donna was formed in 1996. Between June and September 1996 a postal survey was conducted to research the characteristics and activities of the various breast cancer associations in Italy in order to help, inform and promote future initiatives of the Italian forum of Europa Donna. A total of 213 breast cancer associations were sent a postal questionnaire. Ninety-five of them (44.6%) participated in the survey. The results show that the breast cancer associations in Italy vary markedly in terms of their structure and organization. The associations perceive a variety of deficiencies in the prevention, diagnosis and treatment of breast cancer within the Italian National Health Service and they offer a wide range of services themselves. Their views of the relevance of the 10 goals of Europa Donna vary. In this paper, we discuss the implications of the low rate of participation in the survey and the heterogeneity of the breast cancer associations' structures, activities and views for the future activities of the Italian forum of Europa Donna. PMID- 11281876 TI - The role of doctors, patients and managers in priority setting decisions: lessons from the 'Child B' case. AB - The difficulties in treating patients with life threatening illnesses were highlighted by the case of Jaymee Bowen, a 10-year-old girl with leukaemia who was refused funding for a second bone marrow transplant in 1995. Jaymee's case was widely reported at the time and came to epitomize the dilemmas of rationing in the United Kingdom's National Health Service. In reality, the paediatricians who had cared for Jaymee based their decision on clinical rather than financial considerations, and the media reporting of the case failed to reflect the complexities of the issues involved. The case also demonstrated the difficulties of determining the best interests of children and of obtaining their consent to treatment. There were disagreements between Jaymee's father and the paediatricians who had treated her about how her best interests could be served and this led to a breakdown of trust and the search for further opinions. This highlighted the rise of consumerism in health care and the challenge to doctors and managers to justify their decisions and to give reasons for these decisions. The common theme in Jaymee's story is the need for greater openness in decisions on priority setting and stronger safeguards for patients. PMID- 11281875 TI - The relationship between expectations and satisfaction: a qualitative study of patients' experiences of surgery for gynaecological cancer. AB - It is important that a patient perspective is introduced to the identification and measurement of the outcomes of health care. The aim of this study was to use qualitative methods to examine the presence or absence of expectations prior to the experience of health care and the relationship between expectations, satisfaction and dissatisfaction in a group of women undergoing surgery in a large teaching hospital. Nineteen women with a diagnosis of gynaecological cancer were interviewed on two occasions, before and after surgery. A thematic analysis was undertaken. The results suggest that there is not a clear relationship between expectations and satisfaction. Women had different levels of expectations about different types of care and different aspects of care. Unfulfilled expectations did not lead to less satisfaction. The women were able to express satisfaction either with the care overall or with specific aspects of care, as well as being able to distinguish aspects of care with which they were dissatisfied. PMID- 11281877 TI - HELP - the Health Education Library for People, India's first Consumer Health Education Resource Centre. PMID- 11281878 TI - Research and development in the NHS: how can you make a difference? PMID- 11281879 TI - Information for Health. PMID- 11281880 TI - Editorial. PMID- 11281881 TI - The price of autonomy. AB - The randomized controlled trial produces a clash of ethical principles with the need for informed consent (autonomy) in conflict with the principles of beneficence and justice. Informed consent is one of the major rate-limiting factors of recruitment and this delays the discovery of life-saving treatments indirectly. Whilst supporting the concept of non-exploitation we wish to challenge the prevailing dogma by asking the awkward question 'what is the price of autonomy?'. Using breast cancer as an example we have developed a decision model with explicit assumptions allowing numerical values to be fed into a mathematical equation, which calculates the cost in lives. With conservative assumptions we estimate that the price of autonomy is 2500 lives over a 10-year period in the United Kingdom alone. We issue the challenge to health policy makers and ethicists to survey public opinion to determine the value placed on autonomy in the war against cancer. PMID- 11281882 TI - Patient empowerment in the United States: a critical commentary. AB - Whilst there is no consensus amongst analysts regarding how best to define 'patient empowerment', at the very least, this concept entails a re-distribution of power between patients and physicians. Empowered patients attempt to take charge of their own health and their interactions with health care professionals. Empowerment can occur at different levels (micro, meso, and macro) and patients have different ideas about what it means to 'take charge' and 'be empowered'. Some patients simply want to be given information about their conditions whilst others want to have full control over all medical decision-making. Some empirical evidence suggests that active patient participation in health care is associated with better patient outcomes. This field is ripe for future studies which both help to develop theoretical models of patient empowerment and articulate the conditions under which patient empowerment occurs. PMID- 11281883 TI - Patient expectations and health-related quality of life. AB - OBJECTIVE: The measurement of health-related quality of life (HRQL) has become increasingly common in health services research. Whilst useful, its focus on behaviour, capacities and activities means that it remains relatively specific. This paper explores the possibility of extending the evaluation of health by considering the concept of patients' expectations. DESIGN: In-depth and semi structured interviews were used to explore the concept of expectations from the patients' perspective. Patients' expectations were then used in the construction of a two-part questionnaire. SETTINGS AND PARTICIPANTS: Expectations were explored with a group of 33 cardiac patients. The resulting questionnaire was given to 400 cardiac patients in a large teaching hospital in London. RESULTS: Patients identified a range of expectations which related to their health and seemed to represent the desired results of their hospital stay. Comparison of the content of patient expectations with a commonly used generic measure of HRQL, the Short-Form 36 (SF-36), found some overlap but indicated that patients seemed to adopt a broader approach to their health. Expectations that patients identified were used to construct two scales to measure expectations and their evaluation. The internal consistency of these scales was 0.82 and 0.88, respectively. CONCLUSION: The study indicates the potential complexity of the concept of expectations and the need for further exploration. It also demonstrates the feasibility of constructing standardized scales to measure patient expectations. Whilst conceptually different from HRQL such standardized expectations scales could provide a useful adjunct to HRQL measurement and provide a meaningful context for the interpretation of HRQL data. PMID- 11281884 TI - Is 'shared decision-making' feasible in consultations for upper respiratory tract infections? Assessing the influence of antibiotic expectations using discourse analysis. AB - OBJECTIVES: To examine the discourse of consultations in which conflict occurs between parents and clinicians about the necessity of antibiotics to treat an upper respiratory tract infection. To appraise the feasibility of shared decision making in such consultations. DESIGN: A qualitative study using discourse analysis techniques. SETTING: A general practice with 12 500 patients in an urban area of Cardiff, Wales. PARTICIPANTS: Two consultations were purposively selected from a number of audiotaped sessions. The consultations took place during normal clinics in which appointments are booked at 7-minute intervals. The practitioner is known to be interested in involving patients in treatment decisions. METHOD: Discourse analysis was employed to examine the consultation transcripts. This analysis was then compared with the theoretical competencies proposed for 'shared decision-making'. RESULTS: The consultations exhibit less rational strategies than those suggested by the shared decision-making model. Strong parental views are expressed (overtly and covertly) which seem derived from prior experiences of similar illnesses and prescribing behaviours. The clinician responds by emphasizing the 'normality' of upper respiratory tract infections and their recurrence, accompanied by expressions that antibiotic treatment is ineffective in 'viral' illness - the suggested diagnosis. The competencies of 'shared decision-making' are not exhibited. CONCLUSIONS: The current understanding of shared decision-making needs to be developed for those situations where there are dis-agreements due to the strongly held views of the participants. Clinicians have limited strategies in situations where patient treatment preferences are opposed to professional views. Dispelling 'misconceptions' by sharing information and negotiating agreed management plans are recommended. But it seems that communication skills, information content and consultation length have to receive attention if such strategies are to be employed successfully. PMID- 11281885 TI - Pilot study of an information aid for women with a family history of breast cancer. AB - OBJECTIVE: To develop and pilot study an information aid for women with a family history of breast cancer. DESIGN, SETTING AND PARTICIPANTS: The information aid, consisting of a booklet and audiotape, was developed by a multi-disciplinary team of health care professionals, breast cancer survivors and their relatives. Women with no personal history of breast cancer, on the waiting list for a familial breast cancer clinic at either of two centres, who could read English, were eligible for the pilot study which consisted of three sets of mailed questionnaires. MAIN OUTCOME MEASURES: The baseline questionnaires included: demographic information: the Breast Cancer and Heredity Knowledge Scale (BCHK); psychological measures (the State-Trait Anxiety Inventory [STAI], Centre for Epidemiologic Studies Depression Scale [CES-D] and an item about breast cancer worry), and an item about breast cancer risk perception. Immediately after reviewing the information aid, participants completed a satisfaction survey, the risk perception and cancer worry items and a checklist about their personal family history. The third set of questionnaires, completed 2-4 weeks after reviewing the aid, was identical to the first. Patients then attended their scheduled clinic visit and an objective hereditary breast cancer risk assessment was made by the genetic counselling team. RESULTS AND CONCLUSIONS: Of 97 eligible women who were contacted, 67 completed all three sets of questionnaires. Overall, women were very satisfied with the aid and 96% would recommend it to other women. There was a highly significant improvement in their knowledge scores after they reviewed the aid. Anxiety and depression did not change and there was a decline in breast cancer worry. Risk perception did not change significantly. Ninety per cent of women completed their personal family history checklist accurately. Several important improvements have been made in the information aid and it will now be evaluated in the community. PMID- 11281886 TI - Consumer collaboration, patient-defined outcomes and the preparation of Cochrane Reviews. AB - OBJECTIVES: To identify the extent to which the Cochrane Collaboration involves consumers (patients, carers, patient and non-patient members of patient/consumer organizations) as members of Cochrane Review Groups (CRGs); to explore the emphasis CRGs place on identifying and collecting information on outcomes identified by patients as being important indicators of quality and effectiveness of treatment and care ('patient-defined outcomes'). METHOD: A postal questionnaire designed by The College of Health, a UK patient organization, was sent in January 1998 to all CRGs registered with the Cochrane Collaboration on 1 January 1998 (n = 42). RESULTS AND CONCLUSIONS: Replies were received from 35 CRGs, a response rate of 83% and 33 questionnaires (79%) were completed. The survey revealed that CRGs varied in the extent to which they had recruited consumer members: almost one third of respondents said their CRG did not have any consumer members. There was also no apparent consensus across CRGs on the importance attached to identifying and collecting information on patient-defined outcomes or on integrating such information into their activities. It is hoped that differences between CRGs may inform discussions as to whether and how the Cochrane Collaboration might address the issue of patient-defined outcomes in the future. PMID- 11281887 TI - Information is the key to patient choice. PMID- 11281888 TI - Guidebook for Ulcerative Colitis. PMID- 11281889 TI - Inside Out: visual art and emotional decision-making in medicine. PMID- 11281890 TI - Editorial. PMID- 11281891 TI - Reflections on health care consumerism: insights from feminism. AB - Health care consumerism is a movement concerned with patients' interests in health care, crucially those that are repressed or partly repressed by dominant interest-holders. Like feminism, health care consumerism attracts dislike and confusion as well as enthusiasm. But just as the voicing of women's repressed interests leads to their gradual acceptance by dominant interest-holders, so does the voicing of patients' repressed interests. PMID- 11281892 TI - Participation in treatment decision-making by women with early stage breast cancer. AB - OBJECTIVE: This study aimed to assess the way women treated for early stage breast cancer perceived the treatment selection process. The purpose was to understand more fully patients' experiences of the decision process and their preferences for participation in treatment decisions. SETTING AND PARTICIPANTS: The study informants were 40 women, treated at a teaching hospital in Sydney Australia, who were interviewed face to face 1 year after their first treatment for stage I or stage II breast cancer. METHODS: This study used a qualitative approach, based on the analysis of interview transcripts. The main areas covered were how the informants' treatment decisions were made and their preferences for participation in treatment decisions. Content and thematic analyses were conducted with findings presented using verbatim quotations for illustration. RESULTS AND CONCLUSIONS: Many of the informants who preferred not to participate in decisions also failed to recognize the need for value judgements (as well as medical expertise) in the decision-making process. Some informants believed they ought to be responsible for the consequences whilst others did not. Difficulties were identified in patient utilization of medical information for treatment decision-making, and also in establishing preferences for the risks and benefits of treatments where few patients had prior experience of the potential outcomes. The findings indicate that patient participation in treatment decision-making is a more complex issue than simply giving patients information and choices. Ways of enhancing patients' involvement in the treatment selection process are discussed. PMID- 11281893 TI - Patient participation groups in general practice in the National Health Service. AB - General practice remains the organizational hub of first level health services in the United Kingdom. Patient participation groups are probably the most well known model for public participation in this setting and, although still not widespread, they have been a slowly expanding area of development for almost three decades. This paper sets out to critically asses patient participation groups in general practice by considering the context of their development and reviewing the research literature about groups. Critical issues needing more study and key methodological challenges are then discussed. Patient participation groups have been a somewhat shifting and contested phenomenon, embracing trends and changing as policy priorities have changed over the years. There is some evidence to think that they might have potential as a local element within a public participation strategy in the National Health Service. However, the field studies are very limited and more research of a better quality is needed. The state of knowledge is not adequate to be able to say with any confidence if or how such groups should be developed. A better understanding is needed of the public's perspectives on this and other models of participation. There are many other questions to do with patient participation groups' purpose, equitable access, and effectiveness that need to be addressed. The methodological challenges include issues of how to involve all stakeholders in the research process; and how to study less tangible aspects of general practice organization, such as culture and power, that effect the public's participation. PMID- 11281894 TI - Investing in early detection versus intensive treatment for breast cancer: a study of the Israeli public priorities. AB - OBJECTIVE: Public opinion has become one of the main inputs in setting priorities, rationing and allocating health resources. The present study focuses on the priorities of the Israeli public in allocating marginal breast cancer funds between early detection in the healthy majority of the population and intensive treatment of the sick minority in need. DESIGN: A sample of 2030 individuals representing the Israeli Jewish urban population aged 45-75 was interviewed in 1993/4. A full sit-down interview collected information on several health related issues. RESULTS: Sixty-one per cent chose to direct the funds to early detection efforts and 35% chose to direct the funds to treating the sick. Four per cent of the population could not decide. Higher education and better health are related to choosing early detection; religious observance is related to choosing the treatment option; and men and older persons tend, more than women and younger respondents, to be undecided. CONCLUSIONS: Whilst the majority of the population tend to follow a cost-effectiveness rationale in the marginal use of breast cancer funds, for more than a third of the population efficiency is not thus important, and they adhere to entitlement based upon a need-equity principle in allocating health resources. PMID- 11281895 TI - 'It's the best of two evils': a study of patients' perceived information needs about oral steroids for asthma. AB - OBJECTIVES: To explore the sources of patients' knowledge about the potential side-effects of oral steroids prescribed to treat asthma. METHODS: Seventeen in depth interviews were conducted with patients taking prescribed oral steroid medication (prednisolone) for asthma. The interviews were transcribed verbatim and the data organized according to common themes. RESULTS: All the respondents acknowledged they had no choice but to take oral steroids but they wanted to be informed about the potential side-effects. Respondents reported that they had not received sufficient information about side-effects from their general practitioner (GP). Information was sought from both medical sources (pharmacists and asthma nurses) and non-medical sources (friends and family, self-help groups and the media) to supplement their knowledge. The conclusions drawn about the risks of taking oral steroids were also influenced by respondents' existing beliefs. CONCLUSIONS: Respondents drew upon information about oral steroids from a variety of professional and lay sources. The findings add weight to calls for doctors and patients to share their respective knowledge in consultations. Developing an understanding of the views of GPs about the provision of information about side-effects would help to identify any perceived barriers to a more open exchange of information in the consultation. PMID- 11281896 TI - Developing consumer-led maternity services: a survey of women's views in a local healthcare setting. AB - OBJECTIVE: This paper describes a prospective study of women's views and experiences of maternity services. The aim was to examine the way women make choices and decisions about maternity care and the factors which influence decision making, with a view to developing services which best meet the needs of the population. Patient choice issues reviewed included: choice of place of birth, choice of lead professional and choices in labour management. DESIGN, SETTING AND PARTICIPANTS: A cohort of women attending maternity booking clinics, within the catchment area of Peterhead Maternity Unit (PMU) in north-east Scotland, were surveyed by means of postal questionnaires at three stages during their contact with maternity services. A subset of women also took part in in depth interviews. RESULTS: Not all women were given information about all the available options for place of birth and many women were unclear of the differences between them. Factors influencing choice of place of birth can change, with the medical aspects of maternity care becoming more important as the pregnancy progresses. Women rated the importance of seeing the same staff at antenatal visits highly, but were less concerned with their ability to choose which professional to see. More importance was attached to being able to choose a particular midwife rather than a particular obstetrician. Women's choices with regard to labour management were largely met. Insufficient information, however, was provided about choices in pain relief. CONCLUSIONS: The survey revealed the importance of locally based research, involving all stakeholders, in developing services which best meet the needs of a population. PMID- 11281897 TI - The role and activity of an Italian volunteer organization providing information and emotional support for patients with cancer. PMID- 11281898 TI - Editorial. PMID- 11281899 TI - Public involvement in health care priority setting: an overview of methods for eliciting values. AB - There is increasing interest, in the UK and elsewhere, in involving the public in health care priority setting. At the same time, however, there is evidence of lack of clarity about the objectives of some priority setting projects and also about the role of public involvement. Further, some projects display an apparent ignorance of both long-standing theoretical literature and practical experience of methodologies for eliciting values in health care and related fields. After a brief examination of the context of health care priority setting and public involvement, this paper describes a range of different approaches to eliciting values. These approaches are critically examined on a number of dimensions including the type of choice allowed to respondents and the implications of aggregation of values across individuals. Factors which affect the appropriateness of the different techniques to specific applications are discussed. A check-list of questions to be asked when selecting techniques is presented. PMID- 11281900 TI - Public involvement in health care priority setting: an economic perspective. AB - BACKGROUND: Public involvement in health care decision making and priority setting in the UK is being promoted by recent policy initiatives. In 1993, the British Medical Association called for public consultation where rationing of services was to be undertaken. The approach to priority setting advocated by many health economists is the maximization of quality adjusted life years (QALYs). Typically, for a particular health care programme, the QALY calculation takes account of four features: (1) the number of patients receiving the programme, (2) the survival gain, (3) the gain in quality of life and, (4) the probability of treatment success. Only one feature, that relating to quality of life, is based upon public preferences. If the QALY is to be used as a tool for health care resource allocation at a societal level then it should incorporate broader societal preferences. METHODS: This study used an interview-based survey of 91 members of the general public to explore whether the traditional QALY maximization model is a good predictor of public responses to health care priority setting choices. RESULTS AND CONCLUSIONS: Many respondents did not choose consistently in line with a QALY maximization objective and were most influenced by quality of life concerns. There was little support for health care programmes that provided a prognostic improvement but left patients in relatively poor states of health. The level of respondent engagement in the survey exercise was not sensitive to the provision of supporting clinical information. PMID- 11281901 TI - Public participation and marginalized groups: the community development model. AB - OBJECTIVES: To develop ways of reaching house-bound people and enabling them to give their views in planning and monitoring health and social care. STRATEGY: HealthLINK - a project based in a community health council - explored ways of involving older house-bound people in the London Borough of Camden, in planning and monitoring health and social care using community development techniques. RESULTS: HealthLINK set up an infrastructure to enable house-bound people to have access to information and to enable them to give their views. This resulted in access for health and local authorities to the views of house-bound older people and increased the self esteem and quality of life of those who became involved. CONCLUSIONS: Community development approaches that enable an infrastructure to be established may be an effective way of reaching marginalized communities. However, there are tensions in this approach between the different requirements for public involvement of statutory bodies and of users, and between representation of groups and listening to individual voices. PMID- 11281902 TI - A randomized community intervention trial to increase awareness and knowledge of the role of periconceptional folate in women of child-bearing age. AB - OBJECTIVES: To determine the effect of a consumer-directed information campaign to increase knowledge of folate for the prevention of neural tube defects among women of child-bearing age, and to measure women's recall of sources of information and knowledge about folate. DESIGN: A community randomized trial. SETTING: Three matched pairs of geographically distinct Local Government Areas in the state of Victoria, Australia. INTERVENTION: Printed information recommending folate intake to decrease the risk of neural tube defects was disseminated to women of child-bearing age in three of the Local Government Areas selected randomly. MAIN OUTCOME MEASURE: The proportion of women aware of the association between folate and spina bifida. RESULTS: Of 1197 women interviewed prior to the intervention, 12.4% (adjusted for the cluster and population sampling unit) were aware of folate and neural tube defects. After the intervention, there was not only a significant background increase of 3.4% (P=0.02) in folate awareness since the pre-intervention survey (n=603), but also a significant additional increase of 4.0% (P=0.04) owing to the intervention itself (n=603). Only 70% of women who were aware of folate knew the correct timing. CONCLUSIONS: The provision of printed educational material can increase folate awareness among women of child bearing age. A comprehensive, long-term and ongoing health promotion campaign including such material, together with initiatives by relevant health service providers and the food industry, could best address the current low levels of folate awareness among women of child-bearing age. PMID- 11281903 TI - Decision-making role preferences and information needs: a comparison of colorectal and breast cancer. AB - OBJECTIVE: An exploratory study has been carried out to examine decision-making role preferences and information needs for a sample of people with colorectal cancer (n=48). The work replicated a larger study carried out for women with breast cancer (n=150), and this paper compares and contrasts findings for both disease groups. DESIGN: A cross-sectional design was employed, involving structured interviews. The main variables investigated were decision-making preference (using a decisional role preference card sort), perceived decisional role and information need (using an information needs questionnaire). RESULTS: The majority (78%) of the colorectal cancer patients preferred to play a passive role in decision making, in contrast to 52% of women with breast cancer in previous work. Eighty per cent of the colorectal sample and 61% of the women with breast cancer perceived that the doctor had made treatment decisions. Priority information needs for both groups related to cure, spread of disease and treatment options. CONCLUSIONS: The two most striking findings from the comparison between the two disease groups relate to the differences in decision making role preferences and the similarities in information needs. The process of involving people with colorectal cancer in treatment decision making warrants further investigation. The similarity in information needs of the two disease groups has implications for health care professionals providing information to people with cancer. PMID- 11281904 TI - Patient participation in service improvement: the initial Measures Project experience. AB - BACKGROUND: 'Measures' is a 2-year, part-funded European Community project. It began in March 1998 and has, as its primary foci, the aims of continuing to improve the quality and efficiency of patient care by, (1) re-examining healthcare processes and (2) determining how best to take advantage of information technology. Central to this has been patient involvement in the project. Initially the project is focused on aspects of rheumatology, diabetes and spinal injury services based across Pinderfields and Pontefract NHS Hospitals Trust. APPROACH: Patient questionnaires, interactive cross-specialty and individual specialty focus groups have been employed. The re-enforcement of relationships with voluntary organizations has featured prominently. CONCLUSION: Initial perception from patients participating in the process is highly positive and supportive. Specific areas to be considered for change have been addressed. PMID- 11281905 TI - The price of autonomy. PMID- 11281906 TI - Editorial: special conference issue. PMID- 11281907 TI - Clinical decision-making in the context of chronic illness. AB - This paper develops a framework to compare clinical decision making in relation to chronic and acute medical conditions. Much of the literature on patient physician decision making has focused on acute and often life-threatening medical situations in which the patient is highly dependent upon the expertise of the physician in providing the therapeutic options. Decision making is often constrained and driven by the overwhelming impact of the acute medical problem on all aspects of the individual's life. With chronic conditions, patients are increasingly knowledgeable, not only about their medical conditions, but also about traditional, complementary, and alternative therapeutic options. They must make multiple and repetitive decisions, with variable outcomes, about how they will live with their chronic condition. Consequently, they often know more than attending treatment personnel about their own situations, including symptoms, responses to previous treatment, and lifestyle preferences. This paper compares the nature of the illness, the characteristics of the decisions themselves, the role of the patient, the decision-making relationship, and the decision-making environment in acute and chronic illnesses. The author argues for a different understanding of the decision-making relationships and processes characteristic in chronic conditions that take into account the role of trade-offs between medical regimens and lifestyle choices in shaping both the process and outcomes of clinical decision-making. The paper addresses the concerns of a range of professional providers and consumers. PMID- 11281908 TI - The medical visit context of treatment decision-making and the therapeutic relationship. AB - The ascendance of the autonomy paradigm in treatment decision-making has evolved over the past several decades to the point where few bioethicists would question that it is the guiding value driving health-care provider behaviour. In achieving quasi-legal status, decision-making has come to be regarded as a formality largely removed from the broader context of medical communication and the therapeutic relationship within which care is delivered. Moreover, disregard for individual patient preference, resistance, reluctance, or incompetence has at times produced pro forma and useless autonomy rituals. Failures of this kind, have been largely attributed to the psychological dynamics of the patients, physicians, illnesses, and contexts that characterize the medical decision. There has been little attempt to provide a framework for accommodating or understanding the larger social context and social influences that contribute to this variation. Applying Paulo Freire's participatory social orientation model to the context of the medical visit suggests a framework for viewing the impact of physicians' communication behaviours on patients' capacity for treatment decision making. Physicians' use of communication strategies can act to reinforce an experience of patient dependence or self-reliance in regard to the patient physician relationship generally and treatment decision-making, in particular. Certain communications enhance patient participation in the medical visit's dialogue, contribute to patient engagement in problem posing and problem-solving, and finally, facilitate patient confidence and competence to undertake autonomous action. The purpose of this essay is to place treatment decision-making within the broader context of the therapeutic relationship, and to describe ways in which routine medical visit communication can accommodate individual patient preferences and help develop and further patient capacity for autonomous decision making. PMID- 11281909 TI - Doctor-patient communication patterns in breast cancer adjuvant therapy discussions. AB - OBJECTIVE: To identify variables within the patient-oncologist communication pattern that impact overall patient comprehension and satisfaction within the breast cancer adjuvant therapy (AT) setting. SETTING AND PARTICIPANTS: Fifty patients were recruited from a number of academic and community-based oncology practices. Fifteen oncologists participated. MAIN VARIABLES: Three communication variables were identified: percentage of total utterances spoken by the patient, percentage of total physician utterances that were coded as affective (i.e. emotional), and total number of questions asked by the patient during the consultation. Knowledge and satisfaction were assessed by a variety of outcome measures, including knowledge items and satisfaction as measured by VASs, the satisfaction with decision scale and the decisional conflict scale. RESULTS: The level of patient knowledge about breast cancer and satisfaction with the clinical encounter showed a tendency to correlate with the variables measuring aspects of patient-physician communication style. Patients who spoke more or asked more questions tended to be more knowledgeable whilst patients whose physicians used more affective language tended to know less but to be more satisfied with their clinical encounter. CONCLUSIONS: In order to optimize patients' degree of comprehension and satisfaction with their breast cancer adjuvant therapy, physicians need to increase their affective participation in clinical encounters whilst encouraging patients to ask questions and to actively participate in the decision-making process. PMID- 11281910 TI - Involving patients in decisions regarding preventive health interventions using the analytic hierarchy process. AB - Practice guidelines that recommend active patient involvement in decisions about preventive health interventions are becoming increasingly common. These decisions frequently involve difficult trade-offs between competing risks and benefits that require easily accessible information about the expected outcomes, superb doctor patient communication, and effective integration of objective outcome data with individual values and preferences. Successful implementation of recommendations for shared decision-making in preventive health care will require the development of efficient methods for making these complex decisions in busy practice settings. This article describes how the analytic hierarchy process, a multiple criteria decision-making method, could facilitate successful implementation of shared decision-making regarding preventive health care in clinical practice. The method is illustrated using recent guidelines for colorectal cancer screening for average risk patients issued by the American Gastroenterological Association. PMID- 11281911 TI - Practical issues in assisting shared decision-making. AB - To facilitate treatment decision-making, one aims to provide information, present it in a way that makes it as easy as possible to understand, and to help the decision-maker through the cognitive processes that result in a treatment decision. Decision aids aim to accomplish just these goals and this paper identifies practical issues that we have encountered in creating a decision aid for men with early stage prostate cancer. We highlight the results of studies we carried out to provide an empirical basis for the decision aid that we were developing. Several of the studies were designed to identify what information key players (health professionals, patients and family members) thought was important for the decision-making process. Another investigation studied methodological considerations in identifying important information. The final study focused on presentation issues. These studies, designed to explore what information was considered important, found great variability among both health care professionals involved in treating patients with prostate cancer (urologists, radiation oncologists, nurses in cancer clinics, and radiation technologists) and among the patients, themselves. The studies also showed that not all information contained within a typical category is of equal importance. A methodological study showed that the information that patients deem to be important to their decision depends on whether they are rating the information that could be provided, or questions that could be answered. Finally, presentation studies showed that the various formats used in presenting quantitative information are processed with differing degrees of accuracy and ease. Each of the above results has implications for those creating decision aids; these implications are highlighted. PMID- 11281912 TI - A treatment trade-off based decision aid for patients with locally advanced non small cell lung cancer. AB - Purpose To describe the structure and use of a decision aid for patients with locally advanced non-small cell lung cancer (LA-NSCLC) who are eligible for combined-modality treatment (CMT) or for radiotherapy alone (RT). METHODS: The aid included a structured description of the treatment options and trade-off exercises designed to help clarify the patient's values for the relevant outcomes by determining the patient's survival advantage threshold (the increase in survival conferred by CMT over RT that the patient deemed necessary for choosing CMT). Additional outcome measures included each patient's strength of treatment preference, decisional conflict, objective understanding of survival information, decisional role preference, and evaluation of the aid itself. RESULTS: Twenty five patients met the eligibility criteria for study. Of these, seven declined the decision aid because they had a clear treatment preference (four chose CMT and three chose RT). The remaining 18 participants completed the decision aid; 16 chose CMT and two chose RT. All 18 patients wished to participate in the decision to some extent. All patients reported that using the decision support was useful to them and recommended its use for others. No patient or physician reported that the aid interfered with the physician-patient relationship. Patients' 3-year survival advantage thresholds, and their median survival advantage thresholds, were each strongly correlated with their strengths of treatment preference (rho=0.80, P < 0.001 and rho=0.77, P < 0.001, respectively). For all but one patient, either their 3-year or median survival threshold was consistent with their final treatment choice. Eight patients reported a stronger treatment preference after using the decision aid. CONCLUSIONS: We conclude that a treatment trade-off based decision aid for patients with locally advanced non small cell lung cancer is feasible, that it demonstrates internal consistency and convergent validity, and that it is favourably evaluated by patients and their physicians. The aid seems to help patients understand the benefits and risks of treatment and to choose the treatment that is most consistent with their values. PMID- 11281914 TI - Supporting and resourcing treatment decision-making: some policy considerations. AB - This paper considers some of the policy implications of issues raised during a conference about treatment decision-making in the clinical encounter held in Hamilton, Ontario in May 1999. Policies promoting patient participation in treatment decision-making need to be flexible enough to ensure that they are appropriate across the range of contexts in which health care decisions are made and acceptable to people with diverse preferences and abilities. They should also be formulated in consideration of other health policies and of available resources. Policies of informing people and involving them in decisions about their care are unlikely to be simple to implement. Various strategies might be needed to support them. These include the development of appropriate skills among health professionals and in the general population, the use of interventions to encourage people to play more active roles in decisions about their health care, the provision of decision aids for people facing specific decisions and the provision and accreditation of more general information resources and services. If information and other facilitators of patient participation in decision-making are seen as integral to good quality health care, then funding should be made available for them. This will, however, have opportunity costs. Policy makers' decisions about how much health care funding should be invested in which strategies should be underpinned by good research evidence about the effects that different types of intervention have on a range of outcomes for individuals, health care systems and populations. The knowledge on which current policies are based is limited. The development of future policies will be enhanced if policy makers invest in critical conceptual thinking, reflective practice, imaginative development work and good quality evaluative research. PMID- 11281913 TI - Lessons learned from the Decision Board: a unique and evolving decision aid. AB - One session of the conference was devoted to the presentation of different types of decision aids. This paper reports the experience and lessons learned through the development and use of the Decision Board. This is a uniquely interactive decision aid administered by the clinician during the medical consultation. The instrument has been developed in a number of clinical contexts, primarily regarding treatment options for cancer patients. Studies have shown the instrument to improve patient understanding and facilitate the shared decision making process. Randomized trials are ongoing, evaluating the addition of the Decision Board to the traditional medical consultation. The instrument continues to evolve to meet patients' need for information and flexibility in presentation. Computer-based versions of the Decision Board are currently being developed. PMID- 11281915 TI - Editorial. PMID- 11281916 TI - Regulated competition and citizen participation: lessons from Israel. AB - OBJECTIVE: To investigate the relationship between health system structure and citizen participation, in particular whether increased reliance on competition encourages or depresses citizen involvement. SETTING: The case of Israel's ongoing health reform, based on regulated competition among sick funds, is examined. DESIGN: Interviews with government officials and representatives of consumer groups; analysis of policy documents, judicial rulings, public surveys and journalistic accounts. RESULTS: The Israeli reform is based in large measure on a regulated competition model, in which citizens have free choice among highly regulated competing sick funds. At the same time, the reform process has been accompanied by legal, institutional and political frameworks, as well as significant interest group activity, all aimed at increasing public input into processes of health policy making and implementation. The Israeli case, it is argued, lends support to the proposition that citizen participation (voice) and individual choice (exit) are complementary, rather than alternative, modes of ensuring citizen influence over health services. The question is whether the development of multiple avenues for citizen involvement represents disarray or a healthy social learning process regarding the running of the health system. CONCLUSION: This paper expresses cautious optimism that citizen participation is a projection of a healthy social learning process, and suggests directions for public policy to encourage this outcome. PMID- 11281917 TI - Eliciting patients' preferences for adjuvant chemotherapy in breast cancer: development and validation of a bedside decision-making instrument in a French Regional Cancer Centre. AB - In developed countries, the physician-patient relationship is moving from a paternalistic model to new decision-making models that take patient preferences into account. OBJECTIVES: Our aim was to develop a Decision Board (DB) and to test its acceptability in a French Regional Cancer Centre regarding the decision on whether or not to use chemotherapy after surgery in postmenopausal women with breast cancer. This paper presents the development process for this instrument and reports the pretesting phase, as well as the corresponding results. METHODS: A working group was created with oncologists, psychologists and economists. Following the first phase, i.e. the development process, a first version of the instrument was presented to health professionals. Their feedback led to important modifications of the instrument. The DB was then presented to experienced patients, which resulted in slight changes. The second phase consisted of pretesting the comprehension, internal and across-time consistency of the DB on healthy volunteers. RESULTS: The DB was pretested in a group of 40 healthy volunteers. Eighteen respondents chose chemotherapy and 22 chose not to have chemotherapy. Comprehension rates were very high (>/=87.5%). Internal consistency was assessed considering option attitudes based on outcomes and option attitudes based on process. Women shifted their choices in a predictable way. Across-time consistency was appraised using the test-retest method with Visual Analog Scales. The Intraclass Correlation Coefficient was 0.97. DISCUSSION-CONCLUSION: Due to cultural differences, the DB developed in our French Cancer Centre is quite different from the DBs previously developed elsewhere. Our instrument showed good comprehension and consistency properties, which are corroborated by the DB literature. Whether our DB is acceptable for patients with breast cancer must still be tested. Patients' reactions will tell us which type of decision-making model is at work. Further research is needed in order to explore the shared decision-making process and clarify the concept. PMID- 11281918 TI - Five cases, four actors and a moral: lessons from studies of contested treatment decisions. AB - The case of Jaymee Bowen (child B) illustrated the conflict that may arise over treatment decisions in the National Health Service (NHS). This article reviews four further cases involving disagreement between patients and families on the one hand, and health authorities on the other, and a fifth case in which a health authority questioned the treatment decision of a medical specialist. The cases illustrate the rise of consumerism in health care and the challenge for health authorities in weighing the claims of individual patients against the needs of communities. They also demonstrate the increasing role of lawyers and the courts in resolving disputes over treatment decisions. Clinicians were closely involved in all cases, both in recommending treatment options and in serving as independent advisers when disputes arose. The findings presented here indicate that there is a need to strengthen the process of decision-making in cases of this kind and to make greater use of evidence in informing decisions. In future, decision-making needs to be characterized by openness, reason giving, an appeals procedure and regulation of the process to ensure that these conditions are met. The funders of health care also need to consider each individual in his or her own right while also using their resources for the benefit of the population as a whole. PMID- 11281919 TI - Development and evaluation of a decision aid for patients with stage IV non-small cell lung cancer. AB - Although guidelines for treating stage IV non-small cell lung cancer suggest that the patient's values should be considered in decision-making, there are no practical tools available to assist them with their decision-making. OBJECTIVE: To develop and evaluate a decision aid that incorporates patient values. DESIGN AND SAMPLE: (1) Before/after evaluation with patients referred to a regional cancer centre. (2) Mailed survey of thoracic surgeons and respirologists in Ontario. INTERVENTION: An audio-tape guided individuals to review a booklet describing stage IV non-small cell lung cancer, its impact and possible coping strategies, treatment options, benefits and risks, and examples of the decision making of others. Patients then used a worksheet to consider and communicate personal issues involved in the choice, including: personal values using a 'weigh scale'; questions; preferred role in decision-making; and predisposition. MEASURES: (1) Patient questionnaires eliciting knowledge, the decision, decisional conflict and acceptability of the decision aid. (2) Physician questionnaires eliciting attitudes toward the decision aid. RESULTS: (1) Twenty of 30 patients used the aid in decision-making. Users thought that the aid was acceptable and significantly improved their knowledge about options and outcomes (P < 0.001), and reduced their decisional conflict (P < 0.001). (2) The majority of the 29 physicians who reviewed the decision aid found it acceptable, were comfortable providing it to patients and said that they were likely to use it. CONCLUSION: The decision aid is a useful and acceptable adjunct to personal counselling. PMID- 11281920 TI - The use of research-based theatre in a project related to metastatic breast cancer. AB - Research-based theatre represents an innovative approach to disseminating the results of qualitative studies. In this paper, we provide a rationale for the importance of research-based theatre and also review previous work that has been done in the area. We then describe our experience in transforming research data into a dramatic production, Handle with Care? This production was based on two studies - one with women with metastatic breast cancer, and the other with medical oncologists treating breast cancer patients. Results from ongoing assessment of the project are reported. We discuss some of the factors related to the success of Handle with Care? and reflect on what has been learned about the process of developing dramatic pieces related to serious illness. PMID- 11281921 TI - Does provision of an evidence-based information change public willingness to accept screening tests? AB - OBJECTIVE: To investigate whether the willingness of the general population to undergo a screening test of questionable effectiveness for pancreatic cancer is influenced by the quality and the extent of the information provided. DESIGN: Randomised study. SETTING: Switzerland. PARTICIPANTS: Representative sample (N=1000) of the general population aged over 20. INTERVENTIONS: Participants were randomly allocated into two groups (N=500 each), with one group to receive basic and the other extended quality of information. The information was presented in two hypothetical scenarios about implicit and explicit benefits and adverse events of the screening test. Response rates were, respectively, 80.2% (N=401) and 93.2% (N=466). MAIN OUTCOME MEASURES: Stated willingness to undergo the screening test. RESULTS: Out of the 401 participants receiving the basic information scenario, 241 (60%) stated their willingness to accept the test, as compared to the 63/466 (13.5%) exposed to the extended one (P < 0.001). After adjusting for respondent characteristics through a logistic regression model, the 'information effect', expressed in terms of odds-ratio (OR), shows that provision of additional information was related to a 91% (OR 0.09; 95CI: 0.07 - 0.13) relative reduction in the likelihood of accepting the screening test. CONCLUSION: The quality and the extent of the information provided about the implicit and explicit benefits and adverse events on hypothetical scenarios of a screening test may dramatically change the willingness of people to participate in the testing. This study suggests that provision of full information on the yield of health care interventions plays an important role in protecting the public from being exposed to procedures of questionable effectiveness. PMID- 11281923 TI - New UK health information websites. PMID- 11281922 TI - An experiment in patient information using the internet. PMID- 11281924 TI - Editorial: Promoting realistic expectations. PMID- 11281925 TI - Public perceptions about low back pain and its management: a gap between expectations and reality? AB - OBJECTIVE: To compare public perceptions and patient perceptions about back pain and its management with current clinical guidelines. DESIGN: A survey using a quota sampling technique. SETTING: On-the-street in South Derbyshire in the UK. SUBJECTS: 507 members of the general population aged between 20 and 60 years, including a representative subsample of 40% who had experienced back pain in the previous year. SURVEY: To test knowledge and perceptions of back pain and its best management using statements based on The Back Book which was produced in conjunction with the Royal College of General Practitioners and based on best available evidence. In addition expectations of back pain management and outcome were investigated. RESULTS: Forty percent of this sample had experienced back pain during the previous year, more than half of whom had consulted their GP. More than half believed the spine is one of the strongest part of the body, but nearly two thirds incorrectly believed that back pain is often due to a slipped disc or trapped nerve. Two thirds expected a GP to be able to tell them exactly what was wrong with their back, although slightly fewer among those who had consulted. Most expected to have an X-ray, especially if they had consulted. Most recognised that the most important thing a GP can do is offer reassurance and advice. The responses were not related to age, gender or social class. Those who had consulted appeared to have slightly more misconceptions: this could be partly due to people with more severe problems or more misconceptions being more likely to consult, but also suggests either that GPs are still giving inaccurate information or at least failing to correct these misconceptions. CONCLUSIONS: The problem of managing back pain might be reduced by closing the gap between the public's expectations and what is recommended in the guidelines through the promotion of appropriate health education messages. Further professional education of GPs also appears to be needed to update them in the most effective approach to managing back pain. PMID- 11281926 TI - Using focus groups to seek the views of patients dying from cancer about the care they receive. AB - PURPOSE: The prime purpose of the study was to investigate whether focus groups were a practical way of seeking the views of dying people and whether the information collected added to that collected by more established methods. DESIGN: A sample frame of Macmillan nurse patients was collated from which three cluster samples were randomly selected to participate in focus groups. The focus groups were structured with an experienced facilitator and recorder to answer three key questions. SETTING AND PARTICIPANTS: Patients of Macmillan nurses in three NHS Trusts in West Yorkshire who were living at home with incurable cancer. They were aware of their condition and were willing to participate. MAIN VARIABLES STUDIED: The three key questions were; what kind of help are you currently receiving? What sort of help do you want? Of the kind of help you are receiving what kind is most important to you? RESULTS: 17 patients participated in three focus groups. Participants were generally fairly able and living with spouses or relatives. They were of varying age with different types and duration of cancer. They were receiving a range of health and social services of varying importance to them. More help was particularly wanted with support for daily living, support from specialist cancer nurses, help getting out and with housework. Macmillan nurses and general practice services were highly rated in some but not all three trust areas. CONCLUSIONS: We propose that focus groups are a practical way of collecting information about dying patients that can complement other sources of information in planning and auditing the provision of care. PMID- 11281928 TI - Implementing shared decision-making in routine practice: barriers and opportunities. AB - OBJECTIVE: Determine feasibility of shared decision-making programmes in fee-for service hospital systems including physicians' offices and in-patient facilities. DESIGN: Survey and participant observation. Data obtained during Phase 1 of a patient outcome study. SETTINGS AND PARTICIPANTS: Three hospitals in Michigan: one 299-bed rural regional hospital, one 650-bed urban community hospital, one 459-bed urban and suburban teaching hospital. All nurses and physicians who agreed to use the programmes participated in the evaluation (n = 34). INTERVENTION: Two shared decision-making(R) (SDP) multimedia programmes: surgical treatment choice for breast cancer and ischaemic heart disease treatment choice. MAIN OUTCOME MEASURES: (1) clinicians' evaluations of programme quality; (2) challenges in hospital settings; and (3) patient referral rates. RESULTS: SDP programmes were judged to be clear, accurate and about the right length and amount of information. Programmes were judged to be informative and appropriate for patients to see before making a decision. Clinicians were neutral about patients' desire to participate in treatment decision-making. Referral volume to SDPs was lower than expected: 24 patients in 7 months across three hospitals. Implementation challenges centred on time pressures in patient care. CONCLUSIONS: Productivity and time pressure in US health care severely constrain shared decision-making programme implementation. Physician referral may not be a reliable mechanism for patient access. Possible innovations include: (1) incorporation into the informed consent process; (2) provider or payer negotiated requirement in the routine hospital procedure to use the SDP as a quality indicator; and (3) payer reimbursement to professional providers who make SDP programmes available to patients. PMID- 11281927 TI - The use of patients' stories by self-help groups: a survey of voluntary organizations in the UK on the register of the College of Health. AB - OBJECTIVE: First-hand accounts of illness experience are believed to provide important insights for other patients and their carers. We report the results of a survey that explored how patients' stories are being collected and used by self help and voluntary groups. METHODS: The annual College of Health survey contacts 2 458 addresses, which includes many self-help groups and voluntary associations. A brief questionnaire for the self-help groups on the register was attached to the summer 1999 survey on behalf of the DIPEx (database of individual patient experience) project. RESULTS: DIPEx received replies from 309 organizations representing a wide range of interests and conditions. These indicated that 202 (65%) of the groups currently use patients' stories in various ways. A further 59 (19%) of the groups reported that although they are not currently using them, they would like to in the future. Organizations that use patients' stories were invited to describe how they use them and provide examples, if applicable. Content analysis of the free text descriptions revealed 22 distinct uses among the 202 organizations using patient stories. The most frequent uses are the inclusion of patient stories in interviews or articles for the group newsletter (74 or 37%) and the use of stories for inclusion in newspaper articles or media broadcasts (31 or 15%). Some form of database of patients' stories was maintained by 23 groups (12%). CONCLUSIONS: These findings suggest that patients' stories are widely collected and used to support a wide range of the recognized functions of self-help and voluntary groups. This is encouraging to the DIPEx project's efforts to collect and analyse accounts of illness experience, which will be presented with evidence-based information about the effects of treatments. PMID- 11281929 TI - Life after hospital closure: users' views of living in residential 'resettlement' projects. A case study in consumer-led research. AB - OBJECTIVE: To conduct a user-led and focused study of the views and experiences of former psychiatric hospital patients in community-based residential projects four years after hospital closure. The aims of the study were to assess residents' views about their current living arrangements, their opportunities to give their views and their interest in a formal user-group such as a residents' council or citizen advocacy scheme. DESIGN: A small-scale, qualitative study designed to enable users to voice their own views and experiences in their own words, conducted by a project group of psychiatric service users/survivors. SETTING AND PARTICIPANTS: All eight residential 're-provision' projects in the area were included, with a total potential sample of 65 residents. All residents were invited to take part and a total of 26 were interviewed, although a larger number of residents together with residential care staff took part in initial 'house' meetings to discuss the study. METHODS: Semi-structured, open-ended interviews with all residents willing to participate, researcher participation in 'house meetings', researchers' personal reflection and discussion. RESULTS AND CONCLUSIONS: On the whole, residents were content with community living arrangements and preferred these to hospital, although levels of satisfaction varied across different residential projects. Residents lacked awareness of rights to and means of voicing concerns and making choices about major issues in their lives. They showed greater interest in individualized rather than group advocacy. Ideally, research and evaluation, to be truly user-focused, should be long-term and continuous in order to involve participants more fully, and should anticipate the structures and processes needed to act on findings. PMID- 11281930 TI - Measurement of consumer preference for treatments used to induce labour: a willingness-to-pay approach. AB - AIM: The purpose of the study was to assess the acceptability to consumers of two methods of induction of labour using a willingness-to-pay (WTP) approach. The methods compared were amniotomy plus oxytocin and prostaglandin E2 vaginal gel, followed by oxytocin if necessary. METHODS: A description of each method was presented, in questionnaire format, to pregnant women attending a public hospital ante-natal clinic. Women were asked to choose one of the two treatments, then give a valuation in dollar terms for both their preferred treatment and the alternative. RESULTS: It was found that 73.7% of patients preferred gel. The mean maximum WTP for amniotomy plus oxytocin was Aus$133 while that for gel was Aus$178 (P=0.0001). Those who chose amniotomy plus oxytocin were WTP 90% more for their preferred treatment compared with the alternative (Aus$180 vs. Aus$95). Similarly, those who preferred gel were WTP 90% more for their preferred treatment compared with the alternative (Aus$222 vs. Aus$119). CONCLUSION: Consumers were able to assess drug information provided on the two therapies, make an informed choice and to value that choice. Information obtained in this way, combined with information on costs, could be used in policy decision-making. PMID- 11281931 TI - The role of health professionals in informing cancer patients: findings from The Teamwork Project (phase one). AB - BACKGROUND: The Teamwork Project is managed by the National Cancer Alliance (NCA) and funded jointly by the National Lottery Charities Board and the Department of Health. The aim of the Project is to produce a Personal Information File to help people with cancer work in partnership with health professionals. Phase one was carried out between September 1998 and April 2000. The Teamwork Project arose as a direct result of the NCA report, 'Patient-Centred Cancer Services'? - What Patients Say,1 one of a number of studies that found people with cancer want to be involved in decisions about their treatment and care. The study also found that, for this involvement to be successful, health professionals need to support patients in accessing information relevant to their individual needs and help them understand and apply that information. The focus of The Teamwork Project is to help provide a practical solution to meeting this information need. APPROACH: The Teamwork Project has used a wide-range of methods including literature appraisal; patient questionnaires; focus groups; semi-structured interviews and a consultation exercise. Throughout the Project there has been on-going involvement from both patients and professionals. CONCLUSIONS: There may be a divergence of views among health professionals in cancer services regarding their role as providers of patient information. Consequently, there may also be a significant variance in how their patients are informed in practice. This finding needs to be validated and the reasons for this understood if the full potential of the forthcoming National Health Service (NHS) Cancer Information Strategy is to be realised. PMID- 11281932 TI - Editorial. PMID- 11281933 TI - A qualitative study of women's views about how health professionals communicate about infant feeding. AB - OBJECTIVE: To look at how communication by health professionals about infant feeding is perceived by first time mothers. DESIGN: Qualitative semi-structured interviews early in pregnancy and 6-10 weeks after birth. SUBJECTS AND SETTING: Twenty-one white, low income women expecting their first baby were interviewed mostly at home, often with their partner or a relative. RESULTS: The personal and practical aspects of infant feeding which were important to women were seldom discussed in detail in ante-natal interviews. In post-natal interviews women described how words alone encouraging them to breastfeed were insufficient. Apprenticeship style learning of practical skills was valued, particularly time patiently spent watching them feed their baby. Women preferred to be shown skills rather than be told how to do them. Some felt pressure to breastfeed and bottle feeding mothers on post-natal wards felt neglected in comparison. Women preferred their own decision-making to be facilitated rather than being advised what to do. Some women experienced distress exposing their breasts and being touched by health professionals. Continuity of care and forming a personal relationship with a health professional who could reassure them were key factors associated with satisfaction with infant feeding communication. CONCLUSIONS: The infant feeding goal for many women is a contented, thriving baby. In contrast, women perceive that the goal for health professionals is the continuation of breastfeeding. These differing goals can give rise to dissatisfaction with communication which is often seen as 'breastfeeding centred' rather than 'woman centred.' Words alone offering support for breastfeeding were often inadequate and women valued practical demonstrations and being shown how to feed their baby. Spending time with a caring midwife with whom the woman had developed a personal, continuing relationship was highly valued. Women were keen to maintain ownership, control and responsibility for their own decision-making about infant feeding. PMID- 11281934 TI - 'Do I don't I call the doctor': a qualitative study of parental perceptions of calling the GP out-of-hours. AB - The purpose of this study was to investigate how parents use the GP out-of-hours service. There was a lack of information about how parents managed childhood illness and what strategies they put in place to help them to cope before calling the GP. The investigation of parental perceptions was based on a qualitative design using in-depth interviews of 29 families from a semi-rural location in the south-east of England. All parents said they found dealing with a sick child out of-hours stressful and were concerned to make the right decision for their child. Furthermore, parents usually employed a reasonable strategy in attempting to manage the child's illness. This study demonstrated that the decision to call the doctor was not taken lightly. Many parents had implemented useful strategies prior to calling the doctor. However, most parents were also aware of their limitations and feared doing the wrong thing. It would seem that on occasion this fear combined with factors such as a lack of social support and loss of parental confidence resulted in calling the doctor out of hours to seek 'peace of mind'. A rethink is needed among health professionals about the 'problem' of out-of-hours calls. GPs could actively seek to empower parents by educating them about minor illness during visits and consultations. It is not enough to offer reassurance to parents that their children are fine. Health visitors and other health professionals who come into contact with young families may help to educate and empower. PMID- 11281935 TI - Challenges of participatory research: reflections on a study with breast cancer self-help groups. AB - OBJECTIVE: To review and discuss issues related to participatory research, as they apply within the arena of cancer control. DESIGN: A participatory research study with breast cancer self-help groups is referred to for description and discussion purposes. That study employed primarily individual and group interviews to assess benefits and limitations of self-help groups. SETTINGS: Four breast cancer self-help groups in Ontario communities provided the core involvement in the participatory research project. RESULTS: The values and practices of mainstream academic research often conflict with those of research emphasizing participation and control of communities under study, leading to a variety of challenges for the latter approaches. Practical constraints faced by many community groups have important implications for participatory research approaches. CONCLUSIONS: A balance needs to be found for participatory research within cancer control - one that ensures that the core aims of participatory research are maintained, while simultaneously acknowledging the various challenges that make a fully participatory project unrealistic. Steps can be taken to achieve a workable balance. PMID- 11281936 TI - Enhancing prevention in primary care: are interventions targeted towards consumers' and providers' perceived needs? AB - OBJECTIVE: To explore perceived barriers to the implementation of prevention guidelines, with a particular interest to perceived information needs from the point of view of health professionals and consumers. STUDY DESIGN: Focus group. SETTING AND PARTICIPANTS: Eight focus groups were held in three Canadian cities: three with consumer, three with family physician, and two with primary care nurses. ANALYSIS: Inductive analysis based on transcribed interviews. The material was analysed by two of the investigators. Agreement on interpretation was checked independently by three other researchers on 10% of the material. RESULTS: Lack of motivation, discontinuity of care and lack of adequate remuneration were perceived as the strongest barriers to prevention implementation. Computerized information management systems were not perceived by physicians and nurses as strong facilitating factors. Consumers expressed strongly a need for information on non-traditional preventive interventions. Physicians and nurses expressed a need for patient education material more than for practice guidelines. Research evidence was not considered as the first criteria to judge the value of preventive information. CONCLUSIONS: Evidence based medicine has triggered a massive effort to develop technologies to support the dissemination of evidence-based information on the assumption that poor access to such information is an important barrier to implementation of effective practices. Our results suggest that such an assumption may not be correct. Providing only evidence-based information from the realm of traditional medicine will appear restrictive to most users, particularly to consumers, and may not be as valued as anticipated considering the expressed scepticism toward research evidence. PMID- 11281937 TI - Reassurance through surveillance in the face of clinical uncertainty: the experience of women at risk of familial breast cancer. AB - OBJECTIVE: To identify the main issues raised by clinicians when they are counselling women at risk of breast cancer and explore the response of a group of women 1 year after counselling. DESIGN: A qualitative study which involved the thematic analysis of a series of transcripts from clinical consultations, semi structured interviews and focus groups. PARTICIPANTS: First, a series of clinical consultations (n=153), involving seven clinicians, were randomly selected during a Medical Research Council funded study of genetic assessment (TRACE). Second, a group of women (n=43), involved in the TRACE study, were interviewed, or joined a focus group, 1 year after their genetic assessment. CONCLUSIONS: There was evidence that, although the clinical consultations were embedded with multiple messages of uncertainty, the women's accounts did not reflect this. The women talked about the reassurance they had found because they had met with an expert and become members of the surveillance society. The authors highlight the tension that exists because of the difference between lay expectations about on-going surveillance and the realities of collective service provision. PMID- 11281939 TI - Time-course changes in rat cerebral cortex subcellular distribution of the cyclic AMP binding after treatment with selective serotonin reuptake inhibitors. AB - Pharmacological investigations have suggested the involvement of the cAMP transduction pathway in the action of antidepressant drugs and in the pathophysiology of mood disorders. We have extended these studies to determine the time-related effects of two selective serotonin reuptake inhibitors, fluvoxamine (15 mg/kg) and paroxetine (5 mg/kg), on the cAMP-binding in rat cerebral cortex, after short and long-term treatments. Photoaffinity labelling experiments with 8-N(3)-[(32)P]cAMP were carried out in cerebrocortical soluble (S1 or S2) and microtubule fractions. In our conditions, both SSRIs administered for 5 days were unable to affect the cAMP-binding in S1, S2, and in microtubule fractions. After 12 days of treatment, paroxetine and fluvoxamine significantly enhanced the cAMP-binding to the 54 kDa protein, corresponding to the type II regulatory subunit of PKA (RII), in the S1 and microtubule fractions. Any modification in respect to controls was observed in S2, the soluble fraction devoid of microtubules. After 21 days of treatment no changes were observed in the soluble S1 fraction and in microtubules, but the cAMP-binding to the RII subunit was found to be significantly higher in the S2 fraction. The high concentration of RII, demonstrated first in microtubules (12 days) and then in the cytosol (21 days), could be the result of a time-related effect of SSRIs on PKA and its translocation from microtubule compartment to the cytosol. The present findings seem to demonstrate the capacity of SSRIs to modulate the subcellular distribution of PKA and support the involvement of the cAMP pathway in the mechanism of action of these drugs. PMID- 11281940 TI - Effectiveness of ipsapirone, a 5-HT-1A partial agonist, in major depressive disorder: support for the role of 5-HT-1A receptors in the mechanism of action of serotonergic antidepressants. AB - Desensitisation of serotonin 1A (5-HT-1A) receptors is a leading hypothesis for the mechanism of action of antidepressants which block serotonin reuptake. This hypothesis predicts that direct-acting 5-HT-1A agonists should also exhibit anti depressant properties. Here we report the results of the first large-scale controlled study of the efficacy and tolerability of a 5-HT-1A agonist in outpatients with major depressive disorder (MDD). Three hundred and seventy-three subjects meeting DSM-III-R criteria for MDD participated in this randomised, double-blind comparison of the 5-HT-1A partial agonist ipsapirone (5 mg, 7.5 mg and 10 mg t.i.d.) to placebo t.i.d. Improvement in depressive symptoms relative to placebo, as measured by the Hamilton Depression Rating Scale, occurred in the ipsapirone (7.5 mg t.i.d.) group with a magnitude of effect (D=-2.53 points) that was statistically significant (p=0.010). Adverse events occurred in 76% of the placebo patients and 92% of the ipsapirone patients. A dose-related increase in the incidence of adverse events led to discontinuation of treatment with the 10 mg t.i.d. Results of this study demonstrate that ipsapirone, at a dose of 7.5 mg t.i.d., is an effective antidepressant agent in the treatment of MDD, supporting the hypothesised role of 5-HT-1A receptors in the mechanism of action of serotonin reuptake inhibitors. However, as a potential therapeutic agent for depression, ipsapirone shows only a modest magnitude of drug-placebo differences as well as a side-effect profile less favorable than many of the newer antidepressants. PMID- 11281941 TI - The effect of clozapine therapy on frontal lobe dysfunction in schizophrenia: neuropsychology and event-related potential measures. AB - There are several reports that performance-based measures as well as symptom ratings improve with clozapine in patients with schizophrenia who previously responded poorly to typical neuroleptic treatment. It is not clear whether improved cognitive function following initiation of clozapine is simply related to relief of psychotic symptoms and extrapyramidal side-effects associated with prior use of typical neuroleptics, or reflects another dimension of the greater efficacy of clozapine compared with typical neuroleptics. To elucidate this issue and better specify the cognitive changes associated with use of clozapine, the authors have assessed cognitive function psychometrically and using event-related potentials (ERPs), pre- and 8-12 wk post-initiation of clozapine treatment. Patients were rated on the BPRS, the SAPS and the SANS and completed a number of tests tapping aspects of frontal lobe function. ERP recordings were conducted using an auditory task twice, which was repeated under passive and active attention conditions. It was found that clozapine differentially affects tests reflecting executive and planning function, and not stimulus-driven cognitive functions. The results were not consistent with the hypothesis that these effects were simply due to relief of medication side-effects but could be related to the D(1) receptor antagonist actions of clozapine. PMID- 11281938 TI - NRG1 is required for glucose repression of the SUC2 and GAL genes of Saccharomyces cerevisiae. AB - BACKGROUND: Glucose repression of transcription in the yeast, Saccharomyces cerevisiae, has been shown to be controlled by several factors, including two repressors called Mig1 and Mig2. Past results suggest that other repressors may be involved in glucose repression. RESULTS: By a screen for factors that control transcription of the glucose-repressible SUC2 gene of S. cerevisiae, the NRG1 gene was identified. Analysis of an nrg1Delta mutant has demonstrated that mRNA levels are elevated at both the SUC2 and of the GAL genes of S. cerevisiae when cells are grown under normally glucose-repressing conditions. In addition, genetic interactions have been detected between nrg1Delta and other factors that control SUC2 transcription. CONCLUSIONS: The analysis of nrg1Delta demonstrates that Nrg1 plays a role in glucose repression of the SUC2 and GAL genes of S. cerevisiae. Thus, three repressors, Nrg1, Mig1, and Mig2, are involved as the downstream targets of the glucose signaling in S. cerevisiae. PMID- 11281942 TI - Chronic epi-inositol has an anxiolytic-like effect in the plus-maze model in rats. AB - Behavioural effects of myo-inositol have been demonstrated in animal models of psychiatric disorders and in clinical trials in psychiatric patients. These effects of myo-inositol were thought to be consequences of its being a substrate for the enzyme phosphatidylinositol (PI) synthase. Recently, epi-inositol, an unnatural stereoisomer of myo-inositol that is not a substrate for PI synthase, was found to also have effects similar to those of myoinositol to reverse lithium pilocarpine seizures. The present study measured the behaviour of rats in an elevated plus-maze model of anxiety after chronic treatment of 11 daily i.p. injections of epi-inositol, myo-inositol, or control solution. Epi-inositol reduced levels of anxiety-like behaviour in rats compared with controls, and its effect was stronger than that of myo-inositol. The increased relative effect of epi-inositol could be due to slower metabolism or to a different mechanism of action. PMID- 11281944 TI - An association between clozapine-induced agranulocytosis in schizophrenics and HLA-DQB1*0201. AB - A group of 18 Israeli, clozapine-treated, schizophrenia patients underwent molecular and serological HLA typing in order to determine whether the major histocompatibility complex is associated with the development of clozapine induced agranulocytosis. While under treatment with clozapine, 2 of the 18 patients developed agranulocytosis (total white blood cell count <3000/mm(3) and absolute polymorphonuclear count <500/mm(3)) and 3 developed granulocytopenia (total white blood cell count <3500/mm(3) and absolute polymorphonuclear count <1000/mm(3)). HLA-DQB1*0201 was present in all five patients who developed agranulocytosis or granulocytopenia (5/5; 100%), but in only 54% (7/13) of the patients who did not develop those complications. These findings indicate that DQB1*0201 or a gene located nearby could be involved in clozapine-induced agranulocytosis. PMID- 11281943 TI - The effect of nifedipine, Ca(2+) antagonist, on activity of MAO inhibitors, N acetylserotonin and melatonin in the mouse tail suspension test. AB - An antidepressive effect is associated with the A type monoamine oxidase (MAO) inhibitors which selectively stimulate serotonin conversion into N acetylserotonin and melatonin. The current study compared the effect of these compounds with the non-selective MAO inhibitors and selective MAO-B inhibitors on the duration of immobility in the mouse tail suspension test, a variant of the 'behavioural despair' test. Since the Ca(2+) antagonist, nifedipine, potentiated the effect of tricyclic and atypical antidepressants in the tail suspension test, the additional aim of the current study was to evaluate the effect of nifedipine in combination with ineffective doses of MAO inhibitors. Befloxatone, a selective reversible MAO-A inhibitor, N-acetylserotonin and melatonin decreased the duration of immobility. Non-selective MAO inhibitors (phenelzine and niamid), selective MAO-B inhibitors (deprenyl and Ro 196327) and selective MAO-A inhibitors (brofaromine, moclobemide and clorgyline) did not affect the duration of immobility. Nifedipine decreased the duration of immobility in combination with ineffective doses of all tested drugs, except the selective MAO-B inhibitor, Ro 196327. The results of our study suggest the antidepressant-like activity of N acetylserotonin and melatonin and the possibility of the potentiation of the effect of practically all known classes of antidepressants by Ca(2+) antagonist, nifedipine. PMID- 11281945 TI - Pain threshold is reduced in depression. AB - Pain and depression may share common neurochemical substrates, therefore the study of pain sensation in depression might be valuable in the investigation of the pathophysiology of depression itself. In order to investigate the sensation of pain in depression, we measured pain threshold and sensory threshold by means of a dental tester, comparing a group of depressed patients with healthy volunteers. The results showed the presence of a higher sensory threshold and pain threshold in patients than in controls. This may be related to a hyperfunction of the opiate system, which in turn might be primary or secondary to a decreased modulatory function of other neurotransmitters, in particular of serotonin, whose abnormalities in depressive states are well-documented. PMID- 11281946 TI - Evaluation of central serotonergic function in affective and related disorders by the fenfluramine challenge test: a critical review. AB - Plasma prolactin levels following oral administration of the serotonin (5-HT) releasing agent, fenfluramine hydrochloride, have been extensively used to evaluate central serotonergic function in affective and related disorders. Cortisol responses to fenfluramine have generally been a less informative measure. In healthy subjects, prolactin release by fenfluramine is dose dependent, blocked by antagonists of serotonin receptors of the 5-HT-2a/2c type, negatively correlated with age and increased in young females. In major depression, a preponderance of studies have found blunted prolactin responses compared to matched normal controls. Although a significant minority of studies have not found blunting, increased prolactin release has not been observed. The blunted prolactin release is not due to a deficient secretory capacity of pituitary lactotrophs and is congruent with other evidence for reduced central serotonergic function in major depression. Blunting of the prolactin response may be associated with severity of depression and with elevated baseline cortisol levels. Treatment with antidepressant drugs and electroconvulsive therapy has been reported to increase the prolactin response but this has not been replicated in all studies. Blunted prolactin responses to fenfluramine have been fairly consistently associated with impulsive aggression in different personality disorders and with severity of suicide attempts in depressed patients. A number of studies employing the fenfluramine challenge test (FCT) have been conducted in obsessive compulsive disorder but their results have been variable. Prolactin responses to fenfluramine may be enhanced in panic disorder and chronic fatigue syndrome but the number of studies in these conditions is small as is the case for seasonal affective disorder. Since the therapeutic administration of fenfluramine as an appetite suppressant has been suspended because of reports of cardiac complications, further use of this compound as a challenge agent is not anticipated. Future studies are likely to employ agents acting on specific serotonin receptors and should apply methodological insights from the use of the FCT, which are considered in this review. Use of concomitant brain imaging to evaluate the central effects of challenge agents directly is likely to become more prevalent and may supplant neuroendocrine challenge paradigms such as the FCT which have been remarkably heuristic but are limited in scope and methodologically complex. PMID- 11281947 TI - Health aspects of cannabis: revisited. AB - Literature pertaining to the effects of cannabis use and health which has been published during the past 11 years has been reviewed. Many older concerns about adverse effects on health (chromosomal damage, 'cannabinol psychosis', endocrine abnormalities, cardiac events, impaired immunity) no longer seem to elicit much interest. Continuing concerns about the adverse cognitive effects of chronic use indicate that these can be demonstrated by proper testing; some studies suggest that they may be long-lasting. Although cannabis does not produce a specific psychosis, the possibility exists that it may exacerbate schizophrenia in persons predisposed to that disorder. However, evidence from retrospective surveys must always be questioned. Tolerance and dependence have occurred in man, confirming previous findings in many other species. Addiction tends to be mild and is probably less severe than with other social drugs. Driving under the influence of cannabis is impaired acutely; how long such impairments last is still unknown. More exacting tasks, such as flying an airplane, may be impaired for as long as 24 hours. While there is no doubt that marijuana smoke contains carcinogens, an increase in cancer among users has thus far been anecdotal. Because of the long latent period between cancer induction and initiation of cigarette smoking, the full story is yet to be told. Marijuana use during pregnancy is not advised although the consequences are usually not greater than those of smoking cigarettes, and far less than those from alcohol use. Whether smoked marijuana should become a therapeutic agent requires a cost-benefit analysis of the potential benefits versus the adverse effects of such use as we now know them. PMID- 11281948 TI - Comment on 'Health aspects of cannabis: revisited' (Hollister). AB - Dr. Leo Hollister's excellent article begins to address the need for better understanding of the effects of cannabis use on health. The last five years in the US have seen an increase in advocacy groups extolling the medicinal utility of cannabis. On 5 November 1996, this culminated in California (proposition 215) joining the list of states permitting the limited use of cannabis for the medicinal treatment of disorders including intractable pain, glaucoma, nausea induced by chemotherapy for cancer or by AZT or Foscavir for the treatment of AIDS, and for spasticity associated with multiple sclerosis (Burstein, 1997; West and Homi, 1996; Grinspoon and Bakalar, 1995; Nahas and Manger, 1995). Of these potential uses for cannabis, the evidence for the treatment of nausea and the stimulation of appetite in cachetic patients appears most promising (for a review see Voth and Schwartz, 1997). Yet not only do doubts remain about the effectiveness of cannabis for the treatment of these conditions, since definitive controlled clinical studies are typically lacking (Voelker, 1997), but there is concern that any therapeutic advantage is more than offset by its harmful effects. Within this context of increased medical sanction for the use of cannabis in specific disease states for which it may have therapeutic potential, evaluating its risks vs. benefits profile is essential to rational prescribing. In addition, evaluating the public health risks associated with reports of increased risks of cannabis use (Robertson et al., Poulton et al., 1997), is of concern to advocates of its widespread legalization, governmental agencies attempting to limit its promulgation, and to planners and providers of health care charged with providing treatment for its consequences. PMID- 11281949 TI - Comment on 'Health aspects of cannabis: revisited' (Hollister). AB - Twelve years ago Leo Hollister (1986) published a review on health aspects of cannabis which can truly be called a 'classic'. In a field where emotions, prejudices and personal preferences frequently supplant objectivity, Hollister drew a picture not only solidly based on clinical studies - some of which came from his laboratory - but also on his thorough knowledge of drug dependence and medicine and on his common sense. For over a decade, at meetings, 'Hollister says' meant his 1986 review. He has now revisited the field, again demonstrating his mastery of the topic. In the 1986 review a portion was devoted to therapeutic uses. This aspect has not been revisited now, which we can only regret. I certainly would have liked to hear what he has to say on the debate on 'medical marijuana', on the clinical advantages of marijuana versus THC (or vice versa), and on specific issues such as the use of marijuana in multiple sclerosis, which is widely used by patients. Such a use is not supported in the US by the National Multiple Sclerosis Society, while in the UK, the British Medical Association strongly recommends 'carefully controlled trials of cannabinoids in patients with chronic spastic disorders which have not responded to other drugs'. Such trials merit a high priority (British Medical Association, 1997). PMID- 11281950 TI - Knockout mice in neuropsychopharmacology: present and future. AB - The technique of targeted inactivation of individual genes in mice has undoubtedly revolutionized biomedicine. Applications for gene knockout mice in neuropsychopharmacology are manifold: Determination of targets for established treatments is eased, while development of novel drugs is facilitated for a given target. This review discusses advantages and limitations of gene knockout strategies and, in particular, their relevance for the development of novel diagnostic and therapeutic approaches in psychiatric disorders. In addition to the classical targeted disruption of one or several genes simultanously, it specifically emphasizes those novel techniques that allow the inactivation of a gene with spatio-temporal selectivity. PMID- 11281951 TI - Hyperprolactinaemia induced by risperidone. AB - Risperidone is a potent antagonist of both dopamine (D2) and serotonin (5-HT2) receptors, demonstrating improvement of both positive and negative symptoms and a lower propensity for inducing extrapyramidal symptoms (EPS) than typical neuroleptics. Its most common side-effects, found in the Canadian multi-centre trial (Chouinard et al., 1993), were agitation, anxiety, insomnia, EPS, headache and nausea, in order of frequency. With regard to endocrine effects, risperidone causes an increase in prolactin levels similar to that of other neuroleptics (Claus et al., 1992). In open clinical trials (De Cuyper, 1991), the overall incidence of risperidone-induced endocrine side-effects was quite low: 2.9 % for amenorrhoea and 1-2% for galactorrhoea. However, it is assumed that the incidence can vary depending upon the characteristics of patients and the drug regimen, i.e. dosage and titration schedule. In our experience, hyperolactinaemia is likely to occur when prescribing risperidone to female or first-onset psychotic patients: we are reporting 5 cases of risperidone-induced hyperprolactinaemia with these characteristics. PMID- 11281952 TI - D(2)- and 5-HT(2) receptor occupancy in high-dose neuroleptic-treated patients. AB - Individual schizophrenic patients are sometimes reported to benefit from unusually high doses of neuroleptics. Such patients may have poor drug penetration into the brain or ultra-rapid metabolism. Alternately, very high doses may be required to induce occupancy of 5-HT(2) receptors, which have been suggested as mediators of atypical effects. Five schizophrenic patients treated with high doses of fluphenazine decanoate (100-250 mg/wk) and adjunct medications were examined with positron emission tomography and [(11)C]raclopride to measure D(2) receptor occupancy and [(11)C]NMSP to measure 5-HT(2) receptor occupancy. All patients were rated globally as 'markedly' to 'severely' ill and had high scores on all subscales of the Positive and Negative Syndrome Scale for schizophrenia. However, according to retrospective clinical evaluation, there was improved social function and reduced distress following high-dose treatment, an effect that deteriorated after previous explorative dose reduction. Extrapyramidal symptoms were modest. D(2) receptor occupancy was very high (89 97%). 5-HT(2) receptor occupancy was also high (76-105%). Plasma concentrations of fluphenazine were 5-37 nm. No patient had a cytochrome P450 CYP2D6 genotype associated with ultra-rapid drug metabolism. The findings suggest almost complete saturation of D(2) receptors, and do not support poor drug availability in the brain as the basis of the apparent high-dose requirement. The high 5-HT(2) receptor occupancy may have contributed to the apparent clinical improvement and modest degree of EPS. However, it is likely that the treatment used also induced occupancy of other neuroreceptors. PMID- 11281953 TI - Latent inhibition is disrupted by acute and repeated administration of corticosterone. AB - Latent inhibition (LI), namely, a retardation in conditioning to a stimulus, as a consequence of its prior non- reinforced pre-exposure, is disrupted in amphetamine-treated rats and humans and in some subsets of schizophrenic patients. One factor that has been repeatedly implicated in precipitating and/or exacerbating psychotic episodes is stress. Since a principal biological response to stress is the activation of the hypothalamic-pituitary-adrenocortical (HPA) axis, leading, as its end product, to the secretion of corticosterone, the present experiments tested whether increase in corticosterone levels following exogenous corticosterone administration would disrupt LI. Both repeated (Experiment 1) and acute (Experiment 2) administration of corticosterone led to LI disruption, providing evidence for the involvement of the HPA axis alterations in LI and further supporting the viability of disrupted LI as an animal model of psychosis. Both regimens also increased amphetamine-induced activity. We suggest that disrupted LI may reflect a cognitive mechanism whereby prolonged periods of increased corticosterone levels can lead to 'sensory flooding' characteristic of psychosis. PMID- 11281954 TI - Subsensitive melatonin suppression by dim white light: possible biological marker of panic disorder. AB - Light is involved in providing entrainment of circadian rhythms and the suppression of the pineal hormone melatonin. In patients with affective disorders, there have been indications of circadian as well as seasonal variation in illness, which may be reflected in melatonin production. Varying sensitivity to light has been noted within healthy individuals as well as in some patients with affective disorders. Recent evidence suggests that patients with panic disorder may have an altered and phase-delayed melatonin rhythm. The present study examined the nocturnal plasma melatonin rhythm in patients with panic disorder, and also examined their melatonin sensitivity to dim light. The melatonin rhythm was examined in 6 patients with panic disorder and 8 controls. The melatonin sensitivity to dim white light (200 lx) was examined in 8 patients with panic disorder and 63 controls and was compared to that of a group of 7 patients with other anxiety disorders. Patients with panic disorder demonstrated a trend towards higher and delayed peak melatonin levels compared to controls. Patients with panic disorder also had a subsensitive melatonin suppression by dim white light, compared to controls and patients with other anxiety disorders (p<0.005). The phase-delayed circadian rhythm observed in patients with panic disorder may be secondary to the subsensitivity of the melatonin response to light. It is hypothesized that the subsensitivity may be due to abnormal neurotransmitter/receptor systems involved in regulation of melatonin suppression and circadian rhythmicity, and may lead to phase- delayed circadian rhythms. The melatonin subsensitivity to light may be used as a biological marker of panic disorder. PMID- 11281955 TI - Gender difference in m-CPP challenge test in healthy volunteers. AB - Challenge test with oral m-CPP and placebo were performed in 12 healthy, young volunteers (6 male, 6 female) in a double-blind, placebo-controlled, cross-over study to assess possible gender differences in serotonergic responsivity. Women but not men showed significant prolactin and anxiety increase after administration of 0.25 mg/kg oral m-CPPP. Core temperature increased in both sexes compared to placebo. m-CPP induced significant physical symptoms only in men. Gender differences in serotonergic functions may have clinical significance with regard to differences in rates of some psychiatric disorders between the sexes. PMID- 11281956 TI - Dopamine receptor D3 gene and response to lithium prophylaxis in mood disorders. AB - Lithium has established itself as an effective prophylactic agent in mood disorders, but not all patients respond to lithium therapy. It is probable that genetic factors play a substantial role in determining the differences in response to lithium. The aim of this study was to investigate the association between the dopamine receptor D3 (DRD3) gene and prophylactic efficacy of lithium in mood disorders. Fifty-five subjects affected by bipolar (n=43) and major depressive (n=12) disorder were followed prospectively for an average of 49 months and were also typed for their DRD3 variant, using polymerase chain reaction techniques. DRD3 variants were not associated with lithium outcome. Consideration of possible stratification effects, such as gender, polarity, family history, age at onset or duration of lithium treatment, also did not reveal any associations. DRD3 variants are not, therefore, a major factor influencing the prophylactic efficacy of lithium in mood disorders. PMID- 11281957 TI - Double-blind, placebo-controlled, crossover trial of D-cycloserine adjuvant therapy for treatment-resistant schizophrenia. AB - Dysfunction of N-methyl-D-aspartate (NMDA) receptor-mediated glutamatergic neurotransmission may be relevant to the pathogenesis of negative symptoms in schizophrenia. The tuberculostatic compound D-cycloserine (DCS) acts as a partial agonist at the strychnine-insensitive glycine regulatory site on the NMDA receptor complex. Dose-finding trials suggest that DCS doses of 50-100 mg/d may be beneficial in the treatment of negative symptoms in schizophrenia. Nine treatment-resistant chronic schizophrenic patients participated in a double blind, placebo-controlled, adjuvant treatment trial with 50 mg/d DCS. Between treatment-groups differences in symptom changes were not significant. However, a significant (p<0.05) reduction in negative symptoms was registered during treatment with DCS but not placebo. Greater reductions were registered in patients with lower baseline serum glycine levels (p<0.008). No side effects were registered. These preliminary findings indicate a potential role of DCS in the treatment of negative symptoms in schizophrenia. However, the degree of symptom reduction may be modest, at least among treatment-resistant inpatients. PMID- 11281958 TI - An hypothesis of initial conditions: receptor-effector systems as determinants of psychopharmacologic drug response. AB - A variety of biological factors have been sought to explain why psychopharmacologic drug response varies between patients. In addition to variables such as age, race, gender, age of onset, severity, number of previous episodes, prior drug treatment, comorbidity and hormonal state, there have been investigations also into a variety of pharmacokinetic and pharmacodynamic variables such as dose, absorption, metabolism, distribution, globulin/protein binding, excretion and rate of titration, all potentially considered to explain differences in drug response. However, it is clear that, in many cases, such variables do not fully account for differences in drug response. In this work, it is hypothesized that drug response may also be significantly determined by the initial conditions of receptor site kinetics, as well as the sensitivity of downstream receptor-linked responses. This theory of initial conditions suggests that drug-action depends on receptor affinity and capacity, as well as on the conditions determining gene regulatory factors, which may influence transcription of gene products related to the emergent drug properties mediated through the receptor. Under one such scenario, dose response would be related, in the expected direction, to the initial affinity conditions of the receptor. For example, if the net effect of a drug were to decrease the apparent affinity of the receptor for its endogenous ligand, a stronger response could then be expected within a lower dose range, for those cases in which the initial affinity conditions for the receptor are already lower. The principle of initial conditions may apply not only to therapeutic psychotropic drugs, but also to drugs of abuse. This work reviews evidence to date in support of initial receptor conditions determining response to antipsychotic agents, response to antidepressants, and dependence as a response to alcohol, as conceptual examples. PMID- 11281960 TI - Comment - Genes and temperament, a shortcut for unravelling the genetics of psychopathology? AB - Personality is complex and unique, in that people differ greatly from one another in multiple components of personality. Nevertheless, personality can be measured and dissected into a number of quantitative traits. One of the more recent tools is the Temperament and Character Inventory (TCI), which is a self-report personality questionnaire based on Cloninger's psychobiological model of personality (Cloninger et al., 1994). This model consists of four dimensions of temperament and three dimensions of character. Temperament dimensions (novelty seeking, harm avoidance, reward dependence and persistence) refer to automatic emotional responses to stimuli and are moderately heritable, relatively stable throughout life, and invariant despite sociocultural influences (Svrakic et al., 1996). Individuals high in novelty-seeking tend to be quick-tempered, excitable, exploratory, curious, enthusiastic, impulsive and disorderly. Individuals high in harm avoidance tend to be cautious, careful, fearful, tense, apprehensive, nervous, discouraged, insecure, negativistic and pessimistic, even in situations which do not worry other people. People scoring high on reward dependence seek social contact and are open to communication with other people; they are tender hearted, sensitive, dedicated and sociable. Highly persistent individuals tend to be industrious, hard-working, persistent, and stable, despite frustration and fatigue. The character dimensions (self- directedness, cooperativeness and self transcendence) involve differences in social goals and values and seem to be moderately influenced by family environment (Cloninger et al., 1994). Cloninger's psychobiological model of personality shows correlations to other factor models of personality, like the NEO five-factor model. The TCI dimension of novelty seeking shows positive loading on NEO conscientiousness (Cloninger et al., 1994). PMID- 11281959 TI - Genes for personality traits: implications for psychopathology. AB - Although 30-60% of the variance in many personality traits is inherited, until recently, little was known about the genes responsible. Preliminary studies of family history in bipolar disorder and of X-linkage of personality traits in colour-blindness suggested a 'quantitative trait locus' (QTL) approach to the genetics of normal personality. In methodically similar but independent studies of 124 Israeli and 315 American normal volunteers, an association was found between the dopamine D4 receptor gene (D4DR) and the personality trait of novelty seeking. In the Israeli sample there was preliminary evidence for an interaction between the D4DR gene and the serotonin 2C receptor gene (5-HT-2C), with a marked effect on the trait of reward dependence. In addition to receptors, monoamine uptake mechanisms, such as the serotonin transporter (5-HTT), are candidate genes for personality traits. 5-HTT gene transcription is modulated by a frequent polymorphism in its promoter region, with resulting effects on 5-HTT expression and 5-HT uptake. In an extended American sample totalling 505 subjects, the 5-HTT polymorphism was associated with anxiety- and depression-related personality traits. The allelic variation in functional expression of the 5-HTT may also be a susceptibility factor for disorders of the affective spectrum. Further investigation of genes for personality traits may provide additional links between normal personality and psychiatric illness. PMID- 11281961 TI - Polymorphism of CYP2D6 in Black populations: implications for psychopharmacology. AB - Drug-metabolizing enzymes found primarily in the liver (CYP450) are a major determinant of therapeutic drug response. Polymorphism dependent upon race/ethnic origin for CYP2D6 is now well-established. Despite consistent reports of ethnic differences in pharmacologic response to antidepressants and neuroleptics, there is a paucity of data on controlled clinical trials and studies determining polymorphic characteristics of CYP2D6 enzymes in African-Americans. There is little and conflicting information available on black populations (Africans, bushmen, Australian Aborigines or African Americans). The prevalence of poor metabolizers in Black populations has been estimated from 0 to 19%, compared with consistent reports of poor metabolizer status in Caucasians (5-10%) and Asians (0 2%). Within the extensive metabolizer category, Asians have higher metabolic ratios (that is, slower metabolism) than Caucasian extensive metabolizers. A high frequency of a mutant gene, CYP2D6*10 has been associated with the slower metabolic rate in Asians. Previous research suggests that slower metabolic rates compared with Caucasians may also be characteristic of Black populations. Recent reports suggest that a novel gene mutant in Black populations, CYP2D6*17, associated with a slower metabolic rate, may occur in a high frequency in these populations. Common clinical practice, supported by controlled clinical studies in Asians, have led to a reduction in dosage recommendations for many antidepressants and neuroleptics for this ethnic group. It is imperative that the determinants of bioavailability be established in African-Americans in order to establish rational drug therapy guidelines for this population. PMID- 11281962 TI - The adenosine A(2A) receptor knockout mouse: a model for anxiety? AB - The main behavioural features of the adenosine A(2A) receptor knockout mouse include anxiety, aggressiveness in males and a paradoxical response to caffeine. These behavioural characteristics caused by the lack of adenosine A(2A) receptor function in mice correspond to the effects of functional antagonism of adenosine A(2A) receptors in humans and rodents. Increased anxiety in patients with panic disorder and increased psychotic symptomatology in patients with schizophrenia have been observed after caffeine administration. Several hypotheses have been developed suggesting a reduced adenosine A(2A) receptor-mediated transmission as a contributing factor in the pathogenesis of these disorders. Recent genetic studies, in particular of panic disorder, suggest an involvement of adenosine A(2A) receptor gene variation. If future studies prove a pathogenetic role for a genetically determined loss of A(2A) receptor function in psychiatric disorders, the A(2A) receptor knockout mouse will be a valuable model to develop novel pharmacological therapies for these disorders. PMID- 11281963 TI - Pioneers in Psychopharmacology. AB - In pursuing the history of any field, even one in which many of the main exponents are still alive, it can be very difficult to establish facts and priorities. Detailed scrutiny of the events leading to the recognition of the antidepressant effects of iproniazid, in which Nathan Kline was involved, may fail to establish the exact sequence of events or the sources of inspiration for a discovery (Healy, 1997). Quite apart from the 'facts' behind the antidepressant story, Kline's role in the story beautifully illustrates one of the sayings of Francis Galton, to the effect that in history the driving force may not lie with the first discoverer of a new scientific fact, but rather with the individual who was the first to persuade the world of the importance of a particular discovery. This maxim applies with some force to the three individuals who have been honoured by the CINP in 1998 for their pioneering roles in psychopharmacology - Pierre Deniker, Joel Elkes and Heinz Lehmann whose contributions to the field have included original research and also key initiatives to capture the central ground of academic and public opinion for the new science. They have been science makers rather than just scientists. PMID- 11281964 TI - Pharmacological actions of the antidepressant venlafaxine beyond aminergic receptors. AB - The present study examines the effects of the antidepressant venlafaxine, a dual amine reuptake inhibitor, on (a) in vivo regulation of the densities of high- and low-affinity dihydroalprenolol (DHA) binding sites in the cortex of normal and reserpinized Sprague-Dawley rats and (b) targets beyond the beta adrenoceptor. While venlafaxine (30 mg/kg i.p. b.i.d.) administered for 4 d did not alter the DHA-binding parameters in the cortex of normal rats, it significantly reduced, in reserpinized animals, the number of up-regulated low-affinity sites (R(L)) which have been tentatively identified as serotonin(1B) sites. The drug did not influence the up-regulated high-affinity (R(H)) DHA-binding sites (beta adrenoceptor sites). Venlafaxine failed to alter the up-regulated R(L) sites in brains of rats depleted of serotonin (5-HT) by p-chlorophenylalanine (PCPA) indicating that the normalization by venlafaxine of the up-regulated R(L) receptor population is mediated by increased synaptic 5-HT. Venlafaxine, given for a short period of time, thus mimicked the action of fluoxetine. While venlafaxine (20 mg/kg i.p. b.i.d.) given for 10 d did not change protein kinase A activity as assessed by the phosphorylation of kemptide in the 900 g supernatant or particulate fractions, the drug significantly reduced phosphorylated cAMP response-element binding protein (CREB-P) in nuclear lysates of cortex after chronic but not acute administration. Depletion of 5-HT by PCPA did not alter the venlafaxine-induced change in nuclear CREB-P. Lastly, analysis of reverse transcribed cortical CREB mRNA by competitive PCR indicated that the mean steady state levels of CREB mRNA in venlafaxine vs. saline-treated animals were not significantly different. Therefore, since the phosphorylation status of CREB determines its transcriptional activity the reduction of nuclear CREB-P may be venlafaxine's most relevant action beyond the adrenoceptor. PMID- 11281965 TI - Role of CCK in anti-exploratory action of paroxetine, 5-HT reuptake inhibitor. AB - The administration of paroxetine (0.5-8 mg/kg), a selective 5-HT reuptake inhibitor, induced a dose-dependent reduction of exploratory activity of rats in the motility test. In the elevated plus-maze paroxetine was less effective, only 8 mg/kg of paroxetine decreased the exploratory behaviour of rats. The doses of paroxetine (2-8 mg/kg) reducing the exploratory activity in the motility test increased the density of CCK receptors in the frontal cortex, but not in the hippocampus. The treatment of rats with the CCK(B) receptor antagonist LY288,513 (0.01-1 mg/kg) did not change the exploratory activity. However, the reduction of exploratory activity induced by the low dose of paroxetine (2 mg/kg), but not by the higher dose (8 mg/kg), was dose-dependently reversed by the administration of LY288,513. Moreover, LY288,513 did not affect the anti-exploratory action of paroxetine (8 mg/kg) in the elevated plus-maze. Diazepam at doses (0.5-1.0 mg/kg) not suppressing the locomotor activity did not change the anti-exploratory action of paroxetine in the motility test. It is likely that the anti-exploratory action of a low dose of paroxetine (2 mg/kg) is not related to the increase in anxiety, but rather to the reduction of exploratory drive. Evidence exists that this effect of paroxetine is mediated via the activation of CCK-ergic transmission. PMID- 11281966 TI - Pharmacokinetics of trazodone and its major metabolite m-chlorophenylpiperazine in plasma and brain of rats. AB - Sprague-Dawley rats were used as models for single trazodone administration (males), continuous adminstration and dose proportionality experiments (males, females, pregnant females). Plasma and brain tissue were analysed for trazodone and its active metabolite, m-chlorophenylpiperazine (m-CPP). Fetal exposure to trazodone and m-CPP was assessed and differences in their steady-state plasma concentration were sought between adult males and females. Both trazodone and m CPP rapidly appeared in plasma and brain tissue following a single intraperitoneal trazodone dose with brain concentrations exceeding those in plasma. Plasma concentrations of m-CPP were lower than those of trazodone but exceeded them in brain tissue. Chronic administration using osmotic mini-pumps revealed a significant linear relationship between trazodone concentration in plasma and brain at steady-state (r=0.96, p<0.0001). No simple relationship was found between plasma and brain tissue concentration for m-CPP. In contrast to observations following single trazodone administration, m-CPP concentrations at steady-state were lower than trazodone concentrations in brain tissue, suggesting a lack of stationarity in the disposition of trazodone over time. No significant differences in plasma or brain tissue drug concentrations relative to administered trazodone dose were observed between male and female rats, nor between pregnant and non-pregnant females. Trazodone and m-CPP were both detected in fetal and placental tissues, with placenta having the highest concentrations. The data suggest that neuropharmacological studies of trazodone could yield different results depending upon the route and schedule of drug administration. Maternally administered trazodone, like many other antidepressants, is distributed to fetal tissues in rodents, reaffirming the need for caution in treating pregnant women with psychoactive drugs. PMID- 11281967 TI - Effect of bipolar disorder on lymphocyte inositol monophosphatase mRNA levels. AB - The activity of inositol monophosphatase (IMPase), the lithium (Li)-inhibitable enzyme in the phosphatidylinositol (PI) signal transduction system, has recently been found significantly lower in lymphoblastoid cell lines from bipolar (BP) patients, particularly in Li-responders. To probe for possible quick detection of the disease and prediction of the therapeutic response we repeated our study in fresh lymphocytes. Since IMPase in fresh lymphocytes is inhibited in vivo by ongoing Li treatment and its pre-Li activity cannot be evaluated, IMPase mRNA levels were measured. Relative (to beta-actin) mRNA levels were quantified by reverse transcriptase (RT)-PCR in 5 drug-free and 31 drug-treated BP patients compared with 36 control subjects in fresh lymphocytes. In agreement with our findings with IMPase activity, the small group of drug-free BP patients exhibited approximately 2/3 reduction in IMPase relative mRNA levels compared to control subjects. Approximately 2-fold elevation of these levels toward control values was found for patients treated with Li and other mood stabilizers. The study further suggests the possible importance of IMPase in the aetiology of BP disorder and in the mediation of the therapeutic efficacy of Li. It may be that chronic inhibition of IMPase activity by Li results in up-regulation of its gene at the transcriptional level. PMID- 11281968 TI - The influence of rapid-rate transcranial magnetic stimulation (rTMS) parameters on rTMS effects in Porsolt's forced swimming test. AB - To assess the similarity of the behavioural effects of the rapid transcranial magnetic stimulation (rTMS) to those produced by other antidepressant treatments, in particular to repeated electroconvulsive shock (ECS), we carried out experiments on Wistar rats. The effects of a standard ECS procedure (9 daily treatments; the current parameters: 150 mA, 50 Hz, 0.5 s) were compared with 18 d treatment with rTMS of the same field intensity of 1.6 T but with different stimulation frequency (20 or 30 Hz) and a different number of sessions (9 or 18). Twenty-four hours after the last treatment the forced swimming test was carried out and the immobility time was recorded. The standard ECT reduced the immobility by 50%, the intensive rTMS (90 or 104 K impulses for the whole period of treatment) caused a significant effect, although smaller than that induced by ECT (reduction by 20-30%). The stimulation at 20 Hz required 18 treatment sessions to produce a significant effect, while only 9 sessions with stimulation at 30 Hz were sufficient to produce a comparable result. This suggests that the effectiveness of rTMS may be augmented by increasing the number or frequency of rTMS impulses. PMID- 11281969 TI - High dose glycine nutrition affects glial cell morphology in rat hippocampus and cerebellum. AB - Enhancement of N-methyl-D-aspartate (NMDA) receptor-mediated neurotransmission by glycine (Gly) administration may represent a novel pharmacological strategy in schizophrenia. Given the involvement of NMDA receptors in plasticity and excitatory processes, the present study explores effects of Gly on brain cell morphology. Adult rats were randomized to receive, for 2 wk, no dietary supplementation or supplementation with 0.8 or 3.2 g/kg per day Gly. Glial cell morphology was examined using antibodies to glial fibrillary acidic protein (GFAP) and to microglial complement receptor 3 (CR3). Cresyl violet was used for general cellular staining. No evidence of neuronal or microglial pathology was found. Although astrocyte proliferation was not evident in Gly-treated rats, GFAP like immunoreactivity was dose-dependently increased in the hippocampus (p<0.01), whereas in cerebellum, a dose dependent decrease in density of astocytic fibres was demonstrated (p<0.01). These findings demonstrate for the first time that in vivo administration of high dose Gly may induce brain morphology changes. PMID- 11281970 TI - Brain dopamine D(4) receptors: basic and clinical status. AB - Since their discovery in 1991, an extraordinary amount of information has accumulated about the neurobiology and pharmacology of D(4) receptors in the mammalian central nervous system, making it timely to review salient aspects of this rapidly evolving research story and its relevance to clinical neuroscience. Recent progress in the molecular, genetic, anatomical, and functional characterization of D(4) receptors in the animal and human brain is yielding insights into their neurochemical and physiological roles in brain function. The temporal patterns of postnatal D(4) receptor development appear to differ in specific regions of mammalian forebrain. Postmortem neuropathological and clinical genetic studies have generally been disappointing regarding possible relationships of D(4) receptors to the pathophysiology or treatment of schizophrenia, however, they suggest relationships to other neuropsychiatric conditions, including attention deficit hyperactivity disorder, mood disorders, and Parkinson's disease. Several selective agonists and antagonists for D(4) receptors have been developed. Some are employed as experimental D(4) probes. The potential of D(4)-selective ligands as innovative treatments for neuropsychiatric disorders requires further investigation. PMID- 11281971 TI - Strategies for analysing behavioural data in clinical trials involving patients with Alzheimer's disease. AB - Behavioural disturbances are common in Alzheimer's disease and may be affected by medications being developed to enhance cognition or slow disease progression as well as by psychotropic agents developed specifically to affect behaviour. In many cases, Alzheimer's disease patients included in clinical trials are not selected for behavioural attributes and the patient population is heterogeneous at baseline with regard to these symptoms. Analyses of the behavioural data should include assessment of the effects of the agent on patients who were symptomatic at baseline as well as on the incidence of new behaviours in those without symptoms at the time of study initiation. Analyses may focus on symptomatic patients exhibiting specific degrees of improvement (e.g. 50% reduction in symptom severity). The analytic strategy chosen to characterize the behavioural changes occurring in clinical trials involving Alzheimer's disease patients will depend on the hypotheses being explored, the characteristics of the patients at baseline, the size of the population studied, the assessment methodology, and the outcomes of interest. PMID- 11281972 TI - Knockout Corner: Neurobehavioural consequences of a serotonin 5-HT(2C) receptor gene mutation. AB - Studies employing nonselective serotonin 5-HT(2C) receptor agonists and antagonists have implicated this receptor subtype in many of the actions of serotonin. To further examine the function of this receptor, 5-HT(2C) receptor mutant mice were generated; studies of these animals reveal pleiotropic neurobehavioural effects of the mutation. Three examples are described: (1) Mutants exhibit chronically elevated food intake and the development of an obesity syndrome during the 'middle-age' portion of their lifespan. Their potential utility as a model of human obesity is further indicated by their enhanced sensitivity to high-fat feeding, leading to the development of type 2 diabetes. (2) 5-HT(2C) receptor mutants also display infrequent and sporadic spontaneous seizures. Further studies suggested the presence of globally enhanced neuronal network excitability in these mice. These findings raise the possibility that 5-HT(2C) receptors mediate a role for serotonin in the suppression of seizure activity. (3) Behavioural analysis of mutant mice revealed abnormal performance in a spatial learning task and altered exploratory behaviour, associated with perturbed long-term potentiation restricted to the dentate gyrus perforant path synapse. Taken together, the above findings implicate 5-HT(2C) receptors in the serotonergic regulation of feeding, neuronal network excitability, and hippocampal function. PMID- 11281973 TI - Extrastriatal dopamine D2 receptor density and affinity in the human brain measured by 3D PET. AB - The aim of the present study was to quantify the density and affinity of human extrastriatal dopamine D2 receptors using positron emission tomography (PET). [(11)C]FLB-457, a high-affinity dopamine D2 receptor antagonist with various specific radioactivities (SA) was used. Eight healthy male subjects, age 20-35 yr, participated twice or three times at different SAs (1-279 GBq/ umol), and serial dynamic scans were performed in the 3D data acquisition mode. The peak of the specific binding was not well defined with high SA due to the flatness of the curves after 60 min but was observed within the PET measurement. In the experiment with low SA, the peak came earlier than that with high SA. Scatchard analysis was performed using the maximal specific binding value (transient equilibrium) and the radioactivity in the cerebellum as free ligand concentration. The highest density was observed in the thalamus (2.3+/-0.6 pmol/ml), followed by the temporal cortex (1.5+/-0.5 pmol/ml), hippocampus (1.4+/ 0.5 pmol/ml), parietal cortex (0.9+/-0.4 pmol/ml), frontal cortex (0.8+/-0.2 pmol/ml) and occipital cortex (0.7+/-0.3 pmol/ml). There was no significant difference in K(d) values in these six regions. The present results demonstrate that dopamine D2 receptor densities in the extrastriatal regions were only 2-8% of that in the striatum. Although the density of extrastriatal dopamine D2 receptor was low, significant regional differences were observed in the present study, as reported in postmortem studies. PMID- 11281974 TI - Auditory event-related potential indices of fronto-temporal information processing in schizophrenia syndromes: valid outcome prediction of clozapine therapy in a three-year follow-up. AB - Reliability, specificity, and validity of three event-related potential (ERP) indices of non-attended and attended auditory information processing [the mismatch negativity (MMN), the novelty-P3a and the target-P3b] were assessed in 21 healthy subjects and 25 schizophrenic patients while performing a visual discrimination task (Ignore condition) and, subsequently, an auditory discrimination task (Attend condition). Re-test reliability, as measured in a subset of 10 healthy subjects and 9 patients respectively, ranged from r=0.4 to r=0.8. In both groups P3a was found to be smaller in the Ignore than in the Attend condition. Patients had significantly larger P3a amplitudes than healthy subjects while their MMN and P3b amplitudes were smaller. Two ERP factors were extracted in healthy subjects: (1) 'novelty reaction' with high loading scores for P3a and (2) 'deviant detection' with high loading scores for MMN. P3b was predominantly loading on the first factor thus confirming partly overlapping P3a and P3b generators. However, considerable variance was also shared by the second factor, particularly in patients. External validity of the ERP factors was confirmed post hoc. Particularly ERP indices of deviant detection were found to be associated with negative symptoms. In order to assess further clinical relevance, therapy response to clozapine was followed-up over 3 yr in a sub sample of 17 patients. Poor treatment response was associated with high Novelty Factor scores and large P3a amplitudes whereas good response was associated with high Deviant Factor scores and relatively intact MMN. It is concluded that ERP abnormalities provide a severity index of brain impairment and a measure of predicting therapeutic outcome. PMID- 11281975 TI - Differential effects of fluoxetine and citalopram treatments on serotonin 5 HT(2C) receptor occupancy in rat brain. AB - Ex vivo receptor occupancy measurements were performed in order to study the effects of the serotonin reuptake inhibitors fluoxetine and citalopram on serotonin 5-HT(2C) receptors. To determine the degree of 5-HT(2C) receptor occupancy, [(3)H]mesulergine binding in brain sections containing rat choroid plexus was measured at various time-points after drug injection. For comparison, [(3)H]ketanserin binding to frontal cortex 5-HT(2A) receptors was measured. Fluoxetine treatments (10 and 20 mg/kg) resulted in 5-HT(2C) receptor occupancy of up to 25 and 43%, respectively. Fluoxetine (20 mg/kg) caused a persistent effect: at the 24 h time-point, 23% of 5-HT(2C) receptors were still occupied. Citalopram treatment did not result in marked 5-HT(2C) receptor occupancy. Neither drug caused significant 5-HT(2A) receptor occupancy. In conclusion, the results demonstrate pharmacodynamic differences between fluoxetine and citalopram at the level of 5-HT(2C) receptors. These findings provide evidence that direct occupancy of 5-HT(2C) receptors may contribute to the mechanism of action of fluoxetine. PMID- 11281976 TI - Enhancement of serotonin(1A) receptor function following repeated electroconvulsive shock in young rat hippocampal neurons in vitro. AB - Effects of repeated electroconvulsive shock (ECS) treatment on 5 hydroxytryptamine (5-HT) response were investigated to elucidate the ECS-induced changes, which may be related to antidepressant effects, using electrophysiological methods with hippocampal slices in vitro. ECS was applied to Wistar rats once daily for 14 d from 3 wk of age (ECS group). Control animals did not receive ECS (control group). Twenty-four hours after the final ECS treatment, hippocampal slices were prepared for intracellular recording analysis. Application of 5-HT (0.1-30 um) caused a dose-dependent hyperpolarization in hippocampal CA1 neurons. 5-HT-induced hyperpolarization in the ECS group was significantly greater than that in the control group. Furthermore, 8-OH-DPAT [8 hydroxy-2-(di-n-propylamino)tetralin], a 5-HT(1A) receptor agonist, also induced significantly larger hyperpolarization in the ECS group than in the control group. These results suggest that repeated ECS treatment enhances function of the 5-HT(1A) receptor for 5-HT. This supports the hypothesis that enhanced 5-HT(1A) receptor function, at least in part, contributes to the effectiveness of ECS treatment for depression directly and/or indirectly. PMID- 11281977 TI - Serum thyroid-stimulating-hormone concentration as an index of severity of major depression. AB - Alterations in thyroid axis are common in depression and subclinical hypothyroidism may predispose to recurrent depressive episodes and resistance to antidepressants. The same normal reference ranges are used in both depressive and non-psychiatric patients to detect hypothyroidism. We hypothesized that in depressive patients, serum TSH (thyrotropin) elevation within the normal reference range (>/= upper 25th percentile) may be related to patients' characteristics reflecting the severity of the depressive illness. We analysed, in a cross-sectional study, the relationship between serum TSH and serum-free thyroxine (T4) concentrations and different demographic and psychiatric characteristics in 94 depressive in-patients with DSM-III-R criteria for major depression. The frequency of subclinical hypothyroidism (normal serum T4, higher than normal serum TSH) was 5.3 %. In univariate analyses patients who had serum TSH concentrations >/= upper 25th percentile of the normal range were more likely to have recurrent depression, longer disease duration, higher number of episodes of major depression, higher number of previous suicide attempts and higher body mass index than those patients who had serum TSH concentrations < upper 25th percentile of the normal range (age-adjusted p<0.05). Stepwise logistic regression analysis showed that serum TSH >/= upper 25th percentile of the normal range was positively associated with recurrent depression (p=0.0001), presence of somatic disease condition (p=0.04), marital status (p=0.06) and number of suicide attempt (p=0.1). On the other hand, significantly higher serum TSH concentrations were observed in patients with recurrent depression, melancholia and associated somatic disease conditions. Correspondence analysis showed that serum TSH in the higher 25th percentile of the normal reference range projected together with the presence of melancholia, psychiatric and somatic disease conditions, severe major depressive episodes, recurrence of depressive episodes, prescription of at least two antidepressants or non-response to two antidepressants, and previous suicide attempts. Our study suggests that serum TSH concentration in the upper 25th percentile of the normal reference range may be associated with characteristics of severe major depression. Further prospective studies are needed to establish whether serum TSH concentration in the upper 25th percentile of the normal reference range is a contributory causal factor or a consequence of the severity of major depression. PMID- 11281978 TI - Number of episodes and antidepressant response in major depression. AB - Current series of depression suggests that episodes of major depression sensitize a patient to further episodes so that the illness adopts a recurrent course. This suggested pathophysiological process may also lead to increased risk of treatment resistance and a chronic course of illness. This hypothesis has received little empirical support and, if correct, would suggest that greater number of episodes would lead to a decreased response to antidepressants. We examined this in a cohort of outpatients with major depressive disorder. We observed that initial severity of depression and duration of treatment, but not number of previous episodes, was related to treatment outcome. Our findings are discussed in relation to prevailing theories of the pathophysiology of depression and suggestions for further studies are made. PMID- 11281979 TI - Serotonin transporter promoter polymorphism influences topography of inhibitory motor control. AB - The prefrontal cortex participates in motor control and is modulated by serotonergic activity. The serotonin transporter (5-HTT) is a major regulator of serotonergic neurotransmission and may thus influence motor control. The short allele (s) of the 5-HTT linked polymorphic region (5-HTTLPR) is associated with less 5-HTT expression and function than the long variant (l). The neurophysiological parameters termed 'Go- and NoGo- centroid location' represent characteristic brain electrical substrates of the execution and inhibition of motor response elicited by the Continuous Performance Test (CPT). In the present study, the impact of the 5-HTTLPR genotype on the centroid locations was investigated in 23 healthy subjects. The NoGo-centroid, but not the Go-centroid, was located significantly more anteriorly in the short allele group (mean electrode location in s/s and s/l, 2.86+/-0.37) compared to the group with two long alleles (l/l, 3.34+/-0.49; t=2.66, p<0.05). Age, gender, and test performance did not differ between groups. The results indicate that 5-HTTLPR genotype dependent 5-HTT function is associated with the neurophysiologically assessed topography of inhibitory motor control and provides further evidence for a genetic influence on central serotonergic and motor function. PMID- 11281980 TI - Serotonin-selective reuptake inhibitors in the treatment of geriatric depression and related disorders. AB - Depression is a common disorder in late life which can be successfully treated with antidepressant agents. Other disorders such as behavioural agitation associated with dementia may also be treated with antidepressants. In this review, we have examined the use of five serotonin-selective reuptake inhibitors in the treatment of late-life depression and related disorders in the elderly population. Medications examined include citalopram, fluoxetine, fluvoxamine, sertraline and paroxetine. Comparisons of these agents' efficacy in the elderly are made. In addition, comparisons of these agents' side effects, pharmacokinetics and potential for drug-drug interactions are also discussed. PMID- 11281981 TI - Proposed research diagnostic criteria for neuroleptic malignant syndrome. AB - Many sets of diagnostic criteria have been proposed for neuroleptic malignant syndrome (NMS) but there is a lack of uniformity. No universally agreed criteria exist currently for research purposes, thus making comparisons across studies very difficult. Most of them have flaws and detect too many false-positives based on an over-inclusive definition. The estimates of incidence rates of NMS vary because of differences in the sensitivity threshold of the diagnostic criteria used. A new set of diagnostic criteria is proposed for research purposes. It is hoped that with this set of stringent research diagnostic criteria, future epidemiological, aetiological and treatment research studies on NMS will be more meaningful and comparable across studies. For routine clinical purposes, the clinicians should continue to use their clinical acumen, sound clinical judgement and discretion. To avoid premature aetiological closure and broaden treatment options, we also propose renaming this syndrome descriptively as drug-induced hyperthermic catatonia (DIHC). PMID- 11281982 TI - Knockout Corner. AB - Serotonin (5-hydroxytryptamine, 5-HT) is a neurotransmitter involved in a number of physiological functions including sleep, appetite, pain perception, and sexual activity. Several pathological states such as migraine, depression, and anxiety have been linked to the serotonergic system, and serotonergic drugs have been used to treat these disorders. To date, there are 14 known serotonin receptor subtypes through which serotonin exerts its multiple actions. The classic pharmacological approach to study how these individual receptor subtypes contribute to various behaviours has been to use selective drugs that either block or activate certain receptor subtypes, and then study the effects of these compounds on physiology and behaviour. A complementary genetic approach is the technique of gene targeting. Using this technology, we and others have begun to examine the contribution of several serotonin receptor subtypes to complex behaviours through the generation of knockout mice that lack the genes encoding these receptors. In this review, we will describe what we have learned about the serotonergic system and the function of the 5-HT(1B) receptor by the analysis of 5-HT(1B) receptor knockout mice. Furthermore, we will discuss the implications of these findings and our plans for future studies. PMID- 11281983 TI - Treatment of tardive akathisia with clonidine: a case report. AB - Akathisia is characterized by subjective discomfort and motor restlessness. The motor hyperactivity can express itself by frequent changes of posture, constant limb shaking or restless pacing. If symptoms of akathisia are severe, treatment becomes extremely complicated and patients may even become suicidal as seen in the case described here. In the literature, different forms of akathisia are distinguished (Barnes and Braude, 1985): the acute form of neuroleptic-induced akathisia (recent onset, related to an increase in antipsychotic drug dose), pseudoakathisia (motor signs but no subjective symptoms), and chronic or tardive akathisia. The acute form of akathisia is well known and described (Zubenko et al., 1984a,b). In a retrospective analysis of clinical features and therapeutic trials for tardive akathisia, Burke et al. (1989) showed that almost all of the 52 cases developed this chronic form after an average of 4.5 yr following neuroleptic drug initiation, 34% even within 1 yr. Twenty-six of the patients who were able to stop taking dopamine antagonists still had symptoms of akathisia persisting for 0.3-7 yr (mean = 2.7 yr). PMID- 11281984 TI - Differential changes in glutamatergic transmission via N-methyl-D-aspartate receptors in the hippocampus and striatum of rats behaviourally sensitized to methamphetamine. AB - We searched for changes in glutamatergic transmission via N-methyl-D-aspartate (NMDA) receptors in the hippocampus and striatum of rats behaviourally sensitized to methamphetamine (Meth). Prior to being given a challenge dose of Meth (2 mg/kg, s.c.), the rats were given Meth (4 mg/kg, s.c.) five times a week for 3 wk. Seven days after the challenge test, we examined glutamate (Glu) release from hippocampal and striatal slices evoked by 30 mm KCl, and NMDA-evoked dopamine (DA) release from striatal slices. We further immunoquantified NMDAR1, R2A and R2B receptors as well as the fodrin alpha-subunit, a 240 kDa cytoskeletal protein that is cleaved to form 150 kDa limited proteolytic fragments by NMDA receptor stimulation. In the study of KCl-evoked Glu release, Glu release from the hippocampus was 31% lower in the Meth-sensitized rats than in the control rats, while Glu release from the striatum was 34% higher in the Meth-sensitized rats. NMDAR1, R2A and R2B immunoreactivities in the striatum were significantly lower in the Meth-sensitized rats (by 12, 13 and 12%, respectively) than those in the control rats. However, no differences in the immunoreactivities were found for the hippocampus. Immunoquantification of the fodrin alpha-subunit in the hippocampus revealed that 150 kDa fragments were significantly lower (by 10%) in the Meth-sensitized rats than in the control rats. In contrast to the control rats, NMDA-evoked DA release from the striatum was diminished in the Meth sensitized rats. These results indicate that the activity of the Glu system is functionally decreased in the hippocampus of Meth-sensitized rats, whereas the Glu system in the striatum of Meth- sensitized rats shows adaptive and functional changes in the receptors in response to the increased Glu release. PMID- 11281985 TI - Partial generalization of (-)DOM to fluvoxamine in the rat: implications for SSRI induced mania and psychosis. AB - Recent reports have implicated selective serotonin-reuptake inhibitors in the induction of psychosis and mania when SSRIs are given in combination with neuroleptics. We hypothesize that the partial substitution of fluvoxamine for the hallucinogen, (-)DOM, in the rat provides evidence for a 5-HT(2)-mediated effect of fluvoxamine which may in turn account for the adverse effects observed in humans. Male Fischer-344 rats were trained with (-)DOM (0.56 mg/kg) as a discriminative stimulus using standard operant procedures. Tests of generalization were then conducted with fluvoxamine either alone or in combination with the 5-HT(1A) antagonist, WAY-100635, the 5-HT(2) antagonist, pirenperone, and the neuroleptics, fluphenazine, chlorpromazine, thioridazine, loxapine, risperidone, and clozapine. In rats trained with (-)DOM, fluvoxamine at a dose of 20 mg/kg yielded a maximum 58% (-)DOM-appropriate response. This partial generalization was potentiated by treatment with WAY-100635 and antagonized by pirenperone, loxapine, risperidone, and clozapine. The present data are compatible with a 5-HT(2)-mediated effect of fluvoxamine which may play a role in SSRI-induced mania and psychosis. It is predicted by the results of this study that the probability of these adverse effects will be increased by the concurrent use of antagonists at 5-HT(1A) receptors and decreased by neuroleptics with antagonistic activity at 5-HT(2) receptors. PMID- 11281986 TI - Calcitonin gene-related peptide (CGRP) levels and alcohol. AB - Calcitonin gene-related peptide (CGRP) when administered into the brain exerts stress-like effects such as increased pain sensitivity, anorexia, and potentiation of fear-related behaviours. Since alcohol consumption may be related to alcohol's anxiolytic properties, the present study sought to determine if brain CGRP levels were correlated with genetic differences in preference for drinking alcohol and/or affected by alcohol exposure/withdrawal. CGRP-like immunoreactivity (CGRP-LI) was measured by radioimmunoassay (RIA) in amygdala, hippocampus, frontal cortex, hypothalamus, and caudate. In the first experiment, CGRP-LI was compared in alcohol-naive rats [preferring (P) and non-preferring (NP)], lower concentrations were found in the hippocampus (U = 153.5; d.f. = 1,28; p < 0.014) and frontal cortex (U = 183.0; d.f. = 1,28; p < 0.0001) of the P rats. In a second experiment, a group of outbred Wistar rats were exposed to alcohol in vapour chambers, or control conditions. At 7 wk of alcohol exposure there were no differences in exposed rats as compared to controls. However, at 4 wk following ethanol withdrawal, higher concentrations of CGRP-LI were found in the hippocampus (U = 26.5; d.f. = 1,20 p < 0.05), hypothalamus (U = 17.5; d.f. = 1,20; p < 0.009), and caudate-putamen (U = 17.0; d.f. = 1,20; p < 0.009) of the previously exposed animals. These studies suggest that CGRP may modulate alcohol preference and additionally, that exposure/withdrawal from ethanol produces long lasting effects on CGRP-LI. PMID- 11281987 TI - Association between a polymorphism in the promoter region of the dopamine D2 receptor gene and schizophrenia in Japanese subjects: replication and evaluation for antipsychotic-related features. AB - To replicate a previously found negative association between the Del allele of the -141C Ins/Del polymorphism in the 5'-promoter region of the dopamine D2 receptor gene (DRD2) and schizophrenia in Japanese subjects and to examine whether this polymorphism is related to the features of antipsychotic drug treatment, we genotyped 94 control subjects and 234 schizophrenic patients. The schizophrenic patients had a significantly lower frequency of the Del allele (p < 0.05). We found a non-significant trend towards a higher frequency of the Del allele in schizophrenic patients susceptible to neuroleptic-induced extrapyramidal symptoms. The daily dosage of haloperidol, the steady-state concentration of serum haloperidol per daily dosage, and the recent 1-yr cumulative neuroleptic dosage were lower in patients with the Del/Del genotype than in the other patients. These findings support the view that the polymorphism is associated with schizophrenia in Japanese subjects and provide hints for further attempts to establish the relationship between this polymorphism and the features of antipsychotic drug treatment. PMID- 11281988 TI - Cyclic AMP-dependent protein kinase in subtypes of major depression and normal volunteers. AB - We have shown a reduction in beta adrenoceptor-linked, cyclic AMP-dependent protein kinase [protein kinase A (PKA)] activity in fibroblasts of patients with major depression with melancholic features relative to normal volunteers. We evaluated a group of 35 patients with major depression subtyped by DSM-IV criteria as melancholic, atypical, and those not meeting either subtype designation ('non-subtyped') and 21 normal volunteers to ascertain whether or not the PKA activity abnormality was specific to melancholia. The melancholics showed marked reduction in cyclic AMP-stimulated PKA activity relative to normal volunteers. Although the atypicals were statistically significantly lower, almost all fell into the range for the normals. The non-subtyped group fell between the atypicals and the melancholics. Basal activity was significantly lower in atypical and melancholic groups. The data suggest that reduced PKA activity is consistently found in melancholic major depression and may not be seen with other depressive subtypes. PMID- 11281989 TI - Inositol versus placebo augmentation of serotonin reuptake inhibitors in the treatment of obsessive-compulsive disorder: a double-blind cross-over study. AB - Current serotonin reuptake inhibitor (SRI) treatments for obsessive-compulsive disorder (OCD) provide only partial benefit. A previous study suggested that inositol alone is efficacious in OCD. Ten DSM-IV OCD patients completed a study of 18 g inositol or placebo for 6 wk each in addition to ongoing SRI treatment in a double-blind randomized cross-over design. Weekly assessments included the Yale Brown Obsessive-Compulsive Scale (YBOCS) and Hamilton Depression and Anxiety scales. No significant difference was found between the two treatment phases. PMID- 11281990 TI - Dopamine receptors and schizophrenia: contribution of molecular genetics and clinical neuropsychology. AB - Family, twin and adoption studies suggest that genetic factors play an important role in the aetiology of schizophrenia. The mode of inheritance, however, is complex and non-Mendelian. Although the aetiology of schizophrenia is unknown, it has been hypothesized that the necessary conditions for developing the disease are environmental stress and a vulnerability to psychosis. The implication of dopamine receptors to schizophrenia has been greatly studied. Several linkage and association studies have been performed in an attempt to establish the involvement of dopamine receptors in schizophrenia. However, although no conclusive evidence of linkage or association to any gene has been established, some results, suggestive of linkage for chromosomes 6, 22 and 13, await confirmation from other studies. Concerning association studies, it is also of interest that some studies support an association between schizophrenia and homozygosity at D(3). More work in larger samples is required before conclusive linkage hypothesis or association to a dopamine receptor may be established. Schizophrenic patients have been shown to have significant deficits in a wide range of cognitive processes, including memory, attention, reasoning ability and language. Since cognitive deficits are significant symptoms of schizophrenia which require effective treatment, their assessment in schizophrenic patients and during clinical trials of new potential antipsychotics is highlighted. Cognitive impairment in schizophrenia impedes psychosocial performance and is therefore an especially relevant target variable in the development of new therapeutic approaches. It is most prominent in tasks involving attention, memory and executive functions which are thought to reflect involvement of prefrontal and left-temporal brain areas. Semantic networks in schizophrenic patients with a younger age of onset are observed to be more disorganized and differ significantly to those of control subjects. The need to use broader approaches such as neuropsychological-related measures to identify pertinent phenotypes in non-affected subjects carrying vulnerability genes is also emphasized. Since dopamine receptors are the primary targets in the treatment of schizophrenia, improved therapy may be obtained by drugs that selectively target a particular subtype of dopamine receptor. In the development of novel antipsychotics, D(3) and D(4) receptors have received much attention and this is partly related to the fact that these receptors have a high abundance in brain areas associated with cognitive and emotional functions, such as parts of the limbic system and cortex. Recent studies suggest that atypical neuroleptics may significantly improve the cognitive deficits observed in schizophrenic patients and that atypical neuroleptics such as risperidone appear to improve memory and alertness suggesting that further clinical studies are needed to determine the precise influence of antipsychotics on the cognitive system of schizophrenic patients. Such studies could lead to useful insights as to the potential advantages of the newer antipsychotics which appear to have a sparing or beneficial effect on various components of cognitive function. However, the observation that cortical D(2) receptors are important sites of action for antipsychotics, that the cerebral cortex may harbour the common sites of actions of antipsychotics and that the balancing of the opposing actions of D(1) and D(2) receptor regulation may be an appropriate drug treatment suggests that the adjustment of D(1) receptor levels in the cortex may become an important goal of future antipsychotic generation. Such antipsychotics will be able to treat the positive, negative and cognitive deficits of schizophrenia. PMID- 11281991 TI - Does phenylethylamine act as an endogenous amphetamine in some patients? AB - In brain capillary endothelium and catecholaminergic terminals a single decarboxylation step effected by aromatic amino-acid decarboxylase converts phenylalanine to phenylethylamine, at a rate comparable to that of the central synthesis of dopamine. Phenylethylamine, however, is not stored in intra-neuronal vesicles and is rapidly degraded by monoamine oxidase-B. Despite its short half life, phenylethylamine attracts attention as an endogenous amphetamine since it can potentiate catecholaminergic neurotransmission and induce striatal hyperreactivity. Subnormal phenylethylamine levels have been linked to disorders such as attention deficit and depression; the use of selegiline (Deprenyl) in Parkinson's disease may conceivably favour recovery from deficient dopaminergic neurotransmission by a monoamine oxidase-B inhibitory action that increases central phenylethylamine. Excess phenylethylamine has been invoked particularly in paranoid schizophrenia, in which it is thought to act as an endogenous amphetamine and, therefore, would be antagonized by neuroleptics. The importance of phenylethylamine in mental disorders is far from fully elucidated but the evolution of phenylethylamine concentrations in relation to symptoms remains a worthwhile investigation for individual psychotic patients. PMID- 11281992 TI - Knockout Corner: Knockout mice for monoamine oxidase A. AB - A line of transgenic mice was isolated in which transgene integration had caused a deletion in the gene encoding monoamine oxidase A, an enzyme that degrades serotonin and norepinephrine. This has provided an animal model of MAOA deficiency in humans, a condition characterized by borderline mental retardation and impulsive aggression. PMID- 11281993 TI - Blood flow structure related to red cell flow: determinant of blood fluidity in narrow microvessels. AB - The review article deals with phenomena of the blood flow structure (structuring) in narrow microvessels-capillaries and the adjacent arterioles and venules. It is particularly focused on the flow behavior of red blood cells (RBCs), namely, on their specific arrangements of mutual interaction while forming definite patterns of self-organized microvascular flow. The principal features of the blood flow structure in microvessels, including capillaries, include axial RBC flow and parietal plasma layer, velocity profile in larger microvessels, plug (or bolus) flow in narrow capillaries, and deformation and specific behavior of the RBCs in the flow. The actual blood flow structuring in microvessels seems to be a most significant factor in the development of pathological conditions, including arterial hypertension, brain and cardiac infarctions, inflammation, and many others. The blood flow structuring might become a basic concept in determining the blood rheological properties and disorders in the narrow microvessels. No solid theoretical (biorheological) basis of the blood flow structuring in microvessel has been found, but in the future it might become a foundation for a better understanding of the mechanisms of these properties under normal and pathological conditions in the narrowest microvessels 5 to 25 microm large. It is also a topic for further biorheological research directed to find the background of actual physiopathological phenomena in the microcirculation. PMID- 11281994 TI - Changes in cell volume induced by activation of the cyclic amp-dependent chloride channel in guinea-pig cardiac myocytes. AB - The effects of the activation of cyclic AMP-dependent Cl- current (ICl,cAMP) on cell volume were studied at various [K+]o under isosmotic conditions in guinea pig ventricular myocytes. The area of the cell image obtained with videomicroscopy was used as an index of cell volume. I(Cl,cAMP) was activated by adrenaline (5.5 microM). Measurements of the membrane potential (Vm) were performed by the gramicidin-perforated patch-clamp method. At 5.4 mM [K+]o with low [Cl-]o, where Vm was negative to the predicted equilibrium potential of Cl- (ECl), adrenaline sizably decreased the cell area. At high [K+]o with normal [Cl ]o, where Vm was positive to ECl, adrenaline increased the cell area; at 145.4 mM [K+]o the cell area was increased to 110% of control on average (n = 22). The cells swollen in this manner shrank when [Cl-]o was reduced to a low level in the presence of adrenaline. The results indicate that the induction of Cl- influxes (outward I(Cl,cAMP)) or effluxes (inward I(Cl,cAMP)) can lead to a cell swelling or shrinkage, respectively. The addition of BaCl2 (1 mm), a blocker of K+ channels, attenuated the adrenaline-dependent cell swelling, supporting the view that Cl- fluxes must be accompanied by cofluxes of K+ ions to affect the cell volume. The adrenaline-dependent cell swelling was inhibited by antagonizing beta adrenergic stimulation with acetylcholine or by blocking I(Cl,cAMP) channels with glibenclamide, confirming the involvement of I(Cl,cAMP) in the adrenaline response. The results show that the activation of I(Cl,cAMP) can shrink or inflate the cardiac cells under isosmotic conditions, depending on Vm and ECl. PMID- 11281995 TI - Low compliance rather than high reflection of arterial system decreases stroke volume in arteriosclerosis: a simulation. AB - Although various investigators have suggested that the left ventricles of aged subjects suffer from high-frequency reflection, arterial reflection is larger in the low-frequency range because of a larger impedance mismatch. It has not been quantified whether high-frequency reflection rather than low-frequency reflection has larger deleterious effects on stroke volume. We used a computer simulation method to evaluate how increases in high- and low-frequency arterial reflections associated with age-related arterial sclerosis affect left ventricular (LV) pump function. Low-frequency reflections derive principally from the total arterial compliance, and high-frequency reflections result from impedance fluctuations in the high-frequency range. We numerically coupled a time-varying elastance LV model with a variety of arterial impedances to quantitatively evaluate the effects of low- and high-frequency reflections on LV pump performance. When we simultaneously increased low- and high-frequency reflections to levels of sclerotic impedance (type A in Murgo et al., Circulation 62: 105-116, 1980), stroke volume decreased by 4.4%. Further increases of the reflections up to 8 times of the type A impedance lowered stroke volume by 15.9%. This trend was clearly seen with selective increases in low-frequency reflections (3.5 and 20.2% decrease in stroke volume, respectively), but not with those in high-frequency reflections (1.0% decrease and 0.9% increase in stroke volume, respectively). Thus we conclude that the detrimental effect of increases in arterial reflections associated with arterial sclerosis on stroke volume is mild and mainly attributable to decreased compliance rather than to increased high-frequency reflections. PMID- 11281996 TI - Semiquantitative analysis of the expression of GABA-A receptor subunits in the developing embryonic chick brain stem. AB - The expression levels of seven types of gamma-aminobutyric acid-A (GABA-A) receptor subunits (alpha1, beta2, beta3, beta4, gamma1, gamma2, and gamma4) were quantified in the embryonic chick brain stem at 2 to 20 d of incubation (E2 to E20) and just after hatching. The expression level of mRNA was measured by using semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). When property regions were amplified, two products were observed for each of the beta2, beta4, and gamma2 subunits because of alternative splicing. These were named beta2S and beta2L, beta4S and beta4L, and gamma2S and gamma2L for shorter and longer fragments, respectively. Transcripts of alpha1, beta2L, beta2S, beta3, beta4L, beta4S, gamma1, and gamma2S subunits were first detected from E2 to E5 brain stems. The expression level of each subunit increased gradually with development and reached a plateau at E9 to E12. In contrast, a delay occurred in the appearance of both the gamma4 and gamma2L subunits, which were not detected until E8 to E10. The absence of gamma4 and/or gamma2L subunits may explain differences in the pharmacological characteristics of GABA-A receptors at the early stages of development. PMID- 11281997 TI - Effect of VMH lesion on sucrose-fed analgesia in formalin pain. AB - The ingestion of sucrose (ad libitum) produces an immediate analgesic response to phasic noxious stimuli. The underlying mechanism for the analgesic effect of sucrose is attributed to its palatability, which mediates analgesia probably by the release of beta-endorphin in the hypothalamus. The present study was designed to explore the role of ventromedial hypothalamus in the mediation of sucrose-fed analgesia. Adult male albino rats each received (20%) sucrose solution orally through a separate bottle until they had ingested 4-5 ml. Their behavioral responses to tonic noxious stimulus in a formalin test were studied in pre- and postsucrose-fed rats of control and in the VMH lesion groups. The average pain rating of a 60-min session significantly (p < 0.01) decreased after sucrose feeding in control rats, from 1.94 +/- 0.13 to 1.45 +/- 0.14, but sucrose feeding by the VMH lesion rats did not alter their tonic nociceptive response from a 1.70 +/- 0.07 presucrose-fed state to a 1.71 +/- 0.08 postsucrose-fed state. VMH lesion per se did not alter the nociceptive response in comparison with controls. The results suggest that sucrose feeding produces analgesia to tonic noxious stimulus, which is abolished by lesion of the VMH, thereby indicating a significant role of VMH in sucrose-fed analgesia. PMID- 11281998 TI - Effects of hypothermia on blood flow and neural activity in rabbit spinal cord during postischemic reperfusion. AB - The effects of hypothermia on blood flow and neural activity were investigated in rabbit spinal cord during the acute phase of ischemia/reperfusion. Rabbits were exposed to ischemia for 10 or 40 min by occluding the abdominal aorta, using a balloon catheter. The body temperature was maintained either at 38 degrees C (normothermia) or 34 degrees C (hypothermia). Hyperperfusion was observed within 10 min after the cessation of ischemia in all rabbits exposed to ischemia. The magnitude of hyperperfusion in spinal cord blood flow (SCBF) was not significantly different between the 10 and 40 min ischemia rabbits, but the time for 50% recovery from the hyperperfusion was longer in the 40 min ischemia group (26.1 +/- 2.5 min) than in the 10 min group (15.1 +/- 2.1 min). The amplitude of evoked spinal cord potential decreased during ischemia and recovered to the baseline level during 8 h of reperfusion in the 10 min ischemia group. However, in the 40 min ischemia group, the amplitude was 40 +/- 8% of the baseline value after 8 h of reperfusion. Hypothermia prevented the delay of recovery from hyperperfusion and the reduction of evoked spinal cord potential. These results suggest that hypothermia plays a beneficial role in protecting tissue injury in the acute phase of ischemia/reperfusion in the spinal cord by shortening the time for recovery from postischemic hyperperfusion. PMID- 11281999 TI - Effects of long-term acidification of extracellular pH on ATP-induced calcium mobilization in rabbit lens epithelial cells. AB - ATP-induced calcium (Ca2+) mobilization was investigated in rabbit lens epithelial cells that had been cultured in a medium with pH of 7.4 (group 1), 7.2 (group 2), or 7.0 (group 3) for 10 to 21 d. Intracellular free Ca2+ ([Ca2+]i and pH (pHi) were measured by using fluorescent dyes, fura-2 and BCECF, respectively. The long-term acidification decreased the pHi to 7.15 +/- 0.01, from 7.22 +/- 0.01, in group 2 and to 7.09 +/- 0.01 in group 3. The administration of 10 micromol/l ATP produced an initial peak followed by a sustained increase in [Ca2+]i in the lens cells of group 1. Both the initial peak and the sustained increase in [Ca2+]i were enhanced in groups 2 and 3. The initial peak was abolished by pretreatment with 1 micromol/l thapsigargin, an ER Ca2+ pump inhibitor, but was not affected by the removal of extracellular Ca2+. On the other hand, the sustained increase was suppressed either by the thapsigargin treatment or by the Ca2+ removal. Treatment with only thapsigargin caused a sustained increase in [Ca2+]i that was greater in group 3 than in group 1. These results suggest that (1) the ATP-induced initial peak in [Ca2+]i is due to Ca2+ release from the intracellular stores, (2) the sustained increase in [Ca2+]i is mediated through either Ca2+ influx from the extracellular space or Ca2+ release from the store triggered by the Ca2+ influx, and (3) long-term, moderate acidification enhances both the initial peak and the sustained increase in [Ca2+)]i in rabbit lens epithelial cells. One possible mechanism of the ATP induced Ca2+ influx seems to be a capacitative Ca2+ entry pathway. PMID- 11282000 TI - Effects of aging on the electroretinogram during ischemia-reperfusion in rats. AB - The effects of aging on the electroretinogram (ERG) during ischemia-reperfusion were investigated in rats. Flash-elicited ERG (a-wave, b-wave, and oscillatory potentials (OPs)) was recorded in young (4 months old) and aged rats (over 18 months old) before, during, and after exposure to 30- or 120-min ischemia induced by increasing intraocular pressure to 80 mmHg. The choroidal blood flow, measured by means of laser Doppler flowmetry, decreased to 40 to 60% of the baseline value during ischemia. Young rats showed no significant difference in the amplitude of each ERG component during ischemia between 30- and 120-min ischemia groups; 78.0 +/- 4.9 vs. 76.1 +/- 3.6% for a-wave, 63.4 +/- 3.1 vs. 60.6 +/- 3.0% for b-wave, and 59.6 +/- 5.9 vs. 57.5 +/- 6.7% for SigmaOP. In aged rats, however, 120-min ischemia caused a greater decrease, to 56.7 +/- 3.1% of the baseline value, in the a-wave amplitude than 30-min ischemia did, to 70.8 +/- 3.2%. The reduction of each ERG component in both 30- and 120-min ischemia experiments was greater in aged rats than in young rats. The recovery time for the amplitude of each ERG component during reperfusion was longer in aged rats than in young rats. The latency of b-wave and the second component of OPs prolonged during ischemia, and recovery time for the latency was longer in aged rats than in young rats. These results suggest that the electrophysiological function of the retina is less tolerable against ischemia-reperfusion in aged rats than in young rats. PMID- 11282001 TI - Oxygen wasting for Ca2+ extrusion activated by partial inhibition of sarcoplasmic reticulum Ca2+ -atpase by cyclopiazonic acid in rat left ventricles. AB - In the excised Langendorff-perfused rat whole-heart preparation, a linear relation between left ventricular myocardial oxygen consumption per beat (Vo2) and systolic pressure-volume area (PVA, a total mechanical energy per beat) is obtained from a curved end-systolic pressure-volume relation as in the blood perfused preparation. The ordinate Vo2 intercept of the Vo2-PVA relation is composed of Vo2 for total Ca2+ handling in the excitation-contraction coupling and basal metabolism. The Vo2 for total Ca2+ handling is mainly consumed by sarcoplasmic reticulum (SR) Ca2+ -ATPase. The aim of the present study was to investigate, in terms of left ventricular mechanoenergetics, how an inhibition of SR Ca2+ -ATPase by cyclopiazonic acid (CPA; 4 micromol/l) affects Ca2+ handling mechanisms in the excised Langendorff-perfused rat whole-heart preparation. The short-term (for 3 to 6 min after onset of the infusion) CPA infusion decreased Vo2 proportionally to the decrease in PVA. The long-term (for 9 to 12 min after the short-term CPA infusion) CPA infusion gradually increased Vo2 almost to the control level with an increase in PVA. The increases in both Vo2 and PVA during this infusion were completely abolished by a Na+/Ca2+ exchanger inhibitor, 3'9,4'9-dichlorobenzamil, indicating the contribution of Na+/Ca2+ exchanger to the increases in Vo2 and PVA. The O2 cost of left ventricular contractility during the long-term CPA infusion was significantly higher than during the short term CPA infusion. All these results suggest the possibility of the contribution of greater energy-wasting Ca2+ extrusion processes (such as Na+/K+-ATPase coupled to the Na+/Ca2+ exchanger; its stoichiometry is 1 ATP : 1 Ca2+ to the larger oxygen cost of left ventricular contractility. PMID- 11282002 TI - Regional myocardial function at the papillary muscle insertion site. AB - The importance of the mitral apparatus to the global left ventricular (LV) function has been suggested in several clinical studies. One recent study reported that chordal transsection induced an unloading of myocardium at the papillary muscle insertion site. We hypothesized that the regional response for afterloading at this site with intact mitral apparatus was different from that at the free wall. We investigated the end-systolic pressure-regional segment length relations (ESPLR) in two anterior LV sites, free wall (FREE) and the papillary muscle insertion site (PAP), during an increasing afterload by aortic occlusion in 7 anesthetized open-chest dogs. To measure the regional segment length at FREE and PAP, two sets of the pair of sonomicrometer crystals were implanted in the same midwall depth at the same circumferential hoop by using an echocardiographic guide. ESPLR both at FREE and PAP were always highly linear in a physiological range (r > or = 0.9). The slope of this relation at FREE (274 +/- 164 mmHg/mm) was significantly steeper than that at PAP (157 +/- 118 mmHg/mm) for each dog (p < 0.05). These data indicate that the regional response for afterloading at PAP loaded by chordal tension is different from that at FREE in the same heart. PMID- 11282003 TI - Lack of tyrosine protein kinase regulation of L-type Ca2+ channel current in transfected cells stably expressing alpha1C-b Subunit. AB - Tyrosine protein kinase (Tyr-PK) regulation of L-type Ca2+ channel (CaL) current was studied in COS-7 cells expressing vascular smooth muscle-type alpha1C-b with no auxiliary subunit by using a whole-cell voltage clamp. The averaged peak amplitude of CaL currents was -0.33 +/- 0.03 at holding potential of -60 mV. Na(3)VO(4), genistein and phosphorylated p60(c-src) peptide had no effect on the current. Thus the alpha1C-b subunit may not be involved in Tyr-PK regulation of CaL current. PMID- 11282004 TI - Is rabbit CLCA1 related to the basolateral Ca2+ -dependent Cl- channel of gastric parietal cells? AB - An expression of mRNA coding the calcium-activated Cl- channel-1 (CLCA1) in rabbit gastric parietal cells was examined to verify the possibility that the CLCA1 mediates housekeeping Cl- channels in the basolateral membrane. In whole cell voltage-clamp experiments of rabbit parietal cells, A23187 (2 microM), a Ca2+ ionophore, activated the basolateral Cl- channels. The partial cDNA fragment of rabbit CLCA1 could be amplified from the total RNA of tracheal epithelium. A Northern blot analysis showed that rabbit CLCA1 mRNA of 3.4 kb is highly expressed in the tracheal epithelium, but not in the gastric parietal cells. Even in a more sensitive detection of rabbit CLCA1 mRNA by RT-PCR, no signal could be observed in the gastric parietal cells. These results suggest that the CLCA1 protein may not be a subunit of the housekeeping Ca2+ -dependent Cl- channel in the basolateral membrane of rabbit gastric parietal cells. PMID- 11282005 TI - Calcium waves in skinned cardiac myocytes evoked by two-photon excitation photolysis of caged calcium. AB - In rat ventricular myocytes chemically skinned with saponin, a local rise of [Ca2+] was achieved by two-photon excitation photolysis (TPP) of the caged Ca2+ compound 1-(2-nitro-4,5-dimethoxyphenyl)-N,N,N',N'-tetrakis[(oxycarbonyl)methyl] 1,2-ethanediamine (DM-nitrophen). Confocal Ca2+ images, by use of fluo-3, were simultaneously collected. TPP of DM-nitrophen induced Ca2+ waves propagating over the myocyte, and the local rise of [Ca2+] at the site of photolysis sustained for 50-60 ms. These TPP-induced Ca2+ events were completely suppressed by ryanodine (10 microM), suggesting that Ca2+ release resulting from TPP of DM-nitrophen triggered regenerative Ca2+ release from the neighboring sarcoplasmic reticulum. The present techniques should be useful to investigate the interaction of elementary Ca2+ events, the process leading to global Ca2+ movements, in cardiac myocytes and other types of cells. PMID- 11282006 TI - Effect of genotypic resistance on the virological response to highly active antiretroviral therapy in cerebrospinal fluid. AB - Paired plasma and cerebrospinal fluid (CSF) specimens drawn from 15 HIV-infected patients with neurological disease before and after a median 6-week duration of highly active antiretroviral therapy (HAART) were studied to assess the short term virological response of CSF and whether this can be predicted on the basis of baseline resistance mutations. After treatment, the median plasma and CSF viral load (VL) decreased by, respectively, 2.08 log10 (p = 0.0001) and 0.91 log10 copies/ml (p = 0.007) in comparison with baseline. A plasma virological response was observed in all but one patient, whereas the posttreatment CSF VL increased, remained unchanged, or decreased at a substantial lower rate than in plasma of six "CSF non/slow responders" (40%). Direct sequencing of baseline specimens showed that none of these patients had reverse transcriptase (RT) or primary protease resistance mutations in the CSF alone, but two had RT mutations conferring high-level resistance to drugs included in the HAART regimen in both CSF and plasma. The other four patients had no RT or primary protease resistance mutations. There was no significant difference in the nucleotide diversity of the CSF and plasma RT sequences, baseline plasma or CSF VL, the CSF-to-plasma VL ratio, the number of CSF cells, the CD4+ cell counts, or the history of antiretroviral treatment between the CSF non-slow responders and the other patients. During this short-term follow-up and despite a plasma response, a significant proportion of HAART-treated patients with neurological symptoms showed a slow or absent CSF response. Most of these cases were not associated with the presence of resistant HIV strains in the CSF. PMID- 11282007 TI - Multivalent anti-CCR ribozymes for stem cell-based HIV type 1 gene therapy. AB - HIV-1 infection of susceptible cells is mediated by the specific interaction of viral envelope glycoproteins with the cell surface CD4 receptor and a chemokine coreceptor, CCR5 or CXCR4. Individuals with a CCR5 genetic defect show resistance to HIV-1 infection, indicating that downregulation of CCR5 expression on target cells can prevent viral infection. In previous studies we demonstrated the utility of an anti-CCR5 ribozyme targeted to a single cleavage site in downregulating CCR5 expression and consequently providing resistance to viral infection. To improve on the level of downregulation we designed a construct containing an anti-CCR5 ribozyme heterotrimer (R5RbzTM) targeted to three different cleavage sites in CCR5 mRNA. In vitro tests showed that the anti-CCR5 ribozyme heterotrimer could effectively cleave the CCR5 RNA substrates to yield products of the expected sizes. This construct was introduced into various retroviral vectors for stable gene transduction. HOS.CD4/R5 cells stably transduced with this anti-CCR5 heterotrimer showed a marked reduction in the surface expression of CCR5 and a concomitant 70% reduction in macrophage-tropic viral infection. In addition, a retroviral vector containing the anti-CCR5 ribozyme heterotrimer and an anti-HIV-1 tat-rev ribozyme heterodimer was constructed. This construct also showed a similar inhibition of CCR5 surface expression and reduced infectability by the macrophage-tropic HIV-1 vector in HOS.CD4/R5 cells. The trimeric and multimeric ribozyme constructs were transduced into CD34+ hematopoietic progenitor cells to determine their effects on lineage specific differentiation. We show that multivalent ribozyme gene-transduced hematopoietic progenitors differentiated normally into mature macrophages that bear CD14 and CD4 surface markers. Macrophages containing the transgenes expressed ribozymes, and showed resistance to M-tropic HIV-1 infection. These results provide strong support for the use of the trimeric anti-CCR5 ribozyme approach in a gene therapy setting for the treatment of HIV infection. PMID- 11282008 TI - Anti-HIV type 1 activity of 3'-fluoro-3'-deoxythymidine for several different multidrug-resistant mutants. AB - The objective of this work was to test the antiviral activity of a potent nucleoside reverse transcriptase inhibitor, 3'-fluoro-3'-deoxythymidine (FLT), on both a wild-type human immunodeficiency virus (HIV-1) isolate and multidrug resistant HIV-1 patient isolates. Drug-resistant viral isolates were selected on the basis of four different categories of well-characterized and representative multidrug-resistant mutants. The isolates included three variants containing 151M alone or in combination; three variants containing 215Y and 41L, 67N, 184V, 210W, and 219N in combination; two insertion mutant viruses (69 + EA and 69 + SA); and two deletion mutant viruses (del67NG and del67GS), the latter two groups both also containing other significant mutations. The activity of FLT and AZT against these isolates was determined by drug susceptibility assays and by measuring viral antigen p24 by ELISA. The cytotoxicity of FLT and AZT was assessed in PHA stimulated PBMCs. Development of resistant mutants under FLT pressure was attempted by passaging HIV-1 isolates in SupT1 cells and stepwise increasing the concentration of FLT. The multidrug-resistant mutant HIV-1 isolates exhibited 7 fold to >100-fold increased resistance to AZT, but showed IC(50) values for FLT of 0.0014-0.0168 microM, which were lower than or similar to that of wild type (0.0075 microM). The cellular cytotoxicities of FLT and AZT fell into a similar range in PBMCs. The development of HIV mutants resistant to FLT appeared to be slower than for other RT inhibitors. HIV isolates with mutations resulting in multidrug resistance had no evidence of resistance to FLT. FLT may be useful in salvage therapies for patients harboring resistant strains and a reassessment of its therapeutic potential seems required. PMID- 11282010 TI - Molecular evidence of homosexual transmission of HIV type 2 in Spain. AB - Eight HIV-2-infected Caucasian men living in the same geographical area in Gipuzkoa (northern Spain) have been identified in the last 5 years. HIV-2 infection in this area is uncommon, and no other cases of HIV-2 infection have been found after extensive testing for HIV-1/2 antibodies. Epidemiological data suggested a possible link among the identified subjects, with homosexual contact being the most likely way of transmission. A genetic analysis of four of the subjects, from whom specimens were available, was conducted. Phylogenetic and signature pattern studies of the reverse transcriptase (RT) and env genes supported a single source of infection. Interindividual nucleotide variability ranged from 2.4 to 4.8% in the RT region and from 5.2 to 6.1% in the env gene, whereas the mean divergence between patient and control strains was 9.8 and 18.3%, respectively. The nucleotide and amino acid signature patterns were closely related in viruses from the four examined individuals. This is the first report of a cluster of HIV-2 infections with genetic sequence data support. The singularity of this cluster should alert clinicians on the possibility of HIV-2 outside endemic areas. PMID- 11282009 TI - Initiation of therapy during primary HIV type 1 infection results in a continuous decay of proviral DNA and a highly restricted viral evolution. AB - A latent pool of HIV-1 is established early in memory CD4+ T lymphocytes and persists during antiretroviral therapy. Also, viral replication may continue in subjects despite undetectable viremia. However, it remains unclear whether this residual replication results in any significant sequence evolution. We were therefore interested in studying the viral evolution and HIV-1 DNA dynamics in subjects with primary infection receiving or not receiving early potent antiretroviral therapy. In 16 subjects, HIV-1 DNA load was monitored from 1 to 23 days, up to 1253 days, after onset of symptoms. Extensive sequential cloning and sequence analysis of the V3 region was performed in four subjects. In the treated subjects a continuous decline in the proviral load was found, corresponding to a half-life of about 6 months. As expected in newly infected individuals the founder virus populations showed high intrasubject sequence similarity. Also, a limited increase in the viral divergence was detected during the first 6 months in three treated subjects. Thereafter, no significant sequence changes were found despite analysis of a large number of clones. Our data thus suggest that early and successful therapy in compliant subjects with primary HIV-1 infection results in a highly restricted viral evolution and a decline in the proviral load close to the decay rate of human memory T lymphocytes. PMID- 11282011 TI - A new human immunodeficiency virus type 1 circulating recombinant form from Tanzania. AB - It is becoming increasingly important to identify and to study human immunodeficiency virus type 1 (HIV-1) circulating recombinant forms (CRFs) with evidence of epidemic spread, since mosaic strains arise frequently, especially in populations where multiple subtypes cocirculate. We describe the almost complete nucleotide sequence of 3 subtype C and D recombinant viruses, selected from a pool of 13 D(gag)-D/C/D(env) perinatally infected infants from Dar es Salaam, Tanzania. All three genomes had cross-over points with approximately the same genomic localization. The subtype C-like sequences were located within pol, vif, vpr, vpu, the first exons of rev and tat, V3, and the U3-R regions of the LTR. Phylogenetic analyses of the full-length genomic sequences from these viruses showed the formation of a distinct subcluster on the HIV-1 subtype D branch. The pattern of recombination of genomes belonging to this new CRF, named CRF10_CD, might have resulted from independent recombination events occurring at high frequency or from a single source that originated earlier in this population. Future surveys will be needed to determine the potential of this CRF for epidemic spread. PMID- 11282012 TI - No difference in in vitro susceptibility to HIV type 1 between high-risk HIV negative Ethiopian commercial sex workers and low-risk control subjects. AB - Host factors such as increased beta-chemokine production, HIV-1 coreceptor expression level, and HIV-1 coreceptor polymorphism have been thought to influence susceptibility to HIV-1 infection. To determine the protective role of these factors in Ethiopians who remained HIV-1 uninfected, despite multiple high risk sexual exposures, we studied 21 Ethiopian women who had been employed as commercial sex workers (CSWs) for five or more years. The HIV-1-resistant CSWs were compared with low-risk age-matched female controls who had a comparable CD4+ cell percentage and mean fluorescence intensity (MFI). Genetic polymorphism in the CCR5, CCR2b, or SDF-1 genes appeared not to be associated with resistance in the Ethiopian CSWs. Expression levels of CCR5 and CXCR4 on naive, memory, and total CD4+ T cells tended to be higher in the resistant CSWs, while the production of beta-chemokines RANTES, MIP-1alpha, and MIP-1beta by phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMCs) was lower compared with low-risk HIV-1 negative controls. In vitro susceptibility of PHA-stimulated PBMCs to primary, CCR5-restricted, Ethiopian HIV-1 isolates was comparable between resistant CSWs and low-risk controls. In vitro susceptibility was positively correlated to CD4+ cell mean fluorescence intensity and negatively correlated to CCR5 expression levels, suggesting that infection of PBMCs was primarily dependent on expression levels of CD4 and that CCR5 expression, above a certain threshold, did not further increase susceptibility. Our results show that coreceptor polymorphism, coreceptor expression levels, beta-chemokine production, and cellular resistance to in vitro HIV-1 infection are not associated with protection in high-risk HIV-1-negative Ethiopian CSWs. PMID- 11282013 TI - Increased expression of CD23 (Fc(epsilon) receptor II) by peripheral blood monocytes of aids patients. AB - Monocytes expressing the Fcepsilon receptor II (CD23) play important roles in inflammatory and allergic immune responses. We found that peripheral blood monocytes of AIDS patients express increased levels of CD23, compared with monocytes of healthy HIV-1-seronegative individuals (controls) (p < 0.05). We compared expression of monocyte CD23 with expression of monocyte Fcgamma receptors (CD16, CD32, CD64), plasma/serum levels of IgE (also IgM, IgG, IgA), and Th1 (IFN-gamma) and Th2 (IL-4, IL-10) cytokines. We found that monocyte CD23 expression directly correlated with monocyte CD16 expression (p < 0.01, R = 0.58), which was also increased in AIDS patients; there was no correlation with CD32 or CD64 or with soluble factors in plasma/serum (i.e., IgE, IL-4, IL-10, and IFN-gamma). Interestingly, despite the known ability of IL-10 to downregulate monocyte CD23 expression, plasma IL-10 levels were increased in these AIDS patients compared with controls (p < 0.05). We thus evaluated the effect of AIDS and control plasma or rhIL-10 to regulate CD23 expression by monocytes in cultures (24 hr) of healthy human cells +/- treatment with anti-IL-10R blocking antibody. We found that anti-IL-10R blocking antibody treatment had no effect on monocyte CD23 expression in cultures containing AIDS plasma, but increased monocyte CD23 expression in cultures containing control plasma (p < 0.05) or rhIL 10. In conclusion, the identification of increased monocyte CD23 expression in AIDS patients may further characterize the aberrant activated phenotype of monocytes during the immunopathogenesis of HIV-1 disease. Further, monocyte CD23 expression does not appear to be suppressed by the IL-10-enriched environment in AIDS. PMID- 11282014 TI - Evaluation of a sensitive/less-sensitive testing algorithm using the 3A11-LS assay for detecting recent HIV seroconversion among individuals with HIV-1 subtype B or E infection in Thailand. AB - The development of a serologic algorithm to determine recent HIV seroconversion, using sensitive/less-sensitive testing strategies, has generated widespread interest in applying this approach to estimate HIV-1 incidence in various populations around the world. To evaluate this approach in non-B subtypes, longitudinal specimens (n = 522) collected from 90 incident infections among injecting drug users in Bangkok (subtype B infection, n = 18; subtype E infection, n = 72) were tested by the 3A11-LS assay. Standardized optical density (SOD) was calculated, using median values, and the window period between seroconversion as determined by sensitive and less sensitive tests was estimated by a maximum-likelihood model described previously. Our results show that the mean window period of the 3A11-LS assay was 155 days (95% CI, 128-189 days) for subtype B but was 270 days (95% CI, 187-349 days) for subtype E specimens from Thailand. About 4% of individuals with incident subtype E infections remained below the threshold (SOD of 0.75), even 2 years after seroconversion. Among the patients with clinical AIDS and declining antibodies, none of the 7 individuals with subtype B, but 10 (8.7%) of 115 with subtype E infections, were misclassified as recent infections. Lowering the cutoff to an SOD of 0.45 for subtype E specimens resulted in a mean window period of 185 days (95% CI, 154-211 days), with all individuals seroconverting, and reduced the number of subtype E infected patients with AIDS who were misclassified as having recent infection to 2.6%. Our results demonstrate that the 3A11-LS assay has different performance characteristics in detecting recent infections among individuals infected with subtypes B or E. Determining appropriate cutoffs and mean window periods for other HIV-1 subtypes will be necessary before this approach can be reliably implemented in settings where non-B subtypes are common. PMID- 11282015 TI - Variation in simian immunodeficiency virus env V1 region in simian AIDS associated lymphoma. AB - Genetic variation of SIV env during the course of infection provides a large population pool that is continually shaped by selective forces in vivo and may influence the development of clinical disease. SAIDS-associated lymphoma (SAL) in the SIV-infected macaque is typically a clonal or oligoclonal mass of B cell origin, extranodal in anatomic distribution, in which SIV is restricted largely to infiltrating macrophages. To explore the degree of genetic variation in SIV env represented in SAL, a 480-bp DNA fragment containing the V1 region was PCR amplified from seven cases of SAL and from a nonneoplastic lymph node of an SIV infected macaque. The nucleotide sequence of the V1 region was determined from at least 10 clones from multiple independent amplification reactions of each tissue. Overall, the degree of V1 variability within lymphomas was found not to be restricted but to resemble the heterogeneity reported in SIV-infected lymphoid and other tissues. V1 variation in the nonneoplastic lymph node was unexpectedly limited, perhaps related to the unusual disease condition associated with SAIDS in that animal. Unlike observations from SIV-infected tissues of animals without neoplastic disease, no increase was detected in the number of O- or N-linked glycosylation sites in the V1 regions isolated from lymphomas as compared with the original inoculum. These findings suggest that, within the microenvironment of the lymphoma, the immune evasion conferred by increased glycosylation may offer little selective advantage. PMID- 11282016 TI - Recombinant strains of HIV type 1 in the United Kingdom. AB - Twenty-five recombinant (mosaic) HIV-1 genomes were detected among 151 samples comprising 118 non-B subtype sequences and 33 samples containing subtype B sequences. Seven of the 25 mosaic patterns were similar to characterized circulating recombinant forms (two A/E, four A/G, and one D/F) and one was a MAL like A/D recombinant. Eighteen of the recombinants had evidence of subtype A sequences in at least one region of their genome. One sample was found to contain a novel recombinant form (pol F, env K). Two samples could not be characterized unambiguously as recombinant forms and a further one appeared to be a complex C/J/D/A genomic form. The majority of the mosaic genomes were recombinants between gag, pol, or env, whereas the C/J/D/A mosaic had cross-over breakpoints within pol. These findings suggest that almost 20% of non-B subtype isolates of HIV-1 circulating in the United Kingdom have mosaic genomes. This shows the diverse origin of HIV-1 strains circulating in the United Kingdom and may have implications for antiretroviral drug resistance. PMID- 11282018 TI - Insulin signalling. PMID- 11282019 TI - AKAP signaling complexes at the cytoskeleton. AB - Targeting of protein kinases and phosphatases to the cytoskeleton enhances the regulation of signal transduction events. The assembly of cytoskeletal signaling complexes facilitates the relay of messages from membrane receptors to specific sites on the actin cytoskeleton. These signals influence fundamental cell properties, such as shape, movement and division. Targeting of the cAMP-dependent kinase (PKA) to the cytoskeleton is achieved through interaction with A-kinase anchoring proteins (AKAPs). AKAPs maintain multivalent signaling complexes by binding additional enzymes, including kinases and phosphatases. PMID- 11282020 TI - Phosphoinositide 3-kinase signalling pathways. AB - Phosphoinositide 3-kinases (PI3Ks) phosphorylate the 3'-OH position of the inositol ring of inositol phospholipids, producing three lipid products: PtdIns(3)P, PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3). These lipids bind to the pleckstrin homology (PH) domains of proteins and control the activity and subcellular localisation of a diverse array of signal transduction molecules. Three major classes of signalling molecule are regulated by binding of D-3 phosphoinositides to PH domains: guanine-nucleotide-exchange proteins for Rho family GTPases, the TEC family tyrosine kinases such as BTK and ITK in B and T lymphocytes, respectively, and the AGC superfamily of serine/threonine protein kinases. These molecules are activated by a variety of extracellular stimuli and have been implicated in a wide range of cellular processes, including cell cycle progression, cell growth, cell motility, cell adhesion and cell survival. PMID- 11282021 TI - Initiating DNA synthesis: from recruiting to activating the MCM complex. AB - The exact duplication of a genome once per cell division is required of every proliferating cell. To achieve this goal, eukaryotes adopt a strategy that limits every replication origin to a single initiation event within a narrow window of the cell cycle by temporally separating the assembly of the pre-replication complex (pre-RC) from the initiation of DNA synthesis. A key component of the pre RC is the hexameric MCM complex, which is also the presumed helicase of the growing forks. An elaborate mechanism recruits the MCM complex to replication origins, and a regulatory chain reaction converts the poised, but inactive, MCM complex into an enzymatically active helicase. A growing list of proteins, including Mcm10 and Cdt1, are involved in the recruitment process. Two protein kinases, the Cdc7-Dbf4 kinase (DDK) and the cyclin-dependent kinase (CDK), trigger a chain reaction that results in the phosphorylation of the MCM complex and finally in the initiation of DNA synthesis. A composite picture from recent studies suggests that DDK is recruited to the pre-RC during G1 phase but must wait until S phase to phosphorylate the MCM complex. CDK is required for the recruitment of Cdc45 and other downstream components of the elongation machinery. PMID- 11282022 TI - Differential effect of GalNAcalpha-O-bn on intracellular trafficking in enterocytic HT-29 and Caco-2 cells: correlation with the glycosyltransferase expression pattern. AB - Our previous work has shown that long-term treatment of mucus-secreting HT-29 cells with 1-benzyl-2-acetamido-2-deoxy-alpha-D-galactopyranoside (GalNAcalpha-O bn), a competitive inhibitor of O-glycosylation, induced several phenotypic changes, in particular a blockade in the secretion of mucins, which are extensively O-glycosylated glycoproteins. Here, we have analyzed the effects of GalNAcalpha-O-bn upon the intracellular trafficking of basolateral and apical membrane glycoproteins at the cellular and biochemical levels in two types of cells, HT-29 G(-) and Caco-2, differentiated into an enterocyte-like phenotype. In HT-29 G(-) cells, but not in Caco-2 cells, DPP-IV and CD44 failed to be targeted to the apical or basolateral membrane, respectively, and accumulated inside intracytoplasmic vesicles together with GalNAcalpha-O-bn metabolites. We observed a strong inhibition of alpha2,3-sialylation of glycoproteins in HT-29 G( ) cells correlated to the high expression of alpha2,3-sialyltransferases ST3Gal I and ST3Gal IV. In these cells, DPP-IV and CD44 lost the sialic acid residue substituting the O-linked core 1 structure Galbeta1-3GalNAc (T-antigen). In contrast, sialylation was not modified in Caco-2 cells, but a decrease of alpha1,2-fucosylation was observed, in correlation with the high expression of alpha1,2-fucosyltransferases Fuc-TI and Fuc-TII. In conclusion, in HT-29 G(-) cells, GalNAcalpha-O-bn induces a specific cellular phenotype, which is morphologically characterized by the formation of numerous intracellular vesicles, in which are accumulated defectively sialylated O-glycosylproteins originally targeted to basolateral or apical membranes, and GalNAcalpha-O-bn metabolites. PMID- 11282023 TI - Avidin expression during chick chondrocyte and myoblast development in vitro and in vivo: regulation of cell proliferation. AB - Avidin is a major [35S]methionine-labeled protein induced by bacterial lipopolysaccharide (LPS) and interleukin 6 (IL-6) in cultured chick embryo myoblasts and chondrocytes. It was identified by N-terminal sequencing of the protein purified from conditioned culture medium of LPS-stimulated myoblasts. In addition, avidin was secreted by unstimulated myoblasts and chondrocytes during in vitro differentiation; maximal expression being observed in differentiated myofibers and hypertrophic chondrocytes. In developing chick embryos, immunohistochemistry revealed avidin in skeletal muscles and growth plate hypertrophic cartilage. Avidin was secreted into culture as a biologically active tetramer. Exogenous avidin added to the medium of proliferating chondrocytes progressively inhibited cell proliferation, whereas addition of avidin to differentiating chondrocytes in suspension allowed full cell differentiation. No toxic effects for the cells were observed in both culture conditions. Western blots of samples from cytosolic extracts using alkaline-phosphatase-conjugated streptavidin showed three biotin-containing proteins. Acetyl-CoA carboxylase was identified by specific antibodies. Based on these data, we propose that avidin binds extracellular biotin and regulates cell proliferation by interfering with fatty acid biosynthesis during terminal cell differentiation and/or in response to inflammatory stimuli. PMID- 11282024 TI - Synergistic effects of different bone morphogenetic protein type I receptors on alkaline phosphatase induction. AB - Bone morphogenetic proteins (BMPs) are members of the transforming growth factor beta superfamily, which regulate the differentiation of osteoprogenitor cells. Here we show that among members of the BMP family, BMP-4 and growth/differentiation factor 5 (GDF-5) induce osteoblast differentiation through the activation of three receptor-regulated Smads (i.e. Smad1, Smad5 and Smad8). By contrast, BMP-6 and BMP-7 induce alkaline phosphatase activity through Smad1 and Smad5, but not through Smad8. Consistent with these findings, BMP-4 induced phosphorylation and nuclear translocation of Smad1, Smad5 and Smad8, but BMP-6 activated only Smad1 and Smad5. BMP-4 and GDF-5 are known to bind to activin receptor-like kinase 3 (ALK-3) and/or ALK-6 (also termed BMP type IA and type IB receptors, respectively), whereas BMP-6 and BMP-7 preferentially bind to ALK-2. Compared with the effects induced by only one of the type I receptors, the combination of constitutively active forms of ALK-2 and ALK-3 (or ALK-6) more strongly induced alkaline phosphatase activity in C2C12 cells. Moreover, addition of BMP-4 and BMP-6 to C2C12 cells resulted in higher alkaline phosphatase activity than that of only one of these BMPs. The combination of ALK-2 and ALK-3 also induced higher transcriptional activity than either receptor alone. Thus, ALK-2 and ALK-3 (or ALK-6) might synergistically induce osteoblast differentiation of C2C12 cells, possibly through efficient activation of downstream signaling pathways. PMID- 11282025 TI - The Cdk9 and cyclin T subunits of TAK/P-TEFb localize to splicing factor-rich nuclear speckle regions. AB - TAK/P-TEFb is an elongation factor for RNA polymerase II-directed transcription that is thought to function by phosphorylating the C-terminal domain of the largest subunit of RNA polymerase II. TAK/P-TEFb is composed of Cdk9 and cyclin T and serves as the cellular cofactor for the human immunodeficiency virus transactivator Tat protein. In this study, we examined the subcellular distribution of Cdk9 and cyclin T1 using high resolution immunofluorescence microscopy and found that Cdk9 and cyclin T1 localized throughout the non nucleolar nucleoplasm, with increased signal present at numerous foci. Both Cdk9 and cyclin T1 showed only limited colocalization with different phosphorylated forms of RNA polymerase II. However, significant colocalization with antibodies to several splicing factors that identify nuclear 'speckles' was observed for Cdk9 and especially for cyclin T1. The pattern of Cdk9 and cyclin T1 distribution was altered in cells treated with transcription inhibitors. Transient expression of cyclin T1 deletion mutants indicated that a region in the central portion of cyclin T1 is important for accumulation at speckles. Furthermore, cyclin T1 proteins that accumulated at speckles were capable of recruiting Cdk9 and the HIV Tat protein to this compartment in overexpression experiments. These results suggest that cyclin T1 functions to recruit its binding partners to nuclear speckles and raises the possibility that nuclear speckles are a site of TAK/P TEFb function. PMID- 11282026 TI - The life cycle of actin patches in mating yeast. AB - Actin patches are core components of the yeast actin cytoskeleton that undergo redistribution during establishment of cell polarity. Using 4D imaging, we observe the life cycle of actin patches in living yeast for the first time. We observe assembly of actin patches at sites of polarized growth, and disassembly of actin patches concomitant with movement away from those sites. The total lifetime of an actin patch is 10.9+/-4.2 seconds. These findings indicate that actin patches are labile structures, and that the localization of actin patches during establishment of cell polarity occurs by assembly of these structures at sites of polarized cell surface growth. These findings were confirmed and extended by analysis of myosin I proteins and their receptor, verprolin, proteins implicated in actin assembly in yeast. Deletion of type I myosins or their receptor has no effect on the velocity of actin patch movement. However, these mutants show a 65% reduction in number of patch movements and a three-fold increase in patch lifetime. Finally, the actin patch resident proteins Abp1p, fimbrin, and Arp2p show normal association with actin patches in myosin I and verprolin mutants. However, cofilin accumulates in abnormal 'bars' of G-actin in myo3Delta,myo5Delta and vrp1Delta strains, and Las17p/Bee1p is not associated with actin patches in vrp1Delta strains. These findings imply a multi-step process for actin patch assembly. Early events in this process, including assembly of Abp1p, fimbrin and Arp2p with F-actin, can occur throughout the cell and do not require myosin I proteins or their receptor. Later events in this process are myosin I-dependent, and are required for assembly of actin patches at sites of polarized cell surface growth. PMID- 11282027 TI - Pulsed DC electric fields couple to natural NAD(P)H oscillations in HT-1080 fibrosarcoma cells. AB - Previously, we have demonstrated that NAD(P)H levels in neutrophils and macrophages are oscillatory. We have also found that weak ultra low frequency AC or pulsed DC electric fields can resonate with, and increase the amplitude of, NAD(P)H oscillations in these cells. For these cells, increased NAD(P)H amplitudes directly signal changes in behavior in the absence of cytokines or chemotactic factors. Here, we have studied the effect of pulsed DC electric fields on HT-1080 fibrosarcoma cells. As in neutrophils and macrophages, NAD(P)H levels oscillate. We find that weak (approximately 10(-5) V/m), but properly phased DC (pulsed) electric fields, resonate with NAD(P)H oscillations in polarized and migratory, but not spherical, HT-1080 cells. In this instance, electric field resonance signals an increase in HT-1080 pericellular proteolytic activity. Electric field resonance also triggers an immediate increase in the production of reactive oxygen metabolites. Under resonance conditions, we find evidence of DNA damage in HT-1080 cells in as little as 5 minutes. Thus the ability of external electric fields to effect cell function and physiology by acting on NAD(P)H oscillations is not restricted to cells of the hematopoietic lineage, but may be a universal property of many, if not all polarized and migratory eukaryotic cells. PMID- 11282028 TI - Transcription-dependent nucleocytoplasmic distribution of hnRNP A1 protein in early mouse embryos. AB - A unique feature of certain members of the heterogeneous nuclear ribonucleoprotein (hnRNP) family of proteins is that they shuttle continuously between nucleus and cytoplasm and their accumulation in the nucleus is transcription-dependent. An extensively characterised protein of this group is hnRNP A1. To date, most studies addressing the transcription-dependent transport of hnRNP A1 have been performed on cultured cell lines treated with transcription inhibitors. Here we have analysed the nucleocytoplasmic distribution of hnRNP A1 in early mouse embryos, where the haploid pronuclei remain transcriptionally inactive for a period of several hours. Consistent with its small molecular size (36 kDa), the hnRNP A1 protein diffuses passively through the nuclear pores and equilibrates between the nucleus and the cytoplasm of transcriptionally inactive embryos. In contrast, following transcriptional activation the A1 protein becomes accumulated in the nucleus. This accumulation of the A1 protein in the nucleus is blocked by the lectin wheat germ agglutinin (WGA), which binds to nuclear pore proteins and prevents translocation of receptor-cargo complexes through the pores. This indicates that a carrier-mediated transport pathway is required for the concentration of A1 in transcriptionally active nuclei. To further analyse how transcription is coupled to nucleocytoplasmic transport, we transplanted transcriptionally inactive pronuclei into the cytoplasm of transcriptionally active embryos. The results show that the presence of newly synthesised RNAs in the cytoplasm is not sufficient to induce the accumulation of hnRNP A1 in the nucleus. Rather, the appearance of nascent transcripts in the nucleus appears to be the crucial event. Since hnRNP A1 is a shuttling protein, an increase in its steady state nuclear concentration could be the result of either faster nuclear import or slower export to the cytoplasm. We propose that binding of A1 to nascent transcripts retards its export to the cytoplasm and therefore contributes to its concentration in the nucleus. PMID- 11282029 TI - Mechanism for the transcriptional repression by c-Myc on PDGF beta-receptor. AB - c-Myc plays a key role in the cell cycle dependent control of the PDGF beta receptor mRNA. The mouse platelet-derived growth factor (PDGF) beta-receptor promoter contains a CCAAT motif, and NF-Y plays an essential role in its transcription. Coexpression of c-Myc represses PDGF beta-receptor luciferase reporter activity, and the CCAAT motif in the promoter is indispensable for this repression. Here we show that c-Myc binds NF-Y subunits, YB and YC, by immunoprecipitation from cotransfected COS-1 cells. The in vitro-translated c-Myc also binds the glutathione S-transferase (GST)-NF-YB fusion protein and GST-NF YC, but not GST-NF-YA. The most C-terminal region of HAP domains of NF-YB and NF YC, and the Myc homology boxes, but not the C-terminal bHLHZip domain, are indispensable for the coimmunoprecipitation, and also for the repression of PDGF beta-receptor. c-Myc binds NF-Y complex without affecting the efficiency of NF-Y binding to DNA. However, the expression of Myc represses the transcriptional activation of NF-YC when fused to the GAL4 DNA binding domain. Furthermore, this repression was seen only when Myc homology boxes are present, and NF-YC contains the c-Myc binding region. PMID- 11282030 TI - Superactivation of integrin alphavbeta3 by low antagonist concentrations. AB - Integrins are implicated in cell adhesion, migration and homeostasis. An important feature is their ability to adopt different affinity states that can be regulated by a variety of intra- and extracellular factors. To study affinity modulation of the integrin ectodomain by extracellular factors, we produced a soluble recombinant form of mouse integrin alphavbeta3 in a mammalian expression system and isolated it to purity. We show that the two transmembrane truncated integrin subunits stably associate to form a functional receptor, soluble recombinant alphavbeta3. The affinity of this receptor for its ligands vitronectin, fibronectin and fibrinogen can be modulated by the divalent cations magnesium, calcium and manganese. Most importantly, we found that a cyclic RGD peptide has a biphasic effect on rsalphavbeta3and native purified alphavbeta3, with an antagonistic phase at high concentrations, and an agonistic phase at low concentrations. This integrin superactivation by low antagonist concentrations is shown in binding of sralphavbeta3 to immobilized ligands by ELISA, and in adhesion of cells that express the chimaeric integrin ligand KISS31 to immobilized rsalphavbeta3 and native purified alphavbeta3. Our results indicate that low concentrations of the ligand mimetic cyclo-RGD can result in superactivation of the extracellular domain of integrin alphavbeta3 to a comparable level as activation by manganese. PMID- 11282032 TI - N-cadherin is developmentally regulated and functionally involved in early hematopoietic cell differentiation. AB - The cadherins, an important family of cell adhesion molecules, are known to play major roles during embryonic development and in the maintenance of solid tissue architecture. In the hematopoietic system, however, little is known of the role of this cell adhesion family. By RT-PCR, western blot analysis and immunofluorescence staining we show that N-cadherin, a classical type I cadherin mainly expressed on neuronal, endothelial and muscle cells, is expressed on the cell surface of resident bone marrow stromal cells. FACS analysis of bone marrow mononuclear cells revealed that N-cadherin is also expressed on a subpopulation of early hematopoietic progenitor cells. Triple-color FACS analysis defined a new CD34(+) CD19(+) N-cadherin(+) progenitor cell population. During further differentiation, however, N-cadherin expression is lost. Treatment of CD34(+) progenitor cells with function-perturbing N-cadherin antibodies drastically diminished colony formation, indicating a direct involvement of N-cadherin in the differentiation program of early hematopoietic progenitors. N-cadherin can also mediate adhesive interactions within the bone marrow as demonstrated by inhibition of homotypic interactions of bone-marrow-derived cells with N-cadherin antibodies. Together, these data strongly suggest that N-cadherin is involved in the development and retention of early hematopoietic progenitors within the bone marrow microenvironment. PMID- 11282031 TI - N-WASP, WAVE and Mena play different roles in the organization of actin cytoskeleton in lamellipodia. AB - WASP- and Ena/VASP-family proteins have been reported to regulate the cortical actin cytoskeleton as downstream effectors of the Rho-family small G-proteins Rac and Cdc42, but their functions are little understood. We observed the localization of the WASP family proteins, N-WASP and WAVE, and the Ena/VASP family protein, Mena, in protruding lamellipodia. Rat fibroblast cell line 3Y1 protruded lamellipodia on poly-L-lysine-coated substrate without any trophic factor. N-WASP and Cdc42 were concentrated along the actin filament bundles of microspikes but not at the tips. By immunofluorescence and immunoelectron microscopy, both WAVE and Mena were observed to localize at the lamellipodium edge. Interestingly, Mena tended to concentrate at the microspike tips but WAVE did not. At the edge of the lamellipodium, the correlation between the fluorescence from Mena and actin filaments stained with the specific antibody and rhodamine-phalloidin, respectively, was much higher than that between WAVE and actin filament. The Ena/VASP homology 2 (EVH2) domain of avian Ena, an avian homolog of Mena, was localized to the lamellipodium edge and concentrated at the tip of microspikes. The SCAR homology domain (SHD) of human WAVE was distributed along the lamellipodium edge. These results indicate that N-WASP, WAVE and Mena have different roles in the regulation of the cortical actin cytoskeleton in the protruding lamellipodium. WAVE and Mena should be recruited to the lamellipodium edge through SHD and the EVH2 domain, respectively, to regulate the actin polymerization near the cell membrane. N-WASP should regulate the formation of the actin filament bundle in addition to activating Arp2/3 complex in lamellipodium under the control of Cdc42. PMID- 11282033 TI - Phosphatidylinositol 3-kinase mediates integrin-dependent NF-kappaB and MAPK activation through separate signaling pathways. AB - Integrin-mediated signals play an important but poorly understood role in regulating many leukocyte functions. In monocytes and monocytic leukemia cells, beta1 integrin-mediated adhesion results in a strong induction of immediate-early genes that are important in inflammation. To investigate the signaling pathways from integrins in monocytic cells, THP-1 cells were stimulated via beta1 integrins by binding to fibronectin and by crosslinking the integrins with specific monoclonal antibodies. The involvement of MAPK and PI 3-K on nuclear factor kappaB (NF-kappaB) activation was then analyzed. We found that integrins activated both NF-kappaB and MAPK in a PI 3-K-dependent manner, as wortmannin and LY294002 blocked these responses. However, the specific MEK inhibitor PD98059 did not prevent integrin-mediated NF-kappaB activation. In contrast, a dominant negative mutant of Rac completely prevented NF-kappaB activation, but it did not affect MAPK activation. These results indicate that integrin signaling to NF kappaB is not mediated by the MAPK pathway, but rather by the small GTPase Rac. In addition, a dominant negative form of Rho augmented NF-kappaB activation and blocked MAPK activation, implying that these two pathways are in competition with each other. These data suggest that integrins activate different signaling pathways in monocytic cells. One uses PI 3-K and Rac to activate NF-kappaB, while the other uses PI 3-K, MEK, and MAPK to activate other nuclear factors, such as Elk-1. PMID- 11282035 TI - Should the use of smoking cessation products be promoted by dental offices? An evidence-based report. AB - To address the issue of whether dentists should promote the use of smoking cessation products, an evidence-based methodology was applied to find answers to 3 questions: Does tobacco use affect periodontal health? Are dentists effective cessation counsellors? Do smoking cessation products improve the effectiveness of cessation interventions? MEDLINE and manual searches uncovered relevant evidence to use in developing evidence-based recommendations. There is fair evidence that tobacco use is a major factor in the progression and treatment outcome of adult periodontitis and that quitting tobacco use is beneficial to periodontal health. There is good evidence to recommend that oral health professionals provide cessation counselling. There is good evidence to recommend the use of smoking cessation adjuncts. In view of the strong supporting evidence, dental offices should incorporate systematic smoking cessation services into routine patient care and should promote the use of proven cessation products by patients who are attempting to quit. PMID- 11282034 TI - Effects of mutations in potential phosphorylation sites on transcytosis of FcRn. AB - The neonatal Fc receptor, FcRn, transports immunoglobulin G (IgG) across intestinal epithelial cells of suckling rats and mice from the lumenal surface to the serosal surface. In cell culture models FcRn transports IgG bidirectionally, but there are differences in the mechanisms of transport in the two directions. We investigated the effects of mutations in the cytoplasmic domain of FcRn on apical to basolateral and basolateral to apical transport of Fc across rat inner medullary collecting duct (IMCD) cells. Basolateral to apical transport did not depend upon determinants in the cytoplasmic domain. In contrast, an essentially tailless FcRn was markedly impaired in apical to basolateral transport. Using truncation and substitution mutants, we identified serine-313 and serine-319 as phosphorylation sites in the cytoplasmic domain of FcRn expressed in Rat1 fibroblasts. Mutations at Ser-319 did not affect transcytosis across IMCD cells. FcRn-S313A was impaired in apical to basolateral transcytosis to the same extent as tailless FcRn, whereas FcRn-S313D transported at wild-type levels. FcRn-S313A recycled more Fc to the apical medium than the wild-type receptor, suggesting that Ser-313 is required to allow FcRn to be diverted from an apical recycling pathway to a transcytotic pathway. PMID- 11282036 TI - Bladder, Bowel, and Sexual Dysfunction in Multiple Sclerosis. AB - The majority of patients with multiple sclerosis (MS) experience genitourinary and bowel dysfunction over the course of their illness. Lower extremity pyramidal signs are excellent predictors of concurrent bladder dysfunction. Constipation is the most common bowel dysfunction, which results from a range of causes including pelvic floor spasticity, decreased gastro-colic reflex, inadequate hydration, medications, immobility, poor physical conditioning, and weak abdominal muscles. Despite the advent of new therapeutic modalities, the physician and patient commonly overlook sexual dysfunction. A detailed history of the patient is crucial to determine the cause of the dysfunction. Fatigue, pain, mood disorders, spasticity, bowel, and bladder dysfunction can all interfere with normal sexual functioning, and these subjects should be explored in detail in order to plan for proper treatment. Integrated treatment plans, often in conjunction with an urologist, can lead to amelioration of symptoms. PMID- 11282037 TI - Neck and Back Pain in the Elderly. AB - Surgical intervention for neck and back pain in elderly patients without significant comorbidities can significantly improve a patient's symptoms and quality of life when more conservative therapies fail. Current spine literature strongly supports the paradigm of treating elderly patients with stable, chronic neck or back pain with conservative therapies first in order to optimize the risks and benefits of all available treatment options. If less-invasive methods fail to achieve satisfactory outcomes, more aggressive surgical options can, at that time, typically be implemented with excellent results in elderly patients without significant comorbidities. Clinical scenarios threatening to result in spine instability or nerve root or spinal cord compression require immediate intervention, especially in elderly patients, who, in general, have a higher risk of developing such conditions either through falls or trauma or acquired degenerative disease processes or malignancies. When an elderly patient enters a physician's office and asks "doctor, I've had pain for years, but it's getting worse. At my age, is it really worth having surgery?" The answer is a qualified "yes," if conservative treatments have failed and if the patient is otherwise in reasonably good health. Because the vast majority of these patients first interact with the medical system through their primary care doctors and neurologists, early recognition of situations requiring immediate attention, and those requiring referrals to spine specialists, can greatly expedite the appropriate use of scarce healthcare resources. Furthermore, knowledge of the various treatment options available to elderly patients complaining of the very common symptoms of neck or back pain can significantly improve patient care, especially in this new century when older patients will increasingly become a larger and more active force in all aspects of our society. PMID- 11282038 TI - Attention Deficit Hyperactivity Disorder. AB - Management of attention deficit hyperactivity disorder (ADHD) encompasses two general domains: pharmacologic therapies and nonpharmacologic therapies, including educational, cognitive-behavioral, and other psychological and psychiatric approaches. Within the past year there have been two seminal developments in treatment. The first is that the Evidence-based Practice Center at McMaster University, under contract with the Agency for Health Care Policy and Research, produced an evidence based report on the treatment of ADHD. The topic was proposed to the AHCPR by the American Academy of Pediatrics and American Psychiatric Association, who served as partners to the center. The second is the completion of the Multisite Multimodal Treatment Study of Children with Attention Deficit/Hyperactivity Disorder (MTA) study by the National Institutes of Mental Health, a 14-month double-blind placebo trial of medication and behavioral therapy in ADHD. In general, the result of the evidence-based review and the MTA study is that stimulants are the most effective agents for the treatment of ADHD. Results from the MTA study indicate that methylphenidate (MPH) and MPH combined with behavioral therapy are superior to behavioral therapy alone and that all three are superior to community therapy. The evidence-based review indicates that each of the stimulants is superior to placebo and the stimulants (regular and sustained-release MPH as well as d and l isomers of the stimulants) are comparable. As for other agents, tricyclic antidepressants, specifically desipramine, are superior to placebo. Only a few studies compared stimulants directly with tricyclic antidepressants, and these were technically inadequate, leading to the conclusion that more rigorous studies are required. Only five studies were found that examined nonpharmacologic treatment, and all contained major limitations in methodology. Despite the limitations, all showed that stimulants were more effective than the nonpharmacologic therapies, consonant with the results of the MTA study. There was lack of evidence to support the superiority of combination multimodal treatment over stimulant therapy alone, again consonant with the MTA study. Both the evidence-based review and the MTA study examined ADHD in middle childhood. Finally, most studies are relatively short-term, including the MTA study (at 14 months). Some evidence suggests that MPH reduces behavioral disturbance as long as it is taken. PMID- 11282039 TI - Neonatal Seizures. AB - Neonatal seizures are frequently manifested by subtle movements that are referable to brain stem structure, ie, nystagmus, conjugate eye movements, posturing, sucking movements, and so forth. Electroencephalogram (EEG) confirmation of abnormal movements is essential in diagnosing seizures in the neonate. Clinical seizure signs are often a clue to etiology. Metabolic abnormalities must always be considered, and blood gases, calcium, magnesium, glucose, and ammonia obtained. Neonatal seizures are an indication for cerebrospinal fluid examination. Specific metabolic abnormalities are treated with metabolic approaches, not conventional anticonvulsant drugs. Hypertensive encephalopathy is treated by controlling blood pressure, and not through anticonvulsant drugs. Critically ill infants bind anticonvulsants in an unpredictable fashion, and unbound or free anticonvulsant drug concentrations should be used to guide therapy. Phenobarbital is the most commonly used drug in treating nonmetabolic seizures. Doses to achieve free concentrations of at least 35 mg/L should be used. Use in vitro binding determinations with this formula to calculate loading doses. Dose is 25 mg/kg multiplied by volume and distribution (1 L/kg) divided by % free. Phenytoin is the second most commonly used agent, and doses should be calculated to achieve, but not exceed, 3 mg/L. Dose is 3 mg/kg multiplied by volume and distribution (1 L/kg) divided by % free. Benzodiazepines, notably lorazepam and diazepam are used at doses of 0.15 mg/kg and 0.3 mg/kg, respectively. PMID- 11282040 TI - Lysosomal Storage Diseases. AB - Lysosomal storage disorders (LSDs), over 40 different diseases, are now considered treatable disorders. Only a few short years ago, Lysosomal storage disorders were seen as interesting neurodegenerative disorders without any potential for treatment. Effective treatment strategies such as bone marrow transplantation (BMT), enzyme replacement therapy (ERT), and glycolipid synthesis inhibition have been developed in the last 20 years and continue to be researched and evaluated. Bone marrow transplantation began approximately 15 years ago and has shown benefit for some of the lysosomal storage disorders. In order to be effective, the transplant must be performed early in the course of the disease, before the development of irreversible neurologic damage. Diseases such as Hurler appear to respond to BMT, however, improvement in bone disease is much less vigorous than responses in other organs. Krabbe disease responds if the transplant is performed before irreversible signs of neurologic damage appear. Metachromatic leukodystrophy may respond if the transplant can be performed early enough although peripheral nerve findings appear to progress. Other diseases, eg, GM1- and GM2-gangliosidoses do not appear to be altered by BMT. Despite its high cost, ERT has been very effective treatment for type I (non-neuronopathic) Gaucher disease. Enzyme replacement therapy for other LSDs, including ERT for Fabry and Pompe diseases, which are planned to be imminently introduced, and other enzymes such as for Morquio and Hunter diseases that are in the study phases, may be marketed in the very near future. Glycolipid inhibitors, such as N butyldeoxynijirimycin (OGS-918), have been effective in reducing the liver and spleen volume in type I Gaucher disease. These oral inhibitors may prove to be important adjuncts to ERT and provide the advantage of being able to cross the blood/brain barrier, which limits enzyme access to brain. Currently, clinical studies are being conducted on patients with type III Gaucher disease and Fabry disease using OGS-918. Other, potentially more specific, glycolipid inhibitors are being developed. PMID- 11282041 TI - Childhood Migraine Headache Syndromes. AB - The treatment of migraine headache in children depends on the following: a) defining the underlying cause; b) the frequency of the attacks; and c) the severity of the disability produced by the pain. Any medication taken to relieve pain is most effective if taken at maximum dose at the onset of the headache. The dose should be the maximum recommended by weight or age. Triptans are also more effective if used early. Over-the-counter (OTC) analgesics are often effective in relieving pediatric headache and should be tried before prescription drug therapy is attempted. The more frequent a child's headaches are, the greater the danger that repeated doses of pain medications, including those purchased OTC, will lead to a chronic headache syndrome as the medication is reduced. Recurrent severe headaches, occurring more than once a week and resulting in interruption of normal activities or poor concentration, need to be treated with prophylactic medications taken daily so that the number of headaches can be reduced. Amitriptyline, propanolol, and periactin are the most frequently used drugs to block headaches, but valproate, verapamil, or other calcium channel blockers and other antidepressants are also useful. Biofeedback, relaxation, or cognitive therapies can also reduce headache frequency in children with both migraine and tension headaches. Headaches that are intractable to oral medication for the acute relief of pain may respond more rapidly to an efficiently absorbed drug administered by nasal spray or subcutaneously. The initial dose of an injectable drug should be given in a situation where a physician is immediately available. Recurrent headaches that have occurred over more than 6 months and that are associated with a normal neurologic examination are almost never caused by an intracranial lesion. Routine CT and MRI scans or an electroencephalogram (EEG) are generally unnecessary for these patients because these scans are rarely of value in these patients unless there is a history of another neurologic disorder or the headaches are focal, relentless, and worsening over time. PMID- 11282042 TI - Mitochondrial Disease. AB - Mitochondrial diseases are disorders of energy metabolism that include defects of pyruvate metabolism, Krebs cycle, respiratory chain (RC), and fatty acid oxidation (FAO). Treatment of pyruvate metabolism, Krebs cycle, and RC disorders is, in general, disappointing. Therapeutic approaches consist of electron acceptors, enzyme activators, vitamins, coenzymes, free-radical scavengers, dietary measures, and supportive therapy. These treatment assumptions are based on current understanding of the pathophysiology, on anecdotal clinical reports, and on a few controlled clinical trials, which have not been encouraging. Although it is difficult to perform clinical trials in these conditions due to their rarity and genotypic and phenotypic heterogeneity, there is a great need for well-performed double-blind placebo- controlled clinical trials with comparable groups of patients and with sufficient follow-up periods. Treatment options for FAO disorders are, in general, satisfactory and are mainly based on diet, lifestyle recommendations, and administration of L-carnitine and, in some cases, riboflavin. Special conditions that involve primary deficiencies of L carnitine, coenzyme Q(10), and cofactor- and vitamin-responsive enzyme defects must be systematically considered, because supplementation with these substances may be curative or produce dramatic improvements. While awaiting more specific therapies for mitochondrial disorders, it is useful to reach a consensus regarding the management of these patients. The expected outcome is a slowing of the disease process and stabilization of the clinical syndrome. More definitive treatments hopefully will follow in the near future. PMID- 11282043 TI - Infantile Spasms. AB - Infantile spasm is a catastrophic form of epilepsy found only in infants and young toddlers. Onset is before one year of age, with a peak incidence occurring between 4 to 7 months of age. The prevalence is difficult to calculate, but previous reviews have estimated between 1 per 2000 to 6000 live births. There are many causes of infantile spasms, including tuberous sclerosis, malformations of cortical development, hypoxic-ischemic injury, congenital infectious diseases, inborn errors of metabolism, genetic syndromes such as Aicardi's syndrome, and chromosomal abnormalities. A small percentage of patients have idiopathic infantile spasms, with no identifiable cause and premorbid normal growth and development. In order to prevent an ongoing epileptic encephalopathy with its concomitant consequences of cognitive impairment and intractable seizures, treatment should be aggressive and immediate. It is not enough to control the clinical infantile spasms. The underlying "interictal" hypsarrhythmia pattern must also be abolished if the prognosis is to be improved. Otherwise, the immature brain appears to remain hyperexcitable. PMID- 11282044 TI - [Selective use of the diagnostic tests in syncope of unknown origin. Usefulness of implantable Holter]. PMID- 11282045 TI - [Management of acute myocardial infarction in Spain. Current inter-regional differences according to IBERICA Registry]. PMID- 11282046 TI - [Rotational atherectomy: to be or not to be in interventional cardiology]. PMID- 11282047 TI - [Diagnostic accuracy of a protocol in the evaluation of unexplained syncope]. AB - INTRODUCTION AND OBJECTIVES: To assess the diagnostic capacity of a protocol to study syncope of unknown cause in which electrophysiological studies and tilting table tests are selectively used. PATIENTS AND METHODS: The study was performed in 137 consecutive patients (94 men and 43 women, with a mean age of 57.6+/-18.3 years) with syncope of unknown cause after the initial clinical evaluation, who were divided into two groups. Group A consisted of 77 patients meeting any of the following criteria: a) presence of structural heart disease; b) abnormal ECG; c) presence of significant non-symptomatic arrhythmia in the Holter recording, and d) presence of paroxysmal palpitations. These patients initially underwent an electrophysiological study. Group B consisted of 60 patients not meeting any of the above criteria, who were initially submitted to tilting table tests.Results. In group A, the electrophysiological study was positive in 43 patients (55%). In group B, the tilting test was positive in 41 patients (68%). Among patients in group A with a negative study, 20 (59%) were submitted to the tilting table test, with positive results in 7 cases (35%). Five patients from group B with a negative tilting test underwent the electrophysiological study, which was negative in all of them. Overall, a positive diagnosis was achieved in 91 of 137 patients (66%). CONCLUSIONS: In patients with syncope of a non-apparent cause in the initial assessment, selective use of electrophysiological studies or tilting table tests, guided by clinical criteria, allows for a positive diagnosis in over 60% of the cases. Our results suggest that the tilting table test should be performed in cases of group A with a negative electrophysiological study. PMID- 11282048 TI - [Usefulness of the implantable subcutaneous recorder in the diagnosis of recurrent syncope of unknown etiology in patients without structural heart disease and negative tilt test and electrophysiological study]. AB - BACKGROUND AND OBJECTIVES: In up to 38% of the cases, the etiology of syncope difficult to determine. The main obstacle for diagnosis of the causes of syncope lies in the unpredictable frequency of episodes. Development of implantable loop recorders allows long term electrocardiographic monitoring. The aim of this study was to evaluate the usefulness of the implantable loop recorder for the diagnosis of recurrent syncope of unknown origin. PATIENTS AND METHODS: From May 1991 to April 1999, a cohort of 176 patients with recurrent syncope was prospectively assessed. Investigations, including Holter monitoring, Tilt Test and electrophysiological study, allowed the determination of the etiology in 161 patients. The remaining 15 patients, without structural cardiac disease were selected for continuous electrocardiographic monitoring using an implantable loop recorder. RESULTS: During follow up after implant, 15 +/- 2 months (X- +/- SEM), 9 patients showed recurrence of symptoms concordant with prior episodes (time: 105 +/- 30 days). In 7 cases records during symptoms were diagnostic (0.47; CI 95%: 0.21-0.73), in 3 cases a diagnosis with documented arrhythmia was achieved, and in 4 other cases a presumptive clinical diagnosis of non-arrhythmic cause was made. In 8 patients, 6 with no recurrences, diagnosis was not possible. There were no complications related to the use of the device. CONCLUSIONS: The strategy of long term monitoring with the implantable loop recorder is safe and effective in patients with recurrent syncope of unknown etiology. PMID- 11282049 TI - [Variability in the in-hospital management of acute myocardial infarction in Spain. IBERICA Study (Investigacion, Busqueda Especifica y Registro de Isquemia Coronaria Aguda)]. AB - Introduction and objective. Although some in-hospital studies have described the management of acute myocardial infarction (MI) patients in Spain, none has been able to guarantee the exhaustiveness of patient registry. This study sought to determine the clinical characteristics and in-hospital management of patients with MI in eight Spanish population registries.Methods. The IBERICA study is a population-based MI registry carried out in the 25 to 74 year-old population, in eight Spanish regions in 1997. A standardized methodology was used to register and investigate all MI arriving alive to a hospital. Clinical characteristics, cardiovascular risk factors prevalence, pharmacological treatment, invasive and non-invasive procedures performed and complications at 28 days of evolution were recorded. A descriptive analysis was performed and the variation coefficient (VC) was calculated.Results. In 1997, 4,041 MI patients were registered, 79.9% were men with a mean age of 61.1 years. Although 10.9% (95% CI: 9.9-11.9%) were not admitted to the coronary care unit, a large variability existed among different areas (VC = 53%). There was a high variability in the utilization and performance of non-invasive and invasive procedures among regions, as well as in the use of pharmacological treatment. Only the use of antiaggregants (91.5%) and thrombolytic therapy (41.8%) showed a low variability (VC < 10%). Twenty-eight day mortality was 16.2% (95% CI: 15.1-17.4%) with a high variability being observed among the different regions (VC = 20.6%).Conclusion. Patient characteristics vary among the different Spanish regions. The differences in management and prognosis suggest a lack of equality in the health care provided to MI patients in the different regions in Spain. PMID- 11282050 TI - [Inflammation and infection in stable coronary disease and acute coronary syndrome]. AB - OBJECTIVE: To study whether inflammation and infection are related to coronary artery disease. DESIGN: Sixty patients (44 males, mean age 62 +/- 13 years) with acute coronary syndrome and 40 with stable coronary artery disease (31 males, age 64 +/- 10 years) and a control group of 40 individuals (34 males, 53 +/- 5 years) were analyzed. IgG against Chlamydia pneumoniae, Cytomegalovirus and Helicobacter pylori plus C-reactive protein were assessed in all serum samples. In addition, IgM against C. pneumoniae and Cytomegalovirus on admission and C-reactive protein one month later were measured in acute patients. RESULTS: No IgM seropositivity was observed. A high prevalence of IgG seropositivity with no significant differences among the groups was found: C. pneumoniae: acute group 44 (73%), stable group 29 (73%) and control group 25 (63%); Cytomegalovirus: 55 (92%), 37 (92%) and 38 (95%), respectively; and H. pylori, 43 (72%), 32 (80%) and 34 (85%) respectively. There was a high rate of positive C-reactive protein in the acute group: 48 (80%) vs 10 (25%) the stable group and 0% the control group (p < 0.001). C-reactive protein levels were higher in Q-wave infarction than in unstable angina/ non-Q-wave infarction (median 22.65 vs 7.69, p < 0.001). One month later, C-reactive protein levels decreased (median 22.65 vs 3.38, p < 0.001), but were still positive in 40%. CONCLUSIONS: These data suggest that inflammation is detected by the commonly used methods in clinic practice in acute coronary syndromes and to a lesser extent in stable coronary artery disease. It seems that different mechanisms other than infection account for this inflammatory response, at least this being so when infection is assessed by serology. Serology does not appear to be an adequate method to determine the possible relationship among coronary syndromes, infection and inflammation. PMID- 11282051 TI - [In-hospital major complications associated with rotational atherectomy: experience with 800 patients at a single center]. AB - INTRODUCTION: Rotational atherectomy is usually performed in patients with angiographically determined high risk coronary lesions. The aim of this study was to evaluate the rate of major adverse cardiac events (death, Q-wave infarction or new revascularization) after rotational atherectomy, as well as to identify the clinical characteristics associated with this incidence. PATIENTS AND METHODS: The study population included 800 patients treated with rotational atherectomy from 1993 to 1999: 512 (64%), for de novo lesions, and 288 (36%) for restenosis. Balloon dilation and coronary stenting was performed in 95% and 34% of patients, respectively. RESULTS: During hospitalization, 17 patients (2.1%) died, 16 (2%) had a Q-wave infarction, 30 (3.8%) a non-Q infarction, and new revascularization was performed in 28 (3.5%). The incidence of major adverse cardiac events was 6.5% (n = 52), this incidence being higher in the presence of diabetes (8.9 vs. 4.4%; p = 0.01), unstable angina or acute/recent myocardial infarction (7.6 vs. 3.3%; p = 0.02), multivessel disease (8.6 vs. 3.3%; p < 0.01), treated vessel other than right coronary (7.0 vs. 1.7%; p = 0.01), procedure in > 1 vessel (10.7 vs. 4.7%; p < 0.01), angiographic failure (62.5 vs. 5.5%; p < 0.001), and de novo lesions (8.4 vs. 2.5%; p < 0.01), with diabetes and treatment of de novo lesions being independent predictors of major adverse cardiac events. However, age, previous infarction, and left ventricular dysfunction, were not associated with the rate of events. CONCLUSION: Some simple variables are associated with a higher incidence of major adverse cardiac events after rotational atherectomy. Advanced age, previous infarction and left ventricular dysfunction, however, do not necessarily imply a poorer prognosis in these patients. PMID- 11282052 TI - [The Influence of the growth hormone in the profile of blood pressure. Results in adult patients with deficiency in this hormone]. AB - INTRODUCTION AND OBJECTIVES: There is an increasing interest in the relationship between the growth hormone (GH) and the heart since the GH has an important inotropic effect and its use has been tested in patients with severe systolic dysfunction. However, cardiovascular diseases are the main cause of increased morbimortality observed in patients with acromegaly. Growth hormone deficiency has been related to different clinical findings depending on the age of onset. Recent studies have demonstrated that GH deficiency in adults is associated with alterations in blood pressure. The aim of our study was to assess the influence of GH in blood pressure. PATIENTS AND METHODS: We studied 14 adult patients with GH deficiency and 15 healthy subjects, matched for sex and age. The diagnosis of GH deficiency was based on GH response to intravenous insulin tolerance test < 5 ng/ml and IGF-1 levels lower than the normal limit for each age group. In all the patients 24-hour Holter blood pressure monitorization was performed in addition to a treadmill test and echographic evaluation. RESULTS: All patients showed normal systolic and diastolic function in the echocardiographic study. Only one patient had an increased left ventricular mass. Blood pressure was lower in the patients than in the control subjects (p < 0.05). Moreover, the difference remained significant when analysis was based on the time of day. However, the patients showed normal blood pressure response to the effort test with a mean increase of 60%. The length of the exercise on the treadmill test was shorter in the subgroup of GH deficient patients. CONCLUSIONS: Lower systolic blood pressure was observed in GH deficiency patients. The patients studied did not show structural heart alterations. Blood pressure and chronotrophic response to the effort test were similar in both groups. PMID- 11282053 TI - [Guidelines of the Spanish Society of Cardiology for car driving, airplane flying,and underwater activities in subjects with heart disease]. AB - Car driving, airplane piloting and underwater activities by subjects with heart disease may cause sudden incapacitation leading to the loss of the safety margins necessary to avoid accidents. In the case of car driving and airplane piloting the risk affects, not only the driver or pilot, but also passengers and/or bystanders within an accident zone. In the case of diving the risk resides basically in the loss of control of the vital support mechanisms necessary in a very hostile medium. This document reviews the possible causes of unexpected incapacitation, with or without loss of consciousness, in the light of the pathophysiologic consequences of fatigue, hypoxia, stress or barotrauma posed by each activity. Detailed recommendations are made for limiting driving, piloting and diving, based on official Spanish and European regulations and the addresses of specialized centers are provided for consultation. Moreover, recommendations for airplane travel for patients with heart disease are indicated. PMID- 11282054 TI - [Hormone replacement therapy in ischemic heart disease prevention in women. Arguments in favor]. AB - Numerous evidence have suggested a physiologic action of sexual steroids upon the cardiovascular system and the coherence of epidemiological studies have raised the possibility of a positive action of estradiol in preventing cardiovascular disease, specially through atheroma inhibition and other vascular wall-related mechanisms. From an experimental point of view, some clinical trials have demonstrated an improvement in some intermediate clinical variables, such as hypercholesterolemia and hypertension, after the administration of estradiol. Nonetheless, the HERS study, the first secondary prevention trial of estrogen and cardiovascular disease, failed to demonstrate these positive actions suggested by epidemiological studies and the efficacy of estradiol in the treatment of postmenopausal women with cardiovascular disease has been questioned. In spite of this, the HERS study has also been questioned because of different pitfalls in its development and, for some authors, it is inconclusive. Therefore, at present, it is not possible to make an evidence based clinical decision regarding the key question about the real actions of estradiol in the prevention of cardiovascular disease in postmenopausal women. PMID- 11282055 TI - [Hormone replacement therapy in ischemic heart disease prevention in women. Arguments against]. AB - Hormone replacement therapy is one of the most difficult issues women and their doctors face. Epidemiological studies have consistently found that women using hormone replacement therapy are at a substantially lower risk of developing coronary heart disease. Observational data are supported by findings demonstrating that hormone replacement therapy improves several risk factors o coronary heart disease, specially the favourable changes in lipid profile. However, no study has clearly established hormones help prevent heart disease. In women without heart disease, the benefits of hormone replacement therapy are unclear. However, recent clinical trials have sown that the use of hormone replacement therapy does not provide cardiovascular benefits in women with established heart disease. PMID- 11282056 TI - [Genetics and molecular biology in cardiology (IV). Myocardial response to biomechanical stress]. AB - Biomechanical stress of the myocardium is the situation resulting from hypoxia, hypertension, and other forms of myocardial injury, that invariably lead to increased demands for cardiac work and/or loss of functional myocardium. As a consequence of biomechanical stress a number of responses develop involving all the myocardial cells, namely cardiomyocytes. As a result some myocardial phenotypic changes develop that are initially compensatory (i.e., hypertrophy) but which may mediate the eventual decline in myocardial function that occurs with the transition from hypertrophy to failure in conditions of persistent stress (i.e., apoptosis and fibrosis). This review focuses on the steps involved in the response of the myocardium to biomechanical stress and highlights the most recent developments in the molecular mechanisms involved in the development of heart failure. PMID- 11282057 TI - [Quadruple coronary bypass "Y" graft mammary arteries]. PMID- 11282058 TI - [Apical hypertrophic cardiomyopathy with mid-ventricular obstruction and apical necrosis]. PMID- 11282059 TI - [Wolff-Parkinson-White syndrome cured with diagnostic electrode catheter]. AB - Catheter-induced mechanical trauma to accessory pathways is an infrequent, usually transitory phenomenon related to manipulation of the ablation catheter. We describe a patient with Wolff-Parkinson-White syndrome due to a midseptal accessory pathway with recurrent episodes of paroxysmal tachycardias under antiarrhythmic treatment. During the diagnostic electrophysiological study coinciding with manipulation of the His catheter (5 French), mechanical trauma to the accessory pathways occurred, with preexcitation not being recovered in the midterm follow up (28 months). This is a exceptional case demonstrating the complexity of the decision making (expectant or rescue ablation) following accidental catheter-induced mechanical trauma. PMID- 11282060 TI - [Patent foramen oval as a cause of severe hypoxia during the immediate postoperative period following heart transplantation]. AB - We report the case of a 44-year-old man with dilated cardiomyocardiopathy, pulmonary hypertension and high pulmonary resistance who underwent orthotopic heart transplant. Following transplantation severe hypoxia was observed after weaning of cardiopulmonary bypass secondary to a patent foramen oval in the donor heart. This situation does not usually lead to problems in the donor, however taking into account the special hemodynamic conditions of the transplanted patient, with high pressure in the right side, an opening occurred producing a significant right-left shunt causing severe receptor hypoxia. Successful closure of the patent foramen oval was performed. PMID- 11282061 TI - [Ventricular fibrillation secondary to radiofrequency application in ventricular tachycardia ablation]. AB - Radiofrequency catheter ablation has become a first line therapy for several types of tachycardias because of its high efficacy and low complication rate. The development of proarrhythmic complications due to a direct effect of radiofrequency is very unusual. We describe a patient with previous myocardial infarction and well tolerated sustained monomorphic ventricular tachycardia who underwent catheter ablation of the tachycardia substrate. During two of the radiofrequency applications, ventricular fibrillation developed and external defibrillation was required. PMID- 11282063 TI - [Angioedema due to irbesartan]. PMID- 11282062 TI - [Isolated ventricular diverticulum in asymptomatic male]. AB - Ventricular diverticulum are small outpouchings, in the cardiac wall, which are mostly described as a part of malformation syndromes. This finding is infrequent in asymptomatic patients with no pathology in the thoraco abdominal line. The case we present shows a diverticulum in the cardiac apex in a male patient with no cardiological clinic manifestations and with an abnormal electrocardiogram. At present, magnetic resonance is the best diagnostic test, for this kind of malformation, and is also the most reliable in the follow-up of these patients. PMID- 11282064 TI - [Acute coronary syndrome and anaphylactic reaction]. PMID- 11282065 TI - [ECG and right ventricular dysplasia]. PMID- 11282071 TI - Impact factor of Cardiovascular Research in 2000: all time high! PMID- 11282072 TI - On the trail of cardiac specific transcription factors. PMID- 11282073 TI - Signalling in cardiac disease: the molecular deficit at the heart of the problem. PMID- 11282074 TI - Dispersion of ventricular repolarization and refractory period. PMID- 11282075 TI - Identification of novel, cardiac-restricted transcription factors binding to a CACC-box within the human cardiac troponin I promoter. AB - OBJECTIVES: The expression of the human cardiac troponin I (hTnIc) gene is developmentally regulated and tissue-specific. In analysing the putative binding elements within the proximal promoter, a CACC-box sequence overlapping a consensus Sp1 element has been identified. The aim of this study was to characterise the factors binding to this element and to determine their importance in the transcriptional activity of the promoter. METHODS: A combination of supershift and competition electrophoretic mobility shift assays (EMSA) were used to identify the binding of factors to the overlapping CACC box/Sp1 consensus element. The functional importance of this element was tested by transient transfection into primary neonatal rat cardiac myocytes in culture. RESULTS: At least four factors were able to interact with this region including the zinc finger proteins Sp1, Sp3 and two potentially novel factors. Whereas both Sp1 and Sp3 bound to the consensus Sp1 element, and to a lesser extent the CACC box, two of the complexes required the intact CACC-box for binding. Site-directed mutagenesis of this region showed that the CACC-box is essential for hTnIc promoter-reporter activity. Further characterisation using EMSA indicated that the factors binding the hTnIc CACC-box are unlikely to be zinc finger proteins as they are insensitive to the addition of divalent cation chelating agents. They were also unable to bind to other known CACC-box elements. These factors are present in both human and rat cardiac muscle but absent from a number of cell lines including several derived from skeletal muscle. CONCLUSION: The human cardiac troponin I gene promoter requires an upstream CACC-box element for full activity. This element binds at least two complexes which represent novel, tissue restricted DNA-binding activity present in the heart which we have named HCB1 and HCB2 for heart CACC-box binding factors. PMID- 11282076 TI - Characterization of cardiac myocyte and tissue beta-adrenergic signal transduction in rats with heart failure. AB - OBJECTIVE: The cellular basis of alterations in beta-adrenergic signal transduction in rats with chronic heart failure (CHF) remains unclear. The aim of the present study was to examine this signal transduction system in isolated ventricular cardiomyocytes of rats with CHF. We focused on changes in the levels of stimulatory (Gs) and inhibitory G-proteins (Gi). METHODS: CHF was induced in male Wistar rats by coronary artery ligation (CAL). Hemodynamic and biochemical parameters were measured 8 weeks after CAL. Alterations in contractile function and Ca(2+) transients via beta-adrenergic receptor signaling of cardiomyocytes isolated from rats with CHF were characterized by simultaneous measurements of cell shortening and fura-2 fluorescence intensity. RESULTS: Coronary artery ligated rats showed symptoms of CHF, such as decreased contractile function, increased left ventricular volume, decreased chamber stiffness, and about 40% infarct formation of the left ventricle, by 8 weeks after surgery. The contractile function and Ca(2+) dynamics of cardiomyocytes from the rats with CHF remained normal under basal conditions. Only cardiac cell length was increased. The responses of peak shortening, fura-2 fluorescence ratio amplitude, and cAMP content to beta-adrenoceptor stimulation were reduced in cardiomyocytes of the rats with CHF, whereas direct stimulation of adenylate cyclase did not affect the response of these variables. Cardiomyocyte Gsalpha protein was decreased, whereas no changes in Gialpha proteins were seen in these cells. Increases in tissue Gsalpha and Gialpha proteins in the scar zone were detected. The results on tissue levels of collagen and G-proteins in the viable left ventricle appeared to depend on the presence of nonmyocytes. CONCLUSIONS: The results suggest that impaired contractile function of cardiomyocytes is unlikely to account for global LV contractile dysfunction, and that down-regulation of beta-adrenoceptors occurs in cardiomyocytes per se. The difference in changes of G-protein between the cardiomyocyte and myocardial tissue suggests an appreciable contribution of nonmyocytes to myocardial G-protein levels. PMID- 11282077 TI - Altered expression of Bag-1 in Coxsackievirus B3 infected mouse heart. AB - OBJECTIVE: The mechanisms by which Coxsackie B viruses cause myocarditis or dilated cardiomyopathy are not well understood. This study examined changes in the expression of cardiac genes resulting from Coxsackievirus B3 (CVB3) infection of mice. METHODS: Mice (five per group) were experimentally infected with CVB3 or mock-infected with diluent. Altered expression of genes was initially identified by cDNA array, and confirmed by semiquantitative RT-PCR, western blot and immunohistochemistry. RESULTS: Forty-two up-regulated or down-regulated genes were observed in cDNA arrays carrying 588 known mouse genes. Among these, one down-regulated gene, Bag-1, known to be involved in inhibition of apoptosis and modulation of chaperone activity, was investigated further. Semiquantitative RT PCR showed that Bag-1 expression was down-regulated by up to 30% in virus infected mouse heart on day 7 compared to the mock-infected. Cell fractionation and western blot analysis confirmed that Bag-1 isoform p32 was predominant in the cytoplasm of mouse myocardium and down-regulated at 4 days or 7 days after CVB3 infection. In contrast, Bag-1 isoform p50 appeared to increase in the nuclear fraction of mouse heart at 7 days after infection. Down regulated expression and distribution of Bag-1 protein or evidence of apoptosis in the infected mouse heart was demonstrated by immunostaining or histochemistry (TUNEL assay), respectively. CONCLUSION: CVB3 infection induced differential expression of Bag-1 in cytoplasmic and nuclear fractions of mouse heart and apoptosis. This may be important in the pathogenesis of enterovirus heart muscle disease. PMID- 11282078 TI - A3 adenosine receptor stimulation modulates sarcoplasmic reticulum Ca(2+) release in rat heart. AB - OBJECTIVE: Stimulation of A3 adenosine receptors has been shown to protect cardiac myocytes from ischemic injury, but the mechanism of this action is unknown. We evaluated the effect of adenosine agonists and antagonists on the sarcoplasmic reticulum (SR) Ca(2+) channels. METHODS: Isolated rat hearts were perfused with control buffer or different adenosine agonists and antagonists. Hearts were then homogenized and used to determine SR Ca(2+)-induced Ca(2+) release, assayed by quick filtration technique after loading with 45Ca(2+), and the binding of [3H]ryanodine, a specific ligand of the SR Ca(2+) release channel. In parallel experiments, hearts were challenged with 30 min of global ischemia and 120 min of reperfusion, and the extent of tissue necrosis was evaluated by triphenyltetrazolium chloride staining. RESULTS: Perfusion with the A1>A3 agonist R-PIA and the A3>A1 agonist IB-MECA was associated with reduced [3H]ryanodine binding, due to reduced B(max) (by about 20%), whereas K(d) and Ca(2+)-dependence of the binding reaction were unaffected. These actions were abolished by the A3 antagonist MRS 1191, while they were not affected by A1 and A2 antagonists. The rate constant of SR Ca(2+) release decreased by 25-30% in hearts perfused with R PIA or IB-MECA. Tissue necrosis was significantly reduced in the presence of R PIA or IB-MECA. Protection was removed by MRS 1191, and it was not affected by A1 and A2 antagonists. Hearts were also protected by administration of dantrolene, a ryanodine receptor antagonist. In the presence of dantrolene, no further protection was provided by IB-MECA. CONCLUSION: A3 adenosine receptor stimulation modulates the SR Ca(2+) channel. This action might account for the protective effect of adenosine. PMID- 11282080 TI - The class III antiarrhythmic drugs dofetilide and sotalol prevent AF induction by atrial premature complexes at doses that fail to terminate AF. AB - BACKGROUND: Clinical trials suggest that sotalol and dofetilide are much more effective in preventing atrial fibrillation (AF) than in terminating it. This study evaluated potential mechanisms of discordant sotalol and dofetilide effects on AF termination vs. prevention. METHODS: We applied 240-electrode epicardial mapping and programmed stimulation in a vagotonic dog model of AF before and after dofetilide or sotalol. RESULTS: Under control conditions, sustained AF could be induced by single S(2) extrastimuli that caused unidirectional block and macroreentry. Sotalol (2 mg/kg) and dofetilide (0.04 mg/kg) failed to terminate AF in any dog, but prevented AF induction by S(2) stimuli in 19/22 (86%) and 4/5 (80%) of animals, respectively. With sotalol and dofetilide, unidirectional block still occurred, but wavefront reentry failed. The prevention of S(2)-induced reentry was related to large increases in the effective refractory period (ERP) at a BCL of 1000 ms, leading to ERPs that exceeded the conduction delay following S(2). Reverse use-dependent effects resulted in smaller ERP increases at BCLs closer to the AF cycle length. Although the number of zones of reactivation per cycle during sustained AF were decreased by sotalol and dofetilide, the changes were small and insufficient to terminate AF. CONCLUSIONS: Sotalol and dofetilide prevent AF initiation by premature depolarizations at doses that fail to terminate vagotonic AF, by increasing ERP at the basic cycle length beyond the associated conduction delay that leads to reentry. PMID- 11282079 TI - Arachidonic acid protects neonatal rat cardiac myocytes from ischaemic injury through epsilon protein kinase C. AB - OBJECTIVES: Arachidonic acid is a second messenger which activates protein kinase C (PKC) and is released from the heart during ischaemic preconditioning. The purpose of this study was to examine the effect of arachidonic acid on activation of PKC in cardiac myocytes and the cellular consequences. METHODS: Neonatal rat cardiac myocytes were isolated and maintained in culture. Arachidonic acid induced activation of PKC was examined by cell fractionation and western blot analysis. Contraction frequency was measured by visual inspection under a microscope. Ischaemia was simulated by subjecting cells to an atmosphere of lower than 0.5% oxygen in the absence of glucose and cell damage determined by release of cytosolic lactate dehydrogenase or direct cell viability assay. RESULTS: Arachidonic acid resulted in translocation of delta and epsilonPKC but not alpha, betaII, eta or zetaPKC isozymes, indicating activation of only delta and epsilonPKC. Arachidonic acid induced a dose-dependent decrease in spontaneous contraction rate of cardiac myocytes which was blocked by a selective peptide translocation inhibitor of epsilonPKC. Pretreatment with arachidonic acid partially protected cardiac myocytes against ischaemia. Down-regulation of PKC with 24 h 4beta-phorbol,12-myristate,13-acetate treatment, inhibition of PKC by chelerythrine and selective inhibition of epsilonPKC translocation all decreased the protective effect of arachidonic acid. Pretreatment with eicosapentaenoic acid or oleic acid also protected cardiac myocytes against ischaemia. CONCLUSIONS: These results demonstrate that arachidonic acid selectively activates delta and epsilonPKC in neonatal rat cardiac myocytes, leading to protection from ischaemia. We suggest this is a potential mechanism of PKC activation during PC. In addition, our results suggest that different classes of free fatty acid directly exert cardioprotection from ischaemic injury in cardiac myocytes. PMID- 11282081 TI - Effects of verapamil on atrial fibrillation and its electrophysiological determinants in dogs. AB - BACKGROUND: Atrial tachycardia-induced remodeling promotes the occurrence and maintenance of atrial fibrillation (AF) and decreases L-type Ca(2+) current. There is also a clinical suggestion that acute L-type Ca(2) channel blockade can promote AF, consistent with an AF promoting effect of Ca(2+) channel inhibition. METHODS: To evaluate the potential mechanisms of AF promotion by Ca(2+) channel blockers, we administered verapamil to morphine-chloralose anesthetized dogs. Diltiazem was used as a comparison drug and autonomic blockade with atropine and nadolol was applied in some experiments. Epicardial mapping with 240 epicardial electrodes was used to evaluate activation during AF. RESULTS: Verapamil caused AF promotion in six dogs, increasing mean duration of AF induced by burst pacing, from 8+/-4 s (mean+/-S.E.) to 95+/-39 s (P<0.01 vs. control) at a loading dose of 0.1 mg/kg and 228+/-101 s (P<0.0005 vs. control) at a dose of 0.2 mg/kg. Underlying electrophysiological mechanisms were studied in detail in five additional dogs under control conditions and in the presence of the higher dose of verapamil. In these experiments, verapamil shortened mean effective refractory period (ERP) from 122+/-5 to 114+/-4 ms (P<0.02) at a cycle length of 300 ms, decreased ERP heterogeneity (from 15+/-1 to 10+/-1%, P<0.05), heterogeneously accelerated atrial conduction and decreased the cycle length of AF (94+/-4 to 84+/-3 ms, P<0.005). Diltiazem did not affect ERP, AF cycle length or AF duration, but produced conduction acceleration similar to that caused by verapamil (n=5). In the presence of autonomic blockade, verapamil failed to promote AF and increased, rather than decreasing, refractoriness. Neither verapamil nor diltiazem affected atrial conduction in the presence of autonomic blockade. Epicardial mapping suggested that verapamil promoted AF by increasing the number of simultaneous wavefronts reflected by separate zones of reactivation in each cycle. CONCLUSIONS: Verapamil promotes AF in normal dogs by promoting multiple circuit reentry, an effect dependent on intact autonomic tone and not shared by diltiazem. PMID- 11282082 TI - Loss of cyclin A and G1-cell cycle arrest are a prerequisite of ceramide-induced toxicity in human arterial endothelial cells. AB - BACKGROUND: Ceramide is an important messenger of TNF- and lipid-induced apoptosis. We previously demonstrated the adverse effect of TNF in the process of reendothelialization as well as the dependence of its effect on cell-cycle regulation. The current study was designed to investigate the linkage between ceramide induced toxicity and growth arrest in human endothelial cells. METHODS AND RESULTS: Cultured human arterial endothelial cells (HAEC) served as an in vitro model to test the cellular effects of C2-ceramide (C2). C2-induced cell death in HAECs occurred time- and dose-dependently. The LD(50) in subconfluent cells was three times lower than in confluent cell layers (25 vs. 75 microM). C2 caused up to 70% inhibition of BrdU and [3H]thymidine incorporation at non-toxic concentrations as a result of G1 cell-cycle arrest. Downregulation of cyclin A and p21(Cip1/Waf1) protein expression was observed independently of C2-toxicity, while expression of other cell-cycle regulatory genes was not affected. Inhibition of cyclin A protein expression by sequence-specific antisense oligonucleotides was paralleled by significant growth-inhibition. The protein phosphatase inhibitor okadaic acid induced endothelial cell proliferation, which was completely abrogated by C2. In contrast, aphidicolin-synchronized endothelial cells demonstrated elevated cyclin A levels along with 30% higher BrdU incorporation and 70% less C2-toxicity. G1-arrested cells, however, showed significantly enhanced C2-toxicity, lack of cyclin A expression and induction of uncleaved caspase-3 (CPP32). CONCLUSIONS: Ceramide abrogates endothelial cell proliferation independently of apoptosis or necrosis at low concentrations (55%); than that of patients with altered LVEF (LVEF< or =40%); and than that of patients with moderately reduced LVEF (40%A transition in the splice donor site of intron 4 of the sterol 27-hydroxylase gene. Serum cholestanol concentrations were decreased with the HMG-CoA reductase inhibitor simvastatin alone and greater reductions were achieved after the addition of the bile acid chenodeoxycholic acid; suggesting a synergistic effect of this combination. Despite serum cholestanol concentrations remaining within the low-normal range, there has been no significant improvement in mental and physical abilities or in EEG abnormalities with 5 years of treatment. Metabolism of radiolabeled 7-ketocholesterol to aqueous soluble products was absent in CTX derived macrophages. Consistent with this finding, plasma 7 alpha hydroxycholesterol, 7 beta-hydroxycholesterol, and 7-ketocholesterol concentrations were increased in the CTX subject compared with controls. PMID- 11282096 TI - A new quantitative analytical method of serum biotinidase activity using biocytin as a substrate and its clinical significance in Japan. AB - We have developed a new quantitative analytical method of serum biotinidase activity, which uses the native substrate biocytin, and to which can be applied the improved agar plate method of biotin bioassay. Assay characteristics were within acceptable ranges (intra-assay CVs, 4.44% and 1.95% at 1.82+/-0.08 and 3.08+/-0.06 pmol/min/ml; day-to-day CV, 5.92% at 2.68+/-0.16 pmol/min/ml). The enzyme activity with biocytin was stable at 4 degrees C for 90 days. The mean value of the serum biotinidase levels in 129 healthy adults was 2.71+/-0.93 pmol/min/ml. The method was clinically comparable with a colorimetric method for detection of biotinidase deficiency. Biotin supplementation treatment normalized our partial biotinidase deficiency patient's serum biotinidase activity. This normalized phenomenon has not yet been observed in a Caucasian patient. We also found that the distribution of the enzyme activities with biotinyl-p aminobenzoate in 8 of 11 patients with suspected biotin metabolic disorders shifted to a higher level than that of the controls. Although, we have few opportunities to analyze the sera of biotin metabolic disorders in Japan, the new method are suitable for clinical research applications in combination with the colorimetric method. PMID- 11282097 TI - Production and certification of an enzyme reference material for adenosine deaminase 1 (BCR 647). AB - BACKGROUND: We describe the preparation of a lyophilised reference material containing purified human adenosine deaminase 1 and the certification of its catalytic concentration. METHODS: The enzyme was purified from human erythrocytes. RESULTS: The enzyme was >99% pure on polyacrylamide gel electrophoresis. Only trace amounts (<0.4%) of alanine aminotransferase, aspartate aminotransferase and L-lactate dehydrogenase were detected in the purified fraction. The purified adenosine deaminase had a molar mass of 41600 g/mol and an isoelectric pH at 4.7, 4.85 and 5.0. The material was prepared by diluting the purified adenosine deaminase in a matrix containing 50 mmol/l Tris HCl buffer pH 7.4 and 30 g/l human serum albumin; dispensing in vials and freeze drying. The batch was homogeneous and the predicted loss of adenosine deaminase activity per year on the basis of accelerated degradation studies was 0.006% at 20 degrees C and 0.04% at 4 degrees C. The certified value for adenosine deaminase catalytic concentration in the reconstituted reference material is (2.55+/-0.09) microkat/l when measured by the method that uses adenosine as substrate and glutamate dehydrogenase as auxiliary enzyme at 37 degrees C. CONCLUSIONS: The material can be used to verify the comparability of results from different laboratories, for intra-laboratory quality control, or for calibration of the adenosine deaminase catalytic concentration measurements. PMID- 11282098 TI - Plasma chitotriosidase activity in beta-thalassemia major: a comparative study between Sicilian and Sardinian patients. AB - BACKGROUND: Chitotriosidase is a functional chitinase secreted by activated macrophages, which is extremely increased in plasma of patients with Gaucher disease (beta-glucocerebrosidase deficiency). Recently, we found that chitotriosidase plasma levels were increased to a variable extent in Sicilian patients with beta-thalassemia major. The aim of this study is to elucidate the possible mechanisms underlying chitotriosidase overproduction in beta-thalassemia major. METHODS: Plasma chitotriosidase was measured in 134 patients with beta thalassemia major (64 from Sardinia and 70 from Sicily), which are treated chronically by blood transfusions leading to systemic iron overload. They all have peripheral anemia and enormous enlargement of the reticulo-endothelial system. RESULTS: Plasma chitotriosidase activity was found most frequently elevated among Sardinian (48.4%) than Sicilian (17.1%) patients. In either group, the highest levels of plasma chitotriosidase were observed in patients with the highest degree of iron overload, suggesting that this factor could trigger chitotriosidase overproduction. CONCLUSIONS: The higher rate of subjects with increased plasma chitotriosidase values among Sardinian than Sicilian could be related to distinct molecular basis of beta-thalassemia and environmental features. PMID- 11282099 TI - Effect of five flavonoid compounds isolated from Quercus dentata Thunb on superoxide generation in human neutrophils and phosphorylation of neutrophil proteins. AB - Effect of five yellow compounds isolated from Quercus dentata on superoxide generation and protein phosphorylation in human neutrophils was investigated. The five yellow compounds examined were Kaempferol 3-O-beta-D-glucopyranoside (B), quercetin 3-O-beta-D-glucopyranoside (DA), Kaempferol 3-O-(6"-trans-p-coumaroyl) beta-D-glucopyranoside (D1), Kaempferol 3-O-(2"-6"-di-trans-p-coumaroyl)-beta-D glucopyranoside (D7) and Kaempferol 3-O-(2",4"-di-acetyl-3"-cis-p-coumaroyl-6" trans-p-coumaroyl)-beta-D-glucopyranoside (A). D7 suppressed significantly the superoxide generation induced by N-formyl-methionyl-leucyl-phenylalanine (fMLP). D1 and DA suppressed significantly the superoxide generation induced by arachidonic acid (AA). However, the superoxide generation induced by phorbol 12 myristate 13-acetate (PMA) was suppressed by all yellow compounds. When the cells were incubated with fMLP and D7, the tyrosyl phosphorylation of 67, 60, 58 and 38 kDa proteins of the cells was markedly decreased in a concentration-dependent manner. PMID- 11282100 TI - Effect of smoking on serum concentrations of total homocysteine and B vitamins in mid-pregnancy. AB - There are conflicting findings in the literature on the effect of smoking on total homocysteine (tHcy) concentrations in non-pregnant subjects. We evaluated the effect of smoking on serum concentrations of tHcy, folate, vitamin B-12 pyridoxal 5'-phosphate (PLP, a coenzyme form of vitamin B-6) in 196 women at 18 and 30 weeks' gestation. The smokers were defined as those who self-reported cigarette smoking and had serum concentrations of thiocyanate, a biomaker of smoking, in the highest quartiles of the population. Mid-pregnancy serum tHcy concentrations were not significantly different between smokers and non-smokers. Folate, vitamin B-12 and PLP concentrations were generally lower in smokers than non-smokers. In smokers, tHcy concentrations had significant negative correlations with folate at both time points. The multiple regression analyses indicated that serum folate concentration was the most significant factor associated with tHcy concentrations among smokers, whereas thiocyanate concentrations showed no such effect. We conclude that serum tHcy concentrations were most strongly associated with the nutritional status of folate among the B vitamins tested during mid-pregnancy in our subjects. We suggest that it is essential to consider the nutritional status of folate, vitamin B-12 and vitamin B-6 in evaluating the effect of smoking on homocysteine metabolism. PMID- 11282101 TI - Human kallikrein 10: a novel tumor marker for ovarian carcinoma? AB - BACKGROUND: Human kallikrein 10 (hK10, encoded by KLK10 gene) is a recently discovered member of the human kallikrein family. hK10 is a secreted serine protease. With the development of a highly sensitive and specific immunoassay for hK10, quantification of hK10 in the circulation is now feasible. Our aim was to investigate whether hK10 concentration in serum changes in various malignancies. METHODS: We used a highly specific and sensitive immunofluorometric assay to quantify hK10 protein in 374 serum samples from healthy individuals and patients with various malignancies. RESULTS: Serum hK10 concentration was found to be significantly elevated in 56% of the ovarian cancer patients and such an increase was not observed in serum of healthy individuals or in serum of patients with other types of cancer, with the exception of approximately 15% of patients with gastrointestinal cancer. This hK10 elevation does not correlate well with CA 125. We have further demonstrated that hK10 concentration changes during ovarian cancer progression. CONCLUSION: This is the first report describing that hK10 serum concentration is significantly elevated in the majority of ovarian cancer patients. Our results indicate that hK10 may be a potential new serological marker for ovarian cancer diagnosis and monitoring. PMID- 11282102 TI - Leukocyte activation, erythrocyte damage, lipid profile and oxidative stress imposed by high competition physical exercise in adolescents. AB - BACKGROUND: The aim of this study was to evaluate and to compare the lipid profile and the levels of leukocyte activation, red blood cell (RBC) damage and of oxidative stress in two groups of adolescents, with similar body mass index: high competition swimmers and adolescents practising moderate regular physical exercise. METHODS: As markers of leukocyte activation, we measured plasma lactoferrin, elastase and granulocyte-monocyte colony stimulating factor. We studied RBC membrane band 3 profile and membrane-bound hemoglobin, as markers of RBC damage and aging; total and differential leukocyte count and RBC count, hematocrit, hemoglobin concentration and hematimetric indexes were also measured. Lipid profile included the evaluation of triglycerides (TG), total cholesterol (Chol), high-density lipoprotein cholesterol (HDLc), low-density lipoprotein cholesterol (LDLc), apolipoproteins AI and B (Apo AI and B), and lipoprotein (a) (Lp(a)). To evaluate oxidative stress, lipoperoxidation products and total antioxidant capacity were measured. RESULTS: We found that high competition adolescents presented increased plasma levels of leukocyte activation products, increased RBC damage suggesting aging and premature removal, and higher oxidative stress. Lipid profile showed some risk and some protective changes. CONCLUSIONS: Our data suggest that high competition exercise, by imposing a higher and sustained oxidative and proteolytic stress, may contribute in the future to a higher risk of cardiovascular disease. We believe these findings warrant a reevaluation of current views in the intensity, duration and regularity of physical exercise, and that the evaluation of leukocyte activation products, RBC damage, oxidative stress and lipid profile may represent good markers to establish putative protective thresholds. PMID- 11282103 TI - Serum and ascitic fluid nitrate levels in patients with cirrhosis. AB - BACKGROUND: Increased nitric oxide level may play a critical role in the hemodynamic disturbances in patients with cirrhosis. There are few reports investigating the factors related to this increase and their results are controversial. The purpose of this study was to reveal the clinical importance of nitric oxide levels and the possible factors related to this increase in patients with cirrhosis. METHODS: Serum and ascites nitrate levels were studied in 50 patients with cirrhosis and 10 control subjects. RESULTS: All cirrhotic patients (groups 2, 3, 4, 5, 6) showed significant increase in serum nitrate levels in comparison with that in control subjects (group 1) (p<0.001). Serum nitrate levels were significantly higher (282.4+/-111.3 micromol/l; p<0.05) in patients with spontaneous bacterial peritonitis (group 2) when compared with those in cirrhotic patients without spontaneous bacterial peritonitis (group 3) (186.4+/ 87.6 micromol/l). Ascitic fluid nitrate levels were significantly higher (302.4+/ 66 micromol/l; p<0.001) in patients with spontaneous bacterial peritonitis (group 2) when compared with those in cirrhotic patients without spontaneous bacterial peritonitis (group 3) (135.4+/-65.8 micromol/l). Serum nitrate levels were significantly lower in cirrhotic patients without ascites (group 5) when compared with those in cirrhotic patients with ascites (group 3) (98.8+/-52.6 vs. 186.4+/ 87.6 micromol/l; p<0.05). No significant differences were found among patients with severe anemia (groups 4, 6) and other cirrhotic patients (group 3) (174.5+/ 54.5, 168.8+/-63.8 vs. 186.4+/-87.6 micromol/l; p>0.05). Cirrhotic patients with Child--Pugh B and C scores showed higher serum nitrate levels (179.4+/-81.1, 222.5+/-101.7 micromol/l; p<0.001) than did cirrhotic patients with Child--Pugh A score (85.8+/-59.7 micromol/l). CONCLUSION: Our findings suggest that overproduction of nitric oxide in cirrhotic patients may be related to the severity of liver damage and spontaneous bacterial peritonitis but not related to their anemia. PMID- 11282105 TI - Comparison of BD Vacutainer SST Plus Tubes with BD SST II Plus Tubes for common analytes. AB - Serum separator tubes were introduced 25 years ago and are widely used in the clinical laboratory today for routine collection of blood. These tubes have gained widespread acceptance due to the advantage of the barrier gel that facilitates rapid separation of serum from cellular constituents of blood and thus reduces hemolysis. However, there are some limitations associated with gel tubes (i.e., gel stability and analyte incompatibilities). The serum separator BD SST tubes manufactured by BD are widely used in clinical laboratories. Recently, BD has developed a new barrier gel, which is superior to the existing gel. We studied the stability of common analytes when serum specimens were stored in the new BD SST II tubes by comparing the performance with the existing BD SST tubes. We did not observe any significant reduction in concentrations of 42 commonly ordered analytes using the new BD SST II tubes. Significant differences were noted at low serum volumes for estradiol in both tube types over time. We conclude that the new BD SST II tubes are suitable for collection of blood and storage of serum for commonly ordered laboratory tests. PMID- 11282104 TI - Evaluation of standard tyrosinase RT-PCR in melanoma patients by the use of the LightCycler system. AB - BACKGROUND: Haematogenous spread influences outcome in melanoma patients. The clinical relevance of detecting circulating melanoma cells in peripheral blood by tyrosinase mRNA RT-PCR is, however, questioned as rates of positivity considerably vary between studies. Standard tyrosinase-nested RT-PCR was here compared with a real-time PCR technique. METHODS: Forty-three blood samples from 20 stage III--IV melanoma patients were analyzed. Mononuclear cells were isolated using a Ficoll Hypaque gradient technique. Total RNA extracted by the acid guanidinum thiocyanate-phenol-chloroform method was reverse transcribed using random hexamers or specific primers. Standard tyrosinase-nested PCR was performed on Touchdown machine (Hybaid) and real-time PCR on a LightCycler instrument (Roche). RESULTS: Only two samples from stage IV patients (one from random hexamers, one from antisense primers) were found tyrosinase positive with a 100% agreement between the two PCR techniques. A 10-fold dilution of the first-round products improved the PCR kinetic and the final amount of amplified product of positive samples, but not the rate of positivity. CONCLUSIONS: Efficiency of the PCR reaction can be monitored in an online fashion by the LightCycler instrument allowing technical improvements. However, tyrosinase mRNA RT-PCR cannot be yet considered as a useful technique in the monitoring of melanoma patients. PMID- 11282106 TI - Expression of inducible nitric oxide synthase in primary culture of rat bladder smooth muscle cells by plasma from cyclophosphamide-treated rats. AB - Intraperitoneal administration of cyclophosphamide (50-150 mg/kg) for 6 or 12 h induced edema and hemorrhagic changes in rat bladder, which were both dose and time-dependent. Pretreatment with nitric oxide synthase (NOS) inhibitors N(G) nitro-L-arginine methyl ester (L-NAME, 40 mg/kg) or with s-methylisothiourea (40 mg/kg) ameliorated the cyclophosphamide-induced cystitis. Cyclophosphamide administration also produced increases in NO-metabolite levels (nitrate+nitrite) in the urine and plasma of rats. Greater increases in NO metabolites were observed with 150 than with 50 mg/kg of cyclophosphamide, and at 12 than at 6 h after cyclophosphamide injection. Pretreatment with L-NAME and s methylisothiourea significantly reduced cyclophosphamide-induced increases in urine and plasma NO-metabolite levels. To explore the mechanism by which cyclophosphamide increases the expression of inducible NOS (iNOS), primary cultures of rat bladder smooth muscle were developed. Exposure to tumor necrosis factor alpha (TNF-alpha) plus interferon gamma, produced a marked increase in the expression of iNOS and in NO production in the culture medium. However, exposure to cyclophosphamide or to its metabolite acrolein (10(-6)-10(-4) M for 24 h) did not increase iNOS or NO-metabolite levels. On the other hand, incubation of primary cell cultures with plasma from rats treated with cyclophosphamide (150 mg/kg, 12 h) produced a marked increase in iNOS expression and NO production. Taken together, our results indicate that NO plays an important role in the pathogenesis of cyclophosphamide-induced cystitis in rats, and some factors may be released in cyclophosphamide-treated rat plasma which stimulate iNOS expression in primary culture of rat bladder smooth muscle cells. PMID- 11282107 TI - Phenol derivatives accelerate inactivation kinetics in one inactivation-deficient mutant human skeletal muscle Na(+) channel. AB - Altered inactivation kinetics in skeletal muscle Na(+) channels due to mutations in the encoding gene are causal for the alterations in muscle excitability in nondystrophic myotonia. Na(+) channel blockers like lidocaine and mexiletine, suggested for therapy of myotonia, do not reconstitute inactivation in channels with defective inactivation in vitro. We examined the effects of four methylated and/or halogenated phenol derivatives on one heterologously expressed inactivation-deficient Paramyotonia congenita-mutant (R1448H) muscle Na(+) channel in vitro. All these compounds accelerated delayed inactivation of R1448H whole-cell currents during a depolarization and delayed accelerated recovery from inactivation. The potency of these effects paralleled the potency of the drugs to block the peak current amplitude. We conclude that the investigated phenol derivatives affect inactivation-deficient Na(+) channels more specifically than lidocaine and mexiletine. However, for all compounds, the effect on inactivation was accompanied by a substantial block of the peak current amplitude. PMID- 11282108 TI - Steroids affect collateral sensitivity to gemcitabine of multidrug-resistant human lung cancer cells. AB - Gemcitabine is phosphorylated by deoxycytidine kinase and thymidine kinase 2 and during S-phase incorporated into DNA. The steroids cortisol and dexamethasone, which regulate cell proliferation and gene expression, are pumped out of the cell by the membrane efflux pumps P-glycoprotein and multidrug resistance-associated protein (MRP), which are blocked by verapamil. In parental non-small cell lung cancer (NSCLC) cells (SW1573), 5 microM cortisol and 100 nM dexamethasone decreased sensitivity to gemcitabine. However, both cortisol and dexamethasone only decreased sensitivity with verapamil in MRP (2R120) and P-glycoprotein (2R160) overexpressing variants. Cortisol decreased deoxycytidine kinase activity in SW1573 cells and cortisol with verapamil in 2R120 and 2R160 cells. Dexamethasone with verapamil decreased deoxycytidine kinase activity in 2R160. Cortisol decreased thymidine kinase 2 activity in 2R120 and 2R160 cells. Dexamethasone decreased thymidine kinase 2 activity in SW1573, 2R120 and 2R160 cells. In conclusion, since dexamethasone is frequently used to treat side effects of oncolytic therapy, a decrease of sensitivity to gemcitabine by steroids might be clinically relevant. PMID- 11282109 TI - Induction of apoptosis by esculetin in human leukemia cells. AB - Esculetin, a coumarin compound, has been shown to exhibit antioxidant and anti inflammatory effects. In the present study, esculetin was found to inhibit the survival of human promyelocytic leukemia HL-60 cells in a concentration-dependent and time-dependent manner. HL-60 cells underwent internucleosomal DNA fragmentation and morphological changes characteristic of apoptosis after a 24-h treatment with esculetin (100 microM). Flow cytometric analysis showed that the hypodiploid nuclei of HL-60 cells were increased to 40.93% after a 36-h treatment with esculetin (100 microM). Further investigation showed that esculetin induced the release of cytochrome c from mitochondria into cytosol in a time-dependent and concentration-dependent manner. Moreover, esculetin application reduced Bcl-2 protein expression to 58% after 9 h as compared with that time at 0. Cysteine protease 32 kDa proenzyme (CPP32), a caspase 3, was activated and its substrate, poly (adenosine diphosphate-ribose) polymerase, was cleaved after a 24-h treatment of HL-60 cells with esculetin. These data suggest that esculetin induces apoptosis in human leukemia cells by increasing cytosolic translocation of cytochrome c and activation of CPP32. PMID- 11282110 TI - Ligand induced conformational states of the 5-HT(1A) receptor. AB - It has been shown that G-protein coupled receptors have seven transmembrane alpha helices, but the structural changes occurring in a G-protein coupled receptor as a response on agonist stimulus and the molecular events leading to blockade of the signal transduction by antagonists are not well understood. In the present study, the AMBER 5.0 force field was used for comparative molecular dynamics simulations of a 5-HT(1A) receptor model in the absence of ligand, in complex with a 5-HT(1A) receptor agonist (R)-8-hydroxy-2-(di-n-propylamino)tetralin [(R) 8-OH-DPAT], in complex with a selective 5-HT(1A) receptor antagonist (S)-N-tert butyl-3-[4-(2-methoxyphenyl)piperazin-1-yl ]-2-phenylpropanamide [(S)-WAY100135], and in complex with the partial agonist, buspirone. In the simulations, the agonist induced larger conformational changes into transmembrane helix 3 and 6 than into the other helices, while the main conformational differences between the agonist bound receptor and the antagonist bound receptor were in transmembrane helix 5 and 6. During the simulations, all the three ligands constrained the helical movements compared to those observed in the receptor without any ligand. PMID- 11282111 TI - Neuropeptide Y inhibits the protein kinase C-stimulated Cl(-) secretion in the human colonic cell line HT29cl.19A cell line via multiple sites. AB - Neuropeptide Y is known to exert inhibitory effects on ion secretion in the intestine by reducing the activity of adenylyl cyclase. In the human intestinal epithelial cell line HT29cl.19A, it has been previously shown that neuropeptide Y inhibits the electrophysiological phenomena related to Cl(-) secretion, when induced by elevation of cAMP via forskolin. Moreover, the secretion induced via elevation of intracellular calcium levels via muscarinic activation can be inhibited by neuropeptide Y. Part of these inhibitions appeared to be due to lowered calcium activity in the epithelial cells, thereby reducing the basolateral K(+) conductance. The phorbol ester 4-phorbol-12,13-dibutyrate (PDB) can induce secretion in this cell line via activation of protein kinase C as well; however, the effect of neuropeptide Y on this pathway has not yet been studied. In the present experiments, it is shown that neuropeptide Y inhibits the PDB-induced secretion at two sides: one located in the apical membrane and another in the basolateral membrane. It is shown that the latter effect can, at least partially, be explained via a direct effect of neuropeptide Y on the K(+) conductance. This was concluded from the observation that neuropeptide Y could also reduce basolateral K(+) conductance when intracellular calcium was dramatically increased by ionomycin. The observed inhibitory effects suggest that neuropeptide Y is a very powerful antisecretory peptide in human intestinal epithelial cells. PMID- 11282112 TI - Antinociceptive effect of clomipramine in monoarthritic rats as revealed by the paw pressure test and the C-fiber-evoked reflex. AB - The antinociceptive effect of clomipramine was studied in monoarthritic rats by using the paw pressure test and the C-fiber-evoked reflex. Monoarthritis was produced by intra-articular injection of complete Freund's adjuvant into the tibio-tarsal joint. Joint circumference as well as vocalization threshold to graded paw pressure were evaluated weekly during a 14-week period after the intra articular injection. At week 8, monoarthritic and vehicle-injected control rats were given either clomipramine or saline and both the paw pressure threshold and inhibition of the C-fiber-evoked reflex response were evaluated. Results showed that (i) 1.5, 3.0, and 6.0 mg/kg, i.v. of clomipramine induced significantly greater dose-dependent antinociception to paw pressure testing in the monoarthritic group, as compared to the control one; and (ii) 0.75, 1.5, 3.0, and 6.0 mg/kg, i.v. of clomipramine exerted significantly higher dose-dependent inhibition of the C-reflex activity in monoarthritic rats than in controls. Results suggest that the higher sensitivity to clomipramine in monoarthritic rats could be related to adaptive changes occurring in monoamine metabolism or in other neurotransmitter systems during chronic pain. PMID- 11282113 TI - Protective effect of the antioxidant 6-ethoxy-2,2-pentamethylen-1,2 dihydroquinoline (S 33113) in models of cerebral neurodegeneration. AB - In a previous study Dorey et al. [Bio. Org. Chem. Lett., 10 (2000) 935] a series of novel dihydroquinoline compounds were developed, based on the potent antioxidant 6-ethoxy-1,2-dihydro-2,2,4-trimethylquinoline (ethoxyquin), and permitted the selection of the analogue 6-ethoxy-2,2-pentamethylen-1,2 dihydroquinoline (S 33113) lacking the hypothermic effects associated with ethoxyquin at equivalent doses. Herein, an extensive investigation of the neuroprotective capacity of S 33113 in different in vitro and in vivo paradigms of oxidative stress-mediated cellular degeneration was undertaken. In vitro S 33113 was a potent inhibitor (IC(50) = 0.29 microM) of Fenton-reaction-induced lipid peroxidation in mouse cortical membranes. Administration of S 33113 either intraperitoneally (< or =150 mg/kg i.p.) or orally (< or =600 mg/kg p.o.) did not significantly modify body temperature in NMRI mice. Furthermore, S 33113 (150 mg/kg i.p. or 600 mg/kg p.o.) markedly reduced the lethality induced by an intracerebroventricular injection of t-butylhydroperoxide in NMRI (naval medical research institute) mice for up to 5 h. Oral administration of S 33113, significantly attenuated alloxan-mediated hyperglycaemia in NMRI mice at 400 and 600 mg/kg (60%; P < 0.001). Administration of S 33113 (150 mg/kg i.p.) 30 min before transient global ischaemia significantly prevented delayed neuronal cell death in the CA1 region of the rat hippocampal formation, 7 days post-ischaemia (33% cell loss vs. 88% in ischaemia controls; P < 0.001). Similarly, a single pre administration of S 33113 (150 mg/kg i.p.) prevented kainic acid-induced cell death in the CA3 hippocampal region at 7 days post-exposure (17% cell loss vs. 52% in kainate-treated controls; P < 0.01). Furthermore, D-methamphetamine mediated dopamine depletion in the striatum of C57BL/6 mice (39-46%) was significantly prevented with S 33113 administered at either (2 x 150mg/kg i.p.) (11%; P < 0.01) or (2x150 mg/kg p.o.) (17%; P < 0.001). In conclusion, S 33113 represents a novel dihydroquinoline compound with potential for the treatment of cerebral pathologies implicating chronic neurodegeneration. PMID- 11282114 TI - The effect of N-acetyl-L-aspartic acid dilithium salt on dopamine release and synthesis in the rat striatum in vivo. AB - The effect of the dilithium salt of N-acetyl-L-aspartic acid on release and synthesis of dopamine in the striatum was investigated using microdialysis in freely moving rats. Intrastriatal infusion of 1 mM N-methyl-D-aspartate, an NMDA receptor agonist, augmented extracellular dopamine to 215% of baseline, while 1 mM dilithium N-acetyl-L-aspartate increased dopamine release to 190% of baseline in rat striatum. Infusion of DL-2-amino-5-phosphonopentanoic acid, a competitive NMDA receptor antagonist, prior to infusion of dilithium N-acetyl-L-aspartate did not significantly alter basal levels of dopamine, but reversed the dilithium N acetyl-L-aspartate-evoked elevation in extracellular dopamine. Intrastriatal perfusion with 6-cyano-7-nitroquinoxaline-2,3-dione, an AMPA/kainate receptors antagonist, altered neither basal levels of dopamine nor dilithium N acetylaspartate-induced dopamine release. When the striatum was continuously perfused with the inhibitor of L-aromatic amino acid decarboxylase, 3 hydroxybenzylhydrazine dihydrochloride (100 microM), both dilithium N acetylaspartate and NMDA added to the perfusate increased extracellular 3,4 dihydroxyphenyl-L-alanine, reflecting the effect of the compounds on the biosynthesis of dopamine. The data suggest that availability of dilithium N acetyl-L-aspartate to activate dopamine turnover and release in the rat striatum may be mediated by presynaptic NMDA heteroreceptors located at dopaminergic neurons. PMID- 11282115 TI - Spontaneous and precipitated withdrawal with a synthetic cannabinoid, WIN 55212 2. AB - Physical dependence on the synthetic cannabinoid-receptor agonist R(+)-[2,3 dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1 naphthalenyl) methanone mesylate (WIN 55212-2) was demonstrated in rats by the use of a chronic continuous infusion. Spontaneous withdrawal, of moderate intensity, was shown for the first time with this class of drugs of abuse. Behavioral withdrawal signs were also elicited after challenge with (N-(piperidin 1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3 carboxamide.HCl (SR141716A), a specific CB(1) cannabinoid-receptor antagonist. In both instances, the high-dose regimen (4, 8, 16 and 16 mg/kg/day, i.p. on days 1 4, respectively) was sufficient to evoke a typical withdrawal syndrome quantified by the signs wet-dog shakes and facial rubs. These results are discussed relative to those obtained with Delta(9)-tetrahydrocannabinol and anandamide. With Delta(9)-tetrahydrocannabinol, precipitated but not spontaneous or abrupt withdrawal was observed, and this was ascribed to pharmacokinetic properties. Anandamide, which showed little, if any, physical dependence potential, behaved atypically. Possible implications regarding pharmacotherapeutic and human abuse issues are discussed. PMID- 11282116 TI - Receptor binding and biological activity of the dermorphin analog Tyr-D-Arg(2) Phe-Sar (TAPS). AB - The binding of Tyr-D-Arg(2)-Phe-sarcosine(Sar)(4) (TAPS), a proposed mu-opioid receptor-selective tetrapeptide analog of dermorphin to opioid receptors, was studied using selective binding assays for subtypes of mu-, delta- and kappa opioid receptors. Subtype specific mu-opioid receptor binding was further characterized in the presence of sodium and guanosine nucleotides and the activity of TAPS in isolated guinea pig ileum was compared to other mu-opioid receptor-selective ligands. Further, the antinociceptive properties of TAPS following intrathecal (i.t.) administration in rats, as a model of spinal antinociception, were evaluated. The K(i)-values for TAPS at the mu(1)- and mu(2) opioid receptor sites were 0.4 and 1.3 nM, respectively, suggesting high affinity binding to mu-opioid receptor binding sites with an increased selectivity to mu(1)-opioid receptor sites. The attenuated reduction of TAPS binding at the mu(2)-opioid receptor subtype in the presence of the stable guanosintriphosphate analog 5'-guanylylimidodiphosphate and sodium suggests a potential partial antagonist mode of action at this site. PMID- 11282117 TI - Role of cholecystokinin in the reduction of endomorphin-2-induced antinociception in diabetic mice. AB - We examined the role of cholecystokinin in the reduction of endomorphin-2-induced antinociception in diabetic mice. Endomorphin-1 (1-10 microg, i.c.v.) and endomorphin-2 (3-30 microg, i.c.v.) dose dependently inhibited the tail-flick response in non-diabetic and diabetic mice. There was no significant difference between the antinociceptive effect of endomorphin-1 in non-diabetic and diabetic mice. On the other hand, the antinociceptive effect of endomorphin-2 in diabetic mice was significantly less than that in non-diabetic mice. Cholecystokinin octapeptide (CCK-8) dose dependently reduced the antinociceptive effects of endomorphin-1 and endomorphin-2 in non-diabetic mice. However, in diabetic mice, CCK-8 significantly inhibited the antinociceptive effect of endomorphin-1, but not of endomorphin-2. In non-diabetic mice, CI-988 ((R-[R*,R*])-4-([3-1H-indol]-3 yl)-2-methyl-1-oxo-2-([(tricyclo(3.3.1.1)dec-2-yloxy)carbonyl] amino)propylamino 1-phenyl-ethylamino-4-oxybutanoic acid) had no significant effect on either endomorphin-1- or endomorphin-2-induced antinociception. In diabetic mice, while CI-988 had no significant effect on endomorphin-1-induced antinociception, it dose dependently enhanced the antinociceptive effect of endomorphin-2. The results indicated that the reduction of endomorphin-2-induced antinociception in diabetic mice might be due, at least in part, to the activation of CCK(2) receptors. PMID- 11282118 TI - Fibroblast growth factor inhibits locomotor activity as well as feeding behavior of rats. AB - The effects of acute and chronic intracerebroventricular (i.c.v.) administration of basic fibroblast growth factor (bFGF) on behavior were examined in free feeding rats. An i.c.v. injection of bFGF induced behavioral changes, such as an increase in resting and decreases in grooming, moving, and food intake at a dose of 20 or 50 ng. These effects appeared at 4-5 h and lasted at least 11 h after the injection. These changes, as well as inhibition of body weight gain, were also found during a 6-day period of chronic i.c.v. infusion of bFGF at a dose of 20 ng/h. These results indicate that bFGF as both bolus i.c.v. injection and chronic i.c.v. infusion inhibits not only feeding behavior but also locomotor activity in rats. It is suggested that the inhibitory effect of bFGF on food intake may be in part ascribed to the suppression of behavior by bFGF. PMID- 11282119 TI - Carbamazepine prevents imipramine-induced behavioural sensitization to the dopamine D(2)-like receptor agonist quinpirole. AB - Chronic treatment with antidepressants potentiates the behavioural sensitivity to the administration of dopamine receptor agonists. Such supersensitivity might be involved in the mechanism of action of antidepressant drugs, but it has also been suggested to play a role in the mechanisms underlying antidepressant treatment related mania (i.e. antidepressant-induced mood switch and rapid cycling). Consistently to this hypothesis, we have recently shown that lithium salts, which are poorly effective in antidepressant-related mania, fail to prevent the development of imipramine-induced supersensitivity to the locomotor effect of the dopamine D(2)-like receptor agonist quinpirole. In the present paper, we report the ability of carbamazepine, an anticonvulsant with antimanic and mood stabiliser properties, to prevent the development of supersensitivity to the locomotor response to quinpirole induced by chronic treatment with imipramine. The present results, together with the results of our previous study, might contribute to explain the different responsiveness to lithium and carbamazepine observed in some manic patients, and are consistent with the clinical data suggesting that carbamazepine might be more effective than lithium in antidepressant-related mania. PMID- 11282120 TI - Vasopressin-induced contraction in the rat basilar artery in vitro. AB - Vasopressin ([Arg(8)]vasopressin)-induced contraction was characterized using receptor agonists and antagonists for vasopressin and channel blockers in the rat basilar artery ring preparations. Vasopressin induced rhythmic contractions superimposed on a contraction in endothelium-intact preparations but not in denuded ones. Endothelium removal shifted the concentration-response curve for vasopressin leftward and upward. In endothelium-denuded preparations, vasopressin V(1) receptor antagonist shifted the concentration-response curve for vasopressin downward and rightward. Vasopressin V(1) receptor agonist caused contraction but V(2) receptor agonist did not. The contractile response to vasopressin was partly inhibited by nifedipine, SK&F 96365 (1-[beta-[3-(4-methoxyphenyl)propoxy]-4 methoxyphenethyl]-1H-imidazole) and niflumic acid. In the absence of extracellular Ca(2+), vasopressin produced a transient contraction. Charybdotoxin produced an upward and leftward shift of the concentration-response curve for vasopressin. These results suggest that vasopressin elicits contraction due to Ca(2+) influx through voltage-dependent and receptor-operated Ca(2+) channels and to Ca(2+) release from Ca(2+) stores by activating vasopressin V(1) receptors in the rat basilar artery. PMID- 11282121 TI - Pulmonary vascular remodelling in hypoxic rats: effects of amlodipine, alone and with perindopril. AB - This study investigated whether pulmonary vascular remodelling in hypoxic pulmonary hypertensive rats (10% oxygen; 4 weeks) could be prevented by treatment, during hypoxia, with amlodipine (10 mg/kg/day, p.o.), either alone or in combination with the angiotensin converting enzyme inhibitor, perindopril (30 mg/kg/day, p.o.). Medial thickening of pulmonary arteries (30-500 microm o.d.) was attenuated by amlodipine whereas it was totally prevented by the combination treatment (amlodipine plus perindopril); neomuscularisation of small alveolar arteries (assessed from critical closing pressure in isolated perfused lungs) was not affected. Pulmonary vascular resistance (isolated perfused lungs) was reduced by both treatment regimes but only combination treatment reduced right ventricular hypertrophy. Thus, amlodipine has anti-remodelling properties in pulmonary hypertensive rats. The finding that combining amlodipine with another anti-remodelling drug produced effects on vascular structure that were additive raises the question of whether combination therapy with two different anti remodelling drugs may be of value in the treatment of patients with hypoxic (and possibly other forms of) pulmonary hypertension. PMID- 11282122 TI - Troglitazone corrects metabolic changes but not vascular dysfunction in dietary obese rats. AB - Insulin resistance has been attributed to the defect in vascular function associated with obesity, type 2 diabetes and dyslipidaemia. We have investigated vascular effects of chronic (3-week) administration of troglitazone on female Wistar rats with moderate dietary obesity. Compared with lean controls, untreated obese rats had significantly higher body weights, fat pad masses, plasma triglycerides, free fatty acids and leptin levels (for all P < 0.01). These metabolic changes were corrected by troglitazone treatment. In mesenteric arteries, responses to noradrenaline or KCl were similar in all groups. However, in noradrenaline-preconstricted arteries, vasorelaxations to acetylcholine and insulin were significantly (50-60% less than in lean, P < 0.001) attenuated in both untreated and troglitazone-treated obese rats. Relaxations to sodium nitroprusside showed similar but lesser impairment in both untreated and troglitazone-treated obese animals. Our data show that although troglitazone markedly improved obesity-induced metabolic changes, it failed to correct vascular dysfunction associated with obesity in female Wistar rats. PMID- 11282123 TI - Effect of JTH-601, a putative alpha(1L)-adrenoceptor antagonist, on guinea pig nasal mucosa vasculature. AB - The existence of alpha(1)-adrenoceptors with low affinity for prazosin, an alpha(1L) subtype, has been proposed in addition to alpha(1)-adrenoceptor subtypes with high affinity for prazosin, i.e. the alpha(1H) group: alpha(1A), alpha(1B) and alpha(1D) subtypes. In the present study, we investigated the effect of JTH-601 (3-(N-[2-(4-hydroxy-2-isopropyl-5-methylphenoxy)ethyl]-N methylaminomethyl)-4-methoxy-2,5,6-trimethylphenol hemifumarate), a putative alpha(1L)-adrenoceptor antagonist, on the isolated guinea pig nasal mucosa vasculature. JTH-601 (0.01-0.03 microM) competitively antagonized the noradrenaline-induced contraction of the tissue in a concentration-dependent manner. The pA(2) value for JTH-601 was 8.14 +/- 0.04 (means +/- SEM, n = 6). The data suggests that the alpha(1L)-subtype is involved in the noradrenaline-induced contraction of the guinea pig nasal mucosa vasculature. PMID- 11282124 TI - Variable responses to prostaglandin E(2) in human non-pregnant myometrium. AB - Cumulative concentration-effect curves for prostaglandin E(2), sulprostone and butaprost were constructed in matched strips of human non-pregnant myometrium from 14 different donors. All samples were obtained from the mid-lateral wall of the uterus. Prostaglandin E(2) produced four types of concentration-effect curves: monophasic inhibitory (n = 7), monophasic excitatory (n = 2), biphasic consisting of an excitatory phase followed by an inhibitory phase (n = 4), and biphasic consisting of an inhibitory phase followed by an excitatory phase (n = 1). Sulprostone produced excitation of spontaneous contractile activity in all tissues (mean pEC(50) = 9.1+/-0.2, range 8.1-10.1, n = 14). Butaprost produced relaxation of cloprostenol-stimulated contractile activity in all tissues (mean pEC(50) = 5.7 +/- 0.1, range 5.0-6.9, n = 14). The mean pEC(50) value for sulprostone was significantly higher in tissues where prostaglandin E(2) caused some excitation (pEC(50) = 9.4 +/- 0.2, n = 7) compared to those where prostaglandin E(2) caused only inhibition (pEC(50) = 8.8 +/- 0.2, n = 7). Mean pEC(50) values for butaprost were not significantly different between these groups. These data suggest that (a) variability in EP receptor-mediated responses exists within a single anatomical site; (b) both excitatory and inhibitory EP receptor-mediated pathways are always operative in human non-pregnant myometrium, regardless of the type of tissue response to prostaglandin E(2); and (c) regulation of EP receptor-mediated responses occurs predominantly in the excitatory (EP(3) or EP(1) receptor) pathway rather than the inhibitory (EP(2) receptor) pathway. PMID- 11282125 TI - Functional properties of atypical beta-adrenoceptors on the guinea pig duodenum. AB - In this study, we attempted to further characterize atypical beta-adrenoceptors on the guinea pig duodenum. (-)-Enantiomers of isoprenaline and noradrenaline were more potent than its (+)-enantiomers. The isomeric activity ratios ((+)/(-)) were less than those obtained in the guinea pig atria and trachea. The concentration-response curves to catecholamines ((-)-isoprenaline, (-) noradrenaline and (-)-adrenaline), to the selective beta(3)-adrenoceptor agonist, BRL37344 ((R*, R*)-(+/-)-4-[2-[(2-(3-chlorophenyl)-2 hydroxyethyl)amino]propyl]phenoxyacetic acid sodium), and to the non-conventional partial beta(3)-adrenoceptor agonist, (+/-)-CGP12177A ((+/-)-[4-[3-[(1,1 dimethylethyl)amino]-2-hydroxypropoxy]-1,3-dihydro-2H-benzimidazol-2-one] hydrochloride), were resistant to blockade by (+/-)-pindobind, the beta adrenoceptor alkylating agent. (-)-Noradrenaline and (-)-adrenaline were more potent than dopamine and (-)-phenylephrine, respectively. Selective beta(2) adrenoceptor agonists possess agonistic activities at atypical beta adrenoceptors. (+/-)-Propranolol and (+/-)-bupranolol had no agonistic effect, whereas (+/-)-alprenolol, (+/-)-pindolol, (+/-)-nadolol, (+/-)-CGP12177A and (+/ )-carteolol exhibited agonistic activities at atypical beta-adrenoceptors. These results suggest that pharmacological properties of atypical beta-adrenoceptors differ from those of conventional beta(1)- and beta(2)-adrenoceptors on the guinea pig. PMID- 11282127 TI - Changes in paw oedema triggered via bradykinin B(1) and B(2) receptors in streptozotocin-diabetic rats. AB - The present study investigated hind paw oedema mediated by bradykinin B(1) and B(2) receptors in streptozotocin-diabetic rats. Paw oedema induced by intraplantar (i.pl.) injection of bradykinin or the selective bradykinin B(2) receptor agonist, Tyrosine(8)-bradykinin ([Tyr(8)]bradykinin) (both 3 nmol/paw), was significantly reduced at 4 weeks after streptozotocin treatment (34 +/- 8% and 40 +/- 7%). At 6 weeks after streptozotocin, when paw oedema caused by substance P or prostaglandin E(2) (both 10 nmol/paw) was unchanged, inhibition of bradykinin B(2) receptor-mediated oedema was maximal (66 +/- 6% and 72 +/ -2%, for bradykinin and [Tyr(8)]bradykinin, respectively). The selective bradykinin B(1) receptor agonist, [des-Arg(9)]bradykinin (100 nmol/paw), induced only slight paw oedema in non-diabetic controls. Responses to [des-Arg(9)]bradykinin were markedly enhanced 8 weeks after streptozotocin (from 0.09 +/- 0.01 to 0.38 +/- 0.05 ml), less so at 10 weeks (0.22 +/- 0.03 ml), and returning to basal values at 12 weeks (0.11 +/- 0.03 ml). Treatment with insulin protamine zinc (1-3 U/day/7 weeks, s.c.) did not reverse the inhibition of responses to [Tyr(8)]bradykinin or the potentiation of responses to [des-Arg(9)]bradykinin seen at 8 weeks. Thus, streptozotocin-induced diabetes induces long-lasting alterations in oedematogenic responsiveness to kinins in the rat, characterized by marked reduction of oedema involving activation of bradykinin B(2) receptors, associated with enhancement of bradykinin B(1) receptor-mediated oedema. PMID- 11282126 TI - Agonistic activity of SR59230A at atypical beta-adrenoceptors in guinea pig gastric fundus and duodenum. AB - We have recently suggested that atypical beta-adrenoceptors are present in guinea pig gastric fundus and duodenum. In the present study, we have shown that SR59230A (3-(2-ethylphenoxy)-1-[(1S)-1,2,3,4-tetrahydronaphth-1-ylamino]-(2S)-2 propanol oxalate), a selective beta(3)-adrenoceptor antagonist, possesses agonistic activities at atypical beta-adrenoceptors in these tissues. SR59230A caused concentration-dependent relaxations. However, (+/-)-propranolol (1 microM) did not affect SR59230A-induced relaxations. Pretreatment of with a combination of (+/-)-propranolol (1 microM) and the non-selective beta(1)-, beta(2)-, beta(3) and beta(4)-adrenoceptor antagonist, (+/-)-bupranolol (30 microM), significantly antagonized the relaxant effects induced by SR59230A. The results clearly indicate that SR59230A acts as an atypical beta-adrenoceptor agonist on guinea pig gastric fundus and duodenum. PMID- 11282128 TI - Eye-movement recording in freely moving animals. AB - A new method is described for precise recording of eye movements in freely moving animals using Hall-effect devices. This inexpensive system, of small size and low weight, allows the analysis of horizontal and vertical components of saccadic eye movements, optokinetic nystagmus, slow tracking movements, eye vergence, etc., in unrestrained animals. A set of Hall-effect devices mounted in the skull is used to sense variations in the position of high-power miniature magnets fixed to the eye sclera. The output of the Hall-effect devices is amplified by operational amplifiers and collected through an analog-to-digital converter to be displayed on-line in a personal computer and stored for later analysis by specific software. Some examples of simultaneous body- and eye-movement recordings obtained in freely moving goldfish in different experimental situations are presented. This method would be useful in the recording of eye and gaze movements under natural conditions and for behavioural studies in freely moving animals. PMID- 11282130 TI - Effects of stress during pregnancy on maternal behavior in mice. AB - The effects of stress experienced during pregnancy and raising stressed offspring on maternal behavior were investigated in Swiss-Webster mice. Dams were either stressed or not stressed during pregnancy, and raised either prenatally stressed or nonstressed cross-fostered pups. Maternal behaviors such as grooming, nursing, pup retrieval and maternal aggression were assessed during the first 4 days after birth. Nonstressed dams raising stressed pups and stressed dams raising nonstressed pups groomed and nursed their pups significantly less than did control dams (stressed dams raising stressed pups and nonstressed dams raising nonstressed pups). Nonstressed dams raising stressed pups were also the slowest to retrieve both the first and last pup in retrieval tests. Nonstressed dams raising nonstressed pups were significantly less aggressive than other dams. In contrast, stressed dams raising stressed pups exhibited high levels of nursing and grooming, retrieved their pups rapidly and were very aggressive towards an intruder. These results indicate that raising stressed pups, or experiencing stress during a pregnancy can have significant effects on maternal behaviors. Stressed dams raising stressed pups exhibit maternal care comparable to that of nonstressed dams raising nonstressed pups at least for nesting/nurturing behaviors, and show increased levels of aggression and pup retrieval. PMID- 11282129 TI - The circadian system of reptiles: a multioscillatory and multiphotoreceptive system. AB - Many parameters exhibited by organisms show daily fluctuations that may persist when the organisms are held in constant environmental conditions. Rhythms that persist in constant conditions with a period close to 24 h are called circadian. Although nowadays most research in this field is focused on the molecular and genetic aspects--and therefore mostly on two animal models (Drosophila and mouse) -the study of alternative animal models still represent a useful approach to understanding how the vertebrate circadian system is organized, and how this fascinating time-keeping system has changed throughout the evolution of vertebrates. The present paper summarizes the current knowledge of the circadian organization of Reptiles. The circadian organization of reptiles is multioscillatory in nature. The retinas, the pineal, and the parietal eye (and, possibly, the suprachiasmatic nuclei of the hypothalamus, SCN) contain circadian clocks. Of particular interest is the observation that the role these structures play in the circadian organization varies considerably among species and within the same species in different seasons. Another remarkable feature of this class is the redundancy of circadian photoreceptors: retinas of the lateral eyes, pineal, parietal eye, and the brain all contain photoreceptors. PMID- 11282131 TI - Locomotor activity changes following lipopolysaccharide treatment in mice: a multivariate assessment of behavioral tolerance. AB - To determine the effects of repeated, acute endotoxin exposure on locomotor behavior, male laboratory mice were injected intraperitoneally with lipopolysaccharide (LPS: 50, 100 or 200 microg/kg) or saline vehicle on experimental Days 1, 4 and 7. At 2 h after each treatment, locomotor activity was assessed in a nonnovel, automated open-field apparatus (Digiscan) for 30 min. On Day 1, all horizontal and vertical activity measures were significantly reduced to near zero values by each dose of LPS. Behavioral tolerance to LPS formed rapidly, as locomotor activity of the treated groups did not differ from the control group on Days 4 or 7. In a second study, mice were given LPS (50, 100 or 150 microg/kg ip) or saline vehicle on two test days, 28 days apart. Activity was assessed, 1 h after injection, in a novel open field on the first test day and in a nonnovel open field on the second test day. Significant locomotor activity decrements were readily apparent in LPS-treated mice only in the nonnovel open field. This latter finding indicates that environmental novelty mediates, at least partially, the locomotor-reducing effects of LPS in mice. PMID- 11282132 TI - Effects of adrenalectomy and hormone replacement on B6C3F1 mice fed a high-fat diet. AB - Bilateral adrenalectomy (ADX) causes decreased circulating leptin in both obese and lean mice. It remains unclear whether the decreased plasma leptin after ADX is due to decreased adipose tissue or is due to decreased circulating glucocorticoids. The present experiment was performed to test the hypothesis that the absence of glucocorticoids from circulation is sufficient to decrease circulating leptin. Sixty-four adult male B6C3F1 mice were individually housed and fed either Purina rat chow or an experimental diet. After 6 weeks, mice fed the experimental diet gained more weight than mice fed the control diet. Each dietary group was then subdivided into four groups: ADX with cholesterol replacement (ADX-CHOL), ADX with corticosterone (CORT) replacement (ADX+CORT), ADX with aldosterone (ALDO) replacement (ADX+ALDO), and sham operation (SHAM). Two days after surgery, mice were killed and exsanguinated and the carcasses were prepared for gravimetric analyses. Blood was collected and centrifuged and the plasma was assayed for leptin, CORT, and ALDO. Blood glucose was determined using whole blood taken before centrifugation. There was no difference in body weight due to ADX after 2 days. Mice fed the experimental diet had higher circulating leptin than those fed the control diet. The ADX+CORT groups (both experimental and control diets) had higher plasma leptin concentrations than the other groups. No differences were observed between ADX-CHOL and SHAM groups. These results suggest that circulating leptin is not directly controlled by glucocorticoids. The effect of ADX on circulating leptin reported by others may be the consequence of decreased adiposity. PMID- 11282133 TI - Rabbit classical eyeblink conditioning is altered by brief cerebellar cortical stimulation. AB - A pair of studies examined how cortical intracerebellar stimulation (ICS) affects eyeblink conditioning in the rabbit. Rabbits were implanted with chronic bipolar stimulating electrodes in the cell body layers of cerebellar lobule H-VI. Brief (40 ms) trains of intracranial stimulation (100 Hz, 250 microA) were delivered during training trials [forward pairings of a tone-conditioned stimulus (CS) with an air puff unconditioned stimulus (US)]. In Experiment 1, the onset of ICS varied randomly within sessions. US-onset-coincident ICS proved detrimental to the maintenance of conditioning [measured as the percentage of trials on which conditioned responses (CRs) were made] compared to ICS that ended 60 ms before US onset. Based on these findings, a second experiment compared a group trained with ICS consistently delivered at US onset to groups trained with ICS consistently delivered either at CS onset or between the two stimuli, as well as to unstimulated control subjects. Animals receiving CS- or US-coincident ICS learned slowest, whereas animals receiving middle stimulation learned more quickly than all other groups. In both Experiments 1 and 2, highly trained animals produced blinks in direct response to the stimulation. These data are discussed in terms of a new hypothesis concerning interactions between cerebellar cortex and the deep cerebellar nuclei during eyeblink conditioning--a rebound from inhibition hypothesis. PMID- 11282134 TI - A two-choice discrimination method to assess olfactory performance in pigtailed macaques, Macaca nemestrina. AB - Four pigtailed macaques were trained in a new two-choice olfactory discrimination method. They learned the initial task within 3 months, requiring a maximum of 900 trials. After the method was established, we investigated the olfactory threshold of three monkeys for the odors peanut, iso-amyl acetate, and n-pentanoic acid. The animals detected peanut odor in dilutions as low as 1:10000. They were able to perceive iso-amyl acetate up to a 30000-fold dilution (animals F1 and M2), respectively in a 30 Mio-fold dilution (animal M1). The sensitivity for n pentanoic acid ranged between a dilution of 1:30000 (F1), 1:100000 (M2), and 1:300000 (M1). A comparison with the thresholds of other species demonstrates that the olfactory sensitivity of pigtailed macaques is not necessarily inferior to that of species that are believed to have a very keen sense of smell, such as dogs and rats. The sensitivity for certain odors seems to reflect their biological relevance for the tested species. The fact that the threshold for peanut odor obtained in this study is lower than the one found in a previous study with pigtailed macaques using a multiple olfactory discrimination method indicates that the new two-choice discrimination method is a better candidate for the assessment of olfactory abilities in pigtailed macaques. PMID- 11282135 TI - Memory-enhancing effects of DHEAS in aged mice on a win-shift water escape task. AB - The steroid hormone, dehydroepiandrosterone (DHEA) and its sulfate (DHEAS) have been implicated in age-associated deficits in memory. Numerous studies have demonstrated the effectiveness of these neurosteroids to enhance retention and ameliorate the effects of various memory-blocking agents, but few studies have directly assayed their effects on memory in aged animals. The present study investigated the memory-enhancing effects of DHEAS in a win-shift (nonmatching-to sample) task in aged mice using water escape motivation. Sixteen CD-1 mice, 18-20 months old, were trained to a moderate criterion of 7/10 correct trials and were then divided into two equal groups based on acquisition performance. One group received oral administration of DHEAS (1.5 mg/mouse/day) in a vehicle solution (0.0015% methyl salicylate) while the other group received the vehicle alone. DHEAS effects were assessed using a procedure in which delay intervals (0, 120, and 240 s) were interposed between sample and comparison trials over the course of three test sessions. The group receiving DHEAS recorded significantly higher retention scores across 3 days of testing, particularly at the 120-s delay interval, indicating that DHEAS enhanced working memory in these aged animals. PMID- 11282136 TI - Hormonal control of birth behavior in the bandicoot (Perameles gunnii: Marsupialia) and other marsupials. AB - The barred bandicoot is a small marsupial, locally common on the island of Tasmania, Australia. Several marsupial species have been shown to respond to injections of prostaglandin F2alpha or oxytocin by demonstrating birth behavior but that if an inhibitor of prostaglandin synthesis was given prior to oxytocin, no birth behavior was demonstrated. The barred bandicoot has behaved in a similar manner, demonstrating a significant increase in grooming and three other behaviors (crouching/prone, lateral and 'birth') were only seen after injection of PGF2alpha or oxytocin. The fact that after injection of oxytocin there was an increase in the latency till a birth response confirms a previous finding in another species. Subsequently, we have repeated these experiments with other marsupial groups, with similar results, which suggest that prostaglandin F2alpha stimulates birth behavior and oxytocin stimulates prostaglandin production in most marsupials. PMID- 11282137 TI - Does previous protein feeding affect the response of sheep towards foods that differ in their rumen availability, but not content, of nitrogen? AB - The objective of the experiment was threefold: (1) to test whether an 'unlearned appetite' for dietary protein exists in sheep, (2) whether such an appetite and subsequent diet selection depend on the degree of previous protein deprivation, and (3) whether the N-source in the foods offered as choice influences diet selection. Differences in protein deprivation were achieved by feeding sheep with food either with high (HP) or low (LP) protein. Sheep were fed on Food LP, either for the same period of time as sheep on Food HP or until they reached the same live-weight (LW) as sheep on Food HP. Following the feeding regimes that induced differences in animal state, sheep were given a choice between a novel low protein food (T) or Food T supplemented with isonitrogenous amounts with one of three nitrogen sources: urea (U), casein (C), or formaldehyde-treated casein (TFC). Diet selection measured in the short-term or over the first few days did not provide any evidence in support of an unlearned appetite for protein by sheep of different states. In fact, diet selection by all animals was characterized by an avoidance of the foods supplemented with the three nitrogen sources. This avoidance was strongest for the food supplemented with formaldehyde-treated casein. Selection of considerable amounts of supplemented foods was gradual and consistent only after animals gained experience of them, i.e. were allowed to consume a single supplemented food for a period of 7 days. Following this period, animals that had previously consumed Food LP for the same period of time as animals on Food HP selected a higher proportion of the supplemented food than the other sheep. The results support the view that there is no unlearned appetite for protein in sheep, and that control over diet selection is learned. PMID- 11282138 TI - Adrenalectomy eliminates the extinction spike in autoshaping with rats. AB - Experiment 1, using rats, investigated the effect of adrenalectomy (ADX) on the invigoration of lever-contact performance that occurs in the autoshaping situation after a shift from acquisition to extinction (called the extinction spike). Groups of rats with ADX or sham operations were trained under spaced and massed conditions [average intertrial intervals (ITI) of either 15 or 90 s] for 10 sessions and then shifted to extinction. ADX did not affect acquisition training but it eliminated the extinction spike. Plasma corticosterone levels during acquisition were shown in Experiment 2 to be similar in rats trained under spaced or massed conditions. Adrenal participation in the emotional arousal induced by conditions of surprising nonreward (e.g., extinction) is discussed. PMID- 11282139 TI - Estrogen-induced suppression of intake is not mediated by taste aversion in female rats. AB - Estrogen treatment can suppress the intake of a previously presented gustatory conditioned stimulus (CS). This finding has been interpreted as an estrogen induced conditioned taste aversion. However, a distinction must be made between taste aversion and taste avoidance. In particular, tastes are only considered aversive if they elicit a stereotypic behavioral response, otherwise the reduction in intake is classified as an avoidance. Although aversive orofacial responses have been reported in male rats after taste-estrogen pairings, they have not been examined in ovariectomized female rats. The goal of the present investigation, then, was to use similar procedures to determine whether conditioned aversion also mediates the estrogen-induced reduction of intake in female rats. Animals were introduced to a novel 0.1% saccharin solution and immediately thereafter were given a subcutaneous injection of vehicle or estradiol benzoate (10 microg). Responses were assessed using a two-bottle preference test, a one-bottle acceptance test, and a taste reactivity (TR) test. The results confirmed previous reports of a reduced preference for saccharin after saccharin-estradiol pairing using the two-bottle test. The reduction in intake during the one-bottle test, however, was not accompanied by stereotypic aversive responses, such as gaping. Surprisingly, a similar reduction in intake also occurred when using a backward conditioning procedure in which estrogen was injected before, rather than after, CS access. Thus, the present results show that the suppressive effects of estrogen reflect an avoidance, rather than aversion and, moreover, that the reduced intake may be due to an unconditioned, rather than a conditioned, response. PMID- 11282140 TI - Effect of food restriction on acquisition and expression of a conditioned odor discrimination in mice. AB - Level of food restriction was manipulated in mice to assess its importance for the acquisition and expression of a conditioned odor discrimination. In training, animals were exposed to odors (either rose or lemon) presented on a piece of filter paper in a pot covered in bedding. For half of the conditioning trials, group paired received one odor (CS+) with sucrose, the unconditioned stimulus (us), under the bedding. For the remaining trials, they received the other odor (CS-) alone. Group CS-alone was also exposed to both odors, but neither odor was paired with sugar on any of the conditioning trials. During training, Group Paired mice that were food-restricted tended to dig more readily and longer in the odors, especially in the CS+ odor, than animals that were not restricted. Both restricted and nonrestricted PAIRED GROUPS dug more in the CS+ than in the CS- by the end of training, but the CS-alone mice dug very little in either. Following training, mice were exposed to both odors simultaneously in a discrimination test. Half the mice in each training food restriction condition were tested under food restriction, and half were not. Only PAIRED animals that were food-restricted in the test expressed an odor discrimination, digging only in the CS+. This occurred regardless of their previous restriction state in training. These data suggest that both food-restricted and nonrestricted mice can acquire an odor discrimination; however, expression of this odor discrimination depends on food restriction. PMID- 11282141 TI - Can conditioned histamine release occur under urethane anesthesia in guinea pigs? AB - Many clinical and experimental data have shown that learning can occur under general anesthesia. To clarify this possibility with respect to allergic reactions, particularly asthmatic responses, we first established classical conditioned histamine release in response to a neutral odor by using pairings of the odor and an inhaled antigen for five sessions (Experiment 1) and then investigated whether conditioned histamine release into the plasma, bronchoalveolar lavage fluid (BALF), and lung tissue, which followed such a conditioning procedure, would be produced in urethane-anesthetized guinea pigs in the presence or absence of antigen (Experiment 2). Ovalbumin (OA) was used as the unconditioned stimulus (US) and dimethylsulfide (DMS) served as the conditioned stimulus (CS) in both experiments. In Experiment 1, the plasma histamine levels in the conditioned group increased significantly more than those of the unpaired control group in response to the CS during consciousness. In Experiment 2 in the absence of antigen, however, no significant differences in the histamine levels were found regarding the groups (DMS, triethylamine, saline, or unsensitized) or the time course (before, immediately, 5 min, and 10 min after the inhalations) during anesthesia, except for the finding that the histamine levels in the lung tissue specimens from the DMS group were significantly higher than those from the triethylamine group. In Experiment 2 in the presence of antigen, there was a significant increase in the plasma histamine levels after exposure to the US, irrespective of the presence of the CS, however, no significant difference in the histamine levels was observed between the US and the CS+US groups. These results indicated that a classically CS might not induce asthmatic responses under anesthesia. PMID- 11282142 TI - Lateralization of ventral fins use during object exploration in the blue gourami (Trichogaster trichopterus). AB - Blue gourami fish have a pair of modified ventral fins that are used to obtain tactile information about surrounding objects. Use of ventral fins by blue gourami was investigated during initial exploration of novel objects. When exposed to a sequence of novel plastic objects, varying in shape and colour, the blue gourami showed preferential use of the left fin during initial contacts. Laterality apparently depends on the nature of the stimulus: Fish exposed to a randomized series of natural objects showed preferential use of the left fin for inanimate mineral objects, but no asymmetry was apparent for investigating animate objects. This would suggest that some form of 'handedness' may have been present prior to the appearance of tetrapods. On the other hand, measurements of fish monocular viewing revealed that the fin use was strongly associated with preferential use of the ipsilateral eye before the touching of the stimulus took place, thus, suggesting that the asymmetry in fin use may also be related to lateralization of the visual system. PMID- 11282143 TI - Pregnancy is associated with low fear reactions in ewes. AB - The aims of the study were: (1) to test the influence of pregnancy on responses of ewes to several fear-eliciting situations, (2) to compare the first and latest stages of pregnancy, and (3) to investigate possible correlations between fear reactions and progesterone levels. Fear reactions of nonpregnant (NP; N=22) and pregnant (P) Ile-de-France ewes (day 40 of pregnancy: N=43; day 140 of pregnancy: N=19) were compared during three situations classically reported to induce fear in sheep: isolation, surprise, and the presence of a human. P ewes displayed significantly lower fear reactions than NP ewes when isolated and when confronted with a surprise effect combined with the appearance of a novel object. This reduction in fearfulness may be mediated principally through reduced fear of isolation. However, fear of a human remained constant despite pregnancy. Fear reactions of ewes tested during isolation on gestation day 40 or 140 did not differ, suggesting that decreased fear is not restricted to the latest stage of pregnancy. A negative correlation was found between plasma progesterone levels and fear during isolation and surprise tests of ewes with low levels of progesterone. The decrease in fearfulness during pregnancy may have some adaptative value for the survival of the young. PMID- 11282144 TI - Long-term time estimation is influenced by circadian phase. AB - The adaptive significance of a putative time sense in humans remains unclear as do the factors that underlie the capacity to gauge the passage of time. Here we show that the subjective assessment of relatively long durations varies systematically as a function of time of day. Specifically, the subjective clock ran relatively faster when the circadian oscillation of body temperature was on the rise and relatively slower on the declining portion of the temperature curve. The overall result was a rather labile clock that, on average, ran slow relative to physical time. The results provide a glimpse into an underexplored aspect of how humans use their endogenous clocks in the most fundamental way--to gauge the passage of time. PMID- 11282145 TI - Odorant confusion matrix: the influence of patient history on patterns of odorant identification and misidentification in hyposmia. AB - The odorant confusion matrix (OCM) is an odorant identification test in which the number of correct odorant identifications quantifies the level of olfactory function. As with other confusion matrices, the OCM reflects distortions of sensory perception as errors in identification. Previous work with the OCM suggests that, within an individual, hyposmia is associated with a stable shift in odorant perception. The current study examined whether consistent shifts in odorant perception are also characteristic of the various pathologies that lead to an olfactory loss. In a retrospective study, OCM response patterns for 135 hyposmic patients were fit into a five-dimensional space in which the distances between subjects reflected the dissimilarities between their OCM response patterns. Multivariate regression was performed relating position in the five dimensional space to each of 11 factors representing 33 demographic and medical history variables. One factor, named congestion (gathering the variables of past polyposis, current polyposis, and current nasal obstruction due to swelling), was significantly indicative of patterns of responses on the OCM, independent of the level of hyposmia. These data suggest that conductive olfactory loss may be associated with alterations in odorant perception, which are reflected in consistent odorant confusions. Such alterations in perception may eventually serve as a basis for a clinical test to provide differential diagnoses as to the sources of olfactory losses. PMID- 11282147 TI - Description and evaluation of a Newton-based electronic appetite rating system for temporal tracking of appetite in human subjects. AB - This study assessed the reliability and validity of a palm-top-based electronic appetite rating system (EARS) in relation to the traditional paper and pen method. Twenty healthy subjects [10 male (M) and 10 female (F)] - mean age M=31 years (S.D.=8), F=27 years (S.D.=5); mean BMI M=24 (S.D.=2), F=21 (S.D.=5) - participated in a 4-day protocol. Measurements were made on days 1 and 4. Subjects were given paper and an EARS to log hourly subjective motivation to eat during waking hours. Food intake and meal times were fixed. Subjects were given a maintenance diet (comprising 40% fat, 47% carbohydrate and 13% protein by energy) calculated at 1.6xResting Metabolic Rate (RMR), as three isoenergetic meals. Bland and Altman's test for bias between two measurement techniques found significant differences between EARS and paper and pen for two of eight responses (hunger and fullness). Regression analysis confirmed that there were no day, sex or order effects between ratings obtained using either technique. For 15 subjects, there was no significant difference between results, with a linear relationship between the two methods that explained most of the variance (r(2) ranged from 62.6 to 98.6). The slope for all subjects was less than 1, which was partly explained by a tendency for bias at the extreme end of results on the EARS technique. These data suggest that the EARS is a useful and reliable technique for real-time data collection in appetite research but that it should not be used interchangeably with paper and pen techniques. PMID- 11282146 TI - Nutrient preference and diet-induced adiposity in C57BL/6ByJ and 129P3/J mice. AB - Purified carbohydrates and fats are usually palatable to humans and other animals, and their consumption often induces weight gain and accumulation of fat. In this study, we examined consumption of complex carbohydrates (cornstarch and Polycose) and fats (soybean oil and margarine) in mice from two inbred strains, C57BL/6ByJ and 129P3/J. At lower concentrations of liquid nutrients tested using two-bottle tests, when the amounts consumed had negligible energy content, the C57BL/6ByJ mice had higher acceptance of Polycose and soybean oil. This was probably due to strain differences in chemosensory perception of Polycose and oil. At higher concentrations, the mice consumed a substantial part of their daily energy from the macronutrient sources, however, there were no or only small strain differences in nutrient consumption. These small differences were probably due to strain variation in body size. The two strains also did not differ in chow intake. Despite similar energy intakes, access to the nutrients resulted in greater body weight (BW) gain in the C57BL/6ByJ mice than in the 129P3/J mice. The diet-induced weight gain was examined in detail in groups of 2-month-old C57BL/6ByJ and 129P3/J mice given ether chow, or chow and margarine to eat. Access to margarine did not increase total energy consumption of either strain. It increased BW and adiposity of the C57BL/6ByJ mice, but only after they reached the age of approximately 3 months. There were no differences in BW and adiposity between control and margarine-exposed 129P3/J mice. The results suggest that diet induced adiposity in the B6 mice depends on age and does not depend on hyperphagia. PMID- 11282148 TI - Perinatal dietary NaCl level: effect on angiotensin-induced thermal and dipsogenic responses in adult rats. AB - We have shown previously that administration of angiotensin II (Ang II) produces an apparent decrease in thermoregulatory set point. Exposure to high salt diets either perinatally or later in life has been shown to increase pressor responsiveness to administration of Ang II, so in the present studies we examine whether high dietary NaCl would also increase the thermal responsiveness to Ang II. In the first study, we show that exposure to a basal NaCl diet (0.12%) during gestation through 4 weeks postnatally produced very large elevations in plasma renin activity (PRA) and aldosterone concentrations in the offspring. Exposure to high salt diet (3%) did not decrease the levels of these parameters below those fed mid salt diet (1%). In the second study, we show that rats raised through 4 weeks of age on basal diet, but then fed standard chow until adulthood, showed greater changes in tail skin (T(sk)) and colonic (T(c)) temperatures following administration of Ang II (200 microg/kg sc) than either mid- or high-salt-raised groups. In the third study, we confirmed this finding and extended it to show that rats raised on a very high salt diet (6%) also did not differ from the mid salt group. In both studies, acute water intake measured in a separate test following administration of Ang II did not differ as a function of perinatal salt diet. In a fourth study, the period of exposure to the diets was extended from the perinatal period through adulthood and, surprisingly, there was no longer an enhanced thermal response to Ang II in basal diet rats compared with rats fed the very high salt diet. In the final study, rats raised on a regular diet but exposed only as adults to the test diets showed a nonsignificant trend toward a decreased thermal response in the basal group. Thus, dietary salt level may have opposite effects on Ang II effects on adult thermoregulation, depending on the age at the exposure. PMID- 11282149 TI - Multifocal motor neuropathy. AB - Multifocal motor neuropathy (MMN) is a recently identified peripheral nerve disorder characterized by progressive, predominantly distal, asymmetric limb weakness mostly affecting upper limbs, minimal or no sensory impairment, and by the presence on nerve conduction studies of multifocal persistent partial conduction blocks on motor but not sensory nerves. The etiopathogenesis of MMN is not known, but there is some evidence, based mostly on the clinical improvement after immunological therapies, that the disease has an immunological basis. Antibodies, mostly IgM, to the gangliosides GM1, and though less frequently, GM2 and GD1a, are frequently detected in patients' sera, helping in the diagnosis of this disease. Even if there is some experimental evidence that these antibodies may be pathogenic in vitro, their role in the neuropathy remains to be established. Patients with MMN do not usually respond to steroids or plasma exchange, which may occasionally worsen the symptoms, while the efficacy of cyclophosphamide is limited by its relevant side effects. More than 80% of MMN patients rapidly improve with high dose intravenous immunoglobulin therapy (IVIg). The effect of this therapy is, however, transient and improvement has to be maintained with periodic infusions. A positive response to interferon-beta has been recently reported in a minority of patients, some of whom were resistant to IVIg. Even if many progresses have been made on the diagnosis and therapy of MMN, there are still several issues on the nosological position, etiopathogenesis and long-term treatment of this neuropathy that need to be clarified. PMID- 11282150 TI - ATP and adenosine induce ramification of microglia in vitro. AB - Microglial cells in the healthy adult brain possess a characteristic ramified morphology with multiple branched processes, small somata and down-regulated inflammatory properties. In contrast, microglial cells isolated from new-born rat brain inevitably show a non-ramified amoeboid phenotype, which is observed in vivo after pathologic activation or during development. To identify factors that control microglial morphology we investigated the effects of purines alone or in combination with astrocyte-conditioned medium (ACM). Under optimized culture conditions postnatal rat microglial cells developed an amoeboid to ovoid phenotype. Addition of 0.6-1 mM ATP or adenosine induced the outgrowth of numerous processes after 2-3 days that could be observed also in the presence of ACM as previously reported. Culture in ACM plus ATP or adenosine yielded an optimized ramified phenotype. ATP or adenosine, but not ACM alone, also prevented the formation of a flat, amoeboid morphology induced by lipopolysaccharide (LPS); however, at 0.6-1 mM they did not reduce the initial LPS-induced activation of the transcription factor NF-kappaB. By using specific agonists or antagonists the morphological transformations could not be confined to a distinct purinoreceptor subtype, but appeared to be mediated by long-term presence of adenosine in the medium to which phosphorylated purines were rapidly hydrolyzed by microglial cells. Since ACM did not contain sufficient concentrations of ATP or adenosine, purines are not the only ramification-inducing factors present in ACM; however, they are a valuable tool to induce microglial ramification in vitro. PMID- 11282151 TI - The cyclopentone prostaglandin 15-deoxy-Delta(12,14) prostaglandin J2 represses nitric oxide, TNF-alpha, and IL-12 production by microglial cells. AB - Prostaglandins are generally considered pro-inflammatory molecules that contribute to the pathology associated with a variety of immune-mediated diseases including multiple sclerosis. However, recently it has been demonstrated that specific cyclopentone prostaglandin metabolites including 15-deoxy-Delta(12,14) prostaglandin J2 (15d-PGJ2) are capable of repressing the production of pro inflammatory molecules by cells of the monocyte/macrophage lineage. Activated microglia produce nitric oxide (NO) and TNF-alpha, molecules which can be toxic to cells including oligodendrocytes, thus potentially contributing to the pathology associated with multiple sclerosis. The current study demonstrates that 15d-PGJ2 inhibits lipopolysachharide (LPS) induction of NO and TNF-alpha production by rat primary microglia and mouse N9 microglial cells. 15d-PGJ2 also inhibits NO production by microglial cells activated in response to IFN-gamma and TNF-alpha, cytokines believed to be important modulators of multiple sclerosis. IL-12 plays a critical role in stimulating the production of Th1 cells, which are believed to contribute to the pathology associated with multiple sclerosis. The current studies demonstrate that 15d-PGJ2 represses the production of IL-12 by microglial cells. Collectively, these studies demonstrate that the prostaglandin metabolite 15d-PGJ2 represses microglial production of potentially cytotoxic molecules, as well as molecules capable of altering T-cell phenotype. These in vitro studies suggest the possibility that the prostaglandin 15d-PGJ2 may modulate inflammatory diseases including multiple sclerosis. PMID- 11282152 TI - Social stress increases the susceptibility to endotoxic shock. AB - The influence of social disruption stress (SDR) on the susceptibility to endotoxic shock was investigated. SDR was found to increase the mortality of mice when they were challenged with the bacterial endotoxin lipopolysaccharide (LPS). Histological examination of SDR animals after LPS injection revealed widespread disseminated intravascular coagulation in the brain and lung, extensive meningitis in the brain, severe hemorrhage in the lung, necrosis in the liver, and lymphoid hyperplasia in the spleen, indicating inflammatory organ damage. In situ hybridization histochemical analysis showed that the expression of the glucocorticoid receptor mRNA was down-regulated in the brain and spleen of SDR animals while the ratio of expression of AVP/CRH-the two adrenocorticotropic hormone secretagogue, increased. After LPS injection, the expression of pro inflammatory cytokines, IL-1beta and TNF-alpha, was found significantly higher in the lung, liver, spleen, and brain of the SDR mice as compared with the LPS injected home cage control animals. Taken together, these results show that SDR stress increases the susceptibility to endotoxic shock and suggest that the development of glucocorticoid resistance and increased production of pro inflammatory cytokines are the mechanisms for this behavior-induced susceptibility to endotoxic shock. PMID- 11282153 TI - Restraint stress elevates the plasma interleukin-6 levels in germ-free mice. AB - Several recent reports demonstrated that restraint stress elevates plasma IL-6 levels; however, the precise mechanism whereby stress stimuli trigger the production of IL-6 remains to be clarified. In this study, in order to elucidate whether or not the intestinal microflora contribute to the stress-induced IL-6 elevation, the plasma IL-6 response of germ-free (GF) mice, which are indeed devoid of indigenous microflora, was compared to that of specific pathogen-free (SPF) mice. The plasma IL-6 level increased after 1 h of restraint stress and thereafter gradually decreased in GF mice as well as in SPF mice. In addition, such a stress-induced IL-6 elevation was also found in the mice reconstituted with SPF feces. The expression levels of IL-6 mRNA in the liver increased after 1 h of stress in both GF and SPF mice based on the findings of a semiquantitative RT-PCR method, although no such increase was observed in the spleen and kidney of both groups of mice. These results thus indicate that restraint stress is capable of elevating the plasma IL-6 levels independently of the intestinal microflora and the liver is one of the main sources responsible for the increased plasma IL 6 during stress. PMID- 11282154 TI - Complement regulatory proteins and selective vulnerability of neurons to lysis on exposure to acetylcholinesterase antibody. AB - Systemic injection of antibodies against acetylcholinesterase (AChE) induces complement-mediated destruction of preganglionic nerve terminals in paravertebral sympathetic ganglia, but spares other AChE-rich structures, such as nerve terminals in prevertebral sympathetic ganglia, parasympathetic ganglia, and the neuromuscular junction. This pattern of differing sensitivity to "AChE immunolesion" might be explained by a differing expression of proteins that serve to protect host cells from complement activation. Two major complement regulatory proteins in rats are Crry, which interferes with the assembly of C3 convertase, and CD59, which blocks formation of the terminal cytolytic membrane attack complex. The present study used immunohistochemistry to demonstrate an inverse relation between levels of CD59 and Crry expression and sensitivity to AChE immunolesion in several AChE-rich targets. Thus, the most sensitive structures, i.e., preganglionic nerve terminals in the adrenal gland and superior cervical ganglion (SCG), expressed undetectable levels of CD59 and Crry immunoreactivities. By contrast, AChE-rich, but antibody-resistant, cholinergic nerve terminals in the inferior mesenteric ganglia (IMG) and diaphragm muscle expressed significant amounts of CD59 and Crry. Such expression was functionally important because, after membrane-anchored CD59 was removed from explanted IMG with phosphatidylinositol phospholipase C, exposure to AChE antibody and complement caused greater immunolesion. It was concluded that differential expression of regulatory proteins in different parts of the nervous system influences regional vulnerability to complement mediated damage. PMID- 11282155 TI - Changing the chemokine gradient: CINC1 crosses the blood-brain barrier. AB - Chemokines are a large family of small, inducible, secreted, chemoattractant cytokines that are involved in inflammatory processes. It is well known that systemic and CNS infections cause disruption of the blood-brain barrier (BBB); however, it is not clear how chemokines are involved in this process. We studied the pharmacokinetics of the passage of the chemokine cytokine-induced neutrophil chemoattractant-1 (CINC1) from blood to brain after i.v. bolus injection and its efflux out of the brain after i.c.v. injection. Radiolabeled CINC1 was injected i.v. into mice, and the results were determined by multiple-time regression analysis. Using HPLC, we detected intact CINC1 in brain homogenate and blood after i.v. administration. CINC1 accumulated in the cerebral vasculature but also crossed the BBB completely and rapidly. No saturation of the influx was found, suggesting that either CINC1 crossed the BBB by simple diffusion or the dynamic interactions of binding and internalization precluded the self-inhibition typical of a transport system. Furthermore, there was no efflux system, with CINC1 exiting the brain at the same rate as reabsorption of CSF. The CINC1 injected into blood or CSF did not cause any breakdown of the BBB during the course of the experiments. Thus, the influx of CINC1 may alter the "chemokine gradient" across the BBB and therefore affect inflammatory reactions involving the CNS. PMID- 11282156 TI - beta-Endorphin-containing memory-cells and mu-opioid receptors undergo transport to peripheral inflamed tissue. AB - Immunocyte-derived beta-endorphin can activate peripheral opioid receptors on sensory neurons to inhibit pain within inflamed tissue. This study examined mu opioid receptors (MOR) on sensory nerves and beta-endorphin (END) in activated/memory CD4(+) cells (the predominant population homing to inflamed tissue). We found an upregulation of MOR in dorsal root ganglia, an increased axonal transport of MOR in the sciatic nerve and an accumulation of MOR in peripheral nerve terminals in Freund's adjuvant-induced hindpaw inflammation. A large number of CD4(+) cells containing beta-endorphin, but very few naive cells (CD45RC(+)), were observed in inflamed tissue, suggesting that this opioid is mainly present in activated/memory cells (CD4(+)/CD45RC(-)). Taken together, our results indicate an enhanced transport of both MOR and of the endogenous ligand beta-endorphin to injured tissue. This unique simultaneous upregulation of both receptors and ligands may serve to prevent excessive and/or chronic inflammatory pain. PMID- 11282157 TI - Tolerance induction by acylated peptides: suppression of EAE in the mouse with palmitoylated PLP peptides. AB - Treatment of SJL mice either before or after challenge with palmitoylated PLP139 151 (PAL139-151) completely suppressed or considerably reduced both acute and relapsing stages of EAE induced with PLP139-151. In the presence of Pertussis toxin, treatment with PAL139-151 was less effective, but treatment with a mixture of PAL139-151 and PAL178-191, the palmitoylated PLP epitope to which T cell recognition spreads, resulted in almost complete protection. Proliferation of lymphocytes from treated mice were sharply reduced, and adoptive transfer of lymph node lymphocytes from treated mice to naive recipients resulted in the reduction of the acute phase of EAE and in delayed relapses following challenge. The results suggest that treatment with PAL139-151 leads to both anergy and the generation of regulatory cells. PMID- 11282158 TI - The glossopharyngeal nerve as a novel pathway in immune-to-brain communication: relevance to neuroimmune surveillance of the oral cavity. AB - Glossopharyngeal afferents may be the neural channel by which immune challenge of the posterior oral cavity conveys information to the brain. If this is the case, then bilateral transection of the glossopharyngeal nerves (GLOx) should disrupt this communication. Injection of lipopolysaccharide (LPS) or interleukin (IL) 1beta into the soft palate (ISP) of sham-operated rats induced a dose-related febrile response. GLOx significantly attenuated the febrile response induced by ISP injection of both LPS and IL-1beta. In contrast, GLOx did not affect the febrile response when LPS or IL-1beta were injected intraperitoneally, indicating that the effect of GLOx is not systemic. These results provide experimental evidence for a novel neural pathway for immune-to-brain communication. PMID- 11282159 TI - Involvement of substance P and central opioid receptors in morphine modulation of the CHS response. AB - Morphine administration prior to challenge with the antigen 2,4-dinitro fluorobenzene increases the contact hypersensitivity (CHS) response in rats. The present study extended these findings by showing that central, but not systemic, administration of N-methylnaltrexone antagonized the morphine-induced enhancement of the CHS response. The importance of the neuroimmune mediator substance P was shown via the attenuation of the morphine-induced enhancement following both systemic and topical administration of the NK-1 antagonist WIN51,708. Taken together, the findings of the present study provide new data showing that central opioid receptors and peripheral substance P are involved in the morphine-induced enhancement of the CHS response. PMID- 11282160 TI - Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) blockade enhances incidence and severity of experimental autoimmune neuritis in resistant mice. AB - Experimental autoimmune neuritis (EAN), an autoimmune inflammatory demyelinating disease of the peripheral nervous system, represents an animal model of the human Guillain-Barre syndrome. EAN can be induced by active immunization in several animals, including Lewis rats. In contrast, most strains of mice including the widely used C57BL/6 (B6) strain are reputedly resistant to the induction of EAN. In the present study, we demonstrate that in B6 mice, anti-CTLA-4 monoclonal antibody administration in conjunction with immunization with the P0 protein derived peptide 180-199 can induce clinical and pathological definite EAN. Upregulating effects of CTLA-4 blockade on initial and ongoing EAN are demonstrated. CTLA-4 blockade augmented cellular infiltration and enhanced demyelination in the target organ sciatic nerves as well as increased T cell proliferation in lymph node cells. Moreover, serum levels of IFN-gamma and IL-4 were increased. Thus, manipulation of CTLA-4/B7 costimulatory pathway by CTLA-4 blockade can promote autoreactivity and break the relative tolerance to peripheral autoantigen P0 in resistant B6 mice. PMID- 11282161 TI - Mice resistant to experimental autoimmune encephalomyelitis have increased thymic expression of myelin basic protein and increased MBP specific T cell tolerance. AB - The relationship between expression of the autoantigens in thymi and susceptibility to autoimmune disease was determined in the experimental autoimmune encephalomyelitis (EAE) model. In two different sets of MHC congenic strains of mice characterized by differential susceptibility to EAE, levels of expression of MBP were shown to be higher in the more resistant strain. These data raised the possibility that more central tolerance to MBP may occur in more resistant strains. Differential tolerance was then evidenced by a decrease in T cell responses to MBP 83-102 in the more resistant strains. Together, these data indicate that the list of non-MHC genes involved in susceptibility to autoimmune disease should include genes which regulate expression of autoantigens in thymi. PMID- 11282162 TI - Presentation by myoblasts of an epitope from endogenous acetylcholine receptor indicates a potential role in the spreading of the immune response. AB - It is generally considered that myoblasts are unable to prime naive T cell responses without help from professional antigen-presenting cells (APC). However, their ability to present endogenous antigens to previously primed T lymphocytes in the secondary phase of a T cell response has not been well studied. We show here that primary human myoblasts, when stimulated with IFNgamma to express class II MHC, can present an endogenous epitope, probably an acetylcholine receptor (AChR) peptide, to a CD4(+) AChR-specific T helper lymphocyte clone. Presentation leads to secretion of IFNgamma by the T cell clone and, in addition, killing of the myoblast. Our results suggest that, during the effector phase of the immune response, myoblasts could enhance the inflammatory response by presenting endogenous antigen, and thereby become targets for CD4(+) T lymphocyte-induced cytotoxicity; subsequent release of myoblast antigens could then lead to inter- and intra-molecular determinant spreading. PMID- 11282163 TI - In vivo neutralization of endogenous brain fractalkine increases hippocampal TNFalpha and 8-isoprostane production induced by intracerebroventricular injection of LPS. AB - Fractalkine is a chemokine widely and constitutively expressed in the brain and, as suggested by in vitro studies, it is involved in brain inflammatory responses. In this study, we have investigated the in vivo anti-inflammatory potential of fractalkine in a model of neuroinflammation induced by intracerebroventricular injection of lipopolysaccharide (LPS) in rats. LPS induces a rapid and acute production of the pro-inflammatory cytokine, TNFalpha, in hippocampus and cerebrospinal fluid (CSF), and an increase of 8-isoprostane levels, a marker of oxidative stress, in hippocampus. Although intracerebroventricular injection of fractalkine has no effect on TNFalpha and 8-isoprostane production, neutralization of endogenous fractalkine within the brain with a specific anti fractalkine antibody potentiates LPS effects. These data emphasize the involvement of constitutive brain fractalkine in the control of inflammatory reaction in CNS. PMID- 11282164 TI - Intracellular pathways involved in TNF-alpha and superoxide anion release by Abeta(1-42)-stimulated primary human macrophages. AB - In this study, the intracellular signal transduction pathways leading to the production of TNF-alpha and superoxide anions by amyloid-beta-stimulated primary human monocyte-derived macrophages was investigated. Using Western blotting and specific inhibitors it is shown that both ERK 1/2 and p38 MAPK signal transduction pathways as well as PKC are involved in the amyloid-beta-stimulated superoxide anion production. In contrast, only ERK 1/2 MAPK seems to be involved in TNF-alpha production: questioning the connection between PKC and ERK 1/2 activation. Our results suggest the use of ERK 1/2 MAPK inhibitors in the prevention of macrophage activation in the context of Alzheimer's disease. PMID- 11282166 TI - T lymphocytes conditioned with Interferon beta induce membrane and soluble VCAM on human brain endothelial cells. AB - Vascular cell adhesion molecule (VCAM)-1 plays a critical role in mediating inflammatory cell adhesion and migration. Factors regulating the expression of membrane (m)VCAM and its cleaved counterpart soluble (s)VCAM are poorly understood. We previously demonstrated that serum sVCAM levels are increased in multiple sclerosis (MS) patients treated with interferon beta 1b (IFNbeta1b), which correlated with a reduction in gadolinium enhancing lesions on magnetic resonance imaging. However, subsequent studies have shown that IFNbeta does not directly induce VCAM expression on endothelial cells. We demonstrate here that co culture with IFNbeta-conditioned T cells induces mVCAM on human brain endothelial cells (HBEC). Further, rapid shedding of sVCAM occurs, which mirrors the response after in vivo IFNbeta treatment. The VCAM induction is mediated partially through tumor necrosis factor (TNF)alpha and can be abrogated by sTNF receptor. VCAM could also be induced on astroglioma lines using IFNbeta-conditioned T cells, which suggests the effect is not specific for HBEC. Kinetic studies demonstrated an increase in the sVCAM to mVCAM ratio over time, which may contribute to the ultimate therapeutic effect of IFNbeta in patients. These data have important implications for understanding the events occurring at the blood brain barrier in vivo, and for determining the mechanism of action of IFNbeta in MS. PMID- 11282165 TI - Humoral and cellular immune responses to Copolymer 1 in multiple sclerosis patients treated with Copaxone. AB - Humoral and cellular immune responses were followed in multiple sclerosis patients treated with Copolymer 1 (Cop1, glatiramer acetate, Copaxone) who participated in three different clinical trials. All patients (130) developed Cop1 reactive antibodies, which peaked at 3 months after initiation of treatment, decreasing at 6 months and remaining low. IgG1 antibody levels were 2-3-fold higher than those of IgG2. The proliferative response of Peripheral Blood Mononuclear Cells (PBMC) to Cop1 was initially high and gradually decreased during treatment. Antibodies and T cell responses to MBP were low and did not change significantly during the treatment. The humoral and cellular immunological responses to Cop1 do not correlate with the side effects and do not affect its therapeutic activity. The preferential production of IgG1 over IgG2 antibodies may indicate that Th2 responses are involved in mediating the clinical effect of Cop1. PMID- 11282167 TI - Enhanced expression of fractalkine in HIV-1 associated dementia. AB - The CX(3)C chemokine fractalkine was found to be up-regulated in the brain during inflammatory processes. In this study, we tried to assess the role of fractalkine in HIV-1-associated dementia. Fractalkine expression is up-regulated in the brains of AIDS patients with HAD. Fractalkine immunoreactivity was mainly detected in astrocytes. In addition, fractalkine expression was found to be up regulated in cocultures of astrocytes and HIV-infected macrophages. This up regulation was dependent on cell-cell contact. We propose that fractalkine produced during interactions between astrocytes and HIV-infected macrophages plays a role in HAD by regulating the trafficking of monocytic cells in the brain parenchyma. PMID- 11282168 TI - Characterization of the human T cell response against the neuronal protein synapsin in patients with multiple sclerosis. AB - Although multiple sclerosis (MS) is considered primarily as a demyelinating disease, neuronal damage is abundant and correlates with the neurological deficit. Therefore, we investigated the frequency and characteristics of human T cells specific for synapsin-a neuronal protein highly conserved among species. Synapsin specific T cell responses were detected at a frequency similar to that of MBP specific T cells in MS patients, one patient with acute demyelinating encephalomyelitis (ADEM) and controls. Long-term T cell lines specific for synapsin exhibited a CD3(+), CD4(+), CD8(-) phenotype and produced high amounts of tumor-necrosis-factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) after antigen specific stimulation, whereas lymphotoxin (LT), interleukin-4 (IL-4) and interleukin-10 (IL-10) were detectable in smaller quantities. PMID- 11282169 TI - Expression of inducible nitric oxide synthase, interleukin-1 and caspase-1 in HIV 1 encephalitis. AB - Inflammatory cytokines and enzymes such as IL-1 and inducible nitric oxide synthase (iNOS) may play an important role in the pathogenesis of AIDS dementia, a condition associated with infection of the CNS cells by the HIV-1. In this report, we investigated the expression of iNOS, IL-1, and caspase-1 (interleukin 1 converting enzyme) in HIV-1 encephalitis (HIVE) by immunocytochemistry and analyzed their expression with respect to HIV-1 infection and glial activation. In HIVE, all three molecules were expressed at high levels in areas of HIV-1 infection (microglial nodules with HIV-1 p24 immunoreactivity) and in areas of diffuse white matter gliosis. Expression was cell-type specific, with IL-1 and caspase-1 being expressed in macrophages and microglia, and iNOS in activated astrocytes. Multinucleated giant cells, a hallmark of virally infected cells, showed intense staining for both IL-1 and caspase-1, suggesting induction of these molecules by HIV-1. Double immunocytochemistry demonstrated a regional co localization of astrocyte iNOS and microglial IL-1 and caspase-1. These results support the notion that autocrine and paracrine interactions between HIV-1 infected macrophages and microglia, activated microglia, and astrocytes lead to expression of proinflammatory and neurotoxic molecules. iNOS and caspase-1 may provide additional therapeutic targets for HIVE. PMID- 11282170 TI - Serum and CSF levels of MCP-1 and IP-10 in multiple sclerosis patients with acute and stable disease and undergoing immunomodulatory therapies. AB - The two chemokines, monocyte chemoattractant protein (MCP)-1 and gamma-interferon inducible protein (IP)-10, are thought to be involved in the pathogenesis of multiple sclerosis (MS). We measured MCP-1 and IP-10 levels in serum and CSF samples from 38 acute and 25 stable MS patients and from 40 controls. The latter consisted in patients with other inflammatory neurological diseases (OIND) or with non-inflammatory neurological diseases, and healthy controls. CSF MCP-1 levels exceeded those found in serum in all the patients studied as well as in healthy controls. CSF MCP-1 levels were significantly lower in acute MS [468+/ (S.E.M.) 18 pg/ml] than in stable MS (857+/-104 pg/ml). When detectable, serum and CSF IP-10 levels were significantly higher in acute MS (serum 331+/-66 pg/ml; CSF 118+/-16 pg/ml) than in stable MS (serum 69+/-7 pg/ml; CSF 25+/-2 pg/ml). Among OIND patients, those with HIV-1-associated dementia showed high serum and CSF levels of both MCP-1 and IP-10. Those with encephalitis showed high serum and CSF levels of IP-10 and CSF mononuclear pleiocytosis. We also evaluated the effects of 6-methylprednisolone or IFN-beta1a therapy on circulating MCP-1 and IP 10 levels. Neither MCP-1 nor IP-10 post-therapy levels varied significantly from baseline values. Our findings suggest that (a) MCP-1 could be constitutively produced within the brain; (b) MCP-1 and IP-10 CSF levels in acute MS vary significantly from those in stable MS, and these variations are inverse; and (c) current MS therapies do not modify circulating levels of MCP-1 and IP-10. PMID- 11282172 TI - Taenia solium: germinal cell precursors in tapeworms grown in hamster intestine. AB - BACKGROUND: In a previous study, it was shown that growth of evaginated metacestodes occurs in the germinative tissue of the neck by duplication of somatic stem cells. In these specimens, it was not possible to find the mitotic figures required to demonstrate duplication of germ cell lines. METHODS: Taenia solium strobilae were collected from the intestinal lumen of outbred hamsters infected orally with 10 metacestodes dissected from naturally infected pigs. Animals were anesthetized 1-10 days postinfection, the small intestine excised, submerged in PBS, and cut open longitudinally. Live Taenias were incubated for 6 8 h in medium containing colchicine or 3H-thymidine, washed, and embedded for electron microscopy. For light microscopy and autoradiography, longitudinal sections were cut from whole blocks and mounted on glass slides. A population of large cells without nuclear membranes and containing discrete aggregates of chromatin were observed apposed to myofibrils in the germinative tissue of the neck. These cells were confirmed by electron microscopy as metaphase mitotic figures, with chromosomes attached to a microtubular spindle, embedded in cytoplasm, without a nuclear membrane, and with characteristic centrioles. RESULTS: Only tapeworms in which 3H-thymidine was injected directly into the worm tissue by microsyringe were positive by autoradiography, demonstrating that in contrast to evaginated metacestodes, intestinal worms do not transport thymidine across the tegument. CONCLUSIONS: The results show that differentiating T. solium worms have a subset of stem cells that require passage through a mammalian host to go into mitosis, and that tapeworms grown in an experimental animal do not take up 3H-thymidine in vitro. PMID- 11282171 TI - Decreased expression of c-myc family genes in thymuses from myasthenia gravis patients. AB - The thymus is a critical organ for the elimination of autoreactive T cells by apoptosis. We studied the expression of apoptosis-associated genes, bcl-xL, bad, caspase-3, and c-myc family genes in myasthenia gravis (MG) thymuses. We observed that the mRNA levels of myc family genes, c-myc and max, were markedly reduced in MG thymuses. These results indicate that c-myc-mediated signaling is abnormal in MG thymuses. The levels of molecules whose expressions are associated with myc, such as STAM, prothymosin-alpha, and NFkappaB, were also analyzed. PMID- 11282173 TI - Policosanol modulates HMG-CoA reductase activity in cultured fibroblasts. AB - BACKGROUND: Cholesterol biosynthesis is strictly controlled by 3-hydroxy-3 methylglutaryl Coenzyme A (HMG-CoA) reductase. METHODS: Transfer of cultured fibroblasts to a lipid-depleted medium (LDM) up-regulates the enzyme levels. This, in turn, is followed by an accelerated biosynthesis of cholesterol. RESULTS: Exposure of Vero fibroblasts to LDM and policosanol (0.5-50 microg/mL), a new cholesterol-lowering drug purified from sugarcane (Saccharum officinarum L.) wax, decreased in a dose-dependent manner cholesterol biosynthesis from [14C] acetate and 3H-water, but not from [14C]-mevalonate. CONCLUSIONS: This suggests an effect on HMG-CoA reductase, the rate-controlling enzyme in cholesterol biosynthesis. When enzyme activity was measured in the presence of various concentrations of policosanol (0.5-50 microg/mL), reductase was not suppressed. Therefore, there was no evidence for a competitive or noncompetitive inhibition of enzyme activity. However, after treatment of intact cells with policosanol (50 microg/mL) in the presence of LDM, a suppressive effect on enzyme activity was observed, suggesting a modulatory effect of policosanol on reductase activity. The previous inhibition of enzyme up-regulation by policosanol suggests to date a depression of de novo synthesis of HMG-CoA reductase and/or stimulation of its degradation. However, the exact mechanism by which policosanol inhibits the activity of HMG-CoA reductase still remains unclear. Further studies are needed to clarify the precise mechanism of its inhibitory action on cholesterol biosynthesis. PMID- 11282174 TI - Effect of caffeine on antinociceptive action of ketoprofen in rats. AB - BACKGROUND: To assess a possible synergistic antinociceptive interaction, the antinociceptive effects of ketoprofen (KET), and caffeine (CAF) administered either separately or in combinations were determined in a model of arthritic pain. METHODS: Antinociceptive activity was assayed using "ellipsis pain-induced functional impairment in the rat" (PIFIR model). The antinociceptive efficacies were evaluated using several dose-response curves and time courses. The antinociceptive effects from the combination that produced the greater effect were compared with the maximal antinociceptive effect of either morphine, acetylsalicylic acid (ASA), or KET alone. The animals were administered with 0.05 mL intra-articular (i.a.) of uric acid to induce nociception. Groups of six rats received orally either ASA, morphine (MOR), KET, CAF, or a combination KET + CAF (24 combinations). RESULTS: ASA (ED(50) 465.2 +/- 1.5 mg/kg), MOR (ED(50) 71.0 +/ 1.6 mg/kg), and KET (ED(50) 7.2 +/- 1.4 mg/kg) alone induced dose-dependent antinociception, whereas CAF alone showed no activity at the assayed doses. Nine combinations showed various degrees of potentiation (p <0.01), while the remainder exhibited the antinociceptive effect of KET only. Combinations of 17.8 mg/kg CAF with either 1.0, 1.8, 3.2, 5.6, or 10.0 mg/kg KET yielded the highest antinociceptive potentiations. For example, antinociceptive effect was 125.6 +/- 21.4 area units (au) with KET (3.2 mg/kg) alone, but the combination with CAF (17.8 mg/kg) showed 309.5 +/- 10.3 au. The median effective dose (ED(50)) of KET alone was 7.2 +/- 1.4 mg/kg, whereas the ED(50) of KET + CAF 17.8 mg/kg was 0.4 +/- 0.6 mg/kg: KET in the presence of CAF was approximately 18 times more potent than the analgesic drug without CAF. CONCLUSIONS: These results showed that CAF was able to potentiate the analgesia of KET, but only at selected dose combinations: CAF in the doses of 10.0 and 17.8 mg/kg was able to potentiate the analgesic effect of KET, the most efficacious drug combination being CAF 17.8 mg/kg + KET 3.2 mg/kg. The combination of analgesic drugs and CAF can produce better antinociceptive effects than the analgesic drug alone. This knowledge will permit the selection of the therapeutically most effective combination ratio of drugs, employing lower doses of each drug. PMID- 11282175 TI - Repeatability of heart rate variability during simple cardiovascular reflex tests on healthy subjects. AB - BACKGROUND: Our purpose was to determine the repeatability, after 2 weeks, of frequency domain measures of heart rate variability (HRV) during simple cardiovascular reflex tests. METHODS: Twenty healthy volunteers aged 29.3 +/- 2.5 years were assessed twice (at weeks 0 and 2). Continuous electrocardiogram and minute-to-minute blood pressure were recorded during spontaneous and metronome paced breathing (0.2 Hz). Under paced breathing, two tests were performed: 1) active change of posture: 5 min supine position, 5 min seated upright, and 5 min standing up, and 2) cold pressor test: the right hand was immersed in cold water (5 degrees C) for 2 min. RESULTS: Paced breathing elicited a significant increase of the high-frequency (HF) component of HRV. This effect was repeatable on 95% of the subjects. Active change of posture induced a significant increase of the heart rate with an increase of the low-frequency/high-frequency ratio of HRV. Although repeatability was better for the response to being seated upright than for the response to being standing up, it was always higher than 90%. The cold pressor test induced a significant increase of the heart rate and blood pressure, but with variable changes on the HRV measurements (either a decrease or an increase). Repeatability of responses was evident for 95-100% of the subjects. Although repeatability of HRV measurements in the same subject during the tests was higher than 95%, coefficients of repeatability reflected large differences among the subjects. CONCLUSIONS: The results suggest that it is not advisable to use isolated HRV changes to interpret the response to simple cardiovascular reflex tests in groups of healthy subjects. PMID- 11282176 TI - Levels of myosin heavy chain fragment in patients with tissue damage. AB - BACKGROUND: Myosin heavy chain fragments (MHC) levels are observed to be higher in myoskeletal injuries after surgery. MHC could be a helpful supplementary tool in the study of myoskeletal injuries. METHODS: Serum levels of myosin heavy chain fragments (MHC) were assessed in orthopedic patients before operation (OBO) and after operative (OAO) repairs and in the early phase of soft tissue injury (STI) using a radioimmunoassay involving monoclonal antibodies to the human beta-type MHC. RESULTS: Mean (SD) microU/L of MHC in comparison with the control subjects (75.3 +/- 47.1) was higher in OAO (305.8 +/- 38.1) p <0.0001, and no significant changes in MHC were found in STI (67 +/- 77.5). Myoglobin was notably higher in OBO (81.9 +/- 95.0) compared to STI (43.9 +/- 55.9) or controls p <0.05, but there was no further change in the protein after surgery. The mean proportional raised level of myoglobin in OBO was >twofold, and MHC increased by 27%. Neither myoglobin nor MHC increased in the plasma of the STI within 24 h of injury. CONCLUSIONS: These data suggest that the release of MHC could be a helpful supplementary tool in the study of tissue damage in humans. PMID- 11282178 TI - Dopamine D4 receptor (DRD4) gene polymorphism in the first psychotic episode. AB - BACKGROUND: Dopamine D4 receptor (DRD4) has shown some interesting properties at genetic and possibly functional levels. It has been suggested that some molecular variants of the DRD4 gene (e.g., four and seven alleles) could be implicated in the pathogenesis of psychotic disorders. Additionally, the VNTR polymorphism could be implicated in part of the response to treatment with neuroleptics. This study tested the possible association between the 48-bp tandem repeats in exon 3 of the DRD4 gene and patients experiencing their first psychotic episode. METHODS: Patients with a first psychotic episode (FPE, n = 37) were diagnosed and compared with a matched control group (n = 37). The FPE group was subdivided into two categories: those with nonaffective and those with affective psychoses. The variable number of tandem repeats (VNTR) region of the DRD4 gene was amplified by PCR procedures. Chi-square statistics and appropriate corrections and adjustments were used for data analysis. CONCLUSIONS: A significantly lower frequency of the four repeat (4-R) carriers in the FPE group was observed. This association was sustained mainly by the affective psychotic group (chi2 = 9.99 df = 2, p = 0.0073). Although these results require testing with stringent methods, it is suggested that the DRD4-4R allele may confer some protection against psychosis, mainly of the affective subtype. PMID- 11282177 TI - Efficacy of human recombinant DNase in pediatric patients with cystic fibrosis. AB - BACKGROUND: Most respiratory complications in cystic fibrosis (CF) arise from abnormally viscid mucus, and rhDNase has shown to be effective in enhancing mucous clearance. We explored the responses to rhDNase in a Mexican population of CF patients. METHODS: Patients with CF received aerosolized rhDNase (2.5 mg daily) during 3 months, followed by daily aerosolized placebo during 3 months. RESULTS: A total of 21 CF patients entered the study (11.1 +/- 0.5 years of age, mean +/- SEM, 10 girls): 15 patients (71%) had basal forced vital capacity (FVC) higher than the 70% predicted value, and the remainder of the patients had an FVC of between 30 and 70%. As a group, rhDNase progressively increased the forced expiratory flow at 1 sec (FEV1) as well as the FVC, reaching statistical significance (p <0.005) at the end of the third month of treatment. Sputum production and difficulty to expectorate or to breathe also improved during the rhDNase treatment period (p <0.05 to p <0.001). All these changes progressively decreased to basal values after 3 months with aerosolized placebo. Adverse reactions were almost null, with a sole patient reporting dysphonia. CONCLUSIONS: Aerosolized rhDNase was effective in progressively improving respiratory function and symptoms in most CF patients. PMID- 11282180 TI - Left ventricular diastolic dysfunction secondary to hyperglycemia in patients with type II diabetes. AB - Diabetes mellitus type II, a cause of preclinical left ventricular dysfunction that can progress to cardiac insufficiency ventricular dysfunction in diabetic patients, is attributed to systemic arterial hypertension, or ischemic cardiopathy. Diastolic ventricular dysfunction takes place during the course of diabetes mellitus. The purpose of the present article is to report on the influence of hyperglycemia on the left ventricular diastolic dysfunction independently of dyslipidemia, obesity, and systemic arterial hypertension, usually present in diabetic patients. Left ventricular diastolic function was studied by Doppler echocardiography in asymptomatic type II diabetic patients without ischemic or valvular cardiopathies, cardiomegaly, or systemic arterial hypertension. Two groups of patients were integrated: patients with and without left ventricular diastolic dysfunction, i.e., groups A and B, respectively. Glycemia, cholesterol, triglycerides, and body mass index (BMI) were determined in each subject. Bivariate statistical tests (Student t, chi-square, or Mann Whitney U tests) were applied to study the influence of the previously mentioned variables on the ventricular diastolic function. To evaluate the influence of hyperglycemia on ventricular diastolic function separately from dyslipidemia, systemic arterial hypertension, and the influence of obesity, logistic regression, and multivariate statistical analysis were applied. Independently of dyslipidemia and obesity, a relationship was found between hyperglycemia and diastolic dysfunction of the left ventricle in patients belonging to group A (p <0.05, odds ratio [OR] 12.1). No statistical significance was found between glycemia and the diastolic function of the left ventricle in group B patients. Even in type II diabetic patients without cardiopathy, uncontrolled hyperglycemia provokes diastolic left ventricular dysfunction. PMID- 11282179 TI - Persistence of Trypanosoma cruzi in chronic chagasic cardiopathy patients. AB - BACKGROUND: Although patients with chronic chagasic cardiopathy do have a strong immune response against Trypanosoma cruzi, they have transient and low parasitemia as well as tissue amastigote nests. When conventional studies were carried out, demonstration of such abnormalities is minimally achieved. Molecular biology may provide the best tools to demonstrate parasite persistence, which could be pathogenic in this progressive disease. METHODS: We studied 16 patients with chronic chagasic cardiopathy (CCC) at the Instituto Nacional de Cardiologia Ignacio Chavez in Mexico City. Patients had undergone a complete clinical evaluation, and had antibodies against Trypanosoma cruzi. They came from different rural areas in Mexico. Blood samples were obtained and processed for hemoculture and PCR technique. A CCC necropsy case was also sought for the presence of parasite antigen or DNA, using immunohistochemistry and PCR methods in archival tissues. RESULTS: Five of 16 (31%) hemocultures demonstrated circulating T. cruzi; 60% occurred in persons between 25 and 40 years old. In contrast, we found a positive PCR amplification in 81%; therefore, molecular biology tools appear to be more sensitive for demonstrating parasite persistence. There were no correlations between parasitemic state and clinical findings or specific antibody titer. The autopsy case had parasite antigens and DNA in heart tissues. CONCLUSIONS: Chronic chagasic cardiopathy patients do have persistence of parasite even when parasitemia is low or absent. The continuous presence of a parasite load could maintain immune stimulus and perhaps enhance a pathogenic immune or autoimmune tissue damage in susceptible hosts. PMID- 11282181 TI - Fetal amniotic adhesions. Their topographic concordance with regionally clustered malformations. AB - BACKGROUND: The amniotic band disruption complex (ABDC) has been segregated recently into various phenotypes. In view of the pathogenic mechanisms that have been proposed, this study was designed to assess if it is one variable process or is composed of several distinct complexes. METHODS: The 48 cases of fetuses with bands or placenta attached to fetal parts cited in this paper included nine new cases and 39 from the literature. They were organized first according to the embryonal topography of the malformations, then according to the position of the adhesions, and finally by the assessment of distances between the cases and between the malformations using the squared Euclidean distances for binary variables and cluster analysis. RESULTS: In all three analyses, three groups were identified: 1) fetuses with cephalo-thoracic anomalies; 2) fetuses with caudal anomalies, and 3) fetuses with mixed anomalies. Nonetheless, overlap among the three groups was apparent. Thus, while fetuses with amniotic bands form three clusters, it appears that these are part of a spectrum and should be considered as variable manifestations of a single entity resulting from a single pathogenetic mechanism. An association was established between the localization of the adhesions and the malformations in various axes. Abdominoschisis, however, was not particularly related to adhesions at one or the other end of the fetus; a short umbilical cord was an almost universal finding. Single umbilical artery (SUA) was especially related to caudal adhesions and malformations (p = 0.004 and 0.001), as well as abdominoschisis (p = 0.002) and agenesis of the abdominal organs (p = 0.008). CONCLUSIONS: The association between amniotic adhesions to the fetus and multiple malformations occurring predominantly in the same area suggest that the former are the cause of the latter. The association of abdominoschisis, as well as a short umbilical cord, with malformations and adhesions in all areas, suggests that these are secondary phenomena to generalized embryonal and fetal tension. SUA, however, with a specifically regional association, is more likely to be due to disruption from exposure in cases with abdominoschisis, often accompanying the loss of abdominal organs. PMID- 11282182 TI - Clinical manifestations and survival trends during the first 12 years of the AIDS epidemic in Mexico. AB - BACKGROUND: Our objective was to evaluate survival trends (1984-1995), the prevalence of AIDS-defining conditions, and the role of treatment with zidovudine and/or prophylaxis with trimethoprim-sulfamethoxazole (TMP-SMX) in survival following AIDS diagnosis. METHODS: We reviewed the clinical charts and postmortem studies of all patients admitted to the HIV Clinic from 1984-1995. Three groups were identified according to the following dates of HIV diagnosis: 1) 1984-1988; 2) 1989-1992, and 3) 1993-1995. RESULTS: We studied 909 charts. During the study period, 744 (81.6%) patients developed AIDS. Median survival increased from 11.7 months in group 1 to 15.4 and 17.5 months in groups 2 and 3, respectively (p <0.05). We observed the following important changes in the frequency of AIDS defining conditions over the study period: Pneumocystis carinii pneumonia (PCP) decreased from 24.8 to 17 and 14% in groups 1, 2, and 3, respectively, (p = 0.008), and Kaposi's sarcoma (KS), from 31.1 to 10.5 and 13.5% (p <0.001). On the other hand, there was an increase in cytomegalovirus disease with 12.4, 20.4, and 18.6% (p = 0.04) and wasting syndrome with 36, 45, and 57% (p <0.001). In the proportional hazard model for death, zidovudine or TMP-SMX use was associated with a protective effect. CONCLUSIONS: Survival is improving among patients with HIV infection at our institution. The prevalence of AIDS-defining conditions has changed over the last 12 years. There has been a diminution of PCP and KS, whereas cases of CMV disease and wasting syndrome increased. PMID- 11282183 TI - Neonatal outcome of children born to women with tuberculosis. AB - BACKGROUND: As the incidence of tuberculosis (TB) has increased worldwide, it is expected that pregnant women will acquire this infection more frequently. Mycobacterium tuberculosis infection during pregnancy may represent a risk for maternal and neonatal complications. METHODS: We studied the perinatal events of 35 consecutive pregnancies complicated by TB from March 1990 to June 1998; 105 apparently healthy pregnant women were included as controls, matched in age, gestational age upon arrival at the Institute, and socioeconomic status. Frequency and type of neonatal complications were recorded. Relative risk (RR) with 95% confidence interval (CI) was calculated. To control potentially confounding variables, a stratified analysis was performed. RESULTS: Seventeen (48.5%) tuberculous mothers had a pulmonary infection and 18 (51.5%), an extrapulmonar localization of the TB. The neonatal morbidity rate in children born to women with TB was 23% against 3.8% of the children of the control cohort (p <0.05). Average weight of newborn infants of tuberculous mothers was 2,859 +/- 78.5 g, while average weight at birth of control neonates was 3,099 +/- 484 g (p = 0.03). Newborns of women with TB had a higher risk of prematurity (RR 2.1; 95% CI 1-4.3), perinatal death (RR 3.1; 95% CI 1.6-6), and weight at birth less than 2,500 g (RR 2.2; 95% CI 1.1-4.9). Pulmonary localization of the TB and late start of the treatment in the mothers increase the risk of perinatal death and neonatal morbidity. CONCLUSIONS: Children born to women with TB have an increased risk of morbidity and mortality in the neonatal period. PMID- 11282184 TI - Marginal vitamin and mineral intake of Costa Rican adolescents. AB - BACKGROUND: Although a great deal of attention is given to macronutrient and energy intake in the diet, elements essential to dietary metabolic balance include important micronutrients. Very little information exists on vitamin and mineral intake during adolescence, especially in developing countries. METHODS: The micronutrient intake of urban and rural adolescents aged 12-19 years from the Costa Rican capital city, San Jose, was evaluated. Prospective 3-day diet records including 2 weekdays and 1 weekend day were used for dietary data. We evaluated the micronutrient intake to determine the nutrient adequacy ratio. RESULTS: Approximately 30% of adolescents did not meet the 70% level of the daily recommended intake (DRI) for vitamin Bl2. Additionally, 40% of the Costa Rican youngsters presented a vitamin A, E, and iron intake of between 30 and 69% of the DRI or recommended daily dose (RDA) for these nutrients. Likewise, approximately 15% of adolescents did not meet 30% of the DRI or RDA indicators for these nutrients. The micronutrients most at risk for inadequate intake were zinc, calcium, magnesium, and folate: more than 25% of the adolescents did not meet 50% of DRI or RDA indications for these micronutrients. Contrariwise, vitamin C, vitamin B6, thiamin, riboflavin, and niacin were the nutrients less at risk for inadequate intake. Fast foods prepared in school cafeterias provided approximately 15-30% of the DRI for vitamins B1, B2, B6, Bl2, and niacin, approximately 18% of the RDA for iron, and over 40% of the RDA for vitamin E. CONCLUSIONS: Nutritional interventions and educational strategies are needed to promote the adoption of healthful eating habits among adolescents. PMID- 11282185 TI - Prevalence of intimate partner abuse among nurses and nurses' aides in Mexico. AB - BACKGROUND: Nurses are the health professionals most frequently involved in the diagnosis and treatment of victims of family violence (FV). Understanding their personal experience with victimization is the key to shaping an appropriate role as advocates for medical recognition of FV and as integral members of the screening teams. We sought to determine the lifetime prevalence of intimate partner abuse among them and identify its risk factors. METHODS: In our cross sectional study, 1,150 registered nurses and nurses' aides at 11 urban hospitals in Mexico City self-administered an anonymous survey. We calculated descriptive statistics, Fisher exact tests, and multivariate logistic regression models to analyze physical, sexual, and emotional abuse during adulthood. RESULTS: Physical/sexual abuse during adulthood was 13% for nurses' aides and 18% for nurses. Similar proportions (13% of nurses' aides and 14% of nurses) also reported childhood physical/sexual abuse. Additional respondents (39% nurses' aides, 42% nurses) reported emotional abuse during adulthood. Detecting no significant differences in abuse patterns between the two groups, we combined occupations for all subsequent analyses. Being separated or divorced (vs. married) (Apr = 3.41, 95% confidence interval (CI): 1.81-6.44) and having suffered physical/sexual abuse during childhood (Apr = 3.39, 95% CI: 2.26-5.08) were associated with physical/sexual abuse in adulthood. The same variables were associated with adult emotional abuse (separated/divorced: Apr = 5.33, 95% CI: 2.61-10.85, and childhood physical/sexual abuse: Apr = 2.58, 95% CI: 1.79-3.75). Younger women (between the ages of 23 and 28 years) reported more emotional abuse (Apr = 2.10, 95% CI: 1.48-2.98). CONCLUSIONS: Counseling for abused nursing staff may help break the cycle. Physical/sexual partner abuse among nurses appears lower than among the general Mexican population, but remains worrisome. Battling childhood abuse might prevent intimate partner violence. PMID- 11282186 TI - Vaccination with recombinant modified vaccinia virus Ankara protects against measles virus infection in the mouse and cotton rat model. AB - Modified vaccinia virus Ankara (MVA) has been used as an experimental vaccine vector against respiratory infections. We have tested the safety and immunogenicity of a recombinant virus expressing the hemagglutinin of measles virus (MVA-MV-H) using the mouse model of measles virus induced encephalitis and the cotton rat model for respiratory infection. MVA-MV-H proved to induce a TH1 response, neutralizing antibodies and conferred protection against both encephalitis and lung infection. The cotton rat is very sensitive to infection with replication competent vaccinia virus. In these animals MVA-MV-H proved to be a very well tolerated vaccine. However, the efficiency in the presence of MV specific maternal antibodies was low (even using a prime-boost strategy) and therefore might have to be improved. PMID- 11282187 TI - Pathogenesis of RSV lower respiratory tract infection: implications for vaccine development. AB - Respiratory syncytial virus (RSV) infection is the most prevalent cause of severe respiratory disease in infants. It also causes considerable morbidity in older children and adults with underlying risk factors. RSV vaccine development has been complicated by the need to administer the vaccine at a very young age and by enhanced disease observed after vaccination with formalin inactivated RSV. For infants live attenuated vaccines, which may not be expected to predispose for vaccine induced enhanced pathology, hold the greatest promise. However, the balance between attenuation and immunogenicity appears to be delicate. For older risk groups, results with subunit vaccines are most promising. PMID- 11282188 TI - Intra-epithelial vaccination with COPV L1 DNA by particle-mediated DNA delivery protects against mucosal challenge with infectious COPV in beagle dogs. AB - Protection against viral challenge with canine oral papillomavirus (COPV) was achieved by immunisation via particle-mediated DNA delivery (PMDD) of a plasmid encoding the COPV L1 gene to cutaneous and oral mucosal sites in beagle dogs. The initial dose of approximately 9 microg of DNA was followed by two booster doses at 6 week intervals. A similar approach was used to vaccinate a control group of animals with plasmid DNA encoding the Hepatitis B virus S gene. Following challenge at the oral mucosa with COPV all animals vaccinated with the COPV L1 gene were protected against disease. However five of six animals in the control group developed COPV induced papillomas at the oral mucosa. Both cell-mediated lymphoproliferative and humoral antibody responses to the DNA vaccine were observed. Our data indicate that PMDD of plasmid DNA can protect against mucosal challenge with papillomavirus. PMID- 11282189 TI - Mutations of rubella virus vaccine TO-336 strain occurred in the attenuation process of wild progenitor virus. AB - The sequences of the genomes in the TO-336 vaccine strain (TO-336vac) of rubella virus and its wild progenitor virus (TO-336wt) have been determined and compared with each other. There were 21 differences in the nucleotide sequences between the TO-336vac and the TO-336wt: 13 in the nonstructural protein open reading frame (NSP-ORF), five in the structural protein open reading frame (SP-ORF) and three in the untranslated regions (UTRs) (one in each three UTRs). These mutations resulted in amino acid substitutions at ten residues. Of the ten substitutions, eight were in NSP-ORF and two were in the SP-ORF. Of the eight substitutions in NSP-ORF, four (amino acids (aa) 320, 501, 573 and 704) were in the regions of unknown function, two (aa 1154 and 1159) were within the protease motif, and two (aa 1351 and 1559) were within the helicase motif. Both of the two residues (aa 890 and 954) in the SP-ORF were within the E1 gene. The predicted second structure of the 5'UTR of the TO-336vac was identical to that of TO-336wt. Comparing the TO-336 sequences with other four strains, Therien and M33 (wild viruses), and RA27/3 and Cendehill (vaccine viruses), the mutations responsible for attenuation are thought to differ with each vaccine strain. This is the first report of sequencing in a pair of live attenuated rubella vaccines and their wild type parent. PMID- 11282190 TI - Modulation of cellular and humoral immune responses to a multiepitopic HIV-1 DNA vaccine by interleukin-18 DNA immunization/viral protein boost. AB - In this study, the impact of Th1-inducing cytokine gene co-delivery and antigen boosting on humoral and cellular responses induced by multiepitopic DNA immunization in mice have been investigated. Intramuscular injection of mixed DNA constructs encoding for HIV-1 Gag, Tat and Nef proteins, co-administered with the DNA encoding for interleukin-18 (IL-18) have been used. The effect of boosting with the recombinant proteins was also evaluated on the outcome of the responses in DNA-primed mice. It was demonstrated that at least two DNA immunizations were necessary to generate virus specific Th-1 responses detected by the presence of cytotoxic T lymphocyte (CTL) and by the secretion of IL-2 and IFN-gamma, but not IL-4 and IL-10, in antigen-stimulated splenocyte cultures. Interestingly, co delivery of Th-1-inducing IL-18 gene was able to shorten by 2 weeks, the CTL induction time, and to increase the antigen-induced secretion of IL-2 and IFN gamma. Furthermore, IL-18 co-delivery enhanced antigen-specific lymphoproliferative responses, and this was most evident in mice that were primed and boosted with plasmid DNA. However, the induction of detectable antibodies in mice required two DNA vaccinations and a protein boost. In contrast to the effects on cell-mediated immunity, co-administration of IL-18-plasmid resulted in decreased antibody titers against viral proteins. PMID- 11282191 TI - Persistence of anti-HBs 5 years after the introduction of routine infant and adolescent vaccination in Italy. AB - A population survey was conducted to assess the duration of anti-HBs levels > 10 IU/l in vaccinees living in Lazio Region (Italy) 5 years after the introduction (15 June 1991) of compulsory vaccination of new-borns and 11-year-old children. A random sample of 1192 (533 children born in 1991--92 and 659 adolescents born in 1979--81) was selected. In 92.9% of children and 94.1% of adolescents anti-HBs titres were protective (> or = 10 IU/l). These subjects with protective titres were divided into three categories: low responders (anti-HBs titres = 10--500 IU/l), medium responders (anti-HBs titres = 501--2000 IU/l) and high responders (anti-HBs titres > 2000 IU/l). Factors associated with the level of response were analysed, using a multiple politomic logistic regression analysis. Greater age at first dose (11--12 years) was associated with higher titres (OR = 2.1, 95% CI = 1.4--3.2 for medium responders and OR = 3.0, 95% CI = 1.9--4.8 for high responders). Simultaneous administration of DT vaccine was associated with lower titres (OR = 0.4, 95% CI = 0.2-0.8 for medium responders and OR = 0.3, 95% CI = 0.1--0.7 for high responders). PMID- 11282192 TI - Increment of recombinant hepatitis B surface antigen-specific T-cell precursors after revaccination of slow responder children. AB - The aim of the study was to investigate the in vitro T-cell response to recombinant hepatitis B (rHBsAg) in a group of children (defined as "slow responders") vaccinated at birth, presenting antibody levels < 10 mIU/ml after the vaccination schedule, and developing anti-rHBs antibodies after revaccination. T-cell mediated immune response towards rHBsAg was evaluated in 35 healthy children in "bulk" culture experiments (19 responders and 16 slow responders) and by limiting dilution analysis (nine responders and five slow responders) to quantify the frequency of proliferating T lymphocyte-precursors (PTL-p). Before the booster dose, lymphocytes from slow responder children failed to proliferate to rHBsAg, while a normal proliferation was observed in all responders. A statistically significant difference in rHBsAg-specific PTLp frequencies was observed between the two groups. Among the slow responder group, a significant increase of PTLp was observed after the supplementary vaccine dose.Nevertheless, PTLp frequencies remained significantly lower than those measured in responders. These results suggest a role for follow-up of slow responder children over time, in order to perform booster vaccination when inadequate anti-HBs titre is present. PMID- 11282193 TI - Efficacy of a mass hepatitis B immunization program after switching to recombinant hepatitis B vaccine: a population-based study in Taiwan. AB - To study the efficacy of immunization against hepatitis B after plasma-derived vaccine was replaced by recombinant vaccine, 2-year-old Taiwanese children were recruited by stratification random sampling and tested for hepatitis B markers. They were grouped according to maternal infectivity and children's immunization status. Of 2010 children, 2.5% had hepatitis B surface antigen (HBsAg), 94.1% had its antibody (anti-HBs), 6.8% had core antibody, and 3.3% were seronegative. Children of highly infectious mothers immunized with hepatitis B immunoglobulin and vaccine on schedule had a lower HBsAg-positive rate and a higher anti-HBs positive rate than those with vaccine only and off-schedule. The efficacy of the Taiwanese mass hepatitis B immunization was maintained after switching to recombinant hepatitis B vaccine. PMID- 11282194 TI - Mixed immune response induced in rodents by two naked DNA genes coding for mycobacterial glycosylated proteins. AB - Two genes of Mycobacterium tuberculosis, apa (Rv1860) and pro (Rv1796), coding for two glycosylated excreted proteins have been injected to mice and guinea pigs. They produce an extended immunological response of Th1 and Th2 types. Despite the fact that mycobacterial glycosylation is necessary for a high level of delayed-type hypersensitivity (DTH) reaction, plasmids bearing each of the two genes induced an elevated level of DTH sensitization. An inverse relation between the CpG-N hexamer cluster frequency and the protective effect of injected genes is described. A comparison of the strength of several eukaryotic promoters based on the diameter of the DTH reaction shows that CMVIE followed by the ubiquitin promoter are the most efficient among those tested. A significant protective effect (0.7 log unit CFU) in mice was found for the apa gene while the pro gene had no effect. PMID- 11282195 TI - Immunization of pigs against influenza virus infection by DNA vaccine priming followed by killed-virus vaccine boosting. AB - In a previous study of particle-mediated DNA vaccination of pigs, it was found that administration of an influenza virus hemagglutinin (HA) gene elicited low levels of virus-specific antibody, but did not provide significant protection from challenge infection (as evidenced by virus shedding in nasal secretions). However, the vaccinated pigs developed high antibody titers after exposure to the challenge virus, suggesting strong priming of humoral immune responses by DNA vaccination. In the present study, pigs given a conventional, inactivated influenza virus vaccine 4 weeks after a priming dose of HA DNA developed higher levels of virus-specific serum antibodies and were protected from challenge virus infection to a significantly greater degree than pigs that received two doses of DNA vaccine. PMID- 11282196 TI - Nasal vaccination with attenuated Salmonella typhimurium strains expressing the Hepatitis B nucleocapsid: dose response analysis. AB - Nasal vaccination of mice with recombinant attenuated strains of Salmonella typhimurium is more efficient at inducing antibody responses than oral vaccination. However, mortality was observed when high doses [10(9) colony forming unit (CFU)], otherwise safe by the oral route, were administered. This observation was counterbalanced by the fact that nasal vaccination was still highly efficient with lower doses (10(6) CFU), which are inefficient by the oral route and this, without any incidents of mortality. Here, we further analyse in mice the effect of nasal vaccination with differently attenuated S. typhimurium strains expressing the Hepatitis B nucleocapsid (HBc). Surprisingly, as few as 100 CFU were sufficient to induce a maximal HBc specific antibody response, but only if the bacteria were inhaled. Furthermore, we observed no correlation between the inoculum dose and the number of surviving bacteria in cervical lymph nodes and spleen. Examination of lung sections revealed strong inflammation and bronchopneumonia 24 h after nasal vaccination with 10(8) CFU, while only minor signs of inflammation were detected transiently when 10(3) CFU or phosphate buffered saline (PBS) were administered. Our data suggest that the safety issue of nasal vaccination with low doses of the Salmonella vaccine strains should be addressed in humans, as it might be an efficient alternative to oral vaccination. PMID- 11282197 TI - Vaccination with DNA containing tat coding sequences and unmethylated CpG motifs protects cynomolgus monkeys upon infection with simian/human immunodeficiency virus (SHIV89.6P). AB - Recent evidence suggests that a CD8-mediated cytotoxic T cell response against the Tat protein of human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) controls primary infection after pathogenic virus challenge, and correlates with the status of long-term nonprogressor in humans. Due to the presence of unmethylated CpG sequences, DNA vaccination can boost the innate immunity driving more potent T cell-mediated immune responses. Therefore, cynomolgus monkeys were vaccinated with a tat-expressing vector containing defined unmethylated CpG sequences (pCV-tat). Here it is shown that the intramuscular inoculation of the pCV-tat contained primary infection with the highly pathogenic SHIV89.6P virus preventing the CD4(+) T cell decline in all the vaccinated monkeys. Undetectable virus replication and negative virus isolation correlated in all cases with the presence of anti-Tat CTLs. However, a CD8 mediated non cytolytic antiviral activity was also present in all protected animals. Of note, this activity was absent in the controls but was present in the monkey inoculated with the CpG-rich vector alone that was partially protected against viral challenge (i.e. no virus replication but positive virus isolation). These results suggest that a CTL response against Tat protects against primary infection by blocking virus replication at its early stage, in the absence of sterilizing immunity. Nevertheless, the boost of the innate immunity by CpG sequences can contribute to this protection both by driving more potent CTL responses and by inducing other CD8-mediated antiviral activities. Thus, the CpG rich tat DNA vaccine may represent a promising candidate for preventive and therapeutic vaccination against AIDS. PMID- 11282198 TI - Detection of antibodies to HAV 3C proteinase in experimentally infected chimpanzees and in naturally infected children. AB - Commercial assays for the diagnosis of hepatitis A detect antibody to hepatitis A virus (anti-HAV), but they cannot discriminate between antibody resulting from infection and antibody induced by inactivated vaccine. With the licensing and increasing use of inactivated hepatitis A vaccines, there is a need for a test to distinguish between infection and vaccination. Since antibodies to viral non structural proteins are elicited by infection but not by vaccination with inactivated vaccine, we developed and evaluated a test for such antibodies. The antibody response to the non-structural 3C proteinase (anti-3C) of virus HAV was studied by ELISA in chimpanzees experimentally infected with virulent (wild type) or with attenuated HAV strains and in children who received inactivated HAV vaccine or placebo during a vaccination trial in Nicaragua. Anti-3C was detected in 89% of 18 chimpanzees infected with wild-type HAV strains and 27% of 26 chimpanzees infected with attenuated HAV strains. There was a direct correlation between severity of hepatitis and magnitude of the anti-3C response. In the vaccine trial, anti-3C was detected only in children who were infected with HAV during the study; IgG anti-3C persisted for at least 15 months after infection in one child. Vaccinated and uninfected children remained negative for anti-3C. The anti-3C response can be regarded as an indicator of viral replication. Its detection should be useful for distinguishing between antibody acquired in response to HAV infection and antibody induced by immunization with inactivated vaccine. PMID- 11282199 TI - Degree of antigen adsorption in the vaccine or interstitial fluid and its effect on the antibody response in rabbits. AB - The effect of the degree of adsorption of lysozyme by aluminium hydroxide adjuvant on the immune response in rabbits was studied. The surface charge of the adjuvant was modified by pretreatment with phosphate anion to produce five vaccines having degrees of adsorption ranging from 3 to 90%. The degree of adsorption of vaccines exhibiting 3, 35 or 85% adsorption changed to 40% within 1 h after each vaccine was mixed with sheep interstitial fluid to simulate subcutaneous administration. The mean anti-lysozyme antibody titers produced by the vaccines were the same and were four times greater than that produced by a lysozyme solution. Thus, the degree of adsorption of lysozyme in sheep interstitial fluid rather than the degree of adsorption in the vaccine correlated with the immune response. PMID- 11282200 TI - Response, tolerance and ignorance following oral exposure to a single dietary protein antigen in Gallus domesticus. AB - The objective of this investigation was to analyze conditions leading to antibody responses against innocuous dietary protein antigens in the chick. The physical form of antigen was found to be important for immunization of mature chicks: bovine serum albumin (BSA) in solution was a powerful immunogen, while BSA powder was ignored. When BSA was fed to newly hatched chicks, either in solution or as powder, specific oral tolerance was induced. Tolerance increased with the dose of antigen fed, and was most effective in suppressing BSA-specific oral immunization. Hence, immune responses of mature chicks to innocuous dietary proteins are not likely to constitute a health hazard due to (a) prevalence of oral tolerance induced at hatch, and (b) availability of dietary proteins in solid form. PMID- 11282201 TI - Nasal or intramuscular immunization of mice with influenza subunit antigen and the B subunit of Escherichia coli heat-labile toxin induces IgA- or IgG-mediated protective mucosal immunity. AB - Local mucosal IgA antibodies play a central role in protection of the respiratory tract against influenza virus infection. Therefore, new-generation influenza vaccines should aim at stimulating not only systemic, but also local antibody responses. Previously, we demonstrated that the recombinant B subunit of the Escherichia coli heat-labile toxin (LTB) is a potent adjuvant towards nasally administered influenza subunit antigen. Here, we investigated the protection conferred by LTB-supplemented influenza subunit antigen given intranasally (i.n.) or intramuscularly (i.m.) to mice. Both i.n. and i.m. immunization with subunit antigen and LTB completely protected the animals against viral infection. Protection upon i.n. immunization was associated with the induction of antigen specific serum IgG and mucosal IgA, whereas protection upon i.m. immunization correlated with strong serum and mucosal IgG, but not IgA responses. We conclude that LTB-supplemented influenza subunit antigen, given either i.n. or i.m, induces protective antibody-mediated mucosal immunity and thus represents a promising novel flu vaccine candidate. PMID- 11282202 TI - Adjuvantation of epidermal powder immunization. AB - The skin is an immunologically active site and an attractive vaccination route. All current vaccines, however, are administered either orally, intramuscularly, or subcutaneously. We previously reported that epidermal powder immunization (EPI) with an extremely small dose of powdered influenza vaccine induces protective immunity in mice. In this study, we report that commonly used adjuvants can be used in EPI to further enhance the immune responses to an antigen. The IgG antibody response to diphtheria toxoid (DT) following EPI was augmented by 25- and 250-fold, when 1 microg DT was co-delivered with aluminum phosphate (alum) and a synthetic oligonucleotide containing CpG DNA motifs (CpG DNA), respectively. These antibodies had toxin-neutralization activity and were long lasting. Furthermore, EPI using an adjuvant selectively activated different subsets of T helper cells and gave either a Th1 or a Th2 type of immune response. Similar to needle injection into deeper tissues, EPI with alum adsorbed DT promoted a predominantly IgG1 subclass antibody response and elevated level of IL 4 secreting cells. These are indicative of Th2-type immunity. In contrast, co delivery of CpG DNA adjuvant via EPI led to Th-1 type of response as characterized by the increased production of IgG2a antibodies and IFN-gamma secreting cells. This study indicated that EPI using appropriate adjuvants can produce an augmented antibody response and desirable cellular immune responses. EPI is a promising immunization method that may be used to administer a broad range of vaccines including vaccines with adjuvants. PMID- 11282203 TI - Induction of hepatitis B virus-specific cytotoxic T lymphocytes response in vivo by filamentous phage display vaccine. AB - The ability of inducing MHC class I restricted cytotoxic T lymphocytes response in vivo via recombinant filamentous phage was investigated. The recombinant filamentous phage particles that displayed the Hepatitis B virus epitope S(28- 39) were injected into BALB/c (H-2d) mice without adjuvants. A MHC class I restricted HBs specific CTL response was found 8 days after injection. The potentiality of using the recombinant filamentous phage as anti-virus vaccine was discussed. PMID- 11282204 TI - Safety and immunogenicity of three lots of meningococcal serogroup C conjugate vaccine administered at 2, 3 and 4 months of age. AB - The reactogenicity and immunogenicity of meningococcal serogroup C conjugate (MenC) vaccine was assessed in 322 infants vaccinated at 2, 3, and 4 months of age, with concomitant administration of mixed diphtheria-tetanus-whole-cell pertussis vaccine and Haemophilus influenzae type b conjugate vaccine (DTwP-Hib) and oral polio vaccine. All infants in whom post-vaccination meningococcal C anticapsular IgG levels were assayed (n = 265) attained > or = 2 microg ml(-1). Serum bactericidal titres were assayed for a proportion of subjects (n = 171), 98% of whom obtained a reciprocal titres > or = 8. Local reactions were less frequent at the MenC injection site than at the DTP-Hib site. Systemic events were frequent, but consistent with established DTwP-Hib experience. The study demonstrates that MenC vaccine is immunogenic and well tolerated in infants at manufacturing scale production levels. PMID- 11282205 TI - Proinflammatory cytokine expression by Theileria annulata infected cell lines correlates with the pathology they cause in vivo. AB - Control of Theileria annulata is currently best achieved by the use of live attenuated cell line vaccines. However, the mechanisms underlying attenuation are unclear and there is a need to rapidly produce new cell line vaccines, which could safely and effectively vaccinate cattle against tropical theileriosis. There is increasing evidence to suggest that proinflammatory cytokines produced by T. annulata infected cells play a central role in both pathology and immune evasion. This study aimed to test this hypothesis and to evaluate cytokine expression as a marker of virulence. The pathogenicity and protective efficacy of cloned T. annulata cell lines that expressed different levels of proinflammatory cytokines were compared. In two independent trials using different stocks of T. annulata, cell lines that expressed higher levels of proinflammatory cytokines induced severe reactions, and in some cases death, when used to vaccinate groups of cattle. In contrast, low cytokine expressing lines induced low post-vaccinal reactions. The results clearly demonstrated that cytokine expression by T. annulata infected cells could be used as a marker of virulence and provided strong evidence to support a role for cytokines in the induction of pathology. Both high and low cytokine expressing cell lines protected cattle against heterologous challenge infection, offering the possibility of using cytokine expression to rapidly select new safe, potent vaccines against tropical theileriosis without the need for culture attenuation. PMID- 11282206 TI - Unique immunogenicity of hepatitis B virus DNA vaccine presented by live attenuated Salmonella typhimurium. AB - A novel vaccine for hepatitis B virus (HBV) was designed by putting a naked DNA vaccine carrying hepatitis B surface antigen (HBsAg) into live-attenuated Salmonella typhimurium. Mucosal immunization by the oral route in mice showed significantly stronger cytotoxic T lymphocyte (CTL) response than recombinant HBsAg vaccination (P < 0.01 at an effector:target ratio of 100:1), while comparable to intramuscular naked DNA immunization at all effector:target ratios. Contrary to previous reports on naked DNA vaccines given intramuscularly, the IgG antibody response induced by the mucosal DNA vaccine is relatively weak when compared to recombinant HBsAg vaccine (P < 0.001 at day 21). These findings are supported by a high interferon-gamma but a low interleukin-4 level detected in the supernatant of splenic cell cultures obtained from mucosally immunized mice. As distinct to recombinant HBsAg vaccine which is effective for protection, oral mucosal DNA vaccine should be considered as a candidate for therapeutic immunization in chronic HBV infection, donor immunization before adoptive transfer of HBV-specific CTL to HBsAg positive bone marrow transplant recipients, and immunization of non-responders to recombinant HBsAg vaccine. This strongly cellular and relatively absent humoral response may make this vaccine a better candidate as a therapeutic vaccine for chronic HBV carriers than naked DNA vaccines, as the humoral response is relatively less important for the clearance of HBV from hepatocytes, but its presence may lead to side effects such as serum sickness and immune complex deposition in chronic HBV carriers. PMID- 11282207 TI - Immunogenicity of the E1E2 proteins of hepatitis C virus expressed by recombinant adenoviruses. AB - The E1 and E2 proteins of hepatitis C virus (HCV) are believed to be the viral envelope glycoproteins that are major candidate antigens for HCV vaccine development. We reported previously that the replication-competent recombinant adenovirus encoding core-E1-E2 genes of HCV (Ad/HCV) produces serologically reactive E1 and E2 proteins forming a heterodimer in substantial amounts. Here, we examined immunogenicity of the E1E2 proteins copurified from HeLa cells infected with Ad/HCV virus in mice. Furthermore, we constructed a replication defective recombinant adenovirus encoding the core-E1-E2 genes of HCV (Ad.CMV.HCV) and examined immunogenicity of the virus in mice. The mice immunized intraperitoneally with the copurified E1E2 proteins induced mainly antibodies to E2, but not to E1 by Western blot analysis. The sera of mice immunized with the E1E2 inhibited the binding of E2 protein to the major extracellular loop of human CD81. E2-specific cytotoxic T cells (CTLs), but not antibodies to the E1E2 antigens were induced in the mice intramuscularly immunized with Ad.CMV.HCV virus. When immunized with both Ad.CMV.HCV virus and the E1E2, mice elicited E2 specific CTLs and antibodies to the E1E2 antigens. The results suggest that immunization of Ad.CMV.HCV virus combined with E2 protein is an effective modality to induce humoral as well as cellular immune response to E2 antigen. PMID- 11282208 TI - Oral immunisation with peptide and protein antigens by formulation in lipid vesicles incorporating bile salts (bilosomes). AB - The ability of non-ionic surfactant vesicles to induce systemic immune responses in mice following oral immunisation was studied using a standard antigen (bovine serum albumin), a synthetic measles peptide and an influenza sub-unit vaccine. The effectiveness of this formulation was significantly increased by incorporating bile salts (in particular deoxycholate) into the formulation. We have named the resulting vesicles bilosomes. We found that the most effective immunisation protocol was to give two doses of vaccine three days apart and then repeat this protocol two weeks later. Following this method, preparation of measles peptide in bilosomes produced a specific cell mediated response, as measured by splenocyte proliferation and IL-2 production. Of particular significance, these studies demonstrate that oral administration of bilosomes incorporating the influenza sub-unit vaccine could induce as potent an antibody response as the parenterally administered vaccine containing the same quantity of antigen. In addition, the Th1/Th2 balance, as measured by antibody subclasses, was similar whether animals were immunised by the oral or the parenteral vaccine route. As bilosomes are prepared from naturally occurring lipids and have no apparent toxicity associated with their use, they represent a useful modification of conventional lipid vesicle based systems for the oral delivery of proteins and peptides. PMID- 11282209 TI - Qualitative assessment of the humoral immune status against FMDV in post vaccination cattle. AB - Analyses on diluted sera would not measure the complete activity of the natural in-vivo serum environment used by humoral immune responses. Since these humoral defences must react rapidly, serum reactions occurring during 10 and 60 s were analysed. Primo- and multiple-vaccinated, efficiently responsive, and protected animals were differentiated. A variation in the responses, particularly the quality of response, of individual animals was also now discernible. The critical importance of antibody-antigen contact time was demonstrated, wherein natural immunoglobulins and non-immunoglobulin opsonins would influence specific antibody reactivity, through an increased avidity of reaction with antigen. Although not measured directly, the influence of the non-specific serum components would be manifest through increased specific antibody binding. By considering serum as an entity, analysis of all constituent components permits increased qualitative assessment of post-vaccination sera. PMID- 11282210 TI - Quantitative determination of C-polysaccharide in Streptococcus pneumoniae capsular polysaccharides by use of high-performance anion-exchange chromatography with pulsed amperometric detection. AB - Capsular polysaccharides of Streptococcus pneumoniae are used to formulate polyvalent pneumococcal vaccines. A sensitive method, using high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD), has been developed to quantify the contamination of pneumococcal capsular polysaccharides (PnPs) with the C-polysaccharide (C-Ps). As this polysaccharide is highly immunogenic, and since anti C-Ps antibodies are not protective, the need to monitor and reduce its level is of uppermost importance. The method is based on the quantification by HPAEC-PAD of ribitol, which is released by a two step hydrolysis of the PnPs using aqueous hydrofluoric acid (HF) followed by trifluoroacetic acid hydrolysis (TFA). This simple method has been shown to provide both qualitative and quantitative information about the purity of polysaccharide preparations. PMID- 11282211 TI - Enhancement of mucosal immune response against HIV-1 Gag by DNA immunization. AB - In order to examine the feasibility of Gag-expression DNA as a potential candidate for HIV vaccine using a mouse model, we injected DNA into mice either intramuscularly or by using a gene gun. Both methods induced a low level of antibody production. However, after booster immunization with p24 protein emulsified with complete Freund's adjuvant via a footpad, we found that only the preceding intramuscular DNA immunization induced an anti-Gag Th1-type (IgG(2a)) antibody response, in addition to the enhancement of a Th2-type (IgG(1)) antibody response. Importantly, when mice were boosted intranasally with p24 and cholera toxin, intramuscular DNA injection was found to enhance both systemic and mucosal Gag-specific immune responses. These results indicate that intramuscular DNA immunization confers the inducibility of memory cells, which circulate around various mucosal tissues. Therefore, intramuscular DNA priming, followed by a mucosal booster immunization, could be considered as a regimen applicable to HIV vaccine. PMID- 11282212 TI - Effectiveness of a single dose of acellular pertussis vaccine to prevent pertussis in children primed with pertussis whole cell vaccine. AB - We estimated the protection given by one booster dose of acellular pertussis vaccine (aP) given at 18 months or before school entry to children already primed with whole cell vaccine (wP). Case-control studies were conducted in these two age groups. In children who received or were eligible to receive their 18 months booster, the risk of pertussis was 1.4 and 3.6 times higher for those with 4 and 3 wP, respectively, compared to those with 3 wP + 1 aP. In 5 and 6 yr old children, the risk of pertussis among the subjects with 5 and 4 wP, was 1.4 and 2.1 times higher respectively than in those who received 4 wP + 1 aP. A single dose of aP increased the protection against pertussis and this protection was greater than that obtained with a wP booster. PMID- 11282213 TI - Enhanced immune responses to viral epitopes by combining macrophage-inducible expression with multimeric display on a Salmonella vector. AB - In this study, the immunogenicity of chimeric 987P fimbriae on a Salmonella vaccine strain was improved by optimizing fimbrial expression. The constitutive tetA promoter and the in vivo activated nirB and pagC promoters were evaluated for their use to express two epitopes of the transmissible gastroenteritis virus (TGEV) spike protein carried by fimbriae which were displayed on a Salmonella vaccine strain. Constructs with the pagC promoter were shown to drive increased expression of chimeric 987P fimbriae in macrophages as well as in Mg(2+)-poor media, mimicking a major environmental signal found in Salmonella-containing endocytic vacuoles of macrophages. Mice immunized orally with a Salmonella vaccine strain which expressed chimeric fimbriae from the pagC promoter elicited significantly higher mucosal and systemic immune responses to both the 987P fimbriae and the TGEV epitopes than mice immunized with the same strain hosting a tetA or nirB promoter-driven expression plasmid. Moreover, only the Salmonella vaccine strains harboring a plasmid with the pagC promoter, with or without an additional tetA promoter in tandem, elicited neutralizing antibodies to TGEV. This indicated that the pagC promoter can be used successfully to improve epitope display by chimeric fimbriae on Salmonella vaccine strains for the induction of a desired immune response. PMID- 11282214 TI - Low or no antibody responses to human immunodeficiency virus type 1 Nef in infected carriers with subtype E, in contrast to subtype B that showed antibodies preferentially recognizing subtype-specific Nef epitopes. AB - The viral accessory gene product Nef has been shown to play an important role in human immunodeficiency virus type 1 (HIV-1)-induced pathogenesis. Only little information is available regarding the differences in the host immune responses against Nef protein and its function in vivo among different subtypes of HIV-1. In the present study, we showed marked differences in the immune responses to Nef protein between subtypes B and E. The amino acid sequence in subtype E Nef showed 72% homology with that in subtype B. Most murine monoclonal antibodies obtained by immunization with subtype B or E Nef protein showed cross-reactivity with both Nef proteins (80 and 67%, respectively). Next, we focused on the immune responses among infected Japanese and Thai individuals. Subtyping of the individuals into B and E was carried out by enzyme-linked immunosorbent assay (ELISA) using synthetic peptides corresponding to the V3 loop representing the principal neutralizing domain. Most of the sera from these individuals reacted strongly with Gag p24 proteins derived from subtypes B and E at similar levels. However, the immune responses among these individuals to Nef protein were markedly different. Some subtype B-infected Japanese and Thai individuals (40 and 35%, respectively) showed higher levels of anti-Nef antibodies, although these antibodies preferentially recognized epitopes specific to subtype B. On the other hand, most of the subtype E-infected Japanese and Thai individuals showed low or no antibody responses to Nef proteins. Thus, immune responses to Nef were markedly different between subtypes B- and E-infected carriers, suggesting different function(s) for Nef in AIDS pathogenesis. Further, vaccine design must take into account the different subtypes of HIV-1. PMID- 11282215 TI - Insulin effects on the sympathetic contraction of rabbit ear arteries. AB - Electrical field stimulation (4 Hz, 0.2 ms, 70 V supramaximal voltage, 10 s duration) produced contraction of perfused rabbit central ear arteries, and this contraction was reduced by incubation with insulin (0.6--200 mU/ml). This inhibitory effect of insulin was not significantly modified by removing the endothelium, or by treatment with N(W)-nitro-L-arginine (L-NA, 10(-4) M), meclofenamate (10(-5) M), ouabain (10(-6) M), or cocaine (10(-5) M). Insulin (200 mU/ml) did not modify the vascular contraction due to exogenous norepinephrine (10(-8)--10(-4) M) nor the relaxation due to acetylcholine (10(-8)--10(-4) M). This suggests that insulin may reduce vascular contraction by sympathetic stimulation, and this effect is not dependent on endothelial nitric oxide, prostanoids, or Na(+)--K(+) pump activation. PMID- 11282216 TI - Hwansodan protects PC12 cells against serum-deprivation-induced apoptosis via a mechanism involving Ras and mitogen-activated protein (MAP) kinase pathway. AB - Hwansodan has been used as a prescription for senile and vascular dementia in Oriental medicine. We investigated the neuroprotective effects of Hwansodan water extract on the apoptotic death of PC12 cells by serum deprivation. Hwansodan significantly rescued PC12 cells from apoptotic death by serum deprivation in a dose-dependent manner. The nuclear staining of PC12 cells clearly showed that Hwansodan attenuated nuclear condensation and fragmentation, which represents typical neuronal apoptotic characteristics. Hwansodan also prevents DNA fragmentation and caspase-3-like protease activation in serum-deprived PC12 cells and induces the tyrosine phosphorylation of proteins around 44 kDa, which was identified as ERK1 with electrophoretic gel mobility shift by Western blot. In addition, MEK inhibitor PD98059 and Ras inactivator, alpha hydroxyfarnesylphosphonic acid and mevastatin, attenuated the neuroprotective effects of Hwansodan in serum-deprived PC12 cells. These results indicate that Ras/MEK/ERK signaling pathway plays a role in neuroprotective effects of Hwansodan in serum-deprived PC12 cells. PMID- 11282217 TI - Pharmacokinetics of verapamil in lactating rabbits. Prediction of verapamil distribution into rabbit milk. AB - In this work, we have studied the pharmacokinetics and milk penetration of verapamil following intravenous administration in lactating rabbits. Milk-to serum drug concentration ratios (M/B(obs)) have been determined using area under the milk and serum concentration-time profiles, and the resulting values have then been compared with those obtained by theoretical classical diffusion milk transfer models that were described by Fleishaker et al. [J. Pharm. Sci. 76 (1987) 189.], Atkinson and Begg [Br. J. Clin. Pharmacol. 25 (1990) 495.], and Stebler and Guentert [Pharm. Res. 9 (1992) 1299.]. The pharmacokinetic profile of verapamil in lactating rabbits following endovenous administration is described in the form of a two-compartment model. Moreover, we detected an important milk transfer after endovenous administration of verapamil in lactating rabbits. M/B(obs) was near 15. The classical diffusional models mentioned were not able to predict this extensive transfer of verapamil into rabbit milk. However, when the classical Fleishaker equation was modified and a stepwise regression was carried out, we found that the M/B(obs) value could be predicted using the plasma and milk protein binding. PMID- 11282218 TI - The role of cyclic nucleotides and calcium in the relaxation produced by amrinone in rat aorta. AB - (1) The vasorelaxation produced by the phosphodiesterase 3 (PDE3) inhibitor, amrinone was investigated in isolated rat aorta denuded of endothelium. In the presence of extracellular Ca(2+), amrinone, milrinone and 3-isobutyl-1 methylxanthine (IBMX), relaxed endothelium-denuded rat aortic rings constricted with phenylephrine. While the actions of milrinone and IBMX were inhibited by the protein kinase G (PKG) inhibitor, Rp-8-Bromo guanosine-3',5' monophosphothioate (Rp-8-Br-cGMPS; 0.5 mM), that of amrinone was only slightly affected; whereas the protein kinase A (PKA) inhibitor, Rp-adenosine-3',5' cyclic monophosphothioate (Rp-cAMPS; 0.5 mM) had no effect on any agent. (2) Amrinone (100 microM) inhibited (45)Ca(2+) influx through receptor- or store-operated Ca(2+) channels following stimulation with phenylephrine (1 microM) or thapsigargin (1 microM). In contrast, amrinone had no effect on KCl (120 mM)-stimulated Ca(2+) influx. (3) In the absence of extracellular Ca(2+), amrinone (30 microM) inhibited the constriction produced by phenylephrine, 5-hydroxytryptamine (5HT) and U46619, and this effect was not affected by Rp-cAMPS or Rp-8-Br-cGMPS. (4) The intracellular mechanism of action of amrinone may involve the phospholipase C (PLC)-inositol 1,4,5 trisphosphate (IP(3))-intracellular Ca(2+) signal transduction pathway. However, amrinone (100 microM) had no effect on either basal- or noradrenaline (100 microM)-stimulated PLC activity. Similarly, IP(3) stimulated a concentration dependent release of Ca(2+) from rat brain microsomes that was not affected by amrinone (30 and 100 microM). (5) In conclusion, the vasorelaxant action of amrinone does not involve adenosine 3',5' cyclic monophosphate (cAMP) or involve guanosine 3',5' cyclic monophosphate (cGMP) but may include an inhibition of Ca(2+) influx through receptor- or store-operated Ca(2+) channels, although it does not directly affect intracellular Ca(2+) release. PMID- 11282219 TI - Interactions between agmatine and polyamine uptake pathways in rat pulmonary artery endothelial cells. AB - Agmatine, a product of arginine metabolism in vascular endothelial cells, is structurally similar to the natural polyamines, putrescine, spermidine and spermine. To test the hypothesis that agmatine and polyamines interacted at the level of the polyamine transporter, we determined if polyamines competed with agmatine for import and whether interventions modulating polyamine import exerted coordinate effects on agmatine uptake. Multiple lines of evidence were obtained to suggest that agmatine enters pulmonary artery endothelial cells (PAECs) via the polyamine transporter, though its intracellular disposition after uptake appears different from the natural polyamines. PMID- 11282221 TI - Abstracts from 7th Biannual Conference on Vascular Endothelium: source and target of inflammatory mediators; June 24-July 3, 2000, Knossos Royal Village, Crete, Greece. PMID- 11282220 TI - Effect of atorvastatin on endothelium-dependent constrictor factors in dyslipidemic rabbits. AB - Relaxations to acetylcholine and contractions to acetylcholine in the presence of the nitric oxide (NO) synthesis inhibitor (L-N(G)-nitroarginine methyl ester, L NAME) were studied in aortic rings from rabbits fed either a control or a diet containing 0.5% cholesterol+14% coconut oil for 14 weeks and treated or not with atorvastatin (2.5 mg kg(-1) day(-1)). Rings were incubated with the endothelin (ET(A)) receptor antagonist BQ123, and/or the thromboxane A(2) (TXA(2))/prostaglandin H(2) (PGH(2)) receptor antagonist ifetroban. In rabbits, high cholesterol and triglyceride plasma levels were associated with intimal thickening and blunted acetylcholine-relaxation as compared with controls. By contrast, acetylcholine+L-NAME response was higher. Incubation with either ifetroban or BQ123 increased acetylcholine-relaxation in both diet groups and it reduced the constrictor response only in dyslipidemic rabbits. Removal of endothelium reduced acetylcholine+L-NAME contraction in dyslipidemic rabbits, although increased it in control animals. Atorvastatin treatment reduced plasma lipid levels and lesion size in dyslipidemic animals. Likewise, it prevented acetylcholine-relaxation reduction. In addition, atorvastatin reduced constrictor response in dyslipidemic rabbits but only in rings with endothelium. Incubation with either ifetroban or BQ123 did not further modify these responses in atorvastatin-treated animals in any group. These data suggest that ET and TXA(2) availabilities seem to participate in the endothelial dysfunction associated with dyslipidemia. Atorvastatin treatment reduces intimal thickening and improves endothelial dysfunction in rabbits. This effect seems to be a consequence of its ability to act on ET and TXA(2) systems. PMID- 11282222 TI - Blood pressure in the very low birth weight neonate. AB - Few aspects of management of very low birth weight (VLBW; <1500 g) neonates have generated as much controversy as the assessment of blood pressure (BP) and need for treatment of perceived abnormalities of this physiologic variable. The approach to this problem may differ greatly among various institutions and even among clinicians within a given center. The purpose of this manuscript is to review available information regarding physiologic determinants and measurement of BP in VLBW neonates, normative data for BP, clinical factors that may affect BP in these at-risk neonates and studies in which presumed abnormalities of BP resulted in adverse clinical outcomes. Options for treatment of low BP in VLBW neonates also will be discussed. PMID- 11282224 TI - Development of emotional sweating in preterms measured by skin conductance changes. AB - Skin conductance shows the emotional state, as reflected in changes in the sympathetic nervous system. Skin conductance changes (number and amplitude of the waves, as well as mean skin conductance level) were measured in connection with heel prick from 29 weeks gestational age. The purposes of this study were to examine the development of emotional sweating in preterm infants, and to correlate the changes in emotional sweating with the changes in behavioural state. Fifty infants' behavioural state and skin conductance changes were measured for 2 min before, 2 min during, and 2 min after heel prick. Half of the infants were between 29 and 31 weeks gestational age. They were divided into three sub-groups; 0-10, 11-20 and 21-30 days postnatal age. The other half of the infants were between 32 and 34 weeks gestational age and they were divided into three similar sub-groups. They changed their behavioural state 114 times. Infants from 29 weeks gestational age and more than 10 days old showed emotional sweating as measured by the number and amplitude of the waves that were lowest in sleep and highest during crying (p<0.05). The mean skin conductance level mirrored the behavioural state from 34 weeks gestational age (p<0.05).To conclude, skin conductance changes increased with the level of behavioural state from 29 weeks gestational age and more than 10 days postnatal age. PMID- 11282223 TI - Permanent tooth crown dimensions in prematurely born children. AB - AIM: The aim was to examine the effect of preterm birth on permanent tooth crown dimensions. MATERIALS AND METHODS: The data consisted of 328 prematurely born white and black children and 1804 control children who participated in the cross sectional study of the Collaborative Perinatal Project (USA) in the early 1960s and 1970s. The dental examinations were carried out in a standardized fashion at ages varying from 6 to 12 years in 95% of cases. Tooth crown size measurements were performed on the dental casts with an electronic measuring device and readout by two experienced observers according to precise definitions generally quoted in the anthropological and genetic literature. The preterm and control groups were divided by sex and race. RESULTS: The results show both increased and decreased tooth crown dimensions in the prematurely born children. Significantly increased dimensions were found in the means of the intercuspal distances of the first permanent molars in the white boys and in the mesiodistal dimensions (MD) of the lower lateral incisors and the upper left first molar in the black girls. By contrast, there were decreased intercuspal distances, MD and labiolingual (LL) tooth crown dimensions in the white girls and black boys. The statistical method used was the Mann-Whitney's U-test (Willcoxon Rank-Sums test). CONCLUSIONS: The findings partly support previous reports of decreased tooth crown dimensions in preterm infants, but the increased dimensions found in the preterm white boys and black girls differ from earlier reports. Our results indicate the importance of environmental factors including neonatal factors in determining permanent tooth crown dimensions. Growth patterns, the buffering capacity and the timing of sensitive moments in tooth crown volume gain may vary between the sexes and ethnic groups and the possible effect of the accelerated growth period in preterm infants (catch-up growth) may influence the determination of permanent tooth crown dimensions. PMID- 11282225 TI - Secretory IgA, free secretory component and IgD in saliva of newborn infants. AB - AIM: To determine levels of secretory IgA (sIgA), free secretory component (FSC) and IgD in saliva of newborn infants at the age of 1 day and to evaluate the detection patterns, the influence of saliva flow and the relation to serum derived proteins. METHODS: Seventy-three healthy newborn infants were studied. Saliva was obtained from the bottom of the mouth and buccal sulci using a sterile polyethylene tube connected to a syringe. SIgA, FSC, IgD and albumin were measured by radial immunodiffusion. RESULTS: SIgA was detected in 74.0% of all saliva samples, whereas detection rates for FSC and IgD were 94.5% and 75.3%, respectively. Investigation of detection patterns and their relation to saliva flow indicated that secretion of sIgA and FSC into the oral cavity is under similar regulation. Levels of IgD were found to be independent from saliva flow, as well as from concentrations of serum-derived proteins suggesting different regulative mechanisms compared to sIgA and FSC. The flow rate of unstimulated whole saliva in newborn infants was found to be 15 times lower compared to adolescents, emphasizing the role of saliva flow as a limiting factor for secretion of sIgA and FSC. CONCLUSION: SIgA, FSC and IgD can be determined in saliva of newborn infants even in the first day of life. The saliva flow rate has to be considered when evaluating the function and biological relevance of the oral mucosal immune system of newborn infants shortly after birth. PMID- 11282228 TI - Preparation of semisolid drug carriers for topical application based on solid lipid nanoparticles. AB - Aqueous dispersions of solid lipid nanoparticles (SLN) show some interesting features in topical drug delivery. However, to get a semisolid carrier having the appropriate consistency for topical application, the liquid SLN dispersions have to be incorporated in convenient topical dosage forms like hydrogels or creams. This is a time-consuming production process with several disadvantages. A new one step production process delivering a semisolid topical formulation including SLN is presented avoiding these disadvantages. The semisolid SLN dispersions were produced by high-pressure homogenization using an APV Lab 40 homogenizer. The resulting dispersions were characterized concerning their particle size and rheological properties. Despite the high lipid content of the SLN dispersions, they retained their colloidal particle size. Viscoelastic measurements proved the existence of a gel-like structure with a prevailing elastic component. PMID- 11282227 TI - Production and characterisation of mucoadhesive nanosuspensions for the formulation of bupravaquone. AB - Bupravaquone is a new naphthoquinone antibiotic against Cryptosporidium parvum and other parasites. It has attracted interest for the treatment of C. parvum infections, because of the lack of a drug in the treatment of mostly AIDS patients. The bioavailability of bupravaquone is limited when given orally. To overcome the problem of the high elimination rate caused by diarrhoea, typical for C. parvum infections, bupravaquone was formulated as a mucoadhesive nanosuspension, i.e. combining the properties of mucoadhesive drug delivery systems, in this case hydro gels, with nanosuspensions. In this study different polymers/hydro gels were employed to create a prolonged retention time for the drug in the infected gastrointestinal tract (GIT). The second step to improve the bioavailability of bupravaquone was the formulation as nanosuspension. Therefore various concentrations of bupravaquone with different surfactants were tested. The production of these nanosuspensions was carried out by high pressure homogenisation. In addition to the classical stepwise production, about a new one step production method is described. PMID- 11282229 TI - Polyisobutylcyanoacrylate nanocapsules containing an aqueous core for the delivery of oligonucleotides. AB - Antisense oligonucleotides (ODNs) with base sequences complementary to a specific RNA can, after binding to intracellular mRNA, selectively modulate the expression of a gene. However, these molecules are poorly stable in biological fluids and are characterized by a low intracellular penetration. In view of using ODNs as active molecules, the development of nanocapsules containing ODNs in their aqueous core was considered. Nanocapsules were prepared by interfacial polymerization of isobutylcyanoacrylate (IBCA) in a W/O emulsion. After ultracentrifugation and re-suspension in water, the nanocapsules displayed a size of 350+/-100 nm. Oligonucleotide loading did not significantly influence the zeta potential, suggesting that they were located within the core of the nanocapsules. Fluorescence quenching assays confirmed this localization. When encapsulated in the nanocapsules and incubated in the presence of serum, the ODNs were efficiently protected from degradation by nucleases, whereas ODNs adsorbed onto nanospheres were less efficiently protected. This paper describes, for the first time, a nanotechnology able to encapsulate ODNs, rather than adsorbing them at the surface of a solid support. Such a formulation has great potential for oligonucleotide delivery. PMID- 11282230 TI - Degradable dextran microspheres for the controlled release of liposomes. AB - A novel delivery concept based on the encapsulation of liposomes in biodegradable dextran microspheres was developed. The microspheres were prepared using a two phase system, consisting of water/poly(ethylene glycol), and water/methacrylated dextran. Liposomes were encapsulated almost quantitatively and in their intact form, and were released with full preservation of their integrity. The effects of microsphere water content, degree of methacrylate substitution, and type of dextran derivative used on the release rate were investigated. The release of the liposomes from the dextran microspheres was fully controlled by the degradation rate of the spheres. This resulted, after a lag time, in a pulsed release of the liposomes from relatively rapidly degrading microspheres. On the other hand, slower degrading microspheres resulted in sustained release of liposomes over 100 days. The degradation rate of the dextran microspheres, in turn, depended on the water content, the degree of methacrylate substitution, and type of hydrolytically sensitive spacer present in the cross-links. PMID- 11282231 TI - Production and characterisation of highly concentrated nanosuspensions by high pressure homogenisation. AB - Nanosuspensions produced by high-pressure homogenisation are a solution for the formulation of poorly soluble drugs with bioavailability problems. The typical solid concentration of the nanosuspensions is 10%. However, to transfer the nanosuspensions to a dry product (e.g. granulation, tablets, pellets), a higher solid content is required to remove less water. Nanosuspensions with 20 and 30% solid content were produced, the effect of surfactant concentration assessed and their quality (size data) compared with the lower standard concentrations of 1 10% solid. PMID- 11282232 TI - Comparison of two methods of encapsulation of an oligonucleotide into poly(D,L lactic acid) particles. AB - The aim of this study was to compare two methods to encapsulate a 25-mer phosphorothioate oligonucleotide (ODN) into poly(D,L-lactic acid) (PLA) particles. Antisense oligonucleotides belong to a new therapeutic class especially attractive for the treatment of cancers and viral diseases. The development of these new drugs suffers, however, from poor stability in biological media and very low cellular uptake. Polymeric particulate systems display interesting features for ODN delivery. ODN are highly hydrophilic and most encapsulation methods are inappropriate for such molecules. Using poly(D,L lactide) polymer, two methods of encapsulation were compared. First, a double emulsion technique was used to prepare nano- and microparticles. Secondly, the ODN was combined with a quaternary ammonium, the cethyltrimethyl-ammonium bromide (CTAB), to enhance the hydrophobicity of the molecule before entrapment by the emulsification-diffusion method. Both methods led to the formation of individualized and spherical particles loaded with a significant amount of ODN. Similar entrapment efficiencies were obtained for the nanoparticles prepared by both methods (approx. 27% of the theoretical loading) whereas 45% of entrapment efficiency was observed for the microparticles. Seventy five percent of the ODN were released in 60 min with the particles prepared by the emulsification diffusion method, whereas only 7% were released in 60 h when using the double emulsion method. A viability test on U-937 cells showed better survival rates with the particles prepared by the double emulsion technique. The results suggest that the location of the ODN in the polymeric matrix is affected by the encapsulation method. Particles containing CTAB appeared more toxic than the ones obtained by the double emulsion technique, however, these particles can still be used for antisense activity since high oligonucleotide loading can be achieved. PMID- 11282233 TI - Design of triptorelin loaded nanospheres for transdermal iontophoretic administration. AB - Triptorelin is a decapeptide analog of luteinizing hormone releasing hormone, currently used for the treatment of sex-hormones dependents diseases. The aim of this work was to prepare triptorelin-loaded nanospheres useful for transdermal iontophoretic administration. Nanospheres were prepared with the double emulsion/solvent evaporation technique. The effect of three parameters on the encapsulation efficiency has been determined: the role of the pH of the internal and external aqueous phases, the nature of the organic solvent and the effect of three different poly(lactide-co-glycolide) (PLGA) co-polymers. Particle size, zeta potential and release kinetics were also determined. The encapsulation efficiency varied from 4 to 83% reaching the maximum value when both the internal and the external water phases were brought to pH 7 (isoelectric point of the peptide), methylene chloride was used as solvent of the copolymers and PLGA rich in free carboxylic groups was employed. The release profiles obtained with this co-polymer were characterized by the absence of burst effect. This behavior as well as the high encapsulation efficiency was explained by an ionic interaction occurring between the peptide and the co-polymer. This supports the already expressed theory that the release of peptides and proteins from PLGA nanospheres is also governed by the affinity of the encapsulated molecule versus the polymer. The obtained nanoparticles, regarding their size, amount encapsulated and zeta potential, were shown to be suitable for transdermal iontophoretic administration. PMID- 11282234 TI - Tamoxifen encapsulation within polyethylene glycol-coated nanospheres. A new antiestrogen formulation. AB - When dealing with solid tumors in vivo, pegylated long-circulating carrier systems show, after intravenous administration, an attractive extravasation profile with an enhanced localization in the tumoral interstitium. These systems could be of help for the delivery of cancer fighting drugs, such as Tamoxifen, a well known antiestrogen used in breast cancer therapy that possesses an extended biodistribution in vivo. This work aimed at encapsulating Tamoxifen in long circulating poly(MePEGcyanoacrylate-co-hexadecylcyanoacrylate) 1:4 nanospheres. Tamoxifen-loaded poly(MePEGcyanoacrylate-co-hexadecylcyanoacrylate) nanospheres were successfully synthesized and characterized in terms of hydrophilicity/hydrophobicity by a model made up from near infrared spectra using principal component analysis. Zeta potential, drug loading, encapsulation efficiency, as well as biological effect, in vitro release and nanospheres integrity were also investigated. Even though near infrared spectroscopy could not detect Tamoxifen, it revealed that Pluronic F68 was associated with the pegylated nanospheres. HPLC measurements demonstrated that Tamoxifen was encapsulated in the pegylated nanospheres following a partition equilibrium between the polymeric and the aqueous phases. The Tamoxifen encapsulated in the nanospheres still showed a transcription inhibitory activity in ex vivo experiments. However, zeta potential and in vitro release suggested that Tamoxifen was essentially localized at the nanoparticles surface, resulting in an important and immediate drug release. PMID- 11282235 TI - Non-aqueous emulsions: hydrocarbon-formamide systems. AB - There are few reports in the literature on formulation of non-aqueous emulsions. This study was designed to evaluate some design criteria for such systems. Formamide is the closest polar solvent that has the ability to replace water in emulsification when employing established non-ionic surfactants as stabilisers. For the majority of studies, linear alkanes (C6-C16) were dispersed in formamide as the continuous phase were stabilised with polysorbate 20. Initial studies involved gentle emulsification and observing mean globule size. The mean globule size varied in a non-linear fashion with alkyl chain length, the minimum being between C10 and C12. Sonication for 30 s led to smaller differences in the mean globule size. The effect of various parameters such as surfactant concentration and solvophilicity of the surfactant was observed. The surface activities of polysorbate 20, 40, 60 and 80 in formamide and critical micellar concentrations were determined. The latter were several orders of magnitude higher in formamide than in water, and the areas per molecule larger. The addition of water to the dodecane formamide systems did not destabilise the emulsion. Release of the model drug dehydroepiandrosterone from dodecane in formamide emulsions was studied in distilled water, the rate of release being dependent on the volume fraction of dodecane. PMID- 11282236 TI - Interfacial behaviour and micelle formation of novel amphiphilic sequential lipid lysine oligomers. AB - As part of work on the design and synthesis of new supramolecular carrier systems for drugs, a series of novel linear oligomers of alternating alpha-amino tetradecanoic acid and lysine having positively charged groups and lipid chains was synthesised. The smallest member of the series (n=2) is insoluble in water and diluted acid solutions, but the larger members are soluble in acid conditions and poorly soluble in alkaline conditions. Hence, in one series, one can conduct experiments both on the determination of micelle formation and spread monolayer behaviour. The surface pressure-area isotherms revealed limiting surface areas at the air/water interface ranged from 0.04 to 0.9 nm(2) according to the oligomer size, and a linear correlation between the observed area per molecule and that projected by computer-generated molecular models was demonstrated. The surface tension of the soluble members in dilute acid solution fell as the concentration of the oligomers was increased, indicating that all of these polymers were surface active with quite clearly defined critical micelle concentrations. The fluorescence intensity ratio of third to first band in the emission spectra of pyrene as a function of the polymer concentrations demonstrated that, even after normalising the data for the amount of lipid chains in the system, the (n=3) oligomer had fewer accessible hydrophobic sites for pyrene, and the forces of the repulsion between the charged head groups was crucial on the formation of micelles, especially in the case of the n=3 oligomer. Supramolecular fibre-like structures were observed in aqueous solution only when n=3 by transmission electron microscopy (TEM). Cryogenic TEM observation of the (n=3) solution also revealed that the micelles might elongate to form long cylindrical or fibrous structures. The diameter of these structures was estimated to be 6.0-13 nm, although their length varied. PMID- 11282237 TI - Fluorometric method for the simultaneous quantitation of differently-sized nanoparticles in rodent tissue. AB - The oral absorption and systemic translocation of particulate matter via the gastrointestinal tract has been shown by a number of laboratories using a wide variety of particles in different animal species. While there is debate on the magnitude of particle intestinal translocation, which is encumbered by the differing experimental protocols, particularly the method of quantitation of absorbed material, few have sought to examine the pharmacokinetic aspects of particle absorption. We describe in this communication the development of a simple and a rapid fluorometric assay of quantifying tissue-laden fluorescent nanoparticles that is able to isolate, detect and quantify the presence of two or more particle populations differing both in their size and fluorescent label. Six types of polystyrene nanoparticles incorporating different fluorescent markers were spiked in whole livers. The fluorophores were extracted using our previously developed method of freeze-drying the tissue and using chloroform as the extractive solvent. Only two types of particle populations, orange-labelled 40 nm and Fluoroscein-emitting 500 nm nanoparticles, were sufficiently recoverable and provided a high signal-to-noise ratio for further work. The amount of tissue and type of biological tissue type also impacted on the nanoparticle recovery and detection, reflecting, perhaps, the quenching effects of interacting tissue derived molecules. In addition, the results also indicate that the use of nanoparticles incorporating fluorescent dyes that have emission over 500 nm overcome the tissue interfering autofluorescence for low doses of nanoparticles. The use of this fluorometric method has several advantages compared with other modes of quantitation in that it is rapid, non-radioactive and the marker is non leaching. More importantly, it allows the simultaneous detection of multiple fluorophores such that two or more different fluorescent particle populations can be detected in the same sample. This may enable the uncharted area of pharmacokinetic parameters, such as the impedance, augmentation or site of gut uptake of differently sized particles to be studied. PMID- 11282238 TI - Screening effect of PEG on avidin binding to liposome surface receptors. AB - This study investigates the screening effect of poly(ethylene glycol) phospholipids (PE-PEG) on the interaction of avidin with PEGylated liposomes containing surface-bound biotin ligands. The influence of grafting density and lipopolymer chain length is examined. A simple fluorescence assay involving a receptor-mediated fluorescence increase of BODIPY-labeled avidin upon binding to biotinylated lipids is employed to study the screening effect of submicellar concentrations of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylethanolamine-N [poly(ethylene glycol)-2000] (PE-PEG(2000)) and 1,2-dipalmitoyl-sn-glycero-3 phosphatidylethanolamine-N-[poly(ethylene glycol)-5000] (PE-PEG(5000)) incorporated into 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) liposomes. The results show that incorporation of lipopolymers into DPPC lipid bilayers reduces binding of avidin to the biotinylated liposomes, and it is found that the screening effect of PE-PEG(5000) is stronger than that for PE-PEG(2000). Thus, the results reveal that both the grafting density and the polymer length of the PE-PEG lipopolymers are of importance for the ability of water-soluble macromolecules to reach the surface of PEG liposomes. Furthermore, it is found that none of the lipopolymers completely prevents avidin from reaching the surface-bound biotin ligands. PMID- 11282239 TI - Drug delivery by phospholipase A(2) degradable liposomes. AB - The effect of poly(ethylene glycol)-phospholipid (PE-PEG) lipopolymers on phospholipase A(2) (PLA(2)) hydrolysis of liposomes composed of stearoyl oleoylphosphatidylcholine (SOPC) was investigated. The PLA(2) lag-time, which is inversely related to the enzymatic activity, was determined by fluorescence, and the zeta-potentials of the liposomes were measured as a function of PE-PEG lipopolymer concentration. A significant decrease in the lag-time, and hence an increase in enzymatic activity, was observed with increasing amounts of the negatively charged PE-PEG lipopolymers incorporated into the SOPC liposomes. The enhancement of the PLA(2) enzymatic activity might involve a stronger PLA(2) binding affinity towards the negatively charged and polymer covered PEG liposomes. PMID- 11282240 TI - Interaction of cationic partial dendrimers with charged and neutral liposomes. AB - Amphipathic partial dendrimers having three lipidic (C(14)) chains coupled to dendritic lysine head groups with eight, 16 or 32 free terminal amino groups have been synthesised by solid-phase peptide synthesis. Liposomes were prepared with positive, negative and neutral charge using the dehydration-rehydration method and their interaction with the partial dendrimers studied. The interaction efficiency of the partial cationic dendrimers studied was greater than 88%. Interaction of the cationic partial dendrimer converted liposomes with very low or negative charge into positively charged species. Apparent vesicle size increased with the head size of the partial dendrimer but, in the case of negatively charged liposomes, large changes in properties were observed after ultracentrifugation due to the formation of myelin figures. To investigate the mode of interaction with the liposomes, adsorption studies were performed by adding the partial dendrimer after the preparation of dehydration-rehydration vesicles. The results indicated that adsorption is inversely proportional to the head size of the partial dendrimer molecules and the extent of adsorption was similar on both positively and negatively charged liposomes. Adsorption produced liposomes with greater or similar zeta potentials to liposomes that incorporated partial dendrimer through the dehydration-rehydration method. Taking account of the different interaction efficiencies, this suggests there is a degree of partial dendrimer entrappment inside the liposomes. PMID- 11282241 TI - Association of an acylated model peptide with DPPC-DPPS lipid membranes. AB - The interaction between a small positively charged peptide with a N-terminally linked acyl chain and dipalmitoylphosphatidylcholine dipalmitoylphosphatidylserine (DPPC-DPPS) lipid membranes has been studied by means of fluorescence resonance energy transfer. Two different lipid compositions were used: a neutral membrane (100 mol% DPPC), and a negatively charged membrane (30 mol% DPPS in DPPC). The fluorescence resonance energy transfer results reveal that the peptide associates with both types of membranes. Furthermore, it is found that the slope of the titration curve for the negatively charged membranes is much steeper than that for the neutral membranes. This indicates a higher binding affinity of the acylated peptide towards negatively charged lipid membranes as compared with neutral lipid membranes. PMID- 11282242 TI - A new approach for targeting to Cryptosporidium parvum using mucoadhesive nanosuspensions: research and applications. AB - A new strategy to deliver antibiotics to the Cryptosporidium-infected gastrointestinal tract is presented. In an effort to augment the anticryptosporidial effect of clinically used drugs, mucoadhesive nanosuspensions were prepared. They have the ability to reside in the gastrointestinal tract for an extended period. The hydrogel contained bupravaquone nanosuspensions and an adhesive polymer (chitosan) powder dispersed in water. By the development of mucoadhesive nanosuspensions, a potential drug delivery system for poorly soluble drugs has been investigated to overcome bioavailability problems caused by the pathophysiological diarrhoeic situation in patients suffering from cryptosporidiosis. Adapting drug delivery systems to the situation of Cryptosporidium parvum infections in man allows increased retention times with a prolonged action at reduced elimination in the gastrointestinal tract. In this communication, in vivo data are presented to document the efficiency of bupravaquone formulated as mucoadhesive polymers to improve its activity against C. parvum. PMID- 11282243 TI - The role of plasma proteins in brain targeting: species dependent protein adsorption patterns on brain-specific lipid drug conjugate (LDC) nanoparticles. AB - The in vivo organ distribution of particulate drug carriers is decisively influenced by the interaction with plasma proteins after i.v. administration. Serum protein adsorption on lipid drug conjugate nanoparticles, a new carrier system for i.v. application, was investigated by 2-dimensional electrophoresis (2 DE). The particles were surface-modified to target them to the brain. To assess the protein adsorption pattern after i.v. injection in mice prior to in vivo studies, the particles were incubated in mouse serum. Incubation in human serum was carried out in parallel to investigate similarities or differences in the protein patterns obtained from men and mice. Distinct differences were found. Particles incubated in human serum showed preferential adsorption of apolipoproteins A-I, A-IV and E. Previously, preferential adsorption of ApoE was reported as one important factor for targeting of Tween(R)80 modified polybutylcyanoacrylate nanoparticles to the brain. Preferential adsorption of ApoA-I and A-IV took place after incubation in mouse serum, adsorption of ApoE could not be clearly confirmed. In vivo localization of the LDC nanoparticles at the blood-brain barrier and diffusion of the marker Nile Red into the brain could be shown by confocal laser-scanning microscopy. Differences of the obtained adsorption patterns are discussed with regard to their relevance for correlations of in vitro and in vivo data obtained from different species. PMID- 11282244 TI - Activity of mammalian secreted phospholipase A(2) from inflammatory peritoneal fluid towards PEG-liposomes. Early indications. AB - Due to an increase in the activity of phospholipase A(2) (PLA(2)) in various inflammatory diseases, this enzyme may play a key role in the degradation of liposomes and the subsequent release of drug when PEG-liposomes passively target inflammatory tissue. The activity of mammalian secreted phospholipase A(2) (sPLA(2)) in casein stimulated peritoneal fluid was tested toward liposomes of different compositions. Early results indicate only a slight degradation of conventional dipalmitoylphosphatidylcholine (DPPC) liposomes as well as DPPC liposomes incorporated with different concentrations of PEG(2000). However, the DPPC degradation increased to 7% when inclusion of 30 mol% phosphatidylethanolamine (PE) in the lipid bilayer. The increase in degradation may be due to an improvement of the substrate - as it is well known, that PE is a better substrate for the mammalian sPLA(2) than PC. Incorporation of PE into the bilayer may increase the binding properties of the bilayer resulting in improved conditions for the enzymatic attack by sPLA(2). In addition, inhibitory zones of Staphylococcus aureus in an agar diffusion test showed that PLA(2) from Crotalus atrox venom was able to catalyze the release of gentamicin from PEG-liposomes. In conclusion, this study suggest that degradation of the lipid bilayer of PEG liposomes by PLA(2) result in release of incapsulated drug, e.g. gentamicin and inclusion of PE in the liposomal bilayer, may enhance the activity of the mammalian sPLA(2) toward liposomes composed of DPPC. PMID- 11282245 TI - The fate of poly(2-dimethyl amino ethyl)methacrylate-based polyplexes after intravenous administration. AB - Poly(2-dimethyl amino ethyl) methacrylate (pDMAEMA) cationic polymers have been shown to be efficient vectors for gene delivery in vitro. This contribution deals with the in vivo properties of polyplexes based on this polymer. In mice, pDMAEMA/[32P]-pLuc complexes distributed primarily to the lungs. The gene expression profile matched the biodistribution profile. In vitro turbidity experiments in serum showed severe aggregation upon addition of cationic polyplexes, pointing out the involvement of aggregates in the dominant lung uptake of the positively charged polyplexes. Incubations of polyplexes with albumin yielded a decline of the zeta potential of the complexes to negative values, making an electrostatic mechanism in the dominant lung uptake less likely. Hemagglutination experiments showed that the polyplexes induce the formation of extremely large structures when incubated with washed erythrocytes. Altogether, the present data indicate that aggregate formation and trapping of the formed aggregates in the lung capillary bed is probably responsible for the dominant lung uptake and transfection. Poly(ethylene)glycol (PEG) of the polymeric structures prevented the increase in the observed turbidity in serum seen with polyplexes and was also able to reduce interactions with erythrocytes. Currently, the in vivo fate of the PEGylated polyplexes is under investigation. PMID- 11282246 TI - Therapeutic efficacy of liposomal gentamicin in clinically relevant rat models. AB - Sterically stabilized liposomes are able to localize selectively at sites of infection, potentially permitting targeted drug delivery. Up to now, the majority of studies investigating therapeutic efficacy of liposomes have been conducted in animals with an intact host defense infected with high antibiotic-susceptible bacteria. In the present study, the therapeutic efficacy of gentamicin encapsulated in sterically stabilized liposomes, alone or in combination with the free drug was studied in rats with intact host defense as well as leukopenic rats. Rats were inoculated with a high gentamicin-susceptible or low-gentamicin susceptible Klebsiella pneumoniae in the left lung, resulting in an acute unilateral pneumonia. Survival rates demonstrate the valuable therapeutic properties of the liposome-encapsulated drug in these clinically relevant animal models. PMID- 11282247 TI - The neurobiology and genetics of suicide and attempted suicide: a focus on the serotonergic system. AB - Numerous abnormalities have been found in the serotonergic system in suicide attempters and completers. There is considerable evidence that the serotonergic system is partly under genetic control and that as yet unknown genetic factors mediate the risk for suicidal behavior independently of the genetic factors responsible for the heritability of major psychiatric conditions associated with suicide. An argument is made that there is a relationship of genetic variants to intermediate phenotypes, such as impulsivity, psychomotor change, pathological aggression and biological abnormalities including specific gene products. A variety of biological indices have been generated by new approaches using postmortem tissue and in vivo imaging that will provide a rich substrate for further genetic studies. PMID- 11282248 TI - RNA editing of the human serotonin 5-HT2C receptor. alterations in suicide and implications for serotonergic pharmacotherapy. AB - RNA encoding the human serotonin 5-HT2C receptor (5-HT(2C)R) undergoes adenosine to-inosine RNA editing events at five positions, resulting in an alteration of amino acids in the second intracellular loop. Several edited 5-HT(2C)Rs possess a reduced G-protein coupling efficiency compared to the completely non-edited isoform. The current studies show that the efficacy of the hallucinogenic drug lysergic acid diethylamide and of antipsychotic drugs is regulated by RNA editing, suggesting that alterations in editing efficiencies or patterns might result in the generation of a 5-HT(2C)R population differentially responsive to serotonergic drugs. An examination of the efficiencies of RNA editing of the 5 HT(2C)R in prefrontal cortex of control individuals vs. subjects diagnosed with schizophrenia or major depressive disorder revealed no significant differences in RNA editing among the three populations. However, subjects who had committed suicide (regardless of diagnosis) exhibited a statistically significant elevation of editing at the A-site, which is predicted to change the amino acid sequence in the second intracellular loop of the 5-HT(2C)R. These findings suggest that alterations in RNA editing may contribute to or complicate therapy in certain psychiatric disorders. PMID- 11282249 TI - 1-(m-chlorophenyl)piperazine (mCPP) dissociates in vivo serotonin release from long-term serotonin depletion in rat brain. AB - Serotonin (5-HT) releasing agents such as d-fenfluramine are known to cause long term depletion of forebrain 5-HT in animals, but the mechanism of this effect is unknown. In the present study, we examined the relationship between drug-induced 5-HT release and long-term 5-HT depletion in rat brain. The 5-HT-releasing actions of d-fenfluramine and a non-amphetamine 5-HT drug, 1-(m chlorophenyl)piperazine (mCPP), were compared using in vivo microdialysis in the nucleus accumbens. The ability of d-fenfluramine and mCPP to interact with 5-HT transporters was tested using in vitro assays for [3H]5-HT uptake and radioligand binding. Local infusion of d-fenfluramine or mCPP (1-100 microM) increased extracellular 5-HT, with elevations in dopamine occurring at high doses. Intravenous injection of either drug (1-10 micromol/kg) produced dose-related increases in 5-HT without affecting dopamine. d-Fenfluramine and mCPP exhibited similar potency in their ability to stimulate 5-HT efflux in vivo and interact with 5-HT transporters in vitro. When rats received high-dose d-fenfluramine or mCPP (10 or 30 micromol/kg, i.p., every 2 h, 4 doses), only d-fenfluramine treated rats displayed long-term 5-HT depletions. Thus, mCPP is a 5-HT releaser that does not appear to cause 5-HT depletion. Our data support the notion that 5 HT release per se may not be sufficient to produce the long-term 5-HT deficits associated with d-fenfluramine and other amphetamines. PMID- 11282250 TI - The serotonin reuptake inhibitor fluoxetine reduces sex steroid-related aggression in female rats: an animal model of premenstrual irritability? AB - The aggressive behavior displayed by some (but not all) female Wistar rats when an unfamiliar rat is being introduced into their home cage (the resident intruder paradigm) was found to be higher in non-receptive phases (metestrus, diestrus) than in the receptive phases (proestrus, estrus) of the estrus cycle, and effectively reduced by ovariectomy. When removal of the ovaries was followed by administration of estradiol and progesterone, in a regimen mimicking the normal cyclical release of these hormones, aggressive behavior was elicited, two days after estrus, in animals that had displayed aggressive behavior before ovariectomy, but not in those that had not. Short-term administration of a serotonin reuptake inhibitor (fluoxetine hydrochloride; 10 mg/kg, i.p.; 4-5 days) reduced both the aggressive behavior displayed during the diestrus phase by normally cycling rats, and the aggressive behavior elicited by administration of estradiol plus progesterone after ovariectomy. It is suggested that the aggressive behavior displayed by the female Wistar rat in the resident intruder paradigm may serve as an animal model of premenstrual dysphoria. PMID- 11282251 TI - Assessment of the serotonin and norepinephrine reuptake blocking properties of duloxetine in healthy subjects. AB - Duloxetine is a dual inhibitor of norepinephrine (NE) and serotonin (5-HT) uptake. Initial trials conducted in depressed patients using regimens of 20 mg/day or less did not convincingly demonstrate its efficacy as an antidepressant. The aim of this study was to assess the effects of duloxetine on the 5-HT and NE reuptake processes in healthy human volunteers. Twenty-seven healthy young males without a history of psychiatric disorder were randomly assigned to four groups, each group receiving one of the following daily drug regimens: placebo, clomipramine (a potent 5-HT/NE reuptake blocker) 100 mg/day, duloxetine 20 mg/day, or duloxetine 60 mg/day. In order to assess the NE reuptake process, the pressor response to intravenous tyramine (4 and 6 mg) was measured. Determination of the whole blood 5-HT content was used to evaluate the 5-HT reuptake blockade. These measurements were performed at baseline and repeated after 7 and 14 days of drug intake. Both duloxetine, at doses of 20 to 60 mg/day, and clomipramine significantly interfered with the 5-HT reuptake process, as demonstrated by marked decreases in blood 5-HT concentrations. However, the same doses of duloxetine, unlike clomipramine, failed to impede the usual increase in blood pressure that follows a tyramine intravenous infusion, indicating that clomipramine but not duloxetine blocked NE reuptake. At doses tested in a population of healthy volunteers, duloxetine acted as a selective 5-HT reuptake inhibitor, having no clear effect on the NE reuptake process. Nevertheless, given that the highest dose of duloxetine increased supine systolic blood pressure, it is possible that it represents the threshold regimen for NE reuptake inhibition. PMID- 11282252 TI - Serotonin 5-HT1A receptor binding potential declines with age as measured by [11C]WAY-100635 and PET. AB - Positron emission tomography (PET) and [11C]WAY-100635 were used to examine the effect of age on serotonin-1A (5-HT1A) receptor binding potential (BP) in 19 healthy subjects. Regions of interest (ROI) were drawn on the co-registered magnetic resonance imaging (MRI) in orbitofrontal (OFC), dorsolateral prefrontal (DLPFC), anterior cingulate (ACC), lateral (LTC), and mediotemporal (MTC), parietal, occipital and cerebellar cortex, and the raphe nuclei. BP values were calculated using a simplified reference tissue method. In addition, a voxelwise analysis was performed using SPM99. Voxelwise analysis revealed a significant global decrease of 5-HT(1A) BP with age (set level <.001). ROI analysis revealed significant age-related 5-HT(1A) BP decreases in DLPFC (r = -0.56), ACC (r = 0.44), OFC (r = -0.42), LTC (r = -0.40), parietal (r = -0.65), and occipital cortex (r = -0.43), but not in MTC or raphe nuclei. Overall, cortical 5-HT(1A) BP declined by approximately 10% per decade, except for the MTC, where we did not find a significant age effect. Hence, careful age matching may be recommended for future studies using PET and [11C]WAY-100635 to examine 5-HT1A receptors. PMID- 11282253 TI - Effects of antidepressant drugs on the behavioral and physiological responses to lipopolysaccharide (LPS) in rodents. AB - Antidepressants produce various immunomodulatory effects, as well as an attenuation of the behavioral responses to immune challenges, such as lipopolysaccharide (LPS). To explore further the effects of antidepressants on neuroimmune interactions, rats were treated daily with either fluoxetine (Prozac) or saline for 5 weeks, and various behavioral, neuroendocrine, and immune functions were measured following administration of either LPS or saline. Chronic fluoxetine treatment significantly attenuated the anorexia and body weight loss, as well as the depletion of CRH-41 from the median eminence and the elevation in serum corticosterone levels induced by LPS. Chronic treatment with imipramine also attenuated LPS-induced adrenocortical activation. In rats and in mice, which normally display a biphasic body temperature response to LPS (initial hypothermia followed by hyperthermia), chronic treatment with fluoxetine completely abolished the hypothermic response and facilitated and strengthened the hyperthermic response. The effects of antidepressants on the responsiveness to LPS are probably not mediated by their effects on peripheral proinflammatory cytokine production, because LPS-induced expression of TNFalpha and IL-1beta mRNA in the spleen (assessed by semiquantitative in situ hybridization) was not altered following chronic treatment with either fluoxetine or imipramine. The effects of antidepressants on the acute phase response may have important clinical implications for the psychiatric and neuroendocrine disturbances that are commonly associated with various medical conditions. PMID- 11282254 TI - Abnormal kainate receptor expression in prefrontal cortex in schizophrenia. AB - Abnormalities of molecules associated with the glutamate synapse have been implicated in the pathophysiology of schizophrenia. Of the many glutamate receptors, those most commonly suggested to be involved in schizophrenia are the ionotropic subtypes, the NMDA, AMPA, and kainate receptors. Both the NMDA and AMPA subtypes have been extensively studied in postmortem brains of individuals with schizophrenia, but relatively little is known about the expression of the kainate subtype of glutamate receptor. In this study, we have determined cortical and striatal kainate receptor expression in brains from persons with schizophrenia and a comparison group, using both in situ hybridization and receptor autoradiography. At the level of subunit mRNA expression, a shift in subunit stoichiometry was evident in multiple regions of the prefrontal cortex, with increased expression of gluR7 mRNA and decreased expression of KA2 mRNA. Decreased kainate receptor binding was also observed in the subjects with schizophrenia, but was restricted to infragranular laminae of the prefrontal cortex. No differences in kainate receptor binding or subunit mRNA levels were found in striatum or occipital cortex, suggesting that these findings may be restricted to association cortex. These data add to the growing literature implicating ionotropic glutamate receptor disturbances in schizophrenia, and indicate that in addition to AMPA and NMDA receptors, the kainate receptors are also abnormally expressed in this illness. PMID- 11282255 TI - The variable number of tandem repeats polymorphism of the dopamine transporter gene is not associated with significant change in dopamine transporter phenotype in humans. AB - A 40 base polymorphism of a variable number of tandem repeats (VNTR) has been described in the 3' untranslated region of the gene (SLC6A3) coding for the dopamine transporter (DAT). Despite being located in the untranslated region of the gene, this polymorphism has been associated with clinical phenotypes associated with dysregulation of dopamine transmission, such as attention deficit hyperactivity disorder and cocaine-induced paranoia. To examine the neurochemical phenotype associated with this polymorphism, we compared amphetamine-induced dopamine release (measured as displacement of the radiotracer [123I]IBZM) and DAT expression (measured with [123I]beta-CIT) in the striatum with Single Photon Computerized Emission Tomography (SPECT). Our sample included 59 subjects, 31 healthy controls and 29 patients with schizophrenia. No significant association was found between VNTR polymorphism and amphetamine-induced dopamine release or DAT density in the total sample, nor when each diagnostic group was considered separately. Thus, we did not replicate the findings of two previous studies, which had suggested that the 9 repeat allele was associated with either an increased or decreased DAT expression, albeit in different patient populations. PMID- 11282256 TI - Striatal dopaminergic and serotonergic markers in human heroin users. AB - To establish whether chronic opiate exposure might impair brain dopaminergic or serotonergic function in humans, we assessed biochemical indices of monoaminergic neurotransmitter activity and integrity in post mortem striatum of nine chronic heroin users and 14 control subjects. Striatal levels of the vesicular monoamine transporter were normal, suggesting that the density of dopamine nerve terminals is not reduced in heroin users. In nucleus accumbens, levels of tyrosine hydroxylase protein (-25%) and those of the dopamine metabolite homovanillic acid (-33%) were reduced significantly together with a trend for decreased dopamine ( 32%) concentration. These changes could reflect either a compensatory downregulation of dopamine biosynthesis in response to prolonged dopaminergic stimulation caused by heroin, or reduced axoplasmic transport of tyrosine hydroxylase. Striatal levels of serotonin were either normal or elevated whereas concentrations of the serotonin metabolite 5-hydroxyindoleacetic acid were decreased by 27-38%. Our data suggest that chronic heroin exposure might produce a modest reduction in dopaminergic and serotonergic activity that could affect motivational state and impulse control, respectively. PMID- 11282257 TI - Altered HPA axis responsivity to metyrapone testing in methadone maintained former heroin addicts with ongoing cocaine addiction. AB - Metyrapone testing, a provocation of hypothalamic-pituitary-adrenocortical (HPA) axis function, was performed in 39 in-patient subjects: 10 stable methadone maintained former heroin addicts without ongoing drug or alcohol abuse or dependence (MM), eight methadone- maintained former heroin addicts without ongoing drug or alcohol abuse or dependence other than ongoing cocaine dependence (C-MM), and 21 normal volunteers (NV). Plasma adrenocorticotrophic hormone (ACTH) levels were determined in samples drawn at 9A.M., just before administration of 2.25 g metyrapone orally and 4 and 8 hours afterward. Following metyrapone, C-MM had levels of ACTH that were significantly higher than both MM (p < .05) and NV (p < .01); whereas, MM and NV had levels that were comparable. Area under the plasma ACTH curves yielded similar results. This study documents hyper responsivity to removal of glucocorticoid negative feedback associated with cocaine addiction, even in the setting of methadone maintenance for heroin addiction, which here and previously has been shown to be associated with normalization of HPA axis function. PMID- 11282258 TI - Nicotine receptor inactivation decreases sensitivity to cocaine. AB - The reinforcing properties of nicotine and psychomotor stimulants are thought to be mediated through the mesolimbic dopamine (DA) system. This study investigates the role of high affinity nicotinic acetylcholine receptors (nAChRs) in cocaine place preference and examines some neurochemical changes in the mesolimbic DA system that might account for the interaction between nicotine and cocaine. 5 mg/kg is the lowest dose of cocaine able to condition a place preference in C57Bl/6 mice. Co-treatment with the nicotinic antagonist mecamylamine (1.0 mg/kg) disrupted place preference to 5 mg/kg cocaine. In addition, mice lacking the high affinity nAChR containing the beta2 subunit showed decreased place preference to 5 mg/kg cocaine, although higher doses of cocaine could condition a place preference in these knock out animals. In contrast, co-administration of a low dose of nicotine (0.2 mg/kg) potentiated place preference to a subthreshold dose of cocaine (3 mg/kg). DA turnover was monitored in several brain regions using tissue levels of DA and its primary metabolite DOPAC as an indication of DA release. Wild type mice showed decreased DA turnover following treatment with 5 mg/kg cocaine; whereas, this response was not seen in mice lacking the beta2 subunit of the nAChR. Induction of chronic fos-related antigens by cocaine was also reduced in mutant mice as compared to their wild type siblings, implying that downstream actions of cocaine were also affected by inactivation of the high affinity nAChR. These data indicate that activation of the high affinity nAChR may contribute to cocaine reinforcement. PMID- 11282259 TI - Comparison of the effects of ketamine and memantine on prolactin and cortisol release in men. a randomized, double-blind, placebo-controlled trial. AB - N-methyl D-aspartate (NMDA)-antagonists decrease neurotoxicity by inhibiting Ca2+ influx which is of interest for the treatment of acute cerebrovascular insults and chronic neurodegenerative disorders. Currently, there is no surrogate marker for quantification of NMDA-receptor-mediated drug effects, which hampers dose finding clinical studies. As prolactin and cortisol liberation is in part influenced through NMDA-receptors we investigated whether the elevation of prolactin or cortisol plasma levels is a class effect of NMDA-antagonists and might be an appropriate marker for studying NMDA-antagonistic potency. Fifteen healthy male volunteers participated in this placebo-controlled, randomized, three-way crossover trial. Ketamine (0.5mg/kg), memantine (0.16 mg/kg; i.e., a well tolerated standard dose) or placebo were infused over 60 min. Ketamine increased serum prolactin and cortisol levels (p < 0.001), whereas memantine and placebo did not affect hormone levels. Further studies are needed to define whether higher doses of memantine or other NMDA antagonists can induce hormone release. PMID- 11282260 TI - Lipid-based amphotericin B: a review of the last 10 years of use. AB - The last decade has been remarkable for the dramatic increase in the prevalence of serious fungal infections in patients with haematological disorders and neutropenic cancer patients. The mortality rate of deep-seated infection has been in excess of 90% and there is no doubt that this is one of the greatest challenges currently facing haematologists and oncologists. The development of the lipid-based drugs - liposomal amphotericin (AmBisome(R)), amphotericin B lipid complex, ABLC (Abelcet(R)), amphotericin B colloidal dispersion, Amphocil (ABCD(R)), has meant that doses of amphotericin B can be safely escalated for the first time whilst the problems of nephrotoxicity, infusion related reactions (including chills, rigors, fevers and hypoxia) can be reduced. These toxicities are variably reduced with AmBisome more than Abelcet and more than Amphocil and there is little information from randomised trials other than for AmBisome. AmBisome used in the setting of persistent fever and neutropenia not responding after 3-4 days of intravenous antibiotics, is associated with less breakthrough systemic fungal infections. There is also much less need for premedication, including steroids, compared with amphotericin B and Abelcet. The use of intermittent doses of Ambisome given prophylactically is now being explored. A new and exciting era of antifungal therapy is opening up with new compounds, such as itraconazole voriconazole, posaconazole and echinocandins, being investigated and for the first time, we also have options for combination therapy and prophylaxis. PMID- 11282261 TI - Antibiotic resistance in the absence of selective pressure. AB - Antibiotic resistance poses a serious threat to modern medical practice making treatment more difficult and is associated with increased mortality among patients infected with resistant organisms. There is clear evidence that acquisition of resistance is associated with a decrease in the fitness of the organisms at least in the short term. Evidence from in vitro experiments indicates that bacteria have the ability to adapt to this deficit and recover fitness on serial passage. More recent results show that identical organisms isolated from patients in outbreaks have an initial deficit but that adaptation occurs in vivo. Strategies directed towards controlling resistance must move beyond wishful thinking that supposes that these organisms will disappear merely with control of prescribing. In some cases, resistance will not disappear because there is no evolutionary disadvantage in being resistant once adaptation has taken place. It is important, therefore, that we direct our efforts towards preventing primary resistance emerging and in limiting the spread of resistant strains. Ultimately, we must look again to new drug discovery to improve our therapeutic armoury. PMID- 11282262 TI - An open, randomised, multi-centre study comparing the safety and efficacy of sitafloxacin and imipenem/cilastatin in the intravenous treatment of hospitalised patients with pneumonia. AB - This was a phase II, randomised, open-label, multi-centre study to assess the safety, tolerability, and efficacy of sitafloxacin (DU-6859a, 400 mg once daily) compared with imipenem (imipenem/cilastatin, 500 mg three times daily) in the treatment of hospitalised patients with pneumonia. Patients (n=69) were entered into the study in the intent-to-treat group, 35 in the sitafloxacin and 34 in the imipenem group. Patients (n=65) were included in the clinically evaluable population and 42 in the bacteriologically evaluable population. Baseline demographic data and clinical characteristics were similar for both treatment groups and across all patient populations. The incidence, severity and type of adverse events were similar in both treatment groups. The frequency of adverse events, which were considered to be related to the study of drugs was low and generally similar between the two groups. Mild transient increases in alanine aminotransferase and alkaline phosphatase occurred in the sitafloxacin treatment group, but there were no apparent trends in the other serum enzyme levels. The clinical response at the first and second follow-up assessments indicated that 94 97% of patients in the clinically evaluable population and 91% of patients in the intent-to-treat population were classified as cured in both treatment groups. The bacteriological response was classified as satisfactory for all patients (100%) in the bacteriologically evaluable population in the imipenem treatment group and satisfactory for 90 and 95% of cases at the first and second follow-up assessments in the bacteriologically evaluable population in the sitafloxacin treatment group, respectively. In conclusion, for the treatment of pneumonia, sitafloxacin was considered as safe and as tolerable as imipenem and preliminary data from this study suggest that it may have similar efficacy. PMID- 11282263 TI - Management of upper respiratory tract infections in primary care in Italy: a national survey. AB - This prospective study, carried out in Italy during the winter of 1998 by the means of questionnaires, was designed to investigate the diagnostic and therapeutic approach of the Italian general practitioners (GPs) to the management of acute upper respiratory tract infections (URTIs) in adult outpatients. A total of 354 GPs were questioned about ten adult patients each who had visited the surgery with an URTI requiring an antibiotic prescription. Our data showed there was a tendency to prescribe antibiotics only on the basis of clinical diagnosis, microbiological investigations being required very rarely. Orally administered antibiotics were preferred and compliance with the number of daily doses strongly influenced the antibiotic prescription. In patients affected by more severe infections, injectable antibiotics were frequently prescribed. PMID- 11282264 TI - Antibiotic resistance rates and macrolide resistance phenotypes of viridans group streptococci from the oropharynx of healthy Greek children. AB - A total of 200 isolates of viridans group streptococci isolated from the oropharynx of healthy Greek children were studied. Vancomycin, rifampicin, fluoroquinolones and dalfopristin/quinupristin were active against all tested isolates. High level resistance to gentamicin was not seen. Intermediate and high level penicillin resistance was present in 28.5 and 14.5% isolates, respectively, with 41.3% of the latter group, being also resistant to cefotaxime. Resistance rates to other antimicrobials were as follows - erythromycin 38.5%, clarithromycin 33.5%, clindamycin 7.5% and tetracycline 23%. Penicillin resistance occurred more frequently in Streptococcus mitis isolates, while macrolide resistance was more frequent in S. oralis. MLSB resistance phenotype M was dominant (74%) among erythromycin resistant isolates, with phenotypes IR and CR being represented by 6 and 20% of isolates, respectively. PMID- 11282265 TI - In vitro activity of mezlocillin, meropenem, aztreonam, vancomycin, teicoplanin, ribostamycin and fusidic acid against Borrelia burgdorferi. AB - The in vitro susceptibility profile of Borrelia burgdorferi is not yet well defined for several antibiotics. Our study explored the in vitro susceptibility of B. burgdorferi to mezlocillin, meropenem, aztreonam, vancomycin, teicoplanin, ribostamycin and fusidic acid. Minimal inhibitory concentrations (MICs) and minimal borreliacidal concentrations (MBCs) were measured using a standardised colorimetric microdilution method and conventional subculture experiments. MIC values were lowest for mezlocillin (MIC(90), < or =0.06 mg/l) and meropenem (MIC(90), 0.33 mg/l). Vancomycin (MIC(90), 0.83 mg/l) was less effective in vitro. Borreliae proved to be resistant to aztreonam (MIC(90), >32 mg/l), teicoplanin (MIC(90), 6.6 mg/l), ribostamycin (MIC(90), 32 mg/l), and fusidic acid (MIC(90), >4 mg/l). The mean MBCs resulting in 100% killing of the final inoculum after 72 h of incubation were lowest for mezlocillin (MBC, 0.83 mg/l). This study gathered further data on the in vitro susceptibility patterns of the B. burgdorferi complex. The excellent in vitro effectiveness of acylamino penicillin derivatives and their suitability for the therapy of Lyme disease is emphasised. PMID- 11282266 TI - Antimicrobial resistance of Enterococci in Lebanon. AB - There is little information on the types of Enterococcus spp and their antibiotic resistance patterns in Lebanon. One hundred and fifty-three consecutive clinical enterococcal isolates collected between 1998 and 1999 were tested against 11 antimicrobial agents using disk diffusion and the Etest. The isolates were identified by conventional methods and API-Strep and were found to consist of Enterococcus faecalis (72.5%), Enterococcus faecium (22.9%), Enterococcus avium (3.2%) and Enterococcus gallinarum (1.3%). The percent of resistant strains of E. faecalis and E. faecium respectively were, ampicillin 0.9 and 14%, erythromycin 59% and 40%, tetracycline 72% and 34%, chloramphenicol 32 and 11%, rifampin 36% and 57%, ciprofloxacin, 23% and 34%, norfloxacin 22 and 8%. High level aminoglycoside (HLA) resistance was found in 19% E. faecalis and 9% E. faecium for gentamicin and 36% and 26% for streptomycin. Excellent correlation was observed between the high level disk tests and the Etest in the detection of HLA resistance but not with the regular disks. None of the isolates showed resistance to vancomycin or teicoplanin except for one E. gallinarum isolate which showed intermediate resistance (MIC 16 mg/l) to vancomycin. These variable antimicrobial rates of resistance suggest a surveillance programme for antimicrobial resistance in this country would be helpful to help control infection, guide empirical antibiotic therapy and implement a policy of antibiotic usage. PMID- 11282267 TI - Antimicrobial susceptibility of Listeria monocytogenes isolated from meningoencephalitis in sheep. AB - The antimicrobial susceptibility to different antimicrobial agents of 41 Listeria monocytogenes strains isolated from sheep with meningoencephalitis and from feedstuff was tested by both microdilution and disk diffusion methods. Both sets of isolates of L. monocytogenes were susceptible to penicillin G, amoxicillin, cephalothin, erythromycin, vancomycin, rifampicin, gentamicin, kanamycin, trimethoprim, sulfisoxazole, chloramphenicol and ciprofloxacin, but resistant to tetracycline and doxycycline (7.3 and 4.9%, respectively). Tetracycline was the most frequent resistance trait in L. monocytogenes strains of animal origin. Four strains (9.8%) also exhibited reduced susceptibility (MIC 4 mg/l) to doxycycline suggesting the need of surveillance studies to monitor the antimicrobial resistance of Listeria strains of animal origin. PMID- 11282268 TI - Pharmacokinetics of intravenously administered pefloxacin in the prostate; perspectives for its application in surgical prophylaxis. AB - In an attempt to define whether intravenously administered pefloxacin might be appropriate for surgical prophylaxis in prostatectomy, 50 patients undergoing transvesical prostatectomy for benign prostate hyperplasia were given a single intravenous dose of 800 mg; surgery was then performed after 2, 4, 6, 8 or 10 h. Concentrations of pefloxacin were determined in serum and in both the centre and periphery of the prostate adenoma using a microbiological plate assay. Elevated concentrations of pefloxacin were found in the adenoma from 2 h onwards. The central and peripheral concentrations were similar and had a mean value of 4.39 microg/g of tissue. These concentrations were similar to those achieved in serum. Although concentrations of pefloxacin were not determined separately in the intercellular, interstitial or excreted fluid, the tissue levels found were well above the MICs of pefloxacin for the bacteria commonly causing acute and chronic prostatitis. These data suggest the intravenous administration of pefloxacin to be a satisfactory alternative for the surgical prophylaxis before prostatectomy as well as in the therapy of acute prostatitis. PMID- 11282269 TI - Enhancement of antibiotic activity against poly-drug resistant Mycobacterium tuberculosis by phenothiazines. AB - Phenothiazines have been shown to inhibit the in vitro growth of multi-drug resistant (resistant to rifampicin and isoniazid) Mycobacterium tuberculosis (MDRTB). They have been considered as potential adjuvants to regimens employing four or more antibiotics for the management of freshly diagnosed infections of M. tuberculosis in patients from areas known to have a high prevalence of MDRTB. Chlorpromazine has been shown to enhance the activity of antibiotics (except ethambutol) to which M. tuberculosis is susceptible. This might result in a reduction in the dose of some or all of the antibiotics employed without sacrificing the integrity of treatment. Chlorpromazine, thioridazine and promethazine were shown to enhance the activity of rifampicin and streptomycin when used in combinations at concentrations that are minimally effective when employed separately against clinical strains of M. tuberculosis resistant to two or more antibiotics (poly-drug resistant MTB). The phenothiazines had no effect on the activity of isoniazid against poly-drug resistant MTB. PMID- 11282271 TI - Evolution of metronidazole and tetracycline susceptibility pattern in Helicobacter pylori at a hospital in Saudi Arabia. AB - The association of Helicobacter pylori with chronic gastritis and peptic ulcer disease led to new therapeutic approaches including the use of antibiotics. Recently, resistance of H. pylori to antibiotics has emerged as the major cause of treatment failure. This retrospective analysis was aimed at investigating the development of antimicrobial susceptibility patterns amongst H. pylori strains isolated at King Fahd Hospital of the University, Al-Khobar. Susceptibility patterns obtained using isolates from a pilot study (1987-1988) were compared with those subsequently isolated (1990-1996). Metronidazole resistance was estimated to be 35.2% in the first period but more than doubled (78.5%) during the second period. Isolates from females showed a higher resistance rate to metronidazole than those from males. Only one strain was tetracycline resistant. The extremely high resistance rate to metronidazole indicates that it may not be very effective for eradication of H. pylori. Tetracycline should prove a useful component of treatment regimens in this geographical region. PMID- 11282270 TI - In vitro activity of four fluoroquinolones against Mycobacterium tuberculosis. AB - The in vitro activity of ciprofloxacin, ofloxacin, levofloxacin and moxifloxacin against strains of Mycobacterium tuberculosis was studied. Moxifloxacin and levofloxacin showed the greatest activity having an MIC(90) of 1 mg/l. The MIC(90) for ofloxacin was 2 mg/l and for ciprofloxacin 4 mg/l. Further studies should be made to determine the role played by these compounds in the treatment of tuberculosis. PMID- 11282272 TI - An in vitro study of the efficacy of rifampicin and minocycline coated umbilical venous catheters. AB - The use of antibiotic coated catheters has been proposed as a means of reducing catheter related sepsis. In this study, an in vitro comparison of bacterial colonisation rates was made between uncoated umbilical venous catheters and catheters coated with rifampicin and minocycline. The following parameters were determined; the direct antimicrobial effect of coated and uncoated catheter segments against a range of organisms associated with line sepsis, the assessment of the decline in antimicrobial activity in coated catheters immersed in plasma and the inhibitory efficacy of the catheters to colonisation over a 28-day period. Minocycline and rifampicin coated umbilical catheters showed a superior inhibitory effect and prevented colonisation with the commoner line-related organisms, when compared with uncoated catheters. The inhibitory effect declined after 14 days in the human plasma. Resistance to colonisation in vitro may not extend beyond 21 days. PMID- 11282273 TI - Steroid receptors and metastatic potential in endometrial cancers. AB - The relative overexpression of estrogen receptor (ER)-alpha exon 5 splicing variant, the disrupted synchronization of ER-beta and ER-alpha expressions, and the suppression of progesterone receptor (PR) form A expression as a transcriptional repressor might be related to metastatic potential of uterine endometrial cancers, leading to poor patient prognosis related to estrogen refractoriness. PMID- 11282274 TI - Transcription of androgen receptor and 5alpha-reductase II in genital fibroblasts from patients with androgen insensitivity syndrome. AB - Impaired virilisation during embryonic development and pubertal arrest in patients with androgen insensitivity syndromes (AIS) is usually caused by mutations in the androgen receptor (AR)- or the 5alpha-reductase II (5RII) gene. However, identical mutations may lead to strikingly different phenotypes. To investigate whether this may be caused by individually altered transcription rates in fibroblasts from the genital region (GF) from affected patients, we applied competitive reverse transcribed PCRs (competitive RT-PCR) targeting AR- and 5RII-transcripts. We could demonstrate that AR- and 5RII-mRNA concentrations in cells from patients with partial and complete AIS and missense mutations in the AR- or 5RII-gene are normal or only moderately lowered compared to equally aged normal controls. However, in a patient bearing a premature stop-codon in the AR-gene a considerably lowered AR-transcript level was detected. We conclude, that in patients with incomplete virilisation disorders due to missense mutations, transcription regulation of AR and 5RII generally follows normal patterns. Accordingly, the premature stop-codon found in one patient's AR-gene may rather cause reduced transcript stability than an impairment of transcription activity. Therefore, altered AR- and 5RII-transcription rates in fibroblasts from the GF do not seem to be the cause for the variable genotype-phenotype correlation in androgen insensitivity syndrome. PMID- 11282275 TI - Cloning and cellular localization of the canine progesterone receptor: co localization with growth hormone in the mammary gland. AB - The mammary gland has been found to express the gene encoding growth hormone (GH) in several species. Within the mammary gland, it may act as an autocrine/paracrine growth factor for cyclic epithelial changes, and may be a determinant in mammary carcinogenesis. In the dog, progestins enhance mammary GH expression. To elucidate the mechanism of progestin-induced mammary GH expression, the canine progesterone receptor (PR) is characterized and the cellular localization of the PR in normal and tumorous mammary tissues is examined. Sequence analysis of the canine PR revealed two in-frame ATG codons, encoding a putative PR-B protein of 939 amino acids and a putative PR-A protein of 765 amino acids. Western blot analysis indicated that both isoforms occur in uterus and mammary gland issues. Immunohistochemical analysis of the PR revealed that the PR was differentially expressed in mammary tissue, with many PR-positive epithelial cells in the proliferation phase of the glandular tissue and a low number of PR-positive cells in differentiated mammary tissue. Stromal and myoepithelial cells had no specific PR staining. Mammary tumours had a variety of staining patterns, including no staining, normal nuclear staining, marked heterogeneous immunoreactivity and perinuclear staining of tumorous epithelial cells and cytoplasmic-staining of spindle cells. Double staining showed that all GH-producing cells were positive for PR, whereas not all PR containing cells stained for GH. It is concluded that the activated PR may transactivate GH expression in the mammary gland within the same cell and functions as a pre requisite transcription factor. However, during malignant transformation this regulation may be lost. PMID- 11282276 TI - Econazole and miconazole inhibit steroidogenesis and disrupt steroidogenic acute regulatory (StAR) protein expression post-transcriptionally. AB - The imidazole antifungal drugs econazole and miconazole have previously been shown to disrupt steroidogenesis in Leydig and adrenal cells by inhibiting 17alpha-hydroxylase/17,20-lyase (P450c17) enzyme activity, thus reducing the conversion of progesterone to androstenedione. However, a recent study in Y-1 adrenal cells indicated that these compounds may also reduce the availability of cholesterol to the cytochrome P450 side chain cleavage (P450(scc)) enzyme, the first enzyme in the steroidogenic pathway. Since the steroidogenic acute regulatory protein (StAR) mediates the transfer of cholesterol from the outer to the inner mitochondrial membrane where the P450(scc) enzyme resides, an action which constitutes the rate-limiting and acutely-regulated step in steroidogenesis, we hypothesized that these drugs may also reduce StAR expression and/or activity. Our studies demonstrate that these drugs reversibly inhibited (Bu)(2)cAMP-stimulated progesterone production in a dose- and time-dependent manner in MA-10 cells without affecting total protein synthesis or P450(scc) and 3beta-hydroxysteroid dehydrogenase (3beta-HSD) enzyme expression or activity. In contrast, they dramatically decreased (Bu)(2)cAMP-stimulated StAR protein expression post-transcriptionally. This study indicates that StAR protein is susceptible to inhibition by at least some imidazole compounds that inhibit steroidogenesis. PMID- 11282277 TI - Neuroactive steroids, their precursors, and polar conjugates during parturition and postpartum in maternal and umbilical blood: 1. Identification and simultaneous determination of pregnanolone isomers. AB - A rapid method for the identification and measurement of four pregnanolone isomers and their polar conjugates in human plasma was developed using a simple quadrupole GC/MS system with electron impact ionization. Steroid levels were measured in the plasma of 13 and three women at delivery with subarachnoidal and epidural analgesia, respectively, and in corresponding samples of umbilical plasma. A good correlation (r=0.94, P<0.001, n=8) was found between the allopregnanolone in maternal plasma determined by GC/MS and that measured by RIA. Epipregnanolone (3beta-hydroxy-5beta-pregnan-20-one) was identified and measured for the first time in human plasma; its concentration in both maternal and umbilical plasma was much lower than that of other pregnanolone isomers. The levels of 3beta-hydroxy-pregnanolone isomers were significantly higher in the umbilical plasma than in the maternal plasma, while the differences in 3alpha hydroxy-isomers were insignificant. The differences in conjugates were insignificant except in the case of allopregnanolone, the levels of which were lower in umbilical plasma. In all of the pregnanolone isomers, a significantly lower conjugated/unconjugated steroid ratio was found in the umbilical plasma than in the maternal plasma. The possible role of the sulfatation of pregnanolone isomers around parturition is discussed. PMID- 11282278 TI - Steroid sulfatase activity in the rat ovary, cultured granulosa cells, and a granulosa cell line. AB - Direct production of gonadal steroids from sulfated adrenal androgens may be an important alternative or complementary pathway for ovarian steroidogenesis. The conversion of sulfated adrenal androgens, present in serum at micromolar concentrations in adult women, into unconjugated androgens or estrogens requires steroid sulfatase (STS) activity. STS activity has not been characterized in the rat ovary. Substantial STS activity was present in homogenates of rat ovaries, primary cultures of rat granulosa cells, and a granulosa cell line, as determined by conversion of radiolabeled estrone sulfate (E1S) to unconjugated estrone. The potent inhibitor estrone sulfamate eliminated the STS activity. Using E1S as a substrate with microsomes prepared from a granulosa cell line, the K(m) of STS activity was approximately 72 microM, a value in agreement with previously published data for rat STS. Therefore, ovarian cells possess STS and can remove the sulfate from adrenal androgens such as dehydroepiandrosterone sulfate (DHEA S). Using DHEA-S as a steroidogenic substrate represents an alternative model for the production of ovarian steroids versus the "two cell, two gonadotropin" model of ovarian estrogen synthesis, whereby thecal cells produce androgens from substrate cholesterol and granulosa cells convert the androgens into estrogens. The relative contribution of STS activity to ovarian steroidogenesis remains unclear but may have important physiological and pathophysiological implications. PMID- 11282279 TI - Inhibition of steroid sulphatase activity by tricyclic coumarin sulphamates. AB - The identification of the active pharmacophore required for potent inhibition of steroid sulphatase activity, i.e. an aryl-O-sulphamate structure, has led to the synthesis and testing of a large number of 1-4 ring-based inhibitors. 4 Methylcoumarin-7-O-sulphamate (COUMATE) was one of the first non-steroid based inhibitors identified. In an attempt to increase the potency of this class of inhibitor a series of tricyclic COUMATEs (665-6615 COUMATEs) have been synthesised and evaluated. Using placental microsomes as a source of oestrone sulphatase (E1-STS) the size of the third ring of the tricyclic COUMATEs was found to have a marked effect on inhibitor potency. Whereas 665- and 6615 COUMATEs had IC(50)s of 200 and 370 nM, respectively, the most potent inhibitor in vitro in this series was 6610 COUMATE with an IC(50) of 1 nM. Selected inhibitors were tested for their in vivo potency by administration of a single dose (0.1 or 1 mg/kg, p.o.) to female rats. Surprisingly, in vivo 6615 COUMATE proved to be the most active drug, inhibiting rat liver E1-STS activity by 23 and 94% when assayed 24 h after administration of the 0.1 and 1 mg/kg doses. E1-STS activity in brain tissue and white blood cells was also found to be inhibited when selected drugs were tested. These studies have identified a number of tricyclic COUMATEs with therapeutic potential. PMID- 11282281 TI - Genistein affects lipogenesis and lipolysis in isolated rat adipocytes. AB - Genistein is a phytoestrogen found in several plants eaten by humans and food producing animals and exerting a wide spectrum of biological activity. In this experiment, the impact of genistein on lipogenesis and lipolysis was studied in isolated rat adipocytes. Incubation of the cells (10(6) cells/ml in plastic tubes at 37 degrees C with Krebs-Ringer buffer, 90 min) with genistein (0.01, 0.3, 0.6 and 1 mM) clearly restricted (1 nM) [U-14C]glucose conversion to total lipids in the absence and presence of insulin. When [14C]acetate was used as the substrate for lipogenesis, genistein (0.01, 0.1 and 1 mM) exerted a similar effect. Thus, the anti-lipogenetic action of genistein may be an effect not only of alteration in glucose transport and metabolism, but this phytoestrogen can also restrict the fatty acids synthesis and/or their esterification. Incubation of adipocytes with estradiol at the same concentrations also resulted in restriction of lipogenesis, but the effect was less marked. Genistein (0.1 and 1 mM) augmented basal lipolysis in adipocytes. This process was strongly restricted by insulin (1 microM) and H-89 (an inhibitor of protein kinase A; 50 microM) and seems to be primarily due to the inhibitory action of the phytoestrogen on cAMP phosphodiesterase in adipocytes. Genistein at the smallest concentration (0.01 mM) augmented epinephrine-stimulated (1 microM) lipolysis but failed to potentiate lipolysis induced by forskolin (1 microM) or dibutyryl-cAMP (1 mM). These results suggest genistein action on the lipolytic pathways before activation of adenylate cyclase. The restriction of lipolysis stimulated by several lipolytic agents--epinephrine, forskolin and dibutyryl-cAMP were observed when adipocytes were incubated with genistein at highest concentrations (0.1 and 1 mM). These results prove the inhibitory action of this phytoestrogen on the final steps of the lipolytic cascade, i.e. on protein kinase A or hormone sensitive lipase. Estradiol, added to the incubation medium, did not affect lipolysis. It can be concluded that genistein significantly affects lipogenesis and lipolysis in isolated rat adipocytes. PMID- 11282280 TI - Regulation of steroid sulphatase expression and activity in breast cancer. AB - Steroid sulphatase (STS) catalyzes the conversion of oestrone sulphate (E1S) to oestrone (E1) and its action in breast tumours makes a major contribution to in situ oestrogen production in this tissue. Although expression of STS mRNA and STS activity are increased in malignant breast tissues compared with that in non malignant tissues, little is known about the regulation of its expression or activity. In the present study we have used a RT-PCR technique to investigate the regulation of STS mRNA expression in cultured breast tissue fibroblasts and MCF-7 cells. STS mRNA expression was readily detectable in fibroblasts derived from breast tissue proximal to tumours, breast tumour tissue and reduction mammoplasty tissue. For two pre-menopausal subjects, STS mRNA expression was similar in proximal and tumour fibroblasts whereas for a third, post-menopausal subject, expression in breast tumour fibroblasts was 2.4-fold that in proximal fibroblasts. The cytokine tumour necrosis factor alpha (TNFalpha) or the STS inhibitor, 2-methoxyoestrone-3-O-sulphamate, had no effect on STS mRNA expression in fibroblasts. STS mRNA was detectable in MCF-7 cells but neither TNFalpha nor interleukin 6 (IL-6) affected its expression. Transient transfection of COS-1 and MCF-7 cells with a STS cDNA lacking STS 5' and 3' sequences increased activity 17 fold and 2-fold, respectively. TNFalpha plus IL-6 increased STS activity in mock transfected MCF-7 cells and further increased STS activity in transfected MCF-7 cells. This indicates that activation can occur independently of STS promoter and enhancer elements. In conjunction with the lack of regulation of STS mRNA it suggest that TNFalpha and IL-6 may increase STS activity via a post-translational modification of the enzyme or by increasing substrate availability. PMID- 11282282 TI - Immunomodulatory 7-hydroxylated metabolites of dehydroepiandrosterone are present in human semen. AB - Seminal fluid represents a milieu enabling spermatozoa to break the ovum membrane and suppress its immune response and, at the same time, to protect male germ cells against infection. Among constituents of the seminal fluid, various steroids, including dehydroepiandrosterone (DHEA) and its sulphate, were detected. With respect to immunomodulatory and antioxidative properties of the latter steroid and its 7-hydroxylated metabolites, believed to be at least in some instances the locally active species, their presence in seminal fluid is of particular interest. Here for the first time unconjugated 3beta,7alpha-dihydroxy 5-androsten-17-one (7alpha-OH-DHEA) and its 7beta-hydroxyisomer have been detected and quantified in semen. Eight semen samples were extracted with diethyl ether and following evaporation and solvent partition both isomers were detected by gas chromatography-mass fragmentometry using the ions m/z 358 and 343 for quantification. Another portion was separated by HPLC and in the fractions corresponding to 7-OH-DHEA isomers the steroids were measured by recently developed specific radioimmunoassays (RIA). Mean concentrations of 7-OH-DHEA as measured by RIA amounted 5.75+/-1.29 and 5.39+/-0.75 nmol/l (mean+/-SEM) for 7alpha- and 7beta-OH-DHEA, respectively. PMID- 11282283 TI - Activation of AMPA receptors inhibits prolactin and estradiol secretion and delays the onset of puberty in female rats. AB - Previous experiments have evidenced the neuroendocrine role of AMPA receptors. Present studies were carried out to obtain information on the role of these receptors in the control of the onset of puberty. To this end, female rats were i.c.v. injected with vehicle or AMPA (agonist of AMPA receptors: 0.1 or 0.5 nmol/day) between 26 and 30 days (Experiment 1), or 30 and 34 days (Experiment 2) of age. Serum concentrations of PRL, LH and estradiol were measured before drug administration, 10 min after the last injection, at vaginal opening (VO) and at first estrus (FE) presentation. In both experiments, AMPA administration inhibited PRL and estradiol secretion without affecting LH release. When AMPA was administered between 26 and 30 days a significant delay in the day of vaginal opening was observed. These results confirmed the inhibitory effect of AMPA on PRL secretion and suggests a role of AMPA receptors in the control of puberty onset. PMID- 11282284 TI - Glucocorticoids suppress human immunodeficiency virus type-1 long terminal repeat activity in a cell type-specific, glucocorticoid receptor-mediated fashion: direct protective effects at variance with clinical phenomenology. AB - Glucocorticoid administration and/or excess secretion have been associated with increased Human Immunodeficiency Virus Type-1 (HIV-1) replication and AIDS progression. The HIV-1 long terminal repeat (LTR) promoter contains glucocorticoid-responsive element (GRE)-like sequences that could mediate a positive effect of glucocorticoids on HIV-1. In addition, we recently demonstrated that the HIV-1 accessory protein Vpr is a potent coactivator of the glucocorticoid receptor, which, like the host coactivator p300, potentiates the effect of glucocorticoids on GRE-containing, glucocorticoid-responsive genes. Such an effect may increase the sensitivity of several host target tissues to glucocorticoids by several fold, and may, thus, contribute to a positive effect of glucocorticoids on the HIV-1-LTR in infected host cells. In this study, we determined the direct effect of glucocorticoids on HIV-1-LTR by examining the ability of dexamethasone to modulate the activity of this promoter coupled to the luciferase reporter gene in human cell lines. Dexamethasone markedly inhibited Tat-stimulated, p300- or Vpr-enhanced luciferase activities in a cell-type specific, dose-dependent, and glucocorticoid receptor-mediated fashion. This effect of dexamethasone was not potentiated by Vpr, was antagonized by the glucocorticoid receptor antagonist RU 486 and required the DNA-binding domain of the receptor. These data suggest that the inhibitory effect of glucocorticoids on the HIV-1-LTR may be exerted via non-GRE-dependent inhibition of the strongly positive host transcription factor NF-kappaB, which interacts with the DNA- and ligand-binding domains of the receptor. Alternatively, it is also possible that dexamethasone-activated glucocorticoid receptor competes with other transcription factors for their binding sites on the promoter region or squelches transcription factors shared by HIV-1-LTR and glucocorticoid-responsive promoters. We conclude that glucocorticoids suppress, rather than stimulate, the HIV-1 promoter, thus acting, protectively for the host. Their apparent negative clinical association with AIDS is most likely due to immunosuppression of the host. PMID- 11282285 TI - Expression of cytochrome P450(11beta) (11beta-hydroxylase) gene during gonadal sex differentiation and spermatogenesis in rainbow trout, Oncorhynchus mykiss. AB - Androgens and especially 11-oxygenated androgens are known to be potent masculinizing steroids in fish. As a first step to study their physiological implication in gonadal sex differentiation in fish, we cloned a testicular cytochrome P450(11beta) (11beta-hydroxylase) cDNA in the rainbow trout, Oncorhynchus mykiss. We isolated a 1882 bp P450(11beta) cDNA (rt11betaH2, AF217273) which contains an open reading frame encoding a 552 putative amino acids protein. This sequence was highly homologous (98% in nucleotides and 96.5% in amino acids) to another rainbow trout P450(11beta) sequence (AF179894) and also to a Japanese eel P450(11beta) (68% in amino acids). Northern blot analysis detected a single transcript of 2 kb which was highly expressed in the testis (stage II) and to a lesser degree in the anterior kidney (containing the interrenal tissue). No signal was detected in the posterior kidney, brain, liver, skin, intestine and heart. In the testis this transcript was highly expressed at the beginning of spermatogenesis (stages I and II), followed by a decrease during late spermatogenesis (stages III to V). By semi-quantitative reverse transcription polymerase chain reaction, P450(11beta) expression during gonadal differentiation was estimated to be at least 100 times higher in male than in female gonads. This difference was first detected at 55 days post-fertilization (dpf), i.e. 3 weeks before the first sign of histological sex differentiation, and was sustained long after differentiation (127 dpf). Specific P450(11beta) gene expression was also demonstrated before testis differentiation (around 50 dpf) using virtual Northern blot, with no expression detected in female differentiating gonads. From these results, and also based on the already known actions of 11-oxygenated androgens in testicular differentiation in fish, it is now suggested that P450(11beta) gene expression is a key factor for the testicular differentiation in rainbow trout. PMID- 11282286 TI - Inhibitory effects of stress-activated nitric oxide on antioxidant enzymes and testicular steroidogenesis. AB - The messenger role of nitric oxide (NO) in immobilization stress-induced inhibition of testicular steroidogenesis has been previously suggested. In accord with this, here, we show that the intratesticular injection of isosorbide dinitrate (ISDN; 2x2.5 mg/testis), an NO donor, mimicked the action of stress on serum testosterone concentrations and hCG-stimulated testosterone production in rat testicular tissue. When added in vitro, ISDN inhibited testicular 3beta hydroxysteroid dehydrogenase and 17alpha-hydroxylase/lyase. Immobilization stress and injections of ISDN also decreased the activity of catalase, glutathione peroxidase, glutathione transferase, and glutathione reductase in the interstitial compartment of testis. When stressed rats were treated concomitantly with bilateral intratesticular injections of N(omega)-nitro-L-arginine methyl ester, a non-selective NOS inhibitor (2x600 microg/testis), the activities of antioxidative enzymes, as well as serum testosterone concentration, were partially normalized. These results indicate that stress-induced stimulation of the testicular NO signalling pathway leads to inhibition of both steroidogenic and antioxidant enzymes. PMID- 11282287 TI - Possible involvement of ceramide in the regulation of rat Leydig cell function. AB - In the present study, a possible role of a ceramide-dependent pathway in the regulation of Leydig cell function was investigated. Intracellular ceramide levels were increased by: (a) adding ceramide analogs; (b) inhibiting ceramidase activity; and (c) adding sphingomyelinase (SMase). The cell-permeable ceramide analogs N-acetyl-, N-hexanoyl- and N-octanoylsphingosine (C2, C6 and C8) were used. As inhibitor of ceramidase activity 1S,2R-D-erythro-2-(N-myristoylamino)-1 phenyl-1-propanol (MAPP) was used. Sphingomyelinase from S. aureus origin was utilized. Leydig cells were cultured for 3 or 24 h with or without the different drugs (10 microM) and SMase (0.3 U/ml) in the presence or absence of hCG (10 ng/ml). Basal testosterone production was not modified under any of the experimental conditions. A decrease in hCG-stimulated testosterone production was observed at 3 and 24 h in all cases. The inactive analog (N-hexanoyl dihydrosphingosine) did not produce inhibition in hCG-stimulated testosterone production. TNFalpha and IL1beta, two possible inducers of sphingomyelin hydrolysis, produced similar effects on hCG-stimulated testosterone production. In experiments performed in the presence of C6, inhibition in hCG-stimulated cAMP production was observed. The inhibitory effect of ceramide was also observed in dbcAMP-stimulated cultures indicating that this pathway inhibits post-cAMP formation events. To study possible loci for the action of ceramide on the steroidogenic pathway, cells were incubated with C6 and MAPP in the presence of different testosterone precursors. The drugs inhibited testosterone produced from 22(R)-hydroxycholesterol (22R-OHChol), pregnenolone and 17alpha hydroxyprogesterone (17OHP4) but not from androstenedione (Delta4). These results suggest that a ceramide-dependent pathway regulates hCG-stimulated Leydig cell steroidogenesis at the level of cAMP production and at post-cAMP events. PMID- 11282288 TI - Highly sensitive and specific time-resolved fluoroimmunoassay (TR-FIA) of cyproterone acetate and free cyproterone. AB - We have developed a non-isotopic TR-FIA for Cyproterone acetate and Cyproterone in plasma. Synthesis of the biotinylated tracers, biotinylated Cyproterone acetate, and Cyproterone, as well as the preparation of anti-Cyproterone acetate and anti-Cyproterone antisera are reported. The specificity of anti-Cyproterone acetate antiserum resulting from the coupling of link bridge (link bridge between steroid and BSA), on the 3-position on the steroid skeleton, allowed to carry out the Cyproterone acetate assay directly on extracted plasma (without chromatography). On the other hand Cyproterone assays require a purification step, including extraction plus chromatography, because the plasma Cyproterone acetate concentrations in Cyproterone acetate-treated women are 200 times higher than for Cyproterone. Theses plasma TR-FIA of Cyproterone acetate and Cyproterone presented the advantage of needing only small doses of radioactivity for recovery controls, and better practicability related to the only existing RIA described to date. PMID- 11282289 TI - Association between estradiol, estrogen receptors, total lipids, triglycerides, and cholesterol in patients with benign and malignant breast tumors. AB - This study addresses the correlation between the levels of estradiol (E2), total lipids, triglycerides, and cholesterol in serum and tissue samples of age-matched patients with benign (40 cases; 16 were premenopausal and 24 were postmenopausal) and malignant (50 cases; 17 were premenopausal and 33 were postmenopausal) breast tumors. Estradiol levels were determined in serum and cytosol, estrogen receptors (ER) were assayed in cytosol, and total lipids, triglycerides and cholesterol were determined in serum and membrane fractions of all benign and malignant breast disease patients. Serum E2 was significantly higher in malignant cases than benign ones (P<0.05) with a significant reduction (40%) in postmenopausal than premenopausal women. ER-positive tumors were significantly higher in postmenopausal women with malignant breast tumors than benign cases (P<0.05). Tissue levels of total lipids, triglycerides, and cholesterol were highly significantly increased in breast cancer women than women with benign breast diseases (P<0.05, P<0.005 and P<0.05 respectively) and they were also significantly correlated with estradiol levels. It could be concluded that the uptake of lipids from plasma by the tumor tissue is greatly correlated to estradiol and it may confirm the possible role of lipids as risk factor in breast cancer. PMID- 11282290 TI - Androgen and estrogen stimulation of ornithine decarboxylase activity in mouse kidney. AB - In the present work, the activity of mouse renal ornithine decarboxylase (ODC) from CBA female mice was used as a biological marker to detect (anti)androgenic activity of different groups of endocrine disruptors and steroids. Daily injections of testosterone or dihydrotestosterone (DHT) into 60 day old female mice for 4 days increased renal ODC activity in a dose-dependent manner that reached up to 100-fold (testosterone) or 250-fold (DHT) above the baseline when the highest dose, 200 microg/mouse, was used. Administration of flutamide concurrently with testosterone (75 microg/mouse) caused a potent decrease of ODC induction in a dose-dependent manner, suppressing the enzyme activity at the doses of 0.1 and 0.5 mg/mouse by about 88 and 95%, respectively. In contrast, estradiol at the doses of 0.5 and 1 mg/mouse induced a significant stimulation of renal ODC activity in a dose-dependent manner when it was given alone or in combination with testosterone. Using a sensitive increase in ODC activity in response to androgens as an end point, we did not detect an antiandrogenic effect of several antiandrogens, such as cyproterone acetate, spironolactone, p,p'DDE and vinclozolin. Also, none of these antiandrogens were able to change the basal level of renal ODC activity, with the exception of cyproterone acetate that at a dose of 0.1 mg/mouse stimulated ODC activity. The data obtained suggest that mouse renal ODC from CBA females is not strictly androgen-specific and cannot be used for estimation of antiandrogenic effects of compounds having an affinity to different types of receptors. PMID- 11282291 TI - Cholesterol sulphate sulphohydrolase from human placenta microsomes--purification and properties of the dephosphorylated form of enzyme. AB - The procedure for purification of cholesterol sulphate sulphohydrolase (ChS-ase) from human placenta microsomes was elaborated. The highly purified enzyme preparation (specific activity 2000 nmol x min(-1) x mg protein(-1)) exhibited optimal activity at pH 9.0. The K(m) value was established to be 1.5+/-0.85 x 10( 5) M. The high molecular weight form (200 kDa) and the low molecular weight form (20 kDa) of the enzyme were separated. The interconversion of the high molecular weight variant into the low one occurs under the influence of dephosphorylation. Both forms exhibited typical Michaelis-Menten saturation kinetics. The effect of different compounds on the enzyme activity was tested. PMID- 11282292 TI - Eph receptors and ephrin ligands. essential mediators of vascular development. AB - The molecular and cellular mechanisms governing vascular development are still poorly understood. Prominent among the intercellular signals that control the initial establishment of the vascular network (termed vasculogenesis) and the subsequent remodeling process (called angiogenesis) are soluble ligands that signal through receptor tyrosine kinases (RTKs). Recent reports have added cell bound ephrin ligands and their cognate Eph RTKs to the list of key players in vascular development. PMID- 11282293 TI - DGAT and triglyceride synthesis: a new target for obesity treatment? AB - Because triglycerides are considered essential for survival and their synthesis has been thought to occur through a single mechanism, inhibiting triglyceride synthesis has been largely unexplored as a possible target for obesity treatment. However, recent studies indicate that mice lacking acyl CoA:diacylglycerol acyltransferase (DGAT), a key enzyme in triglyceride synthesis, are viable and resistant to diet-induced obesity. Unexpectedly, this resistance is caused by a mechanism involving increased energy expenditure. These findings suggest that inhibiting specific components of triglyceride synthesis, such as DGAT, is feasible and may represent a novel approach to treating obesity. PMID- 11282294 TI - Platelets and the injured vessel wall-- "rolling into action": focus on glycoprotein Ib/V/IX and the platelet cytoskeleton. AB - Blood platelets play a key role in maintaining the integrity of the vascular system through their ability to arrest bleeding (haemostasis) and promote repair of injured blood vessels. Considerable progress has been made in the last few years in our understanding of the adhesion mechanisms utilized by platelets to adhere to sites of vascular injury. Studies have helped define the precise role of von Willebrand factor (vWf), and its platelet receptor, glycoprotein (GP) Ib/V/IX, in initiating platelet-vessel wall and platelet-platelet adhesion contacts. In addition to its adhesive role, recent studies have highlighted the importance of GPIb/V/IX in regulating the cytoskeleton of platelets. GPIb/V/IX not only maintains the normal cytoskeletal architecture of resting platelets but also induces cytoskeletal reorganization following engagement of vWf. Somewhat surprisingly, the physical link between GPIb/V/IX and the membrane skeleton does not appear necessary for GPIb/V/IX-induced cytoskeletal reorganization. In contrast, this linkage appears critical for GPIb/V/IX to maintain cell adhesion under high shear and also for the ability of the cytoskeleton to exert negative regulatory effects on the GPIb-vWf interaction. Thus a complex functional relationship appears to exist between GPIb/V/IX and the membrane skeleton that goes well beyond preserving the normal cytoskeletal architecture of resting platelets. PMID- 11282295 TI - Activation of factor IX by factor XIa. AB - Blood coagulation factor IX is activated during hemostasis by two distinct mechanisms. Activation through factor VIIa/tissue factor occurs early in the course of fibrin clot formation. Activation by factor XIa appears to be important for maintaining the integrity of the clot over time. In general, coagulation proteases are activated on a phospholipid surface in the presence of a protein cofactor. Until recently, activation of factor IX by factor XIa was thought to be the exception to this rule, as phospholipid has no effect on the reaction and no cofactor had been identified. These curious observations suggest that factor IX is activated by factor XIa in the fluid phase. A large amount of new evidence now indicates that factor IX activation by factor XIa occurs on the surface of activated platelets. The data suggest, however, that this reaction differs significantly from other protease-substrate interactions on the platelet surface. This is likely to be due, in part, to the unusual structure of the factor XI molecule. PMID- 11282296 TI - Regulation of the cardiac repolarizing HERG potassium channel by protein kinase A. AB - Protein kinase A is an enzyme that regulates many cellular processes and is activated in many pathological conditions such as stress and various types of heart failure. Recently it has been shown that protein kinase A couples functionally to the HERG cardiac potassium channel, thereby altering repolarization in the heart. This link between a repolarizing potassium channel and the protein kinase system of cardiac cells may contribute to arrhythmogenesis and may become a target for future approaches to antiarrhythmic therapy. PMID- 11282297 TI - Ligands for peroxisome proliferator-activated receptor inhibit monocyte CCR2 expression stimulated by plasma lipoproteins. AB - Substantial evidence supports a causal role for monocyte chemoattractant protein 1 (MCP-1) and its receptor, CCR2, in the recruitment of monocytes from the circulation into atherosclerotic lesions. MCP-1 is produced and secreted by virtually every cellular component of the vessel wall. It generally is assumed that the magnitude of the monocyte chemotactic activity, which is initiated by the functional activation of CCR2 by MCP-1, is directly proportional to the concentration of the chemoattractant. However, we recently demonstrated that an inflammatory response of monocytes is finely regulated and also depends on the expression levels of CCR2. We identified plasma low-density lipoprotein (LDL) as a positive regulator and showed that it greatly increased monocyte CCR2 gene expression. In contrast, activation of peroxisome proliferator-activated receptor by synthetic ligands or components of oxidized LDL reduces monocyte CCR2 expression and blocks chemotaxis mediated by MCP-1. We hypothesized that the excessive monocyte accumulation in the vessel wall during atherogenesis may result in part from an enhanced chemotactic response. These findings suggest CCR2 gene expression in circulating monocytes as a potential additional target for intervention and prevention of atherosclerosis. PMID- 11282298 TI - Cytokines as emerging targets in the treatment of heart failure. AB - Recent studies have identified the importance of biologically active molecules such as neurohormones in disease progression in heart failure. More recently it has become apparent that in addition to neurohormones another portfolio of biologically active molecules termed cytokines are also expressed in the setting of heart failure. This article reviews recent clinical and experimental material which suggest that the cytokines such as tumor necrosis factor (TNF), interleukin 1 (IL-1) and interleukin-6 (IL-6) may represent another class of biologically active molecules that are responsible for the development and progression of heart failure. In addition, we also review the early results from clinical trials that have utilized various targeted anti-cytokine strategies in patients with heart failure. PMID- 11282299 TI - Angiogenesis and bone growth. AB - Vascularization of the growth plate region represents a key mechanism for the coupling of two fundamental processes determining the rate of bone growth, chondrogenesis (cartilage production) and osteogenesis (bone formation). Precise coupling is crucial during periods of rapid bone growth or fracture repair in adults, and changes in the balance might induce pathologic conditions such as osteoarthritis and ectopic bone formation. During the formation of the growth plates of long bones, there is a close and dynamic interaction between developing vascular structures and the cartilage, which is one of the least vascular tissues in the body. Recent experimental findings provide an explanation why the close proximity of cartilage and vasculature is mutually exclusive: vascular invasion of cartilage is associated with chondrocyte apoptosis and consequently, inhibition of angiogenesis in the growth plate delays chondrocyte cell death, resulting in a massive expansion in the number of hypertrophic cartilage cells in the growth plate. The fundamental importance of chondrocytes in the growth, development and repair of the skeleton has led to intense investigation of the mechanisms that regulate chondrocyte maturation and apoptosis. PMID- 11282300 TI - Human urotensin-II, the most potent mammalian vasoconstrictor identified to date, as a therapeutic target for the management of cardiovascular disease. AB - The novel cyclic undecapeptide human urotensin-II (hU-II) and its high-affinity G protein-coupled receptor, GPR14, are both expressed within the human cardiovasculature (vascular smooth muscle, endothelium, myocardium, coronary atheroma, etc.) and may, therefore, contribute to the (patho)physiological regulation of cardiovascular homeostasis in humans. Indeed, hU-II is an efficacious, sustained spasmogen of mammalian isolated blood vessels including those from rats, rabbits, dogs, pigs, non-human primates and humans (where it is one to two orders of magnitude more potent than endothelin(ET)-1). In vivo, hU-II markedly alters systemic hemodynamics in the anesthetized primate (increase cardiac contractility [dP/dt], increase stroke volume, decrease total peripheral resistance) ultimately resulting in fatal cardiovascular collapse. As such, the development of selective hU-II receptor antagonists may be of utility in the management of cardiovascular disorders characterized by aberrant vasoconstriction, myocardial dysfunction and/or cardiac remodeling (e.g., myocardial infarction, congestive heart failure). PMID- 11282301 TI - PPAR signaling in the control of cardiac energy metabolism. AB - Cardiac energy metabolic shifts occur as a normal response to diverse physiologic and dietary conditions and as a component of the pathophysiologic processes which accompany cardiac hypertrophy, heart failure, and myocardial ischemia. The capacity to produce energy via the utilization of fats by the mammalian postnatal heart is controlled in part at the level of expression of nuclear genes encoding enzymes involved in mitochondrial fatty acid beta-oxidation (FAO). The principal transcriptional regulator of FAO enzyme genes is the peroxisome proliferator activated receptor alpha (PPARalpha), a member of the ligand-activated nuclear receptor superfamily. Among the ligand activators of PPARalpha are long-chain fatty acids; therefore, increased uptake of fatty acid substrate into the cardiac myocyte induces a transcriptional response leading to increased expression of FAO enzymes. PPARalpha-mediated control of cardiac metabolic gene expression is activated during postnatal development, short-term starvation, and in response to exercise training. In contrast, certain pathophysiologic states, such as pressure overload-induced hypertrophy, result in deactivation of PPARalpha and subsequent dysregulation of FAO enzyme gene expression, which sets the stage for abnormalities in cardiac lipid homeostasis and energy production, some of which are influenced by gender. Thus, PPARalpha not only serves a critical role in normal cardiac metabolic homeostasis, but alterations in PPARalpha signaling likely contribute to the pathogenesis of a variety of disease states. PPARalpha as a ligand-activated transcription factor is a potential target for the development of new therapeutic strategies aimed at the prevention of pathologic cardiac remodeling. PMID- 11282302 TI - Detailed molecular model of apolipoprotein A-I on the surface of high-density lipoproteins and its functional implications. AB - The major apolipoprotein (apo) A-I containing lipoprotein, high- density lipoprotein, is a negative risk factor for cardiovascular disease. An atomic resolution molecular model for lipid-associated apo A-I was recently proposed in which two apo A-I molecules are wrapped beltwise around a small discoidal patch of phospholipid bilayer. Because of its detailed predictions of lipid-associated apo A-I structure, this molecular belt model, if confirmed, provides a blueprint for understanding the molecular mechanisms of reverse cholesterol transport, and thus for the rational design of new classes of drugs for reversal of atherosclerosis and cardiovascular disease. The details and implications of the model are currently being explored by site-directed mutagenesis. PMID- 11282303 TI - Alpha4 integrins in cardiovascular development and diseases. AB - Alpha4 integrins (alpha4beta1 and alpha4beta7) have a restricted distribution pattern and are critical for the development and diseases of the cardiovascular system. alpha4 integrins support unique biological properties such as promoting cell migration and inhibiting cell spreading and focal adhesion formation. We have found that the alpha4 integrin subunit directly and tightly binds to a signaling adapter molecule, paxillin, and disruption of the alpha4-paxillin interaction interferes with many of alpha4-dependent biological functions. Consequently, the interaction of alpha4 integrins with paxillin may play an important role in regulating alpha4-mediated functions. This review focuses on what we have known about the alpha4-paxillin interaction and discusses the possible mechanism of regulation for this interaction. PMID- 11282304 TI - Left-right determination. AB - Recent advances have given us new insights into the molecular basis of organ position. A gene cascade that determines left-right positioning of organ primordia has emerged. In here we present the current knowledge of the molecular determinants of organ positioning during vertebrate embryogenesis. PMID- 11282305 TI - Regulators of G-protein signaling in receptor complexes. AB - G protein signaling pathways regulate heart development and adult cardiac function. G protein activity is controlled by the interplay between receptor catalyzed activation and the inhibitory regulators of G protein signaling (RGS) proteins. Most RGS proteins are GTPase accelerating proteins (GAPs) for Gi and Gq class G protein alpha subunits, and thereby terminate signaling. However, RGS proteins also provide scaffolding properties to help assemble or maintain signaling complexes. Thus, RGS proteins are kinetic regulators that may sharpen both signal activation and termination. The five subfamilies of mammalian RGS proteins contain a characteristic RGS domain and distinct flanking domains that convey lipid and/or protein interactions within receptor complexes. The RGS domain provides GAP activity and additional interactions with the receptor complex. Distantly related RGS-like (RGL) proteins provide other regulatory and effector functions in G protein signaling pathways. RGS and RGL proteins provide exciting new therapeutic targets to combat cardiovascular diseases. PMID- 11282306 TI - How calcium influx through calcium leak channels is responsible for the elevated levels of calcium-dependent proteolysis in dystrophic myotubes. AB - Duchenne muscular dystrophy patients lack the protein dystrophin which is an essential link in the complex of proteins that connect the cytoskeleton to the extracellular matrix. In mechanically stressed tissues such as muscle, transient sarcolemmal microdisruptions are normal, but in dystrophic muscle cells the frequency of these microdisruptions is greatly increased. Although both normal and dystrophic cells are able to actively repair these microdisruptions, calcium entry through the more frequent sarcolemmal microdisruptions of dystrophic cells results in an increased calcium-dependent proteolysis that alters the activity of the calcium leak channel. The accumulation of abnormally active calcium leak channels over time results in a gradual loss of calcium homeostasis and eventual cell death. PMID- 11282307 TI - Psychopathy and developmental instability. AB - Psychopaths are manipulative, impulsive, and callous individuals with long histories of antisocial behavior. Two models have guided the study of psychopathy. One suggests that psychopathy is a psychopathology, i.e., the outcome of defective or perturbed development. A second suggests that psychopathy is a life-history strategy of social defection and aggression that was reproductively viable in the environment of evolutionary adaptedness (EEA). These two models make different predictions with regard to the presence of signs of perturbations or instability in the development of psychopaths. In Study 1, we obtained data on prenatal, perinatal, and neonatal signs of developmental perturbations from the clinical files of 643 nonpsychopathic and 157 psychopathic male offenders. In Study 2, we measured fluctuating asymmetry (FA, a concurrent sign of past developmental perturbations) in 15 psychopathic male offenders, 25 nonpsychopathic male offenders, and 31 male nonoffenders. Psychopathic offenders scored lower than nonpsychopathic offenders on obstetrical problems and FA; both psychopathic and nonpsychopathic offenders scored higher than nonoffenders on FA. The five offenders from Study 2 meeting the most stringent criteria for psychopathy were similar to nonoffenders with regard to FA and had the lowest asymmetry scores among offenders. These results provide no support for psychopathological models of psychopathy and partial support for life-history strategy models. PMID- 11282308 TI - Facial attractiveness signals different aspects of "quality" in women and men. AB - We explored the relationships between facial attractiveness and several variables thought to be related to genotypic and phenotypic quality in humans (namely fluctuating asymmetry (FA), body mass index (BMI), health, age). To help resolve some controversy around previous studies, we used consistent measurement and statistical methods and relatively large samples of both female (n=94) and male (n=95) subjects (to be evaluated and measured), and female (n=226) and male (n=153) viewers (to rate attractiveness). We measured the asymmetry of 22 traits from three trait families (eight facial, nine body and five fingerprint traits) and constructed composite asymmetry indices of traits showing significant repeatability. Facial attractiveness was negatively related to an overall asymmetry index in both females and males, with almost identical slopes. Female facial attractiveness was best predicted by BMI and past health problems, whereas male facial attractiveness was best predicted by the socioeconomic status (SES) of their rearing environment. Composite FA indices accounted for a small (<4%) but usually significant percentage of the variation in facial attractiveness in both sexes, when factors related to asymmetry were controlled statistically. We conclude that, although facial attractiveness is negatively related to developmental instability (as measured by asymmetry), attractiveness also signals different aspects of "quality" in the two sexes, independent of FA. PMID- 11282309 TI - Hadza meat sharing. AB - In most human foraging societies, the meat of large animals is widely shared. Many assume that people follow this practice because it helps to reduce the risk inherent in big game hunting. In principle, a hunter can offset the chance of many hungry days by exchanging some of the meat earned from a successful strike for shares in future kills made by other hunters. If hunting and its associated risks of failure have great antiquity, then meat sharing might have been the evolutionary foundation for many other distinctively human patterns of social exchange. Here we use previously unpublished data from the Tanzanian Hadza to test hypotheses drawn from a simple version of this argument. Results indicate that Hadza meat sharing does not fit the expectations of risk-reduction reciprocity. We comment on some variations of the "sharing as exchange" argument; then elaborate an alternative based partly on the observation that a successful hunter does not control the distribution of his kill. Instead of family provisioning, his goal may be to enhance his status as a desirable neighbor. If correct, this alternative argument has implications for the evolution of men's work. PMID- 11282310 TI - The influence of whole-body vs. torso pre-cooling on physiological strain and performance of high-intensity exercise in the heat. AB - Little research has been reported examining the effects of pre-cooling on high intensity exercise performance, particularly when combined with strategies to keep the working muscle warm. This study used nine active males to determine the effects of pre-cooling the torso and thighs (LC), pre-cooling the torso (ice-vest in 3 degrees C air) while keeping the thighs warm (LW), or no cooling (CON: 31 degrees C air), on physiological strain and high-intensity (45-s) exercise performance (33 degrees C, 60% rh). Furthermore, we sought to determine whether performance after pre-cooling was influenced by a short exercise warm-up. The 45 s test was performed at different (P<0.05) mean core temperature [(rectal+oesophageal)/2] [CON: 37.3+/-0.3 (S.D.), LW: 37.1+/-0.3, LC: 36.8+/-0.4 degrees C] and mean skin temperature (CON: 34.6+/-0.6, LW: 29.0+/-1.0, LC: 27.2+/ 1.2 degrees C) between all conditions. Forearm blood flow prior to exercise was also lower in LC (3.1+/-2.0 ml 100 ml tissue(-1) x min(-1)) than CON (8.2+/-2.5, P=0.01) but not LW (4.3+/-2.6, P=0.46). After an exercise warm-up, muscle temperature (Tm) was not significantly different between conditions (CON: 37.3+/ 1.5, LW: 37.3+/-1.2, LC: 36.6+/-0.7 degrees C, P=0.16) but when warm-up was excluded, T(m) was lower in LC (34.5+/-1.9 degrees C, P=0.02) than in CON (37.3+/ 1.0) and LW (37.1+/-0.9). Even when a warm-up was performed, torso+thigh pre cooling decreased both peak (-3.4+/-3.8%, P=0.04) and mean power output (-4.1+/ 3.8%, P=0.01) relative to the control, but this effect was markedly larger when warm-up was excluded (peak power -7.7+/-2.5%, P=0.01; mean power -7.6+/-1.2%, P=0.01). Torso-only pre-cooling did not reduce peak or mean power, either with or without warm-up. These data indicate that pre-cooling does not improve 45-s high intensity exercise performance, and can impair performance if the working muscles are cooled. A short exercise warm-up largely removes any detrimental effects of a cold muscle on performance by increasing Tm. PMID- 11282312 TI - Hydration effects on thermoregulation and performance in the heat. AB - During exercise, sweat output often exceeds water intake, producing a water deficit or hypohydration. The water deficit lowers both intracellular and extracellular fluid volumes, and causes a hypotonic-hypovolemia of the blood. Aerobic exercise tasks are likely to be adversely effected by hypohydration (even in the absence of heat strain), with the potential affect being greater in hot environments. Hypohydration increases heat storage by reducing sweating rate and skin blood flow responses for a given core temperature. Hypertonicity and hypovolemia both contribute to reduced heat loss and increased heat storage. In addition, hypovolemia and the displacement of blood to the skin make it difficult to maintain central venous pressure and thus cardiac output to simultaneously support metabolism and thermoregulation. Hyperhydration provides no advantages over euhydration regarding thermoregulation and exercise performance in the heat. PMID- 11282311 TI - Effect of pre-cooling, with and without thigh cooling, on strain and endurance exercise performance in the heat. AB - Body cooling before exercise (i.e. pre-cooling) reduces physiological strain in humans during endurance exercise in temperate and warm environments, usually improving performance. This study examined the effectiveness of pre-cooling humans by ice-vest and cold (3 degrees C) air, with (LC) and without (LW) leg cooling, in reducing heat strain and improving endurance performance in the heat (35 degrees C, 60% RH). Nine habitually-active males completed three trials, involving pre-cooling (LC and LW) or no pre-cooling (CON: 34 degrees C air) before 35-min cycle exercise: 20 min at approximately 65% VO2peak then a 15-min work-performance trial. At exercise onset, mean core (Tc, from oesophagus and rectum) and skin temperatures, forearm blood flow (FBF), heart rate (HR), and ratings of exertion, body temperature and thermal discomfort were lower in LW and LC than CON (P<0.05). They remained lower at 20 min [e.g. Tc: CON 38.4+/-0.2 (+/ S.E.), LW 37.9+/-0.1, and LC 37.8+/-0.1 degrees C; HR: 177+/-3, 163+/-3 and 167+/ 3 b.p.m.), except that FBF was equivalent (P=0.10) between CON (15.5+/-1.6) and LW (13.6+/-1.0 ml.100 ml tissue(-1) x min(-1)). Subsequent power output was higher in LW (2.95+/-0.24) and LC (2.91+/-0.25) than in CON (2.52+/-0.28 W kg( 1), P=0.00, N=8), yet final Tc remained lower. Pre-cooling by ice-vest and cold air effectively reduced physiological and psychophysical strain and improved endurance performance in the heat, irrespective of whether thighs were warmed or cooled. PMID- 11282313 TI - The importance of aerobic fitness in determining tolerance to uncompensable heat stress. AB - When protective clothing is worn that restricts evaporative heat loss, it is not valid to assume that the higher sweat rates associated with improvements in aerobic fitness will increase heat tolerance. An initial study compared thermoregulatory and cardiovascular responses to both compensable and uncompensable heat stress before and after 8 weeks of endurance training in previously sedentary males. Despite a 15% improvement in VO2peak, and lower heart rates and rectal temperature (T(re)) responses while wearing combat clothing, no changes were noted when subjects wore a protective clothing ensemble. Tolerance times were unchanged at approximately 50 min. A subsequent short-term training model that used daily 1-h exercise sessions for 2 weeks also failed to show any benefit when the protective clothing was worn in the heat. Cross-sectional comparisons between groups of high and low aerobic fitness, however, have revealed that a high aerobic fitness is associated with extended tolerance time when the protective clothing is worn. The longer tolerance time is a function of both a lower starting T(re) and a higher T(re) tolerated at exhaustion. Improvements in cardiovascular function with long-term training may allow higher core temperatures to be reached prior to exhaustion. Conversely, elevations in core temperature that occur with normal training sessions may familiarize the more fit subjects to the discomforts of exercise in the heat. Other factors such as differences in body fatness may account for a faster increase in tissue temperature at a given metabolic rate for less fit individuals. PMID- 11282314 TI - Performance enhancement in rally car drivers via heat acclimation and race simulation. AB - To investigate the combined use of an interactive racecar simulator and heat acclimation on psychomotor (driving) performance, eight rally drivers underwent 4 days of repeated heat (50 degrees C) exposure (1 h x day(-1)) during which they performed a simulated rally drive (3x12-min stages each separated by a 2-min break), after first cycling for 15 min at 125 W to induce some degree of fatigue and heat storage prior to beginning the rally. During the rally stages, a generic set of pace notes were read to the subject by a co-driver. In each simulation, sweat loss, heart rate, core (rectal) and skin temperatures were recorded and driving and psychomotor performance were assessed by recording stage times and time to complete a psychomotor test. Levels of physiological and perceived thermal strain were also recorded. Significant decreases in rally stage times (88 s; P<0.005), psychomotor test time (18 s; P<0.01), final core (0.25 degrees C; P<0.001) and skin (0.44 degrees C; P<0.005) temperatures, heart rate (16 beats x min(-1); P<0.05) and physiological (15 W x m(-2); P<0.005) and perceived thermal (3.7 units; P<0.01) strain were evident by the end of the final simulation, and a significant (P<0.05) increase in sweat sensitivity (+0.33 g x h(-1) x degrees C( 1)) was also recorded. These results suggest that both heat acclimation and race simulation can improve the psychomotor performance of rally drivers, although the relative contribution of each factor was not determined here. However, in a practical setting, these factors would not be used in isolation. After performing the acclimation and simulation protocol prior to an actual rally, drivers subjectively reported improvements in tolerating a high thermal load and in their ability to control the rally vehicle. PMID- 11282315 TI - The combined effect of heat and carbon monoxide on the performance of motorsport athletes. AB - Two of the major stressors endured by a motorsport athlete (MSA) during a racing event are the effects of heat and carbon monoxide. To date, there has been little research into their combined effect on driving performance. Using an interactive racecar simulator located within an environmental chamber, subjects drove a simulated race (60 min) in environmental conditions similar to those that develop during a NASCAR Winston Cup oval track race (50 degrees C ambient temperature and 10-12% carboxyhaemoglobin levels). Subjects also completed cool (20 degrees C) and heat only (50 degrees C) race simulations. During the simulations, oxygen consumption, heart rate, core and skin temperatures and psychomotor performance were measured. The results demonstrated that exposure to a racecar micro environment combining both heat and CO produced significantly greater (P<0.05) sweat loss and core temperature change (1.53 kg; 1.06 degrees C) when compared to the heat only (1.14 kg; 0.73 degrees C) and cool conditions (0.35 kg; 0.09 degrees C). Furthermore, a significant decrease (P<0.05) in psychomotor performance was also shown between the heat/CO condition (contact points=38), and both the heat only (25 points) and cool conditions (22 points). It follows that lengthy exposure to these two stressors could produce a substantial decrement in driving performance, thereby endangering the MSA and other race competitors. PMID- 11282316 TI - Evaluation of cognitive performance in the heat by functional brain imaging and psychometric testing. AB - Military operations in tropical environments have imposed a significant challenge to the Australian Defence Forces (ADF). The hot and humid conditions are known to cause debilitating effects on soldiers deployed to northern regions of Australia, with the consequence that the effectiveness and efficiency of operations are severely compromised. While the adverse effects of thermal stress on soldiers' physiological capability are well established, this has not been confirmed for cognitive performance. A select range of psychometric tests were administered and functional brain electrical activity imaging was performed to investigate the impact of thermal stress on cognitive performance. The brain electrical activity of subjects was measured while undertaking a range of cognitive tasks. Steady State Probe Topography (SSPT), a novel brain imaging technology, was employed to monitor the changes in regional brain activity and neural processing speed of subjects under thermal stress. The psychometric test batteries included the following tasks; Rey Auditory Verbal Learning Test; Inspection Time; Digit Span test; a spatial working memory task; and the AX-continuous performance task. These tasks measure a range of cognitive processes including attention, memory, verbal learning, information processing and concentration. The functional brain imaging provided topographical information, which showed changes in electrical activity in response to thermal stress during cognitive performance. These changes in brain electrical activity and neural speed induced by thermal stress may help to identify the type of cognitive functions that are likely to be impaired under operational conditions. Results indicated that subjects experienced increasing cardiovascular strain through thermally neutral to thermally straining conditions. The results from the psychometric test battery showed some promising effects given the small sample size including deficits in working memory, in information retention and in information processing. There was also marked differences in the electrical responses of the brain when subjects were thermally strained. The Steady-State Visual Evoked Potential recordings showed an increase in amplitude and a decrease in latency, suggesting an increase in the utilisation of neural resources or effort by subjects to maintain the same level of performance as under thermally neutral conditions. The data are suggestive of the high sensitivity of brain imaging techniques with high temporal resolution to identify important decrements in cognitive performance in hostile environments. PMID- 11282317 TI - Nutritional needs for exercise in the heat. AB - Although hot conditions are not typically conducive to optimal sports performance, nutritional strategies play an important role in assisting an athlete to perform as well as possible in a hot environment. A key issue is the prevention of hypohydration during an exercise session. Fluid intake strategies should be undertaken in a cyclical sequence: hydrate well prior to the workout, drink as much as is comfortable and practical during the session, and rehydrate aggressively afterwards in preparation for future exercise bouts. There is some interest in hyperhydration strategies, such as hyperhydration with glycerol, to prepare the athlete for a situation where there is little opportunity for fluid intake to match large sweat losses. Recovery of significant fluid losses after exercise is assisted by the simultaneous replacement of electrolyte losses. Carbohydrate (CHO) requirements for exercise are increased in the heat, due to a shift in substrate utilization towards CHO oxidation. Daily food patterns should focus on replacing glycogen stores after exercise, and competition strategies should include activities to enhance CHO availability, such as CHO loading for endurance events, pre-event CHO intake, and intake of sports drinks in events lasting longer than 60 min. Although CHO ingestion may not enhance the performance of all events undertaken in hot weather, there are no disadvantages to the consumption of beverages containing 4-8% CHO and electrolytes. In fact, the palatability of these drinks may enhance the voluntary intake of fluid. Although there is some evidence of increased protein catabolism and cellular damage due to production of oxygen radicals during exercise in the heat, there is insufficient evidence to make specific dietary recommendations to account for these issues. PMID- 11282318 TI - Effects of whole body cooling on sensory perception and manual performance in subjects with Raynaud's phenomenon. AB - Patients with Raynaud's phenomenon (RP) have abnormal digital vasoconstriction in response to cold. The aim of the study was to investigate the effects of cooling on sensory perception and manual performance in healthy male subjects and subjects with RP. There were two groups of subjects with primary RP: 12 subjects fulfilled the criteria of Lewis (L) and the other 12 the more critical criteria of Maricq (M). Control group (C) consisted of 19 healthy men. Subjects were exposed to 5 degrees C for 60 min. Skin temperatures were measured. Finger dexterity, pinch strength, abduction/adduction of fingers, pressure perception threshold and vibration perception threshold were tested during the exposure every 15 min. At the beginning of the exposure the mean (S.E.) finger temperature was 2.5 (1.2) degrees C (P<0.05) lower in M than in C. Manual performance and sensory perception were impaired due to the cooling, the impairment being significantly greater in M than in C. Responses of L were between those of M and C. In a given finger temperature vibration and pressure sensibility and manual performance were lower in M and L than in C. In conclusion, cold exposure decreased sensory perception and manual performance in the subjects with RP to a lower level than in the healthy subjects. Non-thermal factors may also decrease performance in RP. PMID- 11282319 TI - Effects of metabolic rate on thermal responses at different air velocities in -10 degrees C. AB - The effects of exercise intensity on thermoregulatory responses in cold (-10 degrees C) in a 0.2 (still air, NoWi), 1.0 (Wi1), and 5.0 (Wi5) m x s(-1) wind were studied. Eight young and healthy men, preconditioned in thermoneutral (+20 degrees C) environment for 60 min, walked for 60 min on the treadmill at 2.8 km/h with different combinations of wind and exercise intensity. Exercise level was adjusted by changing the inclination of the treadmill between 0 degrees (lower exercise intensity, metabolic rate 124 W x m(-2), LE) and 6 degrees (higher exercise intensity, metabolic rate 195 W x m(-2), HE). Due to exercise increased heat production and circulatory adjustments, the rectal temperature (T(re)), mean skin temperature (Tsk) and mean body temperature (Tb) were significantly higher at the end of HE in comparison to LE in NoWi and Wi1, and T(re) and Tb also in Wi5. Tsk and Tb were significantly decreased by 5.0 m x s(-1) wind in comparison to NoWi and Wi1. The higher exercise intensity was intense enough to diminish peripheral vasoconstriction and consequently the finger skin temperature was significantly higher at the end of HE in comparison to LE in NoWi and Wi1. Mean heat flux from the skin was unaffected by the exercise intensity. At LE oxygen consumption (VO2) was significantly higher in Wi5 than NoWi and Wi1. Heart rate was unaffected by the wind speed. The results suggest that, with studied exercise intensities, produced without changes in walking speed, the metabolic rate is not so important that it should be taken into consideration in the calculation of wind chill index. PMID- 11282320 TI - Exertion-induced fatigue and thermoregulation in the cold. AB - Cold exposure facilitates body heat loss which can reduce body temperature, unless mitigated by enhanced heat conservation or increased heat production. When behavioral strategies inadequately defend body temperature, vasomotor and thermogenic responses are elicited, both of which are modulated if not mediated by sympathetic nervous activation. Both exercise and shivering increase metabolic heat production which helps offset body heat losses in the cold. However, exercise also increases peripheral blood flow, in turn facilitating heat loss, an effect that can persist for some time after exercise ceases. Whether exercise alleviates or exacerbates heat debt during cold exposure depends on the heat transfer coefficient of the environment, mode of activity and exercise intensity. Prolonged exhaustive exercise leading to energy substrate depletion could compromise maintenance of thermal balance in the cold simply by precluding continuation of further exercise and the associated thermogenesis. Hypoglycemia impairs shivering, but this appears to be centrally mediated, rather than a limitation to peripheral energy metabolism. Research is equivocal regarding the importance of muscle glycogen depletion in explaining shivering impairments. Recent research suggests that when acute exercise leads to fatigue without depleting energy stores, vasoconstrictor responses to cold are impaired, thus body heat conservation becomes degraded. Fatigue that was induced by chronic overexertion sustained over many weeks, appeared to delay the onset of shivering until body temperature fell lower than when subjects were rested, as well as impair vasoconstrictor responses. When heavy physical activity is coupled with underfeeding for prolonged periods, the resulting negative energy balance leads to loss of body mass, and the corresponding reduction in tissue insulation, in turn, compromises thermal balance by facilitating conductive transfer of body heat from core to shell. The possibility that impairments in thermoregulatory responses to cold associated with exertional fatigue are mediated by blunted sympathetic nervous responsiveness to cold is suggested by some experimental observations and merits further study. PMID- 11282321 TI - An evaluation of the concept of living at moderate altitude and training at sea level. AB - Despite equivocal findings about the benefit of altitude training, current theory dictates that the best approach is to spend several weeks living at > or =2500 m but training near sea level. This paper summarizes six studies in which we used simulated altitude (normobaric hypoxia) to examine: (i) the assumption that moderate hypoxia compromises training intensity (two studies); and (ii) the nature of physiological adaptations to sleeping in moderate hypoxia (four studies). When submaximal exercise was >55% of sea level maximum oxygen uptake (VO2max), 1800 m simulated altitude significantly increased heart rate, blood lactate and perceived exertion of skiers. In addition, cyclists self-selected lower workloads during high-intensity exercise in hypoxia (2100 m) than in normoxia. Consequently, our findings partially confirm the rationale for 'living high, training low'. In the remaining four studies, serum erythropoietin increased 80% in the early stages of hypoxic exposure, but the reticulocyte response did not significantly exceed that of control subjects. There was no significant increase in haemoglobin mass (Hb(mass)) and VO2max tended to decrease. Performance in exercise tasks lasting approximately 4 min showed a non significant trend toward improvement (1.0+/-0.4% vs. 0.1+/-0.4% for a control group; P=0.13 for group x time interaction). We conclude that sleeping in moderate hypoxia (2650-3000 m) for up to 23 days may offer practical benefit to elite athletes, but that any effect is not likely due to increased Hb(mass) or VO2max. PMID- 11282322 TI - Structure and function of invertebrate 5-HT receptors: a review. AB - Over the last decade, knowledge of invertebrate serotonin receptors has expanded greatly. The first 5-HT receptor from Drosophila was cloned 10 years ago, and subsequently, 11 additional receptor genes have been cloned from Drosophila, molluscs (Lymnaea and Aplysia) and nematodes (Caenorhabditis and Ascaris). Information has also accumulated from physiological and biochemical studies that have used vertebrate serotonergic ligands to characterize endogenous invertebrate receptors. Although the endogenous receptors are often classified according to mammalian-based categories, in many cases the pharmacological properties of vertebrate and invertebrate receptors differ significantly and the actual identity of the latter is questionable. By providing information on the gene structure and amino acid sequence, molecular cloning studies offer a more definitive way to identify and classify invertebrate 5-HT receptors. This review summarizes information on the pharmacological and transductional properties of cloned invertebrate 5-HT receptors, and considers recent studies of endogenous receptors in the light of this new data. PMID- 11282323 TI - Sensitivity threshold and response characteristics of infrared detection in the beetle Melanophila acuminata (Coleoptera: Buprestidae). AB - The minimum detection threshold of the infrared sensitive beetle, Melanophila acuminata, was measured with a helium-neon laser that emitted light at a wavelength of 3.39 microm. Extracellular recordings were taken both at the pit organ responsible for detection and at the interganglionic connectives in the thorax of the beetle. At the pit organ, generator and action potentials from single neurons were measured with a sharpened tungsten electrode. At the connectives that linked the fused second meso-/metathoracic and prothoracic ganglia, compound action potentials were measured with a tungsten hook electrode that encircled the connective. The latter recordings confirmed conveyance of infrared information through specific pathways to rostrally-situated sites in the nervous system of the beetle. The 50% probability irradiance threshold at which action potentials were elicited from the receptor and connectives occurred at 17.3 and 14.6 mW/cm(2), respectively. In addition to sensitivity threshold, several other characteristics of the response were quantified including dependence of generator potential latency, generator potential duration, spike frequency, and spike latency on irradiance, dependence of response strength (spike count) on exposure time, and flicker fusion frequency. The ability to detect infrared radiation is rare in nature, and these results provide valuable information necessary to understand this unique sensitivity. PMID- 11282324 TI - Glycolipoprotein extract (G-90) from earthworm Eisenia foetida exerts some antioxidative activity. AB - Antioxidants protect DNA, proteins and lipids in the body from damage. These types of damages are a major contributor to aging and to degenerative diseases such as cancer, cardiovascular disease, immune-system decline, brain dysfunction and cataracts. The effect of glycolipoprotein extract of Eisenia foetida (G-90) as an antioxidant was investigated in cultured human fibroblasts and epithelial cells. After treatment of the cells with H2O2 for 4 h, G-90 completely allows the cells to recover and stimulated their growth. When the cells were incubated with G-90 48 h before the treatment with H2O2, the oxidative damage of the cells did not occur. Thus, G-90 had an apparent protective effect against the toxicity of H2O2 and stimulated the growth of the cells. Ascorbic acid, a known antioxidant, did not allow the growth of the cells to recover after damage nor did it protect them, unless it was added simultaneously with H2O2. The antioxidative activity of G-90, together with its antibacterial and mitogen activities, could be useful in the study of G-90 as a wound-healing agent. PMID- 11282325 TI - Uncoupling protein1 mRNA, mitochondrial GTP-binding, and T4 5'-deiodinase of brown adipose tissue in euthermic Daurian ground squirrel during cold exposure. AB - Regulation of thermogenic activity and uncoupling protein1 (UCP1) expression in brown adipose tissue (BAT) were studied in euthermic Daurian ground squirrel after acute and chronic cold exposure at 4 degrees C. The UCP1 concentration was indirectly determined by titration with its specific ligand [3H]-labeled GTP, and Ucp1 mRNA was detected by using a [32P]-labeled antisense oligonucleotide probe. Both acute and chronic cold exposure stimulated up-regulation of Ucp1 mRNA. Although UCP1 concentration is not significantly increased after 24 h of cold exposure, it is markedly elevated by 75% in squirrels after 4-week cold adaptation compared with controls raised at 22 degrees C. Changes in T4 5' deiodinase activity were closely associated with variations of Ucp1 mRNA level. Ucp1 gene expression is significantly affected by cold exposure in BAT from euthermic Daurian ground squirrels. In addition, the activation of T4 5' deiodinase may be an important regulatory factor in cold-induced Ucp1 expression. PMID- 11282326 TI - Involvement of cholinergic mechanisms in the central control of respiration in the cane toad, Bufo marinus. AB - Chemical substrates, central sites and central mechanisms underlying the regulation of breathing in lower vertebrates have not been well characterized. The present study was undertaken to determine the effect of pH changes and cholinergic agents on the central control of respiration in the cane toad, Bufo marinus. Adult toads were anesthetized, catheterized and unidirectionally ventilated before exposing the brainstem. An airtight buccal cannula was also inserted through the tympanum to record buccal pressure. The animal was decerebrated, anesthetic removed and the responses to pH changes of solutions bathing the ventral surface of the medulla (VSM) were tested by superfusing the VSM with mock cerebrospinal fluid (mCSF) of pH 7.8-normal, 7.2-acidic and 8.4 basic. The acidic solution increased respiratory activity, the basic solution decreased activity and the normal solution had no effect. In addition, cholinergeric agents (acetylcholine-ACh, physostigmine-Phy, nicotine-Nic, and atropine-Atr) dissolved in mCSF were applied bilaterally onto the VSM using filter paper pledgets. ACh, Phy and Nic increased episodic breathing frequency by 14.3+/-9.7, 9.4+/-5.4 and 29.1+/-11.8 %, respectively, whereas, Atr caused a decrease (-26.6+/-16.6%). These agents had no effect on blood pressure. It is therefore, concluded that the VSM is pH sensitive and a cholinergic mechanism is involved in the central modulation of respiration in Bufo. PMID- 11282327 TI - Effect of cold-acclimation on oxygen uptake and glucose production of perfused duckling liver. AB - The control of hepatic metabolism by substrates and hormones was assessed in perfused liver from young Muscovy ducklings. Studies were performed in fed or 24 h fasted 5-week-old thermoneutral (25 degrees C; TN) or cold-acclimated ducklings (4 degrees C; CA) and results were compared with those obtained in rats. Basal oxygen uptake of perfused liver (LVO2) was higher after cold acclimation both in fed (+65%) and 24-h fasted (+29%) ducklings and in 24-h fasted rats (+34%). Lactate (2 mM), the main gluconeogenic substrate in birds, similarly increased LVO2 in both TN and CA ducklings and the effect was larger after fasting. Both glucagon and norepinephrine dose-dependently increased LVO2 in ducklings and rats, but cold acclimation did not improve liver response and liver sensitivity to norepinephrine in ducklings was even reduced in CA animals. Liver contribution to glucagon-induced thermogenesis in vivo was estimated to be 22% in TN and 12% in CA ducklings. Glucagon stimulated gluconeogenesis from lactate in duckling liver and the stimulation was 2.2-fold higher in CA than in TN fasted birds. These results indicate a stimulated hepatic oxidative metabolism in CA ducklings but hepatic glucagon-induced thermogenesis (as measured by LVO2) was not improved. A role of the liver is suggested in duckling metabolic acclimation to cold through an enhanced hepatic gluconeogenesis under glucagon control. PMID- 11282329 TI - Subunit compositions of crustacean haemocyanins are species-specific: evidence from non-decapod species. AB - Electrophoretic examination of dissociated haemocyanin subunits from a number of amphipod, decapod and isopod crustaceans supports the hypothesis that subunit composition is species-specific, despite marked within-species variation in many species. General patterns of heterogeneity on native PAGE gels were also evident between groupings within the Amphipoda. Gammarid amphipods could be split into two groups; one characterised by a high degree of heterogeneity and the other by a low degree of heterogeneity. The talitrid amphipods generally displayed a low degree of heterogeneity similar to, although still distinct from, the second gammarid category. Haemocyanin from the Hyalidae, a family allied to the talitrids was highly heterogeneous, similar to the first gammarid group and unlike the talitrids. Isopod haemocyanin banding patterns were more similar to one another than to any of the amphipod or decapod species examined. In general, the molecular weights of the amphipod Hcs tended to be greater than those of the isopods, with the decapods being lowest of all. It is suggested that Hc subunit heterogeneity may be a useful tool for investigating speciation and speciation events, and for reliably separating very closely-related species (e.g. Gammarus spp.), purely on the basis of their Hc subunit compositions. PMID- 11282328 TI - On the mechanism of sodium-proton exchange in crayfish. AB - In salt depleted crayfish net sodium and proton fluxes were coupled 1:1 as required by the frog skin-turtle bladder model. In addition, three proton pump inhibitors produced equal reductions of both fluxes. It is concluded that the model operates in these animals. Net Na+ and H+ fluxes were very small in tap water adapted animals, but regression analysis clearly showed that they were coupled, though perhaps not 1:1. Proton pump inhibitors, at concentrations that suppressed fluxes in salt-depleted crayfish, had no measureable effect on proton movement in tap water-adapted animals. Two of them (dicyclocarbodiimide and N ethyl maleimide), caused a small reduction in Na+ influx without affecting proton efflux. These experiments provide no support for operation of the frog-turtle system in adult crayfish adapted to tap water. A 2Na+-H+ exchanger is considered from an energetic point of view. Such a system might be able to couple Na+ and H+ fluxes in dilute, near neutral solutions ([Na+] approximately 1-2 mM; pH 7). PMID- 11282333 TI - Biocatalysis and biotransformation bio-inorganic chemistry. Web alert. AB - A selection of World Wide Web sites relevant to papers published in this issue of Current Opinion in Chemical Biology. PMID- 11282330 TI - The effects of endurance training and exhaustive exercise on mitochondrial enzymes in tissues of the rat (Rattus norvegicus). AB - The aim of the present study was to ascertain the effects of training and exhaustive exercise on mitochondrial capacities to oxidize pyruvate, 2 oxoglutarate, palmitoylcarnitine, succinate and ferrocytochrome c in various tissues of the rat. Endurance capacity was significantly increased (P<0.01) by an endurance training program over a period of 5-6 weeks. The average run time to exhaustion was 214.2+/-23.8 min for trained rats in comparison with 54.5+/-11.7 min for their untrained counterparts. Oxidative capacities were reduced in liver (P<0.05) and brown adipose tissue (P<0.05) as a result of endurance training. On the contrary, the oxidative capacity of skeletal muscle was slightly increased and that of heart almost unaffected except for the oxidation of palmitoylcarnitine, which was significantly reduced (P<0.05) as a result of training. PMID- 11282335 TI - Biocatalytic modification of natural products. AB - Natural products are ideal training compounds for enzymatic catalysis. New transformations have become possible on a preparative scale thanks to molecular biology, which has made many new enzymes available. Additionally, new synthetic pathways have been developed to regenerate expensive cofactors in situ and to improve enzyme selectivity. PMID- 11282336 TI - Synthetic applications of epoxide hydrolases. AB - There have been several recent advances in the area of biocatalysed hydrolytic kinetic resolution of epoxides using 'newly discovered' enzymes (i.e. epoxide hydrolases). These biocatalysts, two of which will become commercially available in the near future, appear to be highly promising tools for fine organic synthesis, as they enable the preparation of various epoxides and/or their corresponding diols in enantiopure form. PMID- 11282337 TI - Dehydrogenases and transaminases in asymmetric synthesis. AB - Improved stereoselectivity in dehydrogenase-mediated reductions has been achieved by rationally designed gene overexpression and knockouts in Saccharomyces cerevisiae cells and by isolating and characterizing novel dehydrogenases from other organisms. Transaminases have been used to prepare unnatural amines and amino acids in good yields, particularly when the equilibria are shifted by selective product removal. PMID- 11282338 TI - Industrial biocatalysis. AB - The number of industrial processes for the synthesis of fine and commodity chemicals, pharmaceutical and agrochemical intermediates and drug substances utilizing biological catalysts continues to grow. The combination of new molecular biology techniques, such as directed evolution and pathway engineering, with new and efficient high-throughput screening methods is poised to bolster this field and further advance the contribution of biocatalysis to the chemical and the pharmaceutical industries. PMID- 11282339 TI - Improved biocatalysts by directed evolution and rational protein design. AB - The efficient application of biocatalysts requires the availability of suitable enzymes with high activity and stability under process conditions, desired substrate selectivity and high enantioselectivity. However, wild-type enzymes often need to be optimized to fulfill these requirements. Two rather contradictory tools can be used on a molecular level to create tailor-made biocatalysts: directed evolution and rational protein design. PMID- 11282340 TI - Enzymes from extremophiles. AB - The industrial application of enzymes that can withstand harsh conditions has greatly increased over the past decade. This is mainly a result of the discovery of novel enzymes from extremophilic microorganisms. Recent advances in the study of extremozymes point to the acceleration of this trend. In particular, enzymes from thermophilic organisms have found the most practical commercial use to date because of their overall inherent stability. This has also led to a greater understanding of stability factors involved in adaptation of these enzymes to their unusual environments. PMID- 11282341 TI - Novel methods for biocatalyst screening. AB - There have been a number of recent advances in catalysis assays applicable for screening biocatalyst libraries in high-throughput format. These include instrumental assays such as high-performance liquid chromatography, mass spectrometry, capillary electrophoresis and IR-thermography, reagent-based assays producing spectroscopic signals (UV/VIS or fluorescence) in response to reaction progress, and assays based on fluorogenic or chromogenic substrates. These fluorogenic substrates enable the assaying of a variety of enzymes in enantioselective and stereoselective manner, including alcohol dehydrogenases, aldolases, lipases, amidases, epoxide hydrolases and phosphatases. PMID- 11282342 TI - Combinatorial biosynthesis of polyketides and nonribosomal peptides. AB - The engineering of polyketide biosynthesis has begun to provide robust targeted libraries for screening against pharmaceutically relevant targets. New technologies that offer methodology for the rapid generation of more structurally diverse libraries have now been demonstrated. PMID- 11282344 TI - Structural aspects of denitrifying enzymes. AB - The reduction of nitrate to nitrogen gas via nitrite, nitric oxide and nitrous oxide is the metabolic pathway usually known as denitrification, a key step in the nitrogen cycle. As observed for other elemental cycles, a battery of enzymes are utilized, namely the reductases for nitrate, nitrite, nitric oxide and nitrous oxide, as well as multiple electron donors that interact with these enzymes, in order to carry out the stepwise reactions that involve key intermediates. Because of the importance of this pathway (of parallel importance to the nitrogen-fixation pathway), efforts are underway to understand the structures of the participating enzymes and to uncover mechanistic aspects. Three dimensional structures have been solved for the majority of these enzymes in the past few years, revealing the architecture of the active metal sites as well as global structural aspects, and possible mechanistic aspects. In addition, the recognition of specific electron-transfer partners raises important questions regarding specific electron-transfer pathways, partner recognition and control of metabolism. PMID- 11282345 TI - Recent advances in bioinorganic spectroscopy. AB - Spectroscopic methods covering many energy regions together provide complementary insight into metalloenzyme active sites. These methods probe geometric and electronic structure and define these contributions to reactivity. Two recent advances--determination of the polarizations of electronic transitions in solution using magnetic circular dichroism, electron paramagnetic resonance and quantum chemistry, and experimental estimation of covalency using metal L-edges and ligand K-edges--are particularly important. PMID- 11282346 TI - New advances in ligand design for synthetic modeling of metalloprotein active sites. AB - Judicious control of ligand steric bulk and auxiliary structural elements enables the construction of novel synthetic complexes that model the properties of metalloprotein active sites, thus providing insight into their structure and function. Major recent developments include the synthesis of a number of unusual and biologically relevant complexes of copper and iron using elaborate N-donor and O-donor ligands. PMID- 11282347 TI - Designing metal-peptide models for protein structure and function. AB - Recent progress in the rational design of metal sites within peptide model systems shows increasing control in the placement of metals within helical bundles and inclusion of sophisticated elements such as second-sphere ligand interactions. A crystallographically characterized two-metal peptide model for diiron proteins represents a major achievement in de novo design methodologies. Increasingly complex and robust models for electron transfer through and between helices, and electrode-supported electron-transfer peptides, have been constructed. Design elements for peptide-supported ferredoxins and mononuclear Fe(II) and Zn(II) sites have been refined. PMID- 11282348 TI - Lanthanide-mediated DNA hydrolysis. AB - Lanthanide ions are remarkably effective catalysts for the hydrolytic cleavage of phosphate ester bonds, including the robust bonds of DNA. This makes Ln(III) and Ce(IV) ions attractive candidates for developing selective and efficient artificial nucleases, which could have many biochemical and clinical applications. Both small-molecule-based and biopolymer-based lanthanide complexes are being pursued. PMID- 11282349 TI - Electrochemistry at DNA-modified surfaces: new probes for charge transport through the double helix. AB - Electrochemistry at DNA-modified surfaces provides an alternative approach to photochemistry or radiation biology for studying charge migration through the double helix. Using short duplexes self-assembled onto gold, electrochemical reduction of redox-active reporter molecules has been observed through DNA films more than 50 A thick, with heterogeneous rate constants as great as approximately 100 s(-1). Though apparently insensitive to base content and sequence, even small distortions in the electronic structure of the pi-stack (caused, for example, by single-base mismatches and other DNA lesions) attenuate the rate of electron transport. Understanding the role of conformational dynamics within the double helix, as well as the cooperative effects of self-assembling individual duplexes into ordered superlattices remain important challenges for theory and experiment. PMID- 11282350 TI - Recent advances in heme-protein sensors. AB - In recent years, an increasing number of proteins have been discovered which utilize heme cofactors to sense oxygen, carbon monoxide and nitric oxide. The identification and characterization of these proteins are revising our understanding of heme-mediated allostery first established in the early 1960s. Biochemical and structural studies are revealing new mechanisms for heme-driven conformational changes distinct from the classical hemoglobin model. PMID- 11282351 TI - The biological chemistry of lead. AB - Recent biophysical studies on the interactions between lead and recombinant proteins and peptides that naturally bind zinc or calcium have provided unparalleled insights into the biological chemistry and molecular toxicology of lead. These studies lay the foundation for the rational design of improved methods for detecting and treating lead poisoning. PMID- 11282352 TI - Barium-stimulated chemosensory activity may not reflect inhibition of background voltage-insensitive K+ channels in the rat carotid body. AB - To test the hypothesis that the voltage-insensitive background leak K+ channel is responsible for the oxygen-sensitive properties of glomus cells in the rat carotid body (CB) we used Ba2+, a non-specific inhibitor of K+ currents. In vitro changes in cytosolic calcium ([Ca2+]c) and chemosensory discharge were studied to measure the effect of Ba2+. In normal Tyrode buffer, Ba2+ (3 and 5 mM) significantly increased carotid sinus nerve (CSN) discharge over baseline firing rates under normoxia (PO2 approximately 120 Torr) from approximately 150 to approximately 600 imp/0.5 s. However, addition of 200 microM Cd2+ which completely blocked increase in CSN activity stimulated by hypoxia (PO2 approximately 30 Torr), hypercapnia (PCO2 approximately 60 Torr, PO2 approximately 120 Torr) and high CO (PCO approximately 550 Torr, PO2 approximately 120 Torr) did not significantly inhibit Ba2+-stimulated CSN discharge. The response to hypoxia is abolished with Ca2+-free tyrode buffer containing 10 mM EGTA. Yet, in the same buffer, Ba2+ increased CSN discharge from approximately 2 to approximately 180 imp/0.5 s. With 200 microM Cd2+ and 10 mM EGTA, Ba2+ still increased CSN discharge from approximately 2 to approximately 150 imp/0.5 s. Oligomycin (2 microg) abolished the hypoxic response. However, in the presence of oligomycin CSN response to Ba2+ was significant. Since Ba2+ increased neural discharge under conditions where hypoxia stimulated CSN discharge is completely abolished, we suggest that the effect of Ba2+ on CSN discharge may not have anything to do with the oxygen sensing mechanism in the CB. PMID- 11282353 TI - The dual effect of a nitric oxide donor in nociception. AB - Low intrathecal (i.t.) doses of the nitric oxide (NO)-donor 3 morpholinosydnonimine (SIN-1) (0.1-2.0 microg/10 microl) reduced, while higher doses had no effect (5 or 100 microg/10 microl) or increased (10 and 20 microg/10 microl) the mechanical allodynia induced by chronic ligature of the sciatic nerve in rats. SIN-1 (0.1-100 microg/10 microl; i.t.) produced only antinociceptive effect in the rat tail flick test. The inhibitor of guanylate cyclase, 1H [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) (4 microg/10 microl; i.t.), abolished the antinociceptive effects of SIN-1 in both tests and reduced the effect of high doses of SIN-1 in neuropathic rats. Hemoglobin (100 microg/10 microl; i.t.), a NO scavenger, inhibited the effect of low dose of SIN-1 and reduced the effect of high dose of SIN-1 in neuropathic rats. 8-Bromo-cGMP (125 500 microg/10 microl; i.t.), reduced the mechanical allodynia in neuropathic rats. The NO-synthase inhibitors, NG-nitro-L-arginine (L-NOARG) and NG-monomethyl L-arginine (L-NMMA) (75-300 microg/10 microl; i.t.) reduced the mechanical allodynia evoked by nerve injury and increased the tail-flick latency, respectively. These effects were reduced and inhibited, respectively, by previous i.t. ODQ. The effect of L-NOARG was enhanced in a non-significant manner by hemoglobin. These results indicate that SIN-1 and NO-synthase inhibitors reduce pain through a spinal mechanism that involves activation of guanylate cyclase. The effects of SIN-1 vary depending on the dose and pain model utilized, but its most sensitive effect seems to be antinociception. However, high doses of the NO donor can intensify ongoing pain. PMID- 11282354 TI - Sequential upregulation of cell adhesion molecules in degenerating rat basal forebrain cholinergic neurons and in phagocytotic microglial cells. AB - To study the functional role of adhesion molecules in neurodegenerative events in vivo, the basal forebrain cholinergic lesion-induced expression of the intercellular adhesion molecule (ICAM)-1 and leukocyte function-associated antigen (LFA)-1 was studied by double immunocytochemistry and Western blot analysis. A single intracerebroventricular application of the cholinergic immunotoxin, 192IgG-saporin, produced a selective cholinergic cell loss in rat basal forebrain nuclei detectable by gradual loss of choline acetyltransferase (ChAT)-immunoreactive cells starting 3 days but being nearly complete 7 days after injection of the toxin. The degeneration of cholinergic neurons was accompanied by a striking appearance of activated microglial cells in the lesioned areas. Four days following injection of 192IgG-saporin, ICAM-1 immunoreactivity was predominantly observed in ChAT-positive neurons and partly in activated microglia in the basal forebrain nuclei, while LFA-1 expression at this time point was restricted to neurons. However, 7 days after cholinergic lesion, only a few, shrunken neuronal somata were found to be immunoreactive for ICAM-1 and LFA-1, while activated microglial cells demonstrated strong immunoreactivity for ICAM-1 and LFA-1 in the lesioned forebrain areas, persisting up to 14 days after lesion while no immunoreactivity was observed in neurons at this time point. Western blot analysis demonstrated increased ICAM-1 level in the basal forebrain already detectable 4 days after surgery but being more pronounced 7 days post lesion. The data suggest that ICAM-1 and LFA-1 may act as intercellular recognition signals by which degenerating cholinergic neurons actively participate in the sequence of events leading to their targeting and elimination by phagocytotic microglia. PMID- 11282355 TI - Gamma-hydroxybutyric acid-induced absence seizures in GluR2 null mutant mice. AB - In this electrophysiological study, we examined the susceptibility of GluR2 mutant null mice to absence seizures in comparison with wild-type controls. The prodrug of (GHB), gamma-butyrolactone (GBL) was given systemically to induce the absence seizures. We also tested the severity and duration of the seizure activity in this model. The results showed that the latency from GBL administration to onset of seizure was significantly prolonged in GluR2(-/-) mice when compared to GluR2(+/+) mice. The duration of spike-and-wave discharges (SWD) was also significantly decreased in the GluR2(-/-) mice. Ninety minutes following GBL administration, wild-type animals continued to exhibit intermittent SWD bursts while GluR2(-/-) mice had returned to baseline. These data suggest that the GluR2 subunit may be involved in the initiation and maintenance of absence seizures induced by GBL. PMID- 11282356 TI - Serotonin, excitatory amino acids and the photic control of melatonin rhythms and SCN c-FOS in the rat. AB - There is a growing acceptance that serotonergic pathways to the suprachiasmatic nucleus play an important role in the mediation and modulation of light entrainment of rhythms. In this study administration of the 5-HT(2A/2C) agonist (+/-)-1-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane (DOI, 0.5 mg/kg) at mid dark caused a phase shift in the onset of the urinary excretion of 6 sulphatoxymelatonin in rats that was sustained for at least 8 days and was blocked by the specific 5-HT(2C) antagonist SB-242084. Administration of DOI (2 mg/kg) across the night resulted in the appearance of c-FOS in the nucleus of cells in the suprachiasmatic nucleus during subjective darkness, but did not cause induction at the time of expected lights on (CT0). By contrast light exposure induced c-fos throughout the night including CT0. Administration of the NMDA receptor antagonist MK-801 (3 mg/kg) prior to light pulses had no effect on c-fos in the first part of the night, but towards the expected time of lights on, became progressively more potent, such that by CT0, light induction of c-fos was almost completely inhibited. These results provide further evidence that serotonin plays a role in the mediation of light effects on rhythms in the rat. PMID- 11282357 TI - The effects of intrahippocampal testosterone and flutamide on spatial localization in the Morris water maze. AB - The high density of the androgen receptors in fundamental centers of learning and memory, such as hippocampus, shows that there must be some relationships between the androgen receptors and cognitive aspects. To determine the role of hippocampal androgen receptors in spatial learning, the current research has been conducted to assess the effect of testosterone enanthate, as the agonist, and flutamide, as the antagonist, of these receptors on spatial discrimination of rats, using the Morris water maze (MWM). Adult male rats were bilaterally cannulated into the CA1 region of their hippocampus. Different groups received different doses of flutamide (2, 5, 10 and 20 microg/0.5 microl) or testosterone enanthate (20, 40 and 80 microg/0.5 microl) through the cannulas 30 min before training for 3 days. The results showed dose-dependent increases in latencies and traveled distances to find the invisible platform both in flutamide- and testosterone-treated groups as compared to the control group, with peak effects at doses of 5 microg/0.5 microl for flutamide and 80 microg/0.5 microl for testosterone. Therefore, it seems that both androgen receptor blockade and exogenous testosterone can effect spatial localization of adult, male rats. PMID- 11282358 TI - Blockade of U50488H on potassium currents of acutely isolated mouse hippocampal CA3 pyramidal neurons. AB - The actions of the opioid agonist U50488H on IA and IK were examined in acutely isolated mouse hippocampal CA3 pyramidal neurons using the whole-cell patch clamp technique. U50488H caused a concentration dependent, rapidly developing and reversible inhibition of voltage-activated IA and IK. The inhibitory actions were still observed in the presence of 30 microM naloxone or 5 microM nor binaltorphimine dihydrochloride. The IC50 values for the blockade of IA and IK were calculated as 20.1.9 and 3.7 microM, respectively. In the presence of 3.3 microM U50488H, repetitive stimulation induced use-dependent inhibition of IA and IK. A 10 microM concentration of U50488H positively shifted the half-activation membrane potential of IA by +11 mV, but negatively shifted IK by -14 mV. These results demonstrate that U50488H can directly inhibit neuronal IA and IK without involvement of the activation of kappa-opioid receptors. PMID- 11282359 TI - Possible role of enhanced microtubule phosphorylation in dichlorvos induced delayed neurotoxicity in rat. AB - The effect of a single subcutaneous dose of 200 mg/kg body weight dichlorvos on neuronal microtubule phosphorylation has been studied in rat following the development of organophosphate induced delayed neurotoxicity (OPIDN). Microtubule associated Ca2+/calmodulin dependent as well as cAMP dependent protein kinases were assayed. Dichlorvos administration led to a consistent increase in the activity of both the kinases at all post exposure intervals (7th, 15th and 21st day) as compared to that of controls. After in vitro phosphorylation using [gamma 32P]ATP, various proteins were resolved on one-dimensional 8% SDS-PAGE, stained with Coomassie Blue and autoradiographed. The amount of 32P incorporated was quantified by microdensitometry. Dichlorvos enhanced the phosphorylation of 55- and 280-kDa proteins. These two proteins were identified as tubulin and microtubule associated protein-2 (MAP-2) by immunoblotting. This study showed that dichlorvos induced hyperphosphorylation of tubulin and MAP-2 which in turn destabilizes microtubule assembly, and may ultimately result in axonal degeneration leading to dichlorvos induced delayed neurotoxicity. PMID- 11282360 TI - A fMRI study of brain activations during non-noxious and noxious electrical stimulation of the sciatic nerve of rats. AB - An acute pain animal model for fMRI study would provide useful spatial and temporal information for studying the supraspinal nociceptive neuronal responses. The aim of the present study was to investigate whether the nociceptive responses in different brain areas can be differentiated by using functional magnetic resonance imaging (fMRI) in anesthetized rats. Functional changes in brain regions activated by noxious or non-noxious stimuli of the sciatic nerve were investigated using fMRI in a 4.7 T MR system in alpha-chloralose anaesthetized rats. To determine the electrical intensity for noxious and non-noxious stimuli, compound action potential recording was employed to reveal the type of fibers activated by graded electrical stimulation of sciatic nerve. It showed that innocuous A-beta fibers were excited by two times the muscle twitch threshold and nociceptive A-delta and C fibers were recruited and excited by 10 and 20 times threshold, respectively. A series of four-slice gradient echo images were acquired during innocuous (two times threshold) and noxious (10 and 20 times threshold) stimuli in a 4.7 T MR system. Contralateral somatosensory cortex was the most prominent brain area activated by innocuous stimuli. Both signal intensity and activated areas were significantly increased in the somatosensory cortex, cingulate cortex, medial thalamus and hypothalamus during noxious stimuli. These four brain areas activated by noxious stimuli were significantly suppressed by prior intravenous injection of morphine (5 mg/kg). The present findings demonstrated that the difference of the innocuous and nociceptive responses in the brain could be detected and localized by an in vivo spatial map using fMRI. Results suggest that fMRI may be an invaluable tool for studying pain in anesthetized animals. PMID- 11282361 TI - Site-specific activation of dopamine and serotonin transmission by aniracetam in the mesocorticolimbic pathway of rats. AB - The effects of aniracetam on extracellular levels of dopamine (DA), serotonin (5 HT) and their metabolites were examined in five brain regions in freely moving stroke-prone spontaneously hypertensive rats (SHRSP) using in vivo microdialysis. Basal DA release in SHRSP was uniformly lower in all regions tested than that in age-matched control Wistar Kyoto rats. 3,4-Dihydroxyphenylacetic acid and homovanillic acid levels were altered in the basolateral amygdala, dorsal hippocampus and prefrontal cortex of SHRSP. While basal 5-HT release decreased in the striatum and increased in the basolateral amygdala, there was no associated change in 5-hydroxyindoleacetic acid levels. Systemic administration of aniracetam to SHRSP enhanced both DA and 5-HT release with partly associated change in their metabolite levels in the prefrontal cortex, basolateral amygdala and dorsal hippocampus, but not in the striatum and nucleus accumbens shell, in a dose-dependent manner (30 and/or 100 mg/kg p.o.). Microinjection (1 and 10 ng) of aniracetam or its metabolites (N-anisoyl-GABA and 2-pyrrolidinone) into the nucleus accumbens shell produced no turning behavior. These findings indicate that SHRSP have a dopaminergic hypofunction throughout the brain and that aniracetam elicits a site-specific activation in mesocorticolimbic dopaminergic and serotonergic pathways in SHRSP, possibly via nicotinic acetylcholine receptors in the ventral tegmental area and raphe nuclei. The physiological roles in the aniracetam-sensitive brain regions may closely link with their clinical efficacy towards emotional disturbances appearing after cerebral infarction. PMID- 11282362 TI - Hippocampal transection attenuates kainic acid-induced amygdalar seizures in rats. AB - Since the dorsal and ventral hippocampus in the rat may act differently from one another in limbic seizures, we studied effects of orthogonal transection between the dorsal and ventral hippocampus upon kainic acid-induced amygdalar seizures. A total of 26 rats were divided into three groups. Ten rats underwent transection using a modified wire knife (transection group); 16 others were untransection group (n=10) and controls (n=6). All the rats then underwent stereotactic implantation of electrodes in the left amygdala (LA), left dorsal hippocampus (LdH), left ventral hippocampus (LvH), and the left sensorimotor cortex (LCx). A stainless steel cannula also was introduced into the LA. Rats except controls later received 1.0 microg of kainic acid (KA) via the cannula. Controls received phosphate buffer solution alone. In the untransection group, multiple spike discharges in the LA immediately propagated concurrently to the LvH and LdH. Propagation involved the LCx to become status epilepticus 1 to 2 h after KA injection. Seizures, characterized by mastication, salivation, facial twitching, forelimb clonus, and sometimes rearing and falling, lasted 1 to 2 days. Microscopic examination revealed severe neuronal cell damage in the LA, LvH, and LdH. In the transection group, multiple spike discharges initiated from the LA and were propagated to LvH, but LdH as well as LCx involvement was slight. Status epilepticus involved only the LA and LvH 1 to 2 h following KA injection. Seizures subsided within 24 h, showing no ictal manifestations except for aggressiveness. Overall, seizures were weak and transient compared with those in controls. Histologically, hippocampal neuronal damage was slight, but damage to amygdalar neurons was similar to that in untransection group. No electroclinical and histological changes were seen in controls. These results indicated that connections between the dorsal and ventral hippocampus are important for full development of KA-induced amygdalar seizures. PMID- 11282363 TI - Increased extracellular ascorbate release reflects glutamate re-uptake during the early stage of reperfusion after forebrain ischemia in rats. AB - Ascorbate is highly concentrated in neuropils, and its extracellular release is closely related to that of the excitatory neurotransmitters. Thus, the extracellular release of ascorbate and glutamate was measured during the early stage of forebrain ischemia-reperfusion in the rat hippocampus using a microdialysis biosensor system. Male Wistar rats were anesthetized with halothane under mechanical ventilation and normothermia. Two probes of the microdialysis biosensor electrode were inserted in the hippocampus bilaterally. One probe was perfused with phosphate-buffered saline (PBS) and the oxidation signal of dialyzed ascorbate was recorded. A second electropolymerized probe was perfused with PBS containing glutamate oxidase for glutamate measurement. Forebrain ischemia-reperfusion was performed by bilateral carotid artery occlusion with hemorrhagic hypotension (MAP=30 mmHg) for 10 min (Group 10, n=10) or 15 min (Group 15, n=10), followed by reperfusion for 60 min. The release of glutamate increased significantly to 294% (Group 10) and 334% (Group 15) during ischemia, and then decreased rapidly. In Group 15, however, it remained significantly higher after reperfusion than in Group 10. The release of ascorbate increased significantly to 504% (Group 10) and 334% (Group 15) after reperfusion. In Group 10, it was significantly higher for 5-15 min after reperfusion than in Group 15. The marked increase of ascorbate during reperfusion was associated with the rapid decrease in glutamate. The extended time of ischemia significantly inhibited glutamate re-uptake and ascorbate release during reperfusion. These findings suggest the extracellular ascorbate release during reperfusion after global ischemia as a marker of glutamate re-uptake. PMID- 11282364 TI - The effects of angiotensin IV analogs on long-term potentiation within the CA1 region of the hippocampus in vitro. AB - Within the brain-renin angiotensin system, it is generally assumed that angiotensin peptide fragments shorter than angiotensins II and III, including angiotensin IV (AngIV), are inactive. This belief has been challenged by the recent discovery that AngIV, and AngIV-like analogs, bind with high affinity and specificity to a putative angiotensin binding site termed AT4. In the brain these sites include the hippocampus, cerebellum, and cerebral cortex, and influence associative and spatial learning tasks. The present study investigated the effects of two AngIV analogs, Nle1-AngIV (an AT4 receptor agonist) and Nle1 Leual3-AngIV (an AT4 receptor antagonist), on long-term potentiation (LTP). Field excitatory postsynaptic potentials (fEPSPs) were recorded from the CA1 stratum radiatum following stimulation of the Schaffer collateral pathway. Activation of AT4 receptors by Nle1-AngIV enhanced synaptic transmission during low-frequency test pulses (0.1 Hz), and increased the level of tetanus-induced LTP by 63% over that measured under control conditions. Paired stimulation before and during infusion of Nle1-AngIV indicated no change in paired-pulse facilitation (PPF) as a result of AT4 receptor activation suggesting that the underlying mechanism(s) responsible for Nle1-AngIV-induced increase in synaptic transmission and LTP is likely a postsynaptic event. Further, applications of Nle1-Leual3-AngIV prior to, but not 15 or 30 min after, tetanization prevented stabilization of LTP. These results extend previous findings from behavioral data in that AT4 receptor agonists and antagonists are capable of activating, and inhibiting, learning and memory pathways in the hippocampus, and suggest that the AT4 receptor subtype is involved in synaptic plasticity. PMID- 11282366 TI - Electrolytic lesion of the anterior cingulate cortex decreases inflammatory, but not neuropathic nociceptive behavior in rats. AB - The present study investigated the effect of lesions of the anterior cingulate cortex (ACC) on mechanical allodynia/hyperalgesia after L5 ligation or on inflammatory nociceptive responses following formalin injection in the rat. For both the neuropathic and inflammatory pain models, three groups of animals were used. The control groups consisted of a group of sham lesioned animals and a group of animals that had unilateral damage to the ACC or unilateral/bilateral damage to surrounding cortical tissue. The third group consisted of animals that had at least 75% bilateral damage of the ACC. Subjects received L5 ligation or a 0.05-ml injection of 1% formalin into the plantar surface of the hindpaw. In contrast to the control groups, bilateral ACC lesions significantly decreased inflammatory nociceptive responses during the prolonged, tonic portion of the formalin test (20-35 min). The difference between the groups was most prevalent in the amount of time spent licking the paw. However, ACC lesions did not significantly attenuate the enhanced mechanical paw withdrawal threshold in the neuropathic nociceptive model. These results suggest a differential role of the ACC in the modulation of different types of pain conditions. PMID- 11282365 TI - Sleep and waking following microdialysis perfusion of the selective 5-HT1A receptor antagonist p-MPPI into the dorsal raphe nucleus in the freely moving rat. AB - The aim of this study was to examine the involvement of the dorsal raphe nucleus (DRN) presynaptic serotonergic 5-HT1A autoreceptors on sleep and waking parameters, in particular rapid eye movement (REM) sleep. In a previous study, the systemic administration of the selective 5-HT1A receptor antagonist p-MPPI reduced REM sleep in a dose-dependent manner suggesting a blockade of the 5-HT1A autoreceptors. In the present study, a blockade by microdialysis perfusion of 10 microM and 100 microM of p-MPPI for 7 h into the DRN in freely behaving rats influenced vigilance state only to a small extent. The administration of 10 microM of p-MPPI induced a reduction of total REM sleep mainly due to a suppression of REM sleep during the third 2 h period of the recording of sleep and waking. Perfusion of 100 microM of p-MPPI decreased total transition type sleep (TRANS) but the effect on REM sleep did not reach significance. There was no change in waking or slow wave sleep (SWS) following any of the doses. The data suggest that 5-HT1A receptor-mediated mechanisms in the DRN may be only moderately important in the serotonergic modulation of REM sleep. PMID- 11282367 TI - Ethanol-regulated gene expression of neuroendocrine specific protein in mice: brain region and genotype specificity. AB - Neuroendocrine specific protein or reticulon 1 (NSP/RTN1) was identified as a putative ethanol-regulated gene using mRNA differential display in mice genetically selected for severe ethanol withdrawal (withdrawal seizure-prone, WSP). One transcript of RTN1 (3.0 kb) showed a statistically significant increase (13%) in relative abundance in whole brain of ethanol-treated WSP mice but not in mice selected for resistance to ethanol withdrawal convulsions (WSR). We hypothesized that ethanol-induced regulation of gene expression of mRTN1 is specific to mice predisposed to exhibit severe ethanol withdrawal and that the gene might be regulated differentially in specific brain regions. WSP and WSR selected lines and DBA/2J and C57BL/6J inbred strains of mice were exposed to ethanol vapor or air for 72 h. mRNA steady-state expression of RTN1 was assessed in hippocampus, cortex, and cerebellum. Results indicated that the pattern of ethanol-induced changes in gene expression was dependent upon transcript size, brain region, and genotype. Modest increases in the relative abundance of both transcripts of RTN1 were observed in the hippocampus and cortex of all ethanol treated mice. Results from cerebellum showed a moderate decrease in expression of RTN1 (3.0 kb transcript) in WSP and DBA/2J mice, but not in the mice resistant to ethanol withdrawal (C57BL/6J and WSR). These results suggest a genotype-specific effect of chronic ethanol exposure on steady-state mRNA levels of RTN1 in the cerebellum. Overall, the results indicate a complex pattern of ethanol-induced regulation of the putative mouse homologue of RTN1 and suggest that specific brain regional changes may be involved in the expression of physical dependence. PMID- 11282368 TI - Two-stage theory of conditioning: involvement of the cerebellum and the amygdala. AB - Classical conditioning is thought to proceed through two successive stages: fast rate emotional conditioning followed by slower motor conditioning. To verify the involvement of the amygdala and the cerebellum in these two stages of learning, rats were subjected to paired tone-airpuff (CS-US) trials. Lick suppression to CS was used as an index of conditioned emotional response (emotional CRs) and head movement was used as an index of motor CRs. The results showed that the fast acquisition of emotional CRs was dependent on the integrity of the amygdala and the slow acquisition of motor CRs was dependent on the integrity of the cerebellar interpositus nucleus. Cerebellar lesions had no effect on the acquisition of the emotional CRs but prevented the extinction of the emotional CRs seen in intact rats after massive conditioning. These findings suggest that the amygdala and the cerebellum provide the neuronal substrates of the fast and slow conditioning systems, respectively, and that conditioning-related cerebellar output interacts with the amygdala-based emotional conditioning. PMID- 11282370 TI - Contribution of the human superior parietal lobule to spatial selection process: an MEG study. AB - The magnetoencephalographic signal was collected in a visually guided response finger selection GO/NOGO task. The minimum norm distributed source analysis identified the sources in bilateral superior parietal lobules (SPL), with stronger activity for contralateral finger movement. Our results suggest that the human SPL plays a role in the spatial selection in a visuomotor task similar to that identified recently in monkeys. PMID- 11282369 TI - RTI-76, an irreversible inhibitor of dopamine transporter binding, increases locomotor activity in the rat at high doses. AB - An earlier study in our laboratory showed that 24 h after intracerebroventricular administration of the irreversible dopamine transporter inhibitor RTI-76, [3H]GBR12935 binding to the dopamine transporter (DAT) protein was inhibited in both the striatum and nucleus accumbens of the rat in a dose-dependent fashion (0.05-5.0 micromol). The rate of return of binding to control levels was used to calculate the half-life of DAT. Since changes in behavior could conceivably influence the half-life, the effects of various doses of RTI-76 on locomotor activity 1 and 3 days after RTI-76 administration were examined. During the first day after i.c.v. administration, 1.25 micromol RTI-76 had no effect on locomotor activity, but 2.5 micromol RTI-76 significantly increased locomotor activity in rats, a time at which this dose inhibited 41 and 42% of [3H]GBR12935 binding in the striatum and in the nucleus accumbens, respectively. These results agree with earlier reports showing that significant blockade of the dopamine transporter protein in the striatum is required for increases in motor activity in rodents. However, 5.0 micromol RTI-76 did not increase locomotor activity, even though binding was inhibited to 38 and 37% of control levels in the striatum and nucleus accumbens, respectively. Furthermore, our present results suggest that locomotor activity does not continue to increase as the blockade of DAT increases. Notably, there were no increases in locomotor activity at the dose of RTI-76 (100 nmol) used to measure DAT half-life. PMID- 11282371 TI - Effect of Agouti-related protein delivered to the dorsomedial nucleus of the hypothalamus on intake of a preferred versus a non-preferred diet. AB - Agouti-related protein (Agrp), a high-affinity antagonist of the melanocortin-3/4 receptors, increases feeding when administered centrally. Previous studies have shown that this increase is long-lasting (at least 24 h) and delayed, unless the animal is first stimulated to feed by fasting or onset of the dark phase. The present studies first demonstrate that long-lasting and delayed increases in food intake are also evident when Agrp is microinjected into the dorsomedial nucleus of the hypothalamus (DMH). Next, the effects of DMH-administered Agrp were assessed on intake of two foods, isocaloric but differing in flavor (with or without sucrose). Following exposure to the two diets, rats were injected via cannula aimed at the DMH with 100 pmol Agrp at 10:00 h and allowed ad libitum access to either: (1) a choice of both diets or (2) one of the diets alone. Food intake was determined at 2, 4, and 24 h post-injection. In the first (choice) paradigm, Agrp only increased intake of the sucrose-containing diet. In the second (no-choice) paradigm, animals on either diet showed an Agrp-induced increase in intake 24 h following injection; only animals on the sucrose containing diet showed an increase in intake 4 h post-injection. The results are discussed in the context of the possible involvement of Agrp/MC4-R in the rewarding characteristics of food intake. PMID- 11282372 TI - Iron overload following manganese exposure in cultured neuronal, but not neuroglial cells. AB - Our previous studies show that manganese (Mn) exposure inhibits aconitase, an enzyme regulating the proteins responsible for cellular iron (Fe) equilibrium. This study was performed to investigate whether Mn intoxication leads to an altered cellular Fe homeostasis in cultured neuronal or neuroglial cells as a result of disrupted Fe regulation. Our results reveal a significant increase in the expression of transferrin receptor (TfR) mRNAs and a corresponding increase in cellular 59Fe net uptake by PC12 cells, but not astrocytes, following Mn exposure. These findings suggest that alteration by Mn of cellular Fe homeostasis may contribute to Mn-induced neuronal cytotoxicity. PMID- 11282373 TI - Phase-dependent modulation of cutaneous reflexes of tibialis anterior muscle during hopping. AB - During human gait, the amplitude of cutaneous reflexes in the leg is modulated as a function of the phase of the step cycle. In tibialis anterior (TA) the responses to sural nerve stimulation are facilitatory at end stance while they are suppressive at end swing. To investigate in how far this modulation is specifically related to alternating locomotion, the modulation of such reflexes was studied during a symmetric rhythmic movement, namely hopping (as equivalent of galloping). The end-stance facilitation was present during hopping while the end-swing suppression was absent. It is concluded that the end-stance facilitation is not specific for alternating gait. The absence of the end-swing suppressive reflexes may be related to the absence of heel strike in hopping. PMID- 11282374 TI - Effect of different concentrations of iontophoretic nociceptin on distinct classes of nociceptive neurons in rat spinal cord. AB - Iontophoretically applied nociceptin (NC) was tested at different concentrations on the activity of spinal nociceptive specific (NS) and wide dynamic range (WDR) neurons. Low NC dosages inhibited the noxious response of NS neurons, higher dosages inhibited the noxious responses of the WDR neurons but had little effect on the non-noxious response. Naloxone did not antagonize the NC effect. Thus, appropriate dosages of NC may be selective, both for neuronal classes and for sensory modalities. PMID- 11282375 TI - FMRI mapping of the somatosensory cortex with vibratory stimuli. Is there a dependency on stimulus frequency? AB - Vibratory stimuli on the skin are mediated by two major receptors: Meissner corpuscles and Pacinian corpuscles. These receptors differ in properties such as density distribution, receptive field size, frequency sensitivity and depth of location. The cortical response to stimulation of these corpuscles can be tested by taking advantage of the differences in frequency discrimination of the receptors. Meissner corpuscles are most sensitive to frequencies around 10-50 Hz (flutter), while Pacinian corpuscles are most sensitive to high frequency (100 300 Hz) vibration. This study compared the neuronal responses (hemodynamic response) generated from vibratory stimuli of 35 Hz and 150 Hz with functional MRI. Group functional activation maps showed differences in the activation pattern for the two stimulus frequencies. PMID- 11282376 TI - Age-related changes in the distribution of Kv1.1 and Kv1.2 channel subunits in the rat cerebellum. AB - We have revealed age-related changes in the expression patterns of Kv1.1 and Kv1.2 in the rat cerebellum for the first time. In the aged rat, immunoreactivity for Kv1.1 was increased in the cell bodies of Purkinje cells, while the staining intensity was significantly decreased in the granule cells. The cell bodies of cerebellar output neurons showed strong Kv1.1 immunoreactivity in the nucleus medialis, interpositus and lateralis of the aged rat. Kv1.2 immunoreactivity was found in some interneurons with their processes in this region of the aged rat. Image analysis demonstrated that immunoreactivities for Kv1.1 and Kv1.2 were increased specifically in the cell bodies of cerebellar output neurons of the aged rat. This study may provide useful data for future investigations on the channels that cause brain diseases and age-related disorders. PMID- 11282377 TI - Voltage-gated K+ channels in chemoreceptor sensory neurons of rat petrosal ganglion. AB - A subpopulation of sensory neurons in the petrosal ganglion transmits information between peripheral chemoreceptors (glomus cells) in the carotid body and relay neurons in the nucleus of the solitary tract. Expression of voltage-gated K+ channels in these neurons was characterized by immunohistochemical localization. Five members of the Kv1 family, Kv1.1, Kv1.2, Kv1.4, Kv1.5 and Kv1.6 and members of two other families, Kv2.1 and Kv4.3, were identified in over 90% of the chemoreceptor neurons. Although the presence of these channel proteins was consistent throughout the population, individual neurons showed considerable variation in K+ current profiles. PMID- 11282378 TI - MK-801 is cytotoxic to microglia in vitro and its cytotoxicity is attenuated by glutamate, other excitotoxic agents and atropine. Possible presence of glutamate receptor and muscarinic receptor on microglia. AB - We examined the cytotoxicity of MK-801 on cultured microglia and demonstrated its cytotoxicity. Cytotoxicity of MK-801 was reduced by the addition of L-glutamate, kainate and NMDA. The action of MK-801 was due to the direct action of microglia. It suggested the existence of glutamate receptor in microglia. Cytotoxicity of MK 801 was reduced by the addition of atropine sulfate which suggested the presence of muscarinic receptor in microglia. PMID- 11282379 TI - The activation of mu opioid receptors promotes a small modulation of calcium currents in rat pallidal neurons. AB - Globus pallidus receives, from dorsal neostriatum, a dense enkephalinergic innervation whose role is still uncertain. We examined the possibility that the activation of mu, delta or k opioid receptors modulate high-voltage-activated calcium currents in isolated GP neurons. Neither dynorphin nor DPEPE inhibited calcium current, whilst DAMGO produced a small (-16%) but consistent response, selectively antagonized by CTOP. The mu-mediated modulation required the activation of G-proteins but was voltage-independent. The pre-incubation in omega conotoxinVIA abolished the response, implying the involvement of N-type calcium channels. These findings suggest that enkephalin may exert a direct influence on GP excitability also through post-synaptic effects. In degenerative conditions as Parkinsonism, an excessive stimulation of mu binding sites might induce a pathological inhibition of calcium signals, thus contributing to modify the GP firing pattern and transmitter release. PMID- 11282381 TI - A single trial analysis of hippocampal theta frequency during nonsteady wheel running in rats. AB - It has been suggested that hippocampal theta rhythm codes some aspects of motor behavior, but previous studies of the correlation between theta frequency and steady whole body locomotion speed using both linear tracks and wheels have provided inconsistent if not contradictory results. Because the relationship between temporal dynamics of theta frequency and non-steady (or dynamic) whole body locomotion speed can help clarify this issue, single trials of hippocampal EEG were analyzed together with nonsteady wheel running speed recorded during rats perform spontaneous normal locomotion in a wheel. Changes in theta frequency within single trials show positive or negative correlation with nonsteady wheel running speed. As the mean running speed increases and the standard deviation of running speed decreases in a single trial, the correlation between temporal dynamics of theta frequency and nonsteady wheel running speed within the single trial tends to be positive. In addition, we found that the amount of deceleration is also related to the polarity of the correlation coefficients. PMID- 11282380 TI - NMDA glutamate receptor stimulation is required for the expression of D2 dopamine mediated responses in apomorphine primed 6-hydroxydopamine lesioned rats. AB - Three priming injections with the D1/D2 dopamine agonist apomorphine permits a challenge with the D2 agonist quinpirole to elicit robust contralateral rotation and ipsilateral striatal Fos expression in 6-hydroxydopamine lesioned rats. Pretreatment with NMDA glutamate antagonists MK-801 or CPP dose-dependently attenuates these quinpirole-mediated responses. These findings suggest that concomitant NMDA receptor stimulation is required for the expression of D2 mediated responses in apomorphine primed dopamine-depleted rats. PMID- 11282382 TI - Organizing sound sequences in the human brain: the interplay of auditory streaming and temporal integration. AB - The present study examined the relationship between two of the early brain processes of sound organization: auditory streaming and the temporal window of integration (TWI). Presented at a fast stimulus delivery rate, two tones alternating in frequency are perceived as separate streams of high and low sounds. However, when two sounds are presented within a ca. 200 ms temporal window, they are often processed as a single auditory event. Both stream segregation and temporal integration occur even in the absence of focused attention as was shown by their effect on the mismatch negativity (MMN) event related potential. The goal of the present study was to determine the precedence between these two sound organization processes by using the stimulus-omission MMN paradigm. Infrequently omitting one stimulus from a homogeneous tone sequence only elicits an MMN when the stimulus onset asynchrony separating successive tones is shorter than 170 ms. This demonstrates the effect of the TWI. Magnetic brain responses elicited by infrequent stimulus omissions appearing in a sequence of two alternating tones were recorded. The magnetic MMN was elicited by tone omission when the alternating tones formed a single stream (with no or only small frequency separation between the two tones) but not when separate high and low streams emerged in perception (large frequency separation between the two alternating tones). This result shows that auditory streaming takes precedence over the processes of temporal integration. PMID- 11282383 TI - The cardiovascular and behavioral response to cat odor in rats: unconditioned and conditioned effects. AB - Cardiovascular and behavioral responses were recorded in rats during exposure to cat odor. Rats were habituated to an open rectangular arena that contained a small enclosed wooden box in which they could hide. On day 1 of the experiment, after 30 min in the apparatus, rats were presented with a piece of fabric collar for 60 min. On day 2, rats were presented with an identical piece of fabric collar, except that it had been worn by a cat and therefore exuded cat odor. On day 3, rats were again presented with an unworn cat collar, to determine any conditioned responses to the environment or stimulus (collar) previously associated with cat odor. Results showed significantly increased blood pressure and decreased activity during exposure to cat odor as well as avoidance of the odor stimulus and an increase in vigilance and risk-assessment measures. No significant change in heart rate was found during cat odor exposure. On day 3, a transient increase in blood pressure was seen as well as reduced activity and a range of defensive behaviors. This suggests some conditioning of fear to a context in which cat odor had previously been experienced. Heart rate was also significantly decreased on day 3. A transient rise in blood pressure was also seen when the unworn cat collar was placed into the apparatus on day 3, suggesting a conditioned response to a stimulus that has been previously associated with cat odor. This study demonstrates that a natural stressful stimulus can induce both unconditioned and conditioned autonomic and behavioral responses. PMID- 11282384 TI - Role of vagal afferents in the reflex effects of capsaicin and lobeline in monkeys. AB - Reflex effects of pulmonary C-fiber receptor stimulation by right atrial injections of capsaicin and lobeline were investigated in conscious monkeys (n=17). Capsaicin injection (15.0+/-1.4 microg/kg) produced apnea mostly (n=15, latency-1.7+/-0.2 s) and bradycardia, which were abolished by vagotomy (n=4). Lobeline administration (142+/-6 microg/kg) produced either apnea (n=7, latency 2.0+/-0.3 s) or excitation of breathing (n=8, latency -3.5+/-0.3 s) and no change in heart rate. After vagotomy (n=4), the apneic response was abolished, but the respiratory excitation persisted. Neither capsaicin nor lobeline produced cough. In the anesthetized monkey also (n=7), lobeline injection (50-150 microg/kg) did not produce any cardiovascular response. However, it produced excitation of breathing, which persisted after vagotomy but was abolished by carotid sinus denervation. It is concluded that in the non-human primate, it is capsaicin that produces reflexes typical of pulmonary C-fiber receptor stimulation, and cough is not a part of this reflex. PMID- 11282385 TI - Alpha2A-adrenoceptor mediated tachypnea in awake goats. AB - To further elucidate the role of alpha2-adrenoceptors (alpha2-ARs) in the control of respiratory rhythm we examined the ventilatory effects of guanfacine (a preferentially selective alpha2A-AR agonist) and clonidine (a non-selective alpha2-AR agonist) in awake adult goats. Systemic administration of guanfacine in cumulative doses (20 microg/kg; 140-180 microg/kg total cumulative dose) increased breathing in all animals in a dose-dependent manner. The excitatory effect was entirely mediated by increases in respiratory frequency. The magnitude of the guanfacine-induced tachypnea was similar to that produced by systemic administration of cumulative doses of clonidine (1-2 microg/kg; 4-10 microg/kg total cumulative dose) in the same animals studied on a separate day. Both guanfacine- and clonidine-induced tachypnea was reversed by the preferentially selective alpha2A-AR antagonist RX821002 (2-6 microg/kg IV). Unlike clonidine however, guanfacine administration did not produce slow arrhythmic breathing episodes (irregular TE intervals and central apneas) that are characteristic of alpha2-AR stimulation with alpha2-AR agonists in the awake goat. The results suggest that alpha2-AR agonist-induced ventilatory excitation (tachypnea) requires the activation of alpha2A-ARs whereas clonidine-induced ventilatory depression (arrhythmic breathing) requires the activation of an alternate alpha2 AR subtype (presumably alpha2C-ARs). The results further demonstrate that alpha2 AR pathways exert an important influence on respiratory rhythm in the awake goat. PMID- 11282386 TI - The influence of NMDA receptor-mediated processes on breathing pattern in ground squirrels. AB - The effects of blockade of N-methyl-D-aspartate (NMDA) type glutamate receptors by a non-competitive antagonist (MK-801) on cortical arousal, breathing pattern and ventilatory responses to hypoxia (10% O2 in N2) and hypercapnia (5% CO2 in air) were assessed in anesthetized (urethane) and unanesthetized golden-mantled ground squirrels (Spermophilus lateralis). Intra-cerebroventricular administration of MK-801 did not alter ventilation during wakefulness, although it did alter the pattern (breathing frequency and tidal volume components) of the hypercapnic ventilatory response, and suppressed the ventilatory response to hypoxia. Animals did not sleep following treatment with MK-801, and intravenous administration of MK-801 prevented expression of the sleep-like state normally observed in anesthetized animals. In anesthetized animals MK-801 elevated breathing frequency to levels observed without anesthesia, and suppressed the hypoxic ventilatory response. These data suggest that NMDA-type glutamatergic receptor-mediated processes influence cortical arousal and facilitate depression of breathing frequency during anesthesia and the hypoxic ventilatory response. Such processes are not essential for the hypercapnic ventilatory response. PMID- 11282387 TI - Vagal feedback is essential for breathing in unanesthetized ground squirrels. AB - The roles of vagal afferent feedback in terminating inspiration and modulating breathing pattern and ventilatory responses to hypoxia and hypercapnia were assessed in the golden-mantled ground squirrel, Spermophilus lateralis, during wakefulness and urethane anesthesia. Hypoxia increased ventilation primarily through increases in breathing frequency (f(R)) while hypercapnia increased ventilation primarily through increases in tidal volume (V(T)) in both anesthetized and unanesthetized animals. Vagotomy resulted in an increase in tidal volume, a decrease in breathing frequency and ventilation, and depressed ventilatory responses to both hypoxia and hypercapnia in anesthetized animals. In unanesthetized animals vagotomy produced a transient 'gasp-like' breathing pattern that rapidly progressed to a non-obstructive central apnea. These data indicate that vagal feedback shapes ventilation on a breath-by-breath basis during anesthesia and is essential for ventilation in unanesthetized animals. The mechanisms that transform the influences of vagal input on breathing between anesthetized and unanesthetized states remain unclear. Changes in breathing pattern induced by the removal of vagal feedback compromise chemoreflexes. PMID- 11282388 TI - Alpha2 adrenergic receptors and the central control of breathing in the cane toad, Bufo marinus. AB - The effect of adrenergic agents on the central control of breathing in the cane toad, Bufo marinus, was tested by applying adrenergic agents to the ventral medullary surface of decerebrate adult toads. Toads were unidirectionally ventilated while recording lung, buccal, and artery blood pressures (BPI), as well as heart rate (HR). Following a control period, filter paper pledgets soaked in the appropriate solution (epinephrine - 0.023, 0.05, 0.10, and 0.23 mM; norepinephrine - 0.002, 0.016, 0.032, and 0.16 mM; clonidine - 0.00375, 0.0375, and 0.375 mM; or yohimbine - 0.43) mM) were placed bilaterally on the ventral medullary surface. Epinephrine significantly increased the number of breaths (26%), lung amplitude (9%), and episode duration (21%), but had no effect on BP or HR. The alpha2-agonist. clonidine, significantly increased respiratory activity at moderate doses (0.0375 mM) and decreased activity at high doses (0.375 mM), however, it failed to elicit significant changes in BP or HR. Pretreatment with the alpha2-adrenoceptor antagonist, yohimbine (0.43 mM), blocked the clonidine induced changes in respiratory activity. Yohimbine had no effect on cardiorespiratory parameters. Norepinephrine had no effect on either cardiovascular or respiratory variables. Thus, it appears that an alpha2 adrenergic mechanism is involved in the central control of respiration in this lower vertebrate. PMID- 11282389 TI - Ventilation is greater in women than men, but the increase during acute altitude hypoxia is the same. AB - We wished to determine whether the previously reported lower arterial or alveolar P(CO2) in women than men, and in luteal (LUT) compared with follicular (FOL) menstrual cycle phase would persist during normal oral contraceptive use and during early altitude exposure. Ventilation and blood gases were measured at baseline (636 mmHg approximately 5400 ft, 1650 m) and during simulated altitude at 426 mmHg ( approximately 16000 ft, 4880 m), after 1 h (A1) and during the 12th h (A12), in 18 men (once) and in 19 women twice, during LUT and FOL and in 20 women twice while on placebo (PLA) or highest progestin dose (PIL) oral contraceptives. At baseline, Pa(CO2) was significantly higher in men than all women by 3.3 mmHg. When progesterone-progestin (PRO) was elevated in women, Pa(CO2) was significantly lower than in FOL and PLA, but the latter were still significantly lower than men. At altitude the P(CO2) differences between men and women and PRO levels persisted, with PA(CO2) falling by 3.6 and 7.3 mmHg at A1 and A12 in all, indicating an equivalent increase in alveolar ventilation. The mean arterial-end tidal P(CO2) difference was never >2 mmHg in the groups, indicating no VA/Q mismatch related to gender, PRO levels or altitude. All women had higher breathing frequency than men, resulting in greater deadspace ventilation. At altitude, the mean Pa(O2) was approximately 44 mmHg (Sa(O2) approximately 79%) for all, indicating equivalent oxygenation, but alveolar arterial P(O2) differences were greater in women than men and higher when PRO was elevated. These results show that, relative to men, women have a compensated respiratory alkalosis, accentuated with elevated PRO. However, the ventilation response to acute altitude is the same in women and men. PMID- 11282390 TI - A comparative study on cockroach and ovalbumin sensitizations and challenge responses in Hartley guinea-pigs. AB - The role of allergens in asthmatic inflammation is not clearly understood. To elucidate the mechanism of cockroach allergen (CRa)-induced airway disease, we studied three groups of Hartley guinea-pigs sensitized to control, ovalbumin (OA) or CRa. Parameters measured were anaphylactic antibodies by allergy skin test (AST), PCA assay and Western blot, changes in specific airway resistance (SRaw), analysis of bronchoalveolar lavage (BAL) and contracture responses of tracheal muscle (TSM) to non-specific and specific stimuli, in vitro. Both OA and CRa animals showed a similar allergic sensitization (AST and PCA), while Western blot identified several reaginic bands in CRa group compared to a single band in OA group. SRaw illustrated that CRa induce dual-asthmatic responses (4/6) in the CRa group, whereas OA induce only an early asthmatic response (3/6) in the OA group (P<0.01). The average total leukocytes in BALF of the CRa were 27.0x10(6), mostly neutrophils and eosinophils, while those of the OA showed 3.5x10(6), mostly eosinophils, respectively (P<0.0001). TSM responses to non-specific stimuli were similar in both groups (P>0.1), while the antigen-specific TSM contractions were more brisk in the OA group than those of CRa group (P<0.001). Thus, the study indicates that both CRa and OA sensitize guinea-pigs, yet CRa induces more severe and persistent late-phase inflammation than OA. This appears to be related to an influx of neutrophils rather than anaphylactic bronchospasm. PMID- 11282391 TI - Inotrophic effects of the K(+) channel blocker TEA on dystrophic (mdx and dy/dy) mouse diaphragm. AB - K(+) channels regulate diaphragm resting membrane potential and action potential duration, and hence force. Certain blockers of these channels, e.g. tetraethylammonium (TEA), increase twitch force of normal diaphragm. To further address whether these agents may be useful in the treatment of diaphragm weakness, studies examined the effects of TEA on force of overtly diseased muscle. Diaphragm from two mouse models of muscular dystrophy (mdx and dy/dy) was studied in vitro. Diaphragm from both models was significantly weaker than diaphragm from control animals. TEA (10 mM) increased twitch force of both mdx diaphragm (P<0.005) and dy/dy diaphragm (P<0.0005), as well as force of diaphragm from non-diseased animals. The percent force increase of mdx diaphragm was at least as great as that of non-diseased muscle (15.3 vs 9.2%, P=0.14), and the percent force increase of dy/dy diaphragm was significantly greater than that of non-diseased muscle (22.7 vs 10.2%, P<0.02). Absolute force increases normalized for cross-sectional area were comparable for healthy and diseased diaphragm, however. These findings indicate that TEA increases force of both dystrophin deficient and merosin-deficient dystrophic mouse diaphragm muscle. PMID- 11282392 TI - Structural and functional properties of membrane and secreted IgD. AB - More than 35 years ago, study of an unknown immunoglobulin (Ig) in the serum from a myeloma patient led to the discovery of IgD. Subsequently, the finding that it also exists as a membrane-bound Ig stimulated a large number of studies during the 70s. Then, the interest on IgD shrank, largely because of the lack of known function of secretory IgD (secIgD) and of a stagnating knowledge of the functions of surface IgD. In the recent years, very significant advances followed the tremendous accumulation of data on the physiology of the B cell receptor, of which IgD is the major component, on the role of secIgD in normal and diseased individuals. This review, which is focused on human IgD but integrates data in the mouse and other species when needed, summarizes present data on the structure, synthesis and functions of both membrane and secIgD, IgD receptors and the involvement of IgD in various diseases, especially the hyperIgD syndrome. PMID- 11282393 TI - The lymphoid-specific cofactor OBF-1 is essential for the expression of a V(H) promoter/HS1,2 enhancer-linked transgene in late B cell development. AB - Mice deficient for the lymphoid-specific cofactor OBF-1 display reduced levels of IgG, IgA and IgE. To examine whether the lowered immunoglobulin expression is linked to reduced activity of IgH cis-regulatory elements, OBF-1(-/-) mice were crossed with mice expressing transgenes driven by a V(H) or beta-globin promoter linked to the HS1,2 enhancer. Here we show that OBF-1 is essential for the induced expression of a V(H) promoter-linked transgene, in contrast to a beta globin promoter-dependent transgene, in LPS/IL-4 or CD40-stimulated splenic B cells. Furthermore, impaired transgene expression is observed in OBF-1(-/-) peritoneal B cells. This deficiency may be linked to OBF-1, as peritoneal cells from normal mice express OBF-1 protein constitutively. Our data link OBF-1 to IgH gene expression in late B lymphoid development. PMID- 11282394 TI - Molecular analysis of cross-reactive anti-myosin/anti-streptococcal mouse monoclonal antibodies. AB - Nucleotide sequences of VH- and VL-genes of anti-myosin/anti-streptococcal monoclonal antibodies (mAbs) were analyzed and compared with their highly detailed antigen binding reactivities. Antigen-specificities of the cross reactive mAbs included myosin, streptococcal M-protein, actin, keratin, N-acetyl beta-D-glucosamine, vimentin, DNA, tropomyosin, troponin, and laminin as previously described. After nucleotide sequence analysis, homology indicated that some of the V gene sequences aligned with antibodies recognizing gangliosides and blood group antigens glycophorin M and N. Therefore, mAb reactivity with gangliosides and glycophorin M and N was identified. The cross-reactive mAbs utilized a heterogeneous group of germline V-heavy genes comprised of nine J558-, four 7183- and two Q52-family VH-genes. Germline V-light genes utilized by the mAbs included six Vkappa4/5-, three Vkappa8-, two Vkappa10-, three Vkappa19- and one Vkappa23-family VL-genes. No preferential VH/VL-chains correlated with any of the 12 different antigen reactivities, even for mAbs with nearly identical cross reactivities. However, we did find that the cross-reactive mAb germline genes within a V gene family shared more homology among themselves than with other germline genes within their V gene families, suggesting convergent mutation. Cross-reactive mAbs with the highest relative avidity for myosin were found in the VH7183 family which contained two cytotoxic mAbs. Antibodies with V gene sequences most homologous to those of our cross-reactive anti-myosin/anti streptococcal mAbs had specificities for laminin, DNA, carbohydrates, or blood group antigens and were reported to cause autoimmune disease in mice. PMID- 11282396 TI - Interaction of dual-specific autoantibodies with dsDNA and a synthetic dimer peptide simulating the hinge region of IgG2a molecules. AB - Anti-dsDNA autoantibodies and immune complex formation are major factors in SLE pathogenesis. Understanding stable immune complex formation is critical in deciphering mechanisms of autoimmune pathogenesis. Previous studies identified a subpopulation of murine lupus monoclonal autoantibodies that exhibited dual specificity (anti-DNA and anti-IgG2a hinge) and formed stable immune complexes [J. Mol. Rec. 10(1997)225]. Two monoclonal autoantibodies, BV 17-45 and BV 16-13, were extensively studied because of their dual specificity. To quantitatively assess the role of each specificity in the formation of stable immune complexes, studies were performed to determine binding affinities for various sized dsDNA fragments (21, 43, 84, and 114 bp) and the covalent dimer of a nine amino acid hinge peptide. Results characterizing BV 17-45 showed that the affinity for dsDNA directly correlated with increased dsDNA size. Results with BV 16-13 revealed a generally lower affinity for the various dsDNA fragments. Binding inhibition studies, using a covalently linked dimer of a nine amino acid synthetic hinge peptide as an inhibitor of the antibody-43 bp dsDNA interaction, yielded relative affinities for the anti-hinge activity. Binding affinities for the synthetic hinge specificity were lower than affinities measured for the anti-dsDNA activity. Collectively, the binding and inhibition studies provided insight into the correlation between dual specificity and avid immune complex formation. A model was proposed based on the concept that large dsDNA fragments caused localization of the dual-specific antibodies through the anti-dsDNA activity, thereby facilitating subsequent binding and cross-linkage via the anti-hinge specificity. These synergistic interactions resulted in the formation of avid immune complexes. PMID- 11282395 TI - Normal V(D)J recombination in cells from patients with Nijmegen breakage syndrome. AB - The majority of antigen receptor diversity in mammals is generated by V(D)J recombination. During this process DNA double strand breaks are introduced at recombination signals by lymphoid specific RAG1/2 proteins generating blunt ended signal ends and hairpinned coding ends. Rejoining of all DNA ends requires ubiquitously expressed DNA repair proteins, such as Ku70/86 and DNA ligase IV/XRCC4. In addition, the formation of coding joints depends on the function of the scid gene encoding the catalytic subunit of DNA-dependent protein kinase, DNA PK(CS), that is somehow required for processing of coding end hairpins. Recently, it was shown that purified RAG1/2 proteins can cleave DNA hairpins in vitro, but the same activity was also described for a protein complex of the DNA repair proteins Nbs1/Mre11/Rad50. This leaves the possibility that either protein complex might be involved in coding end processing in V(D)J recombination. We have therefore analyzed V(D)J recombination in cells from patients with Nijmegen breakage syndrome, carrying a mutation in the nbs1 gene. We find that V(D)J recombination frequencies and the quality of signal and coding joining are comparable to wild-type controls, as analyzed by a cellular V(D)J recombination assay. In addition, we did not detect significant differences in CDR3 sequences of endogenous Ig lambdaL and kappaL chain gene loci cloned from peripheral blood lymphocytes of an NBS patient and of healthy individuals. These findings suggest that the Nbs1/Mre11/Rad50 complex is not involved in coding end processing of V(D)J recombination. PMID- 11282397 TI - Studies on the energy release of rice mitochondria under different conditions by means of microcalorimetry. AB - The thermodynamic and kinetic behaviors of energy release of mitochondria isolated from rice (Oryza sative L.) were studied by using a LKB 2277 Bioactivity Monitor under different conditions. The thermogenesis curves of energy release of the rice mitochondria (which had been kept at 0-3 degrees C for 15 h and 40 day before the determination) were determined respectively at 25 and 30 degrees C, and the difference in shape of the thermogenesis curves and thermodynamic and kinetic characteristics were compared. The thermodynamic and kinetic parameters of energy release of the mitochondria in the thermogenesis increasing stage have been calculated, and the experimental thermokinetic equations of the thermogenesis have been established. The results indicated that the lower the temperature, the slower the energy release of the rice mitochondria. Both the thermogenesis and the energy release rate of the rice mitochondria increased after the mitochondria was kept at lower temperature for 40 days. One can use the methods to characterize the ability of the rice mitochondria to release energy under different conditions. PMID- 11282398 TI - An antigen capture assay for the measurement of serum clusterin concentrations. AB - We have developed and validated a robust antigen capture assay for the measurement of serum clusterin. Increased clusterin expression, and alterations in serum clusterin levels have been associated with a number of disease states. In particular, clusterin has been shown to be associated with tissue regression and apoptosis in the rat ventral prostate in response to androgen ablation or administration of anti-androgens. The object of this study was to determine if changes in human serum clusterin can be used as a diagnostic or prognostic marker to monitor the response to hormonal therapy in patients with prostate cancer, and to determine if clusterin concentrations increase with the progression towards androgen independence. The antigen capture assay was used for an extensive analysis of human serum clusterin concentration in fasting males, and to determine if there is any relationship between clusterin and age or cholesterol levels. The average clusterin level in serum is 101+/-42 microg/ml (n=96). There is no correlation to age or serum cholesterol levels. Analysis of serum clusterin levels in patients with newly diagnosed prostate cancer (n=5), hormone responsive tumors (n=5), and hormone refractory disease (n=5), demonstrates that no significant changes in serum clusterin levels accompany the progression of prostatic disease, or response to hormone therapy. PMID- 11282399 TI - Pepsin digestion of a mouse monoclonal antibody of IgG1 class formed F(ab')(2) fragments in which the light chains as well as the heavy chains were truncated. AB - In the preparation of F(ab')(2) fragments of monoclonal antibodies (mAbs) of IgG class, heavy (H) chains are truncated by pepsin and light (L) chains are remained intact. However, F(ab')(2) fragments formed by pepsin-digestion of a mouse mAb PM373, which was of the IgG1 class and raised against human prostate specific antigen (PSA), indicated that the L chains of 31 kDa were cleaved into 23-kDa fragments as well as the cleavage of H chains of 50 kDa into 28-kDa fragments. On the other hand, F(ab')(2) fragments formed by digesting the mAb by cathepsin D showed that the L chains were intact and the H chains were truncated. The immunoreactivities against PSA of the F(ab')(2) fragments containing the intact L chains and those containing the truncated L chains were almost the same as that of the parental mAb, suggesting that the truncation of the L chains does not affect the interaction of the mAb with its specific antigen. PMID- 11282400 TI - Data plotting of warfarin binding to human serum albumin. AB - The binding of warfarin to human serum albumin was studied by equilibrium dialysis at pH 7.4 in a 67 mM sodium phosphate buffer at 37 degrees C. The equilibrium data were analysed using a computer program for curve fitting. The analysis was made fitting the data to equations for one, two and three classes of binding sites with one, two and three sites at the primary binding site (n(1)=1, 2 or 3). The data fitting was acceptable for two and three classes of binding sites but the best fit was obtained with the equation for two classes of binding sites, allowing us to define the binding by a model with two independent classes of binding sites on the serum albumin molecule. PMID- 11282402 TI - Synthesis and characterization of immobilized dopamine beta-hydroxylase in membrane-bound and solubilized formats. AB - Dopamine beta-hydroxylase (DBH) catalyzes the beta-hydroxylation of dopamine to norepinephrine. The enzyme in chromaffin granules occurs in a soluble form and a form confined to the surrounding membrane. DBH was noncovalently immobilized in the hydrophobic interface of an immobilized artificial membrane (IAM) liquid chromatographic stationary phase and the resulting DBH-IAM stationary phase was enzymatically active and was shown to mimic the membrane-bound form of the enzyme. DBH was also covalently immobilized onto a silica-based support containing, glutaraldehyde-P (Glut-P). The resulting DBH-Glut-P interphase was also enzymatically active, reproducible and shown to display characteristics of the solubilized enzyme. The results demonstrate that the different immobilization methods utilized for the enzyme can be used to quantitatively and qualitatively determine the enzyme kinetic constants associated with enzyme/substrate and enzyme/inhibitor interactions for the two distinct forms of the enzyme. These new entities can be used in basic biochemical studies as well as in high throughput screening of substances for DBH substrate/inhibitor properties. PMID- 11282401 TI - Depth of immersion of fluorescent chromophores in biomembranes studied by quenching with nitroxide radical. AB - A novel method has been developed for measuring depth of immersion of a fluorescent chromophore in biological matrices, such as biomembranes. The method is based on dynamic quenching of chromophore fluorescence by a nitroxide probe freely diffused in solution. Theoretical considerations and experimental evidences relating to the method are discussed. The proposed method was applied to investigation of lecithin liposomes and membranes from Bacillus subtilis modified by the photochrome-fluorescent probe 4,4'-dimethylaminocyanostilbene. PMID- 11282403 TI - A simple method for midkine purification by affinity chromatography with a heavy chain variable domain (VH) fragment of antibody. AB - A DNA fragment for a heavy chain variable domain (VH) was prepared from a hybridoma that produces a monoclonal antibody against human midkine (MK). The antibody fragment was produced in Escherichia coli and its affinity for chemically synthesized full length MK or recombinant midkine c-terminus (MKc half) protein was confirmed by ELISA. An Escherichia coli cell lysate expressing MKc-half was applied to a VH fragment-coupled Sepharose 4B column and eluted with a buffer containing 0.5 M NaCl. SDS-PAGE analysis revealed a high degree of purity of the MKc-half protein in the eluent, showing the utility of a recombinant VH fragment in purification of proteins by affinity chromatography. PMID- 11282404 TI - From genes to effective drugs for neurological and psychiatric diseases. PMID- 11282405 TI - Mining the genome for causes and cures of neurological disease. PMID- 11282407 TI - The grand design. PMID- 11282406 TI - A compendium of specific motifs for diagnosing GPCR subtypes. AB - Analysis of G-protein-coupled receptor (GPCR) subtypes has attracted considerable interest because some drugs that act on GPCRs cause therapeutic problems as a result of their failure to differentiate between subtypes. In this article, an extensive compendium of diagnostic 'fingerprints' for GPCR subtypes and their families will be described. These fingerprints offer new opportunities to investigate correlations between specific sequence motifs and ligand binding or G protein coupling, and are likely to prove valuable both in seeking novel receptors in genome data and in the characterization of orphan receptors. PMID- 11282408 TI - Transcription factors move with the glow. PMID- 11282416 TI - Cysteine mutants as chemical sensors for ligand-receptor interactions. AB - The incorporation of cysteine residues into membrane receptors by mutagenesis has enabled the development of engineered proteins. Chemical modification of the mutant receptor using a wide range of biochemical and biophysical probes has facilitated functional studies of ligand-receptor interactions. In particular, the substituted-cysteine accessibility method (SCAM) represents a successful example of how to probe transmembrane receptor domains after chemical modification of the mutants with sulfydryl-reacting molecules. We propose an extension of this methodology using site-specific affinity probes that react with cysteine mutants to gain reliable structural information on the binding of a ligand in its receptor site. PMID- 11282417 TI - Glutamate signalling in non-neuronal tissues. AB - Since the discovery of its role in the CNS, glutamate, together with its involvement in signalling at synapses, has been the subject of a vast amount of research. More recently, it has become clear that glutamate signalling is also functional in non-neuronal tissues and occurs in sites as diverse as bone, pancreas and skin. These findings raise the possibility that glutamate acts as a more widespread 'cytokine' and is able to influence cellular activity in a range of tissue types. The impact of these discoveries is significant because they offer a rapid way to advance the development of therapeutics. Agents developed for use in neuroscience applications might be beneficial in the modulation of pathology peripherally, impacting on conditions such as osteoporosis, diabetes and wound healing. PMID- 11282418 TI - Purine-mediated signalling in pain and visceral perception. AB - Receptor subtypes for purines have been identified in a variety of tissues, increasing interest in the roles of purine-mediated signalling in pathophysiological processes. Growing evidence supports the involvement of one of the purinoceptor subtypes, P2X3, in nociception. In this article, recent studies of purine-mediated nociception and visceral pain will be discussed. Furthermore, a novel hypothesis is proposed for purine-mediated mechanosensory transduction where ATP released during distension from epithelial cells lining tubes (such as ureter and gut) and sacs (such as the bladder) acts on P2X3 receptors on a subepithelial nerve plexus to initiate impulses that are relayed via the spinal cord to pain centres in the brain. PMID- 11282419 TI - GABA(A) receptor subtypes: dissecting their pharmacological functions. AB - The enhancement of GABA-mediated synaptic transmission underlies the pharmacotherapy of various neurological and psychiatric disorders. GABA(A) receptors are pluripotent drug targets that display an extraordinary structural heterogeneity: they are assembled from a repertoire of at least 18 subunits (alpha1-6, beta1-3, gamma1-3, delta, epsilon, theta, rho1-3). However, differentiating defined GABA(A) receptor subtypes on the basis of function has had to await recent progress in the genetic dissection of receptor subtypes in vivo. Evidence that the various actions of allosteric modulators of GABA(A) receptors, in particular the benzodiazepines, can be attributed to specific GABA(A) receptor subtypes will be discussed. Such discoveries could open up new avenues for drug development. PMID- 11282420 TI - Biological and clinical implications of the MTHFR C677T polymorphism. AB - The enzyme methylenetetrahydrofolate reductase (MTHFR) directs folate species either to DNA synthesis or to homocysteine (Hcy) remethylation. The common MTHFR C677T polymorphism affects the activity of the enzyme and hence folate distribution. Under conditions of impaired folate status, the homozygous TT genotype has been regarded as harmful because it is associated with a high concentration of plasma total Hcy, increased risk of neural tube defects and colorectal neoplasias, and can also predispose individuals to adverse effects from drugs with antifolate effects. The MTHFR C677T polymorphism shows no consistent correlation with cardiovascular risk and longevity but, in combination with positive folate balance, the TT genotype is associated with decreased risk of colorectal neoplasias. Because of the high prevalence of this polymorphism in most populations, the TT variant might represent an ancestral genetic adaptation to living constraints (tissue injury or unbalanced vitamin intake) that has become a determinant of disease profiles in modern times. PMID- 11282421 TI - FGF and VEGF function in angiogenesis: signalling pathways, biological responses and therapeutic inhibition. AB - Angiogenic growth factors such as fibroblast growth factors (FGFs) and vascular endothelial growth factors (VEGFs) are currently targets of intense efforts to inhibit deregulated blood vessel formation in diseases such as cancer. FGFs and VEGFs exert their effects via specific binding to cell surface-expressed receptors equipped with tyrosine kinase activity. Activation of the receptor kinase activity allows coupling to downstream signal transduction pathways that regulate proliferation, migration and differentiation of endothelial cells. Inhibitors of FGF and VEGF signalling are currently in clinical trials. In this article, the current knowledge of FGF- and VEGF-induced signal transduction that leads to specific biological responses will be summarized. Furthermore, the manner in which this knowledge is being exploited to regulate angiogenesis will be discussed. PMID- 11282422 TI - The anti-smoking climate in EU countries and Poland. AB - BACKGROUND: In the fall of 1998, 9095 smokers above 18 years, were interviewed about their smoking behaviour and knowledge and attitudes relating to the smoking. The survey (S) was conducted for the Cancer Commission of the EU and sponsored by SmithKline Beecham. An anti-smoking thermometer that is intended to assess the anti-smoking climate (ASC) in each EU country plus Poland was created. In doing so country owners of the S were asked to choose and rank the five questions in the S they thought best reflected the ASC. The five questions chosen were--smoking is a major cause of death and disease, want to stop smoking, governments should do more, ever made a serious quit attempt and smoke free areas should be provided. METHOD: The smokers comprised a representative sample of smoking cigarette per day, vis-a-vis age, sex and rural or urban area. Face to face interviews were conducted using a semi-structured questionnaire. RESULTS: Poland had the most developed ASC, 368 points, followed closely by Sweden 358. In the bottom were Germany 266 and Austria 258. Large differences were noted on willingness to quit; from the 85% in Sweden to Italy 37%. CONCLUSION: The ASC varies considerably within EU and measures to reduce the death and disease from smoking should be tailored to the situation in each country. PMID- 11282423 TI - A novel polymorphism in the promoter of the RAGE gene is associated with non small cell lung cancer. AB - The receptor for advanced glycosylation endproducts (RAGE) is abundant at both the transcriptional and translational level in normal lung but is not expressed in non-small cell lung cancer (NSCLC). In order to determine whether sequence variations might be responsible for the inactivation of RAGE in NSCLC, we investigated the RAGE gene in primary NSCLCs and in the corresponding normal tissues of nine patients. Although sequence analysis revealed no somatic, tumor associated mutations, six novel sequence variants were identified: T-->A in the promoter region 388 bp upstream of the start codon: T-->A in exon 1 (Ala2Ala), C- >G in exon 3 (Val89Val), C-->T in intron 6, G-->C and C-->G in exon 10 (Arg365Ser and Arg369Gly). In addition, we detected a 63 bp deletion in the promoter region (358-421 bp upstream of the start codon) in one NSCLC patient. The T-->A transversion in the promoter region was detected in three of nine patients. Further analysis of this polymorphic locus in 54 NSCLC patients and 59 non-cancer controls revealed a significant difference in the genotype distribution between NSCLC patients and controls. Interestingly, the AA genotype was more common in NSCLC patients (20.8%) than in controls (3.5%). The cumulative occurrence of the AA variant in NSCLC suggests that this genotype is a putative risk factor for NSCLC development. PMID- 11282424 TI - The clinical value of lung imaging fluorescence endoscopy for detecting synchronous lung cancer. AB - Some patients with non-small cell lung cancer (NSCLC) will also have a synchronous malignant lesion. The lung imaging fluorescence endoscopy (LIFE) has proven better than conventional white light bronchoscopy (WLB) for visualizing premalignant lesions or early stages of lung cancer. In this study, the additional value of LIFE in diagnosing synchronous lung cancers as well as the impact of these findings on definite therapy was analyzed. Seventy-two patients with recently diagnosed NSCLC or pulmonary lesions highly suspect of lung cancer were studied. Patients underwent WLB, followed by LIFE. Apart from the primary lesions, additional abnormal and suspicious lesions seen at WLB and LIFE were scored separately and biopsied. Sixty-nine patients had NSCLC and three patients had small cell lung carcinoma. Apart from the primary lesion, one up to six additional endobronchial lesions were visualized in 48 patients by WLB and/or LIFE. High-grade dysplastic lesions were detected in ten patients, three of whom were eligible for surgery of the primary tumor after completion of the investigations. Three other patients (4.3%) had synchronous cancers (NSCLC). In one patient, the lesion was visualized by LIFE and by WLB. The other two malignant lesions were detected only by LIFE. In these three latter patients, diagnostic work-up and definite treatment was changed, as a result of detection of synchronous lesions. In conclusion, LIFE has additional value in detecting synchronous malignant lesions in patients with primary lung cancer. The detection of these lesions changed diagnostic work-up and definite treatment plan. Therefore, LIFE should be used in the work-up of patients with primary lung cancer. PMID- 11282425 TI - Fluorescence bronchoscopy in the detection of preinvasive bronchial lesions in patients with sputum cytology suspicious or positive for malignancy. AB - BACKGROUND: A new strategy in the treatment of squamous cell carcinoma of the tracheobronchial tree is the detection and eradication of preinvasive bronchial lesions before they become invasive cancers. It is, however, difficult to detect preinvasive lesions by conventional white-light bronchoscopy alone. PURPOSE: we conducted a detailed investigation on the use of fluorescence bronchoscopy in the detection of preinvasive bronchial lesions in patients with sputum cytology suspicious or positive for malignancy. METHODS: 64 participants with sputum cytology suspicious or positive for malignancy were examined with both white light and fluorescence bronchoscopy (LIFE group). Earlier to this study, before fluorescence bronchoscopy became available in our institute, 48 participants having sputum cytology suspicious or positive for malignancy were examined with white light bronchoscopy alone (control group). Biopsy specimens for pathological examinations were taken of all abnormal areas discovered by white light or fluorescence bronchoscopy examination. RESULTS: In sputum cytology suspicious or positive for malignancy, the diagnosis of preinvasive bronchial lesions was greatly enhanced in the LIFE group as compared with the control group (45 vs. 7 lesions). The percentage of participants with preinvasive bronchial lesions was also significantly higher in the LIFE group than in the control group (40.6 vs. 12.5%, P = 0.00087, respectively). CONCLUSIONS: Our study suggests that the use of fluorescence bronchoscopy in addition to conventional white-light examination could greatly enhance the detection and localization of preinvasive bronchial lesions in patients with sputum cytology suspicious or positive for malignancy. PMID- 11282427 TI - Unfavorable prognosis of patients with non-small cell lung carcinoma associated with HLA-A2. AB - BACKGROUND: HLA class I molecules present antigenic peptides to cytotoxic T lymphocytes and, thus, play an important role in immune surveillance. Since 1970s there have been many reports of an increased frequency of one or more HLA haplotype in association with autoimmune disease, and malignancy. We studied types of HLA class I antigens in 204 resected non-small cell lung carcinoma (NSCLC) patients and also examined its correlation with clinicopathologic features and prognosis. METHOD: Serological typing for HLA class I antigens was performed using a microcytotoxicity test. The disease-free survival curves were calculated by the Kaplan-Meier method and then compared using the Logrank test. Multivariate analysis was carried out by Cox's proportional hazard method. RESULTS: The difference in disease-free survival time between the HLA-A2 present group and A2 absent group was significant (P = 0.040). The 3-year disease-free survival rate of all patients was 44% in HLA-A2 present group and 66% in A2 absent group. When a comparison was made within the group with stage I, expression of HLA-A2 was the only independent factor that affected survival time by multivariate analysis (P = 0.0457). CONCLUSIONS: Expression of HLA-A2 was considered as one of the unfavorable prognostic factors in NSCLC patients. Our results suggested expression of HLA-A2 in NSCLC patients was one of the mechanisms of escape from immune surveillance. PMID- 11282426 TI - Mechanisms of G1 checkpoint loss in resected early stage non-small cell lung cancer. AB - Loss of the G1 checkpoint appears to be extremely common among virtually all neoplasms. A variety of genetic and epigenetic mechanisms have been demonstrated to play significant roles in this process. In a consecutive series of early stage non-small cell lung cancer (NSCLC), we have established the loss of expression of the G1 Cdk inhibitors p15INK4b) and p16INK4a by DNA methylation is very common (37%), and methylation of p16INK4a is tightly correlated with loss of expression of p16INK4a protein (P = 0.0018). Furthermore, methylation of p15INK4b and p16INK4a appear inversely correlated, although methylation of p15INK4b is an infrequent event in this cohort (4%). Methylation was detected in all stages of NSCLC equally, and did not correlate with survival in these patients. Evidence for methylation was more frequent in squamous cell carcinomas in comparison to other tumor histologies (P = 0.0156). In addition, over-expression of cyclin D1 was found to be tightly restricted (P = 0.0032) to those tumors that had retained wild-type expression of pRB, and did not correlate with methylation or expression of p16INK4a gene product. Although loss of p16INK4a function remains tightly correlated with pRB expression, loss of other regulatory elements in NSCLC such as p53 mutation and cyclin D1 over-expression appear independent of loss of the p16INK4a gene product. PMID- 11282428 TI - The prognostic significance of a previous malignancy in operable non-small cell lung cancer. AB - There is little data in literature on survival of patients with lung cancer as a second primary (SP) malignancy. This retrospective study was undertaken to investigate whether a previous malignancy has prognostic significance in operable non-small cell lung cancer (NSCLC). Sixty-six patients with SP NSCLC were compared with 75 'first primary' (FP) NSCLC patients without a previous malignancy. All the 141 patients had been surgically treated with curative intent at The Netherlands Cancer Institute (NKI) between 1977 and 1996. Patients who had undergone resections for lung metastases were excluded. Clinical and pathological characteristics were collected and a multivariate analysis on all the 141 patients was carried out. All the previous malignancies were invasive cancers associated with metastatic potential and predominantly located in the aerodigestive tract. Female-male ratio was higher in the SP group (29 vs. 15%, P = 0.06). Tumour diameter was smaller in the SP group (3.0 vs. 4.7 cm, P < 0.0001). Pneumonectomy was performed less frequently in the SP group. Five-year survival rate was higher in the SP group (61 vs. 34%, P = 0.04). Univariate favourable prognostic factors were; small tumour diameter, female gender, favourable pTNM-stage, favourable pT-stage, favourable cTNM-stage, no post operative radiotherapy and a history of previous malignancy. Multivariate analysis showed tumour diameter, female gender and pTNM-stage to be the major potential confounders. When adjustments were made for these three variables, the prognostic advantage of the SP group disappeared. It was concluded that SP NSCLC has a similar prognosis when compared with FP NSCLC. NSCLC diagnosed during the follow-up of a previous malignancy, and deemed operable, therefore, warrants the same diagnostic and therapeutic approach as NSCLC as first malignancy. PMID- 11282429 TI - Prognosis of resected non-small cell lung cancer patients with carcinomatous pleuritis of minimal disease. AB - OBJECTIVE: The purpose of this study was to clarify the prognosis of resected non small cell lung cancer (NSCLC) patients with carcinomatous pleuritis of minimal disease which might be considered as the next advanced stage of positive pleural lavage cytology. METHOD: The data were collected from a questionnaire survey on the survival of the patients with carcinomatous pleuritis found at thoracotomy from 1985 to December 1994 which was conducted by the Japan Clinical Oncology Group (JCOG). RESULTS: Out of 227 patients with carcinomatous pleuritis found at thoracotomy who had available information on a survival, 100 patients who underwent a resection of the primary tumor had carcinomatous pleuritis of minimal disease defined based on the criteria of the Japan Lung Cancer Society. The mean malignant fluid volume (+/-S.E.) was 37.1 (6.3) ml and the mean number of pleural disseminated nodules was 5.6 (0.9). A lobectomy was performed in 79 patients, a pneumonectomy in 11 and a limited resection in ten. The 3- and 5-year survival rates were 31.8 and 22.8%, respectively. CONCLUSIONS: The prognosis of resected NSCLC patients with carcinomatous pleuritis of minimal disease was unexpectedly good. This indicates that no fine line may exist between positive pleural lavage cytology findings and the aforementioned lesion. PMID- 11282430 TI - Complementary roles of pro-gastrin-releasing peptide (ProGRP) and neuron specific enolase (NSE) in diagnosis and prognosis of small-cell lung cancer (SCLC). AB - In this study, we evaluated the clinical usefulness of ProGRP and NSE for diagnosis and prognosis of small-cell lung cancer (SCLC). Serum levels of ProGRP and NSE were determined in 108 healthy subjects, 103 patients with benign pulmonary diseases, 142 with non-small cell lung cancer (NSCLC), and 114 with SCLC. Sensitivity of ProGRP in diagnosis of SCLC was significantly higher than that of NSE (64.9 vs. 43.0%, P < 0.001). The difference was substantial in patients with limited disease (56.5 vs. 20.3%, P < 0.001). However, 11 of 40 SCLC patients with normal levels of serum ProGRP (27.5%) showed elevated levels of serum NSE. In the SCLC patients receiving chemotherapy, the CR rate in patients with elevated NSE levels was significantly lower than in patients with normal levels of NSE (18.5 vs. 61.7%, P < 0.001). Elevation of both ProGRP and NSE was a poor prognostic factor, and patients with elevated levels of either ProGRP or NSE showed shorter survival than those without. From multivariate analysis, NSE was found to have a greater effect on survival of SCLC patients than ProGRP. These findings indicate that ProGRP is more sensitive than NSE for diagnosis of SCLC, while NSE is superior to ProGRP as a prognostic factor. In conclusion, both ProGRP and NSE are useful tumor markers and they have a complementary role for each other in diagnosis and prognosis of SCLC. PMID- 11282431 TI - The curative treatment by radiation therapy alone of Stage I non-small cell lung cancer in a geriatric population. AB - This review was initiated to assess the outcome of treatment with radical radiation therapy with curative intent for elderly patients diagnosed to have Stage I non-small cell lung cancer (NSCLC). The study involved a retrospective review of 347 patients with T1 and T2N0M0 tumours treated at the Queensland Radium Institute (QRI) during the period 1985-1992. The main reasons for not proceeding to surgery included poor performance status, old age or refusal to submit to surgery. The median age for the group was 70 years with the range being 34-90 years. Patients in this group were all treated by a standard technique involving external beam radiation therapy to a dose of 50 Gray Minimum Tumour Dose in 20 fractions over 4 weeks. When the study group was divided into those patients aged < 70 years and those patients age > or = 70 years, the overall survival at 5 years was 22 and 34%, respectively (median survival 22 months for age < 70 years, and 26 months for age > or = 70 years). The same division in terms of recurrence free survival yielded 5-year survival rates of 18% for the age < 70 years group and 30% for the age > or = 70 years group with the median survival being 17 months in both sub-groups. Both the difference in overall survival and recurrence free survival between the two age groups approached, but did not reach, statistical significance at the P < 0.05 level of significance. Further sub-division into 5-year age groups failed to confirm the hypothesis that older age groups had a poorer outcome. The 75-79-year group showed better survival than other age groups with the 5-year overall survival for this group being 53%, while the 5-year recurrence free survival was 45%. We conclude from this large series of Stage I NSCLC that radical radiation therapy with curative intent may be a viable alternative to surgery in those elderly patients who either refuse surgery or are judged to be unfit for operation. In view of the fact that comparable results can be achieved in elderly lung cancer patients at the price of minimal toxicity, a nihilist approach to treatment in the elderly can no longer be justified. PMID- 11282432 TI - High-dose radiation therapy for elderly patients with inoperable or unresectable non-small cell lung cancer. AB - PURPOSE: To evaluate definitive radiation therapy delivering doses in excess of 60 Gy for elderly patients aged 75 years or over with non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: The treatment results for 97 patients aged 75 years or older (mean age 78 years; elderly group) with inoperable or unresectable NSCLC were retrospectively analyzed and compared with those for 206 patients younger than 75 year old (mean age 64 years; younger group). The elderly patients were classified into two groups; 67 patients aged 75-79 years (the elderly A) and 30 patients aged 80 years or older (the elderly B). Most of all patients were treated with a total dose of 60 Gy or more in 2 Gy daily standard fractionation. RESULTS: The overall 2 and 5 year survival rates were 32 and 13% for the elderly A group, and 28 and 4% for the elderly B group, respectively, compared with 36 and 12% for the younger group. There was not a statistically significant difference in survival rates among three groups. In stage I-II NSCLC patients there was also no significant difference in survival curves among the three groups. In patients with stage III disease, however, the survival curve of the elderly B was inferior to those of the younger group and the elderly A group, although the difference was not statistically significant. After the treatment the deterioration rate of the performance status was only 5% in the younger group and 8% in the elderly group. Only three younger and two elderly patients died of late pulmonary insufficiency associated with high-dose irradiation to the proximal bronchus. No other treatment-related event was observed except for mild acceptable acute complications in the elderly groups. CONCLUSIONS: Definitive radiation therapy is recommended to the elderly aged 75 years or older with inoperable or unresectable NSCLC, especially early stage disease, as an acceptable choice of treatment. PMID- 11282433 TI - Phase II study of paclitaxel and oral etoposide in patients with locally advanced or metastatic non-small cell lung cancer. AB - The combination of paclitaxel and etoposide was evaluated in a phase II study in patients with locally advanced or metastatic non small-cell lung cancer (NSCLC). Thirty-five patients, median age 61, received treatment with paclitaxel 200 mg/m (2) intravenous over 3 h on day 1, and oral etoposide, 100 mg daily on days 1-5. Cycles were repeated every 21 days for a maximum of nine cycles, or until progression occurred. Twenty-eight patients had stage IV disease, and seven patients had stage IIIA or B disease. There was one complete and seven partial responses (overall response rate, 23%). Two of these responses were in patients with stage III disease (29%) and six in patients with stage IV disease (21%). Median survival was 8.7 months, and 36% of patients were alive at 1 year. There were no treatment-related deaths and little grade 3 or 4 non-haematological toxicity although grade 3 or 4 neutropenia occurred in 60% of patients (33% of cycles). There were four episodes of febrile neutropenia. The combination of paclitaxel and oral etoposide is active in advanced NSCLC and can be delivered with acceptable toxicity. PMID- 11282434 TI - Basaloid large cell lung carcinoma presenting concurrently with metastatic uveal tumor. AB - A 51-year-old man complaining of cough, hemoptysis, and decreased visual acuity was admitted to our hospital. Chest radiography revealed a left hilar mass and pleural effusion in the left hemithorax. In his ophtalmological examination, there was total retinal detachment in the left eye. Ultrasonographic examination and orbital magnetic resonance imaging (MRI) were reported as choroidal metastasis. A computed tomography (CT) confirmed the mass in the left hilum and multiple mass lesions consistent with metastasis in the liver and in the body of 12th thoracic vertebra. Bronchoscopic biopsies revealed large cell carcinoma with basaloid features. He died after 4 months with rapid progression of the disease in spite of combined chemotherapy. Although primary lung cancer with concurrent eye metastasis is an uncommon entity, it should always be kept in mind for patients with ocular symptoms. PMID- 11282435 TI - The effect of aqueous extracts of Cynomorium coccineum and Withania somnifera on testicular development in immature Wistar rats. AB - The effect of lyophilized aqueous extract of Cynomorium coccineum and Withania somnifera on testicular development and on serum levels of testosterone, ICSH and FSH was studied in immature male Wistar rats. There was a notable increase in testicular weight of animals treated with both extracts. Histological examination revealed an apparent increase in the diameter of seminiferous tubules and the number of seminiferous tubular cell layers in the testes of treated rats as compared with control ones. Extracts of both plants elicited notable spermatogenesis in immature rats but C. coccineum was more effective than W. somnifera in that respect. Serum testosterone and FSH levels were lower in animals treated with plants extracts than controls, whereas ICSH levels was higher in treated animals, specially in those treated with C. coccineum. It was concluded that extracts of both plants have a direct spermatogenic influence on the seminiferous tubules of immature rats presumably by exerting a testosterone like effect. PMID- 11282436 TI - Antifertility investigation and toxicological screening of the petroleum ether extract of the leaves of Mentha arvensis L. in male albino mice. AB - In male albino mice, the petroleum ether extract of the leaves of Mentha arvensis L., at the doses 10 and 20 mg/mouse per day for 20, 40 and 60 days, when administered orally, showed a dose and duration dependent reduction in the number of offspring of the treated male mated with normal females. Negative fertility was observed in both dose regimens after 60 days of treatment. The body weight and libido of the treated ammals remain unaffected. However, a significant decrease in the weight of the testis, epididymis, cauda epididymal sperm count, motility, viability and normal morphology of the spermatozoa was observed. The levels of serum protein, bilirubin, GOT, GPT and acid phosphatase, blood urea and haematological indices were unaltered throughout the course of investigation. All the altered parameters were reversible following withdrawal of treatment. The results suggest that the petroleum ether extract of the leaves of M. arvensis possess reversible antifertility property without adverse toxicity in male mice. PMID- 11282437 TI - Protective effect of Cassia occidentalis L. on cyclophosphamide-induced suppression of humoral immunity in mice. AB - Cassia occidentalis L. (Kasaundi) is a widely used medicinal plant. Earlier, we have shown that it possesses antimutagenic activity against benzo[a]pyrene (BaP) and cyclophosphamide (CP)-induced mutagenicity in mice. In this study, we investigated if this plant could also provide protection against CP-induced immunosuppression in animal models. Swiss albino male mice were treated per os with the aqueous extract of C. occidentalis (100 mg/kg, body weight (b.w.)) for 14 days. Cyclophosphamide was given intraperitoneally in a single dose of 50 mg/kg b.w. Body weight, relative organ weight, lymphoid organ cellularity, hemagglutination titre (HT), plaque forming cell (PFC) assay and quantitative hemolysis of SRBC (QHS) were studied in these animals. CP, as expected, showed suppressive effects on lymphoid organ weight and cellularity and other parameters of humoral immunity. Plant extract treatment itself produced no toxicity. The administration of plant extract to CP-exposed animals resulted in improved humoral responses. C. occidentalis treatment significantly (P<0.01) enhanced PFC response in CP-treated animals. In QHS assay, also C. occidentalis showed protection in CP-treated animals. Bone marrow cell counts, which were reduced in CP-treated animals, were reversed significantly (p<0.01) to normal levels in CP+ plant extract group animals. In our earlier study, we found that C. occidentalis modulated hepatic drug metabolizing enzymes. It is suggested that by a similar mechanism, it may be influencing the hematotoxic and immunotoxic responses of cyclophosphamide. PMID- 11282438 TI - The importance of weeds in ethnopharmacology. AB - Tropical primary forest is often considered to be the most important habitat for traditional peoples to gather medicinal plants. However, the role of weeds, commonly found in disturbed areas, in traditional medicinal floras has been overlooked. Data are presented showing the significant representation of weeds in the medicinal floras of the Highland Maya in Chiapas, Mexico and in the medicinal flora of Native North Americans as a whole. The frequency with which weeds appear in these pharmacopoeias is significantly larger (P<0.0001) than what would be predicted by the frequency of weed species in general. Explanations based on human ecology and biochemical ecology are presented. PMID- 11282439 TI - Curcuma longa activates NF-kappaB and promotes adhesion of neutrophils to human umbilical vein endothelial cells. AB - Upregulation of expression of cell adhesion molecules, such as ICAM-1, VCAM-1 and E-selectin, is important for immune surveillance. Extravasation and migration of body's effector cells to the site of immune activation is controlled by the expression of cell adhesion molecules on endothelial cells. We demonstrate here that an aqueous extract prepared from Curcuma longa (ClAqE), a dietary component, promotes the adhesion of peripheral neutrophils to human umbilical vein endothelial cells. To delineate the mechanism of increased adhesion, we investigated the possibility that ClAqE induces the expression of ICAM-1 and E selectin on endothelial cells. ClAqE increases the steady state transcript levels of ICAM-1, VCAM-1, and E-selectin as determined by RT-PCR. We also show that ClAqE activates nuclear transcription factor NF-kappaB, a major transcription factor involved in the transcription of genes encoding ICAM-1, VCAM-1 and E selectin. These results have implications for the usage of aqueous preparation of C. longa for upregulation of cell adhesion molecule expression and/or NF-kappaB. PMID- 11282440 TI - Antihypertensive and vasorelaxant effects of aqueous extract from Croton schiedeanus Schlecht in rats. AB - Antihypertensive and vasorelaxant effects of treatment with the aqueous extract of Croton schiedeanus Schlecht (AECS) were investigated in anaesthetized spontaneously hypertensive rats (SHR). Intravenous bolus injections of AECS (5 100 mg/kg) elicited dose-dependent decreases in mean arterial pressure (MAP) and heart rate (HR). Additionally, AECS (10(-6)-3x10(-3)g/ml) completely relaxed the contractions induced by high K(+) concentrations in intact rat aortic rings in a concentration-dependent manner. PMID- 11282441 TI - Effects of the ethanolic extract of Spirulina maxima on endothelium dependent vasomotor responses of rat aortic rings. AB - Dietary Spirulina decreases, endothelium-dependently, the responses to vasoconstrictor agonists and increases the endothelium-dependent, agonist induced, vasodilator responses of rat aorta rings. The aim of this study was to analyze, in vitro, the effects of a raw ethanolic extract of Spirulina maxima on the vasomotor responses of rat aortic rings to phenylephrine and to carbachol. On rings with endothelium, the extract produced the following effects: (a) a concentration-dependent (60-1000 microg/ml) decrease of the contractile response to phenylephrine; (b) a rightward shift and a decrease in maximal developed tension, of the concentration--response curve to phenylephrine; (c) a concentration dependent relaxation of phenylephrine-precontracted rings. These effects were blocked by L-NAME, and not modified by indomethacin. The extract had no effect on the concentration-response curve to carbachol of rings with endothelium. On endothelium-denuded rings the extract caused a significant rightward shift of the concentration response curve to phenylephrine without any effect on maximal tension development. In the presence of the extract, indomethacin induced a marked decrease in the maximal phenylephrine-induced tension of endothelium-denuded rings. These results suggest that the extract increases the basal synthesis/release of NO by the endothelium and, also, the synthesis/release of a cyclooxygenase-dependent vasoconstricting prostanoid by vascular smooth muscle cells. PMID- 11282442 TI - An investigation on the biological activity of Combretum species. AB - Leaf extracts of 20 Combretum species, many of which are used in southern African traditional medicine, were screened for anti-inflammatory, anthelmintic, anti bilharzia (antischistosomal) and DNA-damaging activity. Significant activity in more than one bioassay was exhibited by Combretum apiculatum, Combretum hereroense, Combretum molle and Combretum mossambicense. Ethyl acetate extracts were generally most active, followed by acetone and then water extracts. PMID- 11282443 TI - Study of two plants used in traditional medicine in Zimbabwe for skin problems and rheumatism: Dioscorea sylvatica and Urginea altissima. AB - Two plants used in Zimbabwean traditional medicine, Dioscorea sylvatica (Dioscoreaceae) and Urginea altissima (Liliaceae), produce mild inflammation and itching when rubbed on the skin. Investigations of the causes of these cutaneous reactions showed that raphides of calcium oxalate are, at least in part, responsible for the effects. Histamine could not be detected. PMID- 11282444 TI - Neutralisation of lethality, myotoxicity and toxic enzymes of Naja kaouthia venom by Mimosa pudica root extracts. AB - Aqueous and alcoholic extracts of dried roots of Mimosa pudica were tested for their inhibitory activity on lethality, myotoxicity and toxic enzymes of Naja kaouthia venom. The aqueous extract, particularly the normal water extract, displayed a significant inhibitory effect on the lethality, myotoxicity and tested enzyme activities of venom compared with alcoholic extracts. The present finding suggests that aqueous extracts of M. pudica root possess compound(s), which inhibit the activity of cobra venom. PMID- 11282445 TI - Dose-dependent decrease in glial fibrillary acidic protein-immunoreactivity in rat cerebellum after lifelong ethanol consumption. AB - The effects of aging and lifelong ethanol consumption on astrocytic morphology and glial fibrillary acidic protein-immunoreactivity (GFAP-IR) in the cerebellar vermis obtained from ethanol-preferring Alko, Alcohol (AA) rats were analyzed by using computer-assisted image analysis. The ethanol-consuming animals (both male and female) were given ethanol (10%-12%, vol./vol.) as the only available fluid for 21 months (3-24 months), whereas the young (3 months) and the old (24 months) controls received water. In the male rats, but not in the female rats, an age related decrease in GFAP-IR was found in folia II, VII, and X of the molecular layer, and in turn, an age-related increase was found in folium X of the granular layer, indicating opposite changes in GFAP-IR for male rats due to aging in adjacent brain regions. In the female rats, 21 months of daily average ethanol consumption of 6.6 g/kg resulted in decreased GFAP-IR in folium VII of the molecular layer, and the decrease in cerebellar GFAP-IR correlated with the average daily ethanol intake (r=-.886, P=.019) when folia II, IV, VII, and X were analyzed together. No effect of ethanol on GFAP-IR was detected in the granular layer or in the central white matter of the female rats. There was no change in GFAP-IR in any of the three cerebellar layers of the male rats with average daily ethanol consumption of 3.2 g/kg. These results indicate that the Bergmann glial fibers are the GFAP-expressing structures of the cerebellum most sensitive to moderate-to-heavy chronic ethanol exposure and that this effect is dose dependent. PMID- 11282446 TI - Heterogeneity in early onset alcoholism suggests a third group of alcoholics. AB - Data from 141 Brazilian male alcoholics were investigated with the objective of further exploring the heterogeneity of alcoholism and to replicate previous studies. A set of seven variables was studied by different cluster analyses to test hypotheses with two, three, and four groups. The results suggested that the best solution showed three groups of alcoholics, two of them similar to those previously described and a third relatively similar to type 2, but with lower scores in harm avoidance, more positive impact of life events, higher proportion of alcoholic relatives, less frequent use of antianxiety drugs, and less delirium tremens. These results reinforced the model with three groups and may be useful in the delineation of new etiology and treatment studies. PMID- 11282447 TI - Effects of alcohol ingestion following exercise on postprandial lipemia. AB - The study determined the effect of alcohol ingestion postexercise on postprandial lipemia during recovery. The mean values were compared with those obtained in a control experiment during which no alcohol was given. Nineteen normolipidemic subjects (11 males and 8 females) performed two exercise trials at an intensity corresponding to 70% VO2max for 35 min. In a random order, alcoholic (0.7 g/kg) or alcohol-free drinks were given 1 h after the completion of exercise. Venous blood samples were obtained pre- (before breakfast) and postexercise and pre- and postprandially during recovery. Total cholesterol and high-density lipoprotein cholesterol showed no change with exercise or alcohol ingestion. In the control trial, when subjects consumed a standardized lunch, triglycerides showed no significant change, but when alcohol was consumed postexercise triglyceride concentration increased substantially 5 h during recovery in both males and females. The mechanism responsible for the rise in triglyceride concentration during recovery when alcohol was ingested following exercise is not known, but this appears to be a late phenomenon. PMID- 11282448 TI - The pharmacodynamics of anticonvulsant and subconvulsant effects of ethanol in CBA and C57BL/6 mice. AB - A method of determination of minimal effective doses (MEDs) of bicuculline causing clonic-tonic convulsions (CTC) and tonic extension (TE) was used to investigate ethanol pharmacodynamics in C57BL/6 and CBA mice, differing in levels of alcohol predisposition. It is observed that ethanol produces a powerful anticonvulsant action antagonizing convulsant effects of bicuculline. On a long term scale, the pharmacological action of alcohol had two phases in both strains of mice: anticonvulsant (in the interval 5 min to 4 h after ethanol administration) and subconvulsant (4-24 h after ethanol administration). C57BL/6 mice were characterized by a more rapid development of the anticonvulsant effect and its faster decay in comparison to CBA strain. A possibility of correct quantitative evaluation of data allows using the method of MED determination as an express model of an acute alcohol abstinence syndrome, as well as for screening of new antialcohol drugs. PMID- 11282449 TI - Serum cytokine levels in alcohol-related liver cirrhosis. AB - Chronic alcoholism complicated by alcoholic liver disease is characterized by activation of the inflammatory response system. To evaluate the role of cytokines in the progress of alcoholic cirrhosis, we assessed serum level of the proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-6, and IL-8 and the antiinflammatory cytokines IL-2, IL-10, and transforming growth factor (TGF)-beta in patients with compensated and decompensated alcoholic liver cirrhosis. Compensated alcoholic cirrhosis was characterized by increased IL-6 (6.3+/-2.9 vs. HP 2.2+/-1.4 pg/ml in controls) and decreased IL-10 (HP 4.1+/ 3.5 vs. 6.4+/-5.4 pg/ml in controls). TNF-alpha, IL-8, and TGF-beta1 levels were comparable to those found in controls. In sera of patients with decompensated alcoholic liver cirrhosis, besides increased IL-6 (11.2+/-7.7 pg/ml), increased concentrations of TNF-alpha (25.1+/-4.5 vs. 9.1+/-7.0 pg/ml in controls) and IL-8 (171.7+/-294.0 vs. 2.7+/-2.9 pg/ml in controls) were also detected. TGF-beta1 and IL-10 levels were similar to those found in controls. These results strongly indicate that a significant derangement of the balance between proinflammatory and antiinflammatory signals is characteristic of compensated and especially of decompensated alcoholic cirrhosis. PMID- 11282451 TI - Injected muscimol in pedunculopontine tegmental nucleus alters ethanol self administration. AB - The pedunculopontine tegmental nucleus (PPN) has been implicated in a variety of behavioral functions, including stimulus selection. Given the PPN interactions with the mesolimbic system, it was considered important to determine its involvement in ethanol self-administration. Long-Evans male rats were trained to self-administer 10% ethanol by using a sucrose-substitution procedure. After implantation of cannula guides, microinjections of 30, 100, and 300 ng of muscimol into the PPN before the self-administration session were performed. Ethanol self-administration was decreased at the 300-ng dose, in a manner similar to the actions of dopamine agonists microinjected in the nucleus accumbens. It is hypothesized that loss of PPN cholinergic input to the mesolimbic system affects the integrative activity of the nucleus accumbens and underlies the observed change in ethanol self-administration behavior. PMID- 11282450 TI - Acetaldehyde in vitro exposure and apoptosis: a possible mechanism of teratogenesis. AB - Alcohol abuse by pregnant women can result in fetal alcohol effects (FAE) and fetal alcohol syndrome (FAS). Both ethanol itself and its main metabolite, acetaldehyde (Ach), are able to produce specific FAS-related malformations. In previous in vitro studies, we documented that 10-day-old rat embryos exposed to Ach show a characteristic embryonic Ach syndrome, histologically characterized by marked cellular death. As both necrosis and pathological apoptosis are teratological mechanisms, the aim of this work was to evaluate if cellular death, observed in Ach-exposed embryos, can be related to necrotic or apoptotic events. Ten-day-old rat embryos were cultured in the presence of Ach 30-60 microg/ml and stained with the vital dye acridine orange to visualize apoptotic areas. After fixation, the TUNEL [3' terminal deoxynucleotide transferase (TdT)-mediated dUTP biotin nick end labeling] method was used to histologically identify apoptosis. Both acridine orange and TUNEL staining showed signs of physiological apoptosis in controls and abnormal apoptotic regions in Ach-exposed embryos. Our results show a clear correlation between malformed organs and apoptotic embryonic districts, suggesting the role of apoptosis in Ach-induced abnormalities. PMID- 11282452 TI - Regional differences in cell loss associated with binge-like alcohol exposure during the first two trimesters equivalent in the rat. AB - Women who abuse alcohol during pregnancy may deliver offspring who could be diagnosed with fetal alcohol syndrome (FAS) or a less severe deficit involving cognitive and behavioral disorders. The severity of the deficits may involve the interaction of several known risk factors, such as alcohol consumption pattern or duration, the timing of alcohol consumption relative to critical windows of vulnerability, or the inherent differential vulnerability among the various brain regions to alcohol-induced brain injury. In this study, we explore the vulnerability of the different brain regions by making cell counts from multiple brain regions. Specifically, we used stereological cell-counting techniques to estimate the total cell numbers in the cerebellum (Purkinje and granule cells), olfactory bulb (mitral and granule cells), hippocampus (CA1 and CA3 cells), and dentate gyrus (granule cells). Groups of timed-pregnant Sprague-Dawley rats were assigned to one of five treatments: alcohol by intragastric intubation (2.25, 4.5, or 6.5 g/kg/day), nutritional control [pairfed and intubated=Pairfed) and intubated], and normal control (Chow). Treatments began on embryonic day 1 (E1) and continued through E20. On E33 (usually postnatal day 10), all offspring were perfused intracardially with saline followed by fixatives. Representative forebrains, cerebella, and olfactory bulb from each group were processed for cell counting. The optical dissector was used to obtain cell densities, while Cavalieri's principle was used to calculate the reference volume. The product of density and volume gave unbiased estimates of the total neuronal number within each brain region. Overall peak BACs (regardless of sampling day) for the three alcohol groups averaged 136, 290, and 422 mg/dl for the 2.25-, 4.5-, and 6.5-g/kg groups, respectively. The total number of cerebellar Purkinje cells was reduced in the 6.5-g/kg group relative to controls, while the total number of olfactory bulb mitral cells and hippocampal CA1 and CA3 pyramidal cells from all alcohol treated groups was not different from controls. Total numbers of granule neurons were reduced in the cerebellum and olfactory bulb of offspring exposed to 4.5 or 6.5 g/kg/day, but granule cell numbers in the dentate gyrus were not affected by the prenatal alcohol treatment. Taken together with previous findings, these data demonstrate that prenatal alcohol exposure results in regional vulnerability of various brain structures and underscores the variability of deleterious effects of alcohol on brain development. PMID- 11282453 TI - Assessing the omnipotence of inositol hexakisphosphate. AB - This review assesses the authenticity of inositol hexakisphosphate (InsP(6)) being a wide-ranging regulator of many important cellular functions. Against a background in which the possible importance of localized InsP(6) metabolism is discussed, there is the facile explanation that InsP(6) is merely an "inactive" precursor for the diphosphorylated inositol phosphates. Indeed, many of the proposed cellular functions of InsP(6) cannot sustain a challenge from the implementation of a rigorous set of criteria, which are designed to avoid experimental artefacts. PMID- 11282454 TI - Identification of substrate specificity determinants in human cAMP-specific phosphodiesterase 4A by single-point mutagenesis. AB - To identify amino acids that might be involved in discriminating guanosine-3',5' cyclic phosphate (cGMP) towards adenosine-3',5'-cyclic phosphate (cAMP) binding in the cAMP-specific phosphodiesterases, alignments of different human cyclic nucleotide phosphodiesterases (PDEs) were performed. Eight amino acid residues that are highly conserved in the cAMP-hydrolysing phosphodiesterases (PDE1, PDE3, PDE4, PDE7, PDE8) and that did not show any homologies to the cGMP-specific phosphodiesterases (PDE5, PDE6, PDE9) were selected from these alignments. Using the technique of site-directed mutagenesis, derivatives of PDE4A carrying single mutations at these conserved residues (amino acid positions are given according to the human PDE4A isoform HSPDE4A4B; accession number L20965) were generated and expressed in COS1 cells. The expression products were characterised with regard to cAMP and cGMP hydrolysis and sensitivity towards type-specific inhibitors. The mutation of Phe484 toward Tyr, Ala590 toward Cys, Leu391 and Val501 towards Ala had no significant influence on substrate affinity or specificity. However, the exchange of Trp375 and Trp605 for aliphatic residues abolished catalytic activity and the exchange of Pro595 for Ile led to sevenfold decrease of substrate affinity and an 14-fold decrease of the affinity towards the PDE4-specific inhibitor 4-[3-(cyclopentoxyl)-4-methoxyphenyl]-2-pyrrolidone (rolipram). Both effects may provide evidence for a structural importance of Trp375, Trp605 and Pro595 for PDE function. By exchanging the aspartate residue for asparagine or alanine at position 440 of the human PDE4A4B isoform, the substrate specificity was altered from the highly specific cAMP hydrolysis to an equally efficient cAMP and cGMP binding and hydrolysis. In addition, the IC(50) values for common PDE4 specific inhibitors like rolipram, N-(3,5-dichlorpyrid-4-yl)-3-cyclopentyl-oxy-4 methoxy-benzamide (RPR-73401) and 8-methoxy-5-N-propyl-3-methyl-1-ethyl imidazo[1,5-a]-pyrido[3,2-e]-pyrazinone (D-22888) were dramatically increased. These results demonstrate an important role of the aspartate at position 440 in determining substrate specificity and inhibitor susceptibility of PDE4A. The strong conservation of this residue suggests that Asp440 may play a similar role in other cAMP-PDEs. PMID- 11282455 TI - Glucocorticoid-induced insulin resistance associates with activation of protein kinase C isoforms. AB - We studied glucocorticoid-induced insulin resistance and possible role of protein kinase C (PKC). Pretreatment with dexamethasone, prednisolone and corticosterone for 60 min decreased insulin-induced [3H] 2-deoxyglucose (DOG) uptake in isolated rat adipocytes. Preincubation with Go6976, LY379196 or myristoylated PKC pseudosubstrate, conventional PKC inhibitor, but not cycloheximide or RU38486, recovered dexamethasone-induced insulin resistance. Dexamethasone activated immunoprecipitates with anti-PKC alpha, beta, and zeta antibodies. PKC zeta activity in adipocytes increased to 163%, and 264% from basal level (100%) with dexamethasone and insulin treatment, respectively. Dexamethasone provoked redistribution of both PKC beta and zeta from the cytosol to the membrane. These results indicate that dexamethasone activates both conventional and atypical PKC. However, conventional PKC is more important in glucocorticoid-induced insulin resistance. PMID- 11282456 TI - HCO(3)(-) ions modify the role of PKC isoforms in the modulation of rat mast cell functions. AB - PKC and the intracellular calcium signal are two well-known intracellular signaling pathways implicated in the induction of mast cell exocytosis. Both signals are modified by the presence or absence of HCO(3)(-) ions in the external medium. In this work, we studied the regulation of the exocytotic process by PKC isozymes and its relationship with HCO(3)(-) ions and PKC modulation of the calcium entry. The calcium entry, induced by thapsigargin and further addition of calcium, was inhibited by PMA, a PKC activator, and enhanced by 500 nM GF109203X, which inhibits Ca(2+)-independent PKC isoforms. PMA inhibition of the Ca(2+) entry was reverted by 500 and 50 nM GF109203X, which inhibit Ca(2+)-independent and Ca(2+)-dependent isoforms, respectively, and Go6976, a specific inhibitor of Ca(2+)-dependent PKCs. Thus, activation of Ca(2+)-dependent and Ca(2+) independent PKC isoforms inhibit Ca(2+) entry in rat mast cells, either in a HCO(3)(-)-buffered or a HCO(3)(-)-free medium. PMA, GF109203X, Go6976 and rottlerin, a specific inhibitor of PKC delta, were also used to study the role of PKC isoforms in the regulation of exocytosis induced by thapsigargin, ionophore A23187 and PMA. The results demonstrate that Ca(2+)-dependent PKC isoforms inhibit exocytosis in a HCO(3)(-)-dependent way. Moreover, Ca(2+)-independent PKC delta was the main isoform implicated in promotion of Ca(2+)-dependent mast cell exocytosis in the presence or absence of HCO(3)(-). The role of PKC isoforms in the regulation of mast cell exocytosis depends on the stimulus and on the presence or absence of HCO(3)(-) ions in the medium, but it is independent of PKC modulation of the Ca(2+) entry. PMID- 11282457 TI - Resveratrol inhibits the formation of phosphatidic acid and diglyceride in chemotactic peptide- or phorbol ester-stimulated human neutrophils. AB - Resveratrol (trans-3,5,4'-trihydroxystilbene, Res) is a naturally occurring antioxidant found in grape berry skins and red wine. It has anti-inflammatory effects. In this study, we examined the effect of Res on the formation of phosphatidic acid (PA) and diglyceride (DG), in human neutrophils stimulated by formyl-methionyl-leucyl-phenylalanine (fMLP) or by phorbol 12-myristate 13 acetate (PMA). We measured the masses of PA and DG by using a nonradioactive method. Our results showed that Res inhibited the formation of PA in a concentration dependent manner with an IC(50) value of 42.4 and 60.9 microM in fMLP- and PMA-stimulated cells, respectively. Res also suppressed the formation of phosphatidylethanol (PEt), thereby implying inhibition of phospholipase D (PLD) activity. In addition, Res inhibited the formation of both diacylglycerol (DAG) and ether-linked acylglycerol (EAG) induced by fMLP and by PMA. Our results suggest that Res inhibition of PLD activity may contribute to its anti inflammatory effects. PMID- 11282458 TI - Protein kinase C-dependent and -independent inhibition of Ca(2+) influx by phorbol ester in rat pancreatic beta-cells. AB - Phorbol esters were used to investigate the action of protein kinase C (PKC) on insulin secretion from pancreatic beta-cells. Application of 80 nM phorbol 12 myristate 13-acetate (PMA), a PKC-activating phorbol ester, had little effect on glucose (15 mM)-induced insulin secretion from intact rat islets. In islets treated with bisindolylmaleimide (BIM), a PKC inhibitor, PMA significantly reduced the glucose-induced insulin secretion. PMA decreased the level of intracellular Ca(2+) concentration ([Ca(2+)](i)) elevated by the glucose stimulation when tested in isolated rat beta-cells. This inhibitory effect of PMA was not prevented by BIM. PMA inhibited glucose-induced action potentials, and this effect was not prevented by BIM. Further, 4alpha-phorbol 12,13-didecanoate (4alpha-PDD), a non-PKC-activating phorbol ester, produced an effect similar to PMA. In the presence of nifedipine, the glucose stimulation produced only depolarization, and PMA applied on top of glucose repolarized the cell. When applied at the resting state, PMA hyperpolarized beta-cells with an increase in the membrane conductance. Recorded under the voltage-clamp condition, PMA reduced the magnitude of Ca(2+) currents through L-type Ca(2+) channels. BIM prevented the PMA inhibition of the Ca(2+) currents. These results suggest that activation of PKC maintains glucose-stimulated insulin secretion in pancreatic beta-cells, defeating its own inhibition of the Ca(2+) influx through L-type Ca(2+) channels. PKC-independent inhibition of electrical excitability by phorbol esters was also demonstrated. PMID- 11282460 TI - A soluble LAT deletion mutant inhibits T-cell activation: reduced recruitment of signalling molecules to glycolipid-enriched microdomains. AB - The type III transmembrane adaptor protein linker for activation of T cells (LAT) is essential for membrane recruitment of signalling molecules following TCR activation. Here we show that although LAT deleted in the transmembrane domain is completely soluble, it can be tyrosine phosphorylated after anti-CD3 stimulation or pervanadate treatment. Overexpression of this deletion mutant in transiently transfected Jurkat TAg cells inhibits transcriptional activation of nuclear factor of activated T cells (NF-AT)/AP-1 reporter construct in a concentration dependent manner. Furthermore, by selection of transiently transfected cells, a clear reduction of TCR-induced CD69 expression was observed in cells expressing the mutant. These dominant negative effects seemed to be dependent both on the ability of the membrane deletion mutant to reduce phosphorylation of endogenous LAT and to reduce interaction of endogenous LAT with PLC-gamma1 and Grb2. Consistent with this, the redistribution of PLC-gamma1 and Grb2 to glycolipid enriched microdomains, called lipid rafts, after stimulation was inhibited when the soluble form of LAT was overexpressed. We suggest that the dominant negative effect is caused by the ability of the mutant to sequester signalling molecules in cytosol and thereby inhibit redistribution of signalling molecules to lipid rafts upon T-cell activation. PMID- 11282459 TI - Activation of muscarinic acetylcholine receptors induces Ca(2+) mobilization in FRT cells. AB - The effect of the muscarinic receptors agonist carbachol (Cch) on intracellular calcium concentration ([Ca(2+)](i)) and cAMP level was studied in polarized Fischer rat thyroid (FRT) epithelial cells. Cch provoked a transient increase in [Ca(2+)](i), followed by a lower sustained phase. Thapsigargin, a specific microsomal Ca(2+)-ATPase inhibitor, caused a rapid rise in [Ca(2+)](i) and subsequent addition of Cch was without effect. Removal of extracellular Ca(2+) reduced the initial transient response and completely abolished the plateau phase. Ryanodine, an agent that depletes intracellular Ca(2+) stores through stimulation of ryanodine receptors (RyRs), had no effect on [Ca(2+)](i). However, the transitory activation of [Ca(2+)](i) was dose-dependently attenuated in cells pretreated with U73122, a specific inhibitor of phospholipase C (PLC). These data suggest that the Cch-stimulated increment of [Ca(2+)](i) required IP(3) formation and binding to its specific receptors in Ca(2+) stores. Further studies were performed to investigate whether the effect of Cch on Ca(2+) entry into FRT cells was via L-type voltage-dependent Ca(2+) channels (L-VDCCs). Nicardipine, a nonspecific L-type Ca(2+) channel blocker, decreased Cch-induced increase on [Ca(2+)](i), while Bay K-8644, an L-type Ca(2+) channel agonist, slightly increased [Ca(2+)](i) in FRT cells. These data indicate that Ca(2+) entry into these nondifferentiated thyroid cells occurs through an L-VDCC, and probably through another mechanism such as a capacitative pathway. Cch did not affect the intracellular cAMP levels, but its effects on [Ca(2+)](i) were significantly reduced when cells were pretreated with forskolin, suggesting the existence of an intracellular cross-talk between PLC and cAMP mechanisms in the regulation of intracellular Ca(2+) mobilization in neoplastic FRT cells. PMID- 11282463 TI - Web alert. Cell regulation. PMID- 11282465 TI - Transcription elongation complex: structure and function. AB - Our understanding of the mechanisms of transcription has been greatly advanced by recent determination of the X-ray structure of bacterial RNA polymerase. Using crosslinking approaches, extensive mapping of DNA and RNA contacts onto this structure allowed tracking of the path of nucleic acids through the transcription elongation complex. The resulting structural model of the transcription elongation complex is linked to the functional one, which is based on numerous data accumulated during previous studies of RNA synthesis. An integrated structure-function model allows the rational explanation of termination and pausing and provides new insights into the mechanisms of transcription. PMID- 11282466 TI - How sigma docks to RNA polymerase and what sigma does. AB - It is clear that multiple sites of interaction exist between sigmas and core subunits, likely reflecting the changing pattern of interactions that occur sequentially during the complex process of holoenzyme formation, open promoter formation, and initiation of transcription. Recent studies have revealed that a major site of interaction of Escherichia coli sigma factors is the amino acid 260 309 coiled-coil region of the beta' subunit of core RNA polymerase. This region of beta' interacts with region 2.1-2.2 of sigma(70). Binding of this region of beta' to sigma(70) triggers a conformational change in sigma that allows it to bind to a -10 nontemplate promoter DNA strand oligonucleotide. PMID- 11282467 TI - The AraC transcriptional activators. AB - The AraC family of bacterial transcriptional activators regulate diverse genetic systems. Recent X-ray diffraction studies show that the monomeric MarA and Rob activators bind to their asymmetric degenerate DNA sites via two different helix turn-helix elements. Activation by MarA, SoxS or Rob requires a particular orientation of the asymmetric binding sequence (and hence the activator), depending on its distance from the -10 RNAP signal. Genetic studies are beginning to clarify how the activators interact with RNAP. Growing evidence suggests that for the sugar metabolism activators, multiple binding sites upstream of the promoter anchor the activator in a repressing or nonactivating configuration. By interaction with the sugar and/or CRP, the activator is allosterically altered so it can bind a new set of sites that enable it to activate the promoter. Surprisingly, the virulence activator, Rns, must bind to both upstream and downstream sites in order to activate the rns promoter. PMID- 11282468 TI - Transcriptional regulation at a distance in bacteria. AB - Transcriptional enhancers are cis-acting DNA elements that are binding sites for regulatory proteins and function at large distances from promoter elements to stimulate transcription. Once thought to be unique to eukaryotes, enhancer-like elements have been discovered in a wide variety of bacteria. The regulatory proteins that bind to these bacterial enhancers must contact RNA polymerase to activate transcription. In principle, interactions between bacterial enhancer binding proteins and RNA polymerase can occur by either DNA looping or tracking of the enhancer-binding protein along the DNA. Paradigms for each of these methods are found in bacterial systems. Activators of sigma(54)-RNA polymerase holoenzyme contact polymerase by DNA looping, while bacteriophage T4 gp45 functions as a sliding clamp that tracks along DNA until it engages RNA polymerase. Significant advances have been made over the last few years towards understanding the mechanisms by which bacterial enhancer-binding proteins activate transcription, but important aspects of these mechanisms are still poorly defined. PMID- 11282470 TI - Control of transcription by nucleoid proteins. AB - Nucleoid proteins are a group of abundant DNA binding proteins that modulate the structure of the bacterial chromosome. They have been recruited as specific negative and positive regulators of gene transcription and their fluctuating patterns of expression are often exploited to impart an additional level of control with respect to environmental conditions. PMID- 11282471 TI - Revisiting the stringent response, ppGpp and starvation signaling. AB - Microbial adaptation to environmental stress plays an important role in survival. It is necessary to understand the mechanisms underlying the survival of microbes under stress, as they may eventually aid in the successful control of the growth and persistence of these organisms. During nutrient starvation, Escherichia coli elicits a stringent response to conserve energy. The hallmark of the stringent response is the accumulation of guanosine tetra- (ppGpp) and pentaphosphates (pppGpp), which probably bind RNA polymerase to regulate gene expression at certain promoters. Recently, there has been renewed interest in the stringent responses of other microbes, with a view to correlating it with sporulation, virulence and long-term persistence. PMID- 11282469 TI - Mechanisms of transcriptional repression. AB - Transcriptional repressors are usually viewed as proteins that bind to promoters in a way that impedes subsequent binding of RNA polymerase. Although this repression mechanism is found at several promoters, there is a growing list of repressors that inhibit transcription initiation in other ways. For example, several repressors allow the simultaneous binding of RNA polymerase to the promoter, but interfere with subsequent events of the initiation process, eventually inhibiting transcription initiation. The recent increase in the number of repressors for which the repression mechanism has been characterized in detail has shown an amazing variety of strategies to repress transcription initiation. It is not surprising to find that the repression mechanism used is usually exquisitely adapted to the characteristics of the promoter and of the repressor involved. PMID- 11282472 TI - Responses of Gram-negative bacteria to certain environmental stressors. AB - Bacteria in nature are exposed to variations in temperature, and are affected by the availability of nutrients and water and the presence of toxic molecules. Their reactions to these changes require a series of rapid adaptive responses. Although transcriptional regulation is of primary importance in these responses, translational regulation and even activation of 'silenced' enzymes are critical for survival in changing environments. Bacteria have developed a series of mechanisms at the membrane structure level to cope with high concentrations of solvents. In addition, solvent-tolerant strains express highly effective efflux pumps to remove solvents from the cytoplasm. Desiccation tolerance is based on the synthesis and accumulation of osmoprotectants together with changes in fatty acid composition to preserve membrane structure. Both cold shock and heat shock responses are mainly regulated at a post-transcriptional level, translation efficiency in the case of cold shock and mRNA half-life and sigma32 stability in the case of heat shock. PMID- 11282473 TI - Iron and metal regulation in bacteria. AB - In Escherichia coli, the iron regulator Fur is regulated by two oxidative-stress response regulators. The generation of dangerous radicals by oxygen and iron is the basis for this dual regulation, which is also found in eukaryotes. The binding of iron-regulated transcripts to apo-aconitase and results of mRNA half life studies indicate that there is post-transcriptional iron regulation in bacteria, as in eukaryotes. Fur contains two metal-binding sites, one for Zn2+ and one for Fe2+. Zinc uptake systems are regulated by members of the Fur protein family, and zinc is a cofactor. DtxR and related proteins constitute another family of iron regulators, first found in Gram-positive organisms with a high GC content. In organisms with Fur-dependent iron regulation, members of the DtxR family regulate manganese transport. PMID- 11282474 TI - Transcriptional responses to DNA damage. AB - In Escherichia coli, DNA repair and protective responses are regulated at the transcriptional level. Regulatory mechanisms have evolved that allow cells to respond to DNA damage by mounting the appropriate responses. The regulatory proteins controlling these responses are activated when they recognize the presence of a specific DNA damaging agent, the production of specific DNA lesions, or the production of damage intermediates resulting from replication of lesions containing DNA. Transcription of the responses to DNA damage are induced when the activated regulatory proteins stimulate transcription of the genes they control by a variety of complex and unique molecular mechanisms. PMID- 11282475 TI - Quorum sensing as an integral component of gene regulatory networks in Gram negative bacteria. AB - Bacterial cell-to-cell communication (quorum sensing) relies upon the interaction of a small diffusible signal molecule with a sensor or transcriptional activator to couple gene expression with cell population density. In Gram-negative bacteria, it is now clear that N-acylhomoserine lactones bind directly to LuxR homologues and can be synthesized via one of three unrelated bacterial protein families and by transgenic plants. New chemical classes of signal molecules have been identified, some of which exhibit crosstalk with N-acylhomoserine-lactone mediated quorum sensing. As the determinant of cell population density, quorum sensing is emerging as an integral component of bacterial global gene regulatory networks responsible for facilitating bacterial adaptation to environmental stress. N-acylhomoserine lactones are produced during experimental animal and human infections, and a function beyond quorum sensing has been suggested by their intrinsic immunomodulatory and pharmacological activities. PMID- 11282476 TI - Regulatory circuits for plasmid survival. AB - Bacterial plasmids deploy a diverse range of regulatory mechanisms to control expression of the functions they need to survive in the host population. Understanding of the mechanisms by which autoregulatory circuits control plasmid survival functions, in particular plasmid replication, has been advanced by recent studies. At a molecular level, structural understanding of how certain antisense RNAs control replication and stability functions is almost complete. Control circuits linking plasmid transfer functions to the status of the bacterial population have been dissected, uncovering a complex and hierarchical organisation. Coordinate or global regulation of plasmid replication, transfer and stable maintenance functions is becoming apparent across a range of plasmid families. PMID- 11282477 TI - Bacteriophage lambda: alive and well and still doing its thing. AB - The lambda (lambda) family of bacteriophages continues to provide significant insights into the understanding of basic biological processes, as well as useful technological innovations. Areas in which recent advances have occurred include transcription elongation, repressor interactions, genomics and post transcriptional regulation. The homologous lambda recombination functions have been exploited as an efficient in vivo recombinant engineering system for functional genomic studies. The virulence of some pathogenic strains of Escherichia coli is enhanced by the expression of Shiga toxin (stx) genes encoded on a resident lambdoid prophage. Recent work suggests that the phage regulatory network may be a significant contributor to toxin production and release by these pathogenic E. coli. PMID- 11282478 TI - Mechanism and regulation of transcription in archaea. AB - The archaeal basal transcription machinery resembles the core components of the eucaryal RNA polymerase II apparatus. Thus, studies of the archaeal basal machinery over the last few years have shed light on fundamentally conserved aspects of the mechanisms of transcription pre-initiation complex assembly in both eucarya and archaea. Intriguingly, it has become increasingly apparent that regulators of archaeal transcription resemble regulators initially identified in bacteria. The presence of these shared bacterial-archaeal regulators has given insight into the evolution of gene regulatory processes in all three domains of life. PMID- 11282479 TI - Dimorphism in fungal pathogens: Candida albicans and Ustilago maydis--similar inputs, different outputs. AB - The ability to switch between a yeast-like form and a filamentous form is an extended characteristic among several fungi. In pathogenic fungi, this capacity has been correlated with virulence because along the infection process, dimorphic transitions are often required. Two well-known organisms for which dimorphism have been studied are the pathogenic fungi Candida albicans and Ustilago maydis, which infect mammals and corn, respectively. In both cases, several signal transduction pathways have been defined. Not surprisingly, these pathways are similar to the well-known pathways involved in the pseudohyphal differentiation that some Saccharomyces cerevisiae diploid strains show when nutrients are starved. However, in spite of similarities at the molecular level, strikingly, fungi use similar pathways to respond to environmental inputs, but with differing outcomes. PMID- 11282481 TI - Treatment of subacute hepatitis with Lamivudine and intravenous Glycyrrhizin: a pilot study. AB - Subacute hepatitis is a common and distinct clinicopathological entity due to Hepatitis B and E viruses in India. Lamivudine has been established as a safe and effective antiviral agent for the treatment of chronic HBV hepatitis. This drug was administered orally along with intravenous (I/V) Glycyrrhizin, an immunomodulator drug, in an open pilot trial to assess its efficacy in the treatment of subacute hepatitis. The results establish the safety and efficacy of Lamivudine in combination with I.V. Glycyrrhizin in subacute Hepatitis. PMID- 11282482 TI - Clinical usefulness of the branched DNA assay version 2 in predicting the efficacy of Interferon treatment in a group of chronic HCV patients based on serotype. AB - Interferon (IFN) response depends upon pretreatment viral loads and viral genotype/serotype. This study investigated the difference in response to IFN in different viral load groups of 96 chronic hepatitis C patients and compared version 1 (bDNA1.0) and version 2 (bDNA2.0) of the branched DNA assay, according to serotype. On univariate analysis, viral load (P=0.0001, by bDNA 1.0; P=0.0020, by bDNA 2.0,), serotype (P=0.0053) and age (P=0.0073) were significant predictors of IFN response. On multivariate analysis, serotype (odds ratio, 5.44; 95% confidence interval, 1.94-15.24; P<0.01) was a stronger predictor of IFN response than age or viral load (by bDNA2.0). In very high (>6.7 log eq/ml), high (6.0 approximately 6.69 log eq/ml) and low (<6 log eq/ml) viral load groups (by bDNA2.0), complete response was 25, 55 and 92.6% in serotype 2 (sero-2), and 10, 20 and 71.4% in serotype 1 (sero-1), respectively. In sero-2, bDNA2.0 can detect high viral loads underestimated by bDNA1.0. In a low viral load group (by bDNA2.0), IFN response is dependent upon serotype; sero-2 responded better than sero-1. However, in high and very high viral load groups, sensitivities of bDNA1.0 and bDNA2.0 are not effective in clinically distinguishing CR from non response, and aid in patient selection for IFN therapy. PMID- 11282483 TI - Primary biliary cirrhosis with antibody against carbonic anhydrase II associates with distinct immunological backgrounds. AB - Objective: a part of patients with primary biliary cirrhosis (PBC) has anti-human carbonic anhydrase II (CA II) autoantibodies, although several contradictional reports followed. Since immunization of mice with CA II results in cholangitis in a susceptible strain, PBC with anti-CA II antibody may have distinct clinical features. Thus, we tested the sera of patients with PBC for anti-CA II antibodies and compared clinical characteristics of PBC patients with and without anti-CA II antibodies in Japanese patients. Methods: anti-CA II antibodies were detected in nine of 50 (18%) PBC patients by immunoblotting. The evaluation of these patients included various clinical parameters, autoantibodies, and immunological backgrounds. Results: the levels of serum liver tests and the prevalence of serum anti-mitochondrial antibody (77.8 vs. 92.7%) were not different between the patients with and without anti-CA II antibody. However, the prevalence of anti nuclear antibody (ANA) was significantly higher in the patients with anti-CA II antibody than that in the patients without anti-CA II antibody (66.7 vs. 25.6%, P=0.044), although their mean titers were not statistically different. Association of Sjogren's syndrome tended to be more frequent in the patients with anti-CA II antibody than those without it (33.3 vs. 14.6%, P=0.327). Studies of HLA class I allotype revealed that three of five (60.0%) patients with anti-CA II antibodies and one patients from 34 (3.0%) patients without anti-CA II antibodies had HLA B51 allotype; the difference in the prevalence of this allotype was significant (P=0.004, Pc=0.01), and the prevalence of other HLA class I and HLA DR allotypes was similar between the patients with and those without anti-CA II antibody. Administration of ursodeoxycholic acid (600 mg per day) was accompanied by change in liver tests in a similar way between the two patient groups. Conclusions: These results suggest that, although clinical features are not distinctive, PBC patients with anti-CA II antibody may have a genetic background, which may contribute to a susceptibility to immune-mediated cholangitis. PMID- 11282484 TI - Increased plasma homocysteine in liver cirrhosis. AB - Background: Homocysteine (Hcy), is an atherogenic and thrombogenic risk factor which has also been proposed to be involved in hepatic fibrinogenesis. Hcy metabolism, depends on the cofactors folate, vit. B12, and the vit. B6 vitamer pyridoxalphosphate (PLP). Metabolism of these vitamins is frequently disturbed in cirrhotics, but little is known about plasma Hcy levels in these patients. Methods: Plasma levels of Hcy, methionine, serine, cysteine, PLP, vit. B12 and folate, and standard clinical/biochemical parameters of liver disease were measured in 43 postabsorptive patients with biopsy proven cirrhosis of different origin. Results: 74% of the patients had elevated plasma Hcy levels defined as >13.4 umol/l (mean+2SD of healthy age matched controls). Increased plasma Hcy concentrations were seen in alcoholic as well as in non-alcoholic cirrhosis. Excluding patients with impaired renal function (n=7), Hcy concentrations remained elevated in 69% of the patients. We found a high prevalence of pathological plasma vitamin concentrations of 33% for increased vit. B12 levels and 5% and 80% for decreased folate and vit. B6 levels, respectively. Mean plasma vitamin B12 concentrations increased, folate remained unchanged and PLP concentrations decreased with deteriorating liver function. Hcy concentrations were correlated with levels of creatinine (r=0.44, P<0.01), serine (r=-0.46, P<0.01), and cysteine (r=0.38, P<0.05), but showed no association with parameters of liver function and with plasma levels of folate, vit. B12 und vit. B6. This was contrary to data obtained in healthy individuals. In a stepwise multiple regression serine and cysteine best explained the variance in Hcy levels. Conclusions: Elevated basal Hcy-plasma levels are frequently seen cirrhotic patients. Variations of Hcy concentration in liver cirrhosis are not explained by plasma levels of cofactors of Hcy metabolism. PMID- 11282485 TI - Frequent beta-catenin aberration in human hepatocellular carcinoma. AB - Recently, mutations in the beta-catenin gene in hepatocellular carcinoma (HCC) have been reported: approximately 20% of HCCs had activating mutations at the glycogen synthase kinase 3beta phosphorylation sites within the exon 3 of the beta-catenin gene. However, changes in the level of the beta-catenin protein in HCC have not been well studied. We examined, by Western blotting, the expression level of the beta-catenin protein in cancerous tissues in comparison with that in adjacent non-cancerous tissues obtained from 32 cases of HCC with hepatitis C. An increase in the beta-catenin protein level in cancerous tissue compared to that in adjacent non-cancerous tissue was found in 15 (46.9%) of 32 cases of HCC. Mutation in exon 3 of the beta-catenin gene was found in six (18.8%) of the 32 cases, in five of which the beta-catenin protein level was increased. In total, beta-catenin aberration was found in 16 (50.0%) of 32 cases of HCC. It should be noted that beta-catenin aberration was also found in early HCC although it was observed chiefly in advanced HCCs. These results indicate that beta-catenin aberration is a frequent event in the development of HCC and may facilitate the development of HCC in the course of chronic hepatitis. PMID- 11282486 TI - Correlation between p21(waf1) and p16(INK4a) expression in hepatocellular carcinoma. AB - Background: The cyclin dependent kinase p21(waf1) plays a crucial role in the regulation of cell cycle. The family of p53 proteins has the ability to induce p21(waf1), whereas p16(INK4a) modulates post-transcriptionally the expression of p21(waf1). Methods: Total 36 hepatocellular carcinomas (HCCs) and 24 paired adjacent liver tissues were evaluated for the following: (1) expression of p21(waf1) and p16(INK4a); (2) that of p21(waf1), p73 and p63 mRNAs; (3) genomic mutations and the loss of heterozygosity of p73 and p53; and (4) frequency of methylation in the 5'CpG promoter region of p16(INK4a). Results: In HCCs compared with the adjacent non-cancerous liver tissues, the expression of p21(waf1) and p16(INK4a) was reduced. Indeed, p21(waf1) was not detected in 36% (8/22) of HCCs in spite of the presence of p21(waf1) mRNA: among them, mutations of p53 gene were found in 50%, whereas a lack of p16(INK4a) expression in all of them. p21(waf1) and p16(INK4a) were reduced in proportion to the degree of methylation in p16(INK4a) gene. p73 did not mutated, and p63 did not expressed in HCCs. Conclusion: Methylation status of p16(INK4a) gene will play a part for reducing constitutive expression of p16(INK4a) and of p21(waf1) coordinately in HCCs. PMID- 11282487 TI - Loss of heterozygosity of the mannose 6-phosphate/insulin-like growth factor II receptor and p53 genes in human hepatocellular carcinoma. AB - Information about M6P/IGF2R and p53 genes in hepatocarcinogenesis is limited and controversial. We tested the loss of heterozygosity (LOH) of M6P/IGF2R and p53 genes in cirrhotic and neoplastic foci in surgically resected livers of 30 patients with hepatocellular carcinoma (HCC). The DNAs extracted from microdissected specimens were used for polymerase-chain-reaction (PCR)-based assay. LOH of the M6P/IGF2R gene in the primary HCCs was detected in 10 of 22 informative cases (45%). In five of these 10 cases (50%), LOH was detected in cirrhotic lesions adjacent to the HCCs. The allelic loss patterns of M6P/IGF2R in liver cirrhosis (LC) were identical to those in the corresponding HCC, suggesting that HCC could develop from one of the cells in which M6P/IGF2R had been lost. Furthermore, LOH of the p53 gene in HCC was detected in 10 (43%) of 23 informative cases, and p53 loss in cirrhotic foci adjacent to HCC was shown in one of the 10 cases (10%). The pattern of allelic loss of the p53 gene in the cirrhotic foci was identical with that in the corresponding tumor. The LOH of the M6P/IGF2R and p53 genes occurred independently in HCCs. LOH of the M6P/IGF2R locus was a relatively frequent and possibly early event in hepatocarcinogenesis, and LOH of the M6P/IGF2R gene and LOH of the p53 gene occurred independently. PMID- 11282489 TI - Expression of angiogenic factors, basic fibroblast growth factor and vascular endothelial growth factor, in human biliary tract carcinoma cell lines. AB - In order to clarify angiogenic mechanism in biliary tract carcinoma, expressions and functions of basic fibroblast growth factor (bFGF) and its receptors (FGFR-1 4), and vascular endothelial growth factor (VEGF) and its receptors were investigated by using human biliary tract carcinoma cell lines (KMC-1, KMC-2, KMBC and KMG-C). Expression of bFGF was confirmed in KMC-1 and KMC-2, and that of FGFR-1-4 in all the cell lines except no FGFR-2 in KMC-2. Expression of VEGF was detected in all the cell lines, whereas the cell lines did not express VEGF receptors. Addition of anti-bFGF neutralizing antibody to the medium did not suppress cell proliferation, whereas exogenous bFGF with or without heparin accelerated cell proliferation in all cell lines. Addition of anti-bFGF neutralizing antibody or anti-VEGF neutralizing antibody to the co-culture of human umbilical vascular endothelial cells (HUVEC) and KMC-2 suppressed the proliferation of HUVEC. Surgically obtained cholangiocarcinoma tissues (n=7) were immunohistochemically negative to bFGF, while six of the seven were positive to VEGF. These findings suggested that human biliary tract carcinoma cells express both bFGF and VEGF not as autocrine growth factors but as angiogenic factors. On the other hand, expression of VEGF was found at a higher frequency than bFGF both in the cell lines and tissues. PMID- 11282488 TI - Simple quantitative assay of alpha-fetoprotein mRNA in liver tissue using the real-time detection polymerase chain reaction assay - its application for clinical use. AB - Alpha-feto protein (AFP) mRNA levels increase in hepatocellular carcinoma (HCC) cells as compared with non-neoplastic tissue. Therefore, detection of AFP mRNA in blood nuclear cells is useful for the evaluation of treatment efficacy and prognosis of HCC. In this study, simple and reproducible methods were developed to quantify AFP mRNA using the real-time RT-PCR assay (Taq Man assay). By using in vitro synthesized AFP and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) RNA, the sensitivity and dynamic range of the RT-PCR assay were established. AFP mRNA in both HCC and non-neoplastic tissue, as well as in cell lines, were measured using this assay system. The expression of the AFP mRNA level was normalized using the GAPDH house keeping gene product as an endogenous reference. AFP and GAPDH mRNA can be quantified in the range of 10-10(8) copies when using this quantitative assay. Among HCC cell lines, Huh 7 and HepG2 cells, respectively, represented 1.5x10(6) and 6.0x10(5) AFP mRNA/10(6) GAPDH mRNA, in contrast to 6, 23 and 230 AFP mRNA/10(6) GAPDH mRNA for HLE, HLF and PLC/PRF/5 cells, respectively. Other cell lines derived from stomach, pancreas, and colon cancers have 10 AFP mRNA copies/10(6) GAPDH mRNA. In liver tissue from patients with chronic hepatitis, and the non-neoplastic portion of the liver from HCC patients, AFP mRNA distributes from 2.5x10(3) to 5.8x10(4)/10(6) GAPDH transcripts. In contrast, AFP mRNA in tumor cells were more than 100-fold higher than that found in corresponding non-neoplastic portions in two patients who had a high level of AFP in serum. The establishment of the TaqMan quantifying system for AFP mRNA may have important clinical implications. PMID- 11282490 TI - Androgen metabolism in regenerating liver of male rats: evidence for active uptake and utilization of testosterone. AB - The role of androgen in the regenerative process of the liver remains unclear. Male Sprague-Dawley rats were subjected to either 70% hepatectomy or sham operation. Immediately after surgical procedures, rats were injected with 0.3 uCi/g of body weight [3H]-testosterone from the inferior vena cava. The radioactivities of the remnant liver were counted for 0-24 h. For measurement of sex hormones and their metabolizing enzymes activities in the regenerating liver, the same experiments were also conducted, in which the rats were sacrificed up to 120 h. The plasma and hepatic testosterone, dihydrotestosterone(DHT) and estradiol were determined by radioimmunoassays. Uptake of [3H]-testosterone in the regenerating liver was significantly higher during the first 6 h. 5alpha reductase I and DHT were increased in parallel to the hepatocyte proliferation which was assessed by proliferating cell nuclear antigen (PCNA) labeling index. On the other hand, hepatic 3alpha-hydroxysteroid dehydrogenase level did not alter but aromatase activity significantly decreased after 70% hepatectomy. These results suggest that during the early phase of liver regeneration, testosterone was actively uptaken from the plasma by the regenerating liver and was converted into DHT by elevated hepatic 5alpha-reductase enzyme. Thus androgens might play a crucial role in liver regeneration. PMID- 11282491 TI - Effect of bile acids on biliary excretion of cyclosporin A in the rat. AB - Cyclosporin A, a substrate of P-glycoprotein (P-gp), is known to cause cholestasis in humans and in rat experimental models. Tauroursodeoxycholate is reported to be effective in CyA-induced cholestasis in rats. In the present study, to investigate the mechanism of the inhibition of CyA induced cholestasis, effect of bile acids on biliary cyclosporin A excretion was studied in rats. Infusion of both taurocholate and tauroursodeoxycholate at the rate of 0.8 mmol/min per 100 g bodyweight increased bile flow and biliary cyclosporin A excretion, and the extent was more prominent with tauroursodeoxycholate. It was suggested that these findings were caused by the enhanced vesicular targeting of P-gp to the canalicular membrane by bile acids, thus increasing the numbers of P gp in the canalicular membrane. PMID- 11282492 TI - The effect of trimetazidine on intrahepatic cholestasis caused by carmustine in rats. AB - This study investigated the effect of trimetazidine (TMZ), known as an anti oxidant agent, on intrahepatic cholestasis caused by Carmustine (BCNU) in rats. Rats were assigned into four groups. The first group (Saline) consisted of 12 rats, which were injected with 2 ml/kg of saline intraperitoneally (IP) 48 h before the study. The second group (corn oil group, n=15), which were injected with 2 ml/kg of corn oil IP 48 h before the study. The third group (BCNU group, n=16), which were injected with 2 ml/kg of corn oil+25 mg/kg BCNU IP 48 h before the study. The fourth group (TMZ group, n=12), which were injected with 2.5 mg/kg per day of TMZ IP, administered at the same hour of the day as a single-dose. Twelve hour after the first dose of TMZ, corn oil 2 ml/kg+BCNU 25 mg/kg IP were injected, and the rats were included in the study 48 h after the administration of corn oil+BCNU. Following a pentobarbital anaesthesia, abdomen was opened with incision, a cannula was placed into the channel of choledocus, and the amount of bile was measured per hour. Then intracardiac blood sample was taken, and consequently centrifuged to obtain the plasma. Finally, the rats were killed with cervical dislocation, and their livers were removed and weighted. In addition to histopathological examination of liver, the levels of malon dialdehyde (MDA), oxidised glutation (GSSG), and reduced glutation (GSH) were detected. Also the osmolality of bile and plasma was estimated in mOsm/kg. As a result, the biliary flow was seen to decrease in BCNU group (P<0.005), but to be normal in TMZ group. The serum level of conjugated biluribin was higher in BCNU group compared to other groups (P<0.05 for each). Although the level of total glutation was lower (P<0.005) in TMZ group, GSH/GSSG ratio was normal. These findings suggest that TMZ has a protective effect on intrahepatic cholestasis caused by BCNU. PMID- 11282493 TI - Endotoxin increases paracellular permeability of isolated rat hepatocyte couplets. AB - Hyperbilirubinemia is frequently associated with endotoxemia. Regurgitation of bile constituents including bilirubin into the sinusoidal space is prevented by tight junctions which maintain paracellular permeability between hepatocytes. To investigate the mechanism of endotoxin-associated hyperbilirubinemia, we have studied the changes in paracellular permeability of primary hepatocyte couplets treated with endotoxin. In addition, we examined the effects of ursodeoxycholic acid (UDCA), which has been widely used for various liver diseases, on endotoxin associated changes in paracellular permeability. The paracellular permeability of hepatocyte couplets was evaluated by paracellular penetration of fluorescein isothiocyanate (FITC)-dextran with molecular weights of 3, 10 and 70K using confocal laser scanning microscopy. Endotoxin increased the paracellular penetration of FITC-dextran 3 and 10K. These changes were prevented by treatment with UDCA. There was little paracellular penetration of FITC-dextran 70K under any conditions. These results suggested that endotoxin increased the paracellular permeability of hepatocyte couplets and these changes were prevented by treatment with UDCA. Furthermore, bile regurgitation through the paracellular route is involved in endotoxin-associated hyperbilirubinemia, and UDCA might be a potential therapeutic agent for endotoxin-associated hyperbilirubinemia. PMID- 11282494 TI - The Internet: a valuable tool for Roll Back Malaria. PMID- 11282496 TI - Malaria sporozoite rite of passage. PMID- 11282497 TI - Gender bias in grant application rates. PMID- 11282503 TI - Van Hellemond receives Merial Award. PMID- 11282504 TI - Helminths and atopy. PMID- 11282505 TI - Is Necator americanus approaching a mutualistic symbiotic relationship with humans? AB - The hookworm Necator americanus establishes infections of impressive longevity in the immunologically hostile environment of its human host. In the process, it promotes pronounced T-helper 2 (Th2) cell activity, which in turn seemingly affords the host at least a degree of protection. Given the relatively asymptomatic nature of infection, we argue here that Necator americanus might be approaching a mutualistic symbiotic relationship with humans. In our view, infection is controlled by the immune system while being supported by a subtle immune-evasion strategy that is tolerated and possibly beneficial to the host in certain immunological circumstances, such as in counterbalancing potentially damaging Th1 responses. PMID- 11282506 TI - Do helminths play a role in carcinogenesis? AB - Chronic helminthiasis is recognized as a significant factor in cancer development in humans. However, the mechanisms by which helminths initiate and promote malignant transformation of host cells are still not understood fully. Human helminthiasis can cause genetic instability and affect inter- and intracellular communication, ultimately leading to tumour development through inflammation, modulation of the host immune system, and secretion of soluble factors that interact with host cells. PMID- 11282507 TI - The immunology of myiasis: parasite survival and host defense strategies. AB - Infestations by dipterous larvae that feed on dead or living vertebrate tissues for a variable period are known as myiases; these infestations reduce host physiological functions, destroy host tissues and cause significant economic losses to livestock worldwide. Recent advances in understanding the specific and nonspecific immune responses of hosts to infestation by myiasis-causing larvae and the immunological strategies evolved by larvae against the host are reviewed here. The practical implications of immunological knowledge for diagnostic and vaccination strategies are also discussed, with a view to developing environmentally sustainable control methods to be used as an alternative to chemical treatments. PMID- 11282508 TI - The dangers of using single locus markers in parasite epidemiology: Ascaris as a case study. AB - Molecular markers are used widely to discriminate between closely related species of parasites, and in many cases a single locus is used for this purpose. This article aims to show how molecular data derived from a single genetic marker or linkage group - in this case mitochondrial DNA - can lead to ambiguous conclusions and to illustrate how a multilocus approach has enhanced our understanding of the epidemiology of two closely related parasites, the nematodes Ascaris suum, which infects pigs, and Ascaris lumbicoides, which infects humans. PMID- 11282509 TI - Trade-offs, conflicts of interest and manipulation in Plasmodium-mosquito interactions. AB - A long-held view among parasitologists is that infection by malaria parasites does not harm the mosquito vector. One of the reasons for this belief is that the two partners of the association share interests in the most important life history traits of the mosquito. Both partners benefit from increased survival and an increased rate of bloodfeeding the mosquito to increase its reproductive success and the parasite to ensure its transmission. Problems with this line of reasoning appear when one considers possible trade-offs among the mosquito's life history parameters, which constrain the attempts by the mosquito and the parasite to maximize their success. Could these constraints differ between the two partners and thus lead to conflicts of interest and what would be the evolutionary and epidemiological consequences of conflicting interests? These questions will be investigated below. PMID- 11282510 TI - Antibodies and Plasmodium falciparum merozoites. AB - There is considerable interest in using merozoite proteins in a vaccine against falciparum malaria. Observations that antibodies to merozoite surface proteins block invasion are a basis for optimism. This article draws attention to important and varied aspects of how antibodies to Plasmodium falciparum merozoites affect red blood cell invasion. PMID- 11282511 TI - Linking proteome and genome: how to identify parasite proteins. AB - Parasite genome projects are generating an avalanche of sequence data. If this resource is to be exploited effectively for drug and vaccine design, there is an urgent need to make the link between these DNA sequences and the functional proteins of the parasite, which they encode. Here, we seek to demystify the revolutionary advances in protein identification based on mass spectrometry. PMID- 11282515 TI - [Mycobacterium xenopi: epidemiological and bacteriological features]. AB - Mycobacterium xenopi is a scotochromogenic slow-growing atypical mycobacteria, with a thermostable catalase, no production of niacin and whose cell wall contains types I and VI long-chain fatty acids. Cosmopolitan, it is mainly recovered in tap-warm water. The contamination occurs through aerosol inhalation, water ingestion or use of contaminated medical or surgical equipment. M. xenopi is an opportunistic pathogen; the infection is facilitated by the incidental introduction of the bacteria in the body, pre-existing pulmonary lesions and an immunodepression. M. xenopi is mainly involved in infections of lungs, bones and joints. The treatment consists in the combination of three or four antibiotics, among rifampicin, rifabutin, ethambutol, macrolides, amikacin and fluoroquinolones. PMID- 11282516 TI - [Clinical, biological and genetic heterogeneity of the inborn errors of pulmonary surfactant metabolism: SP-B deficiency and alveolar proteinosis]. AB - Pulmonary surfactant is a multimolecular complex located at the air-water interface within the alveolus and to which a bulk of functions has been assigned, physical (surface-active properties) as well as immune or depurant. This complex consists of a surface active lipid layer (mainly phospholipids), and of an aqueous subphase. From discrete surfactant sub-fractions, one can isolate very hydrophobic proteins SP-B and SP-C as well as the collectins SP-A and SP-D, which were shown to have structural, metabolic, or defensive properties. Inborn or acquired abnormalities of surfactant, qualitative or quantitative in nature, account for a number human diseases. Beside hyaline membrane disease of the preterm neonate, a cluster of hereditary or acquired lung diseases have been characterized by the storage of periodic acid Schiff-positive material filling the alveoli. From this heterogeneous nosologic bulk, at least two discrete entities presently seem to emerge: 1) SP-B deficiency, in which an essentially proteinaceous material is stored within the alveoli, and which is a bona fide autosomal recessive Mendelian entity linked to the SFTPB gene (MIM 1786640), generally entailing neonatal respiratory distress with rapid fatal outcome, although partial or transient deficiencies have also been observed; 2) alveolar proteinosis, characterized by the storage of a mixed, protein and lipid material, and which constitutes a relatively heterogeneous clinical biological syndrome, with regards to age at onset (from the neonate through to adulthood) as well as the severity of associated signs. Murine models with a targeted mutation of the gene encoding GM-CSF (Csfgm) or the beta subunit of its receptor (Il3rbl) support the hypothesis of an abnormality of surfactant turnover in which the alveolar macrophage would be a key player. Beside SP-B deficiency, in which a near consensus diagnostic chart can be designed, the ascertainment of other abnormalities of surfactant metabolism is not straightforward. The disentanglement of this disease cluster is however essential, with aim to propose differentiated therapeutic procedure : repeated bronchoalveolar lavages, GM-CSF replacement, bone marrow grafting or lung transplantation. PMID- 11282517 TI - [Recommended algorithms of prescription in the diagnosis of iron deficiency and overload]. AB - In view of the recent development of new tests of biochemistry and molecular biology the assessment of iron status should be reconsidered and updated. The French Society of Clinical Biology (SFBC) and the French Society of Hematology (Cellular Hematology Group) recommend algorithms in the diagnosis of iron deficiency and iron overload bearing in mind the best efficiency and health economy. These recommendations are based on the known sensibility and specificity of each test. The analytical requirements for the determination of the tests as well as the clinical and biological signs evoking an iron deficiency or overload are recalled. PMID- 11282518 TI - [Evaluation of the Immulite 2000 Toxoplasma quantitative IgG et Toxoplasma IgM for the diagnosis of human toxoplasmosis]. AB - Four hundred and ninety five human sera with clinical and biological data were tested for the evaluation of Immulite 2000 Toxoplasma Quantitative IgG and Immulite 2000 Toxoplasma IgM produced by Diagnostic Products Corporation (Los Angeles, USA) for the diagnosis of human toxoplasmosis. The results of these kits were compared to those of the University Hospital of Nancy where the reference assays were Enzygnost Toxoplasmosis IgG and Enzygnost Toxoplasmosis IgM (Berhing Dade, Germany), Toxoscreen (bioMerieux, France), ISAgA Plus (IgM et IgA) (bioMerieux, France). The sensitivity and the specificity of IgG detection by Immulite 2000 Toxoplasma Quantitative IgG were 98% and 100%, respectively. The high sensitivity of IgM detection by Immulite 2000 Toxoplasma IgM was adapted to the early diagnosis of toxoplasmic primo-infection and to the pediatric diagnosis or follow-up of congenital toxoplasmosis but could reveal IgM a long time after primary infection. PMID- 11282519 TI - [Is detection of schizocytes by computerised image analysis accurate?]. AB - Schistocytes result from red cell fragmentation. The identification of the schistocytes is critical for decisions on appropriate management of the patients. Currently, a systematic approach to the counting method remains rewarded. We programmed a computer image analysis device (Q-Win, Leica) in order to detect fragmented red cells. A good correlation between the computer and a well-trained biologist was found after minor modifications of the computer's results. Image analysis should reduced the biologist-to-biologist variation and improve the identification and enumeration of the schistocytes. PMID- 11282520 TI - [Simultaneous quantitative determination of amprenavir and indinavir in human plasma by high-performance liquid chromatography]. AB - A reversed-phase high-performance liquid chromatographic assay for the determination of the HIV protease inhibitors amprenavir (Agenerase) and indinavir (Crixivan) in human plasma is described, using a mobile phase consisting of 0.50 M phosphate buffer (adjusted to pH 5,5) - Milli-Q water - acetonitrile (120: 1,080: 800, v/v/v). A solid-phase extraction using C18 extraction columns (Discovery columns 100 mg, 1 ml Supelco) and a liquid-liquid extraction with 0.5 ml hydrogenocarbonate/carbonate buffer (adjusted to pH 10.6) and 6 ml methyl ter butyl ether have been compared. The liquid-liquid extraction has been chosen to be easier and cheaper. The method has been validated over the range of 60 to 3,000 ng/ml for amprenavir and 20 to 3,000 ng/ml for indinavir using a 0.5 ml sample volume. The specificity, linearity, accuracy and precision have been studied. The limit of detection was respectively for amprenavir and indinavir 15 and 4 ng, and the limit of quantification was 60 and 20 ng/ml. Stability tests under various conditions were performed. This assay can readily be used in a hospital laboratory for the routine monitoring of plasma concentrations of amprenavir in HIV-infected patients. The trough plasma concentrations average has been determined in patients treated by amprenavir and indinavir for seven months. PMID- 11282521 TI - [Molecular tools in the epidemiology of tick-borne bacterial diseases]. AB - Molecular tools have been used to detect rickettsiae in ticks. In Ixodes ricinus ticks collected in France, we detected for the first time there an emerging pathogen, Rickettsia helvetica, and an Ehrlichia sp, closely related to the agent of human granulocytic ehrlichiosis. In Guadeloupe (French West Indies), we described the occurrence of African tick-bite fever due to Rickettsia africae, which had been previously reported in sub-Saharan Africa only. In Africa, we completed our knowledge about the distribution of R. africae (Mali, Niger, Sudan, Burundi), and detected for the first time Rickettsia mongolotimonae, an emerging pathogen. Anaplasma marginale the agent of bovine anaplasmosis was detected in Mali. Rickettsiae of unknown pathogenicity were detected in Mali and Niger. PMID- 11282522 TI - [Factor XI inhibitors : clinical and biological features]. PMID- 11282524 TI - [Evaluation of the XAS test for sera screening with visible interference in the ARS analyzer]. PMID- 11282523 TI - [Total urinary protein assays using a pyrogallol red automatized technique]. PMID- 11282525 TI - [4th Symposium of the Francophone Society of Vitamins and Biological Factors. Dijon, France, 3-4 December 1999]. PMID- 11282526 TI - [Seminar in Clinical Virology: the respiratory viruses. Caen, France, 5-6 October 2000]. PMID- 11282527 TI - [Internal quality control : false-rejections and preliminary period]. PMID- 11282528 TI - [Clinical biology based on proof or evidence-based laboratory medicine]. PMID- 11282529 TI - [Standardization and quality in the clinical laboratory : towards an European system of clinical laboratory sciences]. PMID- 11282530 TI - [The missing link: quality in the identification and preanalytic phase]. PMID- 11282531 TI - [Units in clinical chemistry: traditional or international system?]. PMID- 11282532 TI - [Collaboration between laboratories with regard to instances of food poisoning]. PMID- 11282540 TI - Regulation of interferon-gamma receptor (INF-gammaR) chains: a peculiar way to rule the life and death of human lymphocytes. AB - Interferon-gamma (IFN-gamma) is a lymphokine produced by activated T lymphocytes and NK cells, that plays an important role in host defense mechanisms by exerting pleiotropic activities on a wide range of cell types. Cellular responses to IFN gamma are mediated by its heterodimeric cell surface receptor (IFN-gammaR), which activates downstream signal transduction cascades, ultimately leading to the regulation of gene expression. Several observations suggest that the signals resulting from the binding of IFN-gamma to its receptor depend on the number of surface receptors transducing the IFN-gamma signal. This review summarizes recent advances in the understanding of the fine regulation of the response of human lymphocytes to IFN-gamma through an interplay between surface expression of IFN gammaR and a variety of environmental factors that combine to control their fate. PMID- 11282541 TI - The interleukin-6 cytokine system in embryonic development, embryo-maternal interactions and cardiogenesis. PMID- 11282542 TI - Role of cytokines in the modulation of nitric oxide production by cyclic AMP. AB - Enhanced levels of cyclic AMP (cAMP), resulting from stimulation of adenylyl cyclase through activation of distinct pharmacological receptor systems, have a remarkable impact on the activity of the immune system. Among other responses, production of nitric oxide (NO) is also affected. The effects of cAMP range from stimulation to inhibition (or no effect) of immune-stimulated biosynthesis of NO, with a preponderance of stimulatory interference. cAMP has been shown to be a potent, dual modulator of cytokine expression. It dose-dependently suppresses secretion of major NO up-regulatory cytokines tumor necrosis factor-alpha (TNF alpha) and interferon-gamma (IFN-gamma). On the other hand, production of IL-10, which is known to regulate the inducible NO synthase (iNOS) activation in both a positive and negative direction, is inversely enhanced. It is suggested that the dual effects of cAMP on NO formation are likely to result from the differences in the concentration ratio of these cytokines. The value of this parameter depends on the type and concentration of cAMP-stable derivatives and cAMP-enhancing agents, such as prostaglandins, beta-adrenoceptor agonists, phosphodiesterase inhibitors, forskolin and cholera toxin. The cytokine ratio may be influenced by dynamically developing multiple down- and up-regulatory feedback circuits among cytokines, NO, and cAMP. PMID- 11282543 TI - The 3' untranslated region of tumor necrosis factor-alpha is highly conserved in idiopathic pulmonary fibrosis (IPF). AB - Tumour necrosis factor alpha (TNF-alpha), a pro-inflammatory cytokine essential for the function of the immune system, has been implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). Production of TNF-alpha is regulated at both the transcriptional and post-transcriptional levels by a number of factors including interleukin-10 (IL-10). We have shown that there is significant TNF alpha activity in patients with IPF, despite the presence of significant amounts of IL-10 and Il-10R. IL-10 is thought to influence the tight translational repression of TNF-alpha mRNA in pulmonary macrophages. The essential element in this regulation is the AU-rich element (ARE) present in the 3' untranslated region of TNF-alpha. We hypothesised that polymorphism in the 3' UTR region of TNF-alpha explains the apparent failure of IL-10 to down regulate TNF-alpha in patients with IPF. Using single strand conformation polymorphism (SSCP) analysis, we have screened this region in 96 patients with IPF, using nine sets of overlapping PCR primers. All samples were successfully amplified for each primer set, but SSCP analysis was unable to detect point mutations or polymorphisms in the patients in any of the nine fragments. Results from this study suggest that the 3' UTR region of TNF-alpha is highly conserved in IPF and mutation of this region is unlikely to be involved in the pathogenesis of IPF. PMID- 11282544 TI - Endogenous interferon-gamma impairs bacterial clearance from lungs during Pseudomonas aeruginosa pneumonia. AB - Interferon (IFN-)gamma is thought to play a role in the resistance to various pathogens. To study the role of IFN-gamma in the pathogenesis of Pseudomonas pneumonia, IFN-gamma receptor (R) alpha-subunit-deficient [IFN-gammaR(-/-)] mice and wild type mice were intranasally inoculated with Pseudomonas aeruginosa (10(5) CFU). IFN-gammaR(-/-) mice demonstrated an enhanced clearance of P. aeruginosa from their lungs when compared to normal wild type mice (P < 0.05 at 24 hours after the infection), which was associated with a tendency towards an improved survival. These findings were not accompanied by a more effective activation of several components of the innate immune system known to contribute to host defense against pneumonia, i.e. the lung concentrations of cytokines and chemokines were similar in IFN-gammaR(-/-) and wild type mice, while the influx of neutrophils in bronchoalveolar lavage fluid (BALF) was even higher in wild type mice than in IFN-gammaR(-/-) mice. Remarkably, IFN-gammaR(-/-) mice had higher nitric oxide levels in the BALF at 24 hours after infection (P < 0.05). Endogenous IFN-gamma impairs rather than augments host defense during pneumonia caused by P. aeruginosa. PMID- 11282545 TI - Development of allergic contact dermatitis requires activation of both tumor necrosis factor-receptors. AB - We investigated the role of the TNF receptors, type I (p55TNFR) and type II (p75TNFR), in a mouse model of contact hypersensitivity, i.e., a model of a delayed type hypersensitivity (DTH) allergic reaction. Mice deficient for p55TNFR or p75TNFR were used to investigate the functions of these receptors in development of the DTH reaction. We show that both TNF receptors have a strong influence on the overall outcome of the DTH reaction, with the two TNF receptors exerting distinct functions. Dendritic cells of mice lacking p55TNFR had a defect in allergen uptake but showed normal migration into regional lymph nodes. In contrast, dendritic cells of p75TNFR-deficient mice showed diminished migration into regional lymph nodes after allergen contact, whereas the allergen uptake was independent of the p75TNFR. Thus, both TNF receptors are required for the development of a complete DTH reaction. PMID- 11282546 TI - Transforming growth factor beta-1 and interferon-alpha in the AIDS dementia complex (ADC): possible relationship with cerebral viral load? AB - Mechanisms involved in the pathogenesis of the AIDS-dementia complex are still unclear. The dichotomy between a small number of HIV-infected cells in the brain and their marked dysfunction could be related to a cellular amplification and/or activation of cerebral viral load by several cytokines. This link between cytokines and viral load could play a role in the generation of the clinical dementia syndrome. We have studied cerebral levels of transforming growth factor beta1 and interferon-alpha, both in the mild and severe AIDS-dementia complex and also compared these cytokines with HIV RNA load in patients with different degrees of dementia. Our data indicate that production of different cytokines characterized the expression of clinical dementia. In the mild AIDS-dementia complex, there was a significant inverse correlation between interferon-alpha and transforming growth factor-beta1 (r = - 0.743; p < 0.001), and HIV-RNA was present in inverse proportion to transforming growth factor beta1 (r = - 0.751; p < 0.001). In patients with severe AIDS-dementia, transforming growth factor-beta1 was undetectable, while interferon-alpha level were higher than in mild AIDS dementia and correlated positively to cerebral HIV-RNA. No significant difference was evident between these cytokines in the serum of ADC patients and in the control samples. Our study suggests that a relationship is possible between productive HIV infection in the cerebral nervous system and a heterogenous and different expression of the immune response via a complex interaction of cytokines with a differential modulation of the dementia phenotype. PMID- 11282547 TI - Interferon-beta (INF-beta) antibodies in interferon-beta1a- and interferon-beta1b treated multiple sclerosis patients. Prevalence, kinetics, cross-reactivity, and factors enhancing interferon-beta immunogenicity in vivo. AB - We analysed the role of dosage, route and frequency of administration of clinical grade interferon-beta (IFN-beta) preparations in inducing anti-IFN-beta antibodies (IFN-beta-Abs) in 5 groups of relapsing-remitting multiple sclerosis (RRMS) patients who were respectively treated as follows: 1) weekly intramuscular (i.m.) injections of 30 mg of recombinant IFN-beta1a (Avonex), 2) subcutis (s.c.) injections of 250 mg IFN-beta1b (Betaferon) every other day, 3) weekly i.m. injections of 250 mg IFN-beta1b (Betaferon), 4) s.c. injections of 22 mg of IFN beta1a (Rebif) three times a week, and 5) i.m. injections of 22 mg of IFN-beta1a (Rebif) twice a week. IFN-beta-Abs were determined by ELISA. IFN-beta1b was more immunogenic than IFN-beta1a not only when administered s.c. every other day, but also when administered i.m. at a lower weekly dose; i.m. injection, however, significantly delayed the appearance, and induced lower serum levels of IFN-beta Abs. In patients treated with s.c. IFN-beta1b, Ab levels peaked 3 to 9 months after therapy initiation, and then slowly, but progressively, declined to pre therapy levels that in some patients were reached after three years. Patients treated with i.m. or s.c. IFN-beta1a only rarely developed IFN-beta-Abs, and then at very low titers. Overall, the i.m. weekly administration of IFN-beta1a was the less immunogenic treatment. In IFN-beta1b-treated patients, a wash-out period of two/three months was sufficient to bring the IFN-beta-Ab levels below the cut off. Our findings suggest that the immunogenicity of IFN-beta1a is low, regardless of the route of administration and the dosage, while that of IFN beta1b is high, and is significantly, but not completely reduced by i.m. administration. As IFN-beta-Abs are cross-reactive, a wash-out period is suggested when the preparation is changed from IFN-beta1b to IFN-beta1a in order to maintain the clinical benefits of the therapy. PMID- 11282548 TI - The promoter polymorphism of the interleukin-6 gene regulates interleukin-6 production in neonates but not in adults. AB - In the promoter region of the IL-6 gene there is a single base exchange (G --> C) polymorphism at position -174. Recent findings suggest that this polymorphism may affect the transcription rate of the IL-6 gene and IL-6 plasma levels. To analyse its biological significance, we examined IL-6 plasma levels in cord blood and IL 6 production by neonatal cells after LPS-stimulation in relation to the presence of the IL-6G and IL-6C alleles. We hypothesized that since healthy neonates lack a previous exposure to exogenous antigens, their cytokine production could be genetically regulated. We also assumed that the normal labour-related stress could provide a physiological stimulus for IL-6 production. Cord blood was collected from 50 healthy, full-term neonates after normal vaginal delivery (VD) and from 42 healthy, full-term neonates after elective caesarean section (ECS). Adult samples were obtained from 450 healthy adult controls. The -174 polymorphism was analysed using PCR. IL-6 plasma levels and in vitro IL-6 production were measured using an ELISA method. Generally, IL-6 plasma levels in neonates were significantly higher than those in adults (neonates born by VD versus adults p < 0.001 and neonates born by ECS versus adults p < 0.001); the median value for neonates born by VD was 11.4 pg/ml (4.5-45.9), for neonates born by ECS 2.9 pg/ml (1.9-6.4) and for adults, 1.2 pg/ml (0.7-2.0). Surprisingly, cord blood IL-6 levels after VD differed significantly from those after ECS (p < 0.001). An analysis was carried out to ascertain if there was a genetic association between different IL-6 genotypes and IL-6 plasma levels in neonates. In the group of VD neonates with the CC genotype, non-carriers of the G allele, secreted significantly more IL-6 than carriers of the G allele (p < 0.03); 21.4 pg/ml (9.5-81.3) and 9.6 pg/ml (3.5-36.2) respectively. In line with this, ECS newborns with the CC genotype had higher IL-6 plasma levels than carriers of the G allele (p < 0.02); respective median values were 6.3 pg/ml (2.2-12.9) and 2.7 pg/ml (1.7-4.1). These findings were also supported when in vitro IL-6 production by neonatal mononuclear cells was compared carriers of the G allele and non carriers of the G allele. IL-6 levels were significantly lower in carriers of the G allele than in non-carriers (p < 0.04); respective median values were 6,980 pg/ml (4,175-16,800) and 17,425 pg/ml (11,400-33,900). In vivo or in vitro production of IL-6 of adult controls was not associated with the IL-6 -174 polymorphism. The difference between cord blood IL-6 levels after VD and after ECS suggests that normal labour-related stress induces IL-6 production. Our data also suggest that the -174 polymorphism of the IL-6 gene participates in the regulation of IL-6 responses in both groups of neonates. Furthermore, the naive IL-6 response of stimulated neonatal cells is associated with the -174 polymorphism of the IL-6 gene. In healthy adults, the regulation of IL-6 responses differs from that of healthy neonates, since baseline and inducible IL 6 levels in adults were not associated with this polymorphism. This indicates that the genetic regulation of IL-6 production can be observed in naive cells, while in adult cells previous contact with exogenous antigens probably modifies their responses. PMID- 11282549 TI - The hepatitis C virus (HCV) induces a long-term increase in interleukin-10 production by human CD4+ T cells (H9). AB - Patients with chronic hepatitis C present an imbalance of Th1/Th2 cytokine production. Therefore, we investigated whether the exposure of the CD4+ T cell line H9 to HCV could induce activation of cells through synthesis of IL-10. Three infection protocols were performed to enhance HCV propagation. Viral particles were prepared by ultracentrifugation of serum from patients. From 3 to 81 days post-infection (p.i.), HCV-RNA was monitored both in supernatants and cells by nested RT-PCR, IL-10 protein in medium by ELISA, and IL-10 mRNA in cells by semi quantitative RT-PCR. The expression of tetraspanins was analyzed by flow cytometry. The PKC signal pathway was studied using specific inhibitors. The H9 cells express CD81. HCV-RNA (+) was detected in cells until 21 days p.i, and in culture media over 39 days p.i. Up to day 81 p.i., HCV exposure induced a specific, 2-fold increase of IL-10 production by H9 cells. IL-10 production was inhibited by a PKC inhibitor (Calphostin C). This study shows that even if the infection of H9 T cells did not result in any viral progeny, HCV induced the activation of IL-10 secretion, which supports the role of IL-10 in HCV pathogenesis. PMID- 11282550 TI - Retrovirus-mediated gene transfer of the cytokine genes interleukin-1beta and tumor necrosis factor-alpha into human neuroblastoma cells: consequences for cell line behavior and immunomodulatory properties. AB - We have investigated the value of a gene therapy approach for neuroblastoma (NB), based on retroviral transduction of the IL-1beta or TNF-alpha cytokine genes into human NB lines. Secretion of the corresponding cytokine, was demonstrated in all lines, although with considerable quantitative variations. Cytokine gene expression significantly reduced the proliferation index (p = 0.0001); this effect was associated with either terminal neuronal (one TNF-alpha line) or fibroblast-like differentiation (two IL-1beta lines), leading to growth arrest after a few weeks. Cell surface levels of CD54 and HLA class II remained unaffected, but HLA class I (p < 0.001) and CD58 expression (p = 0.01) increased on SKNSH after TNF-alpha gene transfer. Mononuclear cells from normal allogeneic donors cocultured with both IL-1beta (p < 0.001) and TNF-alpha lines (p < 0.01), showed a significant increase in the proportion of activated T cells (CD3+DR+); however, their cytotoxicity and proliferation rate remained unchanged. Immunotherapy of neuroblastoma will require identification of transduced lines in which cytokine secretion induces phenotypic changes in such a way as to augment their likely immunomodulatory properties without impeding cell growth. Because of the limited efficacy of IL-1beta or TNF-alpha gene transfer alone, further studies should focus on combination with other immunomodulatory agents, to improve their potential efficacy in neuroblastoma. PMID- 11282551 TI - Characterisation of cytokine mRNA expression in tumour-infiltrating mononuclear cells and tumour cells freshly isolated from human colorectal carcinomas. AB - The in situ function of tumour-infiltrating leukocytes (TIL) in human colorectal adenocarcinoma (CRC) is unclear. Local cytokine expression probably regulates the anti-tumour immune response and tumour immune surveillance. We examined the distribution of mRNA for IFN-gamma, TNF-alpha, IL-10 and IL-4 in TIL, and tumour cells freshly isolated from 21 surgically removed primary CRC, using a semiquantitative RT-PCR. Lamina propria-infiltrating leukocytes (LPL) and epithelial cells from normal colon mucosa of 10 CRC patients served as negative controls. Median levels of IFN-gamma and TNF-alpha mRNA were higher in TIL than LPL (p = 0.0002 and 0.0001). IL-10 mRNA was generally observed in TIL and LPL, but no or very small amounts of IL-4 transcripts were detected in TIL and LPL. TNF-alpha and IL-10 mRNA were more abundant in colorectal tumour cells than in the normal epithelial cells (p = 0.0136 and 0.0036). The number of IFN-gamma transcripts in TIL correlated negatively (p = 0.039) and the number of TNF-alpha transcripts in tumour cells correlated positively with the Dukes' stages (p = 0.0147). Our results suggest that TIL are characterized by a type 1 (Th1/Tc1 like) pattern of cytokine expression and function as T cells (and macrophages) in the local, cell-mediated anti-tumour immune response in early stages of CRC. Changes in IFN-gamma and TNF-alpha mRNA in TIL and tumour cells could be related to tumour progress (e.g. by T cell anergy) or forming of metastases, respectively. PMID- 11282552 TI - Analysis of interleukin-18, interleukin-1 converting enzyme (ICE) and interleukin 18-related cytokines in Crohn's disease lesions. AB - A local increase of interleukin-18 (IL-18) expression has been recently demonstrated in Crohn's disease (CD), suggesting a role for mature IL-18 (cleaved by ICE protease) in the induction of proinflammatory cytokines and Th1 polarization observed in CD lesions. The aim of this study was to investigate IL 18 modulation and its potential immune consequences in CD lesions. We showed increased IL-18 production in chronic CD lesions and identified epithelial cells and macrophages as IL-18-producing cells. A twofold increase in ICE alpha, beta, and/or gamma mRNA that encodes for the complete mature peptide was required for ICE activity, and a marked increase in IL-18R-positive immune cells was observed in chronic lesions compared to uninvolved areas or normal control samples. Chronic lesions also displayed intense transcription of IL-18-induced cytokines, IFN-gamma, IL-1beta, TNF-alpha, and IL-8. By contrast, when neither IL-18 nor ICE mRNAs were enhanced (early asymptomatic CD lesions), IL-18-induced cytokines were not up-regulated. These results are in accordance with a putative role of mature IL-18 in the pathogenesis of CD. PMID- 11282553 TI - Daily production of human tumor necrosis factor in lipopolysaccharide (LPS) stimulated ex vivo blood culture assays. AB - Tumor necrosis factor (TNF) is a pleiotropic cytokine with immunological and neuroendocrine activities. A useful tool for studying TNF is the measurement of its in vitro and/or ex vivo over-expression, induced by a variety of stimuli on isolated peripheral mononuclear cells or whole blood, respectively. The capacity to over-express TNF, in ex vivo LPS-stimulated whole blood from 18 normal individuals, showed inter-individual variations ranging from high (3 ng/ml) to low (0.7 ng/ml) producers. Although at a lower level, a similar situation was observed in the spontaneous production of the cytokine. In order to detect cyclic effects in these variations, blood samples were taken at 08:00, 12:00, 16:00 and 20:00 hours, from nine healthy volunteers, and cultured in the ex vivo system. TNF and cortisol were measured by immunometric assays. Both, LPS-stimulated whole blood and plasma showed important, individual variations in TNF levels. Although cortisol levels presented a normal circadian cycle, these individual patterns in TNF production were basically conserved during the day (p > 0.05), and no correlation was observed between the levels of the hormone and those of the cytokine. When total TNF levels were determined at 20:00 hours, a moderate, temporary variation pattern of the cytokine production was found. These results suggest that cortisol does not play a predominant role in determining the ex vivo capacity of blood to produce TNF. Presumably, the variable capacity to produce the cytokine may have a strong genetic component. PMID- 11282554 TI - Inducible nitric oxide synthase attenuates chronic colitis in human histocompatibility antigen HLA-B27/human beta2 microglobulin transgenic rats. AB - Rats transgenic for HLA-B27/human beta2-microglobulin develop a spontaneous multisystem inflammatory disorder that closely mimics human spondyloarthropathies. Prominent features of this disorder are gut inflammation that predominates in the colon, and arthritis. Several mediators such as IFN gamma, IL-1beta, TNF-alpha, and inducible nitric oxide synthase (iNOS) have been found increased in the inflamed colonic mucosa. In the colon of HLA-B27 transgenic rats, iNOS is predominantly expressed by epithelial cells, and iNOS transcripts are detected in the hip cartilage of those rats, but not in nontransgenic littermates. The role of iNOS in this disorder was evaluated by administering the corticosteroid dexamethasone, or the NOS inhibitor L-N6-(1 iminoethyl)lysine (L-NIL) to HLA-B27 transgenic rats with established disease. Treatment with dexamethasone attenuated some aspects of gut inflammation, although it had no effect on iNOS expression. In contrast, treatment with L-NIL effectively inhibited iNOS activity, and resulted in an increase in colitis. Cytokine transcripts in the colon were modified by these treatments: IFN-gamma and IL-1beta were decreased after dexamethasone treatment, whereas administration of L-NIL resulted in decreased IFN-gamma, and TNF-alpha. A trend towards increased IL-1b expression was observed which could have contributed to the L-NIL pro-inflammatory effect. These results suggest that iNOS exerts a protective effect on colitis, in the inflammatory disorder of HLA-B27 transgenic rats. PMID- 11282555 TI - Up-regulation of mucin secretion in HT29-MTX cells by the pro-inflammatory cytokines tumor necrosis factor-alpha and interleukin-6. AB - The pro-inflammatory cytokines IL-6 and TNF-alpha have been implicated in the pathogenesis of otitis media with effusion (OME). A disease where goblet cells proliferate in a modified respiratory epithelium, leading to the accumulation of a mucin-rich effusion in the middle ear cleft. The MUC5AC and MUC5B mucin gene products have been identified as components of these effusions. To determine the effect of IL-6 and TNF-alpha on MUC5AC and MUC5B secretion we have used HT29-MTX goblet cells, which secrete both types of mucins. MUC5AC and MUC5B mucin secretion was measured by an enzyme-linked immunosorbent assay (ELISA) using a specific monoclonal antibody NCL-HGM-45M1 and polyclonal antiserum TEPA, respectively. Time response (0-72 hours) and dose response (1.5-150 ng/ml) studies were carried out. IL-6 and TNF-alpha stimulated MUC5AC and MUC5B mucin secretion in a time dependent manner, both in pre-confluent and post-confluent cells. IL-6 (15 ng/ml and 20 ng/ml) produced a low and prolonged stimulation of mucin secretion that persisted for 72 hours, with peak response at 24 hours after induction. The IL-6-mediated mucin secretion at 24 hours was concentration dependent, with a maximal effect at 15 ng/ml. TNF-alpha (20 ng/ml) induced rapid stimulation of mucin secretion within the first 24 hours, with peak response at 7 hours after induction. IL-6 and TNF-alpha exposure significantly increased MUC5AC secretion, but not MUC5B secretion. Maximal levels of cytokine-induced mucin secretion were detected in pre-confluent cells that showed one and a half- and two-fold increases in MUC5AC secretion after IL-6 and TNF-alpha stimulation, respectively, in comparison with post-confluent cells. The results presented here suggest that IL-6 and TNF-alpha generate a differential up-regulation of mucin secretion and thus contribute to the expression of mucin genes in inflammatory responses. PMID- 11282556 TI - A Jurkat T cell variant resistant to death receptor-induced apoptosis. Correlation with heat shock protein (Hsp) 27 and 70 levels. AB - Ligation of Fas induces an apoptotic program in Jurkat cells (Jd). We describe a Jurkat T cell variant (Jr) which shows total resistance to Fas-mediated apoptosis but which exhibits sensitivity to non-death-receptor pro-apoptotic stimuli such as staurosporine. Resistance to Fas-induced apoptosis in Jr cells is correlated with high expression of Hsps. A prior heat-shock increases Hsp27 and 70 expression and protects Jd and Jr cells from Fas- and staurosporine-induced apoptosis. Staurosporine, but not the anti-Fas antibody CH11, abrogates constitutive Hsp70 expression at 37 degrees C and staurosporine also inhibit Hsp27 expression in Jd and Jr cells at 42 degrees C. These data suggest that constitutive expression of Hsp27 inhibits Fas-mediated apoptosis, but only induced expression of Hsp70 can protect T cells from staurosporine-induced apoptosis. Thus, Hsp27 could play a role in the regulation of death receptor mediated apoptosis, while Hsp70 could regulate mitochondrial-dependent cell death. PMID- 11282557 TI - Increased cell surface expression of histocompatibility class II I-A(d) in c-fos transfected clones of the P388D1 murine macrophage cell line. AB - Numerous biological functions of the monocyte-macrophage lineage are affected by the presence of immunomodulators. Enhancement in transcription of c-fos has been shown in the murine P388D1 macrophage line treated by LPS, TPA, Ca++ ionophore or dibutyryl cAMP. In order to study the effects of an increased c-Fos protein level on macrophage functions, we previously have established stable c-fos overexpressing clones in the P388D1 cell line. Here we report that the expression of class II MHC antigens (I-A(d) antigen) is increased in these clones, particularly after IFN-gamma treatment. No variation in the cell surface expression of IFN-gamma receptor was observed. The increase of I-A(d) cell surface expression was well correlated with the level of I-A(d) mRNA. No inducible NO synthase activity and no increase of TNF-alpha release were observed in c-fos transfected cells. A slight increase of the basal expression of the main class II MHC transcription factor CIITA which is further amplified by IFN-gamma treatment, was observed in the c-Fos overexpressing clones. This suggests that the increased I-A(d) expression in clones could result from a transactivating action of the c-Fos protein on the CIITA factor. PMID- 11282558 TI - Transforming growth factor-beta inhibits interleukin-10 synthesis by human monocytic cells. AB - Transforming growth factor-beta (TGF-beta1) enhances interleukin-10 (IL-10) synthesis by mouse monocytes/macrophages, suggesting a potential role of IL-10 in mediating some of the anti-inflammatory properties of TGF-beta1. Since differences exist between the transcriptional regulation of human and mouse IL 10, the studies reported here examined whether TGF-beta1 up-regulated IL-10 production by human monocytes/macrophages as well. Exposure of PMA-differentiated U-937 promonocytic cells to TGF-beta1 resulted in an unexpected, dose-dependent decrease in IL-10 production as assessed by specific ELISA. TGF-beta1 was effective when added at the time of the PMA stimulus or 6 hours after. In addition, TGF-beta1 suppressed induction of IL-10 by three different stimuli other than PMA. TGF-beta1 inhibition of IL-10 protein release was associated with proportional changes in IL-10 mRNA accumulation as assessed by quantitative kinetic ELISA PCR. This would result from a decrease in IL-10 gene transcription as TGF-beta1 did not affect IL-10 mRNA stability, and TGF-beta1 limited the luciferase activity in cells transfected with reporter gene constructs containing 1,308 bp of the 5' non-coding sequence of human IL-10 gene. Blocking tumour necrosis factor-alpha (TNF-alpha) with neutralizing anti-TNF-alpha antibody did not modify the response to TGF-beta1, indicating the involvement of TNF-alpha independent mechanisms in the overall process. Thus, the present study provides the first evidence that TGF-beta1 prevents IL-10 production by human monocytic cells at a transcriptional level. PMID- 11282559 TI - Evidence for tumor necrosis factor receptors (TNFRs) in human MRC5 fibroblast cells. AB - Previous reports have shown that tumor necrosis factor alpha (TNF-alpha) controls the functions of fibroblasts cells, such as proliferation, cell migration and secretion of factors. The signaling for the biological effects of this and other cytokines is usually exerted through cell surface receptors. In this study, we demonstrated the presence of a surface and soluble TNF receptor in MRC5 fibroblasts. The presence of TNFRI and TNFRII mRNA was demonstrated by reverse transcriptase-PCR (RT-PCR). Both surface TNFRI and TNFRII are expressed and the number of both membrane receptors is 9,251 sites per cell. Using TNF receptor specific antibodies and flow cytometry, we showed that MRC5 cells express mainly TNFRI. PMID- 11282560 TI - Interleukin-9 induces 24P3 lipocalin gene expression in murine T cell lymphomas. AB - Interleukin-9 (IL-9) is a Th2 cytokine whose overexpression is associated with asthma and T cell lymphomagenesis. All the IL-9 activities studied so far are mediated by a specific hemopoietin receptor that activates a Jak/STAT pathway. Searching for genes specifically modulated by IL-9, we observed that the 24P3 mRNA is strongly upregulated in BW5147 T lymphoma cells upon IL-9 stimulation. 24P3 is a member of the lipocalin family, and has been reported to bind N-formyl Met-Leu-Phe, a potent neutrophil chemoattractant, and possibly other lipophilic mediators of inflammation. A similar 24P3 induction was observed in other T cell lymphomas (EL4 and TH201) in response to IL-9, as well as in EL4 cells stimulated with IL-6 or IL-1. By contrast, other IL-9-responsive cells such as mast cell line MC9 and B cell lymphoma A20 showed no 24P3 induction upon IL-9 stimulation. Experiments using IL-9R mutants indicated that STAT transcription factors, particularly STAT3, are involved in this process. However, 24P3 gene induction was slow, reaching a plateau from 36 to 72 hours after stimulation and was inhibited if cells were treated with cycloheximide during the first 8 hours of IL 9 stimulation, suggesting an indirect induction requiring new protein synthesis. PMID- 11282561 TI - Contrasting effect of interleukin-4 on the expression of cytosolic phospholipase A2, 5-lipoxygenase and 5-lipoxygenase-activating protein in peritoneal macrophages from control and ovalbumin-sensitized rats. AB - Interleukin-4 (IL-4), which has been widely described as an anti-inflammatory cytokine, can also exert proinflammatory effects. In this study, we extend these findings to demonstrate, in an allergic model, the dual effect of IL-4 on arachidonic acid (AA) metabolism in macrophages. In peritoneal macrophages from control rats (cPM), IL-4 had no effect on cPLA2 and 5-LO expression, but increased FLAP expression without affecting basal and A23187- or PMA-challenged arachidonic acid (AA) metabolism. In contrast, in peritoneal macrophages from ovalbumin-sensitized rats (sPM), IL-4 decreased cPLA2, 5-LO and FLAP expression and PMA-challenged eicosanoid production. A23187-challenged AA metabolism of sPM was not affected by IL-4 pretreatment. Thus, IL-4 acts differently on cPLA2, 5-LO and FLAP expression and AA metabolism in peritoneal macrophages depending on their resident or sensitization-induced differentiated status. PMID- 11282563 TI - Balance between autocrine interleukin-1beta and caspases defines life versus death of polymorphonuclear cells. AB - The role of endogenous IL-1beta in regulating spontaneous and Fas-triggered apoptosis of human PMN has been studied in relation to the activity of the IL 1beta-generating enzyme ICE (caspase-1), an enzyme also involved in the mechanism of cell death. Upon in vitro culture, PMN undergo spontaneous apoptosis and express increasing levels of IL-1beta, caspase-1- and caspase-3-like enzymes. Endogenous IL-1beta protects PMN from apoptosis, since inhibition of either IL 1beta or caspase-1 activity can accelerate PMN apoptotic death. Thus, in spontaneous PMN apoptosis caspase-1 essentially plays an anti-apoptotic role by inducing maturation of protective IL-1beta, whereas other molecules are responsible of driving apoptosis. Upon Fas triggering, PMN apoptosis is greatly accelerated, in correlation with increased caspase activity, whereas IL-1beta production is not augmented. Inhibition of IL-1beta activity can increase Fas induced apoptosis, whereas caspase-1 inhibitors are without significant effect. It is hypothesized that in Fas-induced PMN apoptosis caspase-1 has a double role: it can protect from apoptosis through generation of protective IL-1beta, as in spontaneous apoptosis, and it can also exert pro-apoptotic activity which counterbalances the protective effect and allows accelerated apoptosis. PMID- 11282562 TI - Fibrinogen cooperates with cytokines to induce interleukin-6 receptor mRNA expression in human hematopoietic CD34+ progenitors. AB - We have previously demonstrated that purified human fibrinogen (Fg), a major plasma component removed during serum preparation, shows mitogenic properties towards lymphoma cells and normal human hematopoietic progenitors. Indeed, adding Fg with IL-3 to a serum-containing medium stimulates growth of human CD34+ progenitors. In this report, we show in serum-free medium, that this stimulating effect only occurs in the presence of IL-6. To clarify the cooperative effect between Fg and IL-6, the kinetics of IL-6 receptor (IL-6R) mRNA expression in CD34+ cells have been analyzed by semi-quantitative in situ hybridization. In the presence of both IL-3 and Fg, more cells express IL-6R mRNA, and this expression per cell is significantly greater than with each factor added separately. These results suggest that Fg does not promote the growth of normal cells by itself, but sensitizes the cells to cytokines. Fg behaves not as a "progression" factor but as a typical "competence" factor, which induces a faster and greater IL-6R expression in early human hematopoietic progenitors by cooperating with other cytokines. PMID- 11282564 TI - Rat interleukin-10: production and characterisation of biologically active protein in a recombinant bacterial expression system. AB - Interleukin-10 is an anti-inflammatory Th1 immunosuppressive cytokine, the active form of which is a non-covalent homodimer, and which exhibits species-specificity both with respect to structure and biological activity. The rat homologue of IL 10 shares 73% identity with human IL-10 at the amino-acid sequence level, and has, in addition to the two disulphide bonds present in human IL-10, a fifth, unpaired cysteine (cys-149). Preparation of rat IL-10 by bacterial expression followed by solubilisation and refolding in a glutathione redox system, results in a molecule in which cys-149 is almost entirely oxidised, existing either as disulphide dimer or as a mixed disulphide with glutathione, and which has less than 1% of the activity of the native (cys-149-SH) form of the molecule. Site directed mutagenesis of rat IL-10 to replace cys-149 with tyrosine produces a molecule which readily adopts the active conformation upon solubilisation and refolding, and which is recoverable in good yield from bacterial expression systems. Comparison of the biological activities of rat IL-10tyr149 and commercial rat IL-10 preparations confirms that the activity of native-sequence rat IL-10 is either reduced or absent. It is proposed therefore that the biosynthetic analogue rat IL-10tyr149 is a more useful molecule to investigate the biological actions of IL-10 in the rat. PMID- 11282565 TI - Orienting attention in time. AB - Temporal information is essential for effective perception and action in the dynamic environment in which we exist. However, our ability to use information about time intervals flexibly to direct attention to an expected point in time has until recently been unexplored. Here we report a series of behavioural, neuroimaging and electrophysiological experiments that investigate and define the ability to orient attention in the temporal domain. These studies reveal that we are able to orient attention selectively to different time intervals, enhancing behavioural performance. These effects are mediated by a left-hemisphere dominant frontal-parietal system, which partially overlaps with the networks involved in spatial orienting. The optimisation of behaviour by temporal orienting appears to be achieved via motor-related mechanisms, in contrast to the typical perceptual enhancements produced by spatial attention. From a more general perspective, these findings illustrate the flexibility of attentional functions in the human brain. PMID- 11282567 TI - Regulation of glucose uptake in differentiated cells. AB - Glucose uptake into the cell is mediated by a family of glycosylated membrane proteins, called glucose transporters (GTs) that are able to facilitate passive hexose transfer across the lipid plasma membrane. The tissue-specific transporter isoforms generally differ in their affinity to the natural substrate D-glucose according to the specific functions of the respective organ. The mechanisms by which external and internal signals regulate glucose uptake into the cells belong to one of the most extensively studied fields of cell physiology. However, in spite of significant progress in identifying the involved molecular components and signaling pathways, the final cellular mechanism responsible for the short term regulation of glucose uptake is still a matter of intense debate. The widely accepted translocation hypothesis, which explains transport regulation by exo- and endocytic modulation of the number of GTs in the plasma membrane, insufficiently accounts for the whole insulin-induced transport stimulation and is insufficient to integrate the wide variety of different transport-modulating signals in differentiated tissue cells into a common mechanistic concept. Some time ago, a novel type of glucose transport regulation has been proposed prevailing in differentiated tissue cells. This mechanism depends on the presence of glucose transporters on microvilli of differentiated cells. The basic framework for this theory was provided by a recently presented novel concept of ion channel regulation via microvillar structures [Lange, K. (1999): Microvillar Ca++ signaling: A new view on an old problem. J. Cell. Physiol. 180, 19-35; Lange, K. (2000): Regulation of cell volume via microvillar ion channels. J. Cell. Physiol. 185, 21-35; Lange, K. (2000): Microvillar ion channels - Cytoskeletal regulation of ion fluxes. J. Theor. Biol. 206, 561-584], earlier studies on glucose transport regulation and a number actual biochemical findings. Here, a survey on both concepts is given and the ability of the novel mechanism of microvillar transport regulation to integrate a large body of experimental data into the common concept of cellular regulation via microvillar pathways is discussed. PMID- 11282566 TI - Simian virus 40 detection in human mesothelioma: reliability and significance of the available molecular evidence. AB - Simian virus 40 was discovered as a contaminant of early poliovirus vaccines that were inadvertently administered to millions of people in Europe and the United States between 1955 and 1963. Although SV40 was proven to be oncogenic in rodents and capable of transforming human and animal cells in vitro, its role in human cancer could not be proven epidemiologically. The matter was forgotten until 1993 when SV40 was accidentally found to cause mesotheliomas in hamsters injected intra-cardially. Subsequently, DNA sequences associated with its powerful oncogenic principal, the large T antigen, were found with high frequency in human pleural mesothelioma using the polymerase chain reaction (PCR). Since then many laboratories have confirmed the human findings. However, a few laboratories have failed to reproduce these data and the authors of the studies have claimed that the detection of SV40 DNA may simply represent PCR contamination artefacts. The controversy raised by this viewpoint is reviewed in this article together with a critical appraisal of the reliability of the molecular techniques used to detect SV40 DNA, in order to evaluate the potential aetiopathogenic role of SV40 in human mesothelioma. PMID- 11282568 TI - Single microassay for matrix degrading enzymes. AB - Matrix degrading enzymes are implicated in several disease processes such as abdominal aortic aneurysms and emphysema, however, monitoring proteolytic activity in a single assay is not well-established. Numerous assays have been developed to measure matrix degrading enzymes, which use artificial substrates or substrates derived from natural substrate protein. We have recently developed an assay for elastolytic activity based on the detection of primary amines, using trinitrobenzene sulfonic acid (TNBSA), following the digestion of succinylated elastin. The assay is also versatile enough to allow the detection of other proteases through the use of succinylated substrate specific for given protease. In order to improve the sensitivity and versatility of the assay we have refined the buffer conditions (PBS pH 7.2/1 mM CaCl2) to provide a 60 % increase in sensitivity to elastolytic activity and also formulated a substrate mixture containing succinylated elastin, collagen and gelatin. The combination of a substrate mixture and an optimum buffer will allow a spectrum of enzymes to be detected in a single reaction, providing a more complete picture of total proteolytic activity in a biological sample. This assay may also provide a tool to use proteolytic activity as a marker to monitor pathologic conditions involving matrix turn-over. PMID- 11282569 TI - A double blind randomized controlled trial of Maharishi Vedic vibration technology in subjects with arthritis. AB - To explore ancient Vedic medical techniques, one hundred and seventy-six subjects with arthritis participated in a controlled study through the non-pharmacologic approach known as the Maharishi Vedic Vibration Technology (MVVT). Using a double blinded and randomized experimental design, the findings showed significant reductions of pain and stiffness, and improvement in range of motion in the study sample. One hundred percent relief of symptoms was the most commonly reported category of improvement due to treatment. For the group as a whole, differences in mean response of treatment and control conditions with respect to relief of pain, limitation of motion, and reduction in stiffness were highly significant: t values ranged from a low of 5.609 in stiffness to a high of 20.950 in pain, p = 0.000009 to <10-49 respectively. Analysis by sub-categories of peripheral arthritis, painful conditions of the spine, and rheumatoid arthritis likewise produced significant results. Mechanisms of action were proposed, drawing on Maharishi Vedic Science, developments in quantum field theory, and specifically the theories of chaos and self-organizing systems as they relate to physiological functioning. The instantaneous relief of pain and improvement in function in such a high proportion of subjects with chronic arthritis is unparalleled in modern medical science PMID- 11282570 TI - Maharishi Vedic vibration technology on chronic disorders and associated quality of life. AB - There is a growing interest for more effective, innovative programs to address the chronic illness suffered by approximately 40 percent of the U.S. population. The purpose of this study was to evaluate the effects of a new Maharishi Vedic Medicine program-the Maharishi Vedic Vibration Technology-on the quality of life of individuals with chronic disorders. A total of 213 individuals took part in the study (mean age=48.55 years; average length of time of chronic illness=18.42 years). Results showed that over three sessions, the average self-reported improvement in chronic illness was 40.97 percent. Conditions related to neck pain improved the most (51.25%), followed by respiratory ailments (48.00%), digestive problems (46.90%), mental health, including anxiety and depression (46.34%), arthritis (41.57%), insomnia (37.38%), back pain (36.32%), headaches (35.83%), cardiovascular conditions (22.31%), and eye problems (21.19%). Findings also showed significant reductions in frequency of discomfort or pain (p<.000001), intensity of discomfort (p<.000001), and disabling effects of the discomfort in daily activity (p<.000001), in addition to overall improvement in mental health (p<.000001) and vitality (p<.000125). Possible mechanisms of action are presented. PMID- 11282571 TI - The role of DNA methyltransferase 1 in growth control. AB - Vertebrate DNA contains in addition to the four bases comprising the genetic information a modified base, 5-methyl cytosine that plays an important role in the epigenome. The methylated bases form a pattern of methylation that is cell specific and is faithfully inherited during cell division. The enzyme DNA methyltransferase 1 DNMT1 is responsible for copying the DNA methylation pattern but other de novo methyltransferase as well as demethylases might also be involved. Multiple mechanisms are in place to ensure the coordinate inheritance of the DNA methylation pattern with DNA replication. There is a bilateral relationship between the cell cycle and DNMT1. The expression of DNMT1 is tightly regulated with the cell cycle while the expression of DNMT1 can affect the cell cycle. DNMT1 protein might regulate cell cycle events by mechanisms that are independent of its DNA methylation activity through its multiple protein-protein interactions. The unique position of DNMT1 in the cell cycle is consistent with the hypothesis that it plays an important role in cancer. PMID- 11282572 TI - Clustering amino acid contents of protein domains: biochemical functions of proteins and implications for origin of biological macromolecules. AB - Structural classes of protein domains correlate with their amino acid compositions. Several successful algorithms (that use only amino acid composition) have been elaborated for the prediction of structural class or potential biochemical significance. This work deals with dynamic classification (clustering) of the domains on the basis of their amino acid composition. Amino acid contents of domains from a non-redundant PDB set were clustered in 20 dimensional space of amino acid contents. Despite the variations of an empirical parameter and non-redundancy of the set, only one large cluster (tens-hundreds of proteins) surrounded by hundreds of small clusters (1-5 proteins), was identified. The core of the largest cluster contains at least 64% DNA (nucleotide)-interacting protein domains from various sources. About 90% of the proteins of the core are intracellular proteins. 83% of the DNA/nucleotide interacting domains in the core belong to the mixed alpha-beta folds (a+b, a/b), 14% are all-alpha (mostly helices) and all-beta (mostly beta-strands) proteins. At the same time, when core domains that belong to one organism (E.coli) are considered, over 80% of them prove to be DNA/nucleotide interacting proteins. The core is compact: amino acid contents of domains from the core lie in relatively narrow and specific ranges. The core also contains several Fe-S cluster-binding domains, amino acid contents of the core overlap with ferredoxin and CO dehydrogenase clusters, the oldest known proteins. As Fe-S clusters are thought to be the first biocatalysts, the results are discussed in relation to contemporary experiments and models dealing with the origin of biological macromolecules. The origin of most primordial proteins is considered here to be a result of co-adsorption of nucleotides and amino acids on specific clays, followed by en-block polymerization of the adsorbed mixtures of amino acids. PMID- 11282574 TI - A six-year study of sapling and large-tree growth and mortality responses to natural and induced variability in precipitation and throughfall. AB - Global climatic change may cause changes in regional precipitation that have important implications for forest growth in the southern United States. In 1993, a stand-level experiment was initiated on Walker Branch Watershed, Tennessee, to study the sensitivity of forest saplings and large trees to changes in soil water content. Soil water content was manipulated by gravity-driven transfer of precipitation throughfall from a dry treatment plot (-33%) to a wet treatment plot (+33%). A control plot was included. Each plot was 6400 m2. Measurements of stem diameter and observations of mortality were made on large trees and saplings of Acer rubrum L., Cornus florida L., Liriodendron tulipifera L., Nyssa sylvatica Marsh, Quercus alba L. and Quercus prinus L. every 2 weeks during six growing seasons. Saplings of C. florida and A. rubrum grew faster and mortality was less on the wet plot compared with the dry and control plots, through 6 years of soil water manipulation. Conversely, diameter growth of large trees was unaffected by the treatments. However, tree diameter growth averaged across treatments was affected by year-to- year changes in soil water status. Growth in wet years was as much as 2-3 times greater than in dry years. Relationships between tree growth, phenology and soil water potential were consistent among species and quantitative expressions were developed for application in models. These field growth data indicate that differences in seasonal patterns of rainfall within and between years have greater impacts on growth than percentage changes in rainfall applied to all rainfall events. PMID- 11282573 TI - Histone acetylation and the cell-cycle in cancer. AB - A number of distinct surveillance systems are found in mammalian cells that have the capacity to interrupt normal cell-cycle progression. These are referred to as cell cycle check points. Surveillance systems activated by DNA damage act at three stages, one at the G1/S phase boundary, one that monitors progression through S phase and one at the G2/M boundary. The initiation of DNA synthesis and irrevocable progression through G1 phase represents an additional checkpoint when the cell commits to DNA synthesis. Transition through the cell cycle is regulated by a family of protein kinase holoenzymes, the cyclin-dependent kinases (Cdks), and their heterodimeric cyclin partner. Orderly progression through the cell cycle checkpoints involves coordinated activation of the Cdks that, in the presence of an associated Cdk-activating kinase (CAK), phosphorylate target substrates including members of the "pocket protein" family. One of these, the product of the retinoblastoma susceptibility gene (the pRB protein), is phosphorylated sequentially by both cyclin D/Cdk4 complexes and cyclin E/Cdk2 kinases. Recent studies have identified important cross talk between the cell cycle regulatory apparatus and proteins regulating histone acetylation. pRB binds both E2F proteins and histone deacetylase (HDAC) complexes. HDAC plays an important role in pRB tumor suppression function and transcriptional repression. Histones are required for accurate assembly of chromatin and the induction of histone gene expression is tightly coordinated. Recent studies have identified an important alternate substrate of cyclin E/Cdk2, NPAT (nuclear protein mapped to the ATM locus) which plays a critical role in promoting cell-cycle progression in the absence of pRB, and contributes to cell-cycle regulated histone gene expression. The acetylation of histones by a number of histone acetyl transferases (HATs) also plays an important role in coordinating gene expression and cell-cycle progression. Components of the cell-cycle regulatory apparatus are both regulated by HATs and bind directly to HATs. Finally transcription factors have been identified as substrate for HATs. Mutations of these transcription factors at their sites of acetylation has been associated with constitutive activity and enhanced cellular proliferation, suggesting an important role for acetylation in transcriptional repression as well as activation. Together these studies provide a working model in which the cell-cycle regulatory kinases phosphorylate and inactivate HDACs, coordinate histone gene expression and bind to histone acetylases themselves. The recent evidence for cross-talk between the cyclin-dependent kinases and histone gene expression on the one hand and cyclin dependent regulation of histone acetylases on the other, suggests chemotherapeutics targeting histone acetylation may have complex and possibly complementary effects with agents targeting Cdks. PMID- 11282575 TI - Ascorbic acid changes the pattern of purine metabolism during germination of white spruce somatic embryos. AB - It has previously been shown that exogenous applications of ascorbic acid (AA) increase the conversion frequency of somatic embryos of white spruce (Picea glauca (Moench) Voss). To determine whether ascorbic acid alters purine metabolism during the early phases of embryo germination, the relative rates of purine salvage and degradation were investigated by following the metabolic fates of exogenously applied [8-14C]adenine, [8-14C]adenosine, and [8-14C]inosine, and the activities of several key enzymes. We demonstrated that both the salvage and the degradation pathways operate during germination. Specifically, adenine and adenosine were mainly salvaged to nucleotides and nucleic acids, whereas an appreciable amount of inosine was degraded to CO2 and ureides. Comparisons of purine metabolism between control and AA-treated embryos showed that exogenous applications of ascorbic acid enhanced the ability of the embryos to take up adenine and adenosine throughout the germination period. Furthermore, the higher enzymatic activities of adenosine kinase and adenine phosphoribosyltransferase were responsible for the larger proportion of adenine and adenosine being salvaged in AA-treated embryos compared with control embryos. Thus, there was a positive correlation between the ability to anabolize purine precursors and successful embryo conversion. PMID- 11282576 TI - Production efficiency of loblolly pine and sweetgum in response to four years of intensive management. AB - We tested the hypothesis that productivity of intensively managed loblolly pine (Pinus taeda L.) and sweetgum (Liquidambar styraciflua L.) stands is dependent not only on leaf area, but also on foliar photosynthetic rate. Effects of irrigation (irrigation treatment), irrigation with a fertilizer solution (fertigation treatment), and fertigation + pest control (loblolly pine only; fertigation + pest control treatment) on leaf physiology and growth were compared with control plots during the third and fourth growing seasons. Complete weed control was maintained on all plots. Aboveground net primary productivity of loblolly pine and sweetgum increased from 16.3 to 40.5 Mg ha(-1) and from 4.2 to 23.9 Mg ha(-1), respectively, in response to the most intensive treatment. Relative to the control treatment, neither fertigation of sweetgum nor fertigation + pest control of loblolly pine had a significant or consistent influence on foliar N concentration, quantum yield, carboxylation efficiency, net photosynthesis, stomatal conductance, or production efficiency (increment in woody biomass per unit leaf area). Irrigation increased predawn leaf water potential and photosynthesis of loblolly pine, but it had no effect on production efficiency. Leaf area was the predominant determinant of maximum productivity in these rapidly growing plantations. PMID- 11282577 TI - Photosynthetic light acclimation in peach leaves: importance of changes in mass:area ratio, nitrogen concentration, and leaf nitrogen partitioning. AB - Photosynthetic light acclimation of leaves can result from (i) changes in mass based leaf nitrogen concentration, Nm, (ii) changes in leaf mass:area ratio, Ma, and (iii) partitioning of total leaf nitrogen among different pools of the photosynthetic machinery. We studied variations in Nm and Ma within the crowns of two peach (Prunus persica L. Batsch) trees grown in an orchard in Portugal, and one peach tree grown in an orchard in France. Each crown was digitized and a 3-D radiation transfer model was used to quantify the intra-crown variations in time integrated leaf irradiance, . Nitrogen concentration, leaf mass:area ratio, chlorophyll concentration, and photosynthetic capacity were also measured on leaves sampled on five additional peach trees in the orchard in Portugal. The data were used to compute the coefficients of leaf nitrogen partitioning among carboxylation, bioenergetics, and light harvesting pools. Leaf mass:area ratio and area-based leaf nitrogen concentration, Na, were nonlinearly related to , and photosynthetic capacity was linearly related to Na. Photosynthetic light acclimation resulted mainly from changes in Ma and leaf nitrogen partitioning, and to a lesser extent from changes in Nm. This behavior contrasts with photosynthetic light acclimation observed in other tree species like walnut (Juglans regia L.) in which acclimation results primarily from changes in Ma. PMID- 11282578 TI - Winter stem xylem pressure in walnut trees: effects of carbohydrates, cooling and freezing. AB - Pressure transducers were attached to twigs of orchard trees and potted trees of walnut (Juglans regia L.) to measure winter stem xylem pressures. Experimental potted trees were partially defoliated in the late summer and early autumn to lower the amount of stored carbohydrates. Potted trees were placed in cooling chambers and subjected to various temperature regimes, including freeze-thaw cycles. Xylem pressures were inversely proportional to the previous 48-h air temperature, but positively correlated with the osmolarity of the xylem sap. Defoliated trees had significantly lower concentrations of stored carbohydrates and significantly lower xylem sap osmolarities than controls. Plants kept at 1.5 degrees C developed xylem pressures up to 40 kPa, just 7% of the theoretical osmotic pressure of the xylem sap. However, exposure to low, nonfreezing temperatures followed by freeze-thaw cycles resulted in pressures over 210 kPa, which was 39% of the theoretical osmotic pressure. A simple osmotic model could account for the modest positive winter pressures at low, nonfreezing temperatures, but not for the synergistic effects of freeze-thaw cycles. PMID- 11282579 TI - Influence of tree internal N status on uptake and translocation of C and N in beech: a dual 13C and 15N labeling approach. AB - Influence of plant internal nitrogen (N) stocks on carbon (C) and N uptake and allocation in 3-year-old beech (Fagus sylvatica L.) was studied in two 15N- and 13C-labeling experiments. In the first experiment, trees were grown in sand and received either no N nutrition (-N treatment) or 4 mM unlabeled N (+N treatment) for 1 year. The -N- and +N-pretreated trees were then supplied with 4 mM 15N and grown in a 13CO2 atmosphere for 24 weeks. In the second experiment, trees were pretreated with 4 mM 15N for 1 year and then supplied with unlabeled N for 24 weeks and the remobilization of stored 15N was monitored. On the whole-plant level, uptake of new C was significantly reduced in -N-pretreated trees; however, partitioning of new C was not altered, although there was a trend toward increased belowground respiration. The amount of N taken up was not influenced by N nutrition in the previous year. In +N-pretreated trees, partitioning of new N was dominated by the fine roots (59.7% at Week 12), whereas in -N-pretreated trees, partitioning of new N favored stem, coarse roots and fine roots (24, 21 and 31.9%, respectively, at Week 12), indicating the formation of N stores. The contribution of previous-year N to leaf N was about 15%. The N remobilized for leaf formation had been stored in stem and coarse roots. We conclude that, within a growing season, the growth of beech is strongly determined by the availability of tree internal N stores, whereas the current N supply is of less importance. PMID- 11282580 TI - Enhanced transpiration in response to wind effects at the edge of a blue gum (Eucalyptus globulus) plantation. AB - In Australia, tree planting has been widely promoted to alleviate dryland salinity and one proposed planting configuration is that of strategically placed interception belts. We conducted an experiment to determine the effect of tree position in a belt on transpiration rate. We also assessed how much the effect of tree position can be explained by advection and environmental conditions. Daily transpiration rates were determined by the heat pulse velocity technique for four edge and 12 inner trees in a 7-year-old Tasmanian blue gum (Eucalyptus globulus) plantation in South Australia. Various climatic variables were logged automatically at one edge of the plantation. The relationship between daily sap flow and sapwood area was strongly linear for the edge trees (r2 = 0.97), but only moderately correlated for the inner trees (r2 = 0.46), suggesting an edge effect. For all trees, sap flow normalized to sapwood area (Qs) increased with potential evaporation (PE) initially and then became independent as PE increased further. There was a fairly close correlation between transpiration of the edge and inner trees, implying that water availability was partially responsible for the difference between inner and edge trees. However, the ratio of edge tree to inner tree transpiration differed from unity, indicating differences in canopy conductance, which were estimated by an inverse form of the Penman-Monteith equation. When canopy conductances were less than a critical value, there was a strong linear relationship between Qs of the edge and inner trees. When canopy conductances of the edge trees were greater than the critical value, the slope of the linear relationship was steeper, indicating greater transpiration of the edge trees compared with the inner trees. This was interpreted as evidence for enhancement of transpiration of the edge trees by advection of wind energy. PMID- 11282581 TI - Within-tree variation in transpiration in isolated evergreen oak trees: evidence in support of the pipe model theory. AB - Within-tree variation in sap flow density (SFD) was measured in two isolated evergreen oak (Quercus ilex L.) trees growing in an oak savannah (dehesa) in southwest Spain. Sap flow was estimated by the constant heating method. Three sensors were installed in the trunk of each tree in three orientations: northeast (NE), northwest (NW) and south (S). Sap flow density was monitored continuously from May 18 to September 27, 1993. Daily values of SFD ranged between 500 and 4500 mm3 mm-2 day-1. There were significant differences in SFD between orientations; SFD was higher in the NE and NW orientations than in the S orientation. These differences were noted on both a daily and seasonal time scale, and were less pronounced on cloudy days and at the end of the drought period, when SFD was relatively low. Our results support the idea that branches of trees can be viewed as a collection of small independent plants. PMID- 11282583 TI - Construction and analysis of photolyase mutants of Pseudomonas aeruginosa and Pseudomonas syringae: contribution of photoreactivation, nucleotide excision repair, and mutagenic DNA repair to cell survival and mutability following exposure to UV-B radiation. AB - Based on nucleotide sequence homology with the Escherichia coli photolyase gene (phr), the phr sequence of Pseudomonas aeruginosa PAO1 was identified from the genome sequence, amplified by PCR, cloned, and shown to complement a known phr mutation following expression in Escherichia coli SY2. Stable, insertional phr mutants containing a tetracycline resistance gene cassette were constructed in P. aeruginosa PAO1 and P. syringae pv. syringae FF5 by homologous recombination and sucrose-mediated counterselection. These mutants showed a decrease in survival compared to the wild type of as much as 19-fold after irradiation at UV-B doses of 1,000 to 1,550 J m(-2) followed by a recovery period under photoreactivating conditions. A phr uvrA mutant of P. aeruginosa PAO1 was markedly sensitive to UV B irradiation exhibiting a decrease in survival of 6 orders of magnitude following a UV-B dose of 250 J m(-2). Complementation of the phr mutations in P. aeruginosa PAO1 and P. syringae pv. syringae FF5 using the cloned phr gene from strain PAO1 resulted in a restoration of survival following UV-B irradiation and recovery under photoreactivating conditions. The UV-B survival of the phr mutants could also be complemented by the P. syringae mutagenic DNA repair determinant rulAB. Assays for increases in the frequency of spontaneous rifampin-resistant mutants in UV-B-irradiated strains containing rulAB indicated that significant UV B mutability (up to a 51-fold increase compared to a nonirradiated control strain) occurred even in the wild-type PAO1 background in which rulAB only enhanced the UV-B survival by 2-fold under photoreactivating conditions. The frequency of occurrence of spontaneous nalidixic acid-resistant mutants in the PAO1 uvrA and uvrA phr backgrounds complemented with rulAB were 3.8 x 10(-5) and 2.1 x 10(-3), respectively, following a UV-B dose of 1,550 J m(-2). The construction and characterization of phr mutants in the present study will facilitate the determination of the roles of light and dark repair systems in organisms exposed to solar radiation in their natural habitats. PMID- 11282584 TI - Long-range (1)H-(15)N heteronuclear shift correlation at natural abundance: a tool to study benzothiazole biodegradation by two rhodococcus strains. AB - The biodegradation of benzothiazole and 2-hydroxybenzothiazole by two strains of Rhodococcus was monitored by reversed phase high-pressure liquid chromatography and by (1)H nuclear magnetic resonance (NMR). Both xenobiotics were biotransformed into a hydroxylated derivative of 2-hydroxybenzothiazole by these two strains. The chemical structure of this metabolite was determined by a new NMR methodology: long-range (1)H-(15)N heteronuclear shift correlation without any previous (15)N enrichment of the compound. This powerful NMR tool allowed us to assign the metabolite structure to 2,6-dihydroxybenzothiazole. PMID- 11282585 TI - Biological activities and structural properties of the atypical bacteriocins mesenterocin 52b and leucocin b-ta33a. AB - The antibacterial spectra and modes of action of synthetic peptides corresponding to mesenterocin 52B and leucocin B-TA33a greatly differ despite their high sequence homology. Circular dichroism experiments establish the capacity of each of these two peptides to partly fold into an amphiphilic helix that might be crucial for their adsorption at lipophilic-hydrophilic interfaces. PMID- 11282586 TI - Bovine rotavirus nonstructural protein 4 produced by Lactococcus lactis is antigenic and immunogenic. AB - Rotavirus nonstructural protein 4 (NSP4) can induce diarrhea in mice. To get insight into the biological effects of NSP4, production of large quantities of this protein is necessary. We first tried to produce the protein in Escherichia coli, but the nsp4 gene proved to be unstable. The capacity of the generally regarded as safe organism Lactococcus lactis to produce NSP4 either intra- or extracellularly was then investigated by using the nisin-controlled expression system. Production of recombinant NSP4 (rNSP4) was observed in L. lactis for both locations. In spite of a very low secretion efficiency, the highest level of production was obtained with the fusion between a lactococcal signal peptide and rNSP4. Cultures of the rNSP4-secreting strain were injected into rabbits, and a specific immune response was elicited. The anti-rNSP4 antibodies produced in these rabbits recognized NSP4 in MA104 cells infected by rotavirus. We showed that L. lactis is able to produce antigenic and immunogenic rNSP4 and thus is a good organism for producing viral antigens. PMID- 11282587 TI - Use of pulsed-field gel electrophoresis and flagellin gene typing in identifying clonal groups of Campylobacter jejuni and Campylobacter coli in farm and clinical environments. AB - Although campylobacters have been isolated from a wide range of animal hosts, the association between campylobacters isolated from humans and animals in the farm environment is unclear. We used flagellin gene typing and pulsed-field gel electrophoresis (PFGE) to investigate the genetic diversity among isolates from animals (cattle, sheep, and turkey) in farm environments and sporadic cases of campylobacteriosis in the same geographical area. Forty-eight combined fla types were seen among the 315 Campylobacter isolates studied. Six were found in isolates from all four hosts and represented 50% of the total number of isolates. Seventy-one different SmaI PFGE macrorestriction profiles (mrps) were observed, with 86% of isolates assigned to one of 29 different mrps. Fifty-seven isolates from diverse hosts, times, and sources had an identical SmaI mrp and combined fla type. Conversely, a number of genotypes were unique to a particular host. We provide molecular evidence which suggests a link between campylobacters in the farm environment with those causing disease in the community. PMID- 11282589 TI - Genetic evidence for a defective xylan degradation pathway in Lactococcus lactis. AB - Genetic and biochemical evidence for a defective xylan degradation pathway was found linked to the xylose operon in three lactococcal strains, Lactococcus lactis 210, L. lactis IO-1, and L. lactis NRRL B-4449. Immediately downstream of the xylulose kinase gene (xylB) (K. A. Erlandson, J.-H. Park, W. El Khal, H.-H. Kao, P. Basaran, S. Brydges, and C. A. Batt, Appl. Environ. Microbiol. 66:3974 3980, 1999) are two open reading frames encoding a mutarotase (xylM) and a xyloside transporter (xynT) and a partial open reading frame encoding a beta xylosidase (xynB). These are functions previously unreported for lactococci or lactobacilli. The mutarotase activity of the putative xylM gene product was confirmed by overexpression of the L. lactis enzyme in Escherichia coli and purification of recombinant XylM. We hypothesize that the mutarotase links xylan degradation to xylose metabolism due to the anomeric preference of xylose isomerase. In addition, Northern hybridization experiments suggested that the xylM and xynTB genes are cotranscribed with the xylRAB genes, responsible for xylose metabolism. Although none of the three strains appeared to metabolize xylan or xylobiose, they exhibited xylosidase activity, and L. lactis IO-1 and L. lactis NRRL B-4449 had functional mutarotases. PMID- 11282588 TI - Chromosomal locus for cadmium resistance in Pseudomonas putida consisting of a cadmium-transporting ATPase and a MerR family response regulator. AB - Pseudomonads from environmental sources vary widely in their sensitivity to cadmium, but the basis for this resistance is largely uncharacterized. A chromosomal fragment encoding cadmium resistance was cloned from Pseudomonas putida 06909, a rhizosphere bacterium, and sequence analysis revealed two divergently transcribed genes, cadA and cadR. CadA was similar to cadmium transporting ATPases known mostly from gram-positive bacteria, and to ZntA, a lead-, zinc-, and cadmium-transporting ATPase from Escherichia coli. CadR was related to the MerR family of response regulators that normally control mercury detoxification in other bacterial systems. A related gene, zntR, regulates zntA in E. coli, but it is not contiguous with zntA in the E. coli genome as cadA and cadR were in P. putida. In addition, unlike ZntA and other CadA homologs, but similar to the predicted product of gene PA3690 in the P. aeruginosa genome, the P. putida CadA sequence had a histidine-rich N-terminal extension. CadR and the product of PA3689 of P. aeruginosa also had histidine-rich C-terminal extensions not found in other MerR family response regulators. Mutational analysis indicated that cadA and cadR are fully responsible for cadmium resistance and partially for zinc resistance. However, unlike zntA, they did not confer significant levels of lead resistance. The cadA promoter was responsive to Cd(II), Pb(II), and Zn(II), while the cadR promoter was only induced by Cd(II). CadR apparently represses its own expression at the transcriptional level. However, CadR apparently does not repress cadA. Homologs of the cadmium-transporting ATPase were detected in many other Pseudomonas species. PMID- 11282590 TI - Comparative study of the cyclization reactions of three bacterial cyclomaltodextrin glucanotransferases. AB - The actions of cyclomaltodextrin glucanotransferases (CGTase; EC 2.4.1.19) from alkalophilic Bacillus sp. strain A2-5a (A2-5a CGTase), Bacillus macerans (Bmac CGTase), and Bacillus stearothermophilus (Bste CGTase) on amylose were investigated. All three enzymes produced large cyclic alpha-1,4-glucans (cycloamyloses) at the early stage of the reaction, but these were subsequently converted into smaller cycloamyloses. However, the rates of this conversion differed among the three enzymes. The product specificity of each CGTase in the cyclization reaction was determined by measuring the amount of each cycloamylose from CD6 to CD31 (CDn, a cycloamylose with a degree of polymerization of n). A2 5a CGTase produced 10 times more CD7, while Bmac CGTase produced 34 times more CD6 than other cycloamyloses. Bste CGTase produced 12 and 3 times more CD6 and CD7 than other cycloamyloses, respectively. The substrate specificities of the linearization reactions of CD6, CD7, CD8, and larger cycloamyloses (a mixture of CD22 to CD50) were investigated, and we found that CD7 and CD8 are extremely poor substrates for both hydrolytic and transglycosidic linearization (coupling) reactions while larger cycloamyloses are linearized at a much higher rate. By repeating these cyclization and linearization reactions, the larger cycloamyloses initially produced are converted into smaller cycloamyloses and finally into mainly CD6, CD7, and CD8. These three enzymes also differ in their hydrolytic activities, which seem to accelerate the conversion of larger cycloamyloses into smaller cycloamyloses. PMID- 11282591 TI - Investigating microbial (micro)colony heterogeneity by vibrational spectroscopy. AB - Fourier transform infrared and Raman microspectroscopy are currently being developed as new methods for the rapid identification of clinically relevant microorganisms. These methods involve measuring spectra from microcolonies which have been cultured for as little as 6 h, followed by the nonsubjective identification of microorganisms through the use of multivariate statistical analyses. To examine the biological heterogeneity of microorganism growth which is reflected in the spectra, measurements were acquired from various positions within (micro)colonies cultured for 6, 12, and 24 h. The studies reveal that there is little spectral variance in 6-h microcolonies. In contrast, the 12- and 24-h cultures exhibited a significant amount of heterogeneity. Hierarchical cluster analysis of the spectra from the various positions and depths reveals the presence of different layers in the colonies. Further analysis indicates that spectra acquired from the surface of the colonies exhibit higher levels of glycogen than do the deeper layers of the colony. Additionally, the spectra from the deeper layers present with higher RNA levels than the surface layers. Therefore, the 6-h colonies with their limited heterogeneity are more suitable for inclusion in a spectral database to be used for classification purposes. These results also demonstrate that vibrational spectroscopic techniques can be useful tools for studying the nature of colony development and biofilm formation. PMID- 11282592 TI - Purification, characterization, and application of a novel dye-linked L-proline dehydrogenase from a hyperthermophilic archaeon, Thermococcus profundus. AB - The distribution of dye-linked L-amino acid dehydrogenases was investigated in several hyperthermophiles, and the activity of dye-linked L-proline dehydrogenase (dye-L-proDH, L-proline:acceptor oxidoreductase) was found in the crude extract of some Thermococcales strains. The enzyme was purified to homogeneity from a hyperthermophilic archaeon, Thermococcus profundus DSM 9503, which exhibited the highest specific activity in the crude extract. The molecular mass of the enzyme was about 160 kDa, and the enzyme consisted of heterotetrameric subunits (alpha(2) beta(2)) with two different molecular masses of about 50 and 40 kDa. The N-terminal amino acid sequences of the alpha-subunit (50-kDa subunit) and the beta-subunit (40-kDa subunit) were MRLTEHPILDFSERRGRKVTIHF and XRSEAKTVIIGGGIIGLSIAYNLAK, respectively. Dye-L-proDH was extraordinarily stable among the dye-linked dehydrogenases under various conditions: the enzyme retained its full activity upon incubation at 70 degrees C for 10 min, and ca. 40% of the activity still remained after heating at 80 degrees C for 120 min. The enzyme did not lose the activity upon incubation over a wide range of pHs from 4.0 to 10.0 at 50 degrees C for 10 min. The enzyme exclusively catalyzed L-proline dehydrogenation using 2,6-dichloroindophenol (Cl2Ind) as an electron acceptor. The Michaelis constants for L-proline and Cl2Ind were determined to be 2.05 and 0.073 mM, respectively. The reaction product was identified as Delta(1)-pyrroline 5-carboxylate by thin-layer chromatography. The prosthetic group of the enzyme was identified as flavin adenine dinucleotide by high-pressure liquid chromatography. In addition, the simple and specific determination of L-proline at concentrations from 0.10 to 2.5 mM using the stable dye-L-proDH was achieved. PMID- 11282593 TI - Degradation of phenanthrene and anthracene by cell suspensions of Mycobacterium sp. strain PYR-1. AB - Cultures of Mycobacterium sp. strain PYR-1 were dosed with anthracene or phenanthrene and after 14 days of incubation had degraded 92 and 90% of the added anthracene and phenanthrene, respectively. The metabolites were extracted and identified by UV-visible light absorption, high-pressure liquid chromatography retention times, mass spectrometry, (1)H and (13)C nuclear magnetic resonance spectrometry, and comparison to authentic compounds and literature data. Neutral pH ethyl acetate extracts from anthracene-incubated cells showed four metabolites, identified as cis-1,2-dihydroxy-1,2-dihydroanthracene, 6,7 benzocoumarin, 1-methoxy-2-hydroxyanthracene, and 9,10-anthraquinone. A novel anthracene ring fission product was isolated from acidified culture media and was identified as 3-(2-carboxyvinyl)naphthalene-2-carboxylic acid. 6,7-Benzocoumarin was also found in that extract. When Mycobacterium sp. strain PYR-1 was grown in the presence of phenanthrene, three neutral metabolites were identified as cis- and trans-9,10-dihydroxy-9,10-dihydrophenanthrene and cis-3,4-dihydroxy-3,4 dihydrophenanthrene. Phenanthrene ring fission products, isolated from acid extracts, were identified as 2,2'-diphenic acid, 1-hydroxynaphthoic acid, and phthalic acid. The data point to the existence, next to already known routes for both gram-negative and gram-positive bacteria, of alternative pathways that might be due to the presence of different dioxygenases or to a relaxed specificity of the same dioxygenase for initial attack on polycyclic aromatic hydrocarbons. PMID- 11282594 TI - Seasonal variation and indirect monitoring of microcystin concentrations in Daechung reservoir, Korea. AB - Physicochemical and biological water quality, including the microcystin concentration, was investigated from spring to autumn 1999 in the Daechung Reservoir, Korea. The dominant genus in the cyanobacterial blooming season was Microcystis. The microcystin concentration in particulate form increased dramatically from August up to a level of 200 ng liter(-1) in early October and thereafter tended to decrease. The microcystin concentration in dissolved form was about 28% of that of the particulate form. The microcystins detected using a protein phosphatase (PP) inhibition assay were highly correlated with those microcystins detected by a high-performance liquid chromatograph (r = 0.973; P < 0.01). Therefore, the effectiveness of a PP inhibition assay for microcystin detection in a high number of water samples was confirmed as easy, quick, and convenient. The microcystin concentration was highly correlated with the phytoplankton number (r = 0.650; P < 0.01) and chlorophyll-a concentration (r = 0.591; P < 0.01). When the microcystin concentration exceeded about 100 ng liter( 1), the ratio of particulate to dissolved total nitrogen (TN) or total phosphorus (TP) converged at a value of 0.6. Furthermore, the microcystin concentration was lower than 50 ng liter(-1) at a particulate N/P ratio below 8, whereas the microcystin concentration varied quite substantially from 50 to 240 ng liter(-1) at a particulate N/P ratio of >8. Therefore, it seems that the microcystin concentration in water can be estimated and indirectly monitored by analyzing the following: the phytoplankton number and chlorophyll-a concentration, the ratio of the particulate and the dissolved forms of N and P, and the particulate N/P ratio when the dominant genus is toxigenic Microcystis. PMID- 11282595 TI - Method for host-independent detection of generalized transducing bacteriophages in natural habitats. AB - Despite an increasing interest in horizontal gene transfer in bacteria, the role of generalized transduction in this process has not been well investigated yet. Certainly one of the reasons is that only a small fraction of general transducing bacteriophages have been characterized, because many bacterial hosts needed for propagation and identification are not culturable or are simply unknown. A method for host-independent detection of transducing bacteriophages was developed. Phage encapsulated DNA was used as a template for PCR amplification of 16S ribosomal DNA using primers specific for the 16S rRNA genes of most eubacteria. Sequencing of the cloned amplification products permits the identification of the host bacteria. The Salmonella phage P22 was used as an example. Applying this method to a sample of the supernatant of the mixed liquor in the aeration tank of an activated sludge treatment works revealed the presence of transducing phages infecting several bacterial species for which such phages have not yet been described. This method is suitable for estimating the contribution of generalized transduction to horizontal gene transfer in different habitats. PMID- 11282597 TI - Identification of fecal Escherichia coli from humans and animals by ribotyping. AB - Fecal pollution of water resources is an environmental problem of increasing importance. Identification of individual host sources of fecal Escherichia coli, such as humans, pets, production animals, and wild animals, is prerequisite to formulation of remediation plans. Ribotyping has been used to distinguish fecal E. coli of human origin from pooled fecal E. coli isolates of nonhuman origin. We have extended application of this technique to distinguishing fecal E. coli ribotype patterns from human and seven individual nonhuman hosts. Classification accuracy was best when the analysis was limited to three host sources. Application of this technique to identification of host sources of fecal coliforms in water could assist in formulation of pollution reduction plans. PMID- 11282596 TI - Occurrence and diversity of tetracycline resistance genes in lagoons and groundwater underlying two swine production facilities. AB - In this study, we used PCR typing methods to assess the presence of tetracycline resistance determinants conferring ribosomal protection in waste lagoons and in groundwater underlying two swine farms. All eight classes of genes encoding this mechanism of resistance [tet(O), tet(Q), tet(W), tet(M), tetB(P), tet(S), tet(T), and otrA] were found in total DNA extracted from water of two lagoons. These determinants were found to be seeping into the underlying groundwater and could be detected as far as 250 m downstream from the lagoons. The identities and origin of these genes in groundwater were confirmed by PCR-denaturing gradient gel electrophoresis and sequence analyses. Tetracycline-resistant bacterial isolates from groundwater harbored the tet(M) gene, which was not predominant in the environmental samples and was identical to tet(M) from the lagoons. The presence of this gene in some typical soil inhabitants suggests that the vector of antibiotic resistance gene dissemination is not limited to strains of gastrointestinal origin carrying the gene but can be mobilized into the indigenous soil microbiota. This study demonstrated that tet genes occur in the environment as a direct result of agriculture and suggested that groundwater may be a potential source of antibiotic resistance in the food chain. PMID- 11282598 TI - Map of the IncP1beta plasmid pTSA encoding the widespread genes (tsa) for p toluenesulfonate degradation in Comamonas testosteroni T-2. AB - The catabolic IncP1beta plasmid pTSA from Comamonas testosteroni T-2 was mapped by subtractive analysis of restriction digests, by sequencing outwards from the tsa operon (toluenesulfonate degradation), and by generating overlapping, long distance-PCR amplification products. The plasmid was estimated to comprise 72 +/- 4 kb. The tsa region was found to be a composite transposon flanked by two IS1071 elements. A cryptic tsa operon was also present in the tsa transposon. Those backbone genes and regions which we sequenced were in the same order as the corresponding genes in resistance plasmid R751, and identities of about 99% were observed. Enrichment cultures with samples from four continents were done to obtain organisms able to utilize p-toluenesulfonate as the sole source of carbon and energy for aerobic growth. Most (15) of the 16 cultures (13 of them isolates) were obtained from contaminated sites and were attributed to three metabolic groups, depending on their metabolism of p-toluenesulfonate. The largest group contained the tsa transposon, usually (six of seven isolates) with negligible differences in sequence from strain T-2. PMID- 11282599 TI - Relationship of hydrogen bioavailability to chromate reduction in aquifer sediments. AB - Biological Cr(VI) reduction was studied in anaerobic sediments from an aquifer in Norman, Okla. Microcosms containing sediment and mineral medium were amended with various electron donors to determine those most important for biological Cr(VI) reduction. Cr(VI) (about 340 microM) was reduced with endogenous substrates (no donor), or acetate was added. The addition of formate, hydrogen, and glucose stimulated Cr(VI) reduction compared with reduction in unamended controls. From these sediments, an anaerobic Cr(VI)-utilizing enrichment was obtained that was dependent upon hydrogen for both growth and Cr(VI) reduction. No methane was produced by the enrichment, which reduced about 750 microM Cr(VI) in less than six days. The dissolved hydrogen concentration was used as an indicator of the terminal electron accepting process occurring in the sediments. Microcosms with sediments, groundwater, and chromate metabolized hydrogen to a concentration below the detection limits of the mercury vapor gas chromatograph. In microcosms without chromate, the hydrogen concentration was about 8 nM, a concentration comparable to that under methanogenic conditions. When these microcosms were amended with 500 microM Cr(VI), the dissolved hydrogen concentration quickly fell below the detection limits. These results showed that the hydrogen concentration under chromate-reducing conditions became very low, as low as that reported under nitrate- and manganese-reducing conditions, a result consistent with the free energy changes for these reactions. The utilization of formate, lactate, hydrogen, and glucose as electron donors for Cr(VI) reduction indicates that increasing the availability of hydrogen results in a greater capacity for Cr(VI) reduction. This conclusion is supported by the existence of an enrichment dependent upon hydrogen for growth and Cr(VI) reduction. PMID- 11282600 TI - Characterization of a novel type II restriction-modification system, Sth368I, encoded by the integrative element ICESt1 of Streptococcus thermophilus CNRZ368. AB - A novel type II restriction and modification (R-M) system, Sth368I, which confers resistance to phiST84, was found in Streptococcus thermophilus CNRZ368 but not in the very closely related strain A054. Partial sequencing of the integrative conjugative element ICESt1, carried by S. thermophilus CNRZ368 but not by A054, revealed a divergent cluster of two genes, sth368IR and sth368IM. The protein sequence encoded by sth368IR is related to the type II endonucleases R.LlaKR2I and R.Sau3AI, which recognize and cleave the sequence 5'-GATC-3'. The protein sequence encoded by sth368IM is very similar to numerous type II 5-methylcytosine methyltransferases, including M.LlaKR2I and M.Sau3AI. Cell extracts of CNRZ368 but not A054 were found to cleave at the GATC site. Furthermore, the C residue of the sequence 5'-GATC-3' was found to be methylated in CNRZ368 but not in A054. Cloning and integration of a copy of sth368IR and sth368IM in the A054 chromosome confers on this strain phenotypes similar to those of CNRZ368, i.e., phage resistance, endonuclease activity of cell extracts, and methylation of the sequence 5'-GATC-3'. Disruption of sth368IR removes resistance and restriction activity. We conclude that ICESt1 encodes an R-M system, Sth368I, which recognizes the sequence 5'-GATC-3' and is related to the Sau3AI and LlaKR2I restriction systems. PMID- 11282601 TI - Cold-adapted beta-galactosidase from the Antarctic psychrophile Pseudoalteromonas haloplanktis. AB - The beta-galactosidase from the Antarctic gram-negative bacterium Pseudoalteromonas haloplanktis TAE 79 was purified to homogeneity. The nucleotide sequence and the NH(2)-terminal amino acid sequence of the purified enzyme indicate that the beta-galactosidase subunit is composed of 1,038 amino acids with a calculated M(r) of 118,068. This beta-galactosidase shares structural properties with Escherichia coli beta-galactosidase (comparable subunit mass, 51% amino sequence identity, conservation of amino acid residues involved in catalysis, similar optimal pH value, and requirement for divalent metal ions) but is characterized by a higher catalytic efficiency on synthetic and natural substrates and by a shift of apparent optimum activity toward low temperatures and lower thermal stability. The enzyme also differs by a higher pI (7.8) and by specific thermodynamic activation parameters. P. haloplanktis beta-galactosidase was expressed in E. coli, and the recombinant enzyme displays properties identical to those of the wild-type enzyme. Heat-induced unfolding monitored by intrinsic fluorescence spectroscopy showed lower melting point values for both P. haloplanktis wild-type and recombinant beta-galactosidase compared to the mesophilic enzyme. Assays of lactose hydrolysis in milk demonstrate that P. haloplanktis beta-galactosidase can outperform the current commercial beta galactosidase from Kluyveromyces marxianus var. lactis, suggesting that the cold adapted beta-galactosidase could be used to hydrolyze lactose in dairy products processed in refrigerated plants. PMID- 11282602 TI - Proline-rich peptide from the coral pathogen Vibrio shiloi that inhibits photosynthesis of Zooxanthellae. AB - The coral-bleaching bacterium Vibrio shiloi biosynthesizes and secretes an extracellular peptide, referred to as toxin P, which inhibits photosynthesis of coral symbiotic algae (zooxanthellae). Toxin P was produced during the stationary phase when the bacterium was grown on peptone or Casamino Acids media at 29 degrees C. Glycerol inhibited the production of toxin P. Toxin P was purified to homogeneity, yielding the following 12-residue peptide: PYPVYAPPPVVP (molecular weight, 1,295.54). The structure of toxin P was confirmed by chemical synthesis. In the presence of 12.5 mM NH(4)Cl, pure natural or synthetic toxin P (10 microM) caused a 64% decrease in the photosynthetic quantum yield of zooxanthellae within 5 min. The inhibition was proportional to the toxin P concentration. Toxin P bound avidly to zooxanthellae, such that subsequent addition of NH(4)Cl resulted in rapid inhibition of photosynthesis. When zooxanthellae were incubated in the presence of NH(4)Cl and toxin P, there was a rapid decrease in the pH (pH 7.8 to 7.2) of the bulk liquid, suggesting that toxin P facilitates transport of NH(3) into the cell. It is known that uptake of NH(3) into cells can destroy the pH gradient and block photosynthesis. This mode of action of toxin P can help explain the mechanism of coral bleaching by V. shiloi. PMID- 11282603 TI - Succession of phenotypic, genotypic, and metabolic community characteristics during in vitro bioslurry treatment of polycyclic aromatic hydrocarbon contaminated sediments. AB - Dredged harbor sediment contaminated with polycyclic aromatic hydrocarbons (PAHs) was removed from the Milwaukee Confined Disposal Facility and examined for in situ biodegradative capacity. Molecular techniques were used to determine the successional characteristics of the indigenous microbiota during a 4-month bioslurry evaluation. Ester-linked phospholipid fatty acids (PLFA), multiplex PCR of targeted genes, and radiorespirometry techniques were used to define in situ microbial phenotypic, genotypic, and metabolic responses, respectively. Soxhlet extractions revealed a loss in total PAH concentrations of 52%. Individual PAHs showed reductions as great as 75% (i.e., acenapthene and fluorene). Rates of (14)C-PAH mineralization (percent/day) were greatest for phenanthrene, followed by pyrene and then chrysene. There was no mineralization capacity for benzo[a]pyrene. Ester-linked phospholipid fatty acid analysis revealed a threefold increase in total microbial biomass and a dynamic microbial community composition that showed a strong correlation with observed changes in the PAH chemistry (canonical r(2) of 0.999). Nucleic acid analyses showed copies of genes encoding PAH-degrading enzymes (extradiol dioxygenases, hydroxylases, and meta cleavage enzymes) to increase by as much as 4 orders of magnitude. Shifts in gene copy numbers showed strong correlations with shifts in specific subsets of the extant microbial community. Specifically, declines in the concentrations of three ring PAH moieties (i.e., phenanthrene) correlated with PLFA indicative of certain gram-negative bacteria (i.e., Rhodococcus spp. and/or actinomycetes) and genes encoding for naphthalene-, biphenyl-, and catechol-2,3-dioxygenase degradative enzymes. The results of this study suggest that the intrinsic biodegradative potential of an environmental site can be derived from the polyphasic characterization of the in situ microbial community. PMID- 11282604 TI - Biotransformation of biphenyl by Paecilomyces lilacinus and characterization of ring cleavage products. AB - We examined the pathway by which the fungicide biphenyl is metabolized in the imperfect fungus Paecilomyces lilacinus. The initial oxidation yielded the three monohydroxylated biphenyls. Further hydroxylation occurred on the first and the second aromatic ring systems, resulting in the formation of five di- and trihydroxylated metabolites. The fungus could cleave the aromatic structures, resulting in the transformation of biphenyl via ortho-substituted dihydroxybiphenyl to six-ring fission products. All compounds were characterized by gas chromatography-mass spectroscopy and proton nuclear magnetic resonance spectroscopy. These compounds include 2-hydroxy-4-phenylmuconic acid and 2 hydroxy-4-(4'-hydroxyphenyl)-muconic acid, which were produced from 3,4 dihydroxybiphenyl and further transformed to the corresponding lactones 4-phenyl 2-pyrone-6-carboxylic acid and 4-(4'-hydroxyphenyl)-2-pyrone-6-carboxylic acid, which accumulated in large amounts. Two additional ring cleavage products were identified as (5-oxo-3-phenyl-2,5-dihydrofuran-2-yl)-acetic acid and [5-oxo-3-(4' hydroxyphenyl)-2,5-dihydrofuran-2-yl]-acetic acid. We found that P. lilacinus has a high transformation capacity for biphenyl, which could explain this organism's tolerance to this fungicide. PMID- 11282605 TI - Identification and characterization of integron-mediated antibiotic resistance among Shiga toxin-producing Escherichia coli isolates. AB - A total of 50 isolates of Shiga toxin-producing Escherichia coli (STEC), including 29 O157:H7 and 21 non-O157 STEC strains, were analyzed for antimicrobial susceptibilities and the presence of class 1 integrons. Seventy eight (n = 39) percent of the isolates exhibited resistance to two or more antimicrobial classes. Multiple resistance to streptomycin, sulfamethoxazole, and tetracycline was most often observed. Class 1 integrons were identified among nine STEC isolates, including serotypes O157:H7, O111:H11, O111:H8, O111:NM, O103:H2, O45:H2, O26:H11, and O5:NM. The majority of the amplified integron fragments were 1 kb in size with the exception of one E. coli O111:H8 isolate which possessed a 2-kb amplicon. DNA sequence analysis revealed that the integrons identified within the O111:H11, O111:NM, O45:H2, and O26:H11 isolates contained the aadA gene encoding resistance to streptomycin and spectinomycin. Integrons identified among the O157:H7 and O103:H2 isolates also possessed a similar aadA gene. However, DNA sequencing revealed only 86 and 88% homology, respectively. The 2-kb integron of the E. coli O111:H8 isolate contained three genes, dfrXII, aadA2, and a gene of unknown function, orfF, which were 86, 100, and 100% homologous, respectively, to previously reported gene cassettes identified in integrons found in Citrobacter freundii and Klebsiella pneumoniae. Furthermore, integrons identified among the O157:H7 and O111:NM strains were transferable via conjugation to another strain of E. coli O157:H7 and to several strains of Hafnia alvei. To our knowledge, this is the first report of integrons and antibiotic resistance gene cassettes in STEC, in particular E. coli O157:H7. PMID- 11282606 TI - Bacterial diversity and community structure in an aerated lagoon revealed by ribosomal intergenic spacer analyses and 16S ribosomal DNA sequencing. AB - We investigated the bacterial community structure in an aerated plug-flow lagoon treating pulp and paper mill effluent. For this investigation, we developed a composite method based on analyses of PCR amplicons containing the ribosomal intergenic spacer (RIS) and its flanking partial 16S rRNA gene. Community percent similarity was determined on the basis of RIS length polymorphism. A community succession was evident in the lagoon, indicated by a progressive community transition through seven sample locations. The most abrupt changes in community structure were associated with a temperature change from 39 to 35 degrees C and with increases in dissolved oxygen. The temporal differences in community structure, based on summer and winter samplings, were greater than the spatial differences during either season. Clone libraries of rDNA-RIS amplicons were constructed from each of three summer samples. Among 90 clones analyzed (30 clones from each sample), 56 phylotypes were distinguished by restriction fragment length polymorphism. Indices of phylotype richness, evenness, and diversity all increased in clone libraries from the beginning to the end of the lagoon. A representative clone of each phylotype was phylogenetically analyzed on the basis of its partial 16S rRNA gene sequence (ca. 450 bp). Phylogenetic analysis confirmed the increase in diversity and further indicated increasing richness of bacterial divisions. Pioneers in the community spatial succession appeared to include thermotolerant, microaerophilic methanol-oxidizing bacteria related to the genus Methylobacillus, as well as thermotolerant, microaerophilic nitrogen-fixing bacteria related to the genus Azospirillum. PMID- 11282607 TI - Quantification of trichothecene-producing Fusarium species in harvested grain by competitive PCR to determine efficacies of fungicides against Fusarium head blight of winter wheat. AB - We developed a PCR-based assay to quantify trichothecene-producing Fusarium based on primers derived from the trichodiene synthase gene (Tri5). The primers were tested against a range of fusarium head blight (FHB) (also known as scab) pathogens and found to amplify specifically a 260-bp product from 25 isolates belonging to six trichothecene-producing Fusarium species. Amounts of the trichothecene-producing Fusarium and the trichothecene mycotoxin deoxynivalenol (DON) in harvested grain from a field trial designed to test the efficacies of the fungicides metconazole, azoxystrobin, and tebuconazole to control FHB were quantified. No correlation was found between FHB severity and DON in harvested grain, but a good correlation existed between the amount of trichothecene producing Fusarium and DON present within grain. Azoxystrobin did not affect levels of trichothecene-producing Fusarium compared with those of untreated controls. Metconazole and tebuconazole significantly reduced the amount of trichothecene-producing Fusarium in harvested grain. We hypothesize that the fungicides affected the relationship between FHB severity and the amount of DON in harvested grain by altering the proportion of trichothecene-producing Fusarium within the FHB disease complex and not by altering the rate of DON production. The Tri5 quantitative PCR assay will aid research directed towards reducing amounts of trichothecene mycotoxins in food and animal feed. PMID- 11282608 TI - Genomic relatedness within five common Finnish Campylobacter jejuni pulsed-field gel electrophoresis genotypes studied by amplified fragment length polymorphism analysis, ribotyping, and serotyping. AB - Thirty-five Finnish Campylobacter jejuni strains with five SmaI/SacII pulsed field gel electrophoresis (PFGE) genotypes selected among human and chicken isolates from 1997 and 1998 were used for comparison of their PFGE patterns, amplified fragment length polymorphism (AFLP) patterns, HaeIII ribotypes, and heat-stable (HS) serotypes. The discriminatory power of PFGE, AFLP, and ribotyping with HaeIII were shown to be at the same level for this selected set of strains, and these methods assigned the strains into the same groups. The PFGE and AFLP patterns within a genotype were highly similar, indicating genetic relatedness. The same HS serotypes were distributed among different genotypes, and different serotypes were identified within one genotype. HS serotype 12 was only associated with the combined genotype G1 (PFGE-AFLP-ribotype). These studies using polyphasic genotyping methods suggested that common Finnish C. jejuni genotypes form genetic lineages which colonize both humans and chickens. PMID- 11282609 TI - Physiological properties of Saccharomyces cerevisiae from which hexokinase II has been deleted. AB - Hexokinase II is an enzyme central to glucose metabolism and glucose repression in the yeast Saccharomyces cerevisiae. Deletion of HXK2, the gene which encodes hexokinase II, dramatically changed the physiology of S. cerevisiae. The hxk2 null mutant strain displayed fully oxidative growth at high glucose concentrations in early exponential batch cultures, resulting in an initial absence of fermentative products such as ethanol, a postponed and shortened diauxic shift, and higher biomass yields. Several intracellular changes were associated with the deletion of hexokinase II. The hxk2 mutant had a higher mitochondrial H(+)-ATPase activity and a lower pyruvate decarboxylase activity, which coincided with an intracellular accumulation of pyruvate in the hxk2 mutant. The concentrations of adenine nucleotides, glucose-6-phosphate, and fructose-6-phosphate are comparable in the wild type and the hxk2 mutant. In contrast, the concentration of fructose-1,6-bisphosphate, an allosteric activator of pyruvate kinase, is clearly lower in the hxk2 mutant than in the wild type. The results suggest a redirection of carbon flux in the hxk2 mutant to the production of biomass as a consequence of reduced glucose repression. PMID- 11282610 TI - Sensitization of Listeria monocytogenes to low pH, organic acids, and osmotic stress by ethanol. AB - The killing of Listeria monocytogenes following exposure to low pH, organic acids, and osmotic stress was enhanced by the addition of 5% (vol/vol) ethanol. At pH 3, for example, the presence of this agent stimulated killing by more than 3 log units in 40 min of exposure. The rate of cell death at pH 3.0 was dependent on the concentration of ethanol. Thus, while the presence 10% (vol/vol) ethanol at pH 3.0 stimulated killing by more than 3 log units in just 5 min, addition of 1.25% (vol/vol) ethanol resulted in less than 1 log unit of killing in 10 min. The ability of 5% (vol/vol) ethanol to stimulate killing at low pH and at elevated osmolarity was also dependent on the amplitude of the imposed stress, and an increase in the pH from 3.0 to 4.0 or a decrease in the sodium chloride concentration from 25 to 2.5% led to a marked reduction in the effectiveness of 5% (vol/vol) ethanol as an augmentative agent. Combinations of organic acids, low pH, and ethanol proved to be particularly effective bactericidal treatments; the most potent combination was pH 3.0, 50 mM formate, and 5 % (vol/vol) ethanol, which resulted in 5 log units of killing in just 4 min. Ethanol-enhanced killing correlated with damage to the bacterial cytoplasmic membrane. PMID- 11282611 TI - Purification and characterization of the recombinant Thermus sp. strain T2 alpha galactosidase expressed in Escherichia coli. AB - The nucleotide sequence of the Thermus sp. strain T2 DNA coding for a thermostable alpha-galactosidase was determined. The deduced amino acid sequence of the enzyme predicts a polypeptide of 474 amino acids (M(r), 53,514). The observed homology between the deduced amino acid sequences of the enzyme and alpha-galactosidase from Thermus brockianus was over 70%. Thermus sp. strain T2 alpha-galactosidase was expressed in its active form in Escherichia coli and purified. Native polyacrylamide gel electrophoresis and gel filtration chromatography data suggest that the enzyme is octameric. The enzyme was most active at 75 degrees C for p-nitrophenyl-alpha-D-galactopyranoside hydrolysis, and it retained 50% of its initial activity after 1 h of incubation at 70 degrees C. The enzyme was extremely stable over a broad range of pH (pH 6 to 13) after treatment at 40 degrees C for 1 h. The enzyme acted on the terminal alpha galactosyl residue, not on the side chain residue, of the galactomanno oligosaccharides as well as those of yeasts and Mortierella vinacea alpha galactosidase I. The enzyme has only one Cys residue in the molecule. para Chloromercuribenzoic acid completely inhibited the enzyme but did not affect the mutant enzyme which contained Ala instead of Cys, indicating that this Cys residue is not responsible for its catalytic function. PMID- 11282612 TI - Regulation of fumonisin B(1) biosynthesis and conidiation in Fusarium verticillioides by a cyclin-like (C-type) gene, FCC1. AB - Fumonisins are a group of mycotoxins produced in corn kernels by the plant pathogenic fungus Fusarium verticillioides. A mutant of the fungus, FT536, carrying a disrupted gene named FCC1 (for Fusarium cyclin C1) resulting in altered fumonisin B(1) biosynthesis was generated. FCC1 contains an open reading frame of 1,018 bp, with one intron, and encodes a putative 319-amino-acid polypeptide. This protein is similar to UME3 (also called SRB11 or SSN8), a cyclin C of Saccharomyces cerevisiae, and contains three conserved motifs: a cyclin box, a PEST-rich region, and a destruction box. Also similar to the case for C-type cyclins, FCC1 was constitutively expressed during growth. When strain FT536 was grown on corn kernels or on defined minimal medium at pH 6, conidiation was reduced and FUM5, the polyketide synthase gene involved in fumonisin B(1) biosynthesis, was not expressed. However, when the mutant was grown on a defined minimal medium at pH 3, conidiation was restored, and the blocks in expression of FUM5 and fumonisin B(1) production were suppressed. Our data suggest that FCC1 plays an important role in signal transduction regulating secondary metabolism (fumonisin biosynthesis) and fungal development (conidiation) in F. verticillioides. PMID- 11282613 TI - Development and application of a most-probable-number-pcr assay to quantify flagellate populations in soil samples. AB - This paper reports on the first successful molecular detection and quantification of soil protozoa. Quantification of heterotrophic flagellates and naked amoebae in soil has traditionally relied on dilution culturing techniques, followed by most-probable-number (MPN) calculations. Such methods are biased by differences in the culturability of soil protozoa and are unable to quantify specific taxonomic groups, and the results are highly dependent on the choice of media and the skills of the microscopists. Successful detection of protozoa in soil by DNA techniques requires (i) the development and validation of DNA extraction and quantification protocols and (ii) the collection of sufficient sequence data to find specific protozoan 18S ribosomal DNA sequences. This paper describes the development of an MPN-PCR assay for detection of the common soil flagellate Heteromita globosa, using primers targeting a 700-bp sequence of the small subunit rRNA gene. The method was tested by use of gnotobiotic laboratory microcosms with sterile tar-contaminated soil inoculated with the bacterium Pseudomonas putida OUS82 UCB55 as prey. There was satisfactory overall agreement between H. globosa population estimates obtained by the PCR assay and a conventional MPN assay in the three soils tested. PMID- 11282614 TI - Prevalence, antibiotic susceptibility, and diversity of Escherichia coli O157:H7 isolates from a longitudinal study of beef cattle feedlots. AB - Prevalence, antibiotic susceptibility, and genetic diversity were determined for Escherichia coli O157:H7 isolated over 11 months from four beef cattle feedlots in southwest Kansas. From the fecal pat (17,050) and environmental (7,134) samples collected, 57 isolates of E. coli O157:H7 were identified by use of bacterial culture and latex agglutination (C/LA). PCR showed that 26 isolates were eaeA gene positive. Escherichia coli O157:H7 was identified in at least one of the four feedlots in 14 of the 16 collections by C/LA and in 9 of 16 collections by PCR, but consecutive positive collections at a single feedlot were rare. Overall prevalence in fecal pat samples was low (0.26% by C/LA, and 0.08% by PCR). No detectable differences in prevalence or antibiotic resistance were found between isolates collected from home pens and those from hospital pens, where antibiotic use is high. Resistant isolates were found for six of the eight antibiotics that could be used to treat E. coli infections in food animals, but few isolates were multidrug resistant. The high diversity of isolates as measured by random amplification of polymorphic DNA and other characteristics indicates that the majority of isolates were unique and did not persist at a feedlot, but probably originated from incoming cattle. The most surprising finding was the low frequency of virulence markers among E. coli isolates identified initially by C/LA as E. coli O157:H7. These results demonstrate that better ways of screening and confirming E. coli O157:H7 isolates are required for accurate determination of prevalence. PMID- 11282615 TI - Molecular screening of Enterococcus virulence determinants and potential for genetic exchange between food and medical isolates. AB - Enterococci are used as starter and probiotic cultures in foods, and they occur as natural food contaminants. The genus Enterococcus is of increased significance as a cause of nosocomial infections, and this trend is exacerbated by the development of antibiotic resistance. In this study, we investigated the incidence of known virulence determinants in starter, food, and medical strains of Enterococcus faecalis, E. faecium, and E. durans. PCR and gene probe strategies were used to screen enterococcal isolates from both food and medical sources. Different and distinct patterns of incidence of virulence determinants were found for the E. faecalis and E. faecium strains. Medical E. faecalis strains had more virulence determinants than did food strains, which, in turn, had more than did starter strains. All of the E. faecalis strains tested possessed multiple determinants (between 6 and 11). E. faecium strains were generally free of virulence determinants, with notable exceptions. Significantly, esp and gelE determinants were identified in E. faecium medical strains. These virulence determinants have not previously been identified in E. faecium strains and may result from regional differences or the evolution of pathogenic E. faecium. Phenotypic testing revealed the existence of apparently silent gelE and cyl genes. In E. faecalis, the trend in these silent genes mirrors that of the expressed determinants. The potential for starter strains to acquire virulence determinants by natural conjugation mechanisms was investigated. Transconjugation in which starter strains acquired additional virulence determinants from medical strains was demonstrated. In addition, multiple pheromone-encoding genes were identified in both food and starter strains, indicating their potential to acquire other sex pheromone plasmids. These results suggest that the use of Enterococcus spp. in foods requires careful safety evaluation. PMID- 11282616 TI - Determination of DNA content of aquatic bacteria by flow cytometry. AB - The distribution of DNA among bacterioplankton and bacterial isolates was determined by flow cytometry of DAPI (4',6'-diamidino-2-phenylindole)-stained organisms. Conditions were optimized to minimize error from nonspecific staining, AT bias, DNA packing, changes in ionic strength, and differences in cell permeability. The sensitivity was sufficient to characterize the small 1- to 2-Mb genome organisms in freshwater and seawater, as well as low-DNA cells ("dims"). The dims could be formed from laboratory cultivars; their apparent DNA content was 0.1 Mb and similar to that of many particles in seawater. Preservation with formaldehyde stabilized samples until analysis. Further permeabilization with Triton X-100 facilitated the penetration of stain into stain-resistant lithotrophs. The amount of DNA per cell determined by flow cytometry agreed with mean values obtained from spectrophotometric analyses of cultures. Correction for the DNA AT bias of the stain was made for bacterial isolates with known G+C contents. The number of chromosome copies per cell was determined with pure cultures, which allowed growth rate analyses based on cell cycle theory. The chromosome ratio was empirically related to the rate of growth, and the rate of growth was related to nutrient concentration through specific affinity theory to obtain a probe for nutrient kinetics. The chromosome size of a Marinobacter arcticus isolate was determined to be 3.0 Mb by this method. In a typical seawater sample the distribution of bacterial DNA revealed two major populations based on DNA content that were not necessarily similar to populations determined by using other stains or protocols. A mean value of 2.5 fg of DNA cell(-1) was obtained for a typical seawater sample, and 90% of the population contained more than 1.1 fg of DNA cell(-1). PMID- 11282617 TI - Biogeochemical and molecular signatures of anaerobic methane oxidation in a marine sediment. AB - Anaerobic methane oxidation was investigated in 6-m-long cores of marine sediment from Aarhus Bay, Denmark. Measured concentration profiles for methane and sulfate, as well as in situ rates determined with isotope tracers, indicated that there was a narrow zone of anaerobic methane oxidation about 150 cm below the sediment surface. Methane could account for 52% of the electron donor requirement for the peak sulfate reduction rate detected in the sulfate-methane transition zone. Molecular signatures of organisms present in the transition zone were detected by using selective PCR primers for sulfate-reducing bacteria and for Archaea. One primer pair amplified the dissimilatory sulfite reductase (DSR) gene of sulfate-reducing bacteria, whereas another primer (ANME) was designed to amplify archaeal sequences found in a recent study of sediments from the Eel River Basin, as these bacteria have been suggested to be anaerobic methane oxidizers (K. U. Hinrichs, J. M. Hayes, S. P. Sylva, P. G. Brewer, and E. F. DeLong, Nature 398:802-805, 1999). Amplification with the primer pairs produced more amplificate of both target genes with samples from the sulfate-methane transition zone than with samples from the surrounding sediment. Phylogenetic analysis of the DSR gene sequences retrieved from the transition zone revealed that they all belonged to a novel deeply branching lineage of diverse DSR gene sequences not related to any previously described DSR gene sequence. In contrast, DSR gene sequences found in the top sediment were related to environmental sequences from other estuarine sediments and to sequences of members of the genera Desulfonema, Desulfococcus, and Desulfosarcina. Phylogenetic analysis of 16S rRNA sequences obtained with the primers targeting the archaeal group of possible anaerobic methane oxidizers revealed two clusters of ANME sequences, both of which were affiliated with sequences from the Eel River Basin. PMID- 11282618 TI - Growth and arginine metabolism of the wine lactic acid bacteria Lactobacillus buchneri and Oenococcus oeni at different pH values and arginine concentrations. AB - During malolactic fermentation (MLF) in grape must and wine, heterofermentative lactic acid bacteria may degrade arginine, leading to the formation of ammonia and citrulline, among other substances. This is of concern because ammonia increases the pH and thus the risk of growth by spoilage bacteria, and citrulline is a precursor to the formation of carcinogenic ethyl carbamate (EC). Arginine metabolism and growth of Lactobacillus buchneri CUC-3 and Oenococcus oeni strains MCW and Lo111 in wine were investigated. In contrast to L. buchneri CUC-3, both oenococci required a higher minimum pH for arginine degradation, and arginine utilization was delayed relative to the degradation of malic acid, the main aim of MLF. This allows the control of pH increase and citrulline formation from arginine metabolism by carrying out MLF with pure oenococcal cultures and inhibiting cell metabolism after malic acid depletion. MLF by arginine-degrading lactobacilli should be discouraged because arginine degradation may lead to the enhanced formation of acids from sugar degradation. A linear relationship was found between arginine degradation and citrulline excretion rates. From this data, strain-specific arginine-to-citrulline conversion ratios were calculated that ranged between 2.2 and 3.9% (wt/wt), and these ratios can be used to estimate the contribution of citrulline to the EC precursor pool from a given amount of initial arginine. Increasing arginine concentrations led to higher rates of growth of L. buchneri CUC-3 but did not increase the growth yield of either oenococcus. These results suggest the use of non-arginine-degrading oenococci for inducing MLF. PMID- 11282619 TI - Phylogenetic diversity of bacterial and archaeal communities in the anoxic zone of the Cariaco Basin. AB - Microbial community samples were collected from the anoxic zone of the Cariaco Basin at depths of 320, 500, and 1,310 m on a November 1996 cruise and were used to construct 16S ribosomal DNA libraries. Of 60 nonchimeric sequences in the 320 m library, 56 belonged to the epsilon subdivision of the Proteobacteria (epsilon Proteobacteria) and 53 were closely related to ectosymbionts of Rimicaris exoculata and Alvinella pompejana, which are referred to here as epsilon symbiont relatives (ESR). The 500-m library contained sequences affiliated with the fibrobacteria, the Flexibacter-Cytophaga-Bacteroides division, the division Verrucomicrobia, the division Proteobacteria, and the OP3 candidate division. The Proteobacteria included members of the gamma, delta, epsilon and new candidate subdivisions, and gamma-proteobacterial sequences were dominant (25.6%) among the proteobacterial sequences. As in the 320-m library, the majority of the epsilon proteobacteria belonged to the ESR group. The genus Fibrobacter and its relatives were the second largest group in the library (23.6%), followed by the delta proteobacteria and the epsilon-proteobacteria. The 1,310-m library had the greatest diversity; 59 nonchimeric clones in the library contained 30 unique sequences belonging to the planctomycetes, the fibrobacteria, the Flexibacter Cytophaga-Bacteroides division, the Proteobacteria, and the OP3 and OP8 candidate divisions. The proteobacteria included members of new candidate subdivisions and the beta, gamma, delta, and epsilon-subdivisions. ESR sequences were still present in the 1,310-m library but in a much lower proportion (8.5%). One archaeal sequence was present in the 500-m library (2% of all microorganisms in the library), and eight archaeal sequences were present in the 1,310-m library (13.6%). All archaeal sequences fell into two groups; two clones in the 1,310-m library belonged to the kingdom Crenarchaeota and the remaining sequences in both libraries belonged to the kingdom Euryarchaeota. The latter group appears to be related to the Eel-TA1f2 sequence, which belongs to an archaeon suggested to be able to oxidize methane anaerobically. Based on phylogenetic inferences and measurements of dark CO(2) fixation, we hypothesized that (i) the ESR are autotrophic anaerobic sulfide oxidizers, (ii) sulfate reduction and fermentative metabolism may be carried out by a large number of bacteria in the 500- and 1,310 m libraries, and (iii) members of the Euryarchaeota found in relatively large numbers in the 1,310-m library may be involved in anaerobic methane oxidation. Overall, the composition of microbial communities from the Cariaco Basin resembles the compositions of communities from several anaerobic sediments, supporting the hypothesis that the Cariaco Basin water column is similar to anaerobic sediments. PMID- 11282620 TI - Evaluation of inoculum addition to stimulate in situ bioremediation of oily sludge-contaminated soil. AB - A full-scale study evaluating an inoculum addition to stimulate in situ bioremediation of oily-sludge-contaminated soil was conducted at an oil refinery where the indigenous population of hydrocarbon-degrading bacteria in the soil was very low (10(3) to 10(4) CFU/g of soil). A feasibility study was conducted prior to the full-scale bioremediation study. In this feasibility study, out of six treatments, the application of a bacterial consortium and nutrients resulted in maximum biodegradation of total petroleum hydrocarbon (TPH) in 120 days. Therefore, this treatment was selected for the full-scale study. In the full scale study, plots A and B were treated with a bacterial consortium and nutrients, which resulted in 92.0 and 89.7% removal of TPH, respectively, in 1 year, compared to 14.0% removal of TPH in the control plot C. In plot A, the alkane fraction of TPH was reduced by 94.2%, the aromatic fraction of TPH was reduced by 91.9%, and NSO (nitrogen-, sulfur-, and oxygen-containing compound) and asphaltene fractions of TPH were reduced by 85.2% in 1 year. Similarly, in plot B the degradation of alkane, aromatic, and NSO plus asphaltene fractions of TPH was 95.1, 94.8, and 63.5%, respectively, in 345 days. However, in plot C, removal of alkane (17.3%), aromatic (12.9%), and NSO plus asphaltene (5.8%) fractions was much less. The population of introduced Acinetobacter baumannii strains in plots A and B was stable even after 1 year. Physical and chemical properties of the soil at the bioremediation site improved significantly in 1 year. PMID- 11282621 TI - Interaction of the PhiHSIC virus with its host: lysogeny or pseudolysogeny? AB - The marine phage PhiHSIC has been previously reported to enter into a lysogenic relationship with its host, HSIC, identified as Listonella pelagia. This phage produces a variety of plaques on its host, including turbid and haloed plaques, from which lysogens were previously isolated. These lysogens were unstable during long-term storage at -80( degrees ) C and were lost. When HSIC was reinfected with phage PhiHSIC, pseudolysogen-like interactions between the phage and its host were observed. The cells (termed HSIC-2 or HSIC-2e) produced high viral titers (10(11) ml(-1)) in the absence of inoculating phage and yet reached culture densities of nearly 10(9) ml(-1). Prophages were not induced by mitomycin C or the polyaromatic hydrocarbon naphthalene in cells harboring such infections. However, such cells were homoimmune to superinfection. Colonies hybridized strongly with a gene probe from a 100-bp fragment of the PhiHSIC genome, while the host did not. Analysis of chromosomal DNA preparations suggested the presence of a chromosomally integrated prophage. Phage adsorption experiments suggested that HSIC-2 was adsorption impaired. Because of the chromosomal prophage integration and homoimmunity, we interpret these results to indicate that PhiHSIC establishes a lysogenic relationship with its host that involves an extremely high level of spontaneous induction. This could be caused by a weak repressor of phage production. Additionally, poor phage adsorption of HSIC-2 compared to the wild type probably helped maintain this pseudolysogen-like relationship. In many ways, pseudolysogenic phage-host interactions may provide a paradigm for phage host interactions in the marine environment. PMID- 11282622 TI - Enterocin P selectively dissipates the membrane potential of Enterococcus faecium T136. AB - Enterocin P is a pediocin-like, broad-spectrum bacteriocin which displays a strong inhibitory activity against Listeria monocytogenes. The bacteriocin was purified from the culture supernatant of Enterococcus faecium P13, and its molecular mechanism of action against the sensitive strain E. faecium T136 was evaluated. Although enterocin P caused significant reduction of the membrane potential (DeltaPsi) and the intracellular ATP pool of the indicator organism, the pH gradient (DeltapH) component of the proton motive force (Deltap) was not dissipated. By contrast, enterocin P caused carboxyfluorescein efflux from E. faecium T136-derived liposomes. PMID- 11282623 TI - Sensitivities of germinating spores and carvacrol-adapted vegetative cells and spores of Bacillus cereus to nisin and pulsed-electric-field treatment. AB - Treatment of Bacillus cereus spores with nisin and/or pulsed-electric-field (PEF) treatment did not lead to direct inactivation of the spores or increased heat sensitivity as a result of sublethal damage. In contrast, germinating spores were found to be sensitive to PEF treatment. Nisin treatment was more efficient than PEF treatment for inactivating germinating spores. PEF resistance was lost after 50 min of germination, and not all germinated spores could be inactivated. Nisin, however, was able to inactivate the germinating spores to the same extent as heat treatment. Resistance to nisin was lost immediately when the germination process started. A decrease in the membrane fluidity of vegetative cells caused by incubation in the presence of carvacrol resulted in a dramatic increase in the sensitivity to nisin. On the other hand, inactivation by PEF treatment or by a combination of nisin and PEF treatments did not change after adaptation to carvacrol. Spores grown in the presence of carvacrol were not susceptible to nisin and/or PEF treatment in any way. PMID- 11282624 TI - Molecular characterization of a theta replication plasmid and its use for development of a two-component food-grade cloning system for Lactococcus lactis. AB - pCD4, a small, highly stable theta-replicating lactococcal plasmid, was used to develop a food-grade cloning system. Sequence analysis revealed five open reading frames and two putative cis-acting regions. None appears to code for undesirable phenotypes with regard to food applications. Functional analysis of the replication module showed that only the cis-acting ori region and the repB gene coding for the replication initiator protein were needed for the stable replication and maintenance of pCD4 derivatives in Lactococcus lactis. A two component food-grade cloning system was derived from the pCD4 replicon. The vector pVEC1, which carries the functional pCD4 replicon, is entirely made up of L. lactis DNA and has no selection marker. The companion pCOM1 is a repB deficient pCD4 derivative that carries an erythromycin resistance gene as a dominant selection marker. The pCOM1 construct can only replicate in L. lactis if trans complemented by the RepB initiator provided by pVEC1. Since only the cotransformants that carry both pVEC1 and pCOM1 can survive on plates containing erythromycin, pCOM1 can be used transiently to select cells that have acquired pVEC1. Due to the intrinsic incompatibility between these plasmids, pCOM1 can be readily cured from the cells grown on an antibiotic-free medium after the selection step. The system was used to introduce a phage resistance mechanism into the laboratory strain MG1363 of L. lactis and two industrial strains. The introduction of the antiphage barrier did not alter the wild-type plasmid profile of the industrial strains. The phenotype was stable after 100 generations and conferred an effective resistance phenotype against phages of the 936 and c2 species. PMID- 11282625 TI - Effect of exogenous siderophores on iron uptake activity of marine bacteria under iron-limited conditions. AB - More than 60% of species examined from a total of 421 strains of heterotrophic marine bacteria which were isolated from marine sponges and seawater were observed to have no detectable siderophore production even when Fe(III) was present in the culture medium at a concentration of 1.0 pM. The growth of one such non-siderophore-producing strain, alpha proteobacterium V0210, was stimulated under iron-limited conditions with the addition of an isolated exogenous siderophore, N,N'-bis (2,3-dihydroxybenzoyl)-O-serylserine from a Vibrio sp. Growth was also stimulated by the addition of three exogenous siderophore extracts from siderophore-producing bacteria. Radioisotope studies using (59)Fe showed that the iron uptake ability of V0210 increased only with the addition of exogenous siderophores. Biosynthesis of a hydroxamate siderophore by V0210 was shown by paper electrophoresis and chemical assays for the detection of hydroxamates and catechols. An 85-kDa iron-regulated outer membrane protein was induced only under iron-limited conditions in the presence of exogenous siderophores. This is the first report of bacterial iron uptake through an induced siderophore in response to exogenous siderophores. Our results suggest that siderophores are necessary signaling compounds for growth and for iron uptake by some non-siderophore-producing marine bacteria under iron-limited conditions. PMID- 11282626 TI - Characterization of fluorescent and nonfluorescent peptide siderophores produced by Pseudomonas syringae strains and their potential use in strain identification. AB - Nonfluorescent highly virulent strains of Pseudomonas syringae pv. aptata isolated in different European countries and in Uruguay produce a nonfluorescent peptide siderophore, the production of which is iron repressed and specific to these strains. The amino acid composition of this siderophore is identical to that of the dominant fluorescent peptide siderophore produced by fluorescent P. syringae strains, and the molecular masses of the respective Fe(III) chelates are 1,177 and 1,175 atomic mass units. The unchelated nonfluorescent siderophore is converted into the fluorescent siderophore at pH 10, and colors and spectral characteristics of the unchelated siderophores and of the Fe(III)-chelates in acidic conditions are similar to those of dihydropyoverdins and pyoverdins, respectively. The nonfluorescent siderophore is used by fluorescent and nonfluorescent P. syringae strains. These results and additional mass spectrometry data strongly suggest the presence of a pyoverdin chromophore in the fluorescent siderophore and a dihydropyoverdin chromophore in the nonfluorescent siderophore, which are both ligated to a succinamide residue. When chelated, the siderophores behave differently from typical pyoverdins and dihydropyoverdins in neutral and alkaline conditions, apparently because of the ionization occurring around pH 4.5 of carboxylic acids present in beta-hydroxyaspartic acid residues of the peptide chains. These differences can be detected visually by pH-dependent changes of the chelate colors and spectrophotochemically. These characteristics and the electrophoretic behavior of the unchelated and chelated siderophores offer new tools to discriminate between saprophytic fluorescent Pseudomonas species and fluorescent P. syringae and P. viridiflava strains and to distinguish between the two siderovars in P. syringae pv. aptata. PMID- 11282627 TI - Metabolism of benzoate, cyclohex-1-ene carboxylate, and cyclohexane carboxylate by "Syntrophus aciditrophicus" strain SB in syntrophic association with H(2) using microorganisms. AB - The metabolism of benzoate, cyclohex-1-ene carboxylate, and cyclohexane carboxylate by "Syntrophus aciditrophicus" in cocultures with hydrogen-using microorganisms was studied. Cyclohexane carboxylate, cyclohex-1-ene carboxylate, pimelate, and glutarate (or their coenzyme A [CoA] derivatives) transiently accumulated during growth with benzoate. Identification was based on comparison of retention times and mass spectra of trimethylsilyl derivatives to the retention times and mass spectra of authentic chemical standards. (13)C nuclear magnetic resonance spectroscopy confirmed that cyclohexane carboxylate and cyclohex-1-ene carboxylate were produced from [ring-(13)C(6)]benzoate. None of the metabolites mentioned above was detected in non-substrate-amended or heat killed controls. Cyclohexane carboxylic acid accumulated to a concentration of 260 microM, accounting for about 18% of the initial benzoate added. This compound was not detected in culture extracts of Rhodopseudomonas palustris grown phototrophically or Thauera aromatica grown under nitrate-reducing conditions. Cocultures of "S. aciditrophicus" and Methanospirillum hungatei readily metabolized cyclohexane carboxylate and cyclohex-1-ene carboxylate at a rate slightly faster than the rate of benzoate metabolism. In addition to cyclohexane carboxylate, pimelate, and glutarate, 2-hydroxycyclohexane carboxylate was detected in trace amounts in cocultures grown with cyclohex-1-ene carboxylate. Cyclohex-1-ene carboxylate, pimelate, and glutarate were detected in cocultures grown with cyclohexane carboxylate at levels similar to those found in benzoate grown cocultures. Cell extracts of "S. aciditrophicus" grown in a coculture with Desulfovibrio sp. strain G11 with benzoate or in a pure culture with crotonate contained the following enzyme activities: an ATP-dependent benzoyl-CoA ligase, cyclohex-1-ene carboxyl-CoA hydratase, and 2-hydroxycyclohexane carboxyl-CoA dehydrogenase, as well as pimelyl-CoA dehydrogenase, glutaryl-CoA dehydrogenase, and the enzymes required for conversion of crotonyl-CoA to acetate. 2 Ketocyclohexane carboxyl-CoA hydrolase activity was detected in cell extracts of "S. aciditrophicus"-Desulfovibrio sp. strain G11 benzoate-grown cocultures but not in crotonate-grown pure cultures of "S. aciditrophicus". These results are consistent with the hypothesis that ring reduction during syntrophic benzoate metabolism involves a four- or six-electron reduction step and that once cyclohex 1-ene carboxyl-CoA is made, it is metabolized in a manner similar to that in R. palustris. PMID- 11282628 TI - Duplication of a truncated paralog of the family B DNA polymerase gene Aa-polB in the Agrocybe aegerita mitochondrial genome. AB - The Agrocybe aegerita mitochondrial genome contains a truncated family B DNA polymerase gene (Aa-polB P1) whose nucleotide sequence is 86% identical to the previously described and potentially functional Aa-polB gene. A tRNA(Met) gene occurs at the 3' end of the Aa-polB P1 gene. The Aa-polB P1 gene could result from reverse transcription of an Aa-polB mRNA primed by a tRNA(Met) followed by the integration of the cDNA after recombination at the mitochondrial tRNA locus. Two naturally occurring alleles of Aa-polB P1 carry one or two copies of the disrupted sequence. In strains with two copies of Aa-polB P1, these copies are inverted relative to one another and separated by a short sequence carrying the tRNA(Met) gene. Both A. aegerita mitochondrial family B DNA polymerases were found to be related to other family B DNA polymerases (36 to 53% amino acid similarity), including the three enzymes of the archaebacterium Sulfolobus solfataricus. If mitochondria originated from a fusion between a Clostridium-like eubacterium and a Sulfolobus-like archaebacterium, then the A. aegerita family B DNA polymerase genes could be remnants of the archaebacterial genes. PMID- 11282629 TI - Novel alpha-amylase that is highly resistant to chelating reagents and chemical oxidants from the alkaliphilic Bacillus isolate KSM-K38. AB - A novel alpha-amylase (AmyK38) was found in cultures of an alkaliphilic Bacillus isolate designated KSM-K38. Based on the morphological and physiological characteristics and phylogenetic position as determined by 16S ribosomal DNA gene sequencing and DNA-DNA reassociation analysis, it was suggested that the isolate was a new species of the genus Bacillus. The enzyme had an optimal pH of 8.0 to 9.5 and displayed maximum catalytic activity at 55 to 60 degrees C. The apparent molecular mass was approximately 55 kDa, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the isoelectric point was around pH 4.2. This enzyme efficiently hydrolyzed various carbohydrates to yield maltotriose, maltohexaose, maltoheptaose, and, in addition, maltose as major end products after completion of the reaction. The activity was not prevented at all by EDTA and EGTA at concentrations as high as 100 mM. Moreover, AmyK38 was highly resistant to chemical oxidation and maintained more than 80% of its original activity even after incubation for 1 h in the presence of excess H2O2 (1.8 M). PMID- 11282630 TI - Phylogenetic diversity of ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit genes from deep-sea microorganisms. AB - The phylogenetic diversity of the ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO, E.C. 4.1.1.39) large-subunit genes of deep-sea microorganisms was analyzed. Bulk genomic DNA was isolated from seven samples, including samples from the Mid-Atlantic Ridge and various deep-sea habitats around Japan. The kinds of samples were hydrothermal vent water and chimney fragment; reducing sediments from a bathyal seep, a hadal seep, and a presumed seep; and symbiont-bearing tissues of the vent mussel, Bathymodiolus sp., and the seep vestimentiferan tubeworm, Lamellibrachia sp. The RuBisCO genes that encode both form I and form II large subunits (cbbL and cbbM) were amplified by PCR from the seven deep-sea sample DNA populations, cloned, and sequenced. From each sample, 50 cbbL clones and 50 cbbM clones, if amplified, were recovered and sequenced to group them into operational taxonomic units (OTUs). A total of 29 OTUs were recorded from the 300 total cbbL clones, and a total of 24 OTUs were recorded from the 250 total cbbM clones. All the current OTUs have the characteristic RuBisCO amino acid motif sequences that exist in other RuBisCOs. The recorded OTUs were related to different RuBisCO groups of proteobacteria, cyanobacteria, and eukarya. The diversity of the RuBisCO genes may be correlated with certain characteristics of the microbial habitats. The RuBisCO sequences from the symbiont-bearing tissues showed a phylogenetic relationship with those from the ambient bacteria. Also, the RuBisCO sequences of known species of thiobacilli and those from widely distributed marine habitats were closely related to each other. This suggests that the Thiobacillus-related RuBisCO may be distributed globally and contribute to the primary production in the deep sea. PMID- 11282631 TI - Purification and characterization of cellobiose dehydrogenase from the plant pathogen Sclerotium (Athelia) rolfsii. AB - Cellobiose dehydrogenase (CDH) is an extracellular hemoflavoenzyme produced by several wood-degrading fungi. In the presence of a suitable electron acceptor, e.g., 2,6-dichloro-indophenol (DCIP), cytochrome c, or metal ions, CDH oxidizes cellobiose to cellobionolactone. The phytopathogenic fungus Sclerotium rolfsii (teleomorph: Athelia rolfsii) strain CBS 191.62 produces remarkably high levels of CDH activity when grown on a cellulose-containing medium. Of the 7,500 U of extracellular enzyme activity formed per liter, less than 10% can be attributed to the proteolytic product cellobiose:quinone oxidoreductase. As with CDH from wood-rotting fungi, the intact, monomeric enzyme from S. rolfsii contains one heme b and one flavin adenine dinucleotide cofactor per molecule. It has a molecular size of 101 kDa, of which 15% is glycosylation, and a pI value of 4.2. The preferred substrates are cellobiose and cellooligosaccharides; additionally, beta-lactose, thiocellobiose, and xylobiose are efficiently oxidized. Cytochrome c (equine) and the azino-di-(3-ethyl-benzthiazolin-6-sulfonic acid) cation radical were the best electron acceptors, while DCIP, 1,4-benzoquinone, phenothiazine dyes such as methylene blue, phenoxazine dyes such as Meldola's blue, and ferricyanide were also excellent acceptors. In addition, electrons can be transferred to oxygen. Limited in vitro proteolysis with papain resulted in the formation of several protein fragments that are active with DCIP but not with cytochrome c. Such a flavin-containing fragment, with a mass of 75 kDa and a pI of 5.1 and lacking the heme domain, was isolated and partially characterized. PMID- 11282632 TI - Does the high nucleic acid content of individual bacterial cells allow us to discriminate between active cells and inactive cells in aquatic systems? AB - The nucleic acid contents of individual bacterial cells as determined with three different nucleic acid-specific fluorescent dyes (SYBR I, SYBR II, and SYTO 13) and flow cytometry were compared for different seawater samples. Similar fluorescence patterns were observed, and bacteria with high apparent nucleic acid contents (HNA) could be discriminated from bacteria with low nucleic acid contents (LNA). The best discrimination between HNA and LNA cells was found when cells were stained with SYBR II. Bacteria in different water samples collected from seven freshwater, brackish water, and seawater ecosystems were prelabeled with tritiated leucine and then stained with SYBR II. After labeling and staining, HNA, LNA, and total cells were sorted by flow cytometry, and the specific activity of each cellular category was determined from leucine incorporation rates. The HNA cells were responsible for most of the total bacterial production, and the specific activities of cells in the HNA population varied between samples by a factor of seven. We suggest that nucleic acid content alone can be a better indicator of the fraction of growing cells than total counts and that this approach should be combined with other fluorescent physiological probes to improve detection of the most active cells in aquatic systems. PMID- 11282633 TI - Purification, characterization, and gene cloning of purine nucleosidase from Ochrobactrum anthropi. AB - A bacterium, Ochrobactrum anthropi, produced a large amount of a nucleosidase when cultivated with purine nucleosides. The nucleosidase was purified to homogeneity. The enzyme has a molecular weight of about 170,000 and consists of four identical subunits. It specifically catalyzes the irreversible N-riboside hydrolysis of purine nucleosides, the K(m) values being 11.8 to 56.3 microM. The optimal activity temperature and pH were 50 degrees C and pH 4.5 to 6.5, respectively. Pyrimidine nucleosides, purine and pyrimidine nucleotides, NAD, NADP, and nicotinamide mononucleotide are not hydrolyzed by the enzyme. The purine nucleoside hydrolyzing activity of the enzyme was inhibited (mixed inhibition) by pyrimidine nucleosides, with K(i) and K(i)' values of 0.455 to 11.2 microM. Metal ion chelators inhibited activity, and the addition of Zn(2+) or Co(2+) restored activity. A 1.5-kb DNA fragment, which contains the open reading frame encoding the nucleosidase, was cloned, sequenced, and expressed in Escherichia coli. The deduced 363-amino-acid sequence including a 22-residue leader peptide is in agreement with the enzyme molecular mass and the amino acid sequences of NH(2)-terminal and internal peptides, and the enzyme is homologous to known nucleosidases from protozoan parasites. The amino acid residues forming the catalytic site and involved in binding with metal ions are well conserved in these nucleosidases. PMID- 11282634 TI - Tests of a cellular model for constant branch distribution in the filamentous fungus Neurospora crassa. AB - The growth of mycelial fungi is characterized by the highly polarized extension of hyphal tips and the formation of subapical branches, which themselves extend as new tips. In Neurospora crassa, tip growth and branching are crucial elements for this saprophyte in the colonization and utilization of organic substrates. Much research has focused on the mechanism of tip extension, but a cellular model that fully explains the known phenomenology of branching by N. crassa has not been proposed. We described and tested a model in which the formation of a lateral branch in N. crassa was determined by the accumulation of tip-growth vesicles caused by the excess of the rate of supply over the rate of deposition at the apex. If both rates are proportional to metabolic rate, then the model explains the known lack of dependence of branch interval on growth rate. We tested the model by manipulating the tip extension rate, first by shifting temperature in both the wild type and hyperbranching (colonial) mutants and also by observing the behavior of both tipless colonies and colonyless tips. We found that temperature shifts in either direction result in temporary changes in branching. We found that colonyless tips also pass through a temporary transition phase of branching. The tipless colonies produced a cluster of new tips near the point of damage. We also found that branching in colonial mutants is dependent on growth rate. The results of these tests are consistent with a model of branching in which branch initiation is controlled by the dynamics of tip growth while being independent of the actual rate of this growth. PMID- 11282636 TI - Pathway of propionate oxidation by a syntrophic culture of Smithella propionica and Methanospirillum hungatei. AB - The pathway of propionate conversion in a syntrophic coculture of Smithella propionica and Methanospirillum hungatei JF1 was investigated by (13)C-NMR spectroscopy. Cocultures produced acetate and butyrate from propionate. [3 (13)C]propionate was converted to [2-(13)C]acetate, with no [1-(13)C]acetate formed. Butyrate from [3-(13)C]propionate was labeled at the C2 and C4 positions in a ratio of about 1:1.5. Double-labeled propionate (2,3-(13)C) yielded not only double-labeled acetate but also single-labeled acetate at the C1 or C2 position. Most butyrate formed from [2,3-(13)C]propionate was also double labeled in either the C1 and C2 atoms or the C3 and C4 atoms in a ratio of about 1:1.5. Smaller amounts of single-labeled butyrate and other combinations were also produced. 1 (13)C-labeled propionate yielded both [1-(13)C]acetate and [2-(13)C]acetate. When (13)C-labeled bicarbonate was present, label was not incorporated into acetate, propionate, or butyrate. In each of the incubations described above, (13)C was never recovered in bicarbonate or methane. These results indicate that S. propionica does not degrade propionate via the methyl-malonyl-coenzyme A (CoA) pathway or any other of the known pathways, such as the acryloyl-CoA pathway or the reductive carboxylation pathway. Our results strongly suggest that propionate is dismutated to acetate and butyrate via a six-carbon intermediate. PMID- 11282635 TI - Characterization of Listeria monocytogenes strains involved in invasive and noninvasive listeriosis outbreaks by PCR-based fingerprinting techniques. AB - A total of 32 Listeria monocytogenes strains (16 from a recent outbreak of invasive listeriosis and 16 from two outbreaks of noninvasive listeriosis, all three occurring in Italy) were characterized by PCR-ribotyping, arbitrarily primed PCR (AP-PCR), and the recently developed infrequent-restriction-site PCR (IRS-PCR). The discriminatory ability of the techniques, first evaluated on 29 unrelated L. monocytogenes food isolates using Simpson's index of diversity, was 0.714 for PCR-ribotyping, 0.690 for AP-PCR, and 0.919 for IRS-PCR. IRS-PCR was also more capable of distinguishing among strains from the invasive listeriosis outbreak: three different clusters were identified by IRS-PCR compared to two clusters identified by both PCR-ribotyping and AP-PCR. Within each of the two outbreaks of noninvasive listeriosis, the patterns were practically identical, as demonstrated by all three techniques. Only IRS-PCR succeeded in clearly discriminating the strains related to noninvasive listeriosis from all of the other strains included in this study, including those from the outbreak of invasive listeriosis. This finding may suggest the presence of unique differences in their DNA sequences. PMID- 11282637 TI - Secretion of recombinant proteins via the chaperone/usher pathway in Escherichia coli. AB - F1 antigen (Caf1) of Yersinia pestis is assembled via the Caf1M chaperone/Caf1A usher pathway. We investigated the ability of this assembly system to facilitate secretion of full-length heterologous proteins fused to the Caf1 subunit in Escherichia coli. Despite correct processing of a chimeric protein composed of a modified Caf1 signal peptide, mature human interleukin-1beta (hIL-1beta), and mature Caf1, the processed product (hIL-1beta:Caf1) remained insoluble. Coexpression of this chimera with a functional Caf1M chaperone led to the accumulation of soluble hIL-1beta:Caf1 in the periplasm. Soluble hIL-1beta:Caf1 reacted with monoclonal antibodies directed against structural epitopes of hIL 1beta. The results indicate that Caf1M-induced release of hIL-1beta:Caf1 from the inner membrane promotes folding of the hIL-1beta domain. Similar results were obtained with the fusion of Caf1 to hIL-1beta receptor antagonist or to human granulocyte-macrophage colony-stimulating factor. Following coexpression of the hIL-1beta:Caf1 precursor with both the Caf1M chaperone and Caf1A outer membrane protein, hIL-1beta:Caf1 could be detected on the cell surface of E. coli. These results demonstrate for the first time the potential application of the chaperone/usher secretion pathway in the transport of subunits with large heterogeneous N-terminal fusions. This represents a novel means for the delivery of correctly folded heterologous proteins to the periplasm and cell surface as either polymers or cleavable monomeric domains. PMID- 11282638 TI - Purification and characterization of an X-prolyl-dipeptidyl peptidase from Lactobacillus sakei. AB - An X-prolyl-dipeptidyl peptidase has been purified from Lactobacillus sakei by ammonium sulfate fractionation and five chromatographic steps, which included hydrophobic interaction, anion-exchange chromatography, and gel filtration chromatography. This procedure resulted in a recovery yield of 7% and an increase in specificity of 737-fold. The enzyme appeared to be a dimer with a subunit molecular mass of approximately 88 kDa. Optimal activity was shown at pH 7.5 and 55 degrees C. The enzyme was inhibited by serine proteinase inhibitors and several divalent cations (Cu(2+), Hg(2+), and Zn(2+)). The enzyme almost exclusively hydrolyzed X-Pro from the N terminus of each peptide as well as fluorescent and colorimetric substrates; it also hydrolyzed X-Ala at the N terminus, albeit at lower rates. K(m) s for Gly-Pro- and Lys-Ala-7-amido-4 methylcoumarin were 29 and 88 microM, respectively; those for Gly-Pro- and Ala Pro-p-nitroanilide were 192 and 50 microM, respectively. Among peptides, beta casomorphin 1-3 was hydrolyzed at the highest rates, while the relative hydrolysis of the other tested peptides was only 1 to 12%. The potential role of the purified enzyme in the proteolytic pathway by catalyzing the hydrolysis of peptide bonds involving proline is discussed. PMID- 11282639 TI - Predictive modeling of the shelf life of fish under nonisothermal conditions. AB - The behavior of the natural microflora of Mediterannean gilt-head seabream (Sparus aurata) was monitored during aerobic storage at different isothermal conditions from 0 to 15 degrees C. The growth data of pseudomonads, established as the specific spoilage organisms of aerobically stored gilt-head seabream, combined with data from previously published experiments, were used to model the effect of temperature on pseudomonad growth using a Belehradek type model. The nominal minimum temperature parameters of the Belehradek model (T(min)) for the maximum specific growth rate (micro(max)) and the lag phase (t(Lag)) were determined to be -11.8 and -12.8 degrees C, respectively. The applicability of the model in predicting pseudomonad growth on fish at fluctuating temperatures was evaluated by comparing predictions with observed growth in experiments under dynamic conditions. Temperature scenarios designed in the laboratory and simulation of real temperature profiles observed in the fish chill chain were used. Bias and accuracy factors were used as comparison indices and ranged from 0.91 to 1.17 and from 1.11 to 1.17, respectively. The average percent difference between shelf life predicted based on pseudomonad growth and shelf life experimentally determined by sensory analysis for all temperature profiles tested was 5.8%, indicating that the model is able to predict accurately fish quality in real-world conditions. PMID- 11282640 TI - Rapid method of determining factors limiting bacterial growth in soil. AB - A technique to determine which nutrients limit bacterial growth in soil was developed. The method was based on measuring the thymidine incorporation rate of bacteria after the addition of C, N, and P in different combinations to soil samples. First, the thymidine incorporation method was tested in two different soils: an agricultural soil and a forest humus soil. Carbon (as glucose) was found to be the limiting substance for bacterial growth in both of these soils. The effect of adding different amounts of nutrients was studied, and tests were performed to determine whether the additions affected the soil pH and subsequent bacterial activity. The incubation time required to detect bacterial growth after adding substrate to the soil was also evaluated. Second, the method was used in experiments in which three different size fractions of straw (1 to 2, 0.25 to 1, and <0.25 mm) were mixed into the agricultural soil in order to induce N limitation for bacterial growth. When the straw fraction was small enough (<0.25 mm), N became the limiting nutrient for bacterial growth after about 3 weeks. After the addition of the larger straw fractions (1 to 2 and 0.25 to 1 mm), the soil bacteria were C limited throughout the incubation period (10 weeks), although an increase in the thymidine incorporation rate after the addition of C and N together compared with adding them separately was seen in the sample containing the size fraction from 0.25 to 1 mm. Third, soils from high-pH, limestone-rich areas were examined. P limitation was observed in one of these soils, while tendencies toward P limitation were seen in some of the other soils. PMID- 11282641 TI - Varied diazotrophies, morphologies, and toxicities of genetically similar isolates of Cylindrospermopsis raciborskii (nostocales, cyanophyceae) from Northern Australia. AB - The potentially toxic freshwater cyanobacterium Cylindrospermopsis raciborskii has become increasingly prevalent in tropical and temperate water bodies worldwide. This paper investigates the effects of different nitrogen sources (NO3 , NH4+, and omission of a fixed form of nitrogen) on the growth rates, morphologies, and cylindrospermopsin (CYL) concentrations (expressed as a percentage of the freeze-dried weight) of seven C. raciborskii isolates obtained from a range of water bodies in northern Australia and grown in batch culture. In general, growth rates were lowest in the absence of a fixed-nitrogen source and highest with NH4+ as the nitrogen source. Conversely, the highest concentrations of CYL were recorded in cultures grown in the absence of a fixed-nitrogen source and the lowest were found in cultures supplied with NH4+. Cultures supplied with NO3- were intermediate with respect to both CYL concentration and growth rate. Different nitrogen sources resulted in significant differences in the morphology of C. raciborskii trichomes. Most notable were the loss of heterocysts and the tapering of end cells in cultures supplied with NH4+ and the statistically significant increase in vegetative cell length (nitrogen depleted < NO3- < NH4+). The morphological changes induced by different nitrogen sources were consistent for all isolates, despite measurable differences in vegetative-cell and heterocyst dimensions among isolates. Such induced morphological variation has implications for Cylindrospermopsis taxonomy, given that distinctions between species are based on minor and overlapping differences in cell lengths and widths. The close phylogenetic association among all seven isolates was confirmed by the high level (>99.8%) of similarity of their 16S rRNA gene sequences. Another genetic technique, analysis of the HIP1 octameric-palindrome repeated sequence, showed greater heterogeneity among the isolates and appears to be a useful method for distinguishing among isolates of C. raciborskii. PMID- 11282642 TI - Isolation and characterization of a slowly milk-coagulating variant of Lactobacillus helveticus deficient in purine biosynthesis. AB - A slowly milk-coagulating variant (Fmc(-)) of Lactobacillus helveticus CRL 1062, designated S1, was isolated and characterized. Strain S1 possessed all the known essential components required to utilize casein as a nitrogen source, which include functional proteinase and peptidase activities as well as functional amino acid, di- and tripeptide, and oligopeptide transport systems. The amino acid requirements of strain S1 were similar to those of the parental strain. However, on a purine-free, chemically defined medium, the growth rate of the Fmc( ) strain was threefold lower than that of the wild-type strain. L. helveticus S1 was found to be defective in IMP dehydrogenase activity and therefore was deficient in the ability to synthesize XMP and GMP. This conclusion was further supported by the observation that the addition of guanine or xanthine to milk, a substrate poor in purine compounds, restored the Fmc(+) phenotype of L. helveticus S1. PMID- 11282643 TI - Impact of biocontrol Pseudomonas fluorescens CHA0 and a genetically modified derivative on the diversity of culturable fungi in the cucumber rhizosphere. AB - Little is known about the effects of Pseudomonas biocontrol inoculants on nontarget rhizosphere fungi. This issue was addressed using the biocontrol agent Pseudomonas fluorescens CHA0-Rif, which produces the antimicrobial polyketides 2,4-diacetylphloroglucinol (Phl) and pyoluteorin (Plt) and protects cucumber from several fungal pathogens, including Pythium spp., as well as the genetically modified derivative CHA0-Rif(pME3424). Strain CHA0-Rif(pME3424) overproduces Phl and Plt and displays improved biocontrol efficacy compared with CHA0-Rif. Cucumber was grown repeatedly in the same soil, which was left uninoculated, was inoculated with CHA0-Rif or CHA0-Rif(pME3424), or was treated with the fungicide metalaxyl (Ridomil). Treatments were applied to soil at the start of each 32-day long cucumber growth cycle, and their effects on the diversity of the rhizosphere populations of culturable fungi were assessed at the end of the first and fifth cycles. Over 11,000 colonies were studied and assigned to 105 fungal species (plus several sterile morphotypes). The most frequently isolated fungal species (mainly belonging to the genera Paecilomyces, Phialocephala, Fusarium, Gliocladium, Penicillium, Mortierella, Verticillium, Trichoderma, Staphylotrichum, Coniothyrium, Cylindrocarpon, Myrothecium, and Monocillium) were common in the four treatments, and no fungal species was totally suppressed or found exclusively following one particular treatment. However, in each of the two growth cycles studied, significant differences were found between treatments (e.g., between the control and the other treatments and/or between the two inoculation treatments) using discriminant analysis. Despite these differences in the composition and/or relative abundance of species in the fungal community, treatments had no effect on species diversity indices, and species abundance distributions fit the truncated lognormal function in most cases. In addition, the impact of treatments at the 32-day mark of either growth cycle was smaller than the effect of growing cucumber repeatedly in the same soil. PMID- 11282644 TI - Quorum-sensing genes in Pseudomonas aeruginosa biofilms: their role and expression patterns. AB - Acylated homoserine lactone molecules are used by a number of gram-negative bacteria to regulate cell density-dependent gene expression by a mechanism known as quorum sensing (QS). In Pseudomonas aeruginosa, QS or cell-to-cell signaling controls expression of a number of virulence factors, as well as biofilm differentiation. In this study, we investigated the role played by the las and rhl QS systems during the early stages of static biofilm formation when cells are adhering to a surface and forming microcolonies. These studies revealed a marked difference in biofilm formation between the PAO1 parent and the QS mutants when glucose, but not citrate, was used as the sole carbon source. To further elucidate the contribution of lasI and rhlI to biofilm maturation, we utilized fusions to unstable green fluorescent protein in concert with confocal microscopy to perform real-time temporal and spatial studies of these genes in a flowing environment. During the course of 8-day biofilm development, lasI expression was found to progressively decrease over time. Conversely, rhlI expression remained steady throughout biofilm development but occurred in a lower percentage of cells. Spatial analysis revealed that lasI and rhlI were maximally expressed in cells located at the substratum and that expression decreased with increasing biofilm height. Because QS was shown previously to be involved in biofilm differentiation, these findings have important implications for the design of biofilm prevention and eradication strategies. PMID- 11282646 TI - Novel approach for improving the productivity of antibiotic-producing strains by inducing combined resistant mutations. AB - We developed a novel approach for improving the production of antibiotic from Streptomyces coelicolor A3(2) by inducing combined drug-resistant mutations. Mutants with enhanced (1.6- to 3-fold-higher) actinorhodin production were detected at a high frequency (5 to 10%) among isolates resistant to streptomycin (Str(r)), gentamicin (Gen(r)), or rifampin (Rif(r)), which developed spontaneously on agar plates which contained one of the three drugs. Construction of double mutants (str gen and str rif) by introducing gentamicin or rifampin resistance into an str mutant resulted in further increased (1.7- to 2.5-fold higher) actinorhodin productivity. Likewise, triple mutants (str gen rif) thus constructed were found to have an even greater ability for producing the antibiotic, eventually generating a mutant able to produce 48 times more actinorhodin than the wild-type strain. Analysis of str mutants revealed that a point mutation occurred within the rpsL gene, which encodes the ribosomal protein S12. rif mutants were found to have a point mutation in the rpoB gene, which encodes the beta-subunit of RNA polymerase. Mutation points in gen mutants still remain unknown. These single, double, and triple mutants displayed in hierarchical order a remarkable increase in the production of ActII-ORF4, a pathway-specific regulatory protein, as determined by Western blotting analysis. This reflects the same hierarchical order observed for the increase in actinorhodin production. The superior ability of the triple mutants was demonstrated by physiological analyses under various cultural conditions. We conclude that by inducing combined drug-resistant mutations we can continuously increase the production of antibiotic in a stepwise manner. This new breeding approach could be especially effective for initially improving the production of antibiotics from wild-type strains. PMID- 11282645 TI - Combined use of 16S ribosomal DNA and 16S rRNA to study the bacterial community of polychlorinated biphenyl-polluted soil. AB - The bacterial diversity assessed from clone libraries prepared from rRNA (two libraries) and ribosomal DNA (rDNA) (one library) from polychlorinated biphenyl (PCB)-polluted soil has been analyzed. A good correspondence of the community composition found in the two types of library was observed. Nearly 29% of the cloned sequences in the rDNA library were identical to sequences in the rRNA libraries. More than 60% of the total cloned sequence types analyzed were grouped in phylogenetic groups (a clone group with sequence similarity higher than 97% [98% for Burkholderia and Pseudomonas-type clones]) represented in both types of libraries. Some of those phylogenetic groups, mostly represented by a single (or pair) of cloned sequence type(s), were observed in only one of the types of library. An important difference between the libraries was the lack of clones representative of the Actinobacteria in the rDNA library. The PCB-polluted soil exhibited a high bacterial diversity which included representatives of two novel lineages. The apparent abundance of bacteria affiliated to the beta-subclass of the Proteobacteria, and to the genus Burkholderia in particular, was confirmed by fluorescence in situ hybridization analysis. The possible influence on apparent diversity of low template concentrations was assessed by dilution of the RNA template prior to amplification by reverse transcription-PCR. Although differences in the composition of the two rRNA libraries obtained from high and low RNA concentrations were observed, the main components of the bacterial community were represented in both libraries, and therefore their detection was not compromised by the lower concentrations of template used in this study. PMID- 11282648 TI - Phylogenetic and morphological diversity of cyanobacteria in soil desert crusts from the Colorado plateau. AB - We compared the community structures of cyanobacteria in four biological desert crusts from Utah's Colorado Plateau developing on different substrata. We analyzed natural samples, cultures, and cyanobacterial filaments or colonies retrieved by micromanipulation from field samples using microscopy, denaturing gradient gel electrophoresis, and sequencing of 16S rRNA genes. While microscopic analyses apparently underestimated the biodiversity of thin filamentous cyanobacteria, molecular analyses failed to retrieve signals for otherwise conspicuous heterocystous cyanobacteria with thick sheaths. The diversity found in desert crusts was underrepresented in currently available nucleotide sequence databases, and several novel phylogenetic clusters could be identified. Morphotypes fitting the description of Microcoleus vaginatus Gomont, dominant in most samples, corresponded to a tight phylogenetic cluster of probable cosmopolitan distribution, which was well differentiated from other cyanobacteria traditionally classified within the same genus. A new, diverse phylogenetic cluster, named "Xeronema," grouped a series of thin filamentous Phormidium-like cyanobacteria. These were also ubiquitous in our samples and probably correspond to various botanical Phormidium and Schizothrix spp., but they are phylogenetically distant from thin filamentous cyanobacteria from other environments. Significant differences in community structure were found among soil types, indicating that soil characteristics may select for specific cyanobacteria. Gypsum crusts were most deviant from the rest, while sandy, silt, and shale crusts were relatively more similar among themselves. PMID- 11282649 TI - Genomic interspecies microarray hybridization: rapid discovery of three thousand genes in the maize endophyte, Klebsiella pneumoniae 342, by microarray hybridization with Escherichia coli K-12 open reading frames. AB - In an effort to efficiently discover genes in the diazotrophic endophyte of maize, Klebsiella pneumoniae 342, DNA from strain 342 was hybridized to a microarray containing 96% (n = 4,098) of the annotated open reading frames from Escherichia coli K-12. Using a criterion of 55% identity or greater, 3,000 (70%) of the E. coli K-12 open reading frames were also found to be present in strain 342. Approximately 24% (n = 1,030) of the E. coli K-12 open reading frames are absent in strain 342. For 1.6% (n = 68) of the open reading frames, the signal was too low to make a determination regarding the presence or absence of the gene. Genes with high identity between the two organisms are those involved in energy metabolism, amino acid metabolism, fatty acid metabolism, cofactor synthesis, cell division, DNA replication, transcription, translation, transport, and regulatory proteins. Functions that were less highly conserved included carbon compound metabolism, membrane proteins, structural proteins, putative transport proteins, cell processes such as adaptation and protection, and central intermediary metabolism. Open reading frames of E. coli K-12 with little or no identity in strain 342 included putative regulatory proteins, putative chaperones, surface structure proteins, mobility proteins, putative enzymes, hypothetical proteins, and proteins of unknown function, as well as genes presumed to have been acquired by lateral transfer from sources such as phage, plasmids, or transposons. The results were in agreement with the physiological properties of the two strains. Whole genome comparisons by genomic interspecies microarray hybridization are shown to rapidly identify thousands of genes in a previously uncharacterized bacterial genome provided that the genome of a close relative has been fully sequenced. This approach will become increasingly more useful as more full genome sequences become available. PMID- 11282647 TI - Community structure of denitrifiers, bacteria, and archaea along redox gradients in Pacific Northwest marine sediments by terminal restriction fragment length polymorphism analysis of amplified nitrite reductase (nirS) and 16S rRNA genes. AB - Steep vertical gradients of oxidants (O(2) and NO(3)(-)) in Puget Sound and Washington continental margin sediments indicate that aerobic respiration and denitrification occur within the top few millimeters to centimeters. To systematically explore the underlying communities of denitrifiers, Bacteria, and Archaea along redox gradients at distant geographic locations, nitrite reductase (nirS) genes and bacterial and archaeal 16S rRNA genes (rDNAs) were PCR amplified and analyzed by terminal restriction fragment length polymorphism (T-RFLP) analysis. The suitablility of T-RFLP analysis for investigating communities of nirS-containing denitrifiers was established by the correspondence of dominant terminal restriction fragments (T-RFs) of nirS to computer-simulated T-RFs of nirS clones. These clones belonged to clusters II, III, and IV from the same cores and were analyzed in a previous study (G. Braker, J. Zhou, L. Wu, A. H. Devol, and J. M. Tiedje, Appl. Environ. Microbiol. 66:2096-2104, 2000). T-RFLP analysis of nirS and bacterial rDNA revealed a high level of functional and phylogenetic diversity, whereas the level of diversity of Archaea was lower. A comparison of T-RFLPs based on the presence or absence of T-RFs and correspondence analysis based on the frequencies and heights of T-RFs allowed us to group sediment samples according to the sampling location and thus clearly distinguish Puget Sound and the Washington margin populations. However, changes in community structure within sediment core sections during the transition from aerobic to anaerobic conditions were minor. Thus, within the top layers of marine sediments, redox gradients seem to result from the differential metabolic activities of populations of similar communities, probably through mixing by marine invertebrates rather than from the development of distinct communities. PMID- 11282651 TI - 16S ribosomal DNA terminal restriction fragment pattern analysis of bacterial communities in feces of rats fed Lactobacillus acidophilus NCFM. AB - 16S ribosomal DNA terminal restriction fragment patterns from rat fecal samples were analyzed to track the dynamics of Lactobacillus acidophilus NCFM and discern bacterial populations that changed during feeding with NCFM. Lactobacillus johnsonii and Ruminococcus flavefaciens were tentatively identified as such bacterial populations. The presence of L. johnsonii was confirmed by isolation from feces. PMID- 11282650 TI - Comparative analysis of methane-oxidizing archaea and sulfate-reducing bacteria in anoxic marine sediments. AB - The oxidation of methane in anoxic marine sediments is thought to be mediated by a consortium of methane-consuming archaea and sulfate-reducing bacteria. In this study, we compared results of rRNA gene (rDNA) surveys and lipid analyses of archaea and bacteria associated with methane seep sediments from several different sites on the Californian continental margin. Two distinct archaeal lineages (ANME-1 and ANME-2), peripherally related to the order Methanosarcinales, were consistently associated with methane seep marine sediments. The same sediments contained abundant (13)C-depleted archaeal lipids, indicating that one or both of these archaeal groups are members of anaerobic methane-oxidizing consortia. (13)C-depleted lipids and the signature 16S rDNAs for these archaeal groups were absent in nearby control sediments. Concurrent surveys of bacterial rDNAs revealed a predominance of delta-proteobacteria, in particular, close relatives of Desulfosarcina variabilis. Biomarker analyses of the same sediments showed bacterial fatty acids with strong (13)C depletion that are likely products of these sulfate-reducing bacteria. Consistent with these observations, whole-cell fluorescent in situ hybridization revealed aggregations of ANME-2 archaea and sulfate-reducing Desulfosarcina and Desulfococcus species. Additionally, the presence of abundant (13)C-depleted ether lipids, presumed to be of bacterial origin but unrelated to ether lipids of members of the order Desulfosarcinales, suggests the participation of additional bacterial groups in the methane-oxidizing process. Although the Desulfosarcinales and ANME-2 consortia appear to participate in the anaerobic oxidation of methane in marine sediments, our data suggest that other bacteria and archaea are also involved in methane oxidation in these environments. PMID- 11282652 TI - Genotypic analysis of Escherichia coli strains from poultry carcasses and their susceptibilities to antimicrobial agents. AB - Plasmid profiling and amplified fragment length polymorphism (AFLP) analysis were used to genotype 50 Escherichia coli strains from poultry carcasses. Thirty different plasmid profiles were evident, and clustering of the AFLP data showed that they were a distinctly heterogeneous group of strains. Susceptibility testing against five antimicrobial agents used in the South African poultry industry showed all strains to be susceptible to danofloxacin and colistin, while the majority (96%) were resistant to two tetracyclines. PMID- 11282653 TI - Growth interactions during bacterial colonization of seedling rootlets. AB - Rootlet elongation and bacterial growth on rootlets were determined after inoculation of cucumber and spinach seedlings with Pseudomonas strains differing in production of siderophores and HCN. Siderophore producers grew more profusely than nonproducers on both species and promoted rootlet elongation on cucumber. Coinoculation of siderophore producers and nonproducers resulted in restricted growth of the latter. The total populations of nonproducers of HCN in the presence of HCN producers were not decreased, but the tenacity of their association with the rootlet surface was altered. PMID- 11282654 TI - Deep desulfurization of extensively hydrodesulfurized middle distillate oil by Rhodococcus sp. strain ECRD-1. AB - Dibenzothiophene (DBT), and in particular substituted DBTs, are resistant to hydrodesulfurization (HDS) and can persist in fuels even after aggressive HDS treatment. Treatment by Rhodococcus sp. strain ECRD-1 of a middle distillate oil whose sulfur content was virtually all substituted DBTs produced extensive desulfurization and a sulfur level of 56 ppm. PMID- 11282655 TI - Microbial growth on dichlorobiphenyls chlorinated on both rings as a sole carbon and energy source. AB - We have isolated bacterial strains capable of aerobic growth on ortho-substituted dichlorobiphenyls as sole carbon and energy sources. During growth on 2,2' dichlorobiphenyl and 2,4'-dichlorobiphenyl strain SK-4 produced stoichiometric amounts of 2-chlorobenzoate and 4-chlorobenzoate, respectively. Chlorobenzoates were not produced when strain SK-3 was grown on 2,4'-dichlorobiphenyl. PMID- 11282656 TI - Lack of cross-resistance to Cry19A from Bacillus thuringiensis subsp. jegathesan in Culex quinquefasciatus (Diptera: Culicidae) resistant to cry toxins from Bacillus thuringiensis subsp. israelensis. AB - Culex quinquefasciatus mosquitoes with high levels of resistance to single or multiple toxins from Bacillus thuringiensis subsp. israelensis were tested for cross-resistance to the Bacillus thuringiensis subsp. jegathesan polypeptide Cry19A. No cross-resistance was detected in mosquitoes that had been selected with the Cry11A, Cry4A and Cry4B, or Cry4A, Cry4B, Cry11A, and CytA toxins. A low but statistically significant level of cross-resistance, three to fourfold, was detected in the colony selected with Cry4A, Cry4B, and Cry11A. This cross resistance was similar to that previously detected with B. thuringiensis subsp. jegathesan in the same colony. These data help explain the toxicity of B. thuringiensis subsp. jegathesan against the resistant colonies and indicate that the Cry19A polypeptide might be useful in managing resistance and/or as a component of synthetic combinations of mosquitocidal toxins. PMID- 11282658 TI - Cloning and expression of a Ralstonia eutropha HF39 gene mediating indigo formation in Escherichia coli. AB - On complex medium Escherichia coli strains carrying hybrid plasmid pBEC/EE:11.0, pSKBEC/BE:9.0, pSKBEC/PP:3.3, or pSKBEC/PP:2.4 harboring genomic DNA of Ralstonia eutropha HF39 produced a blue pigment characterized as indigo by several chemical and spectroscopic methods. A 1,251-bp open reading frame (bec) was cloned and sequenced. The deduced amino acid sequence of bec showed only weak similarities to short-chain acyl-coenzyme A dehydrogenases, and the gene product catalyzed formation of indoxyl, a reactive preliminary stage for production of indigo. PMID- 11282657 TI - Identification of methyl halide-utilizing genes in the methyl bromide-utilizing bacterial strain IMB-1 suggests a high degree of conservation of methyl halide specific genes in gram-negative bacteria. AB - Strain IMB-1, an aerobic methylotrophic member of the alpha subgroup of the Proteobacteria, can grow with methyl bromide as a sole carbon and energy source. A single cmu gene cluster was identified in IMB-1 that contained six open reading frames: cmuC, cmuA, orf146, paaE, hutI, and partial metF. CmuA from IMB-1 has high sequence homology to the methyltransferase CmuA from Methylobacterium chloromethanicum and Hyphomicrobium chloromethanicum and contains a C-terminal corrinoid-binding motif and an N-terminal methyltransferase motif. However, cmuB, identified in M. chloromethanicum and H. chloromethanicum, was not detected in IMB-1. PMID- 11282659 TI - Biodegradation of n-alkylcycloalkanes and n-alkylbenzenes via new pathways in Alcanivorax sp. strain MBIC 4326. AB - The degradation of long-chain n-alkylbenzenes and n-alkylcyclohexanes by Alcanivorax sp. strain MBIC 4326 was investigated. The alkyl side chain of these compounds was mainly processed by beta-oxidation. In the degradation of n alkylcyclohexanes, cyclohexanecarboxylic acid was formed as an intermediate. This compound was further transformed to benzoic acid via 1-cyclohexene-1-carboxylic acid. PMID- 11282660 TI - Methyl t-butyl ether mineralization in surface-water sediment microcosms under denitrifying conditions. AB - Mineralization of [U-(14)C]methyl t-butyl ether (MTBE) to (14)CO(2) without accumulation of t-butyl alcohol (TBA) was observed in surface-water sediment microcosms under denitrifying conditions. Methanogenic activity and limited transformation of MTBE to TBA were observed in the absence of denitrification. Results indicate that bed sediment microorganisms can effectively degrade MTBE to nontoxic products under denitrifying conditions. PMID- 11282661 TI - Inhibition of a glucose-limited sequencing fed-batch culture of Salmonella enterica serovar enteritidis by volatile fatty acids representative of the ceca of broiler chickens. AB - The effects of concentrations of volatile fatty acids on an anaerobic, glucose limited, and pH-controlled growing culture of Salmonella enterica serovar Enteritidis were studied. Suddenly increasing volatile fatty acids to the concentrations representative of the ceca of 15-day-old broiler chickens caused washout of serovar Enteritidis. In contrast, a sudden increase to the volatile fatty acid concentrations representative of the ceca of younger broiler chickens caused a reduction in the biomass but not washout. Gradually increasing volatile fatty acids caused a gradual decrease in the biomass of serovar Enteritidis. We conclude that the concentrations of volatile fatty acids present in the ceca of broilers with a mature microflora can cause washout of serovar Enteritidis in an in vitro system mimicking cecal ecophysiology. PMID- 11282662 TI - Enhanced acid sensitivity of pressure-damaged Escherichia coli O157 cells. AB - Pressure-damaged Escherichia coli O157 cells were more acid sensitive than native cells and were impaired in pH homeostasis. However differences in acid sensitivity were not related to differences in cytoplasmic pH (pH(i)). Cellular beta-galactosidase was more acid labile in damaged cells. Sensitization to acid may thus involve loss of protective or repair functions. PMID- 11282663 TI - Rapid development of brain hypothermia using femoral-carotid bypass. AB - OBJECTIVES: Advances in the field of cardiopulmonary resuscitation have led to an increasing number of patients initially surviving sudden cardiac arrest. Unfortunately, most of these patients do not recover from the resultant anoxic brain insult. Several animal and human trials have suggested that post resuscitative brain hypothermia may improve neurologic recovery after cardiopulmonary arrest. Present cooling methods are slow, induce only brain surface cooling, or result in systemic hypothermia. The authors tested the hypothesis that unilateral hypothermic carotid bypass would induce bilateral brain cooling without evoking systemic hypothermia or hemodynamic instability. METHODS: Anesthetized, ventilated common swine (n = 6, 24-37 kg) underwent right femoral and carotid artery bypass cannulation. Central and peripheral hemodynamic parameters were recorded every 2 minutes throughout the procedure. Thermodynamic parameters included bilateral frontal lobe, bilateral nasopharyngeal, pulmonary artery, and rectal temperatures. Hypothermic femoral-carotid bypass was accomplished by drawing blood from the right femoral artery, cooling it to 24 degrees C, and returning it to the right carotid artery at a flow rate of 5 mL/kg/min for 30 minutes. RESULTS: With initiation of cooling, brain temperatures dropped rapidly from baseline of 37.2 degrees C to 30.6 degrees C (right frontal lobe) and 33.1 degrees C (left frontal lobe) at 30 minutes. Pulmonary artery and rectal temperatures also decreased, but never reached mild hypothermic levels (34 degrees C). There was no significant change in any hemodynamic parameters during brain cooling. CONCLUSIONS: Femoral-carotid hypothermic bypass rapidly induced a state of selective brain hypothermia without causing systemic hypothermia or hemodynamic instability. PMID- 11282664 TI - Direct correlation between diffusion of Loxosceles reclusa venom and extent of dermal inflammation. AB - OBJECTIVES: Envenomation by Loxosceles species (brown recluse) spiders results in large dermal inflammatory lesions. Venom-induced dermal inflammation occurs indirectly via soluble mediators of inflammation. This study aimed to explore whether the anatomic extent of dermonecrotic arachnidism is due to the cascade of soluble proinflammatory mediators elicited by venom deposited at the bite site, or due to diffusion of the venom per se. METHODS: Three New Zealand white rabbits received intradermal L. reclusa venom (3-microg) injections in the flank. At the time of maximum dermal inflammation (24 hr), paired 4-mm dermal biopsies were obtained in 2-cm intervals extending 0 to 12 cm from the inoculation site. Normal dermal tissue was obtained from the opposite flank to serve as a negative control. One biopsy sample from each interval was homogenized and assayed for myeloperoxidase (MPO) activity and for the presence of venom via an enzyme immunoassay (EIA). The other paired dermal biopsy was sectioned, and examined for the presence of polymorphonuclear neutrophils (PMNs) by microscopy. Lesional areas were measured using digital images imported into imaging software. RESULTS: Mean +/- SD lesional diameter 24 hours post inoculation measured 9.18 +/- 0.64 cm. Venom was detected in biopsies 0 to 10 cm from the injection site. As expected, the highest venom concentrations were measured at the inoculation site (4.28 +/- 3.9 ng/4 mm). In addition, PMNs and MPO were detected up to 8 and 10 cm from the inoculation site, respectively. Neither PMNs nor MPO was detected in tissue absent of venom (kappa = 0.88, p < 0.001). CONCLUSIONS: Loxosceles venom diffuses from the envenomation site. The extent of dermal inflammation mirrors the extent of Loxosceles venom diffusion. This observation implies that the venom itself defines the extent and magnitude of tissue injury following Loxosceles envenomation. PMID- 11282665 TI - Impact of technetium-99m sestamibi imaging on the emergency department management and costs in the evaluation of low-risk chest pain. AB - OBJECTIVES: To assess the impact of rest sestamibi scanning on emergency physicians' (EPs') diagnostic certainty and decision making (as assessed by the hypothetical disposition of patients) for 69 consenting stable patients with suspected acute cardiac ischemia and nondiagnostic electrocardiograms. The resultant impact on costs was examined as a secondary outcome. METHODS: Patients with suspected acute cardiac ischemia were injected with 25 mCi of sestamibi within two hours of active pain in one of three emergency department study sites. The probability of acute myocardial infarction (AMI) and unstable angina (UA), and hypothetical disposition decisions were recorded immediately before and after physicians were notified of scan results. Changes in disposition were classified as optimal or suboptimal. For the cost determinations, a cost-based decision support program was used. RESULTS: For the subgroup found to be free of acute cardiac events (ACEs) (n = 62), the EPs' post-sestamibi scan probabilities for AMI decreased by 11% and UA by 18% (p < 0.001 for both conditions). In seven patients with ACEs, the post-scan probabilities of AMI and UA increased, but neither was statistically significant. Scan results led to hypothetical disposition changes in 29 patients (42%), of which 27 (93%) were optimal (nine patients were reassigned to a lower level of care, two to a higher level, and 16 additional patients to "discharge-home" status). The strategy of scanning all patients who were low to moderate risk for acute cardiac ischemia would result in an increase of direct costs of care of $222 per patient evaluated, due to added cost of sestamibi scanning. CONCLUSIONS: Sestamibi scanning results appropriately affected the EPs' estimates of the probability of AMI and UA and improved disposition decisions. Scanning all low-risk patients would likely be associated with increased costs at this medical center. PMID- 11282666 TI - Limited response to cardiac arrest by police equipped with automated external defibrillators: lack of survival benefit in suburban and rural Indiana--the police as responder automated defibrillation evaluation (PARADE). AB - OBJECTIVE: To assess the out-of-hospital cardiac arrest (OHCA) survival advantage after providing police with automated external defibrillators (AEDs) in rural and suburban Indiana. METHODS: An observational evaluation was conducted in six Indiana counties (population: 464,741) before (retrospective) and after (prospective) training and equipping police with AEDs. The primary outcome evaluated was survival to hospital discharge for all cases of ventricular tachycardia/ventricular fibrillation (VT/VF) OHCA. Other factors evaluated include age, gender, race, arrest location, witnessed arrest, bystander cardiopulmonary resuscitation, response intervals, and survival to discharge for all OHCAs. Results are reported using chi-square, Student's t-test, and logistic regression. RESULTS: Police were equipped with 112 AEDs, increasing total defibrillator capability by 43.2%. During the study period, AED-equipped police responded prior to emergency medical services (EMS) in 26 of 388 cases (6.7%). The time intervals from 911 call-to-scene and 911 call-to-shock were shortened by 1.6 minutes (95% confidence interval [95% CI] = 0.0 to 3.1, p = 0.05) and 4.8 minutes (95% CI = 1.3 to 8.3, p = 0.008), respectively, with police response as compared with EMS response. Survival to hospital discharge for VT/VF OHCA was 15.0% (3/20) in cases in which police responded first and 10.0% (16/160) in cases in which EMS responded first (relative risk [RR] 0.63, 95% CI = 0.17 to 2.39, p = 0.45). Survival to hospital discharge for VT/VF OHCA did not improve from the prestudy period (16/204, 7.8%) to after police AED availability (19/180, 10.6%) (RR 0.72, 95% CI = 0.36 to 1.45, p = 0.38). CONCLUSIONS: Out-of-hospital cardiac arrest survival in suburban and rural Indiana did not improve after police were equipped with AEDs, likely related to poor police response. PMID- 11282667 TI - Emergency department right upper quadrant ultrasound is associated with a reduced time to diagnosis and treatment of ruptured ectopic pregnancies. AB - OBJECTIVE: To determine whether the time to diagnosis and treatment of patients with ruptured ectopic pregnancy is significantly less for patients who had emergency department (ED) right upper quadrant (RUQ) ultrasound (US) compared with those who had US in the radiology department. METHODS: The authors conducted a retrospective review of eligible patients presenting to an urban ED between January 1990 and December 1998. Patients were included in the study if they were seen in the ED, had a discharge diagnosis of ruptured ectopic pregnancy, were brought immediately to the operating room after a definitive diagnosis of ectopic pregnancy rupture was made, and had more than 400 mL of intraperitoneal blood found at the time of surgery. The ED, hospital, radiology, and operative records were reviewed to determine presenting vital signs, intraperitoneal blood loss, time to diagnosis, time to treatment, and type of US performed. RESULTS: There were 37 patients enrolled; 16 received ED RUQ US (group I) and 21 had a formal US in radiology (group II). The ages, pulses, systolic blood pressures, and volumes of hemoperitoneum were similar between the two groups. The average time to diagnosis from ED arrival was 58 minutes for group I (SD = 57; 95% CI = 28 to 87) and 197 minutes for group II (SD = 82; 95% CI = 162 to 232) (p < or = 0.0001). The average time to operative treatment was 111 minutes (group I) (SD = 86; 95% CI = 69 to 153) and 322 minutes (group II) (SD = 107; 95% CI = 270 to 364) (p < or = 0.0001), respectively. CONCLUSIONS: Patients with ruptured ectopic pregnancy, who were selected to have RUQ US performed in the ED by emergency physicians, had an average decrease in time to diagnosis of two and a quarter hours, and an average decrease in time to treatment of three and a half hours, compared with those having a formal pelvic US in the radiology department. Further prospective investigation is needed to determine whether ED RUQ US can safely expedite care of patients with suspected ectopic pregnancy. PMID- 11282668 TI - Intranasal lidocaine for the treatment of migraine headache: a randomized, controlled trial. AB - OBJECTIVE: To evaluate the effect of intranasal lidocaine for immediate relief (5 minutes) of migraine headache pain. METHODS: A randomized, double-blind, placebo controlled clinical trial at two university-affiliated community teaching hospitals enrolled patients 18-50 years old with migraine headache as defined by the International Headache Society. Patients who were pregnant, lactating, known to abuse alcohol or drugs, or allergic to one of the study drugs, those who used analgesics within two hours, or those with a first headache were excluded. Statistical significance was assessed by using chi-square or Fisher's exact test for categorical variables and Student's t-test for continuous variables. Patients rated their pain on a 10-centimeter visual analog scale (VAS) prior to drug administration and at 5, 10, 15, 20, and 30 minutes after the initial dose. Medication was either 1 mL of 4% lidocaine or normal saline (placebo) intranasally in split doses 2 minutes apart and intravenous prochlorperazine. Medications were packaged so physicians and patients were unaware of the contents. Successful pain relief was achieved if there was a 50% reduction in pain score or a score below 2.5 cm on the VAS. RESULTS: Twenty-seven patients received lidocaine and 22 placebo. No significant difference was observed between groups in initial pain scores, 8.4 (95% CI = 7.8 to 9.0) lidocaine and 8.6 (95% CI = 8.0 to 9.2) placebo (p = 0.75). Two of 27 patients (7.4%, 95% CI = 0.8, 24.3) in the lidocaine group and three of 22 patients (13.6%, 95% CI = 2.8 to 34.9) in the placebo group had immediate successful pain relief (p = 0.47), with average pain scores of 6.9 (95% CI = 5.9 to 7.8) and 7.0 (95% CI = 5.8 to 8.2), respectively. No difference in pain relief was detected at subsequent measurements. CONCLUSION: There was no evidence that intranasal lidocaine provided rapid relief for migraine headache pain in the emergency department setting. PMID- 11282670 TI - Electrocardiographic ST-segment elevation: correct identification of acute myocardial infarction (AMI) and non-AMI syndromes by emergency physicians. AB - OBJECTIVE: To determine the emergency physician's (EP's) ability to identify the cause of ST-segment elevation (STE) in a hypothetical chest pain patient. METHODS: Eleven electrocardiograms (ECGs) with STE were given to EPs; the patient in each instance was a 45-year-old male with a medical history of hypertension and diabetes mellitus with the chief complaint of chest pain. The EP was asked to determine the cause of the STE and, if due to acute myocardial infarction (AMI), to decide whether thrombolytic therapy (TT) would be administered (the patient had no contraindication to such treatment). Rates of TT administration were determined; appropriate TT administration was defined as that occurring in an AMI patient, while inappropriate TT administration was defined as that in the non-AMI patient. RESULTS: Four hundred fifty-eight EPs completed the questionnaire; levels of medical experience included the following: postgraduate year 2-3, 193 (42%); and attending, 265 (58%). The overall rate of correct interpretation of the study ECGs was 94.9% (4,782 correct interpretations out of 5,038 instances). Acute myocardial infarction with typical STE, ventricular paced rhythm, and right bundle branch block were never misinterpreted. The remaining conditions were misinterpreted with rates ranging between 9% (left bundle branch block, LBBB) and 72% (left ventricular aneurysm, LVA). The overall rate of appropriate thrombolytic agent administration was 83% (1,525 correct administrations out of 1,832 indicated administrations). The leading diagnosis for which thrombolytic agent was given inappropriately was LVA (28%), followed by benign early repolarization (23%), pericarditis (21%), and LBBB without electrocardiographic AMI (5%). Thrombolytic agent was appropriately given in all cases of AMI except when associated with atypical STE, where it was inappropriately withheld 67% of the time. CONCLUSIONS: In this survey, EPs were asked whether they would give TT based on limited information (ECG). Certain syndromes with STE were frequently misdiagnosed. Emergency physician electrocardiographic education must focus on the proper identification of these syndromes so that TT may be appropriately utilized. PMID- 11282669 TI - Analysis of school injuries resulting in emergency department or hospital admission. AB - OBJECTIVE: To describe the epidemiology of school injuries resulting in emergency department (ED) visits, hospital admission, or death. METHODS: Utah statewide school injuries from 1992 to 1996 were probabilistically linked to statewide ED records (1996 only), inpatient hospital records (1992-1996), and death certificate records (1992-1996). RESULTS: There were 43,881 school injuries for the years 1992 through 1996. In 1996, 1,534 of 6,354 total school injuries (17.5%) resulted in ED evaluation. Between 1992 and 1996, 354 school injuries (0.8%) necessitated hospital admission. The overall rates of school injuries (per 1,000 students) of primary (kindergarten-grade 6) and secondary (grades 7-12) school students requiring ED evaluation were 3.29 and 3.28, respectively; for hospital admission, 0.165 and 0.139. Abbreviated Injury Scale-1990 (AIS-90) regions identified in ED patients were the upper extremity (39.2%), face (20.8%), and lower extremity (17.1%), while AIS regions among inpatients were lower extremity (29.1%), upper extremity (26.6%), and head (22.6%). There were a total of 1,123 hospital days, and total charges of $2.16 million. The ED charges totaled $545,000. Median length of hospital stay was 1 day, and median hospital charge was $3,080. There were four fatalities. CONCLUSIONS: This study emphasizes the significance of school injuries and the need for interventions to prevent these injuries PMID- 11282671 TI - Targeted musculoarticular sonography in the detection of joint effusions. AB - This article describes an advanced application for an established technology, specifically the use of bedside sonography in the assessment of the acutely painful joint in the emergency department. The sonographic windows for each of the axial synovial joints are outlined, with a brief discussion of commonly encountered pathologic conditions. PMID- 11282672 TI - Sonographic features of small-bowel intussusception in pediatric patients. AB - OBJECTIVE: Small-bowel intussusception (SBI) for pediatric patients is unusual and difficult to diagnose preoperatively. This study sought to determine the sonographic findings of pediatric SBI. METHODS: The sonographic features and surgical findings of 13 pediatric patients (7 boys, 6 girls; age range 4 months 15 years; average age 4 years and 2 months) with SBI encountered in the authors' hospital over a 12-year period were retrospectively reviewed. RESULTS: Most of the patients presented with nonspecific symptoms, including vomiting, abdominal pain, and/or irritable crying. Sonographic screening in the emergency department revealed a doughnut or crescent-in-doughnut sign, or a multiple-concentric-rings sign for 11 of the 13 patients, and the lesions appeared short. Eight lesions were found in the paraumbilical or left abdominal regions. Sonographic measurement of the size of the lesions from these 11 patients ranged from 2 cm to 3.7 cm (average 2.77 cm). Subsequent barium enemas were performed for these 11 patients, none of which revealed colon lesions. Surgery revealed ileoileal intussusceptions for eight cases, jejunoileal for three, and jejunojejunal for the remaining two. Bowel ischemia or necrosis and pathologic lead points were demonstrated for seven and six patients, respectively, although none were recognized preoperatively. CONCLUSIONS: Small-bowel intussusception is often over looked due to nonspecific clinical presentations. Sonographic demonstration of a 2-3-cm sized, short, doughnut-like lesion, especially in the left abdomen or paraumbilical regions, should lead to strong suspicion of SBI. PMID- 11282673 TI - Clinicopathological conference: a previously healthy 40-year-old woman with hemoptysis. PMID- 11282674 TI - ST-elevation acute myocardial infarction: a critical but difficult electrocardiographic diagnosis. PMID- 11282675 TI - Building a profession. PMID- 11282676 TI - Ethics seminars: the practice of medical procedures on newly dead patients--is consent warranted? AB - The use of the newly dead to teach procedures is widely practiced in training institutions. This model allows a realistic opportunity both to become more familiar with lifesaving maneuvers before they are actually necessary and to maintain proficiency. Whether to notify the next of kin first has been an issue of ethical debate. Some argue a "don't ask, don't tell" policy is justified, while others mandate open consent by family members prior to the practice. Several medical studies have found that patients and families are likely to consent to the procedures but prefer to be asked permission first. Multiple legal cases have addressed the issue of usage of cadavers postmortem without expressed permission. Earlier cases emphasized the concept of "pseudo-property" rights and declared that the next of kin do not have constitutional ownership of the deceased person's body. More recent legal cases are declaring that families do, in fact, possess these rights. In this day and age of increasing recognition of personal autonomy, it is probably prudent to approach the next of kin for permission before performing procedures on the newly deceased. PMID- 11282677 TI - AAEM, CORD, and SAEM reach a landmark position: consensus recommendations to the Federation of State Medical Boards (FSMB) for revisions to the FSMB May 1998 policy statement on physician licensure. American Academy of Emergency Medicine. Council of Emergency Medicine Residency Directors. Society for Academic Medicine. AB - As a result of months of meetings and deliberations coordinated with the Medical Board of California and chaperoned by the California Chapter of the American Academy of Emergency Medicine (CAL/AAEM), the Society for Academic Medicine (SAEM), the Council of Emergency Medicine Residency Directors (CORD), and the American Academy of Emergency Medicine (AAEM) recently reached a landmark agreement on recommendations to the Federation of State Medical Boards (FSMB) pertaining to controversial May 1998 FSMB recommendations regarding physician licensure. Endorsed unanimously by the boards of all three emergency medicine (EM) organizations, the recommendations of this consensus have been forwarded to the FSMB and await its official response. The recommendations will also be forwarded to remaining EM organizations and to the medical community for comment and to enlist their support. PMID- 11282679 TI - Ultrasound-guided breast abscess aspiration in a difficult case. AB - Although ultrasound guidance is occasionally used for abscess detection and aspiration by our radiology colleagues, this is still a very uncommon application in the emergency department (ED). A case is presented of a patient with a difficult-to-drain, recurrent breast abscess. The consulting surgeon was unable to localize the abscess after 15 attempts at aspiration in the ED and requested ultrasound guidance from the attending emergency physician for the procedure. Drainage of the abscess was successfully completed in one attempt with real-time visualization and guidance of the needle. The consulting surgeon requested that ultrasound be available at the patient's follow-up visit to the ED. PMID- 11282678 TI - Curling iron-related injuries presenting to U.S. emergency departments. AB - OBJECTIVE: To describe curling iron-related injuries reported to the National Electronic Injury Surveillance System (NEISS) between January 1, 1992, and December 31, 1996. METHODS: The authors retrospectively reviewed data from NEISS, a weighted probability sample of emergency departments (EDs) developed to monitor consumer product-related injuries. The information reported includes patient demographics, injury diagnosis, body part injured, incident locale, patient disposition, and a brief narrative description. The authors reviewed the narrative in the hair care products category and abstracted records indicating the injury was caused by contact with a curling iron. Also analyzed were the design features of commonly available curling irons purchased from national discount department stores. RESULTS: There were an estimated 105,081 hair care product-related injuries in the five-year period, of which 82,151 (78%) involved a curling iron. Seventy percent of injuries were to females. The patient's median age was 8 years (range 1 month to 96 years). The most commonly occurring injury was thermal burns (97%; 79,912/82,151). Ninety-eight percent of the injuries occurred in the home and 99% of the patients were discharged home from the ED. In patients <4 years old, 56% of burns occurred by grabbing or touching, while in those > or =10 years the burns occurred by contact while in use. In the older group 69% of burns were of the cornea. Most curling irons use small amounts of power, yet there are no standards for temperature settings or control. The cylinder containing the heating element is mostly exposed, and many irons do not have a power switch. CONCLUSIONS: The most common injury resulting from curling irons is thermal burns. The mechanisms and patterns of injury in developmentally distinct age groups suggest that many of these injuries could be prevented by public education and the re-engineering of curling irons. PMID- 11282680 TI - The outstanding medical student in emergency medicine. PMID- 11282681 TI - Prediction rule in opioid overdose. PMID- 11282682 TI - Reliability and validity of diagnostic tests. PMID- 11282683 TI - Progress in eating disorders research. PMID- 11282684 TI - Effectiveness of second-generation antipsychotics in patients with treatment resistant schizophrenia: a review and meta-analysis of randomized trials. AB - OBJECTIVE: The authors conducted a review and meta-analysis of studies that compared the efficacy and tolerability of typical and second-generation antipsychotics for patients with treatment-resistant schizophrenia. METHOD: A systematic search revealed 12 controlled studies (involving 1,916 independent patients), which were included in the review. For the seven studies that compared clozapine to a typical antipsychotic, a meta-analysis was performed to examine clozapine's effects on overall psychopathology, response rate, extrapyramidal symptoms, and tardive dyskinesia. RESULTS: The meta-analysis confirmed that treatment-resistant schizophrenic patients have more favorable outcomes when treated with clozapine rather than a typical antipsychotic, as reflected by Brief Psychiatric Rating Scale total score, categorical response rate, Scale for the Assessment of Negative Symptoms score, Simpson-Angus Rating Scale score, and compliance rate. Clozapine also conferred benefits on the sickest treatment resistant schizophrenic patients. Patients treated with olanzapine also had more favorable outcomes with regard to categorical response and compliance rates. CONCLUSIONS: In the aggregate, the results of a meta-analysis indicated that clozapine exhibits superiority over typical antipsychotics in terms of both efficacy (as measured by improvement in overall psychopathology) and safety (in terms of reduced extrapyramidal side effects). However, the magnitude of the clozapine treatment effect was not consistently robust. Efficacy data for other second-generation antipsychotics in the treatment of patients with refractory schizophrenia were inconclusive. There is, therefore, a growing need to consider new and different treatment strategies, whether they be adjunctive or monotherapeutic, for schizophrenia that continues to be resistant or only partially responsive to treatment. PMID- 11282685 TI - Cognitive neuropsychiatric models of persecutory delusions. AB - OBJECTIVE: The major cognitive theories of persecutory delusion formation and maintenance are critically examined in this article. METHOD: The authors present a comprehensive review of the literature, citing results of relevant functional neuroimaging and neural network studies. RESULTS: People with persecutory delusions selectively attend to threatening information, jump to conclusions on the basis of insufficient information, attribute negative events to external personal causes, and have difficulty in envisaging others' intentions, motivations, or states of mind. Presence of the "reality distortion" cluster of psychotic symptoms correlates with cerebral blood flow in the left lateral prefrontal cortex, ventral striatum, superior temporal gyrus, and parahippocampal region. Social cognitive processing (selective attention to threat, attribution of causation or mental states) in normal subjects involves similar areas. Neural network models of persecutory delusions highlight the importance of disordered neuromodulation in their formation and of disordered neuroplasticity in their maintenance. CONCLUSIONS: Further studies examining the interaction of these cognitive processes, cross-sectionally and longitudinally, at cognitive psychological, neural network, and functional neuroanatomical levels are warranted to establish a comprehensive cognitive neuropsychiatric model of the persecutory delusion. PMID- 11282686 TI - Cognitive function and cerebral perfusion during cocaine abstinence. PMID- 11282688 TI - Personality profiles in eating disorders: rethinking the distinction between axis I and axis II. AB - OBJECTIVE: Like other DSM-IV axis I syndromes, eating disorders are diagnosed without respect to personality, which is coded on axis II. The authors assessed the utility of segregating eating disorders and personality pathology and examined the extent to which personality patterns account for meaningful variation within axis I eating disorder diagnoses. METHOD: One hundred three experienced psychiatrists and psychologists used a Q-sort procedure (the Shedler Westen Assessment Procedure-200) that assesses personality and personality pathology to describe a patient they were currently treating for bulimia or anorexia. Data were subjected to a cluster-analytic procedure (Q-analysis) to determine whether patients clustered into coherent groupings on the basis of their personality profiles. Categorical and dimensional personality diagnoses were then used to predict measures relevant to adaptation and etiology, controlling for axis I diagnosis. RESULTS: Three categories of patients emerged: a high-functioning/perfectionistic group, a constricted/overcontrolled group, and an emotionally dysregulated/undercontrolled group. This categorization demonstrated substantial incremental validity beyond axis I diagnosis in predicting eating disorder symptoms, adaptive functioning (Global Assessment of Functioning scores and history of psychiatric hospitalization), and etiological variables (sexual abuse history). CONCLUSIONS: Axis I symptoms are a useful component, but only one component, in the accurate diagnosis of eating disorders. Classifying patients with eating disorders by eating symptoms alone groups together patients with anorexic symptoms who are high functioning and self critical with those who are highly disturbed, constricted, and avoidant, and groups together patients with bulimic symptoms who are high functioning and self critical with those who are highly disturbed, impulsive, and emotionally dysregulated. These distinctions may be relevant to etiology, prognosis, and treatment. PMID- 11282689 TI - Morbidity risk for obsessive-compulsive spectrum disorders in first-degree relatives of patients with eating disorders. AB - OBJECTIVE: A hypothesis that eating disorders are a phenomenological variant of obsessive-compulsive disorder (OCD) has been proposed. This study was conducted to determine whether anorexia nervosa and bulimia, the two main eating disorders, are familial and whether the risk for obsessive-compulsive spectrum disorders (OCD and tic disorders) is higher in families of patients with eating disorders. METHOD: The morbidity risk for obsessive-compulsive spectrum disorders in first degree relatives of 136 female probands with eating disorders (84 with anorexia nervosa, 52 with bulimia) was compared to that for first-degree relatives of 72 female comparison subjects. RESULTS: The morbidity risk for obsessive-compulsive spectrum disorders was significantly higher among the 436 relatives of the eating disorder probands than among the 358 relatives of the comparison subjects (9.69% versus 0%). This finding was independent of any comorbid diagnosis of an obsessive-compulsive spectrum disorder in the eating disorder probands. The eating disorder group and the comparison group did not differ in familial risk for eating disorders and tic disorders. CONCLUSIONS: To better understand the genetic components of eating disorders, these disorders should be considered as part of the obsessive-compulsive spectrum of disorders. PMID- 11282690 TI - Comparisons of men with full or partial eating disorders, men without eating disorders, and women with eating disorders in the community. AB - OBJECTIVE: The authors compared 62 men who met all or most of the DSM-III-R criteria for eating disorders with 212 women who had similar eating disorders and 3,769 men who had no eating disorders on a wide variety of clinical and historical variables. METHOD: The groups of subjects were derived from a community epidemiologic survey performed in the province of Ontario that used the World Health Organization's Composite International Diagnostic Interview. RESULTS: Men with eating disorders were very similar to women with eating disorders on most variables. Men with eating disorders showed higher rates of psychiatric comorbidity and more psychosocial morbidity than men without eating disorders. CONCLUSIONS: These results confirm the clinical similarities between men with eating disorders and women with eating disorders. They also reveal that both groups suffer similar psychosocial morbidity. Men with eating disorders show a wide range of differences from men without eating disorders; the extent to which these differences are effects of the illness or possible risk factors for the occurrence of these illnesses in men is not clear. PMID- 11282691 TI - Altered pituitary-adrenal axis responses to provocative challenge tests in adult survivors of childhood abuse. AB - OBJECTIVE: Early adverse life events may predispose individuals to the development of mood and anxiety disorders in adulthood, perhaps by inducing persistent changes in corticotropin-releasing factor (CRF) neuronal systems. The present study sought to evaluate pituitary-adrenal responses to standard hypothalamic-pituitary-adrenal axis challenge tests in adult female survivors of childhood abuse with and without major depressive disorder. METHOD: Plasma ACTH and cortisol responses to the administration of 1 microg/kg ovine CRF and plasma cortisol responses to the administration of 250 microg ACTH(1-24) were measured in healthy women without early life stress (N=20), women with childhood abuse without major depressive disorder (N=20), women with childhood abuse and major depressive disorder (N=15), and women with major depression but no early life stress (N=11). RESULTS: Abused women without major depressive disorder exhibited greater than usual ACTH responses to CRF administration, whereas abused women with major depressive disorder and depressed women without early life stress demonstrated blunted ACTH responses. In the ACTH(1-24) stimulation test, abused women without major depressive disorder exhibited lower baseline and stimulated plasma cortisol concentrations. Abused women with comorbid depression more often suffered from posttraumatic stress disorder and reported more recent life stress than abused women without major depressive disorder. CONCLUSIONS: These findings suggest sensitization of the anterior pituitary and counterregulative adaptation of the adrenal cortex in abused women without major depressive disorder. On subsequent stress exposure, women with a history of childhood abuse may hypersecrete CRF, resulting in down-regulation of adenohypophyseal CRF receptors and symptoms of depression and anxiety. PMID- 11282692 TI - Genetic risk, number of previous depressive episodes, and stressful life events in predicting onset of major depression. AB - OBJECTIVE: The association between stressful life events and the onset of major depression decreases as the number of previous depressive episodes increases. How do genetic risk factors for major depression impact on this "kindling" phenomenon? In particular, do those at high genetic risk exhibit an increase in the speed of kindling, or are they "prekindled"? METHOD: Using discrete-time survival analysis, the authors examined the interaction between genetic risk, number of previous depressive episodes, and life event exposure in the prediction of episodes of major depression in female-female twin pairs from a population based registry. The twins were interviewed four times over a 9-year period, producing 92,521 person-months of exposure. RESULTS: The decline in the association between stressful life events and risk for major depression as the number of previous depressive episodes increased was strongest in those at low genetic risk and was weak to absent in those at high genetic risk. In the absence of previous depressive episodes, those at high genetic risk frequently experienced depressive episodes without major environmental stressors. CONCLUSIONS: Genetic risk factors for depression produce a "prekindling" effect rather than increase the speed of kindling. The "kindled" state, wherein depressive episodes occur with little provocation, may be reached by two pathways: many previous depressive episodes, perhaps driven by multiple adversities, and high genetic risk. PMID- 11282693 TI - Gender differences in the rates of exposure to stressful life events and sensitivity to their depressogenic effects. AB - OBJECTIVE: Women are at greater risk for major depression than men. The authors sought to determine whether the gender difference in prevalence for major depression was due to more frequent exposure to stressful life events and/or greater sensitivity to their depressogenic effects. METHOD: Male-male, female female, and male-female twin pairs from a population-based registry were personally interviewed. Each interview assessed the occurrence, to the nearest month, of 18 personal and social network classes of stressful life events and episode onsets of major depression. Standard logistic regression analyses were conducted for the same-sex pairs, and each female twin in the opposite-sex pairs was compared with her male co-twin by using conditional logistic regression. RESULTS: Women consistently reported higher rates of housing problems, loss of confidant, crises and problems getting along with individuals in their proximal network, and illness of individuals within their distal network. In both the same sex and opposite-sex samples, men reported higher rates of job loss, legal problems, robbery, and work problems. Consistent sex differences in the depressogenic effect of stressful life events were seen for three event categories: men were more sensitive to the depressogenic effects of divorce or separation and work problems; women were more sensitive to the depressogenic effects of problems getting along with individuals in their proximal network. None of the gender difference in prevalence of major depression could be explained by differing rates of or sensitivities to stressful life events. CONCLUSIONS: Women reported more interpersonal whereas men reported more legal and work-related stressful life events. Most life event categories influenced the risk for major depression similarly in the two sexes. The results suggest that the greater prevalence of major depression in women versus men is due neither to differences in the rates of reported stressful life events nor to differential sensitivity to their pathogenic effect. PMID- 11282694 TI - Incidence and prediction of posttraumatic stress disorder symptoms in severely injured accident victims. AB - OBJECTIVE: This study was designed to assess the incidence of posttraumatic stress disorder (PTSD) in severely injured accident victims and to predict the presence of PTSD symptoms at a 12-month follow-up. METHOD: A longitudinal, 1-year follow-up study was carried out with 106 consecutive patients with severe accidental injuries who were admitted to the trauma surgeons' intensive care unit at a university hospital. Patients were interviewed within 1 month and 12 months after the accident. Assessments included an extensive clinical interview, the Impact of Event Scale, the Clinician-Administered PTSD Scale, the Sense of Coherence questionnaire, and the Freiburg Questionnaire of Coping With Illness. RESULTS: A total of 13.4 days (SD=6.6) after the accident, five patients (4.7%) met all criteria for PTSD with the exception of the time criterion. A total of 22 other patients (20.8%) had subsyndromal PTSD. At the 1-year follow-up, two patients (1.9%) had PTSD, and 13 (12.3%) had subsyndromal PTSD. Multiple regression analysis explained 34% of the variance of PTSD symptoms 12 months after the accident. Biographical risk factors, the sense of a death threat, symptoms of intrusion, and problem-oriented coping each contributed significantly to the predictive model. CONCLUSIONS: In severely injured accident victims who were healthy before experiencing trauma, the incidence of PTSD was low. One-third of the variance of PTSD symptoms at 1-year follow-up could be predicted by mainly psychosocial variables. PMID- 11282695 TI - Hypnotizability in acute stress disorder. AB - OBJECTIVE: This study investigated the relationship between acute dissociative reactions to trauma and hypnotizability. METHOD: Acutely traumatized patients (N=61) with acute stress disorder, subclinical acute stress disorder (no dissociative symptoms), and no acute stress disorder were administered the Stanford Hypnotic Clinical Scale within 4 weeks of their trauma. RESULTS: Although patients with acute stress disorder and patients with subclinical acute stress disorder displayed comparable levels of nondissociative psychopathology, acute stress disorder patients had higher levels of hypnotizability and were more likely to display reversible posthypnotic amnesia than both patients with subclinical acute stress disorder and patients with no acute stress disorder. CONCLUSIONS: The findings may be interpreted in light of a diathesis-stress process mediating trauma-related dissociation. People who develop acute stress disorder in response to traumatic experience may have a stronger ability to experience dissociative phenomena than people who develop subclinical acute stress disorder or no acute stress disorder. PMID- 11282696 TI - Elevated D8/17 expression on B lymphocytes, a marker of rheumatic fever, measured with flow cytometry in tic disorder patients. AB - OBJECTIVE: Elevated D8/17 expression on B lymphocytes is a known susceptibility marker of rheumatic fever. Previous studies have reported higher than usual D8/17 expression on B lymphocytes of patients with tic disorders. The purpose of this study was to assess D8/17 expression on B lymphocytes of tic disorder patients by using an objective method in which no operator variability was involved. METHOD: D8/17 expression on B lymphocytes was assessed with flow cytometry by using an immunoglobulin M (IgM) monoclonal D8/17-specific antibody in an unselected group of Dutch patients with tic disorders (N=33) and healthy volunteers (N=20). Binding of this monoclonal antibody was compared with binding of an irrelevant IgM monoclonal antibody, and the shift in mean fluorescence intensity of the D8/17-specific antibody compared to that of the irrelevant IgM monoclonal antibody was used as a measure of D8/17 overexpression. For the patients, Yale Global Tic Severity Scale scores were used to assess disease severity. RESULTS: D8/17 overexpression in the patient group (mean=16.8 arbitrary units, SD=30.5) was significantly higher than in the comparison group (mean=3.2, SD=3.0). A significant minority of the patients (N=13, 39.4%), however, had levels of D8/17 overexpression within the range of that of the healthy comparison subjects. Flow cytometric analysis did not indicate a separate subpopulation of D8/17-positive B cells. CONCLUSIONS: These data confirm the utility of D8/17 B cell overexpression as a peripheral blood marker in patients with tic disorders and are compatible with a streptococcus-related pathogenesis for at least a subgroup of patients with tic disorders. PMID- 11282697 TI - Impact of tic disorders on ADHD outcome across the life cycle: findings from a large group of adults with and without ADHD. AB - OBJECTIVE: The impact of tic disorders on the outcome of attention deficit hyperactivity disorder (ADHD) remains a subject of high scientific and clinical interest. To evaluate the impact of comorbid ADHD and tic disorders from a lifespan perspective, the authors systematically examined data from adults with and without ADHD. METHOD: They comprehensively evaluated 312 consecutively referred adults with ADHD and 252 comparison subjects without ADHD. Tic disorders were characterized along with a wide range of neuropsychiatric correlates, including other comorbid disorders as well as indexes of function in the domains of school, cognition, and interpersonal functioning. RESULTS: A significantly greater proportion of adults with ADHD (12%) than those without ADHD (4%) had tic disorders. Tic disorders followed a mostly remitting course and had little impact on functional capacities. In addition, tic disorders were not associated with stimulant use. CONCLUSIONS: These findings in adults with ADHD confirm and extend previous findings in young subjects with ADHD, documenting that although individuals with ADHD are at greater risk for tic disorders, the presence of tic disorders has a limited impact on ADHD outcome. PMID- 11282698 TI - Thalamic volumes in patients with first-episode schizophrenia. AB - OBJECTIVE: The thalamus, a highly evolved sensory and motor gateway to the cortex, has been implicated in the pathophysiology of several illnesses, including schizophrenia. Several studies have suggested thalamic volume differences in patients with schizophrenia, although only a few studies have examined thalamic structure in new-onset patients. METHOD: The authors used magnetic resonance imaging to measure thalamic volumes in previously untreated patients with first-episode schizophrenia (N=16) relative to those of healthy comparison subjects (N=25). The age range of the patients and comparison subjects was 15 to 45 years of age. Thalamic volumes in the right and left hemispheres were segmented and analyzed, both separately and as total thalamic volume, by a rater blind to clinical data. The thalamus was further segmented into regions that roughly reflected individual thalamic nuclei. Analysis of covariance was used to control for intracranial volume. RESULTS: Right, left, and total thalamic volumes of the patients with schizophrenia were significantly smaller than those of the comparison subjects. Significantly smaller volumes were found in the left central medial subdivision of the patients as well as a smaller volume in the right central medial subdivision that approached significance. These regions primarily comprised the dorsomedial nucleus, a thalamic nucleus thought to be an important component of aberrant circuitry in schizophrenia. Significant volume differences were also seen in the left anterior, right anterior, and right posterior medial subdivisions. CONCLUSIONS: These findings suggest significant thalamic volumetric differences between patients with newly diagnosed schizophrenia and healthy comparison subjects. Future analysis of individual thalamic nuclei may reveal important, specific relationships between thalamic abnormalities and schizophrenia. PMID- 11282699 TI - An MRI study of basal ganglia volumes in first-episode schizophrenia patients treated with risperidone. AB - OBJECTIVE: The basal ganglia may contribute to extrapyramidal movement disorders, affective disturbances, and cognitive deficits in schizophrenia. Basal ganglia volumes are putatively affected by antipsychotic medications. The purpose of this study was to determine the long-term effects of risperidone treatment in a cohort of first-episode patients with schizophrenia. METHOD: The subjects were 30 patients with first-episode schizophrenia, 12 patients chronically treated with typical antipsychotics, and 23 healthy comparison subjects. They were scanned by magnetic resonance imaging at baseline. The first-episode patients received 1 year of continuous risperidone treatment, after which they and the comparison subjects were rescanned. Caudate, putamen, and globus pallidus volumes were determined from coronal images. RESULTS: The baseline caudate, putamen, and globus pallidus volumes were significantly larger in the chronically treated patients than in the untreated first-episode subjects and comparison subjects. These volumes did not differ between the first-episode patients and healthy comparison subjects. Basal ganglia volumes were unchanged after 1 year of exposure to risperidone in the first-episode subjects. Extrapyramidal movement disorders were present in the majority of chronically treated patients and more than one-third of the never-medicated first-episode patients at baseline. CONCLUSIONS: This group of first-episode patients did not exhibit abnormalities of basal ganglia volumes, nor were basal ganglia volumes affected by exposure to risperidone. Movement disorders were observed in both first-episode and chronically treated patients, suggesting effects of both illness and medications. PMID- 11282700 TI - Dialectical behavior therapy for bulimia nervosa. AB - OBJECTIVE: The effects of dialectical behavior therapy adapted for the treatment of binge/purge behaviors were examined. METHOD: Thirty-one women (averaging at least one binge/purge episode per week) were randomly assigned to 20 weeks of dialectical behavior therapy or 20 weeks of a waiting-list comparison condition. The manual-based dialectical behavior therapy focused on training in emotion regulation skills. RESULTS: An intent-to-treat analysis showed highly significant decreases in binge/purge behavior with dialectical behavior therapy compared to the waiting-list condition. No significant group differences were found on any of the secondary measures. CONCLUSIONS: The use of dialectical behavior therapy adapted for treatment of bulimia nervosa was associated with a promising decrease in binge/purge behaviors. PMID- 11282701 TI - Effects of sustained-release bupropion and supportive group therapy on cigarette consumption in patients with schizophrenia. AB - OBJECTIVE: The study examined the efficacy, tolerability, and safety of supportive group psychotherapy and adjunctive sustained-release bupropion for nicotine addiction in patients with schizophrenia. METHOD: Eight patients participated in a 14-week open-label trial. End expired breath carbon monoxide level, symptom levels, neuropsychological performance, and suppression of the P50 event-related potential were measured before and after the 14-week trial. RESULTS: Patients showed a decrease in carbon monoxide levels that was not associated with any worsening in symptom, neuropsychological, or P50 suppression measures. CONCLUSIONS: Use of sustained-release bupropion in combination with supportive group therapy may help patients with schizophrenia decrease their cigarette consumption. PMID- 11282702 TI - Postpartum depression in women receiving public assistance: pilot study of an interpersonal-therapy-oriented group intervention. AB - OBJECTIVE: This study investigated whether a preventive intervention based on the principles of interpersonal psychotherapy administered to pregnant women would reduce the risk of postpartum major depression. METHOD: Thirty-seven pregnant women receiving public assistance who had at least one risk factor for postpartum depression were randomly assigned to a four-session group intervention or to a treatment-as-usual condition. Thirty-five of the women completed the study. Structured diagnostic interviews were administered to assess for postpartum major depression. RESULTS: Within 3 months after they gave birth, six (33%) of the 18 women in the treatment-as-usual condition had developed postpartum major depression, compared with none of the 17 women in the intervention condition. CONCLUSIONS: A four-session interpersonal-therapy-oriented group intervention was successful in preventing the occurrence of major depression during a postpartum period of 3 months in a group of financially disadvantaged women. PMID- 11282703 TI - Urinary free cortisol in chronic fatigue syndrome. AB - OBJECTIVE: The authors measured 24-hour urinary free cortisol in a group of well characterized patients with chronic fatigue syndrome. METHOD: They obtained 24 hour urine collections from 121 consecutive clinic patients with chronic fatigue syndrome and 64 comparison subjects without the syndrome. RESULTS: Urinary free cortisol was significantly lower in the subjects with chronic fatigue syndrome regardless of the presence or absence of current or past comorbid psychiatric illness. Lower levels of urinary free cortisol were not related to medication use, sleep disturbance, or disability levels. CONCLUSIONS: There is mild hypocortisolism in chronic fatigue syndrome. Whether a primary feature or secondary to other factors, hypocortisolism may be one factor contributing to the symptoms of chronic fatigue syndrome. PMID- 11282705 TI - Functional magnetic resonance imaging evidence for disrupted basal ganglia function in schizophrenia. AB - OBJECTIVE: This study was an examination of basal ganglia dysfunction in schizophrenia using functional magnetic resonance imaging (fMRI). METHOD: The authors used a motor sequencing task to investigate activation of the caudate, anterior putamen plus globus pallidus, and posterior putamen plus globus pallidus in eight subjects with schizophrenia and 12 group-matched comparison subjects. Differences in activation of the thalamus, the target of direct output from the globus pallidus, were also examined. RESULTS: The schizophrenia subjects showed significant bilateral deficits in the posterior putamen, globus pallidus, and thalamus but not the anterior putamen plus globus pallidus or caudate. Functional connectivity analysis revealed that the deficits in thalamic activation were related to deficits in posterior putamen and globus pallidus activation. CONCLUSIONS: These results provide fMRI evidence for basal ganglia dysfunction in subjects with schizophrenia and suggest that this deficit results in disrupted outflow to the thalamus. These deficits may underlie the behavioral impairments in goal-directed action observed in schizophrenia. PMID- 11282704 TI - Effect of clozapine on caudate nucleus volume in relation to symptoms of schizophrenia. AB - OBJECTIVE: Studies have found that caudate volume increased after treatment with typical antipsychotics in patients with schizophrenia but decreased after treatment was changed to clozapine. In the current study the authors examined whether this volume decrease was related to clinical improvement. METHOD: Twenty eight patients with schizophrenia who had not responded to treatment with typical antipsychotics were included in the study; 22 completed the study. Caudate volume was assessed by using magnetic resonance imaging during treatment with typical antipsychotics and after 24 weeks and 52 weeks of clozapine treatment. Symptoms were assessed just before clozapine treatment and once a month thereafter. RESULTS: Clozapine treatment resulted in a significant reduction in left caudate volume in patients who responded to the drug but not in patients who did not respond to clozapine at 52 weeks of treatment. Overall, the degree of reduction in left caudate volume was significantly related to the extent of improvement in positive and general symptoms but not in negative symptoms. CONCLUSIONS: These findings suggest that the caudate nucleus plays a role in the positive and general symptoms of schizophrenia. PMID- 11282706 TI - Neuroleptic malignant syndrome after addition of haloperidol to atypical antipsychotic. PMID- 11282707 TI - Risperidone-induced retrograde ejaculation. PMID- 11282708 TI - Abuse of topical analgesic. PMID- 11282709 TI - Granulocytopenia with clozapine and quetiapine. PMID- 11282710 TI - Hippocampus and amygdala pathology in depression. PMID- 11282711 TI - Postpartum depression with bipolar disorder. PMID- 11282712 TI - Effectiveness of involuntary outpatient commitment. PMID- 11282714 TI - Effectiveness of involuntary outpatient commitment. PMID- 11282717 TI - Trauma and depersonalization during panic attacks. PMID- 11282718 TI - Ondansetron for tardive dyskinesia. PMID- 11282719 TI - Personality disorders in the literature. PMID- 11282720 TI - Poststroke depression. PMID- 11282724 TI - Verbal working memory impairment in schizophrenia. PMID- 11282740 TI - Visualization of neural activity associated with dyspnea. PMID- 11282741 TI - Ventilator-induced airway dysfunction? PMID- 11282742 TI - Diagnosing latent tuberculosis infection: the 100-year upgrade. PMID- 11282743 TI - Addressing the ethical problems of randomized and placebo-controlled trials of CPAP. PMID- 11282744 TI - Ventilator monitoring, and sharing the data with patients. PMID- 11282745 TI - Spiral computed tomography screening for lung cancer is ready for prime time. PMID- 11282746 TI - Low-dose spiral computed tomography screening for lung cancer: not ready for prime time. PMID- 11282749 TI - The oscillations of HFO. PMID- 11282750 TI - Two transatlantic viewpoints on an ethical quandary. PMID- 11282751 TI - "Reactive airways disease". A lazy term of uncertain meaning that should be abandoned. PMID- 11282752 TI - Rapid detection of Mycobacterium tuberculosis infection by enumeration of antigen specific T cells. AB - There is no reliable means of detecting latent M. tuberculosis infection, and even in patients with active tuberculosis, infection is often unconfirmed. We hypothesized that M. tuberculosis antigen-specific T cells might reliably indicate infection. We enumerated peripheral blood-derived interferon gamma (IFN gamma)-secreting T cells responding to epitopes from ESAT-6, an antigen that is highly specific for M. tuberculosis complex but absent from BCG, in four groups of individuals. Forty-five of 47 patients with bacteriologically confirmed tuberculosis had ESAT-6-specific IFN-gamma-secreting T cells, compared with four of 47 patients with nontuberculous illnesses, indicating that these T cells are an accurate marker of M. tuberculosis infection. This assay thus has a sensitivity of 96% (95% confidence interval [CI] 92-100) for detecting M. tuberculosis infection in this patient population. By comparison, of the 26 patients with tuberculosis who had a diagnostic tuberculin skin test (TST), only 18 (69%) were positive (p = 0.003). In addition, 22 of 26 (85%) TST-positive exposed household contacts had ESAT-6-specific T cells, whereas zero of 26 unexposed BCG-vaccinated subjects responded. This approach enables rapid detection of M. tuberculosis infection in patients with active tuberculosis and in exposed asymptomatic individuals at high risk of latent infection; it also successfully distinguishes between M. tuberculosis infection and BCG vaccination. This capability may facilitate tuberculosis control in nonendemic regions. PMID- 11282753 TI - A method for noninvasive determination of inspiratory resistance during proportional assist ventilation. AB - Currently available noninvasive methods for measuring inspiratory resistance (RI) are difficult to implement or interpret during assisted mechanical ventilation on account of the confounding effect of respiratory efforts (Pmus). We propose a simple method consisting of brief reductions in airway pressure (Paw) in the early part of the inflation phase (pulse). Paw, flow (V), and volume (V) are measured at the beginning of the pulse (T (0)), at the trough of the pulse (TI) and at a point 0.1 s before T(0) (T(-1)). Equations of motion of the form [Pmus + Paw = V. K(1) + V (2). K(2) +V. E] are generated for the data at the three time points (E = elastance, K(1) and K(2) are Rohrer's constants). These three equations can be solved for K(1) and K(2) if it is arranged that the pulse has appropriate configuration and timing, and if it is assumed that DeltaPmus/Deltat is constant over the brief pulse period. The method was tested in 67 patients ventilated with proportional assist ventilation (PAV). The results were compared with those obtained using the interrupter technique during a period of controlled mechanical ventilation (CMV). RI, expressed at a standard flow of 1 L. s(-)(1), was slightly higher during PAV (16.4 +/- 4.9 versus 15.5 +/- 4.5 cm H(2)O. L(-1). s, p < 0.001). The average difference was 0.9 +/- 2.0 cm H(2)O. L(-1). s, corresponding to 5.4 +/- 12.6% of the average of RCMV and RPAV. The correlation coefficient was 0.92 (p = 8E-28) with a slope (1.01) and intercept (0.8) not significantly different from 1.0 and 0, respectively. We conclude that brief negative pulses applied early during the inflation phase can be used to provide reliable estimates of inspiratory resistance during PAV. PMID- 11282754 TI - Glucagon-like peptide-1(7-36) amide stimulates surfactant secretion in human type II pneumocytes. AB - To determine the influence of glucagon-like peptides on the secretion of human pulmonary surfactant, we used human type II pneumocytes. In these cells, GLP-1(7 36) amide and exendin-4 stimulated phosphatidylcholine secretion (PC) and cAMP formation in a concentration-dependent manner; these effects were reversed by exendin(9-39). No changes were observed with other related peptides. The mechanism by which GLP-1(7-36) amide exerts its stimulatory effect was investigated with various agents that are well known to be stimulators or inhibitors of PC secretion. Thus, 8-bromo-cAMP increased and both Rp-cAMPS and H 89, the latter an inhibitor of protein kinase A (PKA), reduced pulmonary surfactant secretion in type II pneumocytes. Also, GLP-1(7-36) amide and TPA exerted additive effects in stimulating PC secretion, and Calph C, a potent inhibitor of protein kinase C (PKC), blocked most of the effect of GLP-1(7-36) amide. By contrast, both the calcium ionophore A23187 and GLP-1(7-36) amide had additive effects in increasing PC secretion, and the specific inhibitor of Ca(2+) calmodulin-dependent protein kinase (Ca-CM-PK), KN-62, inhibited the effect of A23187 but did not alter the stimulatory action of GLP-1(7-36) amide. Our findings suggest that both PKA and PKC are involved in the stimulatory effects of GLP-1(7-36) amide on PC secretion, whereas this peptide has no effect on PC secretion through a Ca-CM-PK mechanism. PMID- 11282755 TI - Longitudinal changes in pulmonary function and respiratory symptoms in cotton textile workers. A 15-yr follow-up study. AB - To evaluate the chronic effects of exposure to cotton dust, a 15-yr follow-up study in cotton textile workers was performed in Shanghai, China from 1981 to 1996. Testing occurred four times during the 15-yr period. The achieved follow-up rates were 76-88% of the original 447 cotton textile workers, and 70-85% of the original 472 silk textile workers (as a control group). Identical questionnaires, equipment, and methods were used throughout the study. The prevalence of byssinosis increased over time in cotton workers, with 15.3% at the last survey versus 7.6% at the baseline, whereas no byssinosis was found in silk workers. More workers in the cotton group consistently reported symptoms than in the silk group, although symptom reporting varied considerably from survey to survey. Cotton workers had small, but significantly greater, adjusted annual declines in FEV(1) and FVC than did the silk workers. Years worked in cotton mills, high level of exposure to endotoxin, and across-shift drops in FEV(1) were found to be significant determinants for longitudinal change in FEV(1), after controlling for appropriate confounders. Furthermore, there were statistically significant associations between excessive loss of FEV(1) and byssinosis, chest tightness at work, and chronic bronchitis in cotton workers. Workers who consistently (three or four of the surveys) reported byssinosis or chest tightness at work had a significantly greater 15-yr loss of FEV(1). We conclude that long-term exposure to cotton dust is associated with chronic or permanent obstructive impairments. Consistent reporting of respiratory symptoms, including byssinosis and chest tightness at work, is of value to predict the magnitude and severity of chronic impairments in textile workers. PMID- 11282757 TI - Effects of exercise on amino acid metabolism in patients with chronic obstructive pulmonary disease. AB - Depletion of fat-free mass (FFM) significantly contributes to decreased skeletal muscle weakness and impaired exercise capacity in patients with chronic obstructive pulmonary disease (COPD). FFM wasting suggests disturbances in intermediary metabolism, confirmed by data showing profound alterations in the skeletal muscle amino acid (AA) status in COPD at rest. To unravel whether there is a role for AAs in the mechanisms for skeletal muscle dysfunction in COPD, basic knowledge of AA metabolism in the muscle during exercise is important. We examined the effects of 20 min of exercise on AA metabolism in 14 patients with COPD and eight control subjects. Arterialized venous blood and a quadriceps femoris muscle biopsy were obtained before and immediately after exercise. FFM was not significantly different between the groups. In COPD, a significant reduction of most muscle AAs was present postexercise, whereas several plasma AAs were increased (p < 0.05). Consequently, sum AAs was reduced in muscle (20%; p < 0.01) and increased in plasma (16%, p < 0.05), suggesting an enhanced AA release from muscle in COPD during exercise. In the COPD group, the increase in plasma alanine and glutamine was even higher postexercise (61%, p < 0.01 and 21%, p < 0.01, respectively), suggesting enhanced nitrogen efflux. This study shows that exercise alters amino acid (intermediary) metabolism in patients with COPD and independent of the presence of FFM wasting. PMID- 11282756 TI - Increased nitrosothiols in exhaled breath condensate in inflammatory airway diseases. AB - Nitrosothiols (RS-NOs) are formed by interaction of nitric oxide (NO) with glutathione and may limit the detrimental effect of NO. Because NO generation is increased in airway inflammation, we have measured RS-NOs in exhaled breath condensate in patients with asthma, cystic fibrosis, or chronic obstructive pulmonary disease (COPD). We also measured exhaled NO and nitrite (NO(2-)) in the same subjects. RS-NOs were detectable in exhaled breath condensate of all subjects. RS-NOs were higher in subjects with severe asthma (0.81 +/- 0.06 microM) when compared with normal control subjects (0.11 +/- 0.02 microM, p < 0.01) and with subjects with mild asthma (0.08 +/- 0.01 microM, p < 0.01). Elevated RS-NOs values were also found in patients with cystic fibrosis (0.35 +/- 0.07 microM, p < 0.01), in those with COPD (0.24 +/- 0.04 microM, p < 0.01) and in smokers (0.46 +/- 0.09 microM, p < 0.01). In current smokers there was a correlation (r = 0.8, p < 0.05) between RS-NOs values and smoking history (pack/year). We also found elevated concentrations of NO(2-) in patients with severe asthma, cystic fibrosis, or COPD, but not in smokers or patients with mild asthma. This suggests that exhaled NO(2-) is less sensitive than exhaled RS-NOs. This study has shown that RS-NOs are detectable in exhaled breath condensate of healthy subjects and are increased in patients with inflammatory airway diseases. As RS-NOs concentrations in exhaled breath condensate vary in the different airway diseases and increase with the severity of asthma, their measurement may have clinical relevance as a noninvasive biomarker of nitrosative stress. PMID- 11282759 TI - Noninvasive versus conventional mechanical ventilation. An epidemiologic survey. AB - A prospective survey was performed over a period of 3 wk among 42 intensive care units to assess the incidence of use and effectiveness of noninvasive mechanical ventilation (NIV) in clinical practice. All patients requiring ventilatory support for acute respiratory failure (ARF), either with endotracheal intubation (ETI) or NIV, were included. Ventilatory support was required in 689 patients, 581 with ETI and 108 (16%) with NIV (35% of patients not intubated on admission). Reasons for mechanical ventilation were coma (30%), cardiogenic pulmonary edema (7%), and hypoxemic (48%) and hypercapnic ARF (15%). NIV was never used for patients in coma (who were excluded from further analysis), but was used in 14% of patients with hypoxemic ARF, in 27% of those with pulmonary edema, and in 50% of those with hypercapnic ARF. NIV was followed by ETI in 40% of cases. The incidence of both nosocomial pneumonia (10% versus 19%, p = 0.03), and mortality (22% versus 41%, p < 0.001) was lower in NIV patients than in those with ETI. After adjusting for differences at baseline, Simplified Acute Physiology Score (SAPS) II (odds ratio [OR] = 1.05 per point; confidence interval [CI]: 1.04 to 1.06), McCabe/Jackson score (OR = 2.18; CI: 1.57 to 3.03), and hypoxemic ARF (OR = 2.30; CI: 1.33 to 4.01) were identified as risk factors explaining mortality; success of NIV was associated with a lower risk of pneumonia (OR = 0.06; CI: 0.01 to 0.45) and of death (OR = 0.16; CI: 0.05 to 0.54). In NIV patients, SAPS II and a poor clinical tolerance predicted secondary ETI. Failure of NIV was associated with a longer length of stay. In conclusion, NIV can be successful in selected patients, and is associated with a lower risk of pneumonia and death than is ETI. PMID- 11282758 TI - Forced expiratory flow in uninfected infants and children born to HIV-infected mothers. AB - The Pediatric Pulmonary and Cardiovascular Complications of Vertically Transmitted HIV (P(2)C(2) HIV) Study is a multicenter study examining pulmonary and cardiac outcomes in offspring of HIV-infected mothers. This portion of the P(2)C(2) study tests the hypothesis that infants exposed to, but uninfected by, maternal HIV have normal maximal expiratory flow at functional residual capacity (V'max,(FRC)). We obtained 500 measurements of V'max,(FRC) by rapid thoracic compression in 285 children ages 6-30 mo in five U.S. centers. The data were compared with those from a healthy cohort of children described elsewhere. V'max,(FRC) rose with height in a linear relationship. The slope of the regression line in the exposed infants did not differ statistically from the slope in the comparison group, but the intercept was about 20% lower (p < 0.001). Height and weight were comparable in the two cohorts, and the differences between intercepts persisted after adjusting for birth weight and gestational age. However, maternal HIV infection cannot be assumed to be the cause as the cohorts may have differed in other variables, such as socioeconomic status and frequency of maternal smoking and drug use. Also, measurements varied substantially within and between our five centers, probably in part because of different racial and ethnic distributions. In summary, maternal HIV infection probably has only a modest effect, if any, on maximal expiratory flow at functional residual capacity in uninfected infants. PMID- 11282760 TI - The effect of salmeterol on markers of airway inflammation following segmental allergen challenge. AB - Inflammation is a critical component of asthma. Drugs that control asthma generally reduce the degree of airway inflammation. There is theoretical controversy surrounding the effects of beta(2)-agonists on airway inflammation, with some studies suggesting an anti-inflammatory effect, and others predicting a proinflammatory influence. We conducted a double-blind, placebo-controlled, crossover study of the effect of the long-acting beta(2)-agonist salmeterol on airway inflammation induced by segmental allergen challenge (SAC). We studied 13 allergic asthmatics controlled with as needed inhaled short-acting beta(2) agonists alone, and used bronchoalveolar lavage 5 min and 48 h after SAC to assess airway inflammation, and the effects of salmeterol on this process. Salmeterol therapy improved FEV(1), but had no significant effect on the immediate or late cellular response to SAC. One measure of superoxide production was reduced, and interleukin-4 (IL-4) was reduced in baseline samples, but other indices of airway inflammation were unchanged by salmeterol therapy. We conclude that salmeterol therapy alone does not meaningfully reduce airway inflammation induced by SAC, but equally importantly, does not result in amplified inflammation. PMID- 11282761 TI - Short-term oral administration of L-arginine improves hemodynamics and exercise capacity in patients with precapillary pulmonary hypertension. AB - We sought to assess the effects of oral supplementation of L-arginine, the precursor of nitric oxide (NO), on hemodynamics and exercise capacity in patients with pulmonary hypertension. Acute hemodynamic responses to oral L-arginine (0.5 g/10 kg body weight) or placebo were examined in 19 patients with primary or precapillary secondary pulmonary hypertension. Cardiopulmonary exercise tests were performed to measure peak oxygen consumption (peak V O(2)) and the ventilatory response to carbon dioxide production (V E-V CO(2) slope) before and 1 wk after treatment with L-arginine (1.5 g/10 kg body weight/d) or placebo. Oral supplementation of L-arginine significantly increased plasma L-citrulline, which indicated enhancement of NO production. Supplemental L-arginine produced a 9% decrease in mean pulmonary arterial pressure (53 +/- 4 to 48 +/- 4 mm Hg, p < 0.05) and a 16% decrease in pulmonary vascular resistance (14.8 +/- 1.5 to 12.4 +/- 1.4 Wood units, p < 0.05). L-arginine modestly decreased mean systemic arterial pressure (92 +/- 4 to 87 +/- 3 mm Hg, p < 0.05). A 1-wk supplementation of L-arginine resulted in a slight increase in peak V O(2) (831 +/- 88 to 896 +/- 92 ml/min, p < 0.05) and a significant decrease in the V E- V CO(2) slope (43 +/- 4 to 37 +/- 3, p < 0.05) without significant systemic hypotension. Hemodynamics and exercise capacity remained unchanged during placebo administration. These results suggest that oral supplementation of L-arginine may have beneficial effects on hemodynamics and exercise capacity in patients with precapillary pulmonary hypertension. PMID- 11282762 TI - Effects of hyperoxia on ventilatory limitation during exercise in advanced chronic obstructive pulmonary disease. AB - We studied interrelationships between exercise endurance, ventilatory demand, operational lung volumes, and dyspnea during acute hyperoxia in ventilatory limited patients with advanced chronic obstructive pulmonary disease (COPD). Eleven patients with COPD (FEV(1.0) = 31 +/- 3% predicted, mean +/- SEM) and chronic respiratory failure (Pa(O(2)) 52 +/- 2 mm Hg, Pa(CO(2 ))48 +/- 2 mm Hg) breathed room air (RA) or 60% O(2) during two cycle exercise tests at 50% of their maximal exercise capacity, in randomized order. Endurance time (T(lim)), dyspnea intensity (Borg Scale), ventilation (V E), breathing pattern, dynamic inspiratory capacity (IC(dyn)), and gas exchange were compared. Pa(O(2)) at end exercise was 46 +/- 3 and 245 +/- 10 mm Hg during RA and O(2), respectively. During O(2), T(lim) increased 4.7 +/- 1.4 min (p < 0.001); slopes of Borg, V E, V CO(2), and lactate over time fell (p < 0.05); slopes of Borg-V E, V E-V CO(2), V E-lactate were unchanged. At a standardized time near end-exercise, O(2) reduced dyspnea 2.0 +/- 0.5 Borg units, V CO(2) 0.06 +/- 0.03 L/min, V E 2.8 +/- 1.0 L/min, and breathing frequency 4.4 +/- 1.1 breaths/min (p < 0.05 each). IC(dyn) and inspiratory reserve volume (IRV) increased throughout exercise with O(2) (p < 0.05). Increased IC(dyn) was explained by the combination of increased resting IRV and decreased exercise breathing frequency (r(2) = 0.83, p < 0.0005). In conclusion, improved exercise endurance during hyperoxia was explained, in part, by a combination of reduced ventilatory demand, improved operational lung volumes, and dyspnea alleviation. PMID- 11282763 TI - Incidence and host determinants of probable occupational asthma in apprentices exposed to laboratory animals. AB - Laboratory animal (LA) workers are frequently affected with allergic sensitization and occupational asthma (OA). The role of preexposure host factors, in particular airway responsiveness, on the incidence of OA has not been satisfactorily studied. A prospective cohort study of 417 apprentices in animal health technology was conducted to investigate the incidence and determinants of probable OA. Questionnaire and skin-prick tests with common and work-specific allergens were administered on entry and at follow-up visits (up to three) from 8 to 44 mo after starting apprenticeship. Responsiveness to inhaled methacholine was assessed at baseline and at follow-up in apprentices who developed a new specific skin sensitization and in control subjects. Preexposure host characteristics and the school attended were compared between cases and all cohort subjects not meeting the criteria for probable OA. Twenty-eight apprentices satisfied the definition for probable OA, i.e., onset of immediate skin reactivity to > 1 occupational inhalant and > 3.2-fold decrease in the provocative concentration causing a 20% reduction in FEV(1) (PC(20)). The incidence of probable OA was 2.7% (28/1,043 person-years). Baseline immediate skin reactivity to pets (rate ratio [RR] 4.1, 95% confidence interval [CI] = 1.6 to 10.8), and bronchial responsiveness (PC(20) < or = 32 versus PC(20) > 32 mg/ ml) (RR = 2.5, 95% CI = 1.0 to 5.8) were associated with an increased risk of probable OA; a lower FEV(1) had an apparent, protective effect (RR = 0.58, 95% CI = 0.43 to 0.78). It is concluded that apprentices in animal health show a high incidence of probable OA, and that preexposure airway caliber and responsiveness as well as sensitization to pets are associated with an increased risk. PMID- 11282764 TI - Changes in respiratory effort sensation over time are linked to the frequency content of diaphragm electrical activity. AB - This study evaluated whether respiratory effort sensation (RES) changes over time when breathing is performed with constant contraction pattern, fixed diaphragm activation, and maintained pressure generation. Another aim was to assess whether there was any association between RES and the power spectrum center frequency of the diaphragm (CFdi) electrical activity. Six healthy subjects performed two 10 min periods targeting diaphragm electrical activation (EAdi) to 40% of maximum using (1) expulsive or Mueller maneuvers at FRC generating a mean transdiaphragmatic (Pdi) pressure of 55.0 +/- 22.7 cm H(2)O (+/- SD) and (2) inspiration to 71.2 +/- 14.1% of inspiratory capacity (IC) generating a Pdi of 21.4 +/- 5.2 cm H(2)O. The Pdi did not decrease over time during either maneuver. During both periods RES increased (p < 0.001) and CFdi decreased (p < 0.001) over time with higher Pdi levels producing larger decreases in CFdi (p = 0.003) and greater increases in RES (p = 0.008). Changes in CFdi and RES were related, and identical slopes were obtained during the two maneuvers. In conclusion, while breathing with a fixed pattern, constant diaphragm activation, and maintained pressure generation, RES increases over time and is associated with CFdi independent of the level of diaphragm pressure generated. PMID- 11282765 TI - Effect of nasal continuous positive airway pressure on neuropsychological function in sleep apnea-hypopnea syndrome. A randomized, placebo-controlled trial. AB - A placebo-controlled, partial cross-over, double-blind, randomized study was performed with 46 adults with sleep apnea-hypopnea syndrome (SAHS) to determine the effect of therapeutic and subtherapeutic (0-1 cm H(2)O) nasal continuous positive airway pressure (CPAP) treatment on polysomnographic and neuropsychological testing. The following neuropsychological tests were administered: Geriatric Depression Scale, Trail Making A and B, Digit Span Test Forward and Backward, Epworth Sleepiness Scale, SteerClear, Digit Symbol, Controlled Oral Word Association, and Complex Figure Recall. Compared with results without CPAP, subtherapeutic CPAP did not affect any measured polysomnographic parameter. Comparison of neuropsychological test results obtained between the initial periods of effective treatment (Group 1, 16.1 d; Group 2, 19.6 d; p = NS) in all subjects showed significant improvements in Digit Symbol, Digit Span Backward, and Complex Figure tests. However, there were no group differences in changes in test results during the period when one group was on effective CPAP and the other on ineffective CPAP (Group 1, 16.1 d; Group 2, 13.9 d; p = NS). The results indicate the feasibility and importance of using ineffective CPAP as a placebo treatment and the importance of including a placebo control in studies evaluating the effect of treatment on neuropsychological function in SAHS. PMID- 11282766 TI - Randomized, double-blind, placebo-controlled crossover trial of modafinil in the treatment of residual excessive daytime sleepiness in the sleep apnea/hypopnea syndrome. AB - Some patients with the sleep apnea/hypopnea syndrome (SAHS) remain subjectively and objectively sleepy despite using effective continuous positive airway pressure (CPAP) therapy. The aim of this single center study was to determine the efficacy and safety of the novel wake-promoting medication modafinil in the treatment of CPAP-resistant daytime sleepiness. Thirty sleep apneics receiving effective CPAP therapy (objective use, 6.5 +/- 1.1 h/night) received daily single doses of 400 mg modafinil or placebo for 2 wk in a double-blind randomized crossover design. Outcome measures were assessed at baseline and at the end of both 2-wk treatment periods. Treatment periods were separated by a 1-wk washout. Modafinil had no effect on sleepiness as measured by the Epworth Sleepiness Scale or the Multiple Sleep Latency Test (p > 0.1); however, significant improvements in alertness were found with the Maintenance of Wakefulness Test (modafinil 18.3 +/- 3.9 min; placebo, 16.6 +/- 5.0 min; p < 0.02). No significant treatment related improvements in cognitive performance or quality of life were found with modafinil (all p > 0.05). There was a significant reduction in CPAP use on modafinil compared to placebo (6.3 +/- 1 h/ night; 6.5 +/- 1, p = 0.03). This study suggests that modafinil may improve some aspects of alertness in patients with SAHS who remain sleepy during CPAP therapy, but further studies are required to assess the significance of the reduction in CPAP use. PMID- 11282767 TI - Clinical predictors of health-related quality of life depend on asthma severity. AB - The National Asthma Education and Prevention Program guidelines define asthma severity before treatment by lung function and symptoms. It has been assumed, but not demonstrated, that improvement in these measures would translate into improvement in health-related quality of life (HRQL). Because HRQL is an important outcome in asthma management, we asked what are the determinants of HRQL? To address this question, we retrospectively analyzed HRQL data, as measured by the Juniper Asthma Quality of Life Questionnaire, in subjects with mild versus moderate-severe asthma from two clinical trials. We examined whether these traditional clinical outcomes have different relationships to HRQL depending on asthma severity. We also assessed whether the relationship between clinical outcomes and HRQL in subjects with moderate-severe asthma would change when subjects improved to mild-moderate disease with controller medication treatment. Lung function was not an independent predictor or determinant of HRQL at any level of asthma severity, whereas intensity of shortness of breath predicted HRQL at all levels of asthma severity. Rescue beta-agonist use independently predicted HRQL in subjects with mild asthma, but not in those with moderate-severe asthma. In subjects with moderate-severe asthma who improved to mild-moderate disease with controller treatment, rescue beta-agonist use predicted HRQL. We conclude that the independent determinants of HRQL vary according to asthma severity and change with asthma treatment. PMID- 11282768 TI - Effect of pulmonary rehabilitation on quadriceps fatiguability during exercise. AB - We have recently shown that patients with chronic obstructive pulmonary disease (COPD) develop contractile fatigue of their quadriceps muscle following endurance exercise. Pulmonary rehabilitation can produce physiological adaptations in patients with COPD. We hypothesized that if pulmonary rehabilitation induces physiological adaptations in the exercising muscle, it should become more fatigue resistant. Twenty one patients with COPD, mean age 69.9 +/- 1.9 yr, FEV(1) 45 +/- 4% predicted, participated in an 8-wk outpatient, supervised pulmonary rehabilitation exercise program. Quadriceps contractile fatigue was detected by a fall in quadriceps twitch force postexercise. Twitch force was measured during magnetic stimulation of the femoral nerve. Because potentiated twitches may be more sensitive at detecting fatigue, both unpotentiated (TwQu) and potentiated (TwQp) twitches were obtained before and 10, 30, and 60 min after constant load cycle exercise. Prerehabilitation, during constant load exercise, patients exercised at 37 +/- 4 W for 11.2 +/- 1.8 min. Prerehabilitation, TwQu fell significantly postexercise down to a minimum value of 82.5 +/- 3.1% of the baseline preexercise value (p < 0.001). Similarly, prerehabilitation, TwQp fell significantly postexercise down to a minimum value of 73.9 +/- 3.9% of baseline (p < 0.001). Postrehabilitation, for the same intensity and duration of exercise, TwQu was not significantly different from baseline at any time postexercise. Postrehabilitation, TwQp fell significantly postexercise but the fall in TwQp with exercise was significantly less postrehabilitation compared with prerehabilitation (p < 0.001). In conclusion, pulmonary rehabilitation resulted in increased fatigue resistance of the quadriceps muscle in patients with COPD. PMID- 11282769 TI - Exercise capacity predicts health status in alpha(1)-antitrypsin deficiency. AB - Resting lung function is only weakly related to health status in chronic obstructive pulmonary disease, reflecting the multifactorial causes of impairment and the heterogeneous nature of the condition. The current study examined whether density mask analysis of high-resolution computed tomography (HRCT) or exercise capacity were better surrogates for health status in a well-defined, homogeneous group of patients with alpha(1)-antitrypsin deficiency (PiZ). Twenty-nine patients with predominantly lower zone emphysema on HRCT were studied. Exercise was assessed by incremental treadmill (V O(2) peak) and shuttle walking tests (ISWT) and health status by the St. George's Respiratory Questionnaire (SGRQ) and SF-36. Although lower zone expiratory HRCT was related to exercise capacity (rho = -0.64 and -0.63 for V O(2) peak and ISWT, respectively, p < 0.001), multiple regression analysis suggested that FEV(1) was a marginally better predictor (rho = -0.64 and -0.65, p < 0.001). HRCT also related significantly to health status (rho = -0.37 for SGRQ activity, p < 0.05), although again FEV(1) showed a stronger relationship (rho = -0.43, p = 0.01). However, exercise capacity was the best predictor of health status with the ISWT accounting for up to 55% of the variability seen in SGRQ total and up to 53% of the SF-36 domain scores (physical functioning). Although both HRCT and lung function relate to health status, exercise capacity is the best predictor of patients disability in these patients with predominantly lower zone emphysema. PMID- 11282770 TI - Development and testing of formal protocols for oxygen prescribing. AB - The absence of standardized assessment protocols with well- defined measurement properties limits comparison of outcomes among those receiving long-term oxygen therapy (LTOT). We describe simple protocols for a hospital test, a simulated home test, and an actual home test, their reliability and relationship to each other. Stable patients with exercise hypoxemia participated. In 74 patients who completed four exercise tests, correlations between tests ranged from 0.85 to 0.78. Of these 27.0% had the same prescription from all four tests. In 46% prescriptions were within 1 L/ min and in 27% within 2 L/min. During exercise the hospital tests suggested slightly higher oxygen prescriptions than did the simulated home tests (2.5 L/min versus 2.0 L/min, p < 0.001). In 23 patients who participated in actual home assessments, the correlations between the home test, the hospital, and the simulated home tests were 0.22 (95% CI -0.24 to 0.67) and 0.27 (95% CI -0.18 to 0.72). In conclusion, standardizing tests for the assessment of LTOT is important. We describe simple hospital and simulated home tests that are reproducible, easy to carry out, and correlate well with each other. PMID- 11282771 TI - The relationship between craniofacial morphology and obstructive sleep apnea in whites and in African-Americans. AB - Previous studies of craniofacial risk factors for obstructive sleep apnea (OSA) have been based predominantly on cephalometry. However, differences in head form (measured by the cranial index [CI]) and facial form (measured by the facial index [FI]) are considered by anthropologists to provide a basis for structural variation in craniofacial anatomy. We assessed the association of head and facial form with the apnea hypopnea index (AHI) in 364 white individuals and 165 African Americans. Data collected included cranial and facial dimensions (using anthropometric calipers), body mass index (BMI), neck circumference, and the AHI. CI and FI differed for whites with OSA (AHI > or = 15) versus those without OSA (AHI < 5) (increased CI and decreased FI in subjects with OSA, p = 0.005 and p = 0.006, respectively). CI and FI did not differ in OSA versus non-OSA groups of African-Americans. In subjects with OSA, the CI in whites was again greater and the FI smaller than those in African-Americans (p = 0.007 and p = 0.004, for CI and FI.) We conclude that brachycephaly is associated with an increased AHI in whites but not in African-Americans. The CI may useful in phenotyping and identifying population subsets with OSA. PMID- 11282772 TI - Neural substrates for the perception of acutely induced dyspnea. AB - Little is currently known about the brain regions involved in central processing of dyspnea. We performed a functional imaging study with positron emission tomography (PET) to assess brain activation associated with an important component of dyspnea, respiratory discomfort during loaded breathing. We induced respiratory discomfort in eight healthy volunteers by adding external resistive loads during inspiration and expiration. Brain activation was characterized by a significant increase in regional cerebral blood flow (rCBF) (Z score of peak activation > 3.09). As compared with the unloaded control condition, high loaded breathing was associated with neural activation in three distinct brain regions, the right anterior insula, the cerebellar vermis, and the medial pons (respective Z scores = 4.75, 4.44, 4.41). For these brain regions, we further identified a positive correlation between rCBF and the perceived intensity of respiratory discomfort (respective Z scores = 4.45, 4.75, 4.74) as well as between rCBF and the mean amplitude of mouth pressure swings (DeltaPm), the index of the main generating mechanism of the sensation (respective Z scores = 4.67, 4.36, 4.31), suggesting a common activation by these two parameters. Furthermore, we identified an area in the right posterior cingulate cortex where neural activation was specifically associated with perceived intensity of respiratory discomfort that is not related to DeltaPm (Z score = 4.25). Our results suggest that respiratory discomfort related to loaded breathing may be subserved by two distinct neural networks, the first being involved in the concomitant processing of the genesis and perception of respiratory discomfort and the second in the modulation of perceived intensity of the sensation by various factors other than its main generating mechanism, which may include emotional processing. PMID- 11282773 TI - Mechanical ventilation-induced air-space enlargement during experimental pneumonia in piglets. AB - Mechanical ventilation-induced air-space enlargement was investigated in a porcine model of multifocal pneumonia. Following the intrabronchial inoculation of Escherichia coli, 9 piglets (22 +/- 2 kg) were ventilated with a tidal volume (VT) of 15 ml/kg for 43 +/- 15 h. Five noninoculated piglets ventilated for 60 h with the same VT served as control animals. Following death, the lungs were fixed and lung morphometry was assessed. In inoculated animals, unventilated infected and normally ventilated noninfected pulmonary lobules coexisted. In normally ventilated lung regions (1) emphysema-like lesions were present, (2) mean alveolar area and mean linear intercept were significantly greater in inoculated than in control animals, and (3) the degree of alveolar distension correlated with the decrease in respiratory compliance. In unventilated lung areas (1) pseudocysts were frequent, (2) alveolar edema was rare, (3) bronchiolectasis was frequent, (4) mean bronchiolar area was greater in inoculated than in control animals, and (5) the degree of bronchiolar distension correlated with the increase in inspiratory plateau pressure. In conclusion, in piglets with severe bronchopneumonia, air-space enlargement rather than pulmonary edema was the major feature of mechanical ventilation-induced lung barotrauma and resembled lesions previously reported in critically ill patients ventilated using high inspiratory pressures. PMID- 11282775 TI - Interleukin-6 changes deformability of neutrophils and induces their sequestration in the lung. AB - Interleukin-6 (IL-6) is an important mediator of both the hepatic and the bone marrow components of the acute-phase response. Previous studies from our laboratory have shown that cells released into the circulation from the marrow preferentially sequester in the lung. The present study was designed to examine the mechanism of this sequestration using a single dose of recombinant human IL-6 to stimulate the marrow in rabbits. Marrow release was monitored by labeling polymorphonuclear leukocyte (PMN) precursors in the marrow with the thymidine analogue, 5'-bromo-2-deoxyuridine (BrdU), 24 h before IL-6 treatment. This treatment caused a neutrophilia that was associated with the increase of circulating BrdU- labeled PMN (PMN(BrdU)) and morphometric studies confirmed that PMN(BrdU) released from the marrow preferentially sequestered in the lung microvessels compared to unlabeled PMN. IL-6 treatment increases PMN F-actin content (p < 0.05) that was not due to cell activation by IL-6. In vitro studies show that IL-6 treatment decreased the deformability of circulating PMN (p < 0.05). These studies confirm that IL-6 treatment causes an accelerated release of PMN from the bone marrow and shows that these newly released PMN have high levels of F-actin, are less deformable, and preferentially sequester in lung microvessels. PMID- 11282774 TI - Association of a promoter polymorphism of the CD14 gene and atopy. AB - Atopy is generally considered to be caused by interaction of genetic and environmental factors. Recently, an association of a C-to-T transition in the promoter region of the CD14 gene on chromosome 5q31.1 and atopic phenotypes was reported in a population study of school children in the United States. The aim of the present study was to investigate the association of the C allele of the CD14/-159 with phenotypes of atopy and asthma in an adult Dutch population in which linkage of total serum IgE and bronchial hyperresponsiveness to chromosome 5q31-33 is present. We studied 159 probands with asthma and 158 spouses as controls. Phenotypes for asthma (e.g., bronchial hyperresponsiveness, physician's diagnosis) and for atopy (e.g., total serum IgE level, intracutaneous skin test, allergic rhinitis) were studied. In this population, homozygotes for the C allele had a higher number of positive skin tests and higher total serum IgE levels (in skin test-positive individuals) and subsequently, more self-reported allergic symptoms including rhinitis and hay fever, compared with subjects with CT and TT alleles. We conclude that the -159 C-to-T promoter polymorphism in the CD14 gene may result in expression of a more severe allergic phenotype. PMID- 11282776 TI - Lipopolysaccharide-induced diaphragmatic contractile dysfunction and sarcolemmal injury in mice lacking the neuronal nitric oxide synthase. AB - In this study we evaluated the role of the neuronal nitric oxide synthase (nNOS) in lipopolysaccharide (LPS)-induced diaphragmatic contractile dysfunction and sarcolemmal injury. Wild-type (WT) mice or mice deficient in the nNOS gene (nNOS( /-)) were injected with either saline (control) or Escherichia coli LPS (LPS groups) and sacrificed 12 h later. The diaphragm was then examined for NOS expression, NOS activity, and in-vitro contractility. We also assessed sarcolemmal injury in isolated muscle strips under resting condition and after 3 min of artificial stimulations. In WT mice, LPS injection reduced maximum force to about 75% of that of control animals and raised total NOS activity significantly due to the induction of the iNOS isoform. Although muscle fiber injury was minimal under resting condition, the percentage of injured fibers in control and LPS-injected mice approached 27% and 40% of total fibers, respectively, in response to artificial stimulation. By comparison, LPS injection in nNOS(-/-) mice elicited a worsening of muscle contractility (maximum force < 60% of control animals) but elicited degrees of sarcolemmal injury similar to those observed in the WT animals. In addition, muscle NOS activity and iNOS protein level in nNOS(-/-) mice injected with LPS reached about 10% and 60% of that of WT animals, respectively (p < 0.05 compared with WT animals). Protein level of endothelial NOS isoform in the diaphragm was not altered by LPS injection in either WT or nNOS(-/-) animals. We conclude that nNOS plays a protective role in attenuating the negative influence of sepsis on diaphragmatic contractility but is not involved in the pathogenesis of sepsis-induced sarcolemmal injury. PMID- 11282777 TI - The effect of aging on nasal mucociliary clearance, beat frequency, and ultrastructure of respiratory cilia. AB - The increased susceptibility of the elderly to lower respiratory tract infection cannot be fully explained. Although mucociliary clearance, which is affected by ciliary beating and ultrastructure, plays a crucial role in the defense of the airways against inhaled microbes, little is known of the effects of aging on these parameters. We studied the nasal mucociliary clearance (NMCC) time, ciliary beat frequency, and ultrastructure of respiratory cilia in a cohort of healthy volunteers (age range 11 to 90 yr). Ciliary beat frequency of ciliated nasal epithelial cells was obtained via an established photometric method, and NMCC time was measured with the saccharine test. There was a correlation of ciliary beat frequency (r = -0.48, p = 0.0001) and NMCC time r = 0.64, p < 0.001) with increasing age. Transmission electron microscopy revealed an increase in the percent of subjects exhibiting microtubular disarrangement and single central microtubules with aging (p = 0.002 and p = 0.005, respectively). Subjects older than 40 yr of age had significantly slower ciliary beat frequency, higher percent of ciliary cross-sections displaying single tubules, and longer NMCC time than their younger counterparts (p < 0.05). These findings may help explain the frequent occurrence of respiratory infection in the elderly. PMID- 11282778 TI - Absorption, distribution, metabolism, and excretion of a respirable antisense oligonucleotide for asthma. AB - EPI-2010 is a respirable antisense oligonucleotide (RASON), which selectively attenuates discordantly overexpressed adenosine A(1) receptors in allergic lung (Nature 1997;385:721). In the present study, aerosolized [(35)S]-labeled EPI-2010 (5 mg exposure; specific activity 0.055 Ci/mmol) was administered to normal rabbits by endotracheal tube to assess biodistribution, route of elimination, and potential cardiovascular toxicity. The animals were killed at 0, 6, 24, 48, and 72 h after inhalation of EPI-2010. Duplicate aliquots from different tissues and samples were solubilized and assessed for radioactivity. Approximately 1.4% of the total aerosolized EPI-2010 was deposited into the lung. The concentration of the drug in the lung at 0, 6, 24, 48, and 72 h was 64.0 +/- 1.5, 67.0 +/- 4.4, 32.0 +/- 3.7, 23.4 +/- 1.4, and 2.1 +/- 0.5 microg equivalents, respectively. Only a small amount of the radioactivity was detected in extrapulmonary tissues. By 72 h, 67.5% of the administered dose was excreted in the urine, which represented the major pathway of elimination. In postlabeling studies, intact full-length EPI-2010 could only be detected in the lung. Autoradiographic analysis after inhalation of [(35)S]-labeled EPI-2010 showed a relatively uniform deposition of drug throughout the lung. The aerosolized EPI-2010 did not have any significant systemic effects on the cardiovascular system as determined by Cardiomax-II analysis. This pattern of distribution and the lack of effect on cardiovascular function support the concept that RASONs offer the potential to safely address respiratory targets for which systemic distribution and systemic bioavailability may be contraindicated. PMID- 11282779 TI - High-resolution computed tomographic evaluation of airway distensibility and the effects of lung inflation on airway caliber in healthy subjects and individuals with asthma. AB - The effects of a deep inspiration (DI) in individuals with asthma differ from those observed in healthy subjects. It has been postulated that the beneficial effect of lung inflation is mediated by airway stretch. One hypothesis to explain the defects in the function of lung inflation in asthma is that a DI may be unable to stretch the airways. This may result from attenuation of the tethering forces between the airways and the surrounding parenchyma. In the current study, we used high-resolution computed tomography (HRCT) to examine the ability of a DI to distend the airways of subjects with asthma (n = 10) compared with healthy subjects (n = 9) at baseline and after increasing airway tone with methacholine (MCh). We found that both at baseline and after the induction of smooth muscle tone with MCh, a DI distended the airways of healthy and asthmatic subjects to a similar extent, indicating that abnormal interdependence between the lung parenchyma and the airways is unlikely to play a major role in the loss or attenuation of the beneficial effect of lung inflation that characterizes asthma. Furthermore, we observed that after constriction had already been induced by MCh, following a DI, bronchodilation occurred in the healthy subjects but further bronchoconstriction occurred in the subjects with asthma. Our findings suggest that an abnormal excitation contraction mechanism in the airway smooth muscle of subjects with mild asthma counteracts the bronchodilatory effect of a DI. Therefore, the mechanism for reduced bronchodilation after DIs in subjects with mild asthma could be intrinsic to the airway smooth muscle. PMID- 11282780 TI - Liver-lung interactions following Escherichia coli bacteremic sepsis and secondary hepatic ischemia/reperfusion injury. AB - We hypothesized that ischemia/reperfusion (I/R) injury of the liver during normotensive gram-negative bacteremic sepsis alters the kinetics of circulating endotoxin, tumor necrosis factor-alpha (TNF-alpha), and coinduced mediators, thereby exacerbating sepsis-induced lung inflammation. Liver and lung dysfunction were studied after hematogenous infection of Sprague-Dawley rats with 10(9) Escherichia coli serotype O55:B5 (EC) and 90 min of secondary hepatic ischemia in EC + I/R and saline-infused (normal saline NS) x I/R rats, followed by brief (1 h) or longer reperfusion (24 h). TNF- alpha:leukotriene interactions in this model were examined using the 5-lipoxygenase-activating protein inhibitor MK-886. Compared with sham-operated EC + Sham animals, peak serum endotoxin, TNF-alpha, alanine aminotransferase, interleukin-6 (IL-6), and hepatic neutrophil (PMN) influx were higher in EC + I/R rats through 24 h (p < 0.05) despite comparable arterial pressure. Lung PMN influx and wet/dry weight ratios were likewise enhanced in EC + I/R versus EC + Sham or NS + I/R rats. MK-886 attenuated TNF alpha concentrations and ischemic liver injury but not mortality. Thus, focal hepatic I/R augments circulating endotoxin, TNF-alpha, and postbacteremic lung inflammation early after normotensive E. coli bacteremic sepsis. PMID- 11282782 TI - Effect of edentulism on spirometric tests. AB - The impact of denture wear in edentulous subjects while performing routine spirometric measurements has never been systematically investigated. We compared the values of FVC, FEV(1), PEFR, FEF(50%), FIV(1), and FIF(50%) recorded with and without dentures in three groups of edentulous subjects: 36 asymptomatic subjects with normal spirometry (N), 22 patients with chronic obstructive pulmonary disease (COPD), and 18 with interstitial lung disease (ILD). In 14 subjects retropharyngeal space with and without dentures was assessed by cephalometry. Subjects with N and ILD had significantly lower airflow rates without dentures, whereas subjects with COPD had no significant difference in spirometric values recorded with or without dentures. The retropharyngeal space was significantly decreased by removing dentures (from 1.52 +/- 0.07 to 1.16 +/- 0.09 cm, SEM, p < 0.0001). These findings indicate that in edentulous subjects with a normal or restrictive pattern, the recording of flow-volume curves with or without dentures produces small but significant differences. Although such differences do not appear to have clinical significance, the fact that when dentures are used some respiratory flows are higher would favor the use of dentures in edentulous subjects during spirometric evaluation. PMID- 11282781 TI - Mechanisms of interleukin 1beta-induced human airway smooth muscle hyporesponsiveness to histamine. Involvement of p38 MAPK NF-kappaB. AB - We have investigated the effect of IL-1beta on histamine H(1)-receptor (H(1)R) mediated inositol phosphate (IP) accumulation in human airway smooth muscle cells (HASMC) and on histamine-induced contraction of human bronchial rings. Stimulation of HASMC for 24 h with IL-1beta resulted in significant loss of histamine-induced IP formation, which was associated with a reduction of histamine- induced contraction of IL-1beta-treated human bronchial rings. An inhibitor of NF-kappaB activation, pyrrolidine dithiocarbamate, and a p38 MAPK inhibitor, blocked the IL-1beta-induced H(1)R desensitization, whereas anisomycin, an SAPK/JNK and p38 MAPK activator, mimicked the effect of IL-1beta. IL-1beta has been demonstrated to induce cox-2 expression and PGE(2) synthesis. In our study, indomethacin a cox antagonist, completely inhibited the effect of IL-1beta on H(1)R, whereas exogenously added PGE(2) was able to desensitize H(1)R. Furthermore, H-89, a selective PKA inhibitor, antagonized the effect of IL 1beta. Here, we have demonstrated that IL-1beta desensitizes H(1)R, which involves the activation of p38 MAPK and NF-kappaB, leading to the expression of cox-2 and the synthesis of PGE(2). PGE(2) increases intracellular cAMP resulting in PKA activation, which phosphorylates and functionally uncouples H(1)R. Our results suggest that IL-1beta protects airway smooth muscle against histamine induced contractile responses and that bronchial hyperreactivity to histamine is not associated with proinflammatory cytokine-induced enhancement in H(1)R signaling. PMID- 11282783 TI - Alveolar epithelial transport. Basic science to clinical medicine. PMID- 11282784 TI - Independent inheritance of serum immunoglobulin E concentrations and airway responsiveness. PMID- 11282785 TI - Serum vascular endothelial growth factor is elevated in cystic fibrosis and decreases with treatment of acute pulmonary exacerbation. PMID- 11282786 TI - A model of obstructive sleep apnea in normal humans. PMID- 11282787 TI - Are US populations appropriate for trials of human immunodeficiency virus vaccine? The HIVNET Vaccine Preparedness Study. AB - Questions exist about whether testing of preventive human immunodeficiency virus (HIV)-1 vaccines, which will require rapid recruitment and retention of cohorts with high HIV-1 seroincidence, is feasible in the United States. A prospective cohort study was conducted in 1995-1997 among 4,892 persons at high risk for HIV infection in nine US cities. At 18 months, with an 88% retention rate, 90 incident HIV-1 infections were observed (1.31/100 person-years (PY), 95% confidence interval (CI): 1.06, 1.61). HIV-1 seroincidence rates varied significantly by baseline eligibility criteria--1.55/100 PY among men who had sex with men, 0.38/100 PY among male intravenous drug users, 1.24/100 PY among female intravenous drug users, and 1.13/100 PY among women at heterosexual risk-and by enrollment site, from 0.48/100 PY to 2.18/100 PY. HIV-1 incidence was highest among those men who had sex with men who reported unprotected anal intercourse (2.01/100 PY, 95% CI: 1.54, 2.63), participants who were definitely willing to enroll in an HIV vaccine trial (1.96/100 PY, 95% CI: 1.41, 2.73), and women who used crack cocaine (1.62/100 PY, 95% CI: 0.92, 2.85). Therefore, cohorts with HIV 1 seroincidence rates appropriate for HIV-1 vaccine trials can be recruited, enrolled, and retained. PMID- 11282788 TI - Vaccine efficacy trials for human immunodeficiency virus/acquired immunodeficiency syndrome are feasible in the United States: a commentary on the HIVNET Vaccine Preparedness Study. PMID- 11282789 TI - Estimating the relative incidence of heroin use: application of a method for adjusting observed reports of first visits to specialized drug treatment agencies. AB - In this paper, the authors propose a method for estimating the incidence of heroin use by adjusting reported numbers of heroin users visiting drug treatment agencies for the time lag between onset of heroin use and first treatment request (lag distribution). The adjusted incidence is relative, since it represents the number of individuals beginning heroin use in each year whose cases will be reported within 8 years of starting use. Users with longer lag times or whose cases are never reported are excluded. Utilizing data from southeastern England (1991--1998), the authors analyzed the effects of covariates (sex, age group, ethnic group, route of consumption, and year of onset of drug use) on the lag distribution. Trends in the adjusted incidence of heroin use were very different for injectors and noninjectors: Incidence among injectors seemed to be stable, while in noninjectors it increased twofold between 1991 and 1996--1997. These results must be interpreted cautiously, especially in relation to the wider context of underlying trends in the population. Potential biases derive from underreporting and from changes in the proportion of heroin users in treatment. The lag correction method adds substantially to the value of routine treatment data, at least for heroin use, and is potentially the best method for obtaining estimates of incidence. PMID- 11282790 TI - Low blood pressure during pregnancy and poor perinatal outcomes: an obstetric paradox. AB - Low blood pressure during pregnancy has been associated with poor perinatal outcomes. However, whether this association is causal or is due to confounding has never been carefully assessed. The authors used data from the Collaborative Perinatal Project, a large prospective cohort study in 12 hospitals in the United States from 1959 to 1966. A total of 28,095 subjects were included. At first glance, it appeared that the lower the baseline blood pressure during pregnancy, the higher the incidence of very premature birth (<34 weeks) and severe small for gestational age (<5th percentile) in a consistent dose-response pattern. However, women with low blood pressure were generally younger, shorter, lighter, leaner, poorer, and more often a minority, and they gained less weight. After the authors controlled for these factors, low blood pressure was not associated with preterm birth (adjusted relative risks ranging from 0.86 to 0.93, p > 0.05) or small for gestational age (relative risks ranging from 0.45 to 2.0). Therefore, the association between low blood pressure during pregnancy and poor perinatal outcomes is largely due to confounding by other risk factors. Low blood pressure by itself does not increase risk of poor perinatal outcomes at a population level. However, this conclusion may not apply to individual patients who also have a compromised plasma volume expansion or pathologic homeostasis. PMID- 11282791 TI - Frequency of eating during pregnancy and its effect on preterm delivery. AB - Frequency of eating or meal patterns during pregnancy may be a component of maternal nutrition relevant to pregnancy outcome. To identify meal patterns of pregnant women and investigate the relation between these meal patterns and preterm delivery, the authors performed an analysis using data from the Pregnancy, Infection, and Nutrition Study (n = 2,065). Women recruited from August 1995 to December 1998 were categorized by meal patterns on the basis of their reported number of meals (breakfast, lunch, and dinner) and snacks consumed per day during the second trimester. An optimal pattern was defined according to the Institute of Medicine recommendation of three meals and two or more snacks per day. In this population, 72 percent of the women met this recommendation, and 235 delivered preterm. Women who consumed meals/snacks less frequently were slightly heavier prior to pregnancy, were older, and had a lower total energy intake. In addition, these women had a higher risk of delivering preterm (adjusted odds ratio = 1.30, 95 percent confidence interval: 0.96, 1.76). There was no meaningful difference in the risk by early versus late preterm delivery, but those who delivered after premature rupture of the membranes (adjusted odds ratio = 1.87, 95 percent confidence interval: 1.02, 3.43) had a higher risk than those who delivered after preterm labor (adjusted odds ratio = 1.11, 95 percent confidence interval: 0.64, 1.89). This study supports previous animal model work of an association between decreased frequency of eating and preterm delivery. PMID- 11282792 TI - Family size, day-care attendance, and breastfeeding in relation to the incidence of childhood asthma. AB - A hypothesis has been suggested stating that children exposed early to infections are less likely to develop atopy or asthma. The authors investigated the relation between risk of childhood asthma and number of siblings as well as day-care attendance, as factors possibly increasing the likelihood of early infections, and breastfeeding as a factor reducing them. A case-control study was carried out in Montreal, Canada, between 1988 and 1995 that included 457 children diagnosed with asthma at 3--4 years of age and 457 healthy controls. Cases followed for 6 years were later classified as persistent or transient by the symptoms and use of medication after diagnosis. Among cases diagnosed at 3--4 years of age, the adjusted odds ratio for asthma was 0.54 (95% confidence interval (CI): 0.36, 0.80) for one sibling and 0.49 (95% CI: 0.30, 0.81) for two or more. The adjusted odds ratio for day-care attendance before 1 year of age was 0.59 (95% CI: 0.40, 0.87). Results were similar with persistent cases. Among transient cases (who possibly had an infection with wheezing at 3--4 years of age), day-care attendance and a short duration of breastfeeding resulted in increased risk. The results support the hypothesis that opportunity for early infections reduces the risk of asthma. PMID- 11282793 TI - Human and pet-related risk factors for household evacuation failure during a natural disaster. AB - This study characterized risk factors for household evacuation failure. A random digit dial telephone survey was conducted of 397 households in Yuba County, California, in July 1997, 6 months after residents had been under evacuation notice due to flooding. Case households failed to evacuate, whereas control households evacuated. The cumulative incidence of household evacuation failure was 19.4%. Fewer households with children (25.8%) failed to evacuate than households without children (45.9%, p < 0.01). More households with pets (20.9%) than households without pets failed to evacuate (16.3%, p = 0.11). With multivariate logistic regression, the risk of household evacuation failure was lower in households with children (odds ratio = 0.4, 95% confidence interval: 0.2, 0.8) compared with households without children. The risk of household evacuation failure increased in pet-owning households without children (odds ratio = 1.3, 95% confidence interval: 1.1, 1.5) compared with pet-owning households with children; the more pets a household owned, the higher the risk of household evacuation failure was. Impediments to pet evacuation, including owning multiple pets, owning outdoor dogs, or not having a cat carrier, explained why many households that owned pets failed to evacuate. Predisaster planning should place a high priority on facilitating pet evacuation through predisaster education of pet owners and emergency management personnel. PMID- 11282794 TI - Incidence of and risk factors for asymptomatic peripheral arterial occlusive disease: a longitudinal study. AB - The current study describes the age- and sex-specific incidence rates and risk factors for asymptomatic and symptomatic peripheral arterial occlusive disease (PAOD) among 2,327 subjects and the incidence of intermittent claudication in asymptomatic PAOD subjects. The study population was selected from 18 general practice centers in the Netherlands. PAOD was assessed with the ankle-brachial blood pressure index, and intermittent claudication was assessed with a modified version of the Rose questionnaire. After 7.2 years, the overall incidence rate for asymptomatic PAOD, using the person-years method, was 9.9 (95% confidence interval (CI): 7.3, 18.8) per 1,000 person-years at risk. The rate was 7.8 (95% CI: 4.9, 20.3) for men and 12.4 (95% CI: 7.7, 24.8) for women. For symptomatic PAOD, the incidence rate was 1.0 (95% CI: 0.7, 7.5) overall, 0.4 (95% CI: 0.3, 10.0) for men, and 1.8 (95% CI: 1.0, 10.3) for women. Multivariate analyses showed that increasing age, smoking, hypertension, and diabetes mellitus were the most important risk factors. The overall incidence rate for intermittent claudication among PAOD subjects who were asymptomatic at baseline was 90.5 per 1,000 person-years at risk (95% CI: 36.4, 378.3). The incidence of asymptomatic PAOD was higher than the incidence of symptomatic PAOD, with women developing PAOD more often than men. In the development of preventive strategies, modification of atherosclerotic risk factors, such as smoking, hypertension, and diabetes, should be the main goals. PMID- 11282795 TI - Methylenetetrahydrofolate reductase 677 C/T genotype and cardiovascular disease mortality in postmenopausal women. AB - Methylenetetrahydrofolate reductase (MTHFR) is involved in the reduction of 5,10 methylenetetrahydrofolate to 5-methyltetrahydrofolate. A 677 C/T single nucleotide polymorphism localized in the MTHFR gene is associated with both thermolability and reduced activity of the enzyme and is associated with increased homocysteine levels. The authors investigated the relation between the MTHFR 677 C/T polymorphism and risk of cardiovascular disease mortality in a cohort study of 12,239 women initially aged 52--67 years with a maximum follow-up time of 18 years (1976--1995; 153,732 woman-years of follow-up). The cardiovascular disease mortality rate was highest among women with the MTHFR 677 CC wild-type genotype and lowest among MTHFR 677 TT homozygotes. In comparison with women with the 677 CC wild-type genotype, age-adjusted rate ratios were 0.7 (95% confidence interval: 0.5, 0.9) for 677 CT heterozygotes and 0.6 (95% confidence interval: 0.4, 1.0) for 677 TT homozygotes. The possibility that this relation is a chance finding must be considered, because the relation is weak and of borderline significance. However, it provides an important argument against the view that increased levels of homocysteine directly raise cardiovascular disease risk. PMID- 11282796 TI - Dietary and other methyl-group availability factors and pancreatic cancer risk in a cohort of male smokers. AB - The authors examined prospectively whether dietary folate and other factors known to influence methyl-group availability were associated with the development of exocrine pancreatic cancer within the Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study cohort. Of the 27,101 healthy male smokers aged 50--69 years who completed a self-administered dietary questionnaire at baseline, 157 developed pancreatic cancer during up to 13 years of follow-up from 1985 to 1997. Cox proportional hazards models were used to estimate the hazards ratios and 95% confidence intervals. The adjusted hazards ratio comparing the highest with the lowest quintile of dietary folate intake was 0.52 (95% confidence interval: 0.31, 0.87; p-trend = 0.05). Dietary methionine, alcohol intake, and smoking history did not modify this relation. No significant associations were observed between dietary methionine, vitamins B(6) and B(12), or alcohol intake and pancreatic cancer risk. Consistent with prior studies, this study shows that cigarette smoking was associated with an increased risk (highest compared with lowest quintile, cigarettes per day: hazards ratio = 1.82; 95% confidence interval: 1.10, 3.03; p-trend = 0.05). These results support the hypothesis that dietary folate intake is inversely associated with the risk of pancreatic cancer and confirm the risk associated with greater cigarette smoking. PMID- 11282797 TI - Association of the B-vitamins pyridoxal 5'-phosphate (B(6)), B(12), and folate with lung cancer risk in older men. AB - A nested case-control study was conducted within the Alpha-Tocopherol, Beta Carotene Cancer Prevention Study cohort to test for associations between selected B-vitamins (folate, vitamin B(6), vitamin B(12)) and incident lung cancer. This trial was conducted in Finland between 1985 and 1993. Serum was analyzed for these nutrients and homocysteine among 300 lung cancer cases and matched controls (1:1). Odds ratios and 95% confidence intervals were determined in conditional and unconditional (controlling for the matching factors) logistic regression models, after adjusting for body mass index, years of smoking, and number of cigarettes smoked per day. No significant associations were seen between serum folate, vitamin B(12), or homocysteine and lung cancer risk. The authors found significantly lower risk of lung cancer among men who had higher serum vitamin B(6) levels. Compared with men with the lowest vitamin B(6) concentration, men in the fifth quintile had about one half of the risk of lung cancer (odds ratio = 0.51; 95% confidence interval: 0.23, 0.93; p-trend = 0.02). Adjusting for any of the other serum factors (folate, B(12), and homocysteine) either alone or jointly did not significantly alter these estimates. This is the first report from a prospectively conducted study to suggest a role for vitamin B(6) in lung cancer. PMID- 11282798 TI - Lung cancer among industrial sand workers exposed to crystalline silica. AB - In 1997, the International Agency for Research on Cancer determined that crystalline silica was a human carcinogen but noted inconsistencies in the epidemiology. There are few exposure-response analyses. The authors examined lung cancer mortality among 4,626 industrial sand workers, estimating exposure via a job-exposure matrix based on 4,269 industrial hygiene samples collected in 1974- 1995. The average length of employment was 9 years, and estimated average exposure was 0.05 mg/m(3) (the National Institute of Occupational Safety and Health Recommended Exposure Limit). Results confirmed excess mortality from silicosis/pneumoconioses (standardized mortality ratio = 18.2, 95% confidence interval: 10.6, 29.1; 17 deaths). The lung cancer standardized mortality ratio was 1.60 (95% confidence interval: 1.31, 1.93; 109 deaths). Limited data suggested that smoking might account for 10--20% of the lung cancer excess. Exposure-response analyses by quartile of cumulative exposure (15-year lag) yielded standardized rate ratios of 1.00, 0.78, 1.51, and 1.57 (p for trend = 0.07). Nested case-control analyses after exclusion of short-term workers, who had high overall morality, yielded odds ratios by quartile of cumulative exposure (15-year lag) of 1.00, 1.35, 1.63, and 2.00 (p for trend = 0.08) and odds ratios by quartile of average exposure of 1.00, 0.92, 1.44, and 2.26 (p = 0.005). These data lend support to the labeling by the International Agency for Research on Cancer of silica as a human carcinogen. There are approximately 2 million US workers exposed to silica; 100,000 are exposed to more than 0.1 mg/m(3). PMID- 11282800 TI - Re: "Clustering of procoagulation, inflammation, and fibrinolysis variables with metabolic factors in insulin resistance syndrome". PMID- 11282799 TI - Association between air pollution and daily consultations with general practitioners for allergic rhinitis in London, United Kingdom. AB - Few published studies have looked at the health effects of air pollution in the primary care setting, and most have concentrated on lower rather than upper respiratory diseases. The authors investigated the association of daily consultations with general practitioners for allergic rhinitis with air pollution in London, United Kingdom. Generalized additive models were used to regress time series of daily numbers of patients consulting for allergic rhinitis against 1992 -1994 measures of air pollution, after control for possible confounders and adjustment for overdispersion and serial correlation. In children, a 10th--90th percentile increase in sulfur dioxide (SO(2)) levels 4 days prior to consultation (13-31 microg/m(3)) was associated with a 24.5% increase in consultations (95% confidence interval: 14.6, 35.2; p < 0.00001); a 10th--90th percentile increase in averaged ozone (O(3)) concentrations on the day of consultation and the preceding 3 days (6--29 parts per billion) was associated with a 37.6% rise (95% confidence interval: 23.3, 53.5; p < 0.00001). For adults, smaller effect sizes were observed for SO(2) and O(3). The association with SO(2) remained highly significant in the presence of other pollutants. This study suggests that air pollution worsens allergic rhinitis symptoms, leading to substantial increases in consultations. SO(2) and O(3) seem particularly responsible, and both seem to contribute independently. PMID- 11282801 TI - Re: "Racial differences in reported Lyme disease incidence". PMID- 11282802 TI - Re: "Comparison of National Death Index and world wide web death searches". PMID- 11282804 TI - Appointed czars, elected presidents and windows of opportunity. PMID- 11282805 TI - Recurrent unipolar depression requires prolonged treatment. PMID- 11282806 TI - Community care for people with mental disorders in developing countries: problems and possible solutions. PMID- 11282807 TI - How antidepressants work: new perspectives on the pathophysiology of depressive disorder. AB - BACKGROUND: New research in animals is beginning to change radically our understanding of the biology of stress and the effects of antidepressant agents. AIMS: To relate recent findings from the basic neurosciences to the pathophysiology of depressive disorder. METHOD: Drawing together findings from molecular and physiological studies in rats, social studies in primates and neuropsychological studies in humans, we review the neurotrophic and neuroplastic effects of antidepressants and stress. RESULTS: Stress and antidepressants have reciprocal actions on neuronal growth and vulnerability (mediated by the expression of neurotrophins) and synaptic plasticity (mediated by excitatory amino acid neurotransmission) in the hippocampus and other brain structures. Stressors have the capacity to progressively disrupt both the activities of individual cells and the operating characteristics of networks of neurons throughout the life cycle, while antidepressant treatments act to reverse such injurious effects. CONCLUSIONS: We propose a central role for the regulation of synaptic connectivity in the pathophysiology of depressive disorder. PMID- 11282808 TI - Prophylactic effect of citalopram in unipolar, recurrent depression: placebo controlled study of maintenance therapy. AB - BACKGROUND: Major depression is highly recurrent. Antidepressant maintenance treatment has proven efficacy against recurrent depression. AIMS: Comparison of prophylactic efficacy of citalopram versus placebo in unipolar, recurrent depression. METHODS: Patients 18-65 years of age with recurrent unipolar major depression (DSM-IV), a Montgomery-Asberg Depression Rating Scale score of > or =22 and two or more previous depressive episodes, one within the past 5 years, were treated openly with citalopram (20-60 mg) for 6-9 weeks and, if responding, continued for 16 weeks before being randomised to double-blind maintenance treatment with citalopram or placebo for 48-77 weeks. RESULTS: A total of 427 patients entered acute treatment and 269 were randomised to double-blind treatment. Time to recurrence was longer in patients taking citalopram than in patients taking placebo (P:<0.001). Prophylactic treatment was well tolerated. CONCLUSIONS: Citalopram (20, 40 and 60 mg) is effective in the prevention of depressive recurrences. Patients at risk should continue maintenance treatment at the dose necessary to resolve symptoms in the acute treatment phase. PMID- 11282809 TI - Light therapy for seasonal affective disorder in primary care: randomised controlled trial. AB - BACKGROUND: Studies of light therapy have not been conducted previously in primary care. AIMS: To evaluate light therapy in primary care. METHOD: Fifty seven participants with seasonal affective disorder were randomly allocated to 4 weeks of bright white or dim red light. Baseline expectations for treatment were assessed. Outcome was assessed with the Structured Interview Guide for the Hamilton Depression Scale, Seasonal Affective Disorder Version. RESULTS: Both groups showed decreases in symptom scores of more than 40%. There were no differences in proportions of responders in either group, regardless of the remission criteria applied, with around 60% (74% white light, 57% red light) meeting broad criteria for response and 31% (30% white light, 33% red light) meeting strict criteria. There were no differences in treatment expectations. CONCLUSIONS: Primary care patients with seasonal affective disorder improve after light therapy, but bright white light is not associated with greater improvements. PMID- 11282810 TI - Chronic benzodiazepine use in general practice patients with depression: an evaluation of controlled treatment and taper-off: report on behalf of the Dutch Chronic Benzodiazepine Working Group. AB - BACKGROUND: Many patients with depression take benzodiazepine drugs long term despite the absence of continuing therapeutic value. AIMS: To evaluate a treatment programme involving gradual discontinuation with or without simultaneous selective serotonin reuptake inhibitor (SSRI) prescribing and to determine the long-term outcome after benzodiazepine withdrawal. METHOD: Patients went through three phases - change to an equivalent dose of diazepam; subsequent randomisation to either 20 mg of paroxetine or placebo; and gradual reduction of diazepam in depression-free patients - with a follow-up after 2 or 3 years. RESULTS: A total of 230 patients were recruited and 75% in the paroxetine group and 61% in the placebo group were successfully treated after 6 weeks (P:=0.067). After 2 or 3 years 13% of patients were still benzodiazepine free: 26% of those who had successfully tapered off benzodiazepine and 6% of the total group. CONCLUSIONS: Transfer to diazepam followed by gradual withdrawal is an effective way of discontinuing chronic benzodiazepine use. The addition of SSRI treatment is of limited value. PMID- 11282811 TI - Efficacy and safety of sildenafil citrate in the treatment of men with mild to moderate erectile dysfunction. AB - BACKGROUND: Erectile dysfunction is a common, multi-factorial disorder. AIMS: To evaluate the efficacy, tolerability and frequency of use of sildenafil citrate in men with mild to moderate erectile dysfunction of no established organic cause. METHOD: This double-blind, randomised, placebo-controlled, flexible-dose, two-way crossover study was conducted at four centres in the UK in 44 men with mild to moderate erectile dysfunction of no clinically obvious organic cause. The study included two 28-day treatment periods, during which time sildenafil or placebo (25-75 mg, based on efficacy) was taken as required. RESULTS: Compared with placebo, sildenafil was associated with increases in frequency of use, erections adequate for sexual intercourse and level of sexual satisfaction (P:<0.0001). More patients receiving sildenafil stated they would use the treatment again compared with those receiving placebo (P:<0.0001). There were no discontinuations due to sildenafil treatment. CONCLUSIONS: Sildenafil is effective and well tolerated in men with mild to moderate erectile dysfunction of no clinically identifiable organic cause. PMID- 11282812 TI - Self-exposure therapy for panic disorder with agoraphobia: randomised controlled study of external v. interoceptive self-exposure. AB - BACKGROUND: Exposure to external phobic cues is an effective therapy for panic/agoraphobia but the value of exposure to interoceptive cues is unclear. AIMS: Randomised controlled comparison in panic/agoraphobia of the effects of (a) external, (b) interceptive or (c) combined external and interoceptive self exposure to (d) control subjects. METHOD: Eighty out-patients were randomised to a control group or to one of three forms of self-exposure treatment (external, interoceptive, or combined). Each treatment included seven sessions over 10 weeks and daily self-exposure homework. Assessments were at pre- and post-treatment and up to 1 year post-entry. Assessors remained blind during treatment. RESULTS: The three self-exposure groups improved significantly and similarly at post-treatment and up to 1-year followup, and significantly more than did the control subjects. Rates of improvement on main outcome measures averaged 60% at post-treatment and 77% at follow-up. CONCLUSIONS: The three methods of self-exposure were equally effective in reducing panic and agoraphobic symptoms in the short- and long-term. PMID- 11282813 TI - Schizophrenia and the frontal lobes: post-mortem stereological study of tissue volume. AB - BACKGROUND: It has been suggested that there is frontal lobe involvement in schizophrenia, and that it may be lateralised and gender-specific. AIMS: To clarify the structure of the frontal lobes in schizophrenia in a post-mortem series. METHOD: The volume of white matter and cortical components of the frontal lobes was measured in brains of controls and patients with schizophrenia using planimetry and the Cavalieri principle. The components measured were: superior frontal gyrus, middle frontal gyrus, a composite of inferior frontal gyrus and orbito-frontal cortex, as well as total frontal lobe cortex and white matter. In addition, the anterior cingulate gyrus was measured. RESULTS: No diagnosis, gender, diagnosis x side, diagnosis x gender or diagnosis x gender x side interactions were observed in the volume of any of the components, the grey matter as a whole or the white matter. No evidence for volumetric inter-group differences was found for the anterior cingulate gyrus. CONCLUSIONS: Such structural abnormalities as are present in the frontal lobes are more subtle than straightforward alterations in tissue volume; they may include changes in shape and the pattern of gyral folding. PMID- 11282814 TI - Handedness, language lateralisation and anatomical asymmetry in schizophrenia: meta-analysis. AB - BACKGROUND: Cerebral lateralisation appears to be decreased in schizophrenia. Results of studies investigating this, however, are equivocal. AIMS: To review quantitatively the literature on decreased lateralisation in schizophrenia. METHOD: Meta-analyses were conducted on 19 studies on handedness, 10 dichotic listening studies and 39 studies investigating anatomical asymmetry in schizophrenia. RESULTS: The prevalence of mixed- and left-handedness ('non-right handedness') was significantly higher in patients with schizophrenia as compared to healthy controls, and also as compared to psychiatric controls. The analysis of dichotic listening studies revealed no significant difference in lateralisation in schizophrenia. However, when analysis was restricted to studies using consonant-vowel or fused word tasks, significantly decreased lateralisation in schizophrenia emerged. Asymmetry of the planum temporale and the Sylvian fissure was significantly decreased in schizophrenia, while asymmetry of the temporal horn of the lateral ventricle was not. CONCLUSION: Strong evidence is provided for decreased cerebral lateralisation in schizophrenia. PMID- 11282815 TI - Relationship between 5-HT function and impulsivity and aggression in male offenders with personality disorders. AB - BACKGROUND: Reduced serotonergic (5-HT) function and elevated testosterone have been reported in aggressive populations. AIMS: To investigate relationships between impulsivity, aggression, 5-HT function and testosterone in male offenders with personality disorders. METHOD: Sixty male offenders with DSM-III-R personality disorders and 27 healthy staff controls were assessed using the Special Hospital Assessment of Personality and Socialisation (SHAPS), impulsivity and aggression ratings, d-fenfluramine challenge and plasma hormone concentrations. RESULTS: The SHAPS non-psychopaths and those with schizoid personality disorders had enhanced 5-HT function (prolactin response to d fenfluramine). Reduced 5-HT function was found in offenders with DSM-III-R borderline personality disorders and those with a history of repeated self-harm or alcohol misuse. The 5-HT function was inversely correlated more strongly with impulsivity than with aggression. Plasma testosterone correlated positively with aggressive acts. The SHAPS primary psychopaths had lower initial cortisol and higher testosterone concentrations than controls. CONCLUSIONS: Future studies are needed to investigate regional brain 5-HT function. PMID- 11282816 TI - Exposure to anaesthetic agents, cognitive functioning and depressive symptomatology in the elderly. AB - BACKGROUND: Anaesthesia could provoke persistent alterations in specific cognitive domains in the elderly where ageing-related neuronal changes may exacerbate pharmacotoxic effects. AIMS: To evaluate anaesthesia effects on the incidence of cognitive dysfunction after orthopaedic surgery in elderly patients. METHOD: A total of 140 patients over the age of 64 years completed a full range of computerised cognitive tests. The study takes into account effects of pre operative cognitive dysfunction, depressive symptomatology and ability to perform activities of daily living. RESULTS: Postoperative cognitive decline persisted for up to 3 months in 56% of subjects. Dysfunction was limited to verbal, visuo spatial and semantic abilities and secondary and implicit memory. Age, low educational level, pre-operative cognitive impairment or depression are risk factors. CONCLUSIONS: Cognitive functions are not equally affected, type of impairment being determined by the risk factors described above and anaesthesia type. PMID- 11282817 TI - Incidence of psychotic disorders in immigrant groups to The Netherlands. AB - BACKGROUND: Previous reports on the incidence of schizophrenia in immigrant groups to The Netherlands were based on hospital data. AIMS: To compare the incidence of psychotic disorders in the immigrant groups to that in natives. METHOD: Two-year first-contact incidence study in The Hague. RESULTS: The risks of schizophrenia, schizophreniform or schizoaffective disorder (DSM-IV criteria) were increased for subjects born in Morocco (gender and age-adjusted relative risk=4.5; 95% Cl 1.4-8.5), Surinam (relative risk=3.2; 1.8-5.7), The Netherlands Antilles (relative risk=2.9; 0.9-9.5) and other non-Western countries (relative risk=2.4; 1.3-4.7). This risk was also increased for Moroccans (relative risk=8.0; 2.6-24.5) and Surinamese (relative risk=5.5; 2.5-11.9) of the second generation. The risks for Turkish immigrants, first or second generation, and for immigrants from Western countries were not significantly increased. CONCLUSIONS: This study indicates that the incidence of schizophrenia is increased in several, but not all, immigrant groups to The Netherlands. It is possible that factors associated with a process of rapid westernisation precipitate schizophrenia in people who are genetically at risk. PMID- 11282818 TI - The Truth and Reconciliation Commission in South Africa: relation to psychiatric status and forgiveness among survivors of human rights abuses. AB - BACKGROUND: The impact on individual survivors of human rights abuses of testifying before South Africa's Truth and Reconciliation Commission (TRC) has not been established. AIMS: To examine the degree to which participation in the TRC is related to current psychiatric status and forgiveness among survivors. METHOD: Survivors (n=134) who gave public, closed or no testimony to the TRC completed instruments measuring exposure to human rights abuses, exposure to other traumatic events, current psychiatric status and forgiveness attitudes towards the perpetrator(s). RESULTS: There was no significant association between TRC participation and current psychiatric status or current forgiveness attitudes, and low forgiveness was associated with poorer psychiatric health. CONCLUSIONS: Truth commissions should form part of, rather than be a substitute for, comprehensive therapeutic interventions for survivors of human rights abuses. Lack of forgiveness may be an important predictor of psychiatric risk in this population. PMID- 11282819 TI - Towards a unitary theory of stigmatisation. PMID- 11282820 TI - Cognitive therapy in schizophrenia. PMID- 11282821 TI - Stigma caused by psychiatrists. PMID- 11282822 TI - Seasonal variation in suicides: hidden not vanished. PMID- 11282823 TI - Soviet-style psychiatry is alive and well in the People's Republic. PMID- 11282824 TI - No long-term benefit for cognitive therapy in acute psychosis: a type II error. PMID- 11282826 TI - Focus on psychiatry in South Africa. PMID- 11282827 TI - Introduction. PMID- 11282844 TI - Medically unexplained symptoms in secondary care. PMID- 11282845 TI - Managing depression in primary care. PMID- 11282846 TI - Beyond Helsinki: a vision for global health ethics. PMID- 11282847 TI - Prescribing warmer, healthier homes. PMID- 11282848 TI - Reducing deaths among drug misusers. PMID- 11282849 TI - Plan to end age discrimination in NHS is launched. PMID- 11282850 TI - NHS told to pay 10m (pounds sterling) to patients infected with hepatitis C. PMID- 11282852 TI - President Bush outlines patients' bill of rights for US. PMID- 11282851 TI - A healthy old age: realistic or futile goal? Older people need to be encourage to exercise. PMID- 11282853 TI - Lung cancer death rates rise 600% in US women. PMID- 11282854 TI - Traditional butchery methods linked to vCJD cluster. PMID- 11282855 TI - Pregnant women cannot be tested for drugs without consent. PMID- 11282856 TI - House of Lords supports first UK genetic database. PMID- 11282857 TI - UK strategy to help increase clean water supplies worldwide. PMID- 11282859 TI - Dietary fat intake and prevention of cardiovascular disease: systematic review. AB - OBJECTIVE: To assess the effect of reduction or modification of dietary fat intake on total and cardiovascular mortality and cardiovascular morbidity. DESIGN: Systematic review. DATA SOURCES: Cochrane Library, Medline, Embase, CAB abstracts, SIGLE, CVRCT registry, and biographies were searched; trials known to experts were included. INCLUDED STUDIES: Randomised controlled trials stating intention to reduce or modify fat or cholesterol intake in healthy adult participants over at least six months. Inclusion decisions, validity, and data extraction were duplicated. Meta-analysis (random effects methodology), meta regression, and funnel plots were performed. RESULTS: 27 studies (30 902 person years of observation) were included. Alteration of dietary fat intake had small effects on total mortality (rate ratio 0.98; 95% confidence interval 0.86 to 1.12). Cardiovascular mortality was reduced by 9% (0.91; 0.77 to 1.07) and cardiovascular events by 16% (0.84; 0.72 to 0.99), which was attenuated (0.86; 0.72 to 1.03) in a sensitivity analysis that excluded a trial using oily fish. Trials with at least two years' follow up provided stronger evidence of protection from cardiovascular events (0.76; 0.65 to 0.90). CONCLUSIONS: There is a small but potentially important reduction in cardiovascular risk with reduction or modification of dietary fat intake, seen particularly in trials of longer duration. PMID- 11282860 TI - The effectiveness of exercise as an intervention in the management of depression: systematic review and meta-regression analysis of randomised controlled trials. AB - OBJECTIVE: To determine the effectiveness of exercise as an intervention in the management of depression. DESIGN: Systematic review and meta-regression analysis of randomised controlled trials obtained from five electronic databases (Medline, Embase, Sports Discus, PsycLIT, Cochrane Library) and through contact with experts in the field, bibliographic searches, and hand searches of recent copies of relevant journals. MAIN OUTCOME MEASURES: Standardised mean difference in effect size and weighted mean difference in Beck depression inventory score between exercise and no treatment and between exercise and cognitive therapy. RESULTS: All of the 14 studies analysed had important methodological weaknesses; randomisation was adequately concealed in only three studies, intention to treat analysis was undertaken in only two, and assessment of outcome was blinded in only one. The participants in most studies were community volunteers, and diagnosis was determined by their score on the Beck depression inventory. When compared with no treatment, exercise reduced symptoms of depression (standardised mean difference in effect size -1.1 (95% confidence interval -1.5 to -0.6); weighted mean difference in Beck depression inventory -7.3 (-10.0 to -4.6)). The effect size was significantly greater in those trials with shorter follow up and in two trials reported only as conference abstracts. The effect of exercise was similar to that of cognitive therapy (standardised mean difference -0.3 (95% confidence interval -0.7 to 0.1)). CONCLUSIONS: The effectiveness of exercise in reducing symptoms of depression cannot be determined because of a lack of good quality research on clinical populations with adequate follow up. PMID- 11282861 TI - Medically unexplained symptoms in frequent attenders of secondary health care: retrospective cohort study. AB - OBJECTIVE: To estimate the prevalence of medically unexplained symptoms in patients who most frequently attend outpatient services. DESIGN: Retrospective cohort study over three years with review of case notes. SETTING: Secondary care services in the South Thames (West) NHS region. PARTICIPANTS: Outpatient attenders with new appointments in 1993. MAIN OUTCOME MEASURES: Number of outpatient appointments, and number of consultation episodes for medically unexplained conditions. RESULTS: Medical records of 361 of 400 sampled frequent attenders were examined, and 971 consultation episodes were recorded. Ninety seven (27%) had one or more consultation episodes in which the condition was medically unexplained; 208 (21%) of the 971 consultation episodes were medically unexplained. Abdominal pain, chest pain, headache, and back pain were commonly found to be medically unexplained. CONCLUSIONS: Medically unexplained symptoms present in most hospital specialties and account for a considerable proportion of consultations by frequent attenders in secondary care. PMID- 11282862 TI - Longitudinal comparison of depression, coping, and turnover among NHS and private sector staff caring for people with dementia. PMID- 11282863 TI - Prenatal growth and subsequent marital status: longitudinal study. PMID- 11282866 TI - 10-minute consultation: Dyspepsia. PMID- 11282864 TI - Antidepressant drugs and generic counselling for treatment of major depression in primary care: randomised trial with patient preference arms. AB - OBJECTIVES: To compare the efficacy of antidepressant drugs and generic counselling for treating mild to moderate depression in general practice. To determine whether the outcomes were similar for patients with randomly allocated treatment and those expressing a treatment preference. DESIGN: Randomised controlled trial, with patient preference arms. Follow up at 8 weeks and 12 months and abstraction of GP case notes. SETTING: 31 general practices in Trent region. PARTICIPANTS: Patients aged 18-70 who met research diagnostic criteria for major depression; 103 patients were randomised and 220 patients were recruited to the preference arms. MAIN OUTCOME MEASURES: Difference in mean Beck depression inventory score; time to remission; global outcome assessed by a psychiatrist using all data sources; and research diagnostic criteria. RESULTS: At 12 months there was no difference between the mean Beck scores in the randomised arms. Combining the randomised and patient preference groups, the difference in Beck scores was 0.4 (95% confidence interval -2.7 to 3.5). Patients choosing counselling did better than those randomised to it (mean difference in Beck score 4.6, 0.0 to 9.2). There was no difference in the psychiatrist's overall assessment of outcome between any of the groups. 221/265 (83%) of participants with a known outcome had a remission. Median time to remission was shorter in the group randomised to antidepressants than the other three groups (2 months v 3 months). 33/221 (15%) patients had a relapse. CONCLUSIONS: Generic counselling seems to be as effective as antidepressant treatment for mild to moderate depressive illness, although patients receiving antidepressants may recover more quickly. General practitioners should allow patients to have their preferred treatment. PMID- 11282867 TI - Science, medicine, and the future: Radiofrequency ablation for atrial fibrillation. PMID- 11282868 TI - Lesson of the week: Acute hyponatraemia in children admitted to hospital: retrospective analysis of factors contributing to its development and resolution. PMID- 11282870 TI - ABC of diseases of liver, pancreas, and biliary system: Liver and pancreatic trauma. PMID- 11282871 TI - Stakeholder health insurance: empowering the poorest patients. PMID- 11282872 TI - Care of older people: Mental health problems. PMID- 11282873 TI - Detection of breast cancer. Mammography should be available. PMID- 11282874 TI - Detection of breast cancer. Self examination contributes to reduction in mortality. PMID- 11282875 TI - NHS Direct Online has important role. PMID- 11282876 TI - Diagnosing suspected ectopic pregnancy. Can we offer completely non-surgical management for ectopic pregnancy? PMID- 11282877 TI - Uncertainty about clinical equipoise. Clinical equipoise and the uncertainty principles both require further scrutiny. PMID- 11282878 TI - Diagnosing suspected ectopic pregnancy. Patients with falling concentrations of beta human chorionic gonadotrophin should be seen regularly. PMID- 11282879 TI - Uncertainty about clinical equipoise. There is another exchange on equipoise and uncertainty. PMID- 11282880 TI - Kitemarking the west wind. Website labels are analogous to food labels. PMID- 11282881 TI - Sexually transmitted infections in people with HIV infection. PMID- 11282882 TI - GMC no longer favours folder of evidence for revalidation. PMID- 11282883 TI - Detection of breast cancer. Clinical breast examination is not an acceptable alternative to mammography. PMID- 11282885 TI - To Cre or not to Cre: the next generation of mouse models of human cardiac diseases. PMID- 11282886 TI - Imaging the murine cardiovascular system with magnetic resonance. PMID- 11282887 TI - Roads to survival: insulin-like growth factor-1 signaling pathways in cardiac muscle. PMID- 11282888 TI - Gene expression patterns in the lungs of patients with primary pulmonary hypertension: a gene microarray analysis. AB - Primary pulmonary hypertension (PPH) is a disease of unknown etiology characterized by lumen-obliterating endothelial cell proliferation and vascular smooth muscle hypertrophy of the small precapillary pulmonary arteries. Because the vascular lesions are homogeneously distributed throughout the entire lung, we propose that a tissue fragment of the lung is representative of the whole lung. RNA extracted from the fragments is likely to provide meaningful information regarding the changes in gene expression pattern in PPH when compared with structurally normal lung tissue. We hypothesize that the lung tissue gene expression pattern of patients with PPH has a characteristic profile when compared with the gene expression pattern of structurally normal lungs and that this characteristic gene expression profile provides new insights into the pathobiology of PPH. Using oligonucleotide microarray technology, we characterized the expression pattern in the lung tissue obtained from 6 patients with primary pulmonary hypertension (PPH)-including 2 patients with the familial form of PPH (FPPH)-and from 6 patients with histologically normal lungs. For the data analysis, gene clusters were generated and the gene expression pattern differences between PPH and normal lung tissue and between PPH and FPPH lung tissue were compared. All PPH lung tissue samples showed a decreased expression of genes encoding several kinases and phosphatases, whereas several oncogenes and genes coding for ion channel proteins were upregulated in their expression. Importantly, we could distinguish by pattern comparison between sporadic PPH and FPPH, because alterations in the expression of transforming growth factor-beta receptor III, bone morphogenic protein 2, mitogen-activated protein kinase kinase 5, RACK 1, apolipoprotein C-III, and the gene encoding the laminin receptor 1 were only found in the samples from patients with sporadic PPH, but not in FPPH samples. We conclude that the microarray gene expression technique is a new and useful molecular tool that provides novel information pertinent to a better characterization and understanding of the pathobiology of the distinct clinical phenotypes of pulmonary hypertension. PMID- 11282889 TI - Dobutamine-stress magnetic resonance microimaging in mice : acute changes of cardiac geometry and function in normal and failing murine hearts. AB - The aim of this study was to assess the capability of MRI to characterize systolic and diastolic function in normal and chronically failing mouse hearts in vivo at rest and during inotropic stimulation. Applying an ECG-gated FLASH-cine sequence, MRI at 7 T was performed at rest and after administration of 1.5 microgram/g IP dobutamine. There was a significant increase of heart rate, cardiac output, and ejection fraction and significant decrease of end-diastolic and end-systolic left ventricular (LV) volumes (P<0.01 each) in normal mice during inotropic stimulation. In mice with heart failure due to chronic myocardial infarction (MI), MRI at rest revealed gross LV dilatation. There was a significant decrease of LV ejection fraction in infarcted mice (29%) versus sham mice (58%). Mice with MI showed a significantly reduced maximum LV ejection rate (P<0.001) and LV filling rate (P<0.01) and no increase of LV dynamics during dobutamine action, indicating loss of contractile and relaxation reserve. In 4 month-old transgenic mice with cardiospecific overexpression of the beta(1) adrenergic receptor, which at this early stage do not show abnormalities of resting cardiac function, LV filling rate failed to increase after dobutamine stress (transgenic, 0.19+/-0.03 microL/ms; wild type, 0.36+/-0.01 microL/ms; P<0.01). Thus, MRI unmasked diastolic dysfunction during dobutamine stress. Dobutamine-stress MRI allows noninvasive assessment of systolic and diastolic components of heart failure. This study shows that MRI can demonstrate loss of inotropic and lusitropic response in mice with MI and can unmask diastolic dysfunction as an early sign of cardiac dysfunction in a transgenic mouse model of heart failure. PMID- 11282890 TI - Functional roles of cardiac and vascular ATP-sensitive potassium channels clarified by Kir6.2-knockout mice. AB - -ATP-sensitive potassium (K(ATP)) channels were discovered in ventricular cells, but their roles in the heart remain mysterious. K(ATP) channels have also been found in numerous other tissues, including vascular smooth muscle. Two pore forming subunits, Kir6.1 and Kir6.2, contribute to the diversity of K(ATP) channels. To determine which subunits are operative in the cardiovascular system and their functional roles, we characterized the effects of pharmacological K(+) channel openers (KCOs, ie, pinacidil, P-1075, and diazoxide) in Kir6.2-deficient mice. Sarcolemmal K(ATP) channels could be recorded electrophysiologically in ventricular cells from Kir6.2(+/+) (wild-type [WT]) but not from Kir6.2(-/-) (knockout [KO]) mice. In WT ventricular cells, pinacidil induced an outward current and action potential shortening, effects that were blocked by glibenclamide, a K(ATP) channel blocker. KO ventricular cells exhibited no response to KCOs, but gene transfer of Kir6.2 into neonatal ventricular cells rescued the electrophysiological response to P-1075. In terms of contractile function, pinacidil decreased force generation in WT but not KO hearts. Pinacidil and diazoxide produced concentration-dependent relaxation in both WT and KO aortas precontracted with norepinephrine. In addition, pinacidil induced a glibenclamide-sensitive current of similar magnitude in WT and KO aortic smooth muscle cells and comparable levels of hypotension in anesthetized WT and KO mice. In both WT and KO aortas, only Kir6.1 mRNA was expressed. These findings indicate that the Kir6.2 subunit mediates the depression of cardiac excitability and contractility induced by KCOs; in contrast, Kir6.2 plays no discernible role in the arterial tree. PMID- 11282891 TI - Autoimmunity against the second extracellular loop of beta(1)-adrenergic receptors induces beta-adrenergic receptor desensitization and myocardial hypertrophy in vivo. AB - Although immunoapheresis removing autoantibodies against the second extracellular domain of beta(1)-adrenergic receptors (ARs) improves cardiac function in patients with cardiomyopathy, the underlying mechanisms have not been defined. We examined the role of autoimmunity against the domain in the development of cardiac dysfunction in vivo. Japanese white rabbits were immunized with a synthetic peptide corresponding to the second extracellular loop of beta(1)-AR once a month with (beta+biso rabbits, n=10) or without (beta rabbits, n=13) bisoprolol treatment (2 mg/kg per day). Control rabbits received vehicle without bisoprolol treatment (n=13). Autoantibodies of IgG isotype against the domain were persistently detected in beta and beta+biso rabbits. Purified IgG from sera of beta and beta+biso rabbits increased cAMP production in a rabbit cardiac membrane preparation, which was blocked by bisoprolol. At 3 months, beta-AR uncoupling with increased G protein-coupled receptor kinase 5 (GRK5) expression was found in beta rabbits. At 6 months, left ventricular hypertrophy was noted with hemodynamic derangements in beta rabbits. This was accompanied by decreased beta(1)-AR density and increased inhibitory G protein and GRK5 expression, which were related to marked decrease in membrane cAMP production. These changes in beta rabbits at 6 months were prevented in beta+biso rabbits. There was no difference in the plasma norepinephrine concentration in the 3 groups over the observation period. Thus, autoimmunity against the second extracellular loop of beta(1)-ARs induced profound beta-AR desensitization and myocardial hypertrophy in vivo, associated with cardiac dysfunction. Sustained sympathomimetic-like actions of autoantibodies against the domain may be partly responsible for these changes. PMID- 11282892 TI - Inducible gene targeting in postnatal myocardium by cardiac-specific expression of a hormone-activated Cre fusion protein. AB - Cardiac-restricted expression of Cre recombinase can provoke lineage-specific gene excision in the myocardium. However, confounding early lethality may still preclude using loss-of-function models to study the postnatal heart. Here, we have tested whether inducible, heart-specific recombination can be triggered after birth by transgenic expression of a Cre fusion protein that incorporates a mutated progesterone receptor ligand binding domain (PR1) that is activated by the synthetic antiprogestin, RU486, but not by endogenous steroid hormones. CrePR1 driven by the alpha-myosin heavy chain (alphaMHC) promoter was expressed specifically in heart. Translocation of CrePR1 from cytoplasm to nuclei in ventricular myocytes was induced by RU486. To establish whether this approach can mediate cardiac-specific, drug-dependent excision between loxP sites in vivo, we mated alphaMHC-CrePR1 mice with a ubiquitously expressed (ROSA26) Cre reporter line. Offspring harboring alphaMHC-CrePR1 and/or the floxed allele were injected with RU486 versus vehicle, and the prevalence of beta-galactosidase (beta-gal) positive cells was determined, indicative of Cre-mediated excision. Little or no baseline recombination was seen 1 week after birth. Cardiac-restricted, RU486 inducible recombination was demonstrated in bigenic mice at age 3 and 6 weeks, using each of 3 independent CrePR1 lines. Recombination in the absence of ligand paralleled the levels of CrePR1 protein expression and was more evident at 6 weeks. Thus, conditional, posttranslational activation of a Cre fusion protein can bypass potential embryonic and perinatal effects on the heart and permits inducible recombination in cardiac muscle. High levels of the chimeric Cre protein, in particular, were associated with progressive recombination in the absence of drug. PMID- 11282893 TI - Decreased flow-dependent dilation in carotid arteries of tissue kallikrein knockout mice. AB - - Flow-dependent dilation is a fundamental mechanism by which large arteries ensure appropriate blood supply to tissues. We investigated whether or not the vascular kallikrein-kinin system, especially tissue kallikrein (TK), contributes to flow-dependent dilation by comparing wild-type and TK-knockout mice in which the presence or absence of TK expression was verified. We examined in vitro changes in the outer diameter of perfused carotid arteries from TK(+/+) and TK(-/ ) mice. In both groups, exogenous bradykinin caused a similar dilation that was abolished by the B(2) receptor antagonist HOE-140, as well as by the NO synthase inhibitor N:(omega)-nitro-L-arginine methyl ester. However, purified kininogen dilated only TK(+/+) arteries, demonstrating the essential role of TK in the vascular formation of kinins. In TK(+/+) arteries, increasing intraluminal flow caused a larger endothelium-dependent dilation than that seen in TK(-/-). In both strains the flow response was mediated by NO and by endothelium-derived hyperpolarizing factor, whereas in TK(-/-) vasoconstrictor prostanoids participated as well. HOE-140 impaired flow-dependent dilation in TK(+/+) arteries while showing no effect in TK(-/-). This compound reduced the flow response in TK(+/+) arteries to a level similar to that in TK(-/-). After NO synthase inhibition, HOE-140 no longer affected the response of TK(+/+). Impaired flow-dependent dilation was also observed in arteries from knockout mice lacking bradykinin B(2) receptors as compared with wild-type animals. This study demonstrates the active contribution of the vascular kallikrein-kinin system to one-third of the flow-dependent dilation response via activation of B(2) receptors coupled to endothelial NO release. PMID- 11282894 TI - Cyclooxygenase-1 participates in selected vasodilator responses of the cerebral circulation. AB - Cyclooxygenase (COX) is a prostanoid-synthesizing enzyme present in 2 isoforms: COX-1 and COX-2. Although it has long been hypothesized that prostanoids participate in cerebrovascular regulation, the lack of adequate pharmacological tools has led to conflicting results and has not permitted investigators to define the relative contribution of COX-1 and COX-2. We used the COX-1 inhibitor SC-560 and COX-1-null (COX-1(-/-)) mice to investigate whether COX-1 plays a role in cerebrovascular regulation. Mice were anesthetized (urethane and chloralose) and equipped with a cranial window. Cerebral blood flow (CBF) was measured by laser Doppler flowmetry or by the (14)C-iodoantipyrine technique with quantitative autoradiography. In wild-type mice, SC-560 (25 micromol/L) reduced resting CBF by 21+/-4% and attenuated the CBF increase produced by topical application of bradykinin (-59%) or calcium ionophore A23187 (-49%) and by systemic hypercapnia (-58%) (P<0.05 to 0.01). However, SC-560 did not reduce responses to acetylcholine or the increase in somatosensory cortex blood flow produced by vibrissal stimulation. In COX-1(-/-) mice, resting CBF assessed by (14)C-iodoantipyrine was reduced (-13% to -20%) in cerebral cortex and other telencephalic regions (P<0.05). The CBF increase produced by bradykinin, A23187, and hypercapnia, but not acetylcholine or vibrissal stimulation, were attenuated (P<0.05 to 0.01). The free radical scavenger superoxide dismutase attenuated responses to bradykinin and A23187 in wild-type mice but not in COX-1(-/-) mice, suggesting that COX-1 is the source of the reactive oxygen species known to mediate these responses. The data provide evidence for a critical role of COX-1 in maintaining resting vascular tone and in selected vasodilator responses of the cerebral microcirculation. PMID- 11282895 TI - Reperfusion-activated Akt kinase prevents apoptosis in transgenic mouse hearts overexpressing insulin-like growth factor-1. AB - Abstract -Hearts of wild-type and insulin-like growth factor-1 overexpressing (Igf-1(+/-)) transgenic mice were subjected to Langendorff perfusions and progressive periods of ischemia followed by reperfusion. Apoptosis was measured by DNA nucleosomal cleavage and a hairpin probe labeling assay to detect single base overhang. Transgenic hearts subjected to 20 minutes of ischemia and 4 hours of reperfusion (I/R) sustained a rate of apoptosis of 1.8+/-0.3% compared with 4.6+/-1.1% for wild-type controls (n=4; P<0.03). Phosphorylation of the protein kinase Akt/protein kinase B was elevated 6.2-fold in transgenic hearts at baseline and increased another 4.4-fold within 10 minutes of reperfusion, remaining elevated for up to 2 hours. I/R activated Akt in wild-type hearts but to a lesser extent (1.6+/-0.3-fold). Pretreatment of transgenic hearts with wortmannin immediately before and during ischemia eliminated reperfusion-mediated activation of Akt and neutralized the resistance to apoptosis. The stress activated kinase p38 was also activated during ischemia and reperfusion in both wild-type and transgenic hearts. Perfusion with the p38 inhibitor SB203580 (10 micromol/L) blocked both p38 activation and phosphorylation of Akt and differentially modulated apoptosis in wild-type and transgenic hearts. Pretreatment with SB203580 reduced apoptosis in wild-type hearts but increased apoptosis in transgenic hearts. These results demonstrate that Akt phosphorylation during I/R is modulated by IGF-1 and prevents apoptosis in hearts that overexpress the IGF-1 transgene. PMID- 11282896 TI - Cellular pathology of atherosclerosis: smooth muscle cells prime cocultured endothelial cells for enhanced leukocyte adhesion. AB - During the development of an atherosclerotic plaque, mononuclear leukocytes infiltrate the artery wall through vascular endothelial cells (ECs). At the same time, arterial smooth muscle cells (SMCs) change from the physiological contractile phenotype to the secretory phenotype and migrate into the plaque. We investigated whether secretory SMCs released cytokines that stimulated ECs in a manner leading to increased leukocyte recruitment and thus might accelerate atheroma formation. SMCs and ECs were established in coculture on the opposite sides of a porous membrane, and the cocultured cells were incorporated into a flow-based assay for studying leukocyte adhesion. We found that coculture primed ECs so that their response to the inflammatory cytokine tumor necrosis factor alpha was amplified. ECs cocultured with SMCs supported greatly increased adhesion of flowing leukocytes and were sensitized to respond to tumor necrosis factor-alpha at concentrations 10 000 times lower than ECs cultured alone. In addition, coculture altered the endothelial selectin adhesion molecules used for leukocyte capture. EC priming was attributable to the cytokine transforming growth factor-beta(1), which was proteolytically activated to a biologically active form by the serine protease plasmin. These results suggest a new role for secretory SMCs in the development of atheromatous plaque. We propose that paracrine interaction between ECs and SMCs has the potential to amplify leukocyte recruitment to sites of atheroma and exacerbate the inflammatory processes believed to be at the heart of disease progression. PMID- 11282897 TI - Adenoviral expression of vascular endothelial growth factor-C induces lymphangiogenesis in the skin. AB - The growth of blood and lymphatic vasculature is mediated in part by secreted polypeptides of the vascular endothelial growth factor (VEGF) family. The prototype VEGF binds VEGF receptor (VEGFR)-1 and VEGFR-2 and is angiogenic, whereas VEGF-C, which binds to VEGFR-2 and VEGFR-3, is either angiogenic or lymphangiogenic in different assays. We used an adenoviral gene transfer approach to compare the effects of these growth factors in adult mice. Recombinant adenoviruses encoding human VEGF-C or VEGF were injected subcutaneously into C57Bl6 mice or into the ears of nude mice. Immunohistochemical analysis showed that VEGF-C upregulated VEGFR-2 and VEGFR-3 expression and VEGF upregulated VEGFR 2 expression at 4 days after injection. After 2 weeks, histochemical and immunohistochemical analysis, including staining for the lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), the vascular endothelial marker platelet-endothelial cell adhesion molecule-1 (PECAM-1), and the proliferating cell nuclear antigen (PCNA) revealed that VEGF-C induced mainly lymphangiogenesis in contrast to VEGF, which induced only angiogenesis. These results have significant implications in the planning of gene therapy using these growth factors. PMID- 11282898 TI - Weekly dosing with the platelet-derived growth factor receptor tyrosine kinase inhibitor SU9518 significantly inhibits arterial stenosis. AB - The platelet-derived growth factor (PDGF) ligands and their receptors have been implicated as critical regulators of the formation of arterial lesions after tissue injury. SU9518 (3[5-(5-bromo-2-oxo-1,2-dihydroindol-3-ylidenemethyl)-2,4 dimethyl-1H-pyrrol-3-yl]propionic acid) is a novel synthetic indolinone that potently and selectively inhibits the cellular PDGF receptor kinase and PDGF receptor-induced cell proliferation. Inhibition of PDGF receptor phosphorylation in cell-based assays occurs within 5 minutes after drug exposure and persists for >6 hours after drug removal. The pharmacokinetics indicate plasma levels that exceeded the effective concentration required to inhibit the PDGF receptor in cells for up to 8 hours or 7 days after a single oral or subcutaneous administration, respectively. In the rat balloon arterial injury-induced stenosis model, once-daily oral or once-weekly subcutaneous administration of SU9518 reduced intimal thickening of the carotid artery (ratio of neointimal to medial area, 1.94+/-0.38 versus 1.03+/-0.29 [P<0.01] 2.21+/-0.32 versus 1.34+/-0.45 [P<0.01], respectively). These studies provide the rationale to evaluate PDGF receptor tyrosine kinase inhibitors, including inhibitors related to the indolinone, SU9518, for the treatment of arterial restenosis. PMID- 11282899 TI - Retinoids inhibit the actions of angiotensin II on vascular smooth muscle cells. AB - Retinoids are derivatives of vitamin A and powerful inhibitors of cell proliferation and inflammation. Angiotensin II (Ang II) contributes to vascular lesions by promoting cell growth of vascular smooth muscle cells (VSMCs). Therefore, we examined whether retinoids interfere with the proproliferative actions of Ang II in VSMCs via AT(1) receptor-dependent or activator protein-1 (AP-1)-dependent mechanisms. VSMCs express retinoid receptor proteins, ie, retinoic acid receptor (RAR) alpha and retinoid X receptor (RXR) alpha. Long-term exposure to 1 micromol/L all-trans retinoic acid (RA) dose-dependently inhibited Ang II-induced cell proliferation (P<0.005) as well as DNA and protein synthesis (P<0.001). All-trans RA blocked Ang II stimulation of transforming growth factor beta(1) mRNA (P<0.005). All-trans RA inhibition of vascular VSMC growth was mediated both via RAR- and RXR-dependent pathways, as shown by receptor-specific synthetic retinoids. Transfection experiments revealed that inhibition of AP-1 dependent gene transcription is one mechanism by which all-trans RA inhibits Ang II action. RARalpha cotransfection enhanced the anti-AP-1 effects of all-trans RA dose-dependently. AP-1 activity was similarly inhibited by cotransfection with either RARalpha or RXRalpha. Ang II-induced gene expression of c-fos was abrogated by all-trans RA treatment (P<0.005). In VSMCs, all-trans RA downregulated AT(1) receptor mRNA (P<0.01) and reduced B(max) (P<0.001). All trans RA repressed Ang II-stimulated AT(1) receptor promoter activity. The all trans RA inhibitory effect was abolished when the AP-1 consensus site on the AT(1) receptor promoter was deleted. Our findings demonstrate that retinoids are potent inhibitors of the actions of Ang II on VSMCs. The findings support the notion that retinoids may interfere with proliferative vascular disease. PMID- 11282900 TI - Inflammation and thrombosis: the clot thickens. PMID- 11282902 TI - Randomized trial of aspirin, sibrafiban, or both for secondary prevention after acute coronary syndromes. AB - BACKGROUND: The first Sibrafiban Versus Aspirin to Yield Maximum Protection From Ischemic Heart Events Post-Acute Coronary Syndromes (SYMPHONY) trial showed no benefit of 2 doses of sibrafiban over aspirin for secondary prevention after acute coronary syndromes. In 2nd SYMPHONY, we compared low-dose sibrafiban plus aspirin (LDS+A), high-dose sibrafiban (HDS), and aspirin alone. METHODS AND RESULTS: When the first SYMPHONY results became known, enrollment in 2nd SYMPHONY was stopped prematurely at 6671 patients who had been treated for a median of 90 days. The primary end point of death, myocardial (re)infarction (MI), or severe recurrent ischemia did not differ significantly between aspirin (9.3%) and LDS+A (9.2%; OR, 0.98; 95% CI, 0.80 to 1.20) or HDS (10.5%; OR, 1.14; 95% CI, 0.9 to 1.39) patients. Secondary end points did not differ significantly between aspirin and LDS+A patients. Death or MI occurred significantly more often with HDS (OR, 1.43; 95% CI, 1.14 to 1.80), as did mortality alone (OR, 1.83; 95% CI, 1.17 to 2.88) and MI (OR, 1.32; 95% CI, 1.03 to 1.69). Major bleeding was significantly more frequent in LDS+A patients (5.7%) versus aspirin alone (4.0%) but not in HDS patients (4.6%). CONCLUSIONS: Combining aspirin with LDS did not improve outcomes after acute coronary syndromes and caused more bleeding compared with aspirin alone. There was a trend toward increased mortality in this group and a significant increase in the high-dose arm. PMID- 11282901 TI - Low-dose metoprolol CR/XL and fluvastatin slow progression of carotid intima media thickness: Main results from the Beta-Blocker Cholesterol-Lowering Asymptomatic Plaque Study (BCAPS). AB - BACKGROUND: Statins reduce cardiovascular events and progression of carotid intima-media thickness (IMT). beta-Blockers are also known to reduce cardiovascular events, but less is known about their effects on carotid IMT. METHODS AND RESULTS: We conducted a randomized, double-blind, placebo-controlled, single-center trial to compare the effects of low-dose metoprolol CR/XL (25 mg once daily) and fluvastatin (40 mg once daily) on the progression of carotid IMT during 36 months of treatment in 793 subjects who had carotid plaque but no symptoms of carotid artery disease. Changes in mean IMT in the common carotid artery and maximal IMT in the bulb were the main outcome variables. Death and cardiovascular events were monitored. Progression of IMT(max) in the carotid bulb at both 18 and 36 months was reduced by metoprolol CR/XL (-0.058 mm/y; 95% CI, 0.094 to -0.023; P=0.004; and -0.023 mm/y; 95% CI, -0.044 to -0.003; P=0.014, respectively). Incidence of cardiovascular events tended to be lower in metoprolol CR/XL-treated patients (5 versus 13 patients, P=0.055). Rate of IMT(mean) progression in the common carotid at 36 months was reduced by fluvastatin (-0.009 mm/y; 95% CI, -0.015 to -0.003; P=0.002). Women in the fluvastatin group had increased frequency of transiently high liver enzymes. CONCLUSIONS: This is the first randomized trial to show that a beta-blocker can reduce the rate of progression of carotid IMT in clinically healthy, symptom-free subjects with carotid plaque. This suggests that beta-blockers may have a favorable effect on atherosclerosis development. PMID- 11282903 TI - Effect of non-insulin-dependent diabetes mellitus on myocardial insulin responsiveness in patients with ischemic heart disease. AB - BACKGROUND: Patients with non-insulin-dependent diabetes mellitus (NIDDM) exhibit poor clinical outcomes from myocardial ischemia. This may reflect an impairment in their cardiac insulin-response system. METHODS AND RESULTS: We used AV balance and intracoronary infusion techniques to compare the intrinsic cardiac responsiveness to insulin in 26 coronary disease patients with (n=13) and without (n=13) NIDDM. During fasting, NIDDM hearts demonstrated lower fractional extraction of glucose from arterial plasma than controls (1.0+/-0.5% versus 2.1+/ 0.5%, P<0.05) despite higher circulating insulin levels (26+/-5 versus 13+/-4 microU. mL, P<0.05). This was compensated for by higher circulating glucose levels, so that net cardiac glucose uptake in the 2 groups was equivalent (5.2+/ 1.1 versus 5.3+/-1.1 micromol. min). Intracoronary insulin infusion produced an approximately 3-fold increase in fractional extraction and net uptake of glucose across the heart in both groups (to 3.7+/-0.4% and 18.3+/-3.5 micromol. min in NIDDM and to 5.4+/-0.7% and 17.7+/-4.3 micromol. min in controls) accompanied by an approximately 30% increase in net lactate uptake, suggesting preserved insulin action on both glucose uptake and glucose oxidation in the NIDDM heart. In nondiabetics, insulin consistently increased coronary blood flow, but this effect was absent in NIDDM. CONCLUSIONS: In contrast to their peripheral tissues and coronary vasculature, the myocardium of patients with NIDDM expresses a competent insulin-response system with respect to glucose metabolism. This suggests that insulin resistance is mediated at the level of individual organs and that different mechanisms are involved in muscle and vascular tissue. PMID- 11282904 TI - Relationship between neointimal thickness and shear stress after Wallstent implantation in human coronary arteries. AB - BACKGROUND: In-stent restenosis by excessive intimal hyperplasia reduces the long term clinical efficacy of coronary stents. Because shear stress (SS) is related to plaque growth in atherosclerosis, we investigated whether variations in SS distribution are related to variations in neointima formation. METHODS AND RESULTS: In 14 patients, at 6-month follow-up after coronary Wallstent implantation, 3D stent and vessel reconstruction was performed with a combined angiographic and intravascular ultrasound technique (ANGUS). The bare stent reconstruction was used to calculate in-stent SS at implantation, applying computational fluid dynamics. The flow was selected to deliver an average SS of 1.5 N/m(2). SS and neointimal thickness (Th) values were obtained with a resolution of 90 degrees in the circumferential and 2.5 mm in the longitudinal direction. For each vessel, the relationship between Th and SS was obtained by linear regression analysis. Averaging the individual slopes and intercepts of the regression lines summarized the overall relationship. Average Th was 0.44+/-0.20 mm. Th was inversely related to SS: Th=(0.59+/-0.24)-(0.08+/-0.10)xSS (mm) (P<0.05). CONCLUSIONS: These data show for the first time in vivo that the Th variations in Wallstents at 6-month follow-up are inversely related to the relative SS distribution. These findings support a hemodynamic mechanism underlying in-stent neointimal hyperplasia formation. PMID- 11282905 TI - Abnormalities of hemorheological, endothelial, and platelet function in patients with chronic heart failure in sinus rhythm: effects of angiotensin-converting enzyme inhibitor and beta-blocker therapy. AB - BACKGROUND: To investigate the hypothesis that abnormalities of hemorheological (fibrinogen, plasma viscosity), endothelial (von Willebrand factor [vWF]), and platelet (soluble P-selectin) function would exist in patients with chronic heart failure (CHF) who are in sinus rhythm, we conducted a cross-sectional study of 120 patients with stable CHF (median ejection fraction 30%). We also hypothesized that ACE inhibitors and beta-blockers would beneficially affect the measured indices. METHODS AND RESULTS: In the cross-sectional analysis, plasma viscosity (P=0.001), fibrinogen (P=0.02), vWF (P<0.0001), and soluble P-selectin (P<0.001) levels were elevated in patients with CHF compared with healthy controls. Women demonstrated greater abnormalities of hemorheological indices and vWF than males (all P<0.05). Plasma viscosity (P=0.009) and fibrinogen (P=0.0014) levels were higher in patients with more severe symptoms (New York Heart Association [NYHA] class III-IV), but there was no relationship with left ventricular ejection fraction. When ACE inhibitors were introduced, there was a reduction in fibrinogen (repeated-measures ANOVA, P=0.016) and vWF (P=0.006) levels compared with baseline. There were no significant changes in hemorheological, endothelial, or platelet markers after the introduction of beta-blocker therapy, apart from a rise in mean platelet count (P<0.001). CONCLUSIONS: Abnormal levels of soluble P selectin, vWF, and hemorheological indices may contribute to a hypercoagulable state in CHF, especially in female patients and in those with more severe NYHA class. Treatment with ACE inhibitors improved the prothrombotic state in CHF, whereas the addition of beta-blockers did not. These positive effects of ACE inhibitors may offer an explanation for the observed reduction in ischemic events in clinical trials. PMID- 11282906 TI - Role of endothelial nitric oxide in shear stress-induced vasodilation of human microvasculature: diminished activity in hypertensive and hypercholesterolemic patients. AB - BACKGROUND: It has been proposed that flow-mediated shear stress regulates vascular tone; however, whether this operates in the human microcirculation is unknown. This study was designed to investigate the effect of shear stress on human microvascular tone, to assess the contribution of nitric oxide (NO), and to determine whether this mechanism is defective in hypertension and in hypercholesterolemia. METHODS AND RESULTS: In 9 normal controls (NC), 11 hypertensive patients (HT), and 12 hypercholesterolemic patients (HChol), arteries (internal diameter 201+/-26 microm) isolated from gluteal fat biopsies were cannulated and perfused in chambers. Shear stress was induced by increasing the flow rate from 1 to 50 microL/min after preconstriction with norepinephrine (NE). Arterial internal diameter was expressed as percent of NE-induced constriction. In NC, shear stress induced flow-dependent vasodilation from 23+/ 9% at 1 microL/min to 53+/-14% at 50 microL/min (P<0.0001), which was abolished by endothelial removal. The NO synthase inhibitor Nomega-nitro-L-arginine (L-NNA) significantly blunted this response (mean vasodilation decreased from 27+/-6% to 6+/-9%; P=0.04). HT had significant impairment of flow-mediated dilation (mean vasodilation 5+/-6%; P=0.01 versus NC), which was not affected by L-NNA. HChol had preserved flow-mediated vasodilation (mean vasodilation 24+/-7%; P=0.56 versus NC), but this was not significantly modified by L-NNA. CONCLUSIONS: In the human microvasculature, shear stress induces endothelium-dependent, NO-mediated vasodilation. This phenomenon is blunted in HT patients because of reduced activity of NO. In contrast, the HChol microvasculature has preserved shear stress-induced dilation despite diminished NO activity. PMID- 11282907 TI - Ischemic mitral regurgitation: long-term outcome and prognostic implications with quantitative Doppler assessment. AB - BACKGROUND: Myocardial infarction (MI) can directly cause ischemic mitral regurgitation (IMR), which has been touted as an indicator of poor prognosis in acute and early phases after MI. However, in the chronic post-MI phase, prognostic implications of IMR presence and degree are poorly defined. METHODS AND RESULTS: We analyzed 303 patients with previous (>16 days) Q-wave MI by ECG who underwent transthoracic echocardiography: 194 with IMR quantitatively assessed in routine practice and 109 without IMR matched for baseline age (71+/ 11 versus 70+/-9 years, P=0.20), sex, and ejection fraction (EF, 33+/-14% versus 34+/-11%, P=0.14). In IMR patients, regurgitant volume (RVol) and effective regurgitant orifice (ERO) area were 36+/-24 mL/beat and 21+/-12 mm(2), respectively. After 5 years, total mortality and cardiac mortality for patients with IMR (62+/-5% and 50+/-6%, respectively) were higher than for those without IMR (39+/-6% and 30+/-5%, respectively) (both P<0.001). In multivariate analysis, independently of all baseline characteristics, particularly age and EF, the adjusted relative risks of total and cardiac mortality associated with the presence of IMR (1.88, P=0.003 and 1.83, P=0.014, respectively) and quantified degree of IMR defined by RVol >/=30 mL (2.05, P=0.002 and 2.01, P=0.009) and by ERO >/=20 mm(2) (2.23, P=0.003 and 2.38, P=0.004) were high. CONCLUSIONS: In the chronic phase after MI, IMR presence is associated with excess mortality independently of baseline characteristics and degree of ventricular dysfunction. The mortality risk is related directly to the degree of IMR as defined by ERO and RVol. Therefore, IMR detection and quantification provide major information for risk stratification and clinical decision making in the chronic post-MI phase. PMID- 11282908 TI - Differential profile and biochemical effects of antiautonomic membrane receptor antibodies in ventricular arrhythmias and sinus node dysfunction. AB - BACKGROUND: The relationship between anti-beta-adrenergic (anti-betaR) and anti M(2)-cholinergic (anti-M2R) receptor antibodies (Abs) and cardiac arrhythmias and their biochemical effects have not been systematically investigated. METHODS AND RESULTS: We studied 41 patients, 28 with ventricular arrhythmias (primary or due to Chagas' heart disease or idiopathic dilated cardiomyopathy; group I), 13 with sinus node dysfunction (primary or caused by Chagas' heart disease; group II), and 10 healthy controls (group III). The chronotropic effects of the IgG and immunopurified anti-beta(1)RAbs or anti-M2RAbs were assessed on cultured cardiomyocytes before and after exposure to atropine and propranolol. The biochemical effects of the IgG from 9 patients from group I, 6 from group II, and 6 controls were evaluated on COS7 cells transfected with genes encoding for beta(1),beta(2)-adrenergic receptors (cAMP increment) or M(2)-cholinergic receptors (phosphatidylinositol increment). The IgG from group I patients exerted a positive chronotropic action, with a high prevalence of anti-betaRAbs (75%) and low prevalence of anti-M2RAbs (10.7%) and induced a clear-cut and long-lasting increment in cAMP. The IgG from group II patients depressed chronotropism, with a high prevalence of anti-M2RAbs (76.9%) and low prevalence of anti-betaRAbs (15.4%) and evoked a marked augmentation of phosphatidylinositol. CONCLUSIONS: Our results demonstrate a strong correlation between anti-betaRAbs and ventricular arrhythmias and anti-M2RAbs and sinus node dysfunction. Anti-betaRAbs increase and anti-M2RAbs inhibit cAMP production. These findings offer new insight into the etiology and pathophysiology of cardiac arrhythmias, with therapeutic implications. PMID- 11282910 TI - Acyl-CoA:cholesterol acyltransferase inhibitor avasimibe reduces atherosclerosis in addition to its cholesterol-lowering effect in ApoE*3-Leiden mice. AB - BACKGROUND: The present study investigated whether the ACAT inhibitor avasimibe can reduce atherogenesis independently of its cholesterol-lowering effect in ApoE*3-Leiden mice. METHODS AND RESULTS: Two groups of 15 female ApoE*3-Leiden mice were put on a high-cholesterol (HC) diet; 1 group received 0.01% (wt/wt) avasimibe mixed into the diet. The HC diet resulted in a plasma cholesterol concentration of 18.7+/-2.6 mmol/L. Addition of avasimibe lowered plasma cholesterol by 56% to 8.1+/-1.2 mmol/L, caused mainly by a reduction of and composition change in VLDL and LDL. In a separate low-cholesterol (LC) control group, plasma cholesterol was titrated to a level comparable to that of the avasimibe group (10.3+/-1.4 mmol/L) by lowering the amount of dietary cholesterol. After 22 weeks of intervention, atherosclerosis in the aortic root area was quantified. Treatment with avasimibe resulted in a 92% reduction of lesion area compared with the HC control group. Compared with the LC control, avasimibe reduced lesion area by 78%. After correction for the slight difference in cholesterol exposure between the LC control and avasimibe groups, the effect of avasimibe on lesion area (73% reduction) remained highly significant. In addition, monocyte adherence to the endothelium, free cholesterol accumulation, and lesion severity were reduced by avasimibe treatment. CONCLUSIONS: Treatment with avasimibe potently lowered plasma cholesterol levels in ApoE*3-Leiden mice and considerably reduced atherosclerotic lesion area in addition to its cholesterol-lowering effect. Because monocyte adherence to the endothelium and lesion severity were also reduced by avasimibe, treatment with avasimibe may result in higher plaque stability and therefore a reduced risk of plaque rupture. PMID- 11282909 TI - RANTES deposition by platelets triggers monocyte arrest on inflamed and atherosclerotic endothelium. AB - BACKGROUND: Circulating platelets and chemoattractant proteins, such as the CC chemokine RANTES, contribute to the activation and interaction of monocytes and endothelium and may thereby play a pivotal role in the pathogenesis of inflammatory and atherosclerotic disease. METHODS AND RESULTS: The binding of RANTES to human endothelial cells was detected by ELISA or immunofluorescence after perfusion with platelets or exposure to their supernatants. Monocyte arrest on endothelial monolayers or surface-adherent platelets was studied with a parallel-wall flow chamber and video microscopy. We show that RANTES secreted by thrombin-stimulated platelets is immobilized on the surface of inflamed microvascular or aortic endothelium and triggers shear-resistant monocyte arrest under flow conditions, as shown by inhibition with the RANTES receptor antagonist Met-RANTES or a blocking RANTES antibody. Deposition of RANTES and its effects requires endothelial activation, eg, by interleukin-1beta, and is not supported by venous endothelium or adherent platelets. Immunohistochemistry revealed that RANTES is present on the luminal surface of carotid arteries of apolipoprotein E deficient mice with early atherosclerotic lesions after wire-induced injury or cytokine exposure. In a mechanistic model of atherogenesis, monocyte adherence on endothelium covering such lesions was studied in murine carotid arteries perfused ex vivo, showing that the accumulation of monocytic cells in these carotid arteries involved RANTES receptors. CONCLUSIONS: The deposition of RANTES by platelets triggers shear-resistant monocyte arrest on inflamed or atherosclerotic endothelium. Delivery of RANTES by platelets may epitomize a novel principle relevant to inflammatory or atherogenic monocyte recruitment from the circulation. PMID- 11282911 TI - Combined and individual mitochondrial HSP60 and HSP10 expression in cardiac myocytes protects mitochondrial function and prevents apoptotic cell deaths induced by simulated ischemia-reoxygenation. AB - BACKGROUND: The mitochondrial heat-shock proteins HSP60 and HSP10 form a mitochondrial chaperonin complex, and previous studies have shown that their increased expression exerts a protective effect against ischemic injury when cardiac myocytes are submitted to simulated ischemia. The more detailed mechanisms by which such a protective effect occurs are currently unclear. We wanted to determine whether HSP60 and HSP10 could exert a protection against simulated ischemia and reoxygenation (SI/RO)-induced apoptotic cell death and whether such protection results from decreased mitochondrial cytochrome c release and caspase-3 activation and from the preservation of ATP levels by preservation of the electron transport chain complexes. In addition, we explored whether increased expression of HSP60 or HSP10 by itself exerts a protective effect. METHODS AND RESULTS: We overexpressed HSP60 and HSP10 together or separately in rat neonatal cardiac myocytes using an adenoviral vector and then subjected the myocytes to SI/RO. Cell death and apoptosis in myocytes were quantified by parameters such as enzyme release, DNA fragmentation, and caspase-3 activation. Overexpression of the combination of HSP60 and HSP10 and of HSP60 or HSP10 individually protected myocytes against apoptosis. This protection is accompanied by decreases in mitochondrial cytochrome c release and in caspase-3 activity and increases in ATP recovery and activities of complex III and IV in mitochondria after SI/RO. CONCLUSIONS: These results suggest that mitochondrial chaperonins HSP60 and HSP10 in combination or individually play an important role in maintaining mitochondrial integrity and capacity for ATP generation, which are the crucial factors in determining survival of cardiac myocytes undergoing ischemia/reperfusion injury. PMID- 11282912 TI - Liquid-filled balloon brachytherapy using (68)Ga is effective and safe because of the short 68-minute half-life: results of a feasibility study in the porcine coronary overstretch model. AB - BACKGROUND: Liquid-filled balloons for coronary brachytherapy provide significant advantages over solid sources in dose homogeneity but carry the risk of life threatening radiointoxication after balloon rupture and laboratory contamination in case of a spill. We hypothesized that the positron emitter (68)Ga, with a half life of only 68 minutes, was well suited to overcome these safety obstacles while providing full therapeutic efficacy. METHODS AND RESULTS: The feasibility, efficacy, and safety of (68)Ga liquid-filled balloon brachytherapy were investigated in the porcine coronary overstretch model. Four groups of 5 balloon induced coronary lesions were irradiated with 8, 12, 16, and 24 Gy targeted to the adventitia. Ten unirradiated lesions served as controls. Segments treated with 16 or 24 Gy exhibited marked suppression of neointimal proliferation at 28 day follow-up, with quantitative parameters of intraluminal proliferation reduced to <20%. This beneficial effect was not compromised by untoward edge effects. Uninjured but irradiated vessels did not show histological signs of radiation damage. The (68)Ga whole-body dose due to balloon rupture was estimated to be 5 rem/50 mCi treatment activity and compared favorably with that of (188)Re (78 rem/50 mCi). CONCLUSIONS: (68)Ga positron radiation suppresses neointimal proliferation at doses of 16 and 24 Gy. This biological efficacy, coupled with the attractive safety profile, suggests the selection of (68)Ga as an attractive isotope for liquid-filled balloon brachytherapy. PMID- 11282913 TI - Anti-inflammatory, antithrombotic, and neuroprotective effects of activated protein C in a murine model of focal ischemic stroke. AB - BACKGROUND: Activated protein C (APC) contributes to systemic anticoagulant and anti-inflammatory activities. APC may reduce organ damage by inhibiting thrombin generation and leukocyte activation. Neutrophils and cerebrovascular thrombosis contribute to ischemic neuronal injury, suggesting that APC may be a potential protective agent for stroke. METHODS AND RESULTS: We examined the effects of APC in a murine model of focal ischemia. After middle cerebral artery occlusion/reperfusion, the average survival time in controls was 13.6 hours. Animals that received purified human plasma-derived APC 2 mg/kg IV either 15 minutes before or 10 minutes after stroke induction survived 24 hours and were killed for neuropathological analysis. APC 2 mg/kg given before or after onset of ischemia restored cerebral blood flow, reduced brain infarct volume (59% to 69%; P:<0.003) and brain edema (50% to 61%; P:<0.05), eliminated brain infiltration with neutrophils, and reduced the number of fibrin-positive cerebral vessels by 57% (P:<0.05) and 25% (nonsignificant), respectively. The neuroprotective effect of APC was dose-dependent and associated with significant inhibition of ICAM-1 expression on ischemic cerebral blood vessels (eg, 61% inhibition with 2 mg/kg APC). Intracerebral bleeding was not observed with APC. CONCLUSIONS: APC exerts anti-inflammatory, antithrombotic, and neuroprotective effects in stroke. Central effects of APC are likely to be related to improved maintenance of the blood brain barrier to neutrophils and to reduced microvascular obstructions and fibrin deposition. PMID- 11282914 TI - In utero remodeling of the fetal lamb ductus arteriosus: the role of antenatal indomethacin and avascular zone thickness on vasa vasorum proliferation, neointima formation, and cell death. AB - BACKGROUND: The ductus arteriosus (DA) of newborn infants exposed in utero to indomethacin is resistant to postnatal indomethacin; we hypothesized that this is due to ductus constriction in utero, with subsequent remodeling of the vessel. METHODS AND RESULTS: Infusion of fetal lambs with indomethacin for 48 hours constricted the DA and increased the thickness of the avascular zone of the DA, which in turn induced the expression of vascular endothelial growth factor, endothelial nitric oxide synthase (due to ingrowth of vasa vasorum), neointima formation, and loss of smooth muscle cells; moderate degrees of DA constriction in utero increased NO production, which inhibited DA contractility. Marked degrees of DA constriction decreased tissue distensibility and contractile capacity. CONCLUSIONS: DA patency is no longer controlled primarily by prostaglandins once it has been exposed to indomethacin in utero. PMID- 11282915 TI - High-sensitivity C-reactive protein: potential adjunct for global risk assessment in the primary prevention of cardiovascular disease. AB - Inflammation plays a major role in atherothrombosis, and measurement of inflammatory markers such as high-sensitivity C-reactive protein (HSCRP) may provide a novel method for detecting individuals at high risk of plaque rupture. Several large-scale prospective studies demonstrate that HSCRP is a strong independent predictor of future myocardial infarction and stroke among apparently healthy men and women and that the addition of HSCRP to standard lipid screening may improve global risk prediction among those with high as well as low cholesterol levels. Because agents such as aspirin and statins seem to attenuate inflammatory risk, HSCRP may also have utility in targeting proven therapies for primary prevention. Inexpensive commercial assays for HSCRP are now available; they have shown variability and classification accuracy similar to that of cholesterol screening. Risk prediction algorithms using a simple quintile approach to HSCRP evaluation have been developed for outpatient use. Thus, although limitations inherent to inflammatory screening remain, available data suggest that HSCRP has the potential to play an important role as an adjunct for global risk assessment in the primary prevention of cardiovascular disease. PMID- 11282916 TI - Traumatic communication between aorta and left atrium diagnosed by transesophageal echocardiography. PMID- 11282917 TI - Can a Mediterranean-style diet reduce heart disease? PMID- 11282918 TI - AHA Science Advisory: Lyon Diet Heart Study. Benefits of a Mediterranean-style, National Cholesterol Education Program/American Heart Association Step I Dietary Pattern on Cardiovascular Disease. PMID- 11282919 TI - Anomalous origin of right coronary artery. PMID- 11282965 TI - Sleepy dogs don't lie: a genetic disorder informative about sleep. PMID- 11282966 TI - Centromere on the move. PMID- 11282967 TI - Characterization of clustered MHC-linked olfactory receptor genes in human and mouse. AB - Olfactory receptor (OR) loci frequently cluster and are present on most human chromosomes. They are members of the seven transmembrane receptor (7-TM) superfamily and, as such, are part of one of the largest mammalian multigene families, with an estimated copy number of up to 1000 ORs per haploid genome. As their name implies, ORs are known to be involved in the perception of odors and possibly also in other, nonolfaction-related, functions. Here, we report the characterization of ORs that are part of the MHC-linked OR clusters in human and mouse (partial sequence only). These clusters are of particular interest because of their possible involvement in olfaction-driven mate selection. In total, we describe 50 novel OR loci (36 human, 14 murine), making the human MHC-linked cluster the largest sequenced OR cluster in any organism so far. Comparative and phylogenetic analyses confirm the cluster to be MHC-linked but divergent in both species and allow the identification of at least one ortholog that will be useful for future regulatory and functional studies. Quantitative feature analysis shows clear evidence of duplications of blocks of OR genes and reveals the entire cluster to have a genomic environment that is very different from its neighboring regions. Based on in silico transcript analysis, we also present evidence of extensive long-distance splicing in the 5'-untranslated regions and, for the first time, of alternative splicing within the single coding exon of ORs. Taken together with our previous finding that ORs are also polymorphic, the presented data indicate that the expression, function, and evolution of these interesting genes might be more complex than previously thought. PMID- 11282968 TI - Identification and functional analysis of mutations in the hypocretin (orexin) genes of narcoleptic canines. AB - Narcolepsy is a sleep disorder affecting animals and humans. Exon skipping mutations of the Hypocretin/Orexin-receptor-2 (Hcrtr2) gene were identified as the cause of narcolepsy in Dobermans and Labradors. Preprohypocretin (Hcrt) knockout mice have symptoms similar to human and canine narcolepsy. In this study, 11 sporadic cases of canine narcolepsy and two additional multiplex families were investigated for possible Hcrt and Hcrtr2 mutations. Sporadic cases have been shown to have more variable disease onset, increased disease severity, and undetectable Hypocretin-1 levels in cerebrospinal fluid. The canine Hcrt locus was isolated and characterized for this project. Only one novel mutation was identified in these two loci. This alteration results in a single amino acid substitution (E54K) in the N-terminal region of the Hcrtr2 receptor and autosomal recessive transmission in a Dachshund family. Functional analysis of previously described exon-skipping mutations and of the E54K substitution were also performed using HEK-293 cell lines transfected with wild-type and mutated constructs. Results indicate a truncated Hcrtr2 protein, an absence of proper membrane localization, and undetectable binding and signal transduction for exon skipping mutated constructs. In contrast, the E54K abnormality was associated with proper membrane localization, loss of ligand binding, and dramatically diminished calcium mobilization on activation of the receptor. These results are consistent with a loss of function for all three mutations. The absence of mutation in sporadic cases also indicates genetic heterogeneity in canine narcolepsy, as reported previously in humans. PMID- 11282969 TI - Genome-scale compositional comparisons in eukaryotes. AB - We examined dinucleotide relative abundances and their biases in recent sequences of eukaryotic genomes and chromosomes, including human chromosomes 21 and 22, Saccharomyces cerevisiae, Arabidopsis thaliana, and Drosophila melanogaster. We found that dinucleotide relative abundances are remarkably constant across human chromosomes and within the DNA of a particular species. The dinucleotide biases differ between species, providing a genome signature that is characteristic of the bulk properties of an organism's DNA. We detail the relations between species genome signatures and suggest possible mechanisms for their origin and maintenance. PMID- 11282970 TI - Comparing genomes within the species Mycobacterium tuberculosis. AB - The study of genetic variability within natural populations of pathogens may provide insight into their evolution and pathogenesis. We used a Mycobacterium tuberculosis high-density oligonucleotide microarray to detect small-scale genomic deletions among 19 clinically and epidemiologically well-characterized isolates of M. tuberculosis. The pattern of deletions detected was identical within mycobacterial clones but differed between different clones, suggesting that this is a suitable genotyping system for epidemiologic studies. An analysis of genomic deletions among an extant population of pathogenic bacteria provided a novel perspective on genomic organization and evolution. Deletions are likely to contain ancestral genes whose functions are no longer essential for the organism's survival, whereas genes that are never deleted constitute the minimal mycobacterial genome. As the amount of genomic deletion increased, the likelihood that the bacteria will cause pulmonary cavitation decreased, suggesting that the accumulation of mutations tends to diminish their pathogenicity. Array-based comparative genomics is a promising approach to exploring molecular epidemiology, microbial evolution, and pathogenesis. PMID- 11282971 TI - Lineage-specific gene expansions in bacterial and archaeal genomes. AB - Gene duplication is an important mechanistic antecedent to the evolution of new genes and novel biochemical functions. In an attempt to assess the contribution of gene duplication to genome evolution in archaea and bacteria, clusters of related genes that appear to have expanded subsequent to the diversification of the major prokaryotic lineages (lineage-specific expansions) were analyzed. Analysis of 21 completely sequenced prokaryotic genomes shows that lineage specific expansions comprise a substantial fraction (approximately 5%-33%) of their coding capacities. A positive correlation exists between the fraction of the genes taken up by lineage-specific expansions and the total number of genes in a genome. Consistent with the notion that lineage-specific expansions are made up of relatively recently duplicated genes, >90% of the detected clusters consists of only two to four genes. The more common smaller clusters tend to include genes with higher pairwise similarity (as reflected by average score density) than larger clusters. Regardless of size, cluster members tend to be located more closely on bacterial chromosomes than expected by chance, which could reflect a history of tandem gene duplication. In addition to the small clusters, almost all genomes also contain rare large clusters of size > or =20. Several examples of the potential adaptive significance of these large clusters are explored. The presence or absence of clusters and their related genes was used as the basis for the construction of a similarity graph for completely sequenced prokaryotic genomes. The topology of the resulting graph seems to reflect a combined effect of common ancestry, horizontal transfer, and lineage specific gene loss. PMID- 11282972 TI - A comparative genomics approach to prediction of new members of regulons. AB - Identifying the complete transcriptional regulatory network for an organism is a major challenge. For each regulatory protein, we want to know all the genes it regulates, that is, its regulon. Examples of known binding sites can be used to estimate the binding specificity of the protein and to predict other binding sites. However, binding site predictions can be unreliable because determining the true specificity of the protein is difficult because of the considerable variability of binding sites. Because regulatory systems tend to be conserved through evolution, we can use comparisons between species to increase the reliability of binding site predictions. In this article, an approach is presented to evaluate the computational predictions of regulatory sites. We combine the prediction of transcription units having orthologous genes with the prediction of transcription factor binding sites based on probabilistic models. We augment the sets of genes in Escherichia coli that are expected to be regulated by two transcription factors, the cAMP receptor protein and the fumarate and nitrate reduction regulatory protein, through a comparison with the Haemophilus influenzae genome. At the same time, we learned more about the regulatory networks of H. influenzae, a species with much less experimental knowledge than E. coli. By studying orthologous genes subject to regulation by the same transcription factor, we also gained understanding of the evolution of the entire regulatory systems. PMID- 11282973 TI - The complex repeats of Dictyostelium discoideum. AB - In the course of determining the sequence of the Dictyostelium discoideum genome we have characterized in detail the quantity and nature of interspersed repetitive elements present in this species. Several of the most abundant small complex repeats and transposons (DIRS-1; TRE3-A,B; TRE5-A; skipper; Tdd-4; H3R) have been described previously. In our analysis we have identified additional elements. Thus, we can now present a complete list of complex repetitive elements in D. discoideum. All elements add up to 10% of the genome. Some of the newly described elements belong to established classes (TRE3-C, D; TRE5-B,C; DGLT-A,P; Tdd-5). However, we have also defined two new classes of DNA transposable elements (DDT and thug) that have not been described thus far. Based on the nucleotide amount, we calculated the least copy number in each family. These vary between <10 up to >200 copies. Unique sequences adjacent to the element ends and truncation points in elements gave a measure for the fragmentation of the elements. Furthermore, we describe the diversity of single elements with regard to polymorphisms and conserved structures. All elements show insertion preference into loci in which other elements of the same family reside. The analysis of the complex repeats is a valuable data resource for the ongoing assembly of whole D. discoideum chromosomes. PMID- 11282974 TI - Centromere emergence in evolution. AB - Evolutionary centromere repositioning is a paradox we have recently discovered while studying the conservation of the phylogenetic chromosome IX in primates. Two explanations were proposed: a conservative hypothesis assuming sequential pericentric inversions, and a more challenging assumption involving centromere emergence during evolution. The complex evolutionary history showed by chromosome IX did not allow us to clearly distinguish between these two hypotheses. Here we report comparative studies of chromosome X in two lemur species: the black lemur and the ringtailed lemur. The X chromosome is telocentric in the black lemur and almost metacentric in the ringtailed lemur. The marker order along these chromosomes, however, was found to be perfectly colinear with humans. Our data unequivocally point to centromere emergence as the most likely explanation of centromere repositioning. PMID- 11282975 TI - Homogeneous assays for single-nucleotide polymorphism typing using AlphaScreen. AB - AlphaScreen technology allows the development of high-throughput homogeneous proximity assays. In these assays, signal is generated when 680 nm laser light irradiates a donor bead in close proximity to an acceptor bead. For the detection of nucleic acids, donor and acceptor beads are brought into proximity by two bridging probes that hybridize simultaneously to a common target and to the generic oligonucleotides attached covalently to the beads. This method allows the detection of as little as 10 amole of a single-stranded DNA target. The combination of AlphaScreen with allele-specific amplification (ASA) and allele specific hybridization (ASH) has allowed the development of two homogenous single nucleotide polymorphism (SNP) genotyping platforms. Both types of assay are very robust, routinely giving accurate genotyping results with < 2 ng of genomic DNA per genotype. An AlphaScreen validation study was performed for 12 SNPs by using ASA assays and seven SNPs by using ASH assays. More than 580 samples were genotyped with accuracy >99%. The two assays are remarkably simple, requiring no post-PCR manipulations. Genotyping has been performed successfully in 96- and 384 well formats with volumes as small as 2 microL, allowing a considerable reduction in the amount of reagents and genomic DNA necessary for genotyping. These results show that the AlphaScreen technology can be successfully adapted to high throughput genotyping. PMID- 11282976 TI - Self-reporting PNA/DNA primers for PCR analysis. AB - We report a new fluorogenic method for sealed-tube PCR analysis using a quencher labeled peptide nucleic acid (Q-PNA) probe. The Q-PNA hybridizes to a complementary tag sequence located at the 5' end of a 5' fluorophore-labeled oligonucleotide primer, quenching the primer's fluorescence. Incorporation of the primer into a doublestranded amplicon causes displacement of the Q-PNA such that the fluorescence of the sample is a direct indication of the amplicon concentration. The Q-PNA is able to quench multiple primers bearing distinct 5' fluorophores in a single reaction. We show realtime quantitative detection of a single-copy gene, K-ras, from human genomic DNA, as well as an endpoint multiplex assay for Chlamydia trachomatis and Neisseria gonorrhoeae targets. Because the Q PNA may be used to quench any primer that contains the 5' tag sequence, it is possible to inexpensively adapt an existing primer set for use in a self reporting fluorescent assay by including the tag sequence in one of the primers. PMID- 11282977 TI - Automated finishing with autofinish. AB - Currently, the genome sequencing community is producing shotgun sequence data at a very high rate, but finishing (collecting additional directed sequence data to close gaps and improve the quality of the data) is not matching that rate. One reason for the difference is that shotgun sequencing is highly automated but finishing is not: Most finishing decisions, such as which directed reads to obtain and which specialized sequencing techniques to use, are made by people. If finishing rates are to increase to match shotgun sequencing rates, most finishing decisions also must be automated. The Autofinish computer program (which is part of the computer software package) does this by automatically choosing finishing reads. Autofinish is able to suggest most finishing reads required for completion of each sequencing project, greatly reducing the amount of human attention needed. sometimes completely finishes the project, with no human decisions required. It cannot solve the most complex problems, so we recommend that Autofinish be allowed to suggest reads for the first three rounds of finishing, and if the project still is not finished completely, a human finisher complete the work. We compared this Autofinish-Hybrid method of finishing against a human finisher in five different projects with a variety of shotgun depths by finishing each project twice--once with each method. This comparison shows that the Autofinish-Hybrid method saves many hours over a human finisher alone, while using roughly the same number and type of reads and closing gaps at roughly the same rate. Autofinish currently is in production use at several large sequencing centers. It is designed to be adaptable to the finishing strategy of the lab--it can finish using some or all of the following: resequencing reads, reverses, custom primer walks on either subclone templates or whole clone templates, PCR, or minilibraries. Autofinish has been used for finishing cDNA, genomic clones, and whole bacterial genomes (see http://www.phrap.org). PMID- 11282978 TI - Sequence evaluation of four pooled-tissue normalized bovine cDNA libraries and construction of a gene index for cattle. AB - An essential component of functional genomics studies is the sequence of DNA expressed in tissues of interest. To provide a resource of bovine-specific expressed sequence data and facilitate this powerful approach in cattle research, four normalized cDNA libraries were produced and arrayed for high-throughput sequencing. The libraries were made with RNA pooled from multiple tissues to increase efficiency of normalization and maximize the number of independent genes for which sequence data were obtained. Target tissues included those with highest likelihood to have impact on production parameters of animal health, growth, reproductive efficiency, and carcass merit. Success of normalization and inter- and intralibrary redundancy were assessed by collecting 6000-23,000 sequences from each of the libraries (68,520 total sequences deposited in GenBank). Sequence comparison and assembly of these sequences was performed in combination with 56,500 other bovine EST sequences present in the GenBank dbEST database to construct a cattle Gene Index (available from The Institute for Genomic Research at http://www.tigr.org/tdb/tgi.shtml). The 124,381 bovine ESTs present in GenBank at the time of the analysis form 16,740 assemblies that are listed and annotated on the Web site. Analysis of individual library sequence data indicates that the pooled-tissue approach was highly effective in preparing libraries for efficient deep sequencing. PMID- 11282979 TI - Signaling through CD38 induces NK cell activation. AB - Human CD38 is a signal transduction molecule, and, concurrently, an ectoenzyme catalyzing the synthesis and degradation of cyclic ADP-ribose (cADPR), a potent Ca2+ mobilizer. One facet of CD38 that has not yet been addressed is its role in NK cells. To this end, the events triggered by CD38 ligation with agonistic mAb were analyzed on freshly purified human NK cells. Ligation was followed by (i) a significant rise in the intracellular level of Ca2+, (ii) increased expression of HLA class II and CD25, and (iii) tyrosine phosphorylation of discrete cytoplasmic substrates. The phosphorylation cascade involved CD3-zeta and FcepsilonRIgamma chains, zeta-associated protein (ZAP)-70 and the proto-oncogene product c-Cbl. NK effector functions were then analyzed: CD38 signaling was able (iv) to induce release of IFN-gamma and, more prominently, of granulocyte macrophage colony stimulating factor, as assessed by measuring both mRNA and protein products; and, lastly, (v) to induce cytolytic effector functions on target cells after IL-2 activation, as shown both by cytotoxicity assays and ultrastructural changes. The tyrosine-phosphorylated substrates and all the effects mediated by CD38 were similar to those observed following triggering via CD16 (FcgammaRIIIA); moreover, Ca2+ mobilization via CD38 no longer operated in NK-derived cell lines lacking CD16. These results suggest that the activation signals transduced by CD38 in NK cells elicit relevant cellular events. The effects are similar to those elicited via CD16 and possibly rely on common signaling pathways. PMID- 11282980 TI - Suicide induced by cytolytic activity controls the differentiation of memory CD8(+) T lymphocytes. AB - Cytotoxic T lymphocytes (CTL) confer protection against intracellular pathogens, yet the mechanism by which some escape activation induced cell death (AICD) and give rise to long-lived memory cells is unclear. We studied the differentiation of transgenic TCR CD8(+) cells into CTL and memory cells using a novel system that allowed us to control cytolytic activity. The perforin/granzyme granules used to lyse targets induced the apoptosis of CTL in a fratricide-independent manner. After adoptive transfer to antigen-free mice, the ability of CTL to give generate memory cells was determined. We found that the extent of cytolysis by a common pool of CTL controlled the differentiation into memory cells, which were only generated under conditions of minimal cytolytic activity. Thus, the differentiation of naive CD8(+) cells into memory cells may not depend on the presence on a subset of committed CTL precursors, but rather is controlled by the extent of granule-mediated cytolysis. PMID- 11282981 TI - Chondrocyte antigen expression, immune response and susceptibility to arthritis. AB - The association of HLA-B27 with certain forms of arthritis implies a role for MHC class I-restricted T cells in the arthritic process. Our aim was to study CD8(+) T cell responses towards specific antigens localized in joint tissue. Known determinants were introduced into chondrocytes of transgenic (TG) mice, under the control of the cis-regulatory sequences of the human type II collagen gene (COL2A1). Two Escherichia coli beta-galactosidase (beta-gal)-expressing lines were derived (CIIL73 and CIIL64) as well as two lines (CIINP) expressing influenza A virus nucleoprotein (NP). Expression of the antigens could be demonstrated in cartilaginous tissues. The TG lines showed variable degrees of responsiveness towards the transgene-introduced antigens; whilst 75% of CIIL73 mice had an impaired cytotoxic T lymphocyte (CTL) response towards beta-gal, the response in CIIL64 mice was essentially normal. However, both lines displayed normal proliferative and antibody responses to beta-gal. A reduced CTL response was seen to NP in the CIINP lines in approximately 65% of the animals. In spite of the persistence of T cell responses to the transgene antigens in these lines, induction of CTL responses alone has so far failed to induce clinical signs of arthritis. Interestingly, some animals expressing beta-gal were susceptible to arthritis following challenge with type II collagen alone, whilst their non-TG littermates and TG mice from other lines remained unaffected. As beta-gal is expressed by E. coli, a component of the normal gut flora, this suggests a possible role for gut-derived immune responses. We believe these lines could form the basis of a model for studying links between intestinal inflammation and arthritis. PMID- 11282982 TI - Chronic exposure to superantigen induces regulatory CD4(+) T cells with IL-10 mediated suppressive activity. AB - The repeated injection of bacterial superantigens (SAg), such as staphylococcus enterotoxin (SE) A or B, has been shown in mice to induce a state of unresponsiveness characterized by the lack of secretion of Th1 lymphokines, such as IL-2 and IFN-gamma, following subsequent SAg challenge. We made the observation, in vivo as well as in vitro, that unresponsiveness to SAg could be transferred from SEA- to SEB-reactive T cells (and reversibly from SEB- to SEA specific T cells) in C57BL/6 mice but not in BALB/c mice. Since C57BL/6 mice, unlike BALB/c mice, possess TCR V(beta)3+ and V(beta)11+ T cells able to react with both SEA and SEB, we hypothesized that SAg-unresponsive V(beta)3(+) and V(beta)11+ T cells could mediate linked suppression of other SAg-reactive T cells. To analyze further this possibility, spleen cells from BALB/c mice made unresponsive to SEB were tested for their capacity to suppress the response of normal BALB/c cells to SEB. The production of both IFN-gamma and IL-2 following SEB stimulation was greatly impaired in co-cultures containing CD4(+) T cells, but not CD8(+) T cells, isolated from unresponsive animals. In vivo, the production of both IFN-gamma and IL-2 responses to SEB was dramatically reduced in animals adoptively transferred with unresponsive spleen cells. This suppression was abrogated in recipients injected with neutralizing anti-IL-10 antibodies. Moreover, in animals made unresponsive to SEB, SAg-reactive CD4(+) T cells were found to express high levels of CTLA-4, a molecule recently described to play an essential role in the suppressive function of regulatory T cells. Taken together these results demonstrate that the repetitive injection of SAg induces the differentiation of regulatory CD4(+) T cells capable of suppressing SAg-reactive naive T cells. PMID- 11282983 TI - Multiple co-stimulatory signals are required for triggering proliferation of T cells from human secondary lymphoid tissue. AB - Vaccine-based therapies are being developed for a variety of cancers and their efficacy will be determined by their ability to stimulate T cells in the secondary lymphoid tissue. We found that T cells isolated from human secondary lymphoid organs (LT-T), in contrast to peripheral blood T cells (PB-T) are hyporesponsive to cross-linked anti-CD3 mAb (CD3c) even in the presence of exogenous IL-2. Using mAb to trigger CD2 and CD28 co-stimulatory molecules, we found that such dual co-stimulation of LT-T induces profound and sustained responses including CD25 expression, IL-2 secretion and proliferation. Different levels of co-stimulation produced a hierarchical pattern of responses in LT-T, which correlated with the degree of CD3-TCR down-regulation. Mature antigen presenting cells (APC) restored the capacity of LT-T to proliferate to stimulation of the CD3-TCR complex. Blocking studies demonstrated that optimal proliferation was critically dependent on co-stimulation via CD2 and CD28 engaged by their ligands on the APC. Therefore, LT-T have increased co-stimulatory requirements as compared to PB-T, i.e. multiple co-stimulatory signals coupled to CD3-TCR triggering. Furthermore, LT-T were found to be dependent on APC for survival, in contrast to PB-T. Clearly, LT-T do not behave in a comparable way to PB-T and in vitro experiments assessing novel cancer vaccines should therefore use LT-T as the most appropriate population of responder T cells. PMID- 11282984 TI - Dendritic cells infected with Mycobacterium bovis bacillus Calmette Guerin activate CD8(+) T cells with specificity for a novel mycobacterial epitope. AB - Although CD4(+) T cells are essential for protective immunity against Mycobacterium tuberculosis infection, recent reports indicate that CD8(+) T cells may also play a critical role in the control of this infection. However, the epitope specificity and the mechanisms of activation of mycobacteria-reactive CD8(+) T cells are poorly characterized. In order to study the CD8(+) T cell responses to the model mycobacterial antigen, MPT64, we used recombinant vaccinia virus expressing MPT64 (VVWR-64) and a panel of MPT64-derived peptides to establish that the peptide MPT64(190-198) contains an H-2D(b)-restricted CD8(+) T cell epitope. A cytotoxic T lymphocyte response to this peptide could be demonstrated in M. bovis bacillus Calmette Guerin (BCG)-infected mice following repeated in vitro stimulation. When bone marrow-derived dendritic cells (DC) were infected with BCG, the expression of MHC class I molecules by DC was up-regulated in parallel with MHC class II and B7-2, whereas CD1d expression level was not modified. Moreover, BCG-infected DC activated MPT64(190-198)-specific CD8(+) T cells to secrete IFN-gamma, although with a lower efficacy than VVWR-64-infected DC. The production of IFN-gamma by MPT64(190-198)-specific CD8(+) T cells was inhibited by antibodies to MHC class I, but not to CD1d. These data suggest that mycobacteria-specific CD8(+) T cells are primed during infection. Therefore, anti mycobacterial vaccine strategies targeting the activation of specific CD8(+) T cells by DC may have improved protective efficacy. PMID- 11282985 TI - NK cells and NKT cells collaborate in host protection from methylcholanthrene induced fibrosarcoma. AB - NK1.1(+) V(alpha)14J(alpha)281(+) (NKT) cells can be induced by IL-12 therapy to mediate tumor rejection; however, methylcholanthrene (MCA)-induced fibrosarcoma is the only tumor model described where NKT cells play a natural role in controlling tumor initiation. From our previous study in C57BL/6 mice it remained unclear whether NK cells were also involved in this natural response. Herein, to discriminate the function of NK and NKT cells, we have evaluated fibrosarcoma development in mice deficient in NKT cells, but not NK cells, and mice deficient in NK cells, but not NKT cells. The results indicate that both NK cells and NKT cells are essential and collaborate in natural host immunity against MCA-induced sarcoma. In contrast, sarcoma incidence and growth rate were reduced using IL-12 therapy, this effect was mediated in the absence of T cells (including NKT cells), but not NK cells. PMID- 11282986 TI - Differential effect of CD8(+) and CD8(-) dendritic cells in the stimulation of secondary CD4(+) T cells. AB - Dendritic cells (DC), in their role in initiation of the adaptive immune response, have been extensively studied for their capacity to interact and stimulate naive T cells. Subsets of mature murine DC isolated directly from the spleen have been shown to differ in their ability to induce proliferative responses in both primary CD4(+) and primary CD8(+) T cells; the myeloid-related CD8alpha(-) DC induce a more intense or prolonged proliferation of naive T cells than do the lymphoid-related DC bearing CD8alpha despite similar expression of MHC and co-stimulatory molecules. Here we examine the interaction of these DC subpopulations with T cells already in the activated or memory state which are known to have greater sensitivity to antigen stimulation and bear receptors with increased capacity for signal transduction. We show that influenza virus-specific CD4(+) T cell clones and splenic T cells from peptide-primed animals proliferated in response to antigen presented by separated splenic CD8(-) DC. In contrast, these T cells showed only weak, if any, proliferation in response to CD8(+) DC despite observable cluster formation in the cultures. The differential between the two DC types in inducing proliferation was even more pronounced than previously seen with primary T cells and did not reflect differential longevity of the DC in culture, altered response kinetics or deviation from IL-2 to IL-4 induction with CD8(+) DC, but was related to the levels of IL-2 induced. The deficiency in the CD8(+) DC was not overcome by using infectious virus rather than synthetic peptide as the antigen source. These results show that lymphoid related CD8(+) splenic DC, despite their mature phenotype, fail to provide appropriate signals to secondary CD4(+) T cells to sustain their proliferation. PMID- 11282987 TI - Predominance of a novel splenic B cell population in mice expressing a transgene that encodes multireactive antibodies: support for additional heterogeneity of the B cell compartment. AB - We generated IgHmudelta transgenic mice using a V(H) gene that in A/J mice encodes multireactive BCR in the preimmune B cell compartment and is predominantly expressed by a memory B cell subpopulation. Most primary splenic B cells in these mice have a size, cell-surface phenotype and in vitro response profile distinct from mature follicular (B2), marginal zone (MZ) or B1 B cells, but are long-lived and appear to be slowly cycling. They reside in conventional B cell areas of the spleen and mount robust foreign antigen-driven germinal center responses, but do not efficiently differentiate to secretory phenotype. We propose that these qualities result from ongoing, low-avidity BCR-self-ligand interactions and promote entry into the memory pathway. Given these data, and the enormous diversity and characteristic multireactivity of the preimmune antibody repertoire, we also suggest that it may be more appropriate to view the primary B cell compartment as a continuum of functional and phenotypic 'layers', rather than as a group of discrete B1, B2 and MZ subsets. PMID- 11282989 TI - Palindromic but not G-rich sequences are targets of class switch recombination. AB - In order to understand the specificity of sequences or structures recognized by a recombinase involved in class switch recombination (CSR), we examined the relative CSR efficiency of various switch sequences in artificial CSR constructs that undergo CSR in CH12F3-2 murine B lymphoma line. Since CSR recombination is not specific to switch regions of different isotypes or orientation of S sequences, we examined the efficiency of S sequences of non-mammalian species and artificial sequences which lack several characters of mammal switch sequences: chicken S(mu), Xenopus S(mu), telomere, multiple cloning site (MCS) and unrelated negative control sequence. CSR occurred in chicken S(mu) and MCS with significantly higher efficiency than the negative control. A common character of these two sequences is that they are rich in palindrome and stem-loop structures. However, telomeres, which are G-rich and repetitive but not palindromic, could not serve as switch sequences at all. The AT-rich Xenopus S(mu) sequence was inefficient but capable of CSR. CSR breakpoint distribution suggests that the cleavage may take place preferentially in the proximity of the junctions (neck) between the loop and stem in the secondary structure of the single-stranded S sequence, which can be formed by palindromic sequences. The results suggest that the secondary structure of S-region sequences which is transiently formed during transcription may be necessary for recognition by class switch recombinase. PMID- 11282988 TI - Bruton's tyrosine kinase is required for signaling the CD79b-mediated pro-B to pre-B cell transition. AB - Formation of the pre-BCR complex is a critical check point during B cell development and induces the transition of pro-B to pre-B cells. CD79b (Igbeta) is a signaling component in the pre-BCR complex, since differentiation to the pre-B phenotype is induced by cross-linking the CD79b expressed on developmentally arrested pro-B cells from recombination-activating gene (RAG)-2-deficient mice. Bruton's tyrosine kinase (BTK) plays important roles in B cell development. However, its molecular mechanisms in early B cell development are not fully understood. To examine whether BTK functions in CD79b-mediated signaling for the pro-B/pre-B transition, we utilized RAG2/BTK double-knockout (DKO) mice. Pro-B cells from RAG2/BTK-DKO mice did not differentiate into pre-B cells following CD79b cross-linking, although tyrosine phosphorylation of cellular proteins including Erk1/2 and phospholipase C-gamma2 was induced in the same manner as RAG2-KO mice. BTK is phosphorylated after cross-linking of CD79b on RAG2 deficient pro-B cells. These findings suggest that BTK-dependent pathways downstream of CD79b are critical for the pro-B/pre-B transition and BTK independent signaling pathways are also activated via the pre-BCR complex. PMID- 11282990 TI - Longitudinal analysis of T cells responding to tetanus toxoid in healthy subjects as well as in pediatric patients after bone marrow transplantation: the identification of identical TCR-CDR3 regions in time suggests long-term stability of at least part of the antigen-specific TCR repertoire. AB - To understand the nature of long-term Th immune responses, we investigated in the present study the TCRBV gene repertoire of CD4(+) T cells specific for the recall antigen tetanus toxoid (TT) in recipients of an allogeneic bone marrow transplantation (allo-BMT) at several time points after transplantation and in their BM donors. We observed that the TCR repertoire of TT-specific CD4(+) Th cells was heterogeneous, and differed between allo-BMT recipients and their respective donors. Some individuals, however, used similar TCR-complementarity determining region (CDR) 3 motifs that could reflect recognition of and selection by similar promiscuous epitopes of TT. Longitudinal analysis of this TT-specific T cell response revealed that T cells with completely identical TCR were present at several time points after the first analysis in allo-BMT recipients, most probably reflecting long-term stability of at least part of the antigen-specific TCR repertoire. Similar stability of the TT-specific TCR repertoire in time was also noted in the allo-BMT donors. These observations reveal that within a given individual the dominant antigen-specific T cell clones persist in time in an otherwise diverse TT-specific CD4(+) T cell immune response. PMID- 11282991 TI - Resistance of activated human Th2 cells to NO-induced apoptosis is mediated by gamma-glutamyltranspeptidase. AB - Activation-induced death of inflammatory cells (AICD) has an important function in immune maintenance. Type 1 Th cells are known to be more susceptible to AICD than Th2 cells. In the current study we examined whether NO-induced apoptosis also preferentially eliminates Th1 cells over Th2 cells. Naive human Th lymphocytes (CD4(+)CD45RO(-)) were activated in vitro for 1 week in the presence of IL-12 plus anti-IL-4 or IL-4 plus anti-IL-12 to generate Th1- and Th2 polarized cultures respectively. Cultures were exposed to the NO donors Spermine nonoate (Sper) and DPTA-nonoate to study NO-induced apoptosis. We found that NO preferentially induced apoptosis in Th1-polarized cells as demonstrated by Annexin staining in the presence of 10 microM Sper (70 +/- 16 versus 23 +/- 4.4% in Th2 cells P: < 0.01) and by DioC6 staining (38 +/- 10 versus 11 +/- 5% in Th2 cells, P: < 0.01). The mechanism of NO-induced apoptosis in Th1/Th2-polarized cells was distinct from AICD and Fas-induced apoptosis. Differential sensitivity between Th1- and Th2-polarized cultures originated at the level of intracellular glutathione (GSH) metabolism. GSH levels were higher in Th2 cells (1.6 +/- 0.2 fold Th1, P: < 0.01). High intracellular GSH in Th2-polarized cells did not account for reduced susceptibility to NO per se, since the inhibition of gamma glutamyltrans-peptidase (gamma-GT), which is involved in GSH import, sensitized Th2 cells to NO-induced apoptosis without GSH depletion. Therefore, higher activity of gamma-GT in Th2 cells (2.1 +/- 0.4-fold Th1, P: < 0.001) specifically protects Th2 cells against NO-induced apoptosis. Preferential NO-induced elimination of human Th1 cells at sites of inflammation may thus select Th2 cells and contribute to immune deviation. PMID- 11282992 TI - CTLA-4-Fas ligand functions as a trans signal converter protein in bridging antigen-presenting cells and T cells. AB - Co-stimulator blockade and trans inhibitory signaling, using agents such as CTLA 4-Ig and Fas ligand (FasL) respectively have been invoked as alternative strategies for suppressing pathogenic T cells. This study describes a novel hetero-bifunctional fusion protein, CTLA-4-FasL, designed to combine within a single protein both co-stimulator blocking and trans inhibitory signaling potentials. A chimeric expression cassette, in which the ectodomain coding sequences for CTLA-4 and FasL were linked in-frame, was used to produce a CTLA-4 FasL fusion protein. CTLA-4-FasL binding to both B7-1/B7-2-expressing Daudi B cells and Fas-expressing Jurkat T cells was documented by immunofluorescence and flow cytometry. The capacity of CTLA-4-FasL to induce apoptosis in Jurkat targets was markedly enhanced by the addition of Daudi and other B7-1/B7-2(+) B cell lines, which provided a membrane platform for the otherwise soluble CTLA-4-fusion protein. Moreover, in dual-chamber experiments, Daudi cells pre-coated with CTLA 4-FasL demonstrated Jurkat inhibitory activity that was cell-contact dependent. Significantly, when used to inhibit in vitro cellular proliferation of peripheral blood mononuclear cells, CTLA-4-FasL was approximately 1000-fold more potent than the extensively characterized CTLA-4-Ig fusion protein. Furthermore, the degree of inhibition induced by CTLA-4-FasL substantially surpassed that observed for CTLA-4-Ig and a soluble FasL when used in combination. CTLA-4-FasL represents the first of a novel class of fusion proteins, designated here as 'trans signal converter proteins', that combine trans signal masking and direct trans signaling functions. PMID- 11282993 TI - Evidence for a role of ganglioside GM1 in antigen presentation: binding enhances presentation of Escherichia coli enterotoxin B subunit (EtxB) to CD4(+) T cells. AB - Successful antigen presentation by antigen-presenting cells is governed by a number of factors including the efficiency of antigen capture by cell-surface receptors, targeting to compartments of antigen processing, surface expression of MHC II-peptide complexes and presence of co-stimulatory signals. Ganglioside GM1 is an important component of membrane glycosphingolipids, and has been implicated in cell differentiation, apoptosis and signal transduction pathways. Using the B subunit of Escherichia coli enterotoxin (EtxB), a potent immunogen that binds GM1 with high affinity, and a non-binding mutant of EtxB, EtxB(G33D), we demonstrate that GM1 is intimately involved in several aspects of antigen presentation. Thus, GM1-mediated presentation of EtxB by B cells and CD11c(+) dendritic cells (DC) significantly enhanced the proliferation and cytokine expression of EtxB-specific CD4(+) T cells. Investigation regarding potential mechanisms revealed that EtxB binding directly augments the expression of MHC class II on B cells, and fractionation of B cells demonstrated that EtxB binding to GM1 results in rapid internalization and targeting to class II-rich compartments. GM1-mediated uptake of antigens and access to class II compartments in B cells can be exploited to significantly enhance the presentation of ovalbumin-conjugated to EtxB. These results demonstrate that GM1 can play an important role in antigen presentation via the MHC II pathway. PMID- 11282994 TI - Alterations in peripheral B cells and B cell progenitors following androgen ablation in mice. AB - The production of B lymphocytes is regulated in part by physiologic levels of androgens and estrogens. While these sex hormones down-regulate B lymphopoiesis, augmentation of B lymphopoiesis occurs under conditions where androgen or estrogen levels are decreased. In this study we examine the effect of androgen ablation of male mice on B lymphopoiesis and on the phenotypic composition of peripheral B lymphocyte populations. Spleen and thymic weights are significantly increased following castration, as is the total number of peripheral blood lymphocytes. However, the absolute numbers of B cells in the periphery are selectively increased following castration; the numbers of T cells, NK cells and granulocytes remain unchanged. The increase in circulating B cells is due largely to increases in the numbers of recent bone marrow emigrants expressing a B220(lo+)CD24(hi+) phenotype and these cells remain significantly elevated in castrated mice for up to 54 days post-castration. Similar increases in the percentages of newly emigrated B cells are observed in mice that lack a functional androgen receptor (TFM:). Finally, assessments of B cell progenitors in the bone marrow revealed significant increases in the relative numbers of IL-7 responsive B cell progenitors, including cells in Hardy fractions B (early pro-B cells), C (late pro-B cells), D (pre-B cells) and E (immature B cells). These findings demonstrate that androgen ablation following castration significantly and selectively alters the composition of peripheral B cells in mice. Further, these alterations result from the potentiating effects of androgen ablation on IL 7-responsive pro-B cell progenitors. PMID- 11282995 TI - Decreased expression of signaling lymphocytic-activation molecule-associated protein (SAP) transcripts in T cells from patients with rheumatoid arthritis. AB - The function of Epstein-Barr virus (EBV)-specific cytotoxic T cells is disturbed in rheumatoid arthritis (RA) patients but the mechanism for this disturbance has remained unknown. In a recent study searching for the causative gene of X-linked lymphoproliferative syndrome, the gene possibly linked to EBV-specific cytotoxic T cells or NK cell-mediated cytotoxic activity to EBV-infected cells was discovered, and its product is now referred to as signaling lymphocytic activation molecule-associated protein (SAP) or Src homology 2 domain-containing protein (SH2D1A). In the present study, we attempted to investigate the involvement of the SAP gene in RA using a quantitative real-time PCR; the expression level of SAP transcripts in peripheral leukocytes or T cells was examined for patients with RA. The expression level of SAP transcripts in peripheral leukocytes of 21 RA patients was significantly lower than that of 13 normal individuals (P = 0.0007), four patients with palindromic RA, 11 with inactive systemic lupus erythematosus (SLE) or 17 with chronic renal diseases. The decreased expression of SAP transcripts in RA patients was also observed in peripheral CD2(+) T cells compared with normal individuals. There was no mutation in the coding region of SAP cDNAs derived from peripheral leukocytes of five RA patients. The decreased expression of SAP transcripts in peripheral leukocytes or T cells of RA patients might lead to the failure of the immune system to eliminate the EBV-infected synovial lining cells in joints of RA patients. Our findings have suggested that decreased expression of the SAP gene might be involved in the onset or progress of RA. PMID- 11282996 TI - Complex expression patterns of lymphocyte-specific genes during the development of cartilaginous fish implicate unique lymphoid tissues in generating an immune repertoire. AB - Cartilaginous fish express canonical B and T cell recognition genes, but their lymphoid organs and lymphocyte development have been poorly defined. Here, the expression of Ig, TCR, recombination-activating gene (Rag)-1 and terminal deoxynucleosidase (TdT) genes has been used to identify roles of various lymphoid tissues throughout development in the cartilaginous fish, Raja eglanteria (clearnose skate). In embryogenesis, Ig and TCR genes are sharply up-regulated at 8 weeks of development. At this stage TCR and TdT expression is limited to the thymus; later, TCR gene expression appears in peripheral sites in hatchlings and adults, suggesting that the thymus is a source of T cells as in mammals. B cell gene expression indicates more complex roles for the spleen and two special organs of cartilaginous fish-the Leydig and epigonal (gonad-associated) organs. In the adult, the Leydig organ is the site of the highest IgM and IgX expression. However, the spleen is the first site of IgM expression, while IgX is expressed first in gonad, liver, Leydig and even thymus. Distinctive spatiotemporal patterns of Ig light chain gene expression also are seen. A subset of Ig genes is pre-rearranged in the germline of the cartilaginous fish, making expression possible without rearrangement. To assess whether this allows differential developmental regulation, IgM and IgX heavy chain cDNA sequences from specific tissues and developmental stages have been compared with known germline-joined genomic sequences. Both non-productively rearranged genes and germline-joined genes are transcribed in the embryo and hatchling, but not in the adult. PMID- 11282997 TI - Apoptosis induced by the antigen receptor and Fas in a variant of the immature B cell line WEHI-231 and in splenic immature B cells. AB - Signaling by the BCR causes proliferation and resistance to Fas-induced apoptosis in mature B cells, but growth arrest and apoptosis in immature B cells. We have identified a variant of the immature B cell line WEHI 231 that retains the apoptotic response to the BCR but has acquired susceptibility to Fas-induced apoptosis. The Fas susceptibility was associated with increased Fas expression on the cell surface and down-regulated IgD expression. These cells exhibited a distinctive functional relationship in response to signals from the BCR, Fas and CD40: BCR stimulation markedly promoted Fas-mediated apoptosis (and vice versa) and Fas-induced apoptosis was not subject to modulation by CD40 signaling. While BCR-induced apoptosis was effectively rescued by CD40, it was not affected by the expression of a dominant-negative FADD. The mechanistic distinctions between BCR- and Fas-induced apoptosis were further characterized by the differential effects of different caspase inhibitors on these two processes which imply the involvement of different subsets of caspases. For BCR-induced apoptosis, we provide evidence that the final apoptotic destruction phase can be inhibited by the pan-caspase inhibitor BOC-Asp-FMK (BD) and that, in the presence of BD, the BCR only induces growth arrest which is reversible. The striking enhancing effects of Fas on BCR-induced apoptosis seen in the variant cells prompted us to examine if a similar cooperation in induction of apoptosis occurs in the highly tolerizable immature B cells of the spleen. We found that the splenic immature B population contains a significant number of Fas-expressing cells, but neither Fas induced apoptosis nor an enhancing effect of Fas on BCR-induced apoptosis of these cells was detected in vitro. PMID- 11282998 TI - CD11b expression as a marker to distinguish between recently activated effector CD8(+) T cells and memory cells. AB - CD8(+) T cells in different activation states have been difficult to identify phenotypically. In this study we have investigated whether Mac-1 (CD11b) expression can be used as a criterion to distinguish between recently activated effector cells and memory cells belonging to the CD8(+) T cell subset. Polyclonal virus-specific effector and memory CD8(+) T cells from lymphocytic choriomeningitis- and vesicular stomatitis virus-infected mice were visualized through staining for intracellular IFN-gamma or binding of MHC-peptide tetramers, and Mac-1 expression was evaluated. Naive T cells and most virus-specific memory CD8(+) T cells express little or no Mac-1 independent of the virus model employed. In contrast, the majority of CD8(+) T cells present during acute infection express a significant level of Mac-1 and, similarly, Mac-1 expression is found on secondary effectors generated in response to viral re-exposure. We therefore suggest that high Mac-1 expression defines a subset of circulating effector cells and that the presence of this marker on antigen-specific CD8(+) T cells signifies recent activation. PMID- 11282999 TI - Subcellular distribution and cytokine- and chemokine-regulated secretion of leukolysin/MT6-MMP/MMP-25 in neutrophils. AB - Leukolysin, originally isolated from human leukocytes, is the sixth member of the membrane-type matrix metalloproteinase (MT-MMP) subfamily with a potential glycosylphosphatidylinositol (GPI) anchor. To understand its biological functions, we screened subpopulations of leukocytes and localized the expression of leukolysin at the mRNA level to neutrophils. Polyclonal and mono-specific antisera raised against a synthetic peptide from its hinge region recognized a major protein species at 56 kDa and several minor forms between 38 and 45 kDa in neutrophil lysates. In resting neutrophils, leukolysin is distributed among specific granules ( approximately 10%), gelatinase granules ( approximately 40%), secretory vesicles ( approximately 30%), and the plasma membrane ( approximately 20%), a pattern distinct from that of neutrophil MMP-8 and MMP-9. Consistent with its membrane localization and its reported GPI anchor, leukolysin partitions into the detergent phase of Triton X-114 and can be released from intact resting neutrophils by glycosylphosphatidylinositol-specific phospholipase C. Phorbol myristate acetate stimulates neutrophils to discharge 100% of leukolysin from specific and gelatinase granules and approximately 50% from the secretory vesicles and plasma membrane, suggesting that leukolysin can be mobilized by physiological signals to the extracellular milieu as a soluble enzyme. Indeed, interleukin 8, a neutrophil chemoattractant, triggered a release of approximately 85% of cellular leukolysins by a process resistant to a mixture of proteinase inhibitors, including aprotinin, BB-94, pepstatin, and E64. Finally, purified recombinant leukolysin can degrade components of the extracellular matrix. These results not only establish leukolysin as the first neutrophil-specific MT-MMP but also implicate it as a cytokine/chemokine-regulated effector during innate immune responses or tissue injury. PMID- 11283000 TI - Biochemical identification of Xenopus Pumilio as a sequence-specific cyclin B1 mRNA-binding protein that physically interacts with a Nanos homolog, Xcat-2, and a cytoplasmic polyadenylation element-binding protein. AB - Translational activation of dormant cyclin B1 mRNA stored in oocytes is a prerequisite for the initiation or promotion of oocyte maturation in many vertebrates. Using a monoclonal antibody against the domain highly homologous to that of Drosophila Pumilio, we have shown for the first time in any vertebrate that a homolog of Pumilio is expressed in Xenopus oocytes. This 137-kDa protein binds to the region including the sequence UGUA at nucleotides 1335-1338 in the 3'-untranslated region of cyclin B1 mRNA, which is close to but does not overlap the cytoplasmic polyadenylation elements (CPEs). Physical in vitro association of Xenopus Pumilio with a Xenopus homolog of Nanos (Xcat-2) was demonstrated by a protein pull-down assay. The results of immunoprecipitation experiments showed in vivo interaction between Xenopus Pumilio and CPE-binding protein (CPEB), a key regulator of translational repression and activation of mRNAs stored in oocytes. This evidence provides a new insight into the mechanism of translational regulation through the 3'-end of mRNA during oocyte maturation. These results also suggest the generality of the function of Pumilio as a translational regulator of dormant mRNAs in both invertebrates and vertebrates. PMID- 11283001 TI - Functional characterization of arginine 30, lysine 40, and arginine 62 in Tn5 transposase. AB - Three N-terminal basic residues of Tn5 transposase, which are associated with proteolytic cleavages by Escherichia coli proteinases, were mutated to glutamine residues with the goal of producing more stable transposase molecules. Mutation of either arginine 30 or arginine 62 to glutamine produced transposase molecules that were more stable toward E. coli proteinases than the parent hyperactive Tn5 transposase, however, they were inactive in vivo. In vitro analysis revealed these mutants were inactive, because both Arg(30) and Arg(62) are required for formation of the paired ends complexes when the transposon is attached to the donor backbone. These results suggest Arg(30) and Arg(62) play critical roles in DNA binding and/or synaptic complex formation. Mutation of lysine 40 to glutamine did not increase the overall stability of the transposase to E. coli proteinases. This mutant transposase was only about 1% as active as the parent hyperactive transposase in vivo; however, it retained nearly full activity in vitro. These results suggest that lysine 40 is important for a step in the transposition mechanism that is bypassed in the in vitro assay system, such as the removal of the transposase molecule from DNA following strand transfer. PMID- 11283002 TI - Identification of amino acid residues in bone morphogenetic protein-1 important for procollagen C-proteinase activity. AB - Bone morphogenetic protein (BMP)-1, which belongs to the tolloid subgroup of astacin-like zinc metalloproteinases, cleaves the C-propeptides of procollagen at the physiologic site and is, therefore, a procollagen C-proteinase (PCP). Cleavage occurs between a specific alanine or glycine residue (depending on the procollagen chain) and an invariant aspartic acid residue in each of the three chains of procollagen. To learn more about how BMP-1 exhibits PCP activity we mapped the primary structure of BMP-1 onto the x-ray crystal structure of astacin and identified residues in the metalloproteinase domain of BMP-1 for subsequent site-directed mutagenesis studies. Recombinant wild-type and mutant BMP-1 were expressed in COS-7 cells and assayed for PCP activity using type I procollagen as the substrate. We showed that substitution of alanine for Glu(94), which occurs in the HEXXH zinc-binding motif of BMP-1, abolishes PCP activity. Furthermore, mutation of residues Lys(87) and Lys(176), which are located in the S1' pocket of the enzyme and are therefore adjacent to the P1' residue in the substrate, reduced the proteolytic activity of BMP-1 by approximately 50%. A surprising observation was that mutation of Cys(66) reduced the activity to 20%, suggesting that this residue is crucial for activity. Further experiments showed that Cys(66) and Cys(63), which are located in the tolloid-specific sequence Cys(63) Gly(64)-Cys(65)-Cys(66) in the active site, most likely form a disulfide bridge. PMID- 11283003 TI - Residues within the N-terminal domain of human topoisomerase I play a direct role in relaxation. AB - All eukaryotic forms of DNA topoisomerase I contain an extensive and highly charged N-terminal domain. This domain contains several nuclear localization sequences and is essential for in vivo function of the enzyme. However, so far no direct function of the N-terminal domain in the in vitro topoisomerase I reaction has been reported. In this study we have compared the in vitro activities of a truncated form of human topoisomerase I lacking amino acids 1-206 (p67) with the full-length enzyme (p91). Using these enzyme forms, we have identified for the first time a direct role of residues within the N-terminal domain in modulating topoisomerase I catalysis, as revealed by significant differences between p67 and p91 in DNA binding, cleavage, strand rotation, and ligation. A comparison with previously published studies showing no effect of deleting the first 174 or 190 amino acids of topoisomerase I (Stewart, L., Ireton, G. C., and Champoux, J. J. (1999) J. Biol. Chem. 274, 32950-32960; Bronstein, I. B., Wynne-Jones, A., Sukhanova, A., Fleury, F., Ianoul, A., Holden, J. A., Alix, A. J., Dodson, G. G., Jardillier, J. C., Nabiev, I., and Wilkinson, A. J. (1999) Anticancer Res. 19, 317-327) suggests a pivotal role of amino acids 191-206 in catalysis. Taken together the presented data indicate that at least part(s) of the N-terminal domain regulate(s) enzyme/DNA dynamics during relaxation most probably by controlling non-covalent DNA binding downstream of the cleavage site either directly or by coordinating DNA contacts by other parts of the enzyme. PMID- 11283004 TI - A single amino acid substitution within a coiled-coil motif changes the assembly of a 53-amino acid protein from two-dimensional sheets to filamentous structures. AB - The bacteriophage phi29 replication protein p1 self-interacts in vitro, generating highly ordered structures. Specifically, the 53-amino acid protein p1DeltaN33, which retains the sequence of p1 spanning amino acids Met(34) to Lys(85), assembles into two-dimensional protofilament sheets. The region of protein p1 located between residues Glu(38) and Asn(65) presumably forms an alpha helical coiled-coil structure. Here we have examined the role of this coiled-coil sequence in the formation of protofilament sheets. Using sedimentation assays and negative-stain electron microscopy analysis, we demonstrate that residues Leu(46), Met(53), and Leu(60), but not Leu(39), are essential for p1DeltaN33 assembly into sheets. Remarkably, replacement of Leu(46) by Val shifts the pathway of molecular assembly, leading to the formation of filamentous polymers approximately 10 nm in diameter. These results show, for the first time, that a short coiled-coil motif can mediate protein assembly into protofilament sheet structures. PMID- 11283005 TI - Interaction of cytochrome bd with carbon monoxide at low and room temperatures: evidence that only a small fraction of heme b595 reacts with CO. AB - Azotobacter vinelandii is an obligately aerobic bacterium in which aerotolerant dinitrogen fixation requires cytochrome bd. This oxidase comprises two polypeptide subunits and three hemes, but no copper, and has been studied extensively. However, there remain apparently conflicting reports on the reactivity of the high spin heme b(595) with ligands. Using purified cytochrome bd, we show that absorption changes induced by CO photodissociation from the fully reduced cytochrome bd at low temperatures demonstrate binding of the ligand with heme b(595). However, the magnitude of these changes corresponds to the reaction with CO of only about 5% of the heme. CO binding with a minor fraction of heme b(595) is also revealed at room temperature by time-resolved studies of CO recombination. The data resolve the apparent discrepancies between conclusions drawn from room and low temperature spectroscopic studies of the CO reaction with cytochrome bd. The results are consistent with the proposal that hemes b(595) and d form a diheme oxygen-reducing center with a binding capacity for a single exogenous ligand molecule that partitions between the hemes d and b(595) in accordance with their intrinsic affinities for the ligand. In this model, the affinity of heme b(595) for CO is about 20-fold lower than that of heme d. PMID- 11283006 TI - The mechanism of DNA cytosine-5 methylation. Kinetic and mutational dissection of Hhai methyltransferase. AB - Kinetic and binding studies involving a model DNA cytosine-5-methyltransferase, M.HhaI, and a 37-mer DNA duplex containing a single hemimethylated target site were applied to characterize intermediates on the reaction pathway. Stopped-flow fluorescence studies reveal that cofactor S-adenosyl-l-methionine (AdoMet) and product S-adenosyl-l-homocysteine (AdoHcy) form similar rapidly reversible binary complexes with the enzyme in solution. The M.HhaI.AdoMet complex (k(off) = 22 s( )1, K(D) = 6 microm) is partially converted into products during isotope partitioning experiments, suggesting that it is catalytically competent. Chemical formation of the product M.HhaI.(Me)DNA.AdoHcy (k(chem) = 0.26 s(-)1) is followed by a slower decay step (k(off) = 0.045 s(-)1), which is the rate-limiting step in the catalytic cycle (k(cat) = 0.04 s(-)1). Analysis of reaction products shows that the hemimethylated substrate undergoes complete (>95%) conversion into fully methylated product during the initial burst phase, indicating that M.HhaI exerts high binding selectivity toward the target strand. The T250N, T250D, and T250H mutations, which introduce moderate perturbation in the catalytic site, lead to substantially increased K(D)(DNA(ternary)), k(off)(DNA(ternary)), K(M)(AdoMet(ternary)) values but small changes in K(D)(DNA(binary)), K(D)(AdoMet(binary)), k(chem), and k(cat). When the target cytosine is replaced with 5-fluorocytosine, the chemistry step leading to an irreversible covalent M.HhaI.DNA complex is inhibited 400-fold (k(chem)(5FC) = 0.7 x 10(-)3 s(-)1), and the Thr-250 mutations confer further dramatic decrease of the rate of the covalent methylation k(chem). We suggest that activation of the pyrimidine ring via covalent addition at C-6 is a major contributor to the rate of the chemistry step (k(chem)) in the case of cytosine but not 5-fluorocytosine. In contrast to previous reports, our results imply a random substrate binding order mechanism for M.HhaI. PMID- 11283007 TI - Functional domains of the yeast splicing factor Prp22p. AB - The essential Saccharomyces cerevisiae PRP22 gene encodes a 1145-amino acid DEXH box RNA helicase. Prp22p plays two roles during pre-mRNA splicing as follows: it is required for the second transesterification step and for the release of mature mRNA from the spliceosome. Whereas the step 2 function of Prp22p does not require ATP hydrolysis, spliceosome disassembly is dependent on the ATPase and helicase activities. Here we delineate a minimal functional domain, Prp22(262-1145), that suffices for the activity of Prp22p in vivo when expressed under the natural PRP22 promoter and for pre-mRNA splicing activity in vitro. The biologically active domain lacks an S1 motif (residues 177-256) that had been proposed to play a role in RNA binding by Prp22p. The deletion mutant Prp22(351-1145) can function in vivo when provided at a high gene dosage. We suggest that the segment from residues 262 to 350 enhances Prp22p function in vivo, presumably by targeting Prp22p to the spliceosome. We characterize an even smaller catalytic domain, Prp22(466-1145) that suffices for ATP hydrolysis, RNA binding, and RNA unwinding in vitro and for nuclear localization in vivo but cannot by itself support cell growth. However, the ATPase/helicase domain can function in vivo if the N terminal region Prp22(1-480) is co-expressed in trans. PMID- 11283008 TI - Glutathione oxidation by hypochlorous acid in endothelial cells produces glutathione sulfonamide as a major product but not glutathione disulfide. AB - Treatment of cells with hypochlorous acid (HOCl) at sublethal doses causes a concentration-dependent loss in reduced glutathione (GSH) levels. We have investigated the products of the reaction of HOCl with GSH in human umbilical vein endothelial cells. Despite a complete loss of GSH, there were only very small increases in intracellular and extracellular glutathione disulfide and glutathione sulfonic acid after exposure to HOCl. (35)S labeling of the GSH pool showed only a minimal increase in protein-bound GSH, suggesting that S-thiolation was not a major contributor to HOCl-mediated loss of GSH in endothelial cells. Rather, the products of the reaction were mostly exported from cells and included a peak that co-eluted with the cyclic sulfonamide that is a product of the reaction of GSH with reagent HOCl. Evidence of this species in endothelial cell supernatants after HOCl treatment was also obtained using electrospray mass spectrometry. In conclusion, exposure to HOCl causes the irreversible loss of cellular GSH with the formation of novel products that are rapidly exported from the cell, and resynthesis of GSH will be required to restore levels. The loss of GSH would alter the redox state of the cell and compromise its defenses against further oxidative stress. PMID- 11283009 TI - The nuclear pore complex as a transport machine. PMID- 11283010 TI - Autoregulation of cell-specific MAP kinase control of the tryptophan hydroxylase promoter. AB - The neurotransmitter serotonin controls a wide range of biological systems, including its own synthesis and release. As the rate-limiting enzyme in serotonin biosynthesis, tryptophan hydroxylase (TPH) is a potential target for this autoregulation. Using the serotonergic neuron-like CA77 cell line, we have demonstrated that treatment with a 5-hydroxytryptamine autoreceptor agonist, CGS 12066A, can lower TPH mRNA levels and promoter activity. We reasoned that this repression might involve inhibition of MAP kinases, since 5-HT1 receptors can increase mitogen-activated protein (MAP) kinase phosphatase levels. To test this hypothesis, we first showed that the TPH promoter can be activated 20-fold by mitogen-activated extracellular-signal regulated kinase kinase kinase (MEKK), an activator of MAP kinases. This activation was then blocked by CGS 12066A. The maximal MAP kinase and CGS repression regulatory region was mapped to between 149 and -45 base pairs upstream of the transcription start site. The activation by MEKK appears to be cell-specific, because MEKK did not activate the TPH promoter in nonneuronal cell lines. At least part, but not all, of the MAP kinase responsiveness was mapped to an inverted CCAAT box that binds the transcription factor NF-Y. These data suggest a model for the autoregulation of serotonin biosynthesis by repression of MAP kinase stimulation of the TPH promoter. PMID- 11283011 TI - Subcellular compartment and molecular subdomain of beta-amyloid precursor protein relevant to the Abeta 42-promoting effects of Alzheimer mutant presenilin 2. AB - Increased production of amyloid beta peptides ending at position 42 (Abeta42) is one of the pathogenic phenotypes caused by mutant forms of presenilins (PS) linked to familial Alzheimer's disease. To identify the subcellular compartment(s) in which familial Alzheimer's disease mutant PS2 (mt PS2) affects the gamma-cleavage of betaAPP to increase Abeta42, we co-expressed the C-terminal 99-amino acid fragment of betaAPP (C100) tagged with sorting signals to the endoplasmic reticulum (C100/ER) or to the trans-Golgi network (C100/TGN) together with mt PS2 in N2a cells. C100/TGN co-transfected with mt PS2 increased levels or ratios of intracellular as well as secreted Abeta42 at similar levels to those with C100 without signals (C100/WT), whereas C100/ER yielded a negligible level of Abeta, which was not affected by co-transfection of mt PS2. To identify the molecular subdomain of betaAPP required for the effects of mt PS2, we next co expressed C100 variously truncated at the C-terminal cytoplasmic domain together with mt PS2. All types of C-terminally truncated C100 variants including that lacking the entire cytoplasmic domain yielded the secreted form of Abeta at levels comparable with those from C100/WT, and co-transfection of mt PS2 increased the secretion of Abeta42. These results suggest that (i) late intracellular compartments including TGN are the major sites in which Abeta42 is produced and up-regulated by mt PS2 and that (ii) the anterior half of C100 lacking the entire cytoplasmic domain is sufficient for the overproduction of Abeta42 caused by mt PS2. PMID- 11283012 TI - Unique pathway of thrombin-induced platelet aggregation mediated by glycoprotein Ib. AB - Thrombin plays a central role in normal and abnormal hemostatic processes. It is assumed that alpha-thrombin activates platelets by hydrolyzing the protease activated receptor (PAR)-1, thereby exposing a new N-terminal sequence, a tethered ligand, which initiates a cascade of molecular reactions leading to thrombus formation. This process involves cross-linking of adjacent platelets mediated by the interaction of activated glycoprotein (GP) IIb/IIIa with distinct amino acid sequences, LGGAKQAGDV and/or RGD, at each end of dimeric fibrinogen molecules. We demonstrate here the existence of a second alpha-thrombin-induced platelet-activating pathway, dependent on GP Ib, which does not require hydrolysis of a substrate receptor, utilizes polymerizing fibrin instead of fibrinogen, and can be inhibited by the Fab fragment of the monoclonal antibody LJIb-10 bound to the GP Ib thrombin-binding site or by the cobra venom metalloproteinase, mocarhagin, that hydrolyzes the extracellular portion of GP Ib. This alternative alpha-thrombin pathway is observed when PAR-1 or GP IIb/IIIa is inhibited. The recognition sites involved in the cross-linking of polymerizing fibrin and surface integrins via the GP Ib pathway are different from those associated with fibrinogen. This pathway is insensitive to RGDS and anti-GP IIb/IIIa antibodies but reactive with a mutant fibrinogen, gamma407, with a deletion of the gamma-chain sequence, AGDV. The reaction is not due to simple trapping of platelets by the fibrin clot, since ligand binding, signal transduction, and second messenger formation are required. The GP Ib pathway is accompanied by mobilization of internal calcium and the platelet release reaction. This latter aspect is not observed with ristocetin-induced GP Ib-von Willebrand factor agglutination nor with GP Ib-von Willebrand factor-polymerizing fibrin trapping of platelets. Human platelets also respond to gamma-thrombin, an autoproteolytic product of alpha-thrombin, through PAR-4. Co-activation of the GP Ib, PAR-1, and PAR-4 pathways elicit synergistic responses. The presence of the GP Ib pathway may explain why anti-alpha-thrombin/anti-platelet regimens fail to completely abrogate thrombosis/restenosis in the cardiac patient. PMID- 11283013 TI - Antizyme regulates the degradation of ornithine decarboxylase in fission yeast Schizosaccharomyces pombe. Study in the spe2 knockout strains. AB - The mechanism of the regulatory degradation of ornithine decarboxylase (ODC) by polyamines was studied in fission yeast, Schizosaccharomyces pombe. To regulate cellular spermidine experimentally, we cloned and disrupted S-adenosylmethionine decarboxylase gene (spe2) in S. pombe. The null mutant of spe2 was devoid of spermidine and spermine, accumulated putrescine, and contained a high level of ODC. Addition of spermidine to the culture medium resulted in rapid decrease in the ODC activity caused by the acceleration of ODC degradation, which was dependent on de novo protein synthesis. A fraction of ODC forming an inactive complex concomitantly increased. The accelerated ODC degradation was prevented either by knockout of antizyme gene or by selective inhibitors of proteasome. Thus, unlike budding yeast, mammalian type antizyme-mediated ODC degradation by proteasome is operating in S. pombe. PMID- 11283014 TI - A role for Wiskott-Aldrich syndrome protein in T-cell receptor-mediated transcriptional activation independent of actin polymerization. AB - Wiskott-Aldrich syndrome protein (WASP) plays a key role in cytoskeletal rearrangement and transcriptional activation in T-cells. Recent evidence links WASP and related proteins to actin polymerization by the Arp2/3 complex. To study whether the role of WASP in actin polymerization is coupled to T-cell receptor (TCR)-mediated transcriptional activation, we made a series of WASP deletion mutants and tested them for actin co-localization, actin polymerization, and transcriptional activation of NFAT. A WASP mutant with a deletion in the C terminal region (WASPDeltaC) that is defective in actin polymerization potentiated NFAT transcription following TCR activation by anti-CD3 and anti CD3/CD28 antibodies, but not by phorbol 12-myristate 13-acetate/ionomycin. Furthermore, cotransfection of a dominant-active mutant (WASP-WH2-C) for Arp2/3 polymerization did not inhibit NFAT activation. Finally, by analyzing a series of WASP double-domain deletion mutants, we determined that the WASP homology-1 domain is responsible for NFAT transcriptional activation. Our results suggest that WASP activates transcription following TCR stimulation in a manner that is independent of its role in Arp2/3-directed actin polymerization. PMID- 11283015 TI - Cell cycle regulation by galectin-12, a new member of the galectin superfamily. AB - Galectins are a family of beta-galactoside-binding animal lectins with conserved carbohydrate recognition domains (CRDs). Here we report the identification and characterization of a new galectin, galectin-12, which contains two domains that are homologous to the galectin CRD. The N-terminal domain contains all of the sequence elements predicted to form the two beta-sheets found in other galectins, as well as conserved carbohydrate-interacting residues. The C-terminal domain shows considerable divergence from the consensus sequence, and many of these conserved residues are not present. Nevertheless, the protein has lactose binding activity, most likely due to the contribution of the N-terminal domain. The mRNA for galectin-12 contains features coding for proteins with growth-regulatory functions. These include start codons in a context that are suboptimal for translation initiation and AU-rich motifs in the 3'-untranslated region, which are known to confer instability to mRNA. Galectin-12 mRNA is sparingly expressed or undetectable in many tissues and cell lines tested, but it is up-regulated in cells synchronized at the G(1) phase or the G(1)/S boundary of the cell cycle. Ectopic expression of galectin-12 in cancer cells causes cell cycle arrest at the G(1) phase and cell growth suppression. We conclude that galectin-12 is a novel regulator of cellular homeostasis. PMID- 11283016 TI - A novel human striated muscle RING zinc finger protein, SMRZ, interacts with SMT3b via its RING domain. AB - The RING domain is a conserved zinc finger motif, which serves as a protein protein interaction interface. Searches of a human heart expressed sequence tag data base for genes encoding the RING domain identified a novel cDNA, named striated muscle RING zinc finger protein (SMRZ). The SMRZ cDNA is 1.9 kilobase pairs in length and encodes a polypeptide of 288 amino acid residues; analysis of the peptide sequence demonstrated an N-terminal RING domain. Fluorescence in situ hybridization localized SMRZ to chromosome 1p33-34. Northern blots demonstrated that SMRZ is expressed exclusively in striated muscle. In the cardiovascular system, SMRZ is more highly expressed in the fetal heart than in the adult heart (slightly higher expression in the ventricle than in the atrium), suggesting that SMRZ is developmentally regulated. SMRZ was found to interact with SMT3b, a ubiquitin-like protein, through the SMRZ-RING domain. This interaction was abolished by mutagenesis of conserved RING domain residues. Transient transfection of SMRZ into C2C12 myoblasts showed localization of SMRZ to the nucleus. These data suggest that SMRZ may play an important role in striated muscle cell embryonic development and perhaps in cell cycle regulation. PMID- 11283017 TI - Molecular cloning and characterization of UDP-GlcNAc:lactosylceramide beta 1,3-N acetylglucosaminyltransferase (beta 3Gn-T5), an essential enzyme for the expression of HNK-1 and Lewis X epitopes on glycolipids. AB - A new member of the UDP-N-acetylglucosamine:beta-galactose beta1,3-N acetylglucosaminyltransferase (beta3Gn-T) family having the beta3Gn-T motifs was cloned from rat and human cDNA libraries and named beta3Gn-T5 based on its position in a phylogenetic tree. We concluded that beta3Gn-T5 is the most feasible candidate for lactotriaosylceramide (Lc(3)Cer) synthase, an important enzyme which plays a key role in the synthesis of lacto- or neolacto-series carbohydrate chains on glycolipids. beta3Gn-T5 exhibited strong activity to transfer GlcNAc to glycolipid substrates, such as lactosylceramide (LacCer) and neolactotetraosylceramide (nLc(4)Cer; paragloboside), resulting in the synthesis of Lc(3)Cer and neolactopentaosylceramide (nLc(5)Cer), respectively. A marked decrease in LacCer and increase in nLc(4)Cer was detected in Namalwa cells stably expressing beta3Gn-T5. This indicated that beta3Gn-T5 exerted activity to synthesize Lc(3)Cer and decrease LacCer, followed by conversion to nLc(4)Cer via endogenous galactosylation. The following four findings further supported that beta3Gn-T5 is Lc(3)Cer synthase. 1) The beta3Gn-T5 transcript levels in various cells were consistent with the activity levels of Lc(3)Cer synthase in those cells. 2) The beta3Gn-T5 transcript was presented in various tissues and cultured cells. 3) The beta3Gn-T5 expression was up-regulated by stimulation with retinoic acid and down-regulated with 12-O-tetradecanoylphorbol-13-acetate in HL-60 cells. 4) The changes in beta3Gn-T5 transcript levels during the rat brain development were determined. Points 2, 3, and 4 were consistent with the Lc(3)Cer synthase activity reported previously. PMID- 11283018 TI - In vitro selection and characterization of Bcl-X(L)-binding proteins from a mix of tissue-specific mRNA display libraries. AB - The covalent coupling of an mRNA to the protein that it encodes (mRNA display) provides a powerful tool for analysis of protein function in the post-genomic era. This coupling allows the selective enrichment of individual members from libraries of displayed proteins and the subsequent regeneration of an enriched library using the RNA moiety. Tissue-specific libraries from poly(A)(+) mRNA were prepared by priming first and second strand cDNA synthesis with oligonucleotides containing nine random 3' nucleotides, the fixed regions of which encoded the requisite sequences for formation of mRNA display constructs and a library specific sequence tag. Starting with a pool of uniquely tagged libraries from different tissues, an iterative selection was performed for binding partners of the anti-apoptotic protein Bcl-X(L). After four rounds of selection, the pool was deconvoluted by polymerase chain reaction amplification with library-specific primers. Subsequent clonal sequence analysis revealed the selection of three members of the Bcl-2 family known to bind to Bcl-X(L). In addition, several proteins not previously demonstrated to interact with Bcl-X(L) were identified. The relative binding affinities of individual selected peptides were determined, as was their susceptibility to competition with a BH3 domain peptide. Based on these data, a putative BH3 domain was identified in most peptides. PMID- 11283019 TI - Intestinal epithelial cell differentiation involves activation of p38 mitogen activated protein kinase that regulates the homeobox transcription factor CDX2. AB - The intracellular signaling pathways responsible for cell cycle arrest and differentiation along the crypt-villus axis of the human small intestine remain largely unknown. p38 mitogen-activated protein kinases (MAPKs) have recently emerged as key modulators of various vertebrate cell differentiation processes. In order to elucidate further the mechanism(s) responsible for the loss of proliferative potential once committed intestinal cells begin to differentiate, the role and regulation of p38 MAPK with regard to differentiation were analyzed in both intact epithelium as well as in well established intestinal cell models recapitulating the crypt-villus axis in vitro. Results show that phosphorylated and active forms of p38 were detected primarily in the nuclei of differentiated villus cells. Inhibition of p38 MAPK signaling by 2-20 microm SB203580 did not affect E2F-dependent transcriptional activity in subconfluent Caco-2/15 or HIEC cells. p38 MAPK activity dramatically increased as soon as Caco-2/15 cells reached confluence, whereas addition of SB203580 during differentiation of Caco 2/15 cells strongly attenuated sucrase-isomaltase gene and protein expression as well as protein expression of villin and alkaline phosphatase. The binding of CDX2 to the sucrase-isomaltase promoter and its transcriptional activity were significantly reduced by SB203580. Pull-down glutathione S-transferase and immunoprecipitation experiments demonstrated a direct interaction of CDX3 with p38. Finally, p38-dependent phosphorylation of CDX3 was observed in differentiating Caco-2/15 cells. Taken together, our results indicate that p38 MAPK may be involved in the regulation of CDX2/3 function and intestinal cell differentiation. PMID- 11283020 TI - Modulation of mitochondrial function by hydrogen peroxide. AB - During normal cellular metabolism, mitochondrial electron transport results in the formation of superoxide anion (O(2)) and subsequently hydrogen peroxide (H(2)O(2)). Because H(2)O(2) increases in concentration under certain physiologic and pathophysiologic conditions and can oxidatively modify cellular components, it is critical to understand the response of mitochondria to H(2)O(2). In the present study, treatment of isolated rat heart mitochondria with H(2)O(2) resulted in a decline and subsequent recovery of state 3 NADH-linked respiration. Alterations in NADH levels induced by H(2)O(2) closely paralleled changes in the rate of state 3 respiration. Assessment of electron transport chain complexes and Krebs cycle enzymes revealed that alpha-ketoglutarate dehydrogenase (KGDH), succinate dehydrogenase (SDH), and aconitase were susceptible to H(2)O(2) inactivation. Of particular importance, KGDH and SDH activity returned to control levels, concurrent with the recovery of state 3 respiration. Inactivation is not because of direct interaction of H(2)O(2) with KGDH and SDH. In addition, removal of H(2)O(2) alone is not sufficient for reactivation. Enzyme activity does not recover unless mitochondria remain intact. The sensitivity of KGDH and SDH to H(2)O(2)-mediated inactivation and the reversible nature of inactivation suggest a potential role for H(2)O(2) in the regulation of KGDH and SDH. PMID- 11283021 TI - Rac1 activity is required for the activation of hypoxia-inducible factor 1. AB - Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that mediates cellular and systemic homeostatic responses (including erythropoiesis, angiogenesis, and glycolysis) to reduced O(2) availability in mammals. Hypoxia induces both the protein expression and transcriptional activity of the HIF 1alpha subunit. However, the molecular mechanisms of sensing and signal transduction by which changes in O(2) concentration result in changes in HIF-1 activity are poorly understood. We report here that the small GTPase Rac1 is activated in response to hypoxia and is required for the induction of HIF-1alpha protein expression and transcriptional activity in hypoxic cells. PMID- 11283022 TI - Insulin regulates alternative splicing of protein kinase C beta II through a phosphatidylinositol 3-kinase-dependent pathway involving the nuclear serine/arginine-rich splicing factor, SRp40, in skeletal muscle cells. AB - Insulin regulates the inclusion of the exon encoding protein kinase C (PKC) betaII mRNA. In this report, we show that insulin regulates this exon inclusion (alternative splicing) via the phosphatidylinositol 3-kinase (PI 3-kinase) signaling pathway through the phosphorylation state of SRp40, a factor required for insulin-regulated splice site selection for PKCbetaII mRNA. By taking advantage of a well known inhibitor of PI 3-kinase, LY294002, we demonstrated that pretreatment of L6 myotubes with LY294002 blocked insulin-induced PKCbetaII exon inclusion as well as phosphorylation of SRp40. In the absence of LY294002, overexpression of SRp40 in L6 cells mimicked insulin-induced exon inclusion. When antisense oligonucleotides targeted to a putative SRp40-binding sequence in the betaII-betaI intron were transfected into L6 cells, insulin effects on splicing and glucose uptake were blocked. Taken together, these results demonstrate a role for SRp40 in insulin-mediated alternative splicing independent of changes in SRp40 concentration but dependent on serine phosphorylation of SRp40 via a PI 3 kinase signaling pathway. This switch in PKC isozyme expression is important for increases in the glucose transport effect of insulin. Significantly, insulin regulation of PKCbetaII exon inclusion occurred in the absence of cell growth and differentiation demonstrating that insulin-induced alternative splicing of PKCbetaII mRNA in L6 cells occurs in response to a metabolic change. PMID- 11283024 TI - Endogenous formation of protein adducts with carcinogenic aldehydes: implications for oxidative stress. AB - In the present study, we characterize the covalent modification of a protein by crotonaldehyde, a representative carcinogenic aldehyde, and describe the endogenous production of this aldehyde in vivo. The crotonaldehyde preferentially reacted with the lysine and histidine residues of bovine serum albumin and generated a protein-linked carbonyl derivative. Upon incubation with the histidine and lysine derivatives, crotonaldehyde predominantly generated beta substituted butanal adducts of histidine and lysine and N(epsilon)-(2,5-dimethyl 3-formyl-3,4-dehydropiperidino)lysine (dimethyl-FDP-lysine) as the putative carbonyl derivatives generated in the crotonaldehyde-modified protein. To verify the endogenous formation of crotonaldehyde in vivo, we raised the monoclonal antibody (mAb82D3) against the crotonaldehyde-modified protein and found that it cross-reacted with the protein-bound 2-alkenals, such as crotonaldehyde, 2 pentenal, and 2-hexenal. The anti-2-alkenal antibody recognized multiple crotonaldehyde-lysine adducts, including dimethyl-FDP-lysine and an unknown product, which showed the greatest immunoreactivity with the antibody. On the basis of the chemical and spectroscopic evidence, the major antigenic product was determined to be a novel Schiff base-derived crotonaldehyde-lysine adduct, N(epsilon)-(5-ethyl-2-methylpyridinium)lysine (EMP-lysine). It was found that the lysine residues that had disappeared in the protein treated with crotonaldehyde were partially recovered by EMP-lysine. The presence of immunoreactive materials with mAb82D3 in vivo was demonstrated in the kidney of rats exposed to the renal carcinogen, ferric nitrilotriacetate. In addition, the observations that the metal-catalyzed oxidation of polyunsaturated fatty acids in the presence of proteins resulted in an increase in the antigenicity of the protein indicated that lipid peroxidation represents a potential pathway for the formation of crotonaldehyde/2-alkenals in vivo. These data suggest that the formation of carcinogenic aldehydes during lipid peroxidation may be causally involved in the pathophysiological effects associated with oxidative stress. PMID- 11283023 TI - The neural recognition molecule L1 is a sialic acid-binding lectin for CD24, which induces promotion and inhibition of neurite outgrowth. AB - Among the recognition molecules that determine a neuron's interaction with other cells, L1 and CD24 have been suggested to cooperate with each other in neurite outgrowth and signal transduction. Here we report that binding of CD24 to L1 depends on alpha2,3-sialic acid on CD24, which determines the CD24 induced and cell type-specific promotion or inhibition of neurite outgrowth. Using knockout mutants, we could show that the CD24-induced effects on neurite outgrowth are mediated via L1, and not GPI-linked CD24, by trans-interaction of L1 with sialylated CD24. This glycoform is excluded together with L1 from raft microdomains, suggesting that molecular compartmentation in the surface membrane could play a role in signal transduction. PMID- 11283025 TI - Directed inhibition of nuclear import in cellular hypertrophy. AB - Each nuclear pore is responsible for both nuclear import and export with a finite capacity for bidirectional transport across the nuclear envelope. It remains poorly understood how the nuclear transport pathway responds to increased demands for nucleocytoplasmic communication. A case in point is cellular hypertrophy in which increased amounts of genetic material need to be transported from the nucleus to the cytosol. Here, we report an adaptive down-regulation of nuclear import supporting such an increased demand for nuclear export. The induction of cardiac cell hypertrophy by phenylephrine or angiotensin II inhibited the nuclear translocation of H1 histones. The removal of hypertrophic stimuli reversed the hypertrophic phenotype and restored nuclear import. Moreover, the inhibition of nuclear export by leptomycin B rescued import. Hypertrophic reprogramming increased the intracellular GTP/GDP ratio and promoted the nuclear redistribution of the GTP-binding transport factor Ran, favoring export over import. Further, in hypertrophy, the reduced creatine kinase and adenylate kinase activities limited energy delivery to the nuclear pore. The reduction of activities was associated with the closure of the cytoplasmic phase of the nuclear pore preventing import at the translocation step. Thus, to overcome the limited capacity for nucleocytoplasmic transport, cells requiring increased nuclear export regulate the nuclear transport pathway by undergoing a metabolic and structural restriction of nuclear import. PMID- 11283026 TI - A role for TAF3B2 in the repression of human RNA polymerase III transcription in nonproliferating cells. AB - RNA polymerase III (Pol III) synthesizes various small RNA species, including the tRNAs and the 5 S ribosomal RNA, which are involved in protein synthesis. Here, we describe the regulation of human Pol III transcription in response to sustained cell cycle arrest. The experimental system used is a cell line in which cell cycle arrest is induced by the regulated expression of the tumor suppressor protein p53. We show that the capacity of cells to carry out Pol III transcription from various promoter types, when tested in vitro, is severely reduced in response to sustained p53-mediated cell cycle arrest. Furthermore, this effect does not appear to be due to direct inhibition by p53. By using complementation assays, we demonstrate that a subcomponent of the Pol III transcription factor IIIB, which contains the proteins TATA-binding protein and TAF3B2, is the target of repression. Moreover, we reveal that TAF3B2 levels are markedly reduced in extracts from cell cycle-arrested cells because of a decrease in TAF3B2 protein stability. These findings provide a novel mechanism of Pol III regulation and yield insights into how cellular biosynthetic capacity and growth status can be coordinated. PMID- 11283027 TI - Identification, purification, and characterization of monoacylglycerol acyltransferase from developing peanut cotyledons. AB - Biosynthesis of diacylglycerols in plants occurs mainly through the acylation of lysophosphatidic acid in the microsomal membranes. Here we describe the first identification of diacylglycerol biosynthetic activity in the soluble fraction of developing oilseeds. This activity was NaF-insensitive and acyl-CoA-dependent. Diacylglycerol formation was catalyzed by monoacylglycerol (MAG) acyltransferase (EC ) that transferred an acyl moiety from acyl-CoA to MAG. The enzyme was purified by successive chromatographic separations on octyl-Sepharose, blue Sepharose, Superdex-75, and palmitoyl-CoA-agarose to apparent homogeneity from developing peanut (Arachis hypogaea) cotyledons. The enzyme was purified to 6,608 fold with the final specific activity of 15.86 nmol min(-1) mg(-1). The purified enzyme was electrophoretically homogeneous, and its molecular mass was 43,000 daltons. The purified MAG acyltransferase was specific for MAG and did not utilize any other acyl acceptor such as glycerol, glycerol-3-phosphate, lysophosphatidic acid, and lysophosphatidylcholine. The K(m) values for 1 palmitoylglycerol and 1-oleoylglycerol were 16.39 and 5.65 micrometer, respectively. The K(m) values for 2-monoacylglycerols were 2- to 4-fold higher than that of the corresponding 1-monoacylglycerol. The apparent K(m) values for palmitoyl-, stearoyl-, and oleoyl-CoAs were 17.54, 25.66, and 9.35 micrometer, respectively. Fatty acids, phospholipids, and sphingosine at low concentrations stimulated the enzyme activity. The identification of MAG acyltransferase in oilseeds suggests the presence of a regulatory link between signal transduction and synthesis of complex lipids in plants. PMID- 11283029 TI - Toward standardization of Epstein-Barr virus DNA load monitoring: unfractionated whole blood as preferred clinical specimen. AB - Epstein-Barr virus (EBV) DNA load monitoring in peripheral blood has been shown to be a useful tool for the diagnosis of aberrant EBV infections. In the present study we compared the relative diagnostic values of EBV DNA load monitoring in unfractionated whole blood and simultaneously obtained serum or plasma samples from Burkitt's lymphoma (BL) patients, transplant recipients, human immunodeficiency virus (HIV)-infected individuals, and infectious mononucleosis (IM) patients by a quantitative competitive PCR (Q-PCR). The EBV DNA load in BL patients was mainly situated in the cellular blood compartment (up to 4.5 x 10(6) copies/ml). EBV DNA loads in unfractionated whole blood and parallel serum samples showed no correlation. In transplant recipients, IM patients, and HIV infected patients, the EBV burden in the circulation was almost exclusively restricted to the cellular blood compartment, because serum or plasma samples from these patients yielded negative results by Q-PCR, despite high viral loads in corresponding whole-blood samples. A 10-fold more sensitive but qualitative BamHI-W-repeat PCR occasionally revealed the presence of EBV at <2,000 copies of EBV DNA per ml of serum. Spiking of 100 copies of EBV DNA in samples with negative Q-PCR results excluded the presence of inhibitory factors in serum or plasma that influenced the Q-PCR result. Serum samples from all populations were often positive for beta-globin DNA, indicating cell damage in vivo or during serum preparation. We conclude that serum is an undesirable clinical specimen for EBV DNA load monitoring because it omits the presence of cell-associated virus and uncontrolled cell lysis may give irreproducible results or overestimation of the DNA load. Unfractionated whole blood is strongly preferred since it combines all blood compartments that may harbor EBV and it best reflects the absolute viral burden in the patient's circulation. PMID- 11283030 TI - PCR-based diagnosis of Helicobacter pylori infection and real-time determination of clarithromycin resistance directly from human gastric biopsy samples. AB - A novel PCR detection assay that amplifies the Helicobacter pylori-specific vacuolating cytotoxin gene (vacA) and thus enables rapid diagnosis of infection is described. Additionally, a real-time probe hybridization melting point analysis assay to detect all three mutations in the 23S rRNA gene associated with clarithromycin resistance was applied directly to antral gastric biopsy samples. Comparison with culture and an alternative PCR assay targeting the 16S rrn gene showed that the vacA assay was sensitive and specific when tested on biopsy samples from 121 patients. Clarithromycin susceptibilities could be determined in the majority (92.3%) of culture-positive gastric biopsy samples analyzed, four of which generated melting peaks indicative of clarithromycin resistance by either an A-->G or A-->C mutation. The presence of the mutations correlated with the clarithromycin disk diffusion sensitivities of matched cultures. This PCR-based system was simple to perform and could be completed in 3 to 4 h, thereby overcoming the delays associated with conventional culture methods for H. pylori identification and susceptibility testing. PMID- 11283032 TI - Novel mycobacterium related to Mycobacterium triplex as a cause of cervical lymphadenitis. AB - The Mycobacterium avium complex (MAC) is an important cause of cervical lymphadenitis in children, and its incidence appears to be increasing in the United States and elsewhere. In areas where Mycobacterium tuberculosis is not prevalent, M. avium causes the vast majority of cases of mycobacterial lymphadenitis, although several other nontuberculous mycobacterial species have been reported as etiologic agents. This report describes the case of a child with cervical lymphadenitis caused by a nontuberculous mycobacterium that could not be identified using standard methods, including biochemical reactions and genetic probes. Direct 16S ribosomal DNA sequencing showed greater than 99% homology with Mycobacterium triplex, but sequence analysis of the 283-bp 16S-23S internal transcribed spacer (ITS) sequence showed only 95% identity, suggesting that it is a novel species or subspecies within a complex of organisms that includes M. triplex. Mycolic acid high-performance liquid chromatography analysis also identified this isolate as distinct from M. triplex, and differences in susceptibility to streptomycin and rifampin between this strain and M. triplex were also observed. These data support the value of further testing of clinical isolates that test negative with the MAC nucleic acid probes and suggest that standard methods used for the identification of mycobacteria may underestimate the complexity of the genus Mycobacterium. ITS sequence analysis may be useful in this setting because it is easy to perform and is able to distinguish closely related species and subspecies. This level of discrimination may have significant clinical ramifications, as closely related organisms may have different antibiotic susceptibility patterns. PMID- 11283031 TI - Molecular evidence of Bartonella spp. in questing adult Ixodes pacificus ticks in California. AB - Ticks are the vectors of many zoonotic diseases in the United States, including Lyme disease, human monocytic and granulocytic ehrlichioses, and Rocky Mountain spotted fever. Most known Bartonella species are arthropod borne. Therefore, it is important to determine if some Bartonella species, which are emerging pathogens, could be carried or transmitted by ticks. In this study, adult Ixodes pacificus ticks were collected by flagging vegetation in three sites in Santa Clara County, Calif. PCR-restriction fragment length polymorphism and partial sequencing of 273 bp of the gltA gene were applied for Bartonella identification. Twenty-nine (19.2%) of 151 individually tested ticks were PCR positive for Bartonella. Male ticks were more likely to be infected with Bartonella than female ticks (26 versus 12%, P = 0.05). None of the nine ticks collected at Baird Ranch was PCR positive for Bartonella. However, 7 (50%) of 14 ticks from Red Fern Ranch and 22 (17%) of 128 ticks from the Windy Hill Open Space Reserve were infected with Bartonella. In these infected ticks, molecular analysis showed a variety of Bartonella strains, which were closely related to a cattle Bartonella strain and to several known human-pathogenic Bartonella species and subspecies: Bartonella henselae, B. quintana, B. washoensis, and B. vinsonii subsp. berkhoffii. These findings indicate that I. pacificus ticks may play an important role in Bartonella transmission among animals and humans. PMID- 11283033 TI - Biochemical identification and characterization of DNA groups within the Proteus vulgaris complex. AB - We placed 43 isolates belonging to the Proteus vulgaris complex into proposed DNA groups (genomovars) using five previously recommended tests (tests for esculin hydrolysis, production of acid from salicin, L-rhamnose fermentation, and elaboration of DNase and lipase). On the basis of the results of these five tests, 49% of the isolates fell into DNA groups 5 and 6, 37% fell into DNA group 2, and the remaining 14% fell into DNA groups 3 and 4. Sequencing of the 16S rRNA genes of 11 members of DNA groups 5 and 6 indicated that 10 of these isolates (91%) could be unambiguously assigned to one of these two genomospecies. Overall expression of selected enzymatic and virulence-associated characteristics did not differ significantly among DNA groups. PMID- 11283034 TI - Phenotypic and genotypic approaches to characterization of isolates of Neisseria meningitidis from patients and their close family contacts. AB - Characterization of isolates of Neisseria meningitidis obtained from patients with meningococcal disease or from pharyngeal swabs of asymptomatic carriers can be achieved by several approaches which provide different levels of discrimination. A total of 45 gram negative, oxidase-positive diplococcus strains isolated from 15 individuals with meningococcal disease and 30 of their family contacts were examined by three approaches: serological typing, multilocus enzyme electrophoresis (MLEE), and multilocus sequence typing (MLST). For 10 of the 15 patient and contact groups, all of the isolates were confirmed as meningococci, and the bacteria obtained from the patients and contacts, including their mother or principal caregiver in the case of children, were indistinguishable by all three methods. In the remaining five groups the isolates from the patients were distinct from those recovered from the contacts, and in three examples, in two separate groups, the contacts were shown by MLST to be carrying strains of Neisseria lactamica. The data obtained from the three techniques were consistent, although complete serological typing was possible for only a minority of isolates. Both MLEE and MLST established the genetic relationships of the isolates and identified members of known hypervirulent lineages, but MLST was faster than MLEE and had the additional advantages that it could be performed on noninfective material distributed by mail and that the results from different laboratories could be compared via the internet (http://mlst.zoo.ox.ac.uk). PMID- 11283035 TI - Phenotypic and genotypic characterization of Pediococcus strains isolated from human clinical sources. AB - Seventy-two strains of pediococci isolated from human clinical sources were characterized by conventional physiological tests, chromogenic enzymatic tests, analysis of whole-cell protein profiles (WCPP) by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and analysis of chromosomal DNA restriction profiles by pulsed-field gel electrophoresis (PFGE). Conventional tests allowed identification of 67 isolates: 52 strains were identified as Pediococcus acidilactici, 15 strains were identified as Pediococcus pentosaceus, and 5 strains were not identified because of atypical reactions. Analysis of WCPP identified all isolates since each species had a unique WCPP. By the WCPP method, the atypical strains were identified as P. acidilactici (two strains) and P. pentosaceus (three strains). The chromogenic substrate test with o-nitrophenyl beta-D-glucopyranoside differentiated all 54 strains of P. acidilactici (negative reactions) and 13 (72%) of 18 strains of P. pentosaceus (positive reactions). Isolates of both species were shown to be nonclonal as revealed by the genetic diversity when chromosomal DNA was analyzed by PFGE. Using WCPP as the definitive identification procedure, P. acidilactici (28 of 54 strains; 51.8%) was more likely than P. pentosaceus (4 of 18 strains; 22.3%) to be isolated from blood cultures. PMID- 11283037 TI - Performance of the applied biosystems ViroSeq human immunodeficiency virus type 1 (HIV-1) genotyping system for sequence-based analysis of HIV-1 in pediatric plasma samples. AB - The ViroSeq HIV-1 Genotyping System is a commercially available, integrated sequence-based system for analysis of human immunodeficiency virus type 1 (HIV-1) drug resistance. We evaluated the performance of this system by analyzing HIV-1 in pediatric plasma samples. Plasma samples from children 4 months to 17 years of age were obtained from a clinical trial protocol (PACTG 377). Children in PACTG 377 were randomized to four treatment arms, including different combinations of antiretroviral drugs. HIV-1 genotyping was performed using samples collected prior to antiretroviral therapy (baseline) and at the time of virologic failure. Performance of the genotyping system was compared in three university laboratories. A total of 196 samples were analyzed, including 135 baseline and 61 failure samples. Plasma volumes ranged from 0.05 to 0.5 ml, and viral loads ranged from 1,084 to 3,484,991 copies/ml. PCR products suitable for sequencing were obtained for 192 of the 196 samples. Complete sequences for protease and reverse transcriptase were obtained for all of these 192 samples. For 180 samples, data were obtained from both DNA strands for the entire region analyzed. There was no evidence of sample cross-contamination based on phylogenetic analysis of HIV-1 sequences. Performance of the genotyping system was similar in three laboratories. This genotyping system performs well for analysis of HIV-1 in pediatric plasma samples, including those with low volume and low viral load. The availability of this system should facilitate studies of HIV-1 drug resistance. PMID- 11283036 TI - Serological, epidemiological, and molecular differences between human T-cell lymphotropic virus Type 1 (HTLV-1)-seropositive healthy carriers and persons with HTLV-I Gag indeterminate Western blot patterns from the Caribbean. AB - To investigate the significance of serological human T-cell lymphotropic virus type 1 (HLTV-1) Gag indeterminate Western blot (WB) patterns in the Caribbean, a 6-year (1993 to 1998) cross-sectional study was conducted with 37,724 blood donors from Guadeloupe (French West Indies), whose sera were routinely screened by enzyme immunoassay (EIA) for the presence of HTLV-1 and -2 antibodies. By using stringent WB criteria, 77 donors (0.20%) were confirmed HTLV-1 seropositive, whereas 150 (0.40%; P < 0.001) were considered HTLV seroindeterminate. Among them, 41.3% (62) exhibited a typical HTLV-1 Gag indeterminate profile (HGIP). Furthermore 76 (50.7%) out of the 150 HTLV seroindeterminate subjects were sequentially retested, with a mean duration of follow-up of 18.3 months (range, 1 to 70 months). Of these, 55 (72.4%) were still EIA positive and maintained the same WB profile whereas the others became EIA negative. This follow-up survey included 33 persons with an HGIP. Twenty-three of them (69.7%) had profiles that did not evolve over time. Moreover, no case of HTLV-1 seroconversion could be documented over time by studying such sequential samples. HTLV-1 seroprevalence was characterized by an age-dependent curve, a uniform excess in females, a significant relation with hepatitis B core (HBc) antibodies, and a microcluster distribution along the Atlantic coast of Guadeloupe. In contrast, the persons with an HGIP were significantly younger, had a 1:1 sex ratio, did not present any association with HBc antibodies, and were not clustered along the Atlantic facade. These divergent epidemiological features, together with discordant serological screening test results for subjects with HGIP and with the lack of HTLV-1 proviral sequences detected by PCR in their peripheral blood mononuclear cell DNA, strongly suggest that an HGIP does not reflect true HTLV-1 infection. In regard to these data, healthy blood donors with HGIP should be reassured that they are unlikely to be infected with HTLV-1 or HTLV-2. PMID- 11283038 TI - Electrophoretic karyotype analysis of sequential Candida parapsilosis isolates from patients with persistent or pecurrent fungemia. AB - We assessed the genetic relatedness of sequential isolates of Candida parapsilosis during persistent or recurrent fungemia and the effect of central venous catheter (CVC) removal. Serial isolates of C. parapsilosis were obtained from 17 patients with persistent or recurrent fungemia over periods of up to 5 months. Forty-eight C. parapsilosis isolates from the blood of 17 patients were analyzed by electrophoretic karyotyping (EK) with pulsed-field gel electrophoresis (PFGE), revealing 25 different karyotypes. The strains sequentially isolated from each of seven patients whose fungemia resolved following CVC removal had the same karyotype. Two patients with fungemia that cleared without CVC removal each had two sequential isolates with different karyotypes. In six (75%) of the eight patients whose fungemia was recurrent even after CVC removal, the karyotypes of the pre- and post-CVC removal isolates were different, implying the emergence of a new strain. Overall, the sequential strains from each patient had identical karyotypes in 53% (9 of 17) of the patients and two different karyotypes in 47% (8 of 17). This study shows that EK with PFGE is useful for investigating persistent or recurrent fungemia due to C. parapsilosis and that recurrent fungemia due to C. parapsilosis is more likely caused by reinfection with a second strain. PMID- 11283040 TI - Detection of Mycobacterium bovis in bovine clinical specimens using real-time fluorescence and fluorescence resonance energy transfer probe rapid-cycle PCR. AB - Nucleic acid sequence capture extraction was coupled with LightCycler PCR amplification and product detection using real-time fluorescence for rapid, definitive detection of Mycobacterium bovis in lymph node specimens from 38 cattle with bovine tuberculosis lesions. PCR amplification of sequence-captured DNA using both a conventional heating block thermocycler and a LightCycler thermocycler was compared with culture and histopathological analyses. Conventional PCR enabled detection of 26 of 28 culture-positive specimens (93%) in approximately 9 h, and the LightCycler PCR detected 20 of 28 culture-positive specimens (71%) in only 30 min. Specific confirmation of Mycobacterium tuberculosis complex DNA was achieved by LightCycler PCR amplification using Syb Green 1 and an M. tuberculosis complex-specific Cy5-labeled fluorescence resonance energy transfer probe. The system described here enabled rapid and specific laboratory confirmation of bovine tuberculosis, and this is the first report of the detection of M. bovis in tissues using LightCycler PCR. The fluorescence technology used in the study has potential to allow development of a high-throughput molecular diagnostic test for bovine tuberculosis. PMID- 11283039 TI - High-throughput detection of West Nile virus RNA. AB - The recent outbreaks of West Nile virus (WNV) in the northeastern United States and other regions of the world have made it essential to develop an efficient protocol for surveillance of WNV. In the present report, we describe a high throughput procedure that combines automated RNA extraction, amplification, and detection of WNV RNA. The procedure analyzed 96 samples in approximately 4.5 h. A robotic system, the ABI Prism 6700 Automated Nucleic Acid workstation, extracted RNA and set up reactions for real-time reverse transcription (RT)-PCR in a 96 well format. The robot extracted RNA with a recovery as efficient as that of a commercial RNA extraction kit. A real-time RT-PCR assay was used to detect and quantitate WNV RNA. Using in vitro transcribed RNA, we estimated the detection limit of the real-time RT-PCR to be approximately 40 copies of RNA. A standard RT PCR assay was optimized to a sensitivity similar to that of the real-time RT-PCR. The standard assay can be reliably used to test a small number of samples or to confirm previous test results. Using internal primers in a nested RT-PCR, we increased the sensitivity by approximately 10-fold compared to that of the standard RT-PCR. The results of the study demonstrated for the first time that the use of an automated system for the purpose of large-scale viral RNA surveillance dramatically increased the speed and efficiency of sample throughput for diagnosis. PMID- 11283041 TI - Serologic detection of Actinobacillus pleuropneumoniae in swine by capsular polysaccharide-biotin-streptavidin enzyme-linked immunosorbent assay. AB - Serologic detection of Actinobacillus pleuropneumoniae infections in swine have been problematic due to antigenic cross-reactivity of Apx toxins, lipopolysaccharide, and outer membrane proteins between A. pleuropneumoniae serotypes and other bacterial species. To maximize serologic specificity and sensitivity, we developed an assay that uses highly purified A. pleuropneumoniae capsular polysaccharide (CP) conjugated to biotin, which is then bound to streptavidin-coated enzyme-linked immunosorbent assay (CP-BS-ELISA) plates. This assay was used to test a panel of 240 serum samples from pigs prior to challenge, after challenge with bacterial species other than A. pleuropneumoniae, or after challenge with A. pleuropneumoniae serotype 1, 5, or 7. Overall assay results for the individual sera tested were reproducible on the same day and on separate days. The sensitivity of the assay was 100% by ELISAs with biotin-CPs of serotypes 1 and 7 and 87.5% by ELISAs with biotin-CP of serotype 5. Specificity was 100% by ELISAs with biotin-CPs of serotypes 1 and 5 and 94.5% by ELISAs with biotin-CP of serotype 7. The biotin-CPs of at least three A. pleuropneumoniae serotypes could be combined for use in a screening assay to detect antibodies to CPs from strains of different serotypes. In conclusion, the CP-BS-ELISA proved to be a serotype-specific and species-specific assay with high sensitivity for the identification of pigs exposed to A. pleuropneumoniae. PMID- 11283043 TI - Comparison of nucleic acid amplification, serology, and microbiologic culture for diagnosis of Rhodococcus equi pneumonia in foals. AB - Recently, a technique was described for amplification of Rhodococcus equi specific chromosomal and vapA DNA from blood and tracheal wash fluids. It was hypothesized that this technique would be more sensitive than standard culture techniques or serology for diagnosis of R. equi pneumonia in foals. Tracheal wash fluid, nasal swabs, whole blood samples, and serum samples from 56 foals with pneumonia were analyzed. Final clinical diagnosis was determined by the attending clinician on the basis of final interpretation of all available information about each foal, including clinical presentation, diagnostic test results, response to therapy, and outcome. Clinical diagnosis was used as a final reference standard for calculation of sensitivity, specificity, and predictive values for PCR, serology using an agar gel immunodiffusion test, and tracheal wash fluid culture. PCR of tracheal wash fluid using primers that recognized the vapA virulence plasmid of R. equi had a diagnostic sensitivity of 100% and specificity of 90.6%. Sensitivity and specificity were 57.1 and 93.8%, respectively, for standard microbiologic culture of tracheal wash fluid and 62.5 and 75.9%, respectively, for serology. PCR of tracheal wash fluid is more sensitive and specific for diagnosis of R. equi pneumonia than are other available diagnostic tests. PMID- 11283042 TI - Serum immunoglobulin G immune response to Helicobacter pylori antigens in Mongolian gerbils. AB - The Mongolian gerbil model for Helicobacter pylori infection is an animal model that mimics human disease. We examined the serum immune response to H. pylori infection in gerbils by enzyme-linked immunosorbent assay (ELISA) and Western blotting, both with whole-cell (H. pylori) extracts. A total of 66 7-week-old specific-pathogen-free male gerbils were inoculated orogastrically with H. pylori strain ATCC 43504. Sera were collected 1, 2, 4, 8, 12, 26, 38, and 52 weeks after H. pylori inoculation. Sixty-nine noninfected gerbils and their sera were used as controls. The specificity of the ELISA was 95.7%. The frequency of seropositivity increased over time: 2 of 10 (20%), 7 of 10 (70%), and 7 of 7 (100%) samples of sera from inoculated gerbils were positive for H. pylori at 2, 4, and 8 weeks postinoculation, respectively. Western blot assays showed that the primary immunoglobulin G (IgG) response against low-molecular-mass (25-, 30-, and 20-kDa) proteins appeared after a lag period of 2 to 8 weeks after inoculation. Antibodies against 160-, 150-, 110-, 120-, 80-, 66-, and 63-kDa proteins were observed 12 weeks after inoculation. The early reactive 30-kDa protein was identified as a urease alpha subunit by N-terminal amino acid sequencing. After 26 weeks, two groups of animals could be distinguished: one group developed ulcers (n = 5), and the other developed hyperplastic polyps without ulcers (n = 19). Gerbils in the gastric ulcer group showed significantly higher serum anti-H. pylori IgG levels than did gerbils in the hyperplastic group (P = 0.001) as measured by ELISA. Furthermore, a higher proportion of animals developed antibodies to H. pylori proteins of 26, 25, and 20 kDa in the ulcer group than those animals with hyperplastic polyps (75 to 100% versus 17 to 50%) in Western blot assays. These results highlight the importance of the immune response of the host in the development of H. pylori-related gastric lesions. PMID- 11283044 TI - Characterization of Lyme borreliosis isolates from patients with erythema migrans and neuroborreliosis in southern Sweden. AB - Southern Sweden is an area of Lyme borreliosis (LB) endemicity, with an incidence of 69 cases per 100,000 inhabitants. The most frequent clinical manifestations are erythema migrans (77%) and neuroborreliosis (16%). There was no record of human Borrelia strains being isolated from patients in this region before the prospective study reported here. Borrelia spirochetes were isolated from skin and cerebrospinal fluid (CSF) from LB patients living in the region. A total of 39 strains were characterized by OspA serotype analysis, species-specific PCR, and signature nucleotide analysis of the 16S rRNA gene. Of 33 skin isolates, 31 (93.9%) were Borrelia afzelii strains and 2 (6.1%) were Borrelia garinii strains. Of six CSF isolates, five (83.3%) were B. garinii and one (16.7%) was B. afzelii. Neither Borrelia burgdorferi sensu stricto strains nor multiple infections were observed. The B. afzelii isolates were of OspA serotype 2. Three B. garinii strains were of OspA serotype 5, and the remaining four strains were of OspA serotype 6. All of the B. garinii strains belonged to the same 16S ribosomal DNA ribotype class. Our findings agree with earlier findings from other geographic regions in Europe where B. afzelii and B. garinii have been recovered predominantly from skin and CSF cultures, respectively. To further study the possible presence in Sweden of the genotype B. burgdorferi sensu stricto, which is known to be present in Europe and to occur predominantly in patients with Lyme arthritis, molecular detection of Borrelia-specific DNA in synovial samples from Lyme arthritis patients should be performed. PMID- 11283045 TI - Type-specific detection of echovirus 30 isolates using degenerate reverse transcriptase PCR primers. AB - Following an approach used to specifically identify polioviruses and enterovirus 71, we have developed reverse transcriptase (RT) PCR primers containing mixed base residues or deoxyinosine at positions of codon degeneracy. These primers permit specific RT-PCR amplification of echovirus 30 (E30) sequences by targeting sites that encode conserved amino acid motifs within the major capsid protein, VP1. All 221 E30 strains tested, isolated in 16 countries over a 44-year period, yielded the predicted 158-bp PCR product. No specific products were obtained by PCR assays containing templates from any of the other 63 EV serotypes. Inosine containing degenerate primers may be widely applicable to the identification of echovirus serotypes by PCR. PMID- 11283046 TI - Quantification of human immunodeficiency virus type 1 proviral load by a TaqMan real-time PCR assay. AB - Proviral human immunodeficiency virus type 1 (HIV-1) DNA could be a useful marker for exploring viral reservoirs and monitoring antiretroviral treatment, particularly when HIV-1 RNA is undetectable in plasma. A new technique was developed to quantify proviral HIV-1 using a TaqMan real-time PCR assay. One copy of proviral HIV-1 DNA could be detected with 100% sensitivity for five copies and the assay had a range of 6 log(10). Reproducibility was evaluated in intra- and interassays using independent extractions of the 8E5 cell line harboring the HIV 1 proviral genome (coefficients of variation [CV], 13 and 27%, respectively) and peripheral blood mononuclear cells (PBMC) from a patient with a mean proviral load of 26 copies per 10(6) PBMC (CV, 46 and 56%, respectively). The median PBMC proviral load of 21 patients, measured in a cross-sectional study, was determined to be 215 copies per 10(6) PBMC (range, <10 to 8,381). In a longitudinal study, the proviral load of 15 out of 16 patients with primary infection fell significantly during 1 year of antiretroviral therapy (P = 0.004). In the remaining patient, proviral HIV-1 DNA was detectable but not quantifiable due to a point mutation at the 5' end of the TaqMan probe. No correlation was observed between proviral load and levels of CD4(+) cells or HIV-1 RNA in plasma. TaqMan PCR is sensitive and adaptable to a large series of samples. The full interest of monitoring proviral HIV-1 DNA can now be ascertained by its application to the routine monitoring of patients. PMID- 11283047 TI - Differentiation of resistance phenotypes among erythromycin-resistant Pneumococci. AB - Laboratory differentiation of erythromycin resistance phenotypes is poorly standardized for pneumococci. In this study, 85 clinical isolates of erythromycin resistant (MIC > or = 1 microg/ml) Streptococcus pneumoniae were tested for the resistance phenotype by the erythromycin-clindamycin double-disk test (previously used to determine the macrolide resistance phenotype in Streptococcus pyogenes strains) and by MIC induction tests, i.e., by determining the MICs of macrolide antibiotics without and with pre-exposure to 0.05 microg of erythromycin per ml. By the double-disk test, 65 strains, all carrying the erm(AM) determinant, were assigned to the constitutive macrolide, lincosamide, and streptogramin B resistance (cMLS) phenotype, and the remaining 20, all carrying the mef(E) gene, were assigned to the recently described M phenotype; an inducible MLS resistance (iMLS) phenotype was not found. The lack of inducible resistance to clindamycin was confirmed by determining clindamycin MICs without and with pre-exposure to subinhibitory concentrations of erythromycin. In macrolide MIC and MIC-induction tests, whereas homogeneous susceptibility patterns were observed among the 20 strains assigned to the M phenotype by the double-disk test, two distinct patterns were recognized among the 65 strains assigned to the cMLS phenotype by the same test; one pattern (n = 10; probably that of the true cMLS isolates) was characterized by resistance to rokitamycin also without induction, and the other pattern (n = 55; designated the iMcLS phenotype) was characterized by full or intermediate susceptibility to rokitamycin without induction turning to resistance after induction, with an MIC increase by more than three dilutions. A triple-disk test, set up by adding a rokitamycin disk to the erythromycin and clindamycin disks of the double-disk test, allowed the easy differentiation not only of pneumococci with the M phenotype from those with MLS resistance but also, among the latter, of those of the true cMLS phenotype from those of the iMcLS phenotype. While distinguishing MLS from M resistance in pneumococci is easily and reliably achieved, the differentiation of constitutive from inducible MLS resistance is far more uncertain and is strongly affected by the antibiotic used to test inducibility. PMID- 11283048 TI - Epidemiological usefulness of anti-opacity factor antibody screening in schoolchildren. AB - The presence of the anti-opacity factor (anti-OF) antibody (Ab) in the serum used for identifying the OF antigen (Ag) type represents previous or current infection with group A streptococci (GAS) of the OF Ag type. Throat cultures were taken from 172 elementary schoolchildren in Chinju, Korea, and venous blood samples were collected at the same time to screen for the frequency of the anti-OF Ab. After isolation of GAS, the OF Ag of each GAS was identified by inhibition of the opacity reaction with recognized anti-OF sera. The anti-OF Abs in the sera were screened with the six most common OF Ag types. OF22 and OF28 were high in prevalence (28.2 and 20.5%, respectively) among OF Ag types, while anti-OF types 4, 28, and 22 were frequently identified (39.5, 29.7, and 15.7%, respectively) in the sera. Thirty-two of 39 (82.1%) OF Ag-producing GAS carriers, 25 of 34 (73.5%) GAS carriers not producing OF Ag, and 72 of 99 (72.7%) throat culture-negative children harbored the anti-OF Ab. Forty-five (26.2%) of 172 children had two different anti-OF Abs, and 11 (6.4%) had more than three anti-OF Abs. Seventy five percent of 172 elementary schoolchildren were shown to be previously or currently infected with GAS. The percentages of children positive for the anti-OF Ab were very high regardless of the result of throat culture or OF Ag production of GAS. We could also demonstrate (i) that the prevalent strains of GAS changed according to the time span by determining the difference between the frequencies of OF Ag and anti-OF Ab and (ii) that repeated infections were not uncommon in schoolchildren, as one-third had more than two different anti-OF Abs. PMID- 11283049 TI - cagA Status and eradication treatment outcome of anti-Helicobacter pylori triple therapies in patients with nonulcer dyspepsia. AB - The differences in eradication rates reported in clinical trials aiming to cure Helicobacter pylori infection cannot be entirely explained by the type of regimen, bacterial resistance, or lack of compliance. Using data from a clinical trial, a logistic regression model was constructed to determine whether cagA status, assessed by PCR, affects the outcome of eradication. Resistance to clarithromycin (10% of the strains) predicted failure perfectly. In the model (n = 156), a cagA-lacking strain (odds ratio [OR] = 2.2; 95% confidence interval [CI], (1.1 to 4.7), tobacco smoking OR = 3.1; 95% CI, 1.3 to 7.0), and a double dose of proton pump inhibitor in the treatment regimen (OR = 0.3; 95% CI, 0.2 to 0.7) were associated with the treatment outcome. The exact role of cagA in the outcome of H. pylori eradication therapy has not been explored. However, the type of histological lesions which it causes in the gastric mucosa may be implicated. Regardless of the mechanism involved, cagA status is a good predictive marker of eradication outcome. PMID- 11283050 TI - Helicobacter pylori infection in an urban African population. AB - We have studied 221 adults drawn from an impoverished urban population with high human immunodeficiency virus (HIV) seroprevalence (35%) to determine the prevalence of gastroduodenal pathology and its relationship to serological markers of Helicobacter pylori virulence proteins and other potential environmental and immunological determinants of disease including HIV infection. Eighty-one percent were H. pylori seropositive, and 35% were HIV seropositive. Urban upbringing and low CD4 count were associated with a reduced likelihood of H. pylori seropositivity, as was current Ascaris infection, in keeping with recent evidence from an animal model. One hundred ninety-one adults underwent gastroduodenoscopy, and 14 had gastroduodenal pathology. Mucosal lesions were a major cause of abdominal pain in this population. While the majority of patients with gastroduodenal pathology (12 of 14) were seropositive for H. pylori, none were seropositive for HIV. Smoking was associated with increased risk of macroscopic pathology, and a history of Mycobacterium bovis BCG immunization was associated with reduced risk. Antibodies to H. pylori lipopolysaccharide were associated with pathology. HIV infection was associated with protection against mucosal lesions, suggesting that fully functional CD4 lymphocytes may be required for the genesis of gastroduodenal pathology. PMID- 11283051 TI - Evaluation of Etest for direct antifungal susceptibility testing of yeasts in positive blood cultures. AB - The performance of the Etest (AB BIODISK, Solna, Sweden) for direct antifungal susceptibility testing of yeasts in positive blood cultures was compared with that of the macrodilution method for determining the MICs of five antifungal agents. Culture broths with blood from bottles positive for yeasts were inoculated directly onto plates for susceptibility testing with the Etest, and the MICs were read after 24 and 48 h of incubation. A total of 141 positive blood cultures (72 cultures of Candida albicans, 31 of Candida tropicalis, 14 of Candida glabrata, 11 of Candida parapsilosis, 3 of Candida krusei, and 3 of Cryptococcus neoformans, 4 miscellaneous yeast species, and 3 mixed cultures) were tested, and the rates of MIC agreement (+/-1 log(2) dilution) between the direct Etest (at 24 and 48 h, respectively) and macrodilution methods were as follows: amphotericin B, 81.8 and 93.5%; flucytosine, 84.8 and 87.7%; fluconazole, 89.4 and 85.5%; itraconazole, 69.7 and 63.8%; ketoconazole, 87.9 and 79.0%. By a large-sample t test, the difference in log(2) dilution between the direct Etest and the macrodilution method was found to be small (P < 0.05). The lone exceptions were ketoconazole at 48 h of incubation and itraconazole at both 24 and 48 h of incubation (P > 0.05). By Tukey's multiple comparisons, the difference between the direct Etest (48 h) and reference methods among different species was found to be less than 1 log(2) dilution. When the MICs were translated into interpretive susceptibility, the minor errors caused by the direct Etest (at 24 and 48 h, respectively) were as follows: flucytosine, 2.3 and 1.4%; fluconazole, 3.0 and 3.6%; itraconazole, 21.2 and 21.3%. Itraconazole also produced an additional 3.0 and 3.6% major errors as determined by the direct Etest at 24 and 48 h, respectively. It was concluded that, except for itraconazole, the Etest method was feasible for direct susceptibility testing of blood cultures positive for yeasts. The method is simple, and the results could be read between 24 and 48 h after direct inoculation, whenever the inhibition zones were discernible. PMID- 11283052 TI - Comparison of quantitative and qualitative PCR assays for cytomegalovirus DNA in plasma. AB - We analyzed the performance characteristics of the qualitative AMPLICOR CMV Test (Roche Molecular Systems, Pleasanton, Calif.) and quantitative COBAS AMPLICOR CMV MONITOR Test (Roche Molecular Systems) assays and compared the performance of the AMPLICOR quantitative assay with an in-house-developed cytomegalovirus (CMV) DNA PCR assay. The quantitative AMPLICOR assay was found to be more sensitive than the qualitative AMPLICOR assay. The quantitative AMPLICOR assay has a lower limit of sensitivity of 400 CMV DNA copies/ml of plasma and is linear to 50,000 CMV DNA copies/ml of plasma. Compared to the in-house PCR assay, the AMPLICOR quantitative assay gave lower viral load values at all concentrations tested, but the difference between the two assays was not consistent across the entire dynamic range of the AMPLICOR quantitative assay. At the lower end of the assay, the viral load values obtained with the in-house PCR assay were three- to fivefold (0.5 to 0.7 log units) higher than those measured with the AMPLICOR assay. At higher input concentrations, the differences between the two assays approached 10-fold. This direct comparison of the in-house assay and the quantitative AMPLICOR assay provides the ability to compare previously published in-house data with an assay widely available for future research and clinical monitoring of patients with CMV infections. PMID- 11283054 TI - Inoculum standardization for antifungal susceptibility testing of filamentous fungi pathogenic for humans. AB - Two methods of inoculum preparation for filamentous fungi were compared: counting with a hematocytometer and spectrophotometric adjustment. One hundred eighty-two filamentous fungi pathogenic for humans were used. Colony counts were done for all inoculum preparations. The agreement between the hematocytometer counts and the colony counts (CFU per milliliter) was 97.2%. The reproducibility between the hematocytometer counts and the colony counts by means of an intraclass correlation coefficient was 0.70. Pearson's correlation index for hematocytometer counts versus colony counts was 0.56, whereas that for optical density versus colony counts was 0.008. Both methods can be used for inoculum size adjustment. However, the use of the spectrophotometric method requires that each species be standardized separately. PMID- 11283053 TI - Assessment of Helicobacter pylori vacA and cagA genotypes and host serological response. AB - Helicobacter pylori strains can be distinguished by genotyping of virulence associated genes, such as vacA and cagA. Because serological discrimination between strain types would reduce the need for endoscopy, 61 patients carrying H. pylori were studied by vacA and cagA genotyping of H. pylori in gastric biopsy specimens and by detection of specific serum antibodies. Serological responses to H. pylori were determined by Helicoblot (versions 2.0 and 2.1). Antibodies to CagA also were determined by a rapid anti-CagA assay (Pyloriset screen CagA) as well as by two noncommercially developed enzyme immunoassays, each using a recombinant CagA protein. Assessment of performance of the Helicoblot assays indicated substantial interobserver variation, with kappa values between 0.20 and 0.93. There was no relationship between the serological profiles on the Helicoblot and the genotypes from the same patients, except for strong associations between the presence of anti-CagA and the cagA-positive and vacA s1 H. pylori genotypes. Detection of anti-CagA by the five different assays varied considerably, with kappa values ranging from 0.21 to 0.78. Using the cagA genotype as the "gold standard," the sensitivity and specificity of the anti-CagA assays varied from 71.4 to 85.7% and from 54.2 to 100%, respectively. Thus, serological profiles of antibodies to H. pylori are heterogeneous and, with the exception of anti-CagA antibodies, show no relation to the H. pylori vacA and cagA genotypes. Detection of anti-CagA antibodies is strongly dependent on the test used. PMID- 11283055 TI - Genotypic, clinical, and demographic characteristics of children infected with Helicobacter pylori. AB - Helicobacter pylori isolates vary between geographic regions. Certain H. pylori genotypes may be associated with disease outcome. Thirty-eight children underwent diagnostic upper endoscopy at four medical centers and were retrospectively analyzed to determine if H. pylori virulence genes were associated with endoscopic disease severity, histologic parameters, and host demographics. The H. pylori virulence genotype was analyzed by a reverse hybridization line probe assay and type-specific PCR. Endoscopic ulcers or erosions were found in 17 (45%) patients, with 13 (34%) of these patients having antral nodularity. Histological gastritis, of varying severity, was present in all children. Four patients harbored more than one H. pylori strain: one subject had both cagA(+) and cagA negative strains, while three patients harbored either two different cagA negative strains (two children) or two cagA(+) strains (one child). There were 28 (74%) cagA(+) isolates; 19 were associated with the vacA s1b genotype, 7 were associated with the vacA s1a genotype, 1 was associated with the vacA s1c genotype, and 1 was associated with the s2 genotype. Of 14 cagA-negative isolates, 6 were vacA s2 genotype, 4 were vacA s1b, 3 were vacA s1a, and 1 was vacA s1c. Nine of ten (90%) Hispanics had similar H. pylori strains (vacA s1b,m1), and all Asian-Canadian children were infected by strains with vacA s1c genotype. No correlation between H. pylori strain and endoscopic or histopathologic abnormalities was found. This study provides a baseline framework of North American children and their H. pylori strains, serving as a powerful epidemiological tool for prospective investigations to better understand the transmission and evolution of diverse disease outcomes. PMID- 11283056 TI - Molecular characterization of invasive and noninvasive Campylobacter jejuni and Campylobacter coli isolates. AB - Campylobacter jejuni is one of the most common causes of bacterial diarrhea worldwide and is the primary bacterial cause of food-borne illness. Adherence to and invasion of epithelial cells are the most important pathogenic mechanisms of Campylobacter diarrhea. Molecular characterization of invasive and noninvasive Campylobacter isolates from children with diarrhea and symptom-free children was performed by random amplified polymorphic DNA techniques (RAPD). A distinct RAPD profile with a DNA band of 1.6 kb was observed significantly more frequently among invasive (63%) than among noninvasive (16%) Campylobacter isolates (P = 0.000005). The 1.6-kb band was named the invasion-associated marker (IAM). Using specifically designed primers, a fragment of 518 bp of the iam locus was amplified in 85% of invasive and 20% of noninvasive strains (P = 0.0000000). Molecular typing with a PCR-restriction fragment length polymorphism assay which amplified the entire iam locus showed a HindIII restriction fragment polymorphism pattern associated mainly with invasive strains. Although cluster analysis of the RAPD fingerprinting showed genetic diversity among strains, two main clusters were identified. Cluster I comprised significantly more pathogenic and invasive isolates, while cluster II grouped the majority of nonpathogenic, noninvasive isolates. These data indicate that most of the invasive Campylobacter strains could be differentiated from noninvasive isolates by RAPD analysis and PCR using specific primers that amplify a fragment of the iam locus. PMID- 11283057 TI - Comparison of the E-test with the NCCLS M38-P method for antifungal susceptibility testing of common and emerging pathogenic filamentous fungi. AB - The National Committee for Clinical Laboratory Standards (NCCLS) M38-P method describes standard parameters for testing the fungistatic antifungal activities (MICs) of established agents against filamentous fungi (molds). The present study evaluated the in vitro fungistatic activities of itraconazole and amphotericin B by the E-test and the NCCLS M38-P microdilution method against 186 common and emerging pathogenic molds (123 isolates of Aspergillus spp. [five species], 16 isolates of Fusarium spp. [two species], 4 Paecilomyces lilacinus isolates, 5 Rhizopus arrhizus isolates, 15 Scedosporium spp., 18 dematiaceous fungi, and 5 Trichoderma longibrachiatum isolates). The agreement between the methods for amphotericin B MICs ranged from 70% for Fusarium solani to > or =90% for most of the other species after the first reading; agreement was dependent on both the incubation time and the species being evaluated. Major discrepancies between the amphotericin B MICs determined by the E-test and the NCCLS M38-P method were demonstrated for three of the five species of Aspergillus tested and the two species of Fusarium tested. This discrepancy was more marked after 48 h of incubation; the geometric mean MICs determined by the E-test increased between 24 and 48 h from between 1.39 and 3.3 microg/ml to between 5.2 and >8 microg/ml for Aspergillus flavus, Aspergillus fumigatus, and Aspergillus nidulans. The agreement between the itraconazole MICs determined by the E-test and the NCCLS M38-P method ranged from 83.3% for A. nidulans to > or =90% for all the other species tested; the overall agreement was higher (92.7%) than that for amphotericin B (87.9%). The agreement was less dependent on the incubation time. Clinical trials need to be conducted to establish the role of the results of either the E-test or the NCCLS M38-P method in vitro for molds with the two agents as predictors of clinical outcome. PMID- 11283058 TI - Chlamydial serology: comparative diagnostic value of immunoblotting, microimmunofluorescence test, and immunoassays using different recombinant proteins as antigens. AB - To improve the reliability of the serodiagnosis of Chlamydia trachomatis infections, an immunoblot analysis, a microimmunofluorescence titration, and different immunoassays using synthetic peptides derived from species-specific epitopes in variable domain IV of the major outer membrane protein or recombinant antigens (heat shock protein 70 [hsp70], hsp60, hsp10, polypeptide encoded by open reading frame 3 of the plasmid [pgp3], macrophage infectivity potentiator, and a fragment of the total lipopolysaccharide) were evaluated. Because cross reactions between chlamydial species have been reported, the microimmunofluorescence tests were also performed with Chlamydia pneumoniae and Chlamydia psittaci used as antigens, and C. pneumoniae-specific antibodies were also determined by immunoassays. Since the presence of antimicrobial antibodies must be interpreted in light of their prevalence in the general population, responses obtained with serum samples from patients with well-defined infection (i.e., with positive urethral or endocervical C. trachomatis DNA amplification) were compared to those obtained with samples from healthy blood donors. The best sensitivity (86%) with a specificity of 81% was obtained for immunoblotting results, when the number of individuals with > or =10 immunoglobulin G (IgG) and/or > or =2 IgM responses to the different C. trachomatis antigens was considered. A 13-kDa antigen was recognized by most of the samples (86% for IgG) from patients with acute urogenital infection but rarely (3%) by those from healthy blood donors (P < 0.0001). The sensitivity and specificity results obtained for serum antibodies to peptides or recombinant antigens were slightly lower than those results obtained for the number of responses to whole C. trachomatis antigens, which were 76 and 77%, respectively, when IgG responses to both recombinant hsp60 and pgp3 were considered. PMID- 11283059 TI - Single rapid real-time monitored isothermal RNA amplification assay for quantification of human immunodeficiency virus type 1 isolates from groups M, N, and O. AB - Because human immunodeficiency virus type 1 (HIV-1) subtypes and circulating recombinant forms (CRFs) are spreading rapidly worldwide and are becoming less confined to a geographical area, RNA assays that can detect and quantify all HIV 1 isolates reliably are in demand. We have developed a fast, real-time monitored RNA assay based on an isothermal nucleic acid sequence-based amplification technology that amplifies a part of the long terminal repeat region of the HIV-1 genome. Real-time detection was possible due to the addition of molecular beacons to the amplification reaction that was monitored in a fluorimeter with a thermostat. The lower level of detection of the assay was 10 HIV-1 RNA molecules per reaction, and the lower level of quantification was 100 copies of HIV-1 RNA with a dynamic range of linear quantification between 10(2) and 10(7) RNA molecules. All HIV-1 groups, subtypes, and CRFs could be detected and quantified with equal efficiency, including the group N isolate YBF30 and the group O isolate ANT70. To test the clinical utility of the assay, a series of 62 serum samples containing viruses that encompassed subtypes A through G and CRFs AE and AG of HIV-1 group M were analyzed, and these results were compared to the results of a commercially available assay. This comparison showed that the quantification results correlated highly (R(2) = 0.735) for those subtypes that could be well quantified by both assays (subtypes B, C, D, and F), whereas improved quantification was obtained for subtypes A and G and CRFs AE and AG. A retrospective study with six individuals infected with either a subtype A, B, C, or D or an AG isolate of HIV-1 group M, who were treated with highly active antiretroviral therapy, revealed that the assay was well suited to the monitoring of therapy effects. In conclusion, the newly developed real-time monitored HIV-1 assay is a fast and sensitive assay with a large dynamic range of quantification and is suitable for quantification of most if not all subtypes and groups of HIV 1. PMID- 11283060 TI - Development of a genomics-based PCR assay for detection of Mycoplasma pneumoniae in a large outbreak in New York State. AB - A genomics-based PCR method was developed and used to test specimens from patients involved in a large outbreak of Mycoplasma pneumoniae in a closed religious community in New York State. New P1 adhesin gene primers were designed to bind to 9 of 10 target sequences in the repetitive-element sequences obtained from the whole genome sequence of M. pneumoniae. This PCR method had a sensitivity of 0.006 CFU and a specificity of 100% for M. pneumoniae. The PCR was validated by testing a subset of patient samples by culture and comparing the results to those obtained by PCR. Of the initial 280 samples tested, 73 were positive by PCR and 22 were positive by culture. All samples positive by culture were also positive by PCR. Follow-up testing of selected patients 3 to 6 weeks after antibiotic treatment revealed that eight samples remained positive by PCR and that three samples remained positive by culture. Additionally, no nonspecific PCR inhibition was detected as a result of the specimen type, transport medium, or sample preparation methodology. The study demonstrates that the PCR described here is a rapid, sensitive, and specific method for the identification of M. pneumoniae and was helpful for the detection and monitoring of the outbreak. PMID- 11283061 TI - Analysis of Clostridium difficile isolates from nosocomial outbreaks at three hospitals in diverse areas of Japan. AB - Clostridium difficile isolates recovered from patients with C. difficile associated diarrhea (CDAD) at three hospitals located in diverse areas of Japan were analyzed by three typing systems, PCR ribotyping, pulsed-field gel electrophoresis (PFGE), and Western immunoblotting. At the three hospitals examined, a single PCR ribotype strain (type smz) was predominant and accounted for 22 (65%) of 34, 18 (64%) of 28, and 11 (44%) of 25 isolates, respectively. All of the 51 isolates that represented PCR ribotype smz were nontypeable by PFGE because of DNA degradation. Since the type smz strain did not react with any of the antisera against 10 different serogroups (A, B, C, D, F, G, H, I, K, and X), we prepared a new antiserum against a type smz isolate. All 51 type smz isolates presented identical banding patterns, reacting with the newly prepared antiserum (designated subserogroup JP-0 of serogroup JP). These results were compared with those of a strain from a hospital outbreak that occurred in New York, which has been identified as type J9 by restriction enzyme analysis and type 01/A by arbitrarily primed PCR but was nontypeable by PFGE because of DNA degradation. This strain was reported to be epidemic at multiple hospitals in the United States. The J9 strain represented a PCR ribotype pattern different from that of a type smz strain and was typed as subserogroup G-1 of serogroup G by immunoblot analysis. A single outbreak type causing nosocomial CDAD in Japan was found to be different from the strain causing multiple outbreaks in the United States, even though the outbreak strains from the two countries were nontypeable by PFGE because of DNA degradation. PMID- 11283062 TI - Detection of phospholipase C in nontuberculous mycobacteria and its possible role in hemolytic activity. AB - Phospholipase C plays a key role in the pathogenesis of several bacterial infections, for example, those caused by Clostridium perfringens and Listeria monocytogenes. Previous studies have reported multiple copies of plc genes homologous to Pseudomonas aeruginosa plcH and plcN genes encoding the hemolytic and nonhemolytic phospholipase C enzymes in the genomes of Mycobacterium tuberculosis, M. marinum, M. bovis, and M. ulcerans. In this study we analyzed the possible relationship between phospholipase C and hemolytic activity in 21 strains of nontuberculous mycobacteria representing nine different species. Detection of phospholipase C enzymatic activity was carried out using thin-layer chromatography to detect diglycerides in the hydrolysates of radiolabeled phosphatidylcholine. DNA sequences of M. kansasii and M. marinum homologous to the genes encoding phospholipase C from M. tuberculosis and M. ulcerans were identified by DNA-DNA hybridization and sequencing. Finally, we developed a direct and simple assay to detect mycobacterial hemolytic activity. This assay is based on a modified blood agar medium that allows the growth and expression of hemolysis of slow-growing mycobacteria. Hemolytic activity was detected in M. avium, M. intracellulare, M. ulcerans, M. marinum, M. tuberculosis, and M. kansasii mycobacteria with phospholipase C activity, but not in M. fortuitum. No hemolytic activity was detected in M. smegmatis, M. gordonae, and M. vaccae. Whether or not phospholipase C enzyme plays a role in the pathogenesis of nontuberculous mycobacterial diseases needs further investigation. PMID- 11283063 TI - Are two Cryptococcus neoformans strains epidemiologically linked? AB - The aim of this study was to standardize a method to determine whether two strains of Cryptococcus neoformans could be considered epidemiologically linked. We hypothesized that strains isolated from the same patient were epidemiologically linked and that those isolated from different patients were unrelated. We used 17 environmental isolates and 97 clinical isolates from 31 patients diagnosed with cryptococcosis (1 to 14 isolates per patient). Using the plasmid pCnTel-1-labeled probe CENTEL, we were able to differentiate some unrelated strains that yielded the same hybridization profile with the C. neoformans middle-repetitive-element CNRE-1 probe. The genetic distances separating the strains isolated from the same patient and those separating the strains isolated from different patients were estimated, and the results obtained with the two probes were compared. Analysis of the results enabled the calculation of two Dice coefficient limits defining the zones containing the pairs of linked strains and the pairs of unrelated strains, as well as an intermediate uncertainty zone for which it was not possible to establish whether the pairs of strains were linked. PMID- 11283064 TI - Pediatric solid-organ transplant recipients carry chronic loads of Epstein-Barr virus exclusively in the immunoglobulin D-negative B-cell compartment. AB - Solid-organ transplant recipients are at risk for development of lymphoproliferative diseases. The purpose of this study was to examine the distribution of Epstein-Barr virus (EBV) load in the peripheral blood of pediatric transplant recipients who had become chronic viral load carriers (>8 copies/10(5) lymphocytes for >2 months). A total of 19 patients with viral loads ranging from 20 to 5,000 viral genome copies/10(5) lymphocytes were studied. Ten patients had no previous diagnosis of posttransplant lymphoproliferative disease (PT-LPD), while nine had recovered from a diagnosed case of PT-LPD. No portion of the peripheral blood viral load was detected in the cell-free plasma fraction. Viral DNA was found in a population of cells characterized as CD19(hi) and immunoglobulin D negative, a phenotype that is consistent with the virus being carried exclusively in the memory B-cell compartment of the peripheral blood. There was no difference in the compartmentalization based upon either the level of the viral load or the past diagnosis of an episode of PT-LPD. These results have implications for the design of tests to detect EBV infection and for the interpretation and use of positive EBV PCR assays in the management of transplant recipients. PMID- 11283065 TI - Biotyping of Penicillium marneffei reveals concentration-dependent growth inhibition by galactose. AB - Thirty-two isolates of the dimorphic fungus Penicillium marneffei were studied for their biochemical properties. All isolates possessed the enzyme urease and were inhibited by 500 mg of cycloheximide per liter. No strain fermented glucose, and thus no strain fermented any of the other five sugars tested. All assimilated glucose, maltose, and cellobiose; only one of the isolates did not assimilate salicin. Totals of 65.6, 84.4, and 71.9% of the isolates assimilated trehalose, xylose, and nitrate, respectively. Twelve strains possessed the enzyme beta galactosidase. Overall, 17 different biotypes were recognized, but no association was found between the human immunodeficiency virus status of the patients and the biotype. A novel finding of concentration-dependent growth inhibition of P. marneffei by galactose is described. Inhibition of growth occurred at a low concentration of galactose (0.015 to 0.25%) when galactose was the sole carbon source in the medium. Morphological changes of the fungal cells were observed in the presence of galactose. PMID- 11283066 TI - Fluconazole and voriconazole multidisk testing of Candida species for disk test calibration and MIC estimation. AB - Fluconazole and voriconazole MICs were determined for 114 clinical Candida isolates, including isolates of Candida albicans, Candida glabrata, Candida krusei, Candida lusitaniae, Candida parapsilosis, and Candida tropicalis. All strains were susceptible to voriconazole, and most strains were also susceptible to fluconazole, with the exception of C. glabrata and C. krusei, the latter being fully fluconazole resistant. Single-strain regression analysis (SRA) was applied to 54 strains, including American Type Culture Collection reference strains. The regression lines obtained were markedly different for the different Candida species. Using an MIC limit of susceptibility to fluconazole of < or =8 microg/ml, according to NCCLS standards, the zone breakpoint for susceptibility for the 25-microg fluconazole disk was calculated to be > or =18 mm for C. albicans and > or =22 mm for C. glabrata and C. krusei. SRA results for voriconazole were used to estimate an optimal disk content according to rational criteria. A 5-microg disk content of voriconazole gave measurable zones for a tentative resistance limit of 4 microg/ml, whereas a 2.5-microg disk gave zones at the same MIC level for only three of the species. A novel SRA modification, multidisk testing, was also applied to the two major species, C. albicans and C. glabrata, and the MIC estimates were compared with the true MICs for the isolates. There was a significant correlation between the two measurements. Our results show that disk diffusion methods might be useful for azole testing of Candida isolates. The method can be calibrated using SRA. Multidisk testing gives direct estimations of the MICs for the isolates. PMID- 11283067 TI - Evaluation of the NucliSens Basic Kit for detection of Chlamydia trachomatis and Neisseria gonorrhoeae in genital tract specimens using nucleic acid sequence based amplification of 16S rRNA. AB - We evaluated a new RNA amplification and detection kit, the NucliSens Basic Kit (Organon Teknika), for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae in genitourinary specimens. The Basic Kit provides an open platform for RNA amplification and detection and contains isolation reagents for nucleic acid extraction, nucleic acid sequence-based amplification (NASBA) reagents (enzymes and buffers), and a generic ruthenium-labeled probe for electrochemiluminescent (ECL) detection of amplified product. Using freshly purified and titrated stocks of C. trachomatis and N. gonorrhoeae and in vitro generated RNA transcripts for sensitivity determinations, the Basic Kit detected 1 inclusion-forming unit of C. trachomatis, 1 CFU of N. gonorrhoeae, and 100 RNA molecules of 16S rRNA for both bacteria. The clinical performance of the Basic Kit was evaluated by testing a total of 250 specimens for N. gonorrhoeae by culture and NASBA and a total of 96 specimens for C. trachomatis by PCR and NASBA. The Basic Kit detected 139 of 142 N. gonorrhoeae culture-positive specimens and gave a negative result for 73 of 74 culture-negative specimens, for a sensitivity and specificity of 97.9 and 98.7%, respectively. For C. trachomatis, the Basic Kit detected 24 of 24 PCR-positive specimens and gave a negative result for 71 of 72 PCR-negative specimens, for a sensitivity and specificity of 100 and 98.6%, respectively. The Basic Kit also detected specimens containing both N. gonorrhoeae and C. trachomatis, using a multiplex NASBA assay using primers for both bacteria. The NucliSens Basic Kit offers a versatile platform for the development of sensitive RNA detection assays for sexually transmitted diseases. PMID- 11283068 TI - Application and evaluation of the interlaboratory reproducibility of tRNA intergenic length polymorphism analysis (tDNA-PCR) for identification of Streptococcus species. AB - The discriminatory power, speed, and interlaboratory reproducibility of tRNA intergenic length polymorphism analysis (tDNA-PCR) combined with capillary electrophoresis was evaluated for the identification of streptococci. This method was carried out in three different laboratories under highly standardized conditions for 54 strains belonging to 18 different species. It was concluded that interlaboratory reproducibility of tDNA fingerprints produced by means of capillary electrophoresis was sufficiently high to permit the exchange between different laboratories and the construction of common libraries which can be consulted for comparison with fingerprints obtained independently in separate laboratories. In a second step, 17 other species were included in the study and examined in one of the participating laboratories. All Streptococcus species studied, except S. mitis, S. oralis, S. parasanguinis, S. pneumoniae, S. thermophilus, and S. vestibularis, showed distinguishable tDNA fingerprints. A database of well-characterized strains was constructed to enable computer-aided identification of unknown streptococcal isolates. PMID- 11283069 TI - Use of a LightCycler gyrA mutation assay for rapid identification of mutations conferring decreased susceptibility to ciprofloxacin in multiresistant Salmonella enterica serotype Typhimurium DT104 isolates. AB - A LightCycler-based PCR-hybridization gyrA mutation assay (GAMA) was developed to rapidly detect gyrA point mutations in multiresistant (MR) Salmonella enterica serotype Typhimurium DT104 with decreased susceptibility to ciprofloxacin (MIC, 0.25 to 1.0 mg/liter). Ninety-two isolates (49 human, 43 animal) were tested with three individual oligonucleotide probes directed against an Asp-87-to-Asn (GAC- >AAC) mutation, an Asp-87-to-Gly (GAC-->GGC) mutation, and a Ser-83-to-Phe (TCC- >TTC) mutation. Strains homologous to the probes could be distinguished from strains that had different mutations by their probe-target melting temperatures. Thirty-seven human and 30 animal isolates had an Asp-87-to-Asn substitution, 6 human and 6 animal isolates had a Ser-83-to-Phe substitution, and 5 human and 2 animal isolates had an Asp-87-to-Gly substitution. The remaining six strains all had mismatches with the three probes and therefore different gyrA mutations. The sequencing of gyrA from these six isolates showed that one human strain and two animal strains had an Asp-87-to-Tyr (GAC-->TAC) substitution and two animal strains had a Ser-83-to-Tyr (TCC-->TAC) substitution. One animal strain had no gyrA mutation, suggesting that this isolate had a different mechanism of resistance. Fifty-eight of the strains tested were indistinguishable by several different typing methods including antibiograms, pulsed-field gel gel electrophoresis, and plasmid profiling, although they could be further subdivided according to gyrA mutation. This study confirmed that MR DT104 with decreased susceptibility to ciprofloxacin from humans and food animals in England and Wales may have arisen independently against a background of clonal spread of MR DT104. PMID- 11283070 TI - Definition of a divergent epitope that allows differential detection of early protein p41 from human herpesvirus 6 variants A and B. AB - The human herpesvirus 6 (HHV-6) early protein, p41, encoded by the U27 gene has been detected in oligodendrocytes of multiple sclerosis (MS) patients by using a monoclonal antibody (MAb to p41/38). We here report the antigenic epitope of HHV 6 p41 recognized by this MAb. First, we established that the MAb to p41/38 recognizes a nuclear antigen in HHV-6A strain GS-infected cells but not in HHV-6B strain Z29-infected cells. Secondly, we compared the reactivity of the MAb to p41/38 to that of another p41-specific MAb (MAb to p41) on immunoblots with purified p41-glutathione S-transferase fusion protein from strains GS and Z29 and GS- and Z29-infected-cell lysates. The two MAbs were tested in an enzyme-linked immunosorbent assay against a panel of synthetic peptides covering the amino acid substitutions between the GS- and Z29-derived p41 proteins, as determined by DNA sequencing of our cloned isolates of the U27 gene. The MAb to p41/38 reacted specifically with a peptide comprising p41 residues 321 to 340 from strain GS. The critical residue in this peptide was serine 328, as the substitution S328N in the Z29 strain rendered the corresponding peptide nonreactive. The p41 S328 marker was present in three of three HHV-6A strains, while four of four sequenced p41 genes from HHV-6B strains had N328. Our findings are of value for the interpretation of previous findings of p41 expression in brains of MS patients and may allow a more detailed analysis of the role of HHV-6 variants in other disorders. PMID- 11283071 TI - Monitoring the emergence of hepatitis B virus polymerase gene variants during lamivudine therapy using the LightCycler. AB - Treatment of chronic hepatitis B virus (HBV) infection with lamivudine is associated with the appearance in the circulation of HBV variants with mutations in the YMDD (tyrosine, methionine, aspartate, aspartate) motif of the polymerase gene. Fluorometric real-time PCR with the LightCycler assay was used for the detection of resistant variants. Differences in the hybridization melting curve kinetics of probes bound to the sequences encoding the wild-type or the mutant YMDD motifs (YIDD or YVDD in which the methionine residue is altered to an isoleucine or a valine, respectively) distinguished the single-base changes responsible for the resistance phenotype. The LightCycler probe hybridization assay was applied to 40 serum specimens from 19 patients, and the results were correlated with the nucleotide sequences determined for the corresponding PCR products. All three variants could be identified in the specimens. PCR clones obtained from four patients early in the course and prior to lamivudine therapy were investigated for the appearance of YIDD and YVDD variants with the LightCycler assay. In one patient, a transient appearance of the YIDD variant was observed 6 weeks into therapy. Subsequently, after 11 months of lamivudine therapy, the YVDD variant emerged in that patient. PMID- 11283072 TI - Molecular epidemiology of Streptococcus uberis isolates from dairy cows with mastitis. AB - Pulsed-field gel electrophoresis and antimicrobial sensitivity testing were used as tools to investigate the epidemiology of Streptococcus uberis mastitis in dairy cows. A total of 62 different strains were found among 138 isolates from the four herds investigated, and between 10 and 26 different strains were found in each herd. There was no strain common to all four herds. Identical strains of S. uberis were detected from different quarters of individual cows and from cows within the same herd, suggesting that transmission from quarter to quarter and cow to cow had occurred. Despite the great variation in S. uberis strains, persistent infection with the same strain within a lactation was observed in most cows. Predominant strains were present in two herds. Preliminary investigations could not clarify why these particular strains might predominate, but in one herd there was a significant difference between the prevalence of clinical mastitis in quarters infected with the predominant strain and that in quarters infected with other strains, suggesting the greater virulence of the predominant strain. The wide variety of S. uberis strains found is consistent with an environmental source of S. uberis. However, evidence of direct transmission, the persistence of infection, and the predominance of particular strains in some herds indicate that S. uberis infections are epidemiologically complex and that the relative importance of these factors in the occurrence of mastitis may differ between herds. PMID- 11283073 TI - Tsukamurella strandjordae sp. nov., a proposed new species causing sepsis. AB - We have isolated a gram-positive, weakly acid-alcohol-fast, irregular rod-shaped bacterium from cultures of blood from a 5-year-old girl with acute myelogenous leukemia. This isolate was compared with 14 other strains including reference strains of Tsukamurella species by a polyphasic approach based on physiological and biochemical properties, whole-cell short-chain fatty acid and mycolic acid analyses, DNA-DNA hybridization, and sequencing of the 16S rRNA gene. This isolate represents a new taxon within the genus Tsukamurella for which we propose the name Tsukamurella strandjordae sp. nov. Our study also revealed that Tsukamurella paurometabola ATCC 25938 represents a misnamed Tsukamurella inchonensis isolate and confirms that Tsukamurella wratislaviensis belongs to the genus Rhodococcus. PMID- 11283074 TI - Resistance of Pseudomonas aeruginosa isolates to hydrogel contact lens disinfection correlates with cytotoxic activity. AB - One of the most common pathogens in infection of hydrogel contact lens wearers is Pseudomonas aeruginosa, which can gain access to the eye via contamination of the lens, lens case, and lens care solutions. Only one strain per species is used in current regulatory testing for the marketing of chemical contact lens disinfectants. The aim of this study was to determine whether P. aeruginosa strains vary in their susceptibility to hydrogel contact lens disinfectants. A method for rapidly screening bacterial susceptibility to contact lens disinfectants was developed, based on measurement of the MIC. The susceptibility of 35 P. aeruginosa isolates to two chemical disinfectants was found to vary among strains. MICs ranged from 6.25 to 100% for both disinfectants at 37 degrees C, and a number of strains were not inhibited by a 100% disinfectant concentration in the lens case environment at room temperature (22 degrees C). Resistance to disinfection appeared to be an inherent rather than acquired trait, since some resistant strains had been isolated prior to the introduction of the disinfectants and some susceptible P. aeruginosa strains could not be made more resistant by repeated disinfectant exposure. A number of P. aeruginosa strains which were comparatively more resistant to short-term disinfectant exposure also demonstrated the ability to grow to levels above the initial inoculum in one chemical disinfectant after long-term (24 to 48 h) disinfectant exposure. Resistance was correlated with acute cytotoxic activity toward corneal epithelial cells and with exsA, which encodes a protein that regulates cytotoxicity via a complex type III secretion system. These results suggest that chemical disinfection solutions may select for contamination with cytotoxic strains. Further investigation of the mechanisms and factors responsible for resistance may also lead to strategies for reducing adverse responses to contact lens wear. PMID- 11283075 TI - Expression and self-assembly in baculovirus of porcine enteric calicivirus capsids into virus-like particles and their use in an enzyme-linked immunosorbent assay for antibody detection in swine. AB - Porcine enteric calicivirus (PEC) causes diarrhea and intestinal lesions in pigs. PEC strain Cowden grows to low to moderate titers in cell culture but only with the addition of intestinal contents from uninfected gnotobiotic pigs (W. T. Flynn and L. J. Saif, J. Clin. Microbiol. 26:206--212, 1988; A. V. Parwani, W. T. Flynn, K. L. Gadfield, and L. J. Saif, Arch. Virol. 120:115--122, 1991). Cloning and sequence analysis of the PEC Cowden full-length genome revealed that it is most closely related genetically to the human Sapporo-like viruses. In this study, the complete PEC capsid gene was subcloned into the plasmid pBlueBac4.5 and the recombinant baculoviruses were identified by plaque assay and PCR. The PEC capsid protein was expressed in insect (Sf9) cells inoculated with the recombinant baculoviruses, and the recombinant capsid proteins self- assembled into virus-like particles (VLPs) that were released into the cell supernatant and purified by CsCl gradient centrifugation. The PEC VLPs had the same molecular mass (58 kDa) as the native virus capsid and reacted with pig hyperimmune and convalescent-phase sera to PEC Cowden in enzyme-linked immunosorbent assay (ELISA) and Western blotting. The PEC capsid VLPs were morphologically and antigenically similar to the native virus by immune electron microscopy. High titers (1:102,400 to 204,800) of PEC-specific antibodies were induced in guinea pigs inoculated with PEC VLPs, suggesting that the VLPs could be useful for future candidate PEC vaccines. A fixed-cell ELISA and VLP ELISA were developed to detect PEC serum antibodies in pigs. For the fixed-cell ELISA, Sf9 cells were infected with recombinant baculoviruses expressing PEC capsids, followed by cell fixation with formalin. For the VLP ELISA, the VLPs were used for the coating antigen. Our data indicate that both tests were rapid, specific, and reproducible and might be used for large-scale serological investigations of PEC antibodies in swine. PMID- 11283077 TI - Evaluation of mycobacteria growth indicator tube for direct and indirect drug susceptibility testing of Mycobacterium tuberculosis from respiratory specimens in a Siberian prison hospital. AB - The manual Mycobacteria Growth Indicator Tube (MGIT) method was evaluated for performing direct and indirect drug susceptibility testing (DST) of Mycobacterium tuberculosis for isoniazid and rifampin on 101 strongly smear-positive sputum specimens in a Siberian prison hospital. Using the indirect method of proportion (MOP) as the "gold standard," the accuracies of isoniazid and rifampin susceptibility testing by the direct MGIT system were 97.0 and 94.1%, respectively. The accuracy of the indirect MGIT system was 98.0% for both drugs. The turnaround times from specimen processing to reporting of the DST results ranged between 4 and 23 (mean, 9.2) days by the direct MGIT method, 9 and 30 (mean, 15.3) days by the indirect MGIT method, and 26 and 101 (mean, 59.6) days by the indirect MOP. MGIT appears to be a reliable, rapid, and convenient method for performing direct and indirect DSTs in low-resource settings, but further studies are required to refine the direct DST protocol. Cost is the only factor prohibiting widespread implementation of MGIT. PMID- 11283076 TI - Development of an immunoassay for rapid detection of ganglioside GM(1) mimicry in Campylobacter jejuni strains. AB - Mimicry of peripheral nerve gangliosides by Campylobacter jejuni lipopolysaccharides (LPSs) has been proposed to induce cross-reacting antiganglioside antibodies in Guillain-Barre syndrome (GBS). Because current methods for LPS characterization are labor-intensive and inhibit the screening of large numbers of strains, a rapid GM(1) epitope screening assay was developed. Biomass from two agar plates of confluent growth yielded sufficient LPS using a novel phenol-water and ether extraction procedure. Extracts of LPS were reacted with cholera toxin (GM(1) ligand), peanut agglutinin (Gal beta1-->3GalNAc ligand), and anti-GM(1) antibodies. After the assay was validated, 12 of 59 (20%) C. jejuni serostrains, including four serotypes that have not previously been associated with GBS, reacted with two or more anti-GM(1) ganglioside reagents. Subsequently, LPS extracts from 5 of 7 (71%) C. jejuni isolates and 2 of 3 (67%) C. jejuni culture collection strains bore GM(1) structures. Overall, the assay system was reliable, efficient, and reproducible and may be adapted for large scale epidemiological studies. PMID- 11283078 TI - Diagnosis and monitoring of murine histoplasmosis by a nested PCR assay. AB - A newly developed nested PCR assay was applied to murine models of histoplasmosis. ICR and BALB/c mice were intravenously infected with Histoplasma capsulatum and sacrificed up to 29 days later. Samples of blood, spleen, and lung homogenates were cultured and examined by the PCR assay. In the ICR mouse model, 265 of 319 organ samples showed concordant results. With 7 samples, the culture was positive and the PCR assay was negative whereas a positive PCR but a negative culture were obtained with 47 samples (P < 0.0001 according to McNemar's test). Organ homogenates and blood samples of either spontaneously cured or treated BALB/c mice were PCR negative. The nested PCR assay performs excellently in the monitoring of spontaneously and treatment-cured murine histoplasmosis. It limits the infection risks of the laboratory staff and might be of diagnostic value for humans. PMID- 11283079 TI - Specific detection and prevalence of Helicobacter heilmannii-like organisms in the human gastric mucosa by fluorescent in situ hybridization and partial 16S ribosomal DNA sequencing. AB - Gastric infection with Helicobacter heilmannii (previously known as Gastrospirillum hominis) is invariably linked with the presence of chronic gastritis and the risk of developing low-grade mucosa-associated lymphoid tissue lymphoma in humans. In contrast to Helicobacter pylori, various H. heilmannii species colonize the stomachs of domestic animals, which might be a reservoir for transmission to humans (zoonosis). To identify the number and prevalence of different H. heilmanni types in humans, we analyzed 89 gastric biopsy samples histologically identified as H. heilmannii positive by fluorescence in situ hybridization. Of these gastric specimens, 84 (94.4%) contained a single H. heilmannii type. In five samples, however, two different H. heilmannii types were detected. The most prevalent species in monoinfected samples is H. heilmannii type 1, found in 78.5% (66 of 84) of the specimens, followed by a novel H. heilmannii-like organism (HHLO), HHLO type 4, identified in 9.6% (8 of 84) of tissue sections. H. heilmannii type 2 and a further HHLO type not described before, type 3, were found in 8.3% (7 of 84) and 1.2% (1 of 84) of the monoinfected samples, respectively. Additionally, HHLO type 5 with a 16S ribosomal DNA sequence identical to that of Helicobacter salomonis was found with a prevalence of 2.4% (2 of 89). Thirteen of these biopsy samples were also investigated by a PCR approach developed for this study that allows a Helicobacter-specific amplification of a variable portion of the 16S rRNA gene and subsequent sequencing. In total, five different types of HHLOs could be identified within these samples. We conclude that humans can be infected by at least five different HHLO types, which presumably have their origin in animal species like dogs, cats, and pigs. PMID- 11283080 TI - Paratuberculosis infection of nonruminant wildlife in Scotland. AB - Recent reports of natural paratuberculosis (or Johne's disease) in rabbits, foxes, and stoats has focused debate on the presence and importance of wildlife reservoirs in the epidemiology of this disease. This paper describes an extensive study investigating 18 nonruminant wildlife species for evidence of paratuberculosis. Using both culture and histopathological analysis, fox, stoat, weasel, crow, rook, jackdaw, rat, wood mouse, hare, and badger were found to harbor Mycobacterium avium subsp. paratuberculosis, the causative organism of paratuberculosis, suggesting that the epidemiology of this disease is more complex than previously realized. PMID- 11283081 TI - High degree of interlaboratory reproducibility of human immunodeficiency virus type 1 protease and reverse transcriptase sequencing of plasma samples from heavily treated patients. AB - We assessed the reproducibility of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) and protease sequencing using cryopreserved plasma aliquots obtained from 46 heavily treated HIV-1-infected individuals in two laboratories using dideoxynucleotide sequencing. The rates of complete sequence concordance between the two laboratories were 99.1% for the protease sequence and 99.0% for the RT sequence. Approximately 90% of the discordances were partial, defined as one laboratory detecting a mixture and the second laboratory detecting only one of the mixture's components. Only 0.1% of the nucleotides were completely discordant between the two laboratories, and these were significantly more likely to occur in plasma samples with lower plasma HIV-1 RNA levels. Nucleotide mixtures were detected at approximately 1% of the nucleotide positions, and in every case in which one laboratory detected a mixture, the second laboratory either detected the same mixture or detected one of the mixture's components. The high rate of concordance in detecting mixtures and the fact that most discordances between the two laboratories were partial suggest that most discordances were caused by variation in sampling of the HIV-1 quasispecies by PCR rather than by technical errors in the sequencing process itself. PMID- 11283082 TI - A combination of two genetic markers is sufficient for restriction fragment length polymorphism typing of Mycobacterium tuberculosis complex in areas with a high incidence of tuberculosis. AB - The incidence of tuberculosis (TB) in Madagascar is 150 cases per 100,000 people. Because of this endemicity, we studied the genetic diversity of Mycobacterium tuberculosis strains isolated in four big cities in 1994 to 1995 with the aim of monitoring TB transmission. Isolates from 316 cases of pulmonary TB (PTM(+)) were typed by Southern hybridization with genetic markers IS6110 and DR. Of the 316 PTM(+) strains, 66 (20.8%) had a single IS6110 band and were differentiated by the DR marker into 33 profiles. Using both markers, 37.7% (119) of the patients were clustered, a proportion similar to that in countries with a high prevalence of TB. There was no significant difference between clustered and nonclustered patients in age, sex, Mycobacterium bovis BCG status, and drug susceptibility of strains. Clustering was significantly greater in the capital, Antananarivo, than in the other cities, suggesting a higher rate of transmission. However, most of the patients in clusters were living in different areas, and, within a distance of 0.7 km, we did not find epidemiologically unrelated strains with the same restriction fragment length polymorphism profile. Despite an apparently low polymorphism, genetic markers such as IS6110 are potentially valuable for monitoring TB transmission. However, the high proportion of Malagasy isolates with a single IS6110 copy makes this marker alone unsuitable for typing. Additional markers such as DR are necessary for the differentiation of the isolates and for epidemiological surveys. PMID- 11283083 TI - Molecular characteristic-based epidemiology of hepatitis B, C, and E viruses and GB virus C/hepatitis G virus in Myanmar. AB - We carried out a molecular characteristic-based epidemiological survey of various hepatitis viruses, including hepatitis B virus (HBV), hepatitis C virus (HCV), hepatitis E virus (HEV), and GB virus C (GBV-C)/hepatitis G virus (HGV), in Myanmar. The study population of 403 subjects consisted of 213 healthy individuals residing in the city of Yangon, Myanmar, and the surrounding suburbs and 190 liver disease patients (155 virus-related liver disease patients and 35 nonviral disease patients). The infection rates of the viruses among the 213 healthy subjects were as follows: 8% for HBV (16 patients), 2% for HCV (4 patients), and 8% for GBV-C/HGV (17 patients). In contrast, for 155 patients with acute hepatitis, chronic hepatitis, liver cirrhosis, or hepatocellular carcinoma, the infection rates were 30% for HBV (46 patients), 27% for HCV (41 patients), and 11% for GBV-C/HGV (17 patients). In the nonviral liver disease group of 35 patients with alcoholic liver disease, fatty liver, liver abscess, and biliary disease, the infection rates were 6% for HBV (2 patients), 20% for HCV (7 patients), and 26% for GBV-C/HGV (9 patients). The most common viral genotypes were type C of HBV (77%), type 3b of HCV (67%), and type 2 of GBV-C/HGV (67%). Moreover, testing for HEV among 371 subjects resulted in the detection of anti HEV immunoglobulin G (IgG) in 117 patients (32%). The age prevalence of anti-HEV IgG was 3% for patients younger than 20 years and 30% or more for patients 20 years of age or older. Furthermore, a high prevalence of anti-HEV IgG (24%) was also found in swine living together with humans in Yangon. These results suggest that these hepatitis virus infections are widespread in Myanmar and have led to a high incidence of acute and chronic liver disease patients in the region. PMID- 11283084 TI - Identification and characterization of phage variants of a strain of epidemic methicillin-resistant Staphylococcus aureus (EMRSA-15). AB - EMRSA-15 is one of the most important strains of epidemic methicillin-resistant Staphylococcus aureus (EMRSA) found in the United Kingdom. It was originally characterized by weak lysis with phage 75 and production of enterotoxin C but not urease. Two variant strains of EMRSA-15 which show a broader phage pattern than the progenitor strain have emerged. A total of 153 recent clinical isolates representing classical EMRSA-15 (55 isolates) or these phage variants (98 isolates) were compared by SmaI macrorestriction profiles in pulsed-field gel electrophoresis (PFGE) as well as by urease and enterotoxin C production. Eight of the 98 isolates were shown to be other unrelated strains by both PFGE and their production of urease, a misidentification rate of 8% by phage typing. Seventy-one EMRSA-15 isolates were enterotoxin C negative, and the majority of these were sensitive to phage 81. Examination of PFGE profiles and Southern blotting studies suggest that the enterotoxin C gene locus is encoded on a potentially mobile DNA segment of ca. 15 kb. After elimination of the eight non EMRSA-15 isolates, the remaining 145 were characterized by PFGE, yielding 22 profiles. All profiles were within five band differences of at least one other profile. Classical EMRSA-15 isolates showed nine PFGE profiles, with the majority of isolates (68%) in profile B1. Six of these nine PFGE profiles were unique to the classical EMRSA-15 isolates. Among the phage variants of EMRSA-15, 16 profiles were seen, but the majority of isolates (83%) fell into 1 of 4 profiles (B2, B3, B4, and B7) which correlated well with phage patterns. The most divergent PFGE profiles among the EMRSA-15 isolates had as many as 12 band differences from one another, suggesting that in examining isolates belonging to such a temporally and geographically disseminated epidemic strain, the range of PFGE profiles must be regarded as a continuum and analyzed by relating the profiles back to the most common or progenitor profile. PMID- 11283085 TI - 16S ribosomal DNA sequence analysis distinguishes biotypes of Streptococcus bovis: Streptococcus bovis Biotype II/2 is a separate genospecies and the predominant clinical isolate in adult males. AB - We characterized 22 human clinical strains of Streptococcus bovis by genotypic (16S rRNA gene sequence analysis [MicroSeq]; Applied Biosystems, Foster City, Calif.) and phenotypic (API 20 Strep and Rapid ID32 Strep systems (bioMerieux Vitek, Hazelton, Mo.) methods. The strains, isolated from blood, cerebrospinal fluid (CSF), and urine, formed two distinct 16S ribosomal DNA sequence clusters. Three strains which were associated with endocarditis urinary tract infection (UTI), and sepsis clustered with the S. bovis type strain ATCC 33317 (cluster 1); other closely related type strains were S. equinus and S. infantarius. Nineteen strains clustered at a distance of about 2.5% dissimilarity to the S. bovis type strain (cluster 2) and were associated with central nervous system (CNS) disease in addition to endocarditis, UTI, and sepsis. All strains were distinct from S. gallolyticus. Within cluster 2, a single strain grouped with ATCC strain 43143 (cluster 2a) and may be phenotypically distinct. All the other strains formed a second subgroup (cluster 2b) that was biochemically similar to S. bovis biotype II/2 (mannitol negative and beta galactosidase, alpha galactosidase, beta glucuronidase, and trehalose positive). The API 20 Strep system identified isolates of cluster 2b as S. bovis biotype II/2, those of cluster 1 as S. bovis biotype II/1, and that of cluster 2a as S. bovis biotype I. There was an excellent correlation of biotype and genotype: S. bovis biotype II/2 isolates form a separate genospecies distinct from the S. bovis, S. gallolyticus, and S. infantarius type strains and are the most common isolates in adult males. PMID- 11283086 TI - Simultaneous detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae in suspected cases of meningitis and septicemia using real-time PCR. AB - A single-tube 5' nuclease multiplex PCR assay was developed on the ABI 7700 Sequence Detection System (TaqMan) for the detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae from clinical samples of cerebrospinal fluid (CSF), plasma, serum, and whole blood. Capsular transport (ctrA), capsulation (bexA), and pneumolysin (ply) gene targets specific for N. meningitidis, H. influenzae, and S. pneumoniae, respectively, were selected. Using sequence-specific fluorescent-dye-labeled probes and continuous real-time monitoring, accumulation of amplified product was measured. Sensitivity was assessed using clinical samples (CSF, serum, plasma, and whole blood) from culture-confirmed cases for the three organisms. The respective sensitivities (as percentages) for N. meningitidis, H. influenzae, and S. pneumoniae were 88.4, 100, and 91.8. The primer sets were 100% specific for the selected culture isolates. The ctrA primers amplified meningococcal serogroups A, B, C, 29E, W135, X, Y, and Z; the ply primers amplified pneumococcal serotypes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10A, 11A, 12, 14, 15B, 17F, 18C, 19, 20, 22, 23, 24, 31, and 33; and the bexA primers amplified H. influenzae types b and c. Coamplification of two target genes without a loss of sensitivity was demonstrated. The multiplex assay was then used to test a large number (n = 4,113) of culture-negative samples for the three pathogens. Cases of meningococcal, H. influenzae, and pneumococcal disease that had not previously been confirmed by culture were identified with this assay. The ctrA primer set used in the multiplex PCR was found to be more sensitive (P < 0.0001) than the ctrA primers that had been used for meningococcal PCR testing at that time. PMID- 11283087 TI - Genetic diversity of Mycobacterium tuberculosis in Sicily based on spoligotyping and variable number of tandem DNA repeats and comparison with a spoligotyping database for population-based analysis. AB - In a previous study, we proposed to associate spoligotyping and typing with the variable number of tandem DNA repeats (VNTR) as an alternative strategy to IS6110 restriction fragment length polymorphism (RFLP) for molecular epidemiological studies on tuberculosis. The aim of the present study was to further evaluate this PCR-based typing strategy and to describe the population structure of Mycobacterium tuberculosis in another insular setting, Sicily. A collection of 106 DNA samples from M. tuberculosis patient isolates was characterized by spoligotyping and VNTR typing. All isolates were independently genotyped by the standard IS6110-RFLP method, and clustering results between the three methods were compared. The totals for the clustered isolates were, respectively, 15, 60, and 82% by IS6110-RFLP, spoligotyping, and VNTR typing. The most frequent spoligotype included type 42 that missed spacers 21 to 24 and spacers 33 to 36 and derived types 33, 213, and 273 that, together represented as much as 26% of all isolates, whereas the Haarlem clade of strains (types 47 and 50, VNTR allele 32333) accounted for 9% of the total strains. The combination of spoligotyping and VNTR typing results reduced the number of clusters to 43% but remained superior to the level of IS6110-RFLP clustering (ca. 15%). All but one IS6110 defined cluster were identified by the combination of spoligotyping and VNTR clustering results, whereas 9 of 15 spoligotyping-defined clusters could be further subdivided by IS6110-RFLP. Reinterpretation of previous IS6110-RFLP results in the light of spoligotyping-VNTR typing results allowed us to detect an additional cluster that was previously missed. Although less discriminative than IS6110-RFLP, our results suggest that the use of the combination of spoligotyping and VNTR typing is a good screening strategy for detecting epidemiological links for the study of tuberculosis epidemiology at the molecular level. PMID- 11283088 TI - Genotypic characterization of Toxoplasma gondii strains associated with human toxoplasmosis in Spain: direct analysis from clinical samples. AB - Genetic analysis of the SAG2 locus was performed to determine the prevalence of the different genotypes of Toxoplasma gondii (strain types I, II, and III) associated with human toxoplasmosis in Spain. This determination was made directly from primary clinical samples, obviating the previous process of isolation in mice or cell culture. A total of 34 isolates of T. gondii, collected from immunocompromised patients and congenital infection cases, were analyzed. Restriction fragment length polymorphism in PCR-amplified SAG2 products was used to group strains into one of the three genotypes of T. gondii. Complete characterization of the SAG2 gene was successful in 76.5% of the cases, demonstrating the feasibility of direct genotype analysis from clinical samples of different origins. Strains of T. gondii type II were the most prevalent in immunocompromised patients, with 52% of cases, while strains of type I were present in 75% of the congenital infection cases. These data differ from previous reports that show type II strains to be mostly associated with all kinds of human toxoplasmosis. These differences might be an effect of selection in the process of culture and isolation of the samples performed by other researchers prior to strain characterization. PMID- 11283089 TI - Quantitation of bacteria in bone marrow from patients with typhoid fever: relationship between counts and clinical features. AB - Enteric fever is the only bacterial infection of humans for which bone marrow examination is routinely recommended. A prospective study of the concentrations of bacteria in the bone marrow and their relationship to clinical features was conducted with 120 Vietnamese patients with suspected enteric fever, of whom 89 had confirmed typhoid fever. Ninety-three percent of the Salmonella enterica serovar Typhi samples isolated were resistant to ampicillin, chloramphenicol, and co-trimoxazole. For 81 patients with uncomplicated typhoid and satisfactory bone marrow aspirates, the number of serovar Typhi CFU in bone marrow aspirates was a median value of 9 (interquartile range [IQR], 1 to 85; range, 0.1 to 1,580) compared to 0.3 (IQR, 0.1 to 10; range, 0.1 to 399) CFU/ml in simultaneously sampled blood. The ratio of individual blood counts to bone marrow counts was 10 (IQR, 2.3 to 97.5). The number of bacteria in blood but not bone marrow was correlated inversely with the duration of preceding fever. Thus, with increasing duration of illness the ratio of bone marrow-to-blood bacterial concentrations increased; the median ratio was 4.8 (IQR, 1 to 27.5) during the first week compared with 158 (IQR, 60 to 397) during the third week. After lysing the host cells, the median ratio of viable bone marrow to blood increased, reflecting the higher concentration of intracellular serovar Typhi in the bone marrow. Effective antibiotic pretreatment had a significantly greater effect in reducing blood counts compared to bone marrow counts (P < 0.001). Thus, bacteria in the bone marrow of typhoid patients are less affected by antibiotic treatment than bacteria in the blood. The numbers of bacteria in bone marrow correlated negatively with the white blood cell (R = -0.3, P = 0.006) and platelet counts (R = -0.32, P = 0.01) and positively with fever clearance time after treatment (R = 0.4, P < 0.001). The bacterial load in bone marrow therefore may reflect the clinical course of the infection, and high levels may suppress neutrophil proliferation. PMID- 11283090 TI - Colonic infection by Bilophila wadsworthia in pigs. AB - Bilophila wadsworthia is a common inhabitant of the human colon and has been associated with appendicitis and other local sites of inflammation in humans. Challenge-exposure or prevalence studies in laboratory and other animals have not been reported. B. wadsworthia is closely related phylogenetically to Desulfovibrio sp. and Lawsonia intracellularis, which are considered colon pathogens. We developed a PCR specific for B. wadsworthia DNA. Samples of bacterial DNA extracted from the feces of pigs on six farms in Australia and four farms in Venezuela were examined. Specific DNA of B. wadsworthia was detected in the feces of 58 of 161 Australian and 2 of 45 Venezuelan pigs, results comprising 100% of the neonatal pigs, 15% of the weaned grower pigs, and 27% of the adult sows tested. Single-stranded conformational polymorphism analysis of PCR product DNA derived from pigs or from known human strains showed an identical pattern. Histologic examination of the intestines of weaned B. wadsworthia-positive pigs found no or minor specific lesions in the small and large intestines, respectively. B. wadsworthia is apparently a common infection in neonatal pigs, but its prevalence decreases after weaning. The possible role of B. wadsworthia as an infection in animals and in human colons requires further study. PMID- 11283092 TI - Persistence of a distinct Corynebacterium diphtheriae clonal group within two communities in the United States and Canada where diphtheria is endemic. AB - Molecular characterization of 53 U.S. and Canadian Corynebacterium diphtheriae isolates by multilocus enzyme electrophoresis, ribotyping, and random amplified polymorphic DNA showed that strains with distinct molecular subtypes have persisted in the United States and Canada for at least 25 years. These strains are endemic rather than imported from countries with current endemic or epidemic diphtheria. PMID- 11283091 TI - Isolation of Helicobacter cinaedi from the colon, liver, and mesenteric lymph node of a rhesus monkey with chronic colitis and hepatitis. AB - On the basis of biochemical, phenotypic, and 16S rRNA analyses, Helicobacter cinaedi was isolated from the colon, liver, and mesenteric lymph nodes of a 2 year-old rhesus monkey with chronic diarrhea. Histologically, the liver had mild to moderate biliary hyperplasia and hypertrophy with periportal inflammation and fibrosis. Colonic and cecal lesions consisted of diffuse chronic inflammation and glandular hyperplasia extending the length of the crypts. This is the first observation of H. cinaedi associated with active hepatitis and colitis in a nonhuman primate. PMID- 11283093 TI - The susceptibility of Mycobacterium tuberculosis to isoniazid and the Arg-->Leu mutation at codon 463 of katG are not associated. AB - A mutation (CCG-->CTG [Arg-->Leu]) in codon 463 of katG (catalase peroxidase) of Mycobacterium tuberculosis has been found in isoniazid (INH)-resistant strains. A PCR restriction endonuclease analysis to detect this mutation was applied to 395 M. tuberculosis isolates from patients in The Netherlands. The proportion of isolates with a detectable mutation was 32% (32 out of 100) and 29% (85 out of 295) among INH-susceptible isolates and INH-resistant or -intermediate isolates, respectively. Sequencing of five INH-susceptible isolates with such mutations showed that all five had the Arg463Leu mutation. We conclude that the Arg463Leu mutation of katG of M. tuberculosis is not a reliable indicator of INH resistance. PMID- 11283094 TI - Use of spoligotyping to study the evolution of the direct repeat locus by IS6110 transposition in Mycobacterium tuberculosis. AB - Based on the variability of 43 spacers within the direct repeat (DR) locus of Mycobacterium tuberculosis complex organisms, spoligotyping is a rapid method that aids in the study of the epidemiology of tuberculosis. It was recently hypothesized that despite its presence in the DR locus, spacer 31 could not be amplified in M. tuberculosis clinical isolates belonging to spoligotype 50 due to the insertion of an extra copy of IS6110 between spacers 31 and 32 that could lead to an asymmetrical split of the primer targets (I. Filliol, C. Sola, and N. Rastogi, J. Clin. Microbiol. 38:1231--1234, 2000). In the present investigation, previous observations were extended to 25 clinical isolates of type 50 showing that the primer set IS6-DRb that selectively amplified the left and central DR regions was indeed able to demonstrate the presence of spacer 31. IS6110 restriction fragment length polymorphism (RFLP) and DR-RFLP showed that type 50 isolates were characterized by the presence of two copies of IS6110 associated with the DR locus and an additional double IS6110 band of 1.4 kb. The primer set IS3-IS6 was then used to selectively amplify a 750-bp inter-IS6110 fragment within the DR locus. The sequencing of the central DR region corroborated our previous findings and showed that the absence of spacer 31 among the type 50 isolates was due to the asymmetric insertion of an extra copy of IS6110 between spacers 31 and 32, leading to an unequal split of the DRa-DRb target into two portions, of 6 and 30 bp, respectively. These results show that the DR locus constitutes an ideal IS6110 preferential locus (ipl), permitting the insertion of two or more copies of IS6110, and provide new clues for epidemiological and phylogenetic interpretation of changes in IS6110-RFLP and spoligotyping profiles. PMID- 11283095 TI - Typing of Candida albicans isolates by sequence analysis of the cytochrome b gene and differentiation from Candida stellatoidea. AB - Including type strains, mitochondrial cytochrome b genes of 32 strains of Candida albicans and 6 strains of Candida stellatoidea, presently treated as a synonym for C. albicans, were partially sequenced. Analysis of 396-bp nucleotide sequences of the strains under investigation divided C. albicans isolates into three types: type I, type II, and type III; however, strains of C. stellatoidea represented distinct type IV isolates. Deduced amino acid sequences of type I, type II, and type III were identical and differed from that of type IV by one amino acid. Genotypes (rDNA type) of the test strains were also checked. Cytochrome b typing did not correlate with genotyping, and different genotypes occurred for one cytochrome b type. This study shows that cytochrome b gene sequences are useful for analyzing the genetic relatedness of C. albicans isolates and effective for differentiating C. stellatoidea from C. albicans. PMID- 11283096 TI - Weissella confusa (basonym: Lactobacillus confusus) bacteremia: a case report. AB - Infection with Lactobacillus is rare, and only a handful of species have been identified as being clinically significant: Lactobacillus casei, Lactobacillus rhamnosus, and Lactobacillus leichmannii. The literature contains one case report of bacteremia caused by Weissella confusa (basonym: Lactobacillus confusus), but the clinical significance of the infection was unclear. We describe a case of W. confusa bacteremia in a 46-year-old man with a history of abdominal aortic dissection and repair. This procedure was complicated by gut ischemia, which necessitated massive small bowel resection. He subsequently developed short-bowel syndrome, which required him to have total parenteral nutrition. He later developed an Enterococcus faecalis aortic valve endocarditis that required a coronary artery bypass graft and aortic root replacement with homograft and 6 weeks of intravenous ampicillin and gentamicin. Three months prior to his most recent admission, he was diagnosed with Klebsiella pneumoniae bacteremia and candidemia. At the present admission, he had fever (T(max), 39.5 degrees C) and chills of 2 days' duration and was admitted to the intensive care unit because of hemodynamic instability. Blood cultures grew K. pneumoniae and W. confusa in four of four blood culture bottles (both aerobe and anaerobe bottles). Imaging studies failed to find any foci of infection. A transesophageal echocardiogram revealed no vegetations. A culture of the patient's Hickman catheter tip was negative. The patient was treated with piperacillin-tazobactam and gentamicin. His condition improved, and he was discharged home, where he completed 4 weeks of piperacillin tazobactam therapy. Lactobacillemia seldom results in mortality; however, it may be a marker of a serious underlying disease. It is usually seen in patients who have a complex medical history or in patients who receive multiple antibiotics. Lactobacillus spp. are generally associated with polymicrobial infections, and when isolated from the blood, they need to be considered possible pathogens. The presence of a vancomycin-resistant, gram-positive coccobacilli on a blood culture should alert clinicians to the possibility of bacteremia caused by W. confusa or other small gram-positive rods. PMID- 11283097 TI - Cryptococcus laurentii fungemia in a premature neonate. AB - Cryptococcus spp. other than Cryptococcus neoformans are generally considered nonpathogenic to humans. There are only 15 case reports of disease in humans caused by Cryptococcus laurentii infection. Underlying diseases and predisposing risk factors seem to play an important role in these cases. Our patient is the first case of an extremely low birth weight infant with C. laurentii fungemia reported in the English literature. In our case, the MIC of amphotericin B for C. laurentii was 0.25 to 1 microg/ml and the patient had a good outcome following the administration of amphotericin B at 10 mg/kg combined with central venous catheter removal. There will undoubtedly be an increasing occurrence of unusual fungal infections accompanying further advances in medicine. A high degree of suspicion and improvements in the techniques for culture and identification will contribute to the earlier diagnosis and treatment of unusual fungal infections. PMID- 11283098 TI - Comparison of Roche MONITOR and Organon Teknika NucliSens assays to quantify human immunodeficiency virus type 1 RNA in cerebrospinal fluid. AB - We compared Roche MONITOR and Organon Teknika NucliSens assays for human immunodeficiency virus type 1 (HIV-1) RNA in cerebrospinal fluid (CSF). Results of 282 assays were highly correlated (r = 0.826), with MONITOR values being 0.29 +/- 0.4 log(10) copies/ml (mean +/- standard deviation) values. Both assays can reliably quantify HIV-1 RNA in CSF. PMID- 11283099 TI - Fluconazole disk diffusion test with methylene blue- and glucose-enriched Mueller Hinton agar for determining susceptibility of Candida species. AB - A 25-microg fluconazole disk diffusion test using a Mueller-Hinton agar plate containing 2% glucose and 5 microg of methylene blue/ml (GM-MH) was compared to the macrodilution reference method for 210 Candida species. The GM-MH agar plate was read at 24 h. The predictive values of disks with susceptible, intermediate, and resistant results on the GM-MH agar plate at 24 h were 97.1, 56.3, and 76.5%, respectively. PMID- 11283100 TI - Evaluation of PCR methods for rapid identification and differentiation of Streptococcus uberis and Streptococcus parauberis. AB - Streptococcus uberis and Streptococcus parauberis reference strains and isolates obtained from routine diagnostics were investigated by PCR with oligonucleotide primers designed according to species-specific parts of the 16S rRNA gene, the 23S rRNA gene, and the 16S-23S rRNA intergenic spacer region of both species. All three primer pairs allowed an identification of 67 isolates as S. uberis and 4 isolates as S. parauberis. PMID- 11283101 TI - Random amplified polymorphic DNA assay as a rapid tool in screening for Neisseria meningitidis serogroup C isolates of electrophoretic type 24. AB - Neisseria meningitidis serogroup C (NMSC) isolates of electrophoretic type 24 (ET 24), as identified by multilocus enzyme electrophoresis, are the main cause of serogroup C meningococcal disease outbreaks and sporadic meningococcal disease in the United States. We evaluated a random amplified polymorphic DNA assay as a rapid tool to screen for isolates of ET-24 by testing 199 NMSC isolates of 51 different ETs. A sensitivity of 88% and a specificity of 87% was achieved in identification of ET-24 isolates when the patterns obtained by two primers, P1 and P5, were analyzed together. PMID- 11283102 TI - Nucleic acid sequence-based amplification of Aspergillus RNA in blood samples. AB - Nucleic acid sequence-based amplification (NASBA), an isothermal amplification technique, was established and evaluated for the detection of Aspergillus RNA and compared with a previously published, well-defined real-time PCR assay amplifying a region of the Aspergillus 18S rRNA gene. NASBA showed a lower detection limit of 1 CFU and detected RNA from five different clinically relevant Aspergillus species, including Aspergillus fumigatus. All 77 blood samples tested by PCR and NASBA showed identical results in both assays. Results with the NASBA technique were obtained within 6 h. Thus, the NASBA technique provided a valuable tool for sensitive, specific, fast, and reliable detection of Aspergillus RNA with potential for routine diagnosis, including the possibility to test the viability of cells. PMID- 11283103 TI - Detection of microsporidia in travelers with diarrhea. AB - We examined stool specimens of 148 returning travelers from an outpatient department for tropical diseases for the appearance of microsporidia using light microscopy and PCR. Intestinal microsporidiosis was diagnosed for five patients by light microscopy and for nine patients by PCR. Some cases were diagnosed only by PCR, indicating that the true prevalence has to be determined by highly sensitive techniques, such as PCR. PMID- 11283104 TI - B-Cell epitope mapping of the VapA protein of Rhodococcus equi: implications for early detection of R. equi disease in foals. AB - Linear B-cell epitopes of the Rhodococcus equi virulence-associated protein (VapA) were mapped using a synthetic peptide bank in this study. The peptides were screened in an enzyme-linked immunosorbent assay (ELISA) with a total of 70 sera from foals with current R. equi disease (51 sera), as well as from foals that had either recovered from R. equi infection 10 months previously (3 sera) or that had no known history of R. equi disease (16 sera). An epitope with the sequence NLQKDEPNGRA was identified and was universally recognized by all 51 sera from foals with R. equi disease and was not recognized by any of the other sera. There was poor reactivity between all sera and peptides relating to other areas of the VapA protein. It is proposed that an ELISA based upon a defined peptide epitope may be used in an improved serological diagnostic test for R. equi infection in foals. PMID- 11283105 TI - PCR-hybridization assay for Mycobacterium avium complex: optimization of detection in peripheral blood from humans. AB - We evaluated the sensitivity of a DNA amplification test for the detection of Mycobacterium avium in blood samples using different blood components and different DNA extraction methods. M. avium-inoculated blood samples were processed to obtain separate blood components: peripheral blood mononuclear cells (PBMCs), polymorphonuclear cells (PMNCs), and whole-blood sodium dodecyl sulfate (SDS)-lysate pellets. The sensitivity for the detection of the lowest mycobacterial load (1 CFU/ml) was significantly greater (P < 0.01) with DNA extracted from SDS-lysate pellets than with DNA extracted from PBMCs or PMNCs. Subsequently, DNA extraction methods based on guanidine NaOH, and proteinase were compared. The sensitivity of the guanidine-based method was significantly greater (P < 0.01) than those of the others. PMID- 11283106 TI - Genetic Variation among Human Isolates of Uninucleated Cyst-Producing Entamoeba Species. AB - Twelve human infections with Entamoeba spp. producing uninucleated cysts were studied. DNA was extracted from infected feces and used to amplify part of the ameba small-subunit rRNA gene. Sequence analysis identified four distinct types of Entamoeba, all of which are related to Entamoeba polecki and E. chattoni and two of which have not been reported previously. Whether these genetic types represent different species is unclear. We propose that the agent of all human infections with uninucleated cyst-producing Entamoeba species be reported as "E. polecki-like." PMID- 11283107 TI - Evaluation of Candida ID, a new chromogenic medium for fungal isolation and preliminary identification of some yeast species. AB - Candida ID, a new chromogenic medium, allows identification of Candida albicans (blue colonies) and preliminary identification into a group of four species (pink colonies). In comparison with Albicans ID2 and Sabouraud gentamicin chloramphenicol on 446 fungal strains, Candida ID allowed the isolation of more species than Albicans ID 2 (95.5% versus 91.2%). PMID- 11283108 TI - Validation of the string test for the recovery of Helicobacter pylori from gastric secretions and correlation of its results with urea breath test results, serology, and gastric pH levels. AB - The efficacy of the string culture test to isolate Helicobacter pylori from gastric secretions of 28 volunteers was studied. With the urea breath test (UBT) as the "gold standard," the string culture test showed a sensitivity of 75% and a specificity of 100%. The results of string culture did not correlate with the UBT results, with serum antibody levels, or with the pH levels of gastric secretions. PMID- 11283109 TI - Isolation of amantadine-resistant influenza a viruses (H3N2) from patients following administration of amantadine in Japan. AB - In Japan, the use of amantadine for treatment of influenza A virus infection was not accepted until November 1998, although it was widely used for treatment of Parkinsonism. Since then, we have monitored the emergence of amantadine-resistant viruses and isolated two viruses from patients on long-term treatment with amantadine. PMID- 11283110 TI - Susceptibility of Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii to antimicrobial agents used in selective media. AB - Several antimicrobial agents used in selective media for the isolation of Arcobacter were found to be inhibitory to strains belonging to this genus. All three species tested were susceptible to colistin and rifampin at concentrations used in selective media. Arcobacter skirrowii was the most susceptible species. 5 Fluorouracil, novobiocin, trimethoprim, and teicoplanin or vancomycin were found to be without any inhibitory effect on the strains tested at concentrations described for the isolation of Arcobacter species. PMID- 11283112 TI - Serum is the preferred clinical specimen for diagnosis of human brucellosis by PCR. AB - Human brucellosis poses a significant public health problem in many developing countries and requires fast and accurate diagnosis. A PCR assay amplifying part of the 31-kDa Brucella abortus antigenic protein gene sequence was developed and applied to whole-blood and serum samples from 31 brucellosis patients and 45 healthy individuals. All patients except one had detectable Brucella DNA in either whole blood or serum (combined sensitivity, 97%), but the assay sensitivity was higher with serum samples (94%) than with whole-blood samples (61%). The assay specificity was excellent (100%). A confirmatory PCR assay targeting another Brucella gene region (omp-2) was also developed but lacked sensitivity. Serum is the optimal specimen for the diagnosis of brucellosis by PCR, a choice that leads to assay simplification and shortens turnaround time. PMID- 11283111 TI - Multilocus genotyping indicates that the ability to invade the bloodstream is widespread among Candida albicans isolates. AB - Multilocus genotyping was used to compare populations of Candida albicans from oral mucosa and blood. No significant differences in allele frequencies between the two samples were detected, and in a dendrogram of genotypic similarities, genotypes from both types of samples were finely interspersed. This is evidence for widespread distribution of invasive potential. PMID- 11283113 TI - Relevance of reactivity in commercially available hepatitis C virus antibody assays. AB - Sera from 2,148 patients were tested with a third-generation microparticle enzyme immunoassay (MEIA), a confirmatory assay, and a reverse transcription-PCR. Overall, 85.6% of reactivities were confirmed, 13.2% were shown to be unspecifically reactive, and 1.2% were indeterminate. The rate of confirmed MEIA reactivities clearly depended on the strength of the reactivity. PMID- 11283114 TI - Low Prevalence of Community-Acquired Methicillin-Resistant Staphylococcus aureus in Adults at a University Hospital in the Central United States. AB - Community-acquired MRSA (CA-MRSA) is potentially a new emerging pathogen with most strains susceptible to many antimicrobials except for beta-lactam antibiotics. We retrospectively reviewed MRSA isolates during a 20-month study period (January 1998 through August 1999) and investigated those that were clindamycin susceptible. Patients were not considered to harbor CA-MRSA if they had been admitted to a hospital within the preceding 2 years or if their isolate had been obtained more than 72 h after admission. There were 2,817 S. aureus isolates, with 1,071 (38%) being MRSA. Of these 1,071 isolates, 161 were clindamycin susceptible; these were recovered from 81 patients. Of these 81 patients, 20 appeared to have community-acquired strains, but only 2 could be confirmed as having CA-MRSA. PMID- 11283115 TI - Turicella otitidis as an unusual agent causing a posterior auricular abscess. AB - A posterior auricular abscess in a 3-year-old girl was confirmed to have been caused by an unusual organism, Turicella otitidis. PMID- 11283116 TI - Isolations of Leclercia adecarboxylata from a patient with a chronically inflamed gallbladder and from a patient with sepsis without focus. AB - Leclercia adecarboxylata was isolated from a patient with a chronically inflamed gallbladder, together with Enterococcus sp. The organism was considered clinically significant and was susceptible to all antibiotics tested. Another strain of L. adecarboxylata was cultured from blood, together with Escherichia hermannii and E. faecalis, from a patient with sepsis. PMID- 11283117 TI - Chronic melioidosis in a patient with cystic fibrosis. AB - Burkholderia pseudomallei, the causative agent of melioidosis, is endemic in Southeast Asia and northern Australia, where it can be found in soil and surface water. We report a case of chronic pulmonary melioidosis in a patient with cystic fibrosis who had traveled to an area where B. pseudomallei is endemic. PMID- 11283118 TI - Vertebral osteomyelitis caused by Enterococcus raffinosus. AB - Enterococcus raffinosus is a rare isolate in clinical specimens. A case of vertebral osteomyelitis caused by E. raffinosus in an elderly patient is described and confirms this organism to be an opportunistic human pathogen. PMID- 11283119 TI - Tamoxifen for the prevention of breast cancer: psychosocial impact on women participating in two randomized controlled trials. AB - PURPOSE: The purpose of this study was to evaluate the psychosocial implications of tamoxifen versus placebo in women who are at increased risk of breast cancer. PATIENTS AND METHODS: The 488 women in the psychosocial study were recruited from participants in two placebo-controlled, double-blind, randomized, controlled trials that investigated the efficacy of tamoxifen in the prevention of breast cancer in women who are at high familial risk. During a 5-year period, repeated assessments were made of anxiety, psychological distress, and sexual functioning using standardized questionnaires before treatment at baseline and at 6-month intervals during the trial. RESULTS: Questionnaire completion over 5 years was good, with 71.1% of women returning at least 8 of 10 follow-up assessments. Although scores from individuals showed considerable fluctuation and variation over time, changes in anxiety, mood, and sexual functioning were not associated with treatment group. The number of symptoms reported at 48 months via a self report checklist were not associated with treatment group, but vasomotor symptoms were more frequent among tamoxifen-treated women. Symptoms of low energy, breast sensitivity, and visual blurring were reported most frequently in the placebo group. CONCLUSION: In general, these results are comparable to those from the National Surgical Adjuvant Breast and Bowel Project psychosocial study despite differences in study populations, methodology, and instruments. The long-term use of tamoxifen and other selective estrogen response modulators as preventive agents in high-risk groups has been questioned, but we found no evidence of treatment-related side effects that affect women's psychosocial and sexual functioning. PMID- 11283120 TI - Clinical evidence for topotecan-paclitaxel non--cross-resistance in ovarian cancer. AB - PURPOSE: A large, randomized study comparing the efficacy and safety of topotecan versus paclitaxel in patients with relapsed epithelial ovarian cancer showed that these two compounds have similar activity. In this study, a number of patients crossed over to the alternative drug as third-line therapy, ie, from paclitaxel to topotecan and vice versa. We therefore were able to assess the degree of non cross-resistance between these two compounds. PATIENTS AND METHODS: Patients who had progressed after one platinum-based regimen were randomized to either topotecan (1.5 mg/m(2)/d) x 5 every 21 days (n = 112) or paclitaxel (175 mg/m(2) over 3 hours) every 21 days (n = 114). A total of 110 patients received cross over therapy with the alternative drug (61 topotecan, 49 paclitaxel) as third line therapy. RESULTS: Response rates to third-line cross-over therapy were 13.1% (8 of 61 topotecan) and 10.2% (5 of 49 paclitaxel; P =.638). Seven patients who responded to third-line topotecan and four patients who responded to paclitaxel had failed to respond to their second-line treatment. Median time to progression (from the start of third-line therapy) was 9 weeks in both groups, and median survival was 40 and 48 weeks for patients who were receiving topotecan or paclitaxel, respectively. The principal toxicity was myelosuppression; grade 4 neutropenia was more frequent with topotecan (81.4% of patients) than with paclitaxel (22.9% of patients). CONCLUSION: Topotecan and paclitaxel have similar activity as second-line therapies with regard to response rates and progression free and overall survival. We demonstrated that the two drugs have a degree of non-cross-resistance. Thus, there is a good rationale for incorporating these drugs into future first-line regimens. PMID- 11283121 TI - Combination chemotherapy with carboplatin and docetaxel in the treatment of cancers of the ovary and fallopian tube and primary carcinoma of the peritoneum. AB - PURPOSE: Standard chemotherapy for advanced ovarian cancer currently includes a platinum agent (usually carboplatin) and paclitaxel. Because docetaxel is an active agent in platinum-resistant ovarian cancer, it is relevant to evaluate both the toxicity and efficacy of the combination of carboplatin and docetaxel in this clinical setting. PATIENTS AND METHODS: The Gynecologic Oncology Program of the Cleveland Clinic Taussig Cancer Center conducted a phase II trial of carboplatin (area under the concentration-versus-time curve of 6) and docetaxel (60 mg/m(2)), delivered every 3 weeks for six courses, in patients with ovarian and fallopian tube cancers and primary carcinoma of the peritoneum who had either received no prior chemotherapy or had experienced a treatment-free interval of greater than 2 years before developing disease recurrence. RESULTS: Fifty patients (median age, 57 years; range, 44 to 81 years) entered the trial (47 had had no prior chemotherapy). Our toxicity findings included the following: grade 4 neutropenia (64% of patients); hypersensitivity reactions (34%, none requiring discontinuation of therapy); peripheral neuropathy (6%). We had objective responses for 32 of 42 (81%) assessable patients. CONCLUSION: The combination of carboplatin and docetaxel is highly active in ovarian cancer, with the major toxicity being bone marrow suppression. Hypersensitivity reactions are frequent but do not prevent continuation of treatment. With the dose and schedule employed in this trial, neurotoxicity is uncommon. Defining a role for this regimen in routine clinical practice will require the conduct of randomized controlled clinical trials. PMID- 11283122 TI - Human papillomavirus and prognosis of invasive cervical cancer: a population based study. AB - PURPOSE: To determine the association between human papillomavirus (HPV) type and prognosis of patients with invasive cervical carcinoma. PATIENTS AND METHODS: Patients diagnosed with International Federation of Gynecology and Obstetrics (FIGO) stage IB to IV cervical cancer between 1986 and 1997 while residents of three Washington State counties were included (n = 399). HPV typing was performed on paraffin-embedded tumor tissue using polymerase chain reaction methods. Patients were observed for a median of 50.8 months. Total mortality (TM) and cervical cancer-specific mortality (CCSM) were determined. Hazards ratios (HR) adjusted for age, stage, and histologic type were estimated using multivariable models. RESULTS: Eighty-six patients had HPV 18-related tumors and 210 patients had HPV 16-related tumors. Cumulative TM among patients with HPV 18-related tumors and among patients with HPV 16-related tumors were 33.7% and 27.6%, respectively; cumulative CCSM in these two groups were 26.7% and 18.1%, respectively. Compared with patients with HPV 16-related cancers, patients with HPV 18-related cancers were at increased risk for TM (HR(TM), 2.2; 95% confidence interval [CI], 1.3 to 3.6) and CCSM (HR(CCSM), 2.5; 95% CI, 1.4 to 4.4). The HPV18 associations were strongest for patients with FIGO stage IB or IIA disease (HR(TM), 3.1; 95% CI, 2.3 to 4.2; and HR(CCSM), 5.8; 95% CI, 3.9 to 8.7), whereas no associations were observed among patients with FIGO stage IIB to IV disease. Virtually identical associations were found in the subset of patients with squamous cell carcinoma (n = 219). CONCLUSION: HPV 18-related cervical carcinomas, particularly those diagnosed at an early stage, are associated with a poor prognosis. Elucidating the mechanism or mechanisms underlying this association could lead to new treatment approaches for patients with invasive cervical carcinoma. PMID- 11283123 TI - Duration of hospitalization as a measure of cost on Children's Cancer Group acute lymphoblastic leukemia studies. AB - PURPOSE: We used duration of hospitalization as a surrogate for cost and event free survival as a measure of effectiveness to estimate the cost-effectiveness ratios of various treatment regimens on Children's Cancer Group trials for acute lymphoblastic leukemia. PATIENTS AND METHODS: The analyses included 4,986 children (2 to 21 years of age) with newly diagnosed acute lymphoblastic leukemia enrolled onto risk-adjusted protocols between 1988 and 1995. Analyses were based on a model of 100 patients. The marginal cost-effectiveness ratio (hospital days per additional patient surviving event-free) was the difference in total duration of hospitalization divided by the difference in number of event-free survivors at 5 years for two regimens. Relapse-adjusted marginal cost of frontline therapy was the difference in total duration of hospitalization for frontline therapy plus relapse therapy divided by the difference in number of event-free survivors at 5 years on the frontline therapy for two regimens. RESULTS: One or two delayed intensification (DI) phases, augmented therapy, and dexamethasone all improved outcome. Marginal cost-effectiveness of these regimens compared with the control regimens was 133 days per patient for DI, 117 days per patient for double DI, and 41 days per patient for augmented therapy. Dexamethasone resulted in 17 fewer days per patient. Relapse-adjusted marginal costs were 68 days per patient for DI and 52 days for double DI. Augmented therapy and dexamethasone-based therapy resulted in 16 and 82 fewer hospital days, respectively. The estimated cost effectiveness for treating any first relapse was 250 days per patient. CONCLUSION: DI, double DI, augmented therapy, and dexamethasone-based therapy are cost-effective strategies compared with current treatment of first relapse. PMID- 11283124 TI - Congestive heart failure after treatment for Wilms' tumor: a report from the National Wilms' Tumor Study group. AB - PURPOSE: We determined the frequency of and risk factors for congestive heart failure following treatment for Wilms' tumor that included doxorubicin. PATIENTS AND METHODS: Flow sheets and medical records were reviewed to identify cases of congestive heart failure in a cohort of patients treated on National Wilms' Tumor Studies (NWTS)-1, -2, -3, and -4. The frequency of congestive heart failure was estimated using the Kaplan-Meier method. A case-control study was conducted to determine the relationship among cumulative doxorubicin dose, site(s), total dose of abdominal and thoracic irradiation, sex, and the frequency of congestive heart failure. RESULTS: The cumulative frequency of congestive heart failure was 4.4% at 20 years after diagnosis among patients treated initially with doxorubicin and 17.4% at 20 years after diagnosis among those treated with doxorubicin for their first or subsequent relapse of Wilms' tumor. The relative risk (RR) of congestive heart failure was increased in females (RR = 4.5; P =.004) and by cumulative doxorubicin dose (RR = 3.3/100 mg/m(2); P <.001), lung irradiation (RR = 1.6/10 Gy; P =.037), and left abdominal irradiation (RR = 1.8/10 Gy; P =.013). CONCLUSION: We conclude that congestive heart failure is a risk of treatment with doxorubicin for Wilms' tumor. Additional follow-up of those children treated on NWTS-4 will be necessary to determine if the decrease in dose to 150 mg/m(2) significantly reduces this risk. PMID- 11283125 TI - Value of high-dose cytarabine during interval therapy of a Berlin-Frankfurt Munster-based protocol in increased-risk children with acute lymphoblastic leukemia and lymphoblastic lymphoma: results of the European Organization for Research and Treatment of Cancer 58881 randomized phase III trial. AB - PURPOSE: The European Organization for Research and Treatment of Cancer 58881 study was designed to test in a prospective multicentric randomized trial the value of high-dose (HD) intravenous (IV) cytarabine (Ara-C) added to HD IV methotrexate (MTX) to reduce the incidence of CNS and systemic relapses in children with increased-risk acute lymphoblastic leukemia (ALL) or stage III and IV lymphoblastic lymphoma treated with a Berlin-Frankfurt-Munster (BFM)-based regimen. PATIENTS AND METHODS: After completion of induction-consolidation phase, children with increased-risk (risk factor > 0.8 or T-lineage) ALL or stage III and IV lymphoblastic lymphoma were randomized to receive four courses of HD MTX (5 g/m(2) over 24 hours every 2 weeks) and four intrathecal administrations of MTX (Arm A) or the same treatment schedule with additional HD IV Ara-C (1 g/m(2) in bolus injection 12 and 24 hours after the start of each MTX infusion) (Arm B). RESULTS: Between January 1990 and January 1996, 653 patients with ALL (593 patients) or lymphoblastic lymphoma (60 patients) were randomized: 323 were assigned to Arm A (without Ara-C) and 330 to Arm B (with Ara-C). A total of 190 events (177 relapses and 13 deaths without relapse) were reported, and the median follow up was 6.5 years (range, 2 to 10 years). The incidence rates of CNS relapse were similar in both arms whether isolated (5.6% and 3.3%, respectively) or combined (5.3% and 4.6%, respectively). The estimated 6-year disease-free survival (DFS) rate was similar (log-rank P =.67) in the two treatment groups: 70.4% (SE = 2.6%) in Arm A and 71.0% (SE = 2.5%) in Arm B. The 6-year DFS rate was similar for ALL and LL patients: 70.2% (SE = 1.9%) versus 76.3% (SE = 5.6%). CONCLUSION: Prevention of CNS relapse was satisfactorily achieved with HD IV MTX and intrathecal injections of MTX in children with increased-risk ALL or stage III and IV lymphoblastic lymphoma treated with our BFM-based treatment protocol in which cranial irradiation was omitted. Disappointingly, with the dose schedule used in this protocol, HD Ara-C added to HD MTX, although well tolerated, failed to further decrease the incidence of CNS relapse or to improve the overall DFS. PMID- 11283126 TI - Multimodal treatment of malignant sacrococcygeal germ cell tumors: a prospective analysis of 66 patients of the German cooperative protocols MAKEI 83/86 and 89. AB - PURPOSE: To evaluate a multimodal approach including surgery and cisplatinum chemotherapy for treatment of children with malignant sacrococcygeal germ cell tumors (GCT) and to compare adjuvant and neoadjuvant strategies in advanced tumors. PATIENTS AND METHODS: Between 1983 and 1995, 71 patients with malignant sacrococcygeal GCT were prospectively enrolled onto the German protocols for nontesticular GCT Maligne Keimzelltumoren 83/86 and 89. Five patients who received no chemotherapy (n = 2) or nonplatinum chemotherapy (n = 2) or who did not undergo tumor resection (n = 1) were excluded from this analysis. Among the 66 patients analyzed were 14 boys and 52 girls. The median age was 17.4 months (range, 7 months to 119 months). Median follow-up was 79 months (range, 4 months to 145 months). RESULTS: Fifty-two patients presented with locally advanced stage T2 tumors, and 30 patients had distant metastases at diagnosis. Patients received a median of eight cycles (range, four to nine cycles) of cisplatinum-based chemotherapy. Thirty-five patients underwent tumor resection at diagnosis and received adjuvant cisplatinum-based chemotherapy (group A). Thirty-one patients received up-front chemotherapy followed by delayed tumor resection (group B). Group B included more metastatic tumors than group A (group B, 19 of 31 patients; group A, 11 of 35 patients, P =.01). Preoperative chemotherapy facilitated complete tumor resections (group B, 20 of 31 patients; group A, five of 35 patients, P <.001) and avoided second-look surgery. Metastases at diagnosis and completeness of the first attempt of tumor resection were significant prognostic predictors; however, metastases were not predictive for patients treated with up front chemotherapy. At 5 years follow-up, event-free survival was 0.76 +/- 0.05 (50 of 66 patients), and overall survival was 0.81 +/- 0.05 (54 of 66 patients). Four patients died as a result of therapy-related complications, and eight patients died of their tumors. Patients with locally advanced and metastatic tumors (T2b M1) fared better with neoadjuvant treatment [overall survival: 0.83 +/- 0.09 (16 of 19 patients) versus 0.45 +/- 0.15 (five of 11 patients), P =.01]. CONCLUSION: Even locally advanced and metastatic sacrococcygeal GCT can be successfully treated with up-front cisplatinum-based chemotherapy followed by delayed but complete tumor resection. PMID- 11283127 TI - Treatment of recurrent malignant sacrococcygeal germ cell tumors: analysis of 22 patients registered in the German protocols MAKEI 83/86, 89, and 96. AB - PURPOSE: To evaluate therapeutic options for recurrent malignant sacrococcygeal germ cell tumors (GCT) following three-agent, cisplatinum-based, first-line chemotherapy and tumor resection. PATIENTS AND METHODS: Twenty-two patients were evaluated in 22 first-, 14 second-, five third-, and two fourth-relapse situations. One patient, who relapsed with pure teratoma, was excluded from the analysis of adjuvant treatment. RESULTS: Seventeen patients presented with an isolated local recurrence, two patients showed a distant relapse, and three patients suffered from a combined local and distant recurrence. Twelve patients achieved complete remission (CR) after surgery (n = 12) and adjuvant platinum chemotherapy (n = 10). Seven of these patients remain in continuous CR, and five patients relapsed. All patients who achieved only a partial remission developed a second relapse. Three of 14 patients could be cured after a second (or further) relapse. Altogether, 10 patients survived disease free, and 12 patients died as a result of tumor progression (n = 11) or therapy-related complications (n = 1). The completeness of salvage surgery and clinical remission status after first salvage treatment were the most important prognostic parameters. In addition, patients in first or second relapse with locally advanced or poorly responding tumors benefited from preoperative chemotherapy in combination with regional hyperthermia (RHT). In some patients after microscopically incomplete resection, irradiation at doses > 45 Gy contributed to a favorable outcome. CONCLUSION: The complete resection of the local recurrence represents the cornerstone of salvage treatment. Preoperative platinum-based chemotherapy, combined with RHT in some patients, facilitates complete tumor resection. Radiotherapy should be reserved for those patients with microscopically incomplete tumor resection. As the chance of cure decreases with further relapses, it is important to establish a stringent therapeutic strategy to avoid significant treatment delays and, most importantly, insufficient local therapy. PMID- 11283128 TI - Concomitant infusional paclitaxel and fluorouracil, oral hydroxyurea, and hyperfractionated radiation for locally advanced squamous head and neck cancer. AB - PURPOSE: To improve local disease control and survival with organ preservation, we conducted a phase II multi-institutional trial with a concomitant taxane-based chemotherapy and hyperfractionated radiation regimen. PATIENTS AND METHODS: Sixty four patients with locally advanced squamous cancers (stage IV, 98%; N2/3, 81%) were treated on an intensive regimen consisting of 5-day (120-hour) infusions of paclitaxel (20 mg/m(2)/d) and fluorouracil (600 mg/m(2)/d), oral hydroxyurea 500 mg every 12 hours for 11 doses, and radiation 1.5 Gy bid (T-FH2X). Chemoradiation was administered concomitantly on days 1 to 5 of each 14-day cycle. A full treatment course consisted of five cycles during a 10-week period to a total radiation dose of 72 to 75 Gy. RESULTS: The median follow-up for the group is 34 months. At 3 years, progression-free survival is 63%, locoregional control is 86%, and systemic control is 79%; overall survival is 60%. Seventeen patients died of recurrent cancer, two died of second primary cancers, and four died of other causes. Side effects observed include anemia (22% required transfusion), leucopenia (34%, grade 3 to 4), and mucositis (84%, grade 3 to 4). Organ preservation principles were maintained. At 1 year posttreatment, 61% of patients had severe xerostomia and 47% had compromised swallowing. There was little disturbance of speech quality in 97% of patients at the same follow-up point. CONCLUSION: T-FH2X is a highly active and tolerable concomitant chemotherapy and hyperfractionated radiation regimen that induces sustained local tumor control and holds promise for improved survival with organ preservation in high-risk patients. Identification of less toxic therapy and improved distant disease control are needed. T-FH2X should be tested in a randomized trial and compared with a less intensive concomitant regimen that uses once-daily radiation fractionation. PMID- 11283129 TI - Esophageal cancer: the mode of lymphatic tumor cell spread and its prognostic significance. AB - PURPOSE: Data on skip metastases and their significance are lacking for esophageal cancer. This issue is important to determine the extent of lymphadenectomy for esophageal resection. In this study we examined the lymphatic spread in esophageal cancer by routine histopathology and by immunohistochemistry. PATIENTS AND METHODS: A total of 1,584 resected lymph nodes were obtained from 86 patients with resected esophageal carcinoma and evaluated by routine histopathology. Additionally, frozen tissue sections of 540 lymph nodes classified as tumor-free by routine histopathology were screened for micrometastases by immunohistochemistry with the monoclonal antibody Ber-EP4. The lymph nodes were mapped according to the mapping scheme of the American Thoracic Society modified by Casson et al. RESULTS: Forty-four patients (51%) had pN1 disease, and 61 patients (71%) harbored lymphatic micrometastases detected by immunohistochemistry. Skip metastases detected by routine histopathology were present in 34% of pN1 patients. Skipping of micrometastases detected by immunohistochemistry was found in 66%. The presence of micrometastases was associated with a significantly decreased relapse-free and overall survival (56.0 v 10.0 months and > 64 v 15 months, P <.0001 and P =.004, respectively). Cox regression analysis revealed the independent prognostic influence of micrometastases in lymph nodes. Lymph node skipping had no significant independent prognostic influence on survival. CONCLUSION: Histopathologically and immunohistochemically detectable skip metastases are a frequent event in esophageal cancer. Only extensive lymph node sampling, in conjunction with immunohistochemical evaluation, will lead to accurate staging. An improved staging system is essential for more individualized adjuvant therapy. PMID- 11283130 TI - No downstaging after short-term preoperative radiotherapy in rectal cancer patients. AB - PURPOSE: In retrospective studies, total mesorectal excision (TME) surgery has been demonstrated to result in a reduction in the number of local recurrences of rectal cancer. Reports on improved local control after preoperative, hypofractionated radiotherapy have led to the introduction of a randomized multicenter trial to evaluate the effect of TME surgery with and without preoperative radiotherapy. Treatment with preoperative radiotherapy might have an effect on the pathologic characteristics that determine staging of rectal cancer. We investigated the occurrence of downstaging in rectal cancer patients treated with and without preoperative radiotherapy. PATIENTS AND METHODS: We analyzed the differences in tumor size, number of examined lymph nodes, tumor-node-metastasis stage, and histopathologic features in 1,321 patients entered onto a randomized trial. The trial compared preoperative radiotherapy (5 x 5 Gy) followed by TME surgery with TME surgery alone. Patients who had an interval of more than 10 days between the start of radiotherapy and surgery were excluded from analysis. RESULTS: Differences were observed in tumor size (P <.001) and total number of examined lymph nodes (P <.001). No difference in tumor or node classification was detected. The irradiated group demonstrated more poorly differentiated tumors as well as more mucinous tumors. CONCLUSION: In rectal cancer patients, short-term, preoperative radiotherapy with 5 x 5 Gy does not lead to downstaging if the interval between the start of radiotherapy and surgery does not exceed 10 days. PMID- 11283131 TI - Phase II study of the cyclin-dependent kinase inhibitor flavopiridol administered to patients with advanced gastric carcinoma. AB - PURPOSE: Flavopiridol is the first cyclin-dependent kinase inhibitor to enter clinical trials. Activity in gastric cancer xenografts and in a patient with gastric cancer on the phase I trial led to this phase II study of flavopiridol in patients with metastatic gastric cancer. PATIENTS AND METHODS: Sixteen patients were entered onto the study, and 14 were assessable for response. Flavopiridol was administered initially at a dose of 50 mg/m(2)/d by continuous infusion for 72 hours every 2 weeks. Assessment of plasma pharmacokinetics was performed in all patients. Peripheral mononuclear cells were collected throughout the 72-hour infusion for determinants of apoptosis. RESULTS: There were no major objective responses (exact confidence interval 0% to 23%). One patient achieved a minor response in his liver metastases, though the primary progressed. Other patients exhibited histologic and radiographic evidence of tumor necrosis. Common toxicities included fatigue in 93% of patients (grade 3 or 4 in 27%) and diarrhea in 73% of patients (grade 3 or 4 in 20%). Five patients (33%) developed venous thromboses at the central catheter tip. The studies performed on peripheral mononuclear cells indicated no induction of apoptosis. CONCLUSION: Flavopiridol administered as a single agent for 72 hours every 14 days is inactive in the treatment of gastric cancer. The drug also induced an unexpected higher incidence of vascular thrombosis and fatigue than was anticipated from the phase I trials. Future development of flavopiridol will depend on other doses and schedules in combination with chemotherapy. PMID- 11283132 TI - Accelerated treatment of breast cancer. AB - PURPOSE: Radiation therapy (RT) restricted to the tumor bed, by means of an interstitial implant, and lasting 4 to 5 days after lumpectomy was prospectively evaluated in early-stage breast cancer patients treated with breast-conserving therapy (BCT). The goals of the study were to determine whether treatment time can be reduced and whether elective treatment of the entire breast is necessary. MATERIALS AND METHODS: Between January 1993 and January 2000, 174 cases of early stage breast cancer were managed with lumpectomy followed by RT restricted to the tumor bed using an interstitial implant. Each brachytherapy patient was matched with one external-beam RT (ERT) patient derived from a reference group of 1,388 patients treated with standard BCT. Patients were matched for age, tumor size, histology, margins of excision, absence of an extensive intraductal component, nodal status, estrogen receptor status, and tamoxifen use. Median follow-up for both the ERT and brachytherapy groups was 36 months. RESULTS: No statistically significant differences were noted in the 5-year actuarial rates of ipsilateral breast treatment failure or locoregional failure between ERT and brachytherapy patients (1% v 0%, P =.31 and 2% v 1%, P =.63, respectively). In addition, there were no statistically significant differences noted in rates of distant metastasis (6% v 3%, P =.24), disease-free survival (87% v 91%, P =.55), overall survival (90% v 93%, P =.66), or cause-specific survival (97% v 99%, P =.28). CONCLUSION: Accelerated treatment of breast cancer using an interstitial implant to deliver radiation to the tumor bed alone over 4 to 5 days seems to produce 5 year results equivalent to those achieved with conventional ERT. Extended follow up will be required to determine the long-term efficacy of this treatment approach. PMID- 11283133 TI - Phase I study of a third-generation selective estrogen receptor modulator, LY353381.HCL, in metastatic breast cancer. AB - PURPOSE: We conducted this phase I trial to determine the safety and toxicity profile of LY353381.HCl-a novel, potent, third-generation selective estrogen receptor modulator (SERM)-because this benzothiophene derivative demonstrated an SERM profile in preclinical studies. PATIENTS AND METHODS: We studied 32 patients with recurrent or metastatic breast cancer. Patients were treated in four cohorts with oral daily doses of 10, 20, 50, and 100 mg. Pharmacokinetic sampling was performed during the first 72 hours following the first dose on day 1 and during the 24 hours after the day 57 dose. Eligibility criteria included Eastern Cooperative Oncology Group performance status of 0 to 2; no significant major organ dysfunction; and at least 3 weeks elapsed since most recent hormonal therapy, chemotherapy, and estrogen replacement therapy. RESULTS: The median patient age was 56 years (range, 30 years to 76 years). The median number of prior chemotherapies for metastatic disease was one (range, zero to four), while the median number of prior hormone regimens for metastatic disease was two (range, zero to five). Receptor status was estrogen receptor (ER) positive and progesterone receptor (PR) positive, 19 patients; ER positive and PR negative, eight patients; ER positive and PR unknown, two patients; and ER and PR unknown, three patients. Dose-limiting toxicity was not observed. Treatment was well tolerated with mild to moderate hot flashes in 18 of 32 patients (56%) at all dose levels. Transvaginal ultrasound performed at baseline and after 12 weeks of treatment showed no endometrial thickening. Of the 32 patients evaluable for response, six patients had stable disease for at least 6 months with a median duration of 7.7 months (range, 6.2 months to 33.8 months). The pharmacokinetics of LY353381.HCl were generally linear with respect to time and studied dose range. CONCLUSION: As predicted in preclinical testing, daily oral LY353381.HCl is safe, is well tolerated at all tested dose levels, and may be clinically beneficial in patients with extensively pretreated metastatic breast cancer. Further studies with LY353381 to evaluate the efficacy in patients with or without prior exposure to tamoxifen and fewer overall prior regimens are under way. PMID- 11283134 TI - S-phase fraction and urokinase plasminogen activator are better markers for distant recurrences than Nottingham Prognostic Index and histologic grade in a prospective study of premenopausal lymph node-negative breast cancer. AB - PURPOSE: Histologic grade, Nottingham Prognostic Index (NPI), estrogen receptor (ER) and progesterone receptor (PgR) status, and tumor size have previously been shown to be important prognostic indicators for distant recurrence of breast cancer. The purpose of this study was to compare the prognostic value of these factors with flow cytometric S-phase fraction (SPF), urokinase plasminogen activator (uPA), and plasminogen activator inhibitor type 1 (PAI-1) in premenopausal patients with lymph node-negative breast cancer. PATIENTS AND METHODS: In 237 consecutive premenopausal patients with lymph node-negative breast cancer and freshly frozen tumor material available, SPF, ER and PgR status, uPA and its inhibitor PAI-1, histologic grade, and NPI were evaluated. RESULTS: SPF was univariately the most powerful prognostic factor for distant recurrence, followed by uPA, histologic grade, PgR, age, ER, NPI, and PAI-1, the latter being nonsignificant. Multivariate analysis revealed that neither NPI nor histologic grade was significant after adjustment for SPF, a fact that may be explained by the strong association between these factors. uPA was, however, an independent prognostic factor in addition to SPF, NPI, or histologic grade. CONCLUSION: In this prospective study, SPF and uPA were found to be independent prognostic factors in premenopausal women with lymph node-negative breast cancer. We suggest that SPF, if performed under standardized conditions, can replace histologic grade as a selection instrument for adjuvant medical treatment. The value of the combination of SPF and uPA needs to be confirmed in an independent prospective trial. PMID- 11283135 TI - Low-volume nodal metastases detected at retroperitoneal lymphadenectomy for testicular cancer: pattern and prognostic factors for relapse. AB - PURPOSE: To determine the incidence, pattern, and predictive factors for relapse in patients with low-volume nodal metastases (stage pN1) at retroperitoneal lymphadenectomy (RPLND) and identify who may benefit from chemotherapy in the adjuvant or primary setting. PATIENTS AND METHODS: Fifty-four patients with testicular nonseminomatous germ cell tumor had low-volume retroperitoneal metastases (pathologic stage pN1, 1997 tumor-node-metastasis classification) resected at RPLND, 50 of whom were managed expectantly without adjuvant chemotherapy. The dissection was bilateral in 12 and was a modified template in 38 patients. Retroperitoneal metastases were limited to microscopic nodal involvement in 14 patients. Follow-up ranged from 1 to 106 months (median, 31.4 months). RESULTS: Eleven patients (22%) suffered a relapse at a median follow-up of 1.8 months (range, 0.6 to 28 months). The most frequent form of recurrence was marker elevation in nine (18%) patients. Persistent marker elevation after orchiectomy and before retroperitoneal lymphadenectomy was a significant independent predictor of relapse (relative risk, 8.0; 95% confidence interval, 2.3 to 27.8; P =.001). Four of five (80%) patients with elevated markers (alpha fetoprotein alone in three, alpha-fetoprotein and beta human chorionic gonadotropin in one) suffered a relapse, compared with seven of 45 (15.6%) patients with normal markers. CONCLUSION: Clinical stage I and IIA patients with normal markers who have low-volume nodal metastases have a low incidence of relapse and can be managed by observation only if compliance can be assured. In contrast, patients with elevated markers before retroperitoneal lymphadenectomy have a high rate of relapse and should be considered for primary chemotherapy. PMID- 11283136 TI - Methylation of p15 and p16 genes in acute promyelocytic leukemia: potential diagnostic and prognostic significance. AB - PURPOSE: To investigate the frequency of p15 and p16 gene promoter methylation in acute promyelocytic leukemia (APL), and to define its value in the detection of minimal residual disease (MRD) and treatment prognostication. PATIENTS AND METHODS: Bone marrow DNA obtained from 26 patients with APL at diagnosis and during follow-up was studied with the methylation-specific polymerase chain reaction (MS-PCR). Serial marrow DNA was studied by MS-PCR for MRD, and disease free and overall survival were correlated with p15 methylation status at diagnosis. RESULTS: MS-PCR for p16 and p15 gene methylation has a maximum sensitivity of 10(-4) and 10(-5). At diagnosis, 19 patients (73.1%) exhibited p15 methylation, whereas only three patients (11.5%) exhibited p16 methylation, all of whom had concomitant p15 methylation. During follow-up, p16 methylation was acquired in two patients, one during the third hematologic relapse, and the other during transformation into therapy-related myelodysplastic syndrome. Six patients were evaluated serially with MS-PCR for p15 methylation at diagnosis and at follow-up examinations. Persistent p15 methylation preceded subsequent hematologic relapses in two patients, and conversion to negative MS-PCR for p15 methylation correlated with prolonged survival in another four patients. The 5 year disease-free survival of patients with p15 methylation was significantly inferior to that of patients without p15 methylation (15% v 62.5%; P =.02), and this remained significant in multivariate analysis. CONCLUSION: In APL, p15 but not p16 gene methylation is frequent. It is possible that p16 methylation is acquired during clonal evolution. p15 methylation is a potential marker of MRD and might be of prognostic significance. PMID- 11283138 TI - Breaking bad news about cancer: patients' preferences for communication. AB - PURPOSE: The goal of this study was to assess patients' preferences regarding the way in which physicians deliver news about their cancer diagnosis and management. PATIENTS AND METHODS: A sample of 351 patients with a variety of cancers completed a measure assessing their preferences for how they would like to be told news about their cancer. Patients rated characteristics of the context and content of the conversation as well as physician characteristics. RESULTS: Factor analysis indicated that patients' preferences for how they would like to be told news regarding their cancer can be grouped into the following three categories: (1) content (what and how much information is told); (2) facilitation (setting and context variables); and (3) support (emotional support during the interaction). Women (P =.02) and patients with higher education (P =.05) had significantly higher scores on the Content scale, women (P =.02) had higher scores on the Support scale, and younger patients (P =.001) and those with more education (P =.02) had higher scores on the Message Facilitation scale. Medical variables were not associated with patients' ratings of the importance of the three subscales. CONCLUSION: Patients rated items addressing the message content as most important, though the supportive and facilitative dimensions were also rated highly. Understanding what is important to patients when told news about their cancer provides valuable information that may help refine how this challenging task is best performed. PMID- 11283137 TI - Complete remission of primary high-grade B-cell gastric lymphoma after cure of Helicobacter pylori infection. AB - PURPOSE: Treatment of low-grade gastric mucosa-associated lymphoid tissue lymphoma by eradication of Helicobacter pylori is reported to result in complete lymphoma remission in approximately 75% of cases. The effect that cure of the infection has on the course of a primary high-grade gastric lymphoma is largely uncertain. The aim of this study was to report the effect of cure of H pylori infection exerted in patients with high-grade B-cell gastric lymphoma. PATIENTS AND METHODS: Eight patients (4 males and 4 females; age range, 26 to 85 years) with H pylori infection and high-grade lymphoma received eradication therapy before planned treatment. The effect of H pylori eradication on the course of high-grade lymphoma was assessed by analysis of surgical specimens (n = 2) or endoscopic biopsies (n = 6). RESULTS: H pylori eradication was successful in all patients and led to complete remission of the lymphoma in seven patients. One patient has experienced partial remission. Two patients were referred to surgery, one of whom (stage II(1E)) had lymph node involvement, and the histologic work-up of the resected stomach revealed residual infiltrates of a low-grade lymphoma, which prompted consolidation chemotherapy. In one patient (initially stage I(1E)), abdominal lymphoma developed 6 months after eradication therapy, which regressed completely after chemotherapy. In four patients, no further treatment was given. Six patients continue in complete remission (range, 6 to 66 months). CONCLUSION: Primary high-grade B-cell gastric lymphoma in stages I(E) through II(E1) associated with H pylori may regress completely after successful cure of the infection. Prospective trials are needed to investigate this treatment in larger numbers of patients. PMID- 11283139 TI - Patients with cancer referred to hospice versus a bridge program: patient characteristics, needs for care, and survival. AB - PURPOSE: The purpose of this study was to compare the characteristics and needs of patients with advanced cancer who were referred to hospice with those referred to a prehospice "bridge" program that is staffed by hospice nurses. PATIENTS AND METHODS: Data were gathered through retrospective review of computerized clinical records using precoded data fields of 284 patients with cancer enrolled in a bridge program and 1,000 who enrolled in a hospice program. Patient characteristics, needs for supportive care at the time of enrollment, and survival were assessed. RESULTS: Bridge patients were less likely to have Medicare or Medicaid (43% v 72%; odds ratio, 0.30; P <.001) and were younger (69 v 73 years, rank sum test; P <.001), more likely to be married (59% v 43%; odds ratio, 1.90; P <.001), and more likely to be in the highest income category (14% v 10%; odds ratio, 1.77; P =.009). Bridge patients had at least as many needs for care as did patients in hospice. Bridge patients lived significantly longer (median, 46 v 19 days; log-rank test of survivor functions, P <.001). CONCLUSION: Patients referred to this bridge program had prognoses that are significantly better than those of patients who enter hospice, but they have needs for supportive care that are at least as great. These findings underscore the importance of initiatives to extend some of the benefits of hospice care to a wider population of patients and should encourage the analysis of similar programs' ability to meet these needs. PMID- 11283140 TI - Self-reported quality of life of individual cancer patients: concordance of results with disease course and medical records. AB - PURPOSE: To investigate the applicability of a standard quality of life (QL) questionnaire to individual cancer patients and to explore the potential for impact of QL information on the process of care by comparing at group level the QL results with the medical records. PATIENTS AND METHODS: One hundred fourteen consecutive patients at the oncology clinics at St James's Hospital, Leeds, United Kingdom, completed the European Organization for the Research and Treatment of Cancer (EORTC) Quality of Life Questionnaire (QLQ)-C30 on a touch screen computer over a 6-month period. The QL results were compared with the corresponding medical records at individual and group level. RESULTS: For individual patients, the serial measurement of QL allowed recognition of patterns over time corresponding to disease course. At group level, a higher proportion of patients reported problems on EORTC QLQ-C30 than were mentioned in the medical records (McNemar paired test, P <.01). Most often clinicians mentioned pain (22% to 39%), and at the initial visit role (66%), and social issues (77%). For the rest of the symptoms and functions, the problems were recorded in between 1% and 25% of the notes, but 20% to 76% of the patients reported QL impairment. Problems that were not recorded in the medical notes tended to be of low severity, with a significant trend observed for pain, fatigue, nausea/vomiting, dyspnea, loss of appetite, and physical function scale (chi(2) test, 11.55 to 34.42, df = 1, P <.001). CONCLUSION: The QL data on individual patients was consistent with the clinical records, thus providing evidence for the validity of these measures in assessment of the individual. The QL profiles had more information on symptoms and particularly on functional issues, such as emotional distress and physical performance. PMID- 11283141 TI - Multicenter phase Ib/II trial of the radiation enhancer motexafin gadolinium in patients with brain metastases. AB - PURPOSE: Motexafin gadolinium is a magnetic resonance imaging (MRI)--detectable redox active drug that localizes selectively in tumor cells and enhances the effect of radiation therapy. This phase Ib/II trial of motexafin gadolinium, administered concurrently with 30 Gy in 10 fractions whole-brain radiation therapy (WBRT), was conducted to determine maximum-tolerated dose (MTD), dose limiting toxicity, pharmacokinetics, and biolocalization in patients with brain metastases. Additional endpoints were radiologic response rate and survival. PATIENTS AND METHODS: Motexafin gadolinium was administered before each radiation treatment in this open-label, multicenter, international trial. In phase Ib, drug dose was escalated until the MTD was exceeded. In phase II, drug was evaluated in a narrow dose range. RESULTS: In phase Ib, the motexafin gadolinium dose was escalated in 39 patients (0.3 mg/kg to 8.4 mg/kg). In phase II, 22 patients received 5 mg/kg to 6.3 mg/kg motexafin gadolinium. Ten once-daily treatments were well tolerated. The MTD was 6.3 mg/kg, with dose-limiting reversible liver toxicity. Motexafin gadolinium's tumor selectivity was established using MRI. The radiologic response rate was 72% in phase II. Median survival was 4.7 months for all patients, 5.4 months for recursive partitioning analysis (RPA) class 2 patients, and 3.8 months for RPA class 3 patients. One-year actuarial survival for all patients was 25%. CONCLUSION: Motexafin gadolinium was well tolerated at doses up to 6.3 mg/kg, was selectively accumulated in tumors, and, when combined with WBRT of 30 Gy in 10 fractions, was associated with a high radiologic response rate. PMID- 11283142 TI - Phase I clinical and pharmacogenetic study of weekly TAS-103 in patients with advanced cancer. AB - PURPOSE: TAS-103 is an inhibitor of both topoisomerase I and II enzymes with broad antitumor activity. It is metabolized to TAS-103-glucuronide (TAS-103-G) predominantly by uridine diphosphate glucuronosyltransferase isoform 1A1 (UGT1A1). We conducted a phase I study to determine the maximum-tolerated dose (MTD) and dose-limiting toxicity (DLT) of TAS-103 when administered on a weekly schedule to patients with advanced cancer. In addition, we evaluated the influence of UGT1A1 genotype on the pharmacokinetics and toxicity of TAS-103. PATIENTS AND METHODS: Thirty-two patients were treated with escalating doses (50 to 200 mg/m(2)) of TAS-103, administered intravenously over 1 hour each week for 3 weeks. Pharmacokinetic analysis was performed at the 130-, 160-, and 200 mg/m(2) dose levels. UGT1A1 genotypes were determined using reverse-transcription polymerase chain reaction techniques. RESULTS: DLT (grade 3 neutropenia) was observed in 5 of 12 patients at 160 mg/m(2) and in 3 of 6 patients at 200 mg/m(2). At 160 mg/m(2), there was a significant correlation between areas under the curve (AUCs) for TAS-103 and TAS-103-G (r = 0.76, P <.05) and an apparent relationship between TAS-103 AUC and D 15 absolute neutrophil count (r = -0.63, P <.05, n = 11, one outlier excluded). UGT1A1 genotype did not influence clearance of TAS-103. CONCLUSION: We recommend a dose of 130 to 160 mg/m(2), or 250 to 300 mg administered using the above weekly schedule for phase II studies. Further studies to characterize the pharmacodynamics and pharmacogenetics of TAS-103 are warranted. PMID- 11283144 TI - Skin lesions in malignancy. Case 1. Chronic myeloid leukemia in lymphoid blast crisis presenting as multiple cutaneous masses. PMID- 11283143 TI - Ondansetron plus metopimazine compared with ondansetron plus metopimazine plus prednisolone as antiemetic prophylaxis in patients receiving multiple cycles of moderately emetogenic chemotherapy. AB - PURPOSE: To compare the antiemetic efficacy and tolerability of ondansetron plus metopimazine with ondansetron plus metopimazine plus prednisolone during nine cycles of moderately emetogenic chemotherapy. PATIENTS AND METHODS: A total of 221 women with stage I or II breast cancer and no prior chemotherapy who were scheduled to receive adjuvant chemotherapy with intravenous cyclophosphamide, fluorouracil and methotrexate or cyclophosphamide, epirubicin, and fluorouracil given every 3 weeks were included in a double-blind parallel trial. Patients were randomized to 3 days of oral treatment with ondansetron plus metopimazine, or ondansetron plus metopimazine plus prednisolone. Ondansetron was administered as 8 mg bid, metopimazine as 30 mg qid, and prednisolone as 50 mg qd. RESULTS: In all, 216 patients (97.7%) were assessable for efficacy during a total of 1,462 cycles. In cycle 1, complete protection from emetic episodes/nausea day 1, days 2 through 5, and days 1 through 5 was achieved in 84.4%/51.4%, 82.6%/41.3%, and 79.8%/34.9% with ondansetron plus metopimazine and in 84.1%/57.0%, 86.8%/53.8%, and 79.4%/43.0% with ondansetron plus metopimazine plus prednisolone, respectively. In cycle 1, the three-drug combination was superior only in the treatment of nausea on days 2 through 5 (P =.0497). The cumulative emetic protection rate after nine cycles was 0.52 with ondansetron plus metopimazine and 0.75 with ondansetron plus metopimazine plus prednisolone. Side effects were generally few and mild with both treatments. Constipation was the only adverse event significantly more frequent with the three-drug combination (P =.029). CONCLUSION: Ondansetron plus metopimazine plus prednisolone is highly effective and superior to ondansetron plus metopimazine during nine cycles of moderately emetogenic chemotherapy. PMID- 11283145 TI - Skin lesions in malignancy. Case 2. Skin metastases from prostate adenocarcinoma. PMID- 11283146 TI - Skin lesions in malignancy. Case 3. Yellow nail syndrome in non-Hodgkin's lymphoma. PMID- 11283147 TI - The narrow path. PMID- 11283148 TI - A comment from a radiotherapeutic point of view regarding the ASCO guidelines on the role of bisphosphonates in breast cancer. PMID- 11283149 TI - Steroidal side effects of exemestane. PMID- 11283150 TI - Does salvage therapy influence outcome? PMID- 11283151 TI - Inhibition of T1/ST2 during respiratory syncytial virus infection prevents T helper cell type 2 (Th2)- but not Th1-driven immunopathology. AB - T cells secreting interleukin (IL)-4 and IL-5 (T helper cell type 2 [Th2] cells) play a detrimental role in a variety of diseases, but specific methods of regulating their activity remain elusive. T1/ST2 is a surface ligand of the IL-1 receptor family, expressed on Th2- but not on interferon (IFN)-gamma-producing Th1 cells. Prior exposure of BALB/c mice to the attachment (G) or fusion (F) protein of respiratory syncytial virus (RSV) increases illness severity during intranasal RSV challenge, due to Th2-driven lung eosinophilia and exuberant Th1 driven pulmonary infiltration, respectively. We used these polar models of viral illness to study the recruitment of T1/ST2 cells to the lung and to test the effects of anti-T1/ST2 treatment in vivo. T1/ST2 was present on a subset of CD4(+) cells from mice with eosinophilic lung disease. Monoclonal anti-T1/ST2 treatment reduced lung inflammation and the severity of illness in mice with Th2 (but not Th1) immunopathology. These results show that inhibition of T1/ST2 has a specific effect on virally induced Th2 responses and suggests that therapy targeted at this receptor might be of value in treating Th2-driven illness. PMID- 11283152 TI - Matrix metalloproteinase 9 protects mice from anti-glomerular basement membrane nephritis through its fibrinolytic activity. AB - Matrix metalloproteinase (MMP)9/gelatinase B is increased in various nephropathies. To investigate its role, we used a genetic approach. Adult MMP9 deficient (MMP9(-/)-) mice showed normal renal histology and function at 3 mo. We investigated the susceptibility of 3-mo-old mice to the accelerated model of anti glomerular basement membrane nephritis, in which fibrin is an important mediator of glomerular injury and renal impairment. Unexpectedly, nephritis was more severe in MMP9(-/)- than in control mice, as attested by levels of serum creatinine and albuminuria, and the extent of crescents and fibrin deposits. Circulating or deposited immunoglobulin G, interleukin (IL)-1beta, or IL-10 were the same in MMP9(-/-) and MMP9(+/+) mice. However, we found that fibrin is a critical substrate for MMP9, and in its absence fibrin accumulated in the glomeruli. These data indicate that MMP9 is required for a novel protective effect on the development of fibrin-induced glomerular lesions. PMID- 11283153 TI - Glucocorticoids attenuate T cell receptor signaling. AB - Glucocorticoids (GCs) affect peripheral immune responses by inhibiting T cell immunity at several stages of the activation cascade, causing impaired cytokine production and effector function. The recent demonstration that the thymic epithelium and possibly thymocytes themselves produce steroids suggests that endogenous GCs also play a role in the control of T cell development. As both peripheral responsiveness and thymic differentiation appear to be regulated by the quantity and quality of intracellular signals issued by antigen-major histocompatibility complex-engaged T cell receptor (TCR) complexes, we investigated the effects of GCs on the signaling properties of T cells stimulated by anti-CD3 monoclonal antibodies or agonist peptides. We demonstrate in this work that dexamethasone, a synthetic GC, inhibits the early signaling events initiated upon TCR ligation, such as tyrosine phosphorylation of several TCR associated substrates including the zeta chain, the ZAP70 kinase, and the transmembrane adapter molecule linker for activation of T cells. Hypophosphorylation was not a consequence of reduced kinase activity of src protein tyrosine kinases, but was correlated with an altered- membrane compartmentalization of these molecules. These observations indicate that in addition to their well-described ability to interfere with the transcription of molecules involved in peripheral responses, GCs inhibit T cell activation by affecting the early phosphorylating events induced after TCR ligation. PMID- 11283154 TI - Autonomous maturation of alpha/beta T lineage cells in the absence of COOH terminal Src kinase (Csk). AB - The deletion of COOH-terminal Src kinase (Csk), a negative regulator of Src family protein tyrosine kinases (PTKs), in immature thymocytes results in the development of alpha/beta T lineage cells in T cell receptor (TCR) beta-deficient or recombination activating gene (rag)-1-deficient mice. The function of Csk as a repressor of Lck and Fyn activity suggests activation of these PTKs is solely responsible for the phenotype observed in csk-deficient T lineage cells. We provide genetic evidence for this notion as alpha/beta T cell development is blocked in lck(-/)-fyn(-/)- csk-deficient mice. It remains unclear whether activation of Lck and Fyn in the absence of Csk uncouples alpha/beta T cell development entirely from engagement of surface-expressed receptors. We show that in mice expressing the alpha/beta TCR on csk-deficient thymocytes, positive selection is biased towards the CD4 lineage and does not require the presence of major histocompatibility complex (MHC) class I and II. Furthermore, the introduction of an MHC class I-restricted transgenic TCR into a csk-deficient background results in the development of mainly CD4 T cells carrying the transgenic TCR both in selecting and nonselecting MHC background. Thus, TCR-MHC interactions have no impact on positive selection and commitment to the CD4 lineage in the absence of Csk. However, TCR-mediated negative selection of csk deficient, TCR transgenic cells is normal. These data suggest a differential involvement of the Csk-mediated regulation of Src family PTKs in positive and negative selection of developing thymocytes. PMID- 11283156 TI - B7-DC, a new dendritic cell molecule with potent costimulatory properties for T cells. AB - Dendritic cells (DCs), unique antigen-presenting cells (APCs) with potent T cell stimulatory capacity, direct the activation and differentiation of T cells by providing costimulatory signals. As such, they are critical regulators of both natural and vaccine-induced immune responses. A new B7 family member, B7-DC, whose expression is highly restricted to DCs, was identified among a library of genes differentially expressed between DCs and activated macrophages. B7-DC fails to bind the B7.1/2 receptors CD28 and cytotoxic T lymphocyte-associated antigen (CTLA)-4, but does bind PD-1, a receptor for B7-H1/PD-L1. B7-DC costimulates T cell proliferation more efficiently than B7.1 and induces a distinct pattern of lymphokine secretion. In particular, B7-DC strongly costimulates interferon gamma but not interleukin (IL)-4 or IL-10 production from isolated naive T cells. These properties of B7-DC may account for some of the unique activity of DCs, such as their ability to initiate potent T helper cell type 1 responses. PMID- 11283155 TI - Attenuation of colon inflammation through activators of the retinoid X receptor (RXR)/peroxisome proliferator-activated receptor gamma (PPARgamma) heterodimer. A basis for new therapeutic strategies. AB - The peroxisome proliferator-activated receptor gamma (PPARgamma) is highly expressed in the colon mucosa and its activation has been reported to protect against colitis. We studied the involvement of PPARgamma and its heterodimeric partner, the retinoid X receptor (RXR) in intestinal inflammatory responses. PPARgamma(1/)- and RXRalpha(1/)- mice both displayed a significantly enhanced susceptibility to 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis compared with their wild-type littermates. A role for the RXR/PPARgamma heterodimer in the protection against colon inflammation was explored by the use of selective RXR and PPARgamma agonists. TNBS-induced colitis was significantly reduced by the administration of both PPARgamma and RXR agonists. This beneficial effect was reflected by increased survival rates, an improvement of macroscopic and histologic scores, a decrease in tumor necrosis factor alpha and interleukin 1beta mRNA levels, a diminished myeloperoxidase concentration, and reduction of nuclear factor kappaB DNA binding activity, c-Jun NH(2)-terminal kinase, and p38 activities in the colon. When coadministered, a significant synergistic effect of PPARgamma and RXR ligands was observed. In combination, these data demonstrate that activation of the RXR/PPARgamma heterodimer protects against colon inflammation and suggest that combination therapy with both RXR and PPARgamma ligands might hold promise in the clinic due to their synergistic effects. PMID- 11283157 TI - Kinetics of acute hepatitis B virus infection in humans. AB - Using patient data from a unique single source outbreak of hepatitis B virus (HBV) infection, we have characterized the kinetics of acute HBV infection by monitoring viral turnover in the serum during the late incubation and clinical phases of the disease in humans. HBV replicates rapidly with minimally estimated doubling times ranging between 2.2 and 5.8 d (mean 3.7 +/- 1.5 d). After a peak viral load in serum of nearly 10(10) HBV DNA copies/ml is attained, clearance of HBV DNA follows a two or three phase decay pattern with an initial rapid decline characterized by mean half-life (t(1/2)) of 3.7 +/- 1.2 d, similar to the t(1/2) observed in the noncytolytic clearance of covalently closed circular DNA for other hepadnaviruses. The final phase of virion clearance occurs at a variable rate (t(1/2) of 4.8 to 284 d) and may relate to the rate of loss of infected hepatocytes. Free virus has a mean t(1/2) of at most 1.2 +/- 0.6 d. We estimate a peak HBV production rate of at least 10(13) virions/day and a maximum production rate of an infected hepatocyte of 200-1,000 virions/day, on average. At this peak rate of virion production we estimate that every possible single and most double mutations would be created each day. PMID- 11283158 TI - CD47-signal regulatory protein alpha (SIRPalpha) regulates Fcgamma and complement receptor-mediated phagocytosis. AB - In autoimmune hemolytic anemia (AIHA), circulating red blood cells (RBCs) opsonized with autoantibody are recognized by macrophage Fcgamma and complement receptors. This triggers phagocytosis and elimination of RBCs from the circulation by splenic macrophages. We recently found that CD47 on unopsonized RBCs binds macrophage signal regulatory protein alpha (SIRPalpha), generating a negative signal that prevents phagocytosis of the unopsonized RBCs. We show here that clearance and phagocytosis of opsonized RBCs is also regulated by CD47 SIRPalpha. The inhibition generated by CD47-SIRPalpha interaction is strongly attenuated but not absent in mice with only residual activity of the phosphatase Src homology 2 domain-containing protein tyrosine phosphatase (SHP)-1, suggesting that most SIRPalpha signaling in this system is mediated by SHP-1 phosphatase activity. The macrophage phagocytic response is controlled by an integration of the inhibitory SIRPalpha signal with prophagocytic signals such as from Fcgamma and complement receptor activation. Thus, augmentation of inhibitory CD47 SIRPalpha signaling may prevent or attenuate RBC clearance in AIHA. PMID- 11283159 TI - L-selectin shedding regulates leukocyte recruitment. AB - The physiologic role of L-selectin shedding is unknown. Here, we investigate the effect of L-selectin shedding on firm adhesion and transmigration. In a tumor necrosis factor alpha-induced model of inflammation, inhibition of L-selectin shedding significantly increased firm adhesion and transmigration by a lymphocyte function-associated antigen (LFA)-1 and intercellular adhesion molecule (ICAM)-1 dependent mechanism. We examined the quality of leukocyte rolling and L-selectin mediated signaling. Blockade of L-selectin shedding significantly reduced the "jerkiness" of leukocyte rolling, defined as the variability of velocity over time. A low level of jerkiness was also observed in the rolling of microbeads conjugated with L-selectin, a model system lacking the mechanism for L-selectin shedding. Inhibition of L-selectin shedding potentiated activation of LFA-1 and Mac-1 induced by L-selectin cross-linking as shown by activation epitope expression and binding of ICAM-1-conjugated beads. We conclude that inhibition of L-selectin shedding increases leukocyte adhesion and transmigration by (a) increasing leukocyte exposure to the inflamed endothelium by decreasing jerkiness and (b) promoting leukocyte activation by outside-in signaling. These observations help to resolve the apparent discrepancy between the minor contribution of L-selectin to rolling and the significant leukocyte recruitment defect in L-selectin knockout mice. PMID- 11283161 TI - V(D)J recombination becomes accessible. PMID- 11283160 TI - Histone acetylation determines the developmentally regulated accessibility for T cell receptor gamma gene recombination. AB - Variable/diversity/joining (V[D]J) recombination of the T cell receptor (TCR) and immunoglobulin (Ig) genes is regulated by chromatin accessibility of the target locus to the recombinase in a lineage- and stage-specific manner. Histone acetylation has recently been proposed as a molecular mechanism underlying the accessibility control. Here, we investigate the role for histone acetylation in the developmentally regulated rearrangements of the mouse TCR-gamma gene, wherein predominant rearrangement is switched from Vgamma3 to Vgamma2 gene during the fetal to adult thymocyte development. Our results indicate that histone acetylation correlates with accessibility, as histone acetylation at the fetal type Vgamma3 gene in accord with germline transcription is relatively high in fetal thymocytes, but specifically becomes low in adult thymocytes within the entirely hyperacetylated locus. Furthermore, inhibition of histone deacetylation during the development of adult bone marrow-derived thymocytes by a specific histone deacetylase inhibitor, trichostatin A, leads to elevated histone acetylation, germline transcription, cleavage, and rearrangement of the Vgamma3 gene. These data demonstrate that histone acetylation functionally determines the chromatin accessibility for V(D)J recombination in vivo and that an epigenetic modification of chromatin plays a direct role in executing a developmental switch in cell fate determination. PMID- 11283162 TI - Investigating the hows and whys of DNA endoreduplication. AB - Endoreduplication is a form of nuclear polyploidization that results in multiple, uniform copies of chromosomes. This process is common in plants and animals, especially in tissues with high metabolic activity, and it generally occurs in cells that are terminally differentiated. In plants, endoreduplication is well documented in the endosperm and cotyledons of developing seeds, but it also occurs in many tissues throughout the plant. It is thought that endoreduplication provides a mechanism to increase the level of gene expression, but the function of this process has not been thoroughly investigated. Numerous observations have been made of endoreduplication, or at least extra cycles of S-phase, as a consequence of mutations in genes controlling several aspects of cell cycle regulation. However, until recently there were few studies directed at the molecular mechanisms responsible for this specialized cell cycle. It is suggested that endoreduplication requires nothing more elaborate than a loss of M-phase cyclin-dependent kinase activity and oscillations in the activity of S-phase cyclin-dependent kinase. PMID- 11283163 TI - Origins and complexes: the initiation of DNA replication. AB - Eukaryotic DNA is organized for replication as multiple replicons. DNA synthesis in each replicon is initiated at an origin of replication. In both budding yeast, Saccharomyces cerevisiae and fission yeast, Schizosaccharomyces pombe, origins contain specific sequences that are essential for initiation, although these differ significantly between the two yeasts with those of S. pombe being more complex then those of S. cerevisiae. However, it is not yet clear whether the replication origins of plants contain specific essential sequences or whether origin sites are determined by features of chromatin structure. In all eukaryotes there are several biochemical events that must take place before initiation can occur. These are the marking of the origins by the origin recognition complex (ORC), the loading onto the origins, in a series of steps, of origin activation factors including the MCM proteins, and the initial denaturation of the double helix to form a replication "bubble". Only then can the enzymes that actually initiate replication, primase and DNA polymerase-alpha, gain access to the template. In many cells this complex series of events occurs only once per cell cycle, ensuring that DNA is not re-replicated within one cycle. However, regulated re-replication of DNA within one cell cycle (DNA endoreduplication) is relatively common in plants, indicating that the "once-per-cycle" controls can be overridden. PMID- 11283164 TI - Identification of causal relationships among traits related to drought resistance in Stylosanthes scabra using QTL analysis. AB - Previous studies have shown that a negative relationship exists between transpiration efficiency (TE) and carbon isotope discrimination (Delta) and between TE and specific leaf area (SLA) in Stylosanthes scabra. A glasshouse experiment was conducted to confirm these relationships in an F(2) population and to study the causal nature of these relationships through quantitative trait loci (QTL) analysis. One hundred and twenty F(2) genotypes from a cross between two genotypes within S. scabra were used. Three replications for each genotype were maintained through vegetative propagation. Water stress was imposed by maintaining plants at 40% of field capacity for about 45 d. To facilitate QTL analysis, a genetic linkage map consisting of 151 RAPD markers was developed. Results from this study show that Delta was significantly and negatively correlated with TE and biomass production. Similarly, SLA showed significant negative correlation with TE and biomass production. Most of the QTL for TE and Delta were present on linkage groups 5 and 11. Similarly, QTL for SLA, transpiration and biomass productivity traits were clustered on linkage groups 13 and 24. One unlinked marker was also associated with these traits. There were several markers coincident between different traits. At all the coincident QTL, the direction of QTL effects was consistent with phenotypic data. At the coincident markers between TE and Delta, high alleles of TE were associated with low alleles of Delta. Similarly, low alleles of SLA were associated with high alleles of biomass productivity traits and transpiration. At the coincident markers between trans-4-hydroxy-N:-methyl proline (MHP) and relative water content (RWC), low alleles of MHP were associated with high alleles of RWC. This study suggests the causal nature of the relationship between TE and Delta. Phenotypic data and QTL data show that SLA was more closely associated with biomass production than with TE. This study also shows that a cause-effect relationship may exist between SLA and biomass production. PMID- 11283165 TI - Auxin and heat shock activation of a novel member of the calmodulin like domain protein kinase gene family in cultured alfalfa cells. AB - A calmodulin like domain protein kinase (CPK) homologue was identified in alfalfa and termed MsCPK3. The full-length sequence of cDNA encoded a 535 amino acid polypeptide with a molecular weight of 60.2 kDa. The deduced amino acid sequence showed all the conserved motifs that define other members of this kinase family, such as serine-threonine kinase domain, a junction region and four potential Ca2+ -binding EF sites. The recombinant MsCPK3 protein purified from E. coli was activated by Ca2+ and inhibited by calmodulin antagonist (W-7) in in vitro phosphorylation assays. The expression of MsCPK3 gene increased in the early phase of the 2,4-D induced alfalfa somatic embryogenesis. Heat shock also activated this gene while kinetin, ABA and NaCl treatment did not result in MsCPK3 mRNA accumulation. The data presented suggest that the new alfalfa CPK differs in stress responses from the previously described homologues and in its potential involvement in hormone and stress-activated reprogramming of developmental pathways during somatic embryogenesis. PMID- 11283166 TI - Ca2+ and phosphate releases from calcified Chara cell walls in concentrated KCl solution. AB - Ca2+ and P(i) (inorganic phosphate) releases from isolated calcified and uncalcified Chara cell walls were measured with a Ca(2+)-selective electrode and colorimetry, and their ionic relations were analysed on the basis of the electroneutrality rule. The results showed that (1) not only Ca2+ but also P(i) can be released from isolated calcified Chara cell walls into pure deionized water and 100 mM KCl solution, and (2) the positive charge due to the Ca2+ released cannot be neutralized only by the negative charge from the simultaneously released P(i). These findings suggest that calcium bands of calcified Chara cell walls are composed of mainly CaCO(3) and CaHPO(4) and some anions other than P(i) should be released simultaneously with the Ca2+ and P(i). More Ca2+ and P(i) can be solubilized from isolated Chara cell walls in 100 mM KCl solution than in pure deionized water. The pH value of 100 mM KCl solution in which isolated uncalcified young Chara cell walls have been immersed is a little lower than that of pure deionized water in which the same isolated uncalcified young Chara cell walls have been immersed, suggesting that some acidic substances are solubilized by 100 mM KCl. To explain this from the viewpoint of solution chemistry, the solubilities of pure CaCO(3) and pure CaHPO(4) in water and 100 mM KCl solution were measured with a Ca2+ -selective electrode and their pH values with a glass pH electrode. The conclusion reached was that the Ca2+ release from isolated Chara cell walls is accompanied by the release of P(i), CO(2-)(3) and acidic substances. This suggests that the so-called calcium bands and/or ionic relations, including ion exchange, in Chara cell walls are chemically or physicochemically more complex than they are currently considered to be. PMID- 11283167 TI - Methyl jasmonate upregulates biosynthetic gene expression, oxidation and conjugation of polyamines, and inhibits shoot formation in tobacco thin layers. AB - The effect of methyl jasmonate (MJ) on de novo shoot formation and polyamine metabolism was investigated in thin layer explants of tobacco (Nicotiana tabacum L. cv. Samsun). A relatively low concentration of MJ (0.1 microM) enhanced explant fresh weight, but had no effect on the final number of shoots per explant while higher concentrations (1 and 10 microM) significantly inhibited organogenesis. The histological study revealed that, with increasing concentrations of MJ, the formation of meristemoids and shoot domes declined and the incidence of cell hypertrophy increased. In explants cultured with 0.1, 1 or 10 microM MJ, the endogenous levels of free putrescine, spermidine and spermine generally declined compared with controls, after 7 and 15 d. Perchloric acid soluble conjugated polyamines accumulated dramatically during culture, but much more so in the presence of MJ than in controls. Acid-insoluble conjugated spermidine alone increased in response to the elicitor. Activities of the putrescine biosynthetic enzymes arginine decarboxylase (ADC, EC 4.1.1.19) and ornithine decarboxylase (ODC, EC 4.1.1.17) in the soluble fraction of MJ-treated explants displayed up to 3-fold increases relative to control explants. However, the most relevant increases in these enzyme activities occurred in the particulate fraction. The activity of S:-adenosylmethionine decarboxylase (SAMDC, EC 4.1.1.21), an enzyme involved in spermidine and spermine biosynthesis, was also stimulated by exposure to MJ. Northern analyses revealed MJ-induced, generally dose-dependent, increases in the mRNA levels of all three enzymes. Diamine oxidase (DAO, EC 1.4.3.6) activity was stimulated by MJ mainly in the cell wall fraction. The upregulation of polyamine metabolism is discussed in relation to the morphogenic behaviour of MJ-treated explants. PMID- 11283168 TI - Endosperm-specific activity of a storage protein gene promoter in transgenic wheat seed. AB - The characterization of the promoter of a wheat (Triticum aestivum) cv. Cheyenne high molecular weight glutenin subunit (HMW subunit) gene, Glu-1D-1 is reported. The nucleotide sequence of the promoter from position -1191 to -650 with respect to the transcription start site was determined, to add to that already determined. Analysis of this region of the promoter revealed the presence of an additional copy of part of the primary enhancer sequence and sequences related to regulatory elements present in other wheat seed protein genes. A chimaeric gene was constructed comprising the 5' flanking region of the Glu-1D-1 gene from position -1191 to +58, the coding region of the UID:A (Gus) gene, and the nopaline synthase (Nos) gene terminator. This chimaeric gene was introduced into wheat (Triticum durum cv. Ofanto) by particle bombardment of inflorescence explants. Two independent transgenic lines were produced, and both showed expression of the Gus gene specifically in the endosperm during mid-development (first detected 10-12 d after anthesis). Histochemical analysis of homozygous T(2) seed confirmed this pattern of expression, and showed that expression was initiated first in the central lobes of the starchy endosperm, and then spread throughout the endosperm tissue, while no expression was detected in the aleurone layer. Native HMW subunit protein was detectable by Western analysis 12-14 d after anthesis, consistent with concurrent onset of activity of the native and introduced HMW subunit gene promoters. PMID- 11283169 TI - Fusicoccin- and IAA-induced elongation growth share the same pattern of K+ dependence. AB - The dependence of growth induced by the fungal toxin fusicoccin (FC) on the K+ content of the incubation medium was investigated in abraded maize coleoptiles. If the divalent ion Ca2+ was included in the bathing medium, no FC-induced growth occurred in the absence of K+, whereas a strong response was detected in presence of K+. The optimal K+ concentration was in the range of 1-10 mM. With the exception of Rb+, none of the other alkali ions (Na+, Li+, Cs+) could replace for K+ in sustaining FC-induced growth. The potassium channel blocker tetraethylammonium (TEA) reversibly inhibited FC-induced growth. As shown earlier for auxin-induced growth, no strict potassium dependence of FC-triggered elongation was observed in Ca2+ -free media. However, TEA abolished this apparently K+ independent FC-induced growth. It is concluded that FC-induced growth, like auxin-induced growth, requires K+ uptake through K+ channels. PMID- 11283170 TI - Hydraulic properties of individual xylem vessels of Fraxinus americana. AB - Studies of the hydraulic properties of xylem vessels have been limited to measurements of whole plant or whole stem segments. This approach allows the longitudinal transport properties of the ensemble of vessels within a stem to be determined, but provides little information on radial transport. Here the xylem of Fraxinus americana L. has been examined using a new method that allows the transport properties of individual vessels to be examined. One goal of this study was to quantify transport parameters relevant to embolism repair. The longitudinal conductivity of vessel segments open at both ends (i.e. no end walls) agreed with values predicted by the Poiseuille equation. Radial specific conductance (conductance per unit area) was approximately six orders of magnitude lower than the longitudinal conductance of the vessel segment normalized by the cross-sectional area of the vessel lumen. There was a step increase in the radial specific conductance of previously gas-filled vessels when the delivery pressure exceeded 0.4 MPa. This is consistent with the idea that positive pressure, required for embolism repair, can be compartmentalized within a vessel if the bordered pit chambers are gas-filled. The diffusion coefficient for the movement of gas from a pressurized air-filled vessel was of the same order of magnitude as that for air diffusing through water (1.95 e(-9) m(2) s(-1)). Estimates of the time needed to displace all of the gas from an air-filled vessel were in the order of 20 min, suggesting that gas removal may not be a major limitation in embolism repair. PMID- 11283171 TI - Restoration of gravitropic sensitivity in starch-deficient mutants of Arabidopsis by hypergravity. AB - Despite the extensive study of plant gravitropism, there have been few experiments which have utilized hypergravity as a tool to investigate gravisensitivity in flowering plants. Previous studies have shown that starch deficient mutants of Arabidopsis are less sensitive to gravity compared to the wild-type (WT). In this report, the question addressed was whether hypergravity could restore the sensitivity of starch-deficient mutants of Arabidopsis. The strains examined include a WT, a starchless mutant and a reduced-starch mutant. Vertical orientation studies with dark-grown seedlings indicate that increased centrifugal acceleration improves orientation relative to the acceleration vector for all strains, even the WT. For starchless roots, growth of seedlings under constant 5 g acceleration was required to restore orientation to the level of the WT at 1 g. In contrast, approximately 10 g was required to restore the orientation of the starchless mutant hypocotyls to a WT level at 1 g. Examination of plastid position in root cap columella cells of the starchless mutant revealed that the restoration of gravitropic sensitivity was correlated with the sedimentation of plastids toward the distal cell wall. Even in WT plants, hypergravity caused greater sedimentation of plastids and improved gravitropic capability. Collectively, these experiments support the hypothesis of a statolith based system of gravity perception in plants. As far as is known, this is the first report to use hypergravity to study the mechanisms of gravitropism in Arabidopsis. PMID- 11283172 TI - Rapid N transport to pods and seeds in N-deficient soybean plants. AB - Non-nodulated soybean (Glycine max (L.) Merr.) plants were cultivated hydroponically under N-sufficient (5 mM NaNO(3)) or N-deficient (0.5 mM NaNO(3)) conditions. (13)N- or (15)N- labelled nitrate was fed to the cut end of the stems, and the accumulation of nitrate-derived N in the pods, nodes and stems was compared. Real-time images of (13)N distribution in stems, petioles and pods were obtained using a Positron Emitting Tracer Imaging System for a period of 40 min. The results indicated that the radioactivity in the pods of N-deficient plants was about 10 times higher than that of N-sufficient plants, although radioactivity in the stems and nodes of N-deficient versus N-sufficient plants was not different. A similar result was obtained by supplying (15)NO(3) to cut soybean shoots for 1 h. The fact that the N translocation into the pods from NO(3) fed to the stem base was much faster in N-deficient plants may be due to the strong sink activity of the pods in N-deficient plants. Alternatively, the redistribution of N from the leaves to the pods via the phloem may be accelerated in N-deficient plants. The temporal accumulation of (13)NO(3) in nodes was suggested in both N-sufficient and N-deficient plants. In one (13)NO(3) pulse chase experiment, radioactivity in the stem declined rapidly after transferring the shoot from the (13)NO(3) solution to non-labelled NO(3); in contrast, the radioactivity in the node declined minimally during the same time period. PMID- 11283173 TI - Abscisic acid induces a decline in nitrogen fixation that involves leghaemoglobin, but is independent of sucrose synthase activity. AB - Sucrose synthase (SS) activity has been suggested to be a key point of regulation in nodule metabolism since this enzyme is down-regulated in response to different stresses which lead to decreased nitrogen fixation. In soybean, a dramatic decline of SS transcripts has been observed within 1 d from the onset of drought. Such a quick response suggests mediation by a signal transduction molecule. Abscisic acid (ABA) is a likely candidate to act as such a molecule as it mediates in a significant number of plant responses to environmental constraints. The hypothesis of ABA controlling nodule metabolism was approached in this work by assessing nodule responses to exogenous ABA supply in pea. Under the experimental conditions, ABA did not affect plant biomass, nodule numbers or dry weight. However, nitrogen fixation rate was reduced by 70% within 5 d and by 80% after 9 d leading to a reduced plant organic nitrogen content. Leghaemoglobin (Lb) content declined in parallel with that of nitrogen fixation. SS activity, however, was not affected by ABA treatment, and neither were the activities of the enzymes aspartate amino transferase, alkaline invertase, malate dehydrogenase, glutamate synthase, uridine diphosphoglucose pyrophosphorylase, isocitrate dehydrogenase, and glutamine synthetase. Nodule bacteroid-soluble protein content was reduced in nodules only after 9 d of ABA treatment. These results do not support the hypothesis that ABA directly regulates SS activity. However, they do suggest the occurrence of at least two different control pathways in nodules under environmental constraints, which include ABA being involved in a Lb/oxygen-related control of nitrogen fixation. PMID- 11283174 TI - Initiation and regulation of water deficit-induced abscisic acid accumulation in maize leaves and roots: cellular volume and water relations. AB - Water deficit-induced ABA accumulation in relation to cellular water relations was investigated in maize root and leaf tissues. While polyethylene glycol (PEG) treatment led to a significant increase of ABA content in both root and leaf tissues, ethylene glycol (EG), a permeable monomer of PEG, had no effect on ABA accumulation at similar or much lower osmotic potentials. A rapid and massive accumulation of ABA in leaf tissues occurred at a specific threshold of PEG 6000 concentration, about 20% (w/v), and closely coincided with the start of the tissue weight loss and the obvious decrease of cellular osmotic potential. Pretreatment with EG lowered the cell sap osmotic potential and also lowered the capability of both root and leaf tissues to accumulate ABA in response to further air-drying or PEG treatment. When samples were dehydrated and incubated under pressure, a method to maintain high water potential and pressure potential during dehydration, ABA accumulation was similar to those dehydrated and incubated under atmospheric pressure. Such results suggest that both the absolute water potential and pressure potential per se had no direct effects on the dehydration-induced ABA accumulation. The results have provided evidence that the initiation of ABA accumulation is related to the weight loss of tissues or changes in cellular volume rather than the cell water relation parameters, and the capability of ABA accumulation can be regulated by cellular osmotic potential. PMID- 11283175 TI - Relationship between changes in the guard cell abscisic-acid content and other stress-related physiological parameters in intact plants. AB - The relationships of guard cell ABA content to eight stress-related physiological parameters were determined on intact Vicia faba L. plants that were grown hydroponically with split-root systems. Continuous stress was imposed by the addition of PEG to part of the root system. The water potentials of roots sampled after the addition of PEG were 0.25 MPa lower than the water potentials of other roots of the same plant, which were similar to the roots of untreated plants. The leaflet water potentials of plants sampled within 2 h of stress imposition were similar to those of control plants. However, leaf conductance was lower in plants sampled after only 20 min of stress imposition, and the root- and leaflet apoplastic ABA concentrations of these plants were higher than those of untreated plants. As the essence of this report, there was a linear relationship between guard cell ABA content and leaf conductance. Leaflet apoplastic ABA concentrations <150 nM were also linearly related to leaf conductance, but higher leaflet apoplastic ABA concentration did not cause equally large further declines in leaf conductance. It is suggested that evaporation from guard cell walls caused ABA to accumulate in the guard cell apoplast and this pool was saturated at high leaflet apoplastic ABA concentrations. PMID- 11283176 TI - Responses of plant growth rate to nitrogen supply: a comparison of relative addition and N interruption treatments. AB - This paper investigates the effects of uptake of nitrate and the availability of internal N reserves on growth rate in times of restricted supply, and examines the extent to which the response is mediated by the different pools of N (nitrate N, organic N and total N) in the plant. Hydroponic experiments were carried out with young lettuce plants (Lactuca sativa L.) to compare responses to either an interruption in external N supply or the imposition of different relative N addition rate (RAR) treatments. The resulting relationships between whole plant relative growth rate (RGR) and N concentration varied between linear and curvilinear (or possibly bi-linear) forms depending on the treatment conditions. The relationship was curvilinear when the external N supply was interrupted, but linear when N was supplied by either RAR methods or as a supra-optimal external N supply. These differences resulted from the ability of the plant to use external sources of N more readily than their internal N reserves. These results show that when sub-optimal sources of external N were available, RGR was maintained at a rate which was dependent on the rate of nitrate uptake by the roots. Newly acquired N was channelled directly to the sites of highest demand, where it was assimilated rapidly. As a result, nitrate only tended to accumulate in plant tissues when its supply was essentially adequate. By comparison, plants forced to rely solely on their internal reserves were never able to mobilize and redistribute N between tissues quickly enough to prevent reductions in growth rate as their tissue N reserves declined. Evidence is presented to show that the rate of remobilization of N depends on the size and type of the N pools within the plant, and that changes in their rates of remobilization and/or transfer between pools are the main factors influencing the form of the relationship between RGR and N concentration. PMID- 11283177 TI - Xylem hydraulic conductivity related to conduit dimensions along chrysanthemum stems. AB - The stem xylem conduit dimensions and hydraulic conductivity of chrysanthemum plants (Dendranthema x grandiflorum Tzvelev cv. Cassa) were analysed and quantified. Simple exponential relations describe conduit length distribution, height dependency of conduit length distribution, and height dependency of stem hydraulic conductivity. These mathematical descriptions can be used to model the xylem water transport system. Within a chrysanthemum stem of 1.0 m, the conduit half-length (the length within which 50% of the conduits have their end) was 0.029 m at soil surface and decreased by half at a height of 0.6 m. With each 0.34 m increase in height up the stem, the hydraulic conductivity decreased by 50%. The resistance calculated from conduit lumen characteristics was 70% of the measured resistance. The remaining unexplained part of the hydraulic resistance is at least partly caused by inter-conduit connections. PMID- 11283178 TI - The effect of phosphorus availability on the carbon economy of contrasting common bean (Phaseolus vulgaris L.) genotypes. AB - A common response to low phosphorus availability is increased relative biomass allocation to roots. The resulting increase in root:shoot ratio presumably enhances phosphorus acquisition, but may also reduce growth rates by diverting carbon to the production of heterotrophic rather than photosynthetic tissues. To assess the importance of increased carbon allocation to roots for the adaptation of plants to low P availability, carbon budgets were constructed for four common bean genotypes with contrasting adaptation to low phosphorus availability in the field ("phosphorus efficiency"). Solid-phase-buffered silica sand provided low (1 microM), medium (10 microM), and high (30 microM) phosphorus availability. Compared to the high phosphorus treatment, plant growth was reduced by 20% by medium phosphorus availability and by more than 90% by low phosphorus availability. Low phosphorus plants utilized a significantly larger fraction of their daytime net carbon assimilation on root respiration (c. 40%) compared to medium and high phosphorus plants (c. 20%). No significant difference was found among genotypes in this respect. Genotypes also had similar rates of P absorption per unit root weight and plant growth per unit of P absorbed. However, P efficient genotypes allocated a larger fraction of their biomass to root growth, especially under low P conditions. Efficient genotypes had lower rates of root respiration than inefficient genotypes, which enabled them to maintain greater root biomass allocation than inefficient genotypes without increasing overall root carbon costs. PMID- 11283179 TI - Effects of calcium on antioxidant activities and water relations associated with heat tolerance in two cool-season grasses. AB - Calcium (Ca2+) may be involved in plant tolerance to heat stress by regulating antioxidant metabolism or/and water relations. This study was designed to examine whether external Ca2+ treatment would improve heat tolerance in two C(3), cool season grass species, tall fescue (Festuca arundinacea L.) and Kentucky bluegrass (Poa pratensis L.), and to determine the physiological mechanisms of Ca2+ effects on grass tolerance to heat stress. Grasses were treated with CaCl(2) (10 mM) or H(2)O by foliar application and then exposed to heat stress (35/30 degrees C) in growth chambers. Some of the Ca2+ -untreated plants were maintained at 20/15 degrees C as the temperature control. Heat stress reduced grass quality, relative water content (RWC), and chlorophyll (Chl) content of leaves in both species, but Ca2+ treatment increased all three factors under heat stress. The Ca2+ concentration in cell saps increased with heat stress and with external Ca2+ treatment in both species. Osmotic potential increased with heat stress, but external Ca2+ treatment had no effect. Osmotic adjustment increased during short term heat stress, but then decreased with a prolonged period of stress; it was not influenced by Ca2+ treatment. The activity of superoxide dismutase (SOD) in both species increased transiently at 12 d of heat stress and then remained at a level similar to that of the control. External Ca2+ treatment had no effect on SOD activity. The activities of catalase (CAT), ascorbate peroxidase (AP), and glutathione reductase (GR) of both species decreased during heat stress. Plants treated with Ca2+ under heat stress had higher CAT, GR and AP activities than untreated plants. Lesser amounts of malondialdehyde (MDA) accumulated in Ca2+ treated plants than in untreated plants during extended periods of heat stress. The results suggested that exogenous Ca2+ treatment enhanced heat tolerance in both tall fescue and Kentucky bluegrass. This enhancement was related to the maintenance of antioxidant activities and a decrease in membrane lipid peroxidation, but not to the regulation of osmotic potential and osmotic adjustment. PMID- 11283180 TI - In situ and in vitro senescence induced by KCl stress: nutritional imbalance, lipid peroxidation and antioxidant metabolism. AB - Sunflower (Helianthus annuus L. cv. SH222) plants and calli were exposed to KCl stress for three weeks. Calli were more tolerant to KCl than plants. KCl stress decreased NO(-)(3), Mn, Fe and B levels in whole plants and P, Ca and Mg in shoots. NO(-)(3), P, Ca, Mg, Mn, and B levels decreased in 100 mM-stressed calli. Chlorophyll content, F:(m) and (F:(m)-F:(0))/F:(m) ratio decreased in stressed leaves, while F:(0) increased only in leaves exposed to severe stress (100 and 150 mM). Membrane permeability and lipid peroxidation increased in plants under all stress conditions and in 100 and 150 mM stressed calli, but remained unchanged in 25 mM stressed calli. Salt stress also induced changes relating to antioxidant enzymes: plants under all stress conditions showed a decrease in catalase, peroxidase and SOD activities. Calli under moderate stress (25 mM KCl) showed an increase of catalase, peroxidase and SOD activities, but the activities of peroxidase and SOD decreased when calli were exposed to higher KCl concentrations. The decrease of antioxidant enzyme activities is in tune with lipid peroxidation and membrane permeability increases. On the other hand, calli adapted for 6 months to 100 mM KCl showed an increase of these enzyme activities compared to unstressed calli, while MDA production and membrane permeability were not significantly affected. PMID- 11283181 TI - Brief exposure to low-pH stress causes irreversible damage to the growing root in Arabidopsis thaliana: pectin-Ca interaction may play an important role in proton rhizotoxicity. AB - The viability of Arabidopsis thaliana (strain Landsberg) roots exposed to a low pH (4.5 or 4.7) solution that contained 100 microM CaCl(2) was examined by staining with fluorescein diacetate-propidium iodide. The elongation zone of growing roots lost viability within 1-2 h following exposure to low pH, but non growing roots showed no damage under the same treatment. Low-pH damage in growing roots was irreversible after 1 h incubation at pH 4.5 as judged by regrowth in growing medium at pH 5.6. Growing lateral roots also lost viability in the same treatment, whereas non-growing lateral roots remained viable during and after the treatment. The low-pH damage was ameliorated by the simultaneous application of calcium, indicating the involvement of a calcium-requiring process in overcoming proton toxicity. At pH 5.0, growing roots required 25 microM of calcium to maintain elongation, and at pH 4.8 and pH 4.5 more than 250 microM and 750 microM, respectively. The low-pH damage was ameliorated by divalent cations in the order of Ba2+, approximately Sr2+>/=Ca2+>Mg2+. The monovalent cation K+ showed no ameliorative effect, but borate showed a strong ameliorative effect with Ca2+. These results indicate that the primary target of proton toxicity may be linked to a disturbance of the stability in the pectic polysaccharide network, where calcium plays a key role in plant roots. PMID- 11283182 TI - A comparison of three methods for determining the stomatal density of pine needles. AB - Alternative methods were compared for determining the stomatal density of needles from two pine species. Densities estimated from air-dried, whole needles using a binocular dissecting scope were compared to densities estimated from vacuum dried, intact needles using a scanning electron microscope and expanded peels (or macerated cuticles) using a compound light microscope. Differences among methods were expected from two sources: (1) expansion and shrinkage as a function of water content, and (2) differences in geometry of the measured surface. Estimates from the dissecting scope were similar to those from scanning electron microscopy (t=0.509, n=21, P:=0.62), presumably because both used dried, but otherwise intact whole needles. Light microscopy estimates, however, were lower than dissecting scope estimates (t=-2.307, n=13, P:=0.04). After adjusting for expansion due to hydration and changes in needle geometry, differences disappeared (t=-1.205, n=13, P:=0.25). These results are an important consideration for researchers reconstructing palaeo-atmospheric conditions and assessing plant response to environmental change. PMID- 11283183 TI - Isolation and expression analysis of gibberellin 20-oxidase homologous gene in apple. AB - To characterize the gibberellin (GA) 20-oxidase gene in apple, the genomic and cDNA clone from "Fuji" apple (accession no. AB037114) was isolated. The deduced amino acid sequence of this cDNA showed 71% and 66% identity to those of GA 20 oxidase cloned from French bean and Arabidopsis, respectively. The transcript of this gene was detected mainly in immature seeds between 1-3 months after full bloom. These results suggested that this apple GA 20-oxidase gene might be involved in GA biosynthesis in developing apple seed. PMID- 11283184 TI - Cloning of a cDNA encoding EIN3-like protein (DC-EIL1) and decrease in its mRNA level during senescence in carnation flower tissues. AB - A cDNA clone encoding a putative EIN3-like protein (DC-EIL1) was obtained from total RNA isolated from senescing carnation (Dianthus caryophyllus L.) petals using RT-PCR and RACE techniques. The cDNA (2382 bp) contained an open reading frame of 1986 bp corresponding to 662 amino acids. The amino acid sequence of the N-terminal half of the protein, ranging from 80-300 amino acid residues, had 84% identity with that of the corresponding regions of Arabidopsis EIN3 and tobacco TEIL, although the overall identity was 49% and 52%, respectively. Northern blot analysis revealed that the amount of mRNA corresponding to DC-EIL1 decreased in flower tissues, especially in petals, during natural senescence and senescence induced by exogenously applied ethylene or ABA. PMID- 11283185 TI - Senescent expression of genes coding collagens, collagen-degrading metalloproteinases, and tissue inhibitors of metalloproteinases in rat vocal folds: comparison with skin and lungs. AB - In humans, vocal tissue stiffness increases with age, suggesting a possible contribution of age-associated variations in vocal fold collagen turnover to voice senescence. The underlying mechanisms remain to be explored. With the use of reverse-transcriptase polymerase chain reaction (RT-PCR), collagen subtypes expressed in rat vocal folds were determined, and messenger RNA (mRNA) levels of collagens (types I, III, IV, and V), collagen-degrading proteinases (collagenase 3, gelatinase A and B), and tissue inhibitors of metalloproteinases (TIMP-1 to TIMP-4) were measured in vocal folds of neonatal, adult, and elderly rats. Collagens I, III-VIII, XV, XVII, and XVIII are abundantly expressed, whereas collagens II, IX, X, and XI are absent in rat vocal folds. Messenger RNA levels of collagens I, III, IV, and V and collagen-degrading proteinases in the vocal folds of the adult rats are significantly lower than those in the neonates. These mRNA levels show further decline in the vocal folds of the elderly rats, but only the decrease in mRNA levels of collagens I and V significantly differ from the adult levels. There are no marked age-related alterations in vocal fold levels of TIMP mRNAs, and the tissue variation in the gene expression of the aforementioned molecules is minute. Rat vocal folds display tissue-specific expression of collagen genes. Diminished gene expression for collagens and proteinases and unchanged gene expression for TIMPs indicate a slowdown in collagen turnover that may increase the cross-linking of collagen molecules. This observation may explain in part the stiffness that occurs with aging in human vocal folds. PMID- 11283186 TI - Free radical defenses in the liver and kidney of human growth hormone transgenic mice: possible mechanisms of early mortality. AB - The long-term effects of growth hormone (GH) administration are unknown. Although limited data on its short-term effects purport health benefits, numerous detrimental effects are the consequence of chronically elevated GH. We used spectrophotometric assay and Western blot to determine the effects of chronic GH excess on hepatic and renal antioxidant enzymes (AOEs) in young and middle-aged PEPCK (phosphoenolpyruvate carboxykinase) hGH (human GH) transgenic mice. In the liver, glutathione peroxidase (GPx) was reduced in transgenics of both age groups, catalase was reduced only in young transgenics, and Cu-Zn superoxide dismutase (SOD) was similar to normal mice, but declined with age. In all groups, hepatic AOE activity correlated significantly with AOE level. In the kidney, AOEs in young transgenics were similar to those of normal mice. However, middle-aged transgenics showed reduced renal SOD and GPx activities when compared with young transgenic or middle-aged normal mice. Similarly, renal SOD and GPx levels in middle-aged transgenics were reduced when compared with those of middle-aged normal mice. AOE activity in the kidney correlated significantly with AOE protein level among middle-aged animals only. These data suggest the following: ((1)) GH excess is associated with early declines in SOD and GPx in the kidney and reductions of hepatic GPx at all ages examined, perhaps increasing the risk of free radical-induced damage to these tissues; ((2)) in the liver of young animals and in the liver and kidney of middle-aged animals, AOE activity reflects the amount of enzyme protein; and ((3)) age-related reductions in GPx in transgenics may be related to the increased incidence of liver tumors and renal failure in these animals. PMID- 11283187 TI - Conditioning of skeletal muscles in adult and old mice for protection from contraction-induced injury. AB - The purpose of this study was to design a conditioning program that protected muscles in both adult and old mice from a protocol of contractions that previously caused a significant number of damaged fibers and a deficit in force. Hind-limb dorsiflexor muscles of adult (7 months) and old (22 months) female B6D2F1 mice were exposed once a week to a protocol of repeated forced stretches while maximally activated in vivo. By week 4, muscles of adult, but not old, mice showed no force deficit. Conditioning was continued for 6 weeks, when both age groups showed no force deficit for two consecutive weeks. Three days after the sixth contraction protocol, when morphological damage and force deficits are most severe, the numbers of damaged fibers in muscles of adult and old mice were not different from those in uninjured control muscles and the force deficits were reduced dramatically compared with unconditioned muscles. We conclude that muscles of both adult and old mice conditioned successfully, but muscles of old mice conditioned more slowly than those of adult mice. PMID- 11283188 TI - Telomere erosion and senescence in human articular cartilage chondrocytes. AB - Aging and the degeneration of articular cartilage in osteoarthritis are distinct processes, but a strong association exists between age and the incidence and prevalence of osteoarthritis. We hypothesized that this association is due to in vivo replicative senescence, which causes age-related declines in the ability of chondrocytes to maintain articular cartilage. For this hypothesis to be tested, senescence-associated markers were measured in human articular chondrocytes from donors ranging in age from 1 to 87 years. These measures included in situ staining for senescence-associated beta-galactosidase activity, (3)H-thymidine incorporation assays for mitotic activity, and Southern blots for telomere length determinations. We found that senescence-associated beta-galactosidase activity increased with age, whereas both mitotic activity and mean telomere length declined. These findings indicate that chondrocyte replicative senescence occurs in vivo and support the hypothesis that the association between osteoarthritis and aging is due in part to replicative senescence. The data also imply that transplantation procedures performed to restore damaged articular surfaces could be limited by the inability of older chondrocytes to form new cartilage after transplantation. PMID- 11283189 TI - Biomarkers of aging: prediction of longevity by using age-sensitive T-cell subset determinations in a middle-aged, genetically heterogeneous mouse population. AB - Seven T-cell subset values were measured in each of 559 mice at 8 months of age, and then again in the 494 animals that reached 18 months of age. The group included virgin males, virgin females, and mated females, and it was produced by using a four-way cross-breeding system that generates genetic heterogeneity equivalent to a very large sibship. An analysis of covariance showed that four T cell subsets-CD4, CD4 memory, CD4 naive, and CD4 cells expressing P:-glycoprotein were significant predictors (p <.003) of longevity when measured at 18 months of age after adjustment for the possible effects of gender and mating. The subset marked by CD4 and P:-glycoprotein expression showed a significant interaction effect: this subset predicted longevity only in males. Among subsets measured when the mice were 8 months of age, only the levels of CD8 memory cells predicted longevity (p =.016); the prognostic value of this subset was largely limited to mated females. A cluster analysis that separated mice into two groups based upon similarity of T-cell subset patterns measured at 18 months showed that these two groups differed in life expectancy. Specifically, mice characterized by relatively low levels of CD4 and CD8 memory cells, high levels of CD4 naive cells, and low levels of CD4 cells with P:-glycoprotein (64% of the total) lived significantly longer (50 days = 6%; p <.0007) than mice in the other cluster. The results are consistent with the hypothesis that patterns of T-cell subsets vary among mice in a manner than can predict longevity in middle age, and they suggest that these subsets may prove to be useful for further studies of the genetics of aging and age-sensitive traits. PMID- 11283190 TI - Bitter-sweet memories: truth and fiction. PMID- 11283191 TI - Flavor enhancement of food improves dietary intake and nutritional status of elderly nursing home residents. AB - BACKGROUND: Taste and smell losses occur with aging. These changes may decrease the enjoyment of food and may subsequently reduce food consumption and negatively influence the nutritional status of elderly persons, especially those who are frail. The objective of this study was to determine if the addition of flavor enhancers to the cooked meals for elderly residents of a nursing home promotes food consumption and provides nutritional benefits. METHODS: We performed a 16 week parallel group intervention consisting of sprinkling flavor enhancers over the cooked meals of the "flavor" group (n = 36) and not over the meals of the control group (n = 31). Measurements of intake of the cooked meals were taken before and after 8 and 16 weeks of intervention. Appetite, daily dietary intake, and anthropometry were assessed before and after the intervention. RESULTS: On average, the body weight of the flavor group increased (+1.1 +/- 1.3 kg; p <.05) compared with that of the control group (-0.3 +/- 1.6 kg; p <.05). Daily dietary intake decreased in the control group (-485 +/- 1245 kJ; p <.05) but not in the flavor group (-208 +/- 1115 kJ; p =.28). Intake of the cooked meal increased in the flavor group (133 +/- 367 kJ; p <.05) but not in the control group (85 +/- 392 kJ). A similar trend was observed for hunger feelings, which increased only in the flavor group. CONCLUSION: Adding flavor enhancers to the cooked meals was an effective way to improve dietary intake and body weight in elderly nursing home residents. PMID- 11283192 TI - Metabolic, psychological, and health correlates of dietary restraint in healthy postmenopausal women. AB - BACKGROUND: Dietary restraint, a term used to describe the intentional control of food intake to prevent weight gain or promote weight loss, is commonly practiced by older adults, but little is known about its effects on physiology and metabolism. METHODS: We therefore compared a wide range of parameters between groups of healthy non-obese postmenopausal women classified psychometrically as unrestrained eaters (body mass index [BMI] 23.8 +/- 0.6 [SEM] kg/m(2), n = 28) or restrained eaters (BMI 24.5 +/- 0.5, n = 39). Measurements were made of reported micronutrient intakes, cardiopulmonary function, hematology, body temperature, skin thickness, bone mass, and immune function; in addition, self-perceived health, mood, and some dimensions of eating behavior were assessed by questionnaire. RESULTS: Macronutrient and micronutrient intakes were not significantly different between restrained and unrestrained eaters reporting energy intake to within 30% of predicted total energy expenditure. Restrained eaters had significantly lower hemoglobin (12.9 +/- 0.1 [SEM] vs 13.2 +/- 0.1 g/dl; p <.05), but values were within the normal range in both groups. In addition, restrained eaters scored significantly higher on the Eating Attitudes Test (p <.01) and drive-for-thinness (p <.001) and maturity fears (p <.05) subscores of the Eating Disorders Inventory, but values were again within the normal range. No other parameter differed significantly between groups. CONCLUSIONS: In this normal-weight population, restrained eating was not associated with detrimental effects in a wide range of physiological, metabolic, and health characteristics. Further work is needed to determine the relevance of these results to the general population. PMID- 11283193 TI - Nutritional parameters, body composition, and progression of disability in older disabled residents living in nursing homes. AB - BACKGROUND: The evaluation of nutritional status is one of the primary components of multidimensional geriatric assessment. We investigated the relationship between some markers of malnutrition and the modifications in functional status in a sample of older disabled residents living in nursing homes. METHODS: Ninety eight subjects who were independent in at least two activities of daily living (ADLs) were enrolled in a 2-year longitudinal study. Anthropometric, nutritional, and metabolic parameters, as well as body composition, were measured at baseline and after 2 years. RESULTS: Deteriorating functional status (> or =2 additional lost ADLs) was associated with baseline albumin levels (Tertile 3 vs Tertile 1; odds ratio [OR] 0.16, 95% confidence interval [CI] 0.04-0.67) and subscapular skinfold thickness (Tertile 3 vs. Tertile 1; OR 0.06, 95% CI 0.006-0.50). After multivariate adjustment, the OR for increasing disability was >4 in subjects with decreasing body cell mass (BCM), compared with subjects with a stable BCM. The degree of BCM reduction was strongly related to the number of additional ADLs lost at follow-up (test for trend, p = .003). CONCLUSIONS: In a sample of older disabled nursing home residents, signs of malnutrition seem to predict further worsening in functional status. Furthermore, BCM declines proportionally to the loss in ADLs, suggesting the existence of a strong relationship between BCM loss and the progressive deterioration of functional status. PMID- 11283194 TI - Hypertension in older adults. AB - The high prevalence of hypertension in older persons (nearly one of two subjects aged 60 years and older) suggests that the recognition and treatment should be a priority for physicians. Although diastolic blood pressure is regarded as an important risk factor, it is now clear that isolated systolic hypertension and elevated pulse pressure also play an important role in the development of cerebrovascular disease, congestive heart failure, and coronary heart disease, which are the major causes of cardiovascular morbidity and mortality in the population aged older than 65 years. Controlled, randomized trials have shown that treatment of systolic as well as systolodiastolic hypertension decreases the incidence of cardiovascular and cerebrovascular complications in older adults. The question of whether treatment of hypertension should be maintained in very old persons, those older than 80 years, is still undecided. PMID- 11283195 TI - Evaluating the effectiveness of a home-based fall risk reduction program for rural community-dwelling older adults. AB - BACKGROUND: We investigated the effectiveness of a low-cost, multifactor fall risk reduction program in a group of rural community-dwelling older adults. The goal of the program was to provide health care workers and communities with a primary prevention tool that can be used to teach seniors about fall-related risks. The long-term goal of this program is to reduce the incidence of falling among community-dwelling older adults. METHODS: Complete data were collected on 37 community-dwelling subjects, aged 67 to 90, who participated in a 10-week fall risk reduction program. The subjects were randomly assigned to an intervention group or to a control group. The intervention group received fall risk education, home-based exercise programming, nutrition counseling, and environmental hazards education. Both groups completed a variety of physiologic, psychometric, and environmental fall-related risk assessments before and after the intervention period. RESULTS: The intervention group showed statistically significant improvement in balance, bicep endurance, lower extremity power, reduction of environmental hazards, falls efficacy, and nutritious food behavior during the study period. CONCLUSIONS: The low-cost, home-based fall risk reduction program for community-dwelling older adults was effective in reducing some of the studied fall-related risk factors over a 10-week period. PMID- 11283196 TI - Stress in caregivers of hospitalized oldest-old patients. AB - BACKGROUND: Stress in caregivers of elderly patients is a well-recognized health care problem. However, little has been published about the stress in caregivers of the oldest-old patients, the most rapidly growing segment of our population. METHODS: A prospective cohort study was conducted in four teaching hospitals. Questionnaires were administered to patients 80 years of age and older and their surrogates (the person who would make decisions if the patient were unable to usually a family member) who identified themselves as the primary caregivers for the patients. Data were abstracted from medical records. RESULTS: Caregivers tended to be female and 50 years of age or older. About one in five described her own health as fair or poor; nearly half of them lived with the patient. About one quarter spent at least 8 h/d caring for the patient, and they had few persons available to help them with care. Most of the caregivers reported mild-to moderate levels of stress. After adjustment, higher stress scores were associated with female caregivers, poorer caregiver health, more hours per day spent caring for the patient, and the presence of patient depression and hearing impairment. CONCLUSION: Stress is common in caregivers of the hospitalized oldest-old patients. Women who are in poor health and spend 8 or more hours every day caring for relatives aged 80 and over are at high risk for caregiver stress. Treatment of patient depression and hearing impairment may ameliorate caregiver stress. PMID- 11283197 TI - High plasma insulin and lipids profile in older individuals: the Italian longitudinal study on aging. AB - BACKGROUND: The inverse relationship of insulin level to high-density lipoprotein (HDL)-cholesterol and its positive association with hypertriglyceridemia has been demonstrated in several studies; however, the relationship of insulin to low density lipoprotein (LDL)-cholesterol in elderly persons is not clear. This study investigates the relationships of fasting plasma insulin and selected metabolic and biological risk factors in an aged population. METHODS: The present study is based on a cross-sectional analysis of the data collected at baseline of the Italian Longitudinal Study on Aging in 1992 on a random sample of 5632 Italians aged 65-84 years. Analyses were performed to compare the distribution of risk factors, such as blood level of lipids, creatinine, albumin, fibrinogen, apolipoprotein A-1 and B, blood pressure, and body mass index (BMI), by quartiles of insulin, in both diabetic and nondiabetic participants. RESULTS: Significantly higher levels of triglycerides and BMI and lower levels of HDL-cholesterol were found in the upper quartile of insulin among nondiabetic individuals. In men, we also found significantly higher levels of systolic and diastolic blood pressure. The same trend for these variables, although not significant for HDL-cholesterol and blood pressure, was seen in diabetic men. In diabetic women, total and LDL cholesterol were significantly lower in the highest insulin quartile (p <.001), while no significant differences were seen in nondiabetic women or in men. We also found higher levels of white blood cells in the highest insulin quartile of diabetic women. CONCLUSIONS: These results, apparently in disagreement with earlier reports on the clustering of cardiovascular disease risk factors in hyperinsulinemic individuals, could be due to the high frequency of chronic inflammation and the high prevalence of urinary infections in older diabetic women. PMID- 11283198 TI - Lower body function and mortality in Mexican American elderly people. AB - BACKGROUND: The purpose of this analysis was to examine the differential impact of performance-based and self-reported lower body measures on 2-year mortality in Mexican American elderly persons. METHODS: Data employed are from the Hispanic Established Population for Epidemiological Studies of the Elderly, a probability survey of 3050 community-dwelling Mexican Americans aged 65 and older from the five Southwestern states interviewed in 1993 and 1994. Of the baseline sample with complete data, 198 persons were confirmed deceased 2 years later. A three task, performance-based, lower body function measure consisting of a short walk, balance, and repeated chair stands tests was used. Self-reported lower body function was measured by a 4-item Activities of Daily Living (ADL) measure involving the lower body. RESULTS: The three-task, lower body function measure was a significant predictor of 2-year mortality. The short walk alone was as predictive as the summary measure. The predictive ability of both measures was minimally reduced by the inclusion of the self-reported ADL measure and life threatening medical conditions. Finally, the ADL measure was not a significant predictor of mortality with all the other variables in the analysis. CONCLUSION: Objective measures of lower body function were significant predicators of mortality in Mexican American elderly persons, as found in the general population. Unlike previous studies, the ADL measure was not an independent predictor of mortality after controlling for the objective measure and other risk factors. Additional research is needed to address why objective measures of function are such strong predictors of death. PMID- 11283199 TI - Validity of the multi-directional reach test: a practical measure for limits of stability in older adults. AB - BACKGROUND: Falls occur not only in the forward direction, but also to the side and backward. The purpose of this study was to develop a portable and valid tool to measure limits of stability in the anterior-posterior and medial-lateral directions. METHODS: Two hundred fifty-four community-dwelling older persons were administered the Berg Balance Test (BBT), the Timed Up & Go Test (TUG), and the Multi-Directional Reach Test (MDRT). For the MDRT, subjects performed maximal reaches with the outstretched arm forward (FR), to the right (RR), to the left (LR), and leaning backward (BR), with feet flat on the floor. Reach was measured by the subject's total hand excursion along a yardstick affixed to a telescoping tripod. RESULTS: Mean scores on the MDRT were FR = 8.89 +/- 3.4 in., BR = 4.64 +/ 3.07 in., RR = 6.15 +/- 2.99 in., and LR = 6.61 +/- 2.88 in. Interclass Correlation (ICC2,1) for the reaches were greater than.92. Reliability analysis (Cronbach's Alpha,.842) demonstrated that directional reaches measure similar but unique aspects of the MDRT. The MDRT demonstrated significant correlation with the BBT sum and significant inverse relationship with the scores on the TUG. Regression analysis revealed that activity level contributed to scores in the forward, right, and left direction and that fear of falling contributed to scores in the backward direction. CONCLUSION: The Multi-Directional Reach Test is an inexpensive, reliable, and valid tool for measuring the limits of stability as derived by reach in four directions. Values obtained on relatively healthy community-dwelling older adults serve as norms for screening patient populations. PMID- 11283200 TI - History and mobility exam index to identify community-dwelling elderly persons at risk of falling. AB - BACKGROUND: Falls are common in community-dwelling elderly persons and are a frequent source of morbidity. Simple indices to prospectively stratify people into categories at different fall-risk would be useful to health care practitioners. Our goal was to develop a fall-risk index that discriminated between people at high and low risk of falling. METHODS: We evaluated the risk of falling over a one-year period in 557 elderly persons (mean age 81.6) living in a retirement community. On the baseline interview, we asked subjects if they had fallen in the previous year and evaluated risk factors in six additional conceptual categories. On the follow-up interview one year later, we again asked subjects if they had fallen in the prior year. We evaluated risk factors in the different conceptual categories and used logistic regression to determine the independent predictors of falling over a one-year period. We used these independent predictors to create a fall-risk index. We compared the ability of a prior falls history with other risk factors and with the combination of a falls history and other risk factors to discriminate fallers from nonfallers. RESULTS: A fall in the previous year (OR = 2.42, 95% CI = 1.49-3.93), a symptom of either balance difficulty or dizziness (OR = 1.83, 95% CI = 1.16-2.89), or an abnormal mobility exam (OR = 2.64, 95% CI = 1.64-4.26) were independent predictors of falling over the subsequent year. These three risk factors together (c statistic =.71) discriminated fallers from nonfallers better than previous history of falls alone (c statistic =.61) or the symptomatic and exam risk factors alone (c statistic =.68). When combined into a risk index, the three independent risk factors stratify people into groups whose risk for falling over the subsequent year ranges from 10% to 51%. CONCLUSION: A history of falling over the prior year, a risk factor that can be obtained from a clinical history (balance difficulty or dizziness), and a risk factor that can be obtained from a physical exam (mobility difficulty) stratify people into groups at low and high risk of falling over the subsequent year. This risk index may provide a simple method of assessing fall risk in community-dwelling elderly persons. However, it requires validation in other subjects before it can be recommended for widespread use. PMID- 11283201 TI - Rett syndrome and the MECP2 gene. PMID- 11283203 TI - Prevalence of mitochondrial DNA mutations in childhood/congenital onset non syndromal sensorineural hearing impairment. AB - Genetic factors are the major causes of childhood hearing impairment. Whereas autosomal recessive mutations account for the majority of prelingual non syndromic sensorineural hearing impairment (NSSHI), the relative contribution of mitochondrial DNA (mtDNA) mutations to childhood onset NSSHI has not been established. We screened 202 subjects with congenital/childhood onset NSSHI, consisting of 110 sporadic cases, 75 sib pairs, and 17 families with affected subjects in more than one generation, in order to determine the prevalence of mtDNA mutations associated with NSSHI.mtDNA mutations were found in three of 10 families (30%) in whom the affected members were related through the maternal lineage. One sporadic case (0.9%) was also found to have a known mtDNA mutation but none was found in the sib pairs. Although the prevalence of mtDNA mutations was low in the group as a whole (2%), we suggest that screening should be considered in cases of childhood hearing impairment when it is progressive and particularly in families where transmission is compatible with maternal inheritance. PMID- 11283202 TI - Angelman syndrome phenotype associated with mutations in MECP2, a gene encoding a methyl CpG binding protein. AB - Angelman syndrome (AS) is a neurodevelopmental disorder characterised by severe mental retardation, absent speech, ataxia, sociable affect, and dysmorphic facial features. Eighty five percent of patients with AS have an identifiable genetic abnormality of chromosome 15q11-13. Mutations within the X linked MECP2 gene have been identified in patients with Rett syndrome (RTT), a neurodevelopmental disorder which affects females almost exclusively and which shares phenotypic overlap with AS. RTT is usually associated with normal development in infancy followed by loss of acquired skills and evolution of characteristic hand wringing movements and episodes of hyperventilation.A panel of 25 female and 22 male patients with a clinical diagnosis of AS and no molecular abnormality of 15q11-13 were screened for MECP2 mutations and these were identified in four females and one male. Following the diagnosis, it was possible to elicit a history of regression in three of these patients, who by then were showing features suggestive of Rett syndrome. In the remaining two subjects the clinical phenotype was still considered to be Angelman-like. These findings illustrate the phenotypic overlap between the two conditions and suggest that screening for MECP2 mutations should be considered in AS patients without a demonstrable molecular or cytogenetic abnormality of 15q11-13. Since MECP2 mutations almost always occur de novo, their identification will substantially affect genetic counselling for the families concerned. PMID- 11283204 TI - Genetic association of an LBP-1c/CP2/LSF gene polymorphism with late onset Alzheimer's disease. AB - OBJECTIVES: The only locus unequivocally associated with late onset Alzheimer's disease (AD) risk is APOE. However, this locus accounts for less than half the genetic variance. A recent study suggested that the A allele of the 3'UTR biallelic polymorphism in the LBP-1c/CP2/LSF gene was associated with reduced AD risk. Samples were diagnosed predominantly by clinical rather than pathological criteria. We have sought to replicate this finding in a series of necropsy confirmed, late onset AD cases and non-demented controls. METHODS: The 3'UTR polymorphism in the LBP-1c/CP2/LSF gene was typed in 216 necropsy confirmed AD cases and 301 non-demented controls aged >73 years. RESULTS: We found different LBP-1c/CP2/LSF allele distributions in our AD cases and controls (p=0.048); the A allele was associated with reduced AD risk. The allele and genotype frequencies observed in our cases and controls were similar to those previously reported. No significant effects emerged when the data were adjusted for age, sex, or apoE epsilon4 carrier status. CONCLUSIONS: Our data support LBP-1c/CP2/LSF as a candidate gene/risk factor for AD and provide justification for future studies to investigate the role of this gene in Alzheimer's disease. PMID- 11283207 TI - No evidence for mosaicism in Silver-Russell syndrome. PMID- 11283205 TI - A polymorphism in the gene for microsomal epoxide hydrolase is associated with pre-eclampsia. AB - OBJECTIVE: Microsomal epoxide hydrolase is an important enzyme involved in the metabolism of endogenous and exogenous toxicants. Polymorphic variants of the human epoxide hydrolase gene vary in enzyme activity. We determined whether genetic variability in the gene encoding for microsomal epoxide hydrolase contributes to individual differences in susceptibility to the development of pre eclampsia with or without the syndrome of Haemolysis, Elevated Liver enzymes, and Low Platelets (HELLP). METHODS: A total of 183 non-pregnant women with a history of pre-eclampsia, 96 of whom had concurrently developed the HELLP syndrome, and 151 healthy female controls were genotyped for the 113Tyr-->His polymorphism in exon 3 and the 139His-->Arg polymorphism in exon 4 of the epoxide hydrolase gene by a polymerase chain reaction-restriction fragment length polymorphism assay. Chi-square analysis was used for statistical evaluation of differences in polymorphic rates. RESULTS: In pre-eclampsia a higher frequency (29%) of the high activity genotype Tyr113 Tyr113 in exon 3 was found as compared to controls (16%, OR 2.0, 95% CI 1.2-3.7). There was no difference between groups for the 139His- >Arg polymorphism. In women with a history of pre-eclampsia, no difference in epoxide hydrolase genotypes was found between women who either did or did not develop the HELLP syndrome. In addition, a significant association was found between predicted EPHX activity and pre-eclampsia. CONCLUSIONS: Women with the high activity genotype in exon 3, which could reflect differences in metabolic activation of endogenous or exogenous toxic compounds, may have enhanced susceptibility to pre-eclampsia. However, polymorphisms in the epoxide hydrolase gene do not seem to influence the risk for concurrent development of the HELLP syndrome. PMID- 11283208 TI - New problems in testing for Huntington's disease: the issue of intermediate and reduced penetrance alleles. PMID- 11283211 TI - Introduction to PET instrumentation. AB - OBJECTIVE: The purpose of this paper is to introduce technologists to the basic principles of PET imaging and to the instrumentation used to acquire PET data. PET imaging is currently being done on a variety of imaging system types, and the technologist will be introduced to these systems and learn about the basic physical image-degrading factors in PET. After reading this article, the technologist should be able to describe the basics of coincidence imaging, identify at least 3 physical degrading factors in PET, and describe 2 different types of PET scanning systems. PMID- 11283212 TI - Single-photon emission computed tomography in the year 2001: instrumentation and quality control. AB - OBJECTIVE: SPECT instrumentation is more complex than that used for whole-body and planar imaging, and requires careful quality control to ensure optimum performance. Conventional and new hybrid SPECT imaging systems (coincidence and SPECT/CT) will be discussed. New imaging detector materials such as LSO and CZT will also be discussed, along with their potential advantages. Finally, basic SPECT quality control will be reviewed. After reading this article, the nuclear medicine technologist should be able to: (a) explain the use of single and multihead gamma cameras for SPECT imaging; (b) have an understanding of the potential of new hybrid SPECT imaging systems; (c) be aware of future developments in SPECT imaging technology; (d) understand the requirements for SPECT quality control, including field uniformity and center of rotation corrections; and (e) explain the benefits of using phantoms to augment SPECT quality control. PMID- 11283213 TI - 99mTc-DMSA absolute uptake: normal pediatric values at 2-4 hours. AB - OBJECTIVE: This study aims to determine if normal absolute uptake values of dimercaptosuccinic acid (DMSA) can be predicted accurately over the optimal time for imaging. METHODS: Eighty-eight normal kidneys were analyzed from 44 children with a median age of 4 y. The mean time between injection and scan was 171.7 min. The absolute uptake of DMSA in milligrams was calculated for each kidney. RESULTS: A strong positive linear relationship was found between the absolute uptake of DMSA and the amount of DMSA injected in milligrams (corr = 0.940 [P< 0.0005]), and the age of the child (corr = 0.770 (P<0.0005)). Multiple linear regression showed that these 2 factors accounted for 92.2% of the change in absolute DMSA uptake. CONCLUSION: The absolute uptake of DMSA can be accurately predicted using a linear regression equation incorporating the amount of DMSA injected in milligrams and the patient's age. Time between injection and scan appears to play little role in the absolute uptake of DMSA over 2-4 h. PMID- 11283214 TI - Lymphoscintigraphy using (99m)Tc filtered sulfur colloid in chylothorax: a case report. AB - OBJECTIVE: A 66-y-old man was diagnosed with esophageal carcinoma and underwent a right thoracotomy and esophagectomy. Postoperatively, a recurring right pleural effusion developed. Because an attempt at lymphangiography failed, lymphoscintigraphy was suggested. Because of the inability to obtain radiolabeled albumin, dextran, or nanocolloid, we used filtered sulfur colloid. (0.1 um). The study confirmed the diagnosis of chylothorax. PMID- 11283215 TI - Idiopathic hepatic uptake of (99m)Tc methylene diphosphonate: a case report. AB - OBJECTIVE: Two sequential (99m)Tc methylene diphosphonate (MDP) scans were performed on a 42-y-old woman with insulin-dependent diabetes mellitus, chronic right pyelonephritis and anemia. The initial scan showed reduced skeletal uptake with intense and diffuse hepatic uptake. Because these findings were similar to those seen when excessive hydrolyzed-reduced (99m)Tc colloids are present in the radiopharmaceutical, the scan was repeated after an adequate time delay. Increased skeletal uptake was evident in the second scan, but the hepatic uptake persisted. Although there are numerous causes of soft tissue activity on (99m)Tc MDP bone scans, the responsible pathologic entity is not always clear. This study reviews several possible reasons for such uptake, although the exact mechanism in this case remains conjectural. PMID- 11283216 TI - Review of computerized NMTCB certification examination. Nuclear Medicine Technology Certification Board. PMID- 11283217 TI - Corporate nuclear medicine: the implementation of a centralized management model. AB - OBJECTIVE: A trend in corporate healthcare is the merging of small community hospitals with larger regional hospitals to expand the patient base. The purpose of this article is to illustrate the benefits of operating several nuclear medicine departments under a centralized management system, rather than operating many decentralized departments. The issues discussed are the development, financial benefits, operations, and structure of a corporate nuclear medicine department. METHODS: Seven nuclear medicine departments were integrated to form one corporate nuclear medicine department from a large hospital organization comprising seven different hospitals. The management team created the concept and advised administration. Training programs were designed and implemented, and committees were formed to ensure the efficient operation of the integrated department. All aspects of the department, such as scheduling and interpretation of studies, are managed at a central location. All technologists rotate to all hospitals. Success was measured by cost savings, study turn-around times, and evaluation of patient and employee satisfaction. RESULTS: It was found that establishing a corporate nuclear medicine department created a greater patient base by servicing a larger geographic area, and resulted in savings of $870,000 annually. Standardizing procedures and protocols allowed for consistency in patient care, an inpatient turnaround time of 24 h, and a dictated report turnaround time of 30 min. Employee relations and satisfaction remained consistent with a 4.76 out of a 5.0 leadership index rating. CONCLUSION: A nuclear medicine department with a centralized management system is a viable option for corporate health care. It is recommended for operations endeavoring to expand the patient base and improve the financial picture. PMID- 11283218 TI - Making slides of nuclear medicine images using a scanner, personal computer, commercial software, and a laser graphic printer. AB - OBJECTIVE: The process of making slides for presentation of nuclear medicine data has historically used methods that require some photographic expertise. This paper describes the use of readily available software and hardware to achieve the same result. Surface images generated for SPECT images of a normal patient and a patient with Alzheimer's dementia were used to develop the technique described. Images were scanned and processed using commercially available software and a personal computer. Slides were generated by a slide printer and compared with electronic presentations of the same images. The method described is easier than the photographic method used in the past, and the quality of the presentation material (either slide or electronic) is readily predictable. PMID- 11283220 TI - What really happens in the SCN at night. PMID- 11283221 TI - Receptor-mediated control of regulatory volume decrease (RVD) and apoptotic volume decrease (AVD). AB - A fundamental property of animal cells is the ability to regulate their own cell volume. Even under hypotonic stress imposed by either decreased extracellular or increased intracellular osmolarity, the cells can re-adjust their volume after transient osmotic swelling by a mechanism known as regulatory volume decrease (RVD). In most cell types, RVD is accomplished mainly by KCl efflux induced by parallel activation of K+ and Cl- channels. We have studied the molecular mechanism of RVD in a human epithelial cell line (Intestine 407). Osmotic swelling results in a significant increase in the cytosolic Ca2+ concentration and thereby activates intermediate-conductance Ca2+-dependent K+ (IK) channels. Osmotic swelling also induces ATP release from the cells to the extracellular compartment. Released ATP stimulates purinergic ATP (P2Y2) receptors, thereby inducing phospholipase C-mediated Ca2+ mobilization. Thus, RVD is facilitated by stimulation of P2Y2 receptors due to augmentation of IK channels. In contrast, stimulation of another G protein-coupled Ca2+-sensing receptor (CaR) enhances the activity of volume-sensitive outwardly rectifying Cl- channels, thereby facilitating RVD. Therefore, it is possible that Ca2+ efflux stimulated by swelling-induced and P2Y2 receptor-mediated intracellular Ca2+ mobilization activates the CaR, thereby secondarily upregulating the volume-regulatory Cl- conductance. On the other hand, the initial process towards apoptotic cell death is coupled to normotonic cell shrinkage, called apoptotic volume decrease (AVD). Stimulation of death receptors, such as TNF receptor and Fas, induces AVD and thereafter biochemical apoptotic events in human lymphoid (U937), human epithelial (HeLa), mouse neuroblastoma x rat glioma hybrid (NG108-15) and rat phaeochromocytoma (PC12) cells. In those cells exhibiting AVD, facilitation of RVD is always observed. Both AVD induction and RVD facilitation as well as succeeding apoptotic events can be abolished by prior treatment with a blocker of volume-regulatory K+ or Cl- channels, suggesting that AVD is caused by normotonic activation of ion channels that are normally involved in RVD under hypotonic conditions. Therefore, it is likely that G protein-coupled receptors involved in RVD regulation and death receptors triggering AVD may share common downstream signals which should give us key clues to the detailed mechanisms of volume regulation and survival of animal cells. In this Topical Review, we look at the physiological ionic mechanisms of cell volume regulation and cell death associated volume changes from the facet of receptor-mediated cellular processes. PMID- 11283222 TI - Two gamma2L subunit domains confer low Zn2+ sensitivity to ternary GABA(A) receptors. AB - The sensitivity of GABAA receptors (GABARs) to Zn2+ inhibition depends on subunit composition. The predominant neuronal forms of mammalian GABARs, alpha(beta)gamma and, alpha(beta)delta are differentially sensitive to Zn2+ inhibition; alpha(beta)gamma receptors are substantially less sensitive than alpha(beta)delta receptors. Recently, functional domains involved in Zn2+ sensitivity have been identified in and subunits. Our aim in the present study was to localize functional domains of low Zn2+ sensitivity within gamma2L subunits. Chimeric subunits were constructed by progressively replacing the rat gamma2L subunit sequence with that of the rat delta subunit sequence. Whole-cell currents were recorded from mouse L929 fibroblasts coexpressing wild-type rat alpha1 and beta3 subunits with a chimeric delta-gamma2L subunit. Unlike alpha and beta subunits, the gamma2L subunit was found to contain a determinant of low Zn2+ sensitivity in the N-terminal extracellular region. In addition, we identified determinants in the M2 segment and the M2-M3 loop of the gamma2L subunit that are homologous to those found in beta and alpha subunits. We postulate that the interface between the latter two domains, which may form the outer vestibule of the channel, represents a single functional domain modulating Zn2+ sensitivity. Thus, the Zn2+ sensitivity of ternary GABARs appears to be determined by two functional domains, one in the N-terminal extracellular region and one near the outer mouth of the channel. PMID- 11283223 TI - Visualization of elementary mechanosensitive Ca2+-influx events, Ca2+ spots, in bovine lens epithelial cells. AB - Local increases in the intracellular Ca2+ concentration ([Ca2+]i) in several regions within the bovine lens epithelial cell during application of mechanical stress were clearly visualized in the presence of lysophosphatidic acid (LPA), a bioactive lysophospholipid, using real-time confocal microscopy. We called the phenomenon 'Ca2+ spots'. Ca2+ spots started in a circular area with a radius of about 1.5 m. These Ca2+ spots spread concentrically, resulting in a mean global increase in [Ca2+]i. The local increase often occurred in a stepwise manner or repetitively at the same region. The spatiotemporal properties of the Ca2+ spots were completely different from those of the Ca2+ wave induced by ATP, a Ca2+ mobilizing agonist. Ca2+ spots were inhibited by decreasing the extracellular Ca2+ concentration or by the presence of Gd3+, an inhibitor of mechanosensitive (MS) channels, but not by thapsigargin, an inhibitor of the endoplasmic reticulum Ca2+ pump, suggesting that Ca2+ spots arise from Ca2+ influx through Gd3+ sensitive MS channels. On the assumption that, in lens epithelial cells, the open probability of the MS channel is 0.4, the membrane potential is 56 mV and the channel conductance is 50 pS, the estimated maximum flux of Ca2+ in a Ca2+ spot (0.4 x 10-17 to 4.7 x 10-17 mol x s(-1)) was comparable to currents of one or a few MS channels. On real-time three-dimensional confocal imaging analysis, which permitted simultaneous imaging of basal and apical planes of cells at 37.6 ms intervals, Ca2+ spots on the apical plane were more clearly visualized than those on the basal plane. From these results, we propose that the Ca2+ spot is an elementary Ca2+-influx event through MS channels directly coupled with the first step in mechanoreception In addition, our results strongly suggest that LPA functions as an endogenous factor affecting mechanotransduction systems. PMID- 11283224 TI - Intracellular calcium reduces light-induced excitatory post-synaptic responses in salamander retinal ganglion cells. AB - The whole-cell patch clamp technique was used to study the effect of intracellular Ca2+ on light-evoked EPSCs in on-off ganglion cells in salamander retinal slices. Both AMPA and NMDA receptors contributed to the light-evoked responses. In the presence of strychnine and picrotoxin, ganglion cells responded to light onset and offset with transient inward currents at -70 mV. These currents were reduced by 35 +/- 3 % when the light stimulus was preceded by a depolarizing step from -70 to 0 mV. The inhibitory effect of depolarization on light-evoked EPSCs was strongly reduced in the presence of 10 mM BAPTA. The degree of EPSC inhibition by the prepulse holding potential followed the current voltage relationship of the Ca2+ current found in the ganglion cell. In the presence of the NMDA receptor antagonist AP-7, glutamate-dependent current was nearly abolished when high Ca2+ was substituted for high Na+ solution. The release of Ca2+ from internal stores by caffeine or inositol trisphosphate reduced the EPSCs by 36 +/- 5 and 38 +/- 11 %, respectively, and abolished the inhibitory effect of depolarization. The inhibitory effect of depolarization on EPSCs was reduced 5-fold in the presence of AP-7, but was not reduced by the AMPA receptor antagonist CNQX. Neither inhibition of Ca2+-calmodulin-dependent enzymes, nor inhibition of protein kinase A or C had any significant effect on the depolarization-induced inhibition of EPSCs. Our data suggest that elevation of [Ca2+]i, through voltage-gated channels or by release from intracellular stores, reduced primarily the NMDA component of the light-evoked EPSCs. PMID- 11283225 TI - An examination of the secretion-like coupling model for the activation of the Ca2+ release-activated Ca2+ current I(CRAC) in RBL-1 cells. AB - One popular model for the activation of store-operated Ca2+ influx is the secretion-like coupling mechanism, in which peripheral endoplasmic reticulum moves to the plasma membrane upon store depletion thereby enabling inositol 1,4,5 trisphosphate (InsP3) receptors on the stores to bind to, and thus activate, store-operated Ca2+ channels. This movement is regulated by the underlying cytoskeleton. We have examined the validity of this mechanism for the activation of I(CRAC), the most widely distributed and best characterised store-operated Ca2+ current, in a model system, the RBL-1 rat basophilic cell line. Stabilisation of the peripheral cytoskeleton, disassembly of actin microfilaments and disaggregation of microtubules all consistently failed to alter the rate or extent of activation of I(CRAC). Rhodamine-phalloidin labelling was used wherever possible, and revealed that the cytoskeleton had been significantly modified by drug treatment. Interference with the cytoskeleton also failed to affect the intracellular calcium signal that occurred when external calcium was re-admitted to cells in which the calcium stores had been previously depleted by exposure to thapsigargin/ionomycin in calcium-free external solution. Application of positive pressure through the patch pipette separated the plasma membrane from underlying structures (cell ballooning). However, I(CRAC) was unaffected irrespective of whether cell ballooning occurred before or after depletion of stores. Pre treatment with the membrane-permeable InsP3 receptor antagonist 2-APB blocked the activation of I(CRAC). However, intracellular dialysis with 2-APB failed to prevent I(CRAC) from activating, even at higher concentrations than those used extracellularly to achieve full block. Local application of 2-APB, once I(CRAC) had been activated, resulted in a rapid loss of the current at a rate similar to that seen with the rapid channel blocker La3+. Studies with the more conventional InsP3 receptor antagonist heparin revealed that occupation of the intracellular InsP3-sensitive receptors was not necessary for the activation or maintenance of I(CRAC). Similarly, the InsP3 receptor inhibitor caffeine failed to alter the rate or extent of activation of I(CRAC). Exposure to Li+, which reduces InsP3 levels by interfering with inositol monophosphatase, also failed to alter I(CRAC). Caffeine and Li+ did not affect the size of the intracellular Ca2+ signal that arose when external Ca2+ was re-admitted to cells which had been pre exposed to thapsigargin/ionomycin in Ca2+-free external solution. Our findings demonstrate that the cytoskeleton does not seem to regulate calcium influx and that functional InsP3 receptors are not required for activation of I(CRAC). If the secretion-like coupling model indeed accounts for the activation of I(CRAC) in RBL-1 cells, then it needs to be revised significantly. Possible modifications to the model are discussed. PMID- 11283226 TI - Direct autocrine inhibition and cAMP-dependent potentiation of single L-type Ca2+ channels in bovine chromaffin cells. AB - Using the cell-attached recording configuration, we found that in adult bovine chromaffin cells there exists a direct membrane-delimited inhibition of single Bay K-modified L-channels mediated by opioids and ATP locally released in the recording pipette. This autocrine modulation is mediated by pertussis toxin (PTX) sensitive G-proteins and causes a 50 % decrease of the open channel probability (Po) and an equivalent percentage increase of null sweeps at +10 mV with no changes to the activation kinetics, single channel conductance and mean open time. The decrease in Po is mainly due to an increase in the occurrence and duration of slow closed times (> 40 ms). Addition of purinergic and opioidergic antagonists (suramin and naloxone) or cell pre-treatment with PTX removes the inhibition while addition of ATP and opioids inside the pipette, but not outside, mimics the effect. Strong pre-pulses (+150 mV, 280 ms) followed by short repolarizations are unable to remove the inhibition at test potential (+10 mV). Increasing the level of cAMP by either direct application of 8-(4 chlorophenylthio)-cAMP (8-CPT-cAMP) or mixtures of forskolin and 1-methyl-3 isobutylxanthine (IBMX) potentiates the activity of L-channels by increasing the mean open time and decreasing the mean closed time and percentage of null sweeps. The cAMP-induced potentiation occurs regardless of whether the G-protein-mediated inhibition is activated by ATP and opioids or inactivated by PTX. Protein kinase inhibitors (H7 and H89) prevent the effects of cAMP without altering the basal autocrine modulation associated with PTX-sensitive G-proteins. Our results provide new evidence for the coexistence of two distinct modulations that may converge on the same neuroendocrine L-channel: a direct G-protein-dependent inhibition and a cAMP-mediated potentiation, which may work in combination to regulate Ca2+ entry during neurosecretion. PMID- 11283227 TI - Volatile anaesthetic effects on Na+-Ca2+ exchange in rat cardiac myocytes. AB - We examined the influence of two clinically relevant concentrations (1 and 2 MAC (minimum alveolar concentration)) of halothane and sevoflurane on both efflux and reverse modes of Na+-Ca2+ exchange (NCX) in enzymatically dissociated adult rat cardiac myocytes. We hypothesised that a volatile anaesthetic-induced decrease in myocardial contractility is mediated by a reduction in intracellular calcium concentration ([Ca2+]i) via inhibition of NCX. Cells were exposed to cyclopiazonic acid and zero extracellular Na+ and Ca2+ to block sacroplasmic reticulum (SR) re-uptake and NCX efflux, respectively. As [Ca2+]i increased under these conditions, extracellular Na+ was rapidly (< 300 ms) reintroduced in the presence or absence of a volatile anaesthetic to selectively promote Ca2+ efflux via NCX. Other cells exposed to cyclopiazonic acid and ryanodine to inhibit SR Ca2+ re-uptake and release were Na+ loaded in zero extracellular Ca2+. The reintroduction of extracellular Ca2+ was used to selectively activate Ca2+ influx via NCX. Compared to controls, both 1 and 2 MAC halothane as well as sevoflurane reduced NCX-mediated efflux. The reduction in NCX-mediated influx was concentration dependent, but comparable between the two anaesthetics. Both anaesthetics at each concentration also shifted the relationship between extracellular Na+ (or extent of Na+ loading) and NCX-mediated efflux (or influx) to the right. These data indicate that despite inhibition of NCX-mediated Ca2+ efflux, volatile anaesthetics produce myocardial depression. However, the inhibition of NCX-mediated Ca2+ influx may contribute to decreased cardiac contractility. The overall effect of volatile anaesthetics on the [Ca2+]i profile is likely to be determined by the relative contributions of influx vs. efflux via NCX during each cardiac cycle. PMID- 11283228 TI - Oxygen-evoked Na+ transport in rat fetal distal lung epithelial cells. AB - Monolayer cultures of rat fetal distal lung epithelial (FDLE) cells generated larger spontaneous short circuit currents (ISC) when maintained (48 h) at neonatal alveolar PO2 (100 mmHg) than at fetal PO2 (23 mmHg). When cells were shifted between these atmospheres in order to impose a rise in PO2 equivalent to that seen at birth, no rise in ISC was seen after 6 h but the response was fully established by 24 h. Studies of basolaterally permeabilised cells revealed a small rise in apical Na+ conductance (GNa) 6 h after PO2 was raised but no further change had occurred by 24 h. A substantial rise was, however, seen after 48 h. Reporter gene assays showed that no activation of the -ENaC (epithelial Na+ channel -subunit) promoter was discernible 24 h after PO2 was raised but increased transcriptional activity was seen at 48 h. Studies of apically permeabilised cells showed that a small rise in Na+ pump capacity was evident 6 h after PO2 was raised and, in common with the rise in ISC, this effect was fully established by 24 h. The rise in ISC thus develops 6-24 h after PO2 is raised and is due, primarily, to increased Na+ pump capacity. The increase in GNa thus coincides with activation of the -ENaC promoter but these effects occur after the rise in ISC is fully established and so cannot underlie this physiological response. The increased transcription may be an adaptation to increased Na+ transport and not its cause. PMID- 11283229 TI - Comparison of cloned Kir2 channels with native inward rectifier K+ channels from guinea-pig cardiomyocytes. AB - The aim of the study was to compare the properties of cloned Kir2 channels with the properties of native rectifier channels in guinea-pig (gp) cardiac muscle. The cDNAs of gpKir2.1, gpKir2.2, gpKir2.3 and gpKir2.4 were obtained by screening a cDNA library from guinea-pig cardiac ventricle. A partial genomic structure of all gpKir2 genes was deduced by comparison of the cDNAs with the nucleotide sequences derived from a guinea-pig genomic library. The cell-specific expression of Kir2 channel subunits was studied in isolated cardiomyocytes using a multi cell RT-PCR approach. It was found that gpKir2.1, gpKir2.2 and gpKir2.3, but not gpKir2.4, are expressed in cardiomyocytes. Immunocytochemical analysis with polyclonal antibodies showed that expression of Kir2.4 is restricted to neuronal cells in the heart. After transfection in human embryonic kidney cells (HEK293) the mean single-channel conductance with symmetrical K+ was found to be 30.6 pS for gpKir2.1, 40.0 pS for gpKir2.2 and 14.2 pS for Kir2.3. Cell-attached measurements in isolated guinea-pig cardiomyocytes (n = 351) revealed three populations of inwardly rectifying K+ channels with mean conductances of 34.0, 23.8 and 10.7 pS. Expression of the gpKir2 subunits in Xenopus oocytes showed inwardly rectifying currents. The Ba2+ concentrations required for half-maximum block at -100 mV were 3.24 M for gpKir2.1, 0.51 M for gpKir2.2, 10.26 M for gpKir2.3 and 235 M for gpKir2.4. Ba2+ block of inward rectifier channels of cardiomyocytes was studied in cell-attached recordings. The concentration and voltage dependence of Ba2+ block of the large-conductance inward rectifier channels was virtually identical to that of gpKir2.2 expressed in Xenopus oocytes. Our results suggest that the large-conductance inward rectifier channels found in guinea-pig cardiomyocytes (34.0 pS) correspond to gpKir2.2. The intermediate-conductance (23.8 pS) and low-conductance (10.7 pS) channels described here may correspond to gpKir2.1 and gpKir2.3, respectively. PMID- 11283230 TI - Somatostatin activates two types of inwardly rectifying K+ channels in MIN-6 cells. AB - Western blotting revealed the presence of five somatostatin receptor types, sst1, sst2, sst3, sst4 and sst5, in the mouse pancreatic -cell line MIN-6. In MIN-6 cells, glucose-induced electrical activity was potently (pEC50 = 12.7) and irreversibly reduced by somatostatin (SRIF-14); this was associated with hyperpolarization of the membrane potential (pEC50 = 11.2) and a decrease in the input resistance (pEC50 = 12.7). The effects of SRIF-14 were mimicked by 100 nM L 362,855 (a partial agonist at sst5 receptors), but not BIM-23027 or NNC-26,9100 (selective agonists at sst2 and sst4 receptors, respectively). CH-275 at 100 nM (a selective agonist at sst1 receptors) partially inhibited electrical activity but without membrane potential hyperpolarization. One hundred nanomolar SRIF-28 activated an inwardly rectifying K+ current (ISRIF) ISRIF was activated neither by 1 M BIM-23056 nor CYN-154806 (antagonists at sst5 and sst2 receptors, respectively). The activation of ISRIF by 100 nM SRIF-28 was, however, inhibited 93 % by BIM-23056; CYN-154806 had no effect. Both 100 nM glibenclamide and 200 M tolbutamide, blockers of the -cell ATP-sensitive K+ channel (K-ATP), reduced ISRIF by ~44 %, whereas 1 mM Ba2+ abolished ISRIF. In cell-attached patches, 100 nM SRIF-14 activated two types of single-channel currents whose properties were consistent with those of K-ATP and GIRK channels. In conclusion, somatostatin can inhibit glucose-induced electrical activity in MIN-6 cells by the combined activation of K-ATP and GIRK channels. Studies with selective agonists and antagonists are consistent with this effect being mediated by the sst5 receptor. PMID- 11283231 TI - Modulation of rat erg1, erg2, erg3 and HERG K+ currents by thyrotropin-releasing hormone in anterior pituitary cells via the native signal cascade. AB - The mechanism of thyrotropin-releasing hormone (TRH)-induced ether-a-go-go related gene (erg) K+ current modulation was investigated with the perforated patch whole-cell technique in clonal somatomammotroph GH3/B6 cells. These cells express a small endogenous erg current known to be reduced by TRH. GH3/B6 cells were injected with cDNA coding for rat erg1, erg2, erg3 and HERG K+ channels. The corresponding erg currents were isolated with the help of the specific erg channel blockers E-4031 and dofetilide and their biophysical properties were determined. TRH (1 M) was able to significantly reduce the different erg currents. The voltage dependence of activation was shifted by 15 mV (erg1), 10 mV (erg2) and 6 mV (erg3) to more positive potentials without strongly affecting erg inactivation. TRH reduced the maximal available erg current amplitude by 12% (erg1), 13% (erg2) and 39% (erg3) and accelerated the time course of erg1 and erg2 channel deactivation, whereas erg3 deactivation kinetics were not significantly altered. The effects of TRH on HERG currents did not differ from those on its rat homologue erg1. In addition, coinjection of rat MiRP1 with HERG cDNA did not influence the TRH-induced modulation of HERG channels. Rat erg1 currents recorded in the cell-attached configuration were reduced by application of TRH to the extra-patch membrane in the majority of the experiments, confirming the involvement of a diffusible second messenger. Application of the phorbol ester phorbol 12-myristate 13-acetate (PMA; 1 M) shifted the voltage dependence of erg1 activation in the depolarizing direction, but it did not reduce the maximal current amplitude. The voltage shift could not be explained by a selective effect on protein kinase C (PKC) since the PKC inhibitor bisindolylmaleimide I did not block the effects of TRH and PMA on erg1. In addition, cholecystokinin, known to activate the phosphoinositol pathway similarly to TRH, did not significantly affect the erg1 current. Various agents interfering with different known TRH-elicited cellular responses were not able to completely mimic or inhibit the TRH effects on erg1. Tested substances included modulators of the cAMP-protein kinase A pathway, arachidonic acid, inhibitors of tyrosine kinase and mitogen-activated protein kinase, sodium nitroprusside and cytochalasin D. The results demonstrate that all three members of the erg channel subfamily are modulated by TRH in GH3/B6 cells. In agreement with previous studies on the TRH-induced modulation of the endogenous erg current in prolactin secreting anterior pituitary cells, the TRH effects on overexpressed erg1 channels are not mediated by any of the tested signalling pathways. PMID- 11283232 TI - Protein kinase C isoform-dependent modulation of ATP-sensitive K+ channels during reoxygenation in guinea-pig ventricular myocytes. AB - ATP-sensitive K+ (KATP) channels activated by glucose-free anoxia close immediately upon reoxygenation in single guinea-pig ventricular myocytes, while KATP channels open persistently during reperfusion in coronary-perfused guinea pig ventricular myocardium. To investigate the reasons behind this discrepancy, we investigated whether protein kinase C (PKC) modulates the opening of KATP channels during anoxia-reoxygenation and ischaemia-reperfusion. Exposure of guinea-pig ventricular cells to glucose-free anoxia shortened the action potential duration at 90% repolarisation (APD90) and evoked the glibenclamide sensitive robust outward current (IK,ATP). Subsequent reoxygenation caused an immediate prolongation of APD90 and a decrease in IK,ATP within approximately 20 s. When the novel (Ca2+-independent) PKC was activated by applying 1,2-dioctanoyl sn-glycerol (1,2DOG, 20 M) with EGTA (20 mM) in the pipette, the APD90 restored gradually after reoxygenation and the extent of recovery was appoximately 80% of the pre-anoxic value. Moreover, IK,ATP decreased slowly and remained opened for up to approximately 4 min after reoxygenation. These results suggest persistent opening of KATP channels during reoxygenation. The persistent activation of KATP channels was augmented when both novel and conventional (Ca2+-dependent) isoforms of PKC were activated by applying 1,2DOG without EGTA in the pipette. In coronary perfused right ventricular myocardium, APD90 remained shortened for up to approximately 30 min of reperfusion. The gradual restoration of APD90 after ischaemia-reperfusion was facilitated by the KATP channel blocker glibenclamide and by the potent PKC inhibitor chelerythrine. Our results provide the first evidence that PKC activation contributes to the persistent opening of KATP channels during reoxygenation and reperfusion. We also conclude that both novel and conventional PKC isoforms co-operatively modulate the opening of KATP channels during the early phase of reoxygenation. PMID- 11283233 TI - Sex differences in the acetylcholine receptor kinetics of postnatal and denervated rat muscle. AB - Single-channel recording from visualised endplates in freshly dissociated muscles from postnatal and denervated rat muscle revealed the presence of a low conductance, fetal type of acetylcholine receptor. Kinetic analysis showed a main component in the burst durations with a mean of 10.8 +/- 2.7 ms (n = 29). Receptors from female rats had an additional 27.3 +/- 5.5 ms (n = 5) kinetic component which was found in one-third of the 15 female endplates. Recordings from male and female denervated muscles gave more homogeneous kinetics with single time constants of 7.2 +/- 1.3 and 7.4 +/- 1.3 ms, respectively. It is concluded that the acetylcholine receptor channels present during early development are different from those of denervated muscle. PMID- 11283234 TI - Enhanced NMDA receptor activity in retinal inputs to the rat suprachiasmatic nucleus during the subjective night. AB - Circadian oscillator mechanisms in the suprachiasmatic nucleus (SCN) can be reset by photic input, which is mediated by glutamatergic afferents originating in the retina. A key question is why light can only induce phase shifts of the biological clock during a restricted period of the circadian cycle, namely the subjective night. One of several possible mechanisms holds that glutamatergic transmission at retinosuprachiasmatic synapses would be altered, in particular the contribution of glutamate receptor subtypes to the postsynaptic response. By studying the contributions of NMDA and non-NMDA glutamate receptors to the retinal input to SCN in whole-cell patch-clamp recordings in acutely prepared slices, we tested the hypothesis that NMDA receptor current evoked by optic nerve activity is potentiated during the subjective night. During the day the NMDA component of the EPSC evoked by optic nerve stimulation was found less frequently and was significantly smaller in magnitude than during the night. In contrast, the non-NMDA component did not show a significant day-night difference. When the magnitude of the NMDA component was normalized to that of the non-NMDA component, the day-night difference was maintained, suggesting a selective potentiation of NMDA receptor conductance. In addition to contributing to electrically evoked EPSCs, the NMDA receptor was found to sustain a small, tonically active inward current during the night phase. No significant tonic contribution by NMDA receptors was detected during the day. These results suggest, first, a dual mode of NMDA receptor function in the SCN and, second, a clock-controlled type of receptor plasticity, which may gate the transfer of photic input to phase shifting mechanisms operating at the level of molecular autoregulatory feedback loops. PMID- 11283235 TI - Initiation of network bursts by Ca2+-dependent intrinsic bursting in the rat pilocarpine model of temporal lobe epilepsy. AB - Chronically epileptic rats, produced by prior injection of pilocarpine, were used to investigate whether changes in intrinsic neuronal excitability may contribute to the epileptogenicity of the hippocampus in experimental temporal lobe epilepsy (TLE). Paired extra-/intracellular electrophysiological recordings were made in the CA1 pyramidal layer in acute hippocampal slices prepared from control and epileptic rats and perfused with artificial cerebrospinal fluid (ACSF). Whereas orthodromic activation of CA1 neurons evoked only a single, stimulus-graded population spike in control slices, it produced an all-or-none burst of population spikes in epileptic slices. The intrinsic firing patterns of CA1 pyramidal cells were determined by intrasomatic positive current injection. In control slices, the vast majority (97%) of the neurons were regular firing cells. In epileptic slices, only 53% the pyramidal cells fired in a regular mode. The remaining 47% of the pyramidal cells were intrinsic bursters. These neurons generated high-frequency bursts of three to six spikes in response to threshold depolarizations. A subgroup of these neurons (10.1% of all cells) also burst fired spontaneously even after suppression of synaptic activity. In epileptic slices, burst firing in most cases (ca 70%) was completely blocked by adding the Ca2+ channel blocker Ni2+ (1 mM) to, or removing Ca2+ from, the ACSF, but not by intracellular application of the Ca2+ chelater 1,2-bis(o-aminophenoxy)ethane N,N,N ',N '-tetra-acetic acid (BAPTA), suggesting it was driven by a Ca2+ current. Spontaneously recurring population bursts were observed in a subset of epileptic slices. They were abolished by adding 2 M 6-cyano-7-nitro-quinoxaline 2,3-dione (CNQX) to the ACSF, indicating that synaptic excitation is critical for the generation of these events. All sampled pyramidal cells fired repetitively during each population burst. The firing of spontaneously active bursters anteceded the population discharge, whereas most other pyramidal cells began to fire conjointly with the first population spike. Thus, spontaneous bursters are likely to be the initiators of spontaneous population bursts in epileptic slices. The dramatic up-regulation of intrinsic bursting in CA1 pyramidal cells, particularly the de novo appearance of Ca2+-dependent bursting, may contribute to the epileptogenicity of the hippocampus in the pilocarpine model of TLE. These findings have important implications for the pharmacological treatment of medically refractory human TLE. PMID- 11283237 TI - Sustained sensitization and recruitment of rat cutaneous nociceptors by bradykinin and a novel theory of its excitatory action. AB - Excitation and sensitization to heat of nociceptors by bradykinin (BK) were examined using an isolated rat skin-saphenous nerve preparation. A total of 52 C fibres was tested: 42 were mechano-heat sensitive (CMH) and 40% of them were excited and sensitized to heat by BK superfusion (10-5 M, 5 min) of their receptive fields; heat responses were augmented by more than five times and heat thresholds dropped to 36.4 degrees C, on average. Sixty per cent of the CMH did not respond to BK itself, but 3/4 of these units showed an increase in their heat responses by more than 100% following BK exposure. Ten high-threshold mechanosensitive C-fibres did not discharge upon BK application but following this five of them responded to heat in a well-graded manner. In all fibres, the sensitizing effect of BK was abolished within 9 min or less of wash-out, and it could be reproduced several times at equal magnitude, whereas the excitatory effect of BK regularly showed profound tachyphylaxis. Sustained superfusion (20 min) of BK induced a desensitizing excitatory response while superimposed heat responses showed constant degrees of sensitization. The large extent and high prevalence of BK-induced sensitization (almost 80% of all fibres tested) and de novo recruitment of heat sensitivity suggest a prominent role of BK not only in hyperalgesia but also in sustained inflammatory pain which may be driven by body or even lower local temperatures acting on sensitized nociceptors. Based on the latter assumption, a hypothesis is put forward that excludes a direct excitatory effect of BK on nociceptors, but assumes a temperature-controlled activation as a result of rapid and profound sensitization. PMID- 11283238 TI - Functional reorganization of sensory pathways in the rat spinal dorsal horn following peripheral nerve injury. AB - Functional reorganization of sensory pathways in the rat spinal dorsal horn following sciatic nerve transection was examined using spinal cord slices with an attached dorsal root. Slices were obtained from animals whose sciatic nerve had been transected 2-4 weeks previously and compared to sham-operated controls. Whole-cell recordings from substantia gelatinosa neurones in sham-operated rats, to which nociceptive information was preferentially transmitted, revealed that dorsal root stimulation sufficient to activate A afferent fibres evoked a mono- and/or polysynaptic EPSC in 111 of 131 (approximately 85%) neurones. This is in contrast to the response following A fibre stimulation, where monosynaptic EPSCs were observed in 2 of 131 (approximately 2%) neurones and polysynaptic EPSCs were observed in 18 of 131 (approximately 14%) neurones. In sciatic nerve-transected rats, however, a polysynaptic EPSC following stimulation of A afferents was elicited in 30 of 37 (81%) neurones and a monosynaptic EPSC evoked by A afferent stimulation was detected in a subset of neurones (4 of 37, approximately 11%). These observations suggest that, following sciatic nerve transection, large myelinated A afferent fibres establish synaptic contact with interneurones and transmit innocuous information to substantia gelatinosa. This functional reorganization of the sensory circuitry may constitute an underlying mechanism, at least in part, for sensory abnormalities following peripheral nerve injuries. PMID- 11283236 TI - Presynaptic function is altered in snake K+-depolarized motor nerve terminals containing compromised mitochondria. AB - Presynaptic function was investigated at K+-stimulated motor nerve terminals in snake costocutaneous nerve muscle preparations exposed to carbonyl cyanide m chlorophenylhydrazone (CCCP, 2 M), oligomycin (8 g x ml(-1)) or CCCP and oligomycin together. Miniature endplate currents (MEPCs) were recorded at -150 mV with two-electrode voltage clamp. With all three drug treatments, during stimulation by elevated K+ (35 mM), MEPC frequencies initially increased to values > 350 s(-1), but then declined. The decline occurred more rapidly in preparations treated with CCCP or CCCP and oligomycin together than in those treated with oligomycin alone. Staining with FM1-43 indicated that synaptic vesicle membrane endocytosis occurred at some CCCP- or oligomycin-treated nerve terminals after 120 or 180 min of K+ stimulation, respectively. The addition of glucose to stimulate production of ATP by glycolysis during sustained K+ stimulation attenuated the decline in MEPC frequency and increased the percentage of terminals stained by FM1-43 in preparations exposed to either CCCP or oligomycin. We propose that the decline in K+-stimulated quantal release in preparations treated with CCCP, oligomycin or CCCP and oligomycin together could result from a progressive elevation of intracellular calcium concentration ([Ca2+]i). For oligomycin-treated nerve terminals, a progressive elevation of [Ca2+]i could occur as the cytoplasmic ATP/ADP ratio decreases, causing energy dependent Ca2+ buffering mechanisms to fail. The decline in MEPC frequency could occur more rapidly in preparations treated with CCCP or CCCP and oligomycin together because mitochondrial Ca2+ buffering and ATP production were both inhibited. Therefore, the proposed sustained elevation of [Ca2+]i could occur more rapidly. PMID- 11283239 TI - Vasodilatation, oxygen delivery and oxygen consumption in rat hindlimb during systemic hypoxia: roles of nitric oxide. AB - We have investigated the relationship between O2 delivery (DO2) and O2 consumption (VO2) in hindlimb muscle of anaesthetised rats during progressive systemic hypoxia. Since muscle vasodilatation that occurs during hypoxia is nitric oxide (NO) dependent, we examined the effects of the NO synthase (NOS) inhibitor nitro-L-arginine methyl ester (L-NAME). In control rats (n = 8), femoral vascular conductance (FVC) increased at each level of hypoxia. Hindlimb DO2 decreased with the severity of hypoxia, but muscle VO2 was maintained until the critical DO2 value (DO2,crit) was reached at 0.64 +/- 0.06 ml O2 min-1 kg-1; below this VO2 declined linearly with DO2. This is a novel finding for the rat but is comparable to the biphasic relationship seen in the dog. In another group of rats (n = 6), L-NAME caused hindlimb vasoconstriction and attenuated the hypoxia-evoked increases in FVC DO2 was so low after L-NAME administration that VO2 was dependent on DO2 at all levels of hypoxia. In a further group (n = 8), femoral blood flow and DO2 were restored after L-NAME by infusion of the NO donor sodium nitroprusside (20 g x kg(-1) x min(-1). Thereafter, hypoxia-evoked increases in FVC were fully restored. Nevertheless, DO2,crit was increased relative to control (0.96 +/- 0.07 ml O2 min(-1) x kg(-1), P < 0.01). As NOS inhibition limited the ability of muscle to maintain VO2 during hypoxia, we propose that hypoxia-induced dilatation of terminal arterioles, which improves tissue O2 distribution, is mediated by NO. However, since the hypoxia-evoked increase in FVC was blocked by L-NAME but restored by the NO donor, we propose that the dilatation of proximal arterioles is dependent on tonic levels of NO, rather than mediated by NO. PMID- 11283240 TI - Thermogenesis induced by osmotic stimulation of the intestines in the rat. AB - Infusion of 5-20% glucose, 1.8-3.6% NaCl, 20% methylglucose, 20% fructose, or 5 10% solutions of various amino acids (10 ml x kg(-1)) into the duodenum induced dose-dependent thermogenesis in urethane-anaesthetized rats. In contrast, infusion of 0.9% NaCl, distilled water, or safflower oil had no effect on the metabolic rate. Infusion of 7.2% urea induced a small and transient increase in the metabolic rate. These results suggested that the thermogenesis was caused mainly by changes in osmolality rather than by a specific action of the different solute molecules. The respiratory exchange ratio increased after the infusion of glucose, fructose, glycine, or serine, did not change after the infusion of NaCl, methylglucose, safflower oil, or distilled water, and decreased after infusion of arginine. Therefore, there was no relationship between substrate utilization and the occurrence of thermogenesis. Intestinal infusion of 3.6% NaCl elevated the plasma osmolality, with a plateau increase of approximately 20 mosmol x kg(-1). However, intravenous infusion of the same amount of NaCl induced a significantly smaller thermogenic response, although it elevated the plasma osmolality with a time course and magnitude similar to those obtained after the intestinal infusion. Infusion of NaCl into the hepatic portal vein or the peritoneal cavity also produced a significantly small thermogenic response. These results suggested an intestinal or mesenteric location for osmoreceptors. To test for possible stimulation of intestinal osmoreceptors after intake of a normal meal, we measured the osmolality of the intestinal contents. The osmolality of the duodeno jejunal contents was 600-800 mosmol kg-1, whereas the plasma osmolality was 306 +/- 1 mosmol x kg(-1), which suggests that the intestinal osmoreceptors are stimulated after meals and are involved in diet-induced thermogenesis. PMID- 11283241 TI - State-dependent hyperpolarization of voltage threshold enhances motoneurone excitability during fictive locomotion in the cat. AB - Experiments were conducted on decerebrate adult cats to examine the effect of brainstem-evoked fictive locomotion on the threshold voltage (Vth) at which action potentials were initiated in hindlimb motoneurones. Measurements of the voltage threshold of the first spike evoked by intracellular injection of depolarizing ramp currents or square pulses were compared during control and fictive locomotor conditions. The sample of motoneurones included flexor and extensor motoneurones, and motoneurones with low and high rheobase currents. In all 38 motoneurones examined, action potentials were initiated at more hyperpolarized membrane potentials during fictive locomotion than in control conditions (mean hyperpolarization -8.0 +/- 5.5 mV; range -1.8 to -26.6 mV). Hyperpolarization of Vth occurred immediately at the onset of fictive locomotion and recovered in seconds (typically < 60 s) following the termination of locomotor activity. The Vth of spikes occurring spontaneously without intracellular current injection was also reduced during locomotion. Superimposition of rhythmic depolarizing current pulses on current ramps in the absence of locomotion did not lower Vth to the extent seen during fictive locomotion. We suggest that Vth hyperpolarization results from an as yet undetermined neuromodulatory process operating during locomotion and is not simply the result of the oscillations in membrane potential occurring during locomotion.The hyperpolarization of Vth for action potential initiation during locomotion is a state-dependent increase in motoneurone excitability. This Vth hyperpolarization may be a fundamental process in the generation of motoneurone activity during locomotion and perhaps other motor tasks. PMID- 11283242 TI - Circadian clock-specific roles for the light response protein WHITE COLLAR-2. AB - To understand the role of white collar-2 in the Neurospora circadian clock, we examined alleles of wc-2 thought to encode partially functional proteins. We found that wc-2 allele ER24 contained a conservative mutation in the zinc finger. This mutation results in reduced levels of circadian rhythm-critical clock gene products, frq mRNA and FRQ protein, and in a lengthened period of the circadian clock. In addition, this mutation altered a second canonical property of the clock, temperature compensation: as temperature increased, period length decreased substantially. This temperature compensation defect correlated with a temperature-dependent increase in overall FRQ protein levels, with the relative increase being greater in wc-2 (ER24) than in wild type, while overall frq mRNA levels were largely unaltered by temperature. We suggest that this temperature dependent increase in FRQ levels partially rescues the lowered levels of FRQ resulting from the wc-2 (ER24) defect, yielding a shorter period at higher temperatures. Thus, normal activity of the essential clock component WC-2, a positive regulator of frq, is critical for establishing period length and temperature compensation in this circadian system. PMID- 11283243 TI - Reconstitution of human beta-globin locus control region hypersensitive sites in the absence of chromatin assembly. AB - The human beta-globin genes are regulated by the locus control region (LCR), an element composed of multiple DNase I-hypersensitive sites (HS sites) located 5' to the genes. Various functional studies indicate that the LCR confers high level, position-independent, and copy number-dependent expression to linked globin genes in transgenic mice. However, the structural basis for LCR function is unknown. Here we show that LCR HS sites can be reconstituted in an erythroid cell-specific manner on chromatin-assembled LCR templates in vitro. Surprisingly, HS2 and HS3 are also formed with erythroid proteins in the absence of chromatin assembly, indicating that sensitivity to nucleases is not simply a consequence of nucleosome reorganization. The generation of LCR HS sites in the absence of chromatin assembly leads to the formation of S1- and KMnO(4)-sensitive regions in HS2 and HS3. These sites are also sensitive to S1 nuclease in erythroid cells in vivo, suggesting a distorted DNA structure in the LCR core enhancer elements. Finally, we show that RNA polymerase II initiates transcription in the HS2 and HS3 core enhancer regions in vitro. Transcription in both HS2 and HS3 proceeds in a unidirectional manner. Taken together, the data suggest that erythroid proteins interact with the core enhancer elements, distort the DNA structure, and recruit polymerase II transcription complexes. These results further our understanding of the structural basis for LCR function and provide an explanation for why the LCR core regions are so extremely sensitive to nucleases in erythroid cells. PMID- 11283244 TI - A step subsequent to preinitiation complex assembly at the ribosomal RNA gene promoter is rate limiting for human RNA polymerase I-dependent transcription. AB - The assembly, disassembly, and functional properties of transcription preinitiation complexes (PICs) of human RNA polymerase I (Pol I) play a crucial role in the regulation of rRNA gene expression. To study the factors and processes involved, an immobilized-promoter template assay has been developed that allows the isolation from nuclear extracts of functional PICs, which support accurate initiation of transcription. Immunoblotting of template-bound factors showed that these complexes contained the factors required to support initiation of transcription, SL1, upstream binding factor (UBF), and Pol I. We have demonstrated that, throughout a single round of transcription, SL1 and UBF remain promoter bound. Moreover, the promoter-bound SL1 and UBF retain the ability to function in transcription initiation. SL1 has a central role in the stable association of the PIC with the promoter DNA. The polymerase component of the PIC is released from the promoter during transcription yet is efficiently recycled and able to reinitiate from "poised" promoters carrying SL1 and UBF, since the PICs captured on the immobilized templates sustained multiple rounds of transcription. Kinetic analyses of initiation of transcription by Pol I revealed that Pol I-dependent transcription is rate limited in a step subsequent to recruitment and assembly of Pol I PICs. The rate of RNA synthesis is primarily determined by the rates at which the polymerase initiates transcription and escapes the promoter, referred to as promoter clearance. This rate-limiting step in Pol I transcription is likely to be a major target in the regulation of rRNA gene expression. PMID- 11283245 TI - Regulation of Ras signaling specificity by protein kinase C. AB - Ras proteins have the capacity to bind to and activate at least three families of downstream target proteins: Raf kinases, phosphatidylinositol 3 (PI 3)-kinase, and Ral-specific guanine nucleotide exchange factors (Ral-GEFs). We have previously shown that the Ras/Ral-GEF and Ras/Raf pathways oppose each other upon nerve growth factor stimulation, with the former promoting proliferation and the latter promoting cell cycle arrest. Moreover, the pathways are not activated equally. While the Ras/Raf/Erk signaling pathway is induced for hours, the Ras/Ral-GEF/Ral signaling pathway is induced for only minutes. Here we show that this preferential down-regulation of Ral signaling is mediated, at least in part, by protein kinase C (PKC). In particular, we show that PKC activation by phorbol ester treatment of cells blocks growth factor-induced Ral activation while it enhances Erk activation. Moreover, suppression of growth factor-induced PKC activation enhances and prolongs Ral activation. PKC does not influence the basal activity of the Ral-GEF designated Ral-GDS but suppresses its activation by Ras. Interestingly, Ras binding to the C-terminal Ras binding domain of Ral-GDS is not affected by PKC activity. Instead, suppression of Ral-GDS activation occurs through the region N terminal to the catalytic domain, which becomes phosphorylated in response to phorbol ester treatment of cells. These findings identify a role for PKC in determining the specificity of Ras signaling by its ability to differentially modulate Ras effector protein activation. PMID- 11283247 TI - Repair of double-strand breaks by homologous recombination in mismatch repair defective mammalian cells. AB - Chromosomal double-strand breaks (DSBs) stimulate homologous recombination by several orders of magnitude in mammalian cells, including murine embryonic stem (ES) cells, but the efficiency of recombination decreases as the heterology between the repair substrates increases (B. Elliott, C. Richardson, J. Winderbaum, J. A. Nickoloff, and M. Jasin, Mol. Cell. Biol. 18:93-101, 1998). We have now examined homologous recombination in mismatch repair (MMR)-defective ES cells to investigate both the frequency of recombination and the outcome of events. Using cells with a targeted mutation in the msh2 gene, we found that the barrier to recombination between diverged substrates is relaxed for both gene targeting and intrachromosomal recombination. Thus, substrates with 1.5% divergence are 10-fold more likely to undergo DSB-promoted recombination in Msh2( /-) cells than in wild-type cells. Although mutant cells can repair DSBs efficiently, examination of gene conversion tracts in recombinants demonstrates that they cannot efficiently correct mismatched heteroduplex DNA (hDNA) that is formed adjacent to the DSB. As a result, >20-fold more of the recombinants derived from mutant cells have uncorrected tracts compared with recombinants from wild-type cells. The results indicate that gene conversion repair of DSBs in mammalian cells frequently involves mismatch correction of hDNA rather than double-strand gap formation. In cells with MMR defects, therefore, aberrant recombinational repair may be an additional mechanism that contributes to genomic instability and possibly tumorigenesis. PMID- 11283246 TI - Thrombopoietin-mediated sustained activation of extracellular signal-regulated kinase in UT7-Mpl cells requires both Ras-Raf-1- and Rap1-B-Raf-dependent pathways. AB - Thrombopoietin (TPO) regulates growth and differentiation of megakaryocytes. We previously showed that extracellular signal-regulated kinases (ERKs) are required for TPO-mediated full megakaryocytic maturation in both normal progenitors and a megakaryoblastic cell line (UT7) expressing the TPO receptor (Mpl). In these cells, intensity and duration of TPO-induced ERK signal are controlled by several regions of the cytoplasmic domain of Mpl. In this study, we explored the signaling pathways involved in this control. We show that the small GTPases Ras and Rap1 contribute together to TPO-induced ERK activation in UT7-Mpl cells and that they do so by activating different Raf kinases as downstream effectors: a Ras-Raf-1 pathway is required to initiate ERK activation while Rap1 sustains this signal through B-Raf. Indeed, (i) in cells expressing wild-type or mutant Mpl, TPO-induced Ras and Rap1 activation correlates with early and sustained phases of ERK signal, respectively; (ii) interfering mutants of Ras and Rap1 both inhibit ERK kinase activity and ERK-dependent Elk1 transcriptional activation in response to TPO; (iii) the kinetics of activation of Raf-1 and B-Raf by TPO follow those of Ras and Rap1, respectively; (iv) RasV12-mediated Elk1 activation was modulated by the wild type or interfering mutants of Raf-1 but not those of B-Raf; (v) Elk1 activation mediated by a constitutively active mutant of Rap1 (Rap1V12) is potentiated by B-Raf and inhibited by an interfering mutant of this kinase. UT7 Mpl cells represent the second cellular model in which Ras and Rap1 act in concert to modulate the duration of ERK signal in response to a growth factor and thereby the differentiation program. This is also, to our knowledge, the first evidence suggesting that Rap1 may play an active role in megakaryocytic maturation. PMID- 11283248 TI - Insulin control of glycogen metabolism in knockout mice lacking the muscle specific protein phosphatase PP1G/RGL. AB - The regulatory-targeting subunit (RGL), also called GM) of the muscle-specific glycogen-associated protein phosphatase PP1G targets the enzyme to glycogen where it modulates the activity of glycogen-metabolizing enzymes. PP1G/RGL has been postulated to play a central role in epinephrine and insulin control of glycogen metabolism via phosphorylation of RGL. To investigate the function of the phosphatase, RGL knockout mice were generated. Animals lacking RGL show no obvious defects. The RGL protein is absent from the skeletal and cardiac muscle of null mutants and present at approximately 50% of the wild-type level in heterozygotes. Both the level and activity of C1 protein are also decreased by approximately 50% in the RGL-deficient mice. In skeletal muscle, the glycogen synthase (GS) activity ratio in the absence and presence of glucose-6-phosphate is reduced from 0.3 in the wild type to 0.1 in the null mutant RGL mice, whereas the phosphorylase activity ratio in the absence and presence of AMP is increased from 0.4 to 0.7. Glycogen accumulation is decreased by approximately 90%. Despite impaired glycogen accumulation in muscle, the animals remain normoglycemic. Glucose tolerance and insulin responsiveness are identical in wild-type and knockout mice, as are basal and insulin-stimulated glucose uptakes in skeletal muscle. Most importantly, insulin activated GS in both wild-type and RGL null mutant mice and stimulated a GS-specific protein phosphatase in both groups. These results demonstrate that RGL is genetically linked to glycogen metabolism, since its loss decreases PP1 and basal GS activities and glycogen accumulation. However, PP1G/RGL is not required for insulin activation of GS in skeletal muscle, and rather another GS-specific phosphatase appears to be involved. PMID- 11283249 TI - Sustained signaling by phospholipase C-gamma mediates nerve growth factor triggered gene expression. AB - In contrast to conventional signaling by growth factors that requires their continual presence, a 1-min pulse of nerve growth factor (NGF) is sufficient to induce electrical excitability in PC12 cells due to induction of the peripheral nerve type 1 (PN1) sodium channel gene. We have investigated the mechanism for this triggered signaling pathway by NGF in PC12 cells. Mutation of TrkA at key autophosphorylation sites indicates an essential role for the phospholipase C gamma (PLC-gamma) binding site, but not the Shc binding site, for NGF-triggered induction of PN1. In concordance with results with Trk mutants, drug-mediated inhibition of PLC-gamma activity also blocks PN1 induction by NGF. Examination of the kinetics of TrkA autophosphorylation indicates that triggered signaling does not result from sustained activation and autophosphorylation of the TrkA receptor kinase, whose phosphorylation state declines rapidly after NGF removal. Rather, TrkA triggers an unexpectedly prolonged phosphorylation and activation of PLC gamma signaling that is sustained for up to 2 h. Prevention of the elevation of intracellular Ca2+ levels using BAPTA-AM results in a block of PN1 induction by NGF. Sustained signaling by PLC-gamma provides a means for differential neuronal gene induction after transient exposure to NGF. PMID- 11283250 TI - UvsW protein regulates bacteriophage T4 origin-dependent replication by unwinding R-loops. AB - The UvsW protein of bacteriophage T4 is involved in many aspects of phage DNA metabolism, including repair, recombination, and recombination-dependent replication. UvsW has also been implicated in the repression of origin-dependent replication at late times of infection, when UvsW is normally synthesized. Two well-characterized T4 origins, ori(uvsY) and ori(34), are believed to initiate replication through an R-loop mechanism. Here we provide both in vivo and in vitro evidence that UvsW is an RNA-DNA helicase that catalyzes the dissociation of RNA from origin R-loops. Two-dimensional gel analyses show that the replicative intermediates formed at ori(uvsY) persist longer in a uvsW mutant infection than in a wild-type infection. In addition, the inappropriate early expression of UvsW protein results in the loss of these replicative intermediates. Using a synthetic origin R-loop, we also demonstrate that purified UvsW functions as a helicase that efficiently dissociates RNA from R-loops. These and previous results from a number of studies provide strong evidence that UvsW is a molecular switch that allows T4 replication to progress from a mode that initiates from R-loops at origins to a mode that initiates from D-loops formed by recombination proteins. PMID- 11283251 TI - Structure and specificity of GATA proteins in Th2 development. AB - Development of Th2 subset of CD4+ T cells involves the interleukin-4 (IL-4)- and Stat6-dependent increase in GATA-3 expression during primary activation. Recently we reported that the phenotypic stability and factor independence of Th2 cells involves acquisition of an intracellular pathway that maintains GATA-3 expression. Evidence from retroviral expression studies implied that this pathway involved an autoactivation of GATA-3 expression, since Stat6-deficient T cells induced endogenous GATA-3 when infected with GATA-3-expressing retroviruses. That study left unresolved the issue of whether GATA-3 autoactivation was direct or indirect. Several other Th2-specific transcription factors have been described, including c-Maf and JunB. We therefore examined the ability of these other transcription factors to induce GATA-3 expression and promote Th2 development. Neither c-Maf nor JunB induced Th2 development in Stat6-deficient CD4+ T cells, in contrast to GATA-3. Consistent with this indication of a possible direct autoactivation pathway, we also observed that heterologous GATA family proteins GATA-1, GATA-2, and GATA-4 were also capable of inducing GATA-3 expression in developing Stat6-deficient T cells and promote Th2 development. Mutational analysis revealed evidence for two distinct mechanisms of GATA-3 action. IL-4 induction by GATA-3 required each of the functional domains to be present, whereas repression of gamma interferon could occur even when mutants of GATA-3 lacking the second transactivation domain, TA2, were expressed. The GATA dependent induction of the GATA-3 but not the other GATA genes in T cells suggests that T-cell-specific cis elements within the GATA-3 locus likely cooperate with a general GATA recognition motif to allow GATA-3-dependent autoactivation. PMID- 11283252 TI - Histone acetylation at promoters is differentially affected by specific activators and repressors. AB - We analyzed the relationship between histone acetylation and transcriptional regulation at 40 Saccharomyces cerevisiae promoters that respond to specific activators and repressors. In accord with the general correlation between histone acetylation and transcriptional activity, Gcn4 and the general stress activators (Msn2 and Msn4) cause increased acetylation of histones H3 and H4. Surprisingly, Gal4-dependent activation is associated with a dramatic decrease in histone H4 acetylation, whereas acetylation of histone H3 is unaffected. A specific decrease in H4 acetylation is also observed, to a lesser extent, at promoters activated by Hap4, Adr1, Met4, and Ace1. Activation by heat shock factor has multiple effects; H4 acetylation increases at some promoters, whereas other promoters show an apparent decrease in H3 and H4 acetylation that probably reflects nucleosome loss or gross alteration of chromatin structure. Repression by targeted recruitment of the Sin3-Rpd3 histone deacetylase is associated with decreased H3 and H4 acetylation, whereas repression by Cyc8-Tup1 is associated with decreased H3 acetylation but variable effects on H4 acetylation; this suggests that Cyc8-Tup1 uses multiple mechanisms to reduce histone acetylation at promoters. Thus, individual activators confer distinct patterns of histone acetylation on target promoters, and transcriptional activation is not necessarily associated with increased acetylation. We speculate that the activator-specific decrease in histone H4 acetylation is due to blocking the access or function of an H4 specific histone acetylase such as Esa1. PMID- 11283253 TI - Yeast NC2 associates with the RNA polymerase II preinitiation complex and selectively affects transcription in vivo. AB - NC2 (Dr1-Drap1 or Bur6-Ydr1) has been characterized in vitro as a general negative regulator of RNA polymerase II (Pol II) transcription that interacts with TATA-binding protein (TBP) and inhibits its function. Here, we show that NC2 associates with promoters in vivo in a manner that correlates with transcriptional activity and with occupancy by basal transcription factors. NC2 rapidly associates with promoters in response to transcriptional activation, and it remains associated under conditions in which transcription is blocked after assembly of the Pol II preinitiation complex. NC2 positively and negatively affects approximately 17% of Saccharomyces cerevisiae genes in a pattern that resembles the response to general environmental stress. Relative to TBP, NC2 occupancy is high at promoters where NC2 is positively required for normal levels of transcription. Thus, NC2 is associated with the Pol II preinitiation complex, and it can play a direct and positive role at certain promoters in vivo. PMID- 11283254 TI - Jun NH2-terminal kinase phosphorylation of p53 on Thr-81 is important for p53 stabilization and transcriptional activities in response to stress. AB - The p53 tumor suppressor protein plays a key role in the regulation of stress mediated growth arrest and apoptosis. Stress-induced phosphorylation of p53 tightly regulates its stability and transcriptional activities. Mass spectrometry analysis of p53 phosphorylated in 293T cells by active Jun NH2-terminal kinase (JNK) identified T81 as the JNK phosphorylation site. JNK phosphorylated p53 at T81 in response to DNA damage and stress-inducing agents, as determined by phospho-specific antibodies to T81. Unlike wild-type p53, in response to JNK stimuli p53 mutated on T81 (T81A) did not exhibit increased expression or concomitant activation of transcriptional activity, growth inhibition, and apoptosis. Forced expression of MKP5, a JNK phosphatase, in JNK kinase-expressing cells decreased T81 phosphorylation while reducing p53 transcriptional activity and p53-mediated apoptosis. Similarly transfection of antisense JNK 1 and -2 decreased T81 phosphorylation in response to UV irradiation. More than 180 human tumors have been reported to contain p53 with mutations within the region that encompasses T81 and the JNK binding site (amino acids 81 to 116). Our studies identify an additional mechanism for the regulation of p53 stability and functional activities in response to stress. PMID- 11283256 TI - Intermolecular and intramolecular interactions regulate catalytic activity of myotonic dystrophy kinase-related Cdc42-binding kinase alpha. AB - Myotonic dystrophy kinase-related Cdc42-binding kinase (MRCK) is a Cdc42-binding serine/threonine kinase with multiple functional domains. We had previously shown MRCKalpha to be implicated in Cdc42-mediated peripheral actin formation and neurite outgrowth in HeLa and PC12 cells, respectively. Here we demonstrate that native MRCK exists in high-molecular-weight complexes. We further show that the three independent coiled-coil (CC) domains and the N-terminal region preceding the kinase domain are responsible for intermolecular interactions leading to MRCKalpha multimerization. N terminus-mediated dimerization and consequent transautophosphorylation are critical processes regulating MRCKalpha catalytic activities. A region containing the two distal CC domains (CC2 and CC3; residues 658 to 930) was found to interact intramolecularly with the kinase domain and negatively regulates its activity. Its deletion also resulted in an active kinase, confirming a negative autoregulatory role. We provide evidence that the N terminus-mediated dimerization and activation of MRCK and the negative autoregulatory kinase-distal CC interaction are two mutually exclusive events that tightly regulate the catalytic state of the kinase. Disruption of this interaction by a mutant kinase domain resulted in increased kinase activity. MRCK kinase activity was also elevated when cells were treated with phorbol ester, which can interact directly with a cysteine-rich domain next to the distal CC domain. We therefore suggest that binding of phorbol ester to MRCK releases its autoinhibition, allowing N-terminal dimerization and subsequent kinase activation. PMID- 11283255 TI - S and G2 phase roles for Cdk2 revealed by inducible expression of a dominant negative mutant in human cells. AB - Cyclin-dependent kinase 2 (Cdk2) is essential for initiation of DNA synthesis in higher eukaryotes. Biochemical studies in Xenopus egg extracts and microinjection studies in human cells have suggested an additional function for Cdk2 in activation of Cdk1 and entry into mitosis. To further examine the role of Cdk2 in human cells, we generated stable clones with inducible expression of wild-type and dominant-negative forms of the enzyme (Cdk2-wt and Cdk2-dn, respectively). Both exogenous proteins associated efficiently with endogenous cyclins. Cdk2-wt had no apparent effect on the cell division cycle, whereas Cdk2-dn inhibited progression through several distinct stages. Cdk2-dn induction could arrest cells at the G1/S transition, as previously observed in transient expression studies. However, under normal culture conditions, Cdk2-dn induction primarily arrested cells with S and G2/M DNA contents. Several observations suggested that the latter cells were in G2 phase, prior to the onset of mitosis: these cells contained uncondensed chromosomes, low levels of cyclin B-associated kinase activity, and high levels of tyrosine-phosphorylated Cdk1. Furthermore, Cdk2-dn did not delay progression through mitosis upon release of cells from a nocodazole block. Although the G2 arrest imposed by Cdk2-dn was similar to that imposed by the DNA damage checkpoint, the former was distinguished by its resistance to caffeine. These findings provide evidence for essential functions of Cdk2 during S and G2 phases of the mammalian cell cycle. PMID- 11283257 TI - Dimerization of sterol regulatory element-binding protein 2 via the helix-loop helix-leucine zipper domain is a prerequisite for its nuclear localization mediated by importin beta. AB - The sterol regulatory element-binding protein 2 (SREBP-2), a transcription factor of the basic helix-loop-helix-leucine zipper (bHLH-Zip) family, is synthesized in the form of a membrane-attached precursor molecule. When cells are deprived of sterols, a two-step proteolytic processing releases the transcriptionally active N-terminal segment of SREBP-2, thereby allowing it to enter the nucleus. In previous studies, we showed that the nuclear import of SREBP-2 occurs via the direct interaction of importin beta with the HLH-Zip domain. In this study, in order to more completely understand the intracellular dynamics of SREBP-2, we focused on the manner by which importin beta recognizes SREBP-2 at the initial step of the import. It was found that the active form of SREBP-2 exists as a stable dimer in solution and that the substitution of leucine residues for alanine in the leucine zipper motif disrupted the dimerization. It was also demonstrated that this mutant protein did not enter the nucleus either in vivo or in vitro. Solution binding assays, which involved the chemical cross-linking of wild-type or mutated SREBP-2 with importin beta, revealed that the import-active complex appeared to be composed of a dimeric form of SREBP-2 and importin beta. In addition, the SREBP-2 binding domain of importin beta corresponded to an overlapping but not identical region for importin alpha binding, which may explain how importin beta is able to recognize the dimeric HLH-Zip directly. These results indicate that dimerization is a prerequisite process for the nuclear import of SREBP-2 mediated by importin beta. PMID- 11283258 TI - Two compound replication origins in Saccharomyces cerevisiae contain redundant origin recognition complex binding sites. AB - While many of the proteins involved in the initiation of DNA replication are conserved between yeasts and metazoans, the structure of the replication origins themselves has appeared to be different. As typified by ARS1, replication origins in Saccharomyces cerevisiae are <150 bp long and have a simple modular structure, consisting of a single binding site for the origin recognition complex, the replication initiator protein, and one or more accessory sequences. DNA replication initiates from a discrete site. While the important sequences are currently less well defined, metazoan origins appear to be different. These origins are large and appear to be composed of multiple, redundant elements, and replication initiates throughout zones as large as 55 kb. In this report, we characterize two S. cerevisiae replication origins, ARS101 and ARS310, which differ from the paradigm. These origins contain multiple, redundant binding sites for the origin recognition complex. Each binding site must be altered to abolish origin function, while the alteration of a single binding site is sufficient to inactivate ARS1. This redundant structure may be similar to that seen in metazoan origins. PMID- 11283259 TI - Temporal recruitment of the mSin3A-histone deacetylase corepressor complex to the ETS domain transcription factor Elk-1. AB - The transcriptional status of eukaryotic genes is determined by a balance between activation and repression mechanisms. The nuclear hormone receptors represent classical examples of transcription factors that can regulate this balance by recruiting corepressor and coactivator complexes in a ligand-dependent manner. Here, we demonstrate that the equilibrium between activation and repression via a single transcription factor, Elk-1, is altered following activation of the Erk mitogen-activated protein kinase cascade. In addition to its C-terminal transcriptional activation domain, Elk-1 contains an N-terminal transcriptional repression domain that can recruit the mSin3A-histone deacetylase 1 corepressor complex. Recruitment of this corepressor is enhanced in response to activation of the Erk pathway in vivo, and this recruitment correlates kinetically with the shutoff of one of its target promoters, c-fos. Elk-1 therefore undergoes temporal activator-repressor switching and contributes to both the activation and repression of target genes following growth factor stimulation. PMID- 11283260 TI - Reduction of target gene expression by a modified U1 snRNA. AB - Although the primary function of U1 snRNA is to define the 5' donor site of an intron, it can also block the accumulation of a specific RNA transcript when it binds to a donor sequence within its terminal exon. This work was initiated to investigate if this property of U1 snRNA could be exploited as an effective method for inactivating any target gene. The initial 10-bp segment of U1 snRNA, which is complementary to the 5' donor sequence, was modified to recognize various target mRNAs (chloramphenicol acetyltransferase [CAT], beta galactosidase, or green fluorescent protein [GFP]). Transient cotransfection of reporter genes and appropriate U1 antitarget vectors resulted in >90% reduction of transgene expression. Numerous sites within the CAT transcript were suitable for targeting. The inhibitory effect of the U1 antitarget vector is directly related to the hybrid formed between the U1 vector and target transcripts and is dependent on an intact 70,000-molecular-weight binding domain within the U1 gene. The effect is long lasting when the target (CAT or GFP) and U1 antitarget construct are inserted into fibroblasts by stable transfection. Clonal cell lines derived from stable transfection with a pOB4GFP target construct and subsequently stably transfected with the U1 anti-GFP construct were selected. The degree to which GFP fluorescence was inhibited by U1 anti-GFP in the various clonal cell lines was assessed by fluorescence-activated cell sorter analysis. RNA analysis demonstrated reduction of the GFP mRNA in the nuclear and cytoplasmic compartment and proper 3' cleavage of the GFP residual transcript. An RNase protection strategy demonstrated that the transfected U1 antitarget RNA level varied between 1 to 8% of the endogenous U1 snRNA level. U1 antitarget vectors were demonstrated to have potential as effective inhibitors of gene expression in intact cells. PMID- 11283261 TI - Novel fluorescence-based screen to identify small synthetic internal ribosome entry site elements. AB - We report here a novel fluorescent protein-based screen to identify small, synthetic internal ribosome entry site (IRES) elements in vivo. A library of bicistronic plasmids encoding the enhanced blue and green fluorescent proteins (EBFP and EGFP) separated by randomized 50-nucleotide-long sequences was amplified in bacteria and delivered into mammalian cells via protoplast fusion. Cells that received functional IRES elements were isolated using the EBFP and EGFP reporters and fluorescence-activated cell sorting, and several small IRES elements were identified. Two of these elements were subsequently shown to possess IRES activity comparable to that of a variant of the encephalomyocarditis virus IRES element in a context-independent manner both in vitro and in vivo, and these elements functioned in multiple cell types. Although no sequence or structural homology was apparent between the synthetic IRES elements and known viral and cellular IRES elements, the two synthetic IRES elements specifically blocked poliovirus (PV) IRES-mediated translation in vitro. Competitive protein binding experiments suggested that these IRES elements compete with PV IRES mediated translation by utilizing some of the same factors as the PV IRES to direct translation. The utility of this fluorescent protein-based screen in identifying IRES elements with improved activity as well as in probing the mechanism of IRES-mediated translation is discussed. PMID- 11283262 TI - The peptide near the C terminus regulates receptor CAR nuclear translocation induced by xenochemicals in mouse liver. AB - In response to phenobarbital (PB) and other PB-type inducers, the nuclear receptor CAR translocates to the mouse liver nucleus (T. Kawamoto et al., Mol. Cell. Biol. 19:6318-6322, 1999). To define the translocation mechanism, fluorescent protein-tagged human CAR (hCAR) was expressed in the mouse livers using the in situ DNA injection and gene delivery systems. As in the wild-type hCAR, the truncated receptor lacking the C-terminal 10 residues (i.e., AF2 domain) translocated to the nucleus, indicating that the PB-inducible translocation is AF2 independent. Deletion of the 30 C-terminal residues abolished the receptor translocation, and subsequent site-directed mutagenesis delineated the PB-inducible translocation activity of the receptor to the peptide L313GLL316AEL319. Ala mutations of Leu313, Leu316, or Leu319 abrogated the translocation of CAR in the livers, while those of Leu312 or Leu315 did not affect the nuclear translocation. The leucine-rich peptide dictates the nuclear translocation of hCAR in response to various PB-type inducers and appears to be conserved in the mouse and rat receptors. PMID- 11283264 TI - Chromosome instability and defective recombinational repair in knockout mutants of the five Rad51 paralogs. AB - The Rad51 protein, a eukaryotic homologue of Escherichia coli RecA, plays a central role in both mitotic and meiotic homologous DNA recombination (HR) in Saccharomyces cerevisiae and is essential for the proliferation of vertebrate cells. Five vertebrate genes, RAD51B, -C, and -D and XRCC2 and -3, are implicated in HR on the basis of their sequence similarity to Rad51 (Rad51 paralogs). We generated mutants deficient in each of these proteins in the chicken B-lymphocyte DT40 cell line and report here the comparison of four new mutants and their complemented derivatives with our previously reported rad51b mutant. The Rad51 paralog mutations all impair HR, as measured by targeted integration and sister chromatid exchange. Remarkably, the mutant cell lines all exhibit very similar phenotypes: spontaneous chromosomal aberrations, high sensitivity to killing by cross-linking agents (mitomycin C and cisplatin), mild sensitivity to gamma rays, and significantly attenuated Rad51 focus formation during recombinational repair after exposure to gamma rays. Moreover, all mutants show partial correction of resistance to DNA damage by overexpression of human Rad51. We conclude that the Rad51 paralogs participate in repair as a functional unit that facilitates the action of Rad51 in HR. PMID- 11283263 TI - The insert region of Rac1 is essential for membrane ruffling but not cellular transformation. AB - The Rho family of Ras-related proteins, which includes Rac1, RhoA, and Cdc42, is distinguished from other members of the Ras superfamily of small GTPases in that its members possess additional sequences positioned between beta-strand 5 and alpha-helix 4, designated the insert region. Previous studies have established the importance of an intact insert region for the transforming, but not actin cytoskeletal reorganization, activities of Cdc42 and RhoA. Similarly, the insert region was determined to be essential for Rac1-mediated mitogenesis. Additionally, an intact insert region was also determined to be required for the antiapoptotic activity of Rac1 as well as for Rac1 activation of reactive oxygen species and the NF-kappaB transcription factor. However, it has not been determined whether the insert region is important for Rac1-mediated growth transformation. In this study, we assessed the requirement for the insert region in Rac1 transformation and signaling in NIH 3T3 cells. Unexpectedly, we found that a mutant of constitutively activated Rac1 that lacked the insert region retained potent transforming activity. The insert region of Rac1 was dispensable for Rac1 stimulation of transcription from the cyclin D1 promoter and for activation of the c-Jun, NF-kappaB, and E2F-1 transcription factors but was essential for Rac1 induction of serum response factor activity. While an intact insert region was dispensable for inducing reactive oxygen species production in vivo, it was required for Rac1 induction of lamellipodia. When taken together, these results show that the insert region of Rac1 serves roles in regulating actin organization and cell growth that are distinct from those of the analogous regions of Cdc42 and RhoA and support its involvement in regulating specific downstream effector interactions. PMID- 11283266 TI - Requirement for the murine zinc finger protein ZFR in perigastrulation growth and survival. AB - The transition from preimplantation to postimplantation development leads to the initiation of complex cellular differentiation and morphogenetic movements, a dramatic decrease in cell cycle length, and a commensurate increase in the size of the embryo. Accompanying these changes is the need for the transfer of nutrients from the mother to the embryo and the elaboration of sophisticated genetic networks that monitor genomic integrity and the homeostatic control of cellular growth, differentiation, and programmed cell death. To determine the function of the murine zinc finger protein ZFR in these events, we generated mice carrying a null mutation in the gene encoding it. Homozygous mutant embryos form normal-appearing blastocysts that implant and initiate the process of gastrulation. Mutant embryos form mesoderm but they are delayed in their development and fail to form normal anterior embryonic structures. Loss of ZFR function leads to both an increase in programmed cell death and a decrease in mitotic index, especially in the region of the distal tip of the embryonic ectoderm. Mutant embryos also have an apparent reduction in apical vacuoles in the columnar visceral endoderm cells in the extraembryonic region. Together, these cellular phenotypes lead to a dramatic development delay and embryonic death by 8 to 9 days of gestation, which are independent of p53 function. PMID- 11283265 TI - Long-range nucleosome ordering is associated with gene silencing in Drosophila melanogaster pericentric heterochromatin. AB - We have used line HS-2 of Drosophila melanogaster, carrying a silenced transgene in the pericentric heterochromatin, to investigate in detail the chromatin structure imposed by this environment. Digestion of the chromatin with micrococcal nuclease (MNase) shows a nucleosome array with extensive long-range order, indicating regular spacing, and with well-defined MNase cleavage fragments, indicating a smaller MNase target in the linker region. The repeating unit is ca. 10 bp larger than that observed for bulk Drosophila chromatin. The silenced transgene shows both a loss of DNase I-hypersensitive sites and decreased sensitivity to DNase I digestion within an array of nucleosomes lacking such sites; within such an array, sensitivity to digestion by MNase is unchanged. The ordered nucleosome array extends across the regulatory region of the transgene, a shift that could explain the loss of transgene expression in heterochromatin. Highly regular nucleosome arrays are observed over several endogenous heterochromatic sequences, indicating that this is a general feature of heterochromatin. However, genes normally active within heterochromatin (rolled and light) do not show this pattern, suggesting that the altered chromatin structure observed is associated with regions that are silent, rather than being a property of the domain as a whole. The results indicate that long-range nucleosomal ordering is linked with the heterochromatic packaging that imposes gene silencing. PMID- 11283267 TI - runt homology domain transcription factors (Runx, Cbfa, and AML) mediate repression of the bone sialoprotein promoter: evidence for promoter context dependent activity of Cbfa proteins. AB - Expression of the bone sialoprotein (BSP) gene, a marker of bone formation, is largely restricted to cells in mineralized tissues. Recent studies have shown that the Cbfa1 (also known as Runx2, AML-3, and PEBP2alphaA) transcription factor supports commitment and differentiation of progenitor cells to hypertrophic chondrocytes and osteoblasts. This study addresses the functional involvement of Cbfa sites in expression of the Gallus BSP gene. Gel mobility shift analyses with nuclear extracts from ROS 17/2.8 osteoblastic cells revealed that multiple Cbfa consensus sequences are functional Cbfa DNA binding sites. Responsiveness of the 1.2-kb Gallus BSP promoter to Cbfa factors Cbfa1, Cbfa2, and Cbfa3 was assayed in osseous and nonosseous cells. Each of the Cbfa factors mediated repression of the wild-type BSP promoter, in contrast to their well known activation of various hematopoietic and skeletal phenotypic genes. Suppression of BSP by Cbfa factors was not observed in BSP promoters in which Cbfa sites were deleted or mutated. Expression of the endogenous BSP gene in Gallus osteoblasts was similarly downregulated by forced expression of Cbfa factors. Our data indicate that Cbfa repression of the BSP promoter does not involve the transducin-like enhancer (TLE) proteins. Neither coexpression of TLE1 or TLE2 nor the absence of the TLE interaction motif of Cbfa1 (amino acids 501 to 513) influenced repressor activity. However, removal of the C terminus of Cbfa1 (amino acids 362 to 513) relieved suppression of the BSP promoter. Our results, together with the evolutionary conservation of the seven Cbfa sites in the Gallus and human BSP promoters, suggest that suppressor activity by Cbfa is of significant physiologic consequence and may contribute to spatiotemporal expression of BSP during bone development. PMID- 11283268 TI - Discoidin domain receptor 1 tyrosine kinase has an essential role in mammary gland development. AB - Various types of collagen have been identified as potential ligands for the two mammalian discoidin domain receptor tyrosine kinases, DDR1 and DDR2. Here, we used a recombinant fusion protein between the extracellular domain of DDR1 and alkaline phosphatase to detect specific receptor binding sites during mouse development. Major sites of DDR1-binding activity, indicative of ligand expression, were found in skeletal bones, the skin, and the urogenital tract. Ligand expression in the uterus during implantation and in the mammary gland during pregnancy colocalized with the expression of the DDR1 receptor. The generation of DDR1-null mice by gene targeting yielded homozygous mutant animals that were viable but smaller in size than control littermates. The majority of mutant females were unable to bear offspring due to a lack of proper blastocyst implantation into the uterine wall. When implantation did occur, the mutant females were unable to lactate. Histological analysis showed that the alveolar epithelium failed to secrete milk proteins into the lumen of the mammary gland. The lactational defect appears to be caused by hyperproliferation and abnormal branching of mammary ducts. These results suggest that DDR1 is a key mediator of the stromal-epithelial interaction during ductal morphogenesis in the mammary gland. PMID- 11283269 TI - RBP1 recruits the mSIN3-histone deacetylase complex to the pocket of retinoblastoma tumor suppressor family proteins found in limited discrete regions of the nucleus at growth arrest. AB - Retinoblastoma (RB) tumor suppressor family pocket proteins induce cell cycle arrest by repressing transcription of E2F-regulated genes through both histone deacetylase (HDAC)-dependent and -independent mechanisms. In this study we have identified a stable complex that accounts for the recruitment of both repression activities to the pocket. One component of this complex is RBP1, a known pocket binding protein that exhibits both HDAC-dependent and -independent repression functions. RB family proteins were shown to associate via the pocket with previously identified mSIN3-SAP30-HDAC complexes containing exclusively class I HDACs. Such enzymes do not interact directly with RB family proteins but rather utilize RBP1 to target the pocket. This mechanism was shown to account for the majority of RB-associated HDAC activity. We also show that in quiescent normal human cells this entire RBP1-mSIN3-SAP30-HDAC complex colocalizes with both RB family members and E2F4 in a limited number of discrete regions of the nucleus that in other studies have been shown to represent the initial origins of DNA replication following growth stimulation. These results suggest that RB family members, at least in part, drive exit from the cell cycle by recruitment of this HDAC complex via RBP1 to repress transcription from E2F-dependent promoters and possibly to alter chromatin structure at DNA origins. PMID- 11283271 TI - The human kinesin-like protein RB6K is under tight cell cycle control and is essential for cytokinesis. AB - Several members of the kinesin superfamily are known to play a prominent role in the motor-driven transport processes that occur in mitotic cells. Here we describe a new mitotic human kinesin-like protein, RB6K (Rabkinesin 6), distantly related to MKLP-1. Expression of RB6K is regulated during the cell cycle at both the mRNA and protein level and, similar to cyclin B, shows a maximum during M phase. Isolation of the RB6K promoter allowed identification of a CDE-CHR element and promoter activity was shown to be maximal during M phase. Immunofluorescence microscopy using antibodies raised against RB6K showed a weak signal in interphase Golgi but a 10-fold higher signal in prophase nuclei. During M phase, the newly synthesized RB6K does not colocalise with Rab6. In later stages of mitosis RB6K localized to the spindle midzone and appeared on the midbodies during cytokinesis. The functional significance of this localization during M phase was revealed by antibody microinjection studies which resulted exclusively in binucleate cells, showing a complete failure of cytokinesis. These results substantiate a crucial role for RB6K in late anaphase B and/or cytokinesis, clearly distinct from the role of MKLP-1. PMID- 11283270 TI - Serum response factor is required for immediate-early gene activation yet is dispensable for proliferation of embryonic stem cells. AB - Addition of serum to mitogen-starved cells activates the cellular immediate-early gene (IEG) response. Serum response factor (SRF) contributes to such mitogen stimulated transcriptional induction of many IEGs during the G0-G1 cell cycle transition. SRF is also believed to be essential for cell cycle progression, as impairment of SRF activity by specific antisera or antisense RNA has previously been shown to block mammalian cell proliferation. In contrast, Srf(-/-) mouse embryos grow and develop up to E6.0. Using the embryonic stem (ES) cell system, we demonstrate here that wild-type ES cells do not undergo complete cell cycle arrest upon serum withdrawal but that they can mount an efficient IEG response. This IEG response, however, is severely impaired in Srf(-/-) ES cells, providing the first genetic proof that IEG activation is dependent upon SRF. Also, Srf(-/-) ES cells display altered cellular morphology, reduced cortical actin expression, and an impaired plating efficiency on gelatin. Yet, despite these defects, the proliferation rates of Srf(-/-) ES cells are not substantially altered, demonstrating that SRF function is not required for ES cell cycle progression. PMID- 11283272 TI - Cyclin A is a mediator of p120E4F-dependent cell cycle arrest in G1. AB - E4F is a ubiquitously expressed GLI-Kruppel-related transcription factor which has been identified for its capacity to regulate transcription of the adenovirus E4 gene in response to E1A. However, cellular genes regulated by E4F are still unknown. Some of these genes are likely to be involved in cell cycle progression since ectopic p120E4F expression induces cell cycle arrest in G1. Although p21WAF1 stabilization was proposed to mediate E4F-dependent cell cycle arrest, we found that p120E4F can induce a G1 block in p21(-/-) cells, suggesting that other proteins are essential for the p120E4F-dependent block in G1. We show here that cyclin A promoter activity can be repressed by p120E4F and that this repression correlates with p120E4F binding to the cyclic AMP-responsive element site of the cyclin A promoter. In addition, enforced expression of cyclin A releases p120E4F arrested cells from the G1 block. These data identify the cyclin A gene as a cellular target for p120E4F and suggest a mechanism for p120E4F-dependent cell cycle regulation. PMID- 11283274 TI - Differential regulation of cell wall biogenesis during growth and development in yeast. PMID- 11283273 TI - The roles of plasmids in phytopathogenic bacteria: mobile arsenals? PMID- 11283275 TI - Feedback control of polyketide metabolism during tylosin production. AB - Tylosin is produced by Streptomyces fradiae via a combination of polyketide metabolism and synthesis of three deoxyhexose sugars, of which mycaminose is the first to be added to the polyketide aglycone, tylactone (protylonolide). Previously, disruption of the gene (tylMII) encoding attachment of mycaminose to the aglycone unexpectedly abolished accumulation of the latter, raising the possibility of a link between polyketide metabolism and deoxyhexose biosynthesis in S. fradiae. However, at that time, it was not possible to eliminate an alternative explanation, namely, that downstream effects on the expression of other genes, not involved in mycaminose metabolism, might have contributed to this phenomenon. Here, it is shown that disruption of any of the four genes (tylMI--III and tylB) specifically involved in mycaminose biosynthesis elicits a similar response, confirming that production of mycaminosyl-tylactone directly influences polyketide metabolism in S. fradiae. Under similar conditions, when mycaminose biosynthesis was specifically blocked by gene disruption, accumulation of tylactone could be restored by exogenous addition of glycosylated tylosin precursors. Moreover, certain other macrolides, not of the tylosin pathway, were also found to elicit qualitatively similar effects. Comparison of the structures of stimulatory macrolides will facilitate studies of the stimulatory mechanism. PMID- 11283276 TI - The signal transducer (BlaRI) and the repressor (BlaI) of the Staphylococcus aureus beta-lactamase operon are inducible. AB - The precise start points for transcription of the blaZ and of the blaRI/blaI genes of the Staphylococcus aureus beta-lactamase operon have been determined by primer extension analysis. Consequently the overlapping promoter sequences were deduced. Northern blots showed that the synthesis of the 2100 nt mRNA from blaRI is inducible and that a blaI probe hybridized to the same mRNA as the blaRI probe. The gene cat, encoding chloramphenicol acetyltransferase, was fused separately to the blaZ and blaRI/blaI promoters, and used to compare their strengths. The promoter for blaZ is about six times stronger than that for blaRI/blaI and the synthesis of chloramphenicol acetyltransferase from both promoters is inducible, supporting the results from the Northern blots. PMID- 11283277 TI - Clostridium botulinum type A haemagglutinin-positive progenitor toxin (HA(+)-PTX) binds to oligosaccharides containing Gal beta1-4GlcNAc through one subcomponent of haemagglutinin (HA1). AB - Haemagglutinin (HA) activity of Clostridium botulinum type A 19S and 16S toxins (HA-positive progenitor toxin; HA(+)-PTX) was characterized. HA titres against human erythrocytes of HA(+)-PTX were inhibited by the addition of lactose, D galactose, N-acetyl-D-galactosamine and D-fucose to the reaction mixtures. A direct glycolipid binding test demonstrated that type A HA(+)-PTX strongly bound to paragloboside and some neutral glycolipids, but did not bind to gangliosides. Type A HA(+)-PTX also bound to asialoglycoproteins (asialofetuin, neuraminidase treated transferrin), but not to sialoglycoproteins (fetuin, transferrin). Although glycopeptidase F treatment of asialofetuin abolished the binding of HA(+)-PTX, endo-alpha-N-acetylgalactosaminidase treatment did not. Thus these results can be interpreted as indicating that type A HA(+)-PTX detects and binds to Gal beta 1-4GlcNAc in paragloboside and the N-linked oligosaccharides of glycoproteins. Regardless of neuraminidase treatment, type A HA(+)-PTX bound to glycophorin A which is a major sialoglycoprotein on the surface of erythrocytes. Both native glycophorin A and neuraminidase-treated glycophorin A inhibited the binding of erythrocytes to type A HA(+)-PTX. Since the N:-linked oligosaccharide of glycophorin A is di-branched and more than 50% of this sugar chain is monosialylated, type A HA(+)-PTX probably bound to the unsialylated branch of the N-linked oligosaccharide of glycophorin A and agglutinated erythrocytes. One subcomponent of HA, designated HA1, did not agglutinate native erythrocytes, although it did bind to erythrocytes, paragloboside and asialoglycoproteins in a manner quite similar to that of HA(+)-PTX. These results indicate that type A HA(+)-PTX binds to oligosaccharides through HA1. PMID- 11283278 TI - Surface exposure and protease insensitivity of Borrelia burgdorferi Erp (OspEF related) lipoproteins. AB - Borrelia burgdorferi can encode numerous lipoproteins of the Erp family. Although initially described as outer surface proteins, the technique used in that earlier study has since been demonstrated to disrupt bacterial membranes and allow labelling of subsurface proteins. Data are now presented from additional analyses indicating that Erp proteins are indeed surface exposed in the outer membrane. Surface localization of these infection-associated proteins indicates the potential for interactions of Erp proteins with vertebrate tissues. Some Erp proteins were resistant to in situ digestion by certain proteases, suggesting that those proteins fold in manners which hide protease cleavage sites, or that they interact with other protective membrane components. Additionally, cultivation of B. burgdorferi in the presence of antibodies directed against Erp proteins inhibited bacterial growth. PMID- 11283279 TI - Analysis of the role of flagellar activity in virulence gene expression in Vibrio cholerae. AB - Expression of virulence factors and motility of VIBRIO: cholerae are intimately linked by an as yet uncharacterized mechanism. Several lines of evidence indicate that the activity of the flagellum of V. cholerae might have an impact on virulence gene regulation, as alterations of the motility phenotype, either by mutation or by inhibitory drugs, result in varied levels of virulence factor production. The Na(+)-driven polar flagella of some VIBRIO: species are proposed to act as mechanosensors, sensing media viscosity. It has been suggested that the V. cholerae flagellum might act as a 'voltmeter', responding to changes in membrane potential, or might sense some environmental conditions that lead to the repression of virulence factors in V. cholerae. To test these hypotheses, beta galactosidase levels of several types of non-motile mutant derivatives of a V. cholerae toxT::lacZ reporter strain were analysed following changes in media viscosity, membrane potential and other environmental conditions. Like the parental strain, the non-motile strain showed increased toxT::lacZ expression in high-viscosity media, suggesting that the sensing of media viscosity does not occur via the flagella. Other molecules that might be able to detect changes in media viscosity could include mechanosensitive (MS) ion channels found in the bacterial membrane. However, a V. cholerae derivative strain mutated in two putative MS channels still showed increased toxT::lacZ expression in high viscosity media, indicating that these putative ion channels of V. cholerae are not involved in the viscosity effect and suggesting an as yet uncharacterized mechanism for sensing of media viscosity. The flagellum does not appear to act as a voltmeter, as beta-galactosidase activities of the non-flagellate derivative strain were found to be similar to those of the parental strain after artificially changing the sodium membrane bioenergetics. Similarly, several environmental conditions known to reduce toxin expression were equally effective in reducing toxT::lacZ expression in the motile or non-motile strains. In conclusion, the flagellum of V. cholerae does not act as a mechanosensor, voltmeter or signal transducer for environmental conditions. Thus, alternative mechanisms for the detection of these conditions must exist that likely do not involve the ToxR molecule, as the sensing of all of the tested parameters occurred when the TcpP/H proteins alone activated the toxT::lacZ reporter gene. PMID- 11283280 TI - Antigenic variation of gonococcal pilin expression in vivo: analysis of the strain FA1090 pilin repertoire and identification of the pilS gene copies recombining with pilE during experimental human infection. AB - Antigenic variation of gonococcal pilin involves a family of variable genes that undergo homologous recombination, resulting in transfer of variant sequences from the pilS silent gene copies into the complete pilE expression locus. Little is known about the specific recombination events that are involved in assembling new variant pilin genes in vivo. One approach to understanding pilin variation in vivo is to carry out experimental human infections with a gonococcal strain having a fully characterized repertoire of pilin genes, so that the specific recombination events occurring in vivo can be determined. To this end, the authors cloned, sequenced and mapped the pilin genes of strain FA1090 of Neisseria gonorrhoeae. This strain contains one pilE locus and 19 silent gene copies that are arranged in five pilS loci; the pilE locus and four of the pilS loci are clustered in a 35 kb region of the chromosome. The general features of the pilin loci in FA1090 are similar to those in strain MS11, in which the mechanism of pilin variation has been extensively studied. However, none of the silent copy sequences are identical in the two strains, which emphasizes the extreme variability in this gene family among gonococci. Three male volunteers were inoculated with the same variant of strain FA1090 and developed urethritis within 2--4 d. The pilE gene sequences from a total of 23 colonies cultured from the subjects were analysed, determining which pilS silent copy donated each portion of the expressed pilE genes. There were 12 different pilin variants, one of which was the original inoculum variant, among the in vivo-expressed pilE gene sequences. The pilE of the inoculum variant was derived entirely from a single silent copy (pilS6c1). However, the pilE genes in the majority of the colonies cultured from the infected subjects were chimeras of sequence derived from two or three silent copies. Recombination to generate new pilE sequences involved exchange of single variable minicassettes, multiple minicassettes, entire silent gene copies, or (rarely) recombination within a minicassette. PMID- 11283281 TI - Type IV O antigen modification genes in the genome of Shigella flexneri NCTC 8296. AB - The genes encoding type IV O antigen glucosylation were characterized from both Escherichia coli and Shigella flexneri. The putative O antigen modification genes from E. coli, o120 o306 o443, were PCR-amplified and introduced into S. flexneri serotype Y strain SFL124. Immunogold labelling and phage sensitivity indicated the presence of both serotype Y and serotype 4a O antigens on the cell surface of the resulting recombinant SFL124 strains, suggesting that only partial serotype conversion was conferred by the E. coli genes. The type IV O antigen modification genes were then isolated and characterized from S. flexneri serotype 4a strain NCTC 8296. A 3.8 kb chromosomal fragment conferred complete conversion to serotype 4a when introduced into SFL124. Sequence analysis of the fragment revealed the presence of three genes, gtrA(IV) gtrB(IV) gtrIV(Sf). DNAs homologous to bacteriophage int and attP were located upstream of gtrA(IV), suggesting that this region of the NCTC 8296 genome may have originated from a bacteriophage; however, a serotype-converting phage could not be induced from this strain nor from other strains used in this study. Comparison of the GtrIV(Sf) and GtrIV(Ec) (o443) proteins revealed that they are 41% identical and 63% similar, which is the highest degree of similarity reported among the S. flexneri O antigen glucosyltransferases. PMID- 11283282 TI - Antibody response against Escherichia coli heat-stable enterotoxin expressed as fusions to flagellin. AB - The heat-stable toxin (ST) produced by enterotoxigenic Escherichia coli strains causes diarrhoea by altering the fluid secretion in intestinal epithelial cells. Here, the effectiveness of a flagellin fusion protein of Salmonella containing a 19-amino-acid sequence derived from the ST sequence (FLA--ST) in generating antibodies capable of neutralizing the toxic activity of ST was evaluated. This fusion protein, and an alternative construction where two cysteine residues in the ST sequence were substituted by alanines (ST(mt)), were delivered to the immune system by three distinct strategies: (i) orally, using an attenuated Salmonella strain expressing FLA--ST; (ii) intraperitoneally, by injection of purified FLA--ST; (iii) orally, using attenuated Salmonella carrying a eukaryotic expression plasmid (pCDNA3) with the gene encoding FLA-ST. The results showed that the flagellin system can be used as a carrier to generate ST-neutralizing antibodies. However, it should be mentioned that humoral immune response against ST was only obtained when the mutated ST sequence was employed. FLA-ST was found to be non-immunogenic when delivered via the oral route with attenuated Salmonella strains. However, a flagellin antibody response was obtained by immunizing mice with Salmonella carrying pCDNA3/FLA-ST(mt). Oral immunization with Salmonella carrying the eukaryotic expression plasmid (pCDNA3/FLA--ST(mt)) seems to be a promising method to elicit an appropriate response against fusions to flagellin. PMID- 11283283 TI - Differences in sialic acid density in pathogenic and non-pathogenic Aspergillus species. AB - ASPERGILLUS: fumigatus is a ubiquitous soil fungus that causes invasive lung disease in the immunocompromised host. The structure of the conidial wall has not been well characterized although it is thought that adhesins present on the surface are involved in attachment of the conidia to host lung cells and proteins, which is a prerequisite for the establishment of infection. Negatively charged carbohydrates on the conidial surface have been previously identified as the molecules responsible for attachment of conidia to extracellular matrix proteins. The aim of this research was to identify carbohydrates on the conidial surface that contribute to its negative charge. Direct chemical analysis and indirect binding assays have demonstrated that A. fumigatus possesses sialic acids on the conidial surface. Pre-treatment of A. fumigatus conidia with sialidase decreased binding of a sialic acid-specific lectin, Limax flavus agglutinin (LFA), to the conidial surface and decreased adhesion of conidia to the positively charged polymer poly L-lysine. Two other sialic acid-specific lectins, Maackia amurensis agglutinin and Sambucus nigra agglutinin, exhibited negligible binding to A. fumigatus conidia indicating that 2,3-alpha- and 2,6 alpha-linked sialic acids are not the major structures found on the conidial surface. Mild acid hydrolysis and purification of conidial wall carbohydrates yielded a product that had the same R(F) as the Neu5Ac standard when analysed by high-performance thin-layer chromatography. A density of 6.7 x 10(5) sialic acid residues per conidium was estimated using a colorimetric assay. Conidia grown on a minimal medium lacking sialic acid also reacted with LFA, indicating that sialic acid biosynthesis occurs de novo. Sialic acid biosynthesis was shown to be regulated by nutrient composition: the density of sialic acids on the surface of conidia grown in minimal media was lower than that observed when conidia were grown on rich, complex media. It has previously been shown that pathogenic Aspergillus species adhere to basal lamina proteins to a greater extent than non pathogenic Aspergillus species. To determine whether the expression of sialic acid on the conidial surface was correlated with adhesion to basal lamina, conidia from other non-pathogenic Aspergillus species were tested for their reactivity towards LFA. Flow cytometric analysis demonstrated that A. fumigatus had a significantly greater sialic acid density than three non-pathogenic Aspergillus species. Sialic acids on the conidial wall may be involved in adhesion to fibronectin, a component of the basal lamina, as binding of A. fumigatus conidia to fibronectin was strongly inhibited in the presence of a sialylated glycoprotein. PMID- 11283284 TI - The role and relevance of phospholipase D1 during growth and dimorphism of Candida albicans. AB - The phosphatidylcholine-specific phospholipase D1 (PLD1) in Saccharomyces cerevisiae is involved in vesicle transport and is essential for sporulation. The gene encoding the homologous phospholipase D1 from Candida albicans (PLD1) was used to study the role of PLD1 in this pathogenic fungus. In vitro and in vivo expression studies using Northern blots and reverse transcriptase-PCR showed low PLD1 mRNA levels in defined media supporting yeast growth and during experimental infection, while enhanced levels of PLD1 transcripts were detected during the yeast to hyphal transition. To study the relevance of PLD1 during yeast and hyphal growth, an essential part of the gene was deleted in both alleles of two isogenic strains. In vitro PLD1 activity assays showed that pld1 mutants produced no detectable levels of phosphatidic acid, the hydrolytic product of PLD1 activity, and strongly reduced levels of diacylglycerol, the product of lipid phosphate phosphohydrolase, suggesting no or a negligible background PLD1 activity in the pld1 mutants. The pld1 mutants showed no growth differences compared to the parental wild-type in liquid complex and minimal media, independent of the growth temperature. In addition, growth rates of pld1 mutants in media with protein as the sole source of nitrogen were similar to growth rates of the wild-type, indicating that secretion of proteinases was not reduced. Chlamydospore formation was normal in pld1 mutants. When germ tube formation was induced in liquid media, pld1 mutants showed similar rates of yeast to hyphal transition compared to the wild-type. However, no hyphae formation was observed on solid Spider medium, and cell growth on cornmeal/Tween 80 medium indicated aberrant morphogenesis. In addition, pld1 mutants growing on solid media had an attenuated ability to invade the agar. In a model of oral candidosis, pld1 mutants showed no attenuation of virulence. In contrast, the mutant was less virulent in two different mouse models. These data suggest that PLD1 is not essential for growth and oral infections. However, they also suggest that a prominent part of the phosphatidic acid and diacylglycerol pools is produced by PLD1 and that the level of these components is important for morphological transitions under certain conditions in C. albicans. PMID- 11283285 TI - Hybrid genotypes in the pathogenic yeast Cryptococcus neoformans. AB - Amplified fragment length polymorphism (AFLP) genotyping of isolates of the pathogenic fungus Cryptococcus neoformans suggested a considerable genetic divergence between the varieties C. neoformans var. neoformans and C. neoformans var. grubii on the one hand versus C. neoformans var. gattii on the other. This divergence is supported by additional phenotypic, biochemical, clinical and molecular differences. Therefore, the authors propose the existence of two species, C. neoformans (Sanfelice) Vuillemin and C. bacillisporus Kwon-Chung, which differ in geographical distribution, serotypes and ecological origin. Within each species three AFLP genotypes occur, which differ in geographical distribution and serotypes. Differences in ecological origin (AIDS patients, non AIDS patients, animals or the environment) were found to be statistically not significant. In C. neoformans as well as in C. bacillisporus one of the genotypes represented a hybrid. The occurrence of hybridization has consequences for the reproductive biology of the species, as new genotypes with altered virulence or susceptibility to antifungal drugs may arise through the exchange of genetic material. PMID- 11283287 TI - The Bacillus subtilis yabQ gene is essential for formation of the spore cortex. AB - An extensive screening for transcripts with probes specific for the genes in a 108 kb region from rrnO to spo0H of the Bacillus subtilis chromosome led to identification of an operon, yabP--yabQ--divIC--yabR, the expression of which was initiated at the second hour of sporulation and in a sigma(E)-dependent manner. Among three y genes in the operon, deletion of the yabQ gene, which is predicted to encode a protein product of 468 residues with five membrane-spanning domains, resulted in a large decrease in numbers of chloroform-, lysozyme- and heat resistant spores, compared to findings with the wild-type strain. Electron microscopy revealed that development of the spore cortex was blocked in the yabQ mutant. In addition, although the spore coat was visible, the inner coat layer of the mutant seemed partially detached from the outer coat. In sporangia of the strains harbouring an in-frame fusion of the green fluorescent protein gene to yabQ, fluorescence was detected around the forespore. This localization did not depend on SpoIVA or on CotE functions, both of which determine proper localization of coat proteins and cortex formation. The yabQ deletion did not affect expression of genes involved in cortex synthesis. These results suggest that the YabQ protein localizes in the membrane of the forespore and plays an important role in cortex formation. PMID- 11283288 TI - Motions caused by the growth of Bacillus subtilis macrofibres in fluid medium result in new forms of movement of the multicellular structures over solid surfaces. AB - Bacillus subtilis macrofibres, highly ordered multicellular structures, undergo twisting and writhing motions when they grow in fluid medium as a result of forces generated by the elongation of individual cells. Macrofibres are denser than the fluid medium in which they are cultured, consequently they settle to the bottom of the growth chamber and grow in contact with it. The ramifications of growth on plastic and glass surfaces were examined. Macrofibres were observed to rotate about a vertical axis near the centre of their length in a chiral-specific direction. Right-handed fibres rotated clockwise on plastic surfaces at approximately 4 degrees min(-1), left-handed structures of lower twist rotate anti-clockwise at about half that rate. Very large ball structures produced late in macrofibre formation perched on many small protruding fibres but rotated only when driven by large fibres attached to their periphery. Closer examination showed that fibres made contact with surfaces at only a few points along their length (between 1 and 6 on glass). The regions in contact with the surface changed periodically as a result of rotation of the fibre shaft caused by growth. Every time the weight of a fibre transferred from one contact point to another, each section of the fibre took a small step approximately proportional to its distance from the fibre mid-point. The net result was a rolling of each section over the surface so that the fibre rotation about a vertical axis was produced. Macrofibres also took large steps when part of the structure rose off the floor, swept through an arc in the fluid and then returned to the floor at a new location. The rate of movement during a large step, measured as the change of angle between the moving and stationary portions of the fibre, was 5 degrees s( 1). These observations reveal that the forces derived from helical growth that lead to macrofibre formation also cause characteristic macrofibre motion that differs from classical motility. PMID- 11283286 TI - Detergent-independent in vitro activity of a truncated Bacillus signal peptidase. AB - The Gram-positive eubacterium Bacillus subtilis contains five chromosomally encoded type I signal peptidases (SPases) for the processing of secretory pre proteins. Even though four of these SPases, denoted SipS, SipT, SipU and SipV, are homologous to the unique SPase I of Escherichia coli, they are structurally different from that enzyme, being almost half the size and containing one membrane anchor instead of two. To investigate whether the unique membrane anchor of Bacillus SPases is required for in vitro activity, soluble forms of SipS of B. subtilis, SipS of Bacillus amyloliquefaciens and SipC of the thermophile Bacillus caldolyticus were constructed. Of these three proteins, only a hexa-histidine tagged soluble form of SipS of B. amyloliquefaciens could be isolated in significant quantities. This protein displayed optimal activity at pH 10, which is remarkable considering the fact that the catalytic domain of SPases is located in an acidic environment at the outer surface of the membrane of living cells. Strikingly, in contrast to what has been previously reported for the soluble form of the E. coli SPase, soluble SipS was active in the absence of added detergents. This observation can be explained by the fact that a highly hydrophobic surface domain of the E. coli SPase, implicated in detergent-binding, is absent from SipS. PMID- 11283289 TI - Frozen motion of gliding bacteria outlines inherent features of the motility apparatus. AB - High-resolution data of actively gliding wild-type bacteria of four different species and of four different gliding mutants of Myxococcus xanthus were obtained from scanning electron micrographs. By shock freezing and freeze drying, motility associated surface patterns could be fixed and consequently distinct intermediate states of motion could be observed for the first time. It is shown that these topographic patterns are immediately lost when gliding motility is stopped by blocking the respiratory chain with potassium cyanide or sodium azide. From the surface topography, the mode of action of the gliding apparatus of all four bacterial species examined can be described as a twisted circularly closed 'band'. During gliding, groups of nodes of the supertwisted apparatus show evidence of travelling like waves along the trichomes. However, the spacing between the nodes is not constant but varies within a certain range. This indicates that they are flexibly modulated as a consequence of the gliding state of the individual trichome. PMID- 11283290 TI - New members of the ctrA regulon: the major chemotaxis operon in Caulobacter is CtrA dependent. AB - The Caulobacter crescentus che promoter region consists of two divergent promoters, directing expression of the major chemotaxis operon and a novel gene cagA (chemotaxis associated gene A). Analyses of start sites by primer extension and alignment of the divergent promoters revealed significant similarities between them at the -35 promoter region. Both mcpA and cagA are differentially expressed in the cell cycle, with maximal activation of transcription in predivisional cells. The main difference between the mcpA and cagA promoters is that, in common with the fljK flagellin, cagA is expressed in swarmer cells. A cagA--lacZ promoter fusion that contains 36 bases of untranslated mRNA has sufficient information to segregate the lacZ transcript to swarmer cells. Expression of mcpA and cagA was dependent on DNA replication. Transcriptional epistasis experiments were performed to identify potential regulators in the flagellar hierarchy. The sigma factor RpoN, which is required for flagellar biogenesis, is not required for mcpA and cagA expression. Mutations in the genes for the MS-ring and the switch complex (flagellar class II mutants) do not affect expression of mcpA and cagA. However, CtrA, an essential response regulator of flagellar gene transcription, is required. PMID- 11283292 TI - The product of the ybdE gene of the Escherichia coli chromosome is involved in detoxification of silver ions. AB - Transcription of the ybcZ--ylcA ylcBCD--ybdE region of the Escherichia coli K38 chromosome was analysed by Northern RNA--DNA hybridization, RT-PCR and primer extension. Transcription of a dicistronic ybcZ--ylcA mRNA and a tetracistronic ylcBCD--ybdE mRNA was induced by silver and was initiated from the sigma-70 promoters ylcAp and ylcBp. Expression of beta-galactosidase activity from a Phi(ylcBp--lacZ) operon fusion was also induced by Ag(+) and Cu(2+), but not by Zn(2+). In-frame deletion of ybdE from the chromosome yielded a silver-sensitive E. coli mutant strain which did not differ in its copper resistance from its wild type strain. On the other hand, deletion of the copA gene for the copper exporting P-type ATPase CopA resulted in copper sensitivity, but not in silver sensitivity. A Delta ybdE Delta copA double mutant strain behaved towards copper as the Delta copA strain and towards silver as the Delta ybdE strain. Thus, in E. coli, the YlcBCD--YbdE system may be involved in silver- but not in copper resistance, and CopA may be involved in copper- but not in silver resistance. PMID- 11283291 TI - A new simvastatin (mevinolin)-resistance marker from Haloarcula hispanica and a new Haloferax volcanii strain cured of plasmid pHV2. AB - The mevinolin-resistance determinant, hmg, encodes the enzyme 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase and is a commonly used selectable marker in halobacterial genetics. Plasmids bearing this marker suffer from instability in Haloferax volcanii because the resistance gene was derived from the genome of this species and is almost identical in sequence to the chromosomal copy. In order to reduce the level of homologous recombination between introduced plasmid vectors and the chromosome of Haloferax, a homologue of the hmg determinant was obtained from the distantly related organism, Haloarcula hispanica. The nucleotide sequences of the wild-type genes (hmgA) of these two species are only 78% identical, and the predicted protein sequences show 71% identity. In comparison to the wild-type hmgA gene, the resistance gene from a mutant resistant to simvastatin (an analogue of mevinolin) showed a single base substitution in the putative promoter. Plasmids constructed using the new resistance determinant were stably maintained under selection in Hfx. volcanii and possessed very low recombination rates with the chromosome of this species. In addition, an improved strain of Hfx. volcanii was developed to overcome the plasmid instability and growth reduction observed in the commonly used WFD11 strain. PMID- 11283293 TI - Transcriptional regulation of styrene degradation in Pseudomonas putida CA-3. AB - The styrene degradative pathway in Pseudmonas putida CA-3 has previously been shown to be divided into an upper pathway involving the conversion of styrene to phenylacetic acid and a lower pathway for the subsequent degradation of phenylacetic acid. It is reported here that expression of the regulatory genes styS and styR is essential for transcription of the upper pathway, but not for degradation of the lower pathway inducer, phenylacetic acid. The presence of phenylacetic acid in the growth medium completely repressed the upper pathway enzymes even in the presence of styrene, the upper pathway inducer. This repression is mediated at the transcription level by preventing expression of the styS and styR regulatory genes. Finally, an examination was made of the various stages of the diauxic growth curve obtained when P. putida CA-3 was grown on styrene together with an additional carbon source and it is reported that catabolite repression may involve a different mechanism to transcriptional repression by an additional carbon source. PMID- 11283294 TI - Classification of rhizobia based on nodC and nifH gene analysis reveals a close phylogenetic relationship among Phaseolus vulgaris symbionts. AB - The nodC and nifH genes were characterized in a collection of 83 rhizobial strains which represented 23 recognized species distributed in the genera Rhizobium, Sinorhizobium, Mesorhizobium and Bradyrhizobium, as well as unclassified rhizobia from various host legumes. Conserved primers were designed from available nucleotide sequences and were able to amplify nodC and nifH fragments of about 930 bp and 780 bp, respectively, from most of the strains investigated. RFLP analysis of the PCR products resulted in a classification of these rhizobia which was in general well-correlated with their known host range and independent of their taxonomic status. The nodC and nifH fragments were sequenced for representative strains belonging to different genera and species, most of which originated from Phaselous vulgaris nodules. Phylogenetic trees were constructed and revealed close relationships among symbiotic genes of the Phaseolus symbionts, irrespective of their 16S-rDNA-based classification. The nodC and nifH phylogenies were generally similar, but cases of incongruence were detected, suggesting that genetic rearrangements have occurred in the course of evolution. The results support the view that lateral genetic transfer across rhizobial species and, in some instances, across Rhizobium and Sinorhizobium genera plays a role in diversification and in structuring the natural populations of rhizobia. PMID- 11283295 TI - Rapid phenotypic change and diversification of a soil bacterium during 1000 generations of experimental evolution. AB - Evolutionary pathways open to even relatively simple organisms, such as bacteria, may lead to complex and unpredictable phenotypic changes, both adaptive and non adaptive. The evolutionary pathways taken by 18 populations of Ralstonia strain TFD41 while they evolved in defined environments for 1000 generations were examined. Twelve populations evolved in liquid media, while six others evolved on agar surfaces. Phenotypic analyses of these derived populations identified some changes that were consistent across all populations and others that differed among them. The evolved populations all exhibited morphological changes in their cell envelopes, including reductions of the capsule in each population and reduced prostheca-like surface structures in most populations. Mean cell length increased in most populations (in one case by more than fourfold), although a few populations evolved shorter cells. Carbon utilization profiles were variable among the evolved populations, but two distinct patterns were correlated with genetic markers introduced at the outset of the experiment. Fatty acid methyl ester composition was less variable across populations, but distinct patterns were correlated with the two physical environments. All 18 populations evolved greatly increased sensitivity to bile salts, and all but one had increased adhesion to sand; both patterns consistent with changes in the outer envelope. This phenotypic diversity contrasts with the fairly uniform increases in competitive fitness observed in all populations. This diversity may represent a set of equally probable adaptive solutions to the selective environment; it may also arise from the chance fixation of non-adaptive mutations that hitchhiked with a more limited set of beneficial mutations. PMID- 11283296 TI - Characterization of lactobacilli from Scotch malt whisky distilleries and description of Lactobacillus ferintoshensis sp. nov., a new species isolated from malt whisky fermentations. AB - Sixty-four strains of Lactobacillus were isolated from fermentation samples from 23 malt whisky distilleries located in the major whisky producing regions of Scotland. The strains were assigned to 26 ribotype patterns. Strains of some ribotype patterns were widely distributed and recovered from distilleries throughout Scotland, while strains representing other ribotypes were particular to a specific region or even a certain distillery. Repeated sampling of a single distillery over a 12 month period showed that the range of bacteria present, as indicated by ribotyping, was stable, but was influenced by changes in malt supply and the period of closure for annual maintenance. Partial 16S rDNA sequence analysis of ribotype representatives revealed Lactobacillus brevis, Lactobacillus fermentum, Lactobacillus paracasei and Lactobacillus pentosus as the major species present in the distilleries; however, four isolates could not be identified by this procedure. Determination of the full 16S rDNA gene sequence from one of these isolates (strain R7-84) revealed >98.5% similarity to Lactobacillus buchneri and its phylogenetic neighbours. DNA from two other strains showed greater than 70% hybridization to DNA from R7-84 under non stringent renaturation conditions and DNA from strain R7-84 shared less than 65% hybridization with members of the L. buchneri group. It is proposed that these three strains should be placed in a new species for which the name Lactobacillus ferintoshensis represented by the type strain R7-84(T) is suggested. PMID- 11283297 TI - Intracellular pH regulation by Mycobacterium smegmatis and Mycobacterium bovis BCG. AB - Mycobacteria are likely to encounter acidic pH in the environments they inhabit; however intracellular pH homeostasis has not been investigated in these bacteria. In this study, Mycobacterium smegmatis and Mycobacterium bovis [Bacille Calmette- Guerin (BCG)] were used as examples of fast- and slow-growing mycobacteria, respectively, to study biochemical and physiological responses to acidic pH. M. smegmatis and M. bovis BCG were able to grow at pH values of 4.5 and 5.0, respectively, suggesting the ability to regulate internal pH. Both species of mycobacteria maintained their internal pH between pH 6.1 and 7.2 when exposed to decreasing external pH and the maximum Delta pH observed was approximately 2.1 to 2.3 units for both bacteria. The Delta pH of M. smegmatis at external pH 5.0 was dissipated by protonophores (e.g. carbonyl cyanide m-chlorophenylhydrazone), ionophores (e.g. monensin and nigericin) and N,N'-dicyclohexylcarbodiimide (DCCD), an inhibitor of the proton-translocating F(1)F(0)-ATPase. These results demonstrate that permeability of the cytoplasmic membrane to protons and proton extrusion by the F(1)F(0)-ATPase plays a key role in maintaining internal pH near neutral. Correlations between measured internal pH and cell viability indicated that the lethal internal pH for both strains of mycobacteria was less than pH 6.0. Compounds that decreased internal pH caused a rapid decrease in cell survival at acidic pH, but not at neutral pH. These data indicate that both strains of mycobacteria exhibit intracellular pH homeostasis and this was crucial for the survival of these bacteria at acidic pH values. PMID- 11283298 TI - Effect of hydrolysable and condensed tannins on growth, morphology and metabolism of Streptococcus gallolyticus (S. caprinus) and Streptococcus bovis. AB - Streptococcus gallolyticus (S. caprinus) was resistant in vitro to at least 7% (w/v) tannic acid and 4% (w/v) acacia condensed tannin, levels 10-fold greater than those tolerated by S. bovis. Growth of S. gallolyticus in liquid medium was characterized by a lag period which increased, and a growth rate which decreased, with increasing tannin concentration. S. gallolyticus was also more tolerant to the presence of simple phenolic acid monomers than was S. bovis, but the lag period was still concentration dependent. Gallate decarboxylase activity in S. gallolyticus was elevated in the presence of tannic acid or gallic acid but not with other phenolic acids. Scanning electron microscopic analysis showed that both the size and shape of S. gallolyticus and S. bovis changed in response to tannin but only S. gallolyticus was surrounded by an extracellular polysaccharide matrix which accumulated in a tannin-concentration-dependent fashion. Washing of the cells to remove extracellular polysaccharide increased the lag period of S. gallolyticus in the presence of 1% (w/v) tannic acid from 4 h to 6 h. In contrast, increasing extracellular polysaccharide synthesis in S. bovis did not increase its tolerance to tannic acid. These data demonstrate that S. gallolyticus has developed a number of mechanisms to reduce the potential effect of tannins on cell growth, and that these mechanisms provide the organism with a selective advantage over S. bovis when grown in the presence of tannins. PMID- 11283299 TI - The glucomannokinase of Prevotella bryantii B(1)4 and its potential role in regulating beta-glucanase expression. AB - Prevotella bryantii B(1)4 has a transport system for glucose and mannose, but beta-glucanase expression is only catabolite-repressed by glucose. P bryantii B(1)4 cell extracts had ATP-dependent gluco- and mannokinase activities, and significant phosphoenolpyruvate- or GTP-dependent hexose phosphorylation was not observed. Mannose inhibited glucose phosphorylation (and vice versa), and activity gels indicated that a single protein was responsible for both activities. Glucose was phosphorylated at a faster rate than was mannose [V(max) 280 nmol hexose (mg protein)(-1) min(-1) versus 60 nmol hexose (mg protein)(-1) min(-1), respectively] and glucose was a better substrate for the kinase (K(m) 0.12 mM versus 1.2 mM, respectively). The purified glucomannokinase (1250-fold) had a molecular mass of 68 kDa, but SDS-PAGE gels indicated that it was a dimer (monomer 34.5 kDa). The N-terminus (25 residues) had an 8 amino acid segment that was homologous to other bacterial glucokinases. The glucomannokinase was competitively inhibited by the nonmetabolizable glucose analogue 2-deoxyglucose (2DG), and cells grown with glucose and 2DG had lower rates of glucose consumption than did cells given only glucose. When the ratio of 2DG to glucose was increased, the glucose consumption rate decreased and the beta-glucanase activity increased. The glucose consumption rate and the glucomannokinase activity of cells treated with 2DG were highly correlated (r(2)=0.98). This result suggested that glucomannokinase activity was either directly or indirectly regulating beta-glucanase expression. PMID- 11283301 TI - Topological investigations of the FomA porin from Fusobacterium nucleatum and identification of the constriction loop L6. AB - Porin FomA in the outer membrane of Fusobacterium nucleatum is a trimeric protein, which exhibits permeability properties similar to that of the well-known enterobacterial diffusion porins. The proposed topology model of the FomA monomer depicts the beta-barrel motif typical of diffusion porins, consisting of 16 antiparallel beta-strands. To investigate the accuracy of the FomA model and assess the topological relationship with other porins, individual deletions of variable size in seven of the eight surface-exposed regions of the porin were genetically engineered. Deletions in the predicted loops L1 to L7 were tolerated by the FomA porins, as judged by a normal assembly in the outer membrane of Escherichia coli and a sustained pore-forming ability. Deletions in the largest proposed external region, loop L6, made the FomA porins considerably more permeable to antibiotics, indicating larger pore channels. The distinctly increased uptake rates and size exclusion limits displayed by the L6 deletion mutant porins, suggest that loop L6 folds back into the beta-barrel thereby constricting the native FomA channel. Thus, the position of the channel constriction loop appears to be shifted towards the C terminus in the FomA porin, as compared to the crystal structures of five non-specific diffusion porins. PMID- 11283300 TI - Defining the mycoplasma 'cytoskeleton': the protein composition of the Triton X 100 insoluble fraction of the bacterium Mycoplasma pneumoniae determined by 2-D gel electrophoresis and mass spectrometry. AB - After treating Mycoplasma pneumoniae cells with the nonionic detergent Triton X 100, an undefined, structured protein complex remains that is called the 'Triton X-100 insoluble fraction' or 'Triton shell'. By analogy with eukaryotic cells and supported by ultrastructural analyses it is supposed that this fraction contains the components of a bacterial cytoskeleton-like structure. In this study, the composition of the Triton X-100 insoluble fraction was defined by electron microscopic screening for possible structural elements, and by two-dimensional (2 D) gel electrophoresis and MS to identify the proteins present. Silver staining of 2-D gels revealed about 100 protein spots. By staining with colloidal Coomassie blue, about 50 protein spots were visualized, of which 41 were identified by determining the mass and partial sequence of tryptic peptides of individual proteins. The identified proteins belonged to several functional categories, mainly energy metabolism, translation and heat-shock response. In addition, lipoproteins were found and most of the proteins involved in cytadherence that were previously shown to be components of the Triton X-100 insoluble fraction. There were also 11 functionally unassigned proteins. Based on sequence-derived predictions, some of these might be potential candidates for structural components. Quantitatively, the most prevalent proteins were the heat shock protein DnaK, elongation factor Tu and subunits alpha and beta of the pyruvate dehydrogenase complex (PdhA, PdhB), but definite conclusions regarding the composition of the observed structures can only be drawn after specific proteins are assigned to them, for example by immunocytochemistry. PMID- 11283302 TI - The gene yghK linked to the glc operon of Escherichia coli encodes a permease for glycolate that is structurally and functionally similar to L-lactate permease. AB - In Escherichia coli the glc operon involved in glycolate utilization is located at 67.3 min and formed by genes encoding the enzymes glycolate oxidase (glcDEF) and malate synthase G (glcB). Their expression from a single promoter upstream of glcD is induced by growth on glycolate and regulated by the activator encoded by the divergently transcribed gene glcC. Gene yghK, located 350 bp downstream of glcB, encodes a hydrophobic protein highly similar to the L-lactate permease encoded by lldP. Expression studies have shown that the yghK gene (proposed name glcA) is transcribed from the same promoter as the other glc structural genes and thus belongs to the glc operon. Characterization of a glcA::cat mutant showed that GlcA acts as glycolate permease and that glycolate can also enter the cell through another transport system. Evidence is presented of the involvement of L lactate permease in glycolate uptake. Growth on this compound was abolished in a double mutant of the paralogous genes glcA and lldP, and restored with plasmids expressing either GlcA or LldP. Characterization of the putative substrates for these two related permeases showed, in both cases, specificity for the 2 hydroxymonocarboxylates glycolate, L-lactate and D-lactate. Although both GlcA and LldP recognize D-lactate, mutant analysis proved that L-lactate permease is mainly responsible for its uptake. PMID- 11283304 TI - The role and regulation of adenosine in the central nervous system. AB - Adenosine is a modulator that has a pervasive and generally inhibitory effect on neuronal activity. Tonic activation of adenosine receptors by adenosine that is normally present in the extracellular space in brain tissue leads to inhibitory effects that appear to be mediated by both adenosine A1 and A2A receptors. Relief from this tonic inhibition by receptor antagonists such as caffeine accounts for the excitatory actions of these agents. Characterization of the effects of adenosine receptor agonists and antagonists has led to numerous hypotheses concerning the role of this nucleoside. Previous work has established a role for adenosine in a diverse array of neural phenomena, which include regulation of sleep and the level of arousal, neuroprotection, regulation of seizure susceptibility, locomotor effects, analgesia, mediation of the effects of ethanol, and chronic drug use. PMID- 11283303 TI - PDZ domains and the organization of supramolecular complexes. AB - PDZ domains are modular protein interaction domains that bind in a sequence specific fashion to short C-terminal peptides or internal peptides that fold in a beta-finger. The diversity of PDZ binding specificities can be explained by variable amino acids lining the peptide-binding groove of the PDZ domain. Abundantly represented in Caenorhabditis elegans, Drosophila melanogaster, and mammalian genomes, PDZ domains are frequently found in multiple copies or are associated with other protein-binding motifs in multidomain scaffold proteins. PDZ-containing proteins are typically involved in the assembly of supramolecular complexes that perform localized signaling functions at particular subcellular locations. Organization around a PDZ-based scaffold allows the stable localization of interacting proteins and enhances the rate and fidelity of signal transduction within the complex. Some PDZ-containing proteins are more dynamically regulated in distribution and may also be involved in the trafficking of interacting proteins within the cell. PMID- 11283305 TI - Localization and globalization in conscious vision. AB - The primate visual brain consists of many separate, functionally specialized processing systems, each consisting of several apparently hierarchical stages or nodes. The evidence reviewed here leads me to speculate (a) that the processing systems are autonomous with respect to one another, (b) that activity at each node reaches a perceptual end point at a different time, resulting in a perceptual asynchrony in vision, and (c) that, consequently, activity at each node generates a microconsciousness. Visual consciousness is therefore distributed in space and time, with the universal organizing principle of abstraction applied separately within each processing system. The consequence of spatially and temporally distributed microconsciousnesses is that their integration is a multistage, nonhierarchical process that may involve a neural "glue." PMID- 11283306 TI - Glial control of neuronal development. AB - Reciprocal interactions between differentiating glial cells and neurons define the course of nervous system development even before the point at which these two cell types become definitively recognizable. Glial cells control the survival of associated neurons in both Drosophila and mammals, but this control is dependent on the prior neuronal triggering of glial cell fate commitment and trophic factor expression. In mammals, the growth factor neuregulin-1 and its receptors of the ErbB family play crucial roles in both events. Similarly, early differentiating neurons and their associated glia rely on reciprocal signaling to establish the basic axon scaffolds from which neuronal connections evolve. The importance of this interactive signaling is illustrated by the action of glial transcription factors and of glial axon guidance cues such as netrin and slit, which together regulate the commissural crossing of pioneer axons at the neural midline. In these and related events, the defining principle is one of mutually reinforced and mutually dependent signaling that occurs in a network of developing neurons and glia. PMID- 11283307 TI - Touch and go: decision-making mechanisms in somatosensation. AB - A complex sequence of neural events unfolds between sensory receptor activation and motor activity. To understand the underlying decision-making mechanisms linking somatic sensation and action, we ask what components of the neural activity evoked by a stimulus are directly related to psychophysical performance, and how are they related. We find that single-neuron responses in primary and secondary somatosensory cortices account for the observed performance of monkeys in vibrotactile discrimination tasks, and that neuronal and behavioral responses covary in single trials. This sensory activity, which provides input to memory and decision-making mechanisms, is modulated by attention and behavioral context, and microstimulation experiments indicate that it may trigger normal perceptual experiences. Responses recorded in motor areas seem to reflect the output of decision-making operations, which suggests that the ability to make decisions occurs at the sensory-motor interface. PMID- 11283308 TI - Synaptic modification by correlated activity: Hebb's postulate revisited. AB - Correlated spiking of pre- and postsynaptic neurons can result in strengthening or weakening of synapses, depending on the temporal order of spiking. Recent findings indicate that there are narrow and cell type-specific temporal windows for such synaptic modification and that the generally accepted input- (or synapse ) specific rule for modification appears not to be strictly adhered to. Spike timing-dependent modifications, together with selective spread of synaptic changes, provide a set of cellular mechanisms that are likely to be important for the development and functioning of neural networks. When an axon of cell A is near enough to excite cell B or repeatedly or consistently takes part in firing it, some growth or metabolic change takes place in one or both cells such that A's efficiency, as one of the cells firing B, is increased. PMID- 11283309 TI - An integrative theory of prefrontal cortex function. AB - The prefrontal cortex has long been suspected to play an important role in cognitive control, in the ability to orchestrate thought and action in accordance with internal goals. Its neural basis, however, has remained a mystery. Here, we propose that cognitive control stems from the active maintenance of patterns of activity in the prefrontal cortex that represent goals and the means to achieve them. They provide bias signals to other brain structures whose net effect is to guide the flow of activity along neural pathways that establish the proper mappings between inputs, internal states, and outputs needed to perform a given task. We review neurophysiological, neurobiological, neuroimaging, and computational studies that support this theory and discuss its implications as well as further issues to be addressed PMID- 11283310 TI - The physiology of stereopsis. AB - Binocular disparity provides the visual system with information concerning the three-dimensional layout of the environment. Recent physiological studies in the primary visual cortex provide a successful account of the mechanisms by which single neurons are able to signal disparity. This work also reveals that additional processing is required to make explicit the types of signal required for depth perception (such as the ability to match features correctly between the two monocular images). Some of these signals, such as those encoding relative disparity, are found in extrastriate cortex. Several other lines of evidence also suggest that the link between perception and neuronal activity is stronger in extrastriate cortex (especially MT) than in the primary visual cortex. PMID- 11283311 TI - Paraneoplastic neurologic disease antigens: RNA-binding proteins and signaling proteins in neuronal degeneration. AB - Studies of the disorders known as paraneoplastic neurologic degenerations exemplify the successful application of modern molecular biological techniques to diseases, yielding, even for these extremely rare disorders, wide-ranging insight into basic neurobiology, tumor immunity, and autoimmune neurologic disease. Immune responses to paraneoplastic neurologic degeneration antigens, also called onconeural antigens, have been exploited to clone and characterize a number of neuron-specific proteins, including several RNA-binding proteins and new kinds of signaling molecules. The biology and functions of these proteins are reviewed, and a model in which their functions are related to the pathogenesis of autoimmune neurologic disease is discussed. PMID- 11283312 TI - Odor encoding as an active, dynamical process: experiments, computation, and theory. AB - We examine early olfactory processing in the vertebrate and insect olfactory systems, using a computational perspective. What transformations occur between the first and second olfactory processing stages? What are the causes and consequences of these transformations? To answer these questions, we focus on the functions of olfactory circuit structure and on the role of time in odor-evoked integrative processes. We argue that early olfactory relays are active and dynamical networks, whose actions change the format of odor-related information in very specific ways, so as to refine stimulus identification. Finally, we introduce a new theoretical framework ("winnerless competition") for the interpretation of these data. PMID- 11283313 TI - Protein synthesis at synaptic sites on dendrites. AB - Studies over the past 20 years have revealed that gene expression in neurons is carried out by a distributed network of translational machinery. One component of this network is localized in dendrites, where polyribosomes and associated membranous elements are positioned beneath synapses and translate a particular population of dendritic mRNAs. The localization of translation machinery and mRNAs at synapses endows individual synapses with the capability to independently control synaptic strength through the local synthesis of proteins. The present review discusses recent studies linking synaptic plasticity to dendritic protein synthesis and mRNA trafficking and considers how these processes are regulated. We summarize recent information about how synaptic signaling is coupled to local translation and to the delivery of newly transcribed mRNAs to activated synaptic sites and how local translation may play a role in activity-dependent synaptic modification. PMID- 11283314 TI - Signaling and transcriptional mechanisms in pituitary development. AB - During the development of the pituitary gland, distinct hormone-producing cell types arise from a common population of ectodermal progenitors, providing an instructive model system for elucidating the molecular mechanisms of patterning and cell type specification in mammalian organogenesis. Recent studies have established that the development of the pituitary occurs through multiple sequential steps, allowing the coordinate control of the commitment, early patterning, proliferation, and positional determination of pituitary cell lineages in response to extrinsic and intrinsic signals. The early phases of pituitary development appear to be mediated through the activities of multiple signaling gradients emanating from key organizing centers that give rise to temporally and spatially distinct patterns of transcription factor expression. The induction of these transcriptional mediators in turn acts to positionally organize specific pituitary cell lineages within an apparently uniform field of ectodermal progenitors. Ultimately, pituitary cell types have proven to be both specified and maintained through the combinatorial interactions of a series of cell-type-restricted transcription factors that dictate the cell autonomous programs of differentiation in response to the transient signaling events. PMID- 11283315 TI - Neuropeptides and the integration of motor responses to dehydration. AB - Drinking and eating are critically important motivated behaviors whose expression is usually tightly linked; under conditions of spontaneous intake, disruption of one usually disturbs the other. This characteristic is exemplified by dehydration induced anorexia in which increasing plasma osmolality leads to a centrally generated reduction in food intake, which is then rapidly reversed as water is again made available. This review discusses, at a systems level, how the brain is organized to generate these behaviors and how dehydration affects the expression of neuropeptides in sets of anatomically defined forebrain circuits that contribute to the integration of motor outputs. These findings are then used to consider how altered neuropeptidergic signaling operates within motor drive networks and how these changes may impact the way neuroendocrine, autonomic, and behavioral motor systems respond to this fundamental homeostatic challenge. PMID- 11283316 TI - The developmental biology of brain tumors. AB - Tumors of the central nervous system (CNS) can be devastating because they often affect children, are difficult to treat, and frequently cause mental impairment or death. New insights into the causes and potential treatment of CNS tumors have come from discovering connections with genes that control cell growth, differentiation, and death during normal development. Links between tumorigenesis and normal development are illustrated by three common CNS tumors: retinoblastoma, glioblastoma, and medulloblastoma. For example, the retinoblastoma (Rb) tumor suppressor protein is crucial for control of normal neuronal differentiation and apoptosis. Excessive activity of the epidermal growth factor receptor and loss of the phosphatase PTEN are associated with glioblastoma, and both genes are required for normal growth and development. The membrane protein Patched1 (Ptc1), which controls cell fate in many tissues, regulates cell growth in the cerebellum, and reduced Ptc1 function contributes to medulloblastoma. Just as elucidating the mechanisms that control normal development can lead to the identification of new cancer-related genes and signaling pathways, studies of tumor biology can increase our understanding of normal development. Learning that Ptc1 is a medulloblastoma tumor suppressor led directly to the identification of the Ptc1 ligand, Sonic hedgehog, as a powerful mitogen for cerebellar granule cell precursors. Much remains to be learned about the genetic events that lead to brain tumors and how each event regulates cell cycle progression, apoptosis, and differentiation. The prospects for beneficial work at the boundary between oncology and developmental biology are great. PMID- 11283317 TI - To eat or to sleep? Orexin in the regulation of feeding and wakefulness. AB - Orexin-A and orexin-B are neuropeptides originally identified as endogenous ligands for two orphan G-protein-coupled receptors. Orexin neuropeptides (also known as hypocretins) are produced by a small group of neurons in the lateral hypothalamic and perifornical areas, a region classically implicated in the control of mammalian feeding behavior. Orexin neurons project throughout the central nervous system (CNS) to nuclei known to be important in the control of feeding, sleep-wakefulness, neuroendocrine homeostasis, and autonomic regulation. orexin mRNA expression is upregulated by fasting and insulin-induced hypoglycemia. C-fos expression in orexin neurons, an indicator of neuronal activation, is positively correlated with wakefulness and negatively correlated with rapid eye movement (REM) and non-REM sleep states. Intracerebroventricular administration of orexins has been shown to significantly increase food consumption, wakefulness, and locomotor activity in rodent models. Conversely, an orexin receptor antagonist inhibits food consumption. Targeted disruption of the orexin gene in mice produces a syndrome remarkably similar to human and canine narcolepsy, a sleep disorder characterized by excessive daytime sleepiness, cataplexy, and other pathological manifestations of the intrusion of REM sleep related features into wakefulness. Furthermore, orexin knockout mice are hypophagic compared with weight and age-matched littermates, suggesting a role in modulating energy metabolism. These findings suggest that the orexin neuropeptide system plays a significant role in feeding and sleep-wakefulness regulation, possibly by coordinating the complex behavioral and physiologic responses of these complementary homeostatic functions. PMID- 11283318 TI - Spatial processing in the brain: the activity of hippocampal place cells. AB - The startling discovery by O'Keefe & Dostrovsky (Brain Res. 1971; 34: 171-75) that hippocampal neurons fire selectively in different regions or "place fields" of an environment and the subsequent development of the comprehensive theory by O'Keefe & Nadel (The Hippocampus as a Cognitive Map. Oxford, UK: Clarendon, 1978) that the hippocampus serves as a cognitive map have stimulated a substantial body of literature on the characteristics of hippocampal "place cells" and their relevance for our understanding of the mechanisms by which the brain processes spatial information. This paper reviews the major dimensions of the empirical research on place-cell activity and the development of computational models to explain various characteristics of place fields. PMID- 11283319 TI - The vanilloid receptor: a molecular gateway to the pain pathway. AB - The detection of painful stimuli occurs primarily at the peripheral terminals of specialized sensory neurons called nociceptors. These small-diameter neurons transduce signals of a chemical, mechanical, or thermal nature into action potentials and transmit this information to the central nervous system, ultimately eliciting a perception of pain or discomfort. Little is known about the proteins that detect noxious stimuli, especially those of a physical nature. Here we review recent advances in the molecular characterization of the capsaicin (vanilloid) receptor, an excitatory ion channel expressed by nociceptors, which contributes to the detection and integration of pain-producing chemical and thermal stimuli. The analysis of vanilloid receptor gene knockout mice confirms the involvement of this channel in pain sensation, as well as in hypersensitivity to noxious stimuli following tissue injury. At the same time, these studies demonstrate the existence of redundant mechanisms for the sensation of heat evoked pain. PMID- 11283320 TI - Prion diseases of humans and animals: their causes and molecular basis. AB - Prion diseases are transmissible neurodegenerative conditions that include Creutzfeldt-Jakob disease (CJD) in humans and bovine spongiform encephalopathy (BSE) and scrapie in animals. Prions appear to be composed principally or entirely of abnormal isoforms of a host-encoded glycoprotein, prion protein. Prion propagation involves recruitment of host cellular prion protein, composed primarily of alpha-helical structure, into a disease specific isoform rich in beta-sheet structure. The existence of multiple prion strains has been difficult to explain in terms of a protein-only infections agent, but recent studies suggest that strain specific phenotypes can be encoded by different prion protein conformations and glycosylation patterns. The ability of a protein to encode phenotypic information has important biological implications. The appearance of a novel human prion disease, variant CJD, and the clear experimental evidence that it is caused by exposure to BSE has highlighted the need to understand the molecular basis of prion propagation, pathogenesis, and the barriers limiting intermammalian transmission. It is unclear if a large epidemic of variant CJD will occur in the years ahead. PMID- 11283321 TI - Viktor Hamburger and Rita Levi-Montalcini: the path to the discovery of nerve growth factor. AB - The announcement in October 1986 that the Nobel Prize for physiology or medicine was to be awarded to Rita Levi-Montalcini and Stanley Cohen for the discoveries of NGF and EGF, respectively, caused many to wonder why Viktor Hamburger (in whose laboratory the initial work was done) had not been included in the award. Now that the dust has settled, the time seems opportune to reconsider the antecedent studies on the relation of the developing nervous system to the peripheral structures it innervates. The studies undertaken primarily to investigate this issue culminated in the late 1950s in the discovery that certain tissues produce a nerve growth-promoting factor that is essential for the survival and maintenance of spinal (sensory) ganglion cells and sympathetic neurons. In this review, the many contributions that Viktor and Rita made to this problem, both independently and jointly, are reexamined by considering chronologically each of the relevant research publications together with some of the retrospective memoirs they have published in the years since the discovery of NGF was first reported. PMID- 11283322 TI - Early days of the nerve growth factor proteins. AB - Adult male mouse submaxillary glands served as the preferred starting material for the isolation of the nerve growth factor (NGF) proteins in most of the isolation studies done. Two types of NGF proteins were isolated from extracts of the gland, a high-molecular-weight 7S NGF complex and a low-molecular-weight protein variously called NGF, betaNGF, or 2.5S NGF. The latter, which mediated all known biological functions of NGF, were closely related forms of a basic NGF dimer in which the N and C termini of two monomers (chains) were modified by proteolytic enzymes to different extents with no effect on biological activity. The betaNGF dimer showed a novel protein structure in which the two chains interacted non-covalently over a wide surface. Correspondingly, the betaNGF dimer was found to be unusually stable and the form through which NGFs actions were mediated at physiological concentrations. The betaNGF dimer was one of three subunits in 7S NGF; the other two were the gamma subunit, an arginine esteropeptidase or kallikrein, and the alpha subunit, an inactive kallikrein. Two zinc ions were also present in the complex and contributed greatly to its stability. There was much debate about whether 7S NGF was a specific protein complex of interacting subunits and, if so, what functions it might play in the biology of NGF. Observations of the inhibition of the enzyme activity of the gamma subunit and of the biological activity of betaNGF in 7S NGF were important in determining that 7S NGF was a naturally occurring complex and the sole source of NGF in the gland extract or in saliva. Specific interactions between the active site of the gamma subunit and the C-terminal arginine residues of the NGF chains, confirmed in the three-dimensional structure of 7S NGF, suggested a role for the gamma subunit in pro-NGF processing during the assembly of 7S NGF. In spite of the detailed knowledge of 7S NGF structure, no information on the role of this complex in the neurobiology of NGF has emerged. With the exception of the submaxillary gland of an African rodent, no other source of NGF has been convincingly shown to synthesize the alpha and gamma subunits, and they may well be irrelevant to NGFs actions. PMID- 11283323 TI - Sinusitis. PMID- 11283324 TI - Head injury. PMID- 11283325 TI - Consultation with the specialist: The Pierre Robin sequence: a concise review for the practicing pediatrician. PMID- 11283326 TI - Early detection of developmental hip dysplasia: synopsis of the AAP Clinical Practice Guideline. PMID- 11283327 TI - Index of suspicion. Case 1. Diagnosis: A fetomaternal hemorrhage (FMH). PMID- 11283328 TI - A modern pediatric fable: the case of the missing thumbtack. PMID- 11283330 TI - Everything in its place. Conservation of gene order among distantly related plant species. PMID- 11283331 TI - The new biology. Genomics fosters a "systems approach" and collaborations between academic, government, and industry scientists. PMID- 11283332 TI - Control of shoot cell fate: beyond homeoboxes. PMID- 11283333 TI - Activation of the Arabidopsis B class homeotic genes by APETALA1. AB - Proper development of petals and stamens in Arabidopsis flowers requires the activities of APETALA3 (AP3) and PISTILLATA (PI), whose transcripts can be detected in the petal and stamen primordia. Localized expression of AP3 and PI requires the activities of at least three genes: APETALA1 (AP1), LEAFY (LFY), and UNUSUAL FLORAL ORGANS (UFO). It has been proposed that UFO provides spatial cues and that LFY specifies competence for AP3 and PI expression in the developing flower. To understand the epistatic relationship among AP1, LFY, and UFO in regulating AP3 and PI expression, we generated two versions of AP1 that have strong transcriptional activation potential. Genetic and molecular analyses of transgenic plants expressing these activated AP1 proteins show that the endogenous AP1 protein acts largely as a transcriptional activator in vivo and that AP1 specifies petals by regulating the spatial domains of AP3 and PI expression through UFO. PMID- 11283334 TI - A cell plate-specific callose synthase and its interaction with phragmoplastin. AB - Callose is synthesized on the forming cell plate and several other locations in the plant. We cloned an Arabidopsis cDNA encoding a callose synthase (CalS1) catalytic subunit. The CalS1 gene comprises 42 exons with 41 introns and is transcribed into a 6.0-kb mRNA. The deduced peptide, with an approximate molecular mass of 226 kD, showed sequence homology with the yeast 1,3-beta-glucan synthases and is distinct from plant cellulose synthases. CalS1 contains 16 predicted transmembrane helices with the N-terminal region and a large central loop facing the cytoplasm. CalS1 interacts with two cell plate--associated proteins, phragmoplastin and a novel UDP-glucose transferase that copurifies with the CalS complex. That CalS1 is a cell plate--specific enzyme is demonstrated by the observations that the green fluorescent protein--CalS1 fusion protein was localized at the growing cell plate, that expression of CalS1 in transgenic tobacco cells enhanced callose synthesis on the forming cell plate, and that these cell lines exhibited higher levels of CalS activity. These data also suggest that plant CalS may form a complex with UDP-glucose transferase to facilitate the transfer of substrate for callose synthesis. PMID- 11283335 TI - A novel UDP-glucose transferase is part of the callose synthase complex and interacts with phragmoplastin at the forming cell plate. AB - Using phragmoplastin as a bait, we isolated an Arabidopsis cDNA encoding a novel UDP-glucose transferase (UGT1). This interaction was confirmed by an in vitro protein--protein interaction assay using purified UGT1 and radiolabeled phragmoplastin. Protein gel blot results revealed that UGT1 is associated with the membrane fraction and copurified with the product-entrapped callose synthase complex. These data suggest that UGT1 may act as a subunit of callose synthase that uses UDP-glucose to synthesize callose, a 1,3-beta-glucan. UGT1 also interacted with Rop1, a Rho-like protein, and this interaction occurred only in its GTP-bound configuration, suggesting that the plant callose synthase may be regulated by Rop1 through the interaction with UGT1. The green fluorescent protein--UGT1 fusion protein was located on the forming cell plate during cytokinesis. We propose that UGT1 may transfer UDP-glucose from sucrose synthase to the callose synthase and thus help form a substrate channel for the synthesis of callose at the forming cell plate. PMID- 11283336 TI - Demonstration in yeast of the function of BP-80, a putative plant vacuolar sorting receptor. AB - BP-80, later renamed VSR(PS-1), is a putative receptor involved in sorting proteins such as proaleurain to the lytic vacuole, with its N-terminal domain recognizing the vacuolar sorting determinant. Although all VSR(PS-1) characteristics and in vitro binding properties described so far favored its receptor function, this function remained to be demonstrated. Here, we used green fluorescent protein (GFP) as a reporter in a yeast mutant strain defective for its own vacuolar receptor, Vps10p. By expressing VSR(PS-1) together with GFP fused to the vacuolar sorting determinant of petunia proaleurain, we were able to efficiently redirect the reporter to the yeast vacuole. VSR(PS-1) is ineffective on GFP either alone or when fused with another type of plant vacuolar sorting determinant from a chitinase. The plant VSR(PS-1) therefore interacts specifically with the proaleurain vacuolar sorting determinant in vivo, and this interaction leads to the transport of the reporter protein through the yeast secretory pathway to the vacuole. This finding demonstrates VSR(PS-1) receptor function but also emphasizes the differences in the spectrum of ligands between Vps10p and its plant equivalent. PMID- 11283337 TI - DNA microarray analysis of cyanobacterial gene expression during acclimation to high light. AB - DNA microarrays bearing nearly all of the genes of the unicellular cyanobacterium Synechocystis sp PCC 6803 were used to examine the temporal program of gene expression during acclimation from low to high light intensity. A complete pattern is provided of gene expression during acclimation of a photosynthetic organism to changing light intensity. More than 160 responsive genes were identified and classified into distinct sets. Genes involved in light absorption and photochemical reactions were downregulated within 15 min of exposure to high light intensity, whereas those associated with CO(2) fixation and protection from photoinhibition were upregulated. Changes in the expression of genes involved in replication, transcription, and translation, which were induced to support cellular proliferation, occurred later. Several unidentified open reading frames were induced or repressed. The possible involvement of these genes in the acclimation to high light conditions is discussed. PMID- 11283338 TI - A katanin-like protein regulates normal cell wall biosynthesis and cell elongation. AB - Fibers are one of the mechanical tissues that provide structural support to the plant body. To understand how the normal mechanical strength of fibers is regulated, we isolated an Arabidopsis fragile fiber (fra2) mutant defective in the mechanical strength of interfascicular fibers in the inflorescence stems. Anatomical and chemical analyses showed that the fra2 mutation caused a reduction in fiber cell length and wall thickness, a decrease in cellulose and hemicellulose contents, and an increase in lignin condensation, indicating that the fragile fiber phenotype of fra2 is a result of alterations in fiber cell elongation and cell wall biosynthesis. In addition to the effects on fibers, the fra2 mutation resulted in a remarkable reduction in cell length and an increase in cell width in all organs, which led to a global alteration in plant morphology. The FRA2 gene was shown to encode a protein with high similarity to katanin (hence FRA2 was renamed AtKTN1), a protein shown to be involved in regulating microtubule disassembly by severing microtubules. Consistent with the putative function of AtKTN1 as a microtubule-severing protein, immunolocalization demonstrated that the fra2 mutation caused delays in the disappearance of perinuclear microtubule array and in the establishment of transverse cortical microtubule array in interphase and elongating cells. Together, these results suggest that AtKTN1, a katanin-like protein, is essential not only for normal cell wall biosynthesis and cell elongation in fiber cells but also for cell expansion in all organs. PMID- 11283339 TI - IAA17/AXR3: biochemical insight into an auxin mutant phenotype. AB - The Aux/IAA genes are rapidly and specifically induced by the plant hormone auxin. The proteins encoded by this gene family are short-lived nuclear proteins that are capable of homodimerizing and heterodimerizing. Molecular, biochemical, and genetic data suggest that these proteins are involved in auxin signaling. The pleiotropic morphological phenotype and altered auxin responses of the semidominant axr3-1 mutant of Arabidopsis result from a single amino acid change in the conserved domain II of the Aux/IAA protein IAA17. Here, we show that the biochemical effect of this gain-of-function mutation is to increase the half-life of the iaa17/axr3-1 protein by sevenfold. Intragenic mutations that suppress the iaa17/axr3-1 phenotype have been described. The iaa17/axr3-1R3 revertant contains a second site mutation in domain I and the iaa17/axr3-1R2 revertant contains a second site mutation in domain III. Transient expression assays show that the mutant forms of IAA17/AXR3 retain the ability to accumulate in the nucleus. Using the yeast two hybrid system, we show that the iaa17/axr3-1 mutation does not affect homodimerization. However, the iaa17/axr3-1 revertants counteract the increased levels of iaa17/axr3-1 protein by decreasing the capacity of the mutant protein to homodimerize. Interestingly, heterodimerization of the revertant forms of IAA17/AXR3 with IAA3/SHY2, another Aux/IAA protein, and ARF1 or ARF5/MP proteins is affected only by changes in domain III. Collectively, the results provide biochemical evidence that the revertant mutations in the IAA17/AXR3 gene affect the capacity of the encoded protein to dimerize with itself, other members of the Aux/IAA protein family, and members of the ARF protein family. By extension, these findings may provide insight into the effects of analogous mutations in other members of the Aux/IAA gene family. PMID- 11283340 TI - Auxin transport promotes Arabidopsis lateral root initiation. AB - Lateral root development in Arabidopsis provides a model for the study of hormonal signals that regulate postembryonic organogenesis in higher plants. Lateral roots originate from pairs of pericycle cells, in several cell files positioned opposite the xylem pole, that initiate a series of asymmetric, transverse divisions. The auxin transport inhibitor N-1-naphthylphthalamic acid (NPA) arrests lateral root development by blocking the first transverse division(s). We investigated the basis of NPA action by using a cell-specific reporter to demonstrate that xylem pole pericycle cells retain their identity in the presence of the auxin transport inhibitor. However, NPA causes indoleacetic acid (IAA) to accumulate in the root apex while reducing levels in basal tissues critical for lateral root initiation. This pattern of IAA redistribution is consistent with NPA blocking basipetal IAA movement from the root tip. Characterization of lateral root development in the shoot meristemless1 mutant demonstrates that root basipetal and leaf acropetal auxin transport activities are required during the initiation and emergence phases, respectively, of lateral root development. PMID- 11283341 TI - The TRANSPARENT TESTA12 gene of Arabidopsis encodes a multidrug secondary transporter-like protein required for flavonoid sequestration in vacuoles of the seed coat endothelium. AB - Phenolic compounds that are present in the testa interfere with the physiology of seed dormancy and germination. We isolated a recessive Arabidopsis mutant with pale brown seeds, transparent testa12 (tt12), from a reduced seed dormancy screen. Microscopic analysis of tt12 developing and mature testas revealed a strong reduction of proanthocyanidin deposition in vacuoles of endothelial cells. Double mutants with tt12 and other testa pigmentation mutants were constructed, and their phenotypes confirmed that tt12 was affected at the level of the flavonoid biosynthetic pathway. The TT12 gene was cloned and found to encode a protein with similarity to prokaryotic and eukaryotic secondary transporters with 12 transmembrane segments, belonging to the MATE (multidrug and toxic compound extrusion) family. TT12 is expressed specifically in ovules and developing seeds. In situ hybridization localized its transcript in the endothelium layer, as expected from the effect of the tt12 mutation on testa flavonoid pigmentation. The phenotype of the mutant and the nature of the gene suggest that TT12 may control the vacuolar sequestration of flavonoids in the seed coat endothelium. PMID- 11283342 TI - Identification of two loci in tomato reveals distinct mechanisms for salt tolerance. AB - Salt stress is one of the most serious environmental factors limiting the productivity of crop plants. To understand the molecular basis for salt responses, we used mutagenesis to identify plant genes required for salt tolerance in tomato. As a result, three tomato salt-hypersensitive (tss) mutants were isolated. These mutants defined two loci and were caused by single recessive nuclear mutations. The tss1 mutant is specifically hypersensitive to growth inhibition by Na(+) or Li(+) and is not hypersensitive to general osmotic stress. The tss2 mutant is hypersensitive to growth inhibition by Na(+) or Li(+) but, in contrast to tss1, is also hypersensitive to general osmotic stress. The TSS1 locus is necessary for K(+) nutrition because tss1 mutants are unable to grow on a culture medium containing low concentrations of K(+). Increased Ca(2)+ in the culture medium suppresses the growth defect of tss1 on low K(+). Measurements of membrane potential in apical root cells were made with an intracellular microelectrode to assess the permeability of the membrane to K(+) and Na(+). K(+) dependent membrane potential measurements indicate impaired K(+) uptake in tss1 but not tss2, whereas no differences in Na(+) uptake were found. The TSS2 locus may be a negative regulator of abscisic acid signaling, because tss2 is hypersensitive to growth inhibition by abscisic acid. Our results demonstrate that the TSS1 locus is essential for K(+) nutrition and NaCl tolerance in tomato. Significantly, the isolation of the tss2 mutant demonstrates that abscisic acid signaling is also important for salt and osmotic tolerance in glycophytic plants. PMID- 11283343 TI - Gene expression profiles during the initial phase of salt stress in rice. AB - Transcript regulation in response to high salinity was investigated for salt tolerant rice (var Pokkali) with microarrays including 1728 cDNAs from libraries of salt-stressed roots. NaCl at 150 mM reduced photosynthesis to one tenth of the prestress value within minutes. Hybridizations of RNA to microarray slides probed for changes in transcripts from 15 min to 1 week after salt shock. Beginning 15 min after the shock, Pokkali showed upregulation of transcripts. Approximately 10% of the transcripts in Pokkali were significantly upregulated or downregulated within 1 hr of salt stress. The initial differences between control and stressed plants continued for hours but became less pronounced as the plants adapted over time. The interpretation of an adaptive process was supported by the similar analysis of salinity-sensitive rice (var IR29), in which the immediate response exhibited by Pokkali was delayed and later resulted in downregulation of transcription and death. The upregulated functions observed with Pokkali at different time points during stress adaptation changed over time. Increased protein synthesis and protein turnover were observed at early time points, followed by the induction of known stress-responsive transcripts within hours, and the induction of transcripts for defense-related functions later. After 1 week, the nature of upregulated transcripts (e.g., aquaporins) indicated recovery. PMID- 11283344 TI - Changes in root cap pH are required for the gravity response of the Arabidopsis root. AB - Although the columella cells of the root cap have been identified as the site of gravity perception, the cellular events that mediate gravity signaling remain poorly understood. To determine if cytoplasmic and/or wall pH mediates the initial stages of root gravitropism, we combined a novel cell wall pH sensor (a cellulose binding domain peptide-Oregon green conjugate) and a cytoplasmic pH sensor (plants expressing pH-sensitive green fluorescent protein) to monitor pH dynamics throughout the graviresponding Arabidopsis root. The root cap apoplast acidified from pH 5.5 to 4.5 within 2 min of gravistimulation. Concomitantly, cytoplasmic pH increased in columella cells from 7.2 to 7.6 but was unchanged elsewhere in the root. These changes in cap pH preceded detectable tropic growth or growth-related pH changes in the elongation zone cell wall by 10 min. Altering the gravity-related columella cytoplasmic pH shift with caged protons delayed the gravitropic response. Together, these results suggest that alterations in root cap pH likely are involved in the initial events that mediate root gravity perception or signal transduction. PMID- 11283345 TI - Circadian changes in ribulose-1,5-bisphosphate carboxylase/oxygenase distribution inside individual chloroplasts can account for the rhythm in dinoflagellate carbon fixation. AB - Previous studies of photosynthetic carbon fixation in the marine alga Gonyaulax have shown that the reaction rates in vivo vary threefold between day and night but that the in vitro activity of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), which catalyzes the rate-limiting step in this process, remains constant. Using protein gel blotting, we confirm that Rubisco protein levels are constant over time. We present simultaneous measurements of the rhythms of CO(2) fixation and O(2) evolution and show that the two rhythms are approximately 6 hr out of phase. We further show that the distribution of Rubisco within chloroplasts varies as a function of circadian time and that this rhythm in Rubisco distribution correlates with the CO(2) fixation rhythm. At times of high carbon fixation, Rubisco is found in pyrenoids, regions of the chloroplasts located near the cell center, and is separated from most of the light-harvesting protein PCP (for peridinin-chlorophyll a--protein), which is found in cortical regions of the plastids. We propose that the rhythm in Rubisco distribution is causally related to the rhythm in carbon fixation and suggest that several mechanisms involving enzyme sequestration could account for the increase in the efficiency of carbon fixation. PMID- 11283346 TI - Loss of FLOWERING LOCUS C activity eliminates the late-flowering phenotype of FRIGIDA and autonomous pathway mutations but not responsiveness to vernalization. AB - The MADS domain--containing transcription factor FLOWERING LOCUS C (FLC) acts as an inhibitor of flowering and is a convergence point for several pathways that regulate flowering time in Arabidopsis. In naturally occurring late-flowering ecotypes, the FRIGIDA (FRI) gene acts to increase FLC levels, whereas the autonomous floral promotion pathway and vernalization act to reduce FLC expression. Previous work has shown that the Landsberg erecta allele of FLC, which is not a null allele, is able to partially suppress the late-flowering phenotype of FRIGIDA and mutations in the autonomous pathway. In this study, using a null allele of FLC, we show that the late-flowering phenotype of FRIGIDA and autonomous pathway mutants are eliminated in the absence of FLC activity. In addition, we have found that the downregulation of SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 by FRI and autonomous pathway mutants also is mediated by FLC. Complete loss of FLC function, however, does not eliminate the effect of vernalization. Thus, FRI and the autonomous pathway may act solely to regulate FLC expression, whereas vernalization is able to promote flowering via FLC dependent and FLC-independent mechanisms. PMID- 11283347 TI - The pore of plant K(+) channels is involved in voltage and pH sensing: domain swapping between different K(+) channel alpha-subunits. AB - Plant K(+) uptake channel types differ with respect to their voltage, Ca(2)+, and pH dependence. Here, we constructed recombinant chimeric channels between KST1, a member of the inward-rectifying, acid-activated KAT1 family, and AKT3, a member of the weakly voltage-dependent, proton-blocked AKT2/3 family. The homologous pore regions of AKT3 (amino acids 216 to 287) and KST1 (amino acids 217 to 289) have been exchanged to generate the two chimeric channels AKT3/(p)KST1 and KST1/(p)AKT3. In contrast to AKT3 wild-type channels, AKT3/(p)KST1 revealed a strong inward rectification reminiscent of that of KST1. Correspondingly, the substitution of the KST1 by the AKT3 pore led to less pronounced rectification properties of KST1/(p)AKT3 compared with wild-type KST1. Besides the voltage dependence, the interaction between the chimera and extracellular H(+) and Ca(2)+ resembled the properties of the inserted rather than the respective wild-type pore. Whereas AKT3/(p)KST1 was acid activated and Ca(2)+ insensitive, extracellular protons and Ca(2)+ inhibited KST1/(p)AKT3. The regulation of the chimeric channels by cytoplasmic protons followed the respective wild-type backbone of the chimeric channels, indicating that the intracellular pH sensor is located outside the P domain. We thus conclude that essential elements for external pH and Ca(2)+ regulation and for the rectification of voltage-dependent K(+) uptake channels are located within the channel pore. PMID- 11283348 TI - Functional characterization of a maize purine transporter by expression in Aspergillus nidulans. AB - We have characterized the function of Leaf Permease1 (LPE1), a protein that is necessary for proper chloroplast development in maize, by functional expression in the filamentous fungus Aspergillus nidulans. The choice of this ascomycete was dictated by the similarity of its endogenous purine transporters to LPE1 and by particular genetic and physiological features of purine transport and metabolism in A. nidulans. When Lpe1 was expressed in a purine transport-deficient A. nidulans strain, the capacity for uric acid and xanthine transport was acquired. This capacity was directly dependent on Lpe1 copy number and expression level. Interestingly, overexpression of LPE1 from >10 gene copies resulted in transformants with pleiotropically reduced growth rates on various nitrogen sources and the absolute inability to transport purines. Kinetic analysis established that LPE1 is a high-affinity (K(m) = 30 +/- 2.5 microM), high capacity transporter specific for the oxidized purines xanthine and uric acid. Competition studies showed that high concentrations of ascorbic acid (>30 mM) competitively inhibit LPE1-mediated purine transport. This work defines the biochemical function of LPE1, a plant representative of a large and ubiquitous transporter family. In addition, A. nidulans is introduced as a novel model system for the cloning and/or functional characterization of transporter genes. PMID- 11283349 TI - Functional genomic analysis of the HY2 family of ferredoxin-dependent bilin reductases from oxygenic photosynthetic organisms. AB - Phytobilins are linear tetrapyrrole precursors of the light-harvesting prosthetic groups of the phytochrome photoreceptors of plants and the phycobiliprotein photosynthetic antennae of cyanobacteria, red algae, and cryptomonads. Previous biochemical studies have established that phytobilins are synthesized from heme via the intermediacy of biliverdin IX alpha (BV), which is reduced subsequently by ferredoxin-dependent bilin reductases with different double-bond specificities. By exploiting the sequence of phytochromobilin synthase (HY2) of Arabidopsis, an enzyme that catalyzes the ferredoxin-dependent conversion of BV to the phytochrome chromophore precursor phytochromobilin, genes encoding putative bilin reductases were identified in the genomes of various cyanobacteria, oxyphotobacteria, and plants. Phylogenetic analyses resolved four classes of HY2-related genes, one of which encodes red chlorophyll catabolite reductases, which are bilin reductases involved in chlorophyll catabolism in plants. To test the catalytic activities of these putative enzymes, representative HY2-related genes from each class were amplified by the polymerase chain reaction and expressed in Escherichia coli. Using a coupled apophytochrome assembly assay and HPLC analysis, we examined the ability of the recombinant proteins to catalyze the ferredoxin-dependent reduction of BV to phytobilins. These investigations defined three new classes of bilin reductases with distinct substrate/product specificities that are involved in the biosynthesis of the phycobiliprotein chromophore precursors phycoerythrobilin and phycocyanobilin. Implications of these results are discussed with regard to the pathways of phytobilin biosynthesis and their evolution. PMID- 11283350 TI - Comparative sequence analysis reveals extensive microcolinearity in the lateral suppressor regions of the tomato, Arabidopsis, and Capsella genomes. AB - A 57-kb region of tomato chromosome 7 harboring five different genes was compared with the sequence of the Arabidopsis genome to search for microsynteny between the genomes of these two species. For all five genes, homologous sequences could be identified in a 30-kb region located on Arabidopsis chromosome 1. Only two inversion events distinguish the arrangement of the five genes in tomato from that in Arabidopsis. Inversions were not detected when the arrangement of the five Arabidopsis genes was compared with the arrangement in the orthologous region of Capsella, a plant closely related to Arabidopsis. These results provide evidence for microcolinearity between closely and distantly related dicotyledonous species. The degree of microcolinearity found can be exploited to localize orthologous genes in Arabidopsis and tomato in an unambiguous way. PMID- 11283352 TI - An algorithm for discovery. PMID- 11283351 TI - A comprehensive two-hybrid analysis to explore the yeast protein interactome. AB - Protein-protein interactions play crucial roles in the execution of various biological functions. Accordingly, their comprehensive description would contribute considerably to the functional interpretation of fully sequenced genomes, which are flooded with novel genes of unpredictable functions. We previously developed a system to examine two-hybrid interactions in all possible combinations between the approximately 6,000 proteins of the budding yeast Saccharomyces cerevisiae. Here we have completed the comprehensive analysis using this system to identify 4,549 two-hybrid interactions among 3,278 proteins. Unexpectedly, these data do not largely overlap with those obtained by the other project [Uetz, P., et al. (2000) Nature (London) 403, 623-627] and hence have substantially expanded our knowledge on the protein interaction space or interactome of the yeast. Cumulative connection of these binary interactions generates a single huge network linking the vast majority of the proteins. Bioinformatics-aided selection of biologically relevant interactions highlights various intriguing subnetworks. They include, for instance, the one that had successfully foreseen the involvement of a novel protein in spindle pole body function as well as the one that may uncover a hitherto unidentified multiprotein complex potentially participating in the process of vesicular transport. Our data would thus significantly expand and improve the protein interaction map for the exploration of genome functions that eventually leads to thorough understanding of the cell as a molecular system. PMID- 11283353 TI - Pressure dependence of the superconducting transition temperature of magnesium diboride. AB - We studied the pressure and temperature dependence of the electrical resistivity of the superconducting compound magnesium diboride (MgB(2)). The superconducting transition temperature decreases monotonically with pressure, being parabolic or linear, depending on samples. The rate of decrease under pressure is higher than in conventional superconductors. We discuss our results in terms of the semimetallic character of the electronic band structure of MgB(2). PMID- 11283354 TI - Role of histone H3 lysine 9 methylation in epigenetic control of heterochromatin assembly. AB - The assembly of higher order chromatin structures has been linked to the covalent modifications of histone tails. We provide in vivo evidence that lysine 9 of histone H3 (H3 Lys9) is preferentially methylated by the Clr4 protein at heterochromatin-associated regions in fission yeast. Both the conserved chromo- and SET domains of Clr4 are required for H3 Lys9 methylation in vivo. Localization of Swi6, a homolog of Drosophila HP1, to heterochomatic regions is dependent on H3 Lys9 methylation. Moreover, an H3-specific deacetylase Clr3 and a beta-propeller domain protein Rik1 are required for H3 Lys9 methylation by Clr4 and Swi6 localization. These data define a conserved pathway wherein sequential histone modifications establish a "histone code" essential for the epigenetic inheritance of heterochromatin assembly. PMID- 11283355 TI - Cooperation and competition in the evolution of ATP-producing pathways. AB - Heterotrophic organisms generally face a trade-off between rate and yield of adenosine triphosphate (ATP) production. This trade-off may result in an evolutionary dilemma, because cells with a higher rate but lower yield of ATP production may gain a selective advantage when competing for shared energy resources. Using an analysis of model simulations and biochemical observations, we show that ATP production with a low rate and high yield can be viewed as a form of cooperative resource use and may evolve in spatially structured environments. Furthermore, we argue that the high ATP yield of respiration may have facilitated the evolutionary transition from unicellular to undifferentiated multicellular organisms. PMID- 11283356 TI - 14C-dead living biomass: evidence for microbial assimilation of ancient organic carbon during shale weathering. AB - Prokaryotes have been cultured from a modern weathering profile developed on a approximately 365-million-year-old black shale that use macromolecular shale organic matter as their sole organic carbon source. Using natural-abundance carbon-14 analysis of membrane lipids, we show that 74 to 94% of lipid carbon in these cultures derives from assimilation of carbon-14-free organic carbon from the shale. These results reveal that microorganisms enriched from shale weathering profiles are able to use a macromolecular and putatively refractory pool of ancient organic matter. This activity may facilitate the oxidation of sedimentary organic matter to inorganic carbon when sedimentary rocks are exposed by erosion. Thus, microorganisms may play a more active role in the geochemical carbon cycle than previously recognized, with profound implications for controls on the abundance of oxygen and carbon dioxide in Earth's atmosphere over geologic time. PMID- 11283357 TI - Selective bond dissociation and rearrangement with optimally tailored, strong field laser pulses. AB - We used strong-field laser pulses that were tailored with closed-loop optimal control to govern specified chemical dissociation and reactivity channels in a series of organic molecules. Selective cleavage and rearrangement of chemical bonds having dissociation energies up to approximately 100 kilocalories per mole (about 4 electron volts) are reported for polyatomic molecules, including (CH3)2CO (acetone), CH3COCF3 (trifluoroacetone), and C6H5COCH3 (acetophenone). Control over the formation of CH(3)CO from (CH3)2CO, CF3 (or CH3) from CH3COCF3, and C6H5CH3 (toluene) from C6H5COCH3 was observed with high selectivity. Strong field control appears to have generic applicability for manipulating molecular reactivity because the tailored intense laser fields (about 10(13) watts per square centimeter) can dynamically Stark shift many excited states into resonance, and consequently, the method is not confined by resonant spectral restrictions found in the perturbative (weak-field) regime. PMID- 11283358 TI - Crystal structure of the ribosome at 5.5 A resolution. AB - We describe the crystal structure of the complete Thermus thermophilus 70S ribosome containing bound messenger RNA and transfer RNAs (tRNAs) at 5.5 angstrom resolution. All of the 16S, 23S, and 5S ribosomal RNA (rRNA) chains, the A-, P-, and E-site tRNAs, and most of the ribosomal proteins can be fitted to the electron density map. The core of the interface between the 30S small subunit and the 50S large subunit, where the tRNA substrates are bound, is dominated by RNA, with proteins located mainly at the periphery, consistent with ribosomal function being based on rRNA. In each of the three tRNA binding sites, the ribosome contacts all of the major elements of tRNA, providing an explanation for the conservation of tRNA structure. The tRNAs are closely juxtaposed with the intersubunit bridges, in a way that suggests coupling of the 20 to 50 angstrom movements associated with tRNA translocation with intersubunit movement. PMID- 11283359 TI - Ecology. The advantages of togetherness. AB - What would be the advantage of unicellular organisms becoming multicellular? For organisms that feed on organic food (heterotrophs), the most efficient way to produce energy is to metabolize the food by aerobic respiration, but the fastest way is to metabolize it by fermentation. In their Perspective, Cox and Bonner discuss a mathematical model (Pfeiffer et al.), which shows that when these two kinds of organisms (respirators and fermenters) compete for a limited food source, the respirators manage best when they are grouped in clusters rather than remaining as separate cells. In this way, multicellularity could have originated. PMID- 11283360 TI - Subatomic features in atomic force microscopy images. PMID- 11283362 TI - Observation of Fermi pressure in a gas of trapped atoms. AB - We report the attainment of simultaneous quantum degeneracy in a mixed gas of bosons (lithium-7) and fermions (lithium-6). The Fermi gas has been cooled to a temperature of 0.25 times the Fermi temperature by thermal collisions with the evaporatively cooled bosons. At this temperature, the spatial size of the gas is strongly affected by the Fermi pressure resulting from the Pauli exclusion principle and gives clear experimental evidence for quantum degeneracy. PMID- 11283361 TI - Dynamics of the vocal imitation process: how a zebra finch learns its song. AB - Song imitation in birds provides good material for studying the basic biology of vocal learning. Techniques were developed for inducing the rapid onset of song imitation in young zebra finches and for tracking trajectories of vocal change over a 7-week period until a match to a model song was achieved. Exposure to a model song induced the prompt generation of repeated structured sounds (prototypes) followed by a slow transition from repetitive to serial delivery of syllables. Tracking this transition revealed two phenomena: (i) Imitations of dissimilar sounds can emerge from successive renditions of the same prototype, and (ii) developmental trajectories for some sounds followed paths of increasing acoustic mismatch until an abrupt correction occurred by period doubling. These dynamics are likely to reflect underlying neural and articulatory constraints on the production and imitation of sounds. PMID- 11283363 TI - Spin chirality, Berry phase, and anomalous Hall effect in a frustrated ferromagnet. AB - An electron hopping on non-coplanar spin sites with spin chirality obtains a complex phase factor (Berry phase) in its quantum mechanical amplitude that acts as an internal magnetic field, and is predicted to manifest itself in the Hall effect when it is not cancelled. The present combined work of transport measurement, neutron scattering, and theoretical calculation provides evidence that the gigantic anomalous Hall effect observed in Nd2Mo2O7, a pyrochlore ferromagnet with geometrically frustrated lattice structure, is mostly due to the spin chirality and the associated Berry phase originating from the Mo spin tilting. PMID- 11283364 TI - Alignment of liquid crystals with patterned isotropic surfaces. AB - The molecules of a nematic liquid crystal exposed to an isotropic surface adopt a mean tilt relative to the normal but have no in-plane alignment-that is, they are free to have any azimuthal orientation in the surface plane. Pursuing the theoretical suggestion by Meyer that, in spite of this azimuthal degeneracy, spatially inhomogeneous isotropic surfaces combine with liquid crystal elastic anisotropy to produce alignment, we show that a boundary line between two isotropic regions that differ in mean tilt does indeed align the liquid crystal. The boundaries on a patterned surface of distinct isotropic regions thus act as a system of lines that the molecular orientation locally follows. This enables the development of liquid crystal alignment surfaces based on printing or lithographic patterning. PMID- 11283365 TI - Quantitative measurement of short-range chemical bonding forces. AB - We report direct force measurements of the formation of a chemical bond. The experiments were performed using a low-temperature atomic force microscope, a silicon tip, and a silicon (111) 7x7 surface. The measured site-dependent attractive short-range force, which attains a maximum value of 2.1 nanonewtons, is in good agreement with first-principles calculations of an incipient covalent bond in an analogous model system. The resolution was sufficient to distinguish differences in the interaction potential between inequivalent adatoms, demonstrating the ability of atomic force microscopy to provide quantitative, atomic-scale information on surface chemical reactivity. PMID- 11283366 TI - Influence of carbonic anhydrase activity in terrestrial vegetation on the 18O content of atmospheric CO2. AB - The oxygen-18 (18O) content of atmospheric carbon dioxide (CO2) is an important indicator of CO2 uptake on land. It has generally been assumed that during photosynthesis, oxygen in CO2 reaches isotopic equilibrium with oxygen in 18O enriched water in leaves. We show, however, large differences in the activity of carbonic anhydrase (which catalyzes CO2 hydration and 18O exchange in leaves) among major plant groups that cause variations in the extent of 18O equilibrium (theta(eq)). A clear distinction in theta(eq) between C3 trees and shrubs, and C4 grasses makes atmospheric C18OO a potentially sensitive indicator to changes in C3 and C4 productivity. We estimate a global mean theta(eq) value of approximately 0.8, which reasonably reconciles inconsistencies between 18O budgets of atmospheric O2 (Dole effect) and CO2. PMID- 11283367 TI - Ancient geodynamics and global-scale hydrology on Mars. AB - Loading of the lithosphere of Mars by the Tharsis rise explains much of the global shape and long-wavelength gravity field of the planet, including a ring of negative gravity anomalies and a topographic trough around Tharsis, as well as gravity anomaly and topographic highs centered in Arabia Terra and extending northward toward Utopia. The Tharsis-induced trough and antipodal high were largely in place by the end of the Noachian Epoch and exerted control on the location and orientation of valley networks. The release of carbon dioxide and water accompanying the emplacement of approximately 3 x 10(8) cubic kilometers of Tharsis magmas may have sustained a warmer climate than at present, enabling the formation of ancient valley networks and fluvial landscape denudation in and adjacent to the large-scale trough. PMID- 11283368 TI - Role of the stratospheric polar freezing belt in denitrification. AB - Homogeneous freezing of nitric acid hydrate particles can produce a polar freezing belt in either hemisphere that can cause denitrification. Computed denitrification profiles for one Antarctic and two Arctic cold winters are presented. The vertical range over which denitrification occurs is normally quite deep in the Antarctic but limited in the Arctic. A 4 kelvin decrease in the temperature of the Arctic stratosphere due to anthropogenic and/or natural effects can trigger the occurrence of widespread severe denitrification. Ozone loss is amplified in a denitrified stratosphere, so the effects of falling temperatures in promoting denitrification must be considered in assessment studies of ozone recovery trends. PMID- 11283369 TI - Biospheric primary production during an ENSO transition. AB - The Sea-viewing Wide Field-of-view Sensor (SeaWiFS) provides global monthly measurements of both oceanic phytoplankton chlorophyll biomass and light harvesting by land plants. These measurements allowed the comparison of simultaneous ocean and land net primary production (NPP) responses to a major El Nino to La Nina transition. Between September 1997 and August 2000, biospheric NPP varied by 6 petagrams of carbon per year (from 111 to 117 petagrams of carbon per year). Increases in ocean NPP were pronounced in tropical regions where El Nino-Southern Oscillation (ENSO) impacts on upwelling and nutrient availability were greatest. Globally, land NPP did not exhibit a clear ENSO response, although regional changes were substantial. PMID- 11283370 TI - Energetic and fitness costs of mismatching resource supply and demand in seasonally breeding birds. AB - By advancing spring leaf flush and ensuing food availability, climatic warming results in a mismatch between the timing of peak food supply and nestling demand, shifting the optimal time for reproduction in birds. Two populations of blue tits (Parus caeruleus) that breed at different dates in similar, but spatially distinct, habitat types in Corsica and southern France provide a unique opportunity to quantify the energetic and fitness consequences when breeding is mismatched with local productivity. As food supply and demand become progressively mismatched, the increased cost of rearing young pushes the metabolic effort of adults beyond their apparent sustainable limit, drastically reducing the persistence of adults in the breeding population. We provide evidence that the economics of parental foraging and limits to sustainable metabolic effort are key selective forces underlying synchronized seasonal breeding and long-term shifts in breeding date in response to climatic change. PMID- 11283371 TI - Sterility of Drosophila with mutations in the Bloom syndrome gene- complementation by Ku70. AB - The Drosophila Dmblm locus is a homolog of the human Bloom syndrome gene, which encodes a helicase of the RECQ family. We show that Dmblm is identical to mus309, a locus originally identified in a mutagen-sensitivity screen. One mus309 allele, which carries a stop codon between two of the helicase motifs, causes partial male sterility and complete female sterility. Mutant males produce an excess of XY sperm and nullo sperm, consistent with a high frequency of nondisjunction and/or chromosome loss. These phenotypes of mus309 suggest that Dmblm functions in DNA double-strand break repair. The mutant Dmblm phenotypes were partially rescued by an extra copy of the DNA repair gene Ku70, indicating that the two genes functionally interact in vivo. PMID- 11283372 TI - Extreme diversity, conservation, and convergence of spider silk fibroin sequences. AB - Spiders (Araneae) spin high-performance silks from liquid fibroin proteins. Fibroin sequences from basal spider lineages reveal mosaics of amino acid motifs that differ radically from previously described spider silk sequences. The silk fibers of Araneae are constructed from many protein designs. Yet, the repetitive sequences of fibroins from orb-weaving spiders have been maintained, presumably by stabilizing selection, over 125 million years of evolutionary history. The retention of these conserved motifs since the Mesozoic and their convergent evolution in other structural superproteins imply that these sequences are central to understanding the exceptional mechanical properties of orb weaver silks. PMID- 11283373 TI - Costs and benefits of high mutation rates: adaptive evolution of bacteria in the mouse gut. AB - We have shown that bacterial mutation rates change during the experimental colonization of the mouse gut. A high mutation rate was initially beneficial because it allowed faster adaptation, but this benefit disappeared once adaptation was achieved. Mutator bacteria accumulated mutations that, although neutral in the mouse gut, are often deleterious in secondary environments. Consistently, the competitiveness of mutator bacteria is reduced during transmission to and re-colonization of similar hosts. The short-term advantages and long-term disadvantages of mutator bacteria could account for their frequency in nature. PMID- 11283374 TI - Virus-assisted mapping of neural inputs to a feeding center in the hypothalamus. AB - We report the development of a pseudorabies virus that can be used for retrograde tracing from selected neurons. This virus encodes a green fluorescent protein marker and replicates only in neurons that express the Cre recombinase and in neurons in synaptic contact with the originally infected cells. The virus was injected into the arcuate nucleus of mice that express Cre only in those neurons that express neuropeptide Y or the leptin receptor. Sectioning of the brains revealed that these neurons receive inputs from neurons in other regions of the hypothalamus, as well as the amygdala, cortex, and other brain regions. These data suggest that higher cortical centers modulate leptin signaling in the hypothalamus. This method of neural tracing may prove useful in studies of other complex neural circuits. PMID- 11283375 TI - Continuous fatty acid oxidation and reduced fat storage in mice lacking acetyl CoA carboxylase 2. AB - Malonyl-coenzyme A (malonyl-CoA), generated by acetyl-CoA carboxylases ACC1 and ACC2, is a key metabolite in the regulation of energy homeostasis. Here, we show that Acc2-/- mutant mice have a normal life span, a higher fatty acid oxidation rate, and lower amounts of fat. In comparison to the wild type, Acc2-deficient mice had 10- and 30-fold lower levels of malonyl-CoA in heart and muscle, respectively. The fatty acid oxidation rate in the soleus muscle of the Acc2-/- mice was 30% higher than that of wild-type mice and was not affected by addition of insulin; however, addition of insulin to the wild-type muscle reduced fatty acid oxidation by 45%. The mutant mice accumulated 50% less fat in their adipose tissue than did wild-type mice. These results raise the possibility that pharmacological manipulation of ACC2 may lead to loss of body fat in the context of normal caloric intake. PMID- 11283376 TI - Conservation conflicts across Africa. AB - There is increasing evidence that areas of outstanding conservation importance may coincide with dense human settlement or impact. We tested the generality of these findings using 1 degree-resolution data for sub-Saharan Africa. We find that human population density is positively correlated with species richness of birds, mammals, snakes, and amphibians. This association holds for widespread, narrowly endemic, and threatened species and looks set to persist in the face of foreseeable population growth. Our results contradict earlier expectations of low conflict based on the idea that species richness decreases and human impact increases with primary productivity. We find that across Africa, both variables instead exhibit unimodal relationships with productivity. Modifying priority setting to take account of human density shows that, at this scale, conflicts between conservation and development are not easily avoided, because many densely inhabited grid cells contain species found nowhere else. PMID- 11283377 TI - G-protein beta3 subunit and alpha-adducin polymorphisms and risk of subclinical and clinical stroke. AB - BACKGROUND AND PURPOSE: Essential hypertension is a significant risk factor for stroke. Genes contributing to interindividual variation in blood pressure levels and essential hypertension status may play a role in the etiology of stroke either through their effects on blood pressure levels or through separate pathways. For this reason, we sought to examine the association between the alpha adducin (ADD1) G/W460 and G-protein beta3 subunit (GNbeta3) 825C/T polymorphisms and subclinical and clinical stroke in the Atherosclerosis Risk in Communities (ARIC) Study. METHODS: Subclinical stroke was determined by cerebral MRI. Subclinical cerebral infarct cases (n=202) were compared with a stratified random sample (MRI-CRS) identified from individuals participating in the MRI examination (n=211). Incidence of clinical ischemic stroke was determined by following the ARIC cohort for an average of 7.2 years for potential cerebrovascular events; 231 validated clinical ischemic strokes were identified. A stratified random sample of the ARIC cohort (CRS) (n=984) was used as the comparison group for the clinical cases. RESULTS: The frequency of the ADD1 W460 allele was determined for the subclinical cases (0.12), MRI-CRS (0.16), clinical cases (0.14), and CRS (0.17). The frequency of the GNbeta3 825T allele was determined in whites and blacks, respectively, for the subclinical cases (0.26, 0.73), MRI-CRS (0.31, 0.75), clinical cases (0.36, 0.72), and CRS (0.30, 0.72). The ADD1 W460 and GNbeta3 825T alleles were not significantly associated with subclinical stroke. The ADD1 W460 allele was also not a significant predictor of clinical stroke. The GNbeta3 825T allele was significantly associated with clinical stroke in whites after adjustment for age and sex (hazard rate ratio, 1.45; 95% CI, 1.05 to 2.00) and after further adjustment for multiple stroke risk factors (hazard rate ratio, 1.68; 95% CI, 1.18 to 2.41). The GNbeta3 825T allele was not significantly associated with clinical stroke in blacks for either adjustment model. CONCLUSIONS: The GNbeta3 gene 825C/T polymorphism is significantly associated with incident clinical ischemic stroke in a white middle-aged American population, but not in blacks. This association does not appear to be mediated by established stroke risk factors, specifically blood pressure levels or hypertension status. PMID- 11283378 TI - Carotid plaque and intima-media thickness assessed by b-mode ultrasonography in subjects ranging from young adults to centenarians. AB - BACKGROUND AND PURPOSE-To investigate relationships among plaque formation, increasing intima-media thickness, and age, we examined ultrasonographically carotid arteries of subjects who had no major atherosclerotic risk factors and who ranged in age from young adults to centenarians. METHODS: We studied 319 healthy subjects (154 men, 165 women; age range, 21 to 105 years) with no history of hypertension, diabetes mellitus, or atherosclerotic disease. Mean intima-media wall thickness (IMT) of common carotid arteries at plaque-free sites and prevalence of plaques were evaluated by B-mode ultrasound. RESULTS: Mean common carotid IMT increased in a linear manner with age for all decades of life, including centenarians [IMT=(0.009xAge)+0.116] (r=0.83). In centenarians (n=30), intima-media complexes were diffusely thickened (mean IMT, 1.01 mm). Plaque prevalence increased up to the tenth decade of life (83.3%, n=30) but decreased in centenarians (60.0%). IMT and plaque prevalence were closely associated in the seventh and eighth decades of life but not at older ages. CONCLUSIONS: The present study indicates that increased IMT is a physiological effect of aging that corresponds to diffuse intimal thickening, especially in very elderly persons, and that IMT is distinct from pathological plaque formation. PMID- 11283379 TI - Comparison of carotid arterial resistive indices with intima-media thickness as sonographic markers of atherosclerosis. AB - Background and Purpose-The intima-media thickness (IMT) of the carotid artery is a (morphological) sonographic parameter that depends on the degree of atherosclerosis. In the renal arteries, the value of the (hemodynamic) resistive index (RI) is correlated with the severity of atherosclerosis. In contrast to the well-known IMT, no study has yet applied the carotid RI to estimate generalized atherosclerosis. METHODS: -The SMART atherosclerosis risk score was determined in 157 patients (94 men and 63 women; mean age 63 [range 19 to 80] years) with at least 1 vascular risk factor or a known vascular disease. Duplex sonography of the common carotid (CCA) and internal carotid artery (ICA) was then performed, with determination of IMT and RI. RESULTS: -The mean risk score of all patients was 8.8+/-3.5 (range 1 to 17), the mean IMT value in the CCA was 0.727+/-0.161 mm, the mean RI in CCA was 0.79+/-0.066, and the mean RI in ICA was 0.661+/ 0.082. Highly significant correlations were found between the score and IMT CCA and the score and RI ICA (r=0.62, P:<0.0001 and r=0.55, P:<0.0001). The score-RI CCA correlation was much less marked (r=0.354, P:<0.0001). The intraobserver and interobserver agreement was less for IMT than for RI CCA and ICA. The areas under the curve of the receiver operating curves to distinguish between low-risk and high-risk patients resulted in values of 0.86, 0.81, and 0.69 for IMT, RI ICA, and RI CCA, respectively. CONCLUSIONS: -Although RI reflects the atherosclerotic process in an indirect manner, the correlation between the RI ICA and the SMART atherosclerosis score as well as the ability to distinguish between low- and high risk patients are comparable to those of the well-known IMT. PMID- 11283380 TI - Elevated white blood cell count and carotid plaque thickness : the northern manhattan stroke study. AB - BACKGROUND AND PURPOSE: Elevated leukocyte count has been associated with cardiovascular and cerebrovascular disease in several epidemiological studies. We sought to determine whether white blood cell count (WBC) is associated with carotid plaque thickness in a stroke-free, multiethnic cohort. METHODS: For this cross-sectional analysis, WBC was measured in stroke-free community subjects undergoing carotid duplex Doppler ultrasound. Maximal internal carotid plaque thickness (MICPT) was measured for each subject. Demographic and potential medical confounding factors were analyzed with linear and logistic regression to calculate the effect of quartile of WBC on MICPT. Odds ratios (ORs) and 95% confidence intervals (CIs) for the effect of quartile of WBC on MICPT >/=75th percentile were calculated. All analyses were stratified by race-ethnicity. RESULTS: The mean age of the 1422 subjects was 68.6+/-10.2 years; 40.0% were men; 24.4% were white, 46.9% Hispanic, and 26.7% black. Among Hispanics, compared with the lowest quartile of WBC, those in the highest quartile had significantly increased MICPT (mean difference=0.30 mm, P:=0.0086) after adjustment for age, sex, and other atherosclerotic risk factors. There was no significant increase for blacks or whites. The OR for MICPT >/=75th percentile (1.9 mm) was significantly increased for Hispanics (OR, 2.8; 95% CI, 1.4 to 5.6), marginally elevated for black non-Hispanics (OR, 1.6; 95% CI, 0.8 to 3.2), and not increased for white non-Hispanics (OR, 0.5; 95% CI, 0.2 to 1.1). CONCLUSIONS: Relative elevation in WBC is associated with carotid atherosclerosis, but this relationship differs by race-ethnicity. The association is strongest in Hispanics, intermediate in black non-Hispanics, and not present in white non Hispanics in this population. Chronic subclinical infection or inflammation may account for this association. PMID- 11283382 TI - Presence of Chlamydia pneumoniae in human symptomatic and asymptomatic carotid atherosclerotic plaque. AB - BACKGROUND: Chlamydia pneumoniae has been identified in atherosclerotic plaques of patients with cerebrovascular and cardiovascular disease. However, the direct causative effect of C pneumoniae infection in the activation of atherosclerotic plaque to a prothrombotic state remains to be established. The aim of the present study is to examine the correlation between intraplaque presence of chlamydiae and symptomatic carotid disease in humans. METHODS: Plaques from 37 symptomatic and 57 asymptomatic consenting patients undergoing carotid endarterectomy were snap-frozen, and the tissue was prepared for polymerase chain reaction analysis for Chlamydia pneumoniae per Institutional Review Board-approved protocol. Blood was drawn from each patient at the time of surgery for serological analysis. RESULTS: The overall rate of plaques positive for C pneumoniae was 14.82%, with 5 of 37 (13.5%) plaques from symptomatic patients and 9 of 57 (15.8%) from asymptomatic patients, which revealed a definitive presence of the organism. No association existed between C pneumoniae presence and symptomatic disease (P:=1.0). Also, no association existed between presence of C pneumoniae and severity of stenosis. Finally, seropositivity for anti-chlamydial IgG, IgA, and IgM anti-chlamydial antibodies did not correlate with identification of C pneumoniae in the plaques. However, high-serum anti-chlamydial IgA levels (>/=1:128) were associated with occurrence of symptomatic disease (P=0.03; odds ratio, 2.86; 95% CI, 1.12 to 7.28). CONCLUSIONS: Presence of C pneumoniae as a single factor does not appear to be sufficient to explain the occurrence of cerebrovascular symptoms. Low sensitivity of seropositivity for IgG, IgA, or IgM associated with PCR-identified C pneumoniae presence in the plaque makes it unlikely to be valuable as the single determining factor for actively infected plaque. Association of high-level anti-chlamydial IgA with symptomatic disease suggests that chronic or acute chlamydial infection anywhere in the body could play a role in atherosclerotic plaque activation and be used as a marker to target populations in future stroke prevention trials. PMID- 11283381 TI - Expression of tissue factor in high-grade carotid artery stenosis: association with plaque destabilization. AB - BACKGROUND AND PURPOSE: The procoagulant protein tissue factor (TF) has been implicated in thromboembolic complications associated with advanced atherosclerosis. In this study, we investigated whether TF expression in high grade stenoses of the internal carotid artery (ICA) is associated with clinical features of plaque destabilization and addressed the relationship between TF expression and plaque inflammation. METHODS: In 36 consecutive patients undergoing surgery for high-grade ICA stenosis, clinical evidence of plaque instability was provided by the recent occurrence of ischemic symptoms attributable to the stenosis and the detection of cerebral microembolism by means of transcranial Doppler ultrasound monitoring of the ipsilateral middle cerebral artery. Endarterectomy specimens were stained immunocytochemically for TF expression as well as macrophage (CD68) and T cell (CD3) infiltration. RESULTS: Morphologically, TF immunoreactivity was codistributed with plaque inflammation and predominantly localized to CD68+ macrophages. Accordingly, statistical analysis revealed a significant association of TF expression with plaque infiltration by macrophages (P<0.0001) and T cells (P=0.013). Plaques extensively stained for TF (median of TF+ total section area >40% in semiquantitative assessment) were more frequent in symptomatic (12/27) than in asymptomatic patients (1/9). Conversely, plaques exhibiting little TF expression (median of TF+ section area <20%) were more frequent in asymptomatic (3/9) than in symptomatic (1/27) patients (P=0.016). Likewise, we found a highly significant association of TF expression with the occurrence of cerebral microembolism (P=0.008). CONCLUSIONS: Induction of TF at sites of plaque inflammation may play an important role in the destabilization of high-grade ICA stenosis. PMID- 11283383 TI - Community use of intravenous tissue plasminogen activator for acute stroke: results of the brain matters stroke management survey. AB - BACKGROUND AND PURPOSE: Little is known of neurologists' viewpoints regarding intravenous tPA use or institutional readiness to evaluate potential thrombolytic candidates. METHODS: Surveys were distributed at the Brain Matters Stroke Management Workshops held in 16 cities in the United States. RESULTS: Intravenous tPA was administered by 46.9% of responding neurologists. Almost 30% (29.9%) of surveyed neurologists were "very convinced" of its efficacy, whereas 61.6% were "very concerned" about the risk of intracranial hemorrhage. Only half of the respondents believed their institutions could meet all NINDS-recommended stroke evaluation time targets. CONCLUSIONS: Neurologists' enthusiasm for the efficacy of intravenous tPA is tempered by their concern about intracranial hemorrhage. Institutional readiness for evaluating acute stroke patients is not optimized. PMID- 11283384 TI - Acute stroke management in the local general hospital. AB - BACKGROUND AND PURPOSE: The majority of stroke patients are treated in local general hospitals. Despite this fact, little is known about stroke care in these institutions. We sought to investigate the status quo of acute stroke management in nonspecialized facilities with limited equipment and resources. METHODS: Four general hospitals located in smaller cities of a rural area in Germany participated in this study. The 4 hospitals were similar in structure and technical equipment; none had a CT scanner in-house. We reviewed the medical records of every stroke patient hospitalized in 1 of the 4 hospitals within a period of 8 weeks within 1 year. RESULTS: We collected data of a total of 95 patients at all 4 hospitals. The frequency of diagnostic tests was low: at least 1 CT scan was obtained in only 36.8% of all cases, whereas diagnostic methods available in-house were used more frequently, such as Doppler ultrasound (49.0%), echocardiography (42.3%), and 24-hour ECG registration (48.4%). Each hospital had a different therapeutic approach. Main therapeutic options were the use of pentoxyfilline (0% to 90.5%), osmodiuretics (0% to 90%), piracetam (0% to 93.3%), and hydroxyethylstarch (4.8% to 30%). Medication for long-term secondary prevention was given to 69.8% of all patients. CONCLUSIONS: This study provides one of the few data samples reflecting stroke care in smaller general hospitals. The findings demonstrate a partially suboptimal level of care in these institutions. To achieve future improvements, extended human and technical resources as well as research for stroke care should not be restricted to academic stroke centers. PMID- 11283386 TI - Cerebrovascular manifestations in 321 cases of hereditary hemorrhagic telangiectasia. AB - BACKGROUND AND PURPOSE: Patients with hereditary hemorrhagic telangiectasia (HHT) are at risk for developing cerebral vascular malformations and pulmonary arteriovenous fistulae. We assessed the risk of neurological dysfunction from these malformations and fistulae. METHODS: Three hundred twenty-one consecutive patients with HHT seen at a single institution over a 20-year period were studied. Any evidence of prior neurological symptoms or presence of an intracranial vascular malformation was recorded. All cases of possible cerebral arteriovenous malformation were confirmed by conventional arteriography. RESULTS: Twelve patients (3.7%) had a history of cerebral vascular malformations. Ten patients had arteriovenous malformations, 1 had a dural arteriovenous fistula, and 1 had a cavernous malformation. Seven patients (2.1%) presented with intracranial hemorrhage, 2 presented with seizures alone, and 3 were discovered incidentally. The average age at the time of symptomatic intracranial hemorrhage was 25.4 years. All patients with a history of intracranial hemorrhage were classified as Rankin grade I or II at a mean follow-up interval of 6.0 years. A history of cerebral infarction or transient ischemic attack was found in 29.6% of patients with HHT and a pulmonary arteriovenous fistula. CONCLUSIONS: The risk of intracranial hemorrhage is low among people with HHT. Furthermore, a majority of these patients have a good functional outcome after hemorrhage. The data do not suggest a compelling indication for routine screening of patients with HHT for asymptomatic cerebral vascular malformations. By comparison, pulmonary arteriovenous fistulae are a much more frequent cause of neurological symptoms in this population. PMID- 11283385 TI - Should mild or moderate stroke patients be admitted to an intensive care unit? AB - BACKGROUND AND PURPOSE: Inhospital placement of patients with mild (National Institutes of Health Stroke Scale [NIHSS] score <8) or moderate (NIHSS 8 through 16) acute strokes is variable. We assessed the outcome of such patients based on intensive care unit (ICU) versus general ward placement. METHODS: We reviewed 138 consecutive patients admitted within 24 hours of stroke onset to 2 physically adjacent hospitals with different admitting practices. Outcome measures included complication rates, discharge Rankin scale score, hospital discharge placement, costs, and length of stay (LOS). RESULTS: Hospital A, a 626-bed university affiliated hospital, admitted 43% of mild and moderate strokes (MMS) to an ICU (26% of mild, 74% of moderate), whereas hospital B, a 618-bed community facility, admitted 18% of MMS to an ICU (3% of mild, 45% of moderate; P<0.004). There were no significant differences in outcomes between the 2 hospitals. Analysis of only patients admitted to hospital A, and of all patients, demonstrated that mild stroke patients admitted to the general ward had fewer complications and more favorable discharge Rankin scale scores than similar patients admitted to an ICU. There was no statistically significant difference in LOS, but total room costs for a patient admitted first to the ICU averaged $15 270 versus $3638 for admission directly to the ward. CONCLUSIONS: While limited by the retrospective nature of our study, routinely admitting acute MMS patients to an ICU provides no cost or outcomes benefits. PMID- 11283387 TI - Hemostatic activation in spontaneous intracerebral hemorrhage. AB - BACKGROUND AND PURPOSE: There is no in-depth information available on the changes in hemostatic systems in patients in the acute phase of spontaneous intracerebral hemorrhage (ICH). This study was conducted to assess the relationships between the changes in hemostatic systems and clinical parameters in patients in acute phase ICH. METHODS: The records of 358 patients admitted within 6 hours of onset of ICH were reviewed to examine the relationships between changes in hemostatic systems and computed tomographic findings and clinical parameters. RESULTS: -The white blood cell counts and the levels of thrombin-antithrombin complex, plasmin antiplasmin complex, and D-dimer in patients with intraventricular extension (IVE) or subarachnoid hemorrhage (SAH) were significantly (P<0.05) higher than those in patients without IVE or SAH. Most of the hemostatic system parameters in patients without IVE or SAH showed no significant differences compared with normal subjects. Multiple linear regression analysis revealed that the levels of thrombin-antithrombin complex significantly increased with an increase in the amount of SAH (P<0.001) and IVE (P<0.001). The levels of thrombin-antithrombin complex were not significantly associated with the volume of intraparenchymal hematoma. The level of the complex, however, was significantly (P<0.001) and independently associated with the presence of IVE or SAH (multiple logistic regression analysis). CONCLUSIONS: The systemic activation of hemostatic systems in ICH patients seems to take place only when blood reaches the subarachnoid space. The intraparenchymal hematoma itself seems unlikely to activate hemostatic systems in peripheral blood, although the hematoma is expected to cause local activation of hemostatic systems. PMID- 11283388 TI - The ICH score: a simple, reliable grading scale for intracerebral hemorrhage. AB - BACKGROUND AND PURPOSE: Intracerebral hemorrhage (ICH) constitutes 10% to 15% of all strokes and remains without a treatment of proven benefit. Despite several existing outcome prediction models for ICH, there is no standard clinical grading scale for ICH analogous to those for traumatic brain injury, subarachnoid hemorrhage, or ischemic stroke. METHODS: Records of all patients with acute ICH presenting to the University of California, San Francisco during 1997-1998 were reviewed. Independent predictors of 30-day mortality were identified by logistic regression. A risk stratification scale (the ICH Score) was developed with weighting of independent predictors based on strength of association. RESULTS: Factors independently associated with 30-day mortality were Glasgow Coma Scale score (P<0.001), age >/=80 years (P=0.001), infratentorial origin of ICH (P=0.03), ICH volume (P=0.047), and presence of intraventricular hemorrhage (P=0.052). The ICH Score was the sum of individual points assigned as follows: GCS score 3 to 4 (=2 points), 5 to 12 (=1), 13 to 15 (=0); age >/=80 years yes (=1), no (=0); infratentorial origin yes (=1), no (=0); ICH volume >/=30 cm(3) (=1), <30 cm(3) (=0); and intraventricular hemorrhage yes (=1), no (=0). All 26 patients with an ICH Score of 0 survived, and all 6 patients with an ICH Score of 5 died. Thirty-day mortality increased steadily with ICH Score (P<0.005). CONCLUSIONS: The ICH Score is a simple clinical grading scale that allows risk stratification on presentation with ICH. The use of a scale such as the ICH Score could improve standardization of clinical treatment protocols and clinical research studies in ICH. PMID- 11283389 TI - Plasma vitamin C levels are decreased and correlated with brain damage in patients with intracranial hemorrhage or head trauma. AB - BACKGROUND AND PURPOSE: Free radical hyperproduction may play an important role in brain hemorrhage and ischemia/reperfusion injury. The aims of this study were to assess whether antioxidant depletion occurs after intracranial hemorrhage (ICH) and head trauma (HT) and to evaluate the relation between the diameter of the brain lesion, the degree of the neurological impairment, and any observed antioxidant changes. METHODS: We measured plasma levels of vitamin C (ascorbic acid, AA), uric acid (UA), vitamin E (alpha-tocopherol), and ubiquinol-10 in 13 patients with ICH and 15 patients with HT on the day of the brain injury and subsequently every other day up to 1 week. Patients were compared with 40 healthy control subjects. RESULTS: ICH and HT patients had significantly lower plasma levels of AA compared with healthy subjects, in contrast to plasma levels of UA, alpha-tocopherol, and ubiquinol-10. AA levels were significantly inversely correlated with the severity of the neurological impairment as assessed by the Glasgow Coma Scale and the National Institutes of Health Stroke Scale. AA levels were also significantly inversely correlated with the major diameter of the lesion. In addition, mean plasma AA levels were lower in jugular compared with peripheral blood samples obtained from 5 patients. CONCLUSIONS: These findings suggest that a condition of oxidative stress occurs in patients with head trauma and hemorrhagic stroke of recent onset. The consequences of early vitamin C depletion on brain injury as well as the effects of vitamin C supplementation in ICH and HT patients remain to be addressed in further studies. PMID- 11283390 TI - Prospective study of depressive symptoms and risk of stroke among japanese. AB - BACKGROUND AND PURPOSE: We sought to examine the relationship between depressive symptoms and the incidence of stroke among Japanese men and women. METHODS: A 10.3-year prospective study on the relationship between depressive symptoms and the incidence of stroke was conducted with 901 men and women aged 40 to 78 years in a rural Japanese community. Depressive symptoms were measured at baseline with the use of the Zung Self-Rating Depression Scale (SDS). The incidence of stroke was ascertained under systematic surveillance. RESULTS: During the 10-year follow up, 69 strokes (39 ischemic strokes, 10 intracerebral hemorrhages, 10 subarachnoid hemorrhages, and 10 unclassified strokes) occurred. Age- and sex adjusted prevalence of mild depression (SDS scores >/=40) at baseline was 25% among subjects with incident stroke and 12% among subjects without stroke (P<0.01). Persons with SDS scores in the high tertile had twice the age- and sex adjusted relative risk of total stroke as those with scores in the low tertile. The excess risk was confined to ischemic stroke. After we adjusted for body mass index, systolic blood pressure level, serum total cholesterol level, cigarette smoking, current treatment with antihypertensive medication, and history of diabetes mellitus, these relative risks remained statistically significant for total stroke (1.9; 95% CI, 1.1 to 3.5) and ischemic stroke (2.7; 95% CI, 1.2 to 6.0). CONCLUSIONS: Depressive symptoms predict the risk of stroke, specifically ischemic stroke among Japanese. PMID- 11283391 TI - Entry criteria and baseline characteristics predict outcome in acute stroke trials. AB - Background and Purpose-We sought to study the range of entry criteria and baseline characteristics in acute stroke trials and to understand their effects on patient outcomes. METHODS: -Randomized, placebo-controlled therapeutic trials in patients with acute ischemic stroke were identified. Entry criteria, baseline clinical characteristics, and outcome were extracted for the placebo group of each trial. The relationship between key variables was then determined. RESULTS: Across 90 placebo groups identified, there was great variation in entry criteria and outcome measures. This was associated with divergent outcomes; for example, in some studies most placebo group patients died, while in other studies nearly all had no disability. Entry criteria were significantly correlated with outcome; for example, higher age cutoff for study entry correlated with 3-month mortality. Entry criteria also predicted baseline clinical characteristics; for example, wider time window for study entry correlated directly with time to treatment and inversely with stroke severity (initial National Institutes of Health Stroke Scale score). Baseline characteristics predicted outcome. Greater stroke severity predicted higher 3-month mortality rate; despite this, successful thrombolytic trials have enrolled more severe strokes than most trials. The mean age of enrollees also predicted 3-month mortality and was inversely related to percentage of patients with 3-month Barthel Index score >/=95. The strongest predictors of 3-month mortality were obtained with multivariate models. CONCLUSIONS: -Acute stroke studies vary widely in entry criteria and outcome measures. Across multiple studies, differences in entry criteria, and the baseline clinical characteristics they predict, influence patient outcomes along a continuum. In some studies, enrolling a specific subset of patients may have improved the chances of identifying a treatment-related effect, while in others, such chances may have been reduced. These findings may be useful in the design of future stroke therapeutic trials. PMID- 11283392 TI - C-reactive protein in ischemic stroke: an independent prognostic factor. AB - BACKGROUND AND PURPOSE: There is growing evidence of the prognostic importance of C-reactive protein (CRP) in ischemic stroke. However, the independent value of CRP at different stages after stroke has not been established. Therefore, we assessed the prognostic values of CRP in ischemic stroke. We also compared the relation of CRP at admission and discharge with 1-year outcome. METHODS: One hundred ninety-three patients were included in a derivation set (n=128) and a validation set (n=65). Serum CRP was measured, within 24 hours after index ischemic stroke, within 48 to 72 hours, and at hospital discharge. We examined the association between the level of CRP at different stages after stroke and outcome. We adjusted for the possible confounding effect using a multivariate Cox proportional hazard model. RESULTS: A cutoff point of 1.5 mg/dL for CRP at discharge provided optimum sensitivity and specificity for adverse outcome, based on the receiver operator curves. CRP at admission (hazard ratio [HR] 2.78, 95% CI 1.45 to 5.33; P=0.0021) and discharge (HR 9.42, 95% CI 4.27 to 19.05; P<0.0001) were predictors of the combined end point of new vascular events or death at 1 year. CRP at hospital discharge was the strongest independent marker of adverse outcome (HR 7.42, 95% CI 2.75 to 20.03; P=0.0001). These results were confirmed in the validation set (HR 15.66, 95% CI 3.36 to 72.97; P=0.0005). CONCLUSIONS: CRP is a marker of increased 1-year risk in ischemic stroke. CRP at discharge is better related to later outcome and could be of greater utility for risk stratification. These findings are consistent with the hypothesis that elevated CRP may predict future cardiovascular events or death. PMID- 11283393 TI - Direct, longitudinal comparison of (1)H and (23)Na MRI after transient focal cerebral ischemia. AB - BACKGROUND AND PURPOSE: (23)Na MRI may offer new insight into the evaluation of tissue injury. We performed a direct, longitudinal, morphological comparison of (1)H T2 relaxation, (1)H apparent diffusion coefficient (ADC), (23)Na content, and histopathology after cerebral ischemia to address the hypotheses that (a) (23)Na MRI is unique in comparison to (1)H MRI, and (b) accumulation of (23)Na is an unambiguous marker for dead tissue. METHODS: Rats underwent 30 minutes of focal ischemia. MRIs of (1)H T2, (1)H ADC, and (23)Na content were acquired from 12 hours up to 1, 2, or 14 days after reperfusion. On excision, brains were stained with triphenyltetrazolium chloride (TTC). RESULTS: In all cases, the region of abnormality increased in size for 2 days. On day 5, both (1)H T2 and ADC temporarily appeared normal despite the presence of TTC-defined infarction. By comparison, the volume of tissue exhibiting abnormally intense (23)Na signal mirrored the TTC-defined infarct at all time points. CONCLUSIONS: Regions of high (23)Na content correlate well with the TTC-defined infarct and may be a quantitative in vivo marker for dead tissue. In contrast, the dynamics of the (1)H T2 and ADC make it difficult to interpret these images without additional information because they may appear normal despite infarction. Neither type of (1)H image delineates dead tissue, and none of these methods predicts the potential infarct size at early time points. PMID- 11283394 TI - Predicting tissue outcome in acute human cerebral ischemia using combined diffusion- and perfusion-weighted MR imaging. AB - BACKGROUND AND PURPOSE: Tissue signatures from acute MR imaging of the brain may be able to categorize physiological status and thereby assist clinical decision making. We designed and analyzed statistical algorithms to evaluate the risk of infarction for each voxel of tissue using acute human functional MRI. METHODS: Diffusion-weighted MR images (DWI) and perfusion-weighted MR images (PWI) from acute stroke patients scanned within 12 hours of symptom onset were retrospectively studied and used to develop thresholding and generalized linear model (GLM) algorithms predicting tissue outcome as determined by follow-up MRI. The performances of the algorithms were evaluated for each patient by using receiver operating characteristic curves. RESULTS: At their optimal operating points, thresholding algorithms combining DWI and PWI provided 66% sensitivity and 83% specificity, and GLM algorithms combining DWI and PWI predicted with 66% sensitivity and 84% specificity voxels that proceeded to infarct. Thresholding algorithms that combined DWI and PWI provided significant improvement to algorithms that utilized DWI alone (P=0.02) but no significant improvement over algorithms utilizing PWI alone (P=0.21). GLM algorithms that combined DWI and PWI showed significant improvement over algorithms that used only DWI (P=0.02) or PWI (P=0.04). The performances of thresholding and GLM algorithms were comparable (P>0.2). CONCLUSIONS: Algorithms that combine acute DWI and PWI can assess the risk of infarction with higher specificity and sensitivity than algorithms that use DWI or PWI individually. Methods for quantitatively assessing the risk of infarction on a voxel-by-voxel basis show promise as techniques for investigating the natural spatial evolution of ischemic damage in humans. PMID- 11283395 TI - A model for multiparametric mri tissue characterization in experimental cerebral ischemia with histological validation in rat: part 1. AB - BACKGROUND AND PURPOSE: After stroke, brain tissue undergoes time-dependent heterogeneous histopathological change. These tissue alterations have MRI characteristics that allow segmentation of ischemic from nonischemic tissue. Moreover, MRI segmentation generates different zones within the lesion that may reflect heterogeneity of tissue damage. METHODS: A vector tissue signature model is presented that uses multiparametric MRI for segmentation and characterization of tissue. An objective (unsupervised) computer segmentation algorithm was incorporated into this model with the use of a modified version of the Iterative Self-Organizing Data Analysis Technique (ISODATA). The ability of the model to characterize ischemic tissue after permanent middle cerebral ischemia occlusion in the rat was tested. Multiparametric ISODATA measurements of the ischemic tissue were compared with quantitative histological characterization of the tissue from 4 hours to 1 week after stroke. RESULTS: The ISODATA segmentation of tissue identified a gradation of cerebral tissue damage at all time points after stroke. The histological scoring of ischemic tissue from 4 hours to 1 week after stroke on all the animals was significantly correlated with ISODATA segmentation (r=0.78, P<0.001; n=20) when a multiparametric (T2-, T1-, diffusion-weighted imaging) data set was used, less correlated (r=0.70, P<0.01; n=20) when a T2- and T1-weighted data set was used, and not correlated (r=-0.12, P>0.47; n=20) when only a diffusion-weighted imaging data set was used. CONCLUSIONS: Our data indicate that an integrated set of MRI parameters can distinguish and stage ischemic tissue damage in an objective manner. PMID- 11283396 TI - Multiparametric MRI tissue characterization in clinical stroke with correlation to clinical outcome: part 2. AB - BACKGROUND AND PURPOSE: Multiparametric MRI generates different zones within the lesion that may reflect heterogeneity of tissue damage in cerebral ischemia. This study presents the application of a novel model of tissue characterization based on an angular separation between tissues obtained with the use of an objective (unsupervised) computer segmentation algorithm implementing a modified version of the Iterative Self-Organizing Data Analysis Technique (ISODATA). We test the utility of this model to identify ischemic tissue in clinical stroke. METHODS: MR parameters diffusion-, T2-, and T1-weighted imaging (DWI, T2WI, and T1WI, respectively) were obtained from 10 patients at 3 time points (30 studies) after stroke: acute (30 000 people suffer a ruptured intracranial aneurysm, resulting in subarachnoid hemorrhage. Despite the high incidence and catastrophic consequences of a ruptured intracranial aneurysm and the fact that there is considerable evidence that predisposition to intracranial aneurysm has a strong genetic component, very little is understood with regard to the pathology and pathogenesis of this disease. METHODS: To begin characterizing the molecular pathology of intracranial aneurysm, we used a global gene expression analysis approach (SAGE-Lite) in combination with a novel data mining approach to perform a high-resolution transcript analysis of a single intracranial aneurysm, obtained from a 3-year-old girl. RESULTS: SAGE-Lite provides a detailed molecular snapshot of a single intracranial aneurysm. These data suggest that, at least in this specific case, aneurysmal dilation results in a highly dynamic cellular environment in which extensive wound healing and tissue/extracellular matrix remodeling are taking place. Specifically, we observed significant overexpression of genes encoding extracellular matrix components (eg, COL3A1, COL1A1, COL1A2, COL6A1, COL6A2, elastin) and genes involved in extracellular matrix turnover (TIMP-3, OSF-2), cell adhesion and antiadhesion (SPARC, hevin), cytokinesis (PNUTL2), and cell migration (tetraspanin-5). CONCLUSIONS: Although these are preliminary data, representing analysis of only one individual, we present a unique first insight into the molecular basis of aneurysmal disease and define numerous candidate markers for future biochemical, physiological, and genetic studies of intracranial aneurysm. Products of these genes will be the focus of future studies in wider sample sets. PMID- 11283410 TI - Authors should be accurate when describing study design. PMID- 11283411 TI - Distribution and synaptology of vasoactive intestinal polypeptide (VIP) immunoreactive structures in the rat periaqueductal grey. AB - Light microscopic analysis of the rat midbrain periaqueductal grey (PAG) showed vasoactive intestinal polypeptide immunoreactive (VIP-ir) neurons localized at the lateral and ventral walls of the aqueduct. Some varicose VIP-ir elements were detected closely associated with the ependyma. While several VIP-ir elements were encountered immediately under the ependyma, in a few cases, VIP-ir cell bodies were seen on the luminal surface of the ependymal cells lining the aqueduct. Electron microscopy revealed that most of these cells possessed the characteristics of a local circuit neuron. All VIP-ir cells had indented nuclei. Two types were distinguished: one with rounded cell body receiving numerous axo somatic synapses established by VIP-negative axons. The other cell type was fusiform and its surface was almost fully isolated from axonal contacts by a glial sheath. The VIP-ir processes were interconnected with other periaqueductal cells by a variety of synaptic contacts. VIP-ir axon terminals formed asymmetric synapses with immunonegative dendritic shafts often in glomerulus-like assemblies. The postsynaptic immunonegative dendrites were of the aspinous, beaded type. We suggest that VIP-ir cells and processes in the midbrain PAG establish connections between the longitudinal functional columns of this region. On the basis of their morphology, VIP-ir cells in the PAG appear to be excitatory, terminating on inhibitory interneurons. Thus, a VIP-stimulated inhibition may be instrumental in the coordination of responses evoked by the stimulation of PAG columns. PMID- 11283412 TI - Ionic permeability of the frog sciatic nerve perineurium: parallel studies of potassium and lanthanum penetration using electrophysiological and electron microscopic techniques. AB - The isolated sciatic nerve of the frog Rana temporaria was used for a parallel electrophysiological and electron microscopic examination of the ionic permeability of the perineurium, one component of the blood-nerve barrier. Nerves mounted in a grease-gap chamber for electrophysiological recording showed negligible changes in DC potential (Delta DC) or compound action potential on challenge with 100 mM K(+) Ringer, evidence that the perineurium was tight to K(+). In preparations then fixed and exposed to 5 mM lanthanum in the fixative, and examined in the electron microscope, electron-dense lanthanum deposits were seen between perineurial lamellae, but lanthanum was not detectable within the endoneurium, confirming that the perineurium was also tight to lanthanum. Absence of lanthanum penetration was confirmed by X-ray analysis of electron microscopic sections. In nerves exposed to 2 mM sodium deoxycholate (DOC) in the recording chamber, then challenged with high [K(+)], a moderate increase in perineurial K(+) permeability (P(K)) was observed, but lanthanum was still excluded. Exposure of nerves to 4 mM DOC caused a greater increase in perineurial potassium permeability, and the two nerves with the greatest permeability (P(K) > 1 x 10( 5) cm x sec(-1)) also showed detectable lanthanum within the endoneurium. The results indicate that DOC causes a dose-dependent increase in tight junctional permeability in the perineurium, and that the electrophysiological monitoring of K(+) penetration is a more sensitive measure of small ion permeability than electron microscopical analysis using lanthanum as tracer. Vesicular profiles observed in perineurial lamellae did not form open channels for ion flux across the perineurium in control nerves, or in those exposed to DOC. In preparations where lanthanum reached the endoneurium, lanthanum was observed in dense deposits in the extracellular spaces around nodes of Ranvier, and in the outer mesaxon cleft, but did not penetrate the internodal periaxonal space, the myelin intraperiod line, or the Schmidt-Lanterman incisures, in contrast to observations in mammalian nerves. The apparent differences in accessibility of the internodal periaxonal space in frog and mammalian axons are discussed in relation to axonal physiology. The study illustrates the value of parallel electrophysiological and electron microscopic examination in elucidating the properties of extracellular ionic pathways and their role in neural function. PMID- 11283413 TI - Cytoarchitectonic subdivisions in the subtectal midbrain of the lizard Gallotia galloti. AB - Contemporary study of molecular patterning in the vertebrate midbrain is handicapped by the lack of a complete topological map of the diverse neuronal complexes differentiated in this domain. The relatively less deformed reptilian midbrain was chosen for resolving this fundamental issue in a way that can be extrapolated to other tetrapods. The organization of midbrain centers was mapped topologically in terms of longitudinal columns and cellular strata on transverse, Nissl-stained sections in the lizard Gallotia galloti. Four columns extend along the whole length of the midbrain. In dorsoventral order: 1) the dorsal band contains the optic tectum, surrounded by three ventricularly prominent subdivisions, named griseum tectale, intermediate area and torus semicircularis, in rostrocaudal order; 2) a subjacent region is named here the lateral band, which forms the ventral margin of the alar plate and also shows three rostrocaudal divisions; 3) the basal band forms the basal plate or tegmentum proper; it appears subdivided into medial and lateral parts: the medial part contains the oculomotor and accessory efferent neurons and the medial basal part of the reticular formation, which includes the red nucleus rostrally; the lateral part contains the lateral basal reticular formation, and includes the substantia nigra caudally; 4) the median band contains the ventral tegmental area, representing the mesencephalic floor plate. The alar regions (dorsal and lateral) show an overall cellular stratification into periventricular, central and superficial strata, with characteristic cytoarchitecture for each part. The lateral band contains two well developed superficial nuclei, one of which is commonly misidentified as an isthmic formation. The basal longitudinal subdivisions are simpler and basically consist of periventricular and central strata. PMID- 11283414 TI - A light and electron microscopic study of the GABA transporter GAT-3 in the monkey basal ganglia and brainstem. AB - The present study aimed to elucidate the distribution of the GABA transporter GAT 3 in the monkey basal ganglia and brainstem. Very dense GAT-3 immunoreactivity was observed in the medial septum, diagonal band, basal nucleus of Meynert, thalamus, globus pallidus, and substantia nigra. Moderate levels were observed in the subthalamic nucleus, periaqueductal grey, spinal trigeminal and vestibular nuclei. A general light level of staining was observed in the remainder of the brainstem regions, and very light staining was observed in the caudate nucleus and putamen. Electron microscopy showed that GAT-3 immunoreactivity was present in cell bodies with light cytoplasm and dense bundles of glial filaments, and features of astrocytes. Large numbers of astrocytic processes were also labeled in the neuropil. The cell bodies and processes of neurons were unlabeled. Further study is necessary to elucidate GAT-3 expression in neurological conditions, including hyperalgesia and Parkinson's disease. PMID- 11283415 TI - Localization of GABA- and glutamate-like immunoreactivity in the cardiac ganglion of the lobster Panulirus argus. AB - Wholemount immunohistochemical methods were used to examine the localization of gamma-aminobutyric acid (GABA) and glutamate within the cardiac system of the Caribbean spiny lobster Panulirus argus. All of the GABA-like immunoreactivity (GABAi) in the cardiac ganglion originated from a single bilateral pair of fibers that entered the heart via the two dorsal nerves. Each GABAi axon bifurcated upon entering the ganglion and gave rise to varicose fibers that surrounded the somata and initial segments of the five large motor neurons. The four small posterior cells did not appear to receive somatic contacts. Double-labeling experiments in which individual motor neurons were injected with Neurobiotin showed that their dendritic processes, which project to muscle bundles adjacent to the ganglion and are thought to respond to stretch, were also accompanied by branches of the GABAi fibers. Glutamate-like immunoreactivity (GLUi) was present in each of the motor neuron cell bodies. In some preparations, GLUi was also detected in large caliber fibers in the major ganglionic nerves. These fibers gave rise to more slender branches that innervated the cardiac muscle bundles. GLUi was also found in the small cell bodies and in fibers surrounding motor neuron somata. Taken together, these findings support previous electrophysiological, pharmacological and anatomical studies indicating that GABA mediates extrinsic inhibition and that glutamate acts as a neuromuscular and intraganglionic transmitter in this system. While axosomatic contacts may play a major role in both transmitter systems, the GABAergic inhibition also appears to involve substantial axodendritic synaptic signaling. PMID- 11283416 TI - Use of a new myocardial centroid for measurement of regional myocardial dysfunction by electron beam computed tomography: comparison with technetium-99m sestamibi infarct size quantification. AB - RATIONALE AND OBJECTIVES: The study compared the performance of conventional endocardial and epicardial centroid algorithms with the new "myocardial" centroid algorithm in patients with anterior myocardial infarction. "Floating" endocardial or epicardial centroid algorithms, commonly used in tomographic imaging methods to assess regional motion, may misrepresent left ventricular regional myocardial function in the presence of markedly asymmetric left ventricular contraction. METHODS: A new centroid algorithm based on regional myocardial mass distribution was tested in 29 patients with a first anterior myocardial infarction and was compared with conventional centroid algorithms. Direct comparisons in 60 equal sectors at one midventricular level per patient were performed between electron beam computed tomography and technetium-99m sestamibi single-photon emission computed tomography. The thresholds of regional myocardial function used to define infarction were varied for regional ejection fraction from 20% to 40% and for regional wall thickening from 0 to 4 mm. Regression and Bland-Altman analysis were used to compare infarct size by regional myocardial function with infarct size by sestamibi single-photon emission computed tomography. RESULTS: The new myocardial centroid showed the least shift toward infarcted myocardium from diastole to systole and had the highest amplitudes of the measurement curves for regional ejection fraction and regional wall thickening. The optimal regional myocardial function thresholds for each centroid algorithm for regional ejection fraction were endocardial, 30% (R = 0.62; mean difference to sestamibi, -0.5% +/- 22.1% tomographic infarct size points); epicardial, 30% (R = 0.79; mean difference, 2.2% +/- 13.1% tomographic infarct size points); and new myocardial, 25% (R = 0.88; mean difference, -0.6% +/- 9.5% tomographic infarct size points). The optimal thresholds for regional wall thickening were endocardial, 1 mm (R = 0.70; mean difference, -2.2% +/- 14.3% tomographic infarct size points); epicardial, 1 mm (R = 0.78; mean difference, -4.6% +/- 12.7% tomographic infarct size points); and new myocardial, 2 mm (R = 0.71; mean difference, 2.1% +/- 14.1% tomographic infarct size points). The best agreement (R = 0.88) between electron beam computed tomography infarct size and sestamibi single-photon emission computed tomography infarct size was achieved with regional ejection fraction and the new myocardial centroid algorithm. CONCLUSIONS: In asymmetrically contracting left ventricles, the new myocardial centroid algorithm is superior to conventional methods for tomographic analysis of regional myocardial function. PMID- 11283417 TI - Comparison of the mechanical thrombectomy efficacy of the Amplatz thrombectomy device and the Cragg thrombolytic brush in vitro. AB - RATIONALE AND OBJECTIVES: To determine the efficacy of thrombectomy (without thrombolytic agents) for the Amplatz thrombectomy device (ATD) and the Cragg thrombolytic brush catheter (CBC) in vitro. METHODS: Thrombectomy was performed with the ATD or CBC (6F) in a flow model. Embolus sizes, weight, remaining thrombus, and activation time were evaluated. RESULTS: No significant difference in the activation time was found. The CBC produced significantly less embolism (3.3% vs. 0.03% in the 5-mm and 89% vs. 0.5% in the 7-mm model), but also much more thrombus remained in the system than with the ATD (1% vs. 41% in the 5-mm and 0.1% vs. 62% in the 7-mm model). CONCLUSIONS: The ATD can remove almost all thrombus (99%), whereas the CBC removes only up to 60%, producing fewer emboli than the ATD. This might be due to the lower rotational speed of the CBC compared with the ATD, which is 20 times greater. The soft nylon brush offers less resistance and shear force toward the thrombus than the stainless-steel impeller of the ATD. Because of the large amount of remaining thrombus, the CBC should not be used without lytic agents. PMID- 11283418 TI - Does the trabecular bone structure depicted by high-resolution MRI of the calcaneus reflect the true bone structure? AB - RATIONALE AND OBJECTIVES: The purpose of this study was to compare trabecular bone structure parameters assessed with high-resolution magnetic resonance imaging (HR-MRI) with those determined in specimen sections. METHODS: High resolution MR images were obtained for 30 calcaneus specimens with a three dimensional, T1-weighted spin-echo sequence (spatial in-plane resolution 0.195 mm, slice thicknesses of 0.3 and 0.9 mm). Thirty-eight sections were obtained from the specimens, and contact radiography was performed. In the corresponding sections, structural parameters analogous to bone histomorphometry were determined. RESULTS: Significant correlations between MRI-derived structural parameters and those derived from macro pathological sections were found: r values of up to 0.75 were obtained (P < 0.01). The highest correlations were found for apparent bone volume/total volume and trabecular thickness. Image thresholding techniques showed a significant impact on these correlations (P < 0.01). The thinner MR sections were less susceptible to the different thresholding algorithms. CONCLUSIONS: Trabecular bone structure depicted by HR-MR images is significantly correlated with that shown in macro sections (P < 0.01); however, a number of limitations have to be considered, including the substantial impact of thresholding techniques and slice thickness. PMID- 11283419 TI - In vitro evaluation of intravascular stent artifacts in three-dimensional MR angiography. AB - RATIONALE AND OBJECTIVES: To evaluate the intraluminal signal characteristics of various stents and stent-grafts in contrast-enhanced three-dimensional MR angiography (3D MRA) in vitro. METHODS: Fourteen stents made of different materials (steel, nitinol, tantalum, cobalt-based alloy, polyethylene) and six stent-grafts were implanted in plastic tubes simulating the common iliac artery. The tubes were filled with gadopentetate dimeglumine in water at a concentration of 25 mmol/L and positioned in a plastic container filled with water. For imaging, the container was placed in the center of the magnet, parallel, orthogonal, and diagonal to the z axis. A 3D gradient-echo sequence (T1-FFE) was acquired with the following parameters: repetition time 5.3 ms, echo time 1.6 ms, flip angle 50 degrees, slice thickness 1.5 mm, and acquisition matrix 256 with zero filling to 512. To evaluate the influence of the frequency-encoding gradient on the appearance of the artifacts, stents were examined with their axes oriented in all three directions both with the frequency-encoding gradient in the feet head and right-left directions. The size and pattern of stent-related artifacts were evaluated semiquantitatively for each measurement. RESULTS: Five different components of artifacts could be distinguished: homogeneous signal reduction inside the stent, narrowing of the stent lumen, structures of various shapes inside the stents, signal reduction or signal increase at the ends of the stents, and shift of the intraluminal signal orthogonal to the longitudinal axis of the vessel. The size of the artifacts depended heavily on the material of the stent. The polyethylene stent showed no artifacts, the tantalum stent only minor artifacts. Nitinol stents were characterized by artifacts at both ends and signal reduction intraluminally. Stents made of steel demonstrated the strongest artifacts, characterized by almost complete signal loss intraluminally. The characteristics of the artifacts of all stents depended on the direction of the stent relative to the frequency-encoding gradient. CONCLUSIONS: Three-dimensional MRA follow-up after stent placement may be applicable for stent patency evaluation in all instances. However, grading of stenoses seems to be unrealistic in steel stents and in most nitinol stents. PMID- 11283420 TI - Curved reconstructions versus three-dimensional surface rendering in the demonstration of cortical lesions in patients with extratemporal epilepsy. AB - RATIONALE AND OBJECTIVES: To compare the visibility and localization of extratemporal cortical lesions in extratemporal epilepsy by using curved reconstruction (CR) and three-dimensional surface rendering (3D SR) of 3D acquired MR images and to study the degree of confidence with which localizations are made, particularly at the gyral level. METHODS: Twenty patients with extratemporal epilepsy, based on seizure symptomatology and/or scalp electroencephalographic registrations, with an extratemporal structural lesion on conventional MR imaging, were selected for this study by a neuroradiologist with extensive experience in the assessment of epilepsy patients. Transverse T2 spin echo, coronal fluid-attenuated inversion recovery, and transverse 3D-acquired/two dimensionally reconstructed T1 MR images were used for the selection. A second neuroradiologist (observer 1) and a radiology resident (observer 2) assessed CR and 3D SR in random order. Both observers were masked to all patient data. The subjective visibility of lesions and gyral location were scored. The interobserver agreements for lesion visibility and localization and for degree of confidence were compared for CR and 3D SR. RESULTS: For both observers, the lesion was visible in 55% of 3D SRs and 95% of CRs. The proportion with "very clearly visible" lesions on 3D SR was 19% (4/20) according to observer 1 and 30% (6/20) according to observer 2. For CR, this proportion was substantially higher: 55% for both observers. This difference was significant for observer 1 but not for observer 2. The interobserver agreement was high for both methods. Agreement on gyral localization was 28% for CR and 40% for 3D SR. The percentage of similar confidence scores for the same gyral localization and for gyral localization with a maximum difference of one gyrus between the observers did not differ significantly for CR or 3D SR. The observers were more often confident in agreed cases in CR and moderately confident in 3D SR. CONCLUSIONS: These results suggest that CRs of the brain surface are superior to 3D SR for the visualization of extratemporal cortical lesions in patients with drug-resistant extratemporal epilepsy. If lesions are seen, no significant difference was found between the two techniques for localization; however, the degree of confidence appears higher for CR at the gyral level. PMID- 11283421 TI - Renal multigated spectral Doppler imaging before and after captopril challenge. AB - RATIONALE AND OBJECTIVES: A new method of measurement, simultaneous multigated spectral Doppler imaging (MSDI), was used to quantify the hemodynamic changes in kidneys after administration of captopril. METHODS: Forty kidneys in 22 hypertensive patients were included in the study. All underwent MSDI and scintigraphy before and after administration of captopril. RESULTS: With scintigraphy used as the gold standard for diagnosing renal artery stenosis (RAS), three kidneys were found to be positive for RAS. Multigated spectral Doppler imaging detected a decrease in the resistive index after administration of captopril in all patients with RAS and correctly excluded RAS in one patient who was diagnosed as having RAS on scintigraphy. The resistive index was increased after captopril administration in normally functioning kidneys and was unchanged in kidneys with impaired function. CONCLUSIONS: With further supportive evidence, MSDI may prove to be a powerful tool for the acquisition of resistive index information and may increase the value of the resistive index as a physiological hemodynamic parameter in the evaluation of normal and abnormal conditions. PMID- 11283422 TI - Mechanical delivery of aerosolized gadolinium-DTPA for pulmonary ventilation assessment in MR imaging. AB - RATIONALE AND OBJECTIVES: To evaluate a new technique with mechanical administration of aerosolized gadolinium (Gd)-DTPA for MR visualization of lung ventilation. METHODS: Ten experimental procedures were performed in six domestic pigs. Gd-DTPA was aerosolized by a small-particle generator. The intubated animals were mechanically aerosolized with the nebulized contrast agent and studied on a 1.5-T MR imager. Respiratory gated T1-weighted turbo spin-echo images were obtained before, during, and after contrast administration. Pulmonary signal intensity (SI) changes were calculated for corresponding regions of both lungs. Homogeneity of aerosol distribution was graded independently by two radiologists. RESULTS: To achieve a comparable SI increase as attained in previous trials that used manual aerosol ventilation, a ventilation period of 20 minutes (formerly 30 minutes) was sufficient. Mean SI changes of 116% were observed after that duration. Contrast delivery was rated evenly distributed in all cases by the reviewers. CONCLUSIONS: The feasibility of applying Gd-DTPA as a contrast agent to demonstrate pulmonary ventilation in large animals has been described before. The results of this refined technique substantiate the potential of Gd-based ventilation MR imaging by improving aerosol distribution and shortening the nebulization duration in the healthy lung. PMID- 11283423 TI - Effects of different sports on bone density and muscle mass in highly trained athletes. AB - PURPOSE: It is known that participating in sports can have a beneficial effect on bone mass. However, it is not well established which sport is more beneficial for increased bone mineral density (BMD) and appendicular muscle mass (AMM). This study investigated the effects of different high-intensity activities on BMD and AMM in highly trained athletes. MATERIALS AND METHODS: Sixty-two male subjects aged 18--25 yr participated in the study. The sample included judo (J; N = 21), karate (K; N = 14), and water polo (W; N = 24) athletes who all competed at national and international level. Twelve age-matched nonathletic individuals served as the control group (C). All athletes exercised regularly for at least 3 h x d(-1), 6 d x wk(-1). Segmental, total BMD, and AMM were measured with a dual energy x-ray (DXA) absorptiometry (Lunar Corp., Madison, WI). DXA analysis also includes bone mineral content (BMC) and fat and lean masses. RESULTS: Total BMD(C) was significantly lower (mean +/- SD: 1.27 +/- 0.06 g x cm(-2), P < 0.05) than either judo or karate athletes (total BMD(J) (1.4 +/- 0.06 g x cm(-2)) and total BMD(K) (1.36 +/- 0.08 g x cm(-2))) but not different from the W athletes (total BMD(W) (1.31 +/- 0.09 g x cm(-2))). AMM was significantly lower in the C group compared with the three athletic groups (P < 0.05). Fat mass was higher in the W versus J and K athletes but not different from the C group (P < 0.05). CONCLUSIONS: This cross-sectional study has shown that athletes, especially those engaged in high-impact sports, have significantly higher total BMD and AMM than controls. These results suggest that the type of sport activity may be an important factor in achieving a high peak bone mass and reducing osteoporosis risk. PMID- 11283424 TI - Effect of energy restriction and exercise on vitamin B-6 status of women during lactation. AB - PURPOSE: Lactation increases vitamin B-6 requirements because its concentration in breast milk is related to maternal intake and it is essential for infants. Exercise may also increase the requirement because it increases utilization and excretion of vitamin B-6. Therefore, the purpose of this study was to determine whether energy restriction and exercise affected vitamin B-6 status of lactating women. METHODS: Breastfeeding women with a body mass index > or = 25 and < or = 30 kg x m(-2) were randomly assigned at 4 wk postpartum to either restrict energy intake by 500 kcal x d(-1) and exercise for 45 min x d(-1), 4 d x wk(-1) (weight loss group, WG) or maintain usual diet and not exercise (control group, CG) for 10 wk. Women were given a supplement containing 2.0 mg of vitamin B-6. Measurements included vitamin B-6 concentrations in breast milk and plasma, plasma pyridoxal 5'-phosphate, and erythrocyte alanine aminotransferase activity. RESULTS: The WG lost more weight (-4.4 +/- 0.4 vs -0.9 +/- 0.5 kg, P < 0.01) than the CG. Cardiovascular fitness increased by 12% in the WG, compared to 3% in the CG (P = 0.09). Milk vitamin B-6 concentrations increased in both groups (161 +/- 107 and 191 +/- 85 nmol x L(-1), WG and CG, respectively, P = 0.05). There were no significant differences in other vitamin B-6 parameters. Weight and length gain (2.06 +/- 0.21 and 1.83 +/- 0.17 kg; 8.6 +/- 0.6 and 7.2 +/- 0.5 cm; WG and CG, respectively) of infants was not significantly different between groups. CONCLUSIONS: Energy restriction and exercise from 4 to 14 wk postpartum in overweight, breastfeeding women consuming adequate dietary intakes and 2.0 mg of supplemental vitamin B-6 does not adversely affect vitamin B-6 status or infant growth. PMID- 11283425 TI - Exercise training and energy restriction decrease neutrophil phagocytic activity in judoists. AB - PURPOSE: To investigate the effects of weight reduction as the result of exercise training and energy restriction on neutrophil function. METHODS: Eighteen male competitive college judoists participated in the study. In a whole blood assay, oxidative burst activity, phagocytic activity, expressions of Fc gamma receptor 3 (CD16), and complement receptor 3 (CD11b) of neutrophils were measured on a per cell basis by flow cytometry at day 20, 5, and 1 before and at day 7 after the competition. RESULTS: The rate of neutrophil producing reactive oxygen species decreased before the competition, whereas the oxidative burst activity per cell increased significantly in all subjects, which resulted in a significant increase of the total oxidative burst activity. However, there were no significant effect of energy restriction on oxidative burst activity. The rate of neutrophils incorporating opsonized zymosan decreased significantly with energy restriction. The total phagocytic activity of 10,000 neutrophils and the phagocytic activity per cell also decreased significantly by severe energy restriction. The surface antigen expressions of CD11b and CD16 were unaffected by weight reduction. CONCLUSIONS: The results suggest that with respect to the management of health conditions, weight reduction for judoists should be composed of exercise training and energy restriction should be moderate. PMID- 11283426 TI - Exercise training in heart failure: recommendations based on current research. AB - A review of methods used for exercise training in stable chronic heart failure patients (CHF) shows a lack of standardization to guide prescription. Previous recommendations have been adopted from fitness training or rehabilitation studies. A model for use in CHF patients requires specific guidelines which respect the various manifestations of this illness. Pathology and exercise tolerance of patients with CHF allow only a few selected activities to be performed, such as walking and cycle ergometer training. Although the steady state method has usually been applied for aerobic exercise, the interval method has been shown to cause greater exercise stimuli to peripheral muscle than that obtained during steady state training methods without inducing greater cardiovascular stress. There is no consensus at present as to an optimal parameter for measuring intensity. An intensity of 40-80% peak oxygen consumption (VO(2)) has been applied successfully. A heart rate reserve of 60-80% or 75% of peak heart rate was used as a guide to exercise intensity without consideration of the impaired force-frequency relationship in myocardial performance. Because intensity, duration, and frequency of exercise are closely interrelated, initial exercise should be kept at 40-50% peak VO(2) with exercise duration of > 3-5 min x session performed several times daily. Progression should be followed in this order: duration, then frequency, then intensity. Resistance training can be recommended when small muscle groups are involved, using short bouts of work phases and small numbers of repetitions. To increase respiratory muscle strength and endurance, resistive inspiratory muscle training at intensity 25--35% maximum inspiratory pressure, and performed 20-30 min x d(-1), is recommended. On the basis of currently available research, supervised inpatient training programs should be preferred. Future research should be performed with respect on statistically sufficient, randomized, and controlled long-term studies that compare different training modes, intensities, frequency/duration ratios, and rates of progression. PMID- 11283427 TI - Effect of strength training on resting metabolic rate and physical activity: age and gender comparisons. AB - PURPOSE: The purpose of this study was to compare age and gender effects of strength training (ST) on resting metabolic rate (RMR), energy expenditure of physical activity (EEPA), and body composition. METHODS: RMR and EEPA were measured before and after 24 wk of ST in 10 young men (20-30 yr), 9 young women (20-30 yr), 11 older men (65-75 yr), and 10 older women (65-75 yr). RESULTS: When all subjects were pooled together, absolute RMR significantly increased by 7% (5928 +/- 1225 vs 6328 +/- 1336 kJ.d-1, P < 0.001). Furthermore, ST increased absolute RMR by 7% in both young (6302 +/- 1458 vs 6719 +/- 1617 kJ x d(-1), P < 0.01) and older (5614 +/- 916 vs 5999 +/- 973 kJ x d(-1), P < 0.05) subjects, with no significant interaction between the two age groups. In contrast, there was a significant gender x time interaction (P < 0.05) for absolute RMR with men increasing RMR by 9% (6645 +/- 1073 vs 7237 +/- 1150 kJ x d(-1), P < 0.001), whereas women showed no significant increase (5170 +/- 884 vs 5366 +/- 692 kJ x d(-1), P = 0.108). When RMR was adjusted for fat-free mass (FFM) using ANCOVA, with all subjects pooled together, there was still a significant increase in RMR with ST. Additionally, there was still a gender effect (P < 0.05) and no significant age effect (P = NS), with only the men still showing a significant elevation in RMR. Moreover, EEPA and TEE estimated with a Tritrac accelerometer and TEE estimated by the Stanford Seven-Day Physical Activity Recall Questionnaire did not change in response to ST for any group. CONCLUSIONS: In conclusion, changes in absolute and relative RMR in response to ST are influenced by gender but not age. In contrast to what has been suggested previously, changes in body composition in response to ST are not due to changes in physical activity outside of training. PMID- 11283428 TI - Skeletal muscle oxidative capacity and exercise tolerance in rats with heart failure. AB - PURPOSE: Past research has shown the development of exercise intolerance after myocardial infarction (MI). The purpose of this study was to test the hypothesis that reductions in oxidative enzyme activity, in a variety of skeletal muscles, coincide with the development of exercise intolerance in a rat model of chronic heart failure (CHF) induced by MI. METHODS: The animals were initially divided into two groups: sham-operated controls (Sham) and animals in which a MI was surgically induced. MI rats were then subdivided into two groups according to left ventricular end-diastolic pressure (LVEDP): <20 mm Hg [small MI (SMI)] and > 20 mm Hg [large MI (LMI)]. Exercise tolerance was measured by performing a progressive run to fatigue test (RTF). Citrate synthase (CS), 3-hydroxyacyl CoA dehydrogenase (HADH), and malate dehydrogenase (MDH) activities were measured in six hindlimb muscles. RESULTS: After approximately 6 wk of recovery, LVEDP differed among groups (P < 0.05): Sham (1 +/- 1 mm Hg, N = 7), SMI (7 +/- 2 mm Hg, N = 7), and LMI (30 +/- 2 mm Hg, N = 6). RTF was 20 +/- 1 min for Sham, 25 +/ 3 min for SMI, and 11 +/- 2 min for LMI (P < 0.05 for LMI vs Sham, SMI). Significant reductions in enzyme activity were found for all three enzymes in the red portion of the gastrocnemius muscles of LMI. However, no significant correlation was found between RTF and CS, HADH, or MDH in any muscle of the three groups of animals. DISCUSSION: The results of the present study demonstrate that severe left ventricular dysfunction is associated with reductions in exercise tolerance and modest decreases in oxidative enzyme activities in selected muscles. It does not appear, however, that the development of exercise intolerance in CHF and oxidative enzyme activities are mechanistically related to one another. PMID- 11283429 TI - Anaerobic exercise induces moderate acute phase response. AB - PURPOSE: It was intended to compare the immune reaction after single and repeated short bouts of anaerobic exercise. METHODS: Twelve unspecifically trained male subjects (27 +/- 2 yr, 75 +/- 2 kg, VO(2peak) 52 +/- 2 mL x min(-1) x kg(-1)) performed one 60-s all-out test (SMT) on a cycling ergometer and the same test followed by eight 10-s all-out tests every 5 min (AN-TS). These tests and one control day (Co-Day) were applied in randomized order. At rest and 15 min, 2 h, and 24 h after cessation of exercise the following venous blood parameters were determined: concentration of neutrophils and (CD16(+ -)) premacrophages (both flow-cytometrically), interleukin 6 and 8 (IL-6, IL-8), C-reactive protein (CRP) and cortisol. RESULTS: Two hours after cessation of exercise the neutrophils increased stronger after AN-TS than after SMT (P < 0.01). The peak in the number of premacrophages occurred earlier after SMT (15 min post; P < 0.01 to Co-Day) than after AN-TS (2 h post; P < 0.05 to Co-Day). IL-6 was elevated at 15 min and 2 h after AN-TS (P < 0.01 to SMT and Co-Day) but only slightly 2 h after SMT (P < 0.01 to Co-Day). There were no significant changes in IL-8. CRP was the only elevated parameter 24 h postexercise exclusively after AN-TS (P < 0.05 to Co Day). CONCLUSIONS Repeated short anaerobic bouts of cycling lead to an acute phase response, which is more pronounced than after a single bout. Athletes should take care in performing such training sessions several times a week because signs of inflammation are detectable even 24 h after cessation of exercise. PMID- 11283430 TI - Exercise training reduces ischemic myocardial dysfunction. AB - PURPOSE: This study tested the hypothesis that exercise training improves myocardial blood flow and regional myocardial contractile function in a lateral border zone located adjacent to the ischemic zone during coronary artery occlusion. METHODS: Fourteen dogs were subjected to either 12 wk of dynamic exercise training or cage rest. Dogs were anesthetized and instrumented to assess regional myocardial contractile function (percent segment length shortening and rate of shortening) and regional myocardial blood flow (tracer microspheres) in the central ischemic, lateral border, and nonischemic zones. Measurements were made preocclusion and at 2 min and 3 h after occlusion of the left anterior descending coronary artery (CAO). RESULTS: Contractile function and regional myocardial blood flow were not affected by CAO in the nonischemic zone in both cage-rested and exercise trained dogs. Regional myocardial contractile function and blood flow in the lateral border zone were significantly higher in exercise trained dogs compared with cage-rested dogs, both at 2 min and 3 h after CAO. Ischemic dysfunction was similar in the central ischemic zone in both cage-rested and exercise trained dogs both at 2 min and 3 h after CAO. Regional myocardial blood flow was similarly reduced in the ischemic zone in both groups after 2 min of CAO, but was significantly higher in the inner (subendocardial) region of the exercised trained hearts after 3 h (P < 0.05). CONCLUSION: These data suggest that there was greater border zone perfusion in exercise trained animals during prolonged CAO, which was associated with significantly improved myocardial contractile function. PMID- 11283431 TI - Effect of aerobic and anaerobic metabolism on free radical generation swimmers. AB - PURPOSE: In this study, changes in antioxidant systems due to free radicals were investigated in short distance (100-m) and long-distance (800-m) swimmers, within whom the anaerobic and aerobic metabolisms dominate, respectively. METHODS: For this study, swimmers aged between 15 and -21 yr swam 800 m (N = 10) and 100 m (N = 9). Venous blood samples were taken before swimming, and at 1-, 20-, and 40-min intervals after swimming. Lactate, catalase (CAT), glutathione peroxidase (GPx), and reduced glutathione (GSH) levels were determined in the blood samples. RESULTS: The increase of lactate levels was statistically significant in the swimmers, both after the 100- and 800-m distances as compared with the preswimming levels (P < 0.001, P < 0.001). Catalase activity was increased in the first minute postswimming as compared with preswimming levels. Catalase activity then decreased at the 20- and 40-min intervals as compared with the 1-min postswimming interval, at both 100- and 800-m distances (P < 0.01, P < 0.001). GPx activity was also increased in the first minute after swimming as compared with preswimming levels. GPx activity then decreased at the 20- and 40-min intervals when compared with the 1-min postswimming level. This occurred in both 100- and 800-m swimmers (P < 0.001, P < 0.001). GSH activity was decreased in the first minute after swimming, compared with the preswimming levels. GSH activity then increased at the 20- and 40-min postswimming intervals, as compared with the first-minute level. Again, this occurred in both the 100- and 800-m swimmers (P < 0.001, P < 0.01). CONCLUSION: We concluded that both long-distance and particularly short-distance (100-m) swimming increased the activities of antioxidant defense enzymes. PMID- 11283432 TI - Increased neuroendocrine response to a repeated bout of endurance exercise. AB - This study was designed to compare a first bout of high-intensity endurance exercise with a second bout of similar exercise on the same day, and thereby test the hypothesis that the endocrine response elicited by a second bout is more pronounced compared with a single bout of exercise. Nine male, elite endurance athletes participated in three trials of 24-h duration: 1) complete bed rest (REST), 2) one bout of exercise (ONE), and 3) two bouts of exercise separated by a 3-h rest period (TWO). Each exercise bout consisted of a 10-min warm-up at 50% of VO(2max) followed by 65 min at 75% of VO(2max) on a cycle ergometer. Exercise was performed between 11:00 a.m. and 12:15 a.m. (only in TWO) and 3:15 and 4:30 p.m. (both ONE and TWO). The subjects rested in bed at all hours except when exercising. Blood was sampled 11 times at identical time-points until 7:30 a.m. the next morning. We observed significantly increased levels of epinephrine, norepinephrine, ACTH, cortisol, and growth hormone, and decreased levels of testosterone during and/or after the second bout of exercise compared with the first bout. No difference was observed for insulin, follicle stimulating hormone, luteinizing hormone, thyroid stimulating hormone, free fraction of thyroxin or insulin-like growth factor 1. Thus, this study demonstrates a more pronounced neuroendocrine response to a second bout of exercise on the same day compared with a first/single bout, involving both the sympatho-adrenal system and the hypothalamo-pituitary-adrenal axes. PMID- 11283433 TI - Physical activity and fat-free and fat mass by bioelectrical impedance in 3853 adults. AB - OBJECTIVE: To determine the effects of regular physical activity on body composition, as measured by bioelectrical impedance analysis (BIA), in a large Caucasian population of healthy subjects between 15 and 64 yr of age, and to observe the cross-sectional changes in body composition with increasing age. DESIGN: Cross-sectional comparison between sedentary and physically active adults (at least 3 h x wk(-1) at moderate or hard intensity level activity) during aging. SUBJECTS: A total of 3853 healthy adults (1036 sedentary and 1019 physically active men, and 1280 sedentary and 518 physically active women) between 15 and 64 yr of age. MEASUREMENTS: Height, weight, body mass index (BMI), and fat-free mass (FFM), fat mass, and % fat mass measured by 50-kHz BIA. RESULTS: Higher weight in older sedentary adults was due to a higher fat mass. In 55- to 64-yr-olds compared with 25- to 34-yr-olds, fat mass was 5.5 kg (P < 0.0001) higher in sedentary and 0.6 kg (P < 0.3) higher in physically active men, and 4.5 kg (P < 0.0001) and 2.0 kg (P < 0.04) higher in sedentary and physically active women, respectively. Physical activity was able to limit fat mass and weight gain in men over 25 yr of age and in women until 54 yr of age. Endurance type physical activity was not associated with increased FFM. For the same BMI, sedentary men and women have < 0.7 kg (P < 0.001) higher fat mass than physically active men and women. CONCLUSION: Therefore, the benefits of physical activity seem to include maintenance or prevention of an increase of BMI that in turn correlates with prevention of a fat mass increase for physically active subjects. PMID- 11283434 TI - Fitness, fatness, and estimated coronary heart disease risk: the HERITAGE Family Study. AB - PURPOSE: To determine the contributions of fatness and fitness to the estimated risk of future coronary heart disease (CHD). METHODS: The sample consisted of 212 black and 411 white adult sedentary participants. Percent body fat (%BF) was measured using densitometry, whereas maximal oxygen uptake (VO(2max)) was measured on a cycle ergometer. Risk of future CHD was estimated using the revised Framingham Heart Study algorithm. RESULTS: For fatness, the odds ratios for risk of future CHD were 1.83 and 1.70 for the moderate and high tertiles, respectively, compared with the low tertile. Similarly, the odds ratios for VO(2max) were 1.29 (NS) and 1.62, for the moderate and low tertiles, respectively. Removing VO(2max) from the full model had no effect; however, removing %BF resulted in a significantly weaker model (chi(2) = 10.38, P < 0.01). CONCLUSION: Both fatness and fitness are important predictors of risk of future CHD, based on the Framingham index. PMID- 11283435 TI - Do 9- to 12 yr-old children meet existing physical activity recommendations for health? AB - PURPOSE: The purpose of this study was to investigate whether a sample of 9- to 12-yr-old children fulfilled existing U.S. and UK physical activity recommendations for health. METHODS: Habitual physical activity levels of 79 pre- and early pubertal children were assessed using continuous heart rate monitoring over a period of 4 d. Mean, daily, cumulative physical activity levels spent above heart rate thresholds were compared with U.S. and UK recommended values. RESULTS: In this study, when cumulative amounts of physical activity were considered, at intensities greater than 120 beats x min(-1) and 75% above resting heart rate, pre- and early pubertal British children appear to engage in sufficient physical activity to meet U.S. and UK minimum daily recommendations. CONCLUSIONS: Interpretations of children's physical activity levels depend on thresholds set for intensity of physical activity and whether cumulative or continuous bouts of activity are included in the analysis. PMID- 11283437 TI - Gender differences in adult foot shape: implications for shoe design. AB - PURPOSE: To analyze gender differences in foot shape in a large sample of young individuals. METHODS: Univariate t-tests and multivariate discriminant analyses were used to assess 1) significant differences between men and women for each foot and leg dimension, standardized to foot length, 2) the reliability of classification into gender classes using the absolute and standardized variable sets, and 3) the relative importance of each variable to the discrimination between men and women. RESULTS: Men have longer and broader feet than women for a given stature. After normalization of the measurements by foot length, men and women were found to differ significantly in two calf, five ankle, and four foot shape variables. Classification by gender using absolute values was correct at least 93% of the time. Using the variables standardized to foot length, gender was correctly classified 85% of the time. CONCLUSIONS: This study demonstrates that female feet and legs are not simply scaled-down versions of male feet but rather differ in a number of shape characteristics, particularly at the arch, the lateral side of the foot, the first toe, and the ball of the foot. These differences should be taken into account in the design and manufacture of women's sport shoes. PMID- 11283436 TI - Familial aggregation of submaximal aerobic performance in the HERITAGE Family study. AB - PURPOSE: This study examines the contribution of genetic factors to submaximal aerobic performance phenotypes measured before and after 20 wk of endurance training. METHODS: Submaximal oxygen consumption (VO(2)) at three power outputs, 50 W (VO(2)50W), 60% (VO(2)60%) and 80% (VO(2)80%) of VO(2max) and power outputs at 60% (PO60%) and 80% (PO80%) of VO(2max) were measured during cycle ergometer exercise tests in 483 subjects from 99 white families participating in the HERITAGE Family study. The baseline phenotypes were adjusted for the effects of age, sex, and body mass using stepwise multiple regression procedures. The response phenotypes, computed as the difference (Delta) between the posttraining and baseline measures, were adjusted for age, sex, and the baseline value. RESULTS: All submaximal exercise phenotypes measured at baseline and in response to training were characterized by a significant familial resemblance. Maximal heritabilities of the baseline phenotypes range from 48% to 74% with significant spouse, sibling, and parent-offspring correlations. The hypothesis of maternal inheritance where mother-offspring and sibling correlations were forced to be equal was found to fit the data for VO(2)60%, VO(2)80% and PO80%. For the response phenotypes, the maximal heritabilities tended to be lower (23--57%) with a significant maternal inheritance for Delta VO(2)60%, Delta PO60%, and Delta PO80%. CONCLUSION: These results suggest that the submaximal working capacities of sedentary subjects and their responses to endurance training are influenced by familial/genetic factors with a significant contribution of maternal inheritance. PMID- 11283438 TI - Muscle function at the wrist after eccentric exercise. AB - PURPOSE: The purpose of this experiment was to investigate the effects of eccentric exercise by the wrist extensor muscles on the function and motor control of synergist wrist extensor muscles and the antagonist wrist flexor muscles. METHODS: Ten subjects were tested repeatedly over a period of 11 d, once before and four times after a bout of strenuous eccentric exercise with the wrist extensor muscles. Tests performed as indicators of muscle injury were wrist extension MVC, ROM, and soreness. Tests performed as measures of function and motor control were maximum joint velocity, ability to sustain a constant torque, and the ability to track a changing torque. RESULTS: Indicators of muscle injury: subjects exhibited a decline in wrist extension MVC and ROM, which peaked on day 1, and reported that muscle soreness was greatest on day 2. All measures returned to baseline values by day 10. Measures of function and motor control: subjects exhibited a greater difficulty sustaining a submaximal contraction and tracking torque after eccentric exercise. Greater torque variances in these tests were most evident at high torque levels. Subjects exhibited the greatest difficulty 24 h after eccentric exercise and had recovered by day 10. There was no change in maximal wrist extension velocity. CONCLUSIONS: Strenuous eccentric exercise by wrist extensors had an effect on function and motor control of the wrist extensor muscles. The effect was most evident during contractions in which high torque was required. The response of all of the wrist extensors after the exercise bout was similar, suggesting that they operated in a synergistic manner. The antagonists wrist flexors showed increased coactivation after eccentric exercise. PMID- 11283439 TI - The reliability of cycling efficiency. AB - PURPOSE: The aim of this experiment was to establish the reproducibility of gross efficiency (GE), delta efficiency (DE), and economy (EC) during a graded cycle ergometer test in seventeen male subjects. METHODS: All subjects performed three identical exercise tests at a constant pedal cadence of 80 rpm on an electrically braked cycle ergometer. Energy expenditure was estimated from measures of oxygen uptake (VO(2)) and carbon dioxide production (VCO(2)) by using stoichiometric equations. RESULTS: The subjects characteristics were age 24 +/- 6 yr, body mass 74.6 +/- 6.9 kg, body fat 13.9 +/- 2.2%, and VO(2max) 61.9 +/- 2.4 mL x kg(-1) x min(-1) (all means +/- SD). Average GE, DE, and EC for the three tests were 19.8 +/- 0.6%, 25.8 +/- 1.5%, and 5.0 +/- 0.1 kJ x L(-1), respectively. The coefficients of variation (confidence limits) were GE 4.2 (3.2-6.4)%, DE 6.7 (5.0 10.0)%, and EC 3.3 (2.4-4.9)%. GE was significantly lower at 95 W and 130 W when compared with 165 W, 200 W, 235 W, 270 W, and 305 W. GE at 165 W was significantly lower (P < 0.05) that GE at 235 W. A weak correlation (r = 0.491; P < 0.05) was found between peak oxygen uptake (VO(2peak)) and GE, whereas no correlations were found between VO(2max) and DE or EC. CONCLUSION: We conclude that a graded exercise test with 3-min stages and 35-W increments is a method by which reproducible measurements of both GE and EC can be obtained, whereas measurements of DE seemed slightly more variable. PMID- 11283440 TI - Evaluation of pharmacological aids on physical performance after a transmeridian flight. AB - PURPOSE: The purpose of this study was to evaluate physical performance (static and dynamic) of U.S. Air Force reservists after an eastbound air travel across seven time zones and to estimate the pharmacological aids slow-release caffeine and melatonin versus placebo in attempt to overcome the decline in performance. METHODS: 27 American volunteers were randomly divided into three groups: caffeine 300 mg, melatonin 5 mg, and placebo (lactose capsules). Two days before the flight and 10 d after, three tests were performed: hand grip strength test (static performance), squat jump test (maximal height), and multiple jump test (power and endurance). All measures were repeated twice a day: morning and afternoon. RESULTS: In placebo conditions, the static performance of the dominant hand decreased significantly during the first three mornings and tended to decrease the fourth morning. Simultaneously, the caffeine group's static performance increased significantly, whereas the melatonin group maintained its levels. No significant differences were observed the afternoons. No statistical differences appeared for the nondominant hand in the mornings or afternoons. Dynamic capacities presented no significant degradation after the travel. In the placebo group, for the squat jump test, performance increased from the fourth day. No real explanation can be given about this result. CONCLUSIONS: We demonstrated that slow-release caffeine and melatonin might be used to compensate for jet-lag troubles and particularly for the static physical performance decrease. The slow-release caffeine seems to be the best treatment, but its effects are only demonstrated on previously damaged performance. These preliminary results need further investigation, but we are the first to report a beneficial effect of slow-release caffeine and melatonin on physical performances after jet-lag. PMID- 11283442 TI - Comparison of incremental treadmill exercise and free range running. AB - PURPOSE: The aim of this study was to compare physiological responses during incremental treadmill exercise and free range running. METHODS: Fifteen competitive cross-country runners performed an incremental treadmill test and an unpaced 1-mile run on an indoor 200-m track. Physiological variables (VO(2peak), HR(peak), VO(2) x HR(-1)(peak), V(Epeak)) were measured using a portable metabolic analyzer. Blood lactate was measured post exercise. Outcome variables were analyzed with repeated measures ANOVA. RESULTS: Although directionally similar to previous studies with cycle ergometry, the observed peak values (track vs treadmill) for VO(2) (63.0 +/- 7.4 vs 61.9 +/- 7.2 mL x kg(-1) x min(-1)), V(E) (147 +/- 37 vs 144 +/- 30 L x min(-1)), HR (188 +/- 5 vs 189 +/- 7 beats.min 1), and VO(2) x HR(-1) (22.1 +/- 4.4 vs 21.5 +/- 4.5) were not significantly different. The observed peak values for blood lactate (14.4 +/- 3.3 vs 11.7 +/- 3.0 mmol x L(-1)) were significantly (P < 0.05) different. CONCLUSIONS: The results are not in full agreement with previous findings from cycling studies with the exception of post exercise blood lactate. Whether this represents a fundamental lack of effect of free range exercise or is related to mode specificity remains to be determined. PMID- 11283441 TI - Low-volume circuit versus high-volume periodized resistance training in women. AB - PURPOSE: The purpose of this investigation was to determine the long-term training adaptations associated with low-volume circuit-type versus periodized high-volume resistance training programs in women. METHODS: 34 healthy, untrained women were randomly placed into one of the following groups: low-volume, single set circuit (SSC; N = 12); periodized high-volume multiple-set (MS; N = 12); or nonexercising control (CON) group (N = 10). The SSC group performed one set of 8 12 repetitions to muscular failure 3 d x wk(-1). The MS group performed two to four sets of 3-15 repetitions with periodized volume and intensity 4 d x wk(-1). Muscular strength, power, speed, endurance, anthropometry, and resting hormonal concentrations were determined pretraining (T1), after 12 wk (T2), and after 24 wk of training (T3). RESULTS: 1-RM bench press and leg press, and upper and lower body local muscular endurance increased significantly (P < or = 0.05) at T2 for both groups, but only MS showed a significant increase at T3. Muscular power and speed increased significantly at T2 and T3 only for MS. Increases in testosterone were observed for both groups at T2 but only MS showed a significant increase at T3. Cortisol decreased from T1 to T2 and from T2 to T3 in MS. Insulin-like growth factor-1 increased significantly at T3 for SSC and at T2 and T3 for MS. No changes were observed for growth hormone in any of the training groups. CONCLUSION: Significant improvements in muscular performance may be attained with either a low-volume single-set program or a high-volume, periodized multiple-set program during the first 12 wk of training in untrained women. However, dramatically different training adaptations are associated with specific domains of training program design which contrast in speed of movement, exercise choices and use of variation (periodization) in the intensity and volume of exercise. PMID- 11283443 TI - Effects of resistance training on insulin-like growth factor-I and IGF binding proteins. AB - PURPOSE: Our goal was to determine the effects resistance training on circulating IGF-I and on two of its major binding proteins, IGFBP-1 and IGFBP-3. Additional goals were to compare the time course of hormonal changes with the time course of strength changes and to determine the effect of training volume on the extent of hormonal changes. METHODS: Thirty-one men and women (mean age = 37 +/- 7 yr) completed a 25-wk, 3 d x wk(-1) program in which they performed single-set resistance training (1-SET, N = 11), multiple-set resistance training (3-SET, N = 11), or no exercise (Control, N = 9). Before training, and after 13 and 25 wk of training, blood hormones were analyzed and strength was assessed as the sum of one-repetition maximum (1-RM) for leg extension and chest press exercises. RESULTS: During the first 13 wk of resistance training, circulating IGF-I increased by approximately 20% in both the 1-SET and 3-SET groups (P = 0.041). No further increases occurred between 13 and 25 wk. In the 3-SET group, IGFBP-3 decreased 20% between 13 and 25 wk (P = 0.008). Training did not alter IGFBP-1. Increases in 1-RM strength occurred mainly during the first 13 wk of training and were significantly higher with 3-SET training compared to 1-SET. CONCLUSIONS: These findings indicate that increased circulating IGF-I may, at least in part, mediate increases in strength that result from resistance training. PMID- 11283444 TI - Cardiovascular, metabolic, and hormonal parameters in professional tennis players. AB - During the past decade, the physical and mental stress in professional tennis has been constantly increasing. The overall intensity in tennis ranges between 60 and 70% of maximum oxygen uptake and the energy requirements are mainly provided by aerobic energy metabolism. Therefore, particularly with respect to the duration of the tournaments and the length of the matches, a good aerobic capacity promotes continuous success in professional tennis. During frequent periods of high intensity, however, muscular energy is derived from anaerobic glycolysis. Therefore, sports-specific conditioning programs in tennis should improve both glycolytic and oxidative muscular metabolism. Years of training and competition induce a number of cardiovascular and metabolic adaptations: an increase in heart size in terms of an athlete's heart, higher oxygen uptake capacity, improved muscular oxidative enzyme activities, reduced baseline catecholamine levels, and a lower resting heart rate. In addition, tennis induces side-specific increments in bone density, bone diameter, and bone length of the upper extremity. Furthermore, structural and functional adaptations of the conducting arteries in the preferred arm could be demonstrated in professional tennis players. In conclusion, tennis is a very complex sport involving strength, power, speed, agility and explosiveness, as well as endurance components. Scientific data on exercise-related cardiovascular and metabolic parameters in professional tennis are important to evaluate the players individual fitness level and will help to improve sports-specific conditioning programs. This in turn will not only enhance performance but also prevent overstrain and burnout syndromes. PMID- 11283445 TI - Cardiovascular response of trained and untrained old men to mental challenge. AB - PURPOSE: The cardiovascular response to mental challenge of trained and untrained older men was examined. METHODS: Blood pressure, heart rate, stroke volume, cardiac output, total peripheral resistance (TPR), rate pressure product, and cardiac contractility of 10 aerobically trained (trained) and 10 untrained (untrained) older men during and recovering from the Stroop task were compared. Fitness was assessed by online, open-circuit spirometry using a bicycle ergometer. RESULTS: Trained compared with untrained men showed significantly lower absolute heart rate during and recovering from the Stroop and greater left ventricular ejection time recovering from Stroop. Delta TPR scores were greater for Trained during and recovering from the Stroop, whereas delta systolic blood pressure was greater recovering from Stroop. CONCLUSION: These results indicate that although aerobically trained men possessed lower absolute heart rate their change in TPR and systolic blood pressure during mental challenge was significantly greater than that of untrained older men. The greater vascular response of the trained to mental challenge may reflect greater sensitivity to alpha-adrenergic stimulation. PMID- 11283447 TI - Stability and convergent validity of three physical activity assessments. AB - PURPOSE: The purpose of this study was to examine the stability and convergent validity of heart rate (HR) monitoring, Caltrac accelerometer, and physical activity recall (PAR) in sixth-grade girls during normal weekday activities. METHODS: 46 sixth-grade girls (age 12 +/- 0.6) wore HR monitors and Caltrac accelerometers for 3 d during school, after school, and evenings. We also obtained a PAR for each day. Data were compared on the basis of kcal x h(-1). RESULTS: Two days' worth of data were analyzed for each participant. Intraclass correlation coefficients obtained by use of repeated measures ANOVA revealed that HR monitoring (r = 0.99) and PAR (r = 0.98) were extremely stable across 2 d, whereas Caltrac was moderately stable (r = 0.76). Pearson correlations between techniques were HR versus PAR, r = 0.50 (P < 0.01), HR versus Caltrac, r = 0.28, and Caltrac versus PAR, r = 0.76 (P < 0.01). Methods comparison plots showed poor individual subject agreement between all three types of assessment. CONCLUSION: HR and PAR were stable across 2 d. PAR underestimated caloric expenditure by approximately 14%. Caltrac showed the least utility in both reliability and validity. PMID- 11283446 TI - Dual x-ray absorptiometry, bioelectrical impedance, and near infrared interactance in obese women. AB - PURPOSE: Whether the evaluation of body composition in obese people using low cost, simple bedside, two-compartment techniques reflects the data obtained by indirect methods such as dual x-ray absorptiometry (DEXA) remains controversial. The aim of this study was to compare the data obtained by three methods of assessment of body composition (DEXA; bioelectrical impedance, BIA; and near infrared interactance, NII). METHOD: Data on body composition obtained in 53 obese women by these three methods were compared, using the Bland and Altman procedure to test the relative validity. RESULTS: Although the correlation coefficients between DEXA and the two other methods were high, there were some major differences (limits of agreement) between data concerning fat and lean mass. CONCLUSIONS: The present study indicates that these methods cannot be considered as interchangeable and raises some questions on the use of BIA and NII as a single method of evaluation of body composition in clinical research and practice in obese populations. PMID- 11283448 TI - VO(2) slow component: to model or not to model? AB - PURPOSE: The purpose of this study was to compare several techniques often used in the literature for measuring the amplitude of the slow component of oxygen uptake kinetics. METHODS: Eight healthy male volunteer cyclists performed two identical bouts of square wave cycle ergometry, from a VO(2) of 60% of the lactic acid threshold (LAT) to 30% of the difference between LAT and VO(2) peak. Predetermined intervals (3--6 and 3--10 min) were chosen to reflect those often used in the literature, namely 3-6 min and 3 min to the end of exercise. Several procedures were used to estimate the 3, 6, and 10-min VO(2) values (20-s averaging, 60-s averaging, and mono-exponential modeling). These were compared with the modeled slow component amplitude using a two-phase model with independent time delays: VO(2)(t) = B VO(2) + A(1)(1 -- e(-(t-TD1)/tau(1)) + A(2)(1 -- e(-(t-TD2)/tau(2)). CONCLUSIONS: The results showed a significant underestimation for all methods of slow component amplitude estimation (P < 0.05) when compared with the actual (modeled) amplitude. In so far as research on oxygen uptake kinetics is used to understand the underlying physiology, it is imperative that the components of the kinetics be determined accurately. The use of a predetermined time frame for estimation of the amplitude of the slow component is not supported by this study. Future investigations should consider these results and make every effort to model the underlying response. PMID- 11283449 TI - What regulates exercise-induced reactive oxidant generation: mitochondrial O(2) flux or PO(2)? PMID- 11283450 TI - [Adult medulloblastoma. Review of 22 patients]. AB - BACKGROUND AND PURPOSE: Medulloblastoma is a malignant neuro-ectodermal tumor classically considered as a pediatric tumor. Adult medulloblastoma is rare. This low incidence results in a lack of data concerning the management of treatment. We report our experience and propose a review of the literature to clarify the main therapeutic options that are nowadays suggested. METHODS: We reviewed 22 adult patients treated for cerebellar medulloblastoma between 1979 and 1999. Actuarial relapse-free and overall survival were determined by the Kaplan-Meier method. Prognosis factors were studied by Log- Rank test. RESULTS: The five years relapse free and overall survival rates were respectively 63.1% and 81.3%. These rates are superior to those reported in the literature. None of the studied factors (age, gender, histological subtype, total or partial surgery, presence of a CSF derivation device, radiotherapy, chemotherapy) were significantly associated to remission or survival. However our statistical results should be interpreted with caution in this small population. CONCLUSION: Adult medulloblastoma prognosis seems to improve since chemotherapy has been introduced in the therapeutic protocols. Prospective and multicentric studies should determine the exact pattern of treatment. PMID- 11283452 TI - [Lumbar interbody fusion with threaded titanium cages. Results on 222 cases]. AB - Retrospective analysis of 222 cases of degenerative disc disease treated by threaded cage fusion. The objective was to determine the safety and efficiency of lumbar interbody fusions using screwed titanium cages and autogenous bone. Two hundred twenty-two patients had lumbar fusion at 243 levels between L2 and S1, at one or two disc spaces. Main indication was discogenic back pain with radicular leg radiation in degenerative discopathy complicated by disc protusion, segmental canal stenosis with chronic instability or spondylolysthesis of the first degree. Previous failed surgery after discectomy, nonunion or biologically cured discitis were other indications in selected cases. Results were classified as good to excellent in 80%, 15% improved but remained disabled, 5% had minimal or no improvement. Fusion rate was 91% at one year and 96% at 2 years. Peroperative dural tears occurred in 10 patients and transient neurological deficits in 9. A superficial infection occurred in one patient. Reoperation in the first three months included a cage revision in one patient and a foraminotomy in another. Two osteoporotic women needed an additional posterior fixation for kyphotic deformity. In conclusion, lumbar interbody fusion with threaded titanium cages appears to be efficacious with an acceptable rate of complications. Experience up to 7 years confirms that impression. Long term observation is needed before recommending this new method. PMID- 11283451 TI - [Neurosurgical treatment of hyperactive bladder in spinal cord injury patients]. AB - OBJECTIVES: We report long-term results of posterior sacral root rhizotomies in combination with Finetech-Brindley anterior sacral root stimulators implanted intradurally in 20 spinal cord injury patients. MATERIAL: and methods: The 14 female and 6 male patients included 14 paraplegics and 6 tetraplegics. All of them initially presented hyperactive bladder, detrusor-sphincter dyssynergia, recurrent urinary tract infection and performed (self) intermittent catheterization. Prior to implantation, an intrathecal test using bupivacaine was performed to confirm the compliances of the bladder. The main indication for implantation was persistent urinary incontinence refractory to medical therapy. RESULTS: After implantation the mean follow-up was 4,5 years. In all, 18 patients used the stimulator alone for bladder emptying and 18 patients were completely continent. The mean bladder capacity increased from 190 ml preoperatively to 460 ml after the operation. The mean residual urinary volume was reduced from 90 ml to 25 ml. No changes were noted by renal isotopic scanning in upper urinary tracts of patients. In 1 patient, a second extradural implant was performed. DISCUSSION: This article also include an overview of a) the different available sites where application of electrical stimulation results in a detrusor contraction, b) the benefits and disadvantages of the sacral posterior rhizotomy, c) selective stimulation techniques that allow selective detrusor activation by sacral root stimulation. CONCLUSION: Sacral anterior root stimulation combined with sacral posterior rhizotomy is a valuable method to restore bladder functions in spinal cord injured patients suffering from hyperactive bladder refractory to medical therapy. PMID- 11283453 TI - [Discal prostheses and arthrodesis for degenerative lumbar spine disease]. PMID- 11283454 TI - [Extraosseous Ewing's sarcoma of the spinal epidural space]. AB - Ewing's sarcoma is found exceptionally as a primary epidural tumor of the spine. Four cases of extraosseous Ewing's sarcoma of the spinal epidural space are presented. Another 17 cases were found in the pertinent literature and are discussed. There were 15 males and 6 females with mean age of 16 years (range, 4 to 30). Symptoms included back pain and/or radicular pain, paresis of one or more limbs, sensory disturbances, and bladder and bowel dysfunction. The mean diagnostic delay was 4,7 months. Each patient underwent surgery but complete resection of the tumor was obtained in only half of the cases. Most patients received radiation therapy and chemotherapy. Twelve patients died, 1 to 54 months (mean, 18) after diagnosis. Even though the number of patients was too small in this series for statistical analysis, partially resected tumors do appear to be associated with a higher mortality. PMID- 11283456 TI - [Atlanto-axial lateral mass osteoarthritis. Three case reports and review of the literature]. AB - This report describes three cases of symptomatic unilateral C1-C2 mass osteoarthritis. The patients were all female aged 67, 62 and 59 years. One patient had a history of rheumatoid arthritis. Unilateral occipital pain was the main symptom. Neuroradiological work-up included open mouth anteroposterior radiograph, flexion/extension lateral radiograph of the cervical spine, CT scan with coronal and sagittal reconstruction and MRI. Findings allowed differential diagnosis with tumoral or infectious disease of upper cervical spine and ruled out C1-C2 instability. CT scan and MRI permitted precise operative planning by determining the course of vertebral artery in the C1 and C2 vertebrae. Two patients were treated by CT scan guided steroid injection. The third patient was treated by C1-C2 arthodesis after failure of conservative treatment. PMID- 11283455 TI - [Vertebral osteoblastoma and scoliosis. Two cases report]. AB - We report two cases of spinal osteoblastoma in two boys aged 16 and 19 years. The lesion was disclosed by scoliosis with signs of thoracic and lumbar neurological compression. The diagnosis was provided by the CT scan and magnetic resonance imaging and was confirmed by the histology study of the surgical specimen. Involvement of the vertebral column has been estimated to range from 30 to 40% for these rare tumors that account for less than 1% of all bone tumors. Localization on the convex aspect of scoliosis is rare. CT-scan provides an analysis of the tumor components and clearly demonstrates intraspinal extension. MRI is superior in visualizing neurological compression. In our experience, function outcome has been favorable after surgical decompression. PMID- 11283457 TI - [Intracavernous epidermoid cyst. Case report and review of the literature]. AB - We report an unusual case of epidermoid cyst located in the cavernous sinus. Only two cases of epidermoid cyst of the cavernous sinus have been reported in the literature. In 1995, a 61-year-old man experienced diplopia, which resolved spontaneously. Imaging findings, particularly magnetic resonance, showed a cavernous sinus tumor. The MR appearance was compatible with epidermoid cyst. Since the patient was, at the time, asymptomatic, we proposed clinical and MR surveillance. In 1999, he developed a right III palsy with V1 and V2 hypoesthesia. The size of the cavernous tumor had increased on the control MR and the patient was operated via an extradural temporopterional approach through the Dolenc lateral triangle. We used a neuroendoscope to perform total resection of the tumor cyst. The extradural approach to the cavernous sinus and use of a neuroendoscope allowed complete removal of the cyst and shortened postoperative care. The patient was discharged on day 5 after surgery. These techniques provided a good view of the tumor without orbitozygomatic or zygomatic osteotomy. The 30 neuroendoscope allowed visual control of tumor removal that was better than a direct microscope view. PMID- 11283459 TI - [Suboccipital tuberculosis: a case report]. AB - Suboccipital tuberculosis is an uncommon localization of Pott's disease. The gravity results from the neurological and life threatening risk. We report a case of suboccipital tuberculosis in a 22-year woman who survived. She was given an anti-tuberculosis antibiotic regimen due to pulmonary and pericardal involvement. The patient interrupted her treatment after four months and was admitted six months later for torticolis and spastic tetraplegia without sphincter disorders. Standard x-rays and MRI of the head confirmed suboccipital Pott's disease. Transcranial evacuation was performed and the patient was again given anti tuberculosis antibiotics. The clinical course was favorable with definitive recovery 45 days later. The patient continued the antibiotic regimen for nine months. An orthopedic supporting device was worn for nine months. The diagnosis of suboccipital tuberculosis can be confirmed on MRI. Appropriate treatment is a subject of debate between exclusive orthopedic or combined orthopedic and surgical treatment. Prognosis depends on the neurological deficit, early diagnosis and prompt treatment. PMID- 11283458 TI - Sphenoidal metastasis from prostate carcinoma. Case report and review of the literature. AB - A case of an uncommon sphenoidal metastasis from prostate carcinoma with cranial nerve involvement is described. Current concepts of metastatic spread of this tumor to the skull base, clinical signs and therapeutic approaches are reviewed in the light of the available literature. PMID- 11283460 TI - [Difficulty in treating spinal chordoma of the thoracic vertebrae]. AB - We report a case of iterative surgery of a spinal chordoma of the 10(th) thoracic vertebra. This kind of neoplasm constitutes 3 to 7% of primary malignant bone tumors. Approximately 50% originate in the sacrum, 35% at the base of the skull and 15% in the true vertebrae. The slow growth delays diagnosis and compromises effective surgical therapy. On the basis of a review of the literature, we advocate an aggressive surgical resection from the beginning, similar to management of solitary vertebral metastasis. PMID- 11283461 TI - [Amyotrophic lateral sclerosis and cognitive disorders: review and analysis of the literature]. AB - In the last ten years, the syndromic nature of amyotrophic lateral sclerosis (ALS) has become more accepted. Together with upper and lower motor neuron signs, sensory or cognitive impairment are not uncommon. The frequency of a multidegenerative profile in ALS with SOD1 mutations is also an argument for this. We reviewed the literature about: PMID- 11283462 TI - [Pregnancy and epilepsy]. AB - Pregnant women with epilepsy risk complications. Perinatal mortality and congenital malformations are more frequent than in the general population. Offspring of parents with epilepsy have a higher risk for seizure later in life. Identification of factors associated with complications may lead to improved pregnancy outcome. Recommendations about neurological, obstetric and pediatric care of pregnant women with epilepsy are proposed. PMID- 11283463 TI - [Noncognitive symptoms in Alzheimer's disease. A study of 150 community-dwelling patients using a questionnaire completed by the caregiver]. AB - We studied the noncognitive symptoms in 150 community-dwelling Alzheimer's patients using a questionnaire completed by the caregiver, the Echelle Psychopathologique de la Demence de Type Alzheimer, EPDTA (Psychopathologic Scale of Dementia of Alzheimer Type). EPDTA is a 44-item questionnaire derived from the BEHAVE-AD and the Depressive Mood Scale, covering many aspects of the behavior, affective and psychiatric disturbances. Each item is rated from 0 (never observed) to 6 (most of the time). Frequency (percentage of symptom present) and severity (mean score when the symptom was present) were assessed for each item. The cognitive status and severity of the disease were assessed by the MMSE and two scales completed by the caregiver assessing the Activities of Daily Living scale (ADL) and the Cognitive Difficulties Scale (CDS). Noncognitive symptoms were present in all patients but remained moderate in severity. A principal component analysis of the 33 items exploring the affective disturbances showed seven clinically relevant factors: apathy, anxiety, anosognosia-irritability, euphoria, dysphoria, emotional incontinence and agitation. The most frequent noncognitive symptoms were the affective disturbances, especially apathy, and the sexual behavioural disturbances. No correlation were found between the overall severity of behavioural disturbances and cognitive status, duration of the disease nor demographic variables. However, a slight negative correlation was found between scores on apathy and on the MMSE. A second evaluation was performed in 59 patients after a mean follow-up of 18,2 months. The patients showed a progression of the disease evidenced by the scores on the MMSE, ADL and CDS scales. However, the frequency and severity of the noncognitive symptoms remained identical except for eating disorders, psychotic symptoms and agitation which were more frequent at the second examination and negatively correlated with the MMSE score. Most patients showed affective disturbances and scored high for apathy and anxiety-emotional incontinence dimensions. Like in a previous study, we found a double dissociation between these two dimensions in some patients, suggesting that they depend from different mechanisms. Agressivity, mostly verbal, was found in three quarters of the patients and was correlated to apathy, anosognosia and psychotic symptoms. CONCLUSION: The relationship between noncognitive manifestations and cognitive deficits in AD is not clear, suggesting that they depend from different biological and psychological mechanisms. Various dimensions may be described in the behavioural disturbances but their relationship with hypothetical biological mechanisms remains difficult. Our study stresses the importance of apathy, which was corelated with various noncognitive psychobehavioral manifestations in AD patients. PMID- 11283465 TI - [Cerebral blood flow disturbances after anterior choroidal artery infarcts. Anatomical and functional correlates]. AB - We have investigated the cortical and subcortical regional cerebral blood flow (rCBF) disorders resulting from infarcts of the anterior choroidal artery (AChA), and correlations with the severity of lesions, the physical and cognitive deficits, and the functional impairment. Eighteen patients presenting with recent anterior choroidal artery infarct without any other brain injury were examined at the secondary phase post-stroke using the single photon emission computed tomography technique and 133 Xenon inhalation. The rCBF and asymmetry indexes (AI) were calculated for 12 symmetrical hemispheric areas, and the cerebellum. The AI values were compared with those of 24 control subjects. The severity of the lesions was evaluated from CT scans or MRI. The neurological status (Orgogozo scale, walking disorders, MMSE, attention impairment, aphasia) and disability (functional independance measure: FIM) were assessed for each patient at the same time period. The relationships between rCBF disorders and brain lesions, and between the results of clinical investigations and rCBF disorders and brain lesions were assessed by linear regression analyses (stepwise variable selections, p=0.05). The AI values were significantly increased in the cerebral hemispheres, and this was most severe in the internal capsule (direct effect of the lesion) and the dorsolateral hemispheric cortex (diaschisis). Individual evaluations showed that AI were significantly increased in 13 patients in at least one ROI of the cerebral hemispheres, and in 3 patients in the internal capsule. Stepwise variable selections revealed that AI were best explained by the severity of the lesions in the internal capsule and the internal temporal area. The AI of the external temporal area and the internal capsule also helped explain the clinical (physical and cognitive) deficits. Thus, AChA infarcts may have relatively large effects on the central part of the lateral and dorsal cortex of the ipsilateral hemisphere. Subcortical and cortical consequences both contribute to explain the motor and cognitive deficits and disability. PMID- 11283464 TI - [Familial orthochromatic leukodystrophy: clinicopathological study of two cases]. AB - This paper reports the clinico-pathological data in a French family with orthochromatic leukodystrophy. The parents were first cousins and had seven children. Among those, two sisters and one brother presented with neurological signs, with onset around the 5(th) decade, including a dementing syndrome of frontal type, a tetrapyramidal syndrome, seizures, and, in one sibling, a cerebellar syndrome. CT scan or MRI showed diffuse involvement of the white matter. The neurological signs worsened progressively leading to death within 11 and 22 months. Neuropathological examination was performed in two cases. It revealed characteristic orthochromatic leukodystrophy. In one case, the presence of pigmented macrophages and astrocytes was suggestive of Van Bogaert and Nyssen disease. However there were some atypical features including the absence of pigmented cells in the second case whose clinical course was shorter, and the cavitary appearance of the white matter changes with a relative increase in the number of oligodendrocytes raising the issue of a possible link between this condition and cavitary orthochromatic leukodystrophies. PMID- 11283466 TI - [Otolith manifestations in Wallenberg syndrome]. AB - Central vestibular pathways issuing from the otolith may be involved in Wallenberg syndrome, resulting in specific symptoms. These "otolith" symptoms are less well known than vestibular symptoms issuing from the canal. We report 15 patients with Wallenberg syndrome who had one or more clinical signs suggestive of otolith pathway involvement. Clinical examination looked for: Eleven patients presented ipsilesional skew deviation; ipsilesional tilt of subjective visual vertical was found in 8 patients; a room tilt illusion was described in 4 patients in either the frontal or the sagittal plane: 9 patients presented axial lateropulsion toward the lesioned side; 8 patients presented ipsilesional ocular lateropulsion, 6 of them in association with axial lateropulsion. Finally, 3 patients presented positional nystagmus evoked by head tilt in the roll or the pitch plane. Pathophysiology of these symptoms and evidence for an otolith pathway involvement are discussed. PMID- 11283467 TI - [Proximal myotonial myopathy (PROMM): clinical and histology study]. AB - We report 13 French patients with proximal myotonic myopathy. PROMM is a recently delineated multisystem disorder with dystrophic myopathy, myotonia and cataracts. This syndrome is genetically distinct from myotonic dystrophy (DM) by the absence of abnormal CTG repeat expansion. The geographical origin varies but 4 families originated from Poland. Of late onset, muscle weakness is diffuse and predominantly affected proximal and axial muscles. Facial involvement and myotonia were moderate or absent, but in all cases myotonic discharges were detected on EMG. 6 patients suffered from myalgia. Cataracts occurred in 11 patients, mainly indistinguishable from those in DM. Cardiac arrythmia occurred in 7 patients. Muscle biopsy revealed rare structural changes of the muscle fibers and selective type I atrophy, common in DM, could not be found on morphometric analysis. PROMM has a distinct clinical spectrum from DM which includes a predominantly proximal muscle weakness, with troubling pain, a more favourable prognosis and a different histopathological pattern. PMID- 11283468 TI - [Subacute neuropathy, multiple cancers, and Gougerot-Sjogren syndrome: contribution of anti-Hu antibody]. AB - We report the case of a patient presenting a subacute, predominantly sensory neuropathy. The work up revealed a Sjogren's syndrome and a breast carcinoma. The presence of anti-Hu antibodies, identified by Western Blot using purified recombinant HuD protein, and the absence of the Hu antigen in the breast carcinoma ruled out the responsibility of the Sjogren's syndrome or breast carcinoma. In this context, the most likely diagnosis was a subacute neuropathy associated with small cell lung cancer, which was indeed discovered 3 years later. PMID- 11283469 TI - [Cytomegalovirus rhombencephalomyelitis in an immunocompetent subject]. AB - We report the clinical and MRI findings of a 31-year-old healthy immuno-competent patient who presented following 48 hours of diffuse headache with progressive and severe rhombencephalomyelitis signs. Cerebral and medullar MRI confirmed the central nervous system improvement. Serology showed relation with cytomegalovirus infection. Spontaneous improvement was observed and favoured by ganciclovir administration. PMID- 11283470 TI - [Parkinsonian syndrome, dystonia and cognitive disorders in a 67-year-old man]. PMID- 11283471 TI - [Neurological history - Charles-Edouard Brown-Sequard]. PMID- 11283472 TI - Prozac weekly. PMID- 11283473 TI - Detrol LA and Diropan XL for overactive bladder. PMID- 11283474 TI - Nateglinide for type 2 diabetes. PMID- 11283475 TI - [Extended total gastrectomy: indications in the 3rd millennium]. AB - BACKGROUND: Total extended gastrectomy (TEG) is indicated in the treatment of gastric cancer for necessity or to achieve an oncologic radicality. By this surgical treatment the stomach and other organs or a part of them involved by primitive tumor are removed. METHODS: The authors report a study about 15 patients, out of 116 cases of gastric cancer, operated by TEG between 1990-1998. The middle-age of this patients was 63 years (range 45-76) and their general conditions were good in 9 cases and not-good in 6. The postoperative total parenteral nutrition (TPN) was carried out in all the patients, while preoperatively only in the most compromised patients. The surgical treatments were: 2 TG (total gastrectomy)+splenecomy; 3 TG+splenectomy+pancreatic resection; 4 TG+splenectomy+pancreatic resection+distal esophageal resection; 1 TG+distal esophageal resection; 2 TG+atypic hepatic resection; 1 TG+ atypic hepatic resection+duodenum resection; 2 TG+large intestine resection. While 10 patients were operated on to obtain radicality, 5 patients had a palliative treatment. RESULTS: There was not perioperative mortality, but we have observed: one dehiscence of the duodenal stump and one pancreatic fistula treated with conservative therapy; one left subfrenic abscess treated with surgical therapy. The survival has been higher in the patients treated with radicality. On the basis of these cases, the authors consider: 1) the possibility to obtain radicality by TEG; 2) the gastric localizations more often associated to extravisceral neoplastic localization; 3) the role of extensive lymph node resection (III and IV level) to obtain oncological radicality or neoplastic reduction. CONCLUSIONS. On the basis of their personal experience and related literature, the authors conclude that TEG is indicated to: 1) obtain a better lymphadenectomy; 2) obtain an oncologic radicality; 3) reduce the neoplastic mass in order to facilitate adjuvant therapy; 4) avoid or treat neoplastic complications; 5) improve the quality of life. PMID- 11283476 TI - [Current diagnosis of gastroesophageal reflux disease: learning experience]. AB - The authors report their study on gastro-esophageal reflux disease, a pathology that has become increasingly common over the past years reflecting both a real increase and the use of new and more sophisticated and reliable diagnostic methods and tests. It can be included in the group of pathologies absorbing the largest proportion of financial resources, even exceeding biliary lithiasic disease according to American studies. The authors start by analysing the symptoms of gastroesophageal reflux disease, drawing a distinction between typical (heartburn, epigastric pain and postprandial regurgitation) and atypical symptoms (laryngotracheal symptoms, bronchopulmonary symptoms and esophageal motor incoordination). They outline the diagnostic iter and tests most widely used today to achieve a correct diagnosis. Lastly, they report their experience of 160 patients attending their esophageal diagnostic unit since January 1999 who underwent a number of different instrumental tests, the results of which are compared. Three different aspects are compared: the presence of symptoms, 24-hour pH-metry and endoscopic tests. All these are necessary for a correct diagnosis of gastroesophageal reflux disease and to evaluate the possibility and efficacy of surgery. They emphasise the diagnostic importance of 24-hour pH-measurement as the only test that can directly reveal gastroesophageal reflux. Positive pH results represent a discriminating element in deciding whether the patient should undergo surgery. PMID- 11283477 TI - [Laparoscopic appendicectomy: an 8-year clinical experience]. AB - BACKGROUND: For more than a century, open appendectomy through a laparotomy has been the golden standard for the surgical removal of the appendix. Nowadays, many surgeons question the utility of laparoscopic surgery to perform appendectomies because it is commonly stated that the appendix can be removed through a small surgical incision carrying a minimal surgical trauma to the patient. Although open appendectomy is really safe, on the other hand it carries a considerable risk of postoperative complications, is associated with postoperative pain and affects patient s normal activity. Laparoscopic appendectomy was first described in 1983 and, in many studies, it is described to be better than open standard technique for the treatment of appendiceal diseases. The aim of the present study is the retrospective analysis of laparoscopic appendectomies performed in a 8 year period. METHODS: The authors report on 129 patients who underwent laparoscopic appendectomy. RESULTS: Conversion rate was 0.7 %, while the laparoscopic procedure was completed in 96 female and 32 male patients. The position of the appendix was behind the cecum in 37 cases, associate diseases were found in 15 cases. Mean operative time was 51 minutes; kind of laparoscopic instrumentation affected the operation time. Histologically there were 71 (55.5 %) focal appendicitis, 22 (17.1 %) suppurative appendicitis, 11 (8.6 %) gangrenous appendicitis, 18 (14.1 %) chronic appendicitis showing signs of previous suppurative episodes and 6 (4.7 %) normal appendix. There were neither in-hospital morbidity nor mortality. Follow-up showed reduced postoperative pain, short hospital stay, fast return to complete social activity. CONCLUSIONS: The authors conclude that laparoscopic technique can be considered a safe and effective procedure for the removal of the appendix as it has the advantage of allowing faster postoperative recovery; moreover the author recommend a wider and routinely use for appendectomy. PMID- 11283478 TI - [Gastroduodenal reflux after laparotomic or laparoscopic cholecystectomy]. AB - BACKGROUND: The authors analyse gastroduodenal reflux (GDR) in the light of the progress made over the past ten years. The good results achieved using mini invasive techniques in cholecystectomy prompted the authors to compare laparotomic and laparoscopic cholecystectomy in order to evaluate the influence of both techniques in determining GDR and clinical symptoms. METHODS: Symptoms were evaluated before and after surgery in 30 patients operated using a laparotomic technique and in 30 patients operated using a laparoscopic technique. Two groups of 10 patients, operated respectively using laparotomic and laparoscopic techniques, were studied both clinically and endoscopically before and after surgery. RESULTS: The analysis of the results shows a lower incidence of GDR and typical symptoms of GDR in patients undergoing laparoscopic surgery. The rationale underlying the lower incidence of GDR and its symptoms in these patients compared to the more conventional group is not completely clear. CONCLUSIONS: These results may be influenced by reduced surgical trauma and the careful selection of case or the laparoscopic technique, hence the exclusion of cases complicated by severe cholecystitis. PMID- 11283480 TI - [Surgical treatment of hemorrhoid disease. A comparison between techniques]. AB - BACKGROUND: The past years have seen the development of outpatient treatment and surgical procedures for the treatment of hemorrhoid disease in an attempt to reduce postoperative pain on the one hand and execution and hospitalisation times on the other. METHODS: This retrospective study compares the results obtained in the treatment of hemorrhoid disease using three different METHODS. Three groups of 30 patients were selected from those operated during the past 10 years using Milligan-Morgan s technique (Group A), Parks technique (Group B) and radiosurgery (Group C). All patients were matched for sex (50% males and 50% females), age (ranging between 30 and 50 years old), diagnosis (4th degree hemorrhoids not associated with another proctologic pathology such as anal fissures, fistulas, etc.) and referred symptoms. RESULTS: The results obtained allow a positive evaluation to be made of all three surgical techniques, but indicate submucous hemorrhoidectomy using radiosurgery as the method of choice for the treatment of grade III and IV hemorrhoids. CONCLUSIONS: The excellent results achieved using this approach in the authors opinion amply justify the relatively difficult execution and longer operating times. PMID- 11283479 TI - [A new conservative approach in the treatment of postoperative digestive-tract fistulas. Mechanical closure by a balloon-catheter]. AB - BACKGROUND: Digestive fistulas represent troublesome complication in patients operated in modern surgical wards where the improved surgical procedures and better intensive care enhance the surgeon to perform more aggressive approaches with a high surgical risk index. The management of a patient presenting a digestive-tract fistula is never easy, being its approach either conservative (TPN) or surgical. We applied an alternative surgical procedure consisting in a mechanical closure of the fistula using a balloon-catheter so as to improve outcome in those patients in whom medical tratment did not show satisfactory RESULTS. METHODS: We treated 7 patients presenting a postoperative fistula following several surgical procedures for neoplasms of the digestive system. These fistulas were closed using a Foley or Fogarthy balloon catheter preceeded by radiological and/or endoscopy controls. Once the catheter was placed, oral nutrition was started and some patients were discharged. A progressive deflation of the balloon was performed until complete removal of the catheter upon approx 10 days. RESULTS: We obtained a complete healing of the fistula in 6 patients, within 10 days since catheter placement. Only one patient required another operation. CONCLUSIONS: Our case-series may seem statistically not significant, but varied concerning location and type of fistulas. We observed an excellent outcome using this procedure which allows very short healing period thanks to an early oral nutrition uptake and a decrease in costs mainly due to a short hospital stay and a minor use of expensive drugs (TPN). PMID- 11283481 TI - [Influence of acute anemia and hemodilution on wound healing. Experimental study in rats]. AB - BACKGROUND: An experimental study was conducted on rats in order to determine the effects of acute anemia and hemodilution on the cicatriztion of the abdominal wall. METHODS: Forty two Wistar rats were divided into 3 groups: control, anemic and hemodiluted. Acute anemia was promoted by removing 3 ml of blood/100 g animal body weight. An equal volume of isotonic saline was injected into animals submitted to hemodilution. The evolution of the abdominal scar was determined at 7 and 14 days by measuring rupture tension and collagen concentration. The resistance gain was similar in all three groups at the level of the skin scars. RESULTS: In contrast, in the scars of the peritoneum-muscle-aponevrotic plane, resistance was similar in the anemic and hemodiluted rats throughout the study period, but when the resistance of the scars of these groups was compared to that of the control on the 7th day it was found to be lower (anemic rats, p=0.0360; hemodiluted rats, p=0.0270). The same was observed on the 14th day, when anemic and hemodiluted rats presented less resistant scars than the controls (p=0.0270). The collagen concentration in the skin scars was lower in the anemic group than in the hemodiluted group on the 7th day, but the difference was nonsignificant when compared to that of the control group. On the 14th day, control and hemodiluted rats had a higher collagen concentration than anemic rats (p=0.0020 and p=0.0390). On 14th day the collagen concentration were lower in the scar peritoneum-muscle-aponevrotic (p<0.0001). CONCLUSIONS: Thus, under the conditions of the experiment, the skin scar did not show a change in resistance although anemic rats had a lower collagen concentration throughout the study period, and the peritoneum-muscle-aponevrotic scars showed a change in resistance on the 14th day and presented lower collagen concentration in anemic rats. PMID- 11283482 TI - [Endoscopic surveillance of precancerous changes in the stomach]. AB - Undoubtedly, one of the most important achievements of gastroenterology is the demonstration that, for many pathological conditions with future neoplastic degeneration risk, a periodical endoscopic surveillance is a determining element for the restraining of possible evolutive complications. Nonetheless, it is to be considered how, during the last years, the prevention and follow-up procedures for the stomach disease have been sometimes emphasized. In fact, various recent evidences originated from precise scientific evaluations have contained same prevention strategic attitudes so as to reach the best cost-benefit ratio. In this survey the various gastric preneoplastic modifications, subdivided in precancerous conditions and lesions are examined and guidelines are proposed in order to offer quick and easily applicable solutions. PMID- 11283483 TI - Lymphadenectomy in well differentiated thyroid carcinoma. AB - In this review, the authors analyze, in relation to the data collected in other literature, the indications and the type of surgical procedure to perform on the neck lymph nodes in cases of differentiated thyroid carcinoma. The authors stress the fact that the surgical procedure must be determined according to the stage in which the diagnosis and prognostic factors are formed and in relation to the natural history of these tumors. PMID- 11283484 TI - [Biliary complications during videolaparoscopic cholecystectomy. Remarks on methodology and indications in the training period]. AB - In our study we have considered the activity of a surgeon working in our Surgery Department during his laparoscopic training period. We focus our attention on a date related to the same complications checked in 27 cases of cholelithiasis operated by the same surgeon. We have observed three cases of biliary cholelithiasis fistulas, all of them during the three last operations. The examination of the above mentioned cases considers the clinical post-surgery situation and the therapeutic standards we adopted to work out the complications. Now we can precisely state, according to our experience, the particular directions for the videolaparoscopic training period. In addition we can propose one simple mathematical formula to value the IRL (Laparoscopic Risk Index) concerning three variables: the experience of the surgeon; the instruments condition; the clinical situation of the patient. The relation of these three factors turned in numbers suggests the chance of success of a videolaparoscopic operation. We conclude our study mentioning the gasless videolaparoscopic technique that seems to have a large indication, according to the same Surgery School. This technique is particulary indicated on those classes of patients in which the CO2 insufflation into abdominal cavity and the increase of endocavity pressure can represent a contraindication to the videolaparoscopic approach: in this case the surgeon will follow the surgical indication to solve the clinical situation, as happens in war surgery. PMID- 11283485 TI - [Peristomal laparocele: particular indications for the use of prosthetic materials]. AB - The authors report the case of a female patient suffering from colic neoplasm and a vast peristomal laparocele, the long-term outcome of a rectal amputation that the patient underwent at a young age. After colic resection, a vast area of surgical mesh in dacron was modelled and positioned in a retromuscular scat, thus allowing the hernia pathology to be resolved relatively simply, owing to its particular location-in correspondence with a preternatural anus-until not long ago this would have represented an absolute contraindication to the use of prosthetic materials. A precise surgical approach, marked by a scrupulous respect for aseptic conditions, is essential in these circumstances owing to the persistent risk of septic complications. PMID- 11283486 TI - [Littoral hemangioma of the spleen]. AB - The authors report a rare case of littoral hemangioma of the spleen (LHS) accompanied by a revision of the literature on the argument. A male 65-year-old patient was referred to their attention with suspected ultrasonographic diagnosis of lymphoma with a splenic localisation. The complete CT diagnosis led to suspected splenic angioma. During surgery, anatomopathological analysis of the biopsy revealed LHS. The pathological anatomy showed lesions ranging in size from small foci to large nodules which almost completely replaced the splenic parenchyma. These areas were made up of vascular canals or axes that imitate splenic sinuses and have irregular lumen, often appearing as papillary projections and cyst-like spaces; they are bordered by high (cylindrical) endothelial cells that project into the vascular lumen and reveal hemophagocytosis; there is very little mitotic activity. The patient was discharged 7 days after surgery. The authors underline the extreme rarity of this neoplasm and the virtual absence of symptoms, although some cases report signs of hypersplenism, including platelet deficiency and anemia. The diagnostic iter must take care to exclude other pathologies affecting the spleen, including lymphoma, metastases and primary malignant splenic tumours. Lastly, a differential diagnosis must be made with the malignant variant, littoral hemangiosarcoma of the spleen. PMID- 11283487 TI - [Carotid body tumour. Case report and literature review]. AB - The tumour of the carotid body is rare. About 1000 cases had been reported in the literature. It may occur sporadically in 90% of cases and it affects both sexes in the same proportion and in the middle age. This tumour may be misdiagnosed if it is not suspected. Ultrasono-graphy and color-Doppler scan show a hypervascular tumour between the internal and external carotid arteries. CT-scan defines the tumour s extent on the surrounding structures. Angiography is the gold standard for diagnosis, showing a hypervascular mass displacing the bifurcation of the carotid arteries. Sometimes radiotherapy and embolization are indicated but the surgical excision of carotid body tumours is the therapy of choice. The surgical approach through incision like carotid artery operation is performed. If the subadventitial plane between tumour and arterial wall is not identified, resection of carotid artery and insertion of a shunt is required. Although the diagnosis and the surgical technique advances, the incidence of postoperative nerve injury is high in the different series. The clinical suspect and the early diagnosis are very important because low morbidity rate occurs with resection of a small chemodectoma. The surgical excision can be followed by postoperative respiratory depression or dyspnea both with regional and general anesthesia. The authors report a case of a medium size tumour operated on and developing a mild transient weakness of cranial nerve VII. Recent trends in evaluation and therapy are analysed and the literature is reviewed. PMID- 11283488 TI - [The use of an underlay polypropylene mesh in complicated incisional hernias: sucessful French surgical technique]. AB - BACKGROUND: Incisional hernia repair with conventional techniques is associated with high recurrence rates of 30-50%. Surgical repair using different prosthetic biomaterials is becoming increasingly popular. On the basis of the favourable results by French surgeons, the results of underlay prosthetic mesh repair using polypropylene mesh in complicated and recurrent incisional hernias have been studied. METHODS: After preparation and excision of the entire hernia sac, the peritoneum and posterior rectus sheath are closed with a continuous looped polyglyconate suture. The prosthesis used for the midline hernias is positioned on the posterior rectus sheath and extends far beyond the borders of the myoaponeurotic defect. The prosthesis for lumbar and subcostal hernias is placed in a prepared space between the transverse and oblique muscles. Intraperitoneal placement of the mesh must be avoided. Between January 1997 and September 1998 a total of 57 incisional hernia repair (25 primary hernias, 32 recurrent hernias) have been performed using this technique (28 women, 29 men, mean age 56+/-13 years). RESULTS: Local complications occurred in 6 patients (11%). One patient suddenly died on the 3rd postoperative day from severe pulmonary embolism (mortality 1,7%). Thirthy-seven patients with a minimum follow-up of 6 months were reexamined clinically (follow up time 6-33 months). Till now one recurrent hernia has been observed. There were only minor complaints like a feeling of tension in the abdominal wall (n = 3) and slight pain under physical stress (n = 9). CONCLUSIONS: The aforementioned technique of underlay prosthetic repair allows an anatomical and consolidated reconstruction of the damaged abdominal wall with excellent results and low complication rates especially in high risk patients and complicated hernias. PMID- 11283489 TI - Meta-analysis of tympanostomy tube sequelae. AB - OBJECTIVE: To estimate the incidence of tympanostomy tube sequelae based on systematic review of published case series and randomized studies. DATA SOURCES: English-language MEDLINE search from 1966 through April 1999 with manual reference search of proceedings, articles, reports, and guidelines. STUDY SELECTION: Cohort studies with otitis media as the primary indication for tube placement. DATA EXTRACTION: Two reviewers independently extracted data from 134 articles. DATA SYNTHESIS: Transient otorrhea occurred in 16% of patients in the postoperative period and later in 26%; recurrent otorrhea occurred in 7.4% of patients and chronic otorrhea in 3.8%. Sequelae of indwelling tubes included obstruction (7% of ears), granulation tissue (5%), premature extrusion (3.9%), and medial displacement (0.5%). Sequelae after tube extrusion included tympanosclerosis (32%), focal atrophy (25%), retraction pocket (3.1%), cholesteatoma (0.7%), and perforation (2.2% with short-term tubes, 16.6% with long-term tubes). Meta-analysis showed that long-term tubes increased the relative risk of perforation by 3.5 (95% CI, 1.5 to 7.1) and cholesteatoma by 2.6 (95% CI, 1.5 to 4.4). Similarly, intubation increased the relative risk of tympanosclerosis by 3.5 (95% CI, 2.6 to 4.9) and focal atrophy by 1.7 (95% CI, 1.1 to 2.7) over nonintubated control ears (baseline tympanosclerosis and atrophy rates of 10% and 14%, respectively). CONCLUSIONS: Sequelae of tympanostomy tubes are common but are generally transient (otorrhea) or cosmetic (tympanosclerosis, focal atrophy). Nonetheless, the high incidence suggests a need for ongoing otologic surveillance of all patients with indwelling tubes and for a reasonable time period after tube extrusion. Long-term tubes should be used on a selective and individualized basis. PMID- 11283491 TI - John Conley lecture on medical ethics. Ethics and the mystery of wholeness. PMID- 11283492 TI - Cost and cost-effectiveness of universal screening for hearing loss in newborns. AB - OBJECTIVE: To estimate the cost and cost-effectiveness of universal newborn hearing screening. STUDY DESIGN AND SETTING: Decision analysis model utilizing the hospital perspective. This model evaluated 4 distinct protocols for screening a fixed and defined hypothetical cohort of newborn infants. OUTCOME MEASURES: Cost of screening and the number of infants with hearing loss identified through universal screening. RESULTS: Otoacoustic emissions testing at birth followed by repeat testing at follow up demonstrated the lowest cost ($13 per infant) and had the lowest cost-effectiveness ratio ($5100 per infant with hearing loss identified). Screening auditory brainstem evoked response testing at birth with no screening test at follow-up was the only protocol with greater effectiveness, but it also demonstrated the highest cost ($25 per infant) and highest cost effectiveness ratio ($9500 per infant with hearing loss identified). These findings were robust to sensitivity analysis, including best-case and worst-case estimation. The prevalence of hearing loss and the fraction of infants returned for follow-up testing had a large impact on the absolute level, but not relative level of protocol cost and cost-effectiveness. CONCLUSION: The otoacoustic emissions testing protocol should be selected by screening programs concerned with cost and cost-effectiveness, although there are certain caveats to consider. SIGNIFICANCE: The most significant barriers to implementation of universal newborn hearing screening programs have been financial, and this study compares the most common protocols currently in use. This study can assist program directors not only in the decision to initiate universal screening but also in their choice of screening protocol. PMID- 11283494 TI - A prospective observational study of 5-, 7-, and 10-day antibiotic treatment for acute otitis media. AB - OBJECTIVE: To compare 5-, 7- and 10-day duration of antibiotic therapy for acute otitis media (AOM) in children. STUDY DESIGN AND SETTING: Prospective nonrandomized 1-year evaluation of 3 treatment durations for AOM in a private pediatric setting. Outcomes assessed at 14 +/- 4 days after start of therapy with clinical response categorized as cure, improvement, or failure. RESULTS: A total of 2172 children were studied; 46.4% were < or =2-years-old. Antibiotics used were amoxicillin (61.9% of patients), trimethoprim/sulfamethoxazole (11.7%), cephalosporins (14.2%), amoxicillin/clavulanate (5.2%), and macrolides/azalides (4.8%). No overall difference in outcome was observed comparing the 5-day (n = 707), 7-day (n = 423), or 10-day (n = 1042) treatments, including children < or =2-years-old. However, in the subset who had an episode of AOM in the preceding month, outcome differed; 5-day treatment was followed by more frequent failure than 10-day treatment (P < 0.001). In logistic regression analysis, variables identified as contributing to a cure were: >2-years-old (P < 0.0001), no AOM in the preceding month (P = 0.07), or preceding 12 months (P = 0.03). CONCLUSIONS: Our study supports the transition from 10 to 5 days for standard AOM antibiotic treatment duration in most patients. A 10-day regimen may be superior in children who have experienced an episode of AOM within the preceding month, a known risk factor for resistant bacterial infection in the otitis-prone patient. PMID- 11283493 TI - Safety and efficacy of topical mitomycin C in myringotomy patency. AB - OBJECTIVE: To develop an alternative method for prolonged middle ear ventilation using topical mitomycin C. STUDY DESIGN AND SETTING: Twenty guinea pigs with normal ears had bilateral myringotomies performed using the argon laser. After myringotomy, either mitomycin C (0.4 mg/mL) or saline pledgets were applied topically. Monitoring consisted of otomicroscopy and distortion-product otoacoustic emissions. RESULTS: Before myringotomy, all tympanic membranes were intact, and distortion-product otoacoustic emissions were measurable. After myringotomy, none (0%) of the saline-treated myringotomies were patent at day 7 as compared with 100% of the mitomycin C-treated myringotomies. At day 42, 10 (52.6%) of 19 mitomycin-treated myringotomies remained patent and 4 (28.6%) of 14 were patent at 131 days. Five (13.1%) ears developed purulent otorrhea; 3 were mitomycin C-treated and 2 were treated with saline solution.- Distortion-product otoacoustic emissions testing did not document any evidence of ototoxicity. CONCLUSION: Topical mitomycin C appears to be safe and effective at prolonging the duration of myringotomy patency in the guinea pig. SIGNIFICANCE: Mitomycin C may be useful as an adjunct for preventing myringotomy closure. PMID- 11283495 TI - Dosage regimens of intranasal aerosolized surfactant on otitis media with effusion in an animal model. AB - OBJECTIVE: To determine optimal dosage regimens of intranasal metered dose aerosolized surfactant with and without other medications in the treatment of otitis media with effusion (OME). STUDY DESIGN: Resolution of experimental OME in gerbils was determined based on otomicroscopy and tympanometry. Experimental intranasal drugs were: surfactant, surfactant with betamethasone, surfactant with phenylephrine, and a normal saline solution placebo. Medications were administered once or twice daily via a metered dose inhaler. RESULTS: For twice daily dosing, mean days to OME resolution were 8.5 for the aerosolized surfactant, 6.3 for the surfactant with betamethasone, 18.7 for the surfactant with phenylephrine, and 16 each for control and placebo. Resolution with the once daily dosage was longer for all conditions. Results were comparable using tympanometry. CONCLUSION: OME resolved faster than the natural course when treated with intranasal surfactant with and without steroids. Twice-daily dosing was statistically superior. SIGNIFICANCE: This study reiterates the effectiveness of OME treatment with an aerosolized synthetic surfactant with and without steroids and establishes a superior twice-daily dosage schedule. PMID- 11283496 TI - Chronic tinnitus as phantom auditory pain. AB - OBJECTIVES: To investigate similarities between patients who experience chronic tinnitus or pain and to formulate treatment strategies that are likely to be effective for patients who experience phantom auditory pain. STUDY DESIGN: A total of 160 patients rated the severity and loudness of their tinnitus and completed the State-Trait Anxiety Inventory (STAI) and an abbreviated version of the Beck Depression Inventory (aBDI). Patients received counseling, audiometric testing, and matched the loudness of their tinnitus to sounds played through headphones. SETTING: A specialized tinnitus clinic within an urban medical center. RESULTS: Tinnitus severity was highly correlated with patients' degree of sleep disturbance, STAI, and aBDI scores. The self-rated (on a 1-to-10 scale)- but not the matched--loudness of tinnitus was correlated with tinnitus severity, sleep disturbance, STAI, and aBDI scores. CONCLUSIONS: The severity of chronic tinnitus is correlated with the severity of insomnia, anxiety, and depression. These relationships are the same for many patients with chronic pain. Treatment recommendations are discussed in reference to these results. PMID- 11283497 TI - Hemostatic alterations in patients with acute, unilateral vestibular paresis. AB - OBJECTIVES: The etiopathogenesis of acute unilateral peripheral vestibulopathy (APV) still remains a matter of debate; ischemic changes in the circulation of the labyrinth may play a role. We consequently looked for possible hemostasis alterations in a group of patients with APV of an unknown nature. METHODS: We evaluated blood parameters known to be involved in circulation disorders, including total and HDL cholesterol, triglycerides, apolipoprotein A and B, lipoprotein(a), homocysteine, folate, prothrombin time, activated partial thromboplastin time, fibrinogen, D-dimer, antithrombin III, protein C, protein S, activated protein C resistance, and anticardiolipin IgG and IgM antibodies. A series of 23 patients affected with APV were consecutively referred to our department, in the acute phase, before treatment, and in the follow-up phase after 4 to 6 weeks of pharmacologic washout. The aforementioned blood parameters were also measured in a series of 15 patients with Meniere's disease. RESULTS: The patients with APV in the acute phase compared with the patients with Meniere's disease in the acute phase exhibited increased plasma levels of fibrinogen (mean, 338.3 +/- 135.9 SD vs 271.3 +/- 69.8 SD mg/dL, P = 0.05), increased plasma levels of D-dimer (mean, 320 +/- 207.8 SD vs 226.7 +/- 138.7 SD NG/mL), enhanced plasma levels of lipoprotein(a) (41.4 +/- 38.6 SD vs 16 +/- 18.2 SD mg/dL, F = 5.67, P = 0.02), high leukocyte count (9.1 +/- 2.7 SD vs 6.5 +/- 1.3 SD x 10(3)/microL; F = 8.42, P < 0.006), and low serum folate concentration (5.3 +/- 1.8 SD vs 7.1 +/- 2.7 NG/mg; F = 4.34, P = 0.04). During follow-up the prothrombin time was prolonged (F = 4.34, P = 0.04) and leukocyte count decreased (F = 7.39, P < 0.019) in the APV patients, whereas fibrinogen, D-dimer, lipoprotein(a), and folate were unchanged. CONCLUSION: Our results provide evidence suggesting an involvement of the hemostatic system in APV. PMID- 11283498 TI - Posterior petrous face meningiomas. AB - OBJECTIVE: To define the clinical presentation, treatment options, and outcomes for a subset of meningiomas of the posterior fossa skull base that arise from the posterior petrous face between the region of the porus acousticus and the sigmoid sinus. STUDY DESIGN AND SETTING: A retrospective chart review from a large skull base surgery practice at a tertiary care institution. RESULTS: This cohort of patients presented with minimal symptoms, yet large tumors, averaging 3.8 cms and causing significant cerebellar compression. Retrosigmoid craniotomies afforded excellent exposure. CONCLUSION AND SIGNIFICANCE: Patients with large tumors emanating from the posterior fossa aspect of the temporal bone should be evaluated on the basis of their site of origin. Patients with tumors emanating from the anterior or ventral portion of the temporal bone have greater symptoms and greater operative complications than those emanating from the posterior petrous face, between the porus acousticus and sigmoid sinus. PMID- 11283499 TI - Action of histamine on eustachian tube function. AB - INTRODUCTION: The role of allergy in eustachian tube dysfunction is controversial. In this study, allergy was simulated by exposure to histamine, and eustachian tube function testing was performed in an experimental rat model. METHODS: Ventilatory function was assessed by measuring passive opening and closing pressures of the eustachian tube after challenge with either transtympanic or intranasal histamine. The mucociliary clearance time of the tubotympanum was assessed by observing dye transport from the middle ear to the nasopharynx after challenge with either transtympanic histamine or control solution. RESULTS: There was a statistically significant increase in passive opening and closing pressures with transtympanic histamine versus intranasal histamine. In addition, mucociliary clearance times of the tubotympanum after transtympanic histamine showed a statistically significant increase when compared with those after transtympanic control solution. CONCLUSIONS: Transtympanic histamine exposure causes eustachian tube dysfunction in the rat by increasing passive opening and closing pressures of the eustachian tube and impairing mucociliary clearance time. PMID- 11283500 TI - A new laser treatment for vocal cord papilloma--585-nm pulsed dye. AB - OBJECTIVES: Microvascular targeting with the 585- nm pulsed dye laser (PDL) may provide a new form of therapy to control symptoms caused by recurrent respiratory papillomatosis (RRP). METHODS: Ten patients with RRP underwent 13 procedures under general anesthesia with the 585-nm PDL. A micromanipulator (11 procedures) and a flexible nasolaryngoscope (2 procedures) were used to deliver the laser pulses. Patients were followed postoperatively according to protocol. RESULTS: Clinical examination revealed regression of papillomas in all patients. Seven patients had complete regression after PDL surgery, and 2 patients had partial response to treatment. One patient was lost to follow-up. No complications were present during this prospective nonrandomized pilot study. CONCLUSION: Patients treated with PDL experienced regression of their papillomas. PDL may provide patients with RRP with an alternative treatment without the risks associated with CO(2) laser surgery. This procedure also has potential to be delivered on an outpatient basis with flexible fiberoptic laryngoscopes. PMID- 11283501 TI - A novel CD44 v3 isoform is involved in head and neck squamous cell carcinoma progression. AB - OBJECTIVES: CD44 comprises a family of isoforms involved in tumorigenesis. Here we investigate the role of CD44 isoforms in head and neck squamous cell carcinoma (HNSCC) progression. MATERIALS AND METHODS: HNSCC specimens underwent reverse transcriptase-polymerase chain reaction (RT-PCR) followed by Southern blot analysis. After surface biotinylation, FaDu (hypopharyngeal HNSCC) and CD44v3 transfected COS-7 cells were CD44 antibody-precipitated and compared by Western blot analysis. FaDu cells underwent double immunofluorescence staining and growth assays. RESULTS: Southern blot analysis suggested differential CD44v3 isoform expression in tumor and normal tissue. Cloning and sequencing revealed 2 novel CD44v isoforms. Western blot analysis suggested CD44v3 expression in COS-7 transfectants and FaDu. Double immunofluorescence staining revealed co localization of CD44v3 and actin in FaDu projections. Anti-CD44v3 antibody decreased FaDu growth. CONCLUSION: HNSCC tissue and FaDu appear to express CD44v3 isoforms. These isoforms may promote tumorigenesis. CLINICAL SIGNIFICANCE: CD44v3 isoforms may be effective tumor markers and targets for HNSCC therapy. PMID- 11283502 TI - Node counts in neck dissection: are they useful in outcomes research? AB - OBJECTIVES: We studied the unilateral nodal yields for procedures reported as standard or modified radical neck dissection (RND) to determine their applicability in outcomes research. METHODS: We analyzed the unilateral nodal yields for all procedures reported as RND for carcinoma of the oral cavity, pharynx, and larynx at our institution from 1985 to 1997 (n = 64, no prior treatment). These included both standard and modified techniques, encompassing levels I through V of the neck. Each side of a bilateral RND was treated as a separate case. This sample was compared with a similar population from the National Cancer Institute's Surveillance, Epidemiology, and End-Results (SEER) registry. Nodal yield was obtained for RND alone and for unspecified neck dissection with primary excision for the same diseases and time period (n = 1499). RESULTS: The mean nodal yield from 64 RND was 30 vs 27 in the SEER data. The standard deviation was 14.7 compared with 17.2 in the SEER data. Values ranged from 7 to 66 nodes whereas the SEER range was from 1 to 97 nodes. Although the SEER data contain nodal yields from regional or selective neck dissection, we corroborate our findings of large variance in nodal yield from our RND sample. CONCLUSIONS: Large variance in nodal yields from RND may have undefined effects on quality of life, cure rate, and survival. Until correlation of nodal yields with outcomes is examined, we cannot know how to relate RND to outcomes. PMID- 11283503 TI - Potent effector function of tumor-sensitized L-selectin(low) T cells against subcutaneous tumors requires LFA-1 co-stimulation. AB - OBJECTIVE: Animal tumor models have demonstrated that adoptive transfer of tumor draining lymph node (TDLN) T lymphocytes can cure established tumors in many anatomic sites. However, subcutaneous tumors are relatively refractory and have required maximally tolerated doses of cells. The goals of this study were to determine whether a subset of TDLN T lymphocytes varying in expression of the cell adhesion molecule L-selectin (CD62L) had augmented therapeutic efficacy and to determine the co-stimulatory requirements for trafficking and anti-tumor effector function. STUDY DESIGN: TDLNs were recovered from mice bearing progressive MCA 205 fibrosarcomas, and the T lymphocytes were segregated into CD62L(low) and CD62L(high) subsets and activated ex vivo with anti-CD3 mAb and IL 2. Mice bearing established subcutaneous MCA 205 tumors were treated with activated T cell subsets and in some experiments with additional mAb against cell adhesion molecules. RESULTS: Adoptive transfer of as few as 5 x 10(6) activated cells cured mice bearing 3-day subcutaneous MCA 205 tumors initiated with 6 x 10(6) cells, and the tumors demonstrated a dense infiltrate of CD62L(low) cells. In marked contrast, adoptive transfer of 10 times as many T cells derived from the reciprocal CD62L(high) compartment had no effect on tumor growth. The effector function of the CD62L(low) T cells was clearly dependent on co stimulation through the cell adhesion molecule LFA-1, because anti-LFA-1 mAb completely abrogated the anti-tumor reactivity of the transferred cells against subcutaneous tumors and inhibited tumor infiltration. In contrast, blockade of ICAM-1, VLA-4, or VCAM-1 had no inhibitory effect on the anti-tumor function. CONCLUSION: These studies demonstrate the high therapeutic activity of the CD62L(low) subset of tumor-draining LN T cells against subcutaneous tumors, a relatively refractory site, and confirm the essential role of LFA-1 for effector T cell function. SIGNIFICANCE: Identification of the phenotype and requirements for effector function of T lymphocytes sensitized to tumor antigens has implications for clinical trials of adoptive immunotherapy for head and neck carcinoma using a similar approach. PMID- 11283504 TI - Expression of nitric oxide synthase type 3 in reflux-induced esophageal lesions. AB - BACKGROUND: The expression of endothelial constitutive nitric oxide synthase (NOS3) by squamous dysplasia and carcinomas of the head and neck has previously been described. We sought to compare NOS3 expression in squamous mucosa, glandular metaplasia, and adenocarcinoma of the esophagus. METHODS: Forty paraffin-embedded specimens from 20 patients with adenocarcinoma were stained with anti-NOS3 monoclonal antibody. The percentage of cells stained and the intensity of staining were determined for squamous epithelium, metaplasia, and adenocarcinoma. Staining characteristics were statistically analyzed according to clinical variables. RESULTS: NOS3 expression was significantly higher in adenocarcinoma and squamous epithelium compared with glandular metaplasia. Among the carcinomas, larger tumor size (T3/4), nodal positivity, and advanced TNM stage (III/IV) significantly correlated with increased NOS3 expression. CONCLUSIONS: NOS3 is expressed in reflux-induced lesions of the esophagus. Glandular metaplasia shows basal levels of NOS3 that significantly increase with malignant transformation and tumor progression. The role of free radicals in carcinogenesis is being actively studied. PMID- 11283506 TI - Association of 8p23 deletions with poor survival in head and neck cancer. AB - OBJECTIVE: Allelic loss at 8p23 occurs frequently in head and neck squamous cell carcinoma. The objective of this study was to determine the prognostic importance of 8p23 loss. STUDY DESIGN AND SETTINGS: We tested 51 primary tumors and 19 lymph node metastases for loss of heterozygosity with 7 microsatellite polymorphisms at 8p23 and correlated the results with disease-free interval and disease-specific survival. RESULTS: The Kaplan-Meier analysis demonstrated statistically significant association of 8p23 allelic loss with both shorter disease-free interval and disease-specific survival. For the pN stage, the log-rank test indicated significance in correlation with the disease-free interval, whereas the pT stage showed a significant correlation with disease-specific survival. Multivariate analysis identified loss of heterozygosity at 8p23 as independent prognostic marker for disease-free interval. CONCLUSION: Our data suggest that 8p23 allelic loss is associated with poor prognosis in head and neck squamous cell carcinoma and could be useful refining diagnosis of these tumors. PMID- 11283505 TI - Occult laryngeal pathology in a community-based cohort. AB - BACKGROUND: Little information is available regarding the prevalence of laryngeal pathology in adults. PURPOSE: To estimate the prevalence of occult laryngeal pathology in a community-based cohort of adults over 40 years of age. METHODS: One hundred consecutive volunteers over age 40 with no history of voice disorders were enrolled. All completed a self-administered laryngeal symptom questionnaire and underwent a comprehensive head and neck examination including transnasal fiberoptic laryngoscopy. RESULTS: The mean age of the cohort was 61 years. Vocal fold bowing (presbylaryngis) was present in 72% of the patients, and findings of laryngopharyngeal reflux were present in 64% of the cohort. In addition, other laryngeal pathology were identified in 21%. Only 12% had a completely normal laryngeal examination. CONCLUSIONS: Occult laryngeal pathology is very common in persons over 40. Findings suggestive of laryngopharyngeal reflux are present in 64%, and vocal fold bowing is present in 72% of persons over 40. PMID- 11283508 TI - Long-term histologic effects of inferior turbinate laser surgery. AB - OBJECTIVE: In this study we sought to define the histologic changes produced by laser treatment of inferior turbinates. STUDY DESIGN: Eight inferior turbinates with prior laser treatment (mean, 26.8 months) were analyzed by light microscopy after turbinectomy for relief of refractory nasal obstruction. Histologic findings were compared with those of a group of 8 hypertrophic inferior turbinates that had no previous laser surgery. RESULTS: Laser-treated areas of the inferior turbinates demonstrated a histologically bland appearance, with marked diminution of seromucinous glands and relative preponderance of connective tissue matrix. Prominence of venous sinusoids was also significantly reduced in the laser-treated areas. Surface epithelium including goblet cells was reconstituted over the areas of laser application. CONCLUSION: Clinical laser surgery of the inferior turbinate produces striking long-term histologic changes. SIGNIFICANCE: The data suggest a differential response of turbinate histologic components to application of laser energy, with the glandular component being particularly sensitive. Further correlative study is needed to clarify the clinical significance of laser-induced histologic changes in inferior turbinates. PMID- 11283507 TI - Reconstruction of large lower lip defects: a new method. AB - Lower lip defects may be due to some congenital or acquired problems, such as congenital naevi, hemangiomas, tumors, traumas, or infectious disease. The degree of substance loss may be skin, muscle, or mucoso, or a combination of one or all layers. In large defects of the lower lip, the challenge is to accomplish a result that meets the criteria of successful reconstruction. We present 3 patients with large defects of the lower lip from tumor ablation who underwent satisfactory reconstruction with a new method not previously described in the English-language literature. PMID- 11283509 TI - Endoscopic ligature of the sphenopalatine artery for severe posterior epistaxis. AB - OBJECTIVE: To present our experience with endoscopic ligature of the sphenopalatine artery in the treatment of severe posterior epistaxis of patients who had previously undergone conservative procedures. METHODS: Eleven patients with severe posterior epistaxis were treated during a 25-month period with an endoscopic ligature of the sphenopalatine artery. The basic principle of the surgical technique is to identify the branches of the sphenopalatine artery through an endoscopic endonasal approach and to apply a titanium clip under direct vision. RESULTS: The endoscopic ligature of the sphenopalatine artery was performed unilaterally in 10 patients and bilaterally in 1 patient, with a total of 12 ligatures. It was possible to identify the sphenopalatine artery in all cases with a successful outcome using this surgical technique alone. CONCLUSION: Endonasal endoscopic ligature of the sphenopalatine artery has been an effective surgical technique for treating severe posterior epistaxis. PMID- 11283510 TI - Kawasaki disease presenting as cervical lymphadenitis or deep neck infection. AB - OBJECTIVE: To describe a group of patients with Kawasaki disease who had cervical lymphadenopathy as their dominant initial presentations. MATERIALS AND METHODS: We retrospectively reviewed the medical records of 14 children who were admitted to Chang-Gung Children's Hospital between May 1996 and July 1998 with the initial impression of cervical lymphadenitis, cellulitis, and/or deep neck infection but for which a diagnosis of Kawasaki disease was established later. RESULTS: Five (35.7%) patients were less than 5 months of age, and 8 (57.1%) patients were more than 53 months of age. The mean duration for establishing a diagnosis of Kawasaki disease from the onset of illness was 8.2 (6 to 20) days. Initially, empiric antibiotics were prescribed in each case with unsatisfactory response. Intravenous immune gamma globulin (2 g/kg) was administered in 13 patients. Three (21.4%) patients developed coronary artery lesions. CONCLUSION: If a child less than 6 months or more than 4 years of age has a fever and an enlarged cervical lymph node and is unresponsive to empiric antibiotics, Kawasaki disease should be considered. PMID- 11283511 TI - Free transverse colon transfer for large pharyngostoma after pharyngolaryngoesophagectomy: a case report. PMID- 11283512 TI - Giant schwannoma of external auditory canal: a case report. PMID- 11283513 TI - Sinonasal neuroendocrine carcinoma presenting as a nasopharyngeal mass. PMID- 11283514 TI - Lymphangiomatous macroglossia causing upper airway obstruction and associated Plummer-Vinson syndrome. PMID- 11283515 TI - Unusual course of the anterior inferior cerebellar artery through the facial nerve in the cerebellopontine angle. PMID- 11283516 TI - Cocaine-induced sinonasal destruction. PMID- 11283517 TI - Nitric oxide and cell survival: megakaryocytes say "NO". PMID- 11283518 TI - Sarcopenia. AB - Sarcopenia is a term utilized to define the loss of muscle mass and strength that occurs with aging. Sarcopenia is believed to play a major role in the pathogenesis of frailty and functional impairment that occurs with old age. Progressive muscle wasting occurs with aging. The prevalence of clinically significant sarcopenia is estimated to range from 8.8% in young old women to 17.5% in old old men. Persons who are obese and sarcopenic (the "fat frail") have worse outcomes than those who are sarcopenic and non-obese. There is a disproportionate atrophy of type IIa muscle fibers with aging. There is also evidence of an age-related decrease in the synthesis rate of myosin heavy chain proteins, the major anabolic protein. Motor units innervating muscle decline with aging, and there is increased irregularity of muscle unit firing. There are indications that cytokines-especially interleukin-1beta, tumor necrosis factor alpha, and interleukin-6-play a role in the pathogenesis of sarcopenia. Similarly, the decline in anabolic hormones-namely, testosterone, dehydroepiandrosterone growth hormone, and insulin-like growth factor-I-is also implicated in the sarcopenic process. The role of the physiologic anorexia of aging remains to be determined. Decreased physical activity with aging appears to be the key factor involved in producing sarcopenia. An increased research emphasis on the factors involved in the pathogenesis of sarcopenia is needed. PMID- 11283519 TI - Lupus nephritis: lessons from experimental animal models. AB - Lupus nephritis is a frequent and severe complication of SLE. In the last decades, animal models for SLE have been studied widely to investigate the immunopathology of this autoimmune disease because abnormalities can be studied and manipulated before clinical signs of the disease become apparent. In this review an overview is given of our current knowledge on the development of lupus nephritis, as derived from animal models, and a hypothetical pathway for the development of lupus nephritis is postulated. The relevance of the studies in experimental models in relationship with our knowledge of human SLE is discussed. PMID- 11283520 TI - Effect of nitric oxide on megakaryocyte growth induced by thrombopoietin. AB - The present study investigated the effect of nitric oxide (NO) on megakaryocyte (Mk) proliferation induced by thrombopoietin (TPO). Low-density mononuclear cells (MNCs) and CD34+ cells from human bone marrow (BM) were cultured in liquid medium in the presence of sodium nitroprusside (SNP) or (Z)-1-[2-(aminoethyl)-N-(2 ammonioethyl) amino] diazen-1-ium-1, 2-diolate (DETA/NO) and then stimulated with TPO. Mk number decreased in both NO donors, as identified by flow cytometry 11 to 13 days after TPO stimulation. Nitrite, cyanide, or the carrier molecule DETA failed to reproduce the inhibition caused by NO donors. When CD34+ cells were treated with DETA/NO, the inhibition of Mk growth was even more pronounced than that in MNCs. Failure of the guanosine 3',5'-cyclic monophosphate (cGMP) analog 8 bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP) to inhibit Mk proliferation suggests that cGMP is not involved in Mk suppression mediated by NO. On the other hand, DNA analysis by flow cytometry showed that apoptosis of CD34+ cells and Mks seemed to be at least one of the mechanisms associated with the cytotoxic DETA/NO effect. Stimulation of MNCs or CD34+ cells with tumor necrosis factor-alpha (TNF alpha) and interferon-gamma (IFN-gamma) increased endogenous NO levels and suppressed Mk growth. Treatment with NO synthesis inhibitors such as L -N(G) monomethyl arginine (L -NMMA) or L -N(G)-nitroarginine methyl ester hydrochloride (L -NAME) partially reversed Mk growth inhibition induced by TNF-alpha and IFN gamma, although increased NO levels returned to normal values. The results presented here strongly indicate that NO regulates the growth of Mks induced by TPO by a direct effect on both progenitors and mature Mks. PMID- 11283521 TI - Mechanisms in the inhibition of neointimal hyperplasia with triflavin in a rat model of balloon angioplasty. AB - RGD-containing peptides are able to inhibit the binding of ligands to certain beta3 integrins, such as alpha(IIb)beta3 and alpha(v)beta3, both of which are involved in neointimal hyperplasia. The present study was designed to elucidate the detailed mechanisms involved in the inhibition of neointimal hyperplasia with triflavin in a rat model of balloon angioplasty. Triflavin (0.25 mg x kg(-1) x d( 1)), an RGD-containing disintegrin, time dependently inhibited both neointimal hyperplasia and lumen occlusion after angioplasty in carotid arteries of rats. Furthermore, electron micrographs highlighted that SMCs were phenotypically different from the typical contractile, spindle-shaped SMCs normally seen in uninjured vessel walls. PDGF-BB was strongly produced in thrombus formation and neointimal SMCs after angioplasty, and triflavin significantly reduced PDGF-BB expression in vessel lumens and neointimal SMCs after angioplasty. Balloon angioplasty caused a significant increase of nitrate and cyclic guanosine monophosphate levels compared with levels found in sham-operated rats, and these were not significantly changed with infusion of triflavin (0.25 mg x kg(-1) x d( 1)). Furthermore, the plasma level of TXB2 obviously increased after angioplasty, and triflavin markedly suppressed the elevation of plasma TXB2 concentration. The results indicate that triflavin effectively prevents neointimal hyperplasia, possibly through the following 2 mechanisms. First, triflavin binds to alpha(IIb)beta3 integrin on platelet membranes, resulting in inhibition of platelet adhesion, secretion, and aggregation in injured arteries, followed by inhibition of TXA2 formation and PDGF-BB release from platelets. Second, triflavin may also bind to alpha(v)beta3 integrin on SMCs, thus subsequently inhibiting cell migration and proliferation. These results provide new insights into the mechanisms of neointimal hyperplasia and have significant implications for disintegrin therapy for the treatment of restenosis and atherosclerosis. PMID- 11283522 TI - Alterations of glomerular and extracellular glutathione peroxidase levels in patients and rats with focal segmental glomerulosclerosis. AB - Glutathione peroxidase (GPX) is an important antioxidant that effectively scavenges hydrogen peroxide. Recent studies have revealed that plasma GPX activity is decreased in patients with chronic kidney failure. However, there have been no reports on renal and urinary GPX levels in patients with kidney disorders. Therefore in this study we have measured the plasma and urinary GPX levels and glomerular GPX immunostaining in patients with focal segmental glomerulosclerosis (FSGS) and normal kidney function as compared with those in patients with minimal change disease (MCD) and those in normal control subjects. Rats with puromycin aminonucleoside-induced FSGS were also studied. The results showed that the plasma GPX level was significantly lower in FSGS patients than in either MCD patients or normal control subjects (both P <.01). The urinary GPX level was also significantly lower in FSGS patients than in either MCD patients (P <.05) or normal control subjects (P <.01). The immunostaining score of glomerular GPX was significantly lower in FSGS patients than in normal control subjects (both P <.05). Serial examinations of glomerular GPX immunostaining in FSGS rats also demonstrate a decrease in the score with the progression of disease. Our results indicate that all the plasma, urinary, and glomerular GPX levels are decreased in FSGS patients, indicating a decreased antioxidant defense in the early stages of chronic glomerular diseases. PMID- 11283523 TI - Effect of metals on Candida albicans growth in the presence of chemical chelators and human abscess fluid. AB - Calprotectin is a calcium- and zinc-binding protein that is present in abscess fluid supernatants and appears to inhibit microbial growth through competition for zinc. In the present study, growth inhibition by chemical chelators was compared with that produced by human abscess fluid to determine whether other chelators, perhaps with different metal specificities, would have the same or different patterns of metal reversibility as abscess fluid. Zinc was found to be more potent than the other metals tested in reversing C. albicans growth inhibition by human abscess fluid and three chemical chelators, even though in some cases the stability constants of certain of these chelators were higher for other metals. For example, in the presence of the chelator diethylenetriaminopentaacetic acid, zinc stimulated Candida growth at a 10-fold lower concentration than did iron, even though this chelator has a stability constant for iron that is almost 10(10) higher than that for zinc. These results suggest that the zinc specificity of calprotectin's C. albicans growth inhibition can best be explained by the marked sensitivity of this organism to zinc deprivation rather than by selective binding of this metal by the protein. PMID- 11283524 TI - Rapid differentiation of five common alpha-thalassemia genotypes by polymerase chain reaction. AB - The alpha-thalassemias are common genetic disorders that arise from reduced synthesis of the alpha-globin chains. At present, large-scale carrier screening and clinically valuable antenatal detection programs have not been established for the congenital disorder alpha-thalassemia (alpha-thal). We have developed a simple nonradioactive polymerase chain reaction (PCR) approach that can detect and differentiate several common alpha-globin gene deletional alpha-thals regardless of the break points. When three primer sets were used--two gene specific sets for the alpha1- and alpha2-globin genes and one set for the beta actin gene (serving as an internal control)--PCR products from genomic DNA were simultaneously amplified and analyzed after coamplification and gel electrophoresis. The number of alpha-globin genes present in the subjects was determined by the intensity of alpha1 and alpha2 bands normalized with that of beta-actin when using densitometry. Our results demonstrate that five common genotypes of deletional alpha-thal are differentiated by the ratios of alpha1/beta-actin and alpha2/beta-actin. We also examined the feasibility of coupling this allele-specific amplification to a color-complementary assay. This easy and reproducible PCR assay is suitable for identifying alpha-thal carriers in screenings of large populations and improving genetic counseling. PMID- 11283525 TI - Polymorphonuclear leukocytes in coagulating whole blood recognize hydrophilic and hydrophobic titanium surfaces by different adhesion receptors and show different patterns of receptor expression. AB - The mechanism of healing or rejection of implant materials is unknown, but the process always starts at the contact with coagulating blood. Here, the initial reactions of clean (hydrophilic) and alkylated (hydrophobic) titanium with blood were investigated by short-term exposure to human blood and detection of polymorphonuclear leukocyte (PMNL) surface antigens with an immunofluorescence technique. The fluorescence intensity was quantitated by computer-aided image analysis. Antibodies specific to CD11b, CD16, CD18, CD62L, and CD162 were used to block PMNL adhesion. The respiratory burst of adhering cells was stimulated with opsonized zymosan and measured by chemiluminiscence. The thrombin dependence of PMNL reactions was studied by using hirudin, a specific thrombin inhibitor. The expression of CD62L decreased with increasing exposure time, and the rate of decrease was faster at the hydrophilic surface. At the hydrophilic surface, the CD16 exposure was high after 8 minutes of blood contact, and it decreased with time. At the hydrophobic surface, a peak in CD16 expression was seen after 32 minutes of blood exposure. At the hydrophobic surface, the expression of CD11b increased slowly with increasing blood exposure time, whereas at the hydrophilic surface, a peak of CD11b expression was seen after 32 minutes of blood exposure. The expression of CD11b and that of CD16 were found to be thrombin dependent. At the hydrophilic surface, adhesion of PMNLs was blocked by CD16 antibodies, whereas adhesion to the hydrophobic surface was blocked by anti-CD162. Mixing blood with antibodies to CD11b, CD18, and CD62L amplified the adhesion of PMNLs to the hydrophilic surface. PMID- 11283526 TI - Efficacy of bactericidal/permeability-increasing protein in experimental otitis media with effusion in rats: a new therapy for mucosal infections. AB - Otitis media with effusion (OME) is characterized by the presence of fluid in the middle ear without signs or symptoms of acute infection and by persistent changes in the middle ear mucosa. These are mainly induced by gram-negative bacterial infection and dysfunction of the eustachian tube (ET). Gram-negative bacteria (GNB) contain lipopolysaccharide (LPS) in their outer membrane that is responsible for inflammatory reactions in the middle ear. In this study we investigated the therapeutic effect of a recombinant LPS-binding protein, bactericidal/permeability-increasing protein (rBPI21), on the repair of mucosal damage in rats with experimentally induced OME. OME was induced by obstruction of the eustachian tube in combination with LPS injection. Twelve weeks after OME induction, secretory cells in the tympanic orifice of the middle ear were increased from an average of 14 +/- 2 to 31 +/- 5, ciliated cells were decreased from 24 +/- 4 to 6 +/- 4, and the number of macrophages in the subepithelial layer increased from 13 +/- 4 to 27 +/- 3. A single dose of rBPI21 was administered directly into the middle ear cavity 2 weeks after the induction of OME. Histologic examination of the middle ear mucosa at 4 and 12 weeks after OME induction showed that mucosal changes were restored by rBPI21 treatment. These results demonstrate that the middle ear mucosa recovers from inflammatory changes associated with OME after treatment with rBPI21. This suggests that rBPI21 may be useful in the treatment of OME and of mucosal infections of the respiratory tract. PMID- 11283527 TI - Effect of laparoscopy on intra-abdominal blood flow. PMID- 11283528 TI - A prospective randomized comparison of laparoscopic appendectomy with open appendectomy: Clinical and economic analyses. AB - BACKGROUND: Previous randomized studies of laparoscopic appendectomy produced conflicting recommendations, and the adequacy of sample sizes is generally unknown. We compared clinical and economic outcomes after laparoscopic and open appendectomy in a sample of predetermined statistical power. METHODS: A pre-study power analysis suggested that 200 randomized patients would yield 80% power to show a mean decrease of 1.3 days' hospitalization. One hundred ninety-eight patients with a preoperative diagnosis of acute appendicitis were randomized prospectively to laparoscopic or open appendectomy. Economic analysis included billed charges, total costs, direct costs, and indirect costs associated with treatment. RESULTS: Laparoscopic appendectomy took longer to perform than open appendectomy (median, 107 vs 91 minutes; P <.01) and was associated with fewer days to return to a general diet (mean, 1.6 versus 2.3 days; P <.01), a shorter duration of parenteral analgesia (mean, 1.6 versus 2.2 days; P <.01), fewer morphine-equivalent milligrams of parenteral narcotic (median, 14 mg versus 34 mg; P =.001), a shorter postoperative hospital stay (mean, 2.6 versus 3.4 days; P <.01), and earlier return to full activity (median, 14 versus 21 days; P <.02). However, operative morbidity and time to return to work were comparable. Billed charges and direct costs were not significantly different in the 2 groups ($7711 versus $7146 and $5357 versus $4945, respectively), but total costs (including indirect costs) of laparoscopic appendectomy were, on average, nearly $2400 less, given the shorter length of stay and abbreviated recuperative period ($11,577 versus $13,965). Subgroup analyses suggested the benefit of a laparoscopic approach for uncomplicated appendicitis and for patients with active lifestyles. CONCLUSIONS: While laparoscopic appendectomy is associated with statistically significant but clinically questionable advantages over open appendectomy, a laparoscopic approach is relatively less expensive. The estimated difference in total costs of treatment (direct and indirect costs) was at least $2000 in more than 60% of the bootstrapped iterations. The economic significance and implications favoring a laparoscopic approach cannot be ignored. PMID- 11283529 TI - Lymphatic spreading pattern of intrahepatic cholangiocarcinoma. AB - BACKGROUND: There have been very few reports on the pattern of lymphatic spread of intrahepatic cholangiocarcinoma. This pattern was elucidated to help define the rational extent of radical lymph node dissection. METHODS: Thirty-nine consecutive patients who underwent hepatectomy with radical lymph node dissection were reviewed retrospectively. RESULTS: Lymph node metastases were detected in 24 of the 39 patients (62%). The metastatic nodes were found in the hepatoduodenal ligament, along the common hepatic artery, around the abdominal aorta, on the posterior surface of the pancreas head, along the left gastric artery, along the superior mesenteric artery, around the celiac artery, along the lesser curvature of the stomach, and around the cardia. The nodal involvements along the left gastric artery, along the lesser curvature, or around the cardia were recognized only in the left peripheral and hilar types of cholangiocarcinoma, while all other sites included both the left or right peripheral type and the hilar type cholangiocarcinoma. CONCLUSIONS: Intrahepatic cholangiocarcinomas, irrespective of their intrahepatic location, mainly spread to the nodes in the hepatoduodenal ligament, then to the para-aortic nodes, retropancreatic nodes, or common hepatic artery nodes. In addition to these spreading routes, the left peripheral type or hilar type of cholangiocarcinoma tends to spread along the left gastric nodes through the lesser curvature. PMID- 11283530 TI - Estimating the prognosis of hepatic resection in patients with metastatic liver tumors from colorectal cancer with special concern for the timing of hepatectomy. AB - BACKGROUND: The effect of the time interval between colorectal and liver resection for metastatic lesions on the patient's survival remains controversial. Pretreatment classification for predicting the prognosis of this disease has not yet been reported. METHODS: Nine clinical factors revealed by preoperative examinations, intraoperative screening before liver resection, and resection margin were examined in 304 patients who underwent hepatic resections for metastatic colorectal carcinoma. The patients were divided according to the timing of hepatectomy and both tumor number and maximum diameter to devise a staging system. RESULTS: Tumor number and maximum tumor size were significant prognostic factors in the metachronous hepatectomy group, and resection margin was significant in the synchronous group. The following staging system was proposed: stage I, n < or = 3 and diameter < 5 cm in the metachronous hepatectomy group (n = 86); stage II, n < or = 3 and diameter > or = 5 cm in the metachronous hepatectomy group (n = 46); and stage III, n > or = 4 and diameter > 5 cm in the metachronous group and the synchronous hepatectomy group (n = 144). CONCLUSIONS: The current study revealed that the factors that influenced the patient's prognosis were different between the synchronous and metachronous groups. It may be useful to develop a staging system that considers this difference. PMID- 11283531 TI - Eight years of experience with transjugular retrograde obliteration for gastric varices with gastrorenal shunts. AB - BACKGROUND AND OBJECTIVES: There is no standard treatment for gastric varices. Transjugular retrograde obliteration (TJO) is one way of obliterating gastric varices with gastrorenal shunts, in which blood flow is abundant. Our aim was to examine our experience with TJO during an 8-year period and to determine the long term effects of this treatment. METHODS: We performed TJO procedures in 52 patients to obliterate gastric varices. All the patients had liver cirrhosis. Sixteen had hepatocellular carcinoma (HCC) without vascular invasion. We inserted an angiographic catheter with an occlusive balloon through the right internal jugular vein into the gastrorenal shunt or the gastric varices. After controlling the other blood-draining routes with a microcoil or absolute ethanol, or both, we injected 5% ethanolamine oleate with iopamidol into the gastric varices under fluoroscopy. RESULTS: The gastric varices were successfully obliterated by TJO in all cases. The complications were all minor and transient. The mortality rate for TJO was 0%. There was no recurrence and no bleeding of gastric varices at all after TJO. Patient survival differed depending on the presence or absence of HCC (P <.05). The development of HCC in the cirrhotic liver was the most common cause of late death. Gastrointestinal bleeding was not a cause of death. The occurrence rate of esophageal varices after TJO was high, but these varices could be treated easily by endoscopic injection sclerotherapy before they bled. CONCLUSIONS: Portal blood flow through the gastrorenal shunt is diverted to the porto-azygos venous system after the gastrorenal shunt is obliterated by TJO. TJO is a safe option that we recommend for treating gastric varices with gastrorenal shunts, provided that the TJO is followed by endoscopic injection sclerotherapy. PMID- 11283532 TI - Impact of repeat hepatectomy on recurrent colorectal liver metastases. AB - BACKGROUND: Hepatic recurrence is seen in approximately 40% of patients undergoing hepatectomy for colorectal metastases. This study was designed to assess the risks and clinical benefits of repeat hepatectomy for those patients. METHODS: Twenty-six patients underwent repeat hepatectomy for hepatic recurrence, and their clinical data were retrospectively reviewed for operative morbidity and mortality, performance level, and survival. RESULTS: There was no operative mortality after repeat hepatectomy. Operative bleeding was significantly increased in the second hepatectomy; but operating time, duration of hospital stay, and performance status after the second hepatectomy were comparable with those of the initial hepatectomy. The median survival time from the second hepatectomy was 31 months, and the 3- and 5-year survival rates were 62% and 32%, respectively. A short disease-free interval (6 months or less) between the initial hepatectomy and diagnosis of hepatic recurrence in the remnant liver was significantly associated with poor survival after the second hepatectomy. CONCLUSIONS: Repeat resection contributed to clinical benefits for selected patients with hepatic recurrence after the initial hepatectomy for colorectal liver metastases. However, appearance of hepatic recurrence within 6 months or less after the initial hepatectomy is a poor prognostic factor for repeat hepatectomy. PMID- 11283533 TI - Occult papillary carcinoma of the thyroid presenting as a cervical cyst. AB - BACKGROUND: A cystic neck mass representing metastatic papillary thyroid cancer to a cervical lymph node may be the presenting symptom in patients with an occult papillary cancer of the thyroid. This cystic change can cause diagnostic problems and not infrequently delay identification of the primary thyroid tumor. This study investigates the frequency, treatment, and pathologic features of this entity. METHODS: All clinical charts and microscopic slides of 136 consecutive patients who underwent thyroid operation for papillary carcinoma (PC) from 1990 to 1995 were reviewed. Hematoxylin-and-eosin and immunohistochemical stains (IMHS) for thyroglobulin also were reviewed. RESULTS: Eight patients (5.8%) presented with a cystic neck mass and no palpable thyroid lesion. In all 8 patients, the diagnosis was made by an excision of the cystic neck mass. In 3 patients, the cyst demonstrated classical features of PC, such as papillae and psammoma bodies. In the remaining 5 (62%), only focal papillae or nuclear features of papillary carcinoma were present. A careful review of the histology and IMHS were necessary to arrive at the correct diagnosis in these 5 patients. CONCLUSIONS: Occult papillary cancer of the thyroid presenting as a cystic neck mass is not uncommon and must be considered in the differential diagnosis. Excision and careful review of the histology and IMHS is necessary to prevent delay of the proper diagnosis. Although the thyroid tumor was less than 1 cm and sometimes only microscopic, the extensive nodal metastasis has led us to favor near total or total thyroidectomy and modified neck dissection in this entity. PMID- 11283534 TI - Predictive factors for perioperative blood transfusions in rectal resection for cancer: A multivariate analysis of a group of 212 patients. AB - BACKGROUND: In colorectal cancer surgery, allogeneic blood transfusions have reportedly been associated with higher rates of postoperative complications and tumor recurrence. However, because of the increased cost of alternative types of blood transfusions (eg, the use of autologous blood or erythropoietin administration), their routine use cannot be recommended. This study evaluated the risk factors for perioperative blood transfusions in resection for rectal cancer in order to identify patients who could benefit from these methods. METHODS: From 1990 to 1997, 212 consecutive patients who underwent elective rectal resection for cancer were reviewed. The associations between perioperative heterologous blood transfusion and 18 patient-, tumor-, surgical-, and treatment related variables were assessed by univariate and multivariate analysis. RESULTS: Of the 212 patients, 72 (34%) received transfusions. Multivariate analysis revealed that 5 preoperative variables were significant risk factors for perioperative blood transfusion: age > 65 years (P =.03), body mass index > 27 kg/m(2) (P =.04), preoperative hemoglobin < or = 12.5 g/dL (P <.0001), American Society of Anesthesiologists status > 2 (P =.024), and additional surgical procedures (P =.018). In patients with anemia, the risk of transfusion was at least 47% in patients with 1 other risk factor or more. In nonanemic patients, the risk of transfusion was under 11% in patients with 1 risk factor or none, but increased to 47% in those with 2 or more risk factors. CONCLUSIONS: Our analysis of risk factors for perioperative blood transfusion in rectal resection for cancer must be considered to constitute guidelines for a more responsible use of the expensive alternatives to allogeneic blood transfusion. PMID- 11283535 TI - Noncardiac surgery in patients with left ventricular assist devices. AB - BACKGROUND: Noncardiac surgery, especially abdominal surgical procedures in patients with long-term mechanical circulatory support and strong anticoagulation, is difficult. METHODS: We report on 14 patients (aged 44 +/- 15 years) with a portable Novacor or HeartMate system, who underwent noncardiac surgical procedures while being supported by the device. RESULTS: The patients underwent 20 procedures for noncardiac reasons; most had an intestinal operation or cholecystectomy. Half of the procedures were performed within 30 days after placement of the device (mean interval, 53 +/- 57 days), only 6 interventions were necessary after 100 days of mechanical support. Complications occurred in 8 patients (57%), 5 of whom had undergone cholecystectomy and had unacceptably high sanguineous drainage losses. CONCLUSIONS: An elective surgical procedure can be performed with an acceptable risk if the operation is carefully managed. Postponing resumption of full anticoagulation is advisable as it may reduce bleeding complications without apparently increasing the risk for thromboembolism. Emergency interventions remain a difficult task. PMID- 11283536 TI - A novel collagen-based composite offers effective hemostasis for multiple surgical indications: Results of a randomized controlled trial. AB - BACKGROUND: Intraoperative bleeding is ubiquitous during open surgical procedures and uniformly effective hemostasis remains elusive. We conducted a randomized controlled trial to determine the effectiveness of a novel collagen-based composite (CoStasis Surgical Hemostat) compared with standard methods of hemostasis during general, hepatic, cardiac, and orthopedic operations. METHODS: Hemostatic treatment was assigned randomly to 347 subjects; 318 subjects (167 CoStasis, 151 controls) underwent operation, received treatment, and provided hemostatic success data. CoStasis was applied to the bleeding site without manual pressure as a sprayable liquid composite of bovine microfibrillar collagen, bovine thrombin, and autologous plasma. Manual compression was used as the control hemostat. Hemostatic success was achieved if bleeding had ceased completely within 10 minutes (3 minutes for cardiac subjects). The time to controlled bleeding (ie, slight oozing) and time to complete hemostasis were recorded for all subjects. RESULTS: Hemostatic success was achieved in more than 90% (153/167) of CoStasis subjects compared with 58% (88/151) of control subjects (P =.01). Superior hemostatic effectiveness with CoStasis was realized in every surgical specialty: general (77/79 vs 49/75, P =.01), hepatic (38/39 vs 20/29, P =.01), cardiac (28/37 vs 17/37, P =.02), and orthopedic (10/12 vs 2/10, P =.01). The duration of bleeding was also significantly shorter with CoStasis. The median time to controlled bleeding (42 seconds vs 150 seconds, P =.0001) and time to complete hemostasis (75 seconds vs 252 seconds, P =.0001) were both markedly longer with the control intervention. There were no serious adverse events related to the use of CoStasis. CONCLUSIONS: CoStasis is more effective at controlling and stopping diffuse intraoperative bleeding than standard methods of hemostasis in 4 distinct surgical indications representing a wide variety of operative interventions. PMID- 11283537 TI - Femorofemoral bypass grafts: Factors influencing long-term patency rate and outcome. AB - BACKGROUND: Crossover femorofemoral bypass graft (CFFBG) was proposed in the early days of modern vascular surgery to treat patients affected with unilateral iliac artery disease who were a high surgical risk. We investigated factors influencing short- and long-term outcomes of CFFBG: METHODS: The study was designed as a retrospective clinical study in a university hospital setting with a base of 228 patients. Of these patients, 154 (67.5%) presented a high surgical risk. The indication for operation was limb-threatening ischemia in 188 (82.5%) patients. All patients underwent CFFBG: The procedure was performed in 150 patients as the primary operation and in 78 patients after previous vascular graft failure or infection, or both. A preoperative percutaneous transluminal angioplasty was performed in 57 patients (25%) to correct donor iliac artery disease. In 127 patients (55.7%), an associated vascular procedure was performed to improve the outflow. Postoperative complications; 5- and 10-year primary, secondary, and limb salvage rates; and factors influencing short- and long-term results were assessed. RESULTS: Thirteen (5.7%) postoperative deaths occurred. Postoperative mortality and morbidity rates were significantly higher in patients aged more than 65 years (7.9% versus 3.5% and 18.6% versus 6.1%, respectively, P <.03). Primary and secondary patency rates at 5 and 10 years were 70.2% and 48.1%, 82.8% and 63.2%, respectively; 5- and 10-year limb salvage and survival rates were 85.5% and 80.1%, 63.3% and 31.0%, respectively. Ten-year primary and secondary patency and limb salvage rates were significantly lower when the procedure was performed after previous vascular graft failures (50.2% versus 26.5%, P <.007; 74.1% versus 44.1%, P <.01; and 84.3% versus 72.5%, P <.03, respectively). Five- and 8-year patency rates of autogenous vein CFFBG (34.3% and 22.8%, respectively, P <.03) were significantly lower than those of expanded polytetrafluoroethylene (71.1% and 59.8%, respectively) and polyester (77.3% and 50.3%, respectively) CFFBG: Moreover, 5- and 10-year primary and secondary patency rates were significantly better when externally supported grafts were used as compared with those without external support (80.1% and 69.9% versus 61.1% and 21.1%, P <.01; 88.8% and 75.9% versus 78.9% and 45.4%, P <.05, respectively). Multivariate analysis showed that the only variable associated with poor primary and secondary patency and limb salvage rates was the operation performed after previous vascular graft failures (P <.04, P <.03 and P<.05, respectively). CONCLUSIONS: CFFBG allows early and long-term results similar to those obtained with reconstructions originating from the aorta when it is performed as a primary operation when an adequate outflow is provided and externally supported prosthetic material is used. PMID- 11283538 TI - Use of an artificial neural network to quantitate risk of malignancy for abnormal mammograms. AB - BACKGROUND: The purpose of this study was to develop a simplified method for standardized categorization of patients with abnormal mammograms by incorporating quantitative risk assessment. METHODS AND PATIENTS: A prospective collection of 1288 outpatient referrals to a surgeon for abnormal mammograms, 185 (14.4%) with malignancy, was studied. Artificial neural network (ANN) and logistic regression (LR) models were developed and compared with the surgeon's clinical impression. The first 490 patients were used as the training set for each model. The ANN and LR were tested on the remaining patients, who were divided into 2 consecutive groups. The main outcome measures were (1) the accuracy (receiver operating characteristic [ROC] curve analysis) of biopsy recommendations based on the surgeon's impression and created by the 2 models and (2) the percentage of cancers that were falsely categorized as benign by the surgeon or the 2 models. RESULTS: Despite the fact that the surgeon's clinical impression showed good discrimination (area under ROC = 0.86), 13 of 708 cases (1.8%) thought to be benign by the surgeon proved to be carcinomas. The neural network (but not the LR model) was statistically superior to the surgeon's impression (ANN: ROC = 0.89, P =.004; LR: ROC = 0.86). Additionally, the computerized models were able to quantitate risk. Those patients predicted to be "benign" by the network (n = 391) had only a 0.5% risk of intraductal carcinoma and no invasive carcinoma, whereas 47% of those patients in the highest risk quartile had cancer. Both computerized models predicted a need for biopsy in 11 of 13 of the lesions (85%) missed by the surgeon's impression. Each model missed only 2 cases of intraductal carcinoma in young women. CONCLUSIONS: Computerized risk stratification models, used in routine practice, may help surgeons with decision making. The use of either model helps quantitate risk, thereby facilitating discussions with patients, and may reduce the number of "missed" cancers. PMID- 11283539 TI - Raxofelast, a hydrophilic vitamin E-like antioxidant, stimulates wound healing in genetically diabetic mice. AB - BACKGROUND: Impaired wound healing is a well-documented phenomenon in experimental and clinical diabetes. Emerging evidence favors the involvement of free radicals in the pathogenesis of diabetes-related healing deficit. This study assessed the effect of systemic administration of raxofelast, a protective membrane antioxidant agent, on wound healing by using healing-impaired (db/db) mice. METHODS: The wound healing effect of raxofelast was investigated by using an incisional skin-wound model produced on the back of female diabetic C57BL/KsJ db+/db+ mice and their healthy littermates (db+/+m). Animals were then randomized to the following treatment: raxofelast (15 mg/kg/d intraperitoneally) or its vehicle (dimethyl sulfoxide/sodium chloride 0.9%, 1:1, vol/vol). The animals were killed on different days, and the wounded skin tissues were used for histologic evaluation and for analysis of malondialdehyde (MDA) level and myeloperoxidase (MPO) activity, wound breaking strength, and collagen content. RESULTS: Diabetic mice showed delayed wound healing together with low collagen content, breaking strength, and increased MDA levels and MPO activity when compared with their healthy littermates. The administration of raxofelast did not modify the process of wound repair in healthy (db/+) mice, but significantly improved impaired wound healing in diabetic mice through the stimulation of angiogenesis, reepithelialization, synthesis, and maturation of extracellular matrix. Furthermore, raxofelast treatment significantly reduced MDA levels, MPO activity, and increased the breaking strength and collagen content of the wound. CONCLUSIONS: The current study provides evidence that raxofelast restores wound healing to nearly normal levels in experimental diabetes-impaired wounds and suggests that an increased lipid peroxidation in diabetic mice may have a role in determining a defect of wound repair. PMID- 11283540 TI - Chronic extrinsic denervation after small bowel transplantation in rat jejunum: Effects and adaptation in nitrergic and non-nitrergic neuromuscular inhibitory mechanisms. AB - BACKGROUND: Extrinsic denervation of the transplanted small bowel could play a substantial role in motor dysfunction of the transplanted gut. We attempted to determine the effect of chronic extrinsic denervation on intestinal contractility. METHODS: Jejunal longitudinal muscle strips were obtained from rats 1 week and 8 weeks after (1) syngeneic small bowel transplantation, (2) ischemia/reperfusion, or (3) gut transection/reanastomosis. Nonoperated rats (naive controls) and sham-operated rats (sham controls), 1 week after celiotomy/gut manipulation, served as controls. We evaluated the effects of exogenous nitric oxide, increasing doses of cholinergic and adrenergic agonists, and electrical field stimulation (EFS) in the presence or absence of N(G) monomethyl-l-arginine, methylene blue, tetraethylammonium, or tetrodotoxin. RESULTS: Spontaneous contractile activity (_chi +/- SEM), when compared with the naive controls (11.3 +/- 2.0 g.5 min/mg), was increased in all 4 groups at 1 week (15.9 +/- 10 to 19.4 +/- 2 g.5 min/mg; P < or =.03 each) but not at 8 weeks postoperatively. The inhibition of contractile activity by nitric oxide was increased in small bowel transplantation in naive controls at 8 weeks to 80% +/- 10% versus 50% +/- 7% (P <.02). EFS induced an inhibition of contractile activity that was tetraethylammonium- and tetrodotoxin-sensitive but N(G)-monomethyl-l arginine- and methylene blue-insensitive; the maximal EFS-induced inhibition was increased at 1 week and 8 weeks but only in the small bowel transplantation groups to 103% +/- 5% and 95% +/- 7%, respectively, versus 72% +/- 8% in naive controls (P A; A148T) but no mutations. Two of the 10 specimens with PanIN revealed an inactivating hypermethylation of the p16(INK4a) promoter. CONCLUSIONS: This study shows for the first time that p16(INK4a) alterations can be observed in a considerable number of PanIN1 in chronic pancreatitis tissues not associated with pancreatic cancer. Therefore, p16(INK4a) alterations, especially promoter methylation, might indicate high-risk precursors in chronic pancreatitis that might progress to cancer. PMID- 11283543 TI - Ampullary carcinoma developing after androgenic steroid therapy for aplastic anemia: Report of a case. AB - Sex steroids influence the development and course of human genital carcinomas including breast, testis, prostata, and ovarian cancers. (1) Other carcinomas such as hepatoma, cholangioma, and pancreatic cancer have also been reported to be related to sex hormones. (2-4) The existence of sex hormone receptors has been demonstrated immunohistochemically in specimens of these diseases. We recently encountered a patient in whom an ampullary carcinoma developed 39 months after the start of androgenic steroid therapy for aplastic anemia. Immunohistochemic analysis of resected tumor specimens of the patient suggested a possible hormonal effect on the tumor oncology. PMID- 11283544 TI - Mediastinoscopic esophagectomy using carbon dioxide insufflation via the neck approach. PMID- 11283545 TI - Pinch grafts and zinc: Memories of Walter Pories. PMID- 11283546 TI - Sclerosing mesenteritis. PMID- 11283547 TI - Pancreatitis secondary to Ascaris lumbricoides infestation. PMID- 11283548 TI - Laparoscopic adhesiolysis in the treatment of chronic abdominal pain. PMID- 11283550 TI - Computed tomography in suspected acute appendicitis. PMID- 11283552 TI - Miracle cures and embryonic science. PMID- 11283553 TI - BioPulse faces US and Mexican malpractice probes. PMID- 11283554 TI - Bush FY 2002 federal budget request vague. PMID- 11283555 TI - New US committee considers xenotransplants. PMID- 11283556 TI - New Zealand GMO debacle undermines green lobby. PMID- 11283557 TI - Italy performs GMO trial about-face. PMID- 11283558 TI - Survey raises concerns about Bt resistance management. PMID- 11283559 TI - GlaxoSmithKline presents a biotech facade. PMID- 11283574 TI - Vapornomics. PMID- 11283575 TI - Precaution without principle. PMID- 11283576 TI - Persistent failures in gene repair. PMID- 11283577 TI - Immunex's Enbrel enrollment program. PMID- 11283578 TI - Growing pains for biopharmaceuticals. PMID- 11283579 TI - Switching nucleic acids for antibodies. PMID- 11283580 TI - A RACHITT for our toolbox. PMID- 11283581 TI - A clearer vision for in vivo imaging. PMID- 11283582 TI - Toward the phosphoproteome. PMID- 11283588 TI - The potential of nucleic acid repair in functional genomics. AB - Chimeric RNA/DNA oligonucleotides have been used successfully to correct point and frameshift mutations in cells as well as in animal and plant models. This approach is one of several nucleic acid repair technologies that will help elucidate the function of newly discovered genes. Understanding the mechanisms by which these different technologies direct gene alteration is essential for progress in their application to functional genomics. PMID- 11283589 TI - Receptor-targeted optical imaging of tumors with near-infrared fluorescent ligands. AB - We report here the in vivo diagnostic use of a peptide-dye conjugate consisting of a cyanine dye and the somatostatin analog octreotate as a contrast agent for optical tumor imaging. When used in whole-body in vivo imaging of mouse xenografts, indotricarbocyanine-octreotate accumulated in tumor tissue. Tumor fluorescence rapidly increased and was more than threefold higher than that of normal tissue from 3 to 24 h after application. The targeting conjugate was also specifically internalized by primary human neuroendocrine tumor cells. This imaging approach, combining the specificity of ligand/receptor interaction with near-infrared fluorescence detection, may be applied in various other fields of cancer diagnosis. PMID- 11283590 TI - A biodegradable polymer as a cytokine delivery system for inducing bone formation. AB - Bone morphogenetic proteins (BMPs) that have the potential to elicit new bone in vivo have been used in a tissue-engineering approach for the repair of bone injuries and bone defects. Although it is now possible to generate large amounts of recombinant human (rh) BMPs for medical use, the major challenge remains in the development of optimal local delivery systems for these proteins. Here we describe the development of a synthetic biodegradable polymer, poly-d,l-lactic acid-p-dioxanone-polyethylene glycol block copolymer (PLA-DX-PEG). This polymer exhibits promising degradation characteristics for BMP delivery systems and good biocompatibility under test conditions. PLA-DX-PEG/rhBMP-2 composite implants induced ectopic new bone formation effectively when tested in vivo, and can repair large bone defects orthotopically. This polymeric delivery system represents an advance in the technology for the enhancement of bone repair. PMID- 11283591 TI - Immobilized RNA switches for the analysis of complex chemical and biological mixtures. AB - A prototype biosensor array has been assembled from engineered RNA molecular switches that undergo ribozyme-mediated self-cleavage when triggered by specific effectors. Each type of switch is prepared with a 5'-thiotriphosphate moiety that permits immobilization on gold to form individually addressable pixels. The ribozymes comprising each pixel become active only when presented with their corresponding effector, such that each type of switch serves as a specific analyte sensor. An addressed array created with seven different RNA switches was used to report the status of targets in complex mixtures containing metal ion, enzyme cofactor, metabolite, and drug analytes. The RNA switch array also was used to determine the phenotypes of Escherichia coli strains for adenylate cyclase function by detecting naturally produced 3',5'- cyclic adenosine monophosphate (cAMP) in bacterial culture media. PMID- 11283592 TI - Expression profiling using microarrays fabricated by an ink-jet oligonucleotide synthesizer. AB - We describe a flexible system for gene expression profiling using arrays of tens of thousands of oligonucleotides synthesized in situ by an ink-jet printing method employing standard phosphoramidite chemistry. We have characterized the dependence of hybridization specificity and sensitivity on parameters including oligonucleotide length, hybridization stringency, sequence identity, sample abundance, and sample preparation method. We find that 60-mer oligonucleotides reliably detect transcript ratios at one copy per cell in complex biological samples, and that ink-jet arrays are compatible with several different sample amplification and labeling techniques. Furthermore, results using only a single carefully selected oligonucleotide per gene correlate closely with those obtained using complementary DNA (cDNA) arrays. Most of the genes for which measurements differ are members of gene families that can only be distinguished by oligonucleotides. Because different oligonucleotide sequences can be specified for each array, we anticipate that ink-jet oligonucleotide array technology will be useful in a wide variety of DNA microarray applications. PMID- 11283593 TI - A motif-based profile scanning approach for genome-wide prediction of signaling pathways. AB - The rapid increase in genomic information requires new techniques to infer protein function and predict protein-protein interactions. Bioinformatics identifies modular signaling domains within protein sequences with a high degree of accuracy. In contrast, little success has been achieved in predicting short linear sequence motifs within proteins targeted by these domains to form complex signaling networks. Here we describe a peptide library-based searching algorithm, accessible over the World Wide Web, that identifies sequence motifs likely to bind to specific protein domains such as 14-3-3, SH2, and SH3 domains, or likely to be phosphorylated by specific protein kinases such as Src and AKT. Predictions from database searches for proteins containing motifs matching two different domains in a common signaling pathway provides a much higher success rate. This technology facilitates prediction of cell signaling networks within proteomes, and could aid in the identification of drug targets for the treatment of human diseases. PMID- 11283594 TI - DNA shuffling method for generating highly recombined genes and evolved enzymes. AB - We introduce a method of in vitro recombination or "DNA shuffling" to generate libraries of evolved enzymes. The approach relies on the ordering, trimming, and joining of randomly cleaved parental DNA fragments annealed to a transient polynucleotide scaffold. We generated chimeric libraries averaging 14.0 crossovers per gene, a several-fold higher level of recombination than observed for other methods. We also observed an unprecedented four crossovers per gene in regions of 10 or fewer bases of sequence identity. These properties allow generation of chimeras unavailable by other methods. We detected no unshuffled parental clones or duplicated "sibling" chimeras, and relatively few inactive clones. We demonstrated the method by molecular breeding of a monooxygenase for increased rate and extent of biodesulfurization on complex substrates, as well as for 20-fold faster conversion of a nonnatural substrate. This method represents a conceptually distinct and improved alternative to sexual PCR for gene family shuffling. PMID- 11283595 TI - Bactericidal antisense effects of peptide-PNA conjugates. AB - Antisense peptide nucleic acids (PNAs) can specifically inhibit Escherichia coli gene expression and growth and hold promise as anti-infective agents and as tools for microbial functional genomics. Here we demonstrate that chemical modification improves the potency of standard PNAs. We show that 9- to 12-mer PNAs, especially when attached to the cell wall/membrane-active peptide KFFKFFKFFK, provide improvements in antisense potency in E. coli amounting to two orders of magnitude while retaining target specificity. Peptide-PNA conjugates targeted to ribosomal RNA (rRNA) and to messenger RNA (mRNA) encoding the essential fatty acid biosynthesis protein Acp prevented cell growth. The anti-acpP PNA at 2 microM concentration cured HeLa cell cultures noninvasively infected with E. coli K12 without any apparent toxicity to the human cells. These results indicate that peptides can be used to carry antisense PNA agents into bacteria. Such peptide PNA conjugates open exciting possibilities for anti-infective drug development and provide new tools for microbial genetics. PMID- 11283596 TI - Single-mismatch detection using gold-quenched fluorescent oligonucleotides. AB - Here we describe a hybrid material composed of a single-stranded DNA (ssDNA) molecule, a 1.4 nm diameter gold nanoparticle, and a fluorophore that is highly quenched by the nanoparticle through a distance-dependent process. The fluorescence of this hybrid molecule increases by a factor of as much as several thousand as it binds to a complementary ssDNA. We show that this composite molecule is a different type of molecular beacon with a sensitivity enhanced up to 100-fold. In competitive hybridization assays, the ability to detect single mismatch is eightfold greater with this probe than with other molecular beacons. PMID- 11283597 TI - Suppression of a P450 hydroxylase gene in plant trichome glands enhances natural product-based aphid resistance. AB - Trichome glands on the surface of many higher plants produce and secrete exudates affecting insects, microbes, and herbivores. Metabolic engineering of gland exudation has potential for improving pest/disease resistance, and for facilitating molecular farming. We identified a cytochrome P450 hydroxylase gene specific to the trichome gland and used both antisense and sense co-suppression strategies to investigate its function. P450-suppressed transgenic tobacco plants showed a > or =41% decrease in the predominant exudate component, cembratriene diol (CBT-diol), and a > or =19-fold increase in its precursor, cembratriene-ol (CBT-ol). Thus, the level of CBT-ol was raised from 0.2 to > or =4.3% of leaf dry weight. Exudate from antisense-expressing plants had higher aphidicidal activity, and transgenic plants with exudate containing high concentrations of CBT-ol showed greatly diminished aphid colonization responses. Our results demonstrate the feasibility of significantly modifying the natural-product chemical composition and aphid-interactive properties of gland exudates using metabolic engineering. The results also have implications for molecular farming. PMID- 11283598 TI - A systematic approach to the analysis of protein phosphorylation. AB - Reversible protein phosphorylation has been known for some time to control a wide range of biological functions and activities. Thus determination of the site(s) of protein phosphorylation has been an essential step in the analysis of the control of many biological systems. However, direct determination of individual phosphorylation sites occurring on phosphoproteins in vivo has been difficult to date, typically requiring the purification to homogeneity of the phosphoprotein of interest before analysis. Thus, there has been a substantial need for a more rapid and general method for the analysis of protein phosphorylation in complex protein mixtures. Here we describe such an approach to protein phosphorylation analysis. It consists of three steps: (1) selective phosphopeptide isolation from a peptide mixture via a sequence of chemical reactions, (2) phosphopeptide analysis by automated liquid chromatography-tandem mass spectrometry (LC-MS/MS), and (3) identification of the phosphoprotein and the phosphorylated residue(s) by correlation of tandem mass spectrometric data with sequence databases. By utilizing various phosphoprotein standards and a whole yeast cell lysate, we demonstrate that the method is equally applicable to serine-, threonine- and tyrosine-phosphorylated proteins, and is capable of selectively isolating and identifying phosphopeptides present in a highly complex peptide mixture. PMID- 11283599 TI - Enrichment analysis of phosphorylated proteins as a tool for probing the phosphoproteome. AB - The current progression from genomics to proteomics is fueled by the realization that many properties of proteins (e.g., interactions, post-translational modifications) cannot be predicted from DNA sequence. Although it has become feasible to rapidly identify proteins from crude cell extracts using mass spectrometry after two-dimensional electrophoretic separation, it can be difficult to elucidate low-abundance proteins of interest in the presence of a large excess of relatively abundant proteins. Therefore, for effective proteome analysis it becomes critical to enrich the sample to be analyzed in subfractions of interest. For example, the analysis of protein kinase substrates can be greatly enhanced by enriching the sample of phosphorylated proteins. Although enrichment of phosphotyrosine-containing proteins has been achieved through the use of high-affinity anti-phosphotyrosine antibodies, the enrichment of phosphoserine/threonine-containing proteins has not been routinely possible. Here, we describe a method for enriching phosphoserine/threonine-containing proteins from crude cell extracts, and for subsequently identifying the phosphoproteins and sites of phosphorylation. The method, which involves chemical replacement of the phosphate moieties by affinity tags, should be of widespread utility for defining signaling pathways and control mechanisms that involve phosphorylation or dephosphorylation of serine/threonine residues. PMID- 11283600 TI - Biotechnology patents in the food sectors. PMID- 11283601 TI - Recent patents in microarrays. PMID- 11283604 TI - What's hot in biotech employment. PMID- 11283605 TI - Phosphorylation of Snapin by PKA modulates its interaction with the SNARE complex. AB - cAMP-dependent protein kinase A (PKA) can modulate synaptic transmission by acting directly on unknown targets in the neurotransmitter secretory machinery. Here we identify Snapin, a protein of relative molecular mass 15,000 that is implicated in neurotransmission by binding to SNAP-25, as a possible target. Deletion mutation and site-directed mutagenetic experiments pinpoint the phosphorylation site to serine 50. PKA-phosphorylation of Snapin significantly increases its binding to synaptosomal-associated protein-25 (SNAP-25). Mutation of Snapin serine 50 to aspartic acid (S50D) mimics this effect of PKA phosphorylation and enhances the association of synaptotagmin with the soluble N ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex. Furthermore, treatment of rat hippocampal slices with nonhydrolysable cAMP analogue induces in vivo phosphorylation of Snapin and enhances the interaction of both Snapin and synaptotagmin with the SNARE complex. In adrenal chromaffin cells, overexpression of the Snapin S50D mutant leads to an increase in the number of release-competent vesicles. Our results indicate that Snapin may be a PKA target for modulating transmitter release through the cAMP-dependent signal transduction pathway. PMID- 11283606 TI - Membrane blebbing during apoptosis results from caspase-mediated activation of ROCK I. AB - The execution phase of apoptosis is characterized by marked changes in cell morphology that include contraction and membrane blebbing. The actin-myosin system has been proposed to be the source of contractile force that drives bleb formation, although the biochemical pathway that promotes actin-myosin contractility during apoptosis has not been identified. Here we show that the Rho effector protein ROCK I, which contributes to phosphorylation of myosin light chains, myosin ATPase activity and coupling of actin-myosin filaments to the plasma membrane, is cleaved during apoptosis to generate a truncated active form. The activity of ROCK proteins is both necessary and sufficient for formation of membrane blebs and for re-localization of fragmented DNA into blebs and apoptotic bodies. PMID- 11283607 TI - Caspase-3-mediated cleavage of ROCK I induces MLC phosphorylation and apoptotic membrane blebbing. AB - Increased phosphorylation of myosin light chain (MLC) is necessary for the dynamic membrane blebbing that is observed at the onset of apoptosis. Here we identify ROCK I, an effector of the small GTPase Rho, as a new substrate for caspases. ROCK I is cleaved by caspase-3 at a conserved DETD1113/G sequence and its carboxy-terminal inhibitory domain is removed, resulting in deregulated and constitutive kinase activity. ROCK proteins are known to regulate MLC phosphorylation, and apoptotic cells exhibit a gradual increase in levels of phosphorylated MLC concomitant with ROCK I cleavage. This phosphorylation, as well as membrane blebbing, is abrogated by inhibition of caspases or ROCK proteins, but both processes are independent of Rho activity. We also show that expression of active truncated ROCK I induces cell blebbing. Thus, activation of ROCK I by caspase-3 seems to be responsible for bleb formation in apoptotic cells. PMID- 11283608 TI - Spatial regulation of the exocyst complex by Rho1 GTPase. AB - Spatial regulation of membrane traffic is fundamental to many biological processes, including epithelial cell polarization and neuronal synaptogenesis. The multiprotein exocyst complex is localized to sites of polarized exocytosis, and is required for vesicle targeting and docking at specific domains of the plasma membrane. One component of the complex, Sec3, is thought to be a spatial landmark for polarized exocytosis. We have searched for proteins that regulate the polarized localization of the exocyst in the budding yeast Saccharomyces cerevisiae. Here we report that certain rho1 mutant alleles specifically affect the localization of the exocyst proteins. Sec3 interacts directly with Rho1 in its GTP-bound form, and functional Rho1 is needed both to establish and to maintain the polarized localization of Sec3. Sec3 is not the only mediator of the effect of Rho1 on the exocyst, because some members of the complex are correctly targeted independently of the interaction between Rho1 and Sec3. These results reveal the action of parallel pathways for the polarized localization of the exocytic machinery, both of which are under the control of Rho1, a master regulator of cell polarity. PMID- 11283609 TI - p190 RhoGAP is the principal Src substrate in brain and regulates axon outgrowth, guidance and fasciculation. AB - The Src tyrosine kinases have been implicated in several aspects of neural development and nervous system function; however, their relevant substrates in brain and their mechanism of action in neurons remain to be established clearly. Here we identify the potent Rho regulatory protein, p190 RhoGAP (GTPase activating protein), as the principal Src substrate detected in the developing and mature nervous system. We also find that mice lacking functional p190 RhoGAP exhibit defects in axon guidance and fasciculation. p190 RhoGAP is co-enriched with F-actin in the distal tips of axons, and overexpressing p190 RhoGAP in neuroblastoma cells promotes extensive neurite outgrowth, indicating that p190 RhoGAP may be an important regulator of Rho-mediated actin reorganization in neuronal growth cones. p190 RhoGAP transduces signals downstream of cell-surface adhesion molecules, and we find that p190-RhoGAP-mediated neurite outgrowth is promoted by the extracellular matrix protein laminin. Together with the fact that mice lacking neural adhesion molecules or Src kinases also exhibit defects in axon outgrowth, guidance and fasciculation, our results suggest that p190 RhoGAP mediates a Src-dependent adhesion signal for neuritogenesis to the actin cytoskeleton through the Rho GTPase. PMID- 11283610 TI - GTP-dependent segregation of H-ras from lipid rafts is required for biological activity. AB - Different sites of plasma membrane attachment may underlie functional differences between isoforms of Ras. Here we show that palmitoylation and farnesylation targets H-ras to lipid rafts and caveolae, but that the interaction of H-ras with these membrane subdomains is dynamic. GTP-loading redistributes H-ras from rafts into bulk plasma membrane by a mechanism that requires the adjacent hypervariable region of H-ras. Release of H-ras-GTP from rafts is necessary for efficient activation of Raf. By contrast, K-ras is located outside rafts irrespective of bound nucleotide. Our studies identify a novel protein determinant that is required for H-ras function, and show that the GTP/GDP state of H-ras determines its lateral segregation on the plasma membrane. PMID- 11283611 TI - Spinal muscular atrophy disrupts the interaction of ZPR1 with the SMN protein. AB - The survival motor neurons (smn) gene in mice is essential for embryonic viability. In humans, mutation of the telomeric copy of the SMN1 gene causes spinal muscular atrophy, an autosomal recessive disease. Here we report that the SMN protein interacts with the zinc-finger protein ZPR1 and that these proteins colocalize in small subnuclear structures, including gems and Cajal bodies. SMN and ZPR1 redistribute from the cytoplasm to the nucleus in response to serum. This process is disrupted in cells from patients with Werdnig-Hoffman syndrome (spinal muscular atrophy type I) that have SMN1 mutations. Similarly, decreased ZPR1 expression prevents SMN localization to nuclear bodies. Our data show that ZPR1 is required for the localization of SMN in nuclear bodies. PMID- 11283612 TI - Skp1 forms multiple protein complexes, including RAVE, a regulator of V-ATPase assembly. AB - SCF ubiquitin ligases are composed of Skp1, Cdc53, Hrt1 and one member of a large family of substrate receptors known as F-box proteins (FBPs). Here we report the identification, using sequential rounds of epitope tagging, affinity purification and mass spectrometry, of 16 Skp1 and Cdc53-associated proteins in budding yeast, including all components of SCF, 9 FBPs, Yjr033 (Rav1) and Ydr202 (Rav2). Rav1, Rav2 and Skp1 form a complex that we have named 'regulator of the (H+)-ATPase of the vacuolar and endosomal membranes' (RAVE), which associates with the V1 domain of the vacuolar membrane (H+)-ATPase (V-ATPase). V-ATPases are conserved throughout eukaryotes, and have been implicated in tumour metastasis and multidrug resistance, and here we show that RAVE promotes glucose-triggered assembly of the V-ATPase holoenzyme. Previous systematic genome-wide two-hybrid screens yielded 17 proteins that interact with Skp1 and Cdc53, only 3 of which overlap with those reported here. Thus, our results provide a distinct view of the interactions that link proteins into a comprehensive cellular network. PMID- 11283613 TI - Repression of p15INK4b expression by Myc through association with Miz-1. AB - Deregulated expression of c-myc can induce cell proliferation in established cell lines and in primary mouse embryonic fibroblasts (MEFs), through a combination of both transcriptional activation and repression by Myc. Here we show that a Myc associated transcription factor, Miz-1, arrests cells in G1 phase and inhibits cyclin D-associated kinase activity. Miz-1 upregulates expression of the cyclin dependent kinases (CDK) inhibitor p15INK4b by binding to the initiator element of the p15INK4b promoter. Myc and Max form a complex with Miz-1 at the p15 initiator and inhibit transcriptional activation by Miz-1. Expression of Myc in primary cells inhibits the accumulation of p15INK4b that is associated with cellular senescence; conversely, deletion of c-myc in an established cell line activates p15INK4b expression. Alleles of c-myc that are unable to bind to Miz-1 fail to inhibit accumulation of p15INK4b messenger RNA in primary cells and are, as a consequence, deficient in immortalization. PMID- 11283614 TI - TGFbeta influences Myc, Miz-1 and Smad to control the CDK inhibitor p15INK4b. AB - Transforming growth factor-beta (TGFbeta) is a cytokine that arrests epithelial cell division by switching off the proto-oncogene c-myc and rapidly switching on cyclin-dependent kinase (CDK) inhibitors such as p15INK4b. Gene responses to TGFbeta involve Smad transcription factors that are directly activated by the TGFbeta receptor. Why downregulation of c-myc expression by TGFbeta is required for rapid activation of p15INK4b has remained unknown. Here we provide evidence that TGFbeta signalling prevents recruitment of Myc to the p15INK4b transcriptional initiator by Myc-interacting zinc-finger protein 1 (Miz-1). This relieves repression and enables transcriptional activation by a TGFbeta-induced Smad protein complex that recognizes an upstream p15INK4b promoter region and contacts Miz-1. Thus, two separate TGFbeta-dependent inputs - Smad-mediated transactivation and relief of repression by Myc - keep tight control over p15INK4b activation. PMID- 11283615 TI - Regulation of death receptor expression and TRAIL/Apo2L-induced apoptosis by NF kappaB. AB - TRAIL (tumour-necrosis factor-related apoptosis ligand or Apo2L) triggers apoptosis through engagement of the death receptors TRAIL-R1 (also known as DR4) and TRAIL-R2 (DR5). Here we show that the c-Rel subunit of the transcription factor NF-kappaB induces expression of TRAIL-R1 and TRAIL-R2; conversely, a transdominant mutant of the inhibitory protein IkappaBalpha or a transactivation deficient mutant of c-Rel reduces expression of either death receptor. Whereas NF kappaB promotes death receptor expression, cytokine-mediated activation of the RelA subunit of NF-kappaB also increases expression of the apoptosis inhibitor, Bcl-xL, and protects cells from TRAIL. Inhibition of NF-kappaB by blocking activation of the IkappaB kinase complex reduces Bcl-x L expression and sensitizes tumour cells to TRAIL-induced apoptosis. The ability to induce death receptors or Bcl-xL may explain the dual roles of NF-kappaB as a mediator or inhibitor of cell death during immune and stress responses. PMID- 11283616 TI - A role for the Pkc1p/Mpk1p kinase cascade in the morphogenesis checkpoint. AB - In many cells the timing of entry into mitosis is controlled by the balance between the activity of inhibitory Wee1-related kinases (Swe1p in budding yeast) and the opposing effect of Cdc25-related phosphatases (Mih1p in budding yeast) that act on the cyclin-dependent kinase Cdc2 (Cdc28p in budding yeast). Wee1 and Cdc25 are key elements in the G2 arrest mediated by diverse checkpoint controls. In budding yeast, a 'morphogenesis checkpoint' that involves Swe1p and Mih1p delays mitotic activation of Cdc28p. Many environmental stresses (such as shifts in temperature or osmolarity) provoke transient depolarization of the actin cytoskeleton, during which bud construction is delayed while cells adapt to environmental conditions. During this delay, the morphogenesis checkpoint halts the cell cycle in G2 phase until actin can repolarize and complete bud construction, thus preventing the generation of binucleate cells. A similar G2 delay can be triggered by mutations or drugs that specifically impair actin organization, indicating that it is probably actin disorganization itself, rather than specific environmental stresses, that triggers the delay. The G2 delay involves stabilization of Swe1p in response to various actin perturbations, although this alone is insufficient to produce a long G2 delay. PMID- 11283617 TI - Polo kinase and Asp are needed to promote the mitotic organizing activity of centrosomes. AB - Interfering with the activity of polo-like kinases can lead to the formation of monopolar spindles. Polo-like kinases also regulate mitotic entry, activation of the anaphase-promoting complex and the necessary preconditions for cytokinesis. Similarities between the phenotypes of the Drosophila mutants asp and polo point towards a common role in spindle pole function. The abnormal spindles of asp mutants are bipolar but have disorganized broad poles at which gamma-tubulin has an abnormal distribution. Moreover, the synergism or of polo1 aspE3 double mutants indicates a possible involvement of these genes in a common process. Asp is a microtubule-associated protein of relative molecular mass 220,000 (Mr 220K) found at the face of the centrosome that contacts spindle microtubules. In partially purified centrosomes, it is required with gamma-tubulin to organize microtubule asters. Here, we show that Asp is the previously identified Mr 220K substrate of Polo kinase. Polo phosphorylates Asp in vitro, converting it into an MPM2 epitope. Polo and Asp proteins immunoprecipitate together and exist as part of a 25-38S complex. Extracts of polo-derived embryos are unable to restore the ability of salt-stripped centrosomes to nucleate microtubule asters. This can be rescued by addition of phosphorylated Asp or active Polo kinase. PMID- 11283618 TI - Substeps within the 8-nm step of the ATPase cycle of single kinesin molecules. AB - Kinesin is a molecular motor that moves processively by regular 8-nm steps along microtubules. The processivity of this movement is explained by a hand-over-hand model in which the two heads of kinesin work in a coordinated manner. One head remains bound to the microtubule while the other steps from the alphabeta-tubulin dimer behind the attached head to the dimer in front. The overall movement is 8 nm per ATPase cycle. To investigate elementary processes within the 8-nm step, we have developed a new assay that resolves nanometre displacements of single kinesin molecules with microsecond accuracy. Our data show that the 8-nm step can be resolved into fast and slow substeps, each corresponding to a displacement of approximately 4 nm. The substeps are most probably generated by structural changes in one head of kinesin, leading to rectified forward thermal motions of the partner head. It is also possible that the kinesin steps along the 4-nm repeat of tubulin monomers. PMID- 11283619 TI - A role for the Adenomatous Polyposis Coli protein in chromosome segregation. AB - Mutations in the Adenomatous Polyposis Coli (APC) gene are responsible for familial colon cancer and also occur in the early stages of sporadic colon cancer. APC functions in the Wnt signalling pathway to regulate the degradation of beta-catenin (reviewed in refs 1-3). APC also binds to and stabilizes microtubules in vivo and in vitro, localizes to clusters at the ends of microtubules near the plasma membrane of interphase cells, and is an important regulator of cytoskeletal function. Here we show that cells carrying a truncated APC gene (Min) are defective in chromosome segregation. Moreover, during mitosis, APC localizes to the ends of microtubules embedded in kinetochores and forms a complex with the checkpoint proteins Bub1 and Bub3. In vitro, APC is a high affinity substrate for Bub kinases. Our data are consistent with a role for APC in kinetochore-microtubule attachment and suggest that truncations in APC that eliminate microtubule binding may contribute to chromosomal instability in cancer cells. PMID- 11283620 TI - Mutations in the APC tumour suppressor gene cause chromosomal instability. AB - Two forms of genetic instability have been described in colorectal cancer: microsatellite instability and chromosomal instability. Microsatellite instability results from mutations in mismatch repair genes; chromosomal instability is the hallmark of many colorectal cancers, although it is not completely understood at the molecular level. As truncations of the Adenomatous Polyposis Coli (APC) gene are found in most colorectal tumours, we thought that mutations in APC might be responsible for chromosomal instability. To test this hypothesis, we examined mouse embryonic stem (ES) cells homozygous for Min (multiple intestinal neoplasia) or Apc1638T alleles. Here we show that Apc mutant ES cells display extensive chromosome and spindle aberrations, providing genetic evidence for a role of APC in chromosome segregation. Consistent with this, APC accumulates at the kinetochore during mitosis. Apc mutant cells form mitotic spindles with an abundance of microtubules that inefficiently connect with kinetochores. This phenotype is recapitulated by the induced expression of a 253 amino-acid carboxy-terminal fragment of APC in microsatellite unstable colorectal cancer cells. We conclude that loss of APC sequences that lie C-terminal to the beta-catenin regulatory domain contributes to chromosomal instability in colorectal cancer. PMID- 11283622 TI - Breaking the language barrier. AB - English has undoubtedly become the universal language of science. But how did this come about and what are the potential benefits? PMID- 11283624 TI - Problems with LAP nomenclature. PMID- 11283625 TI - Apoptosis: caspases orchestrate the ROCK 'n' bleb. PMID- 11283626 TI - Stressing endocytosis. PMID- 11283627 TI - The survival motor neurons protein uses its ZPR for nuclear localization. PMID- 11283628 TI - p27 destruction: Cks1 pulls the trigger. PMID- 11283634 TI - Prospects for Parkinson disease. PMID- 11283635 TI - Circulating smooth muscle progenitor cells contribute to atherosclerosis. PMID- 11283636 TI - Differential hepatocyte toxicity of recombinant Apo2L/TRAIL versions. PMID- 11283638 TI - Scientists battle obesity overload. PMID- 11283637 TI - Avoiding premature apoptosis of normal epidermal cells. PMID- 11283639 TI - HPV vaccine moves into late stage trials. PMID- 11283644 TI - Companies begin tests of HIV vaccines. PMID- 11283641 TI - GSK renews malaria efforts. GlaxoSmithKline Biologicals. PMID- 11283645 TI - South Africa vs. big pharma. PMID- 11283650 TI - Klaus Rajewsky. PMID- 11283651 TI - Can stem cells cross lineage boundaries? PMID- 11283652 TI - Embryonic stem cell research. The case for... PMID- 11283653 TI - Embryonic stem cell research. The case against... PMID- 11283656 TI - Too many vessels? Not enough? The wrong kind? The VEGF debate continues. PMID- 11283657 TI - A new latent HIV reservoir. PMID- 11283658 TI - Connecting Fanconi anemia to BRCA1. PMID- 11283659 TI - Differentiation or leukemia: is C/EBPalpha the answer? PMID- 11283660 TI - Profiling familial breast cancer. PMID- 11283661 TI - A role for complement in transmissible spongiform encephalopathies. PMID- 11283662 TI - Helping the heart to heal with stem cells. PMID- 11283663 TI - Arch enemy. PMID- 11283664 TI - COX-2 as a multifunctional neuronal modulator. PMID- 11283665 TI - The vicious cyclin of addiction. PMID- 11283667 TI - Therapeutic opportunities in polyglutamine disease. AB - Polyglutamine diseases comprise a class of familial neurodegenerative disorders caused by expression of proteins containing expanded polyglutamine tracts. Great progress has been made in elucidating the molecular mechanisms contributing to polyglutamine pathology, and in identifying potential drug targets. Although much remains to be learned, these advances provide an opportunity for rational approaches to target-based drug discovery. PMID- 11283668 TI - Vascular endothelial growth factor enhances atherosclerotic plaque progression. AB - Vascular endothelial growth factor (VEGF) can promote angiogenesis but may also exert certain effects to alter the rate of atherosclerotic plaque development. To evaluate this potential impact on plaque progression, we treated cholesterol-fed mice doubly deficient in apolipoprotein E/apolipoprotein B100 with low doses of VEGF (2 microg/kg) or albumin. VEGF significantly increased macrophage levels in bone marrow and peripheral blood and increased plaque area 5-, 14- and 4-fold compared with controls at weeks 1, 2 and 3, respectively. Plaque macrophage and endothelial cell content also increased disproportionately over controls. In order to confirm that the VEGF-mediated plaque progression was not species specific, the experiment was repeated in cholesterol-fed rabbits at the three week timepoint, which showed comparable increases in plaque progression. PMID- 11283669 TI - Neovascularization of ischemic myocardium by human bone-marrow-derived angioblasts prevents cardiomyocyte apoptosis, reduces remodeling and improves cardiac function. AB - Left ventricular remodeling is a major cause of progressive heart failure and death after myocardial infarction. Although neoangiogenesis within the infarcted tissue is an integral component of the remodeling process, the capillary network is unable to support the greater demands of the hypertrophied myocardium, resulting in progressive loss of viable tissue, infarct extension and fibrous replacement. Here we show that bone marrow from adult humans contains endothelial precursors with phenotypic and functional characteristics of embryonic hemangioblasts, and that these can be used to directly induce new blood vessel formation in the infarct-bed (vasculogenesis) and proliferation of preexisting vasculature (angiogenesis) after experimental myocardial infarction. The neoangiogenesis resulted in decreased apoptosis of hypertrophied myocytes in the peri-infarct region, long-term salvage and survival of viable myocardium, reduction in collagen deposition and sustained improvement in cardiac function. The use of cytokine-mobilized autologous human bone-marrow-derived angioblasts for revascularization of infarcted myocardium (alone or in conjunction with currently used therapies) has the potential to significantly reduce morbidity and mortality associated with left ventricular remodeling. PMID- 11283670 TI - Histone deacetylases induce angiogenesis by negative regulation of tumor suppressor genes. AB - Low oxygen tension influences tumor progression by enhancing angiogenesis; and histone deacetylases (HDAC) are implicated in alteration of chromatin assembly and tumorigenesis. Here we show induction of HDAC under hypoxia and elucidate a role for HDAC in the regulation of hypoxia-induced angiogenesis. Overexpressed wild-type HDAC1 downregulated expression of p53 and von Hippel-Lindau tumor suppressor genes and stimulated angiogenesis of human endothelial cells. A specific HDAC inhibitor, trichostatin A (TSA), upregulated p53 and von Hippel Lindau expression and downregulated hypoxia-inducible factor-1alpha and vascular endothelial growth factor. TSA also blocked angiogenesis in vitro and in vivo. TSA specifically inhibited hypoxia-induced angiogenesis in the Lewis lung carcinoma model. These results indicate that hypoxia enhances HDAC function and that HDAC is closely involved in angiogenesis through suppression of hypoxia responsive tumor suppressor genes. PMID- 11283671 TI - AML1-ETO downregulates the granulocytic differentiation factor C/EBPalpha in t(8;21) myeloid leukemia. AB - The transcription factor CCAAT/enhancer binding protein alpha, or C/EBPalpha, encoded by the CEBPA gene, is crucial for the differentiation of granulocytes. Conditional expression of C/EBPalpha triggers neutrophilic differentiation, and Cebpa knockout mice exhibit an early block in maturation. Dominant-negative mutations of CEBPA have been found in some patients with acute myeloid leukemia (AML), but not in AML with the t(8;21) translocation which gives rise to the fusion gene RUNX1-CBF2T1 (also known as AML1-ETO) encoding the AML1-ETO fusion protein. RUNX1-CBF2T1 positive-AML blasts had eight-fold lower CEBPA RNA levels and undetectable C/EBPalpha protein levels compared with other subgroups of AML patients. Conditional expression of RUNX1-CBF2T1 in U937 cells downregulated CEBPA mRNA, protein and DNA binding activity. AML1-ETO appears to suppress C/EBPalpha expression indirectly by inhibiting positive autoregulation of the CEBPA promoter. Conditional expression of C/EBPalpha in AML1-ETO-positive Kasumi 1 cells results in neutrophilic differentiation. We suggest that restoring C/EBPalpha expression will have therapeutic implications in RUNX1-CBF2T1-positive leukemias. PMID- 11283672 TI - Therapy of human tumors in NOD/SCID mice with patient-derived reactivated memory T cells from bone marrow. AB - In an analysis of 84 primary-operated breast cancer patients and 11 healthy donors, we found that the bone marrow of most patients contained memory T cells with specificity for tumor-associated antigens. Patients' bone marrow and peripheral blood contained CD8+ T cells that specifically bound HLA/peptide tetramers. In short-term culture with autologous dendritic cells pre-pulsed with tumor lysates, patients' memory T cells from bone marrow (but not peripheral blood) could be specifically reactivated to interferon-gamma-producing and cytotoxic effector cells. A single transfer of restimulated bone-marrow T cells into NOD/SCID mice caused regression of autologous tumor xenotransplants associated with infiltration by human T cells and tumor-cell apoptosis and necrosis. T cells from peripheral blood showed much lower anti-tumor reactivity. Our findings reveal an innate, specific recognition of breast cancer antigens and point to a possible novel cancer therapy using patients' bone-marrow-derived memory T cells. PMID- 11283673 TI - Generation of HIV latency during thymopoiesis. AB - The use of combination antiretroviral therapy results in a substantial reduction in viremia, a rebound of CD4+ T cells and increased survival for HIV-infected individuals. However, this treatment does not result in the total eradication of HIV. Rather, the virus is thought to remain latent in a subset of cells, where it avoids elimination by the immune system. In this state the virus is capable of reactivation of productive infection following cessation of therapy. These latently infected cells are very few in number and it has thus been difficult to determine their origin and to study the molecular nature of the latent viral genome. HIV replication is linked to cellular gene transcription and requires target cell activation. Therefore, should an activated, infected cell become transcriptionally inactive prior to cytopathic effects, the viral genome might be maintained in a latent state. We used the SCID-hu (Thy/Liv) mouse model to establish that activation-inducible HIV can be generated at high frequency during thymopoiesis, a process where previously activated cells mature towards quiescence. Moreover, we showed that these cells can be exported into the periphery where the virus remains latent until T-cell receptor stimulation, indicating that the thymus might be a source of latent HIV in humans. PMID- 11283674 TI - Functional alpha4-integrin: a newly identified pathway of neutrophil recruitment in critically ill septic patients. AB - Using a novel flow chamber assay system and whole blood, we show that leukocytes from septic individuals have a four-fold elevation of adhesion, but not rolling, on a P-selectin/beta2-integrin substrate. Most leukocytes from septic patients (but not healthy controls) that bound vascular cell adhesion molecule 1 (VCAM-1) were neutrophils. All adhesion was inhibited with an antibody specific for the VCAM-1 ligand alpha4-integrin. The alpha4-integrin was present on neutrophils from septic patients but not on neutrophils from patients with localized bacterial infections. The plasma milieu of septic patients was sufficient to induce neutrophils from healthy subjects to bind VCAM-1 under flow conditions. This is the first description of alpha4-integrin/VCAM-1 pathway of neutrophil recruitment in human disease. This pathway may provide a new therapeutic target to reduce inappropriate neutrophil adhesion without altering the normal yet critical beta2-integrin-mediated adhesive function of neutrophils. PMID- 11283675 TI - Targeting acute ischemic stroke with a calcium-sensitive opener of maxi-K potassium channels. AB - During ischemic stroke, neurons at risk are exposed to pathologically high levels of intracellular calcium (Ca++), initiating a fatal biochemical cascade. To protect these neurons, we have developed openers of large-conductance, Ca++ activated (maxi-K or BK) potassium channels, thereby augmenting an endogenous mechanism for regulating Ca++ entry and membrane potential. The novel fluoro oxindoles BMS-204352 and racemic compound 1 are potent, effective and uniquely Ca++-sensitive openers of maxi-K channels. In rat models of permanent large vessel stroke, BMS-204352 provided significant levels of cortical neuroprotection when administered two hours after the onset of occlusion, but had no effects on blood pressure or cerebral blood flow. This novel approach may restrict Ca++ entry in neurons at risk while having minimal side effects. PMID- 11283676 TI - Lysosomal ceroid depletion by drugs: therapeutic implications for a hereditary neurodegenerative disease of childhood. AB - Neuronal ceroid lipofuscinoses (NCLs) are the most common hereditary neurodegenerative diseases of childhood. The infantile form, INCL, is caused by lysosomal palmitoyl-protein thioesterase (PPT) deficiency, which impairs the cleavage of thioester linkages in palmitoylated proteins, preventing their hydrolysis by lysosomal proteinases. Consequent accumulation of these lipid modified proteins (constituents of ceroid) in lysosomes leads to INCL. Because thioester linkages are susceptible to nucleophilic attack, drugs with this property may have therapeutic potential for INCL. We report here that two such drugs, phosphocysteamine and N-acetylcysteine, disrupt thioester linkages in a model thioester compound, [14C]palmitoyl approximately CoA. Most importantly, in lymphoblasts derived from INCL patients, phosphocysteamine, a known lysosomotrophic drug, mediates the depletion of lysosomal ceroids, prevents their re-accumulation and inhibits apoptosis. Our results define a novel pharmacological approach to lysosomal ceroid depletion and raise the possibility that nucleophilic drugs such as phosphocysteamine hold therapeutic potential for INCL. PMID- 11283677 TI - Temporary depletion of complement component C3 or genetic deficiency of C1q significantly delays onset of scrapie. AB - Following peripheral exposure to transmissible spongiform encephalopathies (TSEs), infectivity usually accumulates in lymphoid tissues before neuroinvasion. The host prion protein (PrPc) is critical for TSE agent replication and accumulates as an abnormal, detergent insoluble, relatively proteinase-resistant isoform (PrPSc) in diseased tissues. Early PrPSc accumulation takes place on follicular dendritic cells (FDCs) within germinal centers in lymphoid tissues of patients with variant Creutzfeldt-Jakob disease (vCJD), sheep with natural scrapie or rodents following experimental peripheral infection with scrapie. In mouse scrapie models, the absence of FDCs blocks scrapie replication and PrPSc accumulation in the spleen, and neuroinvasion is significantly impaired. The mechanisms by which the TSE agent initially localizes to lymphoid follicles and interacts with FDCs are unknown. Antigens are trapped and retained on the surface of FDCs through interactions between complement and cellular complement receptors. Here we show that in mice, both temporary depletion of complement component C3 or genetic deficiency of C1q significantly delays the onset of disease following peripheral infection, and reduces the early accumulation of PrPSc in the spleen. Thus, in the early stages of infection, C3 and perhaps C1q contribute to the localization of TSE infectivity in lymphoid tissue and may be therapeutic targets. PMID- 11283679 TI - Imaging mass spectrometry: a new technology for the analysis of protein expression in mammalian tissues. PMID- 11283678 TI - Complement facilitates early prion pathogenesis. AB - New-variant Creutzfeldt-Jakob disease and scrapie are typically initiated by extracerebral exposure to the causative agent, and exhibit early prion replication in lymphoid organs. In mouse scrapie, depletion of B-lymphocytes prevents neuropathogenesis after intraperitoneal inoculation, probably due to impaired lymphotoxin-dependent maturation of follicular dendritic cells (FDCs), which are a major extracerebral prion reservoir. FDCs trap immune complexes with Fc-gamma receptors and C3d/C4b-opsonized antigens with CD21/CD35 complement receptors. We examined whether these mechanisms participate in peripheral prion pathogenesis. Depletion of circulating immunoglobulins or of individual Fc-gamma receptors had no effect on scrapie pathogenesis if B-cell maturation was unaffected. However, mice deficient in C3, C1q, Bf/C2, combinations thereof or complement receptors were partially or fully protected against spongiform encephalopathy upon intraperitoneal exposure to limiting amounts of prions. Splenic accumulation of prion infectivity and PrPSc was delayed, indicating that activation of specific complement components is involved in the initial trapping of prions in lymphoreticular organs early after infection. PMID- 11283680 TI - Subtelomeric chromosome rearrangements are detected using an innovative 12-color FISH assay (M-TEL). PMID- 11283681 TI - Ultrahigh-resolution ophthalmic optical coherence tomography. PMID- 11283697 TI - Phenotype-genotype relationships in monogenic disease: lessons from the thalassaemias. AB - The remarkable phenotypic diversity of the beta-thalassaemias reflects the heterogeneity of mutations at the beta-globin locus, the action of many secondary and tertiary modifiers, and a wide range of environmental factors. It is likely that phenotype-genotype relationships will be equally complex in the case of many monogenic diseases. These findings highlight the problems that might be encountered in defining the relationship between the genome and the environment in multifactorial disorders, in which the degree of heritability might be relatively low and several environmental agents are involved. They also emphasize the value of an understanding of phenotype-genotype relationships in designing approaches to gene therapy. PMID- 11283698 TI - The evolution and genetics of innate immunity. AB - The immune system provides protection from a wide range of pathogens. One component of immunity, the phylogenetically ancient innate immune response, fights infections from the moment of first contact and is the fundamental defensive weapon of multicellular organisms. The Toll family of receptors has a crucial role in immune defence. Studies in fruitflies and in mammals reveal that the defensive strategies of invertebrates and vertebrates are highly conserved at the molecular level, which raises the exciting prospects of an increased understanding of innate immunity. PMID- 11283699 TI - Computational studies of gene regulatory networks: in numero molecular biology. AB - Remarkable progress in genomic research is leading to a complete map of the building blocks of biology. Knowledge of this map is, in turn, setting the stage for a fundamental description of cellular function at the DNA level. Such a description will entail an understanding of gene regulation, in which proteins often regulate their own production or that of other proteins in a complex web of interactions. The implications of the underlying logic of genetic networks are difficult to deduce through experimental techniques alone, and successful approaches will probably involve the union of new experiments and computational modelling techniques. PMID- 11283700 TI - To err (meiotically) is human: the genesis of human aneuploidy. AB - Aneuploidy (trisomy or monosomy) is the most commonly identified chromosome abnormality in humans, occurring in at least 5% of all clinically recognized pregnancies. Most aneuploid conceptuses perish in utero, which makes this the leading genetic cause of pregnancy loss. However, some aneuploid fetuses survive to term and, as a class, aneuploidy is the most common known cause of mental retardation. Despite the devastating clinical consequences of aneuploidy, relatively little is known of how trisomy and monosomy originate in humans. However, recent molecular and cytogenetic approaches are now beginning to shed light on the non-disjunctional processes that lead to aneuploidy. PMID- 11283701 TI - Chromosome territories, nuclear architecture and gene regulation in mammalian cells. AB - The expression of genes is regulated at many levels. Perhaps the area in which least is known is how nuclear organization influences gene expression. Studies of higher-order chromatin arrangements and their dynamic interactions with other nuclear components have been boosted by recent technical advances. The emerging view is that chromosomes are compartmentalized into discrete territories. The location of a gene within a chromosome territory seems to influence its access to the machinery responsible for specific nuclear functions, such as transcription and splicing. This view is consistent with a topological model for gene regulation. PMID- 11283702 TI - Emerging technologies in yeast genomics. AB - The genomic revolution is undeniable: in the past year alone, the term 'genomics' was found in nearly 500 research articles, and at least 6 journals are devoted solely to genomic biology. More than just a buzzword, molecular biology has genuinely embraced genomics (the systematic, large-scale study of genomes and their functions). With its facile genetics, the budding yeast Saccharomyces cerevisiae has emerged as an important model organism in the development of many current genomic methodologies. These techniques have greatly influenced the manner in which biology is studied in yeast and in other organisms. In this review, we summarize the most promising technologies in yeast genomics. PMID- 11283703 TI - Legal and public policy issues in DNA forensics. AB - Since the 1980s, when DNA markers for identifying biological samples were first developed, the use of DNA evidence to convict defendants and to exonerate the wrongfully accused and wrongfully imprisoned has greatly increased. But the increase in databanks for storing DNA information on individuals convicted of certain crimes raises important legal and ethical issues on the use, collection and storage of DNA evidence. These issues have been the subject of a recent US National Commission, which will, hopefully, broaden public discourse about the future uses of DNA forensic technology. PMID- 11283704 TI - Solidarity and equity: new ethical frameworks for genetic databases. AB - Genetic database initiatives have given rise to considerable debate about their potential harms and benefits. The question arises as to whether existing ethical frameworks are sufficient to mediate between the competing interests at stake. One approach is to strengthen mechanisms for obtaining informed consent and for protecting confidentiality. However, there is increasing interest in other ethical frameworks, involving solidarity--participation in research for the common good--and the sharing of the benefits of research. PMID- 11283721 TI - The cap-to-tail guide to mRNA turnover. AB - The levels of cellular messenger RNA transcripts can be regulated by controlling the rate at which the mRNA decays. Because decay rates affect the expression of specific genes, they provide a cell with flexibility in effecting rapid change. Moreover, many clinically relevant mRNAs--including several encoding cytokines, growth factors and proto-oncogenes--are regulated by differential RNA stability. But what are the sequence elements and factors that control the half-lives of mRNAs? PMID- 11283722 TI - mRNA localization: message on the move. AB - Cytoplasmic messenger RNA localization is a key post-transcriptional mechanism of establishing spatially restricted protein synthesis. The characterization of cis acting signals within localized mRNAs, and the identification of trans-acting factors that recognize these signals, has opened avenues towards identifying the machinery and mechanisms involved in mRNA transport and localization. PMID- 11283723 TI - Tie receptors: new modulators of angiogenic and lymphangiogenic responses. AB - Angiogenesis is required for normal embryonic vascular development and aberrant angiogenesis contributes to several diseases, including cancer, diabetes and tissue ischaemia. What are the molecular mechanisms that regulate this important process? The Tie family of receptors and their ligands, the angiopoietins, are beginning to provide insight into how vessels make decisions such as whether to grow or regress--processes that are important not only during development but throughout an organism's life. PMID- 11283724 TI - A real-time view of life within 100 nm of the plasma membrane. AB - The plasma membrane is a two-dimensional compartment that relays most biological signals sent or received by a cell. Signalling involves membrane receptors and their associated enzyme cascades as well as organelles such as exocytic and endocytic vesicles. Advances in light microscope design, new organelle-specific vital stains and fluorescent proteins have renewed the interest in evanescent field fluorescence microscopy, a method uniquely suited to image the plasma membrane with its associated organelles and macromolecules in living cells. The method shows even the smallest vesicles made by cells, and can image the dynamics of single protein molecules. PMID- 11283725 TI - Cell signalling at the shoot meristem. AB - The regulation of cell differentiation at meristems is crucial to developmental patterning in plants. Rapid progress has been made in identifying the genes that regulate differentiation and the receptor-mediated signalling events that have a key role in this process. In particular, we are now learning how the CLAVATA receptor kinase signalling pathway promotes stem cell differentiation in balance with the initiation of stem cells by the transcription factor WUSCHEL. PMID- 11283726 TI - Multifunctional strands in tight junctions. AB - Tight junctions are one mode of cell-cell adhesion in epithelial and endothelial cellular sheets. They act as a primary barrier to the diffusion of solutes through the intercellular space, create a boundary between the apical and the basolateral plasma membrane domains, and recruit various cytoskeletal as well as signalling molecules at their cytoplasmic surface. New insights into the molecular architecture of tight junctions allow us to now discuss the structure and functions of this unique cell-cell adhesion apparatus in molecular terms. PMID- 11283727 TI - Cbl: many adaptations to regulate protein tyrosine kinases. AB - Responses to extracellular stimuli are often transduced from cell-surface receptors to protein tyrosine kinases which, when activated, initiate the formation of protein complexes that transmit signals throughout the cell. A prominent component of these complexes is the product of the proto-oncogene c Cbl, which specifically targets activated protein tyrosine kinases and regulates their signalling. How, then, does this multidomain protein shape the responses generated by these signalling complexes? PMID- 11283728 TI - Signal processing and transduction in plant cells: the end of the beginning? AB - Plants have a very different lifestyle to animals, and one might expect that unique molecules and processes would underpin plant-cell signal transduction. But, with a few notable exceptions, the list is remarkably familiar and could have been constructed from animal studies. Wherein, then, does lifestyle specificity emerge? PMID- 11283746 TI - The brainweb: phase synchronization and large-scale integration. AB - The emergence of a unified cognitive moment relies on the coordination of scattered mosaics of functionally specialized brain regions. Here we review the mechanisms of large-scale integration that counterbalance the distributed anatomical and functional organization of brain activity to enable the emergence of coherent behaviour and cognition. Although the mechanisms involved in large scale integration are still largely unknown, we argue that the most plausible candidate is the formation of dynamic links mediated by synchrony over multiple frequency bands. PMID- 11283747 TI - Constructing inhibitory synapses. AB - Control of nerve-cell excitability is crucial for normal brain function. Two main groups of inhibitory neurotransmitter receptors--GABA(A) and glycine receptors- fulfil a significant part of this role. To mediate fast synaptic inhibition effectively, these receptors need to be localized and affixed opposite nerve terminals that release the appropriate neurotransmitter at multiple sites on postsynaptic neurons. But for this to occur, neurons require intracellular anchoring molecules, as well as mechanisms that ensure the efficient turnover and transport of mature, functional inhibitory synaptic receptor proteins. This review describes the dynamic regulation of synaptic GABA(A) and glycine receptors and discusses recent advances in this rapidly evolving field. PMID- 11283748 TI - Development and plasticity of cortical areas and networks. AB - The development of cortical layers, areas and networks is mediated by a combination of factors that are present in the cortex and are influenced by thalamic input. Electrical activity of thalamocortical afferents has a progressive role in shaping cortex. For early thalamic innervation and patterning, the presence of activity might be sufficient; for features that develop later, such as intracortical networks that mediate emergent responses of cortex, the spatiotemporal pattern of activity often has an instructive role. Experiments that route projections from the retina to the auditory pathway alter the pattern of activity in auditory thalamocortical afferents at a very early stage and reveal the progressive influence of activity on cortical development. Thus, cortical features such as layers and thalamocortical innervation are unaffected, whereas features that develop later, such as intracortical connections, are affected significantly. Surprisingly, the behavioural role of 'rewired' cortex is also influenced profoundly, indicating the importance of patterned activity for this key aspect of cortical function. PMID- 11283749 TI - Plasticity of motor systems after incomplete spinal cord injury. AB - Although spontaneous regeneration of lesioned fibres is limited in the adult central nervous system, many people that suffer from incomplete spinal cord injuries show significant functional recovery. This recovery process can go on for several years after the injury and probably depends on the reorganization of circuits that have been spared by the lesion. Synaptic plasticity in pre-existing pathways and the formation of new circuits through collateral sprouting of lesioned and unlesioned fibres are important components of this recovery process. These reorganization processes might occur in cortical and subcortical motor centres, in the spinal cord below the lesion, and in the spared fibre tracts that connect these centres. Functional and anatomical evidence exists that spontaneous plasticity can be potentiated by activity, as well as by specific experimental manipulations. These studies prepare the way to a better understanding of rehabilitation treatments and to the development of new approaches to treat spinal cord injury. PMID- 11283750 TI - Oligomerization of G-protein-coupled transmitter receptors. AB - Examples of G-protein-coupled receptors that can be biochemically detected in homo- or heteromeric complexes are emerging at an accelerated rate. Biophysical approaches have confirmed the existence of several such complexes in living cells and there is strong evidence to support the idea that dimerization is important in different aspects of receptor biogenesis and function. While the existence of G-protein-coupled-receptor homodimers raises fundamental questions about the molecular mechanisms involved in transmitter recognition and signal transduction, the formation of heterodimers raises fascinating combinatorial possibilities that could underlie an unexpected level of pharmacological diversity, and contribute to cross-talk regulation between transmission systems. Because G-protein-coupled receptors are major pharmacological targets, the existence of dimers could have important implications for the development and screening of new drugs. Here, we review the evidence supporting the existence of G-protein-coupled-receptor dimerization and discuss its functional importance. PMID- 11283751 TI - A unified hypothesis on the lineage of neural stem cells. AB - For many years, it was assumed that neurons and glia in the central nervous system were produced from two distinct precursor pools that diverged early during embryonic development. This theory was partially based on the idea that neurogenesis and gliogenesis occurred during different periods of development, and that neurogenesis ceased perinatally. However, there is now abundant evidence that neural stem cells persist in the adult brain and support ongoing neurogenesis in restricted regions of the central nervous system. Surprisingly, these stem cells have the characteristics of fully differentiated glia. Neuroepithelial stem cells in the embryonic neural tube do not show glial characteristics, raising questions about the putative lineage from embryonic to adult stem cells. In the developing brain, radial glia have long been known to produce cortical astrocytes, but recent data indicate that radial glia might also divide asymmetrically to produce cortical neurons. Here we review these new developments and propose that the stem cells in the central nervous system are contained within the neuroepithelial --> radial glia --> astrocyte lineage. PMID- 11283752 TI - Acetylcholinesterase--new roles for an old actor. AB - The discovery of the first neurotransmitter--acetylcholine--was soon followed by the discovery of its hydrolysing enzyme, acetylcholinesterase. The role of acetylcholinesterase in terminating acetylcholine-mediated neurotransmission made it the focus of intense research for much of the past century. But the complexity of acetylcholinesterase gene regulation and recent evidence for some of the long suspected 'non-classical' actions of this enzyme have more recently driven a profound revolution in acetylcholinesterase research. Although our understanding of the additional roles of acetylcholinesterase is incomplete, the time is ripe to summarize the evidence on a remarkable diversity of acetylcholinesterase functions. PMID- 11283755 TI - Conflicts of interest at the industrial/academic interface. PMID- 11283756 TI - Cost effectiveness of HMG-CoA reductase inhibition in Canada. AB - OBJECTIVE: To assess the cost effectiveness of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor therapy, particularly atorvastatin, in primary and secondary prevention of coronary artery disease (CAD) in Canada. METHODS: A Markov model was developed in which costs and effectiveness of atorvastatin were compared with those of other statins and with no drug therapy in primary and secondary prevention of CAD. PATIENTS: Cost effectiveness was assessed for cohorts of patients with risk profiles defined by CAD status, age, sex, pretreatment low density lipoprotein cholesterol level and presence of sentinel coronary risk factors. Coronary risk was estimated by using initial and subsequent event coronary risk equations from the Framingham Heart Study, and risk factors were estimated by using Canadian population survey data. Recent estimates of the costs of CAD-related medical care in Canada were used to assign costs to health states and acute coronary events. INTERVENTIONS: Interventions included atorvastatin 10 mg, simvastatin 10 mg, pravastatin 20 mg, fluvastatin 20 mg, lovastatin 20 mg and no pharmacological therapy. RESULTS: Incremental cost effectiveness ratios (CDN$/year of life gained) relative to no therapy were lowest for atorvastatin and highest for pravastatin across all risk profiles. Atorvastatin was less costly and more effective than lovastatin, pravastatin and simvastatin in primary and secondary prevention, and conferred additional health benefits at a reduced cost per year of life gained compared with fluvastatin. CONCLUSIONS: Atorvastatin was found to be the most cost effective statin in primary and secondary prevention of CAD. PMID- 11283757 TI - [Indigenous peoples' health in Brazil: current perspectives]. PMID- 11283758 TI - [Organization and quality of health care for Kaingang Indians in Rio Grande do Sul, Brazil]. AB - This study assesses the health care provided to Kaingang Indians in Rio Grande do Sul, Brazil. Deaths preventable by primary health care among the Indians and occurring from 1985 to 1995 were compared to the same rates for the State of Rio Grande do Sul as a whole. Secondary data on health care services were supplemented by field interviews with indigenous leaders and with employees from participating institutions. A Geographic Information System (GIS) was used to correlate distribution of deaths and access to health services. The Kaingang settlements are connected by paved roads to counties with at least a public health clinic or even a small hospital in some cases. Secondary referrals are treated in Palmeiras das Missoes and Frederico Westphallen and tertiary care is provided in Passo Fundo. What distinguishes the Indian settlements from the rest of the State are the high rates of deaths preventable by primary health care and those related to ill-defined conditions, malnutrition, tuberculosis, and cancer of the uterine cervix. PMID- 11283759 TI - [Sickness, Healing, and Health Services: social representations, practices, and demands among the Baniwa]. AB - The research for this paper was conducted in Sao Gabriel da Cachoeira, in the northwestern Amazon, with the Baniwa indigenous people, in partnership with indigenous organizations, seeking to understand the relations among the group's cosmology, their system of representations of sickness and healing practices, and their transformation through inter-ethnic contact. The recording of myths showed the origin of the diseases and demonstrated the existence of several traditional categories of sickness, guiding traditional healing practices and the incorporation of biomedical knowledge. The Baniwa's cosmology operates like a reception system for biomedical information, which the people grasp according to the logic of mythical thought. Similar cognitive strategies are used to generate the demands that indigenous leaders submit to the Health Councils and Health Services. PMID- 11283760 TI - [Tuberculosis among indigenous populations in Rondonia, Amazonia, Brazil]. AB - Tuberculosis persists as a serious public health problem in Brazil. Prevalence rates are alarming in certain social groups, including indigenous peoples. This article presents an epidemiological analysis of records in the Rondonia State Tuberculosis Control Program, identifying the disease's profile among indigenous groups, which are socially more vulnerable and have different issues involved in controlling the disease. The study includes a descriptive statistical and multivariate multinomial analysis of cases reported in 1992 and from 1994 to 1998, attempting to identify factors associated with tuberculosis-related deaths, treatment drop-out, and missing data. Associations were identified between variables related to the disease, to the health service, and to treatment results. There is evidence that the indigenous populations in Rondonia have an increased risk of acquiring and dying from tuberculosis as compared to other residents of the State. Attention is called to the need for prevention and control measures specifically tailored to the reality of indigenous peoples. PMID- 11283761 TI - [Suicide among the Tikuna on the Upper Solimoes River: an expression of conflicts]. AB - This study focuses on suicide among the Tikuna Indians on the Upper Solimoes River in the Brazilian Amazonia. The very nature of the object requires an interdisciplinary approach. The ethnography is concerned with capturing links among suicides occurring over the last decade, in the process of confrontation between different factions or groups that express, in the present, an older historical context, the mechanisms of conflict characterizing the ancient malocas, or indigenous lodges. At the very base of such confrontations lies the abandonment inflicted on this population, especially the bankruptcy of the health care model proposed for the Upper Solimoes area. PMID- 11283762 TI - [Human ecology and nutritional anthropometry of adult Xavante Indians in Mato Grosso, Brazil]. AB - This study compares anthropometric and ecological profiles of two Xavante indigenous communities in Mato Grosso, Central Brazil. The research describes time allocation patterns and involves an anthropometric survey (including body mass, stature, and BMI) in adults over 20 years of age. Data from Etenitepa (also known as Pimentel Barbosa) were collected in 1994. Field work at Sao Jose was conducted in 1998 and 1999. Compared with the Sao Jose group, Xavante in Etenitepa do more subsistence activities like farming, fishing, hunting, and gathering. The Sao Jose Xavante do more paid work and generally engage in less physical activity. Average stature in the two communities is similar, but there are major differences in mean body mass and BMI. The Sao Jose group has average BMI values well over those of the Etenitepa group in practically all age brackets. Obesity prevalence rates were high in both men (24.6%) and women (41.3%) in Sao Jose, while in Etenitepa the rates were only 2.5% and 4.8%, respectively. The authors conclude that the different nutritional profiles in the two communities result from specific patterns of social, political, and economic interactions with Brazilian society. PMID- 11283764 TI - [The Yawanawa medical system and its specialists: healing, power, and shamanic initiation]. AB - A wide variety of terms are used in Yawanawa shamanism to name both healers and the relations between these specialists and techniques of healing and aggression, a characteristic that appears heterogeneous and complex. However, there is unity behind this appearance, because all the techniques are based on the same concepts of power and knowledge. To show how these ideas are conceived by the Yawanawa, this article explores the process by which power and knowledge are acquired: shamanic initiation. PMID- 11283763 TI - [Child health and nutrition in a Terena indigenous community, Mato Grosso do Sul, Brazil]. AB - This paper describes the health and nutritional situation of South American Indian children from a Terena community, characterizing their nutritional status, food consumption, and socioeconomic and environmental conditions. The sample included 100 children, ranging from 0 to 59 months of age and living in Aldeia Corrego do Meio, Mato Grosso do Sul. Prevailing nutritional deficits were: 8.0% for the weight-for-age index, 16.0% for height-for-age, and 5.0% for weight-for height. The growth deficit rate was higher than that of the Brazilian population as a whole, probably reflecting the precarious socioeconomic, environmental, and health conditions in this Terena community. Analysis of the average nutrient sufficiency in the infant diet showed that nutritional recommendations for the different groups were not complied with. New studies, characterized as transdisciplinary and longitudinal, are necessary to better understand this process. PMID- 11283765 TI - [Prevalence of risk factors for cardiovascular disease in the Guarani-Mbya population of the State of Rio de Janeiro]. AB - Social change has been involved in the unequal distribution patterns of chronic diseases in several populations. Among Indian communities experiencing life pattern changes, international studies have reported increased prevalence of hypertension and other cardiovascular risk factors. Such increased prevalence was ascertained in a survey conducted in 1999 in selected Guarani-Mbya communities (Sapukai, Paraty-Mirim, and Araponga) in the State of Rio de Janeiro, Brazil. A population census was carried out and interviews and clinical and biochemical evaluations were conducted with 80 men and 71 women. Observed prevalence of selected risk factors in the overall sample, including men and women, was as follows for the three communities: hypertension (4.8%, 2.6%, 7.4%); overweight (26.7%, 19.5%, 34.8%); total cholesterol levels (2.8%, 2.7%, 2.9%), and increased triglyceride levels (12.6%, 9.5%, 15.9%). All prevalence rates were higher among women and at older ages. The results suggest that the Guarani communities have a moderate risk of chronic diseases and that measures to reduce these risk factors should be adopted. PMID- 11283766 TI - [Demographic profile of the Xavante Indian population in Sangradouro-Volta Grande, Mato Grosso]. AB - This paper analyzes the demographic profile of the Xavante population at the Sangradouro-Volta Grande Indigenous Reserve in Mato Grosso, Brazil, from 1993 to 1997. The survey included annual censuses and vital statistics from 7 Xavante villages. Permanent contact with Brazilian national society, established in the 1940s and 50s, caused a population drop due to epidemics and clashes. In 1995 there were 825 individuals in the community. The crude birth rate (57.7/1,000) and death rate (9.1/1,000) were higher than the national averages. The majority (56%) of the population is under 15 years of age (median: 13 years) and the infant mortality rate is high (87.1 per thousand live births), probably resulting from precarious sanitary conditions in the villages. Other results included the persistence of polygyny; low levels of migration; a dynamic of splits and formation of new villages; traditional housing patterns maintained in the old villages and abandoned in the new ones. The recent demographic recovery in the data from Sangradouro-Volta Grande is similar to that observed in the Pimentel Barbosa community. The study highlights the importance of systematically collecting and analyzing demographic data from indigenous populations. PMID- 11283767 TI - [Risk factors for multiple intestinal parasites in an indigenous community of the State of Pernambuco, Brazil]. AB - An investigation into the ethno-epidemiological profile of the Pankararu indigenous group in the State of Pernambuco, Brazil, identified multiple intestinal parasites in nearly all members of the community. To detect possible environmental risk factors, we used the data base from a previous survey to test relations between daily living conditions (housing, sanitation, water supply and treatment, and garbage disposal) and the number of different parasite species found in the same household. The sample consisted of 84 families from the original sample of 112. Selection was based on the number of stool tests performed in the family. The mean number of parasite species was 5.0 per family, for a mean family size of 6.1 members. This number was greater for wattle-and daub houses (mean 6.0 parasite species vs. 4.9 for brick houses; p < 0.03) and when water used in the household was not treated (mean 5.1 parasite species, vs. 4.5 for treated water; p < 0.05). Other household characteristics and hygienic habits did not significantly influence this number. We concluded that multiple intestinal parasitism in the Pernambuco Pankararu community is frequent, to the point of being the rule, and that it relates essentially to water source and treatment. PMID- 11283768 TI - [Oral health among the Xavante Indians in Pimentel Barbosa, Mato Grosso, Brazil]. AB - This study presents the results of an oral health epidemiological survey conducted in 1997, based on WHO criteria, in the Xavante indigenous community of Pimentel Barbosa (or Etenitepa), Mato Grosso State, Central Brazil. The study included 228 individuals (85% of the population) over two years of age. In about half the sample, the DMF index was less than 2, and in the 12-14-year age bracket it was 3.7. The low frequency of fillings in permanent and deciduous teeth suggests limited access to dental care services. Despite the number of sextants with bleeding and tartar, no cases of severe periodontal disease were detected (CPITN). In the community, comparison of the results of this survey with two previous surveys (1962 and 1991) showed a deterioration in oral health conditions over time and alterations in the occlusal pattern (increase in Angle class II and III). Dietary changes due to environmental and socioeconomic alterations resulting from interaction with the surrounding society, along with the lack of preventive programs, are among the causes of this deterioration in oral health among the Xavante. PMID- 11283769 TI - [Chlamydia infection impact among native Indian groups of the Brazilian Amazon region]. AB - Knowledge is limited on the spread of bacteria from genus Chlamydia in Brazil. This study included a sero-epidemiological survey of 2,086 samples from native Indian populations of the Brazilian Amazon region. Sera were screened using indirect immunofluorescence assay for detection of antibodies to C. trachomatis serotype L2, followed by microimmunofluorescence assay using fifteen C. trachomatis and C. pneumoniae serotypes as antigen substrates. Antibody prevalence was 48.6%, but there was a large prevalence range among the groups, including those that had never been challenged with the bacteria, as well as those in which almost all individuals had been infected. Titration of IgG antibodies and detection of specific IgM in high-titer samples showed the persistence of Chlamydia in 6.1% of the reactive individuals, who probably play an important role as reservoirs for dissemination of the bacteria. Specific seroreactivity to C. trachomatis showed the presence of serotypes A, B, Ba, D, E, G, H, I, and L1 in the geographic area surveyed. Furthermore, the survey showed that C. pneumoniae was also infecting these individuals. Both species may be involved in a significant human disease burden that merits further clarification. PMID- 11283770 TI - [Contact, epidemics, and the body as agents of change: a study of AIDS among the Xokleng indians in the State of Santa Catarina, Brazil]. AB - Based on an analysis of AIDS cases among the Xokleng Indians in 1988, this article relates the illness phenomenon to socio-cultural disruptions and transformations in this indigenous group's universe, focusing on the history of their contact with Brazilian national society. The analysis and interpretation of this relationship are based on anthropological theories about the centrality of the body, corporeality, and degenerative bodily processes in Brazilian indigenous societies, according to which the body, society, and macro-situational elements are articulated by social praxis, and should thus be related in socio anthropological studies of health-illness phenomena. The article briefly describes the history of epidemics emerging from contact and attempts to relate them to specific historical contexts. Ethnomedical categories, cosmology, and Xokleng concepts of corporeality are related to their social organization, which are thus connected to the AIDS cases. The latter are presented with a special focus on the relationship between their emergence and the changes occurring in the Xokleng world with the construction of a dam bordering on their land. PMID- 11283771 TI - [Health and disease among Panara (Kreen-Akarore) Indians in Central Brazil after twenty-five years of contact with our world, with an emphasis on tuberculosis]. AB - The Panara, who had previously lived in isolation from Brazilian national society in the Amazon forest, were first contacted in 1973. Two years later they were moved to another area in Central Brazil. During this same period they were reduced to 82 members, the survivors of a population of 400 to 500 in the mid 1960s. In 1995 they returned to a small area in their old territory still not occupied by outsiders. There, three years later, a health survey showed a presumed diagnosis of tuberculosis in 15 individuals out of a population of 181. Further tests in the town of Colider, based on clinical data and chest X-rays, confirmed the diagnosis in 10 Panara (6 children under 10 years of age and 4 adults from 40 to 50 years old). BCG scars were present in the entire population. The nutritional status of Panara children was better than that of other indigenous groups in the Amazon region. The following measures were introduced for Tb control: a) treatment follow-up in the village, under direct supervision by both a nurse and the local indigenous health worker; b) compliance with defined criteria for ending treatment; c) periodic control of contacts and non contacts; c) and establishment of a reference system with the health services in Colider. PMID- 11283772 TI - [Electric power generation and transmission: the impact on indigenous peoples in Brazil]. AB - This paper presents an overview of the effects of electric power generation and transmission on indigenous communities in Brazil. According to data from FUNAI (the Brazilian government's Board of Indian Affairs), there are 156 cases of direct impact, present or future, of the electric power sector on Indian settlements geographically distributed throughout Brazil, 65% of which are located in the Northern Region of the country. The principal complaints by indigenous communities relate to the direct effects of flooding following construction of hydroelectric dams, destruction of sacred sites like cemeteries, mosquito proliferation, and health-related hazards such as malaria and other infectious diseases, decrease in game for hunting, crowding out of farm land, and increased invasion of indigenous lands. Future perspectives include a scenario with further construction of hydroelectric dams, especially in the Amazon region, with possible similar effects on indigenous communities. PMID- 11283773 TI - [Indigenous peoples' health and the implementation of Health Districts in Brazil: critical issues and proposals for a transdisciplinary dialogue]. AB - Based on the authors' experience, this paper discusses a series of problems during the implementation of so-called Indigenous Health Districts (DSEI) in Brazil, related to organization of health services provision as viewed by both health professionals and anthropologists. The authors report on the Health District model's underlying concepts and present the different approaches used to implement the DSEI. The authors' experience refers to the Rio Negro area - in the northwestern Amazon, representing 10% of Brazil's total indigenous population - and the Brazilian Northeast, specifically the State of Pernambuco, with an indigenous population estimated at 20,000. PMID- 11283774 TI - [Evaluation of the nutritional status of the Parkateje indigenous community in Bom Jesus do Tocantins, Para, Brazil]. AB - The nutritional status of the Mae-Maria indigenous community in Bom Jesus do Tocantins, Para State, Brazil, was ascertained in a descriptive study in which ninety percent of the total population (278 individuals) agreed to participate. Weight-for-height and height-for-age indices and Body Mass Index (BMI) were ascertained for children and adults by gender, respectively, as were weight and height means in adolescents. Compared to NCHS curves, overweight and chronic malnutrition were observed, respectively, in 6.7 and 8.6 % of all children under 10 years old (104). Weight means were similar among Indians and NCHS adolescents, while height means were lower among the former. Overweight prevalence (BMI 25 to 29) was 23.7% among male adults and 50.0% among female adults, and obesity (BMI > 30) was observed in 12.5% of adult females. PMID- 11283775 TI - [Health problems among the Kaingang (Xapeco Indigenous Reserve, Santa Catarina) and the health care system]. AB - The second semester of 1999 was a transition period for the implementation of the Special Indigenous Health District on the Xapeco Indigenous Reserve in western Santa Catarina State. The health clinic in the main village provided treatment with a staff including a general practitioner/obstetrician, pediatrician, dentist, nurse, two nursing assistants, and four nursing technicians. This paper presents the preliminary results of research on the organization of these health care services, their use by the community, and the health/disease profile of the Kaingang, using patient files as the source of information. In September 1999, a total of 222 Indians were treated (children and adults), 50.5% of whom residing in the main village. Among the Indians ages 0 to 14 years, infectious and parasitic diseases were the most frequent, supporting the idea that the Kaingang have precarious sanitary and nutritional conditions. Use of the clinic by adults was more varied, since of the 116 who appeared for consultation, 27 were pregnant women (out of a total of 86 women). In addition, prescriptions were written up for children and adults in 85.0% and 81.8% of the consultations, respectively. PMID- 11283776 TI - [Surgical treatment of the partial or complete obstruction of the lacrimonasal duct with a polyurethane stent]. PMID- 11283777 TI - [New systems of drug administration through a topical ocular administration approach]. PMID- 11283778 TI - [Palpebral contraction (lid twitch) in ptosis caused by myasthenia gravis]. PMID- 11283779 TI - [Experimental study about the effects of exploratory endoscopical technique on the posterior orbit]. AB - PURPOSE: To determine the effects of the orbital endoscopic technique in the dog (IOP, vascularization). METHODS: Eight dogs were used to examine the four quadrants of the posterior orbit that we have established as a theoretical division of the orbit. Thus, two examinations were performed for each region. In order to observe the effects of the current technique, a rigorous control on the visual parameters was established, paying special attention to the effects of the blood supply in the vessels of the eyeball, measuring the posterior ciliary arteries (in the dog, the central artery of the retina is very rudimentary) with the Doppler color and Power Doppler echography. RESULTS: The results compiled on the examination of the visual function, ocular mobility, the general examination of reflexes, examination of the ocular fundus and general condition did not show any significant alteration. Conjunctival chemosis occurred in one case, but spontaneously disappeared 76 hours after the operation. The IOP only showed a mild increase after the surgery in two cases. The PSV and EDV blood flow values increased in the immediate post-surgery, above all in the vessel belonging to the region operated in 6 out the 8 subjects, but had decreased at 7 days post surgery. CONCLUSIONS: The results obtained did not show any significant alterations when the orbital endoscopic technique for the posterior orbit was used. A mild increase is observed in the blood velocity values of the posterior ciliary arteries after the surgery that seems to indicate a certain compression. However, these values are reduced significantly when the method is repeated within 7 days. PMID- 11283780 TI - [Effect of tobramycin with topical diclofenac on arachidonic acid in endotoxin induced uveitis]. AB - PURPOSE: We studied if tobramycin associated to sodium diclofenac modifies the drug effect of the latter on arachidonic acid metabolism in an endotoxin induced uveitis (EIU) model in albino rabbits. METHODS: Experimental uveitis was induced by intravitreal injection of a 10 ng lipopolysaccharide A Salmonella typhimurium endotoxin dissolved in 5 microl saline solution in the right eye of experimental animals. We have used 48 animals (4 groups of 12 animals each). The control group (G-I) was injected with saline solution (5 microl); the endotoxin group (G-II) was injected with 5 microl of ET; group III and IV were injected with the same amount of ET and treated with topical sodium diclofenac (1%) (Group III) and tobramycin and sodium diclofenac (0.3%) (Group IV) two hours before the administration of endotoxin and then every two hours until 24 hours, when we determined E2 prostaglandin and B4 leukotriene concentration in aqueous humor obtained by paracentesis of the anterior chamber. RESULTS: We observed how the E2 prostaglandin concentration was reduced in the two treatment groups compared with the endotoxin group (p<0.01). However, there were no differences between groups III and IV (p>0.01). There was a mild increase of B4 leukotriene in both treatment groups, which was not significant in relationship to the endotoxin group (p>0.01). No differences were found between the two treatment groups (p>0.01). CONCLUSION: Using the association of tobramycin does not affect the action of sodium diclofenac in arachidonic acid metabolism in endotoxin induced uveitis. PMID- 11283781 TI - [Evaluation of the work time used in outpatient ophthalmology techniques]. AB - PURPOSE: To determine the work time used for out-patients seen in the ophthalmology out-patient clinic according to treatment for each diagnosis and examination guidelines. Another objective was to discover the diagnostic related outpatient demand as well as the time spent on each technique. METHOD: We have designed a computer database system that includes examinations, treatments and medical services. Their duration was measured in a random sample of 127 ophthalmology outpatients in the General Hospital of Castellon. Sixty five of the patients had come to their first visit and 62 to the follow-up visits. RESULTS: A new patient spends a mean of 42 minutes and a follow-up patient spends a mean of 27 minutes. The most frequent diagnoses are retinal diseases (30%), cataract (18.6%), glaucoma (11.8%) and ocular motility disorders (11.8%). We present the mean times used by the physician in the ophthalmic techniques used in outpatients. CONCLUSIONS: We have modified the standard times of 30 minutes for the first visit and 15 minutes for the follow-up visit. Data on the work time used for all the ophthalmics techniques in our outpatient clinic are presented. This ophthalmology survey can be used for health care managers to establish the <> and health care plan administration to adapt the resources to the demand. PMID- 11283782 TI - [Ciliary ablation with diode laser. Long-term study]. AB - PURPOSE: The effects of contact transscleral cyclophotocoagulation with diode laser for refractory glaucoma have been analyzed. METHODS: A diode laser system was used to treat twenty one eyes with refractory glaucoma. The mean follow-up was 22.24+/-9.89 months. All the eyes were treated with 15 laser spots placed over 270 degrees, 1 mm posterior to the limbus, using 3.0 J (1.0 W X 3.0 sg). RESULTS: The mean preoperative intraocular pressure was 40.95+/-11.76 mmHg and the mean intraocular pressure at the end of the follow-up was 24.28+/-7.80 mmHg (p<0.5). An intraocular pressure below 21 mmHg was obtained in 10 out the 13 eyes (76.92%). Six of the 8 painful eyes had pain relief (75.00%). The principal complications included a decreased visual acuity in 5 eyes and chronic uveitis in 2 patients. CONCLUSIONS: These results suggest that semiconductor diode transscleral cyclophotocoagulation can be used successfully to reduce intraocular pressure in the treatment of refractory glaucoma. PMID- 11283783 TI - [Experimental study of the suture of the intraocular lens sutured to the ciliary sulcus. Needle passage and distance from the limbus]. AB - OBJECTIVE: To discover which technique is best to scleral suture fixation of the lens in the ciliary sulcus, based on two needle direction options: passing the needle from the outer part of the eyeball inwards or from the inner part outwards and to know at what distance the needle must be passed from the sclerocorneal limbus. MATERIAL AND METHODS: We used 40 cadaver eyes conserved in 10% formaldehyde. The needle was passed from the interior to the exterior of the eye, observing whether if it had crossed through the ciliary sulcus and the distance of the needle from the limbus. Stitches were also made, going from the exterior to the interior of the eyeball at a known distance from the limbus, either parallel to the iris or perpendicular to the sclera. RESULTS: When the needle was passed from the interior toward the exterior of the eye, it passed through the ciliary sulcus, exiting the eye at 1.50+/-0.16 mm from the limbus, in 80% of the cases. When the needle was passed from the exterior toward the interior of the globe perpendicularly to the sclera only 32.5% passed through the sulcus and when it was parallel to the iris only, only 40%. There is greater statistical probability that the needle will pass through the sulcus from the interior of the eyeball, exiting the limbus at approximately 1.5 mm. CONCLUSION: Since it is necessary to pass the suture through the sulcus in order to be able to place the lens haptics in the ciliary sulcus when suturing a posterior chamber lens to the sclera, it is best to pass the needle from the interior to the exterior of the eyeball and for the needle to exit at approximately 1.5 mm from the limbus. PMID- 11283784 TI - [Doppler ultrasound in type I diabetes: preliminary results]. AB - PURPOSE: To study the differences of blood flow in type I diabetes patients using a color Doppler ultrasound. MATERIAL AND METHODS: We measured the systolic peak velocity (Vmax), diastolic velocity (Vmin) as well as the resistance index in the central retinal artery (CRA) and in the ophthalmic artery (OA) using a color Doppler ultrasound in 40 diabetic patients. We classified the patients according to the duration of diabetes, which ranged from 1 to 28 years. and whether retinopathy was present. We compared the results against those obtained in the 40 control subjects without vascular pathology. RESULTS: When the results of the two groups were compared, we found that no decrease was observed in the CRA flow rate in diabetics without retinopathy, however there was a significant decrease in flow velocity in patients with initial (p<0.05), moderate (p<0.001) or proliferative retinopathy (p<0.05). The ophthalmic artery presents an increase in the vascular resistance in diabetics with proliferative retinopathy. As the time of diabetes increases, there is a decrease in the blood flow in CRA and an increase in resistance in the OA. CONCLUSIONS: The Doppler ultrasound is a non invasive technique which allows us to perform a hemodynamic study of the orbital vessels. It is essential to understand the correlation between blood flow velocity and the severity of retinopathy in diabetes patients as it manifests the relationship between the velocity of the blood flow with the severity of the retinopathy and that there is a direct relationship with the evolution of the diabetes and the flood flow velocity. PMID- 11283785 TI - [Retinal thrombosis in young patients. Immunological and clinical aspects]. AB - OBJECTIVES: To ascertain preexisting medical conditions, clinical evolution of retinopathy, and associated immunological disorders in a series of young patients suffering from retinal thrombosis, and to determine the prevalence of antiphospholipid antibodies. METHODS: Twenty two patients younger than 50 years, who had presented an acute retinal thrombotic episode, were studied prospectively with a general physical, ophthalmoscopic and immunological examination, placing special emphasis on the detection of antiphospholipid antibodies (anticardiolipin antibodies and lupus anticoagulant). RESULTS: No baseline disease stood out significantly over the others, and the most frequent risk factor found was systemic arterial hypertension (5/22%). No associated risk factor was found in nine cases (41%), and more than two factors were found in six cases (27%). Most of the vascular occlusions affected the venous vessels (18/81%), and five of them were associated with vasculitis. The ophthalmologic follow-up showed a rapid evolution to retinal neovascularization in 11 cases. Our data show many immunologically altered values, there being nine cases (41%) of the series with more than four parameters altered. The antiphospholipid assay showed a high prevalence of anticardiolipin antibodies (5/23%), and two patients were diagnosed of primary antiphospholipid syndrome. The lupus anticoagulant was negative in all patients. CONCLUSIONS: The high prevalence of anticardiolipin antibodies and immunologic abnormalities found in the retinal thrombosis younger patients leads us to recommend the systematic immunological study in these subjects. It has relevant diagnostic and therapeutic implications in a population with no evident associated risk factors and a greater severity of retinopathy. PMID- 11283786 TI - [Anterior optic neuritis do to ulcerative colitis]. AB - PURPOSE/METHODS: We present the case of a 27 year old woman with acute bilateral loss of vision associated with abdominalgia, bloody diarrhea and arthralgia. In the present article, the different forms of ocular complications in relationship to chronic intestinal inflammations and their possible causes are discussed. RESULTS/CONCLUSIONS: On examination, bilateral anterior uveitis associated to a picture of optic nerve lesion compatible with a bilateral anterior optic neuritis was observed. The patient was diagnosed of ulcerative colitis by intestinal biopsy. The ocular affectation resolved after the ulcerative colitis was treated with systemic mesalamine. PMID- 11283787 TI - [Repair of Descemet's membrane detachment after cataract surgery]. AB - PURPOSE/METHODS: We present two cases of Descemet's membrane detachment, the first one after extracapsular cataract extraction with scleral tunnel incision and the other after clear corneal incision used for phacoemulsification cataract extraction. These two cases were treated with intracameral injection of 20% sulfur hexafluoride gas (SF(6)). RESULTS/CONCLUSIONS: Both cases responded satisfactorily to treatment. After four months of follow-up, the corneas remain clear with a visual acuity of 0.9 and 0.8, respectively. The relative facility of the 20% sulfur hexafluoride gas (SF(6)) technique, its safety, and good prognosis, makes it the treatment of choice in this disease. PMID- 11283788 TI - [Ocular treatments in folk medicine in the city of Badajoz]. PMID- 11283789 TI - [The Coat of Arms of Jean Paul Marat]. PMID- 11283791 TI - Linkage and association analysis of angiotensin I-converting enzyme (ACE)-gene polymorphisms with ACE concentration and blood pressure. AB - Considerable effort has been expended to determine whether the gene for angiotensin I-converting enzyme (ACE) confers susceptibility to cardiovascular disease. In this study, we genotyped 13 polymorphisms in the ACE gene in 1,343 Nigerians from 332 families. To localize the genetic effect, we first performed linkage and association analysis of all the markers with ACE concentration. In multipoint variance-component analysis, this region was strongly linked to ACE concentration (maximum LOD score 7.5). Likewise, most of the polymorphisms in the ACE gene were significantly associated with ACE (P<.0013). The two most highly associated polymorphisms, ACE4 and ACE8, accounted for 6% and 19% of the variance in ACE, respectively. A two-locus additive model with an additive x additive interaction of these polymorphisms explained most of the ACE variation associated with this region. We next analyzed the relationship between these two polymorphisms (ACE4 and ACE8) and blood pressure (BP). Although no evidence of linkage was detected, significant association was found for both systolic and diastolic BP when a two-locus additive model developed for ACE concentration was used. Further analyses demonstrated that an epistasis model provided the best fit to the BP variation. In conclusion, we found that the two polymorphisms explaining the greatest variation in ACE concentration are significantly associated with BP, through interaction, in this African population sample. Our study also demonstrates that greater statistical power can be anticipated with association analysis versus linkage, when markers in strong linkage disequilibrium with a trait locus have been identified. Furthermore, allelic interaction may play an important role in the dissection of complex traits such as BP. PMID- 11283790 TI - A major locus for fasting insulin concentrations and insulin resistance on chromosome 6q with strong pleiotropic effects on obesity-related phenotypes in nondiabetic Mexican Americans. AB - Insulin resistance and hyperinsulinemia are strong correlates of obesity and type 2 diabetes, but little is known about their genetic determinants. Using data on nondiabetics from Mexican American families and a multipoint linkage approach, we scanned the genome and identified a major locus near marker D6S403 for fasting "true" insulin levels (LOD score 4.1, empirical P<.0001), which do not crossreact with insulin precursors. Insulin resistance, as assessed by the homeostasis model using fasting glucose and specific insulin (FSI) values, was also strongly linked (LOD score 3.5, empirical P<.0001) with this region. Two other regions across the genome were found to be suggestively linked to FSI: a location on chromosome 2q, near marker D2S141, and another location on chromosome 6q, near marker D6S264. Since several insulin-resistance syndrome (IRS)-related phenotypes were mapped independently to the regions on chromosome 6q, we conducted bivariate multipoint linkage analyses to map the correlated IRS phenotypes. These analyses implicated the same chromosomal region near marker D6S403 (6q22-q23) as harboring a major gene with strong pleiotropic effects on obesity and on lipid measures, including leptin concentrations (e.g., LOD(eq) for traits-specific insulin and leptin was 4.7). A positional candidate gene for insulin resistance in this chromosomal region is the plasma cell-membrane glycoprotein PC-1 (6q22-q23). The genetic location on chromosome 6q, near marker D6S264 (6q25.2-q26), was also identified by the bivariate analysis as exerting significant pleiotropic influences on IRS related phenotypes (e.g., LOD(eq) for traits-specific insulin and leptin was 4.1). This chromosomal region harbors positional candidate genes, such as the insulin-like growth factor 2 receptor (IGF2R, 6q26) and acetyl-CoA acetyltransferase 2 (ACAT2, 6q25.3-q26). In sum, we found substantial evidence for susceptibility loci on chromosome 6q that influence insulin concentrations and other IRS-related phenotypes in Mexican Americans. PMID- 11283792 TI - The primary erythermalgia-susceptibility gene is located on chromosome 2q31-32. AB - Primary erythermalgia is a rare disorder characterized by recurrent attacks of red, warm, and painful hands and/or feet. The symptoms are generally refractory to treatment and persist throughout life. Five kindreds with multiple cases of primary erythermalgia were identified, and the largest was subjected to a genomewide search. We detected strong evidence for linkage of the primary erythermalgia locus to markers from chromosome 2q. The highest LOD score (Z) was obtained with D2S2330 (Z(max) = 6.51). Analysis of recombination events identified D2S2370 and D2S1776 as flanking markers, on chromosome 2q31-32. This defines a critical interval of 7.94 cM that harbors the primary erythermalgia gene. Affected members within the additional families also shared a common haplotype on chromosome 2q31-32, supporting our linkage results. Identification of the primary erythermalgia gene will allow a better clinical classification of this pleomorphic group of disorders. PMID- 11283794 TI - Null RPGRIP1 alleles in patients with Leber congenital amaurosis. AB - We isolated and characterized the entire coding sequence of a human gene encoding a protein that interacts with RPGR, a protein that is absent or mutant in many cases of X-linked retinitis pigmentosa. The newly identified gene, called "RPGRIP1" for RPGR-interacting protein (MIM 605446), is located within 14q11, and it encodes a protein predicted to contain 1,259 amino acids. Previously published work showed that both proteins, RPGR and RPGRIP1, are present in the ciliary structure that connects the inner and outer segments of rod and cone photoreceptors. We surveyed 57 unrelated patients who had Leber congenital amaurosis for mutations in RPGRIP1 and found recessive mutations involving both RPGRIP1 alleles in 3 (6%) patients. The mutations all create premature termination codons and are likely to be null alleles. Patients with RPGRIP1 mutations have a degeneration of both rod and cone photoreceptors, and, early in life, they experience a severe loss of central acuity, which leads to nystagmus. PMID- 11283795 TI - Heteroplasmy of the human mtDNA control region remains constant during life. AB - In a longitudinal, retrospective study, we monitored the level of heteroplasmy at nucleotide position (nt) 309 and nt 16189 of the control region of human mtDNA. As a unique source of DNA, we analyzed multiple cervical-cell samples collected, during 1 or 2 decades, from four women with heteroplasmy at either nt 309 or nt 16189. According to accurate, quantitative analysis by solid-phase minisequencing, the level of heteroplasmy remained stable in the cervical-cell samples from all four women during the time studied. We also analyzed autopsy samples from several different tissues, all containing nt 309 in heteroplasmic form, of one of the women, who was deceased. On the basis of our results, heteroplasmy in the control region of mtDNA seems to be inherited and is not the result of somatic age-related accumulation. PMID- 11283793 TI - Transaldolase deficiency: liver cirrhosis associated with a new inborn error in the pentose phosphate pathway. AB - This article describes the first patient with a deficiency of transaldolase (TALDO1 [E.C.2.2.1.2]). Clinically, the patient presented with liver cirrhosis and hepatosplenomegaly during early infancy. In urine and plasma, elevated concentrations of ribitol, D-arabitol, and erythritol were found. By incubating the patient's lymphoblasts and erythrocytes with ribose-5-phosphate and subsequently analyzing phosphate sugar metabolites, we discovered a deficiency of transaldolase. Sequence analysis of the transaldolase gene from this patient showed a homozygous deletion of 3 bp. This deletion results in absence of serine at position 171 of the transaldolase protein. This amino acid is invariable between species and is located in a conserved region, indicating its importance for enzyme activity. The detection of this new inborn error of pentose metabolism has implications for the diagnostic workup of liver problems of unknown etiology. PMID- 11283796 TI - Disruption of the bipartite imprinting center in a family with Angelman syndrome. AB - Imprinting in 15q11-q13 is controlled by a bipartite imprinting center (IC), which maps to the SNURF-SNRPN locus. Deletions of the exon 1 region impair the establishment or maintenance of the paternal imprint and can cause Prader-Willi syndrome (PWS). Deletions of a region 35 kb upstream of exon 1 impair maternal imprinting and can cause Angelman syndrome (AS). So far, in all affected sibs with an imprinting defect, an inherited IC deletion was identified. We report on two sibs with AS who do not have an IC deletion but instead have a 1-1.5 Mb inversion separating the two IC elements. The inversion is transmitted silently through the male germline but impairs maternal imprinting after transmission through the female germline. Our findings suggest that the close proximity and/or the correct orientation of the two IC elements are/is necessary for the establishment of a maternal imprint. PMID- 11283797 TI - Mortality in neurofibromatosis 1: an analysis using U.S. death certificates. AB - Although neurofibromatosis 1 (NF1) is a relatively common autosomal dominant condition, information about its effect on mortality is limited. We used Multiple Cause Mortality Files, compiled from U.S. death certificates by the National Center for Health Statistics, for 1983 through 1997. We identified 3,770 cases of presumed NF1 among 32,722,122 deaths in the United States, a frequency of 1/8,700, which is one-third to one-half the estimated prevalence. Mean and median ages at death for persons with NF1 were 54.4 and 59 years, respectively, compared with 70.1 and 74 years in the general population. Results of proportionate mortality ratio (PMR) analyses showed that persons with NF1 were 34 times more likely (PMR=34.3, 95% confidence interval [CI] 30.8-38.0) to have a malignant connective or other soft-tissue neoplasm listed on their death certificates than were persons without NF1. Overall, persons with NF1 were 1.2 times more likely than expected (PMR=1.21, 95% CI 1.14-1.28) to have a malignant neoplasm listed on their death certificates, but the PMR was 6.07 (95% CI 4.88-7.45) for persons who died at 10-19 years of age and was 4.93 (95% CI 4.14-5.82) for those who died at 20-29 years of age. Similarly, vascular disease was recorded more often than expected on death certificates of persons with NF1 who died at <30 years of age (PMR=3.26, 95% CI 1.31-6.71 at age <10 years; PMR=2.68, 95% CI 1.38-4.68 at age 10-19 years; and PMR=2.25, 95% CI 1.46-3.32 at 20-29 years) but not in older persons. This study supports previous findings of decreased life expectancy for persons with NF1 and, within the limitations of death certificates, provides population-based data about NF1 morbidity and mortality that are useful to clinicians caring for patients with NF1. PMID- 11283798 TI - Localization of a gene (MCUL1) for multiple cutaneous leiomyomata and uterine fibroids to chromosome 1q42.3-q43. AB - Dominant transmission of multiple uterine and cutaneous smooth-muscle tumors is seen in the disorder multiple leiomyomatosis (ML). We undertook a genomewide screen of 11 families segregating ML and found evidence for linkage to chromosome 1q42.3-q43 (maximum multipoint LOD score 5.40). Haplotype construction and analysis of recombinations permitted the minimal interval containing the locus, which we have designated "MCUL1," to be refined to an approximately 14-cM region flanked by markers D1S517 and D1S2842. Allelic-loss studies of tumors indicated that MCUL1 may act as a tumor suppressor. Identification of MCUL1 should have wide interest, since this gene may harbor low-penetrance variants predisposing to the common form of uterine fibroids and/or may undergo somatic mutation in sporadic leiomyomata. PMID- 11283800 TI - Severe babesiosis in Long Island: review of 34 cases and their complications. AB - Thirty-four consecutive patients were hospitalized with diagnosis of severe Babesia infection over the course of 13 years. The average time from onset of symptoms to diagnosis was 15 days. When compared with uninfected febrile control patients, affected patients complained significantly more often of malaise, arthralgias and myalgias, and shortness of breath (P<.05), and they more often had thrombocytopenia and abnormal liver function (P<.05). Forty-one percent of patients with Babesia developed complications such as acute respiratory failure, disseminated intravascular coagulation, congestive heart failure, and renal failure. Analysis of data revealed that complicated babesiosis was more commonly associated with the presence of severe anemia (hemoglobin level <10 g/dL; P=.01) and higher parasitemia levels (>10%; P=.08). Patients were treated with a combination of drugs that included clindamycin, quinine, atovaquone, or azithromycin. Despite treatment, parasitemia persisted for an average of 8.5 days (range, 3--21 days). Exchange transfusion was performed for 7 patients, and it effectively reduced the high levels of parasitemia. Three patients died. Improved outcomes may result with prompt recognition and treatment of babesiosis. PMID- 11283799 TI - Up-regulation of WNT-4 signaling and dosage-sensitive sex reversal in humans. AB - Wnt-4, a member of the Wnt family of locally acting secreted growth factors, is the first signaling molecule shown to influence the sex-determination cascade. In mice, a targeted deletion of Wnt-4 causes the masculinization of XX pups. Therefore, WNT-4, the human homologue of murine Wnt-4, is a strong candidate gene for sex-reversal phenotypes in humans. In this article, we show that, in testicular Sertoli and Leydig cells, Wnt-4 up-regulates Dax1, a gene known to antagonize the testis-determining factor, Sry. Furthermore, we elucidate a possible mechanism for human XY sex reversal associated with a 1p31-p35 duplication including WNT-4. Overexpression of WNT-4 leads to up-regulation of DAX1, which results in an XY female phenotype. Thus, WNT-4, a novel sex determining gene, and DAX1 play a concerted role in both the control of female development and the prevention of testes formation. These observations suggest that mammalian sex determination is sensitive to dosage, at multiple steps in its pathway. PMID- 11283801 TI - Rotavirus gastroenteritis in Italian children: can severity of symptoms be related to the infecting virus? AB - The aim of our study was to determine whether the severity of rotavirus gastroenteritis may be related to the different characteristics of infecting viral strains. The severity of clinical symptoms in 401 children with acute rotavirus gastroenteritis was assessed using a scoring system for frequency and duration of vomiting, diarrhea, and fever, as well as the patients' requirements for intravenous rehydration. Rotavirus strains were characterized by determining the electropherotype of their double-stranded RNA, the G type and subgroup by a panel of monoclonal antibodies, and the P type by reverse transcription polymerase chain reaction. Strains with a short electropherotype, G2P[4] type, and subgroup I were associated with more-severe gastroenteritis and affected children older than those infected with strains with a long electropherotype, G1P[8] or G4P[8] type, and subgroup II. Minor differences in clinical symptoms were also detected in children infected with different long electropherotypes and with G1P[8] and G4P[8] specificities. PMID- 11283802 TI - Resurgence of blackwater fever in long-term European expatriates in Africa: report of 21 cases and review. AB - Blackwater fever (BWF) is a severe clinical syndrome, characterized by intravascular hemolysis, hemoglobinuria, and acute renal failure that is classically seen in European expatriates chronically exposed to Plasmodium falciparum and irregularly taking quinine. BWF virtually disappeared after 1950, when chloroquine superseded quinine. We report 21 cases of BWF seen in France from 1990 through 1999 in European expatriates who lived in sub-Saharan Africa. All patients had macroscopic hemoglobinuria, jaundice, and anemia. Acute renal failure occurred in 15 patients (71%), 7 of whom required dialysis. The presumed triggers of BWF were halofantrine (38%), quinine (24%), mefloquine (24%), and halofantrine or quinine (14%). Glucose-6-phosphate dehydrogenase (G6PD) activity was normal in the 14 patients who underwent this test. Low-level P. falciparum parasitemia was found in 8 patients. All 21 patients survived. Our data and 13 cases reported in the literature suggest a resurgence of classic BWF among Europeans living in Africa and a need to discuss attendant therapeutic implications. PMID- 11283803 TI - Microbial etiology of community-acquired pneumonia in the adult population of 4 municipalities in eastern Finland. AB - To determine the etiology of community-acquired pneumonia in the adult population of a defined area, specific antibody responses in paired serum samples, levels of circulating pneumococcal immune complexes in serum samples, and pneumococcal antigen in urine were measured. Samples (304 paired serum samples and 300 acute urine samples) were obtained from 345 patients > or =15 years old with community acquired, radiologically confirmed pneumonia, which comprised all cases in the population of 4 municipalities in eastern Finland during 1 year. Specific infecting organisms were identified in 183 patients (including 49 with mixed infection), as follows: Streptococcus pneumoniae, 125 patients; Haemophilus influenzae, 12; Moraxella catarrhalis, 8; chlamydiae, 37 (of which, Chlamydia pneumoniae, 30); Mycoplasma pneumoniae, 30; and virus species, 27. The proportion of patients with pneumococcal infections increased and of those with Mycoplasma infections decreased with age, but for each age group, the etiologic profile was similar among inpatients and among outpatients. S. pneumoniae was the most important etiologic agent. The annual incidence of pneumococcal pneumonia per 1000 inhabitants aged > or =60 years was 8.0. PMID- 11283804 TI - Constancy of distribution of serogroups of invasive pneumococcal isolates among children: experience during 4 decades. AB - Serogroups of pneumococci that caused bacteremia or meningitis in children were examined from 1981 through 1998 at Boston City Hospital/Boston Medical Center. There were 410 episodes of pneumococcal bacteremia (13--36 cases per year), of which 14 occurred in human immunodeficiency virus (HIV)--infected children and 9 occurred in children with sickle-cell disease. The 7 most common serogroups were 14 (30.7% of isolates), 19 (11.7%), 6 (11%), 18 (10.7%), 9 (7.6%), 23 (7.3%), and 4 (5.6%). The rate of episodes due to serogroups 4, 6, 9, 14, 18, 19, and 23 ranged from 80% to 91.9% during the study period. The rate of episodes due to serogroups 4, 6, 14, 18, 19, and 23 was 84.6% among patients with HIV infection, 100% among patients with sickle-cell disease, and 94.1% among the 18 patients for whom cultures of CSF specimens revealed pneumococcal meningitis. The results demonstrate that type 14 was the dominant pneumococcal serogroup responsible for invasive disease throughout the 18-year study period and that serogroup distribution overall remained constant. A comparison of these findings with historical pediatric data from our institution showed serogroup stability dating back to 1957. PMID- 11283805 TI - Extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae: risk factors for infection and impact of resistance on outcomes. AB - The prevalence of antibiotic resistance among extended-spectrum beta-lactamase (ESBL)--producing Escherichia coli and Klebsiella pneumoniae has increased markedly in recent years. Thirty-three patients with infection due to ESBL producing E. coli or K. pneumoniae (case patients) were compared with 66 matched controls. Total prior antibiotic use was the only independent risk factor for ESBL-producing E. coli or K. pneumoniae infection (odds ratio, 1.10; 95% confidence interval, 1.03--1.18; P=.006). Case patients were treated with an effective antibiotic a median of 72 hours after infection was suspected, compared with a median of 11.5 hours after infection was suspected for controls (P<.001). ESBL-producing E. coli or K. pneumoniae infection was associated with a significantly longer duration of hospital stay and greater hospital charges (P=.01 and P<.001, respectively). Finally, many ESBL-producing E. coli and K. pneumoniae isolates were closely related. ESBL-producing E. coli and K. pneumoniae infections have a significant impact on several important clinical outcomes, and efforts to control outbreaks of infection with ESBL-producing E. coli and K. pneumoniae should emphasize judicious use of all antibiotics as well as barrier precautions to reduce spread. PMID- 11283806 TI - Brucellosis in pregnant women. AB - Brucella species occasionally cause spontaneous human abortion, but theories regarding whether they do so more frequently than do other infectious pathogens remain controversial. We reviewed 92 pregnant women who presented with acute brucellosis at a Saudi Arabian hospital. From 1983 through 1995, the cumulative incidence of pregnancy and brucellosis was 1.3 cases per 1000 delivered obstetrical discharges. The incidence of spontaneous abortion in the first and second trimesters was 43%, and the incidence of intrauterine fetal death in the third trimester was 2%. Antepartum antimicrobial therapy with cotrimoxazole or cotrimoxazole/rifampin was protective against spontaneous abortion (relative risk, 0.14; 95% confidence interval, 0.06--0.37; P<.0001). The beneficial effect of treatment occurred in women with febrile illness; vaginal bleeding at presentation usually led to spontaneous abortion. This study demonstrated that the incidence of spontaneous abortion among pregnant women with brucellosis is high and that these women should receive prompt therapy with antimicrobial agents when they present for medical care. PMID- 11283807 TI - A case of cutaneous ulcerative alternariosis: rare association with diabetes mellitus and unusual failure of itraconazole treatment. AB - Alternaria species are ubiquitous dematiaceous fungi that are increasingly recognized as pathogens in immunocompromised patients or those with significant underlying disease, but they are also pathogens in otherwise healthy hosts. We describe a case of dermal cutaneous ulcerative alternariosis in a frail 83-year old patient with diet-controlled diabetes mellitus. Histological analysis revealed hyphal morphology in tissue sections that was initially confused with that of a zygomycetous fungus, and multiple positive culture results were necessary to identify the organism. Treatment with oral itraconazole and surgical debridement were ineffective; clinical improvement was achieved by means of treatment with intravenous amphotericin B lipid complex. We review the literature regarding the role of diabetes mellitus in cutaneous alternariosis and regarding the efficacy of treatment with itraconazole, which has been used very successfully. To our knowledge, this is only the second case report noting diabetes mellitus uncomplicated by steroid administration as a possible predisposing factor for cutaneous infection. PMID- 11283808 TI - Derriere to the future: is it time to rethink how we use antimicrobial agents? PMID- 11283809 TI - Prophylactic antifungal therapy in the intensive care unit. AB - Antifungal prophylaxis is regularly used during treatment of patients with some cancers, as subgroups with high rates of invasive fungal infections are readily identified; for these patients, prophylaxis has been shown to be of value. High risk liver transplant recipients also benefit from antifungal prophylaxis. Although the idea of extending this concept to the prevention of candidal infections in the larger population of critically ill patients who are seen in the intensive care unit (ICU) and who do not have neutropenia is attractive, implementation of this strategy is difficult because of the widely varying characteristics of patients in the ICU. Two studies have shown the benefit of such prophylaxis, but the benefit was shown only in selected groups of patients who had an unusually high risk for invasive candidiasis. Although the concept is sound, broad-scale implementation of antifungal prophylaxis would be premature and costly, both financially and with regard to resistance and toxicity. Investigations are needed to define and prove the utility of predictive tools for the identification of patients in the ICU who would benefit from prophylaxis. PMID- 11283810 TI - Campylobacter jejuni Infections: update on emerging issues and trends. AB - Infection with Campylobacter jejuni is one of the most common causes of gastroenteritis worldwide; it occurs more frequently than do infections caused by Salmonella species, Shigella species, or Escherichia coli O157:H7. In developed countries, the incidence of Campylobacter jejuni infections peaks during infancy and again during early adulthood. Most infections are acquired by the consumption and handling of poultry. A typical case is characterized by diarrhea, fever, and abdominal cramps. Obtaining cultures of the organism from stool samples remains the best way to diagnose this infection. An alarming recent trend is the rapid emergence of antimicrobial agent--resistant Campylobacter strains all over the world. Use of antibiotics in animals used for food has accelerated this trend. It is fortunate that complications of C. jejuni infections are rare, and most patients do not require antibiotics. Guillain-Barre syndrome is now recognized as a post-infectious complication of C. jejuni infection, but its incidence is <1 per 1000 infections. Careful food preparation and cooking practices may prevent some Campylobacter infections. PMID- 11283811 TI - Mortality among human immunodeficiency virus-infected patients with cirrhosis or hepatocellular carcinoma due to hepatitis C virus in French Departments of Internal Medicine/Infectious Diseases, in 1995 and 1997. AB - Several studies have suggested that the progression of hepatitis C virus (HCV) infection is more severe in patients infected by the human immunodeficiency virus (HIV). Two national retrospective multicenter cohort surveys were performed in France that included 17,487 HIV-infected patients during 1995 and 26,497 during 1997. The following data was evaluated: total number of deaths; number of deaths linked to AIDS, cirrhosis, or hepatocellular carcinoma (HCC); and number of deaths related to other (non-HCV--linked) causes. In 1995, the causes of death were as follows: AIDS, 1307 (7.47%); cirrhosis or HCC, 21 (0.12%); and other (non HCV--linked) causes, 99 (0.56%). In 1997, the causes of deaths were as follows: AIDS, 459 (1.73%); cirrhosis or HCC 36 (0.13%); and other (non-HCV--linked) causes, 48 (0.18%). Comparative results between the 1995 and 1997 surveys showed a dramatic decrease in AIDS-related mortality rates (7.47% vs. 1.73%; P<.001) but not in HCV-related mortality rates (0.06% vs. 0.07%; P=.79). In France, despite the high prevalence of HCV infection in HIV-positive patients, the mortality rate in 1995 and 1997 caused by HCV-related cirrhosis or HCC was low. PMID- 11283812 TI - Multicenter case-control study of risk factors for histoplasmosis in human immunodeficiency virus-infected persons. AB - We conducted a multicenter case-control study to identify risk factors for histoplasmosis among persons with acquired immunodeficiency syndrome (AIDS) and to evaluate predictors of a poor outcome (defined as death or admission to the intensive care unit). Patients with histoplasmosis were each matched by age, sex, and CD4 lymphocyte count to 3 controls. From 1996 through 1999, 92 case patients and 252 controls were enrolled. Of the case patients, 81 (89%) were men, 50 (55%) were black, 78 (85%) had a CD4 lymphocyte count of <100 cells/microL, 80 (87%) were hospitalized, and 11 (12%) died. Multivariable analysis found that receipt of antiretroviral therapy and of triazole drugs were independently associated with a decreased risk of histoplasmosis. Chronic medical conditions and a history of infections with herpes simplex virus were associated with poor outcome. Triazoles should be considered for chemoprophylaxis for persons with AIDS, especially those who take part in high-risk activities that involve frequent exposure to soil, who have CD4 lymphocyte counts of <100 cells/microL, and who live in areas where histoplasmosis is endemic. PMID- 11283813 TI - Avascular necrosis of bone in patients with human immunodeficiency virus infection: report of 6 cases and review of the literature. AB - In 1998 and 1999, we diagnosed avascular necrosis of bone in 6 patients in our human immunodeficiency virus clinic practice, an incidence of 0.45%, which is 45 times greater than would be expected in the general population. Antiphospholipid antibodies and hyperlipidemia secondary to protease inhibitor therapy have been implicated as possible etiologies; however, these abnormalities cannot explain all cases of avascular necrosis of bone reported in patients with human immunodeficiency virus infection. PMID- 11283814 TI - Erroneously low or undetectable plasma human immunodeficiency virus type 1 (HIV 1) ribonucleic acid load, determined by polymerase chain reaction, in West African and American patients with non-B subtype HIV-1 infection. AB - The polymerase chain reaction (PCR) assay for plasma human immunodeficiency virus type 1 (HIV-1) ribonucleic acid (RNA) inadequately quantitates virus load for some non-B HIV-1 subtypes because of genetic diversity in the gag region targeted by the PCR primers. Unexpectedly low or undetectable plasma HIV-1 RNA findings by PCR were a clue to non-B HIV-1 infections in patients in whom plasma HIV-1 RNA was found to be substantially higher when determined by a branched-chain deoxyribonucleic acid assay. PMID- 11283815 TI - Effects of virologic rebound on CD4 cell counts. AB - A retrospective study was conducted to assess the effects of various degrees of virologic rebound on CD4 cell counts over time in human immunodeficiency virus infected patients. We found that (1) the higher the degree of virologic rebound, the more rapid the decrease in CD4 cell counts over time, and (2) the magnitude of the virologic rebound was inversely correlated with the CD4 cell counts at the time of rebound. PMID- 11283816 TI - Cryptococcus myelitis: atypical presentation of a common infection. AB - Cryptococcus neoformans is associated with as much as 45% of meningitis in patients admitted for hospital care in Zimbabwe, and it is an important opportunistic infection in patients infected with the human immunodeficiency virus. Cases of cryptococcosis presenting as a spinal cord syndrome have been reported from Zimbabwe and South Africa, but these were all cases of Cryptococcus vertebral osteomyelitis. We describe 3 unusual patients who presented with a myelitis-like syndrome without vertebral osteomyelitis. PMID- 11283817 TI - Fusarium infection after solid-organ transplantation. AB - We describe a case of soft tissue infection caused by Fusarium species in a heart liver transplant recipient, and review the cases of fusarial infection reported among solid-organ transplant (SOT) recipients. Unlike fusarial infection in patients with hematologic malignancies or bone marrow transplants, fusarial infection in SOT recipients tends to be localized, occurs later in the posttransplantation period, and has a better outcome. Surgical resection, when possible, and prolonged treatment with amphotericin provide the most effective form of therapy. PMID- 11283818 TI - Encephalitis without meningitis due to sandfly fever virus serotype toscana. AB - The role of Toscana (TOS) virus in producing encephalitis without meningitis is uncertain. We studied 2 cases of TOS virus encephalitis without meningitis by means of nested polymerase chain reaction assay and DNA sequencing. Findings confirm that TOS virus may directly cause encephalitis and suggest the usefulness of DNA sequencing in investigating relationships between TOS virus molecular patterns and the spectrum of neurological involvement. PMID- 11283821 TI - Visceral abscesses due to Brucella suis infection in a retired pig farmer. AB - A 78-year-old retired pig farmer developed brucellosis and died of liver failure >20 years after her last exposure to infected livestock. This is an exceptionally long incubation period for this infection, which usually presents within weeks of exposure. PMID- 11283820 TI - Probable locally acquired mosquito-transmitted malaria in Georgia, 1999. AB - In July 1999, the Centers for Disease Control and Prevention received notification of a case of malaria in a 32-year-old female native of Colquitt County, Georgia, who had no history of travel into an area where malaria transmission is endemic. An epidemiological investigation confirmed the absence of risk factors, such as blood transfusion, organ transplantation, malariotherapy, needle sharing, or past malaria infection. Active case finding revealed no other infected persons in Colquitt County. Light trapping and larvae dipping failed to identify adult or larval anophelines; however, Colquitt County is known to be inhabited by Anopheles quadrimaculatus, a competent malaria vector. The patient's home was located near housing used by seasonal migrant workers from regions of southern Mexico and Central America where malaria is endemic, one of whom may have been the infection source. The occurrence of malaria in this patient with no risk factors, except for proximity to potentially gametocytemic hosts, suggests that this illness probably was acquired through the bite of an Anopheles species mosquito. PMID- 11283822 TI - Endoscopic treatment of inverted papilloma: safety and efficacy. AB - PURPOSE: Inverted papillomas of the nose and paranasal sinuses are uncommon neoplasms, characterized by their tendency to recur and by their association with malignancy. Over the past 25 years, the standard surgical treatment of inverted papillomas has involved extranasal, en bloc resections because of high recurrence rates with less extensive surgery. The past decade has witnessed an increasing number of reports documenting the successful treatment of inverted papilloma with endoscopic approaches. This investigation examines the role of endoscopic procedures in treating inverted papilloma. MATERIALS AND METHODS: The present article presents a thorough review of the literature relating to the history, pathology, diagnosis, and treatment of inverted papillomas of the nose and paranasal sinuses. It also provides a systematic review of the pertinent medical literature with pooled statistical analysis of data from 33 studies involving 1,426 patients. RESULTS AND CONCLUSIONS: The findings suggest that many cases of inverted papilloma can be treated successfully with low recurrence rates and less morbidity through endoscopic surgery, with results comparable to extranasal procedures. Less aggressive approaches to the sinuses and nonendoscopic intranasal procedures, such as the Caldwell-Luc, have unacceptable rates of recurrence and should be abandoned. A staging system for inverted papilloma and a logical approach to treatment are presented. PMID- 11283823 TI - Endoscopic sinus surgery in previously irradiated patients. AB - PURPOSE: Our purpose was to evaluate the safety and efficacy of endoscopic sinus surgery in irradiated patients with absolute indications for sinus surgery. PATIENTS AND METHODS: During 5 years at a tertiary referral center, more than 200 patients received irradiation to a field that included the paranasal sinuses. Complaints related to the sinuses are common in such patients and often include crusting and increased mucus drainage. Six patients presented with significant sinus infections in the absence of tumor recurrence and failed medical management. Additional problems included epiphora and nasal obstruction caused by cicatricial choanal stricture. Surgical interventions included ethmoidectomy, multiple osteotomies, debridement of scarred or devitalized tissue, and dacryocystorhinostomy. Outcome measures included intraoperative findings and complications, length of hospital stays, endoscopic assessments of the healing over 6 months post-operatively, and improvement or persistence of symptoms over 2 to 3 years of follow-up. RESULTS: Surgery can be technically difficult because of derangements of normal anatomy and dehiscence of important structures. Although bleeding problems, prolonged admission, and delayed healing were noted in certain cases, they did not result in long-term morbidity. CONCLUSIONS: Endoscopic sinus surgery has become an invaluable tool in the treatment of refractory sinusitis. Our literature review has revealed no information, however, regarding endoscopic sinus surgery in previously irradiated patients. Theoretically, such patients are at risk for healing problems and anatomic derangements, which could lead to complications. There is, nevertheless, a theoretical benefit to avoiding external approaches in patients who might heal poorly. PMID- 11283824 TI - Clinical analysis of multiple primary malignancies of the hypopharynx and esophagus. AB - PURPOSE: Because the capability to control squamous cell carcinomas of the head and neck has improved recently, the phenomenon of multiple primary malignancies of that region is now recognized with increasing frequency. We reviewed cases of multiple primary squamous cell carcinomas of the hypopharynx and esophagus with regard to their frequency, incidence, and prognosis. PATIENTS AND METHODS: We reviewed 104 cases of hypopharyngeal cancer to determine (1) if and when esophageal cancer occurred, (2) the classification of multiple tumors as metachronous or synchronous, and (3) tumor histology. RESULTS: In most cases of the metachronous type, esophageal cancer followed hypopharyngeal cancer within less than 3 years. Most cases of hypopharyngeal cancer were at an advanced stage, in contrast to esophageal cancer, which were all early stage. These cases had a poor prognosis despite various treatments causing local disease to be well controlled. Endoscopic esophageal mucosal resection was found to be an effective treatment for esophageal cancer, especially in superficial types. CONCLUSIONS: The prognosis and mild systemic damage after endoscopic esophageal mucosal resection compare favorably with surgery, radiation, or systemic chemotherapy. PMID- 11283825 TI - Intratympanic gentamicin therapy for vertigo in nonserviceable ears. AB - PURPOSE: Intratympanic ototoxic agents have become a widely accepted means of managing vertigo in patients with Meniere's disease while preserving residual hearing. We investigated expanding the indications for intratympanic gentamicin to include control of vertigo in patients without serviceable hearing in the involved ear caused by a variety of end-organ pathologies. MATERIALS AND METHODS: We present a retrospective series of 6 patients suffering from vertigo caused by end-organ disease, in an ear without serviceable hearing. Two patients suffered from delayed endolymphatic hydrops, 3 from Meniere's disease, and 1 from poststapedectomy vertigo. These patients chose unilateral vestibular ablation with serial intratympanic gentamicin injection rather than labyrinthectomy for a variety of reasons. Conventional electronystagmography (ENG) testing and audiometry were completed on all patients. The ENG testing included bithermal calorics and rotational testing. All patients had a magnetic resonance image with gadolinium to exclude retrocochlear or central pathology. Rotational testing was repeated before each injection and at the conclusion of therapy to assess changes in the peripheral vestibular response. The patients' subjective response to therapy was followed. RESULTS: Follow-up has been 10 to 69 months with successful control of vertigo in all patients. CONCLUSIONS: Intratympanic gentamicin therapy offers a minimally invasive, ambulatory, low morbidity, cost-effective means of managing vertigo in patients with nonserviceable hearing. PMID- 11283826 TI - Peripheral eosinophilia in the diagnosis of chronic rhinosinusitis. AB - PURPOSE: To determine the relationship between peripheral blood eosinophilia and chronic rhinosinusitis. MATERIALS AND METHODS: A retrospective review was conducted of consecutive operative cases during 1 calendar year. The preoperative complete blood count (CBC) were tabulated for three groups of patients: those undergoing endoscopic sinus surgery, those undergoing septoplasty with turbinate reduction alone, and a nonrhinologic control group. Statistical analysis was performed to determine differences in the components of the CBC among these three groups of patients and to identify significant associations between abnormal peripheral eosinophil counts and these diagnoses. RESULTS: A total of 87, 32, and 92 patients were identified for the endoscopic sinus surgery (ESS), septoplasty, and control groups, respectively. Significant differences in the percentages of eosinophils, lymphocytes, and neutrophils were noted among the three groups (P <.05). Comparison among groups indicated that ESS patients had significantly higher percent peripheral eosinophilia when compared with both the control group and septoplasty group (P <.001 and P =.010, respectively); no significant difference was noted between the septoplasty group and the control group (P =.627). The sensitivity and specificity of the peripheral eosinophil count for chronic sinusitis were 49.4% and 84.7%, respectively. CONCLUSIONS: The peripheral eosinophil count in chronic sinusitis is elevated compared with both a nonrhinologic control group and a group of patients with septal deviation. Furthermore, abnormally elevated eosinophil counts are associated with chronic sinusitis but not chronic rhinitis alone. However, this association is not strong enough to be used in the diagnosis of chronic sinusitis because of poor sensitivity. PMID- 11283827 TI - Genetic deletions of glutathione-S-transferase as a risk factor in squamous cell carcinoma of the larynx: a preliminary report. AB - PURPOSE: To test whether genetic deletions of glutathione-S-transferase (GST) are associated with squamous cell carcinoma (SCC) of the larynx. GST are a group of detoxifying enzymes that may help reduce the risk of developing cancer in response to environmental carcinogens. Polycyclic aromatic hydrocarbons, found in high concentration in cigarette smoke, are known carcinogens especially for SCC of the larynx. Individuals with absolute or relative deficiency of the GST enzyme system may therefore be at a higher risk of developing laryngeal carcinoma. MATERIALS AND METHODS: Genotyping for GST-M1 and GST-T1 was performed using polymerase chain reaction (PCR) assay on fresh frozen tissue specimens of 20 patients with SCC of the larynx and on 20 control subjects with a similar smoking history. Because this assay results in the absence of a PCR product in individuals expressing the GST-M1/GST-T1 null genotype, oligonucleotide primers that amplify a portion of the albumin gene were included in a multiplex PCR as a positive control for DNA quality and PCR conditions. The chi-square test was used for statistical analysis. RESULTS: GST-M1 gene was deleted in 80% of patients with laryngeal SCC and in 50% of control subjects (P <.05). No statistically significant difference was observed in the incidence of GST-T1 gene deletion in patients with SCC of the larynx and control subjects. CONCLUSION: GST-M1 gene deletion was significantly associated with SCC of the larynx and may produce a risk for this particular disease. PMID- 11283828 TI - What are the nonsurgical treatment options for obstructive sleep apnea syndrome? AB - Obstructive sleep apnea (OSA) syndrome is now recognized as a relatively common cause of excessive daytime sleepiness, with resultant psychosocial impairment and motor vehicle accidents, and it likely contributes to premature cardiovascular disease. Treatment is naturally directed at the upper airway; however, it is also important to identify and correct significant risk factors, such as obesity and hypothyroidism, whenever possible. Oral appliances or nasal continuous positive airway pressure may immediately reverse symptoms caused by OSA and can be used either indefinitely or as a bridge to potentially definitive surgery. PMID- 11283829 TI - Woman presenting with a postauricular mass. PMID- 11283830 TI - Amyloidoma of the nose in a pediatric patient: a case report. AB - Localized amyloidosis in the head and neck is a rare and benign disease. Larynx is the most common site of involvement and accounts for 0.2% to 1.5% of benign laryngeal tumors. The oral cavity and pharynx may also be involved in localized amyloidosis of the head and neck. There are only 7 cases of localized nasopharyngeal amyloidosis, and 8 cases of localized nasal amyloidosis reported to date. Out of these 8 cases of nasal amyloidosis, only 1 of them is a pediatric patient. We present the second reported case of localized nasal amyloidosis in a pediatric patient. This case report describes a 10-year-old girl with a 1-year history of right-sided nasal obstruction and mucoid discharge. Examination revealed an irregular erythematous, waxy gray mass arising from the right inferior turbinate. Computed tomography and magnetic resonance imaging showed the mass arising from the inferior turbinates, whereas other investigations excluded systemic amyloidosis. Histology from surgical excision revealed amyloidosis. This case illustrates that although amyloidoma of the nose is rare, it should also be considered as part of the differentials of a nasal mass even in pediatric patients. [Editorial comment: These authors demonstrate that an index of suspicion and confirmation of that suspicion through biopsy and subsequent pathologic evaluations Maybe the only way to avoid missing this potentially important cause of nasal obstruction.] PMID- 11283831 TI - Osteomyelitis of the temporomandibular joint. AB - Osteomyelitis of the temporomandibular joint is very uncommon, and osteomyelitis as a result of Aspergillus niger infection has not previously been reported. A case report of skull base and condylar osteomyelitis is presented. Previously reported cases of temporomandibular joint osteomyelitis are reviewed, and management is discussed. Because of the significant morbidity possible with infections in this region, otolaryngologists should be familiar with the anatomy, diagnostic modalities, and therapeutic options. [Editorial comment: This unusual case presents unique aspects of the pathophysiology of osteomyelitis of the skull base.] PMID- 11283832 TI - Radiation-induced malignant fibrous histiocytoma of the neck in a patient with laryngeal carcinoma. AB - Fibrohistiocytomas are soft tissue tumors of histiocytic origin that have a variety of histological patterns. Although cases of malignant fibrous histiocytoma (MFH) of the head and neck have been reported with increasing frequency in recent years, they are considered rare. We report a case of the giant cell variant of MFH of the neck in which the patient had been given radiotherapy for T1 glottic cancer. Prognosis of MFH, the use of radiation as primary treatment, and its role in the development of secondary primary tumors in the head and neck region are reviewed. [Editorial comment: The authors stress the important relationship between prior radiation therapy and the induction of new tumors.] PMID- 11283833 TI - Chondromyxoid fibroma of the nasal bone with extension into the frontal and ethmoidal sinuses: report of one case and a review of the literature. AB - Chondromyxoid fibroma is a rare benign tumor that usually occurs in the long bones. A 50-year-old patient presented with chondromyxoid fibroma of the nasal bone with extension into the frontal and ethmoidal sinuses. This is the fourth case reported to date in the literature. The clinical manifestations of the tumor were very limited, and the appearance at rhinoscopy was misleading. Radiologic imaging showed a soft tissue lesion invading the adjacent bony structures and the dura mater. Surgery was performed by a combined team of otorhinolaryngologists and neurosurgeons, and total excision of the tumor was achieved. The histologic diagnosis of this tumor is difficult because of its similarities to chondrosarcoma. [Editorial comment: The authors concisely review management of this rare tumor, emphasizing that complete surgical excision, rather than curettage, is required for long term control.] PMID- 11283834 TI - Sharp foreign bodies in the tracheobronchial tree. AB - Aspiration and/or ingestion of foreign bodies is a common occurrence. Six cases of scarf pin aspiration are described. Scarf pin inhalation as a cultural and ethnic hazard in an Arab woman is highlighted. [Editorial comment: The authors illustrate the dangers of holding a straight pin in the mouth. Management of these sharp, potentially penetrating foreign bodies is described.] PMID- 11283835 TI - Cervical manifestation of blastomycosis. AB - Blastomycosis is a common systemic fungal infection in which the physical and radiographic findings appear far more serious than the subjective signs and symptoms. Although blastomycosis of the head and neck is often difficult to diagnose, clues in the patient's history and a few laboratory tests can establish the diagnosis. Involvement of the skin and soft tissues provides an unusual opportunity for direct access to the organism for culture and pathologic diagnosis. We present a case of blastomycosis presenting in a single abscessed cervical lymph node. The diagnosis was established by fine-needle aspiration of the cervical mass. The primary lung disease was confirmed by chest radiography and computed tomography. The clinical and pathologic features of the disease are discussed. [Editorial comment: Pulmonary blastomycosis may present with cervical adenopathy. This article demonstrates the potential value of fine needle aspiration in establishing this diagnosis.] PMID- 11283836 TI - Facio-cervical transfixion by a metallic rod: a case report. AB - Foreign bodies penetrating into the neck may cause extensive neurovascular and aerodigestive tract injuries. Facio-cervical transfixion by a metallic rod without such injuries is very rare. The patient presented in this article reported to our center 5 hours after the road traffic accident with impacted hollow metallic rod traversing through the tongue, pharynx, and neck after causing fracture to maxilla. Exploration under general anesthesia after tracheostomy resulted in an uneventful extraction of the metallic rod. Postoperatively, the patient recovered completely and had no neurologic deficits. [Editorial comment: This case report demonstrates an approach to an extremely difficult and potentially dangerous clinical problem. The authors decision to forego angiography is based on physical examination with evidence of palpable flow in the superficial temporal artery.] PMID- 11283838 TI - Molecular mimicry and primary biliary cirrhosis: premises not promises. PMID- 11283839 TI - Cholestatic expression pattern of sinusoidal and canalicular organic anion transport systems in primary cultured rat hepatocytes. AB - To maintain ongoing vectorial bile secretion, hepatocytes localize distinct transport systems at their basolateral and canalicular membrane domains. Here we compare the expression of the basolateral Na(+)-taurocholate cotransporter (Ntcp) and organic anion transporting polypeptides 1 and 2 (Oatp1, Oatp2) and the canalicular bile salt export pump (Bsep) and multidrug resistance-associated protein 2 (Mrp2) in primary cultured rat hepatocytes. During 72 hours of culturing time the messenger RNA (mRNA) and protein levels of Ntcp and Oatp1 decreased in parallel to 2% to 7% of initial values at 3 hours. Although Oatp2 mRNA exhibited a similar down-regulation to 4%, Oatp2 protein and function were maintained at 25% to 47% of initial values. Furthermore, Bsep and Mrp2 protein levels were maintained at about 50%, while the Mrp2 mRNA showed a transient up regulation to 154% at 24 and 48 hours. Also, induction of Mrp1 mRNA and protein was observed starting after 24 hours. These results indicate transcriptional down regulation or decreased mRNA stability of Ntcp and Oatp1, transcriptional and posttranslational regulation of Oatp2, Bsep, and Mrp2 and transcriptional up regulation of Mrp1 in primary cultured hepatocytes. Furthermore, and most importantly, the observed changes in transporter expression closely resemble the altered transporter phenotypes of cholestatic and proliferating hepatocytes in vivo, thus indicating that primary cultured hepatocytes acquire a cholestatic phenotype, and that the transporter expression might be a suitable differentiation marker for maintenance of hepatocytes in vitro. PMID- 11283840 TI - Cellular localization and up-regulation of multidrug resistance-associated protein 3 in hepatocytes and cholangiocytes during obstructive cholestasis in rat liver. AB - The hepatic expression of the ATP-dependent conjugate export pump multidrug resistance-associated protein 2 (Mrp2) is diminished in experimentally induced models of cholestasis. In this study we have examined the localization and expression of Mrp3, another member of the multidrug resistance-associated protein family, in normal liver and after obstructive cholestasis in the rat. Indirect immunofluorescence and confocal microscopy were used to determine the tissue localization and Western blot analysis was performed to quantitate the expression. In normal rat liver Mrp3 was found on the basolateral membrane of cholangiocytes and a single layer of hepatocytes surrounding the central vein. Three and 7 days after bile duct ligation Mrp3 expression was significantly increased, predominantly in hepatocytes in the pericentral region. By 14 days all hepatocytes showed basolateral membrane labeling for Mrp3 at a time when apical Mrp2 staining was significantly diminished. Proliferating bile ducts continued to stain positive, although the intensity of staining did not seem to vary. After 14 days Western blot quantitation showed that Mrp3 had increased approximately 30 fold in total liver membranes. Quantitation of Mrp3 in membranes from isolated hepatocytes of livers of sham and common bile duct-ligated (CBDL) animals showed a significant up-regulation beginning at 1 day and continuing to increase through 14 days postligation. This was in contrast to the progressive decrease in Mrp2 protein. Because Mrp3 is capable of transporting toxic bile acids, up-regulation of Mrp3 may compensate for the down-regulation of Mrp2 in obstructive cholestasis. PMID- 11283841 TI - Heterogeneous response of antimitochondrial autoantibodies and bile duct apical staining monoclonal antibodies to pyruvate dehydrogenase complex E2: the molecule versus the mimic. AB - The 2-oxo-acid dehydrogenase complexes and, in particular, the E2 component of the pyruvate dehydrogenase complex (PDC) are the target of antimitochondrial antibodies (AMA). More than 95% of primary biliary cirrhosis (PBC) patients have detectable levels of autoantibodies to PDC-E2 and in general these react with a region of the molecule that contains the prosthetic group lipoic acid (LA). LA is vital to the function of the enzyme, although there is conflicting evidence as to whether its presence is required for PDC-E2 recognition by AMA. Some, but not all, monoclonal antibodies (mAbs) to PDC-E2 produce an intense staining pattern at the apical surface of bile duct epithelial cells (BEC) in patients with PBC, and it has been argued that the molecule at the apical surface of PBC bile duct cells is a modified form of PDC-E2 or a cross-reactive molecule, acting as a molecular mimic. Herein, we characterize the epitopes recognized by 4 anti-PDC-E2 mAbs that give apical staining patterns (3 mouse and 1 human). In particular, by using a combination of recombinant antigens, competitive inhibition assays, and a unique peptide-on-bead assay, we determined that these apically staining mAbs recognize 3 or 4 distinct epitopes on PDC-E2. More importantly, this suggests that a portion spanning the entire inner lipoyl domain of PDC-E2 can be found at the BEC apical surface. In addition, competition assays with patient sera and a PDC-E2-specific mAb showed significant epitope overlap with only 1 of the 3 mouse mAbs and showed a differential response to the peptide bound to beads. These findings further highlight the heterogeneous response of patient autoantibodies to the inner lipoyl domain of PDC-E2. PMID- 11283842 TI - A meta-analysis of endoscopic variceal ligation for primary prophylaxis of esophageal variceal bleeding. AB - Despite publication of several randomized trials of prophylactic variceal ligation, the effect on bleeding-related outcomes is unclear. We performed a meta analysis of the trials, as identified by electronic database searching and cross referencing. Both investigators independently applied inclusion and exclusion criteria, and abstracted data from each trial. Standard meta-analytic techniques were used to compute relative risks and the number needed to treat (NNT) for first variceal bleed, bleed-related mortality, and all-cause mortality. Among 601 patients in 5 homogeneous trials comparing prophylactic ligation with untreated controls, relative risks of first variceal bleed, bleed-related mortality, and all-cause mortality were 0.36 (0.26-0.50), 0.20 (0.11-0.39), and 0.55 (0.43 0.71), with respective NNTs of 4.1, 6.7, and 5.3. Among 283 subjects from 4 trials comparing ligation with beta-blocker therapy, the relative risk of first variceal bleed was 0.48 (0.24-0.96), with NNT of 13; however, there was no effect on either bleed-related mortality (relative risk [RR], 0.61; confidence interval [CI], 0.20-1.88) or all-cause mortality (RR, 0.95; CI, 0.56-1.62). In conclusion, compared with untreated controls, prophylactic ligation reduces the risks of variceal bleeding and mortality. Compared with beta-blockers, ligation reduces the risk for first variceal bleed but has no effect on mortality. Prophylactic ligation should be considered for patients with large esophageal varices who cannot tolerate beta-blockers. Subsequent research should further compare ligation and beta-blockers to determine the effect on mortality, and measure ligation's cost-effectiveness. PMID- 11283843 TI - Mitochondrial oxidative injury and energy metabolism alteration in rat fatty liver: effect of the nutritional status. AB - Hepatic steatosis is associated with mitochondrial oxidative alterations. This study aimed to characterize in a choline-deficient model of rat fatty liver whether this oxidative imbalance is related to an impairment of the capacity of ATP synthesis both under fed conditions and after starvation, which may sensitize mitochondria to oxidative injury. Mitochondria were isolated from normal and fatty livers of fed or 18-hour fasted rats. Oxidative injury was evaluated by measuring the mitochondrial content of thiobarbituric reactive substances, protein carbonyls, glutathione, and protein sulfhydryls. The mitochondrial F(0)F(1)-ATP synthase content, tissue ATP concentration, and liver histology were also determined. Compared with normal liver, under fed conditions, fatty livers showed a greater mitochondrial content of oxidized lipids and proteins together with a low concentration of sulfhydryls and glutathione. The mitochondrial catalytic beta-F(1) subunit of the F(0)F(1)-ATP synthase was about 35% lower in fatty livers. Hepatic ATP was also significantly reduced in fatty liver. Starvation exacerbated mitochondrial oxidative injury in both groups but to a greater extent in fatty livers. In the steatotic group, fasting induced a significant decrease of the ATP levels, which was accompanied by a 70% fall of the catalytic beta-F(1) subunit. These data indicate that the mitochondrial oxidative alterations in fatty livers are associated with an important reduction of the F(0)F(1)-ATP synthase. These changes, which are greatly exacerbated after starvation, may account for the reduced synthesis of the hepatic ATP observed in the presence of fatty infiltration. PMID- 11283844 TI - Endothelin-1 induces vasoconstriction on portal-systemic collaterals of portal hypertensive rats. AB - Portal hypertension is associated with increased hepatic and collateral resistance to an increased portal blood flow. Endothelin-1 (ET-1) can induce intrahepatic vasoconstriction and consequently increase portal pressure. It is unknown if ET-1 also modulates portal pressure by a direct vasoconstrictive effect on collaterals. This study investigated the collateral vascular responses to ET-1, the receptors in mediation, and the regulation of ET-1 action by nitric oxide and prostaglandin. The portal-systemic collaterals of partially portal vein ligated rats were tested by in situ perfusion. The concentration-response curves of collaterals to graded concentrations of ET-1 (10(-10)-10(-7) mol/L) with or without BQ-123 (ET(A) receptor antagonist, 2 x 10(-6) mol/L), BQ-788 (ET(B) receptor antagonist, 10(-7) mol/L) or both were recorded. In addition, the collateral responses to ET-1 with preincubation of n(omega)-nitro-L-arginine (NNA; 100 mol/L), indomethacin (INDO; 10 mol/L), or in combination were performed. ET-1 increased the perfusion pressure of collaterals and its effect was significantly suppressed by BQ-123 alone and BQ-123 plus BQ-788, but not BQ 788 alone (P <.05). Incubation with NNA, INDO, or both significantly enhanced the response of collaterals to ET-1 (P < .05). These results show that ET-1 produces a direct vasoconstrictive effect on the collateral vessels of portal hypertensive rats. This effect is mediated by ET(A,) but not ET(B), receptors. Both nitric oxide and prostaglandin modulate the collateral vascular response to ET-1 and may therefore participate in the development and maintenance of portal hypertension. PMID- 11283845 TI - Effects of blood volume restitution following a portal hypertensive-related bleeding in anesthetized cirrhotic rats. AB - The aim of this study was to investigate the influence of different strategies of blood volume restitution in the outcome of portal hypertension-related bleeding in anesthetized cirrhotic rats. Gastrointestinal hemorrhage was induced by sectioning a first order branch of the ileocolic vein in 38 cirrhotic rats (common bile duct ligation and occlusion). The subsequent hypovolemic shock was treated with no transfusion (n = 17), moderate transfusion (50% of expected blood loss, 5 mL, n = 11), and total transfusion (100% of expected blood loss, 10 mL, n = 10). At the end of the blood transfusion period (minute 15), mean arterial pressure (MAP) partially recovered in rats receiving moderate transfusion or no transfusion but decreased in the 10-mL transfusion group ( downward arrow 12 +/- 43%, P < .05 vs. no transfusion and 5 mL transfusion). After transfusion, groups given no or 5 mL transfusion remained hemodynamically stable. However, rats receiving 10 mL transfusion continued to deteriorate with persistent bleeding and progressive fall in MAP ( downward arrow 65 +/- 12%; P < .05 vs. no transfusion and 5 mL transfusion). Collected blood loss was significantly greater in the 10 mL group (20.0 +/- 1.5 g) than in groups given 5 mL (15.9 +/- 2.8 g; P < .05) or no transfusion (13.2 +/- 2.1 g; P < .05 vs. 10 mL and 5 mL transfusion). Survival in the no transfusion group was 47%. Rats given 5-mL transfusion had 64% survival. The worst survival was observed in the 10-mL transfusion group (0% survival; P < .05). We concluded that a transfusion policy aimed at completely replacing blood loss worsens the magnitude of bleeding and mortality from portal hypertensive-related bleeding in cirrhotic rats. On the contrary, moderate blood transfusion allowed hemodynamic stabilization and increased survival. PMID- 11283846 TI - Altered peripheral vascular responses to exogenous and endogenous endothelin-1 in patients with well-compensated cirrhosis. AB - Plasma endothelin concentrations are elevated in cirrhosis and correlate with disease severity. This study assessed forearm vascular responses to exogenous endothelin-1 (ET-1), and evaluated the contribution of endogenous ET-1 to the maintenance of basal peripheral vascular tone in patients with well-compensated cirrhosis (n = 11) and matched healthy controls (n = 8). Bilateral forearm blood flow (FBF) was measured at baseline and following unilateral, subsystemic, intrabrachial artery infusions of ET-1 (2 and 6 pmol/min); BQ-123, a selective ET(A) receptor antagonist (3 and 10 nmol/min); and BQ-788, a selective ET(B) receptor antagonist (0.3 and 1 nmol/min) using venous occlusion plethysmography. Baseline systemic hemodynamics and plasma ET-1 and big ET-1 concentrations were measured using electrical bioimpedance and radioimmunoassay, respectively. Patients and controls had similar baseline FBF, systemic hemodynamics, and plasma ET-1 and big ET-1 concentrations. In both groups, ET-1 and BQ-788 caused significant vasoconstriction (P < .001) and BQ-123 caused significant vasodilatation (P < .001). Compared with controls, cirrhotic patients had attenuated ET-1 responses (P < .001), augmented BQ-123 responses (P < .001), and similar BQ-788 responses (P = .62). Despite normal systemic hemodynamics and plasma ET-1 concentrations, forearm vascular responses to exogenous ET-1 are reduced in cirrhotic patients. The augmented vasodilatation to BQ-123 in cirrhotic patients is consistent with a compensated vasodilated state, and a greater contribution of ET-1 to the maintenance of basal vascular tone acting through the ET(A) receptor. PMID- 11283847 TI - Identification of differentially expressed genes in hepatocellular carcinoma with cDNA microarrays. AB - Genes expressed in hepatocellular carcinoma (HCC) were analyzed using cDNA microarrays to clarify gene abnormalities in HCC. mRNA was extracted from cancerous and noncancerous tissues of 10 patients with HCC, cDNA labeled with Cy5 and Cy3 fluorescence was prepared, and it was hybridized for each patient with a cDNA microarray consisting of 1,080 elements (930 unique genes). The mRNA expression rate of each element in HCC was evaluated using the level of mRNA expression in noncancerous tissue in each patient as a reference. The expression of 10 genes was enhanced 2 times or more in HCC cancerous tissue compared with noncancerous tissue in 5 or more of the 10 patients. In contrast, 9 genes were expressed at half the level or less in HCC cancerous tissue compared with noncancerous tissue. When hierarchical clustering was performed to identify genes related to clinical phenotypes of the patients, 22 genes showed changes associated with the degree of differentiation of HCC. Thirteen of these genes were transcriptional factors or tissue-specific expression proteins related to cell differentiation or development. Our present analysis clarified a number of genes that characterize HCC. This information based on examination of clinical samples is considered to be useful for clarification of the mechanism of hepatocarcinogenesis and the diagnosis and treatment of HCC. PMID- 11283848 TI - The vascular endothelial growth factor receptor KDR/Flk-1 is a major regulator of malignant ascites formation in the mouse hepatocellular carcinoma model. AB - The vascular endothelial growth factor-A (VEGF-A), also known as the vascular permeability factor (VPF), has been shown to play an important role in malignant ascites formation. The effects of VEGF-A are mediated through flt-1 and kinase insert domain-containing receptor/fetal liver kinase (KDR/Flk-1) receptors. It has been shown that KDR/Flk-1 is a predominant receptor in solid hepatocellular carcinoma (HCC) development, but the role of this receptor in hepatic ascites formation has not yet been elucidated. In this study, we examined the role of KDR/Flk-1 in the murine MH134 hepatic malignant ascites formation by means of VEGF-A- and KDR/Flk-1-specific neutralizing antibodies (VEGF-A nAb and KDR/Flk-1 nAb, respectively). The mean volume of ascites, number of tumor cells in ascites, and the peritoneal capillary permeability were significantly suppressed by VEGF-A nAb and KDR/Flk-1 nAb treatment. These inhibitory effects of KDR/Flk-1 nAb were more potent than those of VEGF-A nAb. The autophosphorylation of KDR/ Flk-1 in the peritoneal wall was almost completely abolished by KDR/ Flk-1 nAb, whereas a certain level of activation was still shown by VEGF-A nAb treatment. Another VEGF family, VEGF-C, which also binds KDR/Flk-1, was detected in the ascites. Furthermore, in the therapeutic experiment, although both VEGF-A nAb and KDR/Flk 1 nAb prolonged the survival rate of ascites-bearing mice, the latter showed a more significant impact on the survival of animals. These results suggest that KDR/Flk-1 is a major regulator of malignant hepatic ascites formation, and that in addition to VEGF-A, VEGF-C may also be involved in the malignant ascites formation via KDR/ Flk-1 activation. PMID- 11283849 TI - Sustained expression of naked plasmid DNA encoding hepatocyte growth factor in mice promotes liver and overall body growth. AB - To understand the physiological functions of exogenous hepatocyte growth factor (HGF) on normal adult animals, we delivered human HGF gene into mice by a hydrodynamics-based in vivo gene transfection approach using a naked plasmid vector. Systemic administration of naked plasmid containing HGF cDNA driven under cytomegalovirus promoter (pCMV-HGF) by rapid injection via the tail vein produced a remarkable level of human HGF protein in the circulation, beginning to appear at 4 hours and peaking at 12 hours following injection. Tissue distribution studies identified the liver as the organ with the highest level of transgene expression. Through weekly repeated injections of plasmid vector, we achieved sustained, long-term, high levels of exogenous HGF expression in mice for 8 weeks. Increases of more than 31% and 16% in liver and body weights were found, respectively, in the mice that received pCMV-HGF plasmid compared with that given the control vector for 8 weeks. Expression of exogenous HGF in vivo activated mitogen-activated protein kinases and induced proliferating cell nuclear antigen expression in normal adult liver and kidneys. These data suggest that systemic administration of naked plasmid vector is a convenient, safe, and highly efficient approach to introduce and maintain exogenous HGF gene expression in vivo in a controllable fashion. Our results also indicate that long-term expression of human HGF in mice markedly activates growth-related signal transduction events, promotes cell proliferation, and leads to liver and overall body growth in whole adult animals. PMID- 11283850 TI - Expression of cyclooxygenase-2 promotes the release of matrix metalloproteinase-2 and -9 in fetal rat hepatocytes. AB - Treatment of primary cultures of fetal hepatocytes with proinflammatory cytokines, lipopolysaccharide (LPS), and hepatocyte growth factor promoted the expression of cyclooxygenase-2 (COX-2) and the synthesis of high amounts of prostaglandins (PGs). Under these conditions, the active forms of the matrix metalloproteinases-2 and -9 (MMPs) were released to the extracellular medium. This process was inhibited when the synthesis of PGs was suppressed pharmacologically with COX-2 inhibitors. Addition to the cell cultures of PGE(2) promoted the release of MMPs through a mechanism that involved the expression of COX-2 and the synthesis of additional PGs. Kinetic analysis of the secretion of MMPs in response to LPS and PGE(2) showed a similar time course, with a lag period of 6 hours, which suggests that PGE(2) does not act directly on the mechanism of MMP processing and release. Inhibitors of protein kinase A, p38 MAP kinase, phosphatidylinositol-3-kinase, and nuclear factor kappaB (NF-kappaB) activation impaired the release of MMPs in response to PGE(2) challenge, indicating the involvement of multiple steps in the process. The ability of fetal hepatocytes to release MMPs in response to growth factors and inflammatory stimuli constitutes a model for the study of the extracellular matrix remodeling that accompanies most liver diseases. PMID- 11283851 TI - Tumor hepatocytes and basement membrane-Producing cells specifically express two different forms of the endostatin precursor, collagen XVIII, in human liver cancers. AB - Endostatin is an endogenous inhibitor of angiogenesis and tumor growth in mice, which may be generated by proteolytic cleavage of collagen XVIII. In normal tissues, 2 variants of the endostatin precursor, namely the SHORT and LONG forms, regulate tissue specificity. We analyzed 53 human liver biopsies (18 hepatocellular carcinomas, 16 metastases of colorectal cancer, 3 cholangiocarcinomas, and 16 controls) by RNA dot blots, double-labeling immunohistochemistry, and in situ hybridization, using common and variant specific probes. Tumor hepatocytes expressed the LONG form, whereas cholangiocarcinoma cells expressed the SHORT form, which was deposited in tumor basement membranes. Metastatic colorectal carcinoma cells did not express collagen XVIII. In the stromal compartment of primary and metastatic cancers, myofibroblasts and vascular endothelial cells expressed the SHORT form. Both basement membrane components, collagen IV and the SHORT collagen XVIII form, were codistributed and their mRNA levels strongly correlated (R =.75, P <.001). In addition, freshly isolated human hepatocytes expressed the LONG form and culture activated stellate cells the SHORT form. Moreover, the full-length LONG form is a plasma protein. Thus, the LONG form is a hepatocyte-specific variant, and the SHORT form is a major component of the tumor extracellular matrix in primary and metastatic liver cancers. In the clinical context, the global expression of the endogenous endostatin precursor, collagen XVIII, in liver cancer results from the combined expression profiles of tumor cells, stromal cells, and nontumor hepatocytes at the advancing edge of the tumor, particular to each type of cancer. PMID- 11283852 TI - CD45RC gammadelta T-cell infiltration is associated with immunologic unresponsiveness induced by prior donor-specific blood transfusion in rat hepatic allografts. AB - Little is known regarding the role of gammadelta(+) T cells in organ transplantation. We previously reported that immunologic unresponsiveness is induced by prior donor-specific blood transfusion (DST) in rat hepatic allografts. We investigated the phenotype and distribution of gammadelta(+) T cells in the hepatic allograft, spleen, and peripheral blood of recipient rats with immunologic unresponsiveness induced by DST. gammadelta(+) T cells were enumerated in allograft livers and spleens by immunostaining and in blood by flow cytometric analysis. The phenotype of gammadelta(+) T cells was determined using CD45RC isoforms derived from alternative mRNA splicing. The cytokine profile of CD45RC(+) and CD45RC(-) gammadelta(+) T cells was analyzed by reverse transcription polymerase chain reaction. The number of gammadelta(+) T cells in hepatic infiltrates in recipient rats pretreated with DST was significantly greater than in untreated animals. This correlated with significantly higher levels of gammadelta T cell receptor (TCR) mRNA in hepatic allografts of DST treated rats as compared with untreated animals. The gammadelta(+) T cell/alphabeta(+) T-cell ratio increased in hepatic infiltrates in DST-treated recipient rats but not in untreated animals. CD45RC(-)gammadelta(+) T cells were predominantly increased in DST-treated hepatic allografts compared with untreated allografts. Most of the intestinal intraepithelial T cells were CD45RC( )gammadelta(+). Interleukin (IL)-10 and IL-4 mRNA were detected more in CD45RC( )gammadelta(+) T cells than CD45RC(+)gammadelta(+) T cells. CD45RC( )gammadelta(+) T cells infiltrating liver allografts produce Th2-type cytokines and are associated with immunologic unresponsiveness induced by DST. PMID- 11283853 TI - A critical role of T-cell receptor gamma/delta cells in antibacterial protection in mice early in life. AB - Although it is generally assumed that T-cell receptor (TCR) gamma/delta cells participate in protection against intracellular microbial pathogens, their impact remains controversial. In our study, young (14-day-old) mice lacking TCRgamma/delta cells were far more susceptible to Listeria monocytogenes than wild-type (WT) mice of the same age. The number of interferon gamma (IFN-gamma) producers responsible for antilisterial resistance was significantly higher among natural killer (NK)1(+) TCRgamma/delta cells than among NK1(-) TCRgamma/delta cells. Endogenous IFN-gamma neutralization increased susceptibility of young WT mice to L. monocytogenes infection. Liver was a major residence of peripheral NK1(+) TCRgamma/delta cells, whereas NK1(-) TCR gamma/delta cells were broadly distributed in various lymphoid organs. Numbers of both NK1(+) and NK1(-) TCRgamma/delta cells increased in the liver of WT mice prior to TCRalpha/beta cells and represented a substantial population in early life (14 days after birth). Virtually all NK1(+) TCRgamma/delta cells expressed activation markers, whereas substantial numbers of NK1(-) TCRgamma/delta cells showed a naive phenotype. We conclude that TCRgamma/delta cells play a critical role in protection against L. monocytogenes in the early life of mice, probably because their TCRalpha/beta cell compartment is not fully competent. For this antibacterial function, we assign NK1(+) TCRgamma/delta cells a more important role than their NK1(-) cognates. PMID- 11283854 TI - Endothelin B receptor-mediated protection against anoxia-reoxygenation injury in perfused rat liver: nitric oxide-dependent and -independent mechanisms. AB - This study aimed to investigate the roles of endothelin (ET) receptors in biliary dysfunction and cell injury in postischemic livers. Rat livers perfused with oxygenated Krebs-Henseleit solution were exposed to reoxygenation following 20 minute hypoxia. The anoxic perfusion decreased bile output and reduced cyclic guanosine monophosphate (cGMP) contents, an index of nitric oxide (NO) generation. Upon reoxygenation, the decreased bile was not fully recovered, and the resistance increased biphasically: an early transient spike accompanied by an elevated release of ET-1 and a rise accompanied by a cGMP elevation in the later period. The initial vasoconstriction appeared to be mediated by both ET(A) and ET(B) receptors, as judged by inhibitory effects of their antagonists, BQ-485 and BQ-788, respectively, while the late elevation of the resistance was not attenuated by these reagents, but rather enhanced by the ET(B) blockade. The BQ 788 treatment attenuated the reoxygenation-induced cGMP elevation and induced bile acid-dependent choleresis. However, such a change upon the ET(B) blockade coincided with dissociation of a recovery of phospholipids and aggravation of cell injury. The BQ-788-elicited deterioration of reoxygenation-elicited changes was attenuated by NO supplement with S-nitroso-N-acetyl penicillamine. N(omega) Nitro-L-arginine methyl ester, an NO synthase inhibitor, mimicked biliary changes elicited by the ET(B) blockade but without causing notable cell injury. Under these circumstances, coadministration of clotrimazole, an inhibitor of cytochrome P450 mono-oxygenases, elicited the injury comparable with that observed under the ET(B) blockade. These results suggest that ET(B)-mediated signaling limits excessive bile acid excretion and plays a protective role against reoxygenation injury through mechanisms involving both NO-dependent and -independent processes. PMID- 11283855 TI - Subcellular site of superoxide dismutase expression differentially controls AP-1 activity and injury in mouse liver following ischemia/reperfusion. AB - Acute damage following ischemia and reperfusion (I/R) in the liver is in part caused by the generation of reactive oxygen species, such as superoxides, during the reperfusion event. Gene therapy directed at attenuating mitochondrial superoxide production following warm I/R injury in the liver has demonstrated great promise in reducing acute hepatocellular damage. In the present study, we have compared the therapeutic effects of ectopic expression of mitochondrial (MnSOD) and cytoplasmic (Cu/ZnSOD) superoxide dismutase using recombinant adenoviral vectors for reducing I/R damage in the liver. Consistent with previous observations, recombinant adenoviral delivery of MnSOD to the liver significantly attenuated both acute liver damage and AP-1 activation following I/R injury to the livers of mice. However, ectopic expression of Cu/ZnSOD diminished neither I/R-induced elevations in serum alanine transaminase (ALT) nor AP-1 activation. Interestingly, baseline activation of AP-1 before I/R-induced injury was seen in livers infected with recombinant Ad.Cu/ZnSOD, but not Ad.MnSOD or Ad.LacZ, vectors. The level of Cu/ZnSOD-induced AP-1 activation was significantly reduced by ablation of Kupffer cells or by coexpression of catalase, suggesting that increased H(2)O(2) production facilitated by Cu/ZnSOD in hepatocytes and/or Kupffer cells may be responsible for AP-1 activation. In vitro reconstitution studies using hepatocyte and macrophage cell lines demonstrated that Cu/ZnSOD overexpression induces AP-1 in both cell types, and that secretion of a Cu/ZnSOD induced macrophage factor is capable of elevating AP-1 in hepatocytes. In summary, our findings demonstrate that subcellular sites of superoxide production in the liver can differentially affect the outcome of I/R injury in the liver and selectively influence AP-1 activation. PMID- 11283856 TI - Normal liver regeneration in p50/nuclear factor kappaB1 knockout mice. AB - Nuclear factor kappaB (NF-kappaB) is rapidly activated during liver regeneration following partial hepatectomy or carbon tetrachloride (CCl(4))-mediated liver injury and is felt to be important in the antiapoptotic and regenerative responses. After partial hepatectomy, livers of mice deficient in the p50 subunit of NF-kappaB (p50(-/-)) showed a loss of NF-kappaB and decreased STAT3 transcription factor DNA binding activities. However, nuclear levels of the NF kappaB p65 subunit were increased and peaked earlier in p50(-/-) livers. Both messenger RNA and cytoplasmic protein levels of the NF-kappaB inhibitor IkappaBalpha were lower in p50(-/-) livers, potentially accounting for the increase in p65 protein. Small effects on gene expression posthepatectomy were observed in p50(-/-) livers, but no effects were seen on hepatocyte DNA synthetic or mitotic responses, serum enzyme levels, or overall liver mass restoration. After CCl(4) treatment, hepatocyte DNA synthesis and mitosis and serum enzyme levels were similar in p50(-/-) and p50(+/+) mice, and histologic analysis indicated a slight decrease in overall damage in p50(-/-) livers. After injection of Fas antibody, p50(-/-) livers showed an earlier onset of nuclear changes consistent with apoptosis. These data indicate that absence of p50 affects certain protein and gene activation pathways following partial hepatectomy, CCl(4), and Fas treatment but does not impair overall liver regeneration. Interleukin 6 (IL-6) levels were reduced but still adequate to support regeneration. We hypothesize that increased levels of the NF-kappaB p65 subunit in p50(-/-) livers may provide compensation for the absence of p50, thereby allowing normal liver regeneration and repair following liver injury. PMID- 11283857 TI - Effect of nitric oxide on shock-induced hepatic heme oxygenase-1 expression in the rat. AB - Recent evidence suggests that the hepatic expression of heme oxygenase-1 (HO-1) may preserve hepatocellular integrity after hemorrhagic shock and resuscitation (HR). Because nitric oxide (NO) has been shown to modulate HO-1 expression in cultured cells in vitro, we determined its potential role in the regulation of HO 1 expression after HR in the rat liver in vivo. HO-1 mRNA and protein were highly induced and HO enzyme activity was higher after HR when compared with time matched sham controls. Administration of the NO donor, molsidomine (MOL) (3 mg. kg(-1)), during resuscitation attenuated the accumulation of HO-1 mRNA and protein and the rise in HO activity. In addition, MOL prevented the shock-induced increase in DNA binding activity of the transcription factor, activator protein-1 (AP-1), but did not alter the activity of nuclear factor-erythroid 2 related factor (Nrf-2), nuclear transcription factor-kappaB (NF-kappaB), and hypoxia inducible factor-1 (HIF-1). The suppressing action of MOL was not confined to HO 1, because the hepatic expression of the 70-kd major heat shock protein (HSP) in response to HR was also diminished. Moreover, MOL prevented the HR-induced increase in the serum activity of alanine transaminase (ALT) and alpha glutathione-S-transferase (alpha-GST) that could otherwise be observed after HR. In contrast, the NO synthase inhibitor, N(omega)-nitro-L-arginine methyl ester (L NAME) (1 mg.kg(-1)), had either no or only minor effects on the primary experimental endpoints. These findings would be consistent with a reduction of shock-induced liver damage by exogenous NO, which in turn prevents the subsequent activation of injury-sensitive transcription factors, thus attenuating the expression of stress-inducible proteins such as HO-1. PMID- 11283858 TI - Plasma membrane Ca2+ release-activated Ca2+ channels with a high selectivity for Ca2+ identified by patch-clamp recording in rat liver cells. AB - Repetitive waves of increased cytoplasmic Ca2+ concentration play a central role in the process by which hormones regulate liver function. Maintenance of these Ca2+ waves requires Ca2+ inflow through store-operated Ca2+ channels. The properties and mechanism(s) of activation of these channels are not well understood. Store-operated Ca2+ channels (SOCs) in the H4-IIE rat liver cell line were studied by whole-cell patch clamping. Depletion of Ca2+ in intracellular stores by intracellular perfusion with either inositol 1,4,5-trisphosphate (InsP(3)) or thapsigargin in the presence of 10 mmol/L ethylene glycol-bis(beta aminoethyl ether)-N,N-tetraacetic acid (EGTA), or with 10 mmol/L EGTA alone, activated an inward current that reversed at a membrane potential above +40 mV. In physiologic extracellular medium, this inward current was carried exclusively by Ca2+ and was blocked by a variety of di- and trivalent cations. In the absence of extracellular Ca2+ and Mg2+, the inward current was carried by monovalent cations. This current was 10 to 30 times larger than that observed in the presence of extracellular Ca2+, and permitted the detection of single-channel events that corresponded to a single-channel conductance of about 40 pS. Both the Ca2+ and Na+ inward currents were blocked by the calmodulin antagonist, N-(6 amino hexyl)-5-chloro-1-naphthalenesulphonamide (W7), but not by calmidazolium or calmodulin-dependent protein kinase II fragment 290-309. It is concluded that liver cells possess plasma membrane Ca2+ channels that have a high selectivity for Ca2+, are activated by a decrease in the concentration of Ca2+ in intracellular stores through a mechanism that is unlikely to involve calmodulin, and are involved in re-filling intracellular Ca2+ stores during Ca2+ signaling. PMID- 11283859 TI - Resistance to fulminant hepatitis and carcinogenesis conferred by overexpression of retinoblastoma protein in mouse liver. AB - Previously, retinoblastoma (Rb) transgenic mice were produced under the control of the Rb gene promoter and showed dwarf characteristics. Here, we created transgenic mice, in which the human Rb gene was controlled by the hepatocyte nuclear factor-1 gene promoter/enhancer and was expressed primarily in the liver. The liver of these novel transgenic mice was normally developed. Intriguingly, these mice showed resistance to fulminant hepatitis induced by anti-Fas antibody as well as resistance to chemical carcinogenesis in the liver. These results show that the Rb protein acts as an anti-apoptotic and anti-oncogenic agent in vivo. Our novel construct may be useful as a gene cassette in gene therapy for prevention of fulminant hepatitis and hepatoma. PMID- 11283860 TI - In vitro and in vivo activation of rat hepatic stellate cells results in de novo expression of L-type voltage-operated calcium channels. AB - Following chronic liver injury, hepatic stellate cells (HSCs) transdifferentiate into myofibroblast-like cells, which develop contractile properties and contribute to increased resistance to blood flow. We investigated whether this phenotypic activation includes changes in the expression of L-type voltage operated Ca2+ channels (VOCC), which mediate Ca2+ influx and regulate cell contraction in vascular cell types. Rat HSCs were studied in the quiescent phenotype and after their activation in vitro (cultured on plastic for 14 days) and in vivo (isolated from rats with CCl(4)-induced cirrhosis). Patch-clamp studies showed Ca2+ currents through L-type VOCC in HSCs activated both in vitro and in vivo, whereas no currents were detected in quiescent HSCs. Moreover, binding studies with (3)H-isradipine, a specific L-type VOCC antagonist, showed a large number of binding sites in activated HSCs, while no specific binding was found in quiescent HSCs. Finally, messenger RNA (mRNA) encoding L-type VOCC was not detected in quiescent HSCs as assessed by reverse transcription-polymerase chain reaction (RT-PCR) and Northern blot analysis, whereas it was present in activated HSCs. Stimulation of L-type VOCC with KCl resulted in a marked increase in [Ca2+](i) followed by cell contraction in HSCs activated both in vitro and in vivo, whereas no effects were observed in quiescent HSCs. We conclude that the activation of HSCs is associated with up-regulation of L-type VOCC that mediate Ca2+ influx and cell contraction. These results may be relevant to the pathogenesis of portal hypertension. PMID- 11283861 TI - Lamivudine treatment can overcome cytotoxic T-cell hyporesponsiveness in chronic hepatitis B: new perspectives for immune therapy. AB - The hepatitis B virus (HBV) cytotoxic T lymphocyte (CTL) response in patients with chronic HBV infection is generally weak or totally undetectable. This inability to mount protective CTL responses is believed to be a crucial determinant of viral persistence, and its correction represents an important objective of immune therapies for chronic hepatitis B. However, amplification of CTL responses in vivo may be ineffective if HBV-specific CD8 cells are either absent or nonresponsive to exogenous stimulation. In this study, we asked whether antiviral treatments able to inhibit viral replication and to reduce viral and antigen load can successfully reconstitute CTL responses creating the appropriate conditions for their therapeutic stimulation. For this purpose, the HBV-specific CTL response before and during lamivudine therapy was studied longitudinally in 6 HLA-A2-positive patients with HBeAg+ chronic hepatitis B. Both HBV-specific cytotoxic T cell activity measured by chromium release assay on peptide stimulation in vitro and CD8+ T cell frequency measured ex vivo by HLA-A2/peptide tetramer staining were significantly augmented by lamivudine therapy. This enhancement followed the reconstitution of CD4 reactivity and the decline of viral load induced by therapy. Our study shows that lamivudine treatment in chronic hepatitis B can restore CTL reactivity, making CTL susceptible to exogenous stimulation. This effect may enhance the probability that T cell-based immune therapies delivered after lamivudine treatment can successfully reconstitute a protective CTL response able to cure chronic HBV infection. PMID- 11283862 TI - Hepatitis B infection in patients with acute liver failure in the United States. AB - Occult hepatitis B virus (HBV) infection has been reported in 30% to 50% of patients with acute liver failure (ALF) in small case series. The aim of this study was to determine the prevalence of occult HBV infection in a large series of ALF patients in the United States and the prevalence of precore and core promoter variants in patients with ALF caused by hepatitis B. Sera from patients in the US ALF study and liver, when available, were tested using nested polymerase chain reaction (PCR) with primers in the HBV S and precore regions. PCR-positive samples were sequenced. Sera and/or liver from 139 patients (39 males, 100 females; mean age, 42 years) enrolled between January 1998 and December 1999 were studied. Twelve patients were diagnosed with hepatitis B, 1 with hepatitis B+C+D coinfection, and 22 had indeterminate etiology. HBV DNA was detected in the sera of 9 (6%) patients; all 9 had ALF caused by hepatitis B. HBV genotypes A, B, C, and D were present in 4, 3, 1, and 1 patients, respectively. Seven of these 9 patients had precore and/or core promoter variants. Liver from 19 patients were examined. HBV DNA was detected in the liver of 3 patients with ALF caused by hepatitis B, but in none of the remaining 16 patients with non-B ALF. Contrary to earlier reports, occult HBV infection was not present in this large series of ALF patients in the United States. HBV precore and/or core promoter variants were common among US patients with ALF caused by hepatitis B. PMID- 11283863 TI - Detection of hepatitis C virus in the bile and bile duct epithelial cells of hepatitis C virus-infected patients. AB - The pathobiology of hepatitis C virus (HCV) in the biliary system has not been clarified yet, although bile duct damage is a histological finding characteristic of chronic hepatitis C. In this study, we examined whether HCV infects bile ducts and is released into the bile. Twelve patients positive for serum HCV antibody were examined in this study, and eight were seropositive for HCV RNA by polymerase chain reaction (PCR). For those who underwent abdominal surgery, the bile was aspirated from the gall bladders by fine needle puncture. Five underwent wedge liver biopsy. Series of saline-diluted bile were assayed for HCV RNA by PCR to determine the HCV RNA titers. To examine HCV expression in the biliary system, the liver specimens were immunostained using monoclonal antibodies to the HCV proteins. HCV RNA was detected in the bile of 5 patients with high serum HCV RNA load (> or = 2.5 Meq/mL). Comparison of viral loads between serum and bile revealed that the HCV RNA level in the bile was as high as that in serum. Furthermore, immunohistological study showed that bile duct epithelial cells were infected with HCV. In contrast, HCV was not found in either the bile or bile duct of patients seronegative for HCV RNA or with low serum HCV load (< or = 1.1 Meq/mL). These findings suggest that the biliary system is involved in the pathobiology of HCV and that the bile is as highly infectious as the serum. PMID- 11283864 TI - Repopulation of mouse liver with human hepatocytes and in vivo infection with hepatitis B virus. AB - Mice containing livers repopulated with human hepatocytes would provide excellent in vivo models for studies on human liver diseases and hepatotropic viruses, for which no permissive cell lines exist. Here, we report partial repopulation of the liver of immunodeficient urokinase-type plasminogen activator (uPA)/recombinant activation gene-2 (RAG-2) mice with normal human hepatocytes isolated from the adult liver. In the transplanted mice, the production of human albumin was demonstrated, indicating that human hepatocytes remained functional in the mouse liver for at least 2 months after transplantation. Inoculation of transplanted mice with human hepatitis B virus (HBV) led to the establishment of productive HBV infection. According to human-specific genomic DNA analysis and immunostaining of cryostat liver sections, human hepatocytes were estimated to constitute up to 15% of the uPA/RAG-2 mouse liver. This is proof that normal human hepatocytes can integrate into the mouse hepatic parenchyma, undergo multiple cell divisions, and remain permissive for a human hepatotropic virus in a xenogenic liver. This system will provide new opportunities for studies on etiology and therapy of viral and nonviral human liver diseases, as well as on hepatocyte biology and hepatocellular transplantation. PMID- 11283865 TI - A randomized trial of amantadine and interferon versus interferon alone as initial treatment for chronic hepatitis C. AB - The aim of this study was to compare, in an open-label study, the efficacy and safety of a combination of interferon (IFN) and amantadine (AMA) with that of IFN alone in previously untreated patients with chronic hepatitis C. A total of 200 patients were randomized to 6 MU of IFN-alpha2a 3 times per week, with 200 mg of AMA daily (n = 99) or to an identical dose of interferon alpha2a (n = 101). Patients were treated for 12 months and observed for 6 months' posttreatment. At the completion of treatment, 28.7% of patients in the monotherapy group and 45.5% in the combination group had a virologic response (P =.014). At 6 months' posttreatment, a sustained virologic response was observed in 16.8% (95% CI: 9 23) of patients with IFN alone versus 29.3% (95% CI: 19-37) of patients who were treated with combination therapy (P =.036). In each of the 2 treatments, genotype was the only predictive parameter for a sustained response. At the logistic regression analysis, therapy and genotype were the only 2 parameters with an independent predictive value. In the combination group, at examination of month 3, hepatitis C virus (HCV)-RNA status had a 97.6% (95% CI: 93-102) positive predictive value and a 50% (95% CI: 37-63) negative predictive value for a sustained virologic clearance. A substantial proportion of naive patients with chronic hepatitis C have an end-of-treatment and end-of-follow-up virologic and biochemical response to a combination of IFN and AMA. This new treatment appears safe and well tolerated. PMID- 11283866 TI - Autoimmune overlap syndromes. PMID- 11283867 TI - Primary prevention of variceal bleeding. What's new? PMID- 11283868 TI - Mouse liver goes human: a new tool in experimental hepatology. PMID- 11283869 TI - Ca2+ and rho signaling pathways: two paths to hepatic stellate cell contraction. PMID- 11283870 TI - Drug-induced liver injury: mechanisms and test systems. PMID- 11283871 TI - What hides behind an intention-to-treat analysis? PMID- 11283873 TI - Neurodevelopmental outcome after congenital diaphragmatic hernia: Extracorporeal membrane oxygenation before and after surgery. AB - BACKGROUND/PURPOSE: Extracorporeal membrane oxygenation (ECMO) as a treatment of last resort for neonates with persistent pulmonary hypertension of the newborn (PPHN) caused by congenital diaphragmatic hernia (CDH) may be used for preoperative stabilization or postoperative rescue. The aim of this study was to examine the acute and long-term morbidity associated with pre- and postoperative ECMO. METHODS: Neonates born with CDH and needing ECMO were classified into 2 groups. Group 1 consisted of neonates placed on ECMO after CDH surgery. Patients in group 2 underwent preoperative ECMO stabilization. Medical records after birth were evaluated. Growth, neuromotor and cognitive development, hearing, and behavior were evaluated. Student t test and chi(2) were used to determine statistical significance between groups. RESULTS: Subjects in group 2 had significantly more days on ECMO and loop diuretics. Alkalosis was induced for a longer duration in group 2. At follow-up 3 to 9 years later, no differences were found between the 2 groups in growth parameters, neuromotor outcome, or behavior. However, in group 1, 2 of 9 children had significant hearing impairment necessitating amplification compared with 6 of 6 subjects in group 2. CONCLUSIONS: Neonates with CDH first stabilized on ECMO (group 2) had a higher incidence of hearing loss compared with those needing ECMO postrepair (group 1). The etiology of this finding is not clear. This may be secondary to the prolonged period of hyperventilation or general intensive care that is part of the protocol for neonates who are electively stabilized on ECMO preoperatively. J Pediatr Surg 36:539-544. PMID- 11283874 TI - Operative management for sacrococcygeal teratoma diagnosed in utero. AB - BACKGROUND/PURPOSE: Sacrococcygeal teratomas (SCT) diagnosed in utero have been reported to be large and associated with high perinatal mortality rate. However, operative management including timing of operation after birth, combined abdominal approach for devascularization, and the position of the patients during resection is not well established. METHODS: A retrospective review of 14 patients with SCT between 1978 and 1999 was performed. To prevent massive bleeding during surgery, the authors used an abdominoperineal resection in the supine position after devascularization. The patients' clinical and sonographic characteristics, prenatal outcome, operative management, and postnatal outcomes were examined. RESULTS: One fetus died in utero. Two patients died within a week, but no late death and no malignant degeneration were noted. A staged operation with devascularization was performed in 2 patients, and 1 death occurred. Surgical management was analyzed between survivors without massive bleeding at surgery (n = 9) and others (n = 4). A significant difference was observed in the subgroup of tumor resection with devascularization or supine position and that of early resection with devascularization or supine position. CONCLUSIONS: Early resection using the abdominoperineal approach supported by close antenatal sonography may be preferable for a favorable outcome. Resection in the supine position after devascularization may have advantages of respiratory management, cardiac resuscitation, and bleeding prevention. J Pediatr Surg 36:545-548. PMID- 11283875 TI - Adrenocortical tumors in children. AB - BACKGROUND/PURPOSE: Etiopathogenesis and management of pediatric adrenocortical tumors (ACTs) is still obscure because of the limited number of cases. The aim of this study is to present a clear picture of the entire spectrum of pediatric ACTs by reviewing one of the largest noncollected pediatric series treated in a single medical center. METHODS: Records of children treated for ACTs in our unit between 1970 and 1999, inclusive, were reviewed. Information recorded for each patient included age, sex, clinical characteristics, diagnostic methods, stage of disease, treatment, pathologic findings, and outcome. The patients were subdivided into 2 groups: group I, patients with adrenocortical carcinoma (ACC) and group II, patients with adrenocortical adenoma (ACA). These groups were analyzed with regard to parameters mentioned above. RESULTS: There were 30 children treated for ACTs in the study period with a mean age of 6.7 +/- 4.2 years (range, 2.5 to 13 years). Of these, 20 had ACC, and 10 had ACA. The tumors were right sided in 22 patients, left sided in 6 and bilateral in 2. Analysis of each group with regard to age and site of tumor showed no significant difference. Endocrine dysfunction was noted in 83% of the patients and virilization was the most common presentation followed by Cushing's syndrome. The most striking difference between 2 groups was the prepondarance of virilization in group II and Cushing's syndrome in group I. In the latter, 14 patients presented with palpable abdominal mass and 3 patients with distant metastases. The mean time from initial symptoms to diagnosis was 8.1 +/- 0.2 months, and this interval was similar in 2 groups, in functional and nonfunctional tumors, and in both sexes. Ultrasound scan, computerized tomography, magnetic resonance imaging, intravenous pyelography, and angiography were used for the diagnosis. All patients with ACA had localized disease, whereas 80% of the patients with ACC had regional or metastatic disease. Total excision was done in all patients with ACA, but only in 13 patients with ACCs. Of the latter, 2 patients underwent ipsilateral nephrectomy, and 1 patient had right hepatic lobectomy plus nephrectomy. Adjuvant chemotherapy consisting of mitotane (n = 12), mitotane plus cisplatin and etoposide (n = 2) was commenced. Seven patients with ACC had distant metastases postoperatively. The presence of regional disease at presentation was associated with a significantly shorter disease-free interval. All patients presenting with nonfunctional ACC (n = 4), functional ACC that have been totally resected (n = 4), and partially resected (n = 3) died of disease within the first 2.5 years after diagnosis. There was no significant difference between the functional and nonfunctional ACCs with regard to survival rate. All patients who had distant metastases postoperatively and who had partial excision died. Of the surviving 9 patients with ACC, there are 6 known long-term survivors who are still alive. CONCLUSIONS: ACAs are treated by total excision satisfactorily without any complication. For the time being, the most important aspect of therapy for ACCs is early diagnosis and total excision. Partial excision and advanced-stage disease are the major determinants of poor outcome. None of the clinical, laboratory, or pathologic features are reliable predictors for recurrence and discrimination of malignancy in ACTs. Because of the steadily increasing incidence of precancerous genetic syndromes of adrenal glands and poor prognosis of ACCs, childhood patients of endocrine disorders should receive a detailed and vigorous diagnostic evaluation and appropriate treatment as given to adults. Patients with ACTs should be entered into multi-institutional trials to adequately assess effective chemotherapy and radiotherapy protocols and molecular mechanisms of oncogenesis. J Pediatr Surg 36:549-554. PMID- 11283876 TI - Extended hiatoplasty: early experience with a simple technique to increase the intraabdominal esophageal length in complicated gastroesophageal reflux. AB - PURPOSE: The aim of this paper is to describe the experience of the Division of Pediatric Surgery of State University of Campinas Medical School with a simple technique of extended hiatoplasty to achieve intraabdominal placement of the distal esophagus in difficult situations. METHODS: From April 1997 to November 1999, 7 patients who had either complicated or recurrent gastroesophageal reflux (GER) underwent open (2 patients) or laparoscopic (5 patients) correction of GER that included an extended hiatoplasty. All had undergone previous unsuccessful clinical or surgical treatment. To investigate the severity of the disease, diagnostic endoscopy or barium swallow were performed before surgery. Postoperatively, these children underwent clinical evaluation or any additional diagnostic procedure deemed necessary. RESULTS: Using the extended hiatoplasty, a good length of intraabdominal esophagus could be achieved in every patient. No complications resulted from the procedure itself. There was a late instance of paraesophageal hernia with recurrence of GER attributable to disruption of the hiatoplasty, which was rerepaired through a laparoscopic approach. Symptomatic improvement was observed in all but the patient with caustic stricture. CONCLUSIONS: Extended hiatoplasty is a simple maneuver that may represent a good option to increase the length of intraabdominal esophagus in patients with a short esophagus secondary to severe GER disease, being associated with a high success rate and low morbidity. J Pediatr Surg 36:555-558. PMID- 11283877 TI - Risk factors for adverse outcome of newborns with gastroschisis in a Brazilian hospital. AB - BACKGROUND/PURPOSE: The aim of this study was to determine the frequency of postoperative death and to identify factors associated with adverse prognosis in cases of gastroschisis managed in a tertiary hospital of Brazil. METHODS: A retrospective transverse study was conducted including all cases of gastroschisis managed at Instituto Materno-Infantil De Pernambuco (IMIP), Recife, Brazil, between January 1995 and December 1999 (n = 31). Prevalence risk (PR) was determined for several prenatal, intraoperative, and postoperative factors. RESULTS: Overall mortality rate was 52% (16 cases), and sepsis was the main cause of death (93.8%). Prenatal diagnosis reduced about 70% the risk of death. Preterm and low birth weight babies had about 3 times increase in the risk of death. Risk of death was increased twice among outborn babies, but there was no association with delivery route. Increase in risk of neonatal death was related to these other factors: birth-to-admission interval longer than 2 hours and birth-to surgery interval longer than 4 hours. Prevalence risk also was greater with staged silo repair, poor clinical conditions before surgery, and when mechanical ventilation was needed. CONCLUSIONS: A high mortality rate was associated with absence of prenatal diagnosis, prematurity, low birth weight, delivery outside the tertiary center, and delayed surgery, worsening clinical conditions that preclude primary closure and increases need of mechanical ventilation. J Pediatr Surg 36:559-564. PMID- 11283878 TI - Absent peritoneal fluid on screening trauma ultrasonography in children: a prospective comparison with computed tomography. AB - BACKGROUND: Although the accuracy of focused abdominal sonography for trauma (FAST) in adults has been demonstrated, results of this technique in children have been conflicting with few comparisons against computed tomography (CT), the imaging gold standard. METHODS: A total of 160 hemodynamically stable pediatric trauma victims referred for abdominal CT initially underwent rapid screening sonography looking for free fluid. Both studies were interpreted in blinded fashion. RESULTS: Forty-four of the 160 patients had an intraabdominal injury on CT, 24 (55%) of which had normal screening sonography. Fifteen of the 44 (34%) had no free fluid on either modality. Accuracy of sonography compared with CT was 76% with a negative predictive value 81%. CONCLUSIONS: Sonography for free fluid alone is not reliable to exclude blunt intraabdominal injury in hemodynamically stable children given the considerable percentage of injured patients without free fluid. J Pediatr Surg 36:565-569. PMID- 11283879 TI - GLP-2alpha accelerates recovery of mucosal absorptive function after intestinal ischemia/reperfusion. AB - BACKGROUND/PURPOSE: The authors' previous laboratory results have shown that rats treated for 3 days with intravenous GLP-2alpha, a synthetic protease-resistant form of glucagonlike peptide-2, showed increased mucosal mass and absorptive function when compared with saline-treated controls after intestinal ischemia/reperfusion (I/R). This study was designed to explore the temporal relationship between injury that occurs secondary to intestinal I/R and recovery of mucosal absorptive function with and without GLP-2alpha treatment. METHODS: Each of 18 male Sprague-Dawley rats (300 to 333 g) was subjected to superior mesenteric artery occlusion for 30 minutes, during which time a jugular venous catheter was placed and connected to a subcutaneous infusion pump. Rats were divided into 4 groups based on the type and duration of infusion as follows: group 1, systemic saline at 1 microL/h for 24 hours (n = 5); group 2, systemic GLP-2alpha at 100 microg/kg/d for 24 hours (n = 5); group 3, systemic saline at 1 microL/h for 72 hours (n = 4); and group 4, systemic GLP-2alpha at 100 microg/kg/d for 72 hours (n = 4). Immediately after the infusions, (14)C galactose and (14)C-glycine absorption was measured using an in vivo, closed recirculation technique and expressed as micromoles per square centimeter intestine +/- SEM. Statistical analysis was performed using analysis of variance. RESULTS: Twenty-four hours after intestinal I/R injury, there was a decrease in substrate absorption but no significant difference between the saline and GLP 2alpha-treated groups (galactose absorption, 1.13 +/- 0.09 in group 1 v 1.35 +/- 0.11 in group 2, P =.15; glycine absorption, 1.18 +/- 0.13 in group 1 v 1.34 +/- 0.15 in group 2, P =.36). However, after the 72-hour infusion, absorption of galactose and glycine was significantly increased in the rats receiving GLP 2alpha as compared with the saline-infused control group (galactose absorption, 1.24 +/- 0.13 in group 3 v 1.88 +/- 0.10 in group 4, P <.01; glycine absorption, 1.64 +/- 0.07 in group 3 v 2.05 +/- 0.08 in group 4, P <.05). Compared with previously determined levels of absorption in normal, uninjured rat intestine (1.50 +/- 0.12 micromol/cm(2) for galactose and 1.85 +/- 0.17 micromol/cm(2) for glycine), after I/R a 72-hour infusion of GLP-2alpha increased galactose absorption 26% (P <.05) and glycine absorption 11% (P =.29) beyond baseline. CONCLUSIONS: When initiated at the time of intestinal I/R, a continuous infusion of GLP-2alpha accelerated recovery of mucosal absorptive function in rats. Remarkably, carbohydrate absorption at 72 hours was increased to a level significantly greater than that in normal, uninjured rat intestine. J Pediatr Surg 36:570-572. PMID- 11283880 TI - Ultrastructural deficiency in autonomic innervation in cremasteric muscle of boys with undescended testis. AB - BACKGROUND/PURPOSE: The cremaster muscles (CM) associated with undescended testis reveal neurogenic alterations that mainly affect type 2 fibers. The ultrastructure of CM has been evaluated to define if further evidence to explain the alterations could be identified. METHODS: CM of 8 boys with inguinal hernia and 8 boys with undescended testis at similar ages were biopsied. Samples were processed for electron microscopic evaluations. Semithin and thin sections were examined under an electron microscope. RESULTS: The CM associated with inguinal hernia showed normal ultrastructure. However, some alterations were encountered in CM associated with undescended testis. Unmyelinated fibers were diminished in number, and myelinated fibers were outnumbering the unmyelinated fibers. Marked disorientation of myofibers, redundant sarcolemma, empty sleeves of basal lamina, disarray of myofibrils, densely packed myofilaments, Z disk streaming, dilated sarcoplasmic reticulum, and dense-irregularly shaped mitochondria were repeatedly encountered. Satellite cells appeared inactive. Most of the fibers were contracted. CONCLUSIONS: The decrease in number of unmyelinated fibers appears to represent a decrease in autonomic nerve fibers. The alterations within muscle fibers may reflect a deficiency in autonomic innervation. Autonomic nervous system is highly responsive to circulating androgens. Factors decreasing the vulnerability of autonomic nervous system against androgenic effects may result in a CM with neurogenic alterations, thus inhibiting testicular descent. J Pediatr Surg 36:573-578. PMID- 11283881 TI - Insertion of an implantable venous access device in the groin using inferior epigastric vein. AB - BACKGROUND: Patients with cystic fibrosis (CF) need reliable venous access that can be provided by implantable venous access devices (IVAD). Such IVADs usually are placed in the upper part of the body, but placing them in this area has 3 disadvantages: a suitable vein may not be available, the portal may be conspicuous, and there may be interference with chest physiotherapy. Positioning the IVAD in the groin by using the inferior epigastric vein (IEV) is an answer to these problems. METHODS: This is a review of 29 patients from a single surgeon's practice from 1984 to 1999. A groin incision was used to implant the IVAD and to introduce the catheter via the IEV to the inferior vena cava. RESULTS: A total of 33 IVAD were inserted in 29 patients (27 with CF). The average age at first operation was 12.4 years. Infection was seen in 5. Venous thrombosis was not seen in any patient. The average longevity of IVAD is 18.5 months. Total experience is 87 IVAD years. CONCLUSION: This is a useful route of first choice for CF patients, with an overall complication rate comparable with techniques in which IVADs are placed in the upper half of body. J Pediatr Surg 36:579-581. PMID- 11283883 TI - Intestinal permeability in newborns with necrotizing enterocolitis and controls: Does the sugar absorption test provide guidelines for the time to (re-)introduce enteral nutrition? AB - BACKGROUND: In necrotizing enterocolitis (NEC), (sub)mucosal edema, hemorrhage, ulceration, or necrosis will disturb intestinal integrity, as reflected by an increased intestinal permeability. Enteral substrate is therefore withheld for a variable period up to 3 weeks (in many clinics). The authors used the sugar absorption test to measure intestinal permeability changes in surgically treated necrotizing enterocolitis patients and surgical controls to evaluate the usefulness of this test in timing the (re-)introduction of enteral feeding in NEC patients as intestinal integrity recovers. METHODS: Changes in intestinal permeability to lactulose and rhamnose were evaluated prospectively in 13 children with NEC and 10 operated control patients. The patients were given 1 mL/kg body weight lactulose/rhamnose solution at different time intervals after admission. The lactulose to rhamnose (L/R) ratio was determined by gaschromatography in 4-hour urine samples. RESULTS: The L/R ratios in NEC patients were increased for prolonged periods of time with a tendency to decrease in the third week after the start of NEC. However, in some cases, the increased L/R ratios even exceeded the 3-week period of starvation. High peaks in the L/R ratio were seen in patients suffering from bowel perforation or sepsis. Compared with necrotizing enterocolitis patients, L/R ratios of control patients were increased only in the first days after surgery and normalized more rapidly. The results of the L/R tests in this study corroborated the clinical condition of the patients. CONCLUSIONS: The sugar absorption test shows an individual variability in the recovery of intestinal permeability in a group of seriously ill newborns with advanced stages of NEC. An individual approach in restarting enteral nutrition seems to be justified; however, the optimal time-point to restart enteral nutrition cannot be determined by the sugar absorption test alone. Combining parameters of intestinal integrity and function could enable a more accurate determination of this optimal timepoint. J Pediatr Surg 36:587-592. PMID- 11283882 TI - Ultrasonographic features of normalization of the pylorus after pyloromyotomy for hypertrophic pyloric stenosis. AB - PURPOSE: The purpose of this study was to describe the time course, early postoperative changes, and morphologic features of normalization of the pylorus after pyloromyotomy for hypertrophic pyloric stenosis. METHODS: The subjects were 17 infants (9 boys, 8 girls) who underwent umbilical incision Ramstedt pyloromyotomy. The pyloric muscle mass was measured immediately before the operation and then at intervals from 3 days to 6 months after the operation using a 7.5-MHz ultrasound probe. RESULTS: In longitudinal section, the dorsal part of the pyloric muscle thickened transiently and then thinned to normal values by 5 months after the operation. It was 5.1 +/- 0.8 mm (mean +/- SD) preoperatively, increased to 6.0 +/- 0.3 mm by day 3 after the operation (P <.05), and thinned to 2.8 +/- 0.2 mm by 5 months after the operation. Concomitantly, the length of the pylorus gradually decreased (from 20.1 +/- 2.9 mm preoperatively to 16.9 +/- 2.7 mm by 3 days postoperatively [P <.05] and to less than 15 mm, by 4 months). In transverse section, the muscle normalized as in the longitudinal section. At the site of the incision it was 4.3 +/- 0.4 mm thick preoperatively, thickened to 4.6 +/- 0.4 mm by 3 days after the operation (P <.05), thinned to 2.1 +/- 0.9 mm by 7 days (P <.05), and then increased slightly, but always was less than 3.0 mm. Morphologically, in transverse section, the incised area looked like a wedge by 3 days after the operation. CONCLUSIONS: After pyloromyotomy for hypertrophic pyloric stenosis, there is an early transient increase in muscle thickness within the first few postoperative days followed by a slow decrease that reaches normal thickness (<3 mm) by 5 months. This decrease in thickness is accompanied by a gradual decrease in length to 75% of the preoperative value by 5 months. The morphologic features in this normalization are first a wedge (day 3), then a flat tire (days 7 and 14), and finally an elongated ring (5 months). J Pediatr Surg 36:582-586. PMID- 11283884 TI - Oxygen-induced vasodilation is blunted in pulmonary arterioles from fetal rats with nitrofen-induced congenital diaphragmatic hernia. AB - BACKGROUND/PURPOSE: Persistent pulmonary hypertension contributes to the high mortality rate associated with congenital diaphragmatic hernia (CDH). Oxygen is an important stimulus for pulmonary vasodilation in the perinatal period. The authors have investigated the responses of isolated pulmonary arterioles from fetal rats with and without CDH to an increase in oxygen tension. METHODS: CDHs were induced in fetal rats by feeding nitrofen to timed-pregnant rats at midgestation. A third-generation pulmonary arteriole was isolated from the right lung at term. Isolated arterioles were pressurized at their "optimal distending pressure." Diameter changes in response to an increase in oxygen tension from 25 to 40 mm Hg ("hypoxic" conditions) to 90 to 150 mm Hg ("normoxic" conditions) were recorded for K(+) preconstricted arterioles from control rats, from rats with nitrofen-induced CDH, and from rats that were nitrofen exposed but did not have a CDH. RESULTS: "Normoxic" exposure reversed the K(+) preconstriction in control arterioles by 124 +/- 26%. In contrast, arterioles from rats with nitrofen-induced CDH dilated significantly less than controls (20 +/- 15% of the K(+) preconstriction). The responses of arterioles from rats that were nitrofen exposed but did not get a CDH were not different (P >.05) from controls. CONCLUSIONS: Oxygen-induced vasodilation is blunted in pulmonary arterioles from rats with nitrofen-induced CDH. Blunted oxygen-induced vasodilation may contribute to persistent pulmonary hypertension in CDH. J Pediatr Surg 36:593 597. PMID- 11283885 TI - Xanthogranulomatous pyelonephritis in childhood. AB - PURPOSE: Demographic data, clinical presentation, associated abnormalities, and radiologic findings were evaluated to outline diagnostic criteria that may lead to the diagnosis of xanthogranulomatous pyelonephritis in children. METHODS: Eleven boys and 8 girls with a mean age of 3.4 +/- 1.7 years were classified into obstructive xanthogranulomatous pyelonephritis (n = 13), which was associated with nephrolithiasis and nonobstructive (n = 6), which mimicked Wilms' tumor. RESULTS: Twelve children with obstructive diffuse involvement of the renal parenchyma, 1 with left-sided obstructive focal involvement in a horseshoe kidney (group 1), and 6 with nonobstructive diffuse xanthogranulomatous pyelonephritis (group 2) showed a male to female ratio of 1.2:1 and 2:1, respectively. Mean age was 4.1 +/- 1.2 years in group 1 versus 1.8 +/- 1.5 years in group 2; P =.001. The common features were renal mass, hematuria, and anemia (100%; P =.07) and leucocytosis (77% v 83%; P =.097). Main differences between the 2 groups were acute inflammatory syndrome (0 v 33%; P =.01), recurrent urinary infection (54% v 17%; P =.05), isolation of Proteus mirabilis as a pathogen (69% v 0; P =.001), and renal stones (100% v 0; P =.001). Preoperative diagnosis was accurate in all 13 (100%) children with obstructive xanthogranulomatous pyelonephritis. Radiologic features that were not consistent with Wilms' tumor in group 2 were absence of sharp definition and encapsulation of the mass, ill-defined margins with inflammatory infiltration of the perinephric fat and focal inflammatory tissue destruction. Nephrectomy was technically difficult because of extensive adhesions to the retroperitoneum, psoas muscle, and surrounding structures in both groups. CONCLUSIONS: Xanthogranulomatous pyelonephritis must be considered in the differential diagnosis of a child presenting with a renal mass, anemia, and elevated inflammatory markers. Treatment by nephrectomy is curative. J Pediatr Surg 36:598-601. PMID- 11283886 TI - Experimental small bowel transplantation using newborn intestine in rats: III. Long-term cryopreservation of rat newborn intestine. AB - BACKGROUND: If long-term organ cryopreservation can be attained, a significant achievement will have been made to address the problem for donor shortage. Fetal intestine has been known to revascularize naturally without vascular anastmosis. The authors have confirmed previously that the newborn intestine also could develop to maturity in the host omentum. Here, the authors examined whether the cryopreserved newborn intestine could revascularize in the syngeneic combination using the 2 different solutions and whether cryopreservation affect their antigenicity in the allogeneic combination. METHODS: Inbred rat strains of LEW (MHC haplotype; RT1(l)) and BN (RT1(n)) were used. LEW newborn intestinal grafts were stored in RPMI-1640 or University of Wisconsin solution with 10% DEMSO (n = 10 in each group). The grafts were placed into a cold (4 degrees C) preservation solution for 30 minutes and then placed into a freezing chamber and cooled to -80 degrees C at -1 degrees C/min after 12 hours quenched to -180 degrees C in liquid nitrogen for longer than 30 days. Then, the cryopreserved grafts under the 2 different solutions were transplanted syngenicaly (LEW to LEW). The cryopreserved BN grafts also were implanted into the LEW omentum pouch. The allotransplantation was received with a 14-day high-dose course of tacrolimus (0.64 mg/kg, intramuscularly). The grafts were evaluated histologically at 4 weeks after transplantation. Fresh newborn intestines implanted in this syngeneic and allogeneic combination were evaluated as each control group. RESULT: In the syngeneic combination, more than 90% of the mature intestine were obtained. There was no significant difference among the different solution and the fresh group. However, in the allogeneic combination, both fresh and cryopreserved grafts were histologically poor. CONCLUSIONS: This is the first report showing that long-term cryopreservation was not harmful for neovascularization of newborn intestine. Long-term cryopreservation did not reduce the antigenicity of the newborn intestine. J Pediatr Surg 36:602-604. PMID- 11283887 TI - Experiences of a parent support group with the long-term consequences of esophageal atresia. AB - PURPOSE: The aim of this study was to study long-term effects of esophageal atresia based on data from the world's largest parent support group. METHODS: A questionnaire was completed by 128 former patients, now aged 10 to 34 years (median, 14 years), who were all members of the support group. RESULTS: Postoperative bougienage was performed in 70% of patients. The most frequently associated anomalies were skeletal (44%), with 18% of patients having a winged scapula. In 30% of patients, food became trapped; this entrapment continues, although with decreasing frequency. In 38% of patients, recurrent episodes of food lodging in the esophagus lasted only a few years, whereas 32% of patients never experienced any swallowing problems. The foods most often trapped were apples, meat, and bread. Most trapped food is removed by the induction of vomiting, without medical help. Unusual findings of this study were the avoidance of chocolate and sweets by 9% of patients and the habit of eating unorthodox food combinations in 13% of patients. Forty-six percent of patients have gastroesophageal reflux, and 7% have Barrett's esophagus. The most successful antireflux prevention was abstinence from eating late in the evening. Fundoplication was performed in 16% of patients; however, only 25% of these fundoplication operations were successful immediately. Eighty percent of patients had more than 4 lung infections or bronchitis episodes per year. Forty-four percent make noises, such as wheezing or whistling, when breathing or straining. Eighty-eight percent are of normal height, and 70% are of normal weight. Eighty nine percent of patients attended regular schools, and 52% were above-average students. CONCLUSIONS: Very few pediatric surgeons have had the opportunity to follow into adulthood such a large group of patients with esophageal atresia. Patients themselves feel lost when they undergo the transition from pediatric to adult medical treatment. A support group is able to provide an information database, which helps overcome the isolation because of the rareness of the disease and the separation of patients into different medical specialties. Surgeons should be aware of the difficulties that patients and parents face in later life and should seek cooperation with a support group. J Pediatr Surg 36:605-610. PMID- 11283888 TI - Functional treatment of physeal and periphyseal injuries of the metacarpal and proximal phalangeal bones. AB - BACKGROUND/PURPOSE: Hand fractures are common injuries in infants. Complications are rare because of potent remodeling dimension and rapid healing of growing bone. There is limited remodeling capacity for angular and rotational deformity so displaced fractures often require open reduction and internal fixation. METHODS: The authors present a splint system for a protected reduction and mobilization program of displaced proximal phalanx and metacarpal fractures. The custom-molded 2-component thermoplastic splint allows motion of the proximal interphalangeal (PIP) and distal interphalangeal (DIP) joints. It has been developed to allow bone healing and recovery of motion at the same time. In this study, the authors evaluated the clinical and radiologic results of a series of 11 consecutive infants with displaced metacarpal fractures and 13 displaced proximal phalanx fractures who received functional treatment. RESULTS: Fracture consolidation and full active motion was achieved simultaneously in 4 weeks in 21 children; 2 infants required physiotherapy, and 1 child was lost to follow-up. No further growth abnormality was seen within a 12-month observation period. CONCLUSION: When there is no damage of soft tissue the functional mobilization program can lead to good results treating displaced physeal and periphyseal hand injuries of proximal phalanx and metacarpal fractures. J Pediatr Surg 36:611-615. PMID- 11283889 TI - Nitric oxide synthase is absent in only a subset of cases of pyloric stenosis. AB - PURPOSE: The aim of this study was to study nitric oxide synthase (NOS) immunohistochemistry in the pyloric muscle and establish the role of nitric oxide in pyloric stenosis. METHODS: Pyloric muscle biopsy specimens were obtained from 20 patients with pyloric stenosis during pyloromyotomy. Ten control specimens without pyloric disease were obtained from autopsy performed less than 4 hours after death on age-matched babies who died of other causes. Tissues were fixed in 4% paraformaldehyde immediately. A monoclonal antibody against the neuronal form of NOS (bNOS) was used for immunohistochemistry. RESULTS: Immunohistochemistry showed activity of bNOS in the control specimens and some pyloric stenosis specimens. This shows that NOS is present in the pylorus in normal cases as well as in a few cases of pyloric stenosis. CONCLUSIONS: NOS deficiency leading to lack of locally available nitric oxide causes a failure of smooth muscle relaxation. This may account for the cause of pyloric stenosis in infants. However, this study shows that this is true probably only in a subset of cases. The etiology of pyloric stenosis may still be multifactorial. Further investigations are required regarding the etiology of pyloric stenosis. J Pediatr Surg 36:616-619. PMID- 11283890 TI - Congenital vulvar teratoma in a newborn. AB - The most common site of teratomas in neonates is in the sacrococcygeal region. Herein the authors describe a congenital teratoma, and to their knowledge it is the first reported case of vulvar site in a newborn. Tumor was removed on the fourth day of life, and she was completely normal 18 months after the operation. Because of possible malignancy or recurrence, complete surgical excision of the tumor and careful follow-up is the treatment of choice. J Pediatr Surg 36:620 621. PMID- 11283891 TI - Congenital absence of the portal vein associated with focal nodular hyperplasia of the liver and congenital choledochal cyst: a case report. AB - A congenital absence of the portal vein (CAPV) is a rare malformation, which almost is always associated with other anomalies such as hepatic tumors and cardiac malformations. This case report describes a 3-year-old girl with a congenital absence of the portal vein, focal nodular hyperplasia (FNH) of the liver, and a congenital choledochal cyst (CCC). Angiography findings showed the mesenteric vein and splenic vein to be joined together to form a common trunk that entered the inferior vena cava directly above the liver. This is the first known reported case of CAPV with concurrent CCC. J Pediatr Surg 36:622-625. PMID- 11283892 TI - Congenital diaphragmatic hernia associated with a gastroesophageal duplication cyst: a case report. AB - Severe left congenital diaphragmatic hernia was diagnosed in a baby prenatally, and she underwent hernia repair on the sixth postnatal day of life. She was found to have a huge symptomatic gastroesophageal duplication cyst on day 24 of life. A thoracoabdominal dissection allowed successful cyst excision. J Pediatr Surg 36:626-628. PMID- 11283893 TI - Esophageal squamous cell carcinoma 38 years after primary repair of esophageal atresia. AB - The authors report a case of a 38-year-old man with an esophageal squamous cell carcinoma after repair of esophageal atresia with tracheoesophageal fistula. This carcinoma occurred at a young age, near to the scar of the old anastomosis, in a patient with no other apparent risk factors. It is hypothesized that stasis caused by impaired esophageal motility may be the underlying cause. A single case is not enough to unequivocally prove a possible relationship between esophageal atresia and the development of esophageal cancer. Now that the first generation of survivors of esophageal atresia is reaching middle aged adulthood, one should, however, be aware of a possible increased incidence in these patients. J Pediatr Surg 36:629-630. PMID- 11283894 TI - Inflammatory pseudotumor of the trachea. AB - Primary tracheal tumors are extremely rare, and the majority of them are malignant. Inflammatory pseudotumor is a benign, tumorlike lesion, most likely of a reactive nature. Its basic morphologic characteristic is spindle cell (myoblasts and fibroblasts) proliferation with a variable number and type of inflammatory cells. A case of intratracheal inflammatory pseudotumor in a 14-year old boy is presented together with a review of all similar lesions in the available literature. The discussion includes the presentation of tracheal tumors, their basic morphologic and immunohistochemical characteristics, and treatment modalities that are available. The surgeon must exercise caution not to perform radical surgery based on the initial pathologic diagnosis from the intraoperative frozen section, because the prognosis of these benign lesions generally is excellent. This is the second reported case of intratracheal inflammatory pseudotumor successfully endoscopically vaporized using a CO(2) laser, which is an excellent alternative in cases in which surgical treatment is feasible. J Pediatr Surg 36:631-634. PMID- 11283895 TI - Association of biliary atresia and urogenital sinus. AB - A case of an association of extrahepatic biliary atresia (EHBA) and urogenital sinus (UGS) anomaly that was diagnosed as an urachal remnant antenatally is reported. Diagnostic laparoscopy of the abdominal cavity in the postnatal period was the most helpful step for making the diagnosis. Biliary atresia and urogenital sinus, which was the cause of bladder outlet obstruction, were treated successfully. To the authors' knowledge, this is the first case of this type of association introduced to the literature. J Pediatr Surg 36:635-637. PMID- 11283896 TI - Gastroschisis and Hirschsprung's disease: a rare combination. AB - The authors report on a neonate with gastroschisis repaired at birth who later had abdominal distension, emesis, feeding intolerance, and an abnormal stooling pattern. Total colon and partial small bowel aganglionosis (TCAS), or Hirschsprung's disease, was diagnosed subsequently. This is the first report of this combination of gastrointestinal anomalies. J Pediatr Surg 36:638-640. PMID- 11283897 TI - Down's syndrome, precocious puberty, and transverse vaginal septum: an unusual cause of abdominal pain. AB - Hematocolpos should be considered in adolescent girls who present with lower abdominal pain, a pelvic mass, and primary amenorrhea. The authors describe a rare case of a young child with Down's syndrome, precocious puberty, and hematocolpos caused by a transverse vaginal septum. The diagnosis was facilitated using a combination of computed tomography and ultrasound scanning. J Pediatr Surg 36:641-643. PMID- 11283898 TI - Milk of calcium cholelithiasis in children. AB - Milk of calcium bile is uncommon and occurs mainly in the adult population. The authors report on 2 children, each having a distinct clinical history and presentation, and each with milk of calcium bile/calculi possessing differing chemical composition and highly notable gross morphology. J Pediatr Surg 36:644 647. PMID- 11283899 TI - Ischemic small bowel strictures in a case of incomplete Kawasaki disease. AB - The authors describe an atypical case of Kawasaki disease in a 9-month-old girl who presented with fever, coronary artery aneurysms, and acquired ischemic stricture of the proximal jejunum. Histology of the surgical specimen was consistent with mesenteric vasculitis. The infant had only some of the typical clinical signs of Kawasaki disease, suggesting that an atypical or incomplete form of the disease was present. To the best of the authors' knowledge this is only the third case to be reported of incomplete Kawasaki disease associated with ischemic bowel stricture. J Pediatr Surg 36:648-650. PMID- 11283900 TI - Multiple intestinal stenoses and peripheral gangrene: a combination of two rare surgical complications in a child with Kawasaki disease. AB - Kawasaki disease (KD) often presents with a challenging variety of clinical symptoms. Severe gastrointestinal complications are rare and mainly appear as pseudo-obstruction. However, the authors report the unique case of a 4-month-old girl with KD suffering from a mechanical ileus. The optimal timing of surgery presented a dilemma, because she received lytic treatment for gangrenes of both her hands and feet and additionally had large coronary artery aneurysms. Finally laparotomy had to be performed while the patient was on an anticoagulant medication, and it showed a 30-cm-long jejunal segment with multiple filiforme stenoses, requiring resection and anastomosis. The postoperative course was uneventful regarding the abdominal situation; however, the left hand and left foot remained gangrenous and had to be amputated. In patients with KD, not only pseudo-obstruction, but irreversible intestinal obliteration has to be encountered. This combination of intestinal stenosis and acral gangrene has not been described before. J Pediatr Surg 36:651-653. PMID- 11283901 TI - Prolonged use of tissue plasminogen activator for bilateral lower limb arterial occlusion in a neonate. AB - The authors report the case of a 29-week-gestation twin with very severe vascular compromise of both lower limbs secondary to premature twin-twin transfusion. A tissue plasminogen activator (tPA) infusion was used locally for 13 days with complete recovery of perfusion to both legs, demonstrated by serial angiograms, thereby avoiding bilateral amputation. There were no side effects as a result of the continuous administration of tPA. The authors therefore suggest that the benefits of thrombolysis from a local infusion of tPA in neonates may outweigh the potential risks. This case report thus supports the view that under certain circumstances infusion of tPA in neonates may offer significant benefits. J Pediatr Surg 36:654-656. PMID- 11283902 TI - Congenital abdominal aortic aneurysm in the infant: case report and review of the literature. AB - Congenital abdominal aortic aneurysms are a distinct entity from acquired aortic aneurysms. The authors present the case of a 6-week-old boy with a 6-cm aneurysm involving the abdominal aorta and common iliac arteries. Three other cases of congenital aortic aneurysms are reviewed, and an approach to these rare patients is discussed. J Pediatr Surg 36:657-658. PMID- 11283903 TI - Living donor liver transplantation for Budd-Chiari syndrome with inferior vena cava obstruction and associated antiphospholipid antibody syndrome. AB - BACKGROUND: Recently, the antiphospholipid antibody syndrome (APLS) has been recognized as the cause of the Budd-Chiari syndrome (BCS) in adults. However, APLS-induced BCS has been seen rarely in children. The surgical strategy for BCS depends on the patency of the inferior vena cava (IVC) and on the primary disease. METHODS: A 10-year-old boy with a diagnosis of BCS complicated with IVC obstruction caused by APLS received medical treatment and radiologic intervention for 2 years. In spite of these treatments, no relief of the symptoms could be achieved. Finally, the patient underwent living donor liver transplantation (LDLT), which required cavoplasty. He has had an uneventful course since the LDLT. CONCLUSIONS: IVC-obstructed BCS associated with APLS seems to be a good indication for LDLT with cavoplasty. Because liver transplantation (LT) itself is not a cure for APLS, the risk of thrombosis cannot be eliminated entirely by LT. Although immunosuppression may influence antiphospholipid antibody production, long-term observation and life-long anticoagulant treatment is needed even after LT. J Pediatr Surg 36:659-662. PMID- 11283904 TI - Inflammatory pseudotumor of the liver: case report and review of the literature. AB - Inflammatory pseudotumor is a rare lesion that generally is considered to be benign in biological behavior, although some may recur or metastasize. The authors report on a patient with inflammatory pseudotumor of the liver whose preoperative radiologic findings resembled those of focal nodular hyperplasia. The biological investigation showed a polyclonality of the cells and diploidy of the DNA content and suggested benign characteristics of the lesion. J Pediatr Surg 36:663-666. PMID- 11283905 TI - Successful heterotopic segmental liver transplantation from a live donor to a patient with Alagille syndrome. AB - Alagille syndrome is characterized by a paucity of bile ducts in the liver. The syndrome is associated with some or all the features of chronic cholestasis, cardiac disease, skeletal abnormalities, ocular defects and a distinctive facial appearance. The most common finding is chronic cholestasis, which causes intractable pruritus, xanthoma, deficiency of certain metabolic nutrients and growth retardation. Cardiac abnormalities are the most common cause of death in these patients. It is unusual to see the clinical picture of hepatic failure resulting in cirrhosis and requiring transplantation, but liver transplantation is indicated in Alagille syndrome patients who have chronic cholestasis. If the disease is diagnosed in childhood, transplantation can improve significantly the patient's prognosis and the quality of life. In recent years, auxiliary liver transplantation has gained popularity for treating both acute and chronic liver disease. Heterotopic segmental liver transplantation is an alternative treatment modality for patients who do not require native liver removal. Individuals with Alagille syndrome are good candidates for this type of treatment. J Pediatr Surg 36:667-671. PMID- 11283906 TI - Re.: "Testicular Sparing Surgery for Benign Testicular Tumours". PMID- 11283907 TI - Re.: "Histology of the Neobladder Mucosa After Sigmoidocolocystoplasty". PMID- 11283909 TI - Characterization of the stability of recombinant protein production in the GS-NS0 expression system. AB - The GS-NS0 system is an important mammalian expression system used largely within industry for the high-level expression of recombinant proteins for therapeutic use. It is essential that the productivity of this system remains stable throughout culture expansion for the successful long-term production of recombinant proteins. Here we present a study of the stability of recombinant protein production from unamplified GS-NS0 cell lines over extended period of continuous culture. The cell lines used in this study were generated by the transfection of NS0 cells with DNA encoding for a secreted recombinant protein and by two subsequent rounds of limiting dilution cloning prior to analysis of stability. The stability of recombinant protein production was assessed at intervals over a period of 134 days using repeated batch culture in shake flasks. Heterogeneous stability was identified. The productivity of some clones remained consistent throughout 134 days of continuous culture. Others exhibit rapid and progressive loss of productivity. Analysis of the causal relationships underlying stability indicates that the initial transfectant determines the susceptibility to loss or retention of productivity. Selection of production clones on the basis of growth and productivity alone will not predict stability during long-term culture. Our research indicates that stable high-producing clones can readily be obtained from use of the GS-NS0 system in the absence of amplification but there may be molecular features of the original transfectants that could serve as very important predictive indicators of the stability of recombinant protein production. PMID- 11283910 TI - Transformation of 2,4,6-trinitrotoluene (TNT) by immobilized Phanerochaete chrysosporium under fed-batch and continuous TNT feeding conditions. AB - The cometabolic transformation of 2,4,6-trinitrotoluene (TNT) by an immobilized Phanerochaete chrysosporium culture was investigated under different TNT and/or glycerol feeding conditions in a 5-L reactor. In the fed-batch feeding mode, as a result of four spiking events at an average feeding rate of 20 mg TNT L(-1) d(-1) and 250 mg glycerol L(-1) d(-1), the initial TNT transformation rate and the glycerol uptake rate of the 7-day-old immobilized cell culture were 2.41 mg L(-1) h(-1) and 16.6 mg L(-1) h(-1), respectively. Thereafter, the TNT fed into the reactor depicted a negative effect on the cell physiology of P. chrysosporium, i.e., both rates decreased constantly. At 32 mg TNT L(-1) d(-1) feeding rate, also in the presence of glycerol (200 mg L(-1) d(-1)), this effect on the fungal cell metabolism was even more significant. When TNT was fed alone at 3.7 mg L(-1) d(-1), it showed an initial 0.75 mg L(-1) h(-1) rate of TNT transformation, i.e., one-third the initial level observed in the presence of glycerol. In contrast, in the continuous feeding mode (dilution rate, D = 0.11 d(-1)), at 5.5 mg TNT L(-1) d(-1) and 220 mg glycerol L(-1) d(-1), the immobilized cell culture exhibited a constant TNT transformation rate for cultivation periods of 50 and 61 days, under uncontrolled and controlled pH conditions, respectively. Thereafter, during the latter experiment, 100% TNT biotransformation was achieved at 1,100 mg L(-1) d( 1) glycerol feeding rate. Immobilized cells (115-day-old), sampled from a continuous TNT feeding experiment, mineralized [(14)C]-TNT to a level of 15.3% following a 41-day incubation period in a microcosm. PMID- 11283911 TI - Dynamic volume-averaged model of heat and mass transport within a compost biofilter: I. Model development. AB - Successful, long-term operation of a biofilter system depends on maintaining a suitable biofilm environment within a porous medium reactor. In this article a mathematical study was conducted of the spatial and temporal changes of biofilter performance due to interphase heat and mass transport. The method of volume averaging was used to spatially smooth the three-phase (solid, liquid, and gas) conservation equations over the biofilter domain. The packing medium was assumed to be inert, removing the solid phase mass continuity equation from the system. The finite element method was used to integrate the resulting nonlinear-coupled partial differential equations, tracking eight state variables: temperature, water vapor, dry air, liquid water, biofilm, gas and liquid phase organic pollutant, and nutrient densities, through time and space. A multiphase, gas and liquid flow model was adapted to the biofilter model from previous studies of unsaturated groundwater flow. Newton's method accelerated by an LU direct solver was used to iterate the model for solutions. Effects of packing media on performance were investigated to illustrate the utility of the model. The moisture dynamics and nutrient cycling are presented in Part II of this article. PMID- 11283912 TI - Wet extrusion of fibronectin-fibrinogen cables for application in tissue engineering. AB - A method for the wet extrusion of human plasma-derived fibronectin-fibrinogen cables is described. Solutions of fibronectin and fibrinogen with and without sodium alginate and carboxymethylcellulose (CMC) are tested. The rheological properties of the protein solutions changed from Newtonian to shear thinning non Newtonian in the presence of small quantities of these additives, the apparent viscosity increased, and the extrusion properties of the protein solutions improved. Cables were prepared using a capillary with a diameter of 1 mm and overall length of 18 mm. Cable diameter was reduced to about 0.5 mm by drawing using a series of rollers. Cables prepared with sodium alginate were found to have suitable properties, and those made with CMC were sticky and difficult to handle. Solutions containing no sodium alginate required a minimum total protein concentration of about 70 mg/mL for extrusion. Extruded cables were prepared with solutions containing 140 mg/mL total protein with 12.9 mg/mL alginate (high protein), and 46 mg/mL total protein with 47.6 mg/mL of sodium alginate (high alginate). The mechanical strength of the extruded cables was within the range suitable for application in tissue engineering. Extrusion of the protein solutions into cables was achieved in a coagulation bath. Cables with a mechanical strength of approximately 30 N/mm(2), suitable for wound repair and nerve regeneration applications, were prepared with a coagulation bath containing 0.25 M HCl, 2% CaCl(2) at a pH of <0.9. These cables also had a large average elongation at break of 52%, and showed an increase in cable length after breakage (permanent set) of 20%, demonstrating the potential for drawing the cables down to a fine diameter. PMID- 11283913 TI - Process for degradation of nitrobenzene: combining electron beam irradiation with biotransformation. AB - Electron beam irradiations of aqueous solutions containing 15-30 mg/L of nitrobenzene at 60 kGy dose removed 78% of the contaminant. Three mononitrophenols were detected as by-products of electron beam treatment of nitrobenzene. A mixed culture enriched on a mixture of 2-, 3-, and 4-nitrophenol degraded both the residual nitrobenzene and the nitrophenol products. Percentage removal of nitrobenzene increased with increasing electron beam dose. This observation led to the conceptual design of a two-stage electron beam microbial process for degradation of nitrobenzene. Three groups of pure isolates were characterized from the mixed culture based on their abilities to grow on cor- responding nitrophenol substrates: Group A, 2NP(-)3NP(-)4NP(+); Group B, 2NP(+)3NP(+)4NP(-); and Group C, 2NP(-)3NP(+)4NP(-). Bacteria that grew on 3-NP transformed nitrobenzene into ammonia in the electron beam-treated nitrobenzene samples. PMID- 11283914 TI - Design of transgenes for efficient expression of active chimeric proteins on mammalian cells. AB - Heterologous proteins expressed on the surface of cells may be useful for eliciting therapeutic responses and engineering new extracellular properties. We examined factors that control the membrane targeting of alpha-fetoprotein (AFP) and a single-chain antibody (scFv). Chimeric proteins were targeted to the plasma membrane by employing the transmembrane domain (TM) and cytosolic tail of murine CD8O (B7-1), the TM of the human platelet-derived growth factor receptor (PDGFR), the glycosylphosphatidylinositol anchor encoded by the C-terminal extension of decay-accelerating factor (DAF), and the TM of the H1 subunit of the human asialoglycoprotein receptor (ASGPR). AFP chimeric proteins containing the B7, DAF, ASGPR, or PDGFR targeting domains displayed half-lives of 12.2, 3.8, 2.4, and 1.6 h, respectively. The newly synthesized B7 chimera was rapidly transported and remained on the cell surface. Glycosylphosphatidylinositol-anchored chimeras reached the surface more slowly and significant amounts were released into the culture medium. PDGFR TM chimeras were rapidly degraded, whereas ASGPR chimeras were retained in the endoplasmic reticulum (ER). The surface expression of both AFP and scFv chimeric proteins followed the order (highest to lowest) of B7 > DAF >> PDGFR. Introduction of a dimerization domain (hinge-CH(2)-CH(3) region of human IgG1) between scFv and TM dramatically reduced cleavage of the chimeric protein, increased surface expression, and produced biologically active scFv. Our results indicate that transgenes designed for the expression of active scFv on cells should incorporate a TM that does not undergo endocytosis, include an intact cytoplasmic domain, and possess a spacer to reduce cleavage and retain biological activity. PMID- 11283915 TI - Adsorption of avidin on microfabricated surfaces for protein biochip applications. AB - The adsorption of the protein avidin from hen egg white on patterns of silicon dioxide and platinum surfaces on a microchip and the use of fluorescent microscopy to detect binding of biotin are described. A silicon dioxide microchip was formed using plasma-enhanced chemical vapor deposition while platinum was deposited using radiofrequency sputtering. After cleaning using a plasma arc, the chips were placed into solutions containing avidin or bovine serum albumin. The avidin was adsorbed onto the microchips from phosphate-buffered saline (PBS) or from PBS to which ammonium sulfate had been added. Avidin was also adsorbed onto bovine serum albumin (BSA)-coated surfaces of oxide and platinum. Fluorescence microscopy was used to confirm adsorption of labeled protein, or the binding of fluorescently labeled biotin onto previously adsorbed, unlabeled avidin. When labeled biotin in PBS was presented to avidin adsorbed onto a BSA-coated microchip, the fluorescence signal was significantly higher than for avidin adsorbed onto the biochip alone. The results show that a simple, low-cost adsorption process can deposit active protein onto a chip in an approach that has potential application in the development of protein biochips for the detection of biological species. PMID- 11283916 TI - An odyssey of hope. PMID- 11283917 TI - Zoledronic acid reduces skeletal-related events in patients with osteolytic metastases. AB - BACKGROUND: This study evaluated the dose-response relation for zoledronic acid, a new generation high potency bisphosphonate, given as a 5-minute infusion in patients with malignant osteolytic disease. METHODS: Two-hundred eighty patients with osteolytic lesions due to metastatic breast carcinoma or multiple myeloma were randomized to double-blind treatment with either 0.4, 2.0, or 4.0 mg of zoledronic acid or 90 mg pamidronate. The primary efficacy endpoint was the proportion of patients receiving radiation to bone. Other skeletal-related events, bone mineral density (BMD), bone markers, Eastern Cooperative Oncology Group performance status, pain and analgesic scores, and safety also were evaluated. RESULTS: Zoledronic acid at doses of 2.0 and 4.0 mg and pamidronate at a dose of 90 mg each significantly reduced the need for radiation therapy to bone (P < 0.05) in contrast with 0.4 mg zoledronic acid, which did not. Skeletal related events of any kind, pathologic fractures, and hypercalcemia also occurred less frequently in patients treated with 2.0 or 4.0 mg zoledronic acid or pamidronate than with 0.4 mg zoledronic acid. Increases in lumbar spine BMD (6.2 9.6%) and decreases in the bone resorption marker N-telopeptide (range, -37.1 to 60.8%) were observed for all treatment groups. Skeletal pain, fatigue, nausea, vomiting, and headache were the most commonly reported adverse events. Adverse events were similar in nature and frequency with zoledronic acid and pamidronate. CONCLUSIONS: A 5-minute infusion of 2.0-4.0 mg zoledronic acid was at least as effective as a 2-hour 90-mg pamidronate infusion in treatment of osteolytic metastases. A 0.4-mg dose of zoledronic acid was significantly less effective. Both zoledronic acid and pamidronate were well tolerated. PMID- 11283918 TI - Prognostic factors and outcome of pelvic, sacral, and spinal chondrosarcomas: a center-based study of 69 cases. AB - BACKGROUND: The surgical treatment of chondrosarcoma of the pelvis, sacrum, and spine is complex and technically demanding. As such, adequate surgical margins have been difficult to achieve, resulting in poor local control and survival. The objective of this study was to assess the outcome of patients with chondrosarcomas in these sites who were treated at a tumor center by using modern, aggressive surgical techniques and to identify prognostic factors. METHODS: Sixty-nine consecutive patients with chondrosarcoma of the pelvis (46 cases), sacrum (11 cases), and mobile spine (12 cases) who were treated at Sahlgrenska University Hospital from 1967 to 1999 were included in this study. Demographic information and follow-up data were obtained and statistically analyzed. RESULTS: There were 53 men and 16 women with a mean age of 45 years and a mean tumor size of 12 cm. There were 61 conventional chondrosarcomas, Grades 1 3 (with 13 arising in a preexisting osteochondroma) and 8 Grade 4 chondrosarcomas (7 dedifferentiated and one mesenchymal). The overall local recurrence rate was 27%, and the estimated overall 5- and 10-year survival rates were 72% and 67%, respectively. In contrast, the observed local recurrence rate was 3% (1 patient) in 31 patients whose conventional chondrosarcomas were resected with adequate surgical margins; 90% of these patients survived and most of them (26 of 31 or 84%) were continuously disease free. Significant factors associated with a worse prognosis with respect to local control and/or survival were high histologic tumor grade, increasing patient age, primary surgery outside of a tumor center, incisional biopsy versus a noninvasive diagnostic procedure, and inadequate surgical margins. CONCLUSIONS: Center-based diagnosis and treatment using modern aggressive surgical techniques significantly improve the prognosis of patients with chondrosarcoma of the pelvis, sacrum, and spine. PMID- 11283919 TI - Significance of epitrochlear lymph node involvement in Hodgkin disease. AB - BACKGROUND: Epitrochlear involvement in Hodgkin disease (HD) is a rare event, with only limited data available describing this unique presentation, its treatment, and long term outcome. METHODS: Between 1968 and 1997, 1180 patients with clinical stage (CS) IA-IIB HD were treated at the Harvard Longwood Area Hospitals, among whom 11 were identified to have presented with epitrochlear lymphadenopathy (1%). Together with 6 CS III-IV patients also with clinically involved epitrochlear lymph nodes at diagnosis, these 17 patients form the basis of the current study. The clinical characteristics, histopathologic distribution, and treatment details are described. Two radiation therapy techniques were used: the "single field" and "separate-field" techniques. The median dose to the epitrochlear region was 3600 centigrays. Survival outcome was calculated by the Kaplan-Meier method. The median follow-up was 17 years. RESULTS: The actuarial 15 year freedom from recurrence, cause specific survival, and overall survival (OS) rates for the 17 patients were 70%, 88%, and 70%, respectively. Among the CS IA IIB patients, the 15-year OS rates of the 1169 patients and 11 patients without and with epitrochlear involvement were 80% and 90%, respectively. Two of the 11 CS IA-IIB and 3 of the 6 CS III-IV patients experienced recurrence. None of the recurrences involved the epitrochlear or ipsilateral brachial region regardless of the treatment technique, and no complications from the local radiation therapy were observed. CONCLUSIONS: Feasible and effective radiation therapy techniques are available for patients with HD with epitrochlear involvement. If appropriately treated, the prognosis of patients with this unique presentation appears to be similar to that of other HD patients. PMID- 11283921 TI - The impact of a multidisciplinary breast cancer center on recommendations for patient management: the University of Pennsylvania experience. AB - BACKGROUND: Advances in the diagnosis and treatment of breast carcinoma have led to a multidisciplinary approach to management for patients with breast carcinoma. To assess the effect of this approach, the authors performed an evaluation for a cohort of patients examined in a multidisciplinary breast cancer center. METHODS: An analysis was performed for the records of 75 consecutive women with 77 breast lesions examined in consultation in a multidisciplinary breast cancer center between January and June 1998. Each patient's case was evaluated by a panel consisting of a medical oncologist, surgical oncologist, radiation oncologist, pathologist, diagnostic radiologist, and, when indicated, plastic surgeon. A comprehensive history and physical examination was performed, and the relevant mammograms, pathology slides, and medical records were reviewed. Treatment recommendations made before this evaluation were compared with the consensus recommendations made by the panel. RESULTS: For the 75 patients, the multidisciplinary panel disagreed with the treatment recommendations from the outside physicians in 32 cases (43%), and agreed in 41 cases (55%). Two patients (3%) had no treatment recommendation before consultation. For the 32 patients with a disagreement, the treatment recommendations were breast-conservation treatment instead of mastectomy (n = 13; 41%) or reexcision (n = 2; 6%); further workup instead of immediate definitive treatment (n = 10; 31%); treatment based on major change in diagnosis on pathology review (n = 3; 9%); addition of postmastectomy radiation treatment (n = 3; 9%); or addition of hormonal therapy (n = 1; 3%). CONCLUSIONS: The multidisciplinary breast cancer evaluation program provided an integrated program in which individual patients were evaluated by a team of physicians and led to a change in treatment recommendation for 43% (32 of 75) of the patients examined. This multidisciplinary program provided important second opinions for many patients with breast carcinoma. PMID- 11283920 TI - Abnormalities of bone marrow mesenchymal cells in multiple myeloma patients. AB - BACKGROUND: The importance of the bone marrow microenvironment in multiple myeloma is receiving increasing attention. Recent studies have suggested the importance of cytokine production and cell-cell contact by bone marrow stromal cells in the survival of myeloma cells. METHODS: In the current study, the authors examined bone marrow mesenchymal progenitor cell (MPC) cultures derived from eight multiple myeloma patients (mean age, 58 years) and nine normal donors (mean age, 61 years), with emphasis on cell surface antigens, cytokine, and growth factor expression. RESULTS: The authors have found, based on analysis of cellular receptors, growth factors, and cytokine expression, that myeloma MPCs are phenotypically and functionally distinguishable from normal donor MPCs. Immunofluorescence analysis of MPC monolayers shows that myeloma MPC cultures expressed reduced cell surface vascular cell adhesion molecule-1 and fibronectin, in contrast with the strong expression found on normal donor MPCs. Furthermore, a subset of myeloma MPCs strongly express intracellular receptor for hyaluronan mediated motility, whereas normal MPCs do not. Cytokine expression in bone marrow MPC cultures was examined by reverse transcription-polymerase chain reaction and enzyme linked immunosorbent assay. Bone marrow MPCs constitutively express interleukin (IL)-1beta, IL-6, granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage (GM)-CSF, stem cell factor (SCF), and tumor necrosis factor (TNF)-alpha. In comparison to normal MPCs, multiple myeloma MPCs express increased basal levels of IL-1beta and TNF-alpha. In vitro exposure of MPC cultures to dexamethasone resulted in the down-regulation of IL-6, G-CSF, and GM CSF in both normal and myeloma MPC cultures. However, dexamethasone treatment significantly increased expression of SCF-1 in myeloma MPCs. CONCLUSIONS: In myeloma, bone marrow stromal cells provide paracrine factors, through cytokine production and cell-cell contact, which play a role in plasma cell growth and survival. The authors' data indicate differences in bone marrow MPCs, which may be biologically relevant to the growth and survival of myeloma plasma cells. PMID- 11283922 TI - Quality of life after breast carcinoma surgery: a comparison of three surgical procedures. AB - BACKGROUND: Because breast-conserving surgery (BCS), mastectomy alone, and mastectomy with reconstruction are equally effective for the treatment of early stage breast carcinoma, women's choice among them often focuses on quality-of life (QOL) issues. Information regarding QOL after these surgical treatments could help women with this decision. METHODS: Participants in this prospective study were women, age 30-85 years, with newly diagnosed breast carcinoma who underwent BCS (n = 103), mastectomy alone (n = 55), or mastectomy with reconstruction (n = 40). Quality of life was assessed after diagnosis (baseline) and at 1, 3, 6, 12, 18, and 24 months after baseline by using the Mischel Uncertainty in Illness Scale, Profile of Mood States, and Functional Assessment of Cancer Therapy for Breast Cancer. RESULTS: In multivariate regression analyses controlling for the QOL score obtained at baseline, age, and type of nonsurgical treatment, women who underwent mastectomy with reconstruction had greater mood disturbance (P = 0.002) and poorer well-being (P = 0.002) after baseline than women who had mastectomy alone; these differences remained 18 months after surgery. Although similar analyses also showed that women who underwent BCS had more mood disturbance than women who had mastectomy alone, this difference was significant only at 12 months after baseline. The BCS and mastectomy-only group did not differ significantly regarding well-being. CONCLUSIONS: Aspects of QOL other than body image are not better in women who undergo BCS or mastectomy with reconstruction than in women who have mastectomy alone. In fact, mastectomy with reconstruction is associated with greater mood disturbance and poorer well-being. PMID- 11283923 TI - Peritoneovenous shunting for nongynecologic malignant ascites. AB - BACKGROUND: The development of malignant ascites has been associated with a poor prognosis. Previous reports have documented high morbidity rates associated with placement of palliative peritoneovenous shunts (PVS). Most study series have included gynecologic malignancies in their analysis, and wide variations in survival time have been reported. Reported data from nongynecologic malignancies and identification of preoperative factors associated with improved outcome were the concerns of the current study, which attempted to identify patients with malignant ascites who might have benefitted from PVS. METHODS: A retrospective chart review was performed and data including age, gender, weight, preoperative laboratory values, cytology on peritoneal fluid aspirates, and complications within 30 days of the operative procedure were obtained and recorded. Discharge date and follow-up status were obtained for all patients. Statistical analysis was done for categorical values by comparing survival times from date of procedure with follow-up times using the log rank test. Significance for numeric values was determined with Cox regression analysis. Multivariate analysis using Cox regression was performed for those values found to be significant on univariate analysis. RESULTS: Fifty- five patients who had undergone PVS from 1980-1996 for ascites on the Gastric and Mixed Tumor service at the Memorial Sloan-Kettering Cancer Center were identified. Two patients with benign disease and two patients with ovarian malignancies were excluded. The remaining 51 patients underwent placement of 53 PVSs for palliation. Median survival time for the entire group was 52 days. Univariate analysis identified preoperative blood urea nitrogen (BUN), creatinine (Cr), BUN to Cr ratio, and diagnosis as significant factors. Preoperative BUN emerged as an independent predictor of survival by multivariate analysis, and those patients who had a BUN value of < = 17 demonstrated a survival advantage over those with a BUN of > 17. The assessable palliation factors were hospital discharge (80% of patients) and weight loss after shunting (68% of patients lost > 1 kg). Ninety-six percent of patients (24 of 25) with a preoperative BUN of < or = 17 were discharged. CONCLUSIONS: The development of nongynecologic malignant ascites is an end stage event for most patients. The placement of PVS for those patients with nongastrointestinal tumor etiologies, a BUN of < 17, a Cr of < or = 1.1, and a BUN to Cr ratio of < 19 yielded the best results. In the current study, palliation was difficult to assess accurately, although most patients were discharged or lost > 1kg of weight after shunting. PMID- 11283924 TI - Phase II study of oral eniluracil, 5-fluorouracil, and leucovorin in patients with advanced colorectal carcinoma. AB - BACKGROUND: The oral administration of 5-fluorouracil (5-FU) is hindered by erratic bioavailability due to catabolism of 5-FU by the enzyme dihydropyrimidine dehydrogenase (DPD) in the gastrointestinal tract. Eniluracil is a potent inactivator of DPD which results in 100% oral bioavailability of 5-FU. Leucovorin (LV) is another biochemical modulator of 5-FU that potentiates inhibition of thymidylate synthase, the primary target of 5-FU. The goal of this study was to determine the antitumor activity and toxicity of an oral regimen containing eniluracil, 5-FU, and LV in patients with colorectal carcinoma. METHODS: Sixty eligible patients who had previously untreated, measurable, metastatic colorectal carcinoma were treated with oral eniluracil 50 mg on Days 1-7, 5-FU 20 mg/m(2) on Days 2-6, and LV 50 mg on Days 2-6. Cycles were repeated at 28-day intervals. RESULTS: The overall response rate was 13% (95% confidence interval [CI] = 6, 25%), with 1 complete response and 7 partial responses. Three additional patients had partial responses that were not confirmed at subsequent evaluations. The median time to progression of disease was 4.4 months (95% CI = 3.45, 7.69) and the median survival time was 12.6 months (95% CI = 9.1, 14.75). Grade 3-5 toxicity (1 toxic death) occurred in 51 patients (85%). Grade 4 neutropenia occurred in 25 patients (42%), and 18 patients (30%) had Grade 3-4 diarrhea. Twenty-one patients (35%) were hospitalized for toxicity, and 12 (20%) had febrile neutropenia. Baseline creatinine clearance was associated inversely with severe toxicity (P = 0.001). CONCLUSIONS: Although antitumor activity was observed, the frequent occurrence of severe toxicity with this regimen limited its clinical utility. Alternate schedules with a more favorable therapeutic index are undergoing clinical testing and should be pursued. The high level of toxicity observed with orally administered low dose 5-FU underscored the potency of eniluracil as a biochemical modulator. PMID- 11283925 TI - Oxaliplatin plus raltitrexed in patients with advanced colorectal carcinoma: results of a Phase I-II trial. AB - BACKGROUND: Oxaliplatin and raltitrexed both are active anticancer agents in the treatment of patients with advanced colorectal carcinoma: They have different mechanisms of action and toxicity profiles and have shown at least additive effects in experimental and preliminary clinical studies. The aim of this disease oriented Phase I-II study was to determine the maximum tolerated dose (MTD), the dose-limiting toxicities (DLT), and the objective response rate of this combination in patients with advanced colorectal carcinoma. METHODS: Between April 1998 and March 1999, 69 patients with measurable metastatic colorectal carcinoma who previously were unexposed to palliative chemotherapy were enrolled. In the Phase I part of the study, 27 patients were treated with 3-weekly courses of a fixed dose of raltitrexed (3 mg/m(2) given as a 15-minute intravenous infusion) followed by a 2-hour infusion of oxaliplatin, which was escalated in consecutive cohorts of three to six patients from 85 mg/m(2) to 100 mg/m(2), 120 mg/m(2), 130 mg/m(2), and 140 mg/m(2). After having defined the toxic dose, 42 additional patients were entered at one dose level below to define the therapeutic index of this combination more precisely. RESULTS: In the Phase I part of the study, during the first three dose levels, only one patient each experienced DLT (Grade 3 increase in transaminases, diarrhea, and stomatitis); at level 4, two of the first six patients entered had Grade 3 neutropenic infection or peripheral neurotoxicity, whereas dose level 5 (oxaliplatin 140 mg/m(2)) constituted the toxic dose with three of three patients experiencing DLT (Grade 3 asthenia, transient amaurosis, and diarrhea with Grade 4 neutropenia). Externally reviewed objective responses were noted in 9 of these 27 patients (33%), and stable disease occurred in 12 patients (44.4%). Among the 42 patients who were treated subsequently at the MTD level (Phase II portion), 20 patients (47.6%) responded (95% confidence interval, 32-62.6%), and 21 patients (50%) had stable disease. Their median progression free survival was 9.0 months, and the median overall survival, with 42 patients (67%) currently alive, is > 14.5 months. Treatment tolerance at the MTD was acceptable, with only 9 of 42 patients (21%) experiencing Grade 3-4 neutropenia; Grade 3 nonhematologic adverse reactions included increase in serum transaminases in 6 patients, peripheral neuropathy in 3 patients, diarrhea in 3 patients, and both stomatitis and emesis in only 1 patient each. CONCLUSIONS: The described objective response and toxicity data, which are in agreement with preliminary results of other Phase I-II studies, support the promising therapeutic potential of this combination in the treatment of patients with advanced colorectal carcinoma. Due to its substantial antitumor activity, tolerance (at the recommended MTD level), and convenient 3-weekly outpatient administration schedule, further evaluation of this regimen seems warranted. PMID- 11283926 TI - Significant correlation between expression of interleukin-1alpha and liver metastasis in gastric carcinoma. AB - BACKGROUND: Interleukin (IL)-1 has been reported to augment the hematogeneous metastasis of some cancers by inducing the expression of adhesion molecules on vascular endothelial cells and also increasing the expression of proteases from tumor cells in vitro. The purpose of this study was to determine the clinical significance of the IL-1alpha expression in primary tumors of gastric carcinoma. METHODS: Paraffin embedded sections of the tumors obtained from 109 patients who underwent a gastrectomy for gastric carcinoma with subserosal invasion were stained for IL-1alpha by using the streptavidin biotin method. Staining was considered positive when 10% or more of the malignant cells displayed cytoplasmic staining. RESULTS: Sixty (55.0%) tumors expressed IL-1alpha. Positive staining for IL-1alpha was more likely in patients with differentiated tumors than in those with undifferentiated tumors. The expression of IL-1alpha showed a significant correlation with liver metastasis. Of the 84 patients receiving a curative resection, those with tumors expressing IL-1alpha had a significantly higher incidence of liver recurrence than those without. A logistic regression analysis revealed positive staining for IL-1alpha to be the best predictive factor of patients who develop liver recurrence. CONCLUSIONS: The authors' results suggest that the immunohistochemical expression of IL-1alpha is a useful predictor of liver recurrence in patients undergoing a curative resection for gastric carcinoma with subserosal invasion. PMID- 11283927 TI - Human macrophage metalloelastase gene expression in colorectal carcinoma and its clinicopathologic significance. AB - BACKGROUND: Human macrophage metalloelastase (HME), also referred as matrix metalloproteinase 12, has been shown to convert plasminogen into angiostatin, an essential inhibitor of tumor angiogenesis. The current study was designed to investigate the association of HME mRNA expression with disease progression of in patients with colorectal carcinoma. METHODS: The expression of HME mRNA was quantified by Northern blot analysis in tumorous (T) and nontumorous (N) tissues obtained from 54 patients with primary colorectal carcinoma, and the ratio of these two tissue types was defined as the HME T/N ratio. The prognostic significance of this ratio after surgery, along with its correlation with disease progression and metastatic potential, was evaluated. The tissues also were subjected to in situ hybridization analysis for HME. The microvessel count after immunohistochemical staining of factor VIII was used to assess angiogenesis. RESULTS: The HME T/N ratio showed an inverse correlation with the depth of the intestinal wall invasion, the lymphatic invasion, and the vascular invasion. The patients with overexpression of HME mRNA (HME T/N ratio > 1.191) significantly demonstrated better survival compared with those patients who did not show overexpression of HME mRNA. In situ hybridization identified the colorectal carcinoma cells as mainly responsible for the signal shown in Northern blot analysis. A considerable but not statistically significant lower microvessel density (MVD) was observed in the patients with HME overexpression. CONCLUSIONS: These findings demonstrate that the HME gene plays an important role in the inhibition of tumor progression in patients with colorectal carcinoma and that its overexpression is correlated closely with a better prognosis. PMID- 11283928 TI - Impact of second opinion pathology in the definitive management of patients with bladder carcinoma. AB - BACKGROUND: The accurate diagnosis, staging, and grading of bladder neoplasms depend heavily on the interpretation of biopsies and transurethral resection (TUR) specimens. Although many centers require review of outside pathologic material before definitive treatment such as radical cystectomy, the authors are unaware of data supporting the utility of this approach in urothelial (transitional cell) carcinoma. The authors therefore examined the clinical and cost impact of pathologic review on patients referred to an academic urology department for treatment of bladder neoplasia. METHODS: The pathologic material from 97 patients referred to an academic center for evaluation of urothelial carcinoma of the bladder from July 1996 to July 1999 was reviewed. This material was received from 30 community hospitals and 4 academic centers. The 97 patients had undergone 131 (mean, 1.35; range, 1-10) biopsies or TUR procedures before referral. Surgical pathologists at the authors' institution reviewed all outside patient material, and discordant cases were rereviewed by one of the authors (S.E.M), an experienced genitourinary pathologist. Follow-up chart review was performed in discordant cases to determine clinical and pathologic outcomes. RESULTS: Upon review at the authors' institution, 24 of 131 (18%) specimens with a referring diagnosis of urothelial carcinoma exhibited significant discrepancies with regard to the diagnosis, stage, grade, or tumor histologic type made at the outside institution. Four tumors (3%) were found to be nonurothelial neoplasms. Five specimens (4%) were judged inadequate for staging because they contained no muscularis propria. Three patients were upstaged, including two patients shown to have muscle invasive disease. Eight patients were downstaged, including two patients referred with purported muscle invasive disease who were determined to have only superficial disease on pathology review. Two patients initially thought to have carcinoma in situ (tumor in situ [Tis]) showed no evidence of Tis on pathology review. One patient with purported muscle invasive disease was shown to have only metaplasia, and one patient had a highly significant change in tumor grade. As a result of the pathology review, five radical cystectomies were avoided, whereas five repeat TUR procedures were recommended for inadequate staging. One patient shown to have muscle invasion on pathology review proceeded directly to cystectomy, avoiding a planned repeat TUR. A cystectomy also was recommended to a second patient who was shown to have invasive disease by the pathology review. Pathology review of 131 specimens resulted in net savings of $86,176 or $658 per TUR reviewed. CONCLUSIONS: The review of bladder pathologic materials before definitive therapy can impact clinical decisions significantly and can reduce overall expenditures for the management of this cohort of bladder carcinoma patients. PMID- 11283929 TI - Prospective study of correlations between biopsy-detected high grade prostatic intraepithelial neoplasia, serum prostate specific antigen concentration, and race. AB - BACKGROUND: High grade prostatic intraepithelial neoplasia (HGPIN), a premalignant lesion of the prostate gland, is more common in black men than in white men. The influence of HGPIN on the serum prostate specific antigen (PSA) concentration is controversial, and correlations between HGPIN and PSA in black men and white men have not been investigated. METHODS: Between January 1992 and December 1998, 411 black men and 639 white men with suspected prostate carcinoma underwent an initial benign prostate biopsy at a single medical center. The presence or absence of HGPIN in the biopsy specimens was determined by one uropathologist. RESULTS: HGPIN was identified in 8.9% of the specimens. When stratified by PSA concentration (< 4.0 ng/mL, 4.0-9.9 ng/mL, and > or = 10.0 ng/mL), HGPIN was associated with an increased PSA concentration only among men with PSA concentrations < 4.0 ng/mL (P = 0.01). The prevalence of HGPIN in the black and white patients was 13.4% and 5.9%, respectively (P < 0.0001), and was significantly greater in black men than in white men with PSA concentrations < 4.0 ng/mL (P = 0.002). Among the patients with PSA concentrations < 4.0 ng/mL, black race was an independent predictor of an increased PSA concentration when adjusted for patient age, prostate volume, and the presence or absence of HGPIN (P = 0.03). CONCLUSIONS: HGPIN is more common in black men than in white men and may produce an increase in the PSA concentration. However, racial differences in the prevalence of HGPIN may not contribute to racial differences in PSA concentrations among men with no clinical or histologic evidence of carcinoma. PMID- 11283930 TI - The impact of chemotherapy on Leydig cell function in long term survivors of germ cell tumors. AB - BACKGROUND: Because patients with germ cell tumors expect an additional life span of around 50 years after successful treatment, attention is now focused on potential long term toxicity. Limited data are available on Leydig cell function in long term survivors. METHODS: The authors measured testosterone, sex hormone binding-globulin (SHBG), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) levels in 244 patients with germ cell tumors. Patients were divided into three groups: Group 1 had received no chemotherapy (n = 58 patients), Group 2 had received cumulative doses of cisplatin < or = 400 mg/m(2) (n = 117 patients), and Group 3 had received cumulative doses of cisplatin > 400 mg/m(2) (n = 69 patients). The median times from chemotherapy were 74 months and 75 months in Groups 2 and 3, respectively. RESULTS: Subnormal testosterone levels (< 10 nmol/L) were found in 5%, 11%, and 20% in Groups 1, 2, and 3, respectively (Group 1 vs. Group 3; P = 0.02). The mean testosterone level and the testosterone/SHBG ratio did not differ significantly between Groups 1 and 2; however, they did differ between Groups 1 and 3 (testosterone: 17.0 nmol/L vs. 14.9 nmol/L, respectively; P = 0.02; testosterone/SHBG ratio: 0.70 vs. 0.59; P < 0.05). There was a significant inverse correlation between the testosterone/SHBG ratio and LH (correlation coefficient [r] = -0.25; P = 0.0002). A significant positive correlation was found for LH and FSH (r = 0.78; P < 0.0001), indicating a strong association between Leydig cell dysfunction and germinal epithelial damage. CONCLUSIONS: Standard doses of cisplatin-based chemotherapy do not lead to a significant deterioration of Leydig cell function in long term survivors of germ cell tumors. In contrast, high cumulative doses of chemotherapy cause a significant and persistent impairment of Leydig cell function. More data are needed regarding the clinical relevance of moderate testosterone deficiency. Further research is necessary to determine whether some patients may benefit from testosterone replacement. PMID- 11283931 TI - Improved long term survival of patients with metastatic nonseminomatous testicular germ cell carcinoma in relation to prognostic classification systems during the cisplatin era. AB - BACKGROUND: The current study reviews chronologic changes in the long term outcome of patients with metastatic nonseminomatous testicular germ cell tumors (NSTGCT) who were treated at a single institution during the past two decades. The 10-year survival of prognostic subgroups according to the classification of the International Germ Cell Consensus Classification Group (IGCCCG) and various other prognostic classifications is examined in time to evaluate whether cumulative experience has led to an improved outcome of patients with metastatic NSTGCT and to explore differences in outcome of prognostic subgroups. METHODS: Two hundred ninety-nine patients with metastatic NSTGCT who were treated with cisplatin-based polychemotherapy during the period from 1977 to 1996 were staged retrospectively according to the Royal Marsden (RM) classification and the following prognostic classifications: IGCCCG, Indiana, Medical Research Council (MRC), and European Organization for Research and Treatment of Cancer (EORTC). The numbers of patients who were treated during the periods 1977-1986 and 1987 1996 were 146 and 153, respectively. Survival curves were constructed using the Kaplan-Meier method, and disease specific 10-year survival rates of prognostic subgroups treated during the two consecutive 10-year periods were compared using the log rank test. RESULTS: The median follow-up of surviving patients during the periods 1977-1986 and 1987-1996 was 14.7 years (range, 0.2-20.6 years) and 7.0 years (range, 0.4-11.4 years), respectively. The actuarial disease specific 10 year survival rate of patients with metastatic NSTGCT increased from 76% during the period 1977-1986 to 88% during the period 1987-1996 (relative risk [RR], 0.51; 95% confidence interval [95% CI], 0.29-0.89; P < 0.05). The 10-year survival rates of patients with good, intermediate, and poor prognoses according to the IGCCCG classification were 95%, 74%, and 37%, respectively, during the period 1977-1986 and 94%, 87%, and 66%, respectively, during the period 1987 1996. Patients with a poor prognosis according to the IGCCCG classification showed the greatest increase in 10-year survival (RR, 0.43; 95% CI, 0.18-1.04; P = 0.06). Analysis using the RM, Indiana, and EORTC classifications also showed an improved 10-year survival rate of patients with a poor prognosis who were treated during 1987-1996 compared with those who were treated during 1977-1986. CONCLUSIONS: The 10-year survival rate of patients with metastatic NSTGCT who were treated with cisplatin-based chemotherapy significantly increased from 76% during the period 1977-1986 to 88% during the period 1987-1996. This improvement during the cisplatin era resulted mainly from an increase in the survival of patients with metastatic disease who had a poor prognosis. These results indicate that the management of patients with NSTGCT is still improving. PMID- 11283932 TI - Phase II study of paclitaxel, ifosfamide, and carboplatin in patients with recurrent or metastatic head and neck squamous cell carcinoma. AB - BACKGROUND: In the current study the authors assessed the antitumor activity (including response rate, duration of response, and survival) and toxicity profile (including anorexia, fatigue, emesis, and peripheral neuropathy) of a combination of paclitaxel, ifosfamide, and carboplatin (TIC) in patients with recurrent or metastatic squamous cell carcinoma of the head and neck (SCCHN). The trial hypothesis was that the TIC therapeutic index would be as high as that of paclitaxel, ifosfamide, and cisplatin (TIP) in this setting, but with less toxicity. METHODS: Patients with recurrent or metastatic SCCHN were treated with 175 mg/m(2) of paclitaxel as a 3-hour infusion on Day 1, 1000 mg/m(2) of ifosfamide as a 2-hour infusion on Days 1-3, 600 mg/m(2) of mesna on Days 1-3, and carboplatin (area under the concentration-time curve of 6) as a 30-minute infusion on Day 1; the regimen was repeated every 3-4 weeks. All patients were premedicated with dexamethasone, diphenhydramine, and cimetidine before paclitaxel infusion. Prophylactic hematopoietic growth factors were not given. RESULTS: Among 56 patients entered onto the study, 55 patients were analyzed for survival rates (locoregional recurrence alone in 56% of patients and distant metastasis with or without locoregional recurrence in 44% of patients). Fifty four patients were evaluable for tumor response and toxicity. A total of 32 patients (59%) had disease that responded to treatment; the complete response rate was 17% (9 of 54 patients). The median duration of the responses was 3.7 months (95% confidence interval [95% CI], 3.4-7.8 months) and that of complete responses was 9.7 months (95% CI, 7.4 months to date of last follow-up). The median duration of follow-up care in all patients was 13.5 months. The median survival time for all patients was 9.1 months (95% CI, 7.9-12.2 months). The regimen was well tolerated. Neutropenic fever developed in 30% of the patients; 1 patient died of neutropenia and sepsis. Other toxic effects included Grade 2-3 anorexia in 13% of patients, Grade 2-3 weight loss in 11% of patients, Grade 2-3 fatigue in 27% of patients, Grade 2-3 nausea/emesis in 13% of patients, and Grade 2-3 peripheral neuropathy in 9% of patients (toxicity grading based on the National Cancer Institute's Common Toxicity Criteria). Red blood cell and platelet transfusions were required in 13% and 7% of patients, respectively. CONCLUSIONS: The TIC regimen had high antitumor activity in patients with recurrent or metastatic SCCHN, with a 59% major response rate (17% complete response rate with relatively durable complete responses). Neutropenic fever developed in 30% of the patients, the incidence of which might have been decreased by prophylactic antibiotics or hematopoietic growth factor support. Other toxic effects included significantly lower rates and less severe instances of anorexia, emesis, fatigue, and peripheral neuropathy than those reported with the previously studied TIP regimen. The TIC regimen currently is being studied as an induction chemotherapy regimen in previously untreated patients with locally advanced SCCHN. The activity of TIC (a novel paclitaxel and ifosfamide-based regimen) in patients with recurrent or metastatic SCCHN should be confirmed in a Phase III randomized trial. PMID- 11283933 TI - Association of trypsin expression with recurrence and poor prognosis in human esophageal squamous cell carcinoma. AB - BACKGROUND: Overexpression of the matrix serine proteinase (MSP) trypsin has been implicated in tumor growth, invasion, and metastasis. The objective of this study was to clarify the clinicopathologic and prognostic significance of trypsin expression in esophageal squamous cell carcinomas (SCC). METHODS: Production of trypsin in tissue extracts was analyzed by immunoblotting and gelatin zymography. The authors analyzed the association between immunohistochemically detected trypsin expression in esophageal SCC and clinicopathologic characteristics, and they investigated whether trypsin is a predictor of recurrence and/or survival. RESULTS: Overproduction and activation of trypsin was observed in 6 of 10 tumor extracts. The trypsin immunoreactivities at the invasive front were more intense than those at the superficial layer. Sections with immunostaining signals in greater than 30% of carcinoma cells at the invasive front, which were observed in 52 (52%) cases, were judged to be positive for trypsin. Trypsin positivity was significantly correlated with the depth of invasion (P < 0.0001), lymph node metastasis (P = 0.0048), advanced pTNM classification (P = 0.0006), recurrence (P = 0.0003), and recurrence within the first postoperative year (P = 0.0005). Patients with trypsin positive carcinoma had significantly shorter disease free and overall survival times than did those with trypsin negative carcinoma (P < 0.0001 and P < 0.0001, respectively). Trypsin retained its significant predictive value for disease free and overall survival in multivariate analysis that included conventional clinicopathologic factors (P = 0.0029 and P = 0.0006, respectively). Patients with concomitant overexpression of trypsin and matrilysin at the invasive front, in which they often were colocalized, had the worst prognosis. CONCLUSIONS: The authors' results suggest that trypsin plays a key role in the progression of esophageal carcinoma. Detection of trypsin expression as well as matrilysin is useful for the prediction of recurrence and poor prognosis. PMID- 11283934 TI - Differing rates of loss of DPC4 expression and of p53 overexpression among carcinomas of the proximal and distal bile ducts. AB - BACKGROUND: Biliary tract carcinomas are clinically heterogeneous. It is not known if molecular heterogeneity underlies the clinical differences. METHODS: The authors evaluated 128 bile duct carcinomas, 88 of the distal common bile duct and 40 of more proximal origin (28 perihilar carcinomas, 12 intrahepatic carcinomas), immunohistochemically for abnormalities in the expression of the products of the DPC4 and p53 tumor-suppressor genes. Prognostic factors were evaluated in the series of distal bile duct carcinomas for which follow-up information was available. RESULTS: The authors found that a significantly higher percentage of distal bile duct carcinomas (55%) demonstrated loss of DPC4 expression than did the proximal bile duct carcinomas (15%; P < 0.001). They also found that a significantly higher percentage of the distal tumors abnormally expressed the p53 gene product (51% vs. 26%; P < 0.001). Among the distal common bile duct carcinomas, the presence of poorly differentiated histology correlated with decreased survival in multivariate analysis, while labeling for p53 or Dpc4, margin status, lymph node status, and tumor dimension did not correlate significantly with survival. CONCLUSIONS: These results demonstrate that abnormalities in DPC4 and p53 gene expression are frequent in distal common bile duct carcinomas, just as they are in pancreatic ductal adenocarcinoma, suggesting that these two tumor types might share a similar molecular pathogenesis. They also show that proximal and distal bile duct carcinomas have different patterns of inactivation of tumor-suppressor genes, indicating that they often arise through different molecular mechanisms likely reflecting their differing etiologies. PMID- 11283935 TI - Percutaneous radiofrequency ablation therapy with combined angiography and computed tomography assistance for patients with hepatocellular carcinoma. AB - BACKGROUND: Radiofrequency ablation (RFA) for patients with hepatocellular carcinoma (HCC) has been reported previously. This technique is superior to percutaneous microwave coagulation therapy (PMCT) for the enlargement of the necrotic area. Therefore, a few treatment sessions of RFA for patients with small HCC lesions measuring < 3 cm in greatest dimension can achieve complete necrosis. To achieve this with a one-treatment RFA session, the authors designed the technique of RFA with angiography combined with computed tomography (angio-CT) assistance. The advantages of this technique are that it is possible to detect small satellite nodules and to evaluate the real-time therapeutic effect immediately after RFA. METHODS: Ten patients with 12 HCC lesions measuring < 4 cm in greatest dimension underwent RFA with angio-CT assistance. The authors performed standard RFA for six patients (seven tumors) and RFA with balloon occlusion of the hepatic artery (balloon-occluded RFA [BoRFA]) for four patients (five tumors). Final therapeutic efficacy was evaluated with dynamic CT scans performed 2 weeks after treatment. RESULTS: On CT arteriography (CTA) obtained immediately after treatment, a hyperattenuating ring around the nonenhanced region was apparent in all patients. On CT scans obtained 2 weeks after treatment, this ring disappeared, and the greatest dimension of the nonenhanced region was slightly larger than that on the CTA obtained immediately after treatment. The authors achieved complete eradication with one treatment session of RFA in 8 of 10 patients (80%). Local recurrence occurred in one patient 10 months after treatment. The greatest dimension of the area coagulated by BoRFA was significantly larger (greatest long-axis dimension, 38.2 +/- 2.8 mm; greatest short-axis dimension, 35.0 +/- 1.7 mm; n = 5 lesions) than without it (greatest long-axis dimension, 30.0 +/- 4.1 mm; greatest short-axis dimension, 27.0 +/- 4.3 mm; n = 4 lesions; greatest long-axis dimension, P = 0.009; greatest short-axis dimension, P = 0.006). No major complications occurred in any patient. CONCLUSIONS: The authors were able to achieve success with a single treatment session in patients with small HCC using RFA with angio-CT assistance. They consider that RFA with angio-CT assistance is a safe and effective technique for the treatment of patients with small HCC. PMID- 11283936 TI - Expression and prognostic significance of catalase in malignant mesothelioma. AB - BACKGROUND: Free radicals and antioxidant enzymes (AOEs) may play a critical role in cell proliferation and in the resistance of malignant cells against cytotoxic drugs and radiation. Malignant mesothelioma is a resistant tumor with high levels of manganese superoxide dismutase, a central superoxide scavenging AOE. In the current study, the authors assessed the expression and prognostic role of catalase, an important hydrogen peroxide scavenging AOE, in malignant pleural mesothelioma. METHODS: Catalase expression was investigated by immunohistochemistry in 5 cases of nonmalignant healthy pleura and in tumor tissue of 32 mesothelioma patients, and by Western blot in 7 continuous human mesothelioma cell lines. The distribution of catalase in mesothelioma cells was assessed by immunoelectron microscopy. Furthermore, to investigate the effect of catalase inhibition in the drug resistance of these cells in vitro, the authors exposed mesothelioma cells with the highest catalase level to epirubicin with and without aminotriazole pretreatment. RESULTS: Nonmalignant mesothelial cells showed no catalase immunoreactivity whereas most mesothelioma cases (24 of 32, 75%) were catalase positive, 17 cases (53%) showing moderate or high expression. Higher catalase expression in mesothelioma was associated with a better prognosis, mean survival rate from diagnosis being 6 and 24 months for negative/low expression and moderate/high expression, respectively. Furthermore, a coordinately high expression of both manganese-superoxide dismutase (Mn-SOD) and catalase predicted even more favorable outcome of the mesothelioma patients. Catalase also could be detected in all mesothelioma cell lines, the most resistant cell line showing the highest protein expression and compartmentalization of catalase mainly to peroxisomes. Aminotriazole inhibition of catalase had a marginal effect on the toxicity caused by epirubicin. CONCLUSIONS: Catalase may have multifactorial effects in malignant cells; high catalase and/or coordinated high expression of Mn-SOD and catalase may decrease tumor progression by modulating the cellular redox state, but enhanced antioxidant capacity of mesothelioma cells also may protect tumor cells against exogenous oxidants, at least in vitro. PMID- 11283937 TI - Second neoplasms in patients with Merkel cell carcinoma. AB - BACKGROUND: Merkel cell carcinoma (MCC) has been associated with a high incidence of other skin tumors and hematological malignancies. The purpose of this study was to analyze data from the Israel Cancer Registry regarding the incidence of second neoplasms in patients with MCC and their impact on survival. METHODS: Sixty-seven patients in whom MCC was diagnosed between 1983 and 1999 were included. Data were collected on age, gender and ethnic origin, dates of diagnosis of MCC and any other neoplasm, and date and cause of death, if applicable. Comparison of MCC-specific survival, estimated by the Kaplan-Meier product limit method, between patients with no other neoplasm and those with second primary tumors was performed by log rank test. Age-specific standardized incidence ratio (SIR) was calculated using 5751 age- and ethnic-matched malignant melanoma patients as a control group. RESULTS: Seventeen patients (25%) had a second neoplasm before, concomitant with, or after the diagnosis of MCC; 2 of them also had a third primary tumor. The SIR was 2.8 (95% CI; range, 1.38-4.22), significantly higher than the control group. Almost half the tumors were squamous cell carcinomas, either skin or head and neck, and most of the remainder were hematological malignancies or breast and ovarian adenocarcinomas. On univariate analysis, the presence of another neoplasm, regardless of its chronology, was associated with higher MCC-specific mortality (65% vs. 40% for patients with MCC only; P = 0.022). Analysis of only those patients in whom a second neoplasm developed during follow-up after treatment for MCC yielded an estimated actuarial risk of developing a second primary of 2.1% for each year of observation. CONCLUSIONS: There is a high incidence of second neoplasms, including noncutaneous solid tumors, in patients with MCC. The presence of these neoplasms, whether they appear before, after, or simultaneously with MCC, is associated with a higher MCC-specific mortality. PMID- 11283938 TI - Multiple primary malignancies in association with soft tissue sarcomas. AB - BACKGROUND: Modern cancer treatment has increased the survival of patients with various malignancies substantially. One of the late sequelae of successful treatment is the development of a second malignant tumor. However, in many cases of second primary tumors, exposure to chemotherapy or radiation therapy is not evident, and it should be postulated that the putative mechanism for the development of the second tumor is different. In the current series, the association between soft tissue sarcoma (STS) in adults and the development of other primary malignancies was studied. METHODS: A retrospective search of the data files of 610 patients with STS or bone sarcomas who were treated at the study institution between January 1995 and December 1999 was performed. All files regarding patients with STS who developed a second malignant tumor were retrieved for analysis. RESULTS: Of 375 patients with STS, 28 (7.5%) developed other malignant neoplasms either before or after the diagnosis of STS. STS as the first tumor occurred in 14 patients (ages 16-72 years). Only three patients were treated with chemotherapy for their sarcoma. Radiation therapy was administered to five patients as an adjuvant to surgery for the first tumor. The second tumor types mainly included STS and renal cell carcinoma. The time interval between the diagnosis of the STS and the second malignancy was 0 (for synchronous tumors) to 21 years. Three patients developed a third primary tumor within 3 years after the diagnosis of the second tumor. The median overall survival was > 78 months. Fourteen patients (ages 35-87 years) had a first primary tumor other than STS (mainly breast carcinoma and genitourinary malignancies). The second tumors (mainly STS) appeared within 0 (for synchronous tumors) to 27 years. The median overall survival for the 14 patients in this group from the time of diagnosis of the first tumor was > 102 months. CONCLUSIONS: The phenomenon of two or three primary neoplasms developing in patients in whom one of the tumors was STS occurs at a rate of 7.5%, a significantly higher rate than that reported for the occurrence of STS among the general cancer population (1%). The majority of cases occur incidentally. The clinical implication includes the need to search for an occult second primary tumor in patients with STS as an integral part of their follow-up. This is especially true in patients with primary malignant fibrous histiocytoma who demonstrate a risk for developing a renal cell carcinoma. PMID- 11283939 TI - Granulocyte colony-stimulating factor (G-CSF) receptor gene expression of ovarian carcinoma does not correlate with G-CSF caused cell proliferation. AB - BACKGROUND: Recombinant human granulocyte colony-stimulating factor (rhG-CSF) is a growth factor commonly used to avoid leukopenia after chemotherapy. Endogenous G-CSF is produced by macrophages and granulocytes that infiltrate tumors. It has been reported that rhG-CSF stimulates the proliferation of several cell lines as well as bladder carcinoma cells. Conversely, in some hematopoietic cell lines such as U-937, WEHI-3B, and K-562 no effect or in some cases a differentiation pattern was found. Moreover, the role of rhG-CSF on the proliferation of solid tumors is not well understood. METHODS: In this study, 10 ovarian carcinoma biopsies were characterized for the presence of G-CSF and G-CSF receptor by reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemical analysis. Proliferation was analyzed by ATP viability assays. RESULTS: Performing RT-PCR, these biopsies and four ovarian carcinoma cell lines were analyzed for endogenous G-CSF production, which was found in some biopsies and in all cell lines. Despite the presence of the G-CSF receptor in all biopsies and cell lines, no proliferation was found after rhG-CSF incubation of the cell lines or the tumor samples for 3 and for 6 days, respectively. CONCLUSIONS: Summarizing the authors' in vitro studies, rhG-CSF does not affect the proliferation of ovarian carcinoma cells in vitro. PMID- 11283941 TI - Alternative medicine use worldwide: the International Union Against Cancer survey. AB - BACKGROUND: In the current study, the authors attempted to surmount the deficiencies of previous surveys and elicit information regarding the use of alternative treatments of cancer worldwide. METHODS: The International Union Against Cancer (UICC), an international, nongovernment volunteer organization, E mailed a questionnaire concerning alternative therapy use to its members. RESULTS: A total of 83 responses from 33 countries were received. Descriptive analyses of this dataset were conducted, indicating the existence of a large and heterogeneous group of unproved remedies used to treat cancer in both developed and developing countries around the world. CONCLUSIONS: Improved public education concerning the importance of early medical attention and the value of documented cancer therapies, the wider availability of useful cancer treatments, and public policies that are sensitive to the patient's need to play a meaningful role in his or her own care are required. PMID- 11283940 TI - Lack of effectiveness of radiotherapy combined with cisplatin in patients with locally advanced pancreatic carcinoma. AB - BACKGROUND: Cisplatin has been reported to enhance the cell-killing effect of radiation. The current study was conducted to evaluate the efficacy and toxicity of radiotherapy combined with cisplatin in patients with locally advanced pancreatic carcinoma. METHODS: Forty-one patients with pancreatic carcinoma that was unresectable but confined to the pancreatic region were treated with external beam radiation (50.4 grays [Gy] in 28 fractions over 5.5 weeks) and daily cisplatin (5 mg/m(2)/day as a 30-minute infusion just before each radiation fraction). Maintenance 5-fluorouracil (5-FU) (500 mg/m(2)) given once weekly was initiated 1 week after the completion of the chemoradiotherapy and continued until disease progression or unacceptable toxicity. RESULTS: Of the 41 patients, 31 (76%) completed the scheduled course of chemoradiotherapy. The median survival time was 7.7 months, and the 1-year survival rate was 36%. The median progression free survival time was 5.8 months. The first site of failure was distant metastases in 25 patients, locoregional recurrence in 6 patients, and both sites in 1 patient. The major toxicity was leukocytopenia and nausea/emesis. CONCLUSIONS: Radiotherapy with daily cisplatin appears to be inferior to conventional chemoradiotherapy using 5-FU in patients with locally advanced pancreatic carcinoma. PMID- 11283942 TI - Extragonadal seminoma: an international multicenter analysis of prognostic factors and long term treatment outcome. AB - BACKGROUND: The objectives of this study were to evaluate the long term outcome of patients with extragonadal seminomatous germ cell tumors (GCT) so that prognostic variables for disease recurrence and patient survival could be identified and to access the efficacy of different treatment modalities. METHODS: Six hundred thirty-five patients with extragonadal GCT who were treated consecutively at 11 centers in the United States and Europe during the cisplatin based chemotherapy era between 1975 and 1996 were evaluated retrospectively. RESULTS: Fifty-two patients with primary retroperitoneal GCT (50%) and 51 patients with primary mediastinal GCT (49%) of pure seminomatous histology were identified (n = 1 patient with a primary cervical lymph node) representing 16.4% of 635 patients with extragonadal GCT who were included in the data base. The median age was 37 years (range, 18-70 years). Treatment consisted of platin-based chemotherapy in 77 patients (74%), radiotherapy in 9 patients (9%), and combined modality in 18 patients (17%). Ninety-two percent of patients (95% confidence interval, 87-97%) achieved a favorable response to primary therapy. After a median follow-up of 61 months (range, 1-211 months), 18 patients (17%) have had recurrent disease: 14% of those who received chemotherapy and 67% of those who received radiation therapy. The 5-year progression free survival rate favored the chemotherapy group, with 87% compared with 33% for irradiated patients (P = 0.006), whereas the overall survival rates were equal (90% vs. 67%; P = 0.13). No differences in overall survival or progression free survival were observed among patients with primary retroperitoneal and mediastinal seminoma. Prognostic factors that were identified to influence survival negatively were liver metastases (P = 0.01) and two or more metastatic sites (P = 0.04). CONCLUSIONS: In patients with extragonadal seminoma, a survival rate of > 90% at 5 years is achieved with adequate cisplatin-based chemotherapy. Compared with patients with nonseminomatous extragonadal GCT, no difference in long term survival exists between patients with primary retroperitoneal or mediastinal seminoma location. Primary radiotherapy seems to be associated with a significantly higher rate of disease recurrence, although most patients will be salvaged by subsequent chemotherapy. PMID- 11283944 TI - Juxtacellular labeling of identified neurons: kiss the cells and make them dye. PMID- 11283943 TI - Lymphoma and lymphoid leukemia incidence in Florida children: ethnic and racial distribution. AB - BACKGROUND: Incidence reports for pediatric lymphoma and lymphoid leukemia in Hispanic subpopulations in the United States are rare. The authors hypothesized that Florida's Hispanic children would have higher risks of lymphoma and lymphoid leukemia compared with non-Hispanic white children. METHODS: All cases of lymphoid leukemia, Hodgkin, non-Hodgkin, and Burkitt lymphoma (SEER International Classification of Diseases for Oncology codes) in children (< 15 years) in the Florida Cancer Data System (FCDS) from 1985 to 1997 were studied. Cases were classified as: 1) white, 2) Hispanic, or 3) black, and stratified by age. Age adjusted rates for the three race-ethnic groups were calculated. Rates for Hispanics and blacks were compared with whites as standardized rate ratios (SRR) with 95% confidence intervals. RESULTS: Seven hundred thirty-one incident cases of pediatric lymphoma and 1231 cases of lymphoid leukemia were identified during the study period. For children with lymphoma, the SRR for Hispanics was 1.32 (95% CI, 1.20-1.44), and for blacks, the SRR was 0.68 (95% CI, 0.63-0.72. For lymphoid leukemia, the SRR for Hispanics was 1.29 (95% CI, 1.28-1.30), and for blacks, the SRR was 0.55 (95% CI, 0.54-0.56). Similar rates were found for the Hodgkin and non-Hodgkin subgroups. CONCLUSIONS: Incidences of Hodgkin and non-Hodgkin lymphoma were significantly higher in Florida's Hispanic children, with 30% increased relative risks, compared with whites. Black children had significantly decreased incidences and risk. Results for lymphoid leukemia were similar. Incidence of lymphoma in Florida's Hispanic children (primarily Cuban and Central American origin) differed from similar reports from Texas and California, where Hispanics are primarily of Mexican origin. PMID- 11283945 TI - Expression of active caspase-3 in mitotic and postmitotic cells of the rat forebrain. AB - Active caspase-3 immunoreactivity was detected in the rat forebrain proliferative regions at birth and remained high in these areas for about 2 weeks, during which period labeled cells were present centroperipherally across the olfactory bulb. By the end of the third postnatal week, only a small number of immunolabeled cells remained in these forebrain structures. Active caspase-3 immunolabeling was localized mostly to cell nuclei and co-localized partially with TuJ1 and NeuN immunoreactivity, but not with glial fibrially acidic protein, OX-42, gamma aminobutyric acid, or terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL)-positive labeling. Active caspase-3 and 5-bromo-2'-deoxyuridine (BrdU) double-labeled nuclei were seen in the proliferative regions after 2 hours and in the periglomerular region of the bulb after 7 days following BrdU injections. Examination of the cells with electron microscopy confirmed that the active caspase-3-containing nuclei in the proliferative regions often had infoldings and appeared to be undergoing division. Some of the cells with active caspase-3-labeled nuclei in the bulb had synapses on their somata or dendrites. Labeled dendritic spines and a few axon terminals were also observed in the olfactory bulb. Taken together, it appears that a wave of active caspase-3 positive cells are dividing in the proliferative zones and then migrating to the bulb as they differentiate into neurons. Therefore, active caspase-3 may play a role in cellular processes such as neuronal differentiation, migration, and plasticity, in addition to its role in cell death. PMID- 11283947 TI - Mu-opioid receptors are present in functionally identified sympathoexcitatory neurons in the rat rostral ventrolateral medulla. AB - Agonists of the mu-opioid receptor (MOR) produce profound hypotension and sympathoinhibition when microinjected into the rostral ventrolateral medulla (RVL). These effects are likely to be mediated by the inhibition of adrenergic and other presympathetic vasomotor neurons located in the RVL. The present ultrastructural studies were designed to determine whether these vasomotor neurons, or their afferents, contain MORs. RVL bulbospinal barosensitive neurons were recorded in anesthetized rats and filled individually with biotinamide by using a juxtacellular labeling method. Biotinamide was visualized by using a peroxidase method and MOR was identified by using immunogold localization of an antipeptide antibody that recognizes the cloned MOR, MOR1. The subcellular relationship of MOR1 to RVL neurons with fast- or slow-conducting spinal axons was examined by electron microscopy. Fast- and slow-conducting cells were not morphologically distinguishable. Immunogold-labeling for MOR1 was found in all RVL bulbospinal barosensitive neurons examined (9 of 9). MOR1 was present in 52% of the dendrites from both types of cells and in approximately half of these dendrites the MOR1 was at nonsynaptic plasmalemmal sites. A smaller portion of biotinamide-labeled dendrites (16%) from both types of cells were contacted by MOR1-containing axons or axon terminals. Together, these results suggest that MOR agonists can directly influence the activity of all types of RVL sympathoexcitatory neurons and that MOR agonists may also influence the activity of afferent inputs to these cells. The heterogenous distribution of MORs within individual RVL neurons indicates that the receptor is selectively targeted to specific pre- and postsynaptic sites. PMID- 11283946 TI - Selective distribution of mu-opioid receptors in C1 adrenergic neurons and their afferents. AB - Agonists of the mu-opioid receptor (MOR) have profound effects on blood pressure, heart rate, and respiration that may be mediated by C1 adrenergic neurons in the rostral ventrolateral medulla (RVL). C1 neurons are sympathoexcitatory and are involved in both tonic and reflex regulation of sympathetic outflow. This study was designed to determine whether C1 neurons, or their afferents, contain MOR. C1 neurons were identified by using an antibody against the epinephrine synthesizing enzyme phenylethanolamine-N-methyl transferase (PNMT), whereas MOR was localized by using an antipeptide antibody that recognizes the cloned MOR, MOR1. Combined immunoperoxidase and immunogold methods were used to examine the cellular distribution of MOR1 relative to PNMT-containing neurons in the RVL. MOR1 was found in 22% of PNMT-containing dendrites (n = 392), whereas MOR1-containing axons or axon terminals contacted 14% of PNMT-containing dendrites. This distribution was heterogenous with regard to dendritic size: PNMT-labeled dendrites containing MOR1 were usually large (60% were >1.2 microm), whereas PNMT containing dendrites that received MOR1-labeled afferents were usually small (79% were <1.2 microm). Individual dendrites rarely contained MOR1 at both pre- and postsynaptic sites. Together these results suggest that MOR agonists may directly influence the activity of C1 neurons, as well as the activity of select afferents to these cells. Plasmalemmal membrane labeling for MOR1 was more frequent in smaller PNMT-containing dendrites, suggesting that postsynaptic receptors are more readily available for ligand binding in small dendrites, although the receptor was more frequently detected in larger PNMT dendrites. The selective distribution of MORs to specific pre- and postsynaptic sites suggests the receptor may be selectively trafficked to positions where it may regulate afferent activity that is heterogeneously distributed along the dendritic tree of C1 neurons. PMID- 11283948 TI - Peripheral patterns of terminal innervation of vestibular primary afferent neurons projecting to the vestibulocerebellum in the gerbil. AB - Retrograde transganglionic labeling techniques with biotinylated dextran amine (BDA) were used to examine the terminal field structure and topographical patterns of innervation within the vestibular sensory end organs of vestibular primary afferent neurons projecting to the cerebellar uvula/nodulus and flocculus lobules in the gerbil. Robust, dark labeling in the cristae ampullares suggested that the vast majority of the terminals of afferent neurons were of the dimorphic type. The majority (94% to the uvula/nodulus and 100% to the flocculus) innervates the peripheral zones of each of the three semicircular canal cristae. Comparison of the type and distribution of terminals across the canalicular sensory neuroepithelium with morphophysiological studies in chinchilla suggests that the labeled population consists predominantly of peripheral terminal fields of lower-to-intermediate gain, more regularly firing, tonic afferents. For otolith organ-related afferents, the uvula/nodulus receives strong inputs from primary otolith afferent neurons identified as dimorphic in type that predominately innervate the peristriolar zones of the utricular and saccular maculae. No direct otolith organ-related inputs to the flocculus were observed. In contrast to the canal afferents, the types and locations of labeled otolith afferent terminals suggest that they largely consist of irregularly firing, high gain, phasic neurons. PMID- 11283949 TI - Structural abnormalities develop in the brain after ablation of the gene encoding nonmuscle myosin II-B heavy chain. AB - Ablation of nonmuscle myosin heavy chain II-B (NMHC-B) in mice results in severe hydrocephalus with enlargement of the lateral and third ventricles. All B(-)/B(-) mice died either during embryonic development or on the day of birth (PO). Neurons cultured from superior cervical ganglia of B(-)/B(-) mice between embryonic day (E) 18 and P0 showed decreased rates of neurite outgrowth, and their growth cones had a distinctive narrow morphology compared with those from normal mice. Serial sections of E12.5, E13.5, and E15 mouse brains identified developmental defects in the ventricular neuroepithelium. On E12.5, disruption of the coherent ventricular surface and disordered cell migration of neuroepithelial and differentiated cells were seen at various points in the ventricular walls. These abnormalities resulted in the formation of rosettes in various regions of the brain and spinal cord. On E13.5 and E15, disruption of the ventricular surface and aberrant protrusions of neural cells into the ventricles became more prominent. By E18.5 and P0, the defects in cells lining the ventricular wall resulted in an obstructive hydrocephalus due to stenosis or occlusion of the third ventricle and cerebral aqueduct. These defects may be caused by abnormalities in the cell adhesive properties of neuroepithelial cells and suggest that NMHC-B is essential for both early and late developmental processes in the mammalian brain. PMID- 11283950 TI - Cellular and subcellular distribution of NMDA receptor subunit NR2B in the retina. AB - Immunocytochemical studies showed the presence of staining for the N-methyl-D aspartate (NMDA)-R2B glutamate receptor subunit at multiple sites in the cat retina. Reaction product in photoreceptor cells was localized at the inner/outer segment junction and in the axon terminals. Staining within the inner retina was limited to ganglion cells and their dendrites ramifying throughout the inner plexiform layer. These cells were seen to receive synaptic input from cone bipolar cells in both sublaminae. As with other glutamate receptor subunits, this immunoreactivity was typically confined to a single postsynaptic element at a cone bipolar dyad complex. Immunocytochemical localization of the NMDA-R1 subunit, considered to be an essential component of functional receptors, showed a widespread distribution across the retina including all the sites where NMDA R2B staining was seen. Immunoprecipitation and Western blot analysis were used to confirm the presence of the NR2B receptor protein and its association with the NR1 subunit in both proximal and distal retinal layers. The findings suggest that NMDA-R2B subunits are positioned for multiple functions within the retina. PMID- 11283951 TI - ErbB transmembrane tyrosine kinase receptors are differentially expressed throughout the adult rat central nervous system. AB - The neuregulin (NRG) family of growth and differentiation factors and their erbB receptors contribute importantly to the development of the nervous system, but their distribution and function in the adult brain are poorly understood. The present study showed that erbB2, erbB3, and erbB4 transcripts and protein are distributed throughout all areas of adult rat brain. These three receptors were differentially expressed in neurons and glia. Some neurons expressed only a subset of erbB kinases, whereas other neurons expressed all three erbB receptors but sequestered each of these polypeptides into distinct cellular compartments. In synapse-rich regions, erbB immunoreactivity appeared as punctate-, axon-, and/or dendrite-associated staining, suggesting that NRGs are involved in the formation and maintenance of synapses in adult brain. ErbB labeling also was present in neuronal soma, indicating that NRGs act at sites in addition to the synapse. Glia in adult brain also differentially expressed erbB3 and erbB4. Approximately half of the erbB3 labeling in white matter was associated with S100beta+/glial fibrillary acidic protein negative macroglia (i.e., oligodendrocytes or glial fibrillary acidic protein negative astrocytes). In contrast, macroglia in gray matter did not express erbB3. The remaining erbB3 immunoreactivity in white matter and erbB4 glial staining seemed to be associated with microglia. These results showed that erbB receptors are expressed widely in adult rat brain and that each erbB receptor subtype has a distinct distribution. The differential distributions of erbB receptors in neurons and glia and the known functional differences between these kinases suggest that NRGs have distinct effects on these cells. The continued expression of NRGs and their erbB receptors in mature brain also implies that these molecules perform important functions in the brain throughout life. PMID- 11283952 TI - Differential distribution of the glutamate transporters GLT-1 and GLAST in tanycytes of the third ventricle. AB - The ventral one-third of the ventricular lining in the hypothalamus is formed by specialized ependymal cells called the tanycytes. These cells may serve a neuroendocrine transport function because of their structural specializations, which include apical microvili on the ventricular surface and long basal processes that terminate on blood vessels or on the glia limitans. Here, we describe the expression of mRNA and protein for the glutamate transporters GLT-1 and GLAST in unique tanycyte populations of the third ventricle in rat brain. Using nonisotopic in situ hybridization, we demonstrate GLAST mRNA labeling in tanycytes of the ventral floor and lateral walls in the tuberal and mammillary recess portions of the third ventricle. This GLAST mRNA labeling had a higher intensity than the labeling intensity observed in regular ependymal cells throughout the ventricular system. Furthermore, we have identified strong GLT-1 mRNA labeling in a population of tanycytes situated in the dorsolateral walls of caudal tuberal and mammillary recess portions. Immunocytochemical staining indicates that both GLT-1 and GLAST protein are expressed in the tanycyte populations as well. These data corroborate previous findings that third ventricle tanycytes are functionally heterogeneous. Furthermore, the GLT-1 expressing tanycytes represent a population of tanycytes that, to date, has not been recognized as functionally distinct. The strong GLAST expression by the ventral tanycytes in the hypophysiotropic area suggests a role of tanycyte mediated glutamate transport in neuroendocrine activity. The functional role of GLT-1 in dorsal wall tanycytes remains to be explored. PMID- 11283953 TI - Reactive astrocytes express estrogen receptors in the injured primate brain. AB - Previous studies have suggested that estrogen may regulate the expression of genes related to the inflammatory response within the nervous system, particularly within glia. In the present study, we examined whether injury induces estrogen sensitivity in reactive glia in the primate brain. Three adult Macaca fascicularis (cynomolgous) monkeys received unilateral fimbria fornix transections followed by chronic intracranial cannula implants through which a vehicle solution was infused intracerebroventricularly for a 4-week period. Astrocytes adjacent to areas of parenchymal disruption caused either by the lesion or by the instrumentation procedure became reactive, as evidenced by cellular hypertrophy and up-regulation of glial fibrillary acidic protein (GFAP) immunolabeling. Of note, specific estrogen receptor-alpha immunolabeling also was induced adjacent to injured regions, and this labeling strictly colocalized with GFAP immunoreactivity upon double fluorescent confocal immunolabeling. Induction of estrogen receptor immunoreactivity in reactive astrocytes occurred in all monkeys examined, whereas nonreactive glia distant from disrupted regions did not exhibit estrogen receptor labeling. Thus, expression of estrogen receptors is up regulated in reactive astrocytes of the primate brain, potentially allowing estrogen to modulate aspects of the central nervous system's inflammatory response to injury. PMID- 11283954 TI - Neuronal turnover in the Xenopus laevis olfactory epithelium during metamorphosis. AB - Metamorphic changes in the amphibian olfactory system present many interesting questions concerning the competing possibilities of neuronal respecification versus replacement. For example, are olfactory neurons retained during this transition with their presumed sensitivity to waterborne versus airborne stimuli respecified, or are olfactory neurons completely replaced? We address this question using the African clawed frog (Xenopus laevis) as a model. The water sensing nose (principal cavity; PC) of larval X. laevis is respecified into an air-sensing cavity in adults, with changes in odorant receptor gene expression, ultrastructure, and site of innervation of the receptor neurons. The vomeronasal organ (VNO) does not appear to change function, structure, or innervation during metamorphosis. We labeled PC and VNO olfactory receptor neurons with injections of retrogradely transported fluorescent microspheres into the main and accessory olfactory bulbs. Injections were performed in larvae, and animals were allowed to survive through metamorphosis. After metamorphosis, few labeled cells were observed in the PC, whereas the VNO and the olfactory bulbs remained heavily labeled. Animals that were killed before metamorphosis always had extensive label in the PC epithelium regardless of how long the beads were present. This suggests that changes in the PC olfactory epithelium that are seen during metamorphosis are due primarily to turnover of the neurons in this epithelium rather than to respecification of existing neurons. These results also are discussed in terms of natural turnover time of olfactory receptor neurons. PMID- 11283955 TI - Regenerating descending axons preferentially reroute to the gray matter in the presence of a general macrophage/microglial reaction caudal to a spinal transection in adult zebrafish. AB - We analyzed pathway choices of regenerating, mostly supraspinal, descending axons in the spinal cord of adult zebrafish and the cellular changes in the spinal cord caudal to a lesion site after complete spinal transection. Anterograde tracing (by application of the tracer rostral to the spinal lesion site) showed that significantly more descending axons (74%) regenerated in the spinal gray matter of the caudal spinal cord than would be expected from random growth. Retrograde tracing (by application of the tracer caudal to the spinal lesion site) showed that, rostral to the lesion, most of these axons (80%) extended into the major white matter tracts. Thus, ventral descending tracts often were devoid of labeled axons caudal to a spinal lesion but contained many axons rostral to the lesion in the same animals, indicating a pathway switch of descending axons from the white matter to the gray matter. Ascending axons of spinal neurons were not observed regrowing to the rostral tracer application site; therefore, they most likely did not contribute to the axonal populations analyzed. A macrophage/microglia response within 2 days of spinal cord transection, along with phagocytosis of myelin, was observed caudal to the transection by immunohistochemistry and electron microscopy. Nevertheless, caudal to the lesion, descending tracts in the white matter were filled with myelin debris during the time of axonal regrowth, at least up to 6 weeks postlesion. We suggest that the spontaneous regeneration of axons of supraspinal origin after spinal cord transection in adult zebrafish may be due in part to the axons' ability to negotiate novel pathways in the spinal cord gray matter. PMID- 11283956 TI - Pyramidal cell axons show a local specialization for GABA and 5-HT inputs in monkey and human cerebral cortex. AB - Various mechanisms are thought to control excitation of pyramidal cells of the cerebral cortex. With immunocytochemical methods, we found that the proximal portions of numerous pyramidal cell axons (Pyr-axons) in the human and monkey neocortex are immunoreactive for the serotonin (5-HT) receptor 5-HT-(1A). With double-labeling experiments and confocal laser microscopy, we found that most (93.4%) of the 5-HT(1A)-immunoreactive Pyr-axons present in layers II and III were innervated by parvalbumin-immunoreactive chandelier cell axon terminals. In addition, Pyr-axons were compartmentalized: 5-HT-(1A) receptors were found proximal to inputs from chandelier cells. Although we found close appositions between GABAergic chandelier cell axon terminals and Pyr-axons, suggesting synaptic connections, we did not observe 5-HT-immunoreactive fibers in close proximity to the Pyr-axons. These results suggested that Pyr-axons are under the influence of 5-HT in a paracrine manner (via 5-HT-(1A) receptors) and, more distally, are under the influence of gamma-aminobutyric acid (GABA) in a synaptic manner (through the axons of chandelier cells). The local axonal specialization might represent a powerful inhibitory mechanism by which the responses of large populations of pyramidal cells can be globally controlled by subcortical serotonin afferents, in addition to local inputs from GABAergic interneurons. PMID- 11283957 TI - Anatomical distribution of serotonin-containing neurons and axons in the central nervous system of the cat. AB - By using a monoclonal antibody to serotonin (5-HT), an immunohistochemical study was undertaken to provide a comprehensive description of the 5-HT-containing neurons and of the distribution of their axonal processes in the cat brain and spinal cord. The localization of cell bodies was comparable to that previously reported in studies using formaldehyde-induced fluorescence and other 5-HT antibodies, with a large proportion of labeled neurons in the raphe nuclei and a minor, yet not negligible number, in the ventral, lateral, and dorsal reticular formation. The ascending efferent non-varicose axons were best visualized in sagittal sections and mainly seen taking a rostroventral direction through the tegmentum. The varicose axons could be grossly classified into thin and large fibers, according to the size and shape of the immunoreactive varicosities, which were elongated (up to 2 microm in length and 1 microm in width) or round (2-4 microm in diameter). Varicose axonal arborizations invaded almost every region of the gray matter and avoided large myelinated bundles except in the spinal cord. Variations in the density of the plexuses of immunoreactive fibers generally followed the anatomical divisions and were also observed within nuclei, especially in laminated structures. Only the superior olivary complex could be regarded as devoid of 5-HT-containing axons. A few areas contained extremely rich fiber plexuses. These were the olfactory tubercle, nucleus accumbens, ventral mesencephalon, periventricular gray from the hypothalamus to the pons, facial nucleus, subdivisions of the inferior olive, and the intermediolateral nucleus in the spinal cord. Varicose axons formed tight pericellular arrays in the neocortex, mainly the ectosylvian gyrus, and in the lateral septum and medullar magnocellular nucleus. These data, combined with those of the literature concerning the synaptic versus non-synaptic mode of termination of the 5-HT immunoreactive varicosities and the high number of distinct receptors, are indicative of the multiple possible actions of serotonin in the central nervous system. PMID- 11283958 TI - Distribution and ultrastructure of tachykinin-like immunoreactivity in the frog (Rana esculenta) spinal cord, notably, the dorsal horn. AB - Tachykinins are involved in pain transmission at the spinal level. In frog, at least four tachykinins [TK] have been isolated from the brain, but their organization in the dorsal horn of the spinal cord is still poorly known. We have reexamined TK distribution by immunocytochemistry using an antibody recognizing the sequence common to all tachykinins in the spinal cord and dorsal root ganglia of the green frog Rana esculenta. A dense tachykinin-like immunoreactivity (TK LI) was observed in the dorsolateral fasciculus or Lissauer's tract running ventromedial to the entry of the dorsal root and in numerous small and medium sized dorsal root ganglion cells showing a primary afferent origin for part of TK LI of the dorsal horn. The observation of numerous cell bodies in the dorsal horn, in addition, suggested a local or propriospinal origin. One group of cells was localized at the entrance of the Lissauer's tract TK-LI fibers into the dorsal horn, and another group was localized in the upper dorsal horn, a region with a low density of TK-LI fibers. It was suggested that the latter group may correspond to neurokinin B. Electron microscopic examination of the Lissauer's tract showed numerous immunoreactive axons, some located at the center of glomerular-like arrangements, suggesting that the information brought by these fibers may be transmitted and most probably modulated before their entry in the dorsal horn. In conclusion, the functional organization of tachykinins in the frog spinal cord seems to be similar to that of mammals, albeit with a different morphological organization. PMID- 11283959 TI - Subcellular localization of alpha-2A-adrenergic receptors in the rat medial nucleus tractus solitarius: regional targeting and relationship with catecholamine neurons. AB - alpha-2A-adrenergic receptor (alpha2A-AR) agonists modulate diverse autonomic functions. These actions are believed to involve functionally specialized, second order neurons in catecholamine-containing portions of the medial nucleus tractus solitarius (mNTS) at both intermediate (NTSi) and caudal (NTSc) levels. However, the cellular mechanisms subserving alpha2A-AR-mediated actions within the mNTS have yet to be established. Immunocytochemistry was employed to examine the subcellular distribution of alpha2A-AR in both the intermediate and caudal mNTS and its association with cells containing the catecholamine-synthesizing enzyme tyrosine hydroxylase (TH). Quantitative regional comparison using immunogold showed that this receptor was distributed differentially to dendrites (NTSi, 46%; NTSc, 31%) and glia (NTSi, 29%; NTSc, 48%) at different levels of the NTS. Somata, axons, and terminals less frequently contained alpha2A-AR. The subcellular distribution of alpha2A-AR relative to catecholaminergic neurons also was similar within both subregions. Approximately 50% of alpha2A-AR-labeled somata also contained TH. In somatic profiles, alpha2A-AR labeling was often found in the cytosol and in association with endoplasmic reticulum and Golgi complexes, sites of receptor synthesis and trafficking. Approximately 20% of alpha2A-AR-immunoreactive dendrites also contained TH, where the receptor was often found on extrasynaptic portions of the plasma membrane near unlabeled terminals, some of which made symmetric contacts. However, TH-labeled terminals and dendrites usually were detected in the neuropil at a short distance (<10 microm) from alpha2A-AR-labeled neurons. alpha2A-AR-labeled glia frequently apposed unlabeled dendrites and terminals and were often located near TH immunoreactive dendrites. These results indicate that, within the mNTS, alpha2A AR is involved in a variety of autonomic processes, including postsynaptic modulation of mostly noncatecholaminergic dendrites, as well as influencing glia functions. PMID- 11283960 TI - Zinc co-localizes with GABA and glycine in synapses in the lamprey spinal cord. AB - The presence of zinc in synaptic terminals in the lamprey spinal cord was examined utilizing a modification of the Timm's sulfide silver method and with the fluorescent marker 6-methoxy-8-quinolyl-p-toluenesulfonamide (TSQ). Axons labeled with a Timm's staining method were predominantly located in the lateral region of the dorsal column. This correlated with a maximum of TSQ fluorescence in this region of the spinal cord. Single labeled terminals accumulating Timm reaction product were also found throughout the gray matter and fiber tracts. At the ultrastructural level, zinc was located in a population of synaptic terminals that co-localized gamma-aminobutyric acid (GABA) and glycine. Possible effects of Zn2+ on neuronal activity were examined. In spinobulbar interneurons, which receive GABAergic input in the dorsal column, zinc potentiated responses to GABA application, but it did not affect responses to GABA in motoneurons. Responses in motoneurons to pressure application of glycine were also not affected by Zn2+. Zinc, however, potentiated monosynaptic glycinergic inhibitory postsynaptic potentials (IPSPs) evoked in motoneurons by inhibitory locomotor network interneurons and increased frequency, but not amplitude of spontaneous miniature IPSPs recorded in the presence of tetrodotoxin (TTX), suggesting presynaptic effects. Glutamate responses and the amplitude of monosynaptic excitatory postsynaptic potentials (EPSPs) in motoneurons were reduced by zinc. These effects appeared to be mediated largely postsynaptically through an effect on the N-methyl-D-aspartate (NMDA) component of the glutamatergic input. Our results thus show that free zinc is present in inhibitory synaptic terminals in the lamprey spinal cord, and that it may function as a modulator of inhibitory synaptic transmission. PMID- 11283961 TI - Neuropeptide expression in rat paraventricular hypothalamic neurons that project to the spinal cord. AB - The paraventricular hypothalamic nucleus (PVH) exerts many of its regulatory functions through projections to spinal cord neurons that control autonomic and sensory functions. By using in situ hybridization histochemistry in combination with retrograde tract tracing, we analyzed the peptide expression among neurons in the rat PVH that send axons to the spinal cord. Projection neurons were labeled by immunohistochemical detection of retrogradely transported cholera toxin subunit B, and radiolabeled long riboprobes were used to identify neurons containing dynorphin, enkephalin, or oxytocin mRNA. Of the spinally projecting neurons in the PVH, approximately 40% expressed dynorphin mRNA, 40% expressed oxytocin mRNA, and 20% expressed enkephalin mRNA. Taken together with our previous findings on the distribution of vasopressin-expressing neurons in the PVH (Hallbeck and Blomqvist [1999] J. Comp. Neurol. 411:201-211), the results demonstrated that the different PVH subdivisions display distinct peptide expression patterns among the spinal cord-projecting neurons. Thus, the lateral parvocellular subdivision contained large numbers of spinal cord-projecting neurons that express any of the four investigated peptides, whereas the ventral part of the medial parvocellular subdivision displayed a strong preponderance for dynorphin- and vasopressin-expressing cells. The dorsal parvocellular subdivision almost exclusively contained dynorphin- and oxytocin-expressing spinal cord projecting neurons. This parcellation of the peptide-expressing neurons suggested a functional diversity among the spinal cord-projecting subdivisions of the PVH that provide an anatomic basis for its various and distinct influences on autonomic and sensory processing at the spinal level. PMID- 11283962 TI - Expression of SCG10 and stathmin proteins in the rat olfactory system during development and axonal regeneration. AB - The membrane-associated protein SCG10 is expressed specifically by neuronal cells. Recent experiments have suggested that it promotes neurite outgrowth by increasing microtubule dynamics in growth cones. SCG10 is related to the ubiquitous but neuron-enriched cytosolic protein stathmin. To better understand the role played by SCG10 and stathmin in vivo, we have analyzed the expression and localization of these proteins in both the olfactory epithelium and the olfactory bulb in developing and adult rats, as well as in adult bulbectomized rats. The olfactory epithelium is exceptional in that olfactory receptor neurons constantly regenerate and reinnervate the olfactory bulb throughout animal life span. SCG10 and stathmin expression in the olfactory receptor neurons was found to be regulated during embryonic and postnatal development and to correlate with neuronal maturation. Whereas SCG10 expression was restricted to immature olfactory receptor neurons (GAP-43-positive, olfactory marker protein-negative), stathmin was also expressed by the basal cells. In the olfactory bulb of postnatal and adult rats, a moderate to strong SCG10 immunoreactivity was present in the olfactory nerve layer, whereas no labeling was detected in the glomerular layer. Olfactory glomeruli also showed no apparent immunoreactivity for several cytoskeletal proteins such as tubulin and microtubule-associated proteins. In unilaterally bulbectomized rats, SCG10 and stathmin were seen to be up-regulated in the regenerating olfactory epithelium at postsurgery stages corresponding to olfactory axon regeneration. Our data strongly suggest that, in vivo, both SCG10 and stathmin may play a role in axonal outgrowth during ontogenesis as well as during axonal regeneration. PMID- 11283963 TI - Morphology and physiology of neurons in the ventral nucleus of the lateral lemniscus in rat brain slices. AB - The ventral nucleus of the lateral lemniscus (VNLL) is a prominent neuronal group that lies within the auditory pathway connecting the auditory lower brainstem and midbrain. Previous physiologic studies showed that VNLL neurons respond mainly to contralaterally presented sounds and display various firing patterns. To understand better the role that VNLL neurons play in transmitting and processing of auditory information, we examined the morphology of VNLL neurons and their cellular physiology in young rat brain slices. We made whole-cell patch-clamp recordings and labeled cells intracellularly with neurobiotin to investigate the relation between morphologic neuronal types, intrinsic membrane properties, and postsynaptic responses. VNLL neurons fell into two distinct morphologic groups, i.e., bushy cells and stellate cells, based on their dendritic patterns. Stellate cells were grouped further into stellate I, II, and elongate cells according to soma shape, dendritic branches, and orientation. Bushy cells showed an onset firing pattern and a nonlinear current-voltage relationship. All three subtypes of stellate cells had a linear current-voltage relationship, but exhibited different firing patterns. Stellate I cells showed regular and onset-pause firing patterns, whereas stellate II cells showed adapting and elongate cells showed burst firing patterns. Bushy cells and stellate cells responded to stimulation of the lateral lemniscus with excitatory and/or inhibitory synaptic potentials. These results suggest that the VNLL is a heterogeneous neuronal group and that it contains many channels for processing different kinds of auditory information. Neuronal morphology and intrinsic membrane properties contribute to the behavior of individual neurons. PMID- 11283964 TI - Sensitivity to naloxone of the behavioral signs of morphine withdrawal and c-Fos expression in the rat CNS: a quantitative dose-response analysis. AB - Several studies have used c-Fos expression to delineate the neural substrate underlying naloxone-precipitated morphine withdrawal (MW). However, because behavioral manifestations of MW depend on both the degree of dependence and the doses of naloxone (NAL), a comprehensive study would require examining c-Fos expression in relation with the degree of MW. Here, changes in behavior and in c Fos-like immunoreactivity (FLI) were studied in the same rats after injection of three doses of NAL to precipitate various degrees of MW. Fifteen established signs of MW were examined for 1 hour after NAL injection, and FLI was quantified in 52 regions of the brain and in the lumbosacral spinal cord. Linear regression analyses were used to examine changes in numbers of signs and FLI neurons with the doses of NAL, and data were considered dose-related for a statistical level of significance of P < 0.05. In summary, autonomic signs of MW increased in a dose-related manner, whereas somatomotor signs did not. After MW, 33 central nervous system regions exhibited significant increases in FLI and were, thus, considered as important neural correlates of MW. Twenty of them displayed dose related increases in c-Fos expression and correspond to regions related to autonomic functions. Low c-Fos expression was detected in some regions involved in motor control or in reward, suggesting either their minor role in MW or a limitation of the technique. This dose-response analysis suggests that the increase in the severity of autonomic manifestations of MW is associated with a gradual activation of major structures of the autonomic nervous system. PMID- 11283965 TI - Does diabetes induced in rats cause malformations? PMID- 11283967 TI - Diabetic embryopathy 2001: moving beyond the "diabetic milieu". PMID- 11283968 TI - Mutation analysis of left-right axis determining genes in NOD and ICR, strains susceptible to maternal diabetes. AB - BACKGROUND: Genetic background of the fetus contributes to the pathogenesis of congenital malformation after teratogen exposure. Such contribution is illustrated in left-right axis malformations observed in the F1 offspring of nonobese diabetic (NOD) mouse dams and sires from different strains. When sires of the NOD, ICR, or C57BL/6J were mated with NOD dams, incidence varied depending on the fetal genotype, with 65% in NOD x NOD, 24% in NOD x ICR, and 7% in NOD x C57BL/6J. METHODS: As a first step in elucidating the molecular basis of the interstrain differences in susceptibility to situs defects, we compared genomic sequences of six genes HNF3beta, Acvr2b, Nodal, ZIC3, Lefty1, and Smad2, which are involved in the normal development of left-right axis among NOD, ICR, and C57BL/6J strains. RESULTS: The outbred strain ICR had 1) a 0.2-kb insertion in the putative promoter region of the isoform E of HNF3beta together with a G to A change that could create a potential splice acceptor in the exon 3 of HNF3beta (gene frequency P = 0.36), 2) five single base substitutions within the 5' controlling element and a proline to serine substitution (P2S) of Lefty1 (P = 0.77), and 3) a tyrosine to histidine substitution within the prodomain of Nodal (P = 0.48). The inbred strain NOD had the same G to A change as ICR and a three base deletion in the putative promoter of isoform E of HNF3beta. CONCLUSIONS: We suggest that sequence variations in HNF3beta, Lefty1, and Nodal might account, in part, for the interstrain differences in susceptibility to situs abnormalities among the offspring of diabetic dams. PMID- 11283969 TI - Placental transfer of vigabatrin (gamma-vinyl GABA) and its effect on concentration of amino acids in the embryo of TO mice. AB - BACKGROUND: The mechanism of the teratogenicity of vigabatrin (VGB) is unknown. The objectives of this study were to determine the placental transfer of VGB and to evaluate the effect of VGB on maternal, placental, and fetal concentrations of amino acids. METHODS: A single dose of 400 mg/kg VGB in physiological saline was administered intraperitoneally to a group of Theiler outbred (TO) mice on gestational day (GD) 10. The controls received a proportionate volume of saline. Maternal blood samples, embryos, and placentas were collected at 3.5, 6.0, and 9.0 hr after treatment and their total amino acid concentrations determined in an ion-exchange amino acid analyzer. RESULTS: At 3.5 hr, there was a decrease in concentrations of some amino acids in the blood, placenta, and embryos of VGB treated mice, but the decrease in methionine was most marked. gamma-aminobutyric acid (GABA) was significantly higher in the VGB group in both the embryos and the placentas at 3.5 hr but at 6.0 and 9.0 hr the differences were not significant. Vigabatrin levels were higher in the placenta than in the embryo at 3.5 hr, but at 6.0 hr there was an overlap of the VGB peak with that of tryptophan with very much lower levels than at 3.5 hr. At 9.0 hr, there was no vigabatrin peak in either the placenta or the embryo. CONCLUSIONS: Maternal exposure to VGB results in peak levels of the drug after 3.5 hr in the placenta and embryo. Methionine concentration is most severely affected in VGB-treated mothers, placentas, and fetuses. We speculate that this deficiency could be a possible mechanism for the teratogenic effects of vigabatrin. PMID- 11283970 TI - Co-localization of active caspase-3 and DNA fragmentation (TUNEL) in normal and hyperthermia-induced abnormal mouse development. AB - BACKGROUND: Previous work has shown that caspase-3 activation and DNA fragmentation, two hallmarks of apoptosis, are induced in day 9 mouse embryos exposed to hyperthermia (43 degrees C); however, the methods used to assess caspase-3 activation (Western blot) and DNA fragmentation (gel electrophoresis) did not allow these apoptotic events to be localized to specific cells within the embryo. METHODS: To co-localize active caspase-3 and DNA fragmentation to specific cells, we used paraffin sections of day 13 mouse limb buds, sections of control and hyperthermia-treated day 9 mouse embryos, and sequential immunohistochemical staining for caspase-3 and TUNEL staining for DNA fragmentation. We used a primary rabbit antibody specific for the active, p17 subunit of caspase-3 and a goat anti-rabbit secondary antibody conjugated to Alexa 594 fluorochrome (red fluorescence) to localize active caspase-3. To co localize DNA fragmentation, we subsequently processed the same sections by the TUNEL method using fluorescein-labeled dUTP (green fluorescence). RESULTS: Using this dual labeling approach, we show that active caspase-3 (caspase-3 positive) and DNA fragmentation (TUNEL positive) occur in a sub-population of interdigital mesenchyme cells of day 13 mouse limb buds. Using the same approach, we detected a small number of caspase-3 positive and TUNEL-positive cells in the central nervous system and in the mesenchyme of the first branchial arch of untreated day 9 mouse embryos. The number of caspase-3 and TUNEL-positive cells are greatly increased 5 hr after a brief exposure to hyperthermia (43 degrees C, 13 min). Caspase-3 and TUNEL-positive cells were most abundant in the neuroepithelium of the developing central nervous system, mesenchyme of the first pharyngeal arch, and somitic mesoderm. In contrast, the heart, mesencephalic mesenchyme, and the visceral yolk sac contained few, if any, caspase-3 and TUNEL-positive cells. CONCLUSIONS: This is the first demonstration that activation of caspase-3 and DNA fragmentation co-localize in cells programmed to die in the interdigital mesenchyme of day 13 limb buds and in the neuroepithelium and branchial arch mesenchyme of day 9 mouse embryos. Similarly, our results represent the first co localization of teratogen-induced activation of caspase-3 and DNA fragmentation in specific cells of early postimplantation mouse embryos, and confirm that cells of the developing central nervous system are acutely sensitive to the cell death inducing potential of hyperthermia, whereas cells of the heart are resistant. Finally, we show for the first time that, like cells of the heart, cells of the mesencephalic mesenchyme and the visceral yolk sac are also resistant to hyperthermia-induced apoptosis. PMID- 11283971 TI - Interactions in developmental toxicology: combined action of restraint stress, caffeine, and aspirin in pregnant mice. AB - BACKGROUND: Stress can result in an increased use of substances such as caffeine and aspirin. The effect of maternal stress on concurrent exposure to caffeine and aspirin on prenatal development was assessed in mice. METHODS: On gestational day 9, mice were assigned to three treatment groups orally exposed to caffeine (30 mg/kg), aspirin (250 mg/kg), or a combination of caffeine (30 mg/kg) and aspirin (250 mg/kg). Three additional groups of pregnant animals received similar caffeine and aspirin doses and were immediately subjected to restraint for 14 hr. Control groups included unrestrained and restrained pregnant mice not exposed to caffeine or aspirin. All dams were euthanized on gestational day 18. Live fetuses were evaluated for sex, body weight, and external, internal, and skeletal malformations and variations. RESULTS: A single oral dose of caffeine or aspirin did not cause significant maternal toxicity. However, coadministration of these drugs with restraint produced some adverse maternal effects (i.e., reduction in maternal weight gain and food consumption on gestational days 9-11). In relation to embryo/fetal toxicity, the incidence of some skeletal defects was significantly increased after exposure to caffeine, aspirin, or maternal restraint, and their binary and ternary combinations. CONCLUSIONS: Although caffeine and aspirin were given in a single dose in this study, the results suggest that prenatal stress could slightly exacerbate the maternal and developmental toxicity of the combination of these drugs in mice. PMID- 11283972 TI - Phenytoin-induced cleft palate: evidence for embryonic cardiac bradyarrhythmia due to inhibition of delayed rectifier K+ channels resulting in hypoxia reoxygenation damage. AB - BACKGROUND: Phenytoin (PHT) teratogenicity has been related to embryonic arrhythmia due to the capacity of PHT to block I(K) channels pharmacologically, resulting in hypoxia-reoxygenation damage. The aim of this study was to further elucidate the proposed mechanism. METHODS: Pregnant CD-1 mice were given PHT (85 mg/kg) or saline intraperitoneally on gestational days 10-11. Embryonic heart rhythm and presence of hemorrhage in orofacial region was recorded on day 12, fetuses were examined for malformations on day 18. Embryonic heart rate was also recorded on individual days after dosing days 9-16. In addition, PHT was given at doses of 10, 25, or 85 mg/kg on day 12 for analysis of plasma concentrations. RESULTS: PTH-induced bradycardia and arrhythmia in approximately 20% of the embryos, 48% showed hemorrhage in the orofacial region; 39% of the fetuses had cleft palate. The region in which hemorrhages were visible in the embryo corresponded with the region where tissue deficiency (cleft palate) was visible in the fetus at term. None of the controls showed hemorrhages, dysrhythmia, or cleft palate. PHT affected embryonic heart rates on days 9-13, but not on days 14 16. Single dose administration on day 12, the most sensitive day, resulted in a dose-dependent decrease in embryonic heart rate (12-34%). Embryonic arrhythmia occurred at 25 and 85, but not at 10 mg/kg or in the controls. Mean maternal free plasma concentrations were 6 and 14 micromol/L in the 10- and 25-mg/kg groups, respectively. CONCLUSIONS: PHT-induced cleft palate was preceded by embryonic dysrhythmia and hemorrhage in the orofacial region. Embryonic heart rhythm was phase specifically affected, as described for selective I(Kr) channel blockers, at clinically relevant concentrations. The results support the idea that PHT teratogenicity is a consequence of pharmacologically induced dysrhythmia and hypoxia-related damage. PMID- 11283973 TI - Recurrent cleft lip and palate in siblings of a patient with malabsorption syndrome, probably caused by hypovitaminosis a associated with folic acid and vitamin B(2) deficiencies. AB - We present a woman with metabolic disorders secondary to malabsorption and renal disease who gave birth to a stillborn male fetus with left unilateral cleft lip and palate and a live born infant with left unilateral cleft lip and palate. We discuss potential cofactors that could be implicated in the abnormal embryonic process. PMID- 11283975 TI - Immigration and hybridization patterns of yellow and anubis baboons in and around Amboseli, Kenya. AB - In 1986, Samuels and Altmann reported evidence for a hybrid zone between Papio anubis and Papio cynocephalus in Amboseli, Kenya, in a baboon population that has been the subject of long-term study since 1971 [Samuels & Altmann, International Journal of Primatology 7:131-138, 1986]. In the current report we document ongoing patterns of hybridization in Amboseli between anubis and yellow baboons. In July 2000, we exhaustively scored living members of study groups for their degree of hybridity, using seven phenotypic characteristics (five in juveniles). We also scored all former members of study groups on the basis of photographic records, field notes, and observer recollections. A total of five anubis males and 11 males with hybrid phenotypes have immigrated into study groups over the course of the long-term study, and immigrations by hybrid males have increased in frequency over time. Further, the increasing frequency of hybrid phenotypes among animals born into study groups indicates that anubis and hybrid males have successfully reproduced in study groups. However, hybrid phenotypes and anubis immigrations were limited to groups in the southwestern portion of the Amboseli basin, with no hybrids occurring in the six eastern groups. Finally, we present evidence that anubis and hybrid males in Amboseli exhibit patterns of natal dispersal that are different from those of yellow males in Amboseli: males with anubis or hybrid phenotypes were significantly more likely to immigrate as juveniles or young subadults than were yellow males. PMID- 11283976 TI - Demography and pedigree structure of an SPF colony of rhesus monkeys (Macaca mulatta). AB - The SPF rhesus colony at the M.D. Anderson Cancer Center in Bastrop, Texas, was analyzed with the aim of determining the demographic and genetic effects of stringent selection for virus-free breeders, permanent quarantine, continued surveillance, and culling of animals that show evidence of viral infection. The analysis shows minimal effects on population viability and loss of genetic variability in comparison with the traditionally managed (non-SPF) portion of the population. PMID- 11283977 TI - Measurement of urinary and fecal steroid metabolites during the ovarian cycle in captive and wild Japanese macaques, Macaca fuscata. AB - We measured the concentration of steroid hormones from urine, feces, and blood samples of two captive Japanese macaques, Macaca fuscata, during nonconceptive ovarian cycles to compare the patterns of the excreted steroids with those of circulating steroids. Urine and feces were analyzed for estrone conjugates (E1C) and pregnanediol-3-glucronide (PdG) using enzyme immunoassays (EIAs), while plasma was analyzed for estradiol-17beta(E2), progesterone (P), and luteinizing hormone (LH) using radioimmunoassays (RIAs). Urinary and fecal E1C and PdG levels were approximately parallel to plasma E2 and P levels, respectively. The E1C profiles of daily urinary and fecal samples revealed a midcycle peak, followed by a sustained PdG increase lasting up to two weeks from the E1C peak. A fecal E1C peak was one day later than the urinary E1C peak. One of the captive females exhibited a discrete plasma LH peak, one indicator that ovulation has occurred, on the day following the urinary E1C peak, i.e., the same day of fecal E1C peak. We measured excreted steroids in nine wild females and determined the timing of ovulation by comparing fecal steroid profiles to those obtained in captive monkeys. Data from wild females indicated that eight of nine females conceived during their first ovulatory cycle of the sampling period, whereas the remaining female failed to conceive during the sampling period even though she ovulated. In the eight females that conceived, E1C increased again following the detected or estimated E1C peak, with levels comparable to the preovulatory peak levels, and sustained elevations of PdG for over 40 days. These data illustrate that the urinary and fecal profiles of ovarian steroid excretion obtained through the application of these noninvasive techniques provide an accurate approach for monitoring conceptive and nonconceptive ovarian cycle in captive and free-living Japanese macaques. PMID- 11283978 TI - Water proton MR properties of human blood at 1.5 Tesla: magnetic susceptibility, T(1), T(2), T*(2), and non-Lorentzian signal behavior. AB - Accurate knowledge of the magnetic properties of human blood is required for the precise modeling of functional and vascular flow-related MRI. Herein are reported determinations of the relaxation parameters of blood, employing in vitro samples that are well representative of human blood in situ. The envelope of the blood (1)H(2)O free-induction decay signal magnitude during the first 100 msec following a spin echo at time TE is well- described empirically by an expression of the form, S(t) = S(o). exp[-R(*)(2). (t - TE) - AR*. (t - TE)(2)]. The relaxation parameters AR* and R(*)(2) increase as a function of the square of the susceptibility difference between red blood cell and plasma and depend on the spin-echo time. The Gaussian component, AR*, should be recognized in accurate modeling of MRI phenomena that depend upon the magnetic state of blood. The magnetic susceptibility difference between fully deoxygenated and fully oxygenated red blood cells at 37 degrees C is 0.27 ppm, as determined independently by MR and superconducting quantum interference device (SQUID) measurements. This value agrees well with the 1936 report of Pauling and Coryell (Proc Natl Acad Sci USA 1936;22:210-216), but is substantially larger than that frequently used in MRI literature. Magn Reson Med 45:533-542, 2001. PMID- 11283979 TI - 17O relaxation time and NMR sensitivity of cerebral water and their field dependence. AB - 17O spin relaxation times and sensitivity of detection were measured for natural abundance H(2)(17)O in the rat brain at 4.7 and 9.4 Tesla. The relaxation times were found to be magnetic field independent (T(2) = 3.03 +/- 0.08 ms, T(*)(2) = 1.79 +/- 0.04 ms, and T(1) = 4.47 +/- 0.14 ms at 4.7T (N = 5); T(2) = 3.03 +/- 0.09 ms, T(*)(2) = 1.80 +/- 0.06 ms, and T(1) = 4.84 +/- 0.18 ms at 9.4T (N = 5)), consistent with the concept that the dominant relaxation mechanism is the quadrupolar interaction for this nucleus. The (17)O NMR sensitivity was more than fourfold higher at 9.4T than at 4.7T, for both the rat brain and a sodium chloride solution. With this sensitivity gain, it was possible to obtain localized (17)O spectra with an excellent signal-to-noise ratio (SNR) within 15 s of data acquisition despite the relatively low gyromagnetic ratio of this nucleus. Such a 15-s 2D (17)O-MRS imaging data set obtained for natural abundance H(2)(17)O in the rat brain yielded an SNR greater than 40:1 for a approximately 16 microl voxel. This approach was employed to measure cerebral blood flow using a bolus injection of H(2)(17)O via one internal carotid artery. These results demonstrate the ability of (17)O-MRS imaging to reliably map the H(2)(17)O dynamics in the brain tissue, and its potential for determining tissue blood flow and oxygen consumption rate changes in vivo. Magn Reson Med 45:543-549, 2001. PMID- 11283980 TI - Temporal frequency analysis of dynamic MRI techniques. AB - Dynamic imaging strategies often involve updating certain areas of k-space (i.e., the low spatial frequencies) more frequently than others. However, important dynamic signal changes may occur anywhere in k-space. In this study, a dynamic k space sampling analysis method was developed to determine the energy error associated with specific dynamic sampling strategies. The method uses the temporal power spectrum of k-space signals to determine the level and k-space locations of sampling errors. The proposed method was used to compare two dynamic sampling strategies (full sequential and keyhole) for a dynamic first-pass bolus simulation and a continuous heart imaging study. The error analysis agreed well with the errors in the reconstructed images. The technique can be used to determine the minimum sampling frequency for any location in the k-space, and may ultimately be used to optimize dynamic sampling strategies. Magn Reson Med 45:550 556, 2001. PMID- 11283981 TI - MRA of hemodialysis access grafts and fistulae using selective contrast injection and flow interruption. AB - MR is a potentially attractive modality for evaluating hemodialysis access anatomy and function. However, the wide range of flow rates in the hemodialysis access complicates interpretation of phase contrast, time-of-flight, and even contrast-enhanced MR angiograms. At high flow rates, signal voids may easily arise at mild narrowings or sharp-angled anastomoses. A method is proposed which visualizes hemodialysis accesses without flow artifacts. Diluted Gd-DTPA is hand injected directly into the access, while a cuff is used to reduce and subsequently interrupt access flow. Filling of the access is monitored using a fast projection technique with complex subtraction. When filling is satisfactory, a 3D acquisition is started. The feasibility of this selective contrast-enhanced MR angiography technique is demonstrated in four Cimino-fistulae and four PTFE grafts. Magn Reson Med 45:557-561, 2001. PMID- 11283982 TI - Myocardial wall tagging with undersampled projection reconstruction. AB - Azimuthally undersampled projection reconstruction (PR) acquisition is investigated for use in myocardial wall tagging with MR using grid tags to provide increased temporal and spatial resolution. PR can provide the high resolution images required for tagging with very few projections, at the expense of artifact. Insight is provided into the PR undersampling artifact, in the context of measuring myocardial motion with tags. For Fourier transform imaging, at least 112 phase-encodings must be collected to image tagging grids spaced 7 pixels apart. PR requires about 80 projections, a 1.4-fold reduction in scan time. Magn Reson Med 45:562-567, 2001. Published 2001 Wiley-Liss, Inc. PMID- 11283983 TI - Lactate discrimination incorporated into echo-planar spectroscopic imaging. AB - A technique for discriminating a lactate signal from overlapping lipid signals in (1)H spectroscopic imaging is presented. It is based on J-coupling between lactate protons and on the broad spectral bandwidth of lipid signal. Measurement parameters used in the technique are determined so that TE is separated from n/J (n: a natural number, J: J-coupling constant) enough to suppress the lipid signal at the time when the lactate signal is strongest. Data processing is used to calculate the lactate signal intensity from the reconstructed spectra. This technique enables lactate to be discriminated in a single measurement and enables spectra of other metabolites to be acquired simultaneously. However, it necessitates a homogeneous magnetic field, long TE, and supplementary lipid suppression. Discrimination of the lactate signal is demonstrated by applying lactate-discriminating echo-planar spectroscopic imaging (EPSI), which combines this discrimination technique with the standard EPSI, to rat focal cerebral ischemia models. Magn Reson Med 45:568-574, 2001. PMID- 11283984 TI - MR microscopy of cobalt-labeled nerve cells and pathways in an invertebrate brain (Sepia officinalis, Cephalopoda). AB - This article describes a novel application of contrast-enhanced MR microscopy to trace nerve cells and pathways through small invertebrate brains. Using the cuttlefish Sepia officinalis (Cephalopoda) as a model, the cells and pathways of one of the brain nerves were labeled with paramagnetic cobalt(II) ions by conventional centripetal cobalt iontophoresis. In MR microscopy, the cobalt labeled cell bodies and pathways became hypointense in 9.4 T spin echo images. Their course and distribution were identical with those seen with conventional histological techniques after cobalt sulphide precipitation (with or without subsequent silver intensification). Magn Reson Med 45:575-579, 2001. PMID- 11283985 TI - Visualization of neural tissue water compartments using biexponential diffusion tensor MRI. AB - The apparent diffusion tensor (ADT) imaging method was extended to account for multiple diffusion components. A biexponential ADT imaging experiment was used to obtain separate images of rapidly and slowly diffusing water fractions in excised rat spinal cord. The fast and slow component tensors were compared and found to exhibit similar gross features, such as fractional anisotropy, in both white and gray matter. However, there were also some important differences, which are consistent with the different structures occupying intracellular and extracellular spaces. Evidence supporting the assignment of the two tensor components to extracellular and intracellular water fractions is provided by an NMR spectroscopic investigation of homogeneous samples of brain tissue. Magn Reson Med 45:580-587, 2001. PMID- 11283986 TI - Imaging brain function in humans at 7 Tesla. AB - This article describes experimental studies performed to demonstrate the feasibility of BOLD fMRI using echo-planar imaging (EPI) at 7 T and to characterize the BOLD response in humans at this ultrahigh magnetic field. Visual stimulation studies were performed in normal subjects using high-resolution multishot EPI sequences. Changes in R(*)(2) arising from visual stimulation were experimentally determined using fMRI measurements obtained at multiple echo times. The results obtained at 7 T were compared to those at 4 T. Experimental data indicate that fMRI can be reliably performed at 7 T and that at this field strength both the sensitivity and spatial specificity of the BOLD response are increased. This study suggests that ultrahigh field MR systems are advantageous for functional mapping in humans. Magn Reson Med 45:588-594, 2001. PMID- 11283987 TI - Neuroimaging at 1.5 T and 3.0 T: comparison of oxygenation-sensitive magnetic resonance imaging. AB - Noise properties, the signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), and signal responses were compared during functional activation of the human brain at 1.5 and 3.0 T. At the higher field spiral gradient-echo (GRE) brain images revealed an average gain in SNR of 1.7 in fully relaxed and 2.2 in images with a repetition time (TR) of 1.5 sec. The tempered gain at longer TRs reflects the fact that the physiological noise depends on the signal strength and becomes a larger fraction of the total noise at 3.0 T. Activation of the primary motor and visual cortex resulted in a 36% and 44% increase of "activated pixels" at 3.0 T, which reflects a greater sensitivity for the detection of activated gray matter at the higher field. The gain in the CNR exhibited a dependency on the underlying tissue, i.e., an increase of 1.8x in regions of particular high activation-induced signal changes (presumably venous vessels) and of 2.2x in the average activated areas. These results demonstrate that 3.0 T provides a clear advantage over 1.5 T for neuroimaging of homogeneous brain tissue, although stronger physiological noise contributions, more complicated signal features in the proximity of strong susceptibility gradients, and changes in the intrinsic relaxation times may mediate the enhancement. Magn Reson Med 45:595-604, 2001. PMID- 11283988 TI - Dynamic (19)F-MRI of pulmonary ventilation using sulfur hexafluoride (SF(6)) gas. AB - A new method for dynamic imaging of pulmonary wash-in and wash-out kinetics of inhaled sulfur hexafluoride (SF(6)) gas was developed. Measurements at the fluorine-19 Larmor frequency were performed in pigs using a gradient echo pulse sequence with 0.5 ms echo time and a measurement time of 9.1 s per image. Dynamic MRI was performed during wash-in and wash-out of SF(6) gas in mechanically ventilated porcine lungs. A postprocessing strategy was developed for quantitative determination of wash-out time constants in the presence of noise. Mean wash-out constants were 4.78 +/- 0.48 breaths vs. 4.33 +/- 0.76 breaths for left and right lung when ventilation was performed with low tidal volume, and 1.73 +/- 0.16 breaths vs. 1.85 +/- 0.11 breaths with high tidal volume ventilation. In conclusion, breath-hold MRI of SF(6) gas is feasible in large animals. Moreover, regional wash-in and wash-out kinetics of SF(6) can be determined noninvasively with this new method. Potential human applications are discussed. Magn Reson Med 45:605-613, 2001. PMID- 11283989 TI - Dynamic contrast-enhanced MRI of experimental spinal cord injury: in vivo serial studies. AB - The progression of experimental spinal cord injury (SCI) was followed with in vivo dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and neurobehavioral studies on postinjury days 0, 2, 4, 7, 10, 14, 17, 21, 28, 35, and 42. Gadopentate dimeglumine (Gd) was administered IV and postcontrast, T(1) weighted, axial images were acquired repetitively for up to 60 min. Images were analyzed to determine the spatial and temporal evolution of the intensity enhancement. A statistical decision mechanism was developed to objectively detect the enhancement. Strong and rapid enhancement was observed at the epicenter of injury, indicating a significant compromise in blood spinal cord barrier. The enhanced regions in each slice were combined to estimate the area and volume of the lesion. On the day of injury, around 85% of the total cord area at the epicenter showed enhancement within the first 15 min of Gd administration. At the same time, the enhanced volumes attained nearly 40% of the total cord volume and extended axially over 8 mm along the cord. These quantities decreased steadily with time, with a concomitant improvement in the motor functions. The volume of enhancement correlated highly with the neurobehavioral tests (r = -0.87). DCE MRIs revealed small hyperintense regions distributed inside white matter about two weeks postinjury. Based on histology, these enhancements appear to represent new vessels with "leaky endothelium." Magn Reson Med 45:614-622, 2001. PMID- 11283990 TI - Reducing oblique flow effects in interleaved EPI with a centric reordering technique. AB - Segmented interleaved echo planar imaging offers a fast and efficient approach to magnetic resonance angiography. Unfortunately, this technique is particularly sensitive to oblique flow in the imaging plane. In this work, a mathematical analysis of oblique flow effects for several types of k-space coverage is presented. The conventional linear acquisition scheme, an alternating centric and a nonalternating centric encoding scheme are compared with respect to their flow properties. It is shown both by simulations and imaging experiments that artifacts from oblique in-plane flow are effectively reduced by both centric reordered phase-encoding schemes. The nonalternating centric acquisition scheme is preferred to the alternating centric scheme due to the smoother phase error transition in k-space in the presence of obliquely-angled flow. Magn Reson Med 45:623-629, 2001. PMID- 11283991 TI - Rapid T(1) mapping using multislice echo planar imaging. AB - Determination of neurological pathology in white matter disease can be made in a semiquantitative way from T(1)- or T(2)-weighted images. A higher level of quantification based on measured T(1) or T(2) values has been either limited to specific regions of interest or to low-resolution maps. Higher-resolution T(1) maps have proved difficult to obtain due to the excessively long scan times required using conventional techniques. In this study, clinically acceptable images are obtained by using single-shot echo planar imaging (EPI) with an acquisition scheme that maximizes signal-to-noise while minimizing the scan time. Magn Reson Med 45:630-634, 2001. PMID- 11283992 TI - Estimation of respiration-induced noise fluctuations from undersampled multislice fMRI data. AB - Functional MRI time series data are known to be contaminated by highly structured noise due to physiological fluctuations. Significant components of this noise are at frequencies greater than those critically sampled in standard multislice imaging protocols and are therefore aliased into the activation spectrum, compromising the estimation of functional activations and the determination of their significance. However, in this work it is demonstrated that unaliased noise information is available in multislice data, and can be used to estimate and reduce noise due to high-frequency respiratory-related fluctuations. Magn Reson Med 45:635-644, 2001. Published 2001 Wiley-Liss, Inc. PMID- 11283993 TI - Motion artifact reduction and vessel enhancement for free-breathing navigator gated coronary MRA using 3D k-space reordering. AB - Breathing-induced bulk motion of the myocardium during data acquisition may cause severe image artifacts in coronary magnetic resonance angiography (MRA). Current motion compensation strategies include breath-holding or free-breathing MR navigator gating and tracking techniques. Navigator-based techniques have been further refined by the applications of sophisticated 2D k-space reordering techniques. A further improvement in image quality and a reduction of relative scanning duration may be expected from a 3D k-space reordering scheme. Therefore, a 3D k-space reordered acquisition scheme including a 3D navigator gated and corrected segmented k-space gradient echo imaging sequence for coronary MRA was implemented. This new zonal motion-adapted acquisition and reordering technique (ZMART) was developed on the basis of a numerical simulation of the Bloch equations. The technique was implemented on a commercial 1.5T MR system, and first phantom and in vivo experiments were performed. Consistent with the results of the theoretical findings, the results obtained in the phantom studies demonstrate a significant reduction of motion artifacts when compared to conventional (non-k-space reordered) gating techniques. Preliminary in vivo findings also compare favorably with the phantom experiments and theoretical considerations. Magn Reson Med 45:645-652, 2001. PMID- 11283994 TI - Steady-state preparation for spoiled gradient echo imaging. AB - While spoiled gradient echo sequences provide a rapid means of acquiring T(1) weighted images, it is often desirable that the magnetization be in the steady state to avoid artifacts. For some applications, this requires many "dummy" repetitions of the pulse sequence prior to data collection, delaying image acquisition. A method is presented in which a saturation pulse, followed by a prescribed recovery period, places longitudinal magnetization levels of all materials near steady state, ready for data acquisition much sooner than when employing only dummy repetitions to achieve steady state. Effects of transverse coherences are studied using configuration theory. The method is shown to be effective in both phantom studies and in vivo applications, including real-time imaging, multiphase cardiac imaging, and triggered contrast-enhanced angiography. Magn Reson Med 45:653-661, 2001. PMID- 11283995 TI - Monitoring blood oxygen state in muscle microcirculation with transverse relaxation. AB - Oxygen uptake from the microcirculation is a direct measure of tissue function. Magnetic resonance is capable of detecting differences between oxygenated and deoxygenated blood due to the paramagnetic properties of deoxyhemoglobin. At the level of the microcirculation, however, imaging methods cannot directly visualize the vessels. Instead, bulk MR parameters are investigated for their ability to monitor blood oxygen saturation (%O(2)) changes in the microcirculation of tissue, specifically skeletal muscle. Experiments in an in vitro model verified the feasibility of detecting changes in exponential decay signals, and also verified the prediction of only two distinct decay components. Experiments in a rabbit model demonstrate that T(2)' and monoexponential T(2) decay are not sensitive to blood oxygen changes, but that the long-T(2) component in a biexponential fit is correlated to the blood oxygen state. Assuming a two-pool model for water protons in muscle, and with knowledge of the T(2)-%O(2) relation, estimates of the microcirculation blood oxygen state can be made with some reasonable assumptions. Magn Reson Med 45:662-672, 2001. PMID- 11283996 TI - Planar strip array (PSA) for MRI. AB - Parallel, spatial-encoded MRI requires a large number of independent detectors that simultaneously acquire signals. The loop structure and mutual coupling in conventional phased arrays limit the number of coils and therefore the potential reduction in minimum scan time achievable by parallel MRI tchniques. A new near field MRI detector array, the planar strip array (PSA), is presented that eliminates the coupling problems and can be extended to a very large number of detectors and high MRI frequencies. Its basic structure is an array of parallel microstrips with a high permittivity substrate and overlay. The electromagnetic (EM) wavelength can be adjusted with the permittivity, and the strip lengths tuned to a preselected fraction of the wavelength of the MRI frequency. EM wave analysis and measurements on a prototype four-element PSA reveal that the coupling between the strips vanishes when the strip length is either an integer times a quarter wavelength for a standing-wave PSA, or a half wavelength for a travelling-wave PSA, independent of the spacing between the strips. The analysis, as well as phantom and human MRI experiments performed by conventional and parallel-encoded MRI with the PSA at 1.5 T, show that the decoupled strips produce a relatively high-quality factor and signal-to-noise ratio, provided that the strips are properly terminated, tuned, and matched or coupled to the preamplifiers. Magn Reson Med 45:673-683, 2001. PMID- 11283997 TI - Signal-to-noise ratio and absorbed power as functions of main magnetic field strength, and definition of "90 degrees " RF pulse for the head in the birdcage coil. AB - Calculations of the RF magnetic (B(1)) field as a function of frequency between 64 and 345 MHz were performed for a head model in an idealized birdcage coil. Absorbed power (P(abs)) and SNR were calculated at each frequency with three different methods of defining excitation pulse amplitude: maintaining 90 degrees flip angle at the coil center (center alpha = pi/2), maximizing FID amplitude (Max. A(FID)), and maximizing total signal amplitude in a reconstructed image (Max. A(image)). For center alpha = pi/2 and Max. A(image), SNR increases linearly with increasing field strength until 260 MHz, where it begins to increase at a greater rate. For these two methods, P(abs) increases continually, but at a lower rate at higher field strengths. Above 215 MHz in MRI of the human head, the use of FID amplitude to set B(1) excitation pulses may result in apparent decreases in SNR and power requirements with increasing static field strength. Magn Reson Med 45:684-691, 2001. PMID- 11283998 TI - Calculations of B(1) distribution, SNR, and SAR for a surface coil adjacent to an anatomically-accurate human body model. AB - Calculations of the radiofrequency magnetic (B(1)) field, SAR, and SNR as functions of frequency between 64 and 345 MHz for a surface coil against an anatomically-accurate human chest are presented. Calculated B(1) field distributions are in good agreement with previously-published experimental results up to 175 MHz, especially considering the dependence of field behavior on subject anatomy. Calculated SNR in the heart agrees well with theory for low frequencies (nearly linear increase with B(0) field strength). Above 175 MHz, the trend in SNR with frequency begins to depend largely on location in the heart. At all frequencies, present limits on local (1 g) SAR levels are exceeded before limits on whole-body average limits. At frequencies above 175 MHz, limits on SAR begin to be an issue in some common imaging sequences. These results are relevant for coils and subjects similar to those modeled here. Magn Reson Med 45:692-699, 2001. PMID- 11283999 TI - Whole body detection and imaging of nitric oxide generation in mice following cardiopulmonary arrest: detection of intrinsic nitrosoheme complexes. AB - Ischemic tissues generate nitric oxide (NO) by direct reduction of tissue nitrite under the acidic conditions that occur during ischemia. In view of the important implications of this enzyme-independent mechanism of NO generation on the pathogenesis and treatment of tissue injury, the NO formation in mice subjected to cardiopulmonary arrest was measured and imaged. Real-time measurement of NO generation was performed by detection of naturally generated NO-heme complexes in tissues using L-band electron paramagnetic resonance (EPR) spectroscopy. To distinguish NO generated from nitrite, animals were labeled with isotopically enriched (15)N-nitrite. Mice were infused with nitrite (70 mg/kg, intravenous), cardiopulmonary arrest induced by an overdose of phenobarbital, and transferred to the EPR resonator. Measurements of NO generation were performed on the intact animal at the levels of the head, thorax, and abdomen. At the end of 3 hr, major organs were isolated and analyzed for their NO signal. The NO complexes were found to have maximum levels in lung, heart, and liver. Three-dimensional spatial mapping of the NO complex in the intact animal subjected to cardiopulmonary arrest was performed using EPR imaging techniques. The images also confirmed the maximum formation in the lungs, heart, and liver. The present data reveal that mice subjected to cardiopulmonary arrest generate large amounts of NO, which is nitrite mediated. The observed signal was largely due to heme-bound NO, which accounted for the high concentrations found in these organs. This increased NO formation during cardiopulmonary arrest could contribute to the difficulty of resuscitation after long periods of arrest. Magn Reson Med 45:700-707, 2001. PMID- 11284000 TI - Improved selectivity of double quantum coherence filtering for the detection of glutathione in the human brain in vivo. AB - An improved double quantum coherence (DQC) filter for the selective in vivo detection of glutathione (GSH) in the human brain at 1.5 Tesla is presented. The goal was to minimize contamination of the DQC-filtered GSH signal at 2.9 ppm with contributions arising from GABA. The modification consists of tailoring the frequency response of the read pulse, which converts DQC into anti-phase single quantum coherence in such a way that the GABA beta and gamma resonances at 3.0 and 1.9 ppm, respectively, remain unaffected. An implementation incorporating a Dante pulse train is used for in vitro tests as well as for in vivo applications. Magn Reson Med 45:708-710, 2001. PMID- 11284001 TI - Cesium-133: a potential reporter of the hepatic uptake of contrast agents. AB - NMR spectroscopy of intracellularly located (133)Cs has been used to monitor the uptake of Gd-EOB-DTPA by the isolated rat liver. As shown by (31)P spectroscopy, accumulation of (133)Cs ions in hepatocytes does not produce detectable effects on the metabolism. The hepatic internalization of Gd-EOB-DTPA was followed by the paramagnetic relaxation enhancement of the intracellular (133)Cs ions, and confirmed by parallel quantitations of Gd and Cs run by inductively coupled plasma (ICP) analysis of liver samples and aliquots of perfusate. The relaxation data significantly underestimate the Gd content, suggesting a potential compartmentation of Cs(+) and/or the contrast agent. Magn Reson Med 45:711-715, 2001. PMID- 11284002 TI - High-resolution blood flow velocity measurements in the human finger. AB - MR phase contrast blood flow velocity measurements in the human index finger were performed with triggered, nontriggered, and cine acquisition schemes. A strong (G(max) = 200 mT/m), small bore (inner diameter 12 cm) gradient system inserted in a whole body 3 Tesla MR scanner allowed high-resolution imaging at short echo times, which decreases partial volume effects and flow artifacts. Arterial blood flow velocities ranging from 4.9-19 cm/sec were measured, while venous blood flow was significantly slower at 1.5-7.1 cm/sec. Taking into account the corresponding vessel diameters ranging from 800 microm to 1.8 mm, blood flow rates of 3.0-26 ml/min in arteries and 1.2-4.8 ml/min in veins are obtained. The results were compared to ultrasound measurements, resulting in comparable blood flow velocities in the same subjects. Magn Reson Med 45:716-719, 2001. PMID- 11284003 TI - T(1) quantification with inversion recovery TrueFISP. AB - A snapshot FLASH sequence can be used to acquire the time course of longitudinal magnetization during its recovery after a single inversion pulse. However, excitation pulses disturb the exponential recovery of longitudinal magnetization and may produce systematic errors in T(1) estimations. In this context the possibility of using the TrueFISP sequence to detect the recovery of longitudinal magnetization for quantitative T(1) measurements was examined. Experiments were performed on different Gd-doped water phantoms and on humans. T(1) values derived from inversion recovery TrueFISP were in excellent agreement with the single point method even for flip angles up to 50 degrees. In terms of T(1) accuracy and SNR, the proposed method seems to be superior to the conventional inversion recovery snapshot FLASH technique. Magn Reson Med 45:720-723, 2001. PMID- 11284004 TI - Integral method for numerical simulation of MRI artifacts induced by metallic implants. AB - Numerical simulation is a valuable tool for the study of magnetic susceptibility artifacts from metallic implants. A major difficulty in the simulation lies in the computation of the magnetic field induced by the metallic implant. A new method has been designed and implemented to compute the magnetic field induced by metallic objects of arbitrary shape. The magnetic field is expressed pointwise in terms of a surface integral. Efficient quadrature schemes are proposed to evaluate this integral. Finally, the method is linked to an artifact reconstruction model to simulate the images. Magn Reson Med 45:724-727, 2001. PMID- 11284005 TI - [Psi(+)] prion generation in yeast: characterization of the 'strain' difference. AB - The yeast cytoplasmically-inherited nonsense suppressor [PSI(+)] determinant is presumed to be a manifestation of the aggregated prion-like state of the Sup35 protein. Overexpression of the Sup35 protein induces generation of [PSI(+)] determinants with various suppressor efficiency and mitotic stabilities. Here, we demonstrate that the relative frequency of appearance of [PSI(+)] with different properties depends on the SUP35 allele used to induce their generation. The difference in properties of [PSI(+)] determinants was preserved after their transmission from one yeast strain to another. This difference correlated with variation in properties of the Sup35 protein. A novel type of prion instability was observed: some [PSI(+)] with weak suppressor efficiency could convert spontaneously into strong suppressor determinants. PMID- 11284006 TI - Chronological lifespan of stationary phase yeast cells; a model for investigating the factors that might influence the ageing of postmitotic tissues in higher organisms. AB - Budding yeast can be considered to have two distinct lifespans: (a) a replicative (budding, non-chronological) lifespan, measured as the number of daughters produced by each actively dividing mother cell; and (ii) a chronological lifespan, measured as the ability of stationary cultures to maintain viability over time. In non-dividing cells, essential components that become damaged cannot be diluted out through cell division but must, of necessity, be turned over and renewed. By elevating stress resistances, many of the activities needed for such renewal should be elevated with commensurate reduction in the steady-state levels of damaged cell components. Therefore, chronological lifespan in particular might be expected to relate to stress resistance. For yeast to attain a full chronological lifespan requires the expression of the general stress response. It is more important, though, that the cells should be efficiently adapted to respiratory maintenance, since it is cultures grown to stationary phase on respiratory media that usually display the longest chronological lifespans. For this reason, respiration-adapted cells potentially provide a better model of chronological ageing than cultures pre-grown on glucose. PMID- 11284007 TI - The activity of plasma membrane H(+)-ATPase is strongly stimulated during Saccharomyces cerevisiae adaptation to growth under high copper stress, accompanying intracellular acidification. AB - For the adaptation of cells of Saccharomyces cerevisiae, a period of latency is necessary before exponential growth is resumed in a medium supplemented with a highly inhibitory concentration of copper. In this work, we have examined some physiological responses occurring during this period of adaptation. The results revealed that plasma membrane H(+)-ATPase (PM-ATPase) activity is strongly stimulated (up to 24-fold) during copper-induced latency in growth medium with glucose, reaching maximal levels when the cells were about to start inhibited exponential growth. This in vivo activation of the ATPase activity by copper was accompanied by the stimulation of the H(+)-pumping activity of the enzyme in vivo and was essentially due to the increase of the apparent V(max) for MgATP. Although the exact molecular basis of the reported plasma membrane ATPase activation was not clarified, no increase in the mRNA levels from the encoding genes PMA1 and PMA2 was apparently detected during copper-induced latency. The physiological response reported here may allow the cells to cope with copper induced lipid peroxidation and consequent decrease in plasma membrane lipid ordering and increase in the non-specific permeability to protons. The consequences of these copper deleterious effects were revealed by the decrease of the intracellular pH (pH(i)) of the yeast population, from approximately pH(i) 6 to pH(i) 5, during copper-induced latency in growth medium at pH 4.3. The time dependent patterns of plasma membrane ATPase activation and of the decrease of pH(i) during the period of adaptation to growth with copper correlate, suggesting that the regulation of this membrane enzyme activity may be triggered by intracellular acidification. Consistent with this idea, when exponential growth under copper stress was resumed and the pH(i) of the yeast population recovered up to physiological values, plasma membrane ATPase activity simultaneously decreased from the highly stimulated level attained during the adaptation period of latency. PMID- 11284008 TI - Assessment of prediction accuracy of protein function from protein--protein interaction data. AB - Functional prediction of open reading frames coded in the genome is one of the most important tasks in yeast genomics. Among a number of large-scale experiments for assigning certain functional classes to proteins, experiments determining protein-protein interaction are especially important because interacting proteins usually have the same function. Thus, it seems possible to predict the function of a protein when the function of its interacting partner is known. However, in vitro experiments often suffer from artifacts and a protein can often have multiple binding partners with different functions. We developed an objective prediction method that can systematically include the information of indirect interaction. Our method can predict the subcellular localization, the cellular role and the biochemical function of yeast proteins with accuracies of 72.7%, 63.6% and 52.7%, respectively. The prediction accuracy rises for proteins with more than three binding partners and thus we present the open prediction results for 16 such proteins. PMID- 11284009 TI - Overexpression of HUT1 gene stimulates in vivo galactosylation by enhancing UDP galactose transport activity in Saccharomyces cerevisiae. AB - Transfer of activated sugar-nucleotides from the cytoplasm to the lumen of the Golgi is an essential requirement for glycosylation of glycoproteins, proteoglycans and glycosphingolipids. Although mannosylation is the major modification in the yeast Saccharomyces cerevisiae, several reports suggest the presence of galactose residues on yeast proteins and sphingolipids. We have detected alpha-galactosylated O-linked chitinase by lectin blotting from cells that functionally express the gma12(+) gene, encoding alpha 1,2 galactosyltransferase from Schizosaccharomyces pombe. This result implies the presence of a UDP-galactose transporter in S. cerevisiae. A conserved gene, HUT1, which encodes a putative multi-transmembrane protein, was cloned and characterized for its possible involvement in galactosylation. The HUT1 gene is not essential and is expressed at a relatively low level under the physiological conditions we examined. The disruption of this gene did not show any apparent impairments in glycosylation. However, a temperature- and concentration-dependent increase in UDP--galactose transport activity was detected from cells overexpressing HUT1 in the presence of gma12(+). The surface of these cells was confirmed to carry galactose residues by staining with FITC-conjugated alpha galactose-specific lectin. These results suggest a role for Hut1p in the transport of UDP--galactose from the cytosol into the Golgi lumen in S. cerevisiae. PMID- 11284010 TI - Hut1 proteins identified in Saccharomyces cerevisiae and Schizosaccharomyces pombe are functional homologues involved in the protein-folding process at the endoplasmic reticulum. AB - The Saccharomyces cerevisiae HUT1 gene (scHUT1) and the Schizosaccharomyces pombe hut1(+) gene (sphut1(+)) encode hydrophobic proteins with approximately 30% identity to a human UDP-galactose transporter-related gene (UGTrel1) product. These proteins show a significant similarity to the nucleotide sugar transporter and are conserved in many eukaryotic species, but their physiological functions are not known. Both scHUT1 and sphut1(+) genes are non-essential for cell growth under normal conditions, and their disruptants show no defects in the modification of O- and N-linked oligosaccharides, but are sensitive to a membrane permeable reducing agent, dithiothreitol (DTT). Consistent with this phenotype, scHUT1 has genetic interaction with ERO1, which plays an essential role in the oxidation of secretory proteins at the endoplasmic reticulum (ER). Overexpression of the MPD1 or MPD2 genes, which were isolated as multicopy suppressors of protein disulphide isomerase (PDI) depletion, could not replace the essential function of PDI in Delta hut1 S. cerevisiae cells. Our results indicate that scHut1p and spHut1p are functional homologues, and their physiological function is to maintain the optimal environment for the folding of secretory pathway proteins in the ER. PMID- 11284011 TI - Isolation of a gene encoding a putative polyamine transporter from Candida albicans, GPT1. AB - A gene encoding a transport protein from the pathogenic yeast, Candida albicans, has been isolated during a complementation experiment utilizing an ornithine decarboxylase-negative (spe1 Delta) strain of Saccharomyces cerevisiae. This gene restores gamma-aminobutyric acid (GABA) transport to a GABA transport-negative mutant of S. cerevisiae and encodes a protein which putatively allows transport of one or more of the polyamines. We have assigned the name GPT1 (GABA/polyamine transporter) to this gene. PMID- 11284012 TI - A new family of yeast vectors and S288C-derived strains for the systematic analysis of gene function. AB - The yeast genome has been shown to contain a significant number of gene families with more than three members. In order to study these families it is often necessary to generate strains carrying deletions of all members of the family, which can require a wide range of auxotrophic markers. To facilitate such studies, we have generated yeast strains containing deletions of a selection of nutritional marker genes (ade2, ade4, ade8, met3 and met14). We have also cloned the corresponding cognate genes, allowing their use in PCR-based gene disruptions. Two new pRS family Saccharomyces cerevisiae-Escherichia coli shuttle vectors containing ADE8 (one low-copy, pRS4110, and one high-copy, pRS4210) have been produced for use in conjunction with the new strains. A system for easier synthetic lethal screening using one of these new markers is also presented. The ADE8 and HIS3 genes have been cloned together on a high-copy vector (pRS4213), providing a plasmid for red-white colour screening in the ade2 Delta 0 ade8 Delta 0 strains we have generated. In contrast to some conventional systems, this plasmid allows for screening using gene libraries constructed in URA3 plasmids. PMID- 11284013 TI - Current awareness on yeast. PMID- 11284014 TI - Expression of the neural RNA-binding protein Musashi1 in human gliomas. AB - Tumor cells arising from a particular tissue may exhibit the same gene expression patterns as their precursor cells. To test this proposition, we have analyzed the expression of a neural RNA-binding protein, Musashi1, in primary human central nervous system (CNS) tumors. In rodents, Musashi1 is expressed predominantly in proliferating multipotent neural precursor cells, but not in newly generated postmitotic neurons. The expression of Musashi1 is downregulated with the successive progression of neurogenesis. In normal adult human tissues, we detected low levels of Musashi1 expression in brain and testis by RT-PCR analysis. In an RNA panel of 32 cancer tissues and cell lines, elevated expression of Musashi1 was seen in all five malignant gliomas studied, in contrast to the slight expression seen in other tumor cells, including those in several melanomas and a prostate cancer. Western blot analysis showed strong Musashi1 expression in malignant gliomas compared with nonneoplastic brain tissue. Glioblastomas, the most malignant form of glioma, showed higher Musashi1 expression than less malignant gliomas by immunohistochemical analysis. Tumors with strong Musashi1 expression tended to have high proliferative activity. Thus, the expression of Musashi1 correlated with the grade of the malignancy and proliferative activity in gliomas. These results suggest that primary CNS tumors may share gene expression patterns with primitive, undifferentiated CNS cells and that Musashi1 may be a useful marker for the diagnosis of CNS tumors. PMID- 11284015 TI - Retroviral labeling of Schwann cells: in vitro characterization and in vivo transplantation to improve peripheral nerve regeneration. AB - Transplantation of Schwann cells (SCs) is a promising treatment modality to improve neuronal regeneration. Identification of the transplanted cells is an important step when studying the development of this method. Genetic labeling is the most stable and reliable method of cell identification, but it is still unclear whether it has deleterious effect on SC characteristics. Our aim was to achieve a stable population of SCs transduced with the lacZ gene at a high frequency using a retroviral vector in vitro, and to follow the labeled SC in vitro to assess their viability and phenotypic marker expression. Furthermore, we transplanted lacZ-labeled SCs in a conduit to repair peripheral nerve to investigate their effect on nerve regeneration in vivo. Rat and human SCs were cultured and transduced with an MFG lacZ nls marker gene, achieving a transduction rate of 80% and 70%, respectively. Rat SCs were kept in culture for 27 weeks and examined every 4 weeks for expression of lacZ, viability, and phenotypic marker expression of GFAP, p75, MHC I and II. Throughout this period, transduced rat SCs remained viable and continued to proliferate. The proportion of cells expressing lacZ dropped only by 10% and the expression of phenotypic markers remained stable. Transduced human SCs were followed up for 4 weeks in culture. They proliferated and continued to express the lacZ gene and phenotypic marker expression of GFAP and p75 was preserved. Primary culture of transduced rat SCs were transplanted, syngeneically, in a conduit to bridge a 10 mm gap in sciatic nerve and the grafts were examined after 3 weeks for the presence and participation of labeled SCs and for axonal regeneration distance. Transplanted transduced rat SCs were clearly identified, taking part in the regeneration process and enhancing the axonal regeneration rate by 100% (at the optimal concentration) compared to conduits without SCs. Thus, retroviral introduction of lacZ gene has no deleterious effect on SCs in vitro and these SCs take part and enhance nerve regeneration in vivo. PMID- 11284016 TI - Role of the astrocytic ET(B) receptor in the regulation of extracellular endothelin-1 during hypoxia. AB - Astrocytes are known to possess an effective endothelin (ET) eliminatory system which involves astrocytic ET(A) and ET(B) receptors and may become particularly relevant under pathophysiological conditions. The present study has therefore been designed to explore the effect of standardized hypoxia on extracellular concentrations of endothelin-1 (ET-1) and on endothelin-converting enzyme (ECE) activity in primary rat astrocytes genetically (sl/sl) or experimentally (dexamethasone) deficient in ET(B) receptors. The results revealed (1) a hypoxia mediated decrease of extracellular ET-1 in wildtype astrocytes (+/+) that was not observed in ET(B)-deficient (sl/sl) cultures; (2) an ET receptor antagonist induced increase in ET-1 in the media of both genotypes with further elevation upon hypoxia in +/+ cultures only; (3) augmentation of the dexamethasone-induced increase in extracellular ET-1 by hypoxia in +/+, but not in sl/sl cultures; (4) synergistic reduction of ET(B) gene transcription by hypoxia and dexamethasone; and (5) significant increases in endothelin-converting enzyme activity in the presence of hypoxia. To conclude, hypoxia stimulates astrocytic release of mature ET-1. This stimulation is (over)compensated for by increased ET-1 binding to functional ET(B) receptors. ET(B) deficiency, whether genetic or experimentally induced, impairs elimination of extracellular ET-1. PMID- 11284017 TI - Visualising the activity of the cystine-glutamate antiporter in glial cells using antibodies to aminoadipic acid, a selectively transported substrate. AB - The cystine-glutamate antiporter is a transport system that facilitates the uptake of cystine, concomitant with the release of glutamate. The cystine accumulated by this transporter is generally considered for use in the formation of the cysteine-containing antioxidant glutathione, which is abundant in many glial cells. This study used the simple strategy of generating an antibody to aminoadipic acid, a selective substrate for the cystine-glutamate antiporter. Stereospecific accumulation of aminoadipic acid into specific cell types in rat brain slice preparations was detected immunocytochemically. Strong accumulation was detected in astroglial cells in all brain regions studied including those in white matter tracts. Strong accumulation into radial glial cells, including the retinal Muller cells and the Bergmann glial cells was also observed. Glial accumulation was observed not only in cells within the blood brain barrier, but also outside such; anterior pituitary folliculostellate cell and intermediate lobe pituitary glial cells exhibited strong accumulation of aminoadipic acid. Interestingly, some glial cells such as the posterior pituitary glial cells (pituicytes) exhibited very little if any accumulation of aminoadipic acid. Within the brain labelling was not uniform. Particularly strong labelling was noted in some regions, such as the glial cells surrounding the CA1 pyramidal cells. By contrast, neurons never exhibited uptake of aminoadipic acid. Because cystine uptake is associated with glutamate release, it is suggested that this antiporter might contribute to release of glutamate from glial cells under some pathophysiological conditions. PMID- 11284018 TI - The neuron-glia signal beta-neuregulin promotes Schwann cell motility via the MAPK pathway. AB - Neuregulins constitute a family of related growth factors that play important roles in Schwann cell development and maturation. We investigated the involvement of beta-neuregulin in Schwann cell migration, using a simple in vitro bioassay. Pure Schwann cells were prepared from the sciatic nerves of 5-day-old rats and were grown in defined medium, with or without serum, until a monolayer of confluent cells was formed. A cell-free area was then generated by inflicting a scratch resulting in a 1-mm-wide gap. Schwann cell migration within the gap was monitored microscopically at given time intervals and was quantified using an image analysis system. The extent of cell proliferation was estimated by BrdU incorporation, and cell migration was quantified both in the absence and presence of cytosine arabinoside. We found that, in the absence of serum, beta-neuregulin at a dose submaximal for proliferation increased the rate of Schwann cell migration by 84%. A more moderate effect was observed when beta-neuregulin was applied in the presence of serum which, however, is by itself responsible for increased Schwann cell motility. To assess the signal transduction pathways involved in this procedure we used one inhibitor of MAPK, PD098059, two inhibitors of PI-3-kinase, wortmannin, and LY0294002, and three different PKC inhibitors. Of these PD098059 inhibited the neuregulin-induced enhancement in Schwann cell migration by 40%, the two PI-3-kinase inhibitors yielded an approximately 20% inhibition while the PKC inhibitors were ineffective. Our data indicate that the action of beta-neuregulin on Schwann cell motility is primarily mediated via the MAPK pathway. PMID- 11284019 TI - pH-sensitive inwardly rectifying chloride current in cultured rat cortical astrocytes. AB - The effect of pH(o) on plasma membrane chloride current of cultured rat cortical astrocytes was investigated using the whole-cell patch-clamp technique. In the presence of intra- and extracellular solutions with symmetrical high Cl(-) content and K(+) channel inhibitors, the cells exhibited an inwardly rectifying current. The current activated slowly at potentials negative to -40 mV and did not display time-dependent inactivation. The current was inhibited by 0.1 mM Cd(2+), 0.1 mM Zn(2+), 1 mM 9-anthracene-carboxylic acid, and 0.2 mM 5-nitro-2-(3 phenylpropylamino)benzoic acid, but not by 10 mM Ba(2+) or 3 mM Cs(+). Reversal potential of the current followed the chloride equilibrium potential and was not influenced by changes in K(+) or Na(+) concentration. The inwardly rectifying chloride current was augmented by extracellular acidosis and reduced by alkalosis. The pH sensitivity was most pronounced in the physiologically relevant pH(o) range of 6.9--7.9. Lowering pH to 6.4 induced no additional increase in steady-state current amplitude compared with pH(o) 6.9, but it substantially slowed the activation kinetics. According to its kinetic and pharmacological properties this chloride current is similar to that found in cultured rat astrocytes after long-term treatment with dibutyryl-cAMP, however, in our cultures it was consistently expressed without any treatment with the drug. Considering that astrocytes possess carbonic anhydrase and Cl(-)/HCO3(-) antiporter, this current may participate in the regulation of the interstitial and astrocyte pH. PMID- 11284020 TI - Serum deprivation and NGF induce and modulate voltage-gated Na(+) currents in human astrocytoma cell lines. AB - Glial tumor cells derived from primary tissue express large voltage-gated Na(+) currents, whereas glioma cell lines usually lack this feature. We studied the effect of serum deprivation on the expression of Na(+) currents in two astrocytoma cell lines (1321N1 and A172). Serum deprivation for more than 2 days sufficed to induce large Na(+) currents in both cell lines; 300 nM of the specific blocker of voltage-gated Na(+) channels, tetrodotoxin, blocked these currents by about 85%. During serum deprivation, the cells also underwent morphological changes that were characterized by cell rounding and outgrowth of processes. Treatment with 100 ng/ml nerve growth factor (NGF) promoted these morphological changes and also accelerated the development of Na(+) currents. In 1321N1 cells, NGF increased the Na(+) current density after short serum deprivation (3--6 d) and changed several gating properties after longer serum deprivation (9--13 d). In comparison with cells from the early culture stage (3- 6 d), the steady-state inactivation of the Na(+) current was shifted by -24 mV in NGF-treated cells from the late (9--13 d) culture stage. In untreated cells, this shift was only -13 mV. NGF accelerated the kinetics of inactivation and shifted the current-voltage relationship in cells from the late culture stage by -14 mV. In A172 cells, most of these effects were present already after short serum deprivation either in presence or absence of NGF. It is concluded that in astrocytoma cells, Na(+) currents are induced by serum deprivation and are modulated by NGF. This result supports the idea that NFG controls Na(+) currents in these cells by autocrine stimulation. PMID- 11284021 TI - Astrocytes contribute to neuronal impairment in beta A toxicity increasing apoptosis in rat hippocampal neurons. AB - Astrocytosis is a common feature of amyloid plaques, the hallmark of Alzheimer's disease (AD), along with activated microglia, neurofibrillary tangles, and beta amyloid (beta A) deposition. However, the relationship between astrocytosis and neurodegeneration remains unclear. To assess whether beta A-stimulated astrocytes can damage neurons and contribute to beta A neurotoxicity, we studied the effects of beta A treatment in astrocytic/neuronal co-cultures, obtained from rat embryonic brain tissue. We found that in neuronal cultures conditioned by beta A treated astrocytes, but not directly in contact with beta A, the number of apoptotic cells increased, doubling the values of controls. In astrocytes, beta A did not cause astrocytic cell death, nor did produce changes in nitric oxide or prostaglandin E(2) levels. In contrast, S-100 beta expression was remarkably increased. Our data show for the first time that beta A--astrocytic interaction produces a detrimental effect on neurons, which may contribute to neurodegeneration in AD. PMID- 11284022 TI - Evaluation of the contribution to enantioselectivity of quinine and quinidine scaffolds in chemically and physically mixed chiral selectors. AB - Three "dimeric" C(9)-carbamates of quinine (QN) and quinidine (QD), that is, QN QN, QD-QD, and QN-QD (chemically prepared mixture of the two cinchona-derived subunits), separated by an ethylene spacer were synthesized and used as chiral selectors for HPLC and capillary electrophoresis (CE) for the resolution of chiral acids. The chiral recognition abilities of these dimers and of several physically prepared mixtures thereof were compared in order to estimate the contribution of every cinchona scaffold to the overall enantioselectivity. The diverse phenomena observed in nonaqueous capillary electrophoresis (NACE), either using the selector added to the background electrolyte (BGE) in the total filling or partial filling mode, led us to rationalize, taking into account the relative mobilities of the chiral selectors in the capillary. The chromatographic and electrophoretic properties were compared with those of the corresponding "monomeric" QN and QD carbamates. PMID- 11284023 TI - Selective antibodies to methadone enantiomers: synthesis of (R)- and (R,S) methadone conjugates and determination by an immunoenzymatic method in human serum. AB - Selective antibodies to (R)-methadone (Mtd) and to its racemate were produced in rabbits by immunization with conjugates of (R)- or (R,S)-hemisuccinyl-methadol bovine serum albumin, respectively. A hapten was first prepared by reduction of (R)- or (R,S)-Mtd with sodium borohydride, followed by esterification with succinic anhydride. The conjugation of hapten with albumin was achieved by the mixed anhydride method. After immunization of rabbits, the titers and specificity of each antibody were determined by ELISA. The antibodies obtained were tested with (R)-, (S)-, (R,S)-Mtd, its major metabolite (EDDP), and some drugs of abuse (morphine, codeine, cocaine). The sensitivities of antibodies to (R)- and (R,S) Mtd were about 1 and 2 ng/ml, respectively. Selective (R)-antibodies recognized (R)-Mtd about 40 times more avidly than the (S)-isomer, while an antiserum against (R,S)-Mtd recognized (R)- and (S)-isomers to about the same degree. Both selective antibodies showed little interference (about 0.5%) with EDDP metabolite and no crossreactivity with morphine, codeine, and cocaine. These two selective antibodies were used to develop an immunoenzymatic method (ELISA) for the determination of (R)- and (R,S)-Mtd in serum samples of patients under maintenance treatment for narcotic addiction. PMID- 11284024 TI - Column selection and method development for the determination of the enantiomeric purity of investigational non-nucleoside reverse transcriptase inhibitors. AB - DPC 961 and DPC 083 are investigational non-nucleoside reversed transcriptase inhibitors (NNRTI) being evaluated for the treatment of HIV infections (Corbett et al., Antimicrob Agents Chemother 1999;43:2893-2897). Both compounds are chiral and are synthesized as single enantiomers by an asymmetric synthetic pathway (Magnus et al., Tetrahedron Lett 2000;41:3015-3019). A chiral method was developed to control the enantiomeric purity of the drug substance and to monitor for any chiral inversion in the drug substance and in the tablet formulation during stability studies. Three columns were evaluated: Chiralpak AD, Chirobiotic V, and Whelk-O. All three columns have broad applicability and can resolve enantiomers of compounds of very diverse molecular structure and polarity. The three columns were evaluated with various mobile phase compositions for their ability to resolve the racemic mixtures of DPC 083, DPC 961, and their respective enantiomers and for their selectivity toward the synthetic impurities of the two drug substances. Nonaqueous mobile phases were selected because the two drugs are poorly water soluble. The separation between the unwanted enantiomer of DPC 083 and DPC 961, which is a major impurity of DPC 083, in particular, was closely monitored. The final method was fully validated and is used for the routine testing of the drug substance and tablets. PMID- 11284025 TI - Melatonin receptor agents: synthesis, resolution by HPLC on polysaccharides chiral stationary phases, absolute configuration, and pharmacology of the enantiomers of (+/-)-N-[[2[(7-fluoro-1,2,3,4-tetrahydronaphthalen-1 yl)ethyl]acetamide. AB - In order to obtain milligram amounts of the enantiomers of tetrahydronaphthalenic derivative 5 to be tested for binding to the melatonin sites, preparative HPLC employed a mobile phase consisting of n-hexane-alcohol and a silica-based cellulose tris-methylbenzoate (Chiralcel OJ) using isocratic conditions and multiple repetitive injections. The preparative separation was optimized by adjusting the sample size from a scale-up of the analytical method. The enantiomeric elution order was reversed by the change from the carbamate type phase (Chiralcel OD-H) to the benzoate type phase (Chiralcel OJ) in analytical mode. The optical rotation and the circular dichroism spectra of the single enantiomers were determined after separation. The absolute stereochemistry of the two enantiomers of (+/-)-N-[2-(7-fluoro-1,2,3,4-tetrahydronaphthalen-1 yl)ethyl]acetamide 5 was established by X-ray crystallographic analysis. The purity obtained was sufficient for a first screen of their biochemical properties: the (-)-(S) enantiomer shows more affinity for melatonin receptors MT1, MT2 and is responsible of the selectivity towards MT2. PMID- 11284026 TI - Intestinal absorption and metabolism of chlorpheniramine enantiomers in rat. AB - Chlorpheniramine (CPAM) is a chiral antihistaminic drug commercialized as a racemic mixture. The intestinal absorption and metabolism of CPAM have been investigated in rat using in vivo (oral and IV administration), in situ (intestinal loop model), and in vitro (everted sac model) experiments. Oral and IV administrations of 20 mg/kg of the racemic mixture show that the pharmacokinetics of CPAM are stereoselective, with higher AUCs for the (+)-S enantiomer compared to its antipode. The monodesmethyl metabolite (DCPM) was quantifiable in blood and its pharmacokinetics are stereoselective after oral but not after IV administration. Experiments using intestinal loops and everted sacs showed that the absorption is not stereoselective and that in vivo stereoselective formation of DCPM is presumably due to stereoselective hepatic metabolism. Moreover, the in vitro and in situ absorption of CPAM are not modified by modulators of P-glycoprotein and cytochromes P450 (cyclosporin A, ketoconazole). PMID- 11284027 TI - First direct discrimination of chiral phosphorus thionate (P=S) derivatives by multinuclear magnetic resonance spectroscopy in the presence of a chiral dirhodium complex. AB - Enantiomeric ratios of compounds with P=S functionalities can be determined by 1H, 13C, and 31P NMR spectroscopic inspection of their diastereomeric complexes with (R)-Rh2(MTPA)4 (MTPA-H identical with methoxytrifluoromethylphenylacetic acid; Mosher's acid). This is the first facile and rapid spectroscopic method for chiral recognition in this class of compounds. Whereas complexation shifts Deltadelta are moderate or even negligible, significant signal dispersions Delta(nu) can be observed. Some rationalization about the complexation mode is presented. The NMR spectral characteristics of the free P=S compounds 1-9 are described in detail. PMID- 11284028 TI - Is (9Z)-"meso"-zeaxanthin optically active? AB - The question raised in the title was answered. (3R, 3'S)-meso-Zeaxanthin was submitted to iodine catalyzed photochemical stereoisomerisation. The enantiomeric (9Z) and (9'Z) geometrical isomers were isolated by semipreparative HPLC and separated as diastereomeric dicarbamates on a chiral column only. Cleavage of the carbamate could not be effected. CD-Spectra of (1"S, 1"S)- and (1"R, 1"R) dicarbamates of geometrical isomers of (3R, 3'R)- and (3R, 3'S)-meso-zeaxanthin were systematically studied and the contribution from the carbamate moieties revealed. It was concluded that (9Z, 3R, 3'S)-"meso"-zeaxanthin, in spite of having no symmetry elements, is optically inactive. The result has been rationalised in line with the current hypothesis on the origin of carotenoid CD spectra. PMID- 11284029 TI - Molecular cytogenetic characterization of early and late renal cell carcinomas in von Hippel-Lindau disease. AB - Deletions of 3p25, gains of chromosomes 7 and 10, and isochromosome 17q are known cytogenetic aberrations in sporadic renal cell carcinoma (RCC). In addition, a majority of RCCs have loss of heterozygosity (LOH) of the Von Hippel-Lindau (VHL) gene located at chromosome band 3p25. Patients who inherit a germline mutation of the VHL gene can develop multifocal RCCs and other solid tumors, including malignancies of the pancreas, adrenal medulla, and brain. VHL tumors follow the two-hit model of tumorigenesis, as LOH of VHL, a classic tumor suppressor gene, is the critical event in the development of the neoplastic phenotype. In an attempt to define the cytogenetic aberrations from early tumors to late RCC further, we applied spectral karyotyping (SKY) to 23 renal tumors harvested from 6 unrelated VHL patients undergoing surgery. Cysts and low-grade solid lesions were near-diploid and contained 1-2 reciprocal translocations, dicentric chromosomes, and/or isochromosomes. A variety of sole numerical aberrations included gains of chromosomes 1, 2, 4, 7, 10, 13, 21, and the X chromosome, although no tumors had sole numerical losses. Three patients shared a breakpoint at 2p21-22, and three others shared a dicentric chromosome 9 or an isochromosome 9q. In contrast to the near-diploidy of the low-grade lesions, a high-grade lesion and its nodal metastasis were markedly aneuploid, revealed loss of VHL by fluorescence in situ hybridization (FISH), and contained recurrent unbalanced translocations and losses of chromosome arms 2q, 3p, 4q, 9p, 14q, and 19p as demonstrated by comparative genomic hybridization (CGH). By combining SKY, CGH, and FISH of multiple tumors from the same VHL kidney, we have begun to identify chromosomal aberrations in the earliest stages of VHL-related renal cell tumors. Our current findings illustrate the cytogenetic heterogeneity of different VHL lesions from the same kidney, which supports the multiclonal origins of hereditary RCCs. Published 2001 Wiley-Liss, Inc. PMID- 11284030 TI - Integration of amplified BCR/ABL fusion genes into the short arm of chromosome 17 as a novel mechanism of disease progression in chronic myeloid leukemia. AB - We describe the cases of two patients with Philadelphia chromosome-positive chronic myeloid leukemia (CML), in whom the extramedullary blastic phase developed during disease progression. The similar clinical presentations of these patients was accompanied by gain of identical secondary chromosome abnormalities, that is, monosomies 9, 14, and 22, and by a clustered amplification of the BCR/ABL fusion gene. The additional copies of the BCR/ABL fusion gene were integrated into the short arm of structurally abnormal chromosomes 17 in both patients. The conformity of these genetic features in two patients with a rare disease manifestation leads us to the assumption that either the clustered amplification of the BCR/ABL fusion gene or the integration of this cluster into the short arm of chromosome 17 or both are associated with extramedullar disease progression in CML. Furthermore, the insertion of amplified BCR/ABL fusion genes into structurally abnormal chromosomes provides a novel mechanism of disease progression in BCR/ABL-positive CML. PMID- 11284031 TI - Identification of extensive genomic loss and gain by comparative genomic hybridisation in malignant astrocytoma in children and young adults. AB - Although astrocytomas are the most common central nervous system tumours in all age groups, there is substantial evidence that tumours arising in young patients (< 25 years of age) do not have the same genetic abnormalities that are characteristic of tumours in older patients. Furthermore, novel, consistent changes have not been identified in astrocytomas in children and young adults. We analysed 13 malignant astrocytomas from young patients using comparative genomic hybridisation. Regions of genomic imbalance were identified in 10 cases. The most common recurrent copy number aberrations were loss of 16p (54% of cases), 17p (38%), 19p (38%), and 22 (38%) and gain on 2q (38%), 12q (38%), 13 (38%), 4q (31%), 5q (31%), and 8q (31%). Seven regions of high copy number amplification were observed at 8q21-22 (three cases), 7q22-23 (two cases), and 1p21-22, 2q22, 12q13-pter, 12q15-21, and 13q11-14 (one case each). This study provides evidence of new characteristic chromosomal imbalances from which potential candidate genes involved in the development of malignant astrocytoma in children and young adults may be identified. PMID- 11284032 TI - Molecular analysis of a familial case of renal cell cancer and a t(3;6)(q12;q15). AB - We identified a novel familial case of clear-cell renal cancer and a t(3;6)(q12;q15). Subsequent cytogenetic and molecular analyses showed the presence of several abnormalities within tumour samples obtained from different patients. Loss of the der(3) chromosome was noted in some, but not all, of the samples. A concomitant VHL gene mutation was found in one of the samples. In addition, cytogenetic and molecular evidence for heterogeneity was obtained through analysis of several biopsy samples from one of the tumours. Based on these results and those reported in the literature, we conclude that loss of der(3) and subsequent VHL gene mutation may represent critical steps in the development of renal cell cancers in persons carrying the chromosome 3 translocation. Moreover, preliminary data suggest that other (epi)genetic changes may be related to tumour initiation. PMID- 11284033 TI - Duplication or amplification of chromosome band 11q23, including the unrearranged MLL gene, is a recurrent abnormality in therapy-related MDS and AML, and is closely related to mutation of the TP53 gene and to previous therapy with alkylating agents. AB - Gene amplification is a rare phenomenon in acute leukemia, but recently amplification of specific chromosome bands containing genes rearranged in leukemia-specific balanced chromosome translocations has been reported in a few cases. We detected duplication or amplification of chromosome band 11q23 with 3-7 copies of the MLL gene by fluorescence in situ hybridization in 12 out of 70 unselected patients with therapy-related myelodysplasia or acute myeloid leukemia (17%). In all but one case, the supernumerary copies of MLL were located to previously unidentified marker chromosomes or unbalanced translocations. In 4 of the 12 patients, 2-6 copies were located together on the same chromosome arm representing amplification, 7 patients had single, extra duplicated copies of MLL, whereas both amplification and duplication were observed in the same cell in 1 patient. Comparative genomic hybridization demonstrated gain of varying, often large parts of 11q in five patients. The MLL gene was shown to be unrearranged in all 12 patients. Seven out of eight patients with duplication or amplification of MLL had mutations of TP53. Patients with supernumerary copies of MLL were in general older (P = 0.007) and had a shorter survival (P < 0.001) compared to other patients. Duplication or amplification of MLL was significantly associated with a complex karyotype (P = 0.002), with deletion or loss of 5q (P = 0.001), and with prior therapy with alkylating agents. These results support the existence of a specific genetic pathway in t-MDS and t-AML with many previously unidentified chromosome aberrations demonstrated to represent extra copies of parts of 11q, including the unrearranged MLL gene. PMID- 11284034 TI - Refinement within single yeast artificial chromosome clones of a minimal region commonly deleted on the short arm of chromosome 7 in Wilms tumours. AB - Cytogenetic and molecular data indicate an involvement of genes mapped to the proximal portion of the short arm of chromosome 7 (7p) in Wilms tumours (WTs). We have analysed 38 WTs using a panel of eight microsatellite markers mapped to proximal 7p. Loss of heterozygosity (LOH) in tumour, compared with matched constitutional DNA, was identified in eight cases. To define better the minimal region commonly deleted in these tumours, they were analysed with nine additional markers, mapped within the region of interest. One tumour (case 30) showed LOH for only one marker (D7S510), while maintaining heterozygosity for the two immediately flanking loci (D7S555 and D7S668). This result was confirmed by fluorescence in situ hybridisation analysis, which showed that in the majority (65%) of nuclei from tumour 30 hybridising with a bacterial artificial chromosome clone containing the D7S510 locus, only one signal was visible. Noticeably, both markers defining the limits of the observed deleted region are simultaneously present within two distinct overlapping yeast artificial chromosome (YAC) clones mapped to chromosome bands 7p13-p14. This suggests that the maximum length of the missing DNA fragment was approximately 1.3 Mb, corresponding to the length of the smaller of the two YAC clones. In all other cases that showed LOH, the deletion encompassed the 7p13-p14 region. For this reason, we speculate that the identified interval contains a gene whose inactivation is important for the development of at least a fraction of WTs. PMID- 11284035 TI - Genetic imbalances in 26 cases of penile squamous cell carcinoma. AB - To obtain more information on chromosomal changes in the up-to-now poorly studied tumor class of penile squamous cell carcinoma (SCC), we performed a comparative genomic hybridization study of 26 cases of this rare tumor. DNA sequence copy number alterations (CNAs) very similar to those detected in other SCC types, such as oral and esophageal SCC, were noted. The most common copy number gains were found in 8q24, 16p11-12, 20q11-13, 22q, 19q13, and 5p15, and the most common deletions were detected in 13q21-22, 4q21-32, and along the X chromosome. Classifying the patients according to the number of CNAs showed a possible correlation with clinical outcome. PMID- 11284036 TI - Characterization of chromosome aberrations associated with soft-tissue leiomyosarcomas by twenty-four-color karyotyping and comparative genomic hybridization analysis. AB - Data on the chromosome aberrations associated with leiomyosarcomas of soft tissues are limited, complex, and incomplete. The aim of this study was to characterize genetic aberrations associated with this tumor group, to identify consistent regions of involvement and to determine correlations with clinical outcome. Chromosomes were prepared from 10 primary soft-tissue leiomyosarcoma samples, and preparations from four of them, plus the cell line SK-LMS-1, were suitable for analysis using 24-color karyotyping by multifluor fluorescence in situ hybridization. This method allowed rearranged chromosomes to be characterized, which would not have been possible by banding analysis alone. The remaining six chromosome preparations were analyzed using standard Giemsa banding. The chromosome imbalances associated with all the samples were determined by comparative genomic hybridization analysis. Taken together, the results show both intra- and intertumor heterogeneity and considerable complexity. Although no highly consistent rearrangements were found, some regions of the genome frequently were involved, including 1q21, 5p14-pter, and 20q13, which likely harbor genes that play a role in the pathogenesis of soft-tissue leiomyosarcomas. There were no obvious correlations between the chromosomal changes identified and available clinical details. PMID- 11284037 TI - A classification efficiency test of spectral karyotyping and multiplex fluorescence in situ hybridization: identification of chromosome homologies between Homo sapiens and Hylobates leucogenys. AB - Two digital fluorescence microscopy systems, spectral karyotyping (SKY) and multiplex fluorescence in situ hybridisation (M-FISH), are used with multicolour probe sets to assist in the detection of chromosome aberrations. We have compared the resolution of the two methods in their ability to identify karyotype rearrangements, which have occurred during the divergence of Homo sapiens and Hylobates leucogenys in evolution. A 24-color human paint kit distinguishes 74 conserved autosomal segments in H. leucogenys, some of which are difficult to resolve. We examined the extent to which the SKY and M-FISH techniques are able to detect the smallest of these bands. We have found this to be a rigorous test of multicolour chromosome classification systems. We conclude from our results that both systems are able invariably to classify the majority of conserved segments but differ in the efficiency of detection of small inserts. PMID- 11284038 TI - Color bar coding the BRCA1 gene on combed DNA: a useful strategy for detecting large gene rearrangements. AB - Genetic linkage data have shown that alterations of the BRCA1 gene are responsible for the majority of hereditary breast and ovarian cancers. BRCA1 germline mutations, however, are found less frequently than expected. Mutation detection strategies, which are generally based on the polymerase chain reaction, therefore focus on point and small gene alterations. These approaches do not allow for the detection of large gene rearrangements, which also can be involved in BRCA1 alterations. Indeed, a few of them, spread over the entire BRCA1 gene, have been detected recently by Southern blotting or transcript analysis. We have developed an alternative strategy allowing a panoramic view of the BRCA1 gene, based on dynamic molecular combing and the design of a full four-color bar code of the BRCA1 region. The strategy was tested with the study of four large BRCA1 rearrangements previously reported. In addition, when screening a series of 10 breast and ovarian cancer families negatively tested for point mutation in BRCA1/2, we found an unreported 17-kb BRCA1 duplication encompassing exons 3 to 8. The detection of rearrangements as small as 2 to 6 kb with respect to the normal size of the studied fragment is achieved when the BRCA1 region is divided into 10 fragments. In addition, as the BRCA1 bar code is a morphologic approach, the direct observation of complex and likely underreported rearrangements, such as inversions and insertions, becomes possible. PMID- 11284039 TI - ALK probe rearrangement in a t(2;11;2)(p23;p15;q31) translocation found in a prenatal myofibroblastic fibrous lesion: toward a molecular definition of an inflammatory myofibroblastic tumor family? AB - A prenatal tumor located in the lumbar paravertebral area was discovered during a routine ultrasound examination at 32 weeks of pregnancy and surgically removed at 4 months of life. The histopathological diagnosis was first suggested to be an infantile desmoid fibromatosis. The tumor karyotype showed a three-way translocation involving both chromosomes 2 and a chromosome 11, t(2;11;2)(p23;p15;q31). Fluorescence in situ hybridization with a probe flanking the ALK gene at 2p23 demonstrated a rearrangement, as previously described in inflammatory myofibroblastic tumors (IMTs). In light of the genetic analysis, the histopathological diagnosis was revised to IMT, although inflammatory cells were scarce. IMTs are pseudosarcomatous inflammatory lesions that primarily occur in the soft tissue and viscera of children and young adults. Our report describes for the first time the occurrence of IMT during prenatal life. The ALK rearrangement may represent the molecular definition of a subgroup of mesenchymal tumors, not always with complete morphological features of IMT, similar to the model of EWS rearrangement in the Ewing sarcoma family of tumors. PMID- 11284040 TI - A deletion/insertion mutation in the BRCA2 gene in a breast cancer family: a possible role of the Alu-polyA tail in the evolution of the deletion. AB - Patients with breast and/or ovarian cancer were screened for gross rearrangements in the BRCA2 gene by Southern hybridization, with exon 10 and a fragment of exon 11 used as probes. One breast cancer patient with a positive family history had a 6.2-kb deletion including exons 12 and 13. The deletion breakpoint in intron 11 was in the 3' polyA tail of an Alu element, where a track of approximately 60 adenine nucleotide residues was inserted. Expansion of the Alu-polyA tail may have resulted from polymerase slippage during replication, representing a novel mechanism in which Alu elements mediate deletion/insertion mutations. PMID- 11284041 TI - Apparent human BRCA1 knockout caused by mispriming during polymerase chain reaction: implications for genetic testing. AB - We report an apparent BRCA1 homozygous knockout that, on further analysis, was found to be an artefact of the polymerase chain reaction. This finding has two important implications. First, it challenges results of a previous study that reported a homozygous knockout associated with the same BRCA1 mutation. Second, our findings suggest that mispriming caused by mismatched primers at the site of single-nucleotide polymorphisms, leading to preferential amplification of one allele, may represent a significant proportion of instances of mutation-detection insensitivity. This may have major implications for the sensitivity of all polymerase chain reaction-based mutation-detection methods in clinical genetic testing laboratories. PMID- 11284042 TI - Nonlinear temporal dynamics of the cerebral blood flow response. AB - The linearity of the cerebral perfusion response relative to stimulus duration is an important consideration in the characterization of the relationship between regional cerebral blood flow (CBF), cerebral metabolism, and the blood oxygenation level dependent (BOLD) signal. It is also a critical component in the design and analysis of functional neuroimaging studies. To study the linearity of the CBF response to different duration stimuli, the perfusion response in primary motor and visual cortices was measured during stimulation using an arterial spin labeling technique with magnetic resonance imaging (MRI) that allows simultaneous measurement of CBF and BOLD changes. In each study, the perfusion response was measured for stimuli lasting 2, 6, and 18 sec. The CBF response was found in general to be nonlinearly related to stimulus duration, although the strength of nonlinearity varied between the motor and visual cortices. In contrast, the BOLD response was found to be strongly nonlinear in both regions studied, in agreement with previous findings. The observed nonlinearities are consistent with a model with a nonlinear step from stimulus to neural activity, a linear step from neural activity to CBF change, and a nonlinear step from CBF change to BOLD signal change. PMID- 11284043 TI - Comparison of neuronal and hemodynamic measures of the brain response to visual stimulation: an optical imaging study. AB - The noninvasive mapping of hemodynamic brain activity has led to significant advances in neuroimaging. This approach is based in part on the assumption that hemodynamic changes are proportional to (and therefore constitute a linear measure of) neuronal activity. We report a study investigating the quantitative relationship between neuronal and hemodynamic measures. This study exploited the fact that optical imaging methods can simultaneously provide noninvasive measures of neuronal and hemodynamic activity from the same region of the brain. We manipulated visual stimulation frequency and measured responses from the medial occipital area of 8 young adults. The results were consistent with a model postulating a linear relationship between the neuronal activity integrated over time and the amplitude of the hemodynamic response. The hemodynamic response colocalized with the neuronal response. These data support the use of quantitative neuroimaging methods to infer the intensity and localization of neuronal activity in occipital areas. PMID- 11284044 TI - Sensitivity of prefrontal cortex to changes in target probability: a functional MRI study. AB - Electrophysiological studies suggest sensitivity of the prefrontal cortex to changes in the probability of an event. The purpose of this study was to determine if subregions of the prefrontal cortex respond differentially to changes in target probabilities using functional magnetic resonance imaging (fMRI). Ten right-handed adults were scanned using a gradient-echo, echo planar imaging sequence during performance of an oddball paradigm. Subjects were instructed to respond to any letter but "X". The frequency of targets (i.e., any letter but X) varied across trials. The results showed that dorsal prefrontal regions were active during infrequent events and ventral prefrontal regions were active during frequent events. Further, we observed an inverse relation between the dorsal and ventral prefrontal regions such that when activity in dorsal prefrontal regions increased, activity in ventral prefrontal regions decreased, and vice versa. This finding may index competing cognitive processes or capacity limitations. Most importantly, these findings taken as a whole suggest that any simple theory of prefrontal cortex function must take into account the sensitivity of this region to changes in target probability. PMID- 11284045 TI - A functional MRI study on the neural substrates for writing. AB - Functional neuroanatomy of writing is relatively unknown compared to that of other linguistic processes. This study aimed at identifying brain regions crucial to the process of writing. Using functional magnetic resonance imaging (fMRI), brain hemodynamic activity was examined during three conditions that differentially engaged visual, linguistic, and/or motor functions: (1) writing names of pictures with the right index finger, (2) naming pictures silently, and (3) visually cued finger tapping. A writing minus naming comparison and a writing minus tapping comparison were performed, and brain regions commonly activated in these two contrasts were detected. Our main finding was that such common activation was observed in the anterior part of the left superior parietal lobule, the posterior part of the middle and superior frontal gyri, and the right cerebellum. The parietal and frontal regions were considered to subserve the process of writing as separated from that of naming and finger movements, which is consistent with the classical notion mainly proposed by studies of selective writing deficits called pure agraphia. The right cerebellar activation, on the other hand, was interpreted as the reflection of the execution of complex finger movements required for writing. PMID- 11284046 TI - Spatial and temporal independent component analysis of functional MRI data containing a pair of task-related waveforms. AB - Independent component analysis (ICA) is a technique that attempts to separate data into maximally independent groups. Achieving maximal independence in space or time yields two varieties of ICA meaningful for functional MRI (fMRI) applications: spatial ICA (SICA) and temporal ICA (TICA). SICA has so far dominated the application of ICA to fMRI. The objective of these experiments was to study ICA with two predictable components present and evaluate the importance of the underlying independence assumption in the application of ICA. Four novel visual activation paradigms were designed, each consisting of two spatiotemporal components that were either spatially dependent, temporally dependent, both spatially and temporally dependent, or spatially and temporally uncorrelated, respectively. Simulated data were generated and fMRI data from six subjects were acquired using these paradigms. Data from each paradigm were analyzed with regression analysis in order to determine if the signal was occurring as expected. Spatial and temporal ICA were then applied to these data, with the general result that ICA found components only where expected, e.g., S(T)ICA "failed" (i.e., yielded independent components unrelated to the "self-evident" components) for paradigms that were spatially (temporally) dependent, and "worked" otherwise. Regression analysis proved a useful "check" for these data, however strong hypotheses will not always be available, and a strength of ICA is that it can characterize data without making specific modeling assumptions. We report a careful examination of some of the assumptions behind ICA methodologies, provide examples of when applying ICA would provide difficult-to-interpret results, and offer suggestions for applying ICA to fMRI data especially when more than one task-related component is present in the data. PMID- 11284047 TI - Crystal structure-based studies of cytosolic sulfotransferase. AB - Sulfation is a widely observed biological reaction conserved from bacterium to human that plays a key role in various biological processes such as growth, development, and defense against adversities. Deficiencies due to the lack of the ubiquitous sulfate donor 3'-phosphoadenosine-5'-phosphosulfate (PAPS) are lethal in humans. A large group of enzymes called sulfotransferases catalyze the transfer reaction of sulfuryl group of PAPS to the acceptor group of numerous biochemical and xenochemical substrates. Four X-ray crystal structures of sulfotransferases have now been determined: cytosolic estrogen, hydroxysteroid, aryl sulfotransferases, and a sulfotransferase domain of the Golgi-membrane heparan sulfate N-deacetylase/N-sulfotransferase 1. These have revealed the conserved core structure of the PAPS binding site, a common reaction mechanism, and some information concerning the substrate specificity. These crystal structures introduce a new era of the study of the sulfotransferases. PMID- 11284048 TI - Localization of estradiol-responsive region in the phenobarbital-responsive enhancer module of mouse Cyp2b-10 gene. AB - The mouse Cyp2b-10 gene is inducible by treatment with estradiol as well as so called phenobarbital (PB)-like inducers. To identify 5'-flanking elements responsible for induction by estradiol, we carried out reporter gene assays using a primary mouse hepatocyte culture system. Cyp2b-10 gene-driven luciferase activities were induced by estradiol as well as PB in this system. Deletion analysis demonstrated that the sequence contained within the region from -2331 bp to -2281 bp was responsible for the estradiol-induced luciferase activity. This region corresponds to the core element of PB-responsive enhancer module (PBREM). Several nucleotide mutations in the putative binding sites of the PBREM core element showed that the NR1 site was required for estradiol induction, and the same element was required for PB induction. These results indicate that estradiol induces Cyp2b-10 gene expression via PBREM. PMID- 11284049 TI - The subpopulation of CF-1 mice deficient in P-glycoprotein contains a murine retroviral insertion in the mdr1a gene. AB - A subpopulation of the CF-1 mouse strain is sensitive to neurotoxicity following exposure to avermectins, a family of structurally related antiparasitic agents. This unusual sensitivity is the result of a deficiency in the mdr1a P glycoprotein that normally contributes to a functional blood-brain barrier. Previous studies demonstrated a correlation between P-glycoprotein levels in the brain, intestine, testis, and placenta with an restriction fragment length polymorphism (RFLP) pattern from DNA isolated from the animals. We have demonstrated that only P-glycoprotein derived from the mdr1a gene is deficient in these mice. In this article, we describe the genetic defect in the subpopulation of CF-1 mice resulting in an absence of P-glycoprotein. The data presented describes a reverse transcription--polymerase chain reaction (RT-PCR) protocol that specifically amplifies mdr1a mRNA from tissue and confirms that the P glycoprotein defect results from a truncated mRNA with a deleted exon 23. Genomic amplification and sequencing of the intron between exon 22 and 23 in Pgp deficient animals reveals an insertion of approximately 8.35 kb of DNA at the exon 23 intron--exon junction corresponding to a murine leukemia virus. This insertion results in the aberrant splicing of the mRNA and the loss of exon 23 during RNA processing. PMID- 11284050 TI - Cloning, sequencing, heterologous expression, and characterization of murine cytochrome P450 3a25*(Cyp3a25), a testosterone 6beta-hydroxylase. AB - A full-length cDNA clone encoding a novel form of the cytochrome P450 3A subfamily (Cyp3a-25) has been isolated from a mouse liver cDNA library. The sequence contained 2010 base pairs and encoded a protein with 503 amino acids. The amino acid sequence shared greater identities with rat CYP3A18 (90%) and golden hamster CYP3A10 (81%) sequences than with known mouse sequences (Cyp3a-11, Cyp3a-13, Cyp3a-16, and Cyp3a-41 [68--70%]). CYP3A25 was expressed in the Escherichia coli PCWori(+) expression vector following slight modifications of the N- and C-terminals of the cDNA. The purified CYP3A25 was recognized on an immunoblot by CYP3A1 antibody and has a molecular weight of 50 kD. CYP3A25 was catalytically active in the 6 beta-hydroxylation of testosterone and the N demethylation of benzphetamine and erythromycin. It was demonstrated by RT-PCR that the CYP3A25 mRNA is present in both fetal and adult tissues, including liver, lung, intestines, kidney, and brain. Northern blotting demonstrated that expression is greatest in the liver and small intestine. PMID- 11284051 TI - The effect of dichloroacetic acid and trichloroacetic acid on DNA methylation and cell proliferation in B6C3F1 mice. AB - The chlorine disinfection by-products, dichloroacetic acid (DCA) and trichloroacetic acid (TCA), are carcinogenic in mouse liver. We have previously reported that DCA and TCA induced DNA hypomethylation in mouse liver. In the present study, we determined the temporal association for DNA hypomethylation and cell proliferation. Female B6C3F1 mice were administered daily doses of 500 mg/kg DCA or TCA by gavage and sacrificed at 24, 36, 48, 72, and 96 hours after the first dose. The proliferating cell nuclear antigen-labeling index in the liver was increased at 72 and 96 hours by both DCA and TCA, that is, at 72 hours the index was 1.00 +/- 0.21, 0.51 +/- 0.11, and 0.095 +/- 0.016 for DCA, TCA, and the vehicle control, respectively. The mitotic index was also significantly increased at 96 hours. The promoter region for the c-myc gene was hypomethylated only at 72 and 96 hours and not at the earlier sacrifices. Similarly, the methylation of the c-myc gene in the kidney and urinary bladder was decreased only at 72 and 96 hours. In summary, enhancement of cell proliferation and decreased methylation of the c-myc gene were first observed simultaneously at 72 hours after the start of exposure. Thus, the results support the hypothesis that DCA and TCA induce DNA hypomethylation by inducing DNA replication and preventing the methylation of the newly synthesized strands of DNA. PMID- 11284052 TI - Peroxisome proliferator perfluorodecanoic acid alters glutathione and related enzymes. AB - Previously we have shown that treatment with the peroxisome proliferator perfluorodecanoic acid (PFDA) significantly increased hepatic reduced glutathione (GSH) content without altering the activity of selenium-glutathione peroxidase. In this study we examined some potential mechanisms by which PFDA treatment increases GSH levels. Male Sprague-Dawley rats were given a single injection of 0, 8.8, 17.5, and 35 mg PFDA in corn oil per kg body weight. Twelve days later the effects of PFDA on the activities of enzymes associated with GSH synthesis, utilization, and regeneration were assessed. The results showed that in a dose dependent manner, PFDA treatment significantly decreased the activity of gamma glutamylcysteine synthetase, while the activities of NADPH-generating enzymes, malic enzyme, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase were increased. PFDA treatment also dose dependently decreased cytosolic, but not microsomal, glutathione S-transferase activity, and the activity of glutathione reductase was decreased by the highest dose of PFDA. The data obtained suggest that increased hepatic GSH levels following PFDA treatment may result from increased regeneration and/or decreased utilization. PMID- 11284053 TI - Comparison of iron-catalyzed DNA and lipid oxidation. AB - Lipid and DNA oxidation catalyzed by iron(II) were compared in HEPES and phosphate buffers. Lipid peroxidation was examined in a sensitive liposome system constructed with a fluorescent probe that allowed us to examine the effects of both low and high iron concentrations. With liposomes made from synthetic 1 stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine or from rat liver microsomal lipid, lipid peroxidation increased with iron concentration up to the range of 10 -20 microM iron(II), but then rates decreased with further increases in iron concentration. This may be due to the limited amount of lipid peroxides available in liposomes for oxidation of iron(II) to generate equimolar iron(III), which is thought to be important for the initation of lipid peroxidation. Addition of hydrogen peroxide to incubations with 1--10 microM iron(II) decreased rates of lipid peroxidation, whereas addition of hydrogen peroxide to incubations with higher iron concentrations increased rates of lipid peroxidation. Thus, in this liposome system, sufficient peroxide from either within the lipid or from exogenous sources must be present to generate equimolar iron(II) and iron(III). With iron-catalyzed DNA oxidation, hydrogen peroxide always stimulated product formation. Phosphate buffer, which chelates iron but still allows for generation of hydroxyl radicals, inhibited lipid peroxidation but not DNA oxidation. HEPES buffer, which scavenges hydroxyl radicals, inhibited DNA oxidation, whereas lipid peroxidation was unaffected since presumably iron(II) and iron(III) were still available for reaction with liposomes in HEPES buffer. PMID- 11284054 TI - Uremic neuropathy and the analysis of electrophysiological changes. AB - Fourteen patients with chronic renal failure were clinically examined and electrodiagnostically tested before and after a single session of hemodialysis. The electrodiagnostic test conducted on the ulnar sensory and the tibial motor parts indicates that the sensory and motor evoked wave amplitudes increase after dialysis (p value < 5%) but there is little change in the nerve conduction velocity. Wave amplitude is related to number of actively participating axons in neural message transference, so a decrease in amplitude means a decrease in the number of active axons involved in the neural message transfer. The present findings indicate that in the process of dialysis some of the previously inactive axons become activated. The reason for the inactivation of the axons may be due to the accumulation of toxic substances in the body as a result of renal failure. After hemodialysis, most the toxic substances are removed from the body and this leads to an increase in the number of active axons. PMID- 11284055 TI - The correlation between cerebrospinal fluid findings and evoked potentials during an acute MS attack. AB - Clinically definite Multiple Sclerosis (MS) cases have an index greater than 0.7. Evoked potentials using geometrical patterns and click stimuli respectively and electrical stimulation of mixed nerve trunks evokes low level electrical impulses along a nerve. Prolonged latencies and changes in amplitude in these evoked potentials are thought to reflect sensory loss. This study is performed to determine whether there is a correlation between cerebrospinal fluid (CSF) findings and evoked potentials in MS during an acute MS attack. 10 male and 10 female patients' (20-55) mean (37.75) were included in to the study. They were diagnosed as definite MS with their clinical and MRI findings. During the acute attack 10 (50%) patients out of 20 had high Ig G index and one (10%) had oligoclonal band positivity. Of these 10 patients 8 (4 M, 4 F) had pathology in their evoked potentials while two had normal evoked potential findings. 8 of (80%) 10 patients whom had normal Ig G index during the acute attack had evoked potential pathology while (20%) had normal Ig G index and evoked potentials during the acute attack. Of the 8 patients which had high Ig G index and pathological evoked potential findings during acute attack were evaluated. VEP, BAEP latencies were found to be prolonged as Ig G index increased. (p < 0.05) However the same correlation can not be found with SEP parameters. PMID- 11284056 TI - Descending and ascending spinal cord evoked potentials from oesophago-vertebral electrical stimulation in normal awake man. AB - Electrical stimulation of the spinal cord at C7D1 evoked triphasic descending spinal cord evoked potentials (DSCEP) from an oesophago-vertebral recording at D8D8 or D1OD1O. Ascending SCEPs (ASCEP) larger and similar in shape were also observed when the orientation of the stimulating and recording dipoles was reversed. Both SCEPs are in part generated by descending and ascending synchronous excitation of neuronal volume-conducted spinal cord dipoles. PMID- 11284057 TI - Effects of heat and cold application on turns and amplitude in surface EMG. AB - The purpose of this study was to investigate the effects of cooling and heating of muscles on parameters of surface EMG (SEMG) under various well defined grades of isometric muscle contraction. In 32 healthy volunteers, aged 20-30 years, turns and amplitude (RMS) analysis was done in SEMG from the hand extensors. Muscle strength was defined by a new developed hand dynamometer. Values for RMS and turns in case of maximal voluntary contraction (MVC) over periods of 20 sec were determined for each volunteer and subsequently RMS and turns were measured during isometric contractions of 10%, 30%, 50% and 80% of MVC, interrupted each by rest periods. This procedure was repeated after cooling and warming the forearm with thermo packs. For control purposes the same investigations with three measurement periods of SEMG were done following the same time schedule without thermic stimuli. With increasing isometric contractions turns increased after heat application and decreased after ice application, compared to the values achieved without thermic stimuli. The differences were significant from a level of 30% of MVC (p < 0.05). No corresponding changes were seen for RMS. In the control experiment values of turns and RMS did not differ in the three measurement periods. It is concluded, that the amplitude in SEMG tracings seems to be a reliable parameter of muscle force, whereas turns are sensitive to temperature. They might be useful markers for therapeutic approaches targeted at the muscle. PMID- 11284058 TI - Anodal excitation of intact peripheral nerves in humans. AB - The median nerves of six normal subjects were electrically stimulated at different locations on the wrist in a bipolar fashion. The evoked compound sensory nerve action potentials (CSNAPs) were recorded at the index finger. Electrical stimuli consisted of constant-current monophasic rectangular pulses. Initially, the nerve was stimulated with a cathodal current and a just supramaximal CSNAP was obtained. Then, at the same stimulus location, the nerve was excited with an anodal current. A just supramaximal response in the anodal CSNAP was obtained, or until a maximum current of 100 mA was delivered. CSNAPs obtained from anodal excitation required several times more electrical stimulus intensity when compared to those derived from cathodal stimulation. Morphologically, they were similar, except the onset latency of the anodal-CSNAPs was approximately 0.2 to 0.3 msec early compared to that derived from cathodal stimulation. This implies that during anodal excitation, the site of nerve depolarization was not located underneath the anode, but was situated at some distance from it. Also, it appears that a minimum nerve length (10 mm or more) should be exposed to the anodal current in order to induce depolarization. A very localized application of the anodal stimulus often failed to elicit nerve depolarization. PMID- 11284059 TI - Changes in serial correlation coefficients and fractional parameters during functional recovery in stroke patients. AB - This study attempted to determine changes in motor unit discharge patterns, characterized by serial correlation coefficients (SCC) and fractional parameters during a functional recovery in stroke patients. Using the surface electrode technique, 171 single motor units were recorded from bilateral upper extremities of 23 stroke patients and 7 normal volunteers. SCCs and fractional parameters were calculated and correlated with Rasch-converted Functional Independence Measurement (FIM) scores by linear regression analysis. No statistically significant correlation was found between FIM scores and SCCs and between FIM scores and fractional parameters recorded during first four weeks post stroke. Only FIM for total motor scores and FIM for locomotion subscore significantly correlated with SCC recorded after 4 weeks post stroke (p < 0.05). Predictive power (R2) of SCC for FIM scores did not change much over time. Fractional parameters, recorded after 4 weeks post stroke, had significant correlation with FIM for total motor scores and all FIM subscores except FIM for sphincter control subscores. Predictive values of fractional parameters for FIM scores was better after 6 months post stroke. There was a better correlation between fractional parameters with FIM scores and a better predictive value of fractional parameters for FIM scores over functional recovery period. This change in the correlation during the course of stroke recovery may indicate that fractional parameters could reflect a more stabilized motor control stage rather than a less stabilized motor functional stage. PMID- 11284060 TI - Spontaneous blinks of Parkinson's disease patients evaluated by EMG and EOG. AB - In a study of spontaneous blinks, both electromyographic (EMG) activities from m. orbicularis oculi which is responsible for initiating closure of the eyelid and electro-oculogram (EOG) of vertical direction to the movement of the eyelid were measured in ten patients with Parkinson's disease and in thirty normal subjects. The aim of this study was to evaluate the generative mechanism of the spontaneous blinks by comparison of both the EMG and the EOG waveforms in the patients with Parkinson's disease and those in the normal subjects. The mean duration and the amplitude of both the EMG and the EOG were evaluated by the averaging of ten waveforms for the spontaneous blinks. The time lag between the onset of the generation of the EMG and the onset of the EOG signal was analyzed. The mean duration of the EMG and the mean amplitude of both the EMG and the EOG in the patients with Parkinson's disease were shorter and smaller than those in the normal subjects by the significant level of 1%, respectively. There was no difference of the time lag between the subject groups. These results suggest that the function of m. orbicularis oculi for the spontaneous blinks is reduced in patients with Parkinson's disease, because the motoneurones of the facial nucleus innervating the m. orbicularis oculi becomes hypoactive due to abnormal output of basal ganglia. PMID- 11284061 TI - Comparison of effects of spinal manipulation and massage on motoneuron excitability. AB - The purpose of this study was to compare the magnitude and duration of motoneuron inhibition occurring as a sequel to spinal manipulation or paraspinal and limb massage. The physiologic mechanisms involved in spinal manipulative therapy and massage therapy are largely unknown. One possible hypothesis is based upon the theory that these two distinct and different modalities may attenuate the activity of alpha motoneurons. Both modalities have been reported to produce short-term inhibition of motoneurons. Asymptomatic volunteers were randomly assigned to either a spinal manipulation, massage, or control group. Baseline tibial nerve H-reflex amplitudes were obtained prior to the application of either lumboscaral spinal manipulation or paralumbar and limb massage. Post interventional H-reflex recordings were recorded immediately following the application of either modality. Spinal manipulation significantly (p < 0.05) attenuated alpha motoneuronal activity immediately post-therapy, as measured by the amplitude of the tibial nerve H-reflex. Massage subjects exhibited no significant reduction in motoneuronal activity immediately following administration. Spinal manipulation produced a transient attenuation of alpha motoneuronal excitability. Paraspinal and limb massage did not inhibit the motoneuron pool as measured immediately post-therapy. These findings support the supposition that spinal manipulation procedures lead to short-term inhibitory effects on motoneuron excitability to a greater magnitude than massage. PMID- 11284062 TI - [Current pathogenetic aspects of Sezary syndrome and mycosis fungoides]. AB - Cutaneous T-cell lymphomas are a heterogeneous group of lymphoproliferative skin disorders whose pathogenesis is poorly understood. Cytokines and chemokines are important factors which can modify the cutaneous microenvironment allowing the accumulation of lymphocytes. Most of these neoplasms seem to be T helper-2 cells. While interferon gamma is the natural inhibitor of clonal T-cell proliferations, interferon resistance has been recently found in these cells. This interferon resistance may be an Achilles heel that can be targeted by molecular therapeutic interventions. PMID- 11284063 TI - [Alternative activation of antigen-presenting cells: concepts and clinical relevance]. AB - Lymphocytes do not just act as immunological effector cells, but also play an important role in the regulation of the immune response. They are able to induce or suppress inflammatory reactions and this balancing function is reflected in the well-known Th1/Th2 concept. Lymphocytes depend on antigen presenting cells (APC) for induction of differentiation and specific activation mediated by antigen capture, processing and presentation. Thus, APC represent a link between innate and acquired immunity. In parallel to the Th1/Th2 dichotomy, APC may be subdivided into (a) pro-inflammatory, classically activated APC such as mature dendritic cells and IFN-gamma-activated effector macrophages, and (b) into anti inflammatory, alternatively activated APC such as IL-10-activated immature dendritic cells and IL-4-induced suppressor macrophages. Alternatively activated APC may mediate induction and maintenance of tolerance towards allergens and environmental substances, control the course of inflammatory reactions, and participate in healing processes by enhancing angiogenesis. Malignant tumors and certain infectious agents may misuse alternatively activated APC for their purposes, thereby requiring counter-action by Th1 lymphocytes and classically activated APC. The concept of alternative activation thus confirms the important role of APC in maintaining the balance between induction and suppression of both inflammation and immunity and it opens new perspectives for the development of specific immunotherapeutic approaches. PMID- 11284064 TI - [Guidelines for cyclosporin therapy in dermatology. Issued by the Consensus Conference on Cyclosporins]. PMID- 11284065 TI - [Anti-CD20 antibodies in primary cutaneous B-cell lymphoma. Initial results in dermatologic patients]. AB - BACKGROUND AND OBJECTIVE: Primary cutaneous B cell lymphomas (pCBCL) are rare extra-cutaneous non-Hodgkin lymphomas which occur on the trunk as follicle center cell lymphoma or on the leg as large B cell lymphoma. The currently accepted therapy of pCBCL (excision and/or radiotherapy, systemic interleukin 2 and interferon alpha 2a, local injection of cisplatin or multiagent chemotherapy, i.e. CHOP) is insufficient for treatment of multifocal pCBCL and secondary extracutaneous involvement. For this reason, the new synthetic chimeric, monoclonal anti-CD20 antibody Rituximab is an alternative treatment for patients with pCBCL. PATIENTS/METHODS: Four patients with pCBCL localized to the trunk or head were treated with Rituximab (375 mg/kg weekly for 4-8 weeks, then maintenance therapy every 4 weeks for 6 months). RESULTS: All 4 patients showed a response (2/4 partial; 2/4 complete). Side effects were minimal. CONCLUSIONS: Rituximab is an alternative immunotherapeutic drug for the treatment of pCBCL. Our initial experience with this new modality are presented and discussed. PMID- 11284066 TI - [Mortality of invasive malignant cutaneous melanoma. A review with special consideration of gender distribution]. AB - BACKGROUND AND OBJECTIVE: The final goals of malignant melanoma prevention are lowering incidence and mortality. We assessed the parameter "survival" for both men and women as the beginning point for future gender-directed prevention campaigns. We compared the periods 1972-1980, 1981-1988, 1989-1996, and determined the influence of age and of Breslow' tumor thickness on survival. PATIENTS/METHODS: We had sufficient follow-up on 10.433 patients. We calculated survival curves according to Kaplan-Meier and defined differences by the logrank test. RESULTS: At all periods of time, survival of women was higher compared with men, but with no impressive changes over time. This was especially true for younger men. The most important prognostic factor was the Breslow tumor thickness. Within all periods of time, its median was higher in men. A trend downwards for both genders could be observed with higher influence on survival in men. CONCLUSIONS: Our findings justify melanoma prevention campaigns addressed to men. Evaluation of such campaigns has to take into account an already existing upwards trend for male survival, which exceeds that of female survival. PMID- 11284067 TI - [Hairdex: a tool for evaluation of disease-specific quality of life in patients with hair diseases]. AB - Because of a strong reduction of life quality in patients with hair loss, a special questionnaire was developed following the general Skindex questionnaire for dermatoses. The Hairdex was evaluated in 75 female hair patients. The aim of the study was to prove the questionnaire's statistical values, reliability, validity, general acceptance and accuracy. The clinical manifestation of hair loss was categorized as "not visible", slightly visible" and "obviously visible". The hair loss lead to great differences in the life quality of both the patients with obvious hair loss and in patients with non-visible hair loss, especially in the categories "emotions", "self-confidence" and "stigmatization". The convergent and discriminant validity of the questionnaire was satisfactory. The acceptance of the questionnaire was very good with 90%. The hairdex-questionnaire represents a reliable and patient-oriented instrument for evaluation of life quality in hair diseases. The reliability of the questionnaire concerning longitudinal therapeutic effects will need to be investigated in further studies. PMID- 11284068 TI - [Granuloma annulare disseminatum: successful therapy with fumaric acid ester]. AB - A 64-year old female patient was treated for a therapy-resistant generalized granuloma annulare with fumaric acid esters (Fumaderm initial). One week following begin of therapy with an initial dose of 30 mg per day, the papules had regressed significantly. A 6-week therapy with a final dose of 90 mg Fumaderm initial per day led to a nearly complete healing of the illness. PMID- 11284069 TI - [40 years to Flegel's disease (hyperkeratosis lenticularis perstans)]. AB - A case report of hyperkeratosis lenticularis perstans (Flegel) is presented. The disease is characterized clinically, histologically and with electron microscope. The evolution of the disease since Flegel described it 40 years ago is considered, with emphasis on etiology, pathogenesis and unanswered questions. PMID- 11284070 TI - [Plantar fibromatosis with marked cutaneous involvement]. AB - Cutaneous involvement in plantar fibromatosis is very rare. The classical finding are nodules in the plantar arch, which can be detected only with palpation. A 55 year-old man presented with a 3 year history of painful plantar nodules and an ulceration. Histopathology showed a fibroblastic proliferation with a pseudosarcomatous configuration. The immunohistochemistry was positive for vimentin, alpha-actin and desmin, a pattern which characterizes a myofibroblast. Intralesional therapy with corticosteroids did not reduce the lesions. After surgical treatment, the lesions recurred. PMID- 11284071 TI - [Follicular mycosis fungoides (FMF): a rare disease]. AB - Follicular mycosis Fungoides (FMF) was first described in 1924. Since the first description, 22 patients with this special form of mycosis fungoides have been published. Clinical features include epidermal cysts as well as follicular papules, nodules and hyperkeratoses. FMF can be confused with acneiform dermatoses. In most cases, infiltrated plaques typical for cutaneous T cell lymphoma are also present. Histology shows a monomorphic CD4+ T cell infiltrate. Treatment and prognosis are similar of those of classical mycosis fungoides. We present one patient with FMF and review the literature of all published cases. PMID- 11284072 TI - [Multiple pigmented basalioma of the scalp in a patient with Curschmann-Steinert myotonia dystrophica. Confirmation of a rare symptom constellation]. AB - A 50-year-old woman presented with myotonic dystrophy (Curschmann-Steinert disease) and multiple pigmented basal cell carcinomas of the scalp. She also had typical androgenetic alopecia seen in this disorder. In 1986 Stieler and Plewig described the first patient with myotonic dystrophy and multiple basal cell carcinomas. There may be a genetic predisposition for cutaneous tumors with follicular origin, as multiple pilomatricomas also occur frequently in such patients. PMID- 11284073 TI - [Successful therapy with tetracycline and nicotinamide in cicatricial pemphigoid]. AB - We describe a case of cicatricial pemphigoid in a 92-year-old female with extensive mucocutaneous involvement. She developed extensive hemorrhagic blistering with severely bleeding lesions, that healed with scarring. The conjunctivae showed extensive synechia. The diagnosis was based on clinical and histopathological features as well as immunofluorescence findings and immunoblot analysis. There was no clinical response to topical corticosteroids. The patient was given tetracycline and nicotinamid and showed rapid improvement of the mucocutaneous lesions within a few weeks. The clinical features, differential diagnosis and various treatment modalities of cicatricial pemphigoid are briefly reviewed, whereby the use of tetracycline and nicotinamide is discussed as an alternative effective and safe therapy for this potentially incapacitating condition. PMID- 11284074 TI - [Comments on the article by B.M. Henz and T. Zuberbier: "Urticaria--new developments and perspectives". Der Hautarzt (2000) 51:302-308]. PMID- 11284075 TI - [Differences in the groups of chronic urticaria as explanation of divergent results in the search for etiology]. PMID- 11284076 TI - [Letter concerning the article by E.C.Bornhovd, E.Schuller, T.Bieber, A.Wollenberg: "Immunosuppressive macrolides and their use in dermatology". Der Hautarzt (2000) 51:646-654]. PMID- 11284077 TI - [Cystic papules on the breast of a 35-year-old woman]. PMID- 11284078 TI - [Immunoglobulin treatment in dermatology]. PMID- 11284079 TI - [Critical dose drugs]. PMID- 11284080 TI - [Sport as medicine]. PMID- 11284081 TI - [Therapeutic strategies in hyperhidrosis]. PMID- 11284082 TI - [Cotton-tipped swabs in the ear canal. Superfluous and dangerous]. PMID- 11284083 TI - [Quality assurance of medical information offered on the Internet]. PMID- 11284084 TI - The concentration of protein-compounds in interstitial tissue of patients with chronic critical limb ischaemia and oedema. AB - BACKGROUND: Many of chronic critical limb ischaemia (CLI) patients have distal leg and foot oedema. Previous electronmicroscopic studies have shown that chronic severe ischaemia may cause hypoxic damage of the capillary endothelium, including morphological changes i.e. multiplicated/thickened basal lamina, and formation of interendothelial gaps. To assess the functional consequences of these morphologic derangements, where proteins can leak through, we investigated the composition of the interstitial fluid in oedematous ischaemic limbs. PATIENTS AND METHODS: Nine female and 3 male patients with a mean age of 79 +/- 7.9 years were included. All had unilateral CLI and peripheral pitting oedema. Leg and foot volume was measured with water displacement volumetry. Blister suction technique was used to collect subcutaneous interstitial fluid. The concentration of albumin, transferrin, immunoglobulin G and alpha 2-macroglobulin in plasma and blister fluid was measured by immunoturbidimetry. Nine patients, 8 women and 1 man with a mean age of 83 +/- 5.5 years with a proximal femur fracture served as an age matched control group. RESULTS: The mean concentration of albumin in blister fluid was significantly lower in the patients, whereas the mean concentration of alpha 2-macroglobulin in blister fluid did not differ between patients and controls. Mean ratio between concentrations in blister and serum of albumin, transferrin and immunoglobulin G in the limbs with CLI and oedema were significantly lower than respective values in the control group. However, there was no significant difference in the ratio of alpha 2-macroglobulin between these groups. CONCLUSION: A higher transcapillary concentration gradient for proteins in CLI limbs signifies an increase in the net osmotic pressure gradient across the capillary wall, which may be a potential oedema limiting factor. PMID- 11284085 TI - Kidney protection in preventing post-ischaemic renal failure during thoracoabdominal aortic aneurysm repair: does prostaglandin E1 together with cooling provide more protection than cooling alone? AB - BACKGROUND: Prostaglandin E1 (PGE1) is known to have a positive effect on kidney function after kidney ischaemia due to aortic clamping. Side effects of PGE1 are a decrease of systemic blood pressure and prevention of thrombocyte clotting, both being undesired during repair of thoracoabdominal aortic aneurysms (TAA). The aim of this study was to evaluate, whether intraoperative and intraarterial kidney perfusion with 4 degrees Ringer's lactate plus 1000 IU of heparin/l plus 20 micrograms PGE1/l is more effective in preventing postischaemic kidney dysfunction than cold perfusion without PGE1. PATIENTS AND METHODS: In the time period from I/1996 until X/1998 58 patients underwent aortic repair for TAA type II, III, or IV (Crawford's classification). Ten patients fulfilled the criteria for this study: renal artery stenosis or occlusion was excluded by angiography pre- and postoperatively. By means of szintigraphy an at least 30% participation in renal function had to be proven for every kidney. Intraoperatively both kidneys had to be excluded from circulation simultaneously. The left kidney in each patient was perfused with 4 degrees Ringer's lactate plus 1000 IU of heparin/l plus 20 micrograms PGE1/l. The right kidney was perfused with a solution of the same temperature plus heparin but without PGE1. RESULTS: There was an intermittent increase of creatinin and/or urea in each patient postoperatively. By renal szintigraphy, which was performed after a mean time of 9 (5-13) days postoperatively, a shift of renal function from one kidney to the other could be excluded. CONCLUSION: In this experimental setting no additional benefit for kidney function could be detected, when under conditions of ischaemia kidneys were perfused with 4 degrees Ringer's lactate plus 1000 IU of heparin/l plus 20 micrograms PGE1/l compared to kidneys perfused with the same solution without PGE1. PMID- 11284086 TI - Patients with persistent APC-resistance without factor V Leiden mutation. AB - BACKGROUND: Activated protein C (APC) resistance and factor V Leiden mutation are major risk factors for deep venous thrombosis. Previous work has led to the view that the coagulation phenotype and the genetic defect are associated in almost all patients. It has been reported about single APC-resistant patients without associated factor V Leiden, but significance and thrombotic risk of this constellation have not yet been established. PATIENTS AND METHODS: We tested 486 consecutive patients with deep venous thrombosis, arterial disease or other than vascular disease for APC-resistance with a factor VIII based assay. RESULTS: 149 patients (31%) showed a pathological APC-ratio. Sensitivity and specificity for detection of factor V Leiden were 100% and 40%, respectively. At 6 months follow up APC-ratio returned to normal in 55% of the patients with initial pathological APC-resistance. At 12 months follow-up 91% of the patients with persistent APC resistance showed a pathological ratio as well. CONCLUSIONS: Patients with APC resistance not due to factor V Leiden can be attributed to one subset with reversible APC-resistance--possibly due to a hypercoagulable state in an acute thrombotic situation, and to another with persistent APC-resistance. PMID- 11284087 TI - Varicosis of the accessorial lateral saphenous vein: evidence of recirculation shown by pre-operative phlebography. AB - BACKGROUND: The incidence of varicosis of the anterior side branch of the long saphenous vein clinically ranges about 8 to 10% of descending venous decompensation, originating at the level of the thigh. Its incidence in women showing significant overweight is clearly pronounced. Mostly torturous and enlarged varicosed segments of the accessorial lateral saphenous vein can be seen clinically crossing the anterior middle of the thigh. The indications for surgical radical extirpation of the entire varicosed side branch depends from pain, phlebitic complications, peripheral venous dysfunction including cosmetic aspects. PATIENTS: In a clientele of 138 patients (females: 114, males: 24) the phlebographic demonstration of the entire recirculation pathways was performed prior to surgery. RESULTS: Ascending leg phlebography was found sufficient in 7.7% of cases only. Combined with varicography however, in further 90.7% of patients a clear demonstration of the upper and lower points of venous insufficiency was possible. The extent of functional disorders was calculated by additional peripheral venous pressure measurements. CONCLUSIONS: This clientele clearly shows that long-term reflux via the varicosed side branch increases the tendency of peripheral functional decompensation along the lower leg. Adequate surgical therapy depends from a clear demonstration of the varicosed veins being involved, and this can be realized by combined phlebography and varicography. PMID- 11284088 TI - [Ultrasound study before surgery of varicose veins]. AB - Ultrasonographic investigation of the various forms of chronic venous insufficiency has substantial advantages compared to diagnosis with the competing phlebogram, particularly preoperatively. Important details such as side branches in the region of the groin, course variations of the small saphenous vein and insufficiency of the perforators in the lower leg are sometimes missed in the antegrade phlebogram. However, it is absolutely necessary to take these into consideration in order to attain a substained good result of surgery. With adequate qualification of the investigator and using all ultrasound techniques, diagnosis by sonography is better than by means of antegrade phlebogram. There are clear specifications for the documentation. With regard to costs and time required, ultrasonographic investigation of chronic venous insufficiency is superior to the antegrade phlebogram and can be repeated at any time. PMID- 11284089 TI - Brain involvement in extracranial internal carotid artery aneurysms. AB - BACKGROUND: No prospective study of extracranial internal carotid artery aneurysms (EICAA) has been reported to date. The aim of this study was to evaluate central nervous system complications associated with surgical intervention for EICAA. PATIENTS AND METHODS: A total of seven patients, representing all cases observed at our institution from December 1997 to December 1998, were entered in this prospective study. Three patients had bilateral involvement. The aneurysms were both atherosclerotic and dysplastic. All patients were males, with mean age of 70 years (range 65 to 74). Internal or common carotid artery to EICAAs diameter ratios were calculated on the angiograms. The transverse diameter as well as the craniocaudal extension of the lesions were accurately measured intraoperatively. Follow-up evaluations were performed at three, six and twelve months postoperatively, and consisted of a clinical evaluation by both a neurologist and a vascular surgeon who were not part of the primary surgical team. RESULTS: Six patients presented with neurological symptoms ranging from non-hemispheric TIAs to hemispheric stroke. One patient was asymptomatic. The severity of symptoms was correlated with the size of the aneurysm. Preoperative symptoms were more severe in EICAAs of > or = 3 cm in transverse diameter. One case had a postoperative stroke, no perioperative deaths occurred. All the internal carotid arteries operated on were patent during follow up evaluations. No new neurologic event was observed during follow-up. CONCLUSIONS: The severity of central neurologic symptoms seems to depend on the size of the aneurysmatic lesion. Prompt surgical management of small EICAAs may reduce the occurrence of severe CNS complications, both preoperatively and postoperatively, due to the lower risk of embolization associated with small aneurysms compared to larger lesions. PMID- 11284091 TI - Is there a place for duplex screening of brachial artery in haemodialysis patients with vascular access? AB - BACKGROUND: Vascular access (VA) stenosis with subsequent thrombosis remains one of the major causes of morbidity and hospitalization in haemodialysis patients. The present cross-sectional study was planned in order to analyze the usefulness of brachial artery duplex ultrasound for detection and prediction of vascular access stenoses. METHODS: Color duplex ultrasound (Apogee Cx200, sectorial probe 7.5 MHz) was used to obtain the anatomical pattern of the VA and flow velocity waveforms of the brachial artery in 77 non-selected VA (47 Ciminio-Brescia fistulae and 30 PTFE grafts). In each VA, the resistance index (RI), the mean blood flow rate (Q) and the blood flow ratio index (QI) (QI = VA flow rate/contralateral flow rate) were calculated at the level of the brachial artery. The sensitivity and specificity of these brachial Doppler parameters were calculated for the detection of VA stenosis. In normal VA, positive (PPV) and negative predictive (NPV) values were calculated for the development of clinical stenotic complications 3 months post ultrasound examination. RESULTS: Thirteen of the 77 VA (17%) were identified as stenosed by duplex ultrasound and confirmed by fistulography and/or during surgical exploration. The best screening tests for VA stenosis detection were a QI threshold < 4.0 with a sensitivity and specificity of 69 and 69% and an RI > 0.55 with a sensitivity and specificity of 62 and 66%, respectively. In the VA considered as normal by ultrasound, the prediction of subsequent stenosis within three months post-ultrasound examination gave a PPV of only 18% and 19% for RI and QI, respectively. NPV for RI and QI were 90% and 88%. CONCLUSIONS: While Doppler ultrasound is a useful non-invasive test for the detection of prevalent VA stenosis, our results do not confirm that abnormal brachial Doppler flow parameters can predict short term development of VA stenosis. PMID- 11284090 TI - Cardiogenic embolism as the main cause of ischemic stroke in a city hospital: an interdisciplinary study. AB - BACKGROUND: It is essential to understand the pathogenesis of ischemic stroke to ensure rational acute therapy and secondary prevention. We wanted to know the distribution of pathogenesis in patients of a city hospital and the differences in risk factors, neurologic deficits, disability, and delay in clinical admittance. PATIENTS AND METHODS: During a period of one year, 222 patients (mean age 76.6 years; 59% women) with complete acute ischemic stroke were admitted and underwent complete clinical and diagnostic procedures: CCT/MRI; Doppler- and color-coded duplex and transcranial sonography; echocardiography; use of the NINCDS stroke scale and the Oxford disability scale; study of risk factors, and exploration of delay in admittance. RESULTS: The following percentages of etiologies were evident: 31% cardiogenic embolism (60% with atrial fibrillation), 13% microangiopathy, 9% macroangiopathy, 11% cerebellar or brain stem infarction, 18% more than one cause and 18% no cause found. The patients with cardiogenic embolism showed significantly the highest scores on the stroke scale and the disability scale and had the shortest delay in admittance (57% were admitted within 3 hours). CONCLUSIONS: In a city hospital, cardiogenic embolism is the main cause of ischemic stroke. These patients suffer significantly the most severe neurologic deficits, dependence, and requirement of daily nursing care. These patients have the shortest delay in clinical admittance and the best chance of benefitting from acute therapy and early secondary prevention. PMID- 11284092 TI - Acute occlusion of the left iliac artery after long-distance-running. AB - We report a case of spontaneous iliac occlusion in a 44-year-old male patient after long-distance running. Atherogenic risk factors like hypertension, diabetes, hypercholesterolemia and smoking were missing. Spontaneous iliac occlusion is extremely rare and only a few cases have been documented. Angiography showed occlusion of the left iliac artery with collateral flow via the obturator artery to the common femoral artery. Thrombectomy was performed but reocclusion occurred. An iliacofemoral bypass, arterial lysis and bypass thrombectomy was necessary within a few months. At the last follow-up visit two years afterwards the patient was symptom-free. This case indicates that exercise dependent blood flow disturbances in long-distance-runners could produce changes of the intima. PMID- 11284093 TI - Vascular tumors simulating occlusive disease. AB - Two cases of vascular tumors of large vessels with intraluminal growth simulating venous thrombosis and arterial occlusive disease are reported. One was a borderline malignant epithelioid hemangioendothelioma of the femoral vein and the other a malignant epithelioid angiosarcoma of the carotid artery. Immunohistochemical studies permitted to classify the tumors. Treatment consisted in surgical resection. No recurrence and no metastasis are noted at 24 months. Uncertainty regarding biological behaviour of vascular tumors and treatment persists. PMID- 11284094 TI - Hitherto undescribed venous vacuolar myopathy without mucoid degeneration in the varicose saphenous vein of a child. AB - Varicose vein abnormalities involving the lesser saphenous vein of a 7-year-old boy are presented. The histopathology of the vein wall revealed a vacuolar degeneration without mucoid deposits of the muscle cells, which has not been described previously in the literature in congenital varices. PMID- 11284095 TI - [Recurrent coumarin necrosis in type II protein S deficiency]. AB - Coumarin necrosis is a rare but clinical very important complication of therapy with coumarin derivatives. We report a patient with congenital protein S deficiency type II, who developed coumarin necrosis during stabilization of phenprocoumon. Diagnostic problems and therapeutic alternatives are discussed considering the recent literature. PMID- 11284096 TI - Guidelines for testing drugs for chronic venous insufficiency. PMID- 11284097 TI - In vivo evaluation of an intravascular receiver coil for MRI at 1.0 Tesla. AB - BACKGROUND: Evaluation of the applicability of intravascular radiofrequency receiver coils by means of a pig in-vivo-experiment for the detection of vessel wall structures at 1.0 Tesla. MATERIALS AND METHODS: The intravascular receiver coil was constructed according to a well evaluated single-loop-design, which was mounted onto a balloon catheter for angioplasty. Under fluoroscopy control the balloon catheter was placed in the common carotid artery of a porcine. Images were obtained in a 1.0 Tesla clinical scanner using a fast-spin-echo-, a gradient echo- and a high resolution spin-echo-sequence. Histological examinations were obtained to detect any vessel wall damage associated with the use of the receiver coil. RESULTS: High quality images with a resolution up to 0.16*0.12 mm2 could be acquired in aquisition times of about 5 minutes. Subtle intra- and extra-vascular structures such as the balloon, irregularities of the vessel wall or bordering structures could be visualized. The histological examination showed no vessel damage due to the application of the intravascular receiver coil. CONCLUSIONS: The application of intravascular receiver coils for the visualization of the vessel wall is feasible at 1.0 Tesla without histologically detectable trauma to the vessel wall. PMID- 11284098 TI - [Xenograft: perpetual future of transplantation...?]. PMID- 11284099 TI - [Laryngeal immobility after thyroidectomy]. AB - One of the most common complications of surgery of the thyroid gland is vocal folds immobility. New advances in its management have been achieved over the last few years. Laryngeal electromyography, stroboscopy, and computerized analysis of the voice help guide diagnosis, allowing differentiation between recurrent nerve paralysis and glottis traumatism due to intubation, and further follow-up of recovery with relevant therapeutic decisions. In case of unilateral vocal fold paralysis, intrafold silicone or injection of autologous fat is more and more routinely used to obtain vocal rehabilitation. In case of bilateral vocal fold paralysis, to avoid tracheotomy, partial posterior cordectomy using laser surgery restores sufficient laryngeal airflow, with minimal vocal sequelae. Early management of these complications by teams of specialists should allow appropriate and less invasive surgery. PMID- 11284100 TI - [Liver transplantation in the presence of a non-functional portal vein: an original technique]. AB - STUDY AIM: The aim of this retrospective study was to report an original technique for heterotopic liver transplantation with the graft in the left hypochondrium, and to discuss the indications and limitations of this technique. PATIENTS AND METHOD: Over the past ten years, four patients were treated by this technique; this constitutes 2% of all liver transplantations carried out during this period. RESULTS: No immediate per- or postoperative mortality related to the surgical procedure was noted. Moreover, no severe hemodynamic complications occurred during the per- or postoperative period. In three out of four cases, hepatic function was fully restored within 48 hours. Long-term survival (50 and 97 months) was observed in two patients. CONCLUSION: Heterotopic liver transplantation in the left hypochondrium is an alternative to orthotopic liver transplantation; it is a technique that is easy, non-aggressive, and with good long-term results. It is indicated in cases where the main portal vein is non functional (following total thrombosis or porto-caval shunt), and orthotopic liver transplantation is therefore not possible. PMID- 11284101 TI - [Radiofrequency ablation of malignant hepatic tumors. Preliminary experience apropos of 25 cases]. AB - STUDY AIM: Radiofrequency (RF) ablation of malignant hepatic tumors is an interesting and recent technique which offers new treatment possibilities. In this study, the preliminary findings have been reported on 25 patients with hepatic tumors who received RF treatment between January 1998 and February 2000. PATIENTS AND METHODS: Twenty-five patients (11 cases of hepatocellular carcinoma, HCC; and 14 cases of liver metastases, LM) underwent RF treatment. Thirty tumors (range: 10 to 54 mm in diameter) out of a total of 63 were destroyed by RF: 13 HCC (average diameter: 32 mm) and 17 LM (average diameter: 26 mm). Treatment consisted of six percutaneous and 19 surgical RF procedures. In the surgical group, there were 11 cases of hepatectomy: right hepatectomy in five patients with segment IV enlargement in one case, and sub-segmentectomy in six other patients. In all cases, hepatic tomodensitometry was performed at one month post treatment and then every three months. RESULTS: Postoperative portal thrombosis occurred in two patients, one of whom died. Other postoperative complications were observed in five patients. During the mean follow-up period of 14 months (range: 2 to 28 months), two patients died (carcinosis, ascitic decompensation), two and four months respectively after RF treatment. In situ recurrence occurred in four HCC and two LM patients. Three HCC and four LM patients developed new hepatic or extra-hepatic lesions. CONCLUSION: RF is a particularly interesting technique for the treatment of bilobar or unresectable metastases. It appears to be equally as efficient as other local treatments for small-sized HCC. However, technical improvements remain necessary to increase the destructive field covered by RF. A more important follow-up is needed so that the long-term efficacy and specific role of this new therapy can be accurately assessed. PMID- 11284102 TI - [Intensive care after digestive surgery: the outcome in elderly patients]. AB - STUDY AIM: In surgical intensive care, the results must be analyzed both in terms of mortality and quality of life; this is particularly important in elderly patients for whom recovery remains uncertain. The aim of this prospective study was to assess the early and late prognosis in elderly patients (aged over 75 years) admitted to a digestive surgical intensive care unit (DSICU) for mortality, quality of life, patient autonomy, and also the predictive factors involved. PATIENTS AND METHODS: Over a one-year period, 182 patients were admitted to a tertiary referral DSICU; 30 of these subjects were over 75 years old, and formed the basis of this study. The following data were analyzed: hospital mortality rate; mortality rate at six months, and quality of life at six months (Kamofsky scale). These factors were correlated with the severity of the patient's state at admission and also with the causal disease, circumstances connected with admission, and duration of stay in the DSICU. RESULTS: The hospital mortality rate of patients was 23% (7/30 patients), and the overall mortality rate at six months was 40% (12/30 patients). Of the 12 patients who stayed in the DSICU for more than ten days with a simplified acute physiology score (APS) = 10, not one was alive at six months post-DSICU admission. The 18 remaining patients were still alive at six months, and 72% of them (13/18 patients) had regained their previous post-operative autonomy. CONCLUSION: These results provide reference data for this patient category. The results concerning long-term survival and the good functional outcome are encouraging. If the prognostic criteria defined in this investigation are confirmed by further studies, they may help in making the sometimes difficult decisions regarding elderly patients hospitalized in a DSICU. PMID- 11284103 TI - [Percutaneous drainage of perisigmoid abscesses of diverticular origin]. AB - STUDY AIM: The aim of this multicentric retrospective study was to report the results on the percutaneous drainage of perisigmoid abscesses during acute sigmoid diverticulitis in 12 patients. PATIENTS AND METHOD: Between January 1993 and March 2000. 12 patients with a perisigmoid diverticular abscess were treated by antibiotic therapy and percutaneous drainage of the abscess. The patient population consisted of eight males and four females (mean age: 50.2 years). The diagnosis was established in two out of seven cases by enema, in four cases out of seven by abdominal ultrasonography, and in eight cases out of 11 by CT scan. Percutaneous drainage was carried out in all cases, and was guided by ultrasonography (n = 3) and CT scan (n = 9). The mean duration of drainage was 6.5 days. RESULTS: No drainage-associated complications were observed. Drainage combined with antibiotic treatment provided satisfactory results in ten out of 12 cases. Two cases of failure of the method occurred, and the patients involved were operated on day 4 and week 5 by colectomy with protective lateral ileostomy. There was an early recurrence of the abscess in three patients, who were treated by the Hartmann procedure in one case, and by one-stage colectomy in two cases. Five patients underwent a secondary one-stage colectomy. Two patients in whom no residual abscess was detected were not operated on at the time of the study. CONCLUSION: Percutaneous drainage of perisigmoid diverticular abscesses combined with antibiotic therapy provided efficient treatment in ten out of 12 cases. Secondary one-stage colectomy was performed in seven out of the eight patients requiring further surgery. PMID- 11284104 TI - [Retrorectal tumors: an assessment of the abdominal approach]. AB - Retrorectal tumors are frequently resected by a posterior trans- or parasacral approach, while the anterior abdominal approach is generally reserved for small tumors situated above the sacral promontory. STUDY AIM: The aim of this retrospective study was to assess the use of the abdominal approach for the treatment of large tumor masses situated in the presacral space, and to evaluate the results in terms of resectability, morbidity, and risk of recurrence. PATIENTS AND METHODS: Between 1986 and 1998, six female patients (age range: 25 to 79 years) with a retrorectal tumor (mean diameter: 7.5 cm) were operated on by abdominal approach. Clinical findings, morphological and histological data, the surgical resection procedure, and post-operative morbidity were studied. RESULTS: Pathological findings showed that all the tumors were benign: neurofibroma (n = 2), dermoid cyst (n = 1), rectal duplication (n = 1), myelolipoma (n = 1), and epithelioid hemangioma (n = 1). Complete tumor resection was obtained macroscopically and microscopically in all cases. The postoperative course was uneventful, with no tumor recurrence detected at a mean follow-up of 31 months. CONCLUSION: The anterior abdominal approach allows the complete resection of a retrorectal large tumor mass, and provides an interesting alternative to the posterior approach, with low morbidity and an absence of functional impairment. PMID- 11284105 TI - [Immunodetection of thyroid peroxidase in the diagnosis of follicular variants of thyroid papillary cancer]. AB - STUDY AIM: The aim of this retrospective study was to assess the role of thyroid peroxidase immunodetection in the cytological diagnosis of follicular variants of thyroid papillary cancer (FVTPC) which are difficult to identify by standard cytology. PATIENTS AND METHODS: Between 1991 and 1998, 3,505 thyroid fine needle aspiration biopsies were performed by thyroid peroxidase immunocytochemistry and 1,576 patients were operated on. Out of a total of 227 thyroid papillary cancers (TPC), 42 (18.5%) were diagnosed as FVTPC. The results of standard cytology and thyroid peroxidase immunodetection were compared with the histological findings. RESULTS: The rate of false negatives for TPC in standard cytology was 11% (25/227 cases), with 40% of these false negatives being FVTPC; ten out of 42 (23.8%) cases of FVTPC were not identified by standard cytology. However, cytology with thyroid peroxidase immunodetection diagnosed 224 out of the 227 TPC (99%), and all the FVTPC were correctly identified (100%). CONCLUSION: FVTCP are the most frequent source of false negatives in standard cytology. Thyroid peroxidase immunodetection allows most of these errors to be avoided, and correctly identifies 99% of TPC including FVTPC. PMID- 11284106 TI - [Clear cell sarcoma (malignant melanoma of soft tissues) of the calf ]. AB - Clear cell sarcoma (malignant melanoma of soft parts) is a rare malignancy that is found in the young adult, and is generally located in the extremities of the limbs. In this study, a new case has been reported in a 24-year old male with no previous history of cutaneous melanoma. The tumor consisted of fusiform or round cells with clear or granulocytic cytoplasm and vesicular nuclei. The patient was treated by surgical resection of the tumor and postoperative radiotherapy. Eight years later, metastatic nodes were detected in the inguinal region. The histogenesis of this tumor has not yet been determined, and it poses a diagnostic problem for pathologists as it can be mistaken for a malignant metastatic cutaneous melanoma. PMID- 11284107 TI - [Retrocaval ureter in a 10-year-old boy]. AB - Retrocaval ureter is an uncommon abnormality of the inferior vena cava, which is rarely detected in the child due to its non-specific symptomatology. Despite a varying state of severity, chronic ureteral obstruction generally leads to the deterioration of renal function during adulthood. Treatment depends on the symptomatology in question, and surgical management should be as conservative as possible. PMID- 11284108 TI - [Video-guided retrosternoscopy: a new approach in retrosternal esophagoplasty]. AB - In this study, the authors described a new videoscopically guided approach for tunneling in esophagoplasty and insertion of an esophageal prosthesis. PMID- 11284109 TI - [Partial laparoscopic resection of splenic epidermoid cysts]. PMID- 11284110 TI - [Pseudotumorous perforation of the cecum by a duck bone]. PMID- 11284111 TI - [Mesenteric desmoplastic tumor with multiple differentiation]. PMID- 11284112 TI - [Typhoid perforation of the small intestine at the Niamey Hospital, Niger]. PMID- 11284113 TI - [Late cutaneous-vaginal fistula after a Bologna bladder neck suspension procedure]. PMID- 11284114 TI - [The problems of Spanish legal regulation on assisted human reproduction: the decision of the Constitutional Court and the first report of the National Commission of Assisted Reproduction (Part II)]. PMID- 11284115 TI - Genetic testing in the workplace. Medical, ethical and legal issues. PMID- 11284116 TI - [Nullification appeal presented by the Dutch Kingdom against the Directive 98/44 on the legal protection of biotechnological inventions]. AB - The author studies the Appeal of revocation by the Kingdom of Holland against the Directive 98/44 concerning the juridical protection of biotechnological inventions. The main reasons of the Appeal are: the election of a mistaken juridical base; the infraction of the principle of subordination; the violation of the principle of juridical security; the non-fulfillment of the obligations of the International Law; the violation of the fundamental rights; and the infraction of the principle of collegiality. PMID- 11284117 TI - A change of attitudes. PMID- 11284118 TI - The moral status of animals in the discussions on xenotransplantation (Part I). PMID- 11284119 TI - A biography and bibliography: the recent trends in bioethics and medical genetics in Japan (Part II). AB - At the present, we have much misunderstandings and prejudices, connected with psychological conflicts for the direct application of cloning and of prenatal diagnosis and selective abortion. So that we should have much debates among general public on further direction of these researchs, in order to study on safe guard and public understandings. Summarinzingly, in order to educate general public in their understanding of ELSI problems, we should remove the boundaries between individuals, in professional and non-professional fields, and in developed and developing countries, and give more time for medical curriculum for medical genetics and bioethics, in order to educate general public, as well as scientific responsibility in our society. Finally, we hope these recommendations and reports will help the International Bioethics Seminars in Fuki on 2000 for the follow up study on Universial Declaration of Human Genome and Rights, with new technologies on pharmacogenomics DNA polymorphisms as well as our ethical debates on international research ethics, proposed at the International Bioethics Summit. PMID- 11284120 TI - Preimplantation genetic diagnosis--the bridge between human genetics and reproductive medicine. PMID- 11284122 TI - Bioethics Declaration of Gijon 2000. PMID- 11284121 TI - [Seminar on the "Biomedicine Convention of Europe: its implementation in the Spanish legal code"]. PMID- 11284123 TI - [Dignity of human beings as regulatory principle in bioethics]. AB - New discoveries and advances in biotechnology are producing new social realities which must be appraised properly from the ethical point of view. Vitally important in this task is the principle of human dignity, which is examined here by the author. Human dignity is crucial in seeking to resolve the conflicts that might arise as a result of the new possibilities opened up by modern biotechnology, such as embryo research, predictive diagnosis, gene therapy, human cloning or the issue of xenotransplants. PMID- 11284124 TI - [The European directive 98/44/CE on legal protection of biotechnologic inventions]. AB - After a lengthy process, a full decade after work first commenced, at last the European Directive on legal protection of biotechnology inventions has been adopted (Directive 98/44/EC, by the European Parliament and Council, 6 July 1998. In this article the author provides an interesting study of the Directive, and discusses how the text arrived at its definitive version by comparing it with previous drafts and proposals put forward since the original one of 21 October 1988. PMID- 11284125 TI - [The Human Genome Project and the right to intellectual property]. AB - The Human Genome Project was designed to achieve two objectives. The scientific goal was the mapping and sequencing of the human genome and the social objective was to benefit the health and well-being of humanity. Although the first objective is nearing successful conclusion, the same cannot be said for the second, mainly because the benefits will take some time to be applicable and effective, but also due to the very nature of the project. The HGP also had a clear economic dimension, which has had a major bearing on its social side. Operating in the midst of these three dimensions is the right to intellectual property (although not just this right), which has facilitated the granting of patents on human genes. Put another way, the carrying out of the HGP has required the privatisation of knowledge of the human genome, and this can be considered an attack on the genetic heritage of mankind. PMID- 11284126 TI - Changes in cerebral blood flow induced by passive and active elbow and hand movements. AB - Transcranial Doppler ultrasonography (TCD) has been widely used to obtain information about changes in cerebral perfusion during motor activity after stroke. This type of application is greatly limited when severe motor deficits are present that impede the performance of an active motor task. In this study, we explored the effect of performing passive arm movements on cerebral perfusion. Twenty healthy subjects were investigated. A bilateral TCD monitoring of the middle cerebral artery (MCA) flow velocity was performed during the following experimental conditions: 1-min of active and passive flexion-extension elbow movement and 1-min of active and passive dorsal extension hand movement. Each task was performed with both left and right arms. The percentage increase in flow velocity from rest to task performance was calculated. Each task produced a significantly greater increase in mean flow velocity in the contralateral MCA with respect to the ipsilateral. When comparing the effect of passive and active tasks, no significant difference in mean flow velocity changes recorded in the ipsilateral and the contralateral MCA was detected regarding either elbow or hand movements. These findings demonstrate the possibility of obtaining information about changes in hemispheric cerebral perfusion during passive movements involving elbow and hand. This type of application deserves further attention in the study of cerebral functional changes following cerebral lesions. PMID- 11284127 TI - Non-convulsive status epilepticus in two patients receiving tiagabine add-on treatment. AB - We report two patients with epileptic syndromes who developed non-convulsive status epilepticus under adjunctive antiepileptic therapy with tiagabine. The paradoxical effect may be the result of a difference in effects between GABAA and GABAB receptors, or between GABA receptors in different regions of the brain. PMID- 11284128 TI - Clinical and genetic analysis of a four-generation family with a distinct autosomal dominant cerebellar ataxia. AB - The autosomal dominant cerebellar ataxias (ADCAs) are a heterogeneous group of neurodegenerative disorders characterised by progressive cerebellar dysfunction in combination with a variety of other associative features. Since 1993 ADCAs have been increasingly characterised in terms of their genetic mutation and are referred to as spinocerebellar ataxias (SCAs). Some families with ADCA cannot be assigned to any of the known genotypes, which implies further genetic heterogeneity. We investigated the clinical symptoms of 12 patients of a four generation family with ADCA and carried out mutation and genetic linkage studies. The family showed a relatively mild cerebellar ataxic syndrome with cognitive impairment, poor performance on the Wisconsin Card Sorting Test, myoclonus, and a postural irregular tremor of slow frequency. Age at disease onset and severity of cerebellar signs and symptoms suggest anticipation. The genetic loci implicated in ADCA were excluded by mutation analyses (SCA 1, 2, 3, 6, 7, 8, 12) and genetic linkage (SCA 4, 5, 6, 10, 11). We conclude that this family represents a clinically and genetically distinct form of SCA. PMID- 11284129 TI - Uric acid levels in sera from patients with multiple sclerosis. AB - The levels of uric acid (UA), a natural peroxynitrite scavenger, were measured in sera from 240 patients with multiple sclerosis (MS) and 104 sex- and age-matched control patients with other neurological diseases (OND). The mean serum UA concentration was lower in the MS than in the OND group, but the difference did not reach the level of statistical significance (P = 0.068). However, the mean serum UA level from patients with active MS (202.6 + 67.1 mumol/l) was significantly lower than that in inactive MS patients (226.5 + 78.6 mumol/l; P = 0.046) and OND controls (P = 0.007). We found a significant inverse correlation of serum UA concentration with female gender (P = 0.0001), disease activity (P = 0.012) and duration (P = 0.017), and a trend towards an inverse correlation with disability as assessed by EDSS score, which did not reach statistical significance (P = 0.067). Finally, multivariate linear regression analyses showed that UA concentration was independently correlated with gender (P = 0.0001), disease activity (P = 0.014) and duration of the disease (P = 0.043) in MS patients. These findings suggest that serum UA might serve as a possible marker of disease activity in MS. They also provide support to the potential beneficial therapeutic effect of radical-scavenging substances in MS. PMID- 11284130 TI - Rapid distinction of acute demyelinating disorders and central nervous system lymphoma by molecular analysis of cerebrospinal fluid cells. AB - Polymerase chain reaction (PCR) based automated high-resolution fragment analysis of rearranged immunoglobulin heavy-chain genes is a highly sensitive means for identifying clonal B-cell responses. We used this technique to distinguish polyclonal inflammatory from monoclonal neoplastic B-cell populations in the cerebrospinal fluid (CSF) of three patients with acute demyelinating disorders of the central nervous system whose clinical, magnetic resonance imaging (MRI) and CSF features did not permit unequivocal exclusion of primary central nervous system lymphoma (pC-NSL). This approach is highly suitable for detecting CNS inflammation particularly when lymphomatous involvement cannot be ruled out by noninvasive diagnostic procedures alone. PMID- 11284131 TI - Preliminary evidence for neuronal damage in cortical grey matter and normal appearing white matter in short duration relapsing-remitting multiple sclerosis: a quantitative MR spectroscopic imaging study. AB - Neuronal damage and loss is likely to underlie irreversible disability in multiple sclerosis (MS). The time of onset, location and extent of neuronal damage in early disease are all uncertain. To explore this issue 16 patients with short duration, mild relapsing-remitting disease (mean disease duration 1.8 years, median EDSS 1) were studied using short echo time proton magnetic resonance spectroscopic imaging (1H-MRSI) to quantify the concentration of the neuronal marker N-acetyl-aspartate (NAA). The data were compared with those from 12 age-matched controls. 1H-MRSI was obtained from a 1.5-cm-thick slice just above the lateral ventricles. The Linear Combination (LC) Model combined with locally developed software allowed automated measurement of absolute metabolite concentrations from lesions, normal-appearing white matter (NAWM) and cortical grey matter (CGM). MS CGM exhibited significantly lower NAA (P = 0.01) and myo inositol (P = 0.04) than control CGM. MS NAWM exhibited a lower concentration of NAA (P = 0.01) and increased myo-inositol (P = 0.03) than control white matter. More marked reductions in NAA and increases in myo-inositol were seen in lesions. The reduced NAA in MS CGM and NAWM suggest that mild but widespread neuronal dysfunction or loss occurs early in the course of relapsing-remitting MS. This preliminary finding should be confirmed in a larger cohort, and follow-up studies are also needed to determine the prognostic and pathophysiological significance of these early changes. PMID- 11284132 TI - Neurocysticercosis: an unusual presentation of a rare disease. PMID- 11284133 TI - Intracerebral haemorrhage associated with sildenafil citrate. PMID- 11284134 TI - Collet-Sicard's syndrome as a result of jugular vein thrombosis. PMID- 11284135 TI - Idiopathic Lambert-Eaton myasthenic syndrome associated with minimal-change glomerulonephritis and psoriatic arthritis. PMID- 11284136 TI - Rapid resolution of nerve conduction blocks after plasmapheresis in Guillain Barre syndrome associated with anti-GM1b IgG antibody. PMID- 11284137 TI - Myasthenia gravis, corticosteroids and osteoporosis prophylaxis. PMID- 11284138 TI - Pioneers in neurology. Santiago Ramon y Cajal (1852-1934). PMID- 11284139 TI - Neprilysin content is reduced in Alzheimer brain areas. PMID- 11284140 TI - Acanthocytosis and neurological disorders. AB - Acanthocytosis occurs because of ultrastructural abnormalities of the erythrocyte membranous skeleton resulting in reduced membrane fluidity. At least three hereditary neurological conditions are associated with it, although as yet the pathogenesis of the neurological features is unknown. In abetalipoproteinaemia, an autosomal recessive condition, vitamin E deficiency results in a progressive spinocerebellar syndrome associated with peripheral neuropathy and retinitis pigmentosa. Neuroacanthocytosis is also probably an autosomal recessive condition and is characterised by chorea, orofaciolingual dyskinesia, dysarthria, areflexia, seizures and dementia. McLeod syndrome is an X-linked recessive disorder usually presenting in males as a benign myopathy with areflexia, in association with a particular abnormality of expression of Kell blood group antigens. However, occasionally the neurological features are more severe and indistinguishable from those of neuroacanthocytosis. Recent advances in molecular genetics may assist better understanding of the disease mechanisms and the search for more effective treatments. PMID- 11284142 TI - ECG of the month. The hare and the tortoise. Atrial flutter with slow ventricular rate. PMID- 11284141 TI - Behcet's disease: diagnostic and prognostic aspects of neurological involvement. AB - This study was conducted to describe clinical and prognostic aspects of neurological involvement in Behcet's disease (BD). Patients referred for neurological evaluation fulfilled the criteria of the International Study Group for Behcet's Disease. We analyzed disability and survival by the Kaplan-Meier method, using Kurtzke's Extended Disability Status Scale (modified for BD) and the prognostic effect of demographic and clinical factors by Cox regression analysis. We studied 164 patients; of the 107 diagnostic neuroimaging studies: 72.1% showed parenchymal involvement, 11.7% venous sinus thrombosis (VST) and the others were normal. CSF studies were performed in 47 patients; all with inflammatory CSF findings (n = 18) had parenchymal involvement. An isolated increase in pressure was compatible with either VST or normal imaging. The final diagnoses were VST (12.2%), neuro-Behcet syndrome (NBS) (75.6%), isolated optic neuritis (0.6%), psycho-Behcet syndrome (0.6%), and indefinite (11%). VST and NBS were never diagnosed together. Ten years from onset of BD 45.1% (all NBS) reached a disability level of EDSS 6 or higher, and 95.7 +/- 2.1% of the patients were still alive. Having accompanying cerebellar symptoms at onset or a progressive course is unfavorable. Onset with headache or a diagnosis of VST is favorable. Two major neurological diagnoses in BD are NBS and VST. These are distinct in clinical, radiological, and prognostic aspects, hence suggesting a difference in pathogenesis. PMID- 11284143 TI - Management of the preauricular sinus. AB - The preauricular sinus is a relatively common physical finding especially in the pediatric population. It is defined as a congenital lesion in which a small skin opening located in front of the external ear communicates with a subcutaneous network of cysts. The vast majority are benign in nature and require no intervention. Draining sinus tracts are prone to infection and should be excised. Complete excision of the pit and sinus tract provides the only definitive cure. To prevent problematic recurrence, we recommend wide exposure of the lesion by the technique described. PMID- 11284145 TI - The journal 150 & 100 years ago. September 1849 and 1899. PMID- 11284144 TI - Radiology case of the month. Left foot mass. Synovial cell sarcoma. PMID- 11284146 TI - The Medical Assistance Programs Integrity Law (Louisiana's own 'anti fraud and kickback law'). PMID- 11284147 TI - Personality type and medical specialty choice. AB - The July 1999 issue of the Journal of the Louisiana State Medical Society reported results of the Myers-Briggs Type Indicator administered to 1,797 students at Louisiana State University School of Medicine in New Orleans, Louisiana from 1988 through 1998. The current follow-up study has as its subjects 1,262 matriculants who entered the School from 1988 through 1995 and completed their medical undergraduate studies by May 1999. In addition to identification of the graduates' Myers-Briggs type, information on their residency choice was also available. In this study, the authors explore possible association between the graduates' personality type and their chosen career, along with possible type differences of those graduates selecting primary care and those choosing non primary care specialties. PMID- 11284148 TI - Cost of medications for elderly in a nursing home. AB - The rising cost of medical care, particularly for elderly patients in nursing homes, is receiving increasing attention. The purpose of this study was to investigate the cost of and the average number of medications taken per patient in a nursing home. The study included 116 residents of a nursing home in New Orleans, Louisiana. Relevant information about the number of medications taken by each patient on a regular basis was obtained by chart review. The cost of medications was calculated from the Red Book (which represents the cost of drugs to the pharmacy), because the medication prices charged by the nursing home were not available. The number of medications used per patient per day expressed as mean +/- S.D. was 8.1 +/- 4.1. The cost of medications per patient per month expressed as mean +/- S.D. was $182 +/- 141. The estimated annual cost of medications per patient was $2,184. It is highly likely that the cost to the patient is higher than the cost shown here. This preliminary study shows that elderly patients take several medications associated with significant expense. PMID- 11284149 TI - Systemic amyloidosis associated with hepatocellular carcinoma. Case report and literature review. AB - We describe the case of a male patient with biopsy-proven non-resectable liver adenoma at age 32 who presented 17 years later with hepatocellular carcinoma and nephrotic syndrome. Autopsy demonstrated systemic amyloidosis A. Review of the medical literature disclosed only three previous published cases of liver tumors associated with systemic amyloidosis. The association of non-hematologic neoplasias with systemic amyloidosis is rare and our literature review revealed only three cases of systemic amyloidosis in patients with liver tumors. We present here the case of a patient with apparent transition of liver adenoma to hepatocellular carcinoma with associated systemic amyloidosis. PMID- 11284150 TI - [Exposure to pesticides in greenhouses and male fertility]. AB - We attempt to study fertility problems among workers exposed to pesticides, comparing the reproductive experience of greenhouse workers and administrative staff working in the Health Local Units of the same geographical area (reference population). Data on reproductive history and time to pregnancy (TTP) at first pregnancy were collected by personal interview. For workers with children, we collected data on TTP in relation to the occupational risk factors. The analysis of TTP was conducted among 127 greenhouse and 173 administrative workers married and aged 20-55. The greenhouse workers reported 232 pregnancies and a mean number of 1.8 children; the controls 270 and 1.6 respectively. For greenhouse workers the mean TTP in months (5.4 with SD 5.6) resulted longer than for controls (3.9 with SD 5.6). The risk for conception delay (beyond 3 months) by exposure category of the man adjusted for age of woman, smoking of man and woman at first pregnancy resulted 2.4 higher for a subgroup of greenhouse workers with higher exposition (CI 95% 1.2-5.1). PMID- 11284151 TI - [Trawl-fishing: assessment of accidents at the largest Italian marine]. AB - Fishing is a work with high accidents risk. The results of European Community Safety Commission show that in the fishermans the fatal injures are highest than in the manufacturing factory. The authors report the data of working accidents in the fishermans of Mazara del Vallo during the years 1989-98. The study shows 1. an increase the number of accidents in the last years with high incidence of accidents in the extracommunity population of fisherman, 2. the most incidence of injures is concentrated on arms. CONCLUSION: the authors shows that the working organization and the mediocre vocational training are the most reason of the accidents. PMID- 11284152 TI - A comprehensive neurobehavioral and neurophysiological study for low level lead exposed workers. AB - A comprehensive neurobehavioral and neurophysiological study was performed to evaluate the adverse effect of low level lead-exposure, and to compare the sensibility, easiness of the test methods utilized. The tests were: WHO recommended Neurobehavioral Core Test Battery (NCTB), Autonomic Nouvers System Function (ANS) Test Battery, Brain Electricity Active Mapping (BEAM), and Nerve Conduction Velocity. 44 lead-exposed workers were selected, with 34 age, education degree, family economic level, smoking and drinking matched referents. RESULT: The mean blood lead concentration of lead-exposed workers was 1.3870 mumol/L, whereas that of referents was 0.6080 mumol/L, the difference was very significant. The negative Profile of Mood State (POMS) score of lead-expose workers was higher than that of referents, whereas the positive POMS score of the referents was higher than that of lead-exposed group, with a covariance analysis. The lead-exposure affected some NCTB test items, such as simple reaction time (SRT), digital symbol (DSY), correct dots (PAC) and total dots (PA). The heart rate response to Valsalva manoeuvre (HR-V), heart-rate response to deep breathing (HR-DB), and blood-pressure response to immediate standing (BP-IS) were lowered in lead-exposed workers significantly. Some abnormal brain electric waves (dominant beta frequency, semetry-diffuse abnormal and non semetry-diffuse abnormal wave distribution, dominant low wave amplitude) appeared in lead-exposed workers. Left ulnar nerve maximal conduction velocity was significantly lowered in lead-exposed group. CONCLUSION: The NCTB (including POMS), and ANS function test should be the regular screening battery for low level lead-exposed workers. The threshold blood lead concentration for health surveillance should be 30 micrograms/dL, or 1.4 mumol/L. PMID- 11284153 TI - [Health protection of workers in the European legislation of the Rome Treaty today]. AB - Occupational hygiene and prevention of occupational accidents and diseases are institutional objectives of the social policy of the European Union as stated in the Treaty establishing the European Community (firstly Treaty of Rome, signed on 25 March 1957). In this report the legislative activity of the European Community aimed at the improvement of the working environment to protect workers' health and safety is considered through its historical development, up to the present situation. The first part of the report briefly describes the Institutions established by the European Community, with specific tasks relating to occupational hygiene and health and safety at work. Then European laws grouped in categories according to their subjects are considered, especially with regard to the development and progress of the legislation, to point out main features and historical trends. PMID- 11284154 TI - [Analysis of risk of biological accidents at a urban health area]. AB - The makers have examined 455 fingerprint cards of accidents at biologic risk needed in a sanitary structure from the month of november 1995 to the month of december 1998. After they have described the protocol of the sanitary supervision applied, the procedure of the accidents, the qualifications, and the departments mainly interested by the subject of study event, they have pointed out the need of a greater vaccinable covering against the virus B, parvying attention to the "non responders" subjects, thinking also it's necessary bigger resources for the personnel, training and information. PMID- 11284155 TI - [Responsibility (according to the legislative act "626/1994") in the implementation of preventive measures in the preparation of antiblastic drugs]. AB - The 7th title of the legislative act "626/1994" (and the other normative indications) identifies cavcingenic agents in occupational uses, and also the protections of workers in exposure. Among these agents there are any antiblastic drugs. This is a difficult problem for a correct application of normative indications: it's no possible to replace cavcingenic agents with no cavcingenic agents, as so as it's impossible to operate on "closed cycle". The authors put the attention on the other very important normative indications: "preparations in bordered and isolated room", "minimal exposure", "severe procedures and internal normations", formal "identification of the responsibilities" in the hospital organization (also with the strictly identification of managers and foremen). PMID- 11284156 TI - [Isokinetic assessment and mid-term work reincorporation of patients surgically treated with the shoulder Latarjet technique]. AB - We studied a group of 12 subjects with chronic posttraumatic scapulo-humeral instability who underwent arthroscopic coracoid transposition using Latarjet's technique. All patients were followed-up after the operation for a mean of 1.7 years (minimum 1 year, maximum 3 years). The follow-ups consisted of a clinico functional evaluation with a apprehension test and isokinetic studies of strength and joint movements during internal and external rotation of the abducted shoulder, both on the healthy side and on the operated side. The isokinetic evaluation consisted of 5 repetitions of internal-external rotation at ana angular velocity of 60 degrees/sec. The results showed that the operated shoulder was slightly weaker than the healthy one, but that the difference was not statistically significant according to Student's t test for paired data. The profile of the trace did not show macroscopic differences between the two sides. At clinical assessment 11 of 12 patients reported a good-excellent outcome of the operation with complete recovery of shoulder function and return to previously performed sporting activities. In conclusion, this technique is shown to have a good medium-term outcome. Although there have been no complications in our series, a longer follow-up to evaluate possible complications would be advisable. PMID- 11284157 TI - [Clodronate in erosive osteoarthrosis of the hand: efficacy for pain and function recovery]. AB - 29 patients suffering from erosive osteoarthritis of the hands (26 women with a mean age of 61.47 years and 3 men with a mean age of 53.3 years) were treated during painful episodes with cyclical intravenous clodronate (300 mg for slow infusion for 7 days). We administered 73 cycles after which the pain, evaluated by VAS' score, decreased from 5.63 +/- 2.14 to 2.93 +/- 1.95 (p = 0.0001); the pain reduction lasted for a mean of 88.33 days. In 16 out of those 73 cycles were measured also: the strength of the hands (mmHg) which increased, on the right from 169.38 +/- 75.85 to 190.31 +/- 81.76, on the left from 180.56 +/- 68.27 to 196.11 +/- 85.55 (p = n.s.); the number of painful joints which decreased from 4.75 +/- 2.52 to 2.56 +/- 1.93 (p = 0.0011); the number of swollen joints which decreased from 3.07 +/- 2.69 to 2.67 +/- 2.61 (p = n.s). These data prove the efficacy of the clodronate in treating painful episodes of erosive osteoarthritis and suggest that bisphoshonates could play an important role in the rehabilitation of this disease. PMID- 11284159 TI - [National health 1998-2000]. PMID- 11284158 TI - [Efficacy of a chair with magnets in the prevention of musculo-skeletal disorders caused by prolonged sitting]. AB - The aim of the present paper was to evaluate the effectiveness of a sitting system based on the application of magnetic fields in the prevention of posture related musculoskeletal disorders. We studied 5 healthy male volunteers during laboratory simulation tests including 60-minute sessions of "driving" and computer-work. Two subjects performed a 60-minute driving test on an isotonic dynamometer with a steering wheel, in which they had to steer for 2 min and drive normally for 3 min, alternatively. They repeated the test with and without the magnet-based sitting system. Three subjects performed a 60-minute computer-work test in a seated position with and without the magnet-based sitting system. EMG activity was registered in the trapezius muscles and at L1 and L5 lumbar level bilaterally with surface electrodes. The EMG trapezius activity continuously recorded was analysed with the APDF method, as proposed by Jonsson. Before and after the tests, the subjects performed a 60-sec isometric back extension at 60% MVC with a specific back dynamometer, while paraspinal EMG was recorded. The slope of decay of the median frequency of the EMG power spectrum was then calculated as an index of localised muscle fatigue. The results showed a decreased myoelectric activity both at shoulder and lumbar level by using the magnet-based sitting system for prolonged seated work tasks. Thus, the system appears to be an effective tool in preventing muscle contractures secondary to prolonged, constrained positions. PMID- 11284160 TI - Glycohistochemistry: the why and how of detection and localization of endogenous lectins. AB - The central dogma of molecular biology limits the downstream flow of genetic information to proteins. Progress from the last two decades of research on cellular glycoconjugates justifies adding the enzymatic production of glycan antennae with information-bearing determinants to this famous and basic pathway. An impressive variety of regulatory processes including cell growth and apoptosis, folding and routing of glycoproteins and cell adhesion/migration have been unravelled and found to be mediated or modulated by specific protein (lectin)-carbohydrate interactions. The conclusion has emerged that it would have meant missing manifold opportunities not to recruit the sugar code to cellular information transfer. Currently, the potential for medical applications in anti adhesion therapy or drug targeting is one of the major driving forces fuelling progress in glycosciences. In histochemistry, this concept has prompted the introduction of carrier-immobilized carbohydrate ligands (neoglycoconjugates) to visualize the cells' capacity to be engaged in oligosaccharide recognition. After their isolation these tissue lectins will be tested for ligand analysis. Since fine specificities of different lectins can differ despite identical monosaccharide binding, the tissue lectins will eventually replace plant agglutinins to move from glycan profiling and localization to functional considerations. Namely, these two marker types, i.e. neoglycoconjugates and tissue lectins, track down accessible binding sites with relevance for involvement in interactions in situ. The documented interplay of synthetic organic chemistry and biochemistry with cyto- and histochemistry nourishes the optimism that the application of this set of innovative custom-prepared tools will provide important insights into the ways in which glycans can act as hardware in transmitting information during normal tissue development and pathological situations. PMID- 11284161 TI - An immunohistochemical study on the endocrine cells in the alimentary tract of the red-eared slider (Trachemys scripta elegans). AB - The regional distribution and relative frequency of endocrine cells in the alimentary tract of the red-eared slider, Trachemys scripta elegans, were investigated by immunohistochemical methods using 10 antisera. Most of the immunoreactive cells in the intestine were spherical or spindle-like in shape (open-type cells), while round cells (closed-type cells) were occasionally found in the stomach. These immunoreactive cells were located in the basal portion of the intestine, including the oesophagus, and in the gastric glands of the stomach. Cg A-immunoreactive cells were restricted to the pylorus and duodenum and were few in number. Serotonin-immunoreactive cells, which were most commonly found in the pylorus, were found in the epithelia throughout the alimentary tract at various frequencies. Gastrin-immunoreactive cells were found in the pylorus, duodenum and jejunum at moderate, low and very low frequencies, respectively. Somatostatin-immunoreactive cells were found throughout the alimentary tract except for the rectum, at various frequencies. Glucagon-immunoreactive cells were detected in the fundus, pylorus, jejunum and ileum at low or very low frequencies. CCK-8-immunoreactive cells were found in the pylorus, fundus and duodenum at very low, low and moderate frequencies, respectively. Bombesin immunoreactive cells were restricted to the fundus and pylorus at low frequencies. No secretin-, BPP- or VIP-immunoreactive cells were found in this study. PMID- 11284162 TI - Skull typology of adult male Kangal dogs. AB - In this study, a total of 16 skulls of the adult male Kangal dog were used. Craniometric measurements for 44 different parts of the skull were made. All investigated features were expressed as mean +/- SD. Cephalic indices and ratios were calculated. These indices and ratios have been compared with the average values of indices calculated for other breeds. A skull index of 50.29 +/- 1.033, a cranial index of 46.05 +/- 2.213, a facial index of 99.62 +/- 3.891, a facial index-1 of 81.67 +/- 3.667, a basal index of 28.71 +/- 1.425, a basal index-1 of 57.91 +/- 1.365, a length-length index-2 of 1.08 +/- 0.045, a length-width index 2 of 1.99 +/- 0.041, a length-width index-4 of 2.18 +/- 0.108, a palatal index-1 of 62.24 +/- 2.528, a palatal index-2 of 65.37 +/- 2.208, a palato-basal ratio of 55.44 +/- 1.975, a palato-basal ratio-1 of 54.47 +/- 1.716, a palato-palatine ratio of 33.71 +/- 0.860, a palato-palatine ratio-1 of 34.30 +/- 0.733, a cranio facial ratio of 107.87 +/- 4.819 and a cranio-facial ratio-1 of 144.17 +/- 8.099 were obtained. When the skull, cranial and facial indices were considered together with the other calculated indices and ratios, it was clear that the skulls of the Kangal dogs have to be regarded as of dolichocephalic type. Kangal dogs, with their mastiff-like appearance and massive head, are shown in this study to be typical of a dolichocephalic breed. PMID- 11284163 TI - Immunohistochemical detection of components of the insulin-like growth factor system during skeletal muscle growth in the pig. AB - The insulin-like growth factor (IGF) system plays an important role in postnatal somatic and skeletal muscle growth in pigs. There is little information on the occurrence and distribution of components of the IGF system in postnatal porcine skeletal muscle. IGF-I, IGF receptor 1 (IGF1R) and the IGF-binding proteins IGFBP 1 and -3 in longissimus dorsi and triceps brachii were localized in muscle biopsies from 12 commercially crossbred pigs aged from 28 to 199 days as well as from the sire generation, by immunohistochemistry. Plasma IGF-I concentrations were also determined using radio-immunoassays. Unlike other species, IGF-I was localized in porcine skeletal muscle fibres. Staining intensity correlated with the highest plasma IGF-I levels and phases of intensive muscle growth from the 11th to 22nd week. The pattern of IGF1R immunostaining, which was strong, correlated with that of IGF-I, IGF1R was also localized in endomysial tissues. IGFBP-1 was not detected within muscle fibres, but was found in the endomysium and vessel walls, while IGFBP-3 was localized with IGF-1 and its receptor. Higher magnification revealed that IGF1R, IGFBP-3 and probably IGF-I appeared in the tubular system. Inhibitory as well as stimulating controls of IGFBP-1 and -3 on IGF functions are discussed, which may maintain a balance between autocrine growth promoting activities of IGF-I and IGF1R. PMID- 11284164 TI - A quantitative study on the trachea of the dog. AB - This study was carried out to record the detailed morphometric structure of the trachea in dogs using 15 female and four male healthy adult mongrel dogs. The diameter and thickness of each tracheal ring were measured, the number of tracheal rings varying from 36 to 45. All data were subjected to statistical analysis which was carried out on individual sections of the trachea, i.e. the cranial cervical, middle cervical, thoracic inlet and the intrathoracic tracheal regions, which consisted of 12, 12, nine and 12 tracheal rings, respectively. Fusion of the tracheal rings was especially obvious in the cranial cervical and thoracic inlet regions as a result of neck movements. The diameter and thickness of the tracheal rings are smallest at the thoracic inlet level because the direction of the trachea changes at this point where the thoracic inlet is relatively small and surrounded by bone. The ratios of inner transverse to inner vertical and outer transverse to outer vertical diameters were almost the same, between 1.14 and 1.25 in all regions, which indicated that the trachea is near circular in shape in the dog. At the thoracic inlet level cross-sectional lumen areas are 7 and 6% smaller than those in the middle cervical and intrathoracic regions, respectively. The thinnest cartilage was seen at the thoracic inlet level where there is a risk of tracheal collapse. PMID- 11284165 TI - Indications for adrenalectomy in the laparoscopic era. AB - The tilt from open to laparoscopic surgery seems to be definite in most adrenal disorders. The aim of this study is to evaluate the current indications for laparoscopy and the persistent indications for open adrenalectomy, as seen in our experience and in the literature data. Between January 1985 and December 1999, 486 patients were operated on for adrenalectomy. Since January 1994, 91 laparoscopic adrenalectomies were performed in 84 patients. The Authors retrospectively evaluated the indications for laparoscopy in 55 patients (45.9%) and for open adrenalectomy in 65 others (54.1%) operated on in the last three years. Exclusion criteria for the laparoscopic approach included clinical suspicion of malignancy and tumour size greater than 6 cm, in the 38.5% and in the 23.0% of cases respectively. In Authors experience the laparoscopic adrenalectomy is the procedure of the choice for the surgical removal of non malignant, unilateral or bilateral tumours under 6 or 7 cm. The laparoscopy is not a radical operation for cancer. Open surgery is always indicated for large and malignant tumours. The fascinating feature of laparoscopy has not to change the indications and the surgeon must plan the appropriate approach for every single patient. PMID- 11284166 TI - [Lymphadenectomy in stomach carcinoma: is this a new trend?]. PMID- 11284167 TI - Gastro-intestinal stromal tumor (GIST): case report. AB - Gastro-intestinal stromal tumors (GISTs), as currently defined, represent the largest category of primary non epithelial neoplasms of the gastrointestinal tract. They arise from mesenchymal cells located in the wall of the organ and show a remarkable variability in their differentiation pathways. For this reason there is relevant degree of confusion in their interpretation. On the basis of immunohistochemical and ultrastructural studies these neoplasms are divided into several categories: leiomyomas, schwannomas and less differentiated tumors referred as GIST. In the small bowel GIST are uncommon. Usually asymptomatic, they could be the cause of surgical emergencies like massive bleeding, obstruction, intussusception or perforation. Generally benign, an higher percentage of malignant cases are described in the small bowel. The Authors report a case of malignant GIST of the small intestine presented with bowel obstruction by ileal invagination. In this case, as usually it happens in malignant GIST, the final diagnosis was obtained by an abdominal surgical exploration. PMID- 11284168 TI - [Gastroduodenal artery pseudoaneurysm ruptured at the pancreatic head]. AB - The presence of splancnic aneurysms associated with pancreatitis represents an uncommon evidence (10%) but extremely formidable for the high mortality related to the elevate risk of rupture (50%). A case of a broken gastroduodenal artery pseudoaneurysm plugged in the pancreatic head in a patient with chronic pancreatitis surgically treated is reported. The Authors believe that in presence or in suspicious of peripancreatic pseudoaneurysm bleeding, showed by abdominal echography or CT scan, is mandatory the execution of splancnic and peripancreatic vessels angiography to determine the correct localization of the aneurysm, essential to determining the best surgical treatment. Gastroduodenal artery before the origin from the right epatic artery has been tied in presence of an anatomic variant of origin and division of the hepatic arteries, previously showed with the angiographic examination. The exclusion and the complete thrombosis of the false aneurysm was demonstrated with the intraoperatory Doppler control and confirmed by CT scan before the dismission. This surgical strategy avoid a pancreatic resection, potentially burdened from an higher risk of mortality and morbidity as than the artery exclusion. PMID- 11284169 TI - [Pancreatic pseudocysts. Our experience with 21 cases treated surgically]. AB - Twenty-one consecutive patients with pancreatic pseudocyst have been reviewed. Nine cases have been treated with internal drainage, 9 with external drainage, while 3 patients have undergone distal pancreatectomy. No mortality was associated to the surgical treatment, while morbidity was of 9.5% due to pancreatic fistulas. Based on their own experience and literature review the Authors describe clinical, diagnostic and therapeutic features of the disease and point out the good results of cistojejunostomy with Roux-en-Y loop. PMID- 11284170 TI - [Defect of the broad ligament as a cause of internal hernia]. AB - Herniation of small bowel trough a defect of the broad ligament is an extremely rare event, more over in women never had surgical operations. Pathogenetic hypothesis are considered. Differential diagnosis may be difficult. The Authors report a recent observation of such small bowel herniation trough the left mesosalpynx. PMID- 11284171 TI - [Ectopic thyroid: report of a case]. AB - The development of thyroid tissue can occur in any moment of the migration of the thyroid along the thyroglossal duct from the tongue, resulting in lingual (at tongue base), sublingual (below the tongue), prelaryngeal (in front of the larynx), and substernal (in the mediastinum) ectopy. Thyroglossal duct cyst is the most common type of clinical abnormality related to thyroid ectopy. Surgical removal of such ectopic tissue is justified since some Authors describe thyroid cancer arising from aberrant thyroid tissue. The Authors report a case of thyroid ectopy in a patient who underwent 20 years before a left thyroid resection with isthmectomy; during the operation the surgeon described a hypertrophic pyramidal lobe which was left in situ and the patients did not receive any hormone suppressing therapy. PMID- 11284172 TI - [Radiofrequency ablation: a new approach in the treatment of hepatocellular carcinoma]. AB - HCC is a tumor with increasing incidence that usually develops on cirrhotic liver; therefore the prognosis depends on both tumor size and liver function. HCC generally shows a slow growth and (not very important) symptoms; so, the periodic surveillance of cirrhotic patients, by using US examination and alpha-fetoprotein level, allows an early diagnosis of the tumor. Several techniques have proved useful in the treatment of HCC but, in comparison with other currently available percutaneous therapies, RF ablation appears to have several advantages. Authors' results suggest that RF ablation is an effective and safe procedure for the therapy of local hepatic neoplasms. However, further studies will be required to demonstrate that RF ablation is more effective than percutaneous ethanol injection (PEI) in the treatment of HCC. PMID- 11284173 TI - [Urgent cholecystectomy in acute cholecystitis: laparoscopy or laparotomy?]. AB - Early cholecystectomy is the best policy in the case of acute cholecystitis. The aim of this retrospective study is to evaluate the current treatment of choice of acute calculous cholecystitis, as seen in our experience and in the literature data. Between January 1997 and July 2000, 150 patients were operated on for cholecystectomy. In the group of 30 patients (20%) with acute cholecystitis, 15 patients (50%) were managed with laparoscopic approach while 15 patients (50%) with traditional operation. At the beginning the Authors chose the open via for understand the pathologic findings of acute cholecystitis, then they always preferred the laparoscopic approach. Comparison between two groups concerned the interval between onset of symptoms and operation, postoperative mortality and morbidity rates, postoperative hospital stay and follow up. Statistical analysis was performed by the Student's t-test and the chi-square test. Both groups were homogeneous with regard to sex, age and onset of symptoms. There were no deaths and morbidity rate in the laparoscopic group was 20% versus 40% (p = ns). The average postoperative hospital stay in the laparoscopic group was 5.6 days versus 10.5 days (p = 0.046). The conversion rate into laparotomy was 6.6% (1 case). There has been one case of incisional hernia in the open group at a mean follow up of 20 month. Early laparoscopic cholecystectomy is the treatment of choice of acute cholecystitis because of a lower postoperative morbidity rate and a significant shorter hospital stay. PMID- 11284174 TI - Risk factors associated with bronchial asthma in school going children of rural Haryana. AB - Bronchial asthma is one of the most common illnesses in children. Factors influencing development of asthma have not been studied in rural population. 2000 school going children from five schools of Chhainsa and Dayalpur Primary Health Centre area in Ballabgarh Block of Haryana state were screened for presence of symptoms of asthma using a questionnaire suggested by International Study of Asthma and Allergy in Children (ISSAC). 40 children were identified as cases of bronchial asthma. For each child with asthma two age and sex matched non asthmatic controls were selected from the study population. History, clinical examination and in-depth interview were carried out for all cases and controls. Factors associated with presence of symptoms of asthma on multivariate analysis were passive smoking (OR 3.33, 95% CI 1.85-7.65), pets at home (OR 5.5, 95% CI 1.04-29.15), and absence of windows in living rooms (OR 4.03, 95% CI 1.17-13.79). Factors such as family history of asthma, history of worm infestation, fuel used for cooking, location of kitchen and food allergy were not significant on statistical analysis. Thus, passive smoking, inadequate ventilation and pets (dogs and cats) at home are significant risk factors associated with presence of symptoms of asthma in rural children. PMID- 11284175 TI - IDDM and early exposure of infant to cow's milk and solid food. AB - Associations studies were attempted between the type of feeding, duration, and time of starting of solid foods in infancy and the incidence of insulin-dependent diabetes mellitus (IDDM). The study subjects comprised 52 IDDM patients and 52 control subjects matched for sex, age, social status, country, geographical location and selected from pediatric departments of different hospitals in Tehran. Diabetic children (21 boys, 31 girls) were of the ages of 1.5 to 14 years. Information about the pattern of their feeding at the first two years of life were collected through questionnaires administered to the mothers. The questionnaire was designed to evaluate the duration of complete or partial breast feeding and the age at which dietary products containing cow's milk were introduced into the diet. A large proportion of the diabetic children rather than the control children had been breast-fed, and the risk of IDDM among children who had not been breast-fed was below unity. No significant difference in the duration of breast-feeding was observed between diabetic and control group. Our data do not support the existence of a protective effect of breast-feeding on the risk of IDDM, nor do the data indicate that early exposure to cow's milk and dairy products has any influence on the development of IDDM. PMID- 11284176 TI - Epilepsy in children with cerebral palsy. AB - This article deals with the clinical profile of children with cerebral palsy and epilepsy, and to study the clinical predictors of response to anti-epileptic drugs. It is a prospective hospital based follow-up study. All the children who presented with cerebral palsy and history of seizure (other than neonatal seizures) over a period of one year were included. Seizures were classified according to ILAE classification. An EEG was obtained in all cases. Neuroimaging was done in all patients. Eighty-five patients were studied and followed for minimum of 12 months. Perinatal factors accounted for 62 (72.3%) cases. The motor deficits seen were quadriparesis (n = 64), hemiplegia (n = 12) and diplegia (n = 9). Associated mental retardation was seen in 80.9% patients with quadriparesis. A predominance of generalised epilepsy was seen with generalised tonic clonic seizures (32.9%) followed by mycolonic seizures (30.6%) and localisation related epilepsy (24.7%). The patients with quadriparesis were more likely to have generalised epilepsy and 52.4% of them required two or more anti-epileptic drugs for control of seizures. Patients with hemiplegia had localisation related epilepsy in 83.3% of cases. On multivariate analysis presence of quadriparesis, microcephaly, mental retardation and myoclonic epilepsy were found to predict the poor response to AED. Epilepsy in patients with cerebral palsy is of severe nature and difficult to control. Presence of quadriparesis, mental retardation and myoclonic seizures was predictive of poor response to anti- epileptic drugs. PMID- 11284177 TI - Teratomas in infancy and childhood. AB - A 10-year-experience with 42 cases of teratomas in paediatric age group is presented. The commonest type of teratoma was sacrococcygeal followed by ovarian and retroperitoneal teratomas. An analysis of clinical profile, malignant potential, management, prognostic factors and follow up is discussed with review of literature. PMID- 11284178 TI - Sister-chromatid exchange analysis on long-term benzathine penicillin for secondary prophylaxis of rheumatic fever. AB - A single intra-muscular injection of 1.2 millions units of benzathine penicillin every 4 weeks is the most widely used method for the antibiotic prophylaxis of rheumatic fever. The aim of this study is to evaluate the effect of long-term benzathine penicillin on DNA in patients with rheumatic fever. Thirty children with confirmed rheumatic fever who were on the benzathine penicillin prophylaxis were enrolled in the study, and 30 similar normal children served as a control group. To detect any DNA damage, SCE analysis were performed in circulating lymphocytes of the subjects. A statistically significant increased frequency of SCE was observed in children on the benzathine penicillin prophylaxis (no = 30, mean SCEs/cell +/- SD 7.54 +/- 1.81) as compared to a control group (no = 30, mean SCEs/cell +/- SD 5.82 +/- 1.40). It has been suggested that the difference in the SCE frequencies was induced by the administration of the benzathine penicillin for a long time, and further investigations are needed to confirm this toxic effect. PMID- 11284179 TI - Organoleptic study of deacidified and deodourised palm oil. AB - Deficiency of vitamin A has long been identified as a serious and preventable nutritional disorder, associated with increased risk of mortality and morbidity amongst children. The present study was conducted with the objectives (i) to perform organoleptic testing of food products cooked in Deacidified and Deodourised Palm Oil (DDPO), by sensory evaluation method and (ii) to compare the characteristics of these food products with the same products cooked in routinely used oil. Eleven commonly used weaning food items were prepared with routinely used oil (Group A). The same recipes were also prepared with DDPO (Group B). A food testing panel conducted the sensory evaluation for assessing the acceptability of the various food items. It was observed that with respect to all characteristics there was no significant difference in the recipes made with the two types of oil. Results Indicated that DDPO can be used in India for preparation of weaning foods which are routinely given to young children. PMID- 11284180 TI - Safe blood transfusion practices. AB - The advent of AIDS has raised a concern regarding transfusion transmitted diseases. Blood transfusion is safer than ever before through continued improvements in safe donor recruitment, screening of donors, testing of donated blood and appropriate clinical use of blood. The risk of residual infections is further reduced through inactivation of pathogens in blood components. Prevention of technical and human errors in blood grouping, avoiding bacterial contamination of blood components and using leuco-depleted products to minimize immunomodulatory effects also increase blood safety. For safety, efficiency and effectivity it is necessary to improve clinical transfusion practices through alternatives to traditional hemotherapy such as autologous transfusion and audit of blood utilization practices by hospital transfusion committees. PMID- 11284181 TI - Transfusion reactions. AB - Blood components are indicated in a wide variety of disease states. Although most transfusion therapies are administered uneventfully, there are a number of potential adverse transfusion reactions, some of which can assume serious dimensions. These reactions could occur during or even days after a transfusion. A brief description of the adverse effects of transfusion therapy has been outlined in this review. The etiopathogenesis, recognition and treatment of the adverse transfusion reactions have been highlighted. It is imperative that each transfusion of blood components has components be monitored carefully. Prompt recognition of an adverse event and early institution of remedial measures would help in decreasing transfusion related morbidity and mortality. PMID- 11284182 TI - Autologous blood transfusions. AB - Due to enormous risks of transfusion-transmitted diseases in allogenic blood transfusions, including dreaded AIDS, there has been constant endeavour to look for a safer alternative. Autologous transfusion which is transfusion of blood/component donated by intended recipient, has proved to be a safe and viable alternative. Initially tried in 1874 in the form of blood salvage, the process has become popular since 1971 with better PVC containers and storage facilities. Due to ignorance and lack of interest the procedure in still unpopular in India. It is very useful for preventing complications of allogenic transfusion and in rare blood group or patients for whom it is difficult to find compatible blood. Since 1980s the procedure is being widely used. Maximally used category is preoperative donation. Strict protocols regarding selection, storage and labelling are essential. There has been tenfold increase in the preoperative autologous donations in many centres. Isovolemic hemodilution and blood salvage are also being used. The procedure has also been used for preteenage and paediatric ages successfully. 1 to 5 units can be collected from single donor. Now the popularity of the procedure has increased due to awareness and interest. PMID- 11284183 TI - Massive blood transfusion. AB - Pediatricians in the hospital setting must frequently treat children who require massive transfusion (MT) in a variety of clinical situations ranging from major trauma to neonatal hyperbilirubinemia. After identifying the need for massive transfusion, the pediatrician must select the appropriate blood components. Different blood components have specific temperature, preservative, and time requirements for their storage. Changes, termed storage lesions, occur over time in blood components during storage; biochemical changes include decreased levels of 2,3-DPG, a decrease in pH, and an increase in supernatant potassium (K+) with a concurrent decrease in intracellular K+. These changes may affect the function and the viability of components. Additionally, physical changes such as deformation of the red cell membrane occur during storage. Knowledge of these storage lesions is necessary for the pediatrician to make the most appropriate decisions regarding the preparation and selection of components during MT. Serious complications of MT include hemostatic abnormalities, biochemical/metabolic abnormalities, hypothermia, mechanical injury and the effect of Rh incompatibility, each of which has a specific management response. Pediatricians need to be aware of the potential complications associated with massive transfusion, to take measures to prevent them when possible, to anticipate additional transfusion requirements, and to know how to manage them in the pediatric patient. PMID- 11284184 TI - Management of hemophilia in developing countries. AB - Coagulation disorder are common in India. In the absence of any epidemiological studies it is expected that there are at least 50,000 severe hemophilics in our country. The factor concentrates are not easily available in developing countries which poses a major problem while managing severe bleeding episodes in these patients. Various strategies which could be useful while managing these cases in developing countries have been reviewed. PMID- 11284185 TI - Blood support for pediatric surgery. AB - Increasingly complicated surgeries are being performed on neonatal and pediatric patients. Provision of a safe and adequate blood supply is essential for the success of many of these surgeries. Depending on the clinical situation, autologous and/or allogeneic blood may be used. However, in all cases, every attempt should be made to minimize the number of donor exposures to reduce the risk of transfusion transmitted infections. Transfusion of neonatal and pediatric patients requires additional considerations too, such as the risk of graft vs host disease, cytomegalovirus infection, the effects of various preservative anticoagulant solutions, electrolyte levels during blood storage, and wheather or not leukoreduced components are indicated. PMID- 11284186 TI - Haemolytic disease of newborn. AB - Hemolytic disease of the newborn (HDN) occurs due to maternal IgG antibodies crossing the placenta thereby producing hemolysis mainly due to Rh, ABO and Kell groups. A systematic approach to the Rh HDN involves an obstetric history of previous isoimmunized baby, timing and regular monitoring of maternal Rh antibodies and pigment assay of amniotic fluid. Timely decision regarding in utero transfusion and early termination of pregnancy based on the maternal monitoring has radically improved the outcome of these babies. Antenatal prophylaxis with anti D has resulted in great reduction in the magnitude of Rh problem. The fetal blood sampling and in-utero intravenous transfusions has made it possible for almost 100% survival of isoimmunized pregnancies without hydrops. Alternative methods--IVIG and plasma exchange are still of limited application. ABO HDN though common is not a serious form of disease and dose not warrants invasive antenatal monitoring. Anti-Kell is found in patients having received multiple transfusions and the rapid progress of hemolysis in them may not allow such systematic follow up as in Rh HDN. PMID- 11284187 TI - Spontaneous expulsion of unusual tracheobronchial foreign body. AB - An extremely rare case of long, thin and sharp pin in a young boy which was inhaled initially and defied removal at branchoscopy was eventually recovered in stool after a long and variable course through alimentary tract has been reported. PMID- 11284189 TI - Pseudoexstrophy in a female child. AB - A six-year-old female patient presenting with a swelling in the infraumbilical part of the abdomen, bulging out on straining, was diagnosed to have pseudoexstrophy bladder. The urinary tract was normal. The patient had bifid clitoris. There was no other associated malformation. Surgical repair of abdominal wall defect was done successfully. A new classification of exstrophy variants is proposed. PMID- 11284188 TI - The use of L-arginine [correction of F-arginine] and phosphodiesterase inhibitor (dipyridamole) to wean from inhaled nitric oxide. AB - A full-term, female neonate developed acute hypoxemic respiratory failure complicated by persistent pulmonary hypertension of the newborn (PPHN), and responded to high-frequency oscillatory ventilation (HFOV) and inhaled nitric oxide (iNO). Discontinuation of iNO was attempted three times and was followed by severe desaturation due to right-to-left shunt through the patent ductus arteriosus and patent foramen ovale. As a result of iNO dependency state and rebound pulmonary hypertension, the neonate was maintained on iNO therapy for dipyridamole alone was unsuccessful. However, successful discontinuation of iNO therapy was achieved by combination of L-Arginine and dipyridamole. Exogeous NO may lead to down regulation of endogenous NO production, and further lead to rapid hydrolization of cyclic guanosine 3', 5' monophosphate (cGMP), the smooth muscle relaxant, by the enzyme phosphodiesterase. Moreover L-Arginine, the precursor for the formation of endogenous NO, has been found to be deficient in neonates with PPHN, so we speculated that by inhibiting phosphodiesterase and administrating L-Arginine smooth muscle relaxation occurred, and consequent weaning from iNO was achieved. PMID- 11284190 TI - Rickets presenting as pseudotumour cerebri and seizures. PMID- 11284191 TI - Substance abuse in children and adolescents. PMID- 11284192 TI - Long-term care in the United Kingdom: community or institutional care? Individual, family, or state responsibility? PMID- 11284193 TI - Flexible work schedules, older workers, and retirement. AB - Older workers in the United States indicate that they would prefer flexible work arrangements rather than abrupt retirement, yet management has done very little to make this possible. A review of two bodies of literature from the late 1980s is presented: social science writings including sociological, gerontological, and economic literature, and business and management literature. There is a clash between the way jobs are traditionally scheduled and the needs of growing numbers of older workers. Workers continue to be subject to obstacles to phased retirement due to the structuring of health care and pension benefits, downsizing, organizational inflexibility, and "corporate culture." Thus, general views among social scientists regarding the desirability of flexible schedules toward retirement will not produce real changes unless management becomes committed to such changes and they are securely embedded in company policies. PMID- 11284194 TI - Nursing home ownership: an historical analysis. AB - The need to care for dependent elderly in the United States has been a constant since colonial times. Today, as in the earliest days, most care is provided at home by family members. Personal and health services outside the home are sometimes provided by nursing homes. The nursing home industry is large, dominated by private, for-profit ownership, and receives much of its income from public funds. Why are nursing homes publicly funded? Why are nursing homes privately rather than publicly owned? Why is ownership for-profit or proprietary rather than not-for-profit or voluntary? The answers to these questions are found in the history of social policies in the United States. PMID- 11284195 TI - Israel's long-term care insurance law after a decade of implementation. AB - Israel's Long-Term Care Insurance (LTCI) law has been in effect for a decade. It is timely to review the effects of this legislation with a view to identifying possible directions for reform and lessons for other countries considering the introduction of a similar social insurance scheme. The paper considers the law's effects in terms of the size and characteristics of the beneficiary population, the coverage of the scheme, its financial standing, the rate of institutionalization of the elderly, the caregiving burden, the service delivery system, and the overall scope of long-term care services for the aged. Israel's experience has lessons for financing arrangements, target efficiency, service delivery arrangements, and the construction of the burden of care. PMID- 11284196 TI - Grandparents raising grandchildren: challenges faced by these growing numbers of families and effective policy solutions. PMID- 11284197 TI - Sweden and the United States: is the challenge of an aging society leading to a convergence of policy? AB - The aging of the population is one of many forces behind a current reconstruction of welfare benefits in both Sweden and the United States. While both countries represent ideological polarities regarding social policy, they are struggling to meet their welfare goals with limited resources, and both are adopting similar strategies, for example, decentralization, targeting, and an increased emphasis on privatization and evaluation. This paper summarizes some of the differences between Sweden and the United States and describes some of the forces at work that are lessening the differences between the two countries in strategies and policy regarding care services for elderly people. PMID- 11284198 TI - A new double hole pencil point atraumatic needle for amniocentesis. PMID- 11284199 TI - Functional properties of Na,K-ATPase, and their structural implications, as detected with biophysical techniques. AB - A full understanding of the molecular mechanism of ion transport and energetics of the Na,K-ATPase will require both structural and functional data. During recent years biophysical methods have provided a number of important pieces of information on ion binding and release and the charge transfer process. This allows the formulation of kinetic models of the transport process. Although a breakthrough has not been obtained due to the lack of detailed knowledge on the three-dimensional structure with a resolution high enough to identify the ion binding sites and the transport pathway, the functional information has structural implications that create constraints on possible mechanisms of active transport. Here we describe briefly contributions of some biophysical methods to our conceptual understanding of the ion transport process. PMID- 11284201 TI - Formation of folds and vesicles by dipalmitoylphosphatidylcholine monolayers spread in excess. AB - Lipid monolayers exist in several biological systems, including the stratum corneum of the skin, the fluid tear film of the eye, the Eustachian tube of the ear, and airway and alveolar pulmonary surfactants. In this paper, the monolayer to-bilayer transition was studied using dipalmitoylphosphatidylcholine (DPPC) as the model. Depositing DPPC organic solvent solutions in excess at an air:buffer interface led to the formation of elongated structures which could be imaged on carbon grids by transmission electron microscopy. The structures appeared to be DPPC folds protruding into the sol. The structures were frequently ordered with respect to one another, suggesting that they arose during lateral compression due to excess DPPC and are characteristic of a type of monolayer collapse phase. In some cases, series of short folds in an extended line and series of vesicles in line or parallel to the folds were observed. This suggests the elongated folds are unstable and can resolve by forming vesicles. Fold formation occurred at defined lipid concentrations above which more vesicles were observed. Surfactant protein-A did not influence fold or vesicle formation but bound to the edges of these structures preferentially. It is concluded that DPPC monolayers can form bilayers spontaneously in the absence of surfactant apoproteins, other proteins or agents. PMID- 11284202 TI - Formation of flat lamellar intramembrane lipid structures in microorganisms. AB - Analysis of freeze-fracture replicas and thin sections of cells of the bacteria Sulfobacillus thermosulfidooxidans and Anaerobacter polyendosporus showed that their cytoplasmic membranes contain some regions in the form of flat lamellar inverted lipid membranes a few tenths of nanometers to a few microns in size. The specific features of these membrane structures are as follows: (i) they contain no familiar intramembrane particles commonly present on freeze-fracture replicas; (ii) in cross thin sections, intramembrane structures are bi-furcate on the periphery and look like thylakoids; and (iii) the leaflets of intramembrane structures in S. thermosulfidooxidans cells are corrugated. These structures were revealed in bacterial cells cultivated under normal growth conditions. The data obtained suggest the occurrence of a complex type of compartmentalization in biological membranes. PMID- 11284200 TI - Gramicidin-perforated patch analysis on HCO3- secretion through a forskolin activated anion channel in rat parotid intralobular duct cells. AB - Forskolin-induced anion currents and depolarization were investigated to clarify the mechanism of HCO3- secretion in the intralobular duct cells of rat parotid glands. Anion currents of the cells were measured at the equilibrium potential of K+, using a gramicidin-perforated patch technique that negligibly affects intracellular anion concentration. The forskolin-induced anion current was sustained and significantly (54%) suppressed by glibenclamide (200 microM), a blocker of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel. The anion current was markedly suppressed by addition of 1 mM methazolamide, a carbonic anhydrase inhibitor, and removal of external HCO3-. Forskolin depolarized the cells in the current-clamp mode. Addition of methazolamide and removal of external HCO3- significantly decreased the depolarizing level. These results suggest that activation of anion channels (mainly the CFTR Cl- channel located in luminal membranes) and production of cytosolic HCO3- induce the inward anion current and resulting depolarization. Inhibition of the Na(+)-K(+)-2Cl- cotransporter and the Cl(-)-HCO3- exchanger had no significant effect on the current or depolarization, indicating that the uptake of Cl- via the Na(+)-K(+)-2Cl- cotransporter or the Cl(-)-HCO3- exchanger is not involved in the responses. Taken together, we conclude that forskolin activates the outward movement (probably secretion) of HCO3- produced intracellularly, but not of Cl- due to lack of active Cl- transport in parotid duct cells, and that the gramicidin-perforated patch method is very useful to analyze anion transport. PMID- 11284203 TI - Regulation of an outwardly rectifying chloride conductance in renal epithelial cells by external and internal calcium. AB - We have used perforated patch clamp and Fura-2 microfluorescence measurements to study Ca(2+)-activated Cl- currents in cultured mouse renal inner medullary collecting duct cells (mIMCD-3). The conductance was spontaneously active under resting conditions and whole cell currents were time and voltage-independent with an outwardly rectifying current-voltage relationship. The channel blockers DIDS, niflumic acid, glybenclamide and NPPB reversibly decreased the basal currents, whereas the sulfhydryl agent, DTT produced an irreversible inhibition. Increasing or decreasing extracellular calcium produced parallel changes in the size of the basal currents. Variations in external Ca2+ were associated with corresponding changes in free cytosolic Ca2+ concentration. Increasing cytosolic Ca2+ by extracellular ATP or ionomycin, further enhanced Cl- conductance, with whole cell currents displaying identical biophysical properties to the basal currents. However, the agonist-stimulated currents were now increased by DTT exposure, but still inhibited by the other channel blockers. Using RT-PCR, three distinct mRNA transcripts belonging to the CLCA family of Ca(2+)-activated Cl- channel proteins were identified, two of which represent novel sequences. Whether different channels underlie the basal and agonist-stimulated currents in mIMCD-3 cells is unclear. Our findings establish a novel link between alterations in external and internal Ca2+ and the activity of Ca(2+)-activated Cl- transport in these cells. PMID- 11284204 TI - Voltage-induced activation of mechanosensitive cation channels of leech neurons. AB - The voltage dependence of stretch-activated cation channels in leech central neurons was studied in cell-free configurations of the patch-clamp technique. We established that stretch-activated channels excised from identified cell bodies of desheathed ganglia, as well as from neurons in culture, were slowly and reversibly activated by depolarizing membrane potentials. Negative pressure stimuli, applied to the patch pipette during a slow periodical modulation of membrane potential, enhanced channel activity, whereas positive pressures depressed it. Voltage-induced channel activation was observed, with soft glass pipettes, both in inside-out and outside-out membrane patches, at negative and positive reference potentials, respectively. The results presented in this study demonstrate that membrane depolarization induces slow activation of stretch activated channels of leech central neurons. This phenomenon is similar to that found in Xenopus oocytes, however, some peculiar features of the voltage dependence in leech stretch-activated channels indicate that specific membrane glass interactions might not necessarily be involved. Moreover, following depolarization, stretch-activated channels in membrane patches from neurons in culture exhibited significantly shorter delay to activation (sec) than their counterparts from neurons of freshly isolated ganglia (hundreds of sec). PMID- 11284205 TI - Actin modulates the gating of Neurospora crassa VDAC. AB - VDAC forms the major pathway for metabolites across the mitochondrial outer membrane. The regulation of the gating of VDAC channels is an effective way to control the flow of metabolites into and out of mitochondria. Here we present evidence that actin can modulate the gating process of Neurospora crassa VDAC reconstituted into membranes made with phosphatidylcholine. An actin concentration as low as 50 nM caused the VDAC-mediated membrane conductance to drop by as much as 85% at elevated membrane potentials. Actin's effect could be quickly reversed by adding pronase to digest the protein. alpha-Actin, from mammalian muscle, has a stronger effect than beta- and gamma-actin from human platelets. The monomeric form of actin, G-actin, is effective. Stabilization of the fibrous form, F-actin, with the mushroom toxin, phalloidin, blocks the effect of actin on VDAC, indicating that F-actin might be ineffective. Cytochalasin B did not interfere with the ability of actin to favor VDAC closure. DNase- did effectively block actin's effect on VDAC, and VDAC decreased actin's inhibitory effect on DNase-I activity, indicating that N. crassa VDAC competes with DNase-I for the same binding site on actin. The actin-VDAC interaction might be a mechanism by which actin regulates energy metabolism. PMID- 11284206 TI - Investigating the surface expression of the renal type IIa Na+/Pi-cotransporter in Xenopus laevis oocytes. AB - We have combined a functional assay, surface labeling and immunocytochemical methods to compare total and surface-exposed renal type IIa Na+/Pi cotransporter protein. The wild-type type cotransporter (NaPi-IIa) and its functionally comparable cysteine mutant S460C were expressed in Xenopus oocytes. S460C contains a novel cysteine residue that, when modified by preincubation with methanethiosulfonate reagents, leads to complete suppression of cotransport function. This allowed surface labeling of the S460C using MTSEA-Biotin and confirmation by electrophysiology on the same cell. Protein was analyzed by Western blotting before and after streptavidin precipitation and by immunocytochemistry and immunogold electronmicroscopy. MTSEA-Biotin treatment resulted in a complete inhibition of S460C-mediated Na+/Pi-cotransport activity, which indicated that all transporters at the surface were biotinylated. After biotinylation, only a small fraction of total S460C protein was precipitated by streptavidin compared with the total amount of S460C protein detected in the lysate. Light- and electron-microscopy analysis of oocytes showed a large amount of WT and S460C transporter protein beneath the oocyte membrane. These data indicate that the apparent weak labeling efficiencies of surface-biotinylation based assays of membrane proteins heterologously expressed in oocytes can be related to diminished incorporation of the protein in the oolemma. PMID- 11284207 TI - The essential role of cytosolic Cl- in Ca2+ regulation of an amiloride-sensitive channel in fetal rat pneumocyte. AB - An amiloride-sensitive, Ca(2+)-activated nonselective cation (NSC) channel in the apical membrane of fetal rat alveolar epithelium plays an important role in stimulation of Na+ transport by a beta adrenergic agonist (beta agonist). We studied whether Ca2+ has an essential role in the stimulation of the NSC channel by beta agonists. In cell-attached patches formed on the epithelium, terbutaline, a beta agonist, increased the open probability (Po) of the NSC channel to 0.62 +/ 0.07 from 0.03 +/- 0.01 (mean +/- SE; n = 8) 30 min after application of terbutaline in a solution containing 1 mM Ca2+. The Po of the terbutaline stimulated NSC channel was diminished in the absence of extracellular Ca2+ to 0.26 +/- 0.05 (n = 8). The cytosolic Ca2+ concentration ([Ca2+]c) in the presence and absence of extracellular Ca2+ was, respectively, 100 +/- 6 and 20 +/- 2 nM (n = 7) 30 min after application of terbutaline. The cytosolic Cl- concentration ([Cl-]c) in the presence and absence of extracellular Ca2+ was, respectively, 20 +/- 1 and 40 +/- 2 mM (n = 7) 30 min after application of terbutaline. The diminution of [Ca2+]c from 100 to 20 nM itself had no significant effects on the Po if the [Cl-]c was reduced to 20 mM; the Po was 0.58 +/- 0.10 at 100 nM [Ca2+]c and 0.55 +/- 0.09 at 20 nM [Ca2+]c (n = 8) with 20 mM [Cl-]c in inside-out patches. On the other hand, the Po (0.28 +/- 0.10) at 20 nM [Ca2+]c with 40 mM [Cl-]c was significantly lower than that (0.58 +/- 0.10; P < 0.01; n = 8) at 100 nM [Ca2+]c with 20 mM [Cl-]c' suggesting that reduction of [Cl-]c is an important factor stimulating the NSC channel. These observations indicate that the extracellular Ca2+ plays an important role in the stimulatory action of beta agonist on the NSC channel via reduction of [Cl-]c. PMID- 11284208 TI - Recommendations on health-related quality of life research to support labeling and promotional claims in the United States. AB - Health-related quality of life (HRQL) outcomes evaluation is becoming an important component of clinical trials of new pharmaceuticals and medical devices. HRQL research provides patients, providers, and decision makers with important information on the impact of disease and treatment on physical, psychological, and social functioning and well-being. These outcomes are also useful to the pharmaceutical and device industries as they attempt to understand and communicate product value to physicians, patients, health insurers and others. HRQL labeling and promotional claims in the US are likely to increase over the next few years. The evidentiary requirements to make such a claim should be based on accepted scientific standards of HRQL evaluation and consistent with the regulatory requirements for clinical efficacy. This report outlines the scientific practices that should be considered in the evaluation of evidence for an HRQL claim, including the selection of appropriate domains, evidence to support the reliability and validity of HRQL measurement, considerations in research design and statistical analyses, and the issue of clinical significance. Representatives from the pharmaceutical and device industries, regulatory agencies, and the HRQL scientific community should work together to make certain the use of HRQL in labeling and promotion are based on sound scientific evidence, and that these messages are clearly and accurately reported to the consumers. PMID- 11284209 TI - The use of the Tobit model for analyzing measures of health status. AB - Self-reported health status is often measured using psychometric or utility indices that provide a score intended to summarize an individual's health. Measurements of health status can be subject to a ceiling effect. Frequently, researchers want to examine relationships between determinants of health and measures of health status. Regression methods that ignore the presence of a ceiling effect, or of censoring in the health status measurements can produce biased coefficient estimates. The Tobit regression model is a frequently used tool for modeling censored variables in econometrics research. The authors carried out a Monte-Carlo simulation study to contrast the performance of the Tobit model for censored data with that of ordinary least squares (OLS) regression. It was demonstrated that in the presence of a ceiling effect, if the conditional distribution of the measure of health status had uniform variance, then the coefficient estimates from the Tobit model have superior performance compared with estimates from OLS regression. However, if the conditional distribution had non-uniform variance, then the Tobit model performed at least as poorly as the OLS model. PMID- 11284210 TI - Censoring in statistical models of health status: what happens when one can do better than '1'. PMID- 11284211 TI - Dealing with ceiling baseline treatment satisfaction level in patients with diabetes under flexible, functional insulin treatment: assessment of improvements in treatment satisfaction with a new insulin analogue. AB - Even a small improvement in satisfaction with treatment for a chronic disease can be valuable. However, sensitive measurements instruments are needed to assess the effects of treatment changes in patients already well satisfied with baseline treatment. Such instruments were thought to be necessary to deal with ceiling effects in scores of satisfaction with functional insulin treatment (FIT) accommodating full flexibility of food intake and lifestyle in diabetes by manipulation of insulin (FIT; Howorka et al. 2000). The Status(S) version of the Diabetes Treatment Satisfaction Questionnaire (DTSQ, Bradley 1994) was extended with items measuring specific components of FIT and its psychometric properties investigated in a validation study with 171 FIT patients with diabetes. A new Change(C) version of DTSQ extended for FIT was used together with the DTSQ(S) by 52 participants in a subsequent randomised cross-over study involving new fast acting lispro vs. regular insulin, where treatment satisfaction was the primary outcome variable. Insulin lispro use improved satisfaction (p < or = 0.001) both, on classical and extended versions of DTSQ(S) and (C), despite high baseline levels of satisfaction. However, the DTSQ(C) augmented the effects of lispro (p = 0.0001) apparent on DTSQ(S) scores and revealed significant treatment effects (p < or = 0.01) not significant with DTSQ(S). In parallel, glycaemic control improved with lispro (e.g., HbA1c, p = 0.023). Improved satisfaction with treatment was more apparent with DTSQ(C) than DTSQ(S) in patients who at baseline were at or near ceiling for treatment satisfaction. PMID- 11284212 TI - Measuring health related quality of life among women living with HIV. AB - Although women had been under recognized in the literature on HIV/AIDS, increasing numbers of studies have focused on the lives and experiences of women living with HIV/AIDS. Areas of research in which the study of women and HIV continues to be noticeably lacking include health related quality of life (HRQOL). This paper describes HRQOL in an inner city sample of 287 HIV positive non-pregnant women, interviewed as part of a larger multi-site CDC funded study of the reproductive health of women. The average age of the respondents was 33 years and women had known their HIV status an average of 41 months. HRQOL was assessed using a 17-item modified version of the Medical Outcomes Study-HIV Health Survey and demonstrated acceptable internal consistency (0.64-0.89) and variability. Women in our study were similar to other HIV-positive female samples and reported lower levels of well-being and functioning than some HIV-positive male samples. Women's responses to individual items and areas of potential need for health care are discussed. Examination of HRQOL in women with HIV can aid in the comparison of how women and men are affected by HIV and can help health care professionals identify needed services and include possible interventions to promote quality of life. PMID- 11284213 TI - Health related quality of life among persons with HIV after the use of protease inhibitors. AB - OBJECTIVE: This study was conducted to determine the effect of the use of HIV protease inhibitors on the quality of life among persons infected with HIV. METHODS: Subjects were participants in the British Columbia Centre for Excellence in HIV/AIDS Drug Treatment Program who had completed two annual participant surveys, one prior to initiating therapy with a protease inhibitor and one after. Quality of life was measured using the Medical Outcomes Study Short Form Health Survey (MOS-SF). Statistical analyses were conducted using parametric and multivariate methods. RESULTS: Our analysis was based on 179 HIV-positive individuals. Compared to quality of life at baseline, we found no statistically significant changes in the health perception, pain, physical, role and social functioning MOS-SF subscale scores at follow-up. The measure of mental health was the only component to decline significantly over time. Subanalyses found significant increases in the measures of health perception (p = 0.004), physical (p = 0.037), role (p < 0.001) and social functioning (p = 0.053) for individuals with a low baseline quality of life. For those with a higher quality of life before starting a protease inhibitor containing regimen we observed a significant decline in the subscales of mental health (p = 0.001), physical (p = 0.007), role (p = 0.021) and social functioning (p = 0.003) over the study period. CONCLUSIONS: Our findings indicate that despite strict dosage requirements and adverse side effects associated with protease inhibitor use patients are on the whole maintaining their quality of life after starting on a protease inhibitor containing regimen. Continued follow-up of this cohort will be required to determine the long term implications of these observations. PMID- 11284214 TI - A prospective investigation of dispositional optimism as a predictor of health related quality of life in head and neck cancer patients. AB - The aim of this study was to investigate the role of dispositional optimism (DO) as a predictor of health-related quality of life (HRQL) in a sample of upper aerodigestive tract cancer (UADT) patients. A prospective observational study design was used with a cohort of patients from one centre. DO was evaluated using a French version of the Life Orientation Test (the FLOT) translated and validated for this study. HRQL was evaluated using the EORTC QLQ-C30 prior to and 3 months following treatment. The association between FLOT ratings and HRQL was evaluated using linear multiple regression analysis and a two-way ANOVA with repeated measures. Baseline data were gathered on 101 subjects and follow-up data on 88 of these. The sample was dichotomized around the median FLOT score creating 'optimist' and 'pessimist' groups. Before treatment, optimists reported better role, cognitive and emotional function, less pain and fatigue and a better global rating of HRQL than did pessimists. Following treatment, optimists reported better role and cognitive functioning, less pain and better global HRQL than did pessimists. Pessimists reported a greater deterioration in the role domain following treatment than did optimists. At no point did pessimists rate HRQL better than optimists. The results suggest that optimism is associated with better HRQL in French UADT cancer patients. PMID- 11284215 TI - Development and psychometric evaluation of the TAPQOL: a health-related quality of life instrument for 1-5-year-old children. AB - The 43-item TNO-AZL Preschool Children Quality of Life (TAPQOL) questionnaire was developed to meet the need for a reliable and valid instrument for measuring parent's perceptions of health-related quality of life (HRQoL) in preschool children. HRQoL was defined as health status in 12 domains weighted by the impact of the health status problems on well-being. The aim of this study was to evaluate the psychometric performance of the TAPQOL. A sample of 121 parents of preterm children completed the TAPQOL questionnaire (response rate 88%) as well as 362 parents of children from the general population (response rate 60%). On the base of Cronbach's alpha, item-rest correlation, and principal component analysis, the TAPQOL scales were constructed from the data for the preterm children sample. The psychometric performance of these scales was evaluated for both the preterm children sample and the general population sample. Cronbach's alpha ranged from 0.66 to 0.88 for the preterm children sample and from 0.43 to 0.84 for the general population sample. The unidimensionality of the separate scales was confirmed by principal component analysis for both the preterm children sample and the general population sample. Spearman's correlation coefficients between scales were, on average, low. T-tests showed that the very preterm children, the children with chronic diseases, the less healthy and the less happy children had lower mean scores on the TAPQOL scales than healthy children, indicating a worse quality of life. This study shows that the TAPQOL is a reliable and valid parent's perception of HRQoL in preschool children. More research is needed to evaluate the psychometric performance of the TAPQOL in different clinical populations. PMID- 11284216 TI - The English 'sweate' (Sudor Anglicus) and Hantavirus pulmonary syndrome. AB - A rapidly fatal viral infectious disease appeared in England in 1485, persisted for the summer months and disappeared as winter approached. This pattern of infection re-appeared in 1508, 1517, 1528, and finally 1551. The epidemic never returned. It had no respect for wealth or rank, and predominantly attacked males between the ages of 15 and 45 years. The incubation period was frighteningly short and the outcome normally fatal. The symptoms of acute respiratory disease and copious sweating were characteristic, providing the name 'the English sweating disease'. It was never in the big league of killer epidemics, such as plague and influenza, but its pockets of instant lethality in communities gave it a special ranking of horror. The infective cause of this disease remained a total mystery until it was compared with Hantavirus pulmonary syndrome (HPS) in 1994. The strength of this theory is examined in this paper, and it is concluded that, although there is a close resemblance, HPS does not match the English sweating disease completely and positive identification of a possible rodent carrier for the latter was not established. PMID- 11284217 TI - Identification of Candida dubliniensis, based on ribosomal DNA sequence analysis. AB - Differentiation of Candida albicans and the recently described C. dubliniensis has proven difficult due to the high degree of phenotypic similarity of these species. The present study examines sequence variations in the ribosomal DNA (rDNA) intergenic transcribed spacer (ITS) regions of C. albicans (n = 5) and C. dubliniensis (n = 7) strains, with a view to identifying sequence differences that would enable consistent differentiation of these two species by restriction fragment length polymorphism (RFLP) analysis. The ITS1 and ITS2 regions, together with the entire 5.8S rRNA gene of the strains, were amplified by the polymerase chain reaction (PCR), using primers ITS1 and ITS4, PCR products from both C. albicans and C. dubliniensis were of similar size (around 540 bp); however, sequence analysis revealed over 20 consistent base differences between the products of the two species. On the basis of sequence variation, the restriction enzyme MspA1 I was selected and used to differentiate the PCR products of C. albicans and C. dubliniensis by RFLP analysis. MspA1 I yielded two discernible fragments from C. albicans PCR products, whilst those from C. dubliniensis appeared undigested, thereby providing an approach to differentiate the two species. PMID- 11284218 TI - Meningococcal disease due to serogroup Y in Scotland, 1992-1999. AB - Meningococcal disease is an important cause of morbidity and mortality. A retrospective analysis was performed of all cases of invasive group-Y disease that were laboratory-confirmed in Scotland between 1992 and 1999. A total of 1881 meningococcal isolates were characterised, 78 of which were serogroup Y. The incidence of non-invasive group-Y disease remained level between 1992 and 1999. Only 12 isolates were from invasive disease, comprising five strain types. Invasive group-Y disease was associated mostly with the young or old. Serogroup-Y meningococcal disease was uncommon and a rare cause of invasive disease in Scotland between 1992 and 1999; however, it is essential that microbiologists are aware of its potential for increasing in incidence due to the recent introduction of the MenC vaccine, and its increased incidence in the USA. PMID- 11284219 TI - False-positive serological tests in acute malaria. AB - A range of serological tests, including rapid plasma reagin (RPR), Widal test, enzyme-linked immunosorbent assay (ELISA) for human immunodeficiency virus (HIV) 1 and -2, direct Coombs' test (DCT), and rheumatoid factor (RF) were performed in a well-characterised cohort of 100 patients with acute malaria (Plasmodium vivax infection: 31 patients; P. falciparum infection: 69 patients). Twenty-five healthy volunteers from a similar area were used as controls. Three patients from the severe P. falciparum group died, the remainder of the patients recovered completely. A large proportion of these patients showed false-positive serological reactions during the acute stage of infection, which became negative on re-testing, four weeks after recovery. In tropical countries such as India, where malaria is endemic, results of serological tests should be interpreted with caution in a patient with pyrexia of unknown origin (PUO). PMID- 11284220 TI - Detection of high-risk subtypes of human papillomavirus in cervical swabs: routine use of the Digene Hybrid Capture assay and polymerase chain reaction analysis. AB - Human papillomaviruses (HPVs) are major causative agents in the pathogenesis of cervical cancer, and more than twenty types are associated with its development. With the introduction of liquid-based preparation systems, it is envisaged that large-scale HPV testing will be established in the near future. Preliminary studies demonstrate the accessibility of these samples for DNA testing using both the Digene Hybrid Capture assay (DHCA) and polymerase chain reaction (PCR) techniques. This study aims to assess the validity and sensitivity of the DHCA system to detect high-risk HPV DNA, using two sets of HPV consensus primers (Gp5+/Gp6+ and MY09/MY11) in tandem with routine assessment of cervical smear and biopsy samples. Results indicate that the combination of DHCA and PCR detects more high-grade lesions than does the DHCA alone. DHCA-negative cases were categorised by subsequent PCR amplification into low-grade HPV-negative (12/16) cervical lesions and high-grade HPV-positive (7/9) cervical lesions. Gp5+/Gp6+ primers were less sensitive in detecting HPV-positive samples than was the MY09/MY11 primer set. These results support the use of high-risk HPV testing by DHCA, with subsequent analysis of DHCA-negative samples by PCR using the MY09/MY11 primers.